PMID- 10848590 TI - Dominant active alleles of RIM101 (PRR2) bypass the pH restriction on filamentation of Candida albicans. AB - Morphological development of the fungal pathogen Candida albicans is profoundly affected by ambient pH. Acidic pH restricts growth to the yeast form, whereas neutral pH permits development of the filamentous form. Superimposed on the pH restriction is a temperature requirement of approximately 37 degrees C for filamentation. The role of pH in development was investigated by selecting revertants of phr2Delta mutants that had gained the ability to grow at acid pH. The extragenic suppressors in two independent revertants were identified as nonsense mutations in the pH response regulator RIM101 (PRR2) that resulted in a carboxy-terminal truncation of the open reading frame. These dominant active alleles conferred the ability to filament at acidic pH, to express PHR1, an alkaline-expressed gene, at acidic pH, and to repress the acid-expressed gene PHR2. It was also observed that both the wild-type and mutant alleles could act as multicopy suppressors of the temperature restriction on filamentation, allowing extensive filamentation at 29 degrees C. The ability of the activated alleles to promote filamentation was dependent upon the developmental regulator EFG1. The results suggest that RIM101 is responsible for the pH dependence of hyphal development. PMID- 10848591 TI - Functional characterization of the X-linked inhibitor of apoptosis (XIAP) internal ribosome entry site element: role of La autoantigen in XIAP translation. AB - X-linked inhibitor of apoptosis protein (XIAP) is a key regulator of programmed cell death triggered by various apoptotic triggers. Translation of XIAP is controlled by a 162-nucleotide (nt) internal ribosome entry site (IRES) element located in the 5' untranslated region of XIAP mRNA. XIAP IRES mediates efficient translation of XIAP under physiological stress and enhances cell protection against serum deprivation and radiation-induced apoptosis. In the present report we describe the assembly of a sequence-specific RNA-protein complex consisting of at least four cytosolic proteins on the XIAP IRES element. We determine that the core binding sequence is approximately 28 nt long and is located 34 nt upstream of the initiation site. Moreover, we identify the La autoantigen as a protein that specifically binds XIAP IRES in vivo and in vitro. The biological relevance of this interaction is further demonstrated by the inhibition of XIAP IRES mediated translation in the absence of functional La protein. The results suggest an important role for the La protein in the regulation of XIAP expression, possibly by facilitating ribosome recruitment to the XIAP IRES. PMID- 10848592 TI - Involvement of Ras and Ral in chemotactic migration of skeletal myoblasts. AB - In skeletal myoblasts, Ras has been considered to be a strong inhibitor of myogenesis. Here, we demonstrate that Ras is involved also in the chemotactic response of skeletal myoblasts. Expression of a dominant-negative mutant of Ras inhibited chemotaxis of C2C12 myoblasts in response to basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and insulin-like growth factor 1 (IGF-1), key regulators of limb muscle development and skeletal muscle regeneration. A dominant-negative Ral also decreased chemotactic migration by these growth factors, while inhibitors for phosphatidylinositol 3-kinase and mitogen-activated protein kinase kinase (MEK) showed no effect. Activation of the Ras-Ral pathway by expression of an activated mutant of either Ras, the guanine nucleotide dissociation stimulator for Ral, or Ral resulted in increased motility of myoblasts. The ability of Ral to stimulate motility was reduced by introduction of a mutation which prevents binding to Ral-binding protein 1 or phospholipase D. These results suggest that the Ras-Ral pathway is essential for the migration of myoblasts. Furthermore, we found that Ras and Ral are activated in C2C12 cells by bFGF, HGF and IGF-1 and that the Ral activation is regulated by the Ras- and the intracellular Ca(2+)-mediated pathways. Taken together, our data indicate that Ras and Ral regulate the chemotactic migration of skeletal muscle progenitors. PMID- 10848593 TI - Binding of HMG-I(Y) imparts architectural specificity to a positioned nucleosome on the promoter of the human interleukin-2 receptor alpha gene. AB - Transcriptional induction of the interleukin-2 receptor alpha-chain (IL-2Ralpha) gene is a key event regulating T-cell-mediated immunity in mammals. In vivo, the T-cell-restricted protein Elf-1 and the general architectural transcription factor HMG-I(Y) cooperate in transcriptional regulation of the human IL-2Ralpha gene by binding to a specific positive regulatory region (PRRII) in its proximal promoter. Employing chromatin reconstitution analyses, we demonstrate that the binding sites for both HMG-I(Y) and Elf-1 in the PRRII element are incorporated into a strongly positioned nucleosome in vitro. A variety of analytical techniques was used to determine that a stable core particle is positioned over most of the PRRII element and that this nucleosome exhibits only a limited amount of lateral translational mobility. Regardless of its translational setting, the in vitro position of the nucleosome is such that DNA recognition sequences for both HMG-I(Y) and Elf-1 are located on the surface of the core particle. Restriction nuclease accessibility analyses indicate that a similarly positioned nucleosome also exists on the PRRII element in unstimulated lymphocytes when the IL-2Ralpha gene is silent and suggest that this core particle is remodeled following transcriptional activation of the gene in vivo. In vitro experiments employing the chemical cleavage reagent 1,10-phenanthroline copper (II) covalently attached to its C-terminal end demonstrate that HMG-I(Y) protein binds to the positioned PRRII nucleosome in a direction-specific manner, thus imparting a distinct architectural configuration to the core particle. Together, these findings suggest a role for the HMG-I(Y) protein in assisting the remodeling of a critically positioned nucleosome on the PRRII promoter element during IL-2Ralpha transcriptional activation in lymphocytes in vivo. PMID- 10848594 TI - Distinct pathways of cell migration and antiapoptotic response to epithelial injury: structure-function analysis of human intestinal trefoil factor. AB - The trefoil peptide intestinal trefoil factor (ITF) plays a critical role in the protection of colonic mucosa and is essential to restitution after epithelial damage. These functional properties are accomplished through coordinated promotion of cell migration and inhibition of apoptosis. ITF contains a unique three-looped trefoil motif formed by intrachain disulfide bonds among six conserved cysteine residues, which is thought to contribute to its marked protease resistance. ITF also has a seventh cysteine residue, which permits homodimer formation. A series of cysteine-to-serine substitutions and a C terminally truncated ITF were made by PCR site-directed mutagenesis. Any alteration of the trefoil motif or truncation resulted in loss of protease resistance. However, neither an intact trefoil domain nor dimerization was required to promote cell migration. This pro-restitution activity correlated with the ability of the ITF mutants to activate mitogen-activated protein (MAP) kinase independent of phosphorylation of the epidermal growth factor (EGF) receptor. In contrast, only intact ITF retained both phosphatidylinositol 3-kinase and the EGF receptor-dependent antiapoptotic effect in HCT116 and IEC-6 cells. The inability to block apoptosis correlated with a loss of trefoil peptide-induced transactivation of the EGF receptor or Akt kinase in HT-29 cells. In addition to defining structural requirements for the functional properties of ITF, these findings demonstrate that distinct intracellular signaling pathways mediate the effects of ITF on cell migration and apoptosis. PMID- 10848595 TI - Evidence for an interaction between ubiquitin-conjugating enzymes and the 26S proteasome. AB - The targeting of proteolytic substrates is accomplished by a family of ubiquitin conjugating (E2) enzymes and a diverse set of substrate recognition (E3) factors. The ligation of a multiubiquitin chain to a substrate can promote its degradation by the proteasome. However, the mechanism that facilitates the translocation of a substrate to the proteasome in vivo is poorly understood. We have discovered that E2 proteins, including Ubc1, Ubc2, Ubc4, and Ubc5, can interact with the 26S proteasome. Significantly, the interaction between Ubc4 and the proteasome is strongly induced by heat stress, consistent with the requirement for this E2 for efficient stress tolerance. A catalytically inactive derivative of Ubc4 (Ubc4(C86A)), which causes toxicity in yeast cells, can also bind the proteasome. Purified proteasomes can ligate ubiquitin to a test substrate without the addition of exogenous E2 protein, suggesting that the ubiquitylation of some proteolytic substrates might be directly coupled to degradation by the proteasome. PMID- 10848596 TI - Peroxisome proliferator-activated receptor gamma-dependent repression of the inducible nitric oxide synthase gene. AB - The peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear receptor superfamily that activates target gene transcription in a ligand-dependent manner. In addition, liganded PPARgamma can inhibit transcription of genes induced by gamma interferon (IFN-gamma) and/or lipopolysaccharides (LPSs), including the inducible nitric oxide synthase (iNOS) gene. Inhibition of the iNOS promoter is achieved partially through antagonizing the activities of NF-kappaB, AP-1, and STAT1, which are known to mediate effects of LPS and IFN-gamma. Previous studies have suggested that transrepression of these factors by nuclear receptors involves competition for limiting amounts of the general coactivators CREB-binding protein (CBP) and p300. CBP and p300 are thought to be recruited to nuclear receptors through bridging factors that include SRC-1, although CBP also interacts directly with PPARgamma through its amino terminus. These observations have raised questions concerning the involvement of SRC-1-like factors in CBP recruitment and transrepression. We here provide evidence that PPARgamma's ability to repress iNOS transcription requires the ligand-dependent charge clamp that mediates interactions with CBP and SRC-1. Single amino acid mutations in PPARgamma that abolished ligand-dependent interactions with SRC-1 and CBP not only resulted in complete loss of transactivation activity but also abolished transrepression. Conversely, a CBP deletion mutant containing the SRC-1 interaction domain but lacking the N terminal PPARgamma interaction domain was inactive as a PPARgamma coactivator and failed to rescue transrepression. Together, these findings are consistent with a model in which transrepression by PPARgamma is achieved by targeting CBP through direct interaction with its N-terminal domain and via SRC-1-like bridge factors. PMID- 10848597 TI - Cross talk of pp125(FAK) and pp59(Lyn) non-receptor tyrosine kinases to insulin mimetic signaling in adipocytes. AB - Signaling molecules downstream from the insulin receptor, such as the insulin receptor substrate protein 1 (IRS-1), are also activated by other receptor tyrosine kinases. Here we demonstrate that the non-receptor tyrosine kinases, focal adhesion kinase pp125(FAK) and Src-class kinase pp59(Lyn), after insulin independent activation by phosphoinositolglycans (PIG), can cross talk to metabolic insulin signaling in rat and 3T3-L1 adipocytes. Introduction by electroporation of neutralizing antibodies against pp59(Lyn) and pp125(FAK) into isolated rat adipocytes blocked IRS-1 tyrosine phosphorylation in response to PIG but not insulin. Introduction of peptides encompassing either the major autophosphorylation site of pp125(FAK), tyrosine 397, or its regulatory loop with the twin tyrosines 576 and 577 inhibited PIG-induced IRS-1 tyrosine phosphorylation and glucose transport. PIG-induced pp59(Lyn) kinase activation and pp125(FAK) tyrosine phosphorylation were impaired by the former and latter peptide, respectively. Up-regulation of pp125(FAK) by integrin clustering diminished PIG-induced IRS-1 tyrosine phosphorylation and glucose transport in nonadherent but not adherent adipocytes. In conclusion, PIG induced IRS-1 tyrosine phosphorylation by causing (integrin antagonized) recruitment of IRS-1 and pp59(Lyn) to the common signaling platform molecule pp125(FAK), where cross talk of PIG-like structures and extracellular matrix proteins to metabolic insulin signaling may converge, possibly for the integration of the demands of glucose metabolism and cell architecture. PMID- 10848598 TI - The Fanconi anemia protein FANCC binds to and facilitates the activation of STAT1 by gamma interferon and hematopoietic growth factors. AB - Hematopoietic progenitor cells from Fanconi anemia (FA) group C (FA-C) patients display hypersensitivity to the apoptotic effects of gamma interferon (IFN-gamma) and constitutively express a variety of IFN-dependent genes. Paradoxically, however, STAT1 activation is suppressed in IFN-stimulated FA cells, an abnormality corrected by transduction of normal FANCC cDNA. We therefore sought to define the specific role of FANCC protein in signal transduction through receptors that activate STAT1. Expression and phosphorylation of IFN-gamma receptor alpha chain (IFN-gammaRalpha) and JAK1 and JAK2 tyrosine kinases were equivalent in both normal and FA-C cells. However, in coimmunoprecipitation experiments STAT1 did not dock at the IFN-gammaR of FA-C cells, an abnormality corrected by transduction of the FANCC gene. In addition, glutathione S transferase fusion genes encoding normal FANCC but not a mutant FANCC bearing an inactivating point mutation (L554P) bound to STAT1 in lysates of IFN-gamma stimulated B cells and IFN-, granulocyte-macrophage colony-stimulating factor- and stem cell factor-stimulated MO7e cells. Kinetic studies revealed that the initial binding of FANCC was to nonphosphorylated STAT1 but that subsequently the complex moved to the receptor docking site, at which point STAT1 became phosphorylated. The STAT1 phosphorylation defect in FA-C cells was functionally significant in that IFN induction of IFN response factor 1 was suppressed and STAT1-DNA complexes were not detected in nuclear extracts of FA-C cells. We also determined that the IFN-gamma hypersensitivity of FA-C hematopoietic progenitor cells does not derive from STAT1 activation defects because granulocyte macrophage CFU and erythroid burst-forming units from STAT1(-/-) mice were resistant to IFN-gamma. However, BFU-E responses to SCF and erythropoietin were suppressed in STAT(-/-) mice. Consequently, because the FANCC protein is involved in the activation of STAT1 through receptors for at least three hematopoietic growth and survival factor molecules, we reason that FA-C hematopoietic cells are excessively apoptotic because of an imbalance between survival cues (owing to a failure of STAT1 activation in FA-C cells) and apoptotic and mitogenic inhibitory cues (constitutively activated in FA-C cells in a STAT1-independent fashion). PMID- 10848599 TI - The Ste20 kinase misshapen regulates both photoreceptor axon targeting and dorsal closure, acting downstream of distinct signals. AB - We have previously shown that the Ste20 kinase encoded by misshapen (msn) functions upstream of the c-Jun N-terminal kinase (JNK) mitogen-activated protein kinase module in Drosophila. msn is required to activate the Drosophila JNK, Basket (Bsk), to promote dorsal closure of the embryo. A mammalian homolog of Msn, Nck interacting kinase, interacts with the SH3 domains of the SH2-SH3 adapter protein Nck. We now show that Msn likewise interacts with Dreadlocks (Dock), the Drosophila homolog of Nck. dock is required for the correct targeting of photoreceptor axons. We have performed a structure-function analysis of Msn in vivo in Drosophila in order to elucidate the mechanism whereby Msn regulates JNK and to determine whether msn, like dock, is required for the correct targeting of photoreceptor axons. We show that Msn requires both a functional kinase and a C terminal regulatory domain to activate JNK in vivo in Drosophila. A mutation in a PXXP motif on Msn that prevents it from binding to the SH3 domains of Dock does not affect its ability to rescue the dorsal closure defect in msn embryos, suggesting that Dock is not an upstream regulator of msn in dorsal closure. Larvae with only this mutated form of Msn show a marked disruption in photoreceptor axon targeting, implicating an SH3 domain protein in this process; however, an activated form of Msn is not sufficient to rescue the dock mutant phenotype. Mosaic analysis reveals that msn expression is required in photoreceptors in order for their axons to project correctly. The data presented here genetically link msn to two distinct biological events, dorsal closure and photoreceptor axon pathfinding, and thus provide the first evidence that Ste20 kinases of the germinal center kinase family play a role in axonal pathfinding. The ability of Msn to interact with distinct classes of adapter molecules in dorsal closure and photoreceptor axon pathfinding may provide the flexibility that allows it to link to distinct upstream signaling systems. PMID- 10848600 TI - Bcl-2 retards cell cycle entry through p27(Kip1), pRB relative p130, and altered E2F regulation. AB - Independent of its antiapoptotic function, Bcl-2 can, through an undetermined mechanism, retard entry into the cell cycle. Cell cycle progression requires the phosphorylation by cyclin-dependent kinases (Cdks) of retinoblastoma protein (pRB) family members to free E2F transcription factors. We have explored whether retarded cycle entry is mediated by the Cdk inhibitor p27 or the pRB family. In quiescent fibroblasts, enforced Bcl-2 expression elevated levels of both p27 and the pRB relative p130. Bcl-2 still slowed G(1) progression in cells deficient in pRB but not in those lacking p27 or p130. Hence, pRB is not required, but both p27 and p130 are essential mediators. The ability of p130 to form repressive complexes with E2F4 is implicated, because the retardation by Bcl-2 was accentuated by coexpressed E2F4. A plausible relevant target of p130/E2F4 is the E2F1 gene, because Bcl-2 expression delayed E2F1 accumulation during G(1) progression and overexpression of E2F1 overrode the Bcl-2 inhibition. Hence, Bcl 2 appears to retard cell cycle entry by increasing p27 and p130 levels and maintaining repressive complexes of p130 with E2F4, perhaps to delay E2F1 expression. PMID- 10848601 TI - The downstream promoter element DPE appears to be as widely used as the TATA box in Drosophila core promoters. AB - The downstream promoter element (DPE) functions cooperatively with the initiator (Inr) for the binding of TFIID in the transcription of core promoters in the absence of a TATA box. We examined the properties of sequences that can function as a DPE as well as the range of promoters that use the DPE as a core promoter element. By using an in vitro transcription assay, we identified 17 new DPE dependent promoters and found that all possessed identical spacing between the Inr and DPE. Moreover, mutational analysis indicated that the insertion or deletion of a single nucleotide between the Inr and DPE causes a reduction in transcriptional activity and TFIID binding. To explore the range of sequences that can function as a DPE, we constructed and analyzed randomized promoter libraries. These experiments yielded the DPE functional range set, which represents sequences that contribute to or are compatible with DPE function. We then analyzed the DPE functional range set in conjunction with a Drosophila core promoter database that we compiled from 205 promoters with accurately mapped start sites. Somewhat surprisingly, the DPE sequence motif is as common as the TATA box in Drosophila promoters. There is, in addition, a striking adherence of Inr sequences to the Inr consensus in DPE-containing promoters relative to DPE less promoters. Furthermore, statistical and biochemical analyses indicated that a G nucleotide between the Inr and DPE contributes to transcription from DPE containing promoters. Thus, these data reveal that the DPE exhibits a strict spacing requirement yet some sequence flexibility and appears to be as widely used as the TATA box in Drosophila. PMID- 10848602 TI - HuD RNA recognition motifs play distinct roles in the formation of a stable complex with AU-rich RNA. AB - Human neuron-specific RNA-binding protein HuD belongs to the family of Hu proteins and consists of two N-terminal RNA recognition motifs (RRM1 and -2), a hinge region, and a C-terminal RRM (RRM3). Hu proteins can bind to AU-rich elements in the 3' untranslated regions of unstable mRNAs, causing the stabilization of certain transcripts. We have studied the interaction between HuD and prototype mRNA instability elements of the sequence UU(AUUU)(n)AUU using equilibrium methods and real-time kinetics (surface plasmon resonance using a BIACORE). We show that a single molecule of HuD requires at least three AUUU repeats to bind tightly to the RNA. Deletion of RRM1 reduced the K(d) by 2 orders of magnitude and caused a decrease in the association rate and a strong increase in the dissociation rate of the RNA-protein complex, as expected when a critical RNA-binding domain is removed. In contrast, deletion of either RRM2 or -3, which only moderately reduced the affinity, caused marked increases in the association and dissociation rates. The slower binding and stabilization of the complex observed in the presence of all three RRMs suggest that a change in the tertiary structure occurs during binding. The individual RRMs bind poorly to the RNA (RRM1 binds with micromolar affinity, while the affinities of RRM2 and -3 are in the millimolar range). However, the combination of RRM1 and either RRM2 or RRM3 in the context of the protein allows binding with a nanomolar affinity. Thus, the three RRMs appear to cooperate not only to increase the affinity of the interaction but also to stabilize the formed complex. Kinetic effects, similar to those described here, could play a role in RNA binding by many multi-RRM proteins and may influence the competition between proteins for RNA-binding sites and the ability of RNA-bound proteins to be transported intracellularly. PMID- 10848603 TI - Role of DBP in the circadian oscillatory mechanism. AB - Transcript levels of DBP, a member of the PAR leucine zipper transcription factor family, exhibit a robust rhythm in suprachiasmatic nuclei, the mammalian circadian center. Here we report that DBP is able to activate the promoter of a putative clock oscillating gene, mPer1, by directly binding to the mPer1 promoter. The mPer1 promoter is cooperatively activated by DBP and CLOCK-BMAL1. On the other hand, dbp transcription is activated by CLOCK-BMAL1 through E-boxes and inhibited by the mPER and mCRY proteins, as is the case for mPer1. Thus, a clock-controlled dbp gene may play an important role in central clock oscillation. PMID- 10848604 TI - Conservation and function of a potential substrate-binding domain in the yeast Clb5 B-type cyclin. AB - Cyclin A contains a region implicated in binding to the p27 inhibitor and to substrates. There is strong evolutionary conservation of surface residues contributing to this region in many cyclins, including yeast B-type cyclins, despite the absence of a yeast p27 homolog. The yeast S-phase B-type cyclin Clb5p interacted with mammalian p27 in a two-hybrid assay. This interaction was disrupted by mutations designed to disrupt hydrophobic interactions (hpm mutation) or hydrogen bonding (Q241A mutation) based on the cyclin A-p27 crystal structure. In contrast, mutation of the Clb5p p27-binding domain only slightly reduced binding and inhibition by the Sic1p Clb-Cdc28p kinase inhibitor. Mutations disrupting the p27-binding domain strongly reduced Clb5p biological activity in diverse assays without reducing Clb5p-associated kinase activity. An analogous hpm mutation in the mitotic cyclin Clb2p reduced mitotic function, but in some assays this mutation increased the ability of Clb2p to perform functions normally restricted to Clb5p. These results support the idea of a modular, structurally conserved cyclin domain involved in substrate targeting. PMID- 10848605 TI - Phosphorylation of tyrosine residues in the kinase domain and juxtamembrane region regulates the biological and catalytic activities of Eph receptors. AB - Members of the Eph family of receptor tyrosine kinases exhibit a striking degree of amino acid homology, particularly notable in the kinase and membrane-proximal regions. A mutagenesis approach was taken to address the functions of specific conserved tyrosine residues within these catalytic and juxtamembrane domains. Ligand stimulation of wild-type EphB2 in neuronal NG108-15 cells resulted in an upregulation of catalytic activity and an increase in cellular tyrosine phosphorylation, accompanied by a retraction of neuritic processes. Tyrosine-to phenylalanine substitutions within the conserved juxtamembrane motif abolished these responses. The mechanistic basis for these observations was examined using the highly related EphA4 receptor in a continuous coupled kinase assay. Tandem mass spectrometry experiments confirmed autophosphorylation of the two juxtamembrane tyrosine residues and also identified a tyrosine within the kinase domain activation segment as a phosphorylation site. Kinetic analysis revealed a decreased affinity for peptide substrate upon substitution of activation segment or juxtamembrane tyrosines. Together, our data suggest that the catalytic and therefore biological activities of Eph receptors are controlled by a two component inhibitory mechanism, which is released by phosphorylation of the juxtamembrane and activation segment tyrosine residues. PMID- 10848606 TI - Identification of amino acid residues in the Caenorhabditis elegans POU protein UNC-86 that mediate UNC-86-MEC-3-DNA ternary complex formation. AB - The POU homeodomain protein UNC-86 and the LIM homeodomain protein MEC-3 are essential for the differentiation of the six mechanoreceptor neurons in the nematode Caenorhabditis elegans. Previous studies have indicated that UNC-86 and MEC-3 bind cooperatively to at least three sites in the mec-3 promoter and synergistically activate transcription. However, the molecular details of the interactions of UNC-86 with MEC-3 and DNA have not been investigated so far. Here we used a yeast system to identify the functional domains in UNC-86 required for transcriptional activation and to characterize the interaction of UNC-86 with MEC 3 in vivo. Our results suggest that transcriptional activation is mediated by the amino terminus of UNC-86, whereas amino acids in the POU domain mediate DNA binding and interaction with MEC-3. By random mutagenesis, we identified mutations that only affect the DNA binding properties of UNC-86, as well as mutations that prevent coactivation by MEC-3. We demonstrated that both the POU specific domain and the homeodomain of UNC-86, as well as DNA bases adjacent to the proposed UNC-86 binding site, are involved in the formation of a transcriptionally active complex with MEC-3. These data suggest that some residues involved in the contact of UNC-86 with MEC-3 also contribute to the interaction of the functionally nonrelated POU protein Oct-1 with Oca-B, whereas other positions have different roles. PMID- 10848607 TI - Assembly of a functional beta interferon enhanceosome is dependent on ATF-2-c-jun heterodimer orientation. AB - Heterodimeric transcription factors, including the basic region-leucine zipper (bZIP) protein ATF-2-c-jun, are well-characterized components of an enhanceosome that mediates virus induction of the human beta interferon (IFN-beta) gene. Here we report that within the IFN-beta enhanceosome the ATF-2-c-jun heterodimer binds in a specific orientation, which is required for assembly of a complex between ATF-2-c-jun and interferon regulatory factor 3 (IRF-3). We demonstrate that correct orientation of the ATF-2-c-jun binding site is required for virus induction of the IFN-beta gene and for IRF-3-dependent activation of a composite ATF-2- c-jun-IRF site in the IFN-beta promoter. We also show that in vitro the DNA-bound ATF-2-c-jun heterodimer adopts a fixed orientation upon the binding of IRF-3 at an adjacent site in the IFN-beta enhancer and that the DNA-binding domain of IRF-3 is sufficient to mediate this effect. In addition, we show that the DNA-binding domain of ATF-2 is necessary and sufficient for selective protein protein interactions with IRF-3. Strikingly, in vivo chromatin immunoprecipitation experiments with IFN-beta reporter constructs reveal that recruitment of IRF-3 to the IFN-beta promoter upon virus infection is dependent on the orientation of the ATF-2-c-jun heterodimer binding site. These observations demonstrate functional and physical cooperativity between the bZIP and IRF transcription factor families and illustrate the critical role of heterodimeric transcription factors in formation of the IFN-beta enhanceosome. PMID- 10848608 TI - Specific protein-protein interaction between basic helix-loop-helix transcription factors and homeoproteins of the Pitx family. AB - Homeoproteins and basic helix-loop-helix (bHLH) transcription factors are known for their critical role in development and cellular differentiation. The pituitary pro-opiomelanocortin (POMC) gene is a target for factors of both families. Indeed, pituitary-specific transcription of POMC depends on the action of the homeodomain-containing transcription factor Pitx1 and of bHLH heterodimers containing NeuroD1. We now show lineage-restricted expression of NeuroD1 in pituitary corticotroph cells and a direct physical interaction between bHLH heterodimers and Pitx1 that results in transcriptional synergism. The interaction between the bHLH and homeodomains is restricted to ubiquitous (class A) bHLH and to the Pitx subfamily. Since bHLH heterodimers interact with Pitx factors through their ubiquitous moiety, this mechanism may be implicated in other developmental processes involving bHLH factors, such as neurogenesis and myogenesis. PMID- 10848609 TI - Bypass of a meiotic checkpoint by overproduction of meiotic chromosomal proteins. AB - The Saccharomyces cerevisiae zip1 mutant, which exhibits defects in synaptonemal complex formation and meiotic recombination, triggers a checkpoint that causes cells to arrest at the pachytene stage of meiotic prophase. Overproduction of either the meiotic chromosomal protein Red1 or the meiotic kinase Mek1 bypasses this checkpoint, allowing zip1 cells to sporulate. Red1 or Mek1 overproduction also promotes sporulation of other mutants (zip2, dmc1, hop2) that undergo checkpoint-mediated arrest at pachytene. In addition, Red1 overproduction antagonizes interhomolog interactions in the zip1 mutant, substantially decreasing double-strand break formation, meiotic recombination, and homologous chromosome pairing. Mek1 overproduction, in contrast, suppresses checkpoint induced arrest without significantly decreasing meiotic recombination. Cooverproduction of Red1 and Mek1 fails to bypass the checkpoint; moreover, overproduction of the meiotic chromosomal protein Hop1 blocks the Red1 and Mek1 overproduction phenotypes. These results suggest that meiotic chromosomal proteins function in the signaling of meiotic prophase defects and that the correct stoichiometry of Red1, Mek1, and Hop1 is needed to achieve checkpoint mediated cell cycle arrest at pachytene. PMID- 10848610 TI - p53 recruitment of CREB binding protein mediated through phosphorylated CREB: a novel pathway of tumor suppressor regulation. AB - CREB binding protein (CBP) is a 270-kDa nuclear protein required for activated transcription of a large number of cellular genes. Although CBP was originally discovered through its interaction with phosphorylated CREB (pCREB), it is utilized by a multitude of cellular transcription factors and viral oncoproteins. Both CREB and the tumor suppressor p53 have been shown to directly interact with the KIX domain of CBP. Although coactivator competition is an emerging theme in transcriptional regulation, we have made the fortuitous observation that protein kinase A-phosphorylated CREB strongly enhances p53 association with KIX. Phosphorylated CREB also facilitates interaction of a p53 mutant, defective for KIX binding, indicating that CREB functions in a novel way to bridge p53 and the coactivator. This is accomplished through direct interaction between the bZIP domain of CREB and the amino terminus of p53; a protein-protein interaction that is also detected in vivo. Consistent with our biochemical observations, we show that stimulation of the intracellular cyclic AMP (cAMP) pathway, which leads to CREB phosphorylation, strongly enhances both the transcriptional activation and apoptotic properties of p53. We propose that phosphorylated CREB mediates recruitment of CBP to p53-responsive promoters through direct interaction with p53. These observations provide evidence for a novel pathway that integrates cAMP signaling and p53 transcriptional activity. PMID- 10848611 TI - Arginine N-methyltransferase 1 is required for early postimplantation mouse development, but cells deficient in the enzyme are viable. AB - Protein arginine N-methyltransferases have been implicated in a variety of processes, including cell proliferation, signal transduction, and protein trafficking. In this study, we have characterized essentially a null mutation induced by insertion of the U3betaGeo gene trap retrovirus into the second intron of the mouse protein arginine N-methyltransferase 1 gene (Prmt1). cDNAs encoding two forms of Prmt1 were characterized, and the predicted protein sequences were found to be highly conserved among vertebrates. Expression of the Prmt1-betageo fusion gene was greatest along the midline of the neural plate and in the forming head fold from embryonic day 7.5 (E7.5) to E8.5 and in the developing central nervous system from E8.5 to E13.5. Homozygous mutant embryos failed to develop beyond E6.5, a phenotype consistent with a fundamental role in cellular metabolism. However, Prmt1 was not required for cell viability, as the protein was not detected in embryonic stem (ES) cell lines established from mutant blastocysts. Low levels of Prmt1 transcripts (approximately 1% of the wild-type level) were detected as assessed by a quantitative reverse transcription-PCR assay. Total levels of arginine N-methyltransferase activity and asymmetric N(G), N(G)-dimethylarginine were reduced by 85 and 54%, respectively, while levels of hypomethylated substrates were increased 15-fold. Prmt1 appears to be a major type I enzyme in ES cells, and in wild-type cells, most substrates of the enzyme appear to be maintained in a fully methylated state. PMID- 10848612 TI - Isoform-specific localization of A-RAF in mitochondria. AB - RAF kinase is a family of isoforms including A-RAF, B-RAF, and C-RAF. Despite the important role of RAF in cell growth and proliferation, little evidence exists for isoform-specific function of RAF family members. Using Western analysis and immunogold labeling, A-RAF was selectively localized in highly purified rat liver mitochondria. Two novel human proteins, which interact specifically with A-RAF, were identified, and the full-length sequences are reported. These proteins, referred to as hTOM and hTIM, are similar to components of mitochondrial outer and inner membrane protein-import receptors from lower organisms, implicating their involvement in the mitochondrial transport of A-RAF. hTOM contains multiple tetratricopeptide repeat (TPR) domains, which function in protein-protein interactions. TPR domains are frequently present in proteins involved in cellular transport systems. In contrast, protein 14-3-3, an abundant cytosolic protein that participates in many facets of signal transduction, was found to interact with C-RAF but not with A-RAF N-terminal domain. This information is discussed in view of the important role of mitochondria in cellular functions involving energy balance, proliferation, and apoptosis and the potential role of A-RAF in regulating these systems. PMID- 10848613 TI - Developmental expression of latent transforming growth factor beta binding protein 2 and its requirement early in mouse development. AB - Latent transforming growth factor beta (TGF-beta) binding protein 2 (LTBP-2) is an integral component of elastin-containing microfibrils. We studied the expression of LTBP-2 in the developing mouse and rat by in situ hybridization, using tropoelastin expression as a marker of tissues participating in elastic fiber formation. LTBP-2 colocalized with tropoelastin within the perichondrium, lung, dermis, large arterial vessels, epicardium, pericardium, and heart valves at various stages of rodent embryonic development. Both LTBP-2 and tropoelastin expression were seen throughout the lung parenchyma and within the cortex of the spleen in the young adult mouse. In the testes, LTBP-2 expression was seen within lumenal cells of the epididymis in the absence of tropoelastin. Collectively, these results imply that LTBP-2 plays a structural role within elastic fibers in most cases. To investigate its importance in development, mice with a targeted disruption of the Ltbp2 gene were generated. Ltbp2(-/-) mice die between embryonic day 3.5 (E3.5) and E6.5. LTBP-2 expression was not detected by in situ hybridization in E6.5 embryos but was detected in E3.5 blastocysts by reverse transcription-PCR. These results are not consistent with the phenotypes of TGF beta knockout mice or mice with knockouts of other elastic fiber proteins, implying that LTBP-2 performs a yet undiscovered function in early development, perhaps in implantation. PMID- 10848614 TI - Nuclear entry of the circadian regulator mPER1 is controlled by mammalian casein kinase I epsilon. AB - The molecular oscillator that keeps circadian time is generated by a negative feedback loop. Nuclear entry of circadian regulatory proteins that inhibit transcription from E-box-containing promoters appears to be a critical component of this loop in both Drosophila and mammals. The Drosophila double-time gene product, a casein kinase I epsilon (CKIepsilon) homolog, has been reported to interact with dPER and regulate circadian cycle length. We find that mammalian CKIepsilon binds to and phosphorylates the murine circadian regulator mPER1. Unlike both dPER and mPER2, mPER1 expressed alone in HEK 293 cells is predominantly a nuclear protein. Two distinct mechanisms appear to retard mPER1 nuclear entry. First, coexpression of mPER2 leads to mPER1-mPER2 heterodimer formation and cytoplasmic colocalization. Second, coexpression of CKIepsilon leads to masking of the mPER1 nuclear localization signal and phosphorylation dependent cytoplasmic retention of both proteins. CKIepsilon may regulate mammalian circadian rhythm by controlling the rate at which mPER1 enters the nucleus. PMID- 10848615 TI - Set domain-dependent regulation of transcriptional silencing and growth control by SUV39H1, a mammalian ortholog of Drosophila Su(var)3-9. AB - Mammalian SET domain-containing proteins define a distinctive class of chromatin associated factors that are targets for growth control signals and oncogenic activation. SUV39H1, a mammalian ortholog of Drosophila Su(var)3-9, contains both SET and chromo domains, signature motifs for proteins that contribute to epigenetic control of gene expression through effects on the regional organization of chromatin structure. In this report we demonstrate that SUV39H1 represses transcription in a transient transcriptional assay when tethered to DNA through the GAL4 DNA binding domain. Under these conditions, SUV39H1 displays features of a long-range repressor capable of acting over several kilobases to silence basal promoters. A possible role in chromatin-mediated gene silencing is supported by the localization of exogenously expressed SUV39H1 to nuclear bodies with morphologic features suggestive of heterochromatin in interphase cells. In addition, we show that SUV39H1 is phosphorylated specifically at the G(1)/S cell cycle transition and when forcibly expressed suppresses cell growth. Growth suppression as well as the ability of SUV39H1 to form nuclear bodies and silence transcription are antagonized by the oncogenic antiphosphatase Sbf1 that when hyperexpressed interacts with the SET domain and stabilizes the phosphorylated form of SUV39H1. These studies suggest a phosphorylation-dependent mechanism for regulating the chromatin organizing activity of a mammalian su(var) protein and implicate the SET domain as a gatekeeper motif that integrates upstream signaling pathways to epigenetic regulation and growth control. PMID- 10848616 TI - Orphan receptor DAX-1 is a shuttling RNA binding protein associated with polyribosomes via mRNA. AB - The DAX-1 (NR0B1) gene encodes an unusual member of the nuclear hormone receptor superfamily which acts as a transcriptional repressor. Mutations in the human DAX 1 gene cause X-linked adrenal hypoplasia congenita (AHC) associated with hypogonadotropic hypogonadism (HHG). We have studied the intracellular localization of the DAX-1 protein in human adrenal cortex and mouse Leydig tumor cells and found it to be both nuclear and cytoplasmic. A significant proportion of DAX-1 is associated with polyribosomes and is found complexed with polyadenylated RNA. DAX-1 directly binds to RNA, two domains within the protein being responsible for cooperative binding activity and specificity. Mutations in DAX-1 found in AHC-HHG patients significantly impair RNA binding. These findings reveal that DAX-1 plays multiple regulatory roles at the transcriptional and posttranscriptional levels. PMID- 10848617 TI - Herpes simplex virus type 1 entry into host cells: reconstitution of capsid binding and uncoating at the nuclear pore complex in vitro. AB - During entry, herpes simplex virus type 1 (HSV-1) releases its capsid and the tegument proteins into the cytosol of a host cell by fusing with the plasma membrane. The capsid is then transported to the nucleus, where it docks at the nuclear pore complexes (NPCs), and the viral genome is rapidly released into the nucleoplasm. In this study, capsid association with NPCs and uncoating of the viral DNA were reconstituted in vitro. Isolated capsids prepared from virus were incubated with cytosol and purified nuclei. They were found to bind to the nuclear pores. Binding could be inhibited by pretreating the nuclei with wheat germ agglutinin, anti-NPC antibodies, or antibodies against importin beta. Furthermore, in the absence of cytosol, purified importin beta was both sufficient and necessary to support efficient capsid binding to nuclei. Up to 60 to 70% of capsids interacting with rat liver nuclei in vitro released their DNA if cytosol and metabolic energy were supplied. Interaction of the capsid with the nuclear pore thus seemed to trigger the release of the viral genome, implying that components of the NPC play an active role in the nuclear events during HSV-1 entry into host cells. PMID- 10848618 TI - Amino- and carboxy-terminal PEST domains mediate gastrin stabilization of rat L histidine decarboxylase isoforms. AB - Control of enzymatic function by peptide hormones can occur at a number of different levels and can involve diverse pathways that regulate cleavage, intracellular trafficking, and protein degradation. Gastrin is a peptide hormone that binds to the cholecystokinin B-gastrin receptor and regulates the activity of L-histidine decarboxylase (HDC), the enzyme that produces histamine. Here we show that gastrin can increase the steady-state levels of at least six HDC isoforms without affecting HDC mRNA levels. Pulse-chase experiments indicated that HDC isoforms are rapidly degraded and that gastrin-dependent increases are due to enhanced isoform stability. Deletion analysis identified two PEST domains (PEST1 and PEST2) and an intracellular targeting domain (ER2) which regulate HDC protein expression levels. Experiments with PEST domain fusion proteins demonstrated that PEST1 and PEST2 are strong and portable degradation-promoting elements which are positively regulated by both gastrin stimulation and proteasome inhibition. A chimeric protein containing the PEST domain of ornithine decarboxylase was similarly affected, indicating that gastrin can regulate the stability of other PEST domain-containing proteins and does so independently of antizyme/antizyme inhibitor regulation. At the same time, endoplasmic reticulum localization of a fluorescent chimera containing the ER2 domain of HDC was unaltered by gastrin stimulation. We conclude that gastrin stabilization of HDC isoforms is dependent upon two transferable and sequentially unrelated PEST domains that regulate degradation. These experiments revealed a novel regulatory mechanism by which a peptide hormone such as gastrin can disrupt the degradation function of multiple PEST-domain-containing proteins. PMID- 10848619 TI - Ribosomal DNA replication fork barrier and HOT1 recombination hot spot: shared sequences but independent activities. AB - In the ribosomal DNA of Saccharomyces cerevisiae, sequences in the nontranscribed spacer 3' of the 35S ribosomal RNA gene are important to the polar arrest of replication forks at a site called the replication fork barrier (RFB) and also to the cis-acting, mitotic hyperrecombination site called HOT1. We have found that the RFB and HOT1 activity share some but not all of their essential sequences. Many of the mutations that reduce HOT1 recombination also decrease or eliminate fork arrest at one of two closely spaced RFB sites, RFB1 and RFB2. A simple model for the juxtaposition of RFB and HOT1 sequences is that the breakage of strands in replication forks arrested at RFB stimulates recombination. Contrary to this model, we show here that HOT1-stimulated recombination does not require the arrest of forks at the RFB. Therefore, while HOT1 activity is independent of replication fork arrest, HOT1 and RFB require some common sequences, suggesting the existence of a common trans-acting factor(s). PMID- 10848620 TI - Intracellular transport, assembly, and degradation of wild-type and disease linked mutant gap junction proteins. AB - More than 130 different mutations in the gap junction integral plasma membrane protein connexin32 (Cx32) have been linked to the human peripheral neuropathy X linked Charcot-Marie-Tooth disease (CMTX). How these various mutants are processed by the cell and the mechanism(s) by which they cause CMTX are unknown. To address these issues, we have studied the intracellular transport, assembly, and degradation of three CMTX-linked Cx32 mutants stably expressed in PC12 cells. Each mutant had a distinct fate: E208K Cx32 appeared to be retained in the endoplasmic reticulum (ER), whereas both the E186K and R142W mutants were transported to perinuclear compartments from which they trafficked either to lysosomes (R142W Cx32) or back to the ER (E186K Cx32). Despite these differences, each mutant was soluble in nonionic detergent but unable to assemble into homomeric connexons. Degradation of both mutant and wild-type connexins was rapid (t(1/2) < 3 h) and took place at least in part in the ER by a process sensitive to proteasome inhibitors. The mutants studied are therefore unlikely to cause disease by accumulating in degradation-resistant aggregates but instead are efficiently cleared from the cell by quality control processes that prevent abnormal connexin molecules from traversing the secretory pathway. PMID- 10848621 TI - The Kex2p proregion is essential for the biosynthesis of an active enzyme and requires a C-terminal basic residue for its function. AB - The Saccharomyces cerevisiae prohormone-processing enzyme Kex2p is biosynthesized as an inactive precursor extended by its N-terminal proregion. Here we show that deletion of the proregion renders Kex2p inactive both in vivo and in vitro. Absence of the proregion impaired glycosylation and stability and resulted in the retention of the enzyme in the endoplasmic reticulum. These phenotypes were partially complemented by expression of the proregion in trans. Trans complementation was specific to Kex2p proregion because expression of any of the seven mammalian prohormone convertase propeptides had no effect. These data are consistent with a model whereby Kex2p proregion functions as an intramolecular chaperone and indicate that covalent linkage to the protein is not an absolute requirement for proregion function. Furthermore, extensive mutagenesis revealed that, in addition to their function as proteolytic recognition sites, C-terminal basic residues play an active role in proregion-dependent Kex2p activation. PMID- 10848622 TI - Proinsulin endoproteolysis confers enhanced targeting of processed insulin to the regulated secretory pathway. AB - Recently, two different prohormone-processing enzymes, prohormone convertase 1 (PC1) and carboxypeptidase E, have been implicated in enhancing the storage of peptide hormones in endocrine secretory granules. It is important to know the extent to which such molecules may act as "sorting receptors" to allow the selective trafficking of cargo proteins from the trans-Golgi network into forming granules, versus acting as enzymes that may indirectly facilitate intraluminal storage of processed hormones within maturing granules. GH4C1 cells primarily store prolactin in granules; they lack PC1 and are defective for intragranular storage of transfected proinsulin. However, proinsulin readily enters the immature granules of these cells. Interestingly, GH4C1 clones that stably express modest levels of PC1 store more proinsulin-derived protein in granules. Even in the presence of PC1, a sizable portion of the proinsulin that enters granules goes unprocessed, and this portion largely escapes granule storage. Indeed, all of the increased granule storage can be accounted for by the modest portion converted to insulin. These results are not unique to GH4C1 cells; similar results are obtained upon PC1 expression in PC12 cells as well as in AtT20 cells (in which PC1 is expressed endogenously at higher levels). An in vitro assay of protein solubility indicates a difference in the biophysical behavior of proinsulin and insulin in the PC1 transfectants. We conclude that processing to insulin, facilitated by the catalytic activities of granule proteolytic enzymes, assists in the targeting (storage) of the hormone. PMID- 10848623 TI - alpha5beta1 integrin protects intestinal epithelial cells from apoptosis through a phosphatidylinositol 3-kinase and protein kinase B-dependent pathway. AB - Renewal of the gastrointestinal epithelium involves a coordinated process of terminal differentiation and programmed cell death. Integrins have been implicated in the control of apoptotic processes in various cell types. Here we examine the role of integrins in the regulation of apoptosis in gastrointestinal epithelial cells with the use of a rat small intestinal epithelial cell line (RIE1) as a model. Overexpression of the integrin alpha5 subunit in RIE1 cells conferred protection against several proapoptotic stimuli. In contrast, overexpression of the integrin alpha2 subunit had no effect on cell survival. The antiapoptotic effect of the alpha5 subunit was partially retained by a mutated version that had a truncation of the cytoplasmic domain. The antiapoptotic effects of the full-length or truncated alpha5 subunit were reversed upon treatment with inhibitors of phosphatidylinositol 3-kinase (PI-3-kinase), suggesting that the alpha5beta1 integrin might interact with the PI-3-kinase/Akt survival pathway. When cells overexpressing alpha5 were allowed to adhere to fibronectin, there was a moderate activation of protein kinase B (PKB)/Akt, whereas no such effect was seen in alpha2-overexpressing cells adhering to collagen. Furthermore, in cells overexpressing alpha5 and adhering to fibronectin, there was a dramatic enhancement of the ability of growth factors to stimulate PKB/Akt; again, this was not seen in cells overexpressing alpha2 subunit and adhering to collagen or fibronectin. Expression of a dominant negative version of PKB/Akt in RIE cells blocked to ability of alpha5 to enhance cell survival. Thus, the alpha5beta1 integrin seems to protect intestinal epithelial cells against proapoptotic stimuli by selectively enhancing the activity of the PI-3-kinase/Akt survival pathway. PMID- 10848624 TI - Identification of a novel family of nonclassic yeast phosphatidylinositol transfer proteins whose function modulates phospholipase D activity and Sec14p independent cell growth. AB - Yeast phosphatidylinositol transfer protein (Sec14p) is essential for Golgi function and cell viability. We now report a characterization of five yeast SFH (Sec Fourteen Homologue) proteins that share 24-65% primary sequence identity with Sec14p. We show that Sfh1p, which shares 64% primary sequence identity with Sec14p, is nonfunctional as a Sec14p in vivo or in vitro. Yet, SFH proteins sharing low primary sequence similarity with Sec14p (i.e., Sfh2p, Sfh3p, Sfh4p, and Sfh5p) represent novel phosphatidylinositol transfer proteins (PITPs) that exhibit phosphatidylinositol- but not phosphatidylcholine-transfer activity in vitro. Moreover, increased expression of Sfh2p, Sfh4p, or Sfh5p rescues sec14 associated growth and secretory defects in a phospholipase D (PLD)-sensitive manner. Several independent lines of evidence further demonstrate that SFH PITPs are collectively required for efficient activation of PLD in vegetative cells. These include a collective requirement for SFH proteins in Sec14p-independent cell growth and in optimal activation of PLD in Sec14p-deficient cells. Consistent with these findings, Sfh2p colocalizes with PLD in endosomal compartments. The data indicate that SFH gene products cooperate with "bypass Sec14p" mutations and PLD in a complex interaction through which yeast can adapt to loss of the essential function of Sec14p. These findings expand the physiological repertoire of PITP function in yeast and provide the first in vivo demonstration of a role for specific PITPs in stimulating activation of PLD. PMID- 10848625 TI - Interaction of the tau2 transcriptional activation domain of glucocorticoid receptor with a novel steroid receptor coactivator, Hic-5, which localizes to both focal adhesions and the nuclear matrix. AB - Hic-5 (hydrogen peroxide-inducible clone-5) is a focal adhesion protein that is involved in cellular senescence. In the present study, a yeast two-hybrid screen identified Hic-5 as a protein that interacts with a region of the glucocorticoid receptor that includes a nuclear matrix-targeting signal and the tau2 transcriptional activation domain. In transiently transfected mammalian cells, overexpression of Hic-5 potentiated the activation of reporter genes by all steroid receptors, excluding the estrogen receptor. The activity of the estrogen receptor and the thyroid hormone receptor was stimulated by Hic-5 in the presence but not in the absence of coexpressed coactivator GRIP1. In biochemical fractionations and indirect immunofluorescence assays, a fraction of endogenous Hic-5 in REF-52 cells and transiently expressed Hic-5 in Cos-1 cells was associated with the nuclear matrix. The C-terminal region of Hic-5, which contains seven zinc fingers arranged in four LIM domains, was required for interaction with focal adhesions, the nuclear matrix, steroid receptors, and the tau2 domain of glucocorticoid receptor. The N-terminal region of Hic-5 possesses a transcriptional activation domain and was essential for the coactivator activity of Hic-5. Given the coexisting cytoplasmic and nuclear distributions of Hic-5 and its role in steroid receptor-mediated transcriptional activation, it is proposed that Hic-5 might transmit signals that emanate at cell attachment sites and regulate transcription factors, such as steroid receptors. PMID- 10848626 TI - Inactivation of lmpA, encoding a LIMPII-related endosomal protein, suppresses the internalization and endosomal trafficking defects in profilin-null mutants. AB - Profilin is a key phosphoinositide and actin-binding protein connecting and coordinating changes in signal transduction pathways with alterations in the actin cytoskeleton. Using biochemical assays and microscopic approaches, we demonstrate that profilin-null cells are defective in macropinocytosis, fluid phase efflux, and secretion of lysosomal enzymes but are unexpectedly more efficient in phagocytosis than wild-type cells. Disruption of the lmpA gene encoding a protein (DdLIMP) belonging to the CD36/LIMPII family suppressed, to different degrees, most of the profilin-minus defects, including the increase in F-actin, but did not rescue the secretion defect. Immunofluorescence microscopy indicated that DdLIMP, which is also capable of binding phosphoinositides, was associated with macropinosomes but was not detected in the plasma membrane. Also, inactivation of the lmpA gene in wild-type strains resulted in defects in macropinocytosis and fluid phase efflux but not in phagocytosis. These results suggest an important role for profilin in regulating the internalization of fluid and particles and the movement of material along the endosomal pathway; they also demonstrate a functional interaction between profilin and DdLIMP that may connect phosphoinositide-based signaling through the actin cytoskeleton with endolysosomal membrane trafficking events. PMID- 10848627 TI - The MAL proteolipid is necessary for the overall apical delivery of membrane proteins in the polarized epithelial Madin-Darby canine kidney and fischer rat thyroid cell lines. AB - The MAL proteolipid has been recently demonstrated as being necessary for correct apical sorting of the transmembrane influenza virus hemagglutinin (HA) in Madin Darby canine kidney (MDCK) cells. The fact that, in contrast to MDCK cells, Fischer rat thyroid (FRT) cells target the majority of glycosylphosphatidylinositol (GPI)-anchored proteins to the basolateral membrane provides us with the opportunity to determine the role of MAL in apical transport of membrane proteins under conditions in which the majority of GPI-anchored proteins are (MDCK cells) or are not (FRT cells) targeted to the apical surface. Using an antisense oligonucleotide-based strategy to deplete endogenous MAL, we have observed that correct transport of apical transmembrane proteins associated (HA) or not (exogenous neurotrophin receptor and endogenous dipeptidyl peptidase IV) with lipid rafts, as well as that of the bulk of endogenous apical membrane, takes place in FRT cells by a pathway that requires normal MAL levels. Even transport of placental alkaline phosphatase, a GPI-anchored protein that is targeted apically in FRT cells, was dependent on normal MAL levels. Similarly, in addition to the reported effect of MAL on HA transport, depletion of MAL in MDCK cells caused a dramatic reduction in the apical delivery of the GPI-anchored gD1 DAF protein, neurotrophin receptor, and the bulk of membrane proteins. These results suggest that MAL is necessary for the overall apical transport of membrane proteins in polarized MDCK and FRT cells. PMID- 10848628 TI - Cytoplasmic dynein-mediated assembly of pericentrin and gamma tubulin onto centrosomes. AB - Centrosome assembly is important for mitotic spindle formation and if defective may contribute to genomic instability in cancer. Here we show that in somatic cells centrosome assembly of two proteins involved in microtubule nucleation, pericentrin and gamma tubulin, is inhibited in the absence of microtubules. A more potent inhibitory effect on centrosome assembly of these proteins is observed after specific disruption of the microtubule motor cytoplasmic dynein by microinjection of dynein antibodies or by overexpression of the dynamitin subunit of the dynein binding complex dynactin. Consistent with these observations is the ability of pericentrin to cosediment with taxol-stabilized microtubules in a dynein- and dynactin-dependent manner. Centrosomes in cells with reduced levels of pericentrin and gamma tubulin have a diminished capacity to nucleate microtubules. In living cells expressing a green fluorescent protein-pericentrin fusion protein, green fluorescent protein particles containing endogenous pericentrin and gamma tubulin move along microtubules at speeds of dynein and dock at centrosomes. In Xenopus extracts where gamma tubulin assembly onto centrioles can occur without microtubules, we find that assembly is enhanced in the presence of microtubules and inhibited by dynein antibodies. From these studies we conclude that pericentrin and gamma tubulin are novel dynein cargoes that can be transported to centrosomes on microtubules and whose assembly contributes to microtubule nucleation. PMID- 10848629 TI - Dynamic regulation of activated leukocyte cell adhesion molecule-mediated homotypic cell adhesion through the actin cytoskeleton. AB - Restricted expression of activated leukocyte cell adhesion molecule (ALCAM) by hematopoietic cells suggests an important role in the immune system and hematopoiesis. To get insight into the mechanisms that control ALCAM-mediated adhesion we have investigated homotypic ALCAM-ALCAM interactions. Here, we demonstrate that the cytoskeleton regulates ALCAM-mediated cell adhesion because inhibition of actin polymerization by cytochalasin D (CytD) strongly induces homotypic ALCAM-ALCAM interactions. This induction of cell adhesion is likely due to clustering of ALCAM at the cell surface, which is observed after CytD treatment. Single-particle tracking demonstrated that the lateral mobility of ALCAM in the cell membrane is increased 30-fold after CytD treatment. In contrast, both surface distribution and adhesion of a glycosylphosphatidylinositol (GPI)-anchored ALCAM mutant are insensitive to CytD, despite the increase in lateral mobility of GPI-ALCAM upon CytD treatment. This demonstrates that clustering of ALCAM is essential for cell adhesion, whereas enhanced diffusion of ALCAM alone is not sufficient for cluster formation. In addition, upon ligand binding, both free diffusion and the freely dragged distance of wild-type ALCAM, but not of GPI-ALCAM, are reduced over time, suggesting strengthening of the cytoskeleton linkage. From these findings we conclude that activation of ALCAM-mediated adhesion is dynamically regulated through actin cytoskeleton-dependent clustering. PMID- 10848630 TI - Histone deacetylase inhibitors trigger a G2 checkpoint in normal cells that is defective in tumor cells. AB - Important aspects of cell cycle regulation are the checkpoints, which respond to a variety of cellular stresses to inhibit cell cycle progression and act as protective mechanisms to ensure genomic integrity. An increasing number of tumor suppressors are being demonstrated to have roles in checkpoint mechanisms, implying that checkpoint dysfunction is likely to be a common feature of cancers. Here we report that histone deacetylase inhibitors, in particular azelaic bishydroxamic acid, triggers a G2 phase cell cycle checkpoint response in normal human cells, and this checkpoint is defective in a range of tumor cell lines. Loss of this G2 checkpoint results in the tumor cells undergoing an aberrant mitosis resulting in fractured multinuclei and micronuclei and eventually cell death. This histone deacetylase inhibitor-sensitive checkpoint appears to be distinct from G2/M checkpoints activated by genotoxins and microtubule poisons and may be the human homologue of a yeast G2 checkpoint, which responds to aberrant histone acetylation states. Azelaic bishydroxamic acid may represent a new class of anticancer drugs with selective toxicity based on its ability to target a dysfunctional checkpoint mechanism in tumor cells. PMID- 10848631 TI - The peroxin pex3p initiates membrane assembly in peroxisome biogenesis. AB - Rat cDNA encoding a 372-amino-acid peroxin was isolated, primarily by functional complementation screening, using a peroxisome-deficient Chinese hamster ovary cell mutant, ZPG208, of complementation group 17. The deduced primary sequence showed approximately 25% amino acid identity with the yeast Pex3p, thereby we termed this cDNA rat PEX3 (RnPEX3). Human and Chinese hamster Pex3p showed 96 and 94% identity to rat Pex3p and had 373 amino acids. Pex3p was characterized as an integral membrane protein of peroxisomes, exposing its N- and C-terminal parts to the cytosol. A homozygous, inactivating missense mutation, G to A at position413, in a codon (GGA) for Gly(138) and resulting in a codon (GAA) for Glu was the genetic cause of peroxisome deficiency of complementation group 17 ZPG208. The peroxisome-restoring activity apparently required the full length of Pex3p, whereas its N-terminal part from residues 1 to 40 was sufficient to target a fusion protein to peroxisomes. We also demonstrated that Pex3p binds the farnesylated peroxisomal membrane protein Pex19p. Moreover, upon expression of PEX3 in ZPG208, peroxisomal membrane vesicles were assembled before the import of soluble proteins such as PTS2-tagged green fluorescent protein. Thus, Pex3p assembles membrane vesicles before the matrix proteins are translocated. PMID- 10848632 TI - Mitochondria-to-nuclear signaling is regulated by the subcellular localization of the transcription factors Rtg1p and Rtg3p. AB - Cells modulate the expression of nuclear genes in response to changes in the functional state of mitochondria, an interorganelle communication pathway called retrograde regulation. In yeast, expression of the CIT2 gene shows a typical retrograde response in that its expression is dramatically increased in cells with dysfunctional mitochondria, such as in rho(o) petites. Three genes control this signaling pathway: RTG1 and RTG3, which encode basic helix-loop-helix leucine zipper transcription factors that bind as heterodimer to the CIT2 upstream activation site, and RTG2, which encodes a protein of unknown function. We show that in respiratory-competent (rho(+)) cells in which CIT2 expression is low, Rtg1p and Rtg3p exist as a complex largely in the cytoplasm, and in rho(o) petites in which CIT2 expression is high, they exist as a complex predominantly localized in the nucleus. Cytoplasmic Rtg3p is multiply phosphorylated and becomes partially dephosphorylated when localized in the nucleus. Rtg2p, which is cytoplasmic in both rho(+) and rho(o) cells, is required for the dephosphorylation and nuclear localization of Rtg3p. Interaction of Rtg3p with Rtg1p is required to retain Rtg3p in the cytoplasm of rho(+) cells; in the absence of such interaction, nuclear localization and dephosphorylation of Rtg3p is independent of Rtg2p. Our data show that Rtg1p acts as both a positive and negative regulator of the retrograde response and that Rtg2p acts to transduce mitochondrial signals affecting the phosphorylation state and subcellular localization of Rtg3p. PMID- 10848634 TI - Sorting of membrane and fluid at the apical pole of polarized Madin-Darby canine kidney cells. AB - When fluid-phase markers are internalized from opposite poles of polarized Madin Darby canine kidney cells, they accumulate in distinct apical and basolateral early endosomes before meeting in late endosomes. Recent evidence suggests that significant mixing of apically and basolaterally internalized membrane proteins occurs in specialized apical endosomal compartments, including the common recycling endosome and the apical recycling endosome (ARE). The relationship between these latter compartments and the fluid-labeled apical early endosome is unknown at present. We report that when the apical recycling marker, membrane bound immunoglobulin A (a ligand for the polymeric immunoglobulin receptor), and fluid-phase dextran are cointernalized from the apical poles of Madin-Darby canine kidney cells, they enter a shared apical early endosome ( or =1 week apart) to exercise continuously at 65% of peak aerobic capacity for 60 min followed by the recovery without (experiment 1) or with phenylephrine infusion (experiment 2) to counteract post-exercise hypotension. Heart rate, arterial pressure (Finapres), plasma volume (PV, Evans blue dye dilution), haematocrit, haemoglobin, plasma total solutes (refractometer), albumin, total proteins (colorimetric method), [Na+] and [K+] were not different prior to the experiments. Exercise decreased PV -13.7% (-521 mL) and -14.2% (-566 mL) at the end of 60 min in experiments 1 and 2, respectively, associated with increases in the concentrations of plasma albumin, total protein and solutes. These changes were similar between the two experiments. Following 30 min recovery in experiment 1 the decreased PV was not significantly different from the baseline. Although the volume restoration was complete at the end of 90 min recovery, the change in the albumin concentration was still above zero, indicating a gain of 11 g albumin (P < 0.05). When phenylephrine was infused during recovery, there was no gain in intravascular albumin associated with a sustained decrease in PV (-7% or -280 mL, P < 0.05) observed at the end of experiment 2. These data suggest that post-exercise hypotension may be the mechanism for a gain of intravascular albumin via the lymph return, which enhances plasma water retention and PV restoration during recovery from exercise induced hypovolaemia, even without rehydration. PMID- 10848642 TI - Relationships between muscle morphology and insulin sensitivity are improved after adjustment for intra-individual variability in 70-year-old men. AB - The purpose of this investigation was to examine to what extent variability in the muscle morphology and insulin sensitivity influence the correlation between them. Reproducibility of muscle characteristics was estimated in duplicate biopsies from the same thigh of 23 subjects from a cohort of 70-year-old men. The coefficient of variation (CV) for different characteristics of muscle morphology was between 11 and 42% in duplicate biopsies. Coefficient of variation for markers of insulin sensitivity ranged between 12 and 39%. The variability reflected by intra-class correlation ranged from 0.23 to 0.60 for muscle morphology and from 0.68 to 0.96 for estimates of insulin sensitivity. The correlation analysis between muscle morphology and insulin resistance was performed in a sample of 515 men from the cohort, correlation coefficients were calculated with (rtrue) and without (r) adjustment for intra-individual variation. Insulin sensitivity showed a positive relationship with percentage of type I fibres (rtrue=0.33, r=0.21; P < 0.0001) and capillary density (rtrue=0.43, r=0.21; P < 0. 0001) and negative correlations with percentage of type IIB fibres (rtrue=-0.35, r=-0.24; P < 0.0001). Capillary density was inversely correlated to insulin. Thus, an obvious improvement of the correlation was seen after correcting intra-individual variation. In conclusion, owing to the low degree of reproducibility of muscle morphology variables and insulin sensitivity, implying a noticeable underestimation of correlations, the r-values should be adjusted for within-subject variation in order to demonstrate a more accurate estimate of the strength of the relationships studied. PMID- 10848643 TI - Calorie restriction increases insulin-stimulated tyrosine phosphorylation of insulin receptor and insulin receptor substrate-1 in rat skeletal muscle. AB - A moderate reduction in calorie intake (calorie restriction, CR) improves insulin stimulated glucose transport in skeletal muscle. Therefore, we studied muscle insulin signalling in ad libitum (AL) and CR ( approximately 60% AL intake for 20 days) fed rats, which received a control injection (sterile water) or an insulin injection (30 U kg-1 body weight). In control (not insulin-treated) rats, there was no detectable tyrosine phosphorylation of insulin receptor (IR), regardless of diet; no diet effect on tyrosine phosphorylation of insulin receptor substrate 1 (IRS1) or IRS1-associated phosphatidylinositol 3-kinase (PI3K) protein and 21% higher IRS1-associated PI3K activity in AL vs. CR. In insulin-treated rats, tyrosine-phosphorylated IR was 79% higher for CR vs. AL; tyrosine-phosphorylated IRS1 was 109% higher for CR vs. AL; IRS1-associated PI3K protein and IRS1 associated PI3K activity were unaffected by diet. Calorie restriction amplifies early insulin signalling steps without changing IRS1-associated PI3K, suggesting enhanced glucose transport is mediated by altering: IRS1-PI3K localization, PI3K associated with proteins other than IRS1 or post-PI3K events. PMID- 10848644 TI - The influence of nitric oxide on in vivo human skeletal muscle properties. AB - We have investigated the action of exogenous nitric oxide (NO) on the strength and contractile properties of human skeletal muscle working in vivo. Maximum isometric voluntary contraction force (MVC) of the quadriceps was measured and superimposed electrical stimulation was used to estimate the level of activation and 'true maximum force' (TMF). Force-frequency relationships were determined to assess changes in contractile properties of the muscle. Subjects in the experimental group (E, n=10) were measured before and during two separate periods of treatment with different doses of glyceryl trinitrate, a NO donor, delivering 100 (GTN100) or 200 (GTN200) microg h-1 as a trans-dermal patch. A control group (C, n=6) was measured during two similar periods whilst taking an oral placebo. There was a significant increase in strength with GTN200 (MVC: +5. 15%; TMF: +3.87%). There was no change in the strength of group C. There was a trend towards reduced forces at submaximal frequencies with GTN administration but the most notable change was a decline in twitch force (approximately 12%, P < 0.05) with GTN100 treatment and this remained depressed throughout the study. No changes were seen in the contractile properties of the control group C. The present results show that GTN treatment increased maximum voluntary strength but decreased twitch tension. The time course and dose-response characteristics indicate that these are two separate actions of NO on human muscle working in vivo. PMID- 10848645 TI - Relationship between decreased oxyhaemoglobin saturation and exhaled nitric oxide during exercise. AB - Decreases in oxyhaemoglobin saturation (SaO2) are frequently observed in highly trained male endurance athletes during heavy work and has been termed exercise induced hypoxaemia (EIH). Ventilation perfusion (VA/Q) mismatching and diffusion limitations are thought to be responsible. Nitric oxide (NO), a potent vasodilator, is present in the exhaled air of resting and exercising humans. Endogenously produced NO is thought to play a role in VA/Q matching and maintenance of low pulmonary vascular resistance. The purpose of this study was to determine the relationship between exhaled NO and EIH. It was hypothesized that athletes with EIH would have lower NO levels compared with non-EIH athletes. Eighteen highly trained male cyclists (VO2max=67.7 +/- 5.2 mL kg-1 min-1, mean +/ SD) were divided into normal (NORM, n=12, SaO2= 93.9 +/- 0.8) or low (LOW, n=6, SaO2=90.3 +/- 1.0) group, based on significantly different peak exercise SaO2 values (P < 0.05). All other descriptive and physiological characteristics were similar between the groups. Subjects performed a ramped cycle test to exhaustion breathing NO-free gas. The concentration (CNO) and production rate (VNO) of NO were determined from mixed gas samples at rest and during exercise at 100, 200, 250, 300, 350, 400 and 450 W using a chemiluminescent analyser. CNO remained unchanged from resting values in all subjects. VNO increased significantly during exercise in all subjects but was not different between LOW and NORM groups. The correlation between change in SaO2 and VNO from rest to maximal exercise was not significant (r=-0.12, P > 0.05). Collectively, these data suggest that exhaled NO is not related to decreased SaO2 during heavy exercise in highly trained male cyclists. PMID- 10848646 TI - The distribution of rest periods affects performance and adaptations of energy metabolism induced by high-intensity training in human muscle. AB - The effect of the distribution of rest periods on the efficacy of interval sprint training is analysed. Ten male subjects, divided at random into two groups, performed distinct incremental sprint training protocols, in which the muscle load was the same (14 sessions), but the distribution of rest periods was varied. The 'short programme' group (SP) trained every day for 2 weeks, while the 'long programme' group (LP) trained over a 6-week period with a 2-day rest period following each training session. The volunteers performed a 30-s supramaximal cycling test on a cycle ergometer before and after training. Muscle biopsies were obtained from the vastus lateralis before and after each test to examine metabolites and enzyme activities. Both training programmes led to a marked increase (all significant, P < 0.05) in enzymatic activities related to glycolysis (phosphofructokinase - SP 107%, LP 68% and aldolase - SP 46%, LP 28%) and aerobic metabolism (citrate synthase - SP 38%, LP 28.4% and 3-hydroxyacyl-CoA dehydrogenase - SP 60%, LP 38.7%). However, the activity of creatine kinase (44%), pyruvate kinase (35%) and lactate dehydrogenase (45%) rose significantly (P < 0.05) only in SP. At the end of the training programme, SP had suffered a significant decrease in anaerobic ATP consumption per gram muscle (P < 0.05) and glycogen degradation (P < 0.05) during the post-training test, and failed to improve performance. In contrast, LP showed a marked improvement in performance (P < 0.05) although without a significant increase in anaerobic ATP consumption, glycolysis or glycogenolysis rate. These results indicate that high-intensity cycling training in 14 sessions improves enzyme activities of anaerobic and aerobic metabolism. These changes are affected by the distribution of rest periods, hence shorter rest periods produce larger increase in pyruvate kinase, creatine kinase and lactate dehydrogenase. However, performance did not improve in a short training programme that did not include days for recovery, which suggests that muscle fibres suffer fatigue or injury. PMID- 10848647 TI - Response of jejunal Na+, K+-ATPase to 5-hydroxytryptamine in young and adult rats: effect of fasting and refeeding. AB - The present study is aimed to evaluate the effects of 5-hydroxytryptamine (5-HT) upon jejunal Na+,K+-ATPase in young (20-day-old) and adult (60-day-old) rats, and determine the effect of food intake on the response of the sodium pump to the amine. Basal Na+,K+-ATPase activity in jejunal epithelial cells from young rats was twice that in adult animals and responded to 5-HT with stimulation. In adult rats, fasting reduced by 25% basal jejunal Na+, K+-ATPase activity, whereas in young rats, no such change was observed. The sensitivity of jejunal Na+,K+-ATPase to 5-HT in young fasted rats was similar to that observed in fed animals. The effect of refeeding in young rats was a 2-fold increase in jejunal Na+, K+-ATPase activity, this being accompanied by insensitivity to 5-HT. In adult rats, refeeding was accompanied by an increase in jejunal Na+,K+-ATPase activity. It is concluded that the stimulatory effect of 5-HT upon jejunal Na+,K+-ATPase activity is a phenomenon dependent on both age and type of diet. In young rats, it is the food intake that plays an important role in development of insensitivity of Na+,K+-ATPase to stimulation by 5-HT, while in adult animals fasting or fasting followed by refeeding does not play a major role in regulating its sensitivity to the amine. PMID- 10848648 TI - Effect of alpha1-adrenergic stimulation of Cl- secretion and signal transduction in exocrine glands (Rana esculenta). AB - In the present work, the effect of stimulation of alpha-adrenergic receptors on Cl- secretion via exocrine frog skin glands was investigated. The alpha adrenergic stimulation was performed by addition of the adrenergic agonist noradrenaline in the presence of the beta-adrenergic antagonist propranolol. In the presence of propranolol, noradrenaline had no effect on the cellular cAMP content. The Cl- secretion was measured as the amiloride-insensitive short circuit current (ISC). Addition of noradrenaline induced a biphasic increase in the ISC. The increase in ISC coincided with an increase in the net 36Cl- secretion. The noradrenaline-induced increase in ISC was dose-dependent with an EC50 of 13 +/- 0.3 microM. Epifluorescence microscopic measurements of isolated, fura-2-loaded frog skin gland acini were used to characterize the intracellular calcium ([Ca2+]i) response. Application of noradrenaline induced a biphasic [Ca2+]i response, which was dose-dependent with an EC50 of 11 +/- 6 microM. The Ca2+ plateau unlike the peak-response was sensitive to removal of Ca2+ from the extracellular medium. The noradrenaline-induced increase in the Cl- secretion as well as in [Ca2+]i was sensitive to the alpha1-adrenergic antagonist prazosine. Ryanodine and caffeine had no effect on [Ca2+]i indicating that the release was independent of ryanodine-sensitive Ca2+ stores. Noradrenaline mediated a significant increase in the cellular inositol 1,4,5-trisphosphate (IP3) content suggesting that the signal transduction pathway leading to the noradrenaline induced increase in Ca2+ involved IP3 and a release of Ca2+ from IP3-sensitive stores. PMID- 10848649 TI - Review article: potential gastrointestinal effects of long-term acid suppression with proton pump inhibitors. AB - This review examines the evidence for the development of adverse effects due to prolonged gastric acid suppression with proton pump inhibitors. Potential areas of concern regarding long-term proton pump inhibitor use have included: carcinoid formation; development of gastric adenocarcinoma (especially in patients with Helicobacter pylori infection); bacterial overgrowth; enteric infections; and malabsorption of fat, minerals, and vitamins. Prolonged proton pump inhibitor use may lead to enterochromaffin-like cell hyperplasia, but has not been demonstrated to increase the risk of carcinoid formation. Long-term proton pump inhibitor treatment has not been documented to hasten the development or the progression of atrophic gastritis to intestinal metaplasia and gastric cancer, although long term studies are required to allow definitive conclusions. At present, we do not recommend that patients be tested routinely for H. pylori infection when using proton pump inhibitors for prolonged periods. Gastric bacterial overgrowth does increase with acid suppression, but important clinical sequelae, such a higher rate of gastric adenocarcinoma, have not been seen. The risk of enteric infection may increase with acid suppression, although this does not seem to be a common clinical problem with prolonged proton pump inhibitor use. The absorption of fats and minerals does not appear to be significantly impaired with chronic acid suppression. However, vitamin B12 concentration may be decreased when gastric acid is markedly suppressed for prolonged periods (e.g. Zolllinger-Ellison syndrome), and vitamin B12 levels should probably be assessed in patients taking high-dose proton pump inhibitors for many years. Thus, current evidence suggests that prolonged gastric acid suppression with proton pump inhibitors rarely, if ever, produces adverse events. Nevertheless, continued follow-up of patients taking proton pump inhibitors for extended periods will provide greater experience regarding the potential gastrointestinal adverse effects of long-term acid suppression. PMID- 10848650 TI - Review article: alginate-raft formulations in the treatment of heartburn and acid reflux. AB - Alginate-based raft-forming formulations have been marketed word-wide for over 30 years under various brand names, including Gaviscon. They are used for the symptomatic treatment of heartburn and oesophagitis, and appear to act by a unique mechanism which differs from that of traditional antacids. In the presence of gastric acid, alginates precipitate, forming a gel. Alginate-based raft forming formulations usually contain sodium or potassium bicarbonate; in the presence of gastric acid, the bicarbonate is converted to carbon dioxide which becomes entrapped within the gel precipitate, converting it into a foam which floats on the surface of the gastric contents, much like a raft on water. Both in vitro and in vivo studies have demonstrated that alginate-based rafts can entrap carbon dioxide, as well as antacid components contained in some formulations, thus providing a relatively pH-neutral barrier. Several studies have demonstrated that the alginate raft can preferentially move into the oesophagus in place, or ahead, of acidic gastric contents during episodes of gastro-oesophageal reflux; some studies further suggest that the raft can act as a physical barrier to reduce reflux episodes. Although some alginate-based formulations also contain antacid components which can provide significant acid neutralization capacity, the efficacy of these formulations to reduce heartburn symptoms does not appear to be totally dependent on the neutralization of bulk gastric contents. The strength of the alginate raft is dependant on several factors, including the amount of carbon dioxide generated and entrapped in the raft, the molecular properties of the alginate, and the presence of aluminium or calcium in the antacid components of the formulation. Raft formation occurs rapidly, often within a few seconds of dosing; hence alginate-containing antacids are comparable to traditional antacids for speed of onset of relief. Since the raft can be retained in the stomach for several hours, alginate-based raft-forming formulations can additionally provide longer-lasting relief than that of traditional antacids. Indeed, clinical studies have shown Gaviscon is superior to placebo, and equal to or significantly better than traditional antacids for relieving heartburn symptoms. Alginate-based, raft-forming formulations have been used to treat reflux symptoms in infants and children, and in the management of heartburn and reflux during pregnancy. While Gaviscon is effective when used alone, it is compatible with, and does not interfere with the activity of antisecretory agents such as cimetidine. Even with the introduction of new antisecretory and promotility agents, alginate-rafting formulations will continue to have a role in the treatment of heartburn and reflux symptoms. Their unique non-systemic mechanism of action provides rapid and long-duration relief of heartburn and acid reflux symptoms. PMID- 10848651 TI - A placebo-controlled study to assess the effects of 7-day dosing with 10, 20 and 40 mg rabeprazole on 24-h intragastric acidity and plasma gastrin in healthy male subjects. AB - AIM: To compare the effects of rabeprazole 10, 20 and 40 mg o.d. on 24-h intragastric acidity and plasma gastrin concentration in a randomized, double blind placebo-controlled trial. METHODS: Twenty-four healthy male volunteers were studied on the 7th day of morning dosing with either placebo or rabeprazole 10, 20 or 40 mg in a crossover fashion. On day 7, hourly intragastric acidity was measured for 24 h from 08.00 hours by gastric aspiration. Plasma gastrin concentrations were also measured hourly from 08.00 to 24.00 hours, and 2-hourly thereafter. RESULTS: Compared with placebo, rabeprazole 10, 20 and 40 mg produced significant dose-related decreases in intragastric acidity (median 24-h integrated acidity=697, 186, 129 and 82 mmol h/L, respectively). This was associated with significant elevation of plasma gastrin concentration (median 24 h integrated gastrin=141, 1184, 1484 and 1763 pmol.h/L, respectively). Rabeprazole 40 mg resulted in significantly decreased acidity compared with both 10 and 20 mg, and in longer times for which intragastric pH was maintained at > 3 (19. 2 h vs. 17.3 h and 17.5 h) and > 4 (17 h vs. 14.2 h and 15.2 h), but was accompanied by significantly increased plasma gastrin. There was a consistent trend for greater antisecretory activity for 20 mg compared with 10 mg, but these differences did not reach statistical significance. The interindividual variability in antisecretory response was greatest with 10 mg. CONCLUSIONS: Rabeprazole 10, 20 and 40 mg produce significant, profound dose-related inhibition of gastric acid secretion. Taking into account reciprocal increases in plasma gastrin and the interindividual variation in antisecretory response, 20 mg appears to be the preferred dose for routine clinical use. PMID- 10848652 TI - Rabeprazole produces rapid, potent, and long-acting inhibition of gastric acid secretion in subjects with Helicobacter pylori infection. AB - AIM: To compare acid inhibiting activity and duration of action of different doses of rabeprazole, a substituted benzimidazole characterized as a highly potent and irreversible H+, K+-ATPase inhibitor, administered for 7 days to subjects infected with Helicobacter pylori. METHODS: A total of 38 subjects (mean age 39.3 years) were enrolled in a single-centre, double-blind, randomized, crossover study. All subjects were confirmed positive for H. pylori by 14C urea breath test and ELISA serologies. Subjects were divided into two groups of 19 to receive two doses of rabeprazole, either 5 and 20 mg or 10 and 40 mg, and placebo, given in random order daily in the morning for 7 days. Peptone stimulated acid, pH, and gastrin measurements were made for 24 h after the 1st dose and for 48 h after the 7th dose. RESULTS: Peptone-stimulated acid secretion rates were decreased from 12.5 to 6.7, 4.0, 1.5, and 0.26 h after initial 5, 10, 20, and 40 mg doses, respectively; to 7.3, 4.3, 2.1, and 1.2 mmol/h 23 h after the initial dose; and to 2.4, 2.6, 0.6, and 0.8 mmol/h 23 h after the 7th dose. After 48 h, stimulated acid secretion had recovered less than 40% for all treatment groups compared to placebo. Median intragastric pH also increased from 2.0 with placebo to 4.9, 6.2, 6.6 and 6.9 during the 24-h period after the 7th dose of 5, 10, 20, and 40 mg. The 20 mg dose of rabeprazole produced equivalent acid inhibition to the 40 mg dose with less increase in plasma gastrin. CONCLUSION: Rabeprazole in doses from 5 to 40 mg was a highly effective inhibitor of gastric acid secretion in subjects infected with H. pylori. The inhibition was rapid, dose-related, and long-acting, with less than 50% recovery of acid by 48 h after the 7th dose. The optimal acid inhibitory dose in these subjects appeared to be 20 mg daily, however 5 mg and 10 mg doses produced potent inhibition of gastric acid secretion. PMID- 10848653 TI - Gastric acidity and acid breakthrough with twice-daily omeprazole or lansoprazole. AB - BACKGROUND: In patients with severe gastro-oesophageal reflux disease (GERD), proton pump inhibitors are being used increasingly in twice-daily regimens to improve control of gastric acidity. Few data exist to compare the ability of the most-often used proton pump inhibitors, omeprazole and lansoprazole, to control gastric acid at twice-daily dosage regimens. Nocturnal acid breakthrough, defined as gastric pH < 4.0 continuously for > 60 min, may compromise treatment goals in patients with GERD. AIM: To compare the effects of omeprazole 20 mg b.d. or lansoprazole 30 mg b.d. on gastric acidity and the relative ability of each dosage regimen to prevent acid breakthrough. METHODS: In a crossover pharmacodynamic study, 20 healthy volunteers (10 male, 10 female, mean age 38 years) were given omeprazole 20 mg b.d. or lansoprazole 30 mg b.d. for 7 days each, in a randomized manner. Each dosage regimen was separated by a minimum 7 day period where no medication was administered. On day 7 of each regimen, 24-h intragastric pH-metry was performed. The percentage of time for which gastric pH was below 4.0 and 3.0, the occurrence of daytime and nocturnal acid breakthrough, and the duration of action of each regimen were compared. Non-parametric statistics for paired data were used. RESULTS: The percentage time for which gastric pH was below 4.0 was significantly lower with omeprazole 20 mg b.d. (median 14.8%) than with lansoprazole 30 mg b. d. (median 24.2; P=0.0372). Fourteen subjects showed more effective acid control when taking omeprazole; these were significantly more often H. pylori-negative patients compared with those for whom acid control was better on lansoprazole (P < 0.001). Nocturnal acid breakthrough occurred in seven patients (35%) on omeprazole and in 10 (50%) on lansoprazole (N.S.). CONCLUSION: In healthy volunteers, twice-daily dosing of omeprazole 20 mg b.d. appears to be significantly more effective than lansoprazole 30 mg b.d. in controlling gastric acidity. The clinical importance of such a difference remains to be defined in GERD patients. PMID- 10848654 TI - A new highly effective short-term therapy schedule for Helicobacter pylori eradication. AB - BACKGROUND: Although triple therapy regimens suggested in the Current European guidelines give fairly good results, several studies have reported an unsatisfactory Helicobacter pylori eradication rate (< 80%). AIM: To evaluate the efficacy of a new short-term treatment sequence on H. pylori eradication. METHODS: A total of 52 patients with H. pylori infection and either non-ulcer dyspepsia (34 patients) or peptic ulcer (18 patients) were enrolled to receive a 10-day therapy: omeprazole 20 mg b.d. plus amoxycillin 1 g b.d. for the first 5 days, followed by omeprazole 20 mg b.d., clarithromycin 500 mg b.d. and tinidazole 500 mg b.d. for the remaining 5 days. H. pylori infection at entry was assessed by rapid urease test and histology on biopsies from the antrum and the corpus. Bacterial eradication was assessed by endoscopy (peptic ulcer patients) or 13C urea breath test (non-ulcer dyspepsia patients) 4-6 weeks after therapy had ended. RESULTS: All patients completed the study. H. pylori eradication was achieved in all but one patient, with an eradication rate of 98% (95% CI: 94.3 100) with intention-to-treat analysis. Patient compliance was good (consumption of prescribed drugs > 95%) for all but one patient, who took the triple therapy regimen for 4 days instead of 5 days. No major side-effects were reported but three (6%) patients complained of mild side-effects. CONCLUSIONS: The use of this 'five plus five' therapy schedule as an initial treatment for H. pylori deserves further investigation. PMID- 10848655 TI - The effectiveness of omeprazole, clarithromycin and tinidazole in eradicating Helicobacter pylori in a community screen and treat programme. Leeds Help Study Group. AB - INTRODUCTION: Helicobacter pylori screening and treatment has been proposed as a cost-effective method of preventing gastric cancer. AIM: To assess, in a randomized controlled trial, the efficacy of therapy in eradicating H. pylori as part of a screening programme, and to report the adverse events associated with this strategy. METHODS: Subjects between the ages of 40-49 years were randomly selected from the lists of 36 primary care centres. Participants attended their local practice and H. pylori status was determined by 13C-urea breath test. Infected subjects were randomized to receive omeprazole 20 mg b.d., clarithromycin 250 mg b.d. and tinidazole 500 mg b.d. for 7 days (OCT) or identical placebos. Eradication was determined by a 13C-urea breath test 6 months and 2 years after the first visit. Successful eradication was defined as two negative 13C-urea breath tests or one negative and one missing test. Adverse events and compliance were assessed at the 6-month visit. RESULTS: A total of 32 929 subjects were invited to attend, 8407 were evaluable, and 2329 (28%) of these were H. pylori-positive. A total of 1161 subjects were randomized to OCT and 1163 to placebo; over 80% returned for a repeat 13C-urea breath test on at least one occasion. The eradication rates in those allocated to OCT were as follows: intention-to-treat, 710 out of 1161 (61%; 95% confidence interval: 58-64%); evaluable 710 out of 967 (73%; 95% CI: 71-76%); took all medication 645 out of 769 (84%; 95% CI: 81-87%). Adverse events occurred in 45% of the treatment group and in 18% of the placebo group (relative risk 2.5; 95% CI: 2.1-2.9). Compliance, male gender, no antibiotic prescription in the subsequent 2 years and experiencing a bitter taste with the medication were independently associated with treatment success. CONCLUSIONS: The OCT regimen has an eradication rate of 61% in intention-to-treat analysis and is therefore less successful in treating H. pylori as part of a screening programme compared with hospital studies in dyspeptic patients. PMID- 10848656 TI - High incidence of reflux oesophagitis after eradication therapy for Helicobacter pylori: impacts of hiatal hernia and corpus gastritis. AB - BACKGROUND: Although several recent studies have shown that the eradication of Helicobacter pylori provokes reflux oesophagitis, the results are conflicting. AIM: To investigate the prevalence of reflux oesophagitis in patients after eradication of H. pylori and consider its association with hiatal hernia and corpus gastritis. METHODS: A total of 286 patients who underwent H. pylori eradication therapy and 286 age- and disease-matched H. pylori-positive controls who did not undergo eradication therapy were followed prospectively. All patients and controls underwent endoscopy at 1-year intervals or when upper gastrointestinal symptoms recurred. The presence of hiatal hernia and histology of the gastric corpus were evaluated at the time of initial endoscopy. RESULTS: The estimated prevalence of reflux oesophagitis within 3 years was 18% after eradication therapy and 0.3% without therapy. Patients who developed reflux oesophagitis after therapy had a greater prevalence of hiatal hernia (P=0.0008) and more severe corpus gastritis (P=0.0005) before therapy. Cumulative prevalence of reflux oesophagitis was 26% in patients with hiatal hernia, 7.7% in those without hiatal hernia, 33% in those with corpus atrophic gastritis and 13% in those without corpus atrophic gastritis. When patients had both hiatal hernia and corpus gastritis, the prevalence of reflux oesophagitis was 37%. The newly developed reflux oesophagitis was classified as mild in 35 out of 36 (97%) patients who developed reflux oesophagitis after eradication therapy. CONCLUSIONS: Eradication of H. pylori increased the prevalence of reflux oesophagitis in our patient group. The presence of hiatal hernia and corpus gastritis are closely related to the development of reflux oesophagitis after H. pylori eradication therapy. PMID- 10848657 TI - Randomized study comparing omeprazole with ranitidine as anti-secretory agents combined in quadruple second-line Helicobacter pylori eradication regimens. AB - BACKGROUND: Few data are available on the efficacy of second-line H. pylori eradication regimens. AIM: To compare the efficacy of either omeprazole or ranitidine in a second-line quadruple regimen in patients with duodenal ulcer or erosive duodenitis. PATIENTS AND METHODS: A total of 37 patients with erosive duodenitis and 119 with duodenal ulcer who have failed eradication of H. pylori with double or triple regimens, without metronidazole, were randomly assigned to receive tripotassium dicitrato bismuthate 600 mg t.d.s. + metronidazole 500 mg t.d.s. + tetracycline hydrochloride 500 mg t.d. s. combined with either omeprazole 20 mg b.d. (group O, 78 patients) or ranitidine 300 mg b.d. (group R, 78 patients) for 14 days. H. pylori eradication was verified by histology, rapid urease test and 13C-urea breath test. STATISTICS: t-test, chi2-test. RESULTS: A total of 143 patients had a post-treatment endoscopy. Eradication rates were: intention-to-treat: group O 77% (67-87), group R 76% (66-85), P=0.85; per protocol analysis: group O 86% (77-95), group R 82 (71-93), P=0.58. Side-effects were frequent but mild. CONCLUSIONS: Omeprazole 20 mg b.d. and ranitidine 300 mg b.d. were equally effective as antisecretory agents combined in a second-line quadruple eradication regimen. PMID- 10848658 TI - Metronidazole containing quadruple therapy for infection with metronidazole resistant Helicobacter pylori: a prospective study. AB - BACKGROUND: Metronidazole remains a key component of H. pylori infection therapy. It has been suggested that despite resistance, metronidazole may be effective when given at high dose with bismuth, tetracycline, and a proton pump inhibitor (quadruple therapy). AIM: To prospectively evaluate metronidazole quadruple therapy for treatment of metronidazole resistant H. pylori infection in the United States. METHODS: Patients infected with metronidazole resistant H. pylori were prospectively prescribed 14 days of quadruple therapy consisting of metronidazole 500 mg t.d.s., tetracycline 500 mg q.d.s., two bismuth subsalicylate tablets q.d.s., and omeprazole 20 mg o.d. RESULTS: A total of 26 patients were entered into the study; 22 for their first treatment and four as re treatment for failed therapy. Of the 26 patients, 24 were cured (cure rate 92%; 95% CI: 78-99%). Both treatment failures reported full compliance to 14 days of therapy. Side-effects were common and resulted in premature discontinuation of therapy in 31%. Premature discontinuation did not reduce the cure rate. CONCLUSION: Quadruple metronidazole combination therapy is effective despite the presence of metronidazole resistance and should be considered as either first line therapy or for failures of twice-a-day combination therapies. PMID- 10848659 TI - Randomized trial of omeprazole and metronidazole with amoxycillin or clarithromycin for Helicobacter pylori eradication, in a region of high primary metronidazole resistance: the HERO study. AB - BACKGROUND: The efficacy of omeprazole-based eradication therapies has been determined mostly in populations with low to moderate prevalence of metronidazole resistant Helicobacter pylori, yet resistance is high in many regions. AIM AND METHODS: The H. pylori eradication and duodenal ulcer healing rates after 1 week of either omeprazole 40 mg mane, amoxycillin 500 mg t.d.s. and metronidazole 400 mg t.d.s. (OAM) or omeprazole 20 mg b.d., metronidazole 400 mg b. d. and clarithromycin 250 mg b.d. (OMC) were compared in a randomized trial in Australia and New Zealand. Patients had a further 1 week of omeprazole 20 mg. Outcome was assessed at 6 weeks with stringent criteria (endoscopy, biopsies and 13C-urea breath test). RESULTS: Of 220 subjects randomized, the H. pylori eradication rates (all patients treated/per protocol) were 82%/85% for OMC and 58%/63% for OAM (P= 0.001). Pre-treatment metronidazole resistance was present in 56% and clarithromycin resistance in 6%. The eradication rate for primary metronidazole resistance isolates treated with OMC was 80% (CI: 65-90%) compared with 45% (CI: 29-62%) for OAM, whereas for sensitive organisms, the eradication rates were 94% (CI: 79-99%) and 79% (CI: 62-91%), respectively. Duodenal ulcer healing was 96% for OMC and 87% for OAM. Compliance was excellent and both treatments were well tolerated. CONCLUSIONS: OMC is a well-tolerated, effective therapy for H. pylori eradication and duodenal ulcer healing in this region despite the high metronidazole resistance rate. OAM is less effective, largely due to the impact of metronidazole resistance. PMID- 10848660 TI - Influence of age on the steady state disposition of drugs commonly used for the eradication of Helicobacter pylori. AB - BACKGROUND: The success of eradication therapy for Helicobacter pylori might be affected by the age of patients. AIM: To investigate whether disposition of drugs commonly used for H. pylori eradication is age-dependent. METHODS: Trough steady state serum levels of lansoprazole or ranitidine, amoxycillin, clarithromycin and metronidazole were monitored in 232 patients during the last dosing interval of a 5-day quadruple H. pylori eradication regimen. Detailed pharmacokinetic analysis was performed in 28 patients. RESULTS: Linear correlations between age and trough serum levels were observed with lansoprazole (r=0.25; P=0.002), ranitidine (r=0. 38; P=0.001) and clarithromycin (r=0.36; P < 0.0001). These associations were also inversely dependent of creatinine clearance for ranitidine (r=0.36; P=0.001) and clarithromycin (r=0.30; P < 0. 0001). Multiple linear regression revealed age as an important factor influencing trough serum levels of lansoprazole, clarithromycin and ranitidine. There were significant inverse relationships between creatinine clearance and area under curve of ranitidine (r=0.88; P < 0.0001) and amoxycillin (r=0.56; P=0.002). Multiple linear regression revealed serum creatinine as the most important factor influencing the area under curve of ranitidine, clarithromycin and amoxycillin. CONCLUSIONS: Age per se has little influence on pharmacokinetics of amoxycillin and ranitidine, which depend more on age-dependent decline in renal function. The influence of age, but not renal function was established for lansoprazole. Age and renal function have independent impacts on clarithromycin disposition. PMID- 10848661 TI - Cyclosporin in the treatment of adults with refractory coeliac disease--an open pilot study. AB - AIM: To evaluate the effect of cyclosporin treatment on clinical and histological parameters in adult patients with refractory coeliac disease. METHODS: Thirteen patients were treated with oral cyclosporin for 2 months, aiming at serum levels of 100-200 ng/mL. Seven extended medication intake up to a maximum of 12 months. Before and after treatment, clinical parameters were monitored and small intestinal biopsies taken. Ten of 13 patients were typed for HLA-DQA1 and -DQB1 alleles. RESULTS: Eight of 13 patients responded histologically to cyclosporin treatment. Normalization of villi was demonstrated in five patients, three after prolonged treatment. Eight patients reported a clinical response, of whom six had concomitant histological improvement. No serious side-effects of cyclosporin were noticed. Nine of 10 patients who were immunogenetically typed carried the coeliac disease associated serologic DQ2 markers, one carried neither DQ2 nor DQ8 markers. CONCLUSION: In our study group of 13 adult refractory coeliac disease patients, cyclosporin in therapeutic doses induced a histological improvement in eight patients (61%), in five of whom (38%) normalization of villi was demonstrated. Thus, we believe that cyclosporin is a therapeutic option in refractory coeliac disease, although we could not confirm earlier reports of unconditional successful treatment. PMID- 10848662 TI - Alosetron, a 5-HT3 receptor antagonist, delays colonic transit in patients with irritable bowel syndrome and healthy volunteers. AB - BACKGROUND: Alosetron is a potent and selective 5-HT3 receptor antagonist, which has been shown to be beneficial in the treatment of female patients with non constipated irritable bowel syndrome. AIMS: To investigate the effect of alosetron on whole gut, small bowel and colonic transit in patients with irritable bowel syndrome (Study 1) and healthy volunteers (Study 2). SUBJECTS: Thirteen patients with irritable bowel syndrome and 12 healthy volunteers. METHODS: Both studies were randomized, double-blind, placebo-controlled with a two-way crossover design, in which each subject received alosetron (2 mg b.d. administered orally) or placebo for 8 days. Mean whole gut transit was determined from the excretion of radio-opaque markers; small bowel transit was determined from rise in breath hydrogen after a meal; and colonic transit and segmental transit were evaluated from abdominal X-ray. In addition, colonic transit was calculated by subtracting small bowel transit time from whole gut transit time. RESULTS: Alosetron increased colonic transit time by prolonging left colonic transit in both patients with irritable bowel syndrome and controls. This resulted in a tendency for the whole gut transit to be delayed in irritable bowel syndrome patients (P=0.128), which was confirmed in controls (P=0.047). CONCLUSION: Alosetron delays colonic transit by prolonging left colonic transit. These results add to the body of evidence suggesting that alosetron should have a therapeutic role in patients with non-constipated irritable bowel syndrome. PMID- 10848663 TI - Towards identifying optimal doses for alpha-2 adrenergic modulation of colonic and rectal motor and sensory function. AB - RATIONALE: Visceral sensation and motility are important in functional gut disorders and are partly controlled by adrenergic innervation. OBJECTIVES: To characterize the alpha2-adrenergic control of motor and sensory function of descending colon and rectum. METHODS: In 32 healthy volunteers, we assessed compliance, fasting and postprandial tone, and sensations of gas, urgency and pain during phasic distentions. Each subject received one agent at clinically approved doses: clonidine (0.05, 0.1, 0.2 or 0.3 mg p.o. ); or the alpha2 antagonist yohimbine (0.0125 mg, 0.05 mg, 0.125 mg or 0.2 mg intravenously and infusion over 2.5 h). RESULTS: Clonidine increased colonic and rectal compliance, and reduced tone, pain, gas sensation and rectal urgency. Clonidine showed large pairwise differences in sensation and motility between 0.05 and 0.1 mg doses, which did not interfere with the colon's motor response to feeding. Conversely, yohimbine dose-dependently altered the compliance curve, increased tone and sensations of gas, pain and urgency. Drug effects in the colon were more marked at low distensions; alpha2 modulation of rectal sensation was observed at all levels of distension. CONCLUSIONS: alpha2-adrenergic mechanisms modulate colorectal sensations and motility; at doses as low as 0.05 mg, clonidine reduced colorectal sensation while the tone response to feeding was preserved. These studies provide insight into the potential use of alpha2 agents in disease states. PMID- 10848664 TI - The effects of meloxicam, indomethacin or NS-398 on eicosanoid synthesis by fresh human gastric mucosa. AB - BACKGROUND: In the stomach, constitutive cyclooxygenase (COX-1) synthesizes prostaglandins that maintain the integrity of the gastric mucosa, while their inhibition contributes to gastric mucosal damage. In contrast COX-2, an inducible enzyme, forms prostanoids involved in pain and inflammation. AIM: To compare prostaglandin synthesis inhibition by meloxicam, a selective COX-2 NSAID reported to have better gastric tolerability, with indomethacin and NS-398 in human gastric mucosa and in whole blood assays. METHODS: Meloxicam, indomethacin or NS 398 were incubated with fresh human gastric mucosa pieces (100 mg in 1 mL phosphate buffered saline, pH 7.4, 37 degrees C, 30 min), clotting human blood (1 mL, 37 degrees C, 60 min) or with lipopolysaccharide-stimulated heparinized blood (1 mL, 37 degrees C, 24 h). Prostanoids were analysed by radioimmunoassay. RESULTS: Meloxicam was a less potent inhibitor of gastric mucosal eicosanoid compared to indomethacin, showing a sixfold difference in IC50 with gastric mucosal prostaglandin E (PGE) (11.8 and 1.8 microM, respectively). In the whole blood assays, the COX-2/COX-1 ratio for meloxicam was 0.2 compared to 0.9 for indomethacin confirming meloxicam's COX-2 selectivity. CONCLUSION: The results with human mucosa pieces would suggest that the better gastric tolerability of meloxicam compared to indomethacin is related to its relatively lower inhibition of gastric mucosal PGE synthesis by COX-1. PMID- 10848665 TI - Open label trial of oral clarithromycin in active Crohn's disease. AB - BACKGROUND: Crohn's disease seems likely to be due in some way to bacteria. Clarithromycin is a broad spectrum macrolide antibiotic with good penetration into macrophages and may be effective in eradicating the organisms that are presumed to be at the centre of the granulomatous reaction in Crohn's disease. METHODS: Twenty-five patients with active Crohn's disease were treated with oral clarithromycin 250 mg b.d. in an open label study. Treatment was for an initial 4 week period, continued to 12 weeks in patients who had shown a partial or complete response. The patients had a median age of 30 years (range 17-72), and disease duration of 5 years (range 2 months-28 years); 14 had ileocolonic, four small bowel, seven colonic disease and 10 had previous resections. Twenty patients were receiving a 5-ASA preparation, 15 corticosteroids (prednisolone median dose 10 mg range 2-30 mg) and nine azathioprine. All patients receiving corticosteroids or azathioprine had been on unchanged treatment for at least 12 weeks. RESULTS: Median pre-treatment Harvey Bradshaw index (HBI) was 9 (range 5 16) and median serum C-reactive protein was 21.5 mg/L (range < 5-117). By 4 weeks the median HBI had decreased to 5 (range 0-18) (P < 0.001) and median CRP to 17 mg/L (range < 5-157) (P=0.16). Sixteen patients (64%) had at least a 3 point fall in HBI and remission (defined as a HBI less than or equal to 4) was achieved in 12 patients (48%). By 12 weeks median HBI was 5 (range 0-18) (P < 0.001) and median CRP was 14.5 mg/L (range < 5-157) (P=0.05). Eleven of the 25 patients studied continued on oral clarithromycin after 12 weeks for a median of 28 weeks (range 20-60). Eight (73%) remained in remission on treatment. When treatment with clarithromycin was stopped three remained in remission and five relapsed after a median of 5 months (range 4-9). Two patients withdrew due to non-serious side-effects. Treatment was well tolerated in the remaining patients. CONCLUSION: This open label study has shown an impressive response to clarithromycin in a group of patients with active Crohn's disease, many of whom had been resistant to other therapy. A formal randomized controlled trial of clarithromycin in active Crohn's disease is needed. PMID- 10848666 TI - Ridogrel, a dual thromboxane synthase inhibitor and receptor antagonist: anti inflammatory profile in inflammatory bowel disease. AB - BACKGROUND: Thromboxanes, prostaglandins, reactive oxygen metabolites and pro inflammatory cytokines are produced in excess in inflammatory bowel disease. Preliminary reports suggest that ridogrel, a thromboxane synthesis inhibitor and receptor blocker, may have therapeutic benefits in ulcerative colitis. AIMS: To investigate the anti-inflammatory profile of ridogrel. METHODS: The effects of ridogrel on the production of eicosanoids, reactive oxygen metabolites and cytokines by cultured inflamed colorectal mucosal biopsies were made using ELISA and chemiluminescence, reactive oxygen metabolite generation in a cell-free system, and platelet activation using flow cytometry. The effects of oral ridogrel on mucosal release of eicosanoids in two patients with active ulcerative colitis were assessed using rectal dialysis. RESULTS: Ridogrel significantly reduced the release of thromboxane B2, but not prostaglandin E2 or tumour necrosis factor-alpha, from biopsies (P < 0.01 for 10 microM ridogrel). Ridogrel showed no direct antioxidant activity but significantly reduced reactive oxygen metabolite production from cultured biopsies (P < 0.01 for 10 microM ridogrel). Platelet activation in vitro was inhibited by ridogrel (P /= 10 microM ridogrel). Mean rectal mucosal thromboxane B2 release was reduced to 86% of pre-treatment levels in two patients treated with oral ridogrel. CONCLUSIONS: Its inhibition of mucosal production of thromboxane B2, reactive oxygen metabolites, and of platelet activation, suggests that ridogrel could have a therapeutic role in inflammatory bowel disease. PMID- 10848667 TI - The effects of smoking and indomethacin on small intestinal permeability. AB - BACKGROUND: Smoking modulates inflammatory bowel disease, protecting from ulcerative colitis on the one hand and worsening the course of Crohn's disease on the other. This influence might occur through changes in intestinal permeability, because permeability is increased in most patients with Crohn's disease. AIM: To study the influence of smoking on small intestinal permeability and its increase induced by indomethacin. METHODS: 50 smokers and 50 nonsmokers underwent a 51Cr EDTA basal permeability test and the same test after challenge with indomethacin 125 mg p.o. RESULTS: Small intestinal permeability was the same in smokers (median 1.22%; IQR 1.00-1.58) and nonsmokers (1.24%; 0.94-1.66). Basal small intestinal permeability was lower in females (1.09%; 0.87-1.33) than in males (1.48%; 1.18-1.88). Indomethacin challenge increased permeability by 110% (71 141) in smokers, vs. 156% (78-220) in the nonsmokers (P=0.04). CONCLUSION: Smoking reduces the effect of NSAID on small intestinal permeability. It is therefore unlikely that the adverse effect of smoking on Crohn's disease is related to its influence on intestinal permeability. PMID- 10848668 TI - Pharmacokinetics of Lactobacillus plantarum NCIMB 8826, Lactobacillus fermentum KLD, and Lactococcus lactis MG 1363 in the human gastrointestinal tract. AB - OBJECTIVES: Genetically modified lactic acid bacteria may be a way to deliver vaccinal epitopes in the gastrointestinal tract. AIM: Three strains of lactic acid bacteria were studied for their pharmacokinetics in the human gastrointestinal tract. METHODS: The survival of the strains was studied up to the ileum in six subjects each, after ingestion of 150 g of fermented milk. The strains and their concentrations in the products were Lactobacillus fermentum KLD (107 cfu/g), Lactobacillus plantarum NCIMB 8826 (108 cfu/g), and Lactococcus lactis MG 1363 (108 cfu/g). Ileal fluid was aspirated by intestinal intubation and immediately cultured. L. plantarum NCIMB 8826, which was found in high concentrations in the ileum, was studied for its survival in the faeces after consumption of 150 g of fermented milk three times daily for 7 days. Faecal samples were collected for culture. RESULTS: The concentration of L. plantarum NCIMB 8826 in the ileum reached 108 cfu/mL after a single dose, with a survival of 7%. L. fermentum KLD and Lc. lactis MG 1363 had lower (0.5 and 1.0%, respectively) and shorter (4 h) survival in the ileum. During the 7-day ingestion period, L. plantarum NCIMB 8826 reached high concentrations (108 cfu/g) in the faeces, with a survival of 25 +/- 29%. None of the strains colonized. CONCLUSIONS: L. plantarum NCIMB 8826 has a promising pharmacokinetic profile as a candidate vaccine vehicle. PMID- 10848669 TI - Botulinum toxin injected in the gastric wall reduces body weight and food intake in rats. AB - BACKGROUND: Botulinum toxin is a powerful, long-acting inhibitor of muscular contractions in both voluntary and smooth muscle. It acts by blocking the release of the neurotransmitter acetylcholine. In the stomach, propulsive contractions of the antrum are necessary for the gastric contents to pass into the duodenum. AIMS: To investigate whether intramuscular injections of botulinum toxin type A into the gastric antrum of rats would cause a reduction in food intake and hence body weight, by inhibition of gastric emptying. MATERIALS AND METHODS: This was a prospective, randomized, 3-way parallel group study in rats. The first group was anaesthetized, laparotomized and given 20 U of botulinum toxin type A by intramuscular injection into the gastric antrum (botulinum toxin type A group, n=14). The second group was anaesthetized, laparotomized and injected with saline (sham group, n=14) and the third group did not have any intervention (control group, n=5). Food intake was measured daily for 7 weeks and body weight was measured daily for 10 weeks. RESULTS: There was a significant difference in loss of body weight between the two treated groups (14.0 +/- 8.2% botulinum toxin type A group, 4.4 +/- 2.7% sham group; P < 0.001). Further, the time to reach the weight nadir was significantly longer in the botulinum toxin type A group (8.7 +/ 3.9 days) compared with the sham group (5.3 +/- 3.8 days; P < 0.04). There were no significant differences between the sham and control groups for any of the body weight parameters. The minimum dietary intake was significantly lower in the botulinum toxin type A group than in the sham group (37.8 +/- 21.8% of the basal value in the botulinum toxin type A group, vs. 65.5 +/- 32.0 in the sham group, P < 0.05). In addition, the time to reach the nadir was significantly prolonged (8.2 +/- 3.5 days, botulinum toxin type A group vs. 4.9 +/- 1.7 days, sham group, P < 0.001). CONCLUSIONS: The parallel reduction of body weight and food intake in botulinum toxin type A treated animals is consistent with a long lasting inhibition of the antral pump. This is probably due to slowed gastric emptying leading to early satiety. Patients with morbid obesity might benefit from endoscopic injections of botulinum toxin type A into the stomach wall. PMID- 10848670 TI - Systemic effect of peanut agglutinin following intravenous infusion into rats. AB - BACKGROUND: Ingested peanut agglutinin stimulates colonic proliferation in humans. In rats, ingested peanut agglutinin stimulates hormone release and proliferation in the small and large intestines. Peanut agglutinin is absorbed into the circulation but little is known about the systemic effect of this lectin. Therefore, we studied the effect of intravenous peanut agglutinin on hormone release and intestinal growth. METHOD: Six rats per group received peanut agglutinin infusion at 0, 2, 20 or 200 microg/rat/day for 6 days via the right jugular vein. Organ weights were measured, pancreatic enzymes, DNA, RNA and protein levels were analysed. Plasma hormones were measured by radioimmunoassay. All tissues were examined histologically. Small intestinal and colonic proliferation rates were estimated by metaphase arrest. RESULTS: High-dose peanut agglutinin significantly reduced the wet weight of the stomach by 7% (P < 0.05) and large intestine by 10% (P < 0.05). Peanut agglutinin dose-dependently released enteroglucagon; low-, medium- and high-dose by 64%, 126% (P < 0.01) and 180% (P < 0.01), respectively, and glucagon-like peptide-1 by 127% (P < 0.01), 169% (P < 0.01) and 315% (P < 0.001), respectively. Peanut agglutinin had no effect on cholesystokinin, gastrin or insulin levels. Peanut agglutinin, low-, medium- and high-dose stimulated proliferation in the mid colon by 42% (P < 0.01), 30% and 38%, respectively. Only high-dose peanut agglutinin stimulated proliferation in the distal colon by 54% (P < 0.01). No histological changes were evident in any tissue. CONCLUSION: Intravenous peanut agglutinin released hormones and stimulated colonic proliferation. Proliferation of the small intestine seen after ingestion of peanut agglutinin in previous studies appears to require luminal contact between enterocytes and the lectin. Possible clinical applications include reversal of atrophy during total parenteral nutrition, anastomotic healing after surgery and restoration of mucosa integrity in colitis. PMID- 10848671 TI - Tepoxalin inhibits inflammation and microvascular dysfunction induced by abdominal irradiation in rats. AB - BACKGROUND: Inflammatory cells contribute to the acute and sub-acute sequelae of radiation therapy. Tepoxalin, an inhibitor of cyclooxygenase and 5-lipoxygenase that suppresses NF-kappaB activation, has potent anti-inflammatory activity. AIMS: To assess the effects of tepoxalin on radiation-induced inflammatory damage, and determine its mechanisms of action. METHODS: Leucocyte rolling, adhesion and emigration, and albumin leakage were determined by intra-vital microscopy in rat mesenteric venules. NF-kappaB activation was measured by electrophoretic mobility shift assays, and endothelial intercellular adhesion molecule-1 expression by the radiolabelled antibody technique. Groups of irradiated rats were treated with tepoxalin, N-acetyl-L-cysteine, zileuton (lipoxygenase inhibitor), or vehicle. RESULTS: Irradiated animals had a marked increase in the number of rolling, adherent and emigrated leucocytes in mesenteric venules, and in microvascular permeability. Tepoxalin prevented leucocyte adhesion and the increase in permeability after radiation. Tepoxalin did not inhibit radiation-induced NF-kappaB activation or intercellular adhesion molecule-1 up-regulation, while N-acetyl-L-cysteine, which attenuated NF-kappaB activation, had no effect on leucocyte recruitment. In contrast, tepoxalin inhibited the increase in leukotriene B4 levels after radiation, and the anti inflammatory effects of the drug were mimicked by zileuton. CONCLUSIONS: Tepoxalin affords significant protection against radiation-induced inflammation and microvascular dysfunction in splanchnic organs through a mechanism dependent on leukotriene synthesis inhibition. PMID- 10848672 TI - Novel online infusate-assisted dialysis system performs microbiologically safely. AB - Administration of adequate amounts of commercial infusion fluids renders modern convective dialysis modalities, such as hemodiafiltration, labor-intensive and costly. Preparation of infusate by cold sterilization of dialysis fluid, which is abundantly available, and its immediate (online) use, in contrast, enables a large volume fluid exchange in a cost-effective manner. Recent developments aimed at more hygienic and user-friendly online systems with increased operational flexibility. As a result the novel ONLINEplus system does not only provide online prepared infusate for convective dialysis therapy, but also for priming and rinsing of the extracorporeal blood circuit, for intradialytic bolus administration, and for re-infusion of patients' blood as well. Production of infusate from potentially impure dialysis fluid containing endotoxins and other pyrogens raises severe concerns of affecting the patients' well-being. To assess its safety, the online system was challenged with microbially contaminated dialysis fluid. Despite high levels of microbial counts (7.5 x 104 +/- 105 CFU/ml), endotoxin concentration (14.1 +/- 7.7 IU/ml and 9.265 +/- 3.000 IU/ml, as measured turbidimetrically and chromogenically, respectively) and cytokine inducing activity (20,827 +/- 3,082 pg IL-1Ra/Mio WBC), we failed to detect contaminants in the final infusate during a 5 week laboratory testing period. In addition, infusate samples complied consistently with the European Pharmacopeia test for sterility. The present online system is comprehensive, operates user friendly, and provides microbiologically safe infusate in large quantities. In this way, both patients and dialysis staff will benefit from improved dialysis therapy and reduced treatment-related labor burden, respectively. Moreover, convective dialysis modalities will become less expensive. PMID- 10848673 TI - Cytokine release and serum lipoprotein (a) alterations during hemodialysis. AB - It has been reported recently that a number of cytokines, mainly tumor necrosis factor alpha (TNFalpha), interleukin (IL)-1beta, and IL-6, can alter lipid metabolism and produce hyperlipidemia. Studies in hemodialysis (HD) patients have demonstrated increased production of these cytokines during HD. In order to investigate any possible relationship between changes of cytokines and lipid concentrations during HD in the serum of 25 uremic patients on chronic HD using modified cellulose membranes, TNFalpha, IL-1beta, IL-6, total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), lipoprotein a (Lp[a]), and total proteins were measured immediately before (pre-HD) and after HD (post-HD), in one session. The post-HD values were corrected according to the hemoconcentration based on the changes in serum total proteins. Serum TNFalpha and IL-1beta levels were significantly increased from 38.24 +/- 17.85 pg/ml and 2. 60 +/- 3.64 pg/ml pre HD to 48.86 +/- 25.21 and 3.49 +/- 4.08 pg/ml post-HD, p < 0.001 and p < 0.05 respectively. Also Lp(a) levels presented a statistically significant increase post-HD and were almost doubled (pre-HD: 15.41 mg/dl, to post-HD: 27.39 mg/dl, p < 0. 05). Serum IL-6 as well as serum TC, TG, HDL-C, and LDL-C did not show any statistically significant alterations during HD. A significant positive correlation was detected between TNFalpha and Lp(a) values post-HD (r: 0.413, p: 0.04), but not between pre-HD values. No further relationship between serum cytokines and the other estimated lipid parameters was observed, either between pre- or post-HD values. Our results indicate that release of TNFalpha and IL 1beta during HD have no effect on serum lipids concentration, except on Lp(a). It seems that the acute rise of this lipoprotein during hemodialysis may be related with the TNFalpha overproduction. PMID- 10848674 TI - Modeling lipid uptake in expanded polytetrafluoroethylene vascular prostheses and its effects on mechanical properties. AB - The radial transport across the wall of expanded polytetrafluoroethylene (ePTFE) arterial prostheses has a significant effect on lipid uptake observed in prostheses implanted in humans, which has been postulated to be one of the causes associated with implant failure. The goal of this study was to stimulate radial transport on a lipidic dispersion across the wall of an ePTFE prosthesis and investigate its effects on the circumferential mechanical properties of the prosthesis. An in vitro model was developed to simulate the lipidic radial transport across the wall. Lipids contained in a phosphatidylcholine dispersion were used as the transported molecules. Lipid concentration profiles were obtained after exposing commercial ePTFE prostheses to various transmural pressure and/or lipidic concentration gradients. Phospholipids gradually accumulated up to the external reinforcing wrap of the prosthesis, which clearly acted as a rigid barrier against lipid infiltration. Tensile tests performed on the virgin samples showed that the wrap was much more rigid than the microporous part of the prosthesis. After the lipid simulation, the rigidity of the wrap decreased with respect to what was observed for the virgin prosthesis. Finally, some clinical implications of this phenomena are discussed. PMID- 10848675 TI - In vitro hydrodynamic characteristics among three bileaflet valves in the mitral position. AB - The non-fully open phenomenon of the advancing standard medical bileaflet heart valves (the ATS valve) are frequently observed in clinical cases, even though there is no problem with their hemodynamic function. The movement of the leaflets was affected easily by the transvalvular flow because of the unique open pivot design of the ATS valve. In this paper, a comparative in vitro hydrodynamic test was conducted among 3 different types of bileaflet valves, and the effect of different shapes of downstream conduits, which induce different transvalvular flow, on hydrodynamic performance was studied. Three bileaflet valves, the ATS valve, CarboMedics valve (CM), and St. Jude Medical valve (SJM), with an annulus diameter of 29 mm for the mitral position were chosen throughout our experiments. First, pressure drops across the valves under steady flow were measured. Then, the valves were tested at the mitral position with our pneumatically driven pulsatile pump. In this pulsatile flow study, 2 different conduits (straight shape and abrupt enlargement shape) were in turn incorporated at the downstream portion of the mitral valve. A high-speed video camera was employed to observe leaflet movements. In a steady-flow test, the ATS and the SJM produced the same pressure drop, but the CM recorded a higher value. In the pulsatile study, it was observed that the ATS leaflets did not open fully in the mitral position when the downstream conduit with an abrupt enlargement shape was incorporated. However, the CM and the SJM always indicated a fully open movement regardless of the shape of downstream conduits. When the straight downstream conduit was incorporated, the ATS produced a similar pressure drop to that of the SJM, which coincided with the steady test results. When the enlargement conduit was incorporated, however, the ATS presented the lowest pressure drop despite the non-fully open movement. The conduit shape at the valve downstream had a significant influence on the closing volume. These findings indicate that the conduit shape at the valve outlet can affect the hydrodynamic characteristics of bileaflet valves. PMID- 10848676 TI - Glycoprotein IIb/IIIa receptor inhibitor attenuates platelet aggregation induced by thromboxane A2 during in vitro nonpulsatile ventricular assist circulation. AB - A recent development in antithrombotic research allows the inhibition of platelet aggregation via protection of the glycoprotein IIb/IIIa receptor on the platelet membrane. We hypothesized that a GP IIb/IIIa receptor inhibitor would inhibit thromboxane-induced platelet aggregation during circulation in our in vitro ventricular assist device (VAD) circuit and preserve long-term platelet function. Twenty-one in vitro nonpulsatile centrifugal VAD circuits were simulated for 4 days using 450 ml of fresh human whole blood with or without glycoprotein IIb/IIIa receptor inhibitor (tirofiban). Platelet aggregation and degranulation were measured in whole blood induced by ristocetin, collagen, ADP, and thromboxane A2 (TXA2). The tirofiban-treated group preserved the platelet count and tended to exert these beneficial effects by inhibiting pathologic platelet aggregation induced by TXA2, collagen, and ADP as well as degranulation. Tirofiban may be useful in preserving platelet number and function during clinical VAD use. PMID- 10848677 TI - Fluid dynamics of a pediatric ventricular assist device. AB - The number of pediatric patients requiring some form of mechanical circulatory assistance is growing throughout the world because of new surgical procedures and the success of pediatric cardiac transplantation. However, the salvage rate for those patients requiring circulatory support may be as low as 25%. Despite the fact that Penn State's 70 cc pneumatic ventricular assist device has been used with a success rate of over 90% in more than 250 patients worldwide, efforts to scale down the pump have encountered difficulties. Animal experiments with a 15 cc version were unsuccessful, with explanted pumps showing extensive thrombus deposition within the pumping chamber. The materials used to fabricate the smaller pump as well as the basic operating principles are identical to the successful adult-sized version. It is therefore believed that reducing the size of the pump altered the internal flow field, and that fluid dynamic factors were responsible for the high degree of thrombus observed with the implanted devices. A dimensional analysis was conducted that revealed significant differences in both Reynolds (Re) and Strouhal (St) numbers between the successful and unsuccessful pumps. Two component laser Doppler velocimetry was then used to characterize the internal flow field quantitatively. Comparison with data from the 70 cc pump showed a reduction in wall shear stress and turbulence levels in the 15 cc pump that would yield an environment conducive to clot formation. PMID- 10848678 TI - A small pulsatile blood pump for ventricular support during end-stage heart failure. AB - A displacement blood pump to support the natural heart of patients for recovery from end-stage heart failure has been developed. This electromechanical pusher plate pump has a very compact and extremely flat design. The design goal was achieved by developing a novel gear system based on the principle of a swash plate. The blood pump and cannulae can be placed within the thoracic cavity between the lungs and ribcage. The first labtype model delivers an output of 3.1 L/min against an aortic pressure of 100 mm Hg at 120 bpm. PMID- 10848679 TI - Computational flow study of the continuous flow ventricular assist device, prototype number 3 blood pump. AB - A computational fluid dynamics study of blood flow in the continuous flow ventricular assist device, Prototype No. 3 (CFVAD3), which consists of a 4 blade shrouded impeller fully supported in magnetic bearings, was performed. This study focused on the regions within the pump where return flow occurs to the pump inlet, and where potentially damaging shear stresses and flow stagnation might occur: the impeller blade passages and the narrow gap clearance regions between the impeller-rotor and pump housing. Two separate geometry models define the spacing between the pump housing and the impeller's hub and shroud, and a third geometry model defines the pump's impeller and curved blades. The flow fields in these regions were calculated for various operating conditions of the pump. Pump performance curves were calculated, which compare well with experimentally obtained data. For all pump operating conditions, the flow rates within the gap regions were predicted to be toward the inlet of the pump, thus recirculating a portion of the impeller flow. Two smaller gap clearance regions were numerically examined to reduce the recirculation and to improve pump efficiency. The computational and geometry models will be used in future studies of a smaller pump to determine increased pump efficiency and the risk of hemolysis due to shear stress, and to insure the washing of blood through the clearance regions to prevent thrombosis. PMID- 10848680 TI - Is vecuronium toxicity abolished by hemodialysis? A case report. AB - Vecuronium is a curaric agent, largely used in anesthesia. Indications as to its employ in uremic patients appear to be debated because of partial renal elimination of the drug. A 52-year-old hemodialyzed woman required transplantectomy for rejection. At awakeness after general anesthesia (induced with fentanyl, propofol, and 6 mg of vecuronium, repeated with a single 2 mg dose 30 min later), she presented diafragmatic and muscular limb weakeness that lasted 180 min in spite of prostigmine administration. A 2 h 30 min predilutional hemofiltration was then performed, which induced rapid disappearance of neuromuscular blockade. Even if vecuronium can be used in dialysis patients, one should remember its possible side effects, especially with repeated doses, in determining prolonged neuromuscular blockade. Cautious use of this drug in renal failure is mandatory. Low dosage must be employed and repeated administration avoided. Neuromuscular blockade seems to be rapidly reversible with dialytic treatment. PMID- 10848681 TI - Effect of hemodialysis on plasma nitric oxide levels. AB - Nitric oxide (NO) is produced in excess in various pathological states, including sepsis and hepatic cirrhosis, and appears to be related to inflammatory status. In uremia, one would expect the levels of NO to increase. We aimed to determine whether hemodialysis (HD) would remove NO from the systemic circulation of uremic patients. Blood was collected before, after, and 1 day after HD from 12 uremic patients. Plasma nitrite and nitrate (NOx-) levels were measured by colorimetric Greiss reaction and cGMP was measured by an enzyme immunoassay kit. Our study demonstrated that uremic patients have high plasma NO levels, and HD led to a significant drop in plasma NOx- level (63 +/- 15% reduction). The level rose back to the pre-HD level on the following day. Plasma cGMP in the patients also decreased significantly after HD (27 +/- 14% reduction). In conclusion, we hypothesized that HD might be a possible approach for the removal of excess NO in pathological conditions such as sepsis and hepatic cirrhosis. PMID- 10848682 TI - Combined effects of inhaled nitric oxide and positive end-expiratory pressure during mechanical ventilation in acute respiratory distress syndrome. AB - We studied the combined effects of inhaled nitric oxide (INO) and positive end expiratory pressure (PEEP) during mechanical ventilation in patients with acute respiratory distress syndrome (ARDS). Eleven patients received 0 and 4 parts per million of INO in random order for 30 min at PEEP levels of 0, 5, and 10 cm H2O. Respiratory and cardiovascular parameters were measured. The addition of INO and PEEP significantly improved arterial oxygenation (p < 0.005 and p < 0.0001, respectively). The combined effect of INO and PEEP on arterial oxygenation was remarkable during 10 cm H2O PEEP. There was synergistic effect on arterial oxygenation by combining INO and 10 cm H2O PEEP. The present study showed that the combination of INO and 10 cm H2O PEEP enhanced arterial oxygenation in patients with ARDS. PMID- 10848683 TI - Design and test of a vascular access device. AB - Transarterial left ventricular assist devices (LVADs), such as the Hemopump, IABP, and PUCA-pump, are meant to be introduced into the body via the femoral or axillary artery without major surgery. For certain applications, introduction is performed directly into the aorta via an open thorax procedure. A prototype of a vascular access device has been realized that allows direct access into the aorta as an alternative for the common surgical graft anastomosis suturing technique. The device consists of a metal tube acting as a circular knife to cut a hole in the aortic wall, a screw to store the removed part of the aortic wall, and a plastic tube that is introduced through the hole and tightly connected to the aortic wall. The device could be placed without aortic clamping. The device has been tested on a slaughterhouse porcine aorta. A low-pressurized aorta appeared to be the worst case; thus, two animal experiments in the low-pressurized pulmonary artery were performed. No leakage occurred for pressures between 40 and 300 mm Hg. PMID- 10848684 TI - Recombinant human erythropoietin stimulates production of interleukin 2 by whole blood cell cultures of hemodialysis patients. PMID- 10848685 TI - Attenuation of reperfusion injury by renal ischaemic preconditioning: the role of nitric oxide. AB - OBJECTIVE: To determine the effect on nitric oxide (NO) release and renal NO synthase (endothelial, eNOS and inducible, iNOS) activity of renal ischaemia reperfusion (I/R) in vivo in an animal model, and to examine the possible involvement of NO in ischaemic preconditioning (IP) of the kidney. MATERIALS AND METHODS: In a right-nephrectomized rat model, 42 animals were randomized in four groups: controls; IP-only (4 min of ischaemia followed by 11 min of reperfusion, total of four cycles); renal warm ischaemia (45 min) and 6 h reperfusion; ischaemia (45 min) preceded by IP pretreatment. Serum NO metabolites were assayed 2 and 6 h after ischaemia or the control equivalent. NOS expression in the kidney was detected immuno-histochemically, and damage assessed morphologically in sections stained with haematoxylin and eosin. Kidney function was assessed by the levels of serum creatinine, urea and electrolytes. RESULTS: Compared with before ischaemia, the concentration of serum NO metabolites at 6 h was increased in the IP-only animals (P = 0. 016) and in the IP + I/R group (P = 0.002). There was greater eNOS expression in the IP-only group (P = 0.009) and in the IP + I/R group than in controls (P = 0.050). iNOS expression was greater in the IP-only animals than in the control group (P = 0.050). Histological assessment showed less evidence of cellular damage in IP + I/R animals than in the I/R-alone group (P = 0.020). Serum creatinine level was not significantly different between the IP-only group and the control. There were no differences after 2 h of reperfusion. CONCLUSION: Ischaemic preconditioning has a protective effect on renal structure and function, which may be produced by increased NO release arising from increased NOS expression by 6 h after reperfusion. PMID- 10848686 TI - Monitoring of renal function in patients with spinal cord injury. AB - OBJECTIVE: To assess the sensitivity of serum creatinine level in detecting clinically important and early deterioration of renal function in patients with spinal cord injury (SCI), and to evaluate the optimal method of determining creatinine clearance in these patients. PATIENTS AND METHODS: The serum creatinine level of 36 patients (25 paraplegics and 11 quadriplegics) was evaluated and compared with the corresponding measured creatinine clearance rate. Correlations were also assessed between the creatinine clearance measured by 24-h endogenous clearance, single-shot 99mTc-labelled diethylenetriamine pentaacetic acid (99mTc-DTPA) clearance technique, and the Cockcroft-Gault formula, to test their validity. RESULTS: Of the 36 patients 11 (31%) had a measured creatinine clearance of < 100 mL/min (mean 84.8) and a corresponding normal serum creatinine level. Creatinine clearance calculated by the Cockcroft-Gault formula did not correlate well with that measured by the 24-h endogenous clearance (r = 0.426) and 99mTc-DTPA clearance (r = 0. 366), overestimating creatinine clearance in all but three patients. The mean (SD) difference between the creatinine clearance measured by the 24-h and DTPA clearance technique was 17.7 (16.5)% and the correlation between these techniques was good (r = 0.71). CONCLUSION: Serum creatinine level is not sensitive in detecting early deterioration of renal function in patients with SCI. The Cockcroft-Gault formula generally significantly overestimates the true creatinine clearance and is not recommended. The 24-h endogenous creatinine clearance measured on appropriately collected urine samples is an acceptable accurate and practical method of determining glomerular filtration rate in patients with SCI. PMID- 10848687 TI - Bacteriological safety and cost-effectiveness of a nonrefluxing valve in the irrigation system during outpatient flexible cystoscopy. AB - OBJECTIVE: To critically evaluate the infection rate associated with the use of a nonrefluxing irrigation system for outpatient flexible cystoscopy, by comparing it with conventional irrigation systems, and to determine the costs and benefits of this system of irrigation delivery. PATIENTS AND METHODS: All patients undergoing flexible cystoscopy on scheduled outpatient lists were considered for inclusion in the study; 143 patients of 220 undergoing cystoscopy fulfilled the study criteria. The study group of patients underwent cystoscopy with a new single-use nonrefluxing valve inserted into the same irrigation delivery system that was used for the whole endoscopy session, and the control group had the complete irrigation system changed after each endoscopic examination. Midstream urine samples were taken for analysis before cystoscopy and again 3-4 days later. Infection was defined as a pure growth of >/= 105 organisms/mL, with associated pyuria defined as >/= 10 pus cells per high power microscopic field. RESULTS: Complete data were available on 133 patients, with a further nine being excluded from the statistical analysis because they had a pre-existing urinary tract infection, leaving 64 patients in the study and 69 in the control groups. The overall infection rate for flexible cystoscopy was 3.2%, with no significant difference between the study and control groups. Cost savings of > 35% can be expected using the nonrefluxing valve method of irrigation delivery. CONCLUSION: The nonrefluxing valve caused no detectable increase in patient morbidity from bacterial infection when compared with conventional irrigation systems for flexible cystoscopy, and can save considerable costs. PMID- 10848688 TI - Rethinking the secondary prevention of superficial bladder cancer: is there a role for retinoids? PMID- 10848689 TI - A prospective pilot evaluation of urinary and immunohistochemical markers as predictors of clinical stage of urothelial carcinoma of the bladder. AB - OBJECTIVE: To determine, in patients newly diagnosed with bladder cancer, whether p53, epidermal growth factor receptor (EGFR), microvessel density (MVD), urinary bladder tumour antigen (BTA TRAKtrade mark, Bion Diagnostic Sciences, Redmond, WA) and cytology were predictive of clinical stage, evaluated as a function of the clinical stage obtained at transurethral resection of the bladder tumour with and without the addition of clinical grade, a known strong predictor of clinical stage. PATIENTS AND METHODS: Between December 1997 and September 1998, 22 men and seven women with a cystoscopic diagnosis of urothelial bladder carcinoma were prospectively enrolled in the study. Urine was collected for cytological and BTA TRAK evaluation before transurethral resection. Tumour grade and clinical stage were obtained from the transurethral resection specimen. MVD was evaluated by computerized calculations of 'optimal MVD' (OMVD) and 'area-weighted MVD' (AWMVD) while p53 and EGFR information was obtained by manual immunohistochemical techniques; 21 patients had sufficient tissue for all immunohistochemical assessments and comprised the study group. Univariate and multivariate comparisons were carried out to determine the contribution of each variable to the prediction of clinical stage. RESULTS: Although there was a trend, cytological analysis and p53 and MVD immunoreactivity did not significantly correlate with clinical stage, while tumour grade, BTA TRAK and EGFR immunoreactivity did. In a univariate analysis, tumour grade and BTA TRAK were related to clinical stage. In a multivariate analysis, grade was the single best predictor of clinical stage. This analysis also showed that the addition of BTA TRAK and MVD information to grade incrementally improved the predictive ability of grade. CONCLUSIONS: This pilot study suggests that BTA TRAK and MVD contribute incremental information to tumour grade in predicting the clinical stage of urothelial carcinomas of the bladder; grade remains the most important predictor. These results suggest that further work with BTA TRAK and MVD in more patients and on biopsy material obtained during clinic cystoscopy is warranted for the future development of less invasive methods of identifying patients with invasive bladder cancer. PMID- 10848690 TI - Morbidity and the impact on daily activities associated with catheter drainage after acute urinary retention. AB - OBJECTIVE: To evaluate the effect on quality of life of being discharged home with a catheter before definitive treatment in patients with acute urinary retention (AUR). PATIENTS AND METHODS: Patients attending the emergency department with AUR were assessed and discharged home with a catheter if they fulfilled predetermined criteria. They were admitted to the day-care unit for urological assessment and completed a disease-specific quality-of-life questionnaire. RESULTS: Of 101 patients presenting to the emergency department in AUR, 84 were sent home after catheterization (83%); 78 (93%) patients completed the questionnaire. The major side-effects reported were urinary leak (46%), mild haematuria (44%), urgency (42%), pain around the penis (42%), painful erection (31%) and catheter blockage (26%). Only 12% of patients felt having a catheter was very inconvenient and 93% would find it acceptable to have a catheter in future. CONCLUSION: A significant minority of patients discharged home with a catheter had side-effects related to their catheter but were not greatly inconvenienced, and their capacity to carry out normal daily activities was not impaired. The immediate discharge of patients in AUR and planned treatment will enable better use of inpatient urology resources. PMID- 10848691 TI - Are men with lower urinary tract symptoms at increased risk of prostate cancer? A systematic review and critique of the available evidence. PMID- 10848692 TI - The usefulness of power Doppler ultrasonography for diagnosing prostate cancer: histological correlation of each biopsy site. AB - OBJECTIVE: To correlate the findings of power Doppler ultrasonography (PDUS) of the prostate with those of site-specific transrectal ultrasonography (TRUS) guided biopsy. PATIENTS AND METHODS: The study comprised 28 patients referred to our institution for TRUS-guided prostate biopsy because of an elevated PSA level and/or abnormal digital rectal examination. PDUS findings were graded 0, 1 or 2; grades 0-1 were considered as negative and grade 2 as positive. The blood volume of each biopsy site was also determined using the mean number (MN) value that represents the average vascularity in a 5-mm square sample. PDUS values were correlated with the histological findings of 147 biopsies with 19 focal lesions. RESULTS: Grade 2 was assigned to 19 sites, grade 1 to 52 sites, and grade 0 to 76 sites. Fourteen of the 19 PDUS findings of grade 2 sites revealed carcinoma and five were grade 1. Ten of 35 TRUS-positive sites were carcinomas, three benign prostatic hyperplasia (BPH) and 22 normal. The MN value for prostatic carcinoma was 4.33, for BPH 11.7 and for normal tissue 4.7. The overall sensitivity of PDUS was 74%, the specificity 96% and the positive predictive value 74%. CONCLUSIONS: Because TRUS alone cannot detect all cancers, PDUS should be used routinely in all patients undergoing TRUS-guided biopsy, to improve the diagnostic yield of prostate cancer. PMID- 10848693 TI - Transrectal power Doppler imaging in the detection of prostate cancer. AB - OBJECTIVES: To evaluate the clinical utility of transrectal power Doppler imaging (PDI) of the prostate for detecting prostate cancer in patients with abnormally high serum levels of prostate specific antigen (PSA). PATIENTS AND METHODS: Patients (107) with abnormally high serum PSA levels were assessed using a digital rectal examination (DRE), transrectal ultrasonography (TRUS) and PDI. Any hypervascular lesion on PDI was graded on a scale of 0-3, where grade 1-3 was considered positive and grade 0 negative. Patients were then diagnosed by prostatic needle biopsy and the results compared with the other detection methods. RESULTS: Needle biopsy confirmed prostate cancer in 41 (24%) of the 170 patients. PDI was positive in 68, of whom 40 (59%) had prostate cancer; all those but one having prostate cancer were positive on PDI. Thus, PDI had a high sensitivity of 98% (40/41) and a negative predictive value of 99% (101/102). PDI could have saved a significant number of patients from undergoing unnecessary biopsies, compared with DRE and TRUS (P < 0.001). CONCLUSION: The use of PDI in detecting prostate cancer might reduce the number of unnecessary needle biopsies of the prostate in patients with abnormally high serum PSA levels. PMID- 10848694 TI - Trends in prostate cancer incidence, mortality and survival in England and Wales 1971-1998. AB - OBJECTIVES: To examine trends in prostate cancer incidence and mortality in England and Wales between 1971 and 1998, using a newly developed and validated national cancer database and the national mortality database. METHODS: Age standardized incidence and death rates were calculated directly and trends in relative survival rates among men with prostate cancer registered during 1971 1990 were examined. RESULTS: The annual number of new cases of prostate cancer registered in England and Wales increased by 179% between 1971 and 1993, from 6174 to 17 210. Directly age-standardized incidence rates increased by 104% between 1971 and 1993, from 29 to 59 per 100 000. The number of deaths from prostate cancer increased by 113% between 1971 and 1998, from 4027 to 8570. Directly age-standardized death rates increased by 49% between 1971 and 1995 and then decreased by 8% between 1995 and 1998, an overall increase of 38% (20 to 27 per 100 000) between 1971 and 1998. The relative survival rate for prostate cancer among men diagnosed during 1986-1990 was 77% at 1 year and 42% at 5 years, compared with 67% and 33%, respectively, for cases diagnosed during 1971-1975. The increase in survival rates was confined to men diagnosed with prostate cancer up to 1985 and no increase was seen for cases diagnosed after 1985. CONCLUSIONS: Prostate cancer is becoming a growing burden on the health service. The explanation for the large increase in prostate cancer incidence and mortality is unclear and needs further investigation. The lack of any improvement in survival rates in cases diagnosed after 1985 is of concern, and suggests that the current management of prostate cancer in both primary and secondary care may need to be reviewed. PMID- 10848695 TI - Mean time to cancer-specific death of apparently clinically localized prostate cancer: policy implications for threshold ages in prostate-specific antigen screening and ablative therapy. AB - OBJECTIVE: To assess the mean time to cancer-specific death in patients with prostate cancer, using a minimum follow-up of 14 years at one institution, and thus evaluate the minimum life-expectancy for eligibility for radical surgery, radiotherapy and, implicitly, prostate specific antigen (PSA) screening. PATIENTS AND METHODS: The tumour registry of the authors' institution was searched for the records of patients with prostate cancer (stages T1 and nonmetastatic T2-3) over the period 1976-1983, chosen to give a maximum follow-up with sufficient numbers of patients. Kaplan-Meier curves and the mean time to death to 1995 for palpable and impalpable cancers were calculated. Deaths not from cancer and from unknown causes were censored. Patients still alive were also censored, except for in the calculation of mean time to death. RESULTS: Patients with both stages of disease had a steep increase in mortality at 16 years. The mean (SEM) time to prostate cancer-specific death for T1 disease was 17 (1.8) years and for T2-T3 (palpable) was 11.7 (1.2 years). CONCLUSION: These results suggest that, if there is to be one at all, the upper age limit for prostate cancer screening should be 62 years. PMID- 10848696 TI - Early catheter removal in 100 consecutive patients undergoing radical retropubic prostatectomy. AB - OBJECTIVES: To investigate the outcome of 100 consecutive patients selected for early catheter removal after radical retropubic prostatectomy (RRP), where urethral catheter drainage is used routinely for 2-3 weeks. PATIENTS AND METHODS: The study included 129 consecutive patients with clinically localized prostate cancer who underwent RRP. Catheters were removed in the clinic (with no radiographic studies) 8-9 days after RRP provided there was no evidence of urine leak, pelvic haematoma, rectal injury or severe obesity. The follow-up (mean 21 months) results were available for 118 patients, 100 of whom were candidates for early catheter withdrawal. Their records were reviewed for evidence of complications, including urinary retention, anastomotic stricture formation and urinary incontinence. RESULTS: Urinary retention developed in two of the 100 patients, requiring simple catheter replacement. Nine patients developed bladder neck contracture requiring dilatation or incision. No patients developed anastomotic disruption, urinary tract infection or pelvic abscess. At the mean follow-up of 21 months, 76% of patients were continent and did not require pads; 19% of patients had mild stress urinary incontinence requiring the use of 4 pads/day. CONCLUSION: With appropriate patient selection as described, catheters can be removed in the clinic (with no radiographic studies) 8-9 days after RRP, with no increased incidence of complications, including anastomotic stricture, retention or incontinence. PMID- 10848697 TI - Stilboestrol plus adrenal suppression as salvage treatment for patients failing treatment with luteinizing hormone-releasing hormone analogues and orchidectomy. AB - OBJECTIVE: To investigate the efficacy of low-dose stilboestrol (SB) with hydrocortisone in patients with advanced prostate cancer refractory to androgen suppression. PATIENTS AND METHODS: Thirty-four consecutive patients (median age 70 years, range 51-83) with metastatic disease who progressed on hormone therapy, as shown by recurrent/worsening symptoms and an increase in prostate-specific antigen (PSA) level, were recruited and discontinued hormonal treatment before starting SB. Patients received SB (1 mg/day) combined with hydrocortisone (40 mg/day). In an attempt to reduce the incidence of thrombo-embolic events, aspirin (75 mg/day) was also added. RESULTS: Stilboestrol was the second-line treatment in 19 patients and the third or fourth in 15. The median (range) duration of treatment with SB was 5 (0.5-21) months and the median follow-up 7.5 months, with 18 patients still alive and 14 still on treatment. Of 29 symptomatic patients, 24 had symptomatic improvement and five had no clear benefit; the median duration of benefit was 6 (2-21) months. The PSA level decreased by 0-50% in six patients, by 50-90% in 13 and by > 90% in eight, while there was symptomatic improvement in these three categories in five, 11 and seven patients, respectively. The median times to PSA nadir and progression were 4 and 6 months, respectively. Some thrombo-embolic events and fluid retention occurred but overall the treatment was well tolerated. CONCLUSION: Low-dose SB with hydrocortisone is effective in refractory prostate cancer, although there is some toxicity. Randomized studies against hydrocortisone or SB alone are needed to establish the cost/benefit ratio of this combination. PMID- 10848698 TI - Benign prostatic hyperplasia and prostate carcinoma in native Africans. AB - OBJECTIVE: To study the factors associated with morbidity and mortality in benign prostatic hyperplasia (BPH) and carcinoma of the prostate in native Africans. PATIENTS AND METHODS: A prospective study was conducted from 1993 to 1998 at the Ahmadu Bello University Teaching Hospitals, Zaria, Nigeria. During this 5-year period 686 patients were investigated and treated for symptoms and signs of prostatism. They were followed up for a mean (range) of 19.5 (1-60) months. RESULTS: BPH was found in 588 and clinical carcinoma in 98 patients. Adequate results, including a histological diagnosis, were available for 640 patients; there were 545 patients with BPH and 95 patients with histologically diagnosed prostate cancer. Treatment consisted of open prostatectomy for BPH, and subcapsular orchidectomy and/or open bladder-neck wedge resection for patients with prostate cancer and bladder neck obstruction. Within 6 months of surgery, four of 545 (0.7%) patients with BPH and 25 of 95 (26. 3%) with prostate cancer had died. Two-thirds of the patients with cancer presented with paraparesis or paraplegia. CONCLUSIONS: BPH and prostate cancer cause significant morbidity and mortality in African men. There is a need for health education about the early recognition of symptoms. Provision of facilities for transurethral prostatectomy would minimize the complications of surgery and ensure better use of the meagre resources available for health care. PMID- 10848699 TI - Would prostate cancer detected by screening with prostate-specific antigen develop into clinical cancer if left undiagnosed? A comparison of two population based studies in Sweden. AB - OBJECTIVE: To assess the risk of over-diagnosing and over-treating prostate cancer if population-based screening with serum prostate-specific antigen (PSA) is instituted. PATIENTS AND METHODS: From a serum bank stored in 1980, PSA was analysed in 658 men with no previously known prostate cancer from a well-defined cohort from Goteborg, Sweden (men born in 1913); the incidence of clinical prostate cancer was registered until 1995. From the same area, and with the same selection criteria, another cohort of 710 men born in 1930-31, who in 1995 accepted an invitation for PSA screening, was also analysed. RESULTS: Of men born in 1913, 18 (2.7%) had died from prostate cancer and the cumulative probability of being diagnosed with clinical prostate cancer was 11.1% (5.0% in those with a PSA level of < 3 ng/mL vs 32.9% in those with a PSA level of > 3 ng/mL, P < 0.01). The mean lead-time from increased PSA (> 3 ng/mL) to clinical diagnosis was 7 years. The prostate cancer detection rate in men born in 1930-31 was 4.4% (22% among those with increased PSA levels) and 30 of 31 detected cancers were clinically localized. CONCLUSIONS: Screening and sextant biopsies resulted in a lower detection rate (22%) than the cumulative risk of having clinical prostate cancer (33%) in men with increased PSA levels, indicating that under-diagnosis rather than over-diagnosis is the case at least with 'one-time' screening. Even if the stage distribution in screening-detected cancers seems promising (and thus may result in reduced mortality) it is notable that screening 67-year-old men will result in treatment a mean of 7 years before clinical symptoms occur and only one in four men anticipated to develop prostate cancer will die from the disease within 15 years. Large randomized screening trials seem mandatory to further explore the benefits and hazards of PSA screening. PMID- 10848700 TI - Comparison of modified one- and two-layer microsurgical vasovasostomy. AB - OBJECTIVE: To compare the outcome of a modified one-layer and two-layer vasovasotomy (VV) in two groups of similar men undergoing vasectomy reversal. PATIENTS AND METHODS: The charts and surgical records of all surgical procedures performed on men undergoing a modified one- or two-layer VV between June 1992 and July 1994 were retrospectively reviewed. A successful outcome (patency) was defined as sperm present at follow-up (mean follow-up 8 weeks). A modified one layer VV was used in 17 men (group 1) and a two-layer VV in 23 (group 2). RESULTS: Sperm were present in both groups if surgery was undertaken after vasal obstruction lasting < 36 months. The modified one- and two-layer VV had equal patency (88% and 90%, respectively) when undertaken after an obstructed interval of 36-96 months; outcomes were poorer if surgery was performed after > 96 months. The mean operative duration was 96 min for a modified one-layer VV and 167 min for the two-layer VV. CONCLUSIONS: The simpler and faster modified one-layer VV provides sufficient accuracy for successful vasectomy reversal in most cases. For most patients, both procedures have equivalent patency. PMID- 10848701 TI - Intravasal azoospermia: a surgical dilemma. AB - OBJECTIVES: To determine the incidence of intravasal azoospermia (IVA) and evaluate which factors before and during surgery influence outcome, by prospectively and intentionally performing bilateral vasovasostomies (VVs) only in men with intraoperative IVA. PATIENTS AND METHODS: Using a multilayer technique, 472 men underwent microsurgical reconstructive procedures. Intravasal fluid was examined for sperm by the surgeon and a pathologist. Strict enrolment criteria included total absence of sperm or sperm parts and bilateral VV as a treatment procedure. Patients were followed up by semen analysis and paternity assessed only by naturally conceived pregnancies. RESULTS: Of the 472 patients, 27 (5.7%) had bilateral IVA; 15 of these patients were available for a follow-up of 1-47 months. Eleven patients had identical gross appearance of intravasal fluid bilaterally. Of these patients, five had sperm in the ejaculate after surgery (three with clear intravasal fluid and two with no fluid). Bilaterally different vasal fluid was found in four men. Unilateral clear fluid was present in three patients, two of whom had sperm in semen analysed after VV. Overall, there was sperm in the ejaculate in seven of 15 patients with IVA; five of these seven had clear fluid in at least one vas deferens. One patient with unilaterally clear fluid achieved paternity by a naturally conceived pregnancy. The difference between the mean (SEM) obstruction interval in men who had sperm in a semen sample after VV, at 16.7 (3. 30) years, and in persistently azoospermic patients, at 15.5 (1.89) years, was not statistically significant (P = 0.741). CONCLUSION: The results of VV in patients with IVA are unsatisfactory; the patency rate is higher in men with copious clear fluid in at least one vas. The obstructive interval in patients with IVA does not appear to influence the outcome of VV. PMID- 10848702 TI - The postejaculatory refractory period: a neurophysiological study in the human male. AB - OBJECTIVE: To investigate changes in the penile sensory threshold, and the variables of cortical somatosensory evoked potential and sacral evoked response tests in the early postejaculatory period in the human male. SUBJECTS AND METHODS: Twenty healthy volunteers (mean age 25.3 years, range 17-32) were evaluated before and after ejaculation for penile sensory threshold values, and the variables of sacral evoked response and cortical somatosensory evoked potential tests. RESULTS: Three subjects were excluded from the statistical analyses because there were significant differences among the repeated tests. In the remaining 17 subjects the penile sensory threshold was significantly greater in the postejaculatory period (P < 0.05). There were no significant changes in the other variables of both the sacral evoked response and cortical somatosensory evoked potential tests after ejaculation (P > 0.05). CONCLUSION: The postejaculatory refractory period in the human male is accompanied by a greater penile sensory threshold but with no change in the values of the sacral evoked response and cortical somatosensory evoked potential. PMID- 10848703 TI - Epididymectomy is an effective treatment for scrotal pain after vasectomy. AB - OBJECTIVE: To investigate the efficacy of epididymectomy in patients with significant scrotal pain after vasectomy. PATIENT AND METHODS: Sixteen patients were identified retrospectively to have undergone epididymectomy for pain after vasectomy; 19 epididymectomies were performed (three bilateral and 13 unilateral). Details from the preoperative investigations, histological examination and follow-up of symptoms were analysed and correlated. Outcomes were initially assessed at the routine outpatient clinic review 3 months after surgery and the long-term outcomes were assessed by a telephone interview 3-8 years after epididymectomy (mean 5.5 years). RESULTS: Of the 16 patients, 14 had excellent initial symptomatic benefit from epididymectomy. At 3-8 years afterward, nine of 10 patients interviewed had a sustained improvement of their scrotal pain. The following were indicators of a poor outcome: atypical symptoms including testicular or groin pain; erectile dysfunction and normal appearance of the epididymis on ultrasonography. Patients with bilateral scrotal pain can have a good outcome after epididymectomy. CONCLUSION: Epididymectomy in well-selected patients is a reliable and effective treatment for pain after vasectomy. PMID- 10848704 TI - Clinical, physical, sperm and hormonal data in 251 adults operated on for cryptorchidism in childhood. AB - OBJECTIVE: To determine the clinical, physical, sperm and hormonal status during adulthood in cryptorchid patients operated on during childhood, and to assess these variables according to the age at surgery and preoperative testicular position. PATIENTS AND METHODS: A letter was sent to all 890 patients > 18 years old who underwent surgery for cryptorchidism during childhood; 274 responded and were assessed using a sexual history, physical examination, pituitary axis study and sperm analysis. Data were complete for 251 patients (mean age 21 years, range 18-30); 196 had unilateral (25 anorchic) and 55 had bilateral cryptorchidism. Clinical, surgical and anatomopathological records at surgery during childhood were also reviewed. RESULTS: The mean (SD) age at orchidopexy was 6.4 (3.3) years and the mean age at assessment 21.1 (2.7) years. Semen samples generally showed abnormalities; 10 patients (4%) with bilateral and 57 (23%) with unilateral cryptorchidism had semen values within the normal range. There was no correlation between the sperm count and age at surgery for unilateral or bilateral cases (Spearman test). There was a significant correlation (r = - 0.41) between the sperm count and level of follicle-stimulating hormone (FSH). There were no significant differences in the sperm count for patients with different testicular locations (ANOVA) or in those treated or untreated with human chorionic gonadotrophin (Student's t-test). The only significant relationship (P < 0.001) was between the sperm count and unilateral or bilateral cryptorchidism. CONCLUSIONS: The sperm quality of adults operated on for cryptorchidism during childhood is independent of the age at surgery or testicular location, but is influenced by whether the cryptorchidism was unilateral or bilateral. FSH levels were negatively correlated with sperm density. PMID- 10848705 TI - The changing urodynamic pattern from infancy to adolescence in boys with posterior urethral valves. AB - OBJECTIVE: To determine whether bladder dysfunction in boys with posterior urethral valves (PUV) changes from a uniform pattern of hypercontractility during infancy to the hypocontractility found in adolescence, by reviewing serial urodynamic studies. PATIENTS AND METHODS: Thirty boys with PUV and no voiding symptoms underwent a total of 86 urodynamic tests (mean 2.8 each). The first urodynamic study was undertaken at 1-4 years of age in 15 boys and at 5-13 years in 15. They were re-evaluated at least 3 years later; 15 patients underwent the first and last urodynamic study, respectively, at a mean age of 2.8 and 7.7 years (group A), 10 boys at 6.2 and 8. 8 years (group B) and five at 9.4 and 15.2 years (group C). In 10 boys aged > 5 years the first and last pressure-flow studies (PFS) were analysed using an advanced analysis (PFA) to better identify hypocontractility. RESULTS: Bladder dysfunction was found in 21 of 30 (70%) boys at the first evaluation and in 18 (60%) at the last. In 25 boys the urodynamic pattern changed. Of the 15 boys in group A, 10 of 12 who had hypercontractility changed to normal (seven), low compliance (one) or hypocontractility (two), and two remained stable; two of the remaining three with normal urodynamic studies changed to hypocontractility, while one was unchanged. Among the 10 boys in group B, six with hypercontractility changed to normal (three) or hypocontractility (three); two with normal urodynamic findings and one with low compliance changed to hypocontractility. Of the five boys in group C who showed severe hypocontractility after puberty, three had a normal pattern, one low compliance and one hypercontractility before puberty. The PFA showed a 'weak' detrusor in four of the seven boys who were considered normal on standard PFS. At the urodynamic follow-up, the PFA pattern changed to a 'weak' detrusor in four boys who had a normal (two) or strong (two) detrusor at the first evaluation. CONCLUSIONS: Bladder dysfunction in boys with PUV changes during childhood and through adolescence. The urodynamic pattern of hypercontractility generally found soon after valve ablation gradually changes to hypocontractility in many boys and this pattern seems to be the rule after puberty. The evidence from this series supports the hypothesis that long-term detrusor hyperactivity in boys with PUV leads to detrusor failure, but a longitudinal 15-year follow-up from birth to puberty is needed to validate this concept. PMID- 10848706 TI - The effect of bladder outlet obstruction on tissue oxygen tension and blood flow in the pig bladder. AB - OBJECTIVE: To investigate the effect of partial bladder outlet obstruction on detrusor blood flow and oxygen tension (PdetO2) in female pigs. MATERIALS AND METHODS: Detrusor-layer oxygen tension and blood flow were measured using oxygen sensitive electrode and radiolabelled microsphere techniques in five female Large White pigs with a partial urethral obstruction and in five sham-operated controls. The effects of chronic outlet obstruction on bladder weight, and cholinergic nerve density and distribution, are also described. RESULTS: In the obstructed bladders, blood flow and oxygen tension were, respectively, 54.9% and 74.3% of control values at low bladder volume, and 47.5% and 42.5% at cystometric capacity. Detrusor blood flow declined by 27.8% and 37.5% in the control and obstructed bladders, respectively, as a result of bladder filling, whilst PdetO2 did not decrease in the controls, but fell by 42.7% in the obstructed bladders. Bladder weight increased whilst cholinergic nerve density decreased in the obstructed animals. CONCLUSION: In pigs with chronic bladder outlet obstruction, blood flow and oxygen tension in the detrusor layer were lower than in control animals. In addition, increasing detrusor pressure during filling caused significantly greater decreases in blood flow and oxygen tension in the obstructed than in the control bladders. PMID- 10848707 TI - The production of nerve growth factor by human bladder smooth muscle cells in vivo and in vitro. AB - OBJECTIVE: To measure the concentrations of nerve growth factor (NGF) in tissue biopsies taken from subjects with a normal bladder and from patients diagnosed to have idiopathic detrusor instability (associated with a reduction in the density of motor nerves), and to use an in vitro model to study the mechanisms of NGF expression. MATERIALS AND METHODS: Biopsy specimens were obtained during endoscopic and open surgery from patients undergoing routine bladder surgery. The patients were divided into two categories based upon urodynamic characterization. The NGF content in samples from 11 normal bladders and seven idiopathic unstable bladders were measured using an enzyme-linked immunosorbent assay. The mechanisms influencing net NGF production were explored using detrusor cells in vitro. RESULTS: The mean (SEM) NGF content was significantly higher in unstable tissues, at 0.96 (0.05) pg/microg protein, than in the normal bladder, at 0.53 (0.05) pg/microg protein. In the cell model, acetylcholine (10 micromol/L), noradrenaline (1 and 10 micromol/L) and ATP (1 micromol/L) caused a significant increase in net NGF production; acetylcholine at 1 micromol/L had no effect. Direct stimulation of protein kinase C (PKC) by phorbol ester (33 ng/mL) or elevation of cAMP using forskolin (10 micromol/L) increased NGF, suggesting that at least two intracellular pathways (PKC- and PKA-dependent) are involved. The expression of c-Fos was increased by phorbol 12-myristate 13-acetate added before NGF, suggesting that c-Fos may be involved in regulating NGF production. CONCLUSION: These data suggest a role for NGF in the physiology and pathophysiology of the human bladder, and indicate some of the possible mechanisms which might regulate NGF production. PMID- 10848708 TI - Gene transfer to urethral strictures in rabbits: a preliminary report. AB - OBJECTIVE: To assess the rationale for virus-mediated gene transfer into the urethra in vivo and in vitro, using a rabbit model, as this is an attractive approach to prevent recurrence after the endoscopic management of urethral strictures. MATERIALS AND METHODS: Primary cultures of rabbit urethral stromal cells were infected with adenoviral and retroviral solutions carrying a nucleus targeted beta-galactosidase (beta-Gal) reporter gene (respectively 109 and 107 plaque-forming units/mL). In addition, to mimic the human clinical situation, a model was developed of thermally induced stricture in rabbit urethra which produced fibrotic stenosis within 15 days. Using a prototype channelled balloon catheter, these strictures were endoscopically dilated and then instilled with the beta-Gal adenoviral or retroviral constructs. RESULTS: The application of recombinant adenovirus and retrovirus harbouring a nucleus-targeted beta-Gal reporter gene to cultured rabbit urethral stromal cells resulted in a high transduction efficiency of up to 90% and 96%, respectively. Five days after infection, histochemical and immunohistochemical staining of the strictured urethrae showed a 3% rate of transfection targeted to stromal cells within the fibrosis, confirmed by polymerase chain reaction (PCR) analysis. Adjacent and distal spread of the virus was excluded by histochemistry, immunohistochemistry and PCR. CONCLUSION: These results represent the first report of endoscopic adenovirus and retrovirus-mediated gene transfer to the urethra. Although at a low rate, transduction reached stromal cells transmurally within the induced strictures and was site-specific. PMID- 10848709 TI - Active properties of the urinary bladder: in vitro comparative studies between adult and neonatal rats. AB - OBJECTIVE: To determine, using in vitro comparative studies, developmental aspects associated with the active properties of the urinary bladder in neonatal and adult rats. MATERIALS AND METHODS: Urinary bladders were removed from neonatal (1-3 days old) and adult (15 weeks old) male Sprague-Dawley rats. Anterior longitudinal muscle strips were obtained from each group and isometric tensions recorded. Nerve-mediated contractions elicited by electrical field stimulation (0.8 ms pulse) or carbachol-induced contractile responses in neonatal and adult bladder strips were compared. Contractile tensions were normalized using the wet weight of the bladder strip or by using the percentage contraction induced by 60 mmol/L KCl. RESULTS: Nerve-mediated contractile responses showed that the muscarinic component predominated in the neonatal rat bladder, which contrasted with the predominant purinergic components in adult bladder. The pattern of spontaneous activity and carbachol-induced contraction differed in the two groups. Small spontaneous contractions in the basal state occurred in adult bladder strips, while short-lived large spontaneous contractions were present in neonatal strips. The amplitude of carbachol-induced contractions generated in the neonatal bladder was larger than that in the adult bladder. In addition, the time to achieve peak contraction elicited by carbachol (5 micromol/L) was shorter in the neonatal bladder. Repetitive carbachol applications induced an attenuation of the contractile response (desensitization), but the neonatal bladder was more resistant to desensitization than the adult bladder. CONCLUSIONS: These results show that nerve-mediated or agonist-induced contractile patterns, and spontaneous activity, in the neonatal bladder differ from those of the adult bladder in rats. The results suggest that in addition to neural immaturity, there are functional differences between the bladders of adult and neonatal rats. PMID- 10848710 TI - Genes upregulated during castration-induced rat prostatic apoptosis: cloning and characterization of new cDNAs. AB - OBJECTIVE: To isolate new cDNAs corresponding to genes whose expression is increased during castration-induced rat prostate apoptosis. MATERIALS AND METHODS: Differential display of mRNAs from 3-day castrated and normal rat ventral prostates was used to identify differentially expressed clones. Northern blots were hybridized to confirm the positive regulation of the candidates and to follow the change in their expression in the involuting rat prostate, and in thymocytes of dexamethazone-treated rats. RESULTS: Five cDNAs were cloned: one encoding ribosomal protein L7, one coding for the insulin-like growth factor binding protein-3 (IGFBP-3), and three whose products are unknown. After castration, all five genes had expression kinetics that closely paralleled the proportion of prostatic epithelial cells undergoing apoptosis. The gene encoding L7 and two of the unknown genes were also upregulated in glucocorticoid-induced programmed death in thymocytes. In addition to the IGFBP-3 gene, those coding for proteins IGFBP-4, -5 and -6 were also overexpressed in the involuting prostate of androgen-deprived rats. CONCLUSION: Five new genes were identified that are up regulated during castration-induced rat prostate apoptosis, three of which are potentially involved in the common intracellular pathway leading to programmed cell death. PMID- 10848711 TI - Possible biphasic changes of free radicals in ethylene glycol-induced nephrolithiasis in rats. AB - OBJECTIVE: To evaluate the possible role of free radicals in nephrolithiasis in rats induced by ethylene glycol, and to examine the correlation between the urinary enzymes N-acetyl-beta-glucosaminidase (NAG), beta-galactosidase (GAL) and neutral endopeptidase (NEP), and free radical production. MATERIALS AND METHODS: Hyperoxaluria was produced in male Wistar rats by adding ethylene glycol to their drinking water. After 7, 21 and 42 days of treatment, urinary oxalate, creatinine clearance and urinary enzymes (NAG, GAL and NEP) were measured. The nitroblue tetrazolium perfusion method was used to locate the sites of free-radical production. Ultrasensitive chemiluminescence was used to directly measure the production of reactive oxygen species (ROS) in vivo. Vitamin E and potassium citrate were fed to rats, in addition to ethylene glycol, to assess their effects on free radical production. RESULTS: Urinary oxalate increased significantly and was associated with an increase in NAG and GAL at all sample times. However, urinary NEP activity was unchanged on day 7, although there was four times as much NEP on days 21 and 42 than in the control groups. Formazan particles in the renal cortex were scored as 3+ to 4+ in rats treated for 7 days with ethylene glycol. Blood ROS levels were also higher in this group than in the controls (P < 0.01). After vitamin E and potassium citrate treatment, blood ROS levels were lower than in rats treated with ethylene glycol alone. CONCLUSION: Free radicals may be produced in the early stages of nephrolithiasis in rats fed with ethylene glycol. Free radicals occurred mainly in blood and might be associated with NEP inactivation. PMID- 10848712 TI - Extravesical transitional cell carcinoma as a result of implantation after perforation of the bladder. PMID- 10848713 TI - Calcified nonfunctional paraganglioma of the urinary bladder mistaken as bladder calculus: a diagnostic pitfall. PMID- 10848714 TI - Clinical pharmacology and the faculty of pharmaceutical medicine. PMID- 10848715 TI - Methods to determine lung distribution of inhaled drugs - could gamma scintigraphy be the gold standard? PMID- 10848716 TI - Can lung deposition data act as a surrogate for the clinical response to inhaled asthma drugs? AB - Studies involving the direct measurement of clinical response to inhaled asthma drugs, especially inhaled corticosteroids, may be very difficult to conduct. However, the deposition of drug in the lungs may be considered as a measure of local bioavailability, and may be quantified by radionuclide imaging techniques, or for some drugs by pharmacokinetic methods. This paper reviews evidence for considering lung deposition data as a surrogate for the clinical response to inhaled asthma drugs, based mainly upon a series of case histories. The appropriate use of lung deposition data in regulatory packages, especially to document the equivalence or comparability of two products, offers the possibility of significant time saving in the drug development process, and hence a faster drug development programme for inhaled asthma products. PMID- 10848717 TI - Antidepressants in the elderly: challenges for study design and their interpretation. AB - Drug treatment of depressive illness in the elderly differs from that in younger patients and there is no clear consensus as to first line treatment in the former. Nor is it possible to extrapolate directly from studies in younger patients to the elderly with these agents. Whilst there are over two dozen antidepressants currently marketed in the U.K., most studies have been on younger adults and have excluded very old and frail patients. Design short-comings of the few trials conducted in elderly patients do not allow accurate interpretation of differences in efficacy or safety between drugs. This paper identifies key deficiencies in the evidence currently available in support of both older and newer antidepressant agents and makes the proposal that specific studies are required in the elderly to determine the efficacy and safety of antidepressants in the treatment of depressive illness. It outlines a Phase II and III clinical trial programme which could be used to provide adequate evidence of efficacy and safety of new agents and which conforms to current European guidelines. Dose finding studies, short-term efficacy, prevention of relapse (continuation therapy) and prevention of recurrence (maintenance therapy) studies are discussed as are key issues to be addressed in the trial protocol. PMID- 10848718 TI - Influence of hydroxychloroquine on the bioavailability of oral metoprolol. AB - AIMS: Hydroxychloroquine (HCQ) is used widely in the treatment of chronic inflammatory diseases such as rheumatoid arthritis. Since there is great interindividual variability in the pharmacokinetics of HCQ and chloroquine is a potent inhibitor of CYP2D6-catalysed pathways in vitro, we wished to study the interaction of HCQ with CYP2D6-mediated metabolism of other drugs in vivo. METHODS: Metoprolol and dextromethorphan (DM) were selected as probe drugs because they are well-studied and widely used test substrates of CYP2D6. In this randomized, double-blind crossover study, seven healthy volunteers with extensive metabolizer phenotype for CYP2D6 ingested either 400 mg hydroxychloroquine or placebo daily for 8 days after which single oral dose pharmacokinetics of metoprolol were investigated. Dextromethorphan metabolic ratio (DM-MR) was also determined at baseline and after the ingestion of HCQ or placebo. RESULTS: Concomitant administration of HCQ increased the bioavailability of metoprolol, as indicated by significant increases in the area under the plasma concentration time curve (65 +/- 4.6%) and maximal plasma concentrations (72 +/- 6.9%) of metoprolol. While the DM-MR values were not significantly changed, the phenotypic classification of one individual, who was heterozygous for a mutant CYP2D6 allele, was converted to a poor metabolizer by HCQ administration. CONCLUSIONS: HCQ inhibits metoprolol metabolism most probably by inhibiting its biotransformation by CYP2D6. The inhibitory effect of HCQ on dextromethorphan metabolism was not apparent when DM-MR was used as an indicator, except in an individual with limited CYP2D6 capacity. PMID- 10848719 TI - Evaluation of the autoinduction of ifosfamide metabolism by a population pharmacokinetic approach using NONMEM. AB - AIMS: This study investigated the population pharmacokinetics of ifosfamide in 15 patients treated for soft tissue sarcoma with 9 or 12 g m-2 ifosfamide by means of a 72 h continuous i.v. infusion. METHODS: A model was developed using nonlinear mixed effects modelling (NONMEM) to describe the nonlinear pharmacokinetics of ifosfamide by linking the ifosfamide plasma concentrations to the extent of the autoinduction. RESULTS: The proposed model revealed the effect of autoinduction on the disposition of ifosfamide. The initial clearance, volume of distribution, rate constant for enzyme degradation, induction half-life of the enzyme and the ifosfamide concentration at 50% of the maximum inhibition of enzyme degradation were estimated at 2.94 +/- 0.27 l h-1, 43.5 +/- 2.9 l, 0.0546 +/- 0. 0078 h-1, 12.7 h and 30.7 +/- 4.8 microM, respectively. Interindividual variabilities of initial clearance, volume of distribution, rate constant for enzyme degradation were 24.5, 23.4 and 22.7%, respectively. Proportional and additive variability not explained by the model were 13.6% and 0.0763 microM, respectively. CONCLUSIONS: The absence of a lag time for the autoinduction of ifosfamide metabolism could be the result of an immediate inhibition of the enzymatic degradation of CYP3A4 by ifosfamide. By application of the autoinduction model individual pharmacokinetic profiles of patients were described with adequate precision. This model may therefore be used in the future development of a model to individualize dose selection in patients. PMID- 10848720 TI - Pharmacokinetics and pharmacodynamic effects of ABT-627, an oral ETA selective endothelin antagonist, in humans. AB - AIMS: Endothelins (ETs) may play a role in the pathogenesis of a variety of cardiovascular diseases. The present study was designed to investigate the pharmacokinetic and pharmacodynamic effects of the orally active ETA selective receptor antagonist ABT-627 in healthy humans. METHODS: Healthy volunteers were included in two studies with cross-over design. Subjects received single or multiple dose (an 8 day period) administration of oralABT-627 or matched placebo, in a dose range of 0.2-40 mg. The pharmacokinetics of ABT-627 were described and its effects on systemic haemodynamics under resting conditions and on forearm vasoconstriction in response to ET-1 were assessed. RESULTS: ABT-627 was generally well tolerated in both studies, with transient headache being the most reported adverse event (in 62% vs 4% during placebo, P < 0.05, for Study 1 and in 42% vs 60%, P = 0.2, for Study 2). ABT-627 was rapidly absorbed, reaching maximum plasma levels at approximately 1 h post dose. Single dose ABT-627, at a dose of 20 and 40 mg, inhibited ET-1 induced forearm vasoconstriction at 8 h post dose. Eight days ABT-627 treatment, at a dose level of 5 mg and above, also effectively blocked forearm vasoconstriction to ET-1. ABT-627 caused a significant reduction in peripheral resistance as compared with placebo (16 +/- 1 vs 19 +/- 1, 18 +/- 2 vs 23 +/- 3, 15 +/- 1 vs 17 +/- 1 AU at 1, 5, 20 mg in Study 2) with only a mild decrease in blood pressure (79 +/- 2 vs 84 +/- 3, 80 +/- 4 vs 90 +/- 5, 75 +/- 3 vs 79 +/- 1 at 1, 5, 20 mg in Study 2). ABT-627 caused a moderate dose-dependent increase in circulating immunoreactive ET levels (a maximal increase of 50% over baseline at the 20 mg dose level). CONCLUSIONS: The oral ETA receptor blocker ABT 627 is well tolerated, rapidly absorbed, effectively blocks ET-1 induced vasoconstriction and causes a decrease in total peripheral resistance and mean arterial pressure. Our data suggest that ABT-627 may be a valuable tool in treatment of cardiovascular disease. PMID- 10848722 TI - A comparison of the risks of venous thromboembolic disease in association with different combined oral contraceptives. AB - AIMS: In October 1995 in response to the results of three studies, the Committee on the Safety of Medicines advised doctors and pharmacists that oral contraceptives containing desogestrel (DSG) and gestodene (GST) were associated with around a two-fold increase in the risk of thromboembolism compared with those containing other progestogens. The objective of this study was to estimate the risk of idiopathic venous thromboembolic disease (VTE) in users of combined oral contraceptives (COCs), to compare the risk between formulations and to examine the effect of using age banding as opposed to matching by exact year of birth. METHODS: A nested case control study was conducted using the General Practice Research Database. Women with a VTE event recorded between 1992 and 1997, who were treated with an anticoagulant, from consideration of their prescription records were likely to have been using a COC prescription on the day of the event and also had no exclusion factors, were deemed cases. For comparison with the previous studies, two nested case control studies were undertaken. Study 1 used controls matched by practice and year of birth. Study 2 used controls matched by practice and within 5 years age bands. RESULTS: We found an incidence of idiopathic VTE amongst users of combined oral contraceptives of 3.8 per 10 000 exposed women years. Incidence rates increased markedly after 35 years of age. The nested case-control study using controls matched by year of birth showed no significant difference in risk between the major COC formulations. With levonorgestrel (LNG) 150 microgram and ethinyloestradiol (EE) 30 microgram as the reference, the adjusted ORs for GST 75 microgram and EE 30 microgram was 1.3 (95% CI 0.8, 2.1), for DSG 150 microgram and EE 30 microgram it was 1.0 (95% CI 0.7, 1.7) and for DSG 150 microgram and EE 20 microgram it was 0.8 (95% CI 0.4, 1.6). Using less rigorous matching criteria, matching controls to cases within 5 years age bands, the ORs increased. When a mixed group of COCs, characterized by having LNG as the progestogen component was used as the reference category, there was an elevation in the ORs for the newer products. We found a significant association between idiopathic VTE and current smoking (OR 2.0 (1.4, 2.7)), BMI over 35 (OR 3.8 (1.8, 8.0)) and asthma (OR 1.9 (1.3, 2.9)). The OR for women who had proxy evidence of general ill health (indicated by the number of prescriptions issued) was 2.2 (1.7, 3.7). CONCLUSIONS: The results of this study indicate that a number of the characteristics of the women taking COCs affect the risk of VTE. There is no evidence to support the hypothesis that there is any difference in risk between COC formulations containing under 50 microg ethinyloestradiol. PMID- 10848721 TI - Topical application of doxepin hydrochloride, capsaicin and a combination of both produces analgesia in chronic human neuropathic pain: a randomized, double-blind, placebo-controlled study. AB - AIMS: To assess the analgesic efficacy of topical administration of 3.3% doxepin hydrochloride, 0.025% capsaicin and a combination of 3. 3% doxepin and 0.025% capsaicin in human chronic neuropathic pain. METHODS: A randomized, double-blind, placebo-controlled study of 200 consenting adult patients. Patients applied placebo, doxepin, capsaicin or doxepin/capsaicin cream daily for 4 weeks. Patients recorded on a daily basis overall pain, shooting, burning, paraesthesia and numbness using a 0-10 visual analogue scale during the week prior to cream application (baseline levels) and for the 4 week study period. Side-effects and desire to continue treatment were also recorded. RESULTS: Overall pain was significantly reduced by doxepin, capsaicin and doxepin/capsaicin to a similar extent. The analgesia with doxepin/capsaicin was of more rapid onset. Capsaicin significantly reduced sensitivity and shooting pain. Burning pain was increased by doxepin and by capsaicin and to a lesser extent by doxepin/capsaicin. Side effects were minor. One patient requested to continue placebo cream, 17 doxepin cream, 13 capsaicin and 9 the combination of doxepin and capsaicin. CONCLUSIONS: Topical application of 3.3% doxepin, 0.025% capsaicin and 3.3% doxepin/0. 025% capsaicin produces analgesia of similar magnitude. The combination produces more rapid analgesia. PMID- 10848723 TI - Validation of the diagnosis of venous thromboembolism in general practice database studies. AB - AIMS: The study was conducted to determine whether the method for selecting cases of venous thromboembolism (VTE) from general practice databases significantly affected the findings of an epidemiological study. METHODS: Cases of VTE were identified from the UK General Practice Research Database (GPRD) by searching for codes for deep vein thrombosis (DVT) and pulmonary embolism (PE). These had to be supported by evidence of anticoagulation and be exposed to a combined oral contraceptive (COC) at the time of the event. Additional information about the event was sought from general practitioners who were requested to complete a questionnaire and to supply anonymised copies of hospital letters and discharge summaries. RESULTS: Of the 285 cases identified from the GPRD, additional information was available for 177 VTE events. This information showed that 84% of those events were supported by hospital investigations or a death certificate. Using only verified cases, rather than all GPRD identified events, did not alter the results of the epidemiological study. CONCLUSIONS: The GPRD provides information of sufficiently high quality to allow valid epidemiological research of VTE events. Excluding cases without a database record of hospital admission would lead to valid events being overlooked, and an under-estimate of the disease incidence. PMID- 10848724 TI - The completeness of medication histories in hospital medical records of patients admitted to general internal medicine wards. AB - AIMS: Accurate recording of medication histories in hospital medical records (HMR) is important when patients are admitted to the hospital. Lack of registration of drugs can lead to unintended discontinuation of drugs and failure to detect drug related problems. We investigated the comprehensiveness of medication histories in HMR with regard to prescription drugs by comparing the registration of drugs in HMR with computerized pharmacy records obtained from the community pharmacy. METHODS: Patients admitted to the general ward of two acute care hospitals were included in the study after obtaining informed consent. We conducted an interview on drugs used just prior to hospitalization and extracted the medication history from the HMR. Pharmacy records were collected from the community pharmacists over a 1 year period before the admission. Drugs in the pharmacy records were defined as possibly used (PU-drugs) when they were dispensed before the admission date and had a theoretical enddate of 7 days before the admission date or later. If any PU-drug was not recorded in the HMR, we asked the patient whether they were using that drug or not. RESULTS: Data were obtained from 304 patients who had an average age of 71 (range 40-92) years. The total number of drugs according to the HMR was 1239, 43 of which were not used. When compared with the pharmacy records we found an extra 518 drugs that were not recorded in the HMR but were possibly in use. After verification with the patients, 410 of these were indeed in use bringing the total number of drugs in use to 1606. The type of drugs in use but not recorded in the HMR covered a broad spectrum and included many drugs considered to be important such as cardiovascular drugs (n = 67) and NSAIDs (n = 31). The percentages of patients with 0, 1, 2, 3, 4, 5-11 drugs not recorded in the HMR were 39, 28, 16, 8, 3.6 and 5.5, respectively. Of the 1606 drugs in use according to information from all sources, only 38 (2.4%) were not retrievable in the pharmacy records when the complete year prior to hospitalization was evaluated. CONCLUSIONS: The medication history in the hospital medical record is often incomplete, as 25% of the prescription drugs in use is not recorded and 61% of all patients has one of more drugs not registered. Pharmacy records from the community pharmacist can be used to obtain more complete information on the medication history of patients admitted to the hospital. PMID- 10848725 TI - Antihypertensive drugs and the risk of idiopathic aplastic anaemia. AB - AIMS: A recent report has raised concern that nifedipine may be associated with an increased risk of aplastic anaemia. This large population-based study evaluated the risk of idiopathic aplastic anaemia in users of calcium channel blockers compared with that of other antihypertensive drugs. METHODS: The study was based on information derived from the General Practice Research Database. We conducted a follow-up study with a nested case-control analysis of 322 448 subjects who received antihypertensive drugs. Cases were people who had a first time diagnosis of aplastic anaemia during January 1, 1988 through September 30, 1997. The risk estimate of aplastic anaemia was calculated for all antihypertensive drugs. For the nested case-control analysis, six controls were matched to each case on age, sex and general practice attended. Odds ratios compared the risk of idiopathic aplastic anaemia for all antihypertensive drugs relative to nonusers. RESULTS: There were 13 cases of newly diagnosed idiopathic aplastic anaemia. The estimated risk of aplastic anaemia per 100 000 users was 0.8 (95% CI 0.1, 4.7) for calcium channel blockers, 1.4 (95% CI 0.5, 4.1) for beta-adrenoceptor blockers, 2.3 (95% CI 0.6, 8.6) for angiotension-converting enzyme (ACE) inhibitors and 5.9 (95% CI 1.6, 21.5) for users of other antihypertensive drugs. In the case-control analysis of 13 cases and 77 controls, the odds ratio was 0.3 (95% CI 0.02, 3.3) for calcium channel blockers, 0.5 (95% CI 0.1, 2.5) for beta-adrenoceptor blockers, 0.7 (95% CI 0.1, 5.6) for ACE inhibitors, 1.2 (95% CI 0.1, 11.8) for users of other antihypertensive drugs and 0.7 (95% CI 0.1, 7.2) for users of multiple drugs with a calcium channel blocker compared with nonusers. CONCLUSIONS: The present study suggests that the use of calcium channel blockers is not associated with an increased risk of aplastic anaemia. PMID- 10848726 TI - The link between sunshine and phototoxicity of sparfloxacin. AB - AIMS: To test the association between reporting rates for sparfloxacin-induced phototoxicity and sunlight u.v. exposure, and the effects of regulatory action. METHODS: The reporting rates for phototoxicity with sparfloxacin to the French Pharmacovigilance System or to the Drug Manufacturer were compared with concurrent national mean u.v. exposure obtained from Meteo-France, before and after the regulatory restrictions and warnings. RESULTS: There were 371 severe phototoxic reaction reports during the first 9 months of marketing (reporting rate of 0.4 per thousand treated patients), approximately four to 25 times that reported for other fluoroquinolones. The reporting rate correlated highly (r = 0.873, P < 0.001) with the mean monthly u.v. exposure from sunlight (from Meteo France). Regulatory action including warnings for physicians, and restricted indications dramatically decreased the number of reports, but not the reporting rate. CONCLUSIONS: This is the first demonstration of a strong association between sunlight exposure in a population and drug-induced phototoxicity. Regulatory action had no effect on the reporting rate (individual exposed patient risk), though it solved the public health issue. PMID- 10848727 TI - Mast cell mediators other than histamine induced pruritus in atopic dermatitis patients - a dermal microdialysis study. PMID- 10848728 TI - Antiphospholipid antibody syndrome: the problems and the promise. PMID- 10848729 TI - Comedogenesis: some new aetiological, clinical and therapeutic strategies. AB - Hypercornification is an early feature of acne and precedes inflammation. It is associated with ductal hyperproliferation and there are many controlling factors such as androgens, retinoids and cytokines. Cycling of normal follicles and of comedones may explain the natural resolution of comedones and, in the longer term, resolution of the disease itself. There is a need to tailor treatment according to comedonal type. Suboptimal therapy can often result from inappropriate assessments of comedones, especially microcomedones, missed comedones, sandpaper comedones, submarine comedones and macrocomedones. Macrocomedones can produce devastating acne flares, particularly if patients are inappropriately prescribed oral isotretinoin. Gentle cautery under topical local anaesthesia is a useful therapy in the treatment of such lesions. The newer retinoids and new formulations of all-trans-retinoic acid show a better benefit/risk ratio. Evidence-based studies are required to allow adequate comparisons. PMID- 10848730 TI - c-Myc expression in human anagen hair follicles. AB - The hair follicle represents a very attractive organ system for studying the precise balance between cell proliferation, growth, differentiation, and death of cells, because it periodically and regularly regenerates, retaining its morphogenetic signals throughout its life. One of the most intriguing oncogenes which is able to induce both cell growth and apoptosis, depending upon the environmental conditions, is c-myc. The aim of the present study was to investigate its presence and localization in human hair follicles by immunohistochemistry and immunofluorescence. Our observations demonstrated the consistent presence of two clusters of c-Myc-expressing cells in anagen follicles, located in two annular regions of the inner root sheath, at the border between cells characterized by putative trichohyalin granules and cells which are keratinized. The lower group belongs to Henle's layer, while the upper group belongs to Huxley's layer. c-Myc oncoprotein seems to favour apoptosis/differentiation and may be a marker for terminal differentiation of trichocytes, at least in the inner root sheath. Our findings agree with the interpretation that the complex morphology of the hair follicle reflects its complex function; the extrusion of a highly organized multicellular structure, the hair shaft, driven by another highly organized multicellular structure, the inner root sheath. PMID- 10848731 TI - A novel PTEN mutation in a Japanese patient with Cowden disease. AB - Cowden disease (CD) is an autosomal dominant syndrome characterized by multiple hamartomatous lesions and an increased risk for malignancies. Recent evidence has indicated that the PTEN gene, encoding a protein tyrosine phosphatase, is the CD susceptibility gene. However, another line of evidence has suggested that CD might be genetically heterogeneous. Clinical features of CD are variable, and there are interfamilial differences in the expression of skin lesions. Therefore, information on PTEN mutations in CD patients should be accumulated to clarify the genotype-phenotype correlation. In the present study, we found heterozygous germline mutations of PTEN in all of three Japanese patients with CD examined, indicating no genetic heterogeneity among our patients. The mutations included two non-sense mutations of R335X and R130X, and a mis-sense mutation of C136R. To the best of our knowledge, the C136R mutation has not previously been reported in CD patients. This novel mutation was located outside the core motif of the phosphatase domain of PTEN protein, where most of the missense mutations previously reported in CD patients were clustered. Mucocutaneous manifestations were far fewer in the patient with this mutation than in the patients with nonsense mutations. Whether the phenotypic difference in mucocutaneous features was due to the different mutations remains unclear. PMID- 10848732 TI - Expression of Fc receptors for IgG during acute and chronic cutaneous inflammation in atopic dermatitis. AB - Atopic dermatitis is an allergic skin disease characterized by elevated total and antigen-specific serum IgE and IgG4 levels. In acute and chronic cutaneous inflammation, large cellular infiltrates including T cells, dendritic cells and macrophages are found, especially in the dermis. These cells play an important part in the regulation of local inflammatory reactions. Receptors binding IgG (FcgammaR) are involved in dendritic cell and macrophage function. In this study, we examined the in vivo distribution and cellular expression of the three classes of leucocyte FcgammaR in human skin during acute and chronic cutaneous inflammation in atopic dermatitis. Atopy patch test skin was used as a model for acute inflammation in atopic dermatitis, while chronic lesional skin was used to investigate FcgammaR expression in chronically inflamed skin. In atopy patch test sites no increase in the number of CD1a+ dendritic cells and a slight increase in macrophages compared with non-lesional skin was observed. Our results showed increased expression of FcgammaRI (CD64) and FcgammaRIII (CD16) in acutely inflamed skin as well as in chronically inflamed lesional skin, compared with healthy and non-lesional atopic dermatitis skin. FcgammaRI was expressed by RFD1+, RFD7+ and CD68+, but not by CD1a+ dermal dendritic cells. RFD1+ dendritic cells and CD68+ macrophages were the main FcgammaRIII-expressing cells during the acute inflammatory reaction. The significant increase in expression of FcgammaRIII (CD16) and FcgammaRI (CD64) probably results from upregulation of the receptors on resident cells. Insight into the presence of FcgammaR+ cells in human skin during inflammation is important both for our understanding of skin immune reactions and the development of new therapeutic concepts. PMID- 10848733 TI - Mast cell mediators other than histamine induce pruritus in atopic dermatitis patients: a dermal microdialysis study. AB - While histamine is the crucial mediator of pruritus in type 1 allergic reactions, its role in atopic dermatitis (AD) is unclear. In this study, the role of mast cell mediators in protein extravasation and pruritus was evaluated using intradermal microdialysis. The microdialysis capillaries were used to apply the mast cell degranulating substance compound 48/80 (C48/80; 0.05%) or histamine (0.01%) and also to deliver H1-blockers (cetirizine, 200 microg mL-1) in nine AD patients and nine controls. Large pore size membranes (3000 kDa) enabled simultaneous analysis of protein extravasation. Itch sensation was measured psychophysically and weal and flare reaction were evaluated planimetrically. Protein extravasation induced by histamine and C48/80 was significantly reduced in AD patients. Blockade of H1-receptors by cetirizine significantly reduced C48/80-induced protein extravasation in AD patients and controls to an identical level. C48/80-induced pruritus was abolished by cetirizine in controls, whereas pruritus in AD patients was unchanged after H1 blockade. We conclude that mast cell mediators others than histamine are involved in C48/80-induced pruritus in AD patients. Whether the reduced capacity of AD patients to induce protein extravasation is of pathophysiological relevance for pruritus remains to be established. PMID- 10848734 TI - N-acetyltransferase 1 and 2 polymorphisms in para-substituted arylamine-induced contact allergy. AB - Sensitization to arylamines such as p-phenylenediamine is frequently diagnosed in patients with allergic contact dermatitis. Reactive metabolites of p phenylenediamine might be produced in the skin by O-acetylation of N hydroxylamines catalysed by local N-acetyltransferases (NATs). In this study, we tested whether genetic polymorphisms of NATs, which are known to affect enzyme activity, may influence the susceptibility to para-substituted arylamine-induced contact eczema. Using polymerase chain reaction and restriction enzyme analysis, the distribution of polymorphisms of NAT1 and NAT2 was investigated in 88 patients sensitized to para-substituted aryl compounds and 123 healthy controls. NAT2 rapid acetylators, i.e. carriers of the NAT2*4 wild-type allele, were more common in the contact allergy (44%) than in the healthy control group [30%; P = 0.042, odds ratio 1.9 (95% confidence interval, CI 1. 05-3.27)]. Slow acetylators carrying the NAT2*5b/2*6a genotype were significantly less frequent among patients [13% vs. 38% in controls; P = 0.009, odds ratio 0.39 (95% CI 0.19 0.78)]. The carriage rate of the NAT1*10 allele, which is supposed to encode for a rapid NAT1 phenotype, was not significantly different between patients and controls [43% vs. 36%; odds ratio 1.5 (95% CI 0.88-2.68)]. Interactions between NAT2*4 and NAT1*10 were suggested by the increased frequency of the NAT2*4/NAT1*10 haplotype in patients (27%) compared with controls [15%; P = 0.039, odds ratio 2.1 (95% CI 1.04-4.04)]. As the NAT1 and NAT2 encoding genes are located in close proximity on chromosome 8p22, the latter finding could at least partly be due to genetic linkage. In fact, a linkage disequilibrium between NAT2*4 and NAT1*10 was observed in the contact allergy (P = 0.0025) and in the control group (P = 0.042). Our data indicate an association between the NAT2*4/NAT1*10 haplotype and contact sensitization to para-substituted aryl compounds. Therefore, acetylation may either enhance contact sensitization or NAT2*4 and NAT1*10 might be linked to an unknown susceptibility factor. PMID- 10848735 TI - Surveillance of occupational skin disease: EPIDERM and OPRA. AB - Consultant dermatologists in the U.K. have been reporting to EPIDERM, a voluntary surveillance scheme for occupational skin disease, since February 1993; reporting by occupational physicians to the scheme began in May 1994 and was superseded in January 1996 by OPRA (Occupational Physicians Reporting Activity). Currently 244 dermatologists and 790 occupational physicians report incident cases to these schemes. During the 6 years to January 1999 a total of 12, 574 new cases of occupational skin disease was estimated from reports by consultant dermatologists and 10,136 cases estimated from occupational physicians (since May 1994). The annual incidence of occupational contact dermatitis using data from both schemes was 12. 9 per 100,000 workers. The incidence of contact dermatitis per 100, 000 workers increased with age in men from 4.9 (age 16-29 years) to 6.6 (age 45-60 years); in women a higher rate (9.5) was apparent in the younger age group, with lower rates in older female workers. High rates in young workers were associated with wet work and in older workers with exposure to oils. For men, high rates of contact dermatitis were seen in reports from both schemes for chemical operatives, machine tool setters and operatives, coach and spray painters and metal workers. For women, high rates were found for hairdressers, biological scientists and laboratory workers, nurses and those working in catering. The most frequent agents for contact dermatitis were rubber chemicals and materials (14.1% of cases reported by dermatologists), soaps and cleaners (12.7%), nickel (11. 9%), wet work (11.1%), personal protective equipment (6.2%), petroleum products (6.3%), cutting oils and coolants (5.6%), and epoxy and other resins (6.1%). In the 1608 estimated cases of skin cancer all but 4% were attributed to ultraviolet radiation. Cases of contact urticaria attributed to latex peaked in 1996, with a decline in cases since that time. PMID- 10848737 TI - Role for tumour necrosis factor-alpha receptors in ultraviolet-induced skin tumours. AB - The biological effects of tumour necrosis factor (TNF)-alpha are mediated through either the TNFR1 or the TNFR2 receptor. In the present study, the effects of ultraviolet (UV) irradiation on skin pathology and tumour promotion were studied in hairless mice deficient in either the TNFR1 or the TNFR2 receptor. SKH-1 hairless mice were crossed with either TNFR1 knockout (KO) mice or TNFR2 KO mice to develop a strain of hairless mice deficient in either of these receptors. Elastosis and other pathological indications of UVB irradiation were not affected by the loss of either receptor. The absence of either receptor, however, resulted in a highly significant reduction in skin tumours in response to UVB irradiation. Inflammatory cell influx following chronic UV irradiation was virtually eliminated in the TNFR1 KO mice, while the TNFR2 KO mice responded to UV irradiation with the normal increase in inflammatory cells throughout the lower and upper dermis. Contact hypersensitivity responses were eliminated in the TNFR2 KO mice, whereas the TNFR1 KO mice retained normal contact hypersensitivity reactions. These studies suggest that TNF-alpha plays no part in the accumulation of excessive elastin in the skin during chronic UVB exposure. However, there appears to be an important role for TNF-alpha in mediating tumorigenesis, distinct from its role in initiating cutaneous immune responses. PMID- 10848736 TI - The use of two substrates to improve the sensitivity of indirect immunofluorescence in the diagnosis of pemphigus. AB - The aim of this study was to re-evaluate indirect immunofluorescence (IIF) comparing two substrates, normal human skin (HS) and monkey oesophagus (MO) using serum from 29 pemphigus patients classified according to the presence of serum autoantibodies to either desmoglein (Dsg) 1 or Dsg3 detected by enzyme-linked immunosorbent assay (ELISA). Overall, the sensitivity of IIF was 83% on HS and 90% on MO. When data from both substrates were combined, the sensitivity increased to 100%. When sera from pemphigus foliaceus (PF) patients were studied, which contained Dsg1 antibodies only, the sensitivity of IIF was greatest on HS and titres were on average 4.8 doubling dilutions higher than on MO. In contrast, when sera containing autoantibodies only to Dsg3 from pemphigus vulgaris (PV) patients was studied, the sensitivity was greatest on MO and titres were on average 4.4 doubling dilutions higher than on HS. There was a significant correlation between Dsg1 antibody levels and IIF titres on HS and between Dsg3 antibody levels and IIF titres on MO. The investigation of immunobullous disorders in the future is likely to move towards antigen-specific techniques such as the Dsg ELISAs used in this study. However, in laboratories which currently rely on IIF for detecting pemphigus autoantibodies, the data presented in this study strongly suggest that two substrates should be used for IIF screening: one rich in Dsg1, such as HS, and the other rich in Dsg3, such as MO. This combination of substrates should not only increase the sensitivity of detecting pemphigus antibodies, but will aid in the differentiation of PV from PF. It is also possible that the data might be more useful for disease monitoring. PMID- 10848738 TI - Mucosal human papillomavirus types in squamous cell carcinomas of the uterine cervix and subsequently on fingers. AB - Human papillomavirus (HPV), especially type 16, is causally involved in the pathogenesis of anogenital cancer. There is an increasing number of reports of HPV infections in squamous cell carcinoma (SCC) of the fingers. A search of the Swedish cancer register covering the period 1958-94 inclusive for women with a history of genital and upper extremity SCC revealed 63 cases. Archival material from both cervical and cutaneous lesions was traced and analysed for the presence of HPV DNA in 32 of these patients. A newly developed 'neighbour primer' polymerase chain reaction (PCR) for HPV 16 DNA, aimed at overcoming the obstacle of cross-linked target DNA, was shown to be superior to conventional general and type-specific HPV PCR tests. HPV DNA was significantly more frequently found in digital tumours than in tumours at other cutaneous sites of the upper extremities [67% (10 of 15) vs. 7% (three of 43); P < 0.001]. Among 13 patients with a history of both cervical and finger SCC, HPV 16 was found in cervical samples from seven patients. From five of these seven patients, HPV 16 was also present in the corresponding finger lesions. The results support the hypothesis of a possible transmission of patients' genital HPV infections to fingers. PMID- 10848739 TI - Prevalence of solar damage and actinic keratosis in a Merseyside population. AB - This study examines the prevalence of sun-related damage to the skin in a caucasian population in north-west England. The importance of constitutional factors (complexion, skin type and age) as well as environmental and occupational exposures for the development of actinic keratosis (AK) and skin cancers was assessed in people over 40 years of age attending outpatient clinics (non dermatology) at four centres in north-west England (Mersey region). Nine hundred and sixty-eight volunteers (531 men and 437 women) were recruited. The overall prevalence of AK was 15.4% in men and 5.9% in women. The prevalence was strongly related to age in both sexes, being 34.1% and 18.2%, respectively, in men and women aged 70 years and above, and was most strongly related to two objective signs of sun exposure, namely degree of solar elastosis and presence of solar lentigines. The prevalence of AK was higher in subjects with red hair and freckles, particularly women. There was no evidence of an increased prevalence of AK in relation to any occupation. There was a high prevalence of seborrhoeic keratosis and viral warts in both sexes, which was age-related in the case of seborrhoeic keratosis. Ten cases of basal cell carcinoma, eight cases of Bowen's disease and one case of malignant melanoma were identified. This study shows that the sun exposure received in 'normal' life in England is sufficient to cause potentially malignant skin damage in a significant proportion of the population. PMID- 10848740 TI - Calcipotriol ointment and cream or their vehicles applied immediately before irradiation inhibit ultraviolet B-induced erythema. AB - Results of ultraviolet (UV) B phototherapy can be improved by the application of calcipotriol, but studies are needed to decide how the two treatments should be combined. We studied the effect of UVB after application of calcipotriol ointment (50 microg g-1) and calcipotriol cream (50 microg g-1) and determined the optimal time of application of calcipotriol when combined with UVB phototherapy (280-350 nm), in a single-blinded randomized vehicle-controlled study of 37 healthy adult volunteers. Calcipotriol ointment or cream was applied randomly on five areas on the back at different time intervals from UVB irradiation. One area was left untreated as the control. Application times were the evening before, the morning before, 2 h before, immediately before, and immediately after irradiation. UVB irradiation was administered by TL20W/12 fluorescent tube lamps at increasing doses (20, 25, 32, 40, 50 and 64 mJ cm-2) to six subunits of each test area. Clinical assessment was performed 24 h after UVB irradiation by a blinded investigator. Calcipotriol ointment and cream were applied in 19 and 18 subjects, respectively, and erythema was measured for each application time quantified. We found that erythemal reactions were significantly smaller when calcipotriol ointment or cream was applied immediately before irradiation compared with all other application times. To explain these findings, a vehicle control study was performed. No difference in erythema was seen between calcipotriol medication and the vehicle controls. Spectrophotometric analysis of the calcipotriol cream and ointment showed no UV absorbance in the UVB range. No signs of photosensitization were noted. In conclusion, the vehicles of the calcipotriol ointment and cream inhibit the induction of erythema by UVB irradiation if applied immediately before phototherapy. Consequently, calcipotriol ointment and cream should not be applied directly before UVB irradiation; however, they may be applied at any time up to 2 h prior to or immediately after UVB irradiation. Possible explanations for this sunscreen activity are discussed. PMID- 10848741 TI - Cost evaluation of the medical management of neurofibromatosis 1: a prospective study on 201 patients. AB - Neurofibromatosis 1 (NF1) is associated with many internal complications as well as skin manifestations, and patients may require a variety of medical and surgical interventions. We aimed to assess the medical needs of NF1 patients, and to evaluate the financial cost of the resources used for them in relation to the severity of the disease. We conducted a prospective analysis on a cohort of 201 patients in our referral centre for adults. Severity of the disease was assessed. Therapeutic management was considered as multidisciplinary if it required more than three different specialists. Plastic and dermatological surgery procedures performed were recorded. Hospital costs were computed over a 3-year period and included all hospitalization days, clinic visits and procedures performed in all departments where the patients were admitted. One hundred and thirty-seven patients had at least one out-patient procedure or one hospitalization during the follow-up period. The mean cost per patient per year was pound810 (median 240; range 0-13, 860). Multidisciplinary procedures were more frequent in moderately and severely affected NF1 patients than in milder cases (P < 0. 0001); hence, the costs for moderate and severe cases were higher than for less severe groups (P = 0.005). Plastic and/or dermatological surgery was performed with the same frequency in the different severity groups (71%). Regardless of the presence of serious intractable complications, the patients' priority is for treatment of the disfigurement due to the disease. The management of these patients can be considered relatively inexpensive from the viewpoint of the healthcare system. PMID- 10848742 TI - Nailfold video capillaroscopy in psoriasis. AB - Changes in the microvasculature are considered to play an important part in the pathogenesis of psoriasis and its associated arthritis. The novel method of nailfold video capillaroscopy is an extension of the technique of widefield nailfold microscopy which has been of diagnostic and predictive use in the in vivo study of the microcirculation in systemic sclerosis and other connective tissue disorders. However, similar studies in patients with psoriasis and psoriatic arthritis and/or nail changes have produced conflicting results. We tested the hypothesis that any abnormalities in nailfold capillaries of either a quantitative or qualitative nature might be observed more readily in subjects with pathology adjacent to the nailfold, i.e. distal interphalangeal (DIP) joint changes and/or nail dystrophy, when using this technique. Forty-four patients with psoriasis were recruited (21 males, 23 females). Twelve patients had psoriasis alone, 13 had psoriasis and nail changes, six had DIP joint involvement with changes of psoriasis elsewhere, and 13 had psoriasis, DIP arthritis and nail changes. Capillary density and standard capillary dimensions were studied and compared with those of 44 age- and sex-matched control subjects. There was a significant (P < 0.05) decrease in capillary loop density in patients with either psoriasis plus nail disease (14.5 +/- 5.7 capillaries per 3 mm field) or psoriasis plus nail and DIP joint disease (14.3 +/- 5. 0) when compared with controls (19.2 +/- 3.8). In patients with psoriatic arthritis affecting the DIP joints, there was a statistically significant (P < 0.05) decrease in arterial and venous capillary limb diameters, and this was also seen in those with arthritis associated with nail changes. However, there was no difference in capillary dimensions between patients with psoriasis and/or nail changes when compared with normal controls. Morphological abnormalities previously described in the literature were not noted in any of our four patient groups. Our findings of diminution in both nailfold capillary bed density and dimensions of the arterial and venous capillary limbs suggest that vascular injury, previously noted in ultrastructural studies, may play a part in the pathogenesis of psoriatic arthritis. However, in contrast to previous studies, we found no specific pattern of a morphological nature of nailfold capillaries in patients with psoriasis with or without nail changes, when compared with normal controls. PMID- 10848743 TI - A randomized trial of amorolfine 5% solution nail lacquer combined with oral terbinafine compared with terbinafine alone in the treatment of dermatophytic toenail onychomycoses affecting the matrix region. AB - In view of recent advances in the development of antifungal agents, this study examined the possible synergy of two new antifungal agents, terbinafine and amorolfine. The study compared two different courses of terbinafine treatment combined with amorolfine 5% solution nail lacquer. Terbinafine was given orally for 6 (AT6 group) or 12 weeks (AT12 group) and amorolfine nail lacquer applied weekly for 15 months. A control group received terbinafine alone for 12 weeks. This was a randomized, prospective, open study of severe dermatophyte toenail onychomycosis with matrix region involvement. Nail samples were taken before the start of the study, at inclusion and at the visits at 6 weeks, 3, 9, 15 and 18 months. To assess the value of such combined therapy we chose an early parameter as the principal outcome variable, which was the result of mycological examination, including direct microscopy and culture, at 3 months (allowing a margin of 15 days). The secondary parameters of success were the mycological results at the later visits, clinical evaluation and a combined mycological clinical response. Safety and tolerance were also assessed. Adverse events were recorded and liver function tests were performed monthly during the terbinafine treatment. Of the 147 patients included in the trial, 121 attended the 3-month visit, within a time limit of 15 days of 3 months after the beginning of treatment: 40 in the AT6 group, 40 in the AT12 group and 41 in the control group. In all, 32 of 121 patients (26. 4%) had negative mycological results on direct microscopy and culture: 14 of 40 (35.0%) in the AT6 group, 11 of 40 (27.5%) in the AT12 group and seven of 41 (17.1%) in the control group. The cure rate for the global (mycological and clinical cure) response measured at 18 months in 145 patients was 44.0% (22 patients) in the AT6 group, 72.3% (34 patients) in the AT12 group and 37.5% (18 patients) in the terbinafine group. These results suggest that the combination of amorolfine and terbinafine may be of value in the treatment of severe onychomycosis. At the same time a pilot pharmacoeconomic analysis was performed demonstrating a better cost per cure ratio for the patients receiving combination treatment. PMID- 10848744 TI - Regressing Merkel cell carcinoma-a case showing replacement of tumour cells by foamy cells. AB - Merkel cell carcinoma (MCC) is a malignant neuroendocrine tumour with a high rate of recurrence and metastasis. However, some cases of spontaneous regression have recently been reported. We describe the clinical course of an 80-year-old Japanese woman with regressing MCC. We also report histological findings of the regressing tumour for the first time. After the patient's first visit to our hospital, the lesion was a rapidly progressive tumour, but suddenly began decreasing in size, and rapidly regressed. The surface of the tumour flattened, the colour changed from red to dark red, and finally the lesion appeared as a small yellowish plaque. Histopathological analysis of the completely regressed tumour revealed that the tumour cells were completely replaced by numerous foamy cells. This is the first report demonstrating the histopathological features of regressing MCC. PMID- 10848745 TI - Two cases of vulval pigmented extramammary Paget's disease: histochemical and immunohistochemical studies. AB - We describe two Japanese female patients with pigmented extramammary Paget's disease (EMPD); one patient had a dark brown plaque and the other had a reddish patch with a pigmented area, both affecting the vulval region. Histochemical and immunohistochemical examinations confirmed EMPD with melanocyte colonization; plump tumour cells with a large nucleus and pale cytoplasm that were positive for CAM 5.2 and CEA proliferated singly or in nests in the epidermis, and scattered among the tumour cells were many dendritic cells with a large amount of melanin that were positive for HMB-45 and S-100 protein. Fontana-Masson (FM) stain showed many positive cells with well-developed dendritic processes within and around tumour nests. Histochemical and immunohistochemical studies of non-pigmented EMPD cases on the same region showed that HMB-45 positive cells were sparse or not detected at all, and that also FM staining-positive cells were decreased or not detected, and their dendritic processes were poorly formed. The present study suggests that there might be heterogeneity in EMPD in terms of relationships between Paget's cells and melanocytes. PMID- 10848746 TI - Treatment of verrucous carcinoma of vulva with acitretin. AB - We describe a 60-year-old patient with verrucous carcinoma of the vulva, which recurred 6 years after simple vulvectomy and radiotherapy. Treatment with acitretin led to significant improvement and ongoing disease control has been achieved with low-dose maintenance therapy of 10 mg acitretin daily. PMID- 10848747 TI - Widespread cutaneous necrosis occurring in association with the antiphospholipid syndrome: a report of two cases. AB - The antiphospholipid syndrome is an acquired prothrombotic state where thrombosis and/or pregnancy loss is related to the presence of antiphospholipid antibodies. Cutaneous necrosis secondary to intravascular thrombosis of small dermal vessels is a recognized but rare association with antiphospholipid syndrome. We report two patients with high circulating levels of anticardiolipin antibodies who developed widespread cutaneous necrosis as the first clinical manifestation of the antiphospholipid syndrome. The exact mechanism by which antiphospholipid antibodies mediate thrombosis is uncertain; however, proposed mechanisms of activity include endothelial cell activation, altered endothelial production of prostacyclin, activation of platelets, and modulation of the protein C and S pathways. PMID- 10848748 TI - Depigmented hypertrichosis following Blaschko's lines associated with cerebral and ocular malformations: a new neurocutaneous, autosomal lethal gene syndrome from the group of epidermal naevus syndromes? AB - The lines of Blaschko represent one of the cutaneous patterns of mosaicism followed by various skin disorders. Developmental abnormalities affecting other tissues derived from the embryonic ectoderm and mesoderm are occasionally associated. We describe a 30-year-old man with depigmented, bilateral hypertrichosis and dilated follicular orifices following Blaschko's lines associated with cerebral and ocular malformations. The findings suggest a previously unreported neurocutaneous, autosomal lethal gene syndrome from the group of epidermal naevus syndromes. PMID- 10848749 TI - Linear eruptions of the nose in childhood: a form of lichen striatus? AB - We report four children with linear eruptions on the nose, with overlapping features of lichen striatus and linear cutaneous lupus erythematosus. However, linear lupus erythematosus has rarely been reported, and lichen striatus, although classically linear, rarely affects the face. The linear distribution of lesions from the glabella to the ala nasi may represent distribution following Blaschko's lines. PMID- 10848750 TI - Linear IgA disease with IgA antibodies directed against 200- and 280-kDa epidermal antigens. AB - We report an 80-year-old man with the lamina lucida type of linear IgA disease, with IgA autoantibodies reactive with 200-kDa and 280-kDa epidermal proteins. The patient presented with widespread bullous lesions on his trunk and extremities without mucosal involvement. Histopathology of lesional skin showed a subepidermal blister with papillary microabscesses of neutrophils and a few eosinophils. Direct immunofluorescence microscopy of perilesional skin showed linear deposits of IgA and C3 at the basement membrane zone. The patient's serum contained IgA autoantibodies that bound exclusively to the epidermal side of 1 mol L-1 NaCl split skin as determined by indirect immunofluorescence microscopy. Circulating IgA autoantibodies to 200- and 280-kDa antigens were detected in the patient's serum by immunoblot analysis using extracts from normal human epidermis and human epidermal keratinocytes. These two antibodies, eluted from individual nitrocellulose membranes, reacted with the epidermal side of 1 mol L-1 NaCl split skin on indirect immunofluorescence microscopy, and bound to hemidesmosomes as determined by immunoperoxidase electron microscopy. This observation suggests the presence of hitherto uncharacterized 200- and 280-kDa hemidesmosomal proteins distinct from BPAG1, BPAG2 and beta4 integrin as target antigens in linear IgA disease. PMID- 10848751 TI - Red facial pseudochromhidrosis. AB - We describe a 9-year-old girl with pseudochromhidrosis simulating apocrine chromhidrosis. Treatment with topical and systemic erythromycin resulted in complete clearance of the reddish discoloration of the face. No relapse or recurrence was observed over a 3-month period. PMID- 10848752 TI - Photosensitive mycosis fungoides or actinic reticuloid? AB - We report two patients who satisfied the diagnostic criteria for actinic reticuloid (AR) on initial presentation, in whom genotypic analysis of early skin biopsies failed to show T-cell gene receptor rearrangements. Both patients progressed to widespread skin involvement associated with histopathological and genotypic features of mycosis fungoides (MF). Arguably, these patients may have had photosensitive MF from the outset, but their clinical features, phototesting, and subsequent demonstration of a T-cell gene receptor rearrangement in the skin could also suggest progression of AR to MF. PMID- 10848753 TI - Pseudoscleroderma associated with lung cancer: correlation of collagen type I and connective tissue growth factor gene expression. AB - Pseudoscleroderma as a paraneoplastic syndrome is a rare disease. We report here a patient with lung cancer (undifferentiated squamous cell carcinoma), who developed acrosclerosis. Using in situ hybridization, marked expression of alpha1(I)-collagen and connective tissue growth factor (CTGF) mRNA was found in fibroblasts scattered throughout the dermis. However, transforming growth factor (TGF)-beta1 expression was not detected. The pattern of CTGF gene expression and collagen synthesis was similar to that in systemic scleroderma. The absence of TGF-beta1 mRNA could indicate that tumour-derived factors induce the expression of CTGF. PMID- 10848754 TI - Langerhans cell histiocytosis in a child presenting as a pustular eruption. PMID- 10848755 TI - Normolipaemic plane xanthomatosis associated with mycosis fungoides. PMID- 10848756 TI - The prevalence of human T-cell lymphotropic virus type I infection in patients with cutaneous malignancies. PMID- 10848757 TI - Spindle cell hemangioendothelioma: successful treatment with recombinant interleukin-2. PMID- 10848758 TI - Subungual angioleiomyoma masquerading as a glomus tumour. PMID- 10848759 TI - Calcification in the apocrine glands of naevus sebaceus. PMID- 10848760 TI - Syringocystadenoma papilliferum presenting as a cutaneous horn. PMID- 10848761 TI - Normal anagen effluvium: a sign of pemphigus vulgaris. PMID- 10848762 TI - Improvement of folliculitis decalvans following shaving of the scalp. PMID- 10848763 TI - Rheumatoid neutrophilic dermatitis associated with pyoderma gangrenosum. PMID- 10848764 TI - Recurrent erythema multiforme and chronic hepatitis C: efficacy of interferon alpha. PMID- 10848765 TI - Should patients with pruritus be tested for hepatitis C virus infection? A case controlled study. PMID- 10848766 TI - Pityriasis rosea, HHV-7 and multiple sclerosis. A coincidence? PMID- 10848767 TI - Confluent and reticulate papillomatosis: successful treatment with azithromycin. PMID- 10848768 TI - Staphyloccocal scalded skin syndrome in healthy adults. PMID- 10848769 TI - Amlodipine induced-photosensitivity presenting as telangiectasia. PMID- 10848770 TI - Strawberry glans penis: a rare manifestation of angiokeratomas involving the glans penis. PMID- 10848771 TI - Grover's disease secondary to ribavirin. PMID- 10848772 TI - Granulomatous nodule on vocal cord possibly induced by etretinate therapy. PMID- 10848773 TI - Fexofenadine-induced QT prolongation: a myth or fact? PMID- 10848774 TI - Cutaneous alternariosis by Alternaria chartarum in a renal transplanted patient. PMID- 10848775 TI - Bilateral congenital adipose plantar nodules. PMID- 10848776 TI - An appetite for apoptotic cells? Controversies and challenges. PMID- 10848777 TI - Topoisomerase II inhibitor-related acute myeloid leukaemia. PMID- 10848778 TI - Prognostic value of serum markers of bone metabolism in untreated multiple myeloma patients. AB - Bone involvement is a central feature of multiple myeloma (MM). We investigated whether serum markers of osteoblastic and osteoclastic activity correlate with the presence of bone disease and survival in 313 MM patients enrolled in a phase III trial (E9486). Five markers were measured, including osteocalcin (OC), carboxy-terminal propeptide of type I collagen (PICP), bone alkaline phosphatase (BAP), carboxy-terminal telopeptide of type I collagen (ICTP) and tartrate resistant acid phosphatase (TRAP). We analysed the relationship between serum levels of these markers and the presence of bone manifestations, and survival. Serum levels of ICTP and BAP correlated significantly with bone pain, lesions and fractures. Serum level of ICTP was also higher in stage II-III compared with stage I disease. The serum level of ICTP was significantly associated with shortened survival in the univariate analysis. The median survival times were 4.1 and 3.5 years for low and high ICTP respectively (P = 0.02). There was a strong relationship between ICTP and beta-2-micrgolobulin (B2M). ICTP stands out as a significant marker of bone disease. Incorporation of these markers into clinical trials assessing the use of bisphosphonates in MM is needed to determine whether they might serve as indicators of effectiveness of these agents. PMID- 10848779 TI - Amplification of cyclin D1 gene in multiple myeloma: clinical and prognostic relevance. AB - Approximately 30% of myeloma patients express cyclin D1 RNA and protein. The low incidence of translocation t(11; 14) detected by conventional cytogenetics suggests that the up-regulation of cyclin D1 protein might result from other mechanisms as well as from gene amplification. Therefore, the frequency and the clinical and prognostic implications of cyclin D1 amplification were examined. We highly purified myeloma cells from bone marrow by magnetic cell sorting and analysed 50 myelomas by fluorescence in situ hybridization (FISH) using probes specific for cyclin D1 and 20 samples by immunoblotting to detect cyclin D1 expression. The amplification of cyclin D1 gene was found in 19 of 50 analysed patients and was associated with expression of cyclin D1 protein. The amplification correlated significantly with the bone marrow infiltration, plasma cell morphology and labelling index as well as serum beta2-microglobulin, C reactive protein (CRP) and creatinine levels. In univariate analysis, the amplification of the cyclin D1 gene was a significantly unfavourable parameter with regard to overall survival (P = 0.0064) and progression-free survival (P = 0. 0005). In multivariate analysis, cyclin D1 amplification and serum beta2 microglobulin were independent and well-suited parameters for predicting survival. The detection of cyclin D1 amplification seems to be of promising prognostic value in multiple myeloma. PMID- 10848780 TI - Interleukin 6, tumour necrosis factor alpha, interleukin 1beta and interleukin 1 receptor antagonist promoter or coding gene polymorphisms in multiple myeloma. AB - Proinflammatory cytokines such as interleukin 6 (IL-6), tumour necrosis factor alpha (TNF-alpha) and IL-1beta are considered to be involved in the pathogenesis of multiple myeloma (MM). In the present study, we examined a G/C polymorphism at position -174 in the promoter region of IL-6, a biallelic polymorphism at position -308 in the promoter region of TNF-alpha, the TaqI restriction fragment length polymorphism in exon 5 of IL-1beta and a variable number of identical tandem repeat polymorphisms in intron 2 of IL-1 receptor antagonist (IL-1Ra) genes. The alleles of these loci are known to influence the level of production of the cytokines and the IL-1Ra. Seventy-three patients with MM, 27 with monoclonal gammopathy of undetermined significance (MGUS) and 129 healthy individuals were included. No difference was found between patients and healthy controls or between MM and MGUS patients in the distributions of genotypes and frequencies of alleles of the IL-6 (-174), TNF-alpha (-308), IL-1beta TaqI and IL 1Ra gene polymorphisms. No associations between the polymorphisms at the loci under study and clinical factors such as age, sex, clinical stage at onset and M protein type were observed. Our results indicate that the cytokine (IL-6, TNF alpha and IL-1beta) and IL-Ra gene polymorphisms do not confer susceptibility to the development of MM. PMID- 10848781 TI - Increased expression of non-functional killer inhibitory receptor CD94 in CD8+ cells of myeloma patients. AB - Different MHC class I-specific killer inhibitory receptors (KIRs) are expressed in vivo by a minor fraction of activated memory CD8+ cells. It has been postulated that KIRs may 'fine-tune' specific responses by altering their threshold of activation by the TCR-CD3 complex. We have previously shown that, in multiple myeloma (MM) patients, a large fraction of peripheral blood CD8+ cells display the phenotype of chronically activated memory T cells (CD38+, HLA-DR+, CD25-, CD45R0+, CD28-). We investigated the expression of KIRs on MM T cells and determined their possible influence on cytolytic responses elicited via the CD3 TCR complex. The expression of CD94, a molecule that is part of a heterodimeric KIR recognizing the non-classical MHC surface HLA-E molecule, was almost threefold higher in MM T cells than in age-matched normal control subjects (P < 0.0001). CD94 expression was preferentially confined to CD8+ cells but not restricted to activated (HLA-DR+) and/or memory (CD45R0+) T cells. Unlike normal T cells, in which CD94 is assembled with glycoproteins of the NKG2 family to form functional receptors with activating or inhibitory properties, most CD94+ MM T cells were devoid of both the NKG2-A and NKG2-C glycoproteins detected in the inhibitory or activating form respectively. CD94 blockade did not significantly affect either T-cell proliferation or cytotoxic T-lymphocyte generation induced by the myeloma-derived cell lines NCI and RPMI 8226. Similarly, the cytolytic activity induced by direct anti-CD3-mediated targeting of MM T cells to FCR+ P815 target cells was unaffected by the addition of anti-CD94 and/or anti-NKG2-A/C monoclonal antibodies (mAbs). These data indicate that the large majority of MM CD8+ cells do not express a functional CD94 receptor. Thus, their ability to 'fine-tune' an appropriate immune response against tumour cells can be impaired. PMID- 10848782 TI - Human bone marrow stroma-dependent cell line MOLP-5 derived from a patient in leukaemic phase of multiple myeloma. AB - The novel multiple myeloma (MM) cell line MOLP-5 and its homologous sister cell line B407, a lymphoblastoid cell line (LCL), were established from the peripheral blood of a 71-year-old Japanese patient with Bence-Jones kappa-type multiple myeloma (stage IIIB with hyperammonaemia and hypercalcaemia). The growth of MOLP 5 cells is constitutively dependent on bone marrow stroma (BST) cells; none of the cytokines tested nor the culture supernatant of the bone marrow stroma cells could support the growth of MOLP-5. Wright-Giemsa-stained MOLP-5 cells showed typical plasma cell morphology with abundant cytoplasm and one to three nuclei. The immunoprofile of MOLP-5 corresponds to that seen typically in primary MM cells: positive for cytoplasmic immunoglobulin (Ig) kappa light chain, CD28, CD29, CD38, CD40, CD44, CD49d, CD54, CD56, CD58, CD71, CD138 and PCA-1; the cells were negative for surface Ig and various other B-cell, T-cell and myelomonocyte associated immunomarkers. Interleukin 6 (IL-6) receptor mRNA was found in the reverse transcriptase polymerase chain reaction (RT-PCR) analysis. IL-6 and IL-10 could induce cellular proliferation in short-term induction experiments. IL-6 or IL-10 production was not detected by specific enzyme-linked immunoabsorbent assay (ELISA). MOLP-5 cells expressed parathyroid hormone-related protein (PTHrP) at the mRNA level. Cytogenetic analysis showed the typical t(11; 14) chromosome abnormality. The novel MOLP-5 cell line together with the B407 B-LCL sister line will be useful model systems in the investigation of the biology of MM. PMID- 10848783 TI - Lack of BLV and PTLV DNA sequences in the majority of patients with large granular lymphocyte leukaemia. AB - The primate T-cell lymphoma/leukaemia viruses (PTLV) and bovine leukaemia virus (BLV) comprise a unique genus of retroviruses, infection with which induces seroreactivity in the host against conserved epitopes in their p24 gag and gp21 env cognate proteins. Herein, we have confirmed this serocrossreactivity. Patients with large granular lymphocyte (LGL) leukaemia have frequent seroreactivity to the p24 and gp21 env proteins of human T-cell lymphoma/leukaemia virus I (HTLV-I), one of the species in the genus. However, only a small minority of patients are actually infected with prototypic HTLV-I or HTLV-II, another species within the group. In an attempt to determine whether LGL leukaemia might be associated with other members of the PTLV/BLV genus, we examined the peripheral blood mononuclear cell DNA of 22 HTLV p24 and/or gp21 seropositive LGL leukaemia patients via PCR using degenerate and specific primer pair/probe systems capable of detecting all known members of the PTLV/BLV genus. None of the samples was positive. These data indicate that although HTLV-II may be associated with some cases of LGL leukaemia most patients are not infected with a PTLV or BLV virus. PMID- 10848784 TI - IgG-secreting lymphoplasmacytoid leukaemia: a B-cell disorder with extensively mutated VH genes undergoing Ig isotype-switching frequently associated with trisomy 12. AB - We investigated 16 patients with elevated serum monoclonal IgG and a leukaemic B cell lymphocytic disorder different from multiple myeloma. Their clinical history was that of a non-aggressive disease with dominant splenomegaly and long survival. Whereas abnormal blood and bone marrow cells were predominantly small lymphocytes with a few lymphoplasmacytoid cells, histopathological features included a lymphoplasmacytic infiltrate in eight cases. Most frequently, abnormal blood cells displayed a CD19+CD5-CD23+/- immunophenotype different from that of chronic lymphocytic leukaemia, except in two cases with a CD19+CD5+CD23+ phenotype. Interestingly, a coexistent serum monoclonal IgM and/or surface IgMG+ with identical light chain was identified in 10 patients, whereas in the remaining six patients only IgG expression was determined. VH gene analysis was performed in eight patients to investigate the clonal origins of tumour cells. All cases utilized the VH3 family, with evidence of extensive somatic mutations and intraclonal homogeneity in all cases. VH gene analysis indicated a clonal relationship between cells expressing IgM and IgG, with one case being biclonal. Cytogenetic evaluation showed a high incidence of trisomy 12 (60%) and 13q14 deletion (40%). In conclusion, we have described an unusual subset of low-grade lymphoma with high-serum IgG and frequent lymphoplasmacytoid features in which tumour cells derive from post-follicular memory B cells undergoing isotype switching with some cases arrested at both the IgM and IgG stage and others as IgG-positive cells only. PMID- 10848785 TI - A UK multicentre phase II study of rituximab (chimaeric anti-CD20 monoclonal antibody) in patients with follicular lymphoma, with PCR monitoring of molecular response. AB - Follicular lymphoma (FL) cells express CD20 and are associated in most cases with the t(14;18) chromosomal translocation. A multicentre study was undertaken between January 1997 and January 1998 to assess the complete response rate (CR) and overall response rate (RR) to rituximab, a chimaeric anti-CD20 monoclonal antibody. Seventy patients with previously treated FL received rituximab (375 mg/m2/week x4, by intravenous infusion). Restaging studies were performed 1 and 2 months after therapy. Molecular monitoring for the presence of cells harbouring the Bcl-2/JH gene rearrangement in the peripheral blood (PB) and bone marrow (BM) was performed before and after treatment using a two-step semi-nested polymerase chain reaction (PCR) assay. The overall RR was 32/70 (46%), being highest in patients who had received only one previous treatment (12/15, 80%). However, only two patients achieved a CR. The median duration of response was 11 months. Thirteen of 21 evaluable 'PCR-positive' patients (62%) became 'PCR-negative' in PB and/or BM samples 1 month after rituximab, although this did not correlate with clinical response. Treatment was generally well tolerated, although one patient developed Stevens-Johnson syndrome. Rituximab was shown to be active in FL, and in some cases PB and/or BM became PCR negative. Studies in combination with cytotoxic chemotherapy to increase the CR rate are warranted. PMID- 10848786 TI - Frequent good partial remissions from thalidomide including best response ever in patients with advanced refractory and relapsed myeloma. AB - Twenty-three patients with advanced and heavily pretreated myeloma were treated with thalidomide. Starting dose was 200 mg/d, and 20 patients had dose escalations up to 400 (n = 5), 600 (n = 12) or 800 mg/d (n = 3), usually in divided doses. Nineteen patients were refractory to recent chemotherapy, and four had untreated relapse after prior intensive therapy. Ten out of 23 patients (43%) achieved partial response (PR; nine with refractory and one with relapsed disease), six patients had minor response or stabilization of the disease and four had disease progression. Another three patients died early from advanced myeloma at less than 3 weeks of thalidomide therapy. Of the 10 patients with PR, seven had a better response than after any prior therapy, despite vincristine doxorubicin-dexamethasone (VAD)-based treatment in all but one and high-dose melphalan with autologous stem cell support in four. Time to achieve PR was rapid in patients receiving thalidomide in divided doses (median 31 d). Responses also included reduced bone marrow plasma cell infiltration and improved general status. Normalized polyclonal gammaglobulin levels were seen in four cases. Six out of 10 patients with PR remained in remission with a median time on treatment of 23 weeks (range 15-50 weeks). Sedation was common but usually tolerable, and some patients continued full- or part-time work. Four patients had skin problems, three patients had pneumonia, one hypothyrosis, one sinus bradycardia and one minor sensory neuropathy. Thalidomide may induce good partial remissions in advanced refractory myeloma with tolerable toxicity, and should be evaluated in other settings for myeloma patients. Divided thalidomide doses seem to reduce time to achieve remission and may improve response rate. PMID- 10848787 TI - Hypermethylation of P16ink4a and P15ink4b genes as a marker of disease in the follow-up of non-Hodgkin's lymphomas. AB - The hypermethylation of p16ink4a and p15ink4b genes have been described as an inactivating mechanism alternative to deletions and mutations that accounts for a relatively high proportion of cancers, including non-Hodgkin's lymphomas (NHLs). To investigate whether detection of abnormal methylation could have clinical applications in the management and follow-up of lymphomas, we have analysed the behaviour and evolution of p16ink4a and p15ink4b methylation in 13 NHL cases undergoing chemotherapy. All cases were also analysed for the presence of monoclonal rearrangements of immunoglobulin or T-cell receptor genes. Six patients showed methylation in at least one of these genes at diagnosis, whereas in two other cases methylation appeared during the treatment. The other five cases were always unmethylated. Methylation was detected when any histological or molecular evidence of disease was present, suggesting a good correlation between methylation and disease. In some cases, we were able to detect methylation in patients at complete remission and without evidence of monoclonal cell population, indicating a high sensitivity of the PCR to detect methylation. These results suggest that p16ink4a and p15ink4b methylation could be good markers of disease and could be helpful in identifying lymphoma patients at risk of relapse. PMID- 10848788 TI - Persistent mediastinal mass is not indicative of recurrence after chemotherapy only in paediatric Hodgkin's disease. AB - Most patients with Hodgkin's disease are treated with chemotherapy in conjunction with radiotherapy, but at the end of treatment a residual mass is often present. After combined therapy, it has been assumed that no additional treatment is needed. However, for children treated without radiotherapy, no data exist on the relevance of a residual mediastinal mass to risk of relapse. We report on the findings of follow-up thorax radiographs of a group of 27 children with initial mediastinal involvement, who were treated with chemotherapy only. We conclude that the regression rate of the mediastinal mass was not related to a later recurrence. Regression after chemotherapy without radiotherapy is probably slower than after combined therapy. We consider chest radiograph examinations to be appropriate for the follow-up of tumour regression. When the data were compared with a group of children with Hodgkin's disease without mediastinal involvement, we found that survival was not related to initial mediastinal involvement. PMID- 10848789 TI - Role of BCL-2 and cell cycle regulatory proteins for corticosensitivity assessment in childhood acute lymphoblastic leukaemia. AB - Results of treatment in childhood acute lymphoblastic leukaemia (ALL) remain unsatisfactory because relapses occur even after high-dose chemotherapy. Corticosensitivity is used in numerous therapeutic trials as a prognostic factor for treatment choice. The aim of this study was to evaluate the role of cell cycle regulatory protein expression before and during the first 48 h of corticotherapy for predicting corticosensitivity. Fifty-two children presenting with ALL were studied at diagnosis and during the first 48 h of treatment for cell proliferation and apoptosis level by measurement of DNA content, and for expression of several cell proliferation regulatory proteins by means of Western blot. Glucocorticoids induced a significant decrease in the percentage of cells in S-phase and in CDK1, CDK4 and CDK6 expression and an increase in the percentage of cells in subG1 peak. Two criteria for corticosensitivity were used: (i) the number of blast cells after 7 d of treatment with a threshold at 1 x 109/l (usual criterion), (ii) the J8/J1 blast cell ratio, which is independent from initial leucocytosis. Bcl-2 expression at diagnosis was the best predictive variable for the usual corticosensitivity criterion in B- and T-cell ALL. For the second criterion, in B-cell ALL, p21waf1 expression at diagnosis was the sole (albeit poorly) predictive variable, whereas bcl-2 remained of high interest in T cell ALL. Interestingly, these proteins, bcl-2 and p21waf1, are associated with prolonged cell lifespan and their increased expression is often linked to poor response to cytotoxic drugs. Such preliminary results call for subsequent studies on large independent sets of T-cell and B-cell lineage ALL in order to confirm the J8/J1 blast cell ratio value as well as the role of bcl-2 and p21waf1 expression in predicting corticosensitivity. PMID- 10848790 TI - No evidence for a major role of heterozygous deletion 657del5 within the NBS1 gene in the pathogenesis of non-Hodgkin's lymphoma of childhood and adolescence. AB - Nijmegen breakage syndrome (NBS) is an autosomal recessive DNA repair disorder with a high predisposition for lymphoid malignancies. The majority of NBS patients carry a homozygous founder mutation (657del5) within the NBS1 gene. The observation of a high incidence of cancer in close relatives of NBS patients suggests a potential pathogenetic role of NBS1 mutations in heterozygotes as well. We assessed the frequency of the 657del5 mutation in 109 paediatric patients with non-Hodgkin's lymphoma (NHL). None of the patients analysed carried a NBS1 allele with the 657del5 mutation, suggesting that this mutation does not play a major role in the pathogenesis of NHL of childhood and adolescence. PMID- 10848791 TI - Haematopoietic cell transplantation in patients with Fanconi anaemia using alternate donors: results of a total body irradiation dose escalation trial. AB - Allogeneic haematopoietic cell transplantation (HCT) is the only therapeutic modality capable of correcting the haematologic manifestations of Fanconi anaemia (FA). However, HCT from alternative donors has been associated with poor survival. Between June 1993 and July 1998, 29 FA patients (median age 12.1 years; range 3.7-48.5 years) were enrolled in a prospective phase I-II dose escalation study. All patients were treated with cyclophosphamide 40 mg/kg, total body irradiation (TBI) 450 cGy or 600 cGy and antithymocyte globulin (ATG), followed by HCT from an alternative donor. Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporin A for 6 months, short course methylprednisolone (2 mg/kg/day) between days +5 and +19 and marrow T-cell depletion by counterflow elutriation. The probability of developing grade III-IV toxicity was 17% (95% CI 3-31%). For the 25 marrow recipients, the probability of neutrophil engraftment (ANC 0.5 x 109/l by day 45) was 63% (95% CI 42-82%). Probabilities of grade II-IV acute GVHD and chronic GVHD were 32% (95%CI 10-54%) and 0% respectively. With a median follow-up of 18 months, the probability of survival for the entire cohort at 1 year was 34% (95% CI 17-51%). The presence of lymphocyte somatic mosaicism [i.e. the presence of diepoxybutane (DEB)-insensitive cells] was associated with a significantly increased risk of graft failure. Disappointingly, the use of higher dose TBI and post-transplant ATG did not improve engraftment. More effective peritransplant immunosuppression, especially in FA patients with somatic mosaicism, was required to overcome the barrier of graft rejection. New conditioning regimens adapted to each individual's alkylator sensitivity are needed to improve the outcome of alternative donor HCT for FA. PMID- 10848792 TI - Relapse after bone marrow transplantation: evidence for distinct immunological mechanisms between adult and paediatric populations. AB - Donor lymphocyte infusions are particularly effective for remission induction in malignant cells in patients who relapse after allogeneic progenitor cell transplantation (PCT) and who remain sensitive to the administration of unprimed donor T and/or natural killer (NK) cells present in donor lymphocyte infusions. To determine whether relapse after unmanipulated PCT could be ascribed to donor T and/or NK cell loss or tolerization, we evaluated the chimeric status of 81 patients with haematological malignancies who were receiving allogeneic unmanipulated PCT. The incidence of mixed chimaerism (MC) in unfractionated mononuclear leucocyte samples decreased rapidly after transplant, and was not detectable 4 months after PCT, even in patients who subsequently relapsed. The chimeric status of immune effector cell subsets was then evaluated in 15 patients at the time of relapse. All adults demonstrated complete donor haematopoiesis (CDH) for all cell lineages, whereas T- and NK-cell MC was only found in patients younger than age 13 years (P = 0.004). MC was not found in T nor NK cells of a control group consisting of age-matched paediatric patients in remission after allogeneic PCT. Thus, in adults, T and NK cell MC disappears early after unmanipulated allogeneic PCT and is absent at the time of relapse. However, the identification of donor T and NK cell loss in the paediatric relapsed but not remission patients suggests a distinct mechanism of relapse. PMID- 10848793 TI - Conditioning regimens in autologous stem cell transplantation for multiple myeloma: a comparative study of efficacy and toxicity from the Spanish Registry for Transplantation in Multiple Myeloma. AB - High-dose chemoradiotherapy conditioning regimens for autologous stem cell transplantation (ASCT) are generally held to give similar results in multiple myeloma (MM), but no specific comparative study has been published. We addressed this issue by comparing the main high-dose chemoradiotherapy regimens used in the Spanish Registry. Patient cohorts included 315 cases treated with 200 mg/m2 melphalan (MEL200), 127 patients with 140 mg/m2 melphalan plus total body irradiation (MEL140 + TBI) and 121 cases with 12 mg/kg busulphan plus 140 mg/m2 melphalan (BUMEL). After ASCT, granulocyte and platelet recovery time was similar in all conditioning groups. There were no differences in transplant-related mortality. All regimens yielded a similar response in reference to pre-ASCT MM status, although BUMEL produced a slightly better overall response when compared with the other regimens (97% vs. 89% and 92%, P = 0.003). The 5-year overall survival (OS) with BUMEL was 47% [95% confidence interval (CI) 26-68] compared with 43% (CI 31-54) for MEL140 + TBI and 37% (CI: 18-56) for MEL200. The median survival for the BUMEL group was 64 months compared with 45 and 37 months for the MEL200 and MEL140 + TBI groups respectively. These differences were non significant (P = 0.2). The median event-free survival (EFS) was better for BUMEL (32 months) than for MEL200 (22 months) or for MEL140 + TBI (20 months). The differences in EFS between BUMEL and the other conditioning regimens reached statistical significance (P = 0.01). Nevertheless, the adjusted multivariate analysis for OS and EFS revealed that the conditioning regimens had no independent prognostic value. We concluded that three different conditioning regimens, commonly used for ASCT in MM, have a similar antimyeloma effect. However, the trend for better results observed in our series with BUMEL requires a prospective trial. PMID- 10848794 TI - Summary curves for patients transplanted for chronic myeloid leukaemia salvaged by a donor lymphocyte infusion: the current leukaemia-free survival curve. AB - A significant number of patients who relapse after allogeneic stem cell transplantation (SCT) for chronic myeloid leukaemia (CML) will achieve sustained molecular remissions after treatment with donor lymphocyte infusions (DLI) from the original stem cell donor. Leukaemia-free survival, defined as survival without evidence of relapse at any time after transplant does not account for patients who are successfully treated with DLI. To summarize adequately the response to treatment, a new summary probability, called the current leukaemia free survival (CLFS), is proposed. This quantity is defined as the probability that a patient is alive and in remission at a given time after transplant. We discuss two statistical methods for estimating CLFS. The first is based on a multistate modelling approach. The second is based on an estimate constructed by looking at appropriate differences between Kaplan-Meier estimates. We compare these estimates using data on 189 consecutive patients who underwent SCT over a 7 year period. PMID- 10848795 TI - Exclusive expression of G-CSF receptor on myeloid progenitors in bone marrow CD34+ cells. AB - Although granulocyte colony-stimulating factor (G-CSF) has been reported to act on cells of neutrophilic lineage, the administration of G-CSF to induce the mobilization of various haematopoietic progenitors into the circulation. We analysed the expression of receptors for G-CSF (G-CSFR) on human bone marrow and G-CSF-mobilized peripheral blood CD34+ cells, and examined the proliferation and differentiation capabilities of sorted CD34+G-CSFR+ and CD34+G-CSFR- cells using methylcellulose clonal culture. Flow cytometric analysis showed that G-CSFR was expressed on 14.9 +/- 4.9% of bone marrow CD34+ cells, most of which were included in CD34+CD33+ and CD34+CD38+ cell fractions. In clonal cultures, CD34+G CSFR+ cells produced only myeloid colonies, whereas CD34+G-CSFR- cells produced erythroid bursts, megakaryocyte and multilineage colonies. When incubated with the cytokine cocktail for 5 d, CD34+G-CSFR- cells generated CD34+G-CSFR+ myeloid progenitors. In G-CSF-mobilized peripheral blood, CD34+ cells contained 10.8 +/- 5.8% of G-CSFR+ cells, most of which were also myeloid progenitors, although CD34+G-CSFR- cells contained a substantial number of myeloid progenitors. These results indicated that the expression of G-CSFR on CD34+ cells is restricted to myeloid progenitors, suggesting that the specific activity of G-CSF on myelopoiesis depends on the exclusive expression of its receptor on myeloid progenitors, and that the mobilization of various haematopoietic progenitors is not a direct effect of G-CSF in humans. PMID- 10848796 TI - Ex vivo expanded mobilized peripheral blood CD34+ cells accelerate haematological recovery in a baboon model of autologous transplantation. AB - To address the value of ex vivo expanded haematopoietic cells for shortening cytopenia in autologous haematopoietic transplantation, we designed an ex vivo expansion protocol based on a cocktail of early acting cytokines and short-term culture and tested it in a baboon model. Expansion involved enriched CD34+ peripheral blood haematopoietic cells cultured for 6 d with a combination of FLT3 L, stem cell factor (SCF), thrombopoietin (TPO) and interleukin (IL)-3 (50 ng/ml each); CD34+ cells, granulocyte-macrophage colony-forming units (GM-CFU) and megakaryocytic colony-forming units (MK-CFU) were amplified, respectively, 10.5-, 20.5- and 17.9-fold. Baboons were submitted to a myeloablative regimen consisting of cyclophosphamide plus total body irradiation (TBI; 6 Gy) and were then grafted with either 2 x 106/kg unmanipulated CD34+ cells (control group, n = 4) or cells cultured from 2 x 106/kg CD34+ cells (expansion group, n = 4). No cytokines were administered after transplantation. All the animals engrafted. The mean times to white blood cell (WBC), granulocyte and platelet recovery were significantly shorter in the expansion group than in the control group: WBC (> 1 x 109/l) and neutrophil (> 0.5 x 109/l) recovery occurred on days 8 (range 6-9) and 9 (range 6 11), respectively, compared with days 12 (range 10-15) and 14 (range 11-16); platelets recovered (> 20 x 109/l) on day 9 (range 7-12) compared with day 13 (range 11-15) in the control group (P < 0.05). No toxicity was observed after reinfusion. No secondary hypoplasia was observed during more than 12 months of follow-up. Functions of both neutrophils and platelets produced from expanded cells were normal in terms of oxidative metabolism, chemotaxis and the bleeding time. This study shows that in comparison with unmanipulated cells peripheral blood haematopoietic cells expanded from similar doses of CD34+ cells, under the conditions defined here, accelerated both neutrophil and platelet recovery without impairing long-term haematopoiesis. PMID- 10848797 TI - Fetal bone marrow as a source of stem cells for in utero or postnatal transplantation. AB - We examined the potential of human fetal bone marrow (FBM) as a source of haematopoietic stem cells for transplantation. The median number of cells obtained between 20 and 24 weeks' gestation was 1.9 x 109 and a median 1.17 x 108 of these cells expressed CD34. Flow cytometry was also used to estimate the content of three different candidate stem cell populations in the tissues older than 20 weeks' gestation. A median 8.8 x 105 CD34++CD38- cells, 1.37 x 106 CD34++CD4+ cells and 2.20 x 106 CD34++CD90+ cells were detected. The content of colony-forming units culture (CFU-C) in the FBM ranged from 2.8 x 104 to 6.0 x 106 per fetus. The CFU-C content could be expanded 50-fold by culture for 1 week in serum-deprived medium and the growth factors kit ligand and granulocyte macrophage colony-stimulating factor. Positive selection of FBM CD34+/++ cells was achieved using the Baxter Isolex 50 device. An average purity of 82% and yield of up to 19% of CD34+/++ cells was achieved. T cells were depleted by 99.84%. Analysis of candidate stem cell populations and primitive CFU-C suggested a preferential enrichment of these cells over the total population of CD34+/++ cells. All FBM samples were found to be free of microbial contamination at the time of harvest and after selection of CD34+/++ cells. Thus, FBM is a safe source of stem cells. The large number of progenitors and candidate stem cells that can be obtained from FBM makes it suitable for in utero and possibly postnatal transplantation. PMID- 10848798 TI - Determination of serotonin release from platelets by enzyme immunoassay in the diagnosis of heparin-induced thrombocytopenia. AB - [14C]-Serotonin release assay (14C-SRA) from platelets is considered to be the most sensitive test for laboratory confirmation of heparin-induced thrombocytopenia (HIT). This study compared 14C-SRA with an enzyme immunoassay (EIA) to determine the release of serotonin from platelets in the presence of heparin and serum from HIT patients. The normal range (median, 2.5 and 97.5 percentiles) of serotonin release from platelets in healthy subjects (n = 149) is 38 ng/ml (19 and 62) measured by EIA-SRA. Serum from HIT patients (n = 42) released 2548 ng/ml (244 and 7987) serotonin in the presence of 0.1 IU/ml heparin and 29 ng/ml (13 and 76) in the presence of 100 IU/ml heparin. One hundred per cent and 15% of HIT samples exhibited an elevated serotonin release from platelets in the presence of 0.1 IU/ml low molecular weight (LMW) heparin, 2100 ng/ml (869 and 5968), or danaparoid, 272 ng/ml (143 and 403), respectively. The sensitivity and specificity of the EIA-SRA was 100% and 97.4% and of the 14C-SRA 100% and 92.9% in HIT patients. No false-positive results were found in patients receiving heparin (n = 28), in patients with elevated levels of bilirubin (n = 5), in patients with antiphospholipid antibody syndrome (n = 10) or in non-HIT patients (n = 78) with both assays. The EIA technique to quantify serotonin release from platelets provides a reliable non-radioactive method to diagnose heparin-induced thrombocytopenia and to assess in vitro crossreactivity of low molecular weight heparins and heparinoid. PMID- 10848799 TI - Monovalent binding of autoantibodies to beta2-glycoprotein I, detected using surface plasmon resonance at low antigen density. AB - The precise mechanism of interaction between autoantibodies and beta2 glycoprotein I (beta2GPI) and the experimental conditions to be used for their detection are still under debate. Until now, these interactions have been studied under static conditions. We have investigated the interactions of purified IgG from 25 lupus anticoagulant-positive patients with immobilized beta2GPI under flow conditions by real-time analysis based on surface plasmon resonance technology. Sensor chips were coated with purified human beta2GPI coupled to dextran via amino groups at low densities (1.4, 1.8 or 2. 4 ng beta2GPI/mm2). Four patients' IgG displayed efficient binding and had the highest so-called antiphospholipid IgG levels by enzyme-linked immunosorbent assay (ELISA) and the highest absorbance values in an anti- beta2GPI ELISA at a beta2GPI density reported to be around 12 ng/mm2. Binding of antibodies to the beta2GPI sensor chips proved to be dependent upon the IgG concentration and beta2GPI density and was inhibited by a rabbit antibody against beta2GPI. Similar association and dissociation profiles were observed for the four efficient binders. The fast rate of dissociation limited the binding of autoantibodies to beta2GPI and was highly suggestive of a monovalent association, confirmed by binding of Fab fragments under similar experimental conditions. In conclusion, monovalent binding of low affinity antibodies to beta2GPI immobilized at a density as low as 1.8 ng/mm2 could be detected using surface plasmon resonance. PMID- 10848800 TI - Effect of factor VIII concentrate on antigen-presenting cell (APC)/T-cell interactions in vitro: relevance to inhibitor formation and tolerance induction. AB - Inhibitor formation in patients with haemophilia receiving factor VIII (FVIII) concentrate is a common problem requiring tolerance induction therapy. Immune tolerance is dependent on defective T cell/antigen-presenting cell (APC) interactions and inhibitor antibody formation is associated with effective T cell/B-cell interaction. We studied the expression of the cell-surface molecules involved with these interactions using multiparameter flow cytometry and a whole blood stimulation assay-phytohaemaglutinin (PHA) to activate T cells and Escherichia coli lipopolysaccharide (LPS) to activate monocytes and B cells. Up regulation of T-cell co-stimulatory receptors CD11a, CD40 ligand (CD40L) and CTLA4 were inhibited in a dose-dependent manner by plasma-derived (pd)FVIII, but CD28 was unchanged. Up-regulation of monocyte and B-cell co-stimulatory ligands CD4O, B7-1 (CD80) and B7-2 (CD86) were also inhibited in a dose-dependent manner by pdFVIII, but LFA-3 (CD58) was unchanged. The combined inhibitory effect of prednisolone, an immunosuppressive agent used in several tolerance induction protocols, with pdFVIII on co-stimulatory molecules, was additive. There was no significant alteration in T-cell/APC adhesion or co-stimulatory molecules noted in the presence of recombinant (rh)FVIII concentrate. The inhibitory effect of pdFVIII on molecules involved in interaction between T cells and APCs may result in immune tolerance in recipients of pdFVIII concentrate. The inhibitory effect of pdFVIII on CD40/CD40L up-regulation may result in defective antibody formation. We now provide evidence that the use of pdFVIII, through interfering with APC/T-cell interactions, may be more appropriate than rhFVIII for tolerance induction. PMID- 10848801 TI - Cell-surface expression of transrearranged Vgamma-cbeta T-cell receptor chains in healthy donors and in ataxia telangiectasia patients. AB - Transrearrangements between the T-cell receptor (TCR) VgammaI family and JbetaCbeta loci occur at increased frequencies in patients with ataxia telangiectasia (AT). We have used an optimized reverse transcriptase polymerase chain reaction (RT-PCR) approach to investigate the occurrence of TCRVgamma JbetaCbeta transrearrangements involving the dominantly used Vgamma element in peripheral blood gammadelta T cells, i.e. Vgamma9. We detected in frame transcripts of Vgamma9-JbetaCbeta transrearrangements in 4/16 AT patients and in 3/13 healthy control donors. A panel of monoclonal antibodies (mAb) against all expressed TCRVgamma elements was used to monitor cell-surface expression of transrearranged TCR. A very low proportion (< 1%) of peripheral blood TCRalphabeta cells expressed Vgamma instead of Vbeta elements. For the first time, we have isolated and molecularly characterized alphabeta T cells with a Vgamma9-JbetaCbeta transrearrangement from two AT patients and we have shown that such TCR are functional. We conclude that functional TCR transrearrangements can also involve Vgamma9, the dominant Vgamma element in peripheral blood gammadelta T cells. PMID- 10848802 TI - Chronic overexpression of membrane-bound flt3 ligand by T lymphocytes in severe aplastic anaemia. AB - Aplastic anaemia (AA) is an immune-mediated bone marrow failure associated with high serum levels of flt3 ligand (FL). We examined expression of the membrane bound isoform of FL in peripheral blood and bone marrow cells from AA patients at diagnosis (n = 16) and after immunosuppressive (IS) treatment (n = 36). Flow cytometry demonstrated strongly increased FL levels on the cell surface of T lymphocytes in AA relative to normal controls (P < 0.0001). T-cell-specific expression of membrane-bound FL was confirmed by confocal microscopy. FL mRNA and total cellular FL protein levels were increased about threefold. Overexpression of FL in AA was observed for up to 20 years after IS treatment. FL levels correlated inversely with CD34+ cell numbers and the colony-forming ability of AA bone marrow (R = -0.68 and -0.85 respectively). Histological examination of spleen specimens and bone marrow biopsies gave no evidence of degeneration or fibrosis due to prolonged exposure to high FL. Levels of membrane-bound FL were not increased in autoimmune diseases (n = 23), including rheumatoid arthritis and lupus erythematosus, nor in graft-versus-host disease (n = 8). Chronic overexpression of FL on the surface of T lymphocytes in AA, but not in other T cell-mediated disorders, suggests that membrane-bound FL plays a role in cell cell interactions in bone marrow failure and may be important for long-term haemopoietic recovery. PMID- 10848803 TI - Apoptosis in factor-dependent haematopoietic cells is linked to calcium-sensitive mitochondrial rearrangements and cytoskeletal modulation. AB - Apoptosis in murine haematopoietic interleukin (IL)3-dependent cell lines is induced within 6-8 h by IL-3 withdrawal. Direct introduction of cytochrome c by electroporation induces apoptosis within 2 h and was inhibited by caspase inhibitors, such as Z-VADfmk and Z-Dfmk. We report here that apoptosis induced by IL-3 withdrawal was refractory to these inhibitors but was accompanied by striking redistribution of mitochondria, which aggregated into an area associated with centrioles without loss of Deltapsim. Both mitochondrial redistribution and apoptosis were inhibited by the calcium ionophore, ionomycin. Nocodozole, an inhibitor of microtubule assembly, also induced apoptosis, which was unaffected by caspase inhibitors. Although nocodozole did not alter mitochondrial distribution, it significantly reduced Deltapsim, and both reduction of Deltapsim and apoptosis were inhibited by ionomycin. Oligomycin, which inhibits the mitochondrial FoF1 ATPase, similarly induced apoptosis, which was unaffected by caspase inhibitors but was inhibited by ionomycin. Further, oligomycin stimulated the novel formation and release of surface membrane-derived vesicles containing mitochondria with intact Deltapsim; ionomycin also inhibited their production. In all these conditions, Bcl-2 protected cells from apoptosis. Our studies show that apoptosis induced by three very different agents shares insensitivity to caspase inhibitors, suppression by ionomycin and effects on mitochondria, which all appear to be linked to cytoskeletal/microtubule activity. They suggest that microtubules and the cytoskeleton play an important role in apoptosis through mechanisms affecting mitochondria but which are independent of cytochrome c release. PMID- 10848804 TI - Mesenchymal progenitor cells in human umbilical cord blood. AB - Haemopoiesis is sustained by two main cellular components, the haematopoietic cells (HSCs) and the mesenchymal progenitor cells (MPCs). MPCs are multipotent and are the precursors for marrow stroma, bone, cartilage, muscle and connective tissues. Although the presence of HSCs in umbilical cord blood (UCB) is well known, that of MPCs has been not fully evaluated. In this study, we examined the ability of UCB harvests to generate in culture cells with characteristics of MPCs. Results showed that UCB-derived mononuclear cells, when set in culture, gave rise to adherent cells, which exhibited either an osteoclast- or a mesenchymal-like phenotype. Cells with the osteoclast phenotype were multinucleated, expressed TRAP activity and antigens CD45 and CD51/CD61. In turn, cells with the mesenchymal phenotype displayed a fibroblast-like morphology and expressed several MPC-related antigens (SH2, SH3, SH4, ASMA, MAB 1470, CD13, CD29 and CD49e). Our results suggest that preterm, as compared with term, cord blood is richer in mesenchymal progenitors, similar to haematopoietic progenitors. PMID- 10848805 TI - Neutropenia and anaemia due to carbimazole-dependent antibodies. AB - Carbimazole-dependent antibodies to erythrocytes were detected in the sera of three anaemic patients who had been treated with carbimazole for hyperthyroidism. By the use of Rhnull-typed erythrocytes, we could show that some of these were directed against the proteins of the Rh complex. Carbimazole-dependent antibodies eluted from erythrocytes showed no binding to other blood cells. One patient also presented with neutropenia and mild thrombocytopenia. Additional carbimazole dependent antibodies against the neutrophil-specific Fcgamma receptor IIIb (FcgammaRIIIb, CD16b) and the broadly expressed platelet endothelial cell adhesion molecule 1 (PECAM-1; CD31) were detected in this patient's serum. Surprisingly, the PECAM-1-reactive drug-dependent antibodies were also detectable in the sera of the other two patients with normal leucocyte and platelet counts. We assume that carbimazole can induce cell lineage-specific drug-dependent antibodies that cause cytopenia and also drug-dependent antibodies against the broadly expressed PECAM-1 molecule that may cause mild but not severe cytopenia. PMID- 10848806 TI - Low prevalence of chronic hepatitis C virus infection in B-cell non-Hodgkin's lymphoma patients from a population with a high prevalence of healthy hepatitis c virus carriers. PMID- 10848807 TI - Short course of busulphan in essential thrombocythaemia: remodelling of an old strategy. PMID- 10848808 TI - Mutation analysis of BCL 10 in acute myeloid leukaemia. PMID- 10848809 TI - Gallium positivity in Hodgkin's disease. PMID- 10848810 TI - The current status of lymphoma classification. PMID- 10848811 TI - Paraffin section immunotyping of leukaemias. PMID- 10848812 TI - Dynamics of telomere shortening in neutrophils and T lymphocytes during ageing and the relationship to skewed X chromosome inactivation patterns. AB - Human haemopoiesis undergoes profound changes throughout life, resulting in compromised regenerative capacity of haemopoietic stem cells. It has been suggested that telomere shortening results in senescence of haemopoietic stem cell subsets and may influence the balance between stem cell renewal and proliferation. Telomere length and telomerase activity was measured in whole blood leucocytes, neutrophils and T cells from cord blood and individuals aged from 1 year to 96 years. Rapid telomere shortening [700 base pairs (bp)] was demonstrated in the first year of life, followed by a gradual decline of 31 bp/year. T cells were shown to have longer telomeres than neutrophils (mean difference 372 bp, P = < 0.001) but demonstrated similar rates of shortening (20 +/- 0.3 bp/year vs. 22 +/- 0.3 bp/year). Telomerase was detectable in T cells but not in neutrophils, suggesting that telomerase is not the rate-limiting step for regulation of telomere length in haemopoietic cells. Stem cell utilization as measured by X chromosome inactivation patterns was found to be independent of telomere length. This supports the concept that age-dependent skewed haemopoiesis is the result of random stem cell loss or X-allelic exclusion rather than telomeric senescence. These studies provide insight into the ageing process and a reference point for evaluating replicative stress in individuals of different age groups. PMID- 10848813 TI - The monoclonal antibody TER-119 recognizes a molecule associated with glycophorin A and specifically marks the late stages of murine erythroid lineage. AB - The antigen specificity of a rat monoclonal antibody TER-119 was investigated. In adult mice, TER-119 reacted with mature erythrocytes, 20-25% of bone marrow cells and 2-3% of spleen cells but not with thymocytes nor lymph node cells. In fetal haematopoietic tissues, 30-40% of d 10 yolk sac cells, 80-90% of d 14 fetal liver cells and 40-50% of newborn liver cells were reactive with TER-119. TER-119+ cells in adult bone marrow expressed significant levels of CD45 but not myeloid (Mac-1, Gr-1) or B-cell (B220) markers. Morphological examination and haematopoietic colony-forming assays for isolated TER-119+ cells revealed that TER-119 reacts with erythroid cells at differentiation stages from early proerythroblast to mature erythrocyte, but not with cells showing typical erythroid blast-forming unit (BFU-E) and erythroid colony-forming unit (CFU-E) activities. Erythroleukaemia cell lines do not express the TER-119 antigen even after stimulation with dimethylsulphoxide. TER-119 immunoprecipitated protein bands with molecular masses of 110 kDa, 60 kDa, 52 kDa and 32 kDa from erythrocyte membrane, whereas only a 52-kDa band was detected by TER-119 in Western blot analysis. Further molecular and cellular analyses indicated that the TER-119 antigen is a molecule associated with cell-surface glycophorin A but not with glycophorin A itself. PMID- 10848814 TI - Activity of interleukin 6 in the differentiation of monocytes to macrophages and dendritic cells. AB - Peripheral blood monocytes are common precursor cells of dendritic cells (DCs) and macrophages. We have searched for factors with the potential to regulate the differentiation of monocytes to DCs and macrophages. When CD14+ monocytes are cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL) 4, the CD14+CD1a- population, which consists of macrophages, was found in the serum-containing cultures but not in the serum-free cultures. Addition of IL-6 receptor-neutralizing monoclonal antibody (mAb) or gp130 neutralizing mAb to the serum-containing cultures resulted in a decreased population of CD14+CD1a- cells. An increase in the CD14+CD1a- population with reduction in CD14-CD1a+ DCs was observed with the addition of IL-6 to cultures, whereas IL-11, leukaemia inhibitory factor, oncostatin M or macrophage colony stimulating factor did not affect the differentiation of monocytes in the presence of GM-CSF plus IL-4. This effect of IL-6 was blocked by tumour necrosis factor alpha (TNF-alpha), lipopolysaccharide (LPS), IL-1beta, CD40 ligand (CD40L) and transforming growth factor beta1 (TGF-beta1). Among these factors, TNF-alpha was most potent in interfering with the action of IL-6. These results suggest that IL-6 inhibits the differentiation of monocytes to DCs by promoting their differentiation toward macrophages, which is modulated by factors such as TNF alpha, LPS, IL-1beta, CD40L and TGF-beta1. PMID- 10848815 TI - Interferon alpha2b directly induces fibroblast proliferation and transforming growth factor beta secretion of macrophages. AB - To elucidate the effects of interferon alpha2b (IFN-alpha) on normal human bone marrow, fibroblasts from patients without haematopoietic pathology were cultivated and used in stimulation experiments. Further, co-cultures with highly enriched fractions of megakaryocytes and bone marrow macrophages were analysed. In this context, the influence of cell-to-cell interactions and humoral factors was determined in transwell and neutralization studies. Finally, secretion of platelet-derived growth factor (PDGF) and transforming growth factor beta1 (TGF beta1) by single megakaryocytes and macrophages was examined by using the reverse haemolytic plaque assay (RHPA). Following these experimental designs, a direct proliferative activity of IFN-alpha on bone marrow fibroblasts could be demonstrated. In the unstimulated co-cultures, the megakaryocyte- but not the macrophage-enriched fraction induced fibroblast growth and [3H]-thymidine uptake. This effect was dependent on cell-to-cell contact and also on the influence of TGF-beta and PDGF. In the megakaryocyte-enriched co-cultures, the fibroblast proliferation was not altered by IFN-alpha, but in the macrophage fibroblast cultures addition of IFN-alpha enhanced fibroblast growth and [3H]-thymidine uptake was distinctively higher than in the monocultures. This effect was not obvious in the transwell or neutralization experiments. Finally, IFN-alpha treatment exerted a significantly elevated TGF-beta1 secretion in single macrophages. Our findings are in keeping with the assumption that several pathomechanisms participate in IFN-alpha-induced myelofibrosis, including direct and indirect effects. PMID- 10848816 TI - Association of extracellular matrix proteins fibulin-1 and fibulin-2 with fibronectin in bone marrow stroma. AB - Extracellular matrix (ECM) molecules, together with growth factors and stromal cells, regulate haematopoietic cell development in bone marrow (BM). We report here expression of ECM proteins fibulin-1 and fibulin-2 in mouse BM. In other tissues, fibulin-1 and fibulin-2 associate with fibronectin and other ECM proteins. Fibulin-2 has also been found to adhere to cells via beta3 integrins. We studied the association of fibulins with fibronectin in BM stroma. By confocal microscopy, fibulin-1 and fibulin-2 immunostainings were co-localized with fibronectin in the adherent layer of long-term BM cultures. In cell adhesion assays using recombinant proteins, mouse fibulin-2 adhered to human erythroid megakaryocytic leukaemia cell line HEL. This adhesion was mediated by beta3 integrins. However, HEL cells did not adhere to human fibulin-2. We therefore studied a possible species-specific cell-adhesive activity of mouse fibulin-2 by using mouse megakaryocytes, obtained by culture of BM cells in the presence of thrombopoietin. These megakaryocytes did not adhere to mouse fibulin-2. Our findings suggested that the functional role of fibulin-1 and fibulin-2 in BM stroma is related to binding to the major cell adhesion protein fibronectin, whereas adhesion of mouse fibulin-2 to human cells containing the integrin beta3 chain is not related to an apparent physiological function of the protein. PMID- 10848817 TI - Ex vivo expansion of mature human neutrophils with normal functions from purified peripheral blood CD34+ haematopoietic progenitor cells. AB - Purified CD34+ haematopoietic progenitor cells were cultivated with stem cell factor, interleukin 3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte CSF (G-CSF) for 7 d, and thereafter non-adherent cells were divided into two groups. Cells in one group (group A) were further cultivated for 7 d with four cytokines, and cells in the other group (group B) were further cultivated for 7 d with G-CSF alone. On day 14, 220-fold and 130 fold increases in the numbers of non-adherent cells were achieved for groups A and B respectively. These cell preparations contained 65% granulocytes for group A and 95% granulocytes for group B. These cells gained the ability to respond effectively with chemotaxis, phagocytosis and superoxide (O2-) release. Cells in group B were appropriately primed by G-CSF, GM-CSF, tumour necrosis factor alpha and IL-1beta for enhanced release of O2 -. The responsiveness of these cells was identical to that of peripheral blood neutrophils, indicating that cells in group B may be in the resting state. In contrast, cells in group A were not primed by these cytokines for enhanced release of O2- and released a large amount of O2- spontaneously, indicating that cells in group A may be in the activated state. These findings indicate that mature neutrophils with normal functions were expanded ex vivo in group B and suggest that these cells could be used for possible autologous neutrophil transfusion to prevent bacterial infections during severe neutropenia after cytotoxic chemotherapy. PMID- 10848818 TI - Hydroxyurea therapy decreases the in vitro adhesion of sickle erythrocytes to thrombospondin and laminin. AB - The adhesion of sickle erythrocytes to the vascular endothelium and subendothelial matrix probably contributes to the pathogenesis of vaso-occlusive disease. The chemotherapeutic agent hydroxyurea (HU) decreases the frequency of vaso-occlusive crises in patients with sickle cell disease. However, the exact mechanism(s) of HU's effect on vaso-occlusive crises is not fully understood. The goal of this study was to determine the effect of HU therapy on the adhesion of sickle erythrocytes to the subendothelial matrix proteins thrombospondin (TSP) and laminin under conditions of flow in vitro. Erythrocytes from patients with severe sickle cell disease on HU therapy (n = 14) had significantly less adhesion to TSP (687 +/- 92 erythrocytes/mm2, mean +/- SE) than untreated patients with severe disease (n = 18, 1176 +/- 117 erythrocytes/mm2, P = 0.003). In addition, there was significantly less adhesion of erythrocytes to immobilized laminin in patients treated with HU (1695 +/- 293 erythrocytes/mm2) than in the untreated patients (2590 +/- 296 erythrocytes/mm2, P = 0.02). Erythrocytes from an additional nine patients with severe sickle cell disease were studied both before and after initiation of HU therapy. Erythrocytes from these patients became less adhesive to both TSP (P = 0.001) and laminin (P = 0.01), a change that was sustained in most patients throughout the duration of the study (2 months to > 12 months). This study suggests that HU modulates the adhesive phenotype of sickle erythrocytes, an effect that may be in addition to, or independent of, other known effects of HU, such as an increase in fetal haemoglobin level. PMID- 10848819 TI - Paroxysmal cold haemoglobinuria in an adult with chicken pox. AB - Paroxysmal cold haemoglobinuria (PCH) is an autoimmune disorder characterized by intravascular haemolysis causing haemoglobinuria. It is due to a biphasic haemolysin known as the Donath-Landsteiner antibody, which binds specifically to the P antigen of red blood cells at low temperatures, leading to complement activation and red cell lysis at 37 degrees C. PCH is a rare disease which predominantly affects the paediatric population, occurring mostly during viral infections. We report on what is possibly the first case of PCH in an adult to be precipitated by chicken pox infection. PMID- 10848820 TI - Thrombopoietin levels in cord blood plasma and amniotic fluid in fetuses with alloimmune thrombocytopenia and healthy controls. AB - To extend our knowledge of the kinetics of fetal thrombopoietin (TPO), we studied TPO levels in cord blood plasma and amniotic fluid collected from 15 fetuses considered to be at risk of fetomaternal alloimmune thrombocytopenia and also from 10 healthy controls at caesarean delivery. In the plasma of all 25 fetuses and newborn infants studied, TPO was detected above the lower limit of detection (7 pg/ml) and correlated inversely with platelet counts (r = -0.53, P = 0.006). At term, TPO detected in amniotic fluid was at significantly lower levels (7 pg/ml; range 0-22 pg/ml) than simultaneously obtained cord plasma TPO (114 pg/ml; range 43-201 pg/ml; P < 0.001). There was no correlation between levels of TPO in amniotic fluid and cord plasma or platelet counts. In the serial samples collected from the five fetuses with HPA-1a alloimmunization before 37 weeks' gestation, the TPO levels in amniotic fluid were significantly higher than at term (P = 0.013): from 22 to 28 weeks' gestation, 42 pg/ml (30-78 pg/ml); from 32 weeks', 24 pg/ml (17-33 pg/ml); at term, 8 pg/ml (4-13 pg/ml), correlating inversely with gestational age (r = -0.81, P = 0.003). Thus, TPO is present in amniotic fluid at levels apparently inversely related to gestational age. Whether these high levels seen early in pregnancy are normal or are associated with the HPA-1 alloimmunization remains to be shown. PMID- 10848821 TI - Affinity purification of heparin-dependent antibodies to platelet factor 4 developed in heparin-induced thrombocytopenia: biological characteristics and effects on platelet activation. AB - Antibodies to heparin platelet factor 4 (H-PF4) complexes were purified from the plasma of three patients with heparin-induced thrombocytopenia (HIT) using affinity chromatography. From each plasma, the largest amount of antibodies was eluted with 2 M NaCl at pH 7.5 (peak 1) and the remainder was obtained using 0.1 M glycine/0. 5 M NaCl at pH 2.5 (peak 2). In an enzyme-linked immunosorbent assay (ELISA), we then showed that each patient had developed antibodies to PF4 displaying different characteristics. In patient 1, peak 1 IgG reacted almost exclusively with H-PF4 complexes, whereas peak 2 IgG had similar reactivity with PF4 whether or not heparin was present. Patient 2 expressed a mixture of IgA, IgM and IgG and both fractions bound to PF4 alone or to H-PF4 complexes. Finally, IgG in patient 3 only bound to H-PF4 and was unreactive with PF4 alone. Using [14C] serotonin release assays, the antibodies developed in the three patients and exhibiting the strongest ability to activate platelets with heparin were those having the highest affinity to H-PF4. These results strongly support the hypothesis that HIT antibodies to PF4 are heterogeneous regarding their affinity and specificity for target antigens and this may greatly influence their ability to activate platelets and their pathogenicity. PMID- 10848822 TI - Plasminogen activator inhibitor 2 and urokinase-type plasminogen activator in plasma and leucocytes in patients with severe sepsis. AB - Proteins influencing plasminogen activation to plasmin, namely plasminogen activators tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) and their principal inhibitors, plasminogen activator inhibitor 1 (PAI-1) and PAI-2, were measured in the plasma, the polymorph and mononuclear cell fractions taken from patients with major sepsis who were entering a general intensive care unit. The purpose of this study was to elucidate the factors favouring the persistence of fibrin in the microvasculature and thus contributing to multiple organ failure. Levels of u-PA antigen in plasma rose in sepsis and u-PA activity, not detectable in normal plasma, appeared. Levels of u-PA antigen in the cell fractions fell concomitantly. t-PA antigen in plasma and in the mononuclear cell fraction rose in sepsis, but t-PA activity was not detectable. Plasma PAI-1 antigen levels were strikingly raised in sepsis, presumably accounting for the complete neutralization of t-PA activity. PAI-2 antigen, not normally detected in plasma, appeared in the plasma of some patients, whereas it disappeared from the cellular fractions. Appearance of PAI-2 in plasma was associated with non-survival of the patient. The observations indicate that all the agents involved in plasminogen activation are released into the plasma in major sepsis. The levels of PAI-1 reached were quantitatively sufficient to suppress all activity of the released t-PA, but the inhibitors did not prevent expression of u-PA activity in the circulation. Circulating active u PA and PAI-2 in the plasma of patients with severe sepsis may represent material originating from leucocytes. Leucocyte release of these agents within fibrin deposits may influence the persistence of fibrin and thus the development of multiple organ failure. PMID- 10848823 TI - A single nucleotide polymorphism at nucleotide -1793 in the von Willebrand factor (VWF) regulatory region is associated with plasma VWF:Ag levels. AB - von Willebrand Factor (VWF) is a large multimeric glycoprotein involved in the transport and protection of factor VIII and in mediating platelet-subendothelium and platelet-platelet interactions. We have documented the presence of a single nucleotide polymorphism (SNP) at nucleotide (nt) -1793 (G 0.36 or C 0.64) in the VWF 5'-flanking sequence. This polymorphism is in strong linkage disequilibrium with the previously reported SNPs at nts -1234, -1185 and -1051 and, like this other group of polymorphisms, shows a significant association with plasma VWF levels. This association is more marked in subjects who are more than 40 years of age. Further, circumstantial evidence to support a role for the -1793 sequence in regulating VWF expression comes from our demonstration of differential binding of endothelial cell nuclear proteins, including the transcription factor NFkappaB, by this sequence. In summary, the association of the -1793 SNP with plasma VWF levels provides additional evidence for the role of the VWF regulatory region between nts -1793 and -1051 in controlling VWF expression. PMID- 10848824 TI - Liver biopsy in Irish hepatitis C-infected patients with inherited bleeding disorders. AB - The majority of patients receiving plasma-derived clotting factor concentrates between 1970s and the mid-1980s are now hepatitis C positive. The progression of hepatitis C is extremely variable and there is frequently a poor correlation among liver biochemistry, viral load and the stage of liver disease. Liver biopsy remains the only definitive way of staging fibrosis and grading necroinflammatory activity. Concerns have been expressed about the safety of the procedure; however, with modern regimes for the correction of coagulopathy in patients with inherited bleeding disorders, normal haemostasis may be maintained during the peribiopsy period. We performed 21 liver biopsies between 1984 and 1997 on patients with factor VIII (FVIII) or IX (FIX) deficiency and von Willebrand's Disease (VWD). Four had concomitant human immunodeficiency virus (HIV) infection, five were thrombocytopenic and one had a prolonged prothrombin time (PT). Haemostasis was achieved using an intermittent bolus of factor concentrate or continuous infusion regimens. One patient with VWD received Desmopressin (DDAVP). There were no bleeding episodes associated with biopsy. We suggest that liver biopsy is a safe procedure in patients with inherited bleeding disorders when the coagulopathy is fully corrected. It is the only definitive method of staging the extent of fibrosis associated with hepatitis C infection, and it is this that defines prognosis. PMID- 10848825 TI - Re-exposure to recombinant (r)-hirudin in antihirudin antibody-positive patients with a history of heparin-induced thrombocytopenia. AB - Patients with a history of heparin-induced thrombocytopenia (HIT) and antibodies to hirudin were re-exposed to recombinant (r)-hirudin for prophylaxis of thromboembolism. Four patients were re-exposed to 2 x 25 mg of subcutaneous r hirudin for 8-27 d. Two patients were re-exposed once, one patient twice and one four times. Re-exposure was well tolerated in all patients and no thromboembolism occurred. Antihirudin IgG (4/4 patients), IgA and IgM (1/4 patients) antibody levels increased. Baseline ecarin clotting times showed high variability. Patients with antibodies to hirudin may be re-exposed but anticoagulant monitoring is mandatory. PMID- 10848826 TI - Prenatal and peripartum management of congenital afibrinogenaemia. AB - We experienced three cases and four successful deliveries with congenital afibrinogenaemia and propose the following guidelines for the prenatal and peripartum management: (i) genital bleeding usually begins at 5 weeks' gestation and spontaneous abortion always occurs at 6-8 weeks' gestation without fibrinogen infusion; (ii) the fibrinogen level must be at least 0.60 g/l and, if possible, higher than 1.0 g/l during the pregnancy; (iii) the necessary amounts of fibrinogen increase as the pregnancy progresses and the preterm labour occurs; (iv) the fibrinogen level under the continuous infusion of fibrinogen during labour must be at least 1.5 g/l and, if possible, higher than 2.0 g/l to prevent placental abruption; (v) the puerperium is usually uneventful with a reduced dose of fibrinogen infusion. PMID- 10848827 TI - Treatment of anaemia in myelodysplastic syndromes with prolonged administration of recombinant human granulocyte colony-stimulating factor and erythropoietin. AB - Treatment with recombinant human erythropoietin (rhEPO) improves anaemia in approximately 20% of the patients with myelodysplastic syndromes (MDS). Recent reports suggest that a combination treatment with rhEPO plus recombinant human granulocyte colony-stimulating factor (rhG-CSF) given for up to 18 weeks may result in a higher erythroid response rate than with rhEPO alone. We investigated the potential advantage of an even more prolonged schedule of combined rhG-CSF and rhEPO treatment to obtain and maintain stable responses. In a phase II study, 33 patients with MDS [17 with refractory anaemia (RA), eight with RA with ringed sideroblasts (RARS), eight with RA with excess blasts (RAEB) with bone marrow blast counts less than 20%] were scheduled to receive at least 36 weeks of combined therapy with rhG-CSF and rhEPO. Seventeen of 28 evaluable patients demonstrated an erythroid response [61%; 95% confidence interval (CI) 41-78] after 12 weeks of treatment. The erythroid response rate was 80% (20 of 25 evaluable patients; 95% CI 59-93) after 36 weeks. Seven of these responses developed between week 12 and week 36, whereas two initially responding patients became refractory. The cytokine therapy was generally well tolerated. Nineteen of the 20 patients responding after 36 weeks continued to be treated with both cytokines. After 1 year and 2 years of continuous combined treatment, 50% of the initially included patients showed a continuing response. Our results suggest that a prolonged combination treatment with rhG-CSF and rhEPO is highly effective in achieving a stable and long-lasting erythroid response in many patients with MDS and low blast count. PMID- 10848828 TI - Signal antonymy unique to myelodysplastic marrows correlates with altered expression of E2F1. AB - Myelodysplastic syndromes (MDS) have previously been reported to show competitively high rates of apoptosis and proliferation in the bone marrow (BM). Using a double-labelling technique in the present study, we demonstrated that a significantly high number of S-phase cells were simultaneously apoptotic (signal antonymy; SA) in MDS (mean +/- s.e.m. 53.5 +/- 6.7%, n = 24, P < 0.001). In contrast, SA was negligible in all other specimens studied, including normal control BM (n = 13) from non-Hodgkin's lymphoma (NHL) patients, BM from patients with de novo acute myelogenous leukaemia (1'AML; n = 5), or secondary AML that had transformed from MDS (2'AML; n = 10), or the solid tumours from patients with NHL (n = 9) or head and neck squamous cell carcinoma (HNSCC; n = 10). Subsequently, the expression of a transcription factor, E2F1, was studied in density-separated BM aspirate mononuclear cells from MDS patients (n = 9) and a normal control. Two separate sets of primers were used that recognized the regulatory retinoblastoma (Rb) protein-binding region and the functional DNA binding region of E2F1. Interestingly, although the latter manifested the expected band (280 bp) in all samples, the Rb-specific primers showed the expected band (380 bp) in the normal and in 4/9 MDS specimens. Two other MDS specimens also showed a smaller band ( approximately 325 bp), whereas 3/9 MDS patients showed exclusively the smaller band. The levels of SA were significantly higher in those MDS cases that showed the smaller Rb-specific band either alone or in addition to the expected band (median 19.5%, n = 4, P = 0.037) than in those showing exclusively the expected band (median 0.4%, n = 3). Our present studies show SA as a characteristic feature of MDS and, importantly, demonstrate its link with an altered expression of E2F1 in some MDS patients. PMID- 10848829 TI - Differential effect of interferon alpha on chronic myelogenous leukaemia and normal haematopoietic progenitors in a stromal cell co-culture context: role of the flt3 ligand. AB - Interferon alpha (IFN-alpha) is used to treat chronic myelogenous leukaemia (CML) patients. However, its target(s) remain(s) unknown. One possibility is that there is a differing sensitivity of the leukaemic from the normal colony-forming cell (CFC) compartments to IFN-alpha. Co-cultures of progenitors with stromal cells provide a valuable tool to dissect direct and indirect activities of IFN-alpha. In this study, we have used endothelial cells (EC) as a source of stromal cells. In co-cultures of normal progenitors with EC, IFN-alpha increased the generation of clonogenic cells, mainly via an increased production of flt3 ligand (FL) by EC. In contrast, in co-cultures of CML progenitors with EC, IFN-alpha inhibited the generation of clonogenic cells, mainly by direct inhibition on the progenitors, the up-regulation of FL production by stromal cells being unable to compensate for the direct inhibitory effects of IFN-alpha. These data provide evidence for a differential effect of IFN-alpha on the growth of CML and normal CFC cells in a stromal context and suggest that an alteration in the response of CML progenitor cells to FL is important in the explanation of this differential effect. PMID- 10848830 TI - Successful modulation of high-dose cytosine arabinoside metabolism in acute myeloid leukaemia by haematopoietic growth factors: no effect of ribonucleotide reductase inhibitors fludarabine and gemcitabine. AB - High-dose cytosine arabinoside (AraC)-containing regimens have shown the highest antileukaemic efficacy of all currently used regimens in the treatment of acute myeloid leukaemia (AML). This study aimed at increasing the antileukaemic potential of high-dose AraC by raising intracellular levels of AraC triphosphate (AraCTP), which is the mediator of cytotoxicity, via biochemical modulation by inhibitors of ribonucleotide reductase (RR) or haematopoietic growth factors (HGFs). Blasts from patients with de novo AML were analysed for their formation of AraCTP under high-dose AraC conditions (20 microM over 3 h) without prior modulation (n = 47) after a 2-h pre-exposure with fludarabine (50 microg/ml) (n = 40) or gemcitabine (30 ng/ml) (n = 40) and after a 48-h pre-exposure to granulocyte colony-stimulating-factor (G-CSF; 100 ng/ml) (n = 27) or granulocyte macrophage colony-stimulating-factor (GM-CSF; 100 U/ml) (n = 28). Unmodulated formation of AraCTP (median 239.8 ng/107 cells) could not be increased via modulation by gemcitabine (232.4 ng/107 cells) or fludarabine (247.8 ng/107 cells). The lack of effect of RR inhibitors was also observed for all other known metabolites of AraC [Ara-cytosine monophosphate (CMP), Ara-cytosine diphosphate (CDP), AraCDP-choline, Ara-uridine monophosphate (UMP), Ara-uridine diphosphate (UDP) and Ara-uridine triphosphate (UTP)]. In contrast, pre-exposure to HGFs led to significant increases in AraCTP formation (G-CSF 556.0 ng/107 cells, 2.31-fold increase, P < 0.001; GM-CSF 447.9 ng/107 cells, 1.87-fold increase, P < 0.0001). To establish the mechanism responsible for these effects, the activity of the rate-limiting enzyme of AraC metabolism, deoxycytidine kinase (dCK), was investigated (n = 33). In vivo exposure to GM-CSF led to increases in dCK activity from unmodulated values at 0 h (29.8 pmol/min/mg protein) to 34.3 pmol/min/mg protein at 24 h (1.15-fold increase) and 54.5 pmol/min/mg protein at 48 h (1. 83-fold increase). The raise in dCK activity over 48 h was significant (P < 0.013). PMID- 10848831 TI - Establishment and characterization of an arsenic-sensitive monoblastic leukaemia cell line (SigM5). AB - Few human monoblastic cell lines have been characterized to date. We have established the SigM5 cell line from a patient with acute monoblastic leukaemia (FAB M5a). Original leukaemic cells had a karyotype of 47,XY,+8, whereas the cell line showed a stemline clone of 81,XX,Y,Y,1,4,6,7,+8,+8,9,10,10,11,13,16,19[cp], with a minor sideline also present. Cytochemical staining was strongly positive with alpha-naphthylbutyrate acetate esterase, particulate positive with Sudan black and weakly positive for myeloperoxidase. Cells were positive for CD13, CD15, CD18, CD23, CD33, CD38, CD45, CD68 and myeloperoxidase. CD14 expression was 3-15%. SigM5 constitutively secreted interleukin (IL)-2, IL-8, IL-10, tumour necrosis factor (TNF)-alpha, ferritin, lysozyme, N-elastase and neopterin upon stimulation with interferon (IFN)-gamma. Cells expressed the proinflammatory mediator macrophage migration inhibitory factor (MIF). All NADPH oxidase subunits were constitutively present, but nitroblue tetrazolium reduction was only detectable upon activation with IFN-gamma. SigM5 monoblasts were sensitive to arsenic trioxide (As2O3) previously not described to induce apoptosis in monoblastic cells. Differing considerably in morphology, immunophenotype and sensitivity to arsenics from the widely used cell lines U937, HL-60 and THP-1, SigM5 is a new monoblastic cell line useful for studying leukaemogenesis, monocyte differentiation and tumour cell susceptibility to arsenic compounds. PMID- 10848833 TI - Absence of herpesvirus DNA sequences in the 5T murine model of human multiple myeloma. AB - Kaposi's sarcoma-associated herpesvirus (KSHV, also known as HHV-8) has been found in patients with multiple myeloma (MM) and postulated to be aetiologically associated with the development of this common plasma cell malignancy. A murine model of MM was previously established in which intravenous transfer of 5T myeloma cells into C57BL/KaLwRij mice resulted in characteristic features of human MM. In the present study, we sought to identify herpesvirus DNA sequences in this murine model of MM through polymerase chain reaction (PCR) analysis using primers specific for KSHV, murine herpesvirus 68 (MHV68) and murine cytomegalovirus (MCMV) as well as consensus primers designed from the highly conserved DNA polymerase genes of the Herpesviridae family. None of the DNA samples from whole bone marrow (n = 6) or dendritic cells enriched by long-term culture (n = 8) of 5T myeloma-bearing mice as well as the 5T myeloma cell lines (n = 3) maintained in long-term culture yielded specific amplification products in any of the PCR assays. Two KSHV-specific serological assays measuring antibodies to KSHV latent and lytic antigens also failed to detect the presence of anti-KSHV antibodies in mice that developed MM. These results suggest that the development of 5T murine MM is unlikely to be involved with KSHV or a KSHV-like murine herpesvirus. PMID- 10848832 TI - Resveratrol induces Fas signalling-independent apoptosis in THP-1 human monocytic leukaemia cells. AB - Resveratrol, a natural product present in wine, has recently been shown to inhibit the growth of a number of cancer cell lines in vitro. In the current study, we have demonstrated that resveratrol inhibits the growth of THP-1 human monocytic leukaemia cells in a dose-dependent manner with a median effective dose of 12 microM. It did not induce differentiation of THP-1 cells and had no toxic effect on THP-1 cells that had been induced to differentiate into monocytes/macrophages by phorbol myristate acetate. A significant fraction of resveratrol-treated cells underwent apoptosis as judged by flow cytometric analysis of DNA content, DNA fragmentation and caspase-specific cleavage of poly(ADP-ribosyl) polymerase. Resveratrol treatment had no effect on the expression of Fas receptor or Fas ligand (FasL) in THP-1 cells, nor did it induce clustering of Fas receptors. In addition, THP-1 cells were resistant to activating anti-Fas antibody, and neutralizing anti-Fas and/or anti-FasL antibodies had no protective effect against resveratrol-induced inhibition of THP 1 cell growth. The effect of resveratrol on THP-1 cells was reversible after its removal from the culture medium. These results suggest that (1) resveratrol inhibits the growth of THP-1 cells, at least in part, by inducing apoptosis, (2) resveratrol-induced apoptosis of THP-1 cells is independent of the Fas/FasL signalling pathway and (3) resveratrol does not induce differentation of THP-1 cells and has no toxic effect on differentiated THP-1 cells. Thus, resveratrol may be a potential chemotherapeutic agent for the control of acute monocytic leukaemia. PMID- 10848834 TI - Lack of BCL10 mutations in Hodgkin's disease-derived cell lines. AB - The pathogenetic events leading to the malignant transformation of Hodgkin-Reed Sternberg cells are unknown. As Hodgkin-Reed-Sternberg cells are resistant to CD95-mediated apoptosis and chromosomal aberrations involving the 1p22 region harbouring the proapoptotic BCL10 gene represent a recurrent event in Hodgkin's disease-derived cell lines, analysis of the BCL10 gene and its transcripts was performed. As transcription of wild-type BCL10 was detected in all Hodgkin's disease-derived cell lines analysed, alterations of the coding sequence of the BCL10 gene are unlikely to contribute to the malignant transformation of the Hodgkin-Reed-Sternberg cell. PMID- 10848835 TI - Expression of the NUP98/HOXA9 fusion transcript in the blast crisis of Philadelphia chromosome-positive chronic myelogenous leukaemia with t(7;11)(p15;p15). AB - The t(7;11)(p15;p15) translocation is a recurrent aberration observed in acute myeloblastic leukaemia (AML) and chronic myelogenous leukaemia (CML). It has been shown that the NUP98 gene at 11p15 is fused with the HOXA9 gene at 7p15 in AML with t(7;11). We report the first case with CML expressing the NUP98/HOXA9 fusion transcript. A 27-year-old Japanese man was initially diagnosed as in the chronic phase of Philadelphia-positive CML. At the diagnosis of myeloid blast crisis, the karyotype evolved to 46, XY, t(7;11)(p15;p15), t(9;22)(q34;q11). Reverse transcriptase polymerase chain reaction identified the NUP98/HOXA9 transcript, suggesting that the NUP98/HOXA9 fusion protein could play a critical role in the progression to blast crisis. PMID- 10848836 TI - Asparagine synthetase activity in paediatric acute leukaemias: AML-M5 subtype shows lowest activity. AB - Lack of sufficient cellular activity of asparagine synthetase (AS) in blast cells compared with normal tissue is thought to be the basis of the antileukaemic effect of L-asparaginase in acute lymphoblastic leukaemia (ALL). Although L asparaginase is routinely used in ALL, its role and value in the treatment of acute myelogenous leukaemia (AML) is still being discussed. To evaluate the pharmacological basis for L-asparaginase treatment, we established pretreatment monitoring of the intracellular AS activity in blast cells of patients with AML and ALL. There was no general difference in AS activity between ALL and AML samples. Significantly lower AS activity, however, was found in the B-lineage ALL subgroups as well as AML-M5. PMID- 10848837 TI - Simultaneous detection and quantification of minimal residual disease in childhood acute lymphoblastic leukaemia using real-time polymerase chain reaction. AB - A number of prospective studies have indicated the clinical utility of measuring minimal residual disease (MRD) in childhood acute lymphoblastic leukaemia (ALL) and have highlighted the need for improved methodology for quantification of residual disease. We describe a novel real-time polymerase chain reaction (PCR) strategy for MRD analysis based on the exonuclease activity of Taq polymerase to cleave a fluorescently labelled probe. Using a consensus probe designed to the framework 2 region of the IgH gene, together with leukaemia-specific primers, the utility of this technique for simultaneous detection and quantification of MRD was demonstrated in samples from six ALL patients. This technique provides a rapid quantitative assay for determining MRD levels which lends itself to the routine monitoring of minimal residual leukaemia. PMID- 10848838 TI - Generation of HLA-DRB1*1501-restricted p190 minor bcr-abl (e1a2)-specific CD4+ T lymphocytes. AB - A small population of cells in acute lymphoblastic leukaemia is characterized by a specific translocation of the c-abl oncogene on chromosome 9 to the break point cluster lesion (bcr) on chromosome 22, t(9; 22)(q34; q11) (e1a2). Theoretically, the junction-spanning sequences of oncogene fusion proteins might be ideal targets for immunotherapy because these are not present in normal cells. In this study, we show for the first time that in vitro immunization with a 17-mer e1a2 peptide representing the p190 minor bcr-abl fusion protein resulted in HLA DRB1*1501-restricted peptide-specific proliferative CD4+ T lymphocytes, using peptide-pulsed monocyte-derived dendritic cells as the antigen-presenting cells. PMID- 10848839 TI - Remission status defined by immunofixation vs. electrophoresis after autologous transplantation has a major impact on the outcome of multiple myeloma patients. AB - We have retrospectively analysed 344 multiple myeloma (MM) patients (202 de novo patients) treated in a non-uniform way in whom high-dose therapy and autologous stem cell transplantation (ASCT) response was simultaneously measured by both electrophoresis (EP) and immunofixation (IF). Patients in complete remission (CR) by EP were further subclassified as CR1 when IF was negative and CR2 when it remained positive. Partial responders (PR) were also subclassified as PR1 (very good PR, > 90% reduction in M-component) or PR2 (50-90% reduction). CR1 patients showed a significantly better event-free survival (EFS) [35% at 5 years, 95% confidence interval (CI) 17-53, median 46 months] and overall survival (OS) (72% at 5 years, CI 57-86, median not reached) compared with any other response group (univariate comparison P < 0.00000 to P = 0. 004). In contrast, comparison of CR2 with PR1 and with PR2 did not define different prognostic subgroups (median EFS 30, 30 and 26 months respectively, P = 0.6; median survival 56, 44 and 42 months respectively, P = 0.5). The non-responding patients had the worst outcome (5-year OS 8%, median 7 months). Multivariate analysis confirmed both the absence of differences among CR2, PR1 and PR2 and the highly discriminatory prognostic capacity of a three-category classification: (i) CR1 (ii) CR2 + PR1 + PR2, and (iii) non-response (EFS P < 0.00000; OS P < 0.00000; both Cox models P < 0.00000). In the logistic regression analysis, the factors significantly associated with failure to achieve CR1 were the use of two or more up-front chemotherapy lines, status of non-response pre-ASCT and inclusion of total body irradiation (TBI) in the preparative regimen. Tandem transplants or the use of multiple agents (busulphan and melphalan) in the preparative regimen resulted in a higher CR1 level; none of the biological factors explored influenced the possibility of achieving CR1. These results confirm that, in MM patients undergoing ASCT, achieving a negative IF identifies the patient subset with the best prognosis; accordingly, therapeutic strategies should be specifically designed to achieve negative IF. PMID- 10848840 TI - Increased expression of Fas (APO-1, CD95) on CD34+ haematopoietic progenitor cells after allogeneic bone marrow transplantation. AB - Up-regulation of Fas/APO-1 (CD95) on haematopoietic progenitors and Fas-mediated apoptosis have been suggested to occur in a possible pathological mechanism in some bone marrow failure syndromes. We examined the expression of Fas antigen and susceptibility to Fas-mediated suppression of donor-derived haematopoietic cells of allogeneic bone marrow transplantation (BMT) recipients. Cytofluorometric analysis revealed low expression of Fas on CD34+ bone marrow cells from marrow donors or healthy controls. However, significantly higher expression of Fas antigen was observed on CD34+ bone marrow cells of BMT recipients, in whom engraftment of donor bone marrow (BM) cells was confirmed. The addition of an agonistic anti-Fas antibody (Ab) (CH-11) to haematopoietic stem cell culture of BM cells more strongly suppressed colony formation from granulocyte-macrophage colony-forming units (GM-CFU) and erythroid burst-forming units (BFU-E) after BMT. Pretreatment by blocking anti-Fas Ab (ZB4) abrogated the Fas-mediated GM-CFU and BFU-E suppression. Purified marrow CD34+ cells from BMT recipients were also susceptible to the Fas-mediated colony suppression. Thus, donor-derived CD34+ haematopoietic cells increased their expression of Fas antigen and were susceptible to Fas-mediated haematopoietic suppression. These findings provide new insight for understanding the haematological condition after BMT. PMID- 10848841 TI - Association between plasma viscosity and all-cause mortality: results from the MONICA-Augsburg Cohort Study 1984-92. AB - Several studies have reported a strong association between various markers of the acute-phase response and death from cardiovascular diseases and all-cause mortality. Inflammation at a low level of intensity may be a common phenomenon associated with the majority of causes of death owing to chronic diseases. We sought to investigate the association of plasma viscosity with all-cause mortality in a cohort of apparently healthy men. The study population consisted of 964 men aged 45-64 years at entry, randomly selected from the general population and taking part in the first MONICA-Augsburg survey 1984-85. The main outcome measure was all-cause mortality. During 8 years of follow-up, there were 81 deaths (37 cardiovascular deaths, 23 deaths from cancer and 21 deaths from other causes). There was a strong positive and statistically significant age adjusted relationship between plasma viscosity and all-cause mortality. The relative risk of death for a one standard deviation increase in plasma viscosity (0.070 mPa/s) was 1.45 [95% confidence interval (CI) 1.19-1.76]. After further adjusting for smoking, total cholesterol, body mass index, blood pressure and education, a relative risk of 1.41 (95% CI 1.14-1.74) resulted. Other risk variables had only negligible confounding effects. The relative risk of the median of the top quintile of the plasma viscosity distribution compared with the median of the bottom quintile, computed from the adjusted model, was 2.68 (95% CI 1.63-4.42). These findings suggest that plasma viscosity may have considerable potential to predict death from all causes in middle-aged men. PMID- 10848842 TI - Long-term survival of infants with idiopathic myelofibrosis. AB - Idiopathic myelofibrosis can develop in children as well as adults. However, the disease appears to be much more aggressive in adults, being characterized by poor survival rates and a high frequency of malignant transformation. Here, we describe three cases of idiopathic myelofibrosis in infants, two of whom were followed for 16 and 22 years after diagnosis. Neither of these patients required more than minimal supportive care, and both have had spontaneous erythropoietic recovery as early as 2-3 years after diagnosis. There have been no indications of malignant transformation or clinical deterioration. Thus, idiopathic myelofibrosis may have a different pathogenesis and clinical course in infants from adults, requiring a more conservative approach to management. PMID- 10848843 TI - Slowing the heart saves lives: advantages of perioperative beta-blockade. PMID- 10848844 TI - SURGICAL PUBLISHING: writing the manuscript PMID- 10848845 TI - COPE: committee on publication ethics PMID- 10848846 TI - Problematic diagnosis of a breast lesion. PMID- 10848847 TI - Evolution of nutritional support in acute pancreatitis. AB - BACKGROUND: Acute pancreatitis is a catabolic illness and patients with the severe form have high metabolic and nutrient demands. Artificial nutritional support should therefore be a logical component of treatment. This review examines the evidence in favour of initiating nutritional support in these patients and the effects of such support on the course of the disease. METHODS: Medline and Science Citation Index searches were performed to locate English language publications on nutritional support in acute pancreatitis in the 25 years preceding December 1999. Manual cross-referencing was also carried out. Letters, editorials, older review articles and most case reports were excluded. RESULTS AND CONCLUSION: There is no evidence that nutritional support in acute pancreatitis affects the underlying disease process, but it may prevent the associated undernutrition and starvation, supporting the patient while the disease continues and until normal and sufficient eating can be resumed. The safety and feasibility of enteral nutrition in acute pancreatitis have been established; enteral nutrition may even be superior to parenteral nutrition. Some patients, however, cannot tolerate enteral feeding and this route may not be practical in others. Parenteral nutrition still has a role, either on its own or in combination with the oral and enteral routes, depending on the stage of the illness and the clinical situation. PMID- 10848849 TI - VASCULAR SURGICAL SOCIETY: vascular surgical society of great britain and ireland PMID- 10848848 TI - Fournier's gangrene: a review of 1726 cases. AB - BACKGROUND: Although there is much consensus, certain controversies exist regarding the management of Fournier's gangrene. METHOD: Publications in English on Fournier's gangrene from January 1950 to September 1999 were obtained through the Medline database and relevant reference lists in publications. It was possible to identify 1726 cases for study. Data extracted for review included country of reported cases, number of patients in each report and relevant clinical features. RESULTS: Fournier's gangrene occurs worldwide. However, its definition has generated considerable controversy as efforts are made to refine the original description in the light of increasingly understood aetiological factors. Attempts to classify the disease into primary and secondary forms have not been successful. The basic pathological process, necrotizing fasciitis, has been identified in the perineum of women and children, although the disease afflicts the male more often than the female. Most reported cases have occurred in the USA and Canada. The major sources of sepsis are the local skin, colon, anus and rectum, and the lower urinary tract. Colonic, anal and rectal sources carry the worst prognosis. Diabetes mellitus is important in aetiological terms. Rare causes include vasectomy and circumcision. Investigations are essential to define the cause of an episode but not for the diagnosis of the disease. Early aggressive treatment of Fournier's gangrene and underlying conditions is essential. Hyperbaric oxygen and honey are treatment modalities yet to be universally adopted. Risk of death, 16 per cent overall in this series, is related to the patient's condition at presentation. CONCLUSION: Controversies over the definition of Fournier's gangrene persist but these do not affect the treatment options. The diagnosis is made on clinical grounds. The occurrence of the disease in women is under-reported and may go unrecognized by some clinicians. Some treatment options, such as hyperbaric oxygenation and radical excision, remain controversial. PMID- 10848850 TI - Reperfusion injury is greater with delayed restoration of venous outflow in concurrent arterial and venous limb injury. AB - BACKGROUND: Complex limb trauma often involves combined arterial and venous injury, and the resultant ischaemia-reperfusion injury (IRI) causes both local and remote organ injury. This study assessed the influence of the timing of restoration of venous drainage on IRI. METHODS: Male New Zealand white rabbits (n = 36) were randomized into six groups: sham operation (group 1) and unilateral hind limb arterial and venous occlusion for 1 h followed by no reflow for 2 h (group 2), arterial and venous reflow for 2 h (group 3), arterial reflow alone for 2 h (group 4), arterial reflow alone for 1 h followed by arterial and venous (delayed) reflow for a further 1 h (group 5), and pretreatment with an enteral combination antioxidant before occlusion of both artery and vein and delayed venous reflow (group 6). Plasma hydroperoxide (HPO) and glutathione peroxidase concentration, hind limb skeletal muscle and lung tissue wet : dry weight ratios and myeloperoxidase (MPO) concentration were measured. RESULTS: The plasma HPO level in the femoral vein effluent was significantly greater after delayed venous reflow (mean(s.e.m.) 2. 02(0.54) micromol/l) than in control animals (0.98(0.10) micromol/l) (P < 0.05). There was also a significantly greater tissue wet : dry weight ratio after delayed venous reflow than in controls, in skeletal muscle (mean(s.e.m.) 6.89(0.14) versus 5.34(0.54); P < 0. 05) and lung (9.20(1.14) versus 7.23(0.38); P < 0.05) tissue. Lung tissue MPO activity was significantly greater after delayed venous reflow compared with controls (3.20(0.28) versus 1.86(0.14) units/g; P < 0.005), and also in comparison to simultaneous arterial and venous reflow (2.40(0.24) units/g; P < 0.05). In the antioxidant pretreatment group there was no significant increase in plasma HPO concentration, tissue MPO level or tissue wet : dry weight ratio compared with the control group. CONCLUSION: In combined major arterial and venous injury of the limb, delayed restoration of venous drainage leads to significantly greater local skeletal muscle injury and remote neutrophil-mediated lung injury. These results support the clinical rationale for early restoration not only of arterial inflow but also venous drainage by means of intraluminal shunts. PMID- 10848851 TI - Risk factors for postoperative death following elective surgical repair of abdominal aortic aneurysm: results from the UK Small Aneurysm Trial. On behalf of the UK Small Aneurysm Trial participants. AB - BACKGROUND: In regional and population studies, the mortality rate within 30 days of elective surgical repair of abdominal aortic aneurysm is approximately 8 per cent. Identification of preoperative factors associated with this mortality risk is important for informing surgical policy and may suggest suitable preoperative interventions. METHODS: In the UK Small Aneurysm Trial, 820 patients aged 60-80 years underwent elective open surgical repair of an abdominal aortic aneurysm. The relationship between 30-day mortality rate and 13 prespecified potential prognostic factors was investigated. The value of a published clinical prediction rule was also evaluated. RESULTS: The postoperative mortality rate was 5.6 per cent overall (46 deaths in 820 patients). Postoperative mortality risk was significantly associated with older age (P = 0. 03), higher serum creatinine level (P = 0.002) and lower forced expiratory volume in 1 s (FEV1) (P = 0.003) in univariate analyses. Evidence of a relationship between age and postoperative death was weakened (P = 0.08) after adjustment for creatinine level and FEV1. The predicted postoperative mortality risk ranged from 2.7 per cent in younger patients with below average creatinine levels and above average FEV1, to 7.8 per cent in older patients with above average creatinine levels and below average FEV1. The published clinical prediction rule did not validate well on these data; observed risk did not correlate with predicted risk except for a small group of high-risk patients. CONCLUSION: Poor preoperative lung and renal function was strongly associated with postoperative death. Age was less important once these two important prognostic factors had been taken into account. The potential for preoperative improvement in lung and renal function to reduce postoperative mortality rates should be addressed in future studies. PMID- 10848852 TI - Population screening reduces mortality rate from aortic aneurysm in men. AB - BACKGROUND: Rupture of an unsuspected abdominal aortic aneurysm is a major cause of death in men over the age of 65 years. A significant reduction in deaths is likely to result only from higher rates of detection and increased numbers of elective aneurysm repairs. Screening of men reaching the age of 65 years has been taking place in the county of Gloucestershire, UK since 1990 and the aim of this study was to investigate any change in the mortality rate from aortic aneurysm in the screened portion of the population. METHODS: Total number of deaths from all aortic aneurysm-related causes in the county's population was calculated from hospital and post-mortem records, together with computerized death certificate records, for the years 1994-1998. The overall number of aneurysm-related deaths in men aged 65-73 years, who have been progressively influenced by the screening programme, was compared with that for men of all other ages. RESULTS: The total number of aneurysm-related deaths in men aged 65-73 years decreased progressively year by year between 1994 and 1998; this reduction is highly statistically significant (P < 0. 001). No such change was observed in the unscreened part of the population. CONCLUSION: Screening for asymptomatic abdominal aortic aneurysm results in a significant reduction in numbers of deaths from all aneurysm-related causes in the screened portion of the male population. PMID- 10848853 TI - Randomized, multicentre, double-blind, placebo-controlled trial of the use of aprotinin in the repair of ruptured abdominal aortic aneurysm. On behalf of the Joint Vascular Research Group. AB - BACKGROUND: The use of aprotinin in cardiac surgery reduces blood transfusion requirements. The aim of this trial was to see whether the same benefit applies in the repair of ruptured abdominal aortic aneurysm (AAA). METHODS: In this prospective, randomized trial, nine centres with local ethics committee approval recruited 77 patients with a ruptured AAA. A bolus of aprotinin 2 x 106 units, followed by an infusion of 0.5 x 106 units every 30 min, was administered to 38 patients, and 39 received a placebo infusion. The quantity of blood products transfused during surgery and in the first 12 h after operation was noted, along with the incidence of complications, mortality rates and length of hospital stay. RESULTS: Seventeen of the 38 patients who received aprotinin and 17 of the 39 given placebo died within 30 days (overall mortality rate 44 per cent). The median amount of blood given to the aprotinin group after operation was 1 (range 0-14) unit, while for the placebo group it was 3 (range 0-13) units (P = 0.02). However, the difference in the total number of units of blood transfused did not reach significance (10 (range 2-29) versus 14 (range 4-38) units respectively). CONCLUSION: The use of high-dose aprotinin during the repair of a ruptured AAA reduced blood transfusion requirements in the first 12 h after operation, but had no significant effect on the overall blood transfusion requirement. PMID- 10848854 TI - Two-year results of a randomized controlled trial of rifampicin-bonded extra anatomic dacron grafts. PMID- 10848855 TI - Apolipoprotein E genotype is associated with differential expansion rates of small abdominal aortic aneurysms. AB - BACKGROUND: The common polymorphism of the apolipoprotein E (APOE) gene is associated with differential risk of atherosclerosis; the gene could be a candidate gene in abdominal aortic aneurysms (AAA). METHODS: APOE genotypes were determined in 57 men aged 65-73 years with a small AAA (30-50 mm). The patients were included in a population ultrasonographic screening programme and were followed with at least two examinations during an interval of 2-4.5 years. The AAA expansion rates in patients with four different APOE genotypes were studied, with adjustment for initial AAA size and smoking. RESULTS: APOE genotype was a significant determinant of AAA expansion rate (P = 0.001). The adjusted mean (95 per cent confidence interval) rate was 2.1 (1.7-2.6) mm/year in 31 men with genotype E3E3, 1.3 (0.7-1.9) mm/year in 17 men with E3E4, 3.1 (2.0-4. 1) mm/year in six men with E2E3 and 4.2 (2.7-5.6) mm/year in three men with E2E4. The mean expansion rate was 2.2 (1.5-2.8) mm/year in non-smokers and 3.0 (2.5-3.6) mm/year in smokers (P = 0.014). CONCLUSION: APOE genotype seems to influence AAA expansion rate, but the effects of the individual genotypes, in particular E3E3 and E3E4, are contradictory when compared with the effects of the genotypes on risk of atherosclerosis. PMID- 10848856 TI - Clinical governance and the vascular surgeon. AB - BACKGROUND: Audit of adverse outcome might allow identification of substandard surgical results. To test this hypothesis statistical modelling was applied to two indicator vascular procedures (elective abdominal aortic aneurysm repair and carotid endarterectomy) with accepted adverse event rates. METHODS: Binomial statistical models for varying adverse event rates were constructed. A power calculation was used in an attempt to predict the case numbers required to determine substandard results for individual surgeons and vascular units. Two scenarios were considered: first a base adverse event rate of 6 per cent and surgical practice with 9, 12 and 24 per cent morbidity rates, and second a base adverse event rate of 3 per cent and surgical practice with 6, 9 and 12 per cent morbidity rates. RESULTS: A mean of 57 elective abdominal aortic aneurysm repairs and 70 carotid endarterectomies were performed per annum. The adverse event rate for both operations was 4 per cent. Power calculations revealed that 130 patients would need to be studied to detect a surgeon with an adverse event rate twice 6 per cent and over 280 patients would be required with an adverse event rate twice 3 per cent. To gather this number of patients 2 years of unit data and between 3 and 22 years of individual data would need to be studied for a base adverse event rate of 6 per cent. A base rate of 3 per cent requires 7-47 years for an individual and 4-65 years for the unit. With a base adverse event rate of 6 per cent, detection of widely variant surgical practice (four times the morbidity rate as base) requires only 21 procedures. CONCLUSION: Statistical modelling demands assumptions about accepted adverse event rates, confidence criteria and what constitutes substandard results. Data from large numbers of patients are required even for common operations with accepted adverse event rates. These data raise serious questions as to the feasibility of performing clinical governance on the basis of morbidity and mortality event rates alone. PMID- 10848857 TI - Use of superficial femoropopliteal vein for suprainguinal arterial reconstruction in the presence of infection. AB - BACKGROUND: Conventional treatment of mycotic aneurysms or graft infections of the aortoiliac segment by in situ or extra-anatomic prosthetic reconstruction has a high mortality and morbidity rate, with a substantial risk of persistent graft infection. The use of autologous vein may reduce this. METHODS: Eleven patients with suprainguinal arterial infections including two with mycotic aortic aneurysms, four with aortic graft infections, four infected femorofemoral grafts and an infected axillofemoral graft were treated by debridement and in situ reconstruction with autologous superficial femoropopliteal vein. All patients received appropriate antibiotic therapy and were followed by regular postoperative duplex imaging. Preoperative femoral vein duplex imaging was performed in eight of the 11 patients. RESULTS: Ten of the 11 patients survived with a functioning graft and without limb loss or evidence of infection at 4-33 months. One patient died from myocardial infarction after operation. Three patients had minor swelling of one leg. Four patients required subsequent angioplasty of anastomotic stenoses detected by duplex surveillance. CONCLUSION: Superficial femoropopliteal vein is an excellent conduit for suprainguinal reconstruction in the presence of infection. Duplex imaging is useful for confirming the suitability of deep veins for use as a graft and for postoperative surveillance. PMID- 10848858 TI - Randomized clinical trial of the effect of needle gauge and local anaesthetic on the pain of breast fine-needle aspiration cytology. AB - BACKGROUND: Breast fine-needle aspiration cytology (FNAC) is an invasive investigation which can be uncomfortable or distressing. This randomized study investigated the discomfort of breast FNAC and the effect of different techniques. METHODS: Some 116 FNAC samples were taken from 98 women with a palpable breast mass. Each patient was randomized to one of four study groups; aspiration was performed using a green-hub (21 G) or blue-hub (23 G) needle, either with or without local anaesthetic. Each patient scored the pain of the whole procedure using a visual analogue scale. RESULTS: A green-hub needle caused significantly more discomfort (mean(s.e.m.) pain score 5.1(0. 4) cm) than a blue hub needle (2.9(0.4) cm), or either a blue- or green-hub needle with local anaesthetic (3.0(0.4) and 2.1(0.4) cm respectively) (F = 10.28, 3112 d.f., P < 0.01, analysis of variance). CONCLUSION: The discomfort of breast FNAC is dependent upon the gauge of the needle and the use of local anaesthetic. A blue hub needle without local anaesthetic should be first choice for breast FNAC. PMID- 10848859 TI - Long-term results of a randomized clinical trial between laparoscopic hernioplasty and shouldice repair. AB - BACKGROUND: At present only short-term follow-up data are available to compare endoscopic and conventional hernia surgery. This paper presents data from a randomized study 6 years after initial recruitment. METHODS: In 1993 a randomized comparative study of transabdominal preperitoneal (TAPP) and Shouldice repair was commenced. Endpoints were rate of recurrence, late complications, complaints and patient satisfaction. RESULTS: The rate of recurrence in the TAPP group was one (2 per cent) of 48 patients and in the Shouldice group two (5 per cent) of 43. Only five patients in the Shouldice and three in the TAPP group reported slight discomfort in the inguinal region at 6-year follow-up. In neither group was chronic pain syndrome observed. Altogether, 46 (96 per cent) of 48 patients in the TAPP group and 35 (81 per cent) of 43 of those having the Shouldice procedure stated complete satisfaction with the hernia repair. CONCLUSION: Long-term evaluation demonstrated greater satisfaction with the result of the repair in the endoscopic group. The difference between the groups in the recurrence rate was not significant, because of the small numbers. The TAPP method appears to be an effective surgical alternative in patients with inguinal hernia. PMID- 10848860 TI - Beneficial effects of arterialization of the portal vein on extended hepatectomy. AB - BACKGROUND: Extended hepatectomy may result in postoperative liver failure. The aim of this study was to evaluate the effects of arterialization of the portal vein on oxygen supply, hepatic energy metabolism and liver regeneration after extended hepatectomy. METHODS: Portal haemodynamics were evaluated 0 or 10 days after arterialization of the portal vein in three experimental groups: 85 per cent partial hepatectomy, 85 per cent partial hepatectomy 10 days after arterialization of the portal vein and 85 per cent partial hepatectomy 10 days after ligation of the hepatic artery. Survival rates, weight of the regenerating liver, levels of adenine nucleotides and hepatic energy charge were assessed. RESULTS: Arterialization of the portal vein caused a significant increase in partial pressure of oxygen and oxygen saturation. Portal blood flow 10 days after arterialization was significantly increased. Survival rate and weight of the regenerating liver in the group with arterialization of the portal vein were significantly higher than those in the other two groups. The group with arterialization of the portal vein showed the highest levels of adenosine 5' triphosphate. CONCLUSION: The increase in portal blood flow and oxygen supply produced by arterialization of the portal vein has beneficial effects on hepatic energy metabolism and liver regeneration, and leads to improved survival after experimental extended hepatectomy. PMID- 10848861 TI - Experimental study of a novel fibrin sealant for achieving haemostasis following partial hepatectomy. AB - BACKGROUND: Ensuring adequate haemostasis is a major difficulty in the field of liver surgery. This study aimed to evaluate a novel fibrin sealant (Vivostat), designed for autologous preparation, in a porcine model of partial hepatectomy. METHODS: Thirty-six Large White Landrace pigs underwent partial left hepatectomy by finger fracture under portal vascular inflow occlusion. Animals were randomized to treatment of the resected surface with either fibrin sealant (Vivostat, n = 12) or regenerated oxidized cellulose gauze (Surgicel, n = 12), or were left untreated (controls, n = 12). Blood loss from the resection margin was measured at 2-min intervals for 10 min, and the time to haemostasis was recorded. Following complete haemostasis the animals recovered for 7 days. RESULTS: Median (range) blood loss in the control group was 94.3 (17.3-467.0) g, and was significantly reduced with Vivostat (13.8 (5.5-150.9) g) and Surgicel (22.8 (5.8 67.3) g). Median (range) time to haemostasis in controls (31 (12-52) min) was also significantly reduced by Vivostat (8 (0-32) min) and Surgicel (10 (0-19) min) (both P < 0.001 versus controls, Kruskal-Wallis test). CONCLUSION: The novel fibrin sealant, Vivostat, is as effective as Surgicel cellulose gauze in achieving haemostasis after porcine partial hepatic lobectomy. PMID- 10848862 TI - Functional outcome after restorative panproctocolectomy for ulcerative colitis decreases an otherwise enhanced quality of life. AB - BACKGROUND: Restorative panproctocolectomy is a favoured operation for ulcerative colitis, but altered bowel habit may adversely affect overall quality of life. METHODS: Specific and generic quality of life questionnaires and an instrument to award money for continuing disability based on government guidelines were sent to 103 patients who had curative surgery for ulcerative colitis between 1995 and 1997. Seventy-one patients returned completed questionnaires: 30 with an ileostomy (representing incontinence and abnormal body image), 11 with a Koch pouch (representing continence and abnormal body image) and 30 with a pelvic pouch (representing continence and normal body image). RESULTS: Patients valued the disability of having an ileostomy similar to that for a Koch pouch or a pelvic pouch: pound 40 000, pound 30 000 and pound 40 000 respectively (P = 0. 97). There was no sex difference. Body image measured with a visual analogue scale (least = 1, worst = 10) was worst with the ileostomy and Koch pouch (8 each) and best with a pelvic pouch (5) (P = 0.06). However, pelvic pouches scored significantly worse than an ileostomy with regard to altered bowel emptying (pelvic pouch, 8; Koch pouch, 7; ileostomy, 5) (P = 0.01). CONCLUSION: Poor function after pelvic pouch surgery offsets any advantage in body image over an ileostomy. Thus, overall quality of life and perceived monetary damage were the same for the two operations. Improved pelvic pouch function is likely to be reflected in better quality of life after restorative panproctocolectomy. PMID- 10848863 TI - Immunoglobulin G and albumin levels in whole gut lavage fluid provide an objective measure of pouch ileitis. AB - BACKGROUND: Gut protein loss is a characteristic of inflammatory bowel disease (IBD), and immunoglobulin (Ig) G, albumin and alpha1-antitrypsin concentrations in whole gut lavage fluid (WGLF) correlate with clinical disease activity. If inflammation in ileoanal pouches is similar to IBD, then measurement of protein losing enteropathy by analysis of WGLF may provide an objective assessment of disease activity in pouches. METHODS: Forty-two patients who had restorative proctocolectomy for ulcerative colitis underwent whole gut lavage with a polyethylene glycol-electrolyte solution. The first clear effluent was filtered, processed by the addition of protease inhibitors and stored at - 70 degrees C. IgG, albumin and alpha1-antitrypsin were assayed in WGLF. The Pouchitis Disease Activity Index (PDAI) was calculated after pouchoscopy and biopsy; the Moskowitz criteria for pouchitis were also applied. RESULTS: There was a significant correlation of the pouchoscopy score and the PDAI with the concentration of WGLF IgG. All patients with 'pouchitis' according to the Moskowitz criteria had a WGLF IgG concentration greater than 10 microg/ml. The WGLF albumin level also showed a significant correlation with the PDAI, but alpha1-antitrypsin concentration did not. CONCLUSION: Analysis of WGLF for IgG and albumin may be useful in the assessment of disease activity in pouch inflammation. PMID- 10848864 TI - On-line quantitative analysis of surface electromyography of the pelvic floor in patients with faecal incontinence. AB - BACKGROUND: Needle electromyography (EMG) remains the 'gold standard' for the assessment of external anal sphincter innervation. It is, however, an invasive and poorly tolerated technique. In this study a quantitative form of surface electromyography was compared with needle EMG of the external anal sphincter. METHODS: Invasive needle EMG to assess mean fibre density and neuromuscular jitter was compared directly with quantitative surface EMG in 37 patients with faecal incontinence and 12 age-matched controls. RESULTS: There was a significant positive correlation between mean fibre density on needle EMG and maximum turns rate on surface EMG (rs = 0.48 (95 per cent confidence interval 0.28-0.76), P = 0.003). Furthermore, surface EMG was able to discriminate between patients with normal neuromuscular jitter and those with increased jitter, a measure of progressive denervation and reinnervation, on the basis of reduced rectified mean surface signal (P = 0.02, Fisher's exact test). CONCLUSION: Quantitative surface EMG may potentially replace invasive needle EMG as the investigation of choice in the assessment of anal sphincter electrophysiology. PMID- 10848865 TI - Value of herniography in the management of occult hernia and chronic groin pain in adults. PMID- 10848866 TI - Prospective study of safety, patient satisfaction and leg ulcer healing following saphenous and subfascial endoscopic perforator surgery. PMID- 10848867 TI - Author's reply PMID- 10848868 TI - Outcome measures after lower extremity bypass surgery: there is more than just patency. PMID- 10848870 TI - The combined pituitary function test is not indicated in the routine investigation of short stature. PMID- 10848869 TI - Hyponatraemia. PMID- 10848871 TI - Inhibitory action on GHRH-induced GH secretion of chronic tamoxifen treatment in breast cancer. AB - OBJECTIVE: Previous in vitro and in vivo studies on animal models have demonstrated that tamoxifen (TAM) inhibits GH secretion. Studies in humans are conflicting. The aim of this study was to evaluate the effect of chronic TAM treatment on GH secretory dynamics in the presence of negligible endogenous oestrogens, in postmenopausal women with breast cancer. PATIENTS: Ten female patients were studied over a 6-12-month period after surgical therapy, before medical therapy, and during chronic treatment with TAM (20 mg/day p.o.). MEASUREMENTS: In all subjects we performed a standard GHRH-test (50 mg i.v. as a bolus) and compared the single time points, the peak response and the areas under the curves (AUC), before and during treatment. In basal samples, we evaluated the circulating levels of IGF-1, IGF-BP3 and their ratio, SHBG, FSH, LH, Oestradiol (E2) and PRL. GH was assayed by Immunoradiometric assay (IRMA). Insulin-like growth factor type I (IGF-I), Insulin-like growth factor-binding protein-3 (IGF BP3), FSH, LH and PRL were measured by Radioimmunoassay (RIA). SHBG was measured by a noncompetitive liquid phase immunoradiometric assay, while E2 was measured directly in plasma by a liquid phase technique. RESULTS: TAM chronic treatment significantly reduced GH response to GHRH at single time point evaluations, GH peak response (mean decrease: 59.8 +/- 7.3%) and GH-AUC (mean decrease 53.8 +/- 8.9%). TAM also significantly reduced plasma IGF-1 levels. No significant variations were found in IGF-BP3 levels or in the IGF-1/IGF-BP3 ratio. A significant inverse correlation between SHBG and IGF-1 circulating levels was noticed during TAM treatment. CONCLUSIONS: Our data show that long-term tamoxifen treatment blocks the response of GH to exogenous GHRH and reduces IGF-1 levels, possibly by a central mechanism other than the demonstrated peripheral action. The results of this study, keeping in mind the demonstrated mitogenic role of IGF 1 in cancer proliferation, can contribute to clarify the mechanism by which TAM exerts its antiproliferative effect. PMID- 10848872 TI - Genetic immunotherapy of established tumours with adenoviral vectors transducing murine interleukin-12 (mIL12) subunits in a rat medullary thyroid carcinoma model. AB - OBJECTIVE: Interleukin-12 (IL12) is a heterodimeric cytokine that plays an important role in the development of cellular immunity. Studies have demonstrated antitumour activity after systemic administration of recombinant IL12. As with other cytokines, with increasing dosage and longer exposure, systemic toxicity is observed. To reduce systemic toxicity and obtain local production of IL12, we developed a replication defective adenovirus transducing two subunits of the murine IL12 (AdCMVmIL12) gene. DESIGN: Two separate cassettes, expressing the p35 or p40 subunit of mIL12, under the control of human cytomeglavirus immediate early promoter, were inserted into the early1 (E1) region of adenovirus 5. Biological activity of virally expressed mIL12 was demonstrated in vitro through its ability to induce proliferation of mouse ConA blast cells. RESULTS: Rat medullary thyroid carcinoma (MTC) cells infected with AdCMVmIL12 lost their tumorigenicity in their syngenic WAG/Rij rat hosts. Efficient antitumour activity was found after direct injection of the AdCMVmIL12 vector into rMTC tumours. After intratumoural treatment with AdCMVmIL12, 86% of tumour bearing animals were apparently cured, and almost all remaining tumours were stabilized. Challenge studies showed that most animals cured after the first treatment remained tumour free after reinjection of wild type rMTC cells, indicating that long-term antitumour immunity developed. CONCLUSIONS: This study demonstrates the construction of an adenoviral vector expressing a functional heterodimeric mIL12 and its efficient antitumour activity after in vivo delivery in an animal model of medullary thyroid carcinoma. PMID- 10848873 TI - Stereotactic conformal radiotherapy for pituitary adenomas: technique and preliminary experience. AB - OBJECTIVE: Stereotactic conformal radiotherapy (SCRT) is a high precision technique of fractionated radiotherapy which ensures accurate delivery of radiation with reduction in the volume of normal tissue irradiated as compared to conventional external beam radiotherapy. We describe the technique and preliminary experience of SCRT in patients with residual and recurrent pituitary adenomas. PATIENTS AND METHODS: Between February 1995 and March 1999, 22 patients (mean age: 45.3, range: 20-67 years) with residual or recurrent pituitary adenomas (13 nonfunctioning, nine secretory) were treated with SCRT. All were immobilized in a relocatable Gill-Thomas-Cosman (GTC) frame and tumour was localized on a postcontrast planning computerized tomography (CT) and MRI scan. The gross tumour volume (GTV) and the critical structures were outlined on contiguous 2-3 mm separated slices. A margin of 5 mm (12 patients) to 10 mm (10 patients) was grown around GTV in three-dimensions (3-D) to generate the planning target volume (PTV). The treatment was delivered by three (five patients) and four (17 patients) maximally separated conformal fixed fields with each field conformed to the shape of the tumour using customized lead alloy blocks (19 patients) or multileaf collimator (three patients). The patients were treated on a 6-MV linear accelerator to a dose of 45 Gy in 25 fractions (18 patients) and 50 Gy in 30 fractions (four patients). RESULTS: The technique of SCRT has become a part of the routine work of the radiotherapy department. The treatment was well tolerated with minimal acute toxicity. One patient developed transient quadrantanopia 2 weeks after treatment with full recovery after a short course of corticosteroids. One patient had a transient visual deterioration 7 months after treatment due to cystic degeneration of the tumour which fully recovered following surgical decompression. Nine of the 15 patients presenting with visual impairment had improvement after treatment and the visual status remained stable in all others. One patient with acromegaly and one with a prolactinoma achieved normalization of elevated hormonal abnormality four and 10 months after SCRT, respectively. The remaining seven patients with a secretory adenoma had declining hormone levels at last follow-up. Newly initiated hormone replacement therapy was required in five patients. At a median follow-up of 9 months (range 1-44 months), the 1 and 2 year actuarial progression free and overall survival were 100%. CONCLUSION: Stereotactic conformal radiotherapy is a high precision technique suitable for the treatment of pituitary adenomas requiring radiotherapy. Preliminary results suggest effective tumour control and low toxicity within the range expected for conventional external beam radiotherapy. While the technique is of potential benefit in reducing the volume of normal brain irradiated, the advantages in terms of sustained tumour control and reduced toxicity over conventional radiotherapy need to be demonstrated in long-term prospective studies. PMID- 10848874 TI - Abnormal LH pulsatility in women with hyperprolactinaemic amenorrhoea normalizes after bromocriptine treatment: deconvolution-based assessment. AB - OBJECTIVE: The present study examines the LH secretory process in hyperprolactinaemic women before, during and after bromocriptine therapy, using restrictive clinical selection criteria as well as improved methodological tools. PATIENTS AND DESIGN: Six women (aged 20-40 years) with microprolactinomas (mean +/- SE prolactin, PRL: 2478 +/- 427 mU/l, range: 1370-3800 mU/l) and four age- and sex-matched healthy controls were admitted to the study. After an overnight fast, all patients and controls had blood samples withdrawn at 10 minute intervals for 6 h (during saline infusion) from 0800 h to 1400 h to determine serum LH and PRL concentrations. After baseline evaluation, patients were treated with bromocriptine, which was started at a daily dose of 1.25 mg for 7 days; the dose was then increased to 2.5 mg daily for the next 7 days and subsequently to 2.5 mg twice daily. PRL levels were evaluated at weekly intervals after the beginning of bromocriptine therapy for the duration of the study. The 6 h pulsatility study was repeated on four patients during treatment at a time when PRL levels were decreased, although not normalized (PRL range: 450-1350 mU/l) and, on four patients, with the attainment of normal serum PRL levels (PRL < 450 mU/l) in the early follicular phase of the menstrual cycle (days 2-5). The LH instantaneous secretion rate was reconstructed by a nonparametric deconvolution method. In addition to pulse analysis made using the program DETECT, the evaluation of the secretion rate yielded the pulse frequency as well as the pulse amplitude distribution. RESULTS: Each time series was submitted to deconvolution analysis using a nonparametric method in order to estimate the instantaneous secretion rate (ISR). Hyperprolactinaemic patients had very few high-amplitude LH pulses above 0.2 IU/(l minutes) before treatment (average frequency: 0.83 +/- 0.40 pulses/6 h) and at the intermediate evaluation (0.25 +/- 0.25 pulses/6 h). In both cases, the pulse frequency was significantly lower than in controls (P < 0.05 and P < 0.01, respectively). When PRL was normalized, the number of high amplitude LH pulses (4.25 +/- 1.03 pulses/6 h), became statistically different from the pulse number before (P < 0.01) and during (P < 0.01) therapy; in particular the pulse frequency after therapy rose to a level not statistically different from that in controls. CONCLUSION: The present study shows the presence of reduced LH pulsatility in hyperprolactinaemic women that recovers completely to within the physiological distribution when PRL levels are normalized by bromocriptine therapy. PMID- 10848875 TI - Effects of free fatty acids and acipimox, a lipolysis inhibitor, on the somatotroph responsiveness to GHRH in anorexia nervosa. AB - OBJECTIVE: Anorexia nervosa is characterized by low IGF-1 and high GH and free fatty acid (FFA) levels. As FFA exerts an inhibitory feedback action on GH secretion in physiological conditions, we hypothesized that somatotroph cells could be less sensitive to the negative feedback action of FFA in anorexia nervosa. PATIENTS: Fifteen patients with anorexia nervosa (AN, age: mean +/- SEM: 20.8 +/- 1.2 years, BMI: 15.9 +/- 0.3 kg/m2) and 12 normal female controls (NW, age 27.2 +/- 2.1 years, BMI 21.2 +/- 2.2 kg/m2). MEASUREMENTS: We studied the effects of lipid-heparin emulsion (Li-He, Intralipid 10% 250 ml + heparin 2500 U iv from -60 to + 90 minutes in seven AN and six NW) or acipimox (ACI, 250 mg p.o. at -60 minutes in eight AN and six NW), a lipolysis inhibitor, on the GH response to GHRH (1 microg/kg iv as a bolus at 0 minutes). RESULTS: Basal IGF-1 levels were lower (P < 0.05) while GH levels were higher (P < 0.05) in AN than in NW. On the other hand, basal FFA levels in the two groups were not significantly different. In both groups Li-He increased FFA levels (P < 0.05), which became higher (P < 0. 02) in AN than in NW. Li-He infusion inhibited (P < 0.05) basal GH levels in AN to levels overlapping those in NW. The GH response to GHRH in the whole AN group was higher than in NW (P < 0.03). Li-He inhibited the somatotroph responsiveness to GHRH in AN (P < 0.03) as well as in NW (P < 0.03) and during Li He the GH response to GHRH in AN became similar to that in NW. Whilst ACI pretreatment enhanced the GH response to GHRH in AN (P < 0.02), it did not significantly increase that in NW. Interestingly, after ACI administration, FFA levels were inhibited in both groups (P < 0.05) persisting higher in AN than in NW (P < 0.05). CONCLUSION: Though GH hypersecretion in anorexia nervosa occurs in presence of enhanced lipolysis, our present findings indicate that the sensitivity of somatotroph cells to the inhibitory feedback action of free fatty acid is preserved. PMID- 10848876 TI - Effect of gonadotrophin-releasing hormone (GnRH) antagonist during the LH surge in normal women and during controlled ovarian hyperstimulation. AB - OBJECTIVE: Several studies have suggested that GnRH is instrumental in triggering the LH surge. The present studies were performed to evaluate the effect of GnRH antagonist administration in women after the beginning of the LH surge. DESIGN: In study one, six normal cycling women were given a GnRH antagonist (20 mg Nal Glu s.c.) during an unstimulated cycle. Nal-Glu was administered when the LH level was higher than 10 U/l and associated with an oestradiol (E2) level higher than 730 pmol/l. In study two, a GnRH antagonist (3 mg Cetrorelix, ASTA-Medica, Frankfurt, Germany) was administered on day 8 of an IVF-ET cycle, in 157 women. Eighteen women among this cohort received the antagonist, when their LH level was higher than 10 U/l. RESULTS: In normal volunteers (study one), Nal-Glu was administered on day 13.7 +/- 1.4 (mean +/- SD) of the cycle when the LH level was 13.7 +/- 3.5 U/l with an E2 plasma level reaching 980 +/- 131 pmol/l. Twenty-four hours after administration of the antagonist, the LH surge had been interrupted in all subjects; it was postponed in three of the women, and abolished in the remaining three. LH levels fell by 68.5%, E2 by 42% and FSH by 53.2%. In study two, LH plasma levels 24 h after the antagonist administration fell by 94%. No premature ovulation occurred in any of the patients treated. Administering the antagonist before (n = 139) or during the LH surge (n = 18) made no statistically significant difference to the results of the IVF-ET attempt. CONCLUSIONS: Our results indicate that GnRH is required throughout the gonadotrophin surge in women, not only for the initiation but also for the maintenance of the LH surge. In addition, in our study, the suppression of the rise in LH, when the antagonist was given during the surge, had no detrimental impact on IVF-ET outcome. This suggests, if confirmed on a larger scale, that late follicular phase GnRH antagonist administration to prevent the LH surge in controlled ovarian hyperstimulation (COH) is a safe and useful treatment. PMID- 10848877 TI - The combined pituitary function test in children: an evaluation of the clinical usefulness of TRH and LHRH stimulation tests through a retrospective analysis of one hundred and twenty six cases. AB - OBJECTIVE: The combined pituitary function test is routinely used in the endocrine investigation of short children. The TRH and luteinising hormone releasing hormone (LHRH) response tests have been shown to be of minimal value in adults. We have evaluated the clinical utility of these tests in the context of combined pituitary function testing in children. DESIGN: A retrospective analysis of basal hormone measurements and pituitary stimulation tests in relation to clinical assessment of pituitary function. PATIENTS: One hundred and twenty-six children, 82 boys and 44 girls, aged 2-17 years, who had undergone pituitary function testing were studied. RESULTS: The TSH response to TRH stimulation correlated directly with basal plasma TSH but not basal plasma total T4. In patients with an impaired response to stimulation, basal TSH concentrations were <2.0 mIU/l and significantly lower than in patients with a normal response (P < 0.0001). An impaired response to TRH stimulation had a positive predictive value of 0.43 and a negative predictive value of 0.90 for the diagnosis of hypopituitarism. A basal TSH concentration of <2.0 mIU/l had a positive predictive value of 0.22 and a negative predictive value of 0.92. A low basal T4 (normal range 60-140 nmol/l) in combination with an inappropriately low or normal basal TSH was always associated with a diagnosis of hypopituitarism. The responses of plasma LH and FSH to LHRH stimulation correlated directly with basal plasma LH and FSH concentrations. Basal gonadotrophin concentrations, basal sex hormone concentrations or response to LHRH stimulation could not distinguish patients with constitutional delay of growth and puberty from those with hypopituitarism. There was no apparent relationship between either basal gonadotrophin concentrations or response to LHRH stimulation and clinical assessment of pituitary function. In patients > or =13 years with constitutional delay of growth and puberty the median and interquartile ranges of basal LH and FSH were 1.4 IU/l (0.7-3.6) and 2.6 IU/l (2.2-5.2) respectively. The three hypopituitary patients in this study with chronological age > or =13 years had undetectable concentrations of both gonadotrophins. The response of LH and FSH to LHRH stimulation was significantly lower in patients > or =13 years with clinical hypopituitarism than in those with intact pituitary function (P <0.02). CONCLUSION: TRH and LHRH tests in children with short stature appear to have little value over and above the baseline hormone measurements. An abnormal response to hormone stimulation is not diagnostic of hypothalamic-pituitary disease. We have demonstrated that neither TRH nor LHRH stimulation tests should be routinely used in the investigation of children with short stature. PMID- 10848878 TI - The use of insulin-like growth factor 1 reference values for the diagnosis of growth hormone deficiency in prepubertal children. AB - OBJECTIVE: This study was done to determine whether the use of reference values obtained in children with idiopathic short stature (ISS) improved the clinical value of serum insulin-like growth factor I (IGF-1) as a tool for diagnosing GH deficiency (GHD) in prepubertal children. PATIENTS AND METHODS: Serum IGF-1 was measured with a new IRMA kit (IGFI-RIA CT, Cis Bio, Gif sur Yvette, France) in 168 prepubertal normal children and in prepubertal children with ISS (n = 68), organic GHD due to a craniopharyngioma (oGHD, n = 15) and permanent idiopathic GHD (iGHD, n = 28). RESULTS: IGF-1 was lower (P < 0.001) in iGHD than in either ISS or oGHD and was below the fifth percentile of the normal range in 29/68 ISS (43%), 8/15 oGHD (53%) and 28/28 (100%) iGHD patients. Three oGHD (20%) and two iGHD (7%) patients had a serum IGF-1 below the fifth percentile of the normal group but above the fifth percentile of the ISS group. Thus, a serum IGF-1 below the fifth percentile of the normal group distinguished between normal children and iGHD with 100% sensitivity, between normal and oGHD with 53% sensitivity and between normal and all GHD (idiopathic + organic) with 84% sensitivity; the overall specificity was only 57%. Conversely, a serum IGF-1 below the fifth percentile of the ISS population distinguished between ISS and iGHD with 93% sensitivity, between ISS and oGHD with 33% sensitivity and between ISS and all GHD with 72% sensitivity; the overall specificity was then 95%. CONCLUSIONS: A serum IGF-1 within the normal range virtually excludes idiopathic GHD but does not rule out organic GHD, whereas an IGF-1 below the ISS range is strongly in favour of GHD, after exclusion of poor nutritional status and/or liver disease. An IGF-1 below the normal range but in the idiopathic short stature range gives no definitive conclusion even when it is associated with a low GH peak. Thus, whereas reference values obtained in normal children must be used to interpret serum IGF-1 in short prepubertal children, reference data obtained in idiopathic short stature children should also be taken into account. PMID- 10848879 TI - Carbohydrate metabolism during growth hormone treatment and after discontinuation of growth hormone treatment in girls with Turner syndrome treated with once or twice daily growth hormone injections. AB - OBJECTIVE: To assess possible side-effects of treatment with supraphysiological GH dosages on carbohydrate (CH) metabolism in girls with Turner syndrome (TS) during GH treatment until adult height is reached as well as after discontinuation of GH treatment. DESIGN: In a prospective, randomized injection frequency-response study, the effect of GH treatment in combination with low dose ethinyl oestradiol on CH metabolism was evaluated, comparing twice daily (BID) with once daily (OD) injections of a total GH dose of 6 U/m2/day until adult height was reached. PATIENTS: Nineteen untreated girls with TS, mean (SD) pretreatment age 13.3 (1.7) (range 11.0-17.6) year. MEASUREMENTS: Glucose and insulin concentrations during oral glucose tolerance tests (OGTT) were measured before and during GH treatment, as well as at 6 months after discontinuation of GH treatment. RESULTS: GH treatment was discontinued after a mean of 43 (range 27 57) months. In one of the 19 girls, a different girl at each time point before, during and after discontinuation of GH treatment, the glucose response to OGTT after 120 minutes was above 7.8 mmol/l but below 11.1 mmol/l, indicating impaired glucose tolerance. None of the girls developed diabetes mellitus. Fasting glucose levels did not significantly change during, or after discontinuation of GH treatment. The 3 h area under the curve for time-concentration adjusted for fasting levels during the OGTT for glucose showed a significant decrease during GH treatment. In contrast to the glucose levels, GH treatment induced considerably higher insulin levels compared to pretreatment values. After discontinuation of GH insulin levels decreased to values comparable with pretreatment levels. None of these observed changes were different between the GH injection frequency groups. The changes in CH variables during and after discontinuation of GH were not related to changes in body mass index. CONCLUSIONS: GH treatment with 6 U/m2/day in combination with low dose ethinyl oestradiol in girls with Turner syndrome aged > or =11 years did not negatively influence glucose levels, but induced higher levels of insulin indicating relative insulin resistance. These changes in insulin levels were independent of the frequency of the GH injections (once vs. twice daily). After discontinuation of GH treatment, insulin values decreased to baseline levels. PMID- 10848880 TI - FHIT and TSG101 in thyroid tumours: aberrant transcripts reflect rare abnormal RNA processing events of uncertain pathogenetic or clinical significance. AB - OBJECTIVE: The chromosomal regions containing the two putative tumour suppressors, fragile histidine triad gene (FHIT) and tumour suppressor gene 101 (TSG101), are deleted frequently in thyroid tumours. We therefore analysed FHIT and TSG101 transcripts in a group of advanced thyroid tumours to establish their role in thyroid tumorigenesis. DESIGN: Retrospective analysis of FHIT and TSG101 mRNA transcripts and genomic DNA from cryo-preserved thyroid tumours. TP53, previously shown at the genomic level not to be mutated in this cohort of tumours, served as a control. PATIENTS: We analysed nine follicular thyroid carcinomas (FTC), six papillary thyroid carcinomas and six follicular adenomas (FA) and histologically normal thyroid tissue from four of the FA patients. MEASUREMENTS: Single stage and nested reverse transcription polymerase chain reaction (RT-PCR) products of FHIT, TSG101, and TP53 were analysed by agarose or polyacrylamide gel electrophoresis and sequenced. Genomic DNA was also analysed by polymerase chain reaction and sequencing (FHIT) or by Southern blotting (TSG101). Clinical data were correlated with the results of the mutation analysis. RESULTS: Truncated FHIT transcripts were observed frequently alongside full length transcripts with nested RT-PCR, most often in FTC, while single stage RT-PCR revealed only normal length transcripts in all tumours. Similar results were obtained for TP53, while abnormal TSG101 transcripts were detectable by single stage RT-PCR. Sequence analysis of the truncated FHIT and TSG101 transcripts revealed mainly exon skipping and alternate RNA processing events. Only a single point mutation (of TSG101) was found. Southern blotting for the TSG101 gene, and PCR amplification and sequencing of the FHIT gene showed no evidence of genomic abnormalities in either case, and there was no evidence of splice site mutations in the FHIT gene, suggesting that the truncated transcripts result from altered RNA processing. There was no relationship between tumour stage, grade or survival and the presence of FHIT or TSG101 abnormalities. CONCLUSIONS: Truncated FHIT and TSG101 transcripts in thyroid tumours reflect alternate mRNA splicing events, rather than genomic deletions. Such abnormal RNA processing seems to be common and widespread in thyroid neoplasms, as similar results were obtained by analysis of transcripts of TP53, which we had previously shown not to be mutated in these specimens. Although a pathogenetic role for these aberrant transcripts remains possible, no correlation was found with stage, histological grade or outcome in this small group of advanced thyroid malignancies. Relaxation of mRNA splice control appears to be a feature of follicular cell-derived thyroid neoplasms. PMID- 10848882 TI - Ablation of the thyroid remnant and 131I dose in differentiated thyroid cancer. AB - AIMS: To compare the efficacy of remnant ablation following a single low dose (specific activity of 131I administered, 1074-1110 MBq) vs. a single high dose (mostly 2775-3700 MBq) of 131I in patients with differentiated thyroid cancer and to determine whether or not the extent of surgery influences outcome. METHODS: Nineteen studies have reported the results of low dose 131I ablation. Of these, 11 met our criteria for a comparative analysis. Two additional cohorts of ours were added and these were analysed in two groups based on the extent of surgery (near-total [NT; Woodhouse1] vs. sub-total [ST; Woodhouse2]). There were 518 low dose and 449 high dose patients in all. RESULTS: The average failure of a single low dose was 46 +/- 28% (SD). Meta-analysis revealed a statistically significant advantage for a single high over a single low dose and a pooled reduction in relative risk of failure of the high dose of about 27% (P < 0.01). From this we estimate that for every seven patients treated one more would be ablated given a high rather than a low dose (assuming a low dose failure risk of 50%). Also, a significantly greater proportion of patients are ablated after a single high or low dose, if they underwent near-total as opposed to sub-total thyroidectomy (summary relative risk (RR) 1.4; P < 0.05). CONCLUSION: High dose 131I is more efficient than low dose for remnant ablation particularly after less than total thyroidectomy. Results suggest that patients with differentiated thyroid cancer should routinely have a total thyroidectomy followed by high dose 131I (2775 3700MBq) for ablation of the remnant. PMID- 10848881 TI - A polymorphism of the 5' flanking region of tumour necrosis factor alpha gene is associated with thyroid-associated ophthalmopathy in Japanese. AB - OBJECTIVE: We have studied the polymorphism of the 5' flanking region of the tumour necrosis factor (TNF)-alpha gene in order to better understand the genetic background of autoimmune thyroid disorders and thyroid-associated ophthalmopathy. PATIENTS AND METHODS: We studied the polymorphism of the 5' flanking region of the TNF-alpha gene at positions - 1031 (T to C change, termed as - 1031C), - 863 (C to A, - 863 A), - 857 (C to T, - 857T), - 308 (G to A, - 308 A) and - 238 (G to A, - 238 A) in Japanese patients with Graves' disease [n = 173, 62 of whom had associated ophthalmopathy (American Thyroid Association (ATA) class III or greater)] and healthy control subjects (n = 575), using a polymerase chain reaction sequence-specific oligonucleotide probe method. RESULTS: The allele frequency of - 857T in the Graves' disease patients (22. 5% vs. 17.7%, OR = 1.35, P = 0.045, corrected P = 0.23) was slightly greater than in the Japanese healthy subjects, respectively. However, the difference was not statistically significant. In Graves' disease patients with evident ophthalmopathy (ATA class III or greater), the allele frequencies of - 1031C and - 863 A were significantly greater than those with no or mild ophthalmopathy (ATA class 0-II) (31.5% vs. 13.5%, OR =2.94, P < 0.0001, corrected P < 0. 0005; 23.4% vs. 11.7%, OR =2.30, P = 0.0044, corrected P = 0.022, respectively) and in control subjects. The strength of the association of the polymorphism - 1031C increased with the severity of ophthalmopathy, with odds ratios of 2.36 for ATA class III, and 5. 43 for ATA class IV-VI, respectively, compared with Graves' disease with no or mild ophthalmopathy (ATA class 0-II). Although the phenotype frequency of DRB1*0901 was not different among Graves' disease patients with or without ophthalmopathy and control subjects, the phenotype frequency of DRB1*0901(-)/-1031C(+) was significantly increased in Graves' disease patients with ophthalmopathy compared to those with no or mild ophthalmopathy (OR = 4.91, P = 0.0005) or control subjects (OR = 4.59, P < 0.0001). CONCLUSIONS: These results suggest that the - 1031C or - 863 A alleles, or a gene in linkage disequilibrium with the TNF-alpha gene, predispose to the development of ophthalmopathy in Japanese patients with Graves' disease. PMID- 10848883 TI - Oral dehydroepiandrosterone (DHEA) replacement therapy in women with Addison's disease. AB - OBJECTIVE: Patients with primary adrenocortical failure (Addison's disease) have abnormally low levels of DHEA and androgens relative to age. To define a suitable dose, the effect of oral dehydroepiandrosterone (DHEA) replacement therapy in women with Addison's disease (n = 9) was evaluated. DESIGN AND MEASUREMENTS: DHEA was administered as a daily oral dose of either 50 mg (n = 5) or 200 mg (n = 4). Blood sampling and measurements of insulin sensitivity (as measured with euglycemic insulin clamp technique) and body composition (as measured by dual energy X-ray absorptiometry) were performed before and during DHEA treatment and at a 3-month follow up. RESULTS: DHEA and DHEA(S) levels were restored to normal in those patients receiving 50 mg whereas DHEA(S) level was slightly above the normal reference value in those receiving 200 mg. Circulating levels of androgens (androstenedione, testosterone and testosterone/SHBG ratio) were normalized in all patients. A slight rise in IGF-1 levels was seen in both groups as was a decrease in the levels of low and high density lipoproteins. No effect on blood glucose levels or insulin sensitivity was seen and no change of body composition was observed. No serious side-effects were seen, but some of the patients experienced increased apocrine sweat secretion (n = 7), itchy scalp (n = 2) and acne (n = 7), all of which were reversed when DHEA was discontinued. CONCLUSION: A daily replacement dose of 50 mg of DHEA results in near physiological levels of DHEA, DHEA(S) androstenedione and testosterone in women with Addison's disease, without severe side-effects. PMID- 10848884 TI - Lack of menstrual cycle effects on hypothalamic-pituitary-adrenal axis response to insulin-induced hypoglycaemia. AB - OBJECTIVE: Limited data are available on the effects of the menstrual cycle on the hypothalamic-pituitary-adrenal axis (HPA) function. This study evaluates HPA axis reactivity to insulin-induced hypoglycaemia over the menstrual cycle. PATIENTS: Twelve normal women were randomized to placebo and evaluated during three successive menstrual cycles. Menstrual phase was documented by menstrual diary and by oestradiol and progesterone levels at the time of each insulin tolerance test (ITT). Six normal men were included as a comparison in the statistical analysis. MEASUREMENTS: Afternoon ITTs were performed initially on the second or third day of menses in women, then seven more ITTs followed at one or two week intervals during the next 10 weeks. Serum measurements of glucose, adrenocorticotrophin (ACTH) and cortisol were obtained. RESULTS: The glucose and ACTH responses to the ITTs were similar between men and women. Cortisol levels at baseline and during the test were higher in men than in women, although the amount of change was similar. Glucose, ACTH and cortisol response to insulin induced hypoglycaemia did not vary over the menstrual cycle or during repeat testing in men or women. CONCLUSIONS: These data show that it is unnecessary to control for menstrual cycle during insulin tolerance tests performed at 1600 hours. It is, however, necessary to control for the effect of sex on cortisol levels. Repeat testing more than one week apart does not appear to influence the glucose, ACTH or cortisol response to insulin stress. PMID- 10848885 TI - Cushing's syndrome associated with a chemodectoma and a carcinoid tumour. AB - We present a case of Cushing's syndrome where 111In-octreotide scanning provided evidence for the presence of two neuroendocrine tumours. Uptake in the right neck corresponded to a chemodectoma, but there was no change in the clinical condition or fall in ACTH levels following surgical resection. Uptake in the left chest was assumed to relate to a bronchial carcinoid, but a tumour could not initially be localized on magnetic resonance imaging (MRI), spiral CT scanning or on selective venous sampling. A 1 cm bronchial carcinoid tumour was identified post-mortem which immunostained for ACTH. This case demonstrates that 111ln-octreotide scanning is a useful technique for identifying the source of ectopic ACTH production in difficult cases of Cushing's syndrome. Reliance should not be placed solely on standard imaging techniques to localize the tumour prior to surgery. Although rare, the possibility of a non-ACTH secreting neuroendocrine tumour should also be considered in patients with ectopic ACTH syndrome, who have positive 111In-octreotide scans. PMID- 10848886 TI - GH replacement therapy in adults with GHD improves vascular reactivity. PMID- 10848887 TI - A reply PMID- 10848888 TI - Radioactive iodine for hyperthyroidism: patient satisfaction survey. PMID- 10848889 TI - Thyroperoxidase immunostaining in evaluation of thyroid nodules. PMID- 10848890 TI - The 1microg Synacthen test in chronic fatigue syndrome. PMID- 10848891 TI - A reply PMID- 10848892 TI - The anti-inflammatory effects of interleukin-10 in allergic disease. PMID- 10848893 TI - The proteolytic activity of Der p 1 selectively enhances IgE synthesis: a link between allergenicity and cysteine protease activity. PMID- 10848894 TI - Matrix proteins and eosinophil mediator release. PMID- 10848895 TI - The chromones: history, chemistry and clinical development. A tribute to the work of Dr R. E. C. Altounyan. PMID- 10848896 TI - Endogenous interleukin-10 suppresses allergen-induced airway inflammation and nonspecific airway responsiveness. AB - BACKGROUND: The airway inflammation observed in asthma is orchestrated by activated Th-2 lymphocytes relevant for the induction of altered airway responsiveness. An increasing body of evidence is accumulating that not only the pro-inflammatory cytokines interleukin (IL)-4 and IL-5 but also the immunomodulating cytokines IL-12 and possibly IL-10 are crucial for regulating the allergic airway inflammation. OBJECTIVE: Since IL-10 is capable of downregulating a broad spectrum of pro-inflammatory cytokines, we wanted to address the role of endogenously produced IL-10 in vivo in allergic asthma. METHODS: Knockout (IL-10(-/-)) mice (C57BL/6-IL10tm1Cgn) and wild-type (WT) counterparts were immunized (day 0) and exposed (day 14-21) to ovalbumin (OVA). Airway inflammation and reactivity (AR), serum allergen-specific IgE responses and cytokine profiles in the bronchoalveolar lavage fluid (BALF) were studied. RESULTS: The IL-10(-/-) mice had more eosinophilic airway inflammation but comparable levels of allergen-specific serum IgE compared to the WT mice after allergen challenge. The AR was comparably increased in the OVA challenged WT and IL-10(-/-) mice vs sham-exposed WT, but not vs sham-exposed IL-10(-/-)mice since these showed a higher baseline AR. IFN gamma, IL-4 and IL-13 were comparable and IL-5 was even lower in the BALF of the in IL-10(-/-) mice compared to the similarly exposed WT mice. CONCLUSION: These results indicate that IL-10 plays an important and possibly direct role in the control of airway inflammation and responsiveness in an in vivo mouse model of allergy. PMID- 10848897 TI - Functional effects of the inhibition of the cysteine protease activity of the major house dust mite allergen Der p 1 by a novel peptide-based inhibitor. AB - BACKGROUND: The house dust mite (HDM) Dermatophagoides pteronyssinus is an important source of allergens, which can cause allergic conditions. The cysteine protease activity of Der p 1 may enhance the potency of this major mite allergen through cleavage of CD23 and CD25 from the surface of immune cells, IgE independent mast cell activation, increases in epithelial cell permeability and inactivation of an endogenous serine protease inhibitor. Inhibition of the enzymatic activity of Der p 1 may therefore be of therapeutic benefit. OBJECTIVE: To examine the activity of PTL11028, a newly developed Der p 1 inhibitor, in a range of assays that directly or indirectly measure Der p 1 protease activity and to compare its activity to endogenous cysteine protease inhibitors. METHODS: The proteolytic activities of purified Der p 1 or HDM extract and inhibitory properties of PTL11028 were examined through cleavage of an artificial peptidyl substrate, cleavage of CD23 from human B cells and permeability studies on primary human bronchial epithelial cells. RESULTS: PTL11028 is a highly potent and specific Der p 1 inhibitor, being effective against both purified protease and Der p 1 within HDM extract. PTL11028 can completely inhibit Der p 1-mediated CD23 cleavage from human B cells and also reduces HDM-induced human bronchial epithelial cell permeability by 50%. Der p 1 is potently inhibited by cystatin A and to a lesser extent by cystatins C and E/M. CONCLUSION: PTL11028 is a highly potent and selective irreversible inhibitor of the cysteine protease activity of Der p 1, an activity that may be modulated in vivo by some human cystatins. PTL11028 prevents the Der p 1-mediated cleavage of CD23 from human B cells and significantly reduces HDM-induced permeabilization of the epithelial barrier. PTL11028 is an important tool to examine the biological effects of Der p 1 in a range of in vitro and in vivo model systems. PMID- 10848898 TI - Regulation of the release of eosinophil cationic protein by eosinophil adhesion. AB - BACKGROUND: Varying release of eosinophil granule proteins depending on the stimulus and environmental factors has previously been reported. OBJECTIVE: To investigate the degranulation from adherent eosinophils by using mixed granulocytes. METHODS: Granulocytes isolated by Percoll gradient centrifugation were incubated on plates coated with plasma and tissue fibronectin, fibrinogen or human serum albumin (HSA) and stimulated with Mn2+, phorbol-myristate-acetate (PMA), formyl-methionyl-leucyl-phenylalanine (f-MLP) and combinations thereof, respectively. The release of eosinophil cationic protein (ECP) was measured by radioimmunoassay. RESULTS: Unstimulated eosinophils incubated in wells coated with plasma and tissue fibronectin, fibrinogen or HSA did not release any ECP. Furthermore, Mn2+ (5 mmol/L) did not induce release of ECP despite the fact that adhesion of eosinophils to these four proteins was induced. PMA stimulated a dose dependent release of ECP. Contemporaneous stimulation of eosinophils with PMA and Mn2+ induced a dramatically increased release of ECP regardless of which protein the eosinophils were adhering to. A small but significant release of ECP was found when eosinophils incubated on plates coated with fibrinogen and HSA were stimulated by f-MLP. Contemporaneous stimulation of eosinophils with f-MLP and Mn2+ did not induce any synergistic effect on the release of ECP. On the contrary, Mn2+ inhibited the release of ECP induced by f-MLP from eosinophils. Serum-opsonized Sephadex particles stimulated a potent increase of the release of ECP up to 12%-14% in the presence of plasma fibronectin and, in particular, fibrinogen. The kinetics of eosinophil adhesion and degranulation showed that the cellular adhesion preceded the degranulation response and that the degranulation patterns depend on the stimuli and environment. CONCLUSION: The present study indicated that cellular adhesion plays an important role in the regulation of eosinophil degranulation, but that adhesion and degranulation can be induced separately. PMID- 10848899 TI - Transepithelial migration of activated eosinophils induces a decrease of E cadherin expression in cultured human nasal epithelial cells. AB - BACKGROUND: The damage of respiratory epithelium in allergic diseases has a close correlation with the extent of eosinophil infiltration. It seems to be a good possibility that eosinophil infiltration could induce the changes in the expression of the epithelial cell adhesion molecules, which play a key role in the maintenance of structural and functional rigidity of epithelium. OBJECTIVE: We observed the expression of E-cadherin in cultured human nasal epithelial cells (HNECs) to study whether could it be affected by transepithelial migration of inflammatory cells, especially eosinophils. METHODS: In vitro study of the transmigration assay was designed using various types of inflammatory cells and HNEC monolayers. Various assays of each experimental group were done under the stimulation of interleukin-5 (IL-5) and/or platelet activating factor (PAF). Subsequently immunohistochemistry for E-cadherin was performed in the HNECs. The intensity of immunofluorescence of E-cadherin was quantified using confocal laser scanning microscopy (CLSM) system and compared before and after the transmigration. RESULTS: The mean intensity of immunofluorescence for E-cadherin decreased significantly after the transmigration of any types of inflammatory cells. Above all, the migration of eosinophils treated with IL-5 and PAF had an eminent effect on the decrease, whereas the degranulation extracts derived from eosinophils activated by IL-5 and secretory IgA (sIgA) did not affect the intensity. CONCLUSION: This work suggests that transepithelial migration of inflammatory cells can directly induce the decrease in epithelial E-cadherin expression. Furthermore, the most prominent change was induced by transmigration of activated eosinophils, which might be caused by some mechanisms independent of the eosinophil contents. The decrease in E-cadherin expression may trigger the damage of epithelial barrier, which contributes to the pathogenesis of allergic diseases. PMID- 10848900 TI - cDNA sequence of two sheep mast cell tryptases and the differential expression of tryptase and sheep mast cell proteinase-1 in lung, dermis and gastrointestinal tract. AB - BACKGROUND: Mast cell tryptases are a family of serine proteinases which are implicated in the proliferation of smooth muscle cells and fibroblasts, upregulation of interleukin-8 synthesis by endothelial cells, and recruitment of neutrophils and eosinophils. Trials in sheep showed that administration of a specific tryptase inhibitor reduced the late-phase response to inhaled allergen. OBJECTIVES: The aim of this study was to characterize the sequence and distribution of sheep tryptase(s), to validate the sheep model of allergic lung disease. METHODS: Reverse transcriptase PCR cloning was used to obtain cDNA sequences for two sheep tryptases. Lung and gut extracts were used as a source of tryptase for partial purification and characterization of the protein. The distribution of tryptase in skin, lung and gut was determined by immunohistochemistry, and compared with the distribution of sheep mast cell proteinase-1 (sMCP-1). RESULTS: Two highly similar cDNA sequences encoding sheep tryptase were found, indicating the presence of a 28 amino acid leader sequence, and a mature peptide of 245 amino acids. Partial purification of a putative sheep tryptase from lung and gut extracts was achieved using heparin-Sepharose affinity chromatography. Rabbit antihuman skin tryptase antiserum recognized the putative sheep tryptase on Western blot (approximate Mr 32-34 000) and paraformaldehyde fixed tissue sections. Tryptase was detected in all lung, skin and gut mast cells by this antibody, and transcripts for tryptase were detected in all three tissues by RT PCR. Sheep mast cell proteinase-1, detected by a specific monoclonal antibody, was present in all intestinal and gastric mucosal mast cells, but was not found in mast cells of the muscularis, thus defining at least two mast cell phenotypes in the gut. Whereas all dermal and pulmonary mast cells were tryptase positive, only a low proportion in the lung, and almost none in the dermis, were positive for sMCP-1. CONCLUSION: In view of the structural and functional similarities of sheep and human tryptases, and their similarity in tissue distribution in normal sheep, the sheep lung appears to be a good model for in vivo studies relating to human tryptase. PMID- 10848901 TI - Evaluation of treatment response in patients with seasonal allergic rhinitis using domiciliary nasal peak inspiratory flow. AB - BACKGROUND: Measurement of domiciliary nasal peak inspiratory flow rate (PIFR) may have a role in the objective assessment of treatment response in seasonal allergic rhinitis (SAR). OBJECTIVE: We wished to evaluate the relationship between domiciliary measurement of nasal PIFR and a variety of symptoms associated with rhinitis. METHODS: Thirty-eight nonasthmatic patients, mean age (SEM) 30 years (1.4), with symptomatic SAR were evaluated in a placebo controlled, single-blind, double-dummy, three way parallel group study. Patients received oral cetirizine 10 mg once daily and were randomized to receive, in addition, either: (i) intranasal mometasone furoate 200 microgram (n = 14); (ii) oral montelukast 10 mg (n = 11); or (iii) placebo (n = 13). All treatments were given once daily for 4 weeks and were preceded by a 1 week placebo period. Domiciliary diary cards were used to record morning (am) and evening (pm) domiciliary nasal PIFR and symptom (nasal, eye, throat) scores and impact on daily activity. A total daily symptom score was then calculated from the sum of these separate symptom scores. RESULTS: Baseline values for symptom scores and PIFR after placebo run-in were not significantly different when comparing the three groups. After 4 weeks of active treatment, there were significant (P < 0.05) improvements in nasal symptoms, total daily symptoms and PIFR with all treatments, with there being no significant confounding effect of pollen count, when analysed as a covariate. There were significant (P < 0.01) correlations for nasal symptom scores vs PIFRam (r = - 0.51) and PIFRpm (r = - 0.56), and similarly for daily activity vs PIFRam (r = - 0.42) and PIFRpm (r = - 0.48). CONCLUSIONS: These results suggest that domiciliary measurements of nasal peak flow correlate significantly with symptoms of seasonal allergic rhinitis and may therefore be a potentially useful objective short-term marker of treatment response. PMID- 10848902 TI - Occupational asthma caused by soybean flour in bakers--differences with soybean induced epidemic asthma. AB - BACKGROUND: Soybean dust has been identified as the causative agent of occupational asthma and asthma epidemics. Two main soybean hull allergens responsible for asthma outbreaks, Gly m 1 and Gly m 2, have been identified and purified. OBJECTIVE: The soybean allergens causing occupational asthma in exposed bakers were investigated and compared with those involved in epidemic asthma. METHODS: We report four bakers or confectioners with work-related respiratory symptoms who were exposed to soybean flour used as a baking additive. The causative role of soybean flour was investigated by immunological tests and specific inhalation challenge tests. Soybean flour allergens causing occupational asthma were characterized by immunoblotting. Immunoglobulin (Ig) E-reactivity to Gly m 1 and Gly m 2 was assessed using enzyme-linked immunosorbent assay. RESULTS: Sensitization to soybean flour was demonstrated by skin and serological tests and was confirmed by positive inhalation tests. Bronchial challenge test to soybean flour extract elicited immediate or dual asthmatic responses. Immunoblotting with soybean flour and soybean hull extracts showed IgE-binding mainly to high molecular weight (MW) allergens. There was an important individually different allergic response to inhalant soybean components. None of the patients showed IgE-reactivity against Gly m 1 and only one patient showed IgE-reactivity to the soybean hull allergen Gly m 2. CONCLUSION: These bakery workers had developed IgE-mediated occupational asthma to soybean flour. The allergens involved in occupational asthma caused by soybean flour are predominantly high MW proteins that are present both in soybean hull and flour, and they are different from the allergens causing asthma outbreaks, which are mainly low MW proteins concentrated in the hull. PMID- 10848903 TI - Infiltration of cytotoxic T cells in drug-induced cutaneous eruptions. AB - BACKGROUND: Previous in vitro data indicate that perforin containing drug specific cytotoxic T cells are involved in cutaneous drug reactions. OBJECTIVE: The aim of this study was to investigate the in situ expression of perforin and granzyme B together with the nature of the inflammatory infiltrate in acute drug induced exanthem. Furthermore, expression of interleukin (IL)-12 and interferon (IFN)-gamma, which are known to stimulate cytotoxic T cells, was investigated. METHODS: Skin biopsy specimens were obtained from 10 patients with a generalized maculopapular exanthem and from nine controls with normal skin. Expression of CD3, CD4, CD8, CD56, CD1a, CD68, CD25, HLA-DR, CD54, perforin, granzyme B, IL-12 and IFNgamma was analysed using immunohistochemistry. RESULTS: In contrast to the controls, the skin of patients with an exanthem was mainly infiltrated by T cells (CD4 > CD8) and showed a marked enhancement of perforin and granzyme B immunostaining. Double immunostaining revealed that perforin and granzyme B were expressed in both CD4+ and CD8+ cells, which were partly located at the dermoepidermal junction and in the epidermis. In addition, strong immunreactivity for IL-12 and IFNgamma was observed in the mononuclear cells infiltrate, indicating that these cytokines may be important in activation of these cytotoxic T cells. CONCLUSION: The increased numbers of perforin and granzyme B containing T cells infiltrating the dermoepidermal junction may contribute to the damage of epidermal cells, which is frequently observed as a typical feature of interface dermatitis in drug-induced exanthem. Our data provide further evidence that cytotoxic T cells play an essential role in cutaneous drug reactions. PMID- 10848904 TI - Temporal patterns of mediator release during developing cutaneous late-phase reactions. AB - BACKGROUND: Several inflammatory mediators have been found released in sites of cutaneous late phase reactions (LPR). However, the temporal pattern of their release during LPR development has not been characterized. OBJECTIVE: Determine hourly accumulation of mediator release in comparison with gross and inflammatory cell responses during developing LPR. METHODS: Skin chamber appended to sites of allergen and diluent control challenge with hourly collections. Then, study of exuding leucocytes in chamber bases. RESULTS: In the allergen-challenged sites, histamine release peaked in the first hour, then low level release over the next 5 h. Lactoferrin release from neutrophils started by the second hour, likely associated with released IL-8. Eosinophil cationic protein levels started increasing slightly later. The percentage of exuding leucocytes which were activated was significantly higher in the allergen challenge sites than in the control challenge sites CONCLUSIONS: Both gross LPR and local inflammatory cell responses in the skin start soon after the immediate mast cell activation in IgE mediated responses. Such inflammatory responses include leucocyte activation and mediator release. PMID- 10848905 TI - Prevalence of sensitization to Tetranychus urticae in greenhouse workers. AB - BACKGROUND: Tetranychus urticae (TU) is a macroscopic mite which infests a large number of plants of economic interest worldwide. It has recently been described as a cause of occupational allergic disease in greenhouse workers. However, there are no epidemiological data concerning the prevalence of TU allergy in an unselected exposed population. OBJECTIVE: The aims were to study the prevalence of TU sensitization among greenhouse workers and its relationship to the working environment and to personal factors. METHODS: We studied 246 consecutive greenhouse workers, recruited directly from the field. A clinical and epidemiological questionnaire, a skin-prick test (SPT) to TU and common allergens and TU-specific IgE (RAST) determinations were performed. Seventy-five healthy volunteers and 152 atopic patients were used as a control group. RESULTS: The prevalence of a positive SPT to TU was of 25%. Forty-five workers (19%) were TU allergic, occurring more often in atopic greenhouse workers (P < 0.0001). Seven per cent showed asymptomatic sensitization. The time of exposure to TU was significantly greater in the TU-allergic patients (P < 0.05). The probability of sensitization to TU was 3.7 times greater in exposed than in non-exposed subjects (P < 0.0001). CONCLUSIONS: In this study, the prevalence of TU sensitization was 25%. There were significant associations with TU allergy and atopy and the time of exposure to TU. PMID- 10848906 TI - Differences in size selective aerosol sampling for pollen allergen detection using high-volume cascade impactors. AB - BACKGROUND: Assessment of the allergen content of airborne particles small enough to reach the bronchiolar airways is important for a better understanding of allergic asthma. OBJECTIVE: In order to test the performance of a high-volume cascade impactor for size-selective sampling of airborne particles, the characteristics of pollen deposition, and the particle size-dependent allergen distribution, were re-examined. METHODS: Two cascade impactors with rectangular slots were run in parallel, one with glass fibre filters on all stages, the other with silicone grease on stage 2 (collection of particles of size 4.2-10.2 micrometer) instead. Pollen was counted using light microscopy and allergens were measured using ELISA techniques. RESULTS: In the impactor without the greased stage 2, a heavy bounce and blow-off for pollen was found. Bounced pollen was deposited mainly on the back-up filter, the sampling stage for particles smaller than 1.4 micrometer. However, if stage 2 was coated with silicone grease, less than 1% of total pollen was found in the lower stages. On the first stage of the sampler (50% cut-point, 10.2 micrometer) where all pollen should impact, pollen had a deposition efficiency of only 24-47%, depending on the abundant pollen species. Much less allergen was found in fine particle fractions than in previous studies in which particles were sampled with similar samplers but without grease as a pollen trap. CONCLUSIONS: The observed bounce and blow-off of pollen without a greased stage 2 leads to serious mistakes when assessing the allergen content of the fractions collecting particles smaller than intact pollen, i.e., below 10.2 micrometer. Pollen allergen concentrations in respirable particle fractions are much lower than in the pollen size fraction. This is of great importance both for symptoms of sensitized allergic asthmatics and for research of asthma-related mechanisms. PMID- 10848907 TI - Allergen-induced airway inflammation and bronchial responsiveness in interleukin 5 receptor alpha chain-deficient mice. AB - OBJECTIVE: The role of IL-5 receptor alpha chain (IL-5Ralpha) in the onset of bronchial hyperresponsiveness (BHR) to acetylcholine was investigated by testing IL-5Ralpha knockout (IL-5Ralpha KO) mice. METHODS: Mice were immunized with antigen at intervals of 12 days. Starting 10 days after the secondary immunization, mice were exposed to antigen three times every fourth day. Twenty four hours after the last antigen challenge, bronchial responsiveness to acetylcholine was measured and bronchoalveolar lavage was carried out. RESULTS: Twenty-four hours after the last antigen inhalation, total and differential cells counts of bronchoalveolar lavage revealed a significant increase in eosinophils and lymphocytes in ovalbumin-exposed wild-type mice. In IL-5Ralpha KO mice, there was little increase of eosinophils in bronchoalveolar lavage fluid (BALF). The production of IL-5 in BALF increased in both mice after repeated antigen challenge, and there was no significant difference between wild-type and IL 5Ralpha KO mice. Similar to the BAL study, histological sections of lung tissue from ovalbumin-exposed wild-type mice exhibited airway eosinophilic inflammation, which was attenuated by the deficiency of IL-5Ralpha chain. There was no significant difference in serum antigen-specific IgE levels between wild-type and IL-5Ralpha KO mice after immunization nor antigen inhalation. Repeated antigen provocation caused BHR to acetylcholine in wild-type mice. In contrast, no BHR was observed in IL-5Ralpha KO mice after repeated inhalation of antigen. CONCLUSION: These findings indicate that IL-5Ralpha plays an important role in the development of antigen-induced airway eosinophilia and BHR in mice. PMID- 10848908 TI - Influence of bronchial allergen challenge on histamine release by human basophils. AB - BACKGROUND: Basophils can be primed by cytokines such as interleukin (IL) -3, IL 5 or granulocyte macrophage-colony stimulating factor (GM-CSF). It has been described that the concentrations of these cytokines are enhanced at sites of allergic inflammation as well as systemic in allergic asthma. OBJECTIVE: To investigate the priming status of basophils as detected by thapsigargin-induced histamine release during bronchial allergen challenge. METHODS: Ten subjects allergic to house dust mite were challenged via an aerosol delivery system. Spontaneous leucocyte histamine release as well as histamine release induced by various stimuli was measured in vitro at several time points. In addition, lung function parameters, serum IL-5 and blood eosinophil counts were evaluated. RESULTS: We found no effect of bronchial allergen challenge upon spontaneous leucocyte histamine release, nor upon histamine release induced by anti immunoglobulin (Ig) E, house dust mite extract, C5a, fMLP, IL-3, PMA+ thapsigargin or IL-3+ thapsigargin. However, the priming status of basophils as measured by thapsigargin-induced histamine release was enhanced at 24 h after bronchial allergen challenge. Analysis of the individual data showed a heterogeneous initial response (30 min, 6 h) followed by a predominant increase at 24 h after allergen challenge. This increase in the thapsigargin-induced histamine release correlated with the increase in serum IL-5 levels at 24 h after allergen challenge. CONCLUSION: The priming status of human basophils as measured by thapsigargin-induced histamine release is enhanced 24 h after allergen challenge. PMID- 10848909 TI - Comparison of the efficacy, safety and quality of life provided by fexofenadine hydrochloride 120 mg, loratadine 10 mg and placebo administered once daily for the treatment of seasonal allergic rhinitis. AB - BACKGROUND: As there have been no previously published studies, this multinational, double-blind, randomized, placebo-controlled, parallel group study compared the efficacy, safety and impact on quality of life (QoL) in seasonal allergic rhinitis patients (SAR) of fexofenadine and loratadine (with placebo), when administered once daily. METHODS: Six hundred and eighty-eight SAR patients were randomized to receive fexofenadine HCl 120 mg, loratadine 10 mg or placebo, once daily for 2 weeks. The key parameters were the change from baseline in: mean 24-h reflective total symptom scores (TSS); sum of four individual symptom scores, excluding nasal congestion; instantaneous TSS; individual symptom scores including nasal congestion; and Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ). Adverse events were recorded. RESULTS: Mean 24-h reflective and instantaneous TSS were significantly reduced by both fexofenadine HCl (both P or = 0.08). Subsequent RV16 infection was associated with a trend towards an increase in ICAM-1 expression in the BUD group (P = 0.07), whereas the increase was significant in the PLAC-group (P = 0.03). However, the increase was not significantly different between the groups (P = 0.74). Epithelial intactness score was not different between the groups before RV16 infection (P > or = 0.07), and no significant changes were observed in either group (P > or = 0.59). Moreover, ICAM-1 score did not correlate significantly with epithelium score in either group, at any time-point (P > or = 0.27). CONCLUSION: We conclude that an RV16 common cold in atopic asthmatic subjects is associated with increased ICAM-1 expression in the bronchial epithelium, which is not related to epithelial intactness. Glucocorticoid treatment does not appear to prevent the RV16-associated increased ICAM-1 expression. This suggests that other treatment modalities are required to protect against the spreading of infection during rhinovirus-induced exacerbations in asthma. PMID- 10848926 TI - Prolonged oral treatment with low doses of allergen conjugated to cholera toxin B subunit suppresses immunoglobulin E antibody responses in sensitized mice. AB - BACKGROUND: Oral tolerance is a long recognized method for inducing systemic immunological tolerance. However, large doses of antigen and frequent administrations are often required. By linking the antigen to the nontoxic mucosa binding B subunit of cholera toxin (CTB), the required amount can be dramatically reduced. We have previously shown that mucosal administration of small amounts of antigens coupled to CTB can suppress peripheral Th1 cell-reactivity and associated inflammatory immunopathology in both naive and systemically-immunized animals. Induction of oral tolerance by repeated feeding of relatively small doses of antigen has, in some cases been shown to involve the generation of regulatory Th2-like CD4+ T cells, and hence could promote rather than suppress type I immunoglobulin (Ig) E-mediated allergic responses. OBJECTIVES: We examined whether oral prophylactic or therapeutic administration of a model allergen coupled to CTB would modulate allergen-specific IgE responses in high IgE responder Balb/c mice. METHODS: Ovalbumin (OVA) was used as a model allergen. Mice were treated perorally with free or CTB-coupled OVA before or after systemic priming with alum-adsorbed OVA. Allergen-specific IgE levels in serum were measured with the passive cutaneous anaphylaxis test at various time-points. RESULTS: Oral administration of a single low dose of CTB-linked OVA, prior to systemic sensitization and challenge with OVA, suppressed allergen-specific serum IgE antibody responses. Treatment with comparable doses of free OVA was much less effective. Most importantly, oral treatment with CTB-OVA conjugate could also suppress an already initiated IgE antibody response, but to achieve such a 'therapeutic effect', administration of multiple low doses of conjugate over a long time was required. Oral treatment with CTB-OVA conjugate could also effectively suppress antigen-specific Th1-mediated delayed-type hypersensitivity. Thus treatment with a CTB-conjugated model allergen can affect a broad range of T cell-driven immune responses, even in antigen-experienced animals. CONCLUSION: These results may impact on the development of therapeutic vaccines against type I allergies. PMID- 10848927 TI - Regulation of Fc epsilonRI expression on human monocytic cells by ligand and IL 4. AB - BACKGROUND: The Fc epsilonRI subunit composition and kinetic of expression differ between antigen-presenting cells and mast cells. Up to now, there has been no human in vitro model available that mimics the characteristics on monocytes. OBJECTIVE: The characterization of a natural human monocytic cell line (THP1), which expresses Fc epsilonRI, and the comparison to primary human monocytes and other monocytic cell lines, which only express Fc epsilonRI after transfection with the human Fc epsilonRI alpha-chain gene. METHODS: Surface receptor expression was characterized by flow cytometry, the human Fc epsilonRI alpha chain gene was introduced by electroporation, and induction of Fc epsilonRI alpha chain message was detected by semiquantitative RT PCR. RESULTS: Here we show that the parental human cell line THP1, but none of the other cell lines tested, displays surface Fc epsilonRI in response to IL-4 or incubation with receptor ligand (IgE, antibody). Transfection of Fc epsilonRI alpha-chain resulted in receptor expression on all cell lines, all of which increased surface Fc epsilonRI in the presence of IgE. Only the THP1-alpha transfectant, however, further increased receptor levels in response to IL-4, resulting from mRNA induction for the Fc epsilonRI-alpha, but not the beta- or gamma-subunit. CONCLUSION: Based on THP1, U937 and HL60 and their alpha-chain transfectants we present a model system for the study of Fc epsilonRI regulation and signalling on human cells. THP1 in particular, due to its responsiveness to both ligand and IL 4, even without prior manipulation, is ideally suited to address questions on Fc epsilonRI modulation in an 'allergic environment'. PMID- 10848928 TI - Identification of peptide motifs recognized by a human IgG autoanti-IgE antibody using a phage display library. AB - BACKGROUND: The potential of murine monoclonal anti-IgE antibodies as long-term therapy for atopic diseases will have to rely, for the time being, on passive antibody administration. There is therefore considerable interest in developing a peptide-based vaccine for active immunization to elicit long-term protective anti IgE antibodies in the patient. It has been shown that some human IgG autoanti-IgE antibodies have the ability to partially block the binding of IgE to Fc receptors such as Fc epsilonRI. Therefore, the epitopes recognized by such antibodies could have vaccine potential. OBJECTIVE: To determine the epitope specificity of one such human IgG anti-IgE antibody. METHODS: A 15-mer phage-peptide library was used to establish the epitope specificity of an IgG anti-IgE antibody isolated from the serum of an asthma patient. RESULTS: The SRPSP sequence, or part of it (i.e. RPS, RPSP, SPS or PSP), was present in all 18 phage-peptides that have been sequenced. This common motif was found to be within the human epsilon chain sequence Ser341-Thr355 near the N-terminus of the C epsilon3 domain. According to the human Fc epsilon model, the most accessible residues in this sequence are Arg342, Ile350, Arg351, Lys352 and Ser353. CONCLUSIONS: The present data should provide the molecular basis for the rational design of a suitable peptide immunogen (vaccine) for boosting the production of protective autoanti-IgE antibodies. PMID- 10848929 TI - Sunscreens: is an ounce of protection worth the hassle? PMID- 10848930 TI - History of liposuction. PMID- 10848931 TI - Histologic subtyping and malignancy assessment of cutaneous squamous cell carcinoma. AB - BACKGROUND: Squamous cell carcinomas (SCCs) of the skin have a wide range of histologic subtypes and there are indications of differences in prognosis. OBJECTIVE: The morphologic variety of SCCs with respect to its biological behavior and the further course of disease is analyzed, with emphasis on histopathologic criteria, briefly quoting the main clinical and pathogenetic aspects. METHODS: Referring to the international tumor classification of the World Health Organization, histologically different carcinoma variants are presented and discussed, based on a review of the literature regarding each subtype, and also including the desmoplastic SCC type. RESULTS: Histologically, common invasive SCCs are most frequently found, while metastases mainly occur in tumors of high thickness and poor differentiation. The immature spindle cell carcinoma type resembles sarcoma and may grow rapidly with an aggressive clinical course. Lymphoepithelioma-like carcinoma of the skin is extremely rare and its histogenesis remains to be elucidated. Thus far, one case with metastasis and lethal outcome has been reported. As details determining the progression ability have so far only been scanty and partially contradictory, more investigations are necessary, especially for acantholytic SCCs and invasive SCCs developing from Bowen's disease, whereas verrucous carcinomas can be categorized as low malignancy neoplasms. Desmoplastic SCCs, especially with large tumor thickness, should be separated from other SCC subtypes due to their high risk of local recurrence and metastatic spread. CONCLUSION: The future outcome of SCCs of the skin is significantly influenced by their histologic grade and tumor thickness. In addition, subtyping represents another valuable histopathologic tool for improving the assessment of malignancy. PMID- 10848932 TI - Metastatic microcystic adnexal carcinoma in an immunocompromised patient. AB - BACKGROUND: Microcystic adnexal carcinoma is an uncommon, locally aggressive cutaneous neoplasm. To date, there are only two reports of histologically proven lymph node involvement with this tumor. We describe a case of a patient with microcystic adnexal carcinoma who developed multiple local metastasis in transit with histologically proven lymph node involvement and was diagnosed with chronic lymphocytic leukemia. OBJECTIVE: To describe the details of our case and to review what is currently known about this tumor. METHODS: Mohs micrographic surgery was utilized for tumor removal. RESULTS: This patient developed multiple tumors of the scalp over the period of a 1 year which were histologically proven to be microcystic adnexal carcinoma. All tumors were noncontiguous and presented on the scalp. During the histologic analysis of the last tumor removed by Mohs micrographic surgery a lymph node was resected which revealed infiltrative microcystic adnexal carcinoma. CONCLUSIONS: We present the case of an immunocompromised patient treated for microcystic adnexal carcinoma with Mohs micrographic surgery who proceeded to develop local metastasis in transit. PMID- 10848933 TI - Extensive tissue necrosis following high-concentration sclerotherapy for varicose veins. AB - BACKGROUND: Tissue necrosis after sclerotherapy has been observed, but is unexplained. OBJECTIVE: To present the complication of extensive tissue necrosis following high-concentration sclerotherapy for varicose veins. METHODS: Cases coming to the attention of the authors are presented briefly with commentary and discussion to explain the mechanisms of tissue destruction. RESULTS: Although the complication of extensive tissue necrosis has been ascribed to intra-arterial injection, in fact, careful study of the cases described here shows that intravenous injection was present in each case. A theory of distribution of the sclerosant into the arterial arborization is proposed. This theory would explain the distribution of sclerosant into the arterial tree and would also explain the causation of extensive tissue necrosis. Mention is made of experimental work in which intra-arterial injection was not the mechanism of causation of tissue necrosis. CONCLUSION: Extensive tissue necrosis following high-concentration sclerotherapy may be rare, but its occurrence is serious and its treatment may be incomplete. PMID- 10848934 TI - Alar rotation flap for small defects of the ala. AB - BACKGROUND: Surgical defects of the alar lobule can be difficult to repair with aesthetically pleasing results. Full-thickness skin grafts are often smoother than the sebaceous skin of the ala. Random patterned flaps from the cheek or proximal nose usually bridge and obliterate the supra-alar crease and may cause nasal valve malfunction. OBJECTIVE: We describe and illustrate a technique to repair subtotal alar lobule defects within the cosmetic unit of the alar lobule. METHODS: Twenty-three consecutive alar lobule rotation flaps for repair of Mohs surgical defects were reviewed by patient examination and interview. RESULTS: Twenty-one of 23 patients were contacted. Patients rated cosmetic results as excellent (18), good (2), or fair (1), and no patients graded their results as poor. Six patients reported "a little" breathing difficulty in the postoperative period that resolved within 6 months. Anesthesia reported by 11 of 21 patients resolved within 5 years in 8 of 9 patients available for follow-up. CONCLUSION: Rotation of the ala combined with cheek advancement is a cosmetically pleasing and functional method to repair deep defects of the ala. PMID- 10848935 TI - Risk of basal cell and squamous cell carcinoma in persons with prior cutaneous melanoma. AB - BACKGROUND: Melanoma has been associated with an overall increase in actinic tumors, including actinic keratoses, as well as with noncutaneous malignancies. OBJECTIVE: Determine the risk of developing basal cell and squamous cell skin cancer in patients with prior cutaneous melanoma (actinic keratoses not encountered). METHODS: This retrospective study included 1396 white patients with prior cutaneous melanoma followed at the Roswell Park Cancer Institute in the period 1977-1978. The control group was the white population of the Detroit area in the same period (1977-1978). RESULTS: A total of 25 patients (18 males, 7 females) developed 35 basal cell and/or squamous cell carcinomas: 18 developed basal cell carcinomas, 2 squamous cell carcinomas, and 5 both. The calculated odds ratio was 3.49 (males 3.67, females 2.86, 95% confidence interval 1.52 8.00). No correlations were found with age, type, anatomic site, and length of follow-up of cutaneous melanoma. CONCLUSION: A history of cutaneous melanoma significantly increases the risk of basal cell and squamous cell skin cancer. PMID- 10848936 TI - Potential role of the new specific COX-2 inhibitors in dermatologic surgery. PMID- 10848937 TI - Multifactorial classification of male and female androgenetic alopecia. AB - BACKGROUND: Various classifications of male androgenetic alopecia have been described. In fact, all the classification schemes proposed so far are only topographic. A more objective, accurate, and detailed approach to classifying baldness is needed. OBJECTIVE: To propose a dynamic multifactorial classification of certain parameters that can be quantitated and computerized. METHODS: A multifactorial classification has been developed to study parameters such as fixed distances of the face, scalp mobility and thickness, and covering power of hair. This includes density, caliber, shape, length, growth rate, and hair color. RESULTS: Classification proved to be efficient during the fluctuations of different parameters in hormonal and minoxidil treatments. It also helps to determine surgical indications of hair transplant and the stage of maximal baldness. CONCLUSION: This approach will lead to a better evaluation of the evolution of androgenetic alopecia in both sexes, either spontaneously or under treatment. PMID- 10848938 TI - Review of closed dressings after laser resurfacing. AB - BACKGROUND: Laser skin resurfacing has become an accepted technique for the treatment of facial rhytides and associated solar skin damage. Achieving a successful result is directly related to proper postoperative wound care during the reepithelialization process. There are open and closed approaches to the treatment of the post-laser resurfacing patient with distinct advantages and disadvantages. OBJECTIVE: To review the most commonly used closed dressings after facial laser skin resurfacing and compare their advantages and disadvantages. To compare clinical findings with a group of patients treated exclusively with an open technique. METHODS: Review of composite foams, polymer film, polymer mesh, and hydrogel products and prospective observations of clinical outcomes of patients treated with each dressing category after facial laser skin resurfacing. We perform a retrospective chart review of a group of patients treated exclusively with an open technique comparing crust formation, comfort, and pruritus with the prospective group of patients treated with closed dressings. RESULTS: The closed dressings available today each have unique structural configurations and adhesive properties intended to maintain an occlusive wound environment. Patient acceptance of these dressings was favorable, with improved comfort compared to the open dressing group. Complications of bacterial infections and contact dermatitis were not observed when closed dressings were used with a protocol for dressing changes performed at 48 hours. Rates of reepithelialization did not vary according to dressing category. Crust formation and postoperative pruritus occurred less frequently when closed occlusive dressings were worn by patients. CONCLUSIONS: When used properly, these dressings improve patient comfort, simplify their postoperative wound care, and do not increase the risk of infection or contact dermatitis. Overall satisfaction was highest with perforated mesh and polymer dressings for full-face wounds. PMID- 10848939 TI - Occlusive dressing versus oxygen mist therapy following CO2 laser resurfacing. AB - BACKGROUND: Oxygen is an essential element for collagen synthesis and reepithelialization. The use of topical oxygen after CO2 laser resurfacing has not been studied. OBJECTIVE: To compare the rate and quality of healing in wounds treated with an oxygen mist to those treated with occlusive dressing following CO2 laser resurfacing. METHODS: Three patients underwent CO2 laser resurfacing to each half of the face 3 weeks apart. Postoperatively, half of the face was treated with an oxygen mist protocol for 5 days, while the other half was treated with occlusive dressing for 4 days. RESULTS: At postoperative day 5, significantly less crusting was observed on the half of the face treated with the oxygen mist protocol (p < 0.05). CONCLUSION: The oxygen mist postoperative protocol may offer patients similar overall healing rates and significantly less crusting compared to occlusive dressing. PMID- 10848940 TI - The use of imiquimod 5% cream for the treatment of superficial basal cell carcinomas in a basal cell nevus syndrome patient. AB - BACKGROUND: Imiquimod 5% cream has been used effectively to treat superficial basal cell carcinomas (BCCs). OBJECTIVE: The purpose of this study is to examine the effectiveness, tolerability, and desirability of imiquimod 5% cream in treating superficial non-facial basal cell carcinomas in a patient with basal cell nevus syndrome. METHODS: Three biopsy-proven nonfacial BCCs were treated for 18 weeks with once daily application of 5% imiquimod cream. The lesions were then removed to search histologically for residual tumor. RESULTS: The two adequately treated tumors revealed no residual BCC upon removal. Our patient reported that he tolerated the treatment but he would not desire this treatment again based on the length of treatment time and the degree of local inflammation at the treatment sites. CONCLUSION: Imiquimod 5% cream appears to be effective in eradicating superficial nonfacial BCCs. The degree of local inflammatory response may affect the patients' tolerability of treatment and therefore patient compliance. PMID- 10848941 TI - Sentinel node biopsy for staging of aggressive digital papillary adenocarcinoma. AB - BACKGROUND: Aggressive digital papillary adenocarcinoma is a rare malignancy with a propensity for metastases and recurrence. The role of lymph node staging in this tumor is poorly defined. We describe the use of sentinel lymph node mapping and biopsy in staging this tumor. OBJECTIVE: To describe and discuss the use of lymphatic mapping in staging aggressive digital papillary adenocarcinoma. METHODS: Sentinel lymph node mapping and biopsy was performed after excision of an aggressive digital papillary adenocarcinoma of the toe. RESULTS: Metastatic tumor cells were absent in sentinel lymph nodes by hematoxylin and eosin staining and immunocytochemistry analysis. CONCLUSION: We describe the first reported case of staging lymph nodes in a patient with aggressive digital papillary adenocarcinoma utilizing sentinel lymph node mapping and biopsy. PMID- 10848942 TI - Mixed capillary/lymphatic malformation with coexisting port-wine stain: treatment utilizing 3D MRI and CT-guided sclerotherapy. AB - BACKGROUND: Lymphatic malformation, a benign malformation of the skin and the subcutaneous tissues, is divided into two major groups: the classical and the localized forms. Pathologically lymphatic malformation often consists of sequestered lymphatic cisterns with thick muscle walls lying deeply in the subcutaneous tissue. Communicating via dermal lymphatic channels with superficial pseudovesicles, they can vary in size depending on the pressure transmitted by the cisterns beneath. METHODS: We present a patient with mixed capillary/lymphatic malformation and coexisting port-wine stain since birth. To demonstrate the anatomic extent and the subcutaneous involvement we performed a 3D reconstruction of a magnetic resonance imaging (MRI). The diagnostic procedures, therapeutic possibilities, and complications regarding this rare appearance are reviewed. RESULTS: Good results could be obtained with CO2 laser vaporization of the superficial lesions and computed tomography (CT)-guided transcutaneous sclerotherapy for the deeper cisterns with doxycycline. CONCLUSION: The combination of CO2 laser treatment and sclerotherapy with doxycycline seems to present a treatment option for cutaneous and subcutaneous lymphangioma circumscriptum with rare side effects. PMID- 10848943 TI - Mycobacterium fortuitum infection following neck liposuction: A case report. PMID- 10848944 TI - Reconstruction conundrum #1. Excision of a basal cell carcinoma. PMID- 10848946 TI - Controversies in dermatologic surgery PMID- 10848945 TI - Interview with Dr. Andre Davy, Honorary President, International Union of Phlebology; Honorary President, French Society of Phlebology; Honorary President, European Society of Phlebectomy. Interview by Jose Antonio Olivencia. AB - Dr. Davy is the Honorary President International Union of Phlebology, Honorary President French Society of Phlebology, Honorary President European Society of Phlebectomy. Dr. Andre Davy was born in 1924 in Basse, Normandy. I met Dr. Davy in 1967 in a World Congress of Phlebology in Amsterdam. I was so impressed with his skills and knowledge that when he later on proposed to be associated in the practice of phlebology, I accepted immediately. That lead to a very long medical and surgical partnership. He was an expert in the Muller phlebectomy and was also interested in chronic venous insufficiency, chronic stasis ulcer and lymphedema. He promoted sclerotherapy as part of the overall treatment of venous disease. In the early seventies, he started a training program that included not only theoretical but also practical knowledge of phlebology. In 1974 he published a book entitled, "Les Varices." He organized the first Franco-British Symposium of Phlebology that was held in Touquet in May 1981. That symposium was a complete success. It was immediately followed by a second Franco-British symposium and later on by the creation of the Venous Forum of the Royal Society of Medicine, the birth of the English journal "Phlebology," the North American Society of Phlebology, and the American Venous Forum. Later, he became Editor in Chief of Phlebologie, the journal of the French Society of Phlebology. He was named President of the French Society of Phlebology in 1986 and in 1989 President of the International Union of Phlebology. This very well educated and calm gentleman with a very outward tranquil appearance and great strength of character has always held strong opinions. He is now retired at his family home in Pont L'Eveque, France and spends his time reading, listening to the music of his favorite composers Verdi and Mahler, continuously visiting old friends such as Jean Van der Stricht, Robert Muller, Claude Gillot, and still remaining very close to his family. PAUL OUVRY, MD France PMID- 10848947 TI - The use of a suspension suture in lieu of a cartilage strut for deep alar defects. PMID- 10848948 TI - Use of the double-bladed scalpel in peripheral margin control of dermatofibrosarcoma protuberans. AB - BACKGROUND: The double-bladed scalpel previously has not been cited in the published literature for use in Mohs micrographic surgery. OBJECTIVE: To allow for maximum tissue conservation and greatest possible intraoperative efficiency for peripheral margin control of a massive tumor. METHODS: We describe the use of the double-bladed scalpel during Mohs surgery for peripheral margin control of a large dermatofibrosarcoma protuberans (DFSP). RESULTS AND CONCLUSION: While not necessary for the majority of routine Mohs cases, utilization of the double bladed scalpel may be timesaving, especially when used for peripheral margin control of a massive tumor. PMID- 10848949 TI - The liposuction apparatus "suspension device": hi-plane liposculpture. AB - BACKGROUND: The improvement of liposuction equipment and techniques has resulted in a cumbersome operating room array of aspirator hoses, irrigation/infiltration tubing, and the insulated command and control wiring of current ultrasonic or power liposuction devices. This situation is further complicated by the presence of a second surgeon, the concomitant use of additional traditional or ultrasonic cannulas, and the suction hose of a second aspirator. OBJECTIVE: To solve the problems of sterility and operating room organization, the authors describe a simple, releasable suspension design. METHOD: This apparatus can be temporarily or permanently installed in minutes and is easily maintained to control numerous sterile hoses and wires required for certain liposuction procedures. RESULTS: This suspension method appeared to enhance the efficiency and ease of use by single and dual surgeons during cases utilizing ultrasonic and nonultrasonic cannulas. The time savings and ease of use was most noticeable in cases of dual surgeons using both ultrasonic and nonultrasonic systems simultaneously. In addition, surgical personnel were freed for other tasks. Single-surgeon, traditional liposuction was less notably facilitated in terms of speed and fewer personnel. However, wiring and tubing control is facilitated in any case. CONCLUSION: A simple liposuction suspension system facilitates the liposuction procedure to varying degrees depending upon the number of surgeons and devices in simultaneous use. PMID- 10848950 TI - Lessons on dermoscopy #4. Poorly defined pigmented lesion. Diagnosis: pigmented BCC. PMID- 10848951 TI - The role of adjuvant interferon in high risk melanoma patients. PMID- 10848952 TI - Comments on the use of sentinel lymph node biopsy in patients with Merkel cell carcinoma. PMID- 10848953 TI - Why would anyone bother with "the lateral femoral cutaneous nerve block" for harvesting split-thickness grafts? PMID- 10848954 TI - Towards a molecular understanding of polycythemia rubra vera. AB - Polycythemia rubra vera (PV) is one of four diseases collectively called the myeloproliferative disorders (MPDs). Each disorder leads to an increased production of one or several hematopoietic cell lineages. MPDs arise from acquired mutations in a pluripotent hematopoietic stem cell. However, the molecular mechanisms leading to the development of these diseases are poorly understood. This review will summarize and evaluate recent advances in our understanding of one particular MPD, PV. PMID- 10848955 TI - Archaeal protein translocation crossing membranes in the third domain of life. AB - Proper cell function relies on correct protein localization. As a first step in the delivery of extracytoplasmic proteins to their ultimate destinations, the hydrophobic barrier presented by lipid-based membranes must be overcome. In contrast to the well-defined bacterial and eukaryotic protein translocation systems, little is known about how proteins cross the membranes of archaea, the third and most recently described domain of life. In bacteria and eukaryotes, protein translocation occurs at proteinaceous sites comprised of evolutionarily conserved core components acting in concert with other, domain-specific elements. Examination of available archaeal genomes as well as cloning of individual genes from other archaeal strains reveals the presence of homologues to selected elements of the bacterial or eukaryotic translocation machines. Archaeal genomic searches, however, also reveal an apparent absence of other, important components of these two systems. Archaeal translocation may therefore represent a hybrid of the bacterial and eukaryotic models yet may also rely on components or themes particular to this domain of life. Indeed, considering the unique chemical composition of the archaeal membrane as well as the extreme conditions in which archaea thrive, the involvement of archaeal-specific translocation elements could be expected. Thus, understanding archaeal protein translocation could reveal the universal nature of certain features of protein translocation which, in some cases, may not be readily obvious from current comparisons of bacterial and eukaryotic systems. Alternatively, elucidation of archaeal translocation could uncover facets of the translocation process either not yet identified in bacteria or eukaryotes, or which are unique to archaea. In the following, the current status of our understanding of protein translocation in archaea is reviewed. PMID- 10848956 TI - Lck protein tyrosine kinase is a key regulator of T-cell activation and a target for signal intervention by Herpesvirus saimiri and other viral gene products. AB - Protein tyrosine kinases (PTKs) are critically involved in signaling pathways that regulate cell growth, differentiation, activation, and transformation. It is not surprising, therefore, that viruses acquire effector molecules targeting these kinases to ensure their own replication and/or persistence. This review summarizes our current knowledge on Lck, a member of the Src family of PTK, and its viral interaction partners. Lck plays a key role in T lymphocyte activation and differentiation. It is associated with a variety of cell surface receptors and is critical for signal transduction from the T-cell antigen receptor (TCR). Consequently, Lck is targeted by regulatory proteins of T-lymphotropic viruses, especially by the Herpesvirus saimiri (HVS) tyrosine kinase interacting protein (Tip). This oncoprotein physically interacts with Lck in HVS transformed T cells and has an impact on its catalytic activity. However, while Tip inhibits Lck activity in stably expressing cell lines, opposite effects were observed in several in vitro systems. At least in part, this complex situation may be related to the bipartite nature of the interaction surface of the two proteins. Studies on the interrelationships between Lck and its viral partners contribute to the understanding of the mechanisms of T-cell growth regulation, in general, and of viral pathogenicity in particular. In addition, understanding the regulation of Lck activity by viral proteins may serve as a basis for the development of new drugs capable of modifying Lck activity in different pathological situations. PMID- 10848957 TI - Recursive use of evolutionary conservation data in molecular modeling of membrane proteins A model of the multidrug H+ antiporter emrE. AB - Membrane proteins are currently the most biomedically important family of proteins, serving as targets for the majority of pharmaceutical agents. It is also clear that they are invariably abundant in all of the genomes sequence so far, representing up to a third of all open reading frames. Finally, and regrettably, it is clear that they are highly resistant to structural elucidation, representing less than 0.2% of the Protein Data Bank. Recent accomplishments in genome sequencing efforts, however, may help offset this imbalance through the availability of evolutionary conservation data. Herein, we develop a novel approach, utilizing a combination of evolutionary conservation data and global searching molecular dynamics simulations to model membrane proteins, deriving a model for the multidrug H+ antiporter EmrE, a transmembrane four-helix bundle. Structures resulting from an extensive, rotational molecular dynamics search, were evaluated by comparing the residue specific interaction energy and the evolutionary conservation data. Subsequent rounds of molecular dynamics, in which confinement of the search space was undertaken in order to achieve a self consistent result, point to a structure that best satisfies the evolutionary conservation data. As the conservation patterns calculated for each of the helices suggested that the different conservation pattern for helix 3 (as well as being the most conserved) might be due to the oligomeric nature of EmrE, a dodecamer of helices was constructed based on the result of a search of helix 3 as a trimer. The resulting interaction energy per residue in the final model is in reasonable agreement with the evolutionary data and consistent with recent site directed mutagenesis experiments, pointing to the strength of this method as a general tool. PMID- 10848958 TI - The role of arginine 47 in the cyclization and coupling reactions of cyclodextrin glycosyltransferase from Bacillus circulans strain 251 implications for product inhibition and product specificity. AB - Cyclodextrin glycosyltransferase (CGTase) (EC 2.4.1.19) is used for the industrial production of cyclodextrins. Its application, however, is hampered by the limited cyclodextrin product specificity and the strong inhibitory effect of cyclodextrins on CGTase activity. Recent structural studies have identified Arg47 in the Bacillus circulans strain 251 CGTase as an active-site residue interacting with cyclodextrins, but not with linear oligosaccharides. Arg47 thus may specifically affect CGTase reactions with cyclic substrates or products. Here we show that mutations in Arg47 (to Leu or Gln) indeed have a negative effect on the cyclization and coupling activities; Arg47 specifically stabilizes the oligosaccharide chain in the transition state for these reactions. As a result, the mutant proteins display a shift in product specificity towards formation of larger cyclodextrins. As expected, both mutants also showed lower affinities for cyclodextrins in the coupling reaction, and a reduced competitive (product) inhibition of the disproportionation reaction by cyclodextrins. Both mutants also provide valuable information about the processes taking place during cyclodextrin production assays. Mutant Arg47-->Leu displayed an increased hydrolyzing activity, causing accumulation of increasing amounts of short oligosaccharides in the reaction mixture, which resulted in lower final amounts of cyclodextrins produced from starch. Interestingly, mutant Arg47-->Gln displayed an increased ratio of cyclization/coupling and a decreased hydrolyzing activity. Due to the decreased coupling activity, which especially affects the production of larger cyclodextrins, this CGTase variant produced the various cyclodextrins in a stable ratio in time. This feature is very promising for the industrial application of CGTase enzymes with improved product specificity. PMID- 10848959 TI - Kinetic properties of the glucose-6-phosphate and 6-phosphogluconate dehydrogenases from Corynebacterium glutamicum and their application for predicting pentose phosphate pathway flux in vivo. AB - The glucose-6-phosphate (Glc6P) and 6-phosphogluconate (6PG) dehydrogenases of the amino-acid-producing bacterium Corynebacterium glutamicum were purified to homogeneity and kinetically characterized. The Glc6P dehydrogenase was a heteromultimeric complex, which consists of Zwf and OpcA subunits. The product inhibition pattern of the Glc6P dehydrogenase was consistent with an ordered bi bi mechanism. The 6PG dehydrogenase was found to operate according to a Theorell Chance ordered bi-ter mechanism. Both enzymes were inhibited by NADPH and the 6PG dehydrogenase additionally by ATP, fructose 1,6-bisphosphate (Fru1,6P2), D glyceraldehyde 3-phosphate (Gra3P), erythrose 4-phosphate and ribulose 5 phosphate (Rib5P). The inhibition by NADPH was considered to be most important, with inhibition constants of around 25 microM for both enzymes. Intracellular metabolite concentrations were determined in two isogenic strains of C. glutamicum with plasmid-encoded NAD- and NADP-dependent glutamate dehydrogenases. NADP+ and NADPH levels were between 130 microM and 290 microM, which is very much higher than the respective Km and Ki values. The Glc6P concentration was around 500 microM in both strains. The in vivo fluxes through the oxidative part of the pentose phosphate pathway calculated on the basis of intracellular metabolite concentrations and the kinetic constants of the purified enzymes determined in vitro were in agreement with the same fluxes determined by NMR after 13C labelling. From the derived kinetic model thus validated, it is concluded that the oxidative pentose phosphate pathway in C. glutamicum is mainly regulated by the ratio of NADPH and NADP+ concentrations and the specific enzyme activities of both dehydrogenases. PMID- 10848960 TI - Molecular cloning and functional expression of triterpene synthases from pea (Pisum sativum) new alpha-amyrin-producing enzyme is a multifunctional triterpene synthase. AB - Ursane type triterpene is one of the most widespread triterpene aglycones found in plants, together with oleanane type, and these two types often occur together in the same plant. Pisum sativum is known to produce both types of triterpenes. Homology based PCRs with degenerate primers designed from the conserved sequences found in the known beta-amyrin synthases have resulted in cloning of two triterpene synthase cDNAs from immature seeds of P. sativum. They show high sequence identities to each other (78%) and also to the known beta-amyrin synthases (70-90%). ORFs of the full-length clones named as PSY (2277 bp, codes for 759 amino acids) and PSM (2295 bp, codes for 765 amino acids) were ligated into the yeast expression vector pYES2 under the control of GAL1 promoter. Heterologous expression in yeast revealed PSY to be a P. sativum beta-amyrin synthase. Surprisingly, however, PSM turned out to be a novel mixed amyrin synthase producing both alpha- and beta-amyrin. Several minor triterpenes were also identified as the PSM byproducts. The presence of such multifunctional triterpene synthase would account for the co-occurence of ursane and oleanane type triterpenes in plants. PMID- 10848961 TI - The role of individual lysine residues in the basic patch on turnip cytochrome f for electrostatic interactions with plastocyanin in vitro. AB - The role of electrostatic interactions in determining the rate of electron transfer between cytochrome f and plastocyanin has been examined in vitro with mutants of turnip cytochrome f and mutants of pea and spinach plastocyanins. Mutation of lysine residues Lys58, Lys65 and Lys187 of cytochrome f to neutral or acidic residues resulted in decreased binding constants and decreased rates of electron transfer to wild-type pea plastocyanin. Interaction of the cytochrome f mutant K187E with the pea plastocyanin mutant D51K gave a further decrease in electron transfer rate, indicating that a complementary charge pair at these positions could not compensate for the decreased overall charge on the proteins. Similar results were obtained with the interaction of the cytochrome f mutant K187E with single, double and triple mutants of residues in the acidic patches of spinach plastocyanin. These results suggest that the lysine residues of the basic patch on cytochrome f are predominantly involved in long-range electrostatic interactions with plastocyanin. However, analysis of the data using thermodynamic cycles provided evidence for the interaction of Lys187 of cytochrome f with Asp51, Asp42 and Glu43 of plastocyanin in the complex, in agreement with a structural model of a cytochrome f-plastocyanin complex determined by NMR. PMID- 10848962 TI - Sulfhydryls of tubulin. A probe to detect conformational changes of tubulin. AB - The 20 cysteine residues of tubulin are heterogeneously distributed throughout its three-dimensional structure. In the present work, we have used the reactivity of these cysteine residues with 5, 5'-dithiobis(2-nitrobenzoic acid) (DTNB) as a probe to detect the global conformational changes of tubulin under different experimental conditions. The 20 sulfhydryl groups can be classified into two categories: fast and slow reacting. Colchicine binding causes a dramatic decrease in the reactivity of the cysteine residues and causes complete protection of 1.4 cysteine residues. Similarly, other colchicine analogs that bind reversibly initially decrease the rate of reaction; but unlike colchicine they do not cause complete protection of any sulfhydryl groups. Interestingly, in all cases we find that all the slow reacting sulfhydryl groups are affected to the same extent, that is, have a single rate constant. Glycerol has a major inhibitory effect on all these slow reacting sulfhydryls, suggesting that the reaction of slow reacting cysteines takes place from an open state at equilibrium with the native. Ageing of tubulin at 37 degrees C leads to loss of self-assembly and colchicine binding activity. Using DTNB kinetics, we have shown that ageing leads to complete protection of some of the sulfhydryl groups and increased reaction rate for other slow reacting sulfhydryl groups. Ageing at 37 degrees C also causes aggregation of tubulin as indicated by HPLC analysis. The protection of some sulfhydryl groups may be a consequence of aggregation, whereas the increased rate of reaction of other slow reacting sulfhydryls may be a result of changes in global dynamics. CD spectra and acrylamide quenching support such a notion. Binding of 8-anilino-1-naphthalenesulfonate (ANS) and bis-ANS by tubulin cause complete protection of some cysteine residues as indicated by the DTNB reaction, but has little effect on the other slow reacting cysteines, suggesting local effects. PMID- 10848963 TI - Energetics of myo-inositol hexasulfate binding to human acidic fibroblast growth factor effect of ionic strength and temperature. AB - The binding of myo-inositol hexasulfate to an N-terminal truncated 132-amino-acid human acidic fibroblast growth factor form was studied by isothermal titration calorimetry. The technique yields values for the enthalpy change and equilibrium constant, from which the Gibbs energy and entropy change can also be calculated. Experiments in different buffers and pH values show that the proton balance in the reaction is negligible. Experiments at pH 7.0 in the presence of 0.2-0.6 M NaCl showed that the enthalpy and Gibbs energy changes parallel behaviour with ionic strength change, with values in the -21 to -11 kJ x mol(-1) range in the first case and in the -31 to -22 kJ x mol(-1) range in the second. No dependence of entropy on ionic strength was found, with a constant value of approximately 35 J x K(-1) x mol(-1) at all ionic strengths studied. The results can be interpreted in molecular terms by a model in which competitive binding of 3-4 chloride ions to the myo-inositol-binding site is assumed. Isothermal titration calorimetry was also performed at different temperatures and yielded a value of 142+/-13 J x K(-1) x mol(-1) for the heat-capacity change at pH 7.0 and 0.4 M NaCl. Using different parametric equations in the literature, changes on ligand binding in the range -100 to -200 A2 in solvent-accessible surface areas, both polar and apolar, were calculated from thermodynamic data. These values suggest a negligible overall conformational change in the protein when the ligand binds and agree closely with calculations performed with NMR structural data, in which it is shown that the most important negative change in total solvent-accessible surface area occurs in the amino acids Ile56, Gln57, Leu58 and Leu149, in the high-affinity receptor-binding region of the protein. PMID- 10848964 TI - Crystal structure of cambialistic superoxide dismutase from porphyromonas gingivalis. AB - The crystal structure of cambialistic superoxide dismutase (SOD) from Porphyromonas gingivalis, which exhibits full activity with either Fe or Mn at the active site, has been determined at 1.8-A resolution by molecular replacement and refined to a crystallographic R factor of 17.9% (Rfree 22.3%). The crystals belong to the space group P212121 (a = 75.5 A, b = 102.7 A, c = 99.6 A) with four identical subunits in the asymmetric unit. Each pair of subunits forms a compact dimer, but not a tetramer, with 222 point symmetry. Each subunit has 191 amino acid residues most of which are visible in electron density maps, and consists of seven alpha helices and one three-stranded antiparallel beta sheet. The metal ion, a 3 : 1 mixture of Fe and Mn, is coordinated with five ligands (His27, His74, His161, Asp157, and water) arranged at the vertices of a trigonal bipyramid. Although the overall structural features, including the metal coordination geometry, are similar to those found in other single-metal containing SODs, P. gingivalis SOD more closely resembles the dimeric Fe-SODs from Escherichia coli rather than another cambialistic SOD from Propionibacterium shermanii, which itself is rather similar to other tetrameric SODs. PMID- 10848965 TI - Trypsin-specific acyl-4-guanidinophenyl esters for alpha-chymotrypsin-catalysed reactions computational predictions, hydrolyses, and peptide bond formation. AB - The function of acyl-4-guanidinophenyl esters as substrate mimetics for the serine protease alpha-chymotrypsin was investigated by protein-ligand docking, hydrolysis, and acyl transfer experiments. On the basis of protein-ligand docking studies, the binding and hydrolysis properties of these artificial substrates were estimated. The predictions of the rational approach were confirmed by steady state hydrolysis studies on 4-guanidinophenyl esters derived from coded amino acids (which alpha-chymotrypsin is not specific for), noncoded amino acids, and even simple carboxylic acid moieties. Enzymatic peptide syntheses qualify these esters as suitable acyl donors for the coupling of acyl components far from the natural enzyme specificity, thus considerably expanding the synthetic utility of alpha-chymotrypsin. PMID- 10848966 TI - A DNA ligase from the psychrophile Pseudoalteromonas haloplanktis gives insights into the adaptation of proteins to low temperatures. AB - The cloning, overexpression and characterization of a cold-adapted DNA ligase from the Antarctic sea water bacterium Pseudoalteromonas haloplanktis are described. Protein sequence analysis revealed that the cold-adapted Ph DNA ligase shows a significant level of sequence similarity to other NAD+-dependent DNA ligases and contains several previously described sequence motifs. Also, a decreased level of arginine and proline residues in Ph DNA ligase could be involved in the cold-adaptation strategy. Moreover, 3D modelling of the N terminal domain of Ph DNA ligase clearly indicates that this domain is destabilized compared with its thermophilic homologue. The recombinant Ph DNA ligase was overexpressed in Escherichia coli and purified to homogeneity. Mass spectroscopy experiments indicated that the purified enzyme is mainly in an adenylated form with a molecular mass of 74 593 Da. Ph DNA ligase shows similar overall catalytic properties to other NAD+-dependent DNA ligases but is a cold adapted enzyme as its catalytic efficiency (kcat/Km) at low and moderate temperatures is higher than that of its mesophilic counterpart E. coli DNA ligase. A kinetic comparison of three enzymes adapted to different temperatures (P. haloplanktis, E. coli and Thermus scotoductus DNA ligases) indicated that an increased kcat is the most important adaptive parameter for enzymatic activity at low temperatures, whereas a decreased Km for the nicked DNA substrate seems to allow T. scotoductus DNA ligase to work efficiently at high temperatures. Besides being useful for investigation of the adaptation of enzymes to extreme temperatures, P. haloplanktis DNA ligase, which is very efficient at low temperatures, offers a novel tool for biotechnology. PMID- 10848967 TI - Activation of phosphatidylinositol 3-kinase by a complex of p59fyn and the receptor tyrosine kinase Xmrk is involved in malignant transformation of pigment cells. AB - Malignant melanoma in the fish Xiphophorus is induced by overexpression of the Xmrk-oncogene, encoding a subclass I receptor tyrosine kinase. The mutationally activated Xmrk protein triggers constitutive mitogenic signalling in fish melanoma cells. In recent studies we showed that in melanoma cells phosphatidylinositol (PtdIns) 3-kinase, as well as p59fyn, has elevated levels of kinase activity. Both bind directly to different phosphotyrosine residues in the Xmrk receptor C-terminus through their SH2 domains. To analyse the mechanism of regulation of these Xmrk-associated kinases in melanoma we characterized the protein-protein interactions between PtdIns 3-kinase, p59fyn and the Xmrk receptor in detail. A ternary complex in which the p85 subunit of PtdIns 3-kinase is associated with p59fyn as well as with Xmrk was identified. Contrary to complexes described for other receptors, the adaptor protein p120Cbl was not involved in these interactions. Thus, we describe here a new mechanism of activation of PtdIns 3-kinase by a receptor of the epidermal growth factor receptor family in which p59fyn acts as an adaptor as well as an activator of PtdIns 3-kinase. Activation of PtdIns 3-kinase activity by fyn was also found in vivo. The fact that this was only detectable in highly transformed Xmrk overexpressing melanomas but not in benign lesions points to the essential role of the Xmrk receptor in this mechanism of regulation. PMID- 10848968 TI - Regulation of -1 ribosomal frameshifting directed by cocksfoot mottle sobemovirus genome. AB - The polyprotein of Cocksfoot mottle virus (CfMV) is encoded by two overlapping open reading frames (ORFs). The putative replicase of CfMV is produced as a part of the polyprotein from ORF2b by the -1 ribosomal frameshifting mechanism. The signals leading to -1 ribosomal frameshifting directed by CfMV RNA are the slippery heptamer UUUAAAC and a stem-loop structure starting seven nucleotides downstream from the heptamer. We studied the effect of different parts of the CfMV genome on the -1 ribosomal frameshifting efficiency using a wheat germ extract transcription/translation system. A point mutation in the slippery heptamer and a mutation deleting the stem-loop structure prevented frameshifting. Seventy nucleotides of CfMV sequence, including the slippery sequence and the stem-loop structure, was found to act as a minimal region for frameshifting. Interestingly, a termination codon introduced into the -1-frame 27 nucleotides downstream of the stem-loop structure increased frameshift efficiency threefold, while a similarly located termination codon in the 0-frame had no effect. Even fourfold to fivefold efficiencies were observed when the polyprotein encoding ORFs were fused together, which led simultaneously to the formation of a termination codon downstream of the frameshift signal. Possible reasons underlying these observations are discussed. PMID- 10848969 TI - Structural requirements for thymosin beta4 in its contact with actin. An NMR analysis of thymosin beta4 mutants in solution and correlation with their biological activity. AB - We examined the conformational preferences of mutants of thymosin beta4, an actin monomer sequestering protein by NMR spectroscopy in 60% (v/v) trifluoroethanol. Under these conditions, the wild-type thymosin beta4 conformation consists of an alpha-helix (helix I) extending from residues 5-16 with a more stable fragment from lysine 11 to lysine 16 and a second alpha-helix (helix II) encompassing residues 31-39. The point mutations studied here are located in helix I or in the LKKTET segment (residues 17-22) that form the two main entities of interaction with the actin molecule. The alpha-1H conformational shifts allow us to investigate the helicity of the polypeptides at the residue level and to correlate these structures with their biological activity. We determine that an extension of helix I at its C-terminal end over the LKK-segment results in loss of activity. The correct termination of this helix is connected to a specific orientation of the polypeptide essential for a cooperative action of the thymosin beta4 binding entities required for full activity. PMID- 10848970 TI - Folding of a beta-hairpin peptide derived from the N-terminus of ubiquitin. Conformational preferences of beta-turn residues dictate non-native beta-strand interactions. AB - The role of the non-native beta-turn sequence (NPDG) in nucleating the folding of a beta-hairpin peptide derived from the N-terminus of ubiquitin, has been examined by NMR and CD spectroscopy. The NPDG sequence, while representing a common two-residue type I turn sequence in proteins, folds to give a G1-bulged type I turn in the context of a beta-hairpin peptide, to the exclusion of other possible conformations. The turn conformation results in misalignment of the two beta strands and a beta hairpin with non-native side chain interactions. A truncated 12-residue analogue of the hairpin, in which the majority of residues in the N-terminal beta strand have been deleted, shows some weak propensity to fold into a G-bulged type I turn conformation in the absence of interstrand stabilizing interactions. The NPDG turn sequence pays some of the entropic cost in initiating folding allowing interstrand interactions, which in this case arise from the non-native pairing of residue side chains, to stabilize a significant population of the folded state. Examination of the relative abundance of the Pro Asp type I turn, with G in the +B1 position, vs. the type I G-bulged turn PXG, in a database of high resolution structures, reveals 48 instances of PXG bulged turns for which X = Asp is by far the most common residue with 20 occurrences. Strikingly, there are no examples of a type I PD turn with G at the +B1 position, in good agreement with our experimental observations that the PDG G-bulged turn is populated preferentially in solution. PMID- 10848971 TI - Zwitterionic and acidic glycosphingolipids of the Drosophila melanogaster embryo. AB - Defining glycosphingolipid structures in species amenable to genetic manipulation, such as Drosophila melanogaster, provides a foundation for investigating mechanisms that regulate glycolipid expression. Therefore, eight of the 12 major glycosphingolipids, accounting for 64% of lipid-linked carbohydrate in Drosophila embryos, were purified after separation into acidic and zwitterionic pools. The zwitterionic lipids possess phosphoethanolamine (PEtn) linked to one or more GlcNAc residues and comprise a family of serially related structures. The longest characterized glycolipid, an octaosylceramide, designated Nz28, has the structure: GalNAcbeta, 4(PEtn 6)GlcNAcbeta,3Galbeta,3GalNAcalpha,4Ga lNAcbeta, 4(PEtn 6)GlcNAcbeta,3Manbeta,4GlcbetaCer. Heptaosyl (Nz7), hexaosyl (Nz6), pentaosyl (Nz5) and tetraosyl (Nz4) forms of Nz28, sequentially truncated from the nonreducing terminus, possess only one PEtn moiety. The major acidic lipid, designated Az29, possesses two PEtn moieties and a glucuronic acid linked to a Gal-extended Nz28. Two other acidic glycolipids, Az9 and Az6, exhibit one PEtn moiety and the same hexose and N-acetylhexosamine composition as Az29 and Nz6, respectively. The fully extended Drosophila core oligosaccharide differs from that of other dipterans in the linkage at a single glycosidic bond, a distinction with significant structural and biosynthetic consequences. Furthermore, acidic species account for a larger proportion of total glycosphingolipid, and PEtn substitution of GlcNAc is more complete in the Drosophila embryo. Divergent characteristics may reflect interspecies variation or stage-specific glycosphingolipid expression in dipterans. PMID- 10848972 TI - Transcriptional control of adrenomedullin induction by phorbol ester in human monocytic leukemia cells. AB - Adrenomedullin is a potent vasodilator peptide that was originally identified from human pheochromocytoma. In this study, we investigated the induction of adrenomedullin gene expression in THP-1 acute monocytic leukemia cells during differentiation into macrophage-like cells by 12-O-tetradecanoylphorbol-13 acetate (TPA), and identified a cis-regulatory region of the human adrenomedullin gene responsible for TPA-induced adrenomedullin expression. Upon treatment with TPA (100 ng x mL(-1)) for 24 h, immunoreactive adrenomedullin concentrations in the culture medium and adrenomedullin mRNA levels were increased more than 10 fold, concomitant with the differentiation of THP-1 cells into macrophage-like cells. Actinomycin D abolished the TPA-induced adrenomedullin expression, indicating that the induction of ADM gene expression by TPA was regulated at the transcriptional level. Transient transfection assay revealed that a cis-acting region (positions -70 to -30) of human adrenomedullin gene was necessary for TPA induced reporter gene expression. This region contains multiple copies of activator protein 2 (AP-2) binding sites, which are bound by purified AP-2 protein, as judged by electrophoretic mobility shift assay. The binding activity to this region was undetectable in nuclear extracts prepared from untreated THP-1 cells, but was increased in extracts prepared from TPA-treated cells. The protein binding was abolished by unlabeled oligonucleotides containing the AP-2 consensus sequence. These results indicate that the region (-70 to -30) of the human ADM gene containing multiple AP-2 binding sites is responsible for TPA-induced adrenomedullin expression in THP-1 cells. PMID- 10848973 TI - Identification of hypoxia-responsive messengers expressed in human microvascular endothelial cells using differential display RT-PCR. AB - In order to identify new genes overexpressed in endothelial cells exposed to hypoxia, differential display RT-PCR was performed on total RNA extracted from human microvascular endothelial cells incubated under hypoxia or under normoxic conditions. Northern blot and reverse Northern blot analyses were used to confirm the results. Sequences corresponding to tissue inhibitor of metalloproteinase-1, prostate tumor inducing factor-1, enolase-alpha and prothymosin-alpha were evidenced as overexpressed in hypoxia. These results were confirmed by Western blot and immunofluorescence experiments. Moreover, several elements homologous to partial sequences of cDNA (expressed sequence tag) were also identified, as well as unknown cDNA sequences. The present study suggests that hypoxia can change the expression of numerous genes in endothelial cells, and that mRNA differential display is useful for cloning known and unknown hypoxia-responsive genes. PMID- 10848974 TI - Assembly of heterodimeric luciferase after de novo synthesis of subunits in rabbit reticulocyte lysate involves hsc70 and hsp40 at a post-translational stage. AB - Heterodimeric luciferase from Vibrio harveyi had been established as a unique model enzyme for direct measurements of the effects of molecular chaperones and folding catalysts on protein folding and subunit assembly after de novo synthesis of subunits in rabbit reticulocyte lysate. It was observed that luciferase assembly can be separated in time from synthesis of the two subunits and that under these post-translational conditions assembly was inhibited by either ATP depletion or inhibition of peptidylprolyl cis/trans isomerases, that is, by addition of cyclosporin A or FK506. Furthermore, it was observed that the inhibitory effect of FK506 on luciferase assembly can be suppressed by addition of purified cyclophilin, thereby providing the first direct evidence for the involvement of peptidylprolyl cis/trans isomerases in protein biogenesis in the eukaryotic cytosol. Here the ATP requirement in luciferase assembly has been characterized. Depletion of either Hsp90 or CCT from reticulocyte lysate did not interfere with luciferase assembly. However, addition of purified Hsc70 stimulated luciferase assembly. While addition of purified Hsp40 did not have any effect on luciferase assembly, the stimulatory effect of Hsc70 was further increased by Hsp40. Thus, after synthesis of the two subunits in reticulocyte lysate assembly of heterodimeric luciferase involves Hsc70 and its co-chaperone Hsp40. Therefore, Hsc70 aids protein biogenesis in the eukaryotic cytosol not only at the levels of nascent polypeptide chains and precursor proteins that have to be kept competent for transport into cell organelles, but also at the level of subunits that have to be kept competent for assembly. PMID- 10848975 TI - Kinetic properties of the 2-oxoglutarate dehydrogenase complex from Azotobacter vinelandii evidence for the formation of a precatalytic complex with 2 oxoglutarate. AB - The 2-oxoglutarate dehydrogenase complex was purified from Azotobacter vinelandii. The complex consists of three components, 2-oxoglutarate dehydrogenase/decarboxylase (E1o), lipoate succinyltransferase (E2o) and lipoamide dehydrogenase (E3). Upon purification, the E3 component dissociates partially from the complex. From reconstitution experiments, the Kd for E3 was found to be 26 nM, about 30 times higher than that for the pyruvate dehydrogenase complex. The Km values for the substrates 2-oxoglutarate, CoA and NAD+ were found to be 0.15, 0.014 and 0.17 mM, respectively. The system has a high specificity for 2-oxoglutarate, which is determined by the action of both E1o and E2o. Above 4 mM substrate inhibition is observed. From steady-state inhibition experiments with substrate analogs, two substrate-binding modes are revealed at different degrees of saturation of the enzyme with 2-oxoglutarate. At low substrate concentrations (10(-6) to 10(-5) M), the binding mainly depends on the interaction of the enzyme with the substrate carboxyl groups. At a higher degree of substrate saturation (10(-4) to 10(-3) M) the relative contribution of the 2 oxo group in the binding increases. A kinetic analysis points to a single binding site for a substrate analog under steady state conditions. Saturation of this site with an analog indicates that two kinetically different complexes are formed with 2-oxoglutarate in the course of catalysis. From competition studies with analogs it is concluded that one of these complexes is formed at the site that is sterically identical to the substrate inhibition site. The data obtained are represented by a minimal scheme that considers formation of a precatalytic complex SE between the substrate and E1o before the catalytic complex ES, in which the substrate is added to the thiamin diphosphate cofactor, is formed. The incorrect orientation of the substrate molecule in SE or the occupation of this site by analogs is supposed to cause substrate or analog inhibition, respectively. PMID- 10848976 TI - Targeting of single-stranded DNA and RNA containing adjacent pyrimidine and purine tracts by triple helix formation with circular and clamp oligonucleotides. AB - The aim of this work was to construct an anti-messenger targeted to the pim-1 oncogene transcript, based on circular or clamp oligodeoxyribonucleotides. The formation of bimolecular triplexes by clamp or circular oligonucleotides was investigated using single-stranded targets of both DNA (5'-CCCTCCTTTGAAGAA-3') and RNA type (5'-CCCUCCUUUGAAGAA-3'). The third, 'Hoogsteen' strand of the triplex was represented by G,T-rich sequences. The secondary structures of the complexes were determined by thermal denaturation, circular dichroism and gel mobility shift experiments and shown to depend on the nature of the target strand. With DNA as target, the sequence of a clamp (or circular) oligonucleotide that formed the triple helix was 3'-GGGAGGAAACTTCTTTT-TTGTTGTTT-TT-GGTGGG-5', where the first TT dinucleotide (in italics) is a linker and the second TT (bold) represents the bridge through which the 'Hoogsteen' strand switches from one strand of the Watson-Crick duplex to the other, once the duplex is formed by the corresponding portion of the anti-messenger (underlined). The portion of the 'Hoogsteen' sequence of the triplex between the two TT dinucleotides binds to the 3' extremity of the target strand and runs parallel to it. The portion situated at the 5' end of the oligonucleotide switches to the purine tract of the complementary strand of the duplex and is antiparallel to it. In contrast, with RNA as target, for a branched clamp oligonucleotide that formed a triple helix over its entire length (5'-TTCTTCAAAGGAGGG-3' 3'-GGGTGGTTT-T-GTTGTT-5') the portion of the 'Hoogsteen' sequence that bound to the 3' extremity of the target strand had to be antiparallel to it. PMID- 10848977 TI - Human herpes virus 8 interleukin-6 homologue triggers gp130 on neuronal and hematopoietic cells. AB - Human herpes virus-8 (HHV8) encodes a cytokine named viral interleukin-6 (vIL-6) that shares 25% amino-acid identity with its human homologue. Human IL-6 is known to be a growth and differentiation factor of lymphatic cells and plays a potential role in the pathophysiology of various lymphoproliferative diseases. vIL-6 is expressed in HHV8-associated-diseases including Kaposi's sarcoma, Body cavity-based-lymphoma and Castleman's disease, suggesting a pathogenetic involvement in the malignant growth of B-cell associated diseases and other malignant tumours. We expressed vIL-6 in Escherichia coli as a fusion protein with recombinant periplasmic maltose binding protein. After cleavage from the maltose binding protein moiety and purification, vIL-6 was shown to be correctly folded using circular dichroism spectroscopy. A rabbit antiserum was raised against the recombinant vIL-6 protein. vIL-6 turned out to be active on cells that expressed gp130 but no IL-6 receptor (IL-6-R) suggesting that, in contrast to human IL-6, vIL-6 stimulated gp130 directly. Accordingly, vIL-6 activity could be inhibited by a soluble gp130 Fc Fusion protein. vIL-6 was shown to induce neuronal differentiation of rat pheochromocytoma cells and to stimulate colony formation of human hematopoietic progenitor cells. Thus, vIL-6 exhibits biologic activity that has only been observed for the IL-6/soluble IL-6-R complex but not for IL-6 alone. These properties are important for the evaluation of the pathophysiological potential of vIL-6. PMID- 10848978 TI - Structure-function relationships in Drosophila melanogaster alcohol dehydrogenase allozymes ADH(S), ADH(F) and ADH(UF), and distantly related forms. AB - Drosophila melanogaster alcohol dehydrogenase (ADH), a paradigm for gene-enzyme molecular evolution and natural selection studies, presents three main alleloforms (ADHS, ADHF and ADHUF) differing by one or two substitutions that render different biochemical properties to the allelozymes. A three-dimensional molecular model of the three allozymes was built by homology modeling using as a template the available crystal structure of the orthologous D. lebanonensis ADH, which shares a sequence identity of 82.2%. Comparison between D. lebanonensis and D. melanogaster structures showed that there is almost no amino-acid change near the substrate or coenzyme binding sites and that the hydrophobic active site cavity is strictly conserved. Nevertheless, substitutions are not distributed at random in nonconstricted positions, or located in external loops, but they appear clustered mainly in secondary structure elements. From comparisons between D. melanogaster allozymes and with D. simulans, a very closely related species, a model based on changes in the electrostatic potential distribution is presented to explain their differential behavior. The depth of knowledge on Drosophila ADH genetics and kinetics, together with the recently obtained structural information, could provide a better understanding of the mechanisms underlying molecular evolution and population genetics. PMID- 10848979 TI - The blue-colored linker polypeptide L55 is a fusion protein of phycobiliproteins in the cyanobacterium synechocystis sp. strain BO 8402. AB - The cyanobacterium Synechocystis sp. strain BO 8402, isolated from Lake Constance, lacks phycobilisomes but instead forms inclusion bodies containing remnants of phycobiliproteins. The inclusion bodies are surrounded by a proteinaceous capsule and contain alpha-phycocyanin and beta-phycocyanin, the rod linker polypeptide L35RPC and a novel blue-colored protein L55 with an apparent molecular mass of 55 kDa. An antibody raised against beta-phycocyanin showed a strong cross-reaction with L55. Mass spectrometry analysis of proteolytic peptides from L55 revealed mass identity to proteolytic peptides derived from L35RPC and beta-phycocyanin. However, analysis of the genome of strain BO 8402 revealed only one cpcBACE operon, encoding the apoproteins of beta-phycocyanin and alpha-phycocyanin, L35RPC and a subunit of the phycocyanin alpha subunit phycocyanobilin lyase, respectively. The gene structure, sequence and transcription of these genes were identical to that of a revertant strain, Synechocystis sp. strain BO 9201, which formed phycobilisomes and did not express L55. Based on these observations, we concluded that L55 did not derive from a particular gene or from a special form of mRNA-processing. We propose that L55 is formed by post-translational fusion of L35RPC and beta-phycocyanin. Cross-linking may stabilize the formation of the large paracrystalline phycocyanin aggregates unique to Synechocystis sp. strain BO 8402. PMID- 10848980 TI - Inhibition by unsaturated fatty acids of type II secretory phospholipase A2 synthesis in guinea-pig alveolar macrophages evidence for the eicosanoid independent pathway. AB - The effect of arachidonic acid (C20:4) on the production of secretory type II phospholipase A2 (sPLA2-II) by guinea-pig alveolar macrophages was investigated. We show that incubation of these cells with 1-30 microM of arachidonic acid inhibits the synthesis of sPLA2-II in a concentration-dependent manner with an IC50 of approximately 7.5 microM. The inhibition by low concentrations (5 microM) of arachidonic acid was partially reduced by pretreatment of alveolar macrophages with cyclooxygenase or cytochrome P450 inhibitors (aspirin and 1 aminobenzotriazole, respectively), but not by lipoxygenase inhibitor, BW A4C. However, these inhibitors failed to interfere with the effect of high concentrations (30 microM) of arachidonic acid, suggesting that the latter may act on the expression of sPLA2-II, at least in part, independently of eicosanoid generation. Indeed, a similar inhibitory effect on sPLA2-II activity and mRNA expression was observed with other unsaturated fatty acids such as eicosapentaenoic (C20:5) and oleic (C18:1) acids, but not with the saturated fatty acid, palmitic acid (C16:0). In addition, arachidonic acid partially reduced the secretion of tumor necrosis factor alpha, an important intermediate in the induction of sPLA2-II synthesis by guinea-pig alveolar macrophages. However, addition of recombinant tumor necrosis factor alpha failed to reverse the inhibitory effect of arachidonic acid on sPLA2-II expression, suggesting that this process occurs downstream of tumor necrosis factor alpha secretion. We conclude that the expression of sPLA2-II in alveolar macrophages is down regulated at the transcriptional level by arachidonic acid either directly or via its cyclooxygenase and cytochrome P450-derived metabolites. PMID- 10848981 TI - On the iron-sulfur clusters in the complex redox enzyme dihydropyrimidine dehydrogenase. AB - Porcine liver dihydropyrimidine dehydrogenase is a homodimeric iron-sulfur flavoenzyme that catalyses the first and rate-limiting step of pyrimidine catabolism. The enzyme subunit contains 16 atoms each of nonheme iron and acid labile sulfur, which are most likely arranged into four [4Fe-4S] clusters. However, the presence and role of such Fe-S clusters in dihydropyrimidine dehydrogenase is enigmatic, because they all appeared to be redox-inactive during absorbance-monitored titrations of the enzyme with its physiological substrates. In order to obtain evidence for the presence and properties of the postulated four [4Fe-4S] clusters of dihydropyrimidine dehydrogenase, a series of EPR monitored redox titrations of the enzyme under a variety of conditions was carried out. No EPR-active species was present in the enzyme 'as isolated'. In full agreement with absorbance-monitored experiments, only a small amount of neutral flavin radical was detected when the enzyme was incubated with excess NADPH or dihydrouracil under anaerobic conditions. Reductive titrations of dihydropyrimidine dehydrogenase with dithionite at pH 9.5 and photochemical reduction at pH 7.5 and 9.5 in the presence of deazaflavin and EDTA led to the conclusion that the enzyme contains two [4Fe-4S]2+,1+ clusters, which both exhibit a midpoint potential of approximately -0.44 V (pH 9.5). The two clusters are most likely close in space, as demonstrated by the EPR signals which are consistent with dipolar interaction of two S = 1/2 species including a half-field signal around g approximately 3.9. Under no circumstances could the other two postulated Fe-S centres be detected by EPR spectroscopy. It is concluded that dihydropyrimidine dehydrogenase contains two [4Fe-4S] clusters, presumably determined by the C-terminal eight-iron ferredoxin-like module of the protein, whose participation in the enzyme-catalysed redox reaction is unlikely in light of the low midpoint potential measured. The presence of two additional [4Fe-4S] clusters in dihydropyrimidine dehydrogenase is proposed based on thorough chemical analyses on various batches of the enzyme and sequence analyses. The N terminal region of dihydropyrimidine dehydrogenase is similar to the glutamate synthase beta subunit, which has been proposed to contain most, if not all, the cysteinyl ligands that participate in the formation of the [4Fe-4S] clusters of the glutamate synthase holoenzyme. It is proposed that the motif formed by the Cys residues at the N-terminus of the glutamate synthase beta subunit, which are conserved in dihydropyrimidine dehydrogenase and in several beta-subunit-like proteins or protein domains, corresponds to a novel fingerprint that allows the formation of [4Fe-4S] clusters of low to very low midpoint potential. PMID- 10848982 TI - Sulfate assimilation in higher plants characterization of a stable intermediate in the adenosine 5'-phosphosulfate reductase reaction. AB - The enzyme catalysing the reduction of adenosine 5'-phosphosulfate (AdoPS) to sulfite in higher plants, AdoPS reductase, is considered to be the key enzyme of assimilatory sulfate reduction. In order to address its reaction mechanism, the APR2 isoform of this enzyme from Arabidopsis thaliana was overexpressed in Escherichia coli and purified to homogeneity. Incubation of the enzyme with [35S]AdoPS at 4 degrees C resulted in radioactive labelling of the protein. Analysis of APR2 tryptic peptides revealed 35SO2-3 bound to Cys248, the only Cys conserved between AdoPS and prokaryotic phosphoadenosine 5'-phosphosulfate reductases. Consistent with this result, radioactivity could be released from the protein by incubation with thiols, inorganic sulfide and sulfite. The intermediate remained stable, however, after incubation with sulfate, oxidized glutathione or AdoPS. Because truncated APR2, missing the thioredoxin-like C terminal part, could be labelled even at 37 degrees C, and because this intermediate was more stable than the complete protein, we conclude that the thioredoxin-like domain was required to release the bound SO2-3 from the intermediate. Taken together, these results demonstrate for the first time the binding of 35SO2-3 from [35S]AdoPS to AdoPS reductase and its subsequent release, and thus contribute to our understanding of the molecular mechanism of AdoPS reduction in plants. PMID- 10848983 TI - Interactions of the C-terminus of viral protein R with nucleic acids are modulated by its N-terminus. AB - The basic viral protein R (Vpr) performs several functions during the human immunodeficiency virus HIV-1 retroviral cycle, including G2 mitosis arrest and nuclear import of the preintegration complex allowing lentivirus to replicate in nondividing cells. Accordingly, this protein was found in the nucleus of infected cells. In the virus, Vpr is incorporated through interaction with both nucleocapsid protein 7 (NCp7) and p6, two small proteins encoded by the C terminal part of the Gag precursor. NCp7 is also involved in genomic RNA encapsidation during the budding process suggesting a possible interaction of Vpr with nucleic acids, either directly or via the NCp7 intermediate. Gel shift experiments were carried out with RNA and DNA using synthetic Vpr and peptide derivatives. The results show that Vpr binds to nucleic-acid inducing aggregates. This process, which requires the C-terminal basic domain of the protein (in particular the helical 70-80 domain), is regulated by the N-terminal region of Vpr. Moreover, NCp7 was shown to enhance RNA recognition by Vpr, a feature that could be required for Vpr encapsidation and during nuclear import of the preintegration complex. PMID- 10848984 TI - A novel NADPH:diamide oxidoreductase activity in arabidopsis thaliana P1 zeta crystallin. AB - The zeta-crystallin (ZCr) gene P1 of Arabidopsis thaliana, known to confer tolerance toward the oxidizing drug 1,1'-azobis(N, N-dimethylformamide) (diamide) to yeast [Babiychuk, E., Kushnir, S., Belles-Boix, E., Van Montagu, M. & Inze, D. (1995) J. Biol. Chem. 270, 26224], was expressed in Escherichia coli to characterize biochemical properties of the P1-zeta-crystallin (P1-ZCr). Recombinant P1-ZCr, a noncovalent dimer, showed NADPH:quinone oxidoreductase activity with specificity to quinones similar to that of guinea-pig ZCr. P1-ZCr also catalyzed the divalent reduction of diamide to 1,2-bis(N,N dimethylcarbamoyl)hydrazine, with a kcat comparable with that for quinones. Two other azodicarbonyl compounds also served as substrates of P1-ZCr. Guinea-pig ZCr, however, did not catalyze the azodicarbonyl reduction. Hence, plant ZCr is distinct from mammalian ZCr, and can be referred to as NADPH:azodicarbonyl/quinone reductase. The quinone-reducing reaction was accompanied by radical chain reactions to produce superoxide radicals, while the azodicarbonyl-reducing reaction was not. Specificity to NADPH, as judged by kcat/Km, was > 1000-fold higher than that to NADH both for quinones and diamide. N-Ethylmaleimide and p-chloromercuribenzoic acid inhibited both quinone-reducing and diamide-reducing activities. Both NADPH and NADP+ suppressed the inhibition, but NADH did not, suggesting that sulfhydryl groups reside in the binding site for the phosphate group on the adenosine moiety of NADPH. The diamide-reducing activity of P1-ZCr accounts for the tolerance of P1-overexpressing yeast to diamide. Other possible physiological functions of P1-ZCr in plants are discussed. PMID- 10848985 TI - Analysis of the domain structure and the DNA binding site of the transcriptional activator FhlA. AB - FhlA is the transcriptional activator of the genes coding for the formate hydrogen lyase system in Escherichia coli. It is activated by the binding of formate and induces transcription by sigma54 RNA polymerase after binding to specific upstream activating sequences (UAS). Sequence comparison had shown that FhlA exhibits a structure composed of three domains, which is typical for sigma54 dependent regulators. By analyzing the N-terminal domain of FhlA of E. coli (amino acids 1-378; FhlA-N) and the rest of the protein (amino acids 379-693; FhlA-C) as separate proteins in vivo and in vitro the functions of the different domains of FhlA were elucidated. The FhlA-C domain is active in ATP hydrolysis and activation of transcription and its activity is neither influenced by the presence of formate nor of the antiactivator HycA. However, it is stimulated in the presence of the FhlA-specific UAS, indicating that this region of FhlA is responsible for DNA binding. FhlA-N is not active itself but able to reduce the activity of full-length FhlA in trans, probably by formation of nonfunctional heterooligomers. The DNA binding site of FhlA was analyzed by hydroxyradical footprinting. Each UAS consists of two binding sites of 16 bp separated by a spacer region. A consensus sequence could be deduced and a model is presented and supported by in vivo data in which a FhlA tetramer binds to the UAS on one side of the DNA helix. Performing an extensive screening we could show that the FhlA regulatory system is conserved in different species of the family Enterobacteriaceae. The analysis of orthologs of FhlA revealed that they are able to functionally replace the E. coli enzyme. PMID- 10848986 TI - Isolation and characterization of RNA aptamers specific for the hepatitis C virus nonstructural protein 3 protease. AB - Nonstructural protein 3 (NS3) from hepatitis C virus (HCV) is a serine protease that provides an essential function in maturation of the virus by cleaving the nonstructural regions of the viral polyprotein. The goal of this work was to isolate RNA aptamers that bind specifically to the NS3 protease active site in the truncated polypeptide DeltaNS3. RNA aptamers were selected in vitro by systematic evolution of ligands by exponential enrichment (SELEX). The RNA pool for SELEX had a 30-nucleotide randomized core region. After nine selection cycles, a pool of DeltaNS3-specific RNA aptamers were obtained. This RNA pool included 45 clones that divided into three main classes (G9-I, II and III). These classes include the conserved sequence GA(A/U)UGGGAC. These aptamers bind to DeltaNS3 with a binding constant of about 10 nM and inhibit approximately 90% of the protease activity of DeltaNS3 and MBP-NS3 (full-length of NS3 fused with maltose binding protein). In addition, these aptamers inhibited approximately 70% of the MBP-NS3 protease activity in the presence of the NS4A peptide P41. G9-I aptamer appeared to be a noncompetitive inhibitor for DeltaNS3 with a Ki approximately 100 nM in the presence of P41. These results suggest that the pool of selected aptamers have potential as anti-HCV compounds. Mutational analysis of the G9-I aptamer demonstrated that the sequences required for protease inhibition are in stem I, stem III and loop III of the aptamer. These regions include the conserved sequence GA(A/U)UGGGAC. PMID- 10848987 TI - Activated phenoloxidase from Tenebrio molitor larvae enhances the synthesis of melanin by using a vitellogenin-like protein in the presence of dopamine. AB - One of the biological functions of activated phenoloxidase in arthropods is the synthesis of melanin around invaded foreign materials. However, little is known about how activated phenoloxidase synthesizes melanin at the molecular level. Even though it has been suggested that the quinone derivatives generated by activated phenoloxidase might use endogenous protein components for melanin synthesis in arthropods, there is no report of protein components engaged in melanin synthesis induced by activated phenoloxidase. In this study, to isolate and characterize proteins involved in melanin synthesis, we prepared in vitro prophenoloxidase activating solution (designated G-100 solution), specifically showing phenoloxidase activity in the presence of Ca2+ and beta-1, 3-glucan, from the hemolymph of larvae of the coleopteran Tenebrio molitor by using a Sephadex G 100 column. When G-100 solution was incubated with dopamine to induce melanin synthesis in the presence of Ca2+ and beta-1,3-glucan, four types of protein (160 kDa, prophenoloxidase, phenoloxidase and 45 kDa) disappeared from SDS/PAGE under reducing conditions. Under identical conditions, but including phenylthiourea as a phenoloxidase inhibitor added to the G-100 solution, three of these proteins (160 kDa, phenoloxidase and 45 kDa) did not disappear. To characterize these melanization-engaging proteins, we first purified the 160-kDa melanization engaging protein to homogeneity and raised a polyclonal antibody against it. Analysis of the cDNA revealed that it consisted of 1439 amino-acid residues and showed partial homology with Caenorhabditis elegans vitellogenin precursor-6 (19.7%). Western blot analysis showed that it disappeared when active phenoloxidase induced melanin synthesis. Furthermore, when the purified 160-kDa melanization-engaging protein was added to a G-100 solution deficient in it, melanin synthesis was enhanced compared with the same solution without the protein. These data support the conclusion that the 160-kDa vitellogenin-like protein is involved in arthropod melanin synthesis. PMID- 10848988 TI - Expression and characterization of recombinant human eosinophil peroxidase. Impact of the R286H substitution on the biosynthesis and activity of the enzyme. AB - Hereditary eosinophil peroxidase deficiency is a genetic abnormality characterized by a decrease or absence of peroxidase activity and a reduction of the granule matrix volume. Recently, we identified two mutations associated with eosinophil peroxidase deficiency in a subject and his siblings, i.e. a base insertion causing the appearance of a premature stop codon and a base transition causing the replacement of an Arg at codon 286 with a His (R286H). In this article we report the stable expression of both the recombinant wild-type and the R286H eosinophil peroxidase precursor in the K-562 cell line, and the effects of the R286H substitution on the structure and function of the eosinophil peroxidase precursor. Heme group incorporation into both the recombinant wild-type and the recombinant R286H eosinophil peroxidase precursor was comparable, as was the stability of both proteins. Instead, the recombinant R286H eosinophil peroxidase precursor exhibited marked alterations of the catalytic properties and an increased sensitivity to four peroxidase inhibitors with respect to both the recombinant wild-type eosinophil peroxidase precursor and the native enzyme. In addition, the recombinant wild-type, but not the R286H, eosinophil peroxidase precursor was immunoprecipitated by two anti-(eosinophil peroxidase) mAbs. Altogether, our results suggest a protein misfolding of the R286H eosinophil peroxidase precursor which might account for its altered catalytic properties and the absence of expression of some epitopes. PMID- 10848989 TI - Molecular characterization of the thermosensitive E1 ubiquitin-activating enzyme cell mutant A31N-ts20. Requirements upon different levels of E1 for the ubiquitination/degradation of the various protein substrates in vivo. AB - According to our current knowledge, protein ubiquitination involves three steps: activation of ubiquitin through formation of an energy-rich bond with an E1 ubiquitin-activating enzyme; and transfer of activated ubiquitin onto E2 ubiquitin-conjugating enzymes, which, in turn, alone, or in combination with E3 ubiquitin-protein ligase enzymes, transfer ubiquitin onto target proteins. A31N ts20 cells are mouse embryo fibroblasts, thermosensitive for E1. We show here that: (a) the enzymatic activity of the enzyme is heat-inactivatable in vitro; and (b) a major mechanism responsible for E1 inactivation in vivo consists of accelerated destruction. Surprisingly, a >90% reduction in E1 abundance little alters the formation of the bulk of protein-ubiquitin conjugates when A31N-ts20 cells are grown at the nonpermissive temperature, indicating that cautious interpretation of results is required when studying ubiquitination of specific substrates using this cell line. Surprisingly, our data also indicate that, in vivo, ubiquitination of the various protein substrates in A31N-ts20 cells requires different amounts of E1, indicating that this mutant cell line can be used for unveiling the existence of differences in the intimate mechanisms responsible for the ubiquitination of the various cell proteins in vivo, and for providing criteria of reliability when developing in vitro ubiquitination assays for specific proteins. PMID- 10848990 TI - Modified amino acids and peptides as substrates for the intestinal peptide transporter PepT1. AB - The binding affinities of a number of amino-acid and peptide derivatives by the mammalian intestinal peptide transporter PepT1 were investigated, using the Xenopus laevis expression system. A series of blocked amino acids, namely N acetyl-Phe (Ac-Phe), phe-amide (Phe-NH2), N-acetyl-Phe-amide (Ac-Phe-NH2) and the parent compound Phe, was compared for efficacy in inhibiting the uptake of the peptide [3H]-D-Phe-L-Gln. In an equivalent set of experiments, the blocked peptides Ac-Phe-Tyr, Phe-Tyr-NH2 and Ac-Phe-Tyr-NH2 were compared with the parent compound Phe-Tyr. Comparing amino acids and derivatives, only Ac-Phe was an effective inhibitor of peptide uptake (Ki = 1.81+/- 0.37 mM). Ac-Phe-NH2 had a very weak interaction with PepT1 (Ki = 16.8+/-5.64 mM); neither Phe nor Phe-NH2 interacted with PepT1 with measurable affinity. With the dipeptide and derivatives, unsurprisingly the highest affinity interaction was with Phe-Tyr (Ki = 0.10+/-0.04 mM). The blocked C-terminal peptide Phe-Tyr-NH2 also interacted with PepT1 with a relatively high affinity (Ki = 0.94+/-0.38 mM). Both Ac-Phe-Tyr and Ac-Phe-Tyr-NH2 interacted weakly with PepT1 (Ki = 8.41+/-0.11 and 9.97+/-4.01 mM, respectively). The results suggest that the N-terminus is the primary binding site for both dipeptides and tripeptides. Additional experiments with four stereoisomers of Ala-Ala-Ala support this conclusion, and lead us to propose that a histidine residue is involved in binding the C-terminus of dipeptides. In addition, a substrate binding model for PepT1 is proposed. PMID- 10848991 TI - Conformational studies of plasminogen activator inhibitor type 1 by fluorescence spectroscopy. Analysis of the reactive centre of inhibitory and substrate forms, and of their respective reactive-centre cleaved forms. AB - The inhibitors that belong to the serpin family are suicide inhibitors that control the major proteolytic cascades in eucaryotes. Recent data suggest that serpin inhibition involves reactive centre cleavage followed by loop insertion, whereby the covalently linked protease is translocated away from the initial docking site. However under certain circumstances, serpins can also be cleaved like a substrate by target proteases. In this report we have studied the conformation of the reactive centre of plasminogen activator inhibitor type 1 (PAI-1) mutants with inhibitory and substrate properties. The polarized steady state and time-resolved fluorescence anisotropies were determined for BODIPY(R) probes attached to the P1' and P3 positions of the substrate and active forms of PAI-1. The fluorescence data suggest an extended orientational freedom of the probe in the reactive centre of the substrate form as compared to the active form, revealing that the conformation of the reactive centres differ. The intramolecular distance between the P1' and P3 residues in reactive centre cleaved inhibitory and substrate mutants of PAI-1, were determined by using the donor-donor energy migration (DDEM) method. The distances found were 57+/-4 A and 63+/-3 A, respectively, which is comparable to the distance obtained between the same residues when PAI-1 is in complex with urokinase-type plasminogen activator (uPA). Following reactive centre cleavage, our data suggest that the core of the inhibitory and substrate forms possesses an inherited ability of fully inserting the reactive centre loop into beta-sheet A. In the inhibitory forms of PAI-1 forming serpin-protease complexes, this ability leads to a translocation of the cognate protease from one pole of the inhibitor to the opposite one. PMID- 10848992 TI - Differential modulation of alpha, beta and gamma enolase isoforms in regenerating mouse skeletal muscle. AB - Nothing is known about the expression of the glycolytic enzyme enolase in skeletal muscle alterations such as myofiber degeneration and regeneration. Enolase is a dimeric enzyme which exhibits cell type specific isoforms. The embryonic form, alphaalpha, remains expressed in most adult tissues, whereas a transition towards specific isoforms occurs during ontogenesis in two cell types with high energy requirements: alphagamma and gammagamma in neurons, alphabeta and betabeta in striated muscle cells. During murine myogenesis, beta enolase transcripts are detected early in the forming muscles, and the beta gene is further upregulated at specific stages of muscle development. The alpha and beta subunits exhibit characteristic developmental microheterogeneity patterns. High levels of beta enolase subunits characterize the glycolytic fast-twitch fibers of adult muscles. We have investigated the expression of enolase subunits in a mouse experimental model of muscle regeneration. Following a single intramuscular injection of the necrotic agent cardiotoxin, we observed a rapid decrease in the level of the major muscle enolase subunit beta, accounting for the drop in total enolase activity that correlated with the degeneration of myofibers. Concomitant with the regeneration of new fibers, beta subunit levels began to increase, reaching normal values by 30 days after injury. Changes in the embryonic and ubiquitous subunit, alpha, mimicked those occurring during development by two aspects: modifications in electrophoretic variants and redistribution between soluble and insoluble compartments of muscle extracts. Imunocytochemical analyses of alpha and beta enolase subunits first revealed a homogeneous labeling within myofibers. Striations characteristic of normal adult muscle tissue were visible again by day 14 after injury. A perinuclear alpha and beta immunoreactivity was often observed in regenerating myofibers but its functional significance remains to be elucidated. Double labeling experiments with anti-gamma enolase and FITC alpha bungarotoxin allowed us to follow the neuromuscular junction remodeling that occurs during muscle regeneration despite the absence of nerve injury. PMID- 10848993 TI - A novel transmembrane serine/threonine protein kinase gene from a rifamycin SV producing amycolatopsis mediterranei U32. AB - Genomic DNA sequencing in the vicinity of methylmalonyl-CoA mutase gene (mutAB) from a rifamycin SV-producing Amycolatopsis mediterranei U32 allowed us to clone, sequence, and identify a gene encoding a novel serine/threonine protein kinase (amk). The sequence contains a complete ORF of 1821 base pairs encoding a predicted protein of 606 amino acids in length. The N-terminal domain of the protein shows significant homology to the catalytic domain of other protein kinases from both prokaryotic and eukaryotic sources. It also contains all the structural features that are highly conserved in active protein kinases, including the Gly-X-Gly-X-X-Gly motif of ATP-binding and the essential amino acids known to be important for the recognition of the correct hydroxyamino acid in serine/threonine protein kinase. This protein kinase gene was expressed in Escherichia coli and was shown to have the ability of autophosphorylation. The autophosphorylated site was found to be the threonine at position 164 by labeled phosphoamino acid analysis and site-directed mutagenesis. The C-terminal half of protein kinase was found to contain strong transmembrane structures by PhoA fusion protein analysis, suggesting that Amk protein kinase is a transmembrane protein. A Southern hybridization experiment showed that this type of protein kinase is distributed ubiquitously and might play significant physiological roles in the various species of streptomycetes. However, overexpression of amk gene in Streptomyces cinnamonensis showed no effect on methylmalonyl-CoA mutase activity, monensin production and the hyphae morphology. Although its biological role is still unknown, Amk protein kinase is the first transmembrane serine/threonine protein kinase described for genus Amycolatopsis. PMID- 10848994 TI - Deletion of the 6-kDa subunit affects the activity and yield of the bc1 complex from Rhodovulum sulfidophilum. AB - The cytochrome bc1 complex from Rhodovulum sulfidophilum purifies as a four subunit complex: the cytochrome b, cytochrome c1 and Rieske iron-sulphur proteins, which are encoded together in the fbc operon, as well as a 6-kDa protein. The gene encoding the 6-kDa protein, named fbcS, has been identified. It is located within the sox operon, which encodes the subunits of sarcosine oxidase. The encoded 6-kDa protein is very hydrophobic and is predicted to form a single transmembrane helix. It shows no sequence homology to any known protein. The gene has been knocked-out of the genome and a three-subunit complex can be purified. This deletion leads to a large reduction in the yield of the isolated complex and in its activity compared to wild-type. The high quinone content found in the wild-type complex is, however, maintained after removal of the 6-kDa protein. Surprisingly, a fourth subunit of approximately 6 kDa is again found to copurify with the Rhv. sulfidophilum bc1 complex when only the fbc operon is expressed heterologously in a near-relative, Rhodobacter capsulatus, which lacks this small subunit in its own bc1 complex. PMID- 10848995 TI - Characterization of a second methylene tetrahydromethanopterin dehydrogenase from Methylobacterium extorquens AM1. AB - Cell extracts of Methylobacterium extorquens AM1 were recently found to catalyze the dehydrogenation of methylene tetrahydromethanopterin (methylene H4MPT) with NAD+ and NADP+. The purification of a 32-kDa NADP-specific methylene H4MPT dehydrogenase (MtdA) was described already. Here we report on the characterization of a second methylene H4MPT dehydrogenase (MtdB) from this aerobic alpha-proteobacterium. Purified MtdB with an apparent molecular mass of 32 kDa was shown to catalyze the oxidation of methylene H4MPT to methenyl H4MPT with NAD+ and NADP+ via a ternary complex catalytic mechanism. The Km for methylene H4MPT was 50 microM with NAD+ (Vmax = 1100 U x mg(-1) and 100 microM with NADP+ (Vmax = 950 U x mg(-1). The Km value for NAD+ was 200 microM and for NADP+ 20 microM. In contrast to MtdA, MtdB could not catalyze the dehydrogenation of methylene tetrahydrofolate. Via the N-terminal amino-acid sequence, the MtdB encoding gene was identified to be orfX located in a cluster of genes whose translated products show high sequence identities to enzymes previously found only in methanogenic and sulfate reducing archaea. Despite its location, MtdB did not show sequence similarity to archaeal enzymes. The highest similarity was to MtdA, whose encoding gene is located outside of the archaeal island. Mutants defective in MtdB were unable to grow on methanol and showed a pronounced sensitivity towards formaldehyde. On the basis of the mutant phenotype and of the kinetic properties, possible functions of MtdB and MtdA are discussed. We also report that both MtdB and MtdA can be heterologously overproduced in Escherichia coli making these two enzymes readily available for structural analysis. PMID- 10848996 TI - Novel processive and nonprocessive glycosyltransferases from Staphylococcus aureus and Arabidopsis thaliana synthesize glycoglycerolipids, glycophospholipids, glycosphingolipids and glycosylsterols. AB - A processive diacylglycerol glucosyltransferase has recently been identified from Bacillus subtilis [Jorasch, P., Wolter, F.P., Zahringer, U., and Heinz, E. (1998) Mol. Microbiol. 29, 419-430]. Now we report the cloning and characterization of two other genes coding for diacylglycerol glycosyltransferases from Staphylococcus aureus and Arabidopsis thaliana; only the S. aureus enzyme shows processivity similar to the B. subtilis enzyme. Both glycosyltransferases characterized in this work show unexpected acceptor specificities. We describe the isolation of the ugt106B1 gene (GenBank accession number Y14370) from the genomic DNA of S. aureus and the ugt81A1 cDNA (GenBank accession number AL031004) from A. thaliana by PCR. After cloning and expression of S. aureus Ugt106B1 in Escherichia coli, SDS/PAGE of total cell extracts showed strong expression of a protein having the predicted size of 44 kDa. Thin-layer chromatographic analysis of the lipids extracted from the transformed E. coli cells revealed several new glycolipids and phosphoglycolipids not present in the controls. These lipids were purified from lipid extracts of E. coli cells expressing the S. aureus gene and identified by NMR and mass spectrometry as 1, 2-diacyl-3-[O-beta-D glucopyranosyl]-sn-glycerol, 1, 2-diacyl-3-[O-beta-D-glucopyranosyl-(1-->6)-O beta-D-glucopyrano-+ ++syl] -sn-glycerol, 1, 2-diacyl-3-[O-beta-D-glucopyranosyl (1-->6)-O-beta-D-glucopyranosyl-( 1-->6)-O-beta-D-glucopyranosyl]-sn-glycerol, sn 3'-[O-beta-D-glucopyranosyl]-phosphatidylglycerol and sn-3'-[O-(6"'-O-acyl)-beta D-glucopyranosyl-(1"'-->6")-O-beta-D-gluco pyranosyl]-sn-2'-acyl-phospha tidylglycerol. A 1, 2-diacyl-3-[O-beta-D-galactopyranosyl]-sn-glycerol was isolated from extracts of E. coli cells expressing the ugt81A1 cDNA from A. thaliana. The enzymatic activities expected to catalyze the synthesis of these compounds were confirmed by in vitro assays with radioactive substrates. Experiments with several of the above described glycolipids as 14C-labeled sugar acceptors and unlabeled UDP-glucose as glucose donor, suggest that the ugt106B1 gene codes for a processive UDP-glucose:1, 2-diacylglycerol-3-beta-D glucosyltransferase, whereas ugt81A1 codes for a nonprocessive diacylglycerol galactosyltransferase. As shown in additional assays with different lipophilic acceptors, both enzymes use diacylglycerol and ceramide, but Ugt106B1 also accepts glucosyl ceramide as well as cholesterol and cholesterol glucoside as sugar acceptors. PMID- 10848997 TI - Phosphoinositides determine specificity of the guanine-nucleotide exchange activity of cytohesin-1 for ADP-ribosylation factors derived from a mammalian expression system. AB - ADP-ribosylation factors (ARFs) are small Ras-like GTPases which play important roles in intracellular vesicle transport and in the remodeling of the actin cytoskeleton. Guanine nucleotide exchange factors (GEFs) for ARFs have recently been identified. One of them, cytohesin-1, a 47-kDa cytoplasmic protein acts as an inside-out signaling molecule and regulates binding of the beta2 integrin leukocyte function antigen 1 (LFA-1) to its ligand intercellular adhesion molecule 1 (ICAM-1). In this study, we address the regulation of the GEF activity of cytohesin-1 by phosphoinositides, using mammalian expression of functional ARF Ig chimeras. The fusion proteins, which can be quantitatively immunoprecipitated on protein A-Sepharose, target to the expected intracellular compartments, and they are readily induced to bind GTP in vitro. We show that both ARF1-Ig and ARF6 Ig chimeras are activated in vitro by cytohesin-1. However, GEF activity towards ARF6 is strongly suppressed by phosphatidylinositol-(3,4,5)-trisphosphate (PtdInsP3). In contrast, cytohesin-1-dependent GTP binding of ARF1 is significantly enhanced by PtdInsP3. We conclude that the membrane phospholipid PtdInsP3 determines the specificity of the GEF activity of cytohesin-1. PMID- 10848998 TI - The assembly pathway of outer membrane protein PhoE of Escherichia coli. AB - The assembly of the wild-type and several mutant forms of the trimeric outer membrane porin PhoE of Escherichia coli was investigated in vitro and in vivo. In in vivo pulse-chase experiments, approximately half of the wild-type PhoE molecules assembled within the 30-s pulse in the native conformation in the cell envelope. The other half of the molecules followed slower kinetics, and three intermediates in this multistep assembly process were detected: a soluble trypsin sensitive monomer, a trypsin-sensitive monomeric form that was loosely associated with the cell envelope and a metastable trimer, which was integrated into the membranes and converted to the stable trimeric configuration within minutes. The metastable trimers disassembled during sample preparation for standard SDS/PAGE into folded monomers. In vitro, the isolated PhoE protein could efficiently be folded in the presence of N,N-dimethyldodecylamine-N-oxide (LDAO). A mutant PhoE protein, DeltaF330, which lacks the C-terminal phenylalanine residue, mainly followed the slower kinetic pathway observed in vivo, resulting in increased amounts of the various assembly intermediates. It appears that the DeltaF330 mutant protein is intrinsically able to fold, because it was able to fold in vitro with LDAO with similar efficiencies as the wild-type protein. Therefore, we propose that the conserved C-terminal Phe is (part of) a sorting signal, directing the protein efficiently to the outer membrane. Furthermore, we analysed a mutant protein with a hydrophilic residue introduced at the hydrophobic side of one of the membrane-spanning amphipathic beta strands. The assembly of this mutant protein was not affected in vivo or in vitro in the presence of LDAO. However, it was not able to form folded monomers in a previously established in vitro folding system, which requires the presence of lipopolysaccharides and Triton. Hence, a folded monomer might not be a true assembly intermediate of PhoE in vivo. PMID- 10848999 TI - A bifunctional delta-fatty acyl acetylenase/desaturase from the moss Ceratodon purpureus. A new member of the cytochrome b5 superfamily. AB - Many plant genes have been cloned that encode regioselective desaturases catalyzing the formation of cis-unsaturated fatty acids. However, very few genes have been cloned that encode enzymes catalyzing the formation of the functional groups found in unusual fatty acids (e.g. hydroxy, epoxy or acetylenic fatty acids). Here, we describe the characterization of an acetylenase from the moss Ceratodon purpureus with a regioselectivity differing from the previously described Delta12-acetylenase. The gene encoding this protein, together with a Delta6-desaturase, was cloned by a PCR-based approach with primers derived from conserved regions in Delta5-, Delta6-fatty-acid desaturases and Delta8 sphingolipid desaturases. The proteins that are encoded by the two cloned cDNAs are likely to consist of a N-terminal extension of unknown function, a cytochrome b5-domain, and a C-terminal domain that is similar to acyl lipid desaturases with characteristic histidine boxes. The proteins were highly homologous in sequence to the Delta6-desaturase from the moss Physcomitrella patens. When these two cDNAs were expressed in Saccharomyces cerevisiae, both transgenic yeast cultures desaturated Delta9-unsaturated C16- and C18-fatty acids by inserting an additional Delta6cis-double bond. One of these transgenic yeast clones was also able to introduce a Delta6-triple bond into gamma-linolenic and stearidonic acid. This resulted in the formation of 9,12,15-(Z,Z,Z)-octadecatrien-6-ynoic acid, the main fatty acid found in C. pupureus. These results demonstrate that the Delta6 acetylenase from C. pupureus is a bifunctional enzyme, which can introduce a Delta6cis-double bond into 9,12,(15)-C18-polyenoic acids as well as converting a Delta6cis-double bond to a Delta6-triple bond. PMID- 10849000 TI - Further analysis of the involvement of the envelope anion channel PIRAC in chloroplast protein import. AB - The ability of preferredoxin to inactivate a 50-pS anion channel of the chloroplast inner membrane in the presence of an energy source was investigated using single-channel recordings. It was found that preferredoxin cannot inactivate the channel when GTP is the only energy source present. From this it is concluded that the precursor has to interact with the, translocon of the inner membrane of chloroplasts (Tic) complex to be able to inactivate the 50-pS anion channel. The ability of two mutants of preferredoxin with deletions in their transit sequence to inactivate the channel was also tested. Both mutants have been shown to have a similar binding affinity for the chloroplast envelope, but only one is able to fully translocate. The mutants were both able to inactivate the channel in a similar manner. From this it is concluded that full translocation is not necessary for the inactivation of the channel. It is also shown that preferredoxin is capable of inactivating the 50-pS anion channel in the chloroplast-attached configuration as was previously found in the inside-out configuration. From this it is concluded that stromal factors do not influence the protein-import-induced inactivation of the 50-pS anion channel of the chloroplast inner membrane. Finally the effect of the anion channel blocker 4, 4' diisothiocyanostilbene-2,2'-disulfonate (DIDS) on the channel activity and on protein import was investigated. It was found that DIDS blocked the channel. Furthermore the addition of the channel blocker reduces the efficiency of import to 52%. This leads to the conclusion that correct functioning of the channel is important for protein import. PMID- 10849001 TI - Calcium-dependent protein kinase from maize seedlings activated by phospholipids. AB - A calcium- and phospholipid-dependent protein kinase of apparent molecular mass 54 kDa (designated ZmCPKp54) was partially purified from etiolated maize seedlings. Activity of ZmCPKp54 is stimulated by phosphatidylserine and phosphatidylinositol, but is not essentially affected by diolein and phorbol esters. The enzyme cross-reacts with polyclonal antibodies against the calmodulin like-domain of the calcium-dependent protein kinase, but not with antibodies against catalytic or regulatory domains of protein kinase C. ZmCPKp54 is not able to phosphorylate the specific substrates of protein kinase C (MARCKS peptide and protein kinase C substrate peptide derived from pseudosubstrate sequence) and its activity is not inhibited by specific PKC inhibitors (bisindolylmaleimide, protein kinase C pseudosubstrate inhibitory peptide). The substrate specificity and sensitivity to the inhibitors of the maize enzyme resembles calcium-dependent protein kinase. The biochemical and immunological properties indicate that ZmCPKp54 belongs to the calcium-dependent protein kinase family. PMID- 10849002 TI - The pro- or anti-apoptotic function of NF-kappaB is determined by the nature of the apoptotic stimulus. AB - To test whether the behaviour of transcription factor NF-kappaB as a promoter or antagonist of apoptosis depends on the apoptotic stimulus, we determined the influence of NF-kappaB on cell killing elicited by a variety of inducers within a given cell type. Inhibition of NF-kappaB by genetic and pharmacological approaches rendered HeLa cells more susceptible to TNF-alpha-induced cell killing, but protected them almost completely from H2O2- and pervanadate-induced apoptosis. TNF-alpha was unable to protect HeLa from H2O2- and pervanadate induced apoptosis and further enhanced the cytotoxicity induced by these two adverse agents. Supernatants from HeLa cells stably overexpressing a transdominant negative form of IkappaB-alpha selectively increased the cytotoxicity of TNF-alpha for HeLa cells, suggesting that the enhanced susceptibility of these cells can be attributed to one or more secretable factors. Supershift experiments showed that the various apoptotic stimuli induced the same subset of DNA-binding subunits. Therefore, the nature of the signals elicited by the respective death inducers determines whether NF-kappaB induction leads to apoptosis or survival, suggesting that the manipulation of NF-kappaB activity may provide a new approach to adjuvant therapy in cancer treatment. PMID- 10849003 TI - Export and folding of signal-sequenceless Bacillus licheniformis beta-lactamase in Escherichia coli. AB - Two genetically engineered variants of the Bacillus licheniformis beta-lactamase gene were expressed in Escherichia coli. One variant coded for the exo-small mature enzyme without the signal peptide. The other coded for the exo-large mature enzyme preceded by 10, mostly polar, residues from an incomplete heterologous signal. As observed following the extraction by a lysozyme-EDTA treatment, the signal-less variant was exported to the periplasm with nearly 20% efficiency, whereas the variant with the N-terminal extension was translocated to a lesser degree; interestingly, nearly all of the former and half of the latter were extracted by osmotic shock, which may be of importance for our understanding of cellular compartments. The fact that a signal-less protein is translocated with substantial yields raises questions about the essential role of signal peptides for protein export. As folding and export are related processes, we investigated the folding in vitro of the two variants. No differences were found between them. In the absence of denaturant, they are completely folded, fully active and have a large DeltaG of unfolding. Under partially denaturing conditions they populate several partially folded states. The absence of significant amounts of a non-native state under native conditions makes a thermodynamic partitioning between folding and export less likely. In addition, kinetic measurements indicated that these B. licheniformis lactamases fold much faster than E. coli beta-lactamase. This behavior suggests that they are exported by a kinetically controlled process, mediated by one or more still unidentified interactions that slow folding and allow a folding intermediate to enter the export pathway. PMID- 10849004 TI - A small-angle X-ray solution scattering study of bovine alpha-crystallin. AB - The native high molecular mass form of alpha-crystallin, the most important soluble protein in the eye lens, and its low molecular mass form obtained at 37 degrees C in dilute solutions were investigated by synchrotron radiation small angle X-ray scattering. The alpha-crystallin solutions are polydisperse and good fits to the experimental data can be obtained using distributions of spheres with radii varying between about 5 and 10 nm. In spite of the polydispersity, two different ab initio methods were used to retrieve low resolution shapes from the scattering data. These shapes correspond to the z-average structure of the oligomers. In the absence of any symmetry constraints, the scattering curves of the two forms of alpha-crystallin yield bean-like shapes. The shape corresponding to the low molecular mass form has about 20% less mass at the periphery. Imposing tetrahedral symmetry on the average structures worsens the fit to the experimental data. We emphasized the apparent contradiction between hydrodynamic and molecular properties of alpha-crystallin. An explanation was put forward based on the presence of solvent-exposed flexible C-terminal extensions. We present two bead models ('hollow globule with tentacles' and 'bean with tentacles') based on NMR and cryo-electron microscopy studies and discuss how well they correspond with our data from X-ray scattering, light scattering and analytical ultracentrifugation. PMID- 10849005 TI - Metabolic characterization of Lactococcus lactis deficient in lactate dehydrogenase using in vivo 13C-NMR. AB - The metabolism of glucose by nongrowing cells of Lactococcus lactis strain FI7851, constructed from the wild-type L. lactis strain MG1363 by disruption of the lactate dehydrogenase (ldh) gene [Gasson, M.J., Benson, K., Swindel, S. & Griffin, H. (1996) Lait 76, 33-40] was studied in a noninvasive manner by 13C NMR. The kinetics of the build-up and consumption of the pools of intracellular intermediates mannitol 1-phosphate, fructose 1,6-bisphosphate, 3 phosphoglycerate, and phosphoenolpyruvate as well as the utilization of [1 13C]glucose and formation of products (lactate, acetate, mannitol, ethanol, acetoin, 2,3-butanediol) were monitored in vivo with a time resolution of 30 s. The metabolism of glucose by the parental wild-type strain was also examined for comparison. A clear shift from typical homolactic fermentation (parental strain) to a mixed acid fermentation (lactate dehdydrogenase deficient; LDHd strain) was observed. Furthermore, high levels of mannitol were transiently produced and metabolized once glucose was depleted. Mannitol 1-phosphate accumulated intracellularly up to 76 mM concentration. Mannitol was formed from fructose 6 phosphate by the combined action of mannitol-1-phosphate dehydrogenase and phosphatase. The results show that the formation of mannitol 1-phosphate by the LDHd strain during glucose catabolism is a consequence of impairment in NADH oxidation caused by a highly reduced LDH activity, the transient production of mannitol 1-phosphate serving as a regeneration pathway for NAD+ regeneration. Oxygen availability caused a drastic change in the pattern of intermediates and end-products, reinforcing the key-role of the fulfilment of the redox balance. The flux control coefficients for the step catalysed by mannitol-1-phosphate dehydrogenase were calculated and the implications in the design of metabolic engineering strategies are discussed. PMID- 10849006 TI - Identification of novel periviscerokinins from single neurohaemal release sites in insects MS/MS fragmentation complemented by Edman degradation. AB - Three novel members of the periviscerokinin family could be identified directly from extracts of single abdominal perisympathetic organs of blaberoid cockroaches by means of electrospray ionization-quadrupole time of flight (ESI-QTOF) MS. Sequences of these periviscerokinins were confirmed by Edman degradation. Their primary structures are GSSGLIPFGRT-NH2 (Lem-PVK-1), GSSGLISMPRV-NH2 (Lem-PVK-2), and GSSGMIPFPRV-NH2 (Lem-PVK-3). Hitherto only known from the American cockroach, this neuropeptide family contains a highly conserved N-terminus whereas, at the C terminus, only the penultimate amino-acid residue (Arg) has been found in all members of this peptide family. The identified periviscerokinins are the only abundant myoactive peptides in abdominal perisympathetic organs of blaberoid cockroches and they appear to be absent in the retrocerebral complex. Screening of extracts of single abdominal perisympathetic organs (70-90 microm in diameter), from five different species of the suborder Blaberoidea, revealed that they all contain the three neuropeptides which are described here for the first time. PMID- 10849007 TI - The involvement of coenzyme A esters in the dehydration of (R)-phenyllactate to (E)-cinnamate by Clostridium sporogenes. AB - Phenyllactate dehydratase from Clostridium sporogenes grown anaerobically on L phenylalanine catalyses the reversible syn-dehydration of (R)-phenyllactate to (E)-cinnamate. Purification yielded a heterotrimeric enzyme complex (130 +/- 15 kDa) composed of FldA (46 kDa), FldB (43 kDa) and FldC (40 kDa). By re chromatography on Q-Sepharose, the major part of FldA could be separated and identified as oxygen insensitive cinnamoyl-CoA:phenyllactate CoA-transferase, whereas the transferase depleted trimeric complex retained oxygen sensitive phenyllactate dehydratase activity and contained about one [4Fe-4S] cluster. The dehydratase activity required 10 microM FAD, 0.4 mM ATP, 2.5 mM MgCl2, 0.1 mM NADH, 5 microM cinnamoyl-CoA and small amounts of cell-free extract (10 microg protein per mL) similar to that known for 2-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans. The N-terminus of the homogenous FldA (39 amino acids) is homologous to that of CaiB (39% sequence identity) involved in carnitine metabolism in Escherichia coli. Both enzymes are members of an emerging group of CoA-transferases which exhibit high substrate specificity but apparently do not form enzyme CoA-ester intermediates. It is concluded that dehydration of (R)-phenyllactate to (E)-cinnamate proceeds in two steps, a CoA-transfer from cinnamoyl-CoA to phenyllactate, catalysed by FldA, followed by the dehydration of phenyllactyl-CoA, catalysed by FldB and FldC, whereby the noncovalently bound prosthetic group cinnamoyl-CoA is regenerated. This demonstrates the necessity of a 2-hydroxyacyl-CoA intermediate in the dehydration of 2-hydroxyacids. The transient CoA-ester formation during the dehydration of phenyllactate resembles that during citrate cleavage catalysed by bacterial citrate lyase, which contain a derivative of acetyl-CoA covalently bound to an acyl-carrier-protein (ACP). PMID- 10849008 TI - Structure and location of a ferritin gene of the yellow fever mosquito Aedes aegypti. AB - We have isolated and sequenced a genomic clone encoding the 24- and 26-kDa ferritin subunits in the mosquito Aedes aegypti (Rockefeller strain). The A. aegypti gene differs from other known ferritin genes in that it possesses an additional intron and an unusually large second intron. The additional intron is located within the 5' untranslated region, between the CAP site and the start codon. The second intron contains numerous putative transposable elements. In addition, unlike the human and rat ferritin genes, the A. aegypti ferritin gene is a single copy gene, located at 88.3% FLpter on the q-arm of chromosome 1. Primer extension analysis indicates that the A. aegypti ferritin gene has multiple transcriptional start sites. A differential usage of these sites is observed with varied cellular iron concentrations. PMID- 10849009 TI - The PKC targeting protein RACK1 interacts with the Epstein-Barr virus activator protein BZLF1. AB - Phorbol esters reactivate Epstein-Barr virus (EBV) from latently infected cells via transcriptional activation of the viral immediate-early gene BZLF1. BZLF1 is a member of the extended AP-1 family of transcription factors that binds to specific BZLF1-binding motifs within early EBV promoters and to consensus AP-1 sites. Regulation of BZLF1's activity is achieved at the transcriptional level as well as through post-translational modifications. Recently, we reported that the transcriptional activity of BZLF1 is augmented by TPA [Baumann, M., Mischak, H., Dammeier, S., Kolch, W., Gires, O., Pich, D., Zeidler, R., Delecluse, H. J. & Hammerschmidt, W., (1998) J. Virol. 72, 8105-8114]. The increase of BZLF1's activity depends on a single serine residue (S186) that is phosphorylated by protein kinase C (PKC) in vitro and in vivo after stimulation with 12-O tetradecanoylphorbol-13-acetate (TPA). Here, we identified RACK1 as a binding partner of BZLF1 in a yeast interaction trap assay. RACK stands for receptor of activated C-kinase and is involved in targeting activated PKCs and other signaling proteins. In vivo, RACK1 binds directly to the transactivation domain of BZLF1. Although a functional relationship between BZLF1 and PKC could be mediated by RACKs, RACK1 did not have a detectable effect on the phosphorylation status of BZLF1 in in vitro or in vivo phosphorylation assays. We suggest that RACK1 may act as a scaffolding protein on BZLF1 independently of activated PKCs. PMID- 10849010 TI - Characterization of the phosphocholine-substituted oligosaccharide in lipopolysaccharides of type b Haemophilus influenzae. AB - Haemophilus influenzae expresses heterogeneous populations of short-chain lipopolysaccharide (LPS) which exhibit extensive antigenic diversity among multiple oligosaccharide epitopes. These LPS oligosaccharide epitopes can carry phosphocholine (PCho) substituents, the expression of which is subject to high frequency phase variation mediated by genes in the lic1 genetic locus. The location and site of attachment of PCho substituents were determined by structural analysis of LPS from two type b H. influenzae strains, Eagan and RM7004. The lic2 locus is involved in phase variation of oligosaccharide expression. LPS obtained from the parent strains, from mutants generated by insertion of antibiotic resistance cassettes in the lic2 genetic locus, and from phase-variants showing high levels of PCho expression was characterized by electrospray ionization-mass spectrometry (ESI-MS) and 1H NMR spectroscopy of derived O-deacylated samples. ESI-MS of O-deacylated LPS from wild-type strains revealed mixtures of related glycoform structures differing in the number of hexose residues. Analysis of LPS from PCho-expressing phase-variants revealed similar mixtures of glycoforms, each containing a single PCho substituent. O Deacylated LPS preparations from the lic2 mutants were much less complex than their respective parent strains, consisting only of Hex3 and/or Hex2 glycoforms, were examined in detail by high-field NMR techniques. It was found that the LPS samples contain the phosphoethanolamine (PEtn) substituted inner-core element, L alpha-D-Hepp-(1-->2)-[PEtn-->6]-L-alpha-D-Hepp-(1--> 3)-L-alpha-D-He pp-(1-->5) alpha-Kdo in which the major glycoforms carry a beta-D-Glcp or beta-D-Glcp-(1- >4)-beta-D-Glcp at the O-4 position of the 3-substituted heptose (HepI) and a beta-D-Galp at the O-2 position of the terminal heptose (HepIII). LPS from the lic2 mutants of both type b strains were found to carry PCho groups at the O-6 position of the terminal beta-D-Galp residue attached to HepIII. In the parent strains, the central heptose (HepII) of the LPS inner-core element is also substituted by hexose containing oligosaccharides. The expression of the galabiose epitope in LPS of H. influenzae type b strains has previously been linked to genes comprising the lic2 locus. The present study provides definitive evidence for the role of lic2 genes in initiating chain extension from HepII. From the analysis of core oligosaccharide samples, LPS from the lic2 mutant strain of RM7004 was also found to carry O-acetyl substituents. Mono-, di-, and tri-O-acetylated LPS oligosaccharides were identified. The major O-acetylated glycoforms were found to be substituted at the O-3 position of HepIII. A di-O acetylated species was characterized which was also substituted at the O-6 postion of the terminal beta-D-Glc in the Hex3 glycoform. This is the first report pointing to the occurrence of O-acetyl groups in the inner-core region of H. influenzae LPS. We have previously shown that in H. influenzae strain Rd, a capsule-deficient type d strain, PCho groups are expressed in a different molecular environment, being attached at the O-6 position of a beta-D-Glcp, which is in turn attached to HepI. PMID- 10849011 TI - Gastric tonometry. PMID- 10849012 TI - Microglia and neurodegeneration. PMID- 10849013 TI - Does sarcoid diathesis represent susceptibility to micro-organisms? PMID- 10849014 TI - The effect of cholesterol lowering on carotid and femoral artery wall stiffness and thickness in patients with familial hypercholesterolaemia. AB - BACKGROUND: Early in the process of atherosclerosis, changes in vessel wall stiffness and thickness may occur. The present study evaluates the effect of cholesterol reduction on artery wall stiffness and intima media thickness in patients with familial hypercholesterolaemia (FH). MATERIALS AND METHODS: Forty five patients with familial hypercholesterolaemia (mean age 46+/-10 years) with untreated LDL cholesterol concentration > 9 mmol L(-1), were studied before and after one year of cholesterol lowering therapy with statins (simvastatin, atorvastatin 40-80 mg day(-1). The distensibility (DC in 10-3 kPa(-1) and compliance (CC in mm2. kPa(-1) of the common carotid artery (CCA) (right and left side) and common femoral artery (CFA) (right side) were determined by a wall track system (Pie Medical). The intima media thickness (IMT) (both right and left) of the CCA, bulb (BUL), internal carotid artery (ICA) and CFA were measured in mm by high-resolution ultrasound (Biosound). RESULTS: The mean concentration of total cholesterol (TC), LDL-cholesterol (LDL-C) and triglycerides (TG) were reduced significantly by 43%, 51% and 25%, respectively, whereas HDL-cholesterol (HDL-C) increased by 13% (P<0.001). In the CFA, the DC and CC increased significantly (DC from 7.9+/-3.0 to 9.1+/-3.7 in 10(-3) kPa(-1); CC 0.5+/-0.2 0.6+/-0.3 in mm2. kPa(-1), whereas the DC and CC did not change in the CCA. In contrast, the IMT of the CCA decreased significantly in both men and women whereas an IMT decrease was also seen in the BUL and ICA in premenopausal women. A LDL-cholesterol reduction of 44.8% and 45.4% was necessary to induce significant decreases in IMT and increases in DC and CC. CONCLUSIONS: One year of cholesterol lowering therapy in FH decreases the wall stiffness in the CFA and the arterial wall thickness in the CCA. PMID- 10849015 TI - Microvascular angina (cardiological syndrome X) per se is not associated with hyperinsulinaemia or insulin resistance. AB - BACKGROUND: Microvascular angina has been found to be associated with insulin resistance. However, many factors known to affect insulin sensitivity were not excluded in patient selection. We aimed to evaluate whether microvascular angina is per se associated with insulin resistance. MATERIALS AND METHODS: We performed a Frequently Sampled Intravenous Glucose Tolerance Test (0.33 g kg(-1) b.w.) in 10 normal weight and normotensive patients with microvascular angina, with normal glucose tolerance and normal plasma lipids. Ten healthy subjects, comparable for age, sex, body mass index, blood pressure and plasma lipids, were used as control group. RESULTS: Fasting serum glucose (4.49+/-0.2 SEM vs. 4.52+/-0.13 mmol L(-1), P = 0.9), insulin (39.46 +/-3.68 SEM vs. 47.12+/-4.6 pmol L(-1), P = 0.21) and C peptide (0.56 +/-0.05 SEM vs. 0.53+/-0.05 nmol L(-1), P = 0. 68) values, as well as estimated parameters of insulin secretion and hepatic insulin extraction were similar in the two groups. Insulin sensitivity values (median, range) were also similar in the patients and control subjects (5.76 (3.39-12.30) vs. 7.54 (3.68 13.89. 10(-4) x min(-1)/(microU/mL), P = 0.97). CONCLUSION: Microvascular angina per se is not associated with hyperinsulinaemia or insulin resistance when other confounding factors are excluded in patient selection. PMID- 10849016 TI - Criteria for mutation analysis in MEN 1-suspected patients: MEN 1 case-finding. AB - BACKGROUND: Multiple endocrine neoplasia type 1 (MEN 1) is an autosomal, dominantly inherited cancer syndrome, with tumours in various endocrine glands. In 1997 the responsible tumour suppressor gene was identified. MEN1 gene germ line mutations are detected in the vast majority of MEN 1 patients, however, with regard to case-finding, unfortunately only at a very low frequency in patients with apparently sporadic MEN 1-related tumours. In order to increase the detection rate of disease gene carriers among patients with apparently sporadic MEN 1-related tumours, clinical criteria were needed. DESIGN AND RESULTS: In this study MEN1 gene germ-line mutations were revealed in 16/16 MEN 1 patients/families (100%). Based on our clinical experience with MEN 1 patients/families we formulated clinical criteria to identify disease gene carriers among patients with apparently sporadic MEN 1-related tumours. The criteria for MEN 1-suspected patients are: young age at onset (< 35 years) and/or multiple MEN 1-related lesions in a single organ or two distinct organs affected. Application of these criteria yielded MEN1 gene germ-line mutations in nine of 15 MEN 1-suspected patients (60%), thus identifying novel MEN 1 families. Follow up was also guaranteed for patients not fulfilling these criteria. CONCLUSIONS: The clinical criteria for MEN 1-suspected patients increase the detection rate of germ-line MEN1 gene mutations among patients with apparently sporadic MEN 1 related tumours. These criteria may be used for (presymptomatic) identification of MEN 1 disease gene-carriers, thus enabling early detection of tumour development and timely treatment, as well as genetic counselling. PMID- 10849017 TI - Sporadic endocrine tumours and their relationship to the hereditary endocrine neoplasia syndromes. AB - In the last years of the previous century the genes involved in the aetiology of five endocrine tumour syndromes have been identified. The tumour-suppressor gene that is responsible for Von Hippel-Lindau Disease was cloned in 1993; multiple endocrine neoplasia (MEN) types 2A and 2B and familial medullary thyroid carcinoma were found to be caused by activating mutations in the ret proto oncogene in 1993 and 1994, and most recently the menin-gene, another tumour suppressor gene, was shown to be associated with MEN-1. As usual, the answer to one question leads to innumerable new questions. And so, now we want to know the extent to which germ-line mutations (de novo, or otherwise previously undetected) in these genes play a role in the occurrence of the various endocrine tumours that are associated with these syndromes in apparently sporadic cases. We also want to know if the nature of the (germ-line) mutation conveys any information about the characteristics (phenotype) of the disease. We want to know the role of somatic mutations in these genes in truly sporadic tumours. And finally we want to know the exact function of the proteins that are encoded by these genes. The paper by Roijers et al. [1] elsewhere in this issue is an example of a small but well-directed step on the way to address some of these questions with respect to the menin-gene. It addresses the problem of patient selection when looking for germ-line mutations in apparently sporadic MEN-1 patients. In this review we want to give a brief summary of the present status with regard to some of the questions mentioned above, in relation to the endocrine tumour syndromes caused by the vhl, ret and menin genes. PMID- 10849018 TI - Gastrotonometry represents dramatic increase in PcO2 after acetazolamide administration. AB - BACKGROUND: We sought to evaluate the parameters of CO2 transport during the administration of acetazolamide in order to assess the role of carbonic anhydrase in CO2 transport. MATERIALS AND METHODS: The partial pressure of carbon dioxide in tissue (PtCO2), arterial blood (PaCO2) and end-tidal gas (PETCO2) were monitored to study the correlation between PaCO2, PtCO2 and PETCO2 in spontaneously breathing healthy volunteers after the intravenous administration of acetazolamide 6 mg kg-1. RESULTS: At 60 min after the administration of acetazolamide, the PtCO2 peaked at more than 60 mmHg, and although it decreased by 90 min, it then remained stable above the baseline value. The PaCO2 did not change and the PETCO2 decreased significantly. The changes in PtCO2 were greater than those of either PaCO2 or PETCO2. The minute ventilation increased progressively throughout the study. CONCLUSIONS: We concluded that gastrotonometry represents a new method for monitoring the dramatic increase in PtCO2 induced by drugs such as acetazolamide clinically, and that it could be a warning against acetazolamide administration in severe patients without keeping a ventilation and circulation reserve. PMID- 10849019 TI - Iron supplementation affects the production of pro-inflammatory cytokines in IL 10 deficient mice. AB - BACKGROUND: Iron supplements may increase disease activity in inflammatory bowel disease through the production of the hydroxyl radical because of its catalytic activity in the Fenton reaction. The purpose of this study was to assess the effect of dietary and locally administered iron in the IL-10 knock-out (-/-) mouse, a model of chronic inflammatory bowel disease. MATERIALS AND METHODS: IL10 /- and wild-type mice received a standard or a high-iron diet (35 mg kg(-1) ferrosulphate vs. 500 mg kg(-1) ferrosulphate) after weaning. After 4 weeks the mice were sacrificed. Furthermore, a group of adult IL-10 knock-out mice was given three iron-containing enema's (0.2 mL of 1 mM ferrous-ammonium sulphate) or phosphate buffered saline. These mice were sacrificed after 1 week. Production of pro-inflammatory cytokines by colon tissue cultures, haematological parameters and histology was determined to assess inflammatory activity. RESULTS: Oral as well as rectal administration of iron resulted in increased pro-inflammatory cytokine production in IL-10-/- mice. Neutrophil counts in IL10-/- on a high iron diet increased as well. No enhanced colonic inflammation was noted on histology after iron supplementation. CONCLUSION: We conclude that dietary or topical administered iron increases pro-inflammatory cytokine production in the colon of IL10-/- mice. No significant increase of histological intestinal inflammation was observed. PMID- 10849020 TI - Fast, simple and highly sensitive double-rounded polymerase chain reaction assay to detect medically relevant fungi in dermatological specimens. AB - BACKGROUND: Early detection of fungal infection is essential for beginning of prompt and specific therapy. In this study we describe a rapid and sensitive procedure to detect, by polymerase chain reaction (PCR) assays, a wide range of medically important opportunistic and pathogenic fungi in dermal specimens from dermatomycoses-affected patients. MATERIALS AND METHODS: Three primer pairs, amplifying fragments of the highly conserved gene coding for small ribosomal RNA (18S rDNA) and the adjacent internal transcribed spacer (ITS) rDNA, previously published by others, were probed on DNA from pure cultures of medically relevant human and animal fungal species. In order to evaluate the specificity of the assay, amplifications of control DNAs from other eukaryotes and prokaryotes were also carried out, and they gave negative results. RESULTS: These primer sets, in single amplification or double-rounded PCR assays, allowed specific amplification when applied to a wide number of fungal DNA from human and animal tissue specimens, including dermatophytes (genera Trichophyton, Microsporum), several yeast species (Candida, Saccharomyces, Cryptococcus, Malassezia) and moulds (Aspergillus, Penicillium). The PCR assay was able to detect as little as 10 pg of fungal DNA, corresponding to approximately 25 fungal genomes per sample specimen. A small-scale DNA extraction method was also developed. This simple, time-saving and sensitive procedure was successfully applied to 40 human and veterinary specimens, and the diagnosis was confirmed with cultural techniques, being shown to work even in the presence of other lesions or contaminating organisms. CONCLUSIONS: This method allows early recognition of fungal pathogen cells in clinical samples as an alternative tool to conventional detection techniques. PMID- 10849021 TI - Plasma membrane polarity of polymorphonuclear leucocytes from children with primary ciliary dyskinesia. AB - BACKGROUND: Polymorphonuclear leucocytes (PMN) from subjects with primary ciliary dyskinesia (PCD) can have abnormal locomotory systems. The locomotory activity of PMN is the result of biochemical events mediated by the plasma membrane. In this study we investigated plasma membrane polarity of PMN from children with PCD. DESIGN: Membrane polarity was studied in 11 children with PCD and in healthy controls by measuring the steady-state fluorescence excitation and emission spectra of 2-dimethylamino[6-lauroyl]naphthalene (Laurdan), which is known to be incorporated at the hydrophobic-hydrophilic interface of the bilayer, displaying spectral sensitivity to the polarity of its surroundings. Laurdan shows a marked steady-state emission red shift in polar solvents, with respect to nonpolar solvents. Moreover, the effect of the microtubule disassembling agent colchicine on PMN membrane polarity was evaluated. RESULT: Our results show a red shift of the fluorescence excitation and emission spectra of Laurdan in PMN from the PCD group with respect to the control group. These data indicate an increase in membrane polarity of PMN from the PCD group. Treatment of PMN with colchicine induced a red shift in the Laurdan excitation and emission spectra with the same trend observed in PMN from the PCD group. CONCLUSION: PMN from children with PCD are characterized by an increased plasma membrane polarity. These changes could be the basis of the modifications in the locomotory activities of PMN. The observed alterations may be attributed to abnormalities in the cytoskeleton. PMID- 10849022 TI - Role of macrophage activation in the pathogenesis of Alzheimer's disease and human immunodeficiency virus type 1-associated dementia. AB - The structure and function of neurons are changed not only during development of the central nervous system but also in certain neurological disorders, such as Alzheimer's disease and human immunodeficiency virus type 1 (HIV-1) -associated dementia. Immunological activation and altered production of neurotoxins and neurotrophins by brain macrophages are thought to play an important role in neuronal structure and function. This review describes the clinical and pathological features of both Alzheimer's disease and HIV-1-associated dementia and tries to interpret the role of the macrophage and astrocytes therein. The consequences of activation of macrophages by amyloid-beta in Alzheimer's disease and HIV infection of macrophages in HIV-1-associated dementia and the similarities between these diseases will be discussed. Although the neuropathology of Alzheimer's disease and HIV-1-associated dementia differs, Alzheimer's disease is a cortical dementia and HIV-1-associated dementia is a subcortical dementia, the process of macrophage activation and the resulting pathways leading to neurotoxicity seem very similar. In both Alzheimer's disease and HIV-1-associated dementia, interaction of macrophages and astrocytes appear to play an important role. PMID- 10849023 TI - Phenylalanine inhibition of the phosphorylation of cytoskeletal proteins from cerebral cortex of young rats is prevented by alanine. AB - BACKGROUND: Phenylalanine has been considered the main responsible agent for the brain damage that occurs in phenylketonuria. METHODS AND RESULTS: In this work we studied the effect of this amino acid on the in vitro phosphorylation of cytoskeletal proteins of the cerebral cortex of rats. We observed that 2 mM phenylalanine, a concentration usually found in the plasma of phenylketonuric patients, decreased the in vitro 32P incorporation into these proteins. In addition, we investigated the effect of alanine on the inhibition of 32P incorporation into cytoskeletal proteins caused by phenylalanine. We observed that 0.5 mM alanine did not alter 32P incorporation but prevented the inhibition provoked by phenylalanine. CONCLUSION: In case the inhibition of cytoskeletal protein phosphorylation by phenylalanine also occurs in human phenylketonuria, it is possible that alanine supplementation to the phenylalanine-restricted diet may be beneficial to these patients. PMID- 10849024 TI - Association between alpha-1-antichymotrypsin polymorphism and susceptibility to chronic obstructive pulmonary disease. AB - BACKGROUND: The antiproteases, including alpha-1-antitrypsin, are supposed to prevent lungs from becoming emphysematous. Genetic susceptibility to smoking injury may confer a risk for the development of chronic obstructive pulmonary disease (COPD). METHODS: We have investigated the association between the polymorphism of alpha-1-antichymotrypsin (AACT), one of the antiproteases, and susceptibility to the development of COPD among heavy smokers. Blood samples obtained from both patients with COPD (n = 53) and control subjects (n = 65) at the Tokyo University Hospital, the Juntendo University Hospital and the Tokyo Kenbikyoin Clinic were used for this genotyping assay. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) were performed to genotype the AACT biallelic polymorphism in the signal peptide (-15 alanine to threonine), and the two polymorphisms of the exon (Pro229Ala and Leu55Pro). RESULTS: The proportion of AACT/Ala-15 homozygotes was significantly higher in the COPD patients than in the control subjects (COPD 37.7% vs. control 18.5%). The odds ratio for AACT/Ala-15 homozygotes vs. all other genotypes was 2.7 (95% CI 1.2-6.2) for the COPD group. We could not find any association between the other two polymorphisms and COPD. CONCLUSIONS: Genetic polymorphism in the signal peptide of AACT may be associated with individual susceptibility to the development of COPD, because the AACT/Ala-15 genotype is predominantly found in patients with COPD. It is suggested that AACT/Ala-15 genotype may be less protective against smoking injury. PMID- 10849025 TI - Mannose-binding lectin promoter and structural gene variants in sarcoidosis. AB - BACKGROUND: Sarcoidosis is a chronic granulomatous disease of unknown aetiology. Studies have suggested that the causative agent may be an infectious micro organism. The mannose binding lectin (MBL) is involved in innate immunity to a wide range of micro-organisms. Mutations in the promoter region and exon 1 of the MBL gene occur with high frequency and are associated with reduced serum levels of MBL and increased susceptibility to microbial diseases. This study investigated whether MBL variants predispose to sarcoidosis by increasing their susceptibility to micro-organisms. METHODS: MBL gene promoter and exon 1 variants were detected by sequence specific primer polymerase chain reaction (SSP-PCR) in 167 UK Caucasian sarcoidosis patients and 164 control subjects. Severity of pulmonary disease outcome among patients was assessed by radiography after a minimum of 4 years from disease onset and classified as mild, moderate, and severe disease categories, accordingly. RESULTS: MBL variant frequencies were similar in patients and controls studied. Among sarcoidosis patients, the frequencies of variants were similar regardless of severity of disease outcome. The average patient ages at time of diagnosis were similar for all MBL genotypes. CONCLUSIONS: MBL gene variants do not appear to influence susceptibility to sarcoidosis, age of disease onset, or severity of disease. PMID- 10849026 TI - Metabolic and inflammatory responses to pulmonary exacerbation in adults with cystic fibrosis. AB - BACKGROUND: We hypothesized that increased resting energy expenditure in adults with cystic fibrosis was related to chronic inflammation secondary to pulmonary infection and could be modified by treatment of the underlying infection. METHOD: To determine the relationship between resting energy expenditure and the inflammatory and metabolic responses, we studied 22 adults with cystic fibrosis and chronic Pseudomonas aeruginosa infection before and after treatment of a respiratory exacerbation. Resting energy expenditure was measured by indirect calorimetry. Spirometry and circulating concentrations of C-reactive protein, neutrophil elastase alpha1-antiproteinase complex, catecholamines, non-esterified fatty acids and glycerol were determined. RESULTS: The mean (95% confidence interval)% predicted FEV1 was 28.5% (20.6, 36.4) and mean body weight 50.7 kg (47.4, 54.1). Following treatment, 1-s forced expiratory volume (FEV1) and weight increased, while C-reactive protein (P<0.0001) and neutrophil elastase alpha1 antiproteinase complex concentrations (P<0.0001) were reduced. Resting energy expenditure decreased from 6.8 (6.3, 7.2) to 6.25 (5.9, 6.6) MJ day-1 by day 15 (P<0.001). Changes in resting energy expenditure and C-reactive protein were related (r = 0.66, P< 0.0001). Weight gain was inversely related to resting energy expenditure (r = 0.43, P = 0.02) and unrelated to energy intake (r = 0.02, P = 0.47). Post-treatment reduction in norepinephrine was related to changes in heart rate (r = 0.57, P<0.01), resting energy expenditure (r = 0.51, P = 0.001) and non-esterified fatty acids (r = 0.42, P< 0.05). CONCLUSIONS: A parallel reduction in the host inflammatory and catabolic responses followed treatment of a respiratory exacerbation and may have contributed to weight gain. PMID- 10849027 TI - Effect of chronic analgesic exposure on the central serotonin system: a possible mechanism of analgesic abuse headache. AB - OBJECTIVE: To investigate the effects of chronic analgesic exposure on the central serotonin system and the relationship between the serotonin system and the analgesic efficacy of nonnarcotic analgesics. METHODS: Paracetamol was administered daily to adult male Wistar rats for a period of 15 or 30 days. Analgesic efficacy was measured by the tail flick test. After completion of the treatment protocol, the rats were humanely killed, and the frontal cortex and brain stem were isolated. Characteristics of the specific binding of the 5-HT2A serotonin receptor and the serotonin transporter were studied using a radioligand binding technique. Platelet serotonin was determined by high-performance liquid chromatography. RESULTS: Chronic paracetamol administration resulted in a significant decrease in the maximum number of 5-HT2A binding sites and an increase in the maximum number of 5-HT transporter binding sites in frontal cortical membrane (P<.001). Changes in the central 5-HT system were associated with a rise in platelet 5-HT levels. The degree of receptor downregulation, as well as transporter upregulation, became less evident after more prolonged drug administration. Readaptation of serotonin receptors and transporters coincided with the decrease in the analgesic efficacy of paracetamol, as well as a fall in platelet 5-HT levels. CONCLUSIONS: These findings provide further evidence in support of an involvement of the 5-HT system in the antinociceptive activity of simple nonnarcotic analgesics. Plasticity of this neurotransmitter system after chronic analgesic exposure may lead to the loss of analgesic efficacy and, in its more extreme form, may produce analgesic-related painful conditions, for example, analgesic abuse headache. PMID- 10849028 TI - Predisposing and provoking factors in childhood headache. AB - OBJECTIVE: To study the differences in predisposing and provoking factors between childhood migraine and nonmigrainous headache. BACKGROUND: Information on the predisposing and provoking factors of headache could help to find ways to prevent it. Differences in predisposing and provoking factors between migraine and nonmigrainous headache are largely unknown. METHODS: An unselected, population based, prospective, follow-up study on the occurrence of headache in schoolchildren was carried out in 1290 children aged from 8 to 9 years. The children who had reported headache during the 6 months prior to the study (n = 725) were sent a more detailed questionnaire about factors that might give rise to headache. Six hundred twenty-two (86%) children returned questionnaires that were completed to an acceptable degree. RESULTS: The occurrence of familial paroxysmal headache and unhappiness in the family independently predicted the occurrence of migraine in children, but this was not the case for nonmigrainous headache. CONCLUSIONS: In particular, the family occurrence of paroxysmal headache increases the risk of migraine in a child. The risk is still greater if their living conditions are experienced as unsatisfactory by the family. PMID- 10849029 TI - Recurrent headache, coping, and quality of life in children: a review. AB - OBJECTIVES: To clarify the concepts of coping with pain and quality of life (QoL) and to present a literature review of the strategies that children with recurrent headaches use to cope with their pain, the impact of recurrent headaches on children's QoL, and the influence of personal and situational variables on headache, coping, and QoL in children. METHODS: The literature search encompassed published articles that were found by means of a CD-ROM search of MEDLINE (1966 to December 1998) and PsycLIT (1974 to December 1998) and the snowball method. RESULTS: In pediatric headache research, only three studies have been found in which children report the use of various coping strategies, and only two studies considered QoL. Demographic factors and psychological variables such as depression, anger, and anxiety influence headache prevalence. The impact of headache-related variables such as headache type, severity, perceived cause, and prior experience on QoL has only been studied in adults. CONCLUSIONS: More research on coping and QoL is needed in pediatric headache. The conceptual model that is presented in this article may serve as a guide. PMID- 10849030 TI - Rizatriptan tablet versus wafer: patient preference. AB - After taking both conventional oral rizatriptan tablets and oral disintegrating rizatriptan tablets in the treatment of migraine with or without aura, patients were permitted to select their formulation preference. All adult patients who had requested continuation of rizatriptan during a 6-month period were included in the study. Of the 367 patients studied, 188 selected the oral disintegrating tablet, while 179 preferred the conventional tablet. Although individual patients had strong preferences for one preparation over the other, no group preference was found. PMID- 10849031 TI - Identification of patients with headache at risk of psychological distress. AB - OBJECTIVE: To test the hypothesis that anxiety and depression are associated with headache frequency, severity, and disability. BACKGROUND: There is significant comorbidity between chronic headache and psychological distress. Headache associated with anxiety or depression tends to be more severe and often requires supplementary psychological treatment in addition to headache therapy. Therefore, the identification of patients with headache who are at risk of psychological distress is important. METHODS: One hundred twenty-seven consecutive patients with headache attending a university headache clinic were evaluated. Questionnaires about headache symptoms and psychological distress were completed. Comparisons were made between psychological distress and headache frequency, severity, and disability. RESULTS: Depression and anxiety were significantly greater in the subjects of this study who had frequent headache (>4 days per week) and frequent headache-associated disability (activities reduced or prohibited because of headache >3 days per week). Subjects who reported their headache severity as typically severe were no more likely to report depression or anxiety than those with mild or moderate headache severity. Quality-of-life measures of physical and social functioning, emotional distress, and general health and vitality were reduced in subjects with frequent episodes of headache associated disability. All areas, with the exception of general health perception, were reduced in subjects with frequent headache. Severe headache was associated with reductions in role and social functioning. CONCLUSIONS: Frequent headache and frequent disability are associated with depression, anxiety, and impaired quality of life. Reports of typical headache severity are less likely to correlate with psychological distress. Therefore, patients with headache who report frequent headache or frequent periods of headache-related disability should be further evaluated for the presence of psychological disturbance. PMID- 10849032 TI - Behavioral self-management in an inpatient headache treatment unit: increasing adherence and relationship to changes in affective distress. AB - OBJECTIVE: To evaluate prospectively the contribution of a psychological self management program to the amelioration of headache-related distress of patients with intractable migraine treated in a comprehensive, multidisciplinary, inpatient program. BACKGROUND: Previous research has shown the effectiveness of this overall inpatient program but did not examine the relationships between the use of relaxation and other headache-related behavioral factors. METHODS: Data from 221 admissions to a Commission on Accreditation of Rehabilitation Facilities accredited, nationally recognized, inpatient treatment unit were analyzed for the current study. On admission and on discharge (average length of stay, 12.9 days), subjects completed a 7-day retrospective, self-report questionnaire assessing health behavior compliance and emotional factors. The intervention consisted of intensive medical therapy in addition to cognitive-behavioral treatment delivered in a group setting. RESULTS: Adherence increased significantly for relaxation practice and life-style modifications of diet, exercise, and sleep regulation for headache prevention (P<.00001). Beck Depression Inventory scores decreased significantly (P<.00001), and a greater decrease in depression by the end of the program was reported by subjects who practiced relaxation most compared with those who practiced relaxation least. CONCLUSIONS: Low baseline adherence rates for health behavior increased significantly during the final week of inpatient treatment. Behavioral self-management variables, not headache reduction, were significantly associated with patients' reduction in affective distress. PMID- 10849033 TI - Influence of disease duration on visual evoked potentials in migraineurs. AB - OBJECTIVE: To determine the possible influence of the duration of migraine on pattern-reversal visual evoked potentials. METHODS: An investigation was conducted in 49 patients with migraine without aura according to the International Headache Society criteria. Twenty-two of these patients had had migraine for 2 years or less (group 1), and the other 27 patients had had the disease for 10 years or more (group 2). The control group consisted of 17 healthy subjects. RESULTS: Comparison of the mean P100 latency and amplitude showed no significant difference among the groups. There was, however, a good negative correlation between age and latency (r = -0.59, P =.003) in group 1, but no such correlation was observed for group 2 or the control group. CONCLUSIONS: It was concluded that the duration of migraine has no influence on pattern-reversal visual evoked potentials and that the pathogenesis of early- and late-onset migraine may be different. PMID- 10849034 TI - Daily migraine with aura: a new migraine variant. AB - The frequency of migraine attacks is not used as a diagnostic criterion, however, it is a very important factor in the evaluation of migraine severity and its treatment. Several studies report the frequency of migraine attacks using the International Headache Society criteria. No investigator, however, has reported daily migraine attacks. In the current report, we present five patients whose headaches transformed from episodic migraine to daily migraine with aura. To the best of our knowledge, this is the first description of this variant of migraine. The symptoms in all the patients described comply strictly with the International Headache Society criteria for the diagnosis of migraine with typical aura. An interesting additional observation concerns the beneficial effect of phenytoin, a drug that has not proved to be effective in migraine but showed some efficacy in our patients. PMID- 10849035 TI - Tolosa-Hunt syndrome preceded by facial palsy. AB - The Tolosa-Hunt syndrome consists of a painful ophthalmoplegia related to a granulomatous inflammatory process in the cavernous sinus, which may be documented by cerebral magnetic resonance imaging with gadolinium enhancement. Two cases of Tolosa-Hunt syndrome preceded by facial palsy observed in 1998 at the Department of Neurosurgery of the Second University of Naples are presented here. Both patients developed Tolosa-Hunt syndrome following an ipsilateral facial palsy that resolved in about 15 days with medical treatment. Cerebral magnetic resonance imaging with gadolinium enhancement showed, in both cases, inflammatory tissue in the cavernous sinus. The patients underwent corticosteroid therapy (prednisolone, 80 mg per day, intravenously) with pain regression. In the first case, the patient experienced recurrence of the syndrome that was definitively resolved with further corticosteroid treatment. The rare reports of facial palsy in patients with Tolosa-Hunt syndrome suggest the inclusion of this disease in the so-called multiple cranial nerve palsy syndrome. It is probable that Tolosa-Hunt syndrome has an inflammatory pathogenesis. PMID- 10849036 TI - Acute intracranial hemorrhage caused by acupuncture. AB - A 44-year-old Chinese man developed severe occipital headache, nausea, and vomiting during acupuncture treatment of the posterior neck for chronic neck pain. Computed tomography of the head showed hemorrhage in the fourth, third, and lateral ventricles. A lumbar puncture confirmed the presence of blood. Magnetic resonance angiography with gadolinium did not reveal any saccular aneurysms or arteriovenous malformations. The patient's headache resolved over a period of 28 days without any neurological deficits. Acupuncture of the posterior neck can cause acute intracranial hemorrhage. PMID- 10849037 TI - Headache relief after sumatriptan in a patient with pituitary macroadenoma. PMID- 10849039 TI - Botulinum toxin type A as a migraine preventive treatment. For the BOTOX Migraine Clinical Research Group. AB - OBJECTIVE: To assess the safety and efficacy of botulinum toxin type A (BOTOX; Allergan, Inc) in the prevention of migraine. BACKGROUND: Current migraine preventive therapies are often unsatisfactory because of their limited efficacy, adverse effects, and drug interactions. Botulinum toxin type A injections often reduce the pain associated with conditions such as cervical dystonia, achalasia, rectal fissures, and myofascial pain syndrome. An open-label, noncontrolled study of botulinum toxin type A suggested benefits for patients with migraine. DESIGN AND METHODS: This was a double-blind, vehicle-controlled study of 123 subjects with a history of two to eight moderate-to-severe migraine attacks per month, with or without aura. Participants were randomized to receive single administrations of vehicle or botulinum toxin type A, 25 U or 75 U, injected into multiple sites of pericranial muscles at the same visit. During a 1-month baseline period and for 3 months following injection, subjects kept daily diaries in which they recorded migraine frequency, migraine severity, and the occurrence of migraine-associated symptoms. RESULTS: Compared with vehicle treatment, subjects in the 25-U botulinum toxin type A treatment group showed significantly fewer migraine attacks per month, a reduced maximum severity of migraines, a reduced number of days using acute migraine medications, and reduced incidence of migraine-associated vomiting. Both the 25-U and 75-U botulinum toxin type A groups were significantly better than the vehicle group on subject global assessment. Botulinum toxin A treatment was well tolerated, with only the 75-U treatment group exhibiting a significantly higher rate of treatment-related adverse events than vehicle. CONCLUSIONS: Pericranial injection of botulinum toxin type A, 25 U, was found to be a safe treatment that significantly reduced migraine frequency, migraine severity, acute medication usage, and associated vomiting. PMID- 10849040 TI - L-5-Hydroxytryptophan in the prophylaxis of chronic tension-type headache: a double-blind, randomized, placebo-controlled study. For the Portuguese Head Society. AB - A parallel, randomized, double-blind trial is reported, in which 78 patients with chronic tension-type headaches were treated with L-5-hydroxytryptophan (5-HTP) (n = 43) or placebo (n = 35) for 8 weeks, after a washout period of 2 weeks and with a follow-up period of a further 2 weeks. Five patients dropped out (1 with placebo and 4 with 5-HTP), 1 was excluded due to noncompliance, and in 7 treatment was suspended due to adverse events (3 with placebo and 4 with 5-HTP), yielding a final number for evaluation of 65 patients. In comparison with the group treated with placebo, there was no statistically significant change in the number of days with headache or in headache intensity in the group treated with 5 HTP, 300 mg per day; however, there was a significant decrease in the consumption of analgesics. During the 2 weeks after treatment, there was a significant decrease in the number of days with headache. Subjective opinion during this latter period was also favorable to 5-HTP. PMID- 10849041 TI - Cosinor analysis of heart rate variability in ambulatory migraineurs. AB - OBJECTIVE: To clarify whether the circadian rhythm of heart rate variability parameters can be identified in patients with migraine during a headache-free period and to identify any specific pattern of the circadian rhythm of heart rate variability, using time-domain and spectral analysis and cosinor rhythm analysis of heart rate variability during normal daily activity. METHODS: Forty-eight-hour Holter electrocardiograms were recorded for 27 patients with migraine during headache-free periods and 24 healthy controls during free activity. The circadian rhythms of heart rate fluctuation parameters, that is, mean interval, standard deviation, root-mean-square successive differences (RMSSD), %RR50, and low (0. 020 to 0.150 Hz) and high frequency (0.150 to 0.400 Hz) heart rate fluctuations were analyzed with the group mean cosinor method. RESULTS: The group mean cosinor analysis and the acrophase-amplitude analysis demonstrated significant differences in circadian rhythm in SD, RMSSD, %RR50, and high frequency between the group with migraine and controls. The amplitudes of SD, RMSSD, %RR50, and high frequency in the group with migraine were smaller than those in controls, which implied parasympathetic hypofunction in migraineurs. There were no significant differences in the MESOR (midline estimating statistic of rhythms) of the analyzed heart rate parameters except for low frequency. The MESOR of low frequency in the migraineurs was significantly smaller than that in the controls. CONCLUSIONS: Patients with migraine have hypofunction in the parasympathetic nervous system during normal daily activity in the headache-free period. PMID- 10849042 TI - Treatment of nonresponders to oral sumatriptan with zolmitriptan and rizatriptan: a comparative open trial. AB - In order to study the effect of zolmitriptan and rizatriptan in oral sumatriptan nonresponders (defined as lack of response in three or more of five attacks), 56 patients were studied in an open trial in a crossover fashion. Both zolmitriptan, 5 mg, and rizatriptan, 10 mg, were effective in the majority of sumatriptan nonresponders. The response to rizatriptan, 10 mg, appeared to be better than to zolmitriptan, 5 mg. Approximately 19% of sumatriptan nonresponders remained nonresponders to both zolmitriptan and rizatriptan. PMID- 10849043 TI - MMPI-2 profiles in chronic daily headache and their relationship to anxiety levels and accompanying symptoms. AB - OBJECTIVES: To examine a group of patients with chronic daily headache using the revised version of the Minnesota Multiphasic Personality Inventory (MMPI-2) and to determine whether the data acquired were related to the anxiety levels of the patients, as detected by the Spielberger State-Trait Anxiety Inventory (STAI) 1, 2 and to the presence of a number of accompanying symptoms that are frequently observed in patients with chronic headache. BACKGROUND: In the last decade, the MMPI-2 was released and its items used to develop 15 "content scales." Recently, this instrument was adapted to the Italian population. METHODS: Five men and 30 women with chronic daily headache had a semistructured interview in which the presence of 21 behavioral or somatic symptoms was recorded. The Italian version of the MMPI-2 and the STAI 1, 2 (Italian version) were employed. A configural analysis of the MMPI profiles was performed, and four types were distinguished: "conversion V" (n = 5), elevation of the "neurotic triad" (n = 5), the "emotionally overwhelmed" with scale elevation of the neurotic triad and of several other scales (n = 18), and "the copers" with no scale elevation above 65 (n = 4). Three patients could not be classified. The pain characteristics, the prevalence of accompanying symptoms, and the STAI 1, 2 scores were assessed in all patients and in the different MMPI groups, and the data were statistically analyzed (ANOVA and chi-square analysis). RESULTS: All patients with no MMPI-2 scale elevation showed a tendency to a conversion V profile: in this group, the chronicity was markedly and significantly lower than in all other groups. Moreover, in this group, the STAI 1, 2 scores and the prevalence of some accompanying symptoms were significantly lower than in the other groups. Migraine characteristics did not differ significantly from group to group. CONCLUSIONS: Hysterical traits were observed in a number of patients with chronic daily headache and might constitute a predisposing factor for this condition. With time, the personality profile deteriorates, either through an increase in the hysterical traits or through its transformation, with a parallel increase in anxiety levels and the presence of accompanying symptoms. PMID- 10849044 TI - Prevalence of migraine, tension-type headache, and other headaches in Hong Kong. AB - OBJECTIVE: To assess the prevalence of migraine and other headaches in Hong Kong in 1998. BACKGROUND: A community-based prevalence survey of headache was carried out from July 1992 to March 1993, and the prevalence rates were 1% for migraine, 2% for tension-type headache, and 1% for other headaches. A similar survey was carried out in May and June 1998 to interview individuals aged 15 years or older. Recurrent headache was defined as having two or more headaches unrelated either to influenza or a common cold within the past 12 months. METHODS: Respondents with recurrent headache were offered a personal interview for clinical validation. Of 3156 randomly selected individuals, 1436 responded. RESULTS: Headache was due to influenza or a common cold in 270 (18.8%) respondents; recurrent headache affected 533 (37.1%) respondents. The overall prevalence rates were 4.7% for migraine, 26.9% for tension-type headache, and 5.5% for other headaches. Clinical validation was available for 72 respondents. After adjustment for possible misclassification, the estimated prevalence rates became 12.5% for migraine, 18.7% for tension-type headache, and 6.0% for other headaches. There was a female preponderance for all types of headache with a peak in the 25- to 34 year-age group for tension-type headache. CONCLUSIONS: All types of headache were more common in the 1998 study, and the prevalence rates were closer to those of Western communities. PMID- 10849045 TI - Bilateral subdural hematomas following routine lumbar diskectomy. AB - Intracranial hypotension is a rare, and possibly underrecognized, cause of headache in middle age. Occurring spontaneously in the vast majority of cases, it has been occasionally reported after certain neurosurgical procedures involving craniectomy. We report a unique situation in which a patient developed severe postural headache typical of intracranial hypotension, which was complicated by bilateral subdural hematomas, immediately following a routine lumbar diskectomy; the headache resolved spontaneously. We suggest that an intraoperative microscopic dural breach was the site of sustained, but self-limited, cerebrospinal fluid leakage that eventually led to intracranial hypotension. PMID- 10849046 TI - LASH: a syndrome of long-lasting autonomic symptoms with hemicrania (A new indomethacin- responsive headache). AB - A patient presented with a unique, stereotypical, episodic headache disorder marked by long-lasting autonomic symptoms with associated hemicrania (LASH). The autonomic symptoms clearly overshadowed the headache as the major component of the syndrome. Indomethacin controlled both the autonomic symptoms and the headache, suggesting that this is a new type of indomethacin-responsive headache. It may also complete the indomethacin-responsive headache spectrum. PMID- 10849047 TI - Migraine-associated seizure: a case of reversible MRI abnormalities and persistent nondominant hemisphere syndrome. AB - The complex relationship between migraine and epilepsy is highlighted by the occurrence of a seizure during a migraine attack without aura. This phenomenon, referred to as migralepsy, suggests an inherent overlap in the underlying pathophysiology of these events. We report the case of a patient who had a generalized seizure, persistent nondominant hemisphere syndrome, and reversible magnetic resonance imaging abnormalities during a prolonged migraine attack without aura. PMID- 10849048 TI - Expert opinion: orgasmic headaches: clinical features, diagnosis, and management. PMID- 10849049 TI - Expert opinion: the management of pseudotumor cerebri during pregnancy. PMID- 10849050 TI - Intranasal lidocaine for migraine using a metered-dose spray. PMID- 10849051 TI - Naratriptan prophylaxis of transformed migraine or hemicrania continua? PMID- 10849053 TI - Helicobacter pylori "test-and-scope" strategy for dyspeptic patients. PMID- 10849054 TI - Different Helicobacter pylori strains colonize the antral and duodenal mucosa of duodenal ulcer patients. AB - BACKGROUND: We have investigated the possibility that the same patients may be colonized by Helicobacter pylori strains of different genotypes or phenotypes in the antrum as compared to in the duodenum. The strains were typed for DNA fingerprints, different lipopolysaccharides (LPS), and Lewis antigen expression on the O-side chains of LPS. MATERIALS AND METHODS: Polymerase chain reaction (PCR) amplifications using primer sequences (i.e., the Enterobacterial Repetitive Intergenic Consensus [ERIC]) and randomly amplified polymorphic DNA (RAPD) elements were performed to asses chromosomal DNA diversity between H. pylori strains. The expression of different LPS types and Lewis antigens in the various H. pylori isolates were determined by whole bacterial enzyme-linked immunosorbent assays using monoclonal antibodies. RESULTS: Duodenal ulcer patients had different H. pylori genotypes in the duodenum as compared to in the antrum as shown by ERIC-PCR (44%) and by RAPD-PCR (75%). Different DNA patterns were found among the strains that were isolated from various regions of the duodenum in 4 of 16 patients (25%) as shown by ERIC-PCR and in 8 of 16 patients (50%) as shown by RAPD-PCR. Sixty-three percent of the duodenal ulcer patients had H. pylori strains with a different Lewis antigen phenotype in the duodenum as compared to in the antrum, and 3 of 16 patients (19%) had strains with different Lewis antigens expressed by strains from different duodenal biopsies from the same patient. CONCLUSION: The results suggest that a mixed population of different H. pylori strains with marked variation, both genotypically and phenotypically, colonize the same patient. PMID- 10849055 TI - Prevalence of Helicobacter pylori resistance to metronidazole, clarithromycin, amoxicillin, tetracycline, and furazolidone in Brazil. AB - BACKGROUND: Helicobacter pylori infection is associated with a wide range of digestive diseases and is very prevalent in developing countries, although few data exist on the susceptibility of H. pylori to antimicrobials commonly used in eradication schedules in these countries. The aim of this study was to evaluate the resistance of H. pylori to metronidazole, clarithromycin, amoxicillin, tetracycline, and furazolidone in dyspeptic Brazilian patients. Material and Methods. Ninety consecutive H. pylori-positive patients were enrolled. Resistance was evaluated by an agar dilution test. RESULTS: Resistance to metronidazole was detected in 38 patients (42%); to amoxicillin in 26 individuals (29%); to clarithromycin in 6 patients (7%); to tetracycline in 6 patients (7%); and to furazolidone in 4 individuals (4%). Thirteen strains were resistant to two agents, and eight strains were resistant to three antimicrobials. CONCLUSIONS: These results confirm the need for culture and susceptibility testing to define H. pylori resistance patterns in particular geographical areas before the general use of an eradication schedule. They also suggest the possibility of resistance to such antimicrobials as amoxicillin or tetracycline in geographical areas with a high prevalence of H. pylori infection and still not fully evaluated for antimicrobial susceptibility. PMID- 10849056 TI - Comparison of seven and fourteen days of lansoprazole, clarithromycin, and amoxicillin therapy for eradication of Helicobacter pylori: a report from India. AB - BACKGROUND: In developed countries, a 1-week regimen of combined proton pump inhibitors and two antibiotics is considered adequate for Helicobacter pylori eradication. However, there is a paucity of reports from developing countries on treatment duration of less than 14 days. We compared efficacy of 7 and 14 days of lansoprazole (L), clarithromycin (C), and amoxicillin (A) combinations for eradication of H. pylori. PATIENTS AND METHODS: Forty-six consecutive patients who presented with upper gastrointestinal symptoms and tested positive for H. pylori infection were included in the study. In every patient, after performance of upper gastrointestinal endoscopy, antral biopsies were obtained. H. pylori infection was diagnosed by positive rapid urease test and identification of organisms on antral histology. Patients were randomly selected to receive lansoprazole, 30 mg once daily, plus clarithromycin, 250 mg twice daily, plus amoxicillin, 500 mg three times daily for 2 weeks (group 1; n = 24; age, 36 +/- 12 years; 18 men) or 1 week (group 2; n = 22; age, 45 +/- 15 years; 12 men). One month after completion of treatment, repeat upper gastrointestinal endoscopy was performed. H. pylori eradication was defined as absence of organism on histopathological examination of both antrum and body of stomach and negative rapid urease test. RESULTS: Eradication rate was higher in group 1 (23 of 24; 96%) as compared to group 2 (12 of 22; 54%; p <.05). One patient in group 1 had diarrhea, and one patient in group two had skin rash and itching. CONCLUSIONS: Fourteen-day therapy with lansoprazole, clarithromycin, and amoxicillin is highly effective in eradication of H. pylori. Reducing duration of therapy to 7 days significantly lowers eradication rates. PMID- 10849057 TI - Addition of metronidazole to rabeprazole-amoxicillin-clarithromycin regimen for Helicobacter pylori infection provides an excellent cure rate with five-day therapy. AB - BACKGROUND: New triple therapy for eradication of Helicobacter pylori based on a proton pump inhibitor (PPI) provides a cure rate of approximately 90% with few adverse effects. Recently, a PPI-based quadruple therapy, which consists of a PPI plus bismuth-based triple therapy for 7 days, has been studied, and a sufficient eradication rate has been achieved. However, a shorter duration results in improved compliance. In this study, newly developed short-term, simple twice daily quadruple therapy consisting of rabeprazole, amoxicillin, clarithromycin, and metronidazole (RACM) was compared with a PPI-based triple-therapy regimen for eradication of H. pylori. PATIENTS AND METHODS: This study was designed as a randomized open, prospective single-center study. Of a total of 105 H. pylori positive patients, 55 received the RACM regimen for 5 days (rabeprazole, 10 mg bid; amoxicillin, 750 mg bid; clarithromycin, 200 mg bid; and metronidazole, 250 mg bid), and 50 received the RAC regimen for 5 days (rabeprazole, 10 mg bid; amoxicillin, 750 mg bid; and clarithromycin, 200 mg bid). Cure of the infection was assessed by HpSA (H. pylori stool antigen immunoassay) 1 month after completion of therapy. RESULTS: The rates of eradication of H. pylori by RACM versus RAC were 94.5% (95% CI, 85-99) versus 80.0% (95% CI, 66-90) by intention to-treat analysis; 98.1% (95% CI, 90-100) versus 87.0% (95% CI, 74-95) by all patients-treated analysis; and 98.1% (95% CI, 90-100) versus 86.7% (95% CI, 73 95) by per-protocol analysis. No major adverse effects were reported, and 98.0% of patients reported complete compliance. CONCLUSIONS: The simple twice-daily and short-term quadruple regimen for only 5 days provided an excellent eradication rate. Compliance with the regimen was high, and serious adverse effects were few. Therefore, the RACM regimen can be considered as safe and effective. PMID- 10849058 TI - Controlled trial of the effect of cinnamon extract on Helicobacter pylori. AB - BACKGROUND: Helicobacter pylori has been associated with the pathogenesis of antral gastritis, duodenal ulcer, and gastric lymphoma. Eradication of H. pylori has been shown to reverse or prevent relapse of these diseases. Antimicrobials employed in the eradication of H. pylori are not without adverse effects. Newer treatment modalities, therefore, are required. In vitro studies have shown the effectiveness of cinnamon extract against H. pylori and its urease. In this pilot study, we tested the activity of an alcoholic extract of cinnamon in a group of patients infected with H. pylori. MATERIALS AND METHODS: Fifteen patients (11 women, 4 men) aged 16 to 79 years were given 40 mg of an alcoholic cinnamon extract twice daily for 4 weeks; eight patients aged 35 to 79 (7 women, 1 man) received placebo. The amount of H. pylori colonization was measured by the 13C urea breath test before and after therapy. RESULTS: The mean urea breath test counts in the study and control groups before and after therapy were 22.1 and 23.9 versus 24.4 and 25.9, respectively. The cinnamon extract was well tolerated, and side effects were minimal. CONCLUSIONS: We concluded that cinnamon extract, at a concentration of 80 mg /day as a single agent, is ineffective in eradicating H. pylori. Combination of cinnamon with other antimicrobials, or cinnamon extract at a higher concentration, however, may prove useful. PMID- 10849059 TI - Factors that affect results of the 13C urea breath test in Japanese patients. AB - BACKGROUND: The 13C urea breath test (UBT) is considered to be the most accurate way of diagnosing Helicobacter pylori infection. Our objective was to investigate the accuracy of the UBT in Japanese patients and the association of UBT values with histological findings. MATERIALS AND METHODS: A total of 169 consecutive patients were studied by endoscopy with histology, by serology with IgG antibody and test serum pepsinogen (PG), and by UBT. The association between UBT values and histological findings and the PG I / II ratio were analyzed in H. pylori positive patients. RESULTS: Of 169 Japanese patients, 135 were H. pylori-positive on both histology and serology analysis, 27 were H. pylori-negative on both histology and serology analysis, and 7 patients showed differing results. Using a cutoff value of 2.5 per thousand, test sensitivity was 100%, while specificity was 96%. Among the 135 H. pylori-positive patients, a significant relation was observed between UBT value and H. pylori colonization density of the corpus and antrum, neutrophil activity of the antrum, atrophy, and intestinal metaplasia of the corpus in the H. pylori-positive patients. Also, UBT values correlated with the PG I /II ratio. In multivariate analysis, the PG I /II ratio was the most important factor related to UBT values (odds ration [OR], 4. 99; 95% confidence interval, 1.60-15.55). CONCLUSIONS: The UBT is an accurate method for detecting H. pylori infection in the Japanese population, which shows a high prevalence of atrophic gastritis. Values are affected by H. pylori infection and by the severity of atrophic gastritis. PMID- 10849060 TI - 13C urea breath test with nondispersive isotope-selective infrared spectrometry: reproducibility and importance of the fasting status. AB - BACKGROUND: The 13C urea breath test (13C-UBT) is the most convenient method for diagnosing Helicobacter pylori infection noninvasively. Nondispersive isotope selective infrared spectrometry (NDIRS) is an inexpensive and easy alternative to mass spectrometry. The objective of this study was to evaluate: (1) the reproducibility of the 13C-UBT as performed by using the NDIRS method; (2) the repeatability of bags analysis and the impact of delayed analysis; and (3) the need for fasting status for the 13C-UBT. METHODS: The 13C-UBT was performed with 75 mg urea labeled with 13C, with breath samples collected at times 0 and 30 minutes. Results are expressed as delta over baseline (0/00). Fifty-three patients underwent two successive 13C-UBTs with an interval of 48 to 72 hours. The 106 collected bags were randomly reanalyzed immediately or 72 hours later. In 26 volunteer subjects, the 13C-UBT was performed both in a fasting condition and after a nonstandardized meal. The reproducibility was assessed by the method of Bland and Altman. RESULTS: The mean of difference between two successive tests was 0. 14 0/00 (standard deviation, 0.90), and the coefficient of repeatability was 1.80 (confidence interval, 95%). The difference between two successive analyses was always less than 2.2% of the initial value. The coefficient of variation between two successive tests for the influence of a meal was 11.24. CONCLUSION: The 13C-UBT as performed by using NDIRS is reproducible, analyses can be delayed up to 72 hours, and the test must be performed in fasting conditions. PMID- 10849061 TI - Clinical usefulness of urine-based enzyme-linked immunosorbent assay for detection of antibody to Helicobacter pylori: a collaborative study in nine medical institutions in Japan. AB - BACKGROUND: A urine-based enzyme-linked immunosorbent assay (ELISA) kit for detection of antibody to Helicobacter pylori has been developed in Japan. Urine samples can be obtained noninvasively and are easier and safer to handle than are serum samples. The aim of this study was to examine the clinical usefulness of this urine-based ELISA kit. MATERIALS AND METHODS: A pair of random, single-void urine and serum samples was collected from each of 1,061 subjects, including 238 patients with gastroduodenal disease. The sensitivity and specificity of the urine-based ELISA was compared with those of three commercially available serum based ELISA kits. For those patients with gastroduodenal disease, the urine- and serum-based ELISA results were also compared with those for other diagnostic methods using endoscopic biopsy specimens, such as culture, histology, and rapid urease tests. RESULTS: Based on the three serum-based ELISA results, the sensitivity, specificity, and accuracy of the urine-based ELISA were 97.7%, 95.6%, and 96.8%, respectively. On the basis of the biopsy test results, the sensitivity (96.2%), specificity (78.9%), and accuracy (91.0%) of the urine-based ELISA were almost equivalent or superior to all three serum-based ELISAs tested. In addition, 10 of the 12 false-positive cases for urine-based ELISA were confirmed to be true positives for antibodies to H. pylori by Western blot analysis and inhibition ELISA. CONCLUSIONS: The urine-based ELISA (URINELISA H. pylori Antibody) is very accurate and should be useful as an alternative to serum based ELISAs for screening of H. pylori infection. PMID- 10849062 TI - The evolution of gene number: are heritable and non-heritable errors equally important? AB - Is there a limit to the number of genes carried by an organism? Two reasons have been. First, as most mutations are deleterious, for a given per locus mutation rate there must exist an upper limit to the number of genes that is consistent with individual survival. Second, the imprecision of the mechanisms governing gene expression might also restrict genomic complexity. As gene expression errors are probably much more common than mutations, it is the latter that are more likely to impose a limit. However, these errors are not heritable and therefore cannot accumulate in populations. Which of the two sorts of effect are more likely to impose a limit? We address this issue in two ways. First, we ask about the load imposed by each sort of error. We show that the harmful effect of non heritable failures is higher than that of heritable mutations, if (p) x (delta) > mu, where p is the rate of non-heritable failures, delta measures the harmful effect of these failures and mu is the rate of heritable mutations. Therefore, although the rate of non-heritable errors might be very high, this does not demonstrate that they are more important than mutations as their impact must be discounted by the strength of their effects. Further, we note that both theory and evidence suggest that the most common errors are of the least importance. Second, we discuss the population genetics of a new gene duplication. Previous attempts to make a connection between error rates and limits on gene number are based on group selection arguments. These fail to show a direct limitation on the spread of gene duplications. We note that empirical evidence indicates that duplication per se tends to result in expression errors that may be heritable. We therefore argue that a hybrid model, one evoking heritable expression errors, is likely to be the most realistic. PMID- 10849063 TI - Genetic structure and colonizing success of a clonal, weedy species, Pilosella officinarum (Asteraceae). AB - Introduced populations of weeds which are polyploid and reproduce primarily by apomixis are generally considered as having low levels of population genetic variation, highly differentiated populations and short evolutionary lifespans. Although polyploidy allows for habitat differentiation and colonization, lack of recombination because of apomixis means that long-term persistence is unlikely. However, variation can be introduced to a colonizing population by evolutionary changes in the mating system, or by somatic mutation and recombination. In this study hypersensitive genetic markers, inter-simple sequence repeats (ISSRs), were used to quantify genetic variation within Pilosella officinarum, a major weed of the New Zealand high country. Pilosella officinarum was introduced from Europe to New Zealand late in the 19th century and only polyploid, apomictic populations are thought to have survived. The combination of introduction history and breeding system has led to the assumption that New Zealand populations are necessarily genetically depauperate. However, our studies reveal variable levels of genetic variation and patterns of clonal distribution which indicate varying levels of sexual reproduction within New Zealand populations. PMID- 10849064 TI - Mating systems of diploid and allotetraploid populations of Tragopogon (Asteraceae). II. Artificial populations. AB - Polyploidization has long been recognized as an important force in the diversification of plants. Theoretical models predict that polyploids may be expected to exhibit higher rates of self-fertilization than do closely related diploid species. Wild populations of the neopolyploid Tragopogon mirus (4n) exhibited slightly higher rates of outcrossing than did populations of one of its progenitors, T. dubius (2n). In the current study, outcrossing rates in populations of T. dubius and T. mirus were estimated using artificial arrays constructed to maximize the chances of detecting outcrossing events. The artificial diploid population is more highly outcrossing (t=0.727; family-level estimates range from 0.00 to 1. 32) than the tetraploid population (t=0.591; family-level estimates range from 0.00 to 1.14), although the difference between them is not statistically significant. The results of this study, combined with those of the previous work on wild populations, suggest that mating systems in these species vary more among populations than between ploidal levels. This could be because of the relatively recent origins of the tetraploid species; there may have been insufficient time since the formations of the tetraploids for shifts in mating systems to occur. PMID- 10849065 TI - Correlated response in reproductive and life history traits to selection on testis length in Drosophila hydei. AB - Flies in the genus Drosophila have undergone striking evolutionary divergence in the size and number of sperm produced. Based on comparative studies of sperm length, testis length, and other reproductive and life history traits, including body size, age at first reproduction, and the number of sperm produced, macroevolutionary trade-offs resulting from the need to produce high-investment testes have been postulated. To understand better the microevolutionary processes underlying these interspecific patterns, we imposed replicated bidirectional selection for testis length for 11-12 generations on D. hydei, a species with 23.5 mm-long sperm and 30 mm-long testes. Testis length exhibited realized heritabilities ranging from 0.45 to 0.72. Following selection, traits were assayed for correlated responses. Thorax length, testis mass, sperm length, egg to-adult development time, and posteclosion maturation time showed consistent positive correlated responses. Numbers of sperm produced and transferred to females, male longevity, female egg productivity, and seminal receptacle length did not show consistent correlated responses to selection on testis length. The pattern of correlated responses to testis length reveal the potential for the evolution of reproductive strategies to alter important life history attributes. PMID- 10849066 TI - Estimating variance components in natural populations using inferred relationships. AB - Until recently, the estimation of the heritability of a trait has required knowledge of the pedigree within a population. In natural populations such knowledge is often unknown. Two techniques have been developed which use marker information to estimate heritabilities without reference to the exact nature of the relationships: a regression-based estimator that regresses phenotypic similarity for a pair of individuals against an estimate of their relationship and a likelihood-based estimator that maximizes the probability of the genotypic and phenotypic data given a known population structure. Computer simulation was used to compare the behaviour of these estimators. Bias in estimates of heritability decreased with increasing marker information, decreasing simulated heritability, increasing relatedness and increasing sample size. The techniques displayed reasonable tolerance to the percentage of missing data. The regression based technique shows least average bias, but largest variance over simulations. Likelihood-based techniques show larger average bias, but smaller variances over estimates. A modified form of the likelihood technique, requiring fewer initial assumptions about population parameters, is presented. The modified form shows less bias in its estimates of heritability than the likelihood technique originally proposed. PMID- 10849067 TI - Optimal marker density for interval mapping in a backcross population. AB - An important question in QTL mapping is the optimal choice of marker density. Using analytical results, it is shown for the case of interval mapping in a backcross population, that the power of QTL detection and the standard errors of genetic effect estimates are little affected by an increase of marker density beyond 10 cM. This finding confirms published simulation results by other authors. PMID- 10849068 TI - The likelihood of homoploid hybrid speciation. AB - New species may be formed through hybridization and without an increase in ploidy. The challenge is for hybrid derivatives to escape the homogenizing effects of gene flow from parental species. The mechanisms hypothesized to underlie this process were modelled using a computer simulation. The model is of recombinational speciation, in which chromosomal rearrangements between parental species result in poor fertility of F1 hybrids, but through recombination, novel homozygous types are formed that have restored fertility. In simulations, stable populations bearing the recombinant karyotypes originated frequently and were maintained when the fertility of F1 hybrids was high. However, this high rate of origination was offset by low genetic isolation, and lower F1 hybrid fertility increased the evolutionary independence of derived populations. In addition, simulations showed that ecological and spatial isolation were required to achieve substantial reproductive isolation of incipient species. In the model, the opportunity for ecological isolation arose as a result of adaptation to extreme habitats not occupied by parental species, and any form of spatial isolation (e.g. founder events) contributed to genetic isolation. Our results confirmed the importance of the combination of factors that had been emphasized in verbal models and illustrate the trade-off between the frequency at which hybrid species arise and the genetic integrity of incipient species. PMID- 10849069 TI - Local drift load and the heterosis of interconnected populations. AB - We use Wright's distribution of equilibrium allele frequency to demonstrate that hybrids between populations interconnected by low to moderate levels of migration can have large positive heterosis, especially if the populations are small in size. Beneficial alleles neither fix in all populations nor equilibrate at the same frequency. Instead, populations reach a mutation-selection-drift-migration balance with sufficient among-population variance that some partially recessive, deleterious mutations can be masked upon crossbreeding. This heterosis is greatest with intermediate mutation rates, intermediate selection coefficients, low migration rates and recessive alleles. Hybrid vigour should not be taken as evidence for the complete isolation of populations. Moreover, we show that heterosis in crosses between populations has a different genetic basis than inbreeding depression within populations and is much more likely to result from alleles of intermediate effect. PMID- 10849070 TI - Mitochondrial DNA and chromosomal studies of wild mice (Mus) from Turkey and Iran. AB - Complete D-loop sequences of 20 Mus from three localities in Turkey and seven in Iran were characterized. These countries are thought to be close to the place of origin of the subspecies Mus musculus domesticus. Five new M. m. domesticus haplotypes were added to the nine already known for the region. Four of these 14 haplotypes were very similar to the consensus D-loop sequence for western Europe defined by Nachman et al. (1994), which may represent the ancestral condition for M. m. domesticus. A divergent mtDNA lineage is found in various parts of Turkey and northern Iran; it has spread into western Europe, but other European lineages were not found in either Turkey or Iran. The other Mus D-loop sequences were of M. m. castaneus and Mus macedonicus and confirmed M. macedonicus as a monotypic species with low nucleotide diversity. The prevalence of the standard 40 chromosome complement in this region is particularly interesting with regards M. m. domesticus, as it is consistent with the in situ origin of Robertsonian karyotypic races (2n < 40) in western Europe. PMID- 10849071 TI - Sexual isolation of genetically differentiated sympatric populations of Drosophila melanogaster in Brazzaville, Congo: the first step towards speciation? AB - Two sympatric populations of Drosophila melanogaster were collected in the Brazzaville area in Congo, one from the suburban countryside and the other from a brewery located in the city. They were compared for several genetically determined traits including morphology, allozymes, microsatellites, cuticular hydrocarbons, and sexual behaviour. The two populations were similar to other African populations for morphological traits, but differed significantly from each other for all other characters. The countryside population resembled other African populations, whereas the urban population was consistently similar to European populations. Mating choice experiments showed incipient reproductive separation between the populations. In agreement with the hypothesis that D. melanogaster originated in Africa and spread to the rest of the world by invading human-modified habitats, we suggest that man-adapted fruit fly populations have returned 'back to Africa', and remained partially isolated from older native stocks. PMID- 10849072 TI - Genetic diversity of barley landrace accessions (Hordeum vulgare ssp. vulgare) conserved for different lengths of time in ex situ gene banks. AB - Large numbers of crop plant accessions from all over the world have been amassed in gene banks to secure a gene pool for future breeding programmes. Maintenance of accessions held as seed samples in cold stores involves frequent rejuvenation cycles to ensure the viability of seeds. The practice of rejuvenation by multiplication of a sample of each accession in small field plots has the potential to create population bottlenecks, leading to loss of genetic diversity and changes in gene frequencies every rejuvenation cycle. In order to determine whether these undesirable effects occur, genetic diversity levels were assessed for morphological and isozyme markers within gene bank accessions of two barley landraces from Syria that had been stored for 10, 40 and 72 years. These were compared with genetic diversity levels for the same markers in barley landraces collected recently at locations in Syria where they are still under cultivation. Average gene diversity (H), alleles per locus (A) and percentage polymorphic loci (P (0.01)) all showed very significant declines with length of time in storage, and genetic differentiation FST among accessions increased over time. If the observed differences in genetic diversity are caused by genetic drift in gene bank accessions rejuvenated every 5.3 years, it was estimated that the effective population size Ne of rejuvenation populations over their period in storage was only 4.7. Implications for gene bank management are discussed. PMID- 10849073 TI - The production and characterization of recombination between chromosome 3N of Aegilops uniaristata and chromosome 3A of wheat. AB - Six wheat lines with recombination between Aegilops uniaristata chromosome 3N and wheat chromosome 3A were produced. These were characterized in terms of exchange points by RFLP analysis. Chromosome 3N carries an undesirable brittle rachis gene and three of the recombinant lines had lost this character. The results also support previously published evidence of a pericentric inversion in chromosome 3N relative to the wheat homoeologous group 3 chromosomes. PMID- 10849074 TI - Estimation of spontaneous genome-wide mutation rate parameters: whither beneficial mutations? AB - Empirical estimates of genome-wide mutation rates and of the distribution of mutational effects are needed to illuminate various topics ranging from evolutionary biology to conservation. Methods for inferring genome-wide mutation parameters are presented, and results stemming from these studies are reviewed. It is argued that, although most if not all mutations detected in mutation accumulation experiments are deleterious, the question of the rate of favourable mutations (and their effects) is still a matter for debate. PMID- 10849075 TI - Evolution and maintenance of stigma-height dimorphism in Narcissus. I. Floral variation and style-morph ratios. AB - An unusual stylar dimorphism occurs in Narcissus, a plant genus of insect pollinated Mediterranean geophytes. To determine the characteristics of the sexual polymorphism, we investigated floral variation in 46 populations of N. assoanus (section Jonquillae) and 21 populations of N. dubius (section Tazettae) in SW France. Flowers possess two stamen levels in each morph that occupy slightly different positions within the floral tube. In long-styled plants (L morph), the stigma is located within or slightly above the upper-level stamens, whereas in short-styled plants (S-morph) the stigma is placed well below the lower-level stamens. The stigma-height dimorphism is distinct from heterostyly because the reciprocity of stigma and anther positions in the two style morphs is only weakly developed and there are no differences between the style morphs in pollen size or production. In both species, mean stigma-anther separation is much greater in the S-morph than the L-morph. In N. assoanus, population style-morph ratios vary from isoplethy (1L:1S) to L-biased, whereas in N. dubius they are usually strongly L-biased or occasionally contain only the L-morph. Populations fixed for the S-morph, or with S-biased morph ratios, were not observed. In N. assoanus, style-morph ratios were associated with population size: large continuous populations always exhibited 1:1 morph ratios, whereas smaller, fragmented populations were often L-biased. This pattern was not evident in N. dubius. We argue that biased style-morph ratios largely result from morph specific differences in assortative mating. PMID- 10849076 TI - Evolution and maintenance of stigma-height dimorphism in Narcissus. II. Fitness comparisons between style morphs. AB - Populations of the insect-pollinated geophytes Narcissus assoanus and N. dubius (Amaryllidaceae) are commonly dimorphic for stigma height. An extensive survey of populations of the two species in SW France revealed a wide range of style-morph frequencies, particularly populations with significantly more long-styled than short-styled plants. Here we employ experimental and theoretical approaches to investigate potential selective mechanisms governing the variation in style-morph frequencies. Controlled pollination of both species demonstrated that N. assoanus is moderately self-sterile whereas N. dubius is highly self-compatible. Both intra- and intermorph crosses of N. assoanus were equally fertile, indicating that the species does not exhibit heteromorphic incompatibility. Estimates of female fertility (fruit- and seed-set) and multilocus estimates of outcrossing using allozyme markers provided no evidence of morph-specific differences in maternal components of reproductive success in natural populations of the two species. This result suggested that differences between the morphs in male fertility may be largely responsible for the observed morph-ratio variation. To investigate this hypothesis we developed a mating model that incorporates the genetics of stigma-height dimorphism and contrasting rates of assortative and disassortative mating in the style morphs. Simulation results demonstrated that stigma-height dimorphism will always be maintained when levels of disassortative mating are greater than assortative mating, and that the observed L-biased populations in Narcissus spp. probably result from greater levels of assortative mating in this morph in comparison with the S-morph. PMID- 10849077 TI - Maximum likelihood analysis of quantitative trait loci under selective genotyping. AB - Selective genotyping is a cost-saving strategy in mapping quantitative trait loci (QTLs). When the proportion of individuals selected for genotyping is low, the majority of the individuals are not genotyped, but their phenotypic values, if available, are still included in the data analysis to correct the bias in parameter estimation. These ungenotyped individuals do not contribute much information about linkage analysis and their inclusion can substantially increase the computational burden. For multiple trait analysis, ungenotyped individuals may not have a full array of phenotypic measurements. In this case, unbiased estimation of QTL effects using current methods seems to be impossible. In this study, we develop a maximum likelihood method of QTL mapping under selective genotyping using only the phenotypic values of genotyped individuals. Compared with the full data analysis (using all phenotypic values), the proposed method performs well. We derive an expectation-maximization (EM) algorithm that appears to be a simple modification of the existing EM algorithm for standard interval mapping. The new method can be readily incorporated into a standard QTL mapping software, e.g. MAPMAKER. A general recommendation is that whenever full data analysis is possible, the full maximum likelihood analysis should be performed. If it is impossible to analyse the full data, e.g. sample sizes are too large, phenotypic values of ungenotyped individuals are missing or composite interval mapping is to be performed, the proposed method can be applied. PMID- 10849078 TI - Multiple nuclear pseudogenes of mitochondrial cytochrome b in Ctenomys (Caviomorpha, rodentia) with either great similarity to or high divergence from the true mitochondrial sequence. AB - A fragment of the mitochondrial cytochrome b gene was studied in 13 species of the South American fossorial rodent Ctenomys using PCR with 'universal' primers and DNA sequencing after cloning. Five different groups of sequences were found, one of which corresponds to the functional mitochondrial gene (mt). The other four groups (A, B, C and D) were believed to be nuclear pseudogenes. Sequences A C were highly divergent from the mt sequences and included substitutions, deletions and insertions such that they could not possibly have coded a functional protein. They all shared a common insertion between positions 15055 and 15056 suggestive of a common origin, although the A, B and C sequences otherwise differed greatly from each other. The D sequences also could not have been functional on the basis of nucleotide sequence, but the differences with the mt sequences were far more subtle and in a more limited study the D sequences could easily have been classified as a true mtDNA sequence. It is suggested that there were two transfers of the cytochrome b gene from the mitochondrion to the nucleus; the first leading to sequences A-C and the second to the D sequence. Subsequent to transfer, a sequence of duplications within the nucleus appears to have generated the full range of pseudogenes that are observed. This study adds to other recent observations suggesting the frequent transfer of mtDNA sequences to the nucleus and reinforces the necessity of great care in interpreting PCR generated sequences, particularly those produced with universal primers. There are now data from several species of mammals and birds relating to PCR-generated nuclear copies of cytochrome b, which we review. PMID- 10849079 TI - Multivariate analysis of male mating success on morphometric traits and chromosome dosage in the grasshopper Sinipta dalmani. AB - Sinipta dalmani is a South American grasshopper that is chromosomally polymorphic for a pericentric inversion in the M4 pair. The inversion affects morphology, showing a negative average effect on body size. In the present work, male mating success was analysed in competition cages and possible correlations examined between this fitness component, body size and inversion polymorphism. The outcome of the study revealed that mated males were larger and had lower inversion dosage than unmated ones. The analysis of standardized selection differentials (S') demonstrated that directional selection operated on chromosome dosage and 3rd tibia, 3rd femur, thorax and tegmina lengths. The analysis of karyotype frequencies showed that mated males had a higher frequency of the standard homozygote karyotype. The analysis of selection gradient (beta') and the transformed logistic regression (alpha) showed an important effect of selection on 3rd tibia length. Identical conclusions were obtained when linear and logistic multiple regression were performed on the first three components of PCA analyses. Our results indicate that differences in some body size-related traits are determined by the karyotype, and that of these length of 3rd tibia provides the greatest contribution to variation in fitness. The selective effects detected on chromosome dosage and the other morphological traits may be considered as indirect ones caused by correlated effects. PMID- 10849080 TI - RAPD of controlled crosses and clones from the field suggests that hybrids are rare in the Salix alba-Salix fragilis complex. AB - The polyploid Salix alba-Salix fragilis hybrid complex is rather difficult to study when using only morphological characters. Most of the features have a low diagnostic value for unambiguously identifying the hybrids, introgression patterns and population structures, though morphological traits have proved to be useful in making a hybrid index. Morphology and molecular variation from RAPDs were investigated in several case studies on willows from Belgium. A thorough screening of full-sib progenies of interspecific controlled crosses was made to select homologous amplification products. The selected amplified products proved to be useful in a principal coordinate analysis for the estimation of variability of hybrid progenies. On the basis of genetic similarities and ordination analysis, a method for the identification of clones in the field was established using presumed pure species and presumed introgressants. The chosen reference clones were checked against additional European samples of putative pure species to ensure the reliability of the method beyond a regional scale. The RAPDs suggested that both species have kept their gene pools well separated and that hybridization actually does not seem to be a dominating process. The observation that molecular markers do not always follow the morphological traits or allozyme data is discussed. PMID- 10849081 TI - Genetic diversity of Chilean and Brazilian alstroemeria species assessed by AFLP analysis. AB - One to three accessions of 22 Alstroemeria species, an interspecific hybrid (A. aurea x A. inodora), and single accessions of Bomarea salsilla and Leontochir ovallei were evaluated using the AFLP-marker technique to estimate the genetic diversity within the genus Alstroemeria. Three primer combinations generated 716 markers and discriminated all Alstroemeria species. The dendrogram inferred from the AFLP fingerprints supported the conjecture of the generic separation of the Chilean and Brazilian Alstroemeria species. The principal co-ordinate plot showed the separate allocation of the A. ligtu group and the allocation of A. aurea, which has a wide range of geographical distribution and genetic variation, in the middle of other Alstroemeria species. The genetic distances, based on AFLP markers, determined the genomic contribution of the parents to the interspecific hybrid. PMID- 10849082 TI - A comparison of spermatogenesis in homozygotes, simple Robertsonian heterozygotes and complex heterozygotes of the common shrew (Sorex araneus L.). AB - Spermatogenesis was studied in 56 shrews (Sorex araneus L.) from two chromosomal hybrid zones in Poland. The hybrid zones were formed between chromosome races that differed in Robertsonian metacentrics. Shrews were compared in four classes: homozygotes, simple Robertsonian heterozygotes, complex heterozygotes forming four-element rings in meiosis I, and complex heterozygotes forming four- or five element chains. There was a significant effect of karyotype on the level of germ cell death and chain-forming complex heterozygotes suffered the greatest germ cell loss. However, the estimated level of germ-cell death is probably insufficient to influence the fertility of these males. PMID- 10849083 TI - Temperatures in the cothill habitat of Panaxia (Callimorpha) dominula L. (the scarlet tiger moth). AB - Temperatures were monitored in precisely the places in Cothill Fen in which larvae of Panaxia dominula pupate. At the critical time in June, the temperatures in the litter layer are diurnally often much higher than those used in previous constant temperature experiments. Moreover, the daily mean temperature in the shaded litter layer is also higher than the shaded air temperature at 0.4 m, both temperatures being higher than the estimate of 12 degrees C used as baseline by others. The effects of fluctuating temperatures in the natural habitat on the phenotypic expression of wing patterns of the moth appear to be minimal and do not parallel the results of laboratory experiments. PMID- 10849084 TI - Quantitative trait loci and gene interaction: the quantitative genetics of metapopulations. AB - Genetic population differentiation is typically viewed as differentiation of population means. However, several theories of evolution and speciation postulate that populations differentiate not only with respect to the population means, but also with respect to the effects of alleles within these populations. I develop herein a measure of population differentiation for the 'local average effects' of alleles, where local average effect is defined as the average effect of an allele in a deme measured as a deviation from the metapopulation mean. The differentiation for local average effects has two components, a component attributable to the population mean and a residual component that is attributable to changes in the local average effects independent of the population mean. The variance in local average effects attributable to the population mean is measured as the variance in the mean local average effect of all alleles. The variance in the residual local average effects is measured as the difference between the variance local average effects of individual alleles and the variance in the mean local average effects of all alleles. Differentiation for population means and differentiation for residual local average effects need not be related. I show that when there is only additive gene action, populations can differentiate for population means, but not for residual local average effects. However, if there is gene interaction then populations can also differentiate for local average effects of alleles. The consequence of this differentiation is that the local average effects of alleles change relative to each other such that an allele that is favoured by selection in one population may be removed by selection in other populations. I discuss the evolutionary consequences of differentiation for local average effects, and the interpretation of QTL data in light of this model. PMID- 10849085 TI - Dieldrin resistance in Lucilia cuprina (the australian sheep blowfly): chance, selection and response. AB - Discrete-generation population cages of Lucilia cuprina were initiated with dieldrin-resistant allele (Rdl ) frequencies of 1 or 5% and maintained for 17 generations on media with concentrations of dieldrin in the range 0-0.006% (w/v). The probability of the initial establishment of the Rdl allele in a population was consistently greater at the 5% frequency and dependent on the concentration of dieldrin in the medium for both starting frequencies. Once the resistant allele was established responses to selection were concentration-dependent. It was concluded that in the absence of dieldrin the susceptible allele was selectively favoured, at 0. 00005% (w/v) concentration selection and random genetic drift influenced changes in allele frequency and at concentrations above this the Rdl allele was at a selective advantage. Fixation of Rdl occurred at the higher concentrations. The influence of random genetic drift and selection on the genetic response during the evolution of insecticide resistance is discussed. PMID- 10849086 TI - Multiple causes of male-killing in a single sample of the two-spot ladybird, Adalia bipunctata (Coleoptera: coccinellidae) from Moscow. AB - Thirty-six matrilines from a single Muscovite sample of Adalia bipunctata were assayed, using appropriate primers, for presence of the four male-killing symbionts known to infect this species of ladybird. All four, a Rickettsia, a Spiroplasma and two different strains of Wolbachia, were found to be present. Vertical transmission efficiencies were assessed from F1 and F2 families from each of the matrilines, and were found to differ significantly between symbionts. Potential explanations of the presence of four different male-killing symbionts within a single population, are considered. PMID- 10849087 TI - Genetic differentiation among northern and southern populations of the moor frog Rana arvalis Nilsson in central Europe. AB - Starch gel electrophoresis and morphometric characters were used to assess the geographical variation between 14 populations of the moor frog, Rana arvalis, from northern and southern areas in Central Europe. Six of the 13 screened allozyme loci were polymorphic (95% criterion). No fixed differences in allele composition between the two regions were found. Some of the alleles were region specific. Genetic variability as measured by expected heterozygosity (He) and number of alleles per locus was significantly lower in the southern samples than in northern ones (He=0.104 and He=0.156, alleles/locus=1.6 and 1.8 respectively). This is interpreted as a consequence of the different past history of these two groups during the Pleistocene. Population subdivision, as measured by FST, was substantial (0.124 and 0.078 for the southern and northern group, respectively); 59.9% of the between-locality variation is attributed to this division into two geographical groups. Isolation-by-distance was detected by significant negative correlation between the estimate of gene flow (log M) and log(geographical distance) only for the southern population groups. This indicates that the northern populations have recently recolonized their contemporary distribution area. The mean genetic distance between the northern and southern group of populations was DN=0.062. Despite the relatively low genetic distance between them, the two population groups form two distinct clusters in the maximum likelihood (ML) tree. Discriminant analysis on 11 size adjusted body measurements showed considerable overlap between populations from different geographical areas. An isolated Romanian Reci population which genetically belongs to the southern group of populations was morphologically situated in an intermediate position between northern and other southern populations. PMID- 10849088 TI - Systematic review and meta-analysis in anatomic pathology. AB - Systematic reviews and meta-analyses are techniques of data retrieval and analysis which complement traditional narrative reviews. They are widely used in clinical medicine and are finding an increasing role in anatomical pathology. Performing high quality systematic review and meta-analysis requires the accumulation of large numbers of cases from well planned and executed studies and is facilitated if data is presented in a standardized manner. Techniques which allow data from individual patients included in a variety of different studies are now being developed indicating that in future research papers may require a more detailed description of results than in the past. This need may be met by posting anonymised data on the Internet. Systematic reviews and meta-analyses are never complete since data are continually contributed and analyses constantly updated. As with any research paper, the results of these techniques require careful evaluation and the role of the expert reviewer is enlarged by these methodologies. PMID- 10849089 TI - Arteriolitis in renal transplant biopsies is associated with poor graft outcome. AB - AIMS: The Banff 1997 classification of renal allograft pathology identifies arteriolitis as a finding of uncertain significance. We sought to improve our understanding of arteriolitis by correlating its occurrence with histopathological and clinical parameters. METHODS AND RESULTS: Twenty allograft kidney biopsies from 19 patients, showing arteriolitis, were identified. Arterioles were defined as small vessels with: (1) wall thickness of 1-3 myocytes; (2) diameter less than one-third of an adjacent glomerulus; and (3) discontinuous or absent elastica. Arteriolitis was defined as mural infiltration by lymphocytes. Other histological findings were categorized according to the Banff 1997 working formulation. Ten biopsies (50%) showed type IIA rejection, seven (35%) showed type I rejection, and three (15%) showed borderline change. Two patients with borderline change had acute rejection in the next biopsy. None of the seven patients with type I rejection had previous or subsequent type II rejection on biopsy. A total 11/20 biopsies (10/19 patients) showing arteriolitis had type IIA rejection in the index or next biopsy. On follow-up, graft loss due to rejection occurred in 5/19 (26%) patients (median 126 days); all had shown type IIA rejection on a previous biopsy. Chronic allograft nephropathy developed in a further 4/19 (21%) patients (median 157 days), of whom three had shown only type I rejection on biopsy. CONCLUSION: Arteriolitis is associated with acute rejection, often type II rejection, and is associated with poor graft outcome. Other causes of arteriolitis were not encountered in this series. PMID- 10849090 TI - Warthin-like tumour of the thyroid gland: RET/PTC expression indicates it is a variant of papillary carcinoma. AB - AIMS: Three cases with features of so-called 'Warthin-like tumour' of the thyroid (WaLTT) are described, in order to evaluate its relationship with papillary carcinoma (PC). METHODS AND RESULTS: We performed an histological and immunohistochemical study with emphasis on RET/PTC expression. The most striking features are represented by marked lymphocytic infiltration in the stalks of papillae and by oxyphilic metaplasia of epithelium, resembling Warthin tumour of the salivary gland. In all cases, we found nuclear features reminiscent of PC. The neoplastic cells were strongly positive for Leu M1 and epithelial membrane antigen (EMA), less for thyroglobulin and negative for calcitonin. The lymphocytic infiltrate was composed of a mixed population of B and T-cells with sparse S100-positive Langerhans cells. An interesting finding was the strong positivity with the antibody against RET/PTC. CONCLUSION: All clinicopathological data along with the presence of the extensive lymphocytic infiltrate could imply a more favourable prognosis. The expression of RET/PTC fusion gene adds support to the hypothesis that this tumour is a variant of PC, probably related to the oncocytic variant of PC. PMID- 10849091 TI - Validation of immunolocalization of the urokinase receptor expression in ductal carcinoma in situ of the breast: comparison with detection by non-isotopic in situ hybridization. AB - AIMS: Ductal carcinoma in situ (DCIS) is a pre-invasive form of mammary carcinoma with no microscopic evidence of cancer cell invasion through the basement membrane. However, for initiation of invasion, tumour cells have to acquire and focus proteolytic activity on to the cell surface in order to infiltrate the surrounding extracellular matrix. The receptor (uPA-R or CD87) for the serine protease urokinase-type plasminogen activator (uPA) plays a central role in invasion and metastasis. This study was performed to determine and localize m-RNA and protein of uPA-R in ductal carcinoma in situ of the breast. METHODS AND RESULTS: We analysed uPA-R mRNA and protein expression by in-situ hybridization and immunohistochemistry, respectively, in 50 formalin-fixed, paraffin-embedded specimens of DCIS. Three different antibodies were used to stain cell-associated uPA-R; chicken polyclonal antibody (pAb) HU277 and monoclonal antibodies (mAb) IID7 and 3936. In all cases, myoepithelial and stromal cells reacted with either antibody. Especially, reaction of macrophage-like cells with mAb 3936 resulted in a well-marked and bright staining. Applying mAb IID7, in 46 of the 50 breast specimens tumour cells showed a positive immunoreaction. Likewise pAb HU277 stained tumour cells in 40 of the 50 cases, whereas mAb 3936 reacted with only 24 of the 50 tissue sections. Endothelial cells were marked by both mAb IID7 and pAb HU277 (46/50 and 35/50, respectively); mAb 3936 did not label at all. All of the cell types stained by mAb IID7 and pAb HU277 also displayed reactivity with uPA-R mRNA-specific antisense oligonucleotides in in-situ hybridization. CONCLUSIONS: Our results reveal the presence of the tumour invasion-related receptor for the protease uPA not only in invasive ductal breast carcinoma but also in different types of DCIS. PMID- 10849092 TI - Pleomorphic carcinoma of the breast: clinicopathological analysis of 26 cases of an unusual high-grade phenotype of ductal carcinoma. AB - AIMS: Pleomorphic carcinoma is a poorly described entity whose phenotype is not well recognized as within the morphological spectrum of breast carcinoma. The purpose of this report is to describe the clinicopathological features of this tumour, and to promote its recognition as an unusual high-grade morphological variant of mammary ductal carcinoma. METHODS AND RESULTS: Histological slides of breast carcinomas (N = 64) coded between 1978 and 1995 as having pleomorphic or anaplastic features were reviewed. Pleomorphic carcinoma (N = 26) was diagnosed when > or = 50% of the tumour manifested a pleomorphic cell population (> sixfold variation in nuclear size). Tumours of lobular origin were excluded. All neoplasms occurred in women with a mean age of 53 years. Patients underwent biopsy and/or mastectomy (n = 24) or lumpectomy (n = 2). The tumours' mean size was 54 mm. All were high-grade neoplasms. The pleomorphic cell population comprised 50-100% of the tumour; 31% had a prominent spindled morphology. Fifty eight per cent of the tumours were initially misclassified by referring pathologists as sarcomas or carcinomas, possibly metastatic. Adjacent DCIS or a transition to classic ductal carcinoma was present in 73%. Five (19%) patients were stage I and three (12%) had stage IV disease. Axillary dissections yielded > or = 3 (mean 7.2) positive lymph nodes in 52%. Most (68%) tumours were aneuploid; a high S-phase (> 10%) was present in 63%. All neoplasms were ER negative and all but one were PR negative. p53 expression was present in 71%; none expressed bcl 2. c-erbB-2 was detected in four (19%) node-positive and in 0 (0%) node-negative cases (P = 0.01). Of 16 patients with follow-up, 6 (38%) were disease-free (mean, 74 months), four (25%) alive with disease (mean, 33 months) and six (38%) dead of disease at a mean of 22 months. CONCLUSIONS: Pleomorphic carcinoma is a prognostically unfavourable lesion and represents the extreme end of the morphological spectrum of grade III infiltrating ductal carcinoma. PMID- 10849094 TI - Immunoreactivity for cadherins, HGF/SF, met, and erbB-2 in pleural malignant mesotheliomas. AB - AIMS: To assess the immunoreactivity of malignant mesotheliomas for N- and E cadherins, hepatocyte growth factor/scatter factor (HGF/SF) and the tyrosine kinase receptors, met and erbB-2. METHODS AND RESULTS: Pleural malignant mesotheliomas were stained using a standard indirect immunoperoxidase method applied to paraffin sections. Malignant mesotheliomas were immunoreactive for N cadherin (26/29; 90%), met (29/29; 100%) and erbB-2 (28/29; 97%). Focal immunoreactivity was present for E-cadherin in epithelioid or mixed tumours (14/25; 56%), and for HGF/SF (9/24; 38%). CONCLUSIONS: Expression of N-cadherin supports the diagnosis of malignant mesothelioma and use of appropriate antibodies would be a useful addition to a diagnostic antibody panel. Focal staining for E-cadherin does not exclude mesothelioma. Signalling pathways mediated via met and erbB-2 may play a role in the growth and spread of malignant mesotheliomas. PMID- 10849093 TI - Expression of steroid hormone receptors, their regulated proteins, and bcl-2 protein in myofibroblastoma of the breast. AB - AIMS: To investigate the possible role of steroid hormones in the pathogenesis of myofibroblastoma (MFB) of the breast, we analysed the immunohistochemical expression of oestrogen, progesterone, androgen receptors, their regulated proteins and bcl-2 protein in a series of this rare tumour. METHODS AND RESULTS: Paraffin-embedded sections from seven cases of MFB of the breast (five male; two female) were immunohistochemically tested for the expression of oestrogen receptor (ER), progesterone receptor (PR), androgen receptor (AR), oestrogen regulated pS2 protein, androgen-regulated prostate-specific antigen (PSA) and bcl 2 protein. Rare cases of benign spindle cell tumours or tumour-like lesions of the breast (primitive fibromatosis, inflammatory pseudotumour, muscular hamartoma) which enter into the differential diagnosis with MFB, were also investigated and compared with MFB. All cases of MFB showed a diffuse (70-90% of neoplastic cells) and strong nuclear labelling with ER and PR, whereas AR was expressed only in three cases (two men and one woman) in about 60-70% of cells. Conversely, no immunostaining was detected for the pS2 protein and PSA. bcl-2 protein immunoreactivity was found in all cases of MFB, although with a variable degree of expression. No expression for steroid hormone receptors, their regulated-proteins and bcl-2 protein was observed in the rare benign spindle cell lesions of the breast included in this study. CONCLUSION: The in-situ detection of ER, PR and AR suggests that steroid hormones and their receptors are implicated in the pathogenesis of breast MFB. The consistent demonstration of bcl 2 protein, associated with a positive ER/PR status, provides evidence that bcl-2 may be an oestrogen-regulated protein also in MFB and that probably plays a role in the tumorigenesis. Finally, we postulate that the ER/PR and bcl-2 positive immunoprofile of MFB of the breast, in contrast to the negative profile of other rare primitive benign spindle cell lesions of the breast herein studied, might be exploited as an ancillary diagnostic aid in differential diagnosis of doubtful cases. PMID- 10849095 TI - Combined dermatofibroma: co-existence of two or more variant patterns in a single lesion. AB - AIMS: Based on a series of 25 cases, we define and characterize combined dermatofibroma, a tumour comprising two or more variant patterns of dermatofibroma in a single lesion. METHOD AND RESULTS: Dermatofibroma may present with a wide variety of architectural, cellular or stromal peculiarities. Architectural peculiarities include deep penetration, atrophy, collarette formation, fascicular to plexiform architecture, massive haemorrhage, prominent haemangiopericytoma-like vascularity and palisading; cellular peculiarities the presence of epithelioid cells, clear cells, granular cells, prominent myofibroblastic differentiation and atypical giant cells ('monster cells'); or stromal peculiarities such as prominent sclerosis, mucin, haemosiderin and cholesterotic deposits. In combined dermatofibromas two or more of these features are seen in complex or inhomogenous combination such as the silhouette of a deep penetrating dermatofibroma with an 'ordinary' storiform pattern in the upper and granular cell differentiation in the lower part of the lesion; or a dermatofibroma with ordinary features in the upper, prominent sclerosis in the middle and clear cells in the lower portion of the lesion; or the characteristic epidermal collarette and cells of epithelial cell histiocytoma with a plexiform ('neurothekeoma-like') architecture surrounded by a myxoid stroma with spindle shaped to stellate cells. Clinically, these lesions preferentially occur on the lower extremities of young to middle-aged females, frequently with the diagnosis of a fibrohistiocytic lesion. Apart from one recurrence follow-up was uneventful in all other cases. Immunohistochemically, lesions are consistently positive with KiM1p, variably positive for factor XIIIa, smooth muscle specific actin and with KP1 (CD68), NK1C3 and E9. CONCLUSION: Recognition of combined dermatofibroma allows the histopathologist to apply a confident benign label to unusual lesions which might otherwise elude diagnosis, or tempt description of 'new' entities and to avoid a misdiagnosis of malignancy. PMID- 10849096 TI - Merkel cell tumour with leiomyosarcomatous differentiation. AB - AIMS: To report and confirm the identity of tumour cells showing leiomyosarcomatous differentiation in a regional lymph node containing metastatic Merkel cell tumour. METHODS AND RESULTS: A 79-year-old woman was found to have metastatic Merkel cell tumour within an axillary lymph node 4 months after excision of the primary tumour from the forearm. The lymph node was effaced by tumour identical to that of the primary tumour but there was an additional focus of loose spindle cells. Immunocytochemical staining showed the coexpression of cytokeratin 20, neurofilament and desmin in these sarcomatous cells. CONCLUSION: This, to the best of our knowledge, is the first report of Merkel cell carcinoma showing sarcomatous differentiation, and adds to the expanding morphological spectrum of malignant biphasic tumours. PMID- 10849097 TI - All infiltrating T-lymphocytes in Hodgkin's disease express immunohistochemically detectable T-cell receptor zeta-chains in situ. AB - AIM: We studied the expression of TCR zeta-chain on tumour-infiltrating lymphocytes in EBV-positive and EBV-negative cases of Hodgkin's disease (HD), to assess whether downregulation of TCR zeta-chain on tumour-infiltrating lymphocytes might be a mechanism for immune escape of the neoplastic cells. METHODS AND RESULTS: By immunohistochemistry we investigated tissue of 27 cases of primary HD, both paraffin embedded and frozen, for the presence of T-cell receptor complex zeta-chain and other T-cell markers on the reactive cells. Strong membranous staining of TCR zeta-chain was present in all cases in frozen tissue. In contrast, in paraffin-embedded material substantial loss of TCR zeta chain was detected in old (> 6 years) tissues. However, no differences in either the number of positive cells or their staining intensity were observed in EBV positive and negative cases of HD as detected in frozen tissue. Storage of paraffin-embedded tissue leads to a rapid and substantial loss of TCR zeta-chain reactivity compared to frozen material of the same HD cases. Staining reactivity of other T-cell markers (CD3, CD4 and CD8) on paraffin-embedded material remained unaffected. Immunofluorescent double-staining confirmed colocalization and coexpression of TCR zeta-chain and CD3. CONCLUSIONS: In frozen biopsies of primary HD TCR zeta-chain was expressed on all reactive CD3-positive cells, both in EBV-positive and EBV-negative cases. This suggests that zeta-chain downregulation is not a likely mechanism whereby neoplastic cells of HD can escape immune surveillance. PMID- 10849098 TI - Relationship between fascicle size and perineurial collagen IV content in diabetic and control human peripheral nerve. AB - AIM: The relationship between perineurial collagen IV content and fascicle size was determined in diabetic and control human peripheral nerve. METHODS AND RESULTS: Age-matched diabetic and control sural nerve samples were immunostained using antibodies to collagen IV. The number of cell layers and the perimeter of the fascicle were measured and the collagen IV content of the perineurium determined. Using this method, a comparison could be made of collagen IV content in the perineuria of fascicles of different size. A positive linear relationship was found between fascicle size and the amount of collagen IV per unit of perineurium. The number of perineurial cell layers and the collagen IV content of the diabetic nerve did not differ from control values. CONCLUSIONS: The linear relationship between fascicle size and perineurial collagen IV content per unit of perineurium underlines the importance of taking fascicle size into account when determining changes in basement membrane components associated with neuropathy. The results indicate that increased deposition of collagen IV is not involved in the early changes in the perineurial cell basement membrane and is thus not the primary factor involved in altered nerve function associated with diabetic neuropathy. PMID- 10849099 TI - What is the significance of muciphages in colorectal biopsies? The significance of muciphages in otherwise normal colorectal biopsies. AB - Muciphages (mucin-containing macrophages), first described in 1966 by Azzopardi & Evans, are a common feature of biopsies of large intestinal mucosa, even in the absence of other abnormalities such as active inflammation or evidence of chronic inflammatory bowel disease. Should they be mentioned in diagnostic reports? Do muciphages reliably indicate previous mucosal disease, now quiescent? In the following articles, Salto-Tellez & Price review what is known about muciphages and conclude that they reflect previous occult and clinically unimportant mucosal damage and that, in an otherwise normal colorectal mucosa, they have no diagnostic significance; and Shepherd draws attention to a wide range of clinically much more significant mucosal infiltrates that could be mistakenly regarded as muciphages and thus overlooked. PMID- 10849100 TI - What is the significance of muciphages in colorectal biopsies? Muciphages and other mucosal accumulations in the colorectal mucosa. AB - Muciphages (mucin-containing macrophages), first described in 1966 by Azzopardi & Evans, are a common feature of biopsies of large intestinal mucosa, even in the absence of other abnormalities such as active inflammation or evidence of chronic inflammatory bowel disease. Should they be mentioned in diagnostic reports? Do muciphages reliably indicate previous mucosal disease, now quiescent? In the following articles, Salto-Tellez & Price review what is known about muciphages and conclude that they reflect previous occult and clinically unimportant mucosal damage and that, in an otherwise normal colorectal mucosa, they have no diagnostic significance; and Shepherd draws attention to a wide range of clinically much more significant mucosal infiltrates that could be mistakenly regarded as muciphages and thus overlooked. PMID- 10849101 TI - Primary cutaneous carcinoid tumour. PMID- 10849102 TI - Mucinous cystadenocarcinoma of the breast showing sulfomucin production. PMID- 10849103 TI - Solitary Langerhans cell histiocytosis in association with primary biliary cirrhosis. PMID- 10849104 TI - Periurethral glomangiomyoma in women: case report and review of the literature. PMID- 10849105 TI - Late malignant transformation of biopsy proven benign synovial chondromatosis: an unexpected pitfall. PMID- 10849106 TI - An overview of cancer immunotherapy. AB - The survival of patients with cancer has improved steadily but incrementally over the last century, with the advent of effective anticancer treatments such as chemotherapy and radiotherapy. However, the majority of patients with metastatic disease will not be cured by these measures and will eventually die of their disease. New and more effective methods of treating these patients are required urgently. The immune system is a potent force for rejecting transplanted organs or microbial pathogens, but effective spontaneous immunologically induced cancer remissions are very rare. In recent years, much has been discovered about the mechanisms by which the immune system recognizes and responds to cancers. The specific antigens involved have now been defined in many cases. Improved adjuvants are available. Means by which cancer cells overcome immunological attack can be exploited and overcome. Most importantly, the immunological control mechanisms responsible for initiating and maintaining an effective immune response are now much better understood. It is now possible to manipulate immunological effector cells or antigen-presenting cells ex vivo in order to induce an effective antitumour response. At the same time, it is possible to recruit other aspects of the immune system, both specific (e.g. antibody responses) and innate (natural killer cells and granulocytes). PMID- 10849107 TI - Improved FACS analysis confirms generation of immature dendritic cells in long term stromal-dependent spleen cultures. AB - Cells produced in spleen stroma-dependent long-term cultures (LTC) have now been clearly defined as dendritic cells (DC). Characterization of cells by antibody staining and FACS analysis has only been possible using a procedure to quench the high autofluorescence of DC produced in LTC (LTC-DC). The population of large cells produced by the established LTC-X1 culture are homogeneously positive for a number of cell-surface markers expressed by DC. These include CD11c, CD11b, Dec 205, Fc receptor and CD86. They also express markers detectable with the F4/80 and 33D1 antibodies. Cells produced in LTC do not uniformly express the MHC II marker, consistent with an immature DC phenotype. Most cells are weakly positive for MHC II with a small subset of highly positive cells. The quenching method involves staining cells with crystal violet dye, which is taken up within the cell. The importance of optimizimg fluorescent antibody staining assays for delineating DC subsets is indicated and the LTC system is established as a valuable and continuous source of DC precursors. PMID- 10849108 TI - Phenotypic and functional characteristics of macrophage-like cells differentiated in pro-inflammatory cytokine-containing cultures. AB - Previous studies have demonstrated an infiltration of monocytes and increased levels of IL-1beta and TNF-alpha in some chronic inflammatory tissues. Interleukin-1beta and TNF-alpha are capable of protecting monocytes from spontaneous apoptosis and thus maintain their viability in vitro. To study the possible effects of these cytokines on the differentiation and function of recruited monocytes, a model has been developed in which monocytes isolated from human peripheral blood were differentiated into macrophages in serum in the presence or absence of IL-1beta or TNF-alpha. Monocytes cultured with IL-1beta and TNF-alpha underwent substantial changes in morphology, similar to those observed in monocytes undergoing differentiation into macrophages. The cultured cells increased in size and vacuolization and their content of acid phosphates increased 10-fold. Although they exhibited the morphological characteristics of macrophages, monocytes matured in the cytokines differed functionally from those cultured in serum in a lower expression of HLA-DR, lower ability for triggering the proliferation of allogeneic lymphocytes, higher expression of mannose receptor and greater production of superoxide and TNF-alpha. This data suggests that IL-1beta and TNF-alpha direct monocyte differentiation into macrophages with a reduced antigen-presenting and an increased pro-inflammatory factor-releasing phenotype. Elevated levels of IL-1beta and TNF-alpha in the inflammatory tissues may therefore not only prolong the survival of recruited monocytes, but maintain them in an inflammatory state. PMID- 10849109 TI - Regulation of T cell cytokine production by dendritic cells. AB - Previous work has established that the dendritic cells (DC) of mouse spleen regulate the IL-2 production, and hence the extent of proliferation, of the CD8 T cells they activate. It is now reported here that interaction of primary CD8 T cells with splenic CD8alpha- DC induced much higher production of IL-3, IFN-gamma and granulocyte-macrophage colony-stimulating factor (GM-CSF), as well as IL-2, than did interaction with CD8alpha+ splenic DC. Furthermore, the CD8alpha- DC also induced higher levels of IL-2, IL-3 and IL-10 production in primary CD4 T cells, compared with that induced by CD8alpha+ DC. These quantitative differences did not involve qualitative shifts in the type of cytokine produced. Interleukin 4 production remained low in all the primary T cell cultures and restimulation experiments in secondary cultures did not reveal any bias in the cytokine production profile. When exogenous IL-2 was added to the primary cultures to ensure equal proliferation in response to CD8alpha- or CD8alpha+ DC, the higher level of production of IL-3, IFN-gamma and GM-CSF induced by CD8alpha- DC was maintained. Thus, this general control of T cell cytokine production by splenic DC involves factors additional to those that govern activation of T cells into cell cycle. PMID- 10849110 TI - Disparity in the kinetics of onset of hypermutation in immunoglobulin heavy and light chains. AB - The present paper describes a comparative analysis of light chains associated with primary and secondary IgM, as well as with secondary IgG antibodies to fluorescein, undertaken in order to explore the relationship between light chain somatic hypermutation and the isotype switch. The data reveal a disparity in the frequency of somatic hypermutation of secondary IgM heavy versus light chains. Among 20 secondary IgM light chains, a mutation frequency of 1/777 nucleotides was defined. In contrast, our previous analysis of the heavy chains of these molecules had identified a mutation frequency of 1/129. Among 17 IgG-derived light chains, obtained from animals killed at the same time point as those from which the secondary IgM antibodies were obtained, we measured a mutation frequency of 1/77. Finally, analysis of 20 light chains derived from primary IgM antibodies revealed a mutation frequency of only 1/1192 nucleotides. These data demonstrate that, prior to the class switch, light chain mutation occurs at a frequency considerably lower than that measured for the associated heavy chain gene. Six additional apparent mutations in the secondary IgM antibody 95B3 were all shared with a set of IgG antifluorescein antibodies belonging to the Vkappa 34 family. It is suggested that these light chains represent the products of a previously uncharacterized germ line gene. PMID- 10849111 TI - Dexamethasone-induced apoptosis of mouse thymocytes: prevention by native 7alpha hydroxysteroids. AB - Dehydroepiandrosterone (DHEA) has been shown to decrease the dexamethasone (DEX) induced apoptosis of thymocytes and to be one of the native 3beta-hydroxysteroids extensively 7alpha-hydroxylated in thymus. This led us to question whether DHEA or 7alpha-hydroxy-DHEA is responsible for the decrease in DEX-induced apoptosis of thymocytes and whether this property is shared with other native 3beta hydroxysteroids and their 7alpha-hydroxylated metabolites. Treatment of mice with DHEA or 7alpha-hydroxy-DHEA prior to DEX led to a smaller decrease in thymus weight than with DEX alone and to a disappearance of the DEX-induced changes in thymocyte phenotypes. Thymocyte apoptosis induced by DEX treatment was significantly lowered in DHEA- and 7alpha-hydroxy-DHEA-treated thymi, even after 18 h culture with additional 10-6 mol/L DEX. Extensive apoptosis of thymocytes cultured with 10-7 mol/L DEX was brought back to control levels when 10-5 mol/L 7alpha-hydroxy-DHEA or 10-5 mol/L 7alpha-hydroxy-epiandrosterone was added. After use of DHEA and epiandrosterone or pregnenolone, less significant and no significant changes were obtained, respectively. These findings imply that the 7alpha-hydroxylation of 3beta-hydroxysteroids may be a prerequisite for an exquisite regulation of the thymocyte-positive selection driven by the glucocorticoids produced in thymic epithelial cells. PMID- 10849112 TI - Lymphocytes from H2 mice produce lower levels of several cytokines than congenic H2 or H2 mice. AB - Inbred mice of congenic strains that differ only in their H2 haplotype were used to examine the effects of MHC genes on production of cytokines. Spleen and lymph node cells were stimulated with mitogens in vitro, and cytokine protein was assessed by ELISA and/or bioassays. Cells from H2b mice synthesized less IL-3, IL 4, IL-5, TNF and IL-10 (less clearly) than the equivalent cells from H2k or H2d mice. Production of IL-6 by H2b spleen and lymph node cells was lower than that by cells from H2d mice. In addition, lower lymphoproliferative responses were observed in lymph node cultures from H2b mice. These effects were evident in congenic B10 and BALB strains. B10 H2b mice stimulated in vivo with anti-CD3 had lower levels of IFN-gamma and IL-5 protein in their serum compared with equivalent H2k and H2d mice. Because class I- or II-mediated antigen presentation was not required in our model, an immunoregulatory gene in the central MHC is implicated. Preliminary studies of MHC recombinant mice suggested that the gene or genes responsible lie telomeric of IEbeta. Evidence that the H2b haplotype carries an immunoregulatory allele with a small but consistent effect on cytokine production warrants further investigation. PMID- 10849113 TI - Pertussis antibody levels in infants immunized with an acellular pertussis component vaccine, measured using whole-cell pertussis ELISA. AB - A commercially available whole-cell pertussis IgG ELISA was used to test the response of 137 2-month-old infants to immunization with a trivalent acellular pertussis vaccine. The pre-immunization geometric mean (GM) IgG index was 6.96 (95% confidence interval (CI) 5.88-8.04) and the postimmunization GM index was 13.16 (95% CI 12. 20-14.11), P < 0.001. Eighty percent of subjects (110/137) had a significant 1.5-fold increase of pertussis IgG index (97/137, 71%) or a postimmunization IgG index > 10 (93/137, 68%). In single antigen ELISA, 83% showed at least a fourfold increase in pertussis toxin-specific IgG (PT-IgG) and 91% showed an increase in IgG specific for filamentous haemagglutinin (FHA-IgG). Four percent had high pre- immunization antibody levels (index > 20), likely to reflect recent maternal exposure to pertussis. This correlated with a smaller increase in pertussis IgG index. A decline in pertussis IgG index postimmunization occurred in 17/24 infants (71%) whose pre-immunization IgG index was > 10. This postimmunization pertussis IgG index was not significantly different to that of infants with a low pre-immunization index. A similar trend was noted with PT-IgG and FHA-IgG results. The whole-cell ELISA can detect a response to acellular pertussis vaccination in most infants if both antibody index and degree of seroconversion are calculated and at least one criterion is satisfied. PMID- 10849114 TI - Associations between HLA-DQB1 high-risk alleles and type I diabetes do not depend on cytomegalovirus antibody status at onset: a case-parent study conducted in Chile. AB - The purpose of the present study is to ascertain whether the associations between HLA-DQB1*0201 and DQB1*0302 alleles and childhood diabetes depend on the presence of antibodies to human cytomegalovirus (CMV). A study of incident type I diabetes cases and parents was conducted in Santiago, Chile. HLA-DQB1 polymorphisms were determined in 85 case-parent trios (255 subjects), while the detection of CMV was carried out only in the incident cases. As expected, HLA-DQB1 polymorphisms are strongly associated with type I diabetes, with crude odds ratios of 3.7 (95% confidence interval (CI) 1.8-7.7) for the DQB1*0201 allele and 10.3 (95% CI 5.0 21.4) for the DQB1*0302 allele. In the subset of families with CMV+ cases, the odds ratios were estimated as 3.7 (95% CI 1.6-8.6) for the DQB1*0201 allele and 11.1 (95% CI 4.8-25.8) for the DQB1*0302 allele. In families with patients who tested negative for CMV antibodies, the odds ratios were calculated as 3.5 (95% CI 0.7-16.8) for the DQB1*0201 allele, and 8.0 (95% CI 1.8-34.7) for the DQB1*0302 allele. There was no evidence of statistical interaction between CMV antibodies and the DQB1*0201 allele (P value = 0.9) or the DQB1*0302 allele (P value = 0.7). In conclusion, alleles DQB1*0302 and DQB1*0201 do not display distinct associations with type I diabetes depending on the presence of antibodies for CMV. PMID- 10849115 TI - Analysis of the functional characteristics of L-selectin and its expression on normal and CD18-deficient bovine neutrophils. AB - In vivo responsiveness to epinephrine, expression of L-selectin on neutrophils, changes in intracellular calcium ([Ca2+]i), sulfatide-induced superoxide production and tyrosine phosphorylation in neutrophils were evaluated to elucidate the role of L-selectin-associated functions of normal and CD18 deficient bovine neutrophils. The number of neutrophils in peripheral blood was significantly increased (P < 0.05) in four normal calves at 5-20 min after in vivo administration of epinephrine; however, no significant increase of neutrophils was found in three calves with bovine leucocyte adhesion deficiency (BLAD). Expression of L-selectin on neutrophils from three calves with BLAD was 61-77% of that of normal calves. Pretreatment of neutrophils with phorbol myristate acetate caused a marked decrease in the expression of L-selectin on neutrophils from both normal and BLAD calves. The sulfatide-induced sustained phase of [Ca2+]i concentration in neutrophils from calves with BLAD was significantly (P < 0.05) decreased. Following stimulation with aggregated IgG, the transient phase of [Ca2+]i in neutrophils from normal and BLAD calves was increased; however, the sustained phase of [Ca2+]i in BLAD neutrophils was significantly lower (P < 0.05) than that of controls. Sulfatide-induced O2- production and chemiluminescent response in neutrophils from calves with BLAD were 48-51% of those of normal calves and were inhibited by genistein and wortmannin, respectively, in a dose-dependent manner. The amount of tyrosine phosphorylated 100 kDa protein in neutrophils from BLAD calves stimulated with sulfatides was 57% of that of controls. The degree of L-selectin expression on neutrophils was correlated with the intracellular signalling events and the related superoxide production. PMID- 10849116 TI - Interleukin-2-producing helper T lymphocyte precursor frequency and rejection: a longitudinal cellular study after cardiac transplantation. AB - Interleukin-2-producing helper T lymphocyte precursors (HTLp) in the recipient recognize donor alloantigen expressed by the transplanted organ. The frequency of these reactive cells in the peripheral blood was determined and correlated with rejection episodes. Endomyocardial biopsy is generally used to quantify cardiac allograft rejection and guide immunotherapy. While non-invasive techniques have been investigated, none of these has demonstrated sufficient sensitivity or specificity to replace myocardial biopsy. Twelve cardiac transplant recipients were assessed over a 1 year period using limiting dilution analysis, to determine the frequency of HTLp in response to cadaver donor splenocytes. The IL-2 dependent mouse cell line CTLL-2 was used to measure the IL-2 present and the precursor frequency was calculated using maximum likelihood estimation. In the months immediately post transplantation, six of the 12 recipients displayed an association with increases in HTLp frequency, which preceded histologically detectable rejection. The second six recipients had fewer rejection episodes and achieved 'acceptance' of their graft sooner. Once 'acceptance' was achieved, the association between IL-2 HTLp frequency and rejection was no longer apparent. The ability to identify two groups of patients on the basis of IL-2 HTLp frequency clearly highlights the heterogenous nature of the response to the graft and this emphasizes the need to monitor other cytokines, which may influence the functioning of effector T cells. PMID- 10849117 TI - Murine histidine-rich glycoprotein: cloning, characterization and cellular origin. AB - Histidine-rich glycoprotein (HRG) is a plasma protein of vertebrates that has been implicated in the regulation of several important biological functions, including the immune response and blood clotting. In the present study, we have isolated and determined the sequence of the cDNAs for both mouse and rat HRG. The deduced amino acid sequences of mouse and rat HRG are 525 and 510 amino acids, respectively, and they show the same three-domain structure that has been predicted for human HRG, with which they share high amino acid identity. Northern blot analysis indicates that the mouse HRG mRNA is 1.7 kb and is localized specifically to the liver. It has been suggested, somewhat controversially, that some immune cells, such as monocytes and megakaryocytes, also synthesize HRG. Reverse transcriptase-polymerase chain reaction analysis has failed to show any HRG mRNA in immune tissues of the mouse, including the spleen, thymus, lymph node, bone marrow and peripheral blood leucocytes. These data suggest that HRG expression by immune cells is due to the acquisition of plasma HRG derived from the liver. Finally, genomic Southern blot analysis of the mouse HRG gene suggests that it is a single copy gene. PMID- 10849118 TI - Modelling of peripheral lymphocyte migration: system identification approach. AB - This is the first application of the prediction error method (PEM) of system identification to modelling lymphocyte migration through peripheral lymphoid tissue. The PEM was applied to the emergence of labelled lymphocytes from the efferent lymphatic of a lymph node following their intravenous administration. Advantages of PEM included the capacity to calculate the response to a unit impulse stimulus, unavailable to direct observation, and to allow for the return to the node of labelled cells that had already recirculated once. Calculation of the system delay (time between introduction of cells into the blood and their first appearance in lymph) indicated 4.67 +/- 1.05 h for the total lymphocyte population. The peak in efferent lymph occurred at 11.91 +/- 4.68 h, much earlier than previous reports, which were affected by cells that had already recirculated. While 75% of labelled cells had emerged in efferent lymph by 20.77 +/- 5.62 h, 86.38 +/- 29.44 h was required for 100% emergence. The considerable heterogeneity in migratory behaviour is likely to reflect frequency and duration of binding of lymphocytes by dendritic cells in paracortical cord corridors. It is proposed that differences in the speed with which lymphocytes pass along corridors depend on their functional status, in particular whether they are naive or memory cells. PMID- 10849119 TI - Investigation of the site of precursors for IgA-producing cells in the chicken intestine. AB - Bursectomized chicks received lymphocyte single cell suspensions harvested from the bursa of Fabricius (BF), ileal lymphoid aggregate (ILA), caecal tonsils (CT), spleen and peripheral blood. Four days after cell transfer, repopulation of the duodenal and CT lamina propria in age-matched recipient bursectomized chickens with IgA-secreting plasma cells was determined. The results indicate the highest level of reconstitution with cells derived from BF, but substantial numbers of IgA-secreting plasma cells were also observed in a number of birds that received lymphocytes originating from the ILA and CT. PMID- 10849120 TI - The effects of drugs on wound healing--part II. Specific classes of drugs and their effect on healing wounds. PMID- 10849121 TI - Desideratum dermatologicum--cause and control of premenstrual acne flare. PMID- 10849122 TI - Minor cutaneous features of atopic dermatitis in South Korea. AB - BACKGROUND: Minor cutaneous features are important in atopic dermatitis (AD) because they are related to the ethnic or genetic background and to the etiopathogenesis of the disease other than atopic allergy. In addition, they can be used as auxiliary diagnostic criteria in patients with uncertain major features. It is our experience that our AD patients have characteristic features that have not been described previously in the literature. METHODS: AD patients (n = 130) and control subjects (n = 198) were examined for the 32 conventional and seven additional minor features (sandpaper-like skin lesions on elbow/knee/lateral malleolus, hangnail, ventral wrist dermatitis, itchy hyperkeratotic papules on the dorsum of the hands, oily skin, fissured heel, and palmar erythema). The frequency of each feature was compared between AD patients and controls. The diagnostic significance of these minor features was analyzed separately in the childhood and adolescent-adult AD groups, and the age-related changes were documented. RESULTS: The seven additional features were significant for the diagnosis of AD in South Korean patients. Many of the other conventional minor features were also significant. Nine features were of diagnostic importance only in the adolescent-adult AD group, and three features were characteristic only in the childhood AD group. CONCLUSIONS: Our data suggest that ethnic backgrounds influence the phenotype of AD and that an additional seven features need to be examined to confirm the ethnic effect. As the general clinical presentation of AD is dependent upon age, the frequency of minor features varied in the different age groups. PMID- 10849123 TI - Retrospective study of Mycobacterium marinum skin infections. AB - BACKGROUND: Although infection by Mycobacterium marinum is well reported in the literature, there have been few epidemiologic studies. The purpose of this study was to review the epidemiology of patients with cutaneous M. marinum infection over a 3-year period at the National Skin Centre in Singapore. METHODS: Patients with a diagnosis of cutaneous M. marinum infection, confirmed histologically, were collated from computerized data from 1995 to 1997. Thirty-eight patients were diagnosed as having cutaneous M. marinum infection based on history, and clinical and histologic features. RESULTS: Out of the 38 cases of M. marinum infection, there were 30 men and eight women. The age range was 14-85 years (mean: 44.7 years). The duration of disease ranged from 1 to 132 months (mean: 19 months). Thirteen patients (34.2%) had fish rearing as a hobby and four patients (10.5%) had occupational exposure to fish. Twelve patients (31.5%) gave a history of trauma to the disease site. All patients had biopsies of the lesions. All showed infective granulomas/granulomatous inflammation on histology. Acid-fast bacilli were identified in five out of 38 patients (13.2%) and mycobacteria were isolated in one out of 35 patients (2.9%). Nineteen patients received treatment with cotrimoxazole-trimethoprim alone, three with minocycline alone, five with minocycline and cotrimoxazole-trimethoprim, seven with various combinations of drugs, one with excision, and three defaulted treatment. The duration of treatment ranged from 4 to 38 weeks (mean: 14.9 weeks). Twenty-six patients (68.4%) showed clinical improvement, two (5.3%) had no response, and 10 (26.3%) were lost to follow-up. None of the patients worsened with treatment. The follow up period ranged from 1 to 20 months (mean: 6.8 months). CONCLUSIONS: The diagnosis of cutaneous M. marinum infection is mainly clinical, with supporting evidence from histologic features and the response to therapy. Risk factors include a history of trauma and water/fish-related hobbies or occupations. There is a poor yield of positive isolates in our experience; however, empirical treatment usually produces a good clinical response. In future, the polymerase chain reaction (PCR) technique may become more widely available as a rapid, sensitive, and specific means of diagnosis. PMID- 10849124 TI - Prevalence of cutaneous manifestations in 200 patients with eating disorders. AB - BACKGROUND: Eating disorders are increasing and show a variety of symptoms. They mainly include anorexia nervosa (AN), bulimia nervosa (BN), and eating disorders not specified (EDNOS). They predominate in females and represent an important danger, especially in teenagers. In serious cases, they may be life-threatening. Objective To determine the prevalence of cutaneous findings in patients with eating disorders and to compare the results with those found in the literature. METHODS: An observational, transverse, and prospective study was performed. Two hundred patients of recent admission to ALUBA (association that fights against BN and AN), a psychiatric unit for eating disorders, were included: 122 BN; 62 AN; 16 EDNOS. RESULTS: Patients with eating disorders show dermatologic manifestations (alopecia, xerosis, hypertrichosis, caries, nail fragility) that are secondary to starvation. Russell's sign, seen as calluses on the dorsal aspect of the hands, is a consequence of self-induced vomiting and the local trauma of the superior incisors. This sign represents a compensatory behavior to overeating and predominates in the BN group. CONCLUSION: The recognition of dermatologic signs could be of immense value and could lead to the early diagnosis and treatment of these eating disorders. PMID- 10849125 TI - Anxiety, depression, and nature of acne vulgaris in adolescents. AB - BACKGROUND: The reported prevalence of acne in adolescence is variable; improved treatment may have modified its prevalence and severity; acne has been related to psychiatric morbidity for many years. METHODS: Two thousand six hundred and fifty seven high school students were examined, and adolescents with acne were interviewed about the subject of acne vulgaris. The severity of acne was graded using the Global Acne Grading System (GAGS). The Hospital Anxiety and Depression (HAD) scale was evaluated for one of every two subjects with acne (n = 308) and for the same number of sex-matched control subjects (n = 308) to determine the prevalence of depression and anxiety. RESULTS: Six hundred and fifteen of the subjects (23. 1%) were determined to have acne. Acne prevalence in girls and boys was 16.1% and 29.2%, respectively (P < 0.001). Two hundred and twenty-five (15.8%) of 1424 boys and only 109 (8.8%) of 1233 girls had moderate or severe/very severe acne (P < 0.001), but the GAGS scores in the groups of boys and girls with acne were not significantly different. The acne and control groups showed no significant differences in the HAD anxiety and depression subscale scores. The HAD anxiety subscale scores of girls were significantly higher than those of boys in the acne group. The severity of acne was not correlated with the HAD anxiety or depression subscale scores. CONCLUSIONS: Acne results in higher anxiety in adolescent girls. Although acne and moderate/severe acne are more common in adolescent boys, the severity of acne was found to be similar in boys and girls with acne. Adolescent girls are more vulnerable than boys to the negative psychological effects of acne. PMID- 10849126 TI - Recurrent aphthous stomatitis and smoking. AB - BACKGROUND: Several studies have reported cigarette smoking to have a beneficial protective effect on recurrent aphthous stomatitis (RAS). In this study, we evaluated once again the incidence of smoking in RAS patients compared with controls. This study differs from most previous ones in that the patients were diagnosed by direct observation of active lesions by a dermatologist. METHODS: Thirty-four patients with RAS who were seen at the dermatology clinic during a period of 2 years were compared with 115 outpatients with other skin diseases and 20 healthy hospital personnel who had no history of aphthae, with regard to their smoking habits. RESULTS: Among the 34 patients with RAS, 8.8% were active smokers compared with a significantly higher percentage (25.2%) among the control subjects. CONCLUSIONS: In agreement with others, we found a negative epidemiologic association between smoking and RAS. This finding can be used to clarify the cause and pathogenesis of the disease, and possibly to identify better treatment or preventive options than those currently available. PMID- 10849127 TI - Degos' disease. AB - A 47-year-old woman presented with slightly pruritic, burning lesions on her submammary area and legs of 3.5 years' duration. Similar lesions had appeared on her arms and feet. She was hospitalized at the Pulmonary Diseases Hospital, Izmir, with a complaint of thoracic pain, and was diagnosed with nonspecific pleuritis by pulmonary X-ray, thoracic computed tomography (CT) scanning, bacterial cultures of phlegm, and pleural biopsy examination. Fifteen days after leaving hospital, she was admitted to the Surgical Clinic of Ataturk Training Hospital, Izmir, with a complaint of severe abdominal pain, and diagnosed with nonspecific peritonitis. On dermatologic examination, she had multiple, scattered papules (2-3 mm in diameter), with a typical, central, white, porcelain-like zone of atrophy, on the trunk (Fig. 1), legs (Fig. 2), and arms. The lesions were encircled by a slightly raised erythematous border. Some of the lesions had coalesced giving polycyclic atrophic areas and ulcerations. Histopathologic examination of the skin biopsy specimen showed hyperkeratosis, epidermal atrophy, dermo-epidermal separation, edema, and necrosis in the papillary dermis (Fig. 3). Fibrinoid necrosis and thrombosis were seen in the papillary dermis and in the vessels below the lesions. The patient was diagnosed with Degos' disease from these clinical and histopathologic findings. She was treated with anti inflammatory drugs. One year later, the patient was hospitalized at the Department of Internal Medicine of Ataturk Training Hospital, Izmir, with a diagnosis of pericarditis and pleuritis. PMID- 10849128 TI - Cutaneous and neurologic manifestations of biotinidase deficiency. PMID- 10849129 TI - Retiform hemangioendothelioma. PMID- 10849130 TI - Microcystic adnexal carcinoma induced by multiple radiation therapy. PMID- 10849131 TI - Irritant contact dermatitis secondary to fiberglass: an unusual presentation. PMID- 10849132 TI - Granulomatous slack skin. PMID- 10849133 TI - Late-onset neurofibromatosis in a liver transplant recipient. PMID- 10849135 TI - Dermatology related advertisements from the 1920s and 1930s. PMID- 10849134 TI - Clinical and immunologic parameters during thalidomide treatment of lupus erythematosus. AB - BACKGROUND: Thalidomide is used as an experimental drug for the treatment of chronic inflammatory diseases with an autoimmune or infectious background. The pharmacologic action involves the downregulation of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) and inhibition of basic fibroblast growth factor (bFGF)-induced angiogenesis; however, not much is known about thalidomide's effect on immunologic parameters in lupus erythematosus (LE). Method This is an open study of a group of five consecutive systemic lupus erythematosus (SLE) patients treated with thalidomide (100 mg/day) and five consecutive cutaneous LE patients not responsive to conventional therapy. The clinical and immunologic parameters (C-reactive protein, immunoglobulin, and complement serum levels, lymphocyte counts) were investigated during thalidomide treatment for up to 2 years in both patient groups. RESULTS: An increase in the absolute peripheral lymphocyte count was observed beginning after 2 weeks of systemic thalidomide treatment in nine out of 10 LE patients, and remained stable throughout thalidomide treatment. Elevated serum levels of C-reactive protein and titers of autoantibodies to double-stranded (ds) DNA decreased in SLE patients. No significant changes were detected in the serum levels of the complement components C3 and C4 and immunoglobulins in all LE patients. Regression of inflammatory skin lesions and regrowth of hair were recorded. As a side-effect, polyneuropathy was observed in four out of 10 patients, with the earliest onset at 3 weeks of thalidomide treatment. CONCLUSIONS: Thalidomide is a potent anti inflammatory drug in patients with SLE and cutaneous LE, possibly interacting with the recruitment of lymphocytes. It leads to the regrowth of hair in LE related alopecia and effluvium. Early symptoms of polyneuropathy should be registered and the drug should be withdrawn. Thalidomide should be restricted to patients who show no response to standard therapeutic regimens and should only be used under strict precautions with regard to its known teratogenic risk. PMID- 10849136 TI - Basal cell carcinoma of the nail bed in a korean woman PMID- 10849137 TI - Non-lethal Wiskott-Aldrich syndrome: atopic dermatitis-like lesions persist after splenectomy. PMID- 10849138 TI - The best interest principle as a standard for decision making in the care of neonates. AB - In neonatal care, decisions are made on behalf of newborn infants by their parents or, in some instances, health professionals. This paper examines how the best interest standard is the most appropriate focus for decision-making concerning neonates. The components of surrogate decision-making are discussed from the perspective of the neonate's interests and the contribution of the various persons involved in caring for neonates. An argument is presented for the use of best interests when making decisions and the interpretation of best interests is explored. By examining the ethical approaches using best interests, an argument is put forward for caring as an expression of best interests. How some nurses use the best interest standard in their practice is described. The unique perspective nurses may have because of their philosophy, circumstances, experience and concern for neonates is discussed. Examples are used from the literature to support the argument for nurses being in roles and relationships where the neonate's interests are the basis of practice. How nurses classify infants on the basis of their future outcome is used to illustrate how nurses can apply the best interest standard. Ethical approaches of care and cure are used to show the best interest standard as applied to neonates. Caring as an expression of best interests is recommended for nursing decisions and actions using infant outcomes as a focus for caring and best interests. PMID- 10849139 TI - Asthma and dualism. AB - The rejection of Cartesian dualism can be taken to imply that the mind is implicated in health and illness to a greater degree than conventional medicine would suggest. Surprisingly, however, there appears to be a train of thought in antidualist nursing theory which takes the opposite view. This paper looks closely at an interesting example of antidualist thinking - an article in which Benner and her colleagues comment on the ways in which people with asthma make sense of their condition - and concludes that it places unduly stringent and arbitrary limits on the mind's role. It then asks how antidualism can lead to such a dogmatic rejection of the idea that states of the body are clinically influenced by states of mind. The answer to this question is that Benner assimilates very different philosophical theories into the same 'tradition'. On this occasion, she has combined Descartes, Kant and the Platonist ascetics into a single package, misleadingly labelled 'Cartesianism', and this move accounts for her unexpected views on the relation between mind and body in asthma. PMID- 10849140 TI - Prevention as intervention. AB - Prevention as intervention is identified as a middle-range theory consistent with the Neuman systems model. Through an examination of anxiety research using Neuman's theory, prevention as intervention is examined for clarity, simplicity, generality, accessibility and importance. Conceptual clarity is in need of improvement to proceed with theory testing research. Until such concepts as the flexible and normal lines of defence, created environment, and the inter relatedness of core system variables are further understood, the theory of prevention as intervention remains unclear. PMID- 10849141 TI - Path analysis of efficacy expectations and exercise behaviour in older adults. AB - The benefits of regular exercise for older adults are well documented and include improvements in physical, functional, as well as psychological, health. The purpose of this descriptive study was to test a theoretically and empirically based model describing the factors that influence exercise behaviour of older adults in the United States of America. The hypothesized model suggested that age, gender, and mental and physical health have an effect on self-efficacy and outcome expectations, and that all these variables influence exercise behaviour. Exercise behaviour was hypothesized to have a reciprocal relationship with self efficacy expectations and mental and physical health. The convenience sample was 187 older adults living in a continuing care retirement community (CCRC) in Baltimore, Maryland. A one-time health interview was conducted which included a measure of self-efficacy and outcome expectations related to exercise, a measure of health status (SF-12), and gathering of information from participants about their actual exercise behaviour. Of the 187, 71 (38%) reported participating in 20 minutes of continuous aerobic exercise at least three times per week over the previous 3 months. Six hypothesized paths were significant. The model fitted the data and accounted for 32% of the variance in exercise behaviour. Interventions that focus on strengthening self-efficacy and outcome expectations can improve exercise behaviour in older adults. PMID- 10849142 TI - Evidence-based practice and health visiting: the need for theoretical underpinnings for evaluation. AB - In this paper we argue that evidence-based practice, which is being introduced throughout the British National Health Service to make decisions about the allocation of limited resources, provides a welcome opportunity for health visitors to demonstrate their efficacy, skills and professionalism. However, the paper argues that to view health visiting as evidence-based is not to reduce health visiting merely to a technology through which scientific solutions are applied to social problems. Rather, health visiting needs to be viewed as a political movement, based on a particular model of society, which shapes the goals which health visitors pursue and influences the strategies they adopt to achieve their goals. The paper describes various models of health visiting as a way of showing how the goals of health visiting are always framed within a particular set of assumptions and causal explanations. The paper then turns to look at the issue of evaluating health visiting services. It is argued that evaluation should properly take account of the models which shape health visitors' goals and intervention strategies, and in turn, health visitors need to be explicit about the theoretical frameworks underpinning their interventions. Finally, it is argued that health visitors' knowledge and understanding of a range of models of society enables them to move between the various models to choose the most appropriate and effective means of intervention. Hence it is concluded that the emphasis on evidence-based practice provides health visitors with a valuable opportunity to show that their unique, professional skills and understanding are the preconditions for effective intervention. PMID- 10849143 TI - Developing a concept analysis of control for use in child and adolescent mental health nursing. AB - The need to help children and young people with significant mental health problems develop a sense of personal control in their everyday lives, in a manner which does not endanger themselves or others, was recognized by nurse practitioners working in an English regional multidisciplinary child and adolescent mental health residential unit. A concept analysis of control was undertaken and used to develop a framework for analysing control. This deductive framework was modified iteratively by nurses who developed new knowledge from a qualitative exploration of current practice and the application of the evolving framework to practice problems. The paper describes this process and highlights three main findings: (i) the evolving attributes of the concept analysis helped nurses steer a course through the complexities of practice; (ii) the research highlighted and enabled nurses to confront the paradoxical nature of control; (iii) the process enabled nurses to recognize the mutuality of feelings aroused simultaneously when both the nurse and the child are challenged to maintain personal control. PMID- 10849144 TI - The Roper, Logan and Tierney (1996) model: perceptions and operationalization of the model in psychiatric nursing within a health board in Ireland. AB - This study aimed to determine whether the Roper, Logan & Tierney model was an appropriate model for planning nursing care for clients who are mentally ill. Data were collected from two sources in one Health Board region in Ireland. A care plan audit was used to evaluate the extent to which the Roper, Logan & Tierney model was used to assess, plan and evaluate nursing care in nursing documentation. Qualitative interviews with nurses explored their experiences of using the model and their perceptions of the model's usefulness and appropriateness for planning care. Both data sets were complementary, the qualitative data often providing contextual information which helped put the findings into perspective. It was found that there was little evidence that the Roper, Logan and Tierney model guided care planning and that goals and nursing interventions were frequently not explicitly documented. Interviews with nurses indicated that they lacked educational preparation for using the model and found the model constraining and physically orientated. The appropriateness of the Roper, Logan and Tierney model for planning care for clients who are mentally ill is questioned. It is suggested that nurses need to be adequately prepared if they are to use a model appropriately. Consideration should be given when selecting a model as to its 'fit' with the needs of the client group and the ward team philosophy. PMID- 10849145 TI - Feeling of 'lacking' as the core of envy: a conceptual analysis of envy. AB - The aim of this paper is to explore the subjective experience of envy through concept analysis. Further, the study on which it is based aimed to answer questions about the composition and manifestations of envy. From the viewpoint of nursing science, the analysis of envy is based on a desire to understand human beings from the perspective of subjective health and illness and thus from a health promotion perspective. Envy is conceived of as a dimension of a person's health and illness. The concept is therefore meaningful from the viewpoint of nursing; it describes a phenomenon which enables us to deepen our understanding in a way relevant to nursing science. In the study the hybrid model developed by Schwartz-Barcott et al. was used for conceptual elaboration. In the theoretical phase of the study the subjective experience of envy was explored from the viewpoints of philosophy, religion, Finnish folklore and psychoanalysis, as well as nursing science. As a synthesis of these, a conceptual analysis of envy adapted from Wilson was conducted and a working definition of envy was proposed. In the fieldwork phase, envy was examined by means of an empirical analysis using a phenomenological approach. As a result, a classification describing the experience of envy was presented. The core experience of envy has been defined as a 'lacking', and the object of envy as something good possessed by someone else. Envy manifests itself in both destructiveness and creativity. The trends of development of envy are inflexibility and emancipation, and the essence of envy is multidimensional. Finally, the working definition of the concept was elaborated on the basis of the empirical phase and a new definition reflecting the composition and manifestations of envy was proposed. PMID- 10849146 TI - Myocardial infarction: survivors' and spouses' stress, coping, and support. AB - Despite evidence that spouses play an important role in the recovery of MI survivors, there have been few studies of pertinent psychosocial factors from the perspectives of both survivors and spouses. Accordingly, the aim of this study was to describe stress, coping strategies and social support experienced by survivors and spouses. This study was limited to first-time MI to focus on a time of uncertainty and transition. Twenty-eight persons (14 couples) participated. Both survivors and their spouses reported similar post-MI stresses: emotional impact, lifestyle changes, encounters with health professionals, and their partners' reactions. Spouses and survivors used diverse strategies to cope with the stresses of MI. Seeking informational support was prevalent. Both spouses and survivors engaged in 'protective buffering' of their partners. Couples described deficient support, conflict and miscarried helping efforts within their relationships. Spouses and survivors referred to inadequate informational support from health professionals. PMID- 10849147 TI - Differences between men and women on the waiting list for coronary revascularization. AB - This study aims to examine the situation for patients on the waiting list for possible coronary revascularization in terms of waiting time, treatment and various aspects of well-being in relation to gender. Patients on the waiting list for coronary angiography, percutaneous transluminal coronary angioplasty or coronary artery bypass grafting in September 1990 were approached with a questionnaire dealing with various aspects as described above. Of the 831 patients who participated in the evaluation, 174 (21%) were women. Although age was similar for men and women, men had a higher prevalence of previous myocardial infarction and a lower prevalence of previous hypertension. In terms of medication, women were more frequently treated with diuretics and sedatives than men. Women reported a higher frequency than men with regard to the following symptoms: chest pain at rest and at night, dyspnoea when walking, tachycardia, tiredness, headache, dizziness and sweating. Women also suffered more frequently from difficulty going to sleep, difficulty waking up, repeated awakening and insomnia. Men, on the other hand, suffered more frequently from restlessness, inability to act and irritability. Among patients on the waiting list for possible coronary revascularization, women differed from men by being more frequently treated with diuretics, reporting a higher frequency of various cardiovascular symptoms including chest pain and dyspnoea and, furthermore, reporting more sleeping disorders. Gender differences were found but they were not consistent. PMID- 10849148 TI - Psychosocial experiences of cardiac patients in early recovery: a community-based study. AB - OBJECTIVE: To report on the nature, incidence and severity of problems commonly experienced by cardiac patients in the early months of recovery, and to test the hypotheses that there exist differences in the incidences of these problems depending on age and sex. METHODS: 1124 emergency cardiac patients discharged from hospital with acute myocardial infarction, unstable angina, stable angina pectoris, chronic ischaemic heart disease or heart failure were surveyed 4 months after discharge. They were asked to indicate how often during the previous 2 weeks they had experienced each of a range of feelings and problems common to cardiac patients. RESULTS: A large proportion of patients reported experiencing problems in the areas of emotional reactions (70%), physical condition (79%), convalescence (67%) and relating to family and friends (63%). Severe problems were experienced especially in the physical and convalescence areas (43% and 44%, respectively). A greater proportion of patients diagnosed with heart failure experienced problems than those with other diagnoses, and these problems were more severe. Amongst myocardial infarction patients, a greater proportion of females than males reported severe problems in the emotional and physical areas, and patients 65 years and over were more likely than younger patients to report experiencing severe problems with physical condition. CONCLUSIONS: Many cardiac patients are experiencing psychosocial problems 4 months after hospital discharge, especially with physical activities and convalescence. A knowledge of the incidence and nature of these problems may help nurses to assist patients to validate their experiences. PMID- 10849149 TI - The introduction of a routine monitoring system in primary care for patients with a first episode of cardiovascular disease. AB - A study at a group general practice in the English midlands found that health promotion advice had not been routinely provided to some patients with cardiovascular disease and stroke. The purpose of this project therefore was to introduce a monitoring system to ensure that health promotion issues were covered systematically with patients following a first episode of cardiovascular disease. Patients with a first episode of a relevant condition would be identified by an automated search on the practice database, and contacted by the health visitor. A checklist would ensure that all appropriate issues were covered. The system was easily introduced at the practice and no difficulties were experienced with its administration. A total of 62 patients were seen during the year. A substantial number of secondary prevention issues were addressed through advice and information leaflets. The project was felt to be a useful addition to care by the workers involved. Although some of the issues may have been addressed in routine care, early organized nurse contact ensures systematic coverage and early referrals where necessary, as well as potential psychological benefit to patients. PMID- 10849150 TI - Sense of coherence: quality of life before and after coronary artery bypass surgery--a longitudinal study. AB - The attention to patient outcome has nowadays extended from morbidity and mortality to an aspect of patients' benefits in terms of quality of life. One factor crucial for quality of life is coping capacity, in this study represented by the sense of coherence concept. Physical status and emotional state (often measured by comprehensive instruments not always suitable for clinical use) are also additionally used to reflect quality of life. The purpose was therefore to study sense of coherence and emotional state as indirect measures of quality of life in relation to coronary artery bypass grafting surgery. One hundred and eleven patients were studied by a developed questionnaire on five occasions in relation to the surgery: the week before the angiography, the day before surgery and then at 3, 6, and 12 months post-operatively. The main findings were: (1) The sense of coherence was changed (more than +/-10%) from before to 1 year after surgery in 41% of the patients, which is contrary to the theory of sense of coherence as a stable personality characteristic in adults. (2) Experience of depressed mood, stress, and anxiety decreased significantly from before to after surgery. (3) Beneficial outcome with regard to sense of coherence was significantly related to less experience of loneliness, depressed mood, stress and anxiety, and to less experience of chest pain 1 year after surgery. In conclusion, sense of coherence and emotional state variables, are suggested to be valuable as measurements of quality of life in relation to coronary artery bypass grafting surgery. PMID- 10849151 TI - Coping with type-2 diabetes: the role of sense of coherence compared with active management. AB - Changes in lifestyle, particularly in dietary and exercise habits, are necessary for the majority of patients with type-2 diabetes but are difficult to carry out. However, Antonovsky describes a salutogenic health perspective grounded in patients' developing what he terms 'a sense of coherence' (SOC). Can a strong SOC help diabetes patients to control the disease? The aim of this study was to analyse the relationship between SOC and treatment results measured as glucolysed haemoglobine (HbA1c) in patients with type-2 diabetes. The aim was further to test the relationship between treatment results and an index of patients' participation in active management and emotional state. Eighty-eight patients answered a questionnaire containing 13 statements about sense of coherence (SOC 13), questions about self-assessed health, diabetes activity such as self management of diet, exercise and self-control of blood sugar and emotional acceptance. There was no direct relationship between SOC-13 and treatment results measured as HbA1c but there was a positive correlation between SOC-13, self assessed health and HbA1c (P < 0.02). Self-assessed health was seen as a mediating factor. The better patients' estimation of their own health, the higher were SOC-13 scores and the lower HbA1c. There was also a strong positive correlation between low levels of HbA1c and high levels of an index of active management and emotional acceptance of diabetes (P < 0.001). PMID- 10849152 TI - Coping strategies and health-related quality of life among spouses of continuous ambulatory peritoneal dialysis, haemodialysis, and transplant patients. AB - In the study reported here 55 spouses of patients living with end-stage renal disease (ESRD) were investigated with respect to coping strategies and health related quality of life. Findings from the study were compared to two random samples of the Swedish general population (n = 454, and n = 1200). The study design was correlational and comparative. Coping was measured by the Jalowiec Coping Scale, and quality of life (QoL) by the Swedish Health-Related Quality of Life Survey (SWED-QUAL). Data were analysed using a number of statistical tests including Pearson's product moment correlations, Student's t-test and two way ANOVAs. The combined sample of spouses used significantly more optimistic and palliative coping than the general population, but less confrontative, self reliant, evasive and emotive coping. In the study fatalistic, evasive and emotive coping was associated with low perceived efficiency in handling various aspects of the partners' situation. The male spouses used significantly less optimistic, supportive and palliative coping than did the female spouses. The spouses of transplant patients had better overall quality of life than the continuous ambulatory peritoneal dialysis and haemodialysis spouse groups, most likely due to the lower age of the former group. The study findings suggest that emotive, evasive and fatalistic coping are less than optimal ways to deal with problems occasioned by the partner's treatment. PMID- 10849153 TI - Spousal perspective of Parkinson's disease in middle life. AB - An interpretive study in America explored the experience of living with a partner who has Parkinson's disease (PD) in middle life. Challenges experienced by eight spouses and their ways of coping with these challenges were examined. The convenience sample of five wives and three husbands were 44-58 years and had been married 4 months to 30 years. Spouses described the most significant challenges as watching their partner struggle and be frustrated; and renegotiating their lives. The coping strategies most frequently used were maintaining their own life, encouraging their partner to stay active and involved, and seeing the challenges they experienced as secondary. The context of living with a partner with Parkinson's disease is illuminated and strategies that spouses find most effective in coping with their partners illness are identified. PMID- 10849154 TI - End-of-life care preferences of Canadian senior citizens with caregiving experience. AB - A grounded theory study of senior citizens' preferences for end-of-life care was conducted in 1998 in Canada. Seniors who had experienced the deaths of others and who had considered their own death and dying were the target population. The sample was 49 seniors who met the study criteria. Participants provided end-of life care in a variety of settings for 1-8 family members or friends. Two concepts identified were expected dependency while dying and appropriate end-of life care. Almost all participants preferred to be cared for at home, yet family caregivers who could provide appropriate end-of-life care when dependent were needed for this to occur. The appropriateness of end-of-life care was contingent upon the place where end-of-life care occurred, as well as the type of care provided. Life prolongation was not desired, 53% even endorsed euthanasia as a way of bringing about the inevitable end to life. PMID- 10849155 TI - Late adolescent female smoking. AB - Although there have been intense efforts to reduce the prevalence of cigarette smoking in the past three decades, smoking continues to be a critical public health problem. An area of particular concern is the increasing number of young women who are smoking. Gender specific research usually does not examine factors affecting smoking behaviours. Information on late adolescent female smoking is not readily found in the literature. The aims of this study were to explore the smoking patterns and processes of late adolescent females and to explore factors which may or may not be helpful in assisting them to stop smoking. A qualitative ethnographic approach was used to uncover the perceptions of 25 adolescent girls of their behaviour. Data were collected using tape-recorded semi-structured interviews and a questionnaire. Items for the questionnaire were derived from the Manitoba Youth Smoking Survey and from the Fagerstrorn Nicotine Tolerance Scale. Qualitative analysis resulted in four stories about smoking: the start story, the smoking story, the quit story, and the looking to the future story. Although many of the findings are congruent with the existing literature, much of the data reflected the purposive nature of smoking in late adolescent girls. Study findings support the need for holistic approaches to health promotion to ameliorate factors affecting smoking behaviour. PMID- 10849156 TI - Perceptions of the role of the school in providing information and support to adolescent children of women with breast cancer. AB - Little is known about the needs of adolescents of women with breast cancer. The purpose of this study was to describe the perceptions of adolescents (ages 12-20) about the role of the school in assisting them in dealing with the cancer experience. An exploratory, qualitative study was done to elicit detailed descriptions of adolescent's needs for information and support in response to their mother's breast cancer. A convenience sample of 31 adolescents of women in five illness phases participated in semi-structured interviews. In addition, two focus group interviews were conducted. Interviews were tape-recorded, transcribed and analysed using content analysis techniques. Findings specific to the adolescents' perceptions of the role of the school were discussed according to content, type, amount, timing, provider and quality of information. Support needs were discussed as type of support, source of support, amount, timing and focus. Although school personnel attempted to be supportive and adolescents received generic information about cancer, overall the needs of the adolescents were not adequately addressed. Recommendations for schools and health services in assisting adolescents to cope with this major life experience are made. PMID- 10849157 TI - Effects of infant and caregiving conditions on an infant's focused exploration of toys. AB - An infant's exploration of toys, in the context of the mother's regulating actions, is a setting for cognitive and social development. The aim of this study was to examine the contribution of infant and caregiving conditions to the quantity of focused exploration of toys for 8-month-old infants. Infant biological conditions were gender and birth weight, including a range of both prematurely and term-born infants. The infant behavioural condition was responsiveness to care. Caregiving conditions included mother's education, the supportive, stimulating and sustaining quality of the home environment, and the attention-directing and -supporting behaviour of the mother during play. The direct and indirect effects of these conditions on focused toy exploration were modelled using multiple regression. The sample of 79 mother-infant dyads included 43 full-term infants and 36 premature infants. The mother's attention-directing behaviour was a significant negative predictor of focused toy exploration. Infant birth weight did not have a significant direct effect on focused toy exploration. Birth weight did not interact significantly with responsiveness to care or with any of the caregiving environment conditions to affect focused toy exploration. Further study of infant responses to the mother's attention-regulating and emotion-regulating behaviour during play is recommended to better understand how the caregiving environment supports or thwarts focused toy exploration. PMID- 10849158 TI - Innovations in the nursing care of the chronically ill: a literature review from an international perspective. AB - This literature review focuses on substitution-related innovations in the nursing care of chronic patients in six western industrialized countries. Differences between primary and secondary care-orientated countries in the kind of innovations implemented are discussed. Health care systems are increasingly being confronted with chronic patients who need complex interventions tailored to their individual needs. However, it seems that today's health care professionals, organizations and budgets are not sufficiently prepared to provide this kind of care. As a result, health care policy in many countries targets innovations which reduce health care costs and, at the same time, improve the quality of care. Frequently, these innovations are related directly to the 'substitution of care' phenomenon, in which care is provided by the most appropriate professional at the lowest cost level, and encompass advanced nursing practice, hospital-at-home care and integrated care. The main conclusion of this paper is that integrated care innovations are implemented in both primary care as well as in secondary care orientated countries. However, innovations in hospital-at-home care and advanced nursing practice are primarily implemented in primary care-orientated countries. Whether these innovations positively influence the quality of care, costs of care or patients' use of health care facilities remains rather unclear. PMID- 10849159 TI - Cognitive ability and functional status. AB - Significant confusion exists in the literature about functional status. Despite its importance, little attention has focused on developing and substantiating frameworks that detail the underpinnings of functional status, which has resulted in lack of agreement about its definition and dimensions. The purpose of this literature review was to examine the development of functional status and to describe the inclusion of its cognitive dimension. Cognition is one key dimension of functional status. One must 'know how' to perform to be successful in an activity. While cognitive capacity is generally considered in relation to functional status, the nature of the cognitive dimension is poorly described and poorly understood. Three databases were selected for review: Citations in Nursing and Allied Health (CINAHL), Psychology Literature (PsychLit), and the Medical data base known as MedLine. Key word searches identified thousands of sources. This analysis includes an extensive sampling of these sources from the 1960s through to 1998. The sources sorted into four primary categories and demonstrate a growing recognition of the cognitive dimension of functional status in the literature. Despite this recognition, the lack of conceptual clarity of both the term functional status and its cognitive dimension limits communication among disciplines and limits comparisons of functional status outcomes across studies. Functional status models are needed that include cognition as a core dimension. Population specific descriptions of the cognitive dimension should be guided by knowledge in the neurosciences. PMID- 10849160 TI - The political role of illness narratives. AB - Cultural criticism is used to describe the political role of autobiographical illness narratives or pathographies. In expressing the subjective experience of illness, authors of pathographies illuminate ideological differences between patient and health care cultures, reveal the dominance of health care ideologies, and explicate patients' moral and political claims. The contributions of these literary works to nursing practice provide direction for relational restructuring. Gadow's concept of the relational narrative is proposed as a way to restore patient subjectivity and agency and establish the dialogue necessary for cultural pluralism in nursing and health care. PMID- 10849161 TI - Methodological issues in grounded theory. AB - Examination of the qualitative methodological literature shows that there appear to be conflicting opinions and unresolved issues regarding the nature and process of grounded theory. Researchers proposing to utilize this method would therefore be wise to consider these conflicting opinions. This paper therefore identifies and attempts to address four key issues, namely, sampling, creativity and reflexivity, the use of literature, and precision within grounded theory. The following recommendations are made. When utilizing a grounded method researchers need to consider their research question, clarify what level of theory is likely to be induced from their study, and then decide when they intend to access and introduce the second body of literature. They should acknowledge that in the early stages of data collection, some purposeful sampling appears to occur. In their search for conceptually dense theory, grounded theory researchers may wish to free themselves from the constraints that limit their use of creativity and tacit knowledge. Furthermore, the interests of researchers might be served by attention to issues of precision including, avoiding method slurring, ensuring theoretical coding occurs, and using predominantly one method of grounded theory while explaining and describing any deviation away from this chosen method. Such mindfulness and the resulting methodological rigour is likely to increase the overall quality of the inquiry and enhance the credibility of the findings. PMID- 10849162 TI - Interviewing in phenomenology and grounded theory: is there a difference? AB - This paper explores the differences and similarities that may exist in respect of using the interview method in phenomenological and grounded theory methodologies. Baker et al. set out to differentiate between method in grounded theory and phenomenology and concluded that it was essential to ensure that the method matches the research question being asked. However, the paper, whilst clear in intent to differentiate between the methodologies of phenomenology and grounded theory, does little to help the researcher in the differences that may exist in carrying out such research using the same method, that is, interviewing. Interviewing has become synonymous with qualitative research and may become the accepted method of data collection irrespective of methodology. We postulate that the interview as a method of data collection may be inconsistent with the underlying principles of the methodology (phenomenology or grounded theory). Should this be the case then the interview as a means of collecting data may be viewed as generic and lack a clear connection to the methodological framework. Such a position could be consistent with a critique of qualitative nursing research on the grounds of rigour. PMID- 10849163 TI - One voice, different tunes: issues raised by dual analysis of a segment of qualitative data. AB - Qualitative data analysis is a complex and contested part of the research process that has received limited theoretical attention. This paper explores the relationship between the way in which data are analysed and the nature of findings that emerge. It does this in response to demands to recognize the multiple voices that inform representations of reality, and debates about whether the interpretation of data reveals or constructs meaning. A small segment of data provided by one informant is subjected to both thematic and narrative analysis and the different perspectives that emerge are discussed with reference to whether different kinds of analysis lead to different kinds of meaning being imputed to the same text. The paper suggests that, rather than provide a unified and ever-more refined version of 'reality', the use of dual or multiple analysis helps to elucidate alternative interpretations of the data which might escape consideration with the use of a single approach. PMID- 10849164 TI - Experiences of AIDS-dedicated nurses in alleviating the stress of AIDS caregiving. AB - From the beginning of the AIDS epidemic, there have been individuals dedicated to the care of patients with AIDS. However, there has been little research regarding their perceptions and experiences of AIDS caregiving and the strategies they use to alleviate the stress and promote their willingness to care. Based on the experiences of 12 nurses at one hospital, who had chosen to work on an AIDS dedicated unit, this exploratory study, conducted in 1998, explored the following: the physical, emotional or spiritual risks and stresses associated with AIDS caregiving; factors that provide resistance to the stresses of AIDS caregiving and promote a willingness to care; and strategies recommended by AIDS dedicated nurses in caring for patients with AIDS. The data reveal important themes related to the physical stress of AIDS caregiving, specifically being aware of risks, but not paralysed by fear, and bombardment of the senses. The coping strategies of nurses included taking the risk in their stride, reframing the risk, and protecting oneself. The emotional stress of AIDS caregiving included witnessing suffering, experiencing unresolved grief, accepting diversity, being emotionally connected, distress from the dismantling of the AIDS unit and work demands, and declining team spirit. Coping strategies included balancing personal and professional life, releasing pain, respecting yet controlling feelings, managing demands, and asking for help. Nurses maintained their spiritual perspective. They experienced through AIDS caregiving a greater sense of shared humanity and a new perspective of life. Findings indicate that AIDS-dedicated nurses use many coping strategies. The experiences of these nurses can assist clinicians, educators and administrators in supporting nurses' caregiving and promoting the quality of care offered to patients with AIDS. PMID- 10849165 TI - A phenomenological study of early nursing experiences in Hong Kong. AB - The experience of primary professional socialization is crucial for neophytes to learn to become a nurse. These early nursing encounters may also have long-term effects on professional development of individual nurses. However, research into the early experiences of nurses has been poorly documented. This study endeavours to reveal the early lived nursing experience amongst a group of nurses in Hong Kong. This study adopts a phenomenological approach which involves the thematic analysis of the critical incidents provided by 77 subjects. Findings revealed that incidents associated with death and dying, and clinical learning embracing interpersonal relations and professional development, were the most memorable events. Nurses were in general not equipped adequately to communicate with the dying and the grieving relatives. The subjects disclosed that positive clinical encounters confirmed their value of nursing work and motivated them to stay in the profession. Conversely, the negative experiences made them seriously consider leaving nursing. A number of implications for nursing education have been drawn from the research findings. PMID- 10849166 TI - Factors related to stress and coping among Chinese nurses in Hong Kong. AB - Few empirical studies have investigated the issues linked to Hong Kong nurses work-related health. The present study investigated factors related to stress and coping among Chinese nurses in Hong Kong. The researchers employed a cross sectional survey and made within-group comparisons of nurses' stress and coping. Using stratified random sampling the researchers selected nurses from the mailing list of a local professional organization. One hundred and sixty-eight (33.6%) nurses responded. Nurses reported lower stress levels than other workers assessed with the same measure. Paediatric nurses reported the highest stress levels. Nurses at the lower grades reported higher stress levels than nurses at the higher grades. Single nurses had marginally higher stress scores than married nurses and females had slightly higher stress scores than males. However, none of these results were statistically significant. The respondents' major sources of stress were related to nursing issues like too much work, interpersonal relationships, and dealing with hospital administration. The respondents coped with their stresses by seeking support from friends and colleagues, using different cognitive strategies and through leisure activities. There was a statistically significant link between the respondents' stress and sickness levels. The results raise issues about the nature of nurses' working experiences. PMID- 10849167 TI - Predictive models of the combined effects of curvaticin 13, NaCl and pH on the behaviour of Listeria monocytogenes ATCC 15313 in broth. AB - Thirty-three strains of Listeria monocytogenes belonging to different serotypes were tested for their sensitivity to curvaticin 13, an antilisterial bacteriocin produced by Lactobacillus curvatus SB13, using the well diffusion method in Institut Pasteur agar plates at 37 degrees C. No relationship between serotype and sensitivity was observed. The sensitivity of this species was strain dependent and a large variation in tolerance to curvaticin 13 was observed. The combined effects of curvaticin 13 (0-160 AU ml-1), NaCl (0-6% w/v), pH values (5.0-8.2) and incubation time (0-24 h) were investigated on L. monocytogenes ATCC 15313 in trypcase soy-yeast extract broth at 22 degrees C. For this study, two Doehlert matrices were used in order to investigate the main effects of these factors and their different interactions. The results were analysed using the Response Surface Methodology. Curvaticin 13 had a major inhibitory effect and the response was NaCl concentration-, time- and pH-dependent. This inhibitory activity was the same at pH values between 6.6 and 8.2. Curvaticin 13 was bactericidic at acidic pH values, but the surviving cells resumed growth. For a short incubation time (12 h), the effectiveness of curvaticin 13 was maximal in the absence of NaCl. For longer incubation times (12-48 h), with high NaCl (6%) and curvaticin 13 concentrations (160 AU ml-1), the inhibition of L. monocytogenes was greater than that observed with NaCl or curvaticin 13 alone. PMID- 10849168 TI - Antibiotic-resistant gram-negative bacteria in a virtually closed water reticulation system. AB - The effect of the effluent from a chicken meat-processing plant on the antibiotic resistant bacterial profile was investigated in an almost closed water reticulation system. Of the 273 faecal coliform isolates 256 (93%) were resistant to one or more of the eight antibiotics tested. The most prevalent isolates were for the beta-lactam antibiotics ampicillin and cephalothin followed by the sulphonamides sulphatriad and cotrimoxazole. Eleven different resistance patterns were identified with a single pattern, comprising of ampicillin-, cephalothin-, streptomycin-, sulphatriad-, cotrimoxazole- and tetracyclin-resistant isolates, dominating the meat-processing effluent. An apparent correlation was observed between the specific use of certain antibiotics and the prevalence of the corresponding resistant bacterial isolates. The drugs used to treat the occasional infections, belonging to the beta-lactam and sulphonamide group of antibiotics, seemed to have a more pronounced effect on the antibiotic-resistant bacterial profile in the primary water source than those drugs used as feed additives, oxytetracyclin and the aminoglycoside flavomycin. PMID- 10849169 TI - Stepwise quantitative risk assessment as a tool for characterization of microbiological food safety. AB - This paper describes a system for the microbiological quantitative risk assessment for food products and their production processes. The system applies a stepwise risk assessment, allowing the main problems to be addressed before focusing on less important problems. First, risks are assessed broadly, using order of magnitude estimates. Characteristic numbers are used to quantitatively characterize microbial behaviour during the production process. These numbers help to highlight the major risk-determining phenomena, and to find negligible aspects. Second, the risk-determining phenomena are studied in more detail. Both general and/or specific models can be used for this and varying situations can be simulated to quantitatively describe the risk-determining phenomena. Third, even more detailed studies can be performed where necessary, for instance by using stochastic variables. The system for quantitative risk assessment has been implemented as a decision supporting expert system called SIEFE: Stepwise and Interactive Evaluation of Food safety by an Expert System. SIEFE performs bacterial risk assessments in a structured manner, using various information sources. Because all steps are transparent, every step can easily be scrutinized. In the current study the effectiveness of SIEFE is shown for a cheese spread. With this product, quantitative data concerning the major risk-determining factors were not completely available to carry out a full detailed assessment. However, this did not necessarily hamper adequate risk estimation. Using ranges of values instead helped identifying the quantitatively most important parameters and the magnitude of their impact. This example shows that SIEFE provides quantitative insights into production processes and their risk-determining factors to both risk assessors and decision makers, and highlights critical gaps in knowledge. PMID- 10849170 TI - Diversity of antifungal and plant-associated Serratia plymuthica strains. AB - A total of 21 plant-associated Serratia plymuthica strains were characterized phenotypically by their nutritional patterns, susceptibility to antibiotics, antifungal and haemolytic properties, and genotypically by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA, PCR fingerprints using BOX primers (BOX-PCR) and pulsed-field gel electrophoresis (PFGE) after digestion with SpeI. All of the investigated strains demonstrated antifungal activity in vitro against fungal pathogens while only six strains produced the antifungal antibiotic prodigiosin. Haemolytic activity and antibiotic resistance patterns were investigated to assess the risk associated with the use of isolates in plant protection. The strains were haemolytic at human-relevant temperatures. The level of resistance to antibiotics was low. This work shows that BOX-PCR and PFGE are useful fingerprinting methods to characterize Ser. plymuthica strains, although the discriminatory effect between the two methods differed. Computer-assisted analysis of phenotypic and genotypic features demonstrated relationships between the origin of isolation, the production of prodigiosin and the molecular fingerprint. PMID- 10849171 TI - Characterization of an extracellular enzyme system produced by Micromonospora chalcea with lytic activity on yeast cells. AB - Growth of Micromonospora chalcea on a defined medium containing laminarin as the sole carbon source induced the production of an extracellular enzyme system capable of lysing cells of various yeast species. Production of the lytic enzyme system was repressed by glucose. Incubation of sensitive cells with the active component enzymes of the lytic system produced protoplasts in high yield. Analysis of the enzyme composition indicated that beta(1-->3) glucanase and protease were the most prominent hydrolytic activities present in the culture fluids. The system also displayed weak chitinase and beta(1-->6) glucanase activities whilst devoid of mannanase activity. Our observations suggest that the glucan supporting the cell wall framework of susceptible yeast cells is not directly accessible to the purified endo-beta(1-->3) glucanase and that external proteinaceous components prevent breakdown of this polymer in whole cells. We propose that protease acts in synergy with beta(1-->3) glucanase and that the primary action of the former on surface components allows subsequent solubilization of inner glucan leading to lysis. PMID- 10849172 TI - Effects of fructooligosaccharides and their monomeric components on bile salt resistance in three species of bifidobacteria. AB - The influence of fructooligosaccharides (FOS) and their monomeric components on bile salt resistance of Bifidobacterium breve ATCC 15700, Bif. longum ATCC 15707 and Bif. animalis ATCC 25527 was examined. The neosugars induced fructofuranosidase activities for the degradation of these saccharides. For the three strains tested the growth was identical and bile salts had the same inhibitory effect on growth whatever the carbohydrate used. The survival of Bif. breve and Bif. longum, in the presence of glycodeoxycholic acid depended, however, on carbohydrates: the toxic effects of the bile salt could be partly alleviated by the addition of a metabolizable C-source. For Bif. animalis, the presence of any carbohydrate in the incubation medium did not enhance the viability of the strain. But in the three deconjugating strains of bifidobacteria studied, the presence of neosugar during the growth led to improved resistance to the bactericidal effect of the bile salt compared with the monomeric components of these neosugars (glucose and fructose). PMID- 10849173 TI - Production and properties of hemicellulases by a Thermomyces lanuginosus strain. AB - Thermophilic fungi producing extremely high beta-xylanase and their associated hemicellulases have attracted considerable attention because of potential industrial applications. Thermomyces lanuginosus strain SSBP isolated from soil, produced beta-xylanase activity of 59 600 nkat ml-1 when cultivated on a medium containing corn cobs as substrate and yeast extract as nitrogen source. Lower beta-xylanase activities were produced after growth on other xylan substrates, sugars and soluble starch. Other hemicellulases were produced extracellularly at significantly lower levels than the beta-xylanase activity produced on corn cobs. No cellulase activity was observed. The optimal conditions for beta-xylanase production were 50 degrees C and pH 6.5, whereas 70 degrees C and between pH 5. 5 and 9.5 were optimal for beta-xylanase activity. The temperature optima for other hemicellulases were less than the xylanase with the exception of beta mannosidase. The pH optima of the other hemicellulases were between 5.0 and 6.5. Xylanase was stable up to 70 degrees C and between pH 5.5 and 9.0 for 30 min whereas the other hemicellulase were less stable. These results suggest that the most suitable conditions for hydrolysis of hemicellulose by these enzymes would be at 50 degrees C and pH 6.0. PMID- 10849174 TI - Reduction and precipitation of chromate by mixed culture sulphate-reducing bacterial biofilms. AB - The ability of sulphate-reducing bacterial biofilms to reduce hexavalent chromium (Cr(VI)) to insoluble Cr(III), a process of environmental and biotechnological significance, was investigated. The reduction of chromate to insoluble form has been quantified and the effects of chromate on the carbon source utilization and sulphate-reducing activity of the bacterial biofilms evaluated. Using lactate as the carbon/energy source and in the presence of sulphate, reduction of 500 micromol l-1 Cr(VI) was monitored over a 48-h period where 88% of the total chromium was removed from solution. Mass balance calculations showed that ca 80% of the total chromium was precipitated out of solution with the bacterial biofilm retaining less than 10% of the chromium. Only ca 12% of the chromate added was not reduced to insoluble form. Although Cr(VI) did not have a significant effect on C source utilization, sulphate reduction was severely inhibited by 500 micromol-1 Cr(VI) and only ca 10% of the sulphate reducing activity detected in control biofilms occurred in the presence of Cr(VI). Low levels of sulphide were also produced in the presence of chromate, with control biofilms producing over 10-times more sulphide than Cr(VI)-exposed biofilms. Sulphide- or other chemically-mediated Cr(VI) reduction was not detected. The biological mechanism of Cr(VI) reduction is likely to be similar to that found in other sulphate reducing bacteria. PMID- 10849175 TI - Physicochemical surface properties of five Listeria monocytogenes strains from a pork-processing environment in relation to serotypes, genotypes and growth temperature. AB - Physicochemical surface properties, related to electrostatic, van der Waals and Lewis acid-base interactions, of five Listeria monocytogenes strains isolated from pork-processing environments were determined after two subcultures at 37 degrees C and a final culture at three temperatures: 37, 10 and 4 degrees C. Three strains (Lm1, Lm114 and Lm191) were genetically related while two were unrelated (Lm25 and Lm74) according to ApaI-macrorestriction and pulsed-field gel electrophoresis (PFGE) typing. Listeria monocytogenes cell surfaces were generally negatively charged regardless of pH and tended to be hydrophilic due to a basic character. However, variable physicochemical surface properties of the five Listeria monocytogenes isolates were observed after growth at 37 degrees C. After growth at 10 degrees C, the three genetically related isolates exhibited similar surface properties and were slightly more hydrophilic and basic than the others. After growth at 4 degrees C, the five isolates displayed the same weak affinity for all kinds of solvents and low electrophoretic mobility values. A sharp decrease of temperature and subsequent growth of various Listeria monocytogenes strains resulted in loss of the physicochemical surface property variability, which may suggest the role of common chill adaptation mechanisms affecting surface properties. PMID- 10849176 TI - The effect of temperature and selective agents on the growth of Yersinia enterocolitica serotype O:3 in pure culture. AB - This study examined the individual and combined effects of the selective agents normally present in Yersinia-selective agar (i.e. cefsulodin, irgasan and novobiocin) on the growth kinetics of plasmid-bearing (P+) and plasmid-cured (P-) Yersinia enterocolitica serotype O:3 at 25 and 37 degrees C. Growth studies were carried out in pure culture, and the data obtained were subjected to linear regression analysis to determine lag phase duration(s) and growth rates of the examined strains. In general, the presence of selective agents increased the duration of the lag phase at 37 degrees C, with longer lag phases noted in all cases in which two or more selective agents were present. Growth rates in CIN broth base (CIN NA) and CIN NA plus commercial supplement (SR 109) (CIN) were faster at 37 than 25 degrees C, but in some cultures of incomplete CIN NA broth with less than three supplements added, growth tended to be faster at 25 than 37 degrees C. Generally, plasmid-bearing strains grew slower than plasmid-cured strains in most media at 37 degrees C due to virulence plasmid expression retarding growth. In some instances at 37 degrees C, it was observed that the growth rates of both plasmid-bearing and plasmid-cured strains were comparable, indicating the influence of added selective agent/s negating any effects associated with virulence plasmid expression. The effects of selective agents, incubation temperature and virulence plasmid carriage on the growth kinetics of Y. enterocolitica are discussed. PMID- 10849177 TI - Evolution of a degradative bacterial consortium during the enrichment of naphtha solvent. AB - A microbial mixed culture able to degrade naphtha solvent, a model of hydrocarbon aromatic mixture, was isolated from a hydrocarbon-polluted soil. Composition of the population was monitored by phenotypic and molecular methods applied on soil DNA, on whole enrichment culture DNA, and on 85 isolated strains. Strains were characterized for their 16S rDNA restriction profiles and for their random amplified polymorphic DNA profiles. Catabolic capabilities were monitored by phenotypic traits and by PCR assays for the presence of the catabolic genes methyl mono-oxygenase ( xylA, M), catechol 2,3 dioxygenase (xylE) and toluene dioxygenase (todC1) of TOL and TOD pathways. Different haplotypes belonging to Pseudomonas putida, Ps. aureofaciens and Ps. aeruginosa were found to degrade aromatic compounds and naphtha solvent. The intrinsic catabolic activity of the microbial population of the polluted site was detected by PCR amplification of the xylE gene directly from soil DNA. PMID- 10849178 TI - A colony immunoblotting method for quantitative detection of a Bifidobacterium animalis probiotic strain in human faeces. AB - A colony immunoblotting method has been developed to allow detection of the probiotic Bifidobacterium animalis strain DN-173 010 in human faecal samples. Rabbits were immunized with heat-killed DN-173 010 bacteria resulting in the production of an antiserum highly specific for bacteria belonging to Bif. animalis species. Of the 89 strains representative of 29 different bifidobacterial species tested, only the 15 strains of the Bif. animalis species could be detected with the antiserum. In Western immunoblotting the serum reacts with a protein of 45-kDa apparent molecular weight. None of the bacteria classically encountered in human faecal samples and able to grow on non-selective Columbia blood agar (enterobacteria, Bacteroides or Lactobacillus for instance) reacted with the antiserum. Taking advantage of the high specificity of the antiserum and of the absence of Bif. animalis bacteria in faeces samples of five human volunteers, we demonstrated that strain DN-173 010 survives the intestinal transit. Being based on a combination of semiselective cultivation and colony immunoblotting techniques, the method allowed detection of the Bif. animalis strain even when it represented only one thousandth of the total bifidobacterial population. PMID- 10849179 TI - Molecular population and virulence factor analysis of Staphylococcus aureus from bovine intramammary infection. AB - Staphylococcus aureus isolates from cows in Ireland (n = 102) and the USA (n = 42) were characterized by RAPD-PCR and analysed for the production of a number of putative virulence factors. Of these strains 63 representative isolates were screened for the corresponding virulence factor genes by PCR or Southern hybridization or both. The isolates were divided into 12 distinct clonal types on the basis of their RAPD fingerprint profiles. Of the isolates, 107 (74.3%) tested positive for clumping factor in a slide agglutination test, all 24 RAPD type 7 isolates being negative for clumping factor. PCR analysis of region R, a repeat region of the clfA gene, revealed eight region-R sizes. There was a strong association between RAPD type and the clfA region-R genotype among Irish isolates. Of the RAPD type 7 isolates, 21 (87.5%) coproduced toxic shock syndrome toxin-1 (TSST-1) and staphylococcal enterotoxin C (SEC). Over 90% of isolates demonstrated haemolytic activity on sheep or rabbit red blood cells and all isolates harboured the gamma-haemolysin (hlg) locus. Of the Irish isolates, all those of RAPD type 7 were sensitive to penicillin G, whereas 86% of RAPD types 4 and 5 strains were resistant. Furthermore, RAPD types 5 and 7 were more likely to be associated with clinical mastitis whereas RAPD type 4 isolates were more often associated with a latent infection. The current study identifies some of the putative virulence factors produced by the predominant clonal types of bovine Staph. aureus that may be considered as components of a vaccine. PMID- 10849180 TI - A novel large plasmid carrying multiple beta-lactam resistance genes isolated from a Klebsiella pneumoniae strain. AB - Klebsiella pneumoniae clinical isolates were selected according to the results of antibiotic susceptibility tests. Most of them were resistant to multiple antibiotics, including ampicillin, ceftazidime, cefotaxime and aminoglycosides. Large plasmids were observed in these Kl. pneumoniae strains by pulsed-field gel electrophoresis with S1 nuclease digestion. The Kl. pneumoniae strains investigated produced one to two extrachromosomal bands with a mobility corresponding to 97 approximately 145 kbp linear DNA molecules. A 100 kbp plasmid, designated pK1, was observed in the multiply resistant strain K250. pK1 had sequences homologous to both the TEM-1 and the aphD probe which were associated with beta-lactam and aminoglycoside resistance. pK1 was transformed into Escherichia coli strain DH5alpha and was found to confer resistance to ampicillin, ceftazidime, cefotaxime and kanamycin. A 8 kbp BamHI DNA fragment of pK1 that carried the ampicillin resistance gene (minimum inhibitory concentration > 1000 microgram ml-1) was cloned into the BamHI site of pACYC184. Sequence determination showed that this cloned fragment carried a TEM-1 gene. These findings suggest that pK1 is novel in that it appears to carry genes for resistance to ampicillin, cefotaxime and ceftazidime, as well as kanamycin. PMID- 10849181 TI - Solubility and antimicrobial efficacy of protamine on Listeria monocytogenes and Escherichia coli as influenced by pH. AB - The antimicrobial efficacy of protamine on Listeria monocytogenes and Escherichia coli was evaluated at concentrations from 50 to 10 000 microgram ml-1 and pH levels from 5.5 to 8.0. The minimum inhibitory concentrations decreased with increasing pH. Protamine inhibited E. coli at all pH values while L. monocytogenes was inhibited at pH 6.5 and above. The antimicrobial efficacy of protamine decreased in the presence of negatively charged gelatine B but remained almost unchanged with addition of the positively charged gelatine A. Binding studies showed that the amount of protamine adsorbed to culture media components in tryptic soy broth and bacterial cells increased with increasing pH values. The increased efficacy of protamine at alkaline pH may be explained on the basis of an increase in electrostatic affinity for the cell surface of target cells. E. coli produced a protamine-degrading enzyme, however, was still susceptible to protamine. PMID- 10849182 TI - Heterogeneity of Lactobacillus plantarum isolates from feta cheese throughout ripening. AB - Thirty-two Lactobacillus plantarum strains isolated from Feta cheese throughout ripening were studied for their phenotypic characteristics, protein profile of cell-free extracts, enzyme profiles, plasmid profiles, proteolytic and acidifying abilities and ability to grow at low pH and in the presence of bile. Results showed that some biotechnologically important characteristics, such as acidifying and proteolytic activities, can differ between strains. In addition, different plasmid profiles suggest the presence of different Lact. plantarum strains in Feta cheese throughout ripening. The results suggest the possibility of choosing strains with specific biotechnologically interesting properties. PMID- 10849183 TI - Influence of phenolic compounds on the physiology of Oenococcus oeni from wine. AB - This study shows that the growth of Oenococcus oeni CECT 4100 in a synthetic medium is affected by phenolic compounds in different ways, depending on their type and concentration. Generally they have no effects at low concentrations, but hydroxycinnamic acids are inhibitory at high concentrations. Malolactic fermentation was stimulated in the presence of catechin and quercetin, but increasingly delayed with increasing amounts of p-coumaric acid. Gallic acid appeared to delay or inhibit the formation of acetic acid from citric acid. This could lead to a better control of malolactic fermentation and suppress the increase in volatile acidity, which is undesirable in the wine-making process. PMID- 10849184 TI - New series - evidence-based pharmacotherapy: case reports. PMID- 10849185 TI - Aminoglycoside dosing in diabetes. AB - Studies have evaluated the comparative efficacy, toxicity and costs associated with extended-interval vs. standard multiple daily dosing of aminoglycosides. In this case, an elderly man with diabetes and good renal function at baseline was switched from standard to extended-interval dosing. During the course of therapy there was evidence of decreased renal function. Pertinent literature was searched for, uncovered and critically evaluated to determine if and what evidence supports using extended dosing of aminoglycosides in this population. No data were found specifically evaluating the different dosing strategies in diabetic patients. However, there were many trials and several meta-analysis located that compared the two dosing strategies, most suggesting at least a cost advantage and possibly less toxicity with extended-interval dosing. Further information is needed to determine whether there is a differential risk for toxicity between these dosing regimens in patient with diabetes. PMID- 10849186 TI - Treating mammalian bite wounds. AB - The incidence of dog, cat and human bites has been increasing steadily and represents an important cause of morbidity and mortality in the United States. Approximately half of all Americans will suffer a bite wound during their lifetime, and the annual medical costs of managing these injuries has been estimated to be over $100 million. Possible complications may include disfigurement, dismemberment and infection. Effective management requires rapid medical evaluation and may necessitate surgical intervention and prophylactic antibiotic therapy. As bite wounds are microbiologically diverse and most often polymicrobial in nature, selection of an appropriate antibiotic regimen requires knowledge of common pathogens. Close clinical follow-up is recommended to minimize the risk of late complications. PMID- 10849187 TI - Clostridium difficile-associated diarrhoea in hospitalised patients. AB - OBJECTIVE: The aim of the present study was to evaluate the incidence, risk factors and cost implications of Clostridium difficile-associated diarrhoea (CDAD) in hospitalized adult patients. METHODS: Eighty-seven hospitalized adult patients, positively identified as having CDAD, were reviewed retrospectively to determine the risk factors and cost implications of CDAD. RESULTS: The clinical manifestations, in addition to diarrhoea, included elevated temperature (= 37.8 degrees C; 42.5%), abdominal pain (63. 2%) and leucocytosis (=12 x 109 cells/l; 52.9%). Eight patients underwent endoscopy, and pseudomembranous colitis was confirmed in all of these patients. Nine patients died during their hospital stay. Cefotaxime and cefuroxime were the agents most commonly associated with CDAD. There was a significant difference (P < 0.001) between the sex distribution of CDAD patients and adult hospital patients (69% of CDAD patients were female vs. 52% of general adult hospital population). Significantly (P < 0.001) more patients with CDAD were admitted from the nursing home (NH) setting. The mean age of patients with CDAD admitted from NHs (n = 19) was older than those cases admitted from the community (n = 68) by 14 years (P < 0.001). The length of hospital stay was significantly (P < 0.001) longer for patients with CDAD (16.9 vs. 3.89 days). No differences (P = 0.306) were found in the response times for CDAD patients treated with either oral metronidazole (n = 39) or oral vancomycin (n = 48). The mean response time was, however, significantly longer in the CDAD patients admitted from NHs (4.2 days) compared with those admitted from the community (2.5 days), although the former patients were older and had significantly more comorbidity (P < 0.001). The mean cost per one treated-case of CDAD (bed, laboratory requests and treatment therapy) was calculated as pound2860. CONCLUSION: Patients admitted from NHs are at increased risk of development of CDAD; receiving cefotaxime or cefuroxime axetil (oral form), being elderly and being female are risk factors for the development of CDAD. Treatment of CDAD with oral metronidazole or oral vancomycin gives rise to similar response times and efficacy. PMID- 10849188 TI - Changes in the management of acute myocardial infarction in Southern Tasmania. AB - BACKGROUND: In recent years, the management of acute myocardial infarction (AMI) has been the subject of many clinical trials. These studies have clearly established the value of using pharmacological agents, including aspirin, beta blockers, thrombolytics and angiotensin converting enzyme (ACE) inhibitors. There have been suggestions, however, that practice has been slow to change in light of the findings of these trials. AIM: To review cases of AMI at the major teaching hospital in Tasmania, Australia, to determine whether the recommendations from the results of the trials had been translated into local clinical practice, and to examine temporal changes in drug usage and clinical outcomes. METHODS: A retrospective review of the medical records of patients admitted to the hospital with an AMI during 1996 and for the first four months of 1998 was performed. An extensive range of demographic and clinical variables was recorded, and differences between the 1996 and 1998 patients and between recipients and non recipients of the different pharmacological agents were statistically evaluated. RESULTS: The patients had a mean age of 65.9 +/- 12.3 years in 1996 (n = 205) and 66.8 +/- 12.3 years in 1998 (n = 71), with males accounting for 64.4% of cases in 1996 and 64.8% of cases in 1998. There were no significant demographic or medical history differences between the two groups. The median time of presentation after the onset of chest pain was 3.5 h in 1996 and 4 h in 1998. The rates of use of major therapeutic interventions post-AMI for 1996 and 1998, respectively, were: aspirin (89.1%, 90.3%), streptokinase (18.5%, 9. 9%), r-tPA (14.1%, 21.1%), intravenous beta-blockers (11.2%, 7.0%), oral beta-blockers (67.2%, 49.3%; P < 0.01), ACE inhibitors (44.4%, 59.2%; P < 0.05), intravenous nitrate (94.1%, 91.6%), oral nitrate (22.9%, 26.8%), calcium channel antagonists (19.5%, 35.2%; P < 0.05), cholesterol lowering agents (26.3%, 40.9%; P < 0.05), antiarrhythmics (21.5%, 25.4%) and warfarin (8.3%, 9.9%). Patients who received therapy with each of aspirin, r-tPA, intravenous beta-blockers, oral beta-blockers, intravenous nitrate and cholesterol lowering agents were significantly younger than the non recipients (all P < 0.01), while patients treated with ACE inhibitors and antiarrhythmics were significantly older than the non-recipients (both P < 0.001). Non-recipients of thrombolytics presented to hospital significantly later, on average, than recipients. The hospital mortality rate was 15.1% in 1996 and 12.7% in 1998, and adverse drug reactions occurred in 21.5% of patients in 1996 and 15.5% in 1998. CONCLUSION: Although there have been substantial increases in the use of ACE inhibitors and cholesterol lowering agents post-AMI in recent years, reductions in the use of thrombolytics and beta-blockers and their possible underuse in the elderly are of concern and warrant intervention. PMID- 10849189 TI - Motivating general practitioners to change their prescribing: the incentive of working together. AB - OBJECTIVE: To determine the extent to which GPs were motivated to change their prescribing upon joining a Primary Care Commissioning Group (PCCG) and how effective certain interventions planned by the PCCG might be as a means to change prescribing. To define the characteristics of GPs less motivated to change their prescribing. DESIGN: A cross-sectional survey of participating general practitioners linked with current prescribing information derived from PACT data. SETTING: General practice covering a geographical locality within inner-city south London. SUBJECTS: All 72 general practitioners who had joined a GP Commissioning Group. MAIN OUTCOME MEASURES: questionnaire responses. RESULTS: 93% of GPs entering the GP Commissioning Group expected their prescribing to change but none expected substantial change. There was no difference between fundholders, singlehanders nor training practices in their expectation of change. GPs in practices with the lowest quality prescribing, as measured by a quality index, were least likely to expect change (Spearman's r = 0.25, P = 0.04). Those in practices with higher prescribing costs were not more likely to expect their prescribing to change, whereas expensive prescribers who were unaware of their practices' prescribing costs were associated with a reduced expectation of prescribing change (P = 0.05). Educational interventions were thought to be the most effective means by which prescribing could be changed, whereas formularies and financial factors were perceived as weaker influences. CONCLUSION: Acceptance of a cash-limited prescribing budget by GPs is accompanied by the expectation of personal prescribing change. The motivation to change prescribing may be related to a strongly developed collectivist perspective amongst GPs who are prepared to consider the prescribing implications for their fellow GPs. It is ironic that those with the least expectation of change should have the lowest quality prescribing, or be unaware of their high cost prescribing. Engendering greater commitment to the professional group may be one way of changing their prescribing. PMID- 10849190 TI - Drug utilization pattern of antiepileptic drugs and traditional Chinese medicines in a general hospital in Taiwan - a pharmaco-epidemiologic study. AB - OBJECTIVE: Studies on antiepileptic drug utilization are important for the optimization of drug therapy and drug control. The present study was to evaluate the drug utilization pattern of standard antiepileptic drugs and traditional Chinese medicines (TCMs) in the treatment of different types of epilepsy in a general hospital in Taiwan. METHOD: The epileptic patients under antiepileptic drug treatment at Veterans General Hospital-Taipei were considered in the analysis. Current diagnosis was obtained by the neurologist in charge of the patient. All patients were interviewed by standard questionnaire designed to provide specific information on the types of antiepileptic drugs and details of their use. The questionnaire also sought to determine whether TCMs were used, and whether patients were using TCMs in combination with hospital standard treatment. The results were analysed by descriptive statistics. RESULTS: 729 patients with epilepsy definitely diagnosed were analysed in the study. 445 patients (61.04%) were prescribed with one antiepileptic drug. Combinations of two antiepileptic drugs were prescribed to 261 patients (35.80%), and combinations of three or more antiepileptic drugs to 23 patients (3.16%). A total of 1039 antiepileptic drugs was prescribed, corresponding to an average 1.42 drugs per patient. The most frequently prescribed antiepileptic drug was carbamazepine (56.93%), followed by phenytoin (31.96%), valproate (30.73%) and clonazepam (14.13%). Among the 729 epileptic patients, 83.68% used standard antiepileptic drugs alone, 16.32% used antiepileptic drugs in combination with TCMs. CONCLUSION: Monotherapy is the type of therapy most frequently used in all types of seizures. The selection of antiepileptic drugs is based on efficacy for specific seizure types and epileptic syndromes. The most frequently prescribed antiepileptic drug was carbamazepine, followed by phenytoin, valproate and clonazepam. As some of the patients used TCMs for treatment of epilepsy even when scientific medicine has been provided, further studies on the possible interactions between TCMs and antiepileptic drugs are in progress. PMID- 10849191 TI - Dipyridamole in the treatment of schizophrenia: adenosine-dopamine receptor interactions. AB - OBJECTIVE: There is growing interest in investigating the adenosine-dopamine interaction in the ventral striatum. Adenosine plays a role opposite to dopamine in the striatum and adenosine antagonists, like caffeine, produce similar effects to increased dopaminergic neurotransmission in the striatum. In particular, a strong antagonistic interaction between adenosine A2A and dopamine D2 receptors takes place in the striopallidal GABAergic neurones. Therefore, adenosine agonists or uptake inhibitors provide a potential new treatment for schizophrenia. We undertook a pilot trial to investigate whether the combination of haloperidol with dipyridamole, an uptake inhibitor of adenosine, was more effective than haloperidol alone. METHODS: Thirty patients who met the DSM IV criteria for schizophrenia completed the study. Patients were allocated in a random fashion, 16 to haloperidol 20 mg/day plus dipyridamole 75 mg/day and 14 to haloperidol 20 mg/day plus placebo. RESULTS: Although both protocols significantly decreased the score of the positive, negative and general psychopathological symptoms over the trial period, the combination of haloperidol and dipyridamole was significantly better than haloperidol alone in decreasing positive and general psychopathology symptoms as well as PANSS total scores. CONCLUSION: Dipyridamole may be of therapeutic benefit in treating schizophrenia in combination with neuroleptics. However, a larger study to confirm our results is warranted. PMID- 10849192 TI - Systematic overview of lithium treatment in acute mania. AB - OBJECTIVE: To evaluate the efficacy of lithium in the treatment of acute mania. METHOD: Systematic overview of the literature and meta-analysis of randomised controlled trials. Estimation of (i) the differences in the reduction in mania severity scores, and (ii) the ratio and difference in improvement response rates. RESULTS: A total of 658 patients from 12 trials were included. Treatment periods ranged from 3 to 4 weeks. The response rate ratio for lithium against placebo was 1.95 (95%CI 1.17-3.23). The mean number needed to treat was five (95%CI 3-20). Patients were twice as likely to obtain remission with lithium than with chlorpromazine (rate ratio = 1.96, 95%CI 1.02-3.77). The mean number needed to treat was four (95%CI 3-9). Neither carbamazepine nor valproate was more effective than lithium. The response rate ratios were 1.01 (95%CI 0.54-1.88) for lithium compared to carbamazepine and 1.22 (95%CI 0.91-1.64) for lithium against valproate. Haloperidol was no better than lithium on the basis of improvement based on assessment of global severity. The differences in effects between lithium and risperidone were -2.79 (95%CI -4.22 to -1.36) in favour of risperidone with respect to symptom severity improvement and -0.76 (95%CI -1.11 to -0.41) on the basis of reduction in global severity of disease. Symptom and global severity was as well controlled with lithium as with verapamil. Lithium caused more side-effects than placebo and verapamil, but no more than carbamazepine or valproate. CONCLUSION: The clinical trial evidence suggests that lithium should remain the first line treatment for acute mania. PMID- 10849193 TI - Introduction PMID- 10849194 TI - Measurement of glutathione levels in intact roots of Arabidopsis. AB - Levels of glutathione were measured for different cell types in roots of intact Arabidopsis seedlings after labelling with monochlorobimane to give fluorescent glutathione S-bimane (GSB) and imaging using confocal laser scanning microscopy with excitation at 442 nm. Labelling increased to a plateau in most cell types after about 15-20 min and the GSB accumulated rapidly in the vacuole. Formation of GSB in the cytoplasm was not affected by treatment with sodium azide; however, vacuolar transport of GSB was substantially inhibited under these conditions. We infer that vacuolar sequestration was mediated by a tonoplast glutathione S conjugate pump. Quantitative estimates of the cytoplasmic glutathione concentration involved correction for the loss in fluorescence signal with depth into the specimen using an empirically determined model derived in situ from a permeabilized root. Correction for the dilution experienced on transport into the vacuole also required an estimate of the amount of cytoplasm present in each cell type. This was achieved in two stages: first, the levels of protein were mapped after fixation, permeabilization and labelling with fluroescein isothiocyanate. Second, the corresponding cytoplasmic volume was determined as 40% for epidermal cells in the elongation zone by manual segmentation of the cytoplasm in serial optical sections. Values of relative cytoplasmic volume for other cells were extrapolated in proportion to their protein content. Using this approach, cytoplasmic glutathione concentrations were found to be 2-3 mM in most cell types. There was a marked difference between the central cells and the neighbouring, rapidly dividing initials, and between the columella cells and the outermost cells of the root cap. In the latter case, the difference was equalized in the presence of azide. This might indicate that additional cell-cell movement and preferential sequestration of GSB can occur during the detoxification process in an intact system. PMID- 10849195 TI - Direct measurement of glutathione in epidermal cells of intact Arabidopsis roots by two-photon laser scanning microscopy. AB - Two-photon laser scanning microscopy (TPLSM) was used to directly measure glutathione (GSH) as its fluorescent glutathione S-bimane conjugate (GSB) in developing root hair cells (trichoblasts) and non-root hair cells (atrichoblasts) of intact Arabidopsis roots. In comparison to confocal microscopy, TPLSM showed more detail deep within the tissue with less signal attenuation. The total level of GSB labelling reached a plateau after 60 min in both trichoblasts and atrichoblasts, reflecting depletion of GSH. GSB was formed initially in the cytoplasm and was subsequently transported into the vacuole. The volume ratio of vacuole to cytoplasm was determined using the Cavalieri estimator of volume and used to calculate the amount of GSB per volume of cytoplasm in each cell type. At the end of the time-course the cytoplasmic concentration of GSB was 2.7 +/- 0.5 mM (n = 5) in trichoblasts and 5.5 +/- 0.8 mM (n = 5) in atrichoblasts. In trichoblasts this value represents the initial concentration of GSH in the cytoplasm. Labelling of roots with monochlorobimane (MCB) on ice led to the formation of GSB in the cytoplasm, but prevented vacuolar sequestration. After washing prelabelled roots and transfer to room temperature, vacuolar transport resumed. Although no free MCB was present the total amount of GSB in atrichoblasts increased further, indicating that the higher values recorded in the atrichoblasts might reflect additional symplastic transport and sequestration of GSB from neighbouring cells. PMID- 10849196 TI - Micromanipulation by laser microbeam and optical tweezers: from plant cells to single molecules. AB - Complete manipulation by laser light allows precise and gentle treatment of plant cells, subcellular structures, and even individual DNA molecules. Recently, affordable lasers have become available for the construction of microbeams as well as for optical tweezers. This may generate new interest in these tools for plant biologists. Early experiments, reviewed in this journal, showed that laser supported microinjection of material into plant cells or tissues circumvents mechanical problems encountered in microinjection by fragile glass capillaries. Plant protoplasts could be fused with each other when under microscopical observation, and it was no major problem to generate a triple or quadruple fusion product. In the present paper we review experiments where membrane material was prepared from root hair tips and microgravity was simulated in algae. As many plant cells are transparent, it is possible to work inside living, intact cells. New experiments show that it is possible to release by optical micromanipulation, with high spatial resolutions, intracellular calcium from caged compounds and to study calcium oscillations. An example for avian cardiac tissue is given, but the technique is also suitable for plant cell research. As a more technical tool, optical tweezers can be used to spatially fix subcellular structures otherwise moving inside a cell and thus make them available for investigation with a confocal microscope even when the time for image formation is extended (for example at low fluorescence emission). A molecular biological example is the handling of chromosomes and isolated individual DNA molecules by laser microtools. For example, chromosomes can be cut along complex trajectories, not only perpendicular to their long axis. Single DNA molecules are cut by the laser microbeam and, after coupling such a molecule to a polystrene microbead, are handled in complex geometries. Here, the individual DNA molecules are made visible with a conventional fluorescence microscope by fluorescent dyes such as SYBRGreen. The cutting of a single DNA molecule by molecules of the restriction endonuclease EcoRI can be observed directly, i.e. a type of single molecule restriction analysis is possible. Finally, mechanical properties of individual DNA molecules can be observed directly. PMID- 10849197 TI - The behaviour of the plasma membrane during plasmolysis: a study by UV microscopy. AB - A high resolution ultraviolet (UV) bright-field microscope was used to analyse the formation of Hechtian strands and the Hechtian reticulation that remain attached to the cell wall after plasmolysis and deplasmolysis of onion inner epidermal cells. In real time video images, UV microscopy allowed a detailed investigation of the dynamic behaviour of the plasma membrane during the processes of osmotic water loss and uptake. Furthermore, the role of cytoskeletal elements as possible linkers of the plasma membrane to the cell wall was probed by application of cytoskeletal drugs during plasmolysis. Microtubules were depolymerized in oryzalin, and latrunculin B was used to destabilize actin microfilaments. The results showed no visible changes in the formation of the Hechtian reticulation or strands. Plasmolysis forms appeared to be normal, indicating stong membrane-to-wall attachments independent of cytoskeletal elements. During re-expansion of the protoplast in deplasmolysis, the plasma membrane incorporated Hechtian strands and subprotoplasts, fused with the Hechtian reticulation and finally realigned at the cell wall. PMID- 10849198 TI - The nucleus: a highly organized but dynamic structure. AB - The nucleus in plants and animals is a highly structured organelle containing several well-defined subregions or suborganelles. These include the nucleolus, interphase chromosome territories and coiled bodies. We have visualized transcription sites in plants at both light- and electron-microscopy level by the incorporation of BrUTP. In the nucleolus many dispersed foci are revealed within the dense fibrillar component, each of which probably corresponds to a single gene copy. In the nucleoplasm there are also many dispersed foci of transcription, but not enough to correspond to one site per transcribed gene. We have shown that in wheat, and probably many other plant species, interphase chromosome territories are organized in a very regular way, with all the chromosomes in the Rabl configuration, all the centromeres clustered at the nuclear membrane and all the telomeres located at the nuclear membrane on the opposite side of the nucleus. However, despite this regular, polarized structure, there is no sign of polarization of transcription sites, or of any preferred location for them with respect to chromosome territorial boundaries. The nucleus is also highly dynamic. As an example, we have shown by the use of a green fluorescent protein fusion to the spliceosomal protein U2B" that coiled bodies move and coalesce within the nucleus, and may act as transport structures within the nucleus and nucleolus. PMID- 10849199 TI - Cell wall secretion in the green alga micrasterias. AB - The monoclonal antibodies JIM 1 against non-arabinogalactan epitopes, JIM 5 and JIM 7 recognizing unesterified and methyl-esterified pectins, and JIM 8 specific for arabinogalactan proteins (AGPs) are used for investigating different stages of cell wall formation in high pressure frozen and freeze substituted Micrasterias cells by means of immunoelectron microscopy. The results show that the septum-forming vesicles and the septum wall consist mainly of methyl esterified pectins which become only partly de-esterified in the septum wall. Arabinogalactan proteins appear at the septum rim at the end of septum growth and are main constituents of the primary cell wall, together with esterified pectins. Only the outermost layer of the primary cell wall is labelled by JIM 5, indicating the presence of unesterified pectins. AGPs, non-AGP epitopes indicated by JIM 1, and pectins are transported together in the contents of the primary wall, forming 'dark vesicles' from the site of their production at the dictyosomes to the plasma membrane. Labelling of exclusively the plasma membrane of the non-growing semicell by JIM 1 and JIM 8 points towards a regulatory mechanism of membrane glycoproteins for vesicle fusion. The secondary cell wall of Micrasterias is not labelled by any of the antibodies used. JIM 4, JIM 15, JIM 84 and MAC 207 do not produce any specific staining in Micrasterias. PMID- 10849200 TI - The cytoskeleton in plant and fungal cell tip growth. AB - Tip-growing cells have a particular lifestyle that is characterized by the following features: (1) the cells grow in one direction, forming a cylindrical tube; (2) tip-growing cells are able to penetrate their growth environment, thus having to withstand considerable external forces; (3) the growth velocity of tip growing cells is among the fastest in biological systems. Tip-growing cells therefore appear to be a system well suited to investigating growth processes. The cytoskeleton plays an important role in cell growth in general, which is why tip-growing cells provide an excellent model system for studying this aspect. The cytoskeletal system comprises structural elements, such as actin filaments and microtubules, as well as proteins that link these elements, control their configuration or are responsible for transport processes using the structural elements as tracks. Common aspects as well as differences in configuration and function of the cytoskeleton in various types of tip-growing cells reveal the general principles that govern the relationship between the cytoskeleton and cell growth. PMID- 10849201 TI - Confocal microscopy of FM4-64 as a tool for analysing endocytosis and vesicle trafficking in living fungal hyphae. AB - Confocal microscopy of amphiphilic styryl dyes has been used to investigate endocytosis and vesicle trafficking in living fungal hyphae. Hyphae were treated with FM4-64, FM1-43 or TMA-DPH, three of the most commonly used membrane selective dyes reported as markers of endocytosis. All three dyes were rapidly internalized within hyphae. FM4-64 was found best for imaging the dynamic changes in size, morphology and position of the apical vesicle cluster within growing hyphal tips because of its staining pattern, greater photostability and low cytotoxicity. FM4-64 was taken up into both the apical and subapical compartments of living hyphae in a time-dependent manner. The pattern of stain distribution was broadly similar in a range of fungal species tested (Aspergillus nidulans, Botrytis cinerea, Magnaporthe grisea, Neurospora crassa, Phycomyces blakesleeanus, Puccinia graminis, Rhizoctonia solani, Sclerotinia sclerotiorum and Trichoderma viride). With time, FM4-64 was internalized from the plasma membrane appearing in structures corresponding to putative endosomes, the apical vesicle cluster, the vacuolar membrane and mitochondria. These observations are consistent with dye internalization by endocytosis. A speculative model of the vesicle trafficking network within growing hyphae is presented. PMID- 10849202 TI - Cytoplasmic cleavage in living zoosporangia of Allomyces macrogynus. AB - We have used the vital fluorescent dye, FM4-64, as a marker of membrane development during zoospore formation in living zoosporangia of Allomyces macrogynus. Membrane development was visualized and documented using standard epifluorescence and laser scanning confocal microscopy. Video-enhanced light microscopy and transmission electron microscopy, using cryopreparation methods, were also employed in this study. In the first 10-12 min after the induction of zoospore formation, only the plasma membrane labelled with FM4-64. During this time, nuclei were strictly located in the cortical cytoplasm with their associated centrosomes positioned immediately adjacent to the plasma membrane (Lowry & Roberson, 1997). Between 12 and 20 min post-induction, increased fluorescence appeared along regions of the plasma membrane adjacent to the nuclei. From these sites, membranes (i.e. cleavage elements) extended laterally within the cortex and then, in conjunction with nuclear migration, rapidly elongated into the sporangial cytoplasm. By 25-35 min post-induction, cleavage elements had ramified throughout the cytoplasm forming a complex, interconnected membranous network. Transmission electron microscopy revealed that cleavage elements were paired membrane sheets with a lumen consisting of an electron opaque, granular matrix. Cleavage elements developed into a highly ordered network by 35-40 min post-induction, which fully delimited zoospore initials into polyhedral-shaped cells. Zoospore discharge occurred between 40 and 50 min post induction. Our results have shown that cleavage elements undergo four stages of development during zoospore formation in A. macrogynus: (i) development of membrane initials, (ii) cortical extension, (iii) cytoplasmic elongation and ramification and (iv) zoospore initial delimitation. PMID- 10849203 TI - Sex differences in the brain. PMID- 10849204 TI - Androgenic regulation of steroid hormone receptor mRNAs in the brain of whiptail lizards. AB - Sex and species differences in androgenic regulation of steroid hormone receptor mRNAs were examined in the diencephalon of two species of whiptail lizards: Cnemidophorus inornatus is a sexual species and the direct evolutionary ancestor to Cnemidophorus uniparens, an all-female parthenogenetic species. Lizards were gonadectomized and treated with different doses of either aromatizable testosterone or nonaromatizable dihydrotestosterone. The relative abundances of androgen-, oestrogen-, and progesterone-receptor mRNAs were compared in various nuclei following in situ hybridization with homologous riboprobes. A diversity of patterns in androgenic regulation was observed, with effects differing according to brain region, the steroid-receptor mRNA being considered and, in some cases, between androgens. In the ancestral sexual species, intact males had lower androgen-receptor mRNA abundances than castrated, blank-implanted males in the medial preoptic area. Testosterone significantly decreased androgen-receptor mRNA abundance in the medial preoptic area of castrated males. Males had higher androgen-receptor mRNA levels in the preoptic area than females generally and neither the sexual or parthenogenetic females showed a decrease in androgen receptor mRNA with androgen treatment. Both testosterone and dihydrotestosterone increased oestrogen-receptor mRNA abundance in the ventromedial hypothalamus of C. inornatus, but no sex differences in this effect were observed. Gonadectomy decreased, whereas androgen treatment increased, progesterone-receptor mRNA abundance in the ventromedial hypothalamus. There was a sex difference in this response to androgen in the sexual species, with males having greater amounts than females in this brain area. The parthenogenetic species exhibited a similar pattern to females of the sexual species, but the levels were higher overall, possibly because Cnemidophorus uniparens is triploid. The periventricular preoptic area showed a different pattern, with testosterone treatment increasing progesterone-receptor mRNA abundance in both sexes of the sexual species and in the parthenogenetic species, while dihydrotestosterone did not. The diversity of patterns in androgen effects indicates that gonadal sex, aromatization of androgen, and perhaps gene dosage all influence the expression of steroid receptor mRNAs in the lizard brain. PMID- 10849205 TI - Expression of proadrenomedullin derived peptides in the mammalian pituitary: co localization of follicle stimulating hormone and proadrenomedullin N-20 terminal peptide-like peptide in the same secretory granules of the gonadotropes. AB - Expression of proadrenomedullin-derived peptides in the rat, cow and human pituitary was studied by a variety of techniques. Immunocytochemical detection showed a widespread expression of adrenomedullin peptide in the adenohypophysis and the neural lobe, with low expression in the intermediate pituitary. Proadrenomedullin N-20 terminal peptide (PAMP)-immunoreactivity was also present in the anterior pituitary but showed a more marked heterogeneous distribution, with cells going from very strong to negative immunostaining. Lower levels of PAMP were found in the neural lobe. Interestingly, the distribution of adrenomedullin and PAMP immunoreactivity in the anterior pituitary did not completely overlap. In the present study, we concentrated our efforts to determine which cell type of the adenohypophysis expresses PAMP. Paraffin and semithin serial sections immunostained for PAMP and the classical pituitary hormones revealed that a subpopulation of the gonadotropes expresses high levels of PAMP-immunoreactive material. Ultrastructural analysis clearly showed PAMP immunoreactivity in the follicle stimulating hormone (FSH)-containing large secretory granules of the gonadotropes, suggesting simultaneous secretion of PAMP and FSH by this cell type. Three mouse adenohypophysis-derived cell lines (AtT20, GH3, and alphaT3-1 derived from corticotropes, lacto/somatotropes and gonadotropes, respectively) were also analysed and showed expression of both proadrenomedullin-derived peptides and their mRNA. Functional studies in these three cell lines showed that neither adrenomedullin nor PAMP was able to stimulate cAMP production in our experimental conditions. Taken together, our results support that proadrenomedullin derived peptides are expressed in the pituitary in cell-specific and not overlapping patterns, that could be explained by differences in postranslational processing. Our data showing costorage of PAMP and FSH in the same secretory granules open a way by which PAMP could be involved in the control of reproductive physiology in a coordinated manner with FSH. PMID- 10849206 TI - Effect of season on neuropeptide Y and galanin within the hypothalamus of the ewe in relation to plasma luteinizing hormone concentrations and the breeding season: an immunohistochemical analysis. AB - Within the hypothalamus, neurones that express neuropeptide Y (NPY) and galanin have been implicated in the regulation of gonadotropin-releasing hormone (GnRH) and gonadotropin secretion. We aimed to determine the extent to which the expression of these two neuronal systems is linked to the seasonal reproductive cycle, and the effect of chronic oestrogen treatment. Immunohistochemical analysis was used to examine changes between the breeding season and anestrus in ovariectomized (OVX) ewes with or without oestrogen treatment (s.c. implants for 2 weeks). Serial blood sampling established plasma luteinizing hormone (LH) profiles, and the ewes were subsequently killed and the brains perfused for immunohistochemistry. In OVX ewes, the amplitude of LH pulses was greater in the nonbreeding season than in the breeding season. Oestrogen treatment caused a marked reduction in plasma LH concentrations during anestrus, but not in the breeding season. The number of cells in the arcuate nucleus/median eminence region (ARC-ME) that stained for NPY was lower in ewes killed in anestrus (September) than in ewes killed in the breeding season (May), but there was no seasonal change in the number of galanin-stained cells. Within season, oestrogen treatment did not affect NPY- or galanin-cell number. There was no effect of season or oestrogen on the area of varicose fibres/terminals for either peptide in the ARC-ME, but galanin immunostaining was more intense during the breeding season. We conclude that the amount of NPY in cell bodies of the ARC-ME is lower in ewes in the nonbreeding season; this could reflect a steroid-independent effect of photoperiod. We also conclude that the long-term negative-feedback effect of oestrogen on GnRH/LH secretion does not appear to be mediated by NPY- or galanin-containing neurones in the ewe. PMID- 10849207 TI - Co-expression of melatonin (MEL1a) receptor and arginine vasopressin mRNAs in the Siberian hamster suprachiasmatic nucleus. AB - Durational melatonin signals, cued by the photoperiod and generated by the pineal gland, are processed in the brain to induce seasonally appropriate physiological and behavioural adaptations. The melatonin receptor subtype MEL1a (also known as mt1) appears to regulate seasonal responses. Single label in situ hybridization for MEL1a receptor mRNA revealed labelled cells in several brain regions of Siberian hamsters, including the suprachiasmatic nucleus, the paraventricular nucleus of the thalamus, and the reuniens nucleus of the thalamus. To characterize suprachiasmatic nucleus cells containing MEL1a receptor mRNA, we used 35S-labelled cRNA probes for MEL1a receptor mRNA in combination with digoxigenin-labelled cRNA probes for vasopressin, somatostatin, or orphan retinoid Z receptor beta (RZRbeta; a putative nuclear melatonin receptor). Cells in the suprachiasmatic nucleus that contained MEL1a receptor mRNA also contained mRNAs for vasopressin and RZRbeta, but not for somatostatin. These data suggest that suprachiasmatic nucleus vasopressin cells may respond to melatonin signals, raising the possibility that suprachiasmatic nucleus vasopressin output mediates some of the effects of melatonin on seasonal or circadian responses. PMID- 10849208 TI - Neural connections of hypothalamic neuroendocrine nuclei in the rat. AB - The secretion of many hormones, including oxytocin, vasopressin and growth hormone, is not constant but shows a day-night rhythm. The suprachiasmatic nucleus (SCN) is thought to generate most mammalian biological rhythms and previous studies have reported suprachiasmatic efferents to the paraventricular nucleus (PVN) and the supraoptic nucleus (SON). We used in vivo extracellular electrophysiological techniques to show that the SCN also sends direct and indirect neural projections to the arcuate nucleus (ARC). This projection consisted of both excitatory and inhibitory components and may contribute to the entrainment of the rhythm in growth hormone secretion to the day-night cycle. Some SCN neurones appear to project to both the SON and the ARC. The SCN in turn receives excitatory and inhibitory inputs from the ARC and the peri-nuclear zone of the SON (peri-SON), which may provide feedback information, as well as allowing nonphotic entrainment of the SCN, for example, in response to feeding. Our data thus suggest extensive two-way connections between the SCN and its target nuclei which may contribute to the generation of day-night neuroendocrine rhythms. They also suggest the existence of indirect retinal projections to the ARC and PVN. We further investigated the retinal projection to the SCN. We were unable to demonstrate a significant difference in retinal input to those suprachiasmatic cells which had efferent projections to particular hypothalamic targets (SON and/or ARC), and those which did not. PMID- 10849209 TI - B219/OB-R 5'-UTR and leptin receptor gene-related protein gene expression in mouse brain and placenta: tissue-specific leptin receptor promoter activity. AB - Leptin receptor (OB-R) splice variants either encode proteins with different 3' cytoplasmic domains or have different 5' untranslated regions (UTR), indicative of dual promoters. The B219/OB-R promoter transcribes only OB-R transcripts, whereas the OB-R/GRP promoter initiates transcription of both OB-R and another protein of unknown function, called the leptin receptor gene-related protein (OB RGRP). We compared expression of B219/OB-R 5'-UTR and OB-RGRP mRNAs by in situ hybridization. We thus assessed, by inference, the contributions of the two promoters to the leptin receptor transcript pool, in murine brain or in placenta, a tissue with abundant leptin receptor mRNA. Expression of B219/OB-R 5'-UTR mRNA (and thus by inference B219/OB-R promoter activity) in brain was similar in both distribution and relative intensity to OB-R mRNA. OB-RGRP mRNA (and thus by inference OB-R/GRP promoter activity) was widely distributed in murine brain, with elevated expression in the hypothalamic regions that express the leptin receptor mRNA, including the paraventricular nucleus. B219/OB-R 5'-UTR mRNA, but not OB-RGRP mRNA, was upregulated in hypothalamus of obese ob/ob mice. In placenta, B219/OB-R 5'-UTR mRNA was restricted to the maternal interface, and transcription of both long and short leptin receptor splice variants in the main body of the tissue thus proceeds via the OB-R/GRP promoter, strongly indicative of tissue-specific promoter usage. PMID- 10849210 TI - Distribution of aromatase mRNA in the ram hypothalamus: an in situ hybridization study. AB - The regional distribution of neurones expressing aromatase mRNA in the ram hypothalamus was examined by in situ hybridization using 33P-labelled cRNA probes. The highest amounts of hybridization signal were observed in the central part of the medial preoptic nucleus and posterior medial part of the bed nucleus of the stria terminalis. Moderate amounts of hybridization signal were observed in the anteroventral periventricular preoptic nucleus, medial preoptic nucleus and a broad band extending between the medial preoptic nucleus and bed nucleus of the stria terminalis. Low levels of hybridization signal were observed in the organum vasculosum of the lamina terminalis, anterior part of the medial preoptic nucleus, and central part of the ventromedial nucleus of the hypothalamus. The presence of aromatase mRNA within neurones of the steroid-sensitive hypothalamic circuit supports a role for aromatization in the mechanism of testosterone action on reproductive function in male sheep. The distribution of aromatase mRNA in the ovine hypothalamus was similar to that described for other vertebrate species, suggesting a high degree of functional conservation across species. PMID- 10849211 TI - Evidence that members of the TGFbeta superfamily play a role in regulation of the GnRH neuroendocrine axis: expression of a type I serine-threonine kinase receptor for TGRbeta and activin in GnRH neurones and hypothalamic areas of the female rat. AB - The present study was designed to determine whether transforming growth factor (TGF)beta and/or activin participate in the regulation of the gonadotropin releasing hormone (GnRH) neuroendocrine axis in vivo. Single-label in situ hybridization histochemistry was used to determine the anatomical distribution of a TGFbeta and activin type I receptor (B1) mRNA, in the adult female rat hypothalamic areas that are known to be important sites for the regulation of reproduction. Dual-label in situ hybridization histochemistry was performed to determine whether B1 mRNA was expressed in GnRH neurones. The results of these studies revealed an extensive distribution of B1 mRNA in the hypothalamic regions, including diagonal bands of Broca, preoptic area, arcuate nucleus and median eminence. In the median eminence, B1 mRNA was detected in tanycytes and in the endothelial cells of the pituitary portal blood capillaries. Dual-label in situ hybridization histochemistry showed that 31+/-5% of GnRH neurones expressed B1 mRNA, thus providing evidence that TGFbeta and/or activin can act directly on GnRH neurones to modulate their activity. Taken together, these data provide morphological arguments in favour of a participation of TGFbeta and/or activin in the regulation of reproduction at the hypothalamic level. PMID- 10849212 TI - Cyclical expression of egr-1/NGFI-A in the rat anterior pituitary: a molecular signal for ovulation? AB - Molecular genetic studies of egr-1/NGFI-A have recently established a key role for this immediate early gene in anterior pituitary development, and particularly in transcriptional regulation of the LH beta-subunit gene. These studies, using null mutant mice, addressed the developmental aspects of gonadotrophin gene regulation by egr-1 and, because of the limitations of this approach, did not address the role of egr-1 in adult reproductive physiology. Using the rat, we have now investigated the expression of egr-1 during the female reproductive cycle, and have found evidence of cyclical changes in anterior pituitary egr-1 expression, at mRNA, protein, and DNA binding activity levels. Specifically, we have shown that egr-1 mRNA is significantly elevated during proestrous, and conversely suppressed on the subsequent day of oestrous. We have also demonstrated significantly raised levels of an approximately 70 kDa Egr-1 immunoreactive protein band during the night of proestrous, together with markedly raised levels of a Egr-1 consensus sequence DNA binding complex. These studies are indicative of both direct egr-1 gene regulation by oestrogen, and also regulation by GnRH. Finally, we have sequenced previously uncharacterized regions of the egr-1 gene and found evidence of a potential oestrogen response element. Our findings are consistent with the hypothesis that Egr-1 forms at least part of the molecular signal for ovulation in the pituitary. PMID- 10849213 TI - Expression of the genes encoding the vasopressin-activated calcium-mobilizing receptor and the dual angiotensin II/vasopressin receptor in the rat central nervous system. AB - The distributions of two newly discovered receptors, the vasopressin-activated calcium-mobilizing receptor (VACM-1) and the dual angiotensin II/vasopressin receptor (AII/AVP), in the central nervous system (CNS) of the rat were determined using reverse transcriptase-polymerase chain reaction and in situ hybridization. The sequence of the rat VACM-1 cDNA was determined and found very homologous to the rabbit and human sequences. Both VACM-1 and AII/AVP receptor genes were widely expressed in the brain, but differed according to the cell type studied. Glial cells were very faintly labelled. The epithelial cells of the choroid plexuses, the ependymal cells and the pia mater were all labelled. Both genes were most active in neurones throughout the CNS. VACM-1 and AII/AVP receptors were detected in neurones previously shown to possess V1a and V1b vasopressin receptors, and/or the AT1 and AT2 angiotensin II receptors in many brain areas. This was the case for the magnocellular neurones of the supraoptic and paraventricular nuclei of the hypothalamus. We suggest that the VACM-1 and AII/AVP receptors may account for the V2-like responses to vasopressin by these neurones which lack a genuine V2 vasopressin receptor. PMID- 10849214 TI - Processing of frameshifted vasopressin precursors. AB - Biosynthesis of the vasopressin (VP) prohormone in magnocellular neurones of the hypothalamo-neurohypophysial system comprises endoplasmic reticulum (ER) transit, sorting into the regulated secretory pathway and subsequent processing in the individual proteins VP, neurophysin and a glycoprotein. These processes are severely disrupted in the homozygous diabetes insipidus (di/di) Brattleboro rat, which expresses a mutant VP precursor due to a single nucleotide deletion in the neurophysin region of the VP gene resulting in VP deficiency. Previous studies have shown the presence of additional frameshift mutations in VP transcripts, in solitary magnocellular neurones of the di/di rat due to a GA dinucleotide deletion resulting in two different mutant VP precursors with partly restored reading frame. Frameshifted VP precursors are also expressed in several magnocellular neurones in wild-type rats. In this study, we determined if the +1 frameshifted precursors from di/di and wild-type rats can lead to biosynthesis of the hormone VP. Therefore, eukaryotic expression plasmids containing the frameshifted VP cDNAs were transiently expressed in peptidergic tumour cell lines, and cells were analysed by reversed phase high-performance liquid chromatography and specific radioimmunoassays, and by immunofluoresence. Neuro2A neuroblastoma cells expressing the +1 frameshifted precursors of di/di rats retained products in the cell body. Only precursor or insignificant quantities of neurophysin-immunoreactive products were detected. In contrast, in AtT20 cells, frameshifted VP precursors were at least partly processed to yield the VP peptide, indicating that they have access to the regulated secretory pathway. Comparison between the two cell lines showed a very slow ER transit of the wild type prohormone combined with inefficient processing in Neuro2A cells. The results show that mutant precursors can reach the regulated secretory pathway if ER transport is sufficiently rapid as in the case of AtT20 cells. This suggests that the di/di rat may regain the capacity to biosynthesize authentic VP through these +1 frameshifted precursors in magnocellular neurones. PMID- 10849215 TI - Localization and physiological regulation of the exocytosis protein SNAP-25 in the brain and pituitary gland of Xenopus laevis. AB - In mammals, the synaptosomal-associated protein of 25 kDa, SNAP-25, is generally thought to play a role in synaptic exocytosis of neuronal messengers. Using a polyclonal antiserum against rat SNAP-25, we have shown the presence of a SNAP-25 like protein in the brain of the South-African clawed toad Xenopus laevis by Western blotting and immunocytochemistry. Xenopus SNAP-25 is ubiquitously present throughout the brain, where its distribution in various identified neuronal perikarya and axon tracts is described. Western blot analysis and immunocytochemistry also demonstrated the presence of SNAP-25 in the neural, intermediate and distal lobes of the pituitary gland. Intensity line plots of confocal laser scanning microscope images of isolated melanotropes indicated that SNAP-25 is produced and processed in the rough endoplasmatic reticulum and Golgi apparatus, and is associated with the plasma membrane. Immunoelectron microscopy substantiated the idea that SNAP-25 is present in the plasma membrane but also showed a close association of SNAP-25 with the bounding membrane of peptide containing secretory granules in both the neurohemal axon terminals in the neural lobe and the endocrine melanotropes in the intermediate lobe. Quantitative Western blotting revealed that adapting Xenopus to a dark background has a clear stimulatory effect on the expression of SNAP-25 in the neural lobe and in the melanotrope cells. This background light intensity-dependent stimulation of SNAP 25 expression was confirmed by the demonstration of increased immunofluorescence recorded by confocal laser scanning microscopy of individual melanotropes of black background-adapted toads. On the basis of this study on Xenopus laevis, we conclude that SNAP-25 (i) plays a substantial role in the secretion of a wide variety of neuronal messengers; (ii) functions in the central nervous system but also in neurohormonal and endocrine systems; (iii) acts at the plasma membrane but possibly also at the membrane of synaptic vesicles and peptide-containing secretory granules; (iv) acts not only locally (as in synapses), but at various sites of the plasma membrane (as in the endocrine melanotrope cell); and (v) can be upregulated in its expression by physiological stimuli that increase the extent of the molecular machinery involved in exocytosis. PMID- 10849216 TI - Personal viewpoint: eclecticism in health services for developmental disorders. AB - The term 'eclectic', as applied to health care for children with developmental disorders, portrays an individualized, adaptive service response to local constraints and pressures. While this may appear appropriate for the local setting, the end result is a broad diversity of health care approaches. This paper discusses three separate processes that interact at a local level, increasing the likelihood of an eclectic local model of health care for this population of children. The first process draws from the direct clinical work. Variable training, knowledge and skills among health care providers, in combination with differing beliefs around the nature of the problems and their management leads to health care which directly reflects the attributes of the local clinicians. A separate, second process fuelling variability is the differing models of departmental responsibility across Australia - which Government departments fund which aspect of care for children with disabilities. The final process relates to funding streams for health care. State public health, federal Medicare and private insurance all support health services for children with disabilities, with the financial incentives (budgets compared to fee-for-service) driving a divergence of practice. This paper concludes that the external political, administrative and financial frameworks within which health care is constructed will continue to promote clinical eclecticism to a degree that would probably be considered unacceptable in other areas of child health care. The solution can only arise from within the clinical work itself, with greater clarity of understanding around the nature of the disorders, the outcomes for which health care takes responsibility, and an increasing focus on an evidence based set of approaches towards achieving these. PMID- 10849217 TI - Persistent or severe back pain and stiffness are ominous symptoms requiring prompt attention. AB - BACKGROUND: Children with severe or persistent back pain and stiffness often have an underlying organic cause but there is a large differential diagnosis, examination may be difficult and the problem is relatively rare in general paediatric practice. These difficulties appeared to lead to delays in diagnosis and management of children with this problem. OBJECTIVES: To provide an approach to the diagnosis and management children with severe or persistent back pain or stiffness based on our clinical experience and the literature. METHODOLOGY: The case histories of 10 children with severe back pain seen by the authors over a 5 year period were reviewed. They were chosen as illustrative examples of the diagnostic and management problems and did not represent a systematic review of all cases seen by the authors over that time. RESULTS: Underlying causes included infection, inflammation, neoplasm, trauma and vascular malformation. Four of the children had spinal cord compression which required urgent decompression. There was one child with a conversion disorder but three children with organic disease were initially felt to have a conversion disorder. Investigations generally proceeded relatively slowly and the problem was not regarded as a semi-urgent situation carrying the risk of permanent paraplegia. Magnetic resonance imaging (MRI) scan of the spine was the investigation of choice. CONCLUSION: Children with severe or persistent back pain and stiffness have a wide variety of underlying causes. The possibility of underlying spinal cord compression should always be considered in children with this presentation. If the diagnosis is not obvious, MRI scan of the spine should be arranged without delay. PMID- 10849218 TI - Intellectual disability in Western Australia. AB - OBJECTIVE: To investigate the prevalence of intellectual disability in Western Australia (WA), its causes, prevention, and trends over time. METHODOLOGY: Data from an administrative database of intellectual disability in WA were used to report on the trends in intellectual disability in childhood. RESULTS: The prevalence of intellectual disability was 8.3 per 1000 live births in 1980-90. For half the cases, there was no known cause for the intellectual disability. Down syndrome accounted for 14 to 15% of all cases. Since the introduction of newborn screening, no WA-born child participating in the screening program has been diagnosed with intellectual disability as a result of either phenylketonuria or congenital hypothyroidism. The rate of autism spectrum disorders rose from three to six per 10 000 in the 1980-83 WA birth cohort to 10-13 per 10 000 for the 1989-92 cohort. CONCLUSIONS: Recent linkage of this administrative database to the WA Maternal and Child Health Research Data Base provides a unique opportunity for more detailed investigation of intellectual disability and its risk factors in a large, well-ascertained population of children. PMID- 10849219 TI - Insulin infusions in the neonatal unit: delivery variation due to adsorption. AB - OBJECTIVE: To assess the extent of the variability in insulin delivery over time, under conditions used in Australian neonatal units, studying the following variables: diluent, preconditioning, flushing, sequential preconditioning and flushing, insulin concentration, flow rate, catheter type, and addition of albumin. METHODOLOGY: A range of simulated insulin infusions was studied. Infusions were run over 22 h, with aliquots of infusate collected at baseline and after 15 min, 30 min, 1 h, 2 h, 6 h and 22 h. Insulin concentration was assayed using a radioimmunoassay. RESULTS: An infusion of 50 mU insulin/mL at 1 mL/h gave negligible insulin delivery until 22 h. However, after preconditioning and flushing the tubing, consistent insulin delivery was attained by 6 h. An infusion of 200 mU insulin/mL at 1 mL/h did not provide consistent delivery until 6 h. At this concentration and rate, consistent insulin delivery was attained within 30 min of the start of the infusion either by preconditioning and flushing the tubing, or by addition of albumin to the infusate. CONCLUSION: Clinically significant variation in intravenous insulin delivery will occur in the neonatal setting unless counter-measures are taken, such as sequential preconditioning and flushing of the delivery system or the addition of albumin to the infusate. PMID- 10849221 TI - Early motor development of blind children. AB - OBJECTIVES: The purpose of this study was to assess the characteristic motor developmental pattern in blind children in Israel. METHODOLOGY: The study compared the developmental data concerning 10 motor skills of 40 blind children to a control group of sighted children and to the motor developmental milestones of the Bayley Developmental Scale and the Revised Denver Developmental Screening Test. RESULTS: The motor development of blind children was delayed, the delay being significant in all 10 motor skills that were examined. This delay emphasizes the major importance of vision as a sensory input modality for the process of sensory- motor development. CONCLUSION: An adequate stimulating environment and proper parental handling could potentially shorten the motor developmental delay but probably not eliminate it entirely. PMID- 10849220 TI - Clinical markers of serious illness in young infants: a multicentre follow-up study. AB - OBJECTIVE: To perform a multicentre follow-up study to determine if previously identified markers of serious illness in early infancy were robust and statistically reliable. METHODS: Infants aged 1 week to 26 weeks presenting to the Emergency Departments of the Royal Children's Hospital and two Melbourne metropolitan hospitals were seen over a 12-month period. Eleven clinical markers as well as their temperature were documented by nursing staff and resident medical officers. Serious illness was defined if infants had a positive body fluid bacterial culture, a positive chest X-ray or if significant treatment was required in hospital. The predictive values, sensitivity and specificity for the individual and the best combination of clinical markers were determined. RESULTS: Assessments (3806) were performed with 312 infants being assessed as seriously ill (8.2%). The combination of either drowsiness on history or examination, pallor on history or examination, breathing difficulty (chest wall recession), temperature above 38 degrees C and a lump being present, identified 82.5% of all babies deemed subsequently to be seriously ill. The positive predictive value of an infant who was febrile, drowsy and pale on examination was 70.7% (previous study 74%). CONCLUSIONS: This study confirmed the high individual predictive value of arousal variables, pallor, and chest wall recession, especially when associated with fever, reaffirming their utility in the recognition of serious illness in infants under 6 months of age. PMID- 10849222 TI - Children of renal transplant recipient mothers. AB - OBJECTIVE: To assess the current physical status and developmental outcome of children born to mothers following renal transplantation. METHODOLOGY: A cross sectional prevalence survey of 48 children born to 34 women transplanted at a single centre from 1971 to 1992 was performed. Data on maternal renal disease, immunosuppression, pregnancy, delivery and child development were collected using hospital records and parental questionnaire. Children underwent physical examination, urinalysis and urinary tract ultrasound examination (US). RESULTS: Maternal renal failure was due to reflux nephropathy/chronic pyelonephritis (16), chronic glomerulonephritis (eight) and other causes (10). All mothers received prednisolone immunosuppression, as sole therapy (one), as part of triple therapy (one). Sixteen (47%) received azathioprine/prednisolone and 16 (47%) cyclosporin/prednisolone. Twenty-three girls and 25 boys aged 9 months to 18 years were studied (median age 5.2 years); 27/48 (56%) were born prematurely, 21/48 (44%) with birthweight (BW) < 2500 g 21/48 (44%) were small for gestation (BW < 10th centile). General health and physical assessment were unremarkable in 45/48 (94%) and 41/43 (95%), respectively. Development was considered normal in 47/48 (98%). Four of 40 (10%) had urinary tract abnormalities on US. CONCLUSIONS: Despite a high incidence of preterm delivery, low birth weight, intrauterine growth retardation and urinary tract abnormalities, the overall outcome for children of renal transplant recipient mothers is good. PMID- 10849223 TI - Dietary fibre intake and constipation in children with severe developmental disabilities. AB - OBJECTIVE: Constipation is a common problem in children with severe developmental disabilities (DD). This study aimed to evaluate fibre intake of severe DD children living in a residential institution, and the possibility of reducing the use of laxatives by increasing their fibre intake. METHODOLOGY: A baseline study was performed to evaluate the fibre and macronutrient intake in a group of severe DD children. Nutrients including fibre for a standard serving in each meal were calculated and daily macronutrients and fibre intake were estimated. An intervention study was then carried out to evaluate whether increasing fibre intake could relieve constipation. A total of 20 children aged between 3 and 17 years were assessed over a 4-month period. In a residential unit for severe DD children, laxatives were routinely prescribed if there was no spontaneous bowel motion for two consecutive days. Fibre intake was increased in stages by adding All-Bran(R) (Kellogg Company, Battle Creek, MI, USA) and desserts. The mean number of laxative usage per week per child in the different stages were then compared. RESULTS: The baseline fibre intake was found to be approximately 2 g/day. The mean number of laxatives required per week per child decreased significantly from a baseline value of 1.22 (about 5 laxatives/month) (standard deviation (SD) = 0.36)) to 0.90 (about 3. 5 laxatives/month) (SD = 0.75) in the first stage, and 0.71 (about 3 laxatives/month) (SD = 0.40) in the second stage. Using paired t-test, the difference was statistically significant when compared with the baseline: P < 0.05 for the first, and P < 0.01 for the second stage of fibre supplementation. CONCLUSION: Very low daily intake of fibre of 2 g/day was documented. Relief of constipation and a significant reduction in the usage of laxatives was demonstrated by increasing fibre intake to 17 g/day (stage 1). Increasing fibre intake further to 21 g/day (stage 2), showed a further reduction in the use of laxatives. There was, however, no statistical significance between stage 1 and stage 2 of fibre supplementation. Alternative ways to further relieve constipation in severe DD children require further studies. PMID- 10849224 TI - Hearing in children after meningococcal meningitis. AB - OBJECTIVE: To review the outcome of sensorineural hearing loss in children who have had confirmed meningococcal meningitis. METHODS: A retrospective audit of children admitted to the Starship Children's Hospital with a confirmed diagnosis of meningococcal meningitis during a 4-year period. RESULTS: Sixty-five children had confirmed meningococcal meningitis. Dexamethasone was administered according to recommended guidelines in 46 children (72%) while 12 children (19%) did not receive steroids at any stage. All children were appropriately referred for hearing assessment. Forty-nine children (75%) had their hearing tested and reliable sensorineural evaluation was obtained in all but one case. Thirty-four children (70%) were seen up to 6 weeks from discharge, 86% by 12 weeks. Sixteen cases (25%) did not attend for audiological assessment. Sensorineural hearing impairment was found in two children (4.2%). CONCLUSIONS: Children with meningococcal meningitis were reliably referred for audiological assessment but 39% either failed to attend for an outpatient hearing evaluation (25%) or had an unacceptable delay between discharge and testing (14%). Of those reliably tested, two children (4.2%) had significant sensorineural hearing loss. PMID- 10849225 TI - Paroxysmal non-epileptic events in children: a retrospective study over a period of 10 years. AB - OBJECTIVE: To determine the frequency, nature and clinical characteristics of paroxysmal non-epileptic events in children diagnosed by video electroencephalogram (EEG) monitoring at a tertiary referral centre. METHODOLOGY: A retrospective study of children with paroxysmal non-epileptic events, aged 2 weeks to 17 years inclusive was undertaken. The study group consisted of children who had video EEG monitoring during a 10-year period (1988-99). Telemetry files, medical charts, events recorded on video and record sheets were reviewed. RESULTS: A total of 666 children were analysed, 269 had epileptic events recorded, 285 had non-epileptic events and 112 had no events recorded. In children with non-epileptic events, 43% were developmentally delayed, 25% had an abnormal neurological examination and 40% had epilepsy. In the study sample an epileptiform interictal EEG was common (24%). The major subgroups of non epileptic events were: staring (34%), sleep phenomena - benign sleep myoclonus (15%), arousals (13%), motor tics (11%) and shuddering (7%). Developmental delay (57%) was common in children who presented with staring spells. A diagnosis of a specific non-epileptic event was reached in 96% of cases. CONCLUSION: Paroxysmal non-epileptic events can cause diagnostic confusion, particularly in children with developmental delay, epilepsy or an epileptiform EEG. Accurate diagnosis can be reached in the majority of cases using video EEG monitoring. PMID- 10849226 TI - Asthma management in indigenous children of a remote community using an indigenous health model. AB - OBJECTIVE: To describe the management of asthma in children in a remote indigenous community and the delivery of subspecialist service through the indigenous health-care model. METHODOLOGY: Children referred by indigenous health care workers were evaluated prospectively by paediatric respiratory physicians, based on a standardized protocol, at a primary health care setting at Thursday Island, Queensland. RESULTS: Forty of the 54 children referred with a provisional diagnosis of asthma did have asthma, with 30% having persistent asthma. Only 59% of parents knew the dose of the medication prescribed and 80% had minimal knowledge of the medications. In 88% of children, the management of asthma was improved by introduction of an appropriate spacer device and changing the dose and type of medications. CONCLUSIONS: The management of children with asthma in the Torres region can be improved substantially by the use of age appropriate delivery devices and medications, and improving knowledge of asthma. Specialist delivery service to remote indigenous communities can be effectively delivered in partnership with the indigenous health service. The high proportion of persistent asthma in the Torres Straits community in comparison to urbanised Australia raises issues of inequity of appropriate medical service delivery to remote indigenous communities. PMID- 10849227 TI - A 7-year review of deaths on the general paediatric wards at John Hunter Children's Hospital, 1991-97. AB - OBJECTIVE: To study the palliative management of children dying on the wards of a tertiary referral centre. In particular to identify areas of difficulty and discuss ways in which these can be improved. METHODOLOGY: All children between the ages 1 and 19 years dying on the general paediatric wards of the John Hunter Children's Hospital (JHCH) (a tertiary referral centre in New South Wales Hunter Valley) between 1991 and 1998 were included in the study. The main outcome measures were the duration of 'terminal care only' treatment; time spent in hospital in the previous year; analgesia used, dose and route of administration; adjuvant medications; quality of life in the final 24 h. RESULTS: Eighteen children (aged between 1 and 19 years) died on the general paediatric wards at JHCH between 1991 and 1998. The most common diagnosis was malignancy (five patients), with cystic fibrosis, cerebral palsy and a degenerative neurological disorder each accounting for four patients, and one child had herpes simplex encephalitis. In all children the terminal nature of their condition was recognized and in all but three patients there was documentation of 'not for resuscitation' (NFR) orders following discussion with the child's parents. The mainstay of analgesia was an intravenous narcotic infusion with 11 (61%) children needing a morphine infusion. Fifteen patients (83%) required other medications including antiemetics, anticonvulsants and corticosteroids. Twelve (66%) children were semiconscious or unconscious on the day of death. Three children were uncomfortable and distressed, one alert and symptom free and in two cases the level of consciousness was not recorded. Three children continued to have seizures until their death. CONCLUSION: The majority of terminally ill children were managed well with intravenous narcotics and adjuvant medications. There may be scope for such children to be managed successfully at home with appropriate support for the general practitioner by paediatric and palliative care specialists. Some cases in which palliation is difficult do remain in hospital and additional measures need to be employed to ensure a symptom free death for these children. PMID- 10849228 TI - Postnatal corticosteroids and sensorineural outcome at 5 years of age. AB - BACKGROUND: Postnatal corticosteroids reduce ventilator dependence in preterm infants, but possible long-term benefits for either survival or sensorineural morbidity are not proved. AIM: The aim of this study was to determine the association between corticosteroid therapy given postnatally and sensorineural outcome in childhood. SUBJECTS: The subjects comprised 346 consecutive livebirths either of birthweight < 1000 g or with gestational age < 28 weeks born in the state of Victoria during 1991 and 1992, and who survived the first week after birth; 120 (34.7%) were given corticosteroids postnatally. RESULTS: Of the 120 children who received corticosteroids, 98 (81.7%) survived to 5 years of age, compared with 200 (88.5%) of the 226 children who did not receive corticosteroids. At 5 years of age, survivors treated with corticosteroids postnatally had significantly higher rates of cerebral palsy (corticosteroids 23%, no corticosteroids 4%), blindness (corticosteroids 4%, no corticosteroids 1%) or an intelligence quotient more than one standard deviation below the mean (corticosteroids 54%, no corticosteroids 32%) compared with children not treated with corticosteroids. The rate of sensorineural disabilities imposed by these impairments was significantly higher in children treated with postnatal corticosteroids, and the association between adverse sensorineural outcome and postnatal corticosteroids remained after adjustments for potentially confounding variables. In a separate case-control analysis of 60 children in each group, the rate of cerebral palsy remained significantly elevated (corticosteroids 22%, no corticosteroids 5%). CONCLUSION: Postnatal corticosteroid therapy is associated with substantial adverse sensorineural outcomes at 5 years of age. PMID- 10849229 TI - The drug epidemic: effects on newborn infants and health resource consumption at a tertiary perinatal centre. AB - OBJECTIVES: Illicit drug taking in Australia, with its attendant social and medical consequences, is increasing and the effects extend to maternity hospitals where infants born to addicted mothers have more health problems in the neonatal period. The aims of this study were to evaluate (1) the patterns of illness of such infants and (2) the burden imposed on the neonatal department of a large tertiary maternity centre. METHODOLOGY: An audit was conducted of all Chemical Dependency Unit (CDU) mothers and babies delivered at the Royal Women's Hospital, Melbourne, Australia during 1997. Data were compared with those from a concurrent control group of mothers and babies randomly generated from the hospital's obstetric database. RESULTS: Ninety-six infants born to CDU mothers were compared with a control group of 200 infant/mother pairs. The majority of women in the CDU clinic were treated for narcotic addiction with methadone (90%) but most continued to use heroin during pregnancy (68%). Infants born to CDU mothers were significantly less mature and lighter than control infants. Fifty-three (55%) CDU infants required admission to the Special Care Nursery either because of neonatal abstinence syndrome (n = 29) or other medical reasons (n = 24). The median length of hospital stay was significantly longer in CDU compared with control infants (8 vs 3 days, P < 0.01). CONCLUSIONS: Infants born to drug dependent mothers have more neonatal problems requiring specialized medical and nursing expertise, compared with control infants. These infants are large consumers of scarce health resources. PMID- 10849230 TI - Small bowel injuries in children. AB - OBJECTIVE: To determine the common features of small bowel injury (SBI) in childhood and the consequences of delayed diagnosis. METHODOLOGY: A retrospective case review was performed of children with traumatic SBI between January 1988 and November 1999. RESULTS: Twenty-eight patients were identified with SBI. Road trauma accounted for 71% of them. Tachycardia was present on admission in 82% of patients with SBI including all but one of the intestinal perforations. SBI was associated with a Chance fracture of the lumbar spine in three patients (11%). An abdominal computed tomography scan with intravenous contrast was abnormal in all patients with a perforation or mesenteric tear. Diagnosis was delayed in six patients, one of whom died as a result of sepsis from a small bowel perforation. CONCLUSIONS: Persistent tachycardia with an appropriate mechanism of injury following blunt abdominal trauma requires active exclusion of SBI. Delayed diagnosis is associated with significant morbidity and mortality. PMID- 10849231 TI - 2: describing and displaying data. PMID- 10849232 TI - Back pain in a 4-year-old boy. PMID- 10849233 TI - A painful limp. AB - We describe a healthy 18-month-old child who developed a painful limp, without a history of trauma or fever. The initial laboratory investigations showed normal results but the radiological findings were suggestive of scurvy. Diagnosis was confirmed by blood tests and by a rapid recovery following replacement therapy. PMID- 10849234 TI - Toxic shock syndrome associated with newly diagnosed type I diabetes. AB - Studies of two post-mortem pancreata of children at the onset of type I diabetes have suggested activation and expansion of islet infiltrating T cells by a superantigen. We present the first reported case of a superantigen mediated disease, toxic shock syndrome (TSS), occurring at the diagnosis of type I diabetes. A 12-year-old girl presented with TSS and newly diagnosed diabetes with ketoacidosis. At presentation she was unconscious, febrile and hypotensive, with a desquamating erythematous rash and Kussmaul breathing. During resuscitation, her renal impairment, diarrhoea, thrombocytopaenia and ketoacidosis resolved. Vaginal discharge and blood cultures grew Staphylococcus aureus. T cell studies at 2 weeks after diagnosis detected a high level of spontaneous and islet antigen specific proliferation with associated interleukin-10 production compared to human leucocyte antigen DR matched controls. PMID- 10849235 TI - Bromocriptine for the management of autonomic dysfunction after severe traumatic brain injury. AB - This case report describes a child with severe traumatic brain injury with clinical features of autonomic dysfunction in the immediate post-traumatic period. A history of severe asthma in this child contraindicated the use of beta blockers, the first line approach, and she was managed with bromocriptine (0.05 mg/kg t.d.s) with good effect. PMID- 10849236 TI - Diffuse neonatal haemangiomatosis. AB - A newborn girl with severe diffuse neonatal haemangiomatosis is described. She was treated with high dose systemic corticosteroids and high dose interferon alpha-2a, but with fatal outcome. A review of the current literature is presented. PMID- 10849237 TI - Inducing jet-lag in older people: directional asymmetry. AB - Twenty healthy elderly subjects (12 female, 8 male; mean age 81 years, range 67 87 years) each experienced a 15-day time isolation protocol in which they lived individually in a special laboratory apartment in which sleep and circadian rhythm measures could be taken. There were two experiments: one (6 females, 4 males) involved a 6-h phase advance of the sleep/wake cycle, and the other (6 females, 4 males) a 6-h phase delay. Each started with 5 baseline days, immediately followed by the phase shift. The subject was then held to the phase shifted routine for the remainder of the study. Rectal temperatures were recorded minute-by-minute throughout the entire experiment and each night of sleep was recorded using polysomnography. A directional asymmetry in phase-shift effects was apparent, with significantly more sleep disruption and circadian rhythm amplitude disruption after the phase advance than after the phase delay. Sleep disruption was reflected in reduced time spent asleep, and in changed REM latency, which increased in the phase advance direction but decreased in the phase delay direction. Although the phase advance led to a significant increase in wakefulness in the first half of the night, the phase delay did not lead to an equivalent increase in wakefulness during the second half of the night. Examination of both raw and 'demasked' circadian rectal temperature rhythms confirmed that phase adjustment was slow in both directions, but was less slow (and more monotonic) after the phase delay than after the phase advance. Subjective alertness suffered more disruption after the phase advance than after the phase delay. PMID- 10849238 TI - Individual differences in the phase and amplitude of the human circadian temperature rhythm: with an emphasis on morningness-eveningness. AB - We studied the relationship between the phase and the amplitude of the circadian temperature rhythm using questionnaires that measure individual differences in personality variables, variables that relate to circadian rhythms, age and sex. The ambulatory core body temperature of 101 young men and 71 young women was recorded continuously over 6 days. The temperature minimum (Tmin) and amplitude (Tamp) were derived by fitting a complex cosine curve to each day's data for each subject. Participants completed the Horne-Ostberg Morningness-Eveningness Questionnaire (MEQ), the Circadian Type Inventory (CTI) and the MMPI-2, scored for the Psychopathology-5 (PSY-5) personality variables. We found that the average Tmin occurred at 03.50 h for morning-types (M-types), 05.02 h for the neither-types and 06.01 h for evening-types (E-types). Figures were presented that could provide an estimate of Tmin given an individual's morningness eveningness score or weekend wake time. The Tmin occurred at approximately the middle of the 8-h sleep period, but it occurred closer to wake in subjects with later Tmin values and increasing eveningness. In other words, E-types slept on an earlier part of their temperature cycle than M-types. This difference in the phase-relationship between temperature and sleep may explain why E-types are more alert at bedtime and sleepier after waking than M-types. The Tmin occurred about a half-hour later for men than women. Another interesting finding included an association between circadian rhythm temperature phase and amplitude, in that subjects with more delayed phases had larger amplitudes. The greater amplitude was due to lower nocturnal temperature. PMID- 10849239 TI - The effect of arousals during sleep onset on estimates of sleep onset latency. AB - It is well established that insomniacs overestimate sleep-onset latency. Furthermore, there is evidence that brief arousals from sleep may occur more frequently in insomnia. This study examined the hypothesis that brief arousals from sleep influence the perception of sleep-onset latency. An average of four sleep onsets was obtained from each of 20 normal subjects on each of two nonconsecutive, counterbalanced, experimental nights. The experimental nights consisted of a control night (control condition) and a condition in which a moderate respiratory load was applied to increase the frequency of microarousals during sleep onset (mask condition). Subjective estimation of sleep-onset latency and indices of sleep quality were assessed by self-report inventory. Objective measures of sleep-onset latency and microarousals were assessed using polysomnography. Results showed that sleep-onset latency estimates were longer in the mask condition than in the control condition, an effect not reflected in objective sleep-stage scoring of sleep-onset latency. Furthermore, an increase in the frequency of brief arousals from sleep was detected in the mask condition, and this is a possible source for the sleep-onset latency increase perceived by the subjects. Findings are consistent with the concept of a physiological basis for sleep misperception in insomnia. PMID- 10849240 TI - The ability to self-monitor performance when fatigued. AB - The present study aimed to systematically investigate the effects of elevated fatigue levels on the ability to self-monitor performance. Eighteen participants, aged 19-26 y, remained awake for a period of 28 h. Neurobehavioural performance was measured at hourly intervals using four tests from a standardized computer test battery. From these four tests, six measures of performance were obtained: grammatical reasoning (accuracy and response latency); vigilance (accuracy and response latency); simple sensory comparison and tracking. In addition, before and after each test, participants completed visual analogue scales which required them to rate their alertness level and the speed and accuracy of their performance. Individual test results for both self-ratings and neurobehavioural performance were converted to z-scores. Planned comparison analysis indicated that scores on four of the six performance measures decreased significantly as hours of wakefulness increased. Similarly, predicted performance scores for all six measures of performance decreased significantly. Analysis revealed moderate correlations between predicted and actual performance for the four parameters affected by fatigue. Furthermore, moderate to high correlations were found between all six performance parameters and their respective post-test self ratings. In addition, moderate to high correlations were found between predicted performance and alertness. Taken together, these findings suggest that as fatigue levels increase, subjects globally assess performance decrements. Results suggest that subjective alertness may in part mediate an individual's global assessment of performance. PMID- 10849241 TI - Time in bed, quality of sleep and school functioning of children. AB - This study describes the relationship of time in bed and quality of sleep with concentration and functioning at school. Neurotic and psychosomatic symptoms have been used as control variables. The sample consisted of 449 Dutch children in the seventh and eighth grades of elementary school. The age of the children varied between 9 y 5 mo and 14 y 5 mo. Seven schools participated in the research, with a total of 18 classes. The results indicated that 43% of the children had difficulty getting up in the morning. Furthermore, 15% of the children reported sleep problems and 25% did not feel rested at school. Time in bed and sleep quality show no relationship with concentration. Sleep quality, feeling rested at school and less distinct bedtimes were clearly related to school functioning. Another result was that children who had no difficulty getting up displayed more achievement motivation. Being open to the teacher's influence and achievement motivation depended mainly on sleep characteristics. Not getting bored at school, self-image as a pupil and control over aggressive behaviour were also influenced by gender, age, neuroticism and neurosomaticism. PMID- 10849242 TI - The relationship between frequency of rapid eye movements in REM sleep and SWS rebound. AB - Previous studies have shown a decrease in rapid eye movement (REM) frequency during desynchronized sleep in recovery nights following total or partial sleep deprivation. This effect has been ascribed to an increase in sleep need or sleep depth consequent to sleep length manipulations. The aims of this study were to assess REM frequency variations in the recovery night after two consecutive nights of selective slow-wave sleep (SWS) deprivation, and to evaluate the relationships between REM frequency and SWS amount and auditory arousal thresholds (AAT), as an independent index of sleep depth. Ten normal males slept for six consecutive nights in the laboratory: one adaptation, two baseline, two selective SWS deprivation and one recovery night. SWS deprivation allowed us to set the SWS amount during both deprivation nights close to zero, without any shortening of total sleep time. In the ensuing recovery night a significant SWS rebound was found, accompanied by an increase in AAT. In addition, REM frequency decreased significantly compared with baseline. This effect cannot be attributed to a variation in prior sleep duration, since there was no sleep loss during the selective SWS deprivation nights. Stepwise regression also showed that the decrease in REM frequency is not correlated with the increase in AAT, the traditional index of sleep depth, but is correlated with SWS rebound. PMID- 10849243 TI - Functional neuroanatomy of human sleep states after zolpidem and placebo: a H215O PET study. AB - Changes in the functional organization of the brain during the course of sleep and waking are reflected by different patterns of regional cerebral blood flow (rCBF). To investigate the effect of the hypnotic zolpidem, a benzodiazepine receptor agonist, drug or placebo were administered to eight young, healthy men prior to bedtime. The subjects were sleep-deprived to promote sleep during the 4 h recording period in the positron emission tomography scanner. Intravenous injections of labelled water were administered during pre-drug wakefulness, and during Stage 2, Stage 4 and rapid eye movement (REM) sleep, each injection being followed by an emission scan. Statistical parametric mapping was used to investigate the effects of treatment and sleep states. During sleep (combined Stages 2 and 4, and REM sleep) relative rCBF was lower after zolpidem than after placebo in the basal ganglia and insula, and higher in the parietal cortex. A 'multiple study' analysis of REM sleep revealed that rCBF in the anterior cingulum was lower after zolpidem than after placebo, whereas rCBF in the occipital and parietal cortex, parahippocampal gyrus and cerebellum was higher. When the pooled data (drug and placebo) of Stages 2 and 4 were compared with wakefulness, rCBF was lower in prefrontal cortex and insula, and higher in the occipital and parietal cortex. The results indicate that some differences in rCBF from wakefulness to non-REM sleep are further augmented by zolpidem. PMID- 10849244 TI - Zolpidem and sleep deprivation: different effect on EEG power spectra. AB - To study the role of GABA-ergic mechanisms in sleep regulation, the combined action of 40 h sleep deprivation and either 20 mg zolpidem or placebo on the sleep electroencephalogram (EEG) were investigated by quantitative EEG analysis in eight young men who participated in a positron emission tomography study. Compared with baseline, sleep deprivation increased low-frequency (1.25-7.0 Hz) EEG power in non-rapid eye movement (NREM) sleep in the placebo night. After administration of zolpidem, power in the 3.75-10.0 Hz range and 14. 25-16.0 Hz band was reduced. The largest decrease was observed in the theta band. Comparison with placebo revealed that zolpidem attenuated power in the entire 1.75-11.0 Hz range. The plasma concentration of zolpidem at 4.5 h after intake showed a positive correlation with the drug-induced difference in power from placebo in the 14.25-16.0 Hz band. Regional EEG analysis based on bipolar derivations along the antero-posterior axis disclosed, for NREM sleep, a drug-induced posterior shift of power in the frequency range of 7.75-9.75 Hz. Zolpidem did not affect rapid eye movemnt sleep spectra. We conclude that sleep deprivation and agonistic modulation of GABAA receptors have separate and additive effects on power spectra and that their effects are mediated by different neurophysiological mechanisms. PMID- 10849245 TI - Transient cardiorespiratory events during NREM sleep: a feline model for human microarousals. AB - Microarousals (MAs) are brief transient events that occur during normal sleep in humans and with increased frequency in disordered sleep, especially in association with sleep apnoea. In a feline model, we discovered transient cardiorespiratory events during nonrapid eye movement (NREM) sleep that exhibited consistent features with similarities to human MAs. It was observed that MAs have two distinct phases. Phase I (MAI) is characterized by an abrupt increase in electromyogram (EMG) amplitude (> 50%), increased electrooculogram (EOG) activity and accelerated frequency of hippocampal electroencephalographic (EEG) activity. MAI lasts 4.1 +/- 0.3 s. Phase II (MAII), lasting 9.8 +/- 0.8 s, is characterized by high frequency EEG activity, but EMG, EOG and hippocampal activity remain at baseline levels. Mean inspiratory rate begins to increase 15 s before the onset of the MA, followed 10 s later by the increase in mean heart rate. Mean respiratory rate decreases sharply through MAII, and returns to baseline levels 15 s after the MA. During MAII mean heart rate decreases quickly; there is increased respiratory irregularity, followed by a prolonged ventilatory overshoot. The abrupt shift in heart rate is coincident with the change in breath timing seen during MAII. Heart rate returns to baseline levels 10 s following the MA. Integrating our findings with those described previously in humans, we propose that MAs may serve as a homeostatic mechanism which is designed to restore cardiorespiratory function allowing the continuity of sleep. PMID- 10849246 TI - Naso-oesophageal probes decrease the frequency of sleep apnoeas in infants. AB - The objective of the study was to determine whether a naso-oesophageal probe modifies sleep and cardiorespiratory patterns in infants with repeated obstructive apnoeas. Two polygraphic recording sessions were conducted in random order for 2 nights on 35 infants suspected to have repeated obstructive sleep apnoeas. One sleep study was performed with a pH probe inserted through the nasal passage down to the distal portion of the oesophagus. The other session was conducted without any naso-oesophageal probe (the baseline study). For the 25 infants who presented repeated obstructive apnoeas during baseline studies, the presence of the probe was associated with a small, but significant, decrease in the number of central apnoeas (median frequency of 18.5 apnoeas per hour without a probe; 16.1 per hour with the probe; P=0.040), and obstructive apnoeas (median of 1.9 apnoeas per hour without a probe; 0.6 per hour with the probe; P=0.016). The presence of the probe was also associated with a small increase in percentage non-rapid eye movement (NREM) sleep frequency. The changes were statistically significant only for infants who had no obstructive apnoea during baseline studies (29 vs. 31%). The presence of a naso-oesophageal probe significantly modifies the infants' respiratory characteristics during sleep. These findings should be considered when reporting and interpreting sleep studies in infants. PMID- 10849247 TI - Reported snoring--does validity differ by age? AB - Snoring is a major sign of obstructive sleep apnoea syndrome. Despite the frequent number of studies based on subjective reports of snoring, self-reported snoring has hardly been validated at all. In some previous epidemiological studies, a significant association between snoring and cardiovascular morbidity and mortality was found only below the age of 50-60 y. This study was performed to investigate whether this is due to a decrease in the validity of reported snoring with increasing age. In a population-based study, 2668 men aged 40-79 y answered a questionnaire including questions on snoring. Those who reported loud and disturbing snoring often or very often were regarded as habitual snorers. Without taking account of reported snoring, an age-stratified sample of these men was selected and their snoring was measured using a microphone for 1 night. Significant snoring was defined as recorded snoring sounds for >/= 10% of the night. The participants were divided into younger (age 40-59, mean +/- SD: 51.8 +/- 4.6 y, n=132) and older (age 60-79, 67.7 +/- 5.4 y, n=99) age groups. When analysing the validity of reported snoring, no significant differences were found between the younger and older age groups in terms of specificity [younger: 82% (95% CI 74-90%), older: 88% (81-95%)] or sensitivity [younger: 40% (26-54%), older: 35% (17-53%)]. These data indicate that, in men aged 40-79 y, the validity of reported snoring is similar in different age groups. The lack of an association between reported snoring and cardiovascular disease at higher ages can, therefore, not be explained by a decrease in the validity of reported snoring. PMID- 10849248 TI - Does the physiological success of CPAP titration predict clinical success? AB - Patients commencing continuous positive airway pressure therapy (CPAP) undergo overnight airway pressure titration in sleep centres to optimize breathing and sleep patterns. We tested the hypothesis that data from formal scoring of the sleep and breathing patterns observed at the best achievable pressure during titration can predict CPAP use and effectiveness, as our clinical experience suggested otherwise. The relationship between CPAP titration scores (apnoea/hypopnoea frequency, arousal frequency and sleep staging) and subsequent CPAP use was examined in 150 sleep apnoea/hypopnoea syndrome patients. One hundred patients were continuing CPAP therapy and 50 were randomly selected patients who had discontinued CPAP. Within the CPAP group, titration scores were compared with CPAP machine use, subjective daytime sleepiness and requirements for airway pressure adjustment. Respiratory irregularities and arousals during titration did not relate to outcome. Sleep-stage analysis revealed a weak relationship between more wakefulness during titration and CPAP discontinuation (P=0.02). There was a correlation between more prolonged Stage 4 sleep during titration and reduced sleepiness on established therapy (P=0.002), but this explained less than 12% of the variance. The absence of rapid eye movement sleep during titration was not associated with poorer outcomes. We conclude that routine scoring of breathing and sleep patterns observed during CPAP titration is of little clinical value, as the results do not predict outcome for individual patients. Satisfactory CPAP therapy may be established even if significant numbers of apnoeas/hypopnoeas or arousals are observed at the optimal pressure during titration. PMID- 10849249 TI - Drug metabolism: in vitro biotransformation of anabolic steroids in canines. AB - Forensic drug testing of anabolic steroids in racing animals is required because of the potential for steroid abuse. Often when the metabolic products of an administered compound have not been identified, the analysis and verification of the administered compound is difficult. The objective of this study was to qualitatively identify the in vitro phase I biotransformation products of anabolic steroids that have a high potential for abuse in canines. The investigated steroids included testosterone, methyltestosterone, mibolerone and boldenone. Steroid biotransformation products were generated using beagle liver microsomes and analysed by high performance liquid chromatography (HPLC)/mass spectrometry (MS) with an electrospray ionization source. Characterization of steroid metabolites was based on HPLC retention, UV and mass spectra. The major testosterone metabolites were identified as androstenedione and 6beta- and 16alpha-hydroxytestosterone. 6beta-Hydroxymethyltestosterone was identified as a major metabolite in the methyltestosterone microsomal incubations. Several mibolerone metabolites were identified as monohydroxylated mibolerones as well as an oxidized mibolerone metabolite. Boldenone metabolites were identified as monohydroxylated boldenones, oxidized boldenone, and testosterone. This information should assist in the determination of anabolic steroid use in canines through the correlation of the urinary metabolites to the administered drug. PMID- 10849250 TI - Influence of interleukin-1beta and hyaluronan on proteoglycan release from equine navicular hyaline cartilage and fibrocartilage. AB - Proteoglycan (PG) release, in response to recombinant human interleukin-1beta (rh IL-1beta), was measured in cartilage explants obtained from the equine distal sesamoid bone (navicular bone). Fibrocartilage from the surface of the navicular bone apposing the deep digital flexor tendon and hyaline cartilage from the surface of the navicular bone articulating with the middle phalanx were labelled with 35SO4. Hyaline cartilage from the distal metacarpus was used as a control tissue. Following radiolabel incorporation, the three cartilage types were treated with rh-IL-1beta (100 U/mL) in the presence of hyaluronan (0.2, 2, 20, 200 and 2000 microgram/mL). rh-IL-1beta-Induced PG release was measured by scintillation assay of PG-bound radiolabel. Increases in PG release of 94% (P < 0.01), 101% (P < 0.05) and 122% (P < 0.05), in response to rh-IL-1beta, were noted in fibrocartilage, navicular hyaline cartilage and metacarpal hyaline cartilage, respectively. Hyaluronan (0.2 microgram/mL) significantly reduced rh IL-1beta-induced PG release in metacarpal hyaline cartilage (P < 0.01). In fibrocartilage and navicular hyaline cartilage, hyaluronan did not reduce PG release and at some concentrations appeared to increase PG release, although this was not statistically significant. These experiments show that (i) fibrocartilage and hyaline cartilage of the navicular bone release PGs in response to rh-IL 1beta, and (ii) hyaluronan does not prevent rh-IL-1beta-induced breakdown of navicular bone cartilage. PMID- 10849251 TI - The pharmacokinetics and effects of intravenously administered carprofen and salicylate on gastrointestinal mucosa and selected biochemical measurements in healthy cats. AB - The pharmacokinetics of carprofen, a propionic acid-derived nonsteroidal anti inflammatory (NSAID), and its effect on gastrointestinal mucosa, complete blood counts (CBC) and biochemical indicators of liver and renal function were investigated in healthy cats using a randomized crossover design. A single dose of 4 mg/kg of carprofen (Zenecarp(R) Injection), normal saline, or 20 mg/kg of DL lysine acetyl salicylate (Vetalgine(R)) was given intravenously (i.v.) to each of five cats with a washout period of 2 weeks between treatments. Endoscopy of the stomach and duodenum 8 h postinjection revealed one acetyl salicylate-(aspirin) treated cat with minor pinpoint erosions. None of the other cats in the three treatment groups had evidence of bleeding or ulceration. Serum biochemistry measurements of blood urea nitrogen (BUN), alanine transferase (ALT) and alkaline phosphatase (ALP) and complete blood counts (CBC) were not significantly altered from pretreatment values by the single dose of salicylate or carprofen (P < 0.05). Early and extended sample time points suggest that the pharmacokinetics of carprofen in the cat fit a 2-compartment model, with a long elimination half-life (t1/2) of 20.1 +/- 16.6 h, an area under the plasma concentration-time curve (AUC) of 637 (+/- 237) microgram.mL/h and a volume of distribution (Vdss) of 0.14 +/- 0.05 L/kg. Intravenously administered aspirin fit a 2-compartment model and had a long elimination half-life (t1/2) of 22.2 +/- 3.1 h, an AUC of 3824.2 +/- 506.7 microgram.mL/h and a volume of distribution (Vdss) of 0.17 +/- 0. 01 L/kg. PMID- 10849252 TI - Pulmonary vascular pressures of thoroughbred horses exercised 1, 2, 3 and 4 h after furosemide administration. AB - Furosemide premedication of horses 4 h prior to exercise significantly attenuates exercise-induced pulmonary capillary hypertension which may help diminish the severity of exercise-induced pulmonary haemorrhage. As pulmonary hemodynamic effects of furosemide may be mediated via a reduction in plasma volume (which is most pronounced 15-30 min postfurosemide administration, with plasma volume recovering thereafter), we hypothesized that administration of furosemide at intervals shorter than 4 h before exertion may be more effective in attenuating the exercise-induced rise in pulmonary capillary blood pressure. Thus, our objective was to determine whether furosemide-induced attenuation of exercise induced pulmonary arterial, capillary and venous hypertension would be enhanced when the drug is administered at intervals shorter than 4 h before exercise. Using established techniques, right atrial, and pulmonary arterial, capillary and wedge (venous) pressures were ascertained in seven healthy, sound, exercise trained Thoroughbred horses in a randomized split-plot experimental design. Measurements were made at rest and during exercise performed at maximal heart rate (217 +/- 3 beats/min) in the control (no medications) experiments and following furosemide administration (250 mg intravenously (i.v.)) at 1, 2, 3 and 4 h before exercise. Sequence of treatments was randomized and 7 days were allowed between experiments on each horse. Although furosemide administration in the four treatment groups caused only insignificant changes in the pulmonary arterial, capillary and wedge pressures of standing horses, furosemide-induced reduction in mean right atrial pressure achieved statistical significance in the 2 h postfurosemide experiments. In the control studies, exercise was attended by statistically significant increments in mean right atrial, as well as pulmonary arterial, capillary and wedge pressures. Although exercise in each of the four furosemide experiments was also attended by significant increments in right atrial as well as pulmonary vascular pressures, in the 1, 2 and 3 h postfurosemide experiments, mean right atrial pressure increased to a significantly lower value than in the control study. Exercise-induced changes in pulmonary vascular pressures in the 1 h postfurosemide experiments were not different from the pressures in the control study. There was a significant attenuation of exercise-induced pulmonary capillary and venous hypertension in the 2, 3 and 4 h postfurosemide experiments, but significant differences among these treatments were not found. Thus, these data did not support the contention that administration of furosemide at intervals shorter than 4 h before exercise is more effective in attenuating exercise-induced pulmonary capillary or venous hypertension in Thoroughbred horses. PMID- 10849253 TI - Ochratoxin A from a toxicological perspective. AB - Ochratoxin A (OTA) is a widespread mycotoxin which is produced mainly by the mould fungi Aspergillus ochraceus and Penicillum verrucosum during the storage of cereals, cereal products and other plant-derived products such as herbs, spices, grapes, etc. By carry over from mouldy fodder, ochratoxin A is also found in pork meat, offal and sausages containing pork blood. When ingested as a food contaminant, OTA is very persistent in human beings with a blood half-life of 35 days after a single oral dosage due to unfavourable elimination toxicokinetics. This renders the toxin among the most frequent mycotoxin contaminants in human blood in the EU, the US, Canada, and elsewhere, where it has been investigated. OTA is neither stored nor deposited in the body, but heterogeneous body distribution may impose serious damage to the kidneys. The toxin was classified a 2B cancer compound, being possibly carcinogenic for humans. It was among the strongest carcinogenic compounds in rats and mice. As the toxicological profile also includes teratogenesis, nephrotoxicity, and immunotoxicity, legislation authorities are currently discussing maximal residue levels (MRL) for OTA in various foodstuffs. In the present article arguments are presented which suggest an acceptable daily intake (ADI) of 1.5 ng OTA/kg body weight and a much lower MRL than 5 microgram OTA/kg cereals and cereal products as has been postulated by the EU commission. PMID- 10849254 TI - Pharmacokinetic study of dicloxacillin sodium following intravenous and intramuscular administration to pigs. PMID- 10849255 TI - Pharmacokinetics of fleroxacin in horses. PMID- 10849256 TI - Pharmacokinetics of a long-acting oxytetracycline-polyethylene glycol formulation in horses. PMID- 10849257 TI - Persistent efficacy of moxidectin against Sarcoptes scabiei in sheep. PMID- 10849258 TI - Biochemical and structural analysis of the NS5B RNA-dependent RNA polymerase of the hepatitis C virus. AB - Hepatitis C virus (HCV), the major causative agent of chronic and sporadic non-A, non-B hepatitis worldwide, is a distinct member of the Flaviviridae virus family. These viruses have in common a plus-strand RNA genome that is replicated in the cytoplasm of the infected cell via minus-strand RNA intermediates. Owing to the lack of reliable cell culture systems and convenient animal models for HCV, the mechanisms governing RNA replication are not known. As a first step towards the development of appropriate in vitro systems, we expressed the NS5B RNA-dependent RNA polymerase (RdRp) in insect cells, purified the protein to near homogeneity and studied its biochemical properties. It is a primer- and RNA template dependent RNA polymerase able to copy long heteropolymeric templates without additional viral or cellular cofactors. We determined the optimal reaction parameters, the kinetic constants and the substrate specificity of the enzyme, which turned out to be similar to those described for the 3D polymerase of poliovirus. By analysing a series of nucleosidic and non-nucleosidic compounds for their effect on RdRp activity, we found that ribavirin triphosphates have no inhibitory effect, providing direct experimental proof that the therapeutic effect observed in patients is not related to a direct inhibition of the viral polymerase. Finally, mutation analysis was performed to map the minimal NS5B sequence required for enzymatic activity and to identify the 'classical' polymerase motifs important for template and NTP binding and catalysis. PMID- 10849259 TI - Apoptotic cell death does not parallel other indicators of liver damage in chronic hepatitis C patients. AB - The mechanisms of hepatocyte damage and the events that lead to high rates of chronic liver disease in hepatitis C virus (HCV) infection remain unclear. Recent in vitro studies have suggested that the HCV core protein may disrupt specific signalling pathways of apoptosis. This prompted us to study patients with chronic HCV infection to: determine the extent of apoptosis in the liver; evaluate whether clinical and biochemical data are correlated with histological findings; and to investigate if apoptosis is related to the histological activity of the disease. Twelve patients with chronic hepatitis C were included in the study. Liver histology was scored by using the histological activity index (HAI) of Knodell et al. DNA fragmentation was assessed in liver tissue by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling (TUNEL) assay. Routine methods were used to determine serum markers of liver disease. Bile acids were measured in serum and liver by gas chromatography. Patients were placed, according to their HAI score, into group A (3.8 +/- 0.3) or group B (7.8 +/- 0.8) (P < 0.01). Liver enzymes tended to be higher in group B patients than in patients of group A. Levels of toxic bile acids in serum were greater in patients than in controls (P < 0.01). Chenodeoxycholic acid values were slightly higher in serum and liver of patients in group A. Liver biopsies with low HAI scores showed an increased rate of apoptosis (18.0 +/- 4.0 apoptotic cells per field) compared to those with higher HAI scores (6.6 +/- 2.1, P < 0.05) or to controls (3.5 +/- 0.4, P < 0.01). Hence, less severe liver disease, associated with lower histological grades and biochemistries, as well as increased levels of chenodeoxycholic acid, induces an expanded apoptotic response. The lower apoptotic rate in advanced liver disease may be associated with the high incidence of hepatocellular dysplasia/neoplasia. PMID- 10849260 TI - Methodological issues in papers on IFN therapy: time for reappraisal. AB - We conducted an analytical review of 194 full papers on interferon (IFN) therapy for chronic hepatitis C to evaluate current methodology (i.e. study design, criteria for evaluating the efficacy of therapy and predictors of response). Of the papers evaluated, 64 were randomized controlled trials (RCT), 40 were non randomized controlled trials (NRCT) and 90 were observational studies (OS). The methodological analysis was focused mainly on clinical trials. The number of patients enrolled in RCT was higher compared with the number enrolled in NRCT. Uniform enrolment criteria were used in less than 50% of the trials. Only 20% of RCT and 2.5% of NRCT used criteria for defining sample size. The response rate was calculated on an intention-to-treat basis in 36 of the RCT and in 14 of the NRCT. The outcome of treatment and the criteria employed to define the response to treatment were found to be far from standardized. In 51.5% of the RCT and 42.5% of the NRCT, normalization of alanine aminotransferase (ALT) level at the end of follow-up was the only marker of response studied. Only 57.6% of the trials considered histological evidence as an important outcome. Among the clinical trials, 71.1% evaluated predictors of good response to IFN therapy. In 51% of the OS, ALT normalization by the end of follow-up was the only criterion for defining response. In conclusion, to ensure a high level of reliability in comparing or combining the results of different studies, some basic general requirements must be followed when planning trials on antiviral therapy. PMID- 10849261 TI - Assessment of hepatitis C virus RNA and genotype from 6807 patients with chronic hepatitis C in the United States. AB - Hepatitis C virus (HCV) RNA status and HCV genotype have become important tools in the diagnosis and monitoring of therapy in chronic HCV infection. To establish a database with respect to HCV genotype and serum HCV RNA concentrations in chronic hepatitis C patients in the United States, we analysed 6807 chronic hepatitis C patients who had HCV RNA and HCV genotype tests conducted at a central laboratory. The HCV RNA concentration cut-off for the lower 25th percentile of this population (low titre) was 0.9 x 106 copies ml-1. The median HCV RNA concentration was 3.5 x 106 copies ml-1 and the cut-off for the upper 25th percentile (high titre) was 5 x 106 copies ml-1. Male patients had a median HCV RNA concentration of 3.9 x 106 copies ml-1, which was significantly higher than the median HCV RNA level for females (2.75 x 106 copies ml-1; P < 0.001). HCV genotype 1 was detected in 73% of patients; genotype 2 in 14%; genotype 3 in 8%; mixed genotype in 4%; and genotypes 4, 5 and 6 with a frequency of < 1%. Patients from the Northeast, Southeast and Midwest had significantly (P < 0.001) more infections with genotype 1 than patients from the Western and Southern regions. African-American patients were more likely to be infected with genotype 1 when compared with Caucasian, Hispanic or Asian Pacific Islanders (P < 0.001). Patients infected with HCV genotype 1 and mixed HCV genotypes had significantly higher serum HCV RNA concentrations when compared with HCV genotypes 2 and 3 (P < 0.001 for all comparisons). PMID- 10849262 TI - Comparison between three quantitative assays in patients with chronic hepatitis C and their relevance in the prediction of response to therapy. AB - To compare three quantitative assays measuring viral load in patients with chronic hepatitis C and to determine their value in predicting response to interferon (IFN) therapy, we analysed serum from 896 patients from eight European Centres using QUANTIPLEXtrade mark bDNA, MONITOR AMPLICORtrade mark and SUPERQUANTtrade mark assays. Analyses were performed on the same sample. Viral genotype was assessed using INNO-LiPA HCV II kits. Intercentre variations were observed that were related to the handling of specimens not processed and stored within 6 h of blood sampling. Among sera with optimal handling, a stronger correlation was observed between bDNA and SUPERQUANT (0.806) than between bDNA and MONITOR (0.677) and between MONITOR and SUPERQUANT (0.632). These discrepancies were greatest with genotype 2 (bDNA/SUPERQUANT= 0.772; bDNA/MONITOR=0. 456; SUPERQUANT/MONITOR= 0.299). This correlation was influenced by viraemia level and was better at lower viral loads. The proportion of sera with undetectable viral load was 15% with bDNA, 9.7% with MONITOR and 7.7% with SUPERQUANT. For the three measurements, the best cut-offs of sustained response to IFN treatment were located at their detection threshold. Among patients with viral load below the detection level, a sustained response was observed in 35% tested with bDNA, 38% with MONITOR and 80% with SUPERQUANT. Hence a stronger correlation was observed between bDNA and SUPERQUANT than between either of these assays and MONITOR. SUPERQUANT was the most sensitive assay and this greater sensitivity was associated with a better predictive value of treatment response. PMID- 10849263 TI - Pretreatment symptoms and dosing regimen predict side-effects of interferon therapy for hepatitis C. AB - We analysed data from a multicentre interferon (IFN) treatment trial to evaluate symptoms in patients with chronic hepatitis C and to identify factors that might predict development of debilitating IFN side-effects. Two hundred and twenty-two patients (120 US, 102 French) received 3 or 5 million units (MU) of IFN-alpha three times weekly (t.i.w.) for 3 months. Those who had detectable hepatitis C virus (HCV) RNA, as detected by the branched DNA signal amplification (bDNA) assay, at 3 months were intensified to daily therapy, while patients who were bDNA negative continued t.i.w. dosing for the subsequent 3 months of treatment. Symptoms were assessed at baseline, and adverse effects were evaluated at 6 months of therapy. Prior to treatment, the most common symptom that interfered with daily functioning was fatigue, occurring in 25% of patients. The frequency of debilitating fatigue, myalgia, arthralgia, headache, the presence of dry eyes and dry mouth, and use of antidepressant medication increased significantly from baseline to 6 months of IFN therapy (all P < 0.01). In multivariate analysis, the development of a debilitating side-effect at 6 months of treatment was associated with the presence of that symptom prior to therapy in all cases. Symptoms and adverse effects varied by gender and country. Compared with patients maintained on t.i.w. dosing, those who were dose intensified to daily IFN reported more debilitating fatigue, malaise, myalgia, arthralgia, fever, nausea, and headache, and the presence of dry mouth (all P < 0.05). In conclusion, patient characteristics, including pretreatment symptoms, gender and nationality, as well as daily IFN dosing are associated with the development of debilitating adverse effects on IFN therapy. PMID- 10849264 TI - Do undefined sources of hepatitis C transmission exist? The Greek study in General Practice. AB - A seroepidemiological study was carried out in 15 primary health care (PHC) centres in rural Greece to determine the prevalence of hepatitis C virus (HCV) in the surgeries of Greek General Practitioners (GPs) and to further clarify the transmission of hepatitis C in Greece. Serum samples were obtained from 1961 subjects (1259 females) aged >/= 15 years, who visited GP surgeries between July 1996 and February 1997 in 15 PHC centres located in three large Greek regions (Macedonia, Attika and Crete). Subjects who participated in the study fulfilled the following criteria: history of blood transfusion; hospital admission of > 7 days' duration without surgical or other intervention; use of intravenous drugs (current or previous); or women with a history of medical or paramedical abortion. Nearly 65% (1263 subjects) of the participants in this study reported hospital admission with a length of stay > 7 days. Antibodies to HCV (anti-HCV) were found in 67 participants (3. 5%), 41 of whom were females and 44 of whom were aged >/= 61 years. The highest prevalence (4.8%) of anti-HCV was found in Crete, and differences among the Greek regions were statistically significant (P < 0.05). Multivariate statistical analysis showed that in addition to regional differences, the following variables had a statistically significant effect on the prevalence of anti-HCV: history of dental surgery; use of intravenous drugs; hospital admission for > 7 days; and the high consumption of alcoholic drinks. Hence there is a significant variability in the prevalence of hepatitis C in well defined PHC areas of Greece. Several risk factors for acquiring HCV infection have been identified. Screening for HCV risk factors may enable Greek GPs to identify HCV-infected patients. PMID- 10849265 TI - A randomized controlled trial of kurorinone versus interferon-alpha2a treatment in patients with chronic hepatitis B. AB - It has recently been shown that a Chinese traditional medicine, kurorinone, extracted from Sophora Flavescens Ait, possesses antiviral properties. We evaluated the efficacy and safety of kurorinone treatment in patients with chronic hepatitis B. Ninety-four patients with abnormal alanine transaminase (ALT) levels and hepatitis B e antigen (HBeAg) and/or hepatitis B virus (HBV) DNA positivity were randomly assigned to receive either kurorinone 400 mg daily (45 patients) or 3 million units (MU) of interferon-alpha (IFN-alpha) (49 patients, daily for 1 month, every other day for 2 months) for 3 months. Patients were followed-up for 12 months. At baseline, both groups were comparable regarding age, gender and serological parameters. At the end of treatment, complete response (defined as ALT normalization and HBeAg and/or HBV DNA loss) occurred in 50% of the kurorinone group and in 61.3% of the IFN-alpha-treated group (P > NS). At the end of the 12-month follow-up period, a complete response (sustained response) occurred in 26.7-36.7% of kurorinone-treated patients with moderate or mild liver damage and in 44.4-46.7% of IFN-alpha-treated patients with similar liver injury. In kurorinone- as well as in IFN-alpha-treated patients, there was no statistical significant difference with respect to complete response rates between HBeAg-positive and hepatitis B e antibody-positive subgroups. Kurorinone had no untoward side-effects except for local pain at injection sites. The results of this trial suggest that kurorinone is able to inhibit HBV replication and improve disease remission in patients with chronic hepatitis B. PMID- 10849266 TI - Transfusion-transmissible virus and hepatocellular carcinoma: a case-control study. AB - Although transfusion-transmissible virus (TTV) is often present in the serum of patients with acute and chronic non-A-C liver diseases, its hepatotropism, pathogenicity to the liver and hepatocarcinogenicity have not been proven. We used a case-control format to compare the prevalence of TTV infection among 148 southern African Blacks with hepatocellular carcinoma and 148 matched hospital based controls, and to test for possible interactive effects between this virus and hepatitis B virus (HBV) and hepatitis C virus (HCV) in the development of the tumour. We also determined the prevalence of TTV in 988 blood donors in Gauteng province of South Africa. The presence of TTV DNA in serum samples was detected by using the polymerase chain reaction, Southern hybridization and nucleotide sequencing. Individuals infected with TTV did not have an increased risk of developing hepatocellular carcinoma (relative risk 1.1; 95% confidence limits 0.5 2.4). Moreover, co-infection with TTV did not further increase the risk of tumour development in patients chronically infected with HBV and/or HCV. TTV was present in the serum of 2.2% of blood donors: 4.0% in Black and 1.5% in White donors. We conclude that TTV is unrelated to the development of hepatocellular carcinoma in Black Africans. PMID- 10849267 TI - Interferon treatment with or without oral ganciclovir in HBeAg-negative chronic hepatitis B: a randomized study. AB - The treatment of HBeAg-negative chronic hepatitis B with alpha interferon alone is unsatisfactory. We evaluated the efficacy of combined administration of interferon-a2a (IFN) with oral ganciclovir, a nucleoside analogue. Forty patients with hepatitis B virus (HBV)-DNA-positive/HBeAg-negative chronic hepatitis B, were randomized to receive 4.5 MU IFN thrice weekly, subcutaneously, alone or in combination with 3 g ganciclovir per os daily for 26 weeks and followed for 12 months after treatment. Mean serum HBV-DNA levels decreased by 4.0 log10 in the combination group (from 5 x 106 to 4.8 x 102 copies/ml) and by 2.2 log10 in the interferon group (from 8 x 106 to 4.8 x 104 copies/ml) by quantitative polymerase chain reaction (PCR). HBV-DNA became undetectable in 11 of 20 (55%) and in three of 20 (15%) patients in the two groups, respectively (P=0.02). The alanine aminotransferase levels became normal in all patients receiving combination therapy, compared to 75% of those in the interferon group. After cessation of therapy, HBV-DNA increased or reappeared in all patients with 85% also relapsing biochemically. One year after treatment, three patients in each group (15%) remained in sustained biochemical remission with very low serum HBV-DNA levels (median 15 700 copies/ml). We conclude that, in HBeAg-negative chronic hepatitis B, 6-month combination therapy with oral ganciclovir and IFN is associated with complete biochemical remission in all treated patients and a 4 log10 decrease in serum HBV-DNA levels. The end of treatment efficacy of this combination scheme is far super- ior to that of IFN monotherapy but sustained responses are few. Further studies are warranted to evaluate the efficacy of prolonged combination schemes with nucleoside analogues and IFN, compared to IFN monotherapy. PMID- 10849268 TI - An efficient extraction method from blood clots for studies requiring both host and viral DNA. AB - The clot from blood is usually discarded after the collection of serum. Yet, it contains nucleated white blood cells and substantial serum. Here, we have compared four methods to enable quick and efficient extraction of human genomic and viral DNA from clotted blood. Two of these methods, a phenol-based in-house method and Tripure isolation reagent, only achieved a low polymerase chain reaction (PCR) yield. In contrast, the QIAamp blood kit and the High Pure Viral Nucleic Acid kit were equally efficient, with similar sensitivity to serum for extraction of viral DNA. PMID- 10849269 TI - Development of 16S rDNA-based PCR assay for detecting centipeda periodontii and selenomonas sputigena. AB - To detect oral motile bacteria directly from dental plaque, specific PCR primers for Centipeda periodontii and Selenomonas sputigena were designed based on the sequence analysis of their 16S rDNA. The primers were specific and sensitive enough to amplify DNA fragments from the available oral bacteria. The detection limit was fewer than 10 bacterial cells per sample. It was also possible to detect these bacteria in dental plaque. The prevalence of these bacteria varied in each sample. The specific primers designed in this study may clarify the epidemiology of periodontal disease. PMID- 10849270 TI - Comparative analysis of denaturing gradient gel electrophoresis and temporal temperature gradient gel electrophoresis in separating Escherichia coli uidA amplicons differing in single base substitutions. AB - A set of Escherichia coli freshwater isolates was chosen to compare the effectiveness of denaturing gradient gel electrophoresis (DGGE) vs temporal temperature gradient gel electrophoresis (TTGE) for separating homologous amplicons from the respective uidA region differing in one to seven single base substitutions. Both methods revealed congruent results but DGGE showed a five to eight times higher spatial separation of the uidA amplicons as compared with TTGE, although the experiments were performed at comparable denaturing gradients. In contrast to TTGE, DGGE displayed clear and focused bands. The results strongly indicated a significantly higher discrimination efficiency of the spatial chemical denaturing gradient as compared with the temporal temperature denaturing gradient for separating the uidA amplicons. Denaturing gradient gel electrophoresis proved to be highly efficient in the differentiation of E. coli uidA sequence types. PMID- 10849271 TI - Solar photo-oxidative disinfection of drinking water: preliminary field observations. AB - The feasibility of using solar photo-oxidation to inactivate faecal bacterial contaminants in drinking water has been evaluated under field conditions in India and South Africa. Freshly drawn samples from all six test water sources were low in dissolved oxygen, at 13-40% of the air saturation value. However, vigorous mixing followed by exposure to full-strength sunlight in transparent plastic containers (1-25 l capacity) caused a rapid decrease in the counts of faecal indicator bacteria, giving complete inactivation within 3-6 h, with no evidence of reactivation. These results demonstrate that solar photo-oxidation may provide a practical, low-cost approach to the improvement of drinking water quality in developing countries with consistently sunny climates. PMID- 10849272 TI - Do stresses encountered during the smoked salmon process influence the survival of the spoiling bacterium Shewanella putrefaciens? AB - The influence of different treatments (i.e. cold, NaCl, phenol and anaerobiosis) encountered during the smoked salmon process was studied by analysing the survival capacity of two Shewanella putrefaciens strains (CIP 69.29 and J13.1). Our results indicated that only the salt stress was critical for the survival of S. putrefaciens. Nevertheless, both strains of S. putrefaciens grown at low temperatures developed a cross-protection to a lethal NaCl treatment. To our knowledge, this is the first report showing that growth at low temperatures induces cross-protection towards NaCl challenge. Moreover, we observed a significant sensitization by moderate salt concentration to a phenol treatment. From our combined data, we propose that control of S. putrefaciens proliferation could take place during the smoked salmon process rather than during storage of the final product. PMID- 10849273 TI - Adenylate kinase amplification of ATP bioluminescence for hygiene monitoring in the food and beverage industry. AB - Fourteen food residues, Escherichia coli O157:H7 and Staphylococcus aureus on stainless steel surfaces were detected using a combined assay with adenylate kinase as a cellular marker and ATP bioluminescence. The limit of sensitivity ranged from 0.02 to 708 microg for minced meat and broccoli, respectively. Both methods gave the same detection limit (105 cfu) for E. coli and Staph. aureus on stainless steel surfaces. The combined adenylate kinase-ATP assay is applicable to monitor the hygiene of work surfaces, especially those prone to contamination by meat and vegetable residues. PMID- 10849274 TI - The differentiation of Carnobacterium divergens using the random amplification of polymorphic DNA polymerase chain reaction technique. AB - The potential of the randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) technique to differentiate Carnobacterium divergens from other members of the genus Carnobacterium was examined. A numerical analysis of the genomic profiles obtained demonstrated that it was possible to differentiate the C. divergens strains from other Carnobacterium strains using this technique. The heterogeneity observed in the representatives of the species C. piscicola adds further weight to the suggestion in other taxonomic studies that subspecies of this species exist. PMID- 10849275 TI - Stimulatory effect of honey on multiplication of lactic acid bacteria under in vitro and in vivo conditions. AB - The effect of honey and sucrose on lactic acid bacteria in vitro and in rat gut was studied to determine whether these organisms were affected differently by honey compared with sucrose. Under in vitro conditions, the number of Lactobacillus acidophilus and Lactobacillus plantarum counts increased 10-100 fold in the presence of honey compared with sucrose. Feeding of honey to rats also resulted in significant increase in counts of lactic acid bacteria. Although there was no significant difference in the counts of lactic acid bacteria in the small and large intestines of different groups, the honey-fed group showed a significant increase (P<0.05) in counts over the control and sucrose-fed animals. The results support the fact that consumption of honey has a beneficial effect on the physiological constitution of animals fed with it. PMID- 10849276 TI - Survey for psychrotrophic bacterial pathogens in minimally processed lettuce. AB - A total of 120 minimally processed, cut and packaged lettuce samples were purchased from retail supermarkets or provided by a salad production facility over an 8-month period. The samples were tested for total aerobic plate counts and for the presence of potentially pathogenic species belonging to the genera of Listeria, Aeromonas and Yersinia. The aerobic plate counts ranged from 103 to 109 colony forming units (cfu) g-1. Most samples (76%) contained between 105 and 107 cfu g-1 total aerobic bacteria. Listeria monocytogenes was isolated from three samples, Aeromonas hydrophila or Aeromonas caviae from 66 samples, and Yersinia enterocolitica from 71 samples. The pathogenic potential of Y. enterocolitica isolates was determined by screening for an array of biochemical, serological and genetic traits (heat-stable enterotoxin gene, the attachment and invasion gene locus, the invasin gene locus and the virulence plasmid). The Y. enterocolitica isolates lacked many of the phenotypic and genetic markers associated with virulence in primary pathogenic strains. As the roles of the reputed virulence factors of Aeromonas spp. in human infection are uncertain, the pathogenic potential of the Aeromonas isolates in lettuce remains unclear. PMID- 10849277 TI - Heat inactivation of Salmonella typhimurium DT104 in beef as affected by fat content. AB - The heat resistance of an eight-strain cocktail of Salmonella typhimurium DT104 was determined at 58-65 degrees C in beef containing 7, 12, 18 or 24% fat. Inoculated beef was packaged in bags completely immersed in a circulating water bath and held at 58, 60, 62.5 and 65 degrees C for a predetermined length of time. The surviving cell population was enumerated by spiral plating heat-treated samples onto tryptic soy agar supplemented with 0.6% yeast extract and 1% sodium pyruvate. Preliminary studies on thermal inactivation of the Salmonellae isolates in chicken broth indicated no correlation between heat resistance and origin of the isolates. While linear survival curves were observed in chicken broth, inactivation kinetics in beef showed deviations from the first order kinetics, represented by an initial lag period or shoulder before any death occurred with time. Overall, increased fat levels in beef resulted in longer lag periods and lower D-values, suggesting that the lag periods must be taken into account and added to the D-values for calculating the time required at a specific temperature for achieving a specific lethality for Salm. typhimurium DT104 in beef. Thermal death times from this study will assist the retail food industry to design cooking regimes that ensure safety of beef contaminated with Salm. typhimurium DT104. PMID- 10849278 TI - A comparison of the bioscreen method and microscopy for the determination of lag times of individual cells of listeria monocytogenes. AB - Lag phase durations (tLag) of individual Listeria monocytogenes cells were analysed using the NightOwl Molecular Imaging System, and results were compared with mean individual cell lag times (tL) obtained from the detection time (td) method using Bioscreen. With Bioscreen, an average tL of 6.39+/-0.89 h was obtained from five separate experiments. With the NightOwl method, an average tLag of 2.73+/-0.06 h was obtained from three experiments consisting of eight total replicates. Lag values from the NightOwl and Bioscreen are related by the equation: tLag = tL + DT, where DT is the doubling time. The equivalent tLag mean value for the Bioscreen method was 7.11+/-0.84 h. Individual lag times measured by both methods were normally distributed (r2 for Bioscreen and NightOwl ranged from 0.951 to 0.999 and from 0.884 to 0.982, respectively). The results suggest that the NightOwl method can provide accurate estimates of individual cell lag times, which will facilitate the development of combined discrete continuous models for bacterial growth. PMID- 10849279 TI - Anti-adhesive activity of sulphated exopolysaccharides of microalgae on attachment of red sore disease-associated bacteria and helicobacter pylori to tissue culture cells. AB - Because of the affinity of certain bacterial species for sulphated glycoconjugates exposed on the epithelial cells of susceptible hosts, we hypothesized that sulphated exopolysaccharides of microalgae can be used in anti adhesive therapies against bacterial infections, both in cold- and warm-blooded animals. In this study we found that adhesion of the human pathogen Helicobacter pylori to the HeLa S3 cell line, and adhesion of the fish pathogens Vibrio campbellii, V. ordalii, Streptococcus saprophyticus, and Aeromonas veronii to spotted sand bass primary tissue culture cells, can be effectively blocked with the various sulphated exopolysaccharides used. PMID- 10849280 TI - Establishing a campylobacter-free pig population through a top-down approach. AB - Fattening pigs are often infected with campylobacter. To eliminate campylobacter from the pig population, a top-down approach, involving the breeding and reproduction farms, seems appropriate. In order to investigate the effectiveness of a top-down approach, sows' faeces from the following farms were analysed for the presence of campylobacter: one specific pathogen free (SPF) farm, three top breeding farms with no connection with SPF breeding, and a breeding farm repopulated with SPF sows after a period of vacancy (farm 5). The faeces samples from the SPF farm were free from campylobacter. The three top-breeding farms provided faeces samples which were 98% positive for campylobacter. However, only 22% of the faeces samples from farm 5 were positive for campylobacter. In a period of 20 months, the percentage of sows infected with campylobacter on farm 5 did not significantly increase. Genetic typing with ERIC-PCR and RFLP of campylobacter isolates from one of the top-breeding farms and from farm 5 showed a high diversity of campylobacter types. The results suggest that a campylobacter free pig population can be established in breeding farms by combining a top-down approach (campylobacter-free top-breeding farms) with a strict regime of hygiene management. PMID- 10849281 TI - Genetic structure of Camellia japonica L. in an old-growth evergreen forest, Tsushima, Japan. AB - The spatial genetic structure of Camellia japonica was investigated, using microsatellite markers, in a 4-ha permanent plot within an old-growth forest. Spatial distribution of individuals was also assessed to obtain an insight into spatial relationships between individuals and alleles. Morisita's index of dispersion showed that 518 C. japonica individuals in the plot were clumped, and Moran's I spatial autocorrelation coefficient revealed weak genetic structure, indicating a low level of allele clustering. Average I correlograms showed that there was stronger genetic structure over short-distance classes. The clumped distribution of individuals and the positive autocorrelation over short-distance classes may result from the limited seed dispersal and microsite heterogeneity of the stand, while the genetic structure may be weakened by overlapping seed shadow and extensive pollen flow, mediated by animal vectors, and the high outcrossing rate found in C. japonica. PMID- 10849282 TI - Phylogeographical pattern correlates with pliocene mountain building in the alpine scree weta (Orthoptera, anostostomatidae). AB - Most research on the biological effects of Pleistocene glaciation and refugia has been undertaken in the northern hemisphere and focuses on lowland taxa. Using single-strand conformation polymorphism (SSCP) analysis and sequencing of mitochondrial cytochrome oxidase I, we explored the intraspecific phylogeography of a flightless orthopteran (the alpine scree weta, Deinacrida connectens) that is adapted to the alpine zone of South Island, New Zealand. We found that several mountain ranges and regions had their own reciprocally monophyletic, deeply differentiated lineages. Corrected genetic distance among lineages was 8.4% (Kimura 2-parameter [K2P]) / 13% (GTR + I + Gamma), whereas within-lineage distances were only 2.8% (K2P) / 3.2% (GTR + I + Gamma). We propose a model to explain this phylogeographical structure, which links the radiation of D. connectens to Pliocene mountain building, and maintenance of this structure through the combined effects of mountain-top isolation during Pleistocene interglacials and ice barriers to dispersal during glacials. PMID- 10849283 TI - Phylogeography and genetic structure of northern populations of the yellow warbler (Dendroica petechia). AB - Phylogeographic patterns of intraspecific variation can provide insights into the population-level processes responsible for speciation and yield information useful for conservation purposes. To examine phylogeography and population structure in a migratory passerine bird at both continental and regional geographical scales, we analysed 344 bp of mitochondrial DNA (mtDNA) control region sequence from 155 yellow warblers (Dendroica petechia) collected from seven locations across Canada and from Alaska. There is a major subdivision between eastern (Manitoba to Newfoundland) and western (Alaska and British Columbia) populations which appears to have developed during the recent Pleistocene. Some localities within these two regions also differ significantly in their genetic composition, suggesting further subdivision on a regional geographical scale. Eastern and western birds form distinct phylogeographic entities and the clustering of all western haplotypes with two eastern haplotypes suggests that the western haplotypes may be derived from an eastern lineage. Analyses based on coalescent models support this explanation for the origin of western haplotypes. These results are consistent with important features of Mengel's model of warbler diversification. From a conservation perspective they also suggest that individual populations of migrant birds may form demographically isolated management units on a smaller scale than previously appreciated. PMID- 10849284 TI - A population genetic analysis of migration: the case of the noctule bat (Nyctalus noctula). AB - Although rarely assessed, the population genetics of hibernating colonies can help to understand some aspects of population structure, even when samples from nursery or mating colonies are not available, or in studies of migration when both types of samples are available and can be compared. Here we illustrate both points in a survey of mitochondrial DNA (mtDNA) control region sequences used to study the population genetics of hibernating colonies of a migrating species, the noctule bat (Nyctalus noctula). Lacking samples from Scandinavian nursery colonies, we use a North European hibernacula to suggest that Scandinavian populations are isolated from Central and East European colonies. Then, we compare genetic diversities of nursery and hibernating colonies. We find a significantly higher haplotype diversity in hibernacula, confirming that they consist of individuals from different nursery colonies. Finally, we show that pairwise comparisons of the haplotype frequencies of nursery and hibernating colonies contain some information on the migration direction of the noctule bat. PMID- 10849285 TI - Mitochondrial haplotype diversity among Portuguese brown trout Salmo trutta L. populations: relevance to the post-pleistocene recolonization of northern Europe. AB - Mitochondrial haplotype diversity in seven Portuguese populations of brown trout, Salmo trutta L., was investigated by sequencing the 5' end of the mitochondrial DNA (mtDNA) control region. Five new haplotypes were described for this species, each two to three mutational steps distant from the common north Atlantic haplotype. Significant population subdivision of mtDNA haplotypes was also apparent. Based on these results, as well as on published data describing the distribution of both mtDNA haplotypes and allozyme alleles throughout Europe, the postglacial recolonization of northern Europe was re-evaluated. It is argued that the available data do not support the contribution of two major glacial refugia (southwest Atlantic and Ponto-Caspian Basin) to this postglacial recolonization, as proposed in a recently published model. The unique genetic architecture of Portuguese brown trout within the Atlantic-basin clade of this species represents a highly valuable genetic resource that should be protected from introgression with nonendemic strains of hatchery fish. PMID- 10849286 TI - Postglacial recolonization routes for Picea abies K. in Italy as suggested by the analysis of sequence-characterized amplified region (SCAR) markers. AB - The routes through which Norway spruce recolonized the Alps after the last ice age were investigated at the genetic level. Seven populations along the Alpine range plus one Apennine population were characterized for seven sequence characterized amplified region (SCAR) loci, detecting an overall FST = 0.118. This rather high value for forest species reflects an uneven distribution of genetic variability, and was analysed through different statistical methods. Alternative hypotheses were tested under the isolation-by-distance model and using the analysis of molecular variance (AMOVA) frame. We conclude that the hypothesis of the existence of a glacial refugium in the Apennines should be rejected, while a putative relict population is identified in the Maritime Alps. The Alpine range of Norway spruce appears to be split in two parts across a north south line. The results are discussed in comparison with data based on morphological markers, isozymes, chloroplast microsatellites and mitochondrial markers. PMID- 10849287 TI - Phylogeography of the bullhead Cottus gobio (Pisces: Teleostei: Cottidae) suggests a pre-pleistocene origin of the major central European populations. AB - The bullhead Cottus gobio is a small, bottom-dwelling fish consisting of populations that have not been subject to transplantations or artificial stocking. It is therefore an ideal model species for studying the colonization history of central European freshwater systems, in particular with respect to the possible influences of the Pleistocene glaciation cycles. We sampled Cottus populations across most of its distribution range, with a special emphasis on southern Germany where the major European drainage systems are in closest contact. Mitochondrial D-loop sequencing of more than 400 specimens and phylogenetic network analysis allowed us to draw a detailed picture of the colonization of Europe by C. gobio. Moreover, the molecular distances between the haplotypes enabled us to infer an approximate time frame for the origin of the various populations. The founder population of C. gobio stems apparently from the Paratethys and invaded Europe in the Pliocene. From there, the first colonization into central Europe occurred via the ancient lower Danube, with a separate colonization of the eastern European territories. During the late Pliocene, one of the central European populations must have reached the North Sea in a second step after which it then started to colonize the Atlantic drainages via coastal lines. Accordingly, we found very distinct populations in the upper and lower Rhine, which can be explained by the fact that the lower Rhine was disconnected from the upper Rhine until approximately 1 million years ago (Ma). More closely related, but still distinct, populations were found in the Elbe, the Main and the upper Danube, all presumably of Pleistocene origin. Intriguingly, they have largely maintained their population identity, despite the strong disturbance caused by the glaciation cycles in these areas. On the other hand, a mixing of populations during postglacial recolonization could be detected in the lower Rhine and its tributaries. However, the general pattern that emerges from our analysis suggests that the glaciation cycles did not have a major impact on the general population structure of C. gobio in central Europe. PMID- 10849288 TI - Molecular ecology and biological control: the mating system of a marsupial pest. AB - Many studies in molecular ecology have focused on the use of repeat DNA markers to determine the nature of mating systems in a wide variety of animal species. Whilst these studies typically have focused on important issues such as the evolutionary consequences of fitness variation among males, genetic studies of mating systems are potentially also important because they can generate information of significance to wider issues in wildlife management. For example, genetically modified, sexually transmitted viral diseases have been suggested as potential agents for the control of vertebrate pest species. An understanding of the epidemiology of such agents requires an intimate knowledge of the sexual contact rates between individuals of the target species. Here, we report the use of minisatellite DNA profiling to reveal the mating system in two New Zealand populations of the introduced Australian brushtail possum. The brushtail possum is New Zealand's most important mammalian pest and a species for which control by a sexually transmitted immunocontraceptive has been proposed. Encouragingly, we report considerable variation in the reproductive success of males at both study sites, with one male siring offspring from four females in one year (mean no. of offspring/reproductively successful male/year at the two sites is 1.95-2.15), while many sired none. This bias in the pattern of reproductive success among males will probably facilitate the spread of an immunocontraceptive agent and thereby increase the power of this approach to biological control. PMID- 10849289 TI - Do hornets have zombie workers? AB - Colonies of the European hornet, Vespa crabro, are typically founded by a single queen mated to a single male. From the resulting colony relatedness pattern we predicted strong worker-queen conflict over male production where both the workers and the queen attempt to produce the colony's males. To test for this conflict, male production was studied in 15 hornet nests using a combination of DNA microsatellite analysis (282 males), worker ovary dissections (500 workers from eight nests) and 50 h of observation (four nests). In contrast to our prediction, the data show that hornet males are queens' sons, that workers never attempt to lay eggs, rarely have activated ovaries, and that there is no direct aggression between the queen and the workers. This contrasts with other data for vespine wasps, which support relatedness predictions. Dolichovespula arenaria has the same kin structure as V. crabro and workers produce males in many colonies. The similarity between these two species makes it difficult to explain why workers do not reproduce in V. crabro. Self-restraint is expected if worker reproduction significantly reduces colony productivity but there is no obvious reason why this should be important to V. crabro but not to D. arenaria. Alternatively, queen control may be important. The absence of expressed queen worker conflict rules out physical control. Indirect pheromonal control is a possibility and is supported by the occurrence of royal courts and queen pheromone in Vespa but not Dolichovespula. Pheromonal queen control is considered evolutionarily unstable, but could result from a queen-worker arms race over reproductive control in which the queen is ahead. The genetic data also revealed diploid males in one colony, the first example in the vespine wasps, and two colonies with double matrilines, suggesting that occasional usurpation by spring queens occurs. PMID- 10849290 TI - Microsatellite analysis of kinkajou social organization. AB - Kinkajou social groups generally consist of one adult female, two males, one subadult and one juvenile. Based on analysis of variation in 11 microsatellite loci, we assess the degree of kinship within and between four social groups totaling 25 kinkajous. We use exclusion and likelihood analyses to assign parents for seven of the eight offspring sampled, five with >/= 95% certainty, and two with >/= 80% certainty. Five of six identified sires of group offspring came from the same social group as the mother and pup. Adult males and females within a group were unrelated and subadults and juveniles were offspring of the group adults, suggesting a family structure. All five identified paternities within a social group were by the dominant male of the group. However, this copulation asymmetry does not necessarily reflect cooperation due to kinship ties between the two adult males within a group as one of two adult male pairs sampled was unrelated. Neighbouring male kinkajous were more closely related to each other than neighbouring female kinkajous, suggesting that females disperse more often or farther than males. PMID- 10849291 TI - Genetic analysis of a documented population bottleneck: introduced Bennett's wallabies (Macropus rufogriseus rufogriseus) in New Zealand. AB - Few bottlenecks of wild populations are sufficiently well-documented to constitute models for testing theories about the impact of bottlenecks on genetic variation, and subsequent population persistence. Relevant details of the Bennett's wallaby (Macropus rufogriseus rufogriseus) introduction into New Zealand were recorded (founder number, source and approximate bottleneck duration) and suggest this may provide a rare opportunity to examine the efficacy of tests designed to detect recent bottlenecks in wild populations. We first assessed the accuracy of historic accounts of the introduction using genetic diversity detected in mitochondrial DNA (mtDNA) and at five microsatellite loci. Phylogenetic analyses of mtDNA D-loop sequence haplotypes were consistent with the reported origin of the founders as Tasmania, rather than one of the Bass Strait islands in which Bennett's wallabies are also found. Microsatellite allele frequencies from the Tasmanian source population were then used to seed bottleneck simulations encompassing varying sizes and numbers of generations, in order to assess the severity of bottleneck consistent with diversity observed in the New Zealand population. The results suggested that the founder number was unlikely to have been as small as the three animals suggested by the account of the introduction. Nonetheless, the bottleneck was probably severe; in the range of three to five pairs of wallabies for one to three generations. It resulted in significantly reduced levels of allelic diversity and heterozygosity relative to the source population. This bottleneck is only detectable under the infinite allele model (IAM) and not under the stepwise mutation model (SMM) or the two phase model (TPM), and possible explanations for this are discussed. PMID- 10849292 TI - Reciprocal hybrid formation of Spartina in San Francisco Bay. AB - Diversity in the tRNALEU1 intron of the chloroplast genome of Spartina was used to study hybridization of native California cordgrass, Spartina foliosa, with S. alterniflora, introduced to San Francisco Bay approximately 25 years ago. We sequenced 544 bases of the tRNALEU1 intron and found three polymorphic sites, a pyrimidine transition at site 126 and transversions at sites 382 and 430. Spartina from outside of San Francisco Bay, where hybridization between these species is impossible, gave cpDNA genotypes of the parental species. S. foliosa had a single chloroplast haplotype, CCT, and this was unique to California cordgrass. S. alterniflora from the native range along the Atlantic coast of North America had three chloroplast haplotypes, CAT, TAA, and TAT. Hybrids were discriminated by random amplified polymorphic DNA (RAPD) phenotypes developed in a previous study. We found one hybrid that contained a cpDNA haplotype unknown in either parental species (TCT). The most significant finding was that hybridization proceeds in both directions, assuming maternal inheritance of cpDNA; 26 of the 36 hybrid Spartina plants from San Francisco Bay contained the S. foliosa haplotype, nine contained haplotypes of the invading S. alterniflora, and one had the cpDNA of unknown origin. Furthermore, cpDNA of both parental species was distributed throughout the broad range of RAPD phenotypes, suggesting ongoing contributions to the hybrid swarm from both. The preponderance of S. foliosa cpDNA has entered the hybrid swarm indirectly, we propose, from F1s that backcross to S. foliosa. Flowering of the native precedes by several weeks that of the invading species, with little overlap between the two. Thus, F1 hybrids would be rare and sired by the last S. foliosa pollen upon the first S. alterniflora stigmas. The native species produces little pollen and this has low viability. An intermediate flowering time of hybrids as well as pollen that is more vigourous and abundant than that of the native species would predispose F1s to high fitness in a vast sea of native ovules. Thus, spread of hybrids to other S. foliosa marshes could be an even greater threat to the native species than introductions of alien S. alterniflora. PMID- 10849293 TI - Genetic subdivision, glacial refugia and postglacial recolonization in the golden striped salamander, Chioglossa lusitanica (Amphibia: urodela). AB - The golden-striped salamander (Chioglossa lusitanica) is an ecologically specialized species, endemic to north-western Iberia. Patterns of genetic variation were assessed at seven polymorphic enzyme loci and one mitochondrial DNA (mtDNA) marker (cytochrome b) in 17 populations across its range. Estimates of enzyme genetic diversity revealed a high degree of genetic subdivision (FST = 0.68), mainly attributable to the existence of two groups of populations. The groups were located, respectively, north and south of the Mondego River, indicating that this river coincided with a major historical barrier to gene flow. A significant decrease in genetic variability from the Mondego northwards was associated with the Douro and Minho rivers. mtDNA sequence variation revealed a congruent pattern of two haplotype groups (d = 2.2%), with a geographical distribution resembling that of allozymes. The pattern and depth of genetic variation is consistent with the following hypotheses: (i) subdivision of an ancestral range of the species prior to the middle Pleistocene; (ii) secondary contact between populations representing historical refugia; (iii) relatively recent range expansion giving rise to the northern part of the species range; and (iv) loss of genetic variation through founder effects during range expansion across major rivers. PMID- 10849294 TI - Are recession populations of the desert locust (Schistocerca gregaria) remnants of past swarms? AB - The desert locust (Schistocerca gregaria) undergoes crowding-induced phase transformation from solitary to gregarious, which involves changes in behaviour, colour, development, morphometry, fecundity and endocrine physiology. During recession, solitary locusts persist in the central, drier part of the species' range in small pocket populations that are prone to extinction. During the intermittent upsurges and the subsequent plagues, gregarious swarms attain huge population size and invade a vast area causing major damage to agriculture. A highly variable nuclear DNA marker, a noncoding 3' end fragment of an antennapedia-class homeobox gene, was screened in locust samples from Eritrea. Despite the homogenizing potential of plague swarms, the last of which was in 1986-89 and originated in this region, the population genetic structure of solitary phase locusts along the Red Sea coast of Eritrea revealed significant divergence. The pattern of divergence indicated that the invasion of the western and northern plains in the summer of 1995 may not, as reported then, have originated in eastern Chad or western Sudan. A number of interrelated hypotheses have been presented to explain the observed genetic heterogeneity between the sampled populations. We conclude, with caution due to the limited sample sizes, that: (i) geographical isolation between breeding sites during plagues and recession; (ii) the marked differences in the flight behaviour of plague swarms and recession populations; (iii) possible failure of gregarious locusts to solitarize and re-establish in recession areas; and (iv) the effect of repeated extinction and recolonization in the meta-population contribute to the maintenance of the genetic structure of recession populations. Potentially productive future research has been identified. PMID- 10849295 TI - Mitochondrial DNA (mtDNA) reveals that female Bechstein's bats live in closed societies. AB - We present a microgeographic analysis of mitochondrial DNA (mtDNA) in Bechstein's bats using three sources of control region sequence variability, including a novel mtDNA microsatellite, to assess individual relatedness both within and among 10 maternity colonies. Comparison of marker variability among 268 adult females revealed little genetic variability within each colony. However, most colonies were clearly distinguished by colony-specific mitochondrial haplotypes (total n = 28). Low intracolony variability and strong haplotype segregation among colonies, was reflected by an extraordinary high FST of 0.68, indicating a very low intercolony dispersal rate of approximately one female in five generations. Haplotype distribution among 18 solitary males showed that males frequently disperse between colony locations, indicating the absence of dispersal barriers. Bechstein's bat maternity colonies are thus closed groups that comprise 20-40 females probably belonging to only one or, at most, two matrilines. The genetic population structure of Bechstein's bats is in agreement with the hypothesis that females seek familiar and, at least, partially related cooperation partners for raising their young. Alternatively strong philopatry might reflect the importance of profound roost or habitat knowledge for successful reproduction in female Bechstein's bats. PMID- 10849296 TI - A retrospective assessment of the accuracy of the paternity inference program CERVUS. AB - CERVUS is a Windows-based software package written to infer paternity in natural populations. It offers advantages over exclusionary-based methods of paternity inference in that multiple nonexcluded males can be statistically distinguished, laboratory typing error is considered and statistical confidence is determined for assigned paternities through simulation. In this study we use a panel of 84 microsatellite markers to retrospectively determine the accuracy of statistical confidence when CERVUS was used to infer paternity in a population of red deer (Cervus elaphus). The actual confidence of CERVUS-assigned paternities was not significantly different from that predicted by simulation. PMID- 10849297 TI - Matrilineal history of the endangered Cape Sable seaside sparrow inferred from mitochondrial DNA polymorphism. AB - Restriction analyses were conducted on mitochondrial DNA (mtDNA) amplified by long-PCR from an endangered bird, the Cape Sable seaside sparrow. The first of several successful mtDNA amplifications was accomplished using the partially digested tissue remains of a transmitter-monitored bird retrieved from the gut of a snake. As many as 91 mtDNA restriction fragments produced by 18 endonucleases were compared in this and four other Cape Sable specimens against mtDNA similarly amplified by long-PCR from other taxonomic forms in the seaside sparrow complex. Results indicate that the Cape Sable birds belong to an 'Atlantic' matrilineal clade, and are highly divergent from other seaside sparrows along the Gulf of Mexico. PMID- 10849298 TI - A simple method for the detection of size homoplasy among amplified fragment length polymorphism fragments. PMID- 10849299 TI - Polymorphism at trinucleotide microsatellite loci in the subterranean termite Reticulitermes flavipes. PMID- 10849300 TI - Isolation and characterization of novel polymorphic tetra-nucleotide microsatellite markers from the blue marlin, Makaira nigricans. PMID- 10849301 TI - Isolation and characterization of 11 microsatellite loci in Atlantic halibut (Hippoglossus hippoglossus L.). PMID- 10849302 TI - Microsatellite DNA markers for the study of Allegheny woodrat (Neotoma magister) populations and cross-species amplification in the genus neotoma. PMID- 10849303 TI - Characterization of microsatellite loci in northern flying squirrels (Glaucomys sabrinus). PMID- 10849304 TI - Isolation of microsatellites in striped bass Morone saxatilis (Teleostei) and their preliminary use in population identification. PMID- 10849305 TI - New developments for parasite immunology PMID- 10849306 TI - Apoptosis: a mechanism of immunoregulation during human Schistosomiasis mansoni. AB - People infected with schistosomes may present with a variety of clinical manifestations ranging from the relatively asymptomatic intestinal (INT) form to the hepatointestinal (HI) or hepatosplenic (HS) forms characterized by hepatomegaly and hepatosplenomegaly with severe portal hypertension, respectively. Flow cytometry analyses were used to evaluate the contribution of apoptosis in specific cell populations from schistosomiasis patients to the development of the different clinical forms of the disease. The results showed that cell death induced by combinations of specific antigen and cytokines corresponds with specific clinical presentations. It was shown that soluble egg antigen (SEA) increased the level of apoptosis only in T cells from INT patients. Stimulation with soluble lung worm antigen preparation (SLAP) did not induce significant differences in the levels of apoptosis in T cells from the patients with the different clinical forms of schistosomiasis. These results suggest for the first time that apoptosis plays an important role in the modulation of the anti-SEA response in INT patients. PMID- 10849307 TI - Protective mechanisms against the intestinal nematode Strongyloides venezuelensis in Schistosoma japonicum-infected mice. AB - Mice infected with Schistosoma japonicum were resistant to the intestinal nematode, Strongyloides venezuelensis. The numbers of adult S. venezuelensis recovered from mice were significantly decreased when infections were given from 6 weeks after S. japonicum infection. Larval recovery from the lungs showed that significant numbers of subcutaneously inoculated S. venezuelensis larvae were eliminated by 3 days in S. japonicum-infected mice (P < 0.0001), while histology revealed that this was associated with massive eosinophilic infiltration in the lungs. In addition, adult S. venezuelensis worms implanted in the duodenum of S. japonicum-infected mice could not establish in the intestine. This failure was associated with mucosal mastocytosis. Activation of eosinophils and intestinal mast cells was correlated with elevated expression of mRNA for interleukin (IL) 3, IL-4, and IL-5 in S. japonicum-infected mice. Sera from S. japonicum-infected mice recognized S. venezuelensis larva antigens as strongly as those from S. venezuelensis-infected mice, although transfer of sera from S. japonicum-infected mice to normal recipient mice did not protect them from S. venezuelensis challenge infection. It was concluded that the mechanisms for larval killing and adult worm expulsion of S. venezuelensis in S. japonicum-infected mice were identical to those seen in infections with S. venezuelensis only. PMID- 10849308 TI - Juvenile Fasciola hepatica are resistant to killing in vitro by free radicals compared with larvae of Schistosoma mansoni. AB - Free radicals have previously been shown to kill the immature stages of the trematode, Schistosoma mansoni but their effect on newly excysted juvenile (NEJ) flukes of Fasciola hepatica has not been established. Using acetaldehyde and xanthine oxidase to chemically generate reactive oxygen intermediates (ROI), up to 61% of NEJ were killed but only when exposed to high levels of ROI. At low concentrations of acetaldehyde and xanthine oxidase as sources of reactive oxygen intermediates, only 6-29% of NEJ were killed compared with 70-92% of schistosomula. Incubation with lipopolysaccharide (LPS)-stimulated rat peritoneal lavage cells (PLCs) killed only 7-15% of NEJ whereas 78-87% of schistosomula were killed under the same conditions by a mechanism dependent on the production of reactive nitrogen intermediates. Relative to immature and adult parasites, NEJ expressed 2.5-20-fold lower levels of superoxide dismutase and glutathione S transferase but no catalase activity was detected. Incubation of NEJ with inhibitors of peroxidases and glutathione metabolism increased the mean killing of NEJ by LPS-stimulated rat PLCs to 40-75%. These results demonstrate that, in comparison to schistosomula of S. mansoni, NEJ of F. hepatica are relatively resistant to killing by free radicals and this resistance could, in part, be due to the activity of oxidant scavenger enzymes of NEJ. PMID- 10849309 TI - Parasite-secreted products regulate the host response to larval Taenia crassiceps. AB - Parasite-induced immunosuppression is believed to play a significant role in the pathology of cysticercosis, a disease caused by the larval stage of cestode parasites. The biochemical basis for immunoregulation by Taenia crassiceps in experimental cysicercosis is unknown. In order to determine whether or not excretory/secretory (E/S) products from the parasite have the ability to regulate host immune function, the activity of these products was examined. Excretory/secretory products from larvae early in the infection were found to suppress T cell proliferative responses in vitro as well as the production of IFN gamma and IL-4. In contrast, E/S products secreted from larvae harvested late in infection were not suppressive. Electrophoretic analysis of E/S products revealed both qualitative and quantitative differences in the pattern of proteins produced by larvae taken from an early infection versus those taken from a chronic infection. The viability of parasites taken from an early infection was greatly reduced compared to those taken from chronically infected mice, suggesting a change in the nature of the host immune response to the parasite during the course of the infection. The proliferative activity and cytokine profiles of host immune cells were examined. Both mesenteric lymph node cells and peritoneal exudate cells were found to produce high levels of both IFN-gamma and IL-4, consistent with the high levels of these cytokines in sera of chronically infected animals. Chronic infection with Taenia crassiceps therefore is characterized by high levels of production of both Th1 and Th2 cytokines by host cells. PMID- 10849310 TI - Apoptosis modulation in mononuclear cells recovered from individuals exposed to Plasmodium falciparum infection. AB - In endemic areas, asymptomatic infection by the malaria parasite Plasmodium falciparum was found associated with elevated percentages of human host's mononuclear cell spontaneous in-vitro apoptosis. In Dielmo, a village where malaria is holoendemic, apoptosis was age-and parasite-dependent. In-vitro exposure of peripheral blood mononuclear cells (PBMC) to the parasite extract induced a marked increase in the mononuclear cell membrane expression of functional CD95 antigen: a 3-h exposure of the mononuclear cells to anti-CD95 antibodies led to a detectable increase in the mean percentage of apoptotic nuclei found in the cultures carried out in the presence of P. falciparum extracts compared to control cultures. IL-2, IL-4, IL-6 and IL-10 promoted the viability of PBMC in cultures while IL-1alpha or IFN-gamma had no obvious impact and TNFalpha gave conflicting results. IL-2 was the most efficient cytokine at rescuing PBMC from cell death and this effect was associated with a strong increase in T cell activation. In contrast, IL-4 and IL-10 had no such effect on T cell activation, hence they acted as survival factors and not through their mitogenic activity. Taken together, these different observations suggested that the levels of in-vitro apoptosis observed were not only associated with parasite infection, but also potentially modulated by the human host through different pathways. PMID- 10849311 TI - New prognostic histological parameter of invasive ductal carcinoma of the breast: clinicopathological significance of fibrotic focus. AB - Immunohistochemistry, DNA ploidy analysis and molecular genetics have made it possible to predict the outcome of breast cancer more precisely than routine histological examination alone. However, in routine practice, it is difficult to incorporate these methodologies in all cases. If certain histological parameters can accurately predict the outcome of patients with breast cancer, they would be more practical for routine use. We showed that the presence of fibrotic focus (FF) in invasive ductal carcinoma (IDC) is closely associated with c-erbB-2 or p53 protein expression, high proliferative activity, and high angiogenesis of the tumors. Furthermore, multivariate analyses with well-known prognostic parameters for IDC demonstrated that the presence of FF is the most useful independent parameter to predict IDC patient outcome. In addition, our data suggested that the interaction between tumor cells and stromal fibroblasts may play an important role in the formation of FF in IDC based on growth factor and growth factor receptor protein expression in the tumor cells and fibroblasts forming FF. Based on the results of our clinicopathological studies, we propose a new prognostic classification scheme for the prediction of IDC patient outcome, which consists of FF, nuclear atypia, and fat invasion. This classification has superior predicting power to existing prognostic classifications. PMID- 10849312 TI - Apoptosis, anoikis and their relevance to the pathobiology of colon cancer. AB - The maintenance of a constant number of cells in an adult organism is a tightly regulated process. This is particularly important in organs where cells are in a constant rate of renewal during the entire lifespan. In these organs, cell number homeostasis is the direct consequence of a balance between cell proliferation and apoptosis. The colonic epithelium is an example of such a site and the high prevalence of colon cancer makes the understanding of cell number homeostasis more important to define. Normal colonic epithelium is organized in crypts where cell proliferation, migration, differentiation and apoptosis are topographically organized in a linear fashion along the crypt axis. Normal colonic crypts are composed of stem cells at the base, a proliferation and a differentiation zone in the lower third of the crypt, a migration zone in the upper two-thirds, and the surface epithelium where senescent cells are eliminated by apoptosis. Globally, apoptosis can be defined as a normal process of cell suicide, critical for development and tissue homeostasis. Colonic epithelial cells migrate from the base of the crypt to the surface epithelium in 6-7 days. The normal architecture of the crypt is maintained by a balance between cell proliferation at the base and apoptosis at the top of the crypt and surface epithelium. PMID- 10849313 TI - A human B-lineage dendritic cell line, HBM-noda and its potential role in human T cell leukemia/lymphoma virus type I infection. AB - An Epstein-Barr virus-transformed B-cell line, HBM-Noda (Noda), that has a dendritic morphology as well as several characteristic features of dendritic cells (DC) has been established. We therefore refer to Noda as B-lineage DC. Although human T-cell leukemia/lymphoma virus type I (HTLV-I) exhibit substantial cellular tropism, the roles of DC in HTLV-I infection remain unknown. To further clarify the characteristics of Noda cells, we performed infection experiments using a concentrated HTLV-I fraction from the adult T-cell leukemia cell line, HPB-ATL-2. Noda, as well as other cell lines examined, were sensitive to HTLV-I infection as detected by proviral DNA using polymerase chain reaction, but most infected Noda cells underwent necrosis within 7 days. The most striking feature of Noda cells was the abundant expression of viral antigen (p19) on the cell surface following infection (approximately day 4), probably due to strong viral adsorption. In cocultivation experiments using Noda cells at day 1 of post infection and peripheral blood activated T cells, we detected a few (1.3%) viral antigen expressing T cells after 5 days of coculture by flow cytometry. These results suggest that B-lineage DC such as Noda cells play a role in the establishment of HTLV-I infection at an early phase. PMID- 10849314 TI - Cerebellar basket cells of Creutzfeldt-Jakob disease: immunohistochemical and ultrastructural study. AB - To elucidate possible abnormalities of cerebellar basket cells of Creutzfeldt Jakob disease (CJD), seven sporadic cases were examined neuropathologically. Recently, parvalbumin-positive, GABAergic cerebral interneurons have been demonstrated to show early, selective loss in CJD, and the phenomenon is postulated as a cause of characteristic neurological symptoms of CJD. In this study, however, we demonstrated that the basket cells, cerebellar counterparts, were resistant even in patients with severe brain atrophy, and their processes showed intense argyrophilia and immunopositivity to phosphorylated neurofilament. They can newly be listed as CJD-resistant neurons similar to those of the hippocampus and brainstem nuclei. The mechanism to escape cell loss is of great interest, and there might be unknown factors modulating susceptibility within parvalbumin-positive neuronal subgroups. Furthermore, one case showed abnormal positivity with hematoxylin, crystal violet and pyronin in the basket cells. The pyronin positivity was reduced after ribonuclease digestion, suggesting that the causative substance was composed of RNA. Ultrastructurally, the fibers contained free ribosomes and amorphous electron-dense deposits. To our knowledge, such a finding has also not been previously reported. PMID- 10849315 TI - Mucin histochemistry in primary adenocarcinoma of the urinary bladder (of urachal or vesicular origin) and metastatic adenocarcinoma originating in the colorectum. AB - In order to evaluate the mucin histochemistry of primary adenocarcinomas (PA) of the urinary bladder and metastatic adenocarcinoma (MA) originating in the colorectum, 52 PA and nine MA were examined. It was determined that the percentage of cases in which more than 25% of the tumor was stained by each of the following: (i) Alcian blue pH 2.5 periodic acid-Schiff (AB-PAS); (ii) high iron diamine-AB (HID-AB); (iii) periodic acid-sodium borohydride-potassium hydroxide-PAS (PA-SB-PH-PAS); (iv) galactose oxidase- Schiff (GOS); and (v) paradoxical concanavalin A stain (PCS). For PA, the values obtained were: 75% of cases (blue, AB-PAS), 85% (magenta, AB-PAS), 71% (black, HID-AB), 75% (blue, HID AB), 0% (PA-SB-PH-PAS), 19% (GOS), 8% (class II concanavalin A (Con A)-reactive mucin)), and 0% (class III Con A-reactive mucin). For MA, the corresponding values were 33, 22, 0, 11, 0, 0, 11, and 0%, respectively. A higher percentage of PA than MA cases showed staining in AB-PAS for acidic and neutral mucins, in HID AB for sialo- and sulfomucins, and in GOS for terminal beta-galactose and beta-N acetylgalactosamine. PA and MA were significantly different in terms of both frequency of staining with AB-PAS and frequency of staining with HID-AB. However, the overlap was such that in practice, it might be difficult, if not impossible, to use mucin histochemistry to inform a differential diagnosis. In view of the differences in AB-PAS and HID-AB positivity between PA and MA, we speculate that MA (originating in the colorectum) may have undergone structural distortion affecting the production and/or secretion of neutral mucins and acidic mucins (sialo- and sulfomucins) during metastasis or invasion. PMID- 10849316 TI - Reactive follicular hyperplasia in the lymph node lesions from systemic lupus erythematosus patients: a clinicopathological and immunohistological study of 21 cases. AB - To clarify the clinicopathological and immunohistological findings of reactive follicular hyperplasia in systemic lupus erythematosus (SLE) lymphadenopathy, we examined 21 such cases, including four males and 17 females. Three main patterns could be delineated: pattern A, histological features of Castleman's disease (n = 6); pattern B, follicular hyperplasia with pronounced arborizing vasculature in the paracortex resembling T-zone dysplasia with hyperplastic follicles (n = 6); and pattern C, follicular hyperplasia without any other specific findings (n = 9). The patients who showed patterns A and B on histology were all female with a median age of 36 years, and presented with the lymphadenopathy within 4 months, some before a definitive diagnosis could be made. The group with pattern C included four males and five females with an age ranging from 20 to 58 years (mean, 37 years). In seven of them, the lymphadenopathy was noted 6 months or more after the therapy had been initiated. In a virological study, a small to moderate number of the lymphoid cells were positive for the Epstein-Barr virus encoded small RNA in five of 10 cases examined. Human herpesvius 8 was not detected in the four cases examined by polymerase chain reaction and immunohistochemistry. The present study demonstrated that SLE lymphadenopathy showed histological variety and occasionally represented histopathological findings of multicentric Castleman's disease or findings similar to T-zone dysplasia with hyperplastic follicles in the biopsied specimens. We emphasize that careful attention to these morphological features, together with clinical and laboratory examinations, should allow a firm diagnosis of SLE to be made, providing information that is pertinent to the treatment of the disease. Moreover, disarray of the follicular dendritic cell (FDC) network, which could be easily detected by immunohistochemistry, was found in approximately 60% of our series. SLE lymphadenopathy should be listed as one of the diseases occasionally associated with disarray of the FDC network, although its clinicopathological significance remains unclear. PMID- 10849317 TI - Primary T-cell non-Hodgkin's malignant lymphoma of the appendix. AB - A case of primary T-cell lymphoma of the appendix in an 84-year-old female was reported. Appendectomy was performed as a result of the clinical diagnosis of acute appendicitis, due to the rebound tenderness of McBurney's point and thickness of the appendix wall as determined from ultra echo sonograph. Grossly, the surgical resected appendix did not have a dominant inflammatory appearance, therefore a tumor was suspected. Microscopic examination showed diffused proliferation of large and medium size lymphoma cells. Immunohistochemical examination further revealed that the lymphoma cells were positive for T-cell markers. To ensure this was a T-cell lymphoma, molecular examination was performed using paraffin-embedded tissue sections, since T-cell lymphoma of the appendix is extremely rare. Polymerase chain reaction (PCR) single-strand conformation polymorphism (SSCP) analysis demonstrated monoclonal T-cell receptor gene rearrangement. T-cell-rich B-cell lymphoma was excluded. To our knowledge, this is the first reported case of primary T-cell lymphoma of the appendix. PCR SSCP analysis in paraffin-embedded tissue section was very useful in the diagnosis of lymphoma cell monoclonality. PMID- 10849318 TI - Adenocarcinoma of the rectum with various grades of atypia in association with Crohn's disease: a case report and immunohistochemistry of p53 and Ki-67. AB - A case of adenocarcinoma of the rectum in a 41-year-old woman, in association with Crohn's disease is presented. The patient had suffered diarrhea and constipation, and Crohn's disease was suspected. Although the endoscopy did not reveal the presence of any tumors, biopsy specimens demonstrated adenocarcinoma. A Miles' operation was performed. The adenocarcinoma was composed of various grades of atypia and had invaded the non-peritonealized perirectal tissues. The infiltration of lymphocytes and plasma cells was moderate at the perimeter of the carcinoma and mild in the distant regions. Epithelioid cell granulomas were found. The p53 labeling index (LI) increased with the grade of atypia over the entire length of the carcinomatous gland. In carcinomas with high grade atypia, the p53 LI was high in both the upper and the lower halves of the gland. In carcinomas with low or moderate grade atypia however, the p53 LI was high in the lower half and low in the upper half of the gland. The Ki-67 LI over the entire gland was higher in carcinomas with high grade atypia than in carcinomas with low or moderate grade atypia. PMID- 10849319 TI - Solitary fibrous tumor of the vagina. AB - The solitary fibrous tumor (SFT) is a rare tumor that most commonly arises in the pleura. Recent evidence has indicated that this tumor originates from mesenchymal, probably fibroblastic, cells and is not restricted to the pleura. However, its occurrence in the female genital tract is extremely rare. We report a case of primary SFT that originated from the vagina in a 34-year-old female. It was a pedunculated polypoid tumor and occurred at the site of scar tissue, caused by laceration during her last labor 7 years previously. Histologically, the tumor was predominantly composed of a random proliferation of spindle cells, intimately admixed with collagen. Immunohistochemically, the cells were strongly positive for CD34, vimentin and bcl-2, but were negative for S-100 protein, neuron specific enolase, smooth muscle actin, desmin, CD68, cytokeratins and epithelial membrane antigen. To the best of our knowledge, this is the first reported case of a primary vaginal SFT in the English literature. Our report suggests to include SFT in the differential diagnosis of a spindle cell neoplasm originating from the vagina. PMID- 10849320 TI - Ovarian gonadoblastoma with mixed germ cell tumor in a woman with 46, XX karyotype and successful pregnancies. AB - An extremely rare case of unilateral gonadoblastoma with mixed germ cell tumor arising in the ovary of a 27-year-old woman with 46,XX karyotype and two successful pregnancies is reported. The mixed germ cell tumor was composed of choriocarcinoma, embryonal carcinoma, yolk sac tumor, immature teratoma and dysgerminoma. The patient has been well, without evidence of disease for over 10 years since her first surgery and adjuvant chemotherapy. PMID- 10849321 TI - Adenomyosis with a sex cord-like stromal element. AB - A case of adenomyosis with a sex cord-like stromal element is described. The element was an incidental, solitary, microscopic finding in a focus of adenomyosis. It was characterized by cord and trabecular arrangements of round to polygonal shaped cells in the endometrioid stroma. The cells were immunohistochemically positive for desmin and alpha-smooth muscle actin but negative for sex cord markers (alpha-inhibin and O13). The element appears to originate from the endometrial stromal cells through smooth muscle metaplasia. PMID- 10849322 TI - Spontaneously regressed Kaposi's sarcoma and human herpesvirus 8 infection in a human immunodeficiency virus-negative patient. AB - Kaposi's sarcoma occurring in a 78-year-old woman, with the absence of the human immunodeficiency virus infection, was correctly diagnosed by immunohistochemistry using anti-human herpesvirus 8 (HHV8) antibody (PA1-73N) for the first time. The patient suffered from chronic respiratory failure and was treated with a low dose of steroids for 2.5 years. After her medication dosage was increased for the exacerbation of the respiratory failure, multiple skin tumors in her feet and legs suddenly developed. Histopathologically, skin tumors were suspected as Kaposi's sarcoma at the first biopsy and reactive angiomatosis at the second biopsy. Polymerase chain reaction and immunohistochemistry, however, revealed the presence of HHV8 DNA fragment and positive staining in the majority of spindle cells in the skin tumors. Serological examination confirmed the positivity of anti-HHV8 antibodies. HHV8 infection and steroid-induced immunosuppression, as well as environmental factors played a role in the development of Kaposi's sarcoma in this patient, because she was born in Okinawa, which is a well-known endemic area of Kaposi's sarcoma in Japan. As her general condition improved, the skin lesions regressed without any specific treatment, and disappeared completely 8 months later, in which regression may be associated with evidence of numerous CD8 cell infiltration in the second biopsy tissues. No recurrence was observed during the following 6 month follow up. PMID- 10849323 TI - Extrauterine Mullerian adenosarcoma of the peritoneum with an extensive rhabdomyosarcomatous element and a marked myxoid change. AB - A case of extrauterine Mullerian adenosarcoma of the peritoneum in a 20-year-old woman is reported. The tumor was widely based on the abdominopelvic wall and there were no unusual features in the genital organs. The cut surface of the tumor showed a marked gelatinous appearance. The tumor was composed of an admixture of benign Mullerian-type epithelium and sarcomatous stroma. The predominant element of the sarcomatous area was rhabdomyosarcoma, which showed a close resemblance to well-differentiated embryonal rhabdomyosarcoma. In another sarcomatous area, fibroblastic cells without myoblastic properties diffusely proliferated in a marked myxoid background with some collagen bundles. Both the mitotic count and Ki-67 proliferative index of these cells were lower than those of rhabdomyoblastic cells. On follow up, the patient was disease free for 1 year postoperatively, without any subsequent treatment. The present case indicates that extrauterine adenosarcoma can also show histological heterogeneity as do uterine adenosarcomas. The remarkable myxoid change of this tumor seemed to be more largely due to a fibromyxoid element than a rhabdomyosarcomatous element, and the coexistence of the former may be related to the less aggressive behavior of this tumor. PMID- 10849324 TI - Host genes controlling the susceptibility and resistance to squamous cell carcinoma of the tongue in a rat model. AB - Development of tongue carcinoma (TC) in rats by 4-nitroquinoline 1-oxide (4NQO), a potent carcinogen, is under host genetic control. The inbred Dark-Agouti (DA) strain rats showed a much higher susceptibility to TC than the Wistar-Furth (WF) strain. The author's previous study on crosses between two strains postulated a susceptibility gene in DA and a resistance gene in WF rats. This hypothesis was confirmed by the genetic analysis of the backcrosses to either parent and F2 with a simple sequence repeat polymorphism analysis. In the crosses between the DA and WF strains of rats, two major independently segregating host loci that influenced the cancer development by application of 4NQO positively or negatively were identified and mapped. DA rats had a semidominant susceptibility gene, Stc1, closely linked with D19Mit9 on chromosome 19, which was on the segment syntenic to human chromosome 16. In contrast, WF rats had a semidominant resistance gene, Rtc1, closely linked with D1Rat320 on chromosome 1, which is syntenic to human chromosome 11. The presence of other susceptibility and resistance genes on some chromosomes of both DA and WF rats was suspected, and they will be clarified in the near future. These findings provide powerful evidence that chemically induced tongue carcinogenesis is a multigenetic event. PMID- 10849325 TI - Overexpression of keratinocyte growth factor in cancer cells and enterochromaffin cells in human colorectal cancer. AB - Keratinocyte growth factor (KGF) is a mitogenic polypeptide that is mainly synthesized by mesenchymal cells. Its actions are dependent on its binding to a specific cell-surface KGF receptor (KGFR), which is localized in epithelial cells. In the present study, the expression level of KGF and KGFR messenger RNA (mRNA), and the localization of these mRNA and proteins in tumor specimens obtained from 12 human colorectal cancer cases were estimated. Competitive reverse transcriptase-polymerase chain reaction (RT-PCR) revealed the expression of KGF and KGFR mRNA in both colorectal cancer and normal colorectal tissues. In specimens from 10 of the 12 cancer cases, the KGF mRNA level was higher in the specimens obtained from the cancerous portions than in those obtained from non cancerous tissues of the same cases. KGFR mRNA was higher in cancerous tissues in eight of 12 cases. To localize the KGF protein in normal and cancerous human colorectal tissues, immunohistochemistry was employed. In normal colorectal tissue, faint KGF immunoreactivity was present in a few fibroblasts. In contrast, strong KGF immunoreactivity was present in many of the neuroendocrine cells present in close proximity to cancer cells, and moderate immunoreactivity was recognized in the cancer cells themselves and adjacent fibroblasts. KGF-positive neuroendocrine cells also showed serotonin immunoreactivity, indicating that they were enterochromaffin cells. By in situ hybridization, both KGF and KGFR mRNA were co-overexpressed in these colorectal cancer cells, and KGF mRNA was recognized in neuroendocrine cells lying in close proximity to the cancer cells. These findings indicate the possibility that KGF acts in both a paracrine and autocrine manner to induce colorectal cancer cell growth in vivo. PMID- 10849326 TI - Mxi1 is a potential cellular target of carcinogens and frequently mutated in experimental rat tumors and tumor cell lines. AB - Mxi1, a member of the Myc family of transcription factors, negatively regulates Myc oncoprotein activity and thus may be a tumor suppressor gene. It is mutated in a few human prostate cancers. Rat Mxi1 was isolated as a selective overexpressive message in rat esophageal cancer induced by N-nitrososarcosine ethyl ester using differential display and polymerase chain reaction cloning. Reverse transcription, single-strand conformation polymorphism analysis and subsequent DNA sequencing were used to screen mutations for the rat Mxi1 coding region including the functional domains, Sin3-interacting, helix-loop-helix and leucine zipper in samples from 24 rat tumor tissues and various cell lines. Seven mutations were revealed to exist in six rat tumors (including two esophageal tumors and a breast cancer), and three rat tumor cell lines: Leydig cell tumor, osteogenic sarcoma, and pituitary tumor. No coding changes were detected in 34 samples of human sporadic gastric adenocarcinoma. A silent base substitution (GAG to GAA) at codon 131 was also identified in six rat tumors as well as in one human gastric cancer. Our results indicate that Mxi1 is often mutated in experimental rat tumors but mutations are rare in human sporadic cancers. The Mxi1 tumor suppressor gene may be a cellular target of strong carcinogens. Considering the frequency of mutations in chemical carcinogen-induced tumors, searches for Mxi1 mutation in human tumors should be directed toward patients with a specific epidemiological background. PMID- 10849327 TI - Alterations of myoepithelial cells in the rat mammary gland during pregnancy, lactation and involution, and after estradiol treatment. AB - Hormone-induced alterations of myoepithelial cells in the mammary gland have not been fully investigated. The aim of the present study was to examine whether myoepithelial cells are altered in response to hormonal conditions. The immunohistochemical findings of smooth muscle actin for myoepithelial cells were studied during pregnancy, lactation and involution, and after estradiol dipropionate (ED) treatment (50, 500, 1000 microg/kg per week for 1-4 weeks) using a total of 71 Wistar female rats. Myoepithelial cells showed a stratified appearance around ducts during pregnancy, extended cytoplasmic processes with wider distance during lactation, and vacuolated cytoplasm after weaning. ED treatment (50-1000 microg/kg per week) for 1 week increased myoepithelial cells to a variable degree, achieving a level similar to that in pregnancy, but ED treatment for 4 weeks reduced them as the dose elevated. The present study showed that the myoepithelial cells became hyperplastic or hypertrophic by low-dose ED treatment within the physiological range, while weaning pups, and excess high dose ED treatment beyond the physiological range or prolonged ED treatment induced reduction of the myoepithelial cells. Results indicate that myoepithelial cells themselves are also altered by hormonal conditions coordinating the mammary gland development. PMID- 10849328 TI - Expression of urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, and plasminogen activator inhibitor-1 and -2 in hepatocellular carcinoma. AB - It has become more and more clear in recent decades that the plasminogen activation system, which includes urokinase-type plasminogen activator (uPA), urokinase-type plasminogen activator receptor (uPAR), plasminogen activator inhibitor (PAI)-1 and PAI-2, plays a very important role in the aggressiveness of cancer. Using immunohistochemistry and enzyme-linked immunosorbent assay (ELISA), the expression of these four components of the uPA system was analyzed in 19 cases of hepatocellular carcinoma (HCC) and 18 cases of the adjacent non-cancer tissues which all had chronic active hepatitis with liver fibrosis or liver cirrhosis. Four cases of normal liver tissues, as controls for immunohistochemical stains, were obtained from the hepatectomized liver of patients with metastatic cancer in the liver. The positive rates of uPA, uPAR, PAI-1 and PAI-2 for immunohistochemical stains in cancer tissues were 78.9, 68.4, 57.9 and 31.6%, respectively. Positive signals were mainly distributed in the cytoplasm of the cancer and in stromal cells. Moreover, the strong stains were chiefly located in the invasive front of the cancer cells. No specific stain was detected in four cases of normal liver tissues. In ELISA, there were significant differences between cancer and non-cancer tissues in concentration of uPA, uPAR and PAI-1 (P < 0.0003, 0.0024 and 0.01, respectively), but there was no significant difference in that of PAI-2 (P = 0.37). These results suggest that uPA, uPAR and PAI-1 are related to invasion of HCC. PMID- 10849329 TI - Peripheral primitive neuroectodermal tumor of the small bowel mesentery: a case showing perforation at onset. AB - A case of peripheral primitive neuroectodermal tumor of the small bowel mesentery with an uncommon clinical onset is reported. A 40-year-old man was admitted to hospital because of acute severe abdominal pain. Chest X-ray revealed a free air sign beneath the diaphragm. At emergency surgery a mass measuring 11.0 x 8.0 cm with perforation was located in the jejunal mesenteric region. Histologically the resected lesion consisted of sheets of undifferentiated small round cells forming abortive Homer Wright rosettes. Some spindle-shaped cells showed perivascular pseudorosettes. Immunohistochemical study revealed that the tumor cells expressed positivity against CD99 (MIC2), neuron-specific enolase, synaptophysin and vimentin. To the authors' knowledge this is the first documentation of peripheral primitive neuroectodermal tumor of the small bowel mesentery with perforation at onset. PMID- 10849330 TI - Ileal carcinoid tumor complicating carcinoid heart disease and secondary retroperitoneal fibrosis. AB - A 70-year-old man with metastatic liver tumors showed carcinoid syndrome with clinical symptoms of facial flushing, palpitation, dyspnea, and an itching sensation. Regurgitation of the tricuspid and aortic valves was observed by echocardiography. An elevated serum level of serotonin and a high urine excretion of serotonin metabolites were confirmed. Autopsy confirmed a serotonin immunoreactive 1 cm ileal carcinoid tumor with metastasis to the liver, bone and peritoneum. The tumor cells were argyrophilic but not argentaffin, and showed erythrophagocytosis in the primary lesion. All the four heart valves and bilateral atrial endocardium showed fibromyxoid thickening, indicating the association of carcinoid heart disease. Desmoplastic reaction with deposition of sulfated acid mucopolysaccharides was also observed in the retroperitoneal space (secondary retroperitoneal fibrosis). Stenosis of the bilateral ureters and inferior mesenteric artery provoked hydronephrosis and lethal ischemic necrosis of the left-sided colon, respectively. Rarity of functioning ileal carcinoid tumor and pathogenesis of systemic fibroplasia are discussed. PMID- 10849331 TI - Clear cell sarcoma arising in the transverse colon. AB - A case of clear cell sarcoma (CCS) arising in the transverse colon is presented. The tumor consisted of sheets or small nests of epithelioid malignant cells possessing pleomorphic nuclei with one or more prominent nucleoli and ample clear or slightly eosinophilic cytoplasm. Some of the tumor cells contained various amounts of melanin pigments that were confirmed by histochemical and ultrastructural examinations. Immunohistochemical examination showed a positive immunoreactivity for HMB45 and S-100 protein. A metastatic nodule, which was found 9 months after surgery, showed similar histological findings to those of the primary one but lacked melanin pigments. Reverse transcriptase- polymerase chain reaction using total ribonucleic acid obtained from metastatic nodule demonstrated the presence of EWS-ATF-1 fusion gene. Based on these findings, the present case tumor is a CCS of the colon. PMID- 10849332 TI - Pathologic, cytologic and immunohistochemical findings of an intra-abdominal desmoplastic small round cell tumor in a 15-year-old male. AB - The intra-abdominal desmoplastic small round cell tumor is a rare neoplasm. It usually occurs in young males and diffusely involves the peritoneum and pursues an aggressive clinical course. The present patient was a 15-year-old male who experienced abdominal pain and abdominal swelling. The patient was diagnosed with an intestinal myogenic sarcoma, and surgery for tumor resection was performed in June 1999. The tumor was a 20 x 15 x 15 cm well-defined mass in the peritoneum involving the transverse colon and stomach with peritoneal disseminations and splenic metastasis. Microscopic findings were well-defined nests composed of small round cells and separated by abundant desmoplastic stroma. Cytologically, the tumor cells consisted of small, round to oval cells with a scant amount of light blue cytoplasm. Immunohistochemically, the tumor cells were positive for anti-epithelial membrane antigen, vimentin, desmin, neuron-specific enolase and WT1 protein antibodies. Similar pathologic features with other small round cell tumors may lead to differential diagnostic difficulties that require the application of ancillary diagnostic methods, such as immunohistochemistry and cytogenetic techniques. PMID- 10849333 TI - Bronchial granular cell tumor with osteopontin and osteonectin expression: a case report. AB - The case of a 52-year-old Japanese man with bronchial granular cell tumors with osteopontin and osteonectin expression is reported here because there have been few investigations of their expression in benign tumors. He was admitted because of sudden hematemesis. A bronchoscopic examination revealed a lobulated polypoid tumor located in the left and right bronchi. Histologically, most tumor cells had abundant granular eosinophilic cytoplasm and were immunoreactive for S-100, neuron-specific enolase (NSE), CD68 and vimentin. Moreover, osteopontin-positive tumor cells were randomly distributed in the tumor tissue, but few stromal cells were positive. In contrast, osteonectin was mainly expressed in the peripheral tumor cells and was also distributed in the stromal cells. Blood vessels at the tumor border in which osteonectin-positive tumor cells were distributed, proliferated moderately. These results suggest that osteopontin and osteonectin may play a role in the progression of granular cell tumors and in the interaction between the tumor and host or angiogenesis around the tumor, respectively. PMID- 10849334 TI - Ciliated foregut cyst of the gallbladder: a case report and review of the literature. AB - A case is presented of a ciliated cyst of the gallbladder in a 36-year-old Korean woman which was incidentally found on ultrasonographic study. A cystic mass measuring 1.5 x 1 x 1 cm was found in the fundus of the gallbladder. The cyst was unilocular and intramural without communication to the lumen. Microscopically, the cyst wall was lined by a single layer of pseudostratified, ciliated, columnar epithelium and goblet cells with underlying smooth muscle layers. This was considered to be the cyst arising from the embryonic foregut and showing differentiation toward respiratory structures. The term 'ciliated foregut cyst of the gallbladder' is suggested here. PMID- 10849335 TI - Solitary squamous cell papilloma of the lung in a 40-year-old woman with recurrent laryngeal papillomatosis. AB - A rare case of recurrent respiratory papillomatosis (RRP) is reported with a review of the literature. A 40-year-old Japanese woman had suffered from RRP since 1 year of age. She developed a pulmonary squamous papilloma with a thin walled cavity, which was suspected as being lung carcinoma. The trachea and bronchi around the tumor were intact, and no malignant transformation was present. Two types of human papillomavirus, 6 and 16, were detected, both in the laryngeal and pulmonary papillomas by in situ hybridization and the polymerase chain reaction method. To date, only 40 cases of juvenile laryngeal papilloma with pulmonary involvement have been reported in the English literature. PMID- 10849336 TI - A new type of mutation in the plant photoreceptor phytochrome B causes loss of photoreversibility and an extremely enhanced light sensitivity. AB - Photoreversibility, reversion of the inductive effect of a brief red light pulse by a subsequent far-red light pulse, is a property of photoresponses regulated by the plant photoreceptor phytochrome B (phyB). We screened for mutants with impaired photoreversibility to gain better insight into the phyB-specific signalling cascade. The phenotype of the mutant described is caused by a single amino acid exchange in a phyB subdomain that is highly conserved in all phytochromes but whose functional significance was unknown. The mutated phyB showed a slower dark reversion but no major alterations in its spectral properties. In addition to its loss of photoreversibility, the mutant also exhibited a hypersensitivity towards continuous red-light irradiation and an altered phenotype of adult plants under short-day conditions. PMID- 10849337 TI - Arabidopsis thaliana poly (A) binding protein 2 (PAB2) functions in yeast translational and mRNA decay processes. AB - The single yeast gene (PAB1) encoding poly (A) binding protein (PABP) has several roles in post-transcriptional processes, including translation initiation and mRNA decay. PABP is encoded by a large gene family in plants. Within Arabidopsis thaliana, the several characterized PABP genes exhibit an extreme degree of sequence divergence and are differentially expressed. Arabidopsis PAB2 is expressed in distinct tissues or during defined developmental windows in most plant organs. In this study we demonstrate that PAB2 restores viability to a yeast pab1 mutant strain. Yeast strains containing wild-type, null (PAB2s) and temperature sensitive (PAB2ts) alleles of PAB2 were used to explore the molecular functions of the plant protein. PAB2 can participate in poly (A) tail shortening, thus demonstrating that it interacts with the yeast poly(A) nuclease complex. PAB2 is required for translation, helping to maintain intact polysome structures. Consistent with its role in translation initiation, poly (A) was found to enhance PAB2 binding to Arabidopsis eIF-iso4G in vitro. In addition, PAB2 can partially restore the linkage between deadenylation, decapping and mRNA decay in yeast. Taken together, our results suggest that Arabidopsis PAB2 participates in many of the same complex post-transcriptional processes identified for yeast PAB1, and is functionally distinct from other characterized Arabidopsis PABPs. PMID- 10849338 TI - Conserved expression of Arabidopsis thaliana poly (A) binding protein 2 (PAB2) in distinct vegetative and reproductive tissues. AB - The poly(A) tails of eukaryotic mRNAs are complexed with poly(A) binding protein (PABP). The poly(A)-PABP complex is central to the efficient translation initiation and control of poly (A) tail length and is required in some pathways of mRNA decay. A large gene family encodes PABPs in Arabidopsis thaliana. In striking contrast to the floral and root specific expression of three previously reported Arabidopsis PABPs, we demonstrate that RNA and protein for one highly diverse member of this family, PAB2, are expressed in roots, stems, leaves, flowers, pollen and siliques of Arabidopsis. However, cell-type specific analysis of a PAB2 reporter gene fusion revealed that PAB2 is spatially and temporally regulated in each organ. For example, strong expression was detected only in the stele and meristem region of roots and a dramatic decrease in expression was observed upon fertilization of ovules. Furthermore, the PAB2-reporter construct gave a nearly identical expression pattern in transgenic tobacco, demonstrating that PAB2 expression is under strong selective constraint. The PAB2-reporter was also strongly expressed in the transmittal tissues of both Arabidopsis and tobacco, raising the possibility of its involvement in the pollination-dependent poly(A) tail shortening of transmittal tissue specific mRNAs previously reported in tobacco (Wang et al. 1996, Plant J. 9, 715-727). In view of its potential role in poly(A) tail shortening, we demonstrated the strong and distinct presence of PAB2 protein in transmittal tissues of Arabidopsis. The evolutionary and functional implications of the expression pattern of PAB2 and its possible functional roles in post-transcriptional regulation in transmittal tissues are discussed. PMID- 10849339 TI - The evolution of C4 plants: acquisition of cis-regulatory sequences in the promoter of C4-type pyruvate, orthophosphate dikinase gene. AB - In a previous study, we identified the C4-like pyruvate, orthophosphate dikinase gene (Pdk) in the C3 plant rice, with a similar structure to the C4-type Pdk in the C4 plant maize. In order to elucidate the differences between C4-type and C4 like Pdk genes in C4 and C3 plants, we have produced chimeric constructs with the beta-glucuronidase (GUS) reporter gene under the control of the Pdk promoters. In transgenic rice, both rice and maize promoters directed GUS expression in photosynthetic organs in a light-dependent manner. However, the maize promoter exhibited a much higher transcriptional activity than the rice promoter did. These results indicate that the rice C4-like Pdk gene resembles the maize C4-type Pdk gene in terms of regulation of expression. We also tested the activity of the rice promoter in transgenic maize. GUS activity was seen in both photosynthetic and non-photosynthetic organs. Thus, the rice promoter does not confer a strict organ-specific gene expression, as the maize promoter does. Moreover, the rice promoter directed GUS expression not only in mesophyll cells but also in bundle sheath cells, whereas the maize promoter directed expression only in mesophyll cells. Taken together, the results obtained from both transgenic maize and rice demonstrate that the rice and maize promoters differ not only quantitatively, but also qualitatively, in terms of their cell- and organ-specificity. Experiments with swapped promoters using the rice and maize promoters further demonstrated that a limited sequence region from -330 to -76 of the maize promoter confers light-regulated, high-level expression to the rice promoter in maize mesophyll protoplasts. We conclude the gain of cis-acting elements conferring high-level expression and mesophyll cell specificity was necessary for establishment of a C4 type Pdk gene during the course of evolution from C3 to C4 plants. PMID- 10849340 TI - Modified lignin in tobacco and poplar plants over-expressing the Arabidopsis gene encoding ferulate 5-hydroxylase. AB - Ferulate 5-hydroxylase (F5H) is a cytochrome P450-dependent monooxygenase that catalyses the hydroxylation of ferulic acid, coniferaldehyde and coniferyl alcohol in the pathways leading to sinapic acid and syringyl lignin biosynthesis. Earlier studies in Arabidopsis have demonstrated that F5H over-expression increases lignin syringyl monomer content and abolishes the tissue-specificity of its deposition. To determine whether this enzyme has a similar regulatory role in plants that undergo secondary growth, we over-expressed the F5H gene in tobacco and poplar. In tobacco, over-expression of F5H under the control of the cauliflower mosaic virus 35S promoter increased lignin syringyl monomer content in petioles, but had no detectable effect on lignification in stems. By contrast, when the cinnamate 4-hydroxylase (C4H) promoter was used to drive F5H expression, there was a significant increase in stem lignin syringyl monomer content. Yields of thioglycolic acid and Klason lignin in C4H-F5H lines were lower than in the wild-type, suggesting that F5H over-expression leads to a reduced deposition or an altered extractability of lignin in the transgenic plants. Histochemical analysis suggested that the novel lignin in C4H-F5H transgenic lines was altered in its content of hydroxycinnamyl aldehydes. Transgenic poplar trees carrying the C4H-F5H transgene also displayed enhanced lignin syringyl monomer content. Taken together, these data show that hydroxylation of guaiacyl-substituted lignin precursors controls lignin monomer composition in woody plants, and that F5H over expression is a viable metabolic engineering strategy for modifying lignin biosynthesis in forest species. PMID- 10849341 TI - Diverse effects of overexpression of LEAFY and PTLF, a poplar (Populus) homolog of LEAFY/FLORICAULA, in transgenic poplar and Arabidopsis. AB - PTLF, the Populus trichocarpa homolog of LEAFY (LFY) and FLORICAULA, was cloned to assess its function in a dioecious tree species. In situ hybridization studies showed that the gene was expressed most strongly in developing inflorescences. Expression was also seen in leaf primordia and very young leaves, most notably in apical vegetative buds near inflorescences, but also in seedlings. Although ectopic expression of the PTLF cDNA in Arabidopsis accelerated flowering, only one of the many tested transgenic lines of Populus flowered precociously. The majority of trees within a population of 3-year-old transgenic hybrid Populus lines with PTLF constitutively expressed showed few differences when compared to controls. However, phenotypic effects on growth rate and crown development, but not flowering, were seen in some trees with strong PTLF expression and became manifest only as the trees aged. Competence to respond to overexpression of LFY varied widely among Populus genotypes, giving consistent early flowering in only a single male P. tremula x P. tremuloides hybrid and causing gender change in another hybrid genotype. PTLF activity appears to be subject to regulation that does not affect heterologously expressed LFY, and is dependent upon tree maturation. Both genes provide tools for probing the mechanisms of delayed competence to flower in woody plants. PMID- 10849342 TI - Changes in GA 20-oxidase gene expression strongly affect stem length, tuber induction and tuber yield of potato plants. AB - Gene StGA20ox1 encoding potato GA 20-oxidase is expressed to relatively high levels in leaves and regulated by daylength. To investigate whether this gene is involved in photoperiodic regulation of tuber formation, we have obtained transgenic potato plants expressing sense and antisense copies of the StGA20ox1 cDNA. Over-expression of this cDNA resulted in taller plants that required a longer duration of a short day photoperiod (SD) to tuberize. Tubers from these plants had a decreased time of dormancy and developed sprouts with elongated internodes. Plants expressing antisense copies of the StGA20ox1 cDNA had shorter stems, a decreased length of the internodes and tuberized earlier than control plants, showing increased tuber yields. Antisense inhibition of this gene had no visible effect on the time of dormancy of the tubers, although at the end of dormancy these formed sprouts with shortened internodes. Decreased levels of endogenous GA20 and GA1 were detected in the apex and first leaves of the antisense lines. These results demonstrate the involvement of the GA 20-oxidase activity encoded by StGA20ox1 in the control of stem elongation and in tuber induction but not in tuber dormancy, indicating that the latter may be regulated by another member of the gene family. PMID- 10849343 TI - Disruption of an Arabidopsis cytoplasmic ribosomal protein S13-homologous gene by transposon-mediated mutagenesis causes aberrant growth and development. AB - We identified a Dissociation (Ds) transposon-inserted Arabidopsis mutant of a gene (AtRPS13A) homologous to cytoplasmic ribosomal protein (RP) S13. We named our mutant pointed first leaf (pfl) 2 because of its similar phenotype to the pfl1 mutant of the RPS18 gene. This mutant caused multiple phenotypic changes, including aberrant leaf and trichome morphology, retarded root growth, and late flowering. Microscopic analysis showed that the first leaf blade of pfl2 contained a significantly reduced number of palisade cells, which suggests that the mutant phenotype was caused by reduced cell division. However, no phenotypic changes were observed during reproductive growth. In Arabidopsis, the RPS13 protein was encoded by a small expressed gene family including AtRPS13A. A pfl1 pfl2 double mutant showed no additive effect. These results suggest that RPS13 functions in quantitative and pleiotropic ways during growth and development, and that mutations at different kinds of RP gene loci are accumulatable without serious growth defects because they belong to small gene families. PMID- 10849344 TI - Technical advance: a high throughput system for transposon tagging and promoter trapping in tomato. AB - We describe new tools for functional analysis of the tomato genome based on insertional mutagenesis with the maize Ac/Ds transposable elements in the background of the miniature cultivar Micro-Tom. 2932 F3 families, in which Ds elements transposed and were stabilized, were screened for phenotypic mutations. Out of 10 families that had a clear mutant phenotype, only one mutant was Ds tagged. In addition, we developed promoter trapping using the firefly luciferase reporter gene and enhancer trapping, using beta-glucuronidase (GUS). We show that luciferase can be used as a non-invasive reporter to identify, isolate and regenerate somatic sectors, to study the time course of mutant expression, and to identify inducible genes. Out of 108 families screened for luciferase activity 55% showed expression in the flower, 11% in the fruit and 4% in seedlings, suggesting a high rate of Ds insertion into genes. Preferential insertion into genes was supported by the analysis of Ds flanking sequences: 28 out of 50 sequenced Ds insertion sites were similar to known genes or to ESTs. In summary, the 2932 lines described here contain 2-3 Ds inserts per line, representing a collection of approximately 7500 Ds insertions. This collection has potential for use in high-throughput functional analysis of genes and promoter isolation in tomato. PMID- 10849345 TI - Two bean cell wall proteins more abundant during water deficit are high in proline and interact with a plasma membrane protein. AB - Two antigenically related glycoproteins, called p33 and p36, accumulate in the soluble fraction of the cell wall in response to water deficit in Phaseolus vulgaris. In this report, we show that p33 and p36 are able to adhere to leaf protoplasts, and that they bind to plasma membrane (PM) vesicles in a divalent cation-dependent manner. Data from the partial amino acid sequence of the p33 and p36 proteins indicate that they contain repeats of the decapeptide POVYKPOVEK; therefore, they are related to proline-rich proteins. Binding assays demonstrate that both proteins specifically bind to an 80 kDa PM protein. This binding is competed with a peptide that contains the RGD motif, as well as with fibronectin, which also includes this sequence, suggesting that the 80 kDa PM protein has an integrin-like function whose natural ligands are p33 and p36. This is the first case where a PM ligand for a higher plant cell wall protein has been identified. PMID- 10849346 TI - Functional analysis of tobacco LIM protein Ntlim1 involved in lignin biosynthesis. AB - The AC-rich motif, Pal-box, is an important cis-acting element for gene expression involved in phenylpropanoid biosynthesis. A cDNA clone (Ntlim1) encoding a Pal-box binding protein was isolated by Southwestern screening. The deduced amino acid sequence is highly similar to the members of the LIM protein family that contain a zinc finger motif. Moreover, Ntlim1 had a specific DNA binding ability and transiently activated the transcription of a beta glucuronidase reporter gene driven by the Pal-box sequence in tobacco protoplasts. The transgenic tobacco plants with antisense Ntlim1 showed low levels of transcripts from some key phenylpropanoid pathway genes such as phenylalanine ammonia-lyase, hydroxycinnamate CoA ligase and cinnamyl alcohol dehydrogenase. Furthermore, a 27% reduction of lignin content was observed in the transgenic tobacco with antisense Ntlim1. PMID- 10849347 TI - Mutations in the Arabidopsis VAR2 locus cause leaf variegation due to the loss of a chloroplast FtsH protease. AB - Variegated plants have green- and white-sectored leaves. Cells in the green sectors contain morphologically normal chloroplasts, whereas cells in the white sectors contain non-pigmented plastids that lack organized lamellar structures. Many variegations are caused by mutations in nuclear genes that affect plastid function, yet in only a few cases have the responsible genes been cloned. We show that mutations in the nuclear VAR2 locus of Arabidopsis cause variegation due to loss of a chloroplast thylakoid membrane protein that bears similarity to the FtsH family of AAA proteins (ATPases associated with diverse cellular activities). Escherichia coli FtsH is a chaperone metalloprotease that functions in a number of diverse membrane-associated events. Although FtsH homologs have been identified in multicellular organisms, their functions and activities are largely unknown; we provide genetic in vivo evidence that VAR2 functions in thylakoid membrane biogenesis. We have isolated four var2 alleles and they have allowed us to define domains of the protein that are required for activity. These include two putative ATP-binding sites. VAR2 protein amounts generally correlate with the severity of the var2 mutant phenotype. One allele lacks detectable VAR2 protein, suggesting that the mechanism of var2 variegation involves the action of a redundant activity in the green sectors. We conclude that redundant activities may be a general mechanism to explain nuclear gene-induced plant variegations. PMID- 10849348 TI - Cell-to-cell movement of TMV RNA is temperature-dependent and corresponds to the association of movement protein with microtubules. AB - The movement protein (MP) of tobacco mosaic virus (TMV) is essential for spread of the viral RNA genome from cell to cell. During infection, the MP associates with microtubules, and it has been proposed that the cytoskeleton transports the viral ribonucleoprotein complex from ER sites of synthesis to plasmodesmata through which infection spreads into adjacent cells. However, microtubule association of MP was observed in cells undergoing late infection rather than in cells undergoing early infection at the leading edge of expanding infection sites where virus RNA cell-to-cell spread occurs. Therefore, alternative roles for microtubules in virus infection have been proposed, including a role in MP degradation. To further investigate the role of microtubules in virus pathogenesis, we tested the efficiency of cell-to-cell spread of infection and microtubule association of the MP in response to changes in temperature. We show that the subcellular distribution of MP is temperature-dependent and that a higher efficiency of intercellular transport of virus RNA at elevated temperatures corresponds to an increased association of MP with microtubules early in infection. PMID- 10849349 TI - Probing expansin action using cellulose/hemicellulose composites. AB - Cellulose-based composite materials containing xyloglucans or mannan-based polysaccharides have been shown to possess organisational features with many characteristics similar to primary plant cell walls. We have tested the effects of a typical alpha-expansin (CsExp1) on these composites using two different mechanical assays. We show that CsExp1 induces very rapid extension in composites containing tamarind xyloglucan under constant load. In contrast, expansin treatment had no effect in constant load extension assays using cellulose-only materials or in those carried out on composites containing glucomannan or galactomannan. We show that the effect of expansins is much smaller on composites made with short chain length xyloglucans than on those containing longer chains. In uniaxial extension tests we found that expansin could double the total extension (before failure) in xyloglucan composites and that the effects were again lower in composites containing shorter xyloglucans. We found no effect of expansin on uniaxial extensions with glucomannan or galactomannan. However, a significant effect of expansin on the uniaxial extension behaviour of cellulose only material was observed. These experiments suggest that the target of CsExp1 in cell walls is probably the cellulose xyloglucan matrix, but that other (1-4) beta-glucan to (1-4) beta-glucan hydrogen bonded contacts can also serve as substrates. PMID- 10849350 TI - Expression of a bacterial serine acetyltransferase in transgenic potato plants leads to increased levels of cysteine and glutathione. AB - The coding sequence of the wild-type, cys-sensitive, cysE gene from Escherichia coli, which encodes an enzyme of the cysteine biosynthetic pathway, namely serine acetyltransferase (SAT, EC 2.3.1. 30), was introduced into the genome of potato plants under the control of the cauliflower mosaic virus 35S promoter. In order to target the protein into the chloroplast, cysE was translationally fused to the 5'-signal sequence of rbcS from Arabidopsis thaliana. Transgenic plants showed a high accumulation of the cysE mRNA. The chloroplastic localisation of the E. coli SAT protein was demonstrated by determination of enzymatic activities in enriched organelle fractions. Crude leaf extracts of these plants exhibited up to 20-fold higher SAT activity than those prepared from wild-type plants. The transgenic potato plants expressing the E. coli gene showed not only increased levels of enzyme activity but also exhibited elevated levels of cysteine and glutathione in leaves. Both were up to twofold higher than in control plants. However, the thiol content in tubers of transgenic lines was unaffected. The alterations observed in leaf tissue had no effect on the expression of O-acetylserine(thiol)-lyase, the enzyme which converts O-acetylserine, the product of SAT, to cysteine. Only a minor effect on its enzymatic activity was observed. In conclusion, the results presented here demonstrate the importance of SAT in plant cysteine biosynthesis and show that production of cysteine and related sulfur-containing compounds can be enhanced by metabolic engineering. PMID- 10849351 TI - A resistance gene product of the nucleotide binding site -- leucine rich repeats class can form a complex with bacterial avirulence proteins in vivo. AB - Resistance (R) genes in plants mediate gene-for-gene disease resistance. The ligand-receptor model, which explains the gene-for-gene specificity, predicts a physical interaction between an elicitor, which is directly or indirectly encoded by an avirulence (avr) gene in the pathogen, and the corresponding R gene product. The nucleotide binding site (NBS) - leucine rich repeats (LRR) class of R genes is the largest known class of R genes. Here we report that an NBS-LRR R protein and its cognate Avr protein form a complex together in the plant cell. The Arabidopsis thaliana R genes RPS2 and RPM1 confer gene-for-gene disease resistance to strains of the phytopathogenic bacterium Pseudomonas syringae carrying the avr genes avrRpt2 and avrB, respectively. Using transient expression of these genes in Arabidopsis leaf mesophyll protoplasts, we first demonstrated that the protoplast system is appropriate for the investigation of the gene-for gene recognition mechanism. Formation of an in vivo complex containing the RPS2 and AvrRpt2 proteins was demonstrated by co-immunoprecipitation of the proteins following expression of the genes in protoplasts. This complex contained at least one additional plant protein of approximately 75 kDa. Unexpectedly, RPS2 also formed a complex with AvrB. We speculate that complex formation between AvrRpt2 and RPS2 is productive and leads to the elicitation of the resistance response, whilst complex formation between AvrB and RPS2 is unproductive and possibly competes with complex formation between AvrRpt2 and RPS2. PMID- 10849352 TI - Isolation and characterization of HsfA3, a new heat stress transcription factor of Lycopersicon peruvianum. AB - Stress-induced transcription of heat shock proteins (Hsps) in eukaryotes is mediated by a conserved class of transcription factors called heat stress transcription factors (Hsfs). Here we report the isolation and functional characterization of HsfA3, a new member of the Hsf family. HsfA3 was cloned from a tomato heat stress cDNA library by yeast two-hybrid screening, using HsfA1 as a bait. HsfA3 is a single-copy gene with all the conserved sequence elements characteristic of a heat stress transcription factor. The constitutively expressed HsfA3 is mainly found in the cytoplasm under control conditions and in the nucleus under heat stress conditions. Functionally, HsfA3 behaves similarly to the already known members of tomato Hsf family. It is able to substitute yeast Hsf for viability functions and is a strong activator of Hsf-dependent reporter constructs both in tobacco protoplasts and yeast. Finally, similar to the AHA motifs in HsfA1 and HsfA2, the activator function depends on four short peptide motifs with a central tryptophan residue found in the C-terminal domain of HsfA3. PMID- 10849353 TI - Ozone treatment rapidly activates MAP kinase signalling in plants. AB - Brief exposure to ozone, a potent cross-inducer of plant stress responses, leads within minutes to activation of an ERK-type MAP kinase (approximately 46 kDa) in tobacco. This activation process is calcium-dependent and can be blocked both by free radical quenchers and by a specific inhibitor of MEK-1 (MAPKK). Hydrogen peroxide and superoxide anion radicals can substitute for ozone as the activation stimulus, which does not appear to require salicylate as an intermediary. The properties of the ozone-induced MAPK suggest that it may be SIPK (salicylate induced protein kinase), a tobacco MAPK that is activated by a variety of stress treatments. The ability of ozone to activate SIPK indicates that this protein kinase acts as a very early transducer of redox stress signals in plant cells. PMID- 10849354 TI - Modulation by phytochrome of the blue light-induced extracellular acidification by leaf epidermal cells of pea (Pisum sativum l.): a kinetic analysis. AB - Blue light induces extracellular acidification, a prerequisite of cell expansion, in epidermis cells of young pea leaves, by stimulation of the proton pumping ATPase activity in the plasma membrane. A transient acidification, reaching a maximum 2.5-5 min after the start of the pulse, could be induced by pulses as short as 30 msec. A pulse of more than 3000 micromol m-2 saturated this response. Responsiveness to a second light pulse was recovered with a time constant of about 7 min. The fluence rate-dependent lag time and sigmoidal increase of the acidification suggested the involvement of several reactions between light perception and activation of the ATPase. In wild-type pea plants, the fluence response relation for short light pulses was biphasic, with a component that saturates at low fluence and one that saturates at high fluence. The phytochrome deficient mutant pcd2 showed a selective loss of the high-fluence component, suggesting that the high-fluence component is phytochrome-dependent and the low fluence component is phytochrome-independent. Treatment with the calmodulin inhibitor W7 also led to the elimination of the phytochrome-dependent high fluence component. Simple models adapted from the one used to simulate blue light induced guard cell opening failed to explain one or more elements of the experimental data. The hypothesis that phytochrome and a blue light receptor interact in a short-term photoresponse is endorsed by model calculations based upon a three-step signal transduction cascade, of which one component can be modulated by phytochrome. PMID- 10849355 TI - Phytochromes confer the photoperiodic control of flowering in rice (a short-day plant). AB - The photoperiodic sensitivity 5 (se5) mutant of rice, a short-day plant, has a very early flowering phenotype and is completely deficient in photoperiodic response. We have cloned the SE5 gene by candidate cloning and demonstrated that it encodes a putative heme oxygenase. Lack of responses of coleoptile elongation by light pulses and photoreversible phytochromes in crude extracts of se5 indicate that SE5 may function in phytochrome chromophore biosynthesis. Ectopic expression of SE5 cDNA by the CaMV 35S promoter restored the photoperiodic response in the se5 mutant. Our results indicate that phytochromes confer the photoperiodic control of flowering in rice. Comparison of se5 with hy1, a counterpart mutant of Arabidopsis, suggests distinct roles of phytochromes in the photoperiodic control of flowering in these two species. PMID- 10849356 TI - Expression patterns of genes encoding HD-ZipIV homeo domain proteins define specific domains in maize embryos and meristems. AB - A family of homeo box genes with cell layer-specific expression patterns defining subdomains of the embryo and certain meristems has been isolated from maize. These genes encode proteins from the class of plant specific homeo domain-leucine zipper (HD-Zip) transcription factors containing the previously described ZmOCL1 protein, and have been designated ZmOCL2, ZmOCL3, ZmOCL4 and ZmOCL5. ZmOCL3, ZmOCL4 and ZmOCL5, like ZmOCL1, showed essentially L1 or epidermis-specific expression. However, each gene was expressed in a distinct region of the embryonic protoderm during early development, with ZmOCL3 showing suspensor specific expression, ZmOCL4 transcripts being localized to the adaxial face of the embryo proper and ZmOCL5 showing a more abaxial expression pattern. All three genes were also expressed in vegetative, inflorescence and floral apices, although ZmOCL3 transcripts were excluded from meristems and very young organ primordia. In contrast, ZmOCL2 expression was entirely meristem-specific and was excluded from the L1 layer, appearing instead to be largely restricted to a cell layer directly beneath the L1, especially in floral meristems. This expression pattern is unprecedented and may indicate that cell-layer organization in maize meristems is more complex than that suggested by the classical L1/L2 (outer cell layer/inner cell mass) model. These differing expression patterns indicate that the members of the HD-ZipIV family of maize may not only play roles in defining different regions of the epidermis during embryonic development, but could also be responsible for maintaining cell-layer identity in meristematic regions. PMID- 10849357 TI - Expression of Brassica juncea 3-hydroxy-3-methylglutaryl CoA synthase is developmentally regulated and stress-responsive. AB - 3-Hydroxy-3-methylglutaryl-coenzyme A synthase (HMGS) is an enzyme in mevalonate biosynthesis. In plants, investigations have focused on HMG CoA reductase (HMGR) and less is known of the preceding enzyme, HMGS. To understand the regulation of HMGS, we have isolated a Brassica juncea cDNA encoding HMGS, BjHMGS1, for use as a hybridization probe in Northern blot analyses. BjHMGS is expressed in all plant organs and shows developmental regulation in flower, seed and seedling, with highest expression in early development. In seedlings, expression is highest in young hypocotyls and is induced during the greening of etiolated cotyledons. BjHMGS is down-regulated by abscisic acid, osmotic stress and dehydration, the effects of which arrested seedling growth. Thus BjHMGS expression shows correlation with rapid cell division and growth, like HMGR. This is not unexpected, as mevalonate is the precursor to many essential isoprenoid compounds, including sterols for membrane biogenesis. Wounding, methyl jasmonate or salicylic acid induce BjHMGS expression, suggesting that, like HMGR, HMGS is involved in defence. As in animals, coordinated regulation of HMGS with HMGR occurred in B. juncea upon germination and in response to salicylic acid. HMGS assays confirmed that Escherichia coli-expressed recombinant BjHMGS1 shows HMGS activity that is inhibited by F244, a specific inhibitor of HMGS. Southern blot analysis revealed gene families encoding HMGS in Brassica species and a summation of homologous genes in the fusion amphidiploid genome of B. juncea, a bi-parental species derived from diploids B. nigra and B. campestris. PMID- 10849358 TI - Fusion genetic analysis of gibberellin signaling mutants. AB - A fusion genetic strategy was used to identify gibberellin (GA) signaling mutants in transgenic Arabidopsis expressing the beta-glucuronidase (GUS) and firefly luciferase (LUC) reporter genes under control of the GA-responsive GASA1 promoter. Initial analyses determined the spatial and temporal patterns of reporter expression, and showed that reporter induction by GA was antagonized by ABA. gamma-Irradiated M2 progeny with altered reporter activities were identified by LUC bioimaging followed by GUS assays and northern hybridization of the endogenous GASA1 mRNA. Genetic analysis showed that three mutants, which overexpressed both reporters and endogenous GASA1, were caused by recessive (goe1 and goe2, for GASA over-expressed) and semi-dominant (goe3) mutations at different loci. These mutants are altered in their sensitivity to GA and the GA biosynthetic inhibitor paclobutrazol, and in the expression of several GA signaling related genes. PMID- 10849359 TI - The TATA motif, the CAA motif and the poly(T) transcription termination motif are all important for transcription re-initiation on plant tRNA genes. AB - The effect of alteration of 5' and 3' flanking sequences on the transcription of plant tRNA genes was analysed using an RNA polymerase III-dependent in vitro transcription system derived from nuclei of cultured tobacco cells. A TATA-like sequence and the CAA motif frequently observed upstream of plant tRNA genes, and the poly(T) stretch usually present downstream, were shown to be necessary for efficient re-initiation of transcription. The CAA motif was shown to be a transcription initiation site. Introduction of the CAA and TATA-like motifs into a gene naturally lacking them greatly enhanced transcription by promoting efficient re-initiation. PMID- 10849360 TI - Transformation of Arabidopsis with the codA gene for choline oxidase enhances freezing tolerance of plants. AB - Arabidopsis thaliana was transformed with the codA gene from Arthrobacter globiformis, which encodes choline oxidase, the enzyme that synthesizes glycinebetaine from choline. The transformation enabled the plants to accumulate glycinebetaine in chloroplasts, and significantly enhanced the freezing tolerance of plants. Furthermore, the photosynthetic machinery of transformed plants was more tolerant to freezing stress than that of wild-type plants. Exogenous application of glycinebetaine also increased the freezing tolerance of wild-type plants, suggesting that the presence of glycinebetaine in transformed plants had enhanced their ability to tolerate freezing stress. Northern blotting analysis revealed that the enhancement of freezing tolerance was not related to the expression of four cold-regulated genes. These results suggest that engineering of the biosynthesis of glycinebetaine by transformation with the codA gene might be an effective method for enhancing the freezing tolerance of plants. PMID- 10849361 TI - Epistatic repression of PHANTASTICA and class 1 KNOTTED genes is uncoupled in tomato. AB - Class 1 KNOTTED genes (KNOX) and PHANTASTICA (PHAN) are both central to meristem establishment and maintenance and, in maize and Antirrhinum, it has been proposed that PHAN acts as an epigenetic repressor of KNOX. In tomato, a distinct spatial distribution of Tkn2 KNOX transcripts compared to Antirrhinum and maize suggests either a different spatial distribution of tomato PHAN (Le-phan) transcripts, or that PHAN alone is insufficient for KNOX repression in tomato. We established the pattern of Le-phan expression, including a first demonstration of PHAN expression in healthy roots, and found Le-phan and Tkn2 transcripts to be temporally and spatially coincidental, with PHAN exhibiting an expression pattern in tomato distinct from that in plants with simple leaves. Our results imply that the expression of Le-phan is insufficient for the repression of Tkn2 in tomato and suggest an expanded role for either gene in the establishment of cell identity in plant development. PMID- 10849362 TI - A transformation vector for the production of marker-free transgenic plants containing a single copy transgene at high frequency. AB - We represent here the GST-MAT vector system. The R recombinase gene of the site specific recombination system R/RS from Zygosaccharomyces rouxii was fused to the chemical inducible promoter of the glutathione-S-transferase (GST-II-27) gene from Zea mays. Upon excision, the isopentenyltransferase (ipt) gene that is used as a selectable marker gene is removed. When the cauliflower mosaic virus 35S promoter (CaMV 35S) was used to express R recombinase, 67% of the marker-free transgenic plants had more than three transgene copies. Because the CaMV 35S promoter transiently and efficiently excised the ipt gene before callus and adventitious bud formation, the frequency of emergence of the ipt-shooty explants with a single T-DNA copy might be reduced. In this study we show that the GST-MAT vector efficiently produced transgenic ipt-shooty explants from 37 (88%) out of 42 differentiated adventitious buds and marker-free transgenic plants containing the GUS gene from five (14%) out of 37 ipt-shooty lines. Furthermore, the GST-MAT vector also induced two marker-free transgenic plants without the production of ipt-shooty intermediates. Southern blot analysis showed that six (86%) out of seven marker-free transgenic plants had a single GUS gene. This result suggests that the GST-MAT vector is useful to generate high frequency, marker-free transgenic plants containing a single transgene. PMID- 10849363 TI - Human T-cell receptor BV6 gene polymorphism in relation to expression level and CD4/CD8 skewness. AB - Using 50 samples of umbilical cord blood lymphocytes from Japanese donors, we analysed two human T-cell receptor beta variable (TCRBV) genes, BV6S4 and BV6S5, for their polymorphism, usage frequencies and CD4/CD8 skewness. They showed contrasting CD4/CD8 skewness, BV6S4 to CD8+ T cells and BV6S5 to CD4+ T cells. Genotyping of the BV6S4 alleles (A1, A2 and A3) revealed two of the six possible BV6S4 genotypes, A1/A2 and A2/A2. Among the two BV6S4 genotypes, no significant difference was detected in usage frequency or CD4/CD8 skewness. On the other hand, genotyping of the BV6S5 alleles (A1 and A2) revealed all three possible BV6S5 genotypes, A1/A1, A1/A2 and A2/A2, and the gene usage frequency was high, in the order A1/A1 > A1/A2 > A2/A2. These results indicate that the amino acid substitutions in BV6S5 (S36R and G70E) are in some way associated with the expression level of this gene. In the analysis of CD4/CD8 skewness, the three BV6S5 genotypes had similar skewness, indicating that A1 alleles are expressed more frequently than A2 alleles in both CD4+ and CD8+ T-cell populations. Although BV6S5 exhibits marked skewness to CD4+ T cells, our results indicate that the higher expression of A1 alleles is not associated with the increased ratio of CD4+ T cells. PMID- 10849364 TI - In situ expression of cytokines and cellular phenotypes in the lungs of mice with slowly progressive primary tuberculosis. AB - The cellular phenotypes and the expression of cytokines were studied in the lungs of mice, using immunohistochemistry, during different phases of slowly progressive primary murine tuberculosis infection. During the first phase the small focal lesions in healthy mice contained predominantly interleukin-2 (IL-2) expressing cells. A small number of tumour necrosis factor-alpha (TNF-alpha)-, monocyte chemoattractant protein-1 (MCP-1)- and IL-10-expressing cells were also present. IL-4-expressing cells were not detected. During the second phase the mice became unwell, but the bacterial counts and the size of focal lesions stabilized. IL-4-expressing cells appeared. The IL-10-, TNF-alpha- and MCP-1 expressing cells increased in number. On progression to phase three, the mice became seriously unwell and died rapidly. The inflammation spread to approximately 80% of the lung parenchyma. There was a marked increase in the number of IL-10-expressing cells. Expression of other cytokines was similar to that observed in the second phase. In the lesions, 3-6% of the macrophages (Mphi) containing mycobacterial antigens expressed high levels of IL-10 and TNF-alpha. The absolute numbers of CD3-, CD4- and CD11b-expressing cells in the lesions increased with the progression of infection. The numbers of CD8+ cells were reduced in the last phase of infection. The kinetics of T-lymphocyte subsets and the pattern of cytokine expression changed with the type and degree of tissue injury. The small number of Mphi with a heavy load of mycobacterial antigens may be the cause of this disturbance in cytokine balance, thus leading to progression of inflammation. PMID- 10849365 TI - Recruitment of SHP-1 protein tyrosine phosphatase and signalling by a chimeric T cell receptor-killer inhibitory receptor. AB - Receptors expressing the immunoreceptor tyrosine-based inhibitory motif (ITIM) in their cytoplasmic tail play an important role in the negative regulation of natural killer and B-cell activation. A subpopulation of T cells expresses the ITIM containing killer cell inhibitory receptor (KIR), which recognize MHC class I molecules. Following coligation of KIR with an activating receptor, the tyrosine in the ITIM is phosphorylated and the cytoplasmic protein tyrosine phosphatase SHP-1 is recruited to the ITIM via its SH2 domains. It is still not clear how SHP-1 affects T-cell receptor (TCR) signalling. In this study, we constructed a chimeric TCR-KIR receptor. We demonstrated that SHP-1 is recruited to the chimeric TCR-KIR receptor following T-cell stimulation with either anti TCR monoclonal antibody (MoAb) or superantigen. However, in spite of this we could not detect any effect of SHP-1 on TCR signalling regarding total protein tyrosine phosphorylation, TCR down-regulation, mobilization of intracellular free calcium, or induction of the activation markers CD69 and CD25. PMID- 10849366 TI - Co-induction of Mad1 and c-Myc in activated normal B lymphocytes. AB - The purpose of the present study was to examine the expression of the Myc network proteins c-Myc, Mad1 and Max in normal cells under different growth and differentiation conditions. A dominant view has been that Mad1 as a c-Myc antagonist plays a role in growth inhibition linked to differentiation. Of particular interest to us was therefore to study the regulation of Mad1 in cells undergoing differentiation in the absence of growth cessation. To do so we utilized normal B lymphocytes isolated from peripheral blood. The cells were induced to concomitant proliferation and differentiation by stimulation with a combination of anti-IgM antibodies (anti-mu) and the phorbol ester TPA. Thus, by 72 h of stimulation the percentage of plasmablasts increased from 3 to 17%, and the percentage of lymphocytes decreased from 89 to 27%. The most intriguing observation we made using this cell system was a pronounced coinduction of Mad1 and c-Myc. The levels of c-Myc and Mad1 mRNAs and proteins increased within 3 h of anti-mu stimulation, and the levels were further enhanced by TPA. Furthermore, the expressions of both c-Myc and Mad1 were reduced by forskolin, which also inhibited the anti-mu + TPA driven growth and differentiation of the B lymphocytes. The level of Max remained virtually unchanged. Taken together, our results indicate that a high level of Mad1 in normal human B cells is linked to differentiation and not to growth inhibition. Furthermore, the results demonstrate that Mad1 and c-Myc are not necessarily expressed in a reciprocal manner, which underlines an independent role of Mad1 unrelated to its function as a c-Myc antagonist. PMID- 10849367 TI - Identification and linkage of the proteasome activator complex PA28 subunit genes in zebrafish. AB - PA28 is an activator of the latent 20S proteasome, a large multisubunit complex involved in intracellular proteolysis. Two forms of hexameric PA28 have been identified, PA28-(alphabeta)3 and PA28-(gamma)6, of which the former is of immunological importance. Both the PA28-alpha and PA28-beta subunits are inducible by interferon-gamma (IFN-gamma) and the PA28-(alphabeta)3 complex enhances the ability of the 20S proteasome to produce peptides suited for binding to major histocompatibility complex (Mhc) class I molecules. To identify the homologues of the PA28 subunits in zebrafish we screened a cDNA library and obtained full-length cDNA sequences of the genes PSME1, PSME2 and PSME3 coding for the PA28-alpha, PA28-beta and PA28-gamma subunits, respectively. Phylogenetic analysis indicates the existence of the ancestors of all three genes prior to the divergence of tetrapods and bony fishes. The IFN-gamma-inducible subunits, PA28 alpha and PA28-beta, evolve faster than the presumably older PA28-gamma subunit. Using zebrafish radiation hybrid panels, the genes PSME2 and PSME3 were mapped to linkage group 12 and shown to be separated by a distance of less than 2.4 cM. This observation suggests that an intrachromosomal duplication event created the precursor of the IFN-gamma-inducible genes from a PA28-gamma-like ancestor prior to their recruitment into the Mhc class I peptide presentation pathway. PMID- 10849368 TI - Identification of seven genes in the major histocompatibility complex class I region of the zebrafish. AB - Physical linkage of genes whose products are involved in similar physiological pathways may have functional significance. The identification of conserved gene linkage in distantly related organisms can therefore strengthen the hypothesis of selection acting towards keeping genes on a chromosome. We used the cDNA selection technique and the polymerase chain reaction (PCR) with generic primers for the identification of new genes on the genomic clones bearing the major histocompatibility complex (Mhc) class I genes of the zebrafish (Danio rerio). We found six new genes (BING1, DAXX, TAPBP, KNSL2, TAP2B and KE6) whose orthologues are known to be linked to the Mhc class II region in humans and mice. In addition, a new zebrafish Mhc class I gene, termed Dare-UFA, was detected. By contrast, a search for the human leucocyte antigen (HLA)-linked BING3, KE3 and SACM2L genes revealed that these loci are not located on the class I clones of the zebrafish. The zebrafish class I region contains repetitive elements with similarity to the DANA, SATA and LINE repeats, as well as Tc1 transposable elements. Our findings indicate a high degree of linkage conservation between the zebrafish class I and the mammalian class II regions. PMID- 10849369 TI - Strain-specific variations in the development of dendritic cells in murine bone marrow cultures. AB - The dendritic cell (DC) is a professional antigen-presenting cell of central importance in immunity. In this paper, we examined DCs generated by 11-day culture of bone-marrow cells from the four mouse strains C57BL/6J, BALB/cA, C3H/HeN and B10.PL-H2u (73NS)/Sn with respect to cell yield as well as surface marker phenotype and morphology. We also investigated the phenotypic changes and the T-cell stimulatory activity of the DCs induced by bacterial lipopolysaccharide (LPS). Morphologically, we observed low levels (5-10%) of granulocyte contamination of the cultures after a culture period of 11 days. Considerable strain-specific differences were found in the expression levels of the surface markers in addition to the differences in the ratio of the immature to mature DCs in the cultures that were not stimulated with LPS. Furthermore, we found that LPS strongly induces maturation of DCs in all strains investigated with the exception of the B10.PL strain. PMID- 10849370 TI - CD4+ T cells of schistosomiasis naturally resistant individuals living in an endemic area produce interferon-gamma and tumour necrosis factor-alpha in response to the recombinant 14KDA Schistosoma mansoni fatty acid-binding protein. AB - Cellular immune responses to recombinant (r) Sm14 were examined in chronic, treated patients and uninfected individuals living in an endemic area for schistosomiasis. The lymphocyte proliferative responses and cytokine profile to this antigen were evaluated. Peripheral blood mononuclear cells (PBMC) of all groups studied proliferated to rSm14. However, the highest proliferation index to rSm14 was detected in uninfected endemic normal (EN) individuals who are naturally resistant to schistosomiasis. Regarding the cytokines produced, the levels of interleukin (IL)-5 and IL-10, known as Th2 cytokines, were not statistically different among all groups studied. In contrast, interferon (IFN) gamma and tumour necrosis factor (TNF)-alpha were produced in significantly higher amounts by PBMC of EN individuals following rSm14 stimulation. Additionally, we have determined by flow cytometry that CD4+ T cells from these individuals are the main lymphocyte subpopulation producing IFN-gamma and TNF alpha. Moreover, we have used rIL-10 or rIFN-gamma, or monoclonal antibodies (MoAb) against these two cytokines to determine their role on cellular reactivity to rSm14. Exogenous IL-10 suppressed T-cell proliferation and neutralization of endogenous IL-10 restored lymphocyte activation and enhanced IFN-gamma and TNF alpha production in chronically infected patients. In contrast, the addition of anti-IFN-gamma totally abrogated the PBMC proliferation within the EN group. This study demonstrated that IL-10 is an important cytokine down-regulating T-cell responses in chronic schistosomiasis, whereas lymphocyte proliferation in the uninfected resistant group is dependent on IFN-gamma. Taken together these results suggest that Th1 type of immune response induced in EN individuals to a specific schistosome antigen might be associated with resistance to infection and also highlighted the importance of Sm14 as a potential vaccine candidate. PMID- 10849371 TI - The amino acid sequence of a monoclonal gamma 3-heavy chain from a patient with articular gamma-heavy chain deposition disease. AB - Abnormal deposition of proteins, including monoclonal immunoglobulin gamma-heavy chains, may cause tissue damage and organ dysfunction. We here report the amino acid sequence of the free gamma-heavy chains present in serum and urine of the first reported case (patient G. L.) of synovial heavy chain deposition disease. The protein was heavily deleted and consisted of the hinge, in addition to the CH2 and CH3 domains, in a dimeric form, thus lacking its variable domain as well as the CH1 domain. The sequence was consistent with the gamma 3 subclass (gamma 3GL). Gm typing revealed the gamma 3 allotypes G3m(b0) and G3m(b1) in accordance with the residues Pro123, Phe128, Thr171 and Phe268 in gamma 3GL. Furthermore, the gamma 3GL molecule was glycosylated at Asn in position 129. Finally, the gamma 3GL protein was shown to contain a typical binding site for the first complement component, C1q, namely the residues Glu150, Lys152 and Lys154, with the potential of binding and activating complement, causing tissue damage following deposition. PMID- 10849372 TI - Fine specificity of anti-fibrillin-1 autoantibodies in primary pulmonary hypertension syndrome. AB - Autoantibodies to fibrillin-1 (Fbn-1) have been found in systemic sclerosis (SSc), calcinosis, Raynaud's esophagael dysmotility, sclerodectyly, and telaengectasia (CREST) and mixed connective tissue disease (MCTD) diseases. The purpose of this study was to determine whether patients with primary pulmonary hypertension (PPH) and appetite-suppressant-associated PPH have anti-Fbn-1 autoantibodies. In addition we assessed the human leucocyte antigen (HLA) class II alleles (DRB1, 3, 4, 5 and DQB1) in these patients in order to determine whether the response is genetically restricted. The frequency of anti-Fbn-1 autoantibodies in patient groups was compared with that of a control group of 88 healthy patients, and HLA was correlated similarly with a group of 51 healthy subjects. Anti-Fbn-1 autoantibodies were found at high frequency in PPH: in 70 of 75 adults with PPH (93%), in 28 of 33 children with PPH (84.8) and in 12 of 18 (67%) patients with appetite-suppressant-associated PPH. Utilization of two Fbn-1 fusion proteins allowed us to determine the dominant determinant region, recognized by anti-Fbn-1 autoantibodies, which may be located on the N-terminal fragment of the Fbn-1 protein. No significant immunogenetic correlations were found when the PPH patient groups were compared with normal controls. This novel category of autoantibodies is found in diseases characterized by endothelial and extracellular matrix protein alterations and fibrosis. PMID- 10849373 TI - Anti-SSA/Ro antibody determination by enzyme-linked immunosorbent assay as a supplement to standard immunofluorescence in antinuclear antibody screening. AB - The aim of this study was to investigate the frequency and possible clinical relevance of SSA/Ro antibodies, as determined by enzyme-linked immunosorbent assay (ELISA), in patient sera not exhibiting a concomitant positive reaction by the standard immunofluorescence (IF) test using HEP-2 cells as substrate. SSA/Ro reactivity, as shown by ELISA, was found in 285 (7%) of 4025 serum samples consecutively remitted for antinuclear antibody (ANA) screening. Seventy-five of these serum samples (26%), derived from 64 patients, were negative by the IF-ANA screening test. Serum samples from all 64 patients exhibiting SSA/Ro reactivity by ELISA without concomitant positivity by IF-ANA were further investigated by IF using transfected HEP-2 cells hyperexpressing the 60,000 MW SSA/Ro antigen (HEP 2000(R)) and by immunodiffusion (ID) and Western blot. In 55 of these 64 patients, SSA/Ro reactivity could be verified by one or more of the other techniques investigated. Twelve of these patients fulfilled four or more American College of Rheumatology (ACR) criteria for systemic lupus erythematosus (SLE) and another five patients exhibited a histologically confirmed cutaneous lupus erythematosus (LE). In four of the 12 IF-ANA-negative patients with a diagnosis of SLE, the SSA/Ro reactivity was only detectable by ELISA and Western blot. In conclusion, the use of a sensitive ELISA assay could provide a clinically important supplement to the routine ANA screening by IF, which does not detect certain anti-SSA/Ro-containing sera among patients with relevant autoimmune diagnoses. Detection of anti-SSA/Ro antibodies, however, does not alone signify cutaneous LE or SLE but adds weight to these diagnoses that should rely heavily on other clinical information. PMID- 10849374 TI - Interleukin-12 induces efficient lysis of natural killer-sensitive and natural killer-resistant human osteosarcoma cells: the synergistic effect of interleukin 2. AB - Previously we demonstrated that some osteosarcoma cell lines varied greatly in their susceptibility to natural killer (NK) cell lysis in vitro. The expression of CD54 and CD58 adhesion molecules on their surface appeared to influence their vulnerability, and the tumour necrosis factor-alpha (TNF-alpha)-induced positive modulation of CD54 increased osteosarcoma susceptibility in vitro. This study investigated whether peripheral blood mononuclear cells from normal healthy donors could be activated by interleukin (IL)-12 and IL-2, separately or in combination, to lyse osteosarcoma cell lines in vitro, as evaluated by using a microcytotoxicity test. In addition, we analysed (by flow cytometry) whether this function correlated with modifications of the CD2, CD11a, CD11b and CD18 molecules, which are involved in the adhesion of effector cells to the counter receptors (CD54 and CD58) on osteosarcomas. This study demonstrates that incubation with IL-12 and/or IL-2 triggered NK cell cytolytic activity against osteosarcoma targets and that cytolytic activity was enhanced to a greater extent when lymphocytes were incubated simultaneously with a combination of IL-12 and IL 2. The density of CD18 and CD2 molecules involved in NK adhesion was also up modulated following cytokine incubation. These changes in the density of adhesion molecules can be involved in the increased lytic activity of effector lymphocytes and in the modification of their binding capacity to osteosarcoma target cells. PMID- 10849375 TI - Azathioprine associated T-cell mutations in insulin-dependent diabetes mellitus. AB - Somatic mutations arise regularly in human T lymphocytes. As these events occur at increased frequencies in several autoimmune disorders, presumably because of increased T-cell proliferation, we investigated if this is also true for insulin dependent diabetes mellitus (IDDM). Mutations of the hypoxanthine guanine phosphoribosyltransferase (hprt) gene measured by 6-thioguanine (TG) selection were studied in 28 patients (60 determinations) enrolled in a prospective double blinded placebo-controlled study of azathioprine immunosuppression: 17 patients (34 determinations) were receiving azathioprine and 11 (26 determinations) placebo. Mean hprt T-cell mutant frequencies (MFs) were elevated in both patient groups, but only in the azathioprine group were elevations large and statistically correlated with the duration of the therapy. These results suggest that the organ-specific antigenic stimulus of the T-cell proliferation in IDDM does increase mutant cells in the peripheral blood, but this increase is relatively small. However, azathioprine, which is converted to 6-mercaptopurine in vivo, selects and amplifies the hprt mutants that do arise. Clinical azathioprine resistance may be explained by hprt mutations arising in T cells relevant to the underlying autoimmune process. Monitoring for these mutations should allow more effective use of this immunosuppressive agent. PMID- 10849376 TI - Rituximab (anti-CD20) therapy of B-cell lymphomas: direct complement killing is superior to cellular effector mechanisms. AB - Rituximab (IDEC-C2B8, Mabthera(R)) is a chimeric (human-mouse) monoclonal antibody (MoAb) against the B-cell specific CD20-antigen. It has been used for the clinical treatment of non-Hodgkin's lymphomas, but variable clinical results suggest that some lymphoma cells remain resistant. In the present study we have evaluated the relative efficiencies of humoral and cell-mediated effector mechanisms complement-dependent cytotoxicity (CDC), antibody-(ADCC), complement (CDCC) dependent cellular cytotoxicity and apoptosis on lymphoma cell killing by rituximab. Rituximab activated the cytolytic complement (C) cascade and induced a strong CDC, but the rituximab-triggered ADCC and CDCC were relatively ineffective. The CDC was strongly enhanced by antibodies against the C inhibitor CD59 (protectin). Neutralization of CD55 (DAF) and CD46 (MCP) had a similar but weaker effect. Rituximab also induced apoptosis but in a cell line-dependent fashion. The results strongly emphasize the role of direct CDC as the major, fast and efficient effector mechanism of rituximab. In the immunotherapeutic treatment of B-cell lymphomas, it is important to consider the role of C-regulatory proteins as an escape mechanism of the malignant cells. Our results suggest that the effect of rituximab therapy could be enhanced by combining it with neutralization of CD59. PMID- 10849377 TI - Quality of transfusion practice beyond the blood transfusion laboratory is essential to prevent ABO-incompatible death. PMID- 10849378 TI - 'Noise' in microbiological screening assays. PMID- 10849379 TI - Low ferritin levels indicate the need for iron supplementation: strategy to minimize iron-depletion in regular blood donors. AB - Iron deficiency is a common problem in regular blood donors which can be prevented by timely iron supplementation. Consequently, these donors should be supplied with oral iron in good time. We evaluated the need to use ferritin rather than or in addition to haemoglobin to screen iron deficiency in blood donors. To this end, serum ferritin was measured routinely every 10th donation in 632 long-term and 171 first-time donors. Furthermore, donors with ferritin < 15 microg L-1 were supplemented with iron. The supplementation efficiency was assessed by follow-up haemoglobin levels over the course of five donations in blood donors with high donation frequency. Our results showed that ferritin decreases after 10 donations and with the increase of donation frequency. In 26% of regular donors, ferritin levels were < 15 microg L-1 and 12% of them were anaemic due to low haemoglobin. After iron supplementation, haemoglobin was raised rapidly in donors with initially low haemoglobin, and thus donor deferment was never indicated. In conclusion, regular ferritin measurement is a useful indicator for iron depletion in blood donors. Our data suggested the usefulness of ferritin screening in first-time donors and regular donors with low haemoglobin levels within the normal range. PMID- 10849380 TI - Bruising following blood donation, its management and the response and subsequent return rates of affected donors. AB - A study was undertaken to determine the incidence of bruising among blood donors and to analyse their response to the management of this complication. A total of 52 510 donors were bled at 476 consecutive donor sessions held by the Brentwood Centre during a 4-month period. Of these, 344 donors (0.66%) were found to have developed bruises following venepuncture. The incidence of bruising among males was 0. 35% and that among females was 0.98%. All bruised donors were managed by the Centre nursing and medical staff. One hundred and sixty-one donors informed the Centre that they were fully satisfied with the way their bruising was managed. Of 329 bruised donors who remained in the panel, 249 (75.7%) attended subsequent blood donor sessions in response to routine invitations, showing that the majority of bruised donors continued to donate blood. This response was compared with that of a control group of donors who did not develop any complications and there was no significant difference in the return rates between the two groups. PMID- 10849381 TI - The BCSH guideline on addressograph labels: experience at a cardiothoracic unit and findings of a telephone survey. AB - In 1998 we implemented a BCSH recommendation that addressograph labels should not be used on blood transfusion specimen tubes. Over a 12-month period before the ban was introduced our laboratory received 5964 red cell transfusion requests, 182 (3.1%) of which contained an error in the identification details (ID) supplied on the request form and/or specimen. Three of these errors were of the 'wrong patient' type, i.e. the sample belonged to a different patient from the one whose ID appeared on the specimen tube and request form. Over the 12 months after the ban was introduced 511 (8. 1%) of 6326 requests contained a labelling error, an increase in error rate of 165%; no wrong-patient errors were identified, however. In a survey, seven (29.2%) of 24 transfusion laboratories in the UK accepted specimens labelled with addressograph stickers; in four of these cases a local blood transfusion committee had agreed that the BCSH guideline should not be followed. We believe the BCSH guideline is valid; its implementation, however, has major financial and workload implications, which probably explains why many hospitals apparently do not comply with it. PMID- 10849382 TI - Improvement in transfusion safety using a specially designed transfusion wristband. AB - Fatal haemolytic transfusion reaction due to ABO incompatibility occurs mainly as a result of clerical error. A blood sample drawn from the wrong patient and labelled as another patient's will not be detected by the blood bank unless there is a previous ABO grouping result. We report here the detection of such clerical error by the use of a specially designed transfusion wristband. The wristband has the following special features: (i) once attached, it cannot be removed except by cutting; (ii) it has a pocket containing a transfusion label; (iii) a unique transfusion barcode is printed on each transfusion label and the corresponding wristband simultaneously by computer technology; (iv) a transfusion label removed from the wristband after attachment to the patient has a characteristic tear-mark distinguishing it from one removed prior to attachment. The blood bank only accepted those specimens bearing the tear-marked transfusion labels. All blood units for this patient were labelled with this unique transfusion code together with the patient's details. The nurses counter-checked the transfusion code on the blood units against the transfusion code on the patient's transfusion wristband prior to transfusion. If the blood sample for compatibility testing was drawn from the 'wrong' patient, the intended patient either did not carry a wristband or the transfusion codes did not match at all. Pretransfusion compatibility tests were performed on 2189 patient samples using this procedure. It was well accepted by both ward and blood bank staff. Two potential mismatched transfusions were avoided. These two clerical errors would not have been detected because neither patient had previous ABO grouping results. PMID- 10849383 TI - Human plasma and tirilazad mesylate protect stored human erythrocytes against the oxidative damage of gamma-irradiation. AB - Transfusion-associated graft-versus-host disease (TA-GVHD) is a serious condition that under certain circumstances can be lethal in immunosuppressed patients. The risk of TA-GVHD can be reduced in this population by gamma irradiation (gammaRad) of blood components. gammaRad results in production of reactive oxygen species which can damage red blood cells (RBC). Tirilazad mesylate (TM) is a member of the 21-aminosteroids (Lazaroids) family and is a powerful antioxidant. We investigated the ability of TM and human plasma (which contain powerful antioxidants) to protect stored human RBC against the oxidative damage of gammaRad. Fresh intact packed RBC obtained from the normal donors, with and without autologous plasma or TM (0.05 mg mL-1 RBC), were exposed to gammaRad (50 Gy) and stored for 28 days at 4 degrees C. Oxidative damage was assessed by osmotic fragility at 65 mM NaCl concentration (expressed by percentage haemolysis in 65 mM NaCl solution) and lipid peroxidation (measured by thiobarbituric acid reactive substances, TBARS). Our results showed that storage and irradiation of untreated intact RBC increased the osmotic fragility at 65 mM NaCl concentration (65.8 +/- 1.3 vs. 51.20 +/- 0.87% haemolysis; irradiated vs. controls, respectively; P = 0.002) and lipid peroxidation (TBARS = 4.47 +/- 0. 12 vs. 3.45 +/- 0.09 microM L-1 RBC; irradiated vs. controls, respectively; P = 0.001). TM protected the intact RBC against radiation-induced haemolysis (35.8 +/- 5.0 vs. 65.8 +/- 1.3% haemolysis; treated vs. untreated irradiated RBC, respectively; P = 0.02) and lipid peroxidation (TBARS = 2.91 +/- 0.2 vs. 4.47 +/- 0.12 microM L-1 RBC; treated vs. untreated irradiated RBC, respectively; P = 0.005). Addition of autologous plasma to packed RBC significantly reduced the extent of radiation induced haemolysis by more than six-fold (12.45 +/- 0.26 vs. 65.8 +/- 2.2% haemolysis; irradiated RBC with versus without plasma, respectively; P = 0.0001). In conclusion, these results show that irradiation and storage of blood damages RBC via oxidative processes and addition of autologous plasma and/or TM protects RBC against such damage and possibly enhances their storage and survival. PMID- 10849384 TI - Comparison between a new PVC platelet storage container (UPX80) and a polyolefin container. AB - During storage of platelet concentrates (PCs), the quality of the platelets deteriorates gradually, partially dependent on gas exchange. UPX80 (JMS, Japan) 1 L platelet storage PVC containers with increased gas transport capacity were compared with 1- and 1. 5-L polyolefin (PO) containers (NPBI, the Netherlands) with filtered PCs stored either in GAC (gluconate-acetate-citrate, < 10% plasma) or in plasma, for 8 days. In total 32 PCs were made (260-330 x 109 platelets per concentrate), equally divided over different bags and storage media. During storage, gas exchange, metabolic, physical and activation parameters were measured. No consistent differences for all parameters were observed between UPX80 and PO containers (1-L or 1.5-L). Blood gas parameters indicated better gas exchange for UPX80 containers compared with PO containers. Good morphology was observed in UPX80 and metabolic functions were not significantly different compared with PO containers. During prolonged storage (after day 6), some significant differences in CD62P and CD63 expression were found, indicating a higher degree of platelet activation in UPX80 containers, especially in GAC. UPX80 PC containers are suitable for storage of PCs. Although in UPX80 better gas exchange is demonstrated, as compared with PO containers, this does not improve the platelet quality during storage for 6 days, indicating that gas exchange above the level of PO containers has no effect on the switch to aerobic metabolism in platelets. PMID- 10849385 TI - Alloimmunization in Chinese with warm autoimmune haemolytic anaemia - incidence and characteristics. AB - We perform a retrospective study to determine the incidence and characteristics of alloimmunization in Chinese patients with warm autoimmune haemolytic anaemia. Among 67 patients studied, clinically significant alloantibodies were found in only eight patients, giving a rate of alloimmunization of 11.3%. The latter contrasts with the much higher rate reported in the Western population. This probably reflects the genetic homogeneity in Chinese with respect to the red cell phenotype. The alloimmunization rate, however, is still significant and therefore comprehensive pretransfusion testing is required for Chinese patients with warm autoantibodies so as to safeguard patient's safety. PMID- 10849386 TI - A murine monoclonal antibody against Kx protein which reacts also with beta spectrin. AB - Kx is a polytopic membrane protein of human erythrocytes carrying the Kx blood group antigen, which is deficient in rare patients with McLeod syndrome. Kx is disulphide bond linked to the Kell glycoprotein, which is a bitopic type II membrane protein carrying the Kell blood group antigen. Mice immunized with a synthetic peptide predicted to be located on the second external loop of Kx produced a monoclonal antibody called 3E12 which does not recognize red cells with common Kell phenotype by agglutination and flow cytometry. 3E12 recognizes the Kx protein and the spectrin beta-chain on western blots, the affinity for these two proteins being lowered with increasing ionic strength. Linear epitopes recognized by 3E12 are E116EIEKE121 and L484AQELEKE491 on the Kx protein and spectrin beta-chain, respectively. To quantify the relative amount of Kx in Empigen BB extracts of red cell membranes, an ELISA for Kx was set up which showed conclusively that (i) there is less Kx in membranes of K0 individuals (lacking the Kell glycoprotein) than in membranes of common individuals, and (ii) that all common individuals, typed as K+k-, K-k+ and K+k+, have the same amount of Kx on their red cell membranes. When an erythrocyte membrane detergent extract from one K0 individual was chromatographed on an immobilized 3E12 column, a minute amount of authentic Kell glycoprotein was recovered in acid eluted fractions, indicating that at least the K0 individual under study may still produce some Kell protein. PMID- 10849387 TI - Looking backward and forward-from the eye of the editor. PMID- 10849388 TI - Dissociated vertical deviation: etiology, mechanism, and associated phenomena. Costenbader Lecture. AB - PURPOSE: The etiology and mechanism of dissociated vertical deviation (DVD) are explored. METHODS: In 6 young adults with DVD, the simultaneous horizontal, vertical, and torsional eye movements for both eyes were recorded by using dual coil scleral search coils. Analysis of the simultaneous vertical and torsional movements that occurred during the DVD response identified the primary muscles acting in the vergences and versions involved. RESULTS: Typically, both horizontal and cyclovertical latent nystagmus developed upon occlusion of either eye. A cycloversion/vertical vergence then occurred, with the fixing eye intorting and tending to depress and the covered eye extorting and elevating. Simultaneously, upward versions occurred for the maintenance of fixation, consisting of various saccades and smooth eye movements, and this led to further elevation of the eye behind the cover. The cyclovertical component of the latent nystagmus became partially damped as the DVD developed. CONCLUSIONS: In patients with an early onset defect of binocular function, the occlusion of one eye, or even concentration on fixing with one eye, produces unbalanced input to the vestibular system. This results in latent nystagmus with a cyclovertical component, sometimes only seen with magnification. A normal, oblique-muscle produced, cycloversion/vertical vergence then comes into play, occurring in an exaggerated form in the absence of binocular vision, probably as a learned response. This cycloversion/vertical vergence helps damp the cyclovertical nystagmus (a cyclovertical "nystagmus blockage" phenomenon), aiding vision in the fixing eye. But this mechanism also produces unavoidable and undesirable elevation and extorsion of the fellow eye, which we call DVD. PMID- 10849389 TI - Botulinum toxin treatment versus conservative management in acute traumatic sixth nerve palsy or paresis. AB - PURPOSE: Botulinum toxin (BTX), injected into the ipsilateral medial rectus muscle, has been advocated for the management of acute traumatic sixth nerve palsy or paresis. We conducted a multicenter, nonrandomized, data collection study to evaluate recovery rates of patients treated with either conservative measures or BTX. METHODS: All members of the American Association for Pediatric Ophthalmology and Strabismus and the North American Neuro-Ophthalmology Society were invited to enroll patients with acute traumatic sixth nerve palsy or paresis during a 2-year period (between March 1996 and February 1998). The BTX group was defined as patients who received a BTX injection within 3 months of injury. Recovery at 6 months from injury was defined as absence of diplopia in the primary position and a distance esotropia of no more than 10 PD in the primary position. Nonrecovered patients with less than 6 months of follow-up (n = 15) were excluded. RESULTS: Eighty-four eligible patients were enrolled by 46 investigators. Sixty-two patients (74%) were treated conservatively and 22 (26%) with BTX. Sixty-two patients (74%) had unilateral palsy, and 22 (26%) had bilateral palsy. Recovery rates were similar between BTX and conservatively treated patients (overall: 73% vs 71%, P = 1.0; unilateral: 81% vs 83%, P = 1.0; bilateral: 50% vs 38%, P = 0.66, respectively). CONCLUSIONS: In this prospective multicenter study of acute traumatic sixth nerve palsy or paresis, patients treated with either BTX or conservative measures had similar high recovery rates. PMID- 10849390 TI - Recognized globe perforation during strabismus surgery: incidence, risk factors, and sequelae. AB - BACKGROUND: Inadvertent perforation of the globe is a well-recognized complication of extraocular muscle surgery. We evaluated the incidence, risk factors, and sequelae of this complication at our institution. METHODS: Medical records of patients who underwent extraocular muscle surgery at King Khaled Eye Specialist Hospital, Saudi Arabia, between September 1983 and April 1997, were reviewed for the occurrence of globe perforation. We documented preoperative visual acuity and refraction, surgical procedure, how the perforation occurred, and immediate management, as well as the sequelae of the perforation, its management, and final outcome. RESULTS: Recognized perforations occurred in 15 of 4886 procedures, for an overall incidence rate of 3/1000. Perforations were 3 times more common in myopic eyes (>-6.00 D, P =.05) and 2 times more common in eyes with previous extraocular muscle surgery. Perforations occurred during muscle reattachment (5 cases), placement of traction sutures at the limbus (4 cases with transient hyphema), muscle disinsertion (3 cases), and placement of sutures at the muscle insertion before disinsertion (3 cases). One patient had a large scleral laceration with uveal prolapse, necessitating scleral patch graft at the time of surgery, and later had retinal detachment surgery with loss of 2 lines of visual acuity. Endophthalmitis, cataract, glaucoma, and phthisis bulbi were not encountered in our review. CONCLUSION: The current incidence of globe perforation is low and only rarely associated with serious sequelae. PMID- 10849391 TI - The use of binocular visual acuity in the assessment of intermittent exotropia. AB - BACKGROUND: It has been suggested that a decrease in distance stereoacuity in patients with intermittent exotropia is a good indicator of diminishing control. However, there has been no adequate explanation for this reported reduction in distance stereoacuity in these patients. We postulate that the decrease in stereoacuity is related to blurred visual acuity created by an increasing demand on accommodation, which these patients use in an attempt to control the exodeviation. This can best be assessed by measuring binocular visual acuity (BVA). Analysis of BVA could provide a useful clinical tool to evaluate control measures used by patients with intermittent exotropia. METHODS: A prospective study of patients with intermittent exotropia, ranging in age from 6 to 60 years, was performed. Only those patients with the presence of either basic or divergence excess (simulated or true) type exodeviation were included in the study. The data analysis included the age of these patients, age at onset of the deviation, monocular and binocular visual acuity, oculomotor and fusional status, and near and distance stereoacuity. RESULTS: Data from 36 patients show that the measurements of BVA correlated well with a corresponding loss of distance stereoacuity but not with the size of the deviation. CONCLUSION: The decrease of stereoacuity reported in patients with exotropia can be explained by increased accommodation and decreased distance BVA. This measurement can be a simple method of quantifying the fusional control of patients with intermittent exotropia. PMID- 10849392 TI - Bilateral idiopathic Brown's syndrome with delayed onset in the second eye. AB - PURPOSE: We describe 6 cases of a previously unreported variation of bilateral Brown's syndrome that presented in congenital form in one eye and developed later in the fellow eye with no underlying cause. METHODS: We reviewed the clinical records of 6 patients from 6 separate practices to determine whether there were any common clinical features on presentation or in their clinical courses. RESULTS: All 6 patients were diagnosed with unilateral congenital Brown's syndrome at the first ophthalmologic assessment but showed no evidence of the syndrome in the fellow eye. In 5 cases the contralateral syndrome developed in the second eye after surgery was performed on the first eye, and in 1 case it developed before any surgery was done. The ages at onset of the syndrome in the second eye ranged from 2 to 8 years. None of the children had any evidence of systemic illness or local orbital disease to explain an acquired Brown's syndrome. CONCLUSION: To our knowledge, this is the first reported series of cases of bilateral Brown's syndrome that manifested sequentially in the eyes with no known causes for an acquired syndrome in the second eye. This finding supports the premise that congenital and acquired Brown's syndrome are on a continuum with a common pathophysiology of restriction of free movement of the superior oblique tendon in the trochlea. PMID- 10849393 TI - Surgical management of strabismus after rupture of the inferior rectus muscle. AB - BACKGROUND: Rupture of an inferior rectus muscle is an uncommon problem. The resulting absence of infraduction and large hypertropia that result when the muscle cannot be repaired are challenging to manage surgically. METHODS: We treated 2 patients who had traumatic rupture of the inferior rectus muscle. Both patients underwent an inferior transposition of the inferior halves of the medial and lateral rectus muscles without disinsertion (modified Jensen transposition procedure). RESULTS: Both patients had a persistent small overcorrection in the primary gaze position. One patient was treated with a second strabismus surgery consisting of a recession of the contralateral superior rectus muscle; the other was treated with prism glasses. Both achieved restoration of depression to approximately 40 degrees and single binocular vision in the primary position at distance, near, and in the reading position. CONCLUSION: This modified Jensen transposition procedure of the horizontal rectus muscles appears to be highly effective in the treatment of the hypertropia and infraduction deficit produced by rupture of the inferior rectus muscle. It also appears to be suitable for use in situations when other rectus muscles are absent or unavailable for surgical manipulation. PMID- 10849394 TI - Anomalous retinal correspondence: neuroanatomic mechanism in strabismic monkeys and clinical findings in strabismic children. AB - BACKGROUND: Anomalous retinal correspondence (ARC) is a neural adaptation to eye misalignment in which non-corresponding retinal points are linked in the visual cortex to provide binocular fusion. ARC within the striate cortex would require that horizontal neurons link right-eye and left-eye ocular dominance columns (ODCs) separated by a distance in the cortex proportional to the angle of strabismus. Two hypothetical mechanisms are possible: (1) The ODCs can be linked by axons of horizontal neurons that project monosynaptically from a right-eye to a left-eye ODC. The further apart the ODCs, the longer the axons; hence, axon length should be greater in subjects with strabismus than in healthy subjects (elongated axon, monosynaptic hypothesis). In this case, the clinical probability of developing ARC should be independent of the angle of strabismus, until an upper-limit angle of strabismus is reached equally to the maximal length of axons available to link nonadjacent ODCs, at which point an abrupt decline of ARC probability should be evident. (2) Alternatively, ODCs can be linked by a chain of horizontal neurons, the number of which increases as the distance among ODCs increases; axon length in subjects with strabismus would be expected to be the same as in healthy subjects (normal axon, polysynaptic hypothesis). In this case, the greater the angle of strabismus, the more horizontal neurons and synapses required for linkage, and the greater the probability of signal degradation. Thus, the clinical probability of developing ARC through a polysynaptic mechanism should be inversely proportional to the angle of strabismus. The purpose of this study was to test these 2 hypotheses neuroanatomically in primates and clinically in children. METHODS: For the neuroanatomic portion of the study, biotinylated dextran amine was injected into ODCs of area V1 to label individual neurons. The length of the horizontal axons from these neurons was then compared in strabismic and normal monkeys. In the clinical portion of the study, the medical records of 192 children with strabismus were reviewed retrospectively. The angle of strabismus (prism cover test) and the presence of ARC (Bagolini striated lenses, Worth/Polaroid 4-dot) were recorded. Plots of the presence of ARC as a function of the angle of strabismus were obtained. RESULTS: There was no significant difference in axon length between healthy (7. 02 +/- 0. 83 mm) and strabismic monkeys (6. 60 +/- 1. 07 mm) (P =.16). In children with strabismus, ARC decreased as the angle of strabismus increased (P <. 05). ARC was more prevalent in children who had primary or postsurgical deviations of /=10(20) eV. Because the expected rate of AIC sources in our Galaxy is very small ( approximately 10(-5) yr(-1)), the corresponding contribution to the flux of UHECRs is negligible and the total flux is given by the integrated contribution from AIC sources produced by the distribution of galaxies located within the distance that is unaffected by the GZK cutoff ( approximately 50 Mpc). We find that reconnection should convert a fraction xi greater, similar0.1 of magnetic energy into UHECRs in order to reproduce the observed flux. PMID- 10849414 TI - The Missing Link: Early Methane ("T") Dwarfs in the Sloan Digital Sky Survey. AB - We report the discovery of three cool brown dwarfs that fall in the effective temperature gap between the latest L dwarfs currently known, with no methane absorption bands in the 1-2.5 um range, and the previously known methane (T) dwarfs, whose spectra are dominated by methane and water. The newly discovered objects were detected as very red objects in the Sloan Digital Sky Survey imaging data and have JHK colors between the red L dwarfs and the blue Gl 229B-like T dwarfs. They show both CO and CH(4) absorption in their near-infrared spectra in addition to H(2)O, with weaker CH(4) absorption features in the H and K bands than those in all other methane dwarfs reported to date. Due to the presence of CH(4) in these bands, we propose that these objects are early T dwarfs. The three form part of the brown dwarf spectral sequence and fill in the large gap in the overall spectral sequence from the hottest main-sequence stars to the coolest methane dwarfs currently known. PMID- 10849415 TI - A Very Fast and Momentum-conserving Tree Code. AB - The tree code for the approximate evaluation of gravitational forces is extended and substantially accelerated by including mutual cell-cell interactions. These are computed by a Taylor series in Cartesian coordinates and in a completely symmetric fashion, such that Newton's third law is satisfied by construction and that therefore momentum is exactly conserved. The computational effort is further reduced by exploiting the mutual symmetry of the interactions. For typical astrophysical problems with N=105 and at the same level of accuracy, the new code is about 4 times faster than the tree code. For large N, the computational costs are found to scale almost linearly with N, which can also be supported by a theoretical argument, and the advantage over the tree code increases with ever larger N. PMID- 10849416 TI - Sub-Saturn Planetary Candidates of HD 16141 and HD 46375. AB - Precision Doppler measurements from the Keck High-Resolution Echelle Spectrograph reveal periodic Keplerian velocity variations in the stars HD 16141 and HD 46375. HD 16141 (G5 IV) has a period of 75.8 days and a velocity amplitude of 11 m s-1, yielding a companion having Msini=0.22 M(JUP) and a semimajor axis of a=0.35 AU. HD 46375 (K1 IV-V) has a period of 3.024 days and a velocity amplitude of 35 m s 1, yielding a companion with Msini=0.25 M(JUP), a semimajor axis of a=0.041 AU, and an eccentricity of 0.04 (consistent with zero). These companions contribute to the rising planet mass function toward lower masses. PMID- 10849417 TI - Search for Transits of a Short-Period, Sub-Saturn Extrasolar Planet Orbiting HD 46375. AB - Precise brightness measurements of HD 46375 have been acquired with an automatic telescope to search for transits of its short-period, sub-Saturn extrasolar planet. Transits of the companion do not occur, indicating that the inclination of the orbit i is less than 83 degrees and sini is less than 0.992. This upper limit on sini still preserves the possibility that the mass of the planet is less than Saturn's. Analysis of the photometry for HD 46375 reveals no photometric variability larger than 0.0001+/-0.0002 mag at the orbital period of the planet. This effectively eliminates starspots and stellar pulsations as the cause of the radial velocity variations used to infer the planet's existence. PMID- 10849418 TI - Supernova Sources and the 92Nb-92Zr p-Process Chronometer. AB - We report new Zr isotope evidence for live (92)Nb (mean life: tau&d1;92Nb=52 Myr) within the early solar system resulting in &parl0;92Nb&solm0;93Nb&parr0;initial approximately 10-3. The meteoritic minerals rutile and zircon have, respectively, very high and very low Nb/Zr ratios and are ideal for exploring the (92)Nb-(92)Zr chronometer. Rutiles exhibit high positive straightepsilon92Zr ( approximately 14 36) while a zircon has a negative straightepsilon92Zr ( approximately -4), as would be expected if (92)Nb was live in the early solar system. The meteoritic rutiles appear to be young, with apparent times of formation of approximately 80 220 Myr subsequent to the origin of the solar system. The initial (92)Nb/(92)Mo for the solar system is broadly compatible with a model of uniform production if the (92)Nb/(92)Mo production ratio for Type II supernova (SNII) sources with neutrino-driven winds is used. Data for all the now extinct p-process nuclides ((92)Nb, (97)Tc, and (146)Sm) are consistent with these isotopes being derived by uniform production from SNII sources and a free decay interval of approximately 10 Myr. Consideration of a range of models indicates that the average p-process production ratio of (92)Nb/(92)Mo needs to be at least in the range of 0.06-0.25. PMID- 10849419 TI - The Radio Spectra of SiCCH, SiCN, and SiNC. AB - Three new silicon-bearing radicals of astrophysical interest, SiCCH and the two nearly isoenergetic isomers SiCN and SiNC, were detected in a laboratory discharge in their X2Pi ground states by Fourier transform microwave and millimeter-wave absorption spectroscopy. Hyperfine structure was observed in the low rotational transitions of the (2)Pi(1/2) ladder, and well-resolved Lambda doubling was observed in both fine-structure ladders. With the spectroscopic constants derived from the laboratory measurements, the spectra of all three can be calculated to an uncertainty of less than 0.1 km s(-1) in equivalent radial velocity over the entire range of interest to radio astronomers. SiCN, with a dipole moment of 2.9 D, is probably the most promising of the three for astronomical discovery. PMID- 10849420 TI - The budding yeast homolog of the human EBNA1-binding protein 2 (Ebp2p) is an essential nucleolar protein required for pre-rRNA processing. AB - The human EBP2 protein was found by two-hybrid analysis to interact with the Epstein-Barr virus nuclear antigen 1 (EBNA1). Homologs of human EBP2 can be found in Caenorhabditis elegans, Schizosaccharomyces pombe, and in Saccharomyces cerevisiae, and they all share a conserved 200-300-amino acid block of residues at their C termini. To understand the cellular function of EBP2, we have begun to study the protein in S. cerevisiae. The yeast Ebp2 protein contains N-terminal, nucleolar-associated KKE motifs, and deletion analysis reveals that the C terminal conserved region is required for the activity of the protein. The EBP2 gene codes for an essential protein that localizes to the nucleolus. Temperature sensitive ebp2-1 mutants become depleted of ribosomes and cease to divide after several generations at the restrictive temperature of 36 degrees C. This decline in ribosome levels is accompanied by a diminution in the levels of the 35 S derived recombinant RNAs (rRNAs) (in particular the 25 S and 5.8 S rRNAs). Pulse chase, Northern, and primer extension analysis of the rRNA biosynthetic pathway indicates that ebp2-1 mutants are defective in processing the 27 SA precursor into the 27 SB pre-rRNA. PMID- 10849421 TI - Stress-activated protein kinases (JNK and p38/HOG) are essential for vascular endothelial growth factor mRNA stability. AB - Stability of the vascular endothelial growth factor (VEGF) mRNA is tightly regulated through its 3'-untranslated region (3'-UTR). Here, we demonstrate that VEGF mRNA levels are increased by anisomycin, a strong activator of stress activated protein kinases. Hence, VEGF mRNA induction is inhibited by SB202190, an inhibitor of JNK and p38/HOG kinase. Furthermore, VEGF mRNA expression is increased in cells that overexpress JNK and p38/HOG by an increase in its stability. We show by two different approaches that anisomycin exerts its effect on the VEGF mRNA 3'-UTR. First, by using an in vitro mRNA degradation assay, the half-life of the VEGF mRNA 3'-UTR region transcript was found to be increased when incubated with extracts from anisomycin-treated cells; and second, the 3' UTR was also sufficient to confer mRNA instability to the Nhe3 (Na(+)/H(+) exchanger 3) heterologous reporter gene, and anisomycin treatment stabilized the chimeric mRNA (Nhe3 fused to the VEGF mRNA 3'-UTR). This chimeric mRNA is also more stable in cells overexpressing p38/HOG and JNK that have been stimulated by anisomycin. We show that such regulation is mediated through an AU-rich region of the 3'-UTR contained within a stable hairpin structure. By RNA electrophoretic mobility shift assays, we show that this region binds proteins specifically induced by anisomycin treatment. These findings clearly demonstrate a major role of stress-activated protein kinases in the post-transcriptional regulation of VEGF. PMID- 10849422 TI - Stem cell factor/c-kit up-regulates cyclin D3 and promotes cell cycle progression via the phosphoinositide 3-kinase/p70 S6 kinase pathway in spermatogonia. AB - Stem cell factor (SCF)/c-kit plays an important role in the regulation of hematopoiesis, melanogenesis, and spermatogenesis. In the testis, the SCF/c-kit system is believed to regulate germ cell proliferation, meiosis, and apoptosis. Studies with type A spermatogonia in vivo and in vitro have indicated that SCF induces DNA synthesis and proliferation. However, the signaling pathway for this function of SCF/c-kit has not been elucidated. We now demonstrate that SCF activates phosphoinositide 3-kinase (PI3-K) and p70 S6 kinase (p70S6K) and that rapamycin, a FRAP/mammalian target of rapamycin-dependent inhibitor of p70S6K, completely inhibited bromodeoxyuridine incorporation induced by SCF in primary cultures of spermatogonia. SCF induced cyclin D3 expression and phosphorylation of the retinoblastoma protein through a pathway that is sensitive to both wortmannin and rapamycin. Furthermore, AKT, but not protein kinase C-zeta, is used by SCF/c-kit/PI3-K to activate p70S6K. Dominant negative AKT-K179M completely abolished p70S6K phosphorylation induced by the constitutively active PI3-K catalytic subunit p110. Constitutively active v-AKT highly phosphorylated p70S6K, which was totally inhibited by rapamycin. Thus, SCF/c-kit uses a rapamycin-sensitive PI3-K/AKT/p70S6K/cyclin D3 pathway to promote spermatogonial cell proliferation. PMID- 10849423 TI - Gene-specific silencing by expression of parallel complementary RNA in Escherichia coli. AB - Gene-specific silencing refers to a phenomenon in which expression of an individual gene can be specifically repressed by different mechanisms on the levels of transcription, RNA splicing, transport, degradation in nuclei or cytoplasm, or blocking of translation. In different species gene-specific silencing was observed by expression or injections of antiparallel double stranded RNA formed by a fragment of mRNA and antisense RNA. Here we show a potent and specific gene silencing in bacteria by expression of RNA, that is complementary in a parallel orientation to Escherichia coli lon mRNA. Moreover, the expression of parallel RNA is more effective at producing interference than expression of antisense RNA corresponding to the same mRNA region. Both effects of interference mediated either by parallel RNA or antiparallel RNA gradually decrease up to the 40th generation. Together with in vitro nuclease protection studies these results indicate that a parallel RNA duplex might be formed in vivo and both types of duplexes, antiparallel or parallel, can induce gene-specific silencing by similar mechanisms. PMID- 10849424 TI - Lipid phosphate phosphatase-1 and Ca2+ control lysophosphatidate signaling through EDG-2 receptors. AB - The serum-derived phospholipid growth factor, lysophosphatidate (LPA), activates cells through the EDG family of G protein-coupled receptors. The present study investigated mechanisms by which dephosphorylation of exogenous LPA by lipid phosphate phosphatase-1 (LPP-1) controls cell signaling. Overexpressing LPP-1 decreased the net specific cell association of LPA with Rat2 fibroblasts by approximately 50% at 37 degrees C when less than 10% of LPA was dephosphorylated. This attenuated cell activation as indicated by diminished responses, including cAMP, Ca(2+), activation of phospholipase D and ERK, DNA synthesis, and cell division. Conversely, decreasing LPP-1 expression increased net LPA association, ERK stimulation, and DNA synthesis. Whereas changing LPP-1 expression did not alter the apparent K(d) and B(max) for LPA binding at 4 degrees C, increasing Ca(2+) from 0 to 50 micrometer increased the K(d) from 40 to 900 nm. Decreasing extracellular Ca(2+) from 1.8 mm to 10 micrometer increased LPA binding by 20 fold, shifting the threshold for ERK activation to the nanomolar range. Hence the Ca(2+) dependence of the apparent K(d) values explains the long-standing discrepancy of why micromolar LPA is often needed to activate cells at physiological Ca(2+) levels. In addition, the work demonstrates that LPP-1 can regulate specific LPA association with cells without significantly depleting bulk LPA concentrations in the extracellular medium. This identifies a novel mechanism for controlling EDG-2 receptor activation. PMID- 10849425 TI - Interaction between the transcription factor SPBP and the positive cofactor RNF4. An interplay between protein binding zinc fingers. AB - The activator of stromelysin 1 gene transcription, SPBP, interacts with the RING finger protein RNF4. Both proteins are ubiquitously expressed and localized in the nucleus. RNF4 facilitates accumulation of specific SPBP-DNA complexes in vitro and acts as a positive cofactor in SPBP-mediated transactivation. SPBP harbors an internal zinc finger of the PHD/LAP type. This domain can form intra chain protein-protein contacts in SPBP resulting in negative modulation of SPBP RNF4 interaction. PMID- 10849426 TI - Execution of apoptosis signal-regulating kinase 1 (ASK1)-induced apoptosis by the mitochondria-dependent caspase activation. AB - ASK1 activates JNK and p38 mitogen-activated protein kinases and constitutes a pivotal signaling pathway in cytokine- and stress-induced apoptosis. However, little is known about the mechanism of how ASK1 executes apoptosis. Here we investigated the roles of caspases and mitochondria in ASK1-induced apoptosis. We found that benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD-fmk), a broad spectrum caspase inhibitor, mostly inhibited ASK1-induced cell death, suggesting that caspases are required for ASK1-induced apoptosis. Overexpression of ASK1DeltaN, a constitutively active mutant of ASK1, induced cytochrome c release from mitochondria and activation of caspase-9 and caspase-3 but not of caspase-8 like proteases. Consistently, caspase-8-deficient (Casp8 (-/-)) cells were sensitive to ASK1-induced caspase-3 activation and apoptosis, suggesting that caspase-8 is dispensable for ASK1-induced apoptosis, whereas ASK1 failed to activate caspase-3 in caspase-9-dificient (Casp9 (-/-)) cells. Moreover, mitochondrial cytochrome c release, which was not inhibited by zVAD-fmk, preceded the onset of caspase-3 activation and cell death induced by ASK1. ASK1 thus appears to execute apoptosis mainly by the mitochondria-dependent caspase activation. PMID- 10849427 TI - The small GTPases Ras, Rac, and Cdc42 transcriptionally regulate expression of human fibroblast growth factor 1. AB - Four distinct promoters (1A, 1B, 1C, and 1D) of fibroblast growth factor 1 (FGF1), spaced up to 70 kilobase pairs apart, direct the expression of alternatively spliced transcript variants (FGF1.A, -1. B, -1.C, and -1.D) that encode FGF1. These FGF1 transcripts can be detected in cultured cells as well as in normal and diseased tissues. These transcripts are differentially regulated in a cell-specific manner. To further delineate the biological function of multiple promoter usage by a single gene, we investigated the transcriptional regulation of these promoters by defined signaling pathways associated with cell proliferation and cell survival. Here we show a specific association of two of the FGF1 promoters, 1C and 1D, with signaling cascades of the Ras superfamily of GTPases. A serum-response element, comprised of the Ets and CArG motifs, present in promoter 1D was shown to be the target of distinct signaling cascades; the Ets motif target of Ras, Rac1, and Cdc42 regulation; and the CArG motif target of de novo protein synthesis-independent cascade. Ras and Rac1 also activated the FGF2 promoter. Further, the transcription factor Ets2 synergistically activated FGF1 gene, but not FGF2, in a Ras- and Rac1-dependent signaling pathway. In support of these conclusions high levels of intracellular FGF1 were detected in cells undergoing cytokinesis. Altogether, our results suggest that FGF1 may play a fundamental role in cell division, spreading, and migration, in addition to cell proliferation. PMID- 10849428 TI - The docking molecule gab2 is induced by lymphocyte activation and is involved in signaling by interleukin-2 and interleukin-15 but not other common gamma chain using cytokines. AB - Interleukin (IL)-2, a critical cytokine with indispensable functions in regulating lymphoid homeostasis, induces the activation of several biochemical pathways. Precisely how these pathways are linked and how they relate to the biological action of IL-2 is incompletely understood. We previously identified SHP-2 (Src homology 2 domain containing phosphatase 2) as an important intermediate in IL-2-dependent MAPK activation and showed its association with a 98-kDa phosphoprotein in response to IL-2. Here, we demonstrate that Gab2, a recently identified adapter molecule, is the major SHP-2 and phosphatidylinositol 3'-kinase-associated 98-kDa protein in normal, IL-2-activated lymphocytes. We further demonstrate that phosphorylation of both Gab2 and SHP-2 is largely dependent upon tyrosine 338 of the IL-2 receptor beta chain. Gab2 can be a substrate of all the three major classes of non-receptor tyrosine kinases associated with the IL-2R, but in terms of IL-2 signaling, JAK3 but not Lck or Syk is essential for Gab2 phosphorylation. We also demonstrate that only IL-2 and IL-15, but not other gammac cytokines induce Gab2 phosphorylation; the ability to phosphorylate Gab2 correlates with Shc phosphorylation and ERK1/ERK2 activation. Finally, we also show that Gab2 levels are regulated by T cell activation, and resting T cells express little Gab2. Therefore, up-regulation and activation of Gab2 may be important in linking the IL-2 receptor to activation of MAPK and may be an important means of achieving specificity in cytokine signaling. PMID- 10849429 TI - Do sequence repeats play an equivalent role in the choline-binding module of pneumococcal LytA amidase? AB - LytA amidase breaks down the N-acetylmuramoyl-l-alanine bonds in the peptidoglycan backbone of Streptococcus pneumoniae. Its polypeptide chain has two modules: the NH(2)-terminal module, responsible for the catalytic activity, and the COOH-terminal module, constructed by six tandem repeats of 20 or 21 amino acids (p1-p6) and a short COOH-terminal tail. The polypeptide chain must contain at least four repeats to efficiently anchor the autolysin to the choline residues of the cell wall. Nevertheless, the catalytic efficiency decreases by 90% upon deletion of the final tail. The structural implications of deleting step by step the two last (p5 and p6) repeats and the final COOH-tail and their effects on choline-amidase interactions have been examined by comparing four truncated mutants with LytA amidase by means of different techniques. Removal of this region has minor effects on secondary structure content but significantly affects the stability of native conformations. The last 11 amino acids and the p5 repeat stabilize the COOH-terminal module; each increases the module transition temperature by about 6 degrees C. Moreover, the p5 motif also seems to participate, in a choline-dependent way, in the stabilization of the NH(2) terminal module. The effects of choline binding on the thermal stability profile of the mutant lacking the p5 repeat might reflect a cooperative pathway providing molecular communication between the choline-binding module and the NH(2)-terminal region. The three sequence motives favor the choline-amidase interaction, but the tail is an essential factor in the monomer <--> dimer self-association equilibrium of LytA and its regulation by choline. The final tail is required for preferential interaction of choline with LytA dimers and for the existence of different sets of choline-binding sites. The p6 repeat scarcely affects the amidase stability but could provide the proper three-dimensional orientation of the final tail. PMID- 10849430 TI - Differential regulation of rat aquaporin-5 promoter/enhancer activities in lung and salivary epithelial cells. AB - Aquaporin-5 (AQP5) is a water channel protein that is selectively expressed in respiratory, salivary, and lacrimal tissues. In order to establish the tissue specific transcriptional programs that underlie its lung- and salivary-specific expression, a 4.5-kilobase pair DNA fragment encompassing the 5'-flanking region of the rat AQP5 gene has been characterized in detail. A major transcription start site utilized in lung and salivary glands has been localized downstream of a TATAA-like motif. Transient transfection assays of -4.3- and -1.7-AQP5 luciferase constructs in AQP5-expressing lung (MLE-15) and salivary (Pa-4) cells and nonexpressing fibroblast (NIH3T3) and epithelial (HeLa) cells demonstrate preferential transcriptional enhancement of reporter activities in MLE-15 and Pa 4 cells. Transient transfection assays of a series of 5' --> 3' deletion constructs of -4.3-AQP5-luciferase suggest that a common salivary and lung enhancer is located between nucleotides -274 and -139, and a lung-specific enhancer is located between nucleotides -894 and -710. There is one putative lung specific repressor located in the region of nucleotides -1003/-894 and a common lung and salivary repressor located at nucleotides -503/-385. Moreover, 3' --> 5' deletions up to -171 and -127 base pairs almost abolish transcriptional activation in salivary and lung cells, respectively. Together, our findings indicate that the combination of enhancer/repressor elements within the proximal 5'-flanking region of rat AQP5 gene dictates its restricted expression in both lung and salivary cells. PMID- 10849431 TI - Activation of platelet-transforming growth factor beta-1 in the absence of thrombospondin-1. AB - Thrombospondin-1 (TSP-1) has been shown to bind and activate transforming growth factor-beta1 (TGF-beta1). This observation raises the possibility that TSP-1 helps to sequester TGF-beta1 in platelet alpha granules and activates TGF-beta1 once both proteins are secreted. Herein, we evaluated the level of active and latent TGF-beta1 in the plasma and in the supernatant of thrombin-treated platelets from TSP-1 null and wild-type mice on two genetic backgrounds (C57BL/6 and 129Sv). The plasminogen activator inhibitor-1/luciferase bioassay and an immunological assay were used to determine active and latent TGF-beta1. No significant differences were observed in the levels of active and latent TGF beta1 in the supernatant of thrombin-treated platelets from TSP-1 null and wild type mice. Active and latent TGF-beta1 were significantly increased in the plasma and platelets of C57BL/6 mice as compared with 129Sv mice. In addition, there was an increase of plasma level of latent TGF-beta1 in TSP-1 null mice as compared with wild-type mice on the C57BL/6 background but not on the 129Sv background. No active TGF-beta1 was observed in the plasma of either TSP-1 null and wild-type mice. These data indicate that TSP-1 does not function as a chaperon for TGF beta1 during platelet production and does not activate significant quantities of secreted TGF-beta1 despite a vast excess in the number of TSP-1 molecules as compared with TGF-beta1 molecules. Because platelet releasates from TSP-1 null mice contain active TGF-beta1, we suggest that other important mechanisms of physiological activation of TGF-beta1 probably exist in platelets. PMID- 10849432 TI - Bone morphogenetic protein-9. An autocrine/paracrine cytokine in the liver. AB - Bone morphogenetic proteins (BMPs) occupy important roles during development serving to direct cells through specific differentiation programs. While several BMPs are essential for embryonic viability, their significance in mediating intercellular communication in the context of adult organ systems remains largely unknown. In the adult rat we characterized the tissue- and cell-specific transcription and translation of BMP-9. Utilizing a ribonuclease protection assay, we determined that in the adult animal, BMP-9 expression occurs predominantly in the liver. Furthermore, we determined that the non-parenchymal cells of the liver, i.e. endothelial, Kupffer, and stellate cells, are the major sources of this message. Western analyses corroborate the ribonuclease protection assay results, confirming that LEC and KC contain an abundance of immunoreactive BMP-9. Using [(125)I]BMP-9, a receptor with specific binding affinity for BMP-9 was characterized in primary cultures of hepatic endothelial cells and Kupffer cells. BMP-9 binding to these cell types was observed to be fully reversible and highly specific for this ligand. Additionally, we demonstrate that BMP-9 is specifically internalized upon binding to its receptor. This may represent a novel BMP receptor and is the first to be characterized in primary cultures of mature liver non-parenchymal cells. Our results depict BMP-9 as a potential autocrine/paracrine mediator in the hepatic reticuloendothelial system. PMID- 10849433 TI - Sustained endothelial nitric-oxide synthase activation requires capacitative Ca2+ entry. AB - Endothelial nitric-oxide synthase (eNOS), a Ca(2+)/calmodulin-dependent enzyme, is critical for vascular homeostasis. While eNOS is membrane-associated through its N-myristoylation, the significance of membrane association in locating eNOS near sources of Ca(2+) entry is uncertain. To assess the Ca(2+) source required for eNOS activation, chimera containing the full-length eNOS cDNA and HA-tagged aequorin sequence (EHA), and MHA (myristoylation-deficient EHA) were generated and transfected into COS-7 cells. The EHA chimera was primarily targeted to the plasma membrane while MHA was located intracellularly. Both constructs retained enzymatic eNOS activity and aequorin-mediated Ca(2+) sensitivity. The plasma membrane-associated EHA and intracellular MHA were compared in their ability to sense changes in local Ca(2+) concentration, demonstrating preferential sensitivity to Ca(2+) originating from intracellular pools (MHA) or from capacitative Ca(2+) entry (EHA). Measurements of eNOS activation in intact cells revealed that the eNOS enzymatic activity of EHA was more sensitive to Ca(2+) influx via capacitative Ca(2+) entry than intracellular release, whereas MHA eNOS activity was more responsive to intracellular Ca(2+) release. When eNOS activation by CCE was compared with that generated by an equal rise in [Ca(2+)](i) due to the Ca(2+) ionophore ionomycin, a 10-fold greater increase in NO production was found in the former condition. These results demonstrate that EHA and MHA chimera are properly targeted and retain full functions of eNOS and aequorin, and that capacitative Ca(2+) influx is the principle stimulus for sustained activation of eNOS on the plasma membrane in intact cells. PMID- 10849434 TI - Inhibition of DNA methyltransferase inhibits DNA replication. AB - Ectopic expression of DNA methyltransferase transforms vertebrate cells, and inhibition of DNA methyltransferase reverses the transformed phenotype by an unknown mechanism. We tested the hypothesis that the presence of an active DNA methyltransferase is required for DNA replication in human non-small cell lung carcinoma A549 cells. We show that the inhibition of DNA methyltransferase by two novel mechanisms negatively affects DNA synthesis and progression through the cell cycle. Competitive polymerase chain reaction of newly synthesized DNA shows decreased origin activity at three previously characterized origins of replication following DNA methyltransferase inhibition. We suggest that the requirement of an active DNA methyltransferase for the functioning of the replication machinery has evolved to coordinate DNA replication and inheritance of the DNA methylation pattern. PMID- 10849435 TI - Characterization of the effects of mutations in the putative branchpoint sequence of intron 4 on the splicing within the human lecithin:cholesterol acyltransferase gene. AB - We have previously identified a point mutation (intervening sequence (IVS) 4: T - > C) in the branchpoint consensus sequence of intron 4 of the lecithin:cholesterol acyltransferase (LCAT) gene in patients with fish-eye disease. To investigate the possible mechanisms responsible for the defective splicing, we made a series of mutations in the branchpoint sequence and expressed these mutants in HEK-293 cells followed by the analysis of pre-mRNA splicing using reverse transcriptase-polymerase chain reaction as well as LCAT activity assay. The results reveal that 1) the mutation of the branchpoint adenosine to any other nucleotide completely abolishes splicing; 2) the insertion of a normal branch site into the intronic sequence of the natural (IVS4-22c) or the branchpoint (IVS4-20t) mutant completely restores splicing; 3) the natural mutation can be partially rescued by making a single nucleotide change (G --> A) within the branchpoint consensus sequence; and 4) other single base changes, particularly around the branchpoint adenosine residue, significantly decrease the efficiency of splicing and thus enzyme activity. Surprisingly, the nucleotide transversion at the last position of the branchpoint sequence (i.e. IVS4-25a or 25g) results in a 2.7-fold increase in splicing efficiency. Therefore, these observations clearly establish the functional significance of the branchpoint sequence of intron 4 for the splicing of the human LCAT mRNA precursors. PMID- 10849436 TI - A1 functions at the mitochondria to delay endothelial apoptosis in response to tumor necrosis factor. AB - Tumor necrosis factor (TNF) does not cause endothelial apoptosis unless the expression of cytoprotective genes is blocked. We have previously demonstrated that one of the TNF-inducible cytoprotective genes is the Bcl-2 family member, A1. A1 is induced by the action of the transcription factor, NFkappaB, in response to inflammatory mediators. In this report we demonstrate that, as with other cell types, inhibition of NFkappaB initiates microvascular endothelial apoptosis in response to TNF. A1 is able to inhibit this apoptosis over 24 h. We demonstrate that A1 is localized to and functions at the mitochondria. Whereas A1 is able to inhibit mitochondrial depolarization, loss of cytochrome c, cleavage of caspase 9, BID, and poly(ADP-ribose) polymerase, it does not block caspase 8 or caspase 3 cleavage. In contrast, A1 is not able to prevent endothelial apoptosis by TNF over 72 h, when NFkappaB signaling is blocked. On the other hand, the caspase inhibitor, benzyloxycarbonyl-VAD-formylmethyl ketone, completely blocks TNF-induced endothelial apoptosis over 72 h. Our findings indicate that A1 is able to maintain temporary survival of endothelial cells in response to TNF by maintaining mitochondrial viability and function. However, a mitochondria-independent caspase pathway eventually results in endothelial death despite mitochondrial protection by A1. PMID- 10849437 TI - Essential role of selenium in the catalytic activities of mammalian thioredoxin reductase revealed by characterization of recombinant enzymes with selenocysteine mutations. AB - Mammalian thioredoxin reductases (TrxR) are dimers homologous to glutathione reductase with a selenocysteine (SeCys) residue in the conserved C-terminal sequence -Gly-Cys-SeCys-Gly. We removed the selenocysteine insertion sequence in the rat gene, and we changed the SeCys(498) encoded by TGA to Cys or Ser by mutagenesis. The truncated protein having the C-terminal SeCys-Gly dipeptide deleted, expected in selenium deficiency, was also engineered. All three mutant enzymes were overexpressed in Escherichia coli and purified to homogeneity with 1 mol of FAD per monomeric subunit. Anaerobic titrations with NADPH rapidly generated the A(540 nm) absorbance resulting from the thiolate-flavin charge transfer complex characteristic of mammalian TrxR. However, only the SeCys(498) - > Cys enzyme showed catalytic activity in reduction of thioredoxin, with a 100 fold lower k(cat) and a 10-fold lower K(m) compared with the wild type rat enzyme. The pH optimum of the SeCys(498) --> Cys mutant enzyme was 9 as opposed to 7 for the wild type TrxR, strongly suggesting involvement of the low pK(a) SeCys selenol in the enzyme mechanism. Whereas H(2)O(2) was a substrate for the wild type enzyme, all mutant enzymes lacked hydroperoxidase activity. Thus selenium is required for the catalytic activities of TrxR explaining the essential role of this trace element in cell growth. PMID- 10849438 TI - Tyrosine-phosphorylated Vav1 as a point of integration for T-cell receptor- and CD28-mediated activation of JNK, p38, and interleukin-2 transcription. AB - In this study we identified tyrosine-phosphorylated Vav1 as an early point of integration between the signaling routes triggered by the T-cell receptor and CD28 in human T-cell leukemia cells. Costimulation resulted in a prolonged and sustained phosphorylation and membrane localization of Vav1 in comparison to T cell receptor activation alone. T-cell stimulation induced the recruitment of Vav1 to an inducible multiprotein T-cell activation signaling complex at the plasma membrane. Vav1 activated the mitogen-activated protein kinases JNK and p38. The Vav1-mediated activation of JNK employed a pathway involving Rac, HPK1, MLK3, and MKK7. The costimulation-induced activation of p38 was inhibited by dominant negative forms of Vav1, Rac, and MKK6. Here we show that Vav1 also induces transcription factors that bind to the CD28RE/AP element contained in the interleukin-2 promoter. A detailed mutational analysis of Vav1 revealed a series of constitutively active and nonfunctional forms of Vav1. Almost all inactive versions were mutated in their Dbl homology domain and behaved as dominant negative mutants that impaired costimulation-induced activation of JNK, p38, and CD28RE/AP-dependent transcription. In contrast to NF-AT-dependent transcription, Vav1-mediated transcriptional induction of the CD28RE/AP element in the interleukin-2 promoter could only partially be inhibited by cyclosporin A, suggesting a dual role of Vav1 for controlling Ca(2+)-dependent and -independent events. PMID- 10849439 TI - p38-dependent enhancement of cytokine-induced nitric-oxide synthase gene expression by heat shock protein 70. AB - Heat shock protein (hsp) 70 protects cells against stress by means of its ability to chaperone denatured proteins and to modulate stress-activated signaling pathways. Because inflammatory processes are often accompanied by hsp expression and because stress and cytokines share several signaling pathways, we investigated the possibility that hsp70 might modulate the cellular response to cytokines. We found that stable cell clones overexpressing hsp70, or cells shortly after transfection with hsp70, produced 2 times more nitric oxide and inducible nitric-oxide synthase (iNOS) protein and mRNA in response to cytokines than control cells expressing undetectable amounts of hsp70. Since mitogen activated protein kinases participate in the activation of iNOS by cytokines, we investigated whether hsp70 affected the activation of these signaling pathways. hsp70 overexpression led to a specific enhancement of the activation of the p38 pathway by cytokines, producing little or no effect on the activation of extracellular signal-regulated kinase or Jun N-terminal kinase. Blocking p38 activity with SB203580 totally abolished the enhancing effect of hsp70 on cytokine-induced endogenous iNOS mRNA accumulation or transcription of an iNOS promoter-driven luciferase gene, while having little effect on the cytokine response observed in control cells. We conclude that the p38 pathway acts as an enhancing factor in the activation of iNOS by cytokines and that hsp70 can modulate the cellular response to cytokines by acting on signaling elements upstream of p38. PMID- 10849440 TI - Interaction of TAFII105 with selected p65/RelA dimers is associated with activation of subset of NF-kappa B genes. AB - TAF(II)105, a substoichiometric coactivator subunit of TFIID, is important for activation of anti-apoptotic genes by NF-kappaB in response to the cytokine tumor necrosis factor (TNF)-alpha. In the present study we have analyzed the mechanism of TAF(II)105 function with respect to its regulation of p65/RelA, a component of NF-kappaB. We found two independent p65/RelA-binding domains within the N terminus of TAF(II)105. One of these domains appears to be crucial for TAF(II)105 mediated anti-apoptotic gene activation in response to TNF-alpha. Analysis of the interaction between TAF(II)105 and different NF-kappaB complexes has revealed substantial differences in the affinity of TAF(II)105 toward different p65/RelA containing dimers. We have identified the TNF-alpha induced anti-apoptotic A20 gene as a target gene of TAF(II)105. A20 has a differential protective effect on cell death induced by TNF-alpha in the presence of either the dominant negative mutant of TAF(II)105 (TAF(II)105DeltaC) or the superdominant IkappaBalpha. The results suggest that the inhibitory effect of TAF(II)105DeltaC on NF-kappaB dependent genes is restricted to a subset of anti-apoptotic genes while the effect of IkappaBalpha is more general. Thus, an interaction between NF-kappaB and a specific coactivator is important for specifying target gene activation. PMID- 10849441 TI - Role of peroxiredoxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. AB - Peroxiredoxins (Prxs) play an important role in regulating cellular differentiation and proliferation in several types of mammalian cells. One mechanism for this action involves modulation of hydrogen peroxide (H(2)O(2)) mediated cellular responses. This report examines the expression of Prx I and Prx II in thyroid cells and their roles in eliminating H(2)O(2) produced in response to thyrotropin (TSH). Prx I and Prx II are constitutively expressed in FRTL-5 thyroid cells. Prx I expression, but not Prx II expression, is stimulated by exposure to TSH and H(2)O(2). In addition, methimazole induces a high level of Prx I mRNA and protein in these cells. Overexpression of Prx I and Prx II enhances the elimination of H(2)O(2) produced by TSH in FRTL-5 cells. Treatment with 500 micrometer H(2)O(2) causes apoptosis in FRTL-5 cells as evidenced by standard assays of apoptosis (i.e. terminal deoxynucleotidyl transferase deoxyuridine triphosphate-biotin nick end labeling, BAX expression, and poly(ADP ribose) polymerase cleavage. Overexpression of Prx I and Prx II reduces the amount of H(2)O(2)-induced apoptosis measured by these assays. These results suggest that Prx I and Prx II are involved in the removal of H(2)O(2) in thyroid cells and can protect these cells from undergoing apoptosis. These proteins are likely to be involved in the normal physiological response to TSH-induced production of H(2)O(2) in thyroid cells. PMID- 10849442 TI - Do structural deviations between toxins adopting the same fold reflect functional differences? AB - Three-finger proteins form a structurally related family of compounds that exhibit a great variety of biological properties. To address the question of the prediction of functional areas on their surfaces, we tentatively conferred the acetylcholinesterase inhibitory activity of fasciculins on a short-chain curaremimetic toxin. For this purpose, we assimilated the three-dimensional structure of fasciculin 2 with the one of toxin alpha. This comparison revealed that the tips of the first and second loops, together with the C terminus residue, deviated most. A first recombinant fasciculin/toxin alpha chimera was designed by transferring loop 1 in its entirety together with the tip of loop 2 of fasciculin 2 into the toxin alpha scaffold. A second chimera (rChII) was obtained by adding the point Asn-61 --> Tyr substitution. Comparison of functional and structural properties of both chimeras show that rChII can accommodate the imposed modifications and displays nearly all the acetylcholinesterase-blocking activities of fasciculins. The three-dimensional structure of rChII demonstrates that rChII adopts a typical three-fingered fold with structural features of both parent toxins. Taken together, these results emphasize the great structural flexibility and functional adaptability of that fold and confirm that structural deviations between fasciculins and short-chain neurotoxins do indeed reflect functional diversity. PMID- 10849443 TI - Transcription factors ets1, NF-kappa B, and Sp1 are major determinants of the promoter activity of the human protein kinase CK2alpha gene. AB - CK2alpha is one of two isoforms of protein kinase CK2, a highly conserved, ubiquitous, and vital phosphotransferase whose expression is kept at constant cellular levels and whose dysregulated expression has been linked to malignant diseases. The upstream sequence of the gene coding for human CK2alpha (CSNK1A1, chromosomal location 20p13) has been examined for promoter location and transcription factor interactions using reporter gene assays (luciferase; HeLa cells), site-directed mutagenesis, electrophoretic mobility shift assays, super shifts, UV cross-linking, Western blotting, and DNA affinity chromatography. Highest promoter activity has been found in a region comprising positions -9 to 46. Factors Sp1, Ets-1, and NF-kappaB have been identified as interaction partners and, by mutation of individual sites and simultaneous mutations of two or more sites, shown to cross-talk to each other. At least two of the factors (Sp1; NF-kappaB) were susceptible to phosphorylation by CK2 holoenzyme, a tetramer composed of two CK2alpha and two regulatory CK2beta proteins, but not by individual CK2alpha. Because the phosphorylation decreases promoter binding and repeated immunoprecipitation reveals presence of "free" CK2beta in cell extracts, it is tempting to speculate that the gene product CK2alpha might readily form CK2 holoenzyme and feed back onto gene transcription. The data represent the first promoter control analysis of a mammalian CK2alpha gene and provide a hypothesis of how the constant expression level of CK2alpha may be achieved. PMID- 10849444 TI - Proteasomes regulate the duration of erythropoietin receptor activation by controlling down-regulation of cell surface receptors. AB - The binding of erythropoietin (Epo) to its receptor leads to the transient phosphorylation of the Epo receptor (EpoR) and the activation of intracellular signaling pathways. Inactivation mechanisms are simultaneously turned on, and Epo induced signaling pathways return to nearly basal levels after 30-60 min of stimulation. We show that proteasomes control these inactivation mechanisms. In cells treated with the proteasome inhibitors N-Ac-Leu-Leu-norleucinal (LLnL) or lactacystin, EpoR tyrosine phosphorylation and activation of intracellular signaling pathways (Jak2, STAT5, phosphatidylinositol 3-kinase) were sustained for at least 2 h. We show that this effect was due to the continuous replenishment of the cell surface pool of EpoRs in cells treated with proteasome inhibitors. Proteasome inhibitors did not modify the internalization and degradation of Epo.EpoR complexes, but they allowed the continuous replacement of the internalized receptors by newly synthesized receptors. Proteasome inhibitors did not modify the synthesis of EpoRs, but they allowed their transport to the cell surface. N-Ac-Leu-Leu-norleucinal, but not lactacystin, also inhibited the degradation of internalized Epo.EpoR complexes, most probably through cathepsin inhibition. The internalized EpoRs were not tyrosine-phosphorylated, and they did not activate intracellular signaling pathways. Our results show that the proteasome controls the down-regulation of EpoRs in Epo-stimulated cells by inhibiting the cell surface replacement of internalized EpoRs. PMID- 10849445 TI - Base stacking and even/odd behavior of hairpin loops in DNA triplet repeat slippage and expansion with DNA polymerase. AB - Repetitions of CAG or CTG triplets in DNA can form intrastrand hairpin loops with combinations of normal and mismatched base pairs that easily rearrange. Such loops may promote primer-template slippage in DNA replication or repair to give triplet-repeat expansions like those associated with neurodegenerative diseases. Using self-priming sequences (e.g. (CAG)(16)(CTG)(4)), we resolve all hairpin loops formed and measure their slippage and expansion rates with DNA polymerase at 37 degrees C. Comparing CAG/CTG loop structures with GAC/GTC structures, having similar hydrogen bonding but different base stacking, we find that CAG, CTG, and GTC triplets predominantly form even-membered loops that slip in steps of two triplets, whereas GAC triplets favor odd-numbered loops. Slippage rates decline as hairpin stability increases, supporting the idea that slippage initiates more easily in less stable regions. Loop stabilities (in low salt) increase in the order GTC < CAG < GAC < CTG, while slippage rates decrease in the order GTC > CAG approximately GAC > CTG. Loops of GTC compared with CTG melt 9 degrees C lower and slip 6-fold faster. We interpret results in terms of base stacking, by relating melting temperature to standard enthalpy changes for doublets of base pairs and mispairs, considering enthalpy-entropy compensation. PMID- 10849446 TI - Big mitogen-activated kinase regulates multiple members of the MEF2 protein family. AB - Big mitogen-activated protein (MAP) kinase (BMK1), a member of the mammalian MAP kinase family, is activated by growth factors. The activation of BMK1 is required for growth factor-induced cell proliferation and cell cycle progression. We have previously shown that BMK1 regulates c-jun gene expression through direct phosphorylation and activation of transcription factor MEF2C. MEF2C belongs to the myocyte enhancer factor 2 (MEF2) protein family, a four-membered family of transcription factors denoted MEF2A, -2B, -2C, and -2D. Here, we demonstrate that, in addition to MEF2C, BMK1 phosphorylates and activates MEF2A and MEF2D but not MEF2B. The blocking of BMK1 signaling inhibits the epidermal growth factor dependent activation of these three MEF2 transcription factors. The sites phosphorylated by activated BMK1 were mapped to Ser-355, Thr-312, and Thr-319 of MEF2A and Ser-179 of MEF2D both in vitro and in vivo. Site-directed mutagenesis reveals that the phosphorylation of these sites in MEF2A and MEF2D are necessary for the induction of MEF2A and 2D transactivating activity by either BMK1 or by epidermal growth factor. Taken together, these data demonstrate that, upon growth factor induction, BMK1 directly phosphorylates and activates three members of the MEF2 family of transcription factors thereby inducing MEF2-dependent gene expression. PMID- 10849447 TI - ADAM 12, a disintegrin metalloprotease, interacts with insulin-like growth factor binding protein-3. AB - Insulin-like growth factor-binding protein (IGFBP)-3 binds the insulin-like growth factors with high affinity and modulates their actions. Proteolytic cleavage of IGFBP-3 may regulate insulin-like growth factor bioavailability. IGFBP-3 is extensively degraded in serum during pregnancy; however, as yet the pregnancy-specific protease, or proteases, have not been identified. We utilized a yeast two-hybrid assay and a human placental cDNA library to investigate IGFBP 3-interacting proteins. A disintegrin and metalloprotease-12 (ADAM 12), a member of a family of metalloprotease disintegrins that is highly expressed in placental tissue, was identified as interacting with IGFBP-3. This interaction involved the cysteine-rich domain of ADAM 12. Unlike other members of this family of disintegrin metalloproteases that are membrane proteins, ADAM 12 exists as an alternatively spliced soluble secreted protein. To verify the interaction between ADAM 12 and IGFBP-3, an expression construct containing an ADAM 12-S cDNA was transfected into COS-1 cells. Co-precipitation was observed when conditioned medium was analyzed by immunoprecipitation with an antibody against either ADAM 12 or IGFBP-3 followed by Western blotting with anti-IGFBP-3 or anti-ADAM 12. Although minimal proteolysis of IGFBP-3 was observed in conditioned medium from control cells, this was increased approximately 4-fold in conditioned medium from ADAM 12-S-transfected cells. Recombinant ADAM 12-S partially purified from conditioned medium on a heparin-Sepharose column also proteolyzed IGFBP-3. The degradation pattern was similar to that seen with pregnancy serum, and the presence of ADAM 12-S in serum during pregnancy was confirmed. The data suggest that ADAM 12-S has IGFBP-3 protease activity, and it may contribute to the IGFBP 3 protease activity present in pregnancy serum. PMID- 10849448 TI - Transformation of myeloid leukemia cells to cytokine independence by Bcr-Abl is suppressed by kinase-defective Hck. AB - Bcr-Abl is the constitutively active protein-tyrosine kinase expressed as a result of the Philadelphia translocation in chronic myelogenous leukemia. Bcr-Abl is coupled to many of the same signaling pathways normally regulated by hematopoietic cytokines. Recent work shows that Hck, a member of the Src tyrosine kinase family with myeloid-restricted expression, associates with and is activated by Bcr-Abl. Here we investigated the mechanism of Hck interaction with Bcr-Abl and the requirement for Hck activation in Bcr-Abl transformation signaling. Binding studies demonstrated that the Hck SH3 and SH2 domains are sufficient for interaction with Bcr-Abl in vitro. Hck binding localizes to the Abl SH2, SH3, and kinase domains as well as the distal portion of the C-terminal tail. To address the requirement for endogenous Src family kinase activation in Bcr-Abl signaling, a kinase-defective mutant of Hck was stably expressed in the cytokine-dependent myeloid leukemia cell line DAGM. Kinase-defective Hck dramatically suppressed Bcr-Abl-induced outgrowth of these cells in the absence of cytokine compared with a control cell line expressing beta-galactosidase. In contrast, kinase-defective Hck did not affect cell proliferation in response to interleukin-3, suggesting that the effect is specific for Bcr-Abl. These data show that Hck interacts with Bcr-Abl through a complex mechanism involving kinase dependent and -independent components and that interaction with Hck or other Src family members is essential for transformation signaling by Bcr-Abl. PMID- 10849449 TI - The unexamined death is not worth dying. PMID- 10849450 TI - Psychiatric evaluation of death-hastening requests. Lessons from dialysis discontinuation. AB - The authors aim to facilitate the psychiatric evaluation of death-hastening decisions, such as cessation of life-support treatment or physician-assisted suicide, by deriving principles for evaluating patients from a literature review and a recently completed prospective study on dialysis discontinuation conducted by consultation psychiatrists. Factors are delineated and suggestions are provided for the evaluation of requests to accelerate dying. Included are the authors' method for determining major depression in the context of terminal illness and their "vector analysis" in assessing patient requests to stop dialysis. As our society heatedly examines the care provided to the terminally ill, psychiatry also needs to reconsider whether actions that foreshorten life can be normative and permissible. Familiarity with competency, psychiatric diagnosis, and ease in communication and negotiation between patient, family, and staff are resources that psychiatrists can bring to these complicated assessments. Challenging areas include diagnosing depression, establishing the adequacy of palliative care, and appreciating issues related to personality features, family dynamics, and ethnic differences. PMID- 10849451 TI - Quo vadis, psychiatry? Problems and potential for the future of medical student education in psychiatry. AB - Psychiatric education occupies a relatively low status level in most academic departments of psychiatry. This problem may be in part because career teachers rarely generate sustained external grant support. Their salaries may be regarded as perpetual drains on the budgets of department chairs. The author explores a possible relationship between the decline in the emphasis on psychiatric education and the decline in the number of medical students entering psychiatric residency programs. Recommendations for improvement in the support of psychiatric education are made that focus on greater accountability of how tuition fees are distributed to support the salaries of faculty educators. The unique role that consultation-liaison (C-L) psychiatry occupies in psychiatric education, the effects of managed care on C-L psychiatry, and the special problems that C-L psychiatry currently faces with its strong emphasis on non-reimbursable educational activities are discussed. Revitalization of the importance of education in undergraduate medical education is vital for the future of psychiatry. PMID- 10849452 TI - Repetitive skin-picking in a student population and comparison with a sample of self-injurious skin-pickers. AB - The prevalence of skin-picking and its associated characteristics were documented in a nonclinical sample of 105 college students. Subjects completed a self-report skin-picking inventory and several paper-and-pencil scales. Students who endorsed skin-picking were compared to a clinical sample of self-injurious skin-pickers (n = 31) reported on previously. Of the student subjects, 78.1% (n = 82) endorsed some degree of skin-picking and four subjects satisfied criteria for severe, self injurious picking. Student subjects significantly differed from the clinical sample-of self-injurious skin-pickers in the duration, focus, and extent of picking, techniques used, reasons for picking, associated emotions, and picking sequelae. PMID- 10849453 TI - Pain location and psychological characteristics of patients with chronic pain. AB - The authors assessed psychological characteristics of 140 medical outpatients with chronic non-malignant pain referred for psychiatric consultation. Subjects completed the Toronto Alexithymia Scale, Somatosensory Amplification Scale (SSAS), and Counter-dependency Scale (CDS). The only psychological measure able to differentiate the chronic pain group from the control subjects was the CDS. However, SSAS scores were significantly higher in subjects having pain involving the head, chest, abdomen, or pelvis than in subjects having pain only in their backs or extremities. The latter subgroup had significantly higher CDS scores. The findings suggest that there are discrete subgroups within the chronic pain population defined by pain location and specific psychological characteristics. PMID- 10849454 TI - Stress and other psychosocial characteristics of patients with psychogenic nonepileptic seizures. AB - Research on psychogenic nonepileptic seizures (PNES) has focused on childhood abuse, but less is known about other stressors and psychosocial risk factors. The authors compared 25 patients with PNES with 33 control subjects with epilepsy on stressful life events and other risk factors for somatoform disorders. Compared with control subjects, patients with PNES reported significantly more prevalent and stressful negative life events (including adulthood abuse) and more current rumination, stress-related diseases, somatic symptoms, bodily awareness, and marginally more anxiety and depression. However, the relationship of many of these variables to PNES was accounted for by life stress. Groups did not differ on illness worry, alexithymia, or psychotic symptoms. The results suggest that PNES are part of a larger pattern of somatic symptoms responses to a wide range of negative events, including stress in adulthood. PMID- 10849455 TI - Chronic pain disorder following physical injury. AB - Pain disorders that are primarily associated with psychological factors are of great clinical concern, but they are difficult to study because of the inability to make valid or reliable diagnoses by structured interview alone. The authors confront this difficulty by using an injured subject population that had extensive psychiatric and medical evaluations. Those who developed somatoform pain disorder (SPD) were compared with a control group who did not. The SPD group had distinctive associated factors: more sites of pain, spread of pain beyond area of original injury, and substantially more opiate and benzodiazepine use. Compensation/litigation influenced symptoms more in the SPD group. Psychotherapists often supported the patient's viewpoint that the pain was physical and to be endured. PMID- 10849456 TI - Multidimensional investigation of the structure of coping among people with amputations. AB - In this study, the responses of 61 people with amputations to a measure of coping strategies were submitted to multidimensional scaling and cluster analysis. Interpretations of the three-dimensional solution, aided by the emergence of five coping clusters, suggested that respondents' perceptions of their coping with amputation-related stress were best explained by the following three dimensions: 1) active/confrontive versus passive/avoidance coping; 2) pessimistic/fatalistic versus optimistic/positivistic coping: and 3) social/emotional versus cognitive coping. PMID- 10849457 TI - Referrals to psychiatrists. Assessing the communication interface between psychiatry and primary care. AB - The Study of Outpatient Referral Patterns was conducted in 1998 to examine the nature of the communication relationship between psychiatrists and primary care physicians regarding outpatient referrals. Nationally representative psychiatrists were surveyed (N = 542) regarding their aggregate experience with outpatient referrals from non-psychiatric physicians in the previous 60 days. Data regarding frequency and type of information and mode of communication were gathered. Results indicate that primary care physicians represent a significant source of referrals to psychiatrists and that psychiatrists are generally satisfied with the communication interface with the referring physicians. Psychiatrists' level of satisfaction was related to the quantity and quality of information provided by referring physicians. PMID- 10849458 TI - A nine-year follow-up of people diagnosed with multiple chemical sensitivities. AB - The authors assessed self-reported health status and clinical symptoms in people reporting multiple chemical sensitivities (MCS) at a 9-year follow-up interview using structured and semistructured instruments and self-report questionnaires. Of the original sample, 18 people (69%) consented to an interview. By use of the best estimate diagnostic method, 15 subjects (83%) met DSM-IV criteria for a lifetime mood disorder, 10 (56%) for a lifetime anxiety disorder, and 10 (56%) for a lifetime somatoform disorder. None of the subjects met the criteria for a substance use disorder (current or lifetime). The Illness Behavior Questionnaire and the Symptom Check-list-90-Revised results showed little change from 1988 and remained significantly different from the control group on many subscales. The authors conclude that the subjects remain strongly committed to the diagnosis of MCS, and although improved since their original interview, many remain symptomatic and continue to report ongoing lifestyle changes. PMID- 10849459 TI - The influence of HIV-related support groups on survival in women who lived with HIV. A pilot study. AB - To determine the effect of support groups on survival, the authors retrospectively studied 21 HIV-seropositive women who died during the course of participation in a natural history study of HIV. Groups were composed of women who self-selected HIV support groups before death (n = 11) and a comparison group (n = 10). Survival analysis found group participation to be associated with increased longevity (73 months vs. 45 months; P = 0.011). Proportional-hazards regression demonstrated that HIV-related support groups and smaller family size significantly influenced survival (P = 0.0002). Factors related to group participation and ways in which support groups might promote longevity are discussed. PMID- 10849460 TI - Personality profiles in patients referred for chest pain. Investigation with emphasis on panic disorder patients. AB - Patients (N = 199) referred to cardiac outpatient investigation because of chest pain were assessed with the Personality Diagnostic Questionnaire (PDQ-4). Thirty nine percent scored positive for any personality disorder. Borderline and avoidant personality disorders were found significantly more often in patients with panic disorder (PD) (n = 72) than in patients without PD (12.5% vs. 2.5%, 23.7% vs. 7.7%, respectively). In PD patients, the presence of any personality disorder was significantly associated with higher scores of self-reported anxiety agoraphobia symptoms, neuroticism, and the presence of suicidal thoughts. These results suggest that personality pathology is important in a subgroup of patients presenting with chest pain and that these patients may require more extensive treatment. PMID- 10849461 TI - Thrombocytopenia possibly associated with olanzapine and subsequently with benztropine mesylate. PMID- 10849462 TI - Dissociative identity disorder presenting with psychogenic purpura. PMID- 10849463 TI - Aortic aneurysm in the differential for panic attacks. PMID- 10849464 TI - Effectiveness of a psychiatric pain clinic. PMID- 10849465 TI - Introduction PMID- 10849466 TI - Global initiative for asthma (GINA) and its objectives. AB - Guidelines for the diagnosis and management of asthma have been available for the past 13 years and are updated regularly. However, asthma guidelines are rarely completely up-to-date because our knowledge about the pathophysiology and treatment of asthma is continually evolving. Guidelines are an increasingly familiar part of clinical practice and may have potential benefits and harms. The potential harms of guidelines can be minimized if they are rigorously developed using evidence-based medicine. Guidelines should identify key decisions and their consequences and should review the consequences of alternative decisions. In addition, guidelines should be simple, user-friendly and widely disseminated. Guidelines evolve over time and their recommendations should be supported by evidence from clinical trials. Evidence-based medicine is the new paradigm but the results and reaction to the recent meta-analysis on house-dust mite control measures highlights the need for care when incorporating such information into guidelines. Newer therapeutic agents such as the leukotriene receptor antagonists are included in the most recent revision of the GINA guidelines, but their position in asthma therapy is not yet fully established. In countries where asthma guidelines have been implemented, there appears to have been a reduction in the prevalence of moderate persistent asthma but no decrease in severe asthma. This indicates that there is still room for improvement. PMID- 10849467 TI - Presentation of new GINA guidelines for paediatrics. The Global Initiative on Asthma. AB - The Global Initiative on Asthma (GINA) has provided guidelines for the management of children with asthma. For a step-wise approach to therapy, asthma is divided into four categories based on severity of symptoms: intermittent, mild persistent, moderate persistent, and severe persistent asthma. Long-term preventive therapy is distinguished from quick relief therapy in each group. Although these guidelines are clear and simple there have been few studies on asthma therapy for infants. Moreover, the existence of different wheezing phenotypes with varying pathogenic mechanisms hampers the interpretation of these studies. Transient wheezers have stopped wheezing by the age of 3 years and there is no relationship to atopy or a family history of asthma. In contrast, persistent wheezers continue to wheeze from the first year of life throughout school-age and have a high risk of atopy. Although they have normal lung function at birth, persistent wheezers develop significant decrements in lung function by the age of 6 years. Whether these impairments are amenable to prevention by early initiation of anti-inflammatory therapy remains to be seen. At present, there are no disease markers to identify the different wheezing phenotypes in infancy, although eosinophil counts and measurements of eosinophil cationic protein in serum may prove to be helpful in distinguishing these conditions. PMID- 10849468 TI - Asthma: individual patient perspective and current unmet needs. AB - Today there is an array of medication options to treat asthma and state-of-the art guidelines to help diagnose and manage this chronic disease. Despite this, asthma morbidity and mortality rates are rising. There is an urgent need to direct educational programmes and health resources to address this disparity. A recent US survey of asthma patients and health-care providers disclosed that a large number of asthma patients do not fully understand what control of asthma really means and what treatments are available. They underestimate the severity of their condition and over-estimate how well their asthma is being controlled. More treatment choices provide the opportunity for better asthma control for more patients. However, more choices also mean an increased need for education so that the practitioners can determine the best treatment plan for each patient. Provider/patient communication must be enhanced to address patient cultural and lifestyle practices, including environmental exposures. These factors are as important as medication options for successful patient compliance in asthma control. This will require a change in clinician educator skills or more extensive use of ancillary asthma education in clinics. An interactive seminar for physicians on how to communicate better led to patient-physician encounters that were of shorter duration, had significant impact on the prescribing and communication behaviour of physicians, led to more favourable patient responses to physicians' actions, and led to reductions in health care services use. Not all patient educational needs can be met in brief medical visits. Patient education and patient advocate organizations are essential to supplement education given by the physician. Patient organizations help develop culturally and linguistically appropriate educational materials and they organize patient assist systems that help patients exchange information, practice advice and emotional support. PMID- 10849469 TI - Reducing IgE levels as a strategy for the treatment of asthma. AB - IgE secretion by B lymphocytes defines the allergic state and nearly all asthmatics have higher than normal IgE levels in serum following adjustment for age and sex. It is thought that allergic mechanisms may be responsible for the increasing prevalence of asthma. In particular, in utero changes may encourage T cells to differentiate into Th2 subtypes. Th2 cells produce cytokines such as IL 4 and IL-5, which can act indirectly via B cells, mast cells and eosinophils to mediate the asthma phenotype. Alternatively, IL-4 and IL-13 may act directly on the airway. Th2 lymphocyte inflammation in asthma predisposes subjects to B cell and IgE-mediated airway inflammation. IgE binds to receptors on the surface of a variety of effector cells causing them to release a variety of mediators that promote airway hyperresponsiveness, mucus secretion and increased vascular permeability. Several strategies for decreasing IgE have been developed as a possible treatment for asthma. For example, anti-IgE monoclonal antibodies such as rhuMAb-E25 and CGP 56901 block binding of IgE to its high-affinity receptor and have been shown to reduce IgE levels in humans without causing anaphylaxis. IgE levels must be nearly completely suppressed. Recent clinical studies in subjects with asthma have shown that rhuMAb-E25 attenuates both the early and late phase responses to inhaled allergen, and reduces the associated increase in eosinophils in induced sputum. rhuMAb-E25 is well tolerated and has shown promising results in improving symptoms and lung function in patients with moderate to severe asthma. Other strategies for decreasing IgE levels include interferon gamma, IL-4 antibodies, IL-4 receptor antibodies and soluble IL-4 receptors. PMID- 10849470 TI - The role of eosinophils and neutrophils in inflammation. AB - The eosinophil is well recognized as a central effector cell in the inflamed asthmatic airway. Eosinophils release toxic basic proteins and lipid mediators such as cysteinyl-leukotrienes that cause bronchial epithelial damage and airflow obstruction. Eosinophil-selective cytokines and chemokines including interleukin (IL)-5, eotaxin and RANTES may represent targets for novel asthma therapies. In contrast, the role of the neutrophil in asthma remains relatively obscure. Recent evidence from the ENFUMOSA project and elsewhere suggests that neutrophils not only contribute to acute asthma exacerbations, but also are present in high numbers in the airways of patients with chronic severe asthma. Production by neutrophils of lipid mediators, reactive oxygen intermediates (ROI) and proteases such as elastase, may contribute to airflow obstruction, epithelial damage and remodelling. Leukotriene B4 and cytokines such as IL-8, granulocyte-macrophage colony stimulating factor (GM-CSF), and tumour necrosis factor (TNF)alpha chemoattract neutrophils and reduce neutrophil apoptosis, and selective agents directed against these may prevent neutrophil influx and accumulation. Airway neutrophilia remains apparent in severe asthma patients even after treatment with high doses of corticosteroids. In vitro, corticosteroids paradoxically enhance neutrophil survival by reducing apoptosis, so corticosteroid therapy may exacerbate neutrophil activity in vivo. Both corticosteroids and cytokines may suppress neutrophil apoptosis by upregulating endogenous synthesis of leukotriene (LT)B4. Specific blockade of LTB4 synthesis or LTB4 receptors may induce neutrophil apoptosis and combat the unwanted effects of high-dose steroids on neutrophil survival. Phagocytosis of apoptotic neutrophils stimulates important signals that down-regulate pro-inflammatory cytokine production by macrophages, allowing resolution and repair processes to prevail. PMID- 10849471 TI - The role of mast cells and basophils in inflammation. AB - Mast cells are positioned in the asthmatic airways so that they are able to respond to the inhaled environment. During active disease, the cells are primed to secrete an array of preformed and newly generated inflammatory mediators including histamine, neutral proteases and heparin sulphate, prostaglandins and cysteinyl leukotrienes as well as an array of cytokines and chemokines that are involved in leucocyte recruitment and activation. These cells are a potent source of mediators in both allergen- and exercise-induced asthma and possibly in asthma provoked by other stimuli such as adenosine and inhaled air pollutants. The important role played by mast cells in maintaining airway dysfunction in asthma is underpinned by the efficacy of mediator inhibitors, such as those interfering with the release or action of the leukotrienes, agents that inhibit mast cell activation such as sodium cromoglycate and the recently studied E-20 humanized monoclonal antibody that binds to and removes IgE. The recent discovery of novel inhibitory pathways involving inhibitory motifs (ITIMS) on critical cell surface signalling molecules has opened up new possibilities for preventing mast cell activation. Future research will focus on more effective ways for inhibiting the mast cell's contribution to asthma and understanding what role this unique cell has in the pathogenesis of airway wall remodelling. PMID- 10849472 TI - Learning from vascular remodelling. AB - The positioning of a hollow silicone collar around the carotid artery of a rabbit induces many changes of early atherosclerosis including intimal proliferation of smooth muscle cells. This occurs below an intact endothelium indicating that endothelial damage is not necessary for smooth muscle cell proliferation. The endothelium may in fact produce substances that control processes occurring in the intima. Vascular endothelial growth factor (VEGF) is an angiogenic agent that is produced by cultured vascular smooth muscle cells. The combination of hypoxia and factors such as platelet-derived growth factor, tumour necrosis factor alpha, basic fibroblast growth factor, and interleukin-1beta lead to synergistic production of VEGF by cultured smooth muscle cells. VEGF receptors are present predominantly on the endothelium and may be an important target for modulating the response to damage, hypoxia and inflammation. Transfection of the gene for VEGF resulted in inhibition or regression of intimal hyperplasia induced by the silicone collar in the rabbit. Studies suggest that the two mediators responsible for this inhibition of smooth muscle cell proliferation are nitric oxide and prostacyclin, which are produced by cultured endothelial cells incubated with VEGF. Thus, VEGF produced by smooth muscle cells in response to hypoxia, damage or inflammation, acts on specific endothelial receptors to produce nitric oxide and prostacyclin, which inhibit smooth muscle cell proliferation. Failure of this process could give rise to intimal hyperplasia. Early clinical studies of VEGF transfection from the outside of human arteries using a biodegradable collar are in progress. PMID- 10849473 TI - Epithelial damage and response. AB - Epithelial damage is a characteristic feature of asthma. The epithelium is not merely a passive barrier but can generate a range of mediators that may play a role in the inflammatory and remodelling responses that occur in the lungs in asthma. For example, the cytokine granulocyte macrophage colony-stimulating factor (GM-CSF), whose principal source is the epithelium, can prolong eosinophil survival while transforming growth factor is a potent profibrogenic cytokine. Deposition of collagen in the epithelial subbasement membrane is a characteristic feature of the remodelling response in asthma. This may be due to abnormal associations between myofibroblasts and epithelium, both of which are involved in early lung development (epithelial-mesenchymal trophic unit). In asthma, there may be a primary defect in the epithelium such that it responds abnormally to various stimuli and cannot undergo the normal repair response. Epidermal growth factor (EGF) appears to be a key factor in bronchial epithelial repair; it stimulates epithelial cell proliferation and migration. The 3v isoform of the adhesion molecule CD44 is overexpressed in damaged epithelium and seems to regulate the repair response by presenting EGF more efficiently to its receptor. Although EGF receptor expression is increased in asthma, it does not lead to an appropriate proliferative response and restitution of normal epithelium. Other factors such as transforming growth factor (TGF)beta which are generated by inflammatory cells and epithelium are also upregulated in asthma. An epithelial/fibroblast co-culture system has shown that following epithelial damage various growth factors are released from the underlying myofibroblasts and are responsible for the proliferative response. The TGFbeta family are most likely responsible for collagen production. In an in vitro study, an EGF receptor inhibitor slowed epithelial repair but enhanced TGFbeta production by the slowly repairing epithelial cells. In conclusion, the interaction between epithelial cells and myofibroblasts, i.e. reactivation of the epithelial-mesenchymal trophic unit appears to be central to the airway wall remodelling response. PMID- 10849474 TI - Fibrosis and airway remodelling. AB - The term 'airway remodelling' is now widely used to refer to the development of specific structural changes in the airway wall in asthma. Particular interest has focused on subepithelial fibrosis, myofibroblast accumulation, airway smooth muscle hyperplasia and hypertrophy, mucous gland and goblet cell hyperplasia, and epithelial disruption. The presence of these features is generally accepted, but further studies are still required to define the changes occurring more precisely at the pathological and ultrastructural levels. Attention also needs to be directed towards the existence of such changes in small airways. The natural history of the response has not been well described: remodelling is present in the airways of asthmatic children and of adults with newly diagnosed asthma, and studies that have attempted to relate the extent of remodelling to disease severity have produced conflicting findings. The role of remodelling in the progressive decline in lung function leading to fixed airflow obstruction seen in some patients is also unclear. Epidemiological studies are currently hindered by the absence of a useful non-invasive marker of remodelling. Airway remodelling is frequently assumed to be a consequence of chronic inflammation, but the precise relation between the remodelling and inflammatory components in asthma is unclear. The cellular and molecular events underlying the remodelling process are also poorly understood. There is therefore a need for the development and characterization of animal models that will allow these issues to be explored. Finally, the ability of currently available anti-asthma therapies to prevent or reverse airway remodelling is uncertain. There is some evidence that early treatment with inhaled corticosteroids can lead to improved outcome in asthma but this needs confirmation. Studies addressing the ability of corticosteroid treatment to reverse established structural changes have not produced consistent findings, and there is little information with regard to other therapies such as theophylline and antileukotriene agents. Effective treatment of airway remodelling may require the development of novel therapies directed against appropriate targets. PMID- 10849475 TI - The role of co-stimulation in airway inflammation. AB - There is considerable evidence to support an important role for co-stimulatory molecules in regulating the proliferation and activation of T cells in the immune response. Of particular relevance is the interaction between CD28 on T cells and B7 expressed on the surface of antigen presenting cells (APCs). CTLA-4, another molecule present on activated T cells may downregulate T cell activity, but its role remains uncertain. CTLA4-Ig, a fusion protein consisting of the extracellular domain of CTLA4 and the Fc portion of human immunoglobulin G1 (IgG1), has been useful for studying the role of CD28/B7 interactions in immune responses. A number of studies have shown that CTLA4-Ig can switch off T cell activation. In an ovalbumin sensitive murine model of asthma, CTLA4-Ig treatment suppressed the response to inhaled allergen (increased airway hyperresponsiveness [AHR], IgE production, recruitment of eosinophils into the lungs, production of IL-4, IL-5, and IL-10 and increased IFNgamma production from CD3-TCR-activated T cells). Anti B7-2 treatment has similar effects suggesting that interaction of B7 2 with CD28 is important in the development of a Th-2 type inflammatory response in mice. Recent observations have been of relevance to human allergic disease. In vitro studies have shown that CTLA4-Ig or anti-B7-2 antibody can inhibit allergen induced proliferation and cytokine production by peripheral blood mononuclear cells from atopic subjects. The role of co-stimulation has been studied in a human bronchial explant model of asthma. CTLA4-Ig fusion protein effectively blocked allergen-induced production of IL-5 and IL-13 in bronchial explants from atopic asthmatics. These studies confirm the requirement for interaction between co-stimulatory molecules in cytokine production and allergic inflammation, and point to the CD28-B7 pathway as being important to the allergen-induced inflammation in asthma. Studies of organ transplantation in primates suggest that CTLA4-Ig is extremely effective in preventing organ rejection. While phase 1 clinical trials have shown CTLA-4-Ig treatment of patients with psoriasis vulgaris to be well tolerated and to result in clinical improvement, its role in asthma management merits further investigation. PMID- 10849476 TI - The bronchial microcirculation in asthma. AB - Airway wall remodelling in asthma involves a number of changes including increased vascularity, vasodilation and microvascular leakage. Evidence suggests that the number and size of bronchial vessels is increased in patients with asthma compared with normal controls. In particular, there may be increased numbers of vessels in patients with fatal asthma. Treatment with inhaled corticosteroids is now known to reduce this vascularity. Bronchial vessels may undergo proliferation in response to inflammatory stimuli. Many factors can induce angiogenesis including a range of mediators and growth factors. Others can cause a vascular response by causing vasodilation and microvascular leakage. Airway wall oedema is likely to be important in asthma but has not yet been quantified. It is thought that mast cells play a key role in modulating these vascular remodelling changes by releasing cytokines and growth factors. Many key issues remain to be resolved before we fully understand the role of the bronchial microcirculation in asthma. In the future, novel therapies may be directed towards angiogenesis, vasodilation and microvascular leakage. PMID- 10849477 TI - Airway smooth muscle as a target in asthma. AB - Traditionally, the contractile properties of airway smooth muscle have been regarded as its sole contribution to the pathogenesis of asthma. However, our understanding of the role that this structural cell plays in asthma is changing. Airway smooth muscle can undergo hyperplasia and/or hypertrophy leading to structural changes in the airway wall which contribute to the development of persistent airway obstruction and increased non-specific airway hyperresponsiveness in chronic severe asthma. Many studies in vitro have characterized airway smooth muscle proliferation induced by various pro inflammatory mediators, growth factors and components of the extracellular matrix, but the mediator(s) responsible for the observed increase in airway wall smooth muscle content in vivo remain to be determined. In addition to geometric obstruction by increased airway wall thickening, proliferating airway smooth muscle cells undergo phenotypic modulation from a contractile to synthetic proliferative state where additional functions of airway smooth muscle such as cytokine/chemokine and extracellular matrix secretion may become more apparent. This may be especially relevant in the diseased airway where the content of airway smooth muscle as a fraction of the total cells present in the airway wall is already increased. Airway smooth muscle cells may also interact by direct contact with immunocytes such as T lymphocytes through expression of cell adhesion molecules with the result that myocyte DNA synthesis is induced. As additional functions of airway smooth muscle are described, a more contemporary view is emerging that airway smooth cells may adopt an immuno-effector role in chronic asthma by proliferating, secreting cytokines, expressing adhesion molecules and by interacting with various inflammatory cells. This may involve changes in the phenotypic status of airway smooth muscle, and as a result, these cells may play an active role in perpetuating and orchestrating airway inflammation in the remodelled airway. An important phase for future airway smooth muscle research will be to determine the extent that these putative mechanisms exist in vivo in the pathogenesis of chronic severe asthma. PMID- 10849478 TI - Neural mechanisms in asthma. AB - Advances in the understanding of neural mechanisms in asthma may provide novel therapeutic approaches in the treatment of asthma. Excessive activity of cholinergic nerves may be important in asthma. Dysfunction of M2 muscarinic receptors in asthma may lead to excessive bronchoconstriction and mucus secretion and can be induced in animal models by a range of stimuli including allergen, viral infection, ozone, eosinophil products and cytokines. Cholinergic mechanisms may be especially important in certain types of patients and anticholinergic agents provide protection against bronchospasm due to psychogenic factors or beta2-blockers. Non-adrenergic non-cholinergic (NANC) mechanisms, both inhibitory (i-NANC) and excitatory (e-NANC), may play a significant role in the pathophysiology of asthma. The putative neurotransmitters, vasoactive interstinal polypeptide (VIP) and nitric oxide (NO), mediate neural bronchodilation in human airways. There does not appear to be a defect in the i-NANC system in moderate or severe asthma. e-NANC is mediated by the sensory neuropeptides substance P (SP) and the more potent bronchoconstrictor neurokinin A (NKA). Various studies suggest that the SP content of human airways is increased in asthma. Tachykinins are not only present in sensory nerves, but also are produced by inflammatory cells such as alveolar macrophages, dendritic cells, eosinophils, lymphocytes and neutrophils. They can be released into the airways by stimuli such as allergen and ozone. Evidence suggests that in addition to smooth muscle contraction, which is mediated mainly by NK2 receptors, tachykinins also cause mucus secretion, plasma extravasation and stimulate inflammatory and immune cells. These effects are mediated by NK1 receptors. Recent studies have shown that NK2 receptor antagonists such as saredutant partially inhibit NKA-induced bronchoconstriction in asthmatics. Thus, tachykinin receptor antagonists have potential as therapies for asthma. PMID- 10849479 TI - Unmet needs in adult asthma. AB - Guidelines have been developed to provide an objective framework for the effective management of asthma. They are based on a mix of sound clinical practice and evidence-based medicine. The aims of asthma management are to ensure that the patient is symptom free and living an unrestricted life with normal physical activity, lung function normalized as much as possible, using minimum therapy, that exacerbations are kept to a minimum and mortality is reduced or abolished. In practice, however, these aims are not being met. Thus, unmet needs exist in asthma and need to be addressed. One particular unmet need is poor delivery of asthma care. Undertreatment of asthma is common, especially in severe asthma. However, studies suggest that some patients with severe asthma do not respond to any available treatments. Many outcome measures have been used in asthma and different outcomes will ensue according to which outcome measure is chosen, which in turn depends on the population studied. Asthma is a heterogeneous disease and there is heterogeneity in the response to treatment. However, at present there are no means to determine which patients will be a responder or non-responder to a particular treatment. In clinical trials of potential new asthma treatment we need to target the patient population more carefully according to the selected outcome measure, which should reflect the patient's perspective. PMID- 10849480 TI - Unmet needs in the treatment of asthmatic children and adolescents: 1. AB - The paediatric asthma guidelines have been successful in providing a uniform approach to the management of asthma for the medical profession as a whole. Unfortunately, the guidelines were generated without input from patients themselves and consequently do not consider issues that are important to patients such as a preference for oral treatment. Asthma is a heterogeneous group of conditions and the guidelines do not sufficiently define subgroups of patients and their particular needs. As a result, there has been a tendency to assume that all wheezing in infancy is asthma and this had led to gross overtreatment in certain patients. In contrast, severe asthma often remains underdiagnosed and undertreated. The most recent revision of the guidelines has classified asthma in terms of the patterns of disease; infrequent episodic, frequent episodic and chronic persistent. The treatment required for each of these groups is clearly defined and there is no need for stepwise therapy. Other changes to the guidelines will occur and are needed. None of the treatments available can modify the natural history of asthma; they control the symptoms not the disease process. Evidence from bronchial biopsies suggests that both inflammation and remodelling occur early, even before the first symptoms appear. We need to look for the factors in early life that predict which children will go on to develop asthma and intervene at that stage. Anti-histamines and leukotriene receptor antagonists may be interesting as interventions in that respect. Two important unresolved issues are to understand what drives remodelling and inflammation, and to look at early life origins of asthma. These approaches may provide effective therapeutic targets and, ultimately, a means of prevention. PMID- 10849481 TI - Unmet needs in the treatment of asthmatic children and adolescents: 2. AB - The five main unmet needs in the treatment of asthma in children and adolescents are the treatment of infants, the treatment of adolescents, the efficacy of medication, the lack of clinical trials in children and the need for prevention. Asthma is a dynamic disease that changes over time and there are several patterns of asthma in children. One of the greatest needs is to identify those children with chronic asthma where undertreatment is more of a problem than overtreatment. Although the treatment of asthma in children is the same in principle as adults, there are several practical difficulties with infants, such as accurately assessing lung function and administering drugs. Teenage asthma creates a distinct management problem. Not only is asthma underdiagnosed but acceptance of diagnosis and compliance is often poor. Existing therapies need to be used more effectively in adolescents. It is particularly important to establish a partnership with teenage asthmatics and motivate them to create their own treatment goals. This requires passing on knowledge. The efficacy of asthma medication may be different in children than in adults and clinical studies should be performed to optimize therapy with existing drugs. The only proven primary preventive measure to prevent wheezing and asthma is to avoid passive tobacco smoke exposure. Possible intervention strategies include early intervention with anti-inflammatory therapy, allergen avoidance and vaccine approaches but more of them have proven efficacious. Although modern asthma treatment has improved patient quality of life and long-term prognosis, there is still a need for further improvement. PMID- 10849482 TI - Hormonal deficiency in elderly males. AB - During the aging process, a number of morphological and neurochemical alterations have been found in the supra-chiasmatic nuclei, which are in part responsible for the age-dependent decrease in plasma testosterone (andropause or PADAM), DHEA (adrenopause), GH/IGF-I (somatopause) and melatonin that develops in most men at about the age of 50 (Perry, 1999; Vermeulen et al., 1999). An important principle in antiaging practice is the employment of the best of available means to prevent the preventable and delay the inevitable. Therefore, some scientists advocate multihormonal replacement therapy and the use of antioxidant drugs that may favourably influence some of the pathological conditions in aging men. PMID- 10849483 TI - Male reproductive health research needs and research agenda: Asian and Pacific perspective. AB - Enhancing male reproductive health, and increasing men's participation in it, involves encouraging a range of positive reproductive health and social behaviour by men to help ensure women's and children's well-being. More intellectual work including research programmes-is urgently needed to clarify the conceptual framework for male reproductive health. At the Asia and the Pacific Symposium 'Intra-regional Cooperation in Reproductive Health Research' (Shanghai, China, 12 13 October 1998) the Symposium participants identified regional research needs and recommended a regional reproductive health research agenda, which addresses six key issues related to male reproductive health: (i) male contraceptive technology; (ii) reproductive tract infections/sexually transmitted diseases and male infertility; (iii) male involvement in reproductive health; (iv) male adolescent reproductive health; (v) male reproductive ageing; and (vi) environment and male reproductive health. One of the major challenges now facing us is the elaboration of a comprehensive, yet realistic, male reproductive health research agenda that reflects the needs and demands of Asian developing countries. Making full use of an interdisciplinary approach is of strategic importance to achieve this. PMID- 10849484 TI - Hormonal male contraception. PMID- 10849485 TI - 7alpha-methyl-19-nortestosterone (MENT): the optimal androgen for male contraception and replacement therapy. PMID- 10849486 TI - Semen banking in patients with cancer: 20-year experience. AB - Modern techniques of banking sperm provide an effective way to preserve the option of future fertility for most teenagers and young men diagnosed with a variety of malignancies that will necessitate treatment with chemotherapy, pelvic surgery, or significant radiation doses to the testes. Results of cumulative data collected at the Cleveland Clinic Foundation from patients with testicular cancer, lymphoma, leukemia, sarcoma, carcinoma and other kinds of malignancy have revealed that: (1) pretreatment semen quality (pre-freeze and post-thaw) in patients with cancer is poorer compared with healthy donors; (2) the percentage decline in semen quality (from pre-freeze to post-thaw) in patients with cancer is similar to that of normal donors. This suggested that the effect of cryodamage on spermatozoa from patients with cancer is similar to that of normal donors. (3) The stage of cancer in patients with testicular cancer and Hodgkin's disease shows no relationship to their semen quality. Based on studies conducted at the Cleveland Clinic Foundation, we recommend that sperm cryopreservation be offered to all men of reproductive age who have malignancies. Cryopreservation is safe and inexpensive, and gives patients a chance to establish pregnancies in the future with an assisted reproductive technique. PMID- 10849487 TI - Genetic screening for patients with azoospermia and severe oligo-asthenospermia. AB - In order to explore the genetic defects of patients with azoospermia or severe oligo-asthenospermia, screening examinations were carried out for the chromosome disorder and gene deletion of the Y chromosome for 220 male infertility patients. The present results show that the total prevalence of genetic defects is 23.6%, including 38 patients (28.4%) with chromosome disorder and 14 patients (16.8%) with gene deletion in the Yq arm. The most prevalent chromosome anomaly is 47XXY (Klinefelter's syndrome), which includes 18 cases of pure type and three cases of mosaic type. Variable autosomal translocations occurred in both the azoospermia group (5.2%) and the oligo-astheno-spermia group (5.8%) with similar prevalence. A total of 22 patients had deletions of the variable, interstitial portion of the Yq arm. These gene deletions are distributed not only inside the AZF region, but also outside of this region. The severity of deletions is not well correlated to the clinical testicular function of the patients. We conclude that chromosome disorder and gene deletions are the causative factors of patients with azoospermia and oligo-asthenospermia. Genetic screening should be a routine examination for them before the use of assisted-reproductive technologies. PMID- 10849488 TI - The effects of isolated lipoproteins and triglyceride, combined oxidized low density lipoprotein (LDL) plus triglyceride, and combined oxidized LDL plus high density lipoprotein on the contractile and relaxation response of rabbit cavernous smooth muscle. AB - The aim of this study was to investigate the effects of isolated lipoproteins and triglyceride (TG), and the effects of combined oxidized low density lipoprotein (LDL) plus TG and the combined oxidized LDL plus high density lipoprotein (HDL) on the contractility and relaxation response of rabbit cavernous smooth muscle. Cavernous muscle strips from New Zealand White rabbits were studied in organ chambers for isometric tension measurement. The strips were exposed to HDL, LDL, oxidized LDL, TG, combined oxidized LDL plus TG and combined oxidized LDL plus HDL for 30 min. Both HDL and LDL did not affect contraction and relaxation responses of the cavernous muscles. The oxidized LDL did not affect norepinephrine (NE)-induced contractility of the strips, but significantly (p < 0.05) decreased the relaxation response to endothelium-dependent agonist, acetylcholine (Ach). Non-specific NO synthase inhibitor (L-NAME) completely inhibited the relaxation response to Ach, and L-arginine partially improved the diminished relaxation. TG did not significantly change the relaxation responses to Ach, but decreased the contractility of cavernous muscle to NE. Neither the combined oxidized LDL plus TG nor oxidized LDL plus HDL had significant synergistic or detoxication effects on the contractility and relaxation responses. In conclusion, oxidized LDL may have acute toxic effects on the endothelium-dependent, NO-mediated relaxation, but not on the contractility, of rabbit cavernous smooth muscle. TG may decrease contractility of the cavernous muscle. There may be neither synergistic nor detoxication effects on the contractility and relaxation response when TG or HDL is added to the oxidized LDL. PMID- 10849489 TI - Expanding our understanding of spermatogenesis: the future genetic tests for infertility. PMID- 10849490 TI - Prostate cancer in Taiwan: epidemiology and risk factors. AB - The incidence of prostate cancer (PC) has been rapidly increasing in the past 10 years in Taiwan. It became the sixth common cancer in males in 1996 and resulted in 540 deaths in 1998. It was estimated that the incidence of PC would be up to 14 per 105 in the year 2000, 3-fold higher than that in 1990. Three factors may be responsible for the increase of PC in Taiwan: use of prostate specific antigen, population aging and high fat diet. A case-control study on the risk factors of PC in a patient population comprised mainly of veterans (63%) in Taiwan showed that PC patients tended to have engaged in more physical activity (adjusted odds ratio (OR): 2.16), have a lower body mass index (OR 2.0) and be less likely to consume vegetables cooked with pork lard (OR 0.47). About half of our patients had locally advanced or metastatic diseases upon diagnosis. This staging distribution was consistent in most major institutions in Taiwan and was fairly unchanged over time in the past two decades. The incidence of latent PC in Taiwan was not investigated until a pathological review of 49 cystoprostatectomy specimens recently revealed unsuspected PC in 32.7% and high-grade prostate intra epithelial neoplasia in 49% of the prostates removed. As the incidence of PC grows rapidly in Taiwan, this disease warrants more attention from the public and the authorities. More efforts should also be directed to the investigation on the risk factors for PC in the new millennium. PMID- 10849491 TI - Prostate cancer in Honolulu, Hawaii: pathological and clinical characteristics. AB - We reviewed trends in prostate cancer at our institution to determine if there are changing characteristics that need to be considered in conducting comparative studies with patients from other institutions. Assessed between 1993 and 1998 were trends in PSA status, race, tumour grade, clinical stage and treatment. The incidence dropped between 1993 and 1994, but has remained stable since, except in Philipinos. Changes in clinical tumour stage and PSA positive rate were not noted; Gleason score distribution changed with marked decline of low grade tumours. This reflects the tendency pathologists had of undergrading prostatic adenocarcinoma in biopsy specimens during the early 1990s. Use of hormonal treatment increased from 21% to 57%. Patients receiving radiation therapy also increased slightly. Comparative studies between institutions would require review of the biopsy material for accurate Gleason scores. Outcome studies would need to control for the changing pattern of prostate cancer therapy. PMID- 10849492 TI - Clinical characteristics of prostate cancer in Gunma prefecture. AB - Prostate cancer clinical data from patients registered at Gunma University Urologic Oncology Study Group were compared between pre-PSA and PSA eras, dividing them between those patients diagnosed at clinical practice and those diagnosed on the Gunma mass-screening programme. In the MS, the number of incidents of prostate cancer increased from 126 out of 16750 screened cases in the pre-PSA era (0. 75%) to 227 out of 19782 screened cases in the PSA era (1.15%). The average age was lower in the PSA era. As for clinical stage, stage B and C cases increased while stage D cases decreased. For the clinical practice cases at the Gunma University Urologic Oncology Study Group, the number of registered incidents of prostate cancer increased from 1164 in the pre-PSA era to 2098 in the PSA era. The average age was similar between the pre-PSA and PSA eras. The percentage of cases who were stage D declined from 46.1 during the pre PSA era to 37.7 in the PSA era. In the PSA era, earlier and more cases of prostate cancer are diagnosed both in clinical practice and in the mass screening program in Gunma Prefecture. PMID- 10849493 TI - Clinical-pathological comparison of clinical prostate cancer between Japanese Americans in Hawaii and Japanese living in Japan. AB - In an attempt to determine the role of environmental factors in the etiology of prostate cancer, we compared the clinical-pathological findings of prostate cancer among Japanese Americans in Hawaii and Japanese living in Japan. Our study showed that prostate cancer in Japanese living in Japan is more advanced than in Japanese Americans. The findings indicate that screening for prostate cancer in Japan is far behind that in the USA. The difference in level of cancer screening also precluded our attempt to analyse environmental factors contributing to prostate cancer progression in the two groups. The variation in method of clinical detection of prostate cancer between the USA and Japan is also likely to contribute to the apparent difference in the incidence of the disease. PMID- 10849494 TI - Endocrine disruptors and male reproductive function--a short review. AB - Semen quality has decline in many countries over the last few decades. There has been an increase in the incidence of testicular cancer world-wide. The incidences of cryptorchidism and hypospadias have also increased in many countries. A biological plausible hypothesis has suggested that man-made chemicals act as endocrine disruptors resulting in altered development of the reproductive tract causing the observed effects. Endocrine disruptors include natural products, pharmaceuticals, industrial products and environmental pollutants. There are limitations in the current in vivo and in vitro assays for the assessment of endocrine disruptors. Epidemiological human studies are necessary to fill in the gap of knowledge. Based on the current knowledge, the impact of endocrine disruptors on the male reproductive function remain to be appreciated. PMID- 10849495 TI - Feasibility of surveillance of changes in human fertility and semen quality. AB - To show that male fertility is declining is not simple. Few men volunteer and recruitment bias may lead to over-representation of the subfertile. Semen analysis has errors arising from counting and poorly standardized criteria, which may be overcome by automation. Time to pregnancy (TTP)-the number of menstrual cycles taken to conceive-measures fertility and allows male recruitment bias to be estimated. We review automated measurement of sperm concentration, motility and morphology and present a preliminary report on a study to assess a retrospective TTP questionnaire, recruitment bias and feasibility for large-scale surveillance of fertility. PMID- 10849496 TI - Myth and methodology in the evaluation of human sperm output. PMID- 10849497 TI - Current status of reproductive function in Japanese fertile men: international collaborative project on a study of partners of pregnant women. AB - The data on reproductive function in 255 Japanese fertile men resident in the Kawasaki/Yokohama area in Japan were described. The sperm concentration was 107.9 +/- 97.4 x 106/mL. The semen volume was 3.2 +/- 1.5 mL and percentage motile spermatozoa (grade A + B in WHO criteria) was 56.8% +/- 14.7%. The evidence for secular changes in semen quality and other changes in male reproductive health is inconclusive, although regional differences would appear to be stronger. The present study is the first large-scaled prospective survey on the reproductive function of Japanese normal men proven fertility, which was planned as an international comparative study. PMID- 10849498 TI - Spermatogonial transplantation technique in spermatogenesis research. AB - The spermatogonial stem cell is the foundation of spermatogenesis and it continues to divide throughout the lifetime of a male. It divides to renew itself as well as to produce daughter cells that finally differentiate to spermatozoa. Transplantation of spermatogonial stem cells from a fertile mouse to the testis of a sterile mouse results in the development of donor cell derived spermatogenesis. Furthermore, spermatogenesis of the rat and hamster occurs in immunodeficient mouse testes following spermatogonial transplantation. Cryopreservation of spermatogonial stem cells has been shown to be applicable in many species for later transplantation. In addition, certain forms of infertile donor germ cell can be transplanted to produce functional sperm in a recipient testis. These new techniques will be useful for basic research of spermatogenesis and for application to reproductive technology in the future. PMID- 10849499 TI - Recent advances in the microinsemination of laboratory animals. AB - Microinsemination techniques were applied to laboratory animals, including mice, Mongolian gerbils, mastomys, guinea pigs and cynomolgus monkeys. After micro insemination with spermatozoa or round spermatids, their oocytes were successfully fertilized and subsequently developed into 4-cells to blastocysts, depending on species. The efficiencies of microinsemination techniques for producing fertilized oocytes were comparable with, or superior to those of IVF in these species. PMID- 10849500 TI - Oocyte activation induced by spermatids and the spermatozoa. AB - It has been reported that a sperm factor (SF) found in spermatozoa plays a critical role in fertilization. However, particulars of the oocyte-activating and Ca2+ oscillation (Ca-Os)-inducing abilities of this SF remain unknown. We examined these abilities of spermatids in mouse, hamster and human by a mouse test (injection of spermatids into mouse oocytes). In mice, the round spermatids (ROS), elongated spermatids (ELS) and spermatozoa activated 0%, 93% and 92% of the oocytes, respectively. ROS injection resulted in no Ca-Os (type C). ELS induced a normal oscillation (type A) at 0% and an abnormal oscillation (type B) at 94%. Mouse spermatozoa induced type A Ca-Os at 90%. For mice, oocyte activating and Ca2+ oscillation-inducing ability arose in different phases of spermiogenesis. We also observed this differential timing for hamster spermatids. Hamster ROS activated 74% of oocyte (ELS: 90%, sperm: 86%). Human ROS activated 64% of oocytes (sperm: 100%), but only 35% of the oocytes showed type A Ca-Os. These results indicate that oocyte activation generally occurs between the ROS and ELS phases, although these phases differ among species. They also indicate that oocyte activation is not necessarily accompanied by Ca-Os. These findings suggest the existence of different thresholds at which the SF induces oocyte activation and Ca2+ oscillation, or of different factors that induce oocyte activation and Ca-Os. We found SF to be clinically impaired in 0.9% of ICSI patients. A combination of artificial oocyte activation and ICSI proved effective with such patients. PMID- 10849501 TI - New concepts in operative andrology: a review. AB - Several recently published, or about to be published, controversial issues in operative andrology are clarified and reviewed in this paper. The microsurgical technique for sperm retrieval for nonobstructive azoospermia, the round spermatid controversy and the varicocoele dilemma (why does everybody keep doing varicocoelectomy for male factor infertility?) are presented with salient points that have recently been presented elsewhere and referenced. Finally, at the end, we review briefly what is known about the likelihood of genetic transmission of infertility from male factor patients to their offspring as a result of the new ICSI technology. PMID- 10849502 TI - Prevalence of erectile dysfunction in Thailand. Thai Erectile Dysfunction Epidemiological Study Group. AB - A study of the prevalence rate of erectile dysfunction (ED) in the Thai population has never been done previously, except for a small study in the hospital. The project was carried out across the whole country, including in the north, south, eastern and central plains, and there were representatives from one small and one large province and the Bangkok metropolitan area. There were 250 males in each area, giving as total of 1250 males. The interviews were carried out in urban areas, so that the questions and answers could produce good data. The interviewer was trained by one of our EDACTT members, before going to the interview locations, and the supervisor were also onsite to clarify any the questions that might occur. The questions and pretest were carried out stringently, to help in term of statistics. All the health questions were asked taking care to accommodate the interviewee's feelings, so as not to cause embarrassment. The interviews were held individually and strictly privately, so that the interviewees could speak freely The interviewees were between 40 and 70 years old, to match with MMAS. The rate of ED in this age group is increasing gradually, and the relationship between ED and hypertension, diabetics or heart disease, and lifestyle factors, including smoking habits, alcohol consumption, caffeine and risk factors is of interest. PMID- 10849503 TI - Paraventricular nucleus of hypothalamus - a brain locus in central neural regulation of penile erection in the rat. PMID- 10849504 TI - Phytochemicals and the breakthrough of traditional herbs in the management of sexual dysfunctions. AB - Traditional herbs have been a revolutionary breakthrough in the management of erectile dysfunction and have become known world-wide as an 'instant' treatment. The modern view of the management of erectile dysfunction subscribes to a single etiology, i.e. the mechanism of erection. A large number of pharmacological agents are orally consumed and vasoactive agents inserted intraurethrally or injected intrapenially to regain good erection. Modern phytochemicals have developed from traditional herbs. Phytochemicals focus their mechanism of healing action to the root cause, i.e. the inability to control the proper function of the whole body system. Hence phytochemicals manage erectile dysfunction in the frame of sexual dysfunction as a whole entity. Protodioscin is a phytochemical agent derived from Tribulus terrestris L plant, which has been clinically proven to improve sexual desire and enhance erection via the conversion of protodioscine to DHEA (De-Hydro-Epi-Androsterone). Preliminary observations suggest that Tribulus terrestris L grown on different soils does not consistently produce the active component Protodioscin. Further photochemical studies of many other herbal plants are needed to explain the inconsistent results found with other herbal plants, such as in diversities of Ginseng, Eurycoma longifolia, Pimpinella pruacen, Muara puama, Ginkgo biloba, Yohimbe etc. PMID- 10849505 TI - Algorithm for diagnosis and treatment of erectile dysfunction in the era of sildenafil citrate. AB - Since Viagra (sildenafil citrate) was released as a pharmaceutical agent for the treatment of erectile dysfunction, it has had a big impact on the work undertaken in our practice. I herewith present an algorithm for the diagnosis and treatment of erectile dysfunction in the era of sildenafil citrate. PMID- 10849506 TI - New therapies for erectile dysfunction. AB - The quest for improving and maintaining sexual function has been going on since time immemorial. The advent of an effective oral drug, sildenafil, has brought about unprecedented open discussion on male erectile dysfunction, and gas accelerated the pace of development of new therapies for erectile dysfunction. New knowledge in the physiology of sexual function has enabled researchers to target drug treatment at the whole network of the central nervous system and the numerous cascadic enzymatic reactions leading to relaxation of the corporal smooth muscle. One of the brightest potential applications of future molecular technology in the study of erectile dysfuction is in the utilization of gene therapy. PMID- 10849507 TI - Enrichment and transplantation of spermatogonial stem cells. AB - Spermatogenesis is a complex, highly organized process originated from stem cell spermatogonia. Because there are very few stem cells and they can only be defined by their function, the identification and isolation of these cells has been very difficult. By using a spermatogonial transplantation assay system, we have identified alpha6-and beta1-integrin expression on stem cells, and cells isolated with these antigens were significantly enriched in stem cells. This is the first demonstration of spermatogonial stem cell-associated antigens. Analysis of two infertile mouse models, Steel/SteelDickie (Sl/Sld) and experimental cryptorchidism, showed that the number of stem cells is reduced in Sl/Sld testis. Whereas cryptorchid testes are greatly enriched for stem cells, and one in 200 cells is a stem cell. These techniques will provide an important starting point for further purification and characterization of spermatogonial stem cells. PMID- 10849508 TI - Temporal control of protein synthesis during spermatogenesis. AB - During oogenesis and spermatogenesis transcription ceases prior to the differentiation of the mature cells. To complete germ cell differentiation and initiate early embryogenesis, proteins are synthesized from pre-existing mRNAs that are stored for several days. It is well established that important regulatory elements functioning in spatial localization, temporal translation or messenger RNA stability are located in the 3' untranslated region (3' UTR) of mRNAs. During mammalian spermatogenesis temporal translational regulation of the protamine 1 (Prm1) mRNA is dependent on a highly conserved sequence located in the distal region of its 3' UTR. The 17-nucleotide translational control element (TCE) mediates translational repression of the Prm1 mRNA. Mutation of the TCE causes premature synthesis of protamine protein and sterility. The Prm1 mRNA is stored as a cytoplasmic ribonucleoprotein (mRNP) particle in spermatids. Contained within the particle are several members of the Y box family of nucleic acid binding proteins. In the yeast three-hybrid system the murine Y box proteins MSY1, MSY2 and MSY4 bind in a sequence-dependent manner to a conserved region in the proximal portion of the Prm1 3' UTR. Sequence-specific binding by MSY4 to the Y box recognition sequence (YRS) is dependent on the highly conserved cold shock domain, possibly through the RNP1 and RNP2 motifs present within it. The Y box proteins may function as translational repressors in vivo. Alternatively, their primary function may be to protect mRNAs from degradation during their extended period of storage. Translational activation of stored mRNAs is essential for the completion of gametogenesis. Proper translational activation of the Prm1 mRNA in elongated spermatids requires the cytoplasmic double-stranded RNA binding protein TARBP2. Tarbp2 is expressed at low levels in many cells but is expressed at robust levels in late stage meiotic cells and in postmeiotic spermatids. Mice mutant for Tarbp2 are defective in proper translational activation of the Prm1 and Prm2 mRNAs and are sterile. Current studies are designed to determine the mechanism by which proteins bound to the 3' UTR communicate with the 5' end of the message to control translational silencing and activation. PMID- 10849509 TI - Predicting coronary events with coronary calcium: pathophysiologic and clinical problems. PMID- 10849510 TI - Pediatric digital echocardiography: a study of the analog-to-digital transition. AB - Limited information is available that describes the practical conversion of a pediatric echocardiography laboratory from videotape to a primarily digital format. To help pediatric echocardiographers begin to make the analog-to-digital transition, we report our pediatric digital acquisition protocol and the acquisition and storage parameters of 1000 unselected, consecutive digitally acquired studies of pediatric patients with known or suspected congenital or acquired heart disease. With the use of our acquisition protocol, a complete normal study requires 46 moving clips and 12 still-frame images. Five hundred consecutive patient studies acquired with "high" JPEG (Joint Photographers Experts Group) compression (group 1) were compared with the next 500 examinations acquired using "medium" JPEG compression (group 2) for number of moving clips, still images, and megabytes of storage space. No intergroup difference was found in the number of moving clips or still images. When JPEG compression was decreased from high to medium, the average clip storage requirement per patient increased, and the number of patients stored per 230-MB magneto optical disk decreased significantly. Non-ECG-triggered timed single-plane clips and still images required significantly more storage space than ECG-triggered single-beat clips and still images. The frequency of multiplane sweeps was.03% and was independent of diagnosis. With the use of high JPEG compression, the digital storage cost per patient was $1.90, which was 6.0 times greater than that for simultaneously recorded 120-minute VHS videotape. Many features of the digital paradigm, including decreased MOD storage space, enhanced serial study comparisons, random image access, and improved image quality, mitigate this cost differential. PMID- 10849511 TI - Quantitative 3-dimensional contrast echocardiographic determination of myocardial mass at risk and residual infarct mass after reperfusion: experimental canine studies with intravenous contrast agent NC100100. AB - Two-dimensional contrast echocardiography has been shown to enable the evaluation of myocardial perfusion abnormalities. However, its ability to quantify a regional myocardial mass is limited. The goal of this study was to examine the quantitative value of 3-dimensional echocardiography (3DE) in the estimation of myocardial mass at risk, salvaged mass, and residual infarct mass after intravenous injection of contrast. We created acute coronary occlusion, followed by reperfusion in 10 dogs. Three-dimensional echocardiographic data were acquired at the end of each stage, and the perfusion defect mass and dysfunctional mass were measured. The true mass at risk and infarct mass were determined by anatomic methods. The anatomic mass at risk (x) (27.1+/-14.6 g or 23.8%+/-9.7% of the left ventricle [%LV]) correlated well with the 3DE-determined perfusion defect mass (y) during coronary occlusion (y = 0.5x+8.9; r = 0.90; P<.001; mean difference 4.8+/-8.1 g; or y = 0.7x + 6.5; r = 0.83, P<.01; mean difference -0.1+/-5.4 %LV). Good correlation was also found between the anatomic infarct mass (x) (9.3+/-8.1 g or 9.1+/-8.8 %LV) and the 3DE perfusion defect mass after reperfusion (y) (y = 1.2x+1.2; r = 0.93; P<.001; mean difference 2.3+/-4.0 g; or y = 1. 3x, r = 0.98, P <.0001; mean difference 2.7+/-3.7 %LV). The salvaged mass was 13.6 +/-11.0 %LV from anatomic methods and 14.2+/-13.0 %LV by 3DE. To conclude, with the use of intravenous contrast, 3DE could quantify the actual mass at risk during acute ischemia, and in the setting of reperfusion, the residual infarct mass and salvaged mass. PMID- 10849512 TI - Noninvasive assessment and necropsy validation of changes in left ventricular mass in ascending aortic banded mice. AB - Although left ventricular (LV) hypertrophy can be induced by aortic banding, noninvasive assessment of changes in LV mass in mice with a banded ascending aorta by using 2-dimensional (2D) images has not been previously performed. In this study we serially assessed changes in LV mass by 2D echocardiography with a newly available 12-MHz transducer in mice with a banded ascending aorta and validated measurements at necropsy. Estimated by echocardiography, LV mass increased from 74+/- 17 mg before banding to 191.08+/-54 mg at 8 weeks after banding (P <.0001), and excellent correlation was shown with postmortem measurements (r = 0.97). Furthermore, with the use of pulsed Doppler 2 dimensionally guided echocardiography, noninvasive measurement of flow velocities in the ascending aorta before and after the band at the various time points was possible. We propose that 2D echocardiography with a 12-MHz transducer is a powerful tool for serial noninvasive evaluations as an adjunct to the study of cardiac hypertrophy in the murine model. PMID- 10849513 TI - Assessment of regional longitudinal myocardial strain rate derived from doppler myocardial imaging indexes in normal and infarcted myocardium. AB - Myocardial deformation properties may be characterized by regional strain rates (SRs) calculated from Doppler myocardial velocity data. In 10 control subjects and 12 patients with established transmural infarcts, longitudinal median segmental SR, strain, and myocardial velocity were analyzed and compared with the corresponding wall motion score. All segments in control subjects and normal segments in infarct patients showed no significant difference in either systolic or diastolic SR (systolic: -1.27+/-0.39 s(-1) versus -1.23+/-0.24 s(-1), not significant [NS]; and isovolumic relaxation [IVR]: 1.23+/-0.38 s(-1) versus 1.95+/-0.62 s(-1), NS; respectively) and strain (-0.21+/-0.06 versus -0.19+/ 0.06, NS). In infarcted segments, peak systolic SR, systolic strain, and early diastolic SR showed the most pronounced reduction (hypokinetic and akinetic) or even inversion (dyskinetic segments: 0.10+/-0.26 s(-1), 0.00+/-0.03, and -1.78+/ 0.67 s(-1), respectively; P<.001). In this study, new myocardial deformation indexes were shown to quantitatively describe the function of normal and chronically infarcted regions. PMID- 10849514 TI - Left ventricular end-diastolic pressure can be estimated by either changes in transmitral inflow pattern during valsalva maneuver or analysis of pulmonary venous flow. AB - We directly compared the transmitral inflow pattern during preload reduction and pulmonary venous flow velocities to determine left ventricular end-diastolic pressure (LVEDP) in 78 patients who underwent left heart catheterization. Transmitral inflow indexes (A-wave duration, ratio of peak flow velocity of early diastole [E] to peak flow velocity of late diastole during atrial contraction [A] [E/A ratio]) at rest and during the Valsalva maneuver (30 mm Hg for 15 seconds) and indexes of pulmonary venous flow (velocity and duration of the atrial reversal) were obtained. Fair correlations existed between LVEDP (mean 15+/-6 mm Hg) and the percentage decrease in the E/A ratio (r = 0.72), increase in duration of A wave during the Valsalva maneuver (r = 0.60), flow velocity of atrial reversal (r = 0.58), and difference of duration of atrial flow reversal and A wave (r = 0.62) (all P<.001). While sensitivity, specificity, and diagnostic accuracy to detect an elevated LVEDP were comparable, technically adequate Doppler recordings were obtained more often for the mitral inflow during the Valsalva maneuver than for the pulmonary venous flow (72 versus 66 patients, P< 0.05). PMID- 10849515 TI - In vivo molecular imaging of stretch-induced tissue factor in carotid arteries with ligand-targeted nanoparticles. AB - Molecular imaging permits tissues to be functionally characterized by identification of specific cell-surface receptors with targeted contrast agents. In our study, a ligand-targeted acoustic nanoparticle system was used to identify the angioplasty-induced expression of tissue factor by smooth muscle cells within the tunica media. Pig carotid arteries were overstretched bilaterally with balloon catheters, treated with a tissue factor-targeted or a control nanoparticle system, and imaged with intravascular ultrasound (20 MHz) before and after treatment. Carotid wall acoustic reflectivities were unaffected by overstretch injury. Tissue factor-targeted nanoemulsion bound and increased the echogenicity of smooth muscle cells expressing tissue factor within the tunica media. The targeted emulsion increased the arterial wall gray scale (99.4+/-14.5; P<.05) relative to pretreatment (41.8+/-11.1, P<0.05) and the control gray scale (pre-emulsion: 49.3+/-9.5; post-emulsion: 43.7+/-6.4; P<.05). The area of acoustic enhancement appeared to coincide with expression of induced tissue factor in the tunica media confirmed by immunohistochemistry. We have demonstrated that this novel nanoemulsion can infiltrate into arterial walls after balloon injury and localize the expression of overstretch-induced tissue factor within pig carotid arteries. Molecular imaging and quantification of complex, biochemical change, such as tissue factor expression after angioplasty, may prove to be a prognostically important predictor of subsequent restenosis. PMID- 10849516 TI - Intraoperative myocardial ischemia recognized by transesophageal echocardiography monitoring in the pediatric population: a report of 3 cases. AB - We used continuous intraoperative transesophageal echocardiography (TEE) monitoring to detect intraoperative myocardial ischemia in children after they had been weaned from cardiopulmonary bypass for cardiac surgery. Three pediatric patients are described here to illustrate the usefulness of such TEE monitoring in surgical procedures involving coronary arteries. The indications for intraoperative TEE monitoring and a simplified scheme for immediate qualitative interpretation are discussed. PMID- 10849517 TI - Transesophageal echocardiography detection of an esophageal sarcoma mimicking aortic dissection. AB - This report shows that transesophageal echocardiography can detect thoracic pathology, in this case esophageal sarcoma, as well as cardiac and aortic abnormalities. Transesophageal echocardiography can help differentiate cardiac from aortic or other intrathoracic pathology when the patient's history and physical examination do not provide enough information. PMID- 10849518 TI - Right ventricular outflow tract obstruction in an older woman: facilitated diagnosis with transesophageal echocardiography-guided biopsy. AB - Fibrosarcoma is a rare primary cardiac malignancy. We report the case of a 70 year-old woman who had signs of right ventricular outflow tract obstruction caused by a fibrosarcoma. The pivotal role of multiplanar transesophageal echocardiography in characterizing masses in this location and in guiding transvenous biopsy is discussed. PMID- 10849519 TI - Myxoma on anterior mitral leaflet presenting with symptoms of cerebellar artery infarction. AB - This report describes a patient who had dizziness and loss of balance. During routine investigation, a mass located on the anterior mitral valve leaflet was detected on transthoracic echocardiography. The patient underwent surgery for a mass located on the mitral valve, and histopathologic examination determined the mass was a myxoma. PMID- 10849520 TI - Performing an echocardiographic examination with a contrast agent: a series on contrast echocardiography, article 2. PMID- 10849521 TI - Echocardiographic features of genetic diseases: part 6. Complex cardiovascular defects. PMID- 10849522 TI - Autologous blood cell vs marrow transplantation for acute myeloid leukemia in complete remission: an EBMT retrospective analysis. AB - In order to compare autologous bone marrow (BMT) and blood cell transplantation (BCT) in patients with acute myeloid leukemia (AML) in first remission (CR1), we retrospectively reviewed the data of 1393 patients registered to EBMT and undergoing either BCT (n = 100), purged (n = 252) or unpurged (n = 1041) BMT. Hematopoietic recovery was significantly quicker after BCT than after either purged or unpurged BMT. The 2-year leukemia-free survival (LFS), relapse incidence (RI) and overall survival for the entire population of patients were 52 +/- 1%, 43 +/- 1% and 58 +/- 1% and were significantly influenced by FAB subtype (M3 vs other) and the intervals between diagnosis and CR1 or CR1 and transplant. After BCT, LFS and RI were 44 +/- 6% and 50 +/- 6% and did not differ significantly from that found for unpurged BMT (49 +/- 2% and 45 +/- 2%; P = NS). However, LFS (57 +/- 3%) and RI (37 +/- 3%) of patients undergoing purged BMT were significantly different from that found for BCT patients (P = 0.01 and P = 0.006). As some characteristics of patients undergoing BCT or purged BMT differed significantly (age, intervals between diagnosis and CR1 or CR1 and transplant), the better outcome observed for purged BMT over BCT patients needs to be prospectively investigated. PMID- 10849523 TI - Roquinimex (Linomide) vs placebo in AML after autologous bone marrow transplantation. AB - Roquinimex, Linomide, a quinoline derivative with pleiotropic immunomodulatory activity, has previously been shown to enhance natural killer (NK) cell number and activity after ABMT in patients with AML. In this study 278 AML patients in remission were randomized to receive Roquinimex 0.2 mg/kg body weight or placebo twice weekly for 2 years following ABMT. Out of 139 patients in each group, 109 Roquinimex patients and 108 placebo patients were in their first CR. Median age at inclusion was 41 years for Roquinimex patients and 39 years for placebo patients. Twelve patients in each group had their marrow purged prior to reinfusion. Relapse and death were study endpoints. Surviving patients were followed for 2.6 to 6. 9 years. The total number of relapses was 60 in the Roquinimex group and 63 in the placebo group (not significant). Leukemia-free and overall survivals were similar in the two groups. Recovery of platelet counts was significantly delayed in the Roquinimex group as compared to placebo. No other significant differences regarding toxicity parameters were recorded. In conclusion, previous findings on NK cells could not be confirmed and the study showed no benefit for Roquinimex over placebo regarding relapse or survival following ABMT for AML in remission. PMID- 10849524 TI - A randomised study of allogeneic transplantation with stem cells from blood or bone marrow. AB - Sixty-one consecutive adult patients with leukaemia, primary myelofibrosis or myelodysplastic syndrome with an HLA-identical or one antigen mismatched family donor were randomised to allogeneic transplantation with PBPC or BM. Progenitor cells were mobilised into the blood by giving the donors 10 microg/kg/day G-CSF subcutaneously for 5-7 days. G-CSF was not given to patients after transplantation. The time to neutrophil counts >0.5 x 109/l was 17 days (95% CI 15.2-18.8 days) in the PBPC group compared to 23 (95% CI 20.3-25.7 days) in the BM group (P = 0.0005). The time to platelet counts >20 x 109/l was 13 days (95% CI 11.7-14.3 days) in the PBPC group and 21 days (95% CI 18.7-23.3 days) in the BM group (P = 0.0005). Acute GVHD of grades II-IV developed in six patients transplanted with PBPC and three patients transplanted with BM. The numbers of patients with chronic GVHD were 15 and 8, respectively. Transplant-related mortality and leukaemia-free survival showed no significant differences. Transplantation with PBPC appears preferable for the recipient due to faster neutrophil and platelet recovery. However, the final conclusion can not be drawn before long-term results on chronic GVHD and relapse incidence in longer randomised trials are available. PMID- 10849525 TI - High-dose ifosfamide and etoposide with filgrastim for stem cell mobilization in patients with advanced ovarian cancer. AB - High-dose chemotherapy combined with autologous peripheral blood stem cell transplantation has shown promise as treatment for recurrent or persistent epithelial ovarian cancer. We evaluated the stem cell mobilization regimen of high-dose ifosfamide plus etoposide in 32 patients with epithelial ovarian cancer, who had a positive second-look laparatomy or recurrent disease. Ifosfamide was given at 10 g/m2 by continuous i.v. from days 1 to 3. Etoposide was given at 150 mg/m2 every 12 h for six doses on days 1-3. Filgrastim was given at 10 microg/kg/d s.c. from day 5 until the completion of peripheral blood stem cell harvest. Fourteen of 32 patients had measurable or evaluable disease before mobilization therapy and were assessed for response. In nine (64%) of the 14 patients, treatment response was demonstrated, and these patients received a second cycle of mobilization therapy. The target CD34+ cell dose (>8 x 106 cells/kg) was achieved with a median of one apheresis (range 1-5). A median of 25.1 (range 8.0-122.5) x 106 CD34+ cells/kg body weight was collected. Non hematologic toxicity was limited to grade 2 renal dysfunction in one patient and grade 2 hepatic dysfunction in three patients. In this patient group, high-dose ifosfamide plus etoposide with filgrastim support was well tolerated, lead to successful stem cell harvest and had antitumor activity. PMID- 10849526 TI - Prospective randomized clinical trial comparing high-dose ifosfamide + GM-CSF vs high-dose cyclophosphamide + GM-CSF for blood progenitor cell mobilization. AB - Between August 1994 and June 1999, 56 patients were prospectively randomized to receive ifosfamide 10 g/m2 + GM-CSF 5 microg/kg/day (IFO+GM-CSF n = 28) and cyclophosphamide 4 g/m2 + GM-CSF 5 microg/kg/day (CY+GM-CSF n = 28). Both groups were comparable for age, gender, diagnosis, disease stage and previous chemotherapy. The IFO+GM-CSF group demonstrated a shorter median interval between therapy and apheresis (10 days (8-14) vs 13 days (8-25) P = 0.002), median number of doses of GM-CSF (9 (7-13) vs 15 (9-31) P = 0.001), median of days with aplasia (0.5 (0-10) vs 6 (0-21) P = 0.001), median days with fever (0 (0-6) vs 3 (0-9) P = 0.006) and median of days using i.v. antibiotics (0 (0-11) vs 7.5 (0-19) P = 0.002). The median MNC yield was similar in both groups. The CD34+ cell yield was better in the CY+GM-CSF group (3.14 (0.9-11.8) vs 5.33 (0. 08-32)) but not at significant levels (P = 0.1). White blood cell hematopoietic recovery was more rapid in the CY+GM-CSF group (16 (10-22) vs 13 (10-24) P = 0.02). Platelet engraftment was similar in both groups. Costs of mobilization and transplantation were almost the same: $28 570 ($18 527-$47 028) and $30 020 ($17 281-$67 591), respectively (P = 0.9). There were no differences in disease-free survival and overall survival between both groups. Mild and transient non-hematological toxicity (hemorrhagic cystitis, decrease in serum creatinine clearance and CNS dysfunction) was seen most frequently in the IFO+GM-CSF group. PMID- 10849527 TI - Mobilisation of haemopoietic progenitors in CML: a second course of intensive chemotherapy does not improve Ph-negativity in stem cell harvests. AB - We collected peripheral blood stem cells (PBSC) in 19 early chronic phase CML patients following each of two consecutive cycles of intensive chemotherapy (CT) to evaluate whether an additional cycle of CT would increase Philadelphia (Ph) negativity of the PBSC harvest. Autologous SCT (autoSCT) was performed if a major cytogenetic response (MCR) of the PBSC harvest was obtained. CT consisted of cytarabine 200 mg/ m2/day (days 1-7)/idarubicin 12 mg/m2/day (days 1-2) (cycle one) and cytarabine 2000 mg/m2/day (days 1-6)/amsacrine 120 mg/m2/day (days 1-3) (cycle two). One patient died of fungal pneumonia after the first cycle. Stem cells were harvested in 18 patients after cycle one and in 16 patients after cycle two. After the first cycle, all patients showed a cytogenetic response of their graft (MCR in eight patients: three complete, five partial), after cycle two, seven patients obtained an MCR (one complete, six partial). Seven patients became eligible for autoSCT. All patients proceeded with IFNalpha maintenance. Currently, 16 patients are alive. At the latest cytogenetic examination of bone marrow, four patients showed an MCR and four a minor response. In conclusion, although a second cycle of CT may contribute to elimination of leukemia residing in the patient, it appeared to be ineffective in improving the Ph-negativity of the PBSC graft. PMID- 10849529 TI - Transplantation of CD34+ peripheral blood cells selected using a fully automated immunomagnetic system in patients with high-risk breast cancer: results of a prospective randomized multicenter clinical trial. AB - Tumor contamination of autologous peripheral blood stem/progenitor cell grafts occurs in a substantial proportion of high-risk breast cancer patients, and the possibility that such contamination may contribute to relapse has focused attention on methods for removal of the contaminating cells prior to transplantation. One such approach is positive selection of CD34+ cells. A fully automated immunomagnetic cell selection system has recently been introduced to facilitate the positive selection process. A multicenter randomized clinical trial was designed to evaluate the capacity of CD34+ cells isolated using the fully automated system to support prompt hematopoietic reconstitution following high-dose chemotherapy in high-risk breast cancer patients, as well as to assess the safety and tolerability of the CD34+ cell transplants. In recipients of isolated CD34+ cells, the median time to an absolute neutrophil count > or =500/microl was 10 days, a value identical to that observed in patients receiving unfractionated apheresis collections. In the isolated CD34+ cell recipients median time to a platelet count > or =20 000/microl was 12 days, compared with 10 days in the unfractionated cell group. There were no statistically significant differences between the groups in median time to neutrophil or platelet engraftment. Infusion of autologous cells was well tolerated by the study groups. There were no inter-group differences in the incidence of infections, need for platelet transfusions, or duration of hospitalization. Isolated CD34+ cells were high in purity and sufficient in number for use in autologous transplantation. The fully automated immunomagnetic cell selection system affords an efficient and time-saving option for isolation of CD34+cells to be used as autologous grafts in high-risk breast cancer patients, and the isolated CD34+ cells support undelayed hematopoietic reconstitution. PMID- 10849528 TI - Enumeration of HPC in mobilized peripheral blood with the Sysmex SE9500 predicts final CD34+ cell yield in the apheresis collection. AB - Enumeration of CD34+ cells in the peripheral blood before apheresis predicts the quantity of those cells collected, although the cytometric techniques used are complex and expensive. We found that a subpopulation of lysis-resistant cells in the peripheral blood, identified by the Sysmex SE9500 and designated as HPC, can serve as a surrogate marker predictive of the yield of CD34+ cells. Spearman's rank statistics were used to examine the correlation between WBC, MNC, HPC and CD34+ cells in the peripheral blood and final CD34+ cell yield for 112 samples of peripheral blood and matching apheresis collections from 66 patients and donors. The results indicate that WBC and MNC in the peripheral blood were poor predictors of CD34 content, while HPC gave a correlation coefficient of 0.62. The positive predictive values of different cutoff levels of HPC in the peripheral blood ranging from 5 to 50 x 106/l increased from 0.80 to 0.93 when the target collection was 1 x 106cells/kg. However, for patients with HPC levels below various cutoff levels, the proportion of the collections not reaching that target goal ranged between 0.36 and 0.43, indicating that most collections will still exceed the target goal of CD34+ cells. When the target collection was 2.5 x 106 CD34+ cells/kg, the positive predictive value was lower and negative predictive value was higher. PMID- 10849530 TI - CD34+ selection of autologous peripheral blood stem cells for transplantation following sequential cycles of high-dose therapy and mobilization in multiple myeloma. AB - A potential problem of autologous transplantation in the treatment of multiple myeloma (MM) is the infusion of tumor cells. CD34+ selection has been used to purge autografts in MM and it is also possible to reduce tumour cell contamination of autografts by cytotoxic drug therapy prior to peripheral blood stem cell (PBSC) collection. To evaluate the effectiveness of a protocol combining multiple cycles of high-dose therapy and CD34+ selection to reduce tumour contamination of PBSC autografts, 34 MM patients were entered on a treatment schedule comprising two sequential cycles of mobilisation, CD34+ selection, and transplantation following high-dose therapy. In the second cycle of mobilisation there was a five-fold reduction in tumour contamination of the stem cell harvest (0.5 x 106/kg) compared with the first cycle (2.5 x 106/kg). In the 97 CD34+ selection procedures performed a median of 185 x 108 mononuclear cells (MNC) were processed yielding a median of 0.98 x 108 CD34+-enriched cells. CD34+ cells were enriched 68-fold from 1. 3% to 88.6%. The median yield of CD34+ cells was 42.2%. Following CD34+ selection the tumour cell contamination of the leukapheresis product was reduced by a median of 2.7 logs. This study demonstrates that in multiple myeloma a significant reduction in the malignant contamination of stem cell autografts can be achieved by combining the in vivo purging effect of cytotoxic therapy with in vitro purging by CD34+ selection. PMID- 10849531 TI - Cavernosal arterial insufficiency is a major component of erectile dysfunction in some recipients of high-dose chemotherapy/chemo-radiotherapy for haematological malignancies. AB - We studied 24 male patients aged 26-62 years (median 41) prospectively presenting over a 5 year period with clinical features of hypogonadism and erectile dysfunction (ED), who had been treated with autologous or allogeneic bone marrow/stem cell transplant for a variety of haematological malignancies and had received either high-dose chemotherapy or high-dose chemotherapy combined with total body irradiation (TBI). Ten healthy adult controls (aged 35-50 years) were also studied. Erectile dysfunction (ED) was assessed clinically and by colour flow Doppler studies of the cavernosal vessels. Testicular function was assessed by testicular volume including orchidometry, FSH, LH and testosterone measurements. Libido and ejaculatory function were also recorded. Patients had severe hypogonadism as evidenced by low mean testicular volume (7.0 +/- 2.4 ml vs 20 +/- 2.0 ml; P < 0.001), elevated gonadotrophins (FSH = 18.54 +/- 7.61 vs 5 IU/l (P < 0.001); LH = 8.02 +/- 2.89 vs 3. 9 IU/l (P < 0.001)) and low normal mean testosterone levels (16.4 nmol/l +/- 9.1 vs 22.4 nmol/l (P < 0.5)). Cavernosal arterial insufficiency was found in 11/14 of TBI-treated and in 3/10 HDC-treated patients, indicative of vasculogenic damage to corpora cavernosal vessels. Patients were given a therapeutic trial with testosterone replacement therapy (TRT). Those who had diminished libido had a marked improvement in their symptoms but the effect of TRT on ED was equivocal. In conclusion, this is the first report to show vasculogenic insufficiency in patients with haematological malignancies treated by BMT. Although hypogonadism can account for diminished libido, arteriogenic insufficiency is likely to be an important factor accounting for ED in these patients, especially those treated by TBI. We recommend a comprehensive assessment including endocrine profile and colour flow Doppler study in formulating the best management plan in recipients of high-dose therapy presenting after transplant with ED. PMID- 10849532 TI - Validation of self-reported complications by bone marrow transplantation survivors. AB - Self-administered questionnaires are commonly used to measure exposures and outcomes in epidemiological research and thus need good validity. With increasing numbers of cancer survivors, there is interest in the ongoing assessment of therapy-related complications. A medical record validation of patient-reported complications following bone marrow transplantation (BMT) was performed using a self-administered questionnaire. The study population consisted of 100 patients who had undergone BMT at the City of Hope. The following self-reported complications were validated using medical records: ocular, endocrine, cardiovascular, musculoskeletal, pulmonary, gastrointestinal, neurological, graft versus-host disease, and subsequent cancers. Using information from medical records as the standard, sensitivities ranged from 52.9% for subsequent cancers to 100% for avascular necrosis and hypothyroidism. Specificities ranged from 75.4% for ocular complications to 100% for avascular necrosis. There was intermediate to excellent agreement (kappa = 0. 4-1.0) for all complications evaluated. Thus, the agreement between self-reporting and medical records was good for complications with clear diagnostic criteria that are easily communicated to the patient, but was diminished for complications with non established diagnostic criteria (xerophthalmia) or a fluctuating course (peripheral neuropathies and hypertension). Overall these results suggest that cancer survivors can self-report serious complications with an acceptable level of accuracy in epidemiological research. PMID- 10849533 TI - Adverse side-effects associated with G-CSF in patients with chronic myeloid leukemia undergoing allogeneic peripheral blood stem cell transplantation. AB - Administration of the myeloid growth factor G-CSF after allogeneic hematopoietic stem cell transplantation is usually well tolerated, and associated with rapid hematopoietic engraftment. We report a high incidence (50%) of side-effects associated with post-transplant G-CSF in patients with chronic phase chronic myeloid leukemia undergoing allogeneic HLA-identical sibling peripheral blood stem cell transplantation. One or more of the following signs and symptoms were observed shortly after the subcutaneous injection of G-CSF: dyspnea, chest pain, nausea, hypoxemia, diaphoresis, anaphylaxis, syncope and flushing. These reactions led to discontinuation of G-CSF in the majority of patients. Predictive factors could not be identified, and the underlying mechanism leading to these reactions is unknown. PMID- 10849534 TI - Chromosomal abnormalities in women with breast cancer after autologous stem cell transplantation are infrequent and may not predict development of therapy-related leukemia or myelodysplastic syndrome. AB - We determined prospectively the incidence of chromosomal abnormalities in patients with high-risk breast cancer (HRBC) after high-dose chemotherapy (HDCT) and autologous stem cell transplantation (ASCT), and correlated the cytogenetic abnormalities with the development of post-transplant myelodysplastic syndrome or acute myeloid leukemia (MDS/AML). From 1990 to 1999, 229 women with HRBC underwent ASCT. Cytogenetic analysis of bone marrow (BM) cells was performed 12 59 months after ASCT in 60 consecutive women uniformly treated with six courses of FAC/FEC followed by HDCT and ASCT. With a median follow-up of 36 months after ASCT, there were no cases of MDS/AML among the 229 patients. In the selected cohort of 60 patients, three (5%) showed clonal chromosomal abnormalities (two single trisomy X and one t(1;6)), whereas two additional patients showed non clonal reciprocal translocations. Two of the patients with clonal aberrations had blood cytopenias as well as subtle dysplastic pictures in BM which were not classifiable as MDS according to the FAB criteria. Similar dysplastic features were also observed in four patients with normal karyotypes. All cytogenetic aberrations were transient and disappeared, except a +X detected by FISH in a residual cell population in one of the patients. Retrospective cytogenetic and FISH studies of samples obtained after six cycles of FAC/FEC and before transplant demonstrated no chromosomal abnormalities in any of the five patients with post-ASCT karyotypic changes. Early changes in karyotype detected in breast cancer patients following ASCT are transient and do not correlate with or predict development of MDS/AML. As these aberrations were not present before ASCT, they may be related to the HDCT regimen or transplant procedure rather than to the prior adjuvant therapy. Our results suggest that ASCT may be less likely to cause MDS or AML in breast cancer patients as compared to other malignancies. Bone Marrow Transplantation (2000) 25, 1203-1208. PMID- 10849535 TI - A Philadelphia chromosome positive acute lymphoblastic leukemia of donor origin after allogeneic bone marrow transplantation for chronic myelogenous leukemia in chronic phase. AB - We report here a case of donor cell leukemia in a female Ph-positive CML patient who received an allogeneic BMT from her HLA-identical brother in the chronic phase and subsequently developed a donor cell Ph-positive ALL. The number of cases of donor cell leukemia after BMT so far reported is less than 20 and in this case, as in the first cases reported by Marmont et aland McCane et al, the original leukemia and donor cell leukemia share the presence of a Ph chromosome. Furthermore, we analyzed the patient during different stages of her disease by RT PCR and determined the type of bcr-abl junctions (M bcr-abl junction; b3a2 transcript, p210) in both the recipient and donor cell leukemia. PMID- 10849536 TI - Cunninghamella infection post bone marrow transplant: case report and review of the literature. AB - Cunninghamella spp., in the class Zygomycete and order Mucorales, are unusual opportunistic pathogens that have been identified with increased frequency in immunocompromised patients. Infections with this group of organisms have been seen most frequently in patients with hematologic malignancy. We describe an allogeneic bone marrow recipient who developed fungal pneumonitis and disseminated fungal dermatitis caused by Cunninghamella spp. To our knowledge, this is the first reported case of Cunninghamella infection in a BMT recipient. The case highlights the mortality associated with opportunistic infections in immunocompromised patients and confirms the risk factors associated with non candida fungal infections after bone marrow transplantation. PMID- 10849538 TI - Response PMID- 10849537 TI - Prompt response to high-dose intravenous immunoglobulins given as first-line therapy in post-transplant thrombotic thrombocytopenic purpura. PMID- 10849539 TI - Imaging of malignant cervical lymphadenopathy. AB - Diagnostic imaging is superior to clinical staging in the detection of malignant cervical lymphadenopathy, and thus helps influence therapy and prognosis. The imaging modalities of CT, MRI, US and PET each have their own diagnostic criteria, accuracy and limitations. Newer innovations such as functional imaging, novel MRI contrast agents and FNAC are being appraised with the aim of identifying the micrometastases which are currently radiologically occult. PMID- 10849540 TI - Fractal properties of bone. AB - OBJECTIVE: To introduce the applications of fractal analysis of bone in medical imaging in general and dental radiographs in particular. RESULTS: Various methods for measuring fractal dimension have been used to compare normal with osteoporotic bone with contradictory results. This disparity may be attributed to differences in the anatomical sites studied and differences in methods used to obtain the 2D images. However, differences in methods used to measure fractal dimension can also be responsible. CONCLUSION: When fractal dimension is used to study bone it must be remembered that all stages in the analytical chain have an impact on the results. Thus, to obtain more conclusive results, studies on the fractal dimension of bone should be carefully designed and individual methods thoroughly evaluated. PMID- 10849541 TI - Assessment of maxillary sinus volume for the sinus lift operation by three dimensional magnetic resonance imaging. AB - OBJECTIVES: To calculate sinus and bone graft volumes and vertical bone heights from sequential magnetic resonance imaging (MRI) examinations in patients undergoing a sinus lift operation. METHODS: MRI scans were obtained pre operatively and at 10 days and 10 weeks post-operatively, using a 0.95 tesla MRI scanner and a three-dimensional (3D) magnetisation prepared, rapid acquisition gradient-echo (MP-RAGE) sequence. RESULTS: Estimates of the bone graft volumes required for a desired vertical bone height were made from the pre-operative MRI scan. Measurements of the graft volumes and bone heights actually achieved were made from the post-operative scans. The MRI appearance of the graft changed between the 10 day and 10 week scans. CONCLUSIONS: We have proposed a technique which has the potential to give the surgeon an estimate of the optimum volume of graft for the sinus lift operation from the pre-operative MRI scan alone and demonstrated its application in a single patient. Changes in the sequential MRI appearance of the graft are consistent with replacement of fluid by a matrix of trabecular bone. PMID- 10849542 TI - Is the morphology of the articular eminence of the temporomandibular joint a predisposing factor for disc displacement? AB - OBJECTIVES: To test the hypothesis that the morphology of the articular eminence of the TMJ is a predisposing factor for disc displacement. METHODS: MR images of 220 TM joints in 151 patients were studied: 47 joints had no disk displacement (NDD), 81 joints disc displacement with reduction (DDWR) and 92 joints disc displacement without reduction (DDWOR). The shape of the articular eminence of the TMJ was classified into four types, box, sigmoid, flattened and deformed. The prevalence of the box and flattened forms in the three groups of joints was compared. RESULTS: There was no statistically significant difference in the prevalence of the box form between the joints with NDD, DDWR, and DDWOR. On the other hand, the prevalence of the flattened-type was statistically lower in the joints with DDWR (goodness test of fit for chi 2, P < 0.05). CONCLUSIONS: It appears that disc displacement is less likely to be found in joints with a shallow articular eminence. PMID- 10849543 TI - A study of the association between the prognosis of carcinoma of the mandibular gingiva and the pattern of bone destruction on computed tomography. AB - OBJECTIVE: To clarify whether the pattern of bone destruction seen on CT is more closely associated with the outcome of carcinoma of the mandibular gingiva than that derived from panoramic radiographs (PR). METHOD: Axial bone window CT scans and PR of 62 patients with carcinoma of the mandibular gingiva were evaluated retrospectively by two oral radiologists for the pattern of bone destruction. Patterns were classified into the three types: erosive, invasive and mixed. The relationship between these patterns with each imaging modality and cumulative recurrence rate, cumulative metastasis rate and cumulative survival rate, calculated by the Kaplan-Meier method, were statistically analysed by the log rank test. RESULTS: The pattern of bone destruction derived from CT was closely associated with the cumulative metastasis rate (P < 0.05), the cumulative recurrence rate and the cumulative survival rate. In contrast, the pattern of bone destruction based on the PR was not associated with the cumulative metastasis rate (P = 0.43), the cumulative recurrence rate (P = 0.44), or the cumulative survival rate (P = 0.5). CONCLUSION: The prognosis of patients with carcinoma of the mandibular gingiva is more closely related to a classification derived from the pattern of bone destruction on CT rather than PR. However, the number of subjects investigated in this study was not large enough to confirm our conclusions statistically. Further studies by other investigators are therefore needed. PMID- 10849544 TI - Bilateral mandibular accessory foramina and canals: a case report and review of the literature. AB - OBJECTIVES: To present a case of bilateral mandibular accessory canals and foramina observed on CT and review the relevant literature. RESULTS: Based on the literature, accessory canals and foramina are prevalent in the posterior mandible and the area of the symphysis and more frequently on the internal than the external surface of the mandible. Bilateral symmetry is common. Variations exist in size and number. Occurrence may change with age and racial origin. Nerves, neurovascular bundles, arterioles and venules have been found to occupy the accessory canals and foramina. No gender differences have been described. CONCLUSION: CT has advantages over two-dimensional radiography in identification of anatomical variations in the mandible. PMID- 10849545 TI - The contribution by medical radiology departments to dental radiology in general dental practice in Scotland. AB - AIM: To determine the availability of the services of medical radiology departments to general dental practitioners in Scotland. METHODS: Fifty-seven hospitals were identified as likely to have services available to general dental practitioners; 41 were within central urbanised areas (the 'Central Belt') and 16 in the more remote rural areas, (the 'Borders, Highlands and Islands'). The available services were identified by questionnaire. RESULTS: All 57 questionnaires were returned. Although there were significantly fewer larger hospitals in the 'Borders, Highlands and Islands', there was no significant difference in availability of services to general dental practitioners between the two parts of Scotland. CONCLUSION: The services of medical radiology departments are generally available to general dental practitioners in both the 'Central Belt' and the 'Borders, Highlands and Islands'. PMID- 10849546 TI - Radiological findings in an unusual osteosarcoma in the maxilla. AB - An unusual case of osteosarcoma of the maxilla, which was initially diagnosed as fibrous dysplasia on the basis of the clinical CT and histopathological findings, is presented. However, panoramic and periapical radiography suggested a malignant neoplasm. After surgery, the tumor was diagnosed histopathologically as a low grade osteosarcoma. We conclude that panoramic and periapical radiographs are important adjuncts and should be included in any investigation of the jaws where CT scanning is unable to differentiate between osteosarcoma and fibrous dysplasia. PMID- 10849547 TI - Parotid lymph node metastasis from adenocarcinoma of the urachus. AB - The parotid gland and its lymph nodes are frequent sites of metastases from head and neck cancers. However, metastasis from a distant primary below the clavicle is unusual. These originate from a variety of sites, most commonly the lung, kidney and breast. A case of a 59-year-old woman with parotid lymph node metastasis from an adenocarcinoma of the urachus, diagnosed on the basis of two discrete periparotid masses on CT and the patient's history, is presented. PMID- 10849548 TI - Profiling of genes which are differentially expressed in mouse liver in response to adenoviral vectors and delivered genes. AB - The effects of transgene delivery by adenoviral vectors were studied by probing a 588 gene, mouse cDNA array with mRNA derived from infected liver. The liver tissues were obtained from naive mice and mice infected with replication deficient adenovirus, adenovirus expressing transforming growth factor beta1 (TGFbeta1), and adenovirus expressing connective tissue growth factor (CTGF). Expression of 98 genes was detected in the array analysis. The increased expression of the transcripts for Stat1, gamma interferon-induced monokine (MIG) and interferon regulatory factor 1 (IRF1) clearly demonstrated the immune response induced by infection with a first generation, replication-incompetent adenovirus. In vivo expression of TGFbeta1 led to a down-regulation of genes involved in the immune response. The increased expression of u-PAR1, laminin receptor and BMP-1 confirms the importance of CTGF and TGFbeta1 in angiogenesis, and tissue repair. Expression of the serine protease inhibitors, Spi 2.4 and Spi 2, is also increased in response to AdTGFbeta1 and AdCTGF. PMID- 10849549 TI - Production and concentration of pseudotyped HIV-1-based gene transfer vectors. AB - Strategies to generate highly concentrated HIV-1 vector pseudotypes involving different envelope (Env) proteins including the vesicular stomatitis virus (VSV) G glycoprotein, the Moloney murine leukemia virus (MLV) 4070A amphotropic Env and the rabies G glycoprotein were established. Virus stocks were prepared by transient transfection using standard cell culture media or serum-free media. Such stocks were concentrated 50- to 300-fold by ultracentrifugation or by ultrafiltration using Centricon Plus-80 units yielding titers of up to 109transducing units per milliliter. There was no loss in titer with any of the pseudotypes tested. Thus, like lentiviral vectors pseudotyped with VSV-G, HIV-1 based vectors pseudotyped with the MLV 4070A amphotropic Env and the rabies G glycoprotein resist inactivation during concentration. This opens up the possibility to generate highly concentrated HIV-1 vector stocks carrying alternative Env proteins on a large scale. PMID- 10849550 TI - Low-speed centrifugation of retroviral vectors absorbed to a particulate substrate: a highly effective means of enhancing retroviral titre. AB - For many gene therapy applications the effective titre of retroviral vectors is a limiting factor both in vitro and in vivo. Purification and concentration of retrovirus from packaging cell supernatant can overcome this problem. To this end we have investigated a novel procedure which involves complexing retrovirus to a dense and particulate substrate followed by a short low-speed centrifugation. The study reported here uses heat-killed, formaldehyde fixed Staphylococcus aureus (Pansorbin) absorbed to PG13 derived retrovirus. This complex was then used to harvest retrovirus from packaging cell supernatant: centrifugation and washing of this complex allows the retrovirus to be both purified and concentrated. This procedure increases the effective titre of retrovirus by up to 7500-fold after an only 200-fold reduction in volume. The affinity of Pansorbin for retrovirus allows concentration regardless of its encoded genes and makes this protocol applicable to other popular packaging cells and envelope proteins. Possible explanations for the marked increase in titre of concentrated virus and the mechanism governing the complexing of retrovirus to Pansorbin are discussed. PMID- 10849551 TI - High throughput production, screening and analysis of adeno-associated viral vectors. AB - Recombinant adeno-associated viruses (rAAV) are promising candidates as gene vectors, as they transduce non-dividing cells and permit lasting transgene expression in a wide spectrum of tissues. In this paper, we describe a robust procedure for the high throughput production, screening and characterization of rAAV vectors. The technology includes the production of rAAV from rapid small scale plasmid preparations and the analysis of virus productivity (physical and infectious particles) and activity (transgene expression, replication). rAAV are produced by triple transfection (rAAV plasmid and AAV- and adenovirus (Ad)-helper plasmids) on 293 human embryo kidney (HEK) cells. The titers of physical and infectious particles are obtained by dot blot hybridization and by a serial dilution assay, followed by either dot blot hybridization or real-time PCR, respectively. rAAV can be produced and characterized from plasmid mixtures containing as little as 1/100 productive molecules. Experiments on rAAV replication kinetics and Ad helper functions are discussed. All steps are performed in 96-well microtiter plates. The process is reproducible, high throughput, linear and ready for automation. PMID- 10849552 TI - Adenovirus-p53-mediated gene therapy of anaplastic large cell lymphoma with t(2;5) in a nude mouse model. AB - Adenovirus-p53-mediated apoptosis has been extensively evaluated in animal xenografts derived from human epithelial tumors and recently began testing in phase I clinical trials, but has not been evaluated for lymphoid malignancies. Cell lines derived from anaplastic large cell lymphoma (ALCL) carrying the t(2;5) translocation are efficiently transduced by adenoviral vector expressing p53 and undergo apoptosis. To test the in vivo efficiency of adenovirus-mediated-p53 expression and apoptosis induction, SUDHL-1 cells (derived from human ALCL) were injected subcutaneously into athymic nude mice. Cells from the xenograft had typical morphology of human ALCL by standard hematoxylin-eosin staining, CD5+, CD45+ and CD30+ immunophenotype, the t(2;5) translocation by PCR. Six tumors from an initial set of mice were evaluated for apoptosis by TUNEL and for necrosis by hematoxylin-eosin staining 48-72 h after injection with 1 x 108 p.f.u. of AdWTp53 (adenoviral vector expressing p53), of AdNull (adenoviral vector backbone) and PBS (mock), respectively. TUNEL staining was positive only in tumors injected with AdWTp53 and was mainly localized around the needle track. Differences of the means of the counts of the necrotic cells were statistically significant at P = 0.02 between AdWTp53 and mock and only borderline between AdWTp53 and AdNull. Twenty-three tumors from a separate set of mice were subsequently injected with AdWTp53, AdNull and PBS and evaluated for in vivo tumor response. Three total injections of viral vectors (1 x 108 p.f.u.) and PBS were given every 48-72 h. Only tumors injected with AdWTp53 showed tumor growth inhibition with a mean final tumor volume that was statistically significantly smaller than AdNull (P = 0.007) and mock (P = 0.002). Based on these results we foresee a potential application of adenovirus-mediated p53 apoptosis as gene therapy of lymphomas. PMID- 10849553 TI - Long-term gene expression in the anterior horn motor neurons after intramuscular inoculation of a live herpes simplex virus vector. AB - To clarify the feasibility of the herpes simplex virus (HSV) vector in expressing the foreign gene in the motor neuron, we inoculated a live attenuated HSV expressing beta-galactosidase (beta-gal) activity under a latency-associated transcript promoter in the right gastrocnemius muscle of rats. Expression of beta gal activity was observed 5 days after inoculation in the bilateral anterior horn cells of the spinal cord that innervates the inoculation muscle. However, the spread of beta-gal activity was not observed in the inoculation muscle. Without significant pathological changes, the spread of beta-gal-expressing neurons was observed in the lumbosacral spinal cord until 14 days after inoculation with staining concentrated in the anterior horn cells. Ninety percent of the anterior horn motor neurons expressed beta-gal activity with expression continuing to at least 182 days after inoculation. Thus beta-gal activity was expressed in the bilateral anterior horn cells at the lumbosacral spinal cord that innervates the inoculated muscle for a long time, possibly a life-long period. This indicates that this recombinant HSV vector system to motor neurons may further improve the understanding and treatment of neurological diseases in motor neurons of the spinal cord. PMID- 10849554 TI - Adenoviral gene transfer of basic fibroblast growth factor promotes angiogenesis in rat brain. AB - Cerebral ischemic disease often causes morbidity and mortality, while the induction of new blood vessels is expected to provide a therapeutic effect in this occlusive cerebrovascular disease. In this study, we utilized two replication-deficient adenoviral vectors containing cDNA from basic fibroblast growth factor (bFGF), a well-known angiogenic factor, and examined whether biological angiogenic activity of adenovirally gene-transferred bFGF could be observed in the rat brain. One vector contained native cDNA from bFGF without the secretory signal sequence and the other contained the same cDNA fused with an interleukin-2 secretory signal sequence. After ventricular administration of these viral vectors, gene-transferred cells demonstrated a high immunoreactivity against the anti-bFGF antibody and a remarkably high concentration of bFGF was detected in the cerebrospinal fluid. A semiquantitative analysis of angiogenic activity revealed that bFGF gene transfer induced angiogenesis in normal rat brains, with a more pronounced angiogenic effect seen with the vector of a secreted form than with the vector without a secretory signal sequence. These results suggest that bFGF gene transfer using these adenoviral vectors might be useful for the treatment of ischemic cerebrovascular disease. PMID- 10849555 TI - Gene transfer of human interferon gamma complementary DNA into a renal cell carcinoma line enhances MHC-restricted cytotoxic T lymphocyte recognition but suppresses non-MHC-restricted effector cell activity. AB - Even though renal cell carcinomas (RCC) are thought to be immunogenic, many tumors express variations in surface molecules and intracellular proteins that hinder induction of optimal antitumor responses. Interferon gamma (IFNgamma) stimulation can correct some of these deficiencies. Therefore, we introduced the complementary DNA (cDNA) encoding human IFNgamma into a well-characterized RCC line that has been selected for development of an allogeneic tumor cell vaccine for treatment of patients with metastatic disease. Studies were performed to determine how endogenous IFNgamma expression influences tumor cell immunogenicity. IFNgamma transductants showed minimal increases in surface expression of MHC class I and adhesion molecules but expression of class II molecules was induced. Proteins of the transporter associated with antigen processing (TAP) and low molecular weight polypeptide (LMP) were constitutively expressed at high levels. The transductants stimulated allospecific cytotoxic T lymphocytes (CTL); however, they were not better than unmodified tumor cells in this capacity. Endogenous IFNgamma expression enhanced tumor cell recognition by MHC-restricted, tumor antigen-specific CTL but suppressed recognition by non-MHC restricted cytotoxic cells. Thus, the functional consequences of IFNgamma expression varied with respect to the type of effector cell and were not always beneficial for tumor cell recognition. PMID- 10849556 TI - Pretreatment with cationic lipid-mediated transfer of the Na+K+-ATPase pump in a mouse model in vivo augments resolution of high permeability pulmonary oedema. AB - Resolution of pulmonary oedema is mediated by active absorption of liquid across the alveolar epithelium. A key component of this process is the sodium-potassium ATPase (Na+K+-ATPase) enzyme located on the basolateral surface of epithelial cells and up-regulated during oedema resolution. We hypothesised that lung liquid clearance could be further up-regulated by lipid-mediated transfer and expression of exogenous Na+K+-ATPase cDNA. We demonstrate proof of this principle in a model of high permeability pulmonary oedema induced by intraperitoneal injection of thiourea (2.5 mg/kg) in C57/BL6 mice. Pretreatment of mice (24 h before thiourea) by nasal sniffing of cationic liposome (lipid #67)-DNA complexes encoding the alpha and beta subunits of Na+K+-ATPase (160 microg per mouse), significantly (P<0.01) decreased the wet:dry weight ratios measured 2 h after thiourea injection compared with control animals, pretreated with an equivalent dose of an irrelevant gene. Whole lung Na+K+-ATPase activity was significantly (P<0.05) increased in mice pretreated with Na+K+-ATPase cDNA compared both with untreated control animals as well as animals pretreated with the irrelevant gene. Nested RT PCR on whole lung homogenates confirmed gene transfer by detection of vector specific mRNA in three of four mice studied 24 h after gene transfer. This demonstration of a significant reduction in pulmonary oedema following in vivo gene transfer raises the possibility of gene therapy as a novel, localised approach for pulmonary oedema in clinical settings such as ARDS and lung transplantation. PMID- 10849557 TI - Amelioration of established collagen induced arthritis by systemic IL-10 gene delivery. AB - A novel formulation of cationic liposomes containing the novel cytofectin ACHx was used for delivery of an anti-inflammatory cytokine gene, IL-10, to mice with established collagen induced arthritis. A single intraperitoneal injection of human IL-10 expression plasmid complexed with liposomes 2 to 4 days after the onset of arthritis was sufficient to give significant and prolonged amelioration of arthritis for 30 days. Preliminary experiments suggested that the therapeutic effect was IL-10 dose-dependent. The distribution of the human IL-10 DNA after injection was widespread, including the inflamed paws. Human IL-10 mRNA was also detected in the paws 24 h after injection. IL-10 protein was below the level of detection in paws and serum but was detected in some tissues up to 10 days after injection. The target cell of transfection was demonstrated to be the macrophage. These results suggest that systemic therapy with plasmid DNA complexed with cationic liposomes merits further development as an alternative method for anti inflammatory treatment of arthritis. PMID- 10849558 TI - Experimental subretinal neovascularization is inhibited by adenovirus-mediated soluble VEGF/flt-1 receptor gene transfection: a role of VEGF and possible treatment for SRN in age-related macular degeneration. AB - Accumulating evidence has shown the importance of vascular endothelial growth factor (VEGF) in chorioretinal angiogenesis. However, whether or not VEGF is indeed critical for the pathogenesis of subretinal neovascularization (SRN) in adulthood, which is a serious complication of age-related macular degeneration, has to be further investigated. We constructed an adenovirus expressing an entire ectodomain of the human VEGF receptor/flt-1 fused to Fc portion of human IgG (Adflt-ExR): this soluble receptor is secreted from Adflt-ExR-transfected cells. We studied the effect of Adflt-ExR on the formation of experimental SRN. Experimental SRN was induced by intense photocoagulation on the retina in pigmented rats and Adflt-ExR was injected into the femoral muscle. The formation of SRN assessed by fluorescein angiography was more significantly inhibited for 7 days in the Adflt-ExR-treated rats than in the control rats who received either an adenovirus vector encoding LacZ gene or balanced salt solution (BSS). The serum concentration of this soluble receptor increased for 7 days and thereafter gradually decreased. An immunohistochemical study disclosed the fibroblast cell proliferation and inflammatory cell infiltration to be reduced in the photocoagulation spot of Adflt-ExR-treated rats. VEGF plays a crucial role in the formation of SRN and VEGF soluble receptor gene transfection can inhibit SRN. This method will contribute to future gene therapy for age-related macular degeneration. PMID- 10849559 TI - A combination of poloxamers increases gene expression of plasmid DNA in skeletal muscle. AB - Intramuscular administration of plasmid DNA is a promising strategy to express therapeutic genes, however, it is limited by a relatively low level of gene expression. We report here that a non-ionic carrier, SP1017, composed of two amphiphilic block copolymers, pluronics L61 and F127, also known as poloxamers, significantly increases intramuscular expression of plasmid DNA. Two reporter genes, luciferase and beta-galactosidase, and one therapeutic gene, erythropoietin, were injected intramuscularly with and without SP1017 into C57Bl/6 and Balb/C mice and Sprague-Dawley rats. SP1017 increased gene expression by about 10-fold and maintained higher gene expression compared with naked DNA. Comparison of SP1017 with polyvinyl pyrrolidone (PVP) showed that SP1017 exhibited a significantly higher efficacy and its optimal dose was 500-fold lower. Experiments with beta-galactosidase using X-gal staining suggested that SP1017 considerably increased plasmid DNA diffusion through the tissue. SP1017 also improved expression of the erythropoietin gene leading to an increase in its systemic level and hematocrits. Previous toxicity studies have suggested that SP1017 has over a 1000-fold safety margin. Poloxamers used in SP1017 are listed in the US Pharmacopeia as inactive excipients and are widely used in a variety of clinical applications. We believe that the described system constitutes a simple and efficient gene transfer method to achieve local or systemic production of therapeutic proteins. PMID- 10849560 TI - Sympathetic pathways and adrenergic innervation of the penis. AB - The sympathetic nervous system is important for penile function: it mediates detumescence and may contribute to the maintenance of the penis in a non-erect state. The sympathetic preganglionic neurons are found in the intermediolateral gray matter of the spinal cord. Postganglionic neurons are located to the sympathetic chain ganglia, the inferior mesenteric, hypogastric and pelvic ganglia, and possibly to ganglia near the target organ. Sympathetic fibres can be found in the pelvic, cavernous, and pudendal nerves. Stimulation of the sympathetic pathways to the penis may, however, also produce erection. It has been suggested that the suprasacral vasodilator pathway is a sympathetic cholinergic pathway, operating through cholinergic neurons in the pelvic plexus. In the penis, there is a rich sympathetic, adrenergic innervation of the corpus cavernosum (CC) and the vasculature, and in particular of the helicine arteries. Sympathetic, adrenergic nerves also contain neuropeptide Y. Parasympathetic cholinergic nerves, which mediate CC relaxation and erection, contain not only acetylcholine, but also vasoactive intestinal polypeptide, nitric oxide synthase, and probably other mediators and/or mediator-synthesizing enzymes. Activation of sympathetic adrenergic nerves causes release of noradrenaline, acting on alpha adrenoceptors in the trabecular smooth muscle of the CC and in penile vessels. The role of interactions between different transmitters and mediators, released from nerves or generated locally, in the regulation of contraction and relaxation of CC and penile vessels, needs further study. International Journal of Impotence Research (2000) 12, Suppl 1, S5-S12 PMID- 10849561 TI - Central noradrenergic control of penile erection. AB - Penile erection is completely dependent on commands from the central nervous system. Spinal centers controlling penile erection are located in the thoracolumbar and lumbosacral spinal cord. These centers are activated by information from the periphery and supraspinal nuclei so as to elicit penile erection in a variety of physiological contexts. A small number of nuclei including the locus coeruleus located in the pons sends noradrenergic fibers to the forebrain and spinal cord, including those areas controlling penile erection. Recent morphological techniques such as in situ hybridization and autoradiography using radioligand binding permit investigation of the brain and spinal pathways utilizing alpha adrenoceptor subtypes. Furthermore, pharmacological experiments suggest a modulatory role for noradrenaline in the control of penile erection either in the brain or in the spinal cord. The most robust evidence is that central inhibition of alpha-2 adrenoceptors facilitates sexual function. Taken together, the data propose new directions in the physiological exploration of penile erection and the therapeutic approach of erectile dysfunction. International Journal of Impotence Research (2000) 12, Suppl 1, S13-S19 PMID- 10849564 TI - Abanoquil, a new alpha-1 adrenoceptor antagonist. In vitro and in vivo effect on erectile tissue. AB - Using an organ bath model with porcine cavernosal tissue strips and an in vivo monkey model we demonstrated that abanoquil, a novel alpha adrenoceptor antagonist, is able to relax contracted tissue strips and induce erectile response when injected intracorporally. The erectile response in the monkeys was not dose-related and compared to the effect of papaverine injections, abanoquil induced a lower level of tumescence and rigidity. Hence, abanoquil might be useful as a facilitator of erection in the pharmacological treatment of erectile dysfunction. International Journal of Impotence Research (2000) 12, Suppl 1, S37 S40 PMID- 10849565 TI - Cyclic AMP regulates mRNA expression of alpha-1d and alpha-2a adrenergic receptors in cultured human corpus cavernosum smooth muscle cells. AB - While the physiological effects of contractile (e.g. norepinephrine) and relaxatory (e.g. PGE1, forskolin) agents on corpus cavernosum smooth muscle tone have been characterized, the regulation of alpha adrenergic receptor mRNA expression in erectile tissue remains to be investigated. The goal of this study was to investigate the modulation of alpha-1 and alpha-2 adrenergic receptor mRNA expression in cultured human corpus cavernosum smooth muscle cells in response to increased intracellular cAMP induced by prostaglandin E1 and forskolin. Human corpus cavernosum smooth muscle cells were incubated for 24 h with or without PGE1 (5.7 uM), forskolin (10 uM) or an admixture of both. Total RNA was prepared from the cultures. Expression of alpha-1d adrenergic receptor, alpha-2a adrenergic receptor and m2 muscarinic acetylcholine receptor was determined by RNase protection assays. Loading was normalized by RNase protection of the housekeeping gene, cyclophilin A. The relative abundance of mRNAs was quantitated by scanning densitometry. Treatment of human corpus cavernosum smooth muscle cells with PGE1 or forskolin resulted in decreased mRNA expression of alpha-1d and alpha-2a adrenergic receptors and m2 muscarinic acetylcholine receptor when compared to untreated cells. Combinations of PGE1 and forskolin produced a more pronounced decrease in mRNA than either agent alone. PGE1 and forskolin increased intracellular levels of cAMP in human corpus cavernosum smooth muscle cells and combinations of both agents produced a more pronounced increase in cAMP synthesis. These results suggest that cAMP modulates the expression of alpha adrenergic receptors, one of the principal contractile receptor systems in the corpora cavernosa. These observations further support the concept that erectile function is a balance between contractile and relaxatory processes, which in turn regulate structure and function of the corpora cavernosa. International Journal of Impotence Research (2000) 12, Suppl 1, S41-S47 PMID- 10849566 TI - Role of alpha adrenergic receptors in erectile function. AB - Penile erection is a complex physiological process in which the regulation of penile hemodynamics depends on signal input from central and peripheral nervous systems, and on the balance and interplay between several local physiological mediators (neurotransmitters, vasoactive agents and endocrine factors). A role for the sympathetic nervous system in attaining or maintaining penile flaccidity and detumescence is well recognized. However, the exact mechanisms of alpha adrenergic receptor mediated erectile function remain poorly defined. The objective of this review is to summarize data presented in the literature and from our laboratory on alpha-adrenergic receptors, and to discuss the conceptual framework by which the alpha-adrenergic receptor pathway modulates penile corpus cavernosum smooth muscle contractility. We will integrate the current state of knowledge of penile erection into one framework encompassing the biochemical and physiological pathways responsible for trabecular smooth muscle relaxation (erection) and contraction (detumescence). We will focus our discussion on local control mechanisms of alpha-adrenergic receptors and explore the following topics: (1) functional activity of alpha-1 and alpha-2 adrenergic receptors in the physiology of human penile erection; (2) identification, classification and characterization of alpha-1 and alpha-2 adrenergic receptor subtypes in human penile erectile tissue; (3) molecular mechanism of action of alpha-1 and alpha-2 adrenergic receptors in human penile erectile tissue; (4) blockade of alpha-1 and alpha-2 adrenergic receptor action by selective and non-selective alpha-1 and alpha-2 adrenergic receptor antagonists (blockers); (5) physiologic (functional) antagonism of alpha-1 and alpha-2 adrenergic receptor activity by vasodilators mediating smooth muscle relaxation; and (6) key areas of consensus, as well as critical gaps in the existing scientific knowledge concerning the rationale and the potential use of alpha-blockers in erectile function. International Journal of Impotence Research (2000) 12, Suppl 1, S48-63 PMID- 10849567 TI - Effects of alpha-2 blockade on sexual response: experimental studies with Delequamine (RS15385). AB - The role of alpha-2 receptors and alpha-2 antagonists in central and peripheral mechanisms of sexual response are discussed. It is concluded that the predominant role of the alpha-2 antagonist centrally is to increase arousal which in certain circumstances, is sexual. It is further concluded that the predominant role of the alpha-2 antagonist peripherally is to modulate (block) the norepinephrine (NE)-induced contractility in the smooth muscle of the penis. How the central arousal mechanisms are specifically linked to sexual response is not understood. Experimental studies with a selective alpha-2 antagonist, delequamine, are briefly reviewed and their complex results discussed. Evidence from sleep studies was consistent with delequamine having both central excitatory and inhibitory effects, dependent on dosage. The possibility that men with psychogenic erectile dysfunction might have increased central alpha-2 tone was considered. The apparent loss of responsiveness to the alpha-2 antagonist in older dysfunctional men was discussed. More questions are raised than answered; further research is needed in this area. International Journal of Impotence Research (2000) 12, Suppl 1, S64-S69 PMID- 10849568 TI - Yohimbine in erectile dysfunction: the facts. AB - Yohimbine, a pharmacologically well-characterized alpha-2-adrenoceptor antagonist with activity in the central and peripheral nervous system, has been used for over a century in the treatment of erectile dysfunction. In-depth, systematic studies in animals have shown that the drug has a remarkable positive effect on sexual performance. Meta-analyses of the few controlled, randomized human studies have consistently shown an advantage of yohimbine over placebo. Despite such a long history and encouraging activity, the drug has not yet been subjected to scientifically rigorous human clinical trials. Although relevant basic pharmacological and animal research information has been available for over 15 y, recent studies were designed with a lack of insight and complete disregard of those fundamental studies. Currently, dose-response investigations are not available, alternative routes of administration (i.e. sublingual) have not been investigated, nor has continuous versus 'on-demand' administration been explored. Synergistic activity with other drugs was last studied nearly four decades ago. Assessment of various populations was carried out in very limited cohorts and only in most general terms. In short, properly designed trials in humans have not been done. Why? Yohimbine is an old drug. As such it does not enjoy patent protection or commercial viability. Until molecular/formulation changes can be brought about (as recently happened with two other agents: phentolamine and apomorphine), serious investigations of yohimbine will remain in limbo. It could be that the nay sayers are right and yohimbine, indeed, lacks clinical activity as a treatment for men with erectile dysfunction. As long as it remains an orphan drug, we will never know. International Journal of Impotence Research (2000) 12, Suppl 1, S70-S74 PMID- 10849569 TI - Oral phentolamine: an alpha-1, alpha-2 adrenergic antagonist for the treatment of erectile dysfunction. AB - Phentolamine mesylate is an alpha-1 and alpha-2 selective adrenergic receptor antagonist which has undergone clinical trials for erectile dysfunction treatment. Biochemical and physiological studies in human erectile tissue have revealed a high affinity of phentolamine for alpha-1 and alpha-2 adrenergic receptors. Based on pharmacokinetic studies, it is suggested that 30-40 min following oral ingestion of 40 or 80 mg of phentolamine (Vasomax), the mean plasma phentolamine concentrations are sufficient to occupy the alpha-1 and -2 adrenergic receptors in erectile tissue and thereby result in inhibition of adrenergic-mediated physiologic activity. In large multi-center, placebo controlled pivotal phase III clinical trials, the mean change in the erectile function domain of the International Index of Erectile Function scores (Questions 1-5 and 15) from screening to the end of treatment was significantly higher following use of active drug (40 mg and 80 mg) compared to placebo. Three to four times as many patients receiving phentolamine reported being satisfied or very satisfied compared with those receiving placebo. At doses of 40 mg and 80 mg respectively, 55% and 59% of men were able to achieve vaginal penetration with 51% and 53% achieving penetration on 75% of attempts. The correction of erectile dysfunction or improvement to a less severe category of dysfunction was experienced by 53% of men with the 80 mg dose and 40% with the 40 mg dose of phentolamine. All trends of response were the same regardless of any concomitant medication. There were no severe adverse events. At 40 mg, 7.7% experienced rhinitis and fewer than 3.1% experienced any other side effect of treatment. Phentolamine is safe, well tolerated and efficacious for the treatment of erectile dysfunction. International Journal of Impotence Research (2000) 12, Suppl 1, S75-S80 PMID- 10849570 TI - Comparison of oral and intracavernosal vasoactive agents in penile erection. AB - This discussion summarizes the potential differences in physiologically and pharmacologically induced erections and highlights the possible differences in the pathways facilitated by oral versus intracavernosal agents in penile erection. Oral agents act in conjunction with sexual stimulation either increasing corporal smooth muscle relaxation or attenuating smooth muscle contraction. However, their efficacy is dependent on sexual stimulation. Intracavernosal administration of phosphodiesterase type 5 inhibitor (sildenafil) or short-acting alpha adrenergic receptor antagonist (phentolamine), in the absence of sexual stimulation, does not initiate penile erection. In contrast, intracavernosal administration of PGE1 or papaverine induces erection independent of sexual stimulation. Thus, oral agents are not direct mediators of smooth muscle relaxation and act to facilitate relaxation in response to sexual stimulation, while intracavernosal agents directly mediate smooth muscle relaxation, independent of sexual stimulation. Although, considerable advances have been made in elucidating the physiology and pharmacology of erectile function, details of the signal transduction pathways affected by these agents in the penile corpus cavernosum are yet to be fully investigated. International Journal of Impotence Research (2000) 12, Suppl 1, S81-S88 PMID- 10849571 TI - Adiposity as a risk determinant for postmenopausal breast cancer. AB - BACKGROUND: Cross-cultural studies have long suggested that the high incidence of breast cancer in developed Western countries may be linked to the high prevalence of obesity and to high total energy intake. Recent studies show that hyperinsulinemia, increased concentration of insulin-like growth factor 1 and greater abdominal fat accumulation are markers of high risk for breast cancer. Increased serum concentrations of free estradiol and free testosterone are similar risk markers and are frequent concomitants of hyperinsulinemia. MECHANISMS: The mechanism by which such metabolic-endocrine abnormality may promote mammary carcinogenesis is uncertain, but its effect is likely to predominate in the years approaching the menopause when obesity is common among Western women. In obese subjects, enlargement of adipose deposits is known to produce an excess of free fatty acids and tumor necrosis factor alpha, both of which may be involved in the pathogenesis of insulin resistance. In middle-aged women, the concomitants of hyperinsulinemia may activate invasive and proliferative activity in preneoplastic mammary lesions such as in situ duct carcinoma of the comedo type. This will enhance the risk of progression to invasive breast cancer, which is likely to manifest clinically mainly after the menopause. CONCLUSION: Weight reduction combined with a program of regular physical exercise has been shown to reduce both estrogen and insulin concentrations in obese women and such a regimen might be tested in clinical trials for an effect on breast cancer risk in obese women. Intervention is best begun around the age of 45 y, this being the age when in situ duct carcinoma would normally begin to show evidence of spontaneous involution in women without clinical evidence of invasive breast cancer. International Journal of Obesity (2000) 24, 527-533 PMID- 10849572 TI - Intentional weight control and food choice habits in a national representative sample of adults in the UK. AB - OBJECTIVE: To establish the association between intentional weight control and specific eating behaviours. DESIGN: An interview-based survey of a representative sample of the UK population including questions about demographic characteristics, weight and height, intentional weight control and eating behaviours. Associations among the eating behaviours, and between weight control and eating behaviours, were examined. SUBJECTS: 1894 men and women completed the interview (70% response rate). RESULTS: Approximately equal proportions of the sample were 'not bothered about weight' (30%), 'watching their weight' (36%), or 'trying to lose weight' (28%). More men were 'not bothered' and more women were 'trying to lose'. People who were trying to lose or were watching their weight were more likely to report restricting fats, sugars, snacks, and the amount eaten at meals, than those who were not bothered, but there were no differences between weight watchers and weight losers. The overall level of restriction among weight losers was modest. There were no group differences in eating breakfast, fruits or vegetables, skipping meals or fasting. CONCLUSIONS: Around two-thirds of the adult population of the UK appear to be concerned about weight control, and this is reflected in somewhat higher than average adherence to recommended restrictive dietary habits. PMID- 10849573 TI - Does obesity influence foot structure in prepubescent children? AB - OBJECTIVE: This study examines the relationship between obesity and foot structure in prepubescent children. DESIGN: Field-based, experimental data on BMI (body mass index) and foot structure were collected for 431 consenting children from 18 randomly selected primary schools. SUBJECTS: Of the 431 participants, 62 obese (BMI>95th percentile) and 62 non-obese (10th percentile90th percentile) children (age = 8.5+/-0.5 y) were selected. MEASUREMENTS: Height and weight were measured to calculate BMI. Static weight-bearing footprints for the right and left foot of each subject were then taken using a pedograph to calculate the Footprint Angle and the Chippaux-Smirak Index as representative measures of the surface area of the foot in contact with the ground. RESULTS: A significant difference was found between the Footprint Angle of the obese and non obese subjects for both the left (t = 3.663; P<0.001) and right (t = 3.742; P<0.001) feet whereby obese subjects displayed a reduced angle. Chippaux-Smirak Index scores were also significantly different for both the left (t = -6.362; P<0.001) and right (t=-5.675; P<0.001) feet between the two subject groups where a greater score for the obese subjects was evident. A decreased footprint angle and an increased Chippaux-Smirak Index are characteristic of structural foot changes that have been associated with compromised foot function. CONCLUSIONS: Excess body mass appears to have a significant effect on the foot structure of prepubescent children whereby young obese children display structural foot characteristics which may develop into problematic symptoms if excessive weight gain continues. Further investigation into possible consequences, particularly any effects on foot function, is warranted. PMID- 10849574 TI - The Trp64Arg polymorphism of the beta3-adrenergic receptor gene and obesity in Chinese subjects with components of the metabolic syndrome. AB - BACKGROUND: In regions such as Hong Kong, rapid economic development has led to lifestyle alterations characterized by increases in energy and fat intake and reduction in physical activity. These changes have been associated with a dramatic increase in the prevalence of diabetes and related diseases of the metabolic syndrome. OBJECTIVE: To investigate if a common polymorphism (Trp64Arg) of the beta3-adrenergic receptor gene, previously implicated as predisposing to type 2 diabetes mellitus or obesity in other populations, has a role in the apparent susceptibility of Hong Kong Chinese to diabetes and related disorders. METHOD: A PCR-based protocol was used to genotype 802 Southern Chinese subjects who were either healthy or had one or more of the metabolic disorders including diabetes, hypertension or dyslipidaemia. RESULTS: The frequencies of the mutant A allele (12.7%) and AA genotype (1.7%) did not differ, by the chi2 test, in any patient group with diabetes, hypertension or dyslipidaemia, alone or in combination, compared to healthy controls. Using the t-test in the 802 subjects, those carrying the mutant A allele had evidence of increased obesity with a significantly (all P<0.05) higher body mass index (BMI, kg/m2) and also lower HDL cholesterol. BMI was also elevated in subjects with the A allele in the separate groups with diabetes, dyslipidaemia or hypertension. Stepwise multiple regression showed this polymorphism to be an independent predictor of BMI. CONCLUSION: These data do not support any direct involvement of the Trp64Arg polymorphism in the development of diabetes, hypertension or dyslipidaemia in Chinese subjects, but do suggest a relationship with obesity. PMID- 10849575 TI - Effects of hyperglycaemia and hyperinsulinaemia on plasma amino acid levels in obese subjects with normal glucose tolerance. AB - OBJECTIVE: To investigate the effects of hyperglycaemia and hyperinsulinaemia on amino acid disposal in human obesity. DESIGN: Four sequential experimental conditions: (1) overnight fasting; (2) hyperglycaemia with hyperinsulinaemia (2 h hyperglycaemic clamp at 11 mmol/l); (3) hyperglycaemia with basal insulin (1 h hyperglycaemic clamp during somatostatin infusion), (4) hyperglycaemia with resuming hyperinsulinaemia (1 h hyperglycaemic clamp after somatostatin discontinuation). SUBJECTS: Seven non-obese and seven obese non-diabetic, normo insulinaemic subjects. MEASUREMENTS: Glucose infused to maintain steady-state hyperglycaemia. Plasma insulin, glucagon, free fatty acid and amino acid concentrations in the last 20 min of the four experimental conditions. Net rates of plasma amino acid disappearance and appearance (micromol/l per hour), calculated as the slopes of the regression of amino acid concentration on time. RESULTS: The amount of glucose infused to maintain hyperglycaemia was reduced by nearly 50% in obese subjects. During hyperinsulinaemia, FFA suppression was lower in obese subjects. In all experimental conditions plasma amino acid levels were slightly, non-significantly higher in obese than in non-obese subjects. In both groups plasma amino acids decreased slightly with ongoing fasting, decreased remarkably during hyperglycaemia-hyperinsulinaemia, rose promptly when insulin concentration was suppressed by somatostatin infusion, and declined again after somatostatin discontinuation. Also the time-course of plasma branched-chain amino acids, which paralleled that of total amino acids, was similar in the two groups. The net rates of amino acid disappearance from plasma did not differ in obese and non-obese subjects both at fasting and during hyperglycaemia-hyperinsulinaemia. Also plasma amino acid appearance during hyperglycaemia with basal insulin was not different in the two groups. CONCLUSION: The net traffic of amino acids to and from plasma in relation to insulin drive and prevailing glucose is not impaired in obese subjects with normal glucose tolerance, in spite of a decreased insulin sensitivity of glucose and lipid metabolism. PMID- 10849576 TI - An autosomal genomic scan for loci linked to plasma leptin concentration in Pima Indians. AB - OBJECTIVE: To identify chromosomal regions linked to plasma leptin concentrations. DESIGN: Autosomal genome-wide scan, including 516 microsatellite markers. Sib-pair (Haseman-Elston) and variance components methods were used to assess genetic linkage. SUBJECTS: 770 Pima Indians comprising 239 nuclear families (for a total of 1199 sibling-pairs). MEASUREMENTS: Plasma leptin concentrations and body mass index (BMI), adjusted for age and sex. RESULTS: The strongest evidence for linkage with plasma leptin concentration was on chromosome 6p logarithm of odds (LOD) = 2.1 by variance components analysis). There was no evidence for linkage to BMI in this region. Additional regions with marginal evidence for linkage to plasma leptin concentration (LOD > or =1.0) were detected on chromosomes 3, 11, 13, 15 and 16. CONCLUSIONS: The results suggest that a locus on chromosome 6p influences plasma leptin concentrations. Replication studies are needed to exclude the possibility that linkage has been falsely detected. PMID- 10849577 TI - The effects of 18 months of intermittent vs. continuous exercise on aerobic capacity, body weight and composition, and metabolic fitness in previously sedentary, moderately obese females. AB - OBJECTIVES: To compare the effects of 18 months of continuous vs intermittent exercise on aerobic capacity, body weight and composition, and metabolic fitness in previously sedentary, moderately obese females. DESIGN: Randomized, prospective, long-term cohort study. Subjects performed continuous exercise at 60 75% of maximum aerobic capacity, 3 days per week, 30 min per session, or exercised intermittently using brisk walking for two, 15 min sessions, 5 days per week. MEASURES: Aerobic capacity, body weight, body composition, and metabolic fitness (blood pressure, lipids, glucose and insulin). RESULTS: Significant improvements for aerobic capacity of 8% and 6% were shown for the continuous and intermittent exercise groups, respectively. Weight loss for the continuous exercise group was significant at 2.1% from baseline weight and the intermittent group was essentially unchanged. The continuous group showed a significant decrease in percentage of body fat and fat weight while the intermittent group did not. HDL cholesterol and insulin were significantly improved for both groups. CONCLUSIONS: In previously sedentary, moderately obese females, continuous or intermittent exercise performed long-term may be effective for preventing weight gain and for improving some measures of metabolic fitness. PMID- 10849578 TI - A case-control study of successful maintenance of a substantial weight loss: individuals who lost weight through surgery versus those who lost weight through non-surgical means. AB - OBJECTIVE: To determine if method of weight loss (surgery; non-surgery) is associated with current levels of psychosocial functioning or current weight maintenance behaviors in individuals who have lost large amounts of weight. DESIGN: Subjects were 67 cases and 67 controls selected from the National Weight Control Registry, a longitudinal study of individuals successful at long-term maintenance of weight loss. Cases had initially lost weight through bariatric surgery while controls had lost weight through non-surgical means. The current psychosocial functioning and weight maintenance behaviors of cases and controls were assessed and compared. RESULTS: Cases and controls were matched on gender, current weight and total weight loss. Surgical cases reported significantly higher fat intake and lower physical activity levels. There were no differences in cases' and controls' reports of the impact of weight loss on other areas of their lives, neither were there differences on measures of depression or binge eating. CONCLUSIONS: Reported improvement in psychosocial functioning did not depend upon how weight was initially lost, but cases and controls appear to be using very different behaviors to maintain their weight losses. PMID- 10849579 TI - Novel polymorphisms in the neuropeptide-Y Y5 receptor associated with obesity in Pima Indians. AB - OBJECTIVE: To investigate whether the neuropeptide Y receptor 5 gene (NPY5R) is associated with obesity in humans. DESIGN: The NPY5R gene was screened for polymorphisms by direct sequencing in two groups of Pima Indians, selected for extremes of body mass index (BMI). Genotype frequencies were analyzed for association with BMI extreme. SUBJECTS: Full-heritage Pima Indians, non-diabetic and not first degree relatives. Obese group: 19 M/24 F, BMI = 49+/-7 kg/m2 (mean+/-s.d.) age = 24+/-2 y, lean group: 16 M/16 F, BMI = 23+/-2 kg/m2, age = 27+/-3 y. MEASUREMENTS: Initially, the entire gene (proximal promoter, exon 1A, coding sequence, 5' and 3' UTRs) was sequenced in a subset of 20 individuals. No variants were found in the coding sequence, however three novel single nucleotide polymorphisms were detected in the non-coding regions: (1) a C-->T transition located within the promoter 28 bp upstream of the exon 1A transcription start site; (2) a T-->C transition 94 bp downstream of the stop codon; and (3) a G-->A transition 432 bp downstream of the stop codon. The polymorphisms were then screened in all 75 subjects. RESULTS: The polymorphisms had mean heterozygosities of 0.34-0.50 and were in strong linkage disequilibrium (P<0.001). Genotype frequencies differed significantly in lean and obese Pimas for P2 (P=0.04) and for a triple haplotype (P=0.02, Bonferroni corrected). CONCLUSION: Considering the importance of this gene in regulation of body weight, the association of these polymorphisms with extremes of BMI in Pima Indians indicates that NPY5R, or a locus nearby, may contribute to susceptibility to obesity in this population. PMID- 10849580 TI - Depot-related and thiazolidinedione-responsive expression of uncoupling protein 2 (UCP2) in human adipocytes. AB - OBJECTIVES: Uncoupling protein 2 (UCP2) is a recently described homologue of the uncoupling protein of brown adipocytes (UCP1), which is expressed at high levels in human white adipose tissue. Studies were undertaken (1) to establish whether the expression of UCP2 mRNA varies in a depot-related manner in isolated human adipocytes, (2) to determine whether thiazolidinedione exposure influences the expression of UCP2 mRNA in cultured human pre-adipocytes, and (3) to determine whether human UCP2 is targeted to mitochondria when transfected into mammalian cells. SUBJECTS: Abdominal subcutaneous and omental adipose tissue biopsies were obtained from adult patients undergoing elective intra-abdominal surgical procedures. MEASUREMENTS: A competitive reverse transcriptase-polymerase chain reaction (RT-PCR) was used to quantify UCP2 mRNA expression in human omental and subcutaneous adipocytes, and in cultured human preadipocytes differentiated in vitro using the thiazolidinedione, BRL49653. Chinese hamster ovary cells were transfected with a vector expressing human UCP2, and its cellular localization was determined by confocal immunofluorescence microscopy. RESULTS: Adipocytes isolated from human omentum consistently expressed more UCP2 mRNA than did subcutaneous adipocytes from the same subjects (mean fold difference 2.92+/-0.44 P<0.001, n=11) with no effect of gender or body mass index being seen. BRL49653 treatment of subcutaneously, but not omentally, derived preadipocytes stimulated expression of UCP2 mRNA (5.1+/-1.1 fold). Transfected human UCP2 was detected exclusively in mitochondria of CHO cells. CONCLUSIONS: Increased expression of UCP2 in human omental adipose tissue relative to subcutaneous adipose tissue is related to the expression levels in adipocytes per se, a finding which may relate to the particular functional attributes of this sub-population of adipocytes. Furthermore, BRL 49653 has site-specific effects of on the expression of UCP2 in human preadipocytes, a finding which may be relevant to the therapeutic effects of such compounds. Finally we present evidence for the mitochondrial localisation of human UCP2. PMID- 10849581 TI - Ethnic differences in perceptions of body size in middle-aged European, Maori and Pacific people living in New Zealand. AB - OBJECTIVES: The aim of this study was to compare perceptions of body size in European, Maori and Pacific Islands people with measured body mass index (BMI), waist-to-hip ratio and change in BMI since age 21 y. Socio-demographic factors that influenced perceptions of body size were also investigated. DESIGN: Cross sectional survey. METHODS: Participants were 5554 workers, aged > or =40 y, recruited from companies in New Zealand during 1988-1990. RESULTS: Prevalences of BMI>25 kg/m2 were: Europeans, 64.7% men, 47.2% women; Maori, 93.2% men, 80.6% women; and Pacific Islanders, 94.1% men, 92.9% women. Similarly, prevalences of BMI >30 kg/m2 were: Europeans, 14.4% men, 14.6% women; Maori, 55.0% men, 41.9% women; and Pacific Islanders, 55.1% men, 71.7% women. At each perception of body size category, Maori and Pacific Islands men and women had a higher BMI than European men and women, respectively. BMI increased with increasing perception of body size in all gender and ethnic groups. Since age 21, increases in BMI were highest in Pacific Islands people and increased with increasing perceptions of body size category in all ethnic and gender groups. BMI adjusted odds (95% CI) of being in a lower perception category for body size were 1.70 (1.38-2.12) in Maori and 8.99 (7.30-11.09) in Pacific people compared to Europeans, 1.27 (1.13-1.42) times higher for people with no tertiary education, 1.41 (1.25-1.59) times higher in people with low socioeconomic status, and 0.94 (0.92- 0.95) for change in BMI since age 21. CONCLUSION: Nutritional programs aimed at reducing levels of obesity should be ethnic-specific, addressing food and health in the context of their culture, and also take into account the socioeconomic status of the group. On the population level, obesity reduction programs may be more beneficial if they are aimed at the maintenance of weight at age 21. PMID- 10849582 TI - The association between television viewing and overweight among Australian adults participating in varying levels of leisure-time physical activity. AB - OBJECTIVES: To investigate the effect of physical activity on the association between television viewing and overweight (body mass index (BMI) > or =25 kg/m2). DESIGN: Cross-sectional study administered by interview to adults randomly selected from the electronic white pages. SUBJECTS: 3392 adults (64% response rate) from a representative population sample in the State of New South Wales, Australia. MEASUREMENTS: Self-reported height and weight, two-week leisure-time physical activity recall, one-week average television viewing recall. RESULTS: BMI and physical activity patterns were both associated with hours of television watched. Compared to those participants who reported watching less than one hour of television per day, those watching 1 to 2.5 hours were 93% more likely to be overweight (BMI>/=25 kg/m2), those watching 2.5 to 4 hours were 183% more likely to be overweight, those watching more than 4 hours per day were four times more likely to be overweight. Physical activity was not directly associated with being overweight, but an interaction between activity and television watching was present. Respondents in the low, moderate and high physical activity categories who reported watching more than 4 hours of television per day were twice as likely to be overweight compared to those who watched less than one hour of television per day, irrespective of physical activity participation. CONCLUSIONS: With approximately half the Australian adult population overweight or obese, these findings indicate that public health strategies to reduce overweight and prevent weight gain may need to focus on reducing sedentary behaviours such as television viewing in addition to increasing physical activity. PMID- 10849583 TI - Change in intra-abdominal adipose tissue volume during weight loss in obese men and women: correlation between magnetic resonance imaging and anthropometric measurements. AB - OBJECTIVE: To compare the changes in intra-abdominal adipose tissue (IAAT) measured by magnetic resonance imaging (MRI) with changes in central abdominal fat (CAF) measured by dual-energy X-ray absorptiometry (DXA) and anthropometric measurements in obese subjects before and after a weight loss programme. DESIGN: Longitudinal, clinical intervention study of a 600 kcal/day deficit diet with 10 mg/day sibutramine per day for six months. SUBJECTS: Nineteen women (age: 42+/ 8.7 y, BMI: 33.3+/-1.9 kg/m2) and 17 men (age: 41.8+/-5.3 y, BMI: 32.6+/-2.4 kg/m2). MEASUREMENTS: MRI was used to measure the effect of weight loss on IAAT. Changes in IAAT were compared with changes in CAF by DXA and anthropometry. RESULTS: The percentage of changes in IAAT was greater than that in subcutaneous adipose tissue (SCAT) in both women and men (P<0.01). Changes in IAAT were significantly correlated with changes in weight and BMI in both women and men. In women and not in men, changes in CAF by DXA, waist circumference and WHR were also significantly correlated with the changes in IAAT. CONCLUSION: Estimation of the change in IAAT was better in obese women than obese men. In both sexes the changes in weight and BMI had the highest correlation coefficients with the change in IAAT. PMID- 10849584 TI - Obesity and abdominal fatness associated with undernutrition early in life in a survey in Rio de Janeiro. AB - BACKGROUND: Undernutrition early in life has been associated with chronic diseases and obesity among adults. Our study tested the hypothesis by examining the association between low stature, a marker of early poor nutrition, with obesity and abdominal fatness among adults. METHODS: A population-based survey was conducted in 1996, among 2040 households, with a non-response rate of 11.2%. Weight, height, waist and hip circumference, and skinfolds were measured at home. RESULTS: Age-adjusted prevalence of body mass index (BMI) greater than 25 kg/m2 was 32% more frequent among adult men, and 60% more frequent among adult women, comparing the first to the fourth quintile of height. A J-shaped curve describes the association between weight and the sum of skinfolds with stature after adjusting for confounding by age, energy intake, physical activity, smoking, age at menarche, and race. The adjusted odds ratio of obesity (BMI>30 kg/m2) for short stature, compared to normal stature, was 1.57 with a 95% confidence interval (CI) = 0.90-2.71 among men and 1.84 with a 95% CI=1.10-3.06 among women. Short stature was associated with the risk of abdominal fatness only among women, with an odds ratio=1.77; 95% CI=1.10-2.83. CONCLUSIONS: Increased risk of obesity and abdominal fatness among women of short stature, a marker for undernutrition early in life, was not explained by racial and socio-economic conditions, energy intake or age at menarche. PMID- 10849585 TI - Serum lipid and leptin concentrations in hypopituitary patients with growth hormone deficiency. AB - OBJECTIVE: To investigate the effects of growth hormone (GH) deficiency on serum lipid and leptin concentrations in hypopituitary patients taking conventional replacement therapy and to determine the relations between leptin and gender and anthropometric and metabolic variables. SUBJECTS: Twenty-one GH deficient adult hypopituitary patients (15 women, six men) and 21 (14 women, seven men) age, sex and body mass index (BMI) matched healthy controls. MEASUREMENTS: After an overnight fast, anthropometric parameters were measured and body composition was determined by a bioelectrical impedance analyser. Venous blood samples were obtained for the measurements of glucose, total cholesterol, high density lipoprotein (HDL) cholesterol, triglyceride, intact insulin, insulin-like growth factor 1 (IGF-1) and leptin concentrations. Serum leptin and hormones were analysed by radioimmunoassay. RESULTS: Hypopituitary patients with GH deficiency showed significantly higher triglyceride, total and low density lipoprotein (LDL) cholesterol and lower HDL cholesterol concentrations on conventional replacement therapy. The unfavourable lipid profile was particularly evident in women. Significantly higher leptin concentrations were found in patients compared with healthy controls with similar body fat content (23. 5+/-11.8 ng/ml vs 11.7+/-6.9 ng/ml, P=0.01). This difference remained significant even when leptin values were expressed in relation to fat mass percentage (0.79+/-0.40 vs. 0.42+/-0.17 ng/ml%, P<0.05) and fat mass kg (1.32+/-0.81 vs 0.66+/-0.30 ng/ml kg, P<0. 05). Significant positive correlations were observed between leptin concentrations and body fat percentage and age in the control group. In patients the sole significant relation between leptin and study parameters was the positive correlation observed between leptin and total cholesterol concentrations. Serum leptin concentrations were significantly higher in women than men in the control group, but not in the patients. No significant gender difference was observed when leptin concentrations were expressed in relation to fat mass (percentage and kg). CONCLUSION: Growth hormone deficient hypopituitary patients (particularly women) on conventional replacement therapy have a more atherogenic lipid profile. Leptin concentrations are increased in GH deficient adults even after adjustment for percentage body fat and body fat mass (kg). Although the nature of our data does not allow us to draw any conclusions on the mechanism(s) of increased leptin concentrations in GH deficiency, decreased central sensitivity to leptin and increased leptin production from per unit fat mass, or alterations in leptin clearance, might be operative. PMID- 10849586 TI - Overweight and obese children have low bone mass and area for their weight. AB - OBJECTIVES: To determine whether girls and boys categorized from body mass index (BMI) values as overweight or obese for their age have lower bone mineral content (BMC) or lower bone area in relation to total body weight than children of normal adiposity. DESIGN: Cross-sectional study in a university bone research unit. SUBJECTS: Two hundred girls and 136 boys aged 3-19 y recruited from the general population by advertisement. MEASUREMENTS: Total body BMC (g) and bone area (cm2) measured by dual energy X-ray absorptiometry (DXA) in relation to body weight (kg), lean tissue mass (kg) and fat mass (kg) in boys and girls of three different BMI percentile groupings: normal weight (BMI<85th percentile); overweight (85 to 94th BMI percentile); obese (> or =95th BMI percentile). RESULTS: Obese children had higher BMC, bone area, and fat mass for chronological age than those of normal body weight (P<0.001). In spite of this the observed values for age-adjusted total body BMC and bone area relative to body weight were each lower than predicted values, in both overweight and obese children (2.5 10.1% less, P<0.05) than in children of lower adiposity. CONCLUSION: In overweight and obese boys and girls there is a mismatch between body weight and bone development during growth: their bone mass and bone area are low for their body weight. PMID- 10849587 TI - Associations between energy density and macronutrient composition in the diets of pre-school children: sugars vs. starch. AB - OBJECTIVE: To investigate the associations between energy density (ED) and macronutrient composition in the National Diet and Nutrition Survey of Children aged 1(1/2)-4(1/2) y, hypothesizing that high-ED diets tend to be high in sugars as well as fat. DESIGN: Further analysis of data from a cross-sectional dietary survey of 1675 children with complete 4-day weighed dietary records. Differences in diet composition and food choice between children with diets of high, medium and low ED (defined as kJ/g of all food and drink) were identified. The possibility of confounding by water, or by soft drinks, was also explored in age adjusted correlations. RESULTS: High-ED diets (>3.7 kJ/g of total diet) were proportionately richer in fat and lower in carbohydrate, compared with diets of low ED (<2.9 kJ/g). In contrast to the hypothesis, high-ED diets were found to be proportionately lower in sugars, and higher in starch. Children with high-ED diets consumed more of a whole range of foods: meat, eggs, potatoes, cereal products, confectionery, sugar/preserves and savoury snacks, but consumed less soft drinks, water and fruit. CONCLUSION: The inverse relationship observed between sugars and energy density may be partly attributable to the reciprocal relationship between sugars and fat, expressed as a proportion of energy. It may also reflect developing preferences in young childhood for a more adult-type, energy-dense, diet. Further work is required to verify ED/macronutrient relationships in other age groups, as the results have potential implications for obesity prevention and for food product development. PMID- 10849588 TI - Development of high fat diet-induced obesity and leptin resistance in C57Bl/6J mice. AB - OBJECTIVE: To investigate the development of high fat diet-induced obesity and leptin resistance. DESIGN: Two experiments were carried out in this study. Firstly, we fed the mice with a high- or low-fat diet for up to 19 weeks to examine a progressive development of high fat diet-induced obesity. Secondly, we examined peripheral and central exogenous leptin sensitivity in mice fed high- or low-fat diets for 1, 8 or 19 weeks. SUBJECTS: A total of 168 C57BL/6J mice (3 weeks old) were used in this study. MEASUREMENTS: In the first experiment, we measured the body weight, energy intake, adipose tissue mass, tibia bone length, and plasma leptin in mice fed either a high- or low-fat diet for 1, 8, 15 and 19 weeks. In the second experiment, body weight change and cumulative energy intake were measured at 6 h intervals for 72 h after leptin injection in mice fed a high or low-fat diet for 1, 8 or 19 weeks. RESULTS: The results from the first experiment suggested that the development of high fat diet-induced obesity in mice could be divided into early, middle and late stages. Compared with the mice fed a low-fat diet, the mice fed a high-fat diet showed a gradually increased body weight (+5.2%), fat storage (epididymal plus perirenal; +6.7%) and plasma leptin (+18%) at 1 week; +11.4%, +68.1%, and +223%, respectively, at 8 weeks; and +30.5%, +141%, and +458%, respectively, at 19 weeks. Energy intake of high fat diet-fed mice was equal to that of low fat diet-fed controls for the first 3 weeks; it fell below control levels over the next 5 week period, but began to increase gradually after 8 weeks of high-fat diet feeding and then increased dramatically from 15 weeks to be 14% higher than that of controls after 19 weeks. The results from our second experiment showed that: (1) after 1 week of feeding, the mice fed a high-fat diet were sensitive to a 2 microg/g (body weight) intraperitoneal (i. p.) injection of leptin, with no differences in body weight change or cumulative energy intake post-injection; (2) after 8 weeks of feeding, the mice fed a high-fat diet were insensitive to 2 microg/g (body weight) i.p. leptin, but were sensitive to a 0.1 microg intracerebroventricular (i.c.v.) injection of leptin; (3) after 19 weeks of feeding, the mice fed a high-fat diet were insensitive to 0. 1 microg i.c.v. leptin, but were sensitive to a high dose of 2 microg i.c.v. leptin. CONCLUSIONS: The present study demonstrated that the development of high fat diet-induced obesity (19 weeks) in C57 B1/6J mice could be divided into three stages: (1) an early stage in response to high-fat diet that mice were sensitive to exogenous leptin; (2) a reduced food intake stage when mice had an increase in leptin production and still retained central leptin sensitivity; and (3) an increased food intake stage, accompanied by a reduction of central leptin sensitivity. PMID- 10849589 TI - Variation in the gene for human peroxisome proliferator activated receptor gamma (PPARgamma) does not play a major role in the development of morbid obesity. AB - OBJECTIVE: To determine the extent of variation in the gene for peroxisome proliferator activated receptor gamma (PPARgamma) in patients with morbid obesity. SUBJECTS: Two hundred morbidly obese patients who underwent gastric banding surgery and 192 healthy blood donors. Diabetics were excluded. EXPERIMENTAL: The frequency of the P115Q and P12A variants in the PPARgamma gene was determined. Single strand conformational polymorphism (SSCP) analysis was performed on all exons, exon/intron boundaries and part of the promoter of the PPARgamma gene on a sub-group of 67 morbid obese patients. RESULTS: None of the morbid patients or the blood donors were carriers of the P115Q mutation. The frequency of the P12A polymorphism did not differ significantly between morbid obese patients and controls and there was no statistically significant association between P12A and BMI. Male blood donors who were A12A homozygotes had statistically significant higher serum leptin concentrations (P = 0.001). Mutation screening revealed that one patient had a T -->G transversion at -208 in the promoter of PPARgamma-2, two had silent mutations, one a T-->C transition in the third base of codon 144 and the other a C-->T transition in codon 297. The fourth patient had a CGC-->TGC transition in codon 316 resulting in the replacement of an arginine with a cysteine. This mutation was not found in any other morbidly obese patient. CONCLUSION: Variation in the PPARgamma gene is unlikely to play a major role in the development of morbid obesity. PMID- 10849590 TI - Reliability of anthropometric measurements in overweight and lean subjects: consequences for correlations between anthropometric and other variables. AB - OBJECTIVE: To estimate the reliability of anthropometric measurements in overweight and lean subjects, and to examine the influence of this reliability on correlations to other variables, since low reliability leads to underestimation of correlations. DESIGN: Replicate measurements by two observers in 26 overweight and 25 lean subjects measured at two occasions. MEASUREMENTS: Sagittal abdominal diameter (SAD), waist circumference (waist), waist-to-hip ratio (W/H) and skinfold measurements. RESULTS: Intra-class correlation coefficients (ICCs) for SAD and waist were higher than for W/H (0.98 vs. 0.90, P<0.001, and 0.97 vs. 0.90, P = 0.001, respectively). For waist, the ICC was lower for overweight than for lean subjects (0.85 vs 0.95, P=0.030), but the ICC values were comparable for SAD and W/H (0.92 vs. 0.95 and 0.78 vs. 0.83, respectively). Intra-observer variations (IOV) for SAD and waist were lower than for W/H (coefficients of variation; 1.6%, 1.4% and 2.3%, respectively), as were intra-subject variations (ISV) (2.7%, 3.0% and 3.4%, respectively). ICC values ranged from 0.84 to 0.93 and were lower for overweight than for lean subjects for biceps, subscapular and umbilical skinfolds (P=0.031, P<0.001 and P=0.048, respectively). Coefficients of variations for skinfold measurements ranged between 7.3% and 16.0% for IOV and between 14.9% and 20.8% for ISV. CONCLUSIONS: The low ICC values imply that correlations can be underestimated in overweight groups. We propose that, because of their higher reliability, SAD and waist have a higher predictive capacity for cardiovascular risk than W/H. SAD is the only measurement with high reliability in both weight groups and its use is recommended. PMID- 10849591 TI - Variation in the body mass index among young adult Polish males between 1965 and 1995. AB - OBJECTIVE: To investigate whether the incidence of overweight and underweight individuals among young adults showed inter-generation changes or social-class differences in Poland between the mid-1960s and mid-1990s. DESIGN: Comparisons of variation in the body mass index and in height among 19-y-old Polish males drawn from three successive birth cohorts. SUBJECTS: Three 10% nation wide random samples of 19-y-old Polish conscripts, examined in 1965, 1986 and 1995, a total of ca. 80,000 individuals. MEASUREMENTS: Body mass index (weight (kg)/height (m2) and height (m). PRINCIPAL RESULT: There has been during the three decades between the mid-1960s and mid-1990s a gradual and significant increase in the proportion of both 'overweight' and of 'underweight' young males, as well as of the very tall and very short ones in the population. CONCLUSION: The above finding seems intriguing. It may suggest that certain elements of individual lifestyles, those influencing the leanness vs fatness variation among young adults, as well as those affecting growth in height, have tended to become in Poland increasingly diversified in terms of between-family differences, irrespective of social-class differences and of the general nationwide changes in living standards. PMID- 10849592 TI - Social desirability and self-reported weight and height. AB - The present study examines the relationship between the desire to conform to perceived societal norms and the misreporting of weight and height. Self-reported and measured weights and heights for 56 young, healthy non-obese volunteers were assessed and compared to scores on the Marlowe Crowne Social Desirability Scale (MCSDS). Discrepancies between actual and self-reported weights for females were directly related to actual weight (r = 0.66, P<0.0001). The same was not true for males (r = 0.03). Height was significantly overreported, regardless of gender (P<0.004). Most importantly, for females, MCSDS scores were significantly correlated with the discrepancy between actual and self-reported weights after statistically adjusting for differences in actual weight (r = 0.51, P<0.0001). Results suggest that the misreporting of weight among young, non-obese women may be directly influenced by the desire to conform to perceived societal norms. PMID- 10849616 TI - Prognostic factors in mesial temporal lobe epilepsy surgery. AB - Eighty-four patients submitted to anterior temporal lobectomy were evaluated retrospectively in order to correlate the different type of simple partial seizure (SPS) and their prognostic implications in patients with mesial temporal sclerosis. The patients were divided in two groups following the classification of Engel; Group 1 (53 patients) included patients Class I (without seizures or of good outcome) and Group 2 (31 patients) included Classes II, III and IV (with seizures or of bad outcome). The two groups were compared and results showed no statistical difference in relation to the demographic aspects as sex, side of surgery, age at onset of seizures and time of the patients' postoperative follow up. Statistical analysis revealed no relationship between type of SPS and outcome. SPS did not show a statistical value in localizing the side of pathology. However, when the two groups were compared statistically in terms of patients' ages at the time of surgery, and the time elapsed from the onset of the seizures to the surgical intervention, it was observed that Group 1 (of good outcome) had seizures for smaller interval (p <0.05) and was operated at an earlier age (p<0.02) than Group 2 (of bad outcome). The presence or the type of SPS can not be used as a prognostic measure; surgical therapy must be considered as soon as clinical resistance is demonstrated. PMID- 10849617 TI - Sexual dysfunction in epilepsy: identifying the psychological variables. AB - In order to evaluate the psychological variables that affect sexual dysfunction (SD) in epilepsy, where compared 60 epileptics (Group 1) with 60 healthy individuals (Group 2), through the State-Trait Anxiety Inventory (Spielberger et al., 1970), Beck Depression Inventory (Beck, 1974) and Sexual Behavior Interview (Souza, 1995). Sexual dysfunction (SD), anxiety and depression were found more frequently in Group 1 than in Group 2 and were not related to sex. Variables such as the onset duration and frequency of seizures as well as the use to medication were not associated with SD. Temporal lobe epilepsy was related to SD (p = 0.035) but not to anxiety or depression. Anxiety and depression were related to SD in both groups. Perception in controlling the seizures was closely related to anxiety (p = 0) and depression (p = 0.009). We conclude that psychological factors play an important role in the alteration of sexual behavior in epileptics and that suitable attention must be given to the control of these variables. PMID- 10849618 TI - Doctors' perspectives and practices regarding epilepsy. AB - OBJECTIVE: The main aim of this study was to evaluate the knowledge, management practices and attitudes towards people with epilepsy (PWE) by a group of general practitioners (GP) and pediatrician (PD) residents. METHODS: A cross-sectional study was carried out in three training hospitals, and had been selected 31 GP and 47 PD who agreed with the study. The collection of data was made by self applied structured questionnaire. RESULTS: Many respondents have positive values about PWE, and recognize prejudice in the population against them. The residents recognize in themselves and in the colleagues lack of knowledge about PWE, and that Medical School do not give enough importance to the study of PWE. The reference of PWE to the neurologist is a common practice among the doctors. Half of them are favorable to the idea of assuming the patients clinical management after an initial clientele appraisal by the neurologist. CONCLUSIONS: The non neurologist doctors do not feel comfortable in managing PWE due to barriers. Our doctors complain about the undergraduate medical training related to the epilepsy. Although, there is not a clear relationship between the undergraduate medical training, referral practices and satisfaction about the management of PWE. The patients care is influenced not only by knowledge, but also by doctors' attitudes. In this way, there are other barriers, perceived or not, to providing care to PWE by the generalists, and they need to be approached in the medical undergraduate curriculum and medical continuing education. PMID- 10849619 TI - Client with epilepsy in a work Brazilian rehabilitation center. AB - INTRODUCTION: People with epilepsy (PWE) may have problems in obtaining or maintaining regular employment because of restrictions related to their handicap, social prejudices and also high rates of unemployment of the population. The main aim of this pilot study was to know the vocational rehabilitation problems involving PWE sent to a vocational rehabilitation center (VRC) in Rio de Janeiro. METHOD: Fifteen PWE were selected unbiased from those seen at the VCR. It was reviewed their records in the search of sociodemographic, health care, employment suitability and work rehabilitation data. RESULTS: Only one person was eligible for the training program, four were ineligible, six were temporarily ineligible, and the other four do not necessitate the rehabilitation, but as the majority, the better seizures control. CONCLUSIONS: The studied sample of selected PWE, but representative of the studied population, do not show any important successful in the vocational rehabilitation carried out at the VRC. PMID- 10849620 TI - Nocturnal sleep pattern in Native Brazilian Terena adults. AB - Social-economic factors influence sleep habits. This research analyzes characteristics of nocturnal sleep in Brazilian Native Terena adults. Sixty-four adults (31 M; 33 F) from 18 to 75 years, with a mean age of 37.0, from the Indian Reservation village of Corrego do Meio, in the central region of Mato Grosso do Sul, an agriculturally oriented group were evaluated. Nocturnal sleep characteristics were evaluated by means of a standard questionnaire applied to each individual. It was observed that reported nocturnal sleep was longer, sleep onset was earlier and wake up time was also earlier than usually described in urban populations. The mean total time in bed was 8.5 h or more, in every age bracket. The seven-day prevalence rate of insomnia was 4.6%, while the seven-day prevalence rate of hypnotic use was 1.5%, both remarkably less than described in urban populations. These findings stress the need to consider ethnic influences on sleep patterns and disorders. PMID- 10849621 TI - Rapid eye movements during paradoxical sleep in patients with cerebrovascular disease. AB - Rapid eye movements that occur during paradoxical sleep are generated from the brainstem and are modulated by cerebral hemispheres. In an attempt to establish the participation of cerebral hemispheres on rapid eye movements, we carried out a quantitative study of eye movements density in patients bearing hemispheres vascular lesions. The polysomnographic recordings of 24 patients were compared to those of 24 healthy volunteers. Density of rapid eye movements was defined as the percentage of eye movements during the respective time of paradoxical sleep. Based on the present results, we concluded that: stroke patients with hemispheric lesions displayed increased density of rapid eye movements; there was no difference on the density of rapid eye movements according to the hemispheric lesion; higher density of rapid eye movements was observed in patients with anterior hemispheric lesion. PMID- 10849622 TI - Evaluation of the areas of neuronal cell bodies and nuclei in the myenteric plexus of the duodenum of adult rats. AB - This study compared the areas of cell body and nucleus profiles of the myenteric neurons in the antimesenteric and intermediate regions of the duodenum of adult rats. Five male rats were used. The duodenum was removed and dissected to whole mount preparations, which were stained by the Giemsa technique. The areas of cell body and nucleus profiles of 100 neurons, 50 from each region, of each animal, were assessed with image analyser. Based on the global mean+/-SD of the areas of cell body profiles, neurons were labelled as small, medium or large. It was observed that the neurons did not differ significantly in size or incidence between the antimesenteric and intermediate regions. However, the nuclei of the small and medium neurons were significantly smaller in the latter region. It is discussed that the smaller nuclear size could be related to the cell bodies being slightly smaller on this region and to a possible smaller biosynthetic activity which would influence nuclear size. PMID- 10849623 TI - Body mass index and carpal tunnel syndrome. AB - Carpal tunnel syndrome (CTS) has been correlated to body mass index (BMI) increase. The present study was done in a Brazilian population to compare BMI values in the following groups: first, CTS vs. controls subjects, and, second CTS groups of increasing median sensory latency (MSL). According to MSL>/=3.7 ms (wrist-index finger, 14 cm), median/ulnar sensory latency difference>/=0.5 ms (ring finger, 14 cm) or median palm-to-wrist (8 cm) latency>/=2.3 ms (all peak measured), 141 cases (238 hands) had CTS confirmation. All were symptomatic; previous surgery and polyneuropathy were excluded; mean age 50.3; 90.8% female. Controls subjects (n=243; mean age 43.0; 96. 7% female) and CTS cases had BMI calculated (kg/m2). Controls subjects had a mean BMI of 25.43+/-4.80 versus 28.38+/-4.69 of all CTS cases, a statistically significant difference (p < 0.001). The CTS groups of increasing MSL severity do not show additional increase in BMI (28.44 for incipient, 28.27 for mild, 28.75 for moderate and 29.0 for severe). We conclude that CTS cases have a significant correlation with higher BMI when compared to controls subjects; however, higher BMI do not represent a statistically significant increasing risk for more severe MSL. PMID- 10849624 TI - [Electrophysiological study of the lateral dorsal cutaneous nerve: technical applicability and normal values]. AB - The distal nerve conduction study of the long nerve in the leg is more efficient to work with so that it can establish the early diagnosis of the majority of polyneuropathies. The main purpose of this study is the technical applicability of the orthodromic neural conduction examination of the dorsal cutaneous branch of the sural nerve (lateral dorsal cutaneous nerve) on healthy people, and define the normal values used as references to compare with the proximal segment. Forty five persons mean age 41.56 years old (range 19-75) were examined, and the sensory nerve action potentials were registered from ninety feet. The active recording superficial electrode was placed below and behind the lateral malleolus and the stimulating electrode was placed 10 cm distal to the recording superficial electrode at the dorsal lateral aspect of the feet. The mean value for the lateral dorsal cutaneous nerve conduction velocity was 47.35 +/- 4.8 m/s and for amplitudes 4.19 +/- 1.9 microV. The sensory conduction velocity in the distal segment was 14% lower than the proximal one. The sensory nerve action potential amplitude of the distal segment was 73% lower than the proximal one. The lower normal limit recommended for conduction velocity of this nerve plus correction for skin temperature of 34 degrees C is 38 m/s. Some differences in amplitude and conduction velocity among group ages are to be considered. PMID- 10849625 TI - [Evidence of cerebral involvement in the chronic stage of Chagas disease obtained using the P300 potential and quantified electroencephalography]. AB - It is already known the involvement of the peripheral nervous system in chronic stages of Chagas disease. Tomographic and neuropsychological evidence of brain compromise has been included recently. In order to evaluate the neurophysiological counterpart of cerebral involvement, we studied P300 evoked potential and quantified EEG (qEEG) of 35 patients (26-55 years), and compared to an equal number of control subjects (29-55 years). We have found increased P300 latency compared to the control group (331.24 +/- 24. 02 vs 318.86 +/- 23.18) (p=0.01716). qEEG showed lower relative Beta 1 power in the patients group (p=1.6E5), and the principal frequency 1 Hz slower in the same group (p=0.01077). Multivariate analysis showed three subpopulations: a normal one, pathological one with higher Alpha power and pathological with Alpha decrement and DeltaTheta increment. Pathological findings represented 20% for the qEEG and 11.43% for cognitive potentials. Cardiac and neurologic involvement were not correlated. We conclude that there is clear electrophysiological evidence of cerebral involvement in chronic Chagas disease, thus reinforcing findings obtained by other methods. PMID- 10849626 TI - [Alpha band coherence analysis of EEG in healthy adult's and Alzheimer's type dementia patients]. AB - We studied the occipital inter-hemispheric coherence (IHCoh) of EEG (electrodes O1-O2) for alpha band (alpha1 - 8,0 to 10,0 Hz and alpha2 - 10,1 to 12,5 Hz) in healthy adults and Alzheimer's type dementia (ATD) subjects, to observe if there is any significant difference between these two groups that could help in the early diagnosis of ATD. We found a decrease of occipital IHCoh in ATD group for both alpha sub-bands. We believe that Coh analysis of EEG is a powerful auxiliary method in ATD diagnosis. PMID- 10849627 TI - [Dose-dependent relationship of chronic use of phenytoin and cerebellar atrophy in patients with epilepsy]. AB - The chronic treatment with phenytoin or the acute intoxication by this drug may cause permanent cerebellar injury with atrophy of cerebellum vermis and hemispheres, which can be detected by neuroimaging studies. The aim of the present study was to investigate the correlation between the dosage and duration of treatment with phenytoin and the occurrence of cerebellar atrophy. Sixty-six patients were studied and had their tomographies analyzed for cerebellar atrophy. Of the 66 patients studied, 18 had moderate/severe atrophy, 15 had mild atrophy and 33 were considered to be normal. The patients with moderate/severe atrophy were those with higher exposure to phenytoin (longer duration of treatment and higher total dosage) showing statistically significant difference when compared to patients with mild atrophy or without atrophy (p=0. 02). Further, the patients with moderate/severe atrophy had serum levels of phenytoin statistically higher than those of patients with mild atrophy or without atrophy (p = 0.008). There was no association between other antiepileptic drugs dosage or duration of treatment and degree of cerebellar atrophy. We also found that older patients had cerebellar atrophy more frequently, indicating that age or duration of the seizure disorder may also be important in the determination of cerebellar degeneration in these patients. We conclude that although there is a possibility that repeated seizures contribute to cerebellar damage, long term exposure to phenytoin, particularly in high doses and toxic serum levels, cause cerebellar atrophy. PMID- 10849628 TI - [Comparative non-randomized study with midazolam versus thiopental in children with refractory status epilepticus]. AB - We evaluated the use of midazolam versus thiopental in 50 children with refractory status epilepticus (RSE), admitted in a pediatric intensive care unit. The study consisted of two groups of patients: Group A - Midazolam, a prospective study, and Group B - Thiopental, a historical group. These patients already had previous medication with benzodiazepin and diphenylhydantoin and other drugs. When there was no effective control of the seizures, the patients of Group A received midazolam of 200 microg/kg intravenous in bolus, being followed by continuous intravenous infusion at the rate 0. 25-15 microg/kg/min. Group B received thiopental 1 mg/kg intravenous in bolus followed by continuous intravenous infusion at the rate of 10-120 microg/kg/min. In relation to the time of seizure control and effectiveness, there was no statistical significance for the two groups. The Midazolam Group had significantly less complications during the treatment: less cyanosis (p=0.00006), and they did not need respiratory support (p<0.00001). When the therapy with midazolam was ceased, 12.5% of the patients from this group showed psychological disorders such as mental confusion, aggressive behavior, restlessness, hallucinations and agitation. PMID- 10849629 TI - [Duration and age range of epilepsy and its correlation with work: prospective ambulatory study in 379 epileptics]. AB - This study aims to evaluate, prospectively, the relationship of the active epilepsy length and the work, of the 379 out-patients from Hospital de Base, Sao Jose do Rio Preto. Epilepsy length was subdivided into 5 time rates and by age, into 3 age-groups. There was no relationship, not considering the age of patient among the epilepsy length and the work, but young patient (14 at 29 years old) and with recent epilepsy (length of 0 at 5 years) lost more work than the adult and elderly epileptic. PMID- 10849630 TI - [Perception of cognitive deficits and behavior disorders in patients with Alzheimer's disease]. AB - Alzheimer's disease (AD) is associated with cognitive decline and, often, the development of behavioural abnormalities. However, patients may have limited insight into the severity and extent of their impairments. This study was designed to investigate agreement between patients and carers on the assessment of cognitive and behavioural symptoms typically associated with AD. Thirty consecutive outpatients meeting criteria for the diagnosis of AD according to DSM IV and their respective carers were invited to participate in this study. An enlarged version of the questionnaire for dementia-anosognosia (QD) was used to assess cognitive and behavioural symptoms according to patients and carers. Cognitive impairment was further assessed with the mini-mental state examination (MMSE). The mean age of patients and carers was 71.38 (CI=68.23 to 74.53) and 52.48 (CI=47.11 to 57.86) respectively. Sixty and 73.3% of patients and carers were women. The mean MMSE score for patients was 14.93 (CI=12.68 to 17.18). Agreement between patients and carers for all QD items was assessed using weighted Kappa - rates were low and ranged from 0 to 0.67. Only 3 of the 42 QD items were associated with Kappa values greater than 0.40. The overall score for questions assessing cognitive abilities (QD-A) was significantly lower according to the evaluation of patients than that of carers (paired t-test = -4.07, p<0.001). The same pattern was observed for questions assessing behaviour (QD B)(paired t-test= -2.27, p=0.032). The Spearman correlation between QD-A and MMSE scores was -0.39 and -0.57 according patients and carers respectively. The association between cognitive anosognosia (difference between QD-A according to carers and patients) and MMSE scores was poor (rho= -0.14). In addition, cognitive anosognosia scores of patients with and without significant depressive symptoms was similar (t= -0.40, p=0.698). These results suggest that AD patients have limited insight into the severity and extent of their cognitive and behavioural problems. In addition, our data shows that this lack of awareness is not influenced by the severity of cognitive impairment or the presence of depressive symptoms. PMID- 10849631 TI - [Comparative analysis of 2 clinical scales for clinical evaluation in multiple sclerosis: review of 302 cases]. AB - Many neurologic scales have been used for clinical evaluation of multiple sclerosis, but there is no consensus about which one is the most appropriate to assess evolution and point to a new relapse. The Expanded Disability Status Scale (EDSS) has been the most commonly used. We analyse the reliability of two scales: the EDSS and Neurologic Rating Scale (NRS) in 302 multiple sclerosis patients. It is shown that NRS is a more sensitive scale than EDSS to disclose clinical changes (22.1% of cases). Changes in NRS were more evident in patients with EDSS 3.0 and 3.5. We comment on these findings and suggest that both scales should be employed in multiple sclerosis treatment trials. PMID- 10849632 TI - [Neurologic disorders in patients submitted to liver transplantation: analysis of 30 consecutive cases]. AB - Neurologic complications are important source of morbi-mortality, in liver transplantation. They result from previous factors, alterations during the surgical procedure, effects from immunosuppressor drugs, coagulopathy and infections. We analyzed, retrospectively, the chronology, causes, and frequencies of neurologic alterations in thirty adult patients submitted to liver transplantation, and our results differ slightly from those registered in other series. PMID- 10849633 TI - [Haemangioblastomas: clinical, epidemiological and pathological findings in 14 cases]. AB - We report the clinical, epidemiological and pathological findings of 14 patients with haemangioblastoma. Sixty-four percent occurred in males, with ages ranging from 16 to 60 years, with an average of 34, 4 years. Most of the tumours were confined to the cerebellum (n=9). The most frequent symptoms were headache (n=7) and dizziness (n=7), with a mean duration of 70 days. Von Hippel-Lindau syndrome (vHL) was diagnosed in 3 patients. Eleven patients were submitted to total surgical removal and in 3 the tumour was partially resected. A relapse rate of 28% in 3 years of follow-up was found. The patients with vHL showed recurrence in 66% of the cases. These findings corroborate those in current medical literature, showing an increased morbidity of this tumor when associated with vHL. PMID- 10849634 TI - [Carotid body tumors: review of 8 cases and surgical approach]. AB - Carotid body tumors are rather uncommon. Even though there is a great amount of literature on the subject, the controversy regarding its biological behavior and therapeutics and mainly, the surgical management still remains. We present eight patients with carotid body tumors (total of 9 tumors, one bilateral) surgically treated at the Neurosurgery Department of Santa Casa in Belo Horizonte, from 1989 to 1999. The age ranged from 11 to 66-years-old (35,6+/-17.7). Four patients were women and four were men. We had satisfactory postoperative results with low morbidity and no deaths. Based on our experience and on the review of the literature, some aspects of this disease are discussed. We conclude that carotid body tumors are uncommon and should be treat with carefully surgery techniques to obtain low morbi-mortality rates. PMID- 10849635 TI - [Tumor of ceruminous gland with intracranial invasion: case report]. AB - Ceruminous glands are modified apocrine glands, confined to the skin lining of the cartilaginous part of the external auditory meatus. Tumors arising from these glands are rare. Controversy exists regarding the term "ceruminoma". Actually this neoplasia should be classified as adenoma, adenocarcinoma, adenoid cystic carcinoma and pleomorphic ceruminous adenoma. We report a 39-year-old woman first seen at Santa Casa of Belo Horizonte, in 1998, presenting with headache, nausea, vertigo, hearing loss and tinnitus on the right for the past two years. CT scan showed a tumor eroding cartilaginous and bony limits with intracranial invasion. She was submitted to multidisciplinary treatment with surgery followed by radiotherapy (6000 cG). Histology showed a ceruminous adenoid cystic carcinoma. The patient manifested a right peripheral facial palsy and had no recovery of the previous deficits. After one year from surgery she is clinically stable. PMID- 10849636 TI - [Cerebral aneurysm and arachnoid cyst: about a case with intracystic hemorrhage]. AB - We report a case of a carotid artery bifurcation aneurysm which ruptured into a silvian fissure arachnoid cyst. In the review of the literature, only three cases were before reported. We discuss about uncommon clinical findings, the surgical aspects and the associations among the lesions. PMID- 10849637 TI - [Can the biologic pattern of cervicogenic headache change after overuse or withdrawal of ergotamine derivatives?]. AB - The transformation of a primary headache into a chronic daily headache (CDH) may or may not be related to the overuse of pain-killers, as their influence on the pathophysiological mechanisms remain inconclusive. We describe three patients (female, aged 65 and 39 years, and male, 46) affected by cervicogenic headache (CH) and CDH linked to the overuse of pain-killers (ergotamine derivatives) that were submitted to the infiltration of the greater occipital nerve (GON). At the end of three days of treatment, a total improvement of the pain symptoms was recorded, which allowed for the withdrawal of the ergotamine derivatives. The CH cannot be ranked with the CDHs, since it presents an organic etiology; however, if the pain is daily and the diagnosis is belated, the indiscriminate and excessive use pain-killers may occur. In the cases described, the overuse of pain killers did not affect the natural evolution of this headache after treatment with the infiltration of the GON, as all the patients who underwent infiltration showed a total improvement of their painful symptoms, without headache resulting from the withdrawal of pain-killers, nor did they show any pharmacological dependence. This is an evidence that the CH presents and organic etiology, not being influenced in its pathophysiology by the overuse of ergotamine derivatives. PMID- 10849638 TI - Fibrodysplasia ossificans progressiva: case report. AB - Fibrodysplasia ossificans progressiva is a rare genetic disease characterized by widespread soft tissue ossification and congenital stigmata of the extremities. We report on a male child followed for ten years since the age of 3 years and 9 months, when the diagnosis was made. He was born with bilateral hypoplasic hallux valgus and ventricular septal defect, corrected by trans-sternal approach when 32 months old. Restriction of neck mobility followed and foci of ectopic ossification appeared. Four crises of disease exacerbation were treated with oral prednisone and/or other antiinflammatory drugs. Sodium etidronate 5 to 10 mg/kg/day was prescribed intermittently during about six years but was discontinued due to osteopenia. The disease course has been relentless, with severe movement restriction including the chest wall. A review showed few similar case reports in the Brazilian literature. We revisit the criteria for diagnosis and the essentials of management and treatment. PMID- 10849639 TI - The Wada test with propofol in a patient with epilepsy. AB - The usual drug used in the Wada test is amobarbital, but it is not available in Brazil. Propofol was already used by Bazin et al. in 1998, and in their report the test resulted good in the absence of any adverse effect. We report the use of propofol as the anesthetic for the Wada test. The test was carried out in a 26 years old woman with temporal medial lobe epilepsy refractory to medical treatment. Language functions and memory were tested after injection in both hemispheres by three procedures (Seattle, Montreal and Interview procedures). Propofol showed to be good to carry on the Wada test. PMID- 10849640 TI - [Hypertensive encephalopathy associated to repetitive seizures: case report]. AB - Five years old, female, who started with tonic-clonic seizures on the right side of the body, with vomits and unconsciousness. The patient had been hospitalized for eight times in the last sixty days because of seizures. At physical exam, she had a severe arterial hypertension (270/140 mmHg). The computerized tomographic scan and magnetic resonance imaging revealed hypodense areas, mainly on the right parietal-temporal region, suggesting presence of edema. The angiography showed stenosis of the right renal artery, that was the cause of arterial hypertension. After the control of arterial hypertension by nephrectomy, the patient had a complete remission of the symptoms, as well as the images anomalies. PMID- 10849641 TI - [Basilar artery dissection: case report]. AB - We report a case of basilar artery dissection. MRI and angiographic study could ascertain the diagnosis. PMID- 10849642 TI - [Behavioral aspects of chronic pain syndromes]. AB - The knowledge of biological pain mechanisms are not sufficient for the understanding of patients with chronic pain syndromes such as low back, cervicobrachial and muscle pain. Psychological and psychosocial aspects play important roles in the setting and perpetuation of symptoms. Mood and anxiety disorders, secondary gains such as early retirement and financial compensations, must all be acknowledged by the physician as possible contributors to the symptoms. Abnormal illness behavior may better characterize patients with chronic pain syndromes. Behavior observation, which is akin to medical practice, is therefore a powerful tool in the diagnosis and management of these syndromes. Physicians ought be very careful in not reinforcing the patients already strong organic convictions regarding their symptoms, avoiding making decisions based on patients complaints and alleged disabilities, and assigning poorly defined and disputable diagnosis labels. Society needs also to refrain from policies that encourage abnormal illness behaviors. PMID- 10849643 TI - [D8/17 marker: implication to neuropsychiatry]. AB - Recent studies suggest that there is a relationship between rheumatic fever (RF) and some neuropsychiatric disorders. Thus, it has been thought that autoimmune mechanisms might be related to the etiology of these neuropsychiatric disorders. It has also been demonstrated that a B cell alloantigen associated to RF is also abnormally overexpressed in patients with such neuropsychiatric disorders. This alloantigen is recognised by a monoclonal antibody known as D8/17. The aim of this article is to introduce the recent work done about D8/17 and its possible implications to the study of neuropsychiatric disorders related or not to RF. PMID- 10849644 TI - [Guidelines for the treatment of migraine crisis. Consensus of the Sociedade Brasileira de Cefaleia. Ad Hoc Committee]. PMID- 10849645 TI - A tribute to Robert W. Torrance. PMID- 10849646 TI - Plasticity and multiplicity in the mechanisms of oxygen sensing. PMID- 10849647 TI - Evolution of human hypoxia tolerance physiology. AB - Analysis of human responses to hypobaric hypoxia in different lineages (lowlanders, Andean natives, Himalayan natives, and East Africans) indicates 'conservative' and 'adaptable' physiological characters involved in human responses to hypoxia. Conservative characters, arising by common descent, dominant and indeed define human physiology, but in five hypoxia response systems analyzed, we also found evidence for 'adaptable' characters at all levels of organization in all three high altitude lineages. Since Andeans and Himalayans have not shared common ancestry with East Africans for most of our species history, we suggest that their similar hypoxia physiology may represent the 'ancestral' condition for humans--an interpretation consistent with recent evidence indicating that our species evolved under 'colder, drier, and higher' conditions in East Africa where the phenotype would be simultaneously advantageous for endurance performance and for high altitude hypoxia. It is presumed that the phenotype was retained in low capacity form in highlanders and in higher capacity form in most lowland lineages (where it would be recognized by most physiologists as an endurance performance phenotype). Interestingly, it is easier for modern molecular evolution theory to account for the origin of 'adaptable' characters through positive selection than for conserved traits. Many conserved physiological systems are composed of so many gene products that it seems difficult to account for their unchanging state (for unchanging structure and function of hundreds of proteins linked in sequence to form the physiological system) by simple models of stabilizing selection. PMID- 10849648 TI - Comparative aspects of high-altitude adaptation in human populations. AB - The conditions and duration of high-altitude residence differ among high-altitude populations. The Tibetan Plateau is larger, more geographically remote, and appears to have been occupied for a longer period of time than the Andean Altiplano and, certainly, the Rocky Mountain region as judged by archaeological, linguistic, genetic and historical data. In addition, the Tibetan gene pool is less likely to have been constricted by small numbers of initial migrants and/or severe population decline, and to have been less subject to genetic admixture with lowland groups. Comparing Tibetans to other high-altitude residents demonstrates that Tibetans have less intrauterine growth retardation better neonatal oxygenation higher ventilation and hypoxic ventilatory response lower pulmonary arterial pressure and resistance lower hemoglobin concentrations and less susceptibility to CMS These findings are consistent with the conclusion that "adaptation" to high altitude increases with time, considering time in generations of high-altitude exposure. Future research is needed to compare the extent of IUGR and neonatal oxygenation in South American high-altitude residents of Andean vs. European ancestry, controlling for gestational age and other characteristics. Another fruitful line of inquiry is likely to be determining whether persons with CMS or other altitude-associated problems experienced exaggerated hypoxia during prenatal or neonatal life. Finally, the comparison of high-altitude populations with respect to the frequencies of genes involved in oxygen sensing and physiologic response to hypoxia will be useful, once candidate genes have been identified. PMID- 10849649 TI - Tibetan and Andean contrasts in adaptation to high-altitude hypoxia. AB - High-altitude environments provide natural experimental settings to investigate adaptation to environmental stress. An important evolutionary and functional question is whether sea-level human biology constrains the adaptive response. This paper presents evidence that indigenous populations of the Tibetan and Andean plateaus exhibit quantitatively different responses to hypobaric hypoxic stress. At the same altitude, Tibetan mean resting ventilation and hypoxic ventilatory response were more than one-half standard deviation higher than Andean Aymara means while Tibetan mean oxygen saturation and hemoglobin concentration were more than one standard deviation below the Andean means. Quantitative genetic analyses of the familial patterning of these traits provided indirect evidence of population differences in genes influencing them. The Tibetan and Andean patterns of oxygen transport appear equally effective functionally as evaluated by birthweight and maximal aerobic capacity across a range of altitudes. PMID- 10849650 TI - A genomic model for differential hypoxic ventilatory responses. AB - Inbred mice are routinely used as genetic models in lung biology. Among many phenotypic differences in lung function and structure, C3H/HeJ (C3) and C57BL/6J (B6) inbred mice also demonstrate a significantly different ventilatory pattern during acute hypoxic challenge. The present study rejects the hypothesis that a genomic basis for differential hypoxic ventilatory responses (HVR) is linked to loci which determine differential breathing pattern at baseline, while proposing an alternative genetic model for HVR variation. Twelve BXH recombinant inbred (RI) strains derived from C3 and B6 progenitors were examined to enumerate the genes regulating differential HVR. In each of 134 mice, HVR was assessed using whole-body plethysmography to measure tidal volume (VT) and breathing frequency (f). With respect to f during hypoxia, three distinct and reproducible phenotypes are evident in the BXH RI strain distribution pattern (SDP). The SDP for hypoxic f is consistent with the hypothesis that parental strain differences are regulated by two genes. Cosegregation analysis suggest that the genetic control of f during hypoxia differs from the genes which control differential baseline f. Although the genetic control of VT appears more complex, differences in the minute ventilation (VE) during hypoxia is determined by VT. Therefore, this study suggests that the phenotypic variation in HVR between C3 and B6 parental strains, especially related to f during hypoxia, is regulated by as few as two major genetic determinants. PMID- 10849651 TI - Regulation of the hypoxia-inducible factor-1 alpha. ARNT is not necessary for hypoxic induction of HIF-1 alpha in the nucleus. AB - Hypoxia-inducible factor-1 (HIF-1) is a master regulator of mammalian oxygen homeostasis. HIF-1 consists of two subunits, HIF-1 alpha and the aryl hydrocarbon receptor nuclear translocator (ARNT). Whereas hypoxia prevents ubiquitination and proteasomal degradation of HIF-1 alpha, ARNT expression is thought to be oxygen independent. We and others previously showed that ARNT is indispensable for HIF-1 DNA-binding and transactivation function. To examine the requirement of ARNT for accumulation and nuclear translocation of HIF-1 alpha in hypoxia, we used ARNT mutant mouse hepatoma and ARNT-deficient embryonic stem cells. As shown by immunofluorescence, HIF-1 alpha accumulation in the nucleus of hypoxic cells did not require ARNT, demonstrating that nuclear translocation is intrinsic to HIF-1 alpha. During biochemical separation, both HIF-1 alpha and ARNT tend to leak from the nuclei in the absence of the corresponding subunit, suggesting that heterodimerization is required for stable association within the nuclear compartment. Nuclear stabilization of the heterodimer might also explain the hypoxically increased total cellular ARNT levels observed in some of the cell lines examined. PMID- 10849652 TI - Intracellular pathways linking hypoxia to activation of c-fos and AP-1. AB - Organisms respond to hypoxia through detection of blood oxygen levels by sensors at peripheral chemoreceptors and by receptors in certain key cells of the body. The pathways over which peripheral chemoreceptor signals are transmitted to respiratory muscles are well established. However, the intracellular pathways that transmit hypoxic stimulus to gene activation are just being identified. Using anti-sense c-fos strategy, we have shown that c-fos is essential for the activation of activator protein-1 transcription factor complex (AP-1) and subsequent stimulation of downstream genes such as tyrosine hydroxylase (TH; Mishra et al. 1998). The purpose of the present study was to identify intracellular pathways that link hypoxia to activation of c-fos. The results of the present study show that hypoxia causes Ca2+ influx through L-type voltage gated Ca2+ channels and that hypoxia-induced c-fos gene expression is Ca2+/calmodulin dependent. We also demonstrate that hypoxia activates the extracellular-regulated kinase (ERK) and p38, but not JNK. Further, phosphorylation of ERK is essential for c-fos activation via SRE cis-element. Further characterization of nuclear signalling pathways provides evidence for the involvement of Src, a non receptor protein tyrosine kinase, and Ras, a small G protein, in the hypoxia-induced c-fos gene expression. These results suggest a possible role for non-receptor protein tyrosine kinases in propagating signals from G-protein coupled receptors to the activation of immediate early genes such as c-fos during hypoxia. PMID- 10849653 TI - Hypoxia-induced regulation of mRNA stability. AB - Because molecular oxygen is essential for generating cellular energy in aerobic organisms, and because survival depends on this fundamental requirement for oxygen, all higher organisms have evolved numerous diversely regulated mechanisms to detect and respond to potentially life-threatening occurrences of decreased oxygen availability (hypoxia). While the oxygen-dependent regulation of gene expression involves both transcriptional- and post-transcriptional mechanisms, investigations have focused mainly on mechanisms working at the transcriptional level. In this review, the focus is on a growing body of work that looks at post transcriptional mechanisms acting at a level of mRNA stability. PMID- 10849655 TI - Gene regulation during hypoxia in excitable oxygen-sensing cells: depolarization transcription coupling. PMID- 10849654 TI - Hypoxia, HIF-1, and the pathophysiology of common human diseases. AB - Hypoxia plays a fundamental role in the pathophysiology of common causes of mortality, including ischemic heart disease, stroke, cancer, chronic lung disease, and congestive heart failure. In these disease states, hypoxia induces changes in gene expression in target organs that either fail to result in adequate adaptation or directly contribute to disease pathogenesis. Hypoxia inducible factor 1 (HIF-1) is a transcriptional activator that is expressed in response to cellular hypoxia and mediates multiple cellular and systemic homeostatic responses to hypoxia. Recent studies have provided evidence that important pathophysiological responses to hypoxia in pulmonary hypertension, myocardial ischemia, and cancer are mediated by HIF-1. Pharmacologic and gene therapy strategies designed to modulate HIF-1 activity may represent a novel and effective therapeutic approach to these common disorders. PMID- 10849656 TI - Regulation of CREB by moderate hypoxia in PC12 cells. AB - The mechanisms by which excitable cells adapt and respond to changes in O2 levels remain largely unknown. We have investigated the effect of hypoxia on the cyclic AMP response element binding protein (CREB) transcription factor. PC12 cells were exposed to moderate levels of hypoxia (5% O2) for various times between 20 min and 6 hr. We found that hypoxia rapidly and persistently induced ser133 phosphorylation of CREB. This effect was more robust than that produced by exposing PC12 cells to either forskolin, KCl, or NGF. This effect was not due to activation of any of the previously known CREB kinases, including PKA, CaMK, PKC, p70s6k, or MAPKAP kinase-2. Thus, hypoxia may induce activation of a novel CREB kinase. To test whether phosphorylation of CREB was associated with an activation of CRE-dependent gene expression, cells were transfected with wild type and mutated regions of the 5'-flanking region of the tyrosine hydroxylase (TH) gene fused to a CAT reporter gene. Mutation of the CRE element in a TH reporter gene reduced, but did not abolish, the effects of hypoxia on TH gene expression. However, hypoxia did not induce transactivation of a GAL4-luciferase reporter by a GAL4-CREB fusion protein. Thus, the mechanism by which hypoxia regulates CREB is distinct, and more complex, than that induced by forskolin, depolarization, or nerve growth factor. PMID- 10849657 TI - Reactive oxygen species as regulators of oxygen dependent gene expression. PMID- 10849658 TI - A glycolytic pathway to apoptosis of hypoxic cardiac myocytes. Molecular pathways of increased acid production. PMID- 10849659 TI - Mitochondrial-nuclear crosstalk is involved in oxygen-regulated gene expression in yeast. AB - The expression of several oxygen-regulated nuclear genes in the yeast Saccharomyces cerevisiae is affected by the mitochondrion. Recent evidence suggests two levels of mitochondrial involvement. On the one hand, mitochondrial respiratory function is essential for the anoxic induction of some hypoxic genes. On the other hand, the mitochondrial genome itself functions independently of its respiratory function, in the optimal expression of some aerobic genes. These findings suggest that the mitochondrion release at least two types of 'signals' that function in the expression of oxygen-regulated genes. PMID- 10849660 TI - Rox1 mediated repression. Oxygen dependent repression in yeast. AB - For a large number of oxygen-regulated genes in the facultative aerobe, Saccharomyces cerevisiae, the presence of oxygen is sensed through the ability to use oxygen for heme biosynthesis. Heme induces the transcription of oxygen induced genes and represses the transcription of hypoxic genes. Repression is mediated by the Rox1 protein in conjunction with the Ssn6/Tup1 general repression complex. The differential repression of hypoxic genes results from a combination of the tightness of Rox1 binding to the regulatory region of specific hypoxic genes and the presence or absence of binding sites for Mot3 which enhances Rox1 repression. PMID- 10849661 TI - Oxygen dependence of expression of cytochrome C and cytochrome C oxidase genes in S. cerevisiae. PMID- 10849662 TI - Hypoxic and redox inhibition of the human cardiac L-type Ca2+ channel. PMID- 10849663 TI - Molecular identification of O2 sensors and O2-sensitive potassium channels in the pulmonary circulation. AB - Small, muscular pulmonary arteries (PAs) constrict within seconds of the onset of alveolar hypoxia, diverting blood flow to better-ventilated lobes, thereby matching ventilation to perfusion and optimizing systemic PO2. This hypoxic pulmonary vasoconstriction (HPV) is enhanced by endothelial derived vasoconstrictors, such as endothelin, and inhibited by endothelial derived nitric oxide. However, the essence of the response is intrinsic to PA smooth muscle cells in resistance arteries (PASMCs). HPV is initiated by inhibition of the Kv channels in PASMCs which set the membrane potential (EM). It is currently uncertain whether this reflects an initial inhibitory effect of hypoxia on the K+ channels or an initial release of intracellular Ca2+, which then inhibits K+ channels. In either scenario, the resulting depolarization activates L-type, voltage gated Ca2+ channels, which raises cytosolic calcium levels [Ca2+]i and causes vasoconstriction. Nine families of Kv channels are recognized from cloning studies (Kv1-Kv9), each with subtypes (i.e. Kv1.1-1.6). The contribution of an individual Kv channel to the whole-cell current (IK) is difficult to determine pharmacologically because Kv channel inhibitors are nonspecific. Furthermore, the PASMC's IK is an ensemble, reflecting activity of several channels. The K+ channels which set EM, and inhibition of which initiates HPV, conduct an outward current which is slowly inactivating, and which is blocked by the Kv inhibitor 4 aminopyridine (4-AP) but not by inhibitors of Ca(2+)- or ATP-sensitive K+ channels. Using anti-Kv antibodies to immunolocalize and inhibit Kv channels, we showed that the PASMC contains numerous types of Kv channels from the Kv1 and Kv2 families., Furthermore Kv1.5 and Kv2.1 may be important in determining the EM and play a role as effectors of HPV in PASMCs. While the Kv channels in PASMCs are the "effectors" of HPV, it is uncertain whether they are intrinsically O2 sensitive or are under the control of an "O2 sensor". Certain Kv channels are rich in cysteine, and respond to the local redox environment, tending to open when oxidized and close when reduced. Candidate sensors vary the PASMC redox potential in proportion to O2. These include Nicotinamide Adenine Dinucleotide Phosphate Oxidase, (NADPH oxidase) and the cytosolic ratio of reduced/oxidized redox couples (i.e. glutathione GSH/GSSG), as controlled by electron flux in the mitochondrial electron transport chain (ETC). Using a mouse that lacks the gp91phox component of NADPH oxidase, we have recently shown that loss of the gp91phox-containing NADPH oxidase as a source of activated oxygen species does not impair HPV. However, inhibition of complex 1 of the mitochondrial electron transport chain mimics hypoxia in that it inhibits IK, reduces the production of activated O2 species and causes vasoconstriction. We hypothesize that a redox O2 sensor, perhaps in the mitochondrion, senses O2 through changes in the accumulation of freely diffusible electron donors. Changes in the ratio of reduced/oxidized redox couples, such as NADH/NAD+ and glutathione (GSH/GSSG) can reduce or oxidize the K+ channels, resulting in alterations of PA tone. PMID- 10849664 TI - Chemosensing at the carotid body. Involvement of a HERG-like potassium current in glomus cells. AB - Currently, it is not clear what type of K+ channel(s) is active at the resting membrane potential (RMP) in glomus cells of the carotid body (CB). HERG channels produce currents that are known to contribute to the RMP in other neuronal cells. The goal of the present study was to determine whether CB glomus cells express HERG-like (HL) K+ current, and if so, to determine whether HL currents regulate the RMP. With high [K+]o, depolarizing voltage steps from -85 mV revealed a slowly deactivating inward tail current indicative of HL K+ current in whole cell, voltage clamped glomus cells. The HL currents were blocked by dofetilide (DOF) in a concentration-dependent manner (IC50 = 13 nM) and high concentrations (1 and 10 mM) of Ba2+. The steady-state activation properties of the HL current (Vh = -45 mV) suggest that it is active at the RMP in glomus cells. Whole-cell, current clamped glomus cells exhibited a RMP of -48 mV. 150 nM DOF caused a significant (14 mV) depolarizing shift in the RMP. In isolated glomus cells, [Ca2+]i increased in response to DOF (1 microM). In an in-vitro CB preparation, DOF increased basal sensory discharge in a concentration-dependent manner and significantly attenuated the sensory response to hypoxia. These results suggest that the HERG-like current is responsible for controlling the RMP in glomus cells of the rabbit CB, and that it is involved in the chemosensory response to hypoxia of the CB. PMID- 10849665 TI - Oxidant signalling and vascular oxygen sensing. Role of H2O2 in responses of the bovine pulmonary artery to changes in PO2. PMID- 10849666 TI - Tissue PO2 and mitochondrial enzymes. Cytochrome C oxidase as O2 sensor. PMID- 10849667 TI - Regulation of Shaker-type potassium channels by hypoxia. Oxygen-sensitive K+ channels in PC12 cells. AB - Little is known about the molecular composition of the O2-sensitive K+ (Ko2) channels. The possibility that these channels belong to the Shaker subfamily (Kv1) of voltage-dependent K+ (Kv) channels has been raised in pulmonary artery (PA) smooth muscle cells. Numerous findings suggest that the Ko2 channel in PC12 cells is a Kv1 channel, formed by the Kv1.2 alpha subunit. The Ko2 channel in PC12 cells is a slow-inactivating voltage-dependent K+ channel of 20 pS conductance. Other Kv channels, also expressed in PC12 cells, are not inhibited by hypoxia. Selective up-regulation by chronic hypoxia of the Kv1.2 alpha subunit expression correlates with an increase O2-sensitivity of the K+ current. Other Kv1 alpha subunit genes encoding slow-inactivating Kv channels, such as Kv1.3, Kv2.1, Kv3.1 and Kv3.2 are not modulated by chronic hypoxia. The Ko2 current in PC12 cells is blocked by 5 mM externally applied tetraethylammonium chloride (TEA) and by charydbotoxin (CTX). The responses of the Kv1.2 K+ channel to hypoxia have been studied in the Xenopus oocytes and compared to those of Kv2.1, also proposed as Ko2 channel in PA smooth muscle cells. Two-electrode voltage clamp experiments show that hypoxia induces inhibition of K+ current amplitude only in oocytes injected with Kv1.2 cRNA. These data indicate that Kv1.2 K+ channels are inhibited by hypoxia. PMID- 10849668 TI - HIF-1 is essential for multilineage hematopoiesis in the embryo. PMID- 10849669 TI - Dual influence of nitric oxide on gene regulation during hypoxia. AB - It is being increasingly recognized that nitric oxide (NO) is associated with many physiological processes, including regulation of gene expression. NO shares certain similarities with molecular oxygen (O2). Previous studies have shown that hypoxia up-regulates c-fos, an immediate early gene, and tyrosine hydroxylase (TH), a late response gene that encodes rate limiting enzyme in catecholamine synthesis. Given the similarities between NO and O2, we hypothesized that NO inhibits hypoxia-induced up-regulation of c-fos and TH. Experiments were performed on rat pheochromocytoma (PC12) cells. c-fos and TH mRNA's were analysed by Northern blot and promoter activities by reporter gene assays, respectively. Hypoxia (1% O2 for 6 h) up-regulated c-fos and TH mRNA and increased c-fos promoter activity. Hypoxia-induced c-fos mRNA expression, and promoter activities were significantly potentiated in presence of spermine nitric oxide (SNO), a NO donor. By contrast, SNO significantly inhibited TH mRNA expression and TH promoter activity during hypoxia. Electrophoretic mobility shift-assay showed increased binding of AP-1 and HIF-1 transcription factors to the TH promoter in cells exposed to hypoxia. SNO abolished the binding of AP-1 and HIF-1 to the TH promoter during hypoxia, suggesting that inhibition of hypoxia-induced TH transcription by NO are due to reduced binding of AP-1 and HIF-1 transcription factors. These result demonstrate that NO has both positive and negative influence on gene regulation by hypoxia and suggest that although NO resembles O2 does not always inhibit gene expression during low oxygen. PMID- 10849670 TI - Hypoxia differentially regulates the mitogen- and stress-activated protein kinases. Role of Ca2+/CaM in the activation of MAPK and p38 gamma. AB - Hypoxic/ischemic trauma is a primary factor in the pathology of various vascular, pulmonary, and cerebral disease states. Yet, the signaling mechanisms by which cells respond and adapt to changes in oxygen levels are not clearly established. The effects of hypoxia on the stress- and mitogen-activated protein kinase (SAPK and MAPK) signaling pathways were studied in PC12 cells. Exposure to moderate hypoxia (5% O2) was found to progressively stimulate phosphorylation and activation of p38 gamma in particular, and also p38 alpha, two isoforms of the p38 family of stress-activated protein kinases. In contrast, hypoxia had no effect on enzyme activity of p38 beta, p38 beta 2, p38 delta, or on JNK, another stress-activated protein kinase. Prolonged hypoxia also induced phosphorylation and activation of p42/p44 MAPK, although this activation was modest when compared to NGF and UV-induced activation. We further showed that activation of p38 gamma and MAPK during hypoxia requires calcium, as treatment with Ca(2+)-free media or the calmodulin antagonist, W13, blocked the activation of p38 gamma and MAPK, respectively. These studies demonstrate that an extremely typical physiological stress (hypoxia) causes selective activation of specific elements of the SAPKs and MAPKs, and identifies Ca+2/CaM as a critical upstream activator. PMID- 10849671 TI - Chairman's summary: mechanisms of oxygen homeostasis, circa 1999. AB - Oxygen sensing is a fundamental physiologic requirement of all living organisms and all cells within the human body. This paper presents a brief summary of recent investigations into the molecular mechanisms underlying oxygen sensing and adaptive responses to hypoxia, with particular reference to other papers in this volume. PMID- 10849672 TI - Oxygen, homeostasis, and metabolic regulation. AB - Even a cursory review of the literature today indicates that two views dominate experimental approaches to metabolic regulation. Model I assumes that cell behavior is quite similar to that expected for a bag of enzymes. Model II assumes that 3-D order and structure constrain metabolite behavior and that metabolic regulation theory has to incorporate structure to ever come close to describing reality. The phosphagen system may be used to illustrate that both approaches lead to very productive experimentation and significant advances are being made within both theoretical frameworks. However, communication between the two approaches or the two 'groups' is essentially nonexistent and in many cases (our own for example) some experiments are done in one framework and some in the other (implying some potential schizophrenia in the field). In our view, the primary paradox and problem which no one has solved so far is that essentially all metabolite concentrations are remarkably stable (are homeostatic) over large changes in pathway fluxes. For muscle cells O2 is one of the most perfectly homeostatic of all even though O2 delivery and metabolic rate usually correlate in a 1:1 fashion. Four explanations for this behavior are given by traditional metabolic regulation models. Additionally, there is some evidence for universal O2 sensors which could help to get us out of the paradox. In contrast, proponents of an ultrastructurally dominated view of the cell assume intracellular perfusion or convection as the main means for accelerating enzyme-substrate encounter and as a way to account for the data which have been most perplexing so far: the striking lack of correlation between changes in pathway reaction rates and changes in concentrations of pathway substrates and intermediates, including oxygen. The polarization illustrated by these two views of living cells extends throughout the metabolic regulation field (and has caused the field to progress along two surprisingly independent paths with minimal communication between them). The time may have come when cross talk between the two fields may be useful. PMID- 10849673 TI - Evidence that nitric oxide plays a role in O2 sensing from tissue NO and PO2 measurements in cat carotid body. PMID- 10849674 TI - Carotid body gap junctions: secretion of transmitters and possible electric coupling between glomus cells and nerve terminals. AB - It is proposed that intercellular coupling between glomus cells and carotid nerve terminals form an integral part of the chemoreceptor process. Coupling is possible because gap junctions occur between these elements. At rest, most glomus cells would be coupled. Stimuli uncouple (or reduce coupling) most glomus cells that extrude their contents toward the nerve terminals. However, other glomus cells do not secrete but recharge and intercellular coupling increases. These phenomena would allow for sustained chemoreceptor activity during prolonged stimulation. Coupling between glomus and sustentacular cells may explain why the behavior of glomus cells in the intact carotid body and when clustered in cultures (when their sustentacular envelope is preserved) is different from that of isolated cells where sustentacular cells are destroyed. The presence of electric synapses between glomus cells and nerve terminals may explain the poor performance of synaptic blockers on natural (hypoxia, hypercapnia, acidity) carotid body stimulation. PMID- 10849675 TI - Short- and long-term regulation of rat carotid body gap junctions by cAMP. Identification of connexin43, a gap junction subunit. AB - Intact and cultured carotid bodies (CBs) of the rat were used in this study. Applications of membrane-permeant db-cAMP to cultured carotid bodies increased electric coupling between most glomus cells (increasing junctional conductance) probably by opening preformed intercellular channels. This a short-term effect of the nucleotide, increasing gating between glomus cells. When cultures and intact carotid bodies were treated with membrane-permeant 8Br-cAMP for 3 h or more (to increase cytosolic cAMP), there was enhanced gap junction formation and better dye spread between carotid body cells. Connexin43 (CX43) was identified by immunocytochemical methods as forming part of the intercellular channels between carotid body cells, and the expression of Cx43 increased by cAMP. This is a long term effect, inducing the formation of gap junctions. Thus, cAMP had short and long-term effects on the intercellular junctions of the carotid body. Long-term formation of gap junctions may be important in modulating carotid body functions during stimulation by chronic hypoxia. PMID- 10849676 TI - Subcellular localization and function of B-type cytochromes in carotid body and other paraganglionic cells. PMID- 10849677 TI - Acetylcholine sensitivity of cat petrosal ganglion neurons. AB - We investigated if neuronal nicotinic acetylcholine receptors (nAChRs) are localized in chemoreceptor afferent neurons in the cat petrosal ganglion (PG) and if acetylcholine (ACh) excites chemoreceptor afferent neurons. Immunocytochemistry revealed that a majority of PG neurons expressed alpha 4 and/or alpha 7 subunits of neuronal nAChRs, and a part of them were tyrosine hydroxylase positive. Excitability of cultured PG neurons was studied with patch clamp techniques (whole cell configuration). ACh and nicotine evoked both inward and outward currents. The inward current was partially blocked by removal of extracellular calcium and by antagonists for alpha 4 beta 2 (dihydro-beta erythroidine) or alpha 7 nAChRs (methyllycaconitine). Outward current was blocked by 4-aminopyridine (4-AP) and sometimes by atropine. ACh-induced membrane potential changes were well correlated with ACh-induced currents. Depolarization and hyperpolarization occurred in response to ACh. Occasionally depolarization was followed by a train of action potentials. The results suggest that heterogeneous neuronal nAChRs are widely distributed in both chemoreceptor and other PG neurons. In some neurons nAChRs may be functionally coupled with outward K+ channels. Further studies are required to determine whether chemoreceptor neurons have a distinct distribution pattern of nAChRs and K+ channels. PMID- 10849678 TI - Responses of petrosal ganglion neurons in vitro to hypoxic stimuli and putative transmitters. PMID- 10849679 TI - The metabolic hypothesis revisited. AB - High levels of CO are used to mimic the stimulatory response of the CSN initiated by hypoxia. Using light of different wavelengths we show that the stimulatory effects of high CO can be pinpointed to the cytochrome c oxidase in the mitochondrial respiratory chain. This supports the metabolic theory of oxygen sensing in the mitochondria. PMID- 10849681 TI - The present status of the mechanical hypothesis for chemoreceptor stimulation. AB - Reasons are given to show why the transmitter based hypothesis for the stimulation of chemoreceptors needs to be reviewed. On the other hand evidence is presented to show that chemoreceptors can be stimulated by various mechanical stimuli and how the local PO2 can be sensed by the type II cell which by getting mechanically deformed causes this cell to shrink. This shrinkage is transmitted to the generator region of the nerve terminal thereby leading to the production of propagated impulses at the regenerative region thus making the whole process of generation of information about the local PO2, similar to the generation of sensory information by other sensory receptors. PMID- 10849680 TI - Effect of adenosine on CO2 chemosensitivity. Functional, cellular, and molecular studies. PMID- 10849682 TI - Identification of an oxygen-sensitive potassium channel in neonatal rat carotid body type I cells. PMID- 10849683 TI - Significance of ROS in oxygen chemoreception in the carotid body chemoreception. Apparent lack of a role for NADPH oxidase. PMID- 10849684 TI - ATP-dependent K+ and voltage-gated Ca2+ channels in endothelial cells of brain capillaries. Effect of hypoxia. PMID- 10849685 TI - Different O2-sensing mechanisms by different K+ channels. PMID- 10849686 TI - Response of intracellular pH to acute anoxia in individual neurons from chemosensitive and nonchemosensitive regions of the medulla. AB - The effect of acute (10 minutes) exposure to anoxia on intracellular pH (pHi) in individual brainstem neurons, in slices from neonatal (P7 to P11) rats, was studied using a fluorescence microscopy imaging technique. Neurons from 4 regions of the medulla were studied, two of which contained chemosensitive neurons (nucleus tractus solitarius, NTS, and ventrolateral medulla, VLM) and two regions which did not contain chemosensitive neurons (hypoglossal, Hyp, and inferior olivary, IO). Acute anoxia caused a rapid and maintained acidification of 0.1-0.3 pH unit that was not different in neurons from chemosensitive vs. nonchemosensitive regions. Blocking the contribution of Na+/H+ exchange (NHE) to pHi regulation by exposing neurons to acute anoxia in the presence of the exchange inhibitor amiloride (1 mM) did not affect the degree of acidification seen in neurons from the NTS and VLM region, but significantly increased acidification (to about 0.35 pH unit) in Hyp and IO neurons. In summary, anoxia induced intracellular acidification is not different between neurons from chemosensitive and nonchemosensitive regions, but NHE activity blunts acidification in neurons from the latter regions. These data suggest that neurons from chemosensitive areas might have a smaller acid load in response to anoxia than neurons from nonchemosensitive regions of the brainstem. PMID- 10849687 TI - Hyperbaric oxygen depolarizes solitary complex neurons in tissue slices of rat medulla oblongata. AB - Hyperbaric oxygen (HBO2) at approximately 3 atmospheres absolute (ATA) pressure is toxic to the mammalian CNS due to excessive O2 free radical production. No study has ever determined the effects of < or = 3 ATA of O2 on the membrane potential and firing rate of neurons in the mammalian brainstem. Likewise, no study has ever determined the effects of < or = 3 ATA pressure per se on brainstem neurons. Accordingly, we initiated intracellular recordings at 1 ATA in solitary complex neurons in slices (300 microns) of rat caudal medulla oblongata that were maintained inside a 72 liter hyperbaric chamber. Helium, which is inert and without narcotic effect at moderate levels of hyperbaria, was used to hydrostatically compress the submerged brain slice to determine the effects of pressure per se. Tissue oxygen tension and extracellular pH were also measured during exposure to hyperbaric gases. Six of 19 neurons were affected by hyperbaric helium; 5 cells were depolarized and 1 cell was hyperpolarized. Input resistance (Rin) either increased (n = 1) or decreased (n = 3). When control perfusate (0.95 ATA O2) was switched to perfusate saturated with 98% O2 (balance CO2, pH = 7.3-7.4, pO2 = 2.5-3.4 ATA; 2-18 minutes of exposure) in a separate pressure vessel, 8 of 13 neurons were depolarized and 5 neurons were insensitive. In the 8 O2-responsive neurons, Rin either increased (n = 5), decreased (n = 2) or was unchanged (n = 1). Three of 8 neurons depolarized by HBO2 were also depolarized by hyperbaric helium, usually with an additional change in Rin. We conclude that hydrostatic (helium) pressure and HBO2 independently increase excitability in certain solitary complex neurons. We hypothesize that these responses contribute, in part, to neural events that either precede or occur during CNS O2 toxicity. PMID- 10849688 TI - Chronic hypoxia induces changes in the central nervous system processing of arterial chemoreceptor input. AB - Chronic hypoxia increases the hypoxic ventilatory response (HVR) in awake rats and the phrenic nerve response to carotid sinus nerve stimulation in anesthetized rats. An increased O2 sensitivity of the arterial chemoreceptors contributes to the increase in the HVR, but changes in the CNS processing of afferent information from arterial chemoreceptors are also involved. Adult male Sprague Dawley rats were exposed to 0-7 days of hypobaric hypoxia (PIO2 = 80 Torr). Ventilation was measured in rats exposed to 0, 2 and 7 days of hypoxia using whole-body plethysmography. Ventilation increased after 2 days and remained elevated after 7 days of hypoxia. Following dopamine D2 receptor (D2-R) blockade in the CNS, frequency significantly decreased after 0 and 7 days of hypoxia, but did not change significantly after 2 days of hypoxia. In anesthetized rats, the phrenic nerve response to carotid sinus nerve stimulation was reduced following systemic D2-R blockade in control rats and those exposed to 7 days of hypoxia. After 2 days of hypoxia, there was no effect of blocking systemic D2-R. To determine whether changes in D2-R mRNA precede physiological changes, competitive RT-PCR was used to quantify D2-R mRNA in micropunches from the nucleus tractus solitarius (NTS) in normoxic and chronically hypoxic rats. In hypoxia, D2-R mRNA in the caudal NTS initially increased (6-12 hours) and then decreased below control levels (24 hours-7 days). These results show that chronic hypoxia causes time-dependent changes in D2-R that could result in changes in the ventilatory response to hypoxia. PMID- 10849689 TI - Acetylcholine is released from in vitro cat carotid bodies during hypoxic stimulation. AB - Previous pharmacological, immunocytochemical, electrophysiological, and microfluorometric studies have suggested that acetylcholine (ACh) is a critically important excitatory transmitter in the chemotransduction of hypoxia by the cat carotid body (CB). With the use of HPLC this study shows that the in vitro cat CB releases ACh under normoxic conditions; this release is increased when the CB is challenged with hypoxia. The preliminary observation that greater amounts of ACh are liberated in the presence of gallamine and AFDX116 suggests the presence of functioning M2 muscarinic receptors on the glomus cells of the CB. PMID- 10849690 TI - Interactions between acetylcholine and dopamine in chemoreception. PMID- 10849691 TI - Interaction between catecholamines and neuropeptides in the carotid body: evidence for dopamine modulation of neutral endopeptidase activity. AB - Carotid body (CB) contains multiple neurochemicals that include catecholamines (CA) and neuropeptides. They are involved in the modulation of sensory response of the carotid body. Based on observations from the central nervous system, we hypothesized that CA modulates neuropeptide metabolism in CB. To test our hypothesis, fetal calf carotid body model was used. Immunocytochemical analysis showed that fetal calf carotid body expresses both tyrosine hydroxylase, and neutral endopeptidase-like immunoreactivity. To assess the effect of CA, thin slices of fetal calf carotid body were incubated with 50-500 microM of dopamine (DA) at 37 degrees C for 1 hour. As an index of neuropeptide metabolism, the activity of neutral endopeptidase (NEP), a major degrading enzyme of neuropeptides in CB was determined in the membrane-enriched and soluble fractions of the carotid body. CBs incubated with medium lacking DA served as control. On average, NEP activities of the membrane-enriched, and soluble fractions of the untreated CB were 4.8 +/- 0.2 and 6.7 +/- 0.2 pmole per hour per mg of CB respectively. At concentrations less than 200 microM, DA enhanced NEP activity of the membrane fraction (approximately 60%) whereas inhibition of NEP was observed in the soluble fraction (approximately 62%). At concentrations > 200 microM, DA inhibited NEP activity of the two fractions. When CBs were incubated with DA in the presence of sodium dithionite, an oxygen scavenger, DA, even at higher concentrations, stimulated NEP activity of the membrane-enriched fraction. The above results demonstrate that DA modulates neuropeptide metabolism in CB via a non-receptor-mediated mechanism involving a direct interaction with NEP. PMID- 10849692 TI - Pharmacological effects of endothelin in rat carotid body. Activation of second messenger pathways and potentiation of chemoreceptor activity. PMID- 10849693 TI - Oxygen and acid chemoreception by pheochromocytoma (PC12) cells. PMID- 10849694 TI - Postnatal changes in cardiovascular regulation during hypoxia. PMID- 10849695 TI - Expression and localization of A2a and A1-adenosine receptor genes in the rat carotid body and petrosal ganglia. A2a and A1-adenosine receptor mRNAs in the rat carotid body. PMID- 10849696 TI - Serotonin and the hypoxic ventilatory response in awake goats. PMID- 10849697 TI - Peripheral chemosensitivity in mutant mice deficient in nitric oxide synthase. AB - Nitric oxide (NO) is endogenously generated from two constitutively expressed nitric oxide synthase (NOS) isoforms, i.e., neuronal (NOS-1) and endothelial (NOS 3). Both isoforms are localized within the carotid body. Previous studies have shown endogenously generated NO modulates carotid body activity. In the present study, we examined the relative contribution of NO generated by NOS-1 and NOS-3 in respiratory reflexes arising from the carotid body. Experiments were performed on mutant mice deficient in NOS-1 or NOS-3. Wild-type (WT) mice, which contained both isoforms, served as controls. Respiration was monitored in unanesthetized mice by plethysmography. In anaesthetized mice, efferent phrenic nerve activity was monitored as index of breathing. We examined the effects of hypoxia (12% O2), cyanide and brief hyperoxia (Dejour's test) on respiration. In NOS-1 mutant mice, the ventilatory response to hypoxia (12% O2) were significantly augmented, compared to wild-type (WT) mice. By contrast, NOS-3 mutant mice displayed significantly blunted respiratory responses to hypoxia compared to WT controls. The responses to cyanide were augmented in NOS-1; whereas they were blunted in NOS-3 mutant mice. Respiratory depression in response to brief hyperoxia was more pronounced in NOS-1, while it was nearly absent in NOS-3 mutant mice. These results demonstrate that NO produced by the neuronal and endothelial NOS isoforms have different modulatory roles in carotid body chemosensitivity. PMID- 10849698 TI - Dopaminergic excitation in goat carotid body may be mediated by serotonin receptors. PMID- 10849699 TI - Augmentation of calcium current by hypoxia in carotid body glomus cells. AB - Several lines of evidence indicate that transduction of the hypoxic stimulus at the carotid body involves an increase in cytosolic Ca2+ ([Ca2+]i) via activation of voltage-gated Ca2+ channels in the glomus cells. However, reported responses to hypoxia include either no effect on or inhibition of Ca2+ current in glomus cells. The apparent discrepancy between the effects of hypoxia on [Ca2+]i and Ca2+ channel activity prompted us to re-examine the effects of low oxygen on Ca2+ currents in glomus cells. Experiments were performed on freshly dissociated glomus cells from rabbit carotid bodies. Ca2+ channel activity was monitored using the whole-cell configuration of the patch clamp technique with Ba2+ as the charge carrier. Hypoxia (pO2 = 40 mmHg) augmented the Ca2+ current by 24% (at 0 mV). This augmentation was seen in a CO2/HCO3- but not in a HEPES buffered extracellular solution. However, when the extracellular pH (pHo) of a HEPES buffered solution is lowered from 7.4 to 7.0, then the Ca2+ current in glomus cells is augmented by hypoxia by 20%. Nisoldipine, an L-type Ca2+ channel blocker (2 microM), prevented augmentation of the Ca2+ current by hypoxia. On the other hand, an N- and P-type Ca2+ channel blocker (2 microM omega-conotoxin MVIIC) did not prevent the augmentation of the Ca2+ current by hypoxia. Protein kinase C (PKC) inhibitors, staurosporine (100 nM) and bisindolylmaleimide (2 microM), prevented augmentation by hypoxia. Okadaic acid (100 nM), an inhibitor of serine/threonine phosphatases also prevented augmentation of Ca2+ current by hypoxia; whereas, norokadaone, an inactive analog of okadaic acid, had no effect. These results suggest that hypoxia augments Ca2+ current through L-type Ca2+ channels via a PKC and/or phosphatase-sensitive pathways. PMID- 10849700 TI - O2-chemosensitivity in developing rat adrenal chromaffin cells. PMID- 10849701 TI - O2-sensing by model airway chemoreceptors. Hypoxic inhibition of K+ channels in H146 cells. PMID- 10849702 TI - Morphological adaptation of the peptidergic innervation to chronic hypoxia in the rat carotid body. PMID- 10849703 TI - Continuous, but not episodic hypoxia, induces CREB phosphorylation in rat carotid body type I cells. PMID- 10849704 TI - Intracellular PO2 of the carotid body. PMID- 10849705 TI - Redox-based inhibition of K+ channel/current is not related to hypoxic chemosensory responses in rat carotid body. PMID- 10849707 TI - Estimation of CO2 chemosensitivity from the carotid body in humans. PMID- 10849706 TI - Effects of 2,4-dinitrophenol (DNP) on the relationship between the chemosensory activities of the rat carotid body and the intracellular calcium of glomus cells. AB - To test the hypothesis that the uncoupler 2,4-dinitrophenol (DNP) increases [Ca2+]i equally well, independent of pHi, we studied the effects of 250 microM DNP on [Ca2+]i and carotid sinus nerve (CSN) activity of rat carotid body (CB). CSN activity was measured in CB perfused and superfused with hypocapnic (pHo 7.80) and normocapnic (pHo 7.42) Tyrode solutions. [Ca2+]i of glomus (type I cells) was assessed by superfusion techniques under identical conditions as for CSN recording experiments. The results indicate that 250 microM DNP increased [Ca2+]i of type I cells as well as CSN activity at both pHos, although alkalosis diminished these responses. Given that pHi will change with pHo, DNP did not make any additional pHi change, although [Ca2+]i changed. We conclude that DNP effects were due to [Ca2+]i change alone, and the relationship between [Ca2+]i and CSN activity are internally consistent. PMID- 10849708 TI - Adenosine-dopamine interactions and ventilation mediated through carotid body chemoreceptors. PMID- 10849709 TI - Carotid body NO-CO interaction and chronic hypoxia. PMID- 10849710 TI - Interplay between the cytosolic Ca2+ increase and potential changes in glomus cells in response to chemical stimuli. PMID- 10849711 TI - Characteristics of carotid body chemosensitivity in the mouse. Baseline studies for future experiments with knockout animals. PMID- 10849712 TI - Role of substance P in neutral endopeptidase modulation of hypoxic response of the carotid body. AB - Carotid body expresses neutral endopeptidase (NEP)-like enzyme activity and phosphoramidon, an inhibitor of NEP augments sensory response of the carotid body to hypoxia (Kumar et al., 1990). NEP hydrolyzes substance P (SP) and methionine enkephalin (Met-ENK) in the nervous system. In the present study, we determined whether NEP hydrolyzes Met-ENK and SP in the carotid body and whether these peptides contribute to the phosphoramidon-induced potentiation of the sensory response to hypoxia. Experiments were performed on carotid bodies excised from anaesthetized adult cats. HPLC analysis showed that both SP and Met-ENK were hydrolyzed by the carotid body. Phosphoramidon (400 microM) markedly inhibited SP (approximately 90%) but had only marginal effect on Met-ENK hydrolysis (approximately 15%). Sensory responses of the carotid body in vitro to hypoxia (pO2, 68 +/- 6 mmHg) and SP (10 nmoles) were potentiated by phosphoramidon by approximately 80% and approximately 275% respectively (p < 0.01). SP-receptor antagonist abolished phosphoramidon-induced potentiation of the sensory response to hypoxia as well as to SP. These results demonstrate that SP is a preferred substrate for NEP in the carotid body and SP plays a major role in the potentiation of the hypoxic response of the carotid body by phosphoramidon. PMID- 10849713 TI - Effect of barium on rat carotid body glomus cell [Ca2+]i and carotid chemosensory response. PMID- 10849714 TI - A dual acid-influx transport system in the carotid body type I cell. Acid influx in carotid body type I cells. PMID- 10849715 TI - L-DOPA and high oxygen influence release of catecholamines from the cat carotid body. AB - Current modelling of carotid body (CB) chemotransduction postulates an essential role for neurotransmitters, including dopamine (DA). Catecholamines (CA) released from incubated/superfused cat CBs has often been reported to diminish rapidly over the course of the exposure. The purpose of the first set of experiments was to determine the effects of including L-dihydroxyphenylalanine (L-DOPA), the immediate precursor to DA, in the incubation medium. CBs were removed from deeply anesthetized cats, cleaned of connective tissue, and placed in separate incubation tubes containing Krebs Ringer Bicarbonate solution (KRB) at 37 degrees C. One tube contained 40 microM L-DOPA. Both tubes were bubbled for 2 hr with a normoxic gas mixture (21% O2/6% CO2). This was followed immediately by a 30 minute exposure to a hypoxic gas mixture (4% O2/5% CO2). The mean amounts of DA, dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and norepinephrine (NE) released during 30 min exposures were always greater when L-DOPA was present. The use of gas mixture like the above normoxic gas mixture in incubation studies has often been considered quasi-hypoxic. Hence, in a second set of experiments we tested the effect of high oxygen mixture (95% O2/5% CO2). All other features of these experiments were the same as the above. The high oxygen environment correlated with lower DA release suggesting a reduced excitation/inhibition. The subsequent exposure to hypoxia, however, provoked a much larger release of DA and NE. The data demonstrate the substantial effect of oxygen on the release of CAs and the apparent need of a DA precursor like L-DOPA to allow detection of the changes in CA release from the CBs upon exposure to a hypoxic stimulus. PMID- 10849716 TI - Effects of a dopamine agonist on cytosolic Ca2+ changes induced by hypoxia in rat glomus cells. PMID- 10849717 TI - Carotid chemoreceptors participation in brain glucose regulation. Role of arginine-vasopressin. PMID- 10849718 TI - Nitric oxide modulation of carotid chemoreception. PMID- 10849719 TI - O2 and respiration in exercising human muscle. The regulation of oxidative phosphorylation in vivo. PMID- 10849720 TI - pH sensitivity in the isolated CNS of newborn mouse. PMID- 10849721 TI - Aortic body chemoreflex of the anaesthetized rat. Electrophysiological, morphological, and reflex studies. PMID- 10849722 TI - Changes in the peptidergic innervation of the rat carotid body a month after the termination of chronic hypoxia. PMID- 10849723 TI - Carotid bodies and the sigh reflex in the conscious and anaesthetised guinea-pig. PMID- 10849724 TI - Immunohistochemical study of the carotid body during hibernation. PMID- 10849725 TI - Neurochemical reorganization of O2 chemoreflex pathway after carotid body denervation in rats. PMID- 10849726 TI - Review: the role of membrane peptidases in immune functions. PMID- 10849727 TI - Structure and function of aminopeptidase N. PMID- 10849728 TI - Modulation of WNT-5A expression by actinonin: linkage of APN to the WNT-pathway? AB - Inhibition of alanyl-aminopeptidase gene expression or enzymatic activity compromises T cell proliferation and function. Molecular mechanisms mediating these effects are not known as yet. Applying the cDNA array technique we identified the proto-oncogen Wnt-5a strongly affected by APN-inhibition. Wnt-5a and other members of the Wnt family of secreted factors are implicated in cell growth and differentiation. Wnt-5a was moderately expressed in resting T cells, but strongly down-regulated in response to activation by OKT3/IL-4/IL-9. Actinonin increased Wnt-5a-mRNA contents as confirmed by RT-PCR. In addition, expression of GSK-3 beta, an inherent component of the Wnt-pathway, was found to be increased in response to activation, but suppressed by actinonin at both the mRNA and protein level. These findings may provide a rationale for the strong growth inhibitory effects resulting from an inhibition of alanyl aminopeptidase expression or activity. PMID- 10849729 TI - Enzymatic activity is not a precondition for the intracellular calcium increase mediated by mAbs specific for aminopeptidase N/CD13. PMID- 10849730 TI - Transforming growth factor-beta increases the expression of aminopeptidase N/CD13 mRNA and protein in monocytes and monocytic cell lines. AB - Aminopeptidase N (APN)/CD13 is a membrane-bound surface ectopeptidase with a ubiquitous distribution. In hematopoiesis, APN/CD13 is expressed on stem cells and during most developmental stages of myeloid cells. Because APN/CD13 has been implicated in the trimming on the cell surface of peptides that protrude out of MHC class II molecules, we wanted to study the regulation of this membrane peptidase in antigen presenting cells by TGF-beta. TGF-beta is a potent inducer of the maturation of monocyte precursors towards a macrophage phenotype. Using competitive RT-PCR and cytofluorimetric analyses, we quantified the modulation of the APN/CD13 mRNA as well as protein expression by TGF-beta 1 and -2 and found a stimulation of the APN/CD13 expression in a time- and dose-dependent manner in monocytic cells. In U937 cells, the time course showed a maximum for APN/CD13 mRNA at 24 hours incubation with TGF-beta. In experiments with actinomycin D- treated cells was found a stabilization of APN/CD13 mRNA by TGF-beta 1. Contrary to the IL-4-induced expression of APN/CD13 as well as of MHC class II in monocytic cells, we could show that TGF-beta is able to augment the APN/CD13 expression but decreases the MHC class II expression. PMID- 10849731 TI - Cell-cell contact between lymphocytes and fibroblast-like synoviocytes induces lymphocytic expression of aminopeptidase N/CD13 and results in lymphocytic activation. PMID- 10849732 TI - Natural substrates of dipeptidyl peptidase IV. PMID- 10849733 TI - Relating structure to function in the beta-propeller domain of dipeptidyl peptidase IV. Point mutations that influence adenosine deaminase binding, antibody binding and enzyme activity. AB - Point mutations in human CD26/DP IV were analysed for adenosine deaminase (ADA) binding, monoclonal antibody (mAb) binding and DP IV enzyme activity. Point mutations at either Leu294 or Val341 ablated ADA binding. Binding by mAbs that inhibit ADA binding was found to involve both Leu340 to Arg343 and Thr440/Lys441. Glu205 and Glu206 were found to be essential for enzyme activity. All residues of interest were mapped onto a model of the beta-propeller domain of DP IV. These data led us to suggest that in DP IV and related peptidases ligand and antibody binding sites are non-linear and that enzyme activity depends on charged sidechains that surround the entrance to the central tunnel of the beta propeller. PMID- 10849734 TI - Development of a tertiary-structure model of the C-terminal domain of DPP IV. AB - Based on the recently published structure of prolyl oligopeptidase (POP) a model of the C-terminal part of dipeptidyl peptidase IV (DPP IV) which contains the active site has been developed. The structure of the model of DPP IV shows considerable similarity to the structure of POP particularly in the active site. A hydrophobic pocket (Tyr666, Tyr670, Tyr 631, Val556) forms the S1-binding site for recognition of proline. Tyr547 may stabilise the oxyanion formed in the tetrahedral intermediates by a strong hydrogen bond. The positively charged N terminus of ligands of DPP IV is recognised by forming a salt bridge with the acidic side chain Glu668. A second hydrophobic pocket (S2' to S5') may represent an important binding site for HIV-1 Tat-protein derivatives, chemokines and others. PMID- 10849735 TI - Post-proline-cleaving peptidases having DP IV like enzyme activity. Post-proline peptidases. AB - DP IV has been studied extensively in disease and in the immune system by the use of enzyme assays which detect hydrolysis of Gly-Pro or Ala-Pro substrate. In addition many studies have used inhibitors of DP IV enzyme activity. The characterisation of a novel DP IV like protein, DPP4R, and of other proteases which have a substrate specificity similar to DP IV or that bind DP IV inhibitors suggests that these studies require further evaluation. PMID- 10849736 TI - A new type of fluorogenic substrates for determination of cellular dipeptidyl peptidase IV (DP IV/CD26) activity. AB - The stability of cell associated fluorescence is an essential requirement for measurements of cellular enzymatic activity via enzyme catalyzed liberation of fluorophores. Rhodamine 110 (R110), a highly fluorescent xanthene dye, was used to synthesize nonfluorescent dipeptidyl peptidase IV (DP IV) substrates Xaa-Pro R110-Y allowing the stable covalent binding of the enzymatically released fluorescent R110-Y on cells. All compounds have been characterized as substrates of isolated DP IV with kcat/Km values of about 10(6) M-1.s-1. The hydrophobicity of the residue Y affects the affinity of the substrate to the catalytic site of DP IV. The compounds are characterized as sensitive substrates of cell surface associated DP IV of DP IV rich U-937 cells. The binding of the enzymatically released R110-Y on cells results in a stable cellular fluorescence. This way, the quantitative determination of cell surface associated DP IV activity is possible. PMID- 10849737 TI - Potent inhibitors of dipeptidyl peptidase IV and their mechanisms of inhibition. AB - Dipeptidyl peptidase IV (DP IV) is a proline specific serine protease which cleaves Xaa-Pro-dipeptides from the N-terminus of longer peptides. A series of product analogous amino acid amides containing different structure modifications like substitution of a ring atom, variation of the ring size and/or the introduction of a thioxo amide bond, phosphono amide bond or reduced amide bond were done to characterize these compounds as inhibitors of DP IV. These compounds are mostly classical reversible inhibitors of DP IV. In contrast amino acyl-2 cyanopyrrolidides inhibit DP IV according to a slow-binding mechanism with inhibition constants in the nanomolare range. On the other hand, diaryl dipeptide phosphonates inhibit irreversibly. In conclusion, this work shows, that the mechanism of inhibition of DP IV depends on the structure of the investigated compounds. PMID- 10849738 TI - N-terminal HIV-1 Tat nonapeptides as inhibitors of dipeptidyl peptidase IV. Conformational characterization. AB - Compared to the N-terminal nonapeptide of the HIV-1 Tat protein as inhibitor of activity of DP IV which is supposed to mediate the immunosuppressive effects of HIV-1 Tat, the Ile5 and Leu6 analogues showed strongly reduced inhibitory activity. Interestingly, replacement of Asp2 with Gly or Lys led to compounds with considerably enhanced inhibition. Therefore, we have applied 1H NMR spectroscopy and restrained molecular dynamics calculations to elucidate the molecular conformation of a series of Tat nonapeptides. Conformational backbone differences of these peptides as well as the nature and the arrangement of the side chains per se at significant positions preventing effective binding to DP IV might explain their different inhibitory activity on DP IV. PMID- 10849739 TI - Signal transduction events induced or affected by inhibition of the catalytic activity of dipeptidyl peptidase IV (DP IV, CD26). AB - DP IV (CD26) represents an accessory surface molecule playing an important role in the process of activation and proliferation of human lymphocytes. The molecular events mediated by this ectoenzyme are only partly established and the necessity of DP IV enzymatic activity for its signalling capacity has been discussed controversial. Focusing on the putative role of the catalytic domain of this peptidase, it could be shown that inhibition of the catalytic activity can provoke many cellular effects, including induction of tyrosine phosphorylations and p38 MAP kinase activation as well as suppression of DNA synthesis and reduced production of various cytokines. TGF-beta 1, the production and secretion of which is increased after DP IV inhibition, supposedly mediates the observed suppressive effects by maintaining p27kip expression levels which leads to a cell cycle arrest in G1. Moreover, anti-CD3-induced signalling pathways, including Ca2+ mobilisation, MEK1-, Erk1/2- and PKB-activation, can be strongly affected by DP IV inhibition. Thus, the enzymatic activity or at least the interaction of effectors with the catalytic domain of CD26 seems to be important for crucial functions of this cell surface antigen. PMID- 10849740 TI - Specific inhibitors of dipeptidyl peptidase IV suppress mRNA expression of DP IV/CD26 and cytokines. AB - The dipeptidyl peptidase IV is an activation marker on T, B and NK cells. Specific inhibitors of DP IV suppress DNA synthesis, as well as cytokine protein production. Here, we describe for the first time the quantitative changes of mRNA expression of IFN-gamma, IL-2, IL-12 and DP IV after inhibition of DP IV. Due to the stimulation of human peripheral blood mononuclear cells (PBMC) with pokeweed mitogen (PWM) both cytokine as well as DP IV mRNA expression is increased significantly. Treatment with DP IV inhibitor suppresses dose-dependently these changes. Importantly, mRNA expression of DP IV itself was inhibited. The presented data are fully compatible with our hypothesis that inhibition of DP IV leads to cell cycle arrest in late G1 due to enhanced TGF-beta 1 expression. PMID- 10849741 TI - Dipeptidyl peptidase IV in inflammatory CNS disease. AB - Current pathogenic concepts of inflammatory demyelinating disorders such as multiple sclerosis (MS) are based on the hypothesis that a T cell-mediated autoimmune response is involved in the disease process. One of the primary goals in the in the development of immunotherapies for autoimmune diseases has been to achieve inactivation of disease-inducing lymphocytes either by direct inhibition or suppression through regulatory cells and/or cytokines. The CD26 antigen is identical with the cell surface ectopeptidase dipeptidyl peptidase IV (DP IV, EC 3.4.14.5) which is involved in regulating T cell activation and growth. Activated T cells, including those specific for myelin antigens, express high levels of CD26/DP IV. In vitro, reversible DP IV inhibitors suppress T cell proliferation and pro-inflammatory cytokine production in response to myelin antigens. Further studies will evaluate the role of DP IV inhibition in T cell-mediated inflammatory disease of the central nervous system. PMID- 10849742 TI - Dipeptidyl peptidase IV (CD26): role in T cell activation and autoimmune disease. AB - The ectoenzyme dipeptidyl peptidase IV (DP IV; EC 3.4.14.5; CD26) has been shown to play a crucial role in T cell activation. In the present study, we show by flow cytometry and by enzymatic DP IV assay that myelin basic protein (MBP) specific, CD4+ T cell clones (TCC) derived from patients with multiple sclerosis (MS) express high levels of DP IV/CD26. The enzymatic activity of resting TCC was found to be three to fourfold higher than on resting peripheral blood T cells and close to that of T cells 48 hours after PHA stimulation. The DP IV inhibitors Lys[Z(NO2)]-thiazolidide and Lys[Z(NO2)]-pyrrolidide suppress in a dose-dependent manner DNA synthesis and IFN-gamma, IL-4, and TNF-alpha production of the antigen stimulated TCC. These data suggest that CD26 plays a role in regulating activation of autoreactive TCC. Further in vivo investigations will clarify, whether the inhibition of the enzymatic activity of DP IV could be a useful tool for therapeutic interventions in MS and/or other autoimmune diseases. PMID- 10849743 TI - Effects of nonapeptides derived from the N-terminal structure of human immunodeficiency virus-1 (HIV-1) Tat on suppression of CD26-dependent T cell growth. AB - The human immunodeficiency virus-1 (HIV-1) transactivator Tat occurs extracellularly and exerts immunosuppressive effects. Interestingly, Tat inhibits dipeptidyl peptidase IV (DP IV) activity of the T cell activation marker CD26. The short N-terminal nonapeptide Tat(1-9), MDPVDPNIE, also inhibits DP IV activity and suppresses DNA synthesis of tetanus toxoid-stimulated peripheral blood mononuclear cells (PBMC). Here, we present the influence of amino acid exchanges in the first three positions of Tat(1-9). For instance, the replacement of D2 of Tat(1-9) by G or K generated peptides, which inhibit DP IV-catalyzed IL 2(1-12) cleavage nearly threefold stronger. Similar effects were observed on the suppression of DNA synthesis of Tetanus toxoid-stimulated PBMC. This correlation suggests that Tat(1-9)-deduced peptides mediate antiproliferative effects at least in part via specific DP IV interactions and supports the hypothesis that CD26 plays a key role in the regulation of lymphocyte growth. PMID- 10849744 TI - DNA synthesis in cultured human keratinocytes and HaCaT keratinocytes is reduced by specific inhibition of dipeptidyl peptidase IV (CD26) enzymatic activity. AB - The ectopeptidase dipeptidyl peptidase IV (DP IV, CD26, EC 3.4.14.5) is present on most mammalian cells. Using specific inhibitors of DP IV, it has been shown that this enzyme is involved in the regulation of DNA synthesis and in production of various cytokines in lymphocytes. The aim of the present work was to investigate the expression of DP IV/CD26 on human keratinocytes and to answer the question, whether the proliferation (DNA synthesis) of human keratinocytes is influenced by inhibition of the enzymatic activity of DP IV. Using flow cytometry, RT-PCR, and specific enzymatic activity assays, expression of DP IV mRNA and CD26 antigen were shown on primary keratinocyte strains and on the HaCaT keratinocyte cell line. The synthetic DP IV inhibitors Lys[Z(NO2)]-thiazolidide and -pyrrolidide suppress the DNA-synthesis of these cells in a dose-dependent manner. These data demonstrate that CD26 is also involved in the regulation of DNA synthesis of keratinocytes and that the enzymatic activity is required for mediating these effects. PMID- 10849745 TI - Attractin: a cub-family protease involved in T cell-monocyte/macrophage interactions. AB - Attractin is a rapidly upregulated membrane-associated molecule on activated T cells. It is a member of the CUB family of extracellular guidance and development proteins, sharing with them a protease activity similar to that of Dipeptidyl peptidase IV (DPPIV/CD26). Most remarkably, and in sharp contrast to CD26, it is released from the T cell and is presumed to be a major source of a soluble serum circulating attractin. Genomic sequencing reveals that the soluble form is not a proteolytic product of the membrane form, but is in fact the result of alternative splicing. Recent results prove that the loss of murine membrane attractin results in the mahogany mutation with severe repercussions upon skin pigmentation and control of energy metabolism. In each of these latter instances, there is a strong likelihood that attractin is moderating the interaction of cytokines with their respective receptors. We propose that attractin is performing a similar function in the immune system through capture and proteolytic modification of the N-terminals of several cytokines and chemokines. This regulatory activity allows cells to interact and form immunoregulatory clusters and subsequently aids in downregulating chemokine/cytokine activity once a response has been initiated. These two properties are likely to be affected by the balance of membrane-expressed to soluble attractin. PMID- 10849746 TI - Analogs of glucose-dependent insulinotropic polypeptide with increased dipeptidyl peptidase IV resistance. PMID- 10849747 TI - Dipeptidyl peptidase IV (DPP IV, CD26) in patients with mental eating disorders. AB - The notion that patients with eating disorders maintain a functional immunosurveillance in spite of severe malnutrition has attracted researchers for years. Dipeptidyl Peptidase IV (DPP IV), a serine protease with broad tissue distribution and known activity in serum, operates in the cascade of immune responses. Membrane-bound DPP IV expressed on lymphocytes, also known as the leukocyte antigen CD26, is considered to participate in T cell activation. We hypothesized that the activity of DPP IV in serum and expression of CD26 in lymphocytes may be altered in patients with eating disorders. Serum DPP IV activity and the number of CD26 (DPP IV)-positive peripheral blood lymphocytes were measured in 44 patients (anorexia nervosa (AN): n = 21, bulimia (B): n = 23) in four consecutive weekly analyses. The analysis of CD26-positive cells included the characterization of CD26-bright and CD26-dim positive subsets. Additionally, the expression of CD25 (IL-2 Receptor alpha chain) was evaluated to estimate the degree of T cell activation. The same analyses were carried out in healthy female volunteers (HC, n = 20). CD26-positive cells were reduced in patients as compared to healthy controls (mean 40.2% (AN) and 41.1% (B) vs. 47.4% (HC), p < 0.01), while the DPP IV activity in serum was elevated (mean 108.4 U/l (AN) and 91.1 U/l (B) vs. 80.3 U/l (HC), p < 0.01). The potential implications of changes in DPP IV expression and serum activity on--and beyond--immune function are discussed. PMID- 10849748 TI - The membrane-bound ectopeptidase CPM as a marker of macrophage maturation in vitro and in vivo. AB - During terminal maturation of human blood monocytes into macrophages, a multitude of phenotypic and functional changes occurs: cells increase in size, they enhance their capacity for phagocytosis and tumor cytotoxicity but decrease their ability for T-lymphocyte stimulation. The pattern of secreted cytokines is shifted as is the profile of surface antigens. We recently identified carboxypeptidase M (CPM) as a macrophage maturation-associated antigen detected by mAb MAX. 1/MAX. 11. CPM, a phosphoinositol-linked ectopeptidase, is able to process a multitude of different substrates, among them immunologically important peptides like bradykinin, anaphylatoxins and enkephalins. It was previously shown to be expressed in placenta, lung, and kidney. CPM as detected by MAX. 1/11 shows a strong expression on monocyte-derived macrophages in vitro and on macrophages in vivo accompanying T-lymphocyte activation like during allogeneic transplant rejection or allergic alveolitis. In contrast, its expression is suppressed on macrophages by some types of tumor cells. CPM expression seems to correlate with macrophage cytotoxic functions. However, the biological importance of CPM expression in human macrophages in vivo is difficult to predict. A wide range of biologically active peptides are cleaved by CPM, and the relevance of CPM peptide processing during an immune reaction is only poorly understood. The generation and analysis of CPM-deficient animals might improve our understanding of CPM function. Therefore we cloned a cDNA for the murine homologue of CPM. However, expression of mCPM was undetectable in murine primary macrophages and macrophage cell-lines, suggesting that CPM expression and function is not conserved between human and mouse macrophages. PMID- 10849749 TI - Matrix metalloproteinases (MMP-8, -13, and -14) interact with the clotting system and degrade fibrinogen and factor XII (Hagemann factor). PMID- 10849750 TI - The neprilysin family in health and disease. AB - The mammalian neprilysin (NEP) family comprises at least seven members: NEP itself, Kell blood group antigen (KELL), the endothelin-converting enzymes (ECE-1 and ECE-2), the enzyme PEX, associated with X-linked hypophosphataemia, "X converting enzyme" (XCE) a CNS-expressed orphan peptidase and a soluble, secreted endopeptidase (SEP). These zinc metallopeptidases are all type II integral membrane proteins. Where identified, these enzymes have roles in the processing or metabolism of regulatory peptides and therefore represent potential therapeutic targets. A distinct feature of ECE-1 species is their existence as distinct isoforms differing in their N-terminal cytoplasmic tails. These tails play a role in enzyme targeting and turnover with di-leucine and tyrosine-based motifs affecting localization. Additional anchorage of these enzymes can also occur through palmitoylation. Bacterial homologues of the neprilysin family exist, for example the products of the pepO genes from L. lactis and S. parasanguis, and a recently described gene product of P. gingivalis which is an ECE-1 homologue that can catalyse the conversion of big endothelin to endothelin. A genomics based approach to understanding the functions of this proteinase family is aided by the completion of the C. elegans and Drosophila genomes, both of which encode multiple copies of NEP-like enzymes. PMID- 10849751 TI - Review: novel cysteine proteases of the papain family. PMID- 10849752 TI - Development and validation of homology models of human cathepsins K, S, H, and F. AB - Models of the tertiary structures of cathepsins K, S, H, and F were constructed by using homology protein modelling methods and refinements by interactive graphics and energy minimisation. The predicted structures yield information regarding their substrate binding sites and indicate the residues surrounding these sites. The ligand binding sites were characterised and compared with each other by means of calculated molecular electrostatic surface potentials. This will allow designing and development of new ligands specific for these cathepsins in future investigations. PMID- 10849753 TI - The function of propeptide domains of cysteine proteinases. AB - The papain-like cysteine proteinases can be divided into cathepsin L-like and cathepsin B-like enzymes because of the extended proregion of the former ones. We performed a series of mutations (alanine scan) in the prodomain of procathepsin S in order to elucidate the function of this extended domain in the L-like cathepsins. One of the most striking results was that the structural stability and the folding of procathepsin S were considerably dependent on an aromatic stack built by the residues Trp 28, Trp 31 and Trp 52. Replacement either of one or of all of these residues by alanine resulted in loss of transport, maturation and secretion of the mutated zymogen. Recombinant propeptides carrying the same mutations are not longer selective and powerful inhibitors of cathepsin S. Therefore we postulated an essential role of the propeptide for proper folding of the whole enzyme. This assumption was further proved by in vitro studies. We investigated the capability of recombinant cathepsin S propeptide to catalyze the renaturation of denatured mature cathepsin S. The experiments showed a 10-25 fold faster renaturation rate in presence than in absence of the propeptide underlining its function as intramolecular chaperone. PMID- 10849754 TI - Human cathepsins W and F form a new subgroup of cathepsins that is evolutionary separated from the cathepsin B- and L-like cysteine proteases. PMID- 10849755 TI - Cathepsin K expression in human lung. AB - Tissue remodeling is crucial in different lung diseases, in the embryonal development as well as in bronchial carcinoma. Cathepsins were proposed to be involved in the degradation of matrix proteins. Cathepsin K is one of the most potent matrix-degrading cysteine proteinases known as yet. The elastinolytic and collagenolytic activity of this papain-like protease is comparable with that of neutrophil elastase. We have investigated the cathepsin K expression in normal adult lung tissues, in embryonal lung tissue and in bronchial carcinoma. With help of specific anti-cathepsin K antibodies it could be shown that cathepsin K was expressed in bronchial epithelial cells. These data could be confirmed at mRNA level using a quantitative RT-PCR as well as by visualisation of the specific enzymatic activity in epithelial cell lines. During the embryonal development cathepsin K was expressed in the epithelial cells of the developing bronchi. The expression seemed to be upregulated in parallel with the development of the bronchial and alveolar lumen. In the later phase of lung development the cathepsin K expression was restricted to bronchial epithelial cells. Furthermore, using quantitative RT-PCR it could be shown that cathepsin K-mRNA was upregulated in lung tumor tissues in comparison to normal tissues from the same patients. These data suggest that cathepsin K may play an important role in matrix remodeling of the lung under physiological and pathological conditions. PMID- 10849756 TI - Expression of cathepsins B and L in human lung epithelial cells is regulated by cytokines. AB - The cathepsins B, L, and H are expressed ubiquitously and represent the major proportion of lysosomal enzymes. They are involved in bulk proteolysis in the lysosomes, processing of proteins and matrix degradation. Under pathological conditions the participation of cathepsins, especially their secreted forms, was observed in inflammation, tumor progression and metastasis. The enzymatic activity of cathepsins is regulated by posttranslational modification, localization, maturation, changes in pH, and their interaction with inhibitors. Regulation at the level of transcription is not well elucidated. The aim of this study was to investigate the effect of IL-1 beta, IL-6, IL-10, TGF-beta 1, and HGF on mRNA expression and protein level in human lung epithelial cell lines A 549 and BEAS-2B. IL-6 leads to a twofold increase in cathepsin L mRNA expression, whereas TGF-beta 1 decreases the amount of cathepsin L mRNA. At protein level, using enzyme immunoassay, it was shown that IL-6 induced increased amounts of cathepsin L but not cathepsin B. In contrast, after incubation of bronchial epithelial cells with TGF-beta 1 the cathepsin L concentration was decreased. In conclusion, gene expression of cathepsins B and L is variable. The cytokines IL-6 and TGF-beta 1 modulate cathepsin gene expression. PMID- 10849757 TI - Functions of cathepsin K in bone resorption. Lessons from cathepsin K deficient mice. AB - Cathepsin K is a cysteine proteinase expressed predominantly in osteoclasts. Cathepsin K cleaves key bone matrix proteins and is believed to play an important role in degrading the organic phase of bone during bone resorption. Pycnodysostosis, an autosomal recessive osteosclerosing skeletal disorder has recently been shown to result from mutations in the cathepsin K gene. Cathepsin K deficient mice generated by targeted disruption of this proteinase phenocopy many aspects of pycnodysostosis. They display an osteopetrotic phenotype with excessive trabeculation of the bone-marrow space accompanied by an altered ultrastructural appearance of the cathepsin K deficient osteoclasts. These cells also demonstrate an impaired resorptive activity in vitro. In contrast to other forms of osteopetrosis, which are due to disrupted osteoclastogenesis, cathepsin K deficiency is associated with an inhibition of osteoclast activity. Taken together the phenotype of cathepsin K knockout mice underlines the importance of this proteinase in bone remodelling. PMID- 10849758 TI - Ceramide as an activator lipid of cathepsin D. AB - We have identified the aspartic protease cathepsin D as a novel intracellular target protein for the lipid second messenger ceramide. Ceramide specifically binds to and induces CTSD proteolytic activity. A-SMase deficient cells derived from Niemann-Pick patients show decreased CTSD activity that was reconstituted by transfection with A-SMase cDNA. Ceramide accumulation in cells derived from A ceramidase defective Farber patients correlates with enhanced CTSD activity. These findings suggest that A-SMase-derived ceramide targets endolysosomal CTSD. PMID- 10849759 TI - Human cathepsin X. A novel cysteine protease with unique specificity. PMID- 10849760 TI - A novel proteolytic mechanism for termination of the CA2+ signalling evoked by proteinase-activated receptor-1 (PAR-1) in rat astrocytes. PMID- 10849761 TI - Natural and synthetic inhibitors of the tumor-associated serine protease urokinase-type plasminogen activator. PMID- 10849762 TI - Processing of interleukin-18 by human vascular smooth muscle cells. PMID- 10849763 TI - Review: peptidases and peptidase inhibitors in the pathogenesis of diseases. Disturbances in the ubiquitin-mediated proteolytic system. Protease-antiprotease imbalance in inflammatory reactions. Role of cathepsins in tumour progression. PMID- 10849764 TI - The role of proteolysis in Alzheimer's disease. AB - Alzheimer's disease is characterised by the progressive deposition of the 4 kDa beta-amyloid peptide (A beta) in extracellular senile plaques in the brain. A beta is derived by proteolytic cleavage of the amyloid precursor protein (APP) by various proteinases termed secretases. alpha-Secretase is inhibited by hydroxamate-based zinc metalloproteinase inhibitors such as batimastat with I50 values in the low micromolar range, and displays many properties in common with the secretase that releases angiotensin converting enzyme. A cell impermeant biotinylated derivative of one such inhibitor completely blocked the release of APP from the surface of neuronal cells, indicating that alpha-secretase cleaves APP at the cell-surface. A range of hydroxamate-based compounds have been used to distinguish between alpha-secretase and tumour necrosis factor-alpha convertase, a member of the ADAMs (a disintegrin and metalloproteinase-like) family of zinc metalloproteinases. Recent data suggests that the presenilins may be aspartyl proteinases with the specificity of gamma-secretase. Although APP and the presenilins are present in detergent-insoluble, cholesterol- and glycosphingolipid-rich lipid rafts, they do not behave as typical lipid raft proteins, and thus it is unclear whether these membrane domains are the sites for proteolytic processing of APP. PMID- 10849766 TI - The role of cysteine proteases in intracellular pancreatic serine protease activation. AB - Autodigestion by proteolytic enzymes is thought to represent the critical mechanism by which acute pancreatitis is initiated. Where and why pancreatic proteases, which are physiologically stored and secreted as inactive precursor zymogens, are activated within the pancreas has remained controversial. Here we present data which indicate that: the lysosomal protease cathepsin B can activate trypsinogen in vitro in a manner that is similar to trypsinogen activation by enterokinase; that cathepsin B colocalizes with trypsinogen in the secretory compartment of the rat pancreas and of the human pancreas; that trypsinogen activation begins in a secretory compartment that is distinct from mature zymogen granules; and that the inhibition of cathepsin B can either increase or decrease premature trypsinogen activation depending on the concentration of the inhibitor, its specificity and its site of action in the pancreatic acinar cell. These observations elucidate some of the complex relations between cysteine and serine proteases in the pancreas with respect to their mechanisms of activation, their subcellular sites of action, and their possible role in the onset of pancreatitis. PMID- 10849765 TI - Observing proteases in living cells. AB - The lysosomal cysteine protease cathepsin B has been implicated in tumor progression and metastasis in part due to its altered trafficking. In order to analyze the trafficking of cathepsin B in living cells, we utilized enhanced green fluorescent protein (EGFP) fused to various cathepsin B constructs for transfecting two cell lines: an invasive human breast adenocarcinoma cell line (BT20) and a cathepsin B deficient mouse embryonic fibroblast cell line (MEF T -/ ). The cells were transiently transfected with four cathepsin B-EGFP fusion constructs: full-length preprocathepsin B-EGFP, cathepsin B preregion-EGFP, cathepsin B prepro region-EGFP, and cathepsin B prepro region-EGFP with a mutation of the glycosylation site in the pro region. The full length construct showed vesicular distribution throughout the cells in both cell lines. In both BT20 and MEF T -/- cells, preregion-EGFP was localized in a ring tightly associated with the cell nucleus, suggesting distribution to the endoplasmic reticulum. The distribution of the prepro region-EGFP construct was similar except that it also included some patchy areas adjacent to the nucleus. This suggested that the cathepsin B prepro region-EGFP might have entered the Golgi. Distribution of the mutated cathepsin B prepro region-EGFP was similar to that of wild-type prepro region-EGFP in the MEF T -/-. In the invasive BT20 cells, however, the mutated prepro region-EGFP showed a vesicular distribution throughout the cytoplasm and in cell processes. This distribution is similar to that of endogenous cathepsin B in these cells. Our results suggest that: 1) tumor cells have an alternative mechanism for trafficking of cathepsin B which is independent of the mannose-6-phosphate receptor pathway, and 2) the pro region of cathepsin B may contain the sorting sequence necessary for its trafficking via this pathway. PMID- 10849767 TI - Peptidases in the asthmatic airways. PMID- 10849768 TI - Inactivation of interleukin-6 by neutrophil proteases at sites of inflammation. Protective effects of soluble IL-6 receptor chains. AB - In contrast to the excessively elevated immunochemically detectable concentrations of interleukin-6 (IL-6) in inflammatory exudates, the IL-6 bioactivities are significantly reduced, suggesting an inactivation of IL-6 at sites of inflammation. Since high amounts of proteases are released by invading neutrophils (PMN) in close temporal correlation to elevated IL-6 concentrations at sites of inflammation, this study focused on effects of the PMN-derived proteases elastase (NE), proteinase 3 (PR 3) and cathepsin G (Cat G) on the bioactivity and molecular integrity of IL-6. Here, we demonstrate that these enzymes play a crucial role in the initiation of the degradation and subsequent inactivation of IL-6 at sites of inflammation. Soluble IL-6 receptor subunits elicit a protective effect against the IL-6 inactivation by Cat G, only. Possible consequences of the proteolytical IL-6 inactivation for local inflammatory processes will be discussed. PMID- 10849769 TI - Antisense inhibition of cathepsin B in a human osteosarcoma cell line. PMID- 10849770 TI - Protease-protease inhibitor balance in the gastric mucosa. Influence of Helicobacter pylori infection. PMID- 10849771 TI - The role of bacterial and host proteinases in periodontal disease. AB - It is abundantly obvious that the uncontrolled degradation and/or activation of host defense pathways is the major pathway by which the periodontal pathogen P. gingivalis promotes its growth and proliferation. By being able to shed host receptors, degrade cytokines, and activate coagulation, complement, and kallikrein/kinin pathways it is clear that this organism has found a mechanism(s) to evade host defense and at the same time develop a system for cannibalizing host proteins for its own nutritional usage (Fig 2). Thus, it seems only logical that the development of inhibitors against these bacterial proteinases would be a useful method for negating their activities and making such pathogens more susceptible to attack by host phagocyte cells. In this respect, the structure of the truncated form of RGP has just been elucidated. Thus, it should only be a question of time before inhibitors to this enzyme will be developed and, hopefully, be used to reduce the pathologies associated with the development of periodontitis and/or eliminate the disease altogether. PMID- 10849772 TI - Multifunctional role of proteases in rheumatic diseases. PMID- 10849773 TI - Evidence of proteolytic activation of transforming growth factor beta in synovial fluid. PMID- 10849774 TI - Matrix metalloproteinases and TACE play a role in the pathogenesis of endometriosis. AB - Endometriosis, a benign gynecologic disorder, occurs in about 10% of women in reproductive age and in up to 50% of women with infertility. The basic etiologic factors causing this disease are unknown as yet. Matrix metalloproteinases (MMP) are involved in degradation of the extracellular matrix (ECM). Their proteolytic activity is regulated by tissue inhibitors of metalloproteinases (TIMPs). Tumor necrosis factor-alpha converting enzyme (TACE) is a membrane-bound disintegrin metalloproteinase that processes the membrane-associated cytokine proTNF-alpha to its mature soluble form. TNF-alpha induces the secretion of several MMPs. In order to study the expression of MMP-1, -2, -3 and -9, TIMP-1 and -2, TACE and TNF-alpha in endometrium and endometriotic tissue, we investigated formalin-fixed paraffin sections of endometriotic tissues and normal endometrium with immunohistochemical techniques and in situ hybridisation. Furthermore, quantitative PCR was used for quantification of TACE-mRNA in fresh tissue. We found in this study significant higher protein expression of MMP-1 and TACE and significant lower protein expression of TMP-1 and -2 in endometriotic tissue compared to endometrium. This data may suggest that high TACE expression causes the increased conversion of membrane-bound proTNF-alpha into its soluble form, which stimulates the increased secretion of MMP-1. The simultaneous deficiency of TIMP-1 and -2 in endometriotic tissue suppose an additional proteinase inhibitor imbalance in endometriosis. PMID- 10849775 TI - Influence of proliferation, differentiation and dedifferentiation factors on the expression of the lysosomal cysteine proteinase cathepsin L (CL) in thyroid cancer cell lines. PMID- 10849776 TI - Fundamental factors affecting biomass enzymatic reactivity. AB - Poplar wood was treated with peracetic acid, KOH, and ball milling to produce 147 model lignocelluloses with a broad spectrum of lignin contents, acetyl contents, and crystallinity indices (CrIs), respectively. An empirical model was identified that describes the roles of these three properties in enzymatic hydrolysis. Lignin content and CrI have the greatest impact on biomass digestibility, whereas acetyl content has a minor impact. The digestibility of several lime-treated biomass samples agreed with the empirical model. Lime treatment removes all acetyl groups and a moderate amount of lignin and increases CrI slightly; lignin removal is the dominant benefit from lime treatment. PMID- 10849777 TI - Fourier transform infrared quantification of sugars in pretreated biomass liquors. AB - The process of converting renewable lignocellulosic biomass to ethanol requires a number of steps, and pretreatment is one of the most important. Pretreatment usually involves a hydrolysis of the easily hydrolyzed hemi-cellulosic component of biomass using some form of thermal/chemical/mechanical action that results in a product that can be further hydrolyzed by cellulase enzymes (the cellulosic portion). The sugars produced can then be fermented to ethanol by fermentative microorganisms. If the pretreatment step is not severe enough, the resultant residue is not as easily hydrolyzed by the cellulase enzyme. More severe pretreatment conditions result in the production of degradation products that are toxic to the fermentative microorganism. In this article, we report the quantitative analysis of glucose, mannose, xylose, and acetic acid using Fourier transform infrared (FTIR) spectroscopy on liquors from dilute-acid-pretreated soft-wood and hard-wood slurries. Comparison of FTIR and high-performance liquid chromatography quantitative analyses of these liquors are reported. Recent developments in infrared probe technology has enabled the rapid quantification of these sugars by FTIR spectroscopy in the batch reactor during optimization of the pretreatment conditions, or interfaced to the computer controlling a continuous reactor for on-line monitoring and control. PMID- 10849778 TI - Floating cultivation of marine cyanobacteria using coal fly ash. AB - The aim of this study was to develop improved methodologies for bulk culturing of biotechnologically useful marine cyanobacteria in the open ocean. We have investigated the viability of using coal fly ash (CFA) blocks as the support medium in a novel floating culture system for marine micro-algae. The marine cyanobacterium Synechococcus sp. NKBG 040607 was found to adhere to floating CFA blocks in liquid culture medium. Maximum density of attached cells of 2.0 x 10(8) cells/cm2 was achieved using seawater. The marine cyanobacterium Synechococcus sp. NKBG 042902 weakly adhered to floating CFA blocks in BG-11 medium. Increasing the concentration of calcium ion in the culture medium enhanced adherence to CFA blocks. PMID- 10849779 TI - A kinetic study of lipase-catalyzed alcoholysis of palm kernel oil. AB - The use of lipases as biocatalysts in interesterification reactions has been the object of growing interest, owing to the importance of esters as emulsifiers, intermediates to produce oleochemicals, and fuel alternatives. We consider in this article a kinetic study of lipase-catalyzed alcoholysis of palm kernel oil, using n-hexane as the solvent. In a first step the ester production was maximized by using a Taguchi design, and then an empirical model was built to determine the influence of the process variables. Taking into account the results obtained in the first step, we performed a kinetic study and developed a simple model for this system. PMID- 10849780 TI - Simultaneous saccharification and fermentation of steam-pretreated spruce to ethanol. AB - Ethanol production was studied in simultaneous saccharification and fermentation (SSF) of steam-pretreated spruce at 42 degrees C, using a thermotolerant yeast. Three yeast strains of Kluyveromyces marxianus were compared in test fermentations. SSF experiments were performed with the best of these on 5% (w/w) of substrate, at a cellulase loading of 37 filter paper units/g of cellulose, and a beta-glucosidase loading of 38 IU/g of cellulose. The detoxification of the substrate and the lack of pH control in the experiments increased the final ethanol concentration. The final ethanol yield was 15% lower compared to SSF with Saccharomyces cerevisiae at 37 degrees C, owing to the cessation of ethanol fermentation after the first 10 h. PMID- 10849781 TI - Cellulose and hemicellulose hydrolysis models for application to current and novel pretreatment processes. AB - Acids catalyze the hydrolysis of cellulose and hemicellulose to produce sugars that organisms can ferment to ethanol and other products. However, advanced low- and no-acid technologies are critical if we are to reduce bio-ethanol costs to be competitive as a pure fuel. We believe carbohydrate oligomers play a key role in explaining the performance of such hydrolysis processes and that kinetic models would help us understand their role. Various investigations have developed reaction rate expressions based on an Arrhenius temperature dependence that is first order in substrate concentration and close to first order in acid concentration. In this article, we evaluate these existing hydrolysis models with the goal of providing a foundation for a unified model that can predict performance of both current and novel pretreatment process configurations. PMID- 10849782 TI - Effect of chip size on steam explosion pretreatment of softwood. AB - Although considerable progress has been made in technology for converting lignocellulosic biomass into ethanol, substantial opportunities still exist to reduce production costs. In biomass pretreatment, reducing milling power is a technological improvement that will substantially lower production costs for ethanol. Improving sugar yield from hemicellulose hydrolysis would also reduce ethanol production costs. Thus, it would be desirable to test innovative pretreatment conditions to improve the economics by reducing electrical power of the milling stage and by optimizing pretreatment recovery of hemicellulose, as well as to enhance cellulose hydrolysis. The objective of this study was to evaluate the effect of chip size (2-5, 5-8, and 8-12 mm) on steam-explosion pretreatment (190 and 210 degrees C, 4 and 8 min) of softwood (Pinus pinaster). PMID- 10849783 TI - Simultaneous saccharification and cofermentation of peracetic acid-pretreated biomass. AB - Previous work in our laboratories has demonstrated the effectiveness of peracetic acid for improving enzymatic digestibility of lignocellulosic materials. The use of dilute alkali solutions as a pre-pretreatment prior to peracetic acid lignin oxidation increased carbohydrate hydrolysis yields in a synergistic as opposed to additive manner. Deacetylation of xylan is easily achieved using dilute alkali solutions under mild conditions. In this article, we evaluate the effectiveness of peracetic acid combined with an alkaline pre-pretreatment through simultaneous saccharification and cofermentation (SSCF) of pretreated hybrid poplar wood and sugar cane bagasse. Respective ethanol yields of 92.8 and 91.9% of theoretical are achieved using 6% NaOH/15% peracetic acid-pretreated substrates and recombinant Zymomonas mobilis CP4/pZB5. Reduction of acetyl groups of the lignocellulosic materials is demonstrated following alkaline pre-pretreatments. Such processing may be helpful in reducing peracetic acid requirements. The influence of deacetylation is more significant in combined pretreatments using lower peracetic acid loadings. PMID- 10849784 TI - Pretreatment of wastepaper and pulp mill sludge by aqueous ammonia and hydrogen peroxide. AB - Pretreatment of two different softwood-based lignocellulosic wastes (newsprint and Kraft pulp mill sludge) was investigated. Pretreatment was done by aqueous ammonia and hydrogen peroxide (H2O2), two delignifying reagents that are environmentally benign. Three different treatment schemes were employed: aqueous ammonia alone (ammonia recycled percolation [ARP]), mixed stream of aqueous ammonia and H2O2, and successive treatment with H2O2 and aqueous ammonia. In all cases there was a substantial degree of delignification ranging from 30 to 50%. About half of the hemicellulose sugars were dissolved into the process effluent. Retention of cellulose after pretreatment varied from 85 to 100% for newspaper feedstock and from 77 to 85% for the pulp mill sludge. After treatment with aqueous ammonia alone (ARP), the digestibility of newspaper and the pulp mill sludge was improved only by 5% (from 40 to 45% for the former and from 68 to 73% for the latter), despite a substantial degree of delignification occurring after the ARP process. The lignin content thus did not correlate with the digestibility for these substrates. Simultaneous treatment with H2O2 and aqueous ammonia did not bring about any significant improvement in the digestibility over that of the ARP. A successive treatment by H2O2 and ARP showed the most promise because it improved the digestibility of the newspaper from 41 to 75%, a level comparable to that of alpha-cellulose. PMID- 10849785 TI - Fermentations of pectin-rich biomass with recombinant bacteria to produce fuel ethanol. AB - Pectin-rich residues from sugar beet processing contain significant carbohydrates and insignificant amounts of lignin. Beet pulp was evaluated for conversion to ethanol using recombinant bacteria as biocatalysts. Hydrolysis of pectin-rich residues followed by ethanolic fermentations by yeasts has not been productive because galacturonic acid and arabinose are not fermentable to ethanol by these organisms. The three recombinant bacteria evaluated in this study, Escherichia coli strain KO11, Klebsiella oxytoca strain P2, and Erwinia chrysanthemi EC 16 pLOI 555, ferment carbohydrates in beet pulp with varying efficiencies. E. coli KO11 is able to convert pure galacturonic acid to ethanol with minimal acetate production. Using an enzyme loading of 10.5 filter paper units of cellulase, 120.4 polygalacturonase units of pectinase, and 6.4 g of cellobiase (per gram of dry wt sugar beet pulp), with substrate addition after 24 h of fermentation, 40 g of ethanol/L was produced. Other recombinants exhibited lower ethanol yields with increases in acetate and succinate production. PMID- 10849786 TI - Oxidative cracking of precipitated hardwood lignin by hydrogen peroxide. AB - Precipitated hard-wood lignin (PHL) is a major byproduct in the biomass-to ethanol process. Oxidative cracking of PHL by hydrogen peroxide in aqueous medium was investigated as a means to produce potentially useful chemicals. The cracking reaction takes place at moderate temperatures (80-160 degrees C), giving mono- and dicarboxylic acids as the main products. The yields of these products are in the range of 30-50% of initial lignin. The reaction mechanism and the product distribution are dependent upon the reaction conditions, especially the pH. The reaction under strong alkaline condition proceeds well even at low reaction temperatures (80-90 degrees C). Under acidic conditions, higher temperatures (130 160 degrees C) are required to attain the same degrees of cracking. The reaction patterns of the oxidative cracking reaction involve the cleavage of lignin ring, aryl ether bond, or other linkages within lignin. By using the findings of this investigation and those of previous work, we have illustrated the reaction pathways for degradation of PHL under alkaline and acidic conditions. Aldehydes and aromatic acids are intermediate products in the oxidative degradation of lignin. However, they were produced only in trace amounts owing to rapid degradation induced by hydrogen peroxide. PMID- 10849787 TI - Optimizing ammonia pressurization/depressurization processing conditions to enhance enzymatic susceptibility of dwarf elephant grass. AB - An ammonia pressurization/depressurization process was investigated to evaluate the potential of producing reducing sugars from dwarf elephant grass, a warm season forage. Moisture, temperature, and ammonia loading affected sugar yield (p < 0.0001). At optimal conditions, ammonia processing solubilized 50.9% of the hemicellulose and raised the sugar yield (percentage of theoretical) from 18 to 83%. Glucose and xylose production were increased 3.2- and 8.2-fold, respectively. The mild processing conditions of the ammonia treatment (90-100 degrees C, 5 min), the low enzyme loading (2 international filter paper units/g), and the short hydrolysis time (24 h), greatly enhance the potential of using forages to produce sugars valuable for several applications. PMID- 10849788 TI - Development of new ethanologenic Escherichia coli strains for fermentation of lignocellulosic biomass. AB - Two new ethanologenic strains (FBR4 and FBR5) of Escherichia coli were constructed and used to ferment corn fiber hydrolysate. The strains carry the plasmid pLOI297, which contains the genes from Zymomonas mobilis necessary for efficiently converting pyruvate into ethanol. Both strains selectively maintained the plasmid when grown anaerobically. Each culture was serially transferred 10 times in anaerobic culture with sugar-limited medium containing xylose, but no selective antibiotic. An average of 93 and 95% of the FBR4 and FBR5 cells, respectively, maintained pLOI297 in anaerobic culture. The fermentation performances of the repeatedly transferred cultures were compared with those of cultures freshly revived from stock in pH-controlled batch fermentations with 10% (w/v) xylose. Fermentation results were similar for all the cultures. Fermentations were completed within 60 h and ethanol yields were 86-92% of theoretical. Maximal ethanol concentrations were 3.9-4.2% (w/v). The strains were also tested for their ability to ferment corn fiber hydrolysate, which contained 8.5% (w/v) total sugars (2.0% arabinose, 2.8% glucose, and 3.7% xylose). E. coli FBR5 produced more ethanol than FBR4 from the corn fiber hydrolysate. E. coli FBR5 fermented all but 0.4% (w/v) of the available sugar, whereas strain FBR4 left 1.6% unconsumed. The fermentation with FBR5 was completed within 55 h and yielded 0.46 g of ethanol/g of available sugar, 90% of the maximum obtainable. PMID- 10849789 TI - Characterization and complementation of a Pichia stipitis mutant unable to grow on D-xylose or L-arabinose. AB - Pichia stipitis CBS 6054 will grow on D-xylose, D-arabinose, and L-arabinose. D Xylose and L-arabinose are abundant in seed hulls of maize, and their utilization is important in processing grain residues. To elucidate the degradation pathway for L-arabinose, we obtained a mutant, FPL-MY30, that was unable to grow on D xylose and L-arabinose but that could grow on D-arabinitol. Activity assays of oxidoreductase and pentulokinase enzymes involved in D-xylose, D-arabinose, and L arabinose pathways indicated that FPL-MY30 is deficient in D-xylitol dehydrogenase (D-XDH), D- and L-arabinitol dehydrogenases, and D-ribitol dehydrogenase. Transforming FPL-MY30 with a gene for xylitol dehydrogenase (PsXYL2), which was cloned from CBS 6054 (GenBank AF127801), restored the D-XDH activity and the capacity for FPL-MY30 to grow on L-arabinose. This suggested that FPL-MY30 is critically deficient in XYL2 and that the D-xylose and L arabinose metabolic pathways have xylitol as a common intermediate. The capacity for FPL-MY30 to grow on D-arabinitol could proceed through D-ribulose. PMID- 10849790 TI - Investigation of the cell-wall loosening protein expansin as a possible additive in the enzymatic saccharification of lignocellulosic biomass. PMID- 10849791 TI - Butanol production using Clostridium beijerinckii BA101 hyper-butanol producing mutant strain and recovery by pervaporation. AB - Clostridium beijerinckii BA101 (mutant strain) and C. beijerinckii 8052 (wild type) were compared for substrate and butanol inhibition. The wild-type strain is more strongly inhibited by added butanol than is the mutant strain. Acetone and butanol were removed from a fed-batch reactor inoculated with C. beijerinckii BA101 by pervaporation using a silicone membrane. In the batch reactor, C. beijerinckii BA101 produced 25.3 g/L of total solvents, whereas in the fermentation-recovery experiment it produced 165.1 g/L of total solvents. Solvent productivity increased from 0.35 (batch reactor) to 0.98 g/L.h (fed-batch reactor). The fed-batch reactor was fed with 500 g/L of glucose-based P2 medium. Acetone selectivities ranged from 2 to 10 whereas butanol selectivities ranged from 7 to 19. Total flux varied from 26 to 31 g/m2.h. PMID- 10849792 TI - Enzyme production of Trichoderma reesei Rut C-30 on various lignocellulosic substrates. AB - Economical production of cellulase enzyme is key for feasible bio-ethanol production from lignocellulosics using an enzyme-based process. On-site cellulase production can be more feasible with the process of separate hydrolysis and fermentation (SHF) than with simultaneous saccharification and fermentation, since the cost of enzyme is more important and a variety of substrates are available for the SHF process. Cellulase production using various biomass substrates available for SHF, including paper sludge, pretreated wood (steam exploded), and their hydrolysis residues, was investigated in shake flasks and a fermenter for their productivities and titers. Among the newspaper sludge, office paper sludge, and steam-exploded woods treated in various ways, the steam exploded wood showed the best properties for substrate in cellulase production. The best titer of 4.29 IU/mL was obtained using exploded wood of 2% (w/v) slurry in the shake flask, and the titer with the same substrate was duplicated to about 4.30 IU/mL in a 3.7-L fermenter. Also, the yield of enzyme reached 215 IU/g of substrate or 363 IU/g of cellulose. Despite various pretreatment attempts, newspaper and office paper substrate was inferior to the exploded-wood substrate for cellulase production. However, hydrolysis residues of papers showed quite promising results. The hydrolysis residue of office paper produced 2.48 IU/mL of cellulase in 7 d. Hence, the utilization of hydrolysis residues for cellulase production will be further investigated in the future. PMID- 10849793 TI - Nitrogen regulation of Saccharomyces cerevisiae invertase. Role of the URE2 gene. AB - The regulation of extracellular enzymes is of great biotechnological interest. We studied the regulatory role of the URE2 gene on the periplasmic invertase of Saccharomyces cerevisiae, because its periplasmic asparaginase is regulated by the URE2/GLN3 system. Enzymatic activity was measured in the isogenic strains P40 1B, the ure2 mutant P40-3C, and the P40-3C strain transformed with the pIC-CS plasmid carrying the URE2 gene. The assays were performed using midlog and stationary phase cells and nitrogen-starved cells from these growth phases. During exponential growth, the level of invertase in both wild-type and ure2 mutant cells was comparable. However, the invertase activity in ure2 mutant cells from stationary phase was sixfold lower than in the wild-type cells. When P40-3C cells were transformed with the pIC-CS plasmid, the wild-type phenotype was restored. On nitrogen starvation in the presence of sucrose, the invertase activity in wild-type cells from midlog phase decreased three times, whereas in stationary cells, the activity decreased eight times. However, invertase activity doubled in ure2 mutant cells from both phases. When these cells were transformed with the aforementioned plasmid, the wild-type phenotype was restored, although a significant invertase decrease in stationary cell was not observed. These results suggested that the URE2 protein plays a role in invertase activity. PMID- 10849794 TI - N-demethylation of methylene blue by lignin peroxidase from Phanerochaete chrysosporium. Stoichiometric relation for H2O2 consumption. AB - Phanerochaete chrysosporium lignin peroxidase (LiP) can degrade synthetic dyes such as heterocyclic, azo, and triphenylmethane on its activation by H2O2. Analysis of the reaction products indicated that N-demethylation reactions are involved in the degradation of crystal violet and methylene blue (MB). We studied LiP oxidation of methylene blue and azure B (AB) in reaction mixtures containing different dye:H2O2 stoichiometric relations aiming at the selective formation of N-demethylated derivatives. High yields, about 70%, of the mono- and didemethylated derivatives, azure B and azure A, were obtained with the use of 1:1 and 1:2 MB:H2O2, respectively. Using azure B as substrate in reaction mixtures containing 1:1 AB:H2O2, a yield of 70% was also observed in azure A. Reaction mixtures containing 1:3 MB:H2O2 and 1:2 AB:H2O2, originated several oxidation products in similar proportions. These results indicated that the process of enzymatic degradation of methylene blue and azure B initiates via N(CH3)2 oxidation. According to the yields that were obtained for azure B and azure A, this enzymatic route can be used for the synthesis of these dyes since these data compare favorably to the chemical route that has a yield of 35%. The use of a dye:H2O2 relation of 1:10 resulted in a decoloration level of about 85%, showing the usefulness of this procedure for wastewater treatment. The reaction products were followed by spectrophotometric analysis within the wavelength of 500-700 nm. The product identifications were performed using a reverse-phase high performance liquid chromatography (HPLC) C-18 column and thin-layer chromatography. PMID- 10849795 TI - Endocellulase and exocellulase activities of two Streptomyces strains isolated from a forest soil. AB - Two Streptomyces strains, M7a and M23, from a Brazilian forest soil were evaluated for the cellulase production of their supernatants after growth in a microcrystalline cellulose medium, using carboxymethylcellulose and filter paper as substrates at different temperatures and pH values. Endoglucanase and exoglucanase activities were compared to a commercial Trichoderma reesei cellulase using fluorogenic conjugated substrates. Similar specific activities were observed for the enzyme preparations of strain M23 and T. reesei. For M7a the activities were about seven times higher than those obtained for T. reesei. Extracellular or cell-associated cellobiase activities were not detected in both strains. PMID- 10849796 TI - Comparative energetics of glucose and xylose metabolism in recombinant Zymomonas mobilis. AB - Recombinant Zymomonas mobilis CP4:pZB5 was grown with pH control in batch and continuous modes with either glucose or xylose as the sole carbon and energy source. In batch cultures in which the ratio of the final cell mass concentration to the amount of sugar in the medium was constant (i.e., under conditions that promote "coupled growth"), maximum specific rates of glucose and xylose consumption were 8.5 and 2.1 g/(g of cell.h), respectively; maximum specific rates of ethanol production for glucose and xylose were 4.1 and 1.0 g/(g of cell.h), respectively; and average growth yields from glucose and xylose were 0.055 and 0.034 g of dry cell mass (DCM)/g of sugar, respectively. The corresponding value of YATP for glucose and xylose was 9.9 and 5.1 g of DCM/mol of ATP, respectively. YATP for the wild-type culture CP4 with glucose was 10.4 g of DCM/mol of ATP. For single substrate chemostat cultures in which the growth rate was varied as the dilution rate (D), the maximum or "true" growth yield (max Yx/s) was calculated from Pirt plots as the inverse of the slope of the best-fit linear regression for the specific sugar utilization rate as a function of D, and the "maintenance coefficient" (m) was determined as the y-axis intercept. For xylose, values of max Yx/s and m were 0.0417 g of DCM/g of xylose (YATP = 6.25) and 0.04 g of xylose/(g of cell.h), respectively. However, with glucose there was an observed deviation from linearity, and the data in the Pirt plot was best fit with a second-order polynomial in D. At D > 0.1/h, YATP = 8.71 and m = 2.05 g of glu/(g of cell.h) whereas at D < 0.1/h, YATP = 4.9 g of DCM/mol of ATP and m = 0.04 g of glu/(g of cell.h). This observation provides evidence to question the validity of the unstructured growth model and the assumption that Pirt's maintenance coefficient is a constant that is independent of the growth rate. Collectively, these observations with individual sugars and the values assigned to various growth and fermentation parameters will be useful in the development of models to predict the behavior of rec Zm in mixed substrate fermentations of the type associated with biomass-to-ethanol processes. PMID- 10849797 TI - Continuous fermentation studies with xylos-utilizing recombinant Zymomonas mobilis. AB - This study examined the continuous cofermentation performance characteristics of a dilute-acid "prehydrolysate-adapted" recombinant Zymomonas 39676:pZB4L and builds on the pH-stat batch fermentations with this recombinant that we reported on last year. Substitution of yeast extract by 1% (w/v) corn steep liquor (CSL) (50% solids) and Mg (2 mM) did not alter the cofermentation performance. Using declared assumptions, the cost of using CSL and Mg was estimated to be 12.5 cents/gal of ethanol with a possibility of 50% cost reduction using fourfold less CSL with 0.1% diammonium phosphate. Because of competition for a common sugar transporter that exhibits a higher affinity for glucose, utilization of glucose was complete whereas xylose was always present in the chemostat effluent. The ethanol yield, based on sugar used, was 94% of theoretical maximum. Altering the sugar ratio of the synthetic dilute acid hardwood prehydrolysate did not appear to significantly change the pattern of xylose utilization. Using a criterion of 80% sugar utilization for determining the maximum dilution rate (Dmax), changing the composition of the feed from 4% xylose to 3%, and simultaneously increasing the glucose from 0.8 to 1.8% shifted Dmax from 0.07 to 0.08/h. With equal amounts of both sugars (2.5%), Dmax was 0.07/h. By comparison to a similar investigation with rec Zm CP4:pZB5 with a 4% equal mixture of xylose and glucose, we observed that at pH 5.0, the Dmax was 0.064/h and shifted to 0.084/h at pH 5.75. At a level of 0.4% (w/v) acetic acid in the CSL-based medium with 3% xylose and 1.8% glucose at pH 5.75, the Dmax for the adapted recombinant shifted from 0.08 to 0.048/h, and the corresponding maximum volumetric ethanol productivity decreased 45%, from 1.52 to 0.84 g/(L.h). Under these conditions of continuous culture, linear regression of a Pirt plot of the specific rate of sugar utilization vs D showed that 4 g/L of acetic acid did not affect the maximum growth yield (0.030 g dry cell mass/g sugar), but did increase the maintenance coefficient twofold, from 0.46 to 1.0 g of sugar/(g of cell.h). PMID- 10849798 TI - Improvement of enantioselectivity of chiral organophosphate insecticide hydrolysis by bacterial phosphotriesterase. AB - The bacterial phosphotriesterase (PTE) isolated from Flavobacterium sp. can catalyze the cleavage of the P-O bond in a variety of organophosphate triesters and has been shown to be an effective catalyst for the degradation of toxic organophosphate esters. Ethyl 4-nitrophenyl phenylphosphonothioate (EPN) is a chiral organophosphate. Optical isomers of EPN show differences in their toxicity. R-EPN is known to be more toxic to hens and houseflies than S-EPN. We determined the Ki value of each enantiomer toward electric eel acetylcholinesterase. R-EPN (Ki = 6 microM) inhibited acetylcholinesterase much more effectively than S-EPN (Ki = 52 microM) did in vitro. Since PTE has been found to hydrolyze only the S-isomer of EPN, we attempted to alter the enantioselectivity of PTE in order to degrade toxic EPN enantiomer effectively. When PTE hydrolyzed EPN in the presence of dimethyl sulfoxide (DMSO), enzymatic activity toward S-EPN decreased linearly, but enzymatic activity toward R-EPN increased as a function of DMSO concentration. At 20% DMSO, the maximum activity was observed. The kinetic parameters of PTE to EPN isomers clearly indicated that in the presence of 20% DMSO, the enantioselectivity of PTE changed. The Km value for R-EPN decreased from 0.24 to 0.03 mM, and the Vmax value increased from 0.25 to 0.60 U/mg of protein. Vmax/Km values indicated that PTE preferred R-EPN over S EPN in the presence of DMSO by a factor of 2. PMID- 10849799 TI - Enhanced alkaline protease production in addition to alpha-amylase via constructing a Bacillus subtilis strain. AB - Bacillus subtilis Bios11 strain was previously isolated and identified. This strain naturally produces a high level of alpha-amylase. The multicopy (pS1) plasmid that carries the complete alkaline protease aprA gene was introduced to this host strain by transformation. The newly constructed strain was found to express the aprA gene and produces a high level of alkaline protease. The level of alpha-amylase production was not affected compared with the parent strain. The pS1 plasmid in the new host was proved to be segregationally and structurally stable, and the multicopy aprA gene was expressed at the stationary phase. This expression did not affect growth rate and sporulation frequency. Moreover, the level of alpha-amylase was maintained. Both alkaline protease and alpha-amylase enzymes were purified using a single-step affinity chromatography column. The use of the newly constructed strain would be valuable to the enzyme industry and would promote recycling of some food-processing wastes. PMID- 10849800 TI - Methods to enhance tolerances of Chlorella KR-1 to toxic compounds in flue gas. AB - Possible methods to minimize the toxic effects of SOx and NOx on the growth of a highly CO2 tolerant and fast-growing microalga, Chlorella sp. KR-1, were investigated. Maintaining the pH of the culturing media at an adequate value was quite important to enhancing the tolerances of the microalgae to SOx and NOx. Controlling the pH by adding an alkaline solution, using a low flow rate of gas fed to the culture, and using a high concentration of inoculating cells were effective methods to maintaining the proper pH of the culture. Controlling the pH was the most effective method but could be applied only for some specific microalgae. PMID- 10849801 TI - Characterization of a high-productivity recombinant strain of Zymomonas mobilis for ethanol production from glucose/xylose mixtures. AB - The fermentation characteristics of a recombinant strain of Zymomonas mobilis ZM4(pZB5) capable of converting both glucose and xylose to ethanol have been further investigated. Previous studies have shown that the strain ZM4(pZB5) was capable of converting a mixture of 65 g/L of glucose and 65 g/L of xylose to 62 g/L of ethanol in 48 h with an overall yield of 0.46 g/g. Higher sugar concentrations (e.g., 75/75 g/L) resulted in incomplete xylose utilization (80 h). In the present study, further kinetic evaluations at high sugar levels are reported. Acetate inhibition studies and evaluation of temperature and pH effects indicated increased maximum specific uptake rates of glucose and xylose under stressed conditions with increased metabolic uncoupling. A high-productivity system was developed that involved a membrane bioreactor with cell recycling. At sugar concentrations of approx 50/50 g/L of glucose/xylose, an ethanol concentration of 50 g/L, an ethanol productivity of approx 5 g/(L.h), and a yield (YP/S) of 0.50 g/g were achieved. Decreases in cell viability were found in this system after attainment of an initial steady state (40-60 h); a slow bleed of concentrated cells may be required to overcome this problem. PMID- 10849802 TI - Nuclear magnetic resonance studies of acetic acid inhibition of rec Zymomonas mobilis ZM4(pZB5). AB - The fermentation characteristics and effects of lignocellulosic toxic compounds on recombinant Zymomonas mobilis ZM4(pZB5), which is capable of converting both glucose and xylose to ethanol, and its parental strain, ZM4, were characterized using 13C and 31P nuclear magnetic resonance (NMR) in vivo. From the 31P NMR data, the levels of nucleoside triphosphates (NTP) of ZM(pZB5) using xylose were lower than those of glucose. This can be related to the intrinsically slower assimilation and/or metabolism of xylose compared to glucose and is evidence of a less energized state of ZM4(pZB5) cells during xylose fermentation. Acetic acid was shown to be strongly inhibitory to ZM4(pZB5) on xylose medium, with xylose utilization being completely inhibited at pH 5.0 or lower in the presence of 10.9 g/L of sodium acetate. From the 31P NMR results, the addition of sodium acetate caused decreased NTP and sugar phosphates, together with acidification of the cytoplasm. Intracellular deenergization and acidification appear to be the major mechanisms by which acetic acid exerts its toxic effects on this recombinant strain. PMID- 10849803 TI - Effects of environmental conditions on xylose reductase and xylitol dehydrogenase production by Candida guilliermondii. AB - The effects of environmental conditions, namely initial pH (2.5-7.0) and temperature (25 and 35 degrees C), on xylose reductase and xylitol dehydrogenase levels, as well as on xylitol production, were evaluated. Although the fermentative parameter values increased with an increase in pH and temperature (the maximum Yp/s and Qp were 0.75 g/g and 0.95 g/[L.h], respectively, both attained at pH 6.0, 35 degrees C), the highest xylose reductase activities (nearly 900 IU/mg of protein) were observed at an initial pH varying from 4.0 to 6.0. Xylitol dehydrogenase was favored by an increase in both initial pH and temperature of the medium. The highest xylitol dehydrogenase specific activity was attained at pH 6.5 and 35 degrees C (577 IU/mg of protein). PMID- 10849804 TI - Construction of recombinant Escherichia coli strains for polyhydroxybutyrate production using soy waste as nutrient. AB - Construction and comparison of recombinant Escherichia coli strains harboring the polyhydroxybutyrate (PHB) operon from Ralstonia eutropha using vectors possessing different promotors, as well as the production of PHB from soy waste by the recombinant strain, are reported. The lac promotor was the most efficient on expression of the phb operon among the three promotors studied: i.e., lac promotor, T7 promotor and the normal sigma 70 promotor. The pKS/PHB was the most efficient plasmid for phb operon expression among the three plasmids used: i.e., pKS-, pAED4, and pJM9131. It was observed that isopropyl-beta-D thiogalactopyranoside was not required for the induction of the expression of phb operon. The cell dry wt and polyhydroxyalkanoate content by E. coli XL-1 Blue (pKS/PHB) were 3.025 g/L and 27.83%, respectively. PMID- 10849806 TI - Self-assembling photosynthetic reaction centers on electrodes for current generation. AB - Photosynthetic reaction centers (RCs) made from photosynthetic organisms can be used in solar batteries because their molecules cause light-induced charge separation. We present a simple immobilization system of the intact RCs from Rhodobacter sphaeroides on an electrode that uses nickel ligand binding by the hexameric histidine tag on H subunit (HHisRC). The binding constant of HHisRC to the nickel-nitrilotriacetic acid (Ni-NTA) chip measured with a surface plasmon resonance instrument was 1.6 x 10(8) M-1. HHisRCs were immobilized on an indium tin oxide electrode overlaid with an Ni-NTA gold substrate. The photoinduced displacement current of this electrode was measured to estimate the orientation of HHisRC on the electrode, and the detachability of HHisRC from the electrode was determined by using an imidazole solution wash. The direction of the flash light-induced displacement current suggested that the H subunit side of the immobilized HHisRC faced the surface of the electrode. The photoinduced current disappeared after the electrode was washed in the imidazole solution. This simple immobilization and detachment of HHisRC to the electrode might be useful for making a reproducible photocurrent device. PMID- 10849805 TI - Effect of temperature on growth of psychrophilic and psychrotrophic members of Rhodotorula aurantiaca. AB - The thermo-dependence of growth kinetic parameters was investigated for the Antarctic psychrophilic strain Rhodotorula aurantiaca and a psychrotrophic strain of the same species isolated in Belgium (Ardennes area). Cell production, maximum growth rate (mu max), and half-saturation constant for glucose uptake (Ks) of both yeasts were temperature dependent. For the two yeasts, a maximum cell production was observed at about 0 degree C, and cell production decreased when temperature increased. The mu max values for both strains increased with temperature up to a maximum of 10 degrees C for the psychrophilic strain and 17 degrees C for the psychrotrophic strain. For both yeasts, Ks for glucose was relatively constant at low temperatures. It increased at temperatures above 10 degrees C for the psychrophilic strain and 17 degrees C for the psychrotrophic strain. Although its glucose affinity was lower, the psychrotrophic strain grew more rapidly than the psychrophilic one. The difference in growth rate and substrate affinity was related to the origin of the strain and the adaptation strategy of R. aurantiaca to environmental conditions. PMID- 10849807 TI - Fabrication of an electrode-viologen-hydrogenase heterogeneous system and the electrochemical hydrogen evolution. AB - An indium tin oxide (ITO) electrode was chemically modified by one layer of viologen (VIO) derivative, which possessed a persistent and reproducible electrochemical response. A monolayer of a thermal stable hydrogenase from Thiocapsa roseopersicina was stabilized on a synthesized poly-L-lysine subphase surface and transferred onto the electrode for fabrication of an ITO-VIO hydrogenase heterogeneous system. Electrochemical properties of both the ITO-VIO monolayer and the heterogeneous ITO-VIO-hydrogenase system have been investigated. Hydrogen evolution could be measured by potentiostating the VIO hydrogenase-covered ITO electrode to "electroplate" [(VIO+)n]surf, and a large increase in hydrogen evolution was observed when using an electrolyte solution containing sodium dithionite. We discuss the possible electron transfer process. PMID- 10849808 TI - Successive construction of cellulase hyperproducers of Trichoderma using hyperpolyploids. AB - When the swollen conidia of Trichoderma reesei QM 6a are treated with 0.1% (w/v) colchicine solution, huge autopolyploid nuclei can be formed in those swollen conidia. When a mycelial mat derived from such a conidum is treated with a haploidizing reagent, benomyl, many fan-shaped sectors are produced from the colony, and cellulase hyperproducers are selected from conidia on the colony. When colchicine and benomyl treatments are repeated on cellulase hyperproducers, new hyperproducers can be constructed successively and systematically. Moreover, when conidia derived from autopolyploids are treated with ethylmethanesulfonate solution, another type of cellulase hyperproducers (polyploids) can be obtained. PMID- 10849809 TI - Effect of light/dark cycle on bacterial hydrogen production by Rhodobacter sphaeroides RV. From hour to second range. AB - Hydrogen production by photosynthetic bacteria provides an efficient energy conversion method under low light intensity. However, under strong illumination, such as midday sunlight, the efficiency drops. This prevents the method from being applied industrially. To overcome this problem, we examined a method to thin out the excessive illumination. Light was given intermittently to reduce the total energy flux. The on/off ratio was set at 1/1 throughout the study, so that the time average of the light energy flux became half the continuous illumination. By keeping the time-average light flux constant (0.6 kW.m-2), the effects of the cycle period were examined in the range of hours to seconds. The hydrogen production rate was greatly affected by the cycle period, but cell growth and substrate consumption rates remained almost constant. The 30-min light/dark cycle (30 min on and 30 min off) provided the highest rate of hydrogen production (22 L.m-2.24 h-1). At the shorter cycles, the rate decreased except that there was a suboptimum at about 40 s. Under excessive light intensity (1.2 kW.m-2), the light-to-hydrogen conversion efficiency was greatly enhanced. The hydrogen production rate during the 30-min cycle was twice as high as during a 12 h cycle under the same conditions. PMID- 10849810 TI - Two-dimensional analysis of proteins specific to the bacterial magnetic particle membrane from Magnetospirillum sp. AMB-1. AB - We report the identification of five proteins expressed specifically on the bacterial magnetic particle (BMP) membrane of Magnetospirillum sp. AMB-1. These proteins are major components of the BMP membrane. The molecular weights were determined to be 12.0, 16.0, 24.8, 35.6, and 66.2 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Of these five, the 16.0-kDa protein was the most abundant in the BMP membrane. Furthermore, the 16.0-kDa protein consisted of two components each of differing pI. The 35.6-kDa protein was the second most abundant protein of the five detected. PMID- 10849811 TI - Salinity-regulated replication of the endogenous plasmid pSY10 from the marine cyanobacterium Synechococcus sp. AB - The endogenous plasmid pSY10 in the marine cyanobacterium Synechococcus sp. NKBG042902 is maintained at a high copy number when cells are grown in seawater and at a low copy number when cultured in freshwater. The mechanism of salinity regulated replication of this plasmid was investigated. Transcription of repA was depressed under freshwater, which was accompanied by a low copy number of pSY10 and the appearance of a new protein that was expressed only in cells cultured in freshwater. This protein was observed to bind to putative repA promoters (Prep1 and Prep2) on pSY10. Moreover, this protein was observed only in Synechococcus sp. NKBG042902. The data suggest that this protein(s) regulates repA transcription in pSY10, stress responsive and encoded by the host chromosome. PMID- 10849812 TI - Production of lactic acid from pulp mill solid waste and xylose using Lactobacillus delbrueckii (NRRL B445). AB - Using the simultaneous saccharification and fermentation (SSF) technique, pulp mill solid waste cellulose was converted into glucose using cellulase enzyme and glucose into lactic acid using NRRL B445. SSF experiments were conducted at various pH levels, temperatures, and nutrient concentrations, and the lactic acid yield ranged from 86 to 97%. The depletion of xylose in SSF was further investigated by inoculating NRRL B445 into a xylose-only medium. On prolonged incubation, depletion of xylose with lactic acid production was observed. An experimental procedure with a nonglucose medium was developed to eliminate the lag phase. From xylose fermentation, Lactobacillus delbrueckii yielded 88-92% lactic acid and 2-12% acetic acid. PMID- 10849813 TI - Design and performance of a fibrous bed bioreactor for odor treatment. AB - Biological processes have become popular for odor treatment. In this study, a novel fibrous bed bioreactor was applied for treatment of odorous gas. The column reactor was packed with spirally wound fibrous sheet material on which a consortium of microorganisms selected from activated sludge was immobilized. The first stage of this work comprised a preliminary study that aimed at investigating the feasibility of the fibrous bed bioreactor for treatment of odorous volatile fatty acids (VFAs). In this stage, the performance of a fibrous bed bioreactor at increasing mass loadings ranging from 9.7 to 104.2 g/(m3.h) was studied. VFA removal efficiencies above 90% were achieved at mass loadings up to 50.3 g/(m3.h). At a mass loading of 104.2 g/(m3.h), removal efficiency was found to be 87.7%. In the second stage of the work, the process was scaled up with design and operational considerations, namely, packing medium, process condition, and configuration selections. A trickling biofilter with synthetic fibrous packing medium was selected. It was operated under countercurrent flow of gas and liquid streams. The effects of inlet concentration and empty bed retention time on bioreactor performance were studied. The bioreactor was effective in treating odorous VFAs at mass loadings up to 32 g/(m3.h), at which VFAs started to accumulate in the recirculation liquid, indicating that the biofilm was unable to degrade all the VFAs introduced. Although VFAs accumulated in the liquid phase, the removal efficiency remained above 99%, implying that the biochemical reaction rate, rather than gas-to-liquid mass transfer rate, was the limiting factor of this process. The bioreactor was stable for long-term operation; no clogging and degeneration of the packing medium was observed during the 4-mo operation. PMID- 10849814 TI - alpha-Amylase production by free and immobilized Bacillus subtilis. AB - The effect of glucose on the alpha-amylase production by Bacillus subtilis ATCC 21556 was studied. Initial glucose concentrations up to 20 g/L were found to be directly proportional to the specific alpha-amylase production in an immobilized cell batch system, whereas a free-cell batch system presented an inversely proportional relationship with the initial glucose concentration. This might be owing to the alpha-amylase repression by the glucose present in the culture medium. Three hundred eighty-five percent of the specific alpha-amylase production with the free-cell system was produced by the immobilized-cell batch culture. PMID- 10849815 TI - Effects of trace levels of copper, chromium, and zinc ions on the performance of activated sludge. AB - The effects of copper, chromium, and zinc ions, at trace levels, on the performance of a simulated activated sludge process were investigated. The results of batch adsorption experiments showed that the adsorption of copper, chromium, and zinc ions followed both the Langmuir and Freundlich isotherms. The presence of trace levels of these three metals not only reduced the adsorption rate of organic matters but also the chemical oxygen demand adsorption capacity (CAC) of the activated sludge. Metal ions competed with the organic substrate for adsorption binding sites on the surfaces of activated sludge bioflocs and reduced the CAC. Studies performed in a sequential batch reactor (SBR) showed that the presence of trace levels of heavy metal ions in wastewater affected the SBR performance to different extents depending on the hydraulic retention time (HRT). When the reactors were operated at short HRTs of 2.5 d or less, the presence of trace levels of heavy metal ions reduced substantially the CAC of activated sludge, which, in turn, affected significantly the performance of the SBR. However, under longer HRTs (e.g., 5 d), the heavy metal ions in the wastewater reduced the CAC but had no significant effect on the chemical oxygen demand removal efficiency. PMID- 10849816 TI - Repeated solid-phase fermentation and extraction for enzyme production. AB - Solid-phase fermentation has been found to have a much higher productivity than the popular liquid submerged fermentation in producing cellulase enzymes. The highest reported productivity in the literature for cellulases by Trichoderma cultures in submerged fermentation is 158 filter paper units (FPU)/(h.L) of fermenting liquid. From preliminary experiments of solid-phase fermentation in 1000-mL flasks, a productivity of 234 FPU of cellulases/(h.L) of solid-bed volume was obtained. When two novel techniques--pressure pulsation and repeated extraction--were applied, a productivity of 806 FPU/(h.L) was achieved. The same techniques also greatly enhanced the productivity of other enzymes by fungal cultures in solid-phase fermentation. PMID- 10849817 TI - Ethanol production from glucose and xylose by immobilized Zymomonas mobilis CP4(pZB5). AB - Fermentation of glucose-xylose mixtures to ethanol was investigated in batch and continuous experiments using immobilized recombinant Zymomonas mobilis CP4(pZB5). This microorganism was immobilized by entrapment in kappa-carrageenan beads having a diameter of 1.5-2.5 mm. Batch experiments showed that the immobilized cells cofermented glucose and xylose to ethanol and that the presence of glucose improved the xylose utilization rate. Batch fermentation of rice straw hydrolysate containing 76 g/L of glucose and 33.8 g/L of xylose gave an ethanol concentration of 44.3 g/L after 24 h, corresponding to a yield of 0.46 g of ethanol/g of sugars. Comparable results were achieved with a synthetic sugar control. Continuous fermentation experiments were performed in a laboratory-scale fluidized-bed bioreactor (FBR). Glucose-xylose feed mixtures were pumped through the FBR at residence times of 2-4 h. Glucose conversion to ethanol was maintained above 98% in all experiments. Xylose conversion to ethanol was highest at 91.5% for a feed containing 50 g/L of glucose and 13 g/L of xylose at a dilution rate of 0.24/h. The xylose conversion to ethanol decreased with increasing feed xylose concentration, dilution rate, and age of the immobilized cells. Volumetric ethanol productivities in the range of 6.5-15.3 g/L.h were obtained. The improved productivities achieved in the FBR compared to other bioreactor systems can help in reducing the production costs of fuel ethanol from lignocellulosic sugars. PMID- 10849818 TI - Coproduction of ethanol and glycerol. AB - Ethanol and glycerol are both metabolic products of yeasts. There are occasions when coproduction of both is considered desirable in industrial operations. In this article, we describe the potential of integrating the two processes. A LORRE Y8 yeast culture isolated from molasses is capable of efficient glycerol production from glucose, and a yeast Culture 1400 is an excellent producer of ethanol. By controlling the process conditions, the ratio of ethanol and glycerol production can be varied. PMID- 10849819 TI - Two-stage dilute-acid pretreatment of softwoods. AB - Whole tree chips obtained from softwood forest thinnings were pretreated via single- and two-stage dilute-sulfuric acid pretreatment. Whole-tree chips were impregnated with dilute sulfuric acid and steam treated in a 4-L steam explosion reactor. In single-stage pretreatment, wood chips were treated using a wide range of severity. In two-stage pretreatment, the first stage was carried out at low severity to maximize hemicellulose recovery. Solubilized sugars were recovered from the first-stage prehydrolysate by washing with water. In the second stage, water-insoluble solids from first-stage prehydrolysate were impregnated with dilute sulfuric acid, then steam treated at more severe conditions to hydrolyze a portion of the remaining cellulose to glucose and to improve the enzyme digestibility. The total sugar yields obtained after enzymatic hydrolysis of two stage dilute acid-pretreated samples were compared with sugar yields from single stage pretreatment. The overall sugar yield from two-stage dilute-acid pretreatment was approx 10% higher, and the net enzyme requirement was reduced by about 50%. Simultaneous saccharification and fermentation using an adapted Saccharomyces cerevisiae yeast strain further improved cellulose conversion yield and lowered the enzyme requirement. PMID- 10849820 TI - Simulated dynamics and control of an extractive alcoholic fermentation. AB - In this study, we investigated the dynamics of a computer simulation of a continuous alcoholic fermentation process combined with a flash column under vacuum. The alcohol was partially extracted in order to maintain its concentration at about 40 kg/m3 in the fermentor. The mathematical model of the fermentation was developed for industrial conditions and considers the effect of the temperature on the kinetic parameters. The performance of the dynamic matrix control algorithm, single input single output and multiple input multiple output, for the control of the extractive process was studied. The concepts of factorial design were used in a simulation study to determine the best control structures for the process. PMID- 10849821 TI - Mathematical modeling of controlled-release systems of herbicides using lignins as matrices. A review. AB - The herbicides applied in soils can be easily lost, owing to leaching, volatilization, and bio- and photodegradation. Controlled-release systems using polymeric matrices claim to solve these problems. The movement of the herbicides in the soil is also an important phenomenon to be studied in order to evaluate the loss processes. The development of mathematical models is a relevant requirement for simulation and optimization of such systems. This study reviews mathematical models as an initial step for modeling data obtained for controlled release systems of herbicides (diuron, 2,4-dichlorophenoxyacetic acid, and ametryn) using sugarcane bagasse lignin as a polymeric matrix. The release kinetic studies were carried out using several acceptor systems including a water bath, soil, and soil-packed columns. Generally, these models take into account phenomena such as unsteady-state mass transfer by diffusion (Fick's law) and convection, consumption by several processes, and partitioning processes, resulting in partial differential equations with respect to time and space variables. PMID- 10849822 TI - Influence of lignocellulose-derived aromatic compounds on oxygen-limited growth and ethanolic fermentation by Saccharomyces cerevisiae. AB - Phenolic compounds released and generated during hydrolysis inhibit fermentation of lignocellulose hydrolysates to ethanol by Saccharomyces cerevisiae. A wide variety of aromatic compounds form from lignin, which is partially degraded during acid hydrolysis of the lignocellulosic raw material. Aromatic compounds may also form as a result of sugar degradation and are present in wood as extractives. The influence of hydroxy-methoxy-benzaldehydes, diphenols/quinones, and phenylpropane derivatives on S. cerevisiae cell growth and ethanol formation was assayed using a defined medium and oxygen-limited conditions. The inhibition effected by the hydroxy-methoxy-benzaldehydes was highly dependent on the positions of the substituents. A major difference in inhibition by the oxidized and reduced form of a diphenol/quinone was observed, the oxidized form being the more inhibitory. The phenylpropane derivatives were examined with respect to difference in toxicity depending on the oxidation-reduction state of the gamma carbon, the presence and position of unsaturated bonds in the aliphatic side chain, and the number and identity of hydroxyl and methoxyl substituents. Transformations of aromatic compounds occurring during the fermentation included aldehyde reduction, quinone reduction, and double bond saturation. Aromatic alcohols were detected as products of reductions of the corresponding aldehydes, namely hydroxy-methoxy-benzaldehydes and coniferyl aldehyde. High molecular mass compounds and the corresponding diphenol were detected as products of quinone reduction. Together with coniferyl alcohol, dihydroconiferyl alcohol was identified as a major transformation product of coniferyl aldehyde. PMID- 10849823 TI - Effect of the oxygen transfer coefficient on xylitol production from sugarcane bagasse hydrolysate by continuous stirred-tank reactor fermentation. AB - The effect of the oxygen transfer coefficient on the production of xylitol by bioconversion of xylose present in sugarcane bagasse hemicellulosic hydrolysate using the yeast Candida guilliermondii was investigated. Continuous cultivation was carried out in a 1.25-L fermentor at 30 degrees C, pH 5.5, 300 rpm, and a dilution rate of 0.03/h, using oxygen transfer coefficients of 10, 20, and 30/h. The results showed that the microbial xylitol production (11 g/L) increased by 108% with the decrease in the oxygen volumetric transfer coefficient from 30 to 20/h. The maximum values of xylitol productivity (0.7 g/[L.h]) and yield (0.58 g/g) were obtained akLa 20/h. PMID- 10849824 TI - Study of biocatalyst to produce ethanol from starch. Coimmobilization of glucoamylase and yeast in gel. AB - This article presents a detailed study on the conditions for achieving a stable biocatalyst to be used in the production of ethanol from starch. Different pellets were used depending on which characteristic of the biocatalyst was being studied: (a) Saccharomyces cerevisiae entrapped in pectin or calcium alginate gel particles; (b) silica containing immobilized glucoamylase entrapped in pectin gel particles; or (c) pectin gel particles, with the silica-enzyme derivative and yeast coimmobilized. The influence of several variables on the mechanical resistance of the particle, on the viability of the microorganism, and on the rate of substrate hydrolysis was studied with biocatalyst. The best conditions found were 6% pectin gel, 2-mm particle diameter, and cure in 0.2M CaCl2.2H2O/60 mM acetate buffer, pH 4.2, for gel preparation; and 6.0 g/L of CaCl2.2H2O in the fermentation medium. Biocatalyst (c) was successfully tested for the production of ethanol from liquefied manioc flour syrup. PMID- 10849825 TI - Study of different media for production of penicillin G acylase from Bacillus megaterium ATCC 14945. AB - In this study, several fermentation media were tested for the production of penicillin G acylase (PGA) using Bacillus megaterium. The carbon sources studied were glucose and lactose. The nitrogen sources studied were enzymatic casein hydrolysates produced with proteases of different specificities. The replacement of glucose with cheese whey and the addition of free amino acids in the PGA production were also tested. The results showed a strong correlation between the nitrogen source and enzyme yield and the presence of glucose repression. The highest enzyme concentration achieved was 138 IU/L using casein hydrolyzed with 0.6 L of Alcalase and cheese whey. PMID- 10849826 TI - High-yield fermentation of pentoses into lactic acid. AB - Lactobacillus species capable of fermenting glucose are generally incapable of utilizing xylose for growth or fermentation. In this study, a novel aspect of a well-known Lactobacillus strain, L. casei subsp. rhamnous (ATCC 10863), was uncovered: it can ferment xylose as efficiently as glucose. This strain is a registered organism, extremely stable on long-term operation. Fermentation by this strain is characterized by an initial lag phase lasting 24-72 h before xylose consumption takes place. The yield (grams/gram) of lactic acid from xylose is in excess of 80% with initial volumetric productivity of 0.38 g/(L.h). Acetic acid is the primary byproduct formed at the level of about 10% of the lactic acid. In addition to xylose, it can ferment all other minor sugars in hemicellulose except arabinose. Subjected to mixed sugar fermentation, this strain consumes glucose first, then mannose, followed by almost simultaneous utilization of xylose and galactose. It shows high tolerance for lactic acid as well as extraneous toxins. It can ferment the mixed sugars present in acid treated hydrolysate of softwood, giving yields similar to that of pure sugar but at a slower rate. PMID- 10849827 TI - Cellulase production of Trichoderma reesei Rut C 30 using steam-pretreated spruce. Hydrolytic potential of cellulases on different substrates. AB - Various techniques are available for the conversion of lignocellulosics to fuel ethanol. During the last decade processes based on enzymatic hydrolysis of cellulose have been investigated more extensively, showing good yield on both hardwood and softwood. The cellulase production of a filamentous fungi, Trichoderma reesei Rut C 30, was examined on carbon sources obtained after steam pretreatment of spruce. These materials were washed fibrous steam-pretreated spruce (SPS), and hemicellulose hydrolysate. The hemicellulose hydrolysate contained, besides water-soluble carbohydrates, lignin and sugar degradation products, which were formed during the pretreatment and proved to be inhibitory to microorganisms. Experiments were performed in a 4-L laboratory fermentor. The hydrolytic capacity of the produced enzyme solutions was compared with two commercially available enzyme preparations, Celluclast and Iogen Cellulase, on SPS, washed SPS, and Solka Floc cellulose powder. There was no significant difference among the different enzymes produced by T. reesei Rut C 30. However, the conversion of cellulose using these enzymes was higher than that obtained with Iogen or Celluclast cellulases using steam-pretreated spruce as substrate. PMID- 10849828 TI - Steam pretreatment of Douglas-fir wood chips. Can conditions for optimum hemicellulose recovery still provide adequate access for efficient enzymatic hydrolysis? AB - Douglas-fir sapwood and heartwood were impregnated with SO2 and steam exploded at three severity levels, and the cellulose-rich, water-insoluble component was enzymatically hydrolyzed. The high-severity conditions resulted in near complete solubilization and some degradation of hemicelluloses and a significant improvement in the efficiency of enzymatic digestibility of the cellulose component. At lower severity, some of the hemicellulose remained unhydrolyzed, and the cellulose present in the pretreated solids was not readily hydrolyzed. The medium-severity pretreatment conditions proved to be a good compromise because they improved the enzymatic hydrolyzability of the solids and resulted in the recovery of the majority of hemicellulose in a monomeric form within the water-soluble stream. Sapwood-derived wood chips exhibited a higher susceptibility to both pretreatment and hydrolysis and, on steam explosion, formed smaller particles as compared to heartwood-derived wood chips. PMID- 10849829 TI - Biotreatment of refinery spent-sulfidic caustic using an enrichment culture immobilized in a novel support matrix. AB - Sodium hydroxide solutions are used in petroleum refining to remove hydrogen sulfide (H2S) and mercaptans from various hydrocarbon streams. The resulting sulfide-laden waste stream is called spent-sulfidic caustic. An aerobic enrichment culture was previously developed using a gas mixture of H2S and methyl mercaptan (MeSH) as the sole energy source. This culture has now been immobilized in a novel support matrix, DuPont BIO-SEP beads, and is used to bio-treat a refinery spent-sulfidic caustic containing both inorganic sulfide and mercaptans in a continuous flow, fluidized-bed column bioreactor. Complete oxidation of both inorganic and organic sulfur to sulfate was observed with no breakthrough of H2S and < 2 ppmv of MeSH produced in the bioreactor outlet gas. Excessive buildup of sulfate (> 12 g/L) in the bioreactor medium resulted in an upset condition evidenced by excessive MeSH breakthrough. Therefore, bioreactor performance was limited by the steady-state sulfate concentration. Further improvement in volumetric productivity of a bioreactor system based on this enrichment culture will be dependent on maintenance of sulfate concentrations below inhibitory levels. PMID- 10849830 TI - Lactobacillus plantarum amylase acting on crude starch granules. Native isoforms and activity changes after limited proteolysis. AB - The microheterogeneous native amylolytic complex secreted by the isolate A6 of Lactobacillus plantarum revealed a selective enzyme specificity loss when submitted to a limited proteolysis under a suboptimum pH condition. A clear electrophoretic profile change toward just one shorter, more acidic, and equally active polypeptide fragment resulted from the pronase E pretreatment. Although the whole enzyme activity remained apparently unaffected for soluble starch, the native parallel activity on intact and non-gelatinized starch granules either from cereals or tubers was dramatically reduced. This phenomenon was more clearly documented by scanning electron microscopy using the easiest accessible native substrate: wheat starch granules. The anion-exchange-purified native enzymes from L. plantarum displayed a different optimum pH curve when compared with the thermotolerant alpha-amylase from Bacillus licheniformis. The alpha-amylases from the lactic-acid-producing A6 isolate presented an electrophoretic profile easily distinguishable from those from B. liqueniformis and B. subtilis species. PMID- 10849831 TI - Development of a modified three-stage methane production process using food wastes. AB - A modified three-stage system was developed for the rapid production of methane from food wastes. The primary stage was a semianaerobic hydrolysis/acidogenic system, in which approx 4100 mg/L of volatile fatty acids (VFAs) was produced at a hydraulic retention time (HRT) of 2 d. The operation temperature and pH were 30 degrees C and 5.0-5.5, respectively. The non-degraded materials were removed through a hole at the bottom of the reactor. The secondary stage was an anaerobic acidogenic system equipped with an upflow anaerobic sludge blanket (UASB) type of fermentor. VFA was accumulated up to 6100 mg/L by the addition of Clostridium butyricum to the reactor at an HRT of 2 d. The optimum temperature and pH range were 35 degrees C and 5.0-5.5, respectively. The tertiary methanogenic stage produced CH4 and CO2 from the VFA in the UASB reactor. Methane content was 72% of the total gas volume, and the yield was 0.45-0.50 m3/kg of volatile solids at an HRT of 12 d. The operation temperature and pH were 41 degrees C and 7.6-7.9, respectively. The three-stage process exhibited an unusually high total chemical oxygen demand reduction rate up to 95%. Total nitrogen decreased to 96% and < 10 mg/L of total phosphorus remained in the final effluent. PMID- 10849832 TI - Solid-state fermentation with Aspergillus niger for cellobiase production. AB - Aspergillus niger NRRL3 was cultivated in a moist wheat bran and ground corncob solid medium supplemented with inorganic minerals for the production of cellobiase (beta-1,4-glucosidase, EC 3.2.1.21). With this method, A. niger NRRL3 was able to produce a high concentration of cellobiase (215 IU/g of solid substrate) after 96 h of incubation. Temperature and moisture content affected final cellobiase titers. The best conditions for cellobiase production from solid substrate by A. niger NRRL3 were determined to be 70% moisture and 35 degrees C. PMID- 10849833 TI - Effects of initial pH on biological synthesis of xylitol using xylose-rich hydrolysate. AB - Sugarcane bagasse, an agricultural residue plentiful in Brazil, was utilized for xylitol production by a biotechnological process. A medium fermentation prepared with this xylose-rich biomass at an oxygen transfer volumetric coefficient of 10/h1 and different initial pH values was inoculated with cells of Candida guilliermondii FTI 20037. The maximum values of xylitol and cell volumetric productivities (Qp = 0.56 g/[L.h] and Qx = 0.11 g/[g.h]), xylitol yield factor (YP/S = 0.79 g/g), and xylose uptake rate (qs = 0.197 g/[g.h]) were attained at pH 7.0 without further pH control. The results show that the yeast performance was influenced by the pH, an important bioengineering parameter in this fermentation process. PMID- 10849834 TI - Effect of shear on human insulin in zinc suspension. AB - Human insulin in zinc suspension was used as a model protein to test the effect of shear on the settling rate of proteins, a possible inference for protein denaturation. The rate of settling was determined directly in a spectrophotometer. Shear effects are important in retaining the activity of proteins and are present in bubble, foam, and droplet protein fractionation processes. A simple test, such as that conducted here, may even be useful for monitoring changes in protein structure caused by commercial shipping of the protein. The settling rate for insulin was continuously monitored in the original bottle by spectrophotometric absorbance changes as a function of time. A settling curve was determined following each shear experiment, which included shaking the "worked" insulin solution in a vortex mixer for different lengths of time. It was determined, when comparing long shaking times with short ones, that the initial settling rate was less for the long-term shaking of the insulin samples and greater for the short-term shaking. The secondary effects of light and heat, along with shaking, apparently did not produce differences from shaking alone. PMID- 10849835 TI - Screening of potential antibiotic action of cellulolytic fungi. AB - Twenty different strains of filamentous fungi were initially selected for evaluation of cellulolytic activity using a single test in a simple mineral salts culture medium with filter paper as the only carbon source. Those fungi strains that were capable of completely breaking the filter paper strip within 4-8 d were assayed also for antimicrobial action, using Staphyloccocus aureus ATCC 6538P according to the so-called agar piece method. We screened three different strains with both capacities: the production of cellulolytic activity and antibiotic action. The experimental results suggest that the fungi Penicillium sp. F0PC01, Aspergillus sp. F0Q001, and Cephalosporium sp. F03800 have both capabilities because they grew rapidly on cellulose as the only carbon source and were able to produce an area of growth inhibition in S. aureus of approx 2.04, 1.57, and 2.39 cm, respectively, on agar plates using the agar piece method. Subsequently, the antibiotic production obtained with those cellulolytic strains was evaluated by submerged fermentation at the flask level, in a simple culture medium containing lactose without biosynthesis precursor, obtaining 3670, 2830, and 4060 antibiotic units/mL, referred to as penicillin G, whereas for cellulolytic activity, the results were 1.34, 1.81 and 0.57 FPU/mL, respectively. PMID- 10849836 TI - Mycelial pellet formation by Rhizopus oryzae ATCC 20344. AB - Factors in a cultivation medium affecting fungal growth morphology and fumaric acid production by Rhizopus oryzae ATCC 20344 were investigated. These factors included the initial pH value and trace metals such as zinc, magnesium, iron, and manganese in the cultivation medium. It was found that a significant change in the growth morphology of R. oryzae ATCC 20344 occurs when the initial pH value is varied. A lower initial pH value in the cultivation medium was inhibitory to fungal growth, and fast growth in the cultivation medium at a higher initial pH value promoted the formation of large pellets or filamentous forms. Trace metals in the cultivation media also had significant effects on pellet formation and fumaric acid fermentation. PMID- 10849837 TI - L-DOPA production by immobilized tyrosinase. AB - The production of L-DOPA using L-tyrosine as substrate, the enzyme tyrosinase (EC 1.14.18.1) as biocatalyst, and L-ascorbate as reducing agent for the o-quinones produced by the enzymatic oxidation of the substrates was studied. Tyrosinase immobilization was investigated on different supports and chemical agents: chitin flakes activated with hexamethylenediamine and glutaraldehyde as crosslinking agent, chitosan gel beads, chitosan gel beads in the presence of glutaraldehyde, chitosan gel beads in the presence of polyvinylpyrrolidone, and chitosan flakes using glutaraldehyde as crosslinking agent. The last support was considered the best using as performance indexes the following set of immobilization parameters: efficiency (90.52%), yield (11.65%), retention (12.87%), and instability factor (0.00). The conditions of immobilization on chitosan flakes were optimized using a two-level full factorial experimental design. The independent variables were enzyme-support contact time (t), glutaraldehyde concentration (G), and the amount of enzyme units initially offered (UC). The response variable was the total units of enzymatic activity shown by the immobilized enzyme (UIMO). The optimal conditions were t = 24 h, G = 2% (v/v), and UC = 163.7 U. Under these conditions the total units of enzymatic activity shown by the immobilized enzyme (UIMO) was 23.3 U and the rate of L-DOPA production rate was 53.97 mg/(L.h). PMID- 10849838 TI - Inhibition of microbial xylitol production by acetic acid and its relation with fermentative parameters. AB - Precipitated sugarcane bagasse hemicellulosic hydrolysate containing acetic acid was fermented by Candida guilliermondii FTI20037 under different operational conditions (pH 4.0 and 7.0, three aeration rates). At pH 7.0 and kLa of 10 (0.75 vvm) and 22.5/h (3.0 vvm) the acetic acid had not been consumed until the end of the fermentations, whereas at the same pH and kLa of 35/h (4.5 vvm) the acid was rapidly consumed and acetic acid inhibition was not important. On the other hand, fermentations at an initial pH of 4.0 and kLa of 22.5 and 35/h required less time for the acid uptake than fermentations at kLa of 10/h. The acetic acid assimilation by the yeast indicates the ability of this strain to ferment in partially detoxified medium, making possible the utilization of the sugarcane bagasse hydrolysate in this bio-process. The effects on xylitol yield and production are reported. PMID- 10849839 TI - Anaerobic digestion from residue of industrial cassava industrialization with acidogenic and methanogenic physical separation phases. AB - A trial was carried out in a continuous regimen, using a completely stirred tank reactor, at acidogenic phase, and a hybrid reactor (upflow anaerobic sludge blanket + fixed bed) at methanogenic phase at room temperature. The residue to be treated came from a flour and cassava meal industry, and the reactors operated for 300 d with affluent chemical oxygen demand (COD) concentrations of 7500, 9000, 11,000, and 14,000 mg/L. The final results showed a biogas production with a content of 80% methane and an average reduction of COD and free cyanide of nearly 96 and 98%, respectively. The separation of phases selected bacterial groups. At acidogenic phase, a predominance of propionic, n-butyric, and n valeric acids, as well as a biomass composed of 95% fermentative bacilli, which were responsible for a 90% reduction in free cyanide concentration, was observed. At methanogenic phase, a predominance of methanogenic bacteria that came only from the Methanothrix genus was observed. The bacteria were responsible for high levels of organic matter removal and methane production. PMID- 10849840 TI - Use of corncob for endoxylanase production by thermophilic fungus Thermomyces lanuginosus IOC-4145. AB - The production of cellulase-free endoxylanase by the thermophilic fungus Thermomyces lanuginosus was investigated in semisolid fermentation and liquid fermentation. Different process variables were investigated in semisolid fermentation, employing corncob as the carbon source. The best results were with the following conditions: grain size = 4.5 mm, solid:liquid ratio = 1:2, and inoculum size = 20% (v/v). Corncob, xylan, and xylose were the best inducers for endoxylanase production. Additionally, organic nitrogen sources were necessary for the production of high endoxylanase activities. The crude enzyme had optimum activity at pH 6.0 and 75 degrees C, displaying a high thermostability. The apparent Km and Vmax were 1.77 mg of xylan/mL and 21.5 U/mg of protein, respectively. PMID- 10849841 TI - Effect of protein denaturation on void fraction in foam separation column. AB - Foam fractionation is a cost-effective process that uses air to extract protein from a liquid (in this case "crude" dilute egg-albumin solution). This article deals with how the void fraction (fraction of air in the aerated solution) of foam is affected by heat denaturation of the protein. A 2-mm glass tube was used to sample the foam-liquid interface fluid in a 35-mm-diameter column in order to detect small changes in void fraction and foam production, which are not easily detected directly from the bulk foam. The main control variable in this study was the protein solution preheating time. As the preheating time increased, the initial void fraction in the column decreased. The initial void fraction of the undenatured solution ranged from about 0.73 to 0.80, and the void fraction for significant preheating times of 5 min ranged from approx 0.68 to 0.72. Furthermore, the period of foam production increased from 5 to 7 min for undenatured proteins in solution to as long 15 min for 5-min preheated solutions. Side-port sampling through a small capillary tube has the potential to be used as a rapid and inexpensive way to determine the level of protein denaturation by directly determining the void fraction and then estimating the effect of denaturation from a protein denaturation calibration curve of the void fraction. PMID- 10849842 TI - Microbial production of polyhydroxyalkanoates by bacteria isolated from oil wastes. AB - A Gram-positive coccus-shaped bacterium capable of synthesizing higher relative molecular weight (M(r)) poly-hydroxybutyrate (PHB) was isolated from sesame oil and identified as Staphylococcus epidermidis (by Microbial ID, Inc., Newark, NJ). The experiment was conducted by shake flask fermentation culture using media containing fructose. Cell growth up to a dry mass of 2.5 g/L and PHB accumulation up to 15.02% of cell dry wt was observed. Apart from using single carbohydrate as a sole carbon source, various industrial food wastes including sesame oil, ice cream, malt, and soya wastes were investigated as nutrients for S. epidermidis to reduce the cost of the carbon source. As a result, we found that by using malt wastes as nutrient for cell growth, PHB accumulation of S. epidermidis was much better than using other wastes as nutrient source. The final dried cell mass and PHB production using malt wastes were 1.76 g/L and 6.93% polymer/cells (grams/gram), and 3.5 g/L and 3.31% polymer/cells (grams/gram) in shake flask culture and in fermentor culture, respectively. The bacterial polymer was characterized by 1H-nuclear magnetic resonance (NMR), 13C-NMR, Fourier transform infrared, and differential scanning calorimetry. The results show that with different industrial food wastes as carbon and energy sources, the same biopolymer (PHB) was obtained. However, the use of sesame oil as the carbon source resulted in the accumulation of PHB with a higher melting point than that produced from other food wastes as carbon sources by this organism under similar experimental conditions. PMID- 10849843 TI - Improvements in titer, productivity, and yield using Solka-floc for cellulase production. AB - Researchers studying cellulase enzymes for the economical production of fuel ethanol envision cellulose as the carbon source. However, submerged Trichoderma reesei cultures grown on cellulose exhibit high run-to-run variability. Thus, an investigation of 30 batch cellulase production experiments was instrumental in determining fermentation conditions that improved enzyme titers, yields, and productivities. Eighteen of the 30 batch experiments experienced minimal process upsets and were classified into eight groups based on agitation rate, gas sparge rate, and the use of oxygen supplementation. Comparing corn steep liquor with yeast extract/peptone also tested the effect of different sources of nitrogen in the media. Average 7-d enzyme titers were doubled from 4 to 8 FPU/mL primarily by increasing aeration. PMID- 10849844 TI - Biodegradability of new engineered fuels compared to conventional petroleum fuels and alternative fuels in current use. AB - Concern with environmental issues such as global climate change has stimulated research into the development of more environmentally friendly technologies and energy sources. One critical area of our economy is liquid transportation fuels. This article presents the results of the biodegradability potential of newly developed engineered fuels and compares the results to the biodegradability of conventional fuels and alternative fuels in current use. Biodegradability potential was determined under both aerobic and anaerobic conditions. Fuels that have a high degree of components derived from renewable sources proved to have a higher degradability potential than those composed of petroleum components. PMID- 10849845 TI - Cloning and expression of the limonene hydroxylase of Bacillus stearothermophilus BR388 and utilization in two-phase limonene conversions. AB - A 3.6-kb fragment of Bacillus stearothermophilus BR388 chromosomal DNA that confers growth on limonene to Escherichia coli has been sequenced, revealing a single open reading frame encoding a single subunit limonene hydroxylase containing 444 amino acid residues. This enzyme proved capable of limonene hydroxylation to a mixture of carveol and perillyl alcohol as well as dehydrogenation of these products to carvone and perillyl aldehyde. Oxygen, FAD, and NADH were found to stimulate the hydroxylation reaction in cell extracts, and NAD+ stimulated the dehydrogenase reaction. In two-phase bioconversions using viable E. coli cells over-expressing the limonene hydroxylase, perillyl alcohol and carvone were the principal products observed. PMID- 10849846 TI - Biosurfactants from potato process effluents. AB - High-solids (HS) and low-solids (LS) potato process effluents were tested as substrates for surfactin production. Tests used effluents diluted 1:10, unamended and amended with trace minerals or corn steep liquor. Heat pretreatment was necessary for surfactin production from effluents due to indigenous bacteria, whose spores remained after autoclaving. Surfactin production from LS surpassed HS in all cases. Surfactin yields from LS were 66% lower than from a pure culture in an optimized potato starch medium. LS could potentially be used without sterilization for surfactin production for low-value applications such as environmental remediation or oil recovery. PMID- 10849847 TI - A kinetic study of synthesis of amoxicillin using penicillin G acylase immobilized on agarose. AB - We present a kinetic model for the synthesis of amoxicillin from p hydroxyphenylglycine methyl ester and 6-aminopenicillanic acid, catalyzed by penicillin G acylase immobilized on agarose, at 25 degrees C. Michaelis-Menten kinetic parameters (with and without inhibition) were obtained from initial velocity data (pH 7.5 and 6.5). Amoxicillin synthesis reactions were used to validate the kinetic model after checking mass transport effects. A reasonable representation of this system was achieved under some operational conditions, but the model failed under others. Nevertheless, it will be useful whenever a simplified model is required, e.g., in model-based control algorithms for the enzymatic reactor. PMID- 10849848 TI - Enzyme electrochemical preparation of a 3-keto derivative of 1,5-anhydro-D glucitol using glucose-3-dehydrogenase. AB - A novel enzymatic organic synthesis was reported, utilizing glucose-3 dehydrogenase (G3DH) and its regeneration via electrochemical methods. We combined the water-soluble G3DH prepared from a marine bacterium, Halomonas sp. alpha-15, and electron mediator with the electrode system in order to regenerate the enzyme. Using this system, the conversion of 1,5-anhydro-D-glucitol (1,5AG), a diabetes marker in human blood, was investigated. The final yield of the product, 3-keto anhydroglucitol (3-ketoAG), which was identified by 13C nuclear magnetic resonance, was 82% based on the initial amount of 1,5AG. The electrochemical yield of the reaction proceeded almost stoichiometrically. The electrochemical conversion rate of 1,5AG was 1.24 mmol/(L.h), and the electrochemical yield of 1,5AG consumption was 80%, whereas that for 3-ketoAG was 60%. PMID- 10849849 TI - Enhancement of selectivity for producing gamma-cyclodextrin. AB - The production of cyclodextrins (CDs) by cyclodextrin-glycosyl-transferase (CGTase) from Bacillus firmus was studied, with respect to the effect of the source of starch upon CD yield and on the selectivity for producing gamma-CD. Cyclodextrin production tests were run for 24 h at 50 degrees C, pH 8.0, and 1 mg/L of CGTase, and substrates were maltodextrin or the starches of rice, potato, cassava, and corn hydrolyzed up to D.E. 10. Cornstarch was the best substrate for producing gamma-CD. Later, glycyrrhizin (2.5% [w/v]), which forms a stable complex with gamma-CD, was added to the cornstarch reaction medium and increased the yield of gamma-CD to about four times that produced with only maltodextrin, but the total yield of CDs remained practically unchanged. Therefore, the results showed that the studied CGTase is capable of giving relatively high yield of gamma-CD in the presence of glycyrrhizin as complexant and cornstarch as substrate. PMID- 10849850 TI - Simultaneous saccharification and fermentation of cellulosic biomass to acetic acid. AB - A strain of Clostridium thermoaceticum (ATCC 49707) was evaluated for its homoacetate potential. This thermophilic anaerobe best produces acetate from glucose at pH 6.0 and 59 degrees C with a yield of 83% of theoretical. Enzyme hydrolysis of two substrates, a-cellulose and a pulp mill sludge, yielded 68% and 70% digestion, respectively. The optimum conditions for the simultaneous saccharification and fermentation (SSF) were substrate dependent: 55 degrees C, pH 6.0 for alpha-cellulose, and 55 degrees C, pH 5.5 for the pulp mill sludge. In the SSF with alpha-cellulose, the overall yield of acetate was strongly influenced by the enzyme loading. In a fed-batch operation of SSF with alpha cellulose, an overall acetic acid yield of 60 wt% was obtained. Among the factors limiting the yields were incomplete digestion by the enzyme and the end-product inhibition. In the SSF of pulp mill sludge, inhibitors present in the sludge severely limited bacterial action. A large accumulation of glucose developed over the entire process, changing the intended SSF operation into a separate hydrolysis and fermentation operation. Despite a long lag phase of microbial growth, a terminal yield of 85% was obtained with this substrate. PMID- 10849851 TI - Effect of C:N molar ratio on monomer composition of polyhydroxyalkanoates produced by Pseudomonas mendocina 0806 and Pseudomonas pseudoalkaligenus YS1. AB - Polyhydroxyalkanoates (PHAs) are biodegradable polymers produced by bacteria. In this study, the effect of C:N molar ratio on the monomer composition of PHAs was investigated, including medium chain length PHA produced by Pseudomonas mendocina 0806 and PHA blends consisting of monomers of 3-hydroxybutyrate and medium chain length hydroxyalkanoate produced by Pseudomonas pseudoalkaligenus YS1. It was observed that there were some fixed ranges of C:N molar ratio that affect the monomer composition of PHA independently of the substrate. For strain 0806, the ranges were C:N < 20, 20 < C:N < 200, and C:N > 200. The monomer composition was constant among these ranges when using glucose and octanoate as the sole substrate. For strain YS1, the ranges were C:N < 20, 20 < C:N < 45, and C:N > 45. These results are useful for controlling monomer composition in PHA production. PMID- 10849852 TI - Optimal production of polyhydroxyalkanoates in activated sludge biomass. AB - Polyhydroxyalkanoates (PHAs) have been recognized as good candidates for biodegradable plastics, but their high price compared with conventional plastics has limited their use. In this study, activated sludge microorganisms from a conventional wastewater treatment process were induced, by controlling the carbon:nitrogen (C:N) ratio in the reactor liquor, to accumulate PHAs. In addition, an intermittent nitrogen feeding program was established to optimize the volumetric PHA productivity in a wastewater treatment process. The optimal overall polymer production yield of 0.111 g of polymer/g of carbonaceous substrate consumed was achieved under a C:N ratio of 96:1 by feeding nitrogen in the reactor liquor once every four cycles. At the same time, the amount of excess sludge generated from the wastewater treatment process was reduced by 22.9%. PMID- 10849853 TI - Polyhydroxybutyrate production from carbon dioxide by cyanobacteria. AB - Genetic characterization and enhancement of polyhydroxybutyrate (PHB) accumulation in cyanobacteria were investigated for efficient PHB production from CO2. The genome DNAs in the PHB-accumulating strains Synechococcus sp. MA19 and Spirulina platensis NIES46 retained the highly homologous region to phaC of Synechocystis PCC6803, whereas low homology was detected in the nonaccumulating strains Synechococcus sp. PCC7942 and Anabaena cylindrica NIES19. Synechococcus sp. MA19, which accumulates PHB up to 30% of dry cell weight from CO2 as the sole carbon source, was mutated by insertion of transposon Tn5 to enhance the PHB accumulation. Genetic and physiological analysis of the mutant indicated that decreased phosphotransacetylase activity could trigger an increase of acetyl coenzyme A leading to enhancement of PHB accumulation. PHB synthase in Synechococcus sp. MA19 was probably attached to thylakoid membrane since PHB granules were associated with pigments. A genetically engineered cyanobacteria retaining soluble PHB synthase from Ralstonia eutropha accumulated pigment-free PHB granules, which is an advantage for the purification of PHB. PMID- 10849854 TI - Production of cyclodextrins in a fluidized-bed reactor using cyclodextrin glycosyl-transferase. AB - Cyclodextrin-glycosyl-transferase (EC2.4.1.19), produced by Wacker (Munich, Germany), was purified by biospecific affinity chromatography with beta cyclodextrin (beta-CD) as ligand, and immobilized into controlled pore silica particles (0.42 mm). This immobilized enzyme (IE) had 4.7 mg of protein/g of support and a specific activity of 8.6 mumol of beta-CD/(min.gIE) at 50 degrees C, pH 8.0. It was used in a fluidized-bed reactor (FBR) at the same conditions for producing cyclodextrins (CDs) with 10% (w/v) maltodextrin solution as substrate. Bed expansion was modeled by the Richardson and Zaki equation, giving a good fit in two distinct ranges of bed porosities. The minimum fluidization velocity was 0.045 cm/s, the bed expansion coefficient was 3.98, and the particle terminal velocity was 2.4 cm/s. The FBR achieved high productivity, reaching in only 4 min of residence time the same amount of CDs normally achieved in a batch reactor with free enzyme after 24 h of reaction, namely, 10.4 mM beta-CD and 2.3 mM gamma-CD. PMID- 10849855 TI - Occlusion of transition metal ions by new adsorbents synthesized from plant polyphenols and animal fibrous proteins. AB - Removing and collecting heavy and rare metal ions from industrial effluents and waste aqueous solutions are important problems. Our previous studies showed that animal fibrous proteins (AFPs) such as hen eggshell membrane, chicken feather (CF), wool, and silk were stable and insoluble proteins and had an excellent ability to bind not only hard (Mn2+ and Fe3+) but also soft (Ag+, Au+, Pd2+, Pt2+, and Hg2+) acids from aqueous solutions. In this study, we synthesized some adsorbents for transition (Cr6+, Mn2+, Co2+, Ni2+, and Cu2+) and heavy (Cd2+) metal ions from AFPs (gelatin, CF, and wool) and plant polyphenols (lignin and tannin) by heating a mixture of AFPs and plant polyphenols under acidic conditions. In batch experiments, pH profile, time dependency, and isotherm analysis were performed to determine binding properties of adsorbents for transition and heavy metal ions. Column experiments were also performed to remove copper ion from aqueous solution. The results showed that the new adsorbents were effective for collecting and removing transition and heavy metal ions from aqueous solutions. PMID- 10849856 TI - Phosphotransacetylase as a key factor in biological production of polyhydroxybutyrate. AB - Phosphotransacetylase (Pta) catalyzes the reversible conversion of acetyl coenzyme A (CoA) to acetyl phosphate. Polyhydroxybutyrate (PHB) synthase and accumulation were compared between a Pta-deficient mutant and the wild-type Escherichia coli, which were transformed with pAE100, coding for 3-ketothiolase, NADPH-dependent acetoacetyl-CoA reductase, and PHB synthase from Ralstonia eutropha. During the growth period, PHB synthase activity in the Pta-deficient mutant was lower than that in the wild type. PHB accumulation in the Pta deficient mutant, however, was higher than that in wild-type cells grown in Luria Bertani (LB) medium containing 1% glucose (high C:N ratio). The Pta-deficient mutant showed PHB accumulation even in LB medium (low C:N ratio), whereas wild type cells showed no PHB accumulation. These data suggest the activation of PHB synthase by acetyl phosphate that is synthesized by Pta. A decrease in Pta activity probably causes some increase in acetyl-CoA as substrate for the PHB synthesis pathway, resulting in increased PHB accumulation. PMID- 10849857 TI - Recovery of volatile products from dilute high-fouling process streams. AB - As biomass hydrolysis and fermentation technologies approach commercial viability, advancements in product recovery technologies will be required. For cases in which fermentation products are more volatile than water, recovery by distillation is often the technology of choice. Distillation technologies that will allow the economic recovery of dilute volatile products from streams containing a variety of impurities have been developed and commercially demonstrated. Distillation tower and tray designs, along with specialized heat exchanger designs, allowing for extended processing intervals on solutions containing lignocellulosic residues, organic acids, and inorganic salts concentrations > 100 g/L are in commercial operation. In the case of ethanol, distillation energy consumption efficiencies for processing solutions containing < 40 g/L of desired product can approach demonstrated energy consumption efficiencies for solutions containing concentrations > 120 g/L. These proprietary technologies have been applied individually at commercial scale, and designs have been developed that incorporate the combined technologies with only a marginal increase in capital investment compared to traditional methods. PMID- 10849858 TI - Adsorption of inulinases in ion-exchange columns. AB - The use of adsorption columns packed with ion-exchange resins for recovering, concentrating and purifying proteins is now widespread. The present work consists of a study on the dynamic behavior of adsorption columns that uses two kinds of adsorbents: a cationic and an anionic resin. A frontal analysis of the columns was performed with experimental data obtained from Fructozyme, a mixture of inulinase enzymes. The parameters of a Langmuir type of isotherm and adsorption kinetics were obtained from experimental tests in a batch system. A numerical technique based on orthogonal collocation and a fourth-order Runge-Kutta method was coupled with a nonlinear optimization method to predict the coefficients of the rate equations, which are fundamental for scale-up purposes. PMID- 10849859 TI - Sonic wave separation of invertase from a dilute solution to generated droplets. AB - It has previously been shown that a droplet fractionation process, simulated by shaking a separatory funnel containing a dilute protein solution, can generate droplets richer in protein than present in the original dilute solution. In this article, we describe an alternative method that can increase the amount of protein transferred to the droplets. The new method uses ultrasonic waves, enhanced by a bubble gas stream to create the droplets. The amount of protein in these droplets increases by about 50%. In this method, the top layer of the dilute protein solution (of the solution-air interface) becomes enriched in protein when air is bubbled into the solution. This concentrating procedure is called bubble fractionation. Once the protein has passed through the initial buildup, this enriched protein layer is transferred into droplets with the aid of a vacuum above the solution at the same time that ultrasonic waves are introduced. The droplets are then carried over to a condenser and coalesced. We found that this new method provides an easier way to remove the protein-enriched top layer of the dilute solution and generates more droplets within a shorter period than the separatory funnel droplet generation method. The added air creates the bubbles and carries the droplets, and the vacuum helps remove the effluent airstream from the condenser. The maximum partition coefficient, the ratio of the protein concentration in the droplets to that in the residual solution (approx 8.5), occurred at pH 5.0. PMID- 10849860 TI - Modeling a protein foam fractionation process. AB - A simple staged model for the protein foam fractionation process is proposed in this article. This simplified model does not detail the complex foam structure and gas-liquid hydrodynamics in the foam phase but, rather, is built on the conventional theoretical stage concept considering upward bubbles with entrained liquid and downward liquid (drainage) as counter-current flows. To simulate the protein concentration distribution in the liquid along the column by the model, the bubble size and liquid hold-up with respect to the position must be known, as well as the adsorption isotherm of the protein being considered. The model is evaluated for one stage by data from the semibatch foam fractionation of egg albumin and data from the continuous foam fractionation of bovine serum albumin. The effect of two significant variables (superficial gas velocity and feed protein concentration) on enrichment is well predicted by the model, especially for continuous operation and semibatch operation when initial concentration is high. PMID- 10849861 TI - beta-Xylosidase recovery by reversed micelles. Use of cationic surfactant. AB - beta-Xylosidase, an enzyme produced by Penicillium janthinellum fungus, was prepurified by fractionated precipitation with ethanol and extracted by reversed micelles of N-benzyl-N-dodecyl-N-bis(2-hydroxyethyl) ammonium chloride (BDBAC) cationic surfactant. A 2(5-1) fractional factorial design was employed to evaluate the influence of the following factors on the enzyme extraction: pH (A), conductivity (B), surfactant concentration (C), cosolvent concentration (D) and temperature (E). A statistical analysis of the results revealed that, of the five variables studied, pH, surfactant concentration, and temperature had significant main effects (p < 0.05) on the recovery of beta-xylosidase (Y). However, the interactions between pH and temperature, conductivity and cosolvent concentration, conductivity and temperature, BDBAC concentration and temperature also had significant influences. A first-order model (R2 = 0.98) expressed by the equation Y = 7.73 - 5.55A - 0.67B + 3.49C - 0.41D + 5.26E + 2.05AB - 3.26AC + 0.96AD - 7.07AE - 3.93BD - 2.19BE + 0.99CD + 0.78CE was proposed to represent the enzyme recovery as a function of the effects that were really significant. This model predicts a recovery value of about 40%, which is similar to that obtained experimentally (39.5%). PMID- 10849862 TI - A novel magnetite-immobilized cell process for heavy metal removal from industrial effluent. AB - The sorption and desorption of copper (II) (Cu[II]) ions from the wastewater by magnetite-immobilized cells of Pseudomonas putida 5-x with acidic pretreatment were studied. Pretreating cells with 0.6 N HCl was found to enhance greatly the adsorption capacity of biomass up to 85.6 mg/g and had no significant effect on the loss of P. putida 5-x cells during biosorbent pretreatment. The biosorption capacity to Cu2+ of magnetite-immobilized cells of P. putida 5-x harvested during various growth phases was also investigated. The experimental results illustrated that the adsorption capacity to Cu2+ of P. putida 5-x cultured in sulfate limiting medium reached maximum during the late stationary growth phase or early death phase, and reached minimum during the log growth phase. The mechanism of copper sequestering by this type of biomass was studied via transmission electron microscopy. A degradation of the peptidoglycan layer of the cell wall was observed in the acidic pretreatment, but no further degradation appeared after the adsorption-desorption cycle. Cu(II) accumulated mostly on the surface of the cell walls and was effectively desorbed by the acidic treatment during the desorption process. PMID- 10849863 TI - Effect of food:microorganism ratio in activated sludge foam control. AB - Foaming is a common operational problem in activated sludge processes that often adversely affects the quality of the treated effluent. Overgrowth of the filamentous Nocardia spp. in the microbial ecosystem was previously identified as the cause of foaming. In the present study, the specific growth rate of Nocardia amarae was found to be much higher than that of nonfilamentous bacteria under food:microorganism (F:M) ratios lower than 0.5 mg of biological oxygen demand (BOD)/(mg of mixed liquor suspended solids [MLSS].d). This indicated that filamentous overgrowth may occur in normal activated sludge processes that are continually operated under the usual F:M range of 0.2-0.6 mg of BOD/(mg of MLSS.d). A novel two-component feast-fast operation (FFO) that capitalized on the sensitivity of filamentous bacteria to F:M ratio was designed to prevent and control foaming problems. The F:M ratio in the "feasting" aeration unit was 0.8 mg of BOD/(mg of MLSS.d) whereas that in the "fasting" aeration unit was 0.2 mg of BOD/(mg of MLSS.d). The FFO resulted in an overall process F:M ratio that still remained within the normal range, while avoiding prolonged exposure of the activated sludge ecosystem to an F:M ratio below 0.5 mg of BOD/(mg of MLSS.d). The FFO suppressed the overgrowth of filamentous bacteria without adversely affecting the organic treatment efficiency of the modified process. PMID- 10849864 TI - Lipase production by Penicillium restrictum using solid waste of industrial babassu oil production as substrate. AB - Lipase, protease, and amylase production by Penicillium restrictum in solid-state fermentation was investigated. The basal medium was an industrial waste of babassu oil (Orbignya oleifera) production. It was enriched with peptone, olive oil, and Tween-80. The supplementation positively influenced both enzyme production and fungal growth. Media enriched with Tween-80 provided the highest protease activity (8.6 U/g), whereas those enriched with peptone and olive oil led to the highest lipase (27.8 U/g) and amylase (31.8 U/g) activities, respectively. PMID- 10849865 TI - Brazilian bioethanol program. AB - Brazil is the largest producer of bioethanol, and sugarcane is the main raw material. Bioethanol is produced by both batch and continuous processes, and in some cases, flocculating yeast is used. This article analyzes the Brazilian Ethanol Program. For the 1996-1997 harvest, Brazil produced 14.16 billion L of ethanol and 13.8 million metric t of sugar, from 286 million metric t of sugarcane. These products were produced by 328 industries in activity, with 101 autonomous ethanol plants producing only ethanol, and 227 sugar mills producing sugar and ethanol. The sugar-ethanol market reaches about 7.5 billion US$/yr, accounting for direct and indirect revenues. PMID- 10849866 TI - Energy metabolism and thyroid hormone levels of growing rats in response to different dietary proteins--soy protein or casein. AB - Energy balances were measured by indirect calorimetry in four experiments on male growing rats, fed restrictively on isoenergetic and isonitrogenous (10% CP) diets based on either casein supplemented with methionine, or soy protein isolate (experiments 1, 2 and 3) and soy protein isolate supplemented with methionine (experiment 0), respectively. At the end of experiments the rats were killed for body analysis and determination of thyroid hormones and lipids in blood as well as mitochondrial respiration in liver and heart. Feeding of non-supplemented soy protein resulted in a lower efficiency of energy utilisation as well as a lower protein utilisation compared to casein-fed and supplemented soy protein-fed rats. Chemical body composition was not markedly different between the dietary groups. After long-term feeding of soy protein (experiment 3) mass and mitochondrial protein content of the interscapular brown adipose tissue were increased compared to casein-fed rats. Serum thyroid hormone levels were not changed (T3 and free T3) or were significantly lowered (T4 and free T4) following soy protein feeding in comparison with casein feeding (except for experiment 2). Cholesterol and triglycerides were decreased on an average in response to soy protein or supplemented soy protein feeding. In two of three experiments a significant lower efficiency of hepatic mitochondrial respiration with succinate as substrate, expressed by the ratio of added ADP to oxygen consumed, was observed in soy protein-fed rats compared to casein-fed rats. PMID- 10849867 TI - Selected animal and plant protein sources affect nutrient digestibility and fecal characteristics of ileally cannulated dogs. AB - The objective of this study was to compare ileal and total tract nutrient digestibilities and fecal characteristics of dogs fed selected animal and plant protein sources incorporated into grain-based diets. Four crude protein sources- soybean meal (SBM), poultry meal (PM), poultry by-product meal (PBPM), and beef and bone meal (BBM)--were fed to four ileal cannulated dogs in a 4 x 4 Latin square design. Intakes of dry matter, organic matter, crude protein, and fat by dogs were similar. Total dietary fiber (TDF) intake was highest for the SBM treatment compared to other treatments. Ileal digestibilities of DM, OM, CP, fat, and TDF were not affected by treatment. Total tract digestibility of DM was lower for the BBM and SBM diets, while OM digestibility was lower for the SBM treatment only. Total tract CP digestibility was similar for BBM, PBPM, and SBM treatments and was higher for the PM treatment. As-is fecal excretion [g/d] was greater when dogs received the SBM treatment. Fecal output on a DM basis was higher for the SBM treatment compared to the other treatments. All diets were well utilized by the dogs as assessed by ileal and total tract digestibility data and fecal characteristics. PMID- 10849868 TI - Influence of the dietary ratio between sulphur containing amino acids and lysine on performance of growing-finishing pigs fed diets with various lysine concentrations. AB - This study was conducted to determine the optimal ratio between sulphur containing amino acids and lysine in diets for growing-finishing pigs. Therefore, a total of five trials was carried out in which growing-finishing pigs (live weight range between 53 and 105 kg) were fed diets with various concentrations of lysine (0.62, 0.70 and 0.78%) and various ratios between sulphur containing amino acids to lysine. The diets contained 12.9 MJ ME per kg and 13.5% CP; the ratio between sulphur containing amino acids to lysine was adjusted by individual supplementation of the diets with DL-methionine. Increasing dietary levels of lysine from 0.62 to 0.78% continuously increased daily body weight gains and improved feed conversion efficiency as well as carcass characteristics. There was no significant interaction between the dietary lysine supply and the ratio between sulphur containing amino acids to lysine on animal performance parameters. This means that the effect of the ratio of sulphur containing amino acids to lysine was similar for various dietary lysine concentrations. The optimum ratio between sulphur containing amino acids to lysine according to quadratic regression analysis was 0.60, for both, growth and feed conversion. Reducing the ratio between sulphur containing amino acids to lysine from 0.59 to 0.53 and 0.47 reduced body weight by 3 and 12%, resp., and elevated the feed conversion ratio by 2 and 12%, resp. An increase of the ratio between sulphur containing amino acids to lysine from 0.59 to 0.65 failed to increase the animal performance. In contrast to animal performance parameters, optimum carcass characteristics (eye muscle area, fat area above eye muscle, meat-fat ratio and lean percentage) were achieved already at a ratio of sulphur containing amino acids to lysine of 0.53. PMID- 10849869 TI - Chromium yeast affects growth performance and plasma traits but not carcass characteristics of growing-finishing pigs depending on the glycemic index. AB - Forty Swiss Large White pigs (barrows with 31.7 kg initial to 103.7 kg final BW) were equally and randomly assigned to one of four treatments (H0, H200, L0, L200) involving a combination of chromium supplementation (0 or 200 micrograms/kg) and glycemic index (high GI (H) or low GI (L)). Growth performance, energy and protein digestibility, carcass composition, and some plasma traits were investigated. The data indicated, that the substitution of dietary carbohydrates with fat and crude fibre (low GI) resulted in lower growth performance due to impaired energy digestibility. Moreover, the strong stimulation of insulin secretion due to the high and rapid availability of carbohydrates of the diets H0 and H200 caused increased carcass fat deposition. Chromium supplementation also affected plasma insulin and glucagon concentrations. Depending on glycemic index, chromium affected the growth performance. Daily gain was reduced in pigs of the L200 treatment compared to the L0 group. This finding indicated that the energy availability expressed as GI is one of several nutritional factors, which determine the efficacy of dietary chromium. We could not corroborate evidences that dietary chromium modifies the chemical composition of the whole carcass, but depending on GI, chemical composition of the longissimus muscle was affected. PMID- 10849870 TI - Utilisation of iodine from different sources in pigs. AB - Balance experiments have demonstrated that growing pigs fed a ration consisting of wheat, barley, extracted soya meal, dicalciumphosphate, and iodine-free feeding salt utilised 48.8% of the received iodine. The tested supplementary iodine sources included potassium iodide (KI), ethylenediamine dihydroiodide (EDDI), iodine humate (HUI) prepared from iodine acid (HIO3), and the product P containing 0.004% iodine in an oil base (P). The amount of the supplemented iodine was in all cases 1 mg per 1 kg feed. The utilisation of iodine from the supplements reached 93.6, 92.6, 90.7, and 67.9% for KI, EDDI, P, and HUI, respectively. The values were significantly higher compared with controls (P < 0.01). Compared with KI and EDDI, the utilisation of iodine from HUI was significantly lower (P < 0.01). The lower availability of iodine from HUI was probably due to the high binding capacity of humate. The amount of urinary iodine excreted by control pigs receiving in the non-supplemented ration 147.5 micrograms iodine per day, was 40.3 micrograms per day (27.3%). In the pigs receiving in the supplemented ration 1647.5 micrograms iodine per day, the amount of urinary iodine reached 734.9 to 805.0 micrograms per day (44.6 to 48.9%). The corresponding values of faecal excretion were 75.6 micrograms iodine per day (51.2%) for the control pigs and 106.2 to 121.1 micrograms iodine per day (6.45 to 7.35%) for the pigs fed the supplemented rations. A high amount of 528.6 micrograms iodine per day (32.1%) was excreted in the faeces by pigs of the group HUI. PMID- 10849871 TI - The chemical composition of Phyllanthus discoideus and its effect on the ruminal ammonia and volatile fatty acid concentration when fed to west African dwarf sheep. AB - The feeding value of Phyllanthus discoideus (also called Margaritaria discoidea) leaves was evaluated using eight two-year-old West African Dwarf sheep fed natural grass hay. Four of the animals were fistulated ruminally and used for ammonia and volatile fatty acid determination in the fluid. Dried leaves of Phyllanthus discoideus were offered at two levels (25% and 50% of DMI, diets D25% and D50%, respectively) as supplements to the basal hay diet. The CP content of the control, D25% and D50% diets were 11.5, 12.6 and 13.6%, respectively, and their digestible energy amounted to 58.2, 61.1 and 56.9%, respectively. Rumen liquor was sampled one hour before and one, three and five hours after the morning feeding. Sheep fed the control diet had a higher ruminal ammonia concentration than those fed diet D25%. Similarly, ruminal ammonia concentration was higher in sheep fed the control diet than those fed the diet D50%. Five hours after feeding the ruminal ammonia concentration was significantly lower than one hour after feeding. The VFA concentrations in rumen fluid of sheep fed the control diet was inferior to those fed diets D25% and D50%. Sheep fed diet D50% showed significantly higher VFA concentrations than those fed diet D25%. Digestibility of organic matter and digestible energy did not show any significant difference. However, a marginal increase in organic matter digestibility of 3.5% was observed in diet D25% compared with the control diet. There was no significant difference in the N-digestibility in sheep fed the control, D25% and D50% diets. Nevertheless, a marginal improvement in N digestibility (1.5%) and N-retention (2.7%) was observed with the highest level of Phyllanthus discoideus (D50%). In conclusion, Phyllanthus discoideus appears as a particularly valuable feedstuff because it contains low levels of condensed tannins (12.8 g/kg), high CP content (156 g/kg) and a relatively high GE content (19.3 kJ/g DM). Although the improvement in N-digestibility and N-retention were only marginal the feeding of Phyllanthus discoideus could be justified under extreme shortage of feed resources during dry season. It should also be mentioned that a much more pronounced effect by supplementation with Phyllanthus discoideus could be expected when poor quality grass hay prevalent in West Africa during the dry season is fed. Phyllanthus discoideus could serve as a supplement to poor quality grass at 25% to 50% of supplementation. PMID- 10849872 TI - Application of noninvasive and invasive tests for risk assessment in patients with ventricular arrhythmias. AB - Sudden cardiac death remains a major public health problem in western society. Because most patients who experience cardiac arrest are not successfully resuscitated, primary prevention of sudden death remains an important challenge. A number of noninvasive risk stratification techniques have been suggested as providing useful information in patients with underlying structural heart defects. Unfortunately, the positive predictive value of most of these techniques has been limited. Left ventricular ejection fraction, the presence of nonsustained ventricular tachycardia on Holter monitoring, and inducible sustained ventricular tachycardia at electrophysiologic testing in patients with coronary artery disease remain the best established prognostic test. However, with the exception of two ICD studies using the combination of these markers, prospective studies have not yet completely validated the use of these and other prognostic markers. Further understanding of the pathophysiology of ventricular fibrillation and other risk stratification techniques will be necessary before a clear algorithm can be developed for application to patients at risk for sudden death. PMID- 10849873 TI - Ventricular arrhythmias in normal hearts. AB - Idiopathic ventricular tachycardia (VT) is characterized by two predominant forms. The most common form originates from the right ventricular outflow tract and presents as repetitive monomorphic VT or exercise-induced VT. The tachycardia is adenosine sensitive and is thought to be because of cAMP-mediated triggered activity. The other major form of idiopathic VT is owing to verapamil-sensitive intrafascicular re-entrant tachycardia, which most often originates in the region of the left posterior fascicle. Both forms of idiopathic VT can be readily treated with radiofrequency catheter ablation. PMID- 10849874 TI - Management of postinfarct ventricular tachycardias. AB - The clinical profile of patients with postinfarct VT has changed in the past two decades. Along with these changes, existing treatments have improved, and entirely new therapeutic approaches have been developed. The expanded range of treatment options has made postinfarct VT a less imposing clinical problem than it once was. Emerging therapies promise to make an even greater beneficial impact. The challenge in treating patients with postinfarct VT has changed from merely keeping patients alive to keeping up with innovations in therapy that can provide them with a better quantity and quality of life. PMID- 10849875 TI - Long QT syndrome. AB - In conclusion, much has been learned in the past several years regarding the molecular biology of LQTS, and this information has been directly applicable to the clinical care of patients with this syndrome. The knowledge also has been of considerable importance for understanding the molecular basis of arrhythmias in general and is providing insights into potential molecular-based therapies for arrhythmias. PMID- 10849876 TI - Nonsustained ventricular tachycardia. AB - The patient with nonsustained ventricular tachycardia represents a common management problem for the cardiologist. The challenges posed by this type of arrhythmia differs from those posed by other arrhythmias, because most instances of nonsustained ventricular tachycardia do not cause symptoms. This article reviews common situations in which nonsustained ventricular tachycardia occurs and their appropriate management. PMID- 10849877 TI - Large clinical trials in the management of ventricular arrhythmias. Applying group results to individual cases. AB - Applying the results of clinical trials to everyday practice in an appropriate manner can be difficult. Earlier clinical trials focused on the secondary prevention of ventricular arrhythmias. Studying patients at high risk of recurrent ventricular arrhythmias is important, but the overall impact on arrhythmic death is low. Recent arrhythmia trials have studied specific patient populations for the primary prevention of ventricular arrhythmias. New trials will expand the investigation to other populations. This article summarizes the results of large clinical trials in the management of ventricular arrhythmias and attempts to provide guidelines for applying their results to everyday clinical practice. PMID- 10849878 TI - Drug therapy of sustained ventricular tachyarrhythmias. Is there still a role? AB - This article provides a review of the risks faced by patients with sustained ventricular tachycardia (VT) or ventricular fibrillation (VF) in the absence of a reversible or transient cause so that the goals of therapy can be clearly defined. The therapeutic approaches that have been proposed to achieve these goals are outlined and evidence comparing these various approaches to therapy is then summarized in order to propose an algorithm for the optimal use of antiarrhythmic drug therapies as primary therapy for selected VT/VF patients. Options for the ancillary uses of antiarrhythmic drug therapies in ICD patients are considered. PMID- 10849879 TI - ICD therapy in the new millennium. AB - Remarkable progress has been made in the 15 years since ICD therapy was approved for human use. The early "shock boxes" had almost no diagnostic capabilities and required thoracotomy for epicardial patch implantation with typical duration of hospitalization of about a week. Pulse-generator longevity was less than 2 years. Modern devices provide detailed information about the morphology and rate of electrocardiographic signals before, during, and after arrhythmia therapy. The down-sizing of pulse generators and improvements in lead design and shock waveforms allow the simplicity of defibrillator implantation to approach that of pacemakers, with defibrillation thresholds comparable with those initially observed with epicardial patches. Despite the marked reduction in size and increase in diagnostic capabilities, device longevity is now longer than 6 years. Routine outpatient ICD implantation is presently feasible and will increase in frequency if ongoing primary prevention trials prove beneficial. Further advances in lead technology and arrhythmia discrimination should increase the efficacy and reliability of therapy. Finally, devices have the capabilities to treat multiple problems in addition to life-threatening ventricular arrhythmias including atrial arrhythmias and congestive heart failure. PMID- 10849880 TI - Hybrid therapy for ventricular arrhythmia management. AB - Optimum arrhythmia management has evolved to couple ICD therapy with catheter ablative and drug therapy to attempt to eliminate or reduce arrhythmia risk. No longer should the clinician approach such therapy as a choice among single alternative strategies only. Optimum patient management includes not only recognition of the indications and benefits of such hybrid therapy but also a complete understanding of potential pitfalls of such therapy. PMID- 10849881 TI - Measurement and cardiology--increasing the usefulness of measurement in the future. PMID- 10849882 TI - Good design practice for medical devices and equipment, Part I: A review of current literature. AB - Changes to the medical device regulations within the past few years have forced medical device manufacturers to take an integrated approach to design and validation in order to ensure that their products are reliable and fit for purpose. Good design practice encourages fitness for purpose within commercial reality. This paper contains a review of the current literature that is relevant to good design practice. The results show that there is inadequate guidance regarding the integration of validation with design. Thus, there is a need for good design practice to include 'design for validation' which is aimed at designing medical devices to make them easier and more economic to validate. Research has been carried out at the Cambridge Engineering Design Centre in order to develop an approach that provides guidance in order to help designers achieve integrated design, development and validation. This will be reported in part II of this paper. PMID- 10849883 TI - Use of telekinesthetic feedback in walking assisted by functional electrical stimulation. AB - A telekinesthetic feedback implemented into functional electrical stimulation (FES) orthosis is described. Single channel FES is used to provoke ankle dorsiflexion during walking. FES is controlled manually by a special lever, built into the handle of the crutch. The angular position of the lever defines the intensity of stimulation and thus the magnitude of the ankle dorsiflexion. The measured joint angle provides the feedback information about the ankle joint position, which is presented to the user as a force feedback applied to the control lever. As the first step in the development of a complex micromechatronic device, a simulated testing environment was prepared. A computer model, comprising dynamic foot characteristics, as well as agonistic and antagonistic muscle groups, substitutes the ankle joint. The model also includes fatiguing of the electrically stimulated muscles. For experimental purposes an actuated control lever was built. The efficacy of the telekinesthetic feedback was evaluated in a group of six healthy persons. PMID- 10849884 TI - A quantitative study of the pixel-shifting, blurring and nonlinear distortions in MRI images caused by the presence of metal implants. AB - Magnetic resonance imaging has found an increasing number of medical applications in recent years due to its technical merits as well as its non-invasive nature. However, its full potential has been severely limited by magnetic susceptibility difference artefacts caused by the presence of ferromagnetic sources such as orthopedic implants, dental work or metallic needles used in neurosurgery. In this study, we propose a method to numerically quantify the distortions resulting from the magnetic susceptibility differences by investigating the phenomena from three perspectives: (1) pixel displacement, (2) blurring and (3) nonlinearity. For this purpose, phantom images obtained from a magnetic resonance scanner were studied. Attempts made to reconstruct an ideal image from its distorted version by appropriately compensating for the three types of distortions yielded encouraging results. PMID- 10849885 TI - Reduction of electromagnetic interference from a commercially available MR compatible flow simulator. AB - Electromagnetic interference (EMI) associated with the electronics of a commercially available computer controlled flow simulator substantially decreases the quality of the MR image. The effect of a custom-built radiofrequency shield on its spectral emission, and the corresponding signal-to-noise ratio measured for the image of a standard phantom, were determined. The results demonstrate the elimination of EMI and a significant improvement in image quality. PMID- 10849886 TI - Hospital bed mattresses: an overview of technical aspects. AB - Mattresses designed for clinical applications are deceptively simple pieces of equipment. Their primary function is to comfortably support the patient in an appropriate position. However, there are many other technical factors that affect the quality of the support surfaces. In this paper we discuss the basic functions that mattresses should fulfil, explain some of the underlying mechanics and discuss CE marking and other standards. PMID- 10849887 TI - Humidity measurements in passive heat and moisture exchangers applications: a critical issue. AB - A reliable, quantitative assessment of humidification performances of passive heat and moisture exchangers in mechanically-ventilated patients is still to be achieved, although relevant efforts have been made to date. One of the major problems to tackle consists in the difficulty of humidity measurements, both in vivo (during either anaesthesia or intensive care unit treatments) and in vitro set-ups. In this paper a review of the basic operation principles of humidity sensors as well as an analysis of their usage within in vivo and in vitro tests are presented. Particular attention is devoted to the limitations arising from the specific measurement set-up, as they may affect the results notably. PMID- 10849888 TI - Enhancement of delayed afterdepolarizations and triggered activity by class III antiarrhythmic drugs: multiple effects of E-4031 and dofetilide. AB - The effects of class III antiarrhythmic agents E-4031 and dofetilide were studied on action potentials and subthreshold delayed afterdepolarizations (DADs) induced by the cardiac glycoside acetylstrophanthidin (AS) in isolated cardiac Purkinje fibers. Action potentials were recorded from cardiac Purkinje fibers using microelectrode techniques. E-4031 and dofetilide consistently increased DAD amplitude, occasionally caused triggered action potentials and shortened action potential duration. The application of E-4031 without prior AS exposure, resulted in the typical class III antiarrhythmic effects of action potential lengthening and the induction of early afterdepolarizations. These findings suggest that under our conditions of AS-induced cell Ca2+ overload, the effects of the "pure" class III antiarrhythmic drugs, E-4031 and dofetilide, are markedly different from those found in non-Ca2+ loaded cells. This may represent an additional electrophysiological mechanism for class III antiarrhythmic drug toxicity. PMID- 10849889 TI - Pharmacokinetic characterization of the indole alkaloid ibogaine in rats. AB - To investigate the pharmacokinetic properties of ibogaine, a putatively anti addictive alkaloid, levels of this drug were quantified in plasma and tissues for up to 3 h following i.v. infusion in rats. Immediately following a 31-35 min infusion (20 mg/kg), mean plasma ibogaine levels were 373 ng/ml; these values declined rapidly thereafter in a biexponential manner. The plasma time course in 5 of 7 animals demonstrated an excellent fit to a two-compartment pharmacokinetic model, with alpha and beta half-lives of 7.3 min and 3.3 h, respectively. Drug clearance was estimated to be 5.9 l/h (n = 7). Ibogaine levels in brain, liver and kidney 3 h after the end of drug infusion were 143-170 ng/g, close to simulated values for the peripheral pharmacokinetic compartment. However, 3-h drug levels in adipose tissue were much higher (3,328 ng/g), implying the need for a more complex pharmacokinetic model. Mechanisms for the initial, rapid disappearance of plasma ibogaine are thought to include metabolic demethylation as well as redistribution to body stores. The sequestration of ibogaine by adipose tissue probably contributes to a protracted persistence of drug in the body. This persistence may be underestimated by the beta half-life reported in the present study. PMID- 10849890 TI - Liver sulphoxidative metabolism of albendazole in rat: enantioselectivity and effect of methimazole. AB - The aim of the present work was to evaluate the effects of methimazole (MTZ) on the enantioselective sulphoxidation of albendazole (ABZ) by rat liver microsomes and tissue slices. Albendazole sulphoxide (ABZSO) was the metabolite recovered after the incubation with ABZ in both liver preparations. MTZ significantly reduced ABZSO production both in microsomes and slices. ABZSO production decreased as a function of MTZ concentration. The sulphoxidation reaction performed by rat liver explants in the presence of MTZ was 65% lower than that observed in controls. The reduction in the production of ABZSO in the presence of MTZ was mainly due to a lower production of (+) ABZSO. The results reported further contribute to the understanding of the enantioselective metabolism of ABZ. In addition, the work presented provides information on the comparison of two different liver tissue preparations for the evaluation of xenobiotic metabolism. PMID- 10849891 TI - Donepezil, a centrally acting acetylcholinesterase inhibitor, alleviates learning deficits in hypocholinergic models in rats. AB - Donepezil is a member of a new class of centrally acting cholinesterase inhibitors which preferentially inhibit acetylcholinesterase rather than butyrylcholinesterase. The effects of donepezil on learning impairments were investigated in some hypocholinergic models in rats. In nucleus basalis magnocellularis (NBM)-lesioned rats, donepezil alleviated deficits in passive avoidance response at a dose of 0.125 mg/kg and higher, while tacrine had only a tendency toward improved performance. Donepezil at 0.5 mg/kg effectively counteracted acquisition impairments in the water maze task induced by lesions of the medial septum; tacrine had no significant effects on impairments in this task. Scopolamine caused an increase of errors in the 8-arm radial maze. Donepezil significantly decreased scopolamine-induced errors in the radial maze at 0.5 mg/kg, whereas tacrine decreased errors at 2 mg/kg. These results suggest that donepezil can clearly minimize learning impairments induced by treatments that cause central cholinergic deficiencies in rats. These findings support the clinical efficacy of donepezil in Alzheimer's disease. PMID- 10849892 TI - Effects of intravenous theophylline, aminophylline and ethylenediamine on antigen induced bronchoconstriction in guinea pigs. AB - The antiasthmatic effect of i.v. injection of theophylline was compared with that of aminophylline by using an antigen-induced bronchoconstriction model in sensitized guinea pigs. Both theophylline and aminophylline showed dose-dependent inhibition of antigen-induced bronchoconstriction. Statistically significant differences were observed at theophylline doses of 20 and 40 mg/kg and at aminophylline doses of 25 and 50 mg/kg. Thus, antiasthmatic effects of both drugs appeared to be similar. In addition, intravenous ethylenediamine did not influence either airflow in normal guinea pigs or bronchoconstriction induced by antigen at doses up to 30 mg/kg. In conclusion, the ethylenediamine in aminophylline may not influence the antiasthmatic action of theophylline, and the therapeutic effects of theophylline and aminophylline are suggested to be similar. PMID- 10849893 TI - Teratogenic and fetal toxicity following intravenous theophylline administration in pregnant rabbits is related to maternal drug plasma levels. AB - This study investigated the teratogenic and fetal toxicity of i.v. theophylline and its relationship to maternal plasma levels in pregnant rabbits. From days 6 18 of gestation, each dose of theophylline (15, 30 and 60 mg/kg/day at a rate of 0.5 ml/kg/min) was administered i.v. to pregnant rabbits using an automatic infusion pump. Theophylline showed reversible toxicity: accelerated respiration, sluggish startle reactions, dilation of the auricular vessels and polyuria were observed in dams treated with 60 mg/kg/day but not in animals given 15 or 30 mg/kg/day. Fetuses from the dam group treated with 60 mg/kg/day exhibited teratogenic toxicity such as cleft palate and skeletal variation of the 13th rib. Fetal toxicity was also observed including abortion, increased number of late deaths and decreased body weight appearing on day 29 of gestation. No toxicity was observed in fetuses from the dam group treated with 15 or 30 mg/kg/day. However, in the 30 and 60 mg/kg/day theophylline-treated groups, maternal plasma concentrations (Cmax) during the treatment period were approximately 56 and 106 micrograms/ml, respectively. It is therefore suggested that the risk of teratogenic and fetal toxicity caused by theophylline is dependent on its dosage. In conclusion, caution should be taken when administering theophylline or aminophylline to pregnant individuals at doses that could result in high neonate peak blood levels. PMID- 10849894 TI - Influence of Ramadan on the pharmacokinetics of a single oral dose of valproic acid administered at two different times. AB - This study was carried out in healthy volunteers in order to examine the influence of changes in eating and rest/activity rhythms during Ramadan on the pharmacokinetics of valproic acid (VPA; Depakine). A single oral dose of 800 mg was administered to the first group of subjects (n = 7) at 8:00 PM and to the second group (n = 5) at 5:00 AM. Each group was submitted to three treatment phases: the first was carried out 3 weeks prior to Ramadan (PR), the second one at the end of the first week of Ramadan (R1) and the last at the end of the third week of Ramadan (R3). The plasma kinetics of VPA were determined for each treatment schedule throughout the 50 h following drug intake. During Ramadan, a significant decrease was observed in the Cmax (56.22 +/- 5.32 mg/l in PR vs. 48.35 +/- 5.07 mg/l in R3; p < 0.05) and in the AUC(0.50 h) (1429.92 +/- 284.23 in PR vs. 1090.26 +/- 277.73 mg.h/l in R3; p < 0.05) for the 8:00 PM intake. For the 5:00 AM intake, a significant decrease was observed in the t1/2 (12.15 +/- 1.45 h in PR vs. 9.55 +/- 1.97 h in R3; p < 0.05) and AUC(0.50 h) values (1241.29 +/- 239.01 mg.h/l in PR vs. 1019.21 +/- 256.86 mg.h/l in R3; p < 0.05). These parameters showed a significant decrease at the end of the third week of Ramadan (R3), compared to the control period (PR). PMID- 10849895 TI - Pharmaco-EEG test dose response predicts cholinesterase inhibitor treatment outcome in Alzheimer's disease. AB - Previous investigations have indicated that a single dose pharmaco-EEG may predict the outcome of 4-7 weeks of tetrahydroaminoacridine (THA) treatment in dementia of the Alzheimer type (DAT). This open trial study further examined the relationship of quantitative EEG in relation to treatment response by assessing 24 probable DAT patients at baseline, 2 h after their first oral dose (30 mg), and after 12 weeks of THA treatment. Compared to EEG norms, patients, in general, evidenced EEG slowing, as shown by excessive slow (theta) and diminished fast (alpha and beta) wave power as well as reduced mean frequencies which were present prior to treatment as well as at the end of treatment. The EEG of patients exhibiting stable or improved scores on the Mini-Mental State examination (MMSE) at 12 weeks showed a significantly faster baseline mean alpha frequency as well as a significant reduction in relative theta power following the single THA test dose compared to deteriorated patients. A discriminant analysis using test dose response EEG variables correctly classified 75-79% of these two patient groups, suggesting that this procedure may be a useful approach for optimizing patient selection for antidementia treatments. PMID- 10849896 TI - Discovery of new antimetastatic agents: review of in vitro and in vivo screening methods. PMID- 10849897 TI - The pilosebaceous unit: a pivotal route for topical drug delivery. AB - The hair follicle, hair shaft and sebaceous gland are collectively known as the pilosebaceous unit. The pilosebaceous unit is a complex, dynamic, 3-D structure, which is the site of unique biochemical, metabolical and immunological events. Ongoing research has focused on the hair follicle as a potential route for both localized and systemic drug delivery. Targeted drug delivery to the specific sites of hair follicle has paved ways to treat several dermatological abnormalities that are known to originate at the hair follicle. In recent studies, topical liposomes have been shown to target the drug to the pilosebaceous unit. This review describes general aspects of pilosebaceous unit, follicular delivery, with particular emphasis on studies which have demonstrated targeted follicular delivery via liposomes. PMID- 10849898 TI - Application of an almost traceless linker in the synthesis of 2 alkylthiobenzimidazole combinatorial libraries. AB - Resin-bound triphenylphosphine was coupled to 4-fluoro-3-nitrobenzyl bromide, and 2-alkylthiobenzimidazoles were synthesized on resin in 4 steps using standard chemistries. Cleavage of the compounds from the resin was achieved with 10% NaOH in MeOH to leave a methyl group at the attachment point. A total of 47 amines and 40 electrophiles were evaluated, defining the scope of the reactions, culminating in the synthesis of an 80-member test library of high purity as determined by HPLC. PMID- 10849899 TI - Synthesis and diversity analysis of lead discovery piperazine-2-carboxamide libraries. AB - A Lead Discovery Library of piperazine-2-carboxamide derivatives was produced for general screening. This paper discloses two novel solid phase synthetic routes used to produce 15,000 single compounds via the Irori directed sorting technique. Computational methods such as reagent clustering and library profiling were used to maximize reagent diversity and optimize pharmacokinetic parameters. The results of a four center pharmacophore analysis revealed the added diversity gained by using two independent synthetic routes. PMID- 10849900 TI - Comprehensive survey of combinatorial libraries with undisclosed biological activity: 1992-1997. PMID- 10849901 TI - [Practical significance of ischemic stroke OCSP (Oxfordshire Community Stroke Project) classification]. AB - OCSP classification based on neurological signs and syndromes contains four subtypes of ischaemic stroke: lacunar infarct (LACI), total anterior circulation infarcts (TACI), partial anterior circulation infarcts (PACI), posterior circulation infarcts (POCI). Literature reports suggest that this classification may be useful in estimation of after stroke complications and prognostication, and can raise the sensitivity of therapeutic clinical tests. The aim of this study was to estimate the occurrence of risk factors, accompanying complications and prognosis in a material of clinical records of 346 hospitalized patients. Attention is drawn to limited importance of brain CT as shown in frequent discordance between clinical syndromes and CT findings in establishing to which stroke subtype a given patient belongs. The results obtained in the study are highly similar to those reported from other clinical centres. It was found that PACI syndrome was most frequently occurring, and that TACI subtype was associated with the highest frequency of complications and risk factors, and worst prognosis. The prognosis was best in the LACI subtype. The subtype of OCSP classification seems to be determining the possibility of complications and prognosis, and could suggest the most effective medical treatment. PMID- 10849902 TI - [Morphology of demyelination plaques vs cognitive and emotional disorders in multiple sclerosis patients]. AB - The interrelationship between psychological examination and MRI findings was studied in 70 patients with MS. The cognitive and emotional functions were examined by a battery of tests: Wechsler Adult Intelligence Scale, Visual Retention Test, Hamilton Depression Scale. In MRI examination the localization, area, and the morphology of the plaques were examined. According to plaque's morphology the patients were divided into two groups: with confluent plaques and those with patchy-shaped ones. The signs of dementia were found significantly more frequently in the group with confluent plaques (p. < 0.04). In this group of patients also single-function disorders like disturbances of verbal memory, attention, visual memory, cause- and effect thinking, abstract thinking, and visual-motor coordination were significantly more frequent (p. < 0.01). In the same group the signs of fatigue syndrome were more frequently encountered (p. < 0.02). The authors conclude that the disturbances found in cognitive function may reflect the symptoms of subcortical dementia in MS patients. PMID- 10849903 TI - [Lamotrigine in add-on therapy: assessment of efficacy in drug resistant epilepsy]. AB - The authors presented the results of treatment with lamotrigine (LTG, Lamictal) in 13 patients with drug resistant epilepsy (add-on therapy). There were 8f, 5m. aged 16-60 years, mean age 28.8 years. Generalized seizures occurred in 8 patients (62%). In this group there was 1 patient (aged 16 years) with the Lennox Gastaut syndrome and 1 patient (aged 20 years) with valproate resistant juvenile myoclonic epilepsy. Complex partial seizures and complex partial with secondary generalization occurred in 5 patients (38%). Before LTG addition mean seizure frequency was from 3/month to several times/day. The mean duration of epilepsy was 16.6 years. The 8 patients were treated with CBZ and VPA, one with PHT and VPA, one CBZ and VGB. Monotherapy with VPA was introduced in 3 patients. After 6 months of treatment with LTG the efficacy was evaluated. 12 patients took LTG with VPA, 1 LTG with CBZ. Complete reduction of seizures was achieved in 3 cases (23%), at least 50% reduction in 3 patients (23%), reduction below 50% in 4 patients (31%). In 3 cases (23%) the results of treatment were negative (increase or no change in seizure frequency). Beneficial psychotropic effect was observed in 9 patients (69%). Adverse effects occurred in 2 patients (15%). Headache, vertigo, sleepness were observed in one case. Rash occurred in 1 patient (treated with LTG and VPA). After 6 months 3 patients were excluded from the study because of negative effects of treatment. LTG is helpful and well tolerated in drug resistant epilepsy. PMID- 10849904 TI - [Rehabilitation after stroke in patients with diabetes mellitus]. AB - Diabetes mellitus is not only one of the fundamental risk factors for stroke, but it may also constitute a significant impediment and limitation of rehabilitation process in diabetic patients after stroke. Insufficiently controlled diabetes or insufficient insulin therapy inadequately carried out, especially during the period of acute cerebrovascular complications, may increase the risk of repeated stroke or increase the size of ischaemic area. In patients suffering from diabetes the course of stroke is more severe, cerebral oedema appears more often and mortality is higher. In the present paper methods of rehabilitation and results achieved in a group of patients with stroke and a group of patients with stroke and diabetes mellitus are described, as well as effects of rehabilitation in both those groups. Fifty one (20%), out of 251 patients with cerebral stroke in the past who were rehabilitated in a Rehabilitation Department in 1997, had diabetes mellitus lasting, on average, 21 years. Individual rehabilitation programme for each patient was arranged collectively, taking into consideration restrictions necessary in the course of diabetes, including education of patients. Multidirectional assessment of initial condition and condition after termination of the rehabilitation was carried out taking into consideration the scale of lesions (the Brunnstrom stages), functional scales and the quality of life index (the Barthel index). More objective parameters, i.e. the index of symmetry of loading of lower extremities and walking speed, were also applied. All the patients with both stroke and diabetes mellitus completed the planned rehabilitation programme, although the initial condition of patients with concomitant diabetes was worse than that of patients with stroke only, as far as most of the applied parameters of assessment were concerned. Also improvement achieved in diabetic patients was not observed in each of the parameters examined and differed from that achieved in the group of patients with stroke alone. We found, that in-hospital rehabilitation of patients with stroke and concomitant diabetes mellitus (even long lasting) resulted in a measurable and significant improvement of their condition evaluated multidirectionally, and that diabetes mellitus coexisting with stroke is not a contraindication to in-hospital rehabilitation. In patients with concomitant diabetes mellitus significant disturbances of during the rehabilitation were not found course diabetes mellitus. However they all required meticulous care and monitoring of carbohydrate metabolism, and the average time of their hospitalisation was longer by about 3 days. PMID- 10849905 TI - [Botulinum toxin in the treatment of focal dystonias: own experience with botulinum toxin]. AB - During the last 5 years, 75 patients with focal dystonias were longitudinally treated with injections of botulinum toxin A. Each patient received 2-6 injections. The improvement was assessed after each injection and estimated as significant after 68.47% of injections, as mediocre after 23.42% of injections and none after 8.11% of the injections. The most significant improvement was obtained in patients with blepharospasm and hemifacial spasm, the worst effect was obtained in spasmodic torticollis. Varying responses were observed following repeated injections of botulinum toxin, the clinically assessed improvement did not decrease after successively applied doses. Side-effects occurred after 18% of the injections and were mostly mild and disappeared after short time. This study confirms the usefulness of botulinum toxin, which is an effective and safe treatment of focal dystonias. PMID- 10849906 TI - [Set of neuropsychological tests for the assessment of language disorders after damage to the right cerebral hemisphere: preliminary results]. AB - A set of five experimental tests was prepared for the assessment of language disturbances in right hemisphere-damaged (RHD) patients. The Right Hemisphere Language Battery by Bryan (1995) was a basis for these tests. The set comprises the Metaphor Picture Test, the Written Metaphor Test, the Inferential Meaning Test, the Humour Test and the Lexical Semantic Test. Thirty-six RHD, 15 left hemisphere damaged (LHD) without aphasia and 30 control (C) subjects took part in the investigation. All test significantly differed between the RHD and C groups. No significant differences were noted between LHD and control groups, except for the Inferential Meaning Test. The quantitative performance of tests was different in all the groups. Results of the study indicated patients with right frontal damages to make more errors than those with right posterior lesions. PMID- 10849907 TI - [Correlation of steroid receptors and growth factors expression with peritumoral edema accompanying cerebral neoplasms]. AB - The aetiology of peritumoral brain oedema still remains unclear, despite investigations, attempted to study the role of various clinical factors and histological features of the tumour. The aim of this investigation was to the evaluate correlation of peritumoral brain oedema with expression of progesterone (PR) and oestrogen (ER) receptors and vascular--endothelial growth factor (VEGF). Eighty nine samples of brain tumours were examined, among them 28 meningiomas, 43 gliomas and 18 metastatic tumours. Expression of steroid receptors was examined with the radioreceptor assay method and expression of VEGF by radioimmune assay. A statistically highly significant correlation was found of VEGF expression with the incidence of diffuse pattern of brain oedema around gliomas and metastatic tumours (p < 0.01). PR expression was associated significantly with the occurrence of localized type of oedema around meningiomas, but in gliomas the expression of this receptor correlated with a diffuse pattern of oedema. No correlation was found between ER expression and peritumoral brain oedema. PMID- 10849908 TI - [Infusion test in the selection of patients with hydrocephalus for the shunting]. AB - The indications for the shunting of the patients with NPH are a matter of discussion. The measurement of the outflow resistance during the infusion test is one of the criteria for shunting. The authors present 38 patients with NPH in whom the lumbar constant-infusion test was carried out. The patients with the outflow resistance more then 13 mmHg/ml/min were selected for shunting. The results at least 4 months after operation are presented. PMID- 10849909 TI - [Posttraumatic vasospasm and its treatment with nimodipine]. AB - The aim of our study was to compare the incidence and the time course of vasospasm as well as outcome of patients with tSAH. In group I there were 15 patients treated with nimodipine. Group II consisted of 20 patients in whom nimodipine was not used. All patients suffered from severe head injury, scored 8 points or less according to Glasgow Coma Scale. The initial CT scan revealed tSAH in all of them. Outcome was evaluated according to Glasgow Outcome Scale (GOS) three months after injury. Results were better (but not significantly) in patients in whom nimodipine was used, time course of vasospasm was less severe and follow-up CT scans did not reveal ischaemic infarcts connected with vasospasm. PMID- 10849910 TI - [Cortico-basal degeneration]. AB - Cortico-basal degeneration (CBD) or cortico-basal ganglionic degeneration is a condition characterised by selective cortical atrophy of parietal and in a lesser extent, frontal lobe associated with dysfunction of the basal ganglia. The clinical symptoms of CBD, predominantly extrapyramidal signs (bradykinesia and rigidity) and apraxia, affect often only one body side in the onset phase, with the left one being more frequent. Neuropathological studies reveal neuronal loss, gliosis, and achromasia chiefly in frontal and parietal cortex, as well as in basal ganglia and substantia nigra. Functional investigations, such as SPECT, disclose similar distribution of abnormalities (hypometabolism). The aetiology and causative treatment of CBD are unknown. The authors highlight the diagnostic difficulties in CBD including a necessity of a prolonged patient's observation in order to ascertain the differential diagnosis of other neurodegenerative disorders, in particular progressive supranuclear palsy, Alzheimer's disease and Parkinson's disease. PMID- 10849911 TI - [The role of gamma-aminobutyric acid in the mechanism of action of anticonvulsant drugs]. AB - Decreased activity of gamma-aminobutyric acid, the major inhibitory neurotransmitter in CNS can be epileptogenic. Manipulation of the GABA system has been a target for development of antiepileptic drugs. The different ways for augmenting gabaergic inhibition by conventional and new AEDs are presented in this paper. Among the I generation, barbiturates and benzodiazepines are potent anticonvulsants that act as GABA modulators in postsynaptic GABA-A receptor complex but their usefulness is limited by dependence and tolerance to antiseizure activity. The II generation drugs vigabatrin and tiagabine, and to some extent gabapentin have been developed by a rationale strategy and none of them exert direct action in GABA receptors. Only two former drugs exhibit selective, strictly defined activity: vigabatrine is an irreversible inhibitor of GABA-aminotransferase and tiagabine acts as a GABA-uptake inhibitor from synaptic cleft into neurons and glia. Gabapentin binds to a novel receptors in epileptogenic areas in CNS and enhances GABA turnover. Drugs with multiple mechanisms of action, felbamate and topiramate not only potentiate gabaergic inhibition in several ways but also diminish the activity of excitatory amino acids at their NMDA or AMPA receptors; the later mechanism seems to be essential for their potential neuroprotective activity in epileptogenesis. None of gabamimetic drugs provide optimal seizure control but better tolerability of newer ones and well-established mechanisms of action provide possible harmless therapy. PMID- 10849912 TI - [The role of stereotactic biopsy in diagnosis of pineal region tumors]. AB - The anatomy, pathophysiology nad treatment of the pineal region tumors are presented. The authors stress the role of stereotactic biopsy in the diagnosis and correct management. Based on the literature and their experiences the risk of complications and incorrect diagnosis is discussed. PMID- 10849913 TI - [Transient patency loss of the internal carotid artery in a young patient]. AB - Ischaemic stroke has been observed more rarely among young people in comparison to old adults. The causes of ischaemic stroke among young adults are numerous. One of them is occlusion of internal carotid artery (ICA). The aetiology of the occlusion is different from arteriosclerotic disorders. Within the last few years the authors have paid much attention to the fact that spontaneous dissection is not such a rare cause of ICA occlusion. In many cases (60-70%) of dissection recanalization and benign outcome occur. Angiography has been considered as a gold standard in establishing the diagnosis. We described a young, 16-year-old patient with ischaemic stroke. The cause occlusion of ICA, probably was dissection. The diagnosis was made by angiography MR. In our patient follow-up angiography MR demonstrated recanalisation. The outcome of the stroke was benign and hemiplegia reversed. PMID- 10849914 TI - [Familial form of Fahr syndrome (report of two cases)]. AB - Two cases of familial form (in brothers) of extensive, symmetric, idiopathic calcifications of the basal ganglia and cerebellum (Fahr syndrome) are described. In discussion of factors predisposing to appearance of such idiopathic calcifications, particular attention was given to meningoencephalitis suffered in childhood and coexistence of a tumour of the sella of prolactinoma type, in case 1. Attention was also called to progressive, copious clinical and radiological symptology of the discussed cases. It was impossible to suggest a probable type of inheritance (parents of the patients were dead). PMID- 10849915 TI - [A case of complex development anomalies and aneurysm of the anterior part of arterial circles of the brain]. AB - A case of complex developmental anomalies and an aneurysm of the anterior part of arterial circle of the brain is reported. Every developmental anomaly was analysed separately. Views on their embryologic conditions are quoted. Haemodynamic disturbances are pointed out as a reason of the occurrence of the aneurysm. PMID- 10849916 TI - [The case of severe craniocervical dislocation due to rheumatoid arthritis]. AB - The authors describe a case of craniocervical dislocation secondary to rheumatoid arthritis producing important canal narrowing: ventrally by migrated odontoid and dorsally by posterior arch of C-1 with medullary compression. Symptoms of hyperreflexia, spasticity and left hemiparesis with Babinski sign were present. SURGICAL PROCEDURE: transoral odontoidectomy was performed followed by suboccipital approach, C-1 laminectomy and occipitocervical fixation (Olerud device and bone graft). Outcome with neurologic improvement. CONCLUSIONS: Transoral odontoidectomy combined with occipitocervical decompression and fixation is effective approach for treatment of severe craniocerebral dislocation. Its advantages: ventral and dorsal decompression combined with immediate stabilisation. PMID- 10849917 TI - [Letter concerning the paper of Z. Mariak published in Polish Neurology Neurosurgery 1999, vol. 33, No. 3, p. 665 [letter]. PMID- 10849918 TI - [Report on the satellite symposium "Treatment of multiple sclerosis in the year 2000: achievement and expectation at the turn of the millennium". Basle, Switzerland, 09.18.99]. PMID- 10849919 TI - Drug treatment of hypertension in older people: relieving the pressure. AB - Hypertension is very common, occurring in over 50% of older people, and is a major risk factor for stroke and ischaemic heart disease. Based on systematic reviews there is evidence to show that drug treatment of hypertension in older people saves lives and prevents unnecessary morbidity. There is also strong evidence to support the use of diuretics as first line agents. Quality of life does not appear to be reduced by antihypertensive drug therapy, although more high quality research is needed. Through the use of drug treatment older people with hypertension can continue to contribute to society and live active lives. PMID- 10849920 TI - The benefits of a hospital based community services liaison pharmacist. AB - OBJECTIVE: To investigate the benefits of a community services liaison pharmacist in addressing medication misuse in elderly patients, which occur on both admission and discharge. DESIGN: Completion of a medication history for each patient on admission by the community liaison pharmacist. On discharge updated medication record sheets were faxed to the patient's GP and community pharmacy; a survey of GPs' and community pharmacists' opinions who were involved in the study was carried out. SUBJECTS AND SETTING: 109 patients over the age of 60 on 4 or more medications admitted by the medical admissions unit of Antrim Area Hospital. MAIN OUTCOME MEASURES: Medication related problems; GP and community pharmacist opinions of the service. RESULTS: Of the 109 patients, 61% had an incomplete medication history on admission, 21% of patients who brought their own drugs were not dealt with appropriately in hospital and 33% of discharged patients had medication-related problems. The service was felt to be very useful by GPs (80%) and community pharmacists (100%). A reduction in readmission rate of 2.4% was seen in these patients compared to the average for this age group. CONCLUSION: The community services liaison pharmacist produced benefits in terms of patient medication management, reduced readmission rates and wastage of patients' own drugs. A more detailed one-year study will now be carried out. PMID- 10849921 TI - Influencing NSAID prescribing in primary care using different feedback strategies. AB - The Audit Commission endorsed the role of the prescribing adviser in promoting safe, rational and cost-effective prescribing by general medical practitioners (GPs). However, whether such roles should involve practice visits, facilitation of educational meetings, production of local bulletins, or a combination of these and other approaches is unclear. Few UK studies have investigated the best methods to influence prescribing on a large scale in primary care. The present study was designed to determine the effectiveness of active compared to passive practice specific prescribing feedback. The programme focused on non-steroidal anti-inflammatory drugs (NSAIDs) since concern has been expressed about their use, which accounts for 4% of volume and 5% of the cost of UK National Health Service drugs prescribed in primary care, Sixty-six of the 91 general medical practices contracted to Gwent Health Authority agreed to participate in the study and were randomly stratified by practice size, locality, fund-holding and dispensing status into 2 groups. Group 1 received active feedback via practice visits from the pharmaceutical prescribing adviser to present prescribing analysis and cost data (PACT) concerning NSAID use. Group 2 received passive feedback, a practice specific prescribing analysis workbook that contained similar information to that given to Group 1 practices. Practices not wishing to enter the study were used as a self selected reference group (Group 3) which received no information on NSAIDs from the prescribing adviser. Practice visits and the distribution and completion of workbooks occurred between September 1993 and March 1994, PACT data for all NSAIDs was used to identify changes in prescribing before and after the programme. A combination of 27 indicators, in terms of items and cost per 1000 patients, were chosen to identify overall changes and potential switches between individual drugs, or to generic alternatives. Comparison of the practices in each of the three groups at analysis revealed similar distribution in terms of stratification criteria. Eleven (38%) Group 2 practices returned completed workbooks. Overview indicators (those not targeted) showed similar trends of either increase or decrease, in cost and volume, across all three groups, whereas targeted indicators demonstrated a more mixed picture between groups. In summary the total number of statistically significant changes for targeted indicators in Groups 1, 2, and 3 were 10, 8 and 1 (changes in items per 1000 patients), and 12, 10 and 3 (changes in cost per 1000 patients) respectively. Targeted indicators revealed more statistically significant changes in Group 1 (active feedback) than Group 2 passive feedback) which showed more changes than Group 3 (reference group). Active feedback was more effective at bringing about a required change than the use of passive feedback; both approaches had more impact than that registered by the reference group. The Group 2 analysis presented included both responders and non responders, thus a more marked benefit of the workbook was perhaps masked. These results have implications for the future provision of prescribing advice to practices. Advisers currently use both active and passive methods to provide prescribing feedback to practices. The high resource intensiveness of practice visits leads many authorities to rely increasingly on less effective methods such as bulletins. Evaluation of the cost effectiveness of methods used to influence prescribing are required. PMID- 10849922 TI - An investigation of the factors affecting community pharmacists' selection of over the counter preparations. AB - A postal survey of pharmacists in a random sample of community pharmacies in Great Britain was carried out to ascertain the factors which influenced their decisions when recommending a product for Over The Counter (OTC) sale. Six categories of condition which regularly present in community pharmacies were selected and the pharmacists were asked to state which first line product they would recommend for each condition, in an uncomplicated situation with no other relevant problems. Of the 1138 questionnaires sent, 635 were suitable for analysis giving a response rate of 56.7%. The recommendations were found to be appropriate in 99.7% of cases. Factors which had the greatest influence on product selection were active ingredients, evidence of product efficacy, ease of use by patient and patient characteristics such as age and concurrent medication. Economic factors such as profit, pressures of excess stock and product promotion by sales representatives were shown to have a significantly greater influence on proprietor pharmacists than the other categories of pharmacist. The results of this study suggest that in the majority of cases pharmacists' decisions on OTC drug therapy are based on clinical and patient factors and whilst commercial factors may be involved they do not compromise the quality of the recommendations. PMID- 10849923 TI - Clinical and economic impact of a pharmacist-intervention to promote sequential intravenous to oral clindamycin conversion. AB - A multicentre, prospective, controlled study compared the clinical efficacy, safety and economic impact of a pharmacist intervention to promote sequential intravenous to oral clindamycin conversion. A total of 473 patients receiving intravenous clindamycin for at least 72 hours were included in the study. Two groups were established: an intervention group (204 patients) in which an informative sheet recommending the sequential treatment was provided, and a control group (269 patients). Clindamycin was prescribed for respiratory infections in 38.9% and for prophylaxis in surgery in 25.4% of the patients (71% were contaminated surgery). No difference between groups regarding sex, infection severity, health status or clinical progress was observed. Both the step-down treatments after 72 hours of intravenous clindamycin and the change to the oral route later on, were significantly increased with the intervention (p < 0.001, p < 0.001 respectively). No significant differences between both groups were found in the number of patients with adverse effects associated with the i.v. therapy, although the incidence tended to be lower in the intervention group (49/204 intervention versus 85/269 control, p = 0.07). Compliance with the recommended clindamycin dosing regimen was significantly higher in the intervention group, in which 1.3 days reduction of intravenous therapy provided an average cost savings of PTA5246 (95% CI 2556-7935) per treatment. A higher reduction of 1.7 days was achieved in those patients candidates for switch therapy on the third day of intravenous clindamycin. A sequential program with clindamycin may provide a cost effective alternative to conventional therapy and the introduction of an information sheet is a cost-effective strategy to promote it. PMID- 10849924 TI - Counselling patients on psychotropic medication: physicians' opinions on the role of community pharmacists. AB - OBJECTIVES: To study physicians' opinions on community pharmacists' involvement in counselling patients on use of psychotropic medication. DESIGN: A postal questionnaire with open-ended questions completed by physicians. SUBJECTS: A six per cent random sample (n = 759) of the members of the Finnish Medical Association representing physicians working in hospitals, community health centres, private practices and occupational health services (response rate 64%, n = 487). MAIN MEASURES: Physicians' opinions concerning community pharmacists' involvement in counselling patients about purpose of the medication and adverse effects of benzodiazepines and neuroleptics. RESULTS: When classifying opinions into five categories, majority (72%) of the physicians regarded community pharmacists as a provider of comprehensive or at least general information about adverse effects of benzodiazepines, but only 43% about the purpose of the medication. Correspondingly, 60% of the physicians regarded community pharmacists as a provider of comprehensive or general information about adverse effects of neuroleptics, but only 35% about the purpose of the medication. There was a strong correlation between physicians' opinions concerning pharmacists' involvement in counselling patients about the purpose of the medication of benzodiazepines and neuroleptics (Spearman's coefficient 0.667), and about adverse effects of both type of medication (0.668). Male physicians had more fixed opinions, both positive and negative, than female physicians. CONCLUSIONS: Finnish physicians are still quite critical about community pharmacists involvement in counselling patients on psychotropic medication. Especially with neuroleptics, physicians feel that pharmacists should be cautious when discussing the purpose of the medication if it is excluded from the prescription. PMID- 10849925 TI - Risk factors for the development of adverse drug events in hospitalized patients. AB - Adverse drug events in hospitalized patients lead to increased morbidity, mortality and costs. Early detection of adverse drug events could aid in the prevention of these adverse outcomes. A cost-effective system for the early detection of adverse drug events should focus on high risk patients. A study was set up with the primary aim to identify characteristics that are associated with the development of adverse drug events (ADEs) in hospitalized patients. ADE reports were gathered from physicians and nurses (spontaneous reports) and from patients after intensive ward interviews by hospital pharmacists. All patients admitted to the internal medicine wards of two Dutch hospitals, during a two month period, were included. The following characteristics were analyzed for their potential relationship to the occurrence of ADEs: age (categorized), gender, number of drugs prescribed during hospital stay, types of drugs used and changes in drug use on admission. Age was found to be inversely associated with the development of ADEs (OR 0.36, CI 0.21-0.61 for age category > 80 years; OR 0.56; CI 0.31-1.02 for age category 75-80 years and OR 0.69; CI 0.42-1.11 for age category 60-74 years). Furthermore, statistically significant associations were found for the number of drugs prescribed per hospitalized patient (for the class of 4-6 drugs per patient OR 2.61, CI 1.32-5.18), for newly prescribed drugs (OR 6.65, CI 2.63-16.81) and for the cessation of drugs on hospital admission (OR 1.50, CI 1.02-2.20). The use of gastrointestinal drugs (OR 2.13, CI 1.32-3.45), central nervous system drugs (OR 1.66, CI 1.07-2.57) and antibiotics (OR 2.44, CI 1.65-3.60) were associated with the development of ADEs, when compared to all other drugs taken by the patients. In this study, the most important risk factors are the number of drugs used per patient and the starting of a new drug during hospitalization. As most hospitalized patients start new drug therapies while in hospital, this seems an inappropriate focus. However, careful monitoring of patients using more than 7 drugs at a time may be possible in a cost-effective system for the early detection of ADEs. PMID- 10849926 TI - Oral bioavailability of phenobarbital: a comparison of a solution in Myvacet 9 08, a suspension, and a tablet. AB - PURPOSE: A three-way crossover study with seven healthy male volunteers was conducted to determine the relative bioavailability of phenobarbital after single dose administration of 100 mg of phenobarbital as oral solution in Myvacet 9-08, and as a suspension, compared with a 100 mg phenobarbital tablet. MATERIALS AND METHODS: At 4-week intervals each subject received the solution in Myvacet 9-08, the suspension and the tablet in randomized order. Blood samples were collected for 48 h after each dose for analysis of phenobarbital. From the individual serum concentration-versus-time curves Cmax and Tmax were determined and AUC0-48 was calculated. RESULTS: All three oral dosage forms of phenobarbital are bioequivalent. No significant differences in Tmax were observed. CONCLUSION: The oral solution in Myvacet 9-08, and the suspension of phenobarbital proved to be bioequivalent to a tablet. PMID- 10849927 TI - [Stress proteins in eukaryotic cells]. AB - The review concerns properties and function of heat shock proteins (hsp) in the eukaryotic cell. The factors inducing Hsps' expression are considered with the emphasis on the fact that most of these agents are able to affect the function of peptide-synthesizing and protein-modifying machineries. The common outcome of this effect is accumulation of wrongly assembled protein structures which results in activation of heat shock transcription factors. The major property of proteins belonging to Hsp70 and Hsp90 is their chaperonic activity or the ability to bind newly synthesized or damaged polypeptides followed by restoration of the native structure of the latter. In this activity Hsp70 and Hsp90 are assisted by other proteins, and the work of such chaperonic systems is described. The function of the major stress protein Hsp70 in the eukaryotic cell was analysed, and on the base of a huge amount of data in has been resumed that proteins of this family constitute one of the most abundant systems of cell protection against cytotoxic factors. In the concluding part of the review we present some ways of application of our knowledge of Hsp in ecology, for analysing biological pollution, and in medicine, for increasing tissue or organismal resistance to injuring factors. PMID- 10849928 TI - [Cytologic characteristics of "dark" and "light" cells in the brain amygdaloid complex]. AB - This paper reports data on cytological peculiarities of neurons of two main zones of sexual dimorphism in brain amygdala (dorsomedial nucleus and anterior cortical nucleus). The main attention was paid to some characteristics of "dark" and "pale" cells found in the amygdaloid complex for the first time. It is supposed that the dark and pale cells are targets for gonadal steroids, whose cyclic changes in concentration in the blood difined their functional states. Though the ultrastructure of dark and pale cells of the amygdaloid complex is similar to that of neurosecretory cells of hypothalamus, there are necessary electron microscopic and cytochemical evidences. PMID- 10849929 TI - [Features of morphofunctional rehabilitation of Mauthner neurons after fatigue]. AB - Structural reorganization of smooth endoplasmic reticulum (SER) in relation to changes in functional state of neurons has been investigated using fatigue and subsequent rhabilitation of the goldfish Mauthner (M-) cells as experimental approach. The recovery of original structure of SER in distal parts of dendrities after its significant proliferation, caused by a 3 h natural stimulation, markedly retarded, as compared with quickly normalized functional activity of M cells. At the same time in somata and proximal parts of dendrites the structural recovery of SER coincided with restoration of the initial function of M-cells. The results suggest that within a single neuron SER with its obvious structural plastisity the neuron functional activoty is supported and restored through regulating the extent of proliferation angmenting Ca(2+)-accumulation in its compartments. Nevertheless SER posseses certain autonomy in structural recovery within somata and dendrites. Such differences of SER plasticity in different parts of the same neuron presumable reflect differences in interaction of its individual compartments with the cytoskeleton and adjacent cytoplasm, or may be caused by different activity of synapses situated on the soma and dendrites. PMID- 10849930 TI - [Toxicity of magnetite-dextran particles: morphological study]. AB - Females of OFI mice were given single repeated intravenous injections of magnetite-dextran nanoparticles (MD3), the total partical diameter being 49 nm, with the magnetic core diameter equal to 10-15 nm. MD3 is a superparamagnetic preparation commonly used for magnetic resonance imaging (MRI). The liver, spleen, heart, kidney, and lung microstructures of these mice were determined after MD3 administration. Both dose- and time-dependent changes in the examined organs were compared after single and repeated MD3 doses. MD3 induces an increse in ferritine and iron levels in all the organs, the appearance of small aggregates of lymphoid cells in the liver, the appearance of iron-containing cell formations in hepatic sinusoids, presumably composed of the Kupffer cells and portal macrophages, splenomegaly, and hemostasis of spleen blood vessels. The pronounced morphological alterations have been revealed primarily in the liver and spleen after a single administration of high MD3 doses and after repeated MD3 injections. The results of The present investigation seem to narrow somewhat the safety limits of superparamagnetic iron oxide particles. Nevertheless, the degree of morphological changes in the liver and spleen in our experiments appeared to be rather low even after a single MD3 dose that exceeds approximately by 200 times a dose necessary for diagnostics in MRI. PMID- 10849931 TI - [Effect of gangliosides on intensity of the lipid peroxidation process and structural changes in neuronal membranes, caused by toxic doses of glutamate]. AB - We studied effects of gangliosides on the level of lipid peroxides and microviscosity of membrane lipid bilayer in primary dissociated cultures of cerebellar granule cells prepared from 8 day-old rats under conditions of neurotoxic effect of glutamate. It was found that glutamate (100 mkM) treatment of primary cultures activated the processes of lipid peroxidation and decreased microviscosity of neuronal membranes determined as a degree of pyrene excimerization. It was also shown that preincubation of granule cells with gangliosides did not prevent the accumulation of TBA-reactive products induced by glutamate. At the same time gangliosides significantly decreased the membrane fluidizing effect caused by glutamate. PMID- 10849932 TI - [Anomalies in cytoskeletal microtubules in a mutant line of sugar beets]. AB - The morphological phenotype of meimutation acd (abnormal cytoskeleton dynamics) in Beta vulgaris was analysed using a classic method of achromatic figure visualization (Navashin fixative). The mutation demonstrates two different phenotypes under different growth conditions. The first variant is characterized by formation of chaotic network of microtubule bundles instead of two bipolar spindles at metaphase II. Such abnormality leads to the formation of polyads and monads. The second variant of phenotype demonstrates desorientation of spindles at metaphase II and partial asynapsis of homologous chromosomes. PMID- 10849933 TI - [Differences in phorbol-dependent phosphorylation of regulatory proteins and contraction of phasic and tonic smooth muscle]. AB - Smooth muscles are divided into slowly contracting tonic and relatively fast phasic muscles. In both cases Ca2+ is a key mediator of the contractile response. However, the appearance of a tonic component during sphincter or arterial muscle contraction and its absence in contracting visceral smooth muscle is characteristic of their difference. We have found that in chicken tissues phorbol 12,13-dibutyrate (PDBu) induces a sustained contraction in carotid arterial muscle, but provokes no contraction in phasic gizzard smooth muscle. Next we were aimed to find differences in PDBu-induced phosphorylation of the key proteins involved in regulation of smooth muscle contraction, i.e. caldesmon, myosin light chain kinase (MLCK), and the myosin light chain kinase-related protein (KRP, also known as telokin). Two correlative differences were observed. 1. PDBu stimulated phosphorylation of MLCK in tonic smooth muscle and had no effect on the level of MLCK phosphorylation in phasic muscle. Phosphopeptide mapping suggests the involvement of mitogen-activated protein (MAP) kinases in phosphorylation of MLCK in situ. 2. PDBu induced phosphorylation of MAP-kinase sites in caldesmon in both types of smooth muscle, but this phosphorylation had no significant effect on caldesmon functional activity in vitro. For the first time we have shown that in gizzard PDBu also stimulates a yet unknown transitory caldesmon-kinase different from protein kinase, C, Ca2+/calmodulin-dependent kinase II and casein kinase CK2. 3. No significant difference was found in the kinetics of PDBu-dependent phosphorylation of KRP in tonic and phasic smooth muscles. KRP was also demonstrated to be a major phosphoprotein in smooth muscle phosphorylated in vivo at several sites located within its N-terminal sequence. Protein kinases able to phosphorylate these sites were identified in vitro. Among them, MAP-kinase was suggested to phosphorylate a serine residue homologous to that phosphorylated in MLCK. 4. p42erk2 and p38 MAP-kinases were found in phasic and tonic smooth muscles. Both were responsive to PDBu in cultured chicken aortic smooth muscle cells, and their role in phosphorylation of MLCK and low molecular weight isoform of caldesmon was evaluated. PMID- 10849934 TI - [Regularities in formation of sister chromatid exchanges (SCE) in three generations after irradiation]. AB - Frequencies of sister chromatid exchanges (SCE) induction in 3 generations of Chinese hamster cells (clone 237-7) after gamma-irradiation with a dose of 3 Gy have been studied. It is shown that the frequency of SCE significantly increases only in the 1st postirradiation cycle; it does not differ from the control in the 2nd and 3nd cycles. Irradiation induces production of cells with levels of SCE, which have practically never been discovered in non-irradiated population. These cells are eliminated from the population during the next division. Furthermore, it is stated that SCE can be produced in the same locus of chromosome during successive cell divisions. These results permit to conclude that the mechanism of formation of radiation-induced SCE differs from that in normal non-irradiated population. PMID- 10849935 TI - [Topography of cell ahesion molecules CD9, CD24, L1 and N-CAM on the surface of neuroblastoma cells studied using chemical cross linking]. AB - Cell-adhesion molecules are thought to play crucial roles in development and plasticity in the nervous system. Four neural cell adhesion molecules CD9, CD24, L1 and N-CAM are associated in the surface membrane of cultured neuroblastoma cells as studied by chemical cross-linking with bifunctional reagent 3,3' dithiobis (sulphosuccinimidyl-propionate) followed by a subsequent immunodetection using antibodies directed against the above molecules. We obtained direct evidence of CD9 and L1, but not CD9 and N-CAM clasterisation, also interactions of CD24 with L1, CD24 with N-CAM and some others. These observations illustrate topography of neural cell adhesion molecules located in the vicinity to each other and imply the basis for their functional cooperativity. PMID- 10849936 TI - [Disruption of male meiosis in the pea Pisum sativum L., caused by ms3 mutation]. AB - Analysis of morphological phenotype of meimutation ms3 in Pisum sativum was made at the light microscopic level with vizualization of MT cytoskeletal structures. This mutation disrupts simultaneously the chromosome cycle, nuclear envelope breakdown, and MT cytoskeleton cycle during meiosis in pollen mother cells. PMID- 10849937 TI - [Dynamics of mitotic index of buckwheat root meristems, developing from seeds subjected to percussive wave treatment]. AB - A method of percussive wave treatment of seeds, promoting activization of physiological processes in plants, was proposed. The influence of physical treatment on cell nuclei and phytohormones was investigated. We examined dynamics of mitotic indices (MI) of the root meristems of buckwheat seedlings 48-58 h after the treatment. The seeds were treated with pressures of 11, 23, 29 MPa. The synchrony of cell divisions was estimated according to the augmentation of maximum MI versus average MI. The maximum and the average MI of control seedlings was 9.6 and 8.1%, resp. After the treatment with the pressure of 11 MPa the average MI increased by 2.6 times, and synchrony raised by 50%. After the treatment by the pressure of 23 MPa the average value of MI did not change versus the control, but the synchrony enhanced by 2.8 times. After the treatment of the pressure of 29 MPa the average MI decreased by 31%, the synchrony increased by 33%. Examination of phase indexes allowed to reveal the damage of cytotomy, transition to metaphase, and mitotic apparatus formation. The MI of root meristems of the test seedling aged 8 days topped the control by 21, 156 and 20%. So, the pressure of 11 MPa promoted MI increase in 2 days, and those of 23 and 29 MPa in 8 days. The investigation of hormone balance allowed to detect a raising level of abscisic acid. Its influence on the plant growth depends on the content of zeatin. A hypothetical model of plant reaction mechanism was proposed, based on the obtained results. A momentary pressing in the percussive wave was a stressor, promoting the accumulation of ABA and "growth rest", that changed for an active growth induced by zeatin. PMID- 10849938 TI - [Do not trust anyone over 30; testosterone for all men over 50?]. PMID- 10849939 TI - [Cortisol in critically ill patients with sepsis: physiologic functions and therapeutic implications]. AB - Modern immunology reveals that cortisol interacts with the immune response at virtually all levels exerting both suppressive and permissive effects. A pre requisite for the defense against severe infections is the functional integrity of the hypothalamic-pituitary-adrenal axis (HPAA) resulting in adequate cortisol production. There is increasing evidence that cortisol physiology and regulation are substantially altered in the course of a septic shock. Patients with septic shock may suffer from relative adrenocortical insufficiency resulting in a relative deficiency of cortisol production. In addition, the number and the affinity of cellular glucocorticoid receptors are decreased by which cortisol action at cellular level is reduced. Since septic shock and adrenal insufficiency are sharing hemodynamic abnormalities such as hyperdynamic circulation and peripheral vasodilation, the administration of stress doses of hydrocortisone appears to be a rational therapeutic approach in patients with septic shock. Controlled studies have shown that stress doses of hydrocortisone attenuate the systemic inflammatory response. In two recent double-blind studies stress doses of hydrocortisone given in patients with septic shock have been demonstrated to reduce the time to shock reversal. The most important hemodynamic effect was an increase in systemic vascular resistance. Earlier weaning from vasopressor therapy was associated with a trend towards improvements in organ function and towards decreased mortality, respectively. Large-scale trials are on the way investigating the benefit of stress doses of hydrocortisone on the mortality of septic shock. The focus of this review are changes in glucocorticoid physiology and regulation during septic shock. Effects of stress doses of hydrocortisone on immune response and vascular tone in the course of a septic shock are being discussed. PMID- 10849940 TI - [Focal proliferation in reactive hyperplastic parathyroid tissue in end-stage renal failure--implication for autotransplantation after total parathyroidectomy]. AB - Total parathyroidectomy with simultaneous autotransplantation may be associated with recurrence of graft-dependent hyperfunction due to excessive proliferation. We performed macroscopic tissue selection with a stereomicroscope prior to autotransplantation, which resulted in very low recurrence rates. As this technique greatly depends on experience, we investigated the possibility of additionally using proliferation staining (PCNA, MIB-1) for the detection of dysfunctional tissue. Selected tissue from 26 patients was investigated. Serial sections of freshly removed parathyroid tissue were correlated with their macroscopic appearance, HE and immunohistochemically stained paraffin sections, and with semithin Epon sections. The asymptotic growth mode of clonal proliferating regions was reflected by highest staining intensity (1-5%) in small to medium sized foci (diffuse, up to 3 mm in diameter) and very low staining in large areas (diffuse or nodular, 5-15 mm in diameter, from 0.03 to 0.003% positive cells). Thus, very large dysfunctional regions with (almost) no proliferation could not be detected by this method. However, they were very evident on macroscopic investigation. In conclusion, multiple fulminant recurrence after parathyroidectomy can be prevented by selecting tissue after proliferation staining. This may allow a delayed autotransplantation after total parathyoidectomy for those surgeons lacking experience in macroscopic tissue classification. PMID- 10849941 TI - Beneficial effects of atorvastatin in the treatment of hyperlipidemia after renal transplantation. AB - Despite the availability of various lipid lowering drugs, the treatment of hyperlipidemia, one of the most important risk factors for morbidity and mortality after organ transplantation, remains a therapeutic challenge. We investigated the safety and efficacy of a new HMG-CoA reductase inhibitor, atorvastatin, in renal transplant patients whose serum lipids were insufficiently controlled by diet and treatment with other lipid lowering drugs. Twenty-four patients (14 males/10 females; mean age 51.2 +/- 2.3 years) were converted to low dose atorvastatin (10 mg/day) at a mean of 67.7 +/- 8.6 months after renal transplantation and prospectively followed for 3 months after initiation of the study drug. HDL, LDL, and total cholesterol, triglycerides, serum creatinine and CPK levels were evaluated pre (-3, -1, 0 months) and post conversion (+1, +3 months). In the eighteen patients who completed the study, low dose atorvastatin therapy led to a significant reduction in total cholesterol (304.6 +/- 13.2 vs. 247.6 +/- 12.0 mg/dl; p = 0.007) and LDL cholesterol (191.9 +/- 9.0 vs. 141.8 +/- 14.7 mg/dl; p < 0.0001) and a modest reduction in serum triglyceride levels at three months after conversion. We conclude that low dose atorvastatin (10 mg/day) can be successfully used and appears to be safe in the treatment of posttransplant hyperlipidemia. Its long-term effects on patient morbidity and mortality as well as graft survival should be investigated in larger and more prolonged prospective trials. PMID- 10849942 TI - Serum levels of progesterone in patients with preeclampsia. AB - In a matched pair-study study we investigated the hitherto controversially discussed serum levels of progesterone in 40 women with severe preeclampsia (PE) and 40 normotensive controls. Serum levels were determined by applying a sandwich enzyme-linked immunosorbent assay (ELISA). Median serum levels of progesterone in preeclamptic women and in controls were not statistically significant (P = 0.73). Our study indicates that the absence of altered serum levels of progesterone may not reflect the potential role of this hormone in preeclampsia. PMID- 10849943 TI - [Fatal methadone poisoning of a child]. AB - The substance methadone is used for substitution therapy since the 1960s in the U.S. Mainly because of the endemic spread of HIV-1 infections among intravenous drug abusers methadone was made legally available through medical prescription in Austria in 1987. Legal authorities today also allow the patient to take home the necessary daily consumption for weekends or public holidays. The drug is distributed as a watery solution in tiny bottles, which are fitted with an ordinary screw cap. This kind of distribution may, however, have fatal consequences. This is demonstrated in the following case of accidental poisoning of an infant: A two-year-old girl whose parents were both participating in the substitution scheme was found dead in her bed in Vienna in 1997. Forensic autopsy revealed a methadone concentration in the liver tissue of 640 ng/g. The criminal investigation determined that the girl had opened a bottle of methadone solution and subsequently had taken the drug. Considering the circumstances of this accident, from the medical point of view safety devices for the screw caps of the methadone bottles should be required by law, in order to avoid future accidental poisoning. PMID- 10849944 TI - [Dr. Nicolaas Tulp. A critical view of Rembrandt's Anatomy Lesson]. AB - Rembrandt's painting 'The Anatomy Lesson' (1632) is revolutionary in its portrayal of members of the Anatomic Guild. It has an entirely new composition and vividly depicts the dynamics of the event and the interest of the participants. However, the structures of the dissected forearm have been taken from a copy and not from the original. The possibility of anatomic errors is discussed here. A short biography of Dr. Tulp is also included. PMID- 10849945 TI - [Pain management in view of current new legislative updates and their practical consequences in Austria]. AB - Although the WHO edited guidelines for pain treatment as early as 1986, practical management has frequently remained inadequate, especially in cancer patients. Traditional adherence to restrictions from the former Austrian Controlled Drug Act which have resulted in ongoing limitations in the prescription of opioids as well as complicated formal regulations in the current law represent two major obstacles. As a consequence, recent legislation of a "state of the art" pain management in Austria facilitates adequate provision of analgesics on the one hand, and may, on the other, even result in claims for indemnity should these be withhold. PMID- 10849946 TI - [Perspectives in geriatrics at the threshold of 21. century]. PMID- 10849947 TI - Prevalence of lower urinary tract symptoms and urinary incontinence in the elderly: recent data from Austria. PMID- 10849948 TI - Living well and dying not too badly: integrating a whole life into a tolerable death. PMID- 10849949 TI - Geriatrics for the 3rd millennium. AB - It is a common knowledge that the population around the world is growing old at an unprecedented rate. This is the success story of increasing life expectancy. The demographic breakthroughs occurred in the 20th century. The quality of life breakthrough is our challenge for the 21st century. The implications of the growing elderly population are many, including: rising total health care expenditures; the increasing needs for long-term care services; and the need for expert and focused health care services. Since health care costs increase with advancing age of populations, these costs will fall on older persons, families, and society generally. There is real value for everyone in meeting the needs of an aging society, especially if seen as part of a social contract. The ability to live independently improves with access to good care, but decreases dramatically for those with age-related disabling conditions. With the decreasing number of informal caregivers around the world, frail elderly will require more formal long term care services. However, due to inadequate attention given to long-term care issues, numerous developed countries have recently started to struggle to develop long-term care service programs that will both meet the rising needs for this service and be cost-effective. Effective medical care requires expertise in functional assessment, interdisciplinary care, and advances in treating symptoms of aging. The field of geriatrics is essential to modern health care, and geriatricians need to have proper training that focuses diagnosing and treating this group of patients. Quality care will not only help the elderly to live productively and independently, but it will also tremendously benefit families and communities. PMID- 10849950 TI - [Adverse effects of psychotropic drugs in the elderly]. AB - As in other organs, homeostatic mechanisms become insufficient in the aging brain, in part because of reduced activity of various neurotransmitter systems. Counter-regulatory processes are therefore reduced and reactions to drugs may be increased. Aside from these pharmacodynamic mechanisms, the increased toxicity of psychotropic agents in the elderly may be a result of pharmacokinetic changes. The total clearance of a number of drugs is reduced in old age, hence steady state plasma concentrations of these compounds will be increased when conventional doses are used. Because of these changes, the central nervous system is especially vulnerable in elderly. With tranquilizers and hypnotics, sedation is increased and cognitive function may decrease. The risk of falls and injuries is enhanced with all psychoactive drugs. In addition, response to agents with anticholinergic properties (classical antidepressants and neuroleptics) is increased in old age and is accompanied by peripheral symptoms such as urinary retention, obstipation, tachycardia and visual disturbances. In the central nervous system there may be impairment of intellectual capabilities, agitation, and ultimately delirium. Further, the undesirable cardiac effects of tricyclic antidepressants should be noted. The selective serotonin reuptake inhibitors (SSRI) and moclobemide have an advantageous pharmacological profile compared to older antidepressants. Of the neuroleptics, the benefit-risk relation of the newer atypical agents appears to be more favorable than that of classical neuroleptics. PMID- 10849951 TI - [Geriatric medicine at European universities]. AB - The demographic data of industrialised nations predict an unavoidable shift towards an increase of the ratio of old people to total population. Thus, senescence presents a challenge to sociology, psychology and medicine. The latter responds by the holistic approach of geriatrics and medical gerontology. The diversity of European nations and their medical schools lead us to assess the respective incentives in the field of geriatric medicine. By means of a questionnaire with a response rate of 41%, by direct contact and personal communication and by consulting published and unpublished papers we obtained a survey of current geriatric activities in European universities. The results of this investigation are summarized. PMID- 10849952 TI - Vertebral osteophytosis and vertebral deformities in an elderly population sample. AB - AIM OF THE STUDY: We investigated the association between vertebral osteophytosis and vertebral deformities in an elderly population sample, and the influence of some risk factors on spinal osteophytosis and deformities. SUBJECTS AND METHODS: A population sample of 280 women and 263 men, all Zagreb residents older than 45 years, participated in the study. Radiographs of the thoracic and lumbar spine were evaluated for the presence of osteophyte formation and vertebral deformities. Osteophyte size was graded on a scale from 0 to 4. Vertebral deformities were determined by the semiquantitative method of McCloskey. The chi square test was used to analyse the association between vertebral osteophytes and deformities, and the influence of several risk factors was investigated by discriminate analysis. RESULTS: The prevalence of vertebral osteophytosis was 47.9% in men (36.5% in the thoracic and 21.3% in the lumbar spine) and 56.0% in women (36.0% in the thoracic and 23.9% in the lumbar spine). The prevalence of vertebral deformities was 8.3% in men (5.3% in the thoracic and 3.4% in the lumbar spine) and 12.5% in women (7.9% in the thoracic and 5.4% in the lumbar segment). There was a significant association between deformities and osteophytosis on the lumbar segment of the spine (P = 0.0240 men, P = 0.0152 women). Analysing the influence of several risk factors, age was found to be the most associated with both vertebral deformities and osteophytosis. Obesity was significantly associated with osteophytosis. CONCLUSIONS: We found a significant association between vertebral osteophytosis and deformities in the lumbar segment and no relationship in the thoracic segment. This implicates different etiologies of vertebral deformities in the thoracic and lumbar spine. PMID- 10849953 TI - [Improvement of rehabilitation outcomes of hip fractures: discharge assessment by patient care team, case management and wound healing]. AB - Fall related hip fractures in elderly persons may substantially deteriorate a previously worthwhile life. Anxiety, isolation, depression and the immediate need for help jeopardize surgery and successful rehabilitation. It was therefore of interest to evaluate the impact of a comprehensive case management guided by a discharge assessment which included medical and social criteria. In a prospective, open study, conducted by a community hospital in Vienna and the Research Department of the Red Cross, 124 carefully selected patients (117 female, 7 male, mean age 81.8 +/- 7.0 years) over a period of six months were assessed one week before hospital discharge by a multiprofessional team. Patients were excluded for mental illness, dementia, disabling neurological diseases and noteworthy surgical complications. Thirty-four patients (mean age 83.7 +/- 7.6 years) were considered as intervention group. Ninety essentially independent patients (mean age 81.1 +/- 6.6 years) were considered as control group. A specialised nurse from the Community of Vienna was responsible for the link between the patients of the intervention group, the rehabilitation unit and the Social Services, for the discharge check lists and the feed back questionnaires (2, 6 and 12 weeks after discharge). All patients were asked for a check up 12 weeks after discharge in order to investigate needs and substantial changes in the ADL or required care. In the control group, nearly all patients reached the pre-traumatic level, whereas in the intervention group a drop out rate of 1/5th and a higher over all need of Social Services care was observed. However, in respect of the higher age, the more compromised health and activities, even this group of patients obviously profits by this case management strategy. In conclusion, surgery and rehabilitation need a thoroughly performed discharge assessment followed by a network of comprehensive Social Services measures to treat successfully high risk elderly patients after fall related hip fractures. PMID- 10849954 TI - [Renaissance of vaginal hysterectomy for cervical carcinoma. 100th anniversary of the first abdominal radical surgery of cervical carcinoma by Ernst Wertheim on November 16, 1898]. AB - In the 100 year long history of the abdominal radical operation of collum carcinoma, due to the continued clinical surgical and scientific work of several generations of physicians, abdominal radical operation with standardized pelvic lymphonodectomy has become the method of choice for surgical treatment of collum carcinoma since 1970. Vaginal radical operation in its various forms has since played only a very restricted role in surgical treatment of collum carcinoma. According to the opinion of the majority of cancer surgeons vaginal radical operation had to be abandoned in view of discontinuous spreading of carcinoma into the regional pelvic lymph nodes. Because of its simplicity vaginal radical operation would still be useful today for very old high-risk patients and very young women with early invasive collum carcinoma detected in cancer screening. In view of our knowledge of the lymph node problem in collum carcinoma, however, this can no longer be the vaginal radical operation of past generations. In order to prevent critical objections to vaginal radical operation, the early attempts of Stoeckel, Suboth Mitra, Bastiaanse, Navratil, Inguilla, and Akashi were resumed. Since 1989 attempts have been made to combine vaginal radical operation of collum carcinoma with laparoscopic pelvic or para-aortic lymphonodectomy. The development has passed the following stages: the development of laparoscopic pelvic and para-aortic lymphonodectomy based on staging criteria the combination of laparoscopic lymphonodectomy with vaginal radical operation of collum carcinoma the combination of laparoscopic lymphonodectomy with complete laparoscopic radical hysterectomy and only subsequent vaginal removal of organs. PMID- 10849955 TI - [Glucose-insulin-potassium (GIK) in prevention and therapy of myocardial ischemia]. AB - Recent times are witnessing a resurge of metabolic interventions in order to preserve myocardial tissue and improve cardiac function after hypoxic or ischaemic events. The basic principle of this therapeutic approach which differs completely from surgical and radiological revascularisation techniques as well as from direct pharmacologic stimulation of contractility by catecholamines or phosphodiesterase inhibitors is illustrated by the effect of glucose-insulin potassium (GIK) infusions. Physiology of myocardial metabolism and its pathophysiology in hypoxia and ischaemia are discussed in their relation to the administration of GIK with its main effect of switching from the preferential fat oxydation under normal conditions to a greater percentage of glucose utilisation. GIK can be used in therapeutic intention after an ischaemic insult (myocardial infarction, preoperatively) or as a preventive measure (pre-ischaemic) e.g. prior to a planned intervention (cardiac surgery). GIK improves myocardial energy supply, economizes oxygen utilisation, stabilizes hypoxic cells, diminishes the area of infarction, and increases myocardial contractility and coronary blood flow. Both in controlled trials and in meta-analyses, GIK administration resulted in reduced mortality from myocardial infarction. In conclusion, metabolic interventions present a promising alternative or adjunctive therapeutic tool for the restitution or maintenance of myocardial integrity and cardiac function impaired or threatened by ischaemic events. PMID- 10849956 TI - Intra- and interindividual reproducibility of heart rate variations in the tilt table test. AB - Various methods are used for the routine diagnosis of autonomic regulation disorders. The evaluation of blood pressure and heart rate response during active orthostasis together with assessment of the 30/15-ratio (Ewing's ratio) was proven to be a valid method. One main disadvantage of these tests is their dependence on the active cooperation of the patient. In contrast, passive orthostasis using the tilt-table may also be carried out in bed-ridden patients. The test comprises a manual tilting manoeuvre of the patient lying on the tilt table, with continuous assessment of heart rate and blood pressure. Until now, the main advantages of the tilt-table test were considered to be its better standardisation and good reproducibility, especially with regard to the diagnosis of vasovagal syncope. Several authors have postulated a specific pattern of initial heart rate response in healthy patients, showing characteristic changes in cases of underlying autonomic neuropathy. However, the diagnostic relevance of these tests is dependent on the reproducibility of the course of initial heart rate response. The objective of our study was to assess the intra- and interindividual reproducibility of initial heart rate response in the tilt-table test in healthy subjects. The tilt-table test was repeated 10 times in all 40 subjects under standardised conditions, and heart rate and blood pressure response were presented in a diagram. Reproducible courses of initial heart rate response were not seen, neither on intra- nor on interindividual comparison. After an initial rise, most subjects showed a rather horizontal course of heart rate, whereas others presented a heart rate response similar to that in the Ewing test, with an initial rise of heart rate around the 15th beat, followed by a decrease, and a maximum around the 30th beat. However, all subjects showed considerable variations in heart rate response within the 10 tilting manoeuvres. A reliable quotient comparable with the 30/15 ratio (Ewing's ratio) in active orthostasis was not seen. Based on these results, we conclude that the initial heart rate response in the tilt-table test is not suitable for routine diagnosis of autonomic regulation disorders, since it is not sufficiently reproducible in healthy individuals. PMID- 10849957 TI - Risk of Pseudomonas aeruginosa cross-colonisation in patients with cystic fibrosis within a holiday camp--a molecular-epidemiological study. AB - OBJECTIVE: A study on the molecular epidemiology of Pseudomonas aeruginosa in patients with cystic fibrosis (CF) from Germany (N = 18) and Israel (N = 12) is presented. The aim is to provide an answer to the question as to whether or not social contact outside the hospital environment involves a potential risk for person-to-person spread of this pathogen. METHODS: Sputa from German and Israeli patients were obtained while these were attending a holiday camp in Israel. The sputum samples were analysed with regard to Pseudomonas aeruginosa. Strains dissimilar in macroscopic appearance and/or antibiotic resistance patterns were genotyped using pulsed-field gel electrophoresis after digestion of genomic DNA with restriction endonuclease Spel. The genetic polymorphism of DNA fragment patterns of all strains (N = 146) was studied for their overall relatedness using a fingerprint software system. RESULTS: Most of the German patients (77.7%) were colonised persistently by a unique clonal type during the four-week screening period. Isolates obtained from Israeli patients displayed a very close clonal relationship and a higher antibiotic resistance as a result of preceding epidemic spread of certain clones before the camp. Additionally, isolates showing identical PFGE patterns were demonstrated once in a single male Israeli patient and in one female German patient, suggesting previous cross-colonisation. CONCLUSION: The occurrence of person-to-person spread through social contact in patients with CF is supported by our findings, but remains a rare event outside the hospital environment, provided appropriate hygienic measures are applied. PMID- 10849958 TI - Successful treatment of high turnover osteoporosis in a patient with adrenocortical insufficiency. AB - We present a case of high-turnover osteoporosis associated with substituted adrenocortical insufficiency (Addison's disease) and its successful treatment with calcitonin and calcitriol. A 43-year-old man presented with markedly reduced bone mineral density (BMD) (lumbar spine BMD -3.46 SD below healthy young adults) after thirteen years of glucocorticoid substitution. Interestingly, his osteocalcin levels indicated an unusually high bone turnover and his alkaline phosphatase levels were also increased. Combination treatment with calcitriol and calcitonin was started. Re-evaluation after twelve months revealed a substantial increase in BMD (+6.8% for the lumbar spine, +15% for the left hip). Alkaline phosphatase levels had normalised and osteocalcin was nearly down to normal. In spite of a thorough evaluation, no other cause of osteoporosis was detected. We discuss these findings in view of the existing literature. PMID- 10849959 TI - [S.S. Korsakov]. PMID- 10849960 TI - [The role of emotional pressure in the development of the early forms of chronic cerebrovascular insufficiency]. AB - The paper presents a comparative analysis of the results of biochemical, neuropsychological and neurophysiological examination of initial chronic cerebrovascular insufficiency in patients without significant psycho-emotional tension (the 1-st group--37 individuals) and with significant emotional tension (the 2-nd group--44 persons). Statistically significant differences between the groups concerned both cerebral hemodynamic parameters and EEG data. On the basis of the results obtained several possible pathogenetic variations of chronic cerebral circulatory insufficiency are suggested on the domination of either primary damages of the structures of the brain and its vessels or the secondary disorder of energetic metabolism and cerebral blood circulation. That, in turn, may be conditioned both by supertension of cerebral systems in chronic stress, and by somatic disturbances. More precise definition of the main pathogenetic factors of initial chronic cerebrovascular insufficiency may be useful for improving efficiency of rehabilitation in these patients. PMID- 10849961 TI - [Specific factors of sex behavior in patients with anorexia nervosa and bulimia]. AB - 65 patients were examined. They were divided into three groups. Group 1 consisted of patients with anorexia nervosa (AN) without bulimia, complicated by cachexia and amenorrhea. Epileptoid personality masculinous body built and behaviour, essential disturbances of sexual self-identification prevailed in this group. 3 patients had homoerotic tendencies, while a syndrome of sex negation developed in 5 cases. In group 2 bulimia was a stage of AN development. The patients had frequently initial endocrinopathy (obesity, dysmenorrhea), experiences of phobia and anxiety, asynchronous disharmonious type of psychosexual ontogenesis. Group 3 of patients was characterized by predomination of bulimia symptoms as a variation of the disease course. Normostenic body built, normal somatoendocrine and psychosexual development were combined with hysteric personal characteristics, mood and sexual fluctuations. PMID- 10849962 TI - [Procedural and developmental aspects of impulse-control disorders in child and adolescent schizophrenia]. AB - 200 schizophrenic patients (130 men, 70 women), which had fallen ill at the age of 4-25 years were examined in outpatient clinic. Either on the separate stages of the disease or during all its course the patients had the following impuls control disorders: of eating--14 patients (7.0%), sexual drive--56 cases (28.0%), self-preservation--37 (18.5%), alcoholism--76 (38.0%), dromomania--38 (19.0%), pyromania--2 (1.0%), kleptomania--4 (2.0%). By the mechanisms of the disorders' development there were impulsive, obsessive and compulsive ones. The author also qualified the morbid impuls-control disorders (87 patients--43.5%) and the evolutive ones (61-30.5%). Morbid signs were considered in terms of the comorbid phenomena combined with the productive and negative symptoms of basic disease. Evolutive manifestations are deviations of ontogenesis which didn't achieve a pathologic form. PMID- 10849963 TI - [The administration of midocalm in the treatment of vertebrogenic algesic syndromes]. PMID- 10849964 TI - [Botulin A toxin: a highly effective drug in the treatment of focal dystonia]. PMID- 10849965 TI - [Effectiveness of instenon ++ in the period of the development of remission in alcoholism]. PMID- 10849966 TI - [EEG mapping in the evaluation of the signs of organic brain damage in patients with mental diseases]. AB - The mapping of EEG was performed in 100 patients with neurosis-like disorders with diffuse organic changes in the brain of hydrocephalic-atrophic character on CT; in 100 patients with organic psychosis and in 50 patients with Altzheimer's type dementia. 100 normals formed a control group. Three informative EEG signs were revealed (space-frequency alpha-rhythm inversion, beta-rhythm reinforsement, alpha-regularity descendence) that allowed to differentiate brain changes in normals and patients with organic brain damage. To accomplish this task a visual EEG evaluation was found insufficiently effective. The accuracy of the differential diagnosis between normals and patients with neurosis-like disorders by means of the method of EEG mapping reached 92%. PMID- 10849967 TI - [Cerebral blood flow in syndrome of vertebral artery]. AB - The paper presents the results of clinical-neurologic study of the indices of cerebral hemodynamics (ultrasonic dopplerography of major arteries of the head, transcranial dopplerography) in 114 patients aged 15-36 years with vertebral artery (VA) syndrome. In the clinical status both subjective manifestations and typical vertebral symptomatology prevailed. There were tendencies to changes of blood flow in extracranial arteries of the carotid system that differed in compression-irritative and reflex-angiospastic forms of VA syndrome. The most significant changes (decrease of blood flow velocity, increase of the indices of the kinematics of blood flow) were observed in vertebral and posterior cerebral arteries (PCA), especially in compression-irritative form of VA syndrome. Homolateral decrease of blood flow in PCA was characteristic for unilateral decrease of blood flow in VA. The results show an important role of the changes of cerebral hemodynamics in pathogenesis of the syndrome of VA. PMID- 10849968 TI - [Specific aspects of thrombocyte system of serotonin in patients with different manifestations of schizoaffective psychosis]. AB - 45 women with different manifestations of schizoaffective psychosis (SAP) were examined. The diagnosis corresponded to ICD-10 (F25). According to the classification elaborated in Mental Health Research Centre of Russian Academy of Medical Sciences, groups of patients were identified with different variants of the psychoses course: a nuclear SAP type; a borderline SAP variation with phasic recurrent course; SAP with progredient variation (schizoaffective variation of schizophrenia). The patients were examined both during the attack and remission. A rate of serotonine uptake (Vmax) in blood platelets, a specific imipramine binding (Bmax) and the level of serotonin in blood platelets were evaluated. It was found that dynamics of both Vmax and the level of serotonin in different SAP types were different, that was related to clinical and biological SAP heterogeneity. A tendency to decreasing of serotonin system functional activity was found in progredient SAP variations, especially during the remission, which was of low quality in these cases. On the contrary, in the borderline variations the indices of the decreased function of serotonin system corresponded well to those of acute psychosis. In nuclear type--a type with the most favourable course of psychosis--any significant changes weren't revealed as compared with the normal parameters. PMID- 10849969 TI - [Control and assessment of the quality of professional competence among physicians and students]. AB - The paper presents a new test method of control which permits to estimate objectively both formal knowledges of students, hospital physicians, post graduates, physicians and their ability for logical reasoning, for determination of cause-effect relationships between separate phenomena. The list of the standards generally accepted for the test's tasks is proposed as well as recommendations for the users are given. 6 variations of the test's tasks are presented as the examples. PMID- 10849970 TI - [The determination of anti-brain antibodies in the blood of patients with brain tumors using hydrosole++ immunoagglutination]. PMID- 10849971 TI - [The study of the influence of picrotoxin on the negative symptoms of schizophrenia]. PMID- 10849972 TI - [Analysis of the association of blood groups ABO and rhesus factor in children with myasthenia gravis]. PMID- 10849973 TI - [Experience with the treatment of depressive patients wtih deprim]. PMID- 10849974 TI - [Specifics of neuroactive amino acid metabolism in acute ischemic stroke]. PMID- 10849975 TI - [Clinical application of opioid agonists for the treatment of opiate dependence]. PMID- 10849976 TI - Imaging the sphenoid sinus: pictorial essay. AB - This paper focuses on the spectrum of sphenoid sinus lesions that may be seen radiologically and the mapping of disease extent. Imaging plays a central role in the assessment of sphenoid sinus disease. Although primary sphenoid sinus disease is uncommon, this sinus is nevertheless affected secondarily by a variety of pathological processes. PMID- 10849977 TI - Deep-seated thoracic and abdominal masses: usefulness of ultrasound and computed tomography guidance in fine needle aspiration cytology diagnosis. AB - Fine needle aspiration cytology (FNAC) was performed under ultrasound and CT guidance in 120 cases. These included abdominal masses (85 cases) and thoracic masses (35 cases) biopsied over a two and a half year period (March 1996 to September 1998). The aim of this study was to assess the contribution of clinico imaging evaluation and image-guided FNAC to the management of patients with deep seated mass lesions. Aspirations in the abdomen were performed from various anatomic sites such as liver (56 cases), lymph nodes (18 cases), gastrointestinal tract (three cases), pancreas (six cases), and kidney (two cases). In the thorax, biopsy was performed in the lung (19 cases) and mediastinum (13 cases). In 112 cases (93.4%) FNAC was diagnostic. Of the lesions that were successfully aspirated, 85% were < or = 5 cm in size. No major complication was encountered. All the successful aspirates could be defined as malignant or non-malignant, but tissue differentiation was possible in 63.7% of malignant lesions and 53.8% of benign lesions. Combined clinical and imaging evaluation for malignancy showed 80% sensitivity and 59% specificity. Although clinicoradiological parameters themselves have certain limitations in diagnosing benign versus malignant lesions, in conjunction with guided FNA they are very accurate and safe in diagnosing deep-seated mass lesions in the thorax and in the abdomen. However, the role of FNA in tissue differentiation of solid lesions such as lymphoma requires further study. PMID- 10849978 TI - Therapeutic embolization for acute haemorrhage in the abdomen and pelvis. AB - Therapeutic embolization for acute haemorrhage is increasingly being utilized. An audit of 34 patients undergoing therapeutic embolization for acute abdominal or pelvic haemorrhage was undertaken, in an attempt to assess the importance of the following variables in determining a successful outcome: coagulation status, transfusion status, time to procedure after onset of circulatory instability, duration of procedure, and the effect of the embolization technique employed. Overall success was 79%, with definitive control of haemorrhage achieved by embolization; 21% required surgical management for rebleeding 4-24 h post embolization. The duration of the procedure and transfusion status of the patient were the most important factors associated with a successful outcome. PMID- 10849979 TI - Clinical and radiological features of biliary papillomatosis. AB - Biliary papillomatosis is a rare disease with strong potential for malignant degeneration. Diagnosis is often not easy and most are made intraoperatively. In the present study, five patients with biliary papillomatosis admitted between 1990 and 1997 were reviewed. Their clinical presentation, radiological and biochemical findings were analysed. The aim of the study was to discern a set of characteristic features that would enable an early diagnosis. All of the five patients presented with recurrent episodes of acute cholangitis and epigastric pain with raised serum alkaline phosphatase. Imaging modalities including ultrasound, CT, endoscopic retrograde cholangio-pancreatogram, MRI and magnetic resonance cholangio-pancreatogram were reviewed. Salient imaging features included a dilated biliary tree with multiple ill-defined and fuzzy filling defects or endoluminal frond-like mass lesions. In conclusion, biliary papillomatosis is a rare but important cause of biliary obstruction with relapsing cholangitis and obstructive jaundice. With a healthy index of suspicion, the diagnosis can be reached when the above features are available. PMID- 10849980 TI - Comparison of transcranial Doppler ultrasound and computed tomography angiography in symptomatic middle cerebral artery stenosis. AB - Transcranial Doppler ultrasound (TCD) and computed tomography angiography (CTA) of 10 patients with middle cerebral artery territory stroke were studied. To obtain data from patients with presumed in situ middle cerebral artery (MCA) stenosis, the study excluded patients with a known source of cardiac emboli, significant carotid stenosis and classical lacunar syndrome. As the gold standard for this study, CTA demonstrated MCA stenosis in all patients (100%), while abnormal TCDs suggesting MCA stenoses were found in only six patients (60%). The stenotic sites differed among patients with normal and abnormal TCDs. Patients with false negative TCDs were found to have more distal lesions (distal M1 or M2 segment) whereas patients with TCD abnormalities tend to have more proximal lesions as demonstrated by CTA. It is concluded that an abnormal TCD is highly suggestive of stenosis of MCA. A normal TCD, however, does not exclude such a lesion, especially in patients with distal M1 or M2 stenoses. Therefore, TCD may not be the best screening test for intracranial vascular stenotic lesion in MCA territory stroke. PMID- 10849981 TI - Acquired immunodeficiency syndrome-related primary cerebral lymphoma: response to irradiation. AB - Acquired immunodeficiency syndrome-related primary cerebral lymphoma (AIDS-PCL) is uncommon. Fourteen cases of presumed AIDS-PCL between 1986 and 1995 were reviewed retrospectively in order to characterize the natural history, and the response to radiotherapy. The median age was 38 years (range 24-65). The median interval between seropositive diagnosis of HIV and AIDS-PCL was 28 months (range 5-113). The median duration of symptoms was 2 weeks (range 0.2-12). At presentation, the Eastern Cooperative Oncology Group performance status (PS) was PS1 (2/14 patients), PS2 (6/14) and PS3 (6/14). The symptoms and signs were non specific and depended on the site and extent of cerebral involvement. There was no characteristic pattern of brain imaging in terms of size, number, location or pattern of contrast enhancement of the cerebral lesions. Nine patients received various fractionation-dose schedules (range 8-50 Gy). Complete and partial responses were seen in 2/9 and 3/9 cases, respectively. Clinical stabilization of neurological symptoms was noted in 3/9 cases and disease progression in 1/9. The median survival times (MST) from presentation for irradiated and non-irradiated patients were 9.3 and 2.1 weeks, respectively (range 0.9-43.1). Although patient selection introduced bias, there appears to be a modest improvement in MST for treated patients. The MST with radiotherapy alone remains poor, but radiotherapy may provide palliation. For some selected patients, a prolonged response is possible. PMID- 10849982 TI - Radiosurgery at the Royal Adelaide Hospital: the first 4 1/2 years' clinical experience. AB - Radiosurgery refers to the treatment of small lesions localized by stereotactic technology using highly focused radiation. This review utilizes prospectively gathered data from the Royal Adelaide Hospital Radiosurgery unit to summarize experience with the first 62 patients (65 lesions) treated between November 1993 and May 1998. This experience included acoustic neuromas (23 patients), arteriovenous malformations (18), brain metastases (12), meningiomas (6), and glomus tumour, subependymoma, dural arteriovenous fistula (1 each). Although follow up is relatively short, the outcome in terms of morbidity and tumour control is thus far comparable with results reported in the literature. Radiosurgery provides a viable alternative to neurosurgery and conventional external beam radiotherapy for several benign and malignant intracranial lesions. PMID- 10849983 TI - Armrest versus vacuum bag immobilization in the treatment of breast cancer by radiation therapy: a randomized comparison. AB - Thirty patients in randomized sequence were simulated in both an arm-rest and a vacuum bag immobilization device and also randomized to treatment in one of the two devices. Overall patient comfort significantly favoured the use of the arm rest, although both were acceptable. Lung exposed in the tangential beams was less in the vacuum bag device. Supraclavicular skin folding was greater in the vacuum bag, but this is probably related to the angle of arm abduction. Treatment times and stability of the setups were not significantly different. This is a small study piloting a technical comparison and the findings require confirmation in a larger sample. PMID- 10849984 TI - Angiomatosis of the hand demonstrated by contrast-enhanced magnetic resonance angiogram. AB - Contrast-enhanced digital subtraction dynamic MR angiography has been applied to lower limb arteries, with accuracy comparable to that of conventional angiography. Application of this technique to hands has not been reported. A 2 year-old girl with a diffuse haemangioma of the right hand elegantly demonstrated by MR angiogram is presented. The feeding vessel and the relationship with surrounding palmar blood vessels were clearly identified. This provided essential preoperative information facilitating surgical resection, yet without the technical difficulties and morbidity associated with conventional angiography. PMID- 10849985 TI - Spinal cord Neurobehcet's disease detected on magnetic resonance imaging. AB - Imaging studies in Neurobehcet's disease have to date focused on the brain, with only four previous case reports of documented spinal cord involvement on MRI being published. A fifth case is documented here, together with a review of the literature. PMID- 10849986 TI - Florid computed tomographic appearance of acute Campylobacter enterocolitis. AB - A 28-year-old male presented with severe abdominal pain and bloody diarrhoea. Computed tomographic scan showed marked swelling of the distal ileum and entire colorectum. The patient recovered and Campylobacter jejuni was subsequently grown from his faeces. PMID- 10849987 TI - Candida jejunitis: a rare cause of intestinal pneumatosis in the immunocompromised patient. AB - A case of fatal necrotizing jejunitis caused by Candida albicans is described in a patient with acute myeloid leukaemia undergoing chemotherapy. The diagnosis was made at autopsy. Computed tomography findings were of small bowel dilatation with pneumatosis. PMID- 10849988 TI - Atrial myxoma demonstrated by cine gradient refocused echo magnetic resonance imaging. AB - A case of atrial myxoma presenting with syncope evaluated by echocardiography and MRI is described. Cine gradient-echo MRI demonstrated atrial myxoma as a very low signal intensity mass indicating the presence of haemosiderin. PMID- 10849989 TI - Segmental infarction of the omentum secondary to torsion: ultrasound and computed tomography diagnosis. AB - Segmental infarction of the omentum is a rare clinical entity that is seldom considered in the differential diagnosis for acute abdominal pain, especially as the clinical findings are so non-specific. Consequently, the diagnosis is usually made intraoperatively. The two cases presented here demonstrate the characteristic appearance of omental infarction on ultrasound and CT, which enables preoperative diagnosis. Preoperative radiological diagnosis may prevent unnecessary surgery. PMID- 10849990 TI - Giant spinal arachnoid cysts: computed tomography, magnetic resonance imaging and magnetic resonance myelography correlation. AB - Two cases of giant spinal arachnoid cysts, one of them being symptomatic, are discussed, along with their CT, MRI and MR myelography findings. PMID- 10849991 TI - Trans-sphenoidal and sphenoethmoidal encephalocele: report of two cases and review of the literature. AB - Basal encephaloceles are the least common form of encephalocele. Due to the critical position of the bony defect, visual and endocrinological abnormalities are frequently associated with basal encephaloceles. There is significant confusion in the classification of basal encephaloceles, particularly among trans sphenoidal, sphenoethmoidal and intrasphenoidal subtypes. Two cases of basal encephaloceles are presented (one trans-sphenoidal and one sphenoethmoidal), along with a review of the literature. The relationship between the basal encephaloceles and endocrinological abnormalities is also emphasized. PMID- 10849992 TI - Rhombencephalosynapsis. AB - A case report of rhombencephalosynapsis in a 34-year-old female is presented and the clinical features and possible pathogenesis of this disorder are reviewed. PMID- 10849993 TI - Adenocarcinoma in an ileal duplication cyst: ultrasound and computed tomography findings. AB - A case of adenocarcinoma in an ileal duplication cyst is presented. The pathological and radiological findings are described. A review of the literature on this rare condition is discussed. PMID- 10849994 TI - Appendiceal rupture during pelvic ultrasound examination. AB - A case of rupture of the appendix during pelvic ultrasound is presented. PMID- 10849995 TI - Primary lymphoma of the breast. AB - Primary lymphoma of the breast is a rare finding. Two cases and a review of the literature are presented. PMID- 10849996 TI - Gastrostomy: an improved technique. AB - Gastrostomy is a commonly performed procedure. The forceps described in this article help make this procedure easier and safer to carry out, reducing the need for complex and costly equipment. The forceps used have been adapted from standard surgical steel instruments by soldering a guidance device to one jaw. This device enables guided placement of a needle alongside the forceps tip. Any hospital technical service department could adapt standard forceps for this procedure. PMID- 10849997 TI - Percutaneous transhepatic changing of an endoscopically placed stent. AB - Biliary stents are commonly positioned for the relief of obstructive jaundice secondary to malignancy. It is occasionally necessary to percutaneously replace a biliary stent placed by the endoscopist. This is usually because there has been progression of the tumour, which has prevented further endoscopic access. The technique described has real merit since it can further extend palliation. PMID- 10849998 TI - Chemical induction of the ataxia-telangiectasia gene. PMID- 10849999 TI - Stratospheric ozone depletion: UVB "effects", the neglected aspect. PMID- 10850000 TI - Ultraviolet-B radiation and stratospheric ozone loss: potential impacts on human health in the arctic. PMID- 10850001 TI - Surface UV retrieval from satellite data. AB - Irradiance of biologically effective ultraviolet radiation on the Earth's surface is retrieved from satellite measurements using radiative transfer models. Retrieval methods are reviewed with a special emphasis on questions pertinent to the Arctic environment. In general, the main problem is the high variability of cloudiness. The most prominent features of high latitudes are the low Sun and the high UV reflectivity of snow-covered terrain. Being white like clouds, snow is a challenge for satellite imagery. The dependence of UV albedo on the properties of snow and ambient conditions is discussed. The use of auxiliary snow cover data from a meteorological model is planned. PMID- 10850002 TI - Penetration of UV radiation into Finnish lakes with different characteristics. AB - Penetration of UV radiation into Finnish lakes was evaluated through field measurements and/or by determining the optical properties of the lake water. The spectral measurements of air and underwater irradiance (280-400 nm) were performed in early September, 1998, in four lakes (Lake Konnevesi, Tuomiojarvi, Jyvasjarvi, and Kopru) representing a variety of humic (DOC 8.1-16.0 mg l-1) and chlorophyll (2.3-14 micrograms l-1) concentrations. A linear relationship was found between vertical attenuation coefficient (Kd) and DOC (r2 > 0.75), whereas no relationship between Kd and chlorophyll a was observed (r2 < 0.20). The depth where the UV-B irradiance is 10% of the value just beneath the surface (z10%) ranged 5-19 cm. Kd based on field measurements, and absorption coefficient (ad) derived from spectrophotometric scanning measurements of filtered lake water showed a linear relationship (r2 = 0.93). Kd values for two lakes where field measurements were not made (DOC 4.2-5.2 mg l-1, chlorophyll a 1.7-1.9 micrograms l-1) were obtained from the relationship between Kd and ad, giving the 10% UV-B penetration depth of 1.3 m to Lake Kuorinka and 0.21-0.24 m to Lake Palosjarvi. PMID- 10850003 TI - UV-B radiation and insect herbivory: preliminary observations on strawberry plants and the leaf beetle Galerucella sagittariae in Finland. AB - Enhanced UV-B (280-315 nm) radiation, as a result of stratospheric ozone depletion is known to increase UV-protective phenolics, especially flavonoids in plant tissues. We tested the hypothesis that the suitability of plant foliage to insect herbivores might be reduced due to higher amount of defence compounds produced by the host plant when exposed to enhanced UV-B radiation. Strawberry plants were exposed to supplementary UV-B and UV-A radiation in an open-field exposure system. The test animal was the leaf beetle (Galerucella sagittariae, Coleoptera, Chrysomelidae), which is known to damage strawberry leaves both in larval and adult stage. The tentative first-year results reported in this paper showed slight, but not statistically significant support for this hypothesis. PMID- 10850004 TI - Vitamin D in an ecological context. AB - Although numerous investigations have been carried out concerning the occurrence of vitamin D (D2 and D3) and their provitamins in different foodstuffs, about the effects of vitamin D intake on the human body as well as the cellular effects of the physiologically active form of vitamin D, there are almost no studies on vitamin D in an ecological context. One source for vitamin D is fish. But fish cannot synthesize vitamin D, nor provitamin D. Both originate at the beginning of the food chain, in phytoplankton. It is likely that the conversion of provitamin D (D2 and D3) to vitamin D can take place only under the influence of ultraviolet B radiation in algae. Therefore the vitamin/provitamin ratio can perhaps be used as a much needed internal "exposure meter" for UV-B radiation. As is well known, provitamin D3 (7-dehydrocholesterol) can be converted to vitamin D3 also in human skin under the influence of ultraviolet-B radiation. Little is known about which species or groups of planktonic algae are capable of vitamin D synthesis, since only natural mixtures of algae and a few defined species have been analyzed. We have observed that reindeer lichen contains vitamin D2 and D3. For plant scientists vitamin D is interesting also because it is synthesized by some (but not all) higher plants, and acts as a growth substance in plants. Provitamin D2 (ergosterol) is synthesized by many fungi, and this fact may explain some traits of plant-fungal symbiosis. PMID- 10850005 TI - Effects of seasonal photoperiod on serum 25-hydroxycholecalciferol and calcium in reindeer, Rangifer tarandus tarandus. AB - In order to understand the effects of solar irradiance on calcium metabolism we measured serum 25-hydroxycholecalciferol and total calcium levels monthly in 6 female nonpregnant reindeer maintained in the Oulu area (65 degrees N). Mean monthly serum total calcium levels varied slightly and the highest levels were seen in October. Serum 25-hydroxycholecalciferol levels were highest in October but the monthly variation was also slight. The small monthly variation of the analytes' abundance and the fact that they peaked not until 4 months after the longest day, (i.e. the main stimulus for vitamin D synthesis) could have something to do with the supplementation of this vitamin in the diet of the captive reindeer. PMID- 10850006 TI - Risks, especially for the eye, emanating from the rise of solar UV-radiation in the Arctic and Antarctic regions. AB - Physical and biological characteristics of solar UV-radiation wavebands A, B, and C are explained and information is provided on UV-levels in particular environments and ocular tissues. The question whether or not the rise in circumpolar UV of the last 20 years or so can be regarded as a threat is briefly addressed and it is concluded that even if no threat to photosynthetic productivity of crops and vegetation exists, there is a danger regarding the status of health of human skin and eyes (in particular the lens). The nature of the UV-induced damage to cornea, lens, and even the retina with its photo receptive cells and pigment epithelium is assessed and a word of caution is sounded with regard to possible injury-potentiating effects of certain chemicals as seen, for instance, in the recent and alarming rise of cataract in Scottish salmon. Finally, because of the multifaceted effects of UV (e.g. at molecular, cellular, tissue, individual, population, and ecosystem level), a plea is made for a concerted, well-funded, international effort to tackle the many remaining problems at all fronts and from all possible angles. PMID- 10850007 TI - Melanoma and other skin cancers in circumpolar areas. AB - During the recent decades, the thickness of the ozone layer over the northern hemisphere has declined by 10 to 40 percent during the winter and spring months. Since ozone is the major barrier protecting the earth from dangerous short wave UV-radiation (UVB), the depletion in the ozone layer consequently increases the amount of UV-radiation reaching the earth's surface. As a rule a 10 percent reduction in the ozone layer causes ca. 20% increase in UV-radiation and a 40% increase in skin cancers. Thus relatively minor changes in ozone layer thickness may a have marked impact on the health of humans. Skin cancer is the most common cancer in humans, i.e. in Finland about 4000 new basal cell carcinomas, 700 other skin cancers, mostly spinous cell carcinomas and 500 melanomas occur yearly. Up to recent years the incidence of skin cancers has steadily increased in northern countries. As an explanation, changes in sunbathing habits have been suggested to play a central role. Due to the high mortality rate in melanoma, and marked morbidity in other skin cancers, it is important to try to prevent skin cancers and inform the public about the risks of excessive sun exposure, and of the ways in which the skin can be protected. Proper clothing and use of sunscreens have been shown to reduce the incidence of both melanomas and other skin cancers. Furthermore, it is important to identify those at high risk for acquiring skin cancers, like individuals with type 1 skin character (fair skin which burns easily), or numerous dysplastic nevi, or a family history of skin cancers. PMID- 10850008 TI - Linkage of Cholestasis Familiaris Groenlandica/Byler-like disease to chromosome 18. AB - In East Greenland (Tasiilaq) a common recessive disease, Cholestasis Familiaris Groenlandica (CFG)/Byler-like disease, occurs in Eskimo children. Samples from 123 persons, from a large consanguineous pedigree in East Greenland including 7 affected and 3 small families from West Greenland with a total of 4 affected children, have been collected for linkage and homozygosity studies. An earlier hint of linkage to chromosome 18q (lod score of 1.5 to blood group JK) is now raised to a multipoint lod score of Z = 3.25 in the area of the DNA markers D18S851 and D18S858. Different haplotypes follow the disease gene among Inuits in West Greenland and a possibility of locus heterogeneity of CFG between East and West Greenland exist. PMID- 10850009 TI - Sleep and mood during a winter in Antarctica. AB - Seasonal variations in sleep characteristics and their association with changes in mood were examined in 91 American men and women who spent the 1991 austral winter at three different research stations in Antarctica. Measures of total hours of sleep over a 24-hr period, duration of longest (i.e., "nighttime") sleep event, number of sleep events, time of sleep onset, and quality of sleep remained unchanged over the course of the austral winter (March through October). However, exposure to total darkness based on station latitude was significantly associated with total hours of sleep, duration of longest sleep event, time of sleep onset, and quality of sleep. Reported vigor the previous month was a significant independent predictor of changes in all five sleep measures; previous month's measures of all six POMS subscales were significant independent predictors of sleep quality. Sleep characteristics were significant independent predictors of vigor and confusion the following month; total sleep, longest sleep event, sleep onset and sleep quality were significant independent predictors of tension anxiety and depression. Changes in mood during the austral winter are preceded by changes in sleep characteristics, but prolonged exposure to the photo-periodicity characteristic of the high latitudes appears to be associated with improved sleep. In turn, mood changes appear to affect certain sleep characteristics, especially sleep quality. PMID- 10850010 TI - A prevalence methodology for mental illness and developmental disorders in rural and frontier settings. AB - A prevalence study methodology developed for use in rural and frontier settings is described. The general method was developed over a 15 year period and has been successfully adapted and used in studies of 14 different childhood onset developmental disorders. Subjects were the 168,000 school aged children from North Dakota who were first surveyed for cases of autism--pervasive developmental disorders in 1985 and 1986. The results of the prevalence study were compared with the results of a 12-year ongoing surveillance of the cohort. The 12-year ongoing surveillance identified one case missed by the original prevalence study. Thus the original prevalence study methodology identified 98% of the cases of autism-pervasive developmental disorder in the population. This methodology may also be useful for studies of other developmental disorders in rural and frontier settings. PMID- 10850011 TI - Osteoporosis issue needs addition of extension exercise findings. PMID- 10850012 TI - Avoid threat of malpractice suits. PMID- 10850013 TI - Opportunities for the osteopathic medical profession to pursue worldwide acclaim and recognition. PMID- 10850014 TI - Progressive inhibition of neuromuscular structures (PINS) technique. AB - Progressive inhibition of neuromuscular structures (PINS) is a technique that can be included in the osteopathic manipulative treatment repertoire. It relies on knowledge of anatomy and neuromuscular physiologic features as well as on standard forms of osteopathic palpatory diagnosis and treatment. It is a variant of the inhibition technique that has been taught as an osteopathic manipulative technique for many years, and it bears some resemblance to other manual medicine techniques. The emphasis of the approach is the determination of the alteration of the tissues due to dysfunction, delivering treatment based on palpatory evaluation and patient feedback. Two related points are initially chosen, followed by a progression from one to the other. Relationships to similar techniques are also discussed. Theoretical as well as selected practical applications are presented. PMID- 10850015 TI - Microsurgical reconstruction of basal cell carcinoma defect of the face: a multidisciplinary approach. AB - This article describes a 73-year-old white man with a history of dizziness secondary to profound anemia who presented with a large basal cell carcinoma of the left front temple region. A multidisciplinary approach to the extirpation and reconstruction of this defect is presented with a review of histopathologic features and outcomes of basal cell carcinoma excision. PMID- 10850016 TI - Increasing osteopathic manipulative treatment skills and confidence through mastery learning. AB - Several recent studies document the declining use of osteopathic manipulative treatment (OMT) in clinical practice. In this article, the authors contend that developing new teaching materials based on the mastery learning approach can augment time-tested methods of teaching OMT and help to stop or reverse this decline. The Spencer technique for shoulder manipulation is used to demonstrate the development and evaluation of OMT mastery learning materials. These materials could be developed as part of a progressive teaching sequence requiring increasing diagnostic acumen, palpatory skill, and therapeutic subtlety. Such a program could be used throughout osteopathic medical training and for continuing medical education to increase skills and confidence in the use of OMT. PMID- 10850017 TI - The treatment of influenza. 1918. PMID- 10850018 TI - Osteopathic success in the treatment of influenza and pneumonia. 1919. PMID- 10850019 TI - One hundred thousand cases of influenza with a death rate of one-fortieth of that officially reported under conventional medical treatment. 1919. PMID- 10850020 TI - Influenza and its osteopathic management. 1937. PMID- 10850021 TI - Asthma. PMID- 10850022 TI - Atopic dermatitis. PMID- 10850023 TI - Contact dermatitis. PMID- 10850024 TI - Current concepts in allergen immunotherapy. PMID- 10850025 TI - Evaluation and management of allergic rhinitis. PMID- 10850026 TI - Food allergy. PMID- 10850027 TI - Immunodeficiency problems in children. PMID- 10850028 TI - The role of allergy in the development of otitis media and sinusitis. PMID- 10850029 TI - Urticaria and angioedema. PMID- 10850030 TI - Stinging insect hypersensitivity. PMID- 10850031 TI - Allergy and immunology. PMID- 10850032 TI - [The journey from genome structure to cure of diseases is long and winding. Research specialists in molecular biology and physiology should cooperate]. PMID- 10850033 TI - [Ventilation in ARDS: respirator, prone position, NO or artificial lung]. AB - This review discusses the treatment of impaired gas exchange in acute respiratory distress syndrome (ARDS) using conventional ventilation, the open lung approach, prone position, nitric oxide (NO) inhalation and extracorporeal membrane oxygenation (ECMO). It is concluded that ventilation with high inspiratory pressures or volumes should be avoided, and that the open lung approach should be used as the first step. If this does not lead to satisfactory results, prone positioning is recommended, and if life-threatening hypoxemia persists, ECMO could be considered. NO inhalation is not recommended. PMID- 10850034 TI - [47-percent 6-months-long survival for intensive care patients over 80]. AB - All 112 patients aged 80 and above treated at the intensive care unit at the University Hospital in Lund, Sweden 1994-1995 were followed-up retrospectively in terms of six-month survival (SMS) and for survivors in terms of quality of life. Overall SMS was the same for both men and women--47%. Patients with the poorest SMS were those aged 90 and above with only one patient out of eleven surviving six months. Patients admitted for severe heart failure also showed a very poor outcome with SMS 27%. Patients were grouped in terms of living conditions prior to admission to the ICU, and a significant difference in six-month survival was noted between those living in their own homes (53%) prior to admission compared to those coming from a nursing home (25%). Patients surviving six months were interviewed by telephone regarding their living situation in March 1997. More than 50% of survivors were living in their own homes with external help no more than once a day. The average APACHE II score was 14.9 +/- 8.2. The average score for patients surviving six months was 13.4 +/- 5.9 and for those not surviving six months 16.8 +/- 5.1. No significant statistical difference in APACHE II scores between these two groups was shown. PMID- 10850035 TI - [Tracheal cannules adapted for special use are manifactured at a respiratory clinic]. PMID- 10850036 TI - [Alcohol drinking habits assessed by the AUDIT test. Reduced maximum levels doubled the number of women with dangerous alcohol drinking]. AB - The Alcohol Use Disorders Identification Test (AUDIT) was completed by 997 persons randomly sampled from the general Swedish population (80 percent response rate). Eighteen percent of the men and 5 percent of the women had hazardous or harmful alcohol use according to the 8+ score criterion. Since women are more sensitive to alcohol than men, a cut-off score of 6+ was suggested for them. The female prevalence of hazardous or harmful alcohol use then increased to nearly 11 percent. Hazardous or harmful alcohol use decreased with increasing age in both genders. The "binge drinking" question explained half of the total AUDIT variance and is thus the best predictor of heavy drinking in the test. The internal and test-retest reliability of the AUDIT was satisfactory. Reference values for different ages and genders are presented. PMID- 10850037 TI - [Computerized medical record--a relic from the last century]. PMID- 10850038 TI - [Problems and possibilities of electronic publishing. It is the responsibility of the research community to carry the development into the right direction]. PMID- 10850040 TI - [A moment in medicine, part 6. "It's not possible to become a good physician without true compassion"]. PMID- 10850039 TI - [Following HUGO. Protein survey will be the real challenge]. PMID- 10850041 TI - [Current medicine requires cooperation--not omnipotence]. PMID- 10850042 TI - [The good will of experts and the free choice of patients. A quality project in connection with the increased number of applications from psychiatric care concerning the support patients are entitled to by the law]. PMID- 10850043 TI - [The power of the word and meaning: "Empowerment"=patient participation?]. PMID- 10850044 TI - [ASCUS, CIN I and vaginal cytological diagnosis--a comment]. PMID- 10850045 TI - [Unsuccessful outcome of a surgical intervention shows the importance of a good knowledge of anatomy]. PMID- 10850046 TI - [New reports on estrogen therapy. Increased risk of breast cancer; no effects on Alzheimer disease]. PMID- 10850047 TI - [Methadone treatment during pregnancy and its effect on the child. Better than continuing drug abuse, should be monitored by a specialized antenatal care center]. AB - Observations concerning methadone treatment in 21 pregnancies of drug addicted women in Stockholm between 1982 and 1997 are presented. Fourteen per cent of new born infants were small for gestational age, with head circumference less than--2 SD. None had congenital malformations, but neonatal abstinence occurred frequently. Mortality due to sudden infant death syndrome was also high. Five infants with severe neonatal abstinence were successfully treated with tinctura opii. It is essential that antenatal care for drug addicted pregnant women and subsequent care for their infants are concentrated at units in which a midwife, an obstetrician and a pediatrician with specific competence in drug abuse during pregnancy cooperate. PMID- 10850048 TI - [Don't forget the risk of fetal abnormalities caused by alcohol use during pregnancy! Panorama of the injuries, mechanisms, prevention, therapeutic principles]. PMID- 10850049 TI - [Specialized maternal health service for drug-abuser parents-to-be: long-term support prevents recurrence and creates a better environment for the growing child]. PMID- 10850050 TI - [Drug addicts with severe mental disorders can be helped by programs using moderate means. Good results when psychiatric, social and drug abuse services cooperate]. AB - Dependency disorders are more common than expected in psychiatric populations. Untreated, dual diagnosis leads to severe social and psychiatric deterioration. Nine treatment resistant, homeless, drug addicts suffering from chronic psychotic disorders were selected to take part in a case management program, integrating social services with regular psychiatric treatment. All but one were greatly improved in general terms as well as regarding their ability to maintain an ordered life style. The need for institutional care decreased dramatically. PMID- 10850051 TI - [Five new antiepileptic agents approved during the 1990's--an observation study on the utilization of the new preparations in routine clinical practice]. AB - Since 1990, five new antiepileptic drugs have been approved in Sweden for add-on therapy of partial epilepsy. The optimal use of these drugs has not yet been established. In a county general hospital, 75 of 382 adult patients with epilepsy were treated with newer antiepileptic drugs. Fifty-two continued treatment with a newer drug for one year or longer mainly because of improved seizure control. The newer drugs represented 18 per cent of total sales of antiepileptic drugs in the area served by the hospital but the corresponding cost was 70 per cent. Despite their higher price, use of the newer drugs seems justifiable when significant improvement of seizure control can be achieved. PMID- 10850052 TI - [Guidelines for management of patients with acute pancreatitis]. AB - During recent years new concepts and methods have been introduced in the management of acute pancreatitis. Severity and risk of complications show wide variation. Outcome is also dependent on the physician's experience and on his local resources. In this light the Swedish Society of Upper Abdominal Surgery has elaborated national guidelines for management. Attention is paid to diagnosis, severity assessment and etiology. Furthermore, guidelines are offered for treatment of mild and severe pancreatitis, as well as for the management of pseudocysts. The role of multidisciplinary intensive care specialist teams in the management of severe disease is emphasized. The guidelines are supported by the Swedish Society of Gastroenterology, the Swedish Society of Gastroenterology, the Swedish Society of Anesthesiology and Intensive Care and by experts from other Nordic countries. PMID- 10850053 TI - [Expert meeting on the effect of sex hormones on the normal breast: moderately increased risk of cancer, but the connection is still not clear]. PMID- 10850054 TI - [Lakartidningen should declare conflict of interest in medical articles]. PMID- 10850055 TI - [Consultation psychiatry in Sweden--time to train the psychiatrists?]. PMID- 10850056 TI - [Time, medical records and quality assurance--the cost-benefit assessment is a must]. PMID- 10850058 TI - [Inadequate comments on the euthanasia debate]. PMID- 10850057 TI - [Incorrectly administered cytostatic agents?]. PMID- 10850059 TI - [Physicians aged 65 and over and the national health insurance]. PMID- 10850060 TI - [Functional genomics--a future method for development of drugs?]. PMID- 10850061 TI - [Medizinische Klinik in the future online in LINK]. PMID- 10850062 TI - [German Society for Internal Medicine at the turn of the millenium: review and perspectives]. PMID- 10850063 TI - [Variability of blood pressure and baroreceptor function; clinical and scientific relevance]. AB - The human arterial blood pressure shows not only exogenic alterations, but also spontaneous fluctuations. This blood pressure variability has both scientific and clinical relevance. A well-known phenomenon is the so-called "white coat hypertension" which may severely affect diagnosis and therapy of essential hypertension. Furthermore, several studies have provided evidence that the degree of end-organ damage in hypertensive patients is closely related to the circadian blood pressure variability. Therefore, it is tempting to suggest that an optimal antihypertensive treatment should take an improvement of the blood pressure variability into consideration. Among other mechanisms modulating the arterial blood pressure, the baroreflex function has to be mentioned. Diseases like chronic renal failure, treatment with drugs like ciclosporine A and also smoking have been shown to reduce the baroreflex sensitivity. As the baroreflex sensitivity mainly reflects interactions between the blood pressure variability on the one hand and the heart rate variability on the other hand, it is not surprising that also a reduced baroreflex function is an independent cardiovascular risk factor. PMID- 10850064 TI - [Long-term problems after kidney transplantations]. AB - Allogenic kidney transplantation is a widely established treatment option for patients with end-stage renal disease to gain independency from dialysis and recovery of excretory and hormonal functions. Transplantation is superior to dialysis with respect to quality of life, morbidity and mortality. Improvements in one-year graft survival have become evident during the last decades, but long term outcome is not yet satisfying. The most common form of late transplant failure is chronic rejection or chronic allograft nephropathy, and as a consequence of higher ages of transplant recipients death of patients with functioning grafts. The histological hallmark of chronic rejection is intimal thickening of small arteries and arterioles, but the involved pathomechanisms are not well understood. Strict normalization of blood pressure and hyperlipidemia by drugs aims to prevent vascular alterations of chronic allograft nephropathy. In addition, in this overview some other late complications of renal transplant recipients are addressed which are associated to chronic immunosuppression, such as viral or malignant diseases. PMID- 10850065 TI - [Alterations of arterial vessel wall properties in hyperparathyroidism]. AB - Alterations of arterial vessel wall function are associated with increased cardiovascular morbidity and can be measured by noninvasive ultrasound techniques. There is increasing evidence that hyperparathyroidism contributes not only to structural changes of the cardiovascular system but to disturbances in functional arterial vessel wall properties. It has been shown that elevated concentrations of parathyroid hormone are associated with decreased arterial distensibility and increased intima-media thickness in patients with renal insufficiency. A permissive role of parathyroid hormone in the pathogenesis of uremic vessel wall changes has been discussed. The endothelium is a newly recognized target organ of parathyroid hormone. In patients with primary hyperparathyroidism endothelial dysfunction has been demonstrated, that seems to be reversible after parathyroidectomy. Effects on arterial vessel wall function may contribute to increased blood pressure observed in hyperparathyroidism. The complex cardiovascular effects of parathyroid hormone and changes in vascular tone, arterial distensibility and endothelial function in hyperparathyroidism are discussed. PMID- 10850066 TI - [Extracorporeal renal replacement therapies in acute renal failure]. AB - The most serious forms of acute renal failure (ARF) are nowadays encountered in the intensive care unit (ICU), where up to 25% of new patients are reported to develop ARF. Lethality rates may reach 50 to 90% when the ARF is part of a multiple organ dysfunction syndrome. A multitude of extracorporeal procedures have been introduced into intensive care medicine. Applied with adequate skills and experience, most of these techniques will suffice to replace excretory renal function. However, because of low efficacy arterio-venous procedures (CAVH and CAVHD) have been abandoned for the veno-venous, pump-driven techniques (CVVH and CVVHD). Up to now, there is no consensus whether continuous or intermittent renal replacement therapy is more advantageous. In many cases, oliguric patients with circulatory instability will be treated by CVVH, even though there is no prospective study to show that in terms of outcome continuous treatment is superior to intermittent hemodialysis. It is equally conceivable to treat such patients with daily, prolonged (intermittent) hemodialysis. Apparently, the dose of replacement therapy, be it continuous filtration (36 to 48 l/24 h) or intermittent hemodialysis (daily 3 to 4 h) with a target BUN of less than 50 mg/dl, is more important than the modality of treatment. Moreover, there is good evidence that the use of biocompatible membranes (no complement- or leukocyte activation) is preferable and that with high-volume hemofiltration bicarbonate containing replacement fluids should be used. However, despite all the technical advances, we firmly believe that the skills and the experience of those physicians and nurses who actually perform renal replacement therapy in the ICU are more important than the modality of treatment applied. PMID- 10850067 TI - [Alterations of the arterial vessel wall in renal failure]. AB - Cardiovascular events are the main cause of death in patients with end-stage renal disease. Functional and structural alterations of the arterial system substantially contribute to the high cardiovascular mortality in these patients. Structural alterations of the arterial wall comprise intima-media thickening and atherosclerotic plaque formation. Moreover, mechanical vessel wall properties of large arteries are significantly disturbed. This is already observed in young patients. Reduced arterial distensibility impairs large artery cushioning function. This results in increased ventricular afterload promoting left ventricular hypertrophy and in reduced coronary perfusion. After renal transplantation, structural alterations of the arterial wall and disturbed mechanical vessel wall properties persist. Moreover, renal failure is characterized by a severe impairment of endothelial function. Disturbed endothelial function results from reduced production of endothelium-dependent endogenous vasodilators and/or blunted vascular effects of these substances. Uremia is associated with the accumulation of an endogenous inhibitor of the endothelial nitric oxide synthase. Impaired endothelial function in renal failure promotes the progression of structural lesions of the arterial wall. Also in renal transplant recipients, substantially impaired endothelial function is observed despite correction of uremia. Hyperparathyroidism commonly observed in renal failure contributes to the disturbed functional vessel wall properties of large arteries. In patients with end-stage renal disease, decreased large artery distensibility is an independent risk factor for increased cardiovascular mortality. This may apply also to intima-media thickening and to impaired endothelial function. PMID- 10850068 TI - [Therapy of hypertensive crises]. AB - Hypertensive crisis is defined as an extreme elevation of arterial blood pressure, with diastolic pressure > 120 mm Hg, and represents an imminent risk to the patient. In such cases, a rapid orientating diagnosis and adequate antihypertensive treatment to avoid sequelae are needed, sometimes even before diagnostic tests are completed. Hypertensive emergencies and hypertensive urgencies can be distinguished. If the critical increase in blood pressure is associated with end-organ damage such as encephalopathy, acute left heart failure and pulmonary edema, angina pectoris, myocardial infarction or dissecting aortic aneurysm, a hypertensive emergency is present, that is an acute threat to the patient's life. A hypertensive emergency requires effective lowering of blood pressure within minutes, but not necessarily to normal range. The choice of suitable antihypertensive agents depends on clinical symptoms, contraindications, duration of pressure elevation and underlying conditions, prior cardiovascular, cerebrovascular and renal disorders. The risk of imminent end-organ damage must be weighed against the risk of rapid blood pressure lowering. In hypertensive urgencies without end-organ complications, blood pressure can be lowered more slowly over several hours, often with oral agents to avoid detrimental fall in blood pressure. The drugs of choice are mainly urapidil i.v. and nitroglycerine. PMID- 10850069 TI - [Diagnosis and therapy of renovascular hypertension]. AB - BACKGROUND: Renovascular hypertension as a secondary form of hypertension can be improved or cured in many cases by interventional radiology or vascular surgery. BASIS: The renin-angiotensin-aldosterone system and the development of hypertension are linked in renovascular hypertension. CLINICAL APPEARANCE: Early clinical symptoms are of special interest in diagnosing renovascular hypertension. DIAGNOSIS AND THERAPY: Nowadays, angioplasty or stenting have mostly replaced surgery in view of treatment of renovascular hypertension. CONCLUSION: Renovascular hypertension, if diagnosed early, can be improved or cured in many cases regarding hypertension and/or renal insufficiency. PMID- 10850070 TI - [Cancer incidence noticeably on the rise. Are we prepared?]. PMID- 10850071 TI - [Fear of opioids by injection and tablets. Patients prefer the patch]. PMID- 10850072 TI - [Deficits in oncology? We're not so bad!. Interview by Christine Vetter]. PMID- 10850073 TI - [Ten million patients with thyroid gland nodules. Excluding a cancer diagnosis]. PMID- 10850074 TI - [Drugs for back pain: caution, liver damage!]. PMID- 10850075 TI - [Improved methods to help drug dependent patients. Drugs against dependence?. Interview by Petra Eiden]. PMID- 10850077 TI - [Drug therapy for pregnant patients. A case for two]. PMID- 10850076 TI - [Drugs tolerated by the fetus. Which antibiotics and antipyretics need not be avoided in pregnancy]. PMID- 10850078 TI - [Internet addresses with specific emphasis. TORCH--risk for the baby]. PMID- 10850079 TI - [When the voice fails. Organic causes for dysphonia]. PMID- 10850080 TI - [Enhanced imaging diagnosis of liver carcinomas. Technical progress is not a substitute for clinical knowledge]. AB - Cholangiocellular carcinomas (CCCs) of the peripheral type (not Klatskin tumors) appear sonographically as solid space-occupying lesions. No typical sonomorphological signs are known. Important hint for the clinician is the knowledge of earlier or accompanying diseases in which CCCs occur more often (primary sclerosing cholangitis, ulcerative colitis, intrahepatic gallstones, parasitic diseases of the bile ducts, choledochal cysts, Caroli's syndrome). Typical Color-Doppler signs are also missing. Definitive diagnostic is left to biopsy. Hepatocellular carcinomas (HCCs) mostly occur in cirrhotic livers. Every newly detected space-occupying lesion in a cirrhotic liver is suspected of HCC until otherwise proven, even when AFP-levels are normal. AFP levels constantly higher than 400 ng/ml make HCC probable, AFP levels of 2000 ng/ml ore more proof HCC. Gray-scale imaging is not uniform and not guiding. Using Color-Doppler nearly all HCCs show markedly hyperperfusion compared to the surrounding liver tissue, vessels are arranged in a chaotic shape. Echo-enhanced Color-Doppler always shows a distinct early arterial hyperperfusion. PMID- 10850081 TI - [Travel pharmacy for the physician. 1: On the road in Europe, North America and Japan]. PMID- 10850082 TI - [Diagnostic quiz. First diarrhea, then dyspnea. Splenic abscess as a complication of systemic salmonellosis]. PMID- 10850083 TI - [Women physicians in Germany. Numerous only in unemployment]. PMID- 10850084 TI - [New antidiabetic in "virtual reality". Eye to eye with insulin resistance]. PMID- 10850085 TI - [Tumor therapy. New concepts for advanced cases]. PMID- 10850086 TI - [Osteoporosis in women. Fewer falls with minerals plus vitamin D]. PMID- 10850087 TI - [Diagnosis and therapy of anal complaints. When can you do yourself and when to consult the surgeon?]. PMID- 10850088 TI - [Teaching based on the case of Lassa fever. What ails our tropical medicine. Interview by Roland Knauer]. PMID- 10850089 TI - [Reduce the risk! Healthy children for epileptic patients]. PMID- 10850090 TI - [Pancreatic carcinoma: the laborious way toward a better prognosis. Current status of therapy and after-care]. PMID- 10850091 TI - [Second opinion in cancer. Not causing conflict with the primary physician!]. PMID- 10850092 TI - [Second opinion in tumor surgery. Valuable--but needing improvement]. AB - In complicated oncological cases, a second opinion is desirable, also in the view of the care-providing surgeon. It serves interdisciplinary therapeutic planning, and helps improve the quality of treatment. In the case of highly consequential interventions, the patient has a legal right to a second opinion. On a practical level, however, the implementation of this possibility encounters problems: organizational shortcomings, incomplete patient documentation, the stresses of patient transportation, loss of valuable time, in particular in the case of postal consultation, following consultation in a center the carrying out of treatment there, whether at the urging of the doctors there or the patient himself, lack of remuneration for the efforts of the consultants. Today, however, all the necessary technical facilities are in place to enable various experts to be consulted, virtually simultaneously, on any case, via video-conferencing. PMID- 10850093 TI - [Two forms of second opinion]. AB - Second Opinions are requested and given for a wide variety of problems in the health care system. It is proposed that such opinions be initially differentiated into "communicative second opinion" (CSO) and "evidence-based second opinion (EBSO). CSOs are frequently set up without a search of the literature, and more in a sense of communication ("that is what I would do too"). In the case of the EBSO, certain demands derived from the principles of evidence-based medicine are expected to be met: examination of the type of problem involved, and the validity and clinical relevance of the clinical data. Of importance is the need to define whether, (on the basis of a scoring system??) for a given problem/question, a proposal threshold (a better options is available) or an exclusion threshold (no data as a basis for an option) is trespassed. PMID- 10850094 TI - [Second opinion as expert consultation--general practitioner as referral source]. AB - The relationship between family doctor and patient is usually one of mutual trust developed over many years. This is why a patient with cancer most likely first raise the question of a second opinion with him, in particular in a "palliative situation". The family doctor, however, can properly deal with this task only if he is included as a link in the network of communication and cooperation between those engaged in the treatment and care of cancer patients. Only then can the family doctor, as the closest and most trusted medical carer who is well acquainted with the patient's overall situation, help the patient to obtain a well-founded second opinion. In so doing they can also additional benefit the patient by sparing him many a fruitless odyssey, dashed false hopes and great disappointments. PMID- 10850095 TI - [No progress without the physician in therapy of obesity]. PMID- 10850096 TI - [Implanted venous catheters and port systems]. PMID- 10850097 TI - [ACE inhibitors for elderly hypertensive patients]. PMID- 10850098 TI - [New heart valve: correctly gauged anticoagulation]. PMID- 10850099 TI - [24-hour blood pressure monitoring]. PMID- 10850100 TI - [Student with homesickness. Wilson disease]. PMID- 10850101 TI - [New findings regarding an "ancient illness"]. PMID- 10850102 TI - [Intravenous thrombolytic therapy: ideal treatment for acute ischemic stroke]. AB - In four patients with symptoms of presumed acute ischaemic stroke intravenous treatment with recombinant tissue plasminogen activator (rtPA) was considered. Two patients indeed received rtPA within 3 hours after onset of symptoms. One of them, a 55-year-old woman, recovered and was able to resume her job as a teacher four months later. The other patient, a 38-year-old man, had a severe bleeding complication that could be stopped, but the patient died several days later because of the massive stroke. The third patient, an 82-year-old woman, could not be treated with rtPA because the time of onset of neurological deficit was uncertain. Nevertheless, she recovered well from her hemiplegia after a few days. The fourth patient, a 24-year-old woman, did not receive rtPA because her symptoms were thought to be the result of a psychogenic disorder. Intravenous thrombolysis increases the risk of intracranial haemorrhage, but should be considered a useful treatment for ischaemic stroke provided there is no doubt about this diagnosis and treatment with rtPA can be started within 3 hours of onset of the neurological deficit. PMID- 10850103 TI - [Thrombolytic therapy of brain infarct: the end of beginning]. AB - Thrombolysis by intravenous application of thrombolytic drugs may improve the outcome of patients with a brain infarct, but it also entails risks. The effect of recombinant tissue plasminogen activator (rtPA) was compared with placebo in three medium-sized randomized controlled clinical trials. One study, performed in North America, showed a clear benefit of rtPA administered within 3 hours after the onset of symptoms. Two European trials showed a less strong effect, but the number of patients who were independent after 3 months' follow-up was also larger after treatment with rtPA within 6 hours. A meta-analysis of all three trials demonstrates a significant advantage of rtPA over placebo for all the usual outcome measures, without significant excess mortality in the rtPA group. The chance of being able to live independently increases by about 8% after treatment with rtPA. In conclusion there is now sufficient evidence to start with thrombolytic treatment for cerebral infarcts in hospitals with a stroke unit, if a number of additional quality standards for the acute diagnosis and treatment of stroke patients are met. PMID- 10850104 TI - [Leukodepletion of blood products: a requirement for improvement of quality and safety]. AB - The presence of leukocytes in blood products has no beneficial effect on the recipient, except in special situations such as for patients being prepared to receive an organ transplantation. On the other hand the leukocytes have a number of untoward side effects such as HLA immunisation, non haemolytic febrile transfusion reactions, virus transmission and postoperative infections. In response to a request of the Minister of Health, Welfare and Sports, the Health Council of the Netherlands prepared a recommendation on the need of routine leukodepletion by filtration of blood. Although the introduction of leukodepletion of blood products is favoured, it is emphasized that only data from selected patient groups are available while the costs of leukodepletion are considerable. Therefore, an evaluation of the benefits and cost effectiveness of blood filtration is recommended. It is argued that leukodepletion, already introduced in a number of countries, is now considered to be 'state of the art'. Furthermore product liability, public opinion about blood safety and the precaution duty of manufacturers should be taken into account. PMID- 10850105 TI - [Fifty years of plastic surgery in the Netherlands. VI. Microsurgery]. AB - Use of the operating microscope created many new possibilities in plastic, reconstructive and hand surgery. Initially most work was done in digital replantation and, somewhat later, in transfers of toes for reconstruction of amputated thumbs. Microvascular surgery, however, appeared to be a technique suitable for more applications. Anatomical research of the blood supply of skin, fascia, muscle and bone identified flaps that could be carried by pedicle vessels. Transfer of these flaps and revascularization by microvascular anastomoses of the pedicle vessels set the stage for free flaps. In a few decades microsurgical techniques in plastic surgery fully matured. With free flap surgery single-stage and complex reconstructions could be achieved leading to earlier mobilization and better restoration of function with a shorter hospital stay. Today, microvascular free tissue transfer is an essential part of plastic and reconstructive surgery. Further advances in microsurgery and free tissue transfers deserve to be mentioned: pre-fabrication of free flaps, reduction of donor site morbidity, development of artificial conduits and instrumentation and finally homologous transplantations. PMID- 10850106 TI - [Fifty years of plastic surgery in the Netherlands. VII. Hand surgery]. AB - The first and second world wars were responsible for a turbulent development of surgery of the hand. The sixties and seventies were marked by the emergence of microsurgery, growing of the knowledge of functional anatomy and use of vascularized skin flaps and joint prostheses. These days hand surgery is mainly concentrated in 'hand centres', with co-operation of a plastic surgeon, physical and occupational therapists and rehabilitation specialists. In the future much can be expected from the genetic and metabolic manipulation of congenital malformations and acquired deformities. PMID- 10850107 TI - [Bovine spongiform encephalopathy and food safety]. AB - The governments of Great Britain and France disagree on the safety of British beef with respect to bovine spongiform encephalopathy (BSE). Eventually the consumer might have the burden to decide whether beef from Britain is safe to eat with regard to the new variant of Creutzfeldt-Jakob disease (vCJD). Outside Britain the incidence of BSE increases. Probably in continental Europe, the use of high risk material, derived from non-British cattle in the subclinical stage of BSE, poses a greater threat than beef imported from Great Britain. The risk to contract vCJD depends on two unknown factors: the susceptibility of man to BSE and the amount of success in keeping infectious tissue of BSE-incubating cattle out of the human food chain. Until the susceptibility of humans to BSE and the extent to which our food is contaminated become known, no clear-cut advice can be given on the safety of certain food items. There appears to be a genetic predisposition to vCJD, i.e. methionine homozygosity at codon 129 of the normal human prion protein gene. The European Community should ban the use of any high risk material for production of food, except in production processes for which inactivation of the BSE agent has been proven. The facts that are known at this moment allow the conclusion that gelatin and beef products of unknown origin and composition pose a greater health threat than eating genuine beef (muscle tissue). PMID- 10850108 TI - [CBO guideline 'Stroke' (revision) Dutch Institute for Healthcare Improvement]. AB - The stroke consensus dating from 1991 had to be revised, because of the introduction of new developments in the treatment of patients with stroke. More than 50 representatives from 25 professions and institutions participated. Under methodological assistance of the Dutch Institute for Healthcare Improvement CBO separate working groups (diagnosis, treatment, organization of care, rehabilitation/education, implementation and cost-effectiveness) studied the literature and translated the results into recommendations with explanatory text. The strength of scientific evidence was classified. During a national public meeting the results were discussed. In the field of guideline development cost effectiveness analyses and specific attention for implementation are new. Care on a stroke unit decreases the risk of mortality, life-long disabilities, and dependence on permanent care with about 20%. Regional stroke services should be instituted, in which continuity and efficient care can be guaranteed. Very early thrombolysis with recombinant tissue plasminogen activator strongly decreases the number of patients dying, or remaining care-dependent in a selected group of appropriate patients. Secondary prevention (lifestyle measures, acetylsalicyclic acid, treatment of hypertension and hypercholesterolaemia, and surgery of the carotids) may decrease the number of residual strokes and myocardial infarctions. In the occurrence of cerebral ischaemia and atrial fibrillation oral anticoagulants are indicated. Early intensive rehabilitation increases the chance of recovery. Silent cognitive defects may hinder rehabilitation. The extensive guideline summarises the scientific literature about treatment of patients with stroke and should serve as a basis for local protocols and appointments. PMID- 10850109 TI - [Thrombolytic therapy in patients with acute brain infarction: favorable preliminary results in Maastricht]. AB - OBJECTIVE: To assess the feasibility of acute thrombolysis for ischaemic stroke in clinical practice. DESIGN: Prospective. METHOD: On July 1st, 1998 thrombolytic therapy for ischaemic stroke was introduced in the University Hospital Maastricht, the Netherlands. All patients admitted with ischaemic stroke were prospectively registered during the first year. Of all patients with ischaemic stroke, it was determined how many were potentially eligible for thrombolysis within 3 hours of stroke symptom onset, and how many of these patients were actually treated with thrombolysis. Furthermore, the reasons for exclusion from thrombolytic therapy were assessed. Several baseline and clinical patient characteristics were noted. RESULTS: During the first year 18 ischaemic stroke patients were treated with thrombolysis within 3 hours of stroke onset. These 18 patients constituted 7% of all 256 ischaemic stroke patients and 18% of the potentially eligible patients who arrived in the hospital within 3 hours. More than 40% of the ischaemic stroke patients were not eligible for thrombolysis due to late arrival in the hospital. There were no major complications in the 18 treated patients: 3 patients developed an asymptomatic haemorrhagic transformation of the infarct. CONCLUSION: Acute thrombolysis for ischaemic stroke within 3 hours from stroke onset is feasible, and can under specific conditions be applied in clinical practice. Only 7% of all ischaemic stroke patients underwent thrombolysis. This percentage of patients could be increased by an earlier presentation of patients to the hospital. PMID- 10850110 TI - [Significant decline of the number of invasive Haemophilus influenzae infections in the first 4 years after introduction of vaccination against H. influenzae type B in children]. AB - OBJECTIVE: To evaluate the effect of four years of immunisation against Haemophilus influenzae type b (Hib) on the occurrence of invasive H. influenzae infection and of vaccine failure in children. DESIGN: Descriptive. METHOD: Through the Nederlands Signalerings-Centrum Kindergeneeskunde (NSCK; Dutch Paediatric Surveillance Unit), invasive H. influenzae infections in children under 15 years of age reported by paediatricians were registered from October 1993 until December 1997. On the basis of the NSCK data the incidence of invasive H. influenzae infections was determined for 1994-1997 in relation to the children's age. RESULTS: The numbers of cases of invasive H. influenzae infection were 129 in 1994, 41 in 1995, 24 in 1996 and 8 in 1997. The decrease mainly concerned type b infections. The mean age at infection increased in the first two years after the introduction of vaccination. No apparent change in the clinical presentation of infection was observed. In four years of surveillance a total of nine cases of vaccine failure was reported. CONCLUSION: The progressive effect of vaccination against Hib was reflected in the strong decline of the number of invasive Hib infections, mainly in the youngest children. In accordance with the high level of voluntary vaccination in the Netherlands (95.5% as of January 1st 1997) and the expected high vaccine efficacy the number of vaccine failures was low. A further beneficial effect for the youngest children may be expected from starting the vaccination scheme at a still earlier age. PMID- 10850111 TI - [Luxatio cordis in blunt-trauma thoraco-abdominal injuries]. AB - A 56-year old woman was admitted to the emergency ward after suffering a blunt thoraco-abdominal high-velocity trauma as she hit the wheel when her car drove into a tree. A laparotomy was performed because of haemodynamic instability and radiographic suspicion of a diaphragmatic rupture. Besides haemorrhage from liver and spleen injuries, an abdominal herniation of the heart through a ruptured pericardium and diaphragm was found. Haemostasis of liver, splenectomy and suturing of defects in pericardium and diaphragm resulted in a haemodynamically stable situation. A high index of suspicion of rupture of pericardium and diaphragm with luxation of the heart in the trauma patient is important to reduce morbidity and mortality due to delay of surgical intervention. PMID- 10850112 TI - [Severe Plasmodium falciparum malaria in pregnancy: a threat to mother and child]. PMID- 10850113 TI - [Frequency of postoperative wound infections: an unsuitable parameter for comparison of hospitals]. PMID- 10850114 TI - [Frequency of postoperative wound infections: an unsuitable parameter for comparison of hospitals]. PMID- 10850115 TI - [Three patients with unrecognized orthostatic intolerance]. PMID- 10850116 TI - [Diagnostic significance of antinuclear antibodies]. PMID- 10850117 TI - [Distal myopathies: clinical and genetic lesson]. PMID- 10850118 TI - [Cognitive deficiency in mild hypothyroidism]. AB - OBJECTIVE: Moderate and severe primary adult hypothyroidism produces cognitive and behavioral disturbances, but the effects of mild hypothyroidism are not well established. We have tried to determine the pattern of cognitive performance and the effect of hormonal treatment in mild hypothyroidism. PATIENTS AND METHODS: 15 patients with subclinical or very mild hypothyroidism were compared with other 15 patients suffering from mild diseases without central nervous system damage, who were matched in age, sex and previous intelligence. A neuropsychological battery, including tests of global cognitive performance (Mini Mental Status Examination, WAIS), attention (reaction time), visual memory (Benton test, Rey's Figure), verbal memory (word learning), verbal fluency and executive functions (Trail Making Test), was applied. RESULTS: Hypothyroid patients had worse performance in some WAIS subtests, reaction time, verbal fluency, free recall and copy of the Rey's figure. After hormonal treatment, a significant improvement and normal performance in almost all cognitive functions were found. CONCLUSIONS: Even mild hypothyroidism produces cognitive defects, mainly in tasks with high attentional requirements, that can and should be corrected with hormonal replacement. PMID- 10850119 TI - [Study of inpatient consultation for the neurological services]. AB - INTRODUCTION: Intrahospitalary interconsultation (IC) to a neurology department is an infrastudied task within daily neurologic health care. AIMS: To evaluate the IC to a neurology department. METHODS: A one-year retrospective study was performed on the IC received in the Department of Neurology in the Hospital General Universitario "Gregorio Maranon" (Spain), to evaluate the reasons for consultation, the urgency of the request, concordance between initial clinical suspicion and final diagnosis, specialties involved, final diagnosis and management/deviation to the IC. RESULTS: A total of 432 IC were analyzed. The most frequent reasons for consultation were cerebrovascular disease (19%) and control of already diagnosed neurological diseases (17%). The most frequent final diagnoses were cerebrovascular disease and acute metabolic encephalopathy (16% in both cases). With regard to management, three percent of the cases were admitted to the ward and 14% were sent to out patients for completing studies and/or follow up. CONCLUSIONS: This study underlines the importance of intrahospitalary IC within the daily tasks and health care carried out in a neurology department in relation to the number of IC undertaken during the study period. It is also an excellent method to evaluate the neurologic manifestations observed in systemic diseases and neurologically study patients with diverse pluripathy. PMID- 10850120 TI - [Schizoencephaly or agenetic porencephaly?]. PMID- 10850121 TI - Late-onset isolated gelastic epilepsy secondary to entrapment of the right temporal horn. AB - The case is reported of a 70 year-old woman with isolated gelastic seizures (GS) secondary to a rare form of focal obstructive hydrocephalus, called entrapment of the lateral horn. Laughing attacks started five years after conservative intracranial surgery for a giant basilar aneurysm. Serial neuro-imaging studies revealed a progressive cystic enlargement of the right temporal horn, damaging the baso-lateral temporal cortex. An ictal EEG recording confirmed the epileptic nature of laughing attacks, and showed that the epileptiform activity originated in the right temporal lobe. Complete seizure control was achieved with current doses of diphenilhydantoin. Analysis of this and other previously reported cases, indicate that symptomatic GS may originate in multiple sites of both cerebral hemispheres, although related to the limbic system. The fact that this case exhibited isolated GS stresses the importance of the baso-lateral temporal cortex in the genesis of this type of seizures. PMID- 10850122 TI - [Progressive anarthria: one case without lingual apraxia]. AB - Progressive anarthria is a focal cortical degenerative disorder characterized by a profound, progressive alteration in speech without impairment in other cognitive domains. The first symptoms consist of an alteration in the articulation of speech producing telegraphic speech. The disorder invariably progress towards anarthria by deprogramming of the phonation and orolingual movements (bucophonetic apraxia). This type of apraxia is usually associated with orolinguofacial apraxia and it has been anatomofunctionally correlated with frontal opercular involvement of left predominance. The patient presented as progressive anarthria in the absence of manifest orolingual apraxia associated with a predominance of cortical atrophy in the right frontal operculum. This semiological dissociation emphasizes the importance of bucophonatory apraxia in the pathophysiology of progressive anarthria. PMID- 10850123 TI - [Relationship between self concept and obesity in children and adolescents in various spheres of daily life]. AB - This article considers the relation between overweight and the self-concept of children and adolescents in different living areas. The visibility and perceived controllability of obesity is considered. Results of previous studies about this topic are inconsistent. Self-esteem as one aspect of the self-concept is measured by the "list of statements for children and adolescents". Comparing a group of obese (N = 56) with a group of children and adolescents who suffered from other chronic illnesses (N = 22), the obese had a lower self-esteem for the public living areas "school" and "leisure time", but not for the private area "family". A further result shows that the visibility of the illness is significantly related to self-esteem; again this relation is only revealed for the public living areas "school" and "leisure time". PMID- 10850124 TI - [Decision of German Supreme Court on July 30, 1999 relating to scientific evidence requirements for psychological expert opinion with respect to credibility of testimony and its consequences for future expert consultation]. AB - Basically nothing has been changed by this significant, clarifying, establishing and, most important, determining decision of the Highest Federal Court concerning the specific methods long since applied by experts in the field of deciding whether or not a testimony is believeable. None the less such a decision through the higher courts was obviously long overdue because numerous opinions of experts utilized in court cases were reached by applying methods not free of fault. The psychological evaluation of the content of truth in a testimony in cases of sexual abuse will therefore still be established with help of a methodical procedure of steps made under the assumption that statements pertaining to personal experiences differ in quality from those resulting from imagination in their characteristics of reality. PMID- 10850125 TI - [There once was a child of divorce: story-telling and retelling as a pedagogic therapeutic tool for younger school children in coping with the experiences of separation and divorce]. AB - The author shows that a rearranged narrative as a pedagogical and therapeutic means to help younger pupils coming to terms with the experiences of separation and divorce. Examples given demonstrate that this method can be a valuable help for these children to cope with their anxiety. PMID- 10850127 TI - Building an international community in environmental and occupational health. PMID- 10850126 TI - [Treatment of children with hyperkinetic disorders (ADHD) based on communications and systems analysis]. AB - Inquired for a therapeutic mode changing hyperactive behavior without application of drugs or diets we analyzed the patterns of cognition, communication and system behavior related to such disturbed children and their families. Following our observations we created a clinically useful therapy basing on a special view to system- and communication theory. We applied our therapeutic mode generally to as hyperactive identified primary school children and collected in N = 46 cases catamnestic data about five years later. In more than 80% out of the sample we found successful developments after application of in means 8 therapeutic sessions during 9 months. About 53% out of the successful treatment children show a splendid school success. Our results are demanding for an extended discussion of relations and family constellations including hyperactive behavior of one child. Searching for the possibilities of change in the view of system theory it seems to be more appropriate to consider relations but individual behavior. PMID- 10850128 TI - [Exposure to lead among 6-12 year-old children]. AB - OBJECTIVE: To identify exposure factors contributing to lead poisoning in school children from Mexico City. MATERIAL AND METHODS: Cross-sectional study of 340 children. A convenience sample of schools and a random sample of children were selected. A questionnaire was filled out and venous blood samples were taken. Lead levels were measured by atomic absorption spectrophotometry. Statistical analysis consisted of comparison of means using Student's t test and ANOVA. Multiple linear regression was used for multivariate analysis. Logarithmic transformation of lead blood levels were used to account for their non-normal distribution. RESULTS: Geometric means for private and public schools were: GM = 8.76 micrograms/dl, 95% CI = 9.1-10.5; GM = 11.5 micrograms/dl, 95% CI = 9.4 13.5. Lead levels were higher among children from public schools who are male, between 6 and 8 years of age, in first and second grade, whose mothers have a profession, who use glazed earthenware utensils, and who live near glazed earthenware shops or factories. CONCLUSIONS: Exposure predictors of lead blood levels are: being between 6 and 8 years of age, having a professional mother, using glazed earthenware utensils, living near glazed earthenware shops or factories, and studying the second grade of elementary school. PMID- 10850129 TI - Lead exposure among young urban women. AB - OBJECTIVE: Blood lead levels have declined among every age group in the United States, but urban minority residents remain at disproportionate risk for elevated lead levels. Our objective was to measure lead burden in young women of childbearing age in New York City. We also describe successful means of recruiting this population into a cohort study. MATERIAL AND METHODS: Healthy women aged 18-25 attending a New York City health care center in 1995-1998 were eligible for participation. Participants were recruited by health care providers, the study coordinator and the participants themselves. Venous blood samples were obtained for whole blood lead, ferritin and hematocrit measurements, and detailed questionnaires were administered. RESULTS: 239 women have been recruited to date. The population is predominately minority: 62% African-American, 33% Hispanic and 5% Caucasian/Asian. The average age of participants is 19.3 years. Recruitment of participants into the study is predominantly (55%) through "word of mouth" from previously enrolled participants. Few participants learned of the study through their health care providers. The mean blood lead level among study participants is 2.1 +/- 1.7 micrograms/dl, which is consistent with the most recent United States national survey. CONCLUSIONS: Blood lead levels are low in young, urban minority women of childbearing age in New York City. In this population, recruitment efforts were substantially enhanced with the help of enrolled participants and the health care community. PMID- 10850130 TI - Methodological issues related to studies of lead mobilization during menopause. AB - While there has been a substantial decline in lead exposure in the United States during the past two decades, mobilization of existing lead stored in bone potentially represents an important endogenous source of exposure for menopausal women. It has been hypothesized that lead may be mobilized from skeletal stores during conditions of high bone turnover, such as during menopause. However, such mobilization has not been documented in prospective studies. This discussion is focussed on some of the methodological difficulties to be anticipated in longitudinal studies of lead mobilization specific to menopause and the issues that need to be taken into account when evaluating the results of such studies. To evaluate whether lead mobilization occurs during menopause, a prospective repeated measures design is needed using X-ray fluorescence analysis of lead in bone and serial measurements of blood lead. Potential confounders and effect modifiers also need to be taken into account in the statistical analysis. PMID- 10850131 TI - Male reproduction and environmental and occupational exposures: a review of epidemiologic methods. AB - Concerns that chemical exposures in the environment have been detrimental to male sexual development and fertility have been heightened by reports of declining sperm counts over the past 50 years. Marked geographic variation has been found in semen quality and in the incidence of testicular cancer and certain urogenital defects. Debate continues over the existence, magnitude and significance of these trends, and how best to evaluate the hypothesis that in utero and childhood exposures to estrogenic compounds may be to blame. Epidemiologic methods for assessing the impact of hazardous substances on male reproductive health have been developed mainly in the area of occupational medicine, and this paper will review the currently recommended methods. These include questionnaires to determine reproductive history and sexual function; reproductive hormone profiles; and semen analyses such as sperm concentration, motility, and morphology. New research tools that show significant promise from the fields of clinical reproductive medicine and reproductive toxicology are discussed as possible additions to epidemiologic studies, including assays of sperm function and genetic integrity, and biomarkers of DNA damage. For population-based studies involving occupational groups or communities with environmental exposures, issues related to the cost, validity, precision and utility of these methods must be carefully considered. PMID- 10850132 TI - [Lead elimination by traditional curing]. AB - OBJECTIVE: To evaluate traditional acidic curing as a preventive method for reducing lead content of glazed ceramic ware. MATERIAL AND METHODS: In 27 ceramic ware pieces from four states in Mexico, the level of residual lead was determined through atomic absorption spectrophotometry after four washings with acetic acid 3%. RESULTS: The lead content of the vinegar washing diminished proportionally with the number of washings, although it remained highly above the permissible levels of this metal (2.5-7.0 p.p.m.). CONCLUSIONS: Traditional acidic curing of glazed ceramic ware is not a useful preventive measure for reducing lead exposure. PMID- 10850133 TI - The application of single cell gel electrophoresis or Comet assay to human monitoring studies. AB - OBJECTIVE: In the search of new human genotoxic biomarkers, the single cell gel electrophoresis assay has been proposed as a sensible alternative. MATERIAL AND METHODS: This technique detects principally single strand breaks as well as alkali-labile and repair-retarded sites. RESULTS: Herein we present our experience using the single cell gel electrophoresis assay in human population studies, both occupationally and environmentally exposed. CONCLUSIONS: We discuss the assay feasibility as a genotoxic biomarker. PMID- 10850134 TI - The epidemiology of Her-2/neu and P53 in breast cancer. AB - Breast cancer is an etiologically heterogeneous disease with marked geographical variations. Joint consideration of the relationship between specific molecular alterations and known or suspected epidemiologic risk factors for this disease should help distinguish subgroups of women that are at elevated risk of developing breast cancer. In this article, we present a comprehensive literature review of the etiologic and prognostic roles of Her-2/neu and P53 among women. In addition, we discuss the advantages and limitations of using biomarkers in epidemiological studies. We conclude that more research is needed to understand the complex relationships between genetic alterations and etiologic risk factors for breast cancer. PMID- 10850135 TI - [Difficulties of the methods for studying environmental exposure and neural tube defects]. AB - OBJECTIVE: To discuss the attitudes in the assessment of environmental exposures as risk factors associated with neural tube defects, and to present the main risk factors studied to date. RESULTS: Environmental exposures have been suggested to have a roll in the genesis of birth defects. However, studies conducted in human populations have found difficulties in the design and conduction to show such an association for neural tube defects (anencephaly, espina bifida and encephalocele) because of problems raised from: a) the frequency measures used to compare time trends and communities, b) the classification of heterogeneous malformations, c) the inclusion of maternal, paternal and fetal factors as an integrated process and, d) the assessment of environmental exposures. CONCLUSIONS: Hypothetically both maternal and paternal environmental exposures can produce damage before and after conception by direct action on the embryo and the fetus-placenta complex. Therefore, in the assessment of environmental exposures we need to take into account: a) both paternal and maternal exposures; b) the critical exposure period, three months before conception for paternal exposures and one month around the conceptional period for maternal exposures; c) quantitatively evaluate environmental exposures when possible, avoiding a dichotomous classification; d) the use of biological markers of exposure is highly recommended as well as markers of genetic susceptibility. PMID- 10850136 TI - [A method for assessing health risks in mining sites]. AB - OBJECTIVE: Considering the health risk associated with mining areas, in this work a methodology for the health assessment of this kind of hazardous sites is proposed. MATERIAL AND METHODS: The methodology includes a toxicological assessment, an environmental monitoring of metals, and the exposure assessment of the high risk population. The scheme was evaluated in the mining area of Villa de la Paz, San Luis Potosi, Mexico. The toxicological studies were done in rats treated with mining waste, biomarkers of effect for liver and central nervous tissue were analyzed. Metals levels in surface soil, household dust and water were studied. Finally, urinary arsenic was quantified in children. RESULTS: Neurotoxicity and hepatotoxicity of the mining waste were shown in rats. Then, arsenic and lead levels were analyzed in surface soil, household dust, and water. In all three media, exposure points, heavily contaminated with both metals, were localized. Finally, high levels of urinary arsenic were found in children living in the vicinity of the mine. CONCLUSIONS: Taking into account all these results, the Mexican authorities concluded that a high health risk is present in Villa de la Paz, and a remediation program is in progress. PMID- 10850137 TI - [Exposure to asbestos, smoking, and neoplasms. 1967]. PMID- 10850138 TI - [Universal coverage: an advance towards the right to health for all]. PMID- 10850139 TI - [National Health Insurance expenditures in 1994 for thirty long-term illness]. AB - OBJECTIVE: The health insurance system carried out a survey in 1994 on the illnesses included in the regulations on the thirty long-term illnesses (ALD 30) exempt from patients' contributions to the costs of medical treatment. One of the objectives of this study was to provide the average amount, per patient with these illnesses, of the total expenditures of payments in kind reimbursed by the health insurance system and their distribution by principal expenditure post and by illness. METHODOLOGY: A sample of 67,828 patients was randomly selected at the rate of 2% of the total patients under ALD 30 in November 1994. The expenditures reimbursed over the course of June through November 1994 were collected from a computerized petition on the Health Insurance Information System. RESULTS: The average annual cost per patient within the regulation is estimated at 35,991 FF (+/- 692). Close to half of these expenditures correspond to public hospital stays. The annual expenditures extrapolated from the total ALD 30 is estimated at 143.7 billion FF, or 35% of the total expenditures of payments in kid reimbursed by the health insurance system. "Mental illnesses" represent the largest portion of these expenditures given to ALD 30 (23%). CONCLUSIONS: The survey allows for an evaluation of the average cost per patient covered 100% by the health insurance system under ALD 30. For the majority of these illnesses, these evaluations are the only benchmarks currently available. PMID- 10850140 TI - [Mothers' perceptions about their children's acute diarrhea in the Quito suburbs]. AB - This study was performed in an urban marginal neighborhood of Quito (Ecuador). It aims to understand how the mothers with children (less than five years old) suffering from diarrhea are using the health center. We did 48 semi-directed interviews with mothers coming to the health center. The results show the various representations of diarrhea (symptomatic patterns and explanatory models: e.g. diarrhea due to supernatural cause, diarrhea related to heat and cold balance...) and the related processes of therapeutic search; For example, when the mother thinks that the diarrhea is due to a supernatural cause, she immediately goes to the traditional practitioner. The existing cultural gap between health practitioners of the center and the population of this neighborhood incite to consider differently the content of health education messages. PMID- 10850141 TI - [Child labor in the artisan sector of Morocco: determinants and health effects]. AB - Despite the fact that child labour is regulated through the work code, and the convention on child rights adopted by the General Assembly of the United Nations in 1989 and ratified by Morocco in 1993, multiple surveys have shown that children are often put to work at a very early age and few employers respect the work conditions laid out in the texts. The aim of this study was to assess the different situations of child labour in the handicraft sector, the reasons and the problems surrounding it, to study its repercussions on health and to propose several preventive measures. From March to July 1997, a retrospective cohort study of working children and children in school was carried out in a small neighbourhood of Casablanca. We interviewed and examined a random sample of two hundred children working in the handicraft sector. The health status of these children was compared to that of the same sample size of children in school, from the same age group and socio-economic status. Each subject was given a standardized questionnaire that was translated into dialectal Arabic and administered by a occupational health doctor and a communications specialist. The results of the study have pointed out the small school network of the working children, the painful conditions of work and the important consequences on their health state with a wide prevalence of pathologies higher than for the children attending school. The misery in addition of the rural exodus, the no-adapted educative and socio-economic systems, the splitting of the family unit often go to generate a submissive childhood without defense and "ready to be used". PMID- 10850142 TI - [Food consumption patterns in the urban milieu of Bamako]. AB - Food consumption patterns in urban environments are changing and diversifying. This longitudinal study of individual food consumption took into account the coexistence of two types of food consumption: within the home and outside the home. This article presents a summary of qualitative and quantitative research carried out in Bamako, Mali in 1995 and 1996 among 74 families from different socio-economic groups. It provides a comprehensive analysis of their food strategies (at the family and individual level) and their nutritional intake. In the home, similarities are found: three meals a day, and a single dish with a "base" ingredient and sauce at each meal. Heterogeneity is expressed in the number of dishes prepared each day, the type of "base", and ingredients in the sauce. Almost all individuals, no matter what their age or socio-economic status, consume food outside the home. This does not substitute meals at home. Street food accounts for 19-27% of food expenses and provides 134-417 Kcal per day per person. It offers a market for local products not often consumed at home and provides a necessary nutritional supplement to people from families of intermediate or low socio-economic status, especially for children. For people from higher socio-economic levels, street food provides mainly gustative and symbolic satisfaction. Following the devaluation of the CFA franc, the most disadvantaged populations seem to have reached a limit in their capacity for adaptation, which includes not only the quality of consumed products but also quantities. In order to ensure food security in urban areas, food and nutritional policies must take into account all the factors that influence food consumption behaviours of different groups of individuals, in particular the behaviours inside and outside the home. The latter is too often overshadowed. PMID- 10850143 TI - [Survey of chemotherapy practice in public and private hospitals in Brittany]. AB - With the objective of planning for cancer research, an assessment of chemotherapy practices in public and private establishments was carried out in 1997 by the Regional Direction of Health and Social Affairs of Brittany. This retrospective survey was carried out during the first trimester of 1997. A questionnaire collected quantitative and qualitative data: skills of mobilised personnel, modes of preparation and dispensation of cytostatic drugs and partner relations. In the 38 establishments reporting that they provide chemotherapy, the data showed varied practices for: the number of patients treated (0 to 579), number of visits per patient (2.07 to 9.4), and skills of the practitioner. The indicators used in this study, with the objective of analysis and aid in planning, will serve in the evaluation phase of the implementation of the regional health programme in cancer research. PMID- 10850144 TI - [Training of field professionals in Switzerland health prevention projects]. AB - Following various difficulties during prevention programs, an interdisciplinary training course in project management was set up for professionals working in the field. Sessions devoted to theoretical aspects alternated with workshops aimed at problem-solving strategies. The intention of the course was to equip the participants with relevant operational tools. The twenty-seven participants came from twelve different professional backgrounds. Fourty-two hours of training were given over a period of nine months. Two physicians specialized in public health accompanied the course throughout, seven invited speakers joined the group for occasional sessions. The evaluation was positive for the teaching methods used and the variety of subjects covered. The analysis of practical experiences has proved to be a powerful learning tool and a vehicle for interdisciplinary exchanges. The course has highlighted the isolation of the professionals in the field, the absence of common points of reference and the resulting difficulties in communication and collaboration. This experience has clearly shown the need for a well-defined standard model in project management and for intensive training of the professionals in the field. PMID- 10850145 TI - [Evaluation of nutritional status in hospitalized aged persons]. AB - The health status of older people strongly depends on their nutritional status. Hospitalisation is a factor that aggravates protein-energy malnutrition among older people. A descriptive study was completed among 71 subjects over 65 years old and hospitalised in a gerontological internal medicine service. The study used a nutritional evaluation scale called the Mini Nutritional Assessment (MNA). The study showed that 21% of subjects had an acceptable nutritional status, 49% were considered to be "at risk of undernourishment", and 31% were malnourished. A relationship was observed between the MNA score, the degree of autonomy (AGGIR grid), cognitive functions (Mini Mental State: MMS) and the existence of an anxious-depressive state. These elements should be taken into consideration to avoid the aggravation of the nutritional status of older people in the hospital, or to improve it. In addition, certain simple biological exams (albumin, pre albumin, reactive protein) are necessary to assess the endogenous or exogenous aspect of undernourishment. PMID- 10850146 TI - [Knowledge and communication in public health. Evolution of scientific knowledge during the dramatic moment of blood contamination. 1--Debates concerning the origin and nature of AIDS]. PMID- 10850147 TI - [Medical and Human Biological Sciences department of the University of Corsica as a tool for public health development]. AB - To overcome the difficulties of access to training in health and the absence of biomedical research, the territorial community of Corsica recommended the implementation of a Department of Medical Sciences and Human Biology within the University of Corsica. It is within the framework of this department that different training courses targeting the full range of actors in the health system were organised, the large majority in agreement with medical faculty from the continent. It is also within this framework that a set of research themes was developed, permitting at full term, a better health management of the population. After four years in existence, we propose an assessment of training actions, and the results of the research work that associated hospital practitioners and university research professors and which allows one to imagine, notably, the development of a research axis in geography and health on the theme, "Genetics Public Health-Environment in the Islands of the Mediterranean". PMID- 10850148 TI - [History of veterinary education in Bern. Fragments on the 100 year jubilee of the faculty of veterinary medicine]. PMID- 10850149 TI - [Development of the faculty]. PMID- 10850150 TI - [The new curriculum of veterinary medicine in Berne]. PMID- 10850151 TI - [The PhD program: an initiative for improving the academic training of veterinarians]. PMID- 10850152 TI - [Virology integrated]. PMID- 10850153 TI - [Research from the Division of Immunology: production of nitric oxide (NO) by macrophages in ruminants]. PMID- 10850154 TI - [Genetic diagnosis in bacteriology]. PMID- 10850155 TI - [Bovine colostrum: more than just an immunoglobulin supplier]. AB - The intake of colostrum in newborn calves not only influences the immune status, but also the development of the gastrointestinal tract and other organs, metabolism and endocrine systems. Besides nutrients bioactive substances of colostrum have additionally to be considered. Metabolic and endocrine effects are transient, indications for metabolic and endocrine imprinting by differences in feeding in the first week of life have so far not been found. Nevertheless, changes observed in association with feeding especially of colostrum are considered important for the critical first week of life. PMID- 10850156 TI - [Genetic investigations for control of hereditary diseases in dogs]. PMID- 10850157 TI - [Infectious keratoconjunctivitis]. PMID- 10850158 TI - [Impairment of cell adhesion in autoimmune bullous skin diseases]. PMID- 10850159 TI - [Species differences in cardiac sodium channels]. PMID- 10850160 TI - [Hypothyroid-associated gait abnormalities in the dog]. PMID- 10850161 TI - [Neospora caninum and neosporosis--basic science at the Institute of Parasitology and possible implications]. PMID- 10850162 TI - Exposure of rainbow trout (Oncorhynchus mykiss) to nonylphenol is associated with an increased chloride cell fractional surface area. AB - Nonylphenol is a biodegradation product of a widely used group of non-ionic detergents. Because of its ubiquitous distribution and persistence, nonylphenol is present in surface waters as a pollutant. Little is known about its biological effects at environmentally relevant concentrations other than its action as a xenoestrogen. The goal of the present paper was to study the trout gill surface epithelium as the major interface between fish and water in view of possible morphological alterations due to exposure to nonylphenol. Rainbow trout were intermittently exposed to 10 micrograms/l nonylphenol and gill samples from experimental and control animals were investigated by scanning electron microscopy. Gill surface epithelium was scrutinised for changes in chloride cell density and their status regarding cell surface modifications. In addition, chloride cell fractional surface area (CCFA) was determined by morphometrical methods. Statistical analysis revealed a highly significant increase of CCFA in animals exposed to nonylphenol as compared to control animals (P = 0.0001). Semi quantitative assessment of the other parameters suggested a higher chloride cell density and a larger proportion of chloride cells bearing microvilli. Taken together, these results provide evidence that exposure of trout to nonylphenol is associated with a substantial increase in the active interface of chloride cells with water. We interpret these findings as being a means to further the fish's capacity for calcium exchange. PMID- 10850163 TI - [Tricuspid valve dysplasia in fifteen dogs]. AB - Clinical findings in fifteen dogs with tricuspid valve dysplasia are described. Thirteen dogs had loud systolic heart murmurs. Eleven of them hat a palpable precordial thrill over the same location. In 14 dogs, right heart enlargement was suspected on thoracic radiographs and electrocardiography. Right atrial dilation was seen echocardiographically in all dogs. Fourteen dogs had additional right ventricular dilatation, some with hypertrophy as well. Doppler echocardiography revealed tricuspid valve regurgitation. Seven dogs remained free of clinical symptoms to date. If symptoms of decompensation develop with tricuspid dysplasia, diuretics, ACE inhibitors and eventually positive inotrope drugs are indicated. Antiarrhythmic drugs may become necessary in cases of supraventricular tachyarrhythmias. PMID- 10850164 TI - [Clinical use of hollow cylinder screw implants for various indications. (Functional mandible reconstruction after tumor resection osteotomy, stable maintenance of spinal column, prosthetics) 20 year experience]. PMID- 10850165 TI - [Synovial osteochondromatosis (SOC) in swine: a case report]. PMID- 10850166 TI - [The treatment of articular and bone infections in large animals with gentamicin impregnated collagen sponges]. PMID- 10850167 TI - [Lung function tests in horses with special reference to ultrasound spirometry/capnography]. PMID- 10850168 TI - [Electrical stunning in cattle before exsanguination. Review of the literature and individual experimental results]. PMID- 10850169 TI - [The osteoplastic efficacy of different forms of hydroxyapatite based on experimental morphological study data]. AB - Time course of healing of induced defects in the femoral bone filled with various forms of hydroxyapatite (HA) was studied in experimental rats on days 30, 60, and 90 after inflicting the defect. HA granulate exerted the best effect on bone repair. In experiments with HA granulate, bone regenerate replaced up to one half of the area of defect by day 90. After insertion of a mixture of granulate and HA powder in bone defects the newly formed bone occupied a little more than one third of the defect area. In experiments with HA powder suspension the size of bone defect by day 90 was the same as after healing under a blood clot. PMID- 10850170 TI - [A marker and prognostic test for phospholipase A2 in periodontal inflammatory diseases]. AB - Phospholipase A2 is one the main enzymes determining cell membranes morphological and functional condition. Close correlation has been revealed between the activity of this enzyme and clinical state of inflammatory process in periodontium. According to the obtained data phospholipase A2 can be a precise diagnostic and prognostic marker of the periodontal tissues condition in pathology. PMID- 10850171 TI - [Caries prevalence and intensity and the health and hygiene habits of oral care in pregnant women]. AB - Examination of the oral status of 664 pregnant women showed that from 77.12 +/- 3.90% at the age of 17-19 years to 100% at the age of 35-39 years were in need of treatment and removal of teeth. An average of 2.46 tooth per woman were to be filled and every other woman was in need of tooth removal. The quality of oral hygiene was poor in the examined patients, the Fedorov-Volodkina index varying from 2.2 +/- 0.05 to 2.6 +/- 0.1. PMID- 10850172 TI - [The blood immunological indices in rapidly progressing periodontitis (preliminary results)]. AB - Clinical and immunological parameters were studied in 20 patients with rapidly progressing periodontitis (RPP). Changes in the immune status were as follows: the count of CD3+ cells in peripheral blood increased, the ratio of regulatory helper to suppressor populations (CD4+/CD8+) decreased, and the level of circulating immune complexes increased. A relationship between some clinical and immunological parameters was detected. The detected changes in the immune status of patients with RPP were virtually the same as those in adult patients with typical periodontitis. Further studies are needed for detecting stable criteria of immunological changes, typical of RPP. PMID- 10850173 TI - [The effect of polychemotherapy on the oral mucosa in patients with acute myeloblastic leukemia]. AB - Oral mucosa was examined in 23 patients with acute myeloblastic leukemia before and after cytostatic polychemotherapy. Lesions of the oral mucosa were observed in all patients after polychemotherapy: its color was changed, atrophic processes developed in it, and salivary secretion was impaired. Destructive ulcerative changes were revealed in 35% cases; terms of their appearance, regression, and epithelialization were established and a clear-cut correlation between their development and severity of leukocytopenia was detected. Ranging of stomatitis severity is proposed, according to which medium-severe stomatitis predominates in this patient population. The study will help develop more effective approaches to prevention and treatment of oral complications and improve the results of therapy of this most grave category of hematological patients. PMID- 10850174 TI - [The differential diagnosis of avascular necrosis of the temporomandibular joint in systemic lupus erythematosus]. AB - Eight patients with facial skull abnormalities with systemic lupus erythematosus (SLE) and temporomandibular involvement were examined. In 4 patients mandibular deformation was caused by erosive arthritis and in 4 others by aseptic necrosis of the temporomandibular joint. Clinical and x-ray characteristics, magnetic resonance and computer tomography findings in the two groups are compared. A detailed protocol for differential diagnosis of temporomandibular joint involvement in SLE is developed. PMID- 10850176 TI - [A comparative evaluation of the immune status of patients with squamous-cell cancer of the oral mucosa at different stages of combined treatment]. AB - Shifts in immunity values were detected in 160 patients with oral cancer before specific therapy and at all stages of multiple-modality treatment. High diagnostic significance of measuring concentrations of circulating immune complexes in the serum for prediction of tumor relapse or metastases is demonstrated. PMID- 10850175 TI - [Immunocorrective treatment in mandibular fractures in inhabitants of the European North]. AB - Consolidation of mandibular fragments in 120 patients with fractures of different location was evaluated by clinical, x-ray, and functional methods (rheography and myography). After conventional treatment the fragments consolidated in 34.5 +/- 0.6 days in unilateral fractures and in 39.3 +/- 0.7 days in bilateral fractures. After combined therapy including immuno-correcting and vasodilating agents recovery was attained 4-5 days earlier. PMID- 10850177 TI - [The possibility for the early diagnosis of suppurative-inflammatory wound complications of the maxillofacial area and neck by using an acoustic skin analyzer]. AB - The acoustic method was used for early diagnosis of pyo-inflammatory complications of wounds of the maxillofacial area and neck. Acoustic skin analyzer was used. The velocities of acoustic wave propagation (Vx, Vy) were notably increased in the pyo-inflammatory focus in comparison with perifocal tissues and with the symmetrical contralateral side. The difference in the absolute values of velocities was about 45-65 m/s. No acoustic anisotropy was observed in the focus of inflammation. Hence, the acoustic method can be used as a method for early diagnosis of pyo-inflammatory complications and for monitoring the wound status in the course of wound process. PMID- 10850178 TI - [The fixation of nonremovable dentures: the rational choice of the material]. AB - Adhesion and marginal permeability of materials used for fixation of whole-cast dentures and surface micro-relief were studied in vitro and in vivo on dogs. Three groups of materials were studied: zinc-phosphate, polycarboxylate, and glass-ionomeric. Polycarboxylate cements effectively fixed dentures irrespective of the status of abutment teeth, and glass-ionomeric cements w V.M. Marker and prognostic phospholipase A2 test in inflammatory diseases of the periodontium. PMID- 10850179 TI - [A hardware and software complex for producing 3D models of the teeth]. AB - A complex of short-basis photogrammetry, methods and software for obtaining and analysis of 3D digital models of relief of dentition and its fragments is presented. This complex is intended for planning and evaluation of tooth preparation and quality of prosthetic treatment. The method was used to assess the correctness of dental shape repair by artificial crowns. The complex can form the base for the CAD/CAM technology. PMID- 10850180 TI - [The characteristics of preparing metalloceramic dentures for patients with cerebrovascular pathology]. AB - 35 patients with aftereffects of cerebral stroke and 25 patients with hypertension were examined. A high rate of parodontal disease was revealed. An orthopedic stomatologic method of making metalloceramic dentures was devised. PMID- 10850181 TI - [Atopic cheilitis in children: the risk factors and clinical symptoms]. AB - General and local factors affecting the development of atopic cheilitis, alone and in association with disseminated neurodermatitis, have been studied. Health disorders were more incident in children with cheilitis concomitant with disseminated neurodermatitis than in those with cheilitis alone (without other skin involvement). The most significant general risk factors are an unfavorable course of antenatal development, exudative catarrhal diathesis, food allergies, gastrointestinal diseases, intestinal dysbacteriosis. Local risk factors were impaired joining of the lips and disordered nasal breathing, long soother sucking, harmful habits, and maxillodental abnormalities. Clinical symptoms in isolated cheilitis and cheilitis concomitant with disseminated neurodermatitis are similar, but cheilitis associated with disseminated neurodermatitis runs a more severe course. PMID- 10850182 TI - [The mechanism of the eruption of the permanent teeth and the causes of the development of anomalies of the maxillodental system]. PMID- 10850183 TI - [The characteristics of the manifestation of syphilis on the mucosa of the mouth and oropharynx in children and adolescents]. AB - Case histories of 81 children and 711 adolescents are analyzed. Specific lesions of the oropharyngeal mucosa have been detected in 43 (53.08%) children and 364 (51.20%) adolescents. Clinical manifestations of diseases of the oral mucosa and location of syphilitic elements and terms of their healing are analyzed. PMID- 10850184 TI - [The problems of dental equipment in Russia--the manufacture of dental units]. PMID- 10850185 TI - [The postgraduate training of dentists]. PMID- 10850186 TI - [An evaluation of the efficacy of immunocorrective treatment with the preparation Imudon in patients with generalized periodontitis against a background of diseases of the internal organs]. PMID- 10850187 TI - [Screening for cervix cancer--where are we going?]. PMID- 10850188 TI - [The Cochrane Hepato-Biliary Group--status after three years]. PMID- 10850189 TI - [Human infection immunology. Basic scientific findings and clinical consequences]. AB - During the last years studies on human immune responses to specific micro organisms have contributed to an increased understanding of the structure and function of the immune system. T lymphocytes with a special antigen receptor (gamma/delta cells) are thus probably important in some infectious diseases such as malaria and tuberculosis. The discovery of T-cell responses to protein-free fractions of mycobacteria and Leishmania parasites has lead to the identification of non-protein T-cell antigens. T-cells that express the CD4 receptor on their surface and are capable of lysing infected target cells have been found in viral infections. It has furthermore been found, that the composition of the signal mediators (cytokines) secreted by activated T-cells can determine the outcome of a number of infections. In the future it may be possible to include enhancement of protective immune reactions in the treatment of infectious diseases to a higher extent than now. PMID- 10850190 TI - [Partial re-screening of all negative smears. A method of quality control of pathology department concerning smear screening against cervix cancer]. AB - Partial rescreening of all negative smears is recommended as the most cost effective method of internal quality control. The method was tested by eight cytotechnologists and the average diagnostic sensitivity, assessed upon their answering of 200 test-smears (150 negative and 50 positive) was 80%, while specificity vas calculated to 95%. Partial rescreening has here-upon been used in daily routine. After one year 21,000 smears have been re-screened. Twenty-nine cases of false negatives, 16 with atypia, eight with koilocytosis, and five cases with dysplasia have been detected, which corresponds to an overall false negative rate on 3%. In five of the 29 false negative cases with dysplasia histological follow-up has shown three cases of CIN III (two carcinoma in situ, one severe dysplasia), one case with CIN II (moderate dysplasia) and one case with CIN I (mild dysplasia). Conclusively, partial rescreening of all negative smears implies an improved quality with reduction of the number of false negative specimens. PMID- 10850191 TI - [Glaucoma patients treated by practicing ophthalmologists in Denmark. Estimated number of patients and the extent of visual field defects]. AB - A test survey based on a questionnaire was carried out among Danish eye practitioners. The response rate was 72%. The purpose was to estimate the minimum number of patients suffering from glaucoma (prevalence). Survey results show a prevalence of 0.65%. Further, the survey shows a clear correlation between glaucoma diagnosis and age. The number of glaucoma patients increase from approximately 0.5% for persons 50 years of age to approximately 5% for persons 80 years of age. Seventy-two percent of the diagnosed glaucoma patients have visual field defects at different stages. Six percent have central islands on both eyes, the equivalent of a prevalence for social blindness of 0.04%. Ninety-three percent of the diagnosed glaucoma patients receive anti-glaucomatous treatment, of these 42% combination therapy. Eighty percent of glaucoma patients in combination therapy received three or more drugs. PMID- 10850192 TI - [Urinary incontinence and pregnancy, vaginal childbirth and obstetric interventions]. AB - The aim was to examine the association between pregnancy, vaginal childbirth (VC) and obstetric techniques, and the prevalence of urinary incontinence (UI). A cross-sectional survey enrolled a random population sample of 6240 women aged 20 59 years, who were mailed a self-administered questionnaire on UI and, among other things, experience of VC and obstetric intervention. More than 75% responded. The present analysis includes 4345 women. Multivariate UI prevalence odds ratios were increased in relation to UI during pregnancy, UI following immediately after a VC, and age 30 or more at the second VC. No multivariate association was found in relation to forceps delivery or vacuum extraction delivery, episiotomy or perineal suturing. In conclusions, not only the process of childbirth itself but also processes during pregnancy seem to be strongly associated with prevalent UI. Perineal suturing may be associated with prevalent UI, whereas other obstetric techniques inspected do not seem to be so. PMID- 10850193 TI - [The effect of a vaginal device on urinary leakage and quality of life of women with stress urinary incontinence]. AB - The aim of this study was to assess the effect of a vaginal device (Continence Guard) on urine leakage and its impact on quality of life (QoL). Fifty-five women participated in a three month study using the Continence Guard. QoL was assessed by an incontinence specific questionnaire (IIQ), two incontinence specific questions and the generic SF-36 health questionnaire. A total of 41 (74.5%) women completed the study. Use of the vaginal device was associated with subjective cure in 11 women (27%) and improvement in 27 (66%). The mean 24-hour pad test leakage decreased significantly. QoL measured by the IIQ and the two incontinence specific questions showed highly significant improvements. The SF-36 questionnaire showed no significant changes. In conclusion treatment with the Continence Guard significantly decreases leakage and improves QoL in women with the symptom of urinary stress incontinence. The SF-36 questionnaire was not sensitive enough to detect alterations in QoL in patients with stress urinary incontinence. PMID- 10850194 TI - [Direct booking of legal abortions by general practitioners to the surgery department--a pilot study]. AB - In order to facilitate the referral procedure for legal abortion, 26 GPs (34% of all GPs in the region) were given the opportunity to book patients directly to the operating unit, without an outpatient' visit to the gynaecology clinic. Thirty-eight patients booked directly to the operating unit by their GP for a legal abortion were compared to 144 patients referred in the same period of time, in the usual procedure to the outpatients' clinic. The number of legal abortions carried out, reasons for abortions cancelled, problems with the referral procedure and complications were registered. There were no differences between the two groups regarding the percentage of legal abortions carried out, the reasons for cancelled operations, problems with the referral procedure or operative complications. Direct-booking of legal abortion by the GP to the operating unit seems to be a safe and convenient alternative to the referral through a gynaecological outpatients' clinic. Further large scale studies are needed in this new field of communication. PMID- 10850195 TI - [Genetic causes of hearing loss--status and perspectives]. AB - Hearing impairment, defined as > or = 40 dB hearing loss, is the most prevalent sensory handicap, present in 1:750 children and in 4-36% of adults, depending on age. Genetic factors are of major importance in more than 50% of all hearing loss. An important distinction is made between syndromic deafness and isolated deafness depending on the presence or not of associated manifestations from other organs. The knowledge about genetic deafness has increased dramatically in the last few years. As of March 1999, at least 53 loci for isolated deafness of different types of monogenic inheritance have been identified. Suspected genetic heterogeneity has thus been confirmed. At least 15 genes for syndromic deafness have been cloned, leading to increased biological insight in shared developmental pathways in different species and leading to better diagnostic tools applicable to patients. The identification of a particularly frequent mutation in a gap junction gene, GJB2 (connexin 26), may turn out to be of particular diagnostic importance in the aetiological evaluation of childhood deafness even in isolated cases. Application of early screening tests, like otoacoustic emission (OAE), in combination with genetic tests will facilitate early and specific diagnosis of hearing impairment and thereby improve audiological rehabilitation. Syndromic deafness involves all organs, and care for and evaluation of affected individuals should be a multiprofessional task. PMID- 10850196 TI - [Primary adenocarcinoma in appendiceal diverticulitis]. AB - A case of primary adenocarcinoma in appendiceal diverticulosis is reported. Such a case has never been mentioned before in the literature. It was not possible to diagnose the case preoperatively. This emphasizes the importance of histological examination of all appendiceal samples. PMID- 10850197 TI - [Blindness in temporal arteritis despite prednisolone treatment]. AB - Giant cell arteritis occurs most commonly in elderly individuals. The lesion is usually restricted to major arteries. Other arteries can be involved e.g. posterior ciliar artery, which is an important complication, because of the risk of blindness. It is commonly accepted, that blindness will not occur after corticosteroid has been initiated. In this article however, a patient is described who became blind on both eyes in spite of initially moderate doses of corticosteroid. PMID- 10850198 TI - [Quality assurance and maintenance of percutaneous nephrostomy drainage]. PMID- 10850199 TI - [Picture of the month. Gallbladder concremont]. PMID- 10850200 TI - [Atherosclerosis, infections and immune response]. PMID- 10850201 TI - [Ambulatory tracing of tuberculosis; a screening project in Copenhagen]. PMID- 10850202 TI - [Inhibition of deformities (disability) in rheumatoid arthritis]. PMID- 10850203 TI - [Estimation of mortality]. PMID- 10850204 TI - [A comment to the memento on breast cancer in the Ugeskrift for Laeger number 11]. PMID- 10850205 TI - [Correct drug administration--but how?]. PMID- 10850206 TI - [Cerebral control of human motor activities]. PMID- 10850207 TI - [Acute disseminated encephalomyelitis in children]. AB - Acute disseminated encephalomyelitis (ADEM) is a demyelinating autoimmune inflammatory disease of the central nervous system. It is most often seen in children a few days to several weeks after certain infections or vaccinations. Clinically, ADEM can hardly be distinguished from primary viral encephalitis. The diagnosis is most frequently established on the basis of characteristic findings on MRI of the brain. Mostly the disease seems to be self-limiting, but in untreated patients the mortality rate may be as high as 20% and the risk of permanent neurological deficits 10-33%. Treatment with glucocorticoids seems to be effective in the majority of patients and should at least be started in all patients with an affected level of consciousness. Treatment with intravenous immunoglobulin has recently been given in a few patients with apparent success. PMID- 10850208 TI - [Roller skating accidents. Registration of roller skating injuries in two emergency departments]. AB - Rising popularity of roller-skates in recent years has resulted in increasing numbers of accidents involving roller-skates. The subject of our study was a consecutive number of patients who were treated at the casualty departments of Svendborg and Middelfart hospitals, Denmark, between July 1997 and June 1998. A total number of 169 patients representing 179 injuries were registered. A high incidence of injuries was found among children of school age. Protective equipment was used, in 31% (n = 52) of cases, which shows an increase compared to previous studies in Denmark (1). Injuries of the distal antebrachium, wrist and metacarpals account for 45% of all injuries. Wrist guards protect the area but do not completely eliminate the risk of fractures and injuries. A larger usage of protective equipment will probably result in a decrease in the number of injuries, and therefore information about protective measures is essential. PMID- 10850209 TI - [Should all younger patients with breast cancer be offered adjuvant cytotoxic chemotherapy?]. AB - The aim of the study was to investigate whether young age at diagnosis is a negative prognostic factor in primary breast cancer and how stage of disease at diagnosis and treatment may influence such an association. It was conducted as a retrospective cohort study based on a population-based data-base of breast cancer diagnosis with detailed information on tumour characteristics, treatment regimens, and vital status and included 10,356 patients with primary breast cancer less than 50 years of age at diagnosis. The main outcome measures were relative risk of dying within the first ten years after diagnosis according to age at diagnosis, adjusted for effect of known prognostic factors and expected mortality. Overall, young patients with low risk disease who did not receive adjuvant treatment had a significantly increasing risk of dying with decreasing age at diagnosis (adjusted relative risk: 45-49 years: 1 (reference); 40-44 years: 1.12 (0.89-1.40); 35-39 years: 1.40 (1.10-1.78); < 35 years: 2.18 (1.64 2.89). However, a similar trend was not seen in young patients who received adjuvant cytotoxic therapy. We found the same difference as above when comparing women receiving no treatment with those receiving adjuvant cytotoxic therapy within strata of node negative patients and patients with the same tumour size. In conclusion, the negative prognostic effect of young age is almost exclusively seen in women diagnosed with low risk disease not receiving adjuvant cytotoxic therapy, whereas young women who receive adjuvant cytotoxic therapy have the same prognosis as middle-aged women. These results suggest that young women with breast cancer, on the basis of age alone, should be regarded as high risk patients and be given adjuvant cytotoxic therapy. PMID- 10850210 TI - [The pattern of medication errors in connection with intravenous administration at an intensive care unit]. AB - In a static study, medication errors in intravenous administration were registered in a multidisciplinary intensive care unit. The most frequent type of errors included administration of incorrect doses, but also administration of diluted drugs over an excessive time period, use of previously opened ampoules, administration of unauthorized drugs, wrong infusion rates, dissolution of drugs in incorrect media, dissolution of drugs in incorrect volumes of dissolution media and wrong dosage form errors were observed. PMID- 10850211 TI - [Nutritional status in hospitalized younger and elderly patients]. AB - The purpose of the study was to compare the nutritional state in a group of hospitalised patients aged over 65 years and a group of younger patients. Information about body height, body weight, body mass index (BMI), weight index (WI), percent weight loss and energy intake was obtained from 89 elderly and 55 younger patients. We found no significant difference in the prevalence of malnutrition (BMI < or = 18.5 kg/m2 and/or WI < or = 80%) or underweight (BMI < 20 kg/m2) among the elderly patients compared to younger. However, significantly more elderly patients had lost more than 5% of their body weight (52.1% vs. 10.0%, p < 0.001). Also, the elderly patients had had a significantly longer hospital stay before the survey started (14 days vs. 4 days, p < 0.001). The prevalence of weight loss was higher (74% vs. 30%, chi 2 = 8.1, p < 0.01) among the elderly with the longest stay (18 days vs. 4 days). Therefore, special attention must be directed towards the nutritional status of the elderly patient in order to initiate nutritional support in time. PMID- 10850212 TI - [Treatment of severe acute disseminated encephalomyelitis with intravenous immunoglobulin]. AB - A three year-old boy with severe acute disseminated encephalomyelitis (ADEM) responding dramatically to treatment with intravenous immunoglobulin (IVIG) 1 g/kg/day is described. Initial treatment with intravenous methylprednisolone 2 mg/kg/day had failed. This and two earlier case reports indicate that IVIG might be efficient in the treatment of ADEM, and we believe that its use should be considered, at least in cases of severe ADEM with insufficient treatment effect of steroids. A synergistic effect of treatment with steroids and IVIG in ADEM is possible. PMID- 10850213 TI - [Break of radial artery catheter resulting in a loose intra-arterial fragment]. AB - An unusual complication after radial artery cannulation is described. Due to difficult fixation the distal part of the catheter broke loose, and was captured intravascularly. This complication has only been reported once before. The present case did not lead to surgical intervention. A suggestion is made as to, how to avoid this complication. PMID- 10850214 TI - [Diagnosis of Clostridium difficile infection in pseudomembranous colitis]. AB - C. difficile is known as the main cause of pseudomembranous colitis, however, some individuals may be asymptomatically colonized. In this paper two patients with diarrhoea had three respectively five negative stool cultures. Endoscopically, one patient had severe colitis consistent with both pseudomembranous colitis and inflammatory bowel disease. In the other case the endoscopic findings were typical for pseudomembranous colitis. In both cases there were positive cultures for C. difficile from biopsies from colon-plaques. We find that culture of biopsy of a colon-plaque may contribute to the detection of C. difficile infection in patients with negative stool cultures. PMID- 10850215 TI - [The Danish Gulf study and mass media]. PMID- 10850217 TI - [Philosophical exercises]. PMID- 10850216 TI - [Occupational environment--an issue for physicians?]. PMID- 10850218 TI - [Interaction between oral contraceptives and antibiotics]. PMID- 10850219 TI - [Parasitologic diagnosis. Schistosomiasis]. PMID- 10850220 TI - [Drug metabolism in the small intestine--the significance for biological availability]. AB - The gut contains drug metabolizing enzymes and drug export proteins, of which the most important are CYP3A4 and P-glycoprotein. P-glycoprotein is localised to the apical membrane and CYP3A4 in the subapical cytoplasmic membranes in the enterocytes. The substrate and modulator specificity overlap between the two systems. The function of the two systems may be integrated. The intestinal first pass effect is the fraction of a drug which is metabolised or excreted during the absorption from the lumen. The significance of the intestinal first pass effect for the bioavailability of the three model drugs, midazolam, cyclosporin and digoxin, has been reviewed. The impact of different disease states in the gastrointestinal tract on the function of intestinal drug metabolising enzymes and P-glycoprotein is far from elucidated. Further insight into the function of these systems may lead to optimisation of drug therapy. PMID- 10850221 TI - [Methods for drug measurements--antibiotics--outside the blood flow]. PMID- 10850222 TI - Chagas disease, Brazil. PMID- 10850223 TI - Resistance to antituberculosis drugs. PMID- 10850224 TI - How do I fire thee? Let me count the ways.... PMID- 10850225 TI - How to fire without fear. PMID- 10850226 TI - Value of developmental age determination in orthodontic treatment planning. PMID- 10850227 TI - Current status of growth hormone treatment for short children. PMID- 10850228 TI - Diagnosis and correction of Class III malocclusion. PMID- 10850229 TI - Collaborative approach for identifying and treating speech, language, and orofacial myofunctional disorders. PMID- 10850230 TI - The dentist's use of normal developmental concepts in making the dental experience a developmentally enhancing one for the child. AB - In all of these dentist-child interactions, the dentist is informing the child that he or she has respect for and knowledge about the child's innate needs and the rules about how these innate needs can be gratified in the dentist's office. When this information is given to the child, the child develops trust in the dentist as a kind, caring, smart, and competent new adult in the child's life. The dentist becomes a new building block in the child's overall developmental growth, and the overall dental experience is deposited in the child's long-term memories as a developmentally enhancing one. PMID- 10850231 TI - Bearbaiting in dentistry: a specialty under siege. PMID- 10850232 TI - Aberrant eruption and its relationship to crowding: the need for early detection and management. AB - A modular, fixed-functional method has been described using a new generation of appliances that are smaller, more comfortable and more patient friendly, while gaining the advantages of a combined fixed-functional and postural therapy. The case has been made for the middle mixed dentition as the optimal starting time. Crowding, overbite and molar rotation are managed simply with a combined use of flexible lip bumpers, bite plane and sequential primary tooth extraction. In conclusion, Phase I Postural Therapy will be shown to have the following advantages: more biologically and patient friendly more conducive to non extraction more efficient and cost effective simple fabrication and reduced chair time reduced Phase II fixed appliance time, and completion at a younger age. PMID- 10850233 TI - Role of developmental and occlusal conditions in timing orthodontic treatment. PMID- 10850234 TI - Interceptive guidance of occlusion with emphasis on diagnosis. PMID- 10850235 TI - Adult orthodontics and alveolar bone changes. PMID- 10850236 TI - Orthodontic management of pseudoprognathism and tooth-size discrepancy. PMID- 10850237 TI - A new material concept for inhibiting the formation of secondary caries. PMID- 10850238 TI - In vitro evaluation of a "smart" dental material for its efficacy in preventing secondary caries using a microbial artificial mouth model. PMID- 10850239 TI - Inhibition of enamel and root dentin demineralization by Ariston pHc: an artificial mouth study. PMID- 10850240 TI - In situ study on the caries-preventive effects of fluoride-releasing materials. PMID- 10850241 TI - Clinical experience with a new tooth-colored dental restorative material (Ariston pHc) on primary teeth. PMID- 10850242 TI - Efficacy of a new caries-inhibitory restorative material and amalgam as control in 12-13 year-old Latvian adolescents with high caries prevalence: 1-year results. PMID- 10850243 TI - Clinical evaluation of Ariston pHc restorations: 1-year results. PMID- 10850244 TI - Bonded restorations for the prevention and treatment of the cracked-tooth syndrome. AB - Several reports revealed that the cracked-tooth syndrome is a common problem in dental practice, which often results in extraction of the affected incompletely fractured teeth. Predominantly restored teeth suffer from these incomplete fractures. Therefore, it is of outstanding importance to stabilize teeth weakened due to cavity preparation. Besides full cuspal coverage by partial or full crowns, bonded restorations have been proposed for internal splinting of restored teeth. Although contradictory data have been published, there is evidence that bonded amalgam or resin-based composite restorations (RBC) do not increase fracture resistance of teeth with wide occlusal-proximal cavities to values similar to sound, unrestored controls. Indirectly fabricated RBC inlays and various ceramic inlays, however, increased fracture strength to levels as high as those of sound caries-free teeth. Therefore, it is recommended that weakened teeth with wide cavities be strengthened by full cuspal coverage with cast or ceramic restorations, by bonded ceramic inlays, or by indirectly-fabricated bonded RBC composite inlays. PMID- 10850245 TI - AFM study of citric acid-ferric chloride etching characteristics of dentin. AB - PURPOSE: To determine the etching characteristics of 10% citric acid etchant with 3% ferric chloride (10-3) compared with related etchants without ferric chloride (10-0), or one of similar pH (36% citric acid). MATERIALS AND METHODS: Atomic force microscopy (AFM) was used to measure the surface recession and morphology of dentin. AFM images were taken of the same samples at seven steps of an etching and drying procedure using 10-3 (pH = 1.09), 10-0 (pH = 1.87) or 36% (pH = 1.08) citric acid solutions: at baseline, after etching 15 s, after 5-s air-drying, after rewetting, after 24-hr desiccation, after 30 s rewetting and after 24 hrs rewetting in filtered and purified water. RESULTS: 10-3 etched surfaces receded more than either 10-0 or 36% etched surfaces, and underwent greater collapse on 5 s air-drying. On rewetting all surfaces re-expanded to near their as-etched level. Following prolonged desiccation (24 hrs) all samples collapsed to the level seen after brief air-drying. 30-s rewetting did not restore the surface, but re-expansion of the collapsed collagen matrix was seen at 24 hrs. PMID- 10850246 TI - Bonding efficiency of single-bottle enamel/dentin adhesives. AB - PURPOSE: To compare the bond strength to dentin of the single-step enamel-dentin adhesive Etch & Prime 3.0 (EP3) to that of two-step systems Clearfil Liner Bond 2 (CL2), which includes a self etching primer and Gluma One Bond (GL1), a one bottle bonding agent, intended for use with the total etch technique. MATERIALS AND METHODS: Shear bond strength after a 24-hour water storage was determined for 10 specimens each bonded with the three adhesives. Maximum, gap widths were recorded microscopically after a 10-minute water storage along the margins of resin-based composite restorations bonded with each adhesive in cylindrical dentin cavities (3.5 mm wide and 1.5 mm deep). Marginal micro-morphology of the specimens bonded with EP3 was observed with the SEM. RESULTS: EP3 had an enamel bond strength similar to that of the other materials, but its bond strength to dentin was approximately only 50% of the bond strength of the two other adhesives. Similarly, the marginal performance of resin restorations bonded with EP3 in cylindrical dentin cavities was poor (average maximum gap width 5 microns) irrespective of the curing conditions tested with EP3. CL2 showed significantly smaller gaps (2 microns) along the cavity margin than EP3, whereas in the GL1 group in 7 of the 10 restorations inspected perfect gap-free margins were observed. An SEM investigation of different steps throughout the EP3 application and processing gave evidence that the product apparently conditions and wets enamel and dentin quite effectively, and that an approximately 0.5 micron thick hybrid layer is formed as a coupling zone between dentin (intertubular and peritubular) and restorative resin. PMID- 10850247 TI - 1-year clinical evaluation of one-step in non-carious cervical lesions. AB - PURPOSE: To evaluate the 1-year clinical retention rate of restorations of non carious cervical lesions placed using a "one-bottle" adhesive (One-Step) and one of two low modulus resin-based composite (RBC) restorative materials (AEliteflo or Bisco Glaze) or a microfilled RBC (Silux Plus). MATERIALS AND METHODS: 105 non carious cervical lesions in 14 patients, mean age 60 years, were restored using either AEliteflo, Bisco Glaze or Silux Plus with One-Step dentin bonding agent. The dentin surface was lightly roughened with a fine diamond stone, cleaned with pumice and water and restored according to the manufacturers' instructions. Patients were recalled at 6 months and 1 year, and the restoration sites examined. RESULTS: At 6 months all restorations were present. At 1 year, five restorations were missing (two Silux, three AEliteflo), three of which were in one patient. A cumulative retention rate of 95% was recorded. A small number of restorations exhibited marginal discoloration, but this was believed to be due to excess RBC on unetched enamel. PMID- 10850248 TI - Bond strengths of new simplified dentin-enamel adhesives. AB - PURPOSE: To compare the in vitro shear bond strengths (SBS) of five simplified dentin adhesives. The tested hypothesis was that the recently introduced simplified adhesive systems would have similar or higher SBS than an existing simplified acetone-based adhesive used as a control. MATERIALS AND METHODS: 100 flat bonding sites were polished to 600-grit on the labial surface of bovine incisors mounted in acrylic resin. 50 teeth were ground to expose enamel, while the remaining 50 specimens were prepared to expose middle dentin. The specimens were randomly divided into five equal groups to be treated with simplified dentin adhesives: Dentastic Uno, EasyBond, Gluma One Bond, One Coat Bond, and One-Step (control). A composite post was bonded to each treatment area. After thermo cycling, enamel and dentin shear bond strengths were determined using an Instron testing machine and the data were submitted to statistical analyses. RESULTS: Mean enamel bond strengths ranged from 14.6-28.4 MPa. One Coat Bond had the highest mean enamel SBS, but it was not significantly higher than those of Gluma One Bond and Dentastic Uno. EasyBond and One-Step had statistically similar mean enamel SBS and these were significantly lower than the mean enamel SBS of the other three adhesives. For dentin, mean SBS ranged from 14.8-21.7 MPa. Dentastic Uno had the highest mean dentin SBS, but it was not significantly greater than those of One Coat Bond and Gluma One Bond. Although One Step had the lowest mean dentin SBS, it was not significantly different from those of either EasyBond or Gluma One Bond. PMID- 10850249 TI - Microleakage of Class V resin-based composite restorations using five simplified adhesive systems. AB - PURPOSE: To evaluate microleakage at enamel and dentin margins of Class V resin based composite (RBC) restorations using five simplified adhesive systems, one self-etching adhesive, three commercial one-bottle adhesives and one experimental one-bottle adhesive. MATERIALS AND METHODS: Class V cavities (3 mm x 2 mm x 2 mm) were prepared in sound human molars with occlusal margins in enamel and gingival margins in dentin/cementum. Etch & Prime 3.0 (Degussa), Single Bond (3M), PQ1 (Ultradent), Prime & Bond NT (Dentsply) and Experimental BEH (Dentsply DeTrey) were applied strictly according to manufacturers instructions. All adhesive systems were applied on etched substrates, except for the self-etching adhesive Etch & Prime 3.0. Cavities were restored with Z100 RBC. After finishing and polishing, teeth were thermo-cycled (x 700, 5-55 degrees C, 60 s dwell time). Specimens were coated with nail varnish, immersed in silver nitrate for 2 hours and sectioned longitudinally with a diamond disc. The extent of leakage was measured and ranked using a 0-4 scale. RESULTS: Statistical analysis using Kruskal-Wallis test revealed significantly higher leakage scores (P < 0.001) for Etch & Prime 3.0 on enamel when compared to all other adhesive systems. Regarding dentin margins none of the systems completely eliminated microleakage. PQ1 had significantly lower scores (P < 0.05) when compared to Etch & Prime 3.0 and Single Bond. No statistically significant difference was observed for the other groups. PMID- 10850250 TI - Water sorption of several bonding resins. AB - PURPOSES: To determine, over the period of 1 month, (a) the degree of water sorption, and (b) the change in dimension, of four bonding resins and one low viscosity resin. MATERIALS AND METHODS: Six specimens of bonding resin (Prisma Universal Bond 3, All-Bond 2, Bondwell LC, and LB Bond from the Clearfil Liner Bond II system) and a low viscosity resin (Protect Liner F) measuring 5 mm long, 5 mm wide and 3 mm thick were made. Each specimen was weighed immediately after curing and placed in distilled water at 37 degrees C. Weights at 1, 2, 3, 7 days and 2, 3 and 4 weeks were obtained. Specimens were measured initially and at the above times. After 4 weeks, specimens were dried for 2 weeks and remeasured for size and weight. Then, the specimens were placed in a vacuum for approximately 4 hours and the weights and dimensions were remeasured. Values were converted to a mean percentage of the original dry weight and dimension and analyzed using one way ANOVA and Fisher's PLSD test. RESULTS: All materials absorbed most water in the first week. Prisma Universal Bond 3 stabilized within 2 days, LB Bond stabilized within the first week, whereas the remaining materials showed statistical increases in weight for 2-3 weeks (P < 0.05). After 4 weeks, LB Bond and All-Bond 2 resins demonstrated an approximate 8% weight increase, and the least increase was observed for Protect Liner F and Prisma Universal Bond 3 at about 3%. After desiccation, Protect Liner F and All-Bond 2 returned close to their original weight, and the remainder weighed less than initially. Dimensional changes were similar to changes in weight; LB Bond showed the greatest dimensional change over the 4 weeks (5%) and Protect Liner F the least change. PMID- 10850251 TI - Effect of water, saliva and blood contamination on bonding of metal brackets with a 4-META/MMA/TBB resin to etched enamel. AB - PURPOSE: To investigate the influence of contamination by water, human saliva, and blood on the bonding of metal brackets with a 4-META/MMA/TBB resin to etched enamel. MATERIALS & METHODS: For compressive shear bond strength measurements, the surfaces of bovine enamel were prepared either by etching with 37% phosphoric acid solution for 10, 30, or 60 s and then dried with oil-free compressed air for 10 s, or by contaminating with water, human saliva, and blood. Brackets were applied with Super Bond under loads of 200, 400, or 600 g. The bonded samples were immersed in water for 1 d or thermo-cycled for 500 cycles, and the mean shear bond strengths were compared using two-way ANOVA and Scheffe's multiple comparisons test at P = 0.05. RESULTS: The bond strengths to enamel etched for 60 s were independent of the variable load, regardless of the type of contamination. A short etching duration provided higher bond strengths than extended etching of samples contaminated with saliva and blood. The bond strengths to enamel etched for 10 s after thermal stress and immersion in water were from 11.4 to 30.4 MPa. The samples contaminated by saliva showed the lowest bond strength, and thermal stress did not reduce the bond strengths. PMID- 10850252 TI - Bonding amalgam to a resin-modified glass-ionomer base. AB - PURPOSE: To determine which surface treatment of a resin-modified glass-ionomer (RMGI) will result in optimal bond strength to amalgam when used with an amalgam adhesive. MATERIALS AND METHODS: Fifty discs of Fuji II LC (4 mm thick) were cured in acrylic testing holders and randomly divided into 5 groups of 10 each. The surface of the glass-ionomer was treated in 1 of 5 ways: (1) no treatment (control) (2) no etch, Amalgambond (AB) primer and AB adhesive (3) citric acid etch, AB primer, and AB adhesive (per manufacturer's instructions) (4) 37.5% phosphoric acid etch, AB primer and AB adhesive, and (5) microetch, 37.5% phosphoric acid etch, AB primer and AB adhesive. Amalgam was condensed onto the treated RMGI surface. Samples were stored in water at 37 degrees C and tested at 24 hours for shear bond strength. The data was analyzed statistically by ANOVA followed by Scheffe post hoc tests for significance. Glass-ionomer discs were also treated as described in each group, sputter-coated and examined using a scanning electron microscope. RESULTS: No significant improvement in bond strengths to amalgam was observed with any of the treatments of the RMGI surface. Amalgambond significantly increased the bond strength of the amalgam to the RMGI. PMID- 10850253 TI - Bonding to caries-affected dentin using self-etching primers. AB - PURPOSE: To evaluate resin bond strengths to caries-affected versus normal dentin using three self-etching primer adhesive systems. MATERIALS AND METHODS: Carious lesions were excavated and then bonded with one of three self-etching primer adhesive systems (Clearfil Liner Bond 2, Clearfil Liner Bond 2V, A.R.T. Bond). A crown was then built up using a resin composite to a height of 4 mm. Five to six serial slabs approximately 0.7 mm thick were sectioned perpendicular to the bonded surface. The bonded caries-affected or normal dentin areas were isolated to create an hourglass configuration with a cross-sectional area of approximately 1 mm2. The specimens were subjected to tensile stress at a crosshead speed of 1.2 mm/min. The bonded interfaces to caries-affected and normal dentin were examined with an SEM. RESULTS: Clearfil Liner Bond 2 and Clearfil Liner Bond 2V produced high bond strengths to normal dentin, but significantly lower bonds than to caries-affected dentin. (P < 0.05). For A.R.T. Bond, there was no statistically significant difference between types of dentin. All three bonding systems produced thin (+/- 0.5-1.5 microns) hybrid layers in normal dentin. When used on caries-affected dentin, all self-etching primer systems produced hybrid layers that were twice as thick as those produced in normal dentin. PMID- 10850254 TI - Protective effects of resin impregnation on demineralization of enamel. AB - PURPOSE: To determine if resin penetrated into the enamel etch pattern, in the absence of a polymerized outer surface film, could reduce the degree of demineralization of enamel subjected to a simulated caries challenge, and to evaluate whether the addition of fluoride to the resin would enhance reductions in demineralization. MATERIALS AND METHODS: Enamel surfaces of extracted human incisors were acid-etched. One-half of the etched area was treated with an unfilled bonding resin, while the other one-half was left untreated as a control. In another group, this same procedure was followed except the unfilled bonding resin contained fluoride. The applied resin was aggressively air thinned to ensure oxygen inhibition throughout the external surface film thickness. The thinned film was visible light-cured and the area was wiped with an ethanol swab to remove the inhibited layer. The specimens were exposed to a buffer solution of pH = 4.7 for 4 days, and were sectioned and examined by polarized light microscopy and microradiography. RESULTS: In each of the two test groups, the demineralization of the resin-treated side was significantly lower than the control side (P < 0.015). Under the conditions of this study, the experimental fluoride resin did not produce statistically significant reductions in demineralization compared with the non-fluoride resin. PMID- 10850255 TI - Indian Health Service: forging a frontier in dental health. PMID- 10850256 TI - Chicago clinic meets native needs. PMID- 10850257 TI - Posts, cores and crowns. PMID- 10850258 TI - Communication breakdown. PMID- 10850259 TI - Searching for consensus. AB - Although caution is expressed by some in relying solely on caries detector dye for diagnosis until more research is done, many aspects of microdentistry are bringing exciting potential to operative dentistry. Perhaps pediatric dentists will find microdentistry the most promising because of its application for long lasting sealants and less traumatic restorations. To dentistry's benefit, fewer injections and less frequent drilling are predicted to create a whole new generation of nonphobic adults. PMID- 10850261 TI - So, you want to buy a practice. PMID- 10850260 TI - Dental historians. Keepers of the flame. PMID- 10850262 TI - Microdentistry: the new standard of care? Part 3. Is air abrasion safe? AB - Practicing microdentistry is not for the weak-spined dentist. It takes guts to face the possible challenges and resistance described in this report. Those who truly believe microdentistry will become the standard of care remain its best advocates. They are better equipped to manage any resistance encountered from peers, boards, insurance companies and patients. But to avoid legal and board troubles, always obtain informed consent from patients of indications, costs, risks of providing and not providing treatment, and alternative treatments and their costs. The debate over whether microdentistry will become the standard of care, replacing traditional restorative dentistry, continues. Most of those who hold onto the tried and true methods of traditional diagnosis and treatment are probably trying to err on the side of caution. Most of those who embrace microdentistry are not unscrupulous opportunists but are probably trying to provide the care that represents the highest quality in their opinion. Best case scenario, traditionalists and microdentists can coexist and dialogue peacefully within their communities and through their dental organizations to navigate the future of optimal treatment. Worst case scenario, attorneys and insurance companies will profit more than ever while the division in dentistry over microdentistry widens. The shakeout will prove interesting. PMID- 10850263 TI - Mediation and peer review. PMID- 10850264 TI - Simple subgingival impressions. PMID- 10850265 TI - All-ceramic restorations: classification and clinical evaluations. AB - In the search for alternative and esthetic restorative materials, many all ceramic systems have been introduced for the general practitioner. They are used as veneers, inlays/onlays, crowns, and as enamel/dentin bonded partial or total coverage without macroretention. This article describes a classification of the different commercial all-ceramic systems and gives a review of their clinical durability. Reasons for failures are given for the different restorations. Fracture is the main reason for failure, especially for all-ceramic crowns and inlays. The frequency of secondary caries contiguous to resin composite luted ceramics is very low. Wear of the luting agent, called ditching, is not an enduring clinical problem. The use of certain ceramic materials as well as luting agents have been shown to be contraindicated, especially in molar teeth. Newer reinforced ceramics showed better durability then the earlier fired ceramic reconstructions. PMID- 10850266 TI - Resin composites in the post-amalgam age. AB - Resin-based composites are now being used as either amalgam substitutes or amalgam alternatives for the direct placement of box-shaped, stress-bearing restorations in posterior teeth. The expected longevity of these restorations is 8 years. With amalgam substitutes, preservation of enamel and dentin and restoration of tooth form and function must be warranted for the full length of the envisaged service life. In addition, with amalgam alternatives, the restoration must be, and must remain, imperceptible at a normal talking distance. The limiting factor with amalgam substitutes is the elevated risk of secondary caries, which is a result of the marginal openings that are unavoidably associated with the nature of the operative technique. Restorations in permanent teeth using amalgam substitutes most likely fail in some critical aspect of the Swiss Dental Society quality guidelines. With amalgam alternatives, the high cost and the demanding operative technique remain the main criticisms. However, amalgam alternatives, if they are placed using a sophisticated operative technique resulting in perfectly adapted restorations, meet the high expectations outlined in the Swiss Dental Society quality guidelines and fit the clinical concept of the post-amalgam age. PMID- 10850267 TI - Dentin bonding--state of the art 1999. AB - The adhesion of restorative materials to the hard components of tooth structure has been a goal pursued by many researchers ever since Buonocore established the foundation for adhesive and preventive dentistry. Based on the industrial use of phosphoric acid to obtain better adhesion of paints and resin coatings to metal surfaces, Buonocore proposed that phosphoric acid could be used to transform the surface of enamel to "render it more receptive to adhesion." Subsequent research indicated that the formation of taglike resin prolongations into the enamel microporosities was the leading bonding mechanism of resin to phosphoric acid etched enamel. The enamel bonding agents of the 1960s and 1970s progressively evolved into complex multibottle or universal adhesives in the early 1990s, which were designed to bond to enamel, dentin, composite, amalgam, porcelain, and non precious metal. Although bonding to enamel has been a dependable technique, bonding to dentin still represents an overwhelming task because dentin is a naturally wet organic tissue penetrated by a tubular maze containing the odontoblastic process, which communicates with the pulp. This intrinsic moisture may actually benefit the chemistry of the newest adhesive systems, which must be applied on moist dentin to be effective. In fact, the collapse of the collagen that occurs on air-drying may prevent the adhesive monomers from penetrating the network of nano-channels formed by the dissolution of hydroxyapatite crystals between collagen fibrils. In view of the complexity of the mechanisms associated with these mechanisms, the objective of this review article is to summarize the most recent concepts in dentin bonding. PMID- 10850268 TI - Indirect composite resin materials for posterior applications. AB - Indirect composite resin restorations were introduced a number of years ago as possible alternatives to traditional metallic or ceramic-based indirect restorations. However, the earlier formulations did not provide evidence of improvement in mechanical and physical properties over chairside-placed direct composite resin materials. Because they required more tooth structure removal than direct restorations, their use became unpopular and was abandoned by most clinicians. Over the past few years, a new class of composite resin indirect materials has surfaced in the profession. Various technologies have been suggested as reinforcement mechanisms. Fibers, matrix modifications, and an assortment of innovations have been proposed for enhancing indirect composite resin restorations. Applications are from inlay restorations all the way to multi unit fixed prostheses. This manuscript summarizes some of the progress made in this area. When available, data is presented to provide clinicians with guidelines and indications for the use of these materials. PMID- 10850269 TI - Resin cements: into the 21st century. AB - Current trends in dentistry are largely dependent on adhesives. The use of indirect restorations in cosmetic dentistry often requires predictable adhesive luting systems. This article discusses the properties and performance of today's resin luting systems. Solubility, wear resistance, strength, and microleakage play an important role in the performance of a cement. Some critical handling issues are addressed and information about current resin cements is provided. PMID- 10850270 TI - Immediate implant surgery: ten-year clinical overview. PMID- 10850271 TI - Optimization of post-replantation healing for avulsed permanent teeth in children. PMID- 10850272 TI - Two radicular cysts associated with endodontically treated primary teeth: rationale for long-term follow-up. AB - Although radicular cysts are relatively rare, they do occur. In addition, it appears that these lesions present in association with endodontically treated primary teeth. As shown in this paper and previous reports, cysts can be associated with a variety of pulp therapies. These lesions can lead to bony expansion and resorption. Radicular cysts may also cause displacement and damage to the developing permanent dentition. Given the severity of these sequelae it is prudent to recommend regular radiographic examination of primary teeth that have undergone pulp therapy. PMID- 10850273 TI - Investigating child abuse. PMID- 10850274 TI - Management of endodontic emergencies. PMID- 10850275 TI - Diagnosis is the key. PMID- 10850276 TI - Dental office urgencies and emergencies--how we would manage them. PMID- 10850277 TI - How we would manage a patient who can't breathe. PMID- 10850278 TI - How we would manage a patient who won't breathe. PMID- 10850279 TI - How we would manage a patient who has too much breathing. PMID- 10850280 TI - Herceptin: breaking new ground. PMID- 10850281 TI - Perceptions of Herceptin: a monoclonal antibody for the treatment of breast cancer. AB - In September 1998 Trastuzumab (Herceptin) became the second monoclonal antibody approved for the treatment of a malignant condition, and the first antibody approved for the treatment of a solid tumor. It is a mouse-human chimeric antibody that produces anti-tumor effects by blocking the HER2-neu receptor, and can also interact with human immune cells to effect antibody dependent cell mediated cytotoxicity. Pivotal trials in breast cancer showed that it has activity as a single agent in a subset of patients whose tumors greatly over express HER2, but results were even more impressive when it was used in combination with chemotherapy. It should also prove to be useful in the treatment of subsets of patients with other adenocarcinomas whose tumors over-express HER2. PMID- 10850282 TI - Review: dendritic cell immunotherapy for melanoma. AB - Melanoma is a particularly aggressive malignant tumour of the skin that is influenced by genetic, environmental and physiological elements. Since current therapy for melanoma is limited and associated with high toxicity and side effects, development of alternative approaches is imperative. The importance of dendritic cells (DCs) in immunity against tumours is now well established. DC immunotherapy for melanoma is possible but must be considered in terms of effectiveness and clinical viability. The source of DCs to be used in adoptive therapy as well as the nature and method of delivery of the priming antigen are important factors. The most suitable DC appears to be cells derived by culture from hemopoietic progenitor cells (HPC) in bone marrow or DC progenitors in peripheral blood. Generation of an effective anti-tumour immune response will be dependent upon the presentation of multiple melanoma-specific antigens by both major histocompatibility complex (MHC) class I and class II molecules and stimulation of both tumour-specific cytotoxic T lymphocytes (Tc) and T helper type 1 (Thl) cells. Different techniques for delivery of the priming antigen offer different advantages. DCs can be pulsed with peptide, protein or tumour cell lysate, transfected with viral vectors or naked nucleic acid and tumour/DC hybridomas can also be generated. Repeated antigen administration into neighbouring lymph nodes appears to be the most effective method for promoting a systemic anti-tumour response. Adjuvant therapies can also enhance immune responses and lead to total tumour clearance. The importance of DC immunotherapy in clinically different stages of disease will also be an important consideration. PMID- 10850283 TI - Dexamethasone, all trans retinoic acid and interferon alpha 2a in patients with refractory multiple myeloma. AB - Few effective regimen are available for patients with refractory multiple myeloma (RMM). Generally, responses are scarce and disease free survival is very short. We developed a new therapeutic option in these patients using dexamethasone (40 mg/m2, i.v., daily, days 1 to 4), all-trans retinoic acid (45 mg/m2, po, daily, days 5 to 14) and interferon alpha 2a (9.0 MU, daily, subcutaneously, days 5 to 14). The treatment was administered every 21 days for 6 cycles. In a pilot study, 12 patients, heavily treated with chemotherapy and radiotherapy and in some cases with interferon, were allocated to receive the afore mentioned treatment. Response was observed in 10 patients (83%). With a median follow-up of 36.1 months (range 27 to 41), seven patients remain alive and disease-free without any treatment. Two patients were failures and have died due to tumor progression. Toxicity was mild and all patients received treatment according to the planned doses of drugs. The use of biological modifiers in combination with dexamethasone offer a safe and effective therapeutic option in patients with refractory multiple myeloma. More studies are warranted to define the role of this type of treatment. PMID- 10850284 TI - Update on the treatment of cancer with multiple immunotherapy. PMID- 10850285 TI - Paclitaxel (Taxol): an inhibitor of angiogenesis in a highly vascularized transgenic breast cancer. AB - Paclitaxel (Taxol), a promoter of microtubule polymerization and a radiosensitizing agent, is one of the more active anticancer drugs in the current treatment of solid tumors. In this study, we show that paclitaxel possesses an antiangiogenic property associated with a down-regulation of vascular endothelial growth factor (VEGF) in a highly-vascularized transgenic murine breast cancer (Met-1). Paclitaxel, at non-cytotoxic doses of 0, 3 and 6 mg/kg/day, was administered intraperitoneally for 5 days to nude mice bearing the Met-1 breast tumor. Extent of intratumoral angiogenesis, as indicated by microvessel tortuosity and microvessel density, was significantly reduced by paclitaxel in a dose-dependent manner. Paclitaxel also suppressed expression of VEGF in the Met-1 cells transplanted in nude mice or maintained in cell culture. These results indicate that antiangiogenesis associated with a down-regulation of VEGF is an additional mode of action of paclitaxel. PMID- 10850286 TI - Carcinoembryonic antigen as a target for radioimmunotherapy of human medullary thyroid carcinoma: antibody processing, targeting, and experimental therapy with 131I and 90Y labeled MAbs. AB - The poor prognosis of patients with advanced medullary thyroid carcinoma (MTC) has prompted a search for new treatment modalities. In this report we explore the characteristics of carcinoembyronic antigen (CEA) as a target for radioimmunotherapy (RAIT) of MTC, with respect to antibody processing, targeting, and experimental therapy. In vitro studies showed a high level of CEA expression on the cell surface of the MTC cell line TT. MAbs bound to the cell were predominantly retained for several days, although there was also a significant level of internalization and catabolism. Immunohistology of frozen sections of tumor xenografts demonstrated that approximately half of the cells were darkly stained, however, some cells expressed little or no CEA. In biodistribution studies in nude mice bearing TT tumors, the mean percent injected dose per gram of tumor observed at three days post injection (time of maximum uptake) of 125I MN-14 was 19.7%. When the MAb was labeled with 88Y, a residualizing label, a much higher accretion, 50.5%, was observed at the time of maximum uptake (7 days). Significant anti-tumor effects were seen at the maximum tolerated doses of 131I- and 90Y-MN-14, compared with relatively rapid tumor growth in untreated animals or those treated with the same dose of control MAbs. Importantly, it was observed that 90Y-MN-14 yielded significantly improved therapeutic efficacy in comparison to 131I-MN-14, which may have important implications for design and conduct of future clinical trials for the treatment of MTC. PMID- 10850287 TI - Adenovirus-mediated TNF-alpha gene transfer induces significant tumor regression in mice. AB - Recombinant adenovirus vectors are highly efficient at in vitro and in vivo gene delivery. The in vitro infection of a mouse colon adenocarcinoma cell line MCA-26 with the adenovirus AdV-LacZ can reach a maximal 75% of infectivity at an MOI of 1000. Intratumoral injection of AdV-LacZ (2X10(9) pfu) resulted in substantial gene transfer in nearly 70% of MCA-26 tumors. After the in vitro infection of AdV TNF-alpha, infected MCA-26 cells showed significant secretion of TNF-alpha (45 ng/ml/10(6) cells) in tissue culture. The secretion peaks at day 2 and is diminished at day 4 following the viral infection. Infected MCA-26 tumor cells secreting TNF-alpha significantly reduced their tumorigenicity in syngeneic BALB/c mice. In mice bearing small tumors, intratumoral injection of 2X10(9) pfu of AdV-TNF-alpha virus with a repeated booster treatment resulted in complete regression of three tumors and significant diminution of the other two with a mean tumor-weight of 0.16 g; this is in contrast to 0.85 and 1.62 g for tumors injected with the control AdV-pLpA and PBS respectively (p < 0.01). Mice with complete tumor regression further developed protective immunity against the second challenge of MCA-26 inoculation. In mice bearing large tumors, this treatment also caused significant inhibition of tumor growth with a mean tumor weight of 0.65 g vis-a-vis 3.05 g for tumors injected with the control AdV-pLpA. On the contrary, in mice bearing large tumors, the treatment of tumors with pCI TNF-alpha delivered by the gene gun did not induce significant tumor inhibition. These results indicate that the adenoviral delivery of TNF-alpha gene is more efficient than the particle-mediated gene gun device, and that adenovirus mediated cytokine gene therapy may be a useful approach in the clinical management of human solid tumors. PMID- 10850288 TI - [111In-DPTA-D-Phe1]-octreotide scintigraphy in the management of patients with advanced renal cell carcinoma. AB - Somatostatin receptor scintigraphy using the 111In-labelled somatostatin analogue octreotide (Octreoscan) was performed in 9 patients with metastatic renal cell carcinoma. In total 11 scintigraphies were performed. Positive tumor uptakes were observed in 9 patients. The results of the octreotide scans were correlated to diagnostic CT and/or X-ray images. Forty (59%) out of 68 known tumor localizations were visualized with the octreotide scan. A second scan following therapy was performed in two patients. These patients showed progressive disease despite treatment and also exhibited intensified uptakes at octreotide scintigraphy. One false positive lesion was observed in the 40 lesions visualized in scintigraphy. It was concluded that renal cell carcinoma expresses somatostatin receptors, as could be visualized with Octreoscan scintigraphy. The scintigraphic technique can be used as an instrument for in vivo characterization of the disease. The data could also form a basis for future investigations regarding the possible therapeutic effect of octreotide in the management of renal cell cancer. PMID- 10850289 TI - Dosimetry for radioimmunotherapy: a rapidly evolving field. PMID- 10850290 TI - The beginnings of radioiodine therapy of metastatic thyroid carcinoma: a memoir of Samuel M. Seidlin, M. D. (1895-1955) and his celebrated patient. AB - Emerging from a stimulating encounter over fifty years ago between Dr. S. M. Seidlin and a celebrated patient at Montefiore Hospital in New York City are a number of findings that bear significantly on the contemporary practice of medicine relating to targeted radioisotope therapy. In 1943, Seidlin administered radioiodine to this patient, who was hyperthyroid although previously thyroidectomized, but who had several metastases from adenocarcinoma of the thyroid which localized the radioisotope. Seidlin recognized early that some thyroid metastases would take up radioiodine (i.e., function), but only after the normal thyroid gland was ablated, an essential preliminary procedure before radioiodine therapy should be administered, the clinical practice followed to this day. He held that removing the normal thyroid increased TSH production and eliminated the gland's competition for radioiodine, inducing the metastases to function. From 1942 until his death in 1955, Seidlin and his group followed many patients having metastatic thyroid carcinoma, conducting fruitful investigations concerned with the induction of function, dosimetry, and the occurrence of leukemia in some massively treated patients. PMID- 10850291 TI - Internal dosimetry in the use of radiopharmaceuticals in therapy--science at a crossroads? AB - Because of expanding therapeutic applications, radionuclide dosimetry is in a stage of reference "man" based system to systems for "patient-specific dosimetry." Although the challenges for the latter system are substantial, patient-specific radiation dosimetry has greater relevance to treatment planning. PMID- 10850292 TI - Preparation and in vitro studies of [125I]IUDR-T101 antibody conjugate. AB - Idoxuridine labeled with 125I was conjugated to polylysine. This conjugate was then coupled to the carbohydrate side chains of T101 monoclonal antibody (anti CD5). The immunoreactivity, cell retention, cytotoxicity, and intracellular localization of this conjugate was tested in CCRF-CEM cells (CD5 positive). The conjugate had 68% immunoreactivity. The retention of 125I by CCRF-CEM cells was higher for the conjugate than for T101 directly labeled with 125I and more of it localized in the nucleus than did the 125I-labeled T101. The 125I IUDR-polylysine T101 conjugate was more cytotoxic than the 125I-labeled T101. In conclusion, the conjugation of [125I]IUDR to T101 is feasible, and preferential targeting of the 125I to the nucleus is obtained. PMID- 10850293 TI - A phase II study of [90Y] yttrium-capromab pendetide in the treatment of men with prostate cancer recurrence following radical prostatectomy. AB - There is currently no curative therapy for men who have disseminated prostate cancer following failed radical prostatectomy. The purpose of this trial was to investigate systemic radioimmunotherapy in these men. Eight patients with occult metastatic prostate cancer following radical prostatectomy as evidenced solely by a rising serum PSA and evidence of soft tissue lesions outside the prostatic fossa detected by an [111I]indiumcapromab pendetide scan received an infusion of 10 mg of capromab pendetide labeled with 9 mCi/m2 of [90Y]yttrium. Serum PSA was used to measure response rate. There were no complete or partial responses by PSA criteria. Significant unexpected bone marrow toxicity developed in the first 6 of 8 patients treated. The last two patients received co-infusion of edetate calcium disodium in an effort to decrease marrow suppression. In these two patients less marrow toxicity was seen. Repeat 111In-capromab pendetide scans were uninterpretable due to grossly altered whole-body biodistribution of the radioimmunoconjugate. Retrospective analysis of serial PSA values after closure of the study showed a decrease in the log slope PSA for seven of eight patients following radioimmunotherapy, with a statistically significant change in the mean log slope (p = 0.01). The clinical significance of this small but measurable change is uncertain. We conclude that radioimmunotherapy for occult metastatic prostate cancer using 90Y-capromab-pendetide at the dose described does not lower serum PSA, is associated with significant hematologic toxicity, and leads to complexation of the immunoconjugate following subsequent capromab pendetide infusion. PMID- 10850294 TI - Preclinical evaluation of chimeric L6 antibody for the treatment of Kaposi's sarcoma with radioimmunotherapy. AB - L6 is a murine IgG2a monoclonal antibody with panadenocarcinoma reactivity. Chimeric L6 (ChL6), the variable region of murine L6 combined with a human IgG1 constant region, has been used in clinical trials for the delivery of radioimmunotherapy to patients with breast cancer. AIDS-associated Kaposi's sarcoma (KS), a malignancy of vascular endothelium, may be an excellent candidate for systemic radioimmunotherapy because KS is well vascularized and radioresponsive. Because ChL6 has been noted to bind vascular endothelium, our hypothesis was that ChL6 will recognize and bind KS tumors making this a potentially useful antibody for the treatment of KS with radioimmunotherapy. To test this hypothesis, 4 human KS spindle cell cultures established from cutaneous punch biopsy specimens (KS-MR, KS-NO, KS-JD and KS 6-3E) and one well characterized human KS cell line (KS Y-1) were assessed for L6 immunoreactivity. All 5 cell cultures were L6 positive by immunohistochemistry. KS Y-1 cells grown as nude mouse xenografts were also L6 positive by immunohistochemistry. Competitive binding assays performed on the KS Y-1 and KS 6-3E cell cultures showed high density and high affinity cell binding. Biodistribution experiments performed on nude mice with KS Y-1 xenografts demonstrate tumor targeting by ChL6. These findings indicate that ChL6 may be a useful antibody for the radioimmunotherapy of KS. Future experiments will assess the therapeutic efficacy of radiolabeled ChL6 with and without concurrent systemic radiosensitizing chemotherapy. PMID- 10850295 TI - In vitro and in vivo enhancement of canine pulmonary alveolar macrophage cytotoxic activity against canine osteosarcoma cells. AB - The combination of chemotherapy with immunotherapy may offer an advantage over either therapy alone and provide a greater potential for total tumor eradication. Monocyte/macrophage-mediated tumor cell killing is a major mechanism of the host's defense against primary and/or metastatic neoplasia. We evaluated the tumoricidal activity against canine osteosarcoma cells of canine pulmonary alveolar macrophages (PAM) exposed in vitro to two recombinant canine (rc) cytokines (rcTNF alpha and rcIFN gamma). We also evaluated the in vivo tumoricidal activity of PAM from dogs treated with the macrophage activator, liposome-encapsulated muramyl tripeptide-phosphatidyl-ethanolamine (L-MTP-PE) alone or in combination with doxorubicin (DOX). This study demonstrated that rcTNF alpha and rcIFN gamma significantly enhance in vitro canine PAM cytotoxicity against canine osteosarcoma cells, and that PAM from dogs treated with DOX + L-MTP-PE have enhanced cytotoxic activity against osteosarcoma cells when compared to dogs treated with DOX or L-MTP-PE alone. These findings support the rationale for combining a chemotherapy agent with an immunotherapy agent for the treatment of metastatic disease, and suggest a role for TNF alpha and IFN gamma as agents for stimulating the antitumor activity of macrophages. PMID- 10850296 TI - Technetium-99m tetrofosmin single photon emission computed tomography in the evaluation of suspected lung cancer. AB - Technetium-99m-tetrofosmin is a radiopharmaceutical employed for myocardial imaging, which has recently emerged as useful in the visualization of tumors. In this study technetium-99m-tetrofosmin was evaluated for its accuracy in differentiating malignant from benign pulmonary lesions, and in detecting mediastinal node metastasis due to lung cancer. Eighty-one patients with a solitary lung lesion on the chest radiograph and/or CT scan were submitted to chest single photon emission computed tomography after technetium-99m-tetrofosmin injection (740 MBq i.v.). The scintigraphic findings were correlated to the final histopathological diagnosis, demonstrating abnormal tracer accumulation in 51 of 54 malignant lesions (sensitivity 94%) and in 4 out of 27 benign conditions (specificity 85%), yielding an accuracy of 91%. Mediastinal lymph-node involvement was evaluated in 35 patients with non small cell lung cancer who underwent mediastinoscopy and/or surgery. Tetrofosmin accuracy (89%) was significantly higher than that of CT (69%, p < 0.05); the false negative scintigraphic results were in nodes sized less than 1 cm. In conclusion, technetium-99m-tetrofosmin imaging is useful in distinguishing malignant from benign pulmonary lesions, and in non-invasively assessing mediastinal node metastases from non small cell lung cancer, especially in patients with enlarged nodes by CT scan. PMID- 10850297 TI - Tumor cell growth suppression by tannic acid. AB - Tannic acid was cultured together with tumor cells that had originated from human malignant tumors (HCT-15 & AGS). Significant suppression of tumor growth was observed. The 50% suppression was observed at the concentration between 50 micrograms/ml and 12.5 micrograms/ml. When tannic acid was injected into HCT-15 cells by electroporation at 1180 mu p and 200 ohm, the suppression rate was 34.3% at the concentration of 50 micrograms/ml in HBS solution. The suppression rate of cells only in contact with tannic acid solution for one hour under the same conditions as used for electroporation, was 26.1%. Histographic findings suggested that tannic acid almost completely blocked the S phase of the cell cycle. PMID- 10850298 TI - Efficient recombination of Lym-1 scFv gene using multiple doubly-restricted DNA fragments. AB - In order to improve radioimmunotherapy of lymphoma, a Lym-1 single-chain antigen binding (scFv) protein molecule was produced. Because the commonly used polymerase chain reaction (PCR) method frequently causes unexpected mutations, we developed a non-PCR method for scFv gene assembly. The method involved a stepwise linkage of doubly-restricted DNA fragments and re-digestion of the resultant concatamers. Using this strategy, the Lym-1 scFv expression gene was readily constructed without mutations. The recombinant gene was cloned into an expression vector and scFv protein was expressed. The method can be used for other genes or DNA recombination. PMID- 10850299 TI - Pretargeted peptide imaging and therapy. PMID- 10850300 TI - Pretargeting with the affinity enhancement system for radioimmunotherapy. AB - The pretargeting technique referred to as the Affinity Enhancement System (AES) uses bispecific antibodies and radiolabeled bivalent haptens that bind cooperatively to target cells in vivo. Experimental and clinical data demonstrate that AES can deliver large radiation doses to tumor cells with high tumor to normal tissue contrast ratios and long activity residence time in tumors. Preliminary clinical results of radioimmunotherapy of medullary thyroid carcinomas and lung cancers look promising. PMID- 10850301 TI - Radioimmunotherapy of acquired immunodeficiency syndrome (AIDS) associated lymphoma. AB - Standard therapy for AIDS associated NHL (AANHL) is toxic and often ineffective. Radioimmunotherapy (RIT) is an appealing alternative to chemotherapy because of the radiosensitivity of NHL and the ability of the Lym-1 monoclonal antibody to target therapeutic irradiation to NHL while relatively sparing normal tissue. A Phase I/II study of 90Y-2IT-BAD-Lym-1 was designed specifically for RIT of AANHL. The first patient has been treated with 15 mCi (7.5 mCi/m2) of 90Y-2IT-BAD-Lym-1, after an imaging dose of 111In-2IT-BAD-Lym-1. Before RIT, AANHL in the maxillary sinus extended into the oral cavity and axillary adenopathy was present. Imaging showed excellent accumulation of 111In-2IT-BAD-Lym-1 in the tumors. Substantial shrinkage of the oral lymphoma was observed 18 hours after the therapy dose of 90Y-2IT-BAD-Lym-1 and axillary adenopathy had disappeared by one week after RIT. Transient Grade IV myelosuppression was the only notable toxicity. Further RIT cycles were precluded by development of an antibody response (HAMA) against Lym 1. This novel preliminary study has shown that Lym-1 can target AANHL and produce significant tumor regression thereby providing encouragement to proceed with additional patients. PMID- 10850302 TI - Synthetic, implantable, biodegradable polymers for controlled release of radiosensitizers. AB - INTRODUCTION: Synthetic, implantable, biodegradable polymers offer the sustained local release of disparate therapeutic agents for the treatment of human malignant brain tumors. The role of polymeric devices for the local delivery of radiosensitizers remains unexplored, however. We therefore quantified the release of the representative radiosensitizers IUdR (5-iodo-2'-deoxyuridine), tirapazamine (3-amino-1,2,4-benzotriazine-1,4-dioxide) and etanidazole [N-(2 hydroxyethyl)-2-nitro-1-imidazole-1-acetamide] from the [(poly(bis(p carboxyphenoxy)-propane) (PCPP):sebacic acid (SA) (PCPP:SA ratio 20:80)] polymer. METHODS: For measurements of controlled release, triplicate polymer discs were incubated for known intervals in 2 ml 0.1 phosphate-buffered saline, pH 7.4, 37 degrees C. Using a predefined schedule, the supernatant fractions were systematically removed and replaced with fresh solution. The supernatant fractions were measured. The cumulative percentage of the loaded drug that appeared in these serial supernatant fractions was plotted vs. time. The percentage of the drug that was loaded into each polymer and that was released vs. time was fit to the power function of the form y = (a) x tb, where y is the cumulative released agent, a and b are constants and t is time (days). RESULTS: The IUdR was released over an interval of approximately one week, while the release of the tirapazimine persisted for over 100 days. The etanidazole was released most rapidly, over a period of hours. Modeling of release showed that regardless of percentage loading of the drug, the monoexponential function showed high correlation of the fit of the plot of the release vs. time. CONCLUSIONS: These results suggest that the hydrophilicity and percentage loading of the drug predominantly determine the rate of release. Based upon these results, IUdR and tirapazamine warrant preclinical testing for radiosensitization of human malignant brain tumors via the synthetic, implantable, biodegradable polymeric devices. PMID- 10850303 TI - Synthetic, implantable polymers for IUdR radiosensitization of experimental human malignant glioma. AB - BACKGROUND: Recently, polymeric controlled delivery of chemotherapy has been shown to improve survival of patients with malignant glioma. We tested the delivery of IUdR via polymers for radiosensitization of experimental intracranial human malignant glioma. To assess efficacy, we measured the in vitro release, the in vivo delivery of IUdR and the resultant radiosensitization of experimental human U251 glioblastoma xenografts. METHODS: In vitro: To measure release, increasing (10%, 30%, 50%) proportions of IUdR in synthetic [(poly(bis(p carboxyphenoxy)-propane) (PCPP):sebacic acid (SA) polymer discs were serially incubated in buffered saline and the supernatant fractions were assayed. In vivo: To compare local vs. systemic delivery, mice bearing flank xenografts had intratumoral or contralateral flank IUdR polymer (50% loading) treatments. Mice bearing intracranial (i.c.) xenografts had i.c. vs. flank IUdR polymer treatments. Four or 8 days after implantation of polymers, mice were sacrificed and the percentage tumor cells that were labeled with IUdR was measured using quantitative microscopic immunohistochemistry. For comparisons of radiosensitization, mice bearing i.c. xenografts had i.c. vs. flank IUdR polymers and cranial fractionated external beam irradiation (2 Gy BID x 4 days). RESULTS: In vitro: Increasing percentage loadings of IUdR resulted in higher percentages of release: 43.7 +/- 0.1, 70.0 +/- 0.2, and 90.2 +/- 0.2 (p < 0.001 ANOVA) for the 10, 30, and 50% loadings, respectively. In vivo: For the flank tumors, both the ipsilateral and contralateral IUdR polymers resulted in similarly high percentages labeling of the tumors vs. time. For the ipsilateral IUdR polymers, the percentages of tumor cellular labeling after 4 vs. 8 days were 45.8 +/- 7.0 vs. 40.6 +/- 3.9 (p = NS. For the contralateral polymer implants, the percentages tumor cellular labeling were 43.9 +/- 10.1 vs. 35.9 +/- 5.2 (p = NS) measured 4 vs. 8 days after implantation. For the i.c. tumors treated with extracranial IUdR polymers, the percentages of tumor cellular labeling were low: 13.9 +/- 8.8 and 11.2 +/- 5.7 measured 4 and 8 days after implantation. For the i.c. tumors having the i.c. IUdR polymers, however, the percentages labeling were comparatively much higher: 34.3 +/- 4.9 and 35.3 +/- 4.0 on days 4 and 8, respectively. For the i.c. tumors, examination of the percentage cellular labeling vs. distance from the implanted IUdR polymer showed labeling was highest closest to the polymer disc. Radiosensitization: For mice bearing i.c. tumors and receiving flank vs. intracranial IUdR polymer treatments, the survival after external beam irradiation was significantly higher for the intracranial treatments: 49 + 8.9 vs. 80 + 4.1 (p = 0.03) days, respectively. CONCLUSIONS: Implantable biodegradable polymers provide the local, controlled release of IUdR and result in the high, local delivery of IUdR to experimental intracranial human malignant glioma. The local delivery and labeling result in improved survival following radiotherapy. This technique holds promise for the local delivery of IUdR for radiosensitization of human brain tumors. PMID- 10850304 TI - Comparison of melanoma antigens in whole tumor vaccine to those from IIB-MEL-J cells. AB - Immunotherapy for melanoma shows promise. Our previous whole tumor (WT) vaccine was noted to have positive clinical effects. We have now developed a new, safer melanoma vaccine that is derived from IIB-MEL-J tissue culture (TC) cells. In this study, we compare by Western blot analyses the antigens in the WT vaccine to antigens in the TC vaccine. Sera from 12 WT vaccine recipients, 8 melanoma patients who received no immunotherapy, and 8 controls served as a source of antibodies to investigate potential antigens in the vaccines. Three major antigenic peptides with approximate molecular weighs of 46, 40, and 36 kDA were present in both vaccines, while two other antigenic peptides with approximate molecular weighs of 68 and 48 kDA were present only in the TC vaccine. The reaction was similar between the patients who received the WT vaccine and those who did not receive the vaccine. Some of the individuals who did not have melanoma showed some reaction, but not to the extent of the melanoma patients. The intensity of immunostaining was greater for the TC vaccine when compared to the WT vaccine, indicating that these proteins are in a higher concentration in the TC vaccine. This new vaccine from IIB-MEL-J tissue culture cells provides a higher yield and a much more consistent source of potentially clinically relevant antigens without risk of infection or contamination by other irrelevant materials. PMID- 10850305 TI - In vivo comparison of CHX-DTPA ligand isomers in athymic mice bearing carcinoma xenografts. AB - Monoclonal antibodies (MAbs) labeled with radiometallonuclides via metal chelators are being investigated in the laboratory for use in clinical trials. The biodistribution of 111In- and 88Y-labeled antibody (MAb B72.3) using two isomeric forms (CHX-A and CHX-B) of the 2-(p-isothiocyanatobenzyl)-cyclohexyl DTPA was compared in athymic mice bearing LS-174T tumors, human colon carcinoma xenografts. CHX-(A or B)-125I-DTPA-B72.3 was co-injected in all athymic mice to assess if the chelate conjugation altered the properties of MAb B72.3. In vitro studies demonstrated maintenance of integrity and immunoreactivity for both radioimmunoconjugates. The in vivo analysis, however, indicated major differences between the two isomer forms. In fact, the 88Y-CHX-A-DTPA radioimmunoconjugate demonstrated over the 7-day study period, a more efficient and stable tumor localization as well as a slower blood clearance rate than the CHX-B-DTPA chelate conjugate, suggesting a greater in vivo stability. Differences were also evident in critical normal organ uptake: no significant increase in liver- and spleen- or bone-to-blood ratios was observed when the CHX-A-DTPA chelate was labeled with indium or yttrium. The results described here demonstrate that the CHX-A-DTPA chelate conjugate can be considered more suitable than the CHX-B-DTPA isomer form when radiometallonuclides are coupled to an MAb. PMID- 10850306 TI - Angiogenesis and growth of murine colon carcinoma are dependent on infiltrating leukocytes. AB - We determined whether the angiogenesis and growth of murine colon carcinomas growing in the wall of the cecum is dependent on infiltrating leukocytes. Syngeneic BALB/c or SCID mice were treated with a myelosuppressive, maximally tolerated dose of doxorubicin. Parental or multidrug resistant CT-26 colon carcinoma cells were implanted into the cecal wall 3 days after the second intravenous injection of doxorubicin. Control mice developed large, well vascularized tumors, whereas doxorubicin-pretreated mice did not. Intravenous injection of spleen cells from normal BALB/c or SCID mice one day prior to tumor cell implantation reversed the decreased vascularity and tumorigenicity. The production of proangiogenic molecules and microvessel density in tumors directly correlated with the number of infiltrating leukocytes, suggesting that tumor infiltrating leukocytes are essential to angiogenesis of murine colon carcinomas. PMID- 10850307 TI - Tannic acid raises survival rate of mice bearing syngeneic tumors. AB - Tannic acid which was found earlier to have growth suppressive activity against cultured tumor cells, was given to Balb/c mice p.o. The mice had been inoculated with syngeneic tumor cells (Meth/A) i.p. When tannic acid was suspended in drinking water and given daily at doses of approximately 875 mg/kg/day and 1750 mg/kg/day, the respective survival rates were 59% and 48%, with that of the control mice being 29%. To study the cytotoxicity of tannic acid, we administered the tannic acid to mice at 875 and 1750 mg/kg/day for 35 days. The weight gain for each dose was lower than that of the control, although the difference was not significant. When tannic acid was given at 8750 mg/kg/day, the weight gain was significantly lower than the control. A histologic study did not show any pathological findings in the kidney, liver, or lungs in the mice given tannic acid at the above doses. PMID- 10850308 TI - About the old and the new: gallium-67 scintigraphy to rituximab for lymphoma. PMID- 10850309 TI - Rituximab immunotherapy. AB - Certain treatments such as allogeneic bone marrow transplantation or the more recent technique of nonablative donor lymphocyte infusions may offer curative opportunities to a small number of patients with low grade lymphoma. For the majority of patients, though, this disease remains incurable. Therefore, management must balance toxicity with efficacy. Rituximab is a new form of treatment with a highly favorable toxicity profile, and its short course of therapy on an outpatient basis has a positive effect on quality of life. Rituximab and related radioimmunotherapy strategies represent new directions for the treatment of non-Hodgkin's lymphoma. Other forms of therapy, including vaccines or modulators of cell activity, promise to further improve the therapeutic index for lymphoma by increasing the specificity of treatment with less toxicity. An added bonus of rituximab is its potential to help unlock some of the mysteries of cellular regulation and dysregulation of normal and malignant B cells. PMID- 10850310 TI - Rituximab immunotherapy for non-Hodgkin's lymphoma. AB - Rituximab, a chimeric monoclonal antibody targeting the CD20 antigen on B lymphocytes, is now the standard treatment for relapsed, low-grade or follicular non-Hodgkin's lymphoma. PMID- 10850311 TI - Gallium-67 scintigraphy in the management: Hodgkin's disease and non-Hodgkin's lymphoma. AB - Gallium scintigraphy has an important role in the management of patients with lymphoma. It contributes to patient management by detecting residual disease or relapse after treatment, monitoring response during therapy, and providing prognostic information. PMID- 10850312 TI - Immunotherapeutic potential of dendritic cells generated in long-term stroma dependent cultures. AB - Long term cultures (LTC) producing dendritic cells (DC) have been established from spleen. A well developed stromal cell layer supported production of DC in numbers suitable for experimentation. Cells had obvious membrane pseudopodia and could be collected from culture every 2-3 days. Large cells produced in LTC stained with fluorescently labelled monoclonal antibodies specific for DC such as 33D1, and M1/70 which is specific for DC and myeloid cells. These staining patterns confirmed the presence of DC within the LTC population. LTC-DC were tested and shown capable of migration in vivo in B10.A(2R) mice following footpad inoculation. Most cells entered the spleen and a small number entered popliteal lymph node. LTC-DC have migratory capacity comparable with control spleen lymphocytes. LTC-DC were tested for capacity to induce an anti-tumour immune response after exposing cells to tumour cell membranes. LTC-DC pulsed with BL/VL3 tumour antigens were able to induce a BL/VL3-specific primary cytotoxic T lymphocyte (CTL) response detectable in popliteal lymph nodes and spleen of C57BL/6J mice within 6 days of priming. BL/VL3 tumour cells grew in sublethally irradiated C57BL/6J mice giving 100% mortality. Adoptive transfer of spleen cells from mice given BL/VL3 antigen-pulsed LTC-DC, two weeks previously, significantly slowed the growth of BL/VL3 tumour cells in mice. DC produced in LTC can function as antigen presenting cells (APC) when adoptively transferred into animals. Their capacity to migrate effectively, to induce a CTL response and to reduce tumour load suggests that DC grown using this in vitro system may have valuable clinical potential in humans. PMID- 10850313 TI - MSC, a new benzoquinone-containing natural product with antimetastatic effect. AB - An orally applicable fermentation product of wheat germ containing 0.04% substituted benzoquinone (MSC) has been invented by Hungarian chemists under the trade name of AVEMAR. Oral administration (3 g/kg body weight) of MSC enhances blastic transformation of splenic lymphocytes in mice. The same treatment shortens the survival time of skin grafts in a co-isogenic mouse skin transplantation model, pointing to the immune-reconstructive effect of MSC. A highly significant antimetastatic effect of MSC has been observed in three metastasis models (3LL-HH, B16, HCR-25). The antimetastatic effect of MSC- besides the immune-reconstitution--may also be due to its cell adhesion inhibitory, cell proliferation inhibitory, apoptosis enhancing, and antioxidant characteristics, also observed in our in vitro experiments. It is even more noteworthy that combined treatment with MSC and one of the following antineoplastic agents (5-FU and DTIC)--both in wide use in every day clinical practice--exhibited a significantly enhanced antimetastatic effect in appropriate metastasis models (established from C38 mouse colon carcinoma and B16 mouse melanoma respectively) as compared to the effect elicited by any component of these therapeutic compositions (MSC + 5-FU and MSC + DTIC) administered alone. The results show that the fermented wheat germ extract (MSC) has more than an additive effect and synergistically enhanced the metastasis inhibitory effect of both antineoplastic agents studied till now. It is also worthy of mention that the synchronous treatment with MSC profoundly decreased the toxic side effects of the applied antineoplastic agents (decreased weight loss etc). Based on the biological effects of MSC--shown to be non-toxic by subacute toxicology studies- this product may be used as an adjuvant in the therapy of malignant neoplasia and other diseases caused by or following immune-deficiency. PMID- 10850314 TI - Immune response to haptenized tumor antigen as possible mechanism of anticancer action of hypoxic bioreductive agents at low doses. AB - Possible anticancer effect mechanism of hypoxic bioreductive agents (HBA) at low nongenotoxic doses are reviewed. Experimental and clinical investigations show the process which can develop step-by-step when injecting HBA into the tumor bearing organism resulting in the stimulation of antitumor immunity: HBA activation in hypoxic tumor tissue, conjugation of activated HBA with proteins of tumor cells, antigen processing, presentation of neoantigen epitops in association with major histocompatibility complex-I and cytolysis of these tumor cells by T-killers. The present process can be a variant of the delayed-type hypersensitivity reaction in the tumor region. The direct regulating influence of HBA on immunocompetent and/or tumor cells as a result of interaction of these drugs with superficial or intercellular receptors is supposed to be realized additively with this process. It is concluded that the ability of HBA to selective activation in tumor tissue and formation of immunogenic conjugates with tumor proteins can be a starting-point for developing drugs with immuno modulation anticancer properties. PMID- 10850315 TI - A hypothesis on the biochemistry of spontaneous remissions of cancer: coupling of oxidative phosphorylation and the remission of cancer. AB - Aggressively growing tumors are highly dependent on the blood glucose supply. Whenever the blood glucose supply is high they grow rapidly and whenever the blood glucose supply is low they grow slowly. In cases of bonafide reports of spontaneous remissions and regressions of cancer, this tumor regulatory mechanism fails and the tumor grows rapidly and steadily despite a low blood glucose supply. This results in the collapse of the tumor system and its removal by the immune system. In lay language, it can be compared to a car that can be driven safely at 60 mph, but if it is driven at 200 mph, it will turn over and stop. Accelerated tumor growth can be induced in many ways such as: (a) reducing tumor mass which will result in a compensatory tumor growth; (b) administering hormones; (c) increasing body temperature; (d) reducing blood cortisol level; and (e) temporarily suppressing cellular immunity. A simple example would be a case of a patient with advanced and widespread cancer. If the tumor mass is reduced at once it will result in compensatory tumor growth and if hypoglycemia is initiated just prior to reducing the tumor mass and the hypoglycemia is maintained, it could possibly be followed with a brief tumor growth and the remission of cancer. The suggested procedure has not been tested or proven to be correct as yet; it is not recommended unless done under careful supervision. PMID- 10850316 TI - Planar gamma camera quantitation of 123I, 99mTc or 111In in the liver and spleen of an abdominal phantom. AB - Three planar, gamma camera methods for quantitating radiopharmaceuticals, such as radiolabeled antibodies, were investigated. Iodine-123 (123I), technetium-99m (99mTc) or indium-111 (111In) in the liver and spleen of an abdominal phantom were assessed in this study. In the first approach, the number of counts detected in a "single image" of the liver or spleen was used to measure radionuclide content using an attenuation correction factor (ACF) calculated from data obtained without radionuclide in the background volume of the phantom. In the other two methods, radionuclide content was derived from either the geometric mean (GM) of counts in conjugate, opposed images ("global conjugate") or in individual, opposed pixels of the conjugate, opposed images ("pixel conjugate") of the liver and spleen. Both of the conjugate image methods were corrected for attenuation with a first order ACF derived from a transmission image. The influence of background radionuclide on the accuracy of quantitation was studied by filling the background volume of the phantom with water containing 7 or 14 percent of the concentration of the radioactive water placed in the liver and spleen. The best estimates of radionuclide content were obtained by quantitation from the GM of counts in conjugate images of the liver and spleen. Radionuclide content of the liver and spleen could be determined from a single image if correction for attenuation was available. In all instances, measurements were less accurate for the spleen and for either organ when 111In was used. These results further validate and extend observations reported by others and provide a basis for radiation dosimetry for these and similar radionuclides and organs in patients. PMID- 10850317 TI - Amifostine. PMID- 10850318 TI - Salivary gland protection by S-2-(3-aminopropylamino)-ethylphosphorothioic acid (amifostine) in high-dose radioiodine treatment: results obtained in a rabbit animal model and in a double-blind multi-arm trial. AB - Since differentiated thyroid cancer has an excellent prognosis, reduction of long term side effects of high-dose radioiodine treatment (HD-RIT), i.e. salivary gland impairment is important. Thus, radioprotective effects of amifostine were studied. Salivary gland function was quantified by scintigraphy both in rabbits and patients. Fifteen rabbits were studied prior to and up to 6 months after HD RIT applying 2 GBq 131I. Ten animals received 200 mg/kg amifostine prior to HD RIT, and five served as controls. Animals were examined histopathologically. Fifty patients with differentiated thyroid cancer were evaluated prospectively prior to and 3 months after HD-RIT with either 3 or 6 GBq 131I in a double-blind, placebo-controlled study. Twenty-five patients were treated with 500 mg/m2 amifostine intravenously prior to HD-RIT, and 25 patients receiving physiological saline solution served as controls. Complete ablation of the thyroid was achieved in all rabbits four weeks after HD-RIT. In control rabbits 6 months after HD-RIT parenchymal function was reduced significantly (p < 0.0001) by 75.3 +/- 5.3% and 53.6 +/- 17.4% in parotid and submandibular glands, respectively. In contrast, in amifostine-treated rabbits parenchymal function was not significantly reduced. Histopathologically, marked lipomatosis was observed in control animals but was negligible in amifostine-treated animals. In control patients, salivary gland function was significantly (p < 0.001) reduced by 40.2 +/- 14.1% and 39.9 +/- 15.3% in parotid and submandibular glands, respectively, three months after HD RIT, and 11 patients developed xerostomia. In 25 amifostine-treated patients, salivary gland function was not significantly reduced (p = 0.691), and xerostomia did not occur. Thus, parenchymal damage in salivary glands induced by high-dose radioiodine therapy can be reduced significantly by amifostine. This may improve quality of life of patients with differentiated thyroid cancer. PMID- 10850319 TI - Improved prognosis in patients with acquired immunodeficiency syndrome-related lymphoma. AB - The purpose of the present study was to evaluate the impact of modern antiretroviral therapy in patients with acquired immunodeficiency syndrome (AIDS) related lymphoma who were treated with standard chemotherapy. Twenty-nine patients with AIDS were treated with triple antiretroviral therapy, including protease inhibitors, were treated with a chemotherapy regimen involving cyclophosphamide 750 mg/m2, i.v. day 1; vincristine 1.4 mg/m2, i.v. day 1; mitoxantrone 10 mg/m2, i.v. day 1; and bleomycin 10 mg/m2, i.v. day 14. Granulocyte colony stimulating factor 5 ug/kg/day, started on day 5 of every cycle was administered to ameliorate the presence of severe myelosuppression. Complete response (CR) was observed in 21 cases (72%, 95% confidence interval; 63% to 83%). At three years the time to treatment failure (TTF) was 85%; disease free survival (DFS) 62%; and overall survival 55%. Eleven patients died secondary to tumor progression and only three patients died secondary to opportunistic infections. Chemotherapy was well tolerated, only 12% of the cycles developed granulocytopenia grade 3 and eleven episodes of infection-related granulocytopenia were observed. Delay on treatment was observed on 39 cycles (22%) N. death secondary to chemotherapy were recorded. CONCLUSION: The use of modern antiretroviral therapy improved the prognosis of patients with AIDS related lymphoma, because patients could receive adequate dose intensity of chemotherapy and the presence of opportunistic infections secondary to AIDS declines with the use of protease inhibitors. Future studies will consider the use of more intensive chemotherapy in an aim to improve the CR rate and overall survival. PMID- 10850320 TI - Expression of co-stimulatory 4-1BB ligand induces significant tumor regression and protective immunity. AB - Co-stimulatory molecules play an important role in initiating antitumor immune responses. Engineered tumor cells expressing co-stimulatory molecules have been used as cancer vaccines in both experimental tumor models and clinical trials. In this study, we cloned a cDNA gene coding for the mouse co-stimulatory molecule 4 1BBL by RTPCR. The expression vector pCI-4-1BBL was constructed by DNA recombinant technology and further transfected into a moderately immunogenic EL4 and a poorly immunogenic BL6-10 tumor cell line. Expression of the co-stimulatory molecule 4-1BBL is able to induce tumor regression of EL4/4-1BBL but not BL6-10/4 1BBL tumor cell line in syngeneic BALB/c mice. The tumor regression which is mainly mediated by CD8+ T cells further leads to protective immunity against the parental EL4 tumor. Our results thus indicate the potential utility of engineered tumor cells expressing co-stimulatory molecule 4-1BBL, especially in combination with other co-stimulatory molecules such as B7-1 in cancer vaccine. PMID- 10850321 TI - Stability of monoclonal antibodies, Lym-1 and ChL6, and 2IT-BAD-Lym-1 immunoconjugate with ultra freezer storage. AB - Use of a single lot of monoclonal antibody (MoAb) or immunoconjugate for clinical studies provides efficiency of scale and consistent characteristics for MoAb based pharmaceuticals. Lym-1, an anti-lymphoma mouse IgG2 alpha chimeric L6 (ChL6), an anti-adenocarcinoma mouse IgG2 alpha-human IgG1 chimera, and the immunoconjugate 2IT-BAD-Lym-1 were examined for stability following storage. METHODS: Lym-1, ChL6, and 2IT-BAD-Lym-1 were aliquotted with filtration and stored at -70 degrees C for up to 8.5 years. 2IT-BAD-Lym-1 stored for 6.3 years (lot A) was compared to freshly prepared 2IT-BAD-Lym-1 (lot B). MoAbs were thawed and examined yearly by gel filtration high performance liquid chromatography (HPLC), polyacrylamide gel electrophoresis (PAGE), cellulose acetate electrophoresis (CAE), and Scatchard analysis of antigen binding. 2IT-BAD-Lym-1 was evaluated by HPLC, CAE, and radioimmunoassay. To assure the in vivo significance of the in vitro studies, 2IT-BAD-Lym-1 lots A and B were labeled with 111In and their pharmacokinetics in BALB/c mice were compared. RESULTS: Lym 1 demonstrated stability over 8.5 years, providing the following ranges of data over the interval: 98-100% chemical purity (HPLC), 96-100% monomeric fraction (CAE), 8.18-8.46 antigen binding pKa (Scatchard). Similar results were obtained for ChL6 for 7.4 years. HPLC and PAGE of Lym-1 and ChL6 have not changed from original manufacturer specifications, and both MoAbs remain sterile and apyrogenic. No significant differences between 2IT-BAD-Lym-1 lots A and B were observed by in vitro evaluation or pharmacokinetics in mice. CONCLUSIONS: Lym-1, ChL6, and 2IT-BAD-Lym-1 as manufactured and stored for 8.5, 7.4, and 6.3 years, respectively, demonstrated retention of structural and functional integrity. PMID- 10850322 TI - Combination vascular targeted and tumor targeted radioimmunotherapy. AB - Rat MAb 201B, which binds to murine thrombomodulin, can deliver up to 50% of the injected dose of attached radioisotopes to the lung vascular endothelium. We have shown previously that intravenous injection of about 30 microCi of 213Bi-MAb 201B, which delivers about 15 Gy of alpha irradiation to the lung, is capable of eradicating small lung colonies (500-1000 cells) of the mammary tumor line, EMT 6. Larger tumors (> 5000 cells) were not completely cured by this vascular targeted radioimmunotherapy (VT-RAIT) approach. We reasoned that VT-RAIT might make the lung vessels serving the tumor cells more permeable, allowing MAb targeted to the tumor cells to extravasate more readily and mediate more efficient standard radioimmunotherapy (RAIT). Distribution experiments with the tumor targeted MAb 13A (RAIT MAb), following VT-RAIT, did not demonstrate a large increase in tumor uptake; however, microautoradiography did indicate that MAb 13A was distributed more evenly throughout the tumor when administered after VT-RAIT. Therapy experiments on lung tumors of approximately 5000 cells each, combining 213Bi-MAb 201B (VT-RAIT) with 213Bi-MAb 13A (RAIT) 24 hours later, resulted in a better outcome (3 cured/10 at risk) than for control groups: RAIT only (0/10), VT RAIT only (1/10), or no therapy (0/10). RAIT therapy delivered 48 hours after VT RAIT had no apparent benefit. 213Bi-MAb 201B VT-RAIT followed by 90Y-MAb 13A Fab' RAIT showed only a slight improvement in tumor cures (2/10) over that in control groups: (0/9), (0/10), (0/10), respectively. These results suggest that optimal timing, dosage, and choice of MAb for RAIT should enhance the double MAb therapy approach significantly. PMID- 10850323 TI - Radiation absorbed dose estimation for 90Y-DOTA-biotin with pretargeted NR-LU 10/streptavidin. AB - Pretargeted radioimmunotherapy permits the administration of doses of 90Y five times higher than is possible with antibodies directly labeled with 90Yttrium (90Y). These high doses of 90Y introduced new issues for dosimetry that were not encountered in prior studies using conventional radioimmunotherapy. We have addressed these issues here and correlated dosimetry estimates with observed toxicity and tumor responses. METHODS: The pretargeted radioimmunotherapy (PRIT) system employed the antibody NR-LU-10 conjugated with streptavidin, a glycoprotein clearing agent and 90Y-DOTA-biotin. A single dose of 90Y was escalated to 140 mCi/m2. Indium-111(111In) (3-5 mCi) DOTA-biotin was co-injected for gamma camera imaging and dosimetry assessment. The effect of bremsstrahlung radiation from increasing 90Y activity levels with a constant dose of 111In was studied using a phantom. Patient images identified the intestinal tract and the kidneys as potential organs at risk of clinically significant radiation toxicity. A method of measuring the activity localized in the intestinal tract was developed, and S values were calculated to estimate intestinal wall dose from radioactivity present in the intestine. Intestinal, bone marrow and renal toxicity were observed. Coefficients were derived for correlating the relationships between observed intestinal and marrow toxicity and the estimated radiation absorbed doses. RESULTS: At an 90Y:111In ratio of 50:1, bremsstrahlung radiation accounted for 12% of the counts in the images. Grade IV diarrhea was observed in patients estimated to have received 6850-14,000 cGy to the large intestinal wall. The correlation coefficient of intestinal toxicity with absorbed dose was 0.64. Myelotoxicity (measured as grade of suppression of absolute neutrophil count) correlated better with marrow dose (r = 0.72) than with the whole body dose, (r = 0.44). Delayed renal toxicity was observed in two patients 8 and 11 months following therapy. Tumor response was seen in the two patients with the highest estimated dose to tumor, 4,000-6,000 cGy. CONCLUSION: Dosimetry is feasible using 111In as a tracer in the presence of high 90Y activity. The absorbed dose estimates derived in the PRIT schema correlated moderately well with clinically observed toxicity and response. PMID- 10850324 TI - The luminol-amplified chemiluminescence of neutrophils and monocytes in patients with gastric cancer after intraoperative radiotherapy using radiosensitizer sanazole. AB - The effect of electron-beam intraoperative radiotherapy (IORT) with radiosensitizer sanazole (drug AK-2123) on the functional activity of neutrophils and monocytes in patients with gastric cancer was studied with the use of luminol amplified chemiluminescence. Sanazole was administered intravenously in a dose of 1.2 g/m2, 30 min before IORT. After resection of the tumor a single dose of 10 Gy was given to the field of regional lymph collectors and the bed of the removed tumor applying a small-size betatron MIB-6E. Cells were separated and chemiluminescence was estimated on the 1-st day before and on the 3-d day and 2-d week after IORT with sanazole. Insignificant effect on the zymosan- or phorbol-12 -myristate-13-acetate (PMA)-stimulated chemiluminescence was observed in this group, although there was found a considerable increase of chemiluminescence of neutrophils in response to zymosan and PMA in 5 from 8 patients, and chemiluminescence of monocytes--in 3 from 8 patients. Correlation between individual indices of zymosan- and PMA-stimulated chemiluminescence of neutrophils before IORT with sanazole was not high, r = 0.71, but it increased significantly to r = 0.91 on the 3-d day and to r = 0.96 on 2-d week after treatment. Correlation between individual indices of stimulated chemiluminescence of monocytes before IORT with sanazole was r = 0.90, and increased to r = 0.99 on the 3-d day and to r = 0.96 on the 2-d week after treatment. So, it is indicative of a correcting effect of the combined treatment including IORT with sanazole administration on the functional activity of neutrophils and monocytes, and the absence of functional disturbances of these cells. PMID- 10850325 TI - Effect of gold on tumor-associated antigens. AB - A gold compound, which is an anti-rheumatoid agent, was clinically applied to patients showing a high level of tumor-associated antigens. Two patients showed no clear evidence of recurrence except for a high level of tumor-associated antigens. The gold compound was injected i.m. every week at the dose of 25 mg/body for 10 times to a patient showing a high CEA level that had arisen eight years after cervical dissection for a tongue carcinoma. It was also administered every other week for 30 times at the dose of 25 mg/body to a patient showing high CA19-9 and SLX levels five years after palliative left pulmonary resection followed by radiation therapy for left pulmonary carcinoma. The CEA level dropped to the normal limit after 10 injection and remained at that level after cessation of the gold treatment. SLX and CA19-9 levels of the second patient showed lower than initial values, although SLX did not fall to the normal range after the 30 injections. No side effects such as lowering of white blood cell count, BUN elevation, or kidney dysfunction were seen. PMID- 10850326 TI - Determination of spleen size by scintigraphy. AB - Methods for determining spleen size in vivo have many clinical applications. Spleen size can be estimated accurately in vivo from measurements of length (L) and width (W) of the spleen silhouette as visualized by a scintillation camera after administration of technetium-99m (99mTc) sulfur colloid to the patient. Assuming mass and volume are proportional, spleen weight will depend on (L x W)3/2. Estimates of spleen size using these parameters obtained from scintigraphs were compared to actual determinations following splenectomy. Within the range of observations, the method had a standard deviation of 45 grams. It is safe, fast, and inexpensive. PMID- 10850327 TI - Fundamentally different. PMID- 10850328 TI - Tc-99m sestamibi scintimammography: where is it now? PMID- 10850329 TI - What to do with IL-2? AB - It has been 15 years since the first positive clinical reports of Interleukin-2 (IL-2) appeared in the medical literature, ten years since moderate dose continuous infusion IL-2 was approved in Europe, and five years since high-dose bolus IL-2 was approved for general use in the United States. IL-2 is accepted as a standard treatment used alone, or in combination with chemotherapy or biotherapy in the management of metastatic melanoma and metastatic renal cell carcinoma. Various physicians utilize high-dose bolus IL-2, moderate-dose continuous infusion IL-2, and low-dose outpatient intravenous or subcutaneous IL 2. There is still no consensus regarding the best way to deliver IL-2 alone in terms of dose and schedule of administration from a risk-to-benefit standpoint. Despite yielding higher tumor response rates, regimens that combine IL-2 with chemotherapy and/or interferon have not produced better long-term survival. PMID- 10850330 TI - Status of scintimammography and its relationship to other detection methods for breast cancer. AB - Breast cancer is the most important malignancy for women of the world's industrialized nations. It is second only to lung cancer in cancer-related mortality. Early detection is the best means of improving survival; the cornerstone of early diagnosis is mammography. Given the endemic nature of breast cancer, screening mammography has secured a routine place in health maintenance for women, although it is less than perfect. To aid in the diagnosis of malignant breast disease, other imaging modalities have evolved: ultrasound, magnetic resonance imaging (MRI), positron emission tomography (PET), and SMM. Scintimammography (SMM) is rapidly with a variety of applications for the management of breast disease. This technology has become a complementary modality to other conventional methods of breast imaging. This review will focus on the science behind SMM and how it is currently used in the management of breast lesions. PMID- 10850331 TI - The feasibility of using short-term cultures of ovarian cancer cells for use as autologous tumor cell vaccines as adjuvant treatment of advanced ovarian cancer. AB - OBJECTIVE: We have tried to establish short-term cultures of autologous tumors from patients with stage III and IV ovarian cancer, which could be used as active specific immunotherapy, (i.e., autologous vaccine) in such patients after debulking surgery & combination chemotherapy. METHODS: Between 5/93 and 11/97 the Hoag cell biology laboratory received 53 ovarian tumor samples that had been surgically excised at the time of laparotomy, and four samples of malignant ascites. Efforts were made to establish short-term tumor cell cultures as confirmed by morphology & phenotype. RESULTS: Short-term proliferating cultures were successfully established from 21/57 samples [37%] which included 8/24 [33%] successes from samples obtained at diagnosis compared to 13/33 [37%] samples obtained at the time of a relapse [p = 0.45]. The probability of successful culture was not related to tumor size for samples with a range of 0.8-34 g (mean 5.8 g). One patient was treated in the setting of metastatic disease and one in the adjuvant setting; both received repeated injections of irradiated autologous tumor cells plus granulocyte macrophage stimulating factor (GM-CSF). In one patient a delayed tumor hypersensitivity skin test converted from negative to positive. CONCLUSIONS: Short-term cultures of autologous tumor cells for use as tumor cell vaccines can be established for about one-third of patients with ovarian cancer using this methodology and the treatment approach is feasible. PMID- 10850332 TI - Combined modality therapy of A431 human epidermoid cancer using anti-EGFr antibody C225 and radiation. AB - BACKGROUND: Monoclonal antibodies (mAb) to epidermal growth factor receptor (EGFr) inhibit tumor cell proliferation and enhance cytotoxicity of chemotherapeutic agents. The purpose of this study was to investigate the interaction of the anti-EGFr antibody C225 combined with radiotherapy (RT) on EGFr expressing A431 human epidermoid cancer cells. METHODS: Cell proliferation, apoptosis, EGFr expression and phosphorylation, and clonogenic survival were assayed in vitro. A431 tumor growth inhibition and immunohistochemistry analysis of EGFr expression and apoptosis were assessed in vivo. RESULTS: C225 plus RT produced greater inhibition of A431 cell proliferation than C225 or RT alone which was corroborated by enhanced apoptosis. Similar clonogenic survival occurred following the addition of C225 to RT, although colonies were smaller in the presence of C225. C225 produced inhibition of EGF-induced phosphorylation of EGFr without concurrent down-regulation of surface receptor, which was not altered by RT. Combined treatment of mice bearing tumors demonstrated enhancement of complete regressions, reduction in time to tumor size doubling, and prolongation of survival. Significant apoptosis occurred in xenograft tumors treated with C225 with or without RT. CONCLUSIONS: These data demonstrate an interaction between C225 and RT. C225-mediated apoptosis and inhibition of EGFr phosphorylation may be critical in the interaction. Studies to define the precise influence of combined modality treatment on the EGFr signal transduction cascade need to be pursued. The combination of growth factor receptor antibodies and RT has potential application in clinical oncology. PMID- 10850333 TI - Relationships between the activity of MMP1/TIMP1 enzymes and the TH1/TH2 cytokine network. AB - The evaluation of the balance between matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) would appear to be important in cancer patients. Since the activity of these enzymes is regulated at the gene level by cytokines, we studied the serum relationships between MMP1/TIMP1 and the network of TH1/TH2 cytokines in healthy subjects to better understand how the physiological network of cytokines regulates MMP1/TIMP1 activity. Such a study could lead to suggestions for follow-up experiments to create prognostic and diagnostic indices for more efficient disease prevention programs and biotherapeutic treatments of patients. For this purpose, we determined serum levels of MMP1, TIMP1 and interleukin (IL)2, interferon (IFN) gamma, IL4 and IL10 in both healthy men and women (men and women were analyzed separately as hormones are one of the non-cytokine regulatory factors of TH1 or TH2 polarization). These cytokines make up our basic network. Cytokines within the physiological network function simultaneously so mathematical models of multivariate statistical methods were used to study MMP1/TIMP1 and TH1/TH2 network relationships. It has been suggested that mathematical modeling is the only effective way of studying the immune system as a whole. The influence of network activation, antigen presenting cells, antibody response and chemokines on MMP1/TIMP1 balance was also studied. Network activation was evaluated by measuring the levels of soluble IL2 receptors (sIL2R) and sIL6R; the influence of antigen presenting cells was evaluated by measuring serum levels of tumor necrosis factor alpha (TNF alpha) and IL1 beta; antibody response was evaluated by measuring IL5 and IL6 serum levels and the influence of chemokines was evaluated by measuring serum levels of IL8. Our overall results suggest that there are relationships between the activity of MMP1/TIMP1 and the TH1/TH2 network in physiological conditions. These data may be useful in gaining a clearer insight into how the two systems interact and hence regulate the physiological homeostasis. Therefore, this paper provides suggestions for experimental studies on MMP1/TIMp1 enzymes and TH1/TH2 cytokines to create clinical and prognostic markers for patient evaluation. PMID- 10850334 TI - Receptor-mediated radionuclide therapy with 90Y-DOTA-D-Phe1-Tyr3-Octreotide: preliminary report in cancer patients. AB - Recent advances in receptor mediated tumor imaging led to the development of a new somatostatin analogue DOTA-D-Phe1-Tyr3-Octreotide. This new compound, named DOTATOC, has shown high affinity for somatostatin receptors, stable labeling with yttrium-90 (90Y) and favourable biodistribution in patients. The aim of this work was to evaluate acute and late toxicity and the response rate in cancer patients administered 90Y-DOTATOC. Twenty patients received three equal i.v. injections of 90Y-DOTATOC. Cohorts of 5 patients were treated starting with 1.1 GBq per cycle in escalating dosage (0.4 GBq increments) in subsequent groups. No patients showed acute or delayed major adverse reactions up to the dose of 2.2 GBq of 90Y DOTATOC per cycle (6.6 GBq total). Maximum tolerated dose has not been determined yet. One patient, after 4.4 GBq total dose, developed delayed kidney grade II toxicity. Complete and partial tumor mass reduction (CR and PR) was measured in 25% of patients along with 55% showing stable disease (SD) and 20% progressive disease (PD). These results indicate that high activities of 90Y-DOTATOC can be administered with low risk of myelotoxicity, although the radiation doses to the kidneys require careful consideration. Tumor doses were high enough in most cases to obtain objective therapeutic responses. PMID- 10850335 TI - Scintigraphic detection of breast cancer xenografts with Tc-99m natural and recombinant human alpha-fetoprotein. AB - Because adenocarcinoma of the breast expresses receptors for alpha-fetoprotein (AFP), we studied Tc-99m AFP as a radiopharmaceutical to detect breast cancer. The biodistribution of Tc-99m radiolabeled natural human AFP (full length) and recombinant domain III (DIII) of human AFP was compared to Tc-99m sestamibi and Tl-201 in a murine model of human breast cancer. Estrogen receptor positive (MCF7, T-47D) and estrogen receptor negative (MDA-MB-231, BT-20) human breast cancer xenografts were grown subcutaneously in the lateral thorax region of immunosuppressed mice (ICR SCID). Quantitative comparisons of percent-injected dose per gram of tissue (%ID/gram) and tumor to thigh ratio (T/Th) were performed at 0-60 minutes and at 24 hours following injection. For most tumors, T/Th for AFP and DIII was significantly greater than T/Th for Tc-99m sestamibi and Tl-201. In all breast cancers (BT-20, MCF7, MDA-MB-231, T-47D), Tc-99m AFP T/Th increased from 60 minutes to 24 hours, suggesting good tumor retention of this radiopharmaceutical. DIII and AFP had significantly higher %ID/gram than either Tl-201 or Tc-99m sestamibi when considered across all tumor types at both 60 minutes and 24 hours. The data suggests that localization of Tc-99m AFP in human breast cancer xenografts is initially rapid, increases with time, and is superior to Tc-99m sestamibi and Tl-201. Given its high uptake by breast cancer cells, its low non-tumor localization and its rapid renal excretion, these Tc-99m AFP preparations may be useful agents to detect human breast carcinoma. PMID- 10850336 TI - Tc-99m-MIBI scintimammography; SPECT versus planar imaging. AB - This study compares single photon emission tomography (SPECT) with planar scintimammography. 16 normal, 54 benign and 80 malignant lesions were studied (total 150). 700 MBq of Tc99m-sestamibi (MIBI) was injected intravenously. Anterior supine, right and left lateral planar images were acquired in prone position at 2 hours post injection. After this a SPECT study (64 projections, 64 x 64 matrix, 30 sec/frame, anterior 180 degrees arc) was acquired in supine position. Attenuation coefficient of 0.12 and Butterworth order 5 Nyquist 0.9 filter was applied for reconstruction. A scoring system for visualization of lesions was devised with scores of 0 or 1 as negative and 2 or 3 as positive for MIBI uptake. Scores for each group of patients were added together and then compared between planar and SPECT studies. RESULTS: Planar imaging missed 10 malignant lesions while SPECT missed only 4. Ten benign lesions showed uptake of MIBI on planar imaging while 11 showed uptake on SPECT. Cumulative scores for normal and benign groups did not show any significant difference between planar and SPECT imaging. The malignant group showed significant increase in score for SPECT. Specificity, false negative fraction and positive predictive value for planar imaging were 85.7%, 14.3% and 87.5%. These values did not show any significant difference for SPECT (84.5%, 15.5% and 87.4% respectively). Sensitivity, false negative fraction and negative predictive value for planar imaging (87.5%, 12.5% and 85.7%) was significantly different (p < 0.005) than SPECT (95%, 5% and 93.7% respectively). MIBI SPECT scintimammography improves the sensitivity, false negative fraction and negative predictive value of the planar imaging. In the absence of MIBI uptake on SPECT imaging, malignancy can be ruled out with great confidence, while only uptake should be further evaluated. PMID- 10850337 TI - Prolonged survival associated with immune response in a patient treated with Lym 1 mouse monoclonal antibody. AB - A patient with aggressive, chemotherapy-resistant non-Hodgkins lymphoma (NHL) was treated with 131I-Lym-1, a mouse antibody, on a protocol designed for serial therapy. Human anti-mouse antibody (HAMA) developed within 1 month of initial therapy. The patient also developed an antibody to the hypervariable region of the Lym-1 antibody (Lym-1 specific). Because the patient was responding to therapy, plasmaphoresis was used to reduce the level of HAMA followed by unlabeled Lym-1 calculated to be sufficient to block residual HAMA. This allowed additional therapy on three subsequent occasions over 5 months. Despite very high HAMA levels, no untoward effects from administrations of Lym-1 were observed. The HAMA response of the patient included anti-Lym-1 specific antibodies containing anti-idiotypic antibodies. The anti-Lym-1 antibody level has been sustained over the 9 year interval since 131I-Lym-1 therapy and has been associated with a uniquely long remission of the patient's disease. These observations demonstrate that, under certain circumstances, radioimmunotherapy (RIT) can be given safely and effectively despite HAMA. Anti-idiotypic antibodies could have induced an immune cascade that contributed to the prolonged disease-free survival of the patient. PMID- 10850338 TI - Phase I study of R24 in patients with metastatic melanoma including evaluation of immunologic parameters. AB - R24 is a mouse IgG3 monoclonal antibody with specificity for the disialoganglioside GD3. Most human melanomas have substantial surface GD3; in addition, a significant proportion of T lymphocytes display surface GD3. In a phase I study, we have investigated the toxicity and effect on selected immunological parameters of three dose levels of R24 given intravenously daily for five days (10 mg/m2/d, 30 mg/m2/d and 50 mg/m2/d) to patients with advanced melanoma. R24 administration neither consistently diminished nor augmented expression of delayed type hypersensitivity (DTH) skin reactions to anergy panel antigens or to a contact allergen dinitrofluorobenzene. R24 was infrequently found on tumor cells, or on lymphocytes from DTH biopsies, despite measurable serum levels of R24. The 30 mg/m2/d dose of R24 produced a statistically significant drop in peripheral blood lymphocytes on treatment Day 5. Likewise, on Day 5 there was a modest but statistically significant decrement in the proportion of circulating cells which were R24+. While there was one mixed response, there were no complete or partial tumor regressions in the R24 treated patients; there was no evident clinical benefit from the R24 therapy. The toxicity of the R24 at the higher dose levels can be very substantial. One patient, on the highest dose level, died on the 4th day of R24 treatment; in the absence of a plausible alternative explanation, a relationship of the death to the administered R24 must be considered. A precipitous drop in serum albumin coincident with R24 administration was found in all cases; this effect has not been previously reported with R24. PMID- 10850339 TI - Spleen peptides (Polyerga) inhibit development of artificial lung metastases of murine mammary carcinoma and increase efficiency of chemotherapy in mice. AB - There are numerous attempts to find novel anticancer drugs or to improve therapeutic protocols based on application of chemotherapeutic agents and immunomodulators (biological response modifiers, cytokines, various plant or bacterial products). Among the preparations that have beneficial effects for the cancer bearing organism are preparations of spleen peptides (Polyerga). Hence, we analyzed if treatment with spleen oligopeptides GP-1 (active substance for the manufacture of Polyerga ampoules' solution injected as 0.5 microgram/kg every second day) if given alone or combined with chemotherapy (Endoxan 50 mg/kg single i.p. dose) of mice bearing artificial lung metastases of mammary carcinoma will have an impact on the metastases count and survival of the animals. The results obtained have shown that chemotherapy reduced metastases count and increased survival of the tumor bearing mice, while the use of GP-1 alone did not affect metastases development. However, combined GP-1 treatment and chemotherapy were more efficient in prevention of the metastases development than chemotherapy alone. Thus, in mice treated with GP-1 and Endoxan, the average metastases count was four times lower than in the mice treated by chemotherapy only, while 2/12 animals were without tumor nodules in the lungs. Finally, all the animals treated by chemotherapy alone died until the 42nd day after tumor transplantation, while at the same time, only 5/10 animals died receiving combined therapy. Thus, these results give an experimental support for the use of the spleen peptides in biotherapy (or combined therapy) of cancer. PMID- 10850340 TI - Effects of a hybrid recombinant human alpha interferon (A/D) on in vitro cytotoxicity and in vivo localization of monoclonal antibody L6-cytosine deaminase conjugate in a colon cancer model. AB - L6 is an IgG2a murine monoclonal antibody which we have demonstrated binds well to HT29 human colon carcinoma cells by flow cytometry, whole cell ELISA, and mixed hemadsorption. In vitro cytotoxicity studies revealed that the monoclonal antibody L6-cytosine deaminase (L6-CD) immunoconjugate plus the nontoxic prodrug, 5-fluorocytosine (5-FC), is equivalent to 5-fluorouracil (5-FU) in its ability to kill HT29 cells. Human alpha-interferon (A/D) was able to enhance this cytotoxic effect. The I.C.50's revealed that very small amounts of L6-CD are needed for this cytotoxic effect (approximately, 5 pg/ml resulted in 50% viability). The limiting factor was the amount of 5-FC employed with L6-CD (3 microM yielded 50% cell viability). alpha-Interferon (A/D) lowered the requirement of 5-FC to 1 microM to achieve 50% cell lethality. In vivo biodistribution experiments indicated that 1 microgram of L6-CD is nonspecifically taken up by the liver and spleen and cleared rapidly from the blood. Significant localization of L6-CD to HT29 tumors occurred only when 99 micrograms of unlabeled L6-CD was added to 1 microgram of 125I-labeled immunoconjugate injected intravenously. Further augmentation of tumor/blood ratios without reduction in percent injected dose per gram of tumor was possible with the intravenous injection of 100 micrograms of anti-idiotypic monoclonal antibody 13B, 24 hours after L6-CD, which bound unreacted L6-CD and cleared it from the blood. The addition of 100,000 U of alpha interferon (A/D) given intraperitoneally every day increased the clearance of L6 CD by the liver and spleen, but impaired tumor localization (percent injected dose per gram). These studies demonstrated that in vivo localization of the L6-CD conjugate to HT29 tumors could be optimized by injecting excess L6-CD followed by an equal amount of L6 anti-idiotype mAb 13B, 24 hours after L6-CD. PMID- 10850341 TI - Dysregulated cytokine production by monocytes from chronic lymphocytic leukemia patients. AB - The serum levels of proinflammatory cytokines (Tumor Necrosis Factor, Interleukin 1, Interleukin-6) in B-cell Chronic Lymphocytic Leukemia (CLL), and secretion of these cytokines by monocytes from CLL patients has been studied in 27 CLL patients and 20 normal subjects under unstimulated and stimulated conditions. It has been observed that monocyte function is impaired and deficient in CLL patients. The cytokine secretion though decreased in unstimulated cultures, was found to reach normal levels as far as IL-6 and TNF are concerned, indicating a possible role of an in vivo suppressive factor. The observed defect in cytokine production may have an important implication for the immune system of the patients. PMID- 10850342 TI - Treatment of drug resistance: biotherapy or chemotherapy? PMID- 10850343 TI - A phase I and pharmacokinetic study of CBT-1 as a multidrug resistance modulator in the treatment of patients with advanced cancer. AB - CBT-1, a natural product, was studied in an escalating dose Phase I clinical trial with doxorubicin at 60 mg/m2. CBT-1 was administered by mouth at doses from 200 mg/m2 to 600 mg/m2. The drug was given for 7 days and doxorubicin administered intravenously on day 6. The MTD was determined to be 500 mg/m2 although some patients did tolerate 600 mg/m2 with moderate nausea and occasional vomiting. Side effects were otherwise mild in the 23 patients treated. Pharmacokinetic determinations in an additional 11 patients demonstrated that CBT 1 did not significantly alter the pharmacokinetics of doxorubicin. In this Phase I study, 25 of 34 patients were evaluable for response and 5 patients demonstrated tumor shrinkage. PMID- 10850344 TI - Chemosensitivity of advanced larynx carcinoma cells in vitro and significance of multidrug resistance markers in these tumors. AB - Thirty cases of previously untreated advanced larynx carcinoma were checked for in vitro chemosensitivity and presence of the resistance markers viz. P glycoprotein (P-gp) glutathione-S-transferase-pi (GST-pi) and protein kinase C (PKC) overexpression. The cytotoxicity testing was done using MTT assay and the resistance markers were checked by immunohistochemical methods using monoclonal antibodies. The drug combinations employed in MIT assay were 5FU* + MTX*, 5FU + cisPt*, 5FU + Mito*, cisPt + Mito and MTX + Mito (*5FU = 5Fluorouracil, MTX methotrexate, cisPt-cisplatin and Mito = mitomycin C). No statistically significant correlation was observed between resistance to the above drug combinations and presence of the resistance markers under consideration. A statistically significant correlation was observed between node positivity and expression of resistance markers which indicates that presence of one or more of these markers in these tumors may be considered as a negative prognosis marker. CisPt-Mito was found to be the most effective drug combination in vitro, in the cases studied. PMID- 10850345 TI - Disease stage prognostic indices in the early clinical screening of colorectal cancer patients. AB - It is a truism to state that in cancer the extent to which the disease has spread (stage) is probably the most important factor determining patient prognosis and must be given prime consideration in evaluating and comparing different therapeutic regimes. Because of this, clinical screening tests (such as a rectal exam proctoscopy and colonoscopy), if tissue that is not normal is found, should include analyses contributing to the patient disease stage classification. However, the difficulty involved in this, principally due to the absence of reliable early prognostic indices of the disease stage, makes the cancer patients' treatment full of problems and risks. By a retrospective statistical study on pre-surgery peripheral blood immunological parameters of our groups of colorectal cancer patients and healthy subjects, we evaluated our previously suggested possibility of defining stage prognostic indices by parameter blood range values, evaluating their ability in the stage classification compared to the pTNM method. We have investigated the serum levels of various cytokines and cytokine receptors, leukocyte surface markers, peripheral blood mononuclear cell (PBMC) cytokine production and PBMC proliferative response. Statistically significant correlations between a variety of these immunological parameters and the disease stage were found, but as a clinical patient screening of all parameters is quite expensive, to identify the greatest stage weighting parameter and the respective blood range values, we performed a multivariate statistical analysis. We found that the blood ranges of IL-4 serum level and the PBMC proliferative response to anti-CD3 monoclonal antibody (mCD3) stimulus may be reliable prognostic indices which may contribute to an early disease stage classification in colorectal cancer patients, since they seem to be valid as the pTNM method. Moreover, as the immunological prognostic indices could be a useful tool to evaluate the patient immune response, they may also improve the definition of the patient's stage classification for the selection of treatment and restaging procedures for the evaluation of the treatment benefit and recurrent disease. PMID- 10850346 TI - Immunosuppressive applications of PHA and other plant mitogens. AB - Phytohemagglutinin was prominent in the evaluation of the immunosuppressive effects of mitogenic lectins that started in 1965 and continued for two decades. Basic studies elucidated the inhibitory actions of PHA on humoral and cellular immune responses in mice, rats, and guinea pigs. Some suppressive effects of this and other mitogens including lentil lectin and Con A were demonstrated on renal, skin, pancreas, and heart allograft rejections in mice, rats, and dogs. In addition to their inherent suppressive activities, these substances have been shown to potently augment the suppression generated by conventional agents. More relevant to tolerance-inducing modulations, the Rigas group showed that in vitro incubation of the parental spleen cells with PHA in the F1 hybrid model before giving them to the F1 mouse virtually abolished the GvH responses. Their explanation was that the donor cells were rendered unresponsive by their reversion to an immature state in which they lost the essential surface receptors. Similar spleen cell suppression was generated by systemic administration of PHA to the parental donor prior to their administration. A method is proposed here for establishing tolerance to either newly introduced or firmly established antigens applying the L4 isolectin of PHA to make non-reactive all T lymphocytes that remain functional in conjunction with full suppression by conventional agents. During the vulnerable period of drug-induced suppression, the host would be protected from infection and bleeding by full nonspecific proliferative activation of the lymphoid and myeloid systems. The establishment of allograft tolerance by preemptive inhibition of responses to newly introduced transplant antigens would be easier to achieve than the reversal of firmly established responses to antigens involved in GvH reactions and autoimmune diseases. The studies of von Boehmer and Kisielow in a transgenic mouse model confirmed that clonal deletion does occur in the thymus whereby corresponding specific thymocytes are destroyed when they encounter self-antigens. Their work suggested that tolerance to self-antigens might be established if immune responses against putative antigen peptides were blocked sufficiently that they would be released to reach the central or peripheral residence sites of their immunologically reactive clones for deletion to occur. This approach to suppressive therapy would be useful for managing the problems of allograft rejection, GvH reactions, and autoimmune disorders even if they fell short of generating complete tolerance. Suppressive treatment with mitogens would also be indicated for the immune-related group of aplastic anemias, but a discussion of these disorders will be deferred to a separate article because of certain aberrations they present. PMID- 10850347 TI - PHA for aplastic anemias: the alpha but not the omega of mitogen therapies. AB - This manuscript updates an analysis of the foremost clinical investigation of PHA for aplastic anemias and other disorders associated with impaired hematopoiesis during the years 1963-1967. Humble displayed remarkable insight in motivating the trials long before the recent era during which these disorders have become much better understood. That investigation actually consisted of a series of small pilot studies totaling only 44 patients involving all categories in the aplastic group and 15 miscellaneous disorders mostly neoplastic or premalignant. The most serious of several faults with the studies was the inadequacy of dosages applied, yet much better results were observed than might have been anticipated. Mitogens such as the L4 isolectin of PHA might be applied to treat aplastic anemias in three different ways. (1) For milder direct cytotoxicity types, a stimulating regimen should be applied to increase the production of growth factors and thereby hasten recovery from aplasia. (2) Where hematopoietic stem cell allo engraftment is needed to replace severely damaged marrow, the mitogen should be included in the preparative regimen. (3) The suppressive protocol of a mitogen that maintains total T cell activation should be added to an established drug regimen such as cyclosporin and antilymphocyte globulin in treating the immuno mediated aplastic anemias not only to provide superior suppression but to help prevent the late emergence of clonal disorders. The models of L4 isolectin application present advantages not fully offered by any of the other immuno therapies currently available: broad scope of activities, low morbidity, ease of administration, favorable cost-effectiveness, assurance of reconstituted immune competence, and high potential for cure. However, a more potent mitogen than PHA L4 not inhibited by serum glycoproteins or immunoglobulins and exhibiting a broader range of modulation would be preferred. A publication by Mann et al. in 1991 suggested that pokeweed mitogen (PWM) showing hundreds of times the potency of PHA-L4 would meet these criteria. Abbreviated reports on two of the studies in the report indicated that PWM had induced lasting remissions of cancers in dogs with nine injections in the mid-range of microgram doses over three weeks. The key study done by Mann himself demonstrated complete disappearance of canine gliomas in a carefully devised humane model applied to five dogs in which the transplantable tumor was established by extradural injection. Also impressive were lasting remissions of autonomous solid neoplasms in four dogs under the care of local veterinarians given PWM supplied by Mann. The critical aspect of these studies, both of which have been extended substantially, is the need keep the dosage within a precisely determined range to avoid loss of efficacy and possibly the occurrence of adverse effects. A review of basic studies now indicates the principle mechanism of response to be binding of PWM with tumor cells that then attract mast cells generated by stem cell factor the production of which is stimulated by the mitogen. Degranulation of mast cell granules releases TNF-alpha and IL-1 at the tumor site with resulting disappearance of neoplastic tissue in the absence of severe systemic effects. For cancer therapy, this represents a highly effective deviation from the stimulation of multiple pathways of immune response usually surmised with mitogens such as PHA-L4. This interpretation illustrates the probability that alternative plant mitogens, each with its unique properties, will likely become available to complement PHA-L4 or displace it from the prime standing as an immunomodulator. PMID- 10850348 TI - Comparison of the effects of Viscum album lectin ML-1 and fresh plant extract (Isorel) on the cell growth in vitro and tumorigenicity of melanoma B16F10. AB - Numerous findings indicate that specific plant lectins acting against cancer could be major active components of Viscum album extracts, although activity of low molecular weight components (peptides, carbohydrates and alkaloids) might be as essential for the beneficial activity of the plain plant extracts, too. Thus, active principle of Viscum album extracts is still not understood, and is difficult to be analysed because of the complex composition of the extracts and uncertainty of the standardised effectiveness (batch consistency) of the extracts. The aims of this study were to compare the concentration dependent effects of the pure mistletoe lectin (ML-1) with the fresh plant Viscum album extract (Isorel) and its different MW components on the in vitro growth of ConA stimulated lymphocytes, on the growth and tumorigenicity (artificial lung metastases development) of murine melanoma B16F10 cells, and to compare concentration dependent effects of the different types of the Viscum album extracts in vitro (applying novel type of MTT assay). The results obtained indicate that the effects of Isorel used at high dose could be result of toxic activity of the mistletoe lectins ("ML-1 like" activity). Unlike ML-1, if used at low concentrations, Isorel selectively inhibited tumor cells, due the activity of the low MW components. On the other hand, the number of tumor nodules was reduced (in comparison to the control) equally in the lungs of mice injected with B16F10 cells pre-treated in vitro with the plain Viscum album extract or any of its modifications or ML-1. Hence, it is supposed that the beneficial therapeutic effects of Isorel might result from the combined biological activity of the high and the low MW components not lectins only. Similarly, in MTT assay low concentrations of all types of the Viscum album extract showed stronger inhibiting activity for B16F10 and HeLa cells than pure ML-1. According to these results we propose a standardisation of aqueous Viscum album extracts by comparing their and ML-1 concentration dependent activity on the tumor cells in vitro applying MTT bioassay described which should be relevant for further evaluation of their active principle and for improvement of biotherapy of cancer. PMID- 10850349 TI - Tumor retention of 186Re-MAG3, 111In-DTPA and 125I labeled monoclonal antibody G250 in nude mice with renal cell carcinoma xenografts. AB - In radioimmunotherapy of solid tumors substantial gain might be achieved by carefully selecting the radionuclide and the linker connecting it to the antibody. In contrast to 131I, radiometals such as 90Y and 111In may be retained in the tumor cell after internalization of the antibody, thereby enhancing the radiation dose to the tumor. The physical properties of 186Re with 1.08 MeV beta emission (71%) and 137 keV gamma-emission (10%) seem ideal for radioimmunotherapy. In this study we investigated in nude mice with s.c. renal cell carcinoma xenografts the biodistribution and the retention in the tumor of 186Re-MAG3 labeled monoclonal antibody (mAb) G250 as compared to 111In-DTPA-G250, to 125I-G250 and to 131I-MN14 (non-specific control mAb). Radiolabeled antibody preparations were i.v. injected. Seventy two hours p.i. the biodistribution of the radiolabel was determined. Blood levels of all mAb G250 preparations were remarkably low (mean: 2.38, 1.40 and 1.43% ID/g for 125I, 111In and 186Re respectively) whereas blood levels of mAb MN14 were significantly higher (mean: 12.3% ID/g), indicating tumor processing of mAb G250. Retention of 111In in the tumor was significantly higher than of 186Re and 125I whereas retention in the tumor of 186Re and 125I did not differ significantly. CONCLUSION: In contrast to other radiometals such as 111In and 90Y, 186Re is not retained in the tumor cell. Therefore 186Re has no additional advantage for radioimmunotherapy with respect to retention in the tumor. PMID- 10850350 TI - Review of recent developments in the molecular characterization of recombinant alfa interferons on the 40th anniversary of the discovery of interferon. AB - Recombinant alfa interferons (IFN-alpha s) are approved worldwide for the treatment of a variety of cancers and diseases of virologic origin. A series of recent advances in the molecular characterization of recombinant IFN-alpha s have allowed the determination of the three-dimensional IFN-alpha 2b structure by high resolution x-ray crystallography. We review here recent developments in our understanding of the molecular and physicochemical properties of recombinant IFN alpha, including our current state of knowledge of the IFN-alpha gene family and the multiple species of human leukocyte IFN. Based on the reported three dimensional structure of IFN-alpha 2b, we propose a molecular model for the IFN alpha 2b receptor complex and predict models for the naturally occurring subtypes IFN-alpha 1 and IFN-alpha 8, as well as the synthetic, non-naturally occurring consensus IFN. Such models provide molecular insights into the mechanism of action of IFN-alpha. PMID- 10850351 TI - A sequential four-drug chemotherapy and biotherapy with interferon alpha and GM CSF--an innovative protocol for the treatment of metastatic melanoma. AB - We present the results of our chemo-biotherapy protocol for patients with metastatic melanoma. The rationale for the design of the combined therapy was induction of systemic anti-tumor immunity by: (a) priming with IFN-alpha for enhancement of tumor and histocompatibility antigen expression, (b) therapy by the 4-drug regimen (BCNU, DTIC, cisplatin and tamoxifen) for maximal tumor destruction, followed by (c) an immunomodulatory, low dose of GM-CSF. Treatment was given in cycles of three weeks: first IFN-alpha (3 x 10(6) U/day on days 1, 3, and 5); then the 4-drug regimen given according to Del Prete et al., (4); followed by GM-CSF (20 micrograms/m2/day on days 15 to 21). All patients were previously untreated by chemotherapy, and had a performance status of ECOG 0-2. Treatment was discontinued upon severe toxicity or disease progression. In responding patients--once maximal response was achieved-IFN alpha treatment (3 x 10(6) U/day, three times weekly) was continued for a period of two years or until disease progression. All 40 patients (28 males and 12 females) who received the above program were evaluable for response and toxicity and received at least two cycles of therapy. At a median follow-up of one year, 50% had achieved an objective response, with 22.5% complete responses (CR), and 27.5% partial remissions (PR). Median duration of response is 11 months (12 for CR and 9 for PR patients). Median survival for all patients is 14 months (range, 7-21), increasing to 22 months (range, 15-29) in responding patients. Activation of patients' peripheral blood Macrophages and Dendritic Cells was observed following GM-CSF treatment. The chemo-biotherapy protocol presented above--while associated with acceptable toxicity--resulted in a relatively high response rate with an increase in the number of durable responses in patients with metastatic melanoma. PMID- 10850352 TI - Clinical experience with autologous tumor cell lines for patient-specific vaccine therapy in metastatic melanoma. AB - Because of their patient specificity and proliferative capacity, tumor cell lines established from autologous metastatic melanoma tumor samples may be an excellent immunogen for patient-specific vaccine therapy. Between October 1990 and July 1996, the Hoag Cancer Center cell biology laboratory received 136 fresh metastatic melanoma samples from 122 different patients. Tumor cell lines were successfully established for 92 of 136 samples (68%), for 87 of 122 patients (71%). Successful cultures were expanded to 10(8) cells (total culture time about 8 weeks), confirmed to be sterile, irradiated, and stored frozen in aliquots of 10(7) cells. Vaccines were prepared from 72 lines, and 62 vaccines were used in 57 different patients. Subcutaneous vaccination took place on weeks 1, 2 and 3, and then monthly for a total of 6 months. A delayed tumor hypersensitivity skin test (DTH) was administered at week zero and week 4. Various adjuvants were co administered including BCG, alpha- or gamma-interferon, and GM-CSF. Patients were monitored for failure-free survival (FFS) and overall survival (OS) from the date of the first vaccination. Follow-up data is available for 52 patients, 27 who had no evident disease (NED) at the time of vaccination and 25 who had metastatic disease at the time of treatment. There were two partial responses which persisted 11.9 and 39.8+ months among the 25 patients who had detectable metastatic disease whun treatment was initiated (8%, 1 to 26%, 95%-Ci). Twenty patients had negative skin tests at week 0 and week 4; six were positive both times, and 13 converted their DTH from negative to positive, for a conversion rate of 13 of 33 (39%). Patients who received interferon-gamma and/or GM-CSF as an adjuvant had a higher rate of DTH conversion compared to patients who received other adjuvants (13 of 20 v 2 of 13, P = 0.003). For patients who were NED, nine of 19 (47%) converted their DTH test compared to four of 14 (29%) patients with metastatic disease (p = 0.33). For patients whose DTH converted from negative to positive after 3 weeks of vaccination, median FFS and OS were superior compared to patients whose DTH remained negative (19.4 v 4.0 months FFS, p = 0.0052 and 39.6 v 18.3 months OS, p = 0.0602). The autologous cell line approach to active specific immunotherapy is feasible for patients who have resectable foci of metastatic disease. Administration of such patient-specific vaccines improves survival for those patients who are NED at the time of vaccination and convert their DTH skin test, compared to those whose DTH test remains negative. PMID- 10850353 TI - Imaging tumor folate receptors using 111In-DTPA-methotrexate. AB - It is known that membrane folic acid receptors are responsible for cellular accumulation of folate and folate analogs, such as methotrexate, and overexpressed on various tumor cells. This study was aimed to develop an 111In labelled DTPA-methotrexate (DTPA-MTX) to image tumor folate receptors in vivo. DTPA-MTX was synthesized by reacting ethylenediamine with MTX. The resulting amino analogue of MTX was reacted with DTPA dianhydride in basic aqueous solution followed by dialysis. Tissue distribution was determined in breast tumor-bearing rats at 0.5, 2, 24, and 48 h (n = 3/time interval). To determine receptor mediated process 111In-DTPA-MTX was co-administrated with varying blocking doses of cold folate to tumor-bearing rats. Planar imaging and whole-body autoradiograms were performed. The data was compared to that using 111In-DTPA. In animal studies, tumor/blood count density ratios at 0.5-48 h gradually increased from 0.8 +/- 0.32 to 2.2 +/- 0.41 with 111In-DTPA-MTX. Conversely, these values showed time-dependent decrease from 1.19 +/- 0.69 to 0.56 +/- 0.10 with 111In DTPA in the same time period. Tumor/muscle and tumor/blood count density ratios significantly decreased with high doses of folic acid co-administration. Planar images and autoradiograms confirmed that the tumors could be visualized acceptably with 111In-DTPA-MTX. The results indicate the feasibility of using 111In-DTPA-MTX to image tumors through a folate receptor-mediated process. PMID- 10850354 TI - Inhibition of cell growth in culture by quinones. AB - Quinones were studied for their growth inhibitory effect on cultured malignant cells. HCT-15 cells derived from human colon carcinoma were used for these experiments. Quinones used were arbutin in the benzoquinone group, juglone and lawsone in the naphthaquinone group, alizarin, emodin, 1,8 dihydroxyanthraquinone, and anthraquinone in the anthraquinone group, and xanthone. Cultured cells were incubated with various concentrations of the quinones for four days in a 5% CO2 incubator, after which cell numbers were counted and significance of differences was analyzed by Student's t test. Anthraquinones and naphthaquinones used in these experiments were more effective than the monocyclic quinone. The 50% suppression dose was less than 12.5 micrograms/ml for them. The number of OH groups seemed to play an important role in the degree of the cell growth inhibition: anthraquinones with 2 or 3 OH groups were more effective than those with no OH group like, 9,10-dioxoanthracene and xanthone. In fact, anthraquinones with no OH group and xanthone were not significantly effective. Flow cytometric histograms revealed a specific pattern; that is, lawsone and juglone in the naphthaquinone group and alizarin and 1,8 dihydroxy-anthraquinone in the anthraquinone group blocked mainly the S phase, and emodin in the anthraquinone group blocked the G1 to S phase of the cell cycle. PMID- 10850355 TI - Antitumor effect of gold as revealed by growth suppression of cultured cancer cells. AB - Gold agents have been widely used for the treatment of rheumatoid arthritis. We studied the growth inhibiting effect of such an agent on malignant cells in vitro. HCT-15, AGS cells derived from a human malignancy, and Meth/A cells from a malignant lymphoma of Balb/C mice were cultured separately with gold agent at concentrations of 2 micrograms/ml. Four days after the cultures had been incubated in a 5% CO2 incubator at 37 degrees C, cell counts were made; and significance of differences was analyzed by Student's t test. Additionally, HCT 15 cells were cultured with gold for two days, and then the cells were analyzed by flow cytometry. The growth of HCT-15, AGS, and Meth/A cells was suppressed by gold. Fifty percent suppression was observed at a concentration between 50 micrograms/ml and 10 micrograms/ml for HCT-15 cells, between 125 micrograms/ml and 50 micrograms/ml for AGS cells, and between 125 micrograms/ml and 50 micrograms/ml for Meth/A cells. Fifty percent suppression of HCT-15 cell growth by cisplatinum was found between 50 micrograms/ml and 10 micrograms/ml. Flow cytometric findings showed a significant rise in the tetraploid peak, a mild rise in the resion between diploid and tetraploid peaks, and an increase in cells with a ploidy greater than four. These data suggest that gold blocks the S phase, G2 to M phase, and M phase as well. To observe the cytotoxicity of gold, each of 10 of 4 week-old Balb/C mice was injected s.c. at a dose of 10 mg/kg or 2 mg/kg every other day for a total of 3 injections, or was administered the gold at 30 mg/kg/day p.o. for 10 days. All mice were still alive after 20 days of observation. Cisplatinum at a dose of 10 mg/kg was also injected s.c. one time into each of 10 mice, and 60% of the animals died within 10 days after the injection. PMID- 10850356 TI - Implications of the analogy between recombinant cytokine toxicities and manifestations of hantavirus infections. AB - The etiologic hantavirus of the 1993 emergence of an acute pulmonary failure syndrome in the area around northwestern New Mexico was quickly recognized as related to the Hantaan virus responsible for the outbreak of Korean epidemic hemorrhagic fever (EHF) among UN troops in 1951. Discovery of the new disease which was named the hantavirus pulmonary syndrome (HPS) and its causative agent the Sine Nombre virus (SNV) inspired detailed comparisons between the two disorders. Major damage to the epithelial cells of the capillaries and arterioles throughout the body leading to extensive capillary leak and subsequent hypotension and shock was the common denominator. The lung capillaries and arterioles were the focus of attack that could lead to rapid pulmonary failure in HPS and the corresponding renal and retroperitoneal vessels that caused a more protracted illness in EHF, but both displayed remarkably similar peripheral blood abnormalities including abnormal mononuclear cells, immature neutrophilia, thrombocytopenia, and hemoconcentration characteristic enough to make blood smear examination a useful tool in early diagnosis. There are evidences that a heavy virus presence in the involved endothelial cells is accompanied by various mononuclear cells capable of generating potent immune response in these areas. Relevant toxic effects of systemically-administered high-dose interleukin-2 for resistant cancers include fever, chills, diarrhea, renal dysfunction, capillary leak syndrome accompanied by hypotension requiring aggressive pressor support, and occasional pleural effusions with diffuse pulmonary infiltrates and hypoxia severe enough to require ventilatory assistance. Peripheral blood mononuclear cells cultured in vitro with IL-2 secrete secondary cytokines such as IL-1, TNF alpha, and interferon-gamma (IFN-gamma). TNF-alpha, implicated in the pathophysiology of septic shock, is capable of inducing adult respiratory distress syndrome (ARDS) in experimental animals and humans. The strong similarity of these effects to the manifestations noted in the hantavirus diseases justifies the conviction that these and other cytokines involved in potent immune responses would constitute the pathogenic toxic substances predicted by perceptive early investigators of EHF. This concept is favored by clear indications that in both diseases active virus infection disappears the first few days and the ages of involvement correlate with periods of immunocompetence. The paradox of systemic injections of IL-2 that risk hantavirus type toxicities for treating renal cell carcinoma and melanoma might be avoided by giving potentially more efficacious plant mitogens like PHA as previously reported. The expanded disclosure of a collaborator's method suggesting superior potential for cancer cure involves a unique application of pokeweed mitogen that delivers various cellular and cytokine responses directly to the tumor. PMID- 10850357 TI - A mechanism of the spontaneous remission and regression of cancer. AB - Some reports of spontaneous remission and regression of cancer are preceded by a severe infection accompanied with a high fever. This paper proposes the mechanism of action of febrile infection in inducing spontaneous remission and regression of cancer and its possible implication in treating cancer patients. PMID- 10850358 TI - A revolution in the treatment of non-Hodgkin's lymphoma. AB - The use of monoclonal antibodies (MoAbs) for immunotherapy and radioimmunotherapy has ushered in a new era in the treatment of non-Hodgkin's lymphoma. PMID- 10850359 TI - Bcl-2 and drugs used in the treatment of cancer: new strategies of biotherapy which should not be underestimated. AB - The theory that an imbalance in the control of the cell cycle contributes to the appearance and progression of neoplastic disease is gaining more ground all the time. This new line of research into tumor disease is a result of the progress made in the comprehension of cell death (apoptosis) and the discovery of alterations in the apoptotic pathway in patients with cancer, which have also been correlated to disease mechanisms. Alterations in the cycle of events that brings about apoptosis can result in tumor cells resistant to chemotherapy. In fact one of the inherent risks of chemotherapy is the generation of new, more aggressive, clonal variants and destruction of healthy cells with deleterious effects on the organism. This review examines the results of studies concerning the identification of the alterations in apoptotic mechanisms in carcinogenesis and the mechanisms governing their regulation. The aim was to evaluate if such data could be of use in identifying drugs able to improve cancer treatment. PMID- 10850360 TI - Low-dose, fractionated radioimmunotherapy for B-cell malignancies using 131I-Lym 1 antibody. AB - PURPOSE: This trial was conducted to assess the toxicity and efficacy of 131I-Lym 1 in patients with either malignant B-cell non-Hodgkin's lymphoma (NHL) or chronic lymphocytic leukemia (CLL) using low-dose, fractionated radioimmunotherapy (RIT). MATERIALS AND METHODS: Thirty adult patients who had advanced B-cell malignancies (25 NHL and 5 CLL) had progressed despite standard therapy; 12 patients entered the trial with Karnofsky performance status (KPS) of equal to or greater than 60. Patients were treated with a series of intravenous doses of 131I-Lym-1 with a goal of reaching a cumulative dose in each patient of at least 300 mCi. All patients were Lym-1 reactive. Clinical responses and immediate toxicity were evaluable in all 30 patients and delayed toxicity in 26. RESULTS: Toxicity to Lym-1 antibody occurred with 28% of the 176 doses and was transient. Human antimouse antibodies (HAMA) were generated in 30% after a mean of 4 doses, but interrupted therapy in only 10% of the patients. Thrombocytopenia was dose-limiting; there were no deaths due to toxicity. Tumor regression occurred in 25 (83%) of the patients and was great enough, and durable enough, in 17 (57%) to qualify them as responders; 13 NHL patients and 4 CLL patients. Advanced disease often interrupted therapy prematurely. However, 18 patients received at least 180 mCi of 131I-Lym-1; 17 (94%) of these responded to the therapy. CONCLUSION: Although advanced disease often interrupted therapy prematurely, the results from 131I-Lym-1 therapy are clearly promising and warrant additional trials. PMID- 10850361 TI - A new chemically modified chimeric TNT-3 monoclonal antibody directed against DNA for the radioimmunotherapy of solid tumors. AB - In the last several years, our laboratory has developed a new approach to the radioimmunotherapy of solid tumors, designated Tumor Necrosis Treatment (TNT), that exploits the presence of degenerating and necrotic cells within tumors by utilizing MAbs directed against universal, intracellular antigens. The first TNT MAb developed by our laboratory, designated TNT-1, was directed against nucleosomal determinants consisting of histone H1 and DNA. Since absolute tumor accretion of MAb is a critical determinant of antitumor efficacy in radioimmunotherapy, we sought to identify new antinuclear antibodies that displayed high tumor localization properties. In the present study, we describe a murine antinuclear antibody, TNT-3, which demonstrates 3-fold higher tumor uptake than TNT-1. Because of this characteristic, a chimeric derivative designated chTNT-3 was developed and evaluated for antigen binding and tumor targeting. ELISA studies using a series of nuclear antigens confirmed that TNT-3 is directed against single-stranded DNA and does not cross react with TNT-1. Immunohistology reveals predominantly nuclear staining reactivity in human tissues and tumors. Since it was shown by our laboratory that charge modification can significantly improve the pharmacokinetic performance of monoclonal antibodies, chTNT-3 was chemically modified with biotin to generate an improved therapeutic reagent designated chTNT-3/B. Comparative studies with unmodified MAb demonstrated that biotinylation significantly shortened its clearance time in mice and produced lower normal tissue levels, while maintaining an equal amount of uptake in tumor xenografts for up to 10 days. These in vivo characteristics suggest that chTNT 3/B is an improved TNT reagent for the radioimmunotherapy of solid tumors. PMID- 10850362 TI - Expression of adhesion molecules in B-cell chronic lymphocytic leukaemia: an analysis in lymphoid compartments--peripheral blood, bone marrow and lymph node. AB - The trafficking or homing of different lymphoid subsets to particular microenvironment is mediated by specific cell adhesion molecules (CAMs) expressed on lymphocytes and endothelial cells. B-cell chronic lymphocytic leukaemia (B CLL) or Non-Hodgkin's lymphoma of small lymphocytic, B-cell type are monoclonal expansions of mature lymphocytes. The relative distribution of the tumor lymphocytes among various lymphoid compartments vary from patient to patient. Very few studies underlying this issue are available. To this effect, we have analysed the expression of LFA-1; VLA-4, ICAM-1; CD44H and CD44v6 (haematopoietic and variant form respectively) on freshly isolated lymphocytes obtained from bone marrow (BM), peripheral blood (PB) and lymph node (LN) by flow cytometry. Overall, we find strong expression of CD44H, low to moderate expression of LFA-1, negative to low expression of VLA-4 and lack of expression of CD44v6. ICAM-1 expression was observed only in patients with prominent lymphadenopathy. Higher expression of CD44H in PB lymphoid cells relative to that of BM lymphoid cells correlated with higher PB lymphocytosis (p < 0.001). Proliferating cell nuclear antigen expression in LN sections correlated inversely with VLA-4 expression on BM and PB lymphoid cells (p < 0.05). There was no significant correlation between expression of CAMs and bcl-2 protein. PMID- 10850363 TI - Susceptibility of natural killer (NK) cells to reactive oxygen species (ROS) and their restoration by the mimics of superoxide dismutase (SOD). AB - Natural killer (NK) cells are susceptible to reactive oxygen species (ROS), and lose the activity by the effects of ROS. Cancer bearing hosts usually suffer from oxidative stress (OS), and the NK-activity decreases to a significantly lower level than normal controls. Superoxide dismutase (SOD)-mimicking substances, such as protein-bound polysaccharide of Coriolus versicolor (Fr) QUEL (PSK) and iron chelating chlorine e6-Na (FeCNa), can restore the NK-activity of cancer bearing hosts, when collaborating with catalase. Incorporation of 3H-thymidine by ROS treated NK-cells is not affected, indicating that these cells are still active in the nucleic acid metabolism. Intraperitoneal administration of anti-Asialo GM1 antibody extinguished the NK-activity. NK-cells affected by ROS lost the adherence to target cancer cells in both in vitro and in vivo. ROS may change the surface charge of NK-cells to anionic, resulting in an inability of adhesion to target cancer cells which usually show the negative surface charge. PMID- 10850364 TI - The expression of cytokeratin 18 in transitional cell carcinoma comparing with hepatoma. AB - The epithelium in kidneys and urinary bladders contain CK18 as in liver cells. The modulation of cytokeratin 18 during tumor transformation in hepatoma had been previously recognized through a series of biochemical and immunological approaches. A 14 KD hepatoma related molecules was found in the previous studies. We would like to utilize the hepatoma transformation model to study the changes in CK18 in transitional cell carcinoma, using immunoblotting and western blotting techniques. The result is that transitional cell carcinoma retain their CK18 molecule. Furthermore, CK18 related molecules similar to those seen in hepatoma also present in transitional cell carcinoma. The conclusions are transitional cell carcinoma contains CK18 related proteins similar to those seen in hepatoma tissues. We suggest that this element would be responsible for the change during the malignant transformation processes. PMID- 10850365 TI - Stabilizing of cytokeratin in PLC/PRF/5 cells. AB - There are two sorted groups of cytokeratin 18 (CK18) in forms of assembly and disassembly in PLC/PRF/5 cells. A subcellular shifting is found in association with conditions of microtubule networks. The finding shows that CK18 mostly in forms of assembly, when microtubule networks are in status. The result also reveals that CK18 is relatively in forms of disassembly, while microtubule networks are disrupted in steps. It indicates that intact microtubule networks are probably a stabilizing factor of assembled CK18. It implies that CK18 is not a stable molecule when the cell is under environmental stress. PMID- 10850366 TI - Zonisamide inhibits nitric oxide synthase activity induced by N-methyl-D aspartate and buthionine sulfoximine in the rat hippocampus. AB - The antiepileptic zonisamide (ZNS) is known to be effective in protecting against epilepsy in a wide variety of animal epilepsy models and in humans with epileptic seizures, with both partial and generalized seizures. ZNS scavenges hydroxyl radicals (OH*) and nitric oxide (NO) in a dose-dependent manner. The mechanism of the antiepileptic effect of ZNS may involve protection of neurons from free radical damage and stabilization of neuronal membranes. In this study, the effect of ZNS on nitric oxide synthase (NOS) activity in the hippocampus of rats induced by N-methyl-D-aspartate (NMDA) with/without L-buthionine-[S, R]-sulfoximine (BSO) was examined. NOS activity was accelerated significantly (plus 93%) in the hippocampus 3 hours after NMDA injection (30 mg/Kg, i.p.), and (plus 220%) 3 hours after NMDA (30 mg/Kg, i.p.) injection into BSO-pretreated rats (150 mg/Kg, i.p.). NOS activity was not affected by ZNS itself. ZNS reduced NOS activity, accelerated by NMDA- with/without BSO-treatment, to the control level in the hippocampus. This suggests that ZNS may inhibit initiation and propagation of seizures by inhibiting NOS activity, and also may protect neurons from free radical damage by NO and/or OH*. PMID- 10850367 TI - In vivo receptor labeling of peripheral benzodiazepine receptor by ex vivo binding of [3H]PK11195. AB - In vivo receptor labeling of the peripheral benzodiazepine receptor was investigated using ex vivo binding of [3H]-(2-chlorophenyl)-N-methyl-N-(1 methylpropyl)-3-isoquinoline-carb ox-amide ([3H]PK11195). In autoradiographic studies, high level specific binding of [3H]PK11195 was observed in the olfactory bulb. Intravenous administration of PK11195 dose-dependently (0.03-3 mg/kg) inhibited ex vivo binding of [3H]PK11195 in the olfactory bulb. Likewise, N-(2,5 dimethoxybenzyl)-N-(5-fluoro-2-phenoxyphenyl)acetamide (DAA1106), a newly identified peripheral benzodiazepine receptor-specific ligand, dose-dependently (0.1-100 mg/kg) reduced ex vivo binding of [3H]PK11195, when administered intraperitoneally. In contrast, clonazepam, a central benzodiazepine receptor specific agonist, had negligible effects on ex vivo binding of [3H]PK11195. We propose that the ex vivo receptor binding technique we used will facilitate determination of in vivo receptor occupancy of the peripheral benzodiazepine receptor. PMID- 10850368 TI - Fas-mediated cytotoxicity by gammadelta T cells during acute rejection in xenotransplantation of spheroidal aggregate-cultured hepatocytes. AB - Xenogeneic transplantation has recently become a subject of interest for the transplantation community due to the current organ shortage, which could be partially or even totally solved by the development of this strategy. However, xenogenetic rejection remains a formidable barrier preventing such use in a clinical setting. The spheroidal aggregate-cultured hepatocytes of WKA rats were injected into the spleen of C3H mice, and quantitative assessment of transplanted xenogeneic hepatocytes using 99mTc-GSA demonstrated that hepatocytes decreased dramatically 2 days after transplantation (day 2) and few viable hepatocytes in spheroids were detected on day 3. The NK activity significantly increased on day 1, and gammadelta receptor/FasL-expressing T cells appeared on day 2. These results suggested that xenogeneic cytotoxicity consisted of gammadelta T cells through the Fas/Fas ligand system, as well as non-T-cell-mediated cellular response, in the MHC-unrestricted pathway in this intrasplenically transplanted xenogeneic hepatocyte model. PMID- 10850369 TI - Differential induction of brain heme oxygenase-1 and heat shock protein 70 mRNA in sepsis. AB - We examined gene expression of heat shock protein 70 (HSP70) and heme oxygenase-1 (HO-1), which is the rate limiting enzyme in heme catabolism and is also known as heat shock protein 32 (HSP32), in the rat brain using a sepsis model induced by bacterial lipopolysaccharide (LPS). Intraperitoneal injection of LPS (10 mg/kg) to rats caused the elevation of body temperature and white blood cell (WBC) counts as well as marked elevation of serum interleukin-6 (IL-6) level, showing the typical pathological characteristics of sepsis. In this model, HO-1 mRNA increased at 6 h after LPS administration and continued to rise until 30 h. In contrast, HSP70 mRNA increased only between 3 h and 6 h after LPS administration, returning completely to the control level by 12 h. HO-1 mRNA was expressed predominantly in the cortex and the medulla oblongata, while HSP70 mRNA was expressed mainly in the striatum. HO-1 and HSP70 mRNA levels thus showed distinctive time courses and tissue distribution in the brain, suggesting that gene expression of these heat shock proteins (HSPs) is separately regulated. PMID- 10850370 TI - Electrocardiographic changes induced by diesel exhaust particles (DEP) in guinea pigs. AB - We have previously reported that diesel exhaust particles (DEP) caused a negative inotropic effect that was followed by cardiac arrest in the isolated atrial preparation of guinea pigs. The purpose of this study was to examine the systemic effects of DEP on electrocardiographic (ECG) changes using guinea-pigs. We found that intravenously administered dimethyl sulfoxide (DMSO) extract of DEP solution induced arrhythmias and deaths via complete atrioventricular (AV) block in guinea pigs. The LD of DEP solution was 132.0 +/- 7.2 mg/kg. The coefficient of variance (CV) of LD measured by the modified Hatcher-Magnus method was relatively small (5.5%). Fractions of DEP extracted by hexane, ethanol or methanol, 4 hydroxyphthalic acid 2-methyl ester, a compound isolated from methanol extract of DEP did not induce significant ECG changes in guinea pigs. As compared with fresh DEP solution, the DMSO/DEP solution used in the present study induced similar cardiac toxicity after being stored in a freezer at 4 degrees C for 3 days. These results suggest that stable and water-soluble fractions of DEP may be responsible for cardiotoxicity. PMID- 10850371 TI - Identification, by cDNA microarray, of A-raf and proliferating cell nuclear antigen as genes induced in rat lung by exposure to diesel exhaust. AB - Diesel exhaust particles (DEP) contain various carcinogens and mutagens, and chronic exposure to diesel exhaust (DE) induces pulmonary cancer in experimental animals. However, the oncogenes involved in pulmonary carcinogenesis have not been identified. After F344 rats were exposed to DE containing 6 mg/m3 DEP for 4 weeks, oncogenes and related genes expressed in their lungs were surveyed using a new technique, cDNA microarray, and the results were confirmed by northern blot analysis. Expression of A-raf and proliferating cell nuclear antigen (PCNA) mRNAs was induced in rat lung by exposure to DE. These results suggest that A-raf and PCNA might contribute to pulmonary carcinogenesis in rats. PMID- 10850372 TI - Effects of dexamethasone on the pharmacokinetics of adriamycin after intravenous administration to rats. AB - Adriamycin was metabolized to adriamycinol by the cytoplasmic aldo-keto reductase and adriamycin and adriamycinol were further metabolized to their deglycosylated aglycones, M3 and M4, respectively, by cytochrome P450. SKF-525A (a cytochrome P450 inhibitor) and dexamethasone (a cytochrome P450 inducer) were pretreated to rats. Adriamycin, 16 mg/kg, was infused over 1-min via the jugular vein of each rat. After pretreatment with SKF-525A, the area under the plasma concentration time curve of adriamycin from time zero to the last measured time in plasma, 8 h (537 versus 155 microg x min/ml) was significantly greater, and the 24-h urinary excretion of M3 (1.65 versus 23.8 microg), and M3, M4, and their glucuronide and/or sulfate conjugates (33.7 versus 3.38 microg) were significantly smaller than those in control rats suggesting that the formation of M3 and M4 were inhibited by SKF-525A. After pretreatment with dexamethasone, the 24-h urinary excretion of M3 (94.5 versus 23.8 microg), M4 (8.43 versus 2.78 microg), and M3, M4, and their glucuronide and/or sulfate conjugates (116 versus 33.7 microg) were significantly greater than those in control rats, suggesting that the formation of M3 and M4 seemed to be induced by dexamethasone. PMID- 10850373 TI - Pharmacodynamic and pharmacokinetic studies of anthraquinone 2-carboxylic acid on passive cutaneous anaphylaxis in rats. AB - The objectives of this study are to describe the inhibitory effect of 9,10 anthraquinone 2-carboxylic acid (AQCA) on IgE-mediated passive cutaneous anaphylaxis (PCA) reaction, and the pharmacokinetics of AQCA. Pharmacodynamic assessments were performed at 0.5, 1 and 2 mg/kg (i.v.) and 5, 10 and 20 mg/kg (p.o) dose levels. In separate groups, pharmacokinetics were assessed at 5 mg/kg (i.v.) and 5, 10, and 20 mg/kg (p.o.) dose levels. Intravenous and oral administration of AQCA inhibited the PCA reaction in rats in a dose-dependent manner. The PCA-inhibitory activity of AQCA (20 mg/kg) lasted more than 12 hrs after oral administration. The oral bio-availability decreased with increasing dosage, from 96% (5 mg/kg) to 81% (10 and 20 mg/kg). The absorption after oral administration was prolonged with Tmax values ranging from 1 to 6 h; while t(1/2) (4.8-16 h) values appeared to be comparable. These results suggest that AQCA has a potent and long acting anti-PCA activity. It is likely to be therapeutically useful in the treatment of asthma. PMID- 10850374 TI - Pharmacokinetics of theophylline and caffeine after intravenous administration of aminophylline to premature neonates in Korea. AB - Theophylline has been widely used to treat apnea of premature neonates. The purpose of this study was to compare the pharmacokinetic parameters of theophylline and caffeine after intravenous administration of aminophylline to seven Korean low-birthweight neonates with apnea to those in other countries. The serum concentrations of theophylline and caffeine were measured simultaneously by high-performance liquid-chromatography (HPLC). The mean (+/- S.E.M.) birth weight and gestational period were 1190 +/- 253 g and 31.5 +/- 1.99 weeks, respectively. The mean (+/- S.E.M.) theophylline maintenance dosage was 1.28 +/- 0.15 mg/kg (given as equivalent aminophylline solution) every six hours. The mean (+/- S.E.M.) volume of distribution, 0.937 +/- 0.232 l/kg, elimination rate constant, 0.0249 +/- 0.0095/h, elimination half-life, 32.1 +/- 12.1 h, and total body clearance, 21.7 +/- 6.18 ml/h/kg, of theophylline in Korean premature neonates were comparable to the values of neonates in other countries. For caffeine, the mean (+/- S.E.M.) elimination half-life was 95.1 +/- 25.4 h and the elimination rate constant was 0.0079 +/- 0.0024/h. The mean (+/- S.E.M.) serum concentrations of theophylline and caffeine on the sixth day after aminophylline infusion were 10.4 +/- 2.28 microg/ml (range, 6.38-13.4 microg/ml) and 2.94 +/- 0.98 microg/ml (range, 1.80-4.44 microg/ml), respectively. The mean (+/- S.E.M.) caffeine to theophylline concentration ratio on the day after discontinuation of aminophylline infusion was 0.71 +/- 0.23 (range, 0.39-1.03). PMID- 10850375 TI - Systemic and coronary hemodynamic effects of JTV-506, a novel potassium channel opener, in conscious dogs: comparison with cromakalim and nicorandil. AB - We compared the coronary and systemic hemodynamic effects of JTV-506, a novel potassium channel opener, with those of cromakalim and nicorandil in chronically instrumented conscious dogs. Experiments were performed in 7 dogs which had undergone the implantation of flow probes on the ascending aorta and left circumflex coronary artery, and of catheters into the descending thoracic aorta and left ventricular cavity, respectively. On different days at least 10 days after the implantation, dogs were randomly assigned to the doses of JTV-506 (2, 5 or 10 microg/kg), cromakalim (10 microg/kg), nicorandil (0.2 mg/kg) or glibenclamide (5 mg/kg) plus JTV-506 (5 microg/kg). Each dose of the three drugs produced dose-related increases in heart rate, coronary blood flow, cardiac output, dP/dt(max) and %SS, and produced decreases in arterial blood pressure, left ventricular systolic pressure, and coronary and systemic vascular resistance. JTV-506 at doses of 2 and 5 microg/kg reduced arterial blood pressure only slightly as compared to its significant coronary vasodilating effect. An increase in myocardial oxygen consumption (as estimated by pressure-work index) was also observed in response to each dose of three drugs. At doses that reduced coronary vascular resistance by 75%, JTV-506 produced a greater increase in coronary blood flow, as compared with cromakalim and nicorandil. JTV-506 has the longest duration of action as compared with cromakalim and nicorandil. Glibenclamide pretreatment completely abolished the effects of JTV-506 on coronary and systemic hemodynamics. These results suggest that JTV-506 is relatively more potent in the coronary bed with minimal systemic influence, and exerts a longer time course of action. PMID- 10850376 TI - Dual effects of quercetin on contraction in cardiac and skeletal muscle preparations. AB - The effects of quercetin, a plant flavonoid, on contraction in cardiac and skeletal muscle preparations were examined. Higher concentrations of quercetin increased mean arterial blood pressure in anesthetized rats. Quercetin showed positive inotropic action on spontaneous contraction in a dose-dependent manner in isolated rat atria. Quercetin prevented the negative inotropism evoked by acetylcholine in isolated rat atria. On the other hand, quercetin inhibited the twitch contraction in isolated rat hemidiaphragm muscles. The addition of Ca2+ to extracellular medium recovered the inhibitory effects of twitch contraction in Ca2+-free medium but did not prevent the inhibitory effects of twitch contraction in Ca2+-free medium containing quercetin. The present results suggest that quercetin have a positive inotropic action in cardiac muscle but inhibitory effects on contraction in skeletal muscle. PMID- 10850377 TI - Hyperammonemia induced by administration of glucose and insulin after hepatopancreatectomy in rats. AB - Massive resection of both the liver and pancreas is performed as a radical procedure in some cases of advanced biliary cancer, but it has been reported that this disease is frequently complicated by hyperbilirubinemia or hepatic insufficiency postoperatively, which is a serious hindrance to performing such extended surgery (Nimura et al., 1991; Nakamura et al., 1992). To investigate the pathogenesis of hepatic dysfunction after hepatopancreatectomy, we performed 4 surgical procedures consisting of 68% hepatectomy, 90% pancreatectomy, 68% hepatectomy plus 90% pancreatectomy (hepatopancreatectomy) and sham-surgery in rats. Then, rats were continuously infused with 5% or 20% glucose solution at a constant speed (50 mL/day) for 24 hours in the fasting state, thus creating a total of 8 groups. During infusion of 20% glucose solution into rats with pancreatectomy or hepatopancreatectomy, insulin (1 U/5 g glucose) was added to the solution to adjust the blood glucose. In rats infused with 20% glucose solution with added insulin after hepatopancreatectomy, the blood glucose level did not differ, but adenosine 5'-triphosphate (ATP) and energy charge levels in the liver tissue were significantly lower, while the blood ammonia level was significantly higher than those in the other 7 groups. These results demonstrate that continuous infusion of high concentrations of glucose solution with added insulin after hepatopancreatectomy in rats reduces hepatic mitochondrial function, resulting in hyperammonemia due to reduced urea synthesis. PMID- 10850378 TI - Differential property of antigenic characterization between piroxicam and ampiroxicam in contact hypersensitivity. AB - Piroxicam (PXM; a non-steroidal anti-inflammatory drug) has been reported to induce photosensitivity. In our previous report, however, ultraviolet-A (UVA) irradiated or non-irradiated PXM did not induce any reactions in the in vivo model of contact hypersensitivity, while positive patch testing was shown by ampiroxicam (APX; a prodrug of PXM). The purpose of the present study was to clarify the influence of protein on the antigenicity of PXM using this model. Animals sensitized by UVA-irradiated 1% APX showed positive patch testing (open application) in UVA-irradiated 1% APX, while they were negative in challenge by UVA-irradiated PXM with or without 5% human serum albumin (HSA). Although animals sensitized by 1% thiosalicylate (TOS), which is thought to be an active hapten of PXM, were cross-reacted with UVA-irradiated 1% APX, they failed to react with UVA irradiated 1% PXM with or without HSA. On the other hand, intra-dermal testing (intra-dermal application) in UVA-irradiated 0.1% PXM with 5% HSA was positive in animals sensitized by UVA-irradiated 1% APX, while 5% HSA alone, 0.1% PXM with 5% HSA and UVA-irradiated 0.1% PXM did not induce any reactions under this condition. Furthermore, concentration of PXM in the presence of HSA was reduced by UVA-irradiation in a time dependent manner, while the degradation of PXM was not observed in the absence of HSA. Finally, PXM almost disappeared at 120 min after the initiation of UVA-irradiation. The degradation of PXM irradiated by UVA was dependent on the concentration of HSA at the range of 0 to 4%. Hence, these results suggest that the presence of protein is necessary for the induction of the antigenic activity of PXM and the antigenic characterization of PXM is different from that of APX in contact hypersensitivity. PMID- 10850379 TI - Evaluation of the effects of pregnyl on pituitary-ovarian hormones and biochemical markers of tissue injury in female Swiss albino mice. AB - Pregnyl (hCG), a preparation of human chorionic gonadotropin, was evaluated for its effects on the endocrinological, biochemical and genotoxic changes in female Swiss albino mice. hCG treatment at different doses (25, 50 and 100 I.U./Kg/day) for 5 days was found to significantly increase the plasma levels of hCG, estradiol and progesterone in a dose-dependent manner, while the concentrations of LH and FSH remained below the detection levels. The plasma levels of ALT, CK MB, creatinine and urea were significantly decreased, whereas the concentrations of AST were significantly increased. The treatment was found to significantly increase and decrease the hepatic concentrations of MDA and NP-SH respectively. The hepatic levels of proteins and DNA were not affected, but there was a significant increase in the concentrations of RNA. In addition, hCG treatment did not show any effect on the frequency of occurrence of micronuclei, whereas the ratio of PCE/NCE was found to be significantly increased. These results demonstrate that the hCG treatment in mice affected the pituitary-ovarian hormones in a similar pattern to that of humans. The treatment increased oxidative stress in hepatic cells without disturbing the functions of the liver as well as other organs. This finding may be of value concerning the safe use of hCG and may contribute to the overall antioxidant balance in the body. PMID- 10850380 TI - Effect of age on rat aortic responses to acetylcholine and nitric oxide donor (NOC-18). AB - The purpose of the present study was to examine the effects of age on vascular responses to acetylcholine (ACh) and NO-releasing compound (NOC-18). The studies were performed in young (4 mo old, n = 8) and old (22 mo old, n = 6) male rats. Responses to ACh and NOC-18 were examined in vitro by using isolated abdominal aortic rings. The maximum relaxation response to ACh, an endothelium-dependent vasodilation, was lower in aortas from old rats. Sensitivity (mean effective concentration; EC50) of ACh in old rats was significantly less than in young rats. There were no differences in maximum NOC-18-induced relaxation, an endothelium-independent vasodilation, in aortas from young and old rats. On the other hand, the concentration-response curve for NOC-18 was shifted to the right and the sensitivity (EC50 to NOC-18) was lower in old rats. These results indicated that both endothelium-dependent vasodilation induced by ACh and endothelium-independent vasodilation induced by NOC-18 are impaired in aorta from old rats. PMID- 10850381 TI - Reduction of oral mucositis by filgrastim (r-metHuG-CSF) in patients receiving chemotherapy. AB - Mucositis, the inflammation and necrosis of mucosal membranes, is a serious and debilitating consequence of many cancer therapies. We were interested in the potential role of filgrastim (recombinant methionyl human granulocyte colony stimulating factor, r-metHuG-CSF) in the reduction of mucositis. Patients with newly diagnosed small-cell lung cancer (SCLC) were treated with CAE chemotherapy (cyclophosphamide, doxorubicin, and etoposide) and placebo or filgrastim. If patients had an episode of febrile neutropenia, they received unblinded filgrastim in subsequent CAE cycles. Oral mucositis was considered to have occurred if a patient reported any clinical sign or symptom of oral mucositis with or without oral candidiasis. Oral mucositis was analyzed using the unadjusted chi-square test, and time to first episode of mucositis was analyzed using the stratified log-rank test as well as the Cox proportional hazards regression model. During cycle 1, placebo-treated patients had more episodes of mucositis (47%) compared with those patients randomized to filgrastim (28%). Across all cycles of treatment, 70% of placebo-treated patients experienced mucositis, compared with 53% of patients randomized to filgrastim. A significant reduction in the incidence of chemotherapy-related oral mucositis occurred across multiple cycles of treatment in patients treated with filgrastim. PMID- 10850382 TI - Differential in vitro maturation of hematopoietic stem cells from wild-type and immunoglobulin transgenic mice. AB - During B-cell lymphopoiesis, hematopoietic stem cells commit to the B-cell lineage as monitored by the expression of phenotypic cell surface antigens and the production of immunoglobulin chains. Two cytokines, interleukin-7 (IL-7) and Flt-3 ligand (FL), appear to act in conjunction to drive this development process. Using an in vitro, stroma-free culture system and these cytokines, the commitment of murine Sca+ Lin- bone marrow cells to the B-cell lineage was examined with stem cells from immunoglobulin (Ig) transgenic and wild-type mice. After 12 days of culture in IL-7 and FL, stem cells from wild-type animals had matured to express surface B220, CD19, CD43, BP-1 and heat-stable antigen (HSA). These cells lacked detectable intracellular mu chains while exhibiting partial D J rearrangement. In contrast, Sca+Lin- cells from Ig transgenic mice that were cultured similarly expressed B220, CD19, IgD, intracellular and surface mu, HSA but not CD43 or BP-1. These results suggest that expression of the Ig transgene during in vitro development overcame a block in B-cell lymphopoiesis and recapitulated in vivo events. Thus, IL-7 and FL treatment allowed uncommitted stem cells to progress to the early pre-B-cell stage while similarly treated Ig transgenic cells progressed completely to the mature B-cell stage. PMID- 10850383 TI - Prophylactic recombinant epoetin alfa markedly reduces the need for blood transfusion in patients with metastatic melanoma treated with biochemotherapy. AB - Treatment of metastatic melanoma with biochemotherapy results in the rapid onset of anemia, requiring blood transfusion in 9 of 13 (69%) patients. Prophylactic use of weekly subcutaneous recombinant epoetin alfa eliminated the need for transfusion in all but 1 of 21 (5%) patients. PMID- 10850384 TI - Utility of hepatitis C virus serotypes in predicting response to treatment of chronic hepatitis C. Consensus Interferon Study Group. AB - Hepatitis C virus (HCV) genotyping has been shown to predict response to interferon, but is expensive. HCV serotyping is less expensive and simpler, and may be similarly useful. Using data from a large, randomized trial comparing consensus interferon (CIFN) and interferon alfa-2b (IFN alfa-2b) in patients with chronic HCV, we evaluated response rates based on HCV serotypes versus genotypes. Patients included in this analysis received subcutaneous injection of 9 microg CIFN (n = 232) or 3 MU IFN alfa-2b (n = 240) three times weekly for 24 weeks followed by 24 weeks of observation. Serum HCV RNA concentrations were measured regularly during treatment and at the end of both the treatment and post treatment periods. Response to interferon was similar for HCV antibody types and their corresponding genotypes. The end-of-treatment HCV RNA rate of response (defined as undetectable serum on two consecutive assessments) was 29% for serotype 1 versus 24% for genotype 1 after CIFN; and 14% versus 15%, respectively, after IFN alfa-2b. Independently of treatment, patients infected with serotype or genotype 2 or 3 had a better therapeutic response than those infected with serotype or genotype 1. Similar results were obtained based on HCV antibody typing and genotyping, suggesting the potential of the former for predicting response to interferon. PMID- 10850385 TI - Genotype does not affect pattern of HCV RNA decrease among responders during interferon treatment of chronic hepatitis C. Consensus Interferon Study Group. AB - We assessed differences in the pattern of HCV RNA decrease for HCV genotypes 1, 2, and 3 during interferon treatment to determine if the lower response rates observed among genotype 1 patients were related to a slower decrease in HCV clearance. Serum HCV RNA values of 472 chronic hepatitis C patients treated with either consensus interferon (CIFN) or interferon alfa-2b (IFN alfa-2b) were evaluated. Neither virological sustained responders nor relapsers differed in the pattern of serum HCV RNA decrease based on genotype. Virological sustained responders infected with genotype 1 cleared HCV RNA as rapidly as sustained responders who were infected with genotype 2 or 3. Relapsers had a slower rate of serum HCV RNA decrease than did virological sustained responders. Nonresponders differed in the pattern of serum HCV RNA decrease based on genotype: HCV genotype 3 patients had the greatest decrease in serum HCV RNA; genotype 2 patients had an intermediate decrease; and genotype 1 patients had the least serum HCV RNA decrease. HCV genotype 1 patients treated with CIFN had a greater decrease in serum HCV RNA during therapy than did patients treated with IFN alfa-2b. However, there was no difference in the magnitude of serum HCV RNA decrease between the two interferon treatments for patients infected with genotype 2 or 3. In summary, both genotype and ultimate response to treatment are determinants of the pattern and rate of serum HCV RNA change during interferon therapy of chronic hepatitis C. PMID- 10850386 TI - Injection of DNA encoding granulocyte-macrophage colony-stimulating factor recruits dendritic cells for immune adjuvant effects. AB - An important issue for effective vaccines is the development of potent adjuvants that can facilitate induction or augmentation of immunity. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a growth factor for myeloid progenitors of monocytes and dendritic cells (DC), which upon maturation are antigen-presenting cells (APC). The adjuvant effects of inoculation of DNA encoding GM-CSF into skin were studied. Initial experiments examined whether the GM-CSF gene injected into the skin of mice could affect the density of epidermal DC (Langerhans cells). DNA encoding GM-CSF delivered by particle bombardment into skin resulted in a significant increase of epidermal DC at the inoculation site. Kinetic analysis of epidermal recruitment after GM-CSF inoculation showed an increase in DC that peaked at seven days. This increase was accompanied by recruitment of DC into draining lymph nodes. The adjuvant effects of DNA encoding GM-CSF inoculated into skin were measured by the ability to augment antibody and T-cell responses against poorly immunogenic tumor antigens. Peptide immunization at skin sites containing epidermal DC newly recruited by GM-CSF DNA elicited T-cell responses against mutant p53, whereas peptide immunization of control skin sites did not elicit any detectable T-cell responses. Likewise, generation of antibodies following immunization with DNA encoding human gp75TRP1, a tyrosinase family member expressed by melanomas, was accelerated and protection from tumor challenge augmented by GM-CSF DNA. PMID- 10850387 TI - Major salivary gland involvement in graft-versus-host disease: considerations related to pathogenesis, the role of cytokines and therapy. AB - Chronic graft-versus-host-disease (GVHD) is an autoimmune-like complication often occurring in patients who have been treated with bone marrow and peripheral blood stem cell transplantation. Various tissues and organs are damaged via the cytotoxicity rendered by the infiltrating donor graft T cells. The mucosal insult is enhanced by the reduced quantity and the altered quality of the saliva, since the salivary glands are a known major target of GVHD. The salivary changes are also expressed by a reduction in related functions, such as anti-infection activity, protection against mechanical and chemical epithelial injuries, assistance in controlling periodontal disease and caries, etc. The purpose of this review is to summarize the data that have been published recently concerning salivary involvement in GVHD and to suggest an underlying mechanism for the disease and its related 'state-of-the-art' therapeutic policy. PMID- 10850388 TI - Catalytic activity of three variants (Ile, Leu, and Thr) at amino acid residue 359 in human CYP2C9 gene and simultaneous detection using single-strand conformation polymorphism analysis. AB - This study evaluated the catalytic activity of three variants (Ile, Leu, and Thr) at codon 359 of CYP2C9 enzymes expressed in a yeast cDNA expression system, and then established single-strand conformation polymorphism (PCR-SSCP) analysis for simultaneous detection as a screening method. Diclofenac was used for the in vitro experiment, and its hydroxy metabolite (4'-hydroxydiclofenac) was measured by HPLC. To discuss the in vivo effect of the Thr359 variant on the pharmacokinetics of phenytoin, a case report is presented. The efficiency of the SSCP method was evaluated by analyzing DNA samples from a homozygote for Ile359 and a heterozygote for Leu359 or Thr359. To evaluate the interaction between the P450 level and reductase activity, two batches of the Thr359 variant with a different P450:reductase activity ratio (1:4.0 and 1:1.4) were used. The in vitro study revealed that recombinant Ile359, Leu359, and Thr359 (2 batches) possessed a mean Km of 2.0, 16.5 and (3.8 and 2.9) micromol and Vmax of 12.4, 17.9 and (4.4 and 5.1) nmol/min/nmol P450, respectively. Although the magnitude of the change in catalytic efficiency for the Thr359 variant was close to that of the Leu359 variant, the effect of the two variants on diclofenac 4'-hydroxylation appears to be different because Leu359 variant was associated with a high Km, and Thr359 with a low Vmax. No significant differences in the kinetic data were observed between the two Thr359 enzymes, suggesting that low reductase activity in the Thr359 enzyme was not a major determinant in the present in vitro experiment. Estimated pharmacokinetic parameters of phenytoin obtained by the Bayesian method in an epileptic patient who was a heterozygote carrier for Thr359 variant were: Km = 6.45 microg/mL, Vmax = 5.77 mg/kg/d, and Vmax/Km = 0.89 L/kg/day. The Vmax/Km value in this patient was similar to the population mean value (0.90 L/kg/day) in Japanese heterozygotes for the Leu359 variant. Results for PCR-SSCP were in complete agreement with those obtained using established methods. Thus, the PCR-SSCP approach is useful for identifying these three variants of the CYP2C9 gene. PMID- 10850389 TI - Effect of a genetic polymorphism of CYP1A2 inducibility on the steady state plasma concentrations of haloperidol and reduced haloperidol in Japanese patients with schizophrenia. AB - The effect of a genetic polymorphism of inducibility of cytochrome P450 (CYP) 1A2 on the steady state plasma concentrations (Css) of haloperidol and reduced haloperidol was studied to clarify if these Css are dependent on the CYP1A2 activity. The subjects were 101 Japanese schizophrenic inpatients receiving oral haloperidol 12 mg/d. The Css of haloperidol and reduced haloperidol were measured in duplicate by high performance liquid chromatographic method, and were corrected to the mean body weight. A point mutation from guanine (wild-type) to adenine (mutated-type) at position -2964 in the 5'-flanking region of CYP1A2 gene was identified by polymerase chain reaction (PCR)-fragment length polymorphism method. Based on the present results, i.e., significant effects of CYP2D6 genotypes on the Css of haloperidol and reduced haloperidol, analyses were separately performed in two groups, i.e., patients with 0 mutated allele of the CYP2D6 (41 cases) and those with 1 or 2 mutated alleles (60 cases). Subjects in each CYP2D6 genotype group consisted of 4 subgroups according to smoking habit and the presence of the mutated allele of the CYP1A2. Neither the Css of haloperidol nor that of reduced haloperidol significantly differed among the 4 subgroups in either CYP2D6 genotype group. The present study thus suggests that the CYP1A2 activity does not play an important role in controlling the Css of haloperidol or reduced haloperidol. PMID- 10850390 TI - Effects of hepatic function on vancomycin pharmacokinetics in patients with cancer. AB - Vancomycin is widely used in the prophylaxis and treatment of infections in neutropenic patients with cancer. The objective of this study was to analyze liver damage effects on vancomycin pharmacokinetics and determine the necessity for liver function evaluation when selecting vancomycin dosing schedules in these patients. A population pharmacokinetic analysis was performed using the global two-stage method. To this purpose serum vancomycin concentrations from 154 cancer patients were measured and individual vancomycin pharmacokinetic parameters were estimated by the Sawchuk and Zaske method. Mean and standard deviation of the vancomycin pharmacokinetic parameters were estimated for various subgroups of patients classified according to the degree of liver damage. Then a multiple linear regression analysis was performed to select the best predictive models for vancomycin clearance (Clvan) and steady state distribution volume (V). Results revealed that Clvan is not influenced by liver failure. Differences in V between patients with and without hepatic failure were initially observed, but these disappeared when patients with ascites were excluded. In conclusion, vancomycin dosing schedule does not need to be modified for patients with liver failure, with the exception of patients with ascites. PMID- 10850391 TI - On the assessment of drug metabolism by assays of codeine and its main metabolites. AB - Codeine and its main metabolites appear to have advantages for assessing drug metabolic phenotypes. The authors have further developed a high-performance liquid chromatography (HPLC) method for the quantification of codeine and six of its metabolites in urine. Quantification was performed by electrochemical detection for morphine, normorphine, morphine-6-glucuronide, and the internal standard 4-O-methyldopamine; and by ultraviolet detection for codeine, norcodeine, and morphine-3-glucuronide. The method had a detection limit of 2 nmol/L(-1) for morphine and normorphine, 4 nmol/L(-1) for morphine-6-glucuronide, 3 nmol/L for the internal standard, 20 nmol/L(-1) for morphine-3-glucuronide, and 60 nmol/L(-1) for codeine and norcodeine. The coefficients of variations were <9% for intraday and <10% for interday analyses. The recovery of codeine and its metabolites ranged from 55% (for morphine-3-glucuronide) to 90% (for codeine, norcodeine, morphine, and morphine-6-glucuronide). Eleven healthy volunteers were phenotyped for CYP2D6 using codeine as well as debrisoquine and dextromethorphan. Ten subjects were extensive metabolizers (EM) and one a poor metabolizer (PM) of codeine, debrisoquine, and dextromethorphan. Significant correlations between the metabolic ratios (MRs) of the different probe drugs were obtained (r2 > 0.95, p < 0.001). This HPLC method is simple, sensitive, accurate, and reproducible for assessing the CYP2D6 phenotype. PMID- 10850392 TI - Mechanisms of cocaine hydrolysis and metabolism in vitro and in vivo: a clarification. AB - There is confusion in the literature concerning the mechanisms by which the cocaine hydrolysis product, benzoylecgonine (BE), is formed in vitro and in vivo. Some authors assume that all BE is formed nonenzymatically. This review summarizes evidence that both enzymatic and nonenzymatic mechanisms exist. In vitro BE is produced exclusively by hydrolysis at alkaline pH, as esterases present in blood or serum do not catalyze formation of BE. In vivo BE is formed both nonenzymatically as well as through the action of esterases found in a number of tissues including hepatocytes. The enzymatic mechanism is the predominant one operating in vivo. PMID- 10850393 TI - Simultaneous determination of human plasma levels of four selective serotonin reuptake inhibitors by high-performance liquid chromatography. AB - A reversed-phase high-performance liquid chromatography (HPLC) method with fluorimetric detection, which allows the simultaneous determination of plasma concentrations of four selective serotonin reuptake inhibitors (SSRIs) is presented. Fluvoxamine, paroxetine, sertraline, and fluoxetine were extracted from plasma with ethyl acetate and then derivatized with dansyl chloride. The analytes were separated using Hypersyl ODS C18 (5 microm) 250 x 4.6 mm column (ThermoQuest, Runcorn, UK). For continuous gradient separation, the mobile phase consists of two eluents, acetonitrile and potassium phosphate buffer (10 mmol/L, pH 7.2) at total flow rate of 1.5 mL/min. Detection was carried out at lambda exc = 366 nm and lambda em = 490 nm. The authors found recoveries of 90% to 95% for fluvoxamine, 94% to 100% for paroxetine, 88% to 95% for sertraline, 93% to 100% for fluoxetine, and 97% to 100% for internal standard (nortriptyline). Imprecision of the method ranged from 2.5% to 8.9%. The assay was linear from 10 to 1500 ng/mL for sertraline, and from 5 to 1500 ng/mL for the other drugs. The authors conclude that this method is suitable for monitoring antidepressant therapy. In addition, the authors report the effects of adding paroxetine to fluvoxamine on plasma levels in a group of patients in combined drug therapy. PMID- 10850394 TI - Distinction of inhaled and oral salbutamol by urine analysis using conventional screening procedures for doping control. AB - Salbutamol administration in athletes is permitted only by inhalation, for the management of asthma. The authors discuss different criteria for suspecting oral use of salbutamol, taking into account the data obtained by application of two conventional screening procedures for doping control: gas chromatography/mass spectrometry (GC/MS) and enzyme-linked immunosorbent assay (ELISA). Urine samples obtained after administration of oral and inhaled salbutamol to asthmatic and nonasthmatic swimmers were analyzed using both analytical approaches. As expected, concentrations obtained by the ELISA test (detection of total salbutamol) were higher than those obtained using the GC/MS procedure (detection of nonsulfated salbutamol). After oral administration, the ELISA test detected significantly higher salbutamol concentrations than those detected after inhalation, reflecting the greater doses administered orally. Urine samples with total salbutamol greater than 1400 ng/mL were obtained after oral doses, but no sample reached this value after inhaled doses. Higher concentrations of nonsulfated salbutamol have also been detected after oral intake, although there is an overlap between the distributions of concentrations after oral and inhaled doses. A cut-off concentration of 500 ng/mL can be used for nonsulfated salbutamol to select suspicious samples, giving 11.8% false negative results and 4.3% false positive results. An additional criterion evaluated was the androsterone-salbutamol peak height ratio, which was lower after oral doses because of the higher concentrations of salbutamol in urine. This ratio was lower than 2 for all the samples collected after oral administration, although 6.8% false positive samples resulted because of low concentrations of androsterone in female urine. Several possibilities for detecting suspicious samples from athletes who have taken prohibited oral salbutamol are available with conventional screening procedures in doping control. PMID- 10850395 TI - Development and application of a high-performance liquid chromatography-based assay for determination of the activity of inosine 5'-monophosphate dehydrogenase in whole blood and isolated mononuclear cells. AB - With the objective of pharmacodynamic monitoring of the immunosuppressive efficacy of mycophenolate mofetil (MMF) (CellCept, Hoffman-LaRoche, Grenzach Wyhlen, Germany), a method for determination of the inosine monophosphate dehydrogenase (IMPDH) activity in whole blood cell (WBC) lysates and mononuclear cells (MNCs) was developed. The assay is based on the incubation of WBC lysates or lysed MNCs in the presence of supplemented inosine 5'-monophosphate (IMP) and nicotimamide adenine dinucleotide (NAD). The formation of xanthosine 5' monophosphate (XMP) was determined by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection. The analytical method was validated, and the obtained data demonstrated that the amount of XMP in WBC and MNC lysates can be reliably determined by this method. Under assay conditions the rate of XMP formation remained constant within the incubation period of 60 minutes and a quantification of product formation at 30 and 60 minutes proved to be sufficient to reliably characterize the IMPDH activity. Applications of this assay with whole blood indicated extremely high IMPDH-activities in samples from patients with renal transplant receiving MMF. IMPDH monitoring within 10 hours after administration of the morning dose demonstrated a marked enzyme inhibition between 2 hours and 3 hours postdosing, but the activities returned to predose levels within one dose interval. The analysis of isolated cell fractions indicated that the IMPDH-activity is predominantly located in erythrocytes. The contribution of MNCs to the whole blood activity remained below 10%. In order to simulate the in vivo exposure of MNCs to mycophenolic acid, an "erythrocyte- and platelet-free" whole blood was reconstituted by resuspension of isolated MNCs with plasma. This strategy allowed for the reliable measurement of IMPDH activity in the target cells of immunosuppression. PMID- 10850396 TI - Rituximab (IDEC-C2B8): validation of a sensitive enzyme-linked immunoassay applied to a clinical pharmacokinetic study. AB - Rituximab is a chimeric monoclonal antibody (MAb) directed against the B-cell CD20 antigen that has been approved for therapy of relapsed and resistant follicular non-Hodgkin's lymphoma (NHL). This study describes the development and validation of a highly sensitive, rapid, accurate, precise enzyme-linked immunosorbent assay (ELISA) to measure Rituximab serum concentrations. This study also describes the application of the ELISA method to a pharmacokinetic study in a homogeneous group of patients with follicular lymphoma who received 4 weekly doses of MAb at the standard dose of 375 mg/m2 as consolidation of chemotherapy. In the patients in this study, the median Rituximab serum concentrations increased during therapy, and showed a slow decline during the posttreatment period. The Rituximab elimination half-life of approximately 20 days accounts for the demonstrated accumulation of MAb in serum samples. Because previous pharmacokinetic studies showed a correlation between Rituximab serum levels and tumor response, the ELISA method used in this study, which allows a precise control of serum concentrations, could be useful for predicting the final response to the MAb and for selecting patients able to benefit from higher dosage or repeated drug administration. PMID- 10850397 TI - Clinical and analytical aspects of pyrrolizidine poisoning caused by South African traditional medicines. AB - In a study carried out in two hospitals in South Africa the authors identified 20 children suffering from hepatic veno-occlusive disease thought to be caused by the administration of traditional remedies. The predominant clinical presentation was ascites of various degrees and hepatomegaly. There was a high morbidity and mortality in the young infants, and in those cases who survived and were followed up the clinical pattern was one of progression to cirrhosis and portal hypertension. Pyrrolizidine alkaloid poisoning is one of the causes of the veno occlusive disease. Therefore there is a need for objective confirmation of this. In four of our cases an on-admission urine specimen was available and in all of these a simple colorimetric screening test confirmed the presence of pyrrolizidine alkaloids. The other cases were admitted from peripheral hospitals and clinics and urine was not obtained until after 72 h, a time at which the levels of pyrrolizidines in urine were below the limit of sensitivity of the screening test. The screening method is helpful for the detection of acute ingestion of pyrrolizidines in large amounts, but is not sufficiently sensitive for the detection of chronic ingestion of smaller amounts. Nevertheless, in those patients who have hepatomegaly and ascites a positive finding of pyrrolizidines is important and may remove the necessity for expensive and invasive investigative measures. PMID- 10850398 TI - Diurnal changes in the pharmacokinetic behavior of amikacin. AB - This retrospective study evaluated possible differences in the pharmacokinetic behavior of amikacin between the morning (AM) and evening (PM). Of 634 patients receiving amikacin therapy, 17 received a dose every 12 hours (an i.v. infusion at 8:00 AM and 8:00 PM) with amikacin serum levels obtained after both the AM and PM infusions. Pharmacokinetic parameter values were estimated by the nonparametric EM algorithm (USC*PACK clinical software) for a one-compartment model. All patient data were analyzed in three ways. The parameter values were estimated by fitting the model first only to the serum levels drawn following the AM dose; second, only to the data following the PM dose; and third, to all serum levels (AM + PM). Parameter values found were (mean, median, SD respectively): AM: Kel = 0.181114 h(-1), 0.224460 h(-1), 0.058820 h(-1); Vol = 23.657507 L; 23.376231 L; 1.353253 L; Cl = 4.326720 L x h(-1), 5.303726 L x h(-1), 1.447731 L x h(-1); PM: Kel = 0.110151 h(-1); 0.121295 h(-1); 0.016860 h(-1); Vol = 28.948043 L; 24.091703 L; 9.266628 L; Cl = 3.081761 L x h(-1), 2.810615 L x h( 1); 0.705874 L x h(-1); AM + PM: Kel = 0.165321 h(-1); 0.131796 h(-1); 0.075425 h(-1); Vol = 25.479043 L; 26.187970 L; 5.367054 L. These findings are in agreement with the known diurnal rhythm of glomerular filtration rate. Because pharmacokinetic parameter values are most often estimated using AM data, this may lead to an overevaluation of these values compared with PM or to values for the entire day. The resulting drug regimens may therefore be overestimated regarding the elimination rate constant and underestimated regarding the volume of distribution. PMID- 10850399 TI - Validation of population pharmacokinetic parameters of phenytoin using the parallel Michaelis-Menten and first-order elimination model. AB - This study was conducted to assess whether the parallel Michaelis-Menten and first-order elimination (MM+FO) model fitted the data better than the Michaelis Menten (MM) model, and to validate the MM+FO model and its parameter estimates. The models were fitted to 853 steady state dose: serum concentration pairs obtained in 332 adults with epilepsy using nonlinear mixed-effects modeling (NONMEM). The MM+FO model fitted the data better than the MM model. The validity of the pharmacokinetic models and the estimated population parameter values was tested using the naive prediction method. The estimation and validation of the pharmacokinetic parameters were undertaken in two separate patient groups (cross validation) obtained by splitting the data set. Patients were randomly allocated to two equally matched groups (groups 1 and 2). The predictive performance was assessed using 770 paired predicted versus actual dose or measured serum concentrations. The population pharmacokinetic parameters estimated by NONMEM in group 1 were validated in group 2 and vice versa. When predicting steady state serum concentration, the MM+FO model was clearly superior to the MM model (mean bias of 0.91 and 8.13 mg/L, respectively). PMID- 10850400 TI - Lidocaine and seizures. AB - Lidocaine has a concentration-dependent effect on seizures. At lower concentrations it has anticonvulsant properties, whereas concentrations above 15 microg/mL frequently result in seizures in laboratory animals and man. Seizures induced by lidocaine in experimental conditions invariably start in the amygdala. Despite the clear focal onset in these experimental models, the seizures emerging in patients given intravenous (i.v.) lidocaine are almost invariably generalized and without any clear signs of focality. Given the prevalence of partial seizures and the frequent use of lidocaine, a higher incidence of partial seizures would be expected with its use. Yet this is clearly not the case. These facts suggest that a history of partial seizures is not a major risk factor for the precipitation of partial seizures in patients treated with intravenous lidocaine. PMID- 10850401 TI - Therapeutic drug monitoring of clozapine: an unexpected outcome. AB - A patient is described who died with the diagnosis of septicemia. After her death the delayed results of a clozapine determination for TDM were sent to the clinician. The clozapine serum level was 4034 microg/L, which was considered to be the primary cause of death. However, a forensic autopsy revealed unexpected metastases of unknown origin with gross liver involvement. Thus the high clozapine levels were judged to be secondary to liver failure. This case is an example of an unexpected outcome of TDM. PMID- 10850402 TI - Goal-oriented, model-based drug regimens: setting individualized goals for each patient. AB - Serum drug concentrations have commonly been described in terms of therapeutic ranges within which most patients have a therapeutic effect and a low incidence of toxicity. However, truly individualized drug dosage regimens cannot be developed without first setting a specific individualized target goal, such as a target serum drug concentration, at a desired target time after the dose (usually at a peak or trough), for each patient. For example, it is well known that the dosage of digoxin, or of any drug with a narrow therapeutic range, should somehow be individualized. One can begin this process by considering each patient as an individual, with his/her own individual need for the drug. If the need is small, so is the upper acceptable risk of toxicity. This would lead to a gently regimen, adjusted to the patient's body weight and renal function, to best achieve that specific target goal. Alternatively, if previous therapy has not sufficed and a significant or urgent need exists, then a higher goal may justifiably be selected, a greater risk of toxicity accepted, and a dosage regimen developed to meet that greater need. After such an individualized target goal is chosen, it should be achieved as precisely as possible. After the regimen is given, serum levels need to be measured and an individualized, patient-specific pharmacokinetic model should be made. Without the model, with only the raw serum level data, one cannot perceive the important exchanges that occur between serum and nonserum compartments of the drug, and we lack the precision given by the combination of the assay and the model to evaluate properly, optimally, the patient's clinical sensitivity to the drug. These concepts have been discussed here for digoxin, but they are general and apply to all drugs. This approach has also been applied to therapy with aminoglycoside antibiotics, vancomycin, lidocaine, theophylline, antiviral agents, a variety of anesthetic agents, psychiatric drugs, and anticancer agents. PMID- 10850403 TI - Recommendations for bioequivalence testing of cyclosporine generics revisited. AB - The immunosuppressant cyclosporine is generally considered a critical-dose drug. The validity of standard criteria to establish bioequivalence between cyclosporine formulations has recently been challenged. Recommendations included establishment of individual bioequivalence rather than average bioequivalence, establishment of bioequivalence in transplant patients and in subgroups known to be poor absorbers, as well as long-term efficacy and safety studies in transplant patients. However, at the moment individual bioequivalence is a theoretical concept, the practical benefits of which have not statistically been proven. The proposed patient pharmacodynamic studies can be expected to require an unrealistically high number of subjects to achieve sufficient statistical power. It is well established that the common practice of blood-concentration-guided dosing of cyclosporine efficiently compensates for interindividual and intraindividual variability and allows for safely switching cyclosporine formulations as bioinequivalent as Sandimmune and Neoral. Recent studies comparing the generic cyclosporine formulation SangCya with Neoral, including individual bioequivalence, bioequivalence in transplant patients, and long-term safety after switching from Sandimmune to SangCya, confirmed that it was valid to conclude bioequivalence of both cyclosporine formulations based on standard average bioequivalence criteria. Present FDA guidelines for approving bioequivalence can be considered adequate and sufficient for generic cyclosporine formulations. PMID- 10850404 TI - Achieving target goals most precisely using nonparametric compartmental models and "multiple model" design of dosage regimens. AB - Multiple model (MM) design and stochastic control of dosage regimens permit essentially full use of all the information contained in either a Bayesian prior nonparametric EM (NPEM) population pharmacokinetic model or in an MM Bayesian posterior updated parameter set, to achieve and maintain selected therapeutic goals with optimal precision (least predicted weighted squared error). The regimens are visibly more precise in the achievement of desired target goals than are current methods using mean or median population parameter values. Bayesian feedback has now also been incorporated into the MM software. An evaluation of MM dosage design using an NPEM population model versus dosage design based on conventional mean population parameter values is presented, using a population model of vancomycin. Further feedback control was also evaluated, incorporating realistic simulated uncertainties in the clinical environment such as those in the preparation and administration of doses. PMID- 10850405 TI - Population pharmacokinetics/pharmacodynamics modeling: parametric and nonparametric methods. AB - As clinicians acquire experience with the clinical and pharmacokinetic behavior of a drug, it is usually optimal to record this experience in the form of a population pharmacokinetic model, and then to relate the behavior of the model to the clinical effects of the drug or to a linked pharmacodynamic model. The role of population modeling is thus to describe and record clinical experience with the behavior of a drug in a certain group or population of patients or subjects. PMID- 10850406 TI - Disposition of propafenone in a poor metabolizer of CYP2D6 with Gilbert's syndrome. AB - Gilbert's syndrome, a genetic deficiency in bilirubin UDP-glucuronosyltransferase (UGT1A1), may dispose to increased toxicity of propafenone in poor metabolizers (PMs) of cytochrome P4502D6 because glucuronidation of propafenone is the major metabolic pathway for drug elimination in PMs. A patient with Gilbert's syndrome who is also PM participated in an interaction study with propafenone and rifampicin along with five otherwise healthy PMs. Using stable isotope techniques, the pharmacokinetics of single doses of 140 mg propafenone i.v. (unlabelled) and 300 mg propafenone p.o. (labelled) were compared between the index patient and the five healthy controls. Propafenone did not accumulate in the plasma of the index patient either before or during induction: AUC(0 infinity) of propafenone in the index patient was within the 95% confidence interval of controls; AUC(0-infinity) of propafenone glucuronide and amount of urinary excretion of propafenone glucuronide in the patient were within or even greater than the 95% confidence intervals of controls. Therefore, individuals with Gilbert's syndrome who also have a PM phenotype appear to be at no higher risk for toxicity of propafenone than otherwise healthy PMs. An indirect conclusion from these in vivo data might be that propafenone is not a substrate of the UGT1A1 isoform. PMID- 10850407 TI - p53 polymorphism in human papillomavirus-associated esophageal cancer. AB - Human papillomavirus type 16/18 (HPV-16/18) is implicated in the pathogenesis of squamous cell carcinoma (SCC) of the cervix and esophagus. The arginine allele at codon 72 of p53 was found to be more susceptible to degradation by HPV E6 protein than is the proline allele in vivo, thus resulting in a high frequency of cervical SCC in individuals homozygous for arginine at the codon. There are controversial results from several clinical studies of cervical SCC. In the present study, encoding regions of p53 codon 72 and HPV-16/18 E6 were directly sequenced, using pairs of primary esophageal SCC tissue and corresponding normal mucosa, which were from 75 patients (Japanese, n = 38; Chinese, n = 37). The arginine allele alone was detected in 70.6% (12 of 17) of HPV-positive cases but only in 43.1% (25 of 58) of HPV-negative cases (P < 0.05). In contrast, such a significant correlation between p53 polymorphism and HPV infection was not evident in corresponding normal mucosae. Because our findings between tumor specimens and the normal mucosae differed, we suggest that the frequent loss of proline allele in HPV-associated carcinogenesis of the esophagus major plays some role. The particular type of p53 polymorphism may indicate a potential candidate for HPV-associated SCC. PMID- 10850408 TI - Amplification in human breast cancer of a gene encoding a c-myc mRNA-binding protein. AB - The coding region determinant-binding protein (CRD-BP) binds in vitro to c-myc mRNA and is thought to stabilize the mRNA and increase c-Myc protein abundance. The CRD-BP gene has 15 exons and 14 introns, is single-copy, and is located on chromosome 11 in mice and 17 in humans, close to HER-2/neu. The CRD-BP gene is moderately amplified in 12 of 40 human breast cancers; it is highly amplified in 2 others (14.4 and 20 copies). Despite their proximity, CRD-BP and HER-2/neu genes can be amplified independently. Amplification of a gene that might up regulate c-Myc abundance could accelerate breast cancer. PMID- 10850409 TI - Detection of exon deletions and duplications of the mismatch repair genes in hereditary nonpolyposis colorectal cancer families using multiplex polymerase chain reaction of short fluorescent fragments. AB - Large genomic deletions within the mismatch repair MLH1 and MSH2 genes have been identified in families with the hereditary nonpolyposis colorectal cancer (HNPCC) syndrome, and their detection represents a technical problem. To facilitate their detection, we developed a simple semiquantitative procedure based on the multiplex PCR of short fluorescent fragments. This method allowed us to confirm in HNPCC families three known deletions of MLH1 or MSH2 and to detect in 19 HNPCC families, in which analysis of mismatch repair genes using classical methods had revealed no alteration, a deletion of exon 5 and a duplication of MSH2 involving exons 9 and 10. The presence of such duplications, the frequency of which is probably underestimated, must be considered in HNPCC families in which conventional screening methods have failed to detect mutations. PMID- 10850410 TI - Functional evidence for a novel human breast carcinoma metastasis suppressor, BRMS1, encoded at chromosome 11q13. AB - We previously showed that introduction of a normal, neomycin-tagged human chromosome 11 reduces the metastatic capacity of MDA-MB-435 (435) human breast carcinoma cells by 70-90% without affecting tumorigenicity, suggesting the presence of one or more metastasis suppressor genes encoded on human chromosome 11. To identify the gene(s) responsible, differential display comparing chromosome 11-containing (neo11/ 435) and parental, metastatic cells was done. We describe the isolation and functional characterization of a full-length cDNA for one of the novel genes, designated breast-cancer metastasis suppressor 1 (BRMS1), which maps to human chromosome 11q13.1-q13.2. Stably transfected MDA-MB-435 and MDA-MB-231 breast carcinoma cells still form progressively growing, locally invasive tumors when injected into mammary fat pads but are significantly less metastatic to lungs and regional lymph nodes. These data provide compelling functional evidence that breast-cancer metastasis suppressor 1 is a novel mediator of metastasis suppression in human breast carcinoma. PMID- 10850411 TI - Telomere erosion varies during in vitro aging of normal human fibroblasts from young and adult donors. AB - The life span of normal fibroblasts in vitro (Hayflick limit) depends on donor age, and telomere shortening has been proposed as a potential mechanism. By quantitative fluorescence in situ hybridization and Southern blot analysis, we show progressive telomere loss to about 5 kb mean telomere restriction fragment length in fibroblasts from two adult donors within 40 population doublings, whereas in fibroblasts from two infant donors, telomere erosion is reduced, leaving a mean telomere restriction fragment length of approximately 7 kb at senescence (after approximately 60 population doublings). Aging of fibroblasts from both infant and adult donors was not accompanied by chromosomal abnormalities but was correlated with increased telomere repeat-binding factor 2 expression at both the protein and transcriptional level. PMID- 10850412 TI - Deletion of 6q16-q21 in human lymphoid malignancies: a mapping and deletion analysis. AB - Two distinct regions of minimal deletion (RMD) have been identified at 6q25-q27 in non-Hodgkin's lymphoma (RMD-1), and at 6q21-q23 in acute lymphoblastic leukemia (ALL; RMD-2) by loss of heterozygosity and fluorescence in situ hybridization studies. In this study, 30 overlapping yeast artificial chromosomes (YACs), 1 expressed sequence tag, and 11 novel YAC ends were identified using bidirectional YAC walks between markers D6S447 (proximal) and D6S246 (distal) in RMD-2. The genes AF6q21, human homologue of the Drosophila tailless (HTLX), CD24 antigen, the Kruppel-like zinc finger BLIMP1, and cyclin C (CCNC), previously mapped to 6q21, were accurately positioned in a telomere-to-centromere orientation. Approximately 3.5 Mb were found to separate the BLIMP1 (adjacent to D6S447) and AF6q21 genes (telomeric to D6S246). Deletions of 6q were investigated in 21 cases of ALL using the newly characterized YAC clones in dual-color fluorescence in situ hybridization studies. A region centromeric to D6S447 (containing marker D6S283) and a region telomeric to marker CHLC.GGAT16CO2 (and containing marker D6S268) were identified as distinct and nonoverlapping regions of deletion in ALL. PMID- 10850413 TI - Differential susceptibility of renal carcinoma cell lines to tumor suppression by exogenous Fhit expression. AB - Hemizygous deletions of the fragile histidine triad (FHIT) gene at human chromosome band 3p14.2 and down-regulation of its gene product are found in the majority of renal cell carcinomas (RCCs). Functional tumor suppressive activity of Fhit in renal cancer cells previously was observed in RCC cell line RC48, which lacks endogenous Fhit expression. To further investigate the potential role of FHIT as a tumor suppressor gene in RCC, we transfected FHIT cDNA expression constructs into RCC cell lines RCC-1 and SN12C, which show low-level expression of endogenous Fhit and reveal an intact von Hippel-Lindau (VHL) gene. Stable transfectants of both cell lines showed no alterations of cell morphology, proliferation kinetics, or cell cycle parameters in vitro. The FHIT gene transfer rate, however, was significantly lower in RCC-1 cells compared with SN12C cells, suggesting a selection against exogenous Fhit expression. In addition, in nude mouse assays, a significant delay of tumor formation was observed for FHIT transfected RCC-1 cell lines, with outgrowing tumors demonstrating loss of Fhit expression in the majority of cells. In contrast, tumorigenicity of FHIT transfected SN12C cell clones was not suppressed, despite stable transgene expression. In conclusion, our results demonstrate a selective tumor suppressive activity of Fhit in RCC cells in vivo and suggest that the susceptibility to suppression is not restricted to cancer cells with complete loss of Fhit expression. PMID- 10850414 TI - Translocation t(10;14)(q11.2:q22.1) fusing the kinetin to the RET gene creates a novel rearranged form (PTC8) of the RET proto-oncogene in radiation-induced childhood papillary thyroid carcinoma. AB - Evaluation of 20 cases of radiation-induced childhood papillary thyroid carcinoma using fluorescence in situ hybridization demonstrated the presence of clonal translocations affecting the RET locus. Semiquantitative reverse transcription PCR indicated overexpression of the RET tyrosine kinase (TK) domain in four cases. In two cases, the RET rearrangements PTC6 and PTC7 were identified and assigned to balanced translocations t(7;10)(q32;q11.2) and t(1;10)(p13;q11.2), respectively. In one case with a balanced translocation t(10;14)(q11.2;q22.1), 5' rapid amplification of cDNA ends revealed a novel type of RET oncogenic activation (PTC8), arising from a fusion of the 5' part of the kinectin (KTN1) gene to the TK domain of the RET gene. The presence of coiled-coil domains in the resulting ktn1/ret fusion protein suggests ligand-independent dimerization and thus constitutive activation of the ret TK domain. PMID- 10850415 TI - Oncolysis of diffuse hepatocellular carcinoma by intravascular administration of a replication-competent, genetically engineered herpesvirus. AB - Herpes simplex virus type 1 (HSV-1) replication within tumors can mediate tumor regression (oncolysis). The genetically engineered, HSV-1 mutant rRp450 does not express viral ribonucleotide reductase and is therefore replication conditional. During the course of infection, rRp450 expresses the cytochrome P450 transgene and HSV-1 thymidine kinase gene, thereby enabling it to bioactivate the prodrugs cyclophosphamide and ganciclovir, respectively. rRp450 replication in hepatocellular carcinoma (HCC) cells is cytotoxic and liberates progeny virion that infect adjacent tumor cells. rRp450-mediated oncolysis is enhanced in the presence of cyclophosphamide, whereas it is inhibited in the presence of ganciclovir. As a consequence of defective viral ribonucleotide reductase expression, the yield of rRp450 progeny virions from infection of HCC cells is 3 to 4 log orders greater than that from infection of normal hepatocytes. This is associated with dramatic tumor reduction of diffuse HCC after a single intravascular administration of rRp450. rRp450 holds the promise of the dual therapeutic benefit of selective oncolysis and P450 transgene delivery. PMID- 10850416 TI - Expression of a repressor of estrogen receptor activity in human breast tumors: relationship to some known prognostic markers. AB - The expression of a specific repressor of estrogen receptor activity (REA) was investigated by a semiquantitative reverse transcription-PCR assay in 40 human breast tumor biopsy samples with respect to steroid hormone receptor status and other known prognostic variables. The data showed that REA expression was positively correlated with estrogen receptor (ER) levels as defined by ligand binding assays (Spearman r = 03231; P = 0.042) and that the median level of REA mRNA was significantly (Mann-Whitney two-tailed test, P = 0.0424) higher in ER+ tumors (median = 94.5; n = 30) compared with ER- tumors (median = 645; n = 10), with no significant differences (P = 0.4988) associated with progesterone receptor status alone. In addition, REA expression was inversely correlated with tumor grade (Spearman r = -0.4375; P = 0.0054). When the tumors were divided into two groups based on grade, REA expression was significantly (Mann-Whitney two tailed test, P = 0.0024) higher in low-grade (median = 97; n = 16) compared with high-grade (median = 76; n = 23) tumors. These results provide preliminary data suggesting that the expression of REA varies among breast tumors and is correlated with known treatment response markers and inversely correlated with a marker of breast cancer progression. REA together with ER status may be an improved marker of endocrine therapy responsiveness in human breast cancer. PMID- 10850417 TI - Aloe-emodin is a new type of anticancer agent with selective activity against neuroectodermal tumors. AB - Here we report that aloe-emodin (AE), a hydroxyanthraquinone present in Aloe vera leaves, has a specific in vitro and in vivo antineuroectodermal tumor activity. The growth of human neuroectodermal tumors is inhibited in mice with severe combined immunodeficiency without any appreciable toxic effects on the animals. The compound does not inhibit the proliferation of normal fibroblasts nor that of hemopoietic progenitor cells. The cytotoxicity mechanism consists of the induction of apoptosis, whereas the selectivity against neuroectodermal tumor cells is founded on a specific energy-dependent pathway of drug incorporation. Taking into account its unique cytotoxicity profile and mode of action, AE might represent a conceptually new lead antitumor drug. PMID- 10850418 TI - A novel antisense oligonucleotide targeting survivin expression induces apoptosis and sensitizes lung cancer cells to chemotherapy. AB - Survivin, an inhibitor of apoptosis protein, deserves attention as a selective target for cancer therapy because it lacks expression in differentiated adult tissues but is expressed in a variety of human tumors. We designed 20-mer phosphorothioate antisense oligonucleotides targeting different regions of survivin mRNA and investigated their ability to down-regulate survivin mRNA and induce apoptosis in the lung adenocarcinoma cell line A549. Oligonucleotide 4003, which targets nucleotides 23-251 of survivin mRNA, was identified as the most potent compound. As measured by real-time PCR, 4003 down-regulated survivin mRNA in a dose-dependent manner with an IC50 of 200 nM. Its maximum effect was achieved at a concentration of 400 nM, at which mRNA was down-regulated by 70%. As revealed by increased caspase-3-like protease activity, nuclear condensation and fragmentation, and trypan blue uptake, treatment with 4003 induced apoptosis and sensitized tumor cells to the chemotherapeutic agent etoposide. Oligonucleotide 4003 did not reduce the viability of normal blood leukocytes with marginal levels of survivin mRNA. PMID- 10850419 TI - Peripheral tolerance to human papillomavirus E7 oncoprotein occurs by cross tolerization, is largely Th-2-independent, and is broken by dendritic cell immunization. AB - The E7 oncoprotein of human papillomavirus 16 functions as a tumor-specific antigen in transformed epithelial cells of the uterine cervix to which immunotherapeutic strategies aimed at CTL induction may be directed. We previously have shown in mice transgenic for the E7 gene driven off an epithelial specific (keratin-14) promoter, that expression of E7 protein in peripheral epithelium is sufficient to tolerize E7-directed CTL precursors (pCTL; Doan et al, J. Virol., 73: 6166-1670, 1999). Here we show that E7 is presented to T cells for tolerization by cells of bone marrow origin ("cross-tolerization"). We demonstrate that tolerization of E7-directed pCTLs occurs within 2 weeks of exposure to E7 in epithelium. It is maintained in the near absence of CD4+ cells and in the absence of the thymus, and is independent of a coexisting E7-directed Th2-type antibody response. Tolerance was broken by immunization with E7 CTL epitope-pulsed dendritic cells. These findings have implications for immunotherapy of patients with human papillomavirus 16-associated cervical carcinoma, whose immune systems may have experienced long-term exposure to E7 expressing epithelial cells. PMID- 10850420 TI - The von Hippel-Lindau tumor suppressor targets to mitochondria. AB - Subcellular localization of von Hippel-Lindau (VHL) tumor suppressor may clarify its role in tumorigenesis. In rat kidney, we observed a granular cytoplasmic immunostaining of VHL, as seen in human tissues. The green fluorescent protein (GFP)-tagged VHL also appeared as cytoplasmic granules in vitro and was colocalized with a mitochondrion-selective dye. Immunogold electron microscopy localized VHL specifically to the mitochondrion. Mitochondria retaining GFP-VHL fusion protein, mimicking an insertional VHL mutant, displayed abnormal phenotypes. Among these, small mitochondria have been observed in clear cell renal carcinomas known to have frequent VHL alterations. Thus, VHL may contribute to tumorigenesis through mitochondria-based action. PMID- 10850421 TI - Role of the von Hippel-Lindau tumor suppressor protein during neuronal differentiation. AB - The von Hippel-Lindau (VHL) tumor suppressor protein down-regulates transcription by transcriptional elongation enhanced by antagonizing elongin B and C. Transcriptional regulation is an important control mechanism for embryogenesis and tumorigenesis. The VHL gene and protein are expressed in neuronal cells of the fetal and adult brain. However, the role of the VHL gene in the central nervous system (CNS) has not been elucidated. The VHL gene might modify the expression of various genes during embryogenesis and tumorigenesis in CNS. We investigated the role of the VHL gene in CNS development using rodent CNS progenitor cells. Here we show that expression of the VHL protein is correlated with neuronal differentiation but not with glial differentiation in CNS progenitor cells, and we also show that VHL gene transduction induces neuronal differentiation. In addition, a VHL mRNA antisense oligonucleotide inhibits differentiation of CNS progenitor cells and up-regulates their cell cycle. In conclusion, the VHL gene plays an essential role in neuronal differentiation as well as transcription. PMID- 10850422 TI - Interaction between Sp1 and cell cycle regulatory proteins is important in transactivation of a differentiation-related gene. AB - The stratified squamous epithelium is a model system in which to define molecular mechanisms underlying the switch from proliferation to differentiation. This can be achieved through the functional dissection of keratin gene promoters. Having previously established the importance of keratin 4 in maintaining the differentiated phenotype in corneal epithelial cells, we investigated the role of Sp1-mediated transactivation of the keratin 4 promoter given the role of Sp1 in differentiation and cell cycle progression. Sp1 transactivation of the keratin 4 promoter was diminished in cyclin D1-overexpressing cells, which may be mediated through a newly described direct interaction between Sp1 and cyclin D1 and opposed by a complex between Sp1 and pRB. PMID- 10850423 TI - Loss of p53 in benzene-induced thymic lymphomas in p53+/- mice: evidence of chromosomal recombination. AB - The purpose of this study was to examine the role of chromosomal recombination in mediating p53 loss in benzene-induced thymic lymphomas in C57BL/6-Trp53 haploinsufficient (N5) mice (p53+/- mice). We characterized loss of heterozygosity (LOH) on chromosome 11 using seven microsatellite markers in 27 benzene-induced and 6 spontaneous thymic lymphomas. Eleven patterns of LOH were found between the induced and spontaneous tumors, with only one pattern being in common between the tumor groups. Nearly 90% (24 of 27) of benzene-induced tumors exhibited loss of the functional p53 allele locus, and 83% (20 of 24) of these tumors retained two copies of the disrupted p53 allele. The results indicate that benzene induces a high frequency of LOH on chromosome 11 in p53+/- mice, likely mediated by aberrant chromosomal recombination. PMID- 10850424 TI - A region of deletion on chromosome 22q13 is common to human breast and colorectal cancers. AB - Chromosomal allelic losses have varying frequency in breast cancer, with key regions including chromosomes 1, 3p, 7q, 9p, 16q, 17, and 22q. Recently, we have been able to map a new target region of allelic loss on chromosome 22q involved in colorectal cancer. The aim of the current investigation was to determine whether this target region may also be involved in human breast carcinogenesis. Thirty-six pairs of matched normal and tumor specimens from breast cancer patients, as well as eight breast cancer-derived cell lines, were genotyped using 17 microsatellite markers spanning chromosome 22q. Allelic deletion was found in 19 of 36 tumors (53%), and the pattern observed in those cases with partial losses was consistent with a region flanked by D22S1171 and D22S928. This interval overlaps that identified in colorectal cancer and comprises nearly 1.1 Mb. This study provides evidence of a common region of deletion on chromosome 22q13 involved in both breast and colorectal cancers and underscores the existence of putative tumor suppressor gene(s) at this location. PMID- 10850425 TI - Expression of bone morphogenetic protein receptors type-IA, -IB and -II correlates with tumor grade in human prostate cancer tissues. AB - Bone morphogenetic proteins (BMPs) are potential regulators of prostate cancer cell growth and metastasis that signal through an interaction with BMP membrane receptors (BMPRs) type I and type II. In the present study, Western blot and immunohistochemical analysis of BMPRs were carried out in benign and malignant human prostate tissues to explain the loss of BMP response in human prostate cancer cells. The results demonstrated that the benign prostate specimens expressed high levels of all three BMPRs. In normal prostate, BMPRs were localized predominantly to epithelial cells. Among prostate cancer specimens, well-differentiated cancers were positive for the expression of BMPR-II, BMPR-IA, and BMPR-IB, for the most part. In contrast, only 1 of 10 poorly differentiated prostate cancer cases was positive for each of the three BMPRs (P < 0.005 for all three receptors). Taken together, these results indicate that human prostate cancer cells frequently exhibit loss of expression of BMPRs and suggest that loss of BMPRs may play an important role during the progression of prostate cancer. PMID- 10850426 TI - RET receptor expression in thyroid follicular epithelial cell-derived tumors. AB - The RET proto-oncogene encodes a receptor tyrosine kinase for transforming growth factor-beta-related neurotrophic factors, which include GDNF and neurturin. The expression of RET proto-oncogene was detected in several tissues, such as spleen, thymus, lymph nodes, salivary gland, and spinal cord, and in several neural crest derived cell lines. RET expression in the thyroid gland was reported to be restricted to neural crest-derived C cells. The presence of RET mRNA or protein has not yet been reported in thyroid follicular cells. We previously demonstrated the expression of oncogenic rearranged versions of RET in papillary thyroid carcinomas: tumors derived from thyroid follicular cells. To assess the expression of the normal RET proto-oncogene in follicular cells, we analyzed its expression in a panel of neoplasias originating from thyroid follicular epithelial cells: papillary carcinomas and both follicular adenomas and carcinomas. We also demonstrated the presence of RET normal transcripts in two follicular thyroid carcinoma lymph node metastases. Moreover, we found the presence of the RET/ELE1 transcript, the reciprocal complementary form of the oncogenic fusion transcript ELE1/RET, in a papillary thyroid carcinoma specimen expressing the RET/PTC3 oncogene, thus demonstrating that the RET promoter is active in those cells after rearrangement. Finally, we show that in a papillary carcinoma-derived cell line expressing the proto-RET receptor and the related GFRalpha2 co-receptor, GDNF treatment induced RET tyrosine phosphorylation and subsequent signal transduction pathway, indicating that RET could be active in thyroid follicular cells. PMID- 10850427 TI - Prostate adenocarcinoma cells release the novel proinflammatory polypeptide EMAP II in response to stress. AB - The proinflammatory protein endothelial monocyte-activating polypeptide II (EMAP II) was first detected in supernatants of murine tumor cells by virtue of its ability to stimulate endothelial-dependent coagulation in vitro. The purified protein has pleiotropic effects on endothelial cells, monocytes, and neutrophils; however, its function in vivo is unknown, and the mechanism whereby it is released from cells is poorly understood. We investigated the expression of EMAP II in human prostate adenocarcinoma specimens by immunohistochemistry and in LNCaP and DU-145 human prostate adenocarcinoma cells by reverse transcription PCR, flow cytometry, and Western blotting. We then examined the effects of chemical and physiological stress on release and processing of EMAP-II by LNCaP and DU-145 cells. These cells constitutively express a Mr 34,000 form of EMAP-II that is retained intracellularly. Exposure to agents that induce apoptosis or, in some cases, necrosis induces the release of the Mr 34,000 form and further processing to the Mr 27,000 and Mr 22,000 forms. Hypoxia, but not heat shock, is a potent inducer of release and processing of biologically active EMAP-II by LNCaP and DU-145 cells. We suggest that release of EMAP-II by prostate adenocarcinoma cells as a consequence of treatment with anticancer agents or as a result of constitutive hypoxia may potentiate the effects of those agents through the localized activation of host effector mechanisms. PMID- 10850428 TI - Impact of global genome repair versus transcription-coupled repair on ultraviolet carcinogenesis in hairless mice. AB - The nucleotide excision repair (NER) system is comprised of two subpathways, i.e., transcription-coupled repair (TCR) and global genome repair (GGR). To establish the relative importance of TCR and GGR for UV effects on the skin, we have used hairless knockout mouse strain lacking either TCR (CSB -/-) or GGR (XPC -/-). In single exposure experiments, we found that CSB -/- mice have a 7-16 times higher susceptibility to sunburn than XPC -/- mice and than heterozygous (+/-) and wild-type (+/+) controls. Exposure to 80 J/m2 UV radiation (i.e., suberythemogenic in CSB -/-) on 10 consecutive days gives rise to epidermal hyperplasia in CSB -/- and XPC -/-, whereas repair-proficient controls do not show epidermal hyperplasia from these exposures. In addition, CSB -/- mice develop marked parakeratosis, whereas XPC -/- mice and controls do not. Under continued exposure to this daily dose, squamous cell carcinomas appear in CSB -/ , XPC -/-, and in the control groups, whereas only in the CSB -/- animals is a fairly high number of benign papillomas also found. The median latency time of squamous cell carcinomas (diameters > or = 1 mm) is 84 days for the XPC -/- mice, 115 days for the CSB -/- mice, and 234-238 days for the heterozygous and wild type control groups. These results indicate that GGR is more important than TCR in protection against UV-induced carcinomas of the skin but not against other UV effects such as sunburn, epidermal thickening, scaling of the stratum corneum, and development of papillomas. These results also indicate that GGR capacity may serve as a better predictor for skin cancer susceptibility than sensitivity to sunburn. The relative cancer susceptibilities of GGR- and TCR-deficient skin could well depend on the balance between an increased mutation rate and the presence (in CSB -/-) or lack (in XPC -/-) of a compensatory apoptotic response. PMID- 10850429 TI - Beta-catenin mutations and protein accumulation in all hepatoblastomas examined from B6C3F1 mice treated with anthraquinone or oxazepam. AB - The molecular pathogenesis of hepatoblastomas in the B6C3F1 mouse is unclear but may involve alterations in the beta-catenin/Wnt signaling pathway as was recently described for chemically induced hepatocellular neoplasms and human liver cancers. The objective of this study was to characterize the mutation frequency and spectrum of beta-catenin mutations and the intracellular localization of beta catenin protein accumulation in chemically induced hepatoblastomas. In this study, beta-catenin mutations were identified in all 19 anthraquinone-induced hepatoblastomas and all 8 oxazepam-induced hepatoblastomas examined. Although several hepatoblastomas had multiple deletion and/or point mutations, the pattern of mutations in the hepatoblastomas did not differ from that identified in hepatocellular neoplasms. In a majority of the hepatoblastomas (six of seven) examined by immunohistochemical methods, both nuclear and cytoplasmic localization of beta-catenin protein were detected, whereas in hepatocellular adenomas, carcinomas, and normal liver only membrane staining was observed. Our data suggest that beta-catenin mutations and the subsequent translocation of beta catenin protein from the cell membrane to the cytoplasm and nucleus may be critical steps in providing hepatocellular proliferative lesions with the growth advantage to progress to hepatoblastoma. PMID- 10850430 TI - Retinoic acid receptor and retinoid X receptor alterations in lung cancer precursor lesions. AB - Smoking prevention will decrease lung cancer incidence in time. However, early detection would improve lung cancer prognosis in subjects at risk provided that specific markers could be identified. We previously reported that retinoic acid receptor (RAR) and retinoid X receptor (RXR) expression was altered in lung tumors. RAR-beta gene status could be derived from corresponding allelotyping and immunohistochemistry data. We now report the continued study on lung cancer precursor lesions. Fluorescence PCR-based assays were used for allelotyping at the RAR/RXR loci of: (a) 66 lung precursor lesions found at the free resection margins of 41 patients undergoing surgery for lung cancer (+ 31 paired tumors); and (b) bronchial cells also found at the free resection margins from 16 current and 8 never smokers operated on for noncancerous diseases. Three microsatellites located at 3p14-21 and 9p21 were also used for interwork comparison. Immunohistochemistry was additionally performed to evaluate P53 and RAR-beta expression in precursor lesions. Chi2 tests showed significant differences (P < 0.05) when comparing the results obtained from never smokers, smokers, squamous metaplasia, dysplasia + in situ carcinoma, and tumors. Microsatellite changes occurred frequently in all samples, but without specificity for any group (P < 0.08-0.52). They were globally correlated with tobacco exposure (P < 0.04), for which the RAR-gamma marker appeared as a preferential target (P < 0.004). Few reparation error phenotypes were observed, mostly at the RXR-alpha and RXR-gamma markers for which combined changes were also linearly increasing from never smokers to dysplasia + in situ carcinoma (P < 0.05 and P < 0.03). RAR-beta marker losses also increased from the first to the last group studied (P < 0.01), with a concomitant decrease in RAR-beta protein expression and correlated p53 increased immunoreactivity (P < 0.02). Losses at 3p14, 3p21, and P16 were frequent, but no significant differences between groups could be found. Unexpectedly, high constitutive homozygosity was observed near the RAR-alpha locus in squamous cell lung cancer cases. RARs/RXRs form homodimers or heterodimers involved in ligand binding. Their added alterations could result in a state of functional vitamin A deficiency in the affected bronchial cells. Further deletion events drawn from a limited repertoire of specific regions such as 3p14-21 and 9p21 could subsequently drive the deficient cells to invasive carcinoma. PMID- 10850431 TI - Genetic resistance to chemical carcinogen-induced preneoplastic hepatic lesions in DRH strain rats. AB - DRH strain rats were established by inbreeding a closed colony of Donryu rats continuously fed the chemical hepatocarcinogen 3'-methyl-4 dimethylaminoazobenzene for over 10 years. They are highly resistant to chemical induction of liver cancer and preneoplastic lesions. We studied the genetic basis of DRH resistance to preneoplastic lesions by analyzing 108 (F344 x DRH)F2 male rats fed 3'-methyl-4-dimethylaminoazobenzene for 7 weeks. Five parameters of preneoplastic liver lesions were selected for quantitative analysis: (a) number of glutathione S-transferase placental form-positive foci per unit area of liver section; (b) percentage area occupied by the foci; (c) average size of foci; (d) glutathione S-transferase placental form mRNA level; and (e) gamma glutamyltranspeptidase mRNA level. Furthermore, O6-methylguanine DNA methyltransferase and mannose 6-phosphatase/insulin-like growth factor 2 receptor mRNA levels were quantified. Composite interval mapping analysis showed that there were two remarkably significant clusters of quantitative trait loci affecting preneoplastic liver lesions on chromosomes 1 and 4. These clusters were designated collectively as Drh1 and Drh2, respectively. The functions of the recessive DRH allele of Drh1 and the semidominant DRH allele of Drh2 were to suppress the phenotypes of precancerous lesions. Each cluster showed two to three subpeaks in linkage likelihood plots, suggesting the presence of several closely linked quantitative trait loci affecting preneoplastic lesions. Possible candidate genes at each locus will be discussed. Expression of O6-methylguanine DNA methyltransferase and mannose 6-phosphatase/insulin-like growth factor 2 receptor did not affect DRH resistance to hepatocarcinogenesis, although they were polymorphic between DRH and F344 rats. PMID- 10850432 TI - In vitro and in vivo studies of methylseleninic acid: evidence that a monomethylated selenium metabolite is critical for cancer chemoprevention. AB - Previous research suggested that the beta-lyase-mediated production of a monomethylated selenium metabolite from Se-methylselenocysteine is a key step in cancer chemoprevention by this agent. In an attempt to affirm the concept, the present study was designed to evaluate the activity of methylseleninic acid, a compound that represents a simplified version of Se-methylselenocysteine without the amino acid moiety, thereby obviating the need for beta-lyase action. The in vitro experiments showed that methylseleninic acid was more potent than Se methylselenocysteine in inhibiting cell accumulation and inducing apoptosis in TM12 (wild-type p53) and TM2H (nonfunctional p53) mouse mammary hyperplastic epithelial cells, and these effects were not attributable to DNA damage, as determined by the comet assay. In general, methylseleninic acid produced a more robust response at one-tenth the concentration of Se-methylselenocysteine. It is possible that these cell lines may have only a modest ability to generate a monomethylated selenium species from Se-methylselenocysteine via the beta-lyase enzyme. In contrast, methylseleninic acid already serves as a preformed active monomethylated metabolite, and this could be an underlying reason why methylseleninic acid acts more rapidly and exerts a more powerful effect than Se methylselenocysteine in vitro. Interestingly, the distinction between these two compounds disappeared in vivo, where their cancer chemopreventive efficacies were found to be very similar to each other [in both methylnitrosourea and dimethylbenz(a)anthracene rat mammary tumor models]. The beta-lyase enzyme is present in many tissues; thus, animals have an ample capacity to metabolize Se methylselenocysteine systemically. Therefore, Se-methylselenocysteine would be expected to behave like methylseleninic acid if beta-lyase is no longer a limiting factor. Taken together, the present in vitro and in vivo results provide strong evidence in support of our earlier hypothesis that a monomethylated selenium metabolite is important for cancer chemoprevention. Methylseleninic acid could be an excellent tool, especially for molecular mechanism studies in cell culture, and some of these attributes are discussed. PMID- 10850433 TI - Sex differences in the activation of tamoxifen to DNA binding species in rat liver in vivo and in rat hepatocytes in vitro: role of sulfotransferase induction. AB - Previous work has indicated that metabolic activation of tamoxifen in rat liver cells involves cytochrome P450-mediated alpha-hydroxylation, followed by sulfate ester formation, mediated by hydroxysteroid sulfotransferase a (rHSTa), a member of the SULT2A subfamily, which efficiently metabolizes dehydroepiandrosterone. Because it is known that the expression of rHSTa and other SULT2A forms is substantially higher in female rats than in males, it might be predicted that tamoxifen would be a more potent liver carcinogen in females than in males. Yet tamoxifen has been shown to be equipotent in both sexes. To investigate this paradox, primary cultures of hepatocytes were prepared from Fischer F-344 rats and treated with tamoxifen (10 microM) or alpha-hydroxytamoxifen (1 microM). Rats were also treated with tamoxifen daily by gavage (0.12 mmol/kg/day) for up to 14 days. DNA was isolated from hepatocytes and liver and analyzed by 32P postlabeling. Liver cytosol fractions were prepared and analyzed for dehydroepiandrosterone sulfotransferase activity and SULT2A protein levels. In tamoxifen-treated hepatocytes and after a single dose of tamoxifen in vivo, DNA adduct formation in male cells was significantly lower than in female cells, 11- and 6-fold, respectively. However, with increasing daily doses of rats with tamoxifen, the adduct level in males increased to a level 89% of that in females by 14 days. Dehydroepiandrosterone sulfotransferase activity in male rat liver cytosols was only 17% of the activity of female cytosols after one dose of tamoxifen but 64% after 14 days of exposure to the compound. This increase in activity correlated with increases in the levels of SULT2A protein, detected by Western blotting. Western blotting did not allow the unambiguous identification of the induced SULT2A form(s). However, by using a specific reverse transcriptase/PCR technique, it was found that it was primarily rHSTa that was induced. Thus, after prolonged exposure to tamoxifen, DNA adduct formation and rHSTa expression in males are significantly closer to the levels in females than they are after initial exposure. These changes explain the similar susceptibility of male and female rats to tamoxifen carcinogenesis. PMID- 10850434 TI - Color Doppler vascularity index can predict distant metastasis and survival in colon cancer patients. AB - The purpose of this study was to investigate the clinical usefulness of the color Doppler vascularity index (CDVI) in patients with colon cancer before surgery. Forty-four patients with sonographically visible tumor mass of colon cancer were investigated. The CDVI of each tumor was determined using transabdominal color Doppler ultrasound. The CDVI was defined as the ratio of the number of the colored pixels within a tumor section to the number of total pixels in that specific tumor section and was calculated by using Encomate software (Electronic Business Machine Co. Ltd., Taipei, Taiwan). The correlation between the CDVI and clinicopathological factors, mode of recurrence, and patient survival was studied. For comparison, microvessel density (the mean number of microvessels in three areas of highest vascular density at x200 magnification) of the tumors of these 44 patients was also evaluated by using immunohistochemical staining of surgical specimens with anti-CD34. The microvessel density was not correlated with Dukes' classification, clinicopathological factors, and survival. The CDVI was significantly higher in the patients with lymph node metastases and vascular invasion than in those without such metastases and invasion (P = 0.006 and P = 0.0098, respectively). Moreover, in patients with a high CDVI (> 15%) and positive vascular invasion, survival was significantly poorer than in those with low CDVI (< or = 15%) and negative invasion (P = 0.0037 and 0.0039, respectively). Multivariate analysis indicated that liver metastasis, vascular invasion, and CDVI are independent prognostic factors in the patients with colon cancer. According to the mode of recurrence in 36 patients who underwent curative resection, the frequency of the distant organ recurrence was significantly higher in the high CDVI group (40%) than in the low CDVI group (0%). The CDVI is a good preoperative indicator of recurrence and patient survival in colon cancer. Thus, the CDVI may be helpful in stratifying patients for adjuvant therapy. PMID- 10850435 TI - Vascular endothelial growth factor (VEGF) in breast cancer: comparison of plasma, serum, and tissue VEGF and microvessel density and effects of tamoxifen. AB - The assessment of angiogenesis in breast cancer is of importance as a key indicator of survival and response to therapy. Circulating vascular endothelial growth factor (VEGF) measurements may provide a less subjective analysis than microvessel density (MVD) or immunohistochemical analysis of VEGF expression; however, most studies have used serum, which is now known to largely reflect platelet-derived VEGF concentrations. This study examined for the first time both plasma (VEGFp) and serum (VEGFs) VEGF concentrations in 201 blood samples from pre- and postmenopausal healthy controls and from patients with benign breast disease, localized breast cancer, breast cancer in remission, or metastatic breast cancer and related these to other clinicopathological markers. VEGFp but not VEGFs concentrations of patients with localized disease were significantly elevated compared with normal controls (P = 0.016). Patients with metastatic disease had higher VEGFp and VEGFs levels than normal controls (P < 0.001, P = 0.044 respectively), and higher VEGFp, but not VEGFs, than patients with benign disease (P = 0.009) and patients with localized disease (P = 0.004). However, the highest VEGFp and VEGFs concentrations were seen in patients in remission compared with normal controls (P < 0.001 and P = 0.008, respectively). VEGFp concentrations in patients in remission were also higher than in patients with benign disease (P = 0.01) or patients with localized disease (P = 0.005). Tamoxifen treatment was significantly associated with higher circulating and platelet-derived VEGF levels. Circulating VEGF did not correlate with any clinicopathological factor, including MVD or VEGF expression. VEGF expression was significantly correlated with estrogen receptor status and inversely correlated with tumor grade. MVD correlated with tumor size. Tamoxifen-induced increases in VEGF may be important in clinical prognosis or associated pathologies. PMID- 10850436 TI - p53 mutations and exposure to environmental tobacco smoke in a multicenter study on lung cancer. AB - Biomarker data may provide a way to strengthen the link between environmental tobacco smoke (ETS) exposure and lung cancer shown in epidemiological studies. We conducted a multicenter case-control study to investigate the association between ETS exposure and lung cancer in never-smokers using p53 mutations as a biomarker of tobacco-related carcinogenesis. Paraffin-embedded tissue or fresh tissue samples from 91 never-smokers and 66 smokers with histologically confirmed lung cancer and interview data about smoking habits and ETS exposure were analyzed for mutations in the p53 gene. Statistical analysis was performed using multivariate logistic regression. Among the lifelong nonsmokers, the overall mutation prevalence was 10% (nine cases). Among 48 never-smokers ever exposed to spousal ETS, 13% (six cases) showed mutations. Smokers exhibited 17 (26%) mutations. A 3 fold [odds ratio, 2.9; 95% confidence interval (CI), 1.2-7.2] increased risk of p53 mutation was observed for smokers as compared with all never-smokers combined (i.e., irrespective of ETS exposure). The increase was 4.4-fold (95% CI, 1.2 16.2) when compared with never-smokers without ETS exposure. Among never-smokers, the risk of mutation was doubled (odds ratio, 2.0; 95% CI, 0.5-8.7) for exposure to spousal ETS only, based on 6 exposed cases with mutation and 42 exposed cases without mutation. The risk was 1.5 (95% CI, 0.2-8.8) for those ever exposed to spousal or workplace ETS as compared with those never exposed to spousal or workplace ETS. For smokers, the most common mutation type was G:C to T:A transversion (31%), whereas G:C to A:T transitions were predominant among never smokers (57%). In conclusion, our study indicates a significant 3-4-fold increased risk of p53 mutation in smoking lung cancer cases, and it suggests that mechanisms of lung carcinogenesis in ETS-exposed never-smokers include mutations in the p53 gene, similar to that seen in smokers. However, the mutation patterns observed also suggest a difference between smokers and never-smokers. Clearly, additional investigations of the role of p53 mutation as a biomarker for tobacco related carcinogenesis, including that related to ETS, are indicated. PMID- 10850437 TI - Enhancement of Fas-mediated apoptosis in renal cell carcinoma cells by adriamycin. AB - Anti-Fas monoclonal antibody (mAb) kills Fas-expressing cells by apoptosis. Several anticancer agents also mediate apoptosis and may share common intracellular pathways leading to apoptosis with Fas. Thus, we reasoned that combination treatment of drug-resistant cells with anti-Fas mAb and drugs might overcome their resistance. We investigated whether anticancer agents enhance Fas mediated apoptosis and cytotoxicity against renal cell carcinoma (RCC) cells. Treatment of ACHN RCC cells with anti-Fas mAb in combination with 5-fluorouracil, vinblastine, IFN-alpha, or IFN-gamma did not overcome resistance to these agents. However, combination treatment with anti-Fas mAb and Adriamycin (ADR) resulted in a synergistic cytotoxic effect. Furthermore, synergy was also obtained even when the exposure time was shortened from 24 h to 8 or 2 h. Synergy was also achieved in four other RCC cell lines and five freshly derived human RCC cells. Treatment with anti-Fas mAb in combination with epirubicin or pirarubicin also resulted in a synergistic cytotoxic effect on ACHN cells. Similar results were achieved with a combination of humanized anti-Fas mAb and ADR. Incubation of ACHN cells with ADR augmented the expression of Fas and p53, but not Bcl-2, Bax, or caspase-3. However, the activity of caspase-3 itself was apparently enhanced after treatment with ADR alone or combined treatment with anti-Fas mAb. The synergy obtained in cytotoxicity with anti-Fas mAb and ADR was also achieved in apoptosis. Exposure of ACHN cells and freshly derived RCC cells to ADR enhanced their susceptibility to lysis by peripheral blood lymphocytes and tumor-infiltrating lymphocytes. This study demonstrates that combination treatment of RCC cells with anti-Fas mAb and ADR might overcome their resistance. The sensitization required a low concentration of ADR and a short exposure time, thus supporting the potential in vivo application of a combination of ADR and anti-Fas mAb or immunotherapy in the treatment of ADR- and/or immunotherapy-resistant RCC. PMID- 10850438 TI - Inhibition of translation initiation mediates the anticancer effect of the n-3 polyunsaturated fatty acid eicosapentaenoic acid. AB - Eicosapentaenoic acid (EPA), an n-3 polyunsaturated fatty acid that is abundant in the fish-based diets of populations that exhibit a remarkably low incidence of cancer, exerts anticancer activity in vitro and in animal models of experimental cancer. Here we define the molecular basis for the anticancer effects of EPA. EPA inhibits cell division by inhibiting translation initiation. This is a consequence of the ability of EPA to release Ca2+ from intracellular stores while inhibiting their refilling via capacitative Ca2+ influx that results in partial emptying of intracellular Ca2+ stores and thereby activation of protein kinase R. Protein kinase R phosphorylates and inhibits eukaryotic initiation factor 2alpha, resulting in inhibition of protein synthesis at the level of translation initiation, preferentially reducing the synthesis and expression of growth regulatory proteins, including G1 cyclins, and causes cell cycle arrest in G1. In a KLN-205 squamous cell carcinoma mouse model, daily oral administration of EPA resulted in a significant reduction of tumor size and expression of cyclin D1 in the tumor tissues. Furthermore, EPA-treated tumors showed a significant increase in the proportion of diploid cells, indicative of cell cycle arrest in G0-G1, and a significant reduction of malignant hypertetraploid cells. These results characterize EPA as a member of an emerging new class of anticancer compounds that inhibit translation initiaton. PMID- 10850439 TI - Blockade of the epidermal growth factor receptor signaling by a novel tyrosine kinase inhibitor leads to apoptosis of endothelial cells and therapy of human pancreatic carcinoma. AB - We determined whether down-regulation of the epidermal growth factor-receptor (EGF-R) signaling pathway by oral administration of a novel EGF-R tyrosine kinase inhibitor (PKI166) alone or in combination with gemcitabine (administered i.p.) can inhibit growth and metastasis of human pancreatic carcinoma cells implanted into the pancreas of nude mice. Therapy beginning 7 days after orthotopic injection of L3.6pl human pancreatic cancer cells reduced the volume of pancreatic tumors by 59% in mice treated with gemcitabine only, by 45% in those treated with PKI166 only, and by 85% in those given both drugs. The combination therapy also significantly inhibited lymph node and liver metastasis, which led to a significant increase in overall survival. EGF-R activation was significantly blocked by therapy with PKI166 and was associated with significant reduction in tumor cell production of VEGF and IL-8, which in turn correlated with a significant decrease in microvessel density and an increase in apoptotic endothelial cells. Collectively, our results demonstrate that oral administration of an EGF-R tyrosine kinase inhibitor decreased growth and metastasis of human pancreatic cancer growing orthotopically in nude mice and increased survival. The therapeutic effects were mediated in part by inhibition of tumor-induced angiogenesis attributable to a decrease in production of proangiogenic molecules by tumor cells and increased apoptosis of tumor-associated endothelial cells. PMID- 10850440 TI - Therapeutic efficacy of the suicide gene driven by the promoter of vascular endothelial growth factor gene against hypoxic tumor cells. AB - We examined whether herpes simplex virus thymidine kinase (HSV-TK) gene expression driven by the promoter of the vascular endothelial growth factor (VEGF) gene that is activated by hypoxia is effective in killing highly metastatic Lewis lung carcinoma A11 cells under hypoxic conditions. We isolated the promoter region encompassing the hypoxia response element (HRE) of the mouse VEGF gene. To assess the hypoxia responsiveness of the VEGF promoter, A11 cells were transiently transfected with luciferase reporter plasmids. Exposure of the transfectants to hypoxia resulted in a 2-3-fold induction of luciferase activity. Deletion of the HRE site abolished VEGF promoter activity under both normoxic and hypoxic conditions. We constructed a retroviral vector harboring the HSV-TK or green fluorescence protein (GFP) gene under the control of the VEGF promoter. A11 cells transfected with vector harboring the VEGF promoter fused to the HSV-TK gene [A11(HRE/TK) cells] were more sensitive to ganciclovir than cells transfected with the control vector harboring the VEGF promoter alone, and the sensitivity of the A11(HRE/TK) cells was increased by exposure to hypoxia followed by reoxygenation. Culturing A11 cells transfected with vector harboring the VEGF promoter fused to the GFP gene under hypoxic conditions resulted in an increase in the expression of GFP. Monitoring GFP expression and vascularity in the A11 transfectant tumors revealed up-regulation of GFP expression in poorly vascularized regions. Administration of ganciclovir to mice bearing s.c. tumors formed by A11(HRE/TK) cells resulted in regression of the tumors. These results suggest a possible application of the suicide gene driven by the VEGF promoter to cancer gene therapy that efficiently targets hypoxic tumor cells. PMID- 10850441 TI - Benzylidene lactam compound, KNK437, a novel inhibitor of acquisition of thermotolerance and heat shock protein induction in human colon carcinoma cells. AB - Cells exposed to heat or other types of stressors transiently synthesize a group of proteins known as heat shock proteins (HSPs). A nonlethal heat treatment can elicit in the cells an ability to resist subsequent lethal heat treatments. We report here that a novel benzylidene lactam compound, KNK437, dose-dependently inhibited the acquisition of thermotolerance and the induction of various HSPs including HSP105, HSP70, and HSP40 in COLO 320DM (human colon carcinoma) cells. The induction of heat-inducible HSP70, which is reported to be involved in the development of thermotolerance, was inhibited at mRNA levels by treatment with KNK437. This compound also inhibited the acquisition of thermotolerance as developed by sodium arsenite. However, it did not increase thermosensitivity in nontolerant cells. The effect of KNK437 was much greater than that of quercetin, a bioflavonoid that was previously reported to inhibit the acquisition of thermotolerance as well as the induction of HSPs. We conclude that this drug is a novel inhibitor of the acquisition of thermotolerance caused by the induction of HSPs. PMID- 10850442 TI - Bisphosphonates inhibit breast and prostate carcinoma cell invasion, an early event in the formation of bone metastases. AB - The molecular mechanisms by which tumor cells metastasize to bone are likely to involve invasion, cell adhesion to bone, and the release of soluble mediators from tumor cells that stimulate osteoclast-mediated bone resorption. Bisphosphonates (BPs) are powerful inhibitors of the osteoclast activity and are, therefore, used in the treatment of patients with osteolytic metastases. However, an added beneficial effect of BPs may be direct antitumor activity. We previously reported that BPs inhibit breast and prostate carcinoma cell adhesion to bone (Boissier et al., Cancer Res., 57: 3890-3894, 1997). Here, we provided evidence that BP pretreatment of breast and prostate carcinoma cells inhibited tumor cell invasion in a dose-dependent manner. The order of potency for four BPs in inhibiting tumor cell invasion was: zoledronate > ibandronate > NE-10244 (active pyridinium analogue of risedronate) > clodronate. In addition, NE-58051 (the inactive pyridylpropylidene analogue of risedronate) had no inhibitory effect, whereas NE-10790 (a phosphonocarboxylate analogue of risedronate in which one of the phosphonate groups is substituted by a carboxyl group) inhibited tumor cell invasion to an extent similar to that observed with NE-10244, indicating that the inhibitory activity of BPs on tumor cells involved the R2 chain of the molecule. BPs did not induce apoptosis in tumor cells, nor did they inhibit tumor cell migration at concentrations that did inhibit tumor cell invasion. However, although BPs did not interfere with the production of matrix metalloproteinases (MMPs) by tumor cells, they inhibited their proteolytic activity. The inhibitory effect of BPs on MMP activity was completely reversed in the presence of an excess of zinc. In addition, NE-10790 did not inhibit MMP activity, suggesting that phosphonate groups of BPs are responsible for the chelation of zinc and the subsequent inhibition of MMP activity. In conclusion, our results provide evidence for a direct cellular effect of BPs in preventing tumor cell invasion and an inhibitory effect of BPs on the proteolytic activity of MMPs through zinc chelation. These results suggest, therefore, that BPs may be useful agents for the prophylactic treatment of patients with cancers that are known to preferentially metastasize to bone. PMID- 10850443 TI - Cationic lipid:bacterial DNA complexes elicit adaptive cellular immunity in murine intraperitoneal tumor models. AB - Previous studies with a mycobacterial heat shock protein (hsp-65) have demonstrated some efficacy using cationic liposome-mediated gene transfer in murine i.p. sarcoma models. To further analyze the efficacy of hsp-65 immunotherapy in clinically relevant models of localized cancer, immunocompetent mice bearing i.p. murine mesothelioma were treated with four i.p. doses of a cationic lipid complexed with plasmid DNA (pDNA) containing hsp65, LacZ, or a null plasmid. We observed >90% long-term survival (median survival, 150 days versus approximately 25 days, treated versus saline control, respectively) in a syngeneic, i.p. murine mesothelioma model (AC29). Long-term survivors were observed in all groups treated with lipid complexed with any pDNA. Lipid alone or DNA alone provided no demonstrable survival advantage. In a more aggressive i.p. model of mesothelioma (AB12), we observed >40% long-term survival in groups treated with lipid:pDNA complexes, again irrespective of the transgene. To ask whether these antitumor effects had led to an adaptive immune response against the tumor cell, we rechallenged long-term survivors in both murine models s.c. with the parental tumor cell line. Specific, long-lasting systemic immunity against the tumor was readily demonstrated in both models (AB12 and AC29). Consistent with these results, splenocytes from long-term survivors specifically lysed the parental tumor cell lines. Depleting the CD8+ T-cells from the splenocyte pool eliminated this lytic activity. Lipid:pDNA treatment of athymic, SCID, and SCID/Beige mice bearing a murine i.p. mesothelioma (AC29) resulted in only a slight survival advantage, but there were no long-term survivors. Treatment of immunocompetent mice depleted of specific immune effector cells demonstrated roles for CD8+ and natural killer cells. Although the exact mechanism(s) responsible for these antitumor effects is unclear, the results are consistent with roles for both innate and adaptive immune responses. An initial tumor cell killing stimulated by cationic lipid:pDNA complexes appears to be translated into long-term, systemic immunity against the tumor cell. These results are the first to demonstrate that adaptive immunity against a tumor cell can be induced by the administration of lipid:pDNA complexes. Multiple administrations of cationic lipid complexed with pDNA lacking an expressed transgene could provide a promising generalized immune-mediated modality for treating cancer. PMID- 10850444 TI - Discovery and characterization of OC144-093, a novel inhibitor of P-glycoprotein mediated multidrug resistance. AB - OC144-093 is a novel substituted diarylimidazole (Mr 495) generated using the OntoBLOCK system, a solid-phase combinatorial chemistry technology, in combination with high-throughput cell-based screening. OC144-093 reversed multidrug resistance (MDR) to doxorubicin, paclitaxel, and vinblastine in human lymphoma, breast, ovarian, uterine, and colorectal carcinoma cell lines expressing P-glycoprotein (P-gp) with an average EC50 of 0.032 microM. Inhibition of MDR by OC144-093 was reversible, but the effect persisted for at least 12 h after removal of compound from the culture medium. OC144-093 had no effect on the response to cytotoxic agents by cells in vitro lacking P-gp expression or expressing a multidrug resistance-associated protein (MRP-1). OC144-093 was not cytotoxic by itself against 15 normal, nontransformed, or tumor cell lines, regardless of P-gp status, with an average cytostatic IC50 of >60 microM. OC144 093 blocked the binding of [3H]azidopine to P-gp and inhibited P-gp ATPase activity. The compound was >50% p.o. bioavailable in rodents and dogs and did not alter the plasma pharmacokinetics of i.v.-administered paclitaxel. OC144-093 increased the life span of doxorubicin-treated mice engrafted with MDR P388 leukemia cells by >100% and significantly enhanced the in vivo antitumor activity of paclitaxel in MDR human breast and colon carcinoma xenograft models, without a significant increase in doxorubicin or paclitaxel toxicity. The results demonstrate that OC144-093 is an orally active, potent, and nontoxic inhibitor of P-gp-mediated multidrug resistance that exhibits all of the desired properties for treatment of P-gp-mediated MDR, as well as for prevention of MDR prior to selection and/or induction of refractory disease. PMID- 10850445 TI - Porphyrin analogues as novel antagonists of fibroblast growth factor and vascular endothelial growth factor receptor binding that inhibit endothelial cell proliferation, tumor progression, and metastasis. AB - Fibroblast growth factors (FGFs) and vascular endothelial growth factor (VEGF) play a pivotal role in the multistep pathway of tumor progression, metastasis, and angiogenesis. We have identified a porphyrin analogue, 5,10,15,20 tetrakis(methyl-4-pyridyl)-21H,23H-porphine-tetra -p-tosylate salt (TMPP), as a potent inhibitor of FGF2 and VEGF receptor binding and activation. TMPP demonstrated potent inhibition of binding of soluble FGF receptor 1 (FGFR1) to FGF2 immobilized on heparin at submicromolar concentrations. TMPP inhibits binding of radiolabeled FGF2 to FGFR in a cell-free system as well as to cells genetically engineered to express FGFR1. Furthermore, TMPP also inhibits the binding of VEGF to its tyrosine kinase receptor in a dose-dependent manner. In an in vitro angiogenic assay measuring the extent of endothelial cell growth, tube formation, and sprouting, TMPP dramatically reduced the extent of the FGF2 induced endothelial cell outgrowth and differentiation. In a Lewis lung carcinoma model, mice receiving TMPP showed a marked inhibition of both primary tumor progression and lung metastases development, with nearly total inhibition of the metastatic phenotype upon alternate daily injections of TMPP at 25 microg/g of body mass. Finally, novel meso-pyridylium-substituted, nonsymmetric porphyrins, as well as a novel corrole-based derivative, with >50-fold increase in activity in vitro, had a significantly improved efficacy in blocking tumor progression and metastasis in vivo. PMID- 10850446 TI - Structure, function, and targeting of interleukin 4 receptors on human head and neck cancer cells. AB - Despite advances in diagnosis and treatment, survival rates for patients with head and neck cancer have remained unchanged for the last 30 years. In an attempt to develop novel therapeutic agents, we have observed that a variety of murine and human carcinoma cells expresses high levels of receptors for interleukin 4 (IL-4) in vitro and in vivo. Here, we demonstrate that 17 head and neck cancer cell lines also express surface IL-4 receptors (IL-4R) and IL-4 binds to IL-4R on one cell line studied with low affinity ((k)d = 37.9 +/- 0.4 nM). The investigation of the subunit structure of IL-4R demonstrated that head and neck cancer cell lines expressed mRNA for IL-4R beta chain (also known as IL-4R alpha) and IL-13R alpha' chain (also known as IL-13R alpha1). However, no cell line expressed IL-2R common gamma-chain, which is known to be shared with IL-4R in immune cells. IL-4R is functional because IL-4 strongly induced activation of signal transducers and activators of transcription 6 (STAT-6) in these cell lines. A fusion protein, IL4(38-37)-PE38KDEL, containing translocation and enzymatic domains of Pseudomonas exotoxin and a circularly permuted human IL-4 was found to be highly and specifically cytotoxic to IL-4R-positive head and neck cancer cells, as determined by protein synthesis inhibition assay and confirmed by clonogenic assay. IL4(38-37)-PE38KDEL induced DNA fragmentation and condensation of nuclei indicative of apoptotic cell death. These results establish uniform expression of IL-4R on head and neck cancer cell lines and IL-4 toxin IL4(38-37)-PE38KDEL as a novel therapeutic agent for the possible treatment of human head and neck cancers. PMID- 10850447 TI - Complete regression of xenografted human carcinomas by camptothecin analogue carboxymethyl dextran conjugate (T-0128). AB - Clinically available camptothecins (CPTs), such as irinotecan (CPT-11) and topotecan, represent one of the most promising classes of antitumor agents, despite their toxicity. To improve their pharmacological profiles, a new macromolecular prodrug, denoted T-0128, was synthesized. This prodrug is a novel CPT analogue (T-2513)-carboxymethyl (CM) dextran conjugate via a triglycine spacer, with a molecular weight of Mr 130,000. This study was designed to test the concept that the rational design of a CPT-polymer conjugate would increase the efficacy of the parent drug. The in vivo antitumor study against Walker-256 carcinoma demonstrated that T-0128 was 10 times as active as T-2513, supporting this concept. Additionally, comparative efficacy studies of T-0128, T-2513, CPT 11, and topotecan were performed using a panel of human tumor xenografts in nude mice, showing the advantage of T-0128 over these CPTs. The maximal tolerated doses (MTDs) of T-0128, T-2513, and CPT-11 were comparable. Even a single i.v. injection of T-0128 at 6 mg/kg (based on the amount of T-2513 bound to CM dextran) induced complete regression of MX-1 mammary carcinoma. T-0128 at 10 mg/kg weekly for 3 weeks (one-tenth of its MTD) cured LX-1 lung carcinoma. Also, T-0128 below its MTD consistently cured or regressed St-4 gastric and HT-29 colorectal tumor xenografts that are highly refractory to CPTs. These demonstrate the broad range of therapeutic doses achieved with T-0128. Pharmacokinetic studies were performed to correlate the efficacy results obtained for T-0128 with plasma and tissue drug concentrations using Walker-256 tumor-bearing rats. Results showed that after i.v. administration of T-0128, the conjugate continued to circulate at a high concentration for an extended period, resulting in tumor accumulation. In the tumor, the sustained release of T-2513 occurred. In contrast, T-2513 disappeared rapidly from the body. The significant increases in the amount and exposure time of released T-2513 in the tumor explain well the enhanced efficacy of T-0128. In conclusion, this study indicated that T-0128 improved the potency of T-2513, demonstrating the proof of the above concept. PMID- 10850448 TI - Inhibition of metastatic renal cell carcinomas expressing somatostatin receptors by a targeted cytotoxic analogue of somatostatin AN-238. AB - The effectiveness of chemotherapy targeted to somatostatin (SST) receptors based on cytotoxic SST analogue AN-238, consisting of 2-pyrrolinodoxorubicin (AN-201) linked to SST carrier octapeptide, was investigated in human renal cell carcinomas (RCCs). SST receptors, which showed high-affinity binding for AN-238, were found in the SW-839 RCC line with sst2A subtype and in the 786-0 RCC line, which expressed the sst5 subtype. CAKI-1 RCC, which does not express sst2A or sst5, was used as a negative control for testing the specificity of SST receptor targeting. Using microsatellite analysis, AN-238 was shown to selectively inhibit the proliferation of 786-0 line, but not the CAKI-1 RCC line in vitro. The effects of three i.v. injections of 150 nmol/kg of AN-238 or AN-201, given on days 1, 8, and 21, were evaluated in groups of nude mice bearing s.c. xenografts of SW-839 and 786-0 RCC. After 5 weeks, the volumes of SW-839 and 786-0 RCC tumors were decreased by 67.2 (P < 0.05) and 78.3% (P < 0.01), respectively, whereas AN-201 had no significant effect on tumor growth. The inhibition of SST receptor-negative CAKI-1 tumors by AN-238 was only marginal. To investigate the efficacy of SST receptor-targeted chemotherapy in metastatic RCC, three i.v. injections of AN-238 or AN-201 at 150 nmol/kg were given at biweekly intervals to nude mice implanted with 786-0 tumors under the renal capsule. After 6 weeks, the weight of orthotopic tumors treated with AN-238 (55.3 +/- 44.3 mg) was significantly lower (87% reduction; P < 0.001) than that in the control group (414.2 +/- 41.0 mg) or in animals given AN-201 (270.2 +/- 603 mg; P < 0.05). Five of six animals (83%), both in the control and the AN-201 group, developed metastases to lymph nodes, but only one of seven mice (14%) given AN-238 showed lymphatic spread. Lung metastases were found in 83% of controls and 50% of AN-201 treated animals, but none occurred in mice treated with AN-238. This study demonstrates that targeted cytotoxic SST analogue AN-238 provides an effective therapy for chemoresistant neoplasms such as RCC. Because most clinical RCCs express SST receptors, this treatment modality might be beneficial to patients with metastatic disease. PMID- 10850449 TI - Sequential loss of cytotoxic T lymphocyte responses to simian virus 40 large T antigen epitopes in T antigen transgenic mice developing osteosarcomas. AB - The role of CTL tolerance in tumor immunity to SV40 large T antigen (T ag) induced tumors was studied using T ag transgenic mice of the line 501 (H2b). 501 mice express SV40 T ag under the influence of the alpha-amylase promoter, which leads to the development of osteogenic osteosarcomas late in life and eventual death between 12 and 17 months of age. We determined the ability of 501 mice to respond to the four H2b-restricted T ag CTL epitopes, which include epitope I (T ag 206-215), epitope II/III (T ag 223-231), the immunorecessive epitope V (T ag 489-497), restricted by H2-Db, and epitope IV (T ag 404-411), restricted by H2 Kb. We demonstrate that 501 mice are partially tolerant to the H2b-restricted T ag epitopes. Immunization of 4-month-old 501 mice with T ag-transformed syngeneic cell lines or a recombinant vaccinia virus expressing full-length T ag elicited CTL responses against the H2-Kb-restricted T ag epitope IV only. In contrast, immunization of 4-month-old 501 mice with recombinant vaccinia viruses expressing individual T ag epitopes as minigenes elicited CTLs against epitopes I, IV, and V, but not against epitope II/III. Complete tolerance to epitopes I, IV, and V developed in 501 mice, but the age when tolerance was detected varied for each epitope. Tolerance to epitope I occurred by 6 months of age and was accelerated in the absence of CD4+ T cells. Tolerance to the immunorecessive epitope V was observed in 12-month-old 501 mice but was independent of the presence of osteosarcomas. In contrast, CTLs specific for epitope IV were detected in mice from 3 to 14 months of age but not in mice that had developed osteosarcomas. Analysis of epitope IV-specific CD8+ cells derived from 3-month-old 501 mice with H2-Kb/epitope IV tetramers revealed decreased numbers of epitope IV-specific CD8+ cells in 501 mice relative to C57BL/6 mice, with a further decrease in older 501 mice. Tumor progression resulted in loss of H2-Kb/epitope IV tetramer staining CD8+ cells. Thus, progression to tolerance to individual T ag CTL epitopes in 501 mice is epitope dependent. PMID- 10850450 TI - CD59 expressed on a tumor cell surface modulates decay-accelerating factor expression and enhances tumor growth in a rat model of human neuroblastoma. AB - It has been hypothesized that complement inhibitors expressed on the surface of tumor cells prevent effective immune-mediated clearance. Whereas there are in vitro data to support this hypothesis, the species-selective activity of complement inhibitors has been a hindrance to investigating the role of membrane bound complement inhibitors in rodent models of human cancer. The CD59-positive LAN-1 human neuroblastoma cell line was significantly more sensitive to lysis by rat complement than by human complement, illustrating the species selectivity of endogenously expressed complement inhibitors. Transfection of LAN-1 cells with rat CD59, an inhibitor of the terminal cytolytic membrane attack complex, effectively protected the cells from lysis by rat complement in vitro. When LAN-1 cells stably expressing rat CD59 were inoculated into immune-deficient rats, the onset of tumor growth and the rate of tumor growth were significantly enhanced compared with those of control-transfected LAN-1 cells. These data show directly that the expression of a complement inhibitor on a tumor cell promotes tumor growth. Flow cytometric analysis revealed that the endogenous expression of decay accelerating factor (DAF), an inhibitor of complement activation, was up regulated on the surface of cells after in vivo growth. Of further interest, higher levels of DAF were present on CD59-transfected cells than on control transfected cells derived from tumors. Increased DAF expression correlated with decreased complement deposition on the tumor cell surface. These results show that expression of complement inhibitors on a tumor cell has functional consequences with regard to complement deposition in vivo and indicate that CD59 can indirectly effect complement activation and C3 deposition in vivo via a link between CD59 and DAF expression. PMID- 10850451 TI - p53 transdominance but no gain of function in mouse brain tumor model. AB - Although p53 mutations in tumors typically result in loss of transactivation of p53 target genes some mutants display gain-of-function activity. The latter has important implications for the design of rational cancer therapy. We previously described a germ-line p53 mutation (deletion of codon 236, Y236delta) associated with a familial brain tumor syndrome. To determine whether this tissue-specific tumor predisposition reflects a gain-of-function activity of Y236delta or an effect of genetic background we have developed a mouse brain tumor model. Primary neuroectodermal cells deficient for p53 (+/- or -/-) and transduced with Y236delta using a retroviral vector were transplanted into the brain of adult wild-type mice. This neurografting paradigm circumvents the problem of early lethal tumors at extracerebral sites associated with germ-line p53 deficiency. Brain tumors arising in this mouse model were highly invasive, reflecting an important feature of the human disease. Tumors arose from p53+/- cells only when transduced with Y236delta. In keeping with in vitro data showing that Y236delta has dominant-negative activity, these tumors retained the endogenous wild-type p53 allele but accumulated high levels of Y236delta. However, the presence of Y236delta in transplanted p53-/- cells had no effect on the tumor frequency, 15% versus 27% without the mutant. In conclusion, Y236delta is transdominant but exerts no gain-of-function activity mediating a more penetrant tumor phenotype. PMID- 10850452 TI - DNA copy number changes in malignant ovarian germ cell tumors. AB - Malignant ovarian germ cell tumors (OGCTs) include immature teratomas (ITs), dysgerminomas (DGs), endodermal sinus tumors (ESTs), choriocarcinomas, and embryonal carcinomas. Knowledge about the genetic changes associated with malignant OGCT development is sparse. We therefore analyzed 25 OGCTs (12 DGs, 4 ESTs, and 9 ITs) for gains and losses by comparative genomic hybridization. In total, more gains than losses were observed, and the number of alterations ranged from 0-20 per tumor. The average number of changes among DGs, ESTs, and ITs was 10, 6, and 1.4, respectively. The most common changes in DGs were gains from chromosome arms 1p (33%), 6p (33%), 12p (67%), 12q (75%), 15q (42%), 20q (50%), 21q (67%), and 22q (58%); gains of the whole of chromosomes 7 (42%), 8 (42%), 17 (42%), and 19 (50%); and losses from 13q (58%). Two of three DGs with a gonadoblastoma component showed gains of 3p21 and loss of 5p, whereas none of the nine pure DGs had these changes, suggesting that they might be characteristic either of gonadoblastoma or of DG developing from a gonadoblastoma. Gain of 12p and gain from 1q were seen in three of four ESTs, whereas gains from 3p, 11q, and Xp and loss from 18q were each found in two tumors. Five of the ITs revealed changes (range, 1-4 changes/tumor), with gains from 1p, 16p, 19, and 22q each being found in two tumors. We conclude that ovarian DGs and ESTs seem to develop via the same genetic pathways that are already known for testicular germ cell tumors. On the other hand, ITs do not exhibit gain of 12p and also typically show fewer changes than other malignant OGCTs, indicating that they arise via different pathogenetic mechanisms. PMID- 10850453 TI - Mitogenic conversion of transforming growth factor-beta1 effect by oncogenic Ha Ras-induced activation of the mitogen-activated protein kinase signaling pathway in human prostate cancer. AB - Elevated expression of transforming growth factor (TGF)-beta1 has been implicated in prostate tumorigenesis despite its growth-inhibitory effect on normal epithelial and carcinoma cells of the prostate. In this study, we identified that G1-to-S transition of the cell cycle is stimulated by TGF-beta1 in the prostate cancer cell line TSU-Pr1. No mutation of signal mediators, including Smads, and induction of PAI-1 transcription indicated that the TGF-beta1 signaling cascade is functionally intact in this cell line. Whereas pharmacological inhibitors of various mitogenic signaling pathways showed no effects, blockade of the mitogen activated protein kinase (MAPK) pathway by the MAPK kinase 1 inhibitor PD98059 restored the growth inhibitory role of TGF-beta1 in TSU-Pr1, which carries an oncogenic mutation in Ha-Ras (V12). Moreover, expression of antisense Ha-Ras or dominant negative Raf-1 abrogated the mitogenic effect of TGF-beta1 in TSU-Pr1, and the TGF-beta1 inhibition of DU145 was switched to stimulation by V12Ha-Ras transfection. Whereas the negative growth regulation by TGF-beta1 was completely inhibited by dominant negative Smad2, Smad3, or Smad4, its mitogenic effect was not affected, suggesting that this action is Smad-independent. Interestingly, whereas the TGF-beta1-mediated up-regulation of p15INK4B and p21WAF1 transcription was abolished in TSU-Pr1 and V12Ha-Ras-transfected DU145, inhibition of the Ras/MAPK pathway restored the TGF-beta1 induction of these genes. Taken together, our data suggest that prostate carcinomas with the Ras/MAPK pathway activation might have a selective growth advantage by autocrine TGF-beta1 production. PMID- 10850454 TI - Allelic loss in the progression of myelodysplastic syndrome. AB - To elucidate the genetic events that may play an important role in the progression of myelodysplastic syndrome (MDS) to acute myeloid leukemia (AML), we performed allelotype analysis of 24 individuals using matched MDS and AML samples from the same patients. Because the evolution can take years to occur, we used DNAs extracted from archival samples. These samples were analyzed with 79 microsatellite markers, which mapped to each of the autosomal arms except the short arms of the acrocentric chromosomes. Loss of heterozygosity on at least one locus was observed in 18 of the 24 cases (75%) as the disease progressed. Frequent allelic loss in >20% of the informative cases was observed on chromosome arms 6q (31%), 7p (23%), 10p (31%), 11q (27%), 14q (25%), and 20q (23%). Although cytogenetic information was available for many of our cases with allelic loss on 6q, 7p, 10p, 11q, 14q, and 20q, no deletions were observed on these arms. Fractional allelic loss, calculated for each sample as the total number of chromosomal arms lost per total number of arms with information, showed a median value of 0.06 and a mean of 0.15 (range, 0-0.59). No microsatellite instability at more than one marker was found in any of the samples. These results suggest that tumor suppressor genes exist on 6q, 7p, 10p, 11q, 14q, and 20q that have an important role in the evolution of MDS to AML when they are mutated. PMID- 10850455 TI - Overexpression of CDC25B phosphatase as a novel marker of poor prognosis of human colorectal carcinoma. AB - There is evidence to suggest that CDC25B phosphatase is an oncogenic protein. To elucidate the role of CDC25B in colorectal carcinoma, we examined the expression of CDC25B at the mRNA and protein levels. Reverse transcription-PCR assay indicated that CDC25B was overexpressed in tumor tissues relative to normal mucosa in 6 of 10 cases. Using immunohistochemistry, we identified high expression of CDC25B in 77 of 181 colorectal cases (43%). Univariate analysis showed that high expression was a significant predictor for poor prognosis compared with low expression (5-year survival rate; 59% versus 82%, respectively; P < 0.0001). Multivariate analysis indicated that CDC25B was an independent prognostic marker (risk ratio for death, 3.7; P < 0.0001) even after controlling for various factors such as lymph node metastasis, tumor size, degree of differentiation, and depth of invasion. Furthermore, the level of CDC25B expression clearly predicted the outcome of patients with Dukes' B and Dukes' C tumors. On the other hand, CDC25A mRNA was overexpressed in 9 of 10 colorectal cancer cases, and immunohistochemistry for CDC25A showed high expression in 52 of 111 cases (47%), but no significant correlation with prognosis. Our findings suggest that CDC25B is a novel independent prognostic marker of colorectal carcinoma and that it may be clinically useful for selecting patients who could benefit from adjuvant therapy. PMID- 10850456 TI - Tumor necrosis factor-related apoptosis-inducing ligand retains its apoptosis inducing capacity on Bcl-2- or Bcl-xL-overexpressing chemotherapy-resistant tumor cells. AB - Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the tumor necrosis factor family and has recently been shown to exert tumoricidal activity in vivo in the absence of any observable toxicity. The signaling pathways triggered by TRAIL stimulation and the mechanisms involved in resistance against TRAIL-mediated apoptosis are still poorly defined. We show here that TRAIL-induced apoptosis involves late dissipation of mitochondrial membrane potential (delta psi(m)) and cytochrome c release. These events follow activation of caspase-8 and caspase-3 and induction of DNA fragmentation. In addition, caspase-8-deficient cells are resistant against TRAIL-induced apoptosis, and inhibition of caspase-8 but not caspase-9 prevents mitochondrial permeability transition and apoptosis. In contrast, various Bcl-2- or Bcl-xL-overexpressing tumor cell lines are sensitive to TRAIL-induced apoptosis; however, they show a delay in TRAIL-induced mitochondrial permeability transition compared with control transfectants. This indicates that TRAIL-induced apoptosis depends on caspase-8 activation rather than on the disruption of mitochondrial integrity. Because most chemotherapeutic drugs used in the treatment of malignancies lead to apoptosis primarily by engagement of the mitochondrial proapoptotic machinery, we tested whether drug-resistant tumor cells retain sensitivity for TRAIL-induced apoptosis. Tumor cells overexpressing Bcl-2 or Bcl-xL become resistant to apoptosis induced by the chemotherapeutic drug etoposide. However, these cells are not protected or are only marginally protected against TRAIL-induced apoptosis. Thus, TRAIL may still kill tumors that have acquired resistance to chemotherapeutic drugs by overexpression of Bcl-2 or Bcl-xL. These data will influence future treatment strategies involving TRAIL. PMID- 10850457 TI - Castration-induced up-regulation of insulin-like growth factor binding protein-5 potentiates insulin-like growth factor-I activity and accelerates progression to androgen independence in prostate cancer models. AB - Although insulin-like growth factor binding protein-5 (IGFBP-5) has been shown to be implicated in prostate cancer progression, the functional role of IGFBP-5 in progression to androgen-independence remains largely undefined. Here, we demonstrate substantial up-regulation of IGFBP-5 during castration-induced regression and androgen-independent (AI) progression in the mouse androgen dependent (AD) Shionogi tumor model. To analyze the functional significance of these changes in IGFBP-5, human AD LNCaP prostate cancer cells were stably transfected with IGFBP-5 gene, and IGFBP-5-overexpressing LNCaP tumors progressed significantly faster to androgen independence after castration compared with controls. Antisense mouse IGFBP-5 oligodeoxynucleotides (ODNs) were then designed that reduced IGFBP-5 expression in Shionogi tumor cells in vitro in a dose dependent and sequence-specific manner. Growth of Shionogi tumor cells was inhibited by antisense IGFBP-5 ODN treatment in a time- and dose-dependent manner, which could be reversed by exogenous IGF-I. However, antisense IGFBP-5 ODN treatment had no additive inhibitory effect on Shionogi tumor cell growth when IGF-I activity was neutralized by anti-IGF-I antibody. Antisense IGFBP-5 ODN treatment resulted in decreased mitogen-activated protein kinase activity and number of cells in the S + G2-M phases of the cell cycle that directly correlated with reduced proliferation rate of Shionogi tumor cells. Systemic administration of antisense IGFBP-5 ODN in mice bearing Shionogi tumors after castration significantly delayed time to progression to androgen independence and inhibited growth of AI recurrent tumors. These findings suggest that up-regulation of IGFBP 5 after castration serves to enhance IGF bioactivity and raise the possibility that the response of prostate cancer to androgen withdrawal can be enhanced by strategies, such as antisense IGFBP-5 ODN therapy, that target IGF signal transduction. PMID- 10850458 TI - Arsenic trioxide-mediated growth inhibition in MC/CAR myeloma cells via cell cycle arrest in association with induction of cyclin-dependent kinase inhibitor, p21, and apoptosis. AB - We investigated the in vitro effect of As2O3 on proliferation, cell cycle regulation, and apoptosis in human myeloma cell lines. As2O3 significantly inhibited the proliferation of all of eight myeloma cell lines examined in a dose dependent manner with IC50 of approximately 1-2 microM. DNA flow cytometric analysis indicated that As2O3 (2 microM) induced a G1 and/or a G2-M phase arrest in these cell lines. To address the mechanism of the antiproliferative effect of As2O3, we examined the effect of As2O3 on cell cycle-related proteins in MC/CAR cells in which both G1 and G2-M phases were arrested. Western blot analysis demonstrated that treatment with As2O3 (2 microM) for 72 h did not change the steady-state levels of CDK2, CDK4, cyclin D1, cyclin E, and cyclin B1 but decreased the levels of CDK6, cdc2, and cyclin A. The mRNA and protein levels of CDKI, p21 were increased by treatment with As2O3, but those of p27 were not. In addition, As2O3 markedly enhanced the binding of p21 with CDK6, cdc2, cyclin E, and cyclin A compared with untreated control cells. Furthermore, the activity of CDK6-associated kinase was reduced in association with hypophosphorylation of Rb protein. The activity of cdc2-associated kinase was decreased, which was accompanied by the up-regulation of cdc2 phosphorylation (cdc2-Tyr15 phosphorylation) resulting from reduction of cdc25B and cdc25C phosphatases. As2O3 also induced apoptosis in MC/CAR cells as evidenced by flow cytometric detection of sub-G1 DNA content and annexin V binding assay. This apoptotic process was associated with down-regulation of Bcl-2, loss of mitochondrial transmembrane potential (delta psi(m)), and an increase of caspase-3 activity. These results suggest that As2O3 inhibits the proliferation of myeloma cells, especially MC/CAR cells, via cell cycle arrest in association with induction of p21 and apoptosis. PMID- 10850459 TI - Expression of polysialic acid and STX, a human polysialyltransferase, is correlated with tumor progression in non-small cell lung cancer. AB - Polysialic acid (PSA) is a carbohydrate composed of a linear homopolymer of alpha 2-8-linked sialic acid residues and is mainly attached to the neural cell adhesion molecule (NCAM). Because of the large negative charge of PSA, presence of PSA attenuates the adhesive property of NCAM and increases the cellular motility. PSA expression on NCAM is developmentally regulated, and PSA plays important roles in formation and remodeling of the neural system through regulation of the adhesive property of NCAM. Expression of the polysialated form of NCAM has been also demonstrated in some malignant tumors, such as Wilms' tumor and small cell lung cancer. Despite the possible importance as an onco developmental antigen, however, significance of PSA expression in most malignant tumors has not been revealed. Therefore, PSA expression in non-small cell lung cancer was assessed in the present study. PSA was expressed only in 5 (20.8%) of 24 pathological stage I cases, whereas it was expressed in most stage IV cases (76.8%, 11 of 14 cases). PSA expression was correlated with nodal metastasis and distant metastasis, but not with local extent of the primary tumor. Next, expression of polysialyltransferase genes (PST and STX genes) which controlled formation of PSA, was examined. The PST gene was constantly expressed in both normal lung tissue and tumor tissue of all cases. In contrast, the STX gene was not expressed in normal lung tissue of any case, and STX gene expression in tumor tissue was closely correlated with tumor progression. The STX gene was expressed only in 1 (4.2%) of 24 stage I cases, whereas it was expressed in most stage IV cases (85.7%, 12 of 14 cases). These results suggested that the PSA and STX genes could be new targets of cancer therapy as well as important clinical markers. PMID- 10850460 TI - Epidermal growth factor receptor vIII enhances tumorigenicity in human breast cancer. AB - Epidermal growth factor receptor vIII (EGFRvIII) is a tumor-specific, ligand independent, constitutively active variant of the EGFR. Its expression has been detected in gliomas and various other human malignancies. To more fully characterize the function and potential biological role of EGFRvIII in regulating cell proliferation and in tumorigenesis, we transfected EGFRvIII cDNA into a nontumorigenic, interleukin 3 (IL-3)-dependent murine hematopoietic cell line (32D cells). We observed 32D cells expressing high levels of EGFRvIII (32D/EGFRvIII P5) to be capable of abrogating the IL-3-dependent pathway in the absence of ligands. In contrast, the parental cells, 32D/EGFR, 32D/ErbB-4, and 32D/ErbB-2+ErbB-3 cells, all depended on IL-3 or EGF-like ligands for growth. 32D/EGFRvIII P5 cells subjected to long-term culture conditions in the absence of IL-3 revealed further elevation of EGFRvIII expression levels. These results suggested that the IL-3-independent phenotype is mediated by EGFRvIII. The level of expression is a critical driving force for the IL-3-independent phenotype. Dose-response analysis revealed 32D/EGFRvIII cells to require 500-fold higher concentrations (50 ng/ml) of EGF to further stimulate the EGF-mediated proliferation than in the 32D/EGFR cells (100 pg/ml). Similar effects were also observed in beta-cellulin-mediated proliferation. Moreover, 32D cells expressing high levels of EGFRvIII formed large tumors in nude mice, even when no exogenous EGF ligand was administered. In contrast, no tumors grew in mice injected with 32D/EGFR, 32D/ErbB-4, and 32D/ErbB-2+ErbB-3 cells or low-expressing clone 32D/EGFRvIII C2 cells or the parental 32D cells. The changes of the ligand specificity support the notion for an altered conformation of EGFRvIII to reveal an activated ligand-independent oncoprotein with tumorigenic activity analogous to v-erbB. These studies clearly demonstrate that EGFRvIII is capable of transforming a nontumorigenic, IL-3-dependent murine hematopoietic cell line (32D cells) into an IL-3-independent and ligand-independent malignant phenotype in vitro and in vivo. To delineate the biological significance of EGFRvIII in human breast cancer, we expressed EGFRvIII in the MCF-7 human breast cancer cell line. Expression of EGFRvIII in MCF-7 cells produced a constitutively activated EGFRvIII receptor. Expression of EGFRvIII in MCF-7 cells also elevated ErbB-2 phosphorylation, presumably through heterodimerization and cross-talk. These MCF 7/EGFRvIII transfectants exhibited an approximately 3-fold increase in colony formation in 1% serum with no significant effect observed at higher percentages of serum. A similar result was also seen in anchorage-dependent assays. Furthermore, EGFRvIII expression significantly enhanced tumorigenicity of MCF-7 cells in athymic nude mice with P < 0.001. Collectively, these results provide the first evidence that EGFRvIII could play a pivotal role in human breast cancer progression. PMID- 10850461 TI - Vascular endothelial growth factor/KDR activated microvessel density versus CD31 standard microvessel density in non-small cell lung cancer. AB - Vascular endothelial growth factor (VEGF) is an important angiogenic factor, linked to poor outcome in human malignancies including non-small cell lung carcinoma (NSCLC). We used the 11B5 monoclonal antibody recognizing the VEGF/KDR complex (R. A. Brekken et al., Cancer Res., 58: 1952-1959, 1998) to assess the VEGF expression in cancer cells and the VEGF/KDR activated microvessel density (aMVD) in early operable NSCLC. The JC70 anti-CD31 monoclonal antibody was used to assess the standard MVD (sMVD). The aMVD was significantly higher in the invading front of the tumors and in the normal lung adjacent to the tumors as compared with normal lung distant to the tumor or to inner tumor areas (P < 0.0002). The sMVD was higher in the normal lung and decreased from the invading front to inner tumor areas (P < 0.0001). However, the vascular activation (aMVD:sMVD) was 4-6 times higher in the tumor areas as compared with lung from normal individuals (36-58% versus 9%; P < 0.0001). Fibroblast 11B5 reactivity, noted in 25% of cases, correlated with high aMVD and sMVD in the inner tumor areas. Multivariate analysis showed that aMVD was the most potent and independent prognostic factor (P = 0.001; t-ratio, 3.28). It is concluded that intense VEGF/KDR angiogenic pathway activation is a tumor-specific feature in more than 50% of NSCLC cases and is associated with poor postoperative outcome. Clinical trials involving targeting of the VEGF/KDR-positive vasculature with specific antibodies, such as 11B5, are, therefore, encouraged. PMID- 10850462 TI - In vivo induction of insulin-like growth factor-I receptor and CD44v6 confers homing and adhesion to murine multiple myeloma cells. AB - One of the main characteristics of multiple myeloma (MM) cells is their specific homing and growth in the bone marrow (BM). Differences between stroma-dependent and -independent MM cell lines may reveal key molecules that play important roles in their homing to the BM. We addressed this topic with a murine MM model, including the in vivo 5T33MM (5T33MMvv) stroma-dependent cell line and its in vitro stroma-independent variant (5T33MMvt). Fluorescence-activated cell-sorting analysis showed expression of insulin-like growth factor (IGF)-I receptor and CD44v6 on all 5T33MMvv cells but not on 5T33MMvt cells. Checkerboard analysis and adhesion assays revealed IGF-I-dependent chemotaxis toward BM-conditioned medium and involvement of CD44v6 in the adhesion to BM stroma of only 5T33MMvv cells. However, when 5T33MMvt cells were injected in vivo (5T33MMvt-vv), after 18 h the MM cells harvested from BM were IGF-I receptor and CD44v6 positive. This up regulation was confirmed in 5T33MMvt-vv cells isolated from terminally diseased animals. These ST33MMvt-vv cells exhibited IGF-I-dependent chemotaxis and CD44v6 dependent adhesion to BM stroma. In vitro culture of the 5T33MMvt-vv cells could completely down-regulate IGF-I receptor and CD44v6. In fact, we could show that direct contact of 5T33MMvt cells with BM endothelial cells is a prerequisite for IGF-I receptor and CD44v6 up-regulation. These data indicate that the BM microenvironment is capable of up-regulating molecules such as IGF-I receptor and CD44v6, which facilitate homing of MM cells to the BM and support their adhesion to BM stroma. PMID- 10850463 TI - Vasoactive intestinal peptide/pituitary adenylate cyclase-activating peptide receptor subtypes in human tumors and their tissues of origin. AB - The evaluation of peptide receptors in man is needed not only to discover the physiological target tissues of a given peptide but also to identify diseases with a sufficient receptor overexpression for diagnostic or therapeutic interventions. Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) receptors have been evaluated in human tumors and in their tissues of origin using in vitro receptor autoradiography with 125I VIP or 125I-acetyl-PACAP-27 in tissue sections. The VIP/PACAP receptor subtypes VPAC1, VPAC2, and PAC1 were evaluated in these tissues by determining the rank order of potencies of VIP and PACAP as well as VPAC1- and VPAC2-selective analogues. The VIP/PACAP receptors expressed in the great majority of the most frequently occurring human tumors, including breast (100% receptor incidence), prostate (100%), pancreas (65%), lung (58%), colon (96%), stomach (54%), liver (49%), and urinary bladder (100%) carcinomas as well as lymphomas (58%) and meningiomas (100%), are predominantly of the VPAC1 type. Their cells or tissues of origin, i.e., hepatocytes, breast lobules and ducts, urothelium, prostate glands, pancreatic ducts, lung acini, gastrointestinal mucosa, and lymphocytes, also predominantly express VPAC1. Leiomyomas predominantly express VPAC2 receptors, whereas paragangliomas, pheochromocytomas, and endometrial carcinomas preferentially express PAC1 receptors. Conversely, VPAC2 receptors are found mainly in smooth muscle (i.e., stomach), in vessels, and in stroma (e.g., of the prostate), whereas PAC1 receptors are present in the adrenal medulla and in some uterine glands. Whereas the very wide distribution of VIP/PACAP receptors in the normal human body is indicative of a key role of these peptides in human physiology, the high VIP/PACAP receptor expression in tumors may represent the molecular basis for clinical applications of VIP/PACAP such as in vivo scintigraphy and radiotherapy of tumors as well as VIP/PACAP analogue treatment for tumor growth inhibition. PMID- 10850464 TI - Elevation of Vernier thresholds during image motion depends on target configuration. AB - Previously we showed that thresholds for abutting Vernier targets are unaffected by motion, as long as the targets are processed by the same spatial-frequency channel at each velocity and remain equally detectable [Invest. Ophthalmol. Visual Sci. (Suppl.) 37, S734 (1996)]. In this study we compared Vernier thresholds for stationary and moving abutting and nonabutting targets (gaps = 0, 18, and 36 arc min) for velocities of 0-16 deg/s. The Vernier targets were spatially filtered vertical lines (peak spatial frequency = 3.3 or 6.6 c/deg), presented at contrast levels of two, four, and eight times the detection threshold of each component line. Unlike the results for abutting targets, Vernier thresholds for nonabutting targets worsen with velocity as well as gap size. The results for abutting Vernier targets are consistent with the hypothesis that thresholds are mediated by oriented spatial filters, whose responses increase proportionally with the stimulus contrast. The velocity-dependent thresholds found for nonabutting Vernier targets can be explained on the basis of local-sign comparisons if the comparison process is assumed to include a small amount of temporal noise. PMID- 10850465 TI - Corneal wave aberration from videokeratography: accuracy and limitations of the procedure. AB - A procedure to calculate the wave aberration of the human cornea from its surface shape measured by videokeratography is presented. The wave aberration was calculated as the difference in optical path between the marginal rays and the chief ray refracted at the surface, for both on- and off-axis objects. The corneal shape elevation map was obtained from videokeratography and fitted to a Zernike polynomial expansion through a Gram-Schmidt orthogonalization. The wave aberration was obtained also as a Zernike polynomial representation. The accuracy of the procedure was analyzed. For calibrated reference surface elevations, a root-mean-square error (RMSE) of 1 to 2 microm for an aperture 4-6 mm in diameter was obtained, and the RMSE associated with the experimental errors and with the fitting method was 0.2 microm. The procedure permits estimation of the corneal wave aberration from videokeratoscopic data with an accuracy of 0.05-0.2 microm for a pupil 4-6 mm in diameter, rendering the method adequate for many applications. PMID- 10850466 TI - Method for recovery of heading from motion. AB - A new method for recovery of heading from motion is developed on the basis of Longuet-Higgins and Prazdny's algorithm [Proc. R. Soc. London Ser. B 208, 385 (1980)]. In the algorithm a radial virtual flow field is generated and the difference between the original velocity field and the virtual radial field is computed. The difference vectors, which are directed to the heading point in the projected plane, allow us to estimate the direction of heading. The simulations of the algorithm were performed, and it was shown that the method estimates the direction of heading accurately. PMID- 10850467 TI - Laser Ray Tracing versus Hartmann-Shack sensor for measuring optical aberrations in the human eye. AB - A comparison and validation study of Laser Ray Tracing (LRT) and Hartmann-Shack wave-front-sensor (to be referred to as H-S) methods was carried out on both artificial and human eyes. The aim of this work was double. First, we wanted to verify experimentally the equivalence of single- and double-pass measurements for both H-S and LRT. This interest is due to the impossibility of making single-pass measurements in human eyes. In addition, we wanted to validate the LRT technique by comparing it with the H-S wave-front sensor, currently used in many physiological optics laboratories. Comparison of the different methods and configurations carried out in the artificial eye yielded basically the same results in all cases, which means a reciprocal validation of both LRT and H-S, in either single- or double-pass configurations. Other aspects, such as robustness against speckle noise or the influence of the size of the entrance (H-S) or exit (LRT) pupil were studied as well. As a global reference, the point-spread function (PSF) of the artificial eye was recorded directly on a CCD camera and compared with simulated PSF's computed from the experimental aberration data. We also applied these two methods to real eyes (double pass), finding again a close match between the resulting aberration coefficients and also between the standard errors for two normal subjects. However, for one myopic eye with an especially low optical quality (RMS wave-front error >2 microm) and asymmetric aberrations, the array of spots recorded with the H-S sensor was highly distorted and too difficult to analyze. PMID- 10850468 TI - Channel selection with non-white-noise masks. AB - It is frequently assumed that the accuracy with which luminance gratings can be detected depends solely on the signal-to-noise ratio at the output of a single linear channel. Proportionality between threshold elevation and power spectral density is implicit in this assumption. I demonstrate that this proportionality does not hold for 1-cycle/degree gratings masked by low-pass noise with a 0.5 cycle/degree cutoff frequency. This implies that different channels can mediate detection, depending on the contrast of masking stimuli. PMID- 10850469 TI - Evidence for the contribution of S cones to the detection of flicker brightness and red-green. AB - We were interested in the question of how cones contribute to the detection of brightness, red-green, and blue-yellow. The linear combination of cone signals contributing to flicker detection was determined by fitting a plane to 64 points (colors) of equal heterochromatic flicker brightness. A small S-cone contribution to flicker brightness of similar amplitude in all five subjects was identified. The ratio of L- to M-cone contribution was found to vary considerably among subjects (1.7-4.1). Chromatic detection thresholds were determined for small patches in the isoluminant plane defined by flicker brightness. These stimuli were presented at an eccentricity of 40 arc min. By using color naming at the detection threshold, one can attribute different segments of the resulting detection ellipses to different chromatic mechanisms. Linear approximation of these segments provided an estimate for the contribution of the different cone types to the detection of red-green and blue-yellow. The results are consistent with the hypothesis that S cones contribute to the red-green mechanism with the same sign as that of the contribution from L cones. The blue-yellow mechanism very probably subtracts S-cone contrast from luminance contrast. The detection ellipse can be mapped into a circle in cone difference space. The base of this canonical transformation is a set of three cone fundamentals that differs from previously published estimates. Projecting the circle onto the three cone difference axes produces sinusoidal changes within the respective excitations. We propose that simultaneous sinusoidal changes of equal increment in the three cone difference excitations generate stimuli differing by equal saliency. PMID- 10850470 TI - Fractional Talbot effect in a tapered gradient-index medium: unit cell AB - A generalization of the fractional Talbot effect to the case of a tapered gradient-index medium for uniform illumination is considered. A unit cell of the fractional Talbot image contains the superposition of unit cell images of the periodic object. PMID- 10850471 TI - Anomalous refractive properties of photonic crystals AB - We describe methods of investigating the behavior of photonic crystals. Our approach establishes a link between the dispersion relation of the Bloch modes for an infinite crystal (which describes the intrinsic properties of the photonic crystal in the absence of an incident field) and the diffraction problem of a grating (finite photonic crystal) illuminated by an incident field. We point out the relationship between the translation operator of the first problem and the transfer matrix of the second. The eigenvalues of the transfer matrix contain information about the dispersion relation. This approach enables us to answer questions such as When does ultrarefraction occur? Can the photonic crystal simulate a homogeneous and isotropic material with low effective index? This approach also enables us to determine suitable parameters to obtain ultrarefractive or negative refraction properties and to design optical devices such as highly dispersive microprisms and ultrarefractive microlenses. Rigorous computations add a quantitative aspect and demonstrate the relevance of our approach. PMID- 10850472 TI - Bessel-like beams modulated by arbitrary radial functions AB - An approximate method for determining the radial and axial intensity of a Bessel like beam is presented for the general case in which a radial Bessel distribution of any order is modulated by an arbitrary function. For Bessel-Gauss, generalized Bessel-Gauss, and Bessel-super-Gauss beams, this simple approximation yields results that are very close to the exact values, while they are exact for Bessel beams. A practical beam that can be generated with a combination of simple lenses is also analyzed and illustrated. PMID- 10850473 TI - Numerical performance of finite-difference modal methods for the electromagnetic analysis of one-dimensional lamellar gratings AB - The numerical performance of a finite-difference modal method for the analysis of one-dimensional lamellar gratings in a classical mounting is studied. The method is simple and relies on first-order finite difference in the grating to solve the Maxwell differential equations. The finite-difference scheme incorporates three features that accelerate the convergence performance of the method: (1) The discrete permittivity is interpolated at the lamellar boundaries, (2) mesh points are located on the permittivity discontinuities, and (3) a nonuniform sampling with increased resolution is performed near the discontinuities. Although the performance achieved with the present method remains inferior to that achieved with up-to-date grating theories such as rigorous coupled-wave analysis with adaptive spatial resolution, it is found that the present method offers rather good performance for metallic gratings operating in the visible and near-infrared regions of the spectrum, especially for TM polarization. PMID- 10850474 TI - Waveletlike basis function approach to the propagation of paraxial beams AB - A semianalytical method is described for calculating the diffraction integral for paraxial propagation through an optical system. The field at the input plane is represented by a linear superposition of nearly Gaussian basis functions that keep a simple analytical form under ABCD propagation. The coefficients of the superposition are obtained by a numerical fit. The flexibility of the basis functions makes the method well suited to dealing with sharp local variations of the input field. PMID- 10850475 TI - Dipole radiation into grating structures AB - We present a detailed electromagnetic analysis for the radiation of an electric source located inside grating structures. Our analysis is based on the differential method and uses the scattering-matrix algorithm. We show that gratings that exhibit periodic modulations along two spatial directions (crossed gratings) enable one to couple out the totality of the light emitted by the source into the guided modes of the structure. This property is investigated through the computation of the far-field radiation patterns for crossed gratings with various etching depths. One key result is the possibility to confine the emitted light in a direction about the sample normal, a property that is of interest in the context of spontaneous emission control by microcavity structures. PMID- 10850476 TI - Marginal phase correction of truncated Bessel beams AB - Approximate analytic expressions are obtained for evaluating the axial intensity and the central-lobe diameter of J0 Bessel beams transmitted through a finite aperture phase filter. A reasonable quality factor governing the axial-intensity behavior of a phase-undistorted truncated Bessel beam is found to be the inverse square root of the Fresnel number defined, for a given aperture, from the axial point of geometrical shadow. Additional drastic reduction of axial-intensity oscillations is accomplished by using marginal phase correction of the beam instead of the well-known amplitude apodization. A procedure for analytically calculating an optimal monotonic slowly varying correction phase function is described. PMID- 10850477 TI - Green dyadic calculations for inhomogeneous optical media AB - A difference equation obtained for the reassociated Green dyadic formalism is exploited to obtain solutions for the case of a multilayered dielectric medium: Computation of limits as layers become progressively thinner leads to a parallel development for the case of a dielectric varying continuously in a single direction. A demonstration example then shows how discrete and continuous techniques can be combined into a hybrid formulation. Finally, numerical computations are presented for the simple case of a dipole, illustrating convergence of the difference equation solutions to the differential equation solution. PMID- 10850478 TI - Frame-theory-based approach for determining the time-frequency distribution characterizing a dense group of reflecting objects AB - An inverse-scattering problem concerning the determination of a time-frequency spreading function is addressed. Such a function characterizes a dense group of reflecting objects at different ranges and moving with different velocities. The problem, arising in radar and other remote-sensing techniques, is a classical inverse problem. The aim is to reconstruct a function of two variables by means of signals (of one variable) reflected from the environment being observed. The proposed approach is developed by recourse to the frame theory in order to provide a reconstruction formula that asymptotically converges to a unique spreading function. The realistic situation with respect to the transmission of a finite number of signals is further considered. In this case the reconstruction formula is shown to yield the orthogonal projection of the spreading function onto a subspace generated by the outgoing signals. PMID- 10850479 TI - Intensity-based modal decomposition of optical beams in terms of Hermite-Gaussian functions AB - We show that when an arbitrary optical beam is decomposed into a superposition of Hermite-Gaussian functions, it is sufficient to record a number of intensity profiles sampled at various transverse planes to uniquely determine the relative modal weights. This result follows from the parity relation and the nature of the Gouy phase, in addition to the orthogonality of the Fourier-transformed intensity profiles associated with the Hermite-Gaussian modes. PMID- 10850480 TI - Irradiance-variance behavior by numerical simulation for plane-wave and spherical wave optical propagation through strong turbulence AB - We have simulated optical propagation through atmospheric turbulence in which the spectrum near the inner scale follows that of Hill and Clifford [J. Opt. Soc. Am. 68, 892 (1978)] and the turbulence strength puts the propagation into the asymptotic strong-fluctuation regime. Analytic predictions for this regime have the form of power laws as a function of beta0(2), the irradiance variance predicted by weak-fluctuation (Rytov) theory, and l0, the inner scale. The simulations indeed show power laws for both spherical-wave and plane-wave initial conditions, but the power-law indices are dramatically different from the analytic predictions. Let sigmaI(2) - 1 = a(beta0(2)/betac(2))-b(l0/Rf)c, where we take the reference value of beta0(2) to be betac(2) = 60.6, because this is the center of our simulation region. For zero inner scale (for which c = 0), the analytic prediction is b = 0.4 and a = 0.17 (0.37) for a plane (spherical) wave. Our simulations for a plane wave give a = 0.234 +/- 0.007 and b = 0.50 +/- 0.07, and for a spherical wave they give a = 0.58 + /- 0.01 and b = 0.65 +/- 0.05. For finite inner scale the analytic prediction is b = 1/6, c = 7/18 and a = 0.76 (2.07) for a plane (spherical) wave. We find that to a reasonable approximation the behavior with beta0(2) and l0 indeed factorizes as predicted, and each part behaves like a power law. However, our simulations for a plane wave give a = 0.57 +/- 0.03, b = 0.33 +/- 0.03, and c = 0.45 +/- 0.06. For spherical waves we find a = 3.3 +/- 0.3, b = 0.45 +/- 0.05, and c = 0.8 +/- 0.1. PMID- 10850481 TI - New modal wave-front sensor: a theoretical analysis AB - We present a new design of a modal wave-front sensor capable of measuring directly the Zernike components of an aberrated wave front. The sensor shows good linearity for small aberration amplitudes and is particularly suitable for integration in a closed-loop adaptive system. We introduce a sensitivity matrix and show that it is sparse, and we derive conditions specifying which elements are necessarily zero. The sensor may be temporally or spatially multiplexed, the former using a reconfigurable optical element, the latter using a numerically optimized binary optical element. Different optimization schemes are discussed, and their performance is compared. PMID- 10850482 TI - Nonlocal-response diffusion model of holographic recording in photopolymer AB - The standard one-dimensional diffusion equation is extended to include nonlocal temporal and spatial medium responses. How such nonlocal effects arise in a photopolymer is discussed. It is argued that assuming rapid polymer chain growth, any nonlocal temporal response can be dealt with so that the response can be completely understood in terms of a steady-state nonlocal spatial response. The resulting nonlocal diffusion equation is then solved numerically, in low-harmonic approximation, to describe grating formation. The effects of the diffusion rate, the rate of polymerization, and a new parameter, the nonlocal response length, are examined by using the predictions of the model. By applying the two-wave coupled-wave model, assuming a linear relationship between polymerized concentration and index modulation, the resulting variation of the grating diffraction efficiency is examined. PMID- 10850483 TI - Gaussian beams in hollow metal waveguides AB - Various families of Gaussian beams have been explored previously to represent the propagation of nearly plane electromagnetic waves in media having at most quadratic transverse variations of the index of refraction and the gain or loss in the vicinity of the beam. However, such beams cannot directly represent the wave solutions for propagation in planar or rectangular waveguides, and sinusoidal mode functions are more commonly used for such waveguides. On the other hand, it is also useful to consider the possibility of recurring Gaussian beams that have an approximately Gaussian transverse profile at certain distinct planes along the propagation path. It is shown here that under some conditions recurring Gaussian beams can describe wave propagation in hollow metal waveguides, and they can also lead to efficient coupling between the waveguide fields and free-space beams. PMID- 10850484 TI - Three-dimensional finite-element beam propagation method: assessments and developments AB - In recent years the finite-element method (FEM) has been widely applied to three dimensional beam propagation analysis, and several FEM propagators have been presented. Up to now, as far as we know, an exhaustive, deep, and comparative analysis of these formulations and of the related algorithms has never been presented. We critically analyze and numerically compare, to our knowledge for the first time, different vectorial, semivectorial, and scalar formulations in order to check their performances, point out weaknesses, and suggest future developments. The results obtained highlight once more the inadequacy of scalar approaches in dealing with actual photonics devices and suggest vector formulations worthy of further development and future research. PMID- 10850485 TI - On the zero-thickness model of diffractive optical elements AB - In a recent paper [J. Opt. Soc. Am. A 16, 113 (1999)] a thin-element approximation of diffractive optical elements was used to describe diffraction of oblique incident wave fronts. This expression motivated by a ray optical analysis is shown to be incorrect. I discuss how the thin-element approximation can be generalized to arbitrary diffraction geometries. This includes an intuitive interpretation of the results. PMID- 10850486 TI - Transcriptional healing. PMID- 10850487 TI - Tripartite management of unfolded proteins in the endoplasmic reticulum. PMID- 10850488 TI - Regulatory T cells: key controllers of immunologic self-tolerance. PMID- 10850489 TI - A TRF1:BRF complex directs Drosophila RNA polymerase III transcription. AB - It has been generally accepted that the TATA binding protein (TBP) is a universal mediator of transcription by RNA polymerase I, II, and III. Here we report that the TBP-related factor TRF1 rather than TBP is responsible for RNA polymerase III transcription in Drosophila. Immunoprecipitation and in vitro transcription assays using immunodepleted extracts supplemented with recombinant proteins reveals that a TRF1:BRF complex is required to reconstitute transcription of tRNA, 5S and U6 RNA genes. In vivo, the majority of TRF1 is complexed with BRF and these two proteins colocalize at many polytene chromosome sites containing RNA pol III genes. These data suggest that in Drosophila, TRF1 rather than TBP forms a complex with BRF that plays a major role in RNA pol III transcription. PMID- 10850490 TI - Identification of human Rap1: implications for telomere evolution. AB - It has been puzzling that mammalian telomeric proteins, including TRF1, TRF2, tankyrase, and TIN2 have no recognized orthologs in budding yeast. Here, we describe a human protein, hRap1, that is an ortholog of the yeast telomeric protein, scRap1p. hRap1 has three conserved sequence motifs in common with scRap1, is located at telomeres, and affects telomere length. However, while scRap1 binds telomeric DNA directly, hRap1 is recruited to telomeres by TRF2. Extending the comparison of telomeric proteins to fission yeast, we identify S. pombe Taz1 as a TRF ortholog, indicating that TRFs are conserved at eukaryotic telomeres. The data suggest that ancestral telomeres, like those of vertebrates, contained a TRF-like protein as well as Rap1. We propose that budding yeast preserved Rap1 at telomeres but lost the TRF component, possibly concomitant with a change in the telomeric repeat sequence. PMID- 10850491 TI - ETS gene Er81 controls the formation of functional connections between group Ia sensory afferents and motor neurons. AB - The connections formed between sensory and motor neurons (MNs) play a critical role in the control of motor behavior. During development, the axons of proprioceptive sensory neurons project into the spinal cord and form both direct and indirect connections with MNs. Two ETS transcription factors, ER81 and PEA3, are expressed by developing proprioceptive neurons and MNs, raising the possibility that these genes are involved in the formation of sensory-motor connections. Er81 mutant mice exhibit a severe motor discoordination, yet the specification of MNs and induction of muscle spindles occurs normally. The motor defect in Er81 mutants results from a failure of group Ia proprioceptive afferents to form a discrete termination zone in the ventral spinal cord. As a consequence there is a dramatic reduction in the formation of direct connections between proprioceptive afferents and MNs. ER81 therefore controls a late step in the establishment of functional sensory-motor circuitry in the developing spinal cord. PMID- 10850492 TI - Transient Notch activation initiates an irreversible switch from neurogenesis to gliogenesis by neural crest stem cells. AB - The genesis of vertebrate peripheral ganglia poses the problem of how multipotent neural crest stem cells (NCSCs) can sequentially generate neurons and then glia in a local environment containing strong instructive neurogenic factors, such as BMP2. Here we show that Notch ligands, which are normally expressed on differentiating neuroblasts, can inhibit neurogenesis in NCSCs in a manner that is completely dominant to BMP2. Contrary to expectation, Notch activation did not maintain these stem cells in an uncommitted state or promote their self-renewal. Rather, even a transient activation of Notch was sufficient to cause a rapid and irreversible loss of neurogenic capacity accompanied by accelerated glial differentiation. These data suggest that Notch ligands expressed by neuroblasts may act positively to instruct a cell-heritable switch to gliogenesis in neighboring stem cells. PMID- 10850493 TI - Tribbles coordinates mitosis and morphogenesis in Drosophila by regulating string/CDC25 proteolysis. AB - Morphogenesis and cell differentiation in multicellular organisms often require accurate control of cell divisions. We show that a novel cell cycle regulator, tribbles, is critical for this control during Drosophila development. During oogenesis, the level of tribbles affects the number of germ cell divisions as well as oocyte determination. The mesoderm anlage enters mitosis prematurely in tribbles mutant embryos, leading to gastrulation defects. We show that Tribbles acts by specifically inducing degradation of the CDC25 mitotic activators String and Twine via the proteosome pathway. By regulating CDC25, Tribbles serves to coordinate entry into mitosis with morphogenesis and cell fate determination. PMID- 10850494 TI - A genetic link between morphogenesis and cell division during formation of the ventral furrow in Drosophila. AB - Stages in development with rapid transitions between mitosis and morphogenesis may require specific mechanisms to coordinate cell shape change. Here we describe a novel mitotic inhibitor that acts during Drosophila gastrulation to counteract String/Cdc25, specifically in the cells that invaginate to form the mesoderm. We have identified two genes, fruhstart and tribbles, that are required for this ventral inhibition. tribbles encodes a kinase-related protein whose RNA, however, is also present outside of the ventral region. Effective inhibition of mitosis in the cells of the ventral furrow depends on the transcription factor Snail that triggers the ventral cell shape changes. When overexpressed in a microinjection assay, Tribbles directly inhibits mitosis. We propose that Fruhstart and Tribbles form a link between the morphogenetic movements and mitotic control. PMID- 10850495 TI - Arabidopsis SGS2 and SGS3 genes are required for posttranscriptional gene silencing and natural virus resistance. AB - Posttranscriptional gene silencing (PTGS) in plants resuits from the degradation of mRNAs and shows phenomenological similarities with quelling in fungi and RNAi in animals. Here, we report the isolation of sgs2 and sgs3 Arabidopsis mutants impaired in PTGS. We establish a mechanistic link between PTGS, quelling, and RNAi since the Arabidopsis SGS2 protein is similar to an RNA-dependent RNA polymerase like N. crassa QDE-1, controlling quelling, and C. elegans EGO-1, controlling RNAi. In contrast, SGS3 shows no significant similarity with any known or putative protein, thus defining a specific step of PTGS in plants. Both sgs2 and sgs3 mutants show enhanced susceptibility to virus, definitively proving that PTGS is an antiviral defense mechanism that can also target transgene RNA for degradation. PMID- 10850496 TI - An RNA-dependent RNA polymerase gene in Arabidopsis is required for posttranscriptional gene silencing mediated by a transgene but not by a virus. AB - Posttranscriptional gene silencing is a defense mechanism in plants that is similar to quelling in fungi and RNA interference in animals. Here, we describe four genetic loci that are required for posttranscriptional gene silencing in Arabidopsis. One of these, SDE1, is a plant homolog of QDE-1 in Neurospora crassa that encodes an RNA-dependent RNA polymerase. The sde1 mutation was specific for posttranscriptional gene silencing induced by transgenes rather than by viruses. We propose that the role of SDE1 is to synthesize a double-stranded RNA initiator of posttranscriptional gene silencing. According to this idea, when a virus induces posttranscriptional gene silencing, the virus-encoded RNA polymerase would produce the double-stranded RNA and SDE1 would be redundant. PMID- 10850497 TI - The SHORT-ROOT gene controls radial patterning of the Arabidopsis root through radial signaling. AB - Asymmetric cell divisions play an important role in the establishment and propagation of the cellular pattern of plant tissues. The SHORT-ROOT (SHR) gene is required for the asymmetric cell division responsible for formation of ground tissue (endodermis and cortex) as well as specification of endodermis in the Arabidopsis root. We show that SHR encodes a putative transcription factor with homology to SCARECROW (SCR). From analyses of gene expression and cell identity in genetically stable and unstable alleles of shr, we conclude that SHR functions upstream of SCR and participates in a radial signaling pathway. Consistent with a regulatory role in radial patterning, ectopic expression of SHR results in supernumerary cell divisions and abnormal cell specification in the root meristem. PMID- 10850498 TI - Osteoporosis prevention: a pediatric challenge. PMID- 10850499 TI - Antioxidant capacity and oxygen radical diseases in the preterm newborn. AB - BACKGROUND: Bronchopulmonary dysplasia, intraventricular hemorrhage, necrotizing enterocolitis, and retinopathy of prematurity may be different manifestations of oxygen radical diseases of prematurity (ORDP). OBJECTIVE: To test the hypothesis that the antioxidant capacity of cord blood serum will predict risk of ORDP. DESIGN: An inception cohort of premature neonates was followed up from birth until discharge or death to determine if outcome was related to cord blood serum antioxidant capacity, as determined by a manual assay measuring the relative inhibition of oxidation of 2,2'-azino-di-(3-ethylbenzthiazoline)-6 sulfonic acid (ABTS). Possible correlations between antioxidant capacity and various perinatal factors were also tested. SETTING: Level 3 newborn intensive care unit. PATIENTS: All inborn very low-birth-weight neonates from whom cord blood was available and for whom maternal consent was obtained were included. Newborns who died in the first week of life or who had major congenital malformations were excluded. A convenience sample of newborns weighing more than 1500 g was used to perfect assay and explore confounders. MAIN OUTCOME MEASURES: Significant ORDP was defined as the presence of intraventricular hemorrhage greater than grade 2, retinopathy of prematurity greater than stage 1, bronchopulmonary dysplasia at the postconceptional age of 36 weeks, or necrotizing enterocolitis with the hypothesis that neonates with ORDP will have lower antioxidant capacity in cord blood serum. RESULTS: Serum antioxidant capacity at birth correlated with gestational age for the entire sample of 41 neonates and for the 26 neonates born before 32 weeks' gestation. After correction for gestational age, cord serum antioxidant capacity did not correlate with maternal smoking, preeclampsia, chorioamnionitis, cord pH Apgar scores, or any of the ORDP studied. CONCLUSION: Cord serum antioxidant capacity correlates with gestational age but does not predict ORDP risk. PMID- 10850500 TI - Parental and self-report of sleep in children with attention deficit/hyperactivity disorder. AB - OBJECTIVE: To determine the prevalence of parent-reported and self-reported sleep disturbances in a sample of school-aged children with attention-deficit/ hyperactivity disorder (ADHD). DESIGN: Cross-sectional survey questionnaire. SETTING: A multidisciplinary ADHD evaluation clinic in a children's teaching hospital (ADHD sample) and 3 elementary schools in southern New England (control sample). PARTICIPANTS: Forty-six unmedicated, school-aged children (mean age, 89.4 +/- 18.7 months; 74% male) diagnosed as having ADHD by Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, criteria who had been screened for marked symptoms of sleep-disordered breathing, and 46 normal control children (mean age, 86.5 +/- 16.9 months; 70% male). INTERVENTION: None. MAIN OUTCOME MEASURE: Sleep habits and sleep disturbances reported by parents and children. RESULTS: Children with ADHD had significantly higher (more sleep disturbed) scores on all sleep subscales of the Children's Sleep Habits Questionnaire (parent measure) than did controls; average sleep duration as reported by parents was also significantly shorter in the ADHD group. Children with ADHD also reported their own sleep to be more disturbed than controls did on the Sleep Self-report, particularly on items relating to bedtime struggles (P range, .05-.001). There was a much higher correlation between parent and child sleep report items for the children with ADHD (mean correlation, 0.55) than for the control children. CONCLUSIONS: Sleep disturbances, particularly at bedtime, are frequently reported by both parents and children with ADHD. Children undergoing evaluation for ADHD should be routinely screened for sleep disturbances, especially symptoms of sleep-disordered breathing. The causes of sleep-onset delay in children with ADHD should be considered in designing intervention strategies for children with difficulty falling and staying asleep. PMID- 10850501 TI - Antibiotic treatment of children with unsuspected meningococcal disease. AB - BACKGROUND: Data from an earlier study suggest that patients with unsuspected meningococcal disease (UMD) cannot be differentiated easily from febrile children with viral syndromes on the basis of physical examinations or peripheral blood counts. Some children with meningococcal disease therefore are treated inadvertently as outpatients. OBJECTIVE: To determine whether antibiotic therapy administered at the outpatient visit prevents complications, permanent sequelae, or death in children with UMD. METHODS: We reviewed the medical records of patients younger than 20 years with invasive meningococcal disease at 7 pediatric referral centers from January 1, 1981, through December 31, 1996. Patients were considered to have UMD if they underwent evaluation and discharge as outpatients and if blood and/or cerebrospinal fluid cultures obtained at evaluation yielded Neisseria meningitidis. We compared the frequency of development of complications (meningitis, sepsis, and pericarditis), permanent sequelae (limb amputation, skin grafting, and persistent neurologic disability) or death between patients who did and did not receive antibiotics at the outpatient visits. RESULTS: Of 58 children with UMD, 19 (33%) received antibiotics and 39 (67%) did not. Complications occurred significantly less frequently in the antibiotic-treated group (7/19 [37%] vs 27/39 [69%]; odds ratio [OR], 0.26; 95% confidence interval [CI], 0.08 0.81; P = .03). There was no significant difference in death or permanent sequelae between groups (0/19 vs 3/39 [8%]; OR, 0; 95% CI, 0-2.61; P=.54). There was insufficient power, however, to exclude the possibility of a clinically meaningful difference between the groups with regard to these latter outcomes. CONCLUSIONS: Antibiotic administration to young patients with UMD at the time of the outpatient visit is associated with a reduction in complications from this disease. Although the routine use of antibiotics in febrile outpatients younger than 20 years cannot be advocated, empirical treatment should be considered in the setting of higher probability of meningococcal disease. PMID- 10850502 TI - Comparing asthma care for Medicaid and non-Medicaid children in a health maintenance organization. AB - OBJECTIVE: To compare ambulatory visit patterns, rates of medication use, and emergency department and hospital utilization for children with asthma covered under Medicaid and commercial payers within the same health maintenance organization (HMO). DESIGN: Retrospective cohort study. SETTING: Eleven staff model pediatric departments of an HMO. PATIENTS: A total of 1928 Medicaid and 11007 non-Medicaid children aged 2 to 18 years with at least 1 encounter with a diagnosis of asthma between October 1, 1991, and September 30, 1996. METHODS: We linked patient-level data from the HMO's automated medical record system for ambulatory encounters, a claims system for emergency department and hospital care, and an automated pharmacy dispensing database. Medicaid and non-Medicaid patients were compared for all encounter types and for prescribing and dispensing of beta-agonist and controller medications (inhaled corticosteroids and cromolyn sodium). Incidence rate ratios were calculated from Poisson regression models to control for age, sex, and, when appropriate, beta-agonist dispensing rate. The number of refills authorized on each prescription and the fraction of medications dispensed as refills compared with new prescriptions were compared for Medicaid and non-Medicaid patients. RESULTS: Medicaid-insured children in the HMO were 1.4 times (95% confidence interval, 1.2-1.5) more likely to receive care in emergency departments and 1.3 times (95% confidence interval, 1.1-1.5) more likely to be hospitalized for their asthma compared with non-Medicaid members. Medicaid and non-Medicaid enrollees had similar yearly rates of nonurgent (1.32 vs 1.17) and urgent (0.38 vs 0.31) ambulatory visits. Beta-agonists were dispensed roughly equally to Medicaid and non-Medicaid members. Although Medicaid patients were less likely to have controller medications dispensed (relative risk, 0.72; 95% confidence interval, 0.69-0.74), they were equally likely to have them prescribed. CONCLUSIONS: Differences in ambulatory contact for Medicaid members do not explain the higher rates of emergency department visits and hospitalization in this population. Reasons for lower rates of dispensing of controller medications should continue to be investigated as one cause of increased morbidity for low-income children with asthma. PMID- 10850503 TI - Weight-related behaviors among adolescent girls and boys: results from a national survey. AB - OBJECTIVE: The study objectives were to assess (1) the prevalence of dieting and disordered eating among adolescents; (2) the sociodemographic, anthropometric, psychosocial, and behavioral correlates of dieting and disordered eating; and (3) whether adolescents report having discussed weight-related issues with their health care providers. DESIGN: Cross-sectional school-based survey. STUDY POPULATION: A nationally representative sample of 6728 adolescents in grades 5 to 12 who completed the Commonwealth Fund surveys of the health of adolescent girls and boys. MAIN OUTCOME MEASURES: Dieting and disordered eating (binge-purge cycling). RESULTS: Approximately 24% of the population was overweight. Almost half of the girls (45%) reported that they had at some point been on a diet, compared with 20% of the boys. Disordered eating was reported by 13% of the girls and 7% of the boys. Strong correlates of these behaviors included overweight status, low self-esteem, depression, suicidal ideation, and substance use. Almost half of the adolescents (38%-53%) reported that a health care provider had at some point discussed nutrition or weight with them. Discussions on eating disorders were reported by lower percentages of girls (24%) and boys (15%). CONCLUSIONS: The high prevalence of weight-related concerns suggests that all youth should be reached with appropriate interventions. Special attention needs to be directed toward youth at greatest risk for disordered eating behaviors, such as overweight youth, youth engaging in substance use behaviors, and youth with psychological concerns such as low self-esteem and depressive symptoms. PMID- 10850504 TI - Hospitalization for community-acquired, rotavirus-associated diarrhea: a prospective, longitudinal, population-based study during the seasonal outbreak. The Greater Toronto Area/Peel Region PRESI Study Group. Pediatric Rotavirus Epidemiology Study for Immunization. AB - OBJECTIVES: To determine the age-specific hospitalization rate for rotavirus associated diarrhea in Canadian children during the seasonal outbreak, and to characterize children and their households, for assessment of the need for a rotavirus vaccine. DESIGN: Prospective multisite cohort study. SETTINGS AND PARTICIPANTS: Children with an admission diagnosis of diarrhea admitted to 18 hospitals serving 132 study census tracts of a major urban region, from November 1, 1997, through June 30, 1998. Prospective centralized testing of stools was performed; research nurses administered a follow-up questionnaire to parents. MAIN OUTCOME MEASURE: Age-specific diarrhea and rotavirus-associated hospitalization rates. RESULTS: Of 224160 children younger than 5 years, the diarrhea hospitalization rate was 4.8 in 1000 (n = 1086) during the seasonal epidemic. Based on testing of 65% of the hospitalized children, the rotavirus associated diarrhea hospitalization rate was 1.3 in 1000; the cumulative incidence to 5 years of age was 1 in 160. Rotavirus-associated diarrhea was reported in 37% of the 1001 hospitalized children undergoing testing inside and outside of the census tracts; in children aged 6 to 35 months, this rose to more than 70% during April and May. Ages of children with rotavirus-associated diarrhea were 0 to 2 months (2%), 3 to 5 months (5%), 6 to 23 months (60%), 24 to 35 months (15%), and 36 months or older (19%). Of children aged 0 to 5 and 6 to 11 months, 4 (19%) of 21 and 6 (10%) of 59, respectively, had been born prematurely; 20 (24%) of 83 younger than 1 year were breastfed at the time of illness. Of children younger than 36 months, 77% were cared for in their homes; 13%, in family day care homes; and 8%, in child care centers. The mean (+/- SD) duration of rotavirus hospitalization based on hospital records and parental questioning was 2.4 +/- 1.7 and 3.1 +/- 1.6 days, respectively; it was significantly longer (P < or = .001) in children with an underlying medical condition. One child required intensive care unit hospitalization. Diarrhea occurred concurrently in 74% of household contacts younger than 3 years; 38%, aged 3 to 18 years; and 29%, older than 18 years. Seventy-six percent of parents were married. Household incomes in Canadian dollars in the 81% reporting were less than $20000 in 20%, $20000 to $60000 in 44%, and greater than $60000 in 36%. Ethnicity was reported as 53% white, 15% black, 10% Asian, 12% East Indian, and 11% other. CONCLUSIONS: Based on testing of 65% of children with diarrhea, rotavirus resulted in hospitalization in a minimum of 1 in 160 children by 5 years of age during the seasonal outbreak. Had 100% of young children with diarrhea undergone testing, the extrapolated cumulative incidence of rotavirus associated diarrhea by 5 years of age may have been 1 in 106. PMID- 10850505 TI - Rotavirus-associated diarrhea in outpatient settings and child care centers. The Greater Toronto Area/Peel Region PRESI Study Group. Pediatric Rotavirus Epidemiology Study for Immunization. AB - OBJECTIVES: To determine the prevalence of rotavirus infection in outpatient and child care center (CCC) settings during the seasonal rotavirus outbreak and to describe associated health care utilization. DESIGN: Prospective, multisite cohort study in various ambulatory settings. SETTINGS AND PARTICIPANTS: Participants were children with diarrhea (1) presenting to hospital emergency departments (EDs) and receiving intravenous (IV; n = 8) or oral (n = 1) hydration, (2) seen in pediatric practices (n=4), or (3) attending CCCs (n = 19) between November 1, 1997, and June 30, 1998. Prospective centralized testing of stool samples for rotavirus was performed using enzyme-linked immunosorbent assay and electron microscopy. Study nurses administered follow-up parent questionnaires for rotavirus-positive children. MAIN OUTCOME MEASURE: Prevalence of rotavirus-associated diarrhea. RESULTS: During the 8-month study, rotavirus was identified in 92 children with diarrhea: ED-IV, 20 (44%) of 45; ED-oral, 9 (47%) of 19; pediatric practices, 30 (20%) of 147; and CCCs, 33 (18%) of 186. Of 226 children with diarrhea in pediatric practices, all 5 who progressed to ED-IV hydration or hospitalization were tested, and 3 (60%) were rotavirus positive. Of 211 children in CCCs with diarrhea, 84% who required no health care visits were tested, and of these 10% were positive; of 56 who went on to require a health care visit and 8 who required ED-IV hydration or hospitalization, all were tested, and 27% and 75%, respectively, were rotavirus positive. Among 16 children with rotavirus followed up with ED-IV hydration, 4 (25%) returned and were hospitalized. Maximal health care intervention among 29 children with rotavirus enrolled in pediatric practices included 22 (76%) seeing the pediatrician only, 5 (17%) seeking further care in the ED, 1 (3%) receiving further ED-IV hydration, and 1 (3%) being hospitalized briefly. Maximal health care intervention for 33 children with rotavirus enrolled in CCCs included 13 (39%) who did not visit a physician, 11 (33%) who did, 3 (9%) who sought care in the ED, 1 (3%) who received ED-IV hydration, and 5 (15%) who were hospitalized. In CCCs, rates of diarrhea per 100 child-months of observation were as follows: ages 0 to 23 months, 6.6 episodes; ages 24 to 35 months, 1.9 episodes; and 3 years and older, 0.07 episodes; rates of rotavirus-associated diarrhea were as follows: ages 0 to 23 months, 1.1 episodes (28 of 2547); ages 24 to 35 months, 0.23 episodes (5 of 2185); and 3 years and older, 0 episodes (0 of 4124). CONCLUSION: Across a variety of outpatient and CCC settings, rotavirus is an important cause of diarrhea and a major cause of health care utilization. PMID- 10850506 TI - Children with febrile seizures do not consume excess health care resources. AB - BACKGROUND: Febrile seizures are benign but so terrifying for parents that they may subsequently view their affected children as "vulnerable". Children viewed as vulnerable may be brought to medical attention more frequently. We examined subsequent hospitalizations and physician visits during a 6- to 7 1/2-year period for a group of children who had participated in a case-control study of initial febrile seizures. METHODS: Individual data from a regional cohort of 75 children with a first febrile seizure and 150 febrile and 150 afebrile controls were linked to 2 comprehensive provincial health services databases-a hospital admissions/ separations database and a physician services database. RESULTS: Linkage was achieved for 98% of the study cohort, with heath care utilization data for 6 to 7 1/2 years available for 96%. Children with febrile seizures had nearly identical rates of subsequent hospitalization compared with age-matched controls (chi2 test, P = .88). An excess of day-surgery visits for primarily otolaryngologic procedures was seen for the febrile seizure patients 0 to 12 months after their initial febrile seizure (chi2 test, P < .001). During the next 6 to 7 1/2 years, the febrile seizure patients had nearly identical rates of physician visits (chi2 test, P = .15); however, they had more visits to otolaryngologists in the first 3 to 9 months after the febrile seizure (chi2 test, P < .001), but fewer visits to pediatricians during the next 1 to 4 years (chi2 test, P < .001). CONCLUSIONS: Children with febrile seizures have nearly identical rates of hospital and physician services utilization compared with controls. This supports the hypothesis that febrile seizures are benign, and that parents recover from their initial anxiety and do not consider their children vulnerable to additional illness in the years that follow. PMID- 10850507 TI - Neurodevelopmental follow-up at 36 months' corrected age of preterm infants treated with prophylactic indomethacin. AB - BACKGROUND: Previous reports have suggested that prophylactic indomethacin decreases cerebral blood flow and may play a role in the development of ischemic brain injury and developmental handicaps. OBJECTIVE: To assess the neurodevelopmental outcome of subjects at 36 months' corrected age (CA) who, as low-birth-weight infants, received prophylactic low-dose indomethacin within the first 24 hours of life to prevent patent ductus arteriosus. SETTING: Newborn intensive care nursery and outpatient follow-up clinic at Children's Hospitals and Clinics of Minneapolis, Minneapolis, Minn. DESIGN: Ninety infants with birth weights of 600 to 1250 g were entered into a prospective, randomized, controlled trial to receive either prophylactic indomethacin, 0.1 mg/kg, or placebo in the first 24 hours and again every 24 hours for 6 doses to prevent patent ductus arteriosus. Nonresponders were treated with standard therapeutic indomethacin or ligation. Neurodevelopmental assessment at approximately 36 months' CA included medical and developmental histories, physical examinations, and developmental testing using the Bayley II Scales of Infant Development on subjects up to 42 months' CA. Subjects were classified as (1) normal, (2) mildly to moderately abnormal, or (3) severely impaired. RESULTS: Forty-two (98%) of 43 subjects who received prophylactic indomethacin survived compared with 46 (98%) of 47 who received placebo. Sixty-six (75%) of 88 survivors were seen for neurodevelopmental assessment at 36 months' CA. This group included 29 (69%) of 42 who received prophylactic indomethacin and 37 (80%) of 46 who received placebo. Twenty-three (79%) of 29 infants in the prophylactic indomethacin group had normal neurodevelopmental assessments at 36 months' CA compared with 26 (70%) of 37 placebo-treated subjects (P = .68). Of 4 significantly impaired subjects treated with prophylactic indomethacin, 1 had spastic diplegia; 1, spastic quadriplegia; 1, cognitive delay; and 1, significant motor delay. Of 8 significantly impaired placebo-treated subjects, 7 had spastic diplegia; 1, microcephaly. CONCLUSION: The use of prophylactic low-dose indomethacin when initiated in the first 24 hours of life in low-birth-weight infants to prevent patent ductus arteriosus is not associated with adverse neurodevelopmental outcome at 36 months' CA. PMID- 10850508 TI - Pyogenic granuloma presenting as a congenital epulis. AB - OBJECTIVE: To describe a clinical approach to the differential diagnosis of oral lesions in neonates. DESIGN: Case report. SETTING: Academic ambulatory care center. PARTICIPANTS: Male infant. RESULTS: A gingival mass in a male infant appeared clinically consistent with a congenital epulis. Following excision and histologic examination, the diagnosis was determined to be a pyogenic granuloma. Careful attention to alternative diagnoses led to the correct etiology. CONCLUSIONS: Primary care pediatricians encounter neonatal oral lesions infrequently. The most common oral lesions in the newborn period are Epstein pearls and Bohn nodules. This case illustrates the importance of formulating a more extensive differential diagnosis on discovery of a neonatal oral mass. PMID- 10850509 TI - Anticipatory guidance about child safety seat misuse: lessons from safety seat "checkups". AB - OBJECTIVES: To quantify the frequency of improper child safety seat use and to identify the most common mistakes in safety seat use, so that priorities for anticipatory guidance about misuse can be identified. DESIGN: Descriptive survey of types and frequency of safety seat misuse. SETTING: Eleven safety seat "checkups" sponsored by the Louisiana SAFE KIDS Coalition in southeastern Louisiana in 1998. PARTICIPANTS: Convenience sample of parents recruited for checkups through local media and sponsoring businesses. Three hundred seventeen child safety seats were checked. RESULTS: Of the 266 forward- and rear-facing seats checked, 250 (94%) were installed incorrectly. Sixty-one (23%) of the seats had minor misuse or were correctly used, 107 (40%) were partially misused, and 98 (37%) were extensively misused. The 3 most frequently found problems were seat not belted into vehicle tightly (142 [88%] of forward-facing seats and 84 [81%] of rear-facing seats), safety seat harness straps not snug (70 [43%] of forward facing seats and 49 [47%] of rear-facing seats), and harness retainer clip not at armpit level (55 [34%] of forward-facing seats and 38 [37%] of rear-facing seats). CONCLUSIONS: As part of the routine anticipatory guidance offered during well-child visits, health care providers (ie, physician, nurse, or nurse practitioner) should counsel parents specifically about these 3 frequent errors in child safety seat use. PMID- 10850510 TI - Teenaged girls, carbonated beverage consumption, and bone fractures. AB - OBJECTIVE: To determine the possible association between carbonated beverage consumption and bone fractures among teenaged girls given the awareness of the concern about the impact of carbonated beverage consumption on children's health. SETTING: An urban high school. METHODS: A cross-sectional (retrospective) study. Four hundred sixty 9th- and 10th-grade girls attending the high school participated in this study by completing a self-administered questionnaire relating to their physical activities and personal and behavioral practices. The school system and the Harvard School of Public Health Institutional Review Boards approved the study. The girls' self-reports on physical activity, carbonated beverage consumption, and bone fractures are analyzed. RESULTS: In the total sample, carbonated beverage consumption and bone fractures are associated: odds ratio = 3.14 (95% confidence limit, 1.45, 6.78), P = .004. Among physically active girls, the cola beverages, in particular, are highly associated with bone fractures: odds ratio = 4.94 (95% confidence limit, 1.79, 13.62), P = .002. CONCLUSIONS: The results reported confirm previous findings, but the mechanism by which cola drinks are associated with bone fractures in physically active girls has neither been fully explored nor determined. Nevertheless, national concern and alarm about the health impact of carbonated beverage consumption on teenaged girls is supported by the findings of this study. The results have policy implications for improving the dietary practices and health of children. PMID- 10850511 TI - Itraconazole pulse therapy for dermatophyte onychomycosis in children. AB - BACKGROUND: Onychomycosis, or fungal infection of the nail, can occur in prepubertal children. However, its diagnosis is often missed or the condition is inappropriately treated with topical medication. Griseofulvin has been the therapy of choice, but even long-term treatment is associated with a poor cure rate and high rate of relapse. Trials with adult patients have shown that itraconazole pulse therapy for onychomycosis requires a shorter duration of total therapy than griseofulvin treatment and is rarely associated with adverse reactions, suggesting that it may be the treatment of choice for pediatric patients with onychomycosis. DESIGN: We retrospectively reviewed the courses of prepubertal patients with dermatophyte onychomycosis who initiated treatment with itraconazole pulse therapy between January 1995 and June 1998. SETTING: Urban and suburban pediatric dermatology clinics of a children's hospital. PATIENTS: Seventeen prepubertal patients met the enrollment and follow-up criteria. These included fungal infection of the nail(s), documented by fungal culture and/or positive potassium hydroxide mounts of nail scrapings; at least 1 follow-up visit; and contact by telephone or clinic visit within 2 months prior to compilation of data. In 59% of patients, a relative living at the home had onychomycosis at the time of diagnosis. INTERVENTION: Patients were treated with daily to twice-daily pulses of itraconazole, administered for 1 week of each of 3 to 5 months. MAIN OUTCOME MEASURES: Clinical cure after itraconazole therapy in patients with documented onychomycosis and clinical and mycologic relapse after initial cure. Fungal cultures were not repeated if clinical cure was noted. RESULTS: All but 1 patient responded fully to therapy, showing improvement within a few months and subsequently clearance (94% clinical cure rate). No patients experienced any clinical adverse reactions. No relapses occurred after clinical cure during a follow-up period of 1 to 4.25 years after initiation of therapy. CONCLUSIONS: Itraconazole pulse therapy is effective and safe for the treatment of onychomycosis in children. The relapse rate in pediatric patients is lower than in adults, although the high frequency of onychomycosis in non-pediatric family members suggests that the recurrence risk is increased if other family members are not treated concomitantly. PMID- 10850512 TI - Combined analgesia and local anesthesia to minimize pain during circumcision. AB - BACKGROUND: Pain of circumcision is only partially relieved by single modalities, such as penile nerve block, lidocaine-prilocaine cream, and sucrose pacifiers. OBJECTIVE: To assess the effectiveness of a combination of interventions on the pain response of infants undergoing circumcision. METHODS: Cohort study. Group 1 included infants circumcised using the Mogen clamp and combined analgesics (lidocaine dorsal penile nerve block, lidocaine-prilocaine, acetaminophen, and sugar-coated gauze dipped in grape juice). Group 2 included infants circumcised using the Gomco clamp and lidocaine-prilocaine. Infants were videotaped during circumcision, and pain was assessed using facial activity scores and percentage of time spent crying. RESULTS: There were 57 infants in group 1 and 29 infants in group 2. Birth characteristics did not differ between groups. Infants in group 1 were older than infants in group 2 (17 days vs 2 days) (P < .001). The mean duration of the procedure was 55 seconds and 577 seconds for infants in group 1 and 2, respectively (P < .001). Facial action scores and percentage of time spent crying were significantly lower during circumcision for infants in group 1 (P < .001). The percentage of time spent crying was 18% and 40% for infants in groups 1 and 2, respectively. No adverse effects were observed in infants in group 1; 1 infant in group 2 had a local skin infection. CONCLUSIONS: Infants circumcised with the Mogen clamp and combined analgesia have substantially less pain than those circumcised with the Gomco clamp and lidocaine-prilocaine cream. Because of the immense pain during circumcision, combined local anesthesia and analgesia using the Mogen clamp should be considered. PMID- 10850513 TI - High prevalence of overweight children and adolescents in the Practice Partner Research Network. AB - OBJECTIVES: To determine the prevalences of overweight children and adolescents seeking care in 49 Practice Partner Research Network (PPRNet) primary care practices and to compare these rates with national population-based surveys. DESIGN AND SETTING: The prevalence of overweight subjects (>95th percentile for age and sex) and subjects at risk for being overweight (>85th percentile for age and sex) was calculated for 30445 children aged 6 through 19 years visiting PPRNet primary care practices from 1995 through 1997. Prevalences were compared with prevalences from the National Health and Nutrition Examination Surveys. Percentile cutoffs from the National Health Evaluation Survey were used as the baseline standard for the comparisons. MAIN OUTCOME MEASURE: Obesity prevalences. RESULTS: Thirty-six percent of boys aged 6 through 11 years and 35% of boys aged 12 through 17 years were either at risk for being overweight or overweight; 20% and 19% were overweight, respectively. Thirty-five percent of girls aged 6 through 11 years and 34% of girls aged 12 through 17 years were either at risk for being overweight or were overweight; 20% and 18% were overweight, respectively. Prevalences of overweight subjects and subjects at risk for being overweight were much greater in patients of PPRNet primary care practices compared with the most recent national survey, the National Health and Nutrition Examination Surveys III. CONCLUSIONS: One in 3 children and adolescents visiting PPRNet primary care practices is at risk for being overweight or is overweight. The prevalence of obesity in children and adolescents visiting primary care practices is much greater than that observed in national population-based surveys. PMID- 10850514 TI - Radiological case of the month. Typhlitis with concurrent appendicitis. PMID- 10850515 TI - Picture of the month. Costello syndrome. PMID- 10850516 TI - Pathological case of the month. Congenital cystic adenomatoid malformation of the lung. PMID- 10850517 TI - Unanticipated reactions to a recent report on alcohol problems among pediatric residents. PMID- 10850518 TI - What about sleep? PMID- 10850519 TI - Who is medicating very young children with attention-deficit/hyperactivity disorder? PMID- 10850520 TI - The case for not omitting evaluation of culture findings for diagnosing tinea capitis. PMID- 10850521 TI - A dose-escalation study of the safety, tolerability, and pharmacokinetics of intravenous gatifloxacin in healthy adult men. AB - STUDY OBJECTIVES: To examine single- and multiple-dose safety, tolerability, and pharmacokinetics of gatifloxacin administered as daily 1-hour intravenous infusions for 14 days, and to determine the effect of gatifloxacin on glucose tolerance, pancreatic beta-cell function, and electrocardiogram (ECG). DESIGN: Randomized, double-blind, placebo-controlled, ascending-dose study. SETTING: Bristol-Myers Squibb, Clinical Pharmacology Unit, Princeton, New Jersey, USA. PATIENTS: Forty healthy male subjects, eight in each of five groups, were enrolled to receive sequential doses of gatifloxacin: 200 mg (10 mg/ml), 200 mg (1 mg/ml), and 400, 600, and 800 mg (2 mg/ml); six subjects per group received active drug and two received placebo. INTERVENTIONS: A single dose of the drug was administered as an intravenous infusion over 1 hour. After a 72-hour washout period, the drug was administered once/day for 14 days by 1-hour intravenous infusion. Physical examinations, ECGs, spirometry, and clinical laboratory tests, including glucose tolerance test (GTT) and assessment of glucose homeostasis, were performed before treatment and on selected dosing days. A safety evaluation was performed before escalating doses. No intrasubject dose escalation was permitted. MEASUREMENTS AND MAIN RESULTS: The pharmacokinetics of gatifloxacin were dose linear and time independent after intravenous administration over the range of 200-800 mg. After daily repeated administration, a predictable, modest accumulation was observed; steady state was reached by the third dose. Approximately 80% of the dose was recovered as unchanged drug in urine. Mean changes (before the first dose to the last dose) after oral GTT and in fasting serum glucose, insulin, and C-peptide concentrations were comparable among the gatifloxacin and placebo treatment groups. A mild, transient decrease in serum glucose was associated with the end of the 1-hour infusion of gatifloxacin. No clinically important changes in QTc interval or spirometry occurred. The most frequent treatment-related adverse effects were local intravenous site reactions, which were associated with dose and/or concentration of intravenous solution. CONCLUSION: Gatifloxacin was safe and well tolerated at intravenous doses of up to 800 mg/day for 14 days. Gatifloxacin pharmacokinetics were linear and time independent. PMID- 10850522 TI - Interchangeability of 400-mg intravenous and oral gatifloxacin in healthy adults. AB - STUDY OBJECTIVE: To evaluate the interchangeability of 400-mg intravenous and oral doses of gatifloxacin. DESIGN: Randomized, open-label, crossover study. SETTING: GFI Pharmaceutical Services, Inc., Evansville, Indiana, USA. SUBJECTS: Twenty-four healthy men and women (12 of each gender), aged 18-42 years. INTERVENTIONS: Subjects received single doses of gatifloxacin 400 mg either by intravenous infusion over 1 hour or a 400-mg tablet orally with 240 ml of water, each dose separated by a 1-week washout. Plasma concentrations of gatifloxacin were determined by a validated high-performance liquid chromatography; pharmacokinetic parameters were calculated using noncompartmental methods. Distributions of pharmacokinetic parameter values were summarized by route of administration and gender. Effects of treatment on pharmacokinetic parameter values of gatifloxacin were assessed by an analysis of variance model suitable for a two-way, two-treatment, crossover design. Clinical evaluations were performed to assess drug safety and tolerability. MEASUREMENTS AND MAIN RESULTS: Intravenous and oral gatifloxacin were considered interchangeable because both routes were bioequivalent with respect to area under the curve (AUC; 90% confidence interval for the ratio of geometric means contained within 0.8-1.25). The plasma concentration-time profile after intravenous administration was similar and comparable in extent of exposure (AUC0-infinity) with that for the oral route when equal doses were administered to men and women. The absolute bioavailability of gatifloxacin after oral administration was 96%, consistent with bioequivalence of the 400-mg intravenous and oral doses. The drug was well tolerated; the frequency of adverse events was comparable after intravenous and oral administration. CONCLUSION: Intravenous and tablet formulations of gatifloxacin are bioequivalent and therefore interchangeable. This permits greater flexibility in choosing oral or parenteral therapy, with the possibility of avoiding hospitalization based on knowledge that oral administration will deliver therapeutic exposure to the drug, or abbreviating hospital stay due to ease of switching from intravenous to oral therapy. PMID- 10850523 TI - Age and gender effects on the pharmacokinetics of gatifloxacin. AB - STUDY OBJECTIVE: To compare the pharmacokinetics and safety of gatifloxacin in elderly (> or = 65 yrs) and young (18-45 yrs) men and women. DESIGN: Open-label, parallel-group, single-dose study. SETTING: GFI Pharmaceutical Services Inc., Evansville, Indiana, USA. SUBJECTS: Forty-eight healthy subjects in four groups of 12 each. INTERVENTIONS: Subjects received single oral doses of gatifloxacin 400 mg. Serial blood and urine samples were collected for 96 hours after dosing to determine drug concentrations. MEASUREMENTS AND MAIN RESULTS: Age and gender had moderate effects on the pharmacokinetics of gatifloxacin. Elderly women had a 21% higher geometric mean peak plasma concentration (Cmax) and a 32% higher area under the plasma concentration-time curve (AUC0-infinity) than young women. Adjustment for creatinine clearance had only a slight effect on Cmax but reduced the estimated effect of age on AUC0-infinity in women from a 32% increase to a 15% increase. Gender effects on pharmacokinetic values were noted among elderly subjects only. Geometric means for Cmax and AUC0-infinity were 21% and 33% higher, respectively, for elderly women and elderly men. Adjustment for body weight reduced these differences to 11% and 20%, respectively. CONCLUSION: The effects of age on gatifloxacin pharmacokinetic values were largely attributed to declining renal function, whereas those of gender were largely attributed to differences in body weight. These modest age- and gender-related differences do not warrant dosage adjustment. PMID- 10850524 TI - Effect of multiple-dose gatifloxacin or ciprofloxacin on glucose homeostasis and insulin production in patients with noninsulin-dependent diabetes mellitus maintained with diet and exercise. AB - STUDY OBJECTIVES: To compare the effects of gatifloxacin and ciprofloxacin on glucose homeostasis, including glucose tolerance test (GTT), pancreatic beta-cell function, and insulin production and secretion in patients with noninsulin dependent (type 2) diabetes mellitus (NIDDM) maintained with diet and exercise; and to evaluate the pharmacokinetics, safety, and tolerability of gatifloxacin. DESIGN: Randomized, double-blind, placebo-controlled, multiple-dose study. SETTING: GFI Pharmaceutical Services, Inc., Evansville, Indiana; Chicago Center for Clinical Research, Chicago, Illinois; and New Orleans Center for Clinical Research, New Orleans, Louisiana, USA. PATIENTS: Forty-eight men and women with NIDDM. INTERVENTIONS: Patients were assigned sequentially at enrollment to receive gatifloxacin 400 mg/day orally, ciprofloxacin 500 mg twice/day orally, or placebo for 10 days. Oral GTTs were performed on specific days throughout the study, as well as measurements of serum glucose, serum insulin, and C-peptide levels. Physical examinations, electrocardiograms, spirometry, and clinical laboratory tests were performed before dosing and on selected dosing days. MEASUREMENTS AND MAIN RESULTS: Gatifloxacin had no significant effect on glucose tolerance and pancreatic beta-cell function, as shown by oral GTT results and insulin and C-peptide levels. Fasting glucose levels 0-6 hours after gatifloxacin administration on days 1 and 10 showed a downward trend, but it was not significant compared with placebo; results were similar with ciprofloxacin. Gatifloxacin also lacked a long-term effect on fasting insulin levels, but this was not shown for a short-term effect, suggesting a modest, transient effect on insulin release. On the other hand, ciprofloxacin had no short-term effect but produced a more sustained effect on insulin release and production. The pharmacokinetics of gatifloxacin in patients with NIDDM were similar to those in healthy subjects. Overall, subjects tolerated treatment well. All reported drug related adverse events were mild to moderate in intensity. The frequency of adverse events was similar in gatifloxacin- and ciprofloxacin-treated patients, and only slightly higher than in placebo-treated patients. CONCLUSION: Gatifloxacin was well tolerated in patients with NIDDM controlled by diet and exercise. It had no significant effect on glucose homeostasis, beta-cell function, or long-term fasting serum glucose levels, but it did cause a brief increase in serum insulin levels. PMID- 10850525 TI - Safety and pharmacokinetics of a single oral dose of gatifloxacin in patients with moderate to severe hepatic impairment. AB - STUDY OBJECTIVES: To assess the safety and pharmacokinetics of oral gatifloxacin 400 mg in subjects with and without hepatic impairment, and the need to modify doses in patients with hepatic dysfunction. DESIGN: Single-dose, nonrandomized, open-label, parallel-group study. SETTING: Clinical Research Center, New Orleans, Louisiana. PATIENTS: Eight subjects with grade B or C hepatic dysfunction (Child Pugh classification) and eight age-, weight-, and gender-matched subjects with normal hepatic function. INTERVENTIONS: After a single oral dose of gatifloxacin 400 mg, blood and urine samples were collected at specified times or intervals over 48 hours to determine drug concentrations. MEASUREMENTS AND MAIN RESULTS: All 16 subjects (7 with grade B and 1 with grade C hepatic impairment, 8 with normal hepatic function) completed the study. Peak plasma concentration (Cmax) and area under the plasma concentration-time curve (AUC0-infinity) for gatifloxacin were 32% and 22% higher, respectively, in subjects with hepatic impairment. Except for Cmax, the ratio of means for AUC satisfied the specified criterion (0.67-1.50) for lack of effect. There were no statistically significant differences in any other pharmacokinetic values except apparent oral clearance (ClT/F). All treatment-emergent adverse events were mild or moderate in intensity and resolved before subjects were discharged from the study. CONCLUSION: Modest increases in Cmax and AUC0-infinity are not anticipated to have a negative effect on the outcome of therapy in hepatically impaired subjects, nor are they anticipated to result in adverse drug reactions. Patients with moderate to severe (Child-Pugh grade B or C) hepatic dysfunction do not require gatifloxacin dose adjustments. In addition, the similarity in half-life (t1/2) for the groups (8.9 hrs for hepatically impaired subjects, 9.3 hrs for controls) suggests that no difference would be anticipated in the extent of drug accumulation after multiple doses. The overall safety and tolerability of a single oral dose of gatifloxacin 400 mg were excellent in both healthy subjects and those with hepatic impairment. PMID- 10850526 TI - Natural pigments: carotenoids, anthocyanins, and betalains--characteristics, biosynthesis, processing, and stability. AB - Pigments are present in all living matter and provide attractive colors and play basic roles in the development of organisms. Human beings, like most animals, come in contact with their surroundings through color, and things can or cannot be acceptable based on their color characteristics. This review presents the basic information about pigments focusing attention on the natural ones; it emphasizes the principal plant pigments: carotenoids, anthocyanins, and betalains. Special considerations are given to their salient characteristics; to their biosynthesis, taking into account the biochemical and molecular biology information generated in their elucidation; and to the processing and stability properties of these compounds as food colorants. PMID- 10850527 TI - A mechanistic analysis of reduced mechanical performance in human heart failure. AB - In failing human hearts (FHH) (NYHA IV) the cardiac output is inadequate to meet the metabolic needs of the peripheral systems. By means of thermo-mechanical analysis we have shown that epicardial strips from FHH (37 degrees C) have a depressed tension independent heat (TIH) and tension independent heat rate (dTIH / dt) liberation that correlates with depression in peak isometric force and the rate of relaxation. Furthermore, in response to a change in frequency of stimulation, FHH shows a severe blunting of the force-frequency relationship resulting in a decrease in myocardial reserve and in the frequency at which optimum force is obtained. We used ventricular ANF as an index of the severity of myocardial disease and demonstrated an inverse relationship between ANF mRNA and the sarcoplasmic reticulum (SR) calcium cycling proteins (SERCA 2, Phospholamban, Ryanodine Receptor) while these latter proteins all had a positive correlation with each other. At the same time there was an increase in sarcolemmal sodium calcium exchange protein. The decrease in SR pump proteins correlates with the decrease in myocardial reserve and optimum frequency of contraction. The latter mechanical changes are explainable in terms of a frequency dependent decrease in calcium concentration (aequorin light) in FHH. PMID- 10850528 TI - Molecular mechanism of mechanical stress-induced cardiac hypertrophy. AB - Mechanical stress is a major cause of cardiac hypertrophy. Although the mechanisms by which mechanical load induces cardiomyocyte hypertrophy have long been a subject of great interest for cardiologists, the lack of a good in vitro system has hampered the understanding of the biochemical mechanisms. For these past several years, however, an in vitro neonatal cardiocyte culture system has made it possible to examine the biochemical basis for the signal transduction of mechanical stress. Passive stretch of cardiac myocytes cultured on silicone membranes activates phosphorylation cascades of many protein kinases including protein kinase C, Raf-1 kinase and extracellular signal regulated kinases, and induces the expression of specific genes as well as an increase in protein synthesis. During that process, the secretion and production of vasoactive peptides such as angiotensin II and endothelin, are increased and they play critical roles in the induction of these hypertrophic responses. Although the involvement of vasoactive peptides in the development of cardiac hypertrophy is clinically important, the "mechanoreceptor" which receives the mechanical stress and converts it into intracellular biochemical signals remained unknown. We have recently obtained evidence suggesting that ion channels and integrins may be the "mechanoreceptor", the activation of which leads to cardiac hypertrophy. PMID- 10850529 TI - Intracoronary serum smooth muscle myosin heavy chain levels following PTCA may predict restenosis. AB - Recently a novel biochemical method that uses an immunoassay to quantitate serum smooth muscle myosin heavy chain (SMMHC) levels was developed for diagnosis of aortic dissection.) The purpose of this study was to determine whether SMMHC released from the coronary arterial wall can be used to predict restenosis after percutaneous transluminal coronary angioplasty (PTCA). Fifty-two consecutive patients undergoing successful PTCA for single vessel disease were examined (40 men, 12 women, 63 +/- 8 years). Intracoronary blood samples were obtained distal to the lesion, and from the femoral artery after PTCA. In 10 patients, blood samples were taken immediately after the final balloon inflation, and 10 and 20 minutes after PTCA. SMMHC levels were measured by ELISA using SMMHC-specific monoclonal antibodies. Follow-up coronary angiography was performed 3 months after PTCA. Intracoronary serum SMMHC levels were significantly higher than those obtained from the femoral artery (10.6 +/- 1.5 vs 2.1 +/- 0.1 ng / ml, p < or = 0.001). Of 40 patients without apparent dissection, the 23 patients who did not develop restenosis in the follow-up study were found to have had higher levels of intracoronary SMMHC levels immediately after PTCA compared to the 17 patients with restenosis (15.2 +/- 2.9 vs 7.1 +/- 1.2 ng /ml, p < or = 0.05). We suggest that elevated intracoronary SMMHC levels after PTCA may reflect the extent of injury to the arterial wall. Intracoronary SMMHC may be a possible biochemical marker for the prediction of restenosis. PMID- 10850530 TI - Usefulness of pulse-wave Doppler tissue sampling and dobutamine stress echocardiography for identification of false positive inferior wall defects in SPECT. AB - False positive inferior wall perfusion defects restrict the accuracy of SPECT in diagnosis of coronary artery disease (CAD). Pulse-Wave Tissue Doppler (PWTD) has been recently proposed to assess regional wall motion velocities. The objectives of this study were to evaluate the presence of CAD by using PWTD during dobutamine stress echocardiography (DSE) in patients with an inferior perfusion defect detected by SPECT and compare PWTD parameters of normal cases with patients who had inferior perfusion defect and CAD. Sixty-five patients (mean age 58 +/- 8 years, 30 men) with a normal LV systolic function at rest according to echocardiographic evaluation with an inferior ischemia determined by SPECT and a control group (CG) of 34 normal cases (mean age 56 +/- 7 years, 16 men) were included in this study. All patients underwent a standard DSE (up to 40 microg / kg / min with additional atropine during sub-maximum heart rate responses). Pulse wave Doppler tissue sampling of inferior wall was performed in the apical 2 chamber view at rest and stress. The coronary angiography was performed within 24 hours. The results were evaluated for the prediction of significant right coronary artery (RCA) and / or left circumflex coronary artery (CX) with narrowing (> or = 50% diameter stenosis, assessed by quantitative coronary angiography). It was observed that the peak stress mean E / A ratio was lower in patients with CAD when compared to patients without CAD (0.78 +/- 0.2 versus 1.29 +/- 0.11 p < 0.0001). Also the peak stress E / A ratio of normal cases was significantly higher than patients who had CAD (1.19 +/- 0.3 versus 0.78 +/- 0.2 p < 0.0001). When the cut off point for the E / A ratio was determined as 1, the sensitivity and specificity of dobutamine stress PWTD E / A were 89% and 86 %, respectively. The peak stress E / A ratio was higher than 1 in all patients with a false positive perfusion defect. Systolic S velocity increase during DSE was significantly lower in patients with CAD (54 % +/- 17 versus 99 % +/- 24 p = 0.01). The analysis of S velocity increase yielded 81% sensitivity and 76 % specificity for prediction of CAD when a 70 % increase was accepted as a cut-off value. Pulse-wave Doppler tissue sampling during DSE may help to identify false positive inferior wall defects detected by SPECT. PMID- 10850531 TI - Magnetic resonance coronary angiography in patients with ischemic heart disease: analysis of coronary arterial blood flow velocity pattern. AB - Only a few reports evaluating coronary arterial blood flow velocity patterns using magnetic resonance (MR) coronary angiography have appeared to date. This study reports an evaluation of coronary arterial blood flow velocity patterns in patients with ischemic heart disease and in healthy subjects using MR coronary angiography. The subjects consisted of 20 patients with ischemic heart disease (IHD group) and 20 normal healthy subjects (N group). Using the fCARD PC method, ECG-gated MR coronary angiography was performed using an anteroposterior opposing phased array coil. Regions of interest were placed on bilateral coronary arteries to measure coronary arterial blood flow velocity patterns. The IHD group was divided into two subgroups, based on the presence (MI group) or absence (AP group) of infarcted myocardium using 99m Tc-methoxyisobutylisonitrile (MIBI) myocardial scintigraphy. Average diastolic peak velocity (ADPV) was lower in the IHD group than in the N group. In addition, the diastolic / systolic velocity ratio (DSVR) was significantly lower in the MI group. Moreover, in the AP group, both the ADPV and DSVR values were significantly increased in those who had undergone percutaneous transluminal coronary angioplasty postoperatively. Different from the Doppler guidewire method, MR coronary angiography facilitates noninvasive evaluation of coronary arterial blood flow velocity. Therefore, these results indicate that MR coronary angiography represents a potentially useful technique for diagnosing lesions of coronary arteries and evaluating their functions. This noninvasive method can be expected to replace the invasive Doppler guidewire method in the near future with development of MR coronary angiography technology. PMID- 10850532 TI - Coronary artery disease and infection with chlamydia pneumonia. AB - The association between chlamydia pneumonia and coronary artery disease is well documented, however less is known about the correlation between chlamydia pneumonia infection and blood inflammatory markers or lipid levels. In 100 patients with proven coronary artery disease (25 females, 61.0 +/- 4.0 years old), and 60 healthy volunteer control cases (15 females, 60.6 +/- 3.4 years old), anti chlamydia pneumonia IgG, blood lipid, C-reactive protein and fibrinogen levels were detected. In cases with coronary artery disease seropositivity for IgG antibodies to chlamydia pneumonia (74% versus 34%, p < 0.0001), C-reactive protein (mg / l) (2.8 +/- 0.6 versus 1.4 +/- 0.6; p < 0.0001), fibrinogen (mg / dl) (317.4 +/- 38.2 versus 256.2 +/- 34.5, p < 0.0001), triglyceride (mg / dl) (217.5 +/- 39.0 versus 191.0 +/- 25.9, p < 0.0001), LDL cholesterol (mg / dl) (126.9 +/- 19.2 versus 110.6 +/- 19.5, p < 0.0001) levels and total cholesterol / HDL-cholesterol ratio (7.7 +/- 1.8 versus 4.4 +/- 1.2, p < 0.0001) were higher but the level of HDL-cholesterol (mg / dl) (26.4 +/- 6.7 versus 47.0 +/- 11.2, p < 0.0001) was lower. The levels of total cholesterol did not differ between the two groups (p=0.9). Levels of triglyceride (r=0.60, p < 0.00001), LDL-cholesterol (r = 0.27, p = 0.0004), C-reactive protein (r = 0.69, p < 0.00001), fibrinogen (r = 0.60, p < 0.00001) and total cholesterol / HDL cholesterol ratio (r = 0.74, p < 0.00001) had a direct relation, but the level of HDL-cholesterol had a negative (r= -0.80, p < 0.00001) relation with the seropositivity for chlamydia pneumonia. As a result, seropositivity for IgG antibodies to chlamydia pneumonia is considered as a risk factor for coronary artery disease by its association with the atherogenic lipid profile and procoagulant activity. PMID- 10850533 TI - QT intervals and heart rate variability in hypertensive patients. AB - Low heart rate variability and increased QT dispersion are risk factors for cardiac mortality in various patient populations. We studied dispersion of QT interval, i.e. an index of inhomogeneity of repolarization, and heart rate variability (HRV) i.e., a measure of cardiac autonomic modulation in 76 essential hypertension cases (45 women, 53.0 +/- 11.1 years, body mass index: 25.1 +/- 1.4 kg/m2) and 70 healthy cases (42 women, 54.0 +/- 10.2 years, body mass index: 25.5 +/- 1.6 kg/m2, p > 0.05). QT-corrected QT intervals and their dispersions were significantly higher in the hypertensive group (p < 0.0001), all showing a direct relation with the level of systolic and diastolic blood pressures, ventricular mass index and high Lown grade ventricular rhythm problems. Time domain measures like standard deviation of RR intervals, standard deviation of the means of all corrected RR intervals calculated at 5 min intervals (p < 0.0001), proportion of adjacent RR intervals differing by > 50 msec (p = 0.005), HRV triangular index (p = 0.007), the square root of the mean squared differences of successive RR intervals (p = 0.011), and the high frequency (HF, 0.16-0.40 Hz, p < 0.0001) part of the frequency domain measure of HRV were all decreased, whereas the low frequency (LF, 0.04-0.15 Hz, p = 0.013) part of the frequency domain measures and LF / HF ratio (p < 0.0001) were increased in hypertensive cases. Time domain and the HF part of frequency domain measures of heart rate variability showed an inverse relation with the increased levels of both systolic and diastolic blood pressures and Lown grading system of ventricular rhythm problems, whereas LF and LF / HF showed direct relations with high levels of systolic and diastolic blood pressures and high Lown grade ventricular rhythm problems. The measures of heart rate variability apart from LF and LF / HF were inversely related with the QT intervals and dispersions, whereas LF / HF was directly related with them. Therefore, we conclude that the levels of both systolic and diastolic blood pressures are related to the generation of ventricular rhythm problems either via increasing left ventricular mass which results in an increase in QT parameter measurements, or by altering heart rate variability measures indicating a disturbance in cardiac autonomic balance in essential hypertension. PMID- 10850534 TI - Polymorphisms of tumor necrosis factor-alpha and interleukin-10 genes in Japanese patients with idiopathic dilated cardiomyopathy. AB - Various cytokines play important roles in the pathogenesis of congestive heart failure. TNF-alpha is one of the pro-inflammatory cytokines, and IL-10 has anti inflammatory actions. The -308 (G / A) polymorphism of the TNF-alpha gene (TNFA1 and A2) and the single base -1082 (G / A) polymorphism of the IL-10 gene (IL-10 1*G and 1*A) have been identified as causing alterations to the in vivo production of TNF-alpha and IL-10, respectively. We examined TNF-alpha and IL-10 gene polymorphisms using a polymerase chain reaction-restriction fragment length polymorphism technique in 48 Japanese patients with idiopathic dilated cardiomyopathy. The frequency of these polymorphisms was compared with 50 healthy Japanese. The clinical courses, such as disease onset, left ventricular function, progression during the follow up period and hospitalization from congestive heart failure, were also analyzed. Serum TNF-alpha levels were measured using an enzyme linked immunosorbent assay (ELISA) technique in the patients with idiopathic dilated cardiomyopathy to reveal the correlation with genotypes. Patients with ischemic cardiomyopathy or other secondary cardiomyopathies were excluded from this study. The allele frequency of TNFA2 in idiopathic dilated cardiomyopathy was significantly higher than that of the healthy group (13.5% and 3.0%, respectively, p = 0.0084). There was no difference in the allele frequency of the IL-10 gene between the two groups. Polymorphism of the TNFA2 gene was not associated with the clinical course. Serum TNF-alpha levels were elevated in the patient group compared with the healthy group. There were no differences in serum TNF-alpha levels between the patients with TNFA1 and those with TNFA2. In conclusion, the TNFA2 allele may be linked to the pathogenesis of idiopathic dilated cardiomyopathy in Japanese patients. PMID- 10850535 TI - Experimental study of the effects of multi-site sequential ventricular pacing on the prophylaxis of ventricular fibrillation. AB - Previous studies report a significant prophylactic effect on the occurrence of atrial fibrillation by simultaneous multi-site atrial pacing. We investigated the effects of multi-site sequential ventricular pacing (MSVP), which may be preferable to simultaneous multi-site pacing in terms of the prophylaxis of the occurrence of ventricular fibrillation (VF). Needle electrodes were inserted at ten different epicardial sites on both ventricles for MSVP in 12 adult beagle dogs. Four premature ventricular extrastimuli (PVE) were introduced to provoke VF reproducibly from a separate electrode in the left ventricle. The 4 PVE were applied to try to provoke VF during MSVP in a comparable fashion to the activation sequence during sinus rhythm. We compared the prophylactic effects of MSVP on the inducibility of VF by changing the number of stimulation sites to either 1, 3, 5, or 10 epicardial sites. We performed a total of 363 trials of induction and suppression of VF. The occurrence rates of VF by the 4 PVE for the various number of epicardial stimulation sites of MSVP, i.e., at 1, 3, 5, and 10 sites, were 0.8263, 0.4286, 0.4450, and 0.2857, respectively (p < 0.05). There was a significant prophylactic effect of MSVP on the inducibility of VF, and this effect became stronger as the number of MSVP sites was increased from 3 to 10. The hemodynamic state was relatively stable during MSVP. MSVP seems to be a promising method with which to reduce the occurrence of VF, and a larger number of stimulation sites would be more effective in terms of the prophylaxis of VF. PMID- 10850536 TI - Screening for cardiac dysfunction in asymptomatic patients by measuring B-type natriuretic peptide levels. AB - Early diagnosis and treatment of heart failure lead to improved survival; pre clinical detection would thus be beneficial. A non-invasive biochemical testing method would indeed be ideal to screen for the condition. In the present study, we sought to determine whether circulating levels of B-type natriuretic peptide (BNP) correlate with cardiac function in asymptomatic subjects. 294 consenting asymptomatic subjects were examined. BNP levels in elevated patients (> 18.4 pg / ml) showed significant correlation with echocardiographic parameters of the systolic and diastolic functions (EF r = -0.51, FS r = -0.50, E/A r = 0.42, p < 0.01). Moderate correlation with the CTR on chest X-ray was also seen (r = 0.23, p < 0.01). Multiple regression analysis showed numerous echocardiographic and hemodynamic parameters including those of systolic and diastolic function in addition to left ventricular wall thickness, blood pressure and serum creatinine levels to be significantly associated with raised BNP levels. Elevated BNP levels reflect cardiac function (both systolic and diastolic) in the asymptomatic population. Detection of cardiac dysfunction by the non-invasive biochemical test may prove useful in early pre-clinical diagnosis of heart failure. PMID- 10850537 TI - Radiofrequency catheter ablation of ventricular tachycardia from the anterobasal left ventricle. AB - Ventricular tachycardia (VT) in coronary artery disease arises mostly from endocardial sites. However, little is known about the site of origin in other diseases. We report two patients who had VT originating from an anterior aspect of the left ventricle just below the mitral annulus, adjacent to the left ventricular outflow tract. The QRS configuration of VT showed an inferior axis and monophasic R waves in all the precordial leads. Radiofrequency current delivered to this site from the endocardial site successfully ablated the tachycardia in both. PMID- 10850538 TI - Patient with atrioventricular node reentrant tachycardia with eccentric retrograde left-sided activation: treatment with radiofrequency catheter ablation. AB - We describe a patient with supraventricular tachycardia with triple atrioventricular (AV) node pathway physiology. A discontinuous curve was present in the antegrade AV nodal function curves. During right ventricular pacing, the earliest retrograde atrial activation was recorded at the left-sided coronary sinus electrode. The retrograde ventricular-atrial interval was long and had decremental conduction. We induced a slow-slow AV node reentrant tachycardia (AVNRT) with eccentric retrograde left-sided activation. After slow pathway ablation, dual AV nodal pathway physiology was present. AVNRT with eccentric retrograde left-sided activation is relatively rare, and our findings suggest that eccentric retrograde left-sided atrial inputs consist partially of a slow pathway and disappear with slow pathway ablation. PMID- 10850539 TI - Occlusion of azygos vein via direct percutaneous puncture of innominate vein following cavopulmonary anastomosis. AB - A 2-year-10-month-old boy was diagnosed with a complex congenital heart disease: right atrial isomerism, left superior vena cava (LSVC), complete atrioventricular septal defect, secundum type atrial septal defect, transposition of the great arteries with pulmonary atresia, patent ductus arteriosus, absence of a right superior vena cava (RSVC), and dextrocardia. He had received a left Blalock Taussig (BT) shunt at the age of 3 months and a left bidirectional Glenn shunt one year after BT shunt. Progressive cyanosis was noted after the second operation and cardiac catheterization showed a functional Glenn shunt with an engorged azygos vein, which was inadvertently skipped for ligation. Because of the absence of RSVC, transcatheter occlusion of the azygos vein was performed successfully via direct puncture of the innominate vein. PMID- 10850540 TI - Cyanosis caused by a huge obstructive right ventricular fibroma. AB - Cardiac fibromas are rare lesions which occur more often in infants and children than in adults. These tumors are benign proliferations of connective tissue most often found in the left ventricular myocardium or septum. In an 8-month-old infant with cyanosis and progressive exertional dyspnea, a huge cardiac tumor obstructing the right ventricular outflow tract (RVOT) was diagnosed by means of 2-dimensional echocardiography and cardiac catheterization. At surgery, a whitish gray solitary tumor measuring 5.0 x 4.5 cm could be well visualized. It was nearly totally resected, and the RVOT was reconstructed with an Equine pericardial patch. Histologic examination classified the tumor as a fibroma. Although surgical mortality in cardiac fibroma with RVOT obstruction is extremely high, early diagnosis and prompt excision of the tumor is mandatory in relieving its dangerous symptoms. PMID- 10850541 TI - Molecular detection of Mycobacterium tuberculosis in tissues showing granulomatous inflammation without demonstrable acid-fast bacilli. AB - Early diagnosis of tuberculosis (TB) is important for early medical intervention and prevention of spread of the bacteria. It is not uncommon to observe granulomatous inflammation but without demonstrable acid-fast bacilli (AFB) on Ziehl-Neelsen (ZN) staining in tissues sent for histologic examination, and the definitive diagnosis of TB cannot be made because no concurrent tissue is sent for TB culture. In this study, the authors explored the feasibility of using polymerase chain reaction (PCR) for early detection of Mycobacterium tuberculosis (Mtb) in formalin-fixed, paraffin-embedded tissues where a definite diagnosis of TB cannot be made. One hundred fifteen patients (131 paraffin blocks of biopsy specimens) with granulomatous inflammation but ZN-negative for AFB were studied. DNA was extracted from paraffin sections and amplified by PCR with the IS6110 primers (specific for the Mtb complex) and the specific 122-base pairs (bp) PCR product was detected by agarose gel electrophoresis. Sixty-eight of the 115 (59%) patients were TB-PCR positive, thus enabling definite diagnosis of TB in significant numbers of these patients in 3 days. The authors conclude that molecular diagnosis by PCR is useful for early detection of TB in histologic material where morphologic features are suggestive but not confirmatory because of negative staining for AFB. PMID- 10850542 TI - Implication of screening for FMR1 and FMR2 gene mutation in individuals with nonspecific mental retardation in Taiwan. AB - Fragile X syndrome (FXS) is the most common form of familial mental retardation (MR), attributable to (CGG)n expansion in the FMR1 gene. FRAXE is less frequent, associated with a similar mutation of the FMR2 gene. This study attempted to ascertain the prevalence of both disorders in Taiwan, as well as to develop a method to effectively find carriers. A total of 321 patients with nonspecific MR were screened for the FMR1 and FMR2 mutation. Four of 206 boys and men (1.9%) and 1 in 115 girls and women (0.9%) were identified as having FXS. All four FXS boys or men could be identified by Southern blot analysis, as well as by a simple nonradioactive polymerase chain reaction analysis. None of the 206 boys or men had FMR2 full mutation. This confirmed the low incidence of FRAXE in Chinese. FXS appears to be more prevalent among patients with mild MR, because 4 of the 5 patients with FXS were from the 115 with mild MR (3.48%) and only 1 was from the other 206 with severe MR (0.49%). All five FXS cases were maternally inherited. Other family members were resistant to further searching for carriers. It is worth noting that none of these mothers had a discernible premarital family history of MR. Thus the negative family history could not preclude the possibility that a woman was a carrier. To identify female carriers of childbearing age, beyond the scope of family history, is thus worthy of further exploration. Screening men for carriers using this inexpensive method is probably feasible, even though normal transmitting men have no immediate risk of producing a child with the disease. Female carriers can then be effectively identified from these normal transmitting men and can take all preventive measures. PMID- 10850543 TI - Diagnosis of quantitative mitochondrial DNA defects by rapidly prepared whole mitochondrial DNA probe. AB - Analysis of disease-causing mutations in mitochondria genome requires rapid and reliable genetic approaches. However, the preparation of mitochondrial DNA (mtDNA) probe used for the determination of quantitative and qualitative mtDNA defects is time-consuming, cumbersome, and requires complicated instrumentation. To overcome the difficulties encountered during isolation and purification of mtDNA, the authors developed an alternative method based on polymerase chain reaction (PCR) amplification of whole mtDNA genome. In this study, they show that PCR-amplified and fluorescein-labeled mtDNA probe makes it possible, through Southern blot analysis, to identify quantitative defect of mtDNA. The results indicate that mtDNA probe can be prepared rapidly by PCR amplification and used to determine the level of mtDNA in the patients with mitochondrial diseases. PMID- 10850544 TI - Five useful approaches for generating more valid gel images of loss of heterozygosity and clonality analysis with an automated 377 DNA sequencer. AB - The recently introduced fluorescence-based gene scan system for assessment of loss of heterozygosity (LOH) and clonality with an automated DNA sequencer has several advantages over the traditional method. However, the production of gel images with this system is subjected to more technical challenges, including the interference of autofluorescence, weaker and less consistent signals that result from the restricted well size and difficulties in sample loading. To minimize the impact of these technical difficulties, several unique strategies were used, including the following: elimination of fabrics or paper towels in the cleaning of gel plates and containers; use of a modified loading buffer; use of more concentrated gels; use of an innovative apparatus to clean gel wells before and after the prerun; and covering the black printer cartridge with a sheet of scotch tape. With these strategies, the authors have been able to consistently obtain gel images that can be presented as either densitometric graphs or as band patterns for direct visual assessment. PMID- 10850545 TI - Comparison of automated short tandem repeat and manual variable number of tandem repeat analysis of chimerism in bone marrow transplant patients. AB - Hematopoietic chimerism can be monitored in bone marrow transplant patients at DNA polymorphic sites. In this study, allele detection and quantification by ethidium bromide-stained agarose gels were compared with automated fluorescent sizing on an artificially mixed system and on chimeric post-transplant whole blood and sorted cell populations. A panel of five variable number of tandem repeats (VNTRs) were amplified and quantified visually on an ethidium bromide stained gel. The ten short tandem repeats (STRs) were amplified as a multiplex polymerase chain reaction (PCR) and fluorescently detected on a DNA sequencer. Fluorescent band intensities were converted to fluorescent peak areas for allele quantification. Using mixed DNA of different proportions, both STRs and VNTRs showed linearity and appeared equally sensitive. However, case studies showed STRs to be more sensitive (<5%) than VNTRs (<10%). The STRs more accurately quantified the minor DNA component at low concentrations. PMID- 10850546 TI - Normalizing complementary DNA by quantitative reverse transcriptase-polymerase chain reaction of beta2-microglobulin: molecular monitoring of minimal residual disease in acute promyelocytic leukemia. AB - Reverse transcription (RT)-polymerase chain reaction (PCR) raises unique methodological matters that may hamper the reliability of the procedure, especially when results should direct therapeutic decisions. One of these matters is represented by the RT step. The present study shows that differences in complementary DNA (cDNA) preparations purposely containing increasing amounts of retrotranscribed RNA were not disclosed by nonquantitative RT-PCR by two different housekeeping genes, leading to fictitious results when the expression of a given gene was quantitatively assessed. To overcome this problem, the following are proposed: 1) to evaluate the efficiency of RT step through the quantification, by competitive RT-PCR, of the expression levels of the housekeeping gene beta2-microglobulin (beta2M); 2) to normalize each cDNA preparation to be comprised within 1 standard deviation of the mean value of beta2M absolute level (3.14 +/- 1.14 attomoles/microg RNA) found by analyzing 33 cell lines of hematopoietic origin. To validate this strategy in a clinical setting, serial cDNA samples from patients were checked by conventional and quantitative RT-PCR for beta2M. Again, only a quantitative evaluation of beta2M levels was allowed to unveil significant differences, otherwise undetected, in the efficiency of RT reactions among these cDNA samples. Normalization of samples to obtain cDNA preparations containing comparable beta2M levels, eventually led to an increased sensitivity in the detection of PML-RARalpha fusion transcripts. This approach seems of great value for the monitoring of minimal residual disease in serial patient samples when a tumor-specific marker is available. PMID- 10850547 TI - Novel polymerase chain reaction approach for full-coding p53 mutation detection in microdissected archival tumors. AB - Evidence suggests that up to 25% of p53 mutations are outside of exons 5-8 and that insertions, deletions, and polymorphic sites in the p53 gene may play a significant role in the process of carcinogenesis. A novel polymerase chain reaction (PCR) approach for the analysis of the entire p53 coding and splice site regions from microdissected, formalin-fixed, paraffin-embedded tumor tissues has been developed which allows multiple genetic analyses to be performed from one primary amplification reaction. The method was initially evaluated using well characterized cell lines. In addition to confirming the published p53 mutations for HT29, Molt 4, A431, and HN5, a 16 base pair (bp) duplication within intron 3 was detected in both the A431 and HT29 cell lines. Analysis of archival samples of ovarian cancer identified the same 16-bp duplication and coding region variations. In all samples, using GenBank submission U94788 as a reference, a C insertion was detected at nucleotide positions 11818 and 11874 within intron 2. At nucleotide position 14168, within intron 7, a T-to-G base change was found. This novel PCR approach has the potential to reduce the amount of clinical material required by up to 95%, thus facilitating retrospective studies on archival tumor collections. Furthermore, a wider analysis of the p53 gene, including splice sites and intronic regions, may yield additional information regarding cancer predisposition, response to therapy, and progression. PMID- 10850549 TI - Effects of ammonia on pentylenetetrazole-induced seizure threshold. AB - The effect of chronic perfusion of ammonia on the seizure threshold against pentylenetetrazol was studied. Ammonia plus sodium bicarbonate and saline (0.9%) was continuously administered to two groups of rats respectively. All animals were tested three times for seizure threshold, and were then decapitated and the brains removed for analysis of the amino acids. The results showed that the infusion of ammonia increased the seizure threshold, and this protective effect was accompanied by selective changes in brain glutamate and glutamine. Thus, continuous infusion of ammonia may cause an imbalance between excitatory and inhibitory systems in favor of inhibitory systems. These findings may provide insights into the basic mechanisms of seizures observed in hepatic failure, in other hyperammonemic states, and in epilepsy. PMID- 10850550 TI - Reversible alterations in brain metabolites during therapy for disseminated nocardiosis using proton magnetic resonance spectroscopy. AB - We report reversible abnormalities in magnetic resonance spectra acquired from a patient with AIDS undergoing antibiotic and corticosteroid therapy for disseminated nocardiosis, a rare opportunistic infection of immunosuppressed patients which can cause cerebral abscess formation. There was no clinical, CT or MRI evidence of HIV-1 encephalitis. MR spectra were acquired before and after treatment using a two-dimensional chemical shift imaging technique (TR 1500ms, TE 130ms). Prior to treatment, a rise in the choline to creatine ratio and a reduction in the N-acetylaspartate to creatine ratio were observed in MR spectra localized to areas of the left anteromedial centrum semiovale that appeared normal on MR imaging. After 16 weeks, the patient had recovered with complete resolution of the cerebral abscesses on MRI. The MR spectral abnormalities also returned to normal. Two months later, the patient had a relapse with focal neurological signs and further abscesses were demonstrated on MRI of the brain. The patient subsequently died and histopathological and microbiological findings at autopsy confirmed the clinical picture of a recurrence of cerebral nocardiosis with no evidence of HIV-1 encephalitis. This case illustrates reversible MR measurable metabolite changes in the brain of an HIV-seropositive patient without HIV-1 encephalitis who underwent treatment for cerebral nocardiosis. PMID- 10850548 TI - Adenoviruses encoding HPRT correct biochemical abnormalities of HPRT-deficient cells and allow their survival in negative selection medium. AB - The Lesch-Nyhan syndrome is an X-linked disorder caused by a virtually complete absence of the key enzyme of purine recycling, hypoxanthine-guanine phosphoribosyltransferase (HPRT). It is characterized by uric acid overproduction and severe neurological dysfunction. No treatment is yet available for the latter symptoms. A possible long-term solution is gene therapy, and recombinant adenoviruses have been proposed as vectors for gene transfer into postmitotic neuronal cells. We have constructed an adenoviral vector expressing the human HPRT cDNA under the transcriptional control of a short human cytomegalovirus major immediate early promoter (RAd-HPRT). Here we show that infection of human 1306, HPRT-negative cells with RAd-HPRT, expressed high enough levels of HPRT enzyme activity, as to reverse their abnormal biochemical phenotype, thus enhancing hypoxanthine incorporation and restoring purine recycling, increasing GTP levels, decreasing adenine incorporation, and allowing cell survival in HAT medium in which only cells expressing high levels of HPRT can survive. Infection of murine STO cells, increased hypoxanthine incorporation and restored purine recycling, thus allowing cell survival in HAT medium, and reduced de novo purine synthesis. Although both cells were able to survive in HAT medium post infection with RAd-HPRT, some of the biochemical consequences differed. In summary, even though adenoviral vectors do not integrate into the genome of target HPRT deficient human or murine cells, RAd-HPRT mediated enzyme replacement corrects abnormal purine metabolism, increases intracellular GTP levels, and allows cells to survive in a negative selection medium. PMID- 10850551 TI - Elevated serum levels of astroglial S100beta in patients with liver cirrhosis indicate early and subclinical portal-systemic encephalopathy. AB - Portal-systemic encephalopathy is the prototype among the neuropsychiatric disorders that fall under the term Hepatic Encephalopathies. Ammonia toxicity is central to the pathophysiology of Portal-systemic encephalopathy, and neuronal ammonia toxicity is modulated by activated astrocytes. The calcium-binding astroglial key protein S100beta is released in response to glial activation, and its measurement in serum only recently became possible. Serum S100beta was determined by an ultrasensitive ELISA in patients (n=36) with liver cirrhosis and transjugular intrahepatic portosystemic stent-shunt. Subclinical portal-systemic encephalopathy and overt portal-systemic encephalopathy were determined by age adjusted psychometric tests and clinical staging, respectively. Serum S100beta, was specifically elevated in the presence of subclinical or early portal-systemic encephalopathy, but not arterial ammonia. S100 levels elevated above a reference value (S100beta < or = 110pg/ml) or the cut off value determined in our group of patients (112pg/ml) predicted subclinical portal-systemic encephalopathy with a specificity and sensitivity of 100 and 56.5%, respectively. Serum S100beta was significantly dependent on liver dysfunction (Child-Pugh score), but was more closely related to cognitive impairments than the score. Serum S100beta seems to be a promising biochemical surrogate marker for mild cognitive impairments due to portal-systemic encephalopathy. PMID- 10850553 TI - Proline administration decreases Na+,K+-ATPase activity in the synaptic plasma membrane from cerebral cortex of rats. AB - Buffered proline was injected subcutaneously into rats twice a day at 8 h intervals from the 6th to the 28th day of age. Control rats received saline in the same volumes. The animals were weighed and killed by decapitation 12 h after the last injection. Cerebral cortex was used for the determination of Na+,K+ ATPase and Mg2+-ATPase activities. Body, whole brain and cortical weights were similar in the two groups. Na+,K+-ATPase activity was significantly reduced (by 20%) in membranes from the proline-treated group compared to the controls, whereas Mg2+-ATPase activity was not affected by proline. In another set of experiments, synaptic plasma membranes were prepared from cerebral cortex of 29 day-old rats and incubated with proline at final concentrations ranging from 0.1 to 2.0 mM. Na+,K+-ATPase activity, but not Mg2+-ATPase activity, was inhibited by 20-30%. Since proline concentrations in plasma of chronically treated rats and of type 11 hyperprolinemic children are of the same order of magnitude as those tested in vitro, the results suggest that reduction of Na+,K+-ATPase activity may contribute to the neurological dysfunction found in some patients affected by type II hyperprolinemia. PMID- 10850552 TI - GABA-transaminase antisense oligodeoxynucleotide modulates cocaine- and pentylenetetrazol-induced seizures in mice. AB - The mechanism of action of many anticonvulsive agents is to increase the function of the GABAergic system. Inhibition of GABA-Transaminase (GABA-T), the degradative enzyme for GABA, increases GABA levels in the brain. In this study, antisense oligodeoxynucleotides (ASO) targeted at the start codon region of GABA Transaminase mRNA were used to modify seizure activity. Mice were treated, by intracerebroventricular injection, with antisense oligos or appropriate controls. At various times after treatment, the animals were challenged with cocaine (70 mg/kg, i.p.) and observed for seizure activity. At 15 hours after treatment, 1.152 and 1.44 nmol antisense oligo blocked cocaine-induced seizures. There was no effect of antisense oligo 8 or 36 hours after treatment. In addition, treatment with 7.2 nmol antisense oligo prevented pentylenetetrazol-induced seizures. These data demonstrate the modulation of seizure threshold using antisense oligodeoxynucleotides to GABA-T. PMID- 10850555 TI - A different view of thrombosis. AB - In this paper we discuss some of the physical difficulties associated with the classical view of thrombosis and conclude that there are problems associated with these concepts. We propose an alternative model based on our published observations that, during ex-vivo thrombus formation, the platelets, which normally do not stain for tissue factor, become tissue factor positive. These hybrid species are capable not only of propagating coagulation, but also of initiating it. Thus thrombi can grow without evoking long-range diffusion -- a very slow and inefficient mechanism, particularly in a flowing system. PMID- 10850554 TI - Effect of portacaval anastomosis on glutamine synthetase protein and gene expression in brain, liver and skeletal muscle. AB - The effects of chronic liver insufficiency resulting from end-to-side portacaval anastomosis (PCA) on glutamine synthetase (GS) activities, protein and gene expression were studied in brain, liver and skeletal muscle of male adult rats. Four weeks following PCA, activities of GS in cerebral cortex and cerebellum were reduced by 32% and 37% (p<0.05) respectively whereas GS activities in muscle were increased by 52% (p<0.05). GS activities in liver were decreased by up to 90% (p<0.01), a finding which undoubtedly reflects the loss of GS-rich perivenous hepatocytes following portal-systemic shunting. Immunoblotting techniques revealed no change in GS protein content of brain regions or muscle but a significant loss in liver of PCA rats. GS mRNA determined by semi-quantitative RT PCR was also significantly decreased in the livers of PCA rats compared to sham operated controls. These findings demonstrate that PCA results in a loss of GS gene expression in the liver and that brain does not show a compensatory induction of enzyme activity, rendering it particularly sensitive to increases in ammonia in chronic liver failure. The finding of a post-translational increase of GS in muscle following portacaval shunting suggests that, in chronic liver failure, muscle becomes the major organ responsible for the removal of excess blood-borne ammonia. PMID- 10850557 TI - Models of blood coagulation. AB - We have used three models to study the process of tissue factor-initiated blood coagulation. These are: synthetic 'plasma' mixtures prepared with the proteins and membranes involved in the reaction and its regulation; mathematical models based on the reaction kinetics, binding constants and stoichiometries of individual procoagulant and inhibitor reactions, and contact pathway-inhibited coagulation of minimally altered whole blood in vitro. In all of these models, the procoagulant process may be divided into two phases: an initiation phase and a propagation phase. The initiation phase is characterized by the appearance of thrombin and other coagulation enzymes, and the activation of pro-cofactors V and VIII. The propagation phase is characterized by explosive and extensive prothrombin activation. During normal blood coagulation, the bulk of thrombin generation occurs after clot formation, while most release of fibrinopeptide A is observed just at the conclusion of the initiation phase. In the case of haemophilia A and B, the initiation phase is slightly extended, while thrombin generation during the propagation phase is significantly suppressed. The clot time, as well as fibrinopeptide release, is delayed in these patients. Data obtained in our laboratory, employing the above models, indicate that they are efficient tools for blood coagulation studies. PMID- 10850556 TI - The effect of factor X level on thrombin generation and the procoagulant effect of activated factor VII in a cell-based model of coagulation. AB - We used a cell-based, in-vitro model of normal hemostasis and hemophilia to address the question of whether factor (F) X concentration affects the hemostatic response to high-dose activated factor VII (FVIIa). Under conditions designed to mimic normal tissue factor-initiated hemostasis in vivo, we found that only a very small amount of FX -- equivalent to about 3% of the normal plasma level -- was required to support a 'normal' level of thrombin generation. This suggests that, under normal conditions in vivo, the level of FX does not significantly affect hemostatic function. By contrast, in experiments designed to mimic the hemophilic condition, the level of FX had a significant effect on the level of thrombin generated in the presence of high-dose FVIIa. This finding suggests that the plasma level of FX could affect the hemostatic response of hemophilic patients to high-dose FVIIa therapy. PMID- 10850558 TI - Structure of human coagulation activated factor VII. AB - Activated factor VII (FVIIa) is a trypsin-like serine protease that plays a key role in the initiation of the blood coagulation cascade. FVIIa, which comprises a light chain (152 residues) and a heavy chain (254 residues) linked by a disulphide bond, is generated by the cleavage of the Arg152-Ile153 peptide bond in factor (F)VII. While a corresponding internal peptide bond cleavage unleashes the activity of other trypsin-like enzymes, FVIIa is unusual in that it remains in a zymogen-like state after cleavage and only becomes an efficient catalyst when associated with tissue factor, its protein cofactor and allosteric regulator. We have determined the structure of free FVIIa lacking the gamma carboxyglutamic acid (Gla) domain as a means to elucidate the molecular reasons for its poor activity when not bound to tissue factor and the conformational changes induced by its association with tissue factor. PMID- 10850560 TI - Management and monitoring of recombinant activated factor VII. AB - Recombinant factor VIIa (rFVIIa) has recently been introduced as a new 'bypassing' agent to improve haemostasis in haemophilia patients with inhibitors to factor (F) VIII or FIX. In noninhibitor patients, levels of circulating FVIII or FIX can be used to assess the quality of substitution therapy. In contrast, laboratory monitoring of haemostatic efficacy in patients treated with rFVIIa has proved more complex. Evaluation of patient samples saved during rFVIIa treatment have revealed some correlation between FVII:C levels and improved haemostasis, while whole blood elasticity, as determined by thromboelastography, has been shown to improve following rFVIIa treatment. The investigation aimed to study the efficacy of rFVIIa in activating FX:C and in shortening the whole blood clotting time (WBCT) using the newly-developed roTEG coagulation instrument. Results indicated a substantial and apparently dose-dependent activation of FX:C by rFVIIa. In addition, the presence of FIX appeared to influence FX:C activation. In-vitro and ex-vivo roTEG thromboelastograph measurements showed that FVIIa shortened WBCT, although normalization did not occur. The results presented here are based on a small number of observations in a few patients and further studies should be planned to test the efficacy of these monitoring principles in clinical treatment practice with rFVIIa. PMID- 10850559 TI - Overall experience with NovoSeven. AB - Recombinant activated factor VII (rFVIIa, NovoSeven) was first developed for treating those haemophilia patients with inhibitors who cannot benefit from conventional therapies. Several clinical trials have clearly demonstrated that rFVIIa is a safe and effective therapy for home treatment of mild-to-moderate bleeding episodes. Its theoretical inability to abnormally activate the coagulation system has also prompted many clinicians to use it in elective surgical procedures. Recommended dose ranges for rFVIIa usually vary from 60 to 120 microg/kg, although 90 microg/kg is generally accepted as an initial treatment dose. If necessary, further rFVIIa can be administered as bolus injections every 2 to 6 h or, alternatively, as a continuous infusion. In patients with congenital haemophilia, this treatment is effective in up to 92% of cases. Recombinant FVIIa has also been successfully used in patients with acquired haemophilia where results have shown a wide safety margin, suggesting that rFVIIa should be considered as first-line therapy. The prospect of extending the indication of rFVIIa exists, and preliminary reports suggest that rFVIIa could be useful in patients with congenital or acquired platelet disorders, thrombocytopenia or liver failure. This drug represents a major advance in the treatment and prevention of bleeding in predisposed patients. PMID- 10850561 TI - Experience with continuous infusion of recombinant activated factor VII in the Asia-Pacific region. AB - There is increasing interest in continuous infusion of recombinant activated factor VII (rFVIIa) as a convenient and safe alternative to intermittent bolus therapy. In the Australian patients reported in this paper, cost savings of up to 25% in the first 12 h of treatment with continuous infusion of rFVIIa have been achieved safely, suggesting that substantial overall savings are possible. However, in the Thai patient reported, a dose reduction of 35% in the first 12 h was associated with poor haemostatic control, suggesting that a dose reduction of >25% may be inadvisable. The indications for treatment in the five Australian patients were: retroperitoneal haemorrhage (n = 3); right forearm compartment syndrome (n = 1); wrist haemarthrosis and median nerve compression (n = 2); sublingual haematoma (n = 1); and cerebral (mid-brain) haemorrhage (n = 1). Treatment was effective in four out of five patients (six bleeding episodes) and there was one treatment failure where treatment had been substantially delayed. The Thai patient was treated as part of a prospective, uncontrolled, observational study of 34 bleeding episodes in 22 patients in the Asia-Pacific region. Treatment was judged ineffective after 24 h, but full haemostatic control was subsequently achieved with intermittent rFVIIa therapy. PMID- 10850562 TI - Treatment of bleeding episodes in patients with hemophilia and an inhibitor: comparison of two treatment protocols with recombinant activated factor VII. AB - Six hemophilia A patients with inhibitors were treated with a continuous infusion of recombinant activated factor VII (rFVIIa) for various bleeding episodes. Bleeding episodes (n = 101) were treated according to a 12 h regular dose protocol or a shortened (6 h) augmented dose protocol. Patient response to therapy was assessed by symptomatic improvement within predefined timeframes. Although patient number was limited, both protocols appeared similar with respect to the proportion of patients responding to therapy; however, the augmented dose protocol appeared to be superior to the regular dose protocol with shorter response time, shorter duration of therapy and possibly lower rFVIIa requirements. Further studies are needed to define the efficacy of rFVIIa augmented dose administered as continuous infusion in treating hemophilia patients with inhibitors during major bleeding episodes, trauma and surgery. PMID- 10850563 TI - NovoSeven in immune tolerance therapy. AB - The development of inhibitors in haemophilia patients is one of the most serious challenges to effective treatment. The effect of factor (F) concentrate and treatment regimen on titre levels was studied in nine patients with haemophilia A or B and inhibitors. Patients with haemophilia A were subdivided into those treated with recombinant activated factor VII (rFVIIa) on demand or those treated prophylactically with activated prothrombin complex concentrate (aPCC) and rFVIIa. A third group comprised haemophilia B patients receiving rFVIIa prophylaxis and on-demand aPCC or rFVIIa. On-demand therapy with rFVIIa proved to be effective and safe treatment for acute bleeding episodes in haemophilic patients. In group 1, inhibitor titres decreased markedly 0.5-21 months prior to immune tolerance therapy (ITT) and remained low for a further 1-19 months. However, use of rFVIIa instead of aPCC as prophylactic treatment in group 2 patients undergoing ITT failed to show favourable results for rFVIIa. This may be due to the short half-life of rFVIIa (2.3-2.9 h). The data presented suggest that exclusive use of rFVIIa in acute bleeding episodes prior to commencing ITT is an effective method of decreasing inhibitor titre, thereby optimizing conditions for ITT. PMID- 10850564 TI - Acute hemarthroses: the benefits of early versus late treatment with recombinant activated factor VII. AB - In hemophiliacs without inhibitors, response to treatment for joint and soft tissue bleeding is much more effective when factor VIII is given early. The same is true in hemophiliacs with inhibitors treated with recombinant activated factor VII (rFVIIa; NovoSeven) for peripheral muscular hemorrhages. In the present report, we analyzed the responses to rFVIIa given to treat acute hemarthroses. We compared the amount of rFVIIa used in the treatment of acute hemarthroses in persons with hemophilia A and B with inhibitors and those with acquired hemophilia in the compassionate use, dose-finding, and United States (US) home treatment studies. We also compared the response rates in each of these studies. As in previous analyses for intramuscular hemorrhages, the average number of doses given for acute joint bleeding was significantly less when treatment was instituted early (as in the US home treatment study). Response rates were also much greater in the US home treatment study than in the dose-finding or compassionate use databases. It is clear that early treatment of acute hemarthroses with rFVIIa results in a greater rate of success, with fewer doses of product being required. Home treatment with rFVIIa results in greater convenience, cost savings and reduced morbidity. PMID- 10850565 TI - Patient/caregiver assessment of convenience in the use of recombinant activated factor VII (rVIIa; NovoSeven) in home therapy. PMID- 10850566 TI - Platelet physiology and function. PMID- 10850567 TI - Recombinant activated factor VII (NovoSeven) treatment of platelet-related bleeding disorders. International Registry on Recombinant Factor VIIa and Congenital Platelet Disorders Group. AB - Recombinant activated factor VII (rFVIIa; NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark), used extensively for the management of hemophilia patients with inhibitors, has also been shown to be effective in the treatment of severe bleeding episodes and for coverage of surgical procedures in patients with platelet disorders. Cases include seven patients with congenital platelet disorders [Glanzmann thrombasthenia (n = 5), Bernard-Soulier syndrome (n = 1), platelet type (pseudo-) von Willebrand disease (n = 1)] and two patients with acquired thrombocytopathy associated with myelodysplastic syndrome and uremia. The clinical efficacy of rFVIIa in functional platelet disorders has been reported as good or excellent, although some cases of ineffectiveness exist. The agent is well tolerated with a single published case of thromboembolism as a postoperative complication. In addition to these reported cases, there are others that remain unreported and unpublished. An International Registry on Recombinant Factor VIIa and Congenital Platelet Disorders (forms in Appendix 1) has been established to obtain more safety and efficacy data on patients with congenital platelet disorders treated with NovoSeven. Analysis of data from this larger population will allow better comprehension of the role of NovoSeven in these disorders, and assist in the design of formal studies to address issues associated with the treatment of these disorders. PMID- 10850568 TI - Coagulation problems in liver disease. AB - Three topics are addressed in this manuscript: (1) the causes of abnormal hemostasis in cirrhosis; (2) the evaluation of hemostasis in cirrhotic patients before invasive procedures or in the presence of bleeding; and (3) the assessment of the effect of recombinant activated factor VII (rFVIIa; NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) on hemostatic function in cirrhotic patients. Laboratory experiments are described that could enhance the understanding of the effect of rFVIIa on blood coagulation during hemostasis in cirrhotic patients. PMID- 10850569 TI - Clinical aspects of bleeding complications in cirrhotic patients. AB - Liver disease is a frequent cause of haemostatic abnormalities, which may lead to overt or occult bleeding. Clinical manifestations of hepatic coagulopathy include upper and lower gastrointestinal haemorrhage, easy bruising and bleeding from gums, nose or the female genital tract. The most significant bleeding problem among patients with chronic liver disease is blood loss due to portal hypertension. About 30% of subjects with oesophageal or gastric varices resulting from cirrhosis have an episode of gastrointestinal bleeding in their lifetime. Risk factors for the first episode of variceal bleeding include the severity of liver dysfunction, large varices, and the presence of endoscopic red colour signs. Bacterial infection in patients with variceal haemorrhage may be critical in triggering bleeding. Nongastrointestinal bleeding events, either spontaneous or induced by minor trauma, are also a common complication of advanced cirrhosis. In women, for instance, dysfunctional uterine bleeding may become so severe that hysterectomy is required. In addition, invasive diagnostic tests (mostly solid tissue biopsies) and surgical procedures have a high risk of haemorrhage and are sometimes withheld in cirrhotic patients for fear of complications. In patients with portal hypertension, surgical procedures aggravate the injury of the hepatic parenchyma and may worsen the condition. PMID- 10850570 TI - Bleeding problems in patients undergoing segmental liver transplantation. AB - Liver transplantation is associated with a great risk of bleeding, which is of both surgical and medical etiopathogenesis. All phases of the transplantation are characterized by a complicated and continuously evolving derangement of pro- and anticoagulant factors, which result in coagulopathy. The magnitude of this coagulopathy varies between patients and therefore must be corrected on an individual basis. During the different phases of the transplantation, treatment must be targeted with the aim of controlling the bleeding according to its pathogenesis. This paper analyzes the problems associated with the monitoring and correction of coagulation during the different phases of liver transplantation, with specific focus on the idiosyncrasies of segmental liver transplantation. PMID- 10850571 TI - Blood loss in orthotopic liver transplantation: a retrospective analysis of transfusion requirements and the effects of autotransfusion of cell saver blood in 164 consecutive patients. AB - Liver transplantation is associated with excessive blood loss. In order to identify factors influencing blood loss and to provide a basis for a pilot study to evaluate recombinant activated factor VII as a haemostatic agent, a retrospective study was performed in 164 consecutive patients with cholestatic or noncholestatic liver disease, who underwent orthotopic liver transplantation at a single centre between 1989 and 1996. Transfusion of allogeneic and autologous (cell saver) blood was used as a measurement of blood loss. Transfusion requirements were associated with age, gender, primary disease, Child-Pugh classification, serum levels of activated partial thromboplastin time, antithrombin III, urea and creatinine, platelet number, year of transplantation, length of cold ischaemia time and autologous blood transfusion. Of these variables, Child-Pugh classification (P = 0.001), urea (P = 0.0007), year of transplantation (P = 0.002), cold ischaemia time (P = 0.01) and autologous blood transfusion (P < 0.0001) were independent predictors of transfusion requirements by multivariate analysis. Thus, blood loss and transfusion requirements depend primarily on the severity of liver disease, quality of the donor liver, experience of the transplantation team and use of autologous (cell saver) blood transfusion. These findings emphasize the need for appropriate drug therapy and a critical reappraisal of current transfusion policy. PMID- 10850572 TI - Measurement of the procoagulant activity of factor VII in patients with liver cirrhosis and normal prothrombin activity: evaluation of the bleeding risk. AB - Patients with liver cirrhosis and diminished prothrombin activity (PA) have decreased levels of factor (F)VII coagulation activity (FVII:C) and an increased bleeding tendency. Whether this is also true of cirrhotic patients with normal PA is unknown. This study measured FVII:C levels in such patients and investigated the correlation between altered FVII:C levels and bleeding tendency. Fifteen of 41 patients (37%) had decreased FVII:C levels. Of these, the Child-Pugh score of liver function was A (n = 9), B (n = 5) and C (n = 1), compared to A (n = 25) and B (n = 1) in patients with normal FVII:C values (chi2 = 8.88, P = 0.012). Bleeding time was significantly prolonged in 9/15 patients (60%) with impaired FVII:C activity, compared to 3/26 (12%) patients with normal FVII:C values (relative risk: 5.2, 95% CI: 1.7-16.6; P = 0.003). In conclusion, liver cirrhosis patients may show impaired FVII:C levels despite normal PA. In those with decreased FVII:C activity, prolonged bleeding time is hypothesized to arise from an alteration in platelet activation due to FVII deficiency and diminished platelet count. Bleeding risk should be evaluated, regardless of platelet count, before these patients are subjected to invasive diagnostic or surgical procedures. PMID- 10850573 TI - Recombinant activated factor VII in children with acute bleeding resulting from liver failure and disseminated intravascular coagulation. AB - Recombinant activated factor VII (rFVIIa) was given to three children with acute bleeding resulting from liver failure and disseminated intravascular coagulation. Cases I and II (girls aged 3 years and 6 years, respectively) were diagnosed with Dengue hemorrhagic fever and prolonged shock. Case III, a boy aged 9 months, underwent left lobe hepatectomy for a hepatoblastoma, during which 60% of his liver was removed. This case was complicated by myoglobinuria, liver and renal impairment and early disseminated intravascular coagulation. All three patients exhibited active bleeding. Cases I and II received rFVIIa combined with other blood component replacement, while Case III received rFVIIa as the only hemostatic agent. A bolus of 40-180 microg/kg b.w. was administered followed by 16.5-33 microg/kg b.w. per h continuous infusion. As a result, bleeding was controlled, the prothrombin time was shortened and FVII clotting activity was significantly increased. In conclusion, rFVIIa has shown some efficacy in controlling acute bleeding in children with liver failure and disseminated intravascular coagulation. PMID- 10850574 TI - NovoSeven as a universal haemostatic agent. AB - Initiation of haemostasis involves the formation of a complex between tissue factor (TF) and activated factor VII (FVIIa) following injury. TF is found in the deeper layers of the vessel wall, in atherosclerotic plaques and in some types of tumour cell and is only exposed to circulating blood after tissue damage. Likewise, FVII is only enzymatically active when complexed with TF (TF/FVIIa). It has recently been shown that the administration of recombinant activated FVII (rFVIIa) in high doses (approximately 100 microg/kg) can induce haemostasis in the absence of FVIII and FIX. In addition, from in-vitro studies it appears that rFVIIa can bind with low affinity to the activated platelet surface and, independently of TF, induce the thrombin burst needed for haemostasis. The ability of rFVIIa to compensate for FVIII/FIX deficiency has been proven clinically in haemophilia patients with life- and limb-threatening bleeds. In addition, patients with congenital FVII deficiency have been successfully treated for bleeds with rFVIIa. Recombinant FVIIa has been used in patients with platelet disorders; five patients with Glanzmann's thrombasthenia and one with Bernard Soulier's thrombasthenia have had bleeding episodes managed effectively. Recombinant FVIIa has also been shown to normalize prothrombin time in patients with liver disease and in warfarin-treated individuals. PMID- 10850575 TI - NovoSeven in warfarin-treated patients. AB - Haemorrhages represent a major complication of treatment with vitamin K antagonists. In cases of severe bleeding, a prompt effect on the increased International Normalized Ratio value is vital to achieve haemostasis. As conventional treatment, that is plasma or plasma-derived concentrates, carries the risk of blood-borne virus transmission, new treatments are needed. An open, multicentre pilot trial is currently under way to determine the effect of recombinant activated factor VII (rFVIIa; NovoSeven) administered to patients experiencing a bleeding episode after receiving vitamin K antagonists. When rFVIIa was given to a patient with a warfarin-induced nosebleed, it had an immediate haemostatic effect and the International Normalized Ratio value virtually normalized. PMID- 10850576 TI - Potential role of NovoSeven in the prevention of rebleeding following aneurysmal subarachnoid haemorrhage. AB - Rebleeding following aneurysmal subarachnoid haemorrhage is a major factor contributing to unfavourable outcome. Antifibrinolytic agents reduce the rate of rebleeding but increase the risk of cerebral ischaemia and infarction and hence provide no overall benefit. To address the theoretical concern that recombinant activated factor VII (NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) might increase the risk of cerebral ischaemia while stabilizing the clot at the site of aneurysmal rupture, an open-label, dose-escalation safety study has been developed in collaboration with the UK Spontaneous Intracranial Haemorrhage Group. The trial design includes the recruitment of 15 patients (aged 18 years or over) in good grade with subarachnoid haemorrhage verified by computerized tomography scan or lumbar puncture. Safety evaluation includes clinical observation, monitoring of laboratory variables, positron emission tomography (PET) scanning (rCBF, rOEF, rCMRO2) and transcranial Doppler ultrasound. To date, ten patients have been recruited [NovoSeven 80 microg/kg single bolus (n = 2), NovoSeven 80 microg/kg single bolus followed by continuous infusion at 3.5 microg/kg per h (n = 2) or 7 microg/kg per h (n = 1), or control (n = 5)]. Clinical observation, transcranial Doppler ultrasound and PET studies revealed no evidence of cerebral ischaemia in the first nine patients treated with NovoSeven. The last patient developed middle cerebral artery branch thrombosis contralateral to the aneurysm. The study is currently suspended pending further investigation. PMID- 10850577 TI - Effect of the administration of recombinant activated factor VII (rFVIIa; NovoSeven) in the management of severe uncontrolled bleeding in patients undergoing heart valve replacement surgery. AB - Recombinant activated factor VII (rFVIIa; NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) is being increasingly used to secure haemostasis in difficult clinical situations. The role of rFVIIa in the treatment of patients undergoing open-heart surgery for valvular heart disease was evaluated in an open pilot study. Study objectives included evaluation of blood loss, haemostatic effect and safety and laboratory parameters following rFVIIa administration. To date, we have treated five patients (one child aged 2.5 years and four adults) undergoing surgical procedures including arterial switch, closure of atrial septal defect and De Vega's procedure (mitral valve replacement with tricuspid valve repair). Four patients received rFVIIa intraoperatively, while the fifth received it postoperatively. Satisfactory haemostasis was achieved with a single dose (30 microg/kg) of rFVIIa. Four hours after treatment mean blood loss was 262.5 ml for adults (220-334 ml) and 85 ml for the child. No significant adverse events were reported. Laboratory parameters indicated a mean 18.5-fold (range 3.7-42) increase in FVII levels at 30 min postinjection and a mean reduction of 12 s (range 3-39 s) in prothrombin time. In conclusion, rFVIIa represents an effective and well-tolerated treatment for serious bleeding episodes both during cardiac surgery and postoperatively. PMID- 10850578 TI - The effect of the administration of recombinant activated factor VII (NovoSeven) on perioperative blood loss in patients undergoing transabdominal retropubic prostatectomy: the PROSE study. AB - Transabdominal retropubic prostatectomy is associated with significant perioperative blood loss, often requiring blood transfusion. However, the administration of allogeneic blood and blood products may induce serious immunological or infectious complications. Several studies show that recombinant activated factor VII (rFVIIa; NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) induces short-term local hemostasis. This ongoing study will evaluate the safety and efficacy of rFVIIa on blood loss in patients with normal coagulation undergoing retropubic prostatectomy. Thirty-six patients will be randomized to three different dose levels and receive either rFVIIa as a single intravenous bolus dose or saline. Perioperative blood loss will be assessed from blood-volume suction containers and drains. Blood sample analysis, physical examination and electrocardiography will be performed postoperatively. Eighteen patients have enrolled in the study. Blood loss was 630-4455 ml (mean = 1698 ml), while the number of red cell transfusions varied between 0 and 4 units (mean = 0.9 units). None of the patients developed venous thromboembolism. An independent committee performed an interim analysis after patient 12 and identified a positive trend between treatment groups (not statistically significant). Although a single bolus injection of rFVIIa appears to decrease perioperative blood loss safely and effectively, definite conclusions must await study completion. PMID- 10850579 TI - Influence of activated factor VII concentrates on thrombin generation in full term and preterm neonates. PMID- 10850580 TI - Active site-blocked activated factor VII as an effective antithrombotic agent: mechanism of action. AB - The tissue factor (TF) coagulation pathway is initiated when circulating factor (F)VII(a) encounters TF, a cell surface glycoprotein, as a result of vascular injury or pathological perturbation. TF-induced coagulation plays a primary role in hemostasis and also in the pathogenesis of various thrombotic disorders. Recent studies suggest that activation of the TF-pathway may also contribute to other pathophysiological processes by altering intracellular responses, either directly or via activated factor X (FXa) and thrombin generation. Therefore, suppression of the aberrant expression of TF/FVIIa on cell surfaces not only prevents thrombotic disorders but may also provide other protective effects. Recent ex-vivo and in-vivo experiments document the effectiveness of active site blocked activated factor VII (FVIIai) in inhibiting TF-mediated injury. It is generally believed that FVIIai exerts its effects by limiting the formation of functional TF/FVIIa complexes by directly competing with plasma FVII(a) for limited available TF sites on cell surfaces. Although such competition can explain the effectiveness of FVIIai immediately after administration, it is not clear how it exerts its prolonged effects. In this manuscript, we summarize the use of FVIIai as an antithrombotic agent in various model systems and discuss potential mechanisms by which FVIIai may exert protective effects. PMID- 10850581 TI - Effect of locally-applied active site-blocked activated factor VII (ASIS) on experimental arterial thrombosis. AB - The starting point of blood coagulation in vivo is the formation of a complex between tissue factor (TF), which is exposed following vascular disease or trauma, and activated blood coagulation factor VII (FVIIa). This blinded, random, paired study evaluates whether active site-blocked activated FVII (FVIIai, ASIS), which binds avidly to TF but is unable to initiate the coagulation processes, inhibits experimental thrombosis. Arteriotomy and deep vessel wall trauma were performed in the central arteries of rabbits' ears. The topical administration of ASIS (0.5 mg in 200 microl vehicle) resulted in a distinct antithrombotic effect compared to vehicle alone. Patency rates at 30 and 120 min after reperfusion were 85% and 75% in the ASIS group and 45% and 30% in the vehicle group, respectively (P = 0.008 and P = 0.004). In contrast, intravenous administration of ASIS (4 mg/kg) produced no antithrombotic effect. Arteriotomy bleeding times were 1.5 min in the ASIS group and 2.0 min in the vehicle group (medians, P = 1). Local application of ASIS produces a pronounced antithrombotic effect in rabbits without giving rise to antihaemostatic side-effects. This mode of treatment may have a potential for a variety of clinical interventions in injured or diseased vessels. PMID- 10850582 TI - Effects of recombinant active site-blocked activated factor VII in rabbit models of carotid stenosis and myocardial infarction. AB - We tested the effects of human recombinant active site-blocked activated factor VII (rFVIIai) in a rabbit model of carotid artery thrombosis. Cyclic flow variations (CFVs), due to recurrent thrombus formation, were obtained in stenotic rabbit carotid arteries with endothelial injury. After 30 min of CFV, the animals received rFVIIai. If CFVs were abolished, animals were observed for 30 additional minutes, after which human recombinant activated factor VII was infused into the carotid artery to determine whether it could displace rFVIIai from tissue factor (TF), thus restoring CFV. An additional group of animals received rFVIIai to determine its duration of action. Recombinant FVIIai abolished CFVs in 8 of 9 rabbits (P < 0.01). This effect was reversible, as rFVIIa administration restored CFVs in all animals. A further study was initiated to assess whether TF-dependent reductions in coronary blood flow might contribute to the occurrence of myocardial injury during postischaemic reperfusion of rabbit hearts. Recombinant FVIIai resulted in significant reductions in both infarct size and no-reflow area, while rFVIIa produced a significant increase in both infarct size and no reflow area. These data suggest that rFVIIai might be beneficial in patients with acute myocardial infarction undergoing reperfusion therapies. PMID- 10850583 TI - Comparison of the factor VII:C clot analysis and a modified activated factor VII analysis for monitoring factor VII activity in patients treated with recombinant activated factor VII (NovoSeven). AB - Bleeding episodes in haemophilia A and B inhibitor patients are now frequently treated with recombinant activated factor VII (NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark). Until now, the FVII:C coagulation assay has been used to monitor NovoSeven-mediated coagulation. However, a new assay (Staclot VIIa-rTF, Diagnostica Stago, France) has been designed to specifically detect activated factor (F)VII. Replacement of the buffer supplied by the manufacturer with a PIPES buffer containing BSA (modified FVIIa assay), resulted in a linear standard curve, greater sample stability and a reduced coefficient of variation. The FVII:C assay and the modified FVIIa assay were compared in a recovery experiment using the International FVIIa standard No 89/688(IS). Recovery of FVIIa was 93 97% for the modified FVIIa assay and 91-115% for the FVII:C assay. However, because samples in the FVII:C assay were not parallel to the standard curve, confidence limits for recovery were as wide as 67-130% compared with 92-106% for the FVIIa assay. In conclusion, a modified version of the Staclot VIIa-rTF assay, suitable for monitoring treatment with NovoSeven, even at low concentrations, has been developed. It provides an alternative to the FVII:C assay, which is not suitable for monitoring FVIIa at low concentrations. PMID- 10850584 TI - Procalcitonin, a new indicator for non-viral infections in heart, lung or liver transplant patients. AB - PCT is a highly specific analyse which shows significant diagnostic validities when non-viral infections are compared with rejection episodes. PCT discriminates inflammatory events such as rejection or viral infections and non-viral infections including bacterial, fungal and protozoal infections. The half-life of PCT is 24 h indicating a clearly competent antibiotic treatment. PCT provides vital information early to clinicians and allows them to improve the management of bacterial/fungal infection in immunocompromised transplant patients. PCT thus facilitates and improves the outcome of survival rate and the quality of life in the postoperative period of patients with heart, lung or liver grafts. PMID- 10850585 TI - Preemptive therapy in CMV-antigen positive patients after liver transplantation- a prospective trial. AB - OBJECTIVE: Preemptive therapy with intravenous ganciclovir and CMV hyperimmunoglobulin in asymptomatic CMV pp65-antigen positive patients was compared with treatment of only symptomatic CMV-disease after liver transplantation in an open prospective study. PATIENTS AND METHODS: 48 out of 200 liver transplant recipients became positive during six weeks follow-up after transplantation. 17 out of these 48 patients who were already symptomatic at the time of positive antigen testing were successfully treated with ganciclovir and CMV-hyperimmunoglobulin. 31 asymptomatic antigen-positive patients were randomised to receive preemptive therapy or to receive therapy only at onset of clinical symptoms. RESULTS: Only two out of 15 patients in this latter group without preemptive therapy developed CMV-syndrome and were successfully treated with intravenous ganciclovir. 13 patients did not experience any clinical symptoms or disease and were therefore spared unneccessary toxicity and costs. The overall incidence of CMV-infection and -disease in the whole study population of 200 liver transplant recipients was 25% and 10%. As expected, CMV-negative patients who received an organ from a seropositive donor were at a higher risk of CMV-infection and -disease, but did not show more severe infections clinically. Patients with IL-2 receptor antibody induction therapy seemed to have a higher risk for CMV-infection and -disease. PMID- 10850586 TI - Factors affecting reactivation of Epstein-Barr virus infection after kidney allograft transplantation. AB - OBJECTIVES: Reactivation of Epstein-Barr virus (EBV) infection in renal transplant recipients may cause significant morbidity and mortality. To evaluate factors associated with activation of EBV replication we followed prospectively a group of 65 recipients of cadaveric kidney for 12 months. METHODS: Sera were collected periodically from these patients and analyzed for the presence of specific anti-EBV antibodies. Control group consisted of renal (n=35) and healthy blood donors (n=35). Enzyme-linked immunoassays based on recombinant EBV proteins were used to detect the following antibody specificities: early antigen (EA) IgA, IgM, and IgG, nuclear antigen (EBNA) IgG. RESULTS: During first year after transplantation, primary infection developed in 4 (6.15%) recipients and reactivation occurred in 18 (27.7%) recipients. Analysis did not show the association of reactivation with type of basic immunosuppressive therapy, prophylactic or therapeutic use of anti-lymphocyte antibodies, as well as acute rejection episodes. There was a borderline association (p=0.068) between the incidence of CMV infection and EBV reactivation. CONCLUSIONS: Our data suggest casual relationship between CMV infection and EBV reactivation. PMID- 10850587 TI - Urine excretion of transforming growth factor-beta1 in chronic allograft nephropathy. AB - Transforming growth factor-beta1 (TGF-beta1) a multifunctional growth cytokine, has been implicated in the pathogenesis of chronic allograft nephropathy (chgn). The primary objective of the present study was to establish whether or not concentration of TGF-beta1 in the sera and urine of transplant patients might be regarded as a chgn diagnostic factor and chgn activity indicator. Another objective was to investigate the response of this growth factor to arterial hypertension and metabolic disorder. Examined were 34 patients with chgn (Group I), 50 patients with a stable allograft function (Group II), and 25 healthy subjects (control). Follow-up since transplantation was 76 +/- 34 months in patients of Group I and 59 +/- 36 months in patients of Group II. Both groups of patients received maintenance triple immunosuppressive therapy. In all the subjects examined, determinations were carried out for serum and urine levels of TGF-beta1 by the immunoenzymatic method. Hypertension was found in all patients of Group I and in 60% of patients of Group II; low levels of HDL cholesterol below I mM were observed in 52% of Group I patients and in 22% of Group II patients. Serum concentrations of TGF-beta1 were similar in all the subjects examined. Patients with chgn showed elevated urine excretion of TGF-beta1, as compared to patients with no graft dysfunction or to the control. Urine excretion of TGF-beta1 was noticeably higher in patients with developed interstitial tissue fibrosis (I-st degree of interstitial fibrosis 6.8 +/- 4.9ng/mg cr. vs. 13.3 +/- 4.7 ng/mg cr. in III-rd degree of fibrosis). Urinary TGF-beta1 levels were correlated with arterial blood pressure, but they had a negative correlation with the HDL cholesterol level. CONCLUSIONS: 1) In chronic renal graft rejection urine secretion of TGF-beta1 was increased; 2) Urine secretion of TGF-beta1 was associated with arterial hypertension, degree of interstitial tissue fibrosis, and progression of graft insufficiency; 3) The negative correlation between HDL level and urine secretion of TGF-beta1 (both in patients with chronic rejection and in recipients with a stable graft function) suggests the influence of dyslipidemia on the secretion of this growth factor. PMID- 10850588 TI - Short-term storage of human haematopoietic cells. Influence of air and deoxyribonuclease I. AB - The aim of this study was to optimize strategy for ex vivo short-term storage of human cord blood and bone marrow haematopoietic cells. We report that the presence of air in the vials (1/2 of their volume), in which hematopoietic cells are stored, improves the survival of clonogenic progenitors. We observed that presence of air prevented a rapid decrease in the pH of storage medium. Similarly, a beneficial effect on cell survival and recovery also had an addition of Deoxyribonuclease I (DNase I). We observed that DNase I efficiently prevented cell clumping, and moreover, did not affect the clonogenecity of the haematopoietic progenitors. Therefore containers, in which haematopoietic cells are stored, should contain enough air (source of oxygen) and the storage medium itself should be supplemented with DNase I. PMID- 10850589 TI - Perioperative single high dose ATG-Fresenius S administration as induction immunosuppressive therapy in cadaveric renal transplantation--preliminary results. AB - Monoclonal and polyclonal antilymphocyte antibodies have been used successfully in organ transplantation as induction therapy and in the treatment of acute graft rejection. Used for induction the medication is generally given for the first 7 10 days. The aim of this study was to assess the safety and efficacy of single high dose (9 mg/kg) ATG Fresenius S given perioperatively, before revascularization, to kidney allograft recipients. During last twelve months seventy six, first cadaveric kidney adult recipients were included into the study in two centers (center A-64, center B-12). All patients received triple drug immunosuppression (Neoral, steroids and Cellcept which was replaced by azathioprine after 4 months), and were randomized to receive ATG or not. The follow-up period ranged from 1 month up to 1 year. The preliminary results are very promising, the rejection rate in bolus group was significantly lower than in control. No significant side effects or serious adverse events in both groups were observed. PMID- 10850590 TI - Vocational rehabilitation following kidney transplantation. AB - Successful kidney transplantation (KT) improves significantly the quality of life as compared with hemodialysis. Vocational rehabilitation and professional activity are the vital part of general rehabilitation and play an important role in determining the quality of life. The aim of this study was to evaluate the degree of vocational rehabilitation of the patients following allogenic kidney transplantation. 114 patients were questionnaired. Evaluation of the data revealed that only 31.6% of the study group returned to work despite good quality of life and satisfactory graft function of more than 75% of the KT patients. Factors influencing re-employment included gender, age and education level. Most of the patients were re-employed within 6 months following KT. In 78.9% of the cases formal disability status was not changed after KT and remained discrepant with good general condition and graft function. Results of the study allow authors to conclude that unless vocational rehabilitation of the KT patients is not improved, the chances of return to full social life given by KT are mostly wasted. PMID- 10850591 TI - Kidney transplantation in recipients with atherosclerotic iliac vessels. AB - The influence of atherosclerotic lesions of hypogastric and iliac arteries of kidney recipients on intraoperative surgical technique and one-year graft and patient survival was investigated. Among 1553 transplanted kidneys atherosclerotic lesions which required surgical intervention were found in 201 (12.9%) recipients. Thrombendarterectomy (TEA) of hypogastric artery was performed in 142 cases with subsequent anastomosis with the renal artery. Occlusion of hypogastric artery was an indication for renal graft artery to external iliac artery anastomosis in 32 patients. Atherosclerotic changes in external and common iliac artery required TEA in 25 patients and anastomosis between renal artery and external artery was performed. Ilio-iliac bypass or Y graft simultaneously with kidney transplantation were performed in two cases. One year survival rate of allografts and patients was 88% and 93%, respectively. No grafts were removed due to kidney artery thrombosis. We conclude that hypogastric artery after TEA provides adequate blood supply to kidney graft. PMID- 10850592 TI - Ureter diverticulum of transplanted kidney. Case report. AB - OBJECTIVES: Authors present a case of successful surgical treatment of a ureter diverticulum observed 3 months after kidney transplantation. METHODS: The fluid collection was detected by ultrasound. Percutaneous drainage was performed, after that the excretion of 4 litres of fluid was observed during 12 hours. This finding clearly indicated the connection with the urinary system. This fact was proved and precisely localised by contrast filling of the lesion under fluoroscopic control. RESULTS: Resection of the ureter diverticulum was performed, and the patient recovered fully, there was no change observed in the graft function during the whole procedure. CONCLUSION: The ureter diverticulum is a rare complication after kidney transplantation: the present case is the only one observed among more then 600 kidney transplantations performed during 20 years in our centre. PMID- 10850593 TI - Kaposi's sarcoma after renal transplantation. Case reports. AB - Immunodeficiency in transplant recipients with chronic immunosuppressive treatment may have influence to developing of virus infection diseases. This publication shows a case report kidney transplant recipient which develops Kaposi's Sarcoma correlated with HLA typing. PMID- 10850594 TI - An unusual case of early pancreas graft loss. AB - Vascular thrombosis is the leading cause of nonimmunologic, technical graft loss following pancreas transplantation. An unusual case of early pancreas graft loss due to dissection of an atherosclerotic plaque -- presumably caused by clamping during implantation -- is described. PMID- 10850595 TI - Cadaveric organ donation in Israel: the facts and the perspectives. AB - OBJECTIVES: Transplantation is limited worldwide by the shortage of organs. The main reasons are a low detection rate of potential donors and a poor motivation and qualification of health care professionals to request for family authorization for organ donation. The aim of our study was divided into two parts: to evaluate the potential and effective organ donor rate in Israel, and to assess the effects of an education program on the knowledge and attitudes of health professionals in regard to organ donation. METHODS: Part 1: We collected data on all potential and effective cadaveric donors from 1991 to 1998. We compared these figures to those obtained from European or American organizations. Part 2: We conducted 7 seminars for a total of 167 health professionals from 10 hospitals. A questionnaire was completed before and after the seminar, and differences in knowledge and attitude were compared. Pearsons correlation and students t-test were used for statistical analysis. RESULTS: Potential organ donors varied from 77 to 201 per year and effective organ donors increased from 4.8 to 14.1 pmp. Multiorgan donation increased from 1991 to 1998 reaching 60% of the cases. Knowledge on brain death, legal and religious aspects, protocols and approach to the family improved significantly (p<0.001) after 2 days of an intensive workshop. CONCLUSIONS: The rate of potential and effective organ donors is low in Israel, in comparison with other developed countries. These results are explained by problems encountered at every step of transplant procurement. We believe that a model, involving experts in brain death determination, maintenance of the donor, family approach and organ procurement could reach results close to other developed countries. PMID- 10850596 TI - Hyperhomocysteinemia as a risk factor for cardiovascular diseases. The association of hyperhomocysteinemia with diabetes mellitus and renal transplant recipients. AB - Hyperhomocysteinemia has been recognised as an independent risk factor for cardiovascular, cerebrovascular and peripheral artery disease. There is strong evidence suggesting that hyperhomocysteinemia accelerates the process of atherogenesis. Possible explanations for this will be shortly reviewed. Recently a growing interest has been focused on the association of hyperhomocysteinemia with diabetes mellitus and with chronic renal disease, including renal transplant recipients. Some clinical aspects of this occurrence, such as interactions with insulin, metformin, and cyclosporine and also with some vitamins, will be described. The issue of hyperhomocysteinemia in heart transplant patients will also be mentioned. Last of all, the interaction of homocysteine concentration with some beverages will be considered. PMID- 10850597 TI - Evidence that human haematopoietic stem cells (HSC) do not reside within the CD34+KIT- cell population. AB - Contradictory reports are published concerning the c-kit receptor (KIT) expression on human haematopoietic stem cells (HHSC). Therefore, the aim of this study was to reevaluate the expression of KIT on human early haematopoietic cells, and to study the distribution of HHSC among bone marrow mononuclear KIT+ and KIT- cells. First, we found that the detection sensitivity of the KIT expression on human HEL cells as well as CD34+ depends on the type of fluorochrome employed for the immunostaining (Cy5 > PE > FITC). Based on this observation, in our strategy for isolating human HHSC we employed a Cy-5 conjugated alpha-KIT MoAbs, which stained CD34+ cells in our preliminary studies the brightest. Accordingly, we labeled human BMMNC with PE-alpha-CD34 and Cy5 alpha-KIT MoAbs and subsequently sorted various subsets of labeled cells (CD34+KIT+, CD34+KIT- and CD34-KIT-). Cells sorted by FACS were then evaluated for their ability to engraft in the immunodeficient SCID mice model. We report here that only CD34+KIT+ cells, but not CD34+KIT- or CD34-KIT- were able to establish a human-murine haematopoietic chimerism in these animals. We found that SCID mice transplanted with CD34+ KIT+ cells, possessed approximately 5-11% of mononuclear cells, which expressed human CD45 antigen 4-5 weeks after transplantation in their bone marrow and, more importantly, early human haematopoietic progenitors from the myeloid and B-lymphoid lineages. Based on these results we conclude that KIT (CD117) is a very useful marker for identifying HHSC, and that HHSC, at least in our hands, are found in the KIT+ population of CD34+ cells. PMID- 10850598 TI - The mechanism of immunological "enhancement" in allografts--distribution of alloantigen and alloantiserum in the recipient. AB - The active or passive "enhancement", by providing the recipient with donor cellular antigens and recipient antiserum against these antigens, has been known for long to prolong the survival time of organ allografts in rodents. Donor specific blood transfusions seem to prolong the graft survival in animals and man evoking a similar protective mechanism. We investigated the mechanism of the effect of administering the recipient donor cells and antibodies against these cells on prolongation of heart graft survival. AUGust rat (RTIc) cells and WIStar rat (RTIu) antibodies against AUG antigens were labeled with 51Cr and I 125, respectively. They were administered i.v. into prospective WIS recipients of AUG hearts -11 and -10 days before grafting, respectively. The distribution and level of specific radioactivity were measured in the recipient lymphoid organs on days 9, -7 and -1 as well as on days +1 and +5 after transplantation. The highest levels of radioactivity of both donor cells and host antibodies against these cells were found on they -9 in recipient spleen but not liver, bone marrow and lymph nodes. After heart or skin transplantation, the radioactivity decreased in spleen to increase in liver and bone marrow. No specific accumulation of antibodies against donor antigens was detected in the grafts. These findings point to a role of antigen-antibody complexes located preferentially in the spleen on the downregulation of in vivo response to donor transplantation antigens. A concept of "continuous alloantigens elimination" is presented in which a systemic process of shed alloantigen inhibition and degradation in concert with blocking of organ surface antigens is presented. PMID- 10850599 TI - DNA from rejecting allografts can be detected in recipient nonlymphoid tissues. AB - The main source of donor DNA in recipients of allograft are "passenger" cells. They are claimed to be responsible for the posttransplantation microchimerism and prolongation of allograft survival. We have noticed that beside of the cellular microchimerism, donor DNA can be found in the recipient tissues at the time of rejection of allograft. In this study we provide evidence for presence in the recipient of both, DNA in "passenger cells" and free DNA in tissues at terminal stage of rejection. Male BN (RTIn) rat heart or skin were transplanted to female LEW (RTII) rats followed by a vascularized bone marrow in hind-limb transplant. CsA was given in a dose of 17mg/kg b.w. for 30 days, then rats were followed until day 100 unless rejection occurred earlier. LEW blood, spleen, mesenteric node and bone marrow cells were stained with moAb OX27 specific for BN but not LEW. Genomic male DNA was isolated and amplified with SRY oligonucleotide. At day 30 and 100 cellular microchimerism was detected in blood, spleen, nodes and bone marrow cells. Donor DNA was detected in recipient skin, liver and heart extracts, beside of lymphoid organs, at the time of rejection of allograft but not when rats were maintained on CsA. Taken together, donor DNA can be detected in recipient tissues at the time of heart or skin rejection. It seems to be released from cells of rejecting grafts and not from "passenger" cells representing only a minor cellular mass compared with the graft. PMID- 10850600 TI - Activity of glutathione S-transferases in the urine of kidney transplant recipients during the first week after transplantation. AB - Glutathione S-Tranferases (GST) are the enzymes which are strictly specific for epithelial cells of the proximal and distal tubules in the kidney. These enzymes are detected in the urine when some tubular damage process is found. In healthy people urine GST is hardly detected. The goal of this study was to evaluate the release of two isoenzymes -- alpha and pi GST in the urine of kidney graft recipients during the first week after kidney transplantation, aiming to differentiate the cause of the delayed function (DF) of transplanted kidney. MATERIAL AND METHODS: 50 cadaveric kidneys were procured using standard technique with "in situ" cooling using UW solution. All kidneys were machine perfused. After preservation kidneys were transplanted to 50 ERSD patients. Standard triple drug immunosuppression was applied (steroids, CsA, Cell-Cept or Aza.). Graft function and the release of alpha and pi GST in the urine were measured 1, 3 and 7 days after transplantation. RESULTS: immediate function (IF) was found in 72% (36pts), DF in 28% (14pts). 5 of DF patients had ATN, 4 had acute rejection (REJ) and the remaining 5 had ATN and acute rejection (see table below). CONCLUSIONS: High alpha and pi GST concentrations were found in pts with DF graft function during the first 7 days after Tx. Elevated pi GST and low alpha GST in the urine indicates acute rejection. High alpha and pi GST in pts with DF should raise suspicion of graft rejection. PMID- 10850601 TI - Disorders of calcium metabolism at various times after renal transplantation. AB - OBJECTIVES: Increased parathyroid hormone (PTH) production and related defects of calcium-phosphorus metabolism could persist even after successful kidney transplantation. Much more serious long term consequences after the transplantation are bone defects caused by immunosuppressive drugs. Many authors consider steroid therapy as one of the factors that maintain this process. Our study aimed to investigate calcium-phosphorus and bone pathological features during the various post transplantation periods, using non-invasive bone research methods (bone ultrasound structurally-densitometric analysis), and also to analyse the risk of hyperparathyroidism and steroid therapy in the development of post transplantation osteopathy. METHODS: 52 patients after successful kidney transplantation were investigated. All patients were divided in three groups according to the time after transplantation. 1st group-patients in the earlier post transplantation period, up to 1 year (n = 12); 2nd group-patients in the period from 1 to 5 years after transplantation (n = 25); 3rd group-patients in later post transplantation period (more than 5 years after the transplantation, n = 15). RESULTS: 8 patients from the 1st group (66.7%), 18 patients from the 2nd group (72%) and 8 patients from the 3rd group (53.3%) had an increased level of serum creatinine. The level of corrected serum Ca was increased (p < 0,05) in the first year after the transplantation. Hypercalcaemia was noted in 5 patients (41.7%) from the 1st group, in 3 patients (12%) from the 2nd group and in 2 patients (13.3%) from the 3rd group. Urine Ca level was lower (p < 0.05) in patients with post transplantation period over 5 years. Serum iPTH level as well as the level of osteocalcin was higher in all groups. The highest iPTH and osteocalcin level (p < 0.05) were observed during the first post transplantation year, but in the later post transplantation period they had a tendency to decrease, but never reached the norm for healthy subjects even in later post transplantation period. The decreased speed of ultrasound in the trabecular bones and osteopenia were noted in 6 patients from the 1st group (50%), osteoporosis -- in 1 patient from the 1st group(8.3%). In the 2nd group 8 patients had osteopenia (32%) and 1 patient had osteoporosis (4%), and in the 3rd group 7 patients had osteopenia (46.7%) and 4 patients -- osteoporosis (26.7%). A negative correlation was noted between patient age and speed of sound in all patient populations (r = 0.39, p<0,01), both in the early post transplantation period (r = -0.67, p<0.01), and during the period 1-5 years after transplantation (r = -0.5, p <0.01). The whole patient population showed negative correlation (r = -0.28, p<0.05) between Z-score and time after the transplantation. Z-score negatively correlates with a cumulative steroid dose in all investigated patients groups(r = -0.35, p<0.02). CONCLUSIONS: Disorders of calcium metabolism and immunosuppression related bone disease are the most common complications after transplantation, especially in patients with an impaired graft function. The mild hyperparathyroidism is usually noted in these patients at various times after transplantation. We also can note hypocalciuria in the later post transplantation period in these patients, which is based on the parathyroid glands hyperfunction and on the negative effects of the steroid therapy. The cumulative steroid dose and patient age are the determining factors for the development of osteopenia in transplantation patients at the stage of 5 or more years after transplantation. PMID- 10850602 TI - Influence of ureterovesical anastomosis technique on the incidence of vesicoureteral reflux in renal transplant recipients. AB - Urological complications of allogenic kidney transplantation include vesicoureteral reflux which can result in graft threatening urinary tract infection. To prevent this complication several ureterovesical anastomosis techniques have been developed. Authors present a comparison of three different techniques: extravesical without antireflux mechanism, extravesical Witzel-Lich with antireflux mechanism and intravesical Laedbetter-Politano with antireflux mechanism. 39 patients were selected randomly from a cohort of 420 allogenic kidney recipients (follow up time 10-147 months). All patients had voiding cystography and urine culture performed. The incidence of vesicoureteral reflux varied from 13.3% to 50%, depending on the anastomosis technique. No correlation between type of anastomosis and urinary tract infection was found. PMID- 10850603 TI - Deleterious effects of telescoped bronchial anastomosis in single and bilateral lung transplantation. AB - PURPOSE: To compare complication rates of telescoped versus end-to-end bronchial anastomoses in single and bilateral lung transplantation. METHODS: One hundred and thirty adult lung transplant recipients were evaluated during a seven-year period for the presence of three types of major bronchial anastomotic complications (ischemia, dehiscence, and severe stenosis). Surgical technique, clinical course, and mortality in all patients were reviewed retrospectively. RESULTS: The three major complications, ischemia, dehiscence, and severe stenosis, were observed in 13 (32%), 10 (24%), and 13 (32%), respectively, of 41 telescoped bronchial anastomoses. In contrast, ischemia, dehiscence, and severe stenosis, occurred in 25 (19%), 14 (10%), and 11 (8%) of 135 end-to end anastomoses. These differences were statistically significant for the occurrence of dehiscence and severe stenosis (p=0.0350 and 0.0004, respectively), and not statistically significant for ischemia (p=0.0846). Five (12%) telescoped anastomoses required stent placement as compared with six (4%) end-to end anastomoses (p=0.1313). Early postoperative pneumonia was more common in the telescoped anastomosis group (57%) as compared to the end-to-end group (35%; p=0.0271). There was a trend to shorter survival in the telescoped anastomosis group (mean survival 1172+/-149 d) as compared to the end-to-end group (mean survival 1542+/-126 d), but these differences did not achieve statistical significance (p=0.2400). CONCLUSION: In single and bilateral lung transplants, telescoped anastomoses are associated with a higher incidence of bronchial anastomotic complications and postoperative pneumonia than end-to-end anastomoses. PMID- 10850604 TI - Kinetics of bone marrow repopulation in lethally irradiated rats after transplantation of vascularized bone marrow in syngeneic hind limb. AB - Grafting of the recipient with bone marrow cell suspension provides only few stromal cells and no autologous environment for cooperation between hemopoietic and stromal cells. Vascularized bone marrow grafts provide the recipient with bone marrow hemopoietic and stromal cells in their natural spatial relationship. It is expected that such a graft will resume its function soon after transplantation and almost immediately repopulate bone marrow cavities of the irradiated recipient as well as supply the sites of cytopoiesis with the necessary growth factors. The aim of the study was to investigate the process of repopulation of bone marrow cavities and lymphoid organs of irradiated recipient by bone marrow cells from syngeneic hind limb transplant. Lewis rats received total body irradiation of 8 Gy followed by orthotopic transplantation of syngeneic limb or i.v. infusion of equivalent amount of bone marrow cells. In control experiments the hind limb was shielded with lead plate during total body irradiation. Ten days after irradiation and hind limb transplantation the yield of nucleated cells from tibia was reaching values of normal animals. In rats receiving i.v. bone marrow cell infusion it was 40% of control values and in rats repopulated from shielded own hind limb it was 60% of controls. In all groups a higher percentage of early and immediate normoblasts and a reduced pool of juvenile and segmented neutrophils was observed. Thirty days after irradiation and repopulation procedures all parameters were returning to normal levels in each group. The results indicate that bone marrow cell transplantation in hind limb graft is highly effective in lethally irradiated animals in reconstituting bone marrow. PMID- 10850605 TI - Usefulness of hybridization and PCR methods in monitoring of CMV infection in renal transplant recipients. AB - The purpose of this work was to compare hybridization and PCR methods as diagnostic tests in diagnosing and monitoring CMV infection. The investigation was performed in a group of 24 renal transplant recipients treated with ATG. The results we obtained suggest that quantitative variant of hybridization is more useful in diagnosing the infection than PCR, because it enables to monitor the infection. DNA CMV level of about 60 pg/ml or the increasing level in the subsequent samples should be a sign to start antiviral therapy. PMID- 10850606 TI - Laparoscopic lymphocele drainage after renal transplantation. AB - The formation of a lymphocele in small pelvis following renal transplantation is a well-known complication. Although various non-operative methods are available, laparotomy with transperitoneal internal drainage has been the gold standard for the treatment of lymphoceles. In 1991, internal drainage of a renal transplant lymphocele was performed for the first time laparoscopically. Nine patients with symptomatic lymphoceles were treated using laparoscopic technique, between July 1995 and November 1999 in the Surgical Department of the District Hospital in Szczecin. In 8 patients, the laparoscopic approach was successful and no further therapy was required. In one case, the videoscopic procedure had to be converted to open surgery. Operative time ranged from 15 to 60 minutes. The postoperative course was uneventful in all nine cases. Laparoscopic method of treatment of a renal transplant lymphocele combines the efficiency of internal surgical drainage, with the minimal invasiveness of non-operative techniques. It reduces postoperative pain, shortens length of hospitalisation and convalescence, and has a similar recurrence rate to open surgery. PMID- 10850607 TI - The influence of the type of anaesthesia on postoperative pain after kidney transplantation. AB - The postoperative pain treatment is one of important factors of a successful outcome after kidney transplantation. Improperly controlled pain leads to agitation, tachycardia, hypertension and increases risk of respiratory complications. Many studies have demonstrated good analgetic effect of morphine delivered by the method of patient controlled analgesia (PCA). Because the intensity of postoperative pain in end-stage kidney insufficiency patients can be modified by the type of received anaesthesia, it was decided to analyze the influence of standardized general anaesthesia on postoperative morphine consumption. 140 (ASA III) patients scheduled for kidney transplantation were included. Patients were divided into four groups; group K (control)- anaesthetised with fentanyl and N2O, group 1--fentanyl, N2O plus halothane, group 2--fentanyl, N2O plus propofol, group 3--fentanyl, N2O plus isoflurane. After operation and initial loading dose, PCA infusion of morphine was started. Bolus doses were set to 30 ug/kg, and lockout interval 10 min. Our results suggest that observed greater morphine consumption after GA with the use of propofol is connected with better psychomotor functions. In that group patients were better oriented and more efficiently controlled the PCA pump and pain. PMID- 10850608 TI - Orthotopic liver transplantation with vena cava preservation and suprahepatic caval anastomosis. PMID- 10850609 TI - Hemodynamic measurement in liver transplantation. Piggyback versus conventional techniques. AB - The piggyback technique in venous outflow tract reconstruction has been proposed as an alternative to the conventional technique in liver transplantation. Maintaining caval flow during the anhepatic phase with hemodynamic stability is regarded as one of the main advantages of this method. Between November 1994 and November 1998, the piggyback technique was used in 47 patients in our Center. Hemodynamic measurements during the operation showed hyperdynamic circulation with an increase in cardiac output (9.3+/-3.5 L/min) and the calculated cardiac index (5.0+/-1.9 L/min/m2). There was a statistically significant increase in heart rate and a decrease in systolic arterial pressure, cardiac output (CO) and cardiac index (CI) during inferior vena cava clamping ( simulated conventional technique). Only a non-significant decrease in CO and CI was observed during the partial clamp on the inferior vena cava (piggyback technique). Out of those two techniques, piggyback proved to be a safer approach to venous outflow tract reconstruction from the hemodynamic point of view. PMID- 10850610 TI - Split liver transplantation (SLT). AB - OBJECTIVES: In the last three decades liver transplantation (LT) has become a standard procedure for terminal liver failure. Anyway the procedure is highly limited by the availability of donor organs. The use of segmental liver grafts from living or cadaver donors are an attractive way to increase the donor pool for LT in adults and children. METHODS: Between 1991 and April 1999 we performed 647 liver transplantations in 416 adults and 231 children. 431 OLT, 124 SLT and 92 LRLT. Commonly used segmental liver grafts are the full right graft, the full left graft, the left lateral lobe graft and the right extended graft from living or cadaver donors respectively. RESULTS: The 1-year survival of elective SLT in adults is 80.5% and in children 84.3% (SLT + LRLT). CONCLUSIONS: Splitting procedures in liver transplantation are a promising completion to whole organ transplantations. The results of split liver (SLT) and living related liver transplantations (LRLT) are comparable to whole organ transplantations. These methods are able to increase the organ pool and thus decrease the pretransplant mortality both in children and adults. PMID- 10850611 TI - Mycophenolate mofetil for prevention of liver allograft rejection: initial results of a controlled clinical trial. AB - Mycophenolate Mofetil (M MF) is a new immunosuppressive agent with proven efficacy for the prevention of kidney allograft rejection. However, only little experience is available with the use of MMF in liver transplant recipients. OBJECTIVES: In this prospective, controlled trial the efficacy and safety of MMF and Azathioprine (AZA) were compared in a Neoral based quadruple immunosuppressive regimen after orthotopic liver transplantation. METHODS: Between 12/96 and 12/98 57 adult patients were enrolled in the study at the University of Hamburg. 28 patients were randomised to MMF, 29 patients to AZA in combination with equivalent doses of lymphocyte antibodies, Neoral and methylprednisolone. RESULTS: After a median follow-up of 10+/-3.2 months patient or graft survival did not differ significantly between the MMF and AZA group. However, MMF treated patients experienced less frequently acute rejection episodes (MMF: 6/28; 21.4% versus AZA: 13/29; 44.8%) (p=0.06). Furthermore, thrombocytopenia (MMF: 6/28; 21.4% versus AZA: 14/29; 48.3%) (p<0.05) and leukopenia (MMF: 2/28; 7.1% versus AZA: 6/29; 20%) (p=0.14) were less often observed under MMF compared to AZA. The incidence of serious bacterial infections and cytomegalovirus infections was almost identical in both groups. These preliminary data suggest that after liver transplantation primary immunosuppression with MMF is advantageous over AZA with regard to safety and efficacy. PMID- 10850612 TI - Liver transplantation in the experience of the centre commencing the program. AB - From 1989 to 1999, 43 orthotopic liver transplantations (OLT) in 40 patients (3 retransplantations) were performed in our Department. The most common indications for OLT were noninflammatory, primary cholestatic liver diseases and postinflammatory liver cirrhosis. Fourty OLT's were done for elective indications, three--on emergency basis, because of fulminant liver failure. The majority of transplantations was performed with classical technique with the excision of retrohepatic vena cava and routine use of the extracorporeal veno venous bypass. Only in 4 patients the piggyback technique was used and performed without temporary portocaval anastomosis. All 3 patients transplanted for fulminant liver failure died in the perioperative period. Twenty four patients are still alive and well, the longest period of observation exceeding 5 years. PMID- 10850613 TI - Liver transplant program in Prague. Results of consecutive first 100 orthotopic liver transplantations. PMID- 10850614 TI - Xenotransplantation of the liver. PMID- 10850615 TI - Microdialysis sampling coupled to HPLC for transdermal delivery study of ondansetron hydrochloride in rats. AB - The transdermal delivery of ondansetron hydrochloride (ON) solution in propylene glycol (PG) with a widely used penetration enhancer, oleic acid (OA), was studied in rats by a microdialysis sampling technique. Dialysate samples collected from the probe were directly injected into the HPLC system without any pre-treatment and no interference occurred in the blank sample. A good linearity between the standard concentrations and peak areas within the calibration range was achieved. In vivo recovery (32.52 +/- 1.8%) of the probe was assessed with the retrodialysis method, which was used to calculate the ON concentration in the dermis. Oleic acid at the concentrations of 2% and 5% (w/v) increased the steady state delivery rate from 0.001 to 0.030 and 0.058 microg/h, respectively. OA proved to be an effective enhancer for transdermal delivery of ON in rats. PMID- 10850616 TI - A method for the determination of the hepatic enzyme activity catalyzing bile acid acyl glucuronide formation by high-performance liquid chromatography with pulsed amperometric detection. AB - A method for the determination of the activity of hepatic glucuronyltransferase catalyzing formation of bile acid 24-glucuronides using high-performance liquid chromatography (HPLC) with pulsed amperometric detection (PAD) has been developed. Bile acid 24-glucuronides were simultaneously separated on a semimicrobore column, Capcell Pak C18UG120, using 20 mM ammonium phosphate (pH 6.0)-acetonitrile (27:10 and 16:10) as the mobile phase in the stepwise gradient elution mode. A 1 M potassium hydroxide solution for the hydrolysis of the 24 glucuronides, which liberates the corresponding bile acids and glucuronic acid, was mixed with the mobile phase in a post-column mode, and the resulting eluant was heated at 90 degrees C, the 24-glucuronides being monitored using a pulsed amperometric detector; the limit of detection was 10 ng. The proposed method was applied to the determination of the hepatic enzyme activity catalyzing bile acid 24-glucuronide formation and the result exhibited the efficient 24-glucuronide formation of the monohydroxylated bile acid, lithocholic acid. PMID- 10850617 TI - High-performance liquid chromatographic determination of naltrexone in plasma of hemodialysis patients. AB - A simple, sensitive, selective and reliable reversed-phase high-performance liquid chromatographic (HPLC) method with UV detection is described for the determination of naltrexone in plasma samples. Naltrexone and the internal standard, naloxone, were isolated from plasma either with a liquid-liquid extraction method using ethyl acetate or with a solid-phase extraction method using Sep-Pack C18 cartridge before chromatography. The extracts were dried under a stream of nitrogen and the samples were reconstituted in the mobile phase, then 20 microL were injected on a Waters Symmetry C18 column (5 microm particle size, 4.6 x 150 mm). The mobile phase consisted of 0.06% triethylamine (pH 2.8) acetonitrile (92:8, v/v) pumped at 1 mL/min. The peak-area ratio versus plasma concentration was linear over the range of 10-500 ng/mL and the detection limit was less than 8 ng/mL. Quantification was by ultra-violet detection at 204 nm. The present method was applied to the determination of the plasma concentration of naltrexone in dialyzed patients. Patients (n = 8) with severe generalized pruritus received 50 mg of naltrexone orally per day for 2 weeks. The variability in the therapeutic response in treated patients required plasma concentration investigations of this opioid antagonist. PMID- 10850618 TI - Laser-induced fluorescence detector for liquid chromatography: applications to protein analysis. AB - A laser-induced fluorescence detector for liquid chromatography was developed. This detector was assessed by utilizing it in conjunction with gel filtration chromatography. Using the 488 nm line of an argon ion laser for excitation and monitoring the emitted fluorescence centering at 535 nm, the limit of detection of fluorescein was 580 fM. Bovine serum albumin labeled with fluorescein was detected at a concentration of 500 fM. PMID- 10850619 TI - Quantitative determination of Cu-thionein from human fluids with application of solid-phase extraction on covalent affinity chromatography with thiol-disulphide interchange support. AB - The aim of our investigation was to carry out quantitative isolation of Cu thionein (Cu-Th) in human body fluids (urine, plasma and breast milk) in order to determine the level of exposure to heavy metals. In the experiment covalent affinity chromatography with thiol-disulphide interchange gel (CAC-TDI) was used as a solid phase extraction (SPE) support for preconcentration of Cu-thionein (Cu Th) protein and Cu bonded with MT from water and human fluids samples. The present paper is a continuation of early experiments on the quantitation of Hg thionein (Hg-Th), Cd-thionein (Cd-Th) and Zn-thionein (Zn-Th) in human body fluids such as urine, plasma and breast milk. PMID- 10850620 TI - Detection in the liquid phase applying chemiluminescence. AB - Several chemiluminescence-based reactions are applicable to the determination of various bio-pharmaceutically important analytes, and they can be applied for monitoring chemiluminescence emission using flow injection, liquid chromatographic and capillary electrophoretic analysis, as well as for the development of chemiluminescence-based sensors or in immunoassays. As in general the emission intensity is linearly proportional to the concentration of any of the reagents, the technique allows the analysis of different species involved in the light-producing reaction, amongst which are the chemiluminescent reagent, oxidants, inhibitors, cofactors, catalysts, some fluorophore, etc. The present overview illustrates some important applications of the last decade on this rather unfamiliar luminescence technique to detectional challenges in the liquid phase. The required instrumentation is limited as no external light source is needed. Also, the technique opens perspectives for increasing detection sensitivity in miniaturized flowing streams. On the other hand, several drawbacks still limit full application, eg dependence of the emission signal upon a number of environmental factors forcing the analyst to make a compromise between separating and measuring conditions, a lack of selectivity in specific cases, the critical detection of the signal at strictly defined periods, especially in the case of sharp emission vs time profiles, and the development of detection devices in capillary electrophoresis. PMID- 10850621 TI - HPLC-OPLC-MS investigation of change of formaldehyde and its generators in human teeth of different physiological stage. AB - Taking into consideration the unquestionable intracellular occurrence of formaldehyde (HCHO) and its generators in cells of plant, animal and human organisms as well as in body fluids it was resolved to determine their levels in hard tissues of physiologically and pathologically changed teeth. The aim of the work was to determine the relationship between the level of HCHO and the levels of its generators in pathologically changed teeth, mainly carietic teeth as tooth caries is still a serious and commonly occurring problem. The occurrence of HCHO (captured as its dimedone adduct) and some of its potential generators was demonstrated in the hard tissues of healthy and pathological human teeth by means of OPLC, HPLC and MS analyses. It was established that the measurable level of HCHO was increased in the carietic teeth in comparison with healthy ones. In the case of paradontic tooth sample, a dramatic increase of HCHO was observed and at the same time the level of betaines was decreased considerably. The obtained results give a new insight into the pathology of hard tissues of teeth in strong correlation with the phases of stress syndrome. PMID- 10850622 TI - Identification of tamoxifen and metabolites in human male urine by GC/MS. AB - Tamoxifen is an antiestrogenic drug which is used in the treatment of breast cancer and nonmalignant breast disorders. It also has a stimulating effect on the secretion of hypofisar gonadotropic hormones and is generally used in the treatment of infertility. In males, tamoxifen causes an increase of endogenous production of androgenic steroids, and therefore is used by athletes. A method for identification of tamoxifen and metabolites in urine, using the gas chromatography and mass spectrometry system (GC/MS) is described. This study also reports the extraction methodology of tamoxifen and metabolites in urine samples of healthy male volunteers and the GC/MS conditions used to identify tamoxifen and its metabolites. PMID- 10850623 TI - Determination of histamine, 1-methylhistamine and N-methylhistamine by capillary electrophoresis with micelles. AB - Urinary histamine and Ngamma-methylhistamine (1-MH), a histamine metabolite, are highly correlated with histamine in plasma. Therefore, allergic reactions can be examined by determination of histamine and 1-MH in urine. We separated histamine, 1-MH and Nalpha-methylhistamine (N-MH) by capillary electrophoresis with UV detection at 210nm, using borate buffer (pH 9) containing 100 mM SDS. The absolute detection limits were 200, 100 and 50 pg for histamine, 1-MH and N-MH, respectively. To purify histamine 1-MH and N-MH in urine, a silica cartridge was used. Recovery rates of histamine, 1-MH and N-MH in physiological saline were 90.0, 91.4 and 95.4%, respectively. We measured histamine and 1-MH levels in urine from a normal female volunteer before and after a meal, and a male bronchial asthma patient. The results showed clearly that the concentrations of histamine and its metabolite rose after eating or asthma attack. N-MH was not detected in the urine. PMID- 10850624 TI - Extraction and chromatographic separation methods in pharmacokinetic studies of Stobadine--an indole-related antioxidant and free-radical scavenger. AB - This overview provides comprehensive information on the most relevant results of Stobadine preclinical disposition studies. In order to investigate pharmacokinetic processes of the drug in rats, dogs and in human volunteers, several bioanalytical assays based on radiometric, spectrofluorometric, as well as chromatographic determination methods were developed and implemented. In small laboratory animals, the drug absorption, distribution, metabolism and elimination were investigated by administering 3H-labeled Stobadine. Spectrofluorometry was used alternatively for the determination of cold/unlabeled Stobadine in extracts of biomaterials sampled from larger animal species. The chromatographic separation methods proved, however, to be the most advantageous for determining details of the drug disposition and fate in the body. PMID- 10850625 TI - High-performance liquid chromatographic determination of phenylephrine and tropicamide in human aqueous humor. AB - A high-performance liquid chromatographic method for the simultaneous determination of phenylephrine and tropicamide in human aqueous humor was developed. After centrifugation, an aliquot of the supernatant was injected onto the column and the eluent was monitored at 280 nm then 254 nm after 5 min. Separation was performed on a CN column with 0.01 M Pic B8 (octane sulfonic acid) acetonitrile (65:35, v/v) as mobile phase. The standard curves were linear in the detection range. The precision of the method (expressed by relative standard deviation) and the accuracy (mean error in per cent) were <5% for both intra- and interassays. PMID- 10850626 TI - High-performance liquid chromatography of monoamines on phenyl-bound sorbents using organic free mobile phases. AB - Tryptophan and its metabolites, 5-hydroxytryptophan, 5-hydroxytryptamine, 5 hydroxyindolacetic acid, as well as dopamine, homovanilic acid and 2,3 dihydroxyphenylacetic acid, were separated on phenyl bound silica gel using isocratic elution with phosphate buffer. The method was successfully transferred to several other phenyl HPLC columns from different manufacturers simply by adjusting the pH of the buffer. The method has been validated by the determination of the level of monoamines in rat hypothalamus. PMID- 10850627 TI - Systemic treatment with epidermal growth factor but not insulin-like growth factor I decreases the involution of the prostate in castrated rats. AB - Epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I) are strong inducers of proliferation to prostate cells cultured in serum-free medium. Accordingly we wanted to study the growth of the prostate gland in castrated rats after treatment with EGF, IGF-I and testosterone. Castrated Wistar rats were treated with growth factors (EGF 35 microg/rat per day; IGF-I 350 microg/rat per day) or testosterone (2 mg/rat per day) for 3 days either immediately after or 10 days after castration. Prostate tissue was examined by stereological and immunohistochemical techniques and by enzyme-linked immunosorbent assay (ELISA). Treatment with EGF inhibited the involution of the prostate (P < 0.05), whereas treatment with IGF-I did not affect the prostate involution as compared to castrated controls. EGF treatment significantly increased the endogenous rat EGF in the ventral prostate, but cellular proliferation was not affected. Testosterone treatment increased the weight of the prostate, by increase of all tissue components of the prostate, and significantly increased cellular proliferation. Systemic administration of EGF but not IGF-I decreased the involution of the rat prostate induced by castration. Compared with testosterone, the effects of EGF treatment on the prostate involution were moderate, and the effects of EGF were not related to cellular proliferation. PMID- 10850628 TI - Differentially expressed mRNAs in androgen-independent but not androgen-dependent Shionogi carcinoma. AB - Recently, a new and highly effective method termed suppressive subtractive hybridization (SSH) has been introduced to clone differentially expressed mRNAs. Genes expressed in androgen-independent but not in androgen-dependent tumors, and vice versa, are obviously significant to delineate the mechanisms of androgen dependency/independency of these tumors. Mouse mammary cancer (Shionogi carcinoma 115) has been extensively used to analyze the mechanism of androgen-dependent cancer growth. METHODS: We cloned androgen-independent and androgen-dependent Shionogi carcinoma-115 specific mRNAs by the SSH method. Cloned sequences were compared with known sequences using NCBI BLAST across the Internet. Two clones were positive for cDNA insert when androgen-independent cDNA was used as tester cDNA, while no clones were positive using the androgen-dependent tester cDNA. One of the former was mouse protein kinase C beta-II while the other was a new DNA sequence. Mouse protein kinase C beta-II mRNA and the new mRNA were shown to be differentially expressed by RT-PCR analysis in androgen-independent but not androgen-dependent Shionogi carcinoma. Two mRNA species differentially expressed in androgen-independent but not androgen-dependent Shionogi carcinoma were cloned by the SSH method. The significance of these mRNAs for androgen dependency/independency of Shionogi carcinoma should be explained in future studies. PMID- 10850629 TI - Establishment and characterization of renal cell carcinoma cell lines with multidrug resistance. AB - Many of the discoveries of multidrug resistance (MDR) have resulted from studies using drug-resistant cultured tumor cell lines as experimental models. To date, there has been no report on the detailed characterization of such a cell line from renal cell carcinoma (RCC). By long-term exposure of an established RCC (RCC8701) to increasing concentrations of adriamycin, we established a series of subcultures that were considerably more resistant to the cytotoxic effect of this drug. Biological morphology and cell cycles were analyzed by morphometry and flow cytometry. The chemoresistance index of cells were measured by methyl tetrazolium assay. For evaluation of the expression of MDR-related protein (MRP), mdr-1, glutathione transferase (GST-pi), and topoisomerase II mRNAs, the reverse transcription-polymerase chain reaction was used. Membranous expression of mdr-1 related p-glycoprotein was analyzed by immunofluorescence cytometry. The intracellular content of both glutathione (GSH) and glucose-6-phosphate dehydrogenase (G-6-PDH) were measured using a capillary electrophoresis method. Compared with parent cells, the resistant sublines had a slower growth rate and lower confluent density. They were smaller and mixed with giant cells in different sizes and with different numbers of nucleoli. Flow cytometric analyses showed that resistant cells had a greater percentage of cells in the G2/M phase. The resistant cells, RCC8701/ADR800, were 122 times more resistant to adriamycin and 238 times more resistant to epirubicin than the parent cells. The resistant cells also demonstrated cross-resistance to cisplatin and 5-fluorouracil. In addition to MRP, the contents of mRNA coding for mdr-1, GST-pi, and topoisomerase II in the MDR sublines were higher than in the native cell line. A higher content of cytoplasmic GSH and G-6-PDH were found in the resistant cells; however, the expression of the MDR-related membranous glycoprotein, p-glycoprotein, was not raised. The adriamycin-induced MDR sublines may be used as an experimental system for the search of a means to overcome drug resistance and elucidate possible mechanisms of acquired MDR involved in human renal cancer. PMID- 10850630 TI - Pulsed-field gel electrophoresis for the analysis of Listeria monocytogenes infection clusters after kidney transplantation. AB - Listeria monocytogenes causes a rare, life-threatening infection in recipients of transplanted organs. We used cultures of blood and cerebrospinal fluid to characterize isolates and to distinguish cases in clusters from what might have been sporadic cases. From December 1994 to November 1995, six systemic L. monocytogenes infections occurred at our renal-transplantation unit. We confirmed the clinical diagnosis with blood and cerebrospinal fluid cultures and characterized the isolates retrospectively with pulsed-field gel electrophoresis (PFGE), phage-typing, and serotyping. We also performed an environmental investigation (food, drug, and stool). We took samples after the first two L. monocytogenes infections and then after cases three and four occurred. All patients recovered completely, and no graft was lost. Four patients had identical or genetically related L. monocytogenes isolates in PFGE (type A) and serotyping (type 1/2b). The other two had PFGE type B and G. L. monocytogenes was not detected in food or drug samples from patients on the renal-transplantation ward or in stool samples from the ward staff. It was concluded that PFGE allows sporadic cases and cluster cases of L. monocytogenes infection to be distinguished. PMID- 10850631 TI - Induction of metallothionein synthesis with preservation of testicular function in rats following long term renal transplantation. AB - Metallothionein (MT), as an acute phase or stress-response protein and free radical scavenger, is related to inflammation and cellular protection from oxidative damage. In order to evaluate long-term testicular damage and the role of MT following renal transplant, nine allogenic (Fisher 344 --> Lewis) and seven isogenic (Lewis --> Lewis) renal transplants were performed and the recipient rats were followed for 140 days when allografts develop chronic transplant rejection. Testicular weight, light microscopic morphology, and lactate dehydrogenase-X enzyme activity were assessed. Testicular MT was determined by Cd heme assay, and was localized immunocytochemically using a polyclonal rabbit antibody. No differences in testis weight, morphology, or LDH-X enzyme activity were found between allograft and isograft recipients. Testicular MT level was significantly increased in the testis of allograft recipients. Testicular zinc (Zn) and copper (Cu) levels, but not iron (Fe) level, were significantly higher in testis with allograft kidney than that with isograft kidney. In addition, Cu/Zn ratio was also significantly high in the allograft group. However, the MT level did not show any significant correlation either with Cu and Zn alone or with Cu/Zn and Fe/Zn ratios. These data suggest that allogenic stimuli may induce MT synthesis in the recipient testis. The increased MT level in an allograft may offer a protective action from oxidative damage in the testis. PMID- 10850632 TI - Histologic and molecular evidence of obstructive uropathy in rats with hereditary congenital hydronephrosis. AB - Partial obstruction of the upper urinary tract, a frequent challenge for the pediatric urologist, leads to renal damage, if deobstruction is delayed. Several but sometimes unsatisfactory animal models have been developed to study this phenomenon. Obstruction created by surgical manipulation lacks adequate correlation with a developing congenital obstruction. In some animals with congenital hydronephrosis, evidence of renal obstruction is absent. A study of the renal morphology of rats with hereditary unilateral hydronephrosis has exhibited clear evidence of renal obstruction distinguishable from renal dilatation. The renal mRNA expression of renin and transforming-growth factor beta1 (TGF-beta1) was measured by a semiquantitative RT-PCR technique. In hydronephrotic kidneys, a marked loss of parenchyma, atrophy and dilation of tubuli and collecting ducts and interstitial fibrosis was observed. The mRNA expression of renin was increased significantly in comparison to controls, whereas the contralateral kidneys showed renin activity below control levels. TGF beta1 expression was markedly increased in hydronephrotic kidneys, whereas contralateral kidneys did not differ significantly from control values. These data suggest the presence of renal obstruction and not only renal dilatation in these rats with congenital hydronephrosis. This colony seems to be a representative animal model to study congenital renal obstruction even in the fetal period without the need of surgical manipulation. PMID- 10850633 TI - Relaxation of human ureteral smooth muscle in vitro by modulation of cyclic nucleotide-dependent pathways. AB - Phosphodiesterases (PDE) are key enzymes regulating intracellular cyclic nucleotide turnover and, thus, smooth muscle tension. Recent reports have indicated the presence of PDE isoenzymes 1, 2, 4, and 5 in cytosolic supernatants prepared from human ureteral smooth muscle homogenates and the ability of second generation inhibitors of PDE 3, 4, and 5 to relax KCl-induced tension of human ureteral muscle in vitro. The aim of the present study was to evaluate the functional effects of recently developed, third-generation isoenzyme-selective PDE inhibitors, the nitric oxide (NO)-donating agents sodium nitroprusside (SNP) and dihydropyridine (DHP), which is also described as an antagonist of L-type calcium channels, and the adenylyl cyclase-stimulating drug forskolin on tissue tension and cyclic nucleotide levels of human ureteral smooth muscle segments in vitro. Relaxant responses of human ureteral smooth muscle were investigated in vitro using the organ bath technique. Cyclic nucleotides cAMP and cGMP were determined by specific radioimmunoassay following time and dose-dependent incubation of the ureteral tissue with the drugs. The most pronounced relaxing effects on KCl-induced tension of ureteral smooth muscle were exerted by nitrovasodilator SNP, PDE4 inhibitor rolipram, and PDE5 inhibitors E 4021 and morpholinosulfonyl-pyrazolopyrimidine (MSPP). Relaxing potency of the drugs was paralleled by their ability to elevate intracellular levels of cGMP and cAMP, respectively. Our data suggest the possibility of using selective inhibitors of PDE isoenzymes 4 and 5 in the treatment of ureteral stones and ureteral colic. PMID- 10850634 TI - Doxazosin modifies serotonin-mediated rabbit urinary bladder contraction. Potential clinical relevance. AB - 5-Hydroxytryptamine (5-HT) induces rabbit detrusor contractions via 5-HT3 receptors. Similarly, 5-HT4 receptors are known to be present in the human bladder. Doxazosin, a non-selective alpha1 antagonist, is used for the symptomatic relief of bladder outflow obstruction. Previous work has shown that doxazosin inhibits 5-HT2-mediated platelet shape change. Hence, the aim of this study was to assess, using organ baths and autoradiography, whether doxazosin has any 5-HT-inhibiting activity in the rabbit detrusor. Detrusor strips from adult New Zealand White rabbits were placed in organ baths; phenoxybenzamine (10(-5) M) was added to block alpha-receptors. After KCl responses were assessed, the tissues were exposed to 10(-3) M 5-HT. Subsequently, the strips were incubated with doxazosin or ondansetron (10(-5) M; 5-HT3 antagonist) followed by a further exposure to 5-HT. In some experiments, after the initial 5-HT-induced contractions, the tissues were washed and then re-exposed to 5-HT. These latter experiments acted as controls. Low-resolution autoradiography was performed on detrusor sections to assess the effect of doxazosin on 5-HT binding. These sections were analyzed densitometrically. Doxazosin and ondansetron produced a significant reduction in 5-HT-mediated contractions. Inhibition by doxazosin was in a concentration-dependent manner. Autoradiography demonstrated a significant reduction in [3H]-5-HT binding by doxazosin. Doxazosin significantly inhibits 5 HT-mediated contractions in the rabbit detrusor. This effect appears to be mainly mediated via 5-HT3 receptor inhibition. Autoradiographic evidence suggests that doxazosin reduces 5-HT binding in the rabbit detrusor. The beneficial effects of doxazosin in bladder outflow obstruction may be due, at least in part, to 5-HT antagonism. PMID- 10850635 TI - Regulation of renal artery smooth muscle tone by alpha1-adrenoceptors: role of voltage-gated calcium channels and intracellular calcium stores. AB - The ischemia induced vasospasm of the renal arterial blood vessels mediated by alpha1-adrenoceptors is of importance for the loss of kidney function. This is based on reduced perfusion of the kidney cortex occurring in kidney transplant and organ preserving surgery. The present study considered the intracellular mechanism of the norepinephrine (NE) induced renal artery vasospasm by using swine renal artery smooth muscle ring. Norepinephrine and phenylephrine (PE) induced dose-dependent and fully reversible isometric contractions with a threshold concentration of 10 nM (n = 7) and 10 nM (n = 4), and an EC50 of 0.3 microM and 1 microM, respectively. The receptor was identified as alpha1A subtype. The contraction was completely inhibited by verapamil (IC50 = 1.51 microM; n = 11) and diltiazem (IC50 = 9.49 microM; n = 8) and 85% by nifedipine (IC50 = 0.13 microM; n = 21). Blockade of the intracellular inositol- 1,4,5 trisphosphate (IP3)-sensitive Ca2+ store by thapsigargin (1 microM, n = 7) or suppression of Ca2+ release from the intracellular Ca2+-sensitive Ca2+ store by ryanodine (100 microM, n = 4) inhibited the PE induced contraction by 39.5% and 47.6%, respectively. The results suggest a key role of voltage-dependent Ca2+ channels and intracellular Ca2+ stores in the alpha1A-adrenoceptor induced contraction of the renal artery. PMID- 10850636 TI - Factors causing differences in voiding parameters between conventional and ambulatory urodynamics. AB - Voiding parameter values measured with ambulatory urodynamic monitoring (AM) are generally found to be different from those measured with conventional cystometry (CMG). The reason for this is unclear, but might be related to differences in the voided volume. To verify this hypothesis, we compared voidings from female patients at an initial bladder volume that was close to the modal volume (that is, the volume most often voided by the patient as derived from frequency/volume charts) with voidings at maximum cystometric capacity during a routine video urodynamic examination. A first group of 35 patients voided at the modal volume before they did at capacity. The order was reversed in a second group of 12 patients. The dependence of the voiding parameters on the voided volume and the order of the measurements were examined. It was found that the maximum flow rate depended significantly on the voided volume, but the associated detrusor pressure did not. Urethral resistance and bladder contraction strength were not volume dependent either. It was concluded that the differences between AM and CMG cannot be explained from possible differences in the voided volume. PMID- 10850637 TI - Failure of short-term gamma-linolenic acid treatment to reduce urinary calcium loss of diabetic rats. AB - Calcium re-absorption in the kidney is impaired in streptozotocin (STZ) diabetic rats, thereby causing hypercalciuria. Increased calcium loss starts within 1-2 days after induction of diabetes and reaches a plateau after 2 weeks. The excessive calcium excretion was previously shown to be reduced by treatment with gamma-linolenic acid (GLA) or evening primrose oil rich in GLA. However, in these studies, the animals were pre-treated for several weeks before injection of STZ. In the present study we investigated whether GLA can reduce calcium excretion when treatment starts at the same time as induction of diabetes. Rats were made diabetic with 60 mg/kg STZ and at the same time food was fortified with 0.4% GLA for the treatment group. A control group was treated with vehicle alone and given standard feed only. Urine was collected from animals in metabolism cages every 3rd day for a period of 26 days. The diabetic group increased their food and water consumption, and urine and faeces production as compared to the control group. The urinary loss of Ca, Mg, Zn, Na, K and creatinine was markedly increased in the diabetic group as compared to the control. GLA treatment, however, did not affect any of these variables. Analysis of fatty acids in kidneys of the rats showed an increased concentration of GLA in the treated group as compared to the two non-treated groups. We conclude that GLA treatment must commence before STZ injection in order to attenuate diabetes-induced hypercalciuria. PMID- 10850638 TI - Effects of phytate and pyrophosphate on brushite and hydroxyapatite crystallization. Comparison with the action of other polyphosphates. AB - This is a comparative study of the effects of phytate and pyrophosphate and other polyphosphates on the crystallization of hydroxyapatite and brushite, the most frequent calcium phosphates involved in calcium oxalate urolithiasis. Brushite and hydroxyapatite crystal formation was studied in synthetic urine, through kinetic-turbidimetric measurements that allowed evaluation of the inhibitory effects on crystallization of insoluble salts. The effectiveness in preventing brushite crystallization decreases in the sequence phytate > polyphosphate > EDTPO > etidronate > pyrophosphate > triphosphate > medronate; whereas the order of effectiveness in preventing hydroxyapatite crystallization was EDTPO > etidronate = pyrophosphate > triphosphate > medronate > polyphosphate > phytate. Phytate, a natural inhibitor in urine, most effectively blocked brushite precipitation (1.21x10(-5) M prevented crystallization during time periods of at least 1 h), and pyrophosphate was the natural inhibitor that most effectively blocked hydroxyapatite precipitation (2.87x10(-6) M prevented crystallization during time periods of at least 1 h). This demonstrates that low excretion of these substances would pose a risk of renal lithiasis. PMID- 10850639 TI - Real-time monitoring of nitric oxide in ischemia-reperfusion rat kidney. AB - In this study we attempted to clarify the release of nitric oxide (NO) and its role in the ischemia-reperfusion rat kidney. After right nephrectomy, male Wistar rats were divided into four groups: one sham operated and three groups who underwent ischemia (30 min) and reperfusion of the left renal artery. Thirty minutes prior to ischemia-reperfusion, two groups were injected intraperitoneally with 10 and 30 mg/kg of NG-nitro-L-arginine methylester (L-NAME). Real-time monitoring of blood flow and NO release in the rat kidney was measured with a laser Doppler flowmeter and an NO-selective electrode, respectively. Serum creatinine and blood urea nitrogen (BUN) levels were measured 1 and 7 days after the induction of ischemia-reperfusion. Clamping of the renal artery decreased blood flow to 1-5% of the basal level measured before clamping. After removal of the clip, the blood flow of the 30 mg/kg L-NAME rats was significantly lower than that of the controls. Immediately following the clipping of the renal artery, NO release rapidly increased. After removing the clip, NO release immediately returned to three-quarters of the basal level. Serum creatinine and BUN levels of the ischemia-reperfusion rats were slightly but not significantly higher and those of 30 mg L-NAME rats were significantly higher than those of the control or ischemia-reperfusion rats 1 day and 7 days after ischemia-reperfusion. Our data suggest that NO acts as a cytoprotective agent in ischemia-reperfusion injury of the rat kidney. PMID- 10850640 TI - Nitric oxide synthase in adult red blood cells: vestige of an earlier age or a biologically active enzyme? PMID- 10850641 TI - Cloning of human stem cells: some broad scientific and philosophical issues. PMID- 10850642 TI - Current concepts and controversies in the pathogenesis, prevention, and treatment of the macrovascular complications of diabetes. PMID- 10850643 TI - Normal circulating adult human red blood cells contain inactive NOS proteins. AB - Human red blood cells (RBCs) are considered to play a significant role in both blood pressure (BP) regulation and tissue oxygenation, because they can bind as well as release previously bound nitric oxide (NO) from hemoglobin (Hb) and other intracellular components. Two reports indicate that the human RBC possesses nitric oxide synthase (NOS) activity-by the accumulation of nitrite across a membraned chamber in one and by the hydrolysis of labeled L-arginine, presumably to labeled L-citrulline, in the other. Furthermore, NOS proteins have been identified by immunoblot in RBCs. If true, the presence of NOS activity would convert the human RBC to a donor with limited ability to bind exogenously generated NO. In considering the importance of the question of the presence or not of NO synthetic capacity of this cell in BP regulation and tissue perfusion, whether human RBCs are, indeed, able to hydrolyze L-arginine to L-citrulline, the coproduct of NO was explored. RBC samples collected from control subjects were assayed for NOS activity by incubation of homogenized cellular fractions with labeled tritiated L-arginine in the presence of 0.5 mmol/L NADPH. By this method, the amino acid coproduct of NO, tritiated L-citrulline, would be recovered in the supernatant after removal of unused substrate by cationic resin treatment. At first, activity appeared to be present in the RBC supernatant but not in the pellet. However, activity was not suppressed by known inhibitors of NOS, whereas activity was suppressed by norvaline, an inhibitor of arginase activity with no known effect on NOS. By contrast, RBC arginase activity was not inhibited by N(omega)-nitro-L-arginine, NG-methyl-L-arginine, or aminoguanidine, known inhibitors of NOS, but was inhibited by norvaline. The label recovered by thin layer chromatography was determined not to be tritiated L-citrulline but was instead tritiated L-ornithine, the product of arginase activity. Thus the enzymatic hydrolysis of L-arginine was not caused by NOS but was a result of the action of the enzyme arginase, which abounds in this cell. However, proteins interacting with antibodies to the endothelial and inducible isoforms of NOS were detected in human RBCs by immunoblot. Together, these findings indicate that human RBCs collected from normal adult individuals possess proteins that react with monoclonal antibodies to the Inducible and endothelial isoforms of NOS, but the proteins are without catalytic activity. PMID- 10850644 TI - Effects of erythropoietin therapy on iron absorption in chronic renal failure. AB - The effect of erythropoietin administration on the absorption of dietary and therapeutic iron was examined in patients with anemia of chronic renal failure on maintenance hemodialysis. Absorption from test meals tagged extrinsically with iron 55, iron 59, or both was determined 2 weeks later by using incorporated red blood cell radioactivity and whole body counting. In an initial study of food iron absorption, the effect of initiating erythropoietin therapy was determined by measuring the absorption of heme and nonheme iron before and 2 weeks after the administration of 64 U/kg body weight erythropoietin (range, 46-85 U/kg body weight) three times weekly. Absorption of heme iron increased 1.6-fold from 18.6% to 30.1% (P < .05), and nonheme iron increased 3.7-fold from 1.3% to 4.9% (P < .01) after erythropoietin therapy. In a second study therapeutic iron absorption was evaluated at baseline and after erythropoietin administration (63 U/kg body weight (range, 48-74 U/kg body weight) three times weekly). The absorption of 50 mg of iron as ferrous sulfate increased 2.4-fold from 3.8% to 9.4% (P < .05) when given without food and 4.2-fold from 1.4% to 5.9% (P < .05) when given with food after erythropoietin administration. After adjusting for changes in iron stores with serum ferritin after erythropoietin therapy, the enhanced erythropoiesis associated with erythropoietin therapy increased absorption about 2-fold, which was similar to the response observed previously in normal subjects. In a final study we examined the absorption of therapeutic iron during the steadystate phase of erythropoietin therapy after an erythroid response to erythropoietin had occurred. The absorption of 50 mg of iron was lower than that occurring with the initiation of erythropoietin therapy at 2.2% when given alone and 1.3% when taken with food. We conclude that iron absorption with or without erythropoietin stimulation is unimpaired in patients with chronic renal failure. PMID- 10850645 TI - Appearance of dissociable and cross-linked hemoglobins in the renal hilar lymph. AB - Unlike unmodified dissociable bovine hemoglobin (UHb), cross-linked hemoglobins do not dissociate into dimers, do not cross the glomerular filter, and are retained in the plasma for a longer time. Renal peritubular capillaries, which are different from the glomerulus, allow the passage of molecules as large as albumin into the renal interstitium. Cross-linked hemoglobins should pass across these capillaries, enter the renal interstitium, and drain through the renal lymphatics. The present experiments were done in anesthetized rats to determine the appearance of UHb, an intramolecularly cross-linked tetrameric hemoglobin (DECHb), and a polymerized bovine hemoglobin (PHb) of larger molecular size into the renal hilar lymph. Renal hilar lymph samples were obtained before and after an isovolemic exchange of 2 mL/100 g rat weight of a 6% solution of each hemoglobin for blood. The behavior of a 5% solution of Evan's blue-labeled albumin was also determined for comparison. After exchange, the Initial plasma concentration of each of the proteins was in excess of 20 mg/mL. UHb appeared both in urine and lymph. DECHb, PHb, and albumin were absent from the urine but appeared promptly in the renal hilar lymph and reached concentrations at least 30% that of plasma. PHb had a significantly smaller lymph clearance (in microliters per minute) and longer plasma half-time than the other nondissociable proteins. These findings indicate that DECHb and PHb, although not filtered, pass across peritubular capillaries and readily enter the renal interstitial space. The passage of the larger molecular-sized PHb may be hindered relative to the other proteins in passage across peritubular and other systemic capillaries. PMID- 10850646 TI - Effects of geranyl-geranyl-acetone administration before heat shock preconditioning for conferring tolerance against ischemia-reperfusion injury in rat livers. AB - The effect of geranyl-geranyl-acetone (GGA) administration before heat shock preconditioning on heat shock protein (HSP) 72 induction and on the acquisition of tolerance against ischemia-reperfusion Injury was studied in rat livers. Male Wistar rats were divided into four groups: a control group (group C); a GGA group (group G); a simple heat shock group (group VH); and a heat shock with GGA premedication group (group GH). Five-, 10-, and 15-minute periods of heat shock preconditioning at 42 degrees C were performed in groups VH and GH. Subgroups were determined according to the period of heat shock exposure. After a 48-hour recovery, rats in groups C, VH5, VH15, and GH5 received a 30-minute period of hepatic ischemia. Induction of HSP72, survival rates, and changes in biochemical and histologic parameters were compared among the groups. Five-minute heat shock preconditioning was not enough to Induce HSP72. However, livers in group GH5 expressed approximately the same amount of HSP72 as those in group VH15. The expression of HSP72 in group GH15 was stronger than that found in group VH15. The degree and location of HSP72 expression were not different between groups GH5 and VH15. Seven-day survival was significantly better in groups GH5 (16/16) and VH15 (15/16) than in group C (8/16) or VH5 (9/16). The recovery of adenosine triphosphate in liver tissue was faster, and the release of liver-related enzymes during reperfusion was lower in groups GH5 and VH15 than in group C or VH5. Administration of GGA before heat shock preconditioning augmented the induction of HSP72 by decreasing the threshold for triggering the stress response. PMID- 10850647 TI - Preservation of intestinal microvascular Po2 during normovolemic hemodilution in a rat model. AB - The effect of hemodilution on the intestinal microcirculatory oxygenation is not clear. The aim of this study was to determine the effect of moderate normovolemic hemodilution on intestinal microvascular partial oxygen pressure (Po2) and its relation to the mesenteric venous Po2 (Pmvo2). Normovolemic hemodilution was performed in 13 anesthetized male Wistar rats. Systemic hemodynamic and intestinal oxygenation parameters were monitored. Intestinal microvascular Po2 was measured by using the oxygen-dependent quenching of palladium-porphyrin phosphorescence. Hemodilution decreased systemic hematocrit from 45.0% +/- 0.1% (average +/- SEM) to 24.6% +/- 1.6%. The mesenteric blood flow did not change from baseline values, resulting in a linear decrease in intestinal oxygen delivery (from 2.77 +/- 0.15 to 1.42 +/- 0.11 mLxkg(-1)xmin(-1)). The intestinal oxygen extraction ratio increased significantly from 24% +/- 1% to 42% +/- 4%. Pmvo2 decreased significantly (from 57 +/- 2 to 41 +/- 2 mm Hg), but intestinal oxygen consumption and microvascular Po2 remained unaffected. As a result, the difference between microvascular Po2 and Pmvo2 increased significantly during hemodilution. Intestinal microvascular Po2 and oxygen consumption were well preserved during moderate normovolemic hemodilution. These results might be explained by the notion of others that hemodilution induces recruitment of capillaries, resulting in redistribution of the intestinal blood flow in favor of the microcirculation, which allows a more efficient extraction of oxygen. These findings further indicate that the use of venous Po2 values as indicators of microvascular oxygenation may be misleading. PMID- 10850648 TI - Delivery of recombinant product from subcutaneous implants of encapsulated recombinant cells in canines. AB - Delivering recombinant therapeutic proteins from a universal microencapsulated cell line is an alternate method for gene therapy. It has proved effective in the treatment of several murine models of human genetic diseases. However, in scaling up to large animal models, intraperitoneal Implantations of these microcapsules in canines were associated with excessive Inflammatory response and rapid degradation. We now show that subcutaneous implantation of microencapsulated cells in canines is effective in delivering recombinant product systemically for extended periods, provides a surgically benign site, leads to less inflammatory response, and permits longer-term survival of microcapsules. Allogeneic MDCK cells engineered to secrete human growth hormone (hGH) were microencapsulated in alginate-poly-L-lysine-alginate and implanted subcutaneously. Systemic delivery of hGH was evident within 4 hours and peaked by day 1 after implantation in all dogs. The gradual decline of hGH in the circulation in the first 2 weeks coincided with the development of anti-hGH antibodies by day 11. The high titer persisted for more than 1 month, demonstrating indirectly the persistent delivery of hGH. Microcapsules retrieved from the subcutaneous implant maintained their structure throughout the experiment and were free of host cellular adhesions. The mechanical integrity of the subcutaneously implanted microcapsules also appeared superior to that of the intraperitoneal implant. Hence the subcutaneously implanted microcapsules required minimal surgical intervention and led to a low level of inflammatory response, and the implant survived for at least 1 month, thus demonstrating the feasibility of systemic delivery of recombinant products via subcutaneous implantation in large animals. PMID- 10850649 TI - Dietary chloride does not correlate with urinary thromboxane in deoxycorticosterone acetate-treated rats. AB - Renal vascular resistance in deoxycorticosterone acetate (DOCA) salt-treated uninephrectomized rats is increased by high dietary chloride. Because DOCA salt hypertensive rats exhibit an increased urinary excretion of thromboxane B2 (TXB2), a metabolite of thromboxane A2 (TXA2), the increased TXB2 excretion by DOCA salt-treated rats could relate to elevated dietary chloride, increased blood pressure, and/or the presence of intact renal tubules. We hypothesized that high NaCl intake, resulting in an elevated tubular chloride excretion, stimulates TXA2 production. A result of that production could be renal vasoconstriction. Baseline blood pressures were measured for 10 days, and then the rats were treated with DOCA (30 mg/kg) and fed (1) normal NaCl, (2) normal sodium with high chloride, or (3) high sodium chloride (NaCl) for 4.5 weeks. Next, the rats were uninephrectomized (1K) or unihydronephrectomized (1KHK) to yield one kidney without an intact tubular system and therefore no macula densa. Two and a half weeks later, urinary excretion of TXB2 was determined. DOCA-high NaCl-fed 1KHK or 1K rats had significant increases in systemic blood pressure to 172 +/- 12 and 190 +/- 5 mm Hg, respectively, compared with no significant increase in blood pressure among the other groups. Urinary TXB2 excretion was increased to 29 +/- 4 pg per 24 hours per gram of body weight in all DOCA-treated 1KHK and 1K animals regardless of diet compared with DOCA-treated animals with two intact kidneys (13 +/- 2 pg per 24 hours per gram of body weight). DOCA treatment in rats with one functional kidney results in the excretion of high levels of urinary TXB2 unrelated to dietary chloride load, blood pressure, or intact renal tubules. PMID- 10850650 TI - Prometheus. PMID- 10850651 TI - 4-1BB: still in the midst of darkness. AB - 4-1BB is a member of the tumor necrosis factor receptor superfamily. The receptor functions mainly as a costimulatory molecule in T lymphocytes. In addition, several lines of evidence have shown that interactions between 4-1BB and its ligand are involved in the antigen presentation process and the generation of cytotoxic T cells. Recent studies, however, have demonstrated that 4-1BB plays more diverse roles: Signals through 4-1BB are important for long-term survival of CD8+ T cells and the induction of helper T cell anergy. Clinically, there is great interest in 4-1BB, because T-cell activation induced by anti-4-1BB monoclonal antibodies is highly efficient in the eradication of established tumor cells in mice. Now, since mice deficient in 4-1BB or the 4-1BB ligand are available, subtle roles played by 4-1BB may be revealed in the near future. PMID- 10850652 TI - Isolation of TC/AG repeat microsatellite sequences for fingerprinting rice blast fungus and their possible horizontal transfer to plant species. AB - Genome fingerprinting has been a major role in characterization of population structure and analysis of the variability in phytopathogenic fungi. In order to characterize Korean rice blast fungal isolates, the genomic DNAs were digested with AluI endonuclease and subsequent PCR amplifications using random decamer primers with combinations of microsatellite primers had been carried out. This Alu-Inter SSR technique revealed high polymorphism among the Korean blast fungal isolates. Then, fragments from the Alu-Inter SSR analysis were isolated to be used as probes in Southern hybridization, which also revealed high polymorphism between isolates to distinguish individuals. The sequences of the isolated fragments contained TC/AG tandem repeats interspersed with a 30 bp direct repeat. In gel blot analysis, the isolated TC/AG repeat microsatellite sequences were proved to be useful for characterizing the isolates in blast fungi in addition to the conventional MGR (Magnaporthe grisea repeat) probes. One interesting point was that the rice blast fungus derived TC/AG repeat microsatellite sequences were abundant in non-rice blast fungi and plant species, but not in other fungi and yeasts. A discussion on the possible horizontal gene transfer between phytopathogenic fungi and host plants is presented. PMID- 10850653 TI - Isolation and characterization of cDNAs encoding ribosome inactivating protein from Dianthus sinensis L. AB - To isolate a ribosome inactivating protein (RIP) gene, six plant species were surveyed for antiviral activity. Crude proteins extracted from these plants were tested for the antiviral activity against tobacco mosaic virus (TMV) in Nicotiana glutinosa. All the plants, Spinacia oleracea, Amaranthus lividus, Dianthus superbus, Dianthus sinensis and Celosia cristata, with an exception of Oenanthe stolonifera, presented 70-90% inhibition of viral infectivity. In an attempt to search for the RIP gene from D. sinensis, partial cDNA was obtained by polymerase chain reaction (PCR) of the poly(A)+ RNA from D. sinensis leaves. DNA gel blot analysis showed that D. sinensis has multi-copy RIP genes. The expression of RIP gene was investigated in the flower, leaf, root and stem of D. sinensis, and was found to be most abundant in the leaf. Using the partial cDNA as a probe, seven full-length cDNAs were isolated from a library prepared from D. sinensis leaves. They were divided into three groups on the basis of their nucleotide sequence homology. The three representative clones, cDsRIP1, cDsRIP2 and cDsRIP3 were completely sequenced. They all had an open reading frame of 882 bp. The cDsRIP2 showed 79% homology with dianthin 30 and saporin genes; 59% with PAP and Mirabilis antiviral protein MAP genes. From the analysis of deduced amino acid sequences, it was predicted that D. sinensis RIP cDNAs might have a putative signal peptide of 23 amino acid residues at their N-terminus. When the cDNA was expressed in E. coli, the bacteria was unable to grow upon IPTG induction, suggesting that expression of the gene renders toxicity to E. coli cells. PMID- 10850654 TI - In vivo gene transfer to the mouse nasal cavity mucosa using a stable cationic lipid emulsion. AB - We evaluate a new cationic emulsion as a mucosal gene carrier and elucidate the relationship between the transfection efficiency and the stability of the carrier/DNA complex. A cationic lipid emulsion was formulated with soybean oil and 1,2-dioleoyl-sn-glycero-3-trimethylammonium-propane (DOTAP) as major components and was used to transfer genes to the epithelial cells of the mouse nasal cavity via intranasal instillation. Correlation between the transfection efficiency and the stability of the carrier/DNA complex was investigated by measuring the carrier size changes and by observing the degree of DNA protection against DNase I digestion in the presence of heparin. The cationic emulsion showed at least 3 times better transfection activity than the liposomal carriers in nasal mucosae. The cationic emulsion was stable in the presence of heparin whereas the liposomal carriers became very unstable. Unlike DNA in liposome/DNA complexes, DNA in the emulsion/DNA complex was resistant to heparin exchange and DNase I digestion. The cationic emulsion was more effective in delivering DNA to nasal mucosae than commercially available liposomal carriers. The transfection activities of the lipid carriers in nasal cavity mucosae are in agreement with the stability of the lipid carriers and their complexes with DNA. PMID- 10850655 TI - Molecular cloning, expression, and purification of nuclear inclusion A protease from tobacco vein mottling virus. AB - The gene encoding the C-terminal protease domain of the nuclear inclusion protein a (NIa) of tobacco vein mottling virus (TVMV) was cloned from an isolated virus particle and expressed as a fusion protein with glutathione S-transferase in Escherichia coli XL1-blue. The 27-kDa protease was purified from the fusion protein by glutathione affinity chromatography and Mono S chromatography. The purified protease exhibited the specific proteolytic activity towards the nonapeptide substrates, Ac-Glu-Asn-Asn-Val-Arg-Phe-Gln-Ser-Leu-amide and Ac-Arg Glu-Thr-Val-Arg-Phe-Gln-Ser-Asp-amide, containing the junction sequences between P3 protein and cylindrical inclusion protein and between nuclear inclusion protein b and capsid protein, respectively. The Km and k(cat) values were about 0.2 mM and 0.071 s(-1), respectively, which were approximately five-fold lower than those obtained for the NIa protease of turnip mosaic potyvirus (TuMV), suggesting that the TVMV NIa protease is different in the binding affinity as well as in the catalytic power from the TuMV NIa protease. In contrast to the NIa proteases from TuMV and tobacco etch virus, the TVMV NIa protease was not autocatalytically cleaved into smaller proteins, indicating that the C-terminal truncation is not a common phenomenon occurring in all potyviral NIa proteases. These results suggest that the TVMV NIa protease has a unique biochemical property distinct from those of other potyviral proteases. PMID- 10850656 TI - Effects of Polycomb group mutations on the expression of Ultrabithorax in the Drosophila visceral mesoderm. AB - The Polycomb group (PcG) genes encode repressors of many developmental regulatory genes including homeotic genes and are known to act by modifying chromatin structure through complex formation. We describe how Ultrabithorax (Ubx) expression is affected by the PcG mutants in the visceral mesoderm. Mutant embryos of the genes extra sex combs (esc), Polycomb (Pc), additional sex combs (Asx) and pleiohomeotic (pho) were examined. In each mutation, Ubx was ectopically expressed outside of their normal domains along the anterior posterior axis in the visceral mesoderm, which is consistent with the effect of PcG proteins repressing the homeotic genes in other tissues. All of these four PcG mutations exhibit complete or partial lack of midgut constriction. However, two thirds of esc mutant embryos did not show Ubx expression in parasegment 7 (PS7). Even in the embryos showing ectopic Ubx expression, the level of Ubx expression in the PcG mutations was weaker than that in normal embryos. We suggest that in PcG mutations the ectopic Ubx expression is caused by lack of PcG repressor proteins, while the weaker or lack of Ubx expression is due to the repression of Ubx by Abd-B protein which is ectopically expressed in PcG mutations as well. PMID- 10850657 TI - Retinoic acid and its geometrical isomers block both growth and fusion of L6 myoblasts by modulating the expression of protein kinase A. AB - All-trans retinoic acid (RA) and its geometrical isomers, such as 9-cis RA, 13 cis RA, and 9,13-di-cis RA, strongly inhibited both growth and fusion of L6 myoblasts. However, illumination of white light diminished their inhibitory activity on membrane fusion with little effect on cell growth. During myogenic differentiation, the intracellular level of cAMP decreased whereas the total activity of protein kinase A as well as the protein level of its regulatory subunit Ialpha (RIalpha) and catalytic subunit (Calpha) increased. RAs raised the intracellular level of cAMP by over 3-fold, but decreased the total activity of protein kinase A. Like RAs, dibutyryl-cAMP inhibited myoblast fusion and reduced the expression of both RIalpha and Calpha subunits. These results suggest that RAs negatively modulate the differentiation of L6 myoblasts by increasing the intracellular level of cAMP, which may in turn down-regulate the expression of protein kinase A and hence its activity. PMID- 10850658 TI - Sustained formation of focal adhesions with paxillin in morphological differentiation of PC12 cells. AB - Differentiation of PC12 cells triggered by nerve growth factor (NGF) is characterized by several well-defined events including induction of a set of neuron-specific genes, gain of membrane excitability, and morphological changes such as neurite outgrowth. Here we report that K252a, a protein kinase inhibitor, converts the proliferation signal of epidermal growth factor (EGF) into the morphological differentiation signal without inducing the sustained activation of ERK and the expression of neurofilament. Major effects of EGF/K252a, found also in the NGF-treated cells, are the sustained mobility shift of paxillin in SDS PAGE and the promoted association of Crk-II with paxillin. These effects explain the prominent and robust development of peripheral focal adhesion assembly and stress fiber-like structures observed in the early stages of PC12 cell differentiation. These results suggest a model that cytoskeletal reorganization via focal adhesion assembly triggered by NGF provides a signal required for the morphological differentiation of PC12 cells. PMID- 10850659 TI - Regulation of Mst57Dc expression in male accessory glands of Drosophila melanogaster. AB - Mst57Dc has been isolated as a male accessory gland transcript of Drosophila melanogaster. Its product is a secretory protein, which is phosphorylated by protein kinase A. In the present study, the expression pattern of Mst57Dc was analyzed. It is preferentially expressed in but not restricted to the male accessory glands. Other than in the accessory glands, it is slightly expressed in other body parts, including the head and female body. In the accessory glands, a high level of expression was detected right after eclosion when the titer of juvenile hormone III (JHIII) reaches a peak. Its accumulation was increased by mating, which has been known to act via JH. In ap56f a JH-deficient mutant, the level of Mst57Dc transcripts was about 60% of the wild type. Moreover a JH responsive element like palindromic sequence and several sequence motifs were found in the 5' and 3' flanking regions of Mst57Dc. Taken together, JH is proposed as a regulator of Mst57Dc gene expression. PMID- 10850660 TI - Immunological detection of serpin in the fall webworm, Hyphantria cunea and its inhibitory activity on the prophenoloxidase system. AB - We previously identified a serine type protease inhibitor (serpin) cDNA, using PCR-based differential display, in the fall webworm which was up-regulated following a bacterial challenge (Shin et al., 1998). The serpin cDNA was inserted into an expression vector and the serpin protein was expressed in Escherichia coli. In order to investigate the action of serpin in vivo, we examined the concentration of serpin protein in the larvae of Hyphantria cunea by Western blot analysis using a polyclonal antibody raised in a rabbit injected with recombinant serpin. H. cunea serpin was found mainly in the plasma with a molecular mass of 56.6 kDa on SDS-PAGE followed by Western blot analysis. The concentration of serpin in the plasma was slightly increased following bacterial challenge. A new 50.5 kDa (approx.) band was detected post E. coli and distilled water injection. Both E. coli and distilled water injection induced increased phenoloxidase (PO) activity in the plasma, although E. coli injection produced a larger increase in activity. Hyphantria serpin probably participates in negative regulation of the prophenoloxidase (proPO) cascade. Recombinant serpin inhibits PO activity in the hemocyte lysate fraction activated by LPS. There is a similarity between the P2 P2' region (NKFG) of the serpin reactive site loop and the S2-S2' region (NRFG) of the insect proPO maturation site. This indicates a form of competitive inhibition of serpin against a protease involved in the activation of proPO. A tyrosine residue in the P11 region of serpin, which is conserved in the S11 regions of all known proPOs maturation sites, provides further support for this hypothesis. PMID- 10850661 TI - Antisense oligonucleotide of clusterin mRNA induces apoptotic cell death and prevents adhesion of rat ASC-17D Sertoli cells. AB - Clusterin has been known to play important roles in cell-cell and/or cell substratum interactions. Recently we reported the transient expression of clusterin in pancreatic endocrine cells during the early developmental stages and suggested a role in aggregating the endocrine cells for islet formation. In the present study, we have investigated the involvement of clusterin in cell substratum interaction by the inhibition of clusterin synthesis using antisense oligonucleotide. The expression of clusterin was transiently increased as early as 2-8 h after plating the ASC-17D Sertoli cells to the culture flask, which was the period of cell attachment. In addition, up-regulation of clusterin mRNA was so much greater when the Sertoli cells were plated on the petri dish for the bacterial culture instead of in a animal cell culture flask that therefore, the cells failed to attach to it. These findings suggested that interruption of cell to plate substratum interaction might lead to over-expression of clusterin from Sertoli cells to induce cell to cell aggregation or, perhaps, to re-establish attachment with the substratum. Transfection of ASC-17D Sertoli cells with a 20 base antisense oligonucleotide against clusterin mRNA resulted in extracellular release of LDH and DNA fragmentation. Sertoli cell death by antisense oligonucleotide of clusterin was sequence specific and dose dependent. Treatment of antisense oligonucleotide induced a marked reduction of synthesis for clusterin protein, but not for clusterin mRNA expression, suggesting the translational suppression of clusterin by antisense oligonucleotide. Further, microscopic observation showed that more noticeable cell death was induced by treating the antisense prior to plating the cells than by treating after cell attachment to the plate. From these results, we speculate that down-regulation of clusterin expression in the anchorage-dependent Sertoli cells prevents them from attaching to the plate, and therefore induces cell death. PMID- 10850662 TI - Physiological and genetic comparison of two aromatic hydrocarbon-degrading Sphingomonas strains. AB - Sphingomonas yanoikuyae strain B1 is able to degrade a wider range of aromatic hydrocarbons than S. paucimobilis strain TNE12 can degrade. Various culture techniques were used to corroborate that B1 used m-xylene, biphenyl, toluene, naphthalene, and phenanthrene as sole carbon and energy sources. In contrast, TNE12 could not mineralize m-xylene, biphenyl, toluene, or naphthalene. However, fluoranthene served as carbon and energy source for TNE12 but not B1. Southern blots were performed using the cloned genomic region (approximately 23 kb) containing the degradative genes for the upstream pathways for biphenyl and m xylene and a TOL plasmid-type meta operon from B1 as a probe against the KpnI restriction-digested total DNA of TNE12. This 23 kb probe hybridized to three KpnI-digested fragments of TNE12 DNA; thus significant homology existed between the aromatic hydrocarbon-degrading genes of B1 and TNE12. Further work with smaller probes revealed, however, that TNE12 DNA fragments did not hybridize with the probe containing the genes encoding for xylene monooxygenase and part of an aromatic dioxygenase. A recombinant plasmid, which contains only the genes for xylene monooxygenase, is able to complement TNE12 on m-xylene. These genes are, therefore, probably missing from TNE12. Hence, TNE12 cannot use monocyclic aromatics whereas B1 can. Pulsed field gel electrophoresis coupled with Southern blotting revealed that the aromatic degradative genes were on an approximately 240 kb plasmid of TNE12; the same genes in B1 are known to be chromosomal. PMID- 10850663 TI - Calcium-binding proteins calbindin D28K, calretinin, and parvalbumin immunoreactivity in the rabbit visual cortex. AB - The distribution and morphology of neurons containing three calcium-binding proteins, calbindin D28K, calretinin, and parvalbumin in the adult rabbit visual cortex were studied. The calcium-binding proteins were identified using antibody immunocytochemistry. Calbindin D28K-immunoreactive (IR) neurons were located throughout the cortical layers with the highest density in layer V. However, calbindin D28K-IR neurons were rarely encountered in layer I. Calretinin-IR neurons were mainly located in layers II and III. Considerably lower densities of calretinin-IR neurons were observed in the other layers. Parvalbumin-IR neurons were predominantly located in layers III, IV, V, and VI. In layers I and II, parvalbumin-IR neurons were only rarely seen. The majority of the calbindin D28K IR neurons were stellate, round or oval cells with multipolar dendrites. The majority of calretinin-IR neurons were vertical fusiform cells with long processes traveling perpendicularly to the pial surface. The morphology of the majority of parvalbumin-IR neurons was similar to that of calbindin D28K: stellate, round or oval with multipolar dendrites. These results indicate that these three different calcium-binding proteins are contained in specific layers and cells in the rabbit visual cortex. PMID- 10850664 TI - Proteolytic processing of oligopeptides containing the target sequences by the recombinant tobacco vein mottling virus NIa proteinase. AB - Tobacco vein mottling virus (TVMV) belongs to the potyviridae that consists of about 200 plant viruses. Potyviruses have RNA genomes of approximately 10,000 bases from which a single polyprotein is expressed from each virus upon infection. The NIa proteinase is known to process the polyprotein at seven distinct junctions between proteins. Kinetic constants were determined for the reactions of the recombinant TVMV NIa protease (27 kDa) with synthetic oligopeptides containing the sequences for the cleavage sites. For optimum activity, the substrate needs to have six amino acids (P6-P1) in the amino region and four (P1'-P4') in the carboxy region, including four conserved amino acids (V R-F-Q) in P4-P1 positions. Mutation of any of four conserved amino acids to Gly made the substrate inert to the enzyme. Among the substrates, the oligopeptides containing the sequences for junctions, P3-6K1, NIa (VPg-Pro), and NIa-NIb were not processed by the NIa protease. Those junctions have Glu at P3, Glu at P1, and Thr at P2. The implications of high substrate specificity and size dependence in polyprotein processing and viral replication are discussed. PMID- 10850665 TI - Cloning of a sesquiterpene cyclase and its functional expression by domain swapping strategy. AB - Sesquiterpene cyclase, the first committed step enzyme from the general isoprenoid building block farnesyl pyrophosphate (FPP) for the synthesis of phytoalexin capsidiol, was isolated from the UV-C treated leaves of Capsicum annuum. This sesquiterpene cyclase, termed as CASC2 showing 77% amino acid identity with the previously cloned sesquiterpene cyclase CASC1, was composed of 560 amino acids with a calculated molecular mass of 64,907. The mRNA expression pattern of CASC2 was very similar to that of CASC1 during the time course of UV-C irradiated leaves of pepper on RNA blot analysis by using each specific probe. The heterologous expression in Escherichia coli using the CASC2 full length failed; however the chimeric construct of CASC2 in which the amino terminal 164 amino acid substituted by the equivalent portion of either CASC1 or tobacco sesquiterpene cyclase was capable of expressing the functional sesquiterpene cyclase activities. The radio-labeled enzymatic products catalyzed by the partially purified chimeric CASC2 were comigrated with authentic radio-labeled sesquiterpene on thin layer chromatography. PMID- 10850666 TI - Analysis of the fit-1 gene encoding a ECM protein, flectin, in Caenorhabditis elegans. AB - Flectin is a new type of extracellular matrix protein and its function was suggested to provide a microenvironment of great elasticity. The C. elegans genome database revealed the presence of a flectin homologue, fit-1, which shows approximately 40% similarity (20% identity) to chick flectin. Here we propose a new gene structure for the fit-1 based on our experiments and the partial cDNA clones obtained from Y. Kohara and further suggest that the previous gene prediction is incorrect. FLT-1 is shown to be expressed in various neurons, hypodermal cells, distal tip cells and vulva epithelial cells. Immunostaining results with anti-FLT-1 antibody, further confirm the FLT-1 expression in vulva epithelial cells. The lipophilic dye, DiI, was used to identify the head neurons expressing GFP and results indicated that none of the head neurons expressing GFP are the 6 chemosensory neurons. In order to determine the function of flt-1 gene, RNA-mediated interference (RNAi) experiments were conducted. PMID- 10850667 TI - Random changes of amino acid residues with expected frequency by saturated point mutagenesis. AB - The yeast transcriptional activator protein, Gcn4p from Saccharomyces cerevisiae binds to the specific sequence in the promoters of many amino acid biosynthetic genes for general control. A new random saturation mutagenesis method was developed to isolate Gcn4p derivatives with only one or two mutations in the DNA binding domain without using radioactive isotope. This will be used to identify the amino acids of Gcn4p involved in protein-protein interactions. Saturation mutagenesis in the DNA binding domain of Gcn4p was performed using spiked degenerate oligonucleotides containing randomized codon bases designed specifically for only one or two base changes in the mutagenized area. These oligonucleotides were synthesized to have two flanking restriction enzyme sites for cloning to the appropriate vector. The 3' ends were mutually primed after hybridization via the palindromic sequences of the restriction enzyme sites. These molecules were then converted to double stranded DNA upon treatment with DNA polymerase. Here, a library collection of 100,680 in an altered Gcn4p pool was generated by cloning a mixed-base oligonucleotide in the place of the sequence coding for the DNA binding domains. The quality of the library was examined by DNA sequencing and found to be in good agreement with the expected statistical values. Calculated mutation frequency was 66% of mutant nucleotide rate and actual sequencing data revealed 68% mutant nucleotide rates from the sequenced library. Thus, among 21 mutants, 16 had one point mutations and 5 had two point mutations. This approach appears to be an effective and general tool for creating proteins with one or two amino acid change(s) in their molecules. PMID- 10850668 TI - A novel technique for the effective production of short peptide analogs from concatameric short peptide multimers. AB - We designed a basic unit of the modified chicken gonadotropin releasing hormone II (cGnRH-II) peptide containing a trypsin cleavable linker peptide at both ends of the original peptide. We made a synthetic DNA coding for the modified cGnRH-II peptide with asymmetric and complementary cohesive ends of linker nucleotides. A tandemly repeated DNA cassette for the expression of concatameric short peptide multimers was constructed by ligating the basic units. The expressed peptide multimers were purified and subject to amino-terminal sequence analysis, which displayed the amino acid sequences expected from the designed nucleotides of the expression cassette. The monomeric cGnRH-II peptide analogs were generated after trypsin digestion. The present results showed that the technique developed for the production of the concatameric peptide multimers with cleavable linker peptides can be generally applicable to the production of short peptide analogs. PMID- 10850669 TI - Expression and high-level secretion of Trichoderma reesei endoglucanase I in Yarrowia lipolytica. AB - The endoglucanase I (EGI) from fungus Trichoderma reesei was cloned, expressed, and secreted from Yarrowia lipolytica using the XPR2 promoter. The signal sequence of EGI transferred from T. reesei was efficiently processed in the Y. lipolytica secretory pathway and directed the secretion of active EGI into the culture medium. However, the recombinant EGI produced from YLCSIn strain was hyperglycosylated and significantly larger than the native enzyme produced by the parent strain. The expression of EGI using XPR2 preproregion has caused secretion of modified proteins that still retained cellulase activity. This resulted from imprecise processing of the N-terminus of recombinant protein. While the batch culture produced 5 mg EGI/L from YLCSIn strain, the EGI yield was increased approx 20-fold when the fed-batch fermentation process strategy in combination with the high-cell density cultivation technique was employed. These results showed that the Y. lipolytica is a useful host organism for production of a large amount of large size heterologous proteins, especially when used in combination with high-cell density and fed-batch culture techniques. PMID- 10850670 TI - Isolation and characterization of a novel feather-degrading bacterial strain. AB - Feather waste, generated in large quantities as a byproduct of commercial poultry processing, is almost pure keratin, which is not easily degradable by common proteolytic enzymes. Feather-degrading bacteria were isolated from a Brazilian poultry industrial waste. Among these isolates, a strain identified as kr2 was the best feather-degrading organism when grown on basal medium containing 10 g/L of native feather as a source of energy, carbon, and nitrogen. The isolate was characterized according to morphological characteristics and biochemical tests belonging to the Vibrionaceae family. Keratinolytic activity of this isolate was monitored throughout the cultivation of the bacterium on raw feather at different temperatures. The optimum temperature for growth was about 30 degrees C, at which maximum enzyme and soluble protein production were achieved. The enzyme had a pH and temperature optima of 8.0 and 55 degrees C, respectively. PMID- 10850671 TI - A sensitive solid-phase fluoroimmunoassay for detection of opiates in urine. AB - An automated flow fluorometer designed for kinetic binding analysis was adapted to develop a solid-phase competitive fluoroimmunoassay for urinalysis of opiates. The solid phase consisted of polymer beads coated with commercial monoclonal antibodies (MAbs) raised against morphine. Fluorescein-conjugated morphine (FL MOR) was used as the fluorescein-labeled hapten. The dissociation equilibrium constant (K(D)) for the binding of FL-MOR to the anti-MOR MAb was 0.23 nM. The binding of FL-MOR to the anti-MOR MAb reached steady state within minutes and was displaced effectively by morphine and other opiates. Morphine-3-glucuronide (M3G), the major urinary metabolite of heroin and morphine, competed effectively with FL-MOR in a concentration-dependent manner for binding to the antimorphine MAb and was therefore used to construct the calibration curve. The sensitivity of the assay was 0.2 ng/mL for M3G. The assay was effective at concentrations of M3G from 0.2 to 50 ng/mL, with an IC50 of 2 ng/mL. Other opiates and heroin metabolites that showed >50% crossreactivity when present at 1 microg/mL included codeine, morphine-6-glucuronide, and oxycodone. Methadone showed very low crossreactivity (<5%), which is a benefit for testing in patients being treated for opiate addictions. The high sensitivity of the assay and the relatively high cutoff value for positive opiate tests allows very small sample volumes (e.g., in saliva or sweat) to be analyzed. A double-blind comparison using 205 clinical urine samples showed good agreement between this single-step competitive assay and a commercially performed enzyme multiplied immunoassay technique for the detection of opiates and benzoylecgonine (a metabolite of cocaine). PMID- 10850672 TI - Continuous production of antibiotics in an airlift fermentor utilizing a transverse magnetic field. AB - In this study, a series of experiments was conducted to demonstrate the feasibility of continuous production of penicillin antibiotic using a three-phase magneto airlift fermentor with immobilized Penicillium chrysogenum. The fermentation processes were carried out in a 2.4-L external loop airlift utilizing a transverse magnetic field. It was found that the application of the magnetic field to a bed of ferromagnetic beads affects both the hydrodynamics of the reactor and the rate of the bioconversion process occurring inside it. One hundred hours after startup, the maximum penicillin concentration increased 48% as the magnetic field intensity increased from 0 to 35 mT, owing to the increased residence time of the substrate in the riser and the positive effect of the magnetic field on the effective fluid-solid interfacial area. In addition, the detached biomass concentration in the liquid phase was found to be only 5% of the immobilized biomass, owing to low shear levels and the absence of friction among the solid-phase particles. PMID- 10850673 TI - Cellular association of glucosyltransferases in Leuconostoc mesenteroides and effects of detergent on cell association. AB - Most glucosyltransferase (GTF) activity in sucrose-grown cultures of some strains of Leuconostoc mesenteroides is found with the cell pellet after centrifugation. GTFs are known to bind to dextrans, and it was traditionally assumed that cell associated GTFs were bound to those dextrans that cosedimented with the cells. We used a mutant strain (LC-17), derived from strain NRRL B-1355, which produced dextransucrase in the absence of dextrans, to investigate the extent to which GTFs were bound to cells or dextrans. Much of the GTF activity in glucose-grown cultures of strain LC-17, which do not produce dextran, was located in the cell pellets. Soluble enzyme activity increased when cell suspensions from glucose- or sucrose-grown cultures were incubated with mild nonionic detergents or zwitterionic reagents. Alternansucrase produced by the parent strain B-1355 was almost entirely associated with cells under conditions in which dextrans were or were not produced. Alternansucrase, but not dextransucrase, tended to be enriched in the particulate fraction of B-1355 cells that had been broken in a French press. The distribution of alternansucrase and the effects of detergents on the distribution of GTFs suggest that soluble GTFs sequestered in the cytoplasm, and GTFs bound or adsorbed to the cell membrane are probably the major contributors to the cell-associated GTF activity. PMID- 10850674 TI - Recognizing hollow strengths in research communities PMID- 10850676 TI - Amphibians come under study. PMID- 10850675 TI - Researchers take US government to court over threat to turtles... PMID- 10850677 TI - Green light for plans to sell off US helium reserve PMID- 10850678 TI - UK labs impeded by old equipment PMID- 10850679 TI - Clinical trials end at gene-therapy institute... PMID- 10850681 TI - France plans marine research port at Brest PMID- 10850680 TI - Harvard keeps its ethics guidelines. PMID- 10850682 TI - Transmutation of nuclear waste branded 'Trojan horse'. PMID- 10850683 TI - Germany edges towards stem-cell accord. PMID- 10850684 TI - French scientists turn to journals in English. PMID- 10850686 TI - Requiem for an observatory PMID- 10850685 TI - World's academies seek a sustainable future. PMID- 10850687 TI - To conserve rainforest, we have to help local people live sustainably. PMID- 10850688 TI - Impact factors aren't relevant to taxonomy. PMID- 10850689 TI - We have moral authority to apologize for the past. PMID- 10850691 TI - Let Armenia show why it's the place for Sesame PMID- 10850690 TI - Offside researchers score an own goal PMID- 10850692 TI - Time for voices to be raised. PMID- 10850693 TI - A double-edged sword. PMID- 10850694 TI - Catching crumbs from the table. In the face of metahuman science, humans have become metascientists PMID- 10850695 TI - Pax Argentinica. PMID- 10850696 TI - Neutralizing noise in gene networks. PMID- 10850697 TI - Glass transition. Hard knock for thermodynamics PMID- 10850699 TI - Quantum decay. A watched pot boils quicker PMID- 10850698 TI - Cognitive neuroscience. Learning how the brain learns. PMID- 10850700 TI - Immunology. The footprint of a killer. PMID- 10850701 TI - Pacific leatherback turtles face extinction. PMID- 10850702 TI - Fossil record of mass moth migration. PMID- 10850704 TI - A massive cool dust torus around eta Carinae? PMID- 10850703 TI - Presenilin-1 mutations in Alzheimer's disease. PMID- 10850705 TI - Inferring the statistical interpretation of quantum mechanics from the classical limit AB - It is widely believed that the statistical interpretation of quantum mechanics cannot be inferred from the Schrodinger equation itself, and must be stated as an additional independent axiom. Here I propose that the situation is not so stark. For systems that have both continuous and discrete degrees of freedom (such as coordinates and spin respectively), the statistical interpretation for the discrete variables is implied by requiring that the system's gross motion can be classically described under circumstances specified by the Schrodinger equation. However, this is not a full-fledged derivation of the statistical interpretation because it does not apply to the continuous variables of classical mechanics. PMID- 10850706 TI - Crystal structure of an NK cell immunoglobulin-like receptor in complex with its class I MHC ligand. AB - Target cell lysis is regulated by natural killer (NK) cell receptors that recognize class I MHC molecules. Here we report the crystal structure of the human immunoglobulin-like NK cell receptor KIR2DL2 in complex with its class I ligand HLA-Cw3 and peptide. KIR binds in a nearly orthogonal orientation across the alpha1 and alpha2 helices of Cw3 and directly contacts positions 7 and 8 of the peptide. No significant conformational changes in KIR occur on complex formation. The receptor footprint on HLA overlaps with but is distinct from that of the T-cell receptor. Charge complementarity dominates the KIR/HLA interface and mutations that disrupt interface salt bridges substantially diminish binding. Most contacts in the complex are between KIR and conserved HLA-C residues, but a hydrogen bond between Lys 44 of KIR2DL2 and Asn 80 of Cw3 confers the allotype specificity. KIR contact requires position 8 of the peptide to be a residue smaller than valine. A second KIR/HLA interface produced an ordered receptor ligand aggregation in the crystal which may resemble receptor clustering during immune synapse formation. PMID- 10850707 TI - Rossby waves on the Sun as revealed by solar 'hills' AB - It is a long-standing puzzle that the Sun's photosphere--its visible surface- rotates differentially, with the equatorial regions rotating faster than the poles. It has been suggested that waves analogous to terrestrial Rossby waves, and known as r-mode oscillations, could explain the Sun's differential rotation: Rossby waves are seen in the oceans as large-scale (hundreds of kilometres) variations of sea-surface height (5-cm-high waves), which propagate slowly either east or west (they could take tens of years to cross the Pacific Ocean). Calculations show that the solar r-mode oscillations have properties that should be strongly constrained by differential rotation. Here we report the detection of 100-m-high 'hills' in the photosphere, spaced uniformly over the Sun's surface with a spacing of (8.7 +/- 0.6) x 10(4) km. If convection under the photosphere is organized by the r-modes, the observed corrugated photosphere is a probable surface manifestation of these solar oscillations. PMID- 10850708 TI - Acceleration of quantum decay processes by frequent observations AB - In theory, the decay of any unstable quantum state can be inhibited by sufficiently frequent measurements--the quantum Zeno effect. Although this prediction has been tested only for transitions between two coupled, essentially stable states, the quantum Zeno effect is thought to be a general feature of quantum mechanics, applicable to radioactive or radiative decay processes. This generality arises from the assumption that, in principle, successive observations can be made at time intervals too short for the system to change appreciably. Here we show not only that the quantum Zeno effect is fundamentally unattainable in radiative or radioactive decay (because the required measurement rates would cause the system to disintegrate), but also that these processes may be accelerated by frequent measurements. We find that the modification of the decay process is determined by the energy spread incurred by the measurements (as a result of the time-energy uncertainty relation), and the distribution of states to which the decaying state is coupled. Whereas the inhibitory quantum Zeno effect may be feasible in a limited class of systems, the opposite effect- accelerated decay--appears to be much more ubiquitous. PMID- 10850709 TI - Absence of thermodynamic phase transition in a model glass former AB - The glass transition can be viewed simply as the point at which the viscosity of a structurally disordered liquid reaches a universal threshold value. But this is an operational definition that circumvents fundamental issues, such as whether the glass transition is a purely dynamical phenomenon. If so, ergodicity gets broken (the system becomes confined to some part of its phase space), but the thermodynamic properties of the liquid remain unchanged across the transition, provided they are determined as thermodynamic equilibrium averages over the whole phase space. The opposite view claims that an underlying thermodynamic phase transition is responsible for the pronounced slow-down in the dynamics at the liquid-glass boundary. Such a phase transition would trigger the dynamic standstill, and then be masked by it. Here we perform Monte Carlo simulations of a two-dimensional system of polydisperse hard disks far within its glassy phase. The approach allows for non-local moves in a way that preserves micro reversibility. We find no evidence for a thermodynamic phase transition up to very high densities; the glass is thus indistinguishable from the liquid on purely thermodynamic grounds. PMID- 10850710 TI - Dynamics of the silicon (111) surface phase transition AB - The manner in which phase transformations occur in solids determines important structural and physical properties of many materials. The main problem in characterizing the kinetic processes that occur during phase transformations is the difficulty of observing directly, in real time, the growth of one phase at the expense of another. Here we use low-energy electron microscopy to study the real-time kinetics of a phase transformation confined to the silicon (111) surface. We show that the transformation is governed by the rate at which material is exchanged between the first layer of the crystal and the surface. In bulk phase transformations, the dynamics are usually governed either by the rate of diffusion of material to the phase boundaries or by the structural rearrangement of atoms at the phase boundary. The kinetic process that we have identified here has no bulk analogue and leads to domain dynamics that are qualitatively different from those expected for bulk systems. PMID- 10850711 TI - Old radiocarbon ages in the southwest Pacific Ocean during the last glacial period and deglaciation AB - Marine radiocarbon (14C) dates are widely used for dating oceanic events and as tracers of ocean circulation, essential components for understanding ocean climate interactions. Past ocean ventilation rates have been determined by the difference between radiocarbon ages of deep-water and surface-water reservoirs, but the apparent age of surface waters (currently approximately 400 years in the tropics and approximately 1,200 years in Antarctic waters) might not be constant through time, as has been assumed in radiocarbon chronologies and palaeoclimate studies. Here we present independent estimates of surface-water and deep-water reservoir ages in the New Zealand region since the last glacial period, using volcanic ejecta (tephras) deposited in both marine and terrestrial sediments as stratigraphic markers. Compared to present-day values, surface-reservoir ages from 11,900 14C years ago were twice as large (800 years) and during glacial times were five times as large (2,000 years), contradicting the assumption of constant surface age. Furthermore, the ages of glacial deep-water reservoirs were much older (3,000-5,000 years). The increase in surface-to-deep water age differences in the glacial Southern Ocean suggests that there was decreased ocean ventilation during this period. PMID- 10850712 TI - Multiple seismic discontinuities near the base of the transition zone in the Earth's mantle AB - The seismologically defined boundary between the transition zone in the Earth's mantle (410-660 km depth) and the underlying lower mantle is generally interpreted to result from the breakdown of the gamma-spinel phase of olivine to magnesium-perovskite and magnesiowustite. Laboratory measurements of these transformations of olivine have determined that the phase boundary has a negative Clapeyron slope and does indeed occur near pressures corresponding to the base of the transition zone. But a computational study has indicated that, because of the presence of garnet minerals, multiple seismic discontinuities might exist near a depth of 660 km (ref. 4), which would alter the simple negative correlation of changes in temperature with changes in the depth of the phase boundary. In particular, garnet minerals undergo exothermic transformations near this depth, acting to complicate the phase relations and possibly effecting mantle convection processes in some regions. Here we present seismic evidence that supports the existence of such multiple transitions near a depth of 660 km beneath southern California. The observations are consistent with having been generated by garnet transformations coupling with the dissociation of the gamma-spinel phase of olivine. Temperature anomalies calculated from the imaged discontinuity depths- using Clapeyron slopes determined for the various transformations--generally match those predicted from an independent P-wave velocity model of the region. PMID- 10850713 TI - Are lemmings prey or predators? AB - Large oscillations in the populations of Norwegian lemmings have mystified both professional ecologists and lay public. Ecologists suspect that these oscillations are driven by a trophic mechanism: either an interaction between lemmings and their food supply, or an interaction between lemmings and their predators. If lemming cycles are indeed driven by a trophic interaction, can we tell whether lemmings act as the resource ('prey') or the consumer ('predator')? In trophic interaction models, peaks of resource density generally have a blunt, rounded shape, whereas peaks of consumer density are sharp and angular. Here we have applied several statistical tests to three lemming datasets and contrasted them with comparable data for cyclic voles. We find that vole peaks are blunt, consistent with their cycles being driven by the interaction with predators. In contrast, the shape of lemming peaks is consistent with the hypothesis that lemmings are functional predators, that is, their cycles are driven by their interaction with food plants. Our findings suggest that a single mechanism, such as interaction between rodents and predators, is unlikely to provide the 'universal' explanation of all cyclic rodent dynamics. PMID- 10850714 TI - Highly fecund mothers sacrifice offspring survival to maximize fitness. AB - Why do highly fecund organisms apparently sacrifice offspring size for increased numbers when offspring survival generally increases with size? The theoretical tools for understanding this evolutionary trade-off between number and size of offspring have developed over the past 25 years; however, the absence of data on the relation between offspring size and fitness in highly fecund species, which would control for potentially confounding variables, has caused such models to remain largely hypothetical. Here we manipulate egg size, controlling for maternal trait interactions, and determine the causal consequences of offspring size in a wild population of Atlantic salmon. The joint effect of egg size on egg number and offspring survival resulted in stabilizing phenotypic selection for an optimal size. The optimal egg size differed only marginally from the mean value observed in the population, suggesting that it had evolved mainly in response to selection on maternal rather than offspring fitness. We conclude that maximization of maternal fitness by sacrificing offspring survival may well be a general phenomenon among highly fecund organisms. PMID- 10850715 TI - Cortical ensemble activity increasingly predicts behaviour outcomes during learning of a motor task. AB - When an animal learns to make movements in response to different stimuli, changes in activity in the motor cortex seem to accompany and underlie this learning. The precise nature of modifications in cortical motor areas during the initial stages of motor learning, however, is largely unknown. Here we address this issue by chronically recording from neuronal ensembles located in the rat motor cortex, throughout the period required for rats to learn a reaction-time task. Motor learning was demonstrated by a decrease in the variance of the rats' reaction times and an increase in the time the animals were able to wait for a trigger stimulus. These behavioural changes were correlated with a significant increase in our ability to predict the correct or incorrect outcome of single trials based on three measures of neuronal ensemble activity: average firing rate, temporal patterns of firing, and correlated firing. This increase in prediction indicates that an association between sensory cues and movement emerged in the motor cortex as the task was learned. Such modifications in cortical ensemble activity may be critical for the initial learning of motor tasks. PMID- 10850716 TI - An electroneutral sodium/bicarbonate cotransporter NBCn1 and associated sodium channel. AB - Two electroneutral, Na+-driven HCO3- transporters, the Na+-driven Cl-/HCO3- exchanger and the electroneutral Na+/HCO3- cotransporter, have crucial roles in regulating intracellular pH in a variety of cells, including cardiac myocytes, vascular smooth-muscle, neurons and fibroblasts; however, it is difficult to distinguish their Cl- dependence in mammalian cells. Here we report the cloning of three variants of an electroneutral Na+/HCO3- cotransporter, NBCn1, from rat smooth muscle. They are 89-92% identical to a human skeletal muscle clone, 55-57% identical to the electrogenic NBCs and 33-43% identical to the anion exchangers. When expressed in Xenopus oocytes, NBCn1-B (which encodes 1,218 amino acids) is electroneutral, Na+-dependent and HCO3(-)-dependent, but not Cl(-)-dependent. Oocytes injected with low levels of NBCn1-B complementary RNA exhibit a Na+ conductance that 4,4-diisothiocyanatostilbene-2,2'-disulphonate stimulates slowly and irreversibly. PMID- 10850717 TI - Cyclin D control of growth rate in plants. AB - The mechanisms by which plants modulate their growth rate in response to environmental and developmental conditions are unknown, but are presumed to involve specialized regions called meristems where cell division is concentrated. The possible role of cell division in influencing meristem activity and overall plant growth rate is controversial, with a prevailing view that cell division is secondary to higher order meristem controls. Here we show that a reduction in the length of the cell-cycle G1 phase and faster cell cycling occur when the rate of cell division in transgenic tobacco plants is increased by the plant D-type cyclin CycD2 (ref. 8). The plants have normal cell and meristem sizes, but elevated overall growth rates, an increased rate of leaf initiation and accelerated development in all stages from seedling to maturity. We conclude that cell division is a principal determinant of meristem activity and overall growth rate, and propose that modulation of plant growth rate is achieved through regulation of G1. PMID- 10850718 TI - Peptides accelerate their uptake by activating a ubiquitin-dependent proteolytic pathway. AB - Protein degradation by the ubiquitin system controls the intracellular concentrations of many regulatory proteins. A protein substrate of the ubiquitin system is conjugated to ubiquitin through the action of three enzymes, E1, E2 and E3, with the degradation signal (degron) of the substrate recognized by E3 (refs 1-3). The resulting multi-ubiquitylated substrate is degraded by the 26S proteasome. Here we describe the physiological regulation of a ubiquitin dependent pathway through allosteric modulation of its E3 activity by small compounds. Ubr1, the E3 enzyme of the N-end rule pathway (a ubiquitin-dependent proteolytic system) in Saccharomyces cerevisiae mediates the degradation of Cup9, a transcriptional repressor of the peptide transporter Ptr2 (ref. 5). Ubr1 also targets proteins that have destabilizing amino-terminal residues. We show that the degradation of Cup9 is allosterically activated by dipeptides with destabilizing N-terminal residues. In the resulting positive feedback circuit, imported dipeptides bind to Ubr1 and accelerate the Ubr1-dependent degradation of Cup9, thereby de-repressing the expression of Ptr2 and increasing the cell's capacity to import peptides. These findings identify the physiological rationale for the targeting of Cup9 by Ubr1, and indicate that small compounds may regulate other ubiquitin-dependent pathways. PMID- 10850719 TI - Recombination signal sequences restrict chromosomal V(D)J recombination beyond the 12/23 rule. AB - The genes encoding the variable regions of lymphocyte antigen receptors are assembled from variable (V), diversity (D) and joining (J) gene segments. V(D)J recombination is initiated by the recombinase activating gene (RAG)-1 and -2 proteins, which introduce DNA double-strand breaks between the V, D and J segments and their flanking recombination signal sequences (RSSs). Generally expressed DNA repair proteins then carry out the joining reaction. The conserved heptamer and nonamer sequences of the RSSs are separated by non-conserved spacers of 12 or 23 base pairs (forming 12-RSSs and 23-RSSs). The 12/23 rule, which is mediated at the level of RAG-1/2 recognition and cutting, specifies that V(D)J recombination occurs only between a gene segment flanked by a 12-RSS and one flanked by a 23-RSS. Vbeta segments are appended to DJbeta rearrangements, with little or no direct Vbeta to Jbeta joining, despite 12/23 compatibility of Vbeta 23-RSSs and Jbeta12-RSSs. Here we use embryonic stem cells and mice with a modified T-cell receptor (TCR)beta locus containing only one Dbeta (Dbeta1) gene segment and one Jbeta (Jbeta1) gene cluster to show that the 5' Dbeta1 12-RSS, but not the Jbeta1 12-RSSs, targets rearrangement of a diverse Vbeta repertoire. This targeting is precise and position-independent. This additional restriction on V(D)J recombination has important implications for the regulation of variable region gene assembly and repertoire development. PMID- 10850720 TI - Intraprotein radical transfer during photoactivation of DNA photolyase. AB - Amino-acid radicals play key roles in many enzymatic reactions. Catalysis often involves transfer of a radical character within the protein, as in class I ribonucleotide reductase where radical transfer occurs over 35 A, from a tyrosyl radical to a cysteine. It is currently debated whether this kind of long-range transfer occurs by electron transfer, followed by proton release to create a neutral radical, or by H-atom transfer, that is, simultaneous transfer of electrons and protons. The latter mechanism avoids the energetic cost of charge formation in the low dielectric protein, but it is less robust to structural changes than is electron transfer. Available experimental data do not clearly discriminate between these proposals. We have studied the mechanism of photoactivation (light-induced reduction of the flavin adenine dinucleotide cofactor) of Escherichia coli DNA photolyase using time-resolved absorption spectroscopy. Here we show that the excited flavin adenine dinucleotide radical abstracts an electron from a nearby tryptophan in 30 ps. After subsequent electron transfer along a chain of three tryptophans, the most remote tryptophan (as a cation radical) releases a proton to the solvent in about 300 ns, showing that electron transfer occurs before proton dissociation. A similar process may take place in photolyase-like blue-light receptors. PMID- 10850722 TI - US Congress encouraged to lay out the welcome mat for skilled foreigners. PMID- 10850721 TI - Engineering stability in gene networks by autoregulation. AB - The genetic and biochemical networks which underlie such things as homeostasis in metabolism and the developmental programs of living cells, must withstand considerable variations and random perturbations of biochemical parameters. These occur as transient changes in, for example, transcription, translation, and RNA and protein degradation. The intensity and duration of these perturbations differ between cells in a population. The unique state of cells, and thus the diversity in a population, is owing to the different environmental stimuli the individual cells experience and the inherent stochastic nature of biochemical processes (for example, refs 5 and 6). It has been proposed, but not demonstrated, that autoregulatory, negative feedback loops in gene circuits provide stability, thereby limiting the range over which the concentrations of network components fluctuate. Here we have designed and constructed simple gene circuits consisting of a regulator and transcriptional repressor modules in Escherichia coli and we show the gain of stability produced by negative feedback. PMID- 10850723 TI - The economic impact of Silicon Valley's immigrant entrepreneurs PMID- 10850724 TI - Cholinesterase inhibitor therapy stabilizes symptoms of Alzheimer disease. AB - Cholinesterase inhibitors tested in clinical trials in Europe, the United States, and Japan include fewer than 10 drugs; however, most of these compounds have advanced to clinical phase III trials. Based on results related to a population of more than 8,000 patients, we conclude that several of these compounds have shown significant clinical efficacy and safety in the treatment of Alzheimer disease. There are, however, differences with regard to side effects. The major clinical effect is stabilization of cognitive function during a 6- to 12-month period with an improvement of behavioral symptoms. The long-term effect of cholinesterase inhibitors extending to a 2-year period was reported. Future applications of these drugs include treatment of other types of dementias such as Lewy bodies dementia, vascular dementia, and Down syndrome dementia. The combination of cholinesterase inhibitors with estrogens, antioxidants, and anti inflammatories may represent a further improvement of the therapy. From an economical point of view, treatment with cholinesterase inhibitors is not cost neutral. PMID- 10850725 TI - Alzheimer disease: current therapy and future therapeutic strategies. AB - Antidementia drugs can be defined as drugs that significantly improve the decreased cognitive functions and/or inhibit the progression of dementia, compared with placebo. The main target of antidementia drugs is Alzheimer disease (AD), and the advent of such drugs is ardently desired. Antidementia drugs that are currently in use or undergoing trial are briefly reviewed. To date, only a few acetylcholine esterase inhibitors have been licensed as antidementia drugs for AD, but more beneficial drugs are being actively sought by many different approaches. The development of additional drugs requires greater basic research on the pathogenesis of AD. Future therapeutic strategies for AD on the basis of recent findings concerning the pathogenesis of AD are also reviewed. PMID- 10850726 TI - Significance of tau phosphorylation and protein kinase regulation in the pathogenesis of Alzheimer disease. AB - The role of the phosphatidylinositol-3 kinase pathway in the hyperphosphorylation of tau protein was investigated in cultured cells. Human kidney 293T-cells were cotransfected with tau and glycogen synthase kinase-3 (GSK-3) genes or tau and protein kinase B genes. The phosphorylation of tau protein was increased by cotransfection with GSK-3; however, it was decreased by cotransfection with protein kinase B. Human neuroblastoma SY5Y cells were treated with wortmannin, an inhibitor of phosphatidylinositol-3 kinase, and only transient (after 1 hour) activation of GSK-3 and hyperphosphorylation of tau protein were observed. However, continuous inactivation of protein kinase B was observed, suggesting the involvement of protein kinases other than protein kinase B in the phosphorylation and inactivation of GSK-3 after 3 hours. In cells treated with wortmannin, protein kinase C delta fragments were observed, and the protein kinase C activity increased after 3 hours, whereas treatment of cells with z-DEVD-fmk, an inhibitor of caspase-3, inhibited fragmentation of protein kinase C delta and induced continuous activation of GSK-3. It is suggested that fragmentation of protein kinase C delta during the process of apoptosis results in the phosphorylation and the inactivation of GSK-3. Those data suggest that, in Alzheimer disease, more complicated mechanisms are involved in the process of phosphorylation of tau protein predominantly regulated by P13K pathway. PMID- 10850727 TI - Familial cerebral amyloid angiopathies and dementia. AB - Amyloidosis is a disorder of protein conformation leading to aggregation. The term defines a diverse group of proteins normally present in body fluids as soluble precursors that can be deposited as insoluble amyloid fibrils in different tissues producing organ dysfunction and cell death. These fibrils are composed of self-assembled, low molecular weight mass peptides adopting beta pleated sheet structure, the conformation responsible for their physicochemical properties and tinctoreal characteristics. So far, 20 different proteins have been identified as subunits of amyloid fibrils (Westermark et al., 1999). Collectively, they are products of normal genes; however, several amyloid precursors contain abnormal amino acid substitutions that can impose an unusual potential for self-aggregation. The molecular mass of the amyloid peptides is within the 4 to 30-kDa range, with heterogeneity at the amino- and carboxyl terminal portions found in most amyloid proteins. Increased levels of amyloid precursors, either in the circulation or locally in sites of deposition, are usually the result of overexpression or defective clearance, or both. Of the 20 amyloid proteins identified, few of them are known to cause amyloid deposition in the central nervous system, which in turn results in cognitive deficits, dementia, stroke, cerebellar and extrapyramidal signs, or a combination of them. PMID- 10850728 TI - Nerve growth factor treatment in dementia. AB - Millions of people are affected by Alzheimer disease. As longevity increases, so will the number of patients with dementia. This has led to an intense search for successful treatment strategies. One area of interest is neurotrophic factors. Brain development and neuronal maintenance, as well as protective efforts, are mediated by a large number of different neurotrophic factors acting on specific receptors. In neurodegenerative disorders, there may be a possibility of rescuing degenerating neurons and stimulating terminal outgrowth with use of neurotrophic factors. The first neurotrophic factor discovered was nerve growth factor (NGF). A wealth of animal studies have shown that cholinergic neurons are NGF sensitive and NGF dependent, which is especially interesting in cognitive disorders, in which central cholinergic projections are important for cognitive function. In Alzheimer disease, cholinergic neurons have been shown to degenerate. This suggests that NGF may be used to pharmacologically counteract cholinergic degeneration and/or induce terminal sprouting in Alzheimer disease. Data from animal studies, as well as from the author's recent clinical trial, in which NGF was infused to the lateral ventricle in patients with Alzheimer disease, will be presented. Effects of NGF on cognition, as well as issues regarding dosage, side effects, and alternative ways of administering NGF, will be discussed. PMID- 10850729 TI - Neurotrophic factor strategies for the treatment of Alzheimer disease. AB - Cholinergic neurons in the nucleus basalis of Meynert are reduced early in the course of Alzheimer disease, and the dysfunction of cholinergic neurons is believed to be primarily responsible for cognitive deficits in the disease. Nerve growth factor has a trophic effect on cholinergic neurons and therefore may have some beneficial effects on the cognitive impairment observed in patients with Alzheimer disease. Experimental studies demonstrated that a continuous infusion of nerve growth factor into the cerebroventricle prevents cholinergic neuron atrophy after axotomy or associated with normal aging and ameliorates cognition impairment in these animals. A clinical study in three patients with Alzheimer disease revealed, however, that a long-term intracerebroventricular infusion of nerve growth factor may have certain potentially beneficial effects, but the continuous intracerebroventricular route of administration is also associated with negative side effects that appear to outweigh the positive effects. Several other strategies have been suggested to provide neurotrophic support to cholinergic neurons. In this article, we review the neurotrophic factor strategies for the treatment of Alzheimer disease. PMID- 10850730 TI - Cell mediators of inflammation in the Alzheimer disease brain. AB - Lesions of Alzheimer disease are associated with low-grade but sustained inflammatory responses. Activated microglia agglomerate in the center of senile plaques. Reactive astrocytes marginate the amyloid beta-protein (A beta) deposits and extend their processes toward the center of plaques. Both microglia and astrocytes are known to secrete a wide variety of molecules involved in inflammation and are potential sources of proinflammatory elements in the brain. Dystrophic neurites occur in senile plaques with such glial reactions, suggesting the relevance of inflammatory responses to the neuronal degeneration in Alzheimer disease. Activated glial cells are, therefore, targets of anti-inflammatory therapy of Alzheimer disease. However, evidence also indicates that these cells eliminate A beta from the brain. A beta is produced continuously in both the normal and the AD brain. Under normal conditions, A beta is removed successfully before it accumulates as extracellular amyloid fibrils. Even in Alzheimer disease, a large portion of A beta may be cleared from the brain with a small portion being left and deposited as neurotoxic senile plaques. Both in vivo and in vitro studies showed the effective uptake of A beta by microglia. Before clinical application, it must be determined whether the treatment that suppresses glial activation and inflammatory responses inhibits A beta removal by glial cells. PMID- 10850731 TI - Neuroinflammation and Alzheimer disease: clinical and therapeutic implications. AB - In Alzheimer disease brains, the amyloid plaques are closely associated with a locally induced, nonimmune-mediated, chronic inflammatory response without any apparent influx of leukocytes from the blood. The present findings indicate that in cerebral A beta diseases (Alzheimer disease, Down syndrome, hereditary cerebral hemorrhage with amyloidosis-Dutch type), the clinical symptoms are determined to a great extent by the site of inflammatory response. It was found that the formation of the amyloid-microglia complex seems to be a relatively early pathogenic event that precedes the process of severe destruction of the neuropil. The idea that inflammation is implicated in Alzheimer pathology has received support from the epidemiologic studies indicating that the use of anti inflammatory drugs can prevent or retard the Alzheimer disease process. In this contribution, we review the relationship between inflammation and clinical manifestation and the opportunities for anti-inflammatory treatments in Alzheimer disease. PMID- 10850732 TI - Oxidative stress, antioxidants, and Alzheimer disease. AB - Recent evidence in the field of Alzheimer disease research has highlighted the importance of oxidative processes in its pathogenesis. Examination of cellular changes shows that oxidative stress is an event that precedes the appearance of neurofibrillary tangles, one of the hallmark pathologies of the disease. Although it is still unclear what the initial source of the oxidative stress is in Alzheimer disease, it is likely that the process is highly dependent on the presence of redox-active transition metals, such as iron and copper. Because of the proximal role that oxidative stress mechanisms seem to play in the pathogenesis of Alzheimer disease, further investigation in this realm may lead to novel therapeutic strategies. PMID- 10850733 TI - Interaction of amyloid precursor proteins and heme oxygenase. AB - Amyloid precursor protein generates the beta-amyloid peptide and is a member of a multigene family that contains at least two other gene products, known as amyloid precursor-like proteins. Heme oxygenase is an enzyme whose products are antioxidant and neuroprotective. The authors report that amyloid precursor protein and amyloid precursor-like protein bind to heme oxygenase to inhibit its enzymatic activity. Our findings suggest that decreased neuroprotection caused by amyloid precursor protein-heme oxygenase interactions influences neurotoxicity in Alzheimer disease. PMID- 10850734 TI - Cerebral hypoperfusion, capillary degeneration, and development of Alzheimer disease. AB - Considerable clinical and experimental data have shown that cerebral perfusion is progressively decreased during increased aging and that this decrease in brain blood flow is significantly greater in Alzheimer disease (AD). The authors propose that advanced aging with a comorbid condition, such as a vascular risk factor, which further decreases cerebral perfusion, promotes a critically attained threshold of cerebral hypoperfusion (CATCH). With time, CATCH induces brain capillary degeneration and suboptimal delivery of energy substrates to neuronal tissue. Because glucose is the main fuel of brain cells, its impaired delivery, with the deficient delivery of oxygen, compromises neuronal stability because the supply for aerobic glycolysis fails to meet brain tissue demand. The outcome of CATCH is a metabolic cascade that involves, among other things, mitochondrial dysfunction, oxidative stress, decreased adenosine triphosphate production, abnormal protein synthesis, cell ionic pump deficiency, signal transduction defects, and neurotransmission failure. These events contribute to the progressive cognitive decline characteristic of patients with AD, as well as regional anatomic pathology, consisting of synaptic loss, senile plaques, neurofibrillary tangles, tissue atrophy, and neurodegeneration. CATCH identifies the clinical heterogeneic pattern that characterizes AD because it provides compelling evidence that any of a multitude of different etiopathophysiologic vascular risk factors, in the presence of advanced aging, can lead to AD. The evidence in support of CATCH as the pathogenic trigger of AD is crystallized in this review. PMID- 10850735 TI - Presynaptic nicotinic acetylcholine receptors as a functional target of nefiracetam in inducing a long-lasting facilitation of hippocampal neurotransmission. AB - Nefiracetam (1-10 microM), a nootropic (or cognition-enhancing) agent, persistently potentiated currents through Torpedo acetylcholine (ACh) receptors expressed in Xenopus oocytes as a result of interacting with a protein kinase C pathway and the ensuing protein kinase C phosphorylation of the receptors. A similar effect was found in neuronal nicotinic ACh receptors (alpha4beta2 and alpha7). In contrast, the other nootropic agents such as piracetam and aniracetam had no potentiating action on the receptors. A sustained enhancement in the activity of nicotinic ACh receptors induced by nefiracetam caused a marked increase in the glutamate release, leading to a long-term potentiation-like facilitation of hippocampal synaptic transmissions. One of the consistent neuropathologic features of the Alzheimer brain is a loss of nicotinic ACh receptors. This fact, together with the results of our study, raises the possibility that the loss of nicotinic ACh receptors may be a key factor in the decline of cognitive function observed in Alzheimer disease and that agents targeting neuronal nicotinic ACh receptors like nefiracetam could, therefore, be of great therapeutic importance. PMID- 10850736 TI - Cellular mechanism of action of cognitive enhancers: effects of nefiracetam on neuronal Ca2+ channels. AB - Cellular mechanisms underlying the cognition-enhancing actions of piracetam-like nootropics were studied by recording Ca2+ channel currents from neuroblastoma x glioma hybrid (NG108-15) cells and Xenopus oocytes expressing Ca2+ channels. In NG108-15 cells, nefiracetam (1 microM) produced a twofold increase in L-type Ca2+ channel currents. A similar, but slightly less potent effect was observed with aniracetam, whereas piracetam and oxiracetam exerted no such effects. Cyclic AMP analogs mimicked the nefiracetam action. N-type Ca2+ channel currents inhibited by leucine (Leu)-enkephalin by means of inhibitory G proteins (Go/Gi) were recovered promptly by nefiracetam, whereas those inhibited by prostaglandin E1 via stimulatory G proteins were not affected by nefiracetam. Cells treated with pertussis toxin (500 ng/mL, > 20 hours) were insensitive to nefiracetam. In Xenopus oocytes functionally expressing N-type (alpha1B) Ca2+ channels and delta opioid receptors, nefiracetam was also effective in facilitating the recovery from Leu-enkephalin-induced inhibition. These results suggest that nefiracetam, and possibly aniracetam, may activate N- and L-type Ca2+ channels in a differential way depending on how they recover from Go/Gi-mediated inhibition. PMID- 10850737 TI - Clinical issues in current drug therapy for dementia. AB - Dementia resulting from Alzheimer disease is one of the most prevalent medical problems. Elaborate expert guidelines for the diagnosis and treatment of Alzheimer disease do not always take sufficient account of the resources available in general practice. The focus on pure Alzheimer disease can be inappropriate for the large proportion of mixed dementia cases in old age. Because of such guidelines, treatment with modern and effective drugs is often delayed until conservative dementia criteria are satisfied. Criteria for the discontinuation of antidementia drugs are highly questionable. Antidementia drug sales in Germany demonstrate that the majority of prescribers hold on to conservative attitudes and prefer Ginkgo biloba and memantine to acetylcholinesterase inhibitors. Disappointment after exaggerated expectations and financial restrictions in the health care sector may aggravate current underprescribing of antidementia drugs. Even though contemporary symptomatic treatments for Alzheimer disease are unsatisfactory, modern medicine has been very successful in the early diagnosis and treatment of other potential causes of dementia. Future strategies will include models for the early identification of individuals carrying a high risk of developing cognitive impairment during their lifetime. PMID- 10850738 TI - Early diagnosis of Alzheimer disease with positron emission tomography. AB - The emergence of drugs that may slow progression of Alzheimer disease, if administered early during its course, has necessitated early diagnosis of the disease itself. Among the functional imaging methods that could assist in early diagnosis, positron emission tomography has an important role in providing quantitative measures of various aspects of brain function affected by the disease. Positron emission tomography studies in patients with Alzheimer disease have revealed a typical pattern of metabolic deficits in the temporal and parietal lobes. Additionally, converging evidence from numerous studies indicates that a similar pattern of deficits can be observed in nondemented subjects who are at risk of developing the disease, such as those with recognized genetic traits such as familial Alzheimer disease with mutations in chromosomes 21 and 14, Down syndrome, subjects with the epsilon4 allele of the apolipoprotein E gene, and individuals with mild cognitive impairment. These findings might have implications for the selection of patients for clinical trials, defining the outcome measures and evaluation of treatment efficacy and responder characteristics, but should be confirmed by prospective studies comprising larger samples and include clinicopathologic correlations. PMID- 10850739 TI - Brain acetylcholinesterase activity in Alzheimer disease measured by positron emission tomography. AB - Brain acetylcholinesterase activity was measured in 14 patients with Alzheimer disease and 14 age-matched control subjects by positron emission tomography with a radioactive acetylcholine analogue. Kinetic analysis was performed to calculate k3, an index of acetylcholinesterase activity. The k3 values were significantly reduced in the neocortex, hippocampus, and amygdala of all patients with Alzheimer disease, suggesting a loss of cholinergic innervation from the basal forebrain. Most profound reductions of k3 values were observed in the temporal ( 30%) and parietal cortices (-31%), although reductions of k3 values were relatively uniform in the cerebral neocortex. This technique may be a powerful tool for early diagnosis of Alzheimer disease and also for therapeutic monitoring of acetylcholinesterase inhibitors in Alzheimer disease. PMID- 10850740 TI - Harmonization of Dementia Drug Guidelines (United States and Europe): a report of the International Working Group for the Harmonization for Dementia Drug Guidelines. AB - The International Working Group for the Harmonization of Dementia Drug Guidelines has been active for more than 5 years attempting to improve the efficiency of international drug development. A virtual organization was formed of clinicians, scientists, industries, participants, and regulators from various nations around the world. This article summarizes a variety of conferences and physicians' papers focusing on aspects of trials ranging from diagnosis, assessment, and trial design to pharmacoeconomics and ethics. PMID- 10850741 TI - Current status of clinical trials for Alzheimer-type dementia in Japan: strategies to facilitate the development of antidementia drugs. AB - Several antidementia drugs are currently being developed in Japan. However, although the clinical trial period is one of the most important factors in drug development, it is not always possible to recruit a sufficient number of eligible patients in each facility for two reasons. One is related to the skill of investigators who are requested to undertake the trial, and another is related to the clinical trial system itself. The author outlines the current status of the development of acetylcholinesterase inhibitors as antidementia drugs and discuss the strategies to facilitate the development of antidementia drugs in Japan. PMID- 10850742 TI - New models for a new age. PMID- 10850743 TI - Heterogeneity in dementia: challenges and opportunities. AB - The major sources of heterogeneity in the manifestation of dementia are analyzed as stemming from three types of sources: initial predisposing factors, lifelong events, and the current condition. Each of these sources is, in turn, comprised of three general domains: genetic/biologic/medical, psychosocial, and environmental. These influences affect the manifestation of dementia in terms of cognitive, behavioral, self-maintenance, and affective functioning. Mapping the sources and manifestations of this heterogeneity can lead to both a more thorough understanding of people with dementia and the development of management settings and interventions that are more responsive to people with dementia. PMID- 10850744 TI - Why we are not ready for a biopsychosocial approach to dementia research and intervention: commentary on "Heterogeneity in dementia: challenges and opportunities". PMID- 10850745 TI - A review of psychosocial models in psychogeriatrics: implications for care and research. AB - Based on an examination of the relevant literature, this article presents a survey of psychosocial models used in the psychogeriatric field. Models with a multidimensional approach to behavior problems in dementia and a focus on the individual were selected. The utility of these psychosocial models as a theoretical framework for emotion-oriented care for people with dementia, especially Alzheimer disease, is examined. In addition to describing the models, this article also reports on the target group for which these models were developed, the degree to which they have been operationalized for psychogeriatrics, and the degree to which they have been subjected to empirical testing. This study shows that all psychosocial models described may be called emotion-oriented, although they also contain, in varying degrees, elements from the consequences model. It was found that the models are used not only as a theoretical framework for research, but also in the provision of care. Despite the fact that most models have been used in the psychogeriatric field, therefore proving their practicability for psychogeriatric practice, only one of the models described has been tested empirically so far. The theoretical validity of the models in question is, therefore, not yet clear. PMID- 10850746 TI - Common pattern of language impairment in vascular dementia and in Alzheimer disease. AB - The authors studied language performance patterns in early stages of vascular dementia and Alzheimer disease. The objective was to clarify to what extent dissolution of language in vascular dementia is similar to that in Alzheimer disease. Both structured language tests (comprehension, repetition, reading, and naming tasks) and nonstructured language tests (object and picture description) were employed. The structured tasks evidenced impairment on complex auditory comprehension and on picture naming for both dementia groups, whereas oral reading and single word repetition did not differentiate the patients from matched control subjects. On the unstructured narrative tasks, both patient groups showed normal fluency, but content analysis revealed that the patients with dementia produced fewer semantic units (themes) than the control subjects. In summary, both patient groups showed impairment, specifically on semantically mediated language tasks. According to the present results, language impairment in vascular dementia resembles that observed in Alzheimer disease. Semantically mediated functions are among the most sensitive language measures in differentiating early stages of both vascular dementia and Alzheimer disease from normal aging. PMID- 10850747 TI - Lessons learned from the Medicare Alzheimer Disease Demonstration. AB - The Medicare Alzheimer Disease Demonstration tested a case management and community care benefit for persons with dementia. The demonstration produced statistically but not clinically significant reductions in caregiver burden and depression. It increased access to community-based long-term care services but did not affect the level of services used. It did not reduce informal caregiver hours spent helping people with dementia. It produced statistically significant but not budget-neutral reductions in Medicare expenditures in that the degree of reduction in regular Medicare expenditures was not enough to offset the added demonstration costs. It did not reduce rates of nursing home placement. Informal care networks providing care to demented enrollees were generally able to function effectively, regardless of whether a professional case manager was involved or whether a long-term care benefit was available. PMID- 10850748 TI - A telemedicine system as a care modality for dementia patients in Korea. AB - Because dementia is a chronic debilitating disease, there are the issues of the difficulty in continuous long-term care and limited accessibility to medical service. We developed the telemedicine system for dementia patients and aimed to examine the acceptance, reliability, and clinical outcome of our telemedicine service. We established the Dementia Telemedicine Center in connection with two recipient sites in 1996. The reliability of the center, which provides telemedicine, tele-education, and telecounseling services, was tested by comparing assessment via our system with in-person assessment, and the clinical outcome was assessed by rating the changes of behavioral symptoms. There have been 140 registered patients for 2 years. The general acceptance of our system by the patients and caregivers was good, and the consistency rates between the assessment via our telemedicine system and in-person assessment ranged from 76% to 89%. A considerable proportion of dementia patients in nursing homes (46%) showed relative clinical improvements through our service. Our telemedicine system seems to be reliable and effective for the assessment and care of dementia patients. Our future direction is to promote our system as a core model of the home-based care system for dementia patients. PMID- 10850749 TI - Propentofylline treatment for Alzheimer disease and vascular dementia: an economic evaluation based on functional abilities. AB - A Canadian economic evaluation of propentofylline (a therapy shown to be effective for patients with mild to moderate Alzheimer disease and/or vascular dementia) versus standard care was conducted. Patients were categorized by functional abilities according to the Alberta Resident Classification System by translating measures that were originally captured through the Gottfries-Brane Steen scale. The Alberta Resident Classification System was then linked to a community dataset of home care costs for a population with dementia. Cost and cost-effectiveness analyses were performed from the perspective of the Ministry of Health, the caregiver, and society using an intent-to-treat analysis for propentofylline versus placebo. Results, limited to the 48-week clinical trial duration, indicated that propentofylline improved health outcomes of persons with dementia as statistically significant treatment effects were found. However, although an incremental cost for the propentofylline intervention was incurred from the Ministry of Health perspective, home care and, to a larger extent, caregiver costs were reduced. Savings in these areas may have partially offset annual treatment medication costs because a non-statistically significant cost difference was observed from a societal perspective. PMID- 10850750 TI - Alzheimer disease: absolute quantification of cerebral metabolites in vivo using localized proton magnetic resonance spectroscopy. AB - In vivo magnetic resonance spectroscopy of brain metabolites such as N acetylaspartate and myo-inositol has been proposed for the diagnosis of Alzheimer disease. Thirty patients with probable Alzheimer disease as well as 22 elderly controls underwent quantitative proton magnetic resonance spectroscopy of parietal gray and white matter with use of a short-echo time localization technique (echo time, 20 ms; repetition times, 6,000 and 3,000 ms, 2.0 Tesla) providing access to the regional concentrations of N-acetylaspartate, creatine, choline-containing compounds, myoinositol, glutamate, glutamine, and lactate. No statistically significant alterations of the metabolites were found in patients relative to controls. There were also no differences between patients with early and late onset of the disease and with respect to the presence of APOE-epsilon4 phenotype. A general trend for slightly decreased levels of N-acetylaspartate and creatine was not observed for their respective concentration ratios. In summary, the spectroscopic findings were in accord with known Alzheimer disease neuropathology, i.e., mild gliosis in white matter as well as mildly enhanced cortical atrophy in comparison to elderly controls. However, cortical atrophy with little or no N-acetylaspartate changes provided no evidence for a major decrease of neuronal density or loss of viable neurons. The data do not support the utility of proton magnetic resonance spectroscopy as an early diagnostic tool for Alzheimer disease. PMID- 10850752 TI - On the relation between W'/W index, hyper-Wiener index, and Wiener numbers AB - It is shown analytically that the W'/W index, the hyper-Wiener index, and the Wiener number are closely related graph-theoretical invariants for acyclic structures. A general analytical expression for the hyper-Wiener index of a tree is derived too. PMID- 10850751 TI - Adult-onset Hallervorden-Spatz syndrome presenting as cortical dementia. AB - The authors examined behavioral and pathophysiologic substrates in a patient with adult-onset Hallervorden-Spatz syndrome who presented with insidious cognitive decline but no motor impairment. The authors combined longitudinal case history and serial neuropsychologic testing with functional neuroimaging (positron emission tomography), structural neuroimaging (magnetic resonance imaging), and brain tissue analyses. Serial assessments of a 29-year-old woman showed progressive dementia. Marked cognitive and behavioral deficits were seen on neuropsychologic testing, corresponding to striking cortical abnormalities on positron emission tomography, magnetic resonance imaging, and histopathologic studies. Typical motor manifestations of the disorder did not emerge until the patient was 34 years old, 5 years after the onset of cognitive symptoms. Hallervorden-Spatz syndrome should be considered in the differential diagnosis of progressive cortical dementia in a young adult, even in the absence of motor dysfunction. PMID- 10850753 TI - On numerical characterization of cyclicity AB - We propose characterizing the cyclicity of molecular graphs by considering their D/DD matrix. Each nondiagonal element of D/DD is a quotient of the corresponding elements of the distance matrix D and the detour matrix DD of a graph. In particular, we are using the leading eigenvalue of the D/DD matrix as a descriptor of cyclicity and are investigating for monocyclic graphs Cn how this eigenvalue depends on the number of vertexes n, as n approaches infinity. PMID- 10850755 TI - Lists of face-regular polyhedra AB - We introduce a new notion that connects the combinatorial concept of regularity with the geometrical notion of face transitivity. This new notion implies finiteness results in the case of bounded maximal face size. We give lists of structures for some classes and investigate polyhedra with constant vertex degrees and faces of only two sizes. PMID- 10850754 TI - Precipitation at equivalence and equilibrium: a method for the determination of equilibrium constants of reaction between multideterminant antigen and specific polyclonal antibodies. AB - A theoretical approach for the determination of the equilibrium constant, Ka, of the reaction between a multideterminant antigen (Ag) and specific polyclonal antibodies (Ab) forming the insoluble Ab/Ag immune complex, is derived. The constant can be expressed as a function of the two accessible experimental parameters, the precipitating concentration of the antigen and the Ab/Ag molar ratio. For this purpose Ab/Ag immune complex must be prepared at equivalence, and equilibrium between precipitated and soluble species must be reached. The proposed method is experimentally tested on the system human serum albumin (HSA) and polyclonal rabbit antibodies. The Ab/Ag precipitates are prepared by the direct mixing of biological fluids in which immunoreacting components naturally occur. Previous separation, purification, or labeling of immunoreacting components are not required. The conditions for the precipitation of Ab/Ag complexes at equivalence, the stoichiometric composition or the average number of Ab molecules bound to one Ag molecule, and the solubility of the immunoprecipitating components are determined by a rectangular two-dimensional double immunodiffusion. Since the solubility determined under the conditions of a double immunodiffusion is a result of the interaction of the global diffusion of the precipitating components and particle growth kinetics, it mostly refers to the dynamic conditions. To find the solubility under equilibrium conditions, it is sufficient to determine the minimal factor by which the solutions of both immunoprecipitating components should be diluted so that no precipitate is formed upon their mixing at equivalence. The dilution factor is determined by a measurement of the laser light scattering of the immunoprecipitating systems prepared with serially diluted Ag and Ab solutions. PMID- 10850756 TI - Dynamical process of excitation fusion in polymers AB - Due to the confinement effect in the nondegenerate polymers and the electron lattice interaction, the electron-hole pair forms a self-trapped single excitation. It is shown that two single excitations in one chain can fuse into one double excitation, and the relaxation time of the fusion process is about 160 fs. PMID- 10850757 TI - Immunohistochemical analysis and prognostic value of cathepsin D determination in laryngeal squamous cell carcinoma. AB - Cathepsin D, a protease with the capability of degrading matrix proteins, is implicated in the process of breast and colorectal cancer invasion and metastasis. Biochemical studies in laryngeal cancer have shown a potential prognostic significance of cathepsin D content determination. We studied immunohistochemical positivity of cathepsin D in tumor epithelium and stroma of 61 surgical specimens of squamous cell laryngeal cancer. Immunohistochemical reaction was quantitatively assessed using a PC-based image analysis system SFORM VAMS. The results were correlated to clinical and morphological parameters and survival. Immunohistochemical positivity was noted in neoplastic cells and tumor stroma. Significant prognostic value for cathepsin D was established separately for epithelial tumor component and tumor stroma using log-rank test, the Cox proportional hazards regression model, and C4.5 machine learning system. In all groups, patients above the median cathepsin D staining showed significantly shorter survival time. C4.5 machine learning system extracted cutoff values for the decision tree that defines the probabilities of patients survival and death with high sensitivity (92.8% alive, 73.6% dead), 100% specificity, and 86.9% accuracy. This makes immunohistochemical cathepsin D estimation an independent prognostic parameter in laryngeal carcinomas within a 5-year period from the time of tumor surgery. PMID- 10850758 TI - Fullerenes as tilings of surfaces AB - If a fullerene is defined as a finite trivalent graph made up solely of pentagons and hexagons, embedding in only four surfaces is possible: the sphere, torus, Klein bottle, and projective (elliptic) plane. The usual spherical fullerenes have 12 pentagons; elliptic fullerenes, 6; and toroidal and Klein-bottle fullerenes, none. Klein-bottle and elliptic fullerenes are the antipodal quotients of centrosymmetric toroidal and spherical fullerenes, respectively. Extensions to infinite systems (plane fullerenes, cylindrical fullerenes, and space fullerenes) are indicated. Eigenvalue spectra of all four classes of finite fullerenes, are reviewed. Leapfrog fullerenes have equal numbers of positive and negative eigenvalues, with 0, 0, 2, or 4 eigenvalues zero for spherical, elliptic, Klein-bottle, and toroidal cases, respectively. PMID- 10850759 TI - Byte structure variable length coding (BS-VLC): a new specific algorithm applied in the compression of trajectories generated by molecular dynamics AB - Molecular dynamics is a well-known technique very much used in the study of biomolecular systems. The trajectory files produced by molecular dynamics simulations are extensive, and the classical lossless algorithms give poor efficiencies in their compression. In this work, a new specific algorithm, named byte structure variable length coding (BS-VLC), is introduced. Trajectory files, obtained by molecular dynamics applied to trypsin and a trypsin:pancreatic trypsin inhibitor complex, were compressed using four classical lossless algorithms (Huffman, adaptive Huffman, LZW, and LZ77) as well as the BS-VLC algorithm. The results obtained show that BS-VLC nearly triplicates the compression efficiency of the best classical lossless algorithm, preserving a near lossless behavior. Compression efficiencies close to 50% can be obtained with a high degree of precision, and the maximum efficiency possible (75%), within this algorithm, can be performed with good precision. PMID- 10850760 TI - Characterizing graph drawing with eigenvectors AB - Two definitions of the problem of graph drawing are considered, and an analytical solution is provided for each of them. The solutions obtained make use of the eigenvectors of the Laplacian matrix of a related structure. The procedures give good results for symmetrical graphs, and they have already been used for drawing fullerene molecules in the literature. The analysis characterizes precisely what problems the two procedures are solving. It also illuminates why they can perform unsatisfactorily on asymmetrical graphs. PMID- 10850761 TI - A model for combinatorial organic chemistry AB - The set of coset representations, CR's, of a group G, [G(/G1), G(/G2), ..., G(/Gs)], where G1 = [I], Gs = G, the marks, m(ij) of subgroup Gj on a given G(/Gi), 1 < or = i < or = s, and the subduction of G(/Gi) by Gj, j < or = i, G(/Gi) precipitates to Gj, are essential tools for the enumeration of stereoisomers and their classification according to their subgroup symmetry (Fujita, S. Symmetry and Combinatorial Enumeration in Chemistry; Springer-Verlag: Berlin, 1991). In this paper, each G(/Gi) is modeled by a set of colored equivalent configurations (called homomers), H = h1, h2, ..., hr, r = G/Gi, such that a given homomer, h(k), remains invariant only under all g epsilon Gi, where g is an element of symmetry. The resulting homomers generate the corresponding set of marks almost by inspection. The symmetry relations among a set H can be conveniently stored in a Cayley-like diagram (Chartrand, G. Graphs as Mathematical Models; Prindle, Weber and Schmidt Incorporated: Boston, MA, 1977; Chapter 10), which is a complete digraph on r vertices so that an arc from vertex v(i) to vertex v(j) is colored with the set Sij of symmetry elements such that h(i)[g(if)]-->h(j),g(ij) epsilon Sij. In addition, each vertex, v(i), is associated with a loop that is colored with a set Sii so that g(ii) epsilon Sii stabilizes h(i). A Cayley-like diagram of a given CR, G[G(/Gi)], leads to graphical generation of G(/Gi) precipitates to Gj for all values of j and also to all m(ij)'s. Several group-theoretical results are rederived and/or became more envisagable through this modeling. The approach is examplified using C2, C3, D2, T, and D3 point groups and is applied to trishomocubane, a molecule that belongs to the D3 point group. PMID- 10850762 TI - MCR XVI. Mathematical support for combinatorial chemistry AB - The algebra of the s- and r-vectors is an adequate formal tool to describe chemical objects in an abstract way. Compounds as well as reactions are represented, including all constitutional and configurational aspects. The stereochemistry of simple organic molecules as well as those of metal-organic compounds may be described in a unique way. Ionic bonds, covalent bonds, aromatics, and electron-deficiency compounds can formally be described without loss of information. Even reaction types and the flow of electrons can be described by this algebra. The biggest benefit of this approach is its intrinsic group theoretical structure. This does not bother the chemist for its use but allows a computer to handle and structure huge amounts of chemical data. This is especially important for combinatorial chemistry, which deals with huge sets of chemically distinct molecules, the so-called molecular libraries. PMID- 10850763 TI - Parallel implementation of a Monte Carlo molecular stimulation program AB - Molecular simulation methods such as molecular dynamics and Monte Carlo are fundamental for the theoretical calculation of macroscopic and microscopic properties of chemical and biochemical systems. These methods often rely on heavy computations, and one sometimes feels the need to run them in powerful massively parallel machines. For moderate problem sizes, however, a not so powerful and less expensive solution based on a network of workstations may be quite satisfactory. In the present work, the strategy adopted in the development of a parallel version is outlined, using the message passing model, of a molecular simulation code to be used in a network of workstations. This parallel code is the adaptation of an older sequential code using the Metropolis Monte Carlo method. In this case, the message passing interface was used as the interprocess communications library, although the code could be easily adapted for other message passing systems such as the parallel virtual machine. For simple systems it is shown that speedups of 2 can be achieved for four processes with this cheap solution. For bigger and more complex simulated systems, even better speedups might be obtained, which indicates that the presented approach is appropriate for the efficient use of a network of workstations in parallel processing. PMID- 10850764 TI - Bounds for the Randic connectivity index AB - For a saturated hydrocarbon with n carbon atoms and m carbon-carbon bonds and with Randic connectivity index chi, two functions, L = L(n,m) and U = U(n,m), are determined, such that L < or = chi < U. These bounds are better than those previously reported; for most chemically relevant values of n and m there exist hydrocarbons for which chi = L or chi = U. PMID- 10850765 TI - On the similarity of DNA primary sequences. AB - We consider numerical characterization of graphical representations of DNA primary sequences. In particular we consider graphical representation of DNA of beta-globins of several species, including human, on the basis of the approach of A. Nandy in which nucleic bases are associated with a walk over integral points of a Cartesian x, y-coordinate system. With a so-generated graphical representation of DNA, we associate a distance/distance matrix, the elements of which are given by the quotient of the Euclidean and the graph theoretical distances, that is, through the space and through the bond distances for pairs of bases of graphical representation of DNA. We use eigenvalues of so-constructed matrices to characterize individual DNA sequences. The eigenvalues are used to construct numerical sequences, which are subsequently used for similarity/dissimilarity analysis. The results of such analysis have been compared and combined with similarity tables based on the frequency of occurrence of pairs of bases. PMID- 10850766 TI - Value of the urinary stone promoters/inhibitors ratios in the estimation of the risk of urolithiasis. AB - An imbalance between urinary-promoting and -inhibiting factors has been suggested as more important in urinary stone formation than a disturbance of any single substance. To investigate the value of promoter/inhibitor ratios for estimation of the risk of urolithiasis, urinary citrate/calcium, magnesium/calcium oxalate, and oxalate/citrate x glycosaminoglycans ratios were determined in 30 children with urolithiasis, 36 children with isolated hematuria, and 15 healthy control children. The cutoff points between normal children and children with urolithiasis, accuracy, specificity, and sensitivity for each ratio were determined and compared with those of the 24-h urine calcium and oxalate excretion and urine saturation calculated with the computer program EQUIL 2. The neural network application (aiNET Artificial Neural Network, version 1.25) was used for the determination of the cutoff points for the classification of normal children and the urolithiasis group. The best test for differentiating stone formers from non-stone formers proved the aiNET determined cutoff values of oxalate/citrate x glycosaminoglycans ratio. The method showed 97.78% accuracy, 100% sensitivity, and 93.33% specificity. Two cutoff points between normal and urolithiasis groups were found showing that the children with urolithiasis had ratio values either above 34.00 or less than 10.16. Increased oxalate excretion was linked to the first cutoff value (34.00), and decreased glycosaminoglycans excretion was typical of the second cutoff value (10.16). PMID- 10850767 TI - Quantitative determination of magnetization exchange rate constants from a series of two-dimensional exchange NMR spectra AB - A method for quantitative determination of magnetization exchange rate constants (cross-relaxation and chemical exchange) from a series of two-dimensional exchange spectra is presented. The method, the least error matrix analysis (LEMA), combines a series of full matrix calculations at different mixing times in a least-squares manner. LEMA embodies the principal advantages of full relaxation matrix analysis (FMA) and initial rate buildup (BU) analysis. Like FMA, it takes into account all the relations among the spectral matrix elements and in analogy to BU makes use of their time evolution. By means of calculations, simulations, and experiments, we have shown that LEMA provides the dynamic matrix from a given set of experimental data with errors that are smaller than in either FMA or BU calculations. PMID- 10850768 TI - Novel canonical coding method for representation of three-dimensional structures AB - A new canonical coding method for representation of three-dimensional structures, CAST (CAnonical representation of STereochemistry), is described. CAST canonically codes stereochemistry around an atom in a molecule. The same CAST notations are given for atoms of molecules in the same conformation. The CAST code is based on the dihedral angles of four atoms that are uniquely defined by a molecular tree structure. CAST has successfully represented similarities and differences between several conformers. PMID- 10850769 TI - Evaluation in quantitative structure--property relationship models of structural descriptors derived from information-theory operators AB - During the search for new structural descriptors we have defined the information theory operators U(M), V(M), X(M), and Y(M), that are computed from atomic invariants and measure the information content of the elements of molecular matrices. Structural descriptors computed with these four information-theory operators are used to develop structure-property models for the boiling temperature, molar heat capacity, standard Gibbs energy of formation, vaporization enthalpy, refractive index, and density of alkanes. The information theory operators were applied to six molecular matrices, namely, the distance D, the reciprocal distance RD, the distance-path Dp, the reciprocal distance-path RDp, the path Szeged Sz(p), and the reciprocal path Szeged RSz(p) matrices. In combination with other topological indices, the information-theory indices offer good structure-property models for all six alkane properties investigated in this study. PMID- 10850770 TI - Quadratic shape descriptors. 1. Rapid superposition of dissimilar molecules using geometrically invariant surface descriptors. AB - In this paper, we present a novel approach to shape-based molecular similarity searching. The method that we introduce is able to superimpose dissimilar molecules by using geometrically invariant molecular surface descriptors. The shape descriptors are calculated by least-squares fitting of a quadratic function to small sections of the molecular surface of a ligand. Invariant geometric properties of the approximated surface patch are then extracted from the fitted quadratic function. The extracted properties are used to quantify the shape and to obtain a canonical orientation for this section of surface. The superimposition algorithm uses these geometric invariants to recognize similar regions of surface shape existing on two molecules and to bring these regions (and consequently the molecules) into registration. Because these geometric descriptors are based upon local surface shape, the superimposing algorithm is insensitive to the connectivity and the relative sizes of the molecules being matched. The capabilities of our algorithm are demonstrated by superimposing dissimilar ligands known to inhibit the same enzyme system. In all cases examined the algorithm generates superpositions that are in agreement with crystallographic results. The algorithm is also applied to align the two different proteins on the basis of the shape of their active sites. PMID- 10850771 TI - Molecular field Topology analysis method in QSAR studies of organic compounds AB - A new method of QSAR analysis for organic compounds, molecular field topology analysis (MFTA), is considered that involves the topological superposition of the training set structures and the construction of a molecular supergraph (MSG). This enables the creation of the uniform descriptor vectors based on the local physicochemical parameters (atom and bond properties) of the molecules. The application of this technique is illustrated by a number of examples, and its features are discussed. The MFTA is especially suitable for solving the problems where the analysis of three-dimensional structure is either unnecessary or complicated. PMID- 10850772 TI - Binary formal inference-based recursive modeling using multiple atom and physicochemical property class pair and torsion descriptors as decision criteria AB - Analysis of a large amount of information, typically generated by high-throughput screening, is a very difficult task. To address this problem, we have developed binary formal inference-based recursive modeling using atom and physicochemical property class pair and torsion descriptors. Recursive partitioning is an exploratory technique for identifying structure in data. The implemented algorithm utilizes a statistical hypothesis testing, similar to Hawkins' formal inference-based recursive modeling program, to separate a data set into two homogeneous subsets at each splitting node. This process is repeated recursively until no further separation can occur. Our implementation of recursive partitioning differs from previously reported approaches by employing a method to extract multiple features at each splitting node. The method was examined for its ability to distinguish random and real data sets. The effect of including a single descriptor and multiple descriptors in the splitting descriptor set was also studied. The method was tested using 27 401 National Cancer Institute (NCI) compounds and their pGI50 (-log(GI50)) against the NCI-H23 cell line. The analyses show that partitioning using multiple descriptors is advantageous in analyzing the structure-activity relationship information. PMID- 10850773 TI - Universal model based on the mobile order and disorder theory for predicting lipophilicity and partition coefficients in all mutually immiscible two-phase liquid systems AB - The quantitative thermodynamic development of the mobile order and disorder theory in H-bonded liquids has been extended in order to predict partition coefficients. The model enables "a priori" estimation of the partition coefficient (log P) of neutral solutes, not only in the conventional 1 octanol/water reference but also in all mutually saturated two-phase systems made up of largely immiscible solvents. The model is obtained from the thermodynamic treatment of the various physicochemical free energy processes encoded in the overall distribution process and accordingly provides a useful tool for better understanding both the origin and the factors, such as the solute molar volume, that determine the partition coefficient of nonelectrolytes in a given system. From the comparison of the relative magnitude of the processes contributing to the log P value, a lot of information can also be gained regarding the variation in log P of the same substance partitioned between different solvent systems. As a demonstration, the model has been successfully applied to predict the log P of a great number of chemicals of varying structure, size, and chemical nature partitioned in a large set of essentially immiscible solvent pairs, differing either by their nonpolar or by their polar phase. In the systems involving water as the polar phase, the hydrophobic effect is always the driving force that governs the distribution process irrespective of the interacting or noninteracting nature of the substances studied. In the other two-phase systems, the partitioning of complexing solutes in particular appears to be ruled rather by their hydrogen-bonding capabilities than by their hydrophobicities. PMID- 10850774 TI - Combinatorial library design: maximizing model-fitting compounds within matrix synthesis constraints AB - The use of combinatorial chemistry has become commonplace within the pharmaceutical industry. Less widespread but gaining in popularity is the derivation of activity models from the high-throughput assays of these libraries. Such models are then used as filters during the design of refined daughter libraries. The design of these second generation libraries, which efficiently test and conform to the derived activity model from the large space of virtual possibilities, remains an area of considerable research. We present here a computationally efficient method for the design of optimally dense (in model matching compounds) synthetic matrices from in silico virtual libraries. PMID- 10850775 TI - Prediction of IC50 values for ACAT inhibitors from molecular structure. AB - A quantitative structure-activity study is performed on several series of compounds derived from N-chlorosulfonyl isocyanate to develop models that relate their structures to IC50 activity for inhibition of acyl-CoA:cholesterol O acyltransferase (ACAT). Numerical descriptors are used to encode topological, electronic, and geometric information from the molecular structures of the inhibitors. A data set of 157 compounds showing triglyceride- and cholesterol lowering activity is used to develop successful linear regression models and nonlinear computational neural network models. The models are validated using an external prediction set. PMID- 10850776 TI - Correction of temperature variations in kinetic-based determinations by use of pruning computational neural networks in conjunction with genetic algorithms. AB - The joint use of genetic algorithms and pruning computational neural networks is shown to be an effective means for selecting the number of inputs required to correct temperature variations in kinetic-based determinations. The genetic algorithm uses a pruning procedure based on Bayesian regularization and is highly efficient as a feature selector; it provides quite good results in the generalization process without the need to use a validation set. The fitness function is defined as the sum of two subfunctions: one controls the learning ability of the network and the other its complexity. The training, pruning, and generalization processes were initially tested with simulated data in order to acquire preliminary information for the ensuing work with real data. The performance of the proposed method was assessed by applying it to the determination of the amino acid L-glycine by its classical spectrophotometric reaction with ninhydrin. A straightforward network topology including temperature as input (40+T:2:1 with 19 connections after the pruning process) was used to estimate the L-glycine concentration from kinetic curves affected by temperature variations over the range 60-75 degrees C, using kinetic data acquired up to only 1.5 half-lives. The trained network estimates this concentration with a standard error of prediction for the testing set of ca. 8%, which is much smaller than those provided by a classical parametric method such as nonlinear regression (even if kinetic data acquired at longer half-lives are used). Finally, a kinetic interpretation of the pruning process is provided in order to better demonstrate its potential for kinetic analysis. PMID- 10850777 TI - Comparison of weighting schemes for molecular graph descriptors: application in quantitative structure-retention relationship models for alkylphenols in gas liquid chromatography AB - Organic compounds containing heteroatoms or multiple bonds can be conveniently represented as vertex- and edge-weighted molecular graphs. These atom and bond parameters can be computed for any organic compound with two parameter sets that we have recently defined, namely, the relative electronegativity X and the relative covalent radius Y weighting schemes. Structural descriptors computed with these two weighting schemes and the previously defined atomic number Z parameter set are used to develop quantitative structure-retention relationship (QSRR) models for alkylphenols in gas-liquid chromatography. The QSRR models are generated with structural descriptors computed with several newly introduced graph operators, namely, the Wiener, hyper-Wiener, minimum eigenvalue, maximum eigenvalue, Ivanciuc-Balaban, and information on distance operators. These molecular graph operators were applied to the distance D and the reciprocal distance RD matrixes. PMID- 10850778 TI - Design and development of computer-aided chemical systems: representation and balance of inorganic chemical reactions AB - A model for the tracking of inorganic chemical reactions is proposed. Designed to acquire, process, and solve a great number of inorganic reactions, this model will hopefully contribute to the development of powerful computer-aided chemistry teaching systems for use within or without the environment of a virtual laboratory. Using full representation of an inorganic reaction to allow the extraction of chemical knowledge, incomplete reactions (where species are absent) may be completed by adding the necessary species, and reactions may be solved and balanced. Various types of reaction are classified, and a layer-based model is defined for the solution of different reaction types, establishing the basis for the construction of a system which, based on a wide set of production rules, is capable of solving an incomplete inorganic chemical reaction. PMID- 10850779 TI - Prediction of inhibition of the sodium ion-proton antiporter by benzoylguanidine derivatives from molecular structure. AB - The use of quantitative structure-activity relationships to predict IC50 values of 113 potential Na+/H+ antiporter inhibitors is reported. Multiple linear regression and computational neural networks (CNNs) are used to develop models using a set of information-rich descriptors. The descriptors encode information about topology, geometry, electronics, and combination hybrids. A five-descriptor CNN model with root-mean-square (rms) errors of 0.278 log units for the training set and 0.377 log units for the prediction set was developed. Examination of data set subclasses showed that systematic structural variations were also well encoded resulting in 100% accuracy of prediction trends. An experiment involving a committee of five CNNs was also performed to examine the effect of network output averaging. This showed improved results decreasing the training and cross validation set rms error to 0.228 log units and the prediction set rms error to 0.296 log units. PMID- 10850780 TI - Quantitative structure-activity relationship studies of progesterone receptor binding steroids. AB - The selection of appropriate descriptors is an important step in the successful formulation of quantitative structure-activity relationships (QSARs). This paper compares a number of feature selection routines and mapping methods that are in current use. They include forward stepping regression (FSR), genetic function approximation (GFA), generalized simulated annealing (GSA), and genetic neural network (GNN). On the basis of a data set of steroids of known in vitro binding affinity to the progsterone receptor, a number of QSAR models are constructed. A comparison of the predictive qualities for both training and test compounds demonstrates that the GNN protocol achieves the best results among the 2D QSAR that are considered. Analysis of the choice of descriptors by the GNN method shows that the results are consistent with established SARs on this series of compounds. PMID- 10850781 TI - Estimation of aqueous solubility for a diverse set of organic compounds based on molecular topology AB - An accurate and generally applicable method for estimating aqueous solubilities for a diverse set of 1297 organic compounds based on multilinear regression and artificial neural network modeling was developed. Molecular connectivity, shape, and atom-type electrotopological state (E-state) indices were used as structural parameters. The data set was divided into a training set of 884 compounds and a randomly chosen test set of 413 compounds. The structural parameters in a 30-12-1 artificial neural network included 24 atom-type E-state indices and six other topological indices, and for the test set, a predictive r2 = 0.92 and s = 0.60 were achieved. With the same parameters the statistics in the multilinear regression were r2 = 0.88 and s = 0.71, respectively. PMID- 10850782 TI - Efficient method for calculating the resonance energy expression of benzenoid hydrocarbons based on the eunumeration of conjugated circuits AB - To reduce the calculating time for the summations over linearly independent and minimal conjugated circuits of benzenoid hydrocarbons (BHs), an approximate method is proposed that counts only the numbers of the first four classes of conjugated circuits R1, R2, R3, and R4, respectively. By representation of BHs as custom-made "ring-block chains" and use of the techniques of Database and visual computing, an application software is realized that is much faster and more powerful than the old one based on an enumeration technique. PMID- 10850783 TI - The E-state as the basis for molecular structure space definition and structure similarity AB - The electrotopological state (E-state) is presented as a representation of molecular structure useful for definition of a space for chemical structures. This E-state representation provides the basis for chemical database management. The E-state formalism is presented along with its extension to the atom-type E state. An approach to database organization, using polychlorobiphenyls (PCBs) as examples, reveals the descriptive power of the E-state paradigm. A well-organized chemical database, as described here, may be searched to find structures similar to a target structure with the expectation that such structures may exhibit properties similar to the target. Searches using the atom-type E-state indices are demonstrated with two example drug molecules. PMID- 10850784 TI - Intermolecular accessibility: the meaning of molecular connectivity AB - The molecular connectivity indices are shown to be the numerical possibilities of a molecule encountering another identical molecule, derived from the bond accessibilites of each. The meaning of the delta values and the significance of all suppressed X-H bonds is described. A new concept of the meaning of molecular connectivity, built around bimolecular encounter accessibility, is presented. PMID- 10850785 TI - Variability of molecular descriptors in compound databases revealed by Shannon entropy calculations AB - A method is introduced to calculate and compare the variability of molecular descriptors in compound databases. Descriptor variability analysis is based on histograms recording the distribution of molecular descriptors and calculation of Shannon entropy (SE), a metric originally applied in digital communication. SE values reflect the variability of descriptor settings. We have calculated a total of 92 molecular descriptors in the ACD and NCI databases and ranked them according to their variability. Significant differences in entropy are observed for a number of descriptors. However, the most variable descriptors are similar in the ACD and NCI databases. Such high-entropy descriptors are preferred tools to discriminate between compounds or account for the diversity of chemical libraries. PMID- 10850786 TI - Molecular descriptors for effective classification of biologically active compounds based on principal component analysis identified by a genetic algorithm. AB - We have evaluated combinations of 111 descriptors that were calculated from two dimensional representations of molecules to classify 455 compounds belonging to seven biological activity classes using a method based on principal component analysis. The analysis was facilitated by application of a genetic algorithm. Using scoring functions that related the number of compounds in pure classes (i.e., compounds with the same biological activity), singletons, and mixed classes, effective descriptor sets were identified. A combination of only four molecular descriptors accounting for aromatic character, hydrogen bond acceptors, estimated polar van der Waals surface area, and a single structural key gave overall best results. At this performance level, approximately 91% of the compounds occurred in pure classes and mixed classes were absent. The results indicate that combinations of only a few critical descriptors are preferred to partition compounds according to their biological activity, at least in the test cases studied here. PMID- 10850787 TI - Two-dimensional arrays in the analysis of trends in series of molecules: strongly subspectral molecular graphs, formula periodic tables, and number of resonance structures AB - Two-dimensional arrays consisting of strongly subspectral molecular graphs and formula periodic tables for polycyclic aromatic hydrocarbons are briefly reviewed. New two-dimensional arrays for free-radical benzenoid hydrocarbons are presented with general analytical expressions for counting their number of resonance structures (SC). The structural origin of the coefficients to these analytical expressions is discussed. PMID- 10850788 TI - Critical point representations of electron density maps for the comparison of benzodiazepine-type ligands. AB - A procedure for the comparison of three-dimensional electron density distributions is proposed for similarity searches between pharmacological ligands at various levels of crystallographic resolution. First, a graph representation of molecular electron density distributions is generated using a critical point analysis approach. Pairwise as well as multiple comparisons between the obtained graphs of critical points are then carried out using a Monte Carlo/simulated annealing technique, and results are compared with genetic algorithm solutions. PMID- 10850789 TI - How hard is mechanism elucidation in catalysis? Combinatorial analysis of C1 chemistry AB - Most chemical reactions occur over multiple steps whose identity is elucidated by experiment, yielding a reaction mechanism. Knowledge of cognitive science suggests that mechanism elucidation can be viewed as a knowledge-guided search within a combinatorial space. The MECHEM computer program searches this space comprehensively for the simplest plausible mechanisms. We use MECHEM to find mechanisms for Fischer-Tropsch chemistry and CO2 re-forming of methane, both heterogeneous catalytic reactions of current importance. The results reveal hundreds of equally simple mechanisms consistent with evidence. Hence, mechanism elucidation in catalysis is a much harder problem than is ordinarily realized. PMID- 10850790 TI - Quantification of the influence of single-point mutations on haloalkane dehalogenase activity: a molecular quantum similarity study. AB - Controlled modifications in certain protein amino acid residues can lead to changes in their function and stability. Amino acid structural features and their relation to these changes were examined by using quantum molecular similarity techniques. The effect of deliberate mutations in position 172 of the haloalkane dehalogenase enzyme, yielding to variations on the dehalogenation of 1,2 dibromoethane, was studied qualitatively and quantitatively using molecular quantum similarity techniques. A valuable classification of the residues according to their effect on activity was obtained by representing the optimal two-dimensional classical scaling solution. In addition, satisfactory quantitative relationships were found, comparable to those attained by previous studies on this same data set using other techniques. Molecular quantum similarity analysis provides a consistent, unbiased, and homogeneous set of molecular descriptors and is a feasible alternative to the use of physicochemical properties. PMID- 10850791 TI - Modeling of ion complexation and extraction using substructural molecular fragments AB - A substructural molecular fragment (SMF) method has been developed to model the relationships between the structure of organic molecules and their thermodynamic parameters of complexation or extraction. The method is based on the splitting of a molecule into fragments, and on calculations of their contributions to a given property. It uses two types of fragments: atom/bond sequences and "augmented atoms" (atoms with their nearest neighbors). The SMF approach is tested on physical properties of C2-C9 alkanes (boiling point, molar volume, molar refraction, heat of vaporization, surface tension, melting point, critical temperature, and critical pressures) and on octanol/water partition coefficients. Then, it is applied to the assessment of (i) complexation stability constants of alkali cations with crown ethers and phosphoryl-containing podands, and of beta cyclodextrins with mono- and 1,4-disubstituted benzenes, and (ii) solvent extraction constants for the complexes of uranyl cation by phosphoryl-containing ligands. PMID- 10850792 TI - Neural network based quantitative structural property relations (QSPRs) for predicting boiling points of aliphatic hydrocarbons AB - Quantitative structural property relations (QSPRs) for boiling points of aliphatic hydrocarbons were derived using a back-propagation neural network and a modified Fuzzy ARTMAP architecture. With the back-propagation model, the selected molecular descriptors were capable of distinguishing between diastereomers. The QSPRs were obtained from four valance molecular connectivity indices (1chiv,2chiv,3chiv,4chiv), a second-order Kappa shape index (2kappa), dipole moment, and molecular weight. The inclusion of dipole moment proved to be particularly useful for distinguishing between cis and trans isomers. A back propagation 7-4-1 architecture predicted boiling points for the test, validation, and overall data sets of alkanes with average absolute errors of 0.37% (1.65 K), 0.42% (1.73 K), and 0.37% (1.54 K), respectively. The error for the test and overall data sets decreased to 0.19% (0.81 K) and 0.31% (1.30 K), respectively, using the modified Fuzzy ARTMAP network. A back-propagation alkene model, with a 7-10-1 architecture, yielded predictions with average absolute errors for the test, validation, and overall data sets of 1.96% (6.79 K), 1.83% (6.45 K), and 1.25% (4.42 K), respectively. Fuzzy ARTMAP reduced the errors for the test and overall data sets to 0.19% (0.73 K) and 0.25% (0.95 K), respectively. The back propagation composite model for aliphatic hydrocarbons, with a 7-9- architecture, yielded boiling points with average absolute errors for the test, validation, and overall set of 1.74% (6.09 K), 1.25% (4.68 K), and 1.37% (4.85 K), respectively. The error for the test and overall data sets using the Fuzzy ARTMAP composite model decreased to 0.84% (1.15 K) and 0.35% (1.35 K), respectively. Performance of the QSPRs, developed from a simple set of molecular descriptors, displayed accuracy well within the range of expected experimental errors and of better accuracy than other regression analysis and neural network-based boiling points QSPRs previously reported in the literature. PMID- 10850793 TI - Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. AB - The solution structure and stability of N-terminally truncated beta2 microglobulin (deltaN6beta2-m), the major modification in ex vivo fibrils, have been investigated by a variety of biophysical techniques. The results show that deltaN6beta2-m has a free energy of stabilization that is reduced by 2.5 kcal/mol compared to the intact protein. Hydrogen exchange of a mixture of the truncated and full-length proteins at microM concentrations at pH 6.5 monitored by electrospray mass spectrometry reveals that deltaN6beta2-m is significantly less protected than its wild-type counterpart. Analysis of deltaN6beta2-m by NMR shows that this loss of protection occurs in beta strands I, III, and part of II. At mM concentration gel filtration analysis shows that deltaN6beta2-m forms a series of oligomers, including trimers and tetramers, and NMR analysis indicates that strand V is involved in intermolecular interactions that stabilize this association. The truncated species of beta2-microglobulin was found to have a higher tendency to self-associate than the intact molecule, and unlike wild-type protein, is able to form amyloid fibrils at physiological pH. Limited proteolysis experiments and analysis by mass spectrometry support the conformational modifications identified by NMR and suggest that deltaN6beta2-m could be a key intermediate of a proteolytic pathway of beta2-microglobulin. Overall, the data suggest that removal of the six residues from the N-terminus of beta2 microglobulin has a major effect on the stability of the overall fold. Part of the tertiary structure is preserved substantially by the disulfide bridge between Cys25 and Cys80, but the pairing between beta-strands far removed from this constrain is greatly perturbed. PMID- 10850794 TI - The structure and dynamics in solution of Cu(I) pseudoazurin from Paracoccus pantotrophus. AB - The solution structure and backbone dynamics of Cu(I) pseudoazurin, a 123 amino acid electron transfer protein from Paracoccus pantotrophus, have been determined using NMR methods. The structure was calculated to high precision, with a backbone RMS deviation for secondary structure elements of 0.35+/-0.06 A, using 1,498 distance and 55 torsion angle constraints. The protein has a double-wound Greek-key fold with two alpha-helices toward its C-terminus, similar to that of its oxidized counterpart determined by X-ray crystallography. Comparison of the Cu(I) solution structure with the X-ray structure of the Cu(II) protein shows only small differences in the positions of some of the secondary structure elements. Order parameters S2, measured for amide nitrogens, indicate that the backbone of the protein is rigid on the picosecond to nanosecond timescale. PMID- 10850795 TI - Binding of Zn2+ to a Ca2+ loop allosterically attenuates the activity of factor VIIa and reduces its affinity for tissue factor. AB - The protease domain of coagulation factor VIIa (FVIIa) is homologous to trypsin with a similar active site architecture. The catalytic function of FVIIa is regulated by allosteric modulations induced by binding of divalent metal ions and the cofactor tissue factor (TF). To further elucidate the mechanisms behind these transformations, the effects of Zn2+ binding to FVIIa in the free form and in complex with TF were investigated. Equilibrium dialysis suggested that two Zn2+ bind with high affinity to FVIIa outside the N-terminal gamma-carboxyglutamic acid (Gla) domain. Binding of Zn2+ to FVIIa, which was influenced by the presence of Ca2+, resulted in decreased amidolytic activity and slightly reduced affinity for TF. After binding to TF, FVIIa was less susceptible to zinc inhibition. Alanine substitutions for either of two histidine residues unique for FVIIa, His216, and His257, produced FVIIa variants with decreased sensitivity to Zn2+ inhibition. A search for putative Zn2+ binding sites in the crystal structure of the FVIIa protease domain was performed by Grid calculations. We identified a pair of Zn2+ binding sites in the Glu210-Glu220 Ca2+ binding loop adjacent to the so-called activation domain canonical to serine proteases. Based on our results, we propose a model that describes the conformational changes underlying the Zn2+ mediated allosteric down-regulation of FVIIa's activity. PMID- 10850796 TI - Conformational transitions and fibrillation mechanism of human calcitonin as studied by high-resolution solid-state 13C NMR. AB - Conformational transitions of human calcitonin (hCT) during fibril formation in the acidic and neutral conditions were investigated by high-resolution solid state 13C NMR spectroscopy. In aqueous acetic acid solution (pH 3.3), a local alpha-helical form is present around Gly10 whereas a random coil form is dominant as viewed from Phe22, Ala26, and Ala31 in the monomer form on the basis of the 13C chemical shifts. On the other hand, a local beta-sheet form as viewed from Gly10 and Phe22, and both beta-sheet and random coil as viewed from Ala26 and Ala31 were detected in the fibril at pH 3.3. The results indicate that conformational transitions from alpha-helix to beta-sheet, and from random coil to beta-sheet forms occurred in the central and C-terminus regions, respectively, during the fibril formation. The increased 13C resonance intensities of fibrils after a certain delay time suggests that the fibrillation can be explained by a two-step reaction mechanism in which the first step is a homogeneous association to form a nucleus, and the second step is an autocatalytic heterogeneous fibrillation. In contrast to the fibril at pH 3.3, the fibril at pH 7.5 formed a local beta-sheet conformation at the central region and exhibited a random coil at the C-terminus region. Not only a hydrophobic interaction among the amphiphilic alpha-helices, but also an electrostatic interaction between charged side chains can play an important role for the fibril formation at pH 7.5 and 3.3 acting as electrostatically favorable and unfavorable interactions, respectively. These results suggest that hCT fibrils are formed by stacking antiparallel beta sheets at pH 7.5 and a mixture of antiparallel and parallel beta-sheets at pH 3.3. PMID- 10850797 TI - Ser45 plays an important role in managing both the equilibrium and transition state energetics of the streptavidin-biotin system. AB - The contribution of the Ser45 hydrogen bond to biotin binding activation and equilibrium thermodynamics was investigated by biophysical and X-ray crystallographic studies. The S45A mutant exhibits a 1,700-fold greater dissociation rate and 907-fold lower equilibrium affinity for biotin relative to wild-type streptavidin at 37 degrees C, indicating a crucial role in binding energetics. The crystal structure of the biotin-bound mutant reveals only small changes from the wild-type bound structure, and the remaining hydrogen bonds to biotin retain approximately the same lengths. No additional water molecules are observed to replace the missing hydroxyl, in contrast to the previously studied D128A mutant. The equilibrium deltaG degrees, deltaH degrees, deltaS degrees, deltaC degrees(p), and activation deltaG++ of S45A at 37 degrees C are 13.7+/-0.1 kcal/mol, -21.1+/-0.5 kcal/mol, -23.7+/-1.8 cal/mol K, -223+/-12 cal/mol K, and 20.0+/-2.5 kcal/mol, respectively. Eyring analysis of the large temperature dependence of the S45A off-rate resolves the deltaH++ and deltaS++ of dissociation, 25.8+/-1.2 kcal/mol and 18.7+/-4.3 cal/mol K. The large increases of deltaH++ and deltaS++ in the mutant, relative to wild-type, indicate that Ser45 could form a hydrogen bond with biotin in the wild-type dissociation transition state, enthalpically stabilizing it, and constraining the transition state entropically. The postulated existence of a Ser45-mediated hydrogen bond in the wild-type streptavidin transition state is consistent with potential of mean force simulations of the dissociation pathway and with molecular dynamics simulations of biotin pullout, where Ser45 is seen to form a hydrogen bond with the ureido oxygen as biotin slips past this residue after breaking the native hydrogen bonds. PMID- 10850798 TI - Conformational flexibility in the apolipoprotein E amino-terminal domain structure determined from three new crystal forms: implications for lipid binding. AB - An amino-terminal fragment of human apolipoprotein E3 (residues 1-165) has been expressed and crystallized in three different crystal forms under similar crystallization conditions. One crystal form has nearly identical cell dimensions to the previously reported orthorhombic (P2(1)2(1)2(1)) crystal form of the amino terminal 22 kDa fragment of apolipoprotein E (residues 1-191). A second orthorhombic crystal form (P2(1)2(1)2(1) with cell dimensions differing from the first form) and a trigonal (P3(1)21) crystal form were also characterized. The structures of the first orthorhombic and the trigonal form were determined by seleno-methionine multiwavelength anomalous dispersion, and the structure of the second orthorhombic form was determined by molecular replacement using the structure from the trigonal form as a search model. A combination of modern experimental and computational techniques provided high-quality electron-density maps, which revealed new features of the apolipoprotein E structure, including an unambiguously traced loop connecting helices 2 and 3 in the four-helix bundle and a number of multiconformation side chains. The three crystal forms contain a common intermolecular, antiparallel packing arrangement. The electrostatic complimentarity observed in this antiparallel packing resembles the interaction of apolipoprotein E with the monoclonal antibody 2E8 and the low density lipoprotein receptor. Superposition of the model structures from all three crystal forms reveals flexibility and pronounced kinks in helices near one end of the four-helix bundle. This mobility at one end of the molecule provides new insights into the structural changes in apolipoprotein E that occur with lipid association. PMID- 10850799 TI - Evidence for phosphorylation-dependent conformational changes in methylesterase CheB. AB - Enhancement of methylesterase activity of the response regulator CheB is dependent upon phosphorylation of the N-terminal regulatory domain of the enzyme. This domain plays a dual role in the regulation of methylesterase activity with an inhibitory effect in the unphosphorylated state and a stimulatory effect in the phosphorylated state. Structural studies of the unphosphorylated state have indicated that the basis for the regulatory domain's inhibitory effect is partial blockage of access of substrate to the active site suggesting that the activation upon phosphorylation involves a repositioning of the two domains with respect to each other. We report in this study evidence for phosphorylation-dependent conformational changes in CheB. Differences in rates of proteolytic cleavage by trypsin between the phosphorylated and unphosphorylated states have been observed at three sites in the protein with one site, 113, within the regulatory domain and two sites, 134 and 148, lying within the interdomain linker. These results support the hypothesis for the mechanism for the activation of CheB wherein phosphorylation of a specific aspartate residue within the N-terminal domain results in a propagated conformational change within the regulatory domain leading to a repositioning of its two domains. Presumably, structural changes in the regulatory domain of CheB facilitate a repositioning of the N- and C-terminal domains, leading to stimulation of methylesterase activity. PMID- 10850800 TI - Alternate modes of binding in two crystal structures of alkaline phosphatase inhibitor complexes. AB - Two high resolution crystal structures of Escherichia coli alkaline phosphatase (AP) in the presence of phosphonate inhibitors are reported. The phosphonate compounds, phosphonoacetic acid (PAA) and mercaptomethylphosphonic acid (MMP), bind competitively to AP with dissociation constants of 5.5 and 0.6 mM, respectively. The structures of the complexes of AP with PAA and MMP were refined at high resolution to crystallographic R-values of 19.0 and 17.5%, respectively. Refinement of the AP-inhibitor complexes was carried out using X-PLOR. The final round of refinement was done using SHELXL-97. Crystallographic analyses of the inhibitor complexes reveal different binding modes for the two phosphonate compounds. The significant difference in binding constants can be attributed to these alternative binding modes observed in the high resolution X-ray structures. The phosphinyl group of PAA coordinates to the active site zinc ions in a manner similar to the competitive inhibitor and product inorganic phosphate. In contrast, MMP binds with its phosphonate moiety directed toward solvent. Both enzyme-inhibitor complexes exhibit close contacts, one of which has the chemical and geometrical potential to be considered an unconventional hydrogen bond of the type C-H...X. PMID- 10850801 TI - Comparative model building of interleukin-7 using interleukin-4 as a template: a structural hypothesis that displays atypical surface chemistry in helix D important for receptor activation. AB - Using a combination of theoretical sequence structure recognition predictions and experimental disulfide bond assignments, a three-dimensional (3D) model of human interleukin-7 (hIL-7) was constructed that predicts atypical surface chemistry in helix D that is important for receptor activation. A 3D model of hIL-7 was built using the X-ray crystal structure of interleukin-4 (IL-4) as a template (Walter MR et al., 1992, J Mol Biol. 224:1075-1085; Walter MR et al., 1992, J Biol Chem 267:20371-20376). Core secondary structures were constructed from sequences of hIL-7 predicted to form helices. The model was constructed by superimposing IL-7 helices onto the IL-4 template and connecting them together in an up-up down-down topology. The model was finished by incorporating the disulfide bond assignments (Cys3, Cys142), (Cys35, Cys130), and (Cys48, Cys93), which were determined by MALDI mass spectroscopy and site-directed mutagenesis (Cosenza L, Sweeney E, Murphy JR, 1997, J Biol Chem 272:32995-33000). Quality analysis of the hIL-7 model identified poor structural features in the carboxyl terminus that, when further studied using hydrophobic moment analysis, detected an atypical structural property in helix D, which contains Cys 130 and Cys142. This analysis demonstrated that helix D had a hydrophobic surface exposed to bulk solvent that accounted for the poor quality of the model, but was suggestive of a region in IL 7 that maybe important for protein interactions. Alanine (Ala) substitution scanning mutagenesis was performed to test if the predicted atypical surface chemistry of helix D in the hIL-7 model is important for receptor activation. This analysis resulted in the construction, purification, and characterization of four hIL-7 variants, hIL-7(K121A), hIL-7(L136A), hIL-7(K140A), and hIL-7(W143A), that displayed reduced or abrogated ability to stimulate a murine IL-7 dependent pre-B cell proliferation. The mutant hIL-7(W143A), which is biologically inactive and displaces [125I]-hIL-7, is the first reported IL-7R system antagonist. PMID- 10850802 TI - The energetics of phosphate binding to a protein complex. AB - The heat of binding the serine protease, porcine pancreatic elastase, by the inhibitor, turkey ovomucoid third domain, is dependent on the presence of inorganic phosphate. This dependence is saturable and can be accurately modeled as the phosphate binding to a single site on the protease-inhibitor complex; thus, the elastase-ovomucoid system provides a unique opportunity to study phosphate-protein interactions. We have used isothermal titration calorimetry to investigate this binding, thereby providing one of the few complete thermodynamic characterizations of phosphate interacting with proteins. The binding is characterized by a small favorable deltaG degrees, a large unfavorable deltaH degrees, and a positive deltaCp, thermodynamics consistent with the release of water being linked to phosphate binding. These measurements provide insight into the binding of phosphotyrosine containing peptides to SH2 domains by suggesting the energetic consequences of binding phosphate free from other interactions. PMID- 10850803 TI - Role of Lys335 in the metastability and function of inhibitory serpins. AB - The native form of inhibitory serpins (serine protease inhibitors) is not in the thermodynamically most stable state but in a metastable state, which is critical to inhibitory functions. To understand structural basis and functional roles of the native metastability of inhibitory serpins, we have been characterizing stabilizing mutations of human alpha1-antitrypsin, a prototype inhibitory serpin. One of the sites that has been shown to be critical in stability and inhibitory activity of alpha1-antitrypsin is Lys335. In the present study, detailed roles of this lysine were analyzed by assessing the effects of 13 different amino acid substitutions. Results suggest that size and architect of the side chains at the 335 site determine the metastability of alpha1-antitrypsin. Moreover, factors such as polarity and flexibility of the side chain at this site, in addition to the metastability, seem to be critical for the inhibitory activity. Substitutions of the lysine at equivalent positions in two other inhibitory serpins, human alpha1-antichymotrypsin and human antithrombin III, also increased stability and decreased inhibitory activity toward alpha-chymotrypsin and thrombin, respectively. These results and characteristics of lysine side chain, such as flexibility, polarity, and the energetic cost upon burial, suggest that this lysine is one of the structural designs in regulating metastability and function of inhibitory serpins in general. PMID- 10850804 TI - Synthesis and NMR solution structure of an alpha-helical hairpin stapled with two disulfide bridges. AB - Helical coiled-coils and bundles are some of the most common structural motifs found in proteins. Design and synthesis of alpha-helical motifs may provide interesting scaffolds that can be useful as host structures to display functional sites, thus allowing the engineering of novel functional miniproteins. We have synthesized a 38-amino acid peptide, alpha2p8, encompassing the alpha-helical hairpin present in the structure of p8MTCP1, as an alpha-helical scaffold particularly promising for its stability and permissiveness of sequence mutations. The three-dimensional structure of this peptide has been solved using homonuclear two-dimensional NMR techniques at 600 MHz. After sequence specific assignment, a total of 285 distance and 29 dihedral restraints were collected. The solution structure of alpha2p8 is presented as a set of 30 DIANA structures, further refined by restrained molecular dynamics, using simulated annealing protocol with the AMBER force field. The RMSD values for the backbone and all heavy atoms are 0.65+/-0.25 and 1.51+/-0.21 A, respectively. Excised from its protein context, the alpha-hairpin keeps its native structure: an alpha-helical coiled-coil, similar to that found in superhelical structures, with two helices spanning residues 4-16 and 25-36, and linked by a short loop. This motif is stabilized by two interhelical disulfide bridges and several hydrophobic interactions at the helix interface, leaving most of its solvent-exposed surface available for mutation. This alpha-helical hairpin, easily amenable to synthetic chemistry and biological expression system, may represent a stable and versatile scaffold to display new functional sites and peptide libraries. PMID- 10850805 TI - Crystal structure of the antibiotic albomycin in complex with the outer membrane transporter FhuA. AB - One alternative method for drug delivery involves the use of siderophore antibiotic conjugates. These compounds represent a specific means by which potent antimicrobial agents, covalently linked to iron-chelating siderophores, can be actively transported across the outer membrane of gram-negative bacteria. These "Trojan Horse" antibiotics may prove useful as an efficient means to combat multi drug-resistant bacterial infections. Here we present the crystallographic structures of the natural siderophore-antibiotic conjugate albomycin and the siderophore phenylferricrocin, in complex with the active outer membrane transporter FhuA from Escherichia coli. To our knowledge, this represents the first structure of an antibiotic bound to its cognate transporter. Albomycins are broad-host range antibiotics that consist of a hydroxamate-type iron-chelating siderophore, and an antibiotically active, thioribosyl pyrimidine moiety. As observed with other hydroxamate-type siderophores, the three-dimensional structure of albomycin reveals an identical coordination geometry surrounding the ferric iron atom. Unexpectedly, this antibiotic assumes two conformational isomers in the binding site of FhuA, an extended and a compact form. The structural information derived from this study provides novel insights into the diverse array of antibiotic moieties that can be linked to the distal portion of iron-chelating siderophores and offers a structural platform for the rational design of hydroxamate-type siderophore-antibiotic conjugates. PMID- 10850806 TI - Peptide and metal ion-dependent association of isolated helix-loop-helix calcium binding domains: studies of thrombic fragments of calmodulin. AB - Calmodulin (CaM), the ubiquitous, eukaryotic, bilobal calcium-binding regulatory protein, has been cleaved by thrombin to create two fragments. TM1 (1-106) and TM2 (107-148). NMR and CD results indicate that TMI and TM2 can associate in the presence of Ca2+ to form a complex similar to native CaM, even though the cleavage site is not in the linker region between two helix-loop-helix domains, but rather within an alpha-helix. Cadmium-113 NMR results show that this complex has enhanced metal-ion binding properties when compared to either TM1 or TM2 alone. This complex can bind several CaM-binding target peptides, as shown by gel bandshift assays, circular dichroism spectra, and 13C NMR spectra of biosynthetically methyl-13C-Met-labeled TM1 and TM2; moreover, gel bandshift assays show that the addition of a target peptide strengthens the interactions between TM1 and TM2 and increases the stability of the complex. Cadmium-113 NMR spectra indicate that the TM1:TM2 complex can also bind the antipsychotic drug trifluoperazine. However, in contrast to CaM:peptide complexes, the TM1:TM2:peptide complexes are disrupted by 4 M urea; moreover, TM1 and TM2 in combination are unable to activate CaM-dependent enzymes. This suggests that TM1:TM2 mixtures cannot bind target molecules as tightly as intact CaM, or perhaps that binding occurs but additional interactions with the target enzymes that are necessary for proper activation are perturbed by the proteolytic cleavage. The results presented here reflect the importance of the existence of helix-loop-helix Ca2+-binding domains in pairs in proteins such as CaM, and extend the understanding of the association of such domains in this class of proteins in general. PMID- 10850807 TI - NMR solution structure of Apis mellifera chymotrypsin/cathepsin G inhibitor-1 (AMCI-1): structural similarity with Ascaris protease inhibitors. AB - The three-dimensional structure of the 56 residue polypeptide Apis mellifera chymotrypsin/cathepsin G inhibitor 1 (AMCI-1) isolated from honey bee hemolymph was calculated based on 730 experimental NMR restraints. It consists of two approximately perpendicular beta-sheets, several turns, and a long exposed loop that includes the protease binding site. The lack of extensive secondary structure features or hydrophobic core is compensated by the presence of five disulfide bridges that stabilize both the protein scaffold and the binding loop segment. A detailed analysis of the protease binding loop conformation reveals that it is similar to those found in other canonical serine protease inhibitors. The AMCI-1 structure exhibits a common fold with a novel family of inhibitors from the intestinal parasitic worm Ascaris suum. The pH-induced conformational changes in the binding loop region observed in the Ascaris inhibitor ATI are absent in AMCI-1. Similar binding site sequences and structures strongly suggest that the lack of the conformational change can be attributed to a Glu-->Gln substitution at the P1' position in AMCI-1, compared to ATI. Analysis of amide proton temperature coefficients shows very good correlation with the presence of hydrogen bond donors in the calculated AMCI-1 structure. PMID- 10850808 TI - Mutants provide evidence of the importance of glycosydic chains in the activation of lipase 1 from Candida rugosa. AB - Sequence analysis of Candida rugosa lipase 1 (LIP1) predicts the presence of three N-linked glycosylation sites at asparagine 291, 314, 351. To investigate the relevance of sugar chains in the activation and stabilization of LIP1, we directed site mutagenesis to replace the above mentioned asparagine with glutamine residues. Comparison of the activity of mutants with that of the wild type (wt) lipase indicates that both 314 and 351 Asn to Gln substitutions influence, although at a different extent, the enzyme activity both in hydrolysis and esterification reactions, but they do not alter the enzyme water activity profiles in organic solvents or temperature stability. Introduction of Gln to replace Asn351 is likely to disrupt a stabilizing interaction between the sugar chain and residues of the inner side of the lid in the enzyme active conformation. The effect of deglycosylation at position 314 is more difficult to explain and might suggest a more general role of the sugar moiety for the structural stability of lipase 1. Conversely, Asn291Gln substitution does not affect the lipolytic or the esterase activity of the mutant that behaves essentially as the wt enzyme. This observation supports the hypothesis that changes in activity of Asn314Gln and Asn351Gln mutants are specifically due to deglycosylation. PMID- 10850809 TI - Penicillopepsin-JT2, a recombinant enzyme from Penicillium janthinellum and the contribution of a hydrogen bond in subsite S3 to k(cat). AB - The nucleotide sequence of the gene (pepA) of a zymogen of an aspartic proteinase from Penicillium janthinellum with a 71% identity in the deduced amino acid sequence to penicillopepsin (which we propose to call penicillopepsin-JT1) has been determined. The gene consists of 60 codons for a putative leader sequence of 20 amino acid residues, a sequence of about 150 nucleotides that probably codes for an activation peptide and a sequence with two introns that codes for the active aspartic proteinase. This gene, inserted into the expression vector pGPT pyrG1, was expressed in an aspartic proteinase-free strain of Aspergillus niger var. awamori in high yield as a glycosylated form of the active enzyme that we call penicillopepsin-JT2. After removal of the carbohydrate component with endoglycosidase H, its relative molecular mass is between 33,700 and 34,000. Its kinetic properties, especially the rate-enhancing effects of the presence of alanine residues in positions P3 and P2' of substrates, are similar to those of penicillopepsin-JT1, endothiapepsin, rhizopuspepsin, and pig pepsin. Earlier findings suggested that this rate-enhancing effect was due to a hydrogen bond between the -NH- of P3 and the hydrogen bond accepting oxygen of the side chain of the fourth amino acid residue C-terminal to Asp215. Thr219 of penicillopepsin JT2 was mutated to Ser, Val, Gly, and Ala. Thr219Ser showed an increase in k(cat) when a P3 residue was present in the substrate, which was similar to that of the wild-type, whereas the mutants Thr219Val, Thr219Gly, and Thr219Ala showed no significant increase when a P3 residue was added. The results show that the putative hydrogen bond alone is responsible for the increase. We propose that by locking the -NH- of P3 to the enzyme, the scissile peptide bond between P1 and P1' becomes distorted toward a tetrahedral conformation and becomes more susceptible to nucleophilic attack by the catalytic apparatus without the need of a conformational change in the enzyme. PMID- 10850810 TI - Characterization of an anti-Borrelia burgdorferi OspA conformational epitope by limited proteolysis of monoclonal antibody-bound antigen and mass spectrometric peptide mapping. AB - Lyme borreliosis is a multisystem disorder caused by the spirochete Borrelia burgdorferi that is transmitted to humans by the tick Ixodes dammini. The immune response against the 31 kDa OspA, which is one of the most abundant B. burgdorferi proteins, appears to be critical in preventing infection and tissue inflammation. Detailed knowledge of the immunological and molecular characteristics of the OspA protein is important for the development of reliable diagnostic assays. In this study, we characterized a new conformational epitope present within the middle part of B. burgdorferi OspA. Our approach used enzymatic proteolyses of the immune complex followed by mass spectrometric identification of the peptides bound to the antibody. It appears to be one of the first reports on the characterization of a discontinuous epitope using mass spectrometry. PMID- 10850811 TI - Nonpolar contributions to conformational specificity in assemblies of designed short helical peptides. AB - A series of designed short helical peptides was used to study the effect of nonpolar interactions on conformational specificity. The consensus sequence was designed to obtain short helices (17 residues) and to minimize the presence of interhelical polar interactions. Furthermore, the sequence contained a heptad repeat (abcdefg), where positions a and d were occupied by hydrophobic residues Leu, Ile, or Val, and positions e and g were occupied by Ala. The peptides were named according to the identities of the residues in the adeg positions, respectively. The peptides llaa, liaa, ilaa, iiaa, ivaa, viaa, lvaa, vlaa, and vvaa were synthesized, and their characterization revealed marked differences in specificity. An experimental methodology was developed to study the nine peptides and their pairwise mixtures. These peptides and their mixtures formed a vast array of structural states, which may be classified as follows: helical tetramers and pentamers, soluble and insoluble helical aggregates, insoluble unstructured aggregates, and soluble unstructured monomers. The peptide liaa formed stable helical pentamers, and iiaa and vlaa formed stable helical tetramers. Disulfide cross-linking experiments indicated the presence of an antiparallel helix alignment in the helical pentamers and tetramers. Rates of amide proton exchange of the tetrameric form of vlaa were 10-fold slower than the calculated exchange rate for unfolded vlaa. In other work, the control of specificity has been attributed to polar interactions, especially buried polar interactions; this work demonstrated that subtle changes in the configuration of nonpolar interactions resulted in a large variation in the extent of conformational specificity of assemblies of designed short helical peptides. Thus, nonpolar interactions can have a significant effect on the conformational specificity of oligomeric short helices. PMID- 10850812 TI - Post-translational modification is essential for catalytic activity of nitrile hydratase. AB - Nitrile hydratase from Rhodococcus sp. N-771 is an alphabeta heterodimer with a nonheme ferric iron in the catalytic center. In the catalytic center, alphaCys112 and alphaCys114 are modified to a cysteine sulfinic acid (Cys-SO2H) and a cysteine sulfenic acid (Cys-SOH), respectively. To understand the function and the biogenic mechanism of these modified residues, we reconstituted the nitrile hydratase from recombinant unmodified subunits. The alphabeta complex reconstituted under argon exhibited no activity. However, it gradually gained the enzymatic activity through aerobic incubation. ESI-LC/MS analysis showed that the anaerobically reconstituted alphabeta complex did not have the modification of alphaCys112-SO2H and aerobic incubation induced the modification. The activity of the reconstituted alphabeta complex correlated with the amount of alphaCys112 SO2H. Furthermore, ESI-LC/MS analyses of the tryptic digest of the reconstituted complex, removed of ferric iron at low pH and carboxamidomethylated without reduction, suggested that alphaCys114 is modified to Cys-SOH together with the sulfinic acid modification of alphaCys112. These results suggest that alphaCys112 and alphaCys114 are spontaneously oxidized to Cys-SO2H and Cys-SOH, respectively, and alphaCys112-SO2H is responsible for the catalytic activity solely or in combination with alphaCys114-SOH. PMID- 10850813 TI - A model for the sickle hemoglobin fiber using both mutation sites. AB - The standard molecular model of the fiber of the sickle hemoglobin (HbS: beta6 Glu-->Val) has been revised to allow both beta6 mutation sites to participate in intermolecular contacts, rather than only one beta6 site as previously thought, for four molecules per 14-molecule fiber cross section. This structure accurately predicts the copolymerization of hybridized mixtures of HbS with HbA or HbC (beta6 Glu-->Lys), which could not be reconciled with prior models in which only half the beta6 sites were required for assembly. This model suggests new contacts within the fiber and raises the question of whether these cross-linked double strands could possess added stability important in such processes as nucleation. PMID- 10850814 TI - Temperature-sensitive suppressor mutations of the Escherichia coli DNA gyrase B protein. AB - Escherichia coli strain LE316 contains a mutation in gyrB that results in the substitution of Val164 to Gly and confers both chlorobiocin resistance and temperature sensitivity. Selection for suppressors of the ts phenotype yielded second-site mutations in GyrB at His38 and Thr157. The properties of proteins bearing these mutations have been characterized, and a mechanism of suppression is proposed based upon structural considerations. PMID- 10850815 TI - NMR characterization of a pH-dependent equilibrium between two folded solution conformations of the pheromone-binding protein from Bombyx mori. AB - NMR spectroscopic changes as a function of pH in solutions of the pheromone binding protein of Bombyx mori (BmPBP) show that BmPBP undergoes a conformational transition between pH 4.9 and 6.0. At pH below 4.9 there is a single "acid form" (A), and a homogeneous "basic form" (B) exists at pH above 6.0. Between pH 5 and 6, BmPBP exists as a mixture of A and B in slow exchange on the NMR chemical shift time scale, with the transition midpoint at pH 5.4. The form B has a well dispersed NMR spectrum, indicating that it represents a more structured, "closed" conformation than form A, which has a significantly narrower chemical shift dispersion. Conformational transitions of the kind observed here may explain heterogeneity reported for a variety of odorant-binding proteins, and it will be of interest to further investigate possible correlations with pH-dependent regulation of ligand binding and release in the biological function of this class of proteins. PMID- 10850816 TI - Mechanobiology in rehabilitation science. PMID- 10850817 TI - Low vision and blindness. PMID- 10850818 TI - Differential effects of embryonic immobilization on the development of fibrocartilaginous skeletal elements. AB - The importance of mechanical influences during skeletal development has been well established in both experimental studies and computer models. Under conditions of embryonic immobilization, it has been observed that the early stages of joint formation proceed normally (up to and including interzone formation), but the later stages of joint cavitation and maintenance are impaired, resulting in fusion of the cartilaginous elements across the presumptive joint line. Two structures in particular are noticeably absent from late-stage synovial joints in immobilized chick embryos: the menisci of the tibiofemoral joint and the plantar tarsal sesamoid of the tibiotarsal joint. Both of these fibrocartilaginous structures are known to serve mechanical functions in postnatal animals, helping to distribute loads within the joint and, in the case of sesamoid structures, to provide a mechanical advantage to muscles acting across the joint. We demonstrate in this study that embryonic immobilization differentially affects the developmental fate of these two distinct fibrocartilages. The absence of the plantar tarsal sesamoid in late-stage immobilized embryos is due to a failure in the initial formation of this structure. In contrast, the early stages of meniscus formation proceed normally. Without the normal mechanical stimuli of skeletal muscle contractions, however, the meniscus fails to mature and ultimately degenerates. PMID- 10850819 TI - Mechanobiology of tendon adaptation to compressive loading through fibrocartilaginous metaplasia. AB - Tendons that wrap around bones often undergo fibrocartilaginous metaplasia. In this paper, we examine the biomechanical causes and consequences of this metaplasia. We propose an adaptation rule in which tissue permeability changes in response to local cyclic hydrostatic pressures associated with physical activity. The proposed rule predicts the development of a low-permeability region corresponding to the fibrocartilaginous region in a representative wrap-around tendon. A poroelastic finite element model is used to examine the time-dependent fluid pressures and compressive stresses and strains in the solid constituents of the tendon's extrafibrillar matrix. The low permeability in the adapted fibrocartilaginous region maintains fluid pressures, protecting the solid constituents of the tendon's extracellular matrix from high compressive stresses and strains that could disrupt the matrix organization. Adaptation through fibrocartilaginous metaplasia therefore allows wrap-around tendons to function effectively over a lifetime without sustaining excessive mechanical damage due to cyclic compressive loading. PMID- 10850820 TI - Mechanobiology in the development, maintenance, and degeneration of articular cartilage. AB - During skeletal development, the establishment of a layer of cartilage at the ends of long bones is intimately linked to the process of endochondral ossification. Previous in vivo studies and computer models suggest that mechanobiological factors can play a key role in modulating cartilage growth and ossification. Specifically, intermittent hydrostatic pressure is thought to maintain cartilage, and shear stresses encourage cartilage destruction and ossification. In the present investigation we examined the combined effects of hydrostatic pressure and shear stress--in the form of an osteogenic index--on the development of a layer of articular cartilage, using an idealized finite element computer model. The results of our analyses provide further support for the view that mechanobiological factors play a key role in regulating the distribution of cartilage thickness and in maintaining a stable cartilage layer at maturity. The model predicts that joints that experience higher contact pressures will have thicker cartilage layers. These predictions are consistent with observations of cartilage thickness in both humans and animals. Variations in articular mechanical load are predicted to modulate cartilage thickness. These results are consistent with the view that the mechanobiological factors responsible for the development of diarthrodial joints eventually lead to cartilage degeneration and osteoarthritis (OA) with aging. PMID- 10850821 TI - Time-dependent effects of intermittent hydrostatic pressure on articular chondrocyte type II collagen and aggrecan mRNA expression. AB - The normal loading of joints during daily activities causes the articular cartilage to be exposed to high levels of intermittent hydrostatic pressure. This study quantified effects of intermittent hydrostatic pressure on expression of mRNA for important extracellular matrix constituents. Normal adult bovine articular chondrocytes were isolated and tested in primary culture, either as high-density monolayers or formed aggregates. Loaded cells were exposed to 10 MPa of intermittent hydrostatic pressure at a frequency of 1 Hz for periods of 2, 4, 8, 12, and 24 hrs. Other cells were intermittently loaded for a period of 4 hrs per day for 4 days. Semiquantitative reverse transcription polymerase chain reaction assays were used to assess mRNA signal levels for collagen types II and I and aggrecan. The results showed that type II collagen mRNA signal levels exhibited a biphasic pattern, with an initial increase of approximately five-fold at 4 and 8 hrs that subsequently decreased by 24 hrs. In contrast, aggrecan mRNA signal increased progressively up to three-fold throughout the loading period. Changing the loading profile to 4 hrs per day for 4 days increased the mRNA signal levels for type II collagen nine-fold and for aggrecan twenty-fold when compared to unloaded cultures. These data suggest that specific mechanical loading protocols may be required to optimally promote repair and regeneration of diseased joints. PMID- 10850822 TI - Methods for evaluating the progression of osteoarthritis. AB - This article discusses methods for evaluating the progression of osteoarthritis through dynamic functional imaging as opposed to current static techniques. Comparison is made between static and dynamic methods of evaluating knee alignment. The correlation between dynamic knee moments during gait and bone mineral content is discussed. Knee loading is considered in terms of high tibial osteotomy, knee braces, pain, and non-steroidal anti-inflammatory drugs. New image-processing techniques for quantitating cartilage loss are described, and computational methods for generating true three-dimensional (3-D) maps of cartilage thickness are developed. Finally, new approaches to cross-correlate magnetic resonance images with kinematic measurements are described. These new techniques promise to become powerful diagnostic tools to detect and characterize pathological load distributions across articular cartilage. PMID- 10850823 TI - In vitro engineering of cartilage. AB - Because adult human cartilage shows poor capacity for repair and regeneration, innovative solutions are required for congenital and acquired degenerative cartilage lesions. Acquired lesions occur in young and old alike, the former being more at risk for sports-related injuries and the latter for age-related degenerative changes. Because cartilage is a relatively simple tissue with respect to its cellular homogeneity and avascularity, it has been a model for research of in vitro engineered tissues. Progress has been slow and obstructed on several levels. The adult chondrocyte has limited capacity for proliferation and has both catabolic and anabolic functions. These metabolic features must be controlled in order for engineered tissue to endure. Use of three-dimensional scaffolds can be combined with regulatory factors (cytokine, extracellular matrix [ECM], and mechanical) to optimize conditions for in vitro engineered cartilage. Cross-disciplinary interactions are likely to accelerate progress and to mediate application of advances made in other fields for consistently successful in vitro engineering of cartilage for all clinical needs. PMID- 10850824 TI - Continuous passive motion (CPM): theory and principles of clinical application. AB - Stiffness following surgery or injury to a joint develops as a progression of four stages: bleeding, edema, granulation tissue, and fibrosis. Continuous passive motion (CPM) properly applied during the first two stages of stiffness acts to pump blood and edema fluid away from the joint and periarticular tissues. This allows maintenance of normal periarticular soft tissue compliance. CPM is thus effective in preventing the development of stiffness if full motion is applied immediately following surgery and continued until swelling that limits the full motion of the joint no longer develops. This concept has been applied successfully to elbow rehabilitation, and explains the controversy surrounding CPM following knee arthroplasty. The application of this concept to clinical practice requires a paradigm shift, resulting in our attention being focused on preventing the initial or delayed accumulation of periarticular interstitial fluids. PMID- 10850825 TI - Osteoporosis and bone functional adaptation: mechanobiological regulation of bone architecture in growing and adult bone, a review. AB - During life, bone is continually optimized for its load-bearing role by a process of functionally adaptive (re)modelling. This process, which is more active in growing bone, is dominated by high-magnitude, high-rate strains, presented in an unusual distribution. Adaptation occurs at an organ level, involving changes in whole bone architecture and bone mass. The repetitive coordinated bone loading associated with habitual activity may have little role in the preservation of bone mass, and may even reduce the osteogenic potential of an otherwise highly osteogenic stimulus. Cells of the osteocyte/osteoblast network are best placed to appreciate mechanical strain. Among the strain-related responses they show, is a reduced rate of apoptosis. This may serve to regulate and target osteoclast activity. A more complete understanding of the stimuli and pathways involved in both the physiology and pathology of this structural homeostatic mechanism will allow the design of more appropriate exercise regimens and targeted pharmacological interventions to limit morbidity and mortality by reducing bone fragility. PMID- 10850826 TI - Mechanobiology of femoral neck structure during adolescence. AB - Understanding femoral neck structure may be critical to preventing fractures at this site. We examined the correlates of changes in the femoral neck during adolescence. Dual energy x-ray absorptiometry measurements of proximal femora were made in 101 Caucasian youths (ages 9 to 26 years). Relationships were examined between developmental parameters (age, pubertal stage, height, body mass, lean mass, and fat mass) and femoral structure (bone mineral content, bone mineral density, neck width, cross-sectional area, and cross-sectional strength). Lean body mass was the best predictor of femoral neck structure, explaining 53-87 percent of the variance, and was independent of gender. Body mass only explained 51-79 percent of the variance. Previously we found body mass to be the strongest predictor of femoral mid-diaphyseal cross-sectional properties. These findings suggest that trabecular bone of the femoral neck may be more responsive to its mechanical environment than the cortical diaphysis. In addition, lean body mass may be a more reliable predictor of muscle loading than body mass. PMID- 10850827 TI - The mechanobiology of cancellous bone structural adaptation. AB - The distinguishing morphological feature of cancellous bone is its high level of porosity relative to cortical bone. This porosity leads to more free surfaces and thus to more of the cellular constituents that inhabit those surfaces. As a result, cancellous bone is often more metabolically active and responsive to stimuli than cortical bone. This extends to the relationship between cancellous bone's internal structure and external mechanical loading. Observational investigations established this relationship as early as the late 19th century. These findings point to the interplay between biology and the cellular mechanical environment, forming the underpinnings of the modern term mechanobiology. Interestingly, it has proven to be more straightforward to assay the biological response than to quantify the precise mechanical environment of cancellous bone and the influence of cancellous bone structure. Despite this concern, significant insights into the nature of cancellous bone mechanobiology can be obtained from computational simulations that allow investigators to determine the morphological consequences of quantitative assumptions about cancellous bone mechanobiology. As the power of computers and the sophistication of these modeling techniques continue to grow, we can expect an increased impact in terms of clinical diagnosis and treatment. The next decade will bring improvements in exercise interventions to prevent and reverse bone loss; improved replacement-joint designs, particularly for those joints currently having poor expected outcomes; and an integration of computer simulation technology with clinical scanners. PMID- 10850828 TI - Tendon and ligament adaptation to exercise, immobilization, and remobilization. AB - This study provides a theoretical and computational basis for understanding and predicting how tendons and ligaments adapt to exercise, immobilization, and remobilization. In a previous study, we introduced a model that described the growth and development of tendons and ligaments. In this study, we use the same model to predict changes in the cross-sectional area, modulus, and strength of tendons and ligaments due to increased or decreased loading. The model predictions are consistent with the results of experimental exercise and immobilization studies performed by other investigators. These results suggest that the same fundamental principles guide both development and adaptation. A basic understanding of these principles can contribute both to prevention of tendon and ligament injuries and to more effective rehabilitation when injury does occur. PMID- 10850829 TI - Bone mineral and geometric changes through the femur with immobilization due to spinal cord injury. AB - This cross-sectional study describes bone mineral and geometric properties of the midshaft and distal femur in a control population and examines effects of immobilization due to spinal cord injury (SCI) at these skeletal sites. The subject populations were comprised of 118 ambulatory adults (59 men and 59 women) and 246 individuals with SCI (239 men and 7 women); 30 of these were considered to have acute injury (SCI duration <1 year). Bone mineral density (BMD) was assessed at the femoral neck, and midshaft and distal femur by dual energy absorptiometry. Geometric properties, specifically cortical area, polar moment of inertia, and polar section modulus, were estimated at the midshaft from cortical dimensions obtained by concurrent radiography. Reduction in BMD was noted in all femoral regions (27%, 25%, and 43% for femoral neck, midshaft, and distal femur, respectively) compared with controls. In contrast, although endosteal diameter was enlarged, geometric properties were not significantly reduced in the midshaft attributable to the age-related increase in periosteal diameter. These results suggest that simultaneous assessment of bone mineral and geometric properties may improve clinically relevant evaluation of skeletal status. PMID- 10850830 TI - A model of mechanobiologic and metabolic influences on bone adaptation. AB - Bone adaptation, the process through which bone mass is modified in the body, plays a key role in the development of osteoporosis. Bone adaptation is known to be influenced by both mechanical and metabolic stimuli. Previous studies have concentrated on changes in bone adaptation caused by mechanical stimuli (mechanobiologic influences), yet current treatments for osteoporosis depend significantly on metabolic influences. We develop a theoretical model of bone adaptation that accounts for both mechanobiologic and metabolic influences. We demonstrate the utility of this model using a simulation of the cellular processes of bone adaptation on a representative volume of cancellous bone. Our long-term objective is the development of a more comprehensive computational model that will aid in the study of osteoporosis and other bone diseases. PMID- 10850831 TI - Musculoskeletal health and the older adult. AB - A decline in muscle mass and function, and in the mass and integrity of the skeletal system, are well-known consequences of aging. These changes impinge on the functional performance required for independent living and contribute to frailty and fracture risk. Resistance exercise has been shown to be an effective mode to circumvent age-related changes in the muscular system, although the benefit of exercise on bone mass in the aging skeleton is comparatively modest at best. This brief review highlights results from several studies that we have undertaken in older adults, examining aspects of the resistance training prescription as well as the potential beneficial role of hormones. What is common to all of these studies is the high degree of residual plasticity that remains in aging skeletal muscle. Risk factors for falls and fracture include reduced bone mass, muscle weakness, impaired balance, and lessened visual acuity. Among these, only muscle strength is reliably enhanced with resistance exercise, which may aid in reducing hip fracture risk as well as improving the ability to undertake daily activities and maintain independence. PMID- 10850832 TI - Gait and speed as exercise components of risk factors associated with onset of fatigue injury of the third metacarpal bone in 2-year-old Thoroughbred racehorses. AB - OBJECTIVE: To determine the degree to which components of the training program of 2-year-old Thoroughbred racehorses influence their susceptibility to fatigue injury of the third metacarpal bone (bucked shins). ANIMALS: 226 two-year-old Thoroughbred racehorses. PROCEDURE: Daily training information and health reports on 2-year-old Thoroughbreds were compiled from records provided from 5 commercial stables. For each horse, data (exercise variables) were collected that comprised distance jogged (approx speed of 5 m/s), galloped (approx 11 m/s), and breezed (approx 15 to 16 m/s) until a single instance of bucked shins was reported. Data were coded for analysis using cross-tabulation, graphic, and survival techniques. RESULTS: Of 226 horses, 56 had bucked shins, 9 completed the observation period without bucked shins, and 161 were lost to follow-up. Distinct training strategies were used at stables resulting in significantly different survival profiles among stables. Mean (+/- SD) allocation of exercise to breezing was 0.15 +/- 0.13 miles/wk (maximum, 0.64 miles/wk), to galloping was 4.47 +/- 1.52 miles/wk (maximum, 9.56 miles/wk), and to jogging was 2.34 +/- 1.70 miles/wk (maximum, 8.53 miles/wk). Survival (ie, lack of bucked shins during 1 year of monitoring) was found to be significantly reduced by exercise allocation to breezing, significantly increased by exercise allocation to galloping, and uninfluenced by exercise allocation to jogging. The log of the hazard ratio was reduced by 4.2 +/- 1.5/mile breezed and increased by 0.3 +/- 0.1/mile galloped. CONCLUSIONS AND CLINICAL RELEVANCE: Relationships between different gaits and speeds in the training regimen influence the incidence of bucked shins. To reduce the incidence of bucked shins, trainers should consider allocating more training effort to regular short-distance breezing and less to long-distance galloping. PMID- 10850834 TI - Changes in kinematic variables observed during pressure-induced forelimb lameness in adult horses trotting on a treadmill. AB - OBJECTIVE: To determine whether kinematic changes induced by heel pressure in horses differ from those induced by toe pressure. ANIMALS: 10 adult Quarter Horses. PROCEDURE: A shoe that applied pressure on the cuneus ungulae (frog) or on the toe was used. Kinematic analyses were performed before and after 2 levels of frog pressure and after 1 level of toe pressure. Values for stride displacement and time and joint angles were determined from horses trotting on a treadmill. RESULTS: The first level of frog pressure caused decreases in metacarpophalangeal (fetlock) joint extension during stance and increases in head vertical movement and asymmetry. The second level of frog pressure caused these changes but also caused decreases in stride duration and carpal joint extension during stance as well as increases in relative stance duration. Toe pressure caused changes in these same variables but also caused maximum extension of the fetlock joint to occur before midstance, maximum hoof height to be closer to midswing, and forelimb protraction to increase. CONCLUSION AND CLINICAL RELEVANCE: Decreased fetlock joint extension during stance and increased head vertical movement and asymmetry are sensitive indicators of forelimb lameness. Decreased stride duration, increased relative stance duration, and decreased carpal joint extension during stance are general but insensitive indicators of forelimb lameness. Increased forelimb protraction, hoof flight pattern with maximum hoof height near midswing, and maximum fetlock joint extension in cranial stance may be specific indicators of lameness in the toe region. Observation of forelimb movement may enable clinicians to differentiate lameness of the heel from lameness of the toe. PMID- 10850833 TI - Concentrations of trace minerals in the spinal cord of horses with equine motor neuron disease. AB - OBJECTIVE: To compare concentrations of trace minerals in the spinal cord of horses with equine motor neuron disease (EMND) with those of horses without neurologic disease (control horses). ANIMALS: 24 horses with EMND and 22 control horses. PROCEDURE: Spinal cord trace mineral concentrations in horses with EMND and control horses were analyzed by use of inductively coupled plasma atomic emission spectroscopy (calcium, phosphorus, sodium, potassium, magnesium, copper, iron, manganese, nickel, zinc, aluminum, cobalt, and chromium), atomic absorption spectrophotometry (lead and cadmium), flameless atomic absorption (mercury), and fluorometry (selenium). RESULTS: Copper concentration was significantly higher in the spinal cord of horses with EMND, compared with control horses; spinal cord concentrations of all other trace minerals were similar between groups. CONCLUSION AND CLINICAL RELEVANCE: Among spinal cord trace minerals investigated in the study, only copper concentrations were significantly different between horses with EMND and horses without neurologic disease, which suggests that copper may be involved in the pathogenesis of EMND. An hypothesis of oxidative injury in this disease is supported by the finding of increased copper concentrations in the spinal cord and by low vitamin E concentrations reported by other researchers. PMID- 10850835 TI - Development and validation of an enzyme-linked immunosorbent assay for feline trypsin-like immunoreactivity. AB - OBJECTIVE: To develop and validate an ELISA for quantitative analysis of feline trypsin-like immunore-activity (fTLI). SAMPLE POPULATION: Purified feline cationic trypsin (fCT) and rabbit anti-fCT antiserum; blood samples from 63 healthy cats. PROCEDURES: A sandwich capture ELISA was developed, using anti-fCT antiserum purified by affinity chromatography that underwent biotinylation. Purified fCT was used for standards. The assay was validated by determination of sensitivity, working range, linearity, accuracy, precision, and reproducibility. A reference range was established by assaying serum samples from the 63 healthy cats. RESULTS: Sensitivity was 1.23 microg/L; working range was 2 to 567 microg/L. Ratios of observed versus expected results for 4 samples tested at various dilutions ranged from 90.0 to 120.7%. Ratios of observed versus expected results for 5 samples spiked with various concentrations of fCT ranged from 82.0 to 101.8%. Intra- and inter-assay coefficients of variability ranged from 9.9 to 11.1% and from 10.2 to 21.7%, respectively. The reference range for serum fTLI measured with this ELISA was 12 to 82 microg/L. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that an ELISA can be used to measure serum fTLI in cats. The ELISA was sufficiently sensitive, linear, accurate, precise, and reproducible for clinical use. PMID- 10850836 TI - Effects of interleukin-1beta and tumor necrosis factor-alpha on expression of matrix-related genes by cultured equine articular chondrocytes. AB - OBJECTIVE: To determine the effects of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) on expression and regulation of several matrix related genes by equine articular chondrocytes. SAMPLE POPULATION: Articular cartilage harvested from grossly normal joints of 8 foals, 6 yearling horses, and 8 adult horses. PROCEDURE: Chondrocytes maintained in suspension cultures were treated with various doses of human recombinant IL-1beta or TNF-alpha. Northern blots of total RNA from untreated and treated chondrocytes were probed with equine complementary DNA (cDNA) probes for cartilage matrix-related genes. Incorporation of 35S-sulfate, fluorography of 14C-proline labeled medium, zymography, and western blotting were used to confirm effects on protein synthesis. RESULTS: IL-1beta and TNF-alpha increased steady-state amounts of mRNA of matrix metalloproteinases 1, 3, and 13 by up to 100-fold. Amount of mRNA of tissue inhibitor of metalloproteinase-1 also increased but to a lesser extent (1.5- to 2-fold). Amounts of mRNA of type-II collagen and link protein were consistently decreased in a dose-dependent manner. Amount of aggrecan mRNA was decreased slightly; amounts of biglycan and decorin mRNA were minimally affected. CONCLUSIONS AND CLINICAL RELEVANCE: Treatment of cultured equine chondrocytes with IL-1beta or TNF-alpha resulted in marked alterations in expression of various matrix and matrix-related genes consistent with the implicated involvement of these genes in arthritis. Expression of matrix metalloproteinases was increased far more than expression of their putative endogenous inhibitor. Results support the suggestion that IL-1beta and TNF-alpha play a role in the degradation of articular cartilage in arthritis. PMID- 10850837 TI - Pharmacokinetics of pentoxifylline in dogs after oral and intravenous administration. AB - OBJECTIVE: To evaluate the pharmacokinetics of pentoxifylline (PTX) and its 5 hydroxyhexyl-metabolite, metabolite 1 (M1), in dogs after IV administration of a single dose and oral administration of multiple doses. ANIMALS: 7 sexually intact, female, mixed-breed dogs. PROCEDURE: A crossover study design was used so that each of the dogs received all treatments in random order. A drug-free period of 5 days was allowed between treatments. Treatments included IV administration of a single dose of PTX (15 mg/kg of body weight), oral administration of PTX with food at a dosage of 15 mg/kg (q 8 h) for 5 days, and oral administration of PTX without food at a dosage of 15 mg/kg (q 8 h) for 5 days. Blood samples were taken at 0.25, 0.5, 1, 1.5, 2, 2.5, and 3 hours after the first and last dose of PTX was administered PO, and at 5, 10, 20, 40, 80, and 160 minutes after PTX was administered IV. RESULTS: PTX was rapidly absorbed and eliminated after oral administration. Mean bioavailability after oral administration ranged from 15 to 32% among treatment groups and was not affected by the presence of food. Higher plasma PTX concentrations and apparent bioavailability were observed after oral administration of the first dose, compared with the last dose during the 5-day treatment regimens. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs, oral administration of 15 mg of PTX/kg results in plasma concentrations similar to those produced by therapeutic doses in humans, and a three-times-a-day dosing regimen is the most appropriate. PMID- 10850838 TI - Effects of warm-up intensity on kinetics of oxygen consumption and carbon dioxide production during high-intensity exercise in horses. AB - OBJECTIVE: To compare effects of low and high intensity warm-up exercise on oxygen consumption (VO2) and carbon dioxide production (VCO2) in horses. ANIMALS: 6 moderately conditioned adult Standard-breds. PROCEDURES: Horses ran for 2 minutes at 115% of maximum oxygen consumption (VO2max), 5 minutes after each of the following periods: no warm-up (NoWU); 10 minutes at 50% of VO2max (LoWU); or 7 minutes at 50% VO2max followed by 45-second intervals at 80, 90, and 100% VO2max (HiWU). Oxygen consumption and VCO2 were measured during exercise, and kinetics of VO2 and VCO2 were calculated. Accumulated O2 deficit was also calculated. RESULTS: For both warm-up trials, the time constant for the rapid exponential increase in VO2 was 30% lower than for NoWU. Similarly, the rate of increase in VCO2 was 23% faster in LoWU and HiWU than in NoWU. Peak values for VO2 achieved during the high-speed test were not significantly different among trials (LoWU, 150.2 +/- 3.2 ml/kg/min; HiWU, 151.2 +/- 4.2 ml/kg/min; NoWU, 145.1 +/- 4.1 ml/kg/min). However, accumulated O2 deficit (ml of O2 equivalents/kg) was significantly lower during LoWU (65.3 +/- 5.1) and HiWU (63.4 +/- 3.9) than during NoWU (82.1 +/- 7.3). CONCLUSIONS AND CLINICAL RELEVANCE: Both the low- and high-intensity warm-up, completed 5 minutes before the start of high-intensity exercise, accelerated the kinetics of VO2 and VCO2 and decreased accumulated O2 deficit during 2 minutes of intense exertion in horses that were moderately conditioned. PMID- 10850839 TI - Effects of treatment with growth hormone and somatostatin on efficacy of diammine [1,1-cyclobutane dicarboxylato (2-)-0,0']-(SP-4-2) in athymic rats with osteosarcoma. AB - OBJECTIVE: To determine the effect of exogenous growth hormone or somatostatin on chemotherapeutic efficacy in athymic (nude) rats with osteosarcoma. ANIMALS: 66 female athymic rats. PROCEDURE: Osteosarcoma was induced at an intratibial site. Rats were randomly allotted to 6 treatment groups. Rats were treated with saline (0.9% NaCl) solution alone, platinum, diammine [1,1-cyclobutane dicaboxylato (2-) 0,0']-(SP-4-2) (CBDCA; ie, carboplatin) plus saline solution, somatostatin alone, somatostatin plus CBDCA, growth hormone alone, or growth hormone plus CBDCA. Variables measured included estimated WBC count and percentage of neutrophils, plasma concentration of insulin-like growth factor I (IGF-I), body weight, tumor volume, weight of primary tumor, survival time, and distant metastasis at time of death. RESULTS: Tumors formed at the injection sites in all rats. Treatment with growth hormone increased, and treatment with somatostatin decreased, plasma IGF-I concentration. Treatment with growth hormone or somatostatin altered CBDCA efficacy, as determined by evaluation of mean and median survival times. Metastatic pulmonary disease developed in 63 of 64 rats. CONCLUSIONS AND CLINICAL RELEVANCE: The technique used here reliably induced local osteosarcomas and metastatic pulmonary disease. Treatment with growth hormone and CBDCA or somatostatin may improve chemotherapeutic efficacy without increasing toxic effects. IMPLICATIONS FOR HUMAN MEDICINE: Results reported here may be useful in the study of osteosarcoma in humans. PMID- 10850840 TI - Comparison of plasma benzodiazepine concentrations following intranasal and intravenous administration of diazepam to dogs. AB - OBJECTIVE: To determine whether plasma concentrations of benzodiazepines (BDZ) in dogs following intranasal (IN) administration of diazepam are comparable to concentrations following IV administration. ANIMALS: 6 (4 male, 2 female) healthy adult Greyhounds. PROCEDURE: Dogs were randomly assigned to 2 groups of 3 dogs in a crossover design. Diazepam (0.5 mg/kg of body weight) was administered intravenously to dogs in group 1 and intranasally to dogs in group 2. Blood was collected from the jugular vein of each dog into tubes containing lithium heparin before and 3, 6, 9, 12, 15, 20, 30, 60, 120, 240, and 480 minutes following diazepam administration. After a 4-day washout period, dogs in group 1 received diazepam intranasally, dogs in group 2 received diazepam intravenously, and blood was again collected. Plasma concentration of BDZ was determined by use of a fluorescence polarization immunoassay. RESULTS: Mean (+/- SD) peak plasma concentration of BDZ following IV administration (1,316 +/- 216 microg/L) was greater than that following IN administration (448 +/- 41 microg/L). Time to peak concentration was < or = 3 minutes following IV administration and 4.5 +/- 1.5 minutes following IN administration. Mean bioavailability of BDZ following IN administration was 80 +/- 9%. CONCLUSIONS AND CLINICAL RELEVANCE: Diazepam is rapidly and efficiently absorbed following IN administration of the parenteral formulation. Plasma concentrations match or exceed the suggested therapeutic concentration (300 microg/L). Intranasal administration of diazepam may be useful for treatment of seizures in dogs by owners or when intravenous access is not readily available. PMID- 10850841 TI - Esophageal transit of capsules in clinically normal cats. AB - OBJECTIVE: To evaluate the esophageal passage of capsules in clinically normal cats and determine the incidence of prolonged transit or entrapment. ANIMALS: 12 clinically normal adult cats. PROCEDURE: Esophageal transit of barium sulfate filled capsules was evaluated fluoroscopically. Each cat was examined 3 times (36 examinations). Esophageal transit times were classified as normal (< or = 30 seconds) or prolonged (> 30 but < or = 240 seconds). Capsules were considered entrapped when transit times were > 240 seconds. RESULTS: Transit times were normal in 10 of the 36 (27.8%) examinations, whereas times were prolonged in 7 (19.4%) examinations. Capsules became entrapped in the midcervical region of the esophagus during 19 (52.8%) examinations. Following termination of each examination, cats with entrapped capsules were fed a small amount (0.5 to 1 ounce) of food; this resulted in passage of the capsule to the stomach. CONCLUSIONS AND CLINICAL RELEVANCE: The possibility of medication-induced esophagitis should be considered when orally administering ulcerogenic drugs to cats. It is recommended that a small volume of food be given following medications to ensure complete esophageal clearance. PMID- 10850842 TI - Development of a method for determination of cyanide concentrations in serum and rumen fluid of cattle. AB - OBJECTIVES: To develop a reliable method for measurement of cyanide concentrations in cattle, using gas chromatography-mass spectrometry (GC-MS), and establish reference ranges of cyanide concentrations in cattle. ANIMALS: 52 Fleckvieh cattle. PROCEDURE: Cattle were allocated to 3 groups; 12 were fed leguminous grass and hay, 36 were fed whole-maize and corn-cob silages, and 4 were fed other feedstuffs. Samples of blood, rumen fluid, and liver were collected at time of slaughter. Serum, rumen fluid, and liver homogenate were assayed for cyanide content, using a derivatization procedure. A technique for analysis by GC-MS that used selected ion monitoring was developed. RESULTS: Compared with a spectrophotometric method, detection of cyanide in serum and rumen fluid by use of GC-MS was selective and sensitive, with a limit of detection of 0.7 microM. Spectrophotometric analysis yielded false-negative and false-positive results. Thus, the GC-MS method was used for subsequent analysis. In all cattle except 1, cyanide concentration ranged from < 0.7 to 35 microM in serum and from < 0.7 to 28 microM in rumen fluid; cyanide concentration in that 1 animal was 206 microM. Cattle fed clover, grass, grass hay, and clover hay had 8.3- to 8.6-fold higher mean cyanide concentrations in rumen fluid and serum than cattle fed whole-maize and corn-cob silages. CONCLUSIONS AND CLINICAL RELEVANCE: Results of this study suggest a reference range that should be useful for aiding in the diagnosis of cyanide poisoning. Also, cattle can apparently accommodate a serum cyanide concentration of 206 microM without adverse effects. PMID- 10850843 TI - Effects of a 98% solution of glycerol or sterilization with ethylene oxide on FeLV in bone allografts and effects on bone incorporation of allografts in cats. AB - OBJECTIVES: To compare virucidal effects and bone incorporation properties of cortical bone allografts transplanted into specific-pathogen-free (SPF) cats. Allografts consisted of untreated bone from a SPF cat (negative-control group) and bone from 5 FeLV-infected cats that was subjected to sterilization with ethylene oxide (ETO), preservation with glycerol, or no treatment (positive control group). SAMPLE POPULATION: Bones from the aforementioned groups and twenty 8-week-old SPF cats (5 cats/group) implanted with an allograft from 1 of the aforementioned groups. PROCEDURE: After implantation, blood samples were collected weekly to monitor FeLV p27 antigen and antibody titers. Quantification of FeLV provirus was performed on blood samples at weeks 0, 4, and 8 and donor bone samples at time of implantation. Cats were euthanatized 8 weeks after transplantation, and graft sites were evaluated. RESULTS: All results for negative-control cats were negative. All ETO group cats had negative results for antigen and provirus in blood, whereas 1 cat had a low antibody titer. Although 3 ETO-treated allografts were positive for provirus, the DNA appeared denatured. One cat in the glycerol group had positive results for all tests in blood samples. All glycerol-preserved allografts were positive when tested for provirus. All results for positive-control group cats were positive. Differences in incorporation of bone grafts were not observed. CONCLUSIONS AND CLINICAL RELEVANCE: Glycerol preservation of FeLV-infected bone allografts did not eliminate transmission of retrovirus to recipients. In contrast, ETO sterilization appeared to denature DNA and prevent infection. Treatments did not affect incorporation of bone grafts in young cats. PMID- 10850844 TI - Comparison of pharmacokinetics of fentanyl after intravenous and transdermal administration in cats. AB - OBJECTIVE: To compare pharmacokinetic and pharmacodynamic characteristics of fentanyl citrate after IV or transdermal administration in cats. ANIMALS: 6 healthy adult cats with a mean weight of 3.78 kg. PROCEDURE: Each cat was given fentanyl IV (25 mg/cat; mean +/- SD dosage, 7.19 +/- 1.17 mg/kg of body weight) and via a transdermal patch (25 microg of fentanyl/h). Plasma concentrations of fentanyl were measured by use of radioimmunoassay. Pharmacokinetic analyses of plasma drug concentrations were conducted, using an automated curve-stripping process followed by nonlinear, least-squares regression. Transdermal delivery of drug was calculated by use of IV pharmacokinetic data. RESULTS: Plasma concentrations of fentanyl given IV decreased rapidly (mean elimination half life, 2.35 +/- 0.57 hours). Mean +/- SEM calculated rate of transdermal delivery of fentanyl was 8.48 +/- 1.7 mg/h (< 36% of the theoretical 25 mg/h). Median steady-state concentration of fentanyl 12 to 100 hours after application of the transdermal patch was 1.58 ng/ml. Plasma concentrations of fentanyl < 1.0 ng/ml were detected in 4 of 6 cats 12 hours after patch application, 5 of 6 cats 18 and 24 hours after application, and 6 of 6 cats 36 hours after application. CONCLUSIONS AND CLINICAL RELEVANCE: In cats, transdermal administration provides sustained plasma concentrations of fentanyl citrate throughout a 5-day period. Variation of plasma drug concentrations with transdermal absorption for each cat was pronounced. Transdermal administration of fentanyl has potential for use in cats for long-term control of pain after surgery or chronic pain associated with cancer. PMID- 10850845 TI - Effect of sodium butyric acid, sodium valerianic acid, and osmolarity on contractility of specimens of intestinal wall obtained from the cecum and spiral colon of healthy cows. AB - OBJECTIVE: To compare the effect of various concentrations of sodium butyric acid and sodium valerianic acid, as well as various osmolarities, on contractility of ex-vivo intestinal wall specimens obtained from the cecum and spiral colon of each of several healthy cows. SAMPLE POPULATION: Full-thickness preparations of intestinal wall, dissected parallel to the longitudinal smooth muscle layers, harvested from freshly slaughtered healthy cows. PROCEDURE: Specimens of intestinal wall were incubated for 5 minutes with various concentrations of sodium butyric acid and sodium valerianic acid as well as various osmolar concentrations of NaCl, using a crossover design. Isometric contractions were induced 7 times with carbachol (CH; 5 X 10(-6) mol/L). Contractility was defined as the maximum amplitude of contraction and the amplitude of contraction 2 minutes after addition of CH. RESULTS: Repeated addition of CH did not result in a significant effect on contractility of specimens from the cecum and spiral colon. Contractility after addition of CH was not significantly affected by prior incubation with various concentrations of sodium butyric acid or sodium valerianic acid or after an increase of osmolarity. Maximum amplitude of contraction was significantly higher in specimens from the spiral colon, compared with specimens from the cecum. CONCLUSIONS: Increases in concentrations of sodium butyric acid or sodium valerianic acid and increases in osmolarity did not inhibit contractility of intestinal wall specimens from the cecum and spiral colon of a group of healthy cows. PMID- 10850846 TI - Pathophysiologic correlates of acute porcine pleuropneumonia. AB - OBJECTIVE: To develop and evaluate an in vivo model to study early events in the pathogenesis of acute porcine pleuropneumonia. ANIMALS: Thirty-six 6- to 8-week old pigs. PROCEDURE: Pigs were inoculated intranasally or endotracheally with Actinobacillus pleuropneumoniae; inoculation routes were compared by evaluation of clinical signs, gross and microscopic lung lesions, hematologic changes, serum zinc, iron, and haptoglobin concentrations, and inflammatory cytokines. RESULTS: The 2 inoculation routes resulted in similar findings, although intranasal inoculation caused unilateral gross lung lesions, whereas endotracheal inoculation caused bilateral gross lesions. Clinical signs of disease were observed < 2 hours after endotracheal inoculation and 6 to 8 hours after intranasal inoculation. Total WBC counts did not differ significantly after inoculation by either inoculation route, although band neutrophils increased significantly. The earliest findings associated with A pleuropneumoniae inoculation, irrespective of route, were decreased serum zinc and iron concentrations. Serum haptoglobin concentrations were significantly increased after inoculation. Inoculation induced rapid influx of macrophages into the lung and local induction of proinflammatory cytokines. Northern blot analysis of total RNA from lung tissue indicated that inoculated pigs had increased concentrations of interleukin (IL)-1beta, IL-1alpha, and IL-8; tumor necrosis factor messenger RNA concentration was not increased. CONCLUSIONS: Endotracheal inoculation with A pleuropneumoniae rapidly and consistently induced diffuse bilateral pneumonia; thus, this method may be useful for the study of acute pathophysiologic changes associated with bacterial pneumonia and may provide an experimental model for testing modalities for prevention and treatment of this and other respiratory tract diseases of pigs. PMID- 10850847 TI - Effect of hyperbaric oxygen treatment on incorporation of an autogenous cancellous bone graft in a nonunion diaphyseal ulnar defect in cats. AB - OBJECTIVE: To determine whether hyperbaric oxygen treatment (HBOT) would affect incorporation of an autogenous cancellous bone graft in diaphyseal ulnar defects in cats. ANIMALS: 12 mature cats. PROCEDURE: Bilateral nonunion diaphyseal ulnar defects were created in each cat. An autogenous cancellous bone graft was implanted in 1 ulnar defect in each cat, with the contralateral ulnar defect serving as a nongrafted specimen. Six cats were treated by use of hyperbaric oxygen at 2 atmospheres absolute for 90 minutes once daily for 14 days, and 6 cats were not treated (control group). Bone labeling was performed, using fluorochrome markers. Cats were euthanatized 5 weeks after implanting, and barium sulfate was infused to evaluate vascularization of grafts. Ulnas were evaluated by use of radiography, microangiography, histologic examination, and histomorphometric examination. RESULTS: Radiographic scores did not differ between treatment groups. Microangiographic appearance of grafted defects was similar between groups, with all having adequate vascularization. Differences were not observed between treated and nontreated groups in the overall histologic appearance of decalcified samples of tissue in grafted defects. Mean distance between fluorescent labels was significantly greater in cats given HBOT than in nontreated cats. Median percentage of bone formation in grafted defects was significantly greater in cats given HBOT. CONCLUSIONS: Hyperbaric oxygen treatment increased the distance between fluorescent labels and percentage of bone formation when incorporating autogenous cancellous bone grafts in induced nonunion diaphyseal ulnar defects in cats, but HBOT did not affect revascularization, radiographic appearance, or qualitative histologic appearance of the grafts. PMID- 10850848 TI - Use of repetitive sequence-based polymerase chain reaction for molecular epidemiologic analysis of Streptococcus equi subspecies equi. AB - OBJECTIVE: To determine whether repetitive sequence-based polymerase chain reaction (rep-PCR) could be used to differentiate Streptococcus equi isolates, to examine S equi isolates from throughout the world, and to determine whether a horse had > 1 subtype of S equi during an outbreak of disease. SAMPLE POPULATION: An initial group of 32 S equi isolates, 63 S equi isolates from various geographic areas, and 17 S equi isolates obtained during outbreaks of disease. PROCEDURE: An aliquot of S equi genomic DNA was amplified, using enterobacterial repetitive intergenic consensus primers. Gel electrophoresis was performed on 1.5% agarose gels, and a computed-assisted program was used to compare rep-PCR results. RESULTS: Use of these primers to analyze 100 ng of S equi genomic DNA resulted in patterns of 6 to 14 bands. The 32 initial isolates were separated into 7 rep-PCR subtypes. There were 30 rep-PCR subtypes found among 29 S equi isolates obtained from Minnesota, Michigan, Canada, and Australia and 34 S equi isolates obtained from Kentucky and other sources. Furthermore, the same clone was identified in several horses during an outbreak of disease. Infected horses on the same farm all had a single clone of S equi. CONCLUSION AND CLINICAL RELEVANCE: Analysis of these results suggests that rep-PCR is useful for delineating S equi into rep-PCR subtypes. Results revealed that isolates with the same geographic source or similar date of collection did not always have the same rep-PCR subtype. A single clone of S equi usually predominated during an outbreak of disease. PMID- 10850849 TI - Pharmacokinetics of enrofloxacin administered intravenously and orally to foals. AB - OBJECTIVE: To determine the pharmacokinetics of enrofloxacin administered IV and orally to foals. ANIMALS: 5 clinically normal foals. PROCEDURE: A 2-dose cross over trial with IV and oral administration was performed. Enrofloxacin was administered once IV (5 mg/kg of body weight) to 1-week-old foals, followed by 1 oral administration (10 mg/kg) after a 7-day washout period. Blood samples were collected for 48 hours after the single dose IV and oral administrations and analyzed for plasma enrofloxacin and ciprofloxacin concentrations by use of high performance liquid chromatography. RESULTS: For IV administration, mean +/- SD total area under the curve (AUC0-infinity) was 48.54 +/- 10.46 microg x h/ml, clearance was 103.72 +/- 0.06 ml/kg/h, half-life (t1/2beta) was 17.10 +/- 0.09 hours, and apparent volume of distribution was 2.49 +/- 0.43 L/kg. For oral administration, AUC0-infinity was 58.47 +/- 16.37 microg x h/ml, t1/2beta was 18.39 +/- 0.06 hours, maximum concentration (Cmax) was 2.12 +/- 00.51 microg/ml, time to Cmax was 2.20 +/- 2.17 hours, mean absorption time was 2.09 +/- 0.51 hours, and bioavailability was 42 +/- 0.42%. CONCLUSIONS AND CLINICAL RELEVANCE: Compared with adult horses given 5 mg of enrofloxacin/kg IV, foals have higher AUC0-infinity, longer t1/2beta, and lower clearance. Concentration of ciprofloxacin was negligible. Using a target Cmax to minimum inhibitory concentration ratio of 1:8 to 1:10, computer modeling suggests that 2.5 to 10 mg of enrofloxacin/kg administered every 24 hours would be effective in foals, depending on minimum inhibitory concentration of the pathogen. PMID- 10850850 TI - Efficacy of difloxacin in calves experimentally infected with Mannheimia haemolytica. AB - OBJECTIVE: To determine the efficacy of difloxacin, a novel fluoroquinolone antibiotic, in calves experimentally infected with Mannheimia haemolytica (formerly Pasteurella haemolytica). ANIMALS: Seventy-two 3-month-old Holstein calves. PROCEDURES: Calves were inoculated with M haemolytica intratracheally; after they developed clinical signs of pneumonic pasteurellosis, they were randomly assigned to 1 of 6 groups (n = 12/group). Calves in each group were treated with 10% difloxacin (2.5 or 5 mg/kg of body weight), 5% difloxacin (2.5 or 5 mg/kg), enrofloxacin (5 mg/kg), or saline (0.9% NaCl) solution (control group), once daily for 5 days, and clinical signs were scored daily. On day 15, calves were euthanatized, and the percentage of diseased lung tissue was calculated. Swab specimens of the lungs were submitted for bacterial culture. RESULTS: Mortality rate and percentage of diseased lung tissue were significantly higher and cure rate and average daily gain were significantly lower for control calves, compared with calves in the treatment groups; however, no significant differences were found among treatment groups. Mannheimia haemolytica was isolated from the lungs of 10 control calves and from at least 2 calves in each of the treatment groups. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that difloxacin and enrofloxacin were equally effective for treatment of calves with experimentally induced pneumonic pasteurellosis. However, treatment of infected calves with difloxacin or enrofloxacin may not eliminate the organism. PMID- 10850851 TI - Concentrations of substance P and prostaglandin E2 in synovial fluid of normal and abnormal joints of horses. AB - OBJECTIVE: To correlate substance P content of synovial fluid with prostaglandin E2 content, radiographic evidence of joint abnormality, and anatomic location of the joint for normal and osteoarthritic joints of horses. SAMPLE POPULATION: Synovial fluid from 46 normal joints in 21 horses and 16 osteoarthritic joints in 10 horses. PROCEDURE: Normal and osteoarthritic joints were identified by clinical and radiographic examination, by response to nerve blocks, during scintigraphy or surgery, or by clinicopathologic evaluation. Substance P and prostaglandin E2 contents of synovial fluid were determined by radioimmunoassay. Radio-graphs of joints were assigned a numeric score reflecting severity of lesions. Joints were assigned a numeric score reflecting anatomic location. RESULTS: Median concentrations of substance P and prostaglandin E2 were significantly increased in osteoarthritic joints, compared with normal joints. A significant correlation was found between concentrations of substance P and prostaglandin E2 in synovial fluid, but a correlation was not detected between substance P concentration in synovial fluid and anatomic location of the joint or between radiographic scores of osteoarthritic joints and concentrations of substance P or prostaglandin E2. CONCLUSIONS AND CLINICAL RELEVANCE: A correlation existed between concentrations of substance P and prostaglandin E2 in synovial fluid obtained from normal and osteoarthritic joints. However, content of substance P in synovial fluid cannot be predicted by the radiographic appearance of the joint or its anatomic location. Substance P and prostaglandin E2 may share an important and related role in the etiopathogenesis of osteoarthritis, lending credence to the importance of neurogenic inflammation in horses. PMID- 10850852 TI - Alveolar type II cell abnormalities and peroxide formation in lungs of rats given IL-1 intratracheally. AB - Acute lung injury (ALI) is characterized by increased lung levels of proinflammatory cytokines, inflammation, oxidative stress, edema, and impaired gas exchange. Notably, ALI patients also exhibit pulmonary surfactant abnormalities, including increased levels of phospholipids in their lung lavages. In the present study, to assess early alterations of the lung surfactant system in ALI, we induced inflammation and acute lung injury in rats by administering interleukin-1alpha (IL-1) intratracheally. Five h after IL-1 instillation, we examined lung tissue ultrastructure by electron microscopy using both routine staining methods and cerium chloride staining to localize hydrogen peroxide (H2O2) histologically. We also measured lung lavage phospholipid levels, lung tissue gamma-glutamyl transpeptidase (GGT) activities (a marker of oxidative stress), and arterial blood oxygen tensions. We observed that lungs of rats given IL-1 intratracheally had increased neutrophil accumulation, increased H2O2 production, and increased alveolar type II (ATII) pneumocyte ultrastructural abnormalities compared to rats given saline intratracheally. Intratracheal instillation of IL-1 also increased phospholipid levels in the bronchoalveolar lavage (BAL), possibly as a consequence of the abnormal discharge of lamellar bodies into the alveolar lumen. In addition, IL-1-insuffated rats had increased lung GGT levels and impaired blood oxygenation compared to saline-insufflated rats. Treatment with mepacrine decreased lung neutrophil accumulation, ultrastructural lung abnormalities, lung lavage phospholipid levels, lung tissue GGT levels, and blood oxygenation impairment in rats given IL-1 intratracheally, suggesting a possible relationship between these events. Our results indicate that IL-1-induced acute lung injury in rats is marked by neutrophil-dependent oxidative stress, ATII cell defects, abnormal discharge of lamellar body phospholipids, and impaired blood oxygenation. PMID- 10850853 TI - IRAK-2 and PI 3-kinase synergistically activate NF-kappaB and AP-1. AB - Antisense interleukin-1 (IL-1) receptor associated kinase-2 (IRAK-2) oligonucleotide (ODN) and antisense p110 PI 3-kinase ODN blocked IRAK-2 and p110 PI 3-kinase expression, respectively. As a result, antisense IRAK-2 ODN or antisense p110 PI 3-kinase ODN inhibited IL-1-induced NF-kappaB and AP-1 activation in HepG2 cells. The inhibition of NF-kappaB activation by antisense IRAK-2 ODN or antisense p110 PI 3-kinase ODN and the inhibition of AP-1 activation by antisense IRAK-2 ODN were incomplete, whereas AP-1 activation could be inhibited by antisense p110 PI 3-kinase ODN completely. These results indicate that IRAK-2 is necessary but insufficient to activate NF-kappaB and AP-1 completely and that although PI 3-kinase is not sufficient for NF-kappaB full activation, it is sufficient to activate AP-1 completely. The effects of IRAK-2 or PI 3-kinase on NF-kappaB and AP-1 activation were confirmed by the results that overexpression of IRAK-2 failed to fully activate NF-kappaB and AP-1 and that overexpression of p110 PI 3-kinase is insufficient for NF-kappaB full activation but sufficient for AP-1 activation. Cotransfection experiments showed that the combination of antisense IRAK-2 ODN and antisense p110 PI 3-kinase ODN resulted in additive inhibition of NF-kappaB as well as AP-1 activation. On the other hand, coexpression of IRAK-2 with p110 PI 3-kinase led to a synergistic activation of NF-kappaB and AP-1. These data suggest that IRAK-2 and PI 3-kinase cooperate to activate NF-kappaB and AP-1. PMID- 10850854 TI - Effects of several glucocorticosteroids and PDE4 inhibitors on increases in total lung eosinophil peroxidase (EPO) levels following either systemic or intratracheal administration in sephadex- or ovalbumin-induced inflammatory models. AB - Representative glucocorticosteroids (GCS) and phosphodiesterase IV (PDE4) inhibitors were compared in several models of pulmonary inflammation ranging in severity. Lung tissue eosinophil peroxidase (EPO) levels rather than bronchoalveolar lavage fluid (BALF) EPO or eosinophil percentages were used to indicate eosinophil recruitment after intratracheal instillation of sephadex beads in rats or nebulized ovalbumin in sensitized guinea pigs. A single oral or intratracheal administration of a GCS was effective against mild and robust sephadex-induced eosinophilia whereas the PDE4 inhibitors evaluated appeared more effective in the milder sephadex models. The GCS were also more effective against sephadex-induced than ovalbumin-induced eosinophilia. The effectiveness of the GCS and PDE4 inhibitors improved when the severity of the ovalbumin-induced eosinophilia was decreased. Multiple day dosing also improved activity. These studies indicated that activity was influenced greatly by administration protocols, the severity of the inflammatory response and possibly the method used for estimating eosinophil recruitment. PMID- 10850855 TI - Evaluation of chemokine- and phlogistin-mediated leukocyte chemotaxis using an in vivo sponge model. AB - We have directly compared the in vivo activity of a number of chemokines and phlogistins using a modified murine in vivo sponge model in which gelatin sponges are soaked with chemoattractant and implanted in the peritoneal cavity. Sponges soaked with murine JE/MCP-1 (monocyte chemoattractant protein-1) or zymosan promoted the chemotaxis of specific leukocyte populations in a time-dependent manner, as judged by multiparameter flow cytometry, with granulocytes predominating in zymosan-soaked sponges and granulocytes and macrophages present in JE/MCP-1-soaked sponges. Smaller numbers of B, T and dendritic cells were identified as well. Eotaxin selectively chemoattracted eosinophils in this model, while MIG induced significant T cell migration relative to other chemokines. Cell migration was inhibited by administration of methotrexate, piroxicam or dexamethasone, and JE/MCP-1-mediated trafficking was impaired by treatment with anti-JE antibody or with IL-10, suggesting a role for pro-inflammatory factors in amplifying the JE/MCP-1-induced response. This amplification phase involves the production of the chemokine KC, since anti-KC antibody significantly attenuated JE/MCP-1-induced chemotaxis. These results indicate that intraperitoneally implanted chemoattractant-soaked gelatin sponges are capable of inducing a pronounced inflammatory response characterized by the selective migration of leukocyte populations, and suggest that this model may be useful for delineating the activity of novel inhibitors of leukocyte chemotaxis. PMID- 10850856 TI - Modulation of hyperoxia-induced TNF-alpha expression in the newborn rat lung by thalidomide and dexamethasone. AB - The effect of high oxygen concentrations on lungs of neonatal rats was studied. In addition, some oxygen-exposed animals were treated with either dexamethasone or thalidomide. No gross histologic changes were noted in the lungs following exposure to 95% oxygen nor were there changes in the total number or the phenotypic distribution of BAL cells obtained from these lungs compared to lungs from air exposed (control) neonatal rats. The majority of the BAL cells were CD45+ leukocytes (macrophages). However, when BAL cells were exposed to LPS in vitro, TNF-alpha production was higher in cells from rats exposed to 95% oxygen compared to cells from rats exposed to ambient air. In addition, lung TNF-alpha and IL-6 mRNA levels were increased after exposure to 95% oxygen. In the lungs of animals treated with either dexamethasone or thalidomide, TNF-alpha mRNA levels were reduced, while only dexamethasone treatment also reduced IL-6 mRNA levels. PMID- 10850857 TI - Nepafenac, a unique nonsteroidal prodrug with potential utility in the treatment of trauma-induced ocular inflammation: I. Assessment of anti-inflammatory efficacy. AB - Nepafenac, the amide analog of 2-amino-3-benzoylbenzeneacetic acid (amfenac), was examined in preclinical models for its potential utility as a topical ocular anti inflammatory agent. Diclofenac was selected as the reference compound. In contrast to diclofenac (IC50 = 0.12 microM), nepafenac exhibited only weak COX-1 inhibitory activity (IC50 = 64.3 microM). However, amfenac was a potent inhibitor of both COX-1 (IC50 = 0.25 microM) and COX-2 activity (IC50 = 0.15 microM). Ex vivo, a single topical ocular dose of nepafenac (0.1%) inhibited prostaglandin synthesis in the iris/ciliary body (85-95%) and the retina/choroid (55%). These levels of inhibition were sustained for 6 h in the iris/ciliary body and 4 h in the retina/choroid. Diclofenac (0.1%) suppressed iris/ciliary body prostaglandin synthesis (100%) for only 20 min, with 75% recovery observed within 6 h following topical dosing. Diclofenac's inhibition of prostaglandin synthesis in the retina/choroid was minimal. Nepafenac's inhibitory efficacy and longer duration of action was confirmed in a trauma-induced rabbit model of acute ocular inflammation monitoring protein or PGE2 accumulation in aqueous humor. Results warrant further assessment of nepafenac's topical ocular efficacy in the treatment of postoperative ocular pain, inflammation, and posterior segment edema. PMID- 10850858 TI - Nepafenac, a unique nonsteroidal prodrug with potential utility in the treatment of trauma-induced ocular inflammation: II. In vitro bioactivation and permeation of external ocular barriers. AB - Nepafenac, the amide analog of the NSAID amfenac, was examined in vitro for its bioactivation by ocular tissue components and its ability to permeate external ocular barriers. Rabbit tissues catalyzed a concentration-dependent conversion of nepafenac to amfenac. The order of specific hydrolytic activity is retina/choroid >> iris/ciliary body. Corneal tissue showed only minimal activity. Similarly, in human ocular cadaver tissue the specific activity of iris/ciliary body was greater than cornea. Continued perfusion of the corneal epithelium demonstrated a nearly six-fold greater permeation coefficient for nepafenac (k(p) = 727 x 10(-6) min(-1)) than for diclofenac (k(p) = 127 x 10(-6) min(-1)). Superior permeation of conjunctival and scleral tissue by nepafenac (k(p) = 128 x 10(-6) min(-1)) compared to diclofenac (k(p) = 80 x 10(-6) min(-1)) was also evident. Short term perfusion (5 min) of the corneal surface with 0.1% nepafenac resulted in sustained flux of drug across the cornea for 6 h. Under identical conditions only 3.3 microM of diclofenac accumulated on the corneal endothelial side compared to 16.7 microM nepafenac. The enhanced permeability of nepafenac, combined with rapid bioactivation to amfenac by the iris/ciliary body and retina/choroid, make it a target specific NSAID for inhibiting prostaglandin formation in the anterior and posterior segments of the eye. PMID- 10850859 TI - Interleukin-1 and the immunogenetics of Alzheimer disease. AB - Established genetic causes of familial Alzheimer disease (AD) involve genes for beta-amyloid precursor protein (betaAPP), presenilin-1, and presenilin-2. For the more common sporadic forms of AD, increased risk has been associated with a number of genes; the most important of which is the epsilon4 allele of apolipoprotein E. Two recent studies, one clinical and one using postmortem material, now show increased risk for AD associated with certain polymorphisms in the genes encoding the alpha and beta isoforms of interleukin-1 (IL-1). IL-1 levels are elevated in Alzheimer brain, and overexpression of IL-1 is associated with beta-amyloid plaque progression. IL-1 interacts with the gene products of several other known or suspected genetic risk factors for AD, including betaAPP, apolipoprotein E, alpha1-antichymotrypsin, and alpha2-macroglobulin. IL-1 overexpression is also associated with environmental risk factors for AD, including normal aging and head trauma. These observations suggest an important pathogenic role for IL-1, and for IL-1-driven cascades, in the pathogenesis of AD. PMID- 10850860 TI - Acute demyelination, neuropathological diagnosis, and clinical evolution. AB - A retrospective analysis of 14 patients who presented with a progressively expanding mass lesion(s) shown at biopsy/autopsy to represent acute demyelination was carried out. The aims of this study were to determine the optimal neuropathological approach to diagnosis and to determine the clinical evolution of this condition. Subsequent investigations and clinical outcome studies confirmed MS in 10 cases. Two patients had received an incorrect neuropathologic diagnosis of astrocytoma resulting in cranial irradiation. Key histologic parameters in establishing a diagnosis of acute demyelination were a predominance of lipid filled macrophages, macrophage alignment along axons, and an absence of oligodendroglial inclusions. Axonal injury was present in all cases and a lymphocytic/plasma cell infiltrate was sparse in areas of demyelination. Neuroimaging revealed single lesions in 10 patients and multiple lesions in 4 patients. Two patients were lost to follow-up, 3 died within 18 months of diagnosis, 8 had a relapsing remitting clinical course, and 1 patient had a chronic progressive course. In conclusion, a dense lymphocytic and plasma cell infiltrate is unusual in acute human demyelination. Although axonal injury is a frequent histologic finding in acute demyelination, it does not preclude a favorable clinical outcome. PMID- 10850861 TI - Alpha-synuclein expression in central nervous system tumors showing neuronal or mixed neuronal/glial differentiation. AB - Alpha-synuclein (alpha-synuclein) is a member of a family of cytoplasmic proteins found predominantly and abundantly in the brain, and concentrated in pre-synaptic nerve terminals, near vesicles. We hypothesized that an antibody to alpha synuclein could be a useful marker of neuronal differentiation in central nervous system (CNS) tumors. Twenty tumors known to have neuronal or mixed neuronal/glial differentiation ( 11 gangliogliomas, 2 anaplastic gangliogliomas, 5 gangliocytomas, and 2 ganglioneuroblastomas), 5 central neurocytomas, and 1 dysembryoplastic neuroepithelial tumor (DNET) were immunostained with a mouse monoclonal antibody raised against human alpha-synuclein. Intense cytoplasmic staining, in some instances extending into the perikarya, was seen in 6 of 11 gangliogliomas, 2 of 2 anaplastic gangliogliomas, and 2 of 2 ganglioneuroblastomas. Alpha-synuclein-positive cells were usually large in size, resembled dysmorphic neurons, and were variably immunoreactive for anti neurofilament and/or anti-synaptophysin antibodies. In contrast, central neurocytomas, gangliocytomas, and the DNET were negative for cytoplasmic alpha synuclein expression. Our findings indicate that alpha-synuclein is expressed within the neuronal component of mixed tumors of the CNS displaying more than 1 histophenotype, and/or showing different degrees of anaplasia. Based on currently available data, we conclude that cytoplasmic alpha-synuclein expression is a marker of maturing neurons in these tumors. PMID- 10850862 TI - Amplification of the platelet-derived growth factor receptor-A (PDGFRA) gene occurs in oligodendrogliomas with grade IV anaplastic features. AB - Activation of the platelet-derived growth factor (PDGF) signaling system has been implicated in the development and malignant progression of diffuse gliomas. Overexpression of PDGF system components, particularly the alpha subtype receptor (PDGFRA), is common in glial tumors, and PDGFRA gene amplification has been reported in glioblastomas. In order to address the incidence of PDGFRA gene amplification in a broad set of diffuse gliomas, we used Southern and fluorescence in situ hybridization (FISH) analyses to examine 167 astrocytic gliomas (20 grade III and 147 grade IV), 41 anaplastic oligodendrogliomas, and 29 anaplastic oligoastrocytomas. PDGFRA gene amplification was identified in 4 anaplastic oligodendrogliomas and in a single case of anaplastic oligoastrocytoma, but in none of the malignant astrocytomas. Each of the 5 tumors with PDGFRA amplification displayed features generally associated with grade IV malignancy in astrocytic tumors. Consequently, our data indicate that this gene alteration is restricted to tumors having oligodendroglial differentiation and highly anaplastic features. PMID- 10850863 TI - Allelic losses in neurofibromatosis 2-associated meningiomas. AB - More than 50% of patients with neurofibromatosis 2 (NF2) develop meningiomas. Recently, a higher proliferative activity, more mitotic figures, and greater nuclear pleomorphism have been described for NF2-associated meningiomas compared with sporadic ones. To analyze whether such histological differences could reflect underlying genetic differences, we examined 30 meningiomas from 22 NF2 patients for allelic losses on those chromosome arms that are frequently affected by deletions in sporadic meningiomas. In addition, we assessed the proliferative activity of the tumors and studied NF2 germline mutations. Twenty-three meningiomas corresponded to WHO grade I (10 fibrous, 6 psammomatous, 4 transitional, 3 meningothelial) and 7 to WHO grade II. The average MIB-1 index was 1.60 +/- 0.85 (WHO grade I: 1.41 +/- 0.80, WHO grade II: 2.13 +/- 0.82). When compared with several published studies of sporadic meningiomas, the MIB-1 index in NF2-associated meningiomas was not higher. Loss of heterozygosity (LOH) flanking or within the NF2 locus at 22q12 was detected in 100% of the tumors. LOH on 1p was the second most frequent abnormality (40%), followed by losses on 10q (27%), 6q and 14q (24%), 18q (23%), and 9p (17%). LOH on 19q and 17p, which is not commonly seen in sporadic meningiomas, was also only rarely detected in NF2 associated meningiomas. NF2 gene mutations were detected in 8 of 15 patients analyzed and were located in exons 2, 5, 6, 7, and 8. We conclude that sporadic and NF2-associated meningiomas share a common spectrum and frequency of allelic deletions as well as, in contrast to previous observations, a similar proliferative activity. PMID- 10850864 TI - A qualitative and quantitative study of grumose degeneration in progressive supranuclear palsy. AB - Grumose degeneration (GD) of the dentate nucleus is a common feature in progressive supranuclear palsy (PSP), but its pathogenesis has not been well studied, and its clinical significance remains unknown. This report describes a quantitative study of GD in 9 cases of PSP using image analysis with single- and double-immunolabeling, as well as histochemical stains for myelin and axons. GD was associated with demyelination, axonal loss, glial tau pathology, and microgliosis in regions juxtaposed to the dentate nucleus (DN). Specifically, demyelination and microgliosis were prominent in the superior cerebellar peduncle (SCP), dentate hilus, and cerebellar hemispheric white matter. Tau pathology and microgliosis were less prominent in the DN itself. The degree of myelin loss correlated with the tau burden in the SCP. GAP-43, which is a phosphoprotein known to be involved in axonal growth and sprouting, was decreased in the DN of PSP, and the degree of GAP-43 loss correlated with severity of GD. These results suggest that GD may be related to progressive pathology in the dentatorubrothalamic tract as well as the cerebellar hemispheric white matter, and that GD may be a consequence of concurrent degeneration in both output from and input to the DN. The results further suggest a possible role for oligodendroglial and myelin pathology in the pathogenesis of PSP. PMID- 10850865 TI - Microvasculitis in non-diabetic lumbosacral radiculoplexus neuropathy (LSRPN): similarity to the diabetic variety (DLSRPN). AB - Diabetic lumbosacral radiculoplexus neuropathy (DLSRPN) has been shown to be due to ischemic injury from microvasculitis. The present study tests whether ischemic injury and microvasculitis are the pathologic cause of non-diabetic lumbosacral radiculoplexus neuropathy (LSRPN), and whether the pathologic alterations are different between LSRPN and DLSRPN. We studied distal cutaneous nerve biopsies of 47 patients with LSRPN and compared findings with those of 14 age-matched healthy controls and 33 DLSRPN patients. In both disease conditions, we found evidence of ischemic injury (multifocal fiber degeneration and loss, perineurial degeneration and scarring, characteristic fiber alterations, neovascularization, and injury neuroma) that we attribute to microvasculitis (mural and perivascular mononuclear inflammation of microvessels, inflammatory separation, fragmentation and destruction of mural smooth muscle, and previous microscopic bleeding [hemosiderin]). Teased nerve fibers in LSRPN showed significantly increased frequencies of axonal degeneration, segmental demyelination, and empty nerve strands. The segmental demyelination appeared to be clustered on fibers with axonal dystrophy. The nerves with abnormal frequencies of demyelination were significantly associated with nerves showing multifocal fiber loss. We reached the following conclusions: 1) LSRPN is a serious condition with much morbidity that mirrors DLSRPN. 2) Ischemic injury from microvasculitis appears to be the cause of LSRPN. 3) Axonal degeneration and segmental demyelination appear to be linked and due to ischemia. 4) The pathologic alterations in LSRPN and DLSRPN are indistinguishable, raising the question whether these 2 conditions have a common underlying mechanism, and whether diabetes mellitus contributes to the pathology or is a risk factor in DLSRPN. 5) Both LSRPN and DLSRPN are potentially treatable conditions. PMID- 10850866 TI - Loss of heterozygosity on chromosome 19 in secondary glioblastomas. AB - Glioblastomas develop rapidly de novo (primary glioblastomas) or slowly through progression from low-grade or anaplastic astrocytoma (secondary glioblastomas). Recent studies have shown that these glioblastoma subtypes develop through different genetic pathways. Primary glioblastomas are characterized by EGFR amplification/overexpression, PTEN mutation, homozygous p16 deletion, and loss of heterozygosity (LOH) on entire chromosome 10, whereas secondary glioblastomas frequently contain p53 mutations and show LOH on chromosome 10q. In this study, we analyzed LOH on chromosomes 19q, 1p, and 13q, using polymorphic microsatellite markers in 17 primary glioblastomas and in 13 secondary glioblastomas that progressed from low-grade astrocytomas. LOH on chromosome 19q was frequently found in secondary glioblastomas (7 of 13, 54%) but rarely detected in primary glioblastomas (1 of 17, 6%, p = 0.0094). The common deletion was 19q13.3 (between D19S219 and D19S902). These results suggest that tumor suppressor gene(s) located on chromosome 19q are frequently involved in the progression from low-grade astrocytoma to secondary glioblastoma, but do not play a major role in the evolution of primary glioblastomas. LOH on chromosome 1p was detected in 12% of primary and 15% of secondary glioblastomas. LOH on 13q was detected in 12% of primary and in 38% of secondary glioblastomas and typically included the RB locus. Except for 1 case, LOH 13q and 19q were mutually exclusive. PMID- 10850867 TI - Comprehensive allelotype and genetic anaysis of 466 human nervous system tumors. AB - Brain tumors pose a particular challenge to molecular oncology. Many different tumor entities develop in the nervous system and some of them appear to follow distinct pathogenic routes. Molecular genetic alterations have increasingly been reported in nervous system neoplasms. However, a considerable number of affected genes remain to be identified. We present here a comprehensive allelotype analysis of 466 nervous system tumors based on loss of heterozygosity (LOH) studies with 129 microsatellite markers that span the genome. Specific alterations of the EGFR, CDK4, CDKN2A, TP53, DMBT1, NF2, and PTEN genes were analyzed in addition. Our data point to several novel genetic loci associated with brain tumor development, demonstrate relationships between molecular changes and histopathological features, and further expand the concept of molecular tumor variants in neuro-oncology. This catalogue may provide a valuable framework for future studies to delineate molecular pathways in many types of human central nervous system tumors. PMID- 10850868 TI - Magnetic resonance arthrography of the canine stifle joint: technique and applications in eleven military dogs. AB - The stifle joints of eleven military working dogs were evaluated using conventional magnetic resonance (MR) imaging and MR arthrography. A protocol optimizing MR imaging of the canine stifle joint is discussed, as well as potential uses for administration of intra-articular gadolinium. The technique for performing MR arthrography is described, and post-contrast image findings are reviewed. MR arthrography was performed by using an intra-articular injection of diluted gadolinium. Consistently good quality images were obtained, and no complications were clinically detected following MR arthrography. Cranial cruciate ligament abnormalities were seen in six dogs, meniscal abnormalities were visualized in nine menisci, and synovitis and medial ligament strain were seen in eight dogs. Surgical and post-mortem confirmation of these findings is discussed in seven dogs. Although MR arthrography adds an invasive procedure to conventional MR imaging, it can provide useful information on pathologic changes in the canine stifle joint. PMID- 10850869 TI - Computed tomographic characteristics of multilobular tumor of bone involving the cranium in 7 dogs and zygomatic arch in 2 dogs. AB - Computed tomography (CT) images of nine dogs with a multilobular tumor of bone of the head were reviewed. The CT characteristics of the neoplasms involving the calvarium (n = 7) were rounded, well defined with a fine granular, nonhomogeneous bone opacity usually in the occipital region. Cranial vault invasion (5 of 7) was commonly found with a significant portion of the mass within the vault. The neoplasms involving the zygomatic arch (n = 2) were also generally rounded and well defined but with a more coarse granular appearance. The common CT findings were best seen when the images were viewed in a bone window. PMID- 10850870 TI - Imaging diagnosis--occipitoatlantoaxial malformation in a miniature horse foal. PMID- 10850871 TI - Radiographic anatomy of the cribriform plate (Lamina cribrosa). AB - The radiographic appearance of the cribriform plate was investigated in 16 canine cadaver heads. The cribriform plate appeared as a "V"-shaped multilinear bone opaque stripe in the caudal nasal region in projections perpendicular to the hard palate in 6 dogs with a skull index between 50.00 and 54.00. In 9 dogs with a skull index between 55.40 and 74.40, the cribriform plate had a more "C"-shaped and sharp appearance. In vertically oblique projections with an obliquity greater than 20 degrees, the cribriform plate lost its sharp outline and finally (40 degrees) disappeared. In lateral projections the cribriform plate appeared as a "C"-shaped interrupted bone-opaque stripe in all 16 dogs. In more brachycephalic dogs frontal bone structures superimposed on the cribriform plate on ventrodorsal and dorsoventral views and accentuated the radiographic appearance of the plate. Vertically oblique views separated both structures to produce two lines. PMID- 10850872 TI - Radiographic detection of defects of the nasal boundaries. AB - The sensitivity of conventional radiography for lesions of the cribriform plate, naso-orbital wall, lateral nasal wall and hard palate was investigated in 13 canine cadaver heads by creating measured defects in these structures. The location of the perforations were marked with a thin copper wire and the radiographic appearance of the defects was evaluated retrospectively by a single reviewer. Despite demarcation cribriform plate destruction of 2 mm could not be detected. Defects of 3 mm were detected in only 2 heads, 4 mm defects in 1 further head, an oblong 4 x 10 mm defect in 7 heads and in the remaining 3 heads only a 10 x 10 mm defect became visible as such. The naso-orbital wall had to be destroyed in its whole vertical length for detection of a defect in ventrodorsal or dorsoventral views. Therefore conventional radiography is of low diagnostic value for these lesions. Defects of 2 mm in the lateral nasal wall and the hard palate could be detected confidently in all heads indicating high sensitivity of conventional radiography. Soft tissue opacification did not alter the detectability of any nasal border structure lesion. PMID- 10850873 TI - Canalographic evaluation of the external ear canal in dogs. AB - Canalography was performed to measure the diameter of the horizontal ear canal and to assess the clinical relevance of the measurements in identifying abnormalities of the ear canal. The diameter of the proximal and distal end of the annular cartilage in the horizontal ear canal was measured using canalography in 222 ears. The tympanic membrane could not be visualized with otoscopic examination in 70 ears even after ear cleansing. These canals were classified as being stenotic. The diameter of the proximal annular cartilage in the stenotic canals ranged from 0.8 to 3.1 mm (mean = 2.6 +/- 0.8 mm). The proximal annular cartilage was consistently smaller in diameter than the distal annular cartilage. The ratio between the diameter of the proximal and distal annular cartilage varied between dogs. In stenotic ear canals (70 ears) the ratio was less than 0.65. Total ear canal ablations and histopathological analysis were performed in 70 ear canals classified as stenotic after canalography. Hyperplasia of the epidermal layer was found in 56 stenotic canals. Stenosis of the canal due to otitis externa was found in the other 14 canals. Three canals with severe otitis externa were also diagnosed with canalography as having a ruptured tympanic membrane. The results suggest that canalography can be used to measure the diameter of the horizontal ear canal and to assess whether or not the tympanic membrane is ruptured prior to conservative or surgical therapy. PMID- 10850874 TI - Contrast medium-related artifacts observed during in vitro radiographic characterization of urocystolith mineral composition. AB - Nine pure mineral types of canine uroliths (bladder or urethral origin only) were exposed to sequential increasing concentrations of iodinated, radiographic contrast medium in petri dishes. The uroliths studied were those composed of 100% magnesium ammonium phosphate, calcium oxalate monohydrate, calcium oxalate dihydrate, calcium phosphate appatite, and calcium hydrogen phosphate dihydrate (Brushite), ammonium acid urate, sodium acid urate, cystine, and silica. Two phenomena were observed. First, there was a tendency for selected urocystoliths to undergo radiopacity augmentation beyond that expected for just contrast medium superimposition. This was termed, contrast medium adhesion, which persisted despite repeated washing of the urocystoliths. Second, there was a tendency for bubbles to form on or near selected urocystolith chemical types. These observations prompted careful scrutiny for their occurrence in subsequent clinical simulation of radiographic procedures using these same urocystoliths in a urinary bladder phantom. Imaging techniques simulated were survey radiography, pneumocystography, double contrast cystography (two iodine concentrations). The contrast medium adhesion occurrence found in the petri dish studies was compared to urocystolith mineral type. Similar comparisons were made for contrast medium adhesion occurrence in the bladder phantom. The detection of contrast medium adhesion in the bladder phantom differed from that observed in the petri dish experiments. While contrast adhesion occurred across a fairly broad range of the urocystolith mineral types in the petri dish studies, it was observed primarily for sodium acid urate and cystine urocystoliths in the bladder phantom. Prompted by the observation of bubbles in association with a limited number of urocystolith types in the petri dish studies, bubble occurrence in the bladder phantom was compared to the urocystolith type. Bubble formation on or near the urocystoliths, although uncommonly observed, was seen only with either cystine or silica urocystoliths. The potential clinical utility and clinical caveat aspects of these phenomena are discussed. PMID- 10850875 TI - Loss of urocystolith architectural clarity during in vivo radiographic simulation versus in vitro visualization. AB - Urocystoliths of 9 mineral types from 434 canine patients submitted to the University of Minnesota Urolith Bank were imaged in a urinary bladder phantom. Imaging techniques simulated were survey radiography and double contrast cystography. Morphologic characteristics visually observed in vitro or by interpretation of high-resolution specimen radiographs were compared to those seen using the simulated in vivo imaging techniques. Shape characteristics that were accurately detected > or = 25% of the time on simulated survey or double contrast radiography were faceted, irregular, jackstone, ovoid, and round. Surface characteristics that were accurately detected > or = 25% of the time on simulated survey or double contrast radiography were rough, smooth, and smooth with blunt tips. Internal architecture characteristics that were accurately detected > or = 25% of the time on simulated survey or double contrast radiography were lucent center, random-nonuniform, and uniform. Shapes such as bosselated, faceted-ovoid, and rosette; surfaces such as botryoidal, and knife edged; and internal architecture characteristics such as dense center, dense shell, laminated, and fissures were of almost no value either due to poor detectability or poor accuracy of recognition. Based on optimized simulated survey and double contrast radiographic procedures, it appears that a number of shape, surface, and internal architecture characteristics may be of limited or no value in discriminating among urocystolith mineral types under clinical circumstances. Shapes and surfaces were more accurately characterized by the simulated double contrast technique, but for internal architecture, the simulated survey radiographic technique seemed slightly superior overall. PMID- 10850876 TI - Radiographic diagnosis--ethmoid hematoma. PMID- 10850877 TI - Bone infection in the bovine appendicular skeleton: a clinical, radiographic, and experimental study. AB - Four hundred and forty-five bovines with bone infection of the appendicular skeleton were selected for clinical and radiographic assessment. A distinction was made between hematogenous and post-traumatic origin (wound/fracture). Bone infection was classified into four types according to the site of infection: type 1 is metaphyseal and/or epiphyseal osteomyelitis close to the growth plate; type 2 is primary subchondral osteomyelitis mostly accompanied by septic arthritis; type 3 is infectious osteoarthritis with subchondral osteomyelitis implicating that infection in the subchondral bone originates from the infection. Type 4 summarizes bone infections which can not be categorized in the previous groups. Hematogenous osteomyelitis is 3.2 times more frequent than post-traumatic osteomyelitis. Within the different groups, the relation of hematogenous to post traumatic infection changed significantly. In type 1 infection the ratio was 5/1, in type 2 the ratio was 8/1 and in type 3 it was 3/1. Epiphyseal or metaphyseal osteomyelitis exhibited early radiographic bone changes, whereas with infection eminating from the joint, the bone lesions were detected later, because the bone was infected as a consequence of a primary septic arthritis. In smaller bones, severe and complete bone destruction was often present. Hematogenous bone infection never involved the entire diaphysis. Actinomyces pyogenes was discovered to be the main etiologic agent, whether or not combined with anaerobes. Bone fragments from the metaphysis of young calves were subjected to the effect of pure cultures of different bacteria. Radiographic changes to the structure of the bone were not identified within 2 weeks. Rapid radiographic changes in osteomyelitis cannot be explained by the direct effect of the bacteria on bone tissue in vivo. General infections of the lungs, disorders of the intestines and other internal organs were rarely responsible for osteomyelitis or septic arthritis. Osteomyelitis and infectious osteoarthritis is probably often induced by external and internal traumatic events to joints and bones in cattle. PMID- 10850878 TI - Ultrasonographic appearance and clinical findings in 14 dogs with gallbladder mucocele. AB - Fourteen dogs with enlarged gallbladders and immobile stellate or finely striated bile patterns on ultrasound are described. Smaller breeds and older dogs were overrepresented, with 4/14 Cocker Spaniels. Most dogs presented for nonspecific clinical signs such as vomiting, anorexia and lethargy. Abdominal pain, icterus and hyperthermia were the most common findings on physical examination. All dogs except one had serum elevation of total bilirubin and/or alkaline phosphatase, alanine aminotransferase and gamma glutamyl transferase. All dogs were diagnosed with a gallbladder mucocele upon histologic and/or macroscopic evaluation. Ultrasonographically, mucoceles are characterized by the appearance of the stellate or finely striated bile patterns and differ from biliary sludge by the absence of gravity dependent bile movement. On ultrasound, gallbladder wall thickness and wall appearance were variable and nonspecific. The cystic or common bile duct were normal sized in 5 dogs although all 5 had evidence of biliary obstruction at surgery or necropsy. Loss of gallbladder wall integrity and/or gallbladder rupture were present in 50% of the dogs, all located in the fundus. Gallbladder wall discontinuity on ultrasound indicated rupture whereas neither bile patterns predicted the likelihood of gallbladder rupture. Pericholecystic hyperechoic fat or fluid were suggestive of but not diagnostic for a gallbladder rupture. Cholecystectomy appears to be an appropriate treatment for mucoceles, if not to treat a gallbladder rupture, at least in most dogs to prevent it since gallbladder wall necrosis was identified by histology in 9 of 10 dogs. Mucosal hyperplasia was present in all gallbladders examined histologically. Positive aerobic bacterial culture was obtained from bile in 6 of 9 dogs. Cholecystitis was diagnosed histologically in 5 dogs and 4 dogs had signs of gallbladder infection solely upon bacterial bile culture. Gallbladder infection was not present with all the mucoceles suggesting that biliary stasis and mucosal hyperplasia may be the primary factors involved in mucocele formation. Based on the results of our study, we suggest two alternate courses of action in the presence of a distended gallbladder with an immobile ultrasonographic stellate or finely striated bile pattern: a cholecystectomy when clinical or biochemical signs of hepatobiliary disease are present or a medical treatment (antibiotics and choleretics) and patient monitoring by follow-up ultrasound examinations when the patient does not have clinical or biochemical abnormalities. An aerobic bile culture should be obtained in all patients, by ultrasound-guided fine needle aspirate or at surgery. PMID- 10850879 TI - Ultrasonographic features of canine gastrointestinal pythiosis. AB - Pythiosis is a chronic pyogranulomatous infection of the gastrointestinal tract or skin caused by the water borne pathogen Pythium insidiosum. The ultrasonographic features of nine dogs with gastrointestinal pythiosis are reported. The stomach, duodenum, jejunum or colon were affected. All dogs had thickening of the gastrointestinal wall and areas with obliteration of the normal layered appearance. In one dog an eccentric mass was found arising from the serosal surface of the wall of the colon with mild diffuse wall thickening. Regional lymph node enlargement was seen in seven of the nine dogs. One dog had invasion of the pancreas and signs compatible with extrahepatic biliary obstruction. When compared to previous reports of gastrointestinal neoplasia, the features of wall thickening, loss of layering and regional lymphadenopathy are not considered specific for gastrointestinal pythiosis. Histological examination of tissue specimens is required for diagnosis. PMID- 10850880 TI - Radiographic and ultrasonographic features of retained surgical sponge in eight dogs. AB - The radiographic and ultrasonographic signs in eight dogs with a surgical or pathologic diagnosis of retained surgical sponge were reviewed. The most frequent previous surgery was ovariohysterectomy, either as an elective procedure or to treat pyometra. The median elapsed time between surgery and diagnosis of retained surgical sponge was 9.5 months (range 4 days to 38 months). Five dogs had a draining sinus; four had a palpable abdominal mass. Radiologic signs included localized, speckled or whirl-like gas lucency, abdominal mass, and non-focal soft tissue swelling. Survey radiography and sinography were considered diagnostic for retained surgical sponge in 4/7 (57%) and 3/5 (60%) dogs, respectively. The combined use of survey radiography and sinography enabled detection of 6/7 (86%) sponges. In each dog that had ultrasonography, a hypoechoic mass was found that had an irregular hyperechoic centre. The possibility of retained surgical sponge should be considered in animals with a history of previous surgery and a sinus or abdominal mass. PMID- 10850881 TI - Doppler ultrasonographic diagnosis and anatomy of congenital intrahepatic arterioportal fistula in a puppy. AB - A dilated, tortuous blood vessel was identified sonographically in the right medial liver lobe in a puppy with severe ascites. This vessel was thought to represent the dilated right medial portal vein branch. Using pulsed wave Doppler ultrasonography, retrograde, abnormally pulsatile flow was detected in both the dilated right medial portal vein branch and the main portal vein. The right medial liver lobe was surgically resected then fixed in formalin. Silicon rubber was injected and outlined the connection between the portal vein and hepatic artery. PMID- 10850882 TI - Radiographic and ultrasonographic evaluation of the normal feline postpartum uterus. AB - The radiographic and ultrasonographic appearance of the normal involution process of the feline postpartum uterus has not been previously described. Six queens were examined to determine the normal radiographic and ultrasonographic appearance of the involuting postpartum uterus. Radiographic and ultrasonographic examinations were performed daily from days one through ten, then on days 12, 14, 18, 24, and 28 postpartum. Radiographically the mean total uterine thickness was 16.5 mm at day one postpartum. By day 14 the mean total uterine thickness was 10.5 mm and by day 24 postpartum the uterus was not radiographically visible. Ultrasonographically at day one postpartum the mean total uterine thickness was 16.6 mm and the mean uterine wall thickness was 2.7 mm. At day 14 postpartum the mean total uterine thickness was 6.2 mm and the mean wall thickness was 2.1 mm. At day 28 postpartum the uterus could still be identified ultrasonographically however individual wall layers were not discernable. PMID- 10850883 TI - Two little ducks went swimming one day... PMID- 10850884 TI - Unexpected occurrence of an intratarsal duck. PMID- 10850885 TI - Photokilling mechanisms induced by zinc(II)-phthalocyanine on cultured tumor cells. AB - The photosensitizing effects of liposomal zinc(II)-phthalocyanine (ZnPc) on HeLa cells, with emphasis on morphological changes and mechanisms for cell death, have been studied. No dark toxicity for ZnPc alone was found. Incubation for 1 h with ZnPc followed by red light irradiation induced a variable decrease in the surviving of cells, which was related to both drug concentration and irradiation time. A lethal photodynamic effect (100% of the cells are killed: LD100) was induced by 5 x 10-6 M ZnPc and 5-min irradiation, whereas a sublethal effect (60% of the cells are killed: LD60) was detected with 10 7 M ZnPc and 3 min of red light. Toluidine blue and Hoechst 33258 staining showed characteristic alterations of cell morphology. Numerous bubbles on the plasma membrane were found immediately after an LD100 treatment, and a necrotic morphology appeared 24 h later. On the contrary, severe cell shrinkage with nuclear fragmentation. characteristic of apoptosis. was observed 8 and 24 h after LD60 treatments. In this case, propidium iodide-acridine orange labeling and the TUNEL assay confirmed the occurrence of apoptosis. The highest amount of apoptotic cells appeared 24 h after LD60 treatments, particularly in detached cells, as revealed by cell counting and DNA electrophoresis. Both apoptotic and necrotic mechanisms for cell death occur in HeLa cells in dependence on the experimental protocol of ZnPc photodynamic treatments. PMID- 10850886 TI - Cell death in paclitaxel-dependent chinese hamster ovary cells is initiated by the loss of telomeric DNA repeats. AB - We have reported earlier that cell death in a metastatic murine melanoma cell line induced by paclitaxel and its water-soluble conjugates is mediated through the extensive erosion of telomeric repeats. The purpose of this study was to investigate if loss of telomeric repeats was also involved in cell death of Tax 18 and Tax-2-4, two paclitaxel-requiring mutant Chinese hamster ovary (CHO) cell lines. Tax-18 and Tax-2-4 cells were grown in paclitaxel-free culture medium for 24, 48, 72, and 96 h at 37 degrees C and then harvested for cytological preparations. Control cultures of both cell lines were grown in paclitaxel supplemented medium and harvested simultaneously. We found that: 1) the frequency of telomeric associations in metaphase preparations was increased with the duration of paclitaxel-depleted culture; 2) Tax-18 cells showed a higher incidence (33.0%) of endoreduplicated metaphases at 24 h of paclitaxel-depleted culture than did Tax-2-4 cells, in which endoreduplicated metaphases were rare; 3) the frequency of polyploid cells was increased after 48, 72, and 96 h of paclitaxel-depleted culture for Tax-18 relative to that for Tax-2-4 cells; 4) both cell lines showed reductions in telomeric signals at chromosomal termini, but not in the interphase nuclei; and 5) both cell lines had shorter terminal telomeric restriction fragments after culture in paclitaxel-depleted medium. These results support our earlier observations and indicate that reduction of telomeric repeats is involved in G2/M cell arrest (endoreduplication) followed by severe DNA fragmentation, and then cell death of two CHO mutant cell lines that require paclitaxel for cell division. PMID- 10850887 TI - Involvement of TNF-alpha in enhancement of invasion and metastasis of colon 26-L5 carcinoma cells in mice by social isolation stress. AB - Psychosocial stress has been implicated in tumor metastasis. We have previously reported that social isolation stress exacerbated liver metastasis of colon 26-L5 by partially suppressing the cellular immunity in male Balb/c mice. To further understand the mechanism underlying the influence of isolation stress on liver metastasis, we investigated the effect of social isolation stress on tumor invasion, which is considered to be a pivotal step of tumor metastasis. The invasion and migration of tumor cells obtained from tumor nodules in the isolated mice were more markedly enhanced than that in the group-housed mice. The mRNA expression of proteolytic proteases, including matrix metalloproteinase (MMP)-2, MMP-9, membrane type 1 (MTI)-MMP, and urokinase-type plasminogen activator (u PA), were increased in the tumor and liver tissues of the isolated mice compared with the control mice. On the other hand, production of plasma TNF-alpha and expression of hepatic TNF-alpha mRNA were elevated in the isolated mice with or without tumor burden. Increased TNF-alpha level was particularly discernible in the liver of tumor-bearing mice. Elevated positive staining for TNF-alpha was immunohistochemically observed within and around tumor mass in the liver from isolated tumor-bearing mice, compared with group-housed mice. In addition, the invasiveness of tumor cells and the expression of proteolytic enzymes, including MMP-9 and u-PA in tumor cells, were enhanced by the treatment of TNF-alpha in vitro. Thus, the data suggested that isolation stress-augmented TNF-alpha may be involved in the enhancement of tumor invasion and metastasis in part by upregulating the proteolytic enzymes such as MMPs and u-PA in tumor and liver tissues. PMID- 10850888 TI - Cytotoxic effects toward human hematopoietic progenitor cells and tumor cell lines of paclitaxel, docetaxel, and newly developed analogues IDN5109, IDN5111, and IDN5127. AB - The growth inhibitory effect of paclitaxel, docetaxel, and newly developed taxanes IDN5109, IDN5111, and IDN5127 was assessed on peripheral blood (PB) CD34+ maintained in liquid culture and on three human cancer cell lines (MDA-MB231, MCF 7 ADRr, CEM VBLr). Concomitantly, DNA analysis was also performed. For unfractionated peripheral blood progenitor cells (PBPC) toxicity was also assessed by clonogenic assay. The cytotoxic effects induced by taxanes toward PBPC as measured by clonogenic assay were correlated with those found for multidrug resistance (MDR)-positive cell lines (IDN5109 > IDN5111 > IDN5127 > docetaxel > paclitaxel). We established a therapeutic index (TI) between the antitumor activity in MDR-positive cells and the toxicity toward PBPC. Paclitaxel and IDN5109, as determined by TI, showed the best value in MDR-negative and MDR positive cells, respectively. The ranking of the cytotoxic effects observed in PB CD34+ was not correlated with that obtained in clonogenic assay and in cancer cells (IDN5127 > IDN5109 > docetaxel > IDN5111). Remarkably, in DNA analysis docetaxel induced the maximal cell cycle blocking activity. Newly developed taxanes IDN5109 and IDN5111 are endowed of a profile of anticancer activity in MDR-bearing cells and toxicity toward hematopoietic progenitors better than that of docetaxel. However, mechanism(s) underlying toxicity toward hematopoietic progenitors could be, at least in part, different from that of docetaxel and likely dependent on the interaction with P-glycoprotein function in PB CD34+ cells. PMID- 10850889 TI - Antimetastatic and anti-invasive ability of phospho-ascorbyl palmitate through intracellular ascorbate enrichment and the resultant antioxidant action. AB - A lipophilic and auto-oxidation-resistant derivative of ascorbic acid (Asc), Asc 2-O-phosphate-6-O-palmitate (Asc2P6Plm), was shown to exert an invasion inhibitory action as promptly as 30-40 min for 50% inhibition and 60-90 min for 80% inhibition after entering fibrosarcoma HT-1080 cells. Invasive inhibition of 95-97% was accomplished by Asc2P6Plm of doses exhibiting no cytotoxicity under the same conditions. Asc2P6Plm was dephosphorylated and esterolyzed to Asc, which enriched the intracellular Asc dose dependently in invasion-suppressed cells, contrasting with no detectable Asc in invasive cells fed without Asc2P6Plm. Intracellular dehydroAsc (DehAsc), unexpectedly, amounted to 1.02-1.65-fold as much as intracellular Asc, suggesting that invasive cells underwent oxidative stress, the repression of which resulted in both inhibition of tumor invasion and oxidative conversion of Asc to DehAsc. Correspondingly, intracellular oxidants fluorometrically detected with a redox indicator CDCFH were more abundant in invasive cells than in invasion-suppressed cells fed with Asc2P6Plm. Invasion inhibitory effects of Asc2P6Plm necessitated the extensive inhibition of the major gelatinases MMP-9 and MMP-2, as shown by zymography and Western blots, but did not necessitate direct expression of the metastasis suppressor gene nm23, taking as long as 6-18 h in contrast to a prompt action of Asc2P6Plm. Antimetastatic effects on melanoma B16BL6 cells were given dose dependently by intravenous administration or pretreatment with Asc2P6Plm. Thus, Asc2P6Plm is anticipated as an antimetastatic agent via the potent antioxidant activity. PMID- 10850890 TI - Laudatio: in celebration of the 80th birthday of Maurice R. Hilleman, Ph.D. PMID- 10850891 TI - Vaccines in historic evolution and perspective: a narrative of vaccine discoveries. AB - The sciences of vaccinology and immunology were created only two centuries ago by Jenner's scientific studies of prevention of smallpox through inoculation with cowpox virus. This rudimentary beginning was expanded greatly by the giants of late 19th- and early 20th-century biomedical sciences. The period from 1930 to 1950 was a transitional era, with the introduction of chick embryos and minced tissues for propagating viruses and rickettsiae in vitro for vaccines. Modern vaccinology began about 1950 as a continuum following notable advances made during the 1940s and World War II. Its pursuit has been based largely on breakthroughs in cell culture, bacterial polysaccharide chemistry, molecular biology, and immunology which have yielded many live and killed viral and bacterial vaccines plus the recombinant-expressed hepatitis B vaccine. The present paper was presented as a lecture given at a Meeting of the Institute of Human Virology entitled A Symposium on HIV-AIDS and Cancer Biology, Baltimore, Maryland, on August 30, 1999 and recounts, by invitation, more than 55 years of vaccine research from the venue of personal experience and attainment by the author. The paper is intentionally brief and truncated with focus only on highlights and limited referencing. Detailed recounting and referencing are given elsewhere in text references 1 and 2. This narration will have achieved its purpose if it provides a background of understanding and guidelines that will assist others who seek to engage in creation of new vaccines. PMID- 10850892 TI - Will a new vaccine be used? AB - Despite the fact that vaccines are successful at preventing disease as well as cost-effective, many factors prevent their uptake. These factors are different for different regions. In the United States and Europe, several factors have been identified. The first of these factors is complacency. Both patients and parents lack the knowledge of disease. In addition, physicians are often apathetic about vaccines. There is a lack of training in public health and preventive medicine in that the mind-set of physicians is not directed toward prevention but rather toward treatment. Younger physicians are often less aware of disease controlled by vaccines than their older colleagues are because they have not seen them. One of the paradoxes of vaccines is that success leads to decreased use. Despite the Vaccine for Children's Program, which has provided ample funding for the purchase of vaccines, there are limits on funding for people, facilities, and systematized approaches to maintaining high levels of immunizations. Finally, and perhaps most important, there is fear, real and unfounded. Additionally, in many developing countries there is a lack of focus on preventive medicine, including immunization, as well as limited funding available. Reasonable steps can be taken to overcome these obstacles. PMID- 10850893 TI - How does HIV induce AIDS? The virus protein hypothesis. AB - The genome, the structural and regulatory proteins of HIV-1, as well as the clinical course of the disease it induces are well studied. Despite this knowledge, it is still unknown how the virus induces AIDS. The disease is characterized by opportunistic infections based on a generalized and nonspecific immunosuppression. I present evidence indicating that the immunosuppressive domain of gp41 of HIV may be causually involved in the induction and progression of AIDS. PMID- 10850894 TI - Is AIDS vaccine research at a turning point? AB - Although initial anti-ENV vaccines have failed, better results may be obtained with "non-ENV" vaccines capable of inducing cell-mediated responses and perhaps mucosal reactions. A turning point in AIDS vaccine research has been reached in that there are three precise questions to answer: 1. Are the protections obtained in monkeys significant and reproducible and could they be made more frequent? 2. Can more frequent, polyepitopic, and long-lasting cell-mediated responses in human subjects be obtained? 3. Is it possible to induce significant mucosal responses? The decision as to whether phase III trials must be launched will depend on answers to these questions. It may be possible to organize international AIDS vaccine research so that these answers could be obtained in 3 to 4 years, but this would require a frank acceleration of the present endeavors. PMID- 10850895 TI - 1999: a time to re-evaluate AIDS vaccine strategies. AB - The field of AIDS vaccine development is in flux. Important new findings were reported in 1999 that led to a rethinking of AIDS vaccine strategies. We have been given the challenging task of providing an overview. Rather than attempting to provide a comprehensive summary, we will restrict our discussion to a few major topics, and we ask for understanding if we can only highlight. PMID- 10850896 TI - Perspectives and needs for health in the 21st century: 20th-century paradigms in 21st-century science. AB - Countries in both the developed and developing world are expected to experience an unprecedented growth in the number of their elderly and decline in the number of their youth. This extraordinarily rapid aging of the world's population is a phenomenon without precedence in human history. Additionally, there is an increasing gap between the rich and poor both among and within nations. Both trends will not only constitute the primary health issues of the first decades of the 21st century but will also profoundly alter the world's economic, political, and social landscapes. PMID- 10850897 TI - Public support for medical research in the 21st century. AB - Key public policies that have contributed to the rise of modern medical research in the 20th Century are reviewed, focusing especially on the United States and the post-World War II period. Drawing on this history, the question is posed: "Are these policies sufficient to insure vigorous medical research in the 21st Century?" Although radical policy changes are not needed, several proposals for policy and medical research portfolio redirection are offered, including a rebalancing of public supported research in all fields of science that contribute to medical advances. Medical research must also invest in a national and international information infrastructure that will allow the linking of researchers, clinical experimenters, practicing physicians, and the public in ways heretofore not imagined. Medical researchers must be leaders and advocates for the whole research enterprise in the 21st Century. PMID- 10850898 TI - Postburn itching, pain, and psychological symptoms are reduced with massage therapy. AB - Twenty patients with burn injuries were randomly assigned to a massage therapy or a standard treatment control group during the remodeling phase of wound healing. The massage therapy group received a 30-minute massage with cocoa butter to a closed, moderate-sized scar tissue area twice a week for 5 weeks. The massage therapy group reported reduced itching, pain, and anxiety and improved mood immediately after the first and last therapy sessions, and their ratings on these measures improved from the first day to the last day of the study. PMID- 10850899 TI - Body image issues for children and adolescents with burns. AB - Body image is a far-reaching, multidimensional, dynamic concept. Because burn injuries threaten the integrity of both the physical and psychologic identity, body-image issues related to burn injuries appear to be a meaningful area of investigation. Little research has been done to directly assess body-image issues for children and adolescents with burns. We reviewed the general findings that body-image adaptation occurs and is influenced by gender, social support, burn severity, overall adjustment, and developmental stage. It is suggested that body image revision, if it occurs, is largely successful, but body-image issues may not be directly related to psychosocial adjustment after a burn injury. PMID- 10850900 TI - A 10-year experience with toxic epidermal necrolysis. AB - Toxic epidermal necrolysis is a devastating medication-induced desquamation disorder with a reported mortality rate of 30% to 60% in adults. Data from previously reported series suggest that age, delay in referral to a burn center, total body surface area (TBSA) involvement, and systemic steroid treatment are poor prognostic indicators. We reviewed the records of 39 patients treated in our burn center over the past 10 years and found that the mortality rate was significantly correlated with age, thrombocytopenia, and delay in presentation. Steroid treatment and TBSA involvement were not significantly related to the mortality rate. Thirty-nine adult patients with greater than 20% TBSA epithelial necrosis were cared for in our center from January 1987 to March 1998. Wounds were treated with topical antimicrobial medications and porcine xenografts in a bacteria-controlled nursing unit. We reviewed the records of these patients for 28 clinical characteristics and looked for clinical correlates of mortality by single analysis of variance. The mortality rate was 44% (17 of 39 patients); the cause of death was most commonly multiple-organ dysfunction syndrome, for which a microbial etiologic agent was not always identified. Autopsies were performed on 11 of the 17 patients who died; there was evidence of multiple-organ damage. The patients who survived and the patients who died did not differ significantly in TBSA epithelial necrosis (66%+/-6% vs 72%+/-5%, respectively), admission platelets, number of nosocomial infections, number of complications, preadmission exposure to steroids, or extent of mucosal involvement. When compared with the patients who died, the patients who survived were (1) 20 years younger (47.5+/ 4.2 years vs 64.5+/-5.3 years), (2) admitted to the hospital sooner after the onset of their rash (3.5+/-0.4 days vs 5.9+/-1.0 days), (3) much less likely to experience early thrombocytopenia (platelet nadir, 154+/-24 vs 70+/-18), (4) more likely to be febrile on presentation, and (5) less likely to have been treated with antibiotics before referral to our unit. These differences were statistically significant. The most common etiologic agents were antibiotics, anticonvulsants, and nonsteroidal anti-inflammatory drugs. Our results for a group of older patients with toxic epidermal necrolysis with extensive skin involvement suggest that age, delay in hospitalization, thrombocytopenia, and early empiric antibiotic treatment are associated with a poor prognosis. PMID- 10850901 TI - Rating the resolving hypertrophic scar: comparison of the Vancouver Scar Scale and scar volume. AB - The increased focus of research interests and clinical documentation on outcomes demands that evaluation tools provide reliable and valid data. The Vancouver Scar Scale (VSS) was developed to provide a more objective measurement of burn scars; however, the validity (a test's ability to measure the phenomenon for which it was designed) of the VSS has not been tested. To examine the construct validity of the VSS, we compared it with scar volume, which has established face validity. Burn scars were evaluated monthly for a minimum of 7 months. Three scar volume measurements were performed on each scar. In addition, 3 independent examiners completed the VSS for the same scar. The data generated by these 2 measurements were used to establish the following: (1) the interrater agreement estimated by interclass correlation coefficient, (2) convergence validity, (3) the sensitivity of the assessments to discriminate changes in the scar over time, and (4) the prevalence of related parameters that are not currently being captured by the VSS. In an attempt to address some of the deficiencies of the VSS, we propose several modifications. We anticipate that these changes will increase the reliability and validity of the VSS through an increase in the awareness that training in the use of this scale is required, through improvement in the quality of the subscales, and through the documentation of additional pertinent information. PMID- 10850902 TI - hsp70, hsp32, and grp78 are increased in thermally injured skin with and without antithrombin(human) concentrate infusion. AB - An acquired deficiency of antithrombin (AT), an anti-inflammatory protein, develops in patients with thermal injuries. Skin thermotolerance is regulated by heat shock protein (hsp) genes. hsp70, hsp32, hsp27, and glucose-regulated protein78 (grp78) were studied in burned and unburned human skin to determine whether correction of the AT deficiency modulated the intensity of expression of these proteins. Fifty-four human skin samples were prepared by Western blot analysis: 11 unburned and 22 burned control skin samples and 7 unburned and 14 burned skin samples from patients treated with AT(Human), or AT(H). The intensity of hsp32 expression in burned AT(H)-treated skin (P < .001) and in burned control skin (P < .01) was significantly increased compared with unburned control skin. The intensity of expression of hsp70 was statistically significant in burned AT(H)-treated skin compared with unburned control skin (P < .02), as was that of grp78 (P < .01). Thermally injured skin with or without AT(H) treatment had an increased expression of hsp70, hsp32, and grp78 compared with unburned control skin. PMID- 10850903 TI - Polyclonal antibodies to rat interleukin-6 attenuate fever in rats after burn injuries: a preliminary report. AB - Rats with 50% total body surface area full-thickness burn injuries have sustained increases in body temperature without evidence of sepsis. In this animal model, the only known endogenous pyrogen consistently present in plasma is interleukin-6 (IL-6). We investigated the hypothesis that IL-6 acts as an endogenous pyrogen and produces the observed increase in body temperature. Body temperature and activity index were recorded every 5 minutes for control rats and rats with burns (50% total body surface area scald burns) for a period before the burn injuries and through the eighth day after the burn injuries. On the fifth day after the burn injuries, between 8 and 9 AM, control animals and animals with burns were given an intra-arterial dose of IL-6 polyclonal neutralizing antibody (IL-6 pAb) that was equivalent to one half of the neutralizing dose of IL-6 pAb. This calculation was based on plasma levels of IL-6 obtained from rats with burns in previous studies and on the neutralizing capability of the IL-6 used. On the sixth and seventh days after the burn injuries, a full neutralizing dose of IL-6 pAb was given. Blood samples were obtained for IL-6 assays at baseline and after treatment with IL-6 pAb but were lost because of the failure of an ultracold freezer. A one half neutralizing dose of IL-6 pAb produced no discernible effect on the body temperature of the control animals or of the animals with burns. On the sixth and seventh days after the burn injury, IL-6 pAb produced 45% and 34% respective decreases in body temperature compared with the animals with burns that received saline solution (P < .05 for both comparisons). On the eighth day after the burn injuries, the effect had completely dissipated. We concluded that our results support the idea that increased circulating levels of IL-6 may be causally related to fever that occurs in nonseptic rats with large burn wounds. PMID- 10850904 TI - Effects of an L-selectin antibody on the pulmonary and systemic manifestations of severe smoke inhalation injuries in sheep. AB - Sheep were treated with either lymphocyte adhesion molecule (LAM)1-3, an antibody against L-selectin, (40 mg 1 hour before smoke inhalation and 35 mg 24 hours after smoke inhalation; n = 6) or equivalent volumes of 0.9% saline solution (n = 6). After the smoke inhalation injuries, the PaO2/FIO2 ratio declined in both groups until 40 hours after the injuries, when a trend toward improvement was noted in the group that received LAM1-3. Lung lymph flow increased in both groups until 36 hours after the smoke inhalation injuries and then significantly decreased in the group that received LAM1-3. Forty-eight hours after the smoke inhalation injuries, there was a significant decrease in the ratio of wet-dry lung weight and in preservation of the reflection coefficient in the group that received LAM1-3 (P < .05). Histopathologic examination showed no differences between the groups in the pulmonary morphology associated with smoke inhalation. A reduction in splanchnic blood flow was noted in the control group (P < .05); this reduction was attenuated by treatment with LAM1-3. The delayed pulmonary effects and improved splanchnic blood flow suggested that LAM1-3 attenuated the development of a systemically induced secondary lung injury rather than of the primary lung injury associated with smoke inhalation. PMID- 10850905 TI - Chromic acid burns: early aggressive excision is the best method to prevent systemic toxicity. AB - Chromium poisoning can occur from the cutaneous absorption of chromium from burns that are as small as 1% of the total body surface area. In this case report, we describe a patient with 10% total body surface area burns caused by hot chromic acid. The amount of chromium removed by peritoneal dialysis and the amount of chromium in the urine were estimated, as well as the chromium content in the excised skin, serum, and red blood cells. The extent of chromium load from this type of injury and subsequent risk of systemic poisoning is not predictable, and we therefore believe that systemic toxicity is best prevented by early excision of the burned skin. PMID- 10850906 TI - Drug-induced linear immunoglobulin A bullous disease that clinically mimics toxic epidermal necrolysis. AB - Drug-induced linear immunoglobulin A bullous disease is a subepidermal blistering disorder that most commonly occurs after exposure to vancomycin. It can clinically mimic toxic epidermolytic necrolysis. We describe an 87-year-old white woman in whom linear immunoglobulin A bullous disease developed while she was taking vancomycin and phenytoin. A few days after the linear immunoglobulin A bullous disease developed, both medications were discontinued. No new bullae developed, and the eruption completely resolved within 2 weeks. The patient was treated with only topical therapy. PMID- 10850907 TI - Hydrogen sulfide inhalation injury. AB - Hydrogen sulfide is a colorless, noxious gas with the distinctive smell of rotten eggs. This compound is a powerful reducing agent that is encountered in a number of industrial processes. When hydrogen sulfide is present, it exposes workers to the potentially lethal effects of the rapid hypoxemia that results from exposure to this agent. The "warning sign" is the characteristic smell of rotten eggs; this smell should alert anyone in the area that a potentially serious risk exists. The immediate removal of the victim and administration of high-flow oxygen is essential. Neurologic sequelae may require anticonvulsants and care must be exercised to observe for cardiac, hepatic, and renal insufficiency. Depending on the concentration, hydrogen sulfide can rapidly overcome a potential victim. PMID- 10850908 TI - Biodebridement: a case report of maggot therapy for limb salvage after fourth degree burns. AB - The wound healing and antimicrobial properties of maggots are well known. Maggot debridement therapy has been used for the treatment of various conditions. For maggot debridement therapy, the larvae of the blowfly are applied over necrotic or nonhealing wounds. We used maggot debridement therapy with the larvae of Phaenicia sericata for limb salvage after bilateral lower extremity fourth-degree burns. PMID- 10850909 TI - Gait variables of patients after lower extremity burn injuries. AB - Functional ambulation is an expected outcome of physical therapy after burn injuries on the lower extremities. The purpose of this study was to document temporal and spatial gait parameters of adult patients with the use of the GAITRite system (CIR Systems Inc, Clifton, NJ) after the patients were burned on their lower extremities and to compare these results with previous data reported for normal subjects. Twenty-five adults with lower extremity burns (19 men and 6 women; mean age, 35.6+/-8.3 years) were evaluated within 5 days of discharge from an acute care facility. The GAITRite system, which consists of an electronic walkway that contains 6 sensor pads encapsulated in a rolled-up carpet, was used to collect temporal and spatial variables. The patients walked at their preferred rate of ambulation and completed 2 passes; the 2 passes were then averaged by the software to determine the patients' gait parameters. A 2-tailed t test was used for comparison of the mean values for the patients and the previously published data. The results indicated that for both men and women, cycle time and base of support were significantly higher (P < or = .01) in the patients with burn injuries than in normal subjects. For men, all of the remaining parameters were significantly lower (P < or = .01) in the patients with burns except stride length, which was not significantly different (P > .05). For women, stance time as a percentage of the gait cycle and cadence, velocity, step length, and stride length, were all significantly lower (P < or = .01) in the patients with burn injuries, whereas double support as a percentage of the gait cycle was not significantly different (P > .05) between the 2 groups. These results indicate that immediately after an acute care hospitalization, patients with lower extremity burns have significantly different gait patterns than gender-and age matched normal subjects. Future studies are necessary to determine whether these impairments in gait limit the functional abilities of a patient. PMID- 10850910 TI - Burns caused by medical therapy. AB - A burn injury may occur as an unexpected consequence of medical treatment. We examined the burn prevention implications of injuries received in a medical treatment facility or as a direct result of medical care. The records of 4510 consecutive admissions to 1 burn center between January 1978 and July 1997 were retrospectively reviewed. A cohort of 54 patients burned as a result of medical therapy was identified and stratified by location (home vs medical facility) and by mechanism of injury. Twenty-two patients were burned in a medical treatment facility, including 12 patients who were burned as a result of careless or unsupervised use of tobacco products. Thirty-two patients were burned as a result of home medical therapy, including 9 patients who had scald injuries from vaporizers, 8 patients who were burned by simultaneous use of cigarettes and home nasal oxygen therapy, and 11 patients who were burned by therapeutic application of heat. In contrast to previous studies, no patient was burned by the use of medical laser devices. To further decrease burn risk from medical therapy we advocate the prohibition of cigarette smoking in any medical facility. Continued tobacco use may represent a contraindication to home oxygen therapy. Given the lack of proof of efficacy combined with the potential for burn injury, the use of vaporizers to treat upper respiratory symptoms should be discouraged. Patients with diminished sensation or altered mental status are at increased risk of burn injury from bathing or topical heat application and merit closer monitoring during these activities. PMID- 10850911 TI - Enteral glutamine administration prevents the decrease in cell energy charge potential produced in ileum after a skin burn in the rat. AB - A skin burn produces a decrease in cell energy charge potential (ECP), a marker of cell function, in the liver and other organs. The process appears to be oxidant induced because reduced glutathione (GSH) appears to be protective. The gut mucosal barrier is also known to be altered after burns and appears to be improved by glutamine, which is a source of energy and GSH production. Neither the relationship between the gut barrier change and the ECP of the gut barrier nor the mechanism of glutamine protection has been defined. The effect of a 20% total body surface area, full-thickness burn in a rat on ECP and GSH content in the ileum and the liver and the effect of oral glutamine (1 g/kg/d) on any changes were studied. Three groups of rats (14 rats per group) were studied: a control group, a group with burns alone, and a group with burns that were given glutamine. Half of the rats were killed 1 day after the burn, and the other half were killed 6 days after the burn. ECP did not decrease 1 day after the burn, but 6 days after the burn, ECP decreased from a control of 0.53+/-0.06 to 0.41+/-0.01 in the liver and from 0.64+/-0.04 to 0.29+/-0.04 in the ileum. In the ileum, glutamine prevented the decrease of the ECP (which increased to 0.58+/-.03), but in the liver, glutamine did not alter the ECP (which remained at 0.42+/-0.05). The GSH content was unchanged after the burn injury in both organs compared with the value before the burn injury. It can be concluded that a delayed decrease in ECP in the ileum occurs after a burn injury. Orally administered glutamine prevents the decrease in ECP, probably by providing cell energy rather than by increasing gut GSH (antioxidant) activity. PMID- 10850912 TI - Implementation of a therapy group at a camp in southern Illinois for children with burn injuries. AB - Pediatric burn injuries result in significant mortality and morbidity. Studies in which the psychosocial effects of burn injuries were investigated have produced mixed results but generally provide some promise that these children can be well adjusted despite serious injuries. However, some children may experience psychologic and social difficulties. To address some of these difficulties, therapy groups were included as part of the programming at a fledgling camp in southern Illinois for children with burn injuries. We present our experiences with this process. Suggestions for future interventions by mental health professionals are also presented. PMID- 10850913 TI - Full-thickness burn to the hand from an automobile airbag. PMID- 10850914 TI - Acute respiratory failure that complicates the resuscitation of pediatric patients with scald injuries. PMID- 10850915 TI - Capsule stabilization for phacoemulsification. PMID- 10850916 TI - Keratectasia after LASIK. PMID- 10850917 TI - Pneumotonometry versus Goldmann tonometry. PMID- 10850918 TI - Photorefractive keratectomy and intraocular pressure measurement. PMID- 10850919 TI - Phacoemulsification and vitrectomy. PMID- 10850920 TI - Endophthalmitis after suture removal. PMID- 10850921 TI - Epithelial deposits after hyperopic LASIK. PMID- 10850922 TI - Problems with unpreserved lignocaine for intraocular use. PMID- 10850923 TI - Clue to complete removal of OVD during phacoemulsification. PMID- 10850924 TI - Hypersensitivity to metallic biomaterials: a review of leukocyte migration inhibition assays. AB - Metal hypersensitivity is a well-established phenomenon occurring in a variety of domestic and workplace settings. Degradation products of metallic biomaterials may mediate metal hypersensitivity. However, little is known about the short- and long-term pharmacodynamics and bioavailability of circulating metal degradation products in vivo. Mechanisms by which in vivo metal sensitivity reactions occur have not been well characterized and the degree to which metal sensitivity may be a predisposing factor for eliciting an overaggressive immune response remains clinically unpredictable. In vitro leukocyte migration inhibition assays have been used for investigating cell-mediated hypersensitivity reactions to biomaterial and biomaterial degradation products. This review provides a historical and technical summary of four in vitro techniques used for determination of leukocyte migration activity: (1) membrane migration or Boyden chamber, (2) capillary tube, (3) leukocyte migration using agarose technique, and (4) collagen gels. It is difficult to determine which, if any, of these techniques is singularly best suited for the investigation of suspected biomaterial-related symptoms in patients. However, Boyden chamber membrane migration testing is recommended for clinical investigations, principally because a high degree of standardized investigator independent materials and methodologies is necessary for compiling and comparing the results of patients tested at various times over the length of an extended study. Ultimately, in vitro migration inhibition testing has the potential to provide a reliable means for predicting some complications and thus enhancing the outcome for patients receiving metallic implants. Continuing improvements in migration inhibition testing methods, used alone or in combination with other immunologic assays, will likely improve assessment of patients susceptible to biomaterial antigen-induced delayed-type hypersensitivity responses. PMID- 10850925 TI - Finite element analysis of thermo-debonding mechanism in dental composites. AB - Finite element method (FEM) has been extensively used for evaluating interfacial status inside biomaterials. This study using FEM was designed to evaluate the thermal stress behavior of a filler-matrix interface. The results were then compared to those of a previous study obtained by a laser thermoacoustic technique (LTAT). The experimental systems (75/25 Bis-GMA/TEGDMA resin reinforced with 0, 25, 50, and 75 wt% 8-microm silanized/unsilanized BaSiO6) as used in the previous study were modeled in this study. The established finite element models were based on coefficient of thermal expansion (CTE) Mismatch Phenomenon. The mechanical properties of the silane coupling agent, such as elastic modulus and thermal expansion coefficient used in the silanized model, were assumed to have optimal heat flux transfer. A third (imaginary) material was proposed to block the transfer of thermal stress between the filler and matrix in the unsilanized model. The thermal load simulation was based on steady-state thermal analysis. The results showed that: (1) The strain energy and interfacial shearing stress calculated from FEM validate the results from the previous LTAT study. (2) Comparing the stress distribution of silanized and unsilanized FEM models, the acoustic signals in LTAT study are mainly derived from debonding of the filler matrix interface of silanized specimens, and from the matrix area of unsilanized specimens. Based on results to date, we conclude that the finite element method may be a powerful tool for exploring thermoacoustic mechanisms of dental composites. PMID- 10850926 TI - Biaxial residual stress states of plasma-sprayed hydroxyapatite coatings on titanium alloy substrate. AB - Biaxial residual stress states of plasma-sprayed hydroxyapatite coatings (HACs) on titanium alloy substrate as a function of plasma power, powder feed rate and coating thickness were studied by X-ray 'sin2 psi' method. The Young's modulus of hydroxyapatite (HA), required for the stress analysis, was measured from the separated free coating by three-point bending test method. It was found that the directions of principal stresses were in proximity to and perpendicular to the spraying direction. The measured Young's moduli of HACs were much lower than the theoretical value reported. The denser, well-melted HAC exhibited a higher residual stress, as compared with the less dense, poor-melting HAC. The denser coatings could be effected by higher plasma power and lower powder feed rate. Significantly, the thicker 200 microm HAC exhibited higher residual stress than that of the thinner 50 microm HAC. The implications of residual stress in HAC for biomaterials are discussed in detail. PMID- 10850927 TI - Chemical gradient in plasma-sprayed HA coatings. AB - The microstructure and inhomogeneous features of plasma-sprayed hydroxyapatite coatings on titanium substrates have been examined using the time-of-flight secondary ion mass spectroscopy (ToF-SIMS), micro-Raman spectroscopy and nano indentation techniques. The crystalline and amorphous areas in coatings can be identified by the elastic modulus difference. The concentration gradient of O and OH ions was detected in the through-thickness direction of coatings. Lack of O and OH ions near the titanium interface implies the existence of phases other than HA, and might result in excessive adsorption of the coatings near the interface in HA-coated Ti implants. PMID- 10850928 TI - Various evaluation techniques of newly formed bone in porous hydroxyapatite loaded with human bone marrow cells implanted in an extra-osseous site. AB - The purpose of this work was to develop qualitative methods for in situ analysis of bone formation in an osteoconductive hydroxyapatite matrix (ENDOBON), loaded with human bone marrow cells (HBMSC) implanted subcutaneously in athymic mice. Samples were taken before implantation (T0), 1, 2, 4 and 6 weeks after implantation. Bone-biomaterial interaction were investigated on undecalcified sections by histological, cytochemical, immunological and molecular biology methodologies. Histological observations were performed in order to observe inflammatory cells, vessels, newly formed bone, woven and lamellar bone. Enzymohistochemistry was carried out to detect positive tartrate resistant acid phosphatase activity (TRAP+). Immunohistochemistry using antibodies against type I collagen and osteocalcin permitted us to characterize the content of the matrix elaborated within the implant. Moreover, in situ hybridization was carried out to discriminate, the implanted human cells from the murine cells, and to evaluate the function of these human cells in osteogenesis. Results demonstrated an early formation of lamellar bone only in the pores of the studied HAP loaded with HBMSC. This bone contained a matrix showing positive reaction for type I collagen and osteocalcin. In situ hybridization identified some of these cells as human cells. At 6 weeks, examination of histological results showed persistance of lamellar bone in the implants. We only found TRAP+ activity in the materials loaded with human bone marrow cells. Molecular hybridization no longer revealed positive cells for the human DNA probe. All these results indicate that the various evaluation techniques performed on undecalcified sections, permit us to evaluate the response of human bone marrow cells in HAP implanted into mice. PMID- 10850929 TI - In vitro surface characterization of a biological patch fixed with a naturally occurring crosslinking agent. AB - The study was designed to characterize the surface properties (including water contact angle, surface tension, protein adsorption, platelet adhesion, and cellular compatibility) of a biological patch fixed with genipin, a naturally occurring crosslinking agent. Fresh and glutaraldehyde-fixed counterparts were used as controls. It was found that both glutaraldehyde and genipin are effective crosslinking agents for biological tissue fixation. Fixation of biological tissue with glutaraldehyde or genipin significantly increased its hydrophilicity and surface tension and reduced its mol ratio of adsorbed fibrinogen to adsorbed albumin as well as the amount of adhered platelet. There were no significant differences in hydrophilicity, surface tension, the mole ratio of adsorbed fibrinogen to adsorbed albumin, and the amount of platelet adhesion between the glutaraldehyde- and genipin-fixed tissues. However, the cellular compatibilities of fresh and the genipin-fixed tissues were significantly superior to the glutaraldehyde-fixed tissue. PMID- 10850930 TI - Gel-impregnated pore membranes with mesh-size asymmetry for biohybrid artificial organs. AB - Membranes based on mechanically supported poly(vinyl alcohol) (PVA) hydrogels with mesh-size asymmetry were developed for potential application in biohybrid artificial organs. The pores of cellulose ester microfiltration membranes were impregnated with a PVA solution, which was lightly crosslinked with glutaraldehyde and then modified under a glutaraldehyde gradient to produce mesh size asymmetry. Permeation experiments were performed with the resulting homogeneous and asymmetric gel-impregnated pore membranes (GIPMs). Creatinine (MW: 113), goat Fab (MW: 50 kD) and human IgG (MW: 150 kD) were used to simulate the molecular size of nutrients, therapeutic proteins, and immunological molecules, respectively. The transport properties of the GIPMs were compared to those of conventional ultrafiltration (UF) and dialysis membranes. Experimental results indicate that GIPMs with mesh-size asymmetry have thickness-normalized creatinine permeabilities that are slightly higher than those in cellulosic UF membranes but as much as 100% greater than those in polysulfone UF or cellulosic dialysis membranes. IgG permeabilities in the GIPMs are from 5 to 50 times lower than those in the UF membranes. Fab permeabilities are 6 to 40 times higher in the UF membranes than those in the GIPMs, but the required permeability for a therapeutic protein is application specific. GIPMs may also be suitable as an alternative for hemodialysis. PMID- 10850931 TI - Effect of maturation on the fluoride release of resin-modified glass ionomer and polyacid-modified composite resin cements. AB - The effect of an early water contact on the fluoride release is studied for the resin-modified glass ionomer cements (RM-GIC) GC Lining LC, PhotacBond, Vitremer and Vitrebond and for the polyacid-modified composite resins (PAM-C) Variglass and Dyract. Six months fluoride release profiles were determined in regularly renewed water (37 degrees C), for the products directly after light curing and after 24 h maturation in a humid atmosphere (85% RH). ANOVA shows that both the short-term and the long-term fluoride release of a RM-GIC are influenced by this maturation. This indicates that direct water contact for this material should be avoided. For the RM-GIC a correlation is found between the initial fluoride release process and the long-term process. For the PAM-C materials, no differences in the fluoride release are found as a function of maturation, indicating that early water contact has no effect. The amounts of fluoride released by PAM-C are low compared to RM-GIC, which can affect their caries preventive potential. The results are explained on the basis of the setting reaction of both types of materials. PMID- 10850932 TI - Light-emitting diode (LED) polymerisation of dental composites: flexural properties and polymerisation potential. AB - The clinical performance of light polymerised dental composites is greatly influenced by the quality of the light-curing unit (LCU) used. Commonly used halogen LCUs have some specific drawbacks such as decreasing of the light output with time. This may result in low degree of monomer conversion of the composites with negative clinical implications. Previous studies have shown that blue-light emitting diode (LED) LCUs have the potential to polymerise dental composites without having the drawbacks of halogen LCUs. Despite the relatively low irradiance of current LED LCUs, their efficiency is close to that of conventional halogen LCUs with more than twice the irradiance. This phenomenon has not been explained fully yet. Hence, more tests of the LED LCU's effectiveness and of the mechanical properties of oral biomaterials processed with LED LCUs need to be carried out. This study investigates the flexural properties of three different composites with three different shades, which were polymerised with either a commercial halogen LCU or an LED LCU, respectively. In most cases no significant differences in flexural strength and modulus between composites polymerised with a halogen LCU or an LED LCU, respectively, were found. A simple model for the curing effectiveness based on the convolution absorption spectrum of the camphorquinone photoinitiator present in composites and the emission spectra of the LCUs is presented. PMID- 10850933 TI - Characterisation of the rheological properties and zeta potential of a range of hydroxyapatite powders. AB - The effect of precipitation temperature, i.e. particle morphology on the rheological properties of a hydroxyapatite (HA) slip was investigated and compared to a commercial HA (batch P120 supplied by Plasma Biotal, Tideswell, Derbyshire, UK). The commercial HA was highly crystalline and had a particle size much larger than the HA precipitated at 60 and 80 degrees C. With no deflocculant addition, the commercial HA had a viscosity much higher compared to the precipitated HA as expected. The commercial HA and the HA precipitated at 60 degrees C showed similar pseudoplastic behaviour, but the HA precipitated at 80 degrees C showed Newtonian behaviour. This was explained by the HA precipitated at 80 degrees C having mean particle size of 82.24 nm, but a much wider particle size distribution. This is confirmed by the electrophoretic mobility measurements which show that the HA precipitated at 80 degrees C has a much lower zeta potential at a 0 wt% addition of deflocculant. Because of the wider particle size distribution, the need to add deflocculant is much reduced. PMID- 10850934 TI - Apoptosis in peri-implant tissue. AB - The authors examined 54 biopsies taken from the tissue surrounding loosened hip joint prostheses. In situ apoptotic cell identification was performed by the detection of single- and double-stranded DNA breaks that occurred in the early stages of apoptosis. Both types of breaks can be revealed by labeling the free 3' OH termini with modified nucleotides (fluoresceine-dUTP) in an enzymatic reaction catalyzed by terminal deoxynucleotidyl transferase (TdT). Results were correlated with the presence of wear debris in the tissue and with the use of bone cement for prosthesis fixation. Apoptotic cells were present in a higher percentage in tissue sections where metal particles were present (24% apoptotic cells) if compared to areas where no wear (6%), or plastic wear (2.8%) or ceramic wear (1.5%) was observed. Apoptosis is neither related to bone cement, nor to the time it takes for the implant to fail. Cell death by apoptosis may be important in implants which release metal ions by corrosion or wear and may have been underestimated up to now, as it is a 'clean' way of cell death, leading to limited damage in the surrounding tissues. PMID- 10850935 TI - Laser-induced acoustic emissions in experimental dental composites. AB - A laser thermoacoustic technique was innovated to evaluate laser-induced acoustic emissions (AEs) in experimental dental composites aged with 75% ethanol solution. Experimental composite systems of 75/25 BisGMA/TEGDMA resin filled with 0, 12.6, 30.0, and 56.5 vol% of 8-microm silanized and unsilanized BaSiO6 were analyzed. The sample size was 4.65 mm (diameter) x 0.5 mm (thick). Aging effects of immersing in 75% ethanol for up to 14 h on AEs were then evaluated. A continuous wave CO2 laser was used to heat the samples. Acoustic emissions were collected as a function of filler fraction, laser power, silanization, and immersion time. Onset of burst-pattern acoustic signals characteristic of fracturing occurred at different laser powers for different tested groups. Acoustic emissions generally increased with laser power, in which lower laser powers produced low-amplitude (45-50 dB) signals; the amplitude distribution (50-85 dB) became more extensive as laser powers increased. After immersion, the lower laser powers could produce the same phenomenon. The higher the filler fraction, the fewer AEs generated. A large percentage AE reduction due to silanization was noted as a function of filler fraction. Unsilanized specimens showed more thermal damages than did silanized ones. PMID- 10850936 TI - Nutritional treatment for acquired immunodeficiency virus-associated wasting using beta-hydroxy beta-methylbutyrate, glutamine, and arginine: a randomized, double-blind, placebo-controlled study. AB - BACKGROUND: The current study was designed to examine whether a combination of three nutrients, consisting of beta-hydroxy-beta-methylbutyrate (HMB), a metabolite of leucine, L-glutamine (Gln) and L-arginine (Arg), each of which has been previously shown to slow muscle proteolysis, could synergistically alter the course of muscle wasting in patients with established acquired immunodeficiency syndrome (AIDS). METHODS: Sixty-eight human immunodeficiency virus (HIV)-infected patients with a documented weight loss of at least 5% in the previous 3 months were recruited from the HIV clinic at Nassau County Medical Center. The subjects were randomly assigned in a double-blind fashion to receive either placebo containing maltodextrin or the nutrient mixture (HMB/Arg/Gln) containing 3 g HMB, 14 g L-glutamine, and 14 g L-arginine given in two divided doses daily for 8 weeks. Body weights (BW) were recorded weekly and lean body mass (LBM) and fat mass (FM) were measured by air displacement plethysmography and by a single computerized tomography (CT) slice through the thigh at 0, 4, and 8 weeks. RESULTS: Forty-three subjects completed the 8-week protocol, (placebo, n = 21; HMB/Arg/Gln, n = 22). At 8 weeks, the subjects consuming the HMB/Arg/Gln mixture gained 3.0 +/- 0.5 kg of BW while those supplemented with the placebo gained 0.37 +/- 0.84 kg (p = .009). The BW gain in the HMB/Arg/Gln-treated subjects was predominantly LBM (2.55 +/- 0.75 kg) compared with the placebo-supplemented subjects who lost lean mass (-0.70 +/- 0.69 kg, p = .003). No significant change in FM gain was observed (0.43 +/- 0.83 kg for the group receiving HMB/Arg/Gln and 1.07 +/- 0.64 kg for the group receiving the placebo, p > .20). Similar percentage changes in muscle mass and fat mass were observed with CT scans. Immune status was also improved as evident by an increase in CD3 and CD8 cells and a decrease in the HIV viral load with HMB/Arg/Gln supplementation. CONCLUSIONS: The data indicate that the HMB/Arg/Gln mixture can markedly alter the course of lean tissue loss in patients with AIDS-associated wasting. PMID- 10850937 TI - Long-term risk of gastrointestinal tumor recurrence after postoperative treatment with recombinant human growth hormone. AB - BACKGROUND: Recombinant human growth hormone (rhGH) promotes protein synthesis, accelerates wound healing, and maintains immune function in the catabolic state. It has also been claimed that rhGH may promote the activation of residual tumor cells, and therefore, increases the risk of tumor recurrence. This study aimed to investigate whether postoperative administration of rhGH increases the long-term risk of tumor recurrences in patients undergoing major gastrointestinal surgery for malignancy. METHODS: Patients (n =104) received three different doses of rhGH (0.075 IU/kg, 0.150 IU/kg, and 0.300 IU/kg) during 5 postoperative days in a placebo-controlled trial. Follow-up was performed for 56-70 months after radical tumor resection. Mean survival period and relapse-free survival were compared with the control group. RESULTS: Complete data were available for 75 patients. Thirty-five percent (n = 20) of all patients treated with rhGH showed tumor recurrences in comparison to 44% (n = 8) of patients given placebo. Mean survival period for rhGH-treated patients was 46 months (median 59 months); in controls, 42 months (median 58 months). The length of relapse-free survival tended to be longer in rhGH-treated patients (2-47 months; median, 21 months) compared with the patients who were given placebo (2-18 months; median, 13 months). CONCLUSIONS: The results demonstrate no evidence for an increased risk of tumor recurrence after rhGH treatment for a short period of time after removal of a gastrointestinal adenocarcinoma. Therefore, the positive metabolic effects of rhGH application can be used safely in the treatment of the postoperative catabolic state in the patient groups investigated. PMID- 10850938 TI - Early versus delayed feeding with an immune-enhancing diet in patients with severe head injuries. AB - BACKGROUND: Although early enteral feeding clearly reduces septic morbidity after blunt and penetrating trauma, data for head-injured patients are conflicting. This study examines the effects of early vs delayed enteral feedings on outcome in patients with severe closed-head injuries with a Glasgow Coma Scale (GCS) score greater than 3 and less than 11. METHODS: Thirty patients were prospectively randomized to receive an immune-enhancing diet (Impact with fiber) early (initiated < 72 hours after trauma) delivered via an endoscopically placed nasoenteric tube (Stay-Put) or late (administered after gastric ileus resolved). This formula was continued for 14 days or until the patient tolerated oral feeding. Goal rate of nutrition was 21 nonprotein cal/kg/d and 0.3 g N/kg/d. RESULTS: Two patients in the early group were excluded due to inability to place the tube, and one patient in the late group died before 72 hours. Five of the remaining 27 died, 1 in the early group and 4 in the late group. There were no significant differences between the groups in length of stay, intensive care unit (ICU) days, significant infection, or GCS score. However, major infection correlated inversely with admission GCS score (R = -0.6, p < .003). Time to reach a GCS score of 14 was significantly longer in patients with significant infections compared with those without (p < .02). CONCLUSIONS: No difference in length of stay or infectious complications is shown in patients with severe closed-head injury when they are given early vs delayed feeding using an immune enhancing formula. Severity of the head injury is closely associated with significant infection. PMID- 10850939 TI - Substrate oxidation in patients with cirrhosis: comparison with other nutritional markers. AB - BACKGROUND: Malnutrition in patients with hepatic cirrhosis is associated with abnormal fuel metabolism marked by reduced glucose oxidation and increased lipid oxidation. A low respiratory quotient (R/Q) indicates reduced glucose and increased lipid oxidation. The aim of this study was to determine if there is an association between substrate oxidation, using indirect calorimetry, and other markers of malnutrition in patients with cirrhosis awaiting liver transplantation. METHODS: Indirect calorimetry (MedGraphics) was used to determine resting energy expenditure and R/Q after an overnight fast. Anthropometric measurements including tricep skinfold thickness (TSF) and midarm muscle circumference (MAMC) were performed and expressed as a percentage of standard values. A 24-hour urinary creatinine excretion was collected to calculate creatinine height index (CHI) and serum albumin. A subjective global assessment (SGA) score was completed on each patient by a dietitian and physician. Spearman rank correlation was used for statistical comparison of R/Q to other nutritional markers. RESULTS: Fifteen patients (7 men, 8 women; mean age, 52 years) were studied. Mean values include: body mass index (BMI) 27.7 kg/m2 +/- 7.3, R/Q 0.78 +/- 0.04, serum albumin 2.97 g/dL +/- 0.56, TSF 71% +/- 27%, MAMC 85% +/- 13%, CHI 75% +/- 18%, and SGA median score A. There was a significant correlation (p < .05) between R/Q and serum albumin, CHI, and SGA score. There was a greater than 90% correlation of SGA estimation by a physician and dietitian. CONCLUSIONS: There is good correlation between R/Q values and serum albumin, CHI, and SGA score. BMI and anthropometric measurements may suggest normal nutrition when, in fact, indirect calorimetry (R/Q) suggests changes consistent with abnormal fuel metabolism and poor nutrition. R/Q can be a useful adjunct in the nutrition assessment of patients with hepatic cirrhosis. PMID- 10850940 TI - Thiamine deficiency in children with congenital heart disease before and after corrective surgery. AB - BACKGROUND: Malnutrition is common in children with congenital heart disease, while thiamine deficiency (TD) is common in malnutrition, in critically ill children, and in adults with congestive heart failure treated with loop diuretics. Our goal was to determine whether children with congenital heart disease had TD and whether treatment with loop diuretics is related to TD in these patients. METHODS: Twelve children with ventricular septal defect (VSD) treated with furosemide, and 10 children with tetralogy of Fallot (TOF) referred for corrective surgery were consecutively enrolled into a prospective study. Data were collected 24 hours before surgery and 5 days after surgery for nutrition evaluation, medications used, anthropometric measurements, and laboratory markers of malnutrition. Thiamine and pyridoxine deficiencies were evaluated using activated enzyme assays. RESULTS: Seven children (32% of patients) did not meet the recommended daily allowance (RDA) for calories and 18% of patients did not meet the RDA for thiamine intake. Anthropometric measurements were low in both groups, more so in those with VSD, although the difference did not reach statistical significance. Overall, 18% (1/12 with VSD and 3/10 with TOF) of children with congenital heart disease had thiamine deficiency before surgery. Three of the four children with TD had adequate intake of thiamine. Six children (27%) had TD 5 days postsurgery (3 children with VSD and 3 children with TOF). CONCLUSIONS: TD is common in children with congenital heart disease (CHD) referred for corrective surgery both before and after surgery. Our results suggest that neither diuretic treatment nor malnutrition can fully explain the development of TD in these children. PMID- 10850941 TI - Fish oil modulates macrophage P44/P42 mitogen-activated protein kinase activity induced by lipopolysaccharide. AB - BACKGROUND: Mitogen-activated protein kinase (MAPK) cascades represent a major signal system to transduce extracellular signals into cellular responses. Overactivity of MAPK has been implicated in the development of many diseases, including cancer and sepsis. This study investigated the hypothesis that fish oil altered the membrane phospholipid composition and modulated MAPK activity. METHODS: RAW 264.7 cells, a mouse macrophage (Mphi) cell line, were grown in eicosapentaenoic acid (EPA)-rich media (114 micromol/L) for 48 hours. Mphi were washed and exposed to Escherichia coli lipopolysaccharide (LPS; 1 microg/mL) for 10 minutes. Both total and activated (phosphorylated) portions of MAPK (P44 and P42) were determined by Western blot assays. AP-1 transcription factor activity was determined by electrophoretic mobility gel shift assays (EMSA). Mphi tumor necrosis factor (TNF) mRNA expression was measured by Northern blot assays. RESULTS: LPS stimulation induced RAW cell phosphorylation of P44/P42. In contrast, RAW cells grown in EPA-rich media had less P44/P42 activation in the presence of LPS. Total P44/P42 were not affected by EPA or LPS. Similarly, EPA also inhibited AP-1 activity. Inhibition of P44/P42 activity with PD98059 reduced both AP-1 activity and TNF mRNA expression of LPS-stimulated Mphi. CONCLUSIONS: Our data suggest that fish oil regulates macrophage proinflammatory gene activation, at least in part, by modulating the MAPK activity. PMID- 10850942 TI - Does the route of feeding modify gastric emptying in rats? AB - BACKGROUND: It has been shown that the pattern of previous nutrient intake can influence gastric emptying. However, the effect of the absence of enteral stimulation in the setting of a normal energy supply on gastric emptying has not been examined. The aim of this study was to determine whether the absence of enteral stimulation during total parenteral nutrition (TPN) could modify gastric emptying in rats. METHODS: Two experiments were performed. First, gastric emptying of a peptone meal was compared between rats receiving TPN, oral liquid diet (same solution as TPN), or regular diet (control group) for 10 days. In the second experiment, gastric emptying of two test meals (40% peptone and 25% glucose) was studied before and after rats received TPN or intragastric nutrition (same solution as TPN) for 10 to 12 days. RESULTS: In experiment 1, gastric emptying of 40% peptone in the TPN and liquid diet groups was slower than that in the control group. This difference was significant between the TPN group and the control group (p < .01) but not between the liquid diet and control groups (p = .076). Gastric emptying of this meal in the TPN and liquid diet groups was similar. In experiment 2, no difference in gastric emptying of 40% peptone or 25% glucose was found between rats receiving TPN and those receiving intragastric nutrition for 10 to 12 days. CONCLUSIONS: The composition of diet not the route of feeding is important in the modification of gastric emptying by the pattern of previous nutrient intake. PMID- 10850943 TI - Foscarnet-induced electrolyte abnormalities in a bone marrow transplant patient receiving parenteral nutrition. AB - Cytomegalovirus (CMV) is a serious complication of immunosuppressed patients receiving bone marrow transplantation. Foscarnet, a pyrophosphate analog, has been used in the treatment of CMV infections. Renal impairment and electrolyte abnormalities are potential adverse reactions associated with the use of foscarnet. We report a case of significant electrolyte changes after initiation of foscarnet in a bone marrow transplant patient receiving parenteral nutrition. PMID- 10850944 TI - Drug-induced electrolyte disorders in patients receiving parenteral nutrition. PMID- 10850945 TI - Treatment of esophageal Crohn's disease by enteral feeding via percutaneous endoscopic gastrostomy. AB - BACKGROUND: Crohn's disease of the esophagus is rare, and medical treatment often ineffective. Complications such as abscess and fistula may arise, and the morbidity of surgery is high. METHODS: Two cases of refractory esophageal Crohn's disease were confirmed by endoscopy and biopsy. Percutaneous endoscopic gastrostomies (PEGs) were inserted and used for enteral nutrition for 9 and 1 month, respectively. RESULTS: The PEGs were well tolerated. Symptoms subsided rapidly, and later gastroscopies confirmed healing of the esophageal ulcers. No complications occurred, and the gastrostomy sites closed quickly after removal of the tubes, with minimal scarring. CONCLUSIONS: Enteral feeding via PEG appears to be safe and well tolerated and may be of great value in the management not only of esophageal Crohn's disease but also of refractory disease at other sites. PMID- 10850946 TI - Sleeve thrombus fibrosis causing neck pain. AB - BACKGROUND: Central venous catheter occlusion due to thrombus formation is the most common cause for malfunction of long-term indwelling catheters. The exact pathology and pathogenesis of this common complication is misunderstood because of an array of terms being used interchangeably in the literature. This article identifies the pathogenesis and symptoms and gives suggestions for the treatment of central venous catheter-related thrombosis. METHODS: Our case report is of a girl who had a percutaneously placed left subclavian central venous line attached to her vascuport. After 9 months she developed severe pain over the catheter tubing on injection into the port. Surgical exploration of the catheter tubing in the infraclavicular region demonstrated that the pain was due to retrograde flow around the catheter within a sleeve thrombus. CONCLUSIONS: In this type of patient, all precautions should be taken to minimize the risk of thrombus formation, including considering the prophylactic use of anticoagulants. PMID- 10850947 TI - Microbiological quality control study of "all-in-one" total parenteral nutrition admixtures. AB - A microbiological quality control study of parenteral nutrition (PN) is presented. The study was carried out in the Pharmacy and Microbiology Services of the Hospital de la Santa Creu i Sant Pau (Barcelona, Spain) from April 1991 to May 1998 and represents more than 25,000 lipid-containing PN admixtures prepared during that 7-year period. A randomized sampling procedure according to the cumulative sum control charts was performed and a 0.45-microm membrane filtration method was used as the microbiological test. Validation showed that the method is sensitive for sterility tests. Bacterial growth was observed on 59 filters (4.58% of the samples). A second control was made for these positive outcomes, and bacterial growth was found in only two cases. The patients' clinical history was reviewed and no relation between the microorganisms of the nutrition and the results obtained from blood cultures was observed. The quality control study revealed a low incidence of contamination in our PN preparation process. PMID- 10850948 TI - pH and concentration of bilirubin in feeding tube aspirates as predictors of tube placement. PMID- 10850949 TI - Promotion of a favorable gut flora in inflammatory bowel disease. AB - Recent evidence suggests that the composition of colonic flora plays a role in intestinal inflammation in inflammatory bowel disease (IBO). This review examines the evidence that altering the concentrations of colonic bacteria might benefit patients with this condition. PMID- 10850950 TI - Glutamine supplementation and intestinal permeability in Crohn's disease. PMID- 10850951 TI - A kinetic study on the interaction between tazobactam (a penicillanic acid sulphone derivative) and active-site serine beta-lactamases. AB - The interaction between tazobactam and several chromosome- and plasmid-encoded (TEM, SHV, PSE types) class A and C beta-lactamases was studied by spectrophotometry. Tazobactam behaved as a competitive inhibitor or inactivator able to restore in several cases the efficiency of piperacillin as a partner beta lactam. A detailed kinetic analysis permitted measurement of the acylation efficiency for some cephalosporinases and broad-spectrum beta-lactamases; the presence of a turn-over of acyl-enzyme complex was also evaluated. PMID- 10850952 TI - Selective inhibition of monoamine oxidase B by aminoethyl substituted benzyl ethers. AB - Aminoethyl 3-chlorobenzyl ether was shown previously (Ding, C.Z. and Silverman, R.B. (1993). Bioorg. Med. Chem. Lett., 3, 2077-2078) to be a potent and selective time-dependent, but reversible inhibitor of monoamine oxidase B (MAO B). Based on this result, a series of novel aminoethyl substituted benzyl ethers was synthesized and the compounds were examined as potential inhibitors of both isozymic forms of MAO. Each compound in the series inhibits both MAO A and MAO B competitively, and IC50 values for each compound were determined. In general, the B isozyme is much more sensitive to these inhibitors than the A isozyme (except for the o- and p-substituted nitro analogues), in some cases by more than two orders of magnitude. The selectivity in favor of MAO B inhibition is relatively high for all of the meta-substituted analogues and quite low for all of the ortho substituted analogues. Having the substituent at the ortho-position is most favorable for MAO A inhibition. With MAO B the meta-analogues were, in general, more potent than the corresponding ortho- and para-analogues with respect to their reversible binding constants. The meta-iodo analogue is the most potent analogue. PMID- 10850953 TI - Carbonic anhydrase inhibitors. Synthesis of topically effective intraocular pressure lowering agents derived from 5-(omega-aminoalkylcarboxamido)-1,3,4 thiadiazole-2-sulfonamide. AB - Reaction of the acyl chlorides of phthalimido-glycine or phthalimido-beta-alanine with 5-amino-1,3,4-thiadiazole-2-sulfonamide afforded after hydrazinolysis and deprotection of the phthalimido group the corresponding 5-(omega aminoalkylcarboxamido)-1,3,4-thiadiazole-2-sulfonamides. Reaction of 5-(beta aminoethylcarboxamido)-1,3,4-thiadiazole-2-sulfonamide with sulfonyl halides or acyl halides afforded a series of compounds possessing beta alkyl/arylsulfonyl/carbonylamidoethylcarboxamido moieties in the 5 position of the thiadiazole-2-sulfonamide ring. The new derivatives were efficient inhibitors of three carbonic anhydrase (CA) isozymes, CA I, II (cytosolic forms) and IV (membrane-bound form), but especially against CA II and CA IV (in nanomolar range), the two isozymes known to play an important role in aqueous humor secretion within the ciliary processes of the eye. Some of the synthesized inhibitors possessed good water solubility (as hydrochlorides or sodium salts) and were applied as 2% solutions directly into the eye of normotensive or glaucomatous albino rabbits. Very strong intraocular pressure (IOP) lowering was observed for many of them for prolonged periods of 1-2 h, and the active drug was detected in eye tissues and fluids indicating that the antiglaucoma effect is due to CA inhibition within the eye. PMID- 10850954 TI - Inhibition of glutathione reductase by isoproterenol oxidation products. AB - Oxidative stress induced by catecholamines is a well recognized toxic event. This effect has been extensively observed in the heart, where high levels of catecholamines cause enzyme inhibition, lipid peroxidation, energy depletion and myocardial necrosis. Catecholamines can be converted into o-quinones and undergo cyclization into aminochromes. This process can occur enzymatically or through autoxidation and involves the formation of free radicals. Aminochromes are highly reactive molecules that can cause oxidation of protein sulfhydryl groups and deamination catalysis, among other deleterious effects; in addition, inhibition of some enzymes has been also reported. We have studied the effects of isoproterenol oxidation products (IOP) on glutathione reductase (GR) activity in vitro. Isoproterenol (ISO) autoxidation was conducted at 37 degrees C in the dark, for 4 h at pH 7.0 and this process was monitored by UV spectrophotometry at both 340 and 490nm. Addition of the autoxidized solution to GR in the presence of oxidized glutathione (GSSG) and NADPH showed that IOP inhibits GR in a competitive mode and that this effect increases during the 4 h incubation period. This inhibitory effect of IOP was partially prevented by the addition of reduced glutathione (GSH), L-cysteine and ascorbic acid to the reaction mixtures. PMID- 10850955 TI - Synthesis and mechanism of action of novel thiocarbamate inhibitors of human leukocyte elastase. AB - Several peptidyl thiocarbamate inhibitors of human leukocyte elastase were synthesized in the molecular weight range of 700-800. Two different sequences with lysine at the P3 and ornithine at the P4 positions were synthesized. Most of the inhibitors with large molecular weights showed high inhibitory capacity with Ki values as low as 10(-8)M. Compounds immobilized on poly,alpha,beta-[N-(2 hydroxyethyl)-D,L-aspartamide] (PHEA) polymers with an average molecular weight of 36,000 showed higher inhibitory capacity than their free forms. PMID- 10850956 TI - Non-enzymatic glycosylation (or glycation) and inhibition of the pig heart cytosolic aspartate aminotransferase by glyceraldehyde 3-phosphate. AB - Glyceraldehyde 3-phosphate (Glyc3P), a glycolytic intermediate, non-enzymatically glycosylated (or glycated) and inhibited the pig heart cytoplasmic aspartate aminotransferase (cAAT). Glyc3P (5.0 mM) decreased cAAT activity by 47% after 1 min at 23 degrees C. cAAT activity remained unchanged after a 24h incubation with either glucose 6-phosphate (5.0 mM) or ribose 5-phosphate (5.0 mM). Increasing the incubation pH from 6.4 to 7.8 or the incubation temperature from 23 degrees C to 50 degrees C enhanced Glyc3P's inhibitory effect on cAAT activity. Glyc3P (250 500 microM) decreased the thermal stability of cAAT as evidenced by lowering the Tm or temperature that caused a 50% irreversible loss of cAAT activity (69 degrees C, control; 58.5 degrees C, 500 microM Glyc3P). Glyc3P decreased cAAT amino group content and increased glycation products, which were measured by adduct formation, fluorescence and protein crosslinking. PMID- 10850957 TI - Interaction of pig kidney and lentil seedling copper-containing amine oxidases with guanidinium compounds. AB - The effect of guanidinium compounds on the catalytic mechanism of pig kidney and lentil seedling amine oxidases has been investigated by polarographic techniques and spectroscopy. Guanidine does not inhibit the lentil enzyme and is a weak inhibitor for pig kidney amine oxidase (Ki=1 mM), whereas aminoguanidine is an irreversible inhibitor of both enzymes, with a Ki value of 10(-6) M. 1,4 Diguanidino butane (arcaine) is a competitive inhibitor for both pig and lentil amine oxidases. Amiloride is a competitive inhibitor for pig enzyme, but upon prolonged incubation with this drug the enzyme gradually loses its activity in an irreversible manner. PMID- 10850958 TI - Stressing Rac, Ras, and downstream heat shock protein 70. PMID- 10850959 TI - Protein kinase C and myocardial biology and function. PMID- 10850960 TI - Apoptosis and heart failure: A critical review of the literature. PMID- 10850961 TI - Series of exon-skipping events in the elastic spring region of titin as the structural basis for myofibrillar elastic diversity. AB - Titins are megadalton-sized filamentous polypeptides of vertebrate striated muscle. The I-band region of titin underlies the myofibrillar passive tension response to stretch. Here, we show how titins with highly diverse I-band structures and elastic properties are expressed from a single gene. The differentially expressed tandem-Ig, PEVK, and N2B spring elements of titin are coded by 158 exons, which are contained within a 106-kb genomic segment and are all subject to tissue-specific skipping events. In ventricular heart muscle, exons 101 kb apart are joined, leading to the exclusion of 155 exons and the expression of a 2.97-MDa cardiac titin N2B isoform. The atria of mammalian hearts also express larger titins by the exclusion of 90 to 100 exons (cardiac N2BA titin with 3.3 MDa). In the soleus and psoas skeletal muscles, different exon skipping pathways produce titin transcripts that code for 3.7- and 3.35-MDa titin isoforms, respectively. Mechanical and structural studies indicate that the exon skipping pathways modulate the fractional extensions of the tandem Ig and PEVK segments, thereby influencing myofibrillar elasticity. Within the mammalian heart, expression of different levels of N2B and N2BA titins likely contributes to the elastic diversity of atrial and ventricular myofibrils. PMID- 10850962 TI - Mechanical stress-induced heat shock protein 70 expression in vascular smooth muscle cells is regulated by Rac and Ras small G proteins but not mitogen activated protein kinases. AB - -Previous studies have documented that acute elevation in blood pressure results in heat shock protein (hsp) 70-mRNA expression followed by hsp70-protein production in rat aortas. In this article, we provide evidence that mechanical forces evoke rapid activation of heat shock transcription factor (HSF) and hsp70 accumulation. In our study, Western blot analysis demonstrated that hsp70-protein induction peaked between 6 and 12 hours after treatment with cyclic stain stress (60 cycles/minute, up to 30% elongation). Elevated protein levels were preceded by hsp70-mRNA transcription, which was associated with HSF1 phosphorylation and activation stimulated by mechanical forces, suggesting that the response was regulated at the transcriptional level. Conditioned medium from cyclic strain stressed vascular smooth muscle cells (VSMCs) did not result in HSF-DNA-binding activation. Furthermore, mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases, c-Jun NH(2)-terminal protein kinases or stress-activated protein kinases, and p38 MAPKs, were also highly activated in response to cyclic strain stress. Inhibition of extracellular signal-regulated kinase and p38-MAPK activation by their specific inhibitors (PD 98059 and SB 202190) did not influence HSF1 activation. Interestingly, VSMC lines stably expressing dominant-negative rac (rac N17) abolished hsp-protein production and HSF1 activation induced by cyclic strain stress, whereas a significant reduction of hsp70 expression was seen in ras N17-transfected VSMC lines. Thus, our findings demonstrate that cyclic strain stress-induced hsp70 expression is mediated by HSF1 activation and regulated by rac and ras GTP-binding proteins. Induction of hsp70 could be important in maintaining VSMC homeostasis during vascular remodeling in response to hemodynamic stimulation. PMID- 10850963 TI - Folic acid reverts dysfunction of endothelial nitric oxide synthase. AB - 5-methyltetrahydrofolate (MTHF), the active form of folic acid, has been reported to restore NO status in hypercholesterolemic patients. The mechanism of this effect remains to be established. We assessed the effects of L- and D-MTHF on tetrahydrobiopterin (BH(4))-free and partially BH(4)-repleted endothelial NO synthase (eNOS). Superoxide production of eNOS and the rate constants for trapping of superoxide by MTHF were determined with electron paramagnetic resonance using 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) as spin trap for superoxide. NO production was measured with [(3)H]arginine citrulline conversion or nitrite assay. The rate constants for scavenging of superoxide by L- and D-MTHF were similar, 1.4 x 10(4) ms(-1). In BH(4)-free eNOS, L- and D-MTHF have no effect on enzymatic activity. In contrast, in partially BH(4)-repleted eNOS, we observe a 2-fold effect of MTHF on the enzymatic activity. First, superoxide production is reduced. Second, NO production is enhanced. In cultured endothelial cells, a similar enhancement of NO production is induced by MTHF. In the present study, we show direct effects of MTHF on the enzymatic activity of NO synthase both in recombinant eNOS as well as in cultured endothelial cells, which provides a plausible explanation for the previously reported positive effects of MTHF on NO status in vivo. PMID- 10850964 TI - Atrial natriuretic factor binding to its receptor is dependent on chloride concentration: A possible feedback-control mechanism in renal salt regulation. AB - Although considerable evidence indicates a role for atrial natriuretic factor (ANF) in renal salt regulation, other studies have found a lack of natriuretic response to high-plasma ANF under certain physiological and pathophysiological conditions. The mechanism for this apparent insensitivity to ANF is unknown. In the present study, it was found that ANF binding to its receptor requires the presence of chloride and occurs in a chloride concentration-dependent manner. ANF binding was measured using the purified recombinant hormone-binding domain of the ANF receptor in the presence of 0.1 mol/L NaCl or other selected salt. High specific binding was detected in the presence of NaCl, KCl, or NH(4)Cl. However, binding was undetectable when the salt was replaced with NaHCO(3), CH(3)COONa, or CH(3)COONH(4), indicating that binding requires the presence of chloride. Chloride dependence was also found with the native receptor in bovine adrenocortical membrane preparations. ANF binding to the recombinant protein was chloride concentration-dependent over a range from 0.05 to 10 mmol/L, and a half maximum binding was attained at approximately 0.6 mmol/L equivalent chloride concentration. Competitive-binding assays at several fixed concentrations of NaCl showed that lowering chloride concentration caused a decrease in maximum binding but did not alter K(d) values, suggesting that a loss of chloride turns off ANF binding rather than reducing affinity for ANF. Saturation-binding studies showed that excess ANF cannot overcome loss of binding caused by low chloride. Chloride dependent ANF-receptor binding may function as a feedback-control mechanism regulating the ANF-receptor action and, hence, renal sodium excretion. PMID- 10850965 TI - Power-law behavior of beat-rate variability in monolayer cultures of neonatal rat ventricular myocytes. AB - It is known that extracardiac factors (nervous, humoral, and hemodynamic) participate in the power-law behavior of heart-rate variability. To assess whether intrinsic properties of cardiac tissue might also be involved, beat-rate variability was studied in spontaneously beating cell cultures devoid of extracardiac influences. Extracellular electrograms were recorded from monolayer cultures of neonatal rat ventricular myocytes under stable incubating conditions for up to 9 hours. The beat-rate time series of these recordings were examined in terms of their Fourier spectra and their Hurst scaling exponents. A non-0 Hurst exponent was found in 21 of 22 preparations (0.29+/-0.09; range, 0.11 to 0.45), indicating the presence of fractal self-similarity in the beat-rate time series. The same preparations exhibited power-law behavior of the power spectra with a power-law exponent of -1.36+/-0.24 (range, -1.04 to -1.96) in the frequency range of 0.001 to 1 Hz. Furthermore, it was found that the power-law exponent was nonstationary over time. These results indicate that the power-law behavior of heart-rate variability is determined not only by extracardiac influences but also by components intrinsic to cardiac tissue. Furthermore, the presence of power-law behavior in monolayer cultures of cardiomyocytes suggests that beat-rate variability might be determined by the complex nonlinear dynamics of processes occurring at the level of the cellular network, eg, interactions among a large number of cell oscillators or metabolic regulatory systems. PMID- 10850966 TI - Investigation of a truncated cardiac troponin T that causes familial hypertrophic cardiomyopathy: Ca(2+) regulatory properties of reconstituted thin filaments depend on the ratio of mutant to wild-type protein. AB - Familial hypertrophic cardiomyopathy (HCM) is caused by mutations in at least 8 contractile protein genes, most commonly beta myosin heavy chain, myosin binding protein C, and cardiac troponin T. Affected individuals are heterozygous for a particular mutation, and most evidence suggests that the mutant protein acts in a dominant-negative fashion. To investigate the functional properties of a truncated troponin T shown to cause HCM, both wild-type and mutant human cardiac troponin T were overexpressed in Escherichia coli, purified, and combined with human cardiac troponins I and C to reconstitute human cardiac troponin. Significant differences were found between the regulatory properties of wild-type and mutant troponin in vitro, as follows. (1) In actin-tropomyosin-activated myosin ATPase assays at pCa 9, wild-type troponin caused 80% inhibition of ATPase, whereas the mutant complex gave negligible inhibition. (2) Similarly, in the in vitro motility assay, mutant troponin failed to decrease both the proportion of actin-tropomyosin filaments motile and the velocity of motile filaments at pCa 9. (3) At pCa 5, the addition of mutant complex caused a greater increase (21.7%) in velocity of actin-tropomyosin filaments than wild-type troponin (12.3%). These data suggest that the truncated troponin T prevents switching off of the thin filament at low Ca(2+). However, the study of thin filaments containing varying ratios of wild-type and mutant troponin T at low Ca(2+) indicated an opposite effect of mutant troponin, causing enhancement of the inhibitory effect of wild-type complex, when it is present in a low ratio (10% to 50%). These multiple effects need to be taken into account to explain the physiological consequences of this mutation in HCM. Further, these findings underscore the importance of studying mixed mutant:wild-type preparations to faithfully model this autosomal-dominant disease. PMID- 10850967 TI - ADP-Ribosyl cyclase in rat vascular smooth muscle cells: properties and regulation. AB - We investigated whether ADP-ribosyl cyclase (ADPR-cyclase) in rat vascular smooth muscle cells (VSMCs) has enzymatic properties that differ from the well characterized CD38-antigen ADPR-cyclase, expressed in HL-60 cells. ADPR-cyclase from VSMCs, but not CD38 ADPR-cyclase from HL-60 cells, was inhibited by gangliosides (10 micromol/L) GT(1B), GD(1), and GM(3). Preincubation of membranes from CD38 HL-60 cells, but not from VSMCs, with anti-CD38 antibodies increased ADPR-cyclase activity; CD38 antigen was detected both in VSMCs and in HL-60 cells. ADPR-cyclase in VSMC membranes was more sensitive than CD38 HL-60 ADPR cyclase to inactivation by N-endoglycosidase F and to thermal inactivation at 45 degrees C. The specific activity of ADPR-cyclase in membranes from VSMCs was >20 fold higher than in membranes from CD38 HL-60 cells. Most importantly, VSMC ADPR cyclase was inhibited by Zn(2+) and Cu(2+) ions; the inhibition by Zn(2+) was dose dependent, noncompetitive, and reversible by EDTA. In contrast, Zn(2+) stimulated the activity of CD38 HL-60 ADPR-cyclase and other known types of ADPR cyclases. Retinoids act either via the nuclear receptor retinoic acid receptor or retinoid X receptor, including all-trans retinoic acid (atRA), and panagonist 9 cis-retinoic acid-upregulated VSMC ADPR-cyclase; the stimulatory effect of atRA was blocked by actinomycin D and cycloheximide. 1,25(OH)(2)-Vitamin D(3) (calciferol) stimulated VSMC ADPR-cyclase dose dependently at subnanomolar concentrations (ED(50) congruent with 56 pmol/L). Oral administration of atRA to rats resulted in an increase of ADPR-cyclase activity in aorta ( congruent with+60%) and, to a lesser degree, in myocardium of left ventricle (+18%), but atRA had no effect on ADPR-cyclases in lungs, spleen, intestinal smooth muscle, skeletal muscle, liver, or testis. Administration of 3,5,3'-triiodothyronine (T(3)) to rats resulted in an increase of ADPR-cyclase activity in aorta ( congruent with+89%), but not in liver or brain. We conclude the following: (1) ADPR-cyclase in VSMCs has enzymatic properties distinct from "classic" CD38 ADPR cyclase, especially sensitivity to inhibition by Zn(2+) and Cu(2+); (2) ADPR cyclase in VSMCs is upregulated by various retinoids, calcitriol, and T(3) in vitro; and (3) administration of atRA and T(3) increases ADPR-cyclase in aorta in vivo. We suggest that the cADPR signaling system plays an important role in the regulation of VSMC functions in response to steroid superfamily hormones. PMID- 10850968 TI - Temporal events underlying arterial remodeling after chronic flow reduction in mice: correlation of structural changes with a deficit in basal nitric oxide synthesis. AB - To define the cellular events of vascular remodeling in mice, we measured blood flow and analyzed the morphology of remodeled vessels at defined points after a flow-reducing remodeling stimulus for 3, 7, 14, and 35 days. Acute ligation of the left external carotid artery reduced blood flow in the left common carotid artery (LC) compared with sham and contralateral right common carotid arteries (RCs). In morphometric analyses, the decrease in diameter in LCs was reversible by vasodilator perfusion 3 days after ligation, whereas ligation for 7 days or greater resulted in a permanent diameter reduction. Coincident with structural remodeling at day 7 was an increase in cell death in remodeled LCs. Functionally, rings from remodeled LCs contracted to prostaglandin F(2alpha) and relaxed to acetylcholine in a manner identical to that of control arteries. However, remodeled LCs were hypersensitive to the nitrovasodilator sodium nitroprusside (at day 7) and exhibited a marked reduction in basal NO synthesis at 7 and 14 days after ligation. The impairment of endothelial NO synthase function was likely due to post-translational mechanisms, given that endothelial NO synthase mRNA and protein levels did not change in remodeled LCs. These data define the ontogeny of flow-triggered luminal remodeling in adult mice and suggest that endothelial dysfunction occurs during reorganization of the vessel wall. PMID- 10850969 TI - Inability to induce hypertension in normotensive rat expressing AT(1) receptor antisense. AB - Our previous studies have shown that neonatal delivery of angiotensin type 1 receptor antisense (AT(1)R-AS) in a retroviral vector prevents spontaneously hypertensive rats from developing hypertension for life but has no effect on blood pressure (BP) in normotensive animals. Based on these results, we hypothesized that AT(1)R-AS transduction in normotensive rats would protect them from developing experimental hypertension. The present study was designed to evaluate this hypothesis. A single intracardiac administration of AT(1)R-AS by a retroviral-mediated delivery system (LNSV-AT(1)R-AS) in 5-day-old normotensive Sprague-Dawley rats resulted in long-term expression of the AT(1)R-AS without an effect on basal BP. However, angiotensin II (Ang II)-induced BP, dipsogenic responses, and renovascular contractility were significantly attenuated in the LNSV-AT(1)R-AS-treated rats. Chronic infusion of low-dose Ang II (55 ng. kg( )(1). min(-)(1)) in LNSV-alone-treated rats caused a modest increase in BP, profound increase in cardiac hypertrophy, and increased vascular contractility. In contrast, the LNSV-AT(1)R-AS-treated rats were protected from developing these changes after Ang II infusion. These data establish that LNSV-AT(1)R-AS pretreatment protects healthy rats from developing Ang II-dependent hypertension. PMID- 10850970 TI - Cardiotrophic effects of protein kinase C epsilon: analysis by in vivo modulation of PKCepsilon translocation. AB - Protein kinase C (PKC) is a key mediator of many diverse physiological and pathological responses. Although little is known about the specific in vivo roles of the various cardiac PKC isozymes, activation-induced translocation of PKC is believed to be the primary determinant of isozyme-specific functions. Recently, we have identified a catalytically inactive peptide translocation inhibitor (epsilonV1) and translocation activator (psiepsilonRACK [receptors for activated C kinase]) specifically targeting PKCepsilon. Using cardiomyocyte-specific transgenic expression of these peptides, we combined loss- and gain-of-function approaches to elucidate the in vivo consequences of myocardial PKCepsilon signaling. As expected for a PKCepsilon RACK binding peptide, confocal microscopy showed that epsilonV1 decorated cross-striated elements and intercalated disks of cardiac myocytes. Inhibition of cardiomyocyte PKCepsilon by epsilonV1 at lower expression levels upregulated alpha-skeletal actin gene expression, increased cardiomyocyte cell size, and modestly impaired left ventricular fractional shortening. At high expression levels, epsilonV1 caused a lethal dilated cardiomyopathy. In contrast, enhancement of PKCepsilon translocation with psiepsilonRACK resulted in selectively increased beta myosin heavy chain gene expression and normally functioning concentric ventricular remodeling with decreased cardiomyocyte size. These results identify for the first time a role for PKCepsilon signaling in normal postnatal maturational myocardial development and suggest the potential for PKCepsilon activators to stimulate "physiological" cardiomyocyte growth. PMID- 10850971 TI - Mechanism and selectivity of the effects of halothane on gap junction channel function. AB - Volatile anesthetics alter tissue excitability by decreasing the extent of gap junction-mediated cell-cell coupling and by altering the activity of the channels that underlie the action potential. In the present study, we demonstrate, using dual whole-cell voltage-clamp techniques, that coexpression of connexin (Cx) 40 and Cx43 rendered cells more sensitive to uncoupling by halothane than cells that express only Cx40 or only Cx43. The halothane-induced reduction in junctional conductance was caused by decreased channel mean open time and increased channel mean closed time. The magnitude of the effect of halothane on channel open time was least for Cx40-like channels and greatest for heteromeric channels. Thus, the data indicate that halothane gates gap junction channels to the closed state in a dose-dependent and connexin-specific manner. One consequence of the selectivity of halothane is that the profile of single-channel events observed in the presence of halothane may not be quantitatively representative of the population of channels contributing to macroscopic conductance in cells that express more than one connexin. In addition, in tissues that express multiple connexins, such as heart and blood vessels, the capacity of the gap junctions to transmit electrical and chemical signals in the presence of halothane could vary according to the pattern of connexin expression. PMID- 10850972 TI - Ten commandments: lessons from the enzymology of DNA replication. PMID- 10850973 TI - Tol1, a fission yeast phosphomonoesterase, is an in vivo target of lithium, and its deletion leads to sulfite auxotrophy. AB - Lithium is the drug of choice for the treatment of bipolar affective disorder. The identification of an in vivo target of lithium in fission yeast as a model organism may help in the understanding of lithium therapy. For this purpose, we have isolated genes whose overexpression improved cell growth under high LiCl concentrations. Overexpression of tol1(+), one of the isolated genes, increased the tolerance of wild-type yeast cells for LiCl but not for NaCl. tol1(+) encodes a member of the lithium-sensitive phosphomonoesterase protein family, and it exerts dual enzymatic activities, 3'(2'),5'-bisphosphate nucleotidase and inositol polyphosphate 1-phosphatase. tol1(+) gene-disrupted cells required high concentrations of sulfite in the medium for growth. Consistently, sulfite repressed the sulfate assimilation pathway in fission yeast. However, tol1(+) gene-disrupted cells could not fully recover from their growth defect and abnormal morphology even when the medium was supplemented with sulfite, suggesting the possible implication of inositol polyphosphate 1-phosphatase activity for cell growth and morphology. Given the remarkable functional conservation of the lithium-sensitive dual-specificity phosphomonoesterase between fission yeast and higher-eukaryotic cells during evolution, it may represent a likely in vivo target of lithium action across many species. PMID- 10850974 TI - Characterization of a bordetella pertussis diaminopimelate (DAP) biosynthesis locus identifies dapC, a novel gene coding for an N-succinyl-L,L-DAP aminotransferase. AB - The functional complementation of two Escherichia coli strains defective in the succinylase pathway of meso-diaminopimelate (meso-DAP) biosynthesis with a Bordetella pertussis gene library resulted in the isolation of a putative dap operon containing three open reading frames (ORFs). In line with the successful complementation of the E. coli dapD and dapE mutants, the deduced amino acid sequences of two ORFs revealed significant sequence similarities with the DapD and DapE proteins of E. coli and many other bacteria which exhibit tetrahydrodipicolinate succinylase and N-succinyl-L,L-DAP desuccinylase activity, respectively. The first ORF within the operon showed significant sequence similarities with transaminases and contains the characteristic pyridoxal-5' phosphate binding motif. Enzymatic studies revealed that this ORF encodes a protein with N-succinyl-L,L-DAP aminotransferase activity converting N-succinyl-2 amino-6-ketopimelate, the product of the succinylase DapD, to N-succinyl-L,L-DAP, the substrate of the desuccinylase DapE. Therefore, this gene appears to encode the DapC protein of B. pertussis. Apart from the pyridoxal-5'-phosphate binding motif, the DapC protein does not show further amino acid sequence similarities with the only other known enzyme with N-succinyl-L,L-DAP aminotransferase activity, ArgD of E. coli. PMID- 10850975 TI - Identification of Sinorhizobium meliloti genes regulated during symbiosis. AB - RNA fingerprinting by arbitrarily primed PCR was used to isolate Sinorhizobium meliloti genes regulated during the symbiotic interaction with alfalfa (Medicago sativa). Sixteen partial cDNAs were isolated whose corresponding genes were differentially expressed between symbiotic and free-living conditions. Thirteen sequences corresponded to genes up-regulated during symbiosis, whereas three were instead repressed during establishment of the symbiotic interaction. Seven cDNAs corresponded to known or predicted nif and fix genes. Four presented high sequence similarity with genes not yet identified in S. meliloti, including genes encoding a component of the pyruvate dehydrogenase complex, a cell surface protein component, a copper transporter, and an argininosuccinate lyase. Finally, five cDNAs did not exhibit any similarity with sequences present in databases. A detailed expression analysis of the nine non-nif-fix genes provided evidence for an unexpected variety of regulatory patterns, most of which have not been described so far. PMID- 10850976 TI - Evidence for a signaling system in Helicobacter pylori: detection of a luxS encoded autoinducer. AB - Helicobacter pylori possesses a homolog of the luxS gene, initially identified by its role in autoinducer production for the quorum-sensing system 2 in Vibrio harveyi. The genomes of several other species of bacteria, notably Escherichia coli, Salmonella enterica serovar Typhimurium, and Vibrio cholerae, also include luxS homologs. All of these bacteria have been shown to produce active autoinducers capable of stimulating the expression of the luciferase operon in V. harveyi. In this report, we demonstrate that H. pylori also synthesizes a functional autoinducer (AI-2) that can specifically activate signaling system 2 in V. harveyi. Maximal activity is produced during early log phase, and the activity is diminished when cells enter stationary phase. We show that AI-2 is not involved in modulating any of the known or putative virulence factors in H. pylori and that a luxS null mutant has a two-dimensional protein profile identical to that of its isogenic parent strain. We discuss the implications of having an AI-2-like quorum-sensing system in H. pylori and suggest possible roles that it may play in H. pylori infection. PMID- 10850977 TI - Cyclic guanosine-3',5'-monophosphate and biopteridine biosynthesis in Nocardia sp. AB - Nocardia sp. strain NRRL 5646 contains a nitric oxide synthase (NOS) enzyme system capable of generating nitric oxide (NO) from arginine and arginine containing peptides. To explain possible roles of the NOS system in this bacterium, guanylate cyclase (GC) and tetrahydrobiopterin (H(4)B) biosynthetic enzymes were identified in cell extracts and in culture media. Cell extracts contained GC activity, as measured by the conversion of GTP to cyclic guanosine 3',5'-monophosphate (cGMP) at 9.56 pmol of cGMP h(-1) mg of protein(-1). Concentrations of extracellular cGMP in culture media were significantly increased, from average control levels of 45 pmol cGMP liter(-1) to a maximum of 315 pmol liter(-1), in response to additions of GTP, L-arginine, H(4)B, and sodium nitroprusside to growing Nocardia cultures. On the other hand, the NOS inhibitor N(G)-nitro-L-arginine and the GC inhibitor 1H-[1,2, 4]oxadiazole[4,3 a]quinoxalin-1-one both dramatically decreased extracellular cGMP levels. Activities for GTP-cyclohydrase-1, 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase, enzymes essential for H(4)B biosynthesis, were present in Nocardia culture extracts at 77.5 pmol of neopterin and 45.8 pmol of biopterin h( 1) mg of protein(-1), respectively. In Nocardia spp., as in mammals, GTP is a key intermediate in H(4)B biosynthesis, and GTP is converted to cGMP by a GC enzyme system that is activated by NO. PMID- 10850978 TI - Characterization of the fructosyltransferase gene of Actinomyces naeslundii WVU45. AB - Oral actinomycetes produce fructosyltransferase (FTF) enzymes which convert sucrose into polymers of D-fructose, known as levans, and these polymers are thought to contribute to the persistence and virulence of the organisms. A gene encoding FTF was isolated from Actinomyces naeslundii WVU45; the deduced amino acid sequence showed significant similarity to known levansucrases of gram negative environmental isolates but was less similar to FTFs from gram-positive bacteria. A transcriptional start site was mapped by primer extension 70 bp 5' from the putative start codon. Promoter fusions to a chloramphenicol acetyltransferase gene were used to confirm that there was a functional promoter driving ftf expression and to show that sequences located 86 to 218 bp upstream of the transcription initiation site were required for optimal ftf expression. Quantitative slot blot analysis against total RNA from cells grown on different sugars or from different growth phases revealed that ftf was constitutively transcribed. Thus, the A. naeslundii FTF is more similar in primary sequence and the regulation of expression to levansucrases of gram-negative bacteria than gram positive bacteria. PMID- 10850979 TI - Elo1p-dependent carboxy-terminal elongation of C14:1Delta(9) to C16:1Delta(11) fatty acids in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae medium-chain acyl elongase (ELO1) mutants have previously been isolated in screens for fatty acid synthetase (FAS) mutants that fail to grow on myristic acid (C14:0)-supplemented media. Here we report that wild-type cells cultivated in myristoleic acid (C14:1Delta(9))-supplemented media synthesized a novel unsaturated fatty acid that was identified as C16:1Delta(11) fatty acid by gas chromatography-mass spectroscopy. Synthesis of C16:1Delta(11) was dependent on a functional ELO1 gene, indicating that Elo1p catalyzes carboxy terminal elongation of unsaturated fatty acids (alpha-elongation). In wild-type cells, the C16:1Delta(11) elongation product accounted for approximately 12% of the total fatty acids. This increased to 18% in cells that lacked a functional acyl chain desaturase (ole1Delta mutants) and hence were fully dependent on uptake and elongation of C14:1. The observation that ole1Delta mutant cells grew almost like wild type on medium supplemented with C14:1 indicated that uptake and elongation of unsaturated fatty acids were efficient. Interestingly, wild-type cells supplemented with either C14:1 or C16:1 fatty acids displayed dramatic alterations in their phospholipid composition, suggesting that the availability of acyl chains is a dominant determinant of the phospholipid class composition of cellular membranes. In particular, the relative content of the two major phospholipid classes, phosphatidylethanolamine and phosphatidylcholine, was strongly dependent on the chain length of the supplemented fatty acid. Moreover, analysis of the acyl chain composition of individual phospholipid classes in cells supplemented with C14:1 revealed that the relative degree of acyl chain saturation characteristic for each phospholipid class appeared to be conserved, despite the gross alteration in the cellular acyl chain pool. Comparison of the distribution of fatty acids that were taken up and elongated (C16:1Delta(11)) to those that were endogenously synthesized by fatty acid synthetase and then desaturated by Ole1p (C16:1Delta(9)) in individual phospholipid classes finally suggested the presence of two different pools of diacylglycerol species. These results will be discussed in terms of biosynthesis of different phospholipid classes via either the de novo or the Kennedy pathway. PMID- 10850980 TI - Purification and characterization of Sa-lrp, a DNA-binding protein from the extreme thermoacidophilic archaeon Sulfolobus acidocaldarius homologous to the bacterial global transcriptional regulator Lrp. AB - Archaea, constituting the third primary domain of life, harbor a basal transcription apparatus of the eukaryotic type, whereas curiously, a large fraction of the potential transcription regulation factors appear to be of the bacterial type. To date, little information is available on these predicted regulators and on the intriguing interplay that necessarily has to occur with the transcription machinery. Here, we focus on Sa-lrp of the extremely thermoacidophilic crenarchaeote Sulfolobus acidocaldarius, encoding an archaeal homologue of the Escherichia coli leucine-responsive regulatory protein Lrp, a global transcriptional regulator and genome organizer. Sa-lrp was shown to produce a monocistronic mRNA that was more abundant in the stationary-growth phase and produced in smaller amounts in complex medium, this down regulation being leucine independent. We report on Sa-Lrp protein purification from S. acidocaldarius and from recombinant E. coli, both identified by N-terminal amino acid sequence determination. Recombinant Sa-Lrp was shown to be homotetrameric and to bind to its own control region; this binding proved to be leucine independent and was stimulated at high temperatures. Interference binding experiments suggested an important role for minor groove recognition in the Sa Lrp-DNA complex formation, and mutant analysis indicated the importance for DNA binding of the potential helix-turn-helix motif present at the N terminus of Sa Lrp. The DNA-binding capacity of purified Sa-Lrp was found to be more resistant to irreversible heat inactivation in the presence of L-leucine, suggesting a potential physiological role of the amino acid as a cofactor. PMID- 10850981 TI - ComP, a pilin-like protein essential for natural competence in Acinetobacter sp. Strain BD413: regulation, modification, and cellular localization. AB - We recently identified a pilin-like competence factor, ComP, which is essential for natural transformation of the gram-negative soil bacterium Acinetobacter sp. strain BD413. Here we demonstrate that transcription and synthesis of the pilin like competence factor ComP are maximal in the late stationary growth phase, whereas competence is induced immediately after inoculation of a stationary-phase culture into fresh medium. Western blot analyses revealed three forms of ComP, one with an apparent molecular mass of 15 kDa, which correlates with the molecular mass deduced from the DNA sequence, one 20-kDa form, which was found to be glycosylated, and one 23-kDa form. The glycosylation of ComP was not required for its function in DNA binding and uptake. The 20-kDa form was present in the cytoplasmic membrane, the periplasm, and the outer membrane, whereas the 23-kDa form was located in the outer membrane and might be due to a further modification. Immunological data suggest that ComP is not a subunit of the pilus structures. Possible functions of ComP in the DNA transformation machinery of Acinetobacter sp. strain BD413 are discussed. PMID- 10850982 TI - Effects of perturbations of the nitrogenase electron transfer chain on reversible ADP-ribosylation of nitrogenase Fe protein in Klebsiella pneumoniae strains bearing the Rhodospirillum rubrum dra operon. AB - The redox state of nitrogenase Fe protein is shown to affect regulation of ADP ribosylation in Klebsiella pneumoniae strains transformed by plasmids carrying dra genes from Rhodospirillum rubrum. The dra operon encodes dinitrogenase reductase ADP-ribosyltransferase and dinitrogenase reductase-activating glycohydrolase, enzymes responsible for the reversible inactivation, via ADP ribosylation, of nitrogenase Fe protein in R. rubrum. In bacteria containing the dra operon in their chromosomes, inactivation occurs in response to energy limitation or nitrogen sufficiency. The dra gene products, expressed at a low level in K. pneumoniae, enable transformants to reversibly ADP-ribosylate nitrogenase Fe protein in response to the presence of fixed nitrogen. The activities of both regulatory enzymes are regulated in vivo as described in R. rubrum. Genetic perturbations of the nitrogenase electron transport chain were found to affect the rate of inactivation of Fe protein. Strains lacking the electron donors to Fe protein (NifF or NifJ) were found to inactivate Fe protein more quickly than a strain with wild-type background. Deletion of nifD, which encodes a subunit of nitrogenase MoFe protein, was found to result in a slower inactivation response. No variation was found in the reactivation responses of these strains. It is concluded that the redox state of the Fe protein contributes to the regulation of the ADP-ribosylation of Fe protein. PMID- 10850983 TI - Identification of an archaeal 2-hydroxy acid dehydrogenase catalyzing reactions involved in coenzyme biosynthesis in methanoarchaea. AB - Two putative malate dehydrogenase genes, MJ1425 and MJ0490, from Methanococcus jannaschii and one from Methanothermus fervidus were cloned and overexpressed in Escherichia coli, and their gene products were tested for the ability to catalyze pyridine nucleotide-dependent oxidation and reduction reactions of the following alpha-hydroxy-alpha-keto acid pairs: (S)-sulfolactic acid and sulfopyruvic acid; (S)-alpha-hydroxyglutaric acid and alpha-ketoglutaric acid; (S)-lactic acid and pyruvic acid; and 1-hydroxy-1,3,4,6-hexanetetracarboxylic acid and 1-oxo-1,3,4, 6 hexanetetracarboxylic acid. Each of these reactions is involved in the formation of coenzyme M, methanopterin, coenzyme F(420), and methanofuran, respectively. Both the MJ1425-encoded enzyme and the MJ0490-encoded enzyme were found to function to different degrees as malate dehydrogenases, reducing oxalacetate to (S)-malate using either NADH or NADPH as a reductant. Both enzymes were found to use either NADH or NADPH to reduce sulfopyruvate to (S)-sulfolactate, but the V(max)/K(m) value for the reduction of sulfopyruvate by NADH using the MJ1425 encoded enzyme was 20 times greater than any other combination of enzymes and pyridine nucleotides. Both the M. fervidus and the MJ1425-encoded enzyme catalyzed the NAD(+)-dependent oxidation of (S)-sulfolactate to sulfopyruvate. The MJ1425-encoded enzyme also catalyzed the NADH-dependent reduction of alpha ketoglutaric acid to (S)-hydroxyglutaric acid, a component of methanopterin. Neither of the enzymes reduced pyruvate to (S)-lactate, a component of coenzyme F(420). Only the MJ1425-encoded enzyme was found to reduce 1-oxo-1,3,4,6 hexanetetracarboxylic acid, and this reduction occurred only to a small extent and produced an isomer of 1-hydroxy-1,3,4,6-hexanetetracarboxylic acid that is not involved in the biosynthesis of methanofuran c. We conclude that the MJ1425 encoded enzyme is likely to be involved in the biosynthesis of both coenzyme M and methanopterin. PMID- 10850984 TI - Analysis of the polar flagellar gene system of Vibrio parahaemolyticus. AB - Vibrio parahaemolyticus has dual flagellar systems adapted for locomotion under different circumstances. A single, sheathed polar flagellum propels the swimmer cell in liquid environments. Numerous unsheathed lateral flagella move the swarmer cell over surfaces. The polar flagellum is produced continuously, whereas the synthesis of lateral flagella is induced under conditions that impede the function of the polar flagellum, e.g., in viscous environments or on surfaces. Thus, the organism possesses two large gene networks that orchestrate polar and lateral flagellar gene expression and assembly. In addition, the polar flagellum functions as a mechanosensor controlling lateral gene expression. In order to gain insight into the genetic circuitry controlling motility and surface sensing, we have sought to define the polar flagellar gene system. The hierarchy of regulation appears to be different from the polar system of Caulobacter crescentus or the peritrichous system of enteric bacteria but is pertinent to many Vibrio and Pseudomonas species. The gene identity and organization of 60 potential flagellar and chemotaxis genes are described. Conserved sequences are defined for two classes of polar flagellar promoters. Phenotypic and genotypic analysis of mutant strains with defects in swimming motility coupled with primer extension analysis of flagellar and chemotaxis transcription provides insight into the polar flagellar organelle, its assembly, and regulation of gene expression. PMID- 10850985 TI - Genetic and environmental factors affecting T-pilin export and T-pilus biogenesis in relation to flagellation of Agrobacterium tumefaciens. AB - The T pilus, primarily composed of cyclic T-pilin subunits, is essential for the transmission of the Ti-plasmid T-DNA from Agrobacterium tumefaciens to plant cells. Although the virB2 gene of the 11-gene virB operon was previously demonstrated to encode the full-length propilin, and other genes of this operon have been implicated as members of a conserved transmembrane transport apparatus, the role of each virB gene in T-pilin synthesis and transport and T-pilus biogenesis remained undefined. In the present study, each virB gene was examined and was found to be unessential for T-pilin biosynthesis, except virB2, but was determined to be essential for the export of the T-pilin subunits and for T-pilus formation. We also find that the genes of the virD operon are neither involved in T-pilin export nor T-pilus formation. Critical analysis of three different virD4 mutants also showed that they are not involved in T-pilus biogenesis irrespective of the A. tumefaciens strains used. With respect to the environmental effects on T-pilus biogenesis, we find that T pili are produced both on agar and in liquid culture and are produced at one end of the A. tumefaciens rod-shaped cell in a polar manner. We also report a novel phenomenon whereby flagellum production is shut down under conditions which turn on T-pilus formation. These conditions are the usual induction with acetosyringone at pH 5.5 of Ti-plasmid vir genes. A search of the vir genes involved in controlling this biphasic reaction in induced A. tumefaciens cells revealed that virA on the Ti plasmid is involved and that neither virB nor virD genes are needed for this reaction. The biphasic reaction therefore appears to be mediated through a two-component signal transducing system likely involving an unidentified vir gene in A. tumefaciens. PMID- 10850986 TI - Characterization of a Snorhizobium meliloti ATP-binding cassette histidine transporter also involved in betaine and proline uptake. AB - The symbiotic soil bacterium Sinorhizobium meliloti uses the compatible solutes glycine betaine and proline betaine for both protection against osmotic stress and, at low osmolarities, as an energy source. A PCR strategy based on conserved domains in components of the glycine betaine uptake systems from Escherichia coli (ProU) and Bacillus subtilis (OpuA and OpuC) allowed us to identify a highly homologous ATP-binding cassette (ABC) binding protein-dependent transporter in S. meliloti. This system was encoded by three genes (hutXWV) of an operon which also contained a fourth gene (hutH2) encoding a putative histidase, which is an enzyme involved in the first step of histidine catabolism. Site-directed mutagenesis of the gene encoding the periplasmic binding protein (hutX) and of the gene encoding the cytoplasmic ATPase (hutV) was done to study the substrate specificity of this transporter and its contribution in betaine uptake. These mutants showed a 50% reduction in high-affinity uptake of histidine, proline, and proline betaine and about a 30% reduction in low-affinity glycine betaine transport. When histidine was used as a nitrogen source, a 30% inhibition of growth was observed in hut mutants (hutX and hutH2). Expression analysis of the hut operon determined using a hutX-lacZ fusion revealed induction by histidine, but not by salt stress, suggesting this uptake system has a catabolic role rather than being involved in osmoprotection. To our knowledge, Hut is the first characterized histidine ABC transporter also involved in proline and betaine uptake. PMID- 10850987 TI - Autodisplay: functional display of active beta-lactamase on the surface of Escherichia coli by the AIDA-I autotransporter. AB - Members of the protein family of immunoglobulin A1 protease-like autotransporters comprise multidomain precursors consisting of a C-terminal autotransporter domain that promotes the translocation of N-terminally attached passenger domains across the cell envelopes of gram-negative bacteria. Several autotransporter domains have recently been shown to efficiently promote the export of heterologous passenger domains, opening up an effective tool for surface display of heterologous proteins. Here we report on the autotransporter domain of the Escherichia coli adhesin involved in diffuse adherence (AIDA-I), which was genetically fused to the C terminus of the periplasmic enzyme beta-lactamase, leading to efficient expression of the fusion protein in E. coli. The beta lactamase moiety of the fusion protein was presented on the bacterial surface in a stable manner, and the surface-located beta-lactamase was shown to be enzymatically active. Enzymatic activity was completely removed by protease treatment, indicating that surface display of beta-lactamase was almost quantitative. The periplasmic domain of the outer membrane protein OmpA was not affected by externally added proteases, demonstrating that the outer membranes of E. coli cells expressing the beta-lactamase AIDA-I fusion protein remained physiologically intact. PMID- 10850988 TI - Temperature-dependent function of the glutamine phosphoribosylpyrophosphate amidotransferase ammonia channel and coupling with glycinamide ribonucleotide synthetase in a hyperthermophile. AB - Genes encoding glutamine phosphoribosylpyrophosphate amidotransferase (GPAT) and glycinamide ribonucleotide synthetase (GARS) from Aquifex aeolicus were expressed in Escherichia coli, and the enzymes were purified to near homogeneity. Both enzymes were maximally active at a temperature of at least 90 degrees C, with half-lives of 65 min for GPAT and 60 h for GARS at 80 degrees C. GPAT activity is known to depend upon channeling of NH(3) from a site in an N-terminal glutaminase domain to a distal phosphoribosylpyrophosphate site in a C-terminal domain where synthesis of phosphoribosylamine (PRA) takes place. The efficiency of channeling of NH(3) for synthesis of PRA was found to increase from 34% at 37 degrees C to a maximum of 84% at 80 degrees C. The mechanism for transfer of PRA to GARS is not established, but diffusion between enzymes as a free intermediate appears unlikely based on a calculated PRA half-life of approximately 0.6 s at 90 degrees C. Evidence was obtained for coupling between GPAT and GARS for PRA transfer. The coupling was temperature dependent, exhibiting a transition between 37 and 50 degrees C, and remained relatively constant up to 90 degrees C. The calculated PRA chemical half-life, however, decreased by a factor of 20 over this temperature range. These results provide evidence that coupling involves direct PRA transfer through GPAT-GARS interaction rather than free diffusion. PMID- 10850989 TI - Role of the dpr product in oxygen tolerance in Streptococcus mutans. AB - We have previously identified and characterized the alkyl hydroperoxide reductase of Streptococcus mutans, which consists of two components, Nox-1 and AhpC. Deletion of both nox-1 and ahpC had no effect on the sensitivity of S. mutans to cumene hydroperoxide or H(2)O(2), implying that the existence of another antioxidant system(s) independent of the Nox-1-AhpC system compensates for the deficiency. Here, a new antioxidant gene (dpr for Dps-like peroxide resistance gene) was isolated from the S. mutans chromosome by its ability to complement an ahpCF deletion mutant of Escherichia coli with a tert-butyl hydroperoxide hypersensitive phenotype. The dpr gene complemented the defect in peroxidase activity caused by the deletion of nox-1 and ahpC in S. mutans. Under aerobic conditions, the dpr disruption mutant carrying a spectinomycin resistance gene (dpr::Spc(r) mutant) grew as well as wild-type S. mutans in liquid medium. However, the dpr::Spc(r) mutant could not form colonies on an agar plate under air. In addition, neither the dpr::Spc(r) ahpC::Em(r)::nox-1 triple mutant nor the dpr::Spc(r) sod::Em(r) double mutant was able to grow aerobically in liquid medium. The 20-kDa dpr gene product Dpr is an iron-binding protein. Synthesis of Dpr was induced by exposure of S. mutans cells to air. We propose a mechanism by which Dpr confers aerotolerance on S. mutans. PMID- 10850990 TI - Cross-pathway regulation in Saccharomyces cerevisiae: activation of the proline utilization pathway by Ga14p in vivo. AB - The Put3p and Gal4p transcriptional activators are members of a distinct class of fungal regulators called the Cys(6) Zn(II)(2) binuclear cluster family. This family includes over 50 different Saccharomyces cerevisiae proteins that share a similar domain organization. Gal4p activates the genes of the galactose utilization pathway permitting the use of galactose as the sole source of carbon and energy. Put3p controls the expression of the proline utilization pathway that allows yeast cells to grow on proline as the sole nitrogen source. We report that Gal4p can activate the PUT structural genes in a strain lacking Put3p. We also show that the activation of PUT2 by Gal4p depends on the presence of the inducer galactose and the Put3p binding site and that activation increases with increased dosage of Gal4p. Put3p cannot activate the GAL genes in the absence of Gal4p. Our in vivo results confirm previously published in vitro data showing that Gal4p is more promiscuous than Put3p in its DNA binding ability. The results also suggest that under appropriate circumstances, Gal4p may be able to function in place of a related family member to activate expression. PMID- 10850991 TI - Marinomonas mediterranea MMB-1 transposon mutagenesis: isolation of a multipotent polyphenol oxidase mutant. AB - Marinomonas mediterranea is a melanogenic marine bacterium expressing a multifunctional polyphenol oxidase (PPO) able to oxidize substrates characteristic for laccases and tyrosinases, as well as produce a classical tyrosinase. A new and quick method has been developed for screening laccase activity in culture plates to detect mutants differentially affected in this PPO activity. Transposon mutagenesis has been applied for the first time to M. mediterranea by using different minitransposons loaded in R6K-based suicide delivery vectors mobilizable by conjugation. Higher frequencies of insertions were obtained by using mini-Tn10 derivatives encoding kanamycin or gentamycin resistance. After applying this protocol, a multifunctional PPO-negative mutant was obtained. By using the antibiotic resistance cassette as a marker, flanking regions were cloned. Then the wild-type gene was amplified by PCR and was cloned and sequenced. This is the first report on cloning and sequencing of a gene encoding a prokaryotic enzyme with laccase activity. The deduced amino acid sequence shows the characteristic copper-binding sites of other blue copper proteins, including fungal laccases. In addition, it shows some extra copper binding sites that might be related to its multipotent enzymatic capability. PMID- 10850992 TI - Cloning, expression, and purification of the K5 capsular polysaccharide lyase (KflA) from coliphage K5A: evidence for two distinct K5 lyase enzymes. AB - The Escherichia coli K5 capsular polysaccharide [-4)-betaGlcA-(1, 4)-alphaGlcNAc (1-] is a receptor for the capsule-specific bacteriophage K5A. Associated with the structure of bacteriophage K5A is a polysaccharide lyase which degrades the K5 capsule to expose the underlying bacterial cell surface. The bacteriophage K5A lyase gene (kflA) was cloned and sequenced. The kflA gene encodes a polypeptide with a predicted molecular mass of 66.9 kDa and which exhibits amino acid homology with ElmA, a K5 polysaccharide lyase encoded on the chromosome of E. coli SEBR 3282. There was only limited nucleotide homology between the kflA and elmA genes, suggesting that these two genes are distinct and either have been derived from separate progenitors or have diverged from a common progenitor for a considerable length of time. Southern blot analysis revealed that kflA was not present on the chromosome of the E. coli strains examined. In contrast, elmA was present in a subset of E. coli strains. Homology was observed between DNA flanking the kflA gene of bacteriophage K5A and DNA flanking a small open reading frame (ORF(L)) located 5' of the endosialidase gene of the E. coli K1 capsule specific bacteriophage K1E. The DNA homology between these noncoding sequences indicated that bacteriophages K5A and K1E were related. The deduced polypeptide sequence of ORF(L) in bacteriophage K1E exhibited homology to the N terminus of KflA from bacteriophage K5A, suggesting that ORF(L) is a truncated remnant of KflA. The presence of this truncated kflA gene implies that bacteriophage K1E has evolved from bacteriophage K5A by acquisition of the endosialidase gene and subsequent loss of functional kflA. A (His)(6)-KflA fusion protein was overexpressed in E. coli and purified to homogeneity with a yield of 4.8 mg per liter of bacterial culture. The recombinant enzyme was active over a broad pH range and NaCl concentration and was capable of degrading K5 polysaccharide into a low-molecular-weight product. PMID- 10850993 TI - Regulation of the furA and catC operon, encoding a ferric uptake regulator homologue and catalase-peroxidase, respectively, in Streptomyces coelicolor A3(2). AB - We isolated the catC gene, encoding catalase-peroxidase in Streptomyces coelicolor, using sequence homology with the katG gene from Escherichia coli. Upstream of the catC gene, an open reading frame (furA) encoding a homologue of ferric uptake regulator (Fur) was identified. S1 mapping analysis indicated that the furA gene was cotranscribed with the catC gene. The transcriptional start site of the furA-catC mRNA was mapped to the translation start codon ATG of the furA gene. The putative promoter contains consensus -10 and -35 elements similar to those recognized by sigma(HrdB), the major sigma factor of S. coelicolor. The transcripts were produced maximally at late-exponential phase and decreased at the stationary phase in liquid culture. The change in the amount of mRNA was consistent with that of CatC protein and enzyme activity. When the furA gene was introduced into S. lividans on a multicopy plasmid, the increased production of catC transcripts and protein product at late growth phase was inhibited, implying a role for FurA as the negative regulator of the furA-catC operon. FurA protein bound to its own promoter region between -59 and -39 nucleotides from the transcription start site. The binding affinity of FurA increased under reducing conditions and in the presence of metals such as Ni(2+), Mn(2+), Zn(2+), or Fe(2+). Addition of these metals to the growth medium decreased the production of CatC protein, consistent with the role of FurA as a metal-dependent repressor. PMID- 10850994 TI - Characterization of the ends and target sites of the novel conjugative transposon Tn5397 from Clostridium difficile: excision and circularization is mediated by the large resolvase, TndX. AB - Tn5397 is a conjugative transposon that was originally isolated from Clostridium difficile. Previous analysis had shown that the central region of Tn5397 was closely related to the conjugative transposon Tn916. However, in this work we obtained the DNA sequence of the ends of Tn5397 and showed that they are completely different to those of Tn916. Tn5397 did not contain the int and xis genes, which are required for the excision and integration of Tn916. Instead, the right end of Tn5397 contained a gene, tndX, that appears to encode a member of the large resolvase family of site-specific recombinases. TndX is closely related to the TnpX resolvase from the mobilizable but nonconjugative chloramphenicol resistance transposons, Tn4451 from Clostridium perfringens and Tn4453 from C. difficile. Like the latter elements, inserted copies of Tn5397 were flanked by a direct repeat of a GA dinucleotide. The Tn5397 target sites were also shown to contain a central GA dinucleotide. Excision of the element in C. difficile completely regenerated the original target sequence. A circular form of the transposon, in which the left and right ends of the element were separated by a GA dinucleotide, was detected by PCR in both Bacillus subtilis and C. difficile. A Tn5397 mutant in which part of tndX was deleted was constructed in B. subtilis. This mutant was nonconjugative and did not produce the circular form of Tn5397, indicating that the TndX resolvase has an essential role in the excision and transposition of Tn5397 and is thus the first example of a member of the large resolvase family of recombinases being involved in conjugative transposon mobility. Finally, we showed that introduction of Tn916 into a strain containing Tn5397 induced the loss of the latter element in 95.6% of recipients. PMID- 10850995 TI - Genetic investigation of the catabolic pathway for degradation of abietane diterpenoids by Pseudomonas abietaniphila BKME-9. AB - We have cloned and sequenced the dit gene cluster encoding enzymes of the catabolic pathway for abietane diterpenoid degradation by Pseudomonas abietaniphila BKME-9. The dit gene cluster is located on a 16.7-kb DNA fragment containing 13 complete open reading frames (ORFs) and 1 partial ORF. The genes ditA1A2A3 encode the alpha and beta subunits and the ferredoxin of the dioxygenase which hydroxylates 7-oxodehydroabietic acid to 7-oxo-11,12-dihydroxy 8, 13-abietadien acid. The dioxygenase mutant strain BKME-941 (ditA1::Tn5) did not grow on nonaromatic abietanes, and transformed palustric and abietic acids to 7-oxodehydroabietic acid in cell suspension assays. Thus, nonaromatic abietanes are aromatized prior to further degradation. Catechol 2,3-dioxygenase activity of xylE transcriptional fusion strains showed induction of ditA1 and ditA3 by abietic, dehydroabietic, and 7-oxodehydroabietic acids, which support the growth of strain BKME-9, as well as by isopimaric and 12, 14-dichlorodehydroabietic acids, which are diterpenoids that do not support the growth of strain BKME-9. In addition to the aromatic-ring-hydroxylating dioxygenase genes, the dit cluster includes ditC, encoding an extradiol ring cleavage dioxygenase, and ditR, encoding an IclR-type transcriptional regulator. Although ditR is not strictly required for the growth of strain BKME-9 on abietanes, a ditR::Km(r) mutation in a ditA3::xylE reporter strain demonstrated that it encodes an inducer-dependent transcriptional activator of ditA3. An ORF with sequence similarity to genes encoding permeases (ditE) is linked with genes involved in abietane degradation. PMID- 10850996 TI - Defining a rob regulon in Escherichia coli by using transposon mutagenesis. AB - The Rob protein of Escherichia coli is a member of the AraC-XylS family of prokaryotic transcriptional regulators and is expressed constitutively. Deletion of the rob gene increases susceptibility to organic solvents, while overexpression of Rob increases tolerance to organic solvents and resistance to a variety of antibiotics and to the superoxide-generating compound phenazine methosulfate. To determine whether constitutive levels of Rob regulate basal gene expression, we performed a MudJ transposon screen in a rob deletion mutant containing a plasmid that allows for controlled rob gene expression. We identified eight genes and confirmed that seven are transcriptionally activated by normal expression of Rob from the chromosomal rob gene (inaA, marR, aslB, ybaO, mdlA, yfhD, and ybiS). One gene, galT, was repressed by Rob. We also demonstrated by Northern analysis that basal expression of micF is significantly higher in wild-type E. coli than in a rob deletion mutant. Rob binding to the promoter regions of most of these genes was substantiated in electrophoretic mobility shift assays. However, Mu insertions in individual Rob-regulated genes did not affect solvent sensitivity. This phenotype may depend on changes in the expression of several of these Rob-regulated genes or on other genes that were not identified. Rob clearly affects the basal expression of genes with a broad range of functions, including antibiotic resistance, acid adaptation, carbon metabolism, cell wall synthesis, central intermediary metabolism, and transport. The magnitudes of Rob's effects are modest, however, and the protein may thus play a role as a general transcription cofactor. PMID- 10850997 TI - Fur positive regulation of iron superoxide dismutase in Escherichia coli: functional analysis of the sodB promoter. AB - In Escherichia coli, the expression of sodB, which encodes iron superoxide dismutase, has been suggested to be activated by Fur, the iron-responsive global regulator initially characterized as a transcriptional repressor. We investigated sodB regulation by functional analysis of the sodB promoter using sodB-lac fusions with various truncated and mutated promoters. Several cis- and trans acting elements involved in sodB regulation have been identified. The beta galactosidase activity of sodB-lacZ reporter fusions and RNA analysis showed sevenfold iron-dependent, Fur-mediated activation of expression. A region just downstream from -10, including a large palindromic sequence encompassing the +1 position followed by a 14-bp AT-rich motif, is the site of Fur positive regulation, and the integrity of both sequences was required for full Fur mediated activation. The life span of sodB mRNA was three times longer in a fur(+) strain, indicating that Fur-mediated activation proceeds, at least in part, at the posttranscriptional level. The H-NS and IHF histone-like factors also affected sodB expression. IHF slightly repressed sodB expression independently of Fur regulation. In contrast, H-NS negative regulation operated only in the absence of Fur. Remarkably, psodB behaved like a "pure extended -10" promoter. Deletion of the -35 region did not affect expression, whereas expression was totally abolished by a TG-to-CC mutation in the extended -10 sequence TGcTACCCT. PMID- 10850998 TI - Physical morphology and surface properties of unsaturated Pseudomonas putida biofilms. AB - Unsaturated biofilms of Pseudomonas putida, i.e., biofilms grown in humid air, were analyzed by atomic force microscopy to determine surface morphology, roughness, and adhesion forces in the outer and basal cell layers of fresh and desiccated biofilms. Desiccated biofilms were equilibrated with a 75.5% relative humidity atmosphere, which is far below the relative humidity of 98 to 99% at which these biofilms were cultured. In sharp contrast to the effects of drying on biofilms grown in fluid, we observed that drying caused little change in morphology, roughness, or adhesion forces in these unsaturated biofilms. Surface roughness for moist and dry biofilms increased approximately linearly with increasing scan sizes. This indicated that the divides between bacteria contributed more to overall roughness than did extracellular polymeric substances (EPS) on individual bacteria. The EPS formed higher-order structures we termed mesostructures. These mesostructures are much larger than the discrete polymers of glycolipids and proteins that have been previously characterized on the outer surface of these gram-negative bacteria. PMID- 10850999 TI - Pheromone-regulated expression of sex pheromone plasmid pAD1-encoded aggregation substance depends on at least six upstream genes and a cis-acting, orientation dependent factor. AB - Conjugative transfer of Enterococcus faecalis-specific sex pheromone plasmids relies on an adhesin, called aggregation substance, to confer a tight cell-to cell contact between the mating partners. To analyze the dependence of pAD1 encoded aggregation substance, Asa1, on pheromone induction, a variety of upstream fragments were fused to an alpha-amylase reporter gene, amyL, by use of a novel promoter probe vector, pAMY-em1. For pheromone-regulated alpha-amylase activity, a total of at least six genes, traB, traC, traA, traE1, orfY, and orf1, are required: TraB efficiently represses asa1 (by a mechanism unrelated to its presumptive function in pheromone shutdown, since a complete shutdown is observed exclusively in the presence of traC); only traC can relieve traB-mediated repression in a pheromone-dependent manner. In addition to traB, traA is required but not sufficient for negative control. Mutational inactivation of traE1, orfY, or orf1, respectively, results in a total loss of alpha-amylase activity for constructs normally mediating constitutive expression. Inversion of a fragment covering traA, P(0), and traE1 without disrupting any gene or control element switches off amyL or asa1 expression, indicating the involvement of a cis-acting, orientation-dependent factor (as had been shown for plasmid pCF10). Unexpectedly, pAD1 represses all pAMY-em1 derivatives in trans, while its own pheromone dependent functions are unaffected. The discrepancy between the new data and those of former studies defining TraE1 as a trans-acting positive regulator is discussed. PMID- 10851000 TI - Secretion of nucleoside diphosphate kinase by mucoid Pseudomonas aeruginosa 8821: involvement of a carboxy-terminal motif in secretion. AB - Nucleoside diphosphate kinase (Ndk) is a ubiquitous enzyme which functions in balancing the nucleotide pool of the cell. We have recently reported that in addition to being intracellular in both mucoid and nonmucoid Pseudomonas aeruginosa, Ndk is also secreted into the extracellular environment by mucoid P. aeruginosa cells. This secreted Ndk has biochemical activity similar to the intracellular Ndk and is 16 kDa in size. To demonstrate that Ndk is indeed secreted and to localize the secretion motif, we constructed an ndk knockout mutant, which lacks both intracellular and extracellular forms of Ndk. In this study, we report the construction of deletion derivatives made from the carboxy terminal region of Ndk. These deletion derivatives were introduced into the ndk::Cm knockout mutant and were examined for the intracellular and extracellular presence of Ndk. It was observed that the carboxy-terminal 8-amino-acid region is required for the secretion of Ndk into the extracellular region. This region has the sequence DXXX, where X is a predominantly hydrophobic residue. Such sequences represent a conserved motif in proteins secreted by the type I secretory pathway in gram-negative microorganisms. To investigate the significance of this motif in the secretion of Ndk, we constructed a fusion protein of Ndk and the blue fluorescent protein (BFP) as well as a fusion protein of mutated Ndk (whose DTEV motif has been changed to AAAA) and the BFP. The presence of extracellular Ndk was detected only in the ndk::Cm knockout mutant harboring the wild-type BFP-Ndk protein fusion. We could not detect the presence of extracellular Ndk in the ndk::Cm knockout mutant containing the mutated BFP-Ndk protein fusion. In addition, we have also used immunofluorescence microscopy to localize the wild type and mutated BFP-Ndk proteins in the cell. The significance of these observations is discussed. PMID- 10851001 TI - An essential two-component signal transduction system in Mycobacterium tuberculosis. AB - The bacterial two-component signal transduction systems regulate adaptation processes and are likely to play a role in Mycobacterium tuberculosis physiology and pathogenesis. The previous initial characterization of an M. tuberculosis response regulator from one of these systems, mtrA-mtrB, suggested its transcriptional activation during infection of phagocytic cells. In this work, we further characterized the mtrA response regulator from M. tuberculosis H37Rv. Inactivation of mtrA on the chromosome of M. tuberculosis H37Rv was possible only in the presence of plasmid-borne functional mtrA, suggesting that this response regulator is essential for M. tuberculosis viability. In keeping with these findings, expression of mtrA in M. tuberculosis H37Rv was detectable during in vitro growth, as determined by S1 nuclease protection and primer extension analyses of mRNA levels and mapping of transcript 5' ends. The mtrA gene was expressed differently in virulent M. tuberculosis and the vaccine strain M. tuberculosis var. bovis BCG during infection of macrophages, as determined by monitoring of mtrA-gfp fusion activity. In M. bovis BCG, mtrA was induced upon entry into macrophages. In M. tuberculosis H37Rv, its expression was constitutive and unchanged upon infection of murine or human monocyte-derived macrophages. In conclusion, these results identify mtrA as an essential response regulator gene in M. tuberculosis which is differentially expressed in virulent and avirulent strains during growth in macrophages. PMID- 10851002 TI - Synechocystis strain PCC 6803 cya2, a prokaryotic gene that encodes a guanylyl cyclase. AB - Synechocystis strain PCC 6803 exhibits similar levels of cyclic AMP (cAMP) and cyclic GMP (cGMP). A thorough analysis of its genome showed that Cya2 (Sll0646) has all the sequence determinants required in terms of activity and purine specificity for being a guanylyl cyclase. Insertional mutagenesis of cya2 caused a marked reduction in cGMP content without altering the cAMP content. Thus, Cya2 represents the first example of a prokaryotic guanylyl cyclase. PMID- 10851003 TI - Positive correlation between virulence of Pseudomonas aeruginosa mutants in mice and insects. AB - Strain PA14, a human clinical isolate of Pseudomonas aeruginosa, is pathogenic in mice and insects (Galleria mellonella). Analysis of 32 different PA14 mutants in these two hosts showed a novel positive correlation in the virulence patterns. Thus, G. mellonella is a good model system for identifying mammalian virulence factors of P. aeruginosa. PMID- 10851004 TI - Mutations in oxyR resulting in peroxide resistance in Xanthomonas campestris. AB - A spontaneous Xanthomonas campestris pv. phaseoli H(2)O(2)-resistant mutant emerged upon selection with 1 mM H(2)O(2). In this report, we show that growth of this mutant under noninducing conditions gave high levels of catalase, alkyl hydroperoxide reductase (AhpC and AhpF), and OxyR. The H(2)O(2) resistance phenotype was abolished in oxyR-minus derivatives of the mutant, suggesting that elevated levels and mutations in oxyR were responsible for the phenotype. Nucleotide sequence analysis of the oxyR mutant showed three nucleotide changes. These changes resulted in one silent mutation and two amino acid changes, one at a highly conserved location (G197 to D197) and the other at a nonconserved location (L301 to R301) in OxyR. Furthermore, these mutations in oxyR affected expression of genes in the oxyR regulon. Expression of an oxyR-regulated gene, ahpC, was used to monitor the redox state of OxyR. In the parental strain, a high level of wild-type OxyR repressed ahpC expression. By contrast, expression of oxyR5 from the X. campestris pv. phaseoli H(2)O(2)-resistant mutant and its derivative oxyR5G197D with a single-amino-acid change on expression vectors activated ahpC expression in the absence of inducer. The other single-amino-acid mutant derivative of oxyR5L301R had effects on ahpC expression similar to those of the wild-type oxyR. However, when the two single mutations were combined, as in oxyR5, these mutations had an additive effect on activation of ahpC expression. PMID- 10851005 TI - Isolation and characterization of canthaxanthin biosynthesis genes from the photosynthetic bacterium Bradyrhizobium sp. strain ORS278. AB - A carotenoid biosynthesis gene cluster involved in canthaxanthin production was isolated from the photosynthetic Bradyrhizobium sp. strain ORS278. This cluster includes five genes identified as crtE, crtY, crtI, crtB, and crtW that are organized in at least two operons. The functional assignment of each open reading frame was confirmed by complementation studies. PMID- 10851006 TI - Role of the Azotobacter vinelandii nitrogenase-protective shethna protein in preventing oxygen-mediated cell death. AB - Azotobacter vinelandii strains lacking the nitrogenase-protective Shethna protein lost viability upon carbon-substrate deprivation in the presence of oxygen. This viability loss was dependent upon the N(2)-fixing status of cultures (N(2)-fixing cells lost viability, while non-N(2)-fixing cells did not) and on the ambient O(2) level. Supra-atmosheric O(2) tensions (40% partial pressure) decreased the viable cell number of the mutant further, and the mutant had a slightly higher spontaneous mutation frequency than the wild type in the high-O(2) conditions. Iron starvation conditions, which resulted in fourfold-reduced superoxide dismutase levels, were also highly detrimental to the viability of the protective protein mutants, but these conditions did not affect the viability of the wild type strain. Nitrogenase or other powerful reductants associated with N(2) fixation may be sources of damaging partially reduced oxygen species, and the production of such species are perhaps minimized by the Shethna protein. PMID- 10851007 TI - Phosphorelay as the sole physiological route of signal transmission by the arc two-component system of Escherichia coli. AB - The Arc two-component system, comprising a tripartite sensor kinase (ArcB) and a response regulator (ArcA), modulates the expression of numerous genes involved in respiratory functions. In this study, the steps of phosphoryl group transfer from phosphorylated ArcB to ArcA were examined in vivo by using single copies of wild type and mutant arcB alleles. The results indicate that the signal transmission occurs solely by His-Asp-His-Asp phosphorelay. PMID- 10851008 TI - Enterococcus faecalis V583 contains a cytochrome bd-type respiratory oxidase. AB - We have cloned an Enterococcus faecalis gene cluster, cydABCD, which when expressed in Bacillus subtilis results in a functional cytochrome bd terminal oxidase. Our results indicate that E. faecalis V583 cells have the capacity of aerobic respiration when grown in the presence of heme. PMID- 10851009 TI - Decoupling of genome size and sequence divergence in a symbiotic bacterium. AB - In contrast to genome size variation in most bacterial taxa, the small genome size of Buchnera sp. was shown to be highly conserved across genetically diverse isolates (630 to 643 kb). This exceptional size conservation may reflect the inability of this obligate mutualist to acquire foreign DNA and reduced selection for genetic novelty within a static intracellular environment. PMID- 10851010 TI - The FtsH protein accumulates at the septum of Bacillus subtilis during cell division and sporulation. AB - The ftsH gene encodes an ATP- and Zn(2+)-dependent metalloprotease which is anchored to the cytoplasmic membrane via two transmembrane segments in such a way that the very short amino- and the long carboxy termini are exposed to the cytoplasm. Deletion of the ftsH gene in Bacillus subtilis results in a pleiotropic phenotype such as filamentous growth. This observation prompted us to ask whether ftsH is involved in cell division. A translational fusion was constructed between the complete coding region of ftsH and gfp(+) the latter carrying five point mutations to obtain enhanced fluorescence. We detected that the FtsH protein accumulates in the midcell septum of dividing cells, and during sporulation first in the asymmetrically located septa of sporulating cells and later in the membrane which engulfs the forespore. These observations revealed a new function of FtsH. PMID- 10851011 TI - Phosphorylated PmrA interacts with the promoter region of ugd in Salmonella enterica serovar typhimurium. AB - The Salmonella PmrA-PmrB system controls the expression of genes necessary for polymyxin B resistance. Four loci were previously identified as part of the regulon, and interaction of PmrA with the promoter region of three of them was observed. Here we characterized the interaction of PmrA with the promoter region of ugd, previously suggested to be regulated indirectly by PmrA. Our results indicate that PmrA controls the expression of ugd by interacting with a specific sequence in the promoter region of this gene. PMID- 10851012 TI - Organization and cell-cell interaction in starved Saccharomyces cerevisiae colonies. AB - Cell growth in yeast colonies is a complex process, the control of which is largely unknown. Here we present scanning electron micrographs of Saccharomyces cerevisiae colonies, showing changes in the pattern of cell organization and cell cell interactions during colony development. In young colonies ( 6.0, the patterns of activation and inhibition changed to mixed-type during the peroxidase oxidation of thioproperazine and triftazine and to competitive inhibition of peroxidase with strophanthin G during the oxidation of aminazine. These effects are suggested to be due to an ionizable enzyme group of pK approximately 6.0. Strophanthin G inhibited free-radical oxidation of o-dianisidine via binding to the enzyme-substrate complex, preventing the generation of a stable semi-oxidized product of o-dianisidine, and thus inhibiting the enzyme by the anticompetitive mechanism. Mechanisms of oxidation of slowly and rapidly oxidizable substrates of peroxidase in the presence of strophanthin G are suggested. PMID- 10851033 TI - Cloning and expression of a new site-specific methyltransferase M.SscL1I from Staphylococcus sp. L1. AB - The gene of the new site-specific methyltransferase M.SscL1I belonging to the same modification-restriction system as the previously described by us site specific endonuclease SscL1I has been cloned from the natural strain Staphylococcus sp. L1. A plasmid to express the methylase gene under control of the T7 phage-specific promotor has been constructed. Conditions were found to express the recombinant methylase M.SscL1I and to purify it to near homogeneity. It is shown that the methylase modifies the adenine base in the recognition site 5;-GANTC-3;. PMID- 10851034 TI - Structural analysis of the coat protein of cucumber green mottle mosaic virus. AB - Using reversed-phase high-performance liquid chromatography, two components of the coat protein of isolate No. 3 of the cucumber green mottle mosaic virus (CGMMV, cucumber strain), Cp1 (minor) and Cp2 (major), were isolated and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). In the Cp2 mass spectrum, two polypeptides with Mr of 16,727.0 and 16,813.5 were detected. By Edman degradation in combination with mass spectrometry, the primary structure of the tryptic peptides of Cp2 comprising in total 150 amino acid residues was determined. Two amino acid substitutions, Val-56-->Ala-56 and Asp-64-->Ser-64, were revealed in Cp2, as compared to the watermelon strain of the virus. Cp1 was shown to consist of three polypeptides with Mr of 10,014.2, 10,224.9, and 10,355.9 corresponding to the N terminal regions of Cp2 (positions 1-92, 1-94, and 1-95). The observed heterogeneity of the coat protein of CGMMV, cucumber strain, may be due to proteolysis during protein isolation. PMID- 10851035 TI - A new inhibitor of the CoQ-dependent redox reactions in mitochondria and chromatophores. AB - The effects of 3,4-dimethoxyphenyl-1-amylketone (DPK) on the CoQ-dependent stages of the electron transport systems in mitochondria and Rhodobacter sphaeroides chromatophores were studied. The two systems contain the complete Q-cycle. The sensitivities of the Q-cycles of two electron transport systems to antimycin, myxothiazole, and other inhibitors are virtually indistinguishable from one another, but these systems have different CoQ reduction processes. The dependence of the inhibition extent of the mitochondrial succinate oxidase on the DPK concentration was studied. The effective concentration of DPK is 0.5-2.5 mM. The presence of the point of inflection in the titration curve indicates that there are two mechanisms of inhibition. The effects of DPK on the extent of reduction of cytochromes b and c1 + c in mitochondria as well as on the electrogenic stages of the Q-cycle in chromatophores were examined. The experiments showed that DPK prevents three CoQ-dependent reactions related to the Q-cycle: electron transport between succinate dehydrogenase and the Q-cycle in mitochondria and functioning of the Z (o) and C (i) sites of the Q-cycle in chromatophores. DPK does not affect the electrogenic reaction associated with protonation of the secondary quinone acceptor QB in the reaction center of chromatophores. The mitochondrial NADH-dehydrogenase is inhibited by DPK at lower but comparable concentrations (C50 = 0.2 mM). PMID- 10851036 TI - Catalytic properties of tryptophanless recombinant horseradish peroxidase. AB - Heme-containing plant peroxidases (EC 1.11.1.7) contain a highly conserved single tryptophan residue. Its replacement with Phe in recombinant horseradish peroxidase (rHRP) increased the stability of the mutant enzyme in acid media. The kinetic properties of native, wild-type, and W117F mutant recombinant horseradish peroxidase in the reactions of ammonium 2, 2;-azino-bis(3-ethylbenzthiazoline-6 sulfonate) (ABTS), guaiacol, and o-phenylenediamine oxidation are very similar. However, significant changes in the reaction rate constant characteristic for the monomolecular rate-limiting step ascribed either to product dissociation from its complex with the enzyme or electron transfer from the substrate to the active site within the Michaelis complex were observed. The data indirectly indicate the participation of the single Trp residue in oxidation of ABTS and guaiacol and possible differences in kinetic mechanisms for oxidation of ABTS, guaiacol, and o phenylenediamine. PMID- 10851037 TI - The natural histidine-containing dipeptide Nalpha-acetylcarnosine as an antioxidant for ophthalmic use. AB - The naturally occurring compound Nalpha-acetylcarnosine is proposed as a prodrug of L-carnosine that is resistant to enzymatic hydrolysis by carnosinase. Eyes of rabbits were treated with 1% Nalpha-acetylcarnosine, L-carnosine, or placebo and extracts of the aqueous humor from the anterior eye chamber were analyzed for imidazole content by reverse-phase analytical high performance liquid chromatography (HPLC) and thin-layer (TLC) and ion-exchange chromatographic techniques. Topical administration of pure L-carnosine to the rabbit eye did not lead to accumulation of this compound in the aqueous humor over 30 min in concentration exceeding that in the placebo-treated matched eye. Nalpha Acetylcarnosine showed dose-dependent hydrolysis in its passage from the cornea to the aqueous humor, releasing L-carnosine after l5-30 min of ocular administration of the prodrug in a series of therapeutic modalities: instillation < or = subconjunctival injection < or = ultrasound-induced phoresis. Different treatment techniques showed excellent toleration of 1%Nalpha-acetylcarnosine by the eye. Once in the aqueous humor, L-carnosine might act as an antioxidant and enter the lens tissue when present at effective concentrations (5-l5 mM). The advantage of the ophthalmic prodrug Nalpha-acetylcarnosine and its bioactivated principle L-carnosine as universal antioxidants relates to their ability to give efficient protection against oxidative stress both in the lipid phase of biological membranes and in aqueous environments. Nalpha-Acetylcarnosine is proposed for treatment of ocular disorders that have a component of oxidative stress in their genesis (cataracts, glaucoma, retinal degeneration, corneal disorders, ocular inflammation, complications of diabetes mellitus, and systemic diseases). PMID- 10851038 TI - Superoxide dismutase: determination of activity by inhibition of photosensitized chemiluminescence of glycyltryptophan. AB - A technique for quantitative determination of superoxide dismutase (SOD) in biological material is described. The technique is based on the riboflavin photosensitized oxidation of Gly-Trp, which is accompanied by chemiluminescence. Formation of the luminescent product is inhibited by SOD. The dependence of SOD activity on reciprocal intensity of chemiluminescence is linear. The concentration for 50% inhibition is 7.5 ng/ml, and the minimum reliably determined concentration is about 2 ng/ml. The reaction is not sensitive to high concentrations of cyanide, which allows the separate determination of Cu,Zn- and Mn-SOD in mixtures. PMID- 10851039 TI - Constitutive biosynthesis and localization of alcohol oxidase in the ethanol insensitive catabolite repression mutant ecr1 of the yeast Pichia methanolica. AB - The activity and localization of alcohol oxidase (EC 1.1.3.13) have been studied in the Pichia methanolica mutant ecr1 defective in ethanol-induced catabolite repression of enzymes of methanol utilization. Ultrasctuctural, immunocytochemical, and biochemical analyses revealed the presence of peroxisomes containing active alcohol oxidase in the mutant grown in media with methanol, ethanol, and a mixture of both substrates. No alcohol oxidase was detected in the wild-type cells (ECR1) grown on ethanol-containing media. Mutant ecr1 growing in medium containing a mixture of different alcohols and the wild-type strain growing on methanol demonstrated similar buoyant density of peroxisomes (1.24 1.27 g/cm3)during isopicnic centrifugation of the organelles in sucrose density gradients. The integrated genetic, immunocytochemical, and biochemical data are in agreement with the model that synthesis, translocation into peroxisomes, and assembly of alcohol oxidase in P. methanolica may not require any regulatory signals induced by methanol. PMID- 10851040 TI - Thermostable DNA polymerase from Thermus thermophilus B35: influence of divalent metal ions on the interaction with deoxynucleoside triphosphates. AB - The interaction of DNA polymerase from Thermus thermophilus B35 (Tte-pol) with deoxynucleoside triphosphates in the presence of different divalent metal ions has been studied. DNA synthesis and competitive inhibition of the polymerase reaction by non-complementary dNTPs are described with corresponding kinetic schemes. The co-factor properties of some metals (Mg2+, Mn2+, Co2+, Ni2+, Cu2+, Ca2+, Cd2+, and Zn2+) were investigated, and their activating concentration ranges were determined. It was found that kcat values are significantly decreased and Km values slowly decrease when Mn2+ displaces Mg2+. The value of Kd for DNA template-primer is Me2+-independent, whereas Kd values for non-complementary dNTPs decrease in the presence of Mn2+. Tte-pol processivity but not DNA synthesis efficiency is Me2+-type independent. PMID- 10851041 TI - Hydrolysis of anandamide and eicosapentaenoic acid ethanolamide in mouse splenocytes. AB - The hydrolysis of anandamide has been studied in mouse splenocytes using tritiated anandamide analogs labeled in the acyl- or ethanolamide parts of the molecule. [3H]Anandamide undergoes rapid (t(1/2) = 2.5 min) uptake and hydrolysis, yielding ethanolamine and arachidonic acid. The anandamide hydrolysis in splenocytes is sensitive to inhibition by phenylmethylsulfonyl fluoride, and it is assumed that the observed activity is due to fatty acid amide hydrolase, which inactivates anandamide in central and peripheral tissues. Eicosapentaenoic acid ethanolamide and the 15-hydroxy-derivative of anandamide are shown to be amidohydrolase substrates as well. The fatty acids derived from hydrolytic cleavage of acylethanolamines undergo rapid oxidation by splenocyte lipoxygenase, yielding the corresponding 12-hydroxy-derivatives. Oxygenated ethanolamide derivatives were not found. The data suggest that polyenoic fatty acid ethanolamides are metabolic precursors of eicosanoids in splenocytes and that amide bond hydrolysis is the key point in switching of biological activity spectra between endocannabinoids and oxylipins. PMID- 10851042 TI - Fumaric acid is a competitive inhibitor of wheat germ lipoxygenase. AB - Fumaric acid is shown to be a competitive inhibitor of wheat germ lipoxygenase. PMID- 10851043 TI - Effect of GABAergic compounds on the anion-sensitive Mg2+-ATPase from bream (Abramis brama L.) brain. AB - Preincubation of plasma membranes from bream brain with 10-8-10-4 M gamma aminobutyric acid (GABA) or muscimol increased the anion-sensitive Mg2+-ATPase activity. The activating effect of neurotransmitters on the Mg2+-ATPase is enhanced with increasing preincubation time of the membranes with the ligands, decreases with increasing Mg2+-ATP concentration in the incubation medium, and is inhibited in the presence of the GABAa-receptor antagonist, bicuculline (90 microgr;M). The anions Cl-, Br-, and I- stimulate the basal Mg2+-ATPase activity, and an effect of 10-4 M GABA in the presence of anions was not found. It is supposed that GABAergic chemicals modify the anion-sensitive Mg2+-ATPase in a receptor-dependent way. PMID- 10851044 TI - Preface: STAT signaling. PMID- 10851045 TI - The role of STATs in transcriptional control and their impact on cellular function. AB - The STAT proteins (Signal Transducers and Activators of Transcription), were identified in the last decade as transcription factors which were critical in mediating virtually all cytokine driven signaling. These proteins are latent in the cytoplasm and become activated through tyrosine phosphorylation which typically occurs through cytokine receptor associated kinases (JAKs) or growth factor receptor tyrosine kinases. Recently a number of non-receptor tyrosine kinases (for example src and abl) have been found to cause STAT phosphorylation. Phosphorylated STATs form homo- or hetero-dimers, enter the nucleus and working coordinately with other transcriptional co-activators or transcription factors lead to increased transcriptional initiation. In normal cells and in animals, ligand dependent activation of the STATs is a transient process, lasting for several minutes to several hours. In contrast, in many cancerous cell lines and tumors, where growth factor dysregulation is frequently at the heart of cellular transformation, the STAT proteins (in particular Stats 1, 3 and 5) are persistently tyrosine phosphorylated or activated. The importance of STAT activation to growth control in experiments using anti-sense molecules or dominant negative STAT protein encoding constructs performed in cell lines or studies in animals lacking specific STATs strongly indicate that STATs play an important role in controlling cell cycle progression and apoptosis. Stat1 plays an important role in growth arrest, in promoting apoptosis and is implicated as a tumor suppressor; while Stats 3 and 5 are involved in promoting cell cycle progression and cellular transformation and preventing apoptosis. Many questions remain including: (1) a better understanding of how the STAT proteins through association with other factors increase transcription initiation; (2) a more complete definition of the sets of genes which are activated by different STATs and (3) how these sets of activated genes differ as a function of cell type. Finally, in the context of many cancers, where STATs are frequently persistently activated, an understanding of the mechanisms leading to their constitutive activation and defining the potential importance of persistent STAT activation in human tumorigenesis remains. Oncogene (2000). PMID- 10851046 TI - STATs in oncogenesis. AB - Since their discovery as key mediators of cytokine signaling, considerable progress has been made in defining the structure-function relationships of Signal Transducers and Activators of Transcription (STATs). In addition to their central roles in normal cell signaling, recent studies have demonstrated that diverse oncoproteins can activate specific STATs (particularly Stat3 and Stat5) and that constitutively-activated STAT signaling directly contributes to oncogenesis. Furthermore, extensive surveys of primary tumors and cell lines derived from tumors indicate that inappropriate activation of specific STATs occurs with surprisingly high frequency in a wide variety of human cancers. Together, these findings provide compelling evidence that aberrant STAT activation associated with oncogenesis is not merely adventitious but instead contributes to the process of malignant transformation. These studies are beginning to reveal the molecular mechanisms leading to STAT activation in the context of oncogenesis, and candidate genes regulated by STATs that may contribute to oncogenesis are being identified. Recent studies suggest that activated STAT signaling participates in oncogenesis by stimulating cell proliferation and preventing apoptosis. This review presents the evidence for critical roles of STATs in oncogenesis and discusses the potential for development of novel cancer therapies based on mechanistic understanding of STAT signaling. Oncogene (2000). PMID- 10851047 TI - STAT signaling in head and neck cancer. AB - The upper aerodigestive tract is predisposed to the formation of multiple primary tumors due to field cancerization. TGF-alpha/EGFR autocrine signaling appears to play an important role in squamous cell carcinoma of the head and neck (SCCHN) and upregulation of TGF-alpha and EGFR is an early event in SCCHN carcinogenesis. STAT proteins, including Stat3, are activated by TGF-alpha and EGFR and strategies that downmodulate TGF-alpha or EGFR inhibit SCCHN cell proliferation and abrogate Stat3 activation. Targeting Stat3 leads to SCCHN growth inhibition, increases apoptosis and a downmodulation of Bcl-xL expression in head and neck tumors. These studies support the role of Stat3 as an oncogene, which is activated early in SCCHN carcinogenesis, and efforts to understand EGFR-mediated Stat3 signaling could facilitate novel strategies that will interfere with this growth promoting pathway. Oncogene (2000). PMID- 10851048 TI - STAT signaling in the pathogenesis and treatment of leukemias. AB - Leukemias continue to cause significant mortality in adults and children, and the use of standard cytotoxic chemotherapy has reached a therapeutic plateau. Thus, there is great interest in treatments directed against inappropriately activated cell signaling pathways which stimulate the uncontrolled growth of neoplastic cells. Increasing evidence suggests that the STAT signaling cascade may be one target of these therapies. Signal transducer and activator of transcription (STAT) proteins are critical in mediating the response of hematopoietic cells to a diverse spectrum of cytokines. Constitutive STAT activation is present in many malignancies and has been especially well characterized in acute and chronic leukemias. While STAT activation is a common characteristic of leukemias, the specific pattern of activated STATs and the manner by which STAT activation occurs vary with each disease. STAT tyrosine phosphorylation can occur through inappropriate Jak activation or by direct activation of an oncoprotein such as Bcr/Abl, and STAT serine phosphorylation may play an important role in leukemias as well. Thus, the STAT signaling pathway is an attractive target for therapeutic intervention, and strategies designed to inhibit STAT activation and STAT mediated gene transcription may play an important role in the next generation of anti-leukemia therapies. Oncogene (2000). PMID- 10851049 TI - Divergent roles of STAT1 and STAT5 in malignancy as revealed by gene disruptions in mice. AB - Stat proteins are latent transcription factors activated by tyrosine phosphorylation downstream of cytokine and growth factor receptors and have been implicated in a variety of cell growth regulatory pathways. Constitutive phosphorylation has also been observed in various transformed cell line and in primary malignant tissue, suggesting that Stat protein activation may contribute to the transformed phenotype. One method to distinguish between a causative role in malignancy as opposed to bystander phosphorylation from the increased tyrosine phosphorylation that accompanies transformation is to investigate cell growth and malignancy in the absence of particular Stat proteins using targeted gene disruptions in transgenic mice. Such studies show that Stat1 primarily mediates growth inhibitory signals and contributes to the host rejection of tumors, and that its activation in transformed cells is not necessary for malignancy. Activation of Stat5 can be both necessary and sufficient for malignant transformation, and single Stat5-target genes have been identified that are critical for heightened proliferation. Nonetheless, some malignancies that are characterized by constitutively phosphorylated Stat5 are not altered by the loss of Stat5 protein. Its role in these cases may be redundant with other transforming events that are in themselves sufficient to cause disease, rendering tyrosine phosphorylation of Stat5 unnecessary in these transformed cells. Oncogene (2000). PMID- 10851050 TI - The role of STATs in myeloid differentiation and leukemia. AB - Myeloid differentiation is a highly regulated process governed by various cytokines, such as EPO, TPO, G-CSF, IL-3, IL-5 and GM-CSF. These cytokines act in part through activation of the STAT transcription factor family. In particular, various isoforms of STAT3 and STAT5 are activated during myeloid differentiation in a cell-type and maturation-state dependent fashion. In vitro studies have shown that STAT proteins are essential for cytokine-regulated processes such as cellular proliferation, differentiation as well as survival. Similarly, various STAT knock-outs have highlighted the role of STATs in myeloid differentiation in vivo. STATs also appear to play an important role in various myeloid malignancies, which are characterized by arrested maturation and cytokine independent proliferation of myeloid progenitors. Constitutive activation of STAT3 and/or STAT5 resulting in enhanced transcription of anti-apoptotic- cell cycle progression genes is likely to contribute to the pathogenesis of various myeloid leukemia's. Oncogene (2000). PMID- 10851051 TI - JAK-STAT signaling activated by Abl oncogenes. AB - The Abl oncoproteins v-Abl and BCR-Abl can activate member of the signal transducers and activators of transcription (STAT) family of signaling proteins. The mechanisms by which these oncoproteins activate STATs appear to differ. In cells transformed by v-Abl, Janus kinase (JAK) tyrosine kinases are constitutively activated. In these cells, the v-Abl oncoprotein and the JAK kinases physically associate. Mapping of the JAK interaction domain in v-Abl demonstrates that amino acids within the carboxyl terminal region of v-Abl bind JAKs through a direct interaction. A mutant of v-Abl lacking this region does not bind or activate JAK 1 in vivo, fails to activate STAT proteins, does not induce cellular proliferation, and is less efficient in cellular transformation. Kinase inactive mutants of JAK 1 inhibit the ability of v-Abl to activate STATs, to induce cytokine-independent proliferation, and to transform bone marrow cells. Interestingly, these effects correlate with defects in the activation of several pathways by v-Abl including Akt, PI3-kinase, STATs, and Ras. These data suggest that Jak kinases may play an important role in v-Abl induced transformation. Oncogene (2000). PMID- 10851052 TI - IL-3 signaling and the role of Src kinases, JAKs and STATs: a covert liaison unveiled. AB - Hematopoiesis is the cumulative result of intricately regulated signal transduction cascades that are mediated by cytokines and their cognate receptors. Proper culmination of these diverse signaling pathways forms the basis for an orderly generation of different cell types and aberrations in these pathways is an underlying cause for diseases such as cancer. Over the past several years, downstream events initiated upon cytokine/growth factor stimulation have been a major focus of biomedical research. As a result, several key concepts have emerged allowing a better understanding of the complex signaling processes. A group of novel transcription factors, termed signal transducers and activators of transcription (STATs) appear to orchestrate the downstream events propagated by cytokine/growth factor interactions with their cognate receptors. Until recently, the JAK proteins were considered to be the tyrosine kinases, which dictated the levels of phosphorylation and activation of STAT proteins, forming the basis of the JAK-STAT model. However, over the past few years, increasing evidence has accumulated which indicates that at least some of the STAT protein activation may be mediated by members of the Src gene family following cytokine/growth factor stimulation. Studies have demonstrated that the Src-family of tyrosine kinases can phosphorylate and activate certain STAT proteins, in lieu of JAK kinases. In such a scenario, JAK kinases may be more crucial to phosphorylation of the cytokine/growth factor receptors and in the process create docking sites on the receptors for binding of SH2-containing proteins such as STATs, Src-kinases and other signaling intermediates. Tyrosine phosphorylation and activation of STAT proteins can be achieved either by JAKs or Src-kinases depending on the nature of STAT that is being activated. This forms the basis for the JAK-Src-STAT model proposed in this review. The concerted action of JAK kinases, members of the Src kinase family and STAT proteins, leads to cell proliferation and cell survival, the end-point of the cytokine/growth factor stimulus. Oncogene (2000). PMID- 10851053 TI - Roles of STAT3 in mediating the cell growth, differentiation and survival signals relayed through the IL-6 family of cytokine receptors. AB - Members of the IL-6 cytokine family are involved in a variety of biological responses, including the immune response, inflammation, hematopoiesis, and oncogenesis by regulating cell growth, survival, and differentiation. These cytokines use gp130 as a common receptor subunit. The binding of ligand to gp130 activates the JAK/STAT signal transduction pathway, where STAT3 plays a central role in transmitting the signals from the membrane to the nucleus. STAT3 is essential for gp130-mediated cell survival and G1 to S cell-cycle-transition signals. Both c-myc and pim have been identified as target genes of STAT3 and together can compensate for STAT3 in cell survival and cell-cycle transition. STAT3 is also required for gp130-mediated maintenance of the pluripotential state of proliferating embryonic stem cells and for the gp130-induced macrophage differentiation of M1 cells. Furthermore, STAT3 regulates cell movement, such as leukocyte, epidermal cell, and keratinocyte migration. STAT3 also appears to regulate B cell differentiation into antibody-forming plasma cells. Since the IL 6/gp130/STAT3 signaling pathway is involved in both B cell growth and differentiation into plasma cells it is likely to play a central role in the generation of plasma cell neoplasias. Oncogene (2000). PMID- 10851054 TI - Jak-Stat signal transduction pathway through the eyes of cytokine class II receptor complexes. AB - Cells of the immune system communicate with each other to initiate, establish and maintain immune responses. The communication occurs through cell-to-cell contact or through a variety of intercellular mediators that include cytokines, chemokines, growth factors and hormones. In the case of cytokines, the signal is transmitted from the outside to the inside of a cell through cell surface receptors specific for each cytokine. At this step the signal is also decoded and amplified: ligand binding causes recruitment and/or activation of numerous cytoplasmic proteins. One cytokine can activate a number of signal transduction pathways leading to regulation of a wide array of biological activities. One of these pathways, the Jak-Stat pathway, is briefly reviewed here with respect to the class II cytokine receptors. Signal transduction through receptors for interferons Type I (IFN-alpha, IFN-beta, IFN-omega) and Type II (IFN-gamma), and interleukin 10 (IL-10) is described in detail. In addition, a complex between tissue factor (TF) and coagulation factor VIIa, and two new receptors related to the class II cytokine receptor family are discussed. Oncogene (2000). PMID- 10851055 TI - The role of Stat5a and Stat5b in signaling by IL-2 family cytokines. AB - The activation of Stat5 proteins (Stat5a and Stat5b) is one of the earliest signaling events mediated by IL-2 family cytokines, allowing the rapid delivery of signals from the membrane to the nucleus. Among STAT family proteins, Stat5a and Stat5b are the two most closely related STAT proteins. Together with other transcription factors and co-factors, they regulate the expression of the target genes in a cytokine-specific fashion. In addition to their activation by cytokines, activities of Stat5a and Stat5b, as well as other STAT proteins, are negatively controlled by CIS/SOCS/SSI family proteins. The outcome of Stat5 activation in regulating expression of target genes varies, depending upon the complexity of the promoter region of target genes and the other signaling pathways that are activated by each cytokine as well. Here, we mainly focus on the IL2-/IL-2 receptor system, as it is one of the best-studied systems that depend on Stat5-mediated signals. We will summarize what we have learned about the molecular mechanisms of how Stat5 is activated by IL-2 family cytokines from in vitro biochemical studies as well as the role that is played by Stat5 in each of the cytokine signaling pathways from in vivo gene-targeting analyses. Oncogene (2000). PMID- 10851056 TI - The biology of Stat4 and Stat6. AB - IL-4 and IL-12 are cytokines that are important regulators of the proliferation, differentiation and functional capacity of lymphocytes. STATs (signal transducers and activators of transcription) are transcription factors that provide a direct link between the cytokine receptors and cytokine induced gene transcription. Stat6 and Stat4 are two STAT family members that specifically mediate signals that emanate from the IL-4 and IL-12 receptors, respectively. Recently a great deal of progress has been made in understanding the specific roles that Stat6 and Stat4 play in lymphocyte function through in vitro as well as in vivo studies using Stat6 and Stat4-deficient mice. This report will summarize and describe the recent advances made in understanding the activation and regulation of Stat6 and Stat4 as well as their roles in the development of an immune response. Oncogene (2000). PMID- 10851057 TI - The role of STAT proteins in growth hormone signaling. AB - Growth hormone (GH) has long been known to be the body's primary regulator of body growth and a regulator of metabolism, yet the mechanisms by which GH regulates the transcription of specific genes required for these processes are just now being delineated. GH binding to its receptor recruits and activates the receptor-associated JAK2 that in turn phosphorylates tyrosines within itself and the GH receptor. These tyrosines form binding sites for a number of signaling proteins, including members of the family of signal transducers and activators of transcription (STAT). Among the known signaling molecules for GH, STAT proteins play a particularly prominent role in the regulation of gene transcription. This paper will review what is currently understood about which STAT proteins are regulated by GH, how they are regulated by GH, the GH-dependent genes they regulate, and discuss current theories about how GH-activated STAT signaling is regulated. Particular attention will be given to the novel role that STAT5 plays in sexually dimorphic gene expression in the liver as determined by the secretory pattern of GH and the role of STAT5 in body growth. Oncogene (2000). PMID- 10851058 TI - The roles of the Drosophila JAK/STAT pathway. AB - The JAK/STAT signal transduction pathway has been conserved throughout evolution such that true structural and functional homologues of components originally identified in vertebrate systems are also present in the model genetic system Drosophila melanogaster. In addition to roles during larval hematopoiesis reminiscent of the requirement for this pathway in mammalian systems, the JAK/STAT pathway in Drosophila is also involved in a number of other developmental events. Recent data has demonstrated further roles for the JAK/STAT pathway in the establishment of sexual identity via the early embryonic expression of Sex lethal, the segmentation of the embryo via the control of pair rule genes including even skipped and the establishment of polarity within the adult compound eye via a mechanism that includes the four jointed gene. Use of the powerful genetics in the model organism Drosophila may identify new components of the JAK/STAT pathway, define new roles for this pathway, and provide insights into the function of this signal transduction system. Here we review the roles of STAT and its associated signaling pathway during both embryonic and adult stages of Drosophila development and discuss future prospects for the identification and characterization of novel pathway components and targets. Oncogene (2000). PMID- 10851059 TI - Roles of STAT3 defined by tissue-specific gene targeting. AB - The physiological role of each individual STAT protein is now being examined through the study of 'knockout' (KO) mice, harboring a null allele for the particular gene. In contrast to other STATs deficient mice that are born alive, STAT3-deficient mice die during early embryogenesis. However, the role of STAT3 in adult tissues can be assessed by utilizing the Cre-loxP recombination system to ablate the gene in later life. Analyses of tissue-specific STAT3-deficient mice indicate that STAT3 plays a crucial role in a variety of biological functions including cell growth, suppression and induction of apoptosis, and cell motility. Oncogene (2000). PMID- 10851060 TI - Control of myeloid differentiation and survival by Stats. AB - Hematopoiesis involves a complex array of growth factors that regulate the survival and proliferation of immature progenitors, influence differentiation commitment, and modulate end-stage cell functions. This mini-review is focused on the role of Stat activation in the development of myeloid cells in response to hematopoietic cytokines. Much of the evidence implicating Stats in these cellular processes comes from studies of mutant cytokine receptors selectively uncoupled from Stat activation, dominant-inhibitory Stat mutants, and mice with targeted disruptions of Stat genes. Together these approaches provide strong evidence that Stat activation, particularly of Stat3 and Stat5, plays an important role in myeloid differentiation and survival. Oncogene (2000). PMID- 10851061 TI - Complex roles of Stat1 in regulating gene expression. AB - Stat1 is a fascinating and complex protein with multiple, yet contrasting transcriptional functions. Upon activation, it drives the expression of many genes but also suppresses the transcription of others. These opposing characteristics also apply to its role in facilitating crosstalk between signal transduction pathways, as it participates in both synergistic activation and inhibition of gene expression. Stat1 is a functional transcription factor even in the absence of inducer-mediated activation, participating in the constitutive expression of some genes. This review summarizes the well studied involvement of Stat1 in IFN-dependent and growth factor-dependent signaling and then describes the roles of Stat1 in positive, negative and constitutive regulation of gene expression as well as its participation in crosstalk between signal transduction pathways. Oncogene (2000). PMID- 10851062 TI - Serine phosphorylation of STATs. AB - Tyrosine phosphorylation regulates the dimerization of STATs as an essential prerequisite for the establishment of a classical JAK-STAT signaling path. However, most vertebrate STATs contain a second phosphorylation site within their C-termini. The phosphorylated residue in this case is a serine contained within a P(M)SP motif, and in the majority of situations its mutation to alanine alters transcription factor activity. This review addresses recent advances in understanding the regulation of STAT serine phosphorylation, as well as the kinases and other signal transducers implied in this process. The biochemical and biological consequences of STAT serine phosphorylation are discussed. Oncogene (2000). PMID- 10851063 TI - Modulation of STAT signaling by STAT-interacting proteins. AB - STATs (signal transducer and activator of transcription) play important roles in numerous cellular processes including immune responses, cell growth and differentiation, cell survival and apoptosis, and oncogenesis. In contrast to many other cellular signaling cascades, the STAT pathway is direct: STATs bind to receptors at the cell surface and translocate into the nucleus where they function as transcription factors to trigger gene activation. However, STATs do not act alone. A number of proteins are found to be associated with STATs. These STAT-interacting proteins function to modulate STAT signaling at various steps and mediate the crosstalk of STATs with other cellular signaling pathways. This article reviews the roles of STAT-interacting proteins in the regulation of STAT signaling. Oncogene (2000). PMID- 10851064 TI - Pharmaceutical intervention in the JAK/STAT signaling pathway. AB - Many cytokines exert their effect via the JAK/STAT signal transduction pathway. Due to the medical relevance of many of these cytokines, they are being exploited, either directly, or through antagonists, as therapeutics for a variety of serious diseases. Currently, these therapeutics consist almost entirely of protein products, with all of their attendant drawbacks. Delineation of the signaling mechanisms for the cytokines, however, has allowed the design and implementation of a variety of cell-based and biochemical screens for small molecule mimics or antagonists of these cytokines. Several successful assays will be described along with the advantages of each type of assay. Use of assays of this type should make it possible to discover numerous small molecule cytokine modulators with significant utility in the clinic. Oncogene (2000). PMID- 10851065 TI - Oncogenic transformation by ras and fos is mediated by c-Jun N-terminal phosphorylation. AB - The nuclear phosphoprotein c-Jun is a major component of the AP-1 transcription factor, whose activity is augmented by many oncogenes. An important mechanism to stimulate AP-1 function is N-terminal phosphorylation of c-Jun at the serine residues 63 and 73 by the c-JunN-terminal kinases (JNKs). Mice and cells harboring a mutant allele of c-jun, which has the JNK phosphoacceptor serines changed to alanines (junAA), were used to determine the function of c-Jun N terminal phosphorylation (JNP) during oncogenic transformation in vitro and in vivo. JunAA immortalized fibroblasts expressing v-ras and v-fos showed reduced tumorigenicity in nude mice, but the efficiency of v-src transformation was unaffected by the lack of JNP. To assess the significance of JNP in tumour development in vivo, two transgenic mouse tumour models were employed. Skin tumour development caused by constitutive activation of the ras pathway by K5-SOS F expression and c-fos-induced osteosarcoma formation were impaired in mice lacking JNP. Inhibition of JNP may, therefore, be a novel therapeutic strategy to inhibit tumour growth in vivo. Oncogene (2000). PMID- 10851066 TI - Interaction between protein tyrosine phosphatase and protein tyrosine kinase is involved in androgen-promoted growth of human prostate cancer cells. AB - Steroid hormones play key roles in regulating cell proliferation and differentiation in targeting tissues. However, in advanced cancers, the steroid hormone regulation is frequently attenuated through a yet unknown mechanism even in the presence of functional steroid hormone receptors. We investigate the functional role of tyrosine phosphorylation signaling in the hormone-refractory growth of human prostate tumors. Initial studies demonstrate that the androgen responsive phenotype of human prostate cancer cells associates with a low phosphotyrosine (p-Tyr) level of ErbB-2, which is regulated by cellular prostatic acid phosphatase (PAcP), a protein tyrosine phosphatase. In prostate cancer cells, the p-Tyr level, but not the protein level, of ErbB-2 inversely correlates with the androgen-responsiveness of cell proliferation. Androgen-stimulated cell growth concurs with a down-regulation of cellular PAcP, an elevated p-Tyr level of ErbB-2, and the activation of mitogen-activated protein kinases. Furthermore, only the ErbB-2 inhibitor AG 879, but not the EGFR inhibitor AG 1478, abolishes androgen-induced cell proliferation. Forced expression of ErbB-2 can also attenuate androgen promotion of cell growth. Data taken collectively conclude that in human prostate cancer cells, the tyrosine phosphorylation of ErbB-2 regulated by cellular PAcP plays a key role in regulating androgen-mediated proliferation signaling. Oncogene (2000). PMID- 10851067 TI - Hepatocellular carcinoma in WHV/N-myc2 transgenic mice: oncogenic mutations of beta-catenin and synergistic effect of p53 null alleles. AB - The intronless N-myc2 gene was originally identified as the major target of hepatitis virus insertion in woodchuck liver tumors. Here we report that transgenic mice carrying the N-myc2 gene controlled by woodchuck hepatitis virus (WHV) regulatory sequences are highly predisposed to liver cancer. In a WHV/N myc2 transgenic line, hepatocellular carcinomas or adenomas arose in over 70% of mice, despite barely detectable expression of the methylated transgene in liver cells. Furthermore, a transgenic founder carrying unmethylated transgene sequences succumbed to a large liver tumor by the age of two months, demonstrating the high oncogenicity of the woodchuck N-myc2 retroposon. Stabilizing mutations or deletions of beta-catenin were found in 25% of liver tumors and correlated with reduced tumor latency (P<0.05), confirming the important role of beta-catenin activation in Myc-induced tumorigenesis. The ability of the tumor suppressor gene p53 to cooperate with N-myc2 in liver cell transformation was tested by introducing a p53-null allele into WHV/N-myc2 transgenic mice. The loss of one p53 allele in transgenic animals markedly accelerated the onset of liver cancer (P=0.0001), and most tumors of WHV/N-myc2 p53+/Delta mice harbored either a deletion of the wt p53 allele or a beta-catenin mutation. These findings provide direct evidence that activation of N-myc2 and reduction of p53 levels act synergistically during multistage carcinogenesis in vivo and suggest that different genetic pathways may underlie liver carcinogenesis initiated by a myc transgene. Oncogene (2000). PMID- 10851068 TI - IGF-I receptor signaling in a prostatic cancer cell line with a PTEN mutation. AB - LNCaP prostatic cancer cells are characterized by having a PTEN mutation, low levels of type 1 insulin-like growth factor receptor (IGF-IR) and no IRS-1, one of the major substrates of the IGF-IR. The absence of IRS-1, an activator of PI3 kinase, is compensated in these cells by the mutation in PTEN, an inhibitor of PI3-kinase. However, IGF-IR signaling in the absence of IRS-1 can cause cell differentiation and growth arrest. We hypothesized that these three characteristics may not be unrelated, specifically that, together, they may favor the metastatic spread of prostatic cancer cells without decreasing their growth potential. In support of this hypothesis, we report here that: (1) IRS-1 expression increases cell adhesion and decreases cell motility; (2) over expression of the IGF-IR, in the absence of IRS-1, causes growth arrest and (3) a combination of IGF-IR and IRS-1 restores the transformed phenotype of LNCaP cells. These findings suggest a mechanism by which prostatic cancer cells can achieve metastatic potential without interfering with their growth potential. Oncogene (2000). PMID- 10851069 TI - Exogenous cdk4 overcomes reduced cdk4 RNA and inhibition of G1 progression in hematopoietic cells expressing a dominant-negative CBF - a model for overcoming inhibition of proliferation by CBF oncoproteins. AB - Core Binding Factor (CBF) is required for the development of definitive hematopoiesis, and the CBF oncoproteins AML1-ETO, TEL-AML1, and CBFbeta-SMMHC are commonly expressed in subsets of acute leukemia. CBFbeta-SMMHC slows the G1 to S cell cycle transition in hematopoietic cells, but the mechanism of this effect is uncertain. We have sought to determine whether inhibition of CBF-mediated trans activation is sufficient to slow proliferation. We demonstrate that activation of KRAB-AML1-ER, a protein containing the AML1 DNA-binding domain, the KRAB repression domain, and the Estrogen receptor ligand binding domain, also slows G1, if its DNA-binding domain is intact. Also, exogenous AML1 overcame CBFbeta SMMHC-induced inhibition of proliferation. Representational difference analysis (RDA) identified cdk4 RNA expression as an early target of KRAB-AML1 activation. Inhibition of CBF activities by KRAB-AML1-ER or CBFbeta-SMMHC rapidly reduced endogenous cdk4 mRNA levels, even in cells proliferating at or near control rates as a result of exogenous cdk4 expression. Over-expression of cdk4, especially a variant which cannot bind p16INK4a, overcame cell cycle inhibition resulting from activation of KRAB-AML1-ER, although cdk4 did not accelerate proliferation when expressed alone. These findings indicate that mutations which alter the expression of G1 regulatory proteins can overcome inhibition of proliferation by CBF oncoproteins. Oncogene (2000). PMID- 10851070 TI - The N-terminal common domain of simian virus 40 large T and small t antigens acts as a transformation suppressor of the HER-2/neu oncogene. AB - Overexpression of HER-2/neu (also known as c-erbB-2) proto-oncogene frequently occurs in many different types of human cancers, including ovarian carcinoma, and is known to enhance tumor metastasis and chemoresistance. Previous studies showed that inhibition of HER-2/neu expression by various agents, such as adenovirus E1A and simian virus 40 large T, can lead to suppression of tumorigenicity of HER 2/neu-overexpressing cancer cells. Here we report that T/t-common, which contains the N-terminal common domain of simian virus 40 large T and small t antigens, could specifically repress the HER-2/neu promoter. When the coding sequence of T/t-common was stably transfected into the HER-2/neu-overexpressing human ovarian carcinoma SK-OV-3 cells, the expression of HER-2/neu was dramatically reduced by the expression of T/t-common. Accordingly the tumorigenic potential of these T/t common-expressing clones, including the ability to grow anchorage-independently and the ability to induce tumor in nu/nu mice, was also drastically suppressed. Furthermore, when T/t-common was transiently cotransfected with the activated genomic neu into NIH3T3 cells, the transforming activity of the latter was suppressed by T/t-common in soft-agarose microcolony formation assays. Taken together, these data suggest that T/t-common may act as a transformation suppressor of the HER-2/neu oncogene. Oncogene (2000). PMID- 10851071 TI - Proneness to UV-induced apoptosis in human fibroblasts defective in transcription coupled repair is associated with the lack of Mdm2 transactivation. AB - The apoptotic response and the level of expression of p53 and of three genes transcriptionally activated by p53 (Mdm2, p21 and bax) were investigated in UV sensitive cells from patients with xeroderma pigmentosum (XP) or Cockayne syndrome (CS). These disorders are due to different genetic defects affecting transcription-coupled repair (TCR) and/or global genome repair (GGR), the nucleotide excision repair subpathways which remove UV-induced lesions from the transcribed strand of active genes or from the rest of the genome, respectively. After 20 J/m2 UV light, normal and GGR-defective XP-C fibroblasts showed rapid increase in p53, late induction of Mdm2 and no evidence of apoptosis even 96 h after irradiation. In contrast, in XP-A (defective in GGR and TCR), CS-A and CS-B (defective only in TCR) fibroblasts, the p53 increase was not followed by Mdm2 induction and the persistence of high levels of p53, due to the lack of its degradation by Mdm2, was associated with the appearance of apoptosis. Besides indicating that the persistence of DNA damage in the transcribed strand of active genes leads to apoptosis, these findings provide the first evidence that the lack of activation of Mdm2 plays a key role in the cascade of events leading to apoptosis. Oncogene (2000). PMID- 10851072 TI - Inhibition of tumor growth and metastasis of human melanoma by intracellular anti ATF-1 single chain Fv fragment. AB - Activating transcription factor-1 (ATF-1) and cAMP-responsive element (CRE) binding protein (CREB) have been implicated in cAMP and Ca2+-induced transcriptional activation. The expression of the transcription factors CREB and ATF-1 is upregulated in metastatic melanoma cells. However, how overexpression of ATF-1/CREB contributes to the acquisition of the metastatic phenotype remains unclear. Here, the effect of disrupting ATF-1 activity was investigated using intracellular expression of an inhibitory anti-ATF-1 single chain antibody fragment (ScFv). Intracellular expression of ScFv anti-ATF-1 in MeWo melanoma cells caused significant reduction in CRE-dependent promoter activation. In addition, expression of ScFv anti-ATF-1 in melanoma cells suppressed their tumorigenicity and metastatic potential in nude mice. ScFv anti-ATF-1 rendered the melanoma cells susceptible to thapsigargin-induced apoptosis in vitro and caused massive apoptosis in tumors transplanted subcutaneously into nude mice, suggesting that ATF-1 and its associated proteins act as survival factor for human melanoma cells. This is the first report to demonstrate the potential of ScFv anti-ATF-1 as an inhibitor of tumor growth and metastasis of solid tumor in vivo. Oncogene (2000). PMID- 10851073 TI - Cell cycle regulation of the endogenous wild type Bloom's syndrome DNA helicase. AB - Bloom's syndrome (BS) is a rare human autosomal recessive disorder characterized by an increased risk to develop cancer of all types. BS cells are characterized by a generalized genetic instability including a high level of sister chromatid exchanges. BS arises through mutations in both alleles of the BLM gene which encodes a 3' - 5' DNA helicase identified as a member of the RecQ family. We developed polyclonal antibodies specific for the NH2- and COOH-terminal region of BLM. Using these antibodies, we analysed BLM expression during the cell cycle and showed that the BLM protein accumulates to high levels in S phase, persists in G2/M and sharply declines in G1, strongly suggestive of degradation during mitosis. The BLM protein is subject to post-translational modifications in mitosis, as revealed by slow migrating forms of BLM found in both demecolcine treated cells and in mitotic cells isolated from non-treated asynchronous populations. Phosphatase treatment indicated that phosphorylation events were solely responsible for the appearance of the retarded moieties, a possible signal for subsequent degradation. Together, these results are consistent with a role of BLM in a replicative (S phase) and/or post-replicative (G2 phase) process. Oncogene (2000). PMID- 10851074 TI - PIK3CA as an oncogene in cervical cancer. AB - Amplification of chromosome arm 3q is the most consistent aberration in cervical cancer, and is implicated in the progression of dysplastic uterine cervical cells into invasive cancer. The present study employed the 'positional candidate gene' strategy to determine the contribution of PIK3CA, which is located in 3q26.3, in cervical tumorigenesis. PIK3CA is known to be involved in the PI 3-kinase/AKT signaling pathway, which plays an important role in regulating cell growth and apoptosis. The results of comparative genomic hybridization show that the 3q26.3 amplification was the most consistent chromosomal aberration in primary tissues of cervical carcinoma, and a positive correlation between an increased copy number of PIK3CA (detected by competitive PCR) and 3q26.3 amplification was found in tumor tissues and in cervical cancer cell lines. In cervical cancer cell lines harboring amplified PIK3CA, the expression of gene product (p110alpha) of PIK3CA was increased, and was subsequently associated with high kinase activity. In addition, transformation phenotypes in these lines, including increased cell growth and decreased apoptosis, were found to be significantly affected by the treatment of specific PI 3-kinase inhibitor, suggesting that increased expression of PIK3CA in cervical cancer may result in promoting cell proliferation and reducing apoptosis. These evidences support that PIK3CA is an oncogene in cervical cancer and PIK3CA amplification may be linked to cervical tumorigenesis. Oncogene (2000). PMID- 10851075 TI - The Rgr oncogene (homologous to RalGDS) induces transformation and gene expression by activating Ras, Ral and Rho mediated pathways. AB - The effects of the 5'-truncated Rgr oncogene, a previously shown specific guanine exchange factor for Ral in vitro, in stimulating proliferation, cell transformation and gene expression were investigated. We have established TetRgr cell lines in which expression of Rgr can be inhibited by the presence of tetracycline in the medium. Using this system, we show that Rgr overexpressing cells are morphologically transformed and grow in a disorganized manner. At the transcriptional level, Rgr enhances the activity of the serum response element and c-Jun. Rgr induces phosphorylation of ERKs, p38 and JNK kinases, and increases the levels of the GTP-bound forms of Ral and Ras. Ras activation could account for the broad spectra of effects displayed by Rgr. The important role of these pathways is confirmed by experiments in which the transcriptional activation events can be blocked by dominant negative versions of Ras, Ral and Rho. Among all the Rgr-induced pathways, the Ras-Raf-MEK-ERK cascade is essential for the transforming properties of Rgr. Additional analysis has shown that the activation of this pathway by Rgr is not due to a feed back mechanism mediated by the Grb2 adaptor protein. Oncogene (2000). PMID- 10851076 TI - Altered natural killer cell differentiation in CD34+ progenitors from chronic myeloid leukemia patients. AB - IL-15 and SCF fail to induce NK differentiation and proliferation of CD34+ hematopoietic progenitors from chronic myeloid leukemia patients in contrast to normal stem cells although, both normal and leukemic CD34+ cells display comparable expression of c-kit or IL-15 receptor subunits. Interestingly, confocal microscopy analysis revealed that leukemic and most normal CD34+ cells produce and secrete IL-15, as shown by its trafficking through the Golgi apparatus and early endosomes. However, only leukemic progenitors express the membrane bound IL-15. Colocalization and internalization of IL-15Rbeta/gammac and IL-15Ralpha/gammac complexes indicated that IL-15 was specifically uptaken by leukemic progenitors. We also demonstrated that in both normal and leukemic progenitors, the signaling kinase Jak3 is constitutively pre-associated with the gammac chain. Anti-IL-15 neutralizing mAb treatment resulted in down-regulation of gammac chain and disruption of gammac/Jak3 interaction in normal but had no effect in leukemic progenitors. Our results suggest the existence in both normal and leukemic CD34+ cells of a constitutive production of a bioactive IL-15 that does not lead to NK differentiation and further indicate that membrane bound IL 15 and constitutive activation of gammac are hallmarks of leukemic progenitors. Oncogene (2000). PMID- 10851077 TI - Initiation of translation from a downstream in-frame AUG codon on BRCA1 can generate the novel isoform protein DeltaBRCA1(17aa). AB - Expression of the breast and ovarian cancer gene BRCA1 is regulated at both the transcriptional and post-transcriptional levels. We found that the expression of the BRCA1 protein may also be regulated at the translational level. In addition to an AUG start codon at position 1, BRCA1 mRNA has a second in-frame AUG (+17) that acts as an alternative start codon to generate a novel BRCA1 protein that lacks the first 17 amino acids (DeltaBRCA1(17aa)). We fused cDNAs encoding the second exon of BRCA1 of the wild-type BRCA1 gene (wt-BRCA1) and a mutated BRCA1 gene (mt-BRCA1), in which the first initiation site and its Kozak consensus sequence were abolished, with the nucleophosmin (NPM) reporter gene and used them for in vitro and in vivo translation assays. In both systems, the wt-BRCA1-NPM constructs produced two distinct proteins (18 and 16 kD) begun from the first and second AUGs. The mt-BRCA1-NPM constructs produced only the shorter 16-kD protein lacking the first 17 amino acids of the BRCA1 gene. Next, we analysed the N terminal protein sequence of purified BRCA1 protein from normal thymocytes and found two different BRCA1 proteins, derived from translation of the first and second in-frame AUGs. Thus, BRCA1 protein expression can be regulated at the translation level in normal cells. Characterization of DeltaBRCA1(17aa) may shed light on the function and regulation of BRCA1 in normal cells as well as the pathogenesis of breast and ovarian cancers. Oncogene (2000). PMID- 10851078 TI - Mlh1 deficiency enhances several phenotypes of Apc(Min)/+ mice. AB - Defects in APC and DNA mismatch repair genes are associated with a strong predisposition to colon cancer in humans, and numerous mouse strains with mutations in these genes have been generated. In this report we describe the phenotype of Min/+ Mlh1-/- mice. We find that these doubly mutant mice develop more than three times the number of intestinal adenomas compared to Min/+ Mlh1+/+ or +/- mice but that these tumors do not show advanced progression in terms of tumor size or histological appearance. Full length Apc protein was not detected in the tumor cells from Min/+ Mlh1-/- mice. Molecular analyses indicated that in many tumors from Min/+ Mlh1-/- mice, Apc was inactivated by intragenic mutation. Mlh1 deficiency in Min/+ mice also led to an increase in cystic intestinal crypt multiplicity as well as enhancing desmoid tumorigenesis and epidermoid cyst development. Thus, Mlh1 deficiency influences the somatic events involved in the development of most of the phenotypes associated with the Min mutation. Oncogene (2000). PMID- 10851079 TI - Angiogenesis is an early event in the generation of myc-induced lymphomas. AB - Angiogenesis was identified as an early consequence of myc gene overexpression in two models of retroviral lymphomagenesis. Avian leukosis virus (ALV) induces bursal lymphoma in chickens after proviral c-myc gene integration, while the HB-1 retrovirus carries a v-myc oncogene and also induces metastatic lymphoma. Immunohistochemical studies of the effects of increased c-myc or v-myc overexpression revealed early angiogenesis within myc-transformed bursal follicles, which persisted in lymphomas and metastases. Abnormal vessel growth was consistently detected within 13 days after transplantation of a few myc overexpressing progenitors into ablated bursal follicles, suggesting that these angiogenic changes may support the initial expansion of tumor precursors, as well as later stage lymphomagenesis. Conditioned media from myc-overexpressing B cell lines promoted proliferation of vascular endothelium in vitro, while media from B cells expressing low myc levels showed little effect. Moreover, ectopic myc overexpression in the low myc B cell lines increased production of the endothelial growth activity, indicating that myc induces secretion of angiogenic factors from B cells. These findings demonstrate that myc overexpression in lymphocytes generates an angiogenic phenotype in vitro as well as in vivo. Oncogene (2000). PMID- 10851080 TI - The concerted regulatory functions of the transcription factors nuclear factor-1 and upstream stimulatory factor on a composite element in the promoter of the hepatocyte growth factor gene. AB - Hepatocyte growth factor (HGF) is an important multifunctional cytokine whose gene expression is regulated mainly at the transcriptional level. Previous studies using transgenic mice as well as in vitro analyses showed that a potential regulatory element(s) exists between -260 to -230 bp in the upstream region of the HGF gene promoter. In the present study, we have discovered that this region is a composite site through which members of the nuclear factor 1 (NF1) and upstream stimulatory factor (USF) families bind to and regulate HGF gene transcription. Gel mobility shift and supershift assays revealed that USF and NF1 have high binding affinity for this region and that the binding sites of the two different transcription factor families overlap. Functional studies showed that NF1 suppresses HGF gene promoter activity and that USF has an activating function. We found that the NF1/X and NF1/Red1 isoforms strongly suppressed HGF promoter activity while the NF1/L variant had no obvious effects. USF1, but not USF2, of the USF family stimulated HGF gene promoter activity. More interestingly, during liver regeneration after partial hepatectomy, a process which activates the HGF gene, we noted that the binding activity of USF to the HGF promoter element increased while that of NF1 decreased. These data provide insight into the molecular mechanisms that govern HGF gene transcription. Oncogene (2000). PMID- 10851081 TI - DNA repair and recombination factor Rad51 is over-expressed in human pancreatic adenocarcinoma. AB - Molecular processes that could contribute to differences in chemo- and radioresistance include variations in DNA repair mechanisms. In mammalian cells, the product of the rad51 gene mediates DNA repair via homologous recombination. We describe that in contrast to conventional monolayer cell systems Rad51 protein accumulates to high-levels in three-dimensional cell culture models as well as in orthotopic xeno-transplants of human pancreatic cancer cells. Strikingly, over expression of wild-type Rad51 was also found in 66% of human pancreatic adenocarcinoma tissue specimens. Functional analysis revealed that Rad51 over expression enhances survival of cells after induction of DNA double strand breaks. These data suggest that perturbations of Rad51 expression contribute to the malignant phenotype of pancreatic cancer. Oncogene (2000). PMID- 10851082 TI - Identification of the TCL6 genes within the breakpoint cluster region on chromosome 14q32 in T-cell leukemia. AB - A region on chromosome 14q32.1 is often involved in chromosomal translocations and inversions with one of the T-cell receptor loci in T-cell lymphoproliferative diseases. The breakpoints of the different rearrangements segregate into two clusters; a cluster due to inversion on the centromeric side and a cluster due to simple balanced translocations on the telomeric side. If the target gene activated by these different types of chromosomal rearrangements is the same, the gene must be localized between the two clusters of breakpoints in a region of around 160 kb. Within this breakpoint cluster region, we isolated two genes; namely, TCL1 and TML1/TCL1b genes. In the course of characterizing the TML1 gene, we further identified a third novel gene, which we named TCL6 (T-cell leukemia/lymphoma 6), from a region 7 kb upstream of the TML1 locus. The TCL6 gene expressed at least 11 isoforms through very complex alternative-splicing, including splicing with the TML1 gene. Those isoforms encode at least five open reading frames (ORFs) with no homology to known sequences. The localization of the proteins corresponding to these ORF was determined by fusing green fluorescence protein at the carboxyl terminal of each ORF. ORF141 and ORF72 were observed in the cytoplasmic region, while ORF105, ORF119, and ORF163 were predominantly localized in the nuclear region. Since the TCL6 gene was expressed in T-cell leukemia carrying a t(14;14)(q11;q32.1) chromosome translocation and was not expressed in normal T-cells (just like the TML1 and TCL1 genes), it is also a candidate gene potentially involved in leukemogenesis. Oncogene (2000). PMID- 10851083 TI - Tracheal development and the von Hippel-Lindau tumor suppressor homolog in Drosophila. AB - von Hippel-Lindau disease is a hereditary cancer syndrome. Mutations in the VHL tumor suppressor gene predispose individuals to highly vascularized tumors. However, VHL-deficient mice die in utero due to a lack of vascularization in the placenta. To resolve the contradiction, we cloned the Drosophila VHL homologue (d VHL) and studied its function. It showed an overall 50% similarity to the human counterpart and 76% similarity in the crucial functional domain: the elongin C binding site. The putative d-VHL protein can bind Drosophila elongin C in vitro. During embryogenesis, d-VHL is expressed in the developing tracheal regions where tube outgrowth no longer occurs. Reduced d-VHL activity (using RNA interference methodology) caused breakage of the main vasculature accompanied by excessive looping of smaller branches, whereas over-expression caused a general lack of vasculature. Importantly, human VHL can induce the same gain-of-function phenotypes. VHL is likely involved in halting cell migration at the end of vascular tube outgrowth. Loss of VHL activity can therefore lead to disruption of major vasculature (as in the mouse embryo), which requires precise cell movement and tube fusion, or ectopic outgrowth from existing secondary vascular branches (as in the adult tumors). Oncogene (2000) 19, 2803 - 2811 PMID- 10851084 TI - Degradation of human Aurora2 protein kinase by the anaphase-promoting complex ubiquitin-proteasome pathway. AB - Human Aurora2 was originally identified by its close homology to yeast IPL1 and fly aurora, which are key regulators of chromosome segregation and a family of serine/threonine kinases. Here we demonstrate that the Aurora2 protein is degraded rapidly after G2/M phase release in mammalian cells. Aurora2 protein has a rapid turnover rate with a half-life of approximately 2 h. In eukaryotic cells, the ubiquitin-proteasome pathway is the major mechanism for the targeted degradation of unstable proteins. The treatment of mammalian cells with proteasome inhibitors blocks Aurora2 degradation. Furthermore, Aurora2 is polyubiquitinated in vivo and in vitro using anaphase-promoting complex (APC). These results demonstrate that Aurora2 protein is turned over through the APC ubiquitin-proteasome pathway. Oncogene (2000) 19, 2812 - 2819 PMID- 10851085 TI - Calcineurin regulation of the mammalian G0/G1 checkpoint element, cyclin dependent kinase 4. AB - Cyclin dependent kinase 4 (cdk4) activity is controlled by the binding of regulatory subunits and inhibitory factors, as well as tyrosine and serine/threonine phosphorylation. More recently the influence of calcium levels have been demonstrated. Using transient transfections in Jurkat cells, we observed specific binding between cdk4 and the calcium and calmodulin activated serine/threonine phosphatase, calcineurin. Furthermore, we demonstrated that the inhibition of the phosphatase activity of calcineurin with FK506 and cyclosporin A resulted in an overall increase in cdk4 kinase activity, suggesting that the phosphatase activity of calcineurin was inhibitory to the kinase activity of cdk4. In contrast, we were not able to observe a similar effect on the kinase activity of either cdk6 or cdk2, indicating that the phosphatase activity of calcineurin was specific for cdk4. In addition, using an in vitro phosphatase assay for calcineurin, we observed that the exogenous addition of calcineurin resulted in the dephosphorylation of cdk4, an event that downregulated the kinase activity of cdk4. Calcineurin could, therefore, play an opposing role to the action of the cyclin activating kinase complex, an enzyme that upregulates the kinase activity of cdk4, an important G0/G1 checkpoint element in mammalian cells. Oncogene (2000) 19, 2820 - 2827 PMID- 10851087 TI - The chicken c-Jun 5' untranslated region directs translation by internal initiation. AB - The 5' untranslated region (UTR) of the chicken c-jun message is exceptionally GC rich and has the potential to form a complex and extremely stable secondary structure. Because stable RNA secondary structures can serve as obstacles to scanning ribosomes, their presence suggests inefficient translation or initiation through alternate mechanisms. We have examined the role of the c-jun 5' UTR with respect to its ability to influence translation both in vitro and in vivo. We find, using rabbit reticulocyte lysates, that the presence of the c-jun 5' UTR severely inhibits translation of both homologous and heterologous genes in vitro. Furthermore, translational inhibition correlates with the degree of secondary structure exhibited by the 5' UTR. Thus, in the rabbit reticulocyte lysate system, the c-jun 5' UTR likely impedes ribosome scanning resulting in inefficient translation. In contrast to our results in vitro, the c-jun 5' UTR does not inhibit translation in a variety of different cell lines suggesting that it may direct an alternate mechanism of translational initiation in vivo. To distinguish among the alternate mechanisms, we generated a series of bicistronic expression plasmids. Our results demonstrate that the downstream cistron, in the bicistronic gene, is expressed to a much higher level when directly preceded by the c-jun 5' UTR. In addition, inhibition of ribosome scanning on the bicistronic message, through insertion of a synthetic stable hairpin, inhibits translation of the first cistron but does not inhibit translation of the cistron downstream of the c-jun 5' UTR. These results are consistent with a model by which the c-jun message is translated through cap independent internal initiation. Oncogene (2000) 19, 2836 - 2845 PMID- 10851086 TI - Cyclin E induction by genotoxic stress leads to apoptosis of hematopoietic cells. AB - Cyclin E is essential for progression through the G1 phase of the cell cycle and initiation of DNA replication by interacting with, and activating its catalytic partner, the cyclin-dependent kinase 2 (Cdk2). We found a substantial increase in cyclin E mRNA, accompanied by increased production of cyclin E protein and cyclin E/Cdk2 kinase activity in multiple myeloma and lymphoma cells following irradiation. Cyclin E expression increased early in a time and dose-dependent manner, with a three-fold induction reached 8 h following gamma-irradiation. Run on analyses indicated a predominantly transcriptional regulation of cyclin E. Stable overexpression of cyclin E, but not cyclin D1, sensitized IM-9 cells to gamma-irradiation-induced apoptosis; in contrast, a dominant-negative Cdk2, prevented apoptosis. Irradiation of cyclin E overexpressing cells led to an enhanced caspase activation and exposure of the phosphatidylserine on the plasma membrane, two key markers of apoptosis, events which were completely abolished in cells expressing a dominant-negative Cdk2. This study identifies cyclin E as a target for activation by ionizing radiation and which plays a functional role in apoptosis of hematopoietic cells. Oncogene (2000) 19, 2828 - 2835 PMID- 10851088 TI - Hyperphosphorylation and increased proteolytic breakdown of c-Myb induced by the inhibition of Ser/Thr protein phosphatases. AB - The c-myb proto-oncogene encodes a nuclear phosphoprotein that plays a crucial role in normal hematopoiesis. It is a short-lived transcription factor rapidly degraded by the 26S proteasome. Although it has been shown that instability determinants reside in its carboxyl terminus, the molecular mechanism of c-Myb degradation is unknown. Here, we report the first evidence that phosphorylation plays a role in targeting the protein to the proteasome. Inhibition of cellular serine/threonine protein phosphatase activity by okadaic acid resulted in hyperphosphorylation of c-Myb and extremely rapid turnover. The hyperphosphorylation resulted in a protein with altered properties that was indicative of conformational changes. Its mobility on gel electrophoresis was altered as well as its recognition by specific monoclonal antibody. The altered hyperphosphorylated protein still bound to DNA with an affinity similar to that of the hypophosphorylated form. Phosphorylation of three previously identified sites, serines 11, 12, and 528, does not appear to be involved in the proposed changes in conformation or stability. However, phosphoamino acid analyses of the hyperphosphorylated form of c-Myb revealed increased c-Myb phosphorylation mainly on threonine residues that correlated with other okadaic acid-induced alterations of c-Myb. These findings indicate that Ser/Thr phosphatases prevent conformational changes that may play an important role in controlled degradation of c-Myb. Oncogene (2000) 19, 2846 - 2854 PMID- 10851089 TI - Yeast homolog of human SAG/ROC2/Rbx2/Hrt2 is essential for cell growth, but not for germination: chip profiling implicates its role in cell cycle regulation. AB - In an attempt to understand the signaling pathway mediating redox-induced apoptosis, we cloned SAG, an evolutionarily conserved zinc RING finger gene that, when overexpressed, protects cells from apoptosis induced by redox agents. Here we report functional characterization of SAG by the use of yeast genetics approach. Targeted disruption of ySAG, yeast homolog of human SAG, and subsequent tetrad analysis revealed that ySAG is required for yeast viability. Complementation experiment showed that the lethal phenotype induced by the ySAG deletion is fully rescued by wildtype SAG, but not by several hSAG mutants. Complementation experiment has also confirmed that ySAG is essential for normal vegetative growth, rather than being required for sporulation. Furthermore, cell death induced by SAG deletion was accompanied by cell enlargement and abnormal cell cycle profiling with an increased DNA content. Importantly, SAG was found to be the second family member of Rbx (RING box protein) or ROC (Regulator of cullins) or Hrt that is a component of SCF E3 ubiquitin ligase. Indeed, like ROC1/Rbx1/Hrt1, SAG binds to Cul1 and SAG-Cul1 complex has ubiquitin ligase activity to promote poly-ubiquitination of E2/Cdc34. This ligase activity is required for complementation of death phenotype induced by ySAG disruption. Finally, chip profiling of the entire yeast genome revealed induction of several G1/S as well as G2/M checkpoint control genes upon SAG withdrawal. Thus, SAG appears to control cell cycle progression in yeast by promoting ubiquitination and degradation of cell cycle regulatory proteins. Oncogene (2000) 19, 2855 - 2866 PMID- 10851090 TI - No requirement for src family kinases for PDGF signaling in fibroblasts expressing SV40 large T antigen. AB - A growing body of literature suggests that the ubiquitously expressed Src family kinases (Src, Fyn and Yes) are required for agents such as platelet-derived growth factor (PDGF) to stimulate DNA synthesis. Yet Klinghoffer and colleagues recently presented evidence that fibroblasts derived from mice null for Src, Fyn and Yes responded normally to PDGF (Klinghoffer et al., 1999, EMBO J., 18: 2459 - 2471). What is the reason for this discrepancy? We noted that Klinghoffer et al. (1999) used SV40 large T antigen (largeT) to facilitate derivation of cell lines from the embryos. We therefore tested the effect of largeT on PDGF receptor signaling. We found that expression of largeT overcame the inhibitory effects of interfering forms of both Ras (N17Ras) and Src (SrcK-). Furthermore, injection of SrcK- or the cst.1 antibody (which inhibits Src, Fyn and Yes) failed to inhibit PDGF-stimulated DNA synthesis in NIH3T3 cells expressing dominant negative p53, and fibroblasts derived from p53 null embryos. These data suggest firstly that caution should be used in interpretation of experiments conducted in cell lines expressing largeT, and secondly that the role of Src family kinases in growth factor signaling may be to oppose the effects of negative growth regulators such as p53. Oncogene (2000) 19, 2867 - 2869 PMID- 10851092 TI - Using generic measures of functional health and well-being to increase understanding of disease burden. PMID- 10851091 TI - Ubiquitin/proteasome-mediated degradation of p19INK4d determines its periodic expression during the cell cycle. AB - Assembly and activity of the proto-oncogenic cyclin D/CDK4(6) complexes, the major driving force of G1 phase progression, is negatively regulated by a family of INK4 CDK inhibitors p16INK4a, p15INK4b, p18INK4c, and p19INK4d. Expression of the INK4 family members is controlled at the transcriptional level, through differential response to environmental and intracellular signals such as cytokines, oncogenic overload, or cellular senescence. Here we show that the periodic oscillation of the p19INK4d protein during the cell cycle is determined by the ubiquitin/proteasome-dependent mechanism, allowing the protein abundance to follow the changes in its mRNA expression. Within the INK4 family, this regulatory mode appears restricted to p19INK4d whose ubiquitination was dependent on the integrity of lysine 62, and binding to CDK4. These results highlight unexpected differences among the INK4 inhibitors, and suggest how p19INK4d may help regulate the rate of cyclin D/CDK4(6) complex formation, and thereby timely progression through the mammalian cell division cycle. Oncogene (2000) 19, 2870 - 2876 PMID- 10851093 TI - 1999 International Society for the Study of the Lumbar Spine Presidential Address: education and quality care: our other missions. PMID- 10851094 TI - Application of nucleus pulposus to the nerve root simultaneously reduces blood flow in dorsal root ganglion and corresponding hindpaw in the rat. AB - STUDY DESIGN: An experimental study to clarify the effects of nucleus pulposus on blood flow in the dorsal root ganglion and hindpaws. OBJECTIVES: To investigate the effects of application of nucleus pulposus to nerve root on blood flow in the dorsal root ganglion and the corresponding hindpaw. SUMMARY OF BACKGROUND DATA: It has been reported experimentally that application of nucleus pulposus into the epidural space induces morphologic and functional changes in the nerve roots and induces compartment syndrome in the dorsal root ganglia. However, it has not been clarified which of these changes induces symptoms in the lower limbs. METHODS: Sixteen adult, female Sprague-Dawley rats had the left L5 nerve root and associated dorsal root ganglions exposed. Autologous nucleus pulposus was applied to the L5 nerve root, just proximal to the dorsal root ganglion (NP group). For control, the same volume of muscle tissue was applied similarly to the neural tissue (control group). Blood flow in the dorsal root ganglion, corresponding hindpaw, and the contralateral hindpaw was continuously monitored by two-channel laser Doppler flowmeter for 3 hours. After measurement of blood flow, the nerve root and dorsal root ganglion were processed for histology and evaluated by light microscope. RESULTS: Blood flow in the NP group was reduced, not only in the dorsal root ganglion, but also in the corresponding hindpaw. These reductions were statistically significant compared with the control group (P < 0.01). Edema was the principal pathologic finding seen consistently in the nerve roots and in many of the associated dorsal root ganglia from nucleus pulposus-treated animals. CONCLUSION: Application of nucleus pulposus to nerve root decreased blood flow in the dorsal root ganglion and corresponding hindpaw. These basic pathophysiologic changes are associated with compression injuries caused by herniated discs and are accepted neuropathologic mechanisms of injury associated with painful neuropathies. These acute observations in the dorsal root ganglion and the hindpaw may be important initial factors in the pathogenesis of radicular leg pain (sciatica) due to disc herniation. PMID- 10851095 TI - Intervertebral disc cell death is dependent on the magnitude and duration of spinal loading. AB - STUDY DESIGN: An in vivo study of the toxic consequences of static compressive stress on the intervertebral disc. OBJECTIVES: To determine whether disc cell death is correlated with the magnitude and duration of spinal compressive loading. SUMMARY OF BACKGROUND DATA: Static compression in vivo has been demonstrated to induce cell death. Cell death, in turn, has been associated with disc degeneration in humans. There are currently no tolerance criteria for the intervertebral disc that combine both biomechanical and biologic factors, although both have been implicated in cases of accelerated degeneration. METHODS: Mouse tail discs were loaded in vivo with an external compression device. Compressive stress was applied at one of two magnitudes (0.4 and 0.8 MPa) for 7 days, and at one additional magnitude (1.3 MPa) for 1, 3, and 7 days. Midsagittal sections of the discs were stained for apoptosis using the TdT-dUTP terminal nick end labeling (TUNEL) reaction. Quantal analysis was used to correlate the extent of cell death to the magnitude and duration of loading. RESULTS: The probit transformation of the percentage of dying cells was proportional to the sum of the logarithmic transformations of the compressive stress and the time of loading. CONCLUSIONS: The results of this study demonstrate the feasibility of developing a quantitative correlation between spinal loading and disc degeneration. Such a correlation may be coupled in the future to existing engineering models that predict spinal loading in response to physical exposures and lead to improved definition of the bounds of healthy and unhealthy spinal loading, and ultimately, refined guidelines for low back safety. PMID- 10851096 TI - Natural history of individuals with asymptomatic disc abnormalities in magnetic resonance imaging: predictors of low back pain-related medical consultation and work incapacity. AB - STUDY DESIGN: Prospective study on individuals with asymptomatic lumbar disc abnormalities detected in magnetic resonance imaging. OBJECTIVES: To determine the natural history of asymptomatic disc abnormalities in magnetic resonance imaging and to identify predictors of future low back pain-related medical consultation and work incapacity. SUMMARY OF BACKGROUND DATA: The natural history of individuals with asymptomatic disc herniations has not been well established, but the high rate of lumbar disc alterations recently detected in asymptomatic individuals by magnetic resonance imaging demands reconsideration of a pathomorphology-based explanation of low back pain and sciatica. METHODS: Forty six asymptomatic individuals who had a high rate of disc herniations (73%) were observed for an average of 5 years (range, 54-72 months). Four classes of variables (medical data including magnetic resonance imaging-identified disc abnormalities, general psychological factors, physical job characteristics, and psychosocial aspects of work) were assessed at baseline and follow-up. RESULTS: Disc herniations and neural compromise did not significantly worsen at follow-up, whereas disc degeneration progressed in 17 individuals (41.5%). Minor episodes of low back pain occurred in 19 individuals (41.3%), 6 of whom had to seek medical treatment and 5 of whom had to stop work temporarily. The requirement for low back pain-related medical consultation was predicted with high accuracy by listlessness, job satisfaction, and working in shifts (P < 0.001). Work incapacity was best predicted by physical job characteristics, job disaffection, and working in shifts (P < 0.01). CONCLUSION: Physical job characteristics and psychological aspects of work were more powerful than magnetic resonance imaging identified disc abnormalities in predicting the need for low back pain-related medical consultation and the resultant work incapacity. However,the conclusions are still preliminary, and replication of the findings in larger and more representative study samples is needed. PMID- 10851097 TI - Magnetic resonance evaluation of recurrent disc herniation: is gadolinium necessary? AB - STUDY DESIGN: Evaluation of magnetic resonance images (MRIs) with surgical reference standard. OBJECTIVES: To determine whether the addition of contrast enhanced MRI scans increases diagnostic efficacy in the evaluation of recurrent disc herniation. SUMMARY OF BACKGROUND DATA: Many centers now routinely use gadolinium-enhanced examinations in the evaluation of recurrent disc herniation. Others, noting the additional expense of contrast injection, advocate a more limited role for contrast injection and emphasize the importance of T2-weighted axial sequences. METHODS: The study included 165 consecutive patients who were referred to the authors' outpatient imaging center and had a history of previous lumbar discectomy and recurrent back and/or leg pain. The scanning protocol included sagittal and axial T1-weighted spin-echo pre- and postcontrast injection images and sagittal and axial T2-weighted fast spin-echo images. Twenty-eight patients (32 vertebral levels) had subsequent surgical exploration of a disc margin that had previously undergone discectomy. The surgical findings formed the reference standard. Three spine radiologists interpreted the MRI examinations without knowledge of the surgical results. They first interpreted the unenhanced studies, indicated whether they felt contrast injection would be helpful in further evaluation, and then (regardless of this determination) read the postcontrast study. RESULTS: On pre- and post-contrast examinations Reader 1 had a sensitivity of 95% (20/21), a specificity of 100% (10/10), and an accuracy of 97% (30/31). Reader 2 had a sensitivity of 95% (20/21), a specificity of 90% (9/10), and an accuracy of 94% (29/31). Reader 3 had a sensitivity of 90% (19/21), a specificity of 100% (10/10), and an accuracy of 94% on the precontrast examinations. His postcontrast performance demonstrated a sensitivity of 86% (18/21), a specificity of 100% (10/10), and an accuracy of 90% (28/31). In the nine interpretations wherein the readers thought that a contrast-enhanced examination might provide useful additional information, they did not change their interpretations in three cases, improved their interpretations in two, and made their interpretations worse in four on the basis of addition of the enhanced images. CONCLUSIONS: Routine use of contrast-enhanced examinations in patients who have had prior lumbar surgery probably adds little diagnostic value and may be confusing. PMID- 10851098 TI - Reoperations after lumbar disc surgery: a population-based study of regional and interspecialty variations. AB - STUDY DESIGN: A follow-up study using nationwide administrative databases. OBJECTIVES: To explore rates of reoperation after lumbar disc surgery and their regional and interspecialty variations. SUMMARY OF BACKGROUND DATA: In many Western countries, rates of lumbar disc surgery display significant geographic variations suggesting varying treatment criteria among operating surgeons. Few population-based studies have explored the risk of reoperation after disc surgery, and regional or interspecialty variations in the reoperations are unknown. METHODS: Patients who underwent lumbar spine surgery from January 1, 1987 through December 31, 1995, were identified in the Finnish Hospital Discharge Register. Data on the patients' initial disc operations, subsequent operations, and cause-of-death records were linked using personal identification codes. The Kaplan-Meier method and proportional hazard model were used to analyze risks of reoperation after initial surgery, according to hospital catchment area rates of disc surgery and for neurosurgical and orthopedic patients of university hospitals. RESULTS: 12.3% of 25,359 surgical patients with herniated lumbar discs underwent subsequent lumbar operations corresponding to the cumulative risk of 18.9% in the 9-year follow-up. Reoperation rates increased during the study period with the recent patient cohorts exhibiting risks. The reoperation risk showed a systematic geographic variation: the higher the regional disc surgery rate, the higher the reoperation risk. Overall, neurosurgical patients had a higher reoperation risk than orthopedic patients (relative risk [RR]: 1.57, 95% confidence interval [CI]: 1.17-2.10), but this was not a uniform finding. CONCLUSIONS: The reoperation risk after disc surgery increased during the study period and was higher in hospital catchment areas with higher overall discectomy rates. The reoperation risks varied among the university hospitals but tended to be higher for neurosurgical rather than for orthopedic patients. PMID- 10851099 TI - The impact of spinal problems on the health status of patients: have we underestimated the effect? AB - STUDY DESIGN: A prospective study of 17,774 patients who consulted spine centers in which the impact of spinal disorders and comorbidities on physical functional status were evaluated. OBJECTIVES: To quantify the effect spinal diagnoses have on patients' physical functional status (SF-36 Physical Component Summary [PCS] score) compared with other common conditions and to quantify the effects of comorbidities on physical functional status in spine patients. SUMMARY OF BACKGROUND DATA: The burden of spinal conditions on a patient's function and the role that comorbidities play in this affliction are poorly quantified in the literature. METHODS: Data from the Health Survey Questionnaire were prospectively gathered through the National Spine Network, a nonprofit consortium of spine focused practices. Each patient's SF-36 score was summarized into a single PCS score. The correlation between diagnosis and comorbidity and PCS score was assessed using multivariate linear regression. RESULTS: The study patients were a mean of 47.5 years of age, 54.7% were female, 52.3% had lumbosacral diagnoses, and 82.0% had had 3 or more months of pain. The population had a mean PCS score of 30.4 +/- 9.95 (SD) compared with 50.0 +/- 10.00 for the general United States population. The more comorbidities in a patient, the lower the PCS score (Spearman rank correlation = -0.27). The five comorbid conditions that lowered the PCS the most included congestive heart failure (CHF), chronic obstructive pulmonary disease (COPD), renal failure, rheumatoid arthritis, and lupus (all P <0.001). In multiple linear regression analysis, age, gender, diagnosis, and comorbidity explained 12.1% of the variance in PCS score. CONCLUSIONS: The PCS score is greatly affected in patients with spinal problems. The study population's PCS (30.4) was lower or similar to the PCS for patients with other illnesses reported in the literature: CHF (31.0), COPD (33.9), SLE (37.1), cancer (38.4), primary total hip arthroplasty (29.0), primary total knee arthroplasty (32.6), and glenohumeral degenerative joint disease (35.2). Further, the presence of comorbidity in spine patients adds to the burden of spinal conditions on functional status. PMID- 10851100 TI - Cauda equina syndrome secondary to lumbar disc herniation: a meta-analysis of surgical outcomes. AB - STUDY DESIGN: A meta-analysis of surgical outcomes of cauda equina syndrome secondary to lumbar disc herniation. OBJECTIVES: To determine the relationship between time to decompression after onset of cauda equina syndrome and clinical outcome, and to identify preoperative variables that were associated with outcomes. SUMMARY OF BACKGROUND DATA: The timing of surgical decompression for cauda equina syndrome is controversial. Although most surgeons recommend emergent decompression, results in certain studies show that delayed surgery may provide a satisfactory outcome. METHODS: A meta-analysis was performed to determine the correlation between timing of decompression and clinical outcome. One hundred four citations were reviewed, and 42 met the inclusion criteria. Preoperative and postoperative data were recorded. Length of time to surgery was broken down into five groups: less than 24 hours, 24-48 hours, 2-10 days, 11 days to 1 month, and more than 1 month. Logistic regression was used to determine the association between preoperative variables and postoperative outcomes. RESULTS: Outcomes were analyzed in 322 patients. Preoperative chronic back pain was associated with poorer outcomes in urinary and rectal function, and preoperative rectal dysfunction was associated with worsened outcome in urinary continence. In addition, increasing age was associated with poorer postoperative sexual function. No significant improvement in surgical outcome was identified with intervention less than 24 hours from the onset of cauda equina syndrome compared with patients treated within 24-48 hours. Similarly, no difference in outcome occurred in patients treated more than 48 hours after the onset of symptoms. Significant differences, however, were found in resolution of sensory and motor deficits as well as urinary and rectal function in patients treated within 48 hours compared with those treated more than 48 hours after onset of symptoms. CONCLUSIONS: There was a significant advantage to treating patients within 48 hours versus more than 48 hours after the onset of cauda equina syndrome. A significant improvement in sensory and motor deficits as well as urinary and rectal function occurred in patients who underwent decompression within 48 hours versus after 48 hours. PMID- 10851101 TI - Can exercise therapy improve the outcome of microdiscectomy? AB - STUDY DESIGN: A prospective randomized controlled trial of exercise therapy in patients who underwent microdiscectomy for prolapsed lumbar intervertebral disc. Results of a pilot study are presented. OBJECTIVE: To determine the effects of a postoperative exercise program on pain, disability, psychological status, and spinal function. SUMMARY OF BACKGROUND DATA: Microdiscectomy is often used successfully to treat prolapsed lumbar intervertebral disc. However, some patients do not have a good outcome and many continue to have low back pain. The reasons for this are unclear but impairment of back muscle function due to months of inactivity before surgery may be a contributing factor. A postoperative exercise program may improve outcome in such patients. METHODS: Twenty patients who underwent lumbar microdiscectomy were randomized into EXERCISE and CONTROL groups. After surgery, all patients received normal postoperative care that included advice from a physiotherapist about exercise and a return to normal activities. Six weeks after surgery, patients in the EXERCISE group undertook a 4 week exercise program that concentrated on improving strength and endurance of the back and abdominal muscles and mobility of the spine and hips. Assessments of spinal function were performed in all patients during the week before surgery and at 6, 10, 26, and 52 weeks after. The assessment included measures of posture, hip and lumbar mobility, back muscle endurance capacity and electromyographic measures of back muscle fatigue. On each occasion, patients completed questionnaires inquiring about pain, disability and psychological status. RESULTS: Surgery improved pain, disability, back muscle endurance capacity and hip and lumbar mobility in both groups of patients. After the exercise program, the EXERCISE group showed further improvements in these measures and also in electromyographic measures of back muscle fatigability. All these improvements were maintained 12 months after surgery. The only further improvement showed by the CONTROL group between 6 and 52 weeks was an increase in back muscle endurance capacity. CONCLUSION: A 4-week postoperative exercise program can improve pain, disability, and spinal function inpatients who undergo microdiscectomy. [Key words: electromyogram median frequency, exercise therapy, intervertebral disc prolapse, microdiscectomy, randomized controlled trial, spinal function. PMID- 10851102 TI - Prospective study of surgical treatment of degenerative spondylolisthesis: comparison between decompression alone and decompression with graf system stabilization. AB - STUDY DESIGN: A prospective study of patients with degenerative spondylolisthesis who underwent decompression of the spine, with and without stabilization using the Graf system. OBJECTIVES: To assess the clinical result of decompression alone and decompression using the Graf system. SUMMARY OF BACKGROUND DATA: The clinical outcome of lumbar stabilization for degenerative spondylolisthesis remains uncertain. There is no prospective study of differences in clinical outcome between patients who undergo decompression alone and those who undergo decompression and stabilization using the Graf system. METHODS: Eighty-eight patients with degenerative spondylolisthesis were included in this study. All patients reported leg symptoms. Decompression alone (Group D) was performed in 42 patients during a 5-year period from 1988 through 1992. Decompression and stabilization with the Graf system (Group G) was performed in 46 patients during a 4-year period from 1993 through 1996. There was no statistical difference regarding sex, the age at operation, compensable cases, and preoperative duration between two groups. The two groups were evaluated at follow-up examinations 1 and 3 years after surgery. The clinical results were evaluated for all patients by means of a 4-grade scale, visual analog scale, recurrence of leg symptoms, and persistent low back pain. The radiographic and clinical findings were examined by an independent investigator. RESULTS: The results according to the 4-grade scale deteriorated with time in both groups. There was no statistical difference between the two groups in the 4-grade scale, visual analog scale, or recurrence of leg symptoms at each follow-up time. Persistent low back pain in Group G was significantly lower than that in Group D at both the 1- and 3-year follow-ups. CONCLUSIONS: Although lumbar Graf stabilization had no effect in preventing the recurrence of leg symptoms, there was a significant effect on reduction of low back pain at the 1- and 3-year follow-ups. PMID- 10851103 TI - Radiation exposure during fluoroscopically assisted pedicle screw insertion in the lumbar spine. AB - STUDY DESIGN: An experimental model to assess radiation exposure during lumbar pedicle screw insertion. OBJECTIVES: To measure skin (patient) and scatter (surgeon) doses of radiation during lumbar spine fluoroscopy to assess safety of the procedure for both the surgeon and patient and determine best practice. SUMMARY OF BACKGROUND DATA: Fluoroscopy assists with accuracy of pedicle screw placement, yet the optimal technique of C-arm use and risk to both patient and operating room staff from radiation exposure are unknown. METHODS: Entry- and scatter-dose recordings were made using a digital dosimeter while screening an anthropomorphic phantom prone on a radiolucent operating table. The source was positioned both superiorly and inferiorly with the height varied in the latter orientation to create a working space under the C-arm. The senior author's fluoroscopy records were reviewed in 140 consecutive cases. RESULTS: In a series of 140 patients who underwent pedicle screw fixation, the fluoroscopy time was 1.4 minutes per case or 0.33 minutes per screw. In the source-superior position, the effective dose received by the patient was approximately 2.3 mSv per case. In the source-inferior position with a working space of 300 mm, the effective dose was 6.8 mSv. Scatter dose to the surgeon was higher in the source-superior position but was still less than 10% of recommended limits for the hand, thyroid, and eyes. CONCLUSIONS: The source-superior position is the preferred position for pedicle screw screening if a working space is required. Patient exposure is minimized, and surgeon dose is well within current recommendations. PMID- 10851104 TI - Morphologic considerations of C2 isthmus dimensions for the placement of transarticular screws. AB - STUDY DESIGN: This study examines the C2 vertebrae using both direct anatomic and computed tomographic measurements. OBJECTIVE: To define the relation of the C2 vertebrae bony elements to the vertebral artery and the spinal canal, to determine individuals at risk for vertebral artery injury during C1-C2 transarticular screw placement. SUMMARY OF BACKGROUND DATA: Recent literature assessing the safety of upper cervical spine transarticular screws has concentrated on technique, including the optimal point of entry and path projection of the screw. The actual dimensions of the C2 isthmus of the pars interarticularis has not been examined in a large number of specimens. METHODS: C2 isthmus width and height measurements were made on 205 human cadaveric C2 vertebrae. Vertebrae predicted to be at risk for vertebral arterial injury were imaged by computed tomography. RESULTS: There were 102 female and 103 male specimens with mean isthmus widths of 8.2 +/- 1.5 mm and 7.2 +/- 1.3 mm, respectively. Five specimens (2.4%) had an isthmus width less than 5 mm. The mean isthmus heights were 8.6 +/- 2.0 mm and 6.9 +/- 1.5 mm for male and female specimens, respectively. In twenty-four specimens (11.7%), one or both isthmi had a height of less than 5 mm. Six of these specimens were male and 18 were female. The right C2 isthmus was generally smaller than the left. Computed tomographic measurements closely approximated those of the actual dimensions of the isthmi. CONCLUSIONS: Placing a 3.5 mm screw in a patient with narrow C2 isthmus dimensions (smaller than 5 mm in either the height or width) is technically difficult. Because of narrow C2 isthmus width and/or height, approximately 10% of patients may be at risk for a vertebral artery injury with placement of C1-C2 transarticular screws. PMID- 10851105 TI - Load-carrying capacity of the human cervical spine in compression is increased under a follower load. AB - STUDY DESIGN: An experimental approach was used to test human cadaveric cervical spine specimens. OBJECTIVE: To assess the response of the cervical spine to a compressive follower load applied along a path that approximates the tangent to the curve of the cervical spine. SUMMARY OF BACKGROUND DATA: The compressive load on the human cervical spine is estimated to range from 120 to 1200 N during activities of daily living. Ex vivo experiments show it buckles at approximately 10 N. Differences between the estimated in vivo loads and the ex vivo load carrying capacity have not been satisfactorily explained. METHODS: A new experimental technique was developed for applying a compressive follower load of physiologic magnitudes up to 250 N. The experimental technique applied loads that minimized the internal shear forces and bending moments, loading the specimen in nearly pure compression. RESULTS: A compressive vertical load applied in the neutral and forward-flexed postures caused large changes in cervical lordosis at small load magnitudes. The specimen collapsed in extension or flexion at a load of less than 40 N. In sharp contrast, the cervical spine supported a load of up to 250 N without damage or instability in both the sagittal and frontal planes when the load path was tangential to the spinal curve. The cervical spine was significantly less flexible under a compressive follower load compared with the hypermobility demonstrated under a compressive vertical load (P < 0.05). CONCLUSION: The load-carrying capacity of the ligamentous cervical spine sharply increased under a compressive follower load. This experiment explains how a whole cervical spine can be lordotic and yet withstand the large compressive loads estimated in vivo without damage or instability. PMID- 10851106 TI - Biomechanical assessment of transoral plate fixation for atlantoaxial instability. AB - STUDY DESIGN: In an experimental study using human cadaver specimens the biomechanical data of anterior atlantoaxial plating according to Harms were evaluated. OBJECTIVES: The purpose of this study was to evaluate this method biomechanically. SUMMARY AND BACKGROUND DATA: The optimum fixation method to achieve fusion at the atlantoaxial joint after odontoid resection is still a matter of discussion. Isolated posterior surgical procedures for treatment of irreducible atlantoaxial kyphosis with spinal cord compression are associated with high rates of morbidity and mortality. Transoral atlantoaxial plate fixation has been designed by Harms as a fixation technique after odontoid resection. In a modification, this procedure has been combined with the posterior wire fusion according to Brooks. METHOD: Eight human cadaver craniocervical specimens were tested in flexion, extension, rotation, and bending with a nondestructive flexibility method using a nonconstrained testing apparatus. Five different groups were examined: 1) control group (intact); 2) unstable group (after dissection of the atlantoaxial ligaments and odontoidectomy), 3) Harms group (transoral atlantoaxial plate fixation) 4) Harms-Brooks group (transoral atlantoaxial plate fixation and dorsal atlantoaxial wire fixation); and 5) Magerl group (transarticular atlantoaxial screw fixation). In a second experimental series, failure loads of the Harms-Brooks and the Magerl fixation methods were determined. RESULTS: The angular displacement of the Harms-Brooks group and the Magerl group was less than in any other group. Stiffness values at 0-3.0 Nm loads in any direction were larger for the Harms-Brooks-and Magerl-fixated specimens than for the Harms, control, or unstable specimens. No statistically significant difference was observed between Harms-Brooks and Magerl reconstruction stiffness. Ultimate failure load in the Harms-Brooks group was higher than in the Magerl group. CONCLUSIONS: Experimentally, isolated anterior atlantoaxial plating was less stable than the combined reconstruction procedures. Transoral plate fixation according to Harms in combination with posterior wire fixation according to Brooks provided a failure load and stiffness equal to transarticular screw fixation according to Magerl. PMID- 10851107 TI - Dural ectasia is associated with back pain in Marfan syndrome. AB - STUDY DESIGN: A cross-sectional age- and sex-matched study comparing the prevalence and size of dural ectasia in two groups of patients with Marfan syndrome. Group I comprised patients with moderate to severe back pain and Group II comprised patients without back pain. OBJECTIVES: To determine whether the presence and size of dural ectasia is associated with back pain in patients with Marfan syndrome. SUMMARY OF BACKGROUND DATA: Dural ectasia is present in more than 60% of patients with Marfan syndrome. Moderate to severe back pain is present in more than 50% of patients with Marfan syndrome. Most cases of significant low back pain in patients with Marfan syndrome do not have a clear cause. It would be useful for the clinician to know whether dural ectasia may be a cause of back pain in patients with Marfan syndrome with no other source. METHODS: Thirty two volunteers aged 30-50 with Marfan syndrome were enrolled as age- and sex-matched pairs with significant back pain (Group I) and without back pain (Group II). A completed questionnaire, physical examination, and magnetic resonance image of the lumbosacral spine were obtained. Dural volume caudal to L5 was calculated from the magnetic resonance data by specially designed software. RESULTS: Dural ectasia was present in 76% of the patients in Group I, and 41% of the patients in Group II. The proportion of patients with dural ectasia was significantly higher in Group I. Furthermore, the mean dural volume was significantly higher in Group I, and a significant correlation between dural volume and Oswestry pain score was noted. CONCLUSIONS: The presence and size of dural ectasia are associated with back pain in the Marfan syndrome. However, a high prevalence of dural ectasia (41%) exists even in patients with Marfan syndrome without back pain. The mere presence of dural ectasia therefore does not necessarily mean the patient will be symptomatic even though the two are associated. PMID- 10851108 TI - Effects of a lifting belt on spine moments and muscle recruitments after unexpected sudden loading. AB - STUDY DESIGN: Ten men and eight women participated in a repeated-measures experiment in which sudden loads were applied unexpectedly to a container held in the hands. Three independent variables were investigated: lifting belt use, preload, and load symmetry. OBJECTIVES: To determine whether a lifting belt would help protect the spine in sudden symmetric and asymmetric loading situations. SUMMARY OF BACKGROUND DATA: Unexpected loading events have long been associated with the onset of back pain. Based on work showing that lifting belts restrict motion of the torso, the hypothesis was that a lifting belt would stiffen the spine, thereby protecting its supporting tissues. METHODS: A weight, equal to 7.5% of the subjects' trunk extension force, was allowed to fall 1 m before the bottom of a box held by blindfolded subjects was pulled. Kinetic and kinematic data, obtained from two force plates and a magnetic motion measurement system, were used in a three-dimensional, dynamic, linked-segment biomechanical model to calculate spine moments. Electromyogram data were simultaneously obtained from eight trunk muscles. RESULTS: The belt reduced the forward bending of the spine during the symmetric loadings. In the men, the belt also reduced the forward flexion moment acting on the spine. The belt restricted lateral bending in the women and men, when the box was preloaded. The peak electromyogram amplitudes from posterior contralateral erector spinae and latissimus dorsi muscles increased during the asymmetric loadings, whereas three ipsilateral muscles were less active. CONCLUSIONS: The conflicting moment and electromyographic results, combined with the influence of load symmetry, preload, and gender make the benefits of the lifting belt difficult to delineate. Although the data support the hypothesis that the belt stiffens the torso's response to sudden loading, the effects are small, and considerable individual differences exist. The findings show that during unexpected sudden loading, a belt may reduce the net external moment loading. At the same time the belt appears to alter the muscle response strategy so that the belt's overall effect on an individual's safety is hard to determine. PMID- 10851109 TI - Treatment of acute low back pain with the COX-2-selective anti-inflammatory drug nimesulide: results of a randomized, double-blind comparative trial versus ibuprofen. AB - STUDY DESIGN: A prospective, randomized double-blind comparative trial. OBJECTIVES: To evaluate the efficacy and tolerability of nimesulide, a cyclooxygenase (COX)-2-selective anti-inflammatory agent versus ibuprofen in patients with acute lumbosacral back pain. SUMMARY OF BACKGROUND DATA: Nonsteroidal anti-inflammatory drugs (NSAIDs) have been more effective than placebo in patients with uncomplicated acute low back pain in previous randomized controlled trials. The efficacy and tolerability of a new COX-2-selective anti inflammatory drug have not yet been established. METHODS: One hundred four patients aged 18-65 years with acute low back pain were enrolled. The patients were randomly allocated either to oral nimesulide (100 mg twice daily for 10 days) or oral ibuprofen (600 mg three times daily for 10 days). Outcome measures on a visual analog scale were an average of the pain intensity and the pain relief, stiffness in the back, functional status, and the results of physical examinations. All side effects were recorded at each visit. RESULTS: With both study therapies, there was a clear improvement in all measured parameters of the pain and back function parameters measured from the third day of treatment onward. The patients' capacity for daily tasks, showed improvement in both groups (P < 0. 001), but a statistically significant difference was found between the two groups in favor of the nimesulide group (P < 0.05) after 10 days. Nimesulide was more effective than ibuprofen in improved lateral bending measurements (P = 0.026). Nimesulide and ibuprofen provided similar degrees of improvement in the modified Schober tests and in the pain intensity and back stiffness scores. More gastrointestinal side effects were reported with ibuprofen than nimesulide, and the comparison showed a trend (P = 0.067). Ten side effects occurred in the nimesulide group in 7 (13%) patients and 13 in the ibuprofen group in 11 (21%) patients. CONCLUSIONS: The results confirmed that the COX-2-selective inhibitor nimesulide is an effective and well-tolerated agent for use in general practices to treat acute low back pain. The incidence of gastrointestinal side effects seems to be lower with nimesulide than with ibuprofen. PMID- 10851110 TI - Ureteral injury during laparoscopy-assisted anterior lumbar fusion. AB - STUDY DESIGN: A case report of ureteral injury as a complication incurred during a laparoscopy-assisted lumbar fusion. OBJECTIVE: To alert orthopedic surgeons to this injury, which may occur during such surgery. SUMMARY OF BACKGROUND DATA: Laparoscopy-assisted lumbar fusion is a minimally invasive surgical technique to accomplish lumbar fusion with excellent patient satisfaction, reduced hospital stay, and decreased rehabilitative time. METHOD AND RESULTS: A case report is presented detailing ureteral injury as a complication of laparoscopy-assisted lumbar fusion and the subsequent treatment of the injury. CONCLUSION: Laparoscopy assisted lumbar fusion is a new, less invasive technique with excellent patient satisfaction; however, ureteral injury may occur, and the surgeon should keep this in mind if a postoperative fluid collection occurs in the pelvis. PMID- 10851111 TI - Aneurysmal bone cyst of the spine with familial incidence. AB - STUDY DESIGN: A report of two cases of aneurysmal bone cysts of the spine occurring in a father and daughter. OBJECTIVE: To present an unusual finding of familial incidence of aneurysmal bone cyst and review the literature. SUMMARY OF BACKGROUND DATA: Aneurysmal bone cysts are benign, expanding, locally aggressive lesions. Up to 20% of cases involve the spine. The cause of primary aneurysmal bone cysts remains unclear. There have been three previous reports of a familial incidence supporting the importance of a hereditary component in the cause of aneurysmal bone cysts. METHODS: A 36-year-old man and a 7-year-old girl were diagnosed with aneurysmal bone cyst involving the spine by clinical manifestations, radiographic features, and histologic evaluation. RESULTS: The father remains recurrence- and symptom-free 6 years after primary resection. Five months after surgery, the daughter was found to have recurrent disease by magnetic resonance imaging and underwent a second procedure within 1 year of the primary resection. CONCLUSION: The occurrence of a primary aneurysmal bone cyst in two family members, occurring at adjacent vertebral levels, is suggestive of a hereditary component to the formation of primary aneurysmal bone cyst. PMID- 10851113 TI - Imagery PMID- 10851112 TI - Acute traumatic myelopathy secondary to a thoracic cyst in a professional football player. AB - STUDY DESIGN: Case report of acute traumatic myelopathy secondary to thoracic synovial cyst in a professional football player. OBJECTIVE: To emphasize examination for myelopathy and describe the radiographic and magnetic resonance findings of a rare source of traumatic myelopathy. BACKGROUND: Magnetic resonance imaging is the best initial evaluation for myelopathy in a traumatic setting. Heightened awareness during evaluation of a player involved in a traumatic incident allowed the diagnosis of potential cord damage and paralysis. METHODS: A subject with symptoms resulting from a direct blow to the back was evaluated for myelopathy, with diagnosis assisted by magnetic resonance imaging used to pinpoint the source of the disorder. RESULTS: The diagnosis allowed a surgical excision of the traumatic synovial cyst and full recovery of the injured football player. CONCLUSIONS: Awareness during examination for myelopathy in an acutely injured athlete is imperative to prompt the clinician to order the proper diagnostic studies and thereby embark on a surgical correction of the problem. PMID- 10851114 TI - Permeation through the CFTR chloride channel. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) protein forms a Cl(-) channel found in the plasma membranes of many epithelial cells, including those of the kidney, gut and conducting airways. Mutation of the gene encoding CFTR is the primary defect in cystic fibrosis, a disease that affects approximately 30 000 individuals in the United States alone. Alteration of CFTR function also plays an important role in the pathophysiology of secretory diarrhea and polycystic kidney disease. The basic mechanisms of permeation in this channel are not well understood. It is not known which portions of the protein contribute to forming the pore or which amino acid residues in those domains are involved in the biophysical processes of ion permeation. In this review, I will discuss (i) the present understanding of ion transport processes in the wild-type CFTR channel, (ii) the experimental approaches currently being applied to investigate the pore, and (iii) a proposed structure that takes into account the present data on mechanisms of ion selectivity in the CFTR channel and on blockade of the pore by open-channel blockers. PMID- 10851115 TI - Catarrhine photopigments are optimized for detecting targets against a foliage background. AB - The colour vision of many primates is trichromatic, whereas that of all other mammals is thought to be dichromatic or monochromatic. Moreover, the triplets of cone pigments in different catarrhines (Old World apes and monkeys) are strikingly similar in their spectral positions. We ask whether the selective advantage of trichromacy lies in an enhanced ability to find edible leaves or fruit. Further, we ask whether any factor in these two search tasks has constrained the particular set of cone spectral sensitivities observed in all catarrhines. We measured the spectral properties of the natural environments of six primate species in Uganda: Pan troglodytes, Cercopithecus mitis, Cercopithecus ascanius, Lophocebus albigena, Colobus guereza and Colobus badius. We concentrated on the fruit and leaves in their diets and the leaves of the trees that make up the background against which these diet items must be found. We plotted these measured stimuli in colour spaces appropriate for each primate species, and found that both frugivory and folivory are facilitated by the extra dimension of colour vision found in catarrhines but lacking in most other mammals. Furthermore, by treating the task of searching for food as a signal detection task, we show that, of all possible combinations of cone sensitivities, the spectral positions of the actual primate pigments are optimal for finding fruit or young leaves against the background of mature leaves. This is because the variance of the chromaticities of the mature leaves is minimised in one channel of the primate's colour vision, so allowing anything that is not a mature leaf to stand out. PMID- 10851116 TI - Chromaticity as a signal of ripeness in fruits taken by primates. AB - We have measured the reflectance spectra of many samples of fruit eaten by chimpanzees and three frugivorous monkey species. If the fruit are plotted in a colour space appropriate for catarrhine primates, several distinct ripening patterns are evident. The degree of ripeness of many species would be discernible by dichromatic primates, but for most fruit a trichromatic consumer would be at an advantage. However, by calculating which set of possible photopigments would maximise the chromatic distance between samples of each fruit species, we show that the spectral positions of the primate long- (L) and middle-wavelength (M) cone pigments are not optimised for this task. PMID- 10851117 TI - Slow muscle function of Pacific bonito (Sarda chiliensis) during steady swimming. AB - The Pacific bonito, Sarda chiliensis, is anatomically intermediate between mackerel and tuna. The specialisations exhibited by tuna are present in the bonito, but to a lesser degree. Slow-twitch muscle strain and activity patterns were determined during steady swimming (tailbeat frequency 1.2-3.2 Hz) at four locations on the body of Sarda chiliensis using sonomicrometry and electromyography. Both strain and the phase of electromygraphic activity were independent of tailbeat frequency. The strain of superficial slow-twitch muscle increased from +/-3.1 % l(0) at 0.35FL to +/-5.8 % l(0) at 0.65FL, where l(0) is muscle resting length and FL is the body length from snout to tail fork. Between 0.35 and 0.65FL, there was a negative phase shift of 16 degrees in the onset of electromygraphic activity in superficial slow-twitch muscle relative to the strain cycle. Muscle activity patterns are comparable with those of tuna. At 0.58FL, the onset of activity in deep slow-twitch muscle was approximately synchronous with the onset of activity in superficial muscle in the same myotome at 0.65FL. The distribution of slow-twitch muscle along the body of Sarda chiliensis and four additional fish species, Anguilla anguilla, Oncorhynchus mykiss, Scomber scombrus and Thunnus albacares, was also measured. Slow-twitch muscle appears to become more concentrated at approximately 0.5FL as swimming kinematics become more thunniform. PMID- 10851118 TI - Effects of catecholamines and purines on luminescence in the brittlestar Amphipholis squamata (Echinodermata). AB - The effects of catecholamines (dopamine, adrenaline, noradrenaline and its derivatives), 5-hydroxytryptamine and purines (adenosine, ATP and their derivatives) on the acetylcholine-induced luminescence of isolated arms and dissociated photocytes of the luminescent ophiuroid Amphipholis squamata were tested. The results showed that catecholamines and 5-hydroxytryptamine (10(-)(5) to 10(-)(3 )mol l(-)(1)) had a strong dose-dependent inhibitory effect on acetylcholine-induced luminescence. In contrast, purines (10(-)(4) and 10(-)(3 )mol l(-)(1)) triggered luminescence in the absence of acetylcholine and/or potentiated acetylcholine-induced luminescence. The results with specific purinergic agonists and antagonists indicated the involvement of P(1)- and P(2) like purinoceptors in the control of luminescence. Our study suggests that, in addition to the previously described cholinergic system in Amphipholis squamata, there may be a purinergic system, acting in synergy with acetylcholine, and an inhibitory neuromodulatory catecholaminergic system, all associated with the control of luminescence. PMID- 10851119 TI - Associative olfactory learning in the moth Manduca sexta. AB - The proboscis extension response conditioning protocol has been used to explore olfactory-based associative learning in an array of insects. We have monitored a different feeding reflex, which involves activation of the cibarial pump, to demonstrate olfactory learning in the moth Manduca sexta. In the first experiment, four different treatment conditions were used to assess associative (Pavlovian) learning. The results indicate that an excitatory cibarial pump response develops and is retained for at least 24 h only when odor is forward paired with the presentation of sucrose. Three control treatments, backward pairing, air (no odor) pairing and random pairing, failed to increase the cibarial pump response. However, an excitatory cibarial pump response developed when the backward- and air-paired groups received forward pairing of odor and sucrose on the following day. In contrast, moths experiencing random pairing on day 1 displayed a slower rate of acquisition during forward pairing on day 2, which may indicate inhibition. The second experiment investigated discrimination learning. Two odors were randomly presented, one odor being forward-paired with sucrose (+), the other presented alone (-) in a counterbalanced design. Again, only when odor was forward-paired with sucrose did learning occur. We discuss the implication of these findings for a broader comparative analysis of learning in insects. PMID- 10851120 TI - Chloride conductance across toad skin: effects of ionic acclimations and cyclic AMP and relationship to mitochondria-rich cell density. AB - The anionic conductance across toad (Bufo viridis) skin was studied using the voltage-clamp technique following long-term (more than 10 days) acclimation to NaCl and KCl solutions. The non-specific baseline conductance was approximately 0.6 mS cm(-)(2) and was similar in skins from all acclimation conditions. The voltage-activated Cl(-) conductance (G(Cl)) was maximal in skins from distilled water- and KCl-acclimated toads (>3 mS cm(-)(2)) and was greatly reduced following acclimation to NaCl solutions. Cyclic AMP (EC(50)=13 micromol l(-)(1)) and isobutylmethyl xanthine (IBMX) (EC(50)=69 micromol l(-)(1)) exerted different effects on the activated conductance. IBMX only sensitized the activated conductance, whereas cyclic AMP (CPTcAMP) at high concentrations induced an increase in anionic conductance that was insensitive to electrical potential. Furthermore, external Cl(-) was not required for the stimulatory effect of cyclic AMP, and the conductive pathway had low selectivity. The effects of the two agonists were reversible and depended on the acclimation conditions. Following electrical measurements, the skin of the toads was removed and stained with silver to measure mitochondria-rich cell density (D(mrc)). There was no correlation between D(mrc) and Cl(-) conductance in the present study. PMID- 10851121 TI - Blood pressure control in a larval amphibian, Xenopus laevis. AB - The regulation of arterial pressure in early vertebrate embryos and larvae with non-innervated hearts is poorly understood. We used nanoliter intravascular injections in anaesthetized Xenopus laevis larvae (stage 49-51) to assess their ability to maintain arterial pressure in the face of a volume load. Injections of saline and hetastarch (a volume expander) were made into the ventricle. Arterial pressure, end-diastolic ventricular volume, end-systolic ventricular volume and heart rate were measured. Injection of 800 nl caused a rapid rise in arterial pressure and stroke volume. There were no changes in heart rate, indicating the absence of an arterial baroreflex. Blood pressure in saline-injected animals recovered quickly (within 5 min), whereas hetastarch injections caused hypertension to be maintained for much longer, for over 40 min in the most extreme case. We surmise that Starling forces at the capillary play an important role in pressure regulation but are not adequate to explain the entire response. Finally, there was ample evidence for a Frank-Starling relationship in the ventricle. PMID- 10851122 TI - The mechanistic basis of aerobic performance variation in red junglefowl. AB - We examined aerobic performance, organ and muscle mass and enzymatic activity in red junglefowl (Gallus gallus). We tested three models of performance limitation (central limits, peripheral limits, symmorphosis) and explored relationships between basal metabolic rate (BMR), aerobic capacity ( V (O2max)) and social rank. Males had a lower BMR, a higher V (O2max) and a greater aerobic scope than females. Females possessed larger peritoneal and reproductive organs, while males had larger hearts, lungs and leg muscles. In females, BMR was correlated with spleen mass and V (O2max) was correlated with hematocrit and large intestine mass. Male BMR was correlated with intestinal tract and lung mass, and V (O2max) was correlated with heart and pectoralis mass. Male citrate synthase activity averaged 57 % higher than that of females and was correlated with V (O2max) (this correlation was not significant in females). Female social status was not correlated with any variable, but male dominance was associated with higher aerobic scope, larger heart and lungs, smaller peritoneal organs and greater leg citrate synthase activity. We conclude that aerobic capacity is controlled by system-wide limitations (symmorphosis) in males, while in females it is controlled by central organs. In neither sex is elevated aerobic capacity associated with increased maintenance costs. PMID- 10851123 TI - Effect of temperature on pH and electrolyte concentration in air-breathing ectotherms. AB - The aim of this study was to determine the effects of temperature upon pH, protein charge and acid-base-relevant ion exchange in air-breathing ectotherms. Plasma and skeletal muscles in cane toads (Bufo marinus) and bullfrogs (Rana catesbeiana) were examined at 30, 20 and 10 degrees C. In addition, skeletal muscle ion concentrations were examined in black racer snakes (Coluber constrictor) at 30 and 10 degrees C. Cooling the amphibians produced a reduction in most of the plasma ion concentrations (Na(+), K(+), Ca(2+), Cl(-), SO(4)(2)( )) and in protein concentration because of increased hydration. Between 30 and 10 degrees C, total plasma osmolality fell by 14 % in the toads and by 5 % in the frogs. Plasma protein charge, calculated using the principle of electroneutrality, was unaffected by temperature, except possibly for the toads at 10 degrees C. The in vivo skeletal muscle capdelta pHi/ capdelta T ratio, where pHi is intracellular pH and T is temperature, between 30 and 20 degrees C averaged -0.014 degrees C(-)(1) in the toads and -0.019 degrees C(-)(1) in the frogs. Between 20 and 10 degrees C, there was no change in pHi in the toads and a -0.005 degrees C(-)(1) change in the frogs. The in vitro skeletal muscle capdelta pHi/ capdelta T averaged -0.011 degrees C(-)(1) in both toads and frogs. In all three species, skeletal muscle inulin space declined with cooling. Intracellular ion concentrations were calculated by subtracting extracellular fluid ion concentrations from whole-muscle ion concentrations. In general, temperature had a large effect upon intracellular ion concentrations (Na(+), K(+), Cl(-)) and intracellular CO(2) levels. The relevance of the changes in intracellular ion concentration to skeletal muscle acid-base status and protein charge and the possible mechanisms producing the adjustments in intracellular ion concentration are discussed. It is concluded that ion-exchange mechanisms make an important contribution to adjusting pH with changes in temperature. PMID- 10851124 TI - RGS proteins inhibit Xwnt-8 signaling in Xenopus embryonic development. AB - RGS family members are GTPase activating proteins (GAPs) that antagonize signaling by heterotrimeric G proteins. Injection of Xenopus embryos with RNA encoding rat RGS4 (rRGS4), a GAP for G(i) and G(q), resulted in shortened trunks and decreased skeletal muscle. This phenotype is nearly identical to the effect of injection of either frzb or dominant negative Xwnt-8. Injection of human RGS2, which selectively deactivates G(q), had similar effects. rRGS4 inhibited the ability of early Xwnt-8 but not Xdsh misexpression to cause axis duplication. This effect is distinct from axin family members that contain RGS-like domains but act downstream of Xdsh. We identified two Xenopus RGS4 homologs, one of which, Xrgs4a, was expressed as a Spemann organizer component. Injection of Xenopus embryos with Xrgs4a also resulted in shortened trunks and decreased skeletal muscle. These results suggest that RGS proteins modulate Xwnt-8 signaling by attenuating the function of a G protein. PMID- 10851125 TI - The role of BicD, Egl, Orb and the microtubules in the restriction of meiosis to the Drosophila oocyte. AB - The oocyte is the only cell in Drosophila that goes through meiosis with meiotic recombination, but several germ cells in a 16-cell cyst enter meiosis and form synaptonemal complexes (SC) before one cell is selected to become the oocyte. Using an antibody that recognises a component of the SC or the synapsed chromosomes, we have analysed how meiosis becomes restricted to one cell, in relation to the other events in oocyte determination. Although BicD and egl mutants both cause the development of cysts with no oocyte, they have opposite effects on the behaviour of the SC: none of the cells in the cyst form SC in BicD null mutants, whereas all of the cells do in egl and orb mutants. Furthermore, unlike all cytoplasmic markers for the oocyte, the SC still becomes restricted to one cell when the microtubules are depolymerised, even though the BicD/Egl complex is not localised. These results lead us to propose a model in which BicD, Egl and Orb control entry into meiosis by regulating translation. PMID- 10851126 TI - Development of chick axial mesoderm: specification of prechordal mesoderm by anterior endoderm-derived TGFbeta family signalling. AB - Two populations of axial mesoderm cells can be recognised in the chick embryo, posterior notochord and anterior prechordal mesoderm. We have examined the cellular and molecular events that govern the specification of prechordal mesoderm. We report that notochord and prechordal mesoderm cells are intermingled and share expression of many markers as they initially extend out of Hensen's node. In vitro culture studies, together with in vivo grafting experiments, reveal that early extending axial mesoderm cells are labile and that their character may be defined subsequently through signals that derive from anterior endodermal tissues. Anterior endoderm elicits aspects of prechordal mesoderm identity in extending axial mesoderm by repressing notochord characteristics, briefly maintaining gsc expression and inducing BMP7 expression. Together these experiments suggest that, in vivo, signalling by anterior endoderm may determine the extent of prechordal mesoderm. The transforming growth factor (beta) (TGFbeta) superfamily members BMP2, BMP4, BMP7 and activin, all of which are transiently expressed in anterior endoderm mimic distinct aspects of its patterning actions. Together our results suggest that anterior endoderm-derived TGFbetas may specify prechordal mesoderm character in chick axial mesoderm. PMID- 10851127 TI - Fate determination of neural crest cells by NOTCH-mediated lateral inhibition and asymmetrical cell division during gangliogenesis. AB - Avian trunk neural crest cells give rise to a variety of cell types including neurons and satellite glial cells in peripheral ganglia. It is widely assumed that crest cell fate is regulated by environmental cues from surrounding embryonic tissues. However, it is not clear how such environmental cues could cause both neurons and glial cells to differentiate from crest-derived precursors in the same ganglionic locations. To elucidate this issue, we have examined expression and function of components of the NOTCH signaling pathway in early crest cells and in avian dorsal root ganglia. We have found that Delta1, which encodes a NOTCH ligand, is expressed in early crest-derived neuronal cells, and that NOTCH1 activation in crest cells prevents neuronal differentiation and permits glial differentiation in vitro. We also found that NUMB, a NOTCH antagonist, is asymmetrically segregated when some undifferentiated crest-derived cells in nascent dorsal root ganglia undergo mitosis. We conclude that neuron glia fate determination of crest cells is regulated, at least in part, by NOTCH mediated lateral inhibition among crest-derived cells, and by asymmetric cell division. PMID- 10851128 TI - Avian neural crest cell migration is diversely regulated by the two major hyaluronan-binding proteoglycans PG-M/versican and aggrecan. AB - It has been proposed that hyaluronan-binding proteoglycans play an important role as guiding cues during neural crest (NC) cell migration, but their precise function has not been elucidated. In this study, we examine the distribution, structure and putative role of the two major hyaluronan-binding proteoglycans, PG M/versicans and aggrecan, during the course of avian NC development. PG M/versicans V0 and V1 are shown to be the prevalent isoforms at initial and advanced phases of NC cell movement, whereas the V2 and V3 transcripts are first detected following gangliogenesis. During NC cell dispersion, mRNAs for PG M/versicans V0/V1 are transcribed by tissues lining the NC migratory pathways, as well as by tissues delimiting nonpermissive areas. Immunohistochemistry confirm the deposition of the macromolecules in these regions and highlight regional differences in the density of these proteoglycans. PG-M/versicans assembled within the sclerotome rearrange from an initially uniform distribution to a preferentially caudal localization, both at the mRNA and protein level. This reorganization is a direct consequence of the metameric NC cell migration through the rostral portion of the somites. As suggested by previous in situ hybridizations, aggrecan shows a virtually opposite distribution to PG M/versicans being confined to the perinotochordal ECM and extending dorsolaterally in a segmentally organized manner eventually to the entire spinal cord at axial levels interspacing the ganglia. PG-M/versicans purified from the NC migratory routes are highly polydispersed, have an apparent M(r) of 1,200 2,000 kDa, are primarily substituted with chondroitin-6-sulfates and, upon chondroitinase ABC digestion, are found to be composed of core proteins with apparent M(r )of 360-530, 000. TEM/rotary shadowing analysis of the isolated PG M/versicans confirmed that they exhibit the characteristic bi-globular shape, have core proteins with sizes predicted for the V0/V1 isoforms and carry relatively few extended glycosaminoglycan chains. Orthotopical implantation of PG M/versicans immobilized onto transplantable micromembranes tend to 'attract' moving cells toward them, whereas similar implantations of a notochordal type aggrecan retain both single and cohorts of moving NC cells in close proximity of the implant and thereby perturb their spatiotemporal migratory pattern. NC cells fail to migrate through three-dimensional collagen type I-aggrecan substrata in vitro, but locomote in a haptotactic manner through collagen type I-PG-M/versican V0 substrata via engagement of HNK-1 antigen-bearing cell surface components. The present data suggest that PG-M/versicans and notochordal aggrecan exert divergent guiding functions during NC cell dispersion, which are mediated by both their core proteins and glycosaminoglycan side chains and may involve 'haptotactic like' motility phenomena. Whereas aggrecan defines strictly impenetrable embryonic areas, PG-M/versicans are central components of the NC migratory pathways favoring the directed movement of the cells. PMID- 10851129 TI - In ovo time-lapse analysis after dorsal neural tube ablation shows rerouting of chick hindbrain neural crest. AB - Previous analyses of single neural crest cell trajectories have suggested important roles for interactions between neural crest cells and the environment, and amongst neural crest cells. To test the relative contribution of intrinsic versus extrinsic information in guiding cells to their appropriate sites, we ablated subpopulations of premigratory chick hindbrain neural crest and followed the remaining neural crest cells over time using a new in ovo imaging technique. Neural crest cell migratory behaviors are dramatically different in ablated compared with unoperated embryos. Deviations from normal migration appear either shortly after cells emerge from the neural tube or en route to the branchial arches, areas where cell-cell interactions typically occur between neural crest cells in normal embryos. Unlike the persistent, directed trajectories in normal embryos, neural crest cells frequently change direction and move somewhat chaotically after ablation. In addition, the migration of neural crest cells in collective chains, commonly observed in normal embryos, was severely disrupted. Hindbrain neural crest cells have the capacity to reroute their migratory pathways and thus compensate for missing neural crest cells after ablation of neighboring populations. Because the alterations in neural crest cell migration are most dramatic in regions that would normally foster cell-cell interactions, the trajectories reported here argue that cell-cell interactions have a key role in the shaping of the neural crest migration. PMID- 10851130 TI - An FGF signal from endoderm and localized factors in the posterior-vegetal egg cytoplasm pattern the mesodermal tissues in the ascidian embryo. AB - The major mesodermal tissues of ascidian larvae are muscle, notochord and mesenchyme. They are derived from the marginal zone surrounding the endoderm area in the vegetal hemisphere. Muscle fate is specified by localized ooplasmic determinants, whereas specification of notochord and mesenchyme requires inducing signals from endoderm at the 32-cell stage. In the present study, we demonstrated that all endoderm precursors were able to induce formation of notochord and mesenchyme cells in presumptive notochord and mesenchyme blastomeres, respectively, indicating that the type of tissue induced depends on differences in the responsiveness of the signal-receiving blastomeres. Basic fibroblast growth factor (bFGF), but not activin A, induced formation of mesenchyme cells as well as notochord cells. Treatment of mesenchyme-muscle precursors isolated from early 32-cell embryos with bFGF promoted mesenchyme fate and suppressed muscle fate, which is a default fate assigned by the posterior-vegetal cytoplasm (PVC) of the eggs. The sensitivity of the mesenchyme precursors to bFGF reached a maximum at the 32-cell stage, and the time required for effective induction of mesenchyme cells was only 10 minutes, features similar to those of notochord induction. These results support the idea that the distinct tissue types, notochord and mesenchyme, are induced by the same signaling molecule originating from endoderm precursors. We also demonstrated that the PVC causes the difference in the responsiveness of notochord and mesenchyme precursor blastomeres. Removal of the PVC resulted in loss of mesenchyme and in ectopic notochord formation. In contrast, transplantation of the PVC led to ectopic formation of mesenchyme cells and loss of notochord. Thus, in normal development, notochord is induced by an FGF-like signal in the anterior margin of the vegetal hemisphere, where PVC is absent, and mesenchyme is induced by an FGF-like signal in the posterior margin, where PVC is present. The whole picture of mesodermal patterning in ascidian embryos is now known. We also discuss the importance of FGF induced asymmetric divisions, of notochord and mesenchyme precursor blastomeres at the 64-cell stage. PMID- 10851131 TI - Progressive restriction in fate potential by neural progenitors during cerebral cortical development. AB - During early stages of cerebral cortical development, progenitor cells in the ventricular zone are multipotent, producing neurons of many layers over successive cell divisions. The laminar fate of their progeny depends on environmental cues to which the cells respond prior to mitosis. By the end of neurogenesis, however, progenitors are lineally committed to producing upper layer neurons. Here we assess the laminar fate potential of progenitors at a middle stage of cortical development. The progenitors of layer 4 neurons were first transplanted into older brains in which layer 2/3 was being generated. The transplanted neurons adopted a laminar fate appropriate for the new environment (layer 2/3), revealing that layer 4 progenitors are multipotent. Mid-stage progenitors were then transplanted into a younger environment, in which layer 6 neurons were being generated. The transplanted neurons bypassed layer 6, revealing that layer 4 progenitors have a restricted fate potential and are incompetent to respond to environmental cues that trigger layer 6 production. Instead, the transplanted cells migrated to layer 4, the position typical of their origin, and also to layer 5, a position appropriate for neither the host nor the donor environment. Because layer 5 neurogenesis is complete by the stage that progenitors were removed for transplantation, restrictions in laminar fate potential must lag behind the final production of a cortical layer. These results suggest that a combination of intrinsic and environmental cues controls the competence of cortical progenitor cells to produce neurons of different layers. PMID- 10851132 TI - Delta signaling mediates segregation of neural crest and spinal sensory neurons from zebrafish lateral neural plate. AB - We examined the role of Delta signaling in specification of two derivatives in zebrafish neural plate: Rohon-Beard spinal sensory neurons and neural crest. deltaA-expressing Rohon-Beard neurons are intermingled with premigratory neural crest cells in the trunk lateral neural plate. Embryos homozygous for a point mutation in deltaA, or with experimentally reduced delta signalling, have supernumerary Rohon-Beard neurons, reduced trunk-level expression of neural crest markers and lack trunk neural crest derivatives. Fin mesenchyme, a putative trunk neural crest derivative, is present in deltaA mutants, suggesting it segregates from other neural crest derivatives as early as the neural plate stage. Cranial neural crest derivatives are also present in deltaA mutants, revealing a genetic difference in regulation of trunk and cranial neural crest development. PMID- 10851133 TI - Persistent expression of HNF6 in islet endocrine cells causes disrupted islet architecture and loss of beta cell function. AB - We used transgenesis to explore the requirement for downregulation of hepatocyte nuclear factor 6 (HNF6) expression in the assembly, differentiation, and function of pancreatic islets. In vivo, HNF6 expression becomes downregulated in pancreatic endocrine cells at 18. 5 days post coitum (d.p.c.), when definitive islets first begin to organize. We used an islet-specific regulatory element (pdx1(PB)) from pancreatic/duodenal homeobox (pdx1) gene to maintain HNF6 expression in endocrine cells beyond 18.5 d.p.c. Transgenic animals were diabetic. HNF6-overexpressing islets were hyperplastic and remained very close to the pancreatic ducts. Strikingly, alpha, delta, and PP cells were increased in number and abnormally intermingled with islet beta cells. Although several mature beta cell markers were expressed in beta cells of transgenic islets, the glucose transporter GLUT2 was absent or severely reduced. As glucose uptake/metabolism is essential for insulin secretion, decreased GLUT2 may contribute to the etiology of diabetes in pdx1(PB)-HNF6 transgenics. Concordantly, blood insulin was not raised by glucose challenge, suggesting profound beta cell dysfunction. Thus, we have shown that HNF6 downregulation during islet ontogeny is critical to normal pancreas formation and function: continued expression impairs the clustering of endocrine cells and their separation from the ductal epithelium, disrupts the spatial organization of endocrine cell types within the islet, and severely compromises beta cell physiology, leading to overt diabetes. PMID- 10851134 TI - A conditional rescue system reveals essential functions for the ecdysone receptor (EcR) gene during molting and metamorphosis in Drosophila. AB - In Drosophila, pulses of the steroid hormone ecdysone trigger larval molting and metamorphosis and coordinate aspects of embryonic development and adult reproduction. At each of these developmental stages, the ecdysone signal is thought to act through a heteromeric receptor composed of the EcR and USP nuclear receptor proteins. Mutations that inactivate all EcR protein isoforms (EcR-A, EcR B1, and EcR-B2) are embryonic lethal, hindering analysis of EcR function during later development. Using transgenes in which a heat shock promoter drives expression of an EcR cDNA, we have employed temperature-dependent rescue of EcR null mutants to determine EcR requirements at later stages of development. Our results show that EcR is required for hatching, at each larval molt, and for the initiation of metamorphosis. In EcR mutants arrested prior to metamorphosis, expression of ecdysone-responsive genes is blocked and normal ecdysone responses of both imaginal and larval tissues are blocked at an early stage. These results show that EcR mediates ecdysone signaling at multiple developmental stages and implicate EcR in the reorganization of imaginal and larval tissues at the onset of metamorphosis. PMID- 10851135 TI - Combinatorial RNA interference indicates GLH-4 can compensate for GLH-1; these two P granule components are critical for fertility in C. elegans. AB - We report that four putative germline RNA helicases, GLHs, are components of the germline-specific P granules in Caenorhabditis elegans. GLH-3 and GLH-4, newly discovered, belong to a multi-gene glh family. Although GLHs are homologous to Drosophila VASA, a polar granule component necessary for oogenesis and embryonic pattern formation, the GLHs are distinguished by containing multiple CCHC zinc fingers. RNA-mediated interference (RNAi) reveals the GLHs are critical for oogenesis. By RNAi at 20 degrees C, when either loss of GLH-1 or GLH-4 alone has no effect, loss of both GLH-1 and GLH-4 results in 97% sterility in the glh 1/4(RNAi) offspring of injected hermaphrodites. glh-1/4(RNAi) germlines are under proliferated and are without oocytes. glh-1/4(RNAi) animals produce sperm; however, spermatogenesis is delayed and the sperm are defective. P granules are still present in glh-1/4(RNAi) sterile worms as revealed with antibodies against the remaining GLH-2 and GLH-3 proteins, indicating the GLHs function independently in P granule assembly. These studies reveal that C.elegans can use GLH-1 or GLH-4 to promote germline development. PMID- 10851136 TI - Endogenous patterns of TGFbeta superfamily signaling during early Xenopus development. AB - Transforming growth factor beta (TGFbeta) superfamily signaling has been implicated in patterning of the early Xenopus embryo. Upon ligand stimulation, TGFbeta receptors phosphorylate Smad proteins at carboxy-terminal SS(V/M)S consensus motifs. Smads 1/5/8, activated by bone morphogenetic protein (BMP) signaling, induce ventral mesoderm whereas Smad2, activated by activin-like ligands, induces dorsal mesoderm. Although ectopic expression studies are consistent with roles for TGFbeta signals in early Xenopus embryogenesis, when and where BMP and activin-like signaling pathways are active endogenously has not been directly examined. In this study, we investigate the temporal and spatial activation of TGFbeta superfamily signaling in early Xenopus development by using antibodies specific for the type I receptor-phosphorylated forms of Smad1/5/8 and Smad2. We find that Smad1/5/8 and two distinct isoforms of Smad2, full-length Smad2 and Smad2(delta)exon3, are phosphorylated in early embryos. Both Smad1/5/8 and Smad2/Smad2(delta)exon3 are activated after, but not before, the mid-blastula transition (MBT). Endogenous activation of Smad2/Smad2(delta)exon3 requires zygotic transcription, while Smad1/5/8 activation at MBT appears to involve transcription-independent regulation. We also find that the competence of embryonic cells to respond to TGF(delta) superfamily ligands is temporally regulated and may be a determinant of early patterning. Levels of phospho Smad1/5/8 and of phospho-Smad2/Smad2(delta)exon3 are asymmetrically distributed across both the animal-vegetal and dorsoventral axes. The timing of the development of these asymmetries differs for phospho-Smad1/5/8 and for phospho Smad2/Smad2(delta)exon3, and the spatial distribution of phosphorylation of each Smad changes dramatically as gastrulation begins. We discuss the implications of our results for endogenous functions of BMP and activin-like signals as candidate morphogens regulating primary germ layer formation and dorsoventral patterning of the early Xenopus embryo. PMID- 10851137 TI - The bHLH gene Hes6, an inhibitor of Hes1, promotes neuronal differentiation. AB - We have isolated the basic helix-loop-helix (bHLH) gene Hes6, a novel member of the family of mammalian homologues of Drosophila hairy and Enhancer of split. Hes6 is expressed by both undifferentiated and differentiated cells, unlike Hes1, which is expressed only by the former cells. Hes6 alone does not bind to the DNA but suppresses Hes1 from repressing transcription. In addition, Hes6 suppresses Hes1 from inhibiting Mash1-E47 heterodimer and thereby enables Mash1 and E47 to upregulate transcription in the presence of Hes1. Furthermore, misexpression of Hes6 with retrovirus in the developing retina promotes rod photoreceptor differentiation, like Mash1, in sharp contrast to Hes1, which inhibits cell differentiation. These results suggest that Hes6 is an inhibitor of Hes1, supports Mash1 activity and promotes cell differentiation. Mutation analysis revealed that Hes1- and Hes6-specific functions are, at least in part, interchangeable by alteration of the loop region, suggesting that the loop is not simply a nonfunctional spacer but plays an important role in the specific functions. PMID- 10851138 TI - The homeodomain-containing gene Xdbx inhibits neuronal differentiation in the developing embryo. AB - The development of the vertebrate nervous system depends upon striking a balance between differentiating neurons and neural progenitors in the early embryo. Our findings suggest that the homeodomain-containing gene Xdbx regulates this balance by maintaining neural progenitor populations within specific regions of the neuroectoderm. In posterior regions of the Xenopus embryo, Xdbx is expressed in a bilaterally symmetric stripe that lies at the middle of the mediolateral axis of the neural plate. This stripe of Xdbx expression overlaps the expression domain of the proneural basic/helix-loop-helix-containing gene, Xash3, and is juxtaposed to the expression domains of Xenopus Neurogenin related 1 and N-tubulin, markers of early neurogenesis in the embryo. Xdbx overexpression inhibits neuronal differentiation in the embryo and when co-injected with Xash3, Xdbx inhibits the ability of Xash3 to induce ectopic neurogenesis. One role of Xdbx during normal development may therefore be to restrict spatially neuronal differentiation within the neural plate, possibly by altering the neuronal differentiation function of Xash3. PMID- 10851139 TI - Mice lacking the transcriptional corepressor TIF1beta are defective in early postimplantation development. AB - TIF1beta, a member of the transcriptional intermediary factor 1 family, has been reported to function as a corepressor for the large class of KRAB domain containing zinc finger proteins of the Kruppel type. To address the biological function of TIF1beta, we have generated TIF1beta-deficient mice by gene disruption. TIF1beta protein was detected in wild-type but not TIF1beta(-/-) blastocysts. Homozygous mutant embryos, which developed normally until the blastocyst stage and underwent uterine implantation, were arrested in their development at the early egg-cylinder stage at about embryonic day (E) 5.5 and were completely resorbed by E8.5. Taken together, these results provide genetic evidence that TIF1beta is a developmental regulatory protein that exerts function(s) essential for early postimplantation development. PMID- 10851140 TI - Mechanisms regulating target gene selection by the homeodomain-containing protein Fushi tarazu. AB - Homeodomain proteins are DNA-binding transcription factors that control major developmental patterning events. Although DNA binding is mediated by the homeodomain, interactions with other transcription factors play an unusually important role in the selection and regulation of target genes. A major question in the field is whether these cofactor interactions select target genes by modulating DNA binding site specificity (selective binding model), transcriptional activity (activity regulation model) or both. A related issue is whether the number of target genes bound and regulated is a small or large percentage of genes in the genome. In this study, we have addressed these issues using a chimeric protein that contains the strong activation domain of the viral VP16 protein fused to the Drosophila homeodomain-containing protein Fushi tarazu (Ftz). We find that genes previously thought not to be direct targets of Ftz remain unaffected by FtzVP16. Addition of the VP16 activation domain to Ftz does, however, allow it to regulate previously identified target genes at times and in regions that Ftz alone cannot. It also changes Ftz into an activator of two genes that it normally represses. Taken together, the results suggest that Ftz binds and regulates a relatively limited number of target genes, and that cofactors affect target gene specificity primarily by controlling binding site selection. Activity regulation then fine-tunes the temporal and spatial domains of promoter responses, the magnitude of these responses, and whether they are positive or negative. PMID- 10851141 TI - Engineering radiation-resistant bacteria for environmental biotechnology. AB - Seventy million cubic meters of ground and three trillion liters of groundwater have been contaminated by leaking radioactive waste generated in the United States during the Cold War. A cleanup technology is being developed based on the radiation-resistant bacterium Deinococcus radiodurans, which is being engineered to express bioremediating functions. PMID- 10851142 TI - Engineering novel carotenoids in microorganisms. AB - A considerable number of microbial and plant carotenoid biosynthesis genes have been cloned over the past few years. Functional heterologous expression of most of these genes has made it possible to engineer carotenoid biosynthesis in non carotenogenic E. coli and yeasts. Recently, gene combination and molecular breeding of pathways have been used to produce novel and rare high-value carotenoids. PMID- 10851143 TI - Forest tree biotechnology. AB - The past year has seen the fruits of biotechnological manipulation of forest trees approach commercial application. Advances in somatic embryogenesis have brought mass clonal propagation of the top commercial trees closer to reality, and efficient gene transfer systems have been developed for a number of conifers and hardwoods. Radical alterations in the quantity and quality of lignin in wood have been shown to be possible in softwoods and hardwoods through identification of naturally occurring mutants, as well as by engineering the lignin biosynthetic pathway with transgenes. The potential environmental and social impacts of the release of transgenic trees have become an increasingly contentious issue that will require more attention if we are to use these technologies to their full advantage. PMID- 10851144 TI - Field applications of genetically engineered microorganisms for bioremediation processes. AB - Genetically engineered microorganisms (GEMs) have shown potential for bioremediation applications in soil, groundwater, and activated sludge environments, exhibiting enhanced degradative capabilities encompassing a wide range of chemical contaminants. However, the vast majority of studies pertaining to genetically engineered microbial bioremediation are supported by laboratory based experimental data. In general, relatively few examples of GEM applications in environmental ecosystems exist. Unfortunately, the only manner in which to fully address the competence of GEMs in bioremediation efforts is through long term field release studies. It is therefore essential that field studies be performed to acquire the requisite information for determining the overall effectiveness and risks associated with GEM introduction into natural ecosystems. PMID- 10851145 TI - Environmental biotechnology informatics. AB - Environmental biotechnology informatics is in its infancy and is growing fast. Computer and information science can assist environmental biotechnology by developing biological databases and building mathematical models of biological systems. Funding and training limitations in this field may, however, hinder its future growth. PMID- 10851146 TI - Aromatic hydrocarbon dioxygenases in environmental biotechnology. AB - Aromatic hydrocarbon dioxygenases belong to a large family of Rieske non-heme iron oxygenases. The dioxygenases have a broad substrate specificity and catalyze enantiospecific reactions with a wide range of substrates. These characteristics make them attractive synthons for the production of industrially and medically important chiral chemicals and also provide essential information for the development of bioremediation technology. PMID- 10851147 TI - Directed evolution of microbial oxidative enzymes. AB - In the past year, a number of oxidative enzymes have been the target of directed evolution. Catalase reaction specificity has been shifted to peroxidase, the high pH, thermal and oxidative stability of a fungal peroxidase has been dramatically improved, and the substrate specificity of cytochrome P450 has been altered to include substrates that the wild-type enzymes are incapable of oxidizing. PMID- 10851148 TI - Engineering bacteria for bioremediation. AB - The treatment of environmental pollution by microorganisms is a promising technology. Various genetic approaches have been developed and used to optimize the enzymes, metabolic pathways and organisms relevant for biodegradation. New information on the metabolic routes and bottlenecks of degradation is still accumulating, enlarging the available toolbox. With molecular methods allowing the characterization of microbial community structure and activities, the performance of microorganisms under in situ conditions and in concert with the indigenous microflora will become predictable. PMID- 10851149 TI - Biocontrol ability of fluorescent pseudomonads genetically dissected: importance of positive feedback regulation. AB - Root diseases caused by fungal pathogens can be suppressed by certain rhizobacteria that effectively colonize the roots and produce extracellular antifungal compounds. To be effective, biocontrol bacteria need to be present at sufficiently high cell densities. These conditions favor the operation of positive feedback mechanisms that control the production of antifungal compounds in biocontrol strains of fluorescent pseudomonads, via both transcriptional and post-transcriptional mechanisms. PMID- 10851150 TI - Bioremedial potential of microbial mechanisms of metal mobilization and immobilization. AB - Microorganisms play important roles in the environmental fate of toxic metals and radionuclides with a multiplicity of mechanisms effecting transformations between soluble and insoluble forms. These mechanisms are integral components of natural biogeochemical cycles and are of potential for both in situ and ex situ bioremedial treatment processes for solid and liquid wastes. PMID- 10851151 TI - Engineering dioxygenases for efficient degradation of environmental pollutants. AB - Dioxygenases have recently been engineered to improve their capabilities for environmental pollutant degradation. The techniques used to achieve this include in vitro DNA shuffling and subunit or domain exchanges between dioxygenases of different bacterial origins. Such evolved enzymes acquire novel and enhanced degradation capabilities of xenobiotic compounds, such as polychlorinated biphenyls, trichloroethylene and a variety of aromatic compounds. Hybrid strains in which the evolved genes are integrated into the chromosomal operons exhibit efficient degradation of xenobiotic chlorinated compounds. PMID- 10851152 TI - International Conference on Harmonization--critical discussion of the biotech 'Specifications' document. AB - Since 1990, the International Conference on Harmonization (ICH) has served as the primary medium for the development of consistent, harmonized and scientifically based international standards that keep abreast of the complexity of rapidly evolving technologies, and health, safety and commerce issues. Of the 45 guidance documents generated to date, influencing the conduct of drug development at various points during its continuum, six are dedicated to biotechnology. 'Specifications', the last in the series, was completed in 1999. It is fully complimentary to the other five guidance documents in the ICH biotechnology compendium. The process of establishing product specifications (principles and applications) is functionally couched within the multifaceted strategy of ensuring quality and consistency, and is proving to be a fundamental resource in crafting future harmonized documents such as the Common Technical Document. PMID- 10851153 TI - Bacterial dormancy and culturability: the role of autocrine growth factors. PMID- 10851154 TI - Novel forms of anaerobic respiration of environmental relevance. AB - Novel forms of anaerobic respiration continue to be discovered. Many of these are environmentally significant as they have important impacts on the fate of organic carbon and the cycling of many inorganic compounds. Furthermore, anaerobic respiration is becoming increasing recognized as a strategy for the remediation of organic and metal contaminants in the subsurface. PMID- 10851155 TI - Microecology of the termite gut: structure and function on a microscale. AB - Long considered simply as anoxic fermentors, termite guts are in fact axially and radially structured environments with physicochemically distinct microhabitats. Recent developments in termite gut microecology, which combined traditional and modern techniques, have focused on the spatial organization of important microbial populations and their in situ activities, and have significantly furthered our understanding of functional interactions within highly structured microenvironments. PMID- 10851156 TI - Microbial physiological state at low growth rate in natural and engineered ecosystems. AB - Microbes often grow in nature and bioreactors at very low growth rates. However, the physiological consequences at low growth rates have not been explored as completely as at faster growth rates or under starvation conditions. Nutrient flux to (and through) the cell surface and non-growth-dependent energy consumption (maintenance) are important considerations under these conditions. The biomass recycle reactor is a system to explore physiological state at low growth rate, and to optimize certain industrial process rates. PMID- 10851157 TI - Lactic acid bacteria: the bugs of the new millennium. AB - Lactic acid bacteria (LABs) are widely used in the manufacturing of fermented food and are among the best-studied microorganisms. Detailed knowledge of a number of physiological traits has opened new potential applications for these organisms in the food industry, while other traits might be beneficial for human health. Important new developments have been made in the research of LABs in the areas of multidrug resistance, bacteriocins and quorum sensing, osmoregulation, proteolysis, autolysins and bacteriophages. Recently, progress has been made in the construction of food-grade genetically modified LABs. PMID- 10851158 TI - Monitoring gene expression using DNA microarrays. AB - The concurrent development of high-density array technologies and the complete sequencing of a number of microbial genomes is providing the opportunity to comprehensively and efficiently survey the transcription profile of microorganisms under different conditions and well-defined genotypes. Microarray based studies are uncovering broad patterns of genetic activity, providing new understanding of gene functions and, in some cases, generating unexpected insight into transcriptional processes and biological mechanisms. One topic that has come to the forefront is how best to effectively manage and interpret the large data sets being generated. Although progress has been made, this remains a challenging opportunity for functional genomics research. PMID- 10851159 TI - Analysis of the microbial proteome. AB - Proteomics has begun to provide insight into the biology of microorganisms. The combination of proteomics with genetics, molecular biology, protein biochemistry and biophysics is particularly powerful, resulting in novel methods to analyse complex protein mixtures. Emerging proteomic technologies promise to increase the throughput of protein identifications from complex mixtures and allow for the quantification of protein expression levels. PMID- 10851160 TI - Bacterial endosymbionts in animals. AB - Molecular phylogenetic studies reveal that many endosymbioses between bacteria and invertebrate hosts result from ancient infections followed by strict vertical transmission within host lineages. Endosymbionts display a distinctive constellation of genetic properties including AT-biased base composition, accelerated sequence evolution, and, at least sometimes, small genome size; these features suggest increased genetic drift. Molecular genetic characterization also has revealed adaptive, host-beneficial traits such as amplification of genes underlying nutrient provision. PMID- 10851161 TI - Environmentally directed mutations and their impact on industrial biotransformation and fermentation processes. AB - Microbial adaptation plays an important role in the selection of improved strains for biotechnological processes and for the maintenance and stability of the selected production strains. Most of the knowledge about adaptation processes and environmentally directed mutations originates from environmental microbiology and from studies on biological evolution. The increasing information on the molecular mechanisms of adapted mutations and on the development of methods frequently used in environmental and evolutionary microbiology, such as the selection in semi continuous cultures or chemostats, can be used as input and tools for the improvement of industrial production organisms. PMID- 10851162 TI - Protein arrays for gene expression and molecular interaction screening. AB - The array format has revolutionised biomedical experimentation and diagnostics, enabling ordered high-throughput analysis. During the past decade, classic solid phase substrates, such as microtitre plates, membrane filters and microscopic slides, were turned into high-density, chip-like structures. The concept of the arrayed library was central to this development which now extends from DNA to protein. The new and versatile protein array technology allows high-throughput screening for gene expression and molecular interactions. As a major platform for functional genomics, it is already on its way into medical diagnostics. PMID- 10851163 TI - Systematic and large-scale two-hybrid screens. AB - The increasing rate at which complete genome sequences become available necessitates rapid and robust methods for investigating the functions of their encoded proteins. Efforts have been made to study protein function by systematically screening large sets of proteins using the two-hybrid method. Analyses of the complete proteomes of baceriophage T7, the mammalian viruses hepatitis C and vaccinia, as well as of several protein complexes including RNA splicing proteins and RNA polymerase III from yeast, have been undertaken. Saccharomyces cerevisiae has been studied extensively by two-hybrid methods, with more than 2500 protein-protein interactions described. Systematic studies on metazoan proteomes are, however, still in their infancy. PMID- 10851164 TI - Genome-wide mutant collections: toolboxes for functional genomics. AB - The sequencing of entire genomes has led to the identification of many genes. A future challenge will be to determine the function of all of the genes of an organism. One of the best ways to ascertain function is to disrupt genes and determine the phenotype of the resulting organism. Novel large-scale approaches for generating gene disruptions and analyzing the resulting phenotype are underway in the budding yeast Saccharomyces cerevisiae and other organisms including flies, Mycoplasma, worms, plants and mice. These approaches and mutant collections will be extremely valuable to the scientific community and will dramatically alter the manner in which science is performed in the future. PMID- 10851165 TI - Detoxification of reactive intermediates during microbial metabolism of halogenated compounds. AB - The reactivity and toxicity of metabolic intermediates that are generated by initial biotransformation reactions can be a major limiting factor for biodegradation of halogenated organic compounds. Recent work on the conversion of haloalkanes, chloroaromatics and chloroethenes indicates that microorganisms may become less sensitive to toxic effects either by using novel pathways that circumvent the generation of reactive intermediates or by producing modified enzymes that decrease the toxicity of such compounds. PMID- 10851166 TI - Long-distance DD-PCR and cDNA microarrays. AB - Analysis of differential gene expression is a classic tool in experimental biology. Broadly applicable new methods to identify and quantitative differential mRNA profiles, such as long distance differential display PCR and cDNA microarrays, promise to greatly accelerate understanding of mechanisms of development, differentiation, and disease. PMID- 10851167 TI - AMPA and NMDA receptors: similarities and differences in their synaptic distribution. AB - Recent technical developments, including antigen-retrieval and electron microscopic immunogold methods, are making it possible to determine some of the basic principles governing the subcellular distribution of ionotropic glutamate receptors. Distinct AMPA and NMDA receptor subtypes are selectively targeted to functionally different synapses of a single cell, resulting in an input-selective fine-tuning and regulation of the postsynaptic responses. The amount, density and variability of AMPA receptors at a given glutamatergic synapse is governed by both pre- and postsynaptic factors, resulting in functionally distinct glutamatergic connections that display characteristic patterns of receptor expression. PMID- 10851168 TI - Neurological diseases caused by ion-channel mutations. AB - During the past decade, mutations in several ion-channel genes have been shown to cause inherited neurological diseases. This is not surprising given the large number of different ion channels and their prominent role in signal processing. Biophysical studies of mutant ion channels in vitro allow detailed investigations of the basic mechanism underlying these 'channelopathies'. A full understanding of these diseases, however, requires knowing the roles these channels play in their cellular and systemic context. Differences in this context often cause different phenotypes in humans and mice. The situation is further complicated by the developmental effects and other secondary effects that might result from ion channel mutations. Recent studies have described the different thresholds to which ion-channel function must be decreased in order to cause disease. PMID- 10851169 TI - CaM-kinases: modulators of synaptic plasticity. AB - Calcium signaling is crucial for several aspects of plasticity at glutamatergic synapses, and studies over the past two to three years have identified key functions for Ca(2+)/calmodulin-dependent protein kinases II and IV (CaM-KII and CaM-KIV). Sustained activation of CaM-KII localized at the postsynaptic density results in phosphorylation of numerous synaptic substrates including ion channels, other signaling molecules and scaffolding proteins, to modulate synaptic transmission within minutes. More prolonged responses may be effected through enhanced dendritic protein synthesis of CaM-KII and regulation of nuclear gene transcription by CaM-KIV. PMID- 10851170 TI - Regulation of transmitter release by Unc-13 and its homologues. AB - Neurotransmitters are released by Ca(2+)-triggered exocytotic fusion of synaptic vesicles. Before fusion, vesicles dock at a specialised presynaptic plasma membrane region, the active zone, where they are primed to a fusion competent state. The nature of this priming reaction has long been enigmatic. Recent evidence demonstrates that priming is an essential and rate-limiting step in secretion from neurons and neuroendocrine cells. Members of the Unc-13 protein family, which are highly conserved during evolution and act as novel targets of the diacylglycerol second-messenger pathway, have been identified to play an essential role in this process. PMID- 10851171 TI - Hebb and homeostasis in neuronal plasticity. AB - The positive-feedback nature of Hebbian plasticity can destabilize the properties of neuronal networks. Recent work has demonstrated that this destabilizing influence is counteracted by a number of homeostatic plasticity mechanisms that stabilize neuronal activity. Such mechanisms include global changes in synaptic strengths, changes in neuronal excitability, and the regulation of synapse number. These recent studies suggest that Hebbian and homeostatic plasticity often target the same molecular substrates, and have opposing effects on synaptic or neuronal properties. These advances significantly broaden our framework for understanding the effects of activity on synaptic function and neuronal excitability. PMID- 10851172 TI - Neurotrophin signal transduction in the nervous system. AB - Neurotrophins use two types of receptors, the Trk tyrosine kinase receptors and the p75 neurotrophin receptor (p75NTR), to regulate the growth, development, survival and repair of the nervous system. These receptors can either collaborate with or inhibit each other's actions to mediate neurotrophin effects. The development and survival of neurons is thus based upon the functional interplay of the signals generated by Trk and p75NTR. In the past two years, the signaling pathways used by these receptors, including Akt and MAPK-induced signaling via Trk, and JNK, p53, and NF-kappaB signaling via p75NTR, have been identified. In addition, a number of novel p75NTR-interacting proteins have been identified that transmit growth, survival, and apoptotic signals. PMID- 10851173 TI - Molecular determinants of presynaptic active zones. AB - The presynaptic cytoskeletal matrix (cytomatrix) assembled at active zones has been implicated in defining neurotransmitter release sites. Munc13, Rim, Bassoon and Piccolo/Aczonin are recently identified presynaptic cytomatrix proteins. These multidomain proteins are thought to organize the exocytotic and endocytotic machinery precisely at active zones. PMID- 10851174 TI - Synaptic kainate receptors. AB - Kainate receptors are a family of ionotropic glutamate receptors with poorly understood functions. Recent evidence firmly establishes kainate receptors as postsynaptic mediators of synaptic transmission. A second, presynaptic, modulatory role of kainate receptors has also been suggested, although the mechanism(s) involved remain controversial. PMID- 10851175 TI - Control of cell behaviour by signalling through Eph receptors and ephrins. AB - Eph receptor tyrosine kinases and ephrins mediate contact-dependent cell interactions that regulate the repulsion and adhesion mechanisms involved in the guidance and assembly of cells. Recent work has revealed a role of overlapping Eph receptor and ephrin expression in modulating neuronal growth cone repulsion, and has shown that bidirectional activation restricts intermingling and communication between cell populations. In addition, progress has been made in understanding how Eph receptors and ephrins control cell adhesion. PMID- 10851176 TI - Downregulation of G protein-coupled receptors. AB - Major advances have been made in understanding mechanisms mediating downregulation of G protein-coupled receptors. Recent studies emphasize the role of multiple proteolytic mechanisms in downregulation. A specific mechanism of downregulation, mediated by endocytosis of receptors via clathrin-coated pits followed by sorting to lysosomes, has been examined in detail. Specific protein interactions that control the specificity of G-protein-coupled receptor trafficking in this pathway are beginning to be elucidated. PMID- 10851177 TI - Sequential steps in clathrin-mediated synaptic vesicle endocytosis. AB - Synaptic vesicles are recycled with remarkable speed and precision in nerve terminals. A major recycling pathway involves clathrin-mediated endocytosis at endocytic zones located around sites of release. Different 'accessory' proteins linked to this pathway have been shown to alter the shape and composition of lipid membranes, to modify membrane-coat protein interactions, and to influence actin polymerization. These include the GTPase dynamin, the lysophosphatidic acid acyl transferase endophilin, and the phosphoinositide phosphatase synaptojanin. Protein perturbation studies in living nerve terminals are now beginning to link the actions of these proteins with morphologically defined steps of endocytosis. PMID- 10851178 TI - Structural insights into the molecular mechanism of Ca(2+)-dependent exocytosis. AB - The fusion of vesicles with target membranes is controlled by a complex network of protein-protein and protein-lipid interactions. Recent structures of the SNARE complex, synaptotagmin III, nSec1, domains of NSF and its adaptor SNAP, along with Rab3 and some of its effectors, provide the framework for developing molecular models of vesicle fusion and for designing experiments to test these models. Ultimately, this knowledge of the structures of higher-order complexes and their dynamic behavior will allow us to obtain a full understanding of the vesicle fusion protein machinery. PMID- 10851179 TI - LTP mechanisms: from silence to four-lane traffic. AB - Brief periods of strong neuronal activity induce long-lasting changes in synaptic function. This synaptic plasticity is thought to play important roles in learning and memory. One example--long-term potentation in the CA1 region of the hippocampus--has been studied extensively, and conflicting views regarding the underlying mechanisms have emerged. Recent findings, regarding basic properties of synaptic transmission, appear to reconcile these diverging views. PMID- 10851180 TI - Roles of Wnt proteins in neural development and maintenance. AB - Many constituents of Wnt signaling pathways are expressed in the developing and mature nervous systems. Recent work has shown that Wnt signaling controls initial formation of the neural plate and many subsequent patterning decisions in the embryonic nervous system, including formation of the neural crest. Wnt signaling continues to be important at later stages of development. Wnts have been shown to regulate the anatomy of the neuronal cytoskeleton and the differentiation of synapses in the cerebellum. Wnt signaling has been demonstrated to regulate apoptosis and may participate in degenerative processes leading to cell death in the aging brain. PMID- 10851181 TI - Recent advances in technology for measuring and manipulating cell signals. AB - Signal transduction research has made some glowing progress in the past 12 months. Recent advances in fluorescent proteins, small molecule fluorophores and imaging technology are generating new ways to investigate signal transduction. PMID- 10851182 TI - Biogenic amine transporters: regulation in flux. AB - Following vesicular release, the biogenic amine neurotransmitters dopamine, norepinephrine and serotonin are actively cleared from extracellular spaces by presynaptic transporters. These transporters interact with multiple psychoactive agents including cocaine, amphetamines and antidepressants. Recent findings indicate that amine reuptake is likely to be a tightly regulated component of synaptic plasticity rather than a constitutive determinant of transmitter clearance. Protein kinase C activation and transporter phosphorylation have been linked to regulatory protein trafficking, and both phosphorylation and trafficking may be influenced by transporter ligands. Recognition that transmitters, antagonists and second messengers can modify the intrinsic activity, surface expression or protein levels of amine transporters raises new questions about the fundamental nature of drug actions in vivo. The theory that dysregulation of transporters may contribute to disease states is supported by the recent discovery that a coding mutation in the human norepinephrine transporter contributes to orthostatic intolerance. PMID- 10851183 TI - Homer: a link between neural activity and glutamate receptor function. AB - The proteins of the Homer family bind to proline-rich sequences in group I metabotropic glutamate receptors, inositol trisphosphate receptors, ryanodine receptors, and Shank family proteins. Homer proteins also self associate and function as adaptors to couple interacting proteins. Recent observations indicate a role for Homer complexes in signal transduction, synaptogenesis and receptor trafficking. PMID- 10851184 TI - Computational genetics: finding protein function by nonhomology methods. AB - During the past year, computational methods have been developed that use the rapidly accumulating genomic data to discover protein function. The methods rely on properties shared by functionally related proteins other than sequence or structural similarity. Instead, these 'nonhomology' methods analyze patterns such as domain fusion, conserved gene position and gene co-inheritance and coexpression to identify protein-protein relationships. The methods can identify functions for proteins that are without characterized homologs and have been applied to genome-wide predictions of protein function. PMID- 10851185 TI - Protein structure prediction in the postgenomic era. AB - As the number of completely sequenced genomes rapidly increases, the postgenomic problem of gene function identification becomes ever more pressing. Predicting the structures of proteins encoded by genes of interest is one possible means to glean subtle clues as to the functions of these proteins. There are limitations to this approach to gene identification and a survey of the expected reliability of different protein structure prediction techniques has been undertaken. PMID- 10851186 TI - Genome data: what do we learn? AB - Genome sequence information has continued to accumulate at a spectacular pace during the past year. Details of the sequence and gene content of human chromosome 22 were published. The sequencing and annotation of the first two Arabidopsis thaliana chromosomes was completed. The sequence of chromosome 3 from Plasmodium falciparum, the second sequenced malaria chromosome, was reported, as was that of chromosome 1 from Leishmania major. The complete genomic sequences of five microbes were reported. Approaches to using data from completely sequenced microbial genomes in phylogenetic studies are being explored, as is the application of microarrays to whole genome expression analysis. PMID- 10851187 TI - Annotating eukaryote genomes. AB - The Genome Annotation Assessment Project tested current methods of gene identification, including a critical assessment of the accuracy of different methods. Two new databases have provided new resources for gene annotation: these are the InterPro database of protein domains and motifs, and the Gene Ontology database for terms that describe the molecular functions and biological roles of gene products. Efforts in genome annotation are most often based upon advances in computer systems that are specifically designed to deal with the tremendous amounts of data being generated by current sequencing projects. These efforts in analysis are being linked to new ways of visualizing computationally annotated genomes. PMID- 10851188 TI - The nature of the universal ancestor and the evolution of the proteome. AB - The past year has seen several attempts to reconstruct the proteome of the universal ancestor of all life on the basis of comparisons of contempory genomes. However, increasing evidence for lateral gene transfer could mean that such attempts are based on an incorrect understanding of evolution. PMID- 10851189 TI - NMR solution structure determination of RNAs. AB - During the past several years, there have been significant advances in NMR solution structure determination of macromolecules. The ability to easily measure residual dipolar couplings, to directly detect NHellipsisN hydrogen bonding interactions and to study much larger macromolecules by the application of heteronuclear experiments that select narrow lines in 2D and 3D spectra of isotopically labeled molecules promises to dramatically improve solution structure determination of nucleic acids. PMID- 10851190 TI - Simulation of the structure and dynamics of nonhelical RNA motifs. AB - Computer simulation methods are increasingly being used to study possible conformations and dynamics of structural motifs in RNA. Recent results of molecular dynamics simulations and continum solvent studies of RNA structures and RNA-ligand complexes show promising agreement with experimental data. Combined with the ongoing progress in the experimental characterization of RNA structure and thermodynamics, these computational approaches can help to better understand the mechanism of RNA structure formation and the binding of ligands. PMID- 10851191 TI - From fold to function. AB - A number of recent advances have been made in deriving function information from protein structure. A fold relationship to an already characterized protein will often allow general information about function to be deduced. More detailed information can be obtained using sequence relationships to already studied proteins. Methods of deducing function directly from structure, without the use of evolutionary relationships, are developing rapidly. All such methods may be used with models of protein structure, rather than with experimentally determined ones, but model accuracy imposes limitations. The rapid expansion of the structural genomics field has created a new urgency for improved methods of structure-based annotation of function. PMID- 10851192 TI - Calculating nucleic acid secondary structure. AB - New results for calculating nucleic acid secondary structure by free energy minimization and phylogenetic comparisons have recently been reported. A complete set of DNA energy parameters is now available and the RNA parameters have been improved. Although databases of RNA secondary structures are still derived and expanded using computer-assisted, ad hoc comparative analysis, a number of new computer algorithms combine covariation analysis with energy methods. PMID- 10851193 TI - Fitting peptides into the RNA world. AB - The structures of several peptide-RNA complexes have been reported in the past year, underscoring the diverse nature of RNA structure and protein interactions. In general, specific peptide conformations are stabilized by the surrounding RNA framework; this is strikingly similar to how peptides are stabilized upon interaction with proteins. PMID- 10851194 TI - Exploitation of gene context. AB - Recently, a number of techniques have been proposed that use completely sequenced genomes for the function prediction of individual proteins encoded therein. They use the fusion of genes, their conserved location in operons or merely their co occurrence in genomes to predict the existence of functional interactions between the proteins they encode. This type of information complements functional features that are predicted by classical homology-based search techniques. PMID- 10851195 TI - The i-motif in nucleic acids. AB - Seven years after the discovery of the DNA i-motif, partial explanations for its occurrence have been uncovered, possibly involving CHellipsisO hydrogen bonds across the narrow grooves. Investigations of its biological significance have been encouraged by the demonstration and description of the intramolecular i motif structure of human telomeric and centromeric sequences, by the recent observation of an intercalated RNA structure and by the discovery of proteins that associate with DNA sequences carrying cytosine repeats. The compatibility of the intercalation with peptide and phosphorothioate DNA analogs is favorable for possible pharmaceutical applications. PMID- 10851196 TI - Allosteric nucleic acid catalysts. AB - Endowing nucleic acid catalysts with allosteric properties provides new prospects for RNA and DNA as controllable therapeutic agents or as sensors of their cognate effector compounds. The ability to engineer RNA catalysts that are allosterically regulated by effector binding has been propelled by the union of modular rational design principles and powerful combinatorial strategies. PMID- 10851197 TI - Single-molecule studies of DNA mechanics. AB - During the past decade, physical techniques such as optical tweezers and atomic force microscopy were used to study the mechanical properties of DNA at the single-molecule level. Knowledge of DNA's stretching and twisting properties now permits these single-molecule techniques to be used in the study of biological processes such as DNA replication and transcription. PMID- 10851198 TI - Structural genomics of microbes: an objective. AB - A comparison of the genome sequences of more than 20 microorganisms reveals that a large fraction of the genes have unknown functions. Determining the structures of the proteins coded by these genes may provide additional key information in an effort to uncover the molecular functions of such proteins and new protein fold patterns. Using existing technology, it is possible to obtain a complete sequence complement and a near complete structural complement for a small microbial genome. Such information may provide a comprehensive view of a small organism, which, in turn, can serve as a platform for understanding more complex organisms. PMID- 10851200 TI - June 13, 2000 PMID- 10851199 TI - Modeling DNA deformations. AB - Recent developments have been made in modeling double-helical DNA at four levels of three-dimensional structure: the all-atom level, whereby an oligonucleotide duplex is surrounded by a shroud of solvent molecules; the base-pair level, with explicit backbone atoms; the mesoscopic level, that is, a few hundred base pairs, with the local duplex conformation described by knowledge-based harmonic energy functions; and the scale of several thousand nucleotides, with the duplex described as an ideal elastic rod. Predictions of the sequence-dependent bending and twisting of the double helix, as well as solvent- and force-induced B-->A and over-stretching conformational transitions, are compared with experimental data. These subtle conformational changes are critical to the functioning of the double helix, including its packaging in the close confines of the cell, the mutual fit of DNA and protein in nucleoprotein complexes, and the effective recognition of base pairs in recombination and transcription. PMID- 10851201 TI - Cytoskeletal abnormalities in the failing heart: out on a LIM? PMID- 10851202 TI - Decreased expression of the cardiac LIM domain protein MLP in chronic human heart failure. AB - BACKGROUND: The cardiac LIM domain protein MLP, a member of the cysteine-rich protein family, is an essential regulator of cardiac muscle development. Mice with a disruption of the MLP gene resemble the morphological and clinical picture of dilated cardiomyopathy and heart failure in humans. We investigated whether altered MLP expression is significant for the pathogenesis of human heart failure. METHODS AND RESULTS: Immunohistochemistry and in situ hybridization confirmed the expression of MLP protein and mRNA in human cardiomyocytes. Western blot analysis revealed that the MLP peptide was present in the contractile protein fraction but not in the cytosolic or membrane fraction and that the binding of MLP to myofibrils required functional zinc finger domains. MLP immunoreactivity was decreased approximately 50% (P<0.05) in the left ventricular myocardium of patients with chronic heart failure due to dilated or ischemic cardiomyopathy compared with non-failing donor hearts. MLP mRNA expression, as assessed by Northern blot experiments, was not significantly different between failing and non-failing control hearts, which suggests that decreased MLP synthesis or increased MLP protein turnover, rather than a decreased number of RNA transcripts, may play a role. CONCLUSIONS: Because MLP may promote myofibril assembly, the down-regulation of this adapter protein might play an essential role in myofibril derangement or impaired myofibril rearrangement in the failing human myocardium. PMID- 10851203 TI - Regulation of angiotensin II receptor subtypes during atrial fibrillation in humans. AB - BACKGROUND: Previous studies have suggested that atrial fibrillation (AF) is associated with the activation of the atrial angiotensin system. However, it is not known whether the expression of angiotensin II receptors changes during AF. The purpose of this study was to determine the atrial expression of angiotensin II type 1 and type 2 receptors (AT(1)-R and AT(2)-R) in patients with AF. METHODS AND RESULTS: Atrial tissue samples from 30 patients undergoing open heart surgery were examined. Eleven patients had chronic persistent AF (> or =6 months; cAF), 8 patients had paroxysmal AF (pAF), and 11 patients were in sinus rhythm. AT(1)-R and AT(2)-R were localized in the atrial tissue by immunohistochemistry and quantified at the protein and mRNA level by Western blotting and quantitative polymerase chain reaction. Both types of AT-R were predominantly expressed in atrial myocytes in all groups. The amount of AT(1)-R was reduced to 34.9% during cAF (P<0.01) and to 51.7% during pAF (P<0.05) compared with patients in sinus rhythm. In contrast, AT(2)-R was increased during cAF (246%; P=NS) and pAF (505%; P<0.01). AT(1)-R/AT(2)-R mRNA content was similar in all groups. CONCLUSIONS: AF is associated with the down-regulation of atrial AT(1)-R and the up-regulation of AT(2)-R proteins. These findings may help define the pathophysiological role of the angiotensin system in the structural remodeling of the fibrillating atria. PMID- 10851204 TI - Predictors of mortality and mortality from cardiac causes in the bypass angioplasty revascularization investigation (BARI) randomized trial and registry. For the BARI Investigators. AB - BACKGROUND: The impact of percutaneous transluminal coronary angioplasty (PTCA) and coronary artery bypass grafting (CABG) on long-term mortality rates in the presence of various demographic, clinical, and angiographic factors is uncertain in the population of patients suitable for both procedures. METHODS AND RESULTS: In the Bypass Angioplasty Revascularization Investigation (BARI) randomized trial and registry, 3610 patients who were eligible to receive PTCA and CABG were revascularized between 1989 and 1992. Multivariate Cox models were used to identify factors associated with 5-year mortality and cardiac mortality, with particular attention to factors that interact with treatment. Diabetic patients receiving insulin had higher mortality and cardiac mortality rates with PTCA compared with CABG (relative risk [RR] 1.78 and 2.63, respectively, P<0.001), and patients with ST elevation had higher cardiac mortality rates with CABG than with PTCA (RR 4.08, P<0.001). Factors most strongly associated with high overall mortality rates were insulin-treated diabetes, congestive heart failure, kidney failure, and older age. Black race was also associated with higher mortality rates (RR 1.49, P=0.019). CONCLUSIONS: A set of variables was identified that could be used to help select a revascularization procedure and to evaluate risk of long-term mortality in the population of patients considering revascularization. PMID- 10851205 TI - High levels of platelet inhibition with abciximab despite heightened platelet activation and aggregation during thrombolysis for acute myocardial infarction: results from TIMI (thrombolysis in myocardial infarction) 14. AB - BACKGROUND: We evaluated platelet activation and aggregation in patients with acute myocardial infarction (AMI) treated with thrombolytic therapy alone or with reduced-dose thrombolysis and concomitant abciximab. METHODS AND RESULTS: The study was performed in 20 control subjects and 51 patients with AMI before and after reperfusion with either alteplase or reteplase or reduced doses of these agents with concomitant abciximab. Platelet activation was assayed by platelet surface expression of P-selectin. Turbidometric platelet aggregation in response to ADP was measured in patients before thrombolytic therapy and 90 minutes and 24 hours after the beginning of thrombolytic therapy. P-selectin expression was greater at baseline in patients than normal control subjects (30.4% versus 9. 8%, P<0.0001) but was identical between the 2 groups after stimulation with ADP (64.4% versus 69.3%, P=0.37). However, at 24 hours, basal P-selectin expression declined in patients (P=0.0025 versus baseline), whereas ADP-stimulated P selectin expression was lower in patients than in control subjects (48% versus 69%, P=0. 0004). When combined with reduced doses of either alteplase or reteplase, abciximab achieved 91% and 83% inhibition of 5 and 20 micromol/L ADP induced platelet aggregation, which decreased to 46% and 40%, respectively, at 24 hours. No appreciable difference in the platelet inhibition profile of abciximab was observed between the 2 thrombolytics. CONCLUSIONS: Platelet activation and aggregation are heightened in the setting of thrombolysis for AMI. Despite this enhanced level of platelet activation, abciximab, combined with a reduced-dose thrombolytic, inhibited platelet aggregation similarly to the level reported in elective settings. PMID- 10851206 TI - Magnetic resonance-based assessment of global coronary flow and flow reserve and its relation to left ventricular functional parameters: a comparison with positron emission tomography. AB - BACKGROUND: Measurement of coronary sinus blood flow (CSF) by phase-contrast magnetic resonance (PC-MR) imaging at rest and during hyperemia may allow noninvasive assessment of global coronary hemodynamics. METHODS AND RESULTS: Sixteen healthy volunteers (age, 22 to 32 years) were examined with MR and PET in random order within 1 to 2 days. At rest and during hyperemia (dipyridamole 0.56 mg/kg), CSF was measured by a cine PC-MR technique (temporal resolution, 40 ms; spatial resolution, 1.25x0.8 mm(2)), and myocardial blood flow (MBF) was measured by [(13)N]NH(3) PET. PET and MR agreed closely for coronary flow reserve (CFR; mean difference, 2.2+/-14.7%; Bland-Altman method). CSF divided by either total left ventricular mass or an estimate of drained myocardium (LVM(drain)) correlated highly with PET flow data (r=0.93 and 0.95, respectively) and with measures of oxygen demand, ie, heart rate, afterload-corrected fiber shortening, and peak systolic stress determined by MR (overall correlation coefficients, 0.81 and 0.87, respectively, multivariate analysis). CSF/LVM(drain) did not differ significantly from PET-derived MBF (difference, 3.6+/-16.6%). In orthotopic heart transplant recipients (n=9), CFR was reduced and blood supply-demand relationships at rest were shifted toward higher flows (P<0.0001). CONCLUSIONS: This integrated MR approach allows comprehensive assessment of autoregulated and hyperemic coronary flow and is suitable for serial measurements in patients. In transplanted hearts, elevated resting flow is the major cause of reduced CFR. PMID- 10851207 TI - Predictors of systolic augmentation from left ventricular preexcitation in patients with dilated cardiomyopathy and intraventricular conduction delay. AB - BACKGROUND: VDD pacing can enhance systolic function in patients with dilated cardiomyopathy and discoordinate contraction; however, identification of patients likely to benefit is unclear. We tested predictors of systolic responsiveness on the basis of global parameters as well as directly assessed mechanical dyssynchrony. METHODS AND RESULTS: Twenty-two DCM patients with conduction delay were studied by cardiac catheterization with a dual-sensor micromanometer to measure LV and aortic pressures during sinus rhythm and LV free-wall pacing. Pacing enhanced isovolumetric (dP/dt(max)) and ejection-phase (pulse pressure, PP) systolic function by 35+/-21% and 16.4+/-11%, respectively, and these changes correlated directly (r=0.7, P=0.001). %DeltadP/dt(max) was weakly predicted by baseline QRS (r=0.6, P<0.02), more strongly by baseline dP/dt(max) (r=0.7, P=0.001), and best by bidiscriminate analysis combining baseline dP/dt(max) < or =700 mm Hg/s and QRS > or =155 ms to predict %DeltadP/dt(max) > or =25% and %DeltaPP > or =10% (P<0.0005, chi(2)), with no false-positives. Benefit could not be predicted by %DeltaQRS. To test whether basal mechanical dyssynchrony predicted responsiveness to LV pacing, circumferential strains were determined at approximately 80 sites throughout the LV by tagged MRI in 8 DCM patients and 7 additional control subjects. Strain variance at time of maximal shortening indexed dyssynchrony, averaging 28.0+/-7.1% in normal subjects versus 201.4+/ 84.3% in DCM patients (P=0.001). Mechanical dyssynchrony also correlated directly with %DeltadP/dt(max) (r=0.85, P=0.008). Conclusions-These results show that although mechanical dyssynchrony is a key predictor for pacing efficacy in DCM patients with conduction delay, combining information about QRS and basal dP/dt(max) provides an excellent tool to identify maximal responders. PMID- 10851208 TI - Sympathetically mediated hypertension in autonomic failure. AB - BACKGROUND: Approximately 50% of patients with primary autonomic failure have supine hypertension. We investigated whether this supine hypertension could be driven by residual sympathetic activity. METHODS AND RESULTS: In patients with multiple system atrophy (MSA) or pure autonomic failure (PAF), we studied the effect of oral yohimbine on seated systolic blood pressure (SBP), the effect of ganglionic blockade (with trimethaphan) on supine SBP and plasma catecholamine levels, and the effect of alpha(1)-adrenoreceptor blockade (phentolamine) on supine SBP. The SBP response to yohimbine was greater in patients with MSA than in those with PAF (area under the curve, 2248+/-543 versus 467+/-209 mm Hg. min; P=0.022). MSA patients with a higher supine SBP had a greater response than those with a lower supine SBP (3874+/-809 versus 785+/-189 mm Hg. min; P=0. 0017); this relationship was not seen in PAF patients. MSA patients had a marked depressor response to low infusion rates of trimethaphan; the response in PAF patients was more variable. Plasma norepinephrine decreased in both groups, but heart rate did not change in either group. At 1 mg/min, trimethaphan decreased supine SBP by 67+/-8 and 12+/-6 mm Hg in MSA and PAF patients, respectively (P<0.0001). Cardiac index and total peripheral resistance decreased in MSA patients by 33.4+/-5.8% and 40.7+/-9.5%, respectively (P=0. 0015). Patients having a depressor response to trimethaphan also had a depressor response to phentolamine. In MSA patients, the pressor response to yohimbine and the decrease in SBP with 1 mg/min trimethaphan were correlated (r=0.98; P=0.001). CONCLUSIONS: Residual sympathetic activity drives supine hypertension in MSA. It contributes to, but does not completely explain, supine hypertension in PAF. PMID- 10851209 TI - Sympathetic denervation of the upper limb improves forearm exercise performance and skeletal muscle bioenergetics. AB - BACKGROUND: Sympathetic activation may limit exercise performance by restraining muscle blood flow or by negatively affecting skeletal muscle metabolic behavior. To test this hypothesis, we studied the effect of thoracoscopic sympathetic trunkotomy (TST) on forearm exercise duration, blood flow, and muscle bioenergetics in 13 patients with idiopathic palmar hyperhidrosis. METHODS AND RESULTS: Heart rate and beat-by-beat mean arterial pressure were recorded at rest and during right and left rhythmic handgrip before and 4 to 7 weeks after right TST. Forearm blood flow was measured bilaterally at rest and on the right during exercise. Right forearm muscle phosphocreatine content and intracellular pH were assessed by (31)phosphorus magnetic resonance spectroscopy. After right TST, exercise duration increased from 8.9+/-1.4 to 13.4+/-1.8 minutes (P<0.0001) with the right forearm and from 5.7+/-0.4 to 7.6+/-0.9 minutes (P<0.05) with the left (P<0.05 for the interaction between treatment and side). Right forearm blood flow at rest was 66% higher (P<0.01) after right TST, but this difference decreased as the exercise progressed. After right TST, a significant reduction occurred in muscle acidification and phosphocreatine depletion during ipsilateral forearm exercise. This was associated with a significantly reduced mean arterial pressure response to right handgrip, whereas the pressor response to left handgrip did not change. CONCLUSIONS: Sympathetic denervation of the upper limb significantly improves forearm skeletal muscle bioenergetics and exercise performance in patients with idiopathic palmar hyperhidrosis. PMID- 10851210 TI - Sotalol in the treatment of fetal dysrhythmias. AB - BACKGROUND: Fetal tachycardia may cause hydrops fetalis and lead to fetal death. No unanimity of opinion exists regarding the optimum treatment. This study evaluates our experience with transplacental sotalol therapy to treat fetal tachycardias in terms of safety and efficacy. METHODS AND RESULTS: The charts of 21 patients who were treated with sotalol for fetal tachycardia were reviewed. Ten fetuses had atrial flutter (AF), 10 had supraventricular tachycardia (SVT), and 1 had VT. Hydrops fetalis was present in 9 fetuses. Drug treatment was successful in establishing sinus rhythm in 8 of 10 fetuses with AF and in 6 of 10 fetuses with SVT. The mortality rate in this study was 19% (4 of 21 fetuses; 3 had SVT and 1 had AF); 3 deaths occurred just days after the initiation of sotalol therapy, and 1 occurred after a dosage increase. At birth, tachycardia was present in 6 infants. Two patients who converted to sinus rhythm in utero suffered from neurologic pathology postnatally. CONCLUSIONS: Fetal tachycardia is a serious condition in which treatment should be initiated, especially in the presence of hydrops fetalis. The high success rate in fetuses with AF suggests that sotalol should be considered a drug of first choice to treat fetal AF. The low conversion rate and the fact that 3 of the 4 deaths in this study occurred in fetuses with SVT indicate that the risks of sotalol therapy outweigh the benefits in this group and that sotalol should, therefore, be limited in the treatment of fetal SVT. PMID- 10851211 TI - Sympathetic reinnervation of the sinus node and exercise hemodynamics after cardiac transplantation. AB - BACKGROUND: Sympathetic cardiac reinnervation occurs variably after cardiac transplantation (CT) in humans. We hypothesized that sinus node reinnervation would partially restore normal chronotropic response to exercise. METHODS AND RESULTS: Thirteen recent CT recipients, 28 late CT recipients (> or =1 year after CT), and 20 control subjects were studied. Sinus node sympathetic reinnervation was determined by heart rate (HR) change after tyramine injection into the artery that perfused the sinus node. HR changes of <5 and > or =15 bpm were defined, respectively, as denervation and marked reinnervation. During treadmill exercise, HR, blood pressure, and expired O(2) and CO(2) were measured. All early transplant recipients exhibited features typical of denervation (basal HR, 88+/-2 bpm; peak HR, 132+/-4 bpm, peaking 1.8+/-0.3 minutes after exercise cessation and slowly declining after exercise). A similar pattern was found in the 12 late transplant recipients with persistent sinus node denervation. However, in patients with marked reinnervation, exercise HR rose more (peak HR, 142+/-4 and 141+/-2 bpm), peaked earlier after cessation of exercise (0.7+/-0.4 and 0. 3+/ 0.1 minute), and fell more rapidly. Exercise duration and maximal oxygen consumption were not related significantly to reinnervation status, but a trend existed for longer exercise time in markedly reinnervated patients. CONCLUSIONS: The present studies suggest that sympathetic reinnervation of the sinus node is accompanied by partial restoration of normal HR response to exercise. Both maximal oxygen consumption and exercise duration were markedly shorter in CT patients than in control subjects, and most of the difference was not related to innervation status. PMID- 10851212 TI - Microvascular obstruction and left ventricular remodeling early after acute myocardial infarction. AB - BACKGROUND: The presence of microvascular obstruction (MO) within infarcted regions may adversely influence left ventricular (LV) remodeling after acute myocardial infarction. This study examined whether the extent of MO directly alters the mechanical properties of the infarcted myocardium. METHODS AND RESULTS: Seventeen dogs underwent 90 minutes of balloon occlusion of the left anterior descending coronary artery, followed by reperfusion. Gadolinium-enhanced perfusion MRI and 3D-tagging were performed 4 to 6 and 48 hours (8 animals) and 10 days (9 animals) after reperfusion. Early increase in LV end-diastolic volume (from 42+/-9 to 54+/-14 mL, P<0.05) between 4 to 6 and 48 hours after reperfusion was predicted by both extent of MO (r=0.89, P<0.01) and infarct size (r=0.83, P<0.01), defined as MRI hypoenhanced and hyperenhanced regions, respectively. Multivariate analysis demonstrated that extent of MO had better and independent value to predict LV volume than overall infarct size. A strong inverse relationship existed between magnitude of first principal strain (r=-0.80, P<0.001) and relative extent of MO within infarcted myocardium. Also, infarcted myocardium involved by extensive areas of MO demonstrated reductions of circumferential (r=-0.61, P<0.01) and longitudinal (r=-0.53, P<0. 05) stretching. Furthermore, significant reductions of radial thickening (9+/-6% versus 14+/-3%, P<0.01) occurred in noninfarcted regions adjacent to infarcts that had increased (>35%) amounts of MO. CONCLUSIONS: In the early healing phase of acute myocardial infarction, the extent of MO in infarcted tissue relates to reduced local myocardial deformation and dysfunction of noninfarcted adjacent myocardium. Such strain alterations might explain the increased remodeling observed in patients with large regions of MO. PMID- 10851213 TI - Attenuation of myocardial ischemia/reperfusion injury by superinduction of inducible nitric oxide synthase. AB - BACKGROUND: Nitric oxide (NO) has been implicated as a mediator in myocardial ischemia/reperfusion (I/R) injury, but its functional properties have been conflicting. We investigated whether NO has a protective role against I/R injury. METHODS AND RESULTS: Using endothelial NO synthase knockout (eNOS KO) mice, inducible NOS KO mice, the NO donor S-nitroso-N-acetylpenicillamine (SNAP), and the NOS inhibitor N-iminoethyl-L-ornithine (L-NIO), we performed studies of isolated perfused hearts subjected to 30 minutes of global ischemia followed by reperfusion. After 60 minutes of reperfusion, nitrite levels in the coronary effluent in the SNAP and eNOS KO groups were significantly elevated compared with other groups. Immunoblot and immunohistochemistry showed that iNOS was markedly induced in the eNOS KO hearts. Under spontaneous beating conditions during reperfusion, increased NO activity was correlated with a prevention of the hyperdynamic contractile response and enhanced myocardial protection, as evidenced by a reduction in myocardial injury and infarct size. During prolonged reperfusion, SNAP-treated hearts were able to preserve contractile functions for 180 minutes, whereas L-NIO-treated hearts showed a sustained deterioration in contractility. CONCLUSIONS: NO protects against I/R injury by preventing the hyperdynamic response of isolated perfused hearts during early reperfusion. In the eNOS KO hearts, a paradoxical increase in NO production was seen, accompanied by a superinduction of iNOS, possibly due to an adaptive mechanism. PMID- 10851214 TI - Na(+)/H(+) exchange inhibition with HOE642 improves postischemic recovery due to attenuation of Ca(2+) overload and prolonged acidosis on reperfusion. AB - BACKGROUND: Na(+)/H(+) exchange inhibition with HOE642 (cariporide) improves postischemic recovery of cardiac function, but the mechanisms of action remain speculative. Because Na(+)/H(+) exchange is activated on reperfusion, it was hypothesized that its inhibition delays realkalinization and decreases intracellular Na(+) and, via Na(+)/Ca(2+) exchange, Ca(2+) overload. Attenuated Ca(2+) overload and prolonged acidosis are known to be cardioprotective. METHODS AND RESULTS: Left ventricular developed and end-diastolic pressures were measured in isolated buffer-perfused rat hearts subjected to 30 minutes of no-flow ischemia and 30 minutes of reperfusion (37 degrees C) with or without 1 micromol/L HOE642 added to the perfusate 15 minutes before ischemia. Intracellular Ca(2+) concentration ([Ca(2+)](i)) and pH(i) were measured with aequorin (n=10 per group) and (31)P NMR spectroscopy (n=6 per group), respectively. HOE642 did not affect preischemic mechanical function, [Ca(2+)](i), or pH(i). Mechanical recovery after 30 minutes of reperfusion was substantially improved with HOE642: left ventricular developed pressure (in percent of preischemic values) was 92+/-3 versus 49+/-7 and left ventricular end-diastolic pressure was 16+/-3 versus 46+/-5 mm Hg (P<0.05 for HOE642-treated versus untreated hearts). End-ischemic [Ca(2+)](i) was significantly lower in HOE642 treated than in untreated hearts (1.04+/-0.06 versus 1.84+/-0. 02 micromol/L, P<0.05). Maximal intracellular Ca(2+) overload during the first 60 seconds of reperfusion was attenuated with HOE642 compared with untreated hearts: 2.0+/-0.3 versus 3.2+/-0.3 micromol/L (P<0.05). pH(i) was not different at end ischemia ( approximately 5.9+/-0.05). Realkalinization was similar in the first 90 seconds of reperfusion and significantly delayed in the next 3 minutes (eg, 6.8+/-0.07 in HOE642-treated hearts compared with 7. 2+/-0.07 in untreated hearts; P<0.05). CONCLUSIONS: HOE642 improves postischemic recovery by reducing Ca(2+) overload during ischemia and early reperfusion and by prolonging postischemic acidosis. PMID- 10851215 TI - Design of a new surgical approach for ventricular remodeling to relieve ischemic mitral regurgitation: insights from 3-dimensional echocardiography. AB - BACKGROUND: Mechanistic insights from 3D echocardiography (echo) can guide therapy. In particular, ischemic mitral regurgitation (MR) is difficult to repair, often persisting despite annular reduction. We hypothesized that (1) in a chronic infarct model of progressive MR, regurgitation parallels 3D changes in the geometry of mitral leaflet attachments, causing increased leaflet tethering and restricting closure; therefore, (2) MR can be reduced by restoring tethering geometry toward normal, using a new ventricular remodeling approach based on 3D echo findings. METHODS AND RESULTS: We studied 10 sheep by 3D echo just after circumflex marginal ligation and 8 weeks later. MR, at first absent, became moderate as the left ventricle (LV) dilated and the papillary muscles shifted posteriorly and mediolaterally, increasing the leaflet tethering distance from papillary muscle tips to the anterior mitral annulus (P<0.0001). To counteract these shifts, the LV was remodeled by plication of the infarct region to reduce myocardial bulging, without muscle excision or cardiopulmonary bypass. Immediately and up to 2 months after plication, MR was reduced to trace-to-mild as tethering distance was decreased (P<0.0001). LV ejection fraction, global LV end-systolic volume, and mitral annular area were relatively unchanged. By multiple regression, the only independent predictor of MR was tethering distance (r(2)=0.81). CONCLUSIONS: Ischemic MR in this model relates strongly to changes in 3D mitral leaflet attachment geometry. These insights from quantitative 3D echo allowed us to design an effective LV remodeling approach to reduce MR by relieving tethering. PMID- 10851216 TI - Hemodynamic stresses induce endothelial dysfunction and remodeling of pulmonary artery in experimental compensated heart failure. AB - BACKGROUND: We hypothesized that, in compensated heart failure (HF), hemodynamic perturbations and their consequences exist in pulmonary artery (PA) despite the absence of any perturbation in thoracic aorta (TA). METHODS AND RESULTS: The left coronary artery was ligated in 20 male Wistar rats with compensated HF. Four months after ligation, these rats were compared with 20 sham-operated control rats. Blood pressure, velocity, viscosity, luminal diameter, and wall tensile and shear stresses were determined in PA and TA. Arterial rings were mounted in a myograph for ex vivo study. Endothelial nitric oxide synthase (eNOS) mRNA expression was determined in lung and aorta. Sections of PA and TA were used for histomorphometric study. In PA from rats with compensated HF, (1) blood pressure and wall tensile stress increased, whereas blood velocity and wall shear stress decreased; (2) contractions to KCl were not altered, but maximal contraction to phenylephrine and EC(50) decreased; (3) endothelium-dependent relaxation to acetylcholine and basal NO activity were blunted, whereas endothelium-independent relaxation was preserved; (4) eNOS mRNA levels and eNOS transcription in lung nuclei decreased; and (5) medial cross-sectional area, thickness, smooth muscle cell number, elastin, and collagen contents increased. Conversely, no such changes were found in TA from rats with compensated HF. CONCLUSIONS: In compensated HF induced by small myocardial infarction, hemodynamics, vascular wall function, and structure are altered in PA but preserved in TA. These results indicate that the pulmonary vascular bed is an early target of regional circulatory alterations in HF. PMID- 10851217 TI - Images in cardiovascular medicine. Magnetic resonance imaging and asymptomatic aortic dissection. PMID- 10851218 TI - PhysioBank, PhysioToolkit, and PhysioNet: components of a new research resource for complex physiologic signals. AB - The newly inaugurated Research Resource for Complex Physiologic Signals, which was created under the auspices of the National Center for Research Resources of the National Institutes of Health, is intended to stimulate current research and new investigations in the study of cardiovascular and other complex biomedical signals. The resource has 3 interdependent components. PhysioBank is a large and growing archive of well-characterized digital recordings of physiological signals and related data for use by the biomedical research community. It currently includes databases of multiparameter cardiopulmonary, neural, and other biomedical signals from healthy subjects and from patients with a variety of conditions with major public health implications, including life-threatening arrhythmias, congestive heart failure, sleep apnea, neurological disorders, and aging. PhysioToolkit is a library of open-source software for physiological signal processing and analysis, the detection of physiologically significant events using both classic techniques and novel methods based on statistical physics and nonlinear dynamics, the interactive display and characterization of signals, the creation of new databases, the simulation of physiological and other signals, the quantitative evaluation and comparison of analysis methods, and the analysis of nonstationary processes. PhysioNet is an on-line forum for the dissemination and exchange of recorded biomedical signals and open-source software for analyzing them. It provides facilities for the cooperative analysis of data and the evaluation of proposed new algorithms. In addition to providing free electronic access to PhysioBank data and PhysioToolkit software via the World Wide Web (http://www.physionet. org), PhysioNet offers services and training via on-line tutorials to assist users with varying levels of expertise. PMID- 10851219 TI - Noninvasive detection of coronary artery stenosis by multislice helical computed tomography. PMID- 10851220 TI - Hormone replacement therapy and cardiac prevention. PMID- 10851221 TI - Coumadin versus warfarin. PMID- 10851222 TI - Impending paradoxical embolism. PMID- 10851223 TI - Risk of acute myocardial infarction in cocaine abusers. PMID- 10851224 TI - Hypertension, diabetes mellitus, hypercholesterolemia, and endothelin B receptor mediated renal nitric oxide release. PMID- 10851225 TI - Shaking the world of gene therapy. PMID- 10851226 TI - Jan L. Breslow, MD, receives 10th Annual Bristol-Myers Squibb Award. PMID- 10851227 TI - Definition of an amino-terminal domain of the human T-cell leukemia virus type 1 envelope surface unit that extends the fusogenic range of an ecotropic murine leukemia virus. AB - Murine leukemia viruses (MuLV) and human T-cell leukemia viruses (HTLV) are phylogenetically highly divergent retroviruses with distinct envelope fusion properties. The MuLV envelope glycoprotein surface unit (SU) comprises a receptor binding domain followed by a proline-rich region which modulates envelope conformational changes and fusogenicity. In contrast, the receptor-binding domain and SU organization of HTLV are undefined. Here, we describe an HTLV/MuLV envelope chimera in which the receptor-binding domain and proline-rich region of the ecotropic MuLV were replaced with the potentially corresponding domains of the HTLV-1 SU. This chimeric HTLV/MuLV envelope was processed, specifically interfered with HTLV-1 envelope-mediated fusion, and similar to MuLV envelopes, required cleavage of its cytoplasmic tail to exert significant fusogenic properties. Furthermore, the HTLV domain defined here broadened ecotropic MuLV envelope-induced fusion to human and simian cell lines. PMID- 10851228 TI - Activation of the particulate and not the soluble guanylate cyclase leads to the inhibition of Ca2+ extrusion through localized elevation of cGMP. AB - We examined whether localized increases in cytosolic cGMP have distinct regulatory effects on the concentration of cytosolic free Ca(2+) in ECV304 cells. Stimulation of the particulate guanylate cyclase by brain-type natriuretic peptide in fura-2-loaded cells caused a profound potentiation of the ATP stimulated and thapsigargin-stimulated rise in cytosolic free Ca(2+). This effect is mediated by the inhibition of Ca(2+) extrusion via the plasma membrane Ca(2+) ATPase pump. Furthermore, the addition of brain-type natriuretic peptide caused the partial inhibition of cation influx in ATP-stimulated cells. In contrast, elevation of cytosolic cGMP by activation of the soluble guanylate cyclase induced by the addition of sodium nitroprusside causes an increased reuptake of Ca(2+) into the intracellular stores without affecting cation influx or Ca(2+) efflux. Thus, localized pools of cGMP play distinct regulatory roles in the regulation of Ca(2+) homeostasis within individual cells. We define a new role for natriuretic peptides in the inhibition of Ca(2+) efflux that leads to the potentiation of agonist-evoked increases in cytosolic free Ca(2+). PMID- 10851229 TI - A p300 protein as a coactivator of GATA-6 in the transcription of the smooth muscle-myosin heavy chain gene. AB - The mechanisms that regulate smooth muscle development and differentiation are poorly understood. Although recent studies have suggested the possible role of a zinc finger transcription factor, GATA-6, in the differentiation of vascular smooth muscle cells (VSMCs), the downstream gene targeted by GATA-6 is unknown. The expression of smooth muscle-myosin heavy chain (Sm-MHC) provides a highly specific marker for the differentiated phenotype of VSMCs as well as the smooth muscle cell lineage. Here, we show that GATA-6 bound to a GATA-like motif (-810/ 805) within the rat Sm-MHC promoter in a sequence-specific manner and activated this promoter through this site. In addition, we show that the transcriptional coactivator p300 associated with GATA-6 during the transcription of the Sm-MHC gene. A p300/GATA-6 complex in VSMCs was up-regulated by induction of the quiescent phenotype. A wild-type E1A, which interferes with endogenous p300, but not a mutant E1A defective for p300 binding, markedly down-regulated the expression of endogenous Sm-MHC in quiescent-phenotype VSMCs. These studies provide the first identification of a functionally important GATA-6 binding site within a smooth muscle-specific promoter and suggest a role for p300 in the maintenance of the differentiated phenotype in VSMCs as a coactivator of GATA-6. PMID- 10851230 TI - Cleavage of holliday junctions by the Escherichia coli RuvABC complex. AB - The Escherichia coli RuvABC proteins process recombination intermediates during genetic recombination and recombinational repair. Although early biochemical studies indicated distinct RuvAB-mediated branch migration and RuvC-mediated Holliday junction resolution reactions, more recent studies have shown that the three proteins act together as a "resolvasome" complex. In this work we have used recombination intermediates made by RecA to determine whether the RuvAB proteins affect the sequence specificity of the RuvC resolvase. We find that RuvAB proteins do not alter significantly the site specificity of RuvC-dependent cleavage, although under certain conditions, they do affect the efficiency of cleavage at particular sites. The presence of RecA also influences cleavage at some sites. We also show that the RuvAB proteins act upon transient strand exchange intermediates made using substrates that have the opposite polarity of those preferred by RecA. Together, our results allow us to develop further a model for the recombinational repair of DNA lesions that lead to the formation of post-replication gaps during DNA replication. The novel features of this model are as follows: (i) the RuvABC resolvasome recognizes joints made by RecA; (ii) resolution by RuvABC occurs at specific sites containing the RuvC consensus cleavage sequence 5'-(A/T)TT downward arrow(G/C)-3'; and (iii) Holliday junction resolution often occurs close to the initiating gap without significant heteroduplex DNA formation. PMID- 10851231 TI - Expression of phosphodiesterase 4D (PDE4D) is regulated by both the cyclic AMP dependent protein kinase and mitogen-activated protein kinase signaling pathways. A potential mechanism allowing for the coordinated regulation of PDE4D activity and expression in cells. AB - Multiple families of cyclic nucleotide phosphodiesterases (PDE) have been described, and the regulated expression of these genes in cells is complex. Although cAMP is known to control the expression of certain PDE in cells, presumably reflecting a system of feedback on cAMP signaling, relatively little is known about the influence of non-cAMP signaling systems on PDE expression. In this study, we describe a novel mechanism by which activators of the protein kinase C (PKC)-Raf-MEK-ERK cascade regulate phosphodiesterase 4D (PDE4D) expression in vascular smooth muscle cells (VSMC) and assess the functional consequences of this effect. Whereas a prolonged elevation of cAMP in VSMC resulted in a protein kinase A (PKA)-dependent induction of expression of two PDE4D variants (PDE4D1 and PDE4D2), simultaneous activation of both the cAMP-PKA and PKC-Raf-MEK-ERK signaling cascades blunted this cAMP-mediated increase in PDE4D expression. By using biochemical, molecular biological, and pharmacological approaches, we demonstrate that this PDE4D-selective effect of activators of the PKC-Raf-MEK-ERK cascade was mediated through a mechanism involving altered PDE4D mRNA stability and markedly attenuated the cAMP-mediated desensitization that results from prolonged activation of the cAMP signaling system in cells. The data are presented in the context of activators of the PKC-Raf-MEK-ERK cascade having both short and long term effects on PDE4D activity and expression in cells that may influence cAMP signaling. PMID- 10851232 TI - Induction and modulation of cerebellar granule neuron death by E2F-1. AB - Growing evidence suggests that certain cell cycle regulators also mediate neuronal death. Of relevance, cyclin D1-associated kinase activity is increased and the retinoblastoma protein (Rb), a substrate of the cyclin D1-Cdk4/6 complex, is phosphorylated during K(+) deprivation-evoked death of cerebellar granule neurons (CGNs). Cyclin-dependent kinase (CDK) inhibitors block this death, suggesting a requirement for the cyclin D1/Cdk4/6-Rb pathway. However, the downstream target(s) of this pathway are not well defined. The transcription factor E2F-1 is regulated by Rb and is reported to evoke death in proliferating cells when overexpressed. Accordingly, we examined whether E2F-1 was sufficient to evoke death of CGNs and whether it was required for death evoked by low K(+). We show that adenovirus-mediated expression of E2F-1 in CGNs results in apoptotic death, which is independent of p53, dependent upon Bax, and associated with caspase 3-like activity. In addition, we demonstrate that levels of E2F-1 mRNA and protein increase during K(+) deprivation-evoked death. The increase in E2F-1 protein is blocked by the CDK inhibitor flavopiridol. Finally, E2F-1-deficient neurons are modestly resistant to death induced by low K(+). These results indicate that E2F-1 expression is sufficient to promote neuronal apoptosis and that endogenous E2F-1 modulates the death of CGNs evoked by low K(+). PMID- 10851233 TI - Extracellular zinc activates p70 S6 kinase through the phosphatidylinositol 3 kinase signaling pathway. AB - We have studied a possible role of extracellular zinc ion in the activation of p70S6k, which plays an important role in the progression of cells from the G(1) to S phase of the cell cycle. Treatment of Swiss 3T3 cells with zinc sulfate led to the activation and phosphorylation of p70S6k in a dose-dependent manner. The activation of p70S6k by zinc treatment was biphasic, the early phase being at 30 min followed by the late phase at 120 min. The zinc-induced activation of p70S6k was partially inhibited by down-regulation of phorbol 12-myristate 13-acetate responsive protein kinase C (PKC) by chronic treatment with phorbol 12-myristate 13-acetate, but this was not significant. Moreover, Go6976, a specific calcium dependent PKC inhibitor, did not significantly inhibit the activation of p70S6k by zinc. These results demonstrate that the zinc-induced activation of p70S6k is not related to PKC. Also, extracellular calcium was not involved in the activation of p70S6k by zinc. Further characterization of the zinc-induced activation of p70S6k using specific inhibitors of the p70S6k signaling pathway, namely rapamycin, wortmannin, and LY294002, showed that zinc acted upstream of mTOR/FRAP/RAFT and phosphatidylinositol 3-kinase (PI3K), because these inhibitors caused the inhibition of zinc-induced p70S6k activity. In addition, Akt, the upstream component of p70S6k, was activated by zinc in a biphasic manner, as was p70S6k. Moreover, dominant interfering alleles of Akt and PDK1 blocked the zinc induced activation of p70S6k, whereas the lipid kinase activity of PI3K was potently activated by zinc. Taken together, our data suggest that zinc activates p70S6k through the PI3K signaling pathway. PMID- 10851234 TI - Phosphorylation site analysis of Semliki forest virus nonstructural protein 3. AB - Nonstructural protein 3 (Nsp3) is an essential subunit of the alphavirus RNA replication complex, although its specific function(s) has yet to be well defined. Previously, it has been shown that Semliki Forest virus Nsp3 (482 amino acids) is a phosphoprotein, and, in the present study, we have mapped its major phosphorylation sites. Mass spectrometric methods utilized included precursor ion scanning, matrix-assisted laser desorption/ionization mass spectrometry used in conjunction with on-target alkaline phosphatase digestions, and tandem mass spectrometry. Two-dimensional peptide mapping was applied to separate tryptic (32)P-labeled phosphopeptides of Nsp3. Radiolabeled peptides were then subjected to Edman sequencing, and phosphoamino acid analysis. In addition, radiolabeled Nsp3 was cleaved successively with cyanogen bromide and trypsin, and microscale iron-chelate affinity chromatography was used to enrich phosphopeptides. By combining these methods, we showed that Nsp3 is phosphorylated on serine residues 320, 327, 332, 335, 356, 359, 362, and 367, and is heavily phosphorylated on peptide Gly(338)-Lys(415), which carries 7-12 phosphates distributed over its 13 potential phosphorylation sites. These analytical findings were corroborated by constructing a Nsp3 derivative devoid of phosphorylation. The results represent the first determination of phosphorylation sites of an alphavirus nonstructural protein, but the approach can be utilized in phosphoprotein analysis in general. PMID- 10851235 TI - Cross-utilization of the beta sliding clamp by replicative polymerases of evolutionary divergent organisms. AB - Chromosomal replicases are multiprotein machines comprised of a DNA polymerase, a sliding clamp, and a clamp loader. This study examines replicase components for their ability to be switched between Gram-positive and Gram-negative organisms. These two cell types diverged over 1 billion years ago, and their sequences have diverged widely. Yet the Escherichia coli beta clamp binds directly to Staphylococcus aureus PolC and makes it highly processive, confirming and extending earlier results (Low, R. L., Rashbaum, S. A. , and Cozzarelli, N. R. (1976) J. Biol. Chem. 251, 1311-1325). We have also examined the S. aureus beta clamp. The results show that it functions with S. aureus PolC, but not with E. coli polymerase III core. PolC is a rather potent polymerase by itself and can extend a primer with an intrinsic speed of 80-120 nucleotides per s. Both E. coli beta and S. aureus beta converted PolC to a highly processive polymerase, but surprisingly, beta also increased the intrinsic rate of DNA synthesis to 240-580 nucleotides per s. This finding expands the scope of beta function beyond a simple mechanical tether for processivity to include that of an effector that increases the intrinsic rate of nucleotide incorporation by the polymerase. PMID- 10851236 TI - The human homolog of Escherichia coli Orn degrades small single-stranded RNA and DNA oligomers. AB - We report here the identification of human homologues to the essential Escherichia coli Orn protein and the related yeast mitochondrial DNA-escape pathway regulatory factor Ynt20. The human proteins appear to arise from alternatively spliced transcripts, and are thus identical, except the human Ynt20 equivalent contains an NH(2)-terminal extension that possesses a predicted mitochondrial protease cleavage signal. In vitro analysis revealed that the smaller human protein exhibits a 3' to 5' exonuclease activity for small (primarily 100-fold relative to wild-type pol beta. Template engineering experiments performed to distinguish among three possible models for deletion formation suggest that most deletions in repetitive sequences by pol beta initiate by strand slippage. However, pol beta also generates deletions by a different mechanism that is strongly enhanced by the substitutions at Arg(283). Analysis of error specificity suggests that this mechanism involves nucleotide misinsertion followed by primer relocation, creating a misaligned intermediate. The structure of pol beta bound to non-gapped DNA also indicates that the templating nucleotide and its downstream neighbor are out of register in the open conformation and this could facilitate misalignment (dNTP or primer terminus) with the next template base. PMID- 10851239 TI - Characterization of the human cysteinyl leukotriene 2 receptor. AB - The contractile and inflammatory actions of the cysteinyl leukotrienes (CysLTs), LTC(4), LTD(4), and LTE(4), are thought to be mediated through at least two distinct but related CysLT G protein-coupled receptors. The human CysLT(1) receptor has been recently cloned and characterized. We describe here the cloning and characterization of the second cysteinyl leukotriene receptor, CysLT(2), a 346-amino acid protein with 38% amino acid identity to the CysLT(1) receptor. The recombinant human CysLT(2) receptor was expressed in Xenopus oocytes and HEK293T cells and shown to couple to elevation of intracellular calcium when activated by LTC(4), LTD(4), or LTE(4). Analyses of radiolabeled LTD(4) binding to the recombinant CysLT(2) receptor demonstrated high affinity binding and a rank order of potency for competition of LTC(4) = LTD(4) LTE(4). In contrast to the dual CysLT(1)/CysLT(2) antagonist, BAY u9773, the CysLT(1) receptor-selective antagonists MK-571, montelukast (Singulair(TM)), zafirlukast (Accolate(TM)), and pranlukast (Onon(TM)) exhibited low potency in competition for LTD(4) binding and as antagonists of CysLT(2) receptor signaling. CysLT(2) receptor mRNA was detected in lung macrophages and airway smooth muscle, cardiac Purkinje cells, adrenal medulla cells, peripheral blood leukocytes, and brain, and the receptor gene was mapped to chromosome 13q14, a region linked to atopic asthma. PMID- 10851240 TI - Possible involvement of aminotelopeptide in self-assembly and thermal stability of collagen I as revealed by its removal with proteases. AB - The functions of aminotelopeptide and N-terminal cross-linking of collagen I were examined. Acetic acid-soluble collagen I (ASC) was purified from neonatal bovine skin and treated with three kinds of proteases. The amino acid sequencing analysis of the N terminus showed that ASC contained a full-length aminotelopeptide. Pepsin and papain cleaved the aminotelopeptide of the alpha1 chain at the same site and the aminotelopeptide of the alpha2 chain at different sites. Proctase-treated ASC lost the whole aminotelopeptide, and the N-terminal sequence began from the tenth residue inside the triple helical region. The rates of fibril formation of pepsin-treated ASC and proctase-treated ASC were the same and were slower than that of ASC. The denaturation temperatures, monitored by CD ellipticity at 221 nm, of ASC, pepsin-treated, or papain-treated collagens were the same at 41.8 degrees C. Proctase-treated ASC showed a lower denaturation temperature of 39.9 degrees C. We also observed the morphology of the collagen fibrils under an electron microscope. The ASC fibrils were straight and thin, whereas the fibrils of pepsin-treated ASC were slightly twisted, and the fibrils from papain- and proctase-treated ASC were highly twisted and thick. When the collagen gel strength was examined by a modified method of viscosity-measurement, ASC was the strongest, followed by pepsin-treated ASC, and papain- and proctase treated ASCs were the weakest. These results suggest that the aminotelopeptide plays important roles in fibril formation and thermal stability. In addition, the functions of intermolecular cross-linking in aminotelopeptides may contribute to the formation of fibrils in the correct staggered pattern and to strengthening the collagen gel. PMID- 10851241 TI - Isolation and characterization of a novel inducible mammalian galectin. AB - A novel mammalian galectin cDNA (ovgal11) was isolated by representational difference analysis from sheep stomach (abomasal) tissue infected with the nematode parasite, Haemonchus contortus. The mRNA is greatly up-regulated in helminth larval infected gastrointestinal tissue subject to inflammation and eosinophil infiltration. Immunohistological analysis indicates that the protein is localized in the cytoplasm and nucleus of upper epithelial cells of the gastrointestinal tract. The protein is also detected in mucus samples collected from infected abomasum but not from uninfected tissue. The restricted and inducible expression of ovgal11 mRNA and limited secretion of the protein support the hypothesis that OVGAL11 may be involved in gastrointestinal immune/inflammatory responses and possibly protection against infection. PMID- 10851242 TI - Receptor for the pain modulatory neuropeptides FF and AF is an orphan G protein coupled receptor. AB - Opiate tolerance and dependence are major clinical and social problems. The anti opiate neuropeptides FF and AF (NPFF and NPAF) have been implicated in pain modulation as well as in opioid tolerance and may play a critical role in this process, although their mechanism of action has remained unknown. Here we describe a cDNA encoding a novel neuropeptide Y-like human orphan G protein coupled receptor (GPCR), referred to as HLWAR77 for which NPAF and NPFF have high affinity. Cells transiently or stably expressing HLWAR77 bind and respond in a concentration-dependent manner to NPAF and NPFF and are also weakly activated by FMRF-amide (Phe-Met-Arg-Phe-amide) and a variety of related peptides. The high affinity and potency of human NPFF and human NPAF for HLWAR77 strongly suggest that these are the cognate ligands for this receptor. Expression of HLWAR77 was demonstrated in brain regions associated with opiate activity, consistent with the pain-modulating activity of these peptides, whereas the expression in adipose tissue suggests other physiological and pathophysiological activities for FMRF amide neuropeptides. The discovery that the anti-opiate neuropeptides are the endogenous ligands for HLWAR77 will aid in defining the physiological role(s) of these ligands and facilitate the identification of receptor agonists and antagonists. PMID- 10851243 TI - Calcium influx factor directly activates store-operated cation channels in vascular smooth muscle cells. AB - Recently, we described a novel 3-pS Ca(2+)-conducting channel that is activated by BAPTA and thapsigargin-induced passive depletion of intracellular Ca(2+) stores and likely to be a native store-operated channel in vascular smooth muscle cells (SMC). Neither Ca(2+) nor inositol 1,4,5-trisphosphate or other second messengers tested activated this channel in membrane patches excised from resting SMC. Here we report that these 3-pS channels are activated in inside-out membrane patches from SMC immediately upon application of Ca(2+) influx factor (CIF) extracted from mutant yeast, which has been previously shown to activate Ca(2+) influx in Xenopus oocytes and Ca(2+) release-activated Ca(2+) current in Jurkat cells. In bioassay experiments depletion of Ca(2+) stores in permeabilized human platelets resulted in the release of endogenous factor, which activated 3-pS channels in isolated inside-out membrane patches excised from SMC and exposed to permeabilized platelets. The same 3-pS channels in excised membrane patches were also activated by acid extracts of CIF derived from human platelets with depleted Ca(2+) stores, which also stimulated Ca(2+) influx upon injection into Xenopus oocytes. Specific high pressure liquid chromatography fractions of platelet extracts were found to have CIF activity when injected into oocytes and activate 3-pS channels in excised membrane patches. These data show for the first time that CIF produced by mammalian cells and yeast with depleted Ca(2+) stores directly activates native 3-pS cation channels, which in intact SMC are activated by Ca(2+) store depletion. PMID- 10851244 TI - Intersectin, an adaptor protein involved in clathrin-mediated endocytosis, activates mitogenic signaling pathways. AB - Intersectin is a member of a growing family of adaptor proteins that possess conserved Eps15 homology (EH) domains as well as additional protein recognition motifs. In general, EH domain-containing proteins play an integral role in clathrin-mediated endocytosis. Indeed, intersectin functions in the intermediate stages of clathrin-coated vesicle assembly. However, recent evidence suggests that components of the endocytic machinery also regulate mitogenic signaling pathways. In this report, we provide several lines of evidence that intersectin has the capacity to activate mitogenic signaling pathways. First, intersectin overexpression activated the Elk-1 transcription factor in an MAPK-independent manner. This ability resides within the EH domains, as expression of the tandem EH domains was sufficient to activate Elk-1. Second, intersectin cooperated with epidermal growth factor to potentiate Elk-1 activation; however, a similar level of Elk-1 activation was obtained by expression of the tandem EH domains suggesting that the coiled-coil region and SH3 domains act to regulate the EH domains. Third, intersectin expression was sufficient to induce oncogenic transformation of rodent fibroblasts. And finally, intersectin cooperated with progesterone to accelerate maturation of Xenopus laevis oocytes. Together, these data suggest that intersectin links endocytosis with regulation of pathways important for cell growth and differentiation. PMID- 10851245 TI - Latrotoxin stimulates secretion in permeabilized cells by regulating an intracellular Ca2+ - and ATP-dependent event: a role for protein kinase C. AB - alpha-Latrotoxin, a component of black widow spider venom, stimulates transmitter release from nerve terminals and intact chromaffin cells and enhances secretion from permeabilized chromaffin cells already maximally stimulated by Ca(2+). In this study we demonstrate that chromaffin cells contain a protein antigenically similar to the cloned Ca(2+)-independent receptor for alpha-latrotoxin. Although this receptor has homology to the secretin family of G-protein-linked receptors, pertussis toxin has no effect on the ability of alpha-latrotoxin to enhance secretion, suggesting that neither G(i) nor G(o) is involved in the response. Furthermore, in the absence of Ca(2+), alpha-latrotoxin does not stimulate polyphosphoinositide-specific phospholipase C. alpha-Latrotoxin specifically enhances ATP-dependent secretion in permeabilized cells. An in situ assay for protein kinase C reveals that alpha-latrotoxin augments the activation of protein kinase C by Ca(2+), and use of protein kinase inhibitors demonstrates that this activation is important for the toxin's enhancing effect. This enhancement of secretion requires Ca(2+) concentrations above 3 microm and is not supported by Ba(2+) or nonhydrolyzable guanine nucleotides, which do not stimulate protein kinase C. We conclude that alpha-latrotoxin stimulates secretion in permeabilized cells by regulating a Ca(2+)- and ATP-dependent event involving protein kinase C. PMID- 10851246 TI - Regulation of human endothelial cell focal adhesion sites and migration by cGMP dependent protein kinase I. AB - cGMP-dependent protein kinase type I (cGK I), a major constituent of the atrial natriuretic peptide (ANP)/nitric oxide/cGMP signal transduction pathway, phosphorylates the vasodilator-stimulated phosphoprotein (VASP), a member of the Ena/VASP family of proteins involved in regulation of the actin cytoskeleton. Here we demonstrate that stimulation of human umbilical vein endothelial cells (HUVECs) by both ANP and 8-(4-chlorophenylthio)guanosine 3':5'-monophosphate (8 pCPT-cGMP) activates transfected cGK I and causes detachment of VASP and its known binding partner (zyxin) from focal adhesions in >60% of cells after 30 min. The ANP effects, but not the 8-pCPT-cGMP effects, reversed after 3 h of treatment. In contrast, a catalytically inactive cGK Ibeta mutant (cGK Ibeta K405A) was incapable of mediating these effects. VASP mutated (Ser/Thr to Ala) at all three of its established phosphorylation sites (vesicular stomatitis virus tagged VASP-AAA mutant) was not phosphorylated by cGK I and was resistant to detaching from HUVEC focal adhesions in response to 8-pCPT-cGMP. Furthermore, activation of cGK I, but not of mutant cGK Ibeta-K405A, caused a 1.5-2-fold inhibition of HUVEC migration, a dynamic process highly dependent on focal adhesion formation and disassembly. These results indicate that cGK I phosphorylation of VASP results in loss of VASP and zyxin from focal adhesions, a response that could contribute to cGK alteration of cytoskeleton-regulated processes such as cell migration. PMID- 10851247 TI - Identification of CRAM, a novel unc-33 gene family protein that associates with CRMP3 and protein-tyrosine kinase(s) in the developing rat brain. AB - Four members of collapsin response mediator proteins (CRMPs) are thought to be involved in the semaphorin-induced growth cone collapse during neural development. Here we report the identification of a novel CRMP3-associated protein, designated CRAM for CRMP3-associated molecule, that belongs to the unc 33 gene family. The deduced amino acid sequence reveals that the CRAM gene encodes a protein of 563 amino acids, shows 57% identity with dihydropyrimidinase, and shows 50-51% identity with CRMPs. CRAM appears to form a large complex composed of CRMP3 and other unidentified proteins in vivo. Indeed, CRAM physically associates with CRMP3 when co-expressed in COS-7 cells. The expression of CRAM is brain-specific, is high in fetal and neonatal rat brain, and decreases to very low levels in adult brain. Moreover, CRAM expression is up regulated during neuronal differentiation of embryonal carcinoma P19 and PC12 cells. Finally, immunoprecipitation analysis of rat brain extracts shows that CRAM is co-immunoprecipitated with proteins that contain protein-tyrosine kinase activity. Taken together, our results suggest that CRAM, which interacts with CRMP3 and protein-tyrosine kinase(s), is a new member of an emerging family of molecules that potentially mediate signals involved in the guidance and outgrowth of axons. PMID- 10851248 TI - A novel human rad54 homologue, Rad54B, associates with Rad51. AB - Members of the SNF2/SWI2 family, characterized with sequence motifs similar to those found in DNA and RNA helicases, play roles in various aspects of cellular fundamental processes such as transcriptional regulation, chromosome stability, nucleotide excision repair, and recombination. We have isolated a novel member of the human SNF2/SWI2 family, RAD54B, which is highly homologous to mammalian RAD54. The RAD54 gene is a member of the RAD52 epistasis group which is involved in the recombinational repair of DNA damage. Here we demonstrate that human Rad54B (hRad54B), like human Rad54 (hRad54), associates with human Rad51 (hRad51). Both hRad54B and hRad54 associate with hRad51 through their NH(2) terminal domains, but there are differences in their ways of association with hRad51. In contrast to Rad54, whose association with Rad51 is induced by ionizing radiation, Rad54B associates with Rad51 constitutively in immunoprecipitation experiments. Also, the failure to detect the interaction between hRad54B and hRad51 in the yeast two-hybrid assay suggests that their interaction, unlike that between hRad54 and hRad51, may be indirect. Immunofluorescence microscopy revealed that hRad54B formed nuclear foci that colocalized with hRad51, hRad54, and BRCA1. These findings suggest that Rad54B may be functionally distinct from Rad54, although it may play an active role in recombination processes in concert with other members of the RAD52 epistasis group. PMID- 10851249 TI - Perfect endings: a review of subtelomeric probes and their use in clinical diagnosis. AB - Chromosomal rearrangements involving the ends of chromosomes (telomeres) are emerging as an important cause of human genetic diseases. This review describes the development of first and second generation sets of telomere specific clones, together with advances in fluorescence in situ hybridisation (FISH) technology, which have made the prospect of screening for telomeric rearrangements a realistic goal. Initial FISH studies using the telomere specific clones indicate that they will be a valuable diagnostic tool for the investigation of mental retardation, the characterisation of known abnormalities detected by conventional cytogenetic analysis, spontaneous recurrent miscarriages, infertility, haematological malignancies, and preimplantation diagnosis, as well as other fields of clinical interest. In addition, they may help investigate telomere structure and function and can be used in the identification of dosage sensitive genes involved in human genetic disease. PMID- 10851250 TI - Over-representation of PPARgamma sequence variants in sporadic cases of glioblastoma multiforme: preliminary evidence for common low penetrance modifiers for brain tumour risk in the general population. AB - PPARgamma, the gamma isoform of a family of peroxisome proliferator activated receptors, plays a key role in adipocyte differentiation. Recently, its broad expression in multiple tissues and several epithelial cancers has been shown. Further, somatic loss of function mutations in PPARgamma have been found in primary colorectal carcinomas. We sought to determine if somatic high penetrance mutations in this gene might also play a role in glioblastoma multiforme (GBM). We also examined this gene to determine if common low penetrance polymorphic alleles might lend low level susceptibility to GBM in the general population. No somatic high penetrance mutations were detected in 96 sporadic GBMs. However, polymorphic alleles at codons 12 and 449 were significantly over-represented among the 27 unrelated American patients with sporadic GBM compared to 80 race matched controls. While nine (33%) were heterozygous for the P12A variant, c.34C/G (cytosine to guanine change at nucleotide 34), 12 (15%) controls were heterozygous for P12A (p<0.05). Similarly, 13 of 26 (50%) glioblastoma patients compared to 10 of 80 (12%) normal controls were found to have the heterozygous H449H polymorphism (p<0.001). The over-representation of H449H in glioblastoma patients was confirmed with a second validation set of American patients. When both American series were combined, polymorphic H449H was over-represented among cases versus controls (p<0.001) and there was a similar trend (p=0.07) for P12A. The precise mechanism for this association is unknown but these PPARgamma polymorphisms may be acting in a low penetrance predisposing manner. However, these associations were not found in a German population, possibly arguing that if these variants are in linkage disequilibrium with a third locus, then this effect is relatively new, after the settlement of the American colonies. PMID- 10851251 TI - FRAXA and FRAXE: the results of a five year survey. AB - We report the results of a five year survey of FRAXA and FRAXE mutations among boys aged 5 to 18 with special educational needs (SEN) related to learning disability. We tested their mothers using the X chromosome not transmitted to the son as a control chromosome, and the X chromosome inherited by the son to provide information on stability of transmission. We tested 3738 boys and 2968 mothers and found 20 FRAXA and one FRAXE full mutations among the boys and none among the mothers. This gives an estimated prevalence of full mutations in males of 1 in 5530 for FRAXA and 1 in 23 423 for FRAXE. We found an excess of intermediate and premutation alleles for both FRAXA and FRAXE. For FRAXA this was significant at the 0.001 level but the excess for FRAXE was significant only at the 0.03 level. We conclude that the excess of intermediate and premutation sized alleles for FRAXA may well be a contributing factor to the boys' mental impairment, while that for FRAXE may be a chance finding. We studied approximately 3000 transmissions from mother to son and found five instabilities of FRAXA in the common or intermediate range and three instabilities of FRAXE in the intermediate range. Thus instabilities in trinucleotide repeat size for FRAXA and FRAXE are rare, especially among alleles in the common size range. PMID- 10851252 TI - Molecular characterisation of congenital glaucoma in a consanguineous Canadian community: a step towards preventing glaucoma related blindness. AB - Glaucoma is a leading cause of irreversible blindness in Canada. Congenital glaucoma usually manifests during the first years of life and is characterised by severe visual loss and autosomal recessive inheritance. Two disease loci, on chromosomes 1p36 and 2p21, have been associated with various forms of congenital glaucoma. A branch of a large six generation family from a consanguineous Amish community in south western Ontario was affected with congenital glaucoma and was studied by linkage and mutational analysis to identify the glaucoma related genetic defects. Linkage analysis using the MLINK component of the LINKAGE package (v 5.1) showed evidence of linkage to the 2p21 region (Zmax=3.34, theta=0, D2S1348 and D2S1346). Mutational analysis of the primary candidate gene, CYP1B1, was done by direct cycle sequencing, dideoxy fingerprinting analysis, and fragment analysis. Two different disease causing mutations in exon 3, 1410del13 and 1505G-->A, both segregated with the disease phenotype. The two different combinations of these alleles appeared to result in a variable expressivity of the phenotype. The compound heterozygote appeared to have a milder phenotype when compared to the homozygotes for the 13 bp deletion. The congenital glaucoma phenotype for this large inbred Amish family is the result of mutations in CYP1B1 (2p21). The molecular information derived from this study will be used to help identify carriers of the CYP1B1 mutation in this community and optimise the management of those at risk of developing glaucoma. PMID- 10851253 TI - Circadian rhythm abnormalities of melatonin in Smith-Magenis syndrome. AB - BACKGROUND: Smith-Magenis syndrome (SMS) is a multiple congenital anomalies/mental retardation syndrome associated with a hemizygous deletion of chromosome 17, band p11.2. Characteristic features include neurobehavioural abnormalities such as aggressive and self-injurious behaviour and significant sleep disturbances. The majority of patients have a common deletion characterised at the molecular level. Physical mapping studies indicate that all patients with the common deletion are haploinsufficient for subunit 3 of the COP9 signalosome (COPS3), which is conserved from plants to humans, and in the plant Arabidopis thaliana regulates gene transcription in response to light. Haploinsufficiency of this gene is hypothesised to be potentially involved in the sleep disturbances seen in these patients. Melatonin is a hormone secreted by the pineal gland. SMS patients are reported to have fewer sleep disturbances when given a night time dose of this sleep inducing hormone. METHODS: Urinary excretion of 6 sulphatoxymelatonin (aMT6s), the major hepatic metabolite of melatonin, in 19 SMS patients were measured in conjunction with 24 hour sleep studies in 28 SMS patients. Five of the 28 patients did not have the common SMS deletion. To investigate a potential correlation of COPS3 haploinsufficiency and disturbed melatonin excretion, we performed fluorescence in situ hybridisation (FISH) using two BACs containing coding exons of COPS3. RESULTS: All SMS patients show significant sleep disturbances when assessed by objective criteria. Abnormalities in the circadian rhythm of aMT6s were observed in all but one SMS patient. Interestingly this patient did not have the common deletion. All patients studied, including the one patient with a normal melatonin rhythm, were haploinsufficient for COPS3. CONCLUSIONS: Our data indicate a disturbed circadian rhythm in melatonin and document the disturbed sleep pattern in Smith-Magenis syndrome. Our findings suggest that the abnormalities in the circadian rhythm of melatonin and altered sleep patterns could be secondary to aberrations in the production, secretion, distribution, or metabolism of melatonin; however, a direct role for COPS3 could not be established. PMID- 10851254 TI - Five cases of isolated glycerol kinase deficiency, including two families: failure to find genotype:phenotype correlation. AB - Little is understood of the genotype/phenotype correlations in X linked glycerol kinase deficiency (GKD) where most cases are caused by extensive deletions of Xp21, which often include genes flanking the GK locus. Few cases of isolated GKD have been investigated where the phenotype is not influenced by neighbouring genes. In this paper, we present the mutation data from four confirmed and one suspected case of non-deletion, isolated, X linked GKD and therefore extend the base of patients that can allow an assessment of genotype/phenotype correlations for this disease. The mutations found were two terminations leading to premature truncation of the GK polypeptide chain, one insertion, and an amino acid substitution. Phenotypic variation was observed in two families, where there was more than one affected subject carrying the same mutation, confirming previous studies that suggest there is no correlation between disease severity and genotype. Furthermore, the nature of the mutation in different families does not appear to influence the spectrum of phenotypic variation. In addition, one coding polymorphism in exon 3 has been found. The characterisation of the gene structure has been completed and shows that instead of 19 there are 21 exons. PMID- 10851255 TI - Delineation of a complex karyotypic rearrangement by microdissection and CGH in a family affected with split foot. AB - We report on a male patient and members of his family with additional material in chromosome 3. This derivative chromosome 3 was transmitted from his mother who had a complex rearrangement between chromosomes 2, 3, and 7. It was possible to delineate her chromosomal rearrangement by microdissection and reverse painting and to exclude these aberrations from being responsible for neonatal deaths and several abortions in this family. Two members of this family suffer from ectrodactyly or split hand/foot malformations (SHFM) of the feet which possibly correlates with the derivative chromosome 7 containing a breakpoint in the SHFM1 critical region involving several homeobox genes. PMID- 10851256 TI - Tietz syndrome (hypopigmentation/deafness) caused by mutation of MITF. AB - Patients with Tietz syndrome have congenital profound deafness and generalised hypopigmentation, inherited in a fully penetrant autosomal dominant fashion. The pigmentary features and complete penetrance make this syndrome distinct among syndromes with pigmentary anomalies and deafness, which characteristically have patchy depigmentation and variable penetrance. Only one family has been reported with the exact features described in the original report of this syndrome. This family was reascertained and a missense mutation was found in the basic region of the MITF gene in family members with Tietz syndrome. Mutations in other regions of this gene have been found to produce Waardenburg syndrome type 2 (WS2), which also includes pigmentary changes and hearing loss, but in contrast to Tietz syndrome, depigmentation is patchy and hearing loss is variable in WS2. PMID- 10851257 TI - Rapid detection of microdeletions using fluorescence in situ hybridisation (FISH) on buccal smears. PMID- 10851258 TI - Mutation analysis of GABRR1 and GABRR2 in autosomal recessive retinitis pigmentosa. PMID- 10851259 TI - Lim2(To3) transgenic mice establish a causative relationship between the mutation identified in the lim2 gene and cataractogenesis in the To3 mouse mutant. AB - PURPOSE: Lim2 is the gene encoding the ocular lens-specific intrinsic membrane protein MP19. We previously reported finding a single nonconservative G->T transversion in exon two of the Lim2 gene. This mutation was linked to the cataract in the To3 (Total opacity number 3) mouse mutant, confirming Lim2 as an ideal candidate gene for the To3 cataract. The aim of the present study was to substantiate a causative relationship between the mutation in the Lim2 gene and cataractogenesis in the To3 mouse mutant. To this end a Lim2To3 transgene cassette was engineered and introduced into fertilized normal mouse embryos to test its ability to induce cataractogenic lens development. METHODS: A Lim2 genomic clone was isolated and purified from a murine 129/SvJ genomic library. A restriction endonuclease map of the gene was generated using classical Southern techniques. The murine Lim2 promoter was characterized by transfecting primary chicken lens epithelial cells with Lim2 promoter-CAT reporter constructs and assaying promoter activity and specificity. This genomic clone was then used in conjunction with PCR to generate a Lim2To3 transgene cassette. After sequencing of the PCR engineered portion, the Lim2To3 transgene was then used to generate Lim2To3 transgenic mice via pronuclear injection. Founder mice and their offspring from outcrosses and intercrosses were characterized by ophthalmic examination, PCR and Southern DNA analysis, RT-PCR mRNA analysis, and histology of lens sections. RESULTS: Two mice, from independent microinjections, were identified as positive for presence of the Lim2To3 transgene cassette as well as presence of bilateral congenital cataracts and reduced eye size and mass. One of these founders was incapable of germline transmission of the transgene to offspring and was not characterized further. The other was capable of germline transmission and was characterized as described above. PCR DNA analysis revealed a perfect concordance between presence of the Lim2To3 transgene cassette and congenital cataract in offspring of this founder. Transgenic hemizygotes exhibited cataract and a reduction in eye and lens size and mass, while transgenic "homozygotes" presented with a more severe cataract and microphthalmic reduction in eye and lens size and mass. Southern analysis revealed approximately 2 copies of the transgene cassette integrated into a single chromosomal site in the founder and all hemizygous offspring. RT-PCR analysis revealed a very low ratio of Lim2To3 transgenic mRNA compared to endogenous normal Lim2. Finally, histology revealed that lens development was abnormal in mutant transgenic animals by embryonic day E15. By E19, just prior to birth, gross disorganization of secondary fibers was observed in mutants. CONCLUSIONS: These transgenic experiments firmly establish a causative relationship between the previously identified mutation in the Lim2 gene and cataractogenesis in the To3 mouse mutant. The low levels of mutant mRNA produced by the transgene cassette as compared to endogenous levels of normal Lim2 mRNA provides evidence that this dominant mutation results in a mutant MP19 protein with altered function rather than simply loss of function. PMID- 10851260 TI - Immune effector mechanisms in myocardial pathologies. AB - It is now well established that immune effector mechanisms contribute to cardiac dysfunction in several heart diseases, including myocarditis and the associated dilated cardiomyopathy, heart transplant rejection and Chagas' disease. These and other pathologies, in which cellular immunity plays an important role, contribute to morbidity and mortality world-wide. As a result of numerous studies performed in this exciting field, two major mechanisms of lymphocytotoxicity have been proposed: a secretory mechanism in which perforin and granzymes are key players, and a non-secretory mechanism involving Fas/FasL activation. While the common notion is that CTL-myocyte interaction, perforin- or Fas-based, inevitably results in target cell apoptotic death, the objective of this review is to consider the concept of non-apoptotic consequences of CTL-target cell interaction. It is proposed that depending on the myocyte status as well as on the fine balance between pro- and anti-apoptotic factors, CTL-myocyte interaction may result in a non-apoptotic, potentially reversible sustained damage to the myocytes, thus contributing to immune-mediated cardiac dysfunction. PMID- 10851261 TI - Adeno-associated viral vectors as gene delivery vehicles. AB - Adeno-associated virus (AAV), a non-pathogenic human parvovirus, is gaining attention for its potential use as a human gene therapy vector. One of the most attractive features of recombinant AAV vectors is the ability to be stably maintained in host cells as integrated proviruses. This property is particularly desireable for therapies requiring long-term correction of a genetic defect. This review highlights recent advances made in the AAV field and will discuss some limitations of rAAV vector integration. A novel method for enhancing the integration efficiency of these vectors will be presented. PMID- 10851262 TI - The role of bacterial DNA in septic arthritis. AB - Unmethylated CpG motifs are frequently found in bacterial DNA and have recently been shown to exert immunostimulatory effects on leukocytes. Bacteria produce severe septic arthritis; bacterial DNA may be involved in this process. We injected intraarticularly bacterial DNA and oligonucleotides containing unmethylated CpG motifs into knee joints of mice. Arthritis was seen by histopathology within 2 h and lasted for at least 14 days, and was characterized by an influx of monocytic, Mac-1+ cells and by a lack of T lymphocytes. Macrophages and their products such as tumor necrosis factor (TNF) alpha are essential for development of arthritis triggered by bacterial DNA containing CpG motifs. In contrast, neurophils, NK cells, and T/B cells were not instrumental in this condition. This review demonstrates that bacterial DNA containing unmethylated CpG motifs induces arthritis and indicates an important pathogenic role for bacterial DNA in septic arthritis. PMID- 10851263 TI - Molecular genetic alterations and viral presence in ophthalmic pterygium. AB - Pterygium is a lesion of the corneoscleral limbus which tends to grow in size, often recurs after surgical excision and is associated with exposure to solar light. Additionally, a family history is frequently reported. Loss of heterozygosity (LOH), increased P53 expression and the presence of oncogenic viruses, such as human papilloma virus (HPV) and herpes simplex virus (HSV), have been detected in pterygia, supporting the possible neoplastic nature of the lesion. Co-infection by HSV and HPV as well as LOH at some loci have also been correlated with clinical features, such as postoperative recurrence and history of conjunctivitis. A possible model of pterygium formation is proposed, in which genetic predisposition, environmental factors and viral infection(s) participate in a multi-step process. Future research may lead to new ways of pterygium treatment such as anti-viral or gene therapy. PMID- 10851264 TI - Sesamin and episesamin induce apoptosis in human lymphoid leukemia Molt 4B cells. AB - The exposure of human lymphoid leukemia Molt 4B cells to sesamin and episesamin which were isolated from unroasted sesame seed oil and identified by MS, and 1H and 13C-NMR, led to both growth inhibition and the induction of programmed cell death (apoptosis). Morphological change showing apoptotic bodies was observed in the Molt 4B cells treated with sesamin and episesamin. The fragmentations by sesamin and episesamin of DNA to oligonucleosomal-sized fragments that are characteristics of apoptosis were observed to be concentration-dependent, respectively. Moreover, the amount of the DNA fragments in the sesamin-treated cells was increased from 2 days, while that in the episesamin-treated cells was elevated at 3 days after addition of the compounds. These findings suggest that growth inhibitions by sesamin and episesamin of Molt 4B cells result from the induction of apoptosis in the cells. PMID- 10851265 TI - Expression of PTEN and PTEN pseudogene in endometrial carcinoma. AB - PTEN is a tumor suppressor gene and its mutation is frequently found in endometrial carcinoma. Recently, the pseudogene of PTEN has been reported to be actively transcribed in a number of cells and tissues and a potential for translation is suggested. For further understanding of the involvement of PTEN in endometrial carcinogenesis, we analysed the expressions of PTEN and the pseudogene in 36 endometrial carcinomas with special reference to the genetic status of PTEN. Mutations of PTEN were found in 42% (15/36) of the endometrial carcinomas. The transcript of the pseudogene was expressed in 6 samples (17%) of 36 endometrial carcinomas, but in none of the normal endometria. Western blot analysis showed no translated protein of the pseudogene in any of the cases. Steady level of PTEN protein expression was observed in all the cases examined. Expression level was consistent among the proliferative endometria, secretory endometria and endometrial carcinomas as long as PTEN protein was not truncated. These results indicate that PTEN is a constitutive protein in the endometrium, so that the somatic mutation of PTEN exerts a crucial effect on the endometrial carcinogenesis. In addition, the presence of the PTEN pseudogene transcript urges us caution for the mutational analysis of PTEN as well as careful choice of the probe for the detection of PTEN transcript. PMID- 10851266 TI - Modulation of complement component (C3 and factor B) biosynthesis by a histone deacetylase inhibitor in human intestinal epithelial cells. AB - Sodium butyrate enhances TNF-alpha-induced complement C3 secretion but suppresses TNF-alpha-induced factor B secretion in intestinal epithelial cells. To further evaluate the mechanism underlying these responses, we assessed the effects of trichostatin A, a compound structurally unrelated to butyrate and a potent inhibitor of histone deacetylase. The C3 and factor B secretion was evaluated by enzyme-linked immunosorbent assay (ELISA) and Northern blot, and the activation of transcription factor was assessed by an electrophoretic gel mobility shift assay (EMSA). Like sodium butyrate, trichostatin A enhanced TNF-alpha-induced C3 secretion, but suppressed TNF-alpha-induced factor B secretion. These effects were also observed at the level of mRNA. EMSAs indicated that trichostatin A weakly suppressed TNF-alpha-induced NF-kappaB and NF-IL6 activation. These observations differ from previous reports that sodium butyrate potently suppressed NF-kappaB activation but enhanced NF-IL6 activation. Trichostatin A modulated TNF-alpha-induced C3 and factor B secretion in a manner similar to that induced by sodium butyrate, suggesting that both sodium butyrate and trichostatin A exert certain counter-regulatory effects associated with histone hyperacetylation. However, it remains to be determined which factors other than histone acetylation are responsible for the counter-regulation of TNF-alpha induced C3 and factor B gene expression. PMID- 10851267 TI - Adenovirus-mediated E2F-1 gene transfer induces an apoptotic response in human gastric carcinoma cells that is enhanced by cyclin dependent kinase inhibitors. AB - E2F -1 is a transcription factor that regulates cell cycle progression into S phase. Deregulation of E2F-1 activity has been associated with cellular commitment to apoptosis. Also critical in the regulation of S-phase are the actions of the cyclin dependent kinases, Cdk2 and cdc2. Inhibition of these cyclin dependent kinases has been similarly associated with disrupting orderly S phase progression and causing subsequent apoptosis in certain cancer cells. In this study, we examine the ability of adenovirus-mediated E2F-1 overexpression to induce apoptosis in human gastric carcinoma cells. Furthermore, we investigate the effect of the cyclin dependent kinase inhibitors, olomoucine and roscovitine, on E2F-1-mediated apoptosis in human gastric carcinoma cells. AGS and SNU-1 gastric adenocarcinoma cells were infected with adenoviral vectors expressing E2F 1 (Ad5CMVE2F-1) or control viruses expressing beta-galactosidase (Ad5CMVLacZ) or lacking a transgene (Ad5). Gastric adenocarcinoma cells were then independently treated with roscovitine or olomoucine. Finally, gastric adenocarcinoma cells were infected with the various adenoviral vectors in combination with roscovitine or olomoucine. E2F-1 overexpression resulted in an 85% reduction in cell viability at 72 h compared to controls. Combining E2F-1 overexpression with roscovitine resulted in >99% reduction in cell viability by 72 h. Overexpression of E2F-1 resulted in premature S-phase entry and G2/M arrest at 24 h, followed by apoptosis by 72 h. Combining E2F-1 overexpression with roscovitine resulted in an earlier G2/M arrest, followed by a more complete, widespread apoptotic response by 24 h. Caspase 3/CPP32 activation and PARP cleavage in response to E2F-1 overexpression, alone and in combination with roscovitine, implicate the caspase cascade in E2F-1-mediated apoptosis of gastric cancer cells. Bax levels also increased in response to E2F-1 gene transfer, alone and in combination with roscovitine. E2F-1 overexpression induces widespread apoptosis in human gastric carcinoma cells. Combining E2F-1 overexpression with cyclin-dependent kinase inhibitors results in an enhanced apoptotic response, causing nearly complete gastric tumor cell death within 72 h. E2F-1 gene therapy in combination with cyclin dependent kinase inhibitors is a potentially active chemogene therapy strategy for the treatment of human gastric cancer. PMID- 10851268 TI - Fate of 6-deoxy-6-[123I]iodo-D-glucose in normal and diabetic rats. AB - The D-glucose analog 6-deoxy-6-?123Iiodo-D-glucose (6-DIG) was recently proposed as a potential tracer for the in vivo characterization of D-glucose transport in distinct cell types. In this study, the validity of such a proposal was investigated in both control and streptozotocin-induced diabetic rats. 6-DIG was injected intravenously in either control or diabetic rats. The fate of 6-DIG was assessed by scintigraphy of the injected animals, blood and urine sampling, and measurement of tissue radioactivity at the time of sacrifice, 140 min after 6-DIG injection. The half-life for 6-DIG in plasma and its accumulation in kidney and urinary bladder indicated that it was mainly eliminated from the body by glomerular filtration. The urinary elimination of 6-DIG was accelerated, however, in the polyuric diabetic rats. Bile formation also apparently contributed to the clearance of 6-DIG. Its uptake by liver, heart and muscles yielded values lower than blood concentration. The usefulness of 6-DIG as a tracer for D-glucose transport in selected organs in the perspective of clinical application, e.g. by single photon emission computed tomography, requires further investigations. PMID- 10851269 TI - Decreased fluorouracil cytotoxic effect on EB-virus transformed lymphocytes from hereditary orotic aciduria. AB - 5-Fluorouracil (5-FU), used widely for malignancies, phosphorylate mostly by uracil phosphoribosyl transferase (UPRT). Patients with hereditary orotic aciduria lack the orotate phosphoribosyl transferase (OPRT) activity. In the cancer cells, the OPRT activity is paralleled with the UPRT activity. This study shows that the UPRT activity of the hereditary orotic aciduria homozygote decreased about 40% of normal controls. Moreover, we investigated the 5-FU cytotoxic effects on hereditary orotic aciduria (one homozygote, 4 heterozygotes and 7 normal controls), using EB-virus transformed lymphocytes (EB-LC). 5-FU was addded to the culture medium at concentrations ranging from 0 to 10.0 micromol/l. The 5-FU cytotoxic effects on the homozygote were milder than those on controls at each 5-FU concentration. The 5-FU cytotoxic effects in the heterozygotes were at intermediate levels between the homozygote and controls. We speculate that 5 FU cytotoxic effects, both anti-tumor effects and adverse reactions, would be weak when a patient with hereditary orotic aciduria was treated with 5-FU. PMID- 10851270 TI - Characterization of the human peroxisome proliferator activated receptor delta gene and its expression. AB - Peroxisome proliferator activated receptors (PPARs) are nuclear receptors regulating the expression of genes involved in lipid and glucose metabolism. Three different PPARs; alpha (PPARA), gamma (PPARG) and delta (PPARD) have been characterized and they are distinguished from each other by tissue distribution and cell activation. In this study, the structure and detailed chromosomal localization of the human PPARD gene was determined. Three genomic clones containing the PPARD gene was isolated from a human P1 library. The gene spans approximately 85 kb of DNA and consists of 9 exons and 8 introns with exons ranging in size from 84 bp to 2.3 kb and introns ranging from 180 bp to 50 kb. All splice acceptor and donor sites conform to the consensus sequences including the AG-GT motif. Although PPARD lacks a TATA box, the gene is transcribed from a unique start site located 380 bp upstream of the ATG initiation codon. The 5' and 3' ends were mapped by rapid amplification of cDNA ends and the mRNA size of PPARD based upon the structure of the gene is 3803 bp. In addition, the chromosomal sublocalization of PPARD was determined by radiation hybrid mapping. The PPARD gene is located at 14 cR from the colipase gene and 15 cR from the serine kinase gene at chromosomal region 6p21.2. PMID- 10851271 TI - MHC-I expression in HTLV-1-positive and -negative cells. AB - The expression level of major histocompatibility class I (MHC-I) and the extent of down-regulation of MHC-I after an anti-MHC-I antibody treatment in numerous human T-cell leukemia virus type 1 (HTLV-1)-positive and -negative lymphocytic cell lines were examined. While there was no clear correlation between the expression level of MHC-I and the presence of HTLV-1 genome, a relatively low level of MHC-I down-regulation was generally induced in HTLV-1-positive cells by the antibody. The results may suggest the potential involvement of MHC-I in HTLV 1 leukemogenesis. PMID- 10851272 TI - Elongation factor-1alpha as a homologous complement activator of Jurkat cells. AB - Although increasing evidence exist suggesting that apoptotic cells activate homologous complement, the homologous complement activators are only poorly characterized. We found that cell lysate of Jurkat cells contained a homologous complement activator of 50 kDa. Digestion of the 50 kDa activator with lysylendopeptidase yielded peptide fragments, with sequences identical to those of EF-1alpha. The 50 kDa activator was removed by immunoadsorption with anti-EF 1alpha, suggesting that the 50 kDa activator is EF-1alpha. The homologous complement activation did not proceed with EGTA-serum. In addition, C4b, a fragment produced by activation of the classical or lectin pathways was found to bind with EF-1alpha. These results suggest that EF-1alpha activates the homologous complement through the classical or lectin pathway. PMID- 10851273 TI - AZT inhibition of the ADP/ATP antiport in isolated rat heart mitochondria. AB - In order to gain insight into the mechanism by which AZT affects mitochondrial metabolism in heart, leading to the ATP deficiency syndrome, the capability of AZT to affect certain mitochondrial translocators was checked in coupled mitochondria isolated from rat heart. AZT was found to strongly inhibit the ADP/ATP antiport, in a competitive manner (Ki 7 microM), as photometrically measured. Contrarily, the rate of the succinate/malate exchange via the dicarboxylate carrier, of the oxaloacetate uptake via the oxodicarboxylate carrier and of cis-aconitate uptake via the tricarboxylate carrier was found to be unchanged in the presence of AZT. PMID- 10851274 TI - Mutational analysis of OB gene in obese and type 2 diabetes affected subjects. AB - Peripheral blood DNA from 12 subjects affected by familial obesity and from 35 subjects affected by type 2 diabetes were analysed for mutations in the coding sequence of the OB gene. Mutational analysis, conducted using the single strand conformation polymorphism (SSCP) technique, followed by direct sequencing did not reveal the presence of nucleotide variants in the coding region of the OB gene. The lack of mutations in the coding sequence is consistent with previous data suggesting that mutations in the coding sequence of the OB gene are not common in human familial obesity. In 2 samples displaying a non-informative pattern of SSCP and in 8 additional samples the nucleotide sequence of portion of the intron 2 bordering the coding sequence of exon 2 identified a G in the positions +14IVS and +18IVS, according to a sequence reported previously, but in contrast with some others. All samples were homozygous for these intron variants. PMID- 10851276 TI - The backlog PMID- 10851275 TI - Modulation of endothelin-1 expression in pulmonary epithelial cell line (A549) after exposure to RSV. AB - Respiratory syncytial virus (RSV) is one of the most important respiratory tract pathogens in infants and young children. The airway epithelial cells are the primary target cells for RSV infection. The airway epithelial layer is not only a physical barrier, but also plays a role in a synthesis of a variety of major inflammatory cytokines (IL-6, IL-8, GM-CSF etc.) as previously reported. Endothelin-1 (ET-1) is a potent bronchoconstrictor and vasoconstrictor factor, and involved in pathogenesis of various diseases of the respiratory tract. We hypothesized that RSV may induce the release of ET-1 from the bronchial epithelial cell line. No previous data is available regarding association between RSV infection and ET-1 release. We evaluated the effect of RSV with different concentrations of RSV (MOI 0.1, 1 and 3 pfu/cell) on bronchial epithelial cell line (A549) and measured the production of ET-1 at both protein and mRNA level. A549 cells were treated with different conditions by using LPS, heat-inactivated RSV, RSV or medium alone as control. We observed time-dependent ET-1 release by RSV-infected A549 cells at 4 h, 24 h and maximum at 72 h. ET-1 was expressed in unstimulated A549 cells and was further increased by RSV. RSV with concentration MOI 0.1 (pfu/cell) and LPS appeared to have strongest stimulation on production of ET-1. In addition, ET-1 mRNA was increased significantly by 16 h and decreased to relatively low-level at 24 h. These experiments suggested that airway epithelial cells might play a role in the local airway smooth muscle tone through the production of endothelin-1 during RSV infection. PMID- 10851277 TI - Test and treat strategies for Helicobacter pylori in uninvestigated dyspepsia: a Canadian economic analysis. AB - BACKGROUND: Recognition of the pivotal role of Helicobacter pylori in the pathogenesis of peptic ulcer disease has revolutionized primary care approaches to dyspepsia. Decision analysis was used to compare the cost effectiveness of empirical ranitidine with a test and treat strategy using either H pylori serology or the 13carbon-urea breath test (13C-UBT). PATIENTS AND METHODS: A cohort of patients under age 50 years presenting with uninvestigated dyspepsia was evaluated. Three initial strategies were compared with respect to direct medical costs and effectiveness in curing H pylori-related ulcers - empirical ranitidine, H pylori serology and UBT. A one-year time horizon and third-party payer perspective were adopted in a Canadian health care setting. RESULTS: UBT was more costly than either serology or ranitidine but was the most effective strategy and required the fewest endoscopies. No strategy demonstrated dominance over another in the base case. The incremental cost effectiveness ratio (ICER) of serology versus ranitidine was $118/cure, and sensitivity analysis induced dominance of serology in several plausible scenarios. The baseline ICER of UBT versus serology was $885/cure but showed substantial variation in sensitivity analysis. Each ICER was highly sensitive to variation in the cost of the tests themselves. At a serology cost of $25, UBT became dominant when its cost fell to $39. CONCLUSIONS: In low risk patients with uninvestigated dyspepsia, testing for H pylori using serology appears to be economically attractive. 13C-UBT may be a cost effective alternative to serology if local conditions closely approximate the model parameters. Future changes in the costs of serology and 13C-UBT may determine the optimal approach. PMID- 10851278 TI - Insulin induces cation fluxes and increases intracellular calcium in the HTC rat hepatoma cell line. AB - BACKGROUND/AIMS: Rat hepatoma HTC cells were used to study conductive pathways implicated in insulin-induced cation and calcium influx into liver cells. METHODS: Membrane potentials and currents were measured by whole-cell patch clamp. Cytosolic calcium was measured using FURA-2 fluorescence. RESULTS: Insulin induced a gradual and reversible depolarization of 5.7+/-0.8 mV. Insulin-induced currents showed a linear slope conductance of 663 pS and a reversal potential of 17.9 mV. Ion substitution experiments showed that these currents were composed mainly of a nonselective cation component. In FURA-2 experiments, insulin caused a slow monophasic rise in HTC cell calcium, which depended on the presence of extracellular calcium. Insulin also induced significant increases of 1.58- and 1.54-fold in basal calcium influx when studied by external calcium withdrawal and readmission, or by the manganese quench method, respectively. Using the latter approach, we found that 100 microM gadolinium and 10 microM SKF96365 blocked the rise of the basal manganese quench rate induced by insulin whereas 100 microM verapamil was without effect. CONCLUSIONS: Insulin induces inward cation currents that depolarize HTC cell membrane potentials and participate in increased calcium influx. PMID- 10851279 TI - Prospective study of biliary strictures to determine the predictors of malignancy. AB - BACKGROUND: There have been few prospective studies regarding the investigation of biliary strictures, principally because of rapid technological change. The present study was designed to determine the sensitivity of various imaging studies for the detection of biliary strictures. Serum biochemistry and imaging studies were evaluated for their role in distinguishing benign from malignant strictures. METHODS: Thirty-one patients with suspected noncalculus biliary obstruction were enrolled consecutively in the study. A complete biochemical profile, ultrasound, Disida scan and cholangiogram (endoscopic retrograde cholangiopancreatography [ERCP] or percutaneous cholangiogram) were obtained at study entry. Stricture etiology was determined based on cytology, biopsy and/or clinical follow-up at one year. RESULTS: Twenty-nine of 31 patients had biliary strictures, of which 15 were malignant. The mean age of the malignant cohort was 73.9 years versus 53.9 years in the benign cohort (P<0.001). Statistically significant differences between the malignant and benign groups, respectively, were as follows: alanine transaminase 235.2 versus 66.9 U/L (P=0.004), aspartate transaminase 189.8 versus 84.5 U/L (P=0.011), alkaline phosphatase 840.2 versus 361.1 U/L (P=0.002), bilirubin 317.8 versus 22.1 micromol/L (P<0. 001) and bile acids 242.5 versus 73.2 micromol/L (P=0.001). Threshold analysis using receiver operative characteristic (ROC) curves demonstrated that a bilirubin level of 75 micromol/L was most predictive of malignant strictures. Intrahepatic duct dilation was present in 93% of malignant strictures versus 36% of benign strictures (P=0.002). Common hepatic duct dilation was less discriminatory (malignant 13.5 versus benign 9.6 mm; P=0.11). Ultrasound was highly sensitive (93%) in the detection of the primary tumour in the bile duct or pancreas, or in the visualization of nodal or liver metastases. In benign disease, ultrasound failed to detect evidence of intrahepatic or extrahepatic biliary dilation in most cases. Disida scans were not able to distinguish between malignant or benign strictures and could not accurately localize the level of obstruction. The sensitivity of Disida scan for the diagnosis of obstruction was 50%. Cholangiographic characterization of strictures revealed an equal distribution of smooth (eight of 13) and irregular (five of 13) strictures in the malignant group. Ten of 13 benign strictures were characterized as smooth. Malignant strictures were significantly longer than benign ones - 30.3 versus 9.2 mm (P=0.001). Threshold analysis using ROC curves showed that strictures greater than or equal to 14 mm were predictive of malignancy (sensitivity 78%, specificity 75%, log odds ratio 11.23). CONCLUSIONS: A serum bilirubin level of 75 micromol/L or higher, or a stricture length of greater than 14 mm was highly predictive of malignancy in patients with a biliary stricture. Ultrasound was useful in predicting malignant strictures by detecting either intrahepatic duct dilation or by visualizing the tumour (primary or metastases). Strictures with a 'benign' cholangiographic appearance are frequently malignant. Disida scan did not add additional information. ERCP is necessary to diagnose benign strictures, which tend to be less extensive at presentation. PMID- 10851280 TI - Measurement of gallbladder volume with ultrasonography in pregnant women. AB - Fasting and postprandial gallbladder volumes were investigated using ultrasonography in three groups (10 subjects in each) of healthy women: third trimester pregnant women, postpartum women up to 10 days after giving birth and nonpregnant controls. The scans were performed at 09:00 after a 12 h fast. After the basal measurement was taken, gallbladder volumes were rescanned in 15 min intervals for 60 mins. At the end of this period, all volunteers received a standard liquid test meal, and scans were performed again for 1 h. The mean basal gallbladder volume was 22.2+/-4.2 mL in the nonpregnant (control) group. In the third trimester group, the basal volume was 37.8+/-10.5 mL -70.5% higher than in the nonpregnant group (P<0.001). In the postpartum group, the mean basal volume was 37.9% lower (27.4+/-6.5 mL) than that of the third trimester group (P<0.02). This basal volume was 23.6% greater than that of the control group (P<0.05). After administration of a test meal, the postprandial gallbladder volumes decreased during the first few minutes compared with baseline values. The volumes decreased by 10.2% to 39.8% (23.5+/-7.3 to 34.0+/-10.2; P<0.01) in the third trimester group, by 14.9% to 43.2% (16.6+/-4.3 to 23.3+/-5.5; P<0.01, 0.001) in the postpartum group and by 19.2% to 51.6% (11.9+/-3.5 to 17.9+/-3.6; P<0.02, 0.05, 0.01, 0.001) in the control group. Postprandial mean gallbladder volumes of the third trimester (P<0. 02) and postpartum groups (P<0.02 to 0.01) were significantly different from those of the control group. In conclusion, incomplete emptying of the gallbladder after eating during the third trimester of pregnancy may contribute to cholesterol-gallstone formation, and pregnancy may thus increase the risk of gallstones. PMID- 10851281 TI - Achalasia: treatment options revisited. AB - The aim of all current forms of treatment of achalasia is to enable the patient to eat without disabling symptoms such as dysphagia, regurgitation, coughing or choking. Historically, this has been accomplished by mechanical disruption of the lower esophageal sphincter fibres, either by means of pneumatic dilation (PD) or by open surgical myotomy. The addition of laparoscopic myotomy and botulinum toxin (BTX) injection to the therapeutic armamentarium has triggered a recent series of reviews to determine the optimal therapeutic approach. Both PD and BTX have excellent short term (less than three months) efficacy in the majority of patients. New data have been published that suggest that PD and BTX (with repeat injections) can potentially obtain long term efficacy. PD is still considered the first-line treatment by most physicians; its main disadvantage is risk of perforation. BTX injection is evolving as an excellent, safe option for patients who are considered high risk for more invasive procedures. Laparoscopic myotomy with combined antireflux surgery is an increasingly attractive option in younger patients with achalasia, but long term follow-up studies are required to establish its efficacy and the potential for reflux-related sequelae. PMID- 10851282 TI - Sphincter of Oddi function and dysfunction. AB - The sphincter of Oddi (SO) is situated at the junction of the bile and pancreatic ducts where they enter the duodenum, and it serves to regulate the flow of bile and pancreatic juices as well as to prevent the reflux of duodenal contents into the pancreatobiliary system. SO dysfunction relates to either the biliary or pancreatic portions of the sphincter. Distinct clinical syndromes relating to either sphincter segment are recognized. The mechanism of dysfunction remains uncertain, but disruption of neural pathways involved in sphincter function seems likely. SO dysfunction is best diagnosed by manometry, which is able to correctly stratify patient groups and determine therapy. Biliary scintigraphy, which is noninvasive, has shown promise as a screening tool for patients with suspected SO dysfunction. Division of the sphincter is an effective treatment for patients with manometrically proven SO stenosis for either the biliary or pancreatic form of the disorder. Other forms of SO dysfunction may benefit from pharmacotherapy. PMID- 10851283 TI - Preservation and restoration of sphincter function in patients with rectal cancer. AB - Radical resection of rectal cancer is the standard treatment for curing this disease. Half of these tumours are located in the rectosigmoid region or the upper third of the rectum and are, therefore, easily resectable with preservation of the sphincter muscles, thus guaranteeing acceptable continence in most patients. However, tumours that originate in the lower parts of the rectum have been accompanied with the need for an abdominoperineal resection and the threat of a permanent colostomy. In the past 20 years, sphincter-saving surgery has become increasingly common in the treatment of tumours of the middle and low rectum due to the knowledge of tumour growth, the use of stapling devices, and the knowledge of the physiology of the pelvic floor and the sphincter muscles, respectively. Recent surgical techniques of resection of the 'ultralow' rectum (intersphincteric resection) and the reconstruction by coloanal anastomosis are reviewed. Functional problems following ultralow resections are emphasized, as well as the possibility of sphincter restoration after abdominoperineal resection by use of dynamic graciloplasty. Taking all surgical options into account, a permanent colostomy for rectal cancer can be avoided in most curatively and electively operated patients. PMID- 10851284 TI - Target proteins in human autoimmunity: cytochromes P450 and UDP- glucuronosyltransferases. AB - Cytochromes P450 (CYPs) and UDP-glucuronosyltransferases (UGTs) are targets of autoantibodies in several hepatic and extrahepatic autoimmune diseases. Autoantibodies directed against hepatic CYPs and UGTs were first detected by indirect immunofluorescence as antiliver and/or kidney microsomal antibodies. In autoimmune hepatitis (AIH) type 2, liver and/or kidney microsomal (LKM) type 1 autoantibodies are detected and are directed against CYP2D6. About 10% of AIH-2 sera further contain LKM-3 autoantibodies directed against family 1 UGTs. Chronic infections by hepatitis C virus and hepatitis delta virus may induce several autoimmune phenomena, and multiple autoantibodies are detected. Anti-CYP2D6 autoantibodies are detected in up to 4% of patients with chronic hepatitis C, and anti-CYP2A6 autoantibodies are detected in about 2% of these patients. In contrast, 14% of patients with chronic hepatitis delta virus infections generate anti-UGT autoantibodies. In a small minority of patients, certain drugs are known to induce immune-mediated, idiosyncratic drug reactions, also known as 'druginduced hepatitis'. Drug-induced hepatitis is often associated with autoantibodies directed against hepatic CYPs or other hepatic proteins. Typical examples are tienilic acid-induced hepatitis with anti-CYP2C9, dihydralazine hepatitis with anti-CYP1A2, halothane hepatitis with anti-CYP2E1 and anticonvulsant hepatitis with anti-CYP3A. Recent data suggest that alcoholic liver disease may be induced by mechanisms similar to those that are active in drug-induced hepatitis. Autoantibodies directed against several CYPs are further detected in sera from patients with the autoimmune polyglandular syndrome type 1. Patients with autoimmune polyglandular syndrome type 1 with hepatitis often develop anti-CYP1A2; patients with adrenal failure develop anti-CYP21, anti- CYP11A1 or CYP17; and patients with gonadal failure develop anti-CYP11A1 or CYP17. In idiopathic Addison disease, CYP21 is the major autoantigen. PMID- 10851285 TI - Achalasia: dilation, injection or surgery? AB - Achalasia results from irreversible alterations of the esophageal myenteric plexus. The target of treatment in this setting is to reduce lower esophageal sphincter resistance to passage of the bolus. Definitive treatment of the disease requires pneumatic dilation or Heller myotomy. Although no controlled studies comparing modern endoscopic and surgical techniques are available, laparoscopic surgery is emerging as the initial intervention of choice. PMID- 10851286 TI - Primary biliary cirrhosis and hemolytic anemia confusing serum bilirubin levels. AB - Hemolysis is observed in more than 50% of patients with cirrhosis. However, there has been little documentation of the association of primary biliary cirrhosis with autoimmune hemolytic anemia. Two cases, found within a single practice, of primary biliary cirrhosis coexisting with autoimmune hemolysis and a third case coexisting with hereditary spherocytosis are presented. Anemia in such patients is commonly attributed to chronic disease, and hyperbilirubinemia is attributed to primary biliary cirrhosis. These patients were considered for liver transplantation until the diagnosis of a comorbid hemolytic process was established. This association may be more prevalent than previously recognized. A diagnosis of comorbid hemolysis must always be considered in context with anemia and serum bilirubin levels that rise out of proportion to the severity of the primary biliary cirrhosis. PMID- 10851287 TI - Esophageal involvement in Wegener's granulomatosis: a case report and review of the literature. AB - Wegener's granulomatosis is characterized by a granulomatous arteritis involving the upper and lower respiratory tracts, progressive glomerulonephritis and systemic symptoms attributable to small vessel vasculitis. Although multisystemic manifestations are frequent, involvement of the gastrointestinal tract is uncommon. Cases have been reported of intestinal perforation, ulceration and hemorrhage. A patient whose initial presentation of Wegener's granulomatosis was odynophagia secondary to esophageal vasculitis is described. Endoscopy revealed multiple punched out ulcerations in the esophagus, which resolved with standard therapy for systemic Wegener's granulomatosis. There are only two previous reports of symptomatic esophageal vasculitis in patients with Wegener's granulomatosis. These reports illustrate the need to consider odynophagia as a reflection of disease activity as opposed to complications of immunosuppressive therapy. PMID- 10851288 TI - Nonceliac diaphragm disease of the duodenum. AB - A 58-year-old, Indo-Canadian man presented with upper abdominal pain and the clinical features of a partial, gastric outlet obstruction. Subsequent studies, including endoscopic examination, revealed diaphragm-like strictures in the descending duodenum. Other reported causes such as celiac disease and drug induced small bowel diaphragms were excluded. Possibly, the changes seen in this patient were related to ethnic food-induced, mucosal injury to the upper gastrointestinal tract. Further studies are needed to evaluate potential toxicity or protective effects of different ethnic diets and their relationship with the development of different intestinal diseases. PMID- 10851289 TI - Absence of HinfI Restriction Abnormalities in Renal Oncocytoma Mitochondrial DNA. AB - Renal oncocytomas are characterized by bland-appearing eosinophilic cells with a profusion of mitochondria. Previous work has suggested that these tumors possess a mutation in the 16.5-kbp circular mitochondrial DNA (mtDNA) manifested by an abnormal restriction fragment pattern after digestion with HinfI (Welter et al, Genes Chromosomes Cancer 1989;1:7-82). To better characterize this mtDNA abnormality in renal oncocytomas, we amplified the entire mitochondrial genome from five paired normal and oncocytoma specimens and subjected the amplified fragments to digestion with the restriction enzyme HinfI. No somatically acquired alterations were detected in the mtDNA from any of the five renal oncocytomas. One specimen displayed a known HinfI polymorphism in the mtDNA from both the normal and oncocytoma tissues. Our data do not support the existence of somatically acquired mitochondrial genome abnormalities in renal oncocytomas. PMID- 10851290 TI - Effects of Unilateral Ischemia on the Contractile Response of the Bladder: Protective Effect of Tadenan (Pygeum africanum Extract). AB - Recent studies indicate that localized (regional) ischemia may play an important role in the etiology of the contractile dysfunctions secondary to partial outlet obstruction. Pretreatment with Tadenan(r) (Pygeum africanum extract) has been shown to protect the rabbit bladder against the development of both contractile and biochemical dysfunctions induced by partial outlet obstruction, possibly by protecting the bladder against ischemic injury. The current study was designed to determine if Tadenan pretreatment of rabbits protected the bladder against the development of contractile dysfunctions induced by unilateral ischemia and to correlate effects in the presence and absence of Tadenan with the molecular response in relation to two of the most prominent genes activated by both ischemia and partial outlet obstruction: Hsp-70 and c-myc. Male New Zealand rabbits were separated into four groups of six rabbits each. Three rabbits of each group were treated for 3 weeks with Tadenan. The other three rabbits of each group were untreated controls. Each rabbit in Groups 2, 3, and 4 was anesthetized, and all major arteries entering on the right side of the bladder were li-gated. Rabbits in Group 1 were nonischemic. After 30, 60, and 120 minutes of unilateral ischemia, the rabbits in groups 2, 3, and 4, respectively, were euthanized. The bladders were removed, and contractility studies were performed on strips of detrusor muscle isolated from both the ischemia and nonischemic sides. The balance of the ischemic and nonischemic sides were frozen and stored in liquid nitrogen until analyzed for the expression of Hsp-70 and for c-myc. Tadenan pretreatment protected the nonischemic side of the bladder from the development of contractile dysfunctions, and unilateral ischemia resulted in a 10 fold increase in the expression of both Hsp-70 and c-myc in the bladders isolated from the Tadenan-treated rabbits. These results indicate that Tadenan has a protective effect against ischemic damage to the bladder. This protection may involve enhanced expression of Hsp-70 and other genetic factors. PMID- 10851291 TI - Mechanism of Brefeldin A-Induced Growth Inhibition and Cell Death in Human Prostatic Carcinoma Cells. AB - The mechanism of growth inhibition and triggering of cell death by the antibiotic brefeldin A (BFA) was investigated in human prostatic cancer DU-145 cells. After cells were cultured with various concentrations of BFA, cell number and viability were determined at specified times. Compared with untreated cells, a drastic growth reduction (>80%) with approximately 50% cell death was observed in the cells cultured with BFA (30 ng/mL) for 72 h. Cell-cycle analysis using flow cytometry revealed that such growth inhibition was associated with approximately 85% reduction in the S-phase population, indicating the inhibition of the G(1)-S phase progression. Western blots further showed that cell-cycle-dependent kinases (cdk2 and cdk4), cyclin D(1), and p53 were all downregulated, whereas WAF1 (p21) was upregulated with BFA treatment. Possible induction of apoptosis by BFA was also assessed by TUNEL assay and by DNA analysis using agarose gel electrophoresis. The TUNEL assay demonstrated the positive staining of BFA treated cells, and gel electrophoresis confirmed nucleosomal DNA ladder formation. Thus, these results suggest that growth inhibition of DU-145 cells by BFA is attributable mainly to a G1 cell-cycle arrest through the modulation of specific cell-cycle regulators. The accompanying cell death may follow a p53 independent apoptotic pathway. PMID- 10851292 TI - Protein Kinase C Mediates Prolactin Regulation of Mitochondrial Aspartate Aminotransferase Gene Expression in Prostate Cells. AB - Prolactin stimulates citrate accumulation in prostate cells by increasing the expression of mitochondrial aspartate aminotransferase (mAAT). In this study, we further investigated the mechanism of prolactin regulation of mAAT expression in rat lateral prostate and LNCaP and PC-3 prostate cancer cells. Prolactin and 12-O tetra-decanoylphorbol 13-acetate (TPA) increased the mAAT mRNA level twofold to fourfold. In addition, prolactin and TPA increased protein kinase C (PKC) activity in prostate cells 20% to 60% and 40% to 210%, respectively. The effects of both prolactin and TPA on mAAT mRNA were eliminated by downregulation of PKC. The effect of prolactin and TPA on gene transcription was determined using mAAT chloramphenicol acetyltransferase (CAT) reporter-gene constructs, transiently transfected into PC-3 cells. The 59 untranslated region of the precursor form (pmAAT) of the mAAT gene contains five sequences that are homologous to the consensus TPA response elements (TRE). Reporter constructs with various combinations of these sequences were used to assay prolactin stimulation of CAT transcription in PC-3 cells. Prolactin increased CAT expression in PC-3 cells transfected with a reporter gene containing four of the TRE consensus sequences. Another CAT reporter gene, which contained two of the putative TREs, was also stimulated by prolactin, but a third reporter, containing the two other TRE sequences, was not induced by prolactin. These results suggest that prolactin regulates mAAT at the transcriptional level. Moreover, because both prolactin and TPA induced PKC activity, and because the effects of prolactin and TPA were eliminated when PKC was downregulated, we postulate that the prolactin effect on mAAT expression is mediated via the diacylglycerol PKC signal transduction pathway in rat lateral prostate and human prostate cancer cells. PMID- 10851293 TI - Cryosurgical Modeling: Sequence of Freezing and Cytotoxic Agent Application Affects Cell Death. AB - Cryosurgery is now being used to eradicate a variety of tumors, including those that are found in the kidney. Because it is well known that cancer treatments are often most effective when combined with adjunctive therapies, we developed a model kidney cell culture system to determine: (1) if 5-fluorouracil (5-FU), a compound known to be a poor kidney tumor chemotherapeutic agent, can be more effective when combined with cryosurgery; and (2) how kidney cells die through freezing-induced injury that might occur during cryosurgery. The DNA isolated from kidney cells that died during an exposure to -15 degrees C showed nonrandom cleavage when separated on an agarose gel, indicating that cell death was attributable, in part, to apoptosis, whereas DNA isolated from kidney cells that died during exposure to -75 degrees C showed random cleavage, indicating that cell death was attributable, in part, to necrosis. The apoptotic protease inhibitor, caspase 1 inhibitor V, was able to prevent freezing-induced cell death, supporting the idea that apoptosis may be a mechanism of cell death in the periphery of the iceball created by cryosurgical procedures in vivo. Because 5-FU is known to induce apoptosis, this drug was used in combination with various freezing regimes to determine if the combination might be a better method of killing cells than either treatment alone. Whereas the addition of 5-FU at the same time or 2 days after freezing resulted in a synergistic lethal effect, many cells survived this combination treatment. When cells were treated with 5-FU 2 days prior to freezing, however, there was an apparent complete loss of viability. The reason may be that, by an unknown mechanism, freezing enhances the ability of 5-FU-treated cells to move more effectively through the apoptosis pathway. Kidney tumors may have robust antiapoptotic mechanisms that make them refractory to 5-FU, but these mechanisms might be overridden by freezing. In either case, the data suggest that chemotherapy may be more effective when followed by cryosurgery. PMID- 10851294 TI - Unsuspected Prostate Carcinoma and Prostatic Intraepithelial Neoplasm in Taiwanese Patients Undergoing Cystoprostatectomy. AB - Although the incidence and death rates for cancer of the prostate (CaP) in Taiwan have been among the lowest in the world, they have increased remarkably in recent years. Because of the very low autopsy rate in this country, prostate specimens obtained via cystoprostatectomy may provide a unique opportunity to study the incidence and status of latent cancer. From January 1992 to December 1997, 49 prostate specimens were obtained from patients with transitional-cell carcinoma of the urinary bladder (48 cases) or pelvic melanoma (one case). Patients' ages ranged from 47 to 89, with a mean age of 67.8 years. No patient had any clinical indication of CaP, as assessed by digital rectal examination. Each prostate was prepared with whole-mount transverse serial sections at 3-mm intervals from the apex to the bladder neck. The stained slides reviewed by two pathologists to evaluate the frequency and pathological status of acinar cancer lesions and high grade prostatic intraepithelial neoplasia (PIN). Of the 49 patients evaluated, 16 (33%) had evidence of adenocarcinoma, and 24 (49%) had high-grade PIN. The incidence of unsuspected CaP in patients aged 40 to 59, 60 to 69, and >/=70 years was 25%, 32%, and 37%, respectively. The frequency of high-grade PIN in patients aged 40 to 59, 60 to 69, and >/=70 years was 25%, 42%, and 64%, respectively. The incidence of high-grade PIN in the 16 patients with unsuspected CaP was significantly higher than in the 31 patients without this early cancer (75% v 36%). Of the 16 patients with unsuspected cancer, 5 had multiple tumors (3 patients with two and 2 with multiple foci). The mean volume of the 24 tumors was 0.0786 cm(3), with a range of 0.008 to 0.393 cm(3), but only 6 tumors exceeded 0.1 cm(3) in volume (0.112, 0.112, 0.164, 0.245, 0.262, and 0.393 cm(3)). Eighty eight percent of these early cancers were low grade (Gleason score 2-4). All unsuspected CaP were organ confined. The frequency of unsuspected CaP in Taiwanese men is relatively higher than in Chinese, as previous reported by Dr. Gu. However, the incidence of this latent cancer is comparable to that of U.S. men of the same age. These findings, together with the high incidence of high grade PIN, suggest that the initial step in the induction of CaP in indigenous Taiwanese is similar to that in U.S. men. The lower number of reports of CaP in Taiwan might be attributable to: (1) lower volume of latent cancer in the Taiwanese compared with U.S. men; (2) underestimation of the incidence rate of CaP in Taiwan; or (3) different genetic or environmental status leading to a different progression rate. PMID- 10851295 TI - Prognostic Role of in Vitro Survival Efficiency in Human Transitional-Cell Carcinoma. AB - The purpose of this study was to assess the correlation of in vitro growth features of transitional-cell carcinoma (TCC) specimens with the clinical behavior of the respective tumors. We also analyzed the impact of depth of tumor invasion, histologic differentiation, morphologic characteristics, and nuclear p53 accumulation of tumors on the in vitro survival efficiency of microtumor cultures and the significance of these factors in predicting recurrence and progression of bladder cancer. The tumor cell lines derived from surgical specimens were cultured at 37 degrees C in 5% CO(2) and constant humidity. Microtumor cultures were classified into three groups according to their in vitro lifespan. Our results indicate that higher survival efficiency implies a propensity for aggressive clinical behavior of the tumor in vivo. Factors that imply a poorer prognosis account for longer lifespans for microtumour cultures. These prognostic indicators are also associated with higher rates of recurrence and progression for tumors that exhibit higher survival efficiency in vitro. PMID- 10851296 TI - WT1 Gene Expression in Human Testicular Germ-Cell Tumors. AB - The Wilms' tumor 1 gene (WT1) is a tumor suppressor gene whose alterations are linked to the genesis of Wilms1 tumor. The gene is expressed in the urogenital organs and plays a key role in their development. Recent studies have shown that the WT1 gene product acts as a growth promoter of human leukemic cells. Because WT1 has been reported to be important in testicular development, we have investigated WT1 messenger RNA (mRNA) expression in testicular germ-cell tumors. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to examine the levels of WT1 mRNA in 34 patients with testicular germ-cell tumor, including 25 low-stage and 9 high-stage tumors. There were 23 seminomas and 11 nonseminomas. The WT1 mRNA was highly expressed in 6 of 9 high-stage lesions (67%) but only 5 of 25 low-stage cases (20%). A significant correlation was observed between the extent of WT1 mRNA expression and tumor stage (P = 0.017). There was no significant difference in WT1 mRNA expression between seminomas and nonseminomatous tumors. These results suggest that WT1 may be causal for the progression of testicular germ-cell tumors. PMID- 10851297 TI - Utility of Competitive Reverse Transcription-Polymerase Chain Reaction Analysis of Specific CD44 Variant RNA for Detecting Upper Urinary Tract Transitional-Cell Carcinoma. AB - Recently, we developed a novel molecular approach, CD44 v8-10/CD44 v10 competitive reverse transcription-polymerase chain reaction (CC-RT-PCR), to detect a sparse population of cancer cells overexpressing CD44v8-10 among a much larger population of nonneoplastic cells in body fluids by the measurement of the transcriptional level of CD44v8-10 relative to that of CD44v10. We have shown the utility of CC-RT-PCR in diagnosing disease using urine samples from patients with bladder cancer. In this study, we initially examined the expression of CD44 splice variants in human upper urinary tract transitional-cell carcinomas (UUT TCCs) and their adjacent normal urinary tissues by RT-PCR. Any CD44 variant isoforms were barely detectable in normal urinary tissues, whereas CD44v8-10 was predominantly expressed in 21 of the 25 UUT-TCC specimens (84%). We then applied CC-RT-PCR to spontaneously voided urine samples from 40 patients with UUT-TCC and 40 patients with benign urologic diseases. The CC-RT-PCR analysis revealed that all of the samples from patients with benign diseases presented a predominant expression of the CD44v10 transcript, whereas 26 of the 40 samples from patients with UUT-TCC dominantly expressed the CD44v8-10 transcript. In addition, the positive rate obtained by the CC-RT-PCR analysis was significantly higher than that obtained by cytologic examination, especially in patients with low-grade UUT TCC. These findings strongly suggest that CC-RT-PCR is a useful noninvasive tool for the diagnosis of UUT-TCC. PMID- 10851298 TI - Expression of Transforming Growth Factor-beta Receptors and Related Cell-Cycle Components in Transitional-Cell Carcinoma of the Bladder. AB - Exogenous transforming growth factor-beta (TGF-beta) is a potent inhibitor of normal epithelial cell growth but does not generally inhibit the growth of cell lines of transitional-cell carcinoma (TCC) of the bladder. In addition, a lack of the TGF-beta2 transcript and a marked reduction of the TGF-beta1 transcript have been reported in some high-stage TCCs. The purpose of this investigation was to examine the steady-state expression of TGF-beta receptor I and TGF-beta receptor II and a downstream target, p27(KIP1), as well as cyclin E in normal bladder and superficial and invasive TCC in order to better understand the role of TGF-beta downstream targets in TCC insensitivity to TGF-beta. Quantitative RT-PCR was employed to study the expression of TGF-beta receptor I and receptor II. p27(KIP1), and cyclin E in normal bladder and superficial and invasive TCC lesions. Steady-state levels of p27(KIP1), TGF-beta receptors, and cyclin E mRNAs were similar in superficial TCC samples and normal bladder mucosa. There was a significant decrease in p27(KIP1) and TGF-beta receptor II mRNA expression in invasive lesions compared with superficial tumors (P < 0.004; P < 0.02). In contrast, there was no significant difference in the expression of TGF-beta receptor I mRNA between normal bladder and superficial and invasive TCC. There was a significant increase in the expression of cyclin E mRNA in invasive TCC compared with superficial TCC or normal bladder (P < 0.015). These results suggest that aberrant expression of these genes contributes to the phenotype of invasive bladder cancer. PMID- 10851299 TI - Multifocal Sarcoma of the Urinary Bladder. AB - Sarcomas of the urinary bladder are rare and typically present as solitary, unifocal neoplasms. This is the second report of a primary multifocal sarcoma of the bladder. Like the first reported case, the tumor was highly aggressive with a short clinical course. The tumor apparently arose in stroma of the lamina propria and showed a variety of histologic patterns but without evidence of differentiation. PMID- 10851300 TI - Therapeutic potential of curcumin in human prostate cancer. II. Curcumin inhibits tyrosine kinase activity of epidermal growth factor receptor and depletes the protein. AB - PURPOSE: In a search for alternative and preventive therapies for prostate cancer, attention was focused on the ways in which curcumin (Turmeric), used in food and medicine in India for centuries, could interfere with the growth factor signaling pathways in both androgen-dependent and androgen-independent prostate cancer cells, as exemplified by the epidermal growth factor receptor (EGF-R) signaling. MATERIALS AND METHODS: The androgen-sensitive LNCaP and androgen insensitive PC-3 cell lines were grown in 5 to 50 microM curcumin and analyzed for EGF-R protein by Western blotting and for EGF-R tyrosine kinase activity. RESULTS: Curcumin was a potent inhibitor of EGF-R signaling, and it accomplished this effect by three different means (1) down regulating the EGF-R protein; (2) inhibiting the intrinsic EGF-R tyrosine kinase activity; and (3) inhibiting the ligand-induced activation of the EGF-R. CONCLUSIONS: These results, taken together with our previous results that curcumin can induce apoptosis in both androgen-dependent and androgen-independent prostate cancer cells, support our view that curcumin may be a novel modality by which one can interfere with the signal transduction pathways of the prostate cancer cell and prevent it from progressing to its hormone-refractory state. PMID- 10851301 TI - Induction of apoptosis in human prostatic cancer cells with beta-glucan (Maitake mushroom polysaccharide). AB - PURPOSE: To explore more effective treatment for hormone-refractory prostate cancer, we investigated the potential antitumor effect of beta-glucan, a polysaccharide of the Maitake mushroom, on prostatic cancer cells in vitro. MATERIALS AND METHODS: Human prostate cancer PC-3 cells were treated with various concentrations of the highly purified beta-glucan preparation Grifron-D(R) (GD), and viability was determined at 24 h. Lipid peroxidation (LPO) assay and in situ hybridization (ISH) were performed to unravel the antitumor mechanism of GD. RESULTS: A dose-response study showed that almost complete (>95%) cell death was attained in 24 h with GD > or = 480 microg/mL. Combinations of GD in a concentration as low as 30 to 60 microg/mL with 200 microM vitamin C were as effective as GD alone at 480 microg/mL, inducing >90% cytotoxic cell death. Simultaneous use with various anticancer drugs showed little potentiation of their efficacy except for the carmustine/GD combination (approximately 90% reduction in cell viability). The significantly (twofold) elevated LPO level and positive ISH staining of GD-treated cells indicated oxidative membrane damage resulting in apoptotic cell death. CONCLUSION: A bioactive beta-glucan from the Maitake mushroom has a cytotoxic effect, presumably through oxidative stress, on prostatic cancer cells in vitro, leading to apoptosis. Potentiation of GD action by vitamin C and the chemosensitizing effect of GD on carmustine may also have clinical implications. Therefore, this unique mushroom polysaccharide may have great a potential as an alternative therapeutic modality for prostate cancer. PMID- 10851302 TI - Adverse effects of oxidative stress on renal cells and its prevention by antioxidants. AB - BACKGROUND AND PURPOSE: Recent reports suggest that reactive oxygen species; e.g., hydrogen peroxide (H(2)O(2)), could be the primary cause of various drug induced renal injuries. We investigated the effects of H(2)O(2) on renal cells to understand its mode of action and to explore cytoprotection from such a fatal injury. MATERIALS AND METHODS: Renal proximal tubular LLC-PK(1) cells were exposed to various concentrations of H(2)O(2), and cell viability was determined at specified times. Lipid peroxidation assay and Western blot analysis of heat shock proteins (Hsp70 and Hsp90) were performed to assess the cellular effects. RESULTS: The dose-response study showed that H(2)O(2) > or = 100 microM was severely cytotoxic. Even a 1-h exposure was sufficient to induce >95% cell death in 24 h. Lipid peroxidation was significantly (>50%) increased, while Hsp90, but not Hsp70, was partially degraded, to an approximately 85-kDa fragment, after a 3 h H(2)O(2) exposure. However, such cytotoxic cell death was remarkably ( approximately 90%) prevented by the antioxidants pyruvate or N-acetylcysteine (NAC), and Hsp90 remained intact. CONCLUSION: Hydrogen peroxide-induced renal cell death involves increased lipid peroxidation and partial degradation of Hsp90. Both pyruvate and NAC are capable of detoxifying H(2)O(2) to maintain cell viability and Hsp90 integrity. Acute renal injuries associated with oxidative stress might preventable by appropriate antioxidants. PMID- 10851303 TI - Nephron-sparing surgery is still controversial for patients with renal cell carcinoma and normal contralateral kidney: risks predictable by AgNOR counts in satellite lesions. AB - PURPOSE: To determine whether nephron-sparing surgery is appropriate for patients with renal cell carcinoma (RCC) and a normal contralateral kidney. MATERIALS AND METHODS: We prepared whole-area histologic sections from 58 cases of RCC to examine the features of satellite tumor lesions (STLs), the proliferative potential of which was evaluated by counting argyrophilic nucleolar organizing regions (AgNORs). RESULTS: We found three types of microscopic lesions: STLs, adenomas, and dysplastic foci, the latter two of which appeared to be preneoplastic. Of the 58 cases, STLs were observed in 27 (47%) and either adenomas or dysplastic foci in 19 (33%). At least one of the three types of lesion was found in 37 cases (64%). No correlation was found between the incidence of STL and the size of the main tumor, but the incidence tended to be higher in lesions of higher grade or stage. The STLs were located >2 cm from the margin of the main tumor in 6 of the 27 cases (22%). The mean number of AgNORs per high-power field was 5.09 +/- 1.53 (SD) in the main tumors, 4.21 +/- 1.36 in the STLs, and 2.27 +/- 0.07 in the adenomas and dysplastic foci. CONCLUSIONS: These findings suggest that nephron-sparing surgery risks leaving STLs, which have considerable proliferative potential. Thus, nephron-sparing surgery must still be considered adventurous treatment in elective cases but can be recommended for patients who require nephron preservation. PMID- 10851304 TI - Zinc induces mitochondria apoptogenesis in prostate cells. AB - BACKGROUND AND PURPOSE: Prostate secretory epithelial cells have the unique function and capability of accumulating extremely high intracellular levels of zinc. One of the effects of this accumulation is inhibition of cell growth due, in part, to an increase in apoptosis. The present studies were conducted to determine if this zinc-induced apoptosis involves stimulation of mitochondrial apoptogenesis. MATERIALS AND METHODS: The PC-3 a human malignant prostate cell line, which is zinc accumulating, was exposed to medium supplemented with physiologic levels of zinc. RESULTS: By 24 h, zinc treatment resulted in the translocation of cytochrome c from the mitochondria to the cytosol, the activation of caspase-9 and caspase-3, and eventually, the cleavage of nuclear poly(ADP)-ribose polymerase (PARP). Under similar conditions, exposure of freshly prepared rat ventral prostate cells (which are also zinc accumulating) resulted in increased apoptosis following translocation of cyochrome c and activation of caspases-9 and 3. The human prostate PZ-HPV-7 cells, which do not accumulate zinc, did not exhibit any apoptotic effect from zinc treatment. CONCLUSION: The accumulation of high intracellular levels of zinc by prostate cells induces mitochondrial apoptogenesis. This represents a newly identified physiological effect of zinc in the regulation of prostate cell growth. PMID- 10851305 TI - Central Nervous System Innervation of the Penis, Prostate, and Perineal Muscles: A Transneuronal Tracing Study. AB - The spinal and brain neurons that innervate the male sex organs (penis, perineal muscles, and prostate) were identified using the transneuronal tracing technique. Rats were anesthetized with ketamine/xylazine, and pseudorabies virus (PRV) was injected into either the corpus cavernosum, ventral prostate, or bulbospongiosus or ischiocavernosus muscle. After a 2- to 5-day survival period, rats were perfused transcardially under deep anesthesia, and viral-labeled neurons were identified using immunohistochemistry methods. After a 2- to 3-day survival, prostate or corpus cavernous injections resulted in labeling of sympathetic and parasympathetic preganglionic neurons. Spinal pudendal motoneurons were found to be labeled after perineal muscle injections. Spinal interneurons were found in the medial gray in the dorsal gray commissure and in the lateral gray of T13-S1 after injection at all sites. In the brain, the regions consistently labeled after all injections were the ventrolateral medulla, raphe magnus, raphe pallidus, A5, Barrington1s nucleus, central gray, and paraventricular nucleus of the hypothalamus. Therefore, there appears to be a large degree of overlap of the spinal and brain circuits that regulate the sex organs. PMID- 10851306 TI - Nitric Oxide and the Cyclic GMP System in the Penis. AB - Since the discovery that nitric oxide is the mediator of penile erection, considerable evidence has been accumulated in experimental animals and men regarding the pharmacology and molecular biology of the nitric oxide/cGMP pathway in the penis. This review examines the expression and tissue localization of nitric oxide synthase isoforms (NOS) in this organ and the functional significance of their variants. The disregulation of the balance between nitric oxide synthesis and the compliance of the corpora cavernosa smooth muscle in relation to erectile dysfunction in animal models is discussed. The possible up regulation of NOS levels and the interaction of NOS variants with protein modulators of their activity is analyzed in the context of novel concepts of gene therapy of erectile dysfunction. PMID- 10851307 TI - Ion Channels and Gap Junctions: Their Role in Erectile Physiology, Dysfunction, and Future Therapy. AB - A flurry of research and clinical activity during this past decade has documented that the tonicity and synchronicity of the corporal smooth muscle cells of the penis are major determinants of erectile capacity and function. Specifically, the effects of diverse and bifurcating intracellular signal transduction pathways on the activity of nonjunctional ion channels such as potassium (K(+)), calcium (Ca(2+)), and chloride (C(1-)) govern the former, whereas intercellular communication through gap junctions provides the anatomic substrate for the latter. Recent studies at the tissue, cellular, subcellular, and molecular levels have verified this supposition and provided important insight into how subtle alterations in the balance between contraction and relaxation of the corporal smooth muscle cells can predispose a man to erectile failure. This report reviews the available information concerning the participation of gap junctions and K(+), Ca(2+), and C(1-) channels in the erectile process and describes their importance as potential molecular targets for the future therapy of erectile dysfunction (ED). It is argued that a major goal should now be to proceed on at least two fronts simultaneously: (1) to capitalize on these new mechanistic insights by developing novel treatments for ED centered on the modulation of ion channel activity; and (2) simultaneously to take advantage of the unique therapeutic opportunities afforded by the presence and ubiquitous distribution of gap junction channels in the human corpora. One strategy that fulfils both criteria will be briefly reviewed, that is, gene therapy with the maxi-K(+) channel subtype. PMID- 10851308 TI - The Role of Andorgens in the Erectile Response: A 1999 Perspective. AB - Evidence from several laboratories strongly supports a critical role for androgens in the maintenance of the mammalian erectile response. In animal studies, androgens appear to act at the end-organ level (i.e., corporal tissue and vasculature), as well as in the portions of the nervous system which mediate erection. Particularly in the rat model, androgens act centrally to support copulatory behavior and peripherally to maintain the production of nitric oxide and support the veno-occlusive mechanisms. Other studies suggest that alternative, non-NO-dependent, pathways may also be androgen sensitive. However, despite this expanding knowledge base about how androgens act in the erectile response in laboratory animals, the recent studies have not greatly clarified the role of androgens in human penile erection. There does not seem to be a strong cause and effect relation between blood androgen concentrations and erectile function; even in severely hypogonadal men, the erectile response is not always lost, and testosterone treatment of hypogonadal men with erectile dysfunction does not necessarily restore lost erectile function. In addition, different types of erection (nocturnal, in response to visual sexual stimulation, in response to sexual partner) may require different degrees of androgenic support. PMID- 10851309 TI - Sexual Endocrinopathies. AB - There is compelling experimental and clinical evidence that for adequate sexual function, there must be complex and efficient cross-communication between the endocrine and nervous systems, both centrally and peripherally. The central nervous system (CNS) controls the release of sex steroids, while at the same time, these hormones are primary modulators of neurologic activity. The investigation and treatment of men suspected of hypogonadism is simple, but careful monitoring is essential. Unfortunately, sexual dysfunction is frequently multifactorial, and in many patients, the dysfunction does not improve with hormonal replacement. However, hypotestosteronemia is associated with a variety of alterations in different organ systems beyond sexual activity. It is important to the practice of urology to be familiar with neuroendocrine interactions and to maintain an adequate knowledge of the evaluation and treatment of hormonal deficiencies, particularly in men beyond middle age, in whom hypogonadism is prevalent. PMID- 10851310 TI - Pathophysiology of Erectile Dysfunction. AB - Physiology of erection and pathophysiology erectile dysfunction is reviewed. Analysis is obtained from basic and clinical research including animals studies, anatomical studies, and molecular and cellular research on corporal tissue obtained during penile prosthesis implantation. Supraspinal influences and spinal influence on penile erection has been learned from spinal cord injury patient. Corporal smooth muscle relaxation of penile arteries and corpus cavernosum leads to penile erection, results from parasympathetic/nonadrenergic noncholinergic neural pathway activation and simultaneous inhibition of sympathetic outflow. Anatomical studies taught understanding of the mechanism for restriction of blood outflow from the corpora cavernosa. The change of smooth muscle tone has emerged as a key factor in erection and detumescence. Many independent factors converge on the modulation of corporal smooth muscle tone. Neuronal and local neurotransmitter effects via gap junction, potassium channels, and calcium channel. A nitric oxide/cyclic guanosine monophosphate mechanism as well as cyclic aminomonophosphate has an important role in mediating the corporal smooth muscle relaxation necessary for erectile function. Erectile dysfunction can be due to vasculogenic, neurogenic, hormonal, veno-occlusive, psychogenic and/or pharmacogenic factors as well as alterations in the nitric oxide/cyclic guanosine monophosphate (cGMP) or cyclic aminophosphate (cAMP) pathway or other regulatory mechanisms including gap junction or ionic channel resulting in an imbalance in corporal smooth muscle contraction and relaxation. Our present knowledge of the hemodynamics, functional anatomy, neurophysiology, and neuropharmacology of penile erection and dysfunction at the cellular and molecular level has led to better understanding of physiology and pathophysiology of erectile dysfunction. PMID- 10851311 TI - Loss of TGFbeta, Apoptosis, and Bcl-2 in Erectile Dysfunction and Upregulation of p53 and HIF-1alpha in Diabetes-Associated Erectile Dysfunction. AB - Vasculogenic erectile dysfunction (ED) is associated with collagen replacement of the cavernosal smooth muscle, mediated by an increase in transforming growth factor (TGF)-production secondary to hypoxemia. We tested the hypothesis that human ED is the result of an increase in apoptosis of the cavernosal smooth muscle cells with replacement by collagen, mediated by the TGFbeta upregulation. We also examined the tissue for proteins associated with apoptosis. Human cavernosal tissue was procured from impotent men at the time of prosthesis insertion. Normal corpous cavernosum served as a control. The TUNEL assay was used to assess apoptosis. Immunohistochemistry staining was used to detect TGFbeta and Bcl-2 expression, while Western blot analysis was used to detect expression of Bcl-2, p53, and hypoxia-inducible factor (HIF)-1a. Immunohistochemistry showed downregulation of TGFbeta protein expression in the corpus cavernosum of men with ED. Apoptotic nuclei were noted in cavernosal smooth muscle from a potent man but were not found in cavernosal tissue from men with ED. To gain insight into the possible mechanism of apoptosis in men with ED, the proto-oncogene Bcl-2, a potential inhibitor of apoptosis, was examined. Both immunohistochemistry and Western analysis revealed the presence of Bcl-2 in the cavernosal nerve of a potent man but its absence in cavernosal tissue from men with ED. Thus, loss of Bcl-2 expression correlated with the loss of apoptosis. In contrast, Western blotting demonstrated upregulation of p53 and HIF-1a expression in the cavernosal tissues from the men with ED and diabetes. Male ED follows an active process characterized by a loss of TGFb expression, apoptosis, and Bcl-2 expression. However, there is upregulation of p53 and HIF-1a in men with diabetes. These data support the possibility of hypoxia-mediated ED in diabetes via upregulation of p53 and HIF-1a but does not substantiate a role for TGFbeta in ED. PMID- 10851312 TI - Changes in Penile Morphometrics in Men with Erectile Dysfunction after Nerve Sparing Radical Retropubic Prostatectomy. AB - There have been anecdotal reports of a decrease in penile size in men with erectile dysfunction (ED) after nerve-sparing radical retropubic prostatectomy (NSRRP). Penile circumference and length measurements are obtained by one physician from 100 men, age 47 to 74, who presented at various intervals (1.7&endash;27.6 months) for the treatment of ED after NSRRP from 1994 through 1997. All patients were asked to complete a brief male sexual function inventory at their initial visit. Penile measurements were obtained both in the flaccid and erect states, with erections being induced with intracorporal injections of Trimix. The sexual inventory scores were compared with those of an age-matched control cohort of 130 men presenting for evaluation of ED during the same time period and 132 age-matched men who completed the inventory at the time of a prostate screening. By self-report, men experiencing ED after NSRRP had better libido but more severe ED than men presenting with ED of other causes. There was a decrease in all penile dimensions after NSRRP. The flaccid and erect measurements of length and circumference decreased 8% and 9%, respectively after surgery (p > 0.05). The most substantial change occurred between the first 4 and 8 months postoperatively. The average change in volume between the first 4 and 8 months was 19% to 22% in the flaccid and erect state, respectively. There is a significant decrease in penile size in men with ED after NSRRP. The etiology may be denervation smooth muscle atrophy, apoptosis, or hypoxia-induced damage to the corpora. Further research is needed to elucidate the nature of these postoperative changes. PMID- 10851313 TI - Experimental Chronic Renal Failure-Associated Erectile Dysfunction: Molecular Alterations in Nitric Oxide Synthase Pathway and IGF-I System. AB - Chronic renal failure causes wide-ranging disturbances in male erectile function, and the dysfunction usually is not corrected by hemodialysis. In this study, erectile function and expression of nitric oxide synthase (NOS) isoforms, insulin like growth factor-I (IGF-I), and IGF-I binding proteins were assessed in rats in which uremia had been produced by 5/6 nephrectomy. In the animals that suffered renal failure, there was a significantly lower rise in intracavernosal pressure in response to electrical stimulation, and the mean patency period after cavernous nerve stimulation was significantly increased. The amount of neuronial NOS mRNA was significantly higher in the penile tissues and major pelvic ganglia (MPG) of uremic rats than in control animals. There consistently were higher levels of nNOS and endothelial NOS in the penile tissues and MPG of rats with renal failure, and there was a significant decrease in the amount of IGF-I gene expression in the MPG of these animals. Expression of the IGF binding proteins 2 and 4 but not 5 also was reduced. This preliminary work demonstrates that impairment of erection in chronic renal failure in the rat is attributable to a disturbance in NOS gene expression with concomitant changes in IGF-I. These discoveries may permit an improved therapeutic approach to men with erectile dysfunction associated with chronic renal failure. PMID- 10851314 TI - Sildenafil Citrate (Viagra(R)): An Update. AB - Sildenafil has had a revolutionary impact on the medical, societal, and economic aspects of erectile dysfunction. The author reviews the clinical trial and field experience with respect to the efficacy and safety of this drug. It is clear that early in the new millenium, erectile dysfunction will not only be effectively treated, it may even be prevented. PMID- 10851315 TI - Transurethral Therapy for the Treatment of Erectile Dysfunction: Infant or Dinosaur? AB - Transurethral pharmacotherapy (TUPT) for the treatment of patients with erectile dysfunction has demonstrated moderate efficacy with an excellent safety profile. Unfortunately, the initial clinical efficacy of transurethral alprostadil (TUA) has led to physician and patient dissatisfaction with this treatment regimen. An understanding of the mechanism of TUPT, and a review of the available literature, suggests that improvements in treatment efficacy and local side effects are in the near future. PMID- 10851316 TI - Penile Prostheses in the Sildenafil Era. AB - The first effective treatment for physical erectile difficulties occurred with the introduction of penile implants more than 25 years ago. The market share these devices hold in the treatment of erectile dysfunction (ED) has declined steadily as less aggressive treatments have been introduced. However, there remains a subset of patients who do well with implants, which includes those with penile scarring and the nonresponders to more conservative treatments. Two main categories of implants are available: the semirigid rods and the inflatable devices. The semirigid rods are inserted easily and are positionable with minimal effort. However, they give a permanently rigid penis and may be difficult to conceal during daily activities. The inflatable models are more challenging to insert but offer a more natural appearance and function to the penis. Repair rates for these devices have been reasonable, in the range of 5% to 10% in the first 5 years. Of all the treatments of EDs patient and partner satisfaction has been the highest with penile implants, in the range of 80% or more. These devices provide a predictable and reliable method, usable on demand, for restoring erectile function. PMID- 10851317 TI - Differential Glycosylation of Cellular Prostate Specific Antigen and the Regulatory Mechanism of Its Secretion. AB - Although serum prostate specific antigen (PSA), derived from cellular PSA through secretion, is widely used as a marker for prostate cancer (CAP), the exact regulatory mechanism of its secretion is not fully understood. To explore the regulation of serum PSA concentration, we examined whether the glycosylation state of cellular PSA might be associated with its secretion, thus determining its serum concentration. Blood and prostate tissue specimens were obtained from patients undergoing radical prostatectomy. Following preparation of cell extracts by tissue homogenization, the concentrations of serum and cellular PSA were determined using the Tandem-E PSA kit. The extent of cellular PSA glycosylation was then assessed by Western blot and affinoblott analyses. Neither serum nor cellular PSA concentrations correlated with the Gleason scores. Similarly, no direct relation between serum and cellular PSA levels was observed. However, the Western blots showed that the cellular PSA proteins were converted to the deglycosylated forms with glycosidase treatment, indicating differential glycosylation of cellular PSA. Affinoblotting further revealed that the various amounts of PSA glycosylation were associated wtih the serum PSA levels, with an inverse correlation between serum PSA and cellular PSA glycosylation: the greater the PSA glycosylation, the lower the serum PSA, and vice versa. The present study demonstrates that cellular PSAs in CAP specimens are differentially glycosylated and that such difference correlates well with the serum PSA concentration. Therefore, the concentrations of serum PSA appear to depend in part on a selective secretion of cellular PSA, which could be regulated primarily by its glycosylation state. PMID- 10851318 TI - Introduction of Human Chromosome 13 into Retinoblastoma-Negative Metastatic Human Prostate Cancer Cells Increases Their Sensitivity to Growth Inhibition by Transforming Growth Factor-&be;1. AB - Like many other carcinomas, prostate cancer develops resistance to inhibition by transforming growth factor (TGF)-&be;1 during oncogenesis. One proposed mechanism of TGF-&be;1 action posits action of the retinoblastoma protein (pRb) to suppress c-myc transcription to inhibit cellular proliferation. A metastatic human prostate cancer cell line, DU145, has both nonfunctional pRb and markedly reduced sensitivity to TGF-&be;1 growth inhibition. The defective rb gene in DU145 cells was replaced by a normal rb allele by microcell fusion of chromosome 13. Two subclones, DU145-Cl-I and DU145-Cl-II, were studied in vitro to determine whether the pRb restoration increased sensitivity to the inhibitory effects of TGF-&be;1. By reverse transcriptase-polymerase chain reaction, increased sensitivity to the inhibitory effects of TGF-&BE;1. By reverse transcriptase-polymerase chain reaction, parental DU145 cells had TGF-b receptors of Type I and Type II. Introduction of chromosome 13 reduced the growth rate and prolonged the G1 phase compared with the parental DU145 cell line. Moreover, responsiveness to TGF-&be;1 growth inhibition was restored in a dose-dependent manner. Transcription of c-myc was not altered by TGF-&be;1 growth inhibition. Thus, DU145 cells presumably required the presence of wildtype rb to become growth inhibited that is independent of c-myc transcription. As the entire chromosome 13 was introduced, unknown tumor suppressor genes, not only rb, may be responsible for the restoration of TGF-&be;1 growth inhibition. PMID- 10851319 TI - MUC1 Expression in Prostate Carcinoma: Correlation with Grade and Stage. AB - Mucins have been implicated in the biologic behavior and progression of several types of cancer. The aims of this study were to define the expression pattern of one particular mucin, MUC1, in benign and malignant human prostate tissue and to determine if MUC1 expression correlates with tumor grade and stage. Immunohistochemical staining utilizing an anti-MUC1 monoclonal antibody was performed on 4 fetal prostates, 4 specimens of benign prostatic hyperplasia (BPH), and 34 radical prostatectomy specimens. In human fetal and BPH specimens, there was an apical pattern of MUC1 expression, similar to that reported in other normal and benign tissues. Ninety-four percent of the prostate cancers were MUC1 positive. A high percentage of prostate cancer specimens (62%) demonstrated a diffuse, cytoplasmic staining pattern. There was a statistically significant correlation between diffuse MUC1 staining and Gleason pattern, with a diffuse/total staining percentage of 9% in Gleason 2, 64% in Gleason 3, 80% in Gleason 4, and 100% in Gleason 5. More diffuse staining was also seen in samples from patients with high pathologic stage: 21% in T(2), 75% in T(3), and 67% in N(1) disease. These data indicate that MUC1 expression is prevalent in prostate cancer and that diffuse cytoplasmic staining correlates with advanced Gleason pattern and advanced pathologic stage. PMID- 10851320 TI - Fourth International Conference on Neoadjuvant Hormonal Therapy of Prostate Cancer: Overview Consensus Statement. PMID- 10851321 TI - Hormonal Therapy in the Management of Prostate Cancer: An Historical Overview. AB - This article reviews the history of hormonal therapy for prostate cancer, beginning with the studies of Huggins and Hodges completed in the 1940s. Although early clinical reports suggested that major improvements and even cure could be obtained, later randomized investigations showed that hormonal treatments were palliative rather than curative. Later investigators demonstrated that prostate cancer is under the trophic influence of male hormones and that ablation of androgens could cause cancer regression. More recently, neoadjuvant and adjuvant hormonal therapy has been shown to improve outcomes for higher-risk patients who receive radiation as definitive local therapy. Numerous studies have attempted to devise hormonal therapy regimens that decrease the adverse physiologic consequences of the currently existing agents and to define the patient population and stage of prostate cancer that most benefit from the use of hormonal therapy, either alone or in association with additional agents. New hormonal agents currently in clinical trials may increase the options available for patients who have metastatic cancer or are at increased risk of recurrence after surgery or radiation. PMID- 10851322 TI - Who Needs to be Treated? AB - Controversy surrounds the appropriate treatment of clinically localized prostate cancer. Physicians debate not only which treatment is best but also who needs to be treated. Men at high risk of developing symptomatic metastatic disease are clearly in need of therapy designed to relieve symptoms and, if possible, prolong life. Risk factors that predict disease progression include high-grade tumor (Gleason score 7-10), tumor volume >0.5 cc, younger age at diagnosis (<72 years), and the absence of competing medical hazards. These are the men destined to die from prostate cancer and who should be enrolled in randomized studies designed to optimize treatment strategies. PMID- 10851323 TI - Pathologic Handling and Reporting of Prostate Tissue Specimens in Patients Receiving Neoadjuvant Hormonal Therapy: Report of the Pathology Committee. AB - Prior meetings of the International Conference on Neoadjuvant Hormonal Therapy in Prostate Cancer identified areas of concern in the pathologic analysis of post treatment specimens. In response, a pathology subcommittee was formed to address the questions that had been raised. Recommendations are made here concerning pretreatment biopsy review and reporting, handling and reporting of radical prostatectomy specimens, Gleason scoring of treated cancer, grading of therapy effect, role of immunohistochemistry procedures, and the significance of high grade prostatic intraepithelial neoplasia. Neoadjuvant trials with both surgery and radiation therapy were considered in drawing up these recommendations. PMID- 10851325 TI - PSA Control Following Definitive Therapy for Clinically Localized Prostate Cancer. AB - The two goals of treatment for clinically localized prostate cancer are disease control and a good quality of life. At present, the options are radical surgery, conformal or conventional external-beam radiation, and interstitial radiation. This article reviews the effectiveness of these techniques, as judged by serum prostate specific antigen monitoring, with special emphasis on magnetic resonance guided interstitial brachytherapy. PMID- 10851324 TI - Neoadjuvant Hormonal Therapy Prior to Radical Prostatectomy. AB - The US T(2b) study of 3 months of neoadjuvant hormonal therapy (NHT) showed a reduction in the likelihood of positive margins from 48% (control group) to 18% in the treated patients. Follow-up at 42 months shows that the cumulative relapse rate (prostate specific antigen) for 129 patients having NHT was not statistically different from that of the 126 control patients. Because the T(2b) study has been criticized for lacking central pathology review, we present a review of a series involving only one surgeon (MS) and one pathologist (FC) of NHT plus prostatectomy (109 patients) v prostatectomy alone (145 patients) with 24 months' follow-up. Positive margins were decreased from 38% in the untreated to 28% in the treated group, the only statistically significant difference in the results. Biochemical recurrence (PSA >0.2 ng/mL) was higher in the treated group, reflecting selection of more aggressive tumors for NHT, but the difference was not statistically significantly. The incidence of extracapsular extension, seminal vesicle invasion, and lymph node metastasis was similar in the two groups. The largest nonrandomized experience with NHT shows a decrease in the incidence of positive surgical margins when used in high-risk patients with clinically localized carcinoma of the prostate. However, it does not have an impact on disease-free survival at a mean 24-month follow-up. PMID- 10851326 TI - Long-Term Combined Androgen Blockade Alone for Localized Prostate Cancer. AB - The effect of long-term continuous combined androgen blockade (CAB) administered alone has been studied in 26 patients with stage B(2)/T(2) and 115 with stage C/T(3) prostate cancer who did not accept or were not candidates for other forms of therapy. In the 26 patients with stage T(2) disease who have received continuous CAB with an LHRH agonist and flutamide, progression of cancer, as evidenced by rising serum prostate specific antigen (PSA), was observed in only one patient receiving CAB, occurring after 7.3 years of continuous CAB treatment. Treatment with CAB was then stopped in 20 patients with stage T(2) cancer after a median period of 7.2 years. Failure or PSA rise occurred in 2 of these patients after a median follow-up of 3.1 years following cessation of CAB, and one died from prostate cancer. The benefits observed with CAB alone in localized disease compare favorably with those obtained by radical prostatectomy or radiotherapy alone. Twenty-six patients with stage T(3) cancer have been treated with CAB continuously for a median of 9.9 years. After a median follow-up of 4.4 years (range 0.3-7.0 years) after cessation of CAB in this group of patients, PSA failure was observed in 5 patients (19%), who had been treated continuously for 3.8, 5.0, 5.0, 9.9, and 10.9 years. Thus, in 85% (39 of 46) patients with stage T(2)-T(3) disease who were treated continuously with CAB for a median of 7.2 and 9.9 years, respectively, PSA remained undetectable up to a median of 3.1 (stage T(2)) and 4.4 (stage T(3)) years of follow-up. The present data, while showing the high efficacy of CAB in localized prostate cancer, clearly indicate the need for long-term treatment, similar to the 5 years of tamoxifen required for control of breast cancer. PMID- 10851327 TI - Hormonal Therapy and Radiation Therapy in the Treatment of Locally Advanced Prostate Cancer. AB - Adjuvant hormonal therapy with radiation may improve local control or survival in patients with locally advanced prostate cancer, as shown by EORTC 22863 and RTOG 85-31. Another trial, RTOG 86-10, showed better local control with adjuvant hormonal therapy and radiation but no difference in survival from patients receiving radiation alone. There are many unanswered questions concerning the optimal timing of androgen ablation and radiation, the duration and type of androgen ablation, and the population of patients who will obtain the greatest benefit. PMID- 10851328 TI - Neoadjuvant Androgen Ablation Combined with External-Beam Radiation Therapy and Permanent Interstitial Brachytherapy Boost in Localized Prostate Cancer. AB - Androgen ablation therapy has been combined with permanent interstitial brachytherapy in order to downsize the gland prior to seed implantation. It also has been employed in an attempt to improve the effectiveness of therapy in patients with a poor prognosis. We report on 50 patients consecutively treated and prospectively followed. All received neoadjuvant hormonal therapy (NHT) and 45 Gy of external-beam therapy to a limited pelvic field, followed by permanent implantation of (125)I or (103)Pd seeds. The median follow-up is 42.1 months (range 9.0-90.8 months). The prostate specific antigen (PSA) progression-free survival rate (<1.0 ng/mL) was 76% at 5 years (Kaplan-Meier method). Local control was achieved in 100% of the patients and distant disease-free survival in 85%. High-risk patients treated contemporaneously with these patients, who received external-beam radiation and a seed boost without NHT, had a 62% rate of 5-year PSA progression-free survival. Although the modest improvement in PSA progression-free survival is not statistically significant at 5 years (P = 0.5), the patients treated with NHT in addition to combined radiotherapy presented with significantly higher serum PSA concentrations (mean 21.0 ng/mL; median 17.0 ng/mL) than those treated with combination radiotherapy alone (mean 15.6 ng/mL; median 10.6 ng/mL) and thus had a worse prognosis. PMID- 10851329 TI - Neoadjuvant Hormonal Therapy Improves the Outcomes of Patients Undergoing Radioactive Seed Implantation for Localized Prostate Cancer. AB - Neoadjuvant hormonal therapy (NHT) has been extensively studied in patients undergoing radical prostatectomy and external-beam irradiation for prostate cancer. While there are a few reports in the literature on its use in men undergoing brachytherapy, little information exists about its beneficial effects in such patients. In this report, we describe the effects of NHT on prostate volume (PV) prior to seed implantation and on the prostate specific antigen (PSA) and postimplant biopsy outcomes of patients who presented with high-risk features. Hormone therapy (leuprolide and flutamide 750 mg/day) was given to 145 patients for 3 months prior to and for 3 months after permanent iodine-125 (160 Gy) or palladium-103 (115 Gy) seed implantation. Of these, 28 (19%) received NHT because of a preimplant PV >50 cc, and 117 patients received NHT because they had a PSA >10 ng/mL, Gleason score >/=7, or clinical stage >/=T(2b). All patients underwent implantation using the real-time intraoperative method, and no patients received external-beam irradiation. Of the 145 patients treated, 67 (46%) had a PSA >10 ng/mL (range 1.9-57 ng/mL; mean 12.2 ng/mL), 50 (35%) had Gleason score >/=7, and 80 (55%) had stage >/=T(2b) disease. Prostate volume was measured in 106 patients prior to NHT and 3 months later immediately prior to the seed implant. The mean PV was 50.4 cc (range 17-150 cc), whereas the mean PV after NHT was 31 cc (range 11.7-73.7 cc). The mean PV reduction was 35% (range 2%-62%). Volume reduction was compared in those patients who presented with a PV <40 cc (N = 51) and those with a PV >/=40 cc (N = 56). The mean reduction for the smaller glands was 29% (range 2%-54%) compared with 41% (range 7%-62%) for the larger glands (P < 0.05). Patients were followed for a minimum of 1 year (range 1.0-6.4; mean 2.2 years). The 4-year actuarial rate of freedom from PSA failure (PSA >1.0 ng/mL with two consecutive elevations) was 85%. There was no difference in rates of freedom from PSA failure for those with initial Gleason 2-4 (96%), 5-6 (78%), 7 (80%), or 8-9 (83%; P = 0.5). Control rates were 85% for patients with PSA 20 ng/mL (P = 0.8). There was a trend to decreased control rates with higher stage disease (98% for T(1)-T(2a) v 68% for T(2c)), but these differences were likewise not significant (P = 0.12). The control rates for the 28 low-risk patients with enlarged prostate glands were compared with those of the 117 with high-risk features and were not different (100% v 82%; P = 0.1). There were 62 patients who agreed to eight-core prostate biopsies 2 years after implantation, and 60 (97%) were negative for tumor. This trial shows that NHT can reduce PV an average of 35% prior to seed implantation with the greatest reduction found in patients with larger prostates (41%). Hormonal therapy also appears to improve biochemical (PSA) control and local control (prostate biopsy) in patients with high-risk disease, yielding results similar to those in men with low-risk prostate cancer. PMID- 10851330 TI - Microwave Thermoablation for Localized Prostate Cancer After Failed Radiation Therapy: Role of Neoadjuvant Hormonal Therapy. AB - The treatment of residual prostate cancer after irradiation is often associated with significant morbidity and a high failure rate. Percutaneous transperineal interstitial microwave thermoablation is a minimally invasive procedure used experimentally in our institution to treat selected patients with failures of external-beam radiation therapy for prostate cancer. The aim is to ablate all residual intraprostatic cancer thermally. Patients were treated under general or epidural anesthesia with transrectal ultrasound guidance of transperineal placement of the microwave antennas. The rectum, urethra, and a developed space between the prostate and surrounding tissues were actively cooled. The minimal target temperature of the prostate was 65 degrees C for 15 min. The temperature was measured in all cases with interstitial prostatic thermosensors and in selected cases with online magnetic resonance scanning. Thirty-seven patients with apparently localized prostate cancer after failure of treatment for cure with external-beam therapy were subjected to this treatment, and 20 of these patients have at least 12 months of follow-up. The initial prostate specific antigen (PSA) concentration ranged from 0.2 to 120 ng/mL. At 12 months, 12 of 20 patients had no biochemical or histologic evidence of disease, and 11 of 14 patients with initial PSA concentration <10 ng/mL had no evidence of disease. Five of the thirty-seven patients were treated with 3 months of neoadjuvant androgen ablation because the volume of their prostates precluded adequate heating. The average volume decline was 28%, which allowed all men to be treated. Two of these patients have been followed for at lease 1 year, and neither shows evidence of recurrence. Side effects of treatment in all patients were modest. Preliminary results suggest that this treatment might be useful in selected patients as a salvage therapy after failure of radiation therapy for localized prostate cancer. PMID- 10851331 TI - Neoadjuvant Hormone Therapy Before Radical Prostatectomy: Update on the Memorial Sloan-Kettering Cancer Center Trials. AB - We report here the latest follow-up of the Phase II and III trials evaluating pathologic results and relapse-free survival, as judged by serum prostate specific antigen (PSA), in patients with localized prostate cancer who had radical prostatectomy performed at the Memorial Sloan-Kettering Center (MSKCC) either with or without neoadjuvant hormone therapy (NHT). Pelvic lymphadenectomy (PLND), radical prostatectomy, or both with or without NHT was performed in 141 patients enrolled in a Phase II trial comparing patients receiving NHT with concurrent controls and 140 patients in a randomized Phase III trial. In the Phase II study, there was a significant difference in the pathologic results, with only 35 (49%) of the 72 patients in the control group having organ-confined margin-negative disease compared with 48 (70%) of the 69 patients in the NHT arm (P = 0.0057; chi(2) test). With a median follow-up of 57 months, there was no significant difference in the PSA relapse rates in the two arms (P = 0.92; log rank test). In the Phase III study, 39 (59%) of the 66 patients in the control arm had organ-confined margin-negative disease compared with 52 (70%) of the 74 patients in the NHT arm (P = 0.17; chi(2) test). However, the positive-margin rate was significantly lower in the NHT arm (19%) than in the control arm (37%) (P = 0.023; chi(2) test). With a median follow-up of 35 months, there was no significant difference in the PSA relapse rates in the two arms (P = 0.73; log rank test). Thus, although NHT improves the pathologic results, further follow-up is necessary to determine if this marked reduction in the positive-margin rate will translate into improved disease-free survival. PMID- 10851332 TI - Neoadjuvant Hormonal Therapy in Stage C Adenocarcinoma of the Prostate and Incidence of Biochemical Recurrence. AB - This report describes the effects of neoadjuvant hormonal therapy (NHT) in 31 patients with clinical stage T3 adenocarcinoma of the prostate (CaP) who underwent radical prostatectomy (RP) and the incidence of biochemical recurrence, as evidenced by serum prostate specific antigen (PSA) >0.2 ng/mL after RP. Twenty six patients received combined androgen blockade consisting of a gonadotropin releasing hormone (GnRH) agonist (leuprolide) and an antiandrogen (flutamide), and the remaining five received flutamide alone. The mean Gleason score was 7.1 +/- 0.3. Five patients were found to have pathologic stage T(2c), 14 stage T(3a), 7 stage T(3b), and 4 stage T(3c) disease. One specimen had no evidence of adenocarcinoma in the whole-mount specimen. The mean PSA concentration decreased from 25.4 ng/mL (Hybritech method) to 1.18 ng/mL. The prostate volume decreased a mean of 47%, and pathologic effects of hormonal deprivation were observed in all patients. Approximately 39% of the patients (12/31) demonstrated evidence of biochemical recurrence, with the mean time to biochemical failure being 28.9 months after NHT. Follow-up ranged from 5 to 83 months with a mean of 52.7 months. Of the variables studied (pretreatment PSA, Gleason score, final pathologic stage, and nadir PSA before surgery), pretreatment PSA (p = 0.001, Fisher's exact test) and pathologic stage T(3c) (p = 0.012, Fisher's exact test) were found to have a statistically significant correlation with biochemical recurrence. Although there was a 39% biochemical recurrence rate in our study, we conclude that NHT offers an alternative mode of management for patients with stage T(3) CaP. PMID- 10851333 TI - Neoadjuvant Hormonal Therapy and Radical Prostatectomy for Locally Advanced Prostate Cancer. AB - The management of locally advanced prostate cancer remains controversial. Cause specific survival rates are high for organ-confined disease, but there is a disturbingly high incidence of positive margins in radical prostatectomy specimens. Thus, there has been much interest in neoadjuvant hormonal therapy as a means of shrinking the primary tumor. Both nonrandomized and randomized trials have revealed reductions in tumor size, but the effect on tumor stage and patient survival is less clear. Because the long-term value is not clear, and because neoadjuvant hormonal therapy does have side effects and increases treatment costs, it is at present not advisable outside the clinical research setting. PMID- 10851334 TI - Immunohistochemical Analysis of Radical Prostatectomy Specimens After 8 Months of Neoadjuvant Hormonal Therapy. AB - Neoadjuvant hormone therapy (NHT) prior to radical prostatectomy (RP) results in residual foci of atrophic glands, which can be difficult to identify with hematoxylin-eosin staining, raising the possibility that the low positive-margin rates are an artifact of pathologic understaging. The purpose of this study was to evaluate changes in prostate specific antigen (PSA) and prostatic acid phosphatase (PAP), as well as proliferation marker Ki-67 and Bcl-2 oncoprotein, by immunostaining after 8 months of NHT. Twenty-nine men with clinically confined prostate cancer who received 8 months of NHT and had both pretreatment biopsy and RP specimens obtained at Vancouver Hospital constituted the treatment group. The control group consisted of 23 RP specimens from patients not receiving NHT. Sections were stained with antibodies against PSA, PAP, proliferation marker Ki 67, and the antiapoptosis protein Bcl-2. The PSA and PAP immunostaining were graded for percentage of positive tumor cells and intensity of staining, while Ki 67 and Bcl-2 staining was graded according to the percentage of positive tumor cells. Absent or low percentage-positive PSA and PAP staining was observed in 40% to 50% of the NHT-treated RP specimens but none of the needle biopsy or untreated control RP specimens. Low-intensity PSA and PAP staining was detected only in RP specimens after NHT treatment, and then in only 20% of cases. Low or absent Ki-67 staining was noted in 78% of the NHT- treated RP specimens, compared with only 13% of the matched pre-NHT needle biopsies and 26% of untreated RP specimens. The percentage of specimens with high (>5%) Ki-67 staining decreased from 37% in the pre-NHT needle biopsies to 8% in the NHT-treated RP specimens. Bcl-2 staining increased after treatment with NHT, with 20% of the needle biopsies having high (>5%) Bcl-2 staining compared with 53% of the NHT-treated RP specimens. The frequency of low Bcl-2 staining (<1%) decreased from 53% in the pre-NHT needle biopsies to 27% in the NHT-treated RP specimens. Although PAP and PSA staining decreased after NHT, both markers remain sufficiently positive to be used as epithelial markers to help detect residual foci of prostate cancer that are difficult to identify on H&E-stained slides after NHT. Increased Bcl-2 after NHT, even in early-stage disease, is consistent with its role in the prevention of apoptosis and progression to androgen independence. Low levels of Ki-67 staining indicates a low probability of proliferation and outgrowth of androgen independent clones after 8 months of NHT. PMID- 10851335 TI - Clinical Experience with Intermittent Androgen Suppression in Prostate Cancer: Minimum of 3 Years' Follow-Up. AB - This report reviews the long-term follow-up of a prospective Phase II evaluation of intermittent androgen suppression in the treatment of prostate cancer. A total of 87 patients have been entered in this protocol. At the time of this report, 50 men have been followed for a minimum of 3 years. Treatment was initiated with combined androgen blockade and continued for at least 6 months until a serum PSA nadir was observed. Medication was then withheld until the serum PSA increased to mean values between 10 and 20 ng/mL. This cycle of treatment and no treatment was repeated until the regulation of PSA became androgen independent. The total time in the study ranges from 40 to 126 months, with a mean of 65.5 months. The off treatment period in the first cycle for the men with long-term follow-up was associated with an improvement in the sense of well-being and recovery of libido and potency in men who reported normal or near-normal sexual function before the start of therapy. The average time off therapy (percentage time off therapy) for cycles 1, 2, 3, and 4 was 15 months (54%), 10 months (48%), 8 months (45%), and 7 months (40%), respectively. The study group included 9 patients treated because of a rising PSA concentration after radiation therapy for locally advanced cancer. These patients have been off therapy for an average of 22 and 13 months in treatment cycles 1 and 2, respectively. Six patients with rising PSA values after radical prostatectomy and with follow-up exceeding 36 months have been off therapy for an average of 19 and 11 months in treatment cycles 1 and 2, respectively. Of the 87 patients, 23 have had their disease progress to androgen independence at a median of 32 months of treatment, and 13 have died cancer specific deaths at a median of 48 months. Prostate cancer is amenable to control by intermittent androgen suppression. This approach affords an improved quality of life when the patient is off therapy, with reduced toxicity and costs. Phase II trials suggest that there is not a negative impact on patient outcome. Randomized protocols are currently in progress to determine whether survival is affected in a beneficial or adverse way by such treatment in men with locally recurrent or metastatic cancer. PMID- 10851336 TI - DNA Microarrays for Analysis of Gene Expression. AB - Transformation of the prostatic epithelium is accompanied by changes in the expression of many genes, as reflected in altered steady-state levels of the mRNAs derived from them. The acquisition of androgen independence is likewise accompanied by changes in gene expression. Using DNA microarray technology, it is now possible to measure the levels of thousands of different mRNAs in a single hybridization step. This technique is particularly powerful for comparing gene expression in the same tissue under different environmental conditions; for example, comparing mRNA levels in malignant cells growing in the presence or absence of a hormone such as testosterone. Sufficient cDNA for hybridization to a microarray can be produced from as little as 1 mg of tissue. Coupled with DNA sequence information emerging from the Human Genome Project, this technique has the potential to rapidly identify genes that are differentially expressed in situations with direct relevance to prostate cancer. It is reasonably likely that diagnostic patterns of gene expression can be found for normal v benign proliferative malignant prostatic epithelium, androgen-dependent v androgen independent tumors, and drug-sensitive v drug-resistant tumors. Once identified, informative cDNAs can be included in diagnostic microarrays with potential for clinical application. PMID- 10851337 TI - Progress in Two-Dimensional and Three-Dimensional Ultrasonic Tissue-Type Imaging of the Prostate Based on Spectrum Analysis and Nonlinear Classifiers. AB - Spectrum analysis of radiofrequency (RF) ultrasonic echo signals often can sense tissue differences that are not visible on conventional ultrasonic images. Spectrum-analysis parameter values combined with other variables, such as serum prostate specific antigen (PSA) concentration, can be classified by neural networks to distinguish effectively between cancerous and noncancerous prostate tissues. Images based on neural network classification of spectral parameters and clinical variables can be advantageous for biopsy guidance, staging, and treatment planning and monitoring. A study based on 644 biopsies from 137 patients showed that these methods are significantly superior to B-mode image interpretation for differentiating cancerous from noncancerous prostate tissues. Using the histologic determination of tissue types as the gold standard, the area under the receiver-operator characteristic (ROC) curve for neural network classification based on spectrum analysis and PSA value for the 644 biopsies was 0.87 +/- 0.04, and the ROC curve are for a level-of-suspicion (LOS) assignment based on B-mode imaging was 0.64 +/- 0.04. Color-encoded and gray-scale images derived from neural network assignment of suspicion for cancer at each pixel location showed remarkable detail and suggested potential clinical value for biopsy guidance using real-time two-dimensional (2D) images and staging, treatment planning, and monitoring using three-dimensional (3D) images. PMID- 10851338 TI - Prostate-Specific Membrane Antigen: Much More Than a Prostate Cancer Marker. AB - Prostate cancer continues to be the most common cancer and second leading cause of cancer-related death among men. The use of markers, particularly serum-based prostate specific antigen (PSA), has contributed to the rapid rise in diagnosed cases in the late 1980s and early 1990s, but new diagnostic and possible therapeutic markers are needed and are currently being evaluated. One of these, prostate-specific membrane antigen (PSMA), is an approximately 100-kDa type II transmembrane protein originally thought to be highly selectively expressed in all types of prostatic tissue, with expression being upregulated in androgen depleted or androgen-independent states. The radioimmunoconjugate form of the anti-PSMA monoclonal antibody (mAb) 7E11 is currently being used to diagnose prostate cancer metastasis and recurrence. In addition, Phase I and II trials have started utilizing PSMA in different therapeutic ways, with promising results. Recent exciting work has demonstrated PSMA expression in endothelial cells of vessels restricted to the tumor-associated neovasculature. This finding expands the possible beneficial uses of PSMA, as new anti-PSMA mAbs continue to be developed. PMID- 10851339 TI - Rehabilitation of the Impotent Patient: An Update. AB - Erectile dysfunction (ED) affects as many as 90% of patients after treatment for prostate cancer. Denervation appears to be the primary mechanism after either surgery or radiation; venooclusion or smooth-muscle dysfunction may follow. There may also be a vasculogenic component. Changes in surgical technique and pharmacologic prophylaxis may reduce the likelihood of ED. In men who desire reversal of ED, a stepwise approach beginning with oral sildenafil and, if that drug is ineffective, moving to intraurethral or intracavernous therapy and, finally, to a surgical approach is advisable. PMID- 10851340 TI - Regulatory Issues in Prostate Cancer Studies. AB - Quality of life and informed consent are two of the important issues for designers of clinical trials. The former has physical, psychological, economic, and social components. Investigators must understand the features of the various measurement instruments and select one that is appropriate to the circumstances. It also is necessary to identify what changes are significant. Statistical significance is not identical to clinical significance. Ethical guidelines for the protection of human subjects is another important issue. Information on regulations is available on many Web sites, including that of the Food and Drug Administration. PMID- 10851341 TI - Survey of UsToo Members and Other Prostate Cancer Patients to Evaluate the Efficacy and Safety of PC-SPES. AB - Prostate cancer survivors frequently seek natural remedies for elevated or rising PSA concentrations to forestall the necessity for more definitive modalities. PC SPES is one of the most widely used of the complementary medicines. It consists of eight Chinese herbs, and although it contains no estrogen, it does exhibit some estrogenic effects. For this reason, UsToo was anxious to determine just how successful the product is and whether any side effects are present. A four-page survey form was designed and pretested on a dozen patients. After refinement, the form was sent to 200 PC-SPES users, mostly UsToo members, with anonymity assured. In only five cases did respondents not identify themselves. After 102 responses had been received, a compilation form was designed to simplify computer database entry of the survey results. This produced a spreadsheet of all 102 respondents' categorized answers. Final analysis followed, with emphasis on prostate specific antigen (PSA) concentrations before and after PC-SPES use, quality of life (QoL), side effects, dosage, and relation to concurrent treatment modalities. Graphs were then constructed on each respondent for study of slope data and PSA changes. Twenty respondents provided insufficient data for analysis; the remaining 82 surveys gave us a good picture of PC-SPES usage and results, as well as information on commingling of other modalities with PC-SPES. Beneficial effects were reported by 77% of the respondents, with 23% reporting more limited or marginal results. Side effects reported were breast tenderness and lowered libido, with three respondents also reporting leg edema. There were no reported cases of circulatory problems or thrombosis. Declines in PSA were reported of as much as 70 ng/mL that were sustained for as long as the respondents have been using PC-SPES, approaching 2 years in some cases. No clinically significant adverse effects were observed. For some men, PC-SPES provides an alternative to hormonal therapy; has a palliative effect when used by patients with advanced, metastatic disease; and overall has a reported 77% effectiveness, with 87% effectiveness when recommended dosages are adhered to. PMID- 10851342 TI - Structure of the intramural nerves of the rat bladder. AB - The bladder of adult female rats receives approximately 16,000 axons (i.e., is the target of that many ganglion neurons) of which at least half are sensory. In nerves containing between 40 and 1200 axons cross-sectional area is proportional to number of axons; >99% of axons are unmyelinated. A capsule forms a seal around nerves and ends abruptly where nerves, after branching, contain approximately 10 axons. A single blood vessel is present in many of the large nerves but never in nerves of <600 axons. The number of glial cells was estimated through the number of their nuclei. There is a glial nucleus profile every 76 axonal profiles. Each glial cell is associated with many axons and collectively covers approximately 1, 000 microm of axonal length. In all nerves a few axonal profiles contain large clusters of vesicles independent of microtubules. The axons do not branch; they alter their relative position along the nerve; they vary in size along their length; none has a circular profile. All the axons are fully wrapped by glial cells and never contact each other. The volume of axons is larger than that of glial cells (55%-45%), while the surface of glial cell is twice as extensive as that of axons; there are approximately 2.27 m(2) of axolemma and approximately 4.60 m(2) of glial cell membrane per gram of nerve. Of the mitochondria of a nerve approximately 3/4 are in axons and approximately 1/4 in glial cells. PMID- 10851343 TI - An investigation of astroglial morphology in torpedo and scyliorhinus. AB - The distribution and morphology of GFAP-immunoreactive cells was investigated in two elasmobranch species, Scyliorhinus canicula and Torpedo marmorata, in an attempt to distinguish between Horstmann's (1954) hypothesis that the presence of cells resembling mammalian astrocytes is a function of the thickness of the ventricular walls, and Cajal's (1911) hypothesis that astrocytes are a phylogenetic novelty found only in birds and mammals. Two types of GFAP-reactive elements were observed, but the distribution of these differed markedly between the two species. In Scyliorhinus, radial glial cells were predominant and astrocytes relatively rare. In Torpedo, on the other hand, a species in which the ventricles are atrophied and the ventricular walls extremely thick, the overwhelming majority of GFAP-labelled structures strongly resembled astrocytes; occasionally, GFAP-positive cells were observed in the ependyma of the spinal cord. These findings, together with previous results obtained by others in hagfish, provide strong evidence in favour of Horstmann's hypothesis. PMID- 10851344 TI - Regional and cellular distribution of neural visinin-like protein immunoreactivities (VILIP-1 and VILIP-3) in human brain. AB - Neural visinin-like proteins (VILIPs) are members of the neuronal subfamily of intracellular EF-hand calcium sensor proteins termed the NCS family, which are thought to play important roles in cellular signal transduction. While numerous studies suggest a wide but uneven distribution of these proteins in rat and chicken brain, their location in, and possible significance for, the human brain, remains to be established. We used specific polyclonal antisera to map the human brain for VILIP-1 and VILIP-3 immunoreactivities. VILIP-1 was detected in cortical pyramidal cells and interneurons, septal, subthalamic and hippocampal neurons (subfields CA1 and CA4 pyramidal cells and especially hilar interneurons) as well as in cerebellar Golgi, basket, granule, stellate and dentate nucleus neurons. Purkinje cells were free of immunoreaction. VILIP-3 was more restricted in its distribution. It was identified in cerebellar Purkinje cells and a subpopulation of granule neurons. Further, neurons belonging to different nuclei of the brain stem and multiple subcortical nerve cells stained for visinin-like protein 3. A weak immunoreaction appeared in cortical and hippocampal neurons. Intracellularly the immunoreactivity appeared in the perikarya, dendrites and some axons. Sometimes, immunostaining was found in the neuropil. Glia did not express visinin-like proteins. Our findings support, from a neuroanatomical viewpoint, the idea that these calcium sensor proteins may be of relevance for neuronal signalling in the human CNS. PMID- 10851345 TI - Tooth pulp tissue promotes neurite outgrowth from rat trigeminal ganglia in vitro. AB - The mammalian tooth pulp becomes innervated by nociceptive and sympathetic axons relatively late during development, when part of the root has formed. In the adult, regenerating axons from an injured tooth nerve or sprouting axons from uninjured nerves in the vicinity rapidly reinnervate denervated tooth pulps. These observations indicate that tooth pulp tissue can use molecular factors to attract pulpal axons from local nerve trunks. The present study examines the hypothesis that these factors include nerve growth factor (NGF), brain derived neurotrophic factor (BDNF) and glial cell line derived neurotrophic factor (GDNF). Explants of trigeminal ganglia from neonatal rat pups showed a distinct neurite outgrowth when co-cultured with pulpal explants collected from molar teeth of 12-day old pups, or after application of a pulpal extract. Control cultures, containing single ganglionic explants, or explants co-cultured with heat-treated pulpal tissue, exhibited a sparse neurite outgrowth. Exogenous NGF and/or GDNF, but not exogenous BDNF, stimulated neurite outgrowth from ganglionic explants. Unexpectedly, application of antibodies against NGF, BDNF and/or GDNF to co-cultures of ganglionic and pulpal explants did not inhibit neuritogenesis. Control experiments showed that IgG molecules readily penetrate the gel used for culture and that even very high concentrations of NGF and GDNF antibodies in combination failed to block neurite growth. On the basis of these data we suggest that other as yet unknown neurite-promoting factors might be present and active in TG/pulpal co-cultures. PMID- 10851346 TI - Microtubule alterations in cultured taiep rat oligodendrocytes lead to deficits in myelin membrane formation. AB - The taiep rat is a myelin mutant in which hypomyelination and progressive demyelination of the CNS are accompanied by an accumulation of microtubules within oligodendrocytes. To investigate whether and how the myelin defects were caused by microtubule abnormalities, we have established a taiep oligodendrocyte culture system in which mutant cells produce abnormally high levels of tubulin and microtubule-associated proteins and exhibit myelin defects. The studies show that abnormal microtubule accumulation and tight microtubule bundles developed in the taiep oligodendrocytes, with a higher ratio of minus-end-distal to plus-end distal microtubules in their processes. Initially, in culture, immature taiep oligodendrocytes which have higher levels of tubulin than controls extend roughly twice as much membrane sheet as controls. The membrane sheets of the mature taiep oligodendrocytes which display the microtubule accumulation, however, grew much less rapidly compared to controls. By the fifth day in culture, a majority of the taiep oligodendrocytes had ceased the expansion of their membrane sheets and in some cases the sheets retracted. The levels of the myelin proteins, proteolipid protein and myelin-associated glycoprotein, were also markedly diminished in the mature taiep oligodendrocytes. Treatment with the microtubule depolymerizing drug nocodazole prevented not only the accumulation of microtubules but also restored the normal distribution of proteolipid proteins within the taiep oligodendrocytes. These data demonstrate that myelin synthesis in the oligodendrocyte cultures relies on the formation of a normal microtubule array, and the microtubule abnormalities are directly responsible for the myelin deficit in the taiep oligodendrocytes. PMID- 10851347 TI - Differential response of macrophage subpopulations to myelin degradation in the injured rat sciatic nerve. AB - Molecular mechanisms of myelin removal by macrophages were explored by examining the immunophenotypes of macrophages following injury of rat sciatic nerve, using a combined method of immunohistochemistry and confocal laser microscopy. In the crush injury model, the involvement in myelin clearance of a cytoplasmic antigen specific for monocytes/macrophages, ED1, was evident. The obvious recruitment of ED1-immunoreactive (-ir) cells was detected first at the crush injury site and then in the distal stump within which Wallerian degeneration had occurred. Double labelling revealed that the ED1-ir cells, except for monocyte-like round cells, always phagocytosed myelin basic protein-ir myelin debris. On the other hand, the expression of ED2, a surface antigen specific for resident macrophages, was significantly different; ED2-ir cells also increased while myelin removal was progressing from day 3 to day 7, but only some of the cells were engaged in myelin phagocytosis. The poor capacity of myelin phagocytosis by ED2-ir cells was supported by the transection model, in which the proximal stump was ligated to suppress regeneration. ED2 may be involved in events other than myelin removal, providing a local environment conducive to axonal regeneration. Our findings thus seem to suggest that ED1 is one of the most reliable markers for cells carrying out myelin phagocytosis, whereas ED2 may participate in entirely different functions. The expression of complement receptor type 3, OX42, was similar to that of ED1 in terms of the swift recruitment of immunopositive cells, their distribution with close association to myelin debris and their high phagocytotic capacity. This supports previously reported in vitro evidence that myelin phagocytosis by macrophages may be complement-mediated. PMID- 10851348 TI - [Pre-and postoperative arthroscopic and MRI findings in arthroscopic anterior cruciate ligament reconstruction of the knee]. PMID- 10851349 TI - Perineal musculature and its innervation by spinal motoneurons in the male rabbit: effects of testosterone. AB - Striated muscles in the perineum, which include the ischiocavernosus (IC), bulbocavernosus (BC), and levator ani (LA), were identified in an attempt to understand motor regulation of penile erection in the male rabbit. The IC surrounded the corpus spongiosum of the penis whereas the BC attached to the dorsum of the penis at the midline. The LA encircled the rectum and attached to the base of the penis. This anatomy suggested that the IC plays the primary role in the erection in the rabbit, whereas the BC may cause flips of the erected penis. Spinal motoneurons that innervate the IC were identified by retrograde labeling by horseradish peroxidase (HRP). As in other mammals, spinal labels from the IC appeared ipsilaterally in the ventral horn that encompassed the sixth lumbar (L6) and the first sacral segments. HRP injections into the BC labeled a small number of cells bilaterally at the same spinal levels. Rabbits are peculiar in having the IC motoneurons scattered among other motoneurons, unlike the rat and other rodents that have their IC motoneurons aggregated to form a spinal nucleus. Castration caused significant decreases in both the wet weight of IC muscles and the size of IC motoneurons within 2 weeks. Testosterone supplement following castration maintained the IC muscle weight and the neuronal size. Neither castration nor testosterone supplement induced changes in the number of IC motoneurons. PMID- 10851350 TI - Knee functions and a return to sports activity in competitive athletes following anterior cruciate ligament reconstruction. AB - We investigated knee functions and a return to sports in 50 competitive athlete patients treated with arthroscopic anterior cruciate ligament reconstruction using double-looped STG augmented by woven polyester at a 1-year follow-up. There were 25 males and 25 females with a mean age of 24.3 years (range: 19-39 years). The majority of preinjury sports were basketball, volleyball and soccer. Athletic rehabilitation including agility training and sports-specific training was started at 12 weeks. Fourty patients (80%) was rated as normal or nearly normal on the assessment of International Knee Documentation Commitee postoperatively. Fourty-eight patients (96%) obtained full range of motion, and the mean quadriceps muscle strength of the injured side was 91.3%of that of the uninjured side. As for a return to sports, 46 patients (92%) were able to do fully competitive sports at a mean of 8.1 postoperative months. These results suggest that arthroscopic reconstruction using augmented double-looped STG allows early athletic rehabilitation, and lead satisfactory outcome as well as a reliable and early return to preinjury level of sport activity for the majority of the competitive athlete patients. PMID- 10851351 TI - Overexpression and localization of heat shock proteins mRNA in pancreatic carcinoma. AB - In the present study we examined the localization and overexpression of heat shock proteins (hsps), mainly hsp90, in pancreatic carcinoma tissue compared with control tissue (including chronic pancreatitis and normal pancreas tissue), with the aid of immunohistochemical staining, in situ hybridization and reverse transcriptase polymerase chain reaction. Hsp90 alpha mRNA was overexpressed more highly in pancreatic carcinoma than in the control tissue. The proliferating-cell nuclear-antigen labeling index was also high in pancreatic carcinoma tissue compared with the other tissue. These findings suggest that the overexpression of hsp90 alpha mRNA in carcinomas may be correlated with cell proliferation. However, hsp90 beta was constitutively overexpressed almost equally in all groups of pancreatic tissue including pancreatic carcinoma, chronic pancreatitis and normal pancreas tissue. Immunohistochemical staining demonstrated a differentiation in the expression of hsp90 between histological types of pancreatic carcinoma. These findings suggest that hsp90 alpha is involved in carcinogenesis and that hsp90 beta is correlated to structural conformation. Hsp90 alpha and hsp90 beta seem to perform different functions in tissue containing malignant cells. P53, MDM2 and WAF1, that were cell-cycle-related oncogene product were more strongly expressed in the nuclei of the cancer cells of the cancer tissue. Especially, MDM2 was more strongly expressed in mucinous carcinoma and the mucin secreting tissues surrounding pancreatic carcinoma tissue. The expression of MDM2 protein might also be correlated to secretion systems during structural conformation and be correlated to hsp90 beta. PMID- 10851352 TI - An epidemiologic study of occupational low back pain in truck drivers. AB - The factors in volved in occupational low back pain occurring in professional drivers were investigated epidemiologically with questionnaires (92 items) including low back symptoms, personal factors and occupational factors. The responses of one hundred fifty-three of one hundred eighty-one truck drivers who work in a large chemical industry corporation were analyzed after they had completely filled in questionnaires. As analysis of the results shows, the prevalence of LBP in one month of the survey was 50.3%. Correlating among data of personal factors and LBP, the prevalence of LBP was significantly higher in the drivers (Odd's ratio of 2.7) who answered "yes" to the item "shortage of spending time with family than in the drivers who didnt answer "yes. The occupational factors, working load and working environment showed no correlation with the prevalence of LBP. In contrast, 3 items of the working format related significantly to the prevalence of LBP: "irregular duty time (Odd's ratio of 3.0), "short resting time (2.4), and "long driving time in a day (2.0). Eighty one of the 153 drivers (52.9%) pointed out the relationship between LBP and work, especially work which muolves vibration or road shock. Our results and the results from previous published studies suggested that vibration is an obvious risk factor for LBP. From the viewpoint of prophylaxis, an improvement in working conditions reduces the incidence of drivers' LBP to some extent. PMID- 10851353 TI - A case report of synovitis, acne, pustulosis, hyperostosis and osteitis syndrome presenting with spondylodiscitis. AB - SAPHO syndrome stands for synovitis, acne, pustulosis, hyperostosis and osteitis. The common site of skeletal lesions in this syndrome is the sternocostoclavicular area. Spondylodiscitis is rarely described in published studies. In general, skin lesions develop before the onset of skeletal lesions. We report a case of SAPHO syndrome in which spondylodiscitis developed more than 1 year before the onset of pustulosis. PMID- 10851354 TI - A case of congenital pseudarthrosis of the tibia treated with pulsing electromagnetic fields. 17-year follow-up. AB - Congenital pseudarthrosis of the tibia presents surgeons with one of the most challenging of all orthopedic problems. Various surgical treatments have succeeded only rarely. We report long-term follow-up of a patient with congenital pseudarthrosis of the tibia treated with pulsed electromagnetic fields (PEMF) and bone grafting. In this severe case, Bassett type III and Boyd type II, encouraging results were achieved with Boyd's dual onlay grafts and PEMF. Seven years after surgery, skeletal maturity was complete and an unacceptable degree of leg shortening had been avoided. PMID- 10851355 TI - [Risk factors of cardiovascular disease and those managements, especially for acute coronary syndrome]. PMID- 10851356 TI - [Two cases of myocardial bridge associated with myocardial ischemia]. PMID- 10851357 TI - Injecting rationale into interferon-beta therapy. PMID- 10851358 TI - Estrogen to treat Alzheimer's disease: too little, too late? So what's a woman to do? PMID- 10851359 TI - Remedies for HIV-associated peripheral neuropathy. PMID- 10851360 TI - Practice parameter: the role of corticosteroids in the management of acute monosymptomatic optic neuritis. Report of the Quality Standards Subcommittee of the American Academy of Neurology. PMID- 10851361 TI - The molecular and genetic basis of AD: the end of the beginning: the 2000 Wartenberg lecture. PMID- 10851362 TI - A comparative study of the relative bioavailability of different interferon beta preparations. AB - BACKGROUND: Three different recombinant interferon beta (IFNbeta) preparations are currently available for the treatment of MS, two IFNbeta-1a products (Rebif and Avonex) and one IFNbeta-1b product (Betaferon). These products differ with respect to the recommended dose, the dosage regimen, and the injection route. This study compared the relative biologic activity of these three products in vitro and in vivo. METHODS: Blood samples were collected from 237 patients with MS (170 on IFNbeta therapy and 67 control subjects receiving no therapy). Samples with neutralizing antibodies were excluded. Levels of the antiviral protein MxA and soluble vascular cell adhesion molecule-1 (sVCAM) in the four groups were measured by ELISA. In the in vitro investigation, untreated blood was incubated for 24 hours with increasing concentrations of the three IFNs from a dose of 1 IU/mL to 1000 IU/mL, after which levels of MxA were measured. RESULTS: A difference between the groups was observed in vitro, with a significant change from baseline in MxA levels being observed at 10 IU for Betaferon compared with 100 IU for Rebif and Avonex. However, this might be due to the different methods used for the determination of IU by the manufacturers. At the single-dose level there were no significant differences between IFNbeta preparations. In vivo, there were significantly different levels of MxA in the four groups. In the Betaferon group, the median value for MxA was 2.29 ng/105 peripheral blood leukocytes (PBL), compared with 1.00 ng/105 PBL for Rebif, 0.57 ng/105 PBL for Avonex, and 0.14 ng/105 PBL for the control group. Some significant differences between the groups were also apparent with respect to levels of sVCAM, which were higher with Betaferon than with Rebif. CONCLUSION: IFNbeta-1b induces higher levels of the above markers of IFNbeta bioactivity than either of the two IFNbeta 1a preparations. Moreover, there is a less striking difference between the two IFNbeta-1a preparations in favor of subcutaneous IFNbeta-1a. PMID- 10851363 TI - Effects of estrogen on cognition, mood, and cerebral blood flow in AD: a controlled study. AB - OBJECTIVE: To examine the effects of estrogen therapy on cognition, mood, and cerebral blood flow in patients with AD. BACKGROUND: Some studies have suggested estrogen may be effective in the treatment of AD. However, most of these studies were not controlled adequately. METHODS: Fifty female AD patients were recruited in a randomized, double-blind, placebo-controlled 12-week trial. Each member of the estrogen-treated group received conjugated estrogen (Premarin) 1.25 mg/day. The primary outcome measures were the Cognitive Ability Screening Instrument (CASI), Clinical Dementia Rating (CDR), and Clinician Interview-Based Impression of Change (CIBIC-plus). The secondary outcome measures were Behavioral Pathology in Alzheimer's Disease (BEHAVE-AD), Hamilton Anxiety Rating Scale (HARS), Hamilton Depression Rating Scale (HDRS), and 99mTc hexamethylpropylene amine oxime SPECT of the brain. RESULTS: No meaningful differences were found between the outcome measures (CASI, CDR, CIBIC-plus, BEHAVE-AD, HARS, HDRS, and cerebral blood flow) taken from the estrogen-treated group and those from the control group. CONCLUSION: A 1.25-mg/day dose of Premarin administered for 12 consecutive weeks does not produce a meaningful effect on cognitive performance, dementia severity, behavior, mood, and cerebral perfusion in female AD patients. Because estrogen therapy has been suspected of yielding adverse effects, and its therapeutic effectiveness is in doubt, additional evaluation of its role in AD treatment ought to be conducted. PMID- 10851364 TI - Reduced prevalence of AD in users of NSAIDs and H2 receptor antagonists: the Cache County study. AB - OBJECTIVE: To test the hypothesis that nonsteroidal anti-inflammatory drugs (NSAIDs) and histamine H2 receptor antagonists (H2RAs) are associated with a decreased risk of AD in late life. BACKGROUND: Sustained use of non-aspirin NSAIDs has been repeatedly associated with a reduced occurrence of AD. Similar effects with aspirin have been weaker. One prior study showed a strong association between use of H2RAs and reduced AD prevalence. METHODS: In a population study of AD in Cache County, UT, we used a sequenced plan of sampling and case ascertainment to identify 201 cases of AD and 4425 participants with no indication of cognitive impairment. Independently, an interview and medicine chest inventory assessed use of several medicines including aspirin, non-aspirin NSAIDs, H2RAs, and three classes of "control" drugs not thought to be associated with AD. Follow-up questioning probed possible indications for use of these drugs. RESULTS: Compared with cognitively intact individuals, the AD cases had significantly less reported current use of NSAIDs, aspirin, and H2RAs. Stronger associations appeared when subjects reported use of both NSAIDs and aspirin (no H2RAs), two different NSAIDs (no H2RAs), or two different H2RAs (with neither aspirin nor NSAIDs). There was little or no such association with use of the control medicines. Adjustment for usage indication did not influence these findings, and there was no appreciable variation with number of APOE epsilon4 alleles. CONCLUSIONS: As predicted, use of NSAIDs and aspirin were specifically associated with reduced occurrence of AD. Notably, a previous observation of an inverse association of AD and use of H2RAs was also affirmed. Definitive evidence for a preventive action of these agents will require randomized prevention trials. PMID- 10851365 TI - Age-specific incidence rates of Alzheimer's disease: the Baltimore Longitudinal Study of Aging. AB - OBJECTIVE: To estimate age-specific incidence rates of AD in the Baltimore Longitudinal Study of Aging (BLSA). BACKGROUND: The BLSA is a volunteer cohort of normal subjects followed longitudinally with biennial evaluations at the Gerontology Research Center of the National Institute on Aging. METHODS: Subjects are 1236 participants (802 men, 434 women) in the BLSA with longitudinal follow up between January 1985 and May 1998. The average length of follow-up was 7.5 years, with participants evaluated every 2 years by physical, neurologic, and neuropsychological examinations. Using Diagnostic and Statistical Manual of Mental Disorders, 3rd ed., revised and National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria, the authors diagnosed dementia and AD. RESULTS: The authors diagnosed 155 cases of dementia, of which 114 (74%) were AD. Incidence rates of AD increased with age from an estimated 0.08% per year (95% CI 0.00 to 0.43) in the 60 to 65 age group to an estimated 6.48% per year (95% CI 5.01 to 8.38) in the 85+ age group for men and women combined. The doubling time of incidence rates was estimated to be approximately 4.4 years and the median time of conversion from mild cognitive impairment to diagnosis of AD was estimated to be 4.4 years. There was a trend for women to have higher incidence rates than men and for fewer years of education to be associated with higher incidence rates; however, these effects were not significant. CONCLUSION: Incidence rates for AD in the BLSA are consistent with published rates in other studies. The longitudinally followed subjects of the BLSA offer a unique opportunity to prospectively investigate the antecedents of AD. PMID- 10851366 TI - TNF gene polymorphism and its relation to intracerebral production of TNFalpha and TNFbeta in AD. AB - OBJECTIVE: To analyze the extent of tumor necrosis factor-alpha (TNFalpha) and TNFbeta gene polymorphism in patients with AD and to relate it to intrathecal levels of these cytokines. METHODS: Analyses of TNFalpha and TNFbeta gene polymorphism were performed using PCR in 52 patients with AD and in 25 control subjects, and the levels of corresponding cytokines were analyzed using ELISA. RESULTS: Patients with AD displayed significantly higher intrathecal levels of TNFalpha, but not TNFbeta, compared with the control subjects. The levels of these cytokines did not differ significantly in patients displaying different alleles of the TNF gene. CONCLUSIONS: Results indicate that increased intrathecal production of TNFalpha in AD is preferentially controlled by environmental stimuli rather than genetic makeup. PMID- 10851367 TI - Is APOE--epsilon4 a risk factor for cognitive impairment in normal aging? AB - OBJECTIVES: To examine the relationship between APOE genotype and cognitive functioning in normal aging, and to determine whether this relationship was moderated by age or the presence of a number of disease conditions, including cardiovascular disease and diabetes. METHODS: The sample was drawn from the Charlotte County Healthy Aging Study, a community-based, cross-sectional study of randomly selected older adults in Charlotte County, FL. A total of 413 older adults (mean age = 72.90 years) were examined in the current study. Participants completed tasks that indexed a variety of dimensions of cognitive functioning, including episodic memory, implicit memory, psychomotor speed, and attention. In addition, participants provided self-reported and objective indices of health status and were genotyped for APOE. RESULTS: Mean-level results indicated that groups with and without the APOE-epsilon4 allele performed similarly on all domains of cognitive functioning. Significant age group differences were observed in episodic memory, psychomotor speed, and attention but not implicit memory. Significant gender differences were present for episodic memory and the Stroop test. Analyses also indicated that participants' age did not exert an impact on the relationship between APOE-epsilon4 and cognitive functioning. Further, the presence of cardiovascular disease or diabetes did little to moderate the relationship between APOE-epsilon4 and cognition. CONCLUSIONS: The authors found no evidence for a relationship between presence of the APOE-epsilon4 allele and cognitive functioning. Further, age or the presence of a number of chronic conditions did not significantly moderate the effect of APOE genotype on cognitive performance. These results indicate that the presence of the epsilon4 allele is not a risk factor for cognitive impairment in normal aging. PMID- 10851368 TI - Do transient ischemic attacks have a neuroprotective effect? AB - OBJECTIVE: To determine whether TIAs have a neuroprotective effect. BACKGROUND: Ischemic tolerance or preconditioning, which protects the brain against stroke, has been demonstrated in animal models of cerebral ischemia. Because TIA may represent a clinical model of ischemic tolerance, patients with TIA before cerebral infarction (CI) may therefore have a better outcome than patients without TIA before CI. METHODS: A total of 2,490 patients admitted consecutively to a primary care center for first-ever CI in the anterior circulation were divided into two groups on the basis of the presence or absence of prior ipsilateral TIAs. Duration of TIA was classified into three groups (<10 minutes, 10 to 20 minutes, and >20 minutes). The severity of the neurologic picture on admission and functional disability after stroke were compared between patients with and without TIAs. RESULTS: A total of 293 (12%) of the 2,490 patients had prior ipsilateral TIAs before CI. Risk factors did not differ between patients with or without TIAs, whereas the topography and etiology of ischemic stroke did differ (p < 0.001). Patients without prior TIAs had a more severe clinical picture on admission, with a greater reduction of consciousness (p = 0.009). Patients with previous TIAs had a more favorable outcome than those without TIAs (67% versus 58%, p = 0.004). After adjustment for confounding variables, TIAs lasting 10 to 20 minutes were still associated with a favorable outcome (odds ratio, 1.98; 95% confidence interval, 1.27 to 3.08; p = 0.002). The interval between TIA and CI influenced the outcome (p = 0.007). CONCLUSIONS: This study suggests that ischemic tolerance may play a role in patients with ipsilateral TIAs before CI, allowing better recovery from a subsequent ischemic stroke. PMID- 10851369 TI - Long-term changes of hemodynamics and metabolism after carotid artery occlusion. AB - OBJECTIVE: To investigate whether in selected patients with internal carotid artery (ICA) occlusion and initially normal oxygen extraction fraction (OEF) measured with PET, subsequent changes of cerebral hemodynamics and metabolism occur during long-term follow-up and, if so, whether the changes are associated with atrophy of the corpus callosum or subsequent ischemic strokes. BACKGROUND: The course of the changes in cerebral hemodynamics and metabolism after ICA occlusion remain unclear. After ICA occlusion, an increase in OEF may increase the risk of cerebral ischemia, and an increase in cortical ischemia would cause progression of callosal atrophy. METHODS: The authors used PET and MRI to examine twice seven medically treated patients with unilateral ICA occlusion and initially normal OEF at intervals ranging from 24 to 64 (mean +/- SD, 42 +/- 17) months. No intervening ischemic attacks occurred between the two examinations. RESULTS: In the hemisphere with ICA occlusion, OEF increased and blood flow decreased during follow-up. At the follow-up evaluation, abnormally increased OEF values were found in three patients, in whom ipsilateral ischemic strokes occurred during subsequent follow-up (18 +/- 6 months). A decrease in oxygen metabolism also occurred and was significantly correlated with the decrease of callosal size. CONCLUSIONS: These preliminary findings in a small, selected patient sample suggest that in patients with ICA occlusion and initially normal OEF, deteriorations of cerebral hemodynamics and metabolism during long-term follow-up may be associated with callosal atrophy or subsequent ischemic strokes. PMID- 10851371 TI - Progressive micrographia in a migraineur PMID- 10851370 TI - Incidence of stroke in relation to cognitive function and dementia in the Kungsholmen Project. AB - OBJECTIVE: To investigate whether cognitive function is related to incidence of stroke. METHODS: A population-based cohort of 1551 subjects with no clinical history or signs of stroke, age 75 years and over at baseline, were followed up for 3 years. Individuals with a first-ever stroke event that was recorded in the Stockholm inpatient register after the date of baseline interview were considered as incident stroke patients. Diagnosis of stroke followed the International Classification of Disease, 9th Revision (ICD-9). Diagnosis of dementia was made according to the Diagnostic and Statistical Manual of Mental Disorders, 3rd ed., revised (DSM-III-R). Cognitive functioning was assessed with the Mini-Mental State Examination. RESULTS: During the 4102 person-years of follow-up, 110 events were recorded, giving an overall incidence of stroke of 26.8 per 1000 person years. Subjects with mild dementia had a relative risk of 2.6 (95% CI, 1.2 to 5.7) of developing stroke after controlling for the potential confounders. The corresponding figure for subjects with cognitive impairment was 2.0 (95% CI, 1.0 to 3.8; p = 0.05). There was a tendency for subjects who developed stroke to be more likely to have vascular factors (systolic blood pressure >180 mm Hg, heart disease, or diabetes mellitus) than those who did not. CONCLUSIONS: Mild dementia and cognitive impairment are associated with an increased incidence of stroke among subjects age 75 years old and over. Because stroke increases risk of dementia and prior stroke increases risk of a subsequent stroke, mild dementia and cognitive impairment may be a manifestation of clinically unrecognized stroke. PMID- 10851372 TI - Biobehavioral characteristics of nondemented older adults with subclinical brain atrophy. AB - OBJECTIVE: To characterize the risk factors and neuropsychological performance of two subgroups of community-dwelling, white elderly men free of severe cognitive impairment (n = 383; mean age, 72.9 +/- 3.0 years) who differ on volumetric measurements of total brain parenchyma and white matter hyperintensity (WMH) volumes. METHODS: Group comparisons were made of cerebrovascular disease risk factors measured at the time of imaging and at three prior examinations extending over 25 years of adult life. Measures of verbal memory and speed psychomotor processing at the time of imaging and 10 years before imaging were also available. RESULTS: Compared with those in the "nonatrophy" group, individuals in the subgroup with "atrophy" (defined by low total brain volume and high WMH volume) were older, reported a higher level of depressive symptomatology, experienced a steeper decline in diastolic blood pressure (DBP) and a steeper increase in pulse pressure, were less physically active, had smoked for more years, and had a higher prevalence of several cardiovascular disease indicators, including an ankle/arm systolic blood pressure ratio less than 0.9, and hypertension. After multivariate analysis, the 25-year decline in DBP, the number of years smoked, and an ankle/arm index of less than 0.9 remained significant discriminators of the two groups. Lower levels of speeded performance at the time of imaging and a steeper 10-year decline in cognitive performance on selected tests were also observed in the atrophic group. CONCLUSION: Community-dwelling older adults with volumetric brain measurements associated with accelerated aging are distinguishable on the basis of several health-related characteristics. These individuals also perform less well on certain tasks involving executive functioning. PMID- 10851373 TI - Access www.neurology.org now for full-text articles PMID- 10851374 TI - A placebo-controlled trial of lamotrigine for painful HIV-associated neuropathy. AB - OBJECTIVE: To investigate the analgesic efficacy of lamotrigine in the treatment of painful HIV-associated distal sensory polyneuropathy (DSP). BACKGROUND: The pathogenesis of HIV-associated DSP is unknown and there is no effective treatment. A novel anticonvulsant, lamotrigine, blocks voltage-sensitive sodium channels and inhibits the release of glutamate and aspartate. There have been anecdotal reports of efficacy of lamotrigine in the treatment of painful neuropathy and trigeminal neuralgia. METHODS: In a multicenter, randomized, double-blind, placebo-controlled study, lamotrigine was initiated at 25 mg per day and slowly titrated over 7 weeks to 300 mg per day. Study duration was 14 weeks. The primary outcome measure was change in pain on the modified Gracely scale with secondary outcome measures including change in neurologic examination, use of concomitant analgesic medications, and global pain relief. RESULTS: Of 42 enrolled subjects, 13 did not complete the 14-week study endpoint. In five of these, rash was the cause for dropout. In the remaining 29 evaluable subjects, 20 patients received placebo and 9 received lamotrigine. The pain scores at baseline were not significantly different. The reduction in average pain from baseline to week 14 was greater (p = 0.03) in the lamotrigine group (-0.55) than in the placebo group (-0.18), adjusting for baseline levels of pain. There was no difference between the groups on the change in peak worst pain. CONCLUSIONS: In this small trial, lamotrigine showed promise in the treatment of pain associated with HIV-related DSP. The frequency of rash was greater than in lamotrigine studies in epilepsy. A larger controlled study of lamotrigine is warranted. PMID- 10851375 TI - Antiretroviral therapy may improve sensory function in HIV-infected patients: a pilot study. AB - OBJECTIVE: To evaluate thermal and nociceptive function in a prospective, longitudinal study of 49 consecutive HIV-1-infected patients before and at 1, 4, and 8 months after initiation of highly active antiretroviral therapy. METHODS: Quantitative assessments of thermal perception thresholds for warmth (dWT), cold (dCT), and heat pain (HPT) were performed. CD4+ cell levels in blood and HIV-1 RNA levels in plasma were determined. Depending on the virologic response to treatment, the patients were divided into two groups: responders (37 of 49, 76%) and nonresponders (12 of 49, 24%). RESULTS: Before treatment, impairment of dWT was found in 26 of 49 patients, of dCT in 33 of 49 patients, and of HPT in 19 of 49 patients. Improvements of perception thresholds for dWT (p < 0.0001), dCT (p < 0.001), and HPT (p < 0.01) were observed after 8 months of treatment in the responder group but not in the nonresponders. Within the responder group, improved thermal perception thresholds was associated with higher pretreatment CD4+ levels than in patients without improvement. CONCLUSIONS: Virologically successful antiretroviral combination therapy of HIV-1-infected patients has a capacity to improve function of the thermal and nociceptive systems, especially in patients with less advanced immunodeficiency. PMID- 10851376 TI - Poor verbal working memory across intellectual level in boys with Duchenne dystrophy. AB - OBJECTIVE: To determine whether all boys with Duchenne muscular dystrophy (DMD) have a similar verbal and memory profile of skills, or whether only a subset is affected, and to determine whether the weak areas in their profile are substantially different from a control group. METHODS: Performance of patients with DMD on neuropsychological tests of verbal and memory skills was examined in two ways. Standardized test scores for 80 boys with DMD (estimated IQ range, 70 to 160) were ranked individually from worst to best, and the individual rankings were compared across the group using Friedman rank analysis. Additionally, performance of 41 boys with DMD was compared with that of their sibling control subjects of similar age and estimated IQ using multivariate analysis of variance. RESULTS: Individual cognitive profiles were significantly similar among the subjects with DMD, such that for most subjects digit span, story recall, and comprehension were the tests on which each performed most poorly. This finding remained true regardless of whether they were of high or low intellectual function. In contrast, no significant cognitive profile was found among their sibling control subjects, and when compared with their siblings, the DMD group scored significantly more poorly on digit span, comprehension, and story recall, but not on other verbal and memory measures. CONCLUSIONS: Boys with DMD have a specific cognitive profile, regardless of their general level of cognitive function. Specifically, boys with DMD performed more poorly on tests requiring attention to complex verbal information than they did on other verbal or memory measures. The possibility that the missing dystrophin brain products may contribute to selective cognitive processing is considered. PMID- 10851377 TI - A three-sister sibship of Gerstmann-Straussler-Scheinker disease with a CJD phenotype. AB - OBJECTIVE: To describe a rare phenotypic variant of P102L Gerstmann-Straussler Scheinker disease (GSS). BACKGROUND: Classic GSS is characterized by an early age at onset, prominent cerebellar signs with a slowly evolving dementia, and a neuropathology including multifocal PrP-positive plaques and variable but usually modest spongiform change. METHODS: Clinical, neuropathologic, immunohistochemical, and molecular genetic analysis of three sisters in a Hungarian family was performed. RESULTS: The clinical course of all three sisters was indistinguishable from sporadic Creutzfeldt-Jakob disease (CJD). Neuropathologic examination revealed spongiform changes, PrP (prion)-positive unicentric "kuru" or multicentric plaques, and abundant beta-A4-positive senile plaques. Molecular genetic analysis of the PRNP gene showed the heterozygous codon P102L mutation of classic GSS, with the methionine encoding allele of a heterozygous codon 129 coupled to the mutant 102 allele. CONCLUSION: The authors report the second recorded example of a sporadic CJD phenotype occurring in association with the P102L GSS genotype, and the first instance in which the phenotype was the rule rather than the exception, or was associated with prominent beta-A4 plaque formation. PMID- 10851378 TI - Charles Skinner Hallpike and the beginnings of neurotology. AB - Although the symptom of vertigo had been well-recognized for several thousand years, it was not until the pioneering work of Prosper Meniere in the mid-19th century that it was appreciated that vertigo could originate from damage to the inner ear. Before that time, patients with vertigo (regardless of the cause) were said to have "cerebral congestion," a condition resulting from excessive blood filling the brain. Bloodletting and leeches to relieve the congestion were the treatment of choice. The discovery of endolymphatic hydrops in temporal bone specimens from patients with Meniere's disease by Hallpike and Cairns in 1938 marked the beginning of modern neurotology. For the first time, vertigo was correlated with specific pathophysiology. Propelled by his temporal bone work, Charles Hallpike received an appointment at the National Hospital at Queen Square, where he developed an internationally renowned neurotology clinic. His bithermal caloric test and positional tests are still routinely used in evaluation of the vertiginous patient. PMID- 10851379 TI - Multiple sclerosis flares associated with recombinant granulocyte colony stimulating factor. AB - Four of 10 patients who were enrolled on protocols of high-dose immunosuppression with peripheral blood stem cell rescue for MS experienced neurologic worsening while receiving recombinant human granulocyte colony-stimulating factor. There was improvement when methylprednisolone was given to three of the patients, but one patient died of respiratory failure. The mechanism of the neurologic worsening is uncertain. PMID- 10851380 TI - Hippocampal sulcal cavities on MRI: relationship to age and apolipoprotein E genotype. AB - Hippocampal sulcal cavities are usually considered incidental findings on brain MRI. In a group of 92 elderly volunteers, the authors graded the number and size of hippocampal sulcal cavities with brain MRI to obtain a cavity score. Cavity scores increased with age, and were higher in subjects carrying the APOE epsilon4 or epsilon2 alleles than in subjects with the epsilon3/3 genotype. Age-related morphologic changes in the hippocampus may be mediated by the APOE genotype. PMID- 10851382 TI - Activate your online subscription PMID- 10851381 TI - Status epilepticus associated with cefepime. AB - Cefepime is a fourth-generation cephalosporin widely used for gram-negative sepsis. The authors report two patients in whom nonconvulsive status epilepticus developed while they were on treatment with cefepime for Pseudomonas aeruginosa infection. The status epilepticus resolved completely once the drug was withdrawn. Cefepime therapy can result in status epilepticus, especially if given in higher doses than required. PMID- 10851383 TI - Interobserver variability of cisternal blood on CT after aneurysmal subarachnoid hemorrhage. AB - Interobserver variability in the prediction of delayed cerebral ischemia by means of blood on CT was investigated in 159 patients with aneurysmal subarachnoid hemorrhage, admitted within 72 hours after the bleed. The authors found considerable interobserver variability in the assessment of the amount of blood in the individual cisterns. A high sum score was an independent predictor for delayed cerebral ischemia only for rater 1 (rater 1: hazard ratio, 3.26; 95% confidence interval [CI], 1.14 to 7.75; rater 2: hazard ratio, 1.72; 95% CI, 0.72 to 4.09). The authors conclude that interobserver variability limits the predictive power of the amount of blood on CT for the occurrence of cerebral ischemia. PMID- 10851384 TI - Intravenous tPA in acute ischemic stroke related to internal carotid artery dissection. AB - The authors describe the outcomes in 11 patients who had acute ischemic stroke related to internal carotid artery (ICA) dissection and were treated with IV tissue plasminogen activator (tPA). One symptomatic intracerebral hemorrhage occurred 36 hours after tPA was given. The mean day 90 modified Rankin Scale (m RS) score was 2.4 (+/-1.6). No death was observed at 3 months. Four patients of 11 (36.4%) made an excellent recovery (day 90 m-RS score: 0 to 1). This study demonstrates the feasibility of IV thrombolysis with tPA (0.8 mg/kg) in ischemic stroke related to ICA dissection within the first 7 hours. PMID- 10851385 TI - Gabapentin's effects on hot flashes and hypothermia. AB - The author describes six cases in which gabapentin treatment reduced the frequency of hot flashes. In addition, gabapentin treatment enhanced the frequency of hypothermic episodes in a separate patient with known hypothalamic dysfunction. Gabapentin may act directly upon temperature regulatory centers. PMID- 10851386 TI - Status epilepticus with neuron-reactive serum antibodies: response to plasma exchange. AB - The authors report a patient with partial and secondarily generalized status epilepticus who required 70 days of general anesthesia for seizure control. Although antiepileptic medications failed to control the seizures, they resolved with plasma exchange. The patient's serum reacted with rat cerebral cortex, hippocampus, and cerebellum, but not with cells expressing the glutamate receptor GluR3. These findings suggest an immune response against neuronal antigens other than GluR3. PMID- 10851387 TI - Temporal lobectomy outcome in older versus younger adults. AB - A total of 340 patients age 50 years and older were compared with 30 patients younger than 50 years, all of whom had anterior temporal lobectomy for refractory epilepsy. Seizure outcome, neuropsychological test scores, and change in driving status were analyzed. Age and duration of epilepsy were related independently to outcome, but laterality of interictal sharp waves (an early epilepsy risk factor) and presence of tumor were not. Sixteen patients (52%) in the older group and 257 patients (75.6%) in the younger group (p < 0.008) were seizure free. Postoperative neuropsychological outcome and driving status were similar in older and younger patients. PMID- 10851388 TI - Successful epilepsy surgery in catastrophic postencephalitic epilepsy. AB - The authors studied prognostic factors for surgical treatment in 22 patients with intractable postencephalitic epilepsy. A small subgroup of patients (9/22) had a positive outcome after resective surgery. They had a higher functional level after encephalitis as measured by Glasgow Outcome Score, a longer time interval between encephalitis and onset of seizures, and localization of ictal EEG to one temporal lobe. The other patients had devastating seizures with poor outcome after surgery. PMID- 10851389 TI - Autosomal dominant partial epilepsy with auditory features: defining the phenotype. AB - The authors previously reported linkage to chromosome 10q22-24 for autosomal dominant partial epilepsy with auditory features. This study describes seizure semiology in the original linkage family in further detail. Auditory hallucinations were most common, but other sensory symptoms (visual, olfactory, vertiginous, and cephalic) were also reported. Autonomic, psychic, and motor symptoms were less common. The clinical semiology points to a lateral temporal seizure origin. Auditory hallucinations, the most striking clinical feature, are useful for identifying new families with this synome. PMID- 10851390 TI - Lambert-Eaton myasthenic syndrome: electrodiagnostic findings and response to treatment. AB - The authors reviewed the incidence of cancer, repetitive nerve stimulation findings, and response to treatment in 73 patients with Lambert-Eaton myasthenic syndrome. Thirty-one patients (42%) had lung cancer, 29 small cell. Doubling of the compound motor action potential amplitude in three tested distal muscles was seen in only 41% of patients. Treatment with 3, 4-diaminopyridine produced moderate to marked self-reported functional improvement in 79% of the 53 treated patients. PMID- 10851391 TI - Temperature-sensitive sodium channelopathy with heat-induced myotonia and cold induced paralysis. AB - The authors report a Japanese family with dominantly inherited heat-induced myotonia and cold-induced paralysis with hypokalemia. This phenotype is associated with a novel mutation in the voltage-dependent skeletal muscle sodium channel alpha subunit (SCN4A). This Pro1158Ser mutation is localized between the fourth and fifth transmembrane segments of domain III in SCN4A and may give rise to a new function; that is, thermosensitive permeability changes of the sodium channel. PMID- 10851392 TI - Ineffective subthalamic nucleus stimulation in levodopa-resistant postischemic parkinsonism. AB - The authors report a patient with postischemic parkinsonism who responded neither to levodopa nor to bilateral subthalamic nucleus (STN) stimulation. MRI revealed bilateral lesions of the substantia nigra, the striatum, the external pallidum, and part of the internal pallidum. PET showed reduced striatal dopa-decarboxylase activity, D2 receptor binding, and glucose metabolism. Perioperative microrecording showed low-frequency activity of STN cells. This case suggests that parkinsonian patients who do not have a good response to levodopa or in whom a postsynaptic dopaminergic lesion can be shown may not be good candidates for STN surgery. PMID- 10851393 TI - Pseudoulnar palsy from a small infarct of the precentral knob. PMID- 10851394 TI - Myositis during long-term interferon-alpha treatment. PMID- 10851395 TI - Scoliosis in a dopa-responsive dystonia family with a mutation of the GTP cyclohydrolase I gene. PMID- 10851396 TI - New variant Creutzfeldt-Jakob disease presenting as localization-related epilepsy. PMID- 10851397 TI - Intravenous valproate in pediatric epilepsy patients with refractory status epilepticus. PMID- 10851398 TI - Long-term follow-up of aneurysms developed during extracranial internal carotid artery dissection. PMID- 10851399 TI - Tongue and limb myokymia in amyotrophic lateral sclerosis. PMID- 10851400 TI - Suppression of pendular nystagmus by smoking cannabis in a patient with multiple sclerosis. PMID- 10851401 TI - Defective slow inactivation of sodium channels contributes to familial periodic paralysis. PMID- 10851402 TI - Serum vitamin A is elevated in idiopathic intracranial hypertension. PMID- 10851403 TI - MRI of spinal cord infarction. PMID- 10851404 TI - Health care funding and surgical practice. PMID- 10851405 TI - A plea for early surgical intervention. PMID- 10851406 TI - Soft-tissue images. Insulinoma. PMID- 10851407 TI - Musculoskeletal images. Bone and soft-tissue tumours. PMID- 10851408 TI - Soft-tissue case 33. Hyperparathyroidism. PMID- 10851409 TI - Musculoskeletal case 10. Neuropathic foot. PMID- 10851410 TI - The role of the macrophage in periprosthetic bone loss. AB - Aseptic loosening after total joint replacement remains the most common reason for long-term implant failure. Macrophages activated by submicron wear particles of the polyethylene liner used in joint replacement have been shown to be the source of periprosthetic bone loss. Understanding the role of material chemistry in macrophage activation and the subsequent effects that macrophage-derived enzymes play in the degradation of implanted biomaterials is key to developing methods for prolonging the lifespan of implantable materials. PMID- 10851411 TI - The treatment of femoral shaft fractures in children: a systematic overview and critical appraisal of the literature. AB - OBJECTIVE: Through a critical systematic overview of the literature on the treatment of pediatric femoral shaft fractures to determine if any method of treatment can be recommended over others. DATA SOURCES: A MEDLINE search was performed for all cohort and randomized clinical trials for the years 1966 to 1996. STUDY SELECTION: Of 1217 identified articles, 15 cohort studies (where 2 or more treatments were compared in the same study) reported the treatment of children with femoral fractures. DATA EXTRACTION: Information was abstracted and articles rated for quality blind to author, institution and journal. DATA SYNTHESIS: Children having early application of a hip spica cast had an average hospital stay of 11 days (range from 5 to 29 days), average charges of $5784 (range from $590 to $11,800), average rates of limb-length discrepancy (greater than 2 cm) of 3% (range from 0 to 25%), angulatory malunion rates (greater than 10 degrees) of 8% (range from 0 to 19%), and rotational malunion rates (greater than 10 degrees) of 13% (range from 0 to 5%). The costs and malunion rates of early application of a hip spica cast were lower than for traction. Internal fixation (including intramedullary nails) had low angulatory malunion rates compared with early application of a hip spica cast but higher over-lengthening rates (greater than 2 cm) of 25% (range from 5% to 100%) and mean rotational malunion rates (greater than 10 degrees) of 25% (range from 11% to 32%). CONCLUSION: Early application of a hip spica cast had lower costs and malunion rates than traction. PMID- 10851412 TI - Lymph-node staining with activated carbon CH40: a new method for axillary lymph node dissection in breast cancer. AB - OBJECTIVE: To demonstrate the usefulness of activated carbon particles (CH40) as a vital staining dye for visualizing lymphatic vessels and lymph nodes in breast cancer. DESIGN: A retrospective evaluation. SETTING: Department of Surgery in Sendai National Hospital, Japan, a 716-bed teaching hospital. METHODS: To identify as many lymph nodes as possible in the axillary fat, by which we might decrease the possibility of the presence of undetected metastatic nodes, an emulsion of activated carbon particles (CH40) was injected into the centre of the mammary gland, close to the tumour site, 3 days before radical surgery. MAIN OUTCOME MEASURE: The number of lymph nodes found by the traditional method and by the CH40-injection method were recorded. RESULTS: After injection, the CH40 was readily adsorbed into regional lymphatics and streamed along with the lymph flow to blacken regional lymph nodes. The CH40-guided method increased the mean number of nodes per case found in the axilla from 8.4, by the traditional method, to 14.0 nodes per case. CONCLUSIONS: The use of the CH40 technique has two technical advantages; one is that it allows surgeons to locate the blackened lymph nodes at the time of surgery and the other is that it allows pathologists to look for the nodes in fatty tissue. Lymph-node dissection with the aid of activated carbon particles is inexpensive, easy to perform and enables the smallest lymph nodes to be easily recognized. CH40 is the technique of choice for the detection of axillary lymph nodes in cases where the number of lymph nodes detected by the traditional method is too small for accurate surgery. In conclusion, the present study demonstrates that CH40 could be an appropriate tool for more accurate staging of breast cancer axillary specimens. PMID- 10851413 TI - Acetabular blood flow during total hip arthroplasty. AB - OBJECTIVE: To determine the immediate effect of reaming and insertion of the acetabular component with and without cement on peri-acetabular blood flow during primary total hip arthroplasty (THA). DESIGN: A clinical experimental study. SETTING: A tertiary referral and teaching hospital in Toronto. PATIENTS: Sixteen patients (9 men, 7 women) ranging in age from 30 to 78 years and suffering from arthritis. INTERVENTION: Elective primary THA with a cemented (8 patients) and non-cemented (8 patients) acetabular component. All procedures were done by a single surgeon who used a posterior approach. MAIN OUTCOME MEASURE: Acetabular bone blood-flow measurements made with a laser Doppler flowmeter before reaming, after reaming and after insertion of the acetabular prosthesis. RESULTS: Acetabular blood flow after prosthesis insertion was decreased by 52% in the non cemented group (p < 0.001) and 59% in the cemented group (p < 0.001) compared with baseline (pre-reaming) values. CONCLUSION: The significance of these changes in peri-acetabular bone blood flow during THA may relate to the extent of bony ingrowth, peri-prosthetic remodelling and ultimately the incidence of implant failure because of aseptic loosening. PMID- 10851414 TI - Effect of mycophenolate mofetil in heart transplantation. AB - OBJECTIVE: To study the effect of mycophenolate mofetil (MMF), a new immunosuppressive drug that acts by inhibiting de novo pathways of purine synthesis, and rabbit antithymocyte globulin (RATG) on the lymphocyte subpopulation after heart transplantation. DESIGN: A review of clinical and laboratory records. SETTING: The Montreal Heart Institute. PATIENTS: Thirty-one patients who underwent heart transplantation. In 9 patients, neoral cyclosporine, prednisone and azathioprine were administered (group 1). In 14 patients RATG was added during the first 3 postoperative days (group 2) and in 8 patients RATG and combination immunosuppression was given, but MMF was used instead of azathioprine (group 3). The demographic characteristics of donors and recipients were similar among the 3 groups. MAIN OUTCOME MEASURES: The proportion of CD2, CD4 and CD8 receptor-positive lymphocytes, expressed as a mean (and standard deviation) percentage of the total lymphocyte population, measured at 7, 15 and 30 days and 6 months after transplantation. RESULTS: At 7 days after transplantation, CD2 lymphocytes averaged 55% (18%), 16% (15%) and 14% (11%) in groups 1, 2 and 3 respectively (p < 0.05), CD4 averaged 36% (11%), 9% (12%) and 7% (8%) in groups 1, 2 and 3 (p < 0.05), and CD8 averaged 14% (6%), 4% (3%) and 4% (3%) in groups 1, 2 and 3 (p < 0.05). At 15 days after transplantation CD2 averaged 69% (10%), 42% (16%) and 47% (20%) in groups 1, 2 and 3 respectively (p < 0.05), and CD8 averaged 16% (7%), 16% (6%) and 19% (7%) (p = NS). At 30 days after transplantation the percentages of CD2, CD4 and CD8 lymphocytes were similar among the groups. The freedom rate from acute rejection averaged 22% (14%), 9% (8%) and 50% (18%) (p < 0.05) in groups 1, 2 and 3 at 6 months after transplantation, and the freedom rate from infection averaged 56% (17%), 36% (13%) and 38% (17%) for the 3 groups at this time period (p = NS). CONCLUSIONS: A short course of RATG causes severe, transitory depletion of CD2, CD4 and CD8 lymphocyte subpopulations. MMF decreases the incidence of early acute rejection after heart transplantation without affecting the lymphocyte subpopulation when compared with azathioprine. PMID- 10851415 TI - Is clinical examination an accurate indicator of raised intra-abdominal pressure in critically injured patients? AB - OBJECTIVES: To determine the rate of elevated intra-abdominal pressure (IAP) and to evaluate the accuracy of clinical abdominal examination in the assessment of IAP in the critically injured trauma patient. DESIGN: A prospective blinded study. SETTING: The medical-surgical critical care unit of a university affiliated regional adult trauma centre. PATIENTS: Forty-two adult blunt trauma victims, who had a mean injury severity score of 36. INTERVENTIONS: Urinary bladder pressure was measured daily and classified as normal (10 mm Hg or less), elevated (more than 10 mm Hg) or significantly elevated (more than 15 mm Hg). A blinded clinical assessment of abdominal pressure was concurrently performed and recorded as elevated or normal. MAIN OUTCOME MEASURES: The sensitivity, specificity and accuracy and the positive and negative predictive values of the 2 interventions in identifying elevated IAP. RESULTS: Twenty-one patients (50%) had an elevated IAP at some point during the study. Of the 147 bladder pressure measurements done in these 42 patients, 47 (32%) were more than 10 mm Hg and 16 (11%) were more than 15 mm Hg. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy of clinical abdominal examination for identifying elevated IAP were 40%, 94%, 76%, 77% and 77%, respectively. Clinical abdominal examination had a sensitivity, specificity, positive predictive value, negative predictive value and accuracy of 56%, 87%, 35%, 94% and 84% respectively, for significantly elevated IAP. CONCLUSIONS: Urinary bladder pressure was commonly elevated among our population of critically injured adults. Compared with bladder pressure measurements, clinical abdominal assessment showed poor sensitivity and accuracy for elevated IAP. These findings suggest that more routine measurements of bladder pressure in patients at risk for intra-abdominal hypertension should be performed. PMID- 10851416 TI - Perioperative blood transfusion and albumin administration are independent risk factors for the development of postoperative infections after colorectal surgery. AB - OBJECTIVES: To determine whether transfused colorectal surgery patients were at increased risk for postoperative infections in a tertiary care teaching hospital and whether transfusion alone was the only significant risk factor. DESIGN: A retrospective study. SETTING: A single tertiary care teaching hospital. PATIENTS: All patients admitted to St. Boniface General Hospital, Winnipeg, for colorectal surgery during the period Apr. 1, 1995, through Mar. 31, 1996, were studied (N = 154). RESULTS: The overall infection rate was 17%: nontransfused patients, 13%, and transfused patients, 28% (p < 0.038). Patients who received albumin perioperatively had a significantly higher infection rate (38%) than those who did not (13%) (p < 0.001). Stepwise logistic regression analysis identified transfusion and albumin administration as the only independent risk factors for postoperative infection. CONCLUSION: Perioperative transfusion or albumin administration significantly increases the risk of postoperative infection in colorectal surgery patients. PMID- 10851417 TI - Osteochondritis dissecans of the talar dome treated with an osteochondral autograft. PMID- 10851418 TI - Ex vivo liver resection. PMID- 10851419 TI - Umbilical pilonidal sinus. PMID- 10851420 TI - Compartment syndrome of the right anterior thigh after primary total hip arthroplasty. PMID- 10851421 TI - A surgical sabbatical in France. AB - During my stay in France I had the unique opportunity to meet surgical professors from all over the world and made many friends and contacts in the field of hepatobiliary surgery. Brittany is a beautiful province of France, having unique way of life and approach to social and societal problems. The cultural enrichment that I received from my year there will last a lifetime, as well the many fond memories of the people, the culinary delights and the spectacular seashore. PMID- 10851422 TI - Metastatic carcinoma masquerading as primary carcinoma. PMID- 10851423 TI - Biotherapy of cancer marches on! PMID- 10851424 TI - Rhenium-188--a generator-derived radioisotope for cancer therapy. AB - Rhenium-188 (188Re) is an important therapeutic radioisotope which is obtained on demand as carrier-free sodium perrhenate by saline elution of the tungsten 188/rhenium-188 generator system. With a half-life of 16.9 hours and emission of a high energy beta particle (maximal energy of 2.12 MeV) and a gamma photon (155 keV, 15%) for imaging, 188Re can be provided at reasonable costs for routine preparation of radiopharmaceuticals for cancer treatment. PMID- 10851425 TI - Combined modality radioimmunotherapy with Taxol and 90Y-Lym-1 for Raji lymphoma xenografts. AB - Despite effective therapies for non-Hodgkin's lymphoma (NHL), the majority of patients are not cured. Radioimmunotherapy (RIT) has shown good results in preclinical and clinical trials even in patients that are non-responsive to standard chemotherapy. To make RIT more effective, agents such as paclitaxel (Taxol), that can enhance radiation effects, are being tested. Nude mice bearing human Burkitt's lymphoma (Raji) xenografts were treated with: 1) 150 or 200 microCi (5.5 or 7.3 MBq) of 90Y-2IT-BAD-Lym-1 alone, 2) 600 micrograms of Taxol alone, 3) 150 or 200 microCi of 90Y-2IT-BAD-Lym-1 plus 600 micrograms of Taxol given 24 hours after RIT, or 4) no treatment. Tumor size, survival, mouse weight and blood counts were monitored to assess efficacy and toxicity. Survival for mice treated in this 84 day trial was: 71% for 90Y-2IT-BAD-Lym-1 (200 microCi) plus Taxol, 29% for Taxol alone, 6% for 90Y-2IT-BAD-Lym-1 (200 microCi) alone and 14% in the untreated group. Average tumor volume in the 90Y-2IT-BAD-Lym-1 (200 microCi) plus Taxol group was reduced by 89 and 99% compared to the RIT alone and Taxol alone groups, respectively. Mice treated with 150 microCi had less toxicity than those treated with 200 microCi of 90Y-2IT-BAD-Lym-1, however, the higher radiation dose, and Taxol, were required for improved survival. Mouse weights and myelotoxicity in the combined modality (RIT plus Taxol) groups were similar to those receiving the same dose of RIT alone. In the Raji tumored nude mouse model, addition of Taxol to 90Y-2IT-BAD-Lym-1, in doses clinically achievable in humans, provided therapeutic synergy without increased or excessive toxicity. PMID- 10851426 TI - Liposomal muramyl tripeptide (CGP 19835A lipid) therapy for resectable melanoma in patients who were at high risk for relapse: an update. AB - Liposome-encapsulated muramyl tripeptide-phosphatidyl ethanolamine (L-MTP-PE) was used in a pilot study for resectable melanoma patients who were at high risk for relapse. We entered 18 evaluable patients. The patient group included: (a) patients with stage III disease and clinically measurable regional metastases at presentation as confirmed by needle biopsy and (b) patients with stage IV disease presenting with measurable and resectable distant metastases confirmed by needle biopsy and limited to lungs, lymph nodes and subcutaneous tissues. L-MTP-PE was given for 4 weeks prior to surgical resection and for an additional 20 weeks postoperatively. Disease-free intervals were then determined based on the date of surgery. A preliminary report published in 1993 indicated an average disease-free interval of 18 months (range 8-33 months). This article presents an updated report on the long-term, disease-free survival status of these patients and shows that of the 18 evaluable patients, 4 remain free of disease for more than 5 years after surgical resection and therapy. The period of survival for these patients ranged from 69 months to more than 91 months (average 80.5 months). Although this was only a pilot study, we believe that the duration of survival indicates that L MTP-PE may produce significant biologic activity in patients with melanoma, resulting in long-term benefits in terms of tumor eradication. PMID- 10851427 TI - Loss of VLA-3 (CD49c/CD29) expression in two multidrug resistant Burkitt's lymphoma cell lines. AB - Expression of P-glycoprotein (P-gp) mediated multidrug resistance (MDR) has been suggested to be associated with an impaired clinical outcome in several malignancies. In contrast to P-gp itself, further phenotypical and functional alterations related to MDR are poorly characterized. In this in vitro study, we analyzed two Burkitt's lymphoma cell lines (Raji and Daudi) for the beta 1 integrin phenotype prior to and after induction of MDR via co-cultivation with vincristine. A significant loss of the VLA-3 (CD49c/CD29) adhesion receptor was observed whereas all other intergins analyzed lacked considerable changes. We conclude that induction of P-gp mediated MDR does not only affect resistance to cytotoxic drugs but also induces cellular changes with potential relevance for migratory and/or adhesive properties of malignant cells. PMID- 10851428 TI - Constitutive expression of interferon beta in differentiated epithelial cells exposed to environmental stimuli. AB - The body's first line of defense against external challenge are the epithelial cells that line the skin and the respiratory, digestive, and genitourinary tracts. Inasmuch as interferon-beta (IFN-beta) participates in host defense against viral, bacterial, and parasitic infections and tumors, we hypothesized that this secreted protein is expressed in various murine epithelial cell types that line portals of entry to the body. We used immunohistochemistry and in situ hybridization techniques to measure IFN-beta expression in the various epithelial cell types and in internal murine organs sheltered from environmental stimuli. The epithelial cell types lining the skin, digestive tract, urinary tract, reproductive tract, and upper respiratory tract constitutively expressed IFN beta. Specifically, all differentiated epithelial cells at risk of environmental exposure expressed IFN-beta (protein and mRNA) with the exception of the ciliated epithelial cells lining the lower respiratory tract. Epithelial cells of internal organs that are not directly exposed to external pathogens did not express IFN beta. PMID- 10851429 TI - Elevated plasma levels of interleukin-1 receptor antagonist are associated with decreased cellular BCL-2 oncoprotein expression in B-chronic lymphocytic leukemia. AB - Plasma IL-1Ra levels and cellular BCL-2 oncoprotein expression were measured in a total of forty blood samples obtained from twenty-eight B-CLL patients and four healthy subjects. High IL-1Ra plasma levels (as defined by mean + three times standard deviation of normal controls) were observed in eleven samples (ten patients) which showed a significantly decreased cellular expression of BCL-2 protein (14.7 +/- 16.3% of cells as determined by immunofluorescence) when compared to B-CLL samples with no elevated IL-1Ra (BCL-2, 31.0 +/- 18.6%; p < or = 0.0115). Albeit correlational only, our results may encourage further experiments to elucidate potential regulatory effects of IL-1Ra for cellular BCL 2 oncoprotein expression. PMID- 10851430 TI - Lovastatin inhibits tumor growth and metastasis development of a rat fibrosarcoma. AB - HMG-CoA (3-hydroxy-3-methylglutaryl coenzyme A) reductase, the rate limiting enzyme in cholesterol synthesis, catalyses mevalonate production and, hence, influence the synthesis of isoprenoid metabolites. It has already been demonstrated that products of the mevalonate pathway play an important role in the progress of the cell cycle and cell survival. Lovastatin (LOV) competitively inhibits HMG-CoA reductase, blocking the synthesis of mevalonic acid and the generation of non-sterol isoprenoids, such as farnesyl residues. The posttranslational farnesylation of p21ras protein is essential for its binding to the membrane and, therefore, for its transforming activity. Considering that p21ras protein was reported to have a significant rol in metastatic behavior of tumor cells, we decided to study LOV as an antimetastatic agent on a rat fibrosarcoma. We demonstrated that a short treatment with LOV diminished primary tumor growth and the number and size of lung experimental metastasis. PMID- 10851431 TI - Impaired proliferation and DNA synthesis of a human tumor cell line (HeLa) caused by short treatment with the anti-anemic drug jectofer (ferric-sorbitol-citrate) and the lipid peroxidation product 4-hydroxynonenal. AB - Anti-anaemic drug, ferric-sorbitol-citrate complex (FSC), inhibit tumour cell growth through the mechanisms which are complex and not entirely understood. The probable mechanisms of described effects of iron is iron-induced oxidative stress of the treated cells. Hence, the effects of FSC on HeLa cell growth in vitro were compared with the biological activity of one of the major mediators of the oxygen free radicals--aldehyde 4-hydroxinonenal (HNE), to see if the effects of FSC and of HNE resemble each other. Impaired proliferative ability and DNA synthesis of HeLa cells was observed after treatment with anti-anaemic drug FSC for 24 hours. After treatment with FSC and culturing of HeLa cells in fresh medium for 24 or 96 hours the cells did not proliferate at all, DNA synthesis was transiently recovered and then diminished again. HNE blocked cell proliferation during the time the aldehyde was present in culture and 24 h later. Afterwards, the cells proliferated as control non-treated cells. HNE did not inhibit DNA synthesis during treatment, but intensity of 3H-thymidine incorporation was lower after preincubation. Thus, both FSC and HNE interfere with the basic mechanisms of the cell growth regulation, while antitumour activity of FSC resembles, but does not necessarily include iron induced lipid peroxidation. PMID- 10851432 TI - Effect of gold on survival of tumor-bearing mice. AB - Sodium aurothiomalate (monogold monosodium mono-hydrogen sulfidobutanedioate), an anti-rheumatoid gold agent, was injected s.c. every other day for three times or given in drinking water daily for two weeks to Balb/C mice that were inoculated with syngeneic Meth/A cells i.p. As the control, cisplatinum was injected s.c. Survival curve was then observed and significance was determined by Wilcoxon's method. Statistically significant effects on survivals was obtained by 30 mg/kg/day s.c. or 75 mg/kg/day p.o. of gold. A dose of less than 12.5 mg/kg/day s.c. adversely affected the survival. Cisplatinum was significantly effective at the dose of 12.5 mg/kg/day, but markedly toxic at the dose of 125 mg/kg and adversely effective at 6 mg/kg/day s.c. The range of dose effectiveness of cisplatinum on survival was narrower than that of gold. Gold was effective when given s.c. or p.o. No significant toxicity of gold was observed at doses up to 125 mg/kg/day. PMID- 10851433 TI - Bone pain palliation. PMID- 10851434 TI - Palliative therapy with bone seeking radiopharmaceuticals. AB - Pain palliation with unsealed source therapy is an effective and cost effective treatment for patients with cancer metastatic to bone who present with progressive pain. Used in the appropriate setting this form of therapy can make an enormous contribution to a very large number of patients. PMID- 10851435 TI - 99mTc-ethylenedicysteine-folate: a new tumor imaging agent. Synthesis, labeling and evaluation in animals. AB - It is known that membrane folic acid receptors are responsible for cellular accumulation of folate and folate analogs such as methotrexate and overexpressed on various tumor cells. However, these receptors are highly restricted in normal differentiated tissues. Results of limited in vitro and in vivo animal studies suggest that folate receptors could be a potential target for tumor imaging. This study aimed to develop a 99mTc-labeled folic acid using ethylenedicysteine (EC) as a chelator and evaluate its labeling efficiency and potential use as a tumor seeking agent. Tissue distribution of 99mTc-EC-folate was determined in breast tumor-bearing rats at 20 min, 1, 2, and 4 h (n = 3/time interval, 370 KBq/rat, i.v.). Blocking study was employed to determine receptor-mediated process; 99mTc EC-folate was co-administrated with 50 and 150 mumol/kg of cold folic acid to tumor-bearing rats. Planar imaging and whole-body autoradiograms were performed. The data was compared to that using 99mTc-EC (control). In animal studies, tumor/blood count density ratios at 20 min-4 h increased from 0.81 +/- 0.09 to 1.23 +/- 0.13 with 99mTc-EC-folate. Conversely, these values showed time dependent decrease from 0.77 +/- 0.32 to 0.65 +/- 0.01 with 99mTc-EC in the same time period. Tumor/muscle and tumor/blood count density ratios significantly decreased with folic acid co-administrations. Planar images and autoradiograms confirmed that the tumors could be visualized clearly with 99mTc-EC-folate. PMID- 10851436 TI - Antitumor effect of oxycellulose as a hemostatic during operation. AB - Oxycellulose, a hemostatic agent used in operation showed antitumor effect in vitro on a murine hepatic cell carcinoma (MH 134), a murine fibrosarcoma (Meth A) and a murine colon carcinoma (Colon 26). The effect was also confirmed in vivo by the survival of mice inoculated with Meth A or MH 134. Eighty milligrams per mouse of this agent, however, showed a toxicity rather than an antitumor effect. The antitumor effect of oxycellulose on Meth A did not compare with that of etoposide or mitomycin C in vivo. The antitumor effect on MH 134 was equal to that of etoposide but not mitomycin C. Oxycellulose inhibited tritium thymidine uptake into Colon 26 cells to the same extent as 5-fluorouracil and mitomycin C and it caused 51Cr-labelled Colon 26 cells but not from 5-fluorouracil or mitomycin C. Oxycellulose decreased a larger number of viable tumor cells than 5 fluorouracil or mitomycin C when the tumor cells were incubated for 24 hours at 37 degrees C. DNA histogram with MH 134 cells showed oxycellulose decreased a ratio of tumor cells in S-phase. These results suggest that the antitumor effect of oxycellulose is cytocidal and phase-specific. PMID- 10851437 TI - Anti-tumor effect of methanol extracts from red and white wines. AB - Crude methanol extracts of red and white wines were added to diethyl ether in order to divide them into the anthocyanin fraction (insoluble in diethyl ether) and fractions containing other flavonoids and their derivatives (soluble in diethyl ether). However, the white wine did not contain anthocyanins (all of the methanol extract was soluble in diethyl ether). When HCT-15 cells, derived from human colon cancer or AGS cells, derived from human gastric cancer, were cultured with these fractions, the anthocyanin fraction from the red wine and the non anthocyanic substances extracted from red and white wines suppressed the growth of the cells, and the suppression rate by the anthocyanin fraction was significantly higher than that of the other fractions. Thin-layer chromatographic analysis revealed mostly delphinidin in the anthocyanin fraction. The other fractions contained mostly flavonoids and their derivatives. The sugars in all fractions were mainly glucose, fucose, and fructose. Flow cytometric study suggested that the anthocyanin fraction blocked mostly S, G2, and M phase, and the non-anthocyanic flavonoids also blocked these phases, although the histographic pattern varied depending on the fractions. Methanol insoluble but water soluble fractions (mostly free sugars) of red and white wines did not show such suppressive effects. PMID- 10851438 TI - Suppression by radiosensitizer AK-2123 of early phorbol myristate acetate stimulated chemiluminescence response of human neutrophils induced by gamma irradiation (short communication). AB - The influence of gamma-irradiation and radiosensitizer AK-2123 on secretion of reactive oxygen species (ROS) of human neutrophils was studied. gamma-Irradiation (from 5 to 25 Gy) of the neutrophil suspension inhibited spontaneous chemiluminescence and activated phorbol-12-myristate-13-acetate (PMA)-stimulated chemiluminescence. The time of maximum PMA-stimulated chemiluminescence amplitude was decreased with the dose range from 2 to 25 Gy. The addition of radiosensitizer AK-2123 depressed the early PMA-stimulated chemiluminescence response to gamma-irradiation. The obtained results suggest that this effect is connected with influence of AK-2123 on ion canals of neutrophils and may prevent the radiation-induced damage of blood cells. PMID- 10851440 TI - Cancer and diabetes: are there similarities? PMID- 10851439 TI - Evaluation in a mouse model of a thyroid-blocking protocol for 131I antibody therapy (short communication). AB - PURPOSE: We describe a murine model to evaluate variations of a published multicenter thyroid blocking protocol described for 131I antibody therapy, using doses of blocking agents proportional to those used in man. Variables include duration of super-saturated potassium iodide (SSKI) pretreatment and use of supplemental KClO4. MATERIALS AND METHODS: Whole-body activity measurements were performed 0, 24, 48 and 72 hours following 131I-NaI administration in control and thyroid-blocked mice. Retained whole-body activity was calculated as a percentage of the injected dose (%ID), primarily reflecting radioiodine sequestered in the thyroid gland. In blocked groups, SSKI was begun one or 7 days preceding 131I-NaI therapy, and was supplemented in one half of the cases with KClO4 from time of therapy. RESULTS: In control mice, %ID was 11.23 +/- 1.47%, 10.15 +/- 1.11% and 9.29 +/- 1.50% at 24, 48 and 72 hrs respectively. %IDs of blocked groups were markedly lower than controls (p = .0001). In the one day SSKI pretreatment group, %ID was reduced from 1.73 +/- 0.58, 1.42 +/- 0.45 and 1.20 +/- 0.38 at 24, 48 and 72 hours to 0.49 +/- 0.08, 0.50 +/- 0.07 and 0.44 +/- 0.06 with addition of supplemental KClO4. In the 7 day SSKI pretreatment group, %ID was reduced from 1.87 +/- 0.73, 1.48 +/- 0.49 and 1.36 +/- 0.57 at 24, 48 and 72 hours to 0.60 +/- 0.36, 0.45 +/- 0.13 and 0.41 +/- 0.14 with addition of supplemental KClO4. %IDs in the 7 day pretreatment animals were not statistically different from those in the one day pretreatment groups (all p >> 0.05). CONCLUSION: SSKI reduces retention of radioiodide approximately six-fold whereas supplemental KClO4 enhances thyroid blocking an additional three-fold. Seven day SSKI pretreatment appears no more effective than one day pretreatment. PMID- 10851441 TI - Hybrid high-dose bolus/continuous infusion interleukin-2 in patients with metastatic renal cell carcinoma: a phase II trial of the National Biotherapy Study Group. AB - BACKGROUND: Interleukin-2 (IL-2) is an active agent for the treatment of renal cell carcinoma. In animal studies, polyethylene glycol conjugated (PEG) IL-2 was found to be effective in certain IL-2-resistant models. When bolus/infusion IL-2 was administered to approximate the pharmacokinetics of PEG-IL-2, resistance was also overcome in these models. Based on these observations, the National Bio therapy Study Group (NBSG) previously had conducted a pilot study (NBSG 90-01) and then a phase I trial of a hybrid regimen of bolus IL-2 followed by continuous IL-2 (NBSG 91-04). METHODS: In the current study, NBSG 92-09, a phase II trial was conducted in patients with metastatic renal cell carcinoma using IL-2 at a dose of 36 MIU/m2 followed by a 72-hour continuous infusion of IL-2 at 18 MIU/m2 per day, so that over 3 days a total of 90 MIU/m2 of IL-2 were delivered; the same amount as previously given during 5 days of continuous intravenous (i.v.) IL 2 at 18 MIU/m2 per day. This was repeated every 2 weeks for 2 months, and then monthly for up to 4 months. RESULTS: Thirty-one patients with a median age of 62 years were enrolled in this trial. During the first 4 biweekly treatments, the percentages of planned IL-2 administered were 98% for 31 patients, 99% for 27, 98% for 23, and 99% for 20 patients. Toxicities were qualitatively the same as those seen with other IL-2 regimens. During the first 2 months, 4 patients ceased treatment because of rapidly progressive disease while 7 patients stopped because of toxicity; 5 of the 7 were > 65 years of age. At the time of formal reassessment after 2 months of treatment, 7 additional patients had progressive disease for a treatment failure rate of 55% prior to monthly maintenance therapy. There were two partial responses among 22 patients who had measurable disease for a response rate of 9% (1 to 29%, 95% CI). Median survival was 10.2 months and failure-free survival (FFS) 3.4 months for the entire group. CONCLUSION: The response rate seen with this regimen is similar to those of other schedules of IL 2 requiring more prolonged hospitalization. This hybrid bolus/continuous infusion IL-2 schedule appears to be an equally effective and less expensive schedule of IL-2 administration than previously reported inpatient regimens. However, it is not likely that this regimen is superior to outpatient combination biotherapy regimens which are currently under investigation. PMID- 10851442 TI - Non-Hodgkin's lymphoma: cytotoxic T lymphocyte mediated activity correlated with histopathological classification and staging. AB - Non-Hodgkin's lymphomas (NHLs) constitute a heterogeneous group of lymphoid tumors and a majority of them in India are of B-cell phenotype. It has been postulated that immunoregulatory dysfunctions may be involved in the pathogenesis of NHL. Hence, peripheral blood mononuclear cells obtained from twenty six untreated patients were assessed for cytotoxic T lymphocyte mediated (CTL) activity in 51Cr release assay. Patients were classified according to Revised European American Lymphoma classification. B-cell small lymphocytic lymphoma patients showed lower CTL activity than NHL patients of other histopathological subtypes and healthy individuals. Diffuse large B cell lymphomas showed CTL activity comparable to healthy individuals. However, within the same histopathological subgroup, the CTL activity did not correlate with the stage of the patients. PMID- 10851443 TI - Apoptosis of tumor cells induced by substances of the circulatory system. AB - Despite global abnormalities of the immune system, such as in AIDS, the incidence of only a few kinds of tumor increases, and even in the development of these tumors the degree of immunosuppression seems not to be a critical factor. This means that the known immune system has no significant role in the tumor preventing mechanism. Thus, the fact that tumors do not develop in the majority of the population during their lifetime, indicates the existence of an additional defense mechanism of the immune system. We demonstrated previously that this defense is produced by the synergistic action of certain substances of the circulatory system. Here we report that the substances taking part in the defense induced, but only when they were used together, the apoptosis of tumor cells, but not normal cells, as was detected by different methods. Other substances of the circulatory system did not show similar effects. These results further support the existence of the mentioned defense mechanism called by us the Passive Antitumor Defense System. PMID- 10851444 TI - Improved immunostimulatory function of bone marrow derived macrophages transduced with the granulocyte-macrophage colony stimulating factor gene. AB - Professional antigen presenting cells (APCs) play a prominent role in generation of antitumor immunity. Granulocyte macrophage colony stimulator factor (GM-CSF) has been shown to increase antigen presenting capacity of macrophages and dendritic cells. We examined whether retroviral mediated gene transfer of the murine GM-CSF cDNA into bone marrow precursor cells would result in generation of mature APCs with improved immunostimulatory function. We show that murine bone marrow cells can be stably transduced to produce GM-CSF (200-300 pg/mL per 10(6) cells in 24 hours). These cells proliferated in the absence of exogenous growth factor for 25 days and expressed the macrophage markers Mac-1, Mac-3 and F4/80. GM-CSF transduced bone marrow cells had enhanced stimulatory capacity in a primary mixed lymphocyte-APC reaction and improved antigen presenting function in a T helper clone proliferation assay. These data demonstrate that bone marrow cells can be genetically engineered to secrete GM-CSF resulting in expansion of effective APCs. GM-CSF transduced APCs may be used as natural adjuvants in stimulating immune responses in vivo. PMID- 10851445 TI - Patterns of hepatic and splenic colonization by an attenuated strain of Salmonella typhimurium containing the gene for human interleukin-2: a novel anti tumor agent. AB - Currently, there is no effective treatment for unresectable hepatic malignancies. Salmonella sp. are known to naturally track to the liver during active infection. A live biological vector was developed for delivery of Interleukin-2 (IL-2) to the liver for anti-tumor purposes. The avirulent and highly immunogenic c4550 strain of Salmonella typhimurium was used to express the IL-2 protein [renamed c4550(pIL-2)]. We have previously demonstrated that the c4550(pIL-2) produces biologically active IL-2 (up to 46.2 IU/ml) and that a single gavage feeding of 10(7) colony forming units (cfu) of c4550(pIL-2) significantly reduced the number of hepatic metastases when compared to animals fed salmonella lacking the IL-2 gene or non-treated controls. The goal of the current studies was to determine the pattern of splenic and hepatic colonization of Salmonella-IL2. Hepatic and splenic colonization was determined following administration of 10(7) cfu of c4550(pIL-2) and c4550(pYA292) via a single gavage feeding to C57BL/6 mice. Five experiments of antibiotic regimen administration were conducted where splenic and hepatic homogenates were cultured after 14 days of parenteral and/or oral antibiotics. The natural history of hepatic and splenic colonization was also determined for animals without antibiotic treatment. Despite administration of various antibiotic regimens using different routes, eradication of salmonella with and without IL-2 was not achieved. Salmonella, however, was not cultured from hepatic and splenic tissue at 4 months after a single gavage feeding of salmonella with no specific treatment. In conclusion, oral administration of c4550(pIL-2) may represent a novel form of in vivo biotherapy for unresectable hepatic malignancies. Antibiotics do not accelerate eradication of this bacteria and it appears that c4550(pIL-2) follows the natural pathophysiological of salmonella infection in which eradication from the splenic and hepatic tissue occurs over a period of 2-4 months. PMID- 10851446 TI - Suppression of growth of cultured malignant cells by allomelanins, plant-produced melanins. AB - Allomelanins, which belong to the melanins produced by higher plants and fungi, were studied with respect to their growth suppressive effects on cultured HCT-15 cells derived from human intestinal carcinoma and Meth/A cells derived from a Balb/C mouse lymphoma. Allomelanins were extracted from black soy beans and black sesame seeds with 0.2% NaOH in water. Proteins bonded to the allomelanins were removed by boiling the extracts for 20 hours in 30% NaOH. Although native allomelanins conjugated with protein did not suppress the growth of the cells, protein-free allomelanins did suppress it. That is, 50% suppression of HCT- 15 cell growth was found at a concentration between 100 and 200 micrograms/ml of the melanin prepared from the black soy beans, and between 25 and 100 micrograms/ml of that from the sesame seeds. For Meth/A cells, 50% suppression by allomelanin from the black beans was approximately 100 micrograms/ml; and that by the allomelanin preparation from sesame seeds, between 25 and 100 micrograms/ml. Thus, protein-free allomelanins seemingly suppress cultured mammalian and animal tumor cells. PMID- 10851447 TI - Tumor cell growth suppression by chalcone (1,3-diphenyl-2-propen-1-one). AB - Chalcone or 1,3-diphenyl-2-propen-1-one which is widely distributed in higher plants and is considered to be a precursor of all flavonoids, was studies in terms of its anti-tumor growth activity in vitro and in vivo. Mice given the chalcone, which was suspended in the drinking water, showed a statistically significant rise in percent survival: 66% at the 35th day for the chalcone treated animals vs. 30% for the control ones. The chalcone also gave significant suppression of growth of HCT-15 and Meth/A cells in vitro. The dose of 50% reduction in growth was between 2.5 micrograms/ml and 0.6 microgram/ml for the HCT-15 cells and less than 10 micrograms/ml for the Meth/A cells. Histograms obtained by flow cytometric examination showed an elevated region between the diploid and the tetra-ploid nucleoidal peaks. A third peak heavier than the tetra ploid peak was also observed, which peak was not observed in the control. The percentage of cells in the S phase was significantly raised. Microscopically, large nucleated cells were observed following addition of the chalcone. These results suggest that the chalcone induced abnormal DNA synthesis and mitosis in the cultured cells. PMID- 10851448 TI - Synthesis and biodistribution of peptide based 99mTc/186Re-MAGIPG-D612 monoclonal antibody in nude mice bearing colon cancer xenografts. AB - A new bifunctional chelating agent MAGIPG was synthesized in good yield using Boc p-nitro-phenylalanine as the starting material. 99mTc-labeled-MAGIPG-D612 conjugates reactive with human colon cancer were prepared in 3% to 15% yield with a radiochemical purity of 63% to 97%. 186Re-labeled MAGIPG-D612 conjugates were prepared with a 7% to 30% yield and a radiochemical purity of 82% to 98%. The biodistribution results demonstrate that these radioimmuno-conjugates selectively localized in LS174T human colon cancer xenografts. PMID- 10851449 TI - Tumor localization by tumor infiltrating lymphocytes labeled with indium-111 in patients with metastatic renal cell carcinoma, melanoma, and colorectal cancer. AB - BACKGROUND: Adoptive immunotherapy with autologous tumor infiltrating lymphocytes (TIL) is a promising approach for cancer bio-therapy. One issue, however, is whether such cells actually migrate to sites of tumor after intravenous infusion. There have been several reports of tumor uptake of radiolabeled TIL in patients with metastatic melanoma, but efforts to visualize tumor with radiolabeled TIL in other tumor types reportedly have been unsuccessful. METHODS: Eight patients with metastatic cancer (5 renal, 2 melanoma, 1 colon) received an intravenous infusion of 2 to 100 billion autologous TIL, including 50 million TIL which had been conjugated to 500 microCi Indium-111, co-administered with interleukin-2 (IL-2). One patient received 1 gm/m2 of cyclophosphamide one day prior to TIL; seven patients received interferon alpha 2b for 4 days prior to receiving TIL. Total body gamma camera imaging, including single photon emission computerized tomography (SPECT), was performed at 24 and 48 hours. RESULTS: All eight patients had demonstrable uptake of 111-Indium-labeled TIL into one or more known sites of tumor. There were no known sites of tumor which were not imaged. Metastatic sites imaged included bone, brain, mediastinal and perihilar lymph nodes, lung and liver parenchyma, abdominal periaortic nodes, and a pelvic mass. One patient served as a negative control in that the TIL scan was negative at a time when she had no evident disease, but a few weeks later had a positive TIL scan which lead to a diagnosis of axillary recurrence. CONCLUSION: Uptake of radiolabeled TIL, whether CD8+ or CD4+, by metastatic renal cell carcinoma and other carcinomas was similar to that previously reported in melanoma. Pretreatment with cyclophosphamide was not a prerequisite for imaging, and TIL uptake did not predict tumor response. PMID- 10851450 TI - Various functions of PBMC from colon cancer patients are not decreased compared to healthy blood donors. AB - The immune surveillance hypothesis suggests impaired immune responses to participate in development of cancer. This may partly be due to increased amounts of PGE2 and histamine, which inhibit cellular immunity. These effects are mediated by cAMP, which is increased and thereby may down-regulate IL-2 and its receptor proteins in T helper cells. The proliferative responses and IL-2 synthesis of PBMC have earlier been shown to be reduced in patients with colon cancer. Recently immune modulating agents have been demonstrated to increase the proliferative response of PBMC in vitro, probably by inhibition of adenylate cyclase activity and induction of IL-2 mRNA expression. We have therefore studied the proliferative responses of PBMC from colon cancer patients to PWM and tested the effect of immune modulating agents, such as Serotonin, Sumatriptan, and Buspirone on these PBMC. We found no difference in levels of intracellular cAMP, IL-2 mRNA expression, IL-2R mRNA expression, or proliferative responses of PBMC from colon cancer patients compared to healthy blood donors. There was no effect of the immune modulating agents on PBMC from colon cancer patients. PMID- 10851451 TI - Correlation of toxicity with treatment parameters for 131I-CC49 radioimmunotherapy in three phase II clinical trials. AB - Analyses were performed on 40 patients with TAG-72 expressing metastatic cancer who were entered into three phase II clinical trials. The dose selected was the maximum tolerated dose in phase I studies. Patients all had unresectable metastatic colon or prostate cancer and had recovered from prior therapies. Patients in trials #1 and #2 received 75 mCi/m2 131I-CC49 antibody whereas those in trial #3 received a total of 75 mCi/m2 with equal amounts of 131I-CC49 and 131I-COL-1. The three trials have resulted in a reproducible degree of reversible marrow suppression; 72.5% of patients experienced moderate or severe toxicity. Comparisons were made between demographic, clinical and pharmacokinetical variables and the grade of WBC toxicity, platelet toxicity and the sum of the two as total toxicity. Whole body radiation dose had a statistically significant relationship with platelet toxicity (r = 0.38, p = 0.015) and total toxicity (r = 0.34, p = 0.035). The bone marrow radiation dose is significantly related to all toxicity indicators with correlation coefficients with WBC and platelet toxicities of 0.47 (p = 0.002) and 0.34 (p = 0.033), respectively. Plasma half life had the strongest correlation with WBC toxicity and combined toxicities. Multivariate models were developed to help describe the simultaneous effect of these variables on toxicity. The results show that the MTD dose was safely given to patients who varied in age, disease burden and degree of marrow compromise. This supports the contention that a fixed dose of radiolabeled antibody per body mass or m2 can be given to a diverse group of non-lymphoma patients with a predictable toxicity range. PMID- 10851452 TI - Comparison between transversal SPECT, 3-dimensional rendering and 3-dimensional rendering plus clipping in the diagnosis of somatostatin receptor distribution in malignancies using 111In-DTPA-D-Phe1-octreotide scintigraphy. AB - Two hundred and seven patients were investigated with [111In-DTPA-D-Phe1] octreotide Single Photon Emission Computerized Tomography (SPECT) scintigraphy. A comparison was carried out of the diagnostic accuracy of the three display modalities, viz. transversal SPECT (trvSPECT), three dimensional volume rendering (3Dvr) and three dimensional volume rendering plus clipping (3Dvr + c) in the rendered volume. TrvSPECT could visualize a greater number of lesions in 85 (41%) cases when compared with 3Dvr. In 48 (23%) cases, trvSPECT could visualize a greater number of lesions when compared with 3Dvr + c. The differences were caused by radioactivity in intestines and the gallbladder and in lesions shaped like an hour-glass. In conclusion, 3Dvr + c imaging was regarded as the reference since it combines both transversal and volume information. An overall sensitivity of 0.998 was found for trvSPECT and the corresponding value for 3Dvr was 0.75. The specificity was 0.45 for trvSPECT and 0.80 for 3Dvr. The accuracy was 0.87 for trvSPECT and 0.76 for 3Dvr, when compared with 3Dvr + c. The value of 3Dvr is in high-contrast images, where it provides additional anatomical information, whereas in low-contrast images, 3Dvr was found to have low accuracy. In low contrast images, trvSPECT was found to be almost equal in accuracy to 3Dvr + c imaging. However, in high-contrast images, trvSPECT was found to give more false positives than 3Dvr + c in the assessment of SPECT studies using [111In-DTPA-D Phe1]-octreotide. The interpretation was that trvSPECT has to be used in conjunction with 3Dvr + c and, in high-contrast images, 3Dvr provides additional value. PMID- 10851453 TI - Reduction in carboplatin hematopoietic toxicity in tumor bearing mice: comparative mechanisms and effects of interleukin-1 beta and corticosteroids. AB - Increasing clinical evidence suggests that treatment of certain cancers is more effective with high dose chemotherapy compared to standard dose chemotherapy. Efforts at reduction of high dose chemotherapy hematotoxicity have focused on post-chemotherapy administration of hematopoietic growth factors or stem cells. A pretreatment strategy to induce hematopoietic resistance has not been extensively examined experimentally or clinically. Pretreatment with agents can provide an alternative or additional method to reduce high-dose chemotherapy hematotoxicity. We have previously described a murine model in which normal mice were injected with high dose carboplatin (CB, 600 mg/m2, intravenously). Within 7-12 days post therapy, severe granulocytopenia and thrombocytopenia were induced and resulted in a granulocytopenic mortality of 70-80%. Utilizing this model, we observed that pretreatment of mice with interleukin (IL)-1 beta and/or a corticosteroid effectively reduced CB hematotoxicity. In the current studies, we demonstrated that C3H/HeJ mice bearing 0.25-0.5 cm RIF-1 tumors, exhibited a tumor associated decrease in hematopoietic tolerance to CB compared to normal mice. However, IL-1 beta (1 ug/mouse/day for 7 days), cortisone acetate (CA 0.5 mg/mouse/day for 7 days) or both CA and IL-1 beta, decreased CB induced mortality rates (control = 73%, IL-1 beta = 46%, CA = 30%, IL-1 beta + CA = 5%). Pretreatment with IL-1 beta, CA, or both ameliorated CB-induced absolute granulocyte, lymphocyte and platelet count nadirs. Bone marrow granulocyte-macrophage colony forming units (CFU-GM) from mice treated with IL-1 beta demonstrated increased ex-vivo resistance to cisplatin, without altering its sensitivity to high specific activity 3H-thymidine (a measure of cell fraction in S-phase). Bone marrow CFU-GM from mice treated with CA exhibited increased resistance to both cisplatin and to high specific activity 3H-thymidine. Pretreatment with IL-1 beta, CA or both did not alter tumor sensitivity to CB in vivo. These data suggest that IL-1 beta, CA or the combination may be clinically useful in reducing the hematotoxicity of CB. PMID- 10851454 TI - Influence of flavonoids on cell cycle phase as analyzed by flow-cytometry. AB - Flavonoids were previously reported to have cytostatic activity in vitro and in vivo. In the present study each phase of the cell cycle was observed by flow cytometry after HCT-15 cells had been cultured with flavonoids for 2 days. In the control group, two peaks were observed on the histogram, the first peak representing double strands of DNA, and the second, tetra strands. Chalcone, which is known to be a precursor of all flavonoids, seemed to block the passage from S to G2 and M phase. Flavonone, which is freely transformed into chalcone in vivo, showed a similar histogram as chalcone. Among the anthocyanins most effective in suppressing the tumor cell growth, cyanin and pelargonidin made a mild rise between the 1st and 2nd peak suggesting block between G1 and S phase. Delphinidin gave a high percent of cells in G1 phase, suggesting prolongation of G1 phase. When quercetin or kaempherol, both of which are flavonols were added to the culture, the second peak rose significantly, indicating a block between G2 and M phases. Thus, the site of these cytostatic agents seems to differ depending on the kind of flavonoid. PMID- 10851455 TI - Synergistic cytotoxicity between a protein kinase C inhibitor, UCN-01, and monoclonal antibody to the epidermal growth factor receptor on MDA-468 cells. AB - Cytotoxic effect of the monoclonal antibody to the epidermal growth factor receptor (anti-EGFR MAb) given alone or in combination with UCN-01, a selective protein kinase C inhibitor, was investigated in vitro. MDA-468 human breast cancer cells expressing both a large amount of EGF receptors and its ligand, transforming growth factor a, were used. A given number of the cells were plated and treated with the MAb and/or UCN-01 for 48 h. These cells were replated and incubated for colony formation assay. Cytotoxicity of the anti-EGFR MAb alone was hardly detected. However, when cells were treated with both anti-EGFR MAb and UCN 01, the combined cytotoxic effect was synergistic. PMID- 10851456 TI - Antitumor vaccination with gene-transduced tumor cells expressing a fusion protein RM4/IFN-tau. AB - Our previous studies showed that secretion of a fusion protein RM4/IFN-tau from a mouse myeloma cell line VKCK/RM4-IFN-tau curtailed its tumorigenicity. Inoculation of VKCK/RM4-IFN-tau tumor cells further induced a protective immunity against a secondary challenge of parental VKCK tumor cells, in which the predominant immune cellular components are CD8+ T cells. In this study, VKCK/RM4 IFN-tau cell line was again used to further characterize the protective immunity. We found that the reduced tumorigenicity of VKCK/RM4-IFN-tau was directly related to the amount of fusion protein secreted by tumor cells, and that CD8+ T cells derived from mice experienced with VKCK/RM4-IFN-tau tumor regression played an important role in the protective immunity in a chromium release assay in vitro and in an animal study in vivo by using T-cell subset depleted mice. Our animal studies also showed that not only the cytotoxic but also the memory T cells against the secondary challenge of parental VKCK cells could be adoptively transferred to normal BALB/c mice. In addition, our animal studies further showed that local vaccination of irradiated VKCK/RM4-IFN-tau cells was able to significantly inhibit established tumors in early stages in vivo. This study thus highlights the potential utility of this engineered VKCK/RM4-IFN-tau tumor cells secreting the fusion protein RM4/IFN-tau in cancer gene therapy. PMID- 10851457 TI - Inhibition of HeLa cell proliferation by 4-hydroxynonenal is associated with enhanced expression of the c-fos oncogene. AB - Previous studies have shown that the highly reactive aldehyde 4-hydroxynonenal (HNE), a mediator of oxidative stress, can either stimulate or inhibit cell proliferation, depending on the concentration of the aldehyde and the presence of serum. HNE can also induce differentiation of tumour cells in vitro and inhibit the tumour development in vivo. The aim of the study presented was to find out more details about the basic mechanisms by which HNE influences cell growth behaviour. Therefore we analysed the effect of HNE on the transcription of the c fos gene in HeLa cells, to clarify the pathway by which the aldehyde modulates gene transcription and growth behaviour of the cells. At a supraphysiological concentration (50 microM) the aldehyde caused an enhanced c-fos transcription (as measured by the reverse transcriptase/polymerase chain reaction assay), while it inhibited cell proliferation markedly. Therefore, we assume that among the "early" effects of HNE on cellular growth regulation might be an altered expression of the "early response" genes (c-fos), while a "late" effect might be an altered autocrine/paracrine growth regulation of the cells. This finding on the possible basic mechanisms of the biological effects of HNE together with the already described high toxicity of the aldehyde for cancer cells give support for the further evaluation of the possible use of HNE in cancer biotherapy. PMID- 10851458 TI - Regulatory hierarchies. PMID- 10851459 TI - Safety, feasibility and therapeutic activity of intrapleural autologous lymphocyte conditioned medium followed by systemic recombinant interleukin-2 and interferon-alpha in a patient with malignant pleural mesothelioma. PMID- 10851460 TI - Clinical and in vitro response to 13-cis-retinoic acid in interferon-alpha resistant renal cell carcinoma. AB - Retinoids are known to control many important biological processes, including differentiation, morphogenesis, growth and tissue homeostasis. More recently, clinical and pre-clinical results provide evidence for an antiproliferative effect of 13-cis-retinoic acid (13cRA) in interferon-alpha (IFN-alpha) treated renal cell carcinoma patients. The manner in which 13cRA augments antitumor effects and modulates biologic and clinical responses of renal cell carcinoma to IFN-alpha remains elusive. In the present study, we report induction of apoptosis and objective tumor regression in response to 13cRA in advanced renal cell carcinoma patients refractory to IFN-alpha. Among 21 patients treated there were one complete and four partial remissions (objective response rate, 24%; median response duration 8+ months). Preliminary evidence suggests that 13cRA acid may reverse IFN-alpha resistance in renal cell carcinoma. PMID- 10851461 TI - Dose distribution of low-dose subcutaneous recombinant interleukin-2 affects the secondary release of interferon-gamma in vivo. AB - Human recombinant interleukin-2 (rIL-2) induced interferon-gamma (IFN-gamma) release in vivo was studied in 16 renal cell carcinoma patients treated with low dose s.c. rIL-2. The s.c. administration of rIL-2 resulted in a significant increase in circulating IFN-gamma in all patients within 6 to 8 hours as measured by enzyme-linked immunosorbent assay (ELISA). Total IFN-gamma release, as expressed by the area under the concentration curve (AUC), and IFN-gamma serum peaks following repetitive s.c. rIL-2 injection showed a direct dose distribution dependancy, whereby significantly higher levels of secondary IFN-gamma were achieved in patients treated with 10 million IU rIL-2/m2 q 12 hours when compared with patients treated with 20 million IU rIL-2/m2 q 24 hours. IFN-gamma release was suppressed significantly in one patient who had been pretreated with corticosteroids, while prior immunotherapy with rIL-2 had no measurable effect on secondary IFN-gamma release in this study. Cumulative secondary IFN-gamma secretion, as expressed by the AUC, and IFN-gamma serum peak concentrations in response to s.c. rIL-2 did not correlate with response to therapy or survival of rIL-2 treated renal cell carcinoma patients. PMID- 10851462 TI - Active specific immunotherapy of renal cell carcinoma: cellular and humoral immune responses. AB - We investigated the effect of vaccinating renal cell carcinoma (RCC) patients with irradiated autologous or allogeneic tumor cells and Newcastle disease virus (NDV) as adjuvant on cellular and humoral antitumor immunity. By Western blot analysis, we found that vaccination induced antibody formation in 33 of 34 patients against NDV proteins but not against tumor cell related antigens. NDV proteins detected had molecular weights of 53 kDa, 55-56 kDa, and 66 kDa. ADCC by patients' isolated PBMC and patients' sera against autologous or allogeneic tumor cells was not enhanced after vaccine treatment in a nonradioactive cytotoxicity assay. Target cells infected with NDV were lysed more effectively (p < 0.05) in ADCC after vaccination than noninfected targets. Natural cellular cytotoxicity of patients' isolated PBMC was not altered during vaccine treatment. Specific lysis rates against autologous and allogeneic RCC cells not exceeded 10% (effector:target ratio 50:1). Specific lysis of K-562 cells was > 20%; a slight decrease in lysis during vaccination was not significant. Numbers of lymphocyte subsets from patients' peripheral blood analyzed by FACS revealed significant expression of CD20+ (p < 0.02) and CD39+ (p < 0.03) cell numbers by vaccine therapy. Cytokine detection in patients' sera by ELISA showed significant increases (p < 0.05) for IFN-gamma and TNF-alpha but not for IFN-alpha four h post vaccination. Thus, immunomodulation with autologous or allogeneic RCC tumor cell vaccines is mainly due to cytokine induction, whereas tumor specific humoral or cellular responses are not detectable in patients' peripheral blood. PMID- 10851463 TI - Use of retroviral vectors encoding murine inducible nitric oxide synthase gene to suppress tumorigenicity and cancer metastasis of murine melanoma. AB - The purpose of this study was to determine whether retrovirus-mediated transfer of murine macrophage inducible nitric oxide synthase (iNOS) can produce inhibition of tumorigenicity and metastasis. Retroviral vectors encoding macrophage iNOS constructed in pLXSN, a retroviral vector with the iNOS gene under the control of a long terminal repeat promoter, were stably transfected into PA317 cells. Medium harvested from confluent monolayers of the virus producing cell lines was used for infection of the murine K-1735 melanoma cells. Expression of iNOS was confirmed by northern and Western blot analyses. Functional iNOS protein expression was confirmed by bioassay of nitrite accumulation in the culture supernatant. Cells infected by a control iNOS negative retrovirus produced fast-growing subcutaneous tumors and many lung metastases in nude mice, whereas iNOS-transduced cells produced slow-growing tumors and few lung metastases, showing that the infection of murine tumor cells by retroviruses harboring the iNOS gene can suppress tumorigenicity and metastasis. PMID- 10851464 TI - Chimeric anti-CD20 (IDEC-C2B8) monoclonal antibody sensitizes a B cell lymphoma cell line to cell killing by cytotoxic drugs. AB - More than 50% of patients with aggressive B lymphomas and the majority of patients with low grade lymphomas are not cured by current therapeutic strategies. The lymphomas express the B cell antigen CD20 on the cell surface and this antigen serves as target for antibody-directed therapies. Clinical studies with encouraging results have been underway with the use of a chimeric anti-CD20 antibody (IDEC-C2B8), consisting of human IgG1-6 constant regions and variable regions from the murine monoclonal anti-CD20 antibody IDEC-2B8. This study investigated the potential anti-tumor therapeutic value of combination treatment with anti-C2B8 and cytotoxic drugs. The in vitro study examined the sensitizing effect of C2B8 antibody on the DHL-4 B lymphoma line to various cytotoxic agents. Cytotoxicity was determined by the MTT assay. Surface and cytoplasmic proteins were determined by flow cytometry. Pretreatment of DHL-4 with C2B8 resulted in inhibition of cell proliferation and cell death and a fraction of the cells underwent apoptosis. While the DHL-4 tumor cells were relatively resistant to several cytotoxic drugs, pretreatment with C2B8 rendered the cells sensitive to TNF-alpha, ricin, diphtheria toxin (DTX), adriamycin and cisplatin but not to VP 16. Chemosensitization of DHL-4 tumor cells was not due to downmodulation of either the MDR-1 or bcl-2 gene products. However, treatment of DHL-4 with C2B8 inhibited TNF-alpha secretion. These findings demonstrate that C2B8 antibody potentiates the sensitivity of DHL-4 tumor cells to several cytotoxic agents. Further, the findings suggest that combination treatments with C2B8 antibody and drugs may be of clinical benefit in the treatment of patients with resistant aggressive B lymphomas. PMID- 10851465 TI - Radioimmunodetection of neuroblastoma cells with I-131-radiolabelled antibodies. AB - The murine anti-neuroblastoma monoclonal antibodies 15/7 and 19/1/4 should be tested for specific radiolocalization of neuroblastoma by immunoscintigraphic imaging of this tumour growing in mice. Radioiodination of both antibodies was done by chloramine-T method resulted in an immunoreactivity of 75%. The calculated specific activity varied from 51.1 to 126.2 kBq/microgram IgG. In each case, about 500 kBq of labeled antibodies were intraperitoneally injected into human neuroblastoma (SK-N-MC, SK-PN-DW and IMR 5) xenografted severe complete immunodeficient (SCID) mice. Whole-body scintigraphy was performed daily by a scintiscanner to localize the tumour site. After last scanning principal organs were removed and their I-131-uptake was determined by measuring the impulse rate. The best scintigrams were done with I-131-19/1/4 at the second day after antibody injection. Radioconjugates were accumulated at highest in the tumour at the third day after application of 15/7 and 19/1/4 with a tumour uptake of 0.4 and 2.2 per cent of injected dose per gram (%ID/g), respectively. The 15/7-moAbs was accumulated approximately 9-fold higher in the SK-N-MC and SK-PN-DW grafts than in principal organs, whereas the tumour/non-tumour-ratio of the 19/1/4 moAb was 3:1. The results indicate the efficacy of these two neuroblastoma antibodies in radiolabelling and their usefulness for tumour imaging of neuroblastoma engrafted SCID-mice. PMID- 10851466 TI - Therapeutic immunostimulating effects of plant mitogens exemplified by the L4 isolectin of PHA. AB - A sizable body of evidence has accumulated showing that the characteristics of certain plant mitogens should endow them with valuable immunomodulating effects. Immune stimulation would be the fundamental function operating through the broad consequences of nonspecific T cell activation following binding with their CD3 or CD2 molecules. Based on in vitro evidences that PHA, operating through the LDCC pathway, might kill any tumor target if it remains present in adequate concentration, the administration of mitogens for cancer therapy would be rational, and the same mechanism should also justify these agents for treatment of certain infections. Being nonerythroagglutinating, although leukoagglutinating in higher concentrations, PHA-L4 serves as a suitable model for immunostimulating activities of the mitogens that can be applied directly or as in vitro activators of adoptive leukocytes. The PHA skin test can be utilized to gauge the serum levels of inhibitory glycoproteins that become elevated in a number of the disorders treatable by the mitogen, thus facilitating necessary dosage modifications. While PHA is the only mitogenic lectin that has been tested clinically, Con A and PWM are the two most widely studied among the alternatives, with others also available and many undoubtedly awaiting discovery. The development of practical methods for producing industrial quantities of nonagglutinating mitogens in pure form should be the goal, and accomplishing the latter might also answer certain hypersensitivity concerns. PMID- 10851467 TI - Theoretical benefits of mitogen applications for HIV-1 infections. AB - Ideal treatment of HIV-1 infections should include an agent that can reverse the capacity of the virus to evade destruction by hiding in sanctuaries and by frequently mutating the epitopes it displays. The rapid proliferation of virions during the years of symptomatic quiescence obligates rapid replacement of CD4+ lymphocytes that leads to a gradual attrition of the T lymphocytes needed to control infections. In vitro evidences suggest that, given systematically, certain mitogenic lectins would interfere with HIV-1 invasion of CD4+ cells by blocking gp120 molecules on the viral membrane before activating T lymphocytes subsequent to binding with their Ti/CD3 molecules. The nonspecific nature of antiviral effector cells generated by this activation should circumvent HIV-1 mutations at the same time it reconstitutes depleted T lymphocytes, stimulates myelopoiesis, and reinforces resistance to malignancies and infections prevalent with the immunodeficiency state. Properly coordinating these effects with appropriate combinations of reverse transcriptase and protease inhibitors could theoretically expedite complete elimination of HIV in a timely fashion that shorten the required treatment duration and excludes the detrimental effects of virus mutations. The proper sequence of this treatment should be maximum reduction of the HIV-1 load with drug combinations, control of complicating infection by other means to reduce mitogen-induced tissue necrosis, and addition of systemic PHA-L4 administration regulated to maintain a 5-10 micrograms/mL serum concentration. The antiviral regimen should be continued an undetermined time beyond when HIV-1 is no longer detectable, and systemic L4 administration until satisfactory immunologic and hematologic competences are re-established. Partially-matched mitogen-activated adoptive leukocyte therapy might be additionally helpful. PMID- 10851468 TI - Magic bullets at last! Finally--approval of a monoclonal antibody for the treatment of cancer!!! PMID- 10851469 TI - A new technique for the 21st century: outpatient, high dose, tumor selective, irradiation by infusion. PMID- 10851470 TI - The Cancer Biotherapy Research Group [CBRG], formerly the National Biotherapy Study Group [NBSG]: a 10-year perspective. AB - The Cancer Biotherapy Research Group [CBRG], formerly known as the National Biotherapy Study Group [NBSG], celebrated its 10th anniversary in 1997. CBRG is a not-for-profit cancer clinical trials group of community oncologists whose activities for the most part have been funded by their associated community hospitals. From its inception the Group has focused on clinical investigations of biological agents in the treatment of patients with advanced cancer. This article reviews the history of the Group, its structure, accomplishments, and its current objectives and challenges. PMID- 10851471 TI - Long-term survival after continuous infusion interleukin-2. AB - PURPOSE: Between 1987-1990, 612 patients received high-dose continuous intravenous interleukin-2 (IL-2) in phase II clinical trials of the National Biotherapy Study Group (NBSG). The purpose of this analysis was to determine the long-term survival rates associated with such therapy and the correlation, if any, between objective tumor response, and survival. METHODS: Patients who are known to have survived at least 3 years or more were identified. Actual and actuarial survival rates were determined for various malignancies and by tumor response. RESULTS: At least 37 (6.0%) survived > or = 3 years from the initiation of IL-2 therapy, and it is possible the 3-year survival rate is as high as 20%. This included 14/168 (8%) of patients with renal cell carcinoma, 10/175 (6%) melanoma, 2/51 (4%) lung cancer, 0/61 colorectal, 0/36 breast, 1/17 sarcoma, 1/14 pancreas, 1/19 ovary, 3/11 lymphoma, 2 adenocystic carcinomas, 2 carcinoid, and one Hodgkin's disease. Four hundred ninety-one of 612 (80%) are known to have died, 121 were still alive at the time of the last follow-up. Median survival was 7.9 months. Among 547 evaluable patients, there were eight complete responses (CR) and 42 partial responses (PR), for an objective response rate of 9%. An additional 32 patients had mixed or minimal responses (MR) for a total response rate of 15%. The 3-year survival rates were 25% for PR, 17% for PR, 16% for MR, 8% for stable disease (SD), and 2% for patients with progressive disease (PD). Responders made up a higher proportion of patients who survived > or = 3 years than of patients who survived < or = 3 years (14/37 = 38% vs 68/500 = 14%; p < .0001, X2). A higher proportion of responders survived > or = 3 years than non responders (14/82 = 17% vs 23/230 = 4%, p < .0001, X2). Patients with CR, PR, or MR had a > or = 3-year survival rate (14/82 = 17%) than patients who had SD (20/249 = 8%; p = .02, x2, who in turn had a > or = 3-year survival rate that was greater than patients who had PD (3/206 = 2%; p = .001, X2). For individual trials in which 10 or more patients were enrolled, the percentage of patients surviving > or = 3 years ranged between 4% and 8%. CONCLUSION: We conclude that in the setting of IL-2 therapy for metastatic cancer, the probability of surviving > or = 3 years was approximately 12 times greater for responders, and five times greater for patients with SD; as compared to patients who had PD. Furthermore, despite diagnoses of metastatic cancer, often in settings in which disease had been refractory to standard therapy there was an actual 3-year survival rate of at least 6% to 8% for patients with metastatic melanoma and renal cell carcinoma. PMID- 10851472 TI - Hybrid high-dose bolus/continuous infusion interleukin-2 in patients with metastatic melanoma: a phase II trial of the Cancer Biotherapy Research Group (formerly the National Biotherapy Study Group). AB - PURPOSE: Interleukin-2 (IL-2) is an active agent for the treatment of melanoma. In animal studies, polyethylene glycol conjugated (PEG) IL-2 was found to be effective in certain IL-2-resistant models. Bolus/infusional IL-2 administered to approximate the pharmacokinetics of PEG-IL-2 also overcame resistance in these models. Based on these observations, the Cancer Biotherapy Research Group (CBRG) [formerly the National Biotherapy Study Group (NBSG)] previously had conducted a pilot study and then a phase I trial of bolus IL-2 followed by continuous IL-2 (NBSG 90-01). METHODS: In the current study, NBSG 92-09, a phase II trial was conducted using IL-2 at a dose of 36 MIU/m2 followed by a 72-hour continuous infusion of IL-2 at 18 MIU/m2/day, so that over 3 days a total of 90 MIU/m2 of IL 2 were delivered; the same amount as previously given during 5 days of continuous i.v. IL-2 at 18 MIU/m2/day. This schedule was repeated every 2 weeks for 2 months, and then monthly for up to 6 months. RESULTS: Twenty-two patients with metastatic melanoma were enrolled in this trial. Toxicities were qualitatively similar to those seen with other IL-2 regimens, but grade 3 and 4 toxicities were observed only in patients who received at least four cycles of treatment; only one patient went off study because of toxicity. For 18 patients with measurable disease, there were two complete and two partial responses in patients ages 32, 66, 72 and 83 years, for a response rate of 22% (6% to 48%; 95% confidence interval [Ci]). The median survival for all 21 evaluable patients enrolled in the trial was 8.5 months. CONCLUSION: The hybrid schedule of drug delivery in NBSG 92 09 allowed the same dose and intensity of IL-2 to be delivered over 3 days instead of 5 days, which resulted in fewer days of hospitalization and therefore decreased cost; but with no increase in toxicity and no decrease in efficacy in patients with metastatic melanoma. PMID- 10851473 TI - The sCEA molecule suppressive role in NK and TH1 cell functions in colorectal cancer. AB - The soluble form of carcinoembryonic antigen (sCEA), an oncofetal glycoprotein, is frequently produced by human epithelial-tumor cells, particularly of colorectal origin, and evaluated as a prognostic index of tumor progression and patient survival. sCEA molecules are often present at high concentrations in the peripheral blood of colorectal cancer patients, but the function and significance of this are not well understood. Reported data have demonstrated that sCEA can interfere in NK-cell/tumor-cell interaction by drastically reducing the lysis of tumor cells in a dose-dependent manner and can also suppress T and B cell functions. The aim of our study was to evaluate this situation in colorectal cancer by determining peripheral blood immunological parameters in a group of patients and healthy subjects. We evaluated the interleukin (IL)-2, interferon (IFN) gamma, IL-4, sIL-2R and IL-10 levels in the serum and the release of IFN gamma, IL-4 and IL-10 from peripheral blood mononuclear cells (PBMC); the PBMC expression of CD3, CD16 and CD19 phenotypic antigens; the PBMC proliferative responses to IL-2, IL-2 + anti-CD3 monoclonal antibody (mCD3) and mCD3. The statistical evaluation of our overall results strongly indicates that the high level of the sCEA molecules in the patient's serum might act as a suppressive factor for NK and TH1 immunocompetent cells. This may be the cause of sCEA involvement in tumor progression, and indicates the possibility of an improvement in cancer treatment through its manipulation. PMID- 10851474 TI - IL-10 and sIL-2R serum levels as possible peripheral blood prognostic markers in the passage from adenoma to colorectal cancer. AB - The pathogenesis of cancer is currently under intense investigation to identify reliable prognostic indices for the early detection of disease. Adenomas have been identified as precursors of colorectal cancer and tumor establishment, and disease progression has been found to reflect a malfunction of the immune system. We previously indicated the investigation of cytokine serum levels in these patients as a useful and non-invasive tool for the study of the disease progression and an imbalance at TH1 and TH2 cell levels was also found. Moreover, the soluble form of interleukin (IL) 2 receptor (sIL-2R) level is an in vivo marker of T cell activation and is used to monitor the activation of the immune system. We therefore performed an immunological study on a group of healthy subjects, subjects with adenomas, and colorectal cancer patients to identify peripheral blood invasiveness markers in the progression from normal mucosa through adenoma to tumor. In this paper we evaluated the relationships between serum levels of interleukin IL-2, sIL-2R, interferon (IFN) gamma, IL-4, IL-6, IL 10 and sICAM-1 and their networks. Our overall data indicate that in the normal mucosa through adenoma to tumor progression, the host immune response proceeded from a TH1 cell-mediated immune response type (healthy subjects) to a type with TH2 suppressive characteristics (adenoma subjects and cancer patients). However, in the adenoma subjects there was no IL-10 or sIL-2R involvement, while these parameters were implicated in the cancer patients' immune responses. Moreover, a concurrent augmentation of sIL-2R and IL-10 levels seems to be prognostic for the passage from adenoma to cancer, and the sIL-2R and sICAM-1 molecules appear to be involved in the invasiveness mechanisms. PMID- 10851475 TI - Effect of allomelanin on tumor growth suppression in vivo and on the cell cycle phase. AB - Allomelanins are nitrogen-free macromolecular polymers of simple phenols produced by higher plants and fungi. In an earlier work we found that allomelanins were effective in suppressing the growth of cultured tumor cells. In the present study we examined the effect of these polymers on the survival curve of Balb/C mice inoculated i.p. with Meth/A cells (3 x 10(4) cells/mouse), which originated from a malignant lymphoma. Allomelanins were extracted from black sesame seeds and black soybeans, and given p.o. via the drinking water. The dose of allomelanins was approximately 3 mg/mouse/day. The percent of survivals was significantly higher in the experimental groups than in the control. Also, when HCT-15 cells were cultured for 2 days in medium containing 400 micrograms/ml of protein-free allomelanins, histograms obtained from flow-cytometry data showed a significant increase in the fraction between the diploid and tetraploid peaks, indicating blockage of the S phase of the cell cycle. These results suggest that allomelanins suppressed the tumor cell growth in vitro and in vivo, and that the effect was cytostatic, mostly by blockage of the S phase. PMID- 10851476 TI - Antitumor effect of anthocyanin fractions extracted from red soybeans and red beans in vitro and in vivo. AB - Many bioflavonoids extracted from petals of higher plants and from fruit rinds, as well as purified flavonoids, have been reported to have antitumor effects in vitro and in vivo. Bioflavonoids extracted from red soybeans are mostly cyanin conjugated with glucose and rhamnose, whereas bioflavonoids of red beans are cyanin conjugated with rhamnose as revealed by thin-layer chromatogram. Flavonoids extracted from red soybeans were effective in inhibiting the growth of HCT-15 cells in vitro. On the other hand, flavonoids from red beans were not effective, although their hydrolyzed sugar-free forms were growth inhibitory. Sugar-bonded bioflavonoids extracted from both red soybeans and red beans were effective in prolonging the survival of Balb/C mice bearing syngeneic tumor Meth/A cells, when they were dissolved in drinking water and given at a dose of approximately 500 micrograms/mouse/day. PMID- 10851477 TI - Theoretical and literary evidence for the existence of the passive antitumor defence system. AB - It is well documented that despite global abnormalities of the immune system in AIDS and other immune deficiency diseases or in immunosuppressed patients the incidence of only a few kinds of tumour increases and even in the development of tumours in question the degree of immunosuppression seems not to be a critical factor. It results from this that the known immune system has no significant role in the mechanism that prevents the development of tumours. Consequently, the fact that tumours do not develop in the majority of the population during their lifetime, indicates the existence of other defence systems. We assumed that the defence is made by small substances of the circulatory system. Substantial evidence exists that the uptake of the majority of these substances is increased and unregulated by tumour cells and proportional to their availability. This feature of tumour cells may be fatal when the number of cells is still low and there are abundant substances in their environment because some substances may be toxic if their concentrations can reach high level in the cells. Thus, the arising cancer cells die in the majority of the population during their lifetime if the number of cells arisen is not too high, or the concentrations of the required substances are not too low. To our hypothesis, the above effects of the physiological mixture of the given molecules in the blood form the Passive Antitumor Defence System (PADS). This hypothesis is confirmed by our experiments and supported by epidemiological, clinical observations and other literary data. PMID- 10851478 TI - The intraoperative detection of ovarian adenocarcinoma using radiolabeled CC49 monoclonal antibody and a hand-held gamma-detecting probe. AB - A phase II study to evaluate the safety and efficacy of the 125I-radiolabeled anti-TAG-72 monoclonal antibody, CC49, as a component of a system for the intraoperative detection of occult ovarian cancer deposits was carried out at the University of Nebraska Medical Center. Patients entered into the study were to have surgery for evaluation of their disease status. The primary objective of this study was to determine the ability of a gamma-detecting probe (GDP), the Neoprobe 1000, to intraopertively localize sites of disease not identified by traditional surgical or radiographic evaluation. It was postulated that improved detection of cancer foci might allow for therapeutic excision or might result in an alteration of subsequent treatment. Ten patients were enrolled in the study between May 1993 and March 1994. Nine of the patients were undergoing second-look surgery after completing primary chemotherapy. The remaining patient was having surgery to assess possible cancer recurrence. All patients received an intravenous injection of 2 mCi/1 mg 125I-radiolabeled CC49 without complication. After a mean of 24.5 days, the patients' background radiation counts were deemed low enough for accurate intraoperative cancer localization, and surgery was performed. Any visibly or palpably abnormal areas were biopsied after being evaluated with the GDP. Any areas suspicious for malignancy by GDP evaluation were also biopsied. Two patients without evident disease by radiographic or surgical examination had histologically confirmed metastases localized by the GDP. Four patients had obvious disease at surgery which was variably confirmed by the GDP; two of these patients had baseline elevations in circulating TAG-72 antigen levels that may have affected binding of antibody to the tumor. This system of radioimmunoguided surgery was well tolerated and practical in its application, and it permitted disease detection that resulted in potentially beneficial changes in patient management. PMID- 10851479 TI - Cancer biotherapy: when, why and how long? PMID- 10851480 TI - The role of the soluble CD30 serum level in colorectal cancer: a possible marker for a patient subset which could benefit from IL-2 biotherapy. AB - On the basis of our previous data suggesting an impairment in host immune response in colorectal cancer caused by an inappropriate switch from TH1 towards TH2 cells, we investigated the role of the soluble CD30 (sCD30) in this disease, as this molecule was found related to immune responses characterized by the activation of a prevalence of TH2 cells. We studied a group of healthy subjects and colorectal cancer patients determining the sCD30 serum level and the following immunological parameters: s interleukin-2 receptor (sIL-2R), IL-2, interferon (IFN) gamma, IL-6, IL-4 and IL-10 levels in the serum and peripheral blood mononuclear cell (PBMC) production; PBMC proliferative responses to IL-2, anti-CD3 monoclonal antibody (CD3) and IL-2 + CD3. Our overall data indicate that in colorectal cancer the sCD30 serum level is also linked to a prevalence of the TH2 immune response activation. However, the Multivariate statistical study underlines that the sCD30 level is principally related to the IL-6 TH2 cytokine. Moreover, it suggests that in colorectal cancer, the sCD30 level might be a marker for identifying a patient subset in which IL-2 biotherapy treatment could contribute to the restoration of the impaired immune system. PMID- 10851481 TI - A CDR-grafted (humanized) domain-deleted antitumor antibody. AB - While several murine monoclonal antibodies (MAbs) directed against carcinoma associated antigens have shown excellent tumor targeting properties in clinical trials, the use of radiolabeled MAbs for both diagnostic and therapeutic applications has been hindered by two factors: (a) the induction of host anti immunoglobulin (Ig) responses and (b) slow plasma clearance of unbound radiolabeled MAb, resulting in bone marrow toxicity for therapeutic application, and long intervals between MAb administration and tumor detection for diagnostic applications. This report describes the development of the first recombinant Ig with properties designed to reduce or eliminate both of the above problems: a complementarity determining region (CDR)-grafted humanized (Hu) MAb with a CH2 domain deletion (delta CH2). The MAb chosen for engineering was CC49, which is directed against a pancarcinoma antigen designated TAG-72 that is expressed on the majority of colorectal, gastric, breast, ovarian, prostate, pancreatic and lung carcinomas. When characterized for antigen binding in solid phase competition radioimmunoassays, the HuCC49 delta CH2 MAb completely inhibited the binding of murine (mu) CC49 and HuCC49 for TAG-72. The relative affinity constants (Ka) of MAbs HuCC49 delta CH2, HuCC49 and muCC49 were 5.1 x 10(-9), 2.1 x 10(-9) and 2.3 x 10(-9), respectively. The plasma clearance of 131I-HuCC49 delta CH2 was significantly faster than that of intact 125I-HuCC49 after either i.v. or i.p. administration in athymic mice (p(2)0.05). Biodistribution studies in athymic mice bearing human colon carcinoma xenografts after i.v. or i.p. administration of 131I-HuCC49 delta CH2 and 125I-HuCC49 demonstrated the efficient tumor localization and substantially lower percent of the injected dose (%ID/g) of the HuCC49 delta CH2 in normal tissues. This is reflected in the significantly higher radiolocalization indices (%ID/g in tumor divided by %ID/g in normal tissue) observed with the HuCC49 delta CH2 for most normal tissues tested (p(2)0.05). The differential between the rate of plasma clearance of HuCC49 delta CH2 and HuCC49 was even more pronounced in SCID mice, which have been shown to be an appropriate model to study the metabolism of human IgG. These studies thus describe the development of a recombinant Ig molecule which, for the first time, combines 1) the properties of more rapid blood clearance than an intact humanized Ig molecule--without loss of antigen binding affinity--and 2) reduced potential for eliciting a human anti-murine antibody (HAMA) response in patients. These studies also demonstrate the potential utility of HuCC49 delta CH2 for i.p. as well as i.v. radioimmunodiagnosis and radioimmunotherapy in patients with TAG-72 positive tumors. PMID- 10851482 TI - Culture medium induced morphological changes of melanoma cells associated with change in sensitivity to lysis by lymphokine-activated killer cells. AB - Three sublines (Clones 1, 2 and 7) of the human melanoma CaCL 73-36 cell line with different cellular morphology, growth patterns, melanin content and surface antigenic profile were maintained in RPMI-1640 medium plus 10% fetal bovine serum (abbreviated as RPMI). Each subline was divided into two groups: one grown in RPMI and the other in Dulbecco's modified Eagle's medium plus 10% fetal bovine serum (abbreviated as DMEM) for 96 h. Phenotypically, Clone 2 expressed Class I and II MHC and ICAM-1 on the surface and in the cytoplasm, while Clones 1 and 7 failed to express these antigens in both the cytoplasm and on the cell surface. Melanotic Clones 1 and 7 cells became even more pigmented, had slower growth rates, and exhibited lower saturation densities when incubated in DMEM than when they were incubated in RPMI. On the other hand, Clone 2 cells maintained in RPMI were grossly amelanotic, contained defective-like melanosomes detected ultrastructurally, and had distinct clusters of microvilli polarly located in most of the cells. Such specialized ultrastructures were not affected by medium conditions. Analysis of sensitivity of the clonal sublines to cytolysis by allogeneic effector cells revealed that in spite of low levels of natural killer (NK) cytotoxicity noted, DMEM produced a 2- to 14-fold increase in sensitivity to NK cells, irrespective of which medium was used. Different levels of lymphokine activated killer (LAK) cytolytic activity were clearly observed in sublines maintained in RPMI, with Clone 2 being the most sensitive and both Clones 1 and 7 being less sensitive. Cells grown in DMEM exhibited significantly higher levels of sensitivity to LAK cytolysis than cells grown in RPMI as revealed by their differences in lytic units (p < 0.05). This was likely due to the high levels of surface ICAM-1 expression in cells incubated in DMEM vs little expression of this adhesion molecule by cells grown in RPMI. Taken together, these results demonstrate the presence of heterogeneous subpopulations within the CaCL 73-36 melanoma cell line regarding their pigmentary status, antigenic profile, growth pattern and responsiveness to NK/LAK cytolysis. The results also call attention to the importance of utilizing a same medium in short- and long-term cultures of melanomas for biological studies and response evaluations of therapeutic agents such as LAK cells, when multiple cell targets from different patients or multi metastatic cell lines from individual patients are to be compared. Finally, these melanoma sublines may be valuable for further elucidation of the relationship between MHC expression, and increased sensitivity to LAK cytolysis, and the role of the components of DMEM in the mechanism for the observed induction of cell differentiation and enhanced LAK cytolysis. PMID- 10851483 TI - Expression of inflammatory cytokines by murine macrophages activated with a new synthetic lipopeptide JT3002. AB - The present study was undertaken to investigate the effect of JT3002, a new synthetic analogue of a lipoprotein from the outer wall of a gram-negative bacterium on the production of cytokines by mouse peritoneal macrophages. Multilamellar liposomes containing different concentrations of JT3002 induced production of the inflammatory cytokines tumor necrosis factor-alpha, interleukin 1 alpha, and interleukin-6 by macrophages in dose- and time-dependent manners. The presence of interferon-gamma enhanced production of tumor necrosis factor alpha by macrophages exposed to lower concentrations of JT3002 and induced the release of nitric oxide, a potent cytolytic molecule of activated macrophages. Unlike lipopolysaccharide, JT3002 activated macrophages independently of serum, but like lipopolysaccharide, it required protein tyrosine kinase. PMID- 10851484 TI - Direct tumor growth suppressive effect of melanoidin extracted from immunomodulator-PSK. AB - Melanoidin, which belongs to the melanin group of molecules, was extracted from the polysaccharide biological response modifier PSK. Melanoidin was cultured together with HCT-15 cells derived from human colon cancer and with AGS cells derived from human gastric carcinoma. After four days of culture, cell count was compared with that of the control cells. Significant suppression was observed, that is, 50% suppression was shown at concentrations of melanoidin between 200 and 100 micrograms/ml. A histogram generated by flow cytometry showed elevation of the tetraploid peak and of that between diploid and tetraploid peaks, suggesting blockage of S phase and G2 to M phase of the cell cycle. Thus, melanoidins contained in the immunomodulator PSK revealed to have a direct tumor cell growth inhibitory effect. PMID- 10851485 TI - Pilot ascending dose tolerance study of parenterally administered 4-(2 hydroxyethyl)-1-piperazine ethane sulfonic acid (TVZ-7) in dogs. AB - We report a pilot toxicity study in healthy beagle dogs which revealed no significant adverse events for TVZ-7 given at i.v. doses up to 520 mg/kg/day. All treated dogs displayed calm behavior and maintained normal clinical laboratory values throughout the study. Increased bone marrow hypercellularity and extramedullary hematopoiesis was also noted in these dogs. Three other dogs with advanced spontaneously occurring tumors have also been dosed. No serious adverse events were reported in any of the dogs. This pilot study suggests that 4-(2 Hydroxyethyl)-1-piperazine ethane sulfonic acid may be used safely in planned human clinical trials. PMID- 10851486 TI - Preliminary evaluation of a fixed dose of zwitterionic piperazine (TVZ-7) in clinical cancer. AB - One of the zwitterion buffers that has shown significant therapeutic value in the treatment of pain due to cancer, immunologically mediated diseases, and the pain associated with these conditions is in the class of N-substituted amino-sulfonic acids known as "Good Buffers." Zwitterion molecules have neither a negative nor a positive charge; thus, they are neutral. 4-(2 Hydroxyethyl)-1-piperazine ethane sulfonic acid has been used for several decades in artificial biological systems (tissue culture) as a buffer. We have been exploring the therapeutic value of these zwitterionic buffers. Pilot animal studies have demonstrated that zwitterionic piperazine increases bone marrow hypercellularity and induces extramedullary hematopoiesis. We report the initial human use to explore dose toxic and physiologic effects of a fixed dose of the zwitterionic piperazine molecule. There appears to be potential therapeutic value in the treatment of pain due to cancer, and there are preliminary indications that tumor activity and tumor size are reduced. Immunologically mediated diseases may also be affected. Toxicity is low and there appear to be minimal side effects. PMID- 10851487 TI - Modeling of internal dose distributions during SR-89 treatment of a patient with bone metastases. AB - METHODS: A model of strontium biodistribution similar to the one created by the International Commission on Radiological Protection (ICRP) was applied for activity and absorbed dose calculations in a patient with bone metastases treated with Sr-89 strontium chloride. Metastases are represented just like all other organs and tissues collecting strontium. Data from the ICRP's standard Reference Man were used. RESULTS: Results include calculated time-activity data for all model compartments and for relevant target organs. Absorbed doses per unit administered activity were calculated for blood (0.036 cGy/MBq), soft tissues (0.046 cGy/MBq), bone marrow (1.15 cGy/MBq), bone surface (1.45 cGy/MBq), upper large intestine (ULI) (0.13 cGy/MBq), lower large intestine (LLI) (0.38 cGy/MBq), bladder (0.12 cGy/MBq), and metastases (37.5 cGy/MBq). CONCLUSIONS: Results of the absorbed dose calculations are comparable with results presented in references for specific clinical cases. Discrepancies in dose values may be effected by the size of metastases and the patient's condition. PMID- 10851488 TI - Interleukin-2: dose, schedule and route PMID- 10851489 TI - Phase I-II study of gamma interferon and 5-fluorouracil for patients with metastatic renal cell carcinoma AB - We have completed a phase I/II trial to evaluate the toxicity and efficacy of the combination of gamma interferon and 5-fluorouracil in metastatic renal cell carcinoma. Gamma interferon was administered at a weekly dose of 100 micrograms. 5-fluorouracil was given as a 5 day continuous infusion days 1-5 of each 21 day cycle. In the phase I portion of the trial, the gamma interferon dose was held constant, while the 5-fluorouracil was escalated from 500 to 800 mg/m2/day. Serum neopterin and beta 2 microglobulin were measured prior to and 48 hours after each dose of gamma interferon for the first six weeks of treatment. Dose limiting toxicity was not encountered in the phase I part of the trial; therefore the phase II study was initiated at the 800 mg/m2/day dose of 5-fluorouracil. No responses were seen among 34 patients treated on the phase II trial. Forty-six percent of patients experienced disease stabilization and the remainder progressed through treatment. In the phase I trial, increments in neopterin and beta 2 microglobulin levels differed significantly between patients treated with lower and higher doses of 5-fluorouracil. We conclude that the addition of 5 fluorouracil to gamma interferon does not appear to enhance the cytokines clinical activity. Incremental increases in macrophage activation markers with escalating 5-fluorouracil doses suggests a role for 5-fluorouracil beyond its usual proposed cytotoxic activity and warrants further investigation into potential immunologic effects of this drug. PMID- 10851490 TI - Neoadjuvant chemoimmunotherapy with cisplatin and low-dose interleukin-2 for locally advanced esophageal carcinoma AB - After a single dose of cisplatin, the ability of peripheral blood mononuclear cells (PBMC) to generate lymphokine-activated killer (LAK) cells was significantly augmented in cancer patients. Based on this clinical finding, the patients with locally advanced esophageal carcinoma were thus treated with a combination of cisplatin and low-dose interleukin-2 (IL-2) in a neoadjuvant setting. Four patients with squamous cell carcinoma of the esophagus (T3 or T4 disease) were preoperatively treated with a regimen consisting of 50 mg/m2 cisplatin on day 1, followed by IL-2 from day 4 through day 8, when the ability of PBMC to generate LAK cells had been shown to be significantly augmented. After two to four courses of the preoperative therapy, one patient achieved a histologic CR, one showed PR and one MR. No severe toxicity was encountered. All patients thereafter underwent surgery. The median survival of these patients was 47.5 months and three of the patients had been disease free for 43 to 62 months after the initiation of the therapy. The combination of cisplatin and low-dose IL 2 administered in a neoadjuvant setting seems to result in an improved survival of locally advanced squamous cell carcinoma of the esophagus. PMID- 10851491 TI - Synthesis, rhenium-188 labeling and biodistribution studies of a phenolic ester derivative of trisuccin AB - Our previous results indicated that the trihydroxamate ligand, trisuccin, was a promising bifunctional chelating agent (BCA) for radiometal labeling of monoclonal antibodies with rhenium and technetium. An interest was developed to evaluate structural modifications of this compound from both synthetic and biological points of view. In this report we describe the synthesis of an esterified trisuccin (referred to as trisester), and conjugation of this new derivative to MAb CC49, radiolabeling of this conjugate with rhenium-188 (188Re), and biodistribution of the labeled conjugate in athymic nude mice. Thus, trisuccin (1) was esterified with benzyl 4-hydroxybenzoate in a DCC/DMAP reaction followed by removal of all benzyl protecting groups with catalytic hydrogenation. The resulting product was conjugated to CC49 by the active ester technique, through formation of the 2-nitrophenyl ester 6, and the conjugate was radiolabeled with generator-produced 188Re. The lead molecule trisuccin 1 was also conjugated to CC49 through the active ester 5 and the conjugate was radiolabeled by the same procedure to serve as the control conjugate. Biodistributions of the labeled conjugates were studied in athymic nude mice, transplanted s.c. with LS174T human colon cancer xenografts. Although an increase in the radiolabeling yield was observed for the esterified ligand-CC49 conjugate, as well as some increase in its immunoreactivity, as compared to those for the parent trisuccin molecule, there were no significant differences in their biodistribution. This new compound therefore may be useful in improving the conjugation and radiolabeling chemistries of this trihydroxamate ligand system. PMID- 10851492 TI - Immunological implications of alterations in the c-Ki-ras and p53 genes in the stepwise progression of colorectal cancer: indications for the improvement of prognosis, biotherapy treatment and tumor biology understanding AB - Alterations in gene structure and functions involving the c-Ki-ras and p53 genes have been shown to play an important role in the various stages of human colorectal carcinogenesis. However, how these gene alterations cooperate with tumoral mechanisms at an immunological level is not known. To this aim an immunological study of a group of healthy subjects, patients with p53 gene deletions (53D), with c-Ki-ras mutations (KrM) and no gene alterations (53D-KrM-) have made. In a previous study we found that a disregulation between TH1/Th2 cell functions seems to be implicated in the establishment and progression of colorectal cancer disease and that soluble interleukin (IL)-2Receptor (sIL-2R) serum level is involved in this. On this basis we investigated the immunological implications of p53 and c.Ki-ras gene alterations, evaluating the relationhips in the immune network between sIL-2R levels in the serum and immunological parameters (IL-2, IL-4 serum levels; CD3, CD16 and CD19 expression on the surface of peripheral blood mononuclear cells--PBMC). Our results suggest that, in the stepwise progression of colorectal cancer, the c-Ki-ras gene alteration is involved in a switch of the host immune response to a suppressive type which, as we have previously reported, may be a determining or concurrent cause of malignant transformation. Alteration in the p53 gene does not appear to ulteriorly impair the patients' immunological response. Our data supports the role of c-Ki-ras gene mutations and p53 deletions as prognostic markers in the passage of normal tissue to adenoma and adenoma to carcinoma respectively. Moreover, the evaluation of the mechanisms involved in the alterations of c-Ki ras gene seems to be more important than that of p53 suppressor gene for the improvement of prevention, biotherapy treatment and tumor biology understanding. PMID- 10851493 TI - E4, a new monoclonal antibody identifying a human prostatic cell surface antigen AB - The monoclonal antibody E4 (IgG2a, kappa) was raised by immunizing mice with dispersed cells obtained from human benign prostatic hyperplasia (BPH). The antibody identifies an antigen abundantly expressed in normal prostate epithelial cells, in benign epithelial prostatic cells and in well- and moderately well differentiated adenocarcinomas of the prostate, whereas poorly differentiated prostatic adenocarcinomas display somewhat less expression. Investigation of the human prostatic adenocarcinoma cell line DU 145 revealed E4 immunoreactivity localized to the cell surface. SDS-PAGE analysis under reducing conditions demonstrated an approximate molecular weight of 70,000 for the antigen. The highly specific reactivity with prostate tissue, as well as intense surface staining, especially in well- and moderately well differentiated prostatic adenocarcinomas, makes the E4 antibody a useful immunohistochemical marker and a possible candidate for future immunoscintigraphy and/or targeted radiotherapy. PMID- 10851494 TI - Tumor cell growth-inhibiting effect of melanoidins extracted from miso and soy sauce AB - Melanoidins, a group of plant-produced melanins, were extracted from miso and shoyu (soy sauce), which are products of soybeans and widely consumed as food in oriental countries. The molecular weight of these melanoidins was approximately 5600 as revealed by Sepharose CL-4B column chromatography. The melanoidins were cultured together with HCT-15 cells derived from human colon carcinoma and AGS cells derived from human gastric carcinoma. Significant cell growth suppression was observed by incubating the cells with these melanoidins. The 50% growth suppression dose ranged between 100 micrograms/ml and 25 micrograms/ml, that is, between 2 x 10(-5) mM/ml and 0.5 x 10(-5) mM/ml. Flow cytometric analysis suggested that these melanoidins blocked the S phase and G2 to M phase of the cell cycle. PMID- 10851495 TI - Augmentation of cisplatin cytotoxicity using cytokines on cervical carcinoma cell lines. AB - We investigated whether the biological response modifiers like IFN-alpha, IFN gamma and TNF-alpha could enhance the cytotoxic action of cisplatin on cervical carcinoma cell lines in vitro. The sensitivity of three cell lines SiHa, ME 180 and C33A to these agents was tested using colorimetric MTT assay as well as tritiated thymidine uptake. All the three cell lines demonstrated range of sensitivity to cisplatin and cytokines. Interferons and TNF when used in combination with lower dose of cisplatin showed a significant enhancement of cytotoxic action of the drug in all the cell lines. Thus these data indicate that cytokines in concert with the drug may have a potential to improve the 'in vivo' therapy in these patients. PMID- 10851496 TI - Prognostic significance of immunological evaluation in colorectal cancer. AB - According to the concept that tumour establishment and progression generally reflects a malfunction of the immune system, we have investigated the prognostic significance of immunological parameters in correlation to stage progression in colorectal cancer. In patients and healthy subjects as control group, we determined: serum levels of interleukin (IL)-2, interferon (IFN) gamma, IL-4, IL 6, IL-7, IL-8, tumor necrosis factor (TNF) alpha cytokines and soluble IL-2 receptor (sIL-2R), CD30 (sCD30), ICAM-1 (sICAM-1) molecules, phenotype of peripheral blood mononuclear cells (PBMC); PBMC proliferative response to IL-2, IL-4 and anti-CD3 monoclonal antibody (anti-CD3) variously combined. Our results show that, compared to healthy controls, the group of all patients, but interestingly, also the groups of patients at the various stages of the disease, seem to have different values of these immunological parameters. Since tumour invasion and metastasis are the major causes of cancer treatment failure the early recognition of preinvasive states could lead to an improvement in prognosis. For this purpose our results might be especially useful in making prognostic and diagnostic indices in this neoplasy to identify patients at risk for tumour detention and the patient condition concerning disease progression by a non-invasive method. Moreover, this evaluation which contributes to identify the damage in the patient immune response to tumor could be helpful in identifying the therapeutic substances which might switch this response from being unproductive to productive. Thus, our data leads us to indicate that it might be possible to define reliable prognostic and diagnostic indices in colorectal cancer from the extension of this immunological study by the evaluation of these and other parameters. PMID- 10851497 TI - Maintenance of intestinal epithelium structural integrity and mucosal leukocytes during chemotherapy by oral administration of muramyl tripeptide phosphatidylethanolamine. AB - The systemic administration of doxorubicin (DXR) decreases the number of epithelial cells and leukocytes in the small intestine of mice. Oral administration of muramyl tripeptide phosphatidylethanolamine (MTP-PE) prevented both disruption of intestinal architecture, and a decrease in the number of macrophages, and it induced the expression of IL-6, G-CSF, GM-CSF, and TNF-alpha in the intestinal tissue. The data suggest that the oral administration of MTP-PE can prevent chemotherapy-induced toxicity to the intestinal mucosa and hence infections due to translocation of aerobic bacteria from the intestine to the blood. PMID- 10851498 TI - Necessity of biotherapeutic treatments inducing TH1 cell functions in colorectal cancer. AB - Our previous data on colorectal cancer suggest that there are faults at the level of mechanisms of the proliferative responses of patients peripheral blood mononuclear cells (PBMC) to the interleukin (IL)-2 and IL-2 PBMC production, which increase with the stage advancement. The damages in the proliferative response seem to be eliminated by the costimulator effects of the signals produced by the anti-CD3 monoclonal antibody (antiCD3), and the disregulation in TH subsets of CD4+ T cells with a malfunction of TH1 cells and an expansion of TH2, might contribute to this situation. So, by using biotherapeutic treatments to allow the generation of productive immune response in these patients it is essential to identify the defect in their immune system to discover how these mechanisms should be appropriately manipulated in vivo to switch their immune response from a non-productive to a productive one. We have studied this in a group of patients and healthy subjects as the control group, performing their immunological evaluation by determining these parameters: serum levels of IL-2, interferon (IFN) gamma, IL-4, IL-6, IL-7, IL-8, tumour necrosis factor (TNF) alpha, soluble IL-2 receptor (sIL-2R), intercellular adhesion molecule 1 (sICAM 1) and CD30 (sCD30) molecules; PBMC phenotypic antigens expression (CD3, CD4, CD8, CD19, CD16, CD56, CD57, CD25) on peripheral blood mononuclear cells (PBMC); proliferative response of PBMC to IL-2, IL-4 and anti-CD3 monoclonal antibody (antiCD3). Moreover, since mutant c-Ki-ras oncogene is a very frequent finding in colorectal cancers and there are indications which suggest its involvement in tumour progression, the analysis of c-ki-ras codon 12 and 13 were determined and the statistical evaluation of the above immunological parameters were performed by comparing the patient groups with (M+) and without (M-) these mutations with each other, and with the healthy group. The results underline the necessity of biotherapeutic treatments inducing TH1 cell functions in these patients. Moreover in M+ it seems also important to solve the problem of the switch from B to macrophage cells as immune cells which present antigens, and the possible involvement of c-Ki-ras gene mutations in the impairment of T cell receptor activation (TCR). PMID- 10851499 TI - Significance of cytotoxic activity of peripheral blood lymphocytes against autologous tumor cells in patients with bladder cancer. AB - Cell-mediated immunity is an important and central mechanism of host resistance to cancer. Most reported studies have used cultured tumor cell lines as targets to assess antitumor cell-mediated cytotoxicity. However, it is difficult to translate the data generated from the cytotoxic activity against cultured tumor cell lines to cytotoxicity against autologous tumors. In a recent study, we have reported on the prognostic significance of circulating cytotoxic lymphocytes against autologous tumor cells in patients with bladder cancer. In this study, we examined whether established bladder cancer cell line like T24 or NK-sensitive K562 target cells can be substituted for autologous bladder cancer cells. The cytotoxic activity of peripheral blood lymphocytes (PBL) against freshly isolated autologous tumor cells, the T24 human bladder cancer cell line and the NK sensitive K562 human myelogenous leukemia cell line was studied in 63 patients with primary initial bladder cancer by a 12-h 51Cr release assay. The mean percent cytotoxic activity of PBL directed against autologous tumor cells, T24 cells and K562 cells were 11.3%, 18.2% and 29.4%, respectively, using an E:T of 40:1. The cytotoxic activity against T24 cells in patients with bladder cancer was higher than that in normal individuals. The anti-K562 and the anti-T24 cytotoxic activities in patients with low-stage or low-grade bladder cancer were relatively higher than those in patients with high-stage or high-grade cancer, but not statistically significant. There was no correlation between the anti autologous tumor cytotoxic activity and either the histologic grade or stage in patients with bladder cancer. The extent of the anti-autologous tumor cytotoxic activity was not paralleled with that of either the anti-K562 or the anti-T24 cytotoxic activity. In contrast, the anti-K562 cytotoxic activity correlated positively with the anti-T24 cytotoxic activity. Separation of PBL revealed that the anti-K562 and the anti-T24 cytotoxic activities were mediated mainly by the NK cells, whereas the anti-autologous tumor cytotoxic activity was mediated by both the NK cells and the T lymphocytes. These findings demonstrate that cytotoxicity against T24 or K562 cells is not of prognostic value. The magnitude of the anti-autologous tumor cytotoxic activity of PBL derived from bladder cancer patients might represent an independent and important immunological parameter to monitor disease progression. PMID- 10851500 TI - Immunotherapy with low-dose interleukin-2 and a polysaccharopeptide derived from Coriolus versicolor. AB - The purpose of the present study was to evaluate the therapeutic efficacy of locally administered low-dose interleukin-2 (IL-2) and a polysaccharopeptide (PSP) derived from Cariolous versicolor in a herpes virus Type 2-transformed murine tumor (H238) model and to determine possible mechanisms of action. BALB/c mice were inoculated subcutaneously (s.c.) with H238 tumor cells and randomized into groups: a) no tumor and no treatment control, b) tumor and no treatment control, c) tumor + IL-2 at 0 to 4 days, d) tumor + PSP at 0 to 10 days, e) tumor + IL-2 at 0 to 4 days + PSP at 0 to 10 days, and f) tumor + IL-2 at 15 to 19 days + PSP at 15 to 25 days. The IL-2 was administered s.c. at 2 x 10(4) i.u./mouse/injection; PSP was given s.c. at 2 mg/mouse/injection. No obvious toxicity was noted during the treatments. IL-2 and, to a lesser extent, PSP significantly slowed (p < 0.05) tumor progression when given alone immediately after tumor cell injection. The combination of the two modalities did not significantly enhance the antitumor effect of IL-2 alone. However, mice receiving both agents had IL-2 in the plasma, their tumors expressed low levels of transforming growth factor-beta, and their splenocyte response to mitogenic stimulation was significantly higher than in untreated controls. Changes in blood leukocyte populations and splenic oxidative burst capacity were associated with tumor presence, but not with the type of treatment. In vitro assays showed that both IL-2 and PSP can suppress the uptake of 3H-thymidine by tumor cells and that the effect is more pronounced whent the agents are used in combination. These results indicate that IL-2 and PSP can slow progression of H238 tumors and that the mechanisms of action may be related to their direct cytotoxic effects, as well to their immunomodulatory properties. PMID- 10851501 TI - Preparation and protein conjugation of a divinyl sulphone derivatized bifunctional chelating agent. AB - A new bifunctional chelating agent with a novel linking arm, 2-[p-?N-benzyl-N-(2 vinylsulfoethyl)?- (aminobenzyl)?-1,3-propane-diamine-N,N,N',N'-tetraacetic acid (VS-PDTA) was synthesized and was conjugated to protein for the purpose of attaching radiometals to monoclonal antibodies (MAbs). The effect of various parameters such as ligand concentration, protein concentration, pH, temperature and reaction period on the conjugation have been examined using chromatographic (SE and TLC) analysis after labeling with 111In. The parameters and chemical variables studied have significant effects on the efficiency and rate of protein conjugation. PMID- 10851502 TI - Pharmacokinetics of iodine-125 CC49 monoclonal antibody in patients with colon cancer. AB - Radioimmunoguided Surgery techniques which use radiolabeled tumor specific markers and an intraoperative detector in an attempt to improve therapy and survival in patients with cancer have been under development for over fifteen years. Monoclonal antibody (MAb) CC49 is a second-generation murine IgG1 which has improved localization properties over its predecessor, MAb B72.3, and has been studied in a number of patients. In order to determine the pharmacokinetics of iodine-125 (125I) CC49 MAb, size-exclusion, high-performance liquid chromatography (HPLC) was used to assess radioactive components in serum and urine following administration of the drug to colon cancer patients. METHODS: Five patients received an intravenous infusion of 10 mg of MAb CC49 labeled with 2 mCi 125I. Following infusion, serum and urine specimens were collected from patients at predetermined time intervals prior to surgery. HPLC analysis of these specimens was completed to determine the radioactive species in each sample. RESULTS: Serum and urine specimens showed that serum levels of CC49 decrease exponentially and become unmeasurable by day 14 (half-life 1.89 days, +/- 0.19), with a steady, low-level of free 125I measurable in postinjection serum until day 21 after infusion. There was no evidence of MAb fragmentation or antibody:antigen (Ab:Ag) complex formation in serum, and no evidence of whole MAb, F(ab')2, or Fab fragment excretion in urine. Preinjection sera with MAb added in vitro also failed to demonstrate Ab:Ag complex formation. Analysis of urine showed low level excretion of free 125I which peaked by day 1 and declined exponentially through day 21, with a very low molecular weight (< 1 kDa) MAb fragment excreted in urine between 1 and 21 days. CONCLUSION: Radioiodinated 125I CC49 MAb remains in serum of cancer patients approximately 14 days, and tissue radioactivity beyond this time may reflect tissue sequestered MAb and/or free 125I and not "bloo pool" radioactivity. CC49 MAb appears to be deiodinated in small but significant quantities before it is metabolized, and clearance of radioactivity is mainly in free 125I form in urine. Measurable quantities of a < 1 Kda MAb fragment in urine and not serum may suggest a renal mechanism of MAb metabolism, but may also represent a metabolic end product of MAb metabolism with a very short serum half life (T1/2) which accumulates in urine. PMID- 10851503 TI - Progress in the monoclonal antibody therapy of cancer. PMID- 10851504 TI - The role of interleukin-2 in cancer therapy. PMID- 10851505 TI - Prognostic markers in interleukin-2 therapy. PMID- 10851506 TI - Outpatient-based subcutaneous interleukin-2 monotherapy in advanced renal cell carcinoma: an update. AB - To minimize interleukin-2-related toxicity while retaining its efficacy, a treatment schedule utilizing subcutaneous IL-2 was evaluated in a phase II setting. Eighty unselected, consecutive patients with metastatic or recurrent renal cell carcinoma (RCC), mean age 58 years (range, 21 to 76), received IL-2 on an outpatient basis, 5 days per week for 4 or 6 consecutive weeks. During the first 5-day cycle, a dose of 18 million IU IL-2 was administered once a day; during subsequent cycles the dose in the first two days was reduced to 9 million IU. Two 6-week or three 4-week courses were given maximally. Patients who had completed at least one full course were considered evaluable. To circumvent flu like symptoms, all patients received a maximum oral dose of 3 g acetaminophen daily. Seventy-seven patients were assessable for response. Three (4%) complete responses (CR) and 6 (8%) partial responses (PR) were observed, and 44 (57%) patients had stable disease (SD). Response durations were 64, 29, 29+ months for the CR and 2, 6, 8, 11, 32, 47 months for the PR. The median length of survival of all patients was 12 months, whereas the median survival of responders and non responders was 35+ and 10+ months, respectively (P < 0.001). Side effects included fever, chills, nausea, vomiting, and transient inflammation and induration at the injection sites. These complications were acceptable, even in the patients with concomitant disease, and completely disappeared after cessation of IL-2. Subcutaneous IL-2 mediates antitumor responses, has limited side effects and is also suitable for elderly RCC patients with concomitant disease. PMID- 10851507 TI - Interleukin-2 in outpatients with renal cell carcinoma: SCAPP1 trial. PMID- 10851508 TI - Adjuvant treatment with IL-2 or interferon-alpha in renal cell carcinoma: a French multicentric study. PMID- 10851509 TI - Interleukin-2 in neuroblastoma: clinical perspectives based on biological studies. AB - Stage IV neuroblastoma (NB) is a disease with a poor prognosis. Chemotherapeutical intensification and hematological rescue with autologous bone marrow transplantation (ABMT) achieve some complete remissions (CR), but most patients relapse during the first year. Immunotherapy could be an alternative in this situation of high risk of relapse due to residual disease and ABMT-related immunodepression. Ten stage IV NB patients in CR or very good partial remission have been treated with recurrent 5-day cycles of high doses of Interleukin-2 (IL2) after ABMT throughout one year (usually 5-6 cycles). Natural killer (NK) and lymphokine-activated killer (LAK) cytotoxic activities, as well as phenotype and number of circulating NK cells were determined, before and after each course of IL2 treatment. The effects promoted by IL2 varied during treatment: early cycles of IL2 induced a great extent of cell expansion, mainly on CD3-/CD16 /CD56+bright and CD8+dim cell phenotypes; conversely, late courses of IL2 promoted higher NK cytotoxic activity but a lesser increase on circulating NK cells. The induction of LAK activity did not significantly differ from early and late IL2 treatments. Clinical results are still inconclusive due to the small number of patients. The median follow-up of patients treated with IL2 is 24 months and the disease free survival (DFS) probability is 0.80 +/- 0.12 vs 0.16 +/- 0.15 from a historical control with identical treatment, but in the absence of IL2 treatment (p < 0.005). IL2 treatment-related toxicity was mild and no interruption of the treatment was required. Extremely accurate hydric control was carried out to avoid, as much as possible, the consequences of vascular leak syndrome, one of the most important toxic effects of IL2 treatment. The results presented here suggest an evolution of NK activity during IL2 treatment after ABMT, which should be taken into account for the designing of new immunotherapeutical protocols and opens a promising perspective in treatment of stage IV neuroblastoma. PMID- 10851510 TI - Cytokines in combination with chemotherapy for advanced renal carcinoma--the importance of patient selection. AB - An outpatient regimen of interferon-alpha (IFN-alpha), interleukin-2 (IL-2) and 5 fluorouracil (5-FU) was previously reported to have significant activity (response rate 48.6%) in patients with advanced renal cell carcinoma (RCC). The patient group reported were generally of good performance status (PS), had undergone previous nephrectomy and would be considered of good prognosis with respect to response and survival after treatment with IL-2. The characteristics of patients with RCC referred to specialist units in the UK differ from that patient group in that many patients present with metastatic disease, are of poor PS and are considered unfit for nephrectomy. We tested the three drug regimen in a representative patient group of 55 patients who had: median PS of 1 (range 0 2); median time from diagnosis to treatment of 2.7 months (0.2-113); and median number of sites of disease 3 (1-5). 22/55 had not had prior nephrectomy and 31 were considered of poor risk, 15 moderate risk and only 9 of good risk. Treatment consisted of an 8 week cycle of IFN-alpha 6 MU/m2 day 1 weeks 1 and 4 and thrice weekly weeks 2-3 and 9 MU/m2 thrice weekly, weeks 5-8. IL-2 20 MU/m2 days 3-5, weeks 1 and 4 and 5 MU/m2 thrice weekly weeks 2-3. 5-FU 750 mg/m2 day 1 of weeks 5-8. There were no complete responses (CR), 9 (17%) partial responses (PR) and 13 patients (24%) had stable disease. Sixteen patients withdrew early from treatment and were not evaluable for response. Amongst 25 evaluable patients who had undergone nephrectomy the response rate was 32% (95% CI: 14-50%). Only 1 response was seen in patients who had not undergone nephrectomy. Survival was predicted by PS, nephrectomy, number of sites of metastasis and risk group. Most patients experienced significant toxicity of grade I/II but few grade III/IV toxicities were seen as compared to intravenous IL-2 regimens. These data are part of a large data set that has been submitted for publication in The British Journal of Urology. The regimen has been shown to have activity but this is seen predominantly in patients of good PS, with prior nephrectomy and limited sites of disease. Patients of poor risk are likely to experience significant toxicity without benefit and should be offered alternative palliative therapies. PMID- 10851511 TI - Cytokines and tumor vaccination. AB - Patients with locally advanced stages of renal cell carcinoma are at high risk of relapse or progress even after initial radical surgery. Based on the proven efficacy of adoptive and active immunotherapeutic approaches of metastatic renal cell carcinoma, a phase II trial was started in 1989 using autologous, Newcastle disease virus modified and lethally irradiated tumor cell vaccines in combination with low-dose recombinant interleukin-2 (1.8 million U) and recombinant interferon-alpha 2a (1.0 million U) for a surgical adjuvant treatment. Patients were vaccinated (subcutaneous injection) once a week for 8-10 weeks and the treatment was started about 4-10 weeks after surgery. Up to now more than 208 patients with locally advanced renal cell carcinoma (stages pT2-3a, N1-2, M0; pT3b-4, N0-2, M0) were vaccinated after initial radical surgery (tumor nephrectomy with lymph node dissection and ipsilateral adrenalectomy and if necessary in combination with en bloc removal of venous extensions). We overview a follow-up of 203 evaluable patients with a median disease-free survival of 21 months (range of 2-64 months). During this observation period 18 relapses (8.9%) were diagnosed with 3 local relapses (1.5%), 10 lymph node metastases (5%) and/or distant organ metastases in 9 cases (4.5%). These progressive patients' disease was treated by surgery and/or combined immunochemotherapy. Toxicity encountered on this tumor cell vaccination was mild (WHO grade 1) and was characterized by flu-like symptoms and fever up to 38.8 degree Celsius for some hours beginning at 4 hours after the vaccine/cytokine application. Occasionally a transient local inflammation at the site of injection was observed. The comparison of the risk factor-adapted group of adjuvant treated renal cell carcinoma patients (locally advanced stages) with historical data gave evidence for an improvement in disease free survival on vaccination treatment. Although this was not a prospective randomized trial, we can summarize that the surgical adjuvant treatment of autologous tumor vaccines in combination with low-dose cytokines may improve relapse-free and overall survival in patients with locally advanced renal cell cancer. PMID- 10851512 TI - Linomide and interleukin-2 in patients with advanced renal cell carcinoma. AB - Quinoline-3-carboxamide (Linomide) is a novel, synthetic immunomodulator acting via immunologic and non-immunologic mechanisms. It has shown efficacy against various malignancies, experimental autoimmune encephalomyelitis, and septic shock in animal models and has been investigated for clinical use in minimal residual myeloid leukemia with promising results. Interleukin-2 has shown considerable efficacy in palliative anti-tumor-treatment of advanced renal cell cancer, revealing remission rates of up to 40% in combination therapy regimens. Linomide is reported to exhibit synergistic effects with interleukin-2. Here we report on a clinical phase I/II study examining tolerance and efficacy of a combination therapy schedule of SQ interleukin-2 and PO Linomide in advanced renal cell cancer. Seventeen patients received 10 IU/m2 interleukin-2 per week for 8 weeks, resting interleukin-2 for another 8 weeks. In week 5 they started 5 mg Linomide daily, continued with 10 mg from week 7 to 16. No objective remissions were observed. Among 15 patients evaluable for response, 10 (66.7%) were progredient during the study. Three patients died during the observation period, including two not evaluable for response. Median survival was 4.0 months, median progression-free survival 2.5 months with a Kaplan-Meier estimate of 3.63 months. Fever, reduced general condition, nausea/vomiting, dyspnea, anorexia, chills and hypotension were the most common side effects, reaching WHO grade 3 in 6 and grade 4 in 2 cases. In summary, Linomide in combination with interleukin-2 provides no advantages in efficacy or toxicity over other therapy regimens employing interleukin-2. PMID- 10851513 TI - Detection of pseudomyxoma peritonei by radioimmunohistochemistry and radioimmunoscintigraphy. AB - Pseudomyxoma peritonei (PP) is a local slowly progressing disease with typical abdominal swelling. Treatment is uneffective and the long-term prognosis is poor. Conventional radiology provides usually only a delineation of low density area relating gelatinous masses accumulating in the peritoneal cavity. In this study, immunohistochemistry based on digital quantitative autoradiography utilizing radiolabelled monoclonal antibody B72.3 (MoAb) recognizing TAG-72 antigen on epithelial carcinomas was used for diagnosis of pseudomyxoma (7 patients). The PP patients were studied with radioiodinated I-131-labeled MoAb after intravenous (2 patients) and intraperitoneal (7 patients) injections. Radioactivities of MoAbs varied considerably in the tumors. Both intra- and extracellular staining pattern was observed by immunohistochemistry. Gamma imaging at 1, 3, 7 and 14 days after i.v. injection (2 patients) revealed targeting of all known lesions. The intraperitoneally injected MoAb (7 patients) retained long time in the peritoneal cavity, specific tumour targeting was seen up to 16 days by an antibody-SPECT, while maximum blood radioactivity was measured between 8-12 hrs. Radiolabelled B72.3 MoAb recognizing TAG-72 antigen is also present within pseudomyxoma cells. It can be used for radioimmunohistochemistry of PP. Accurate imaging of PP is possible by MoAb suggesting earlier diagnosis and more accurate location of residual disease after operations, and evaluating treatment response. Estimated tumour dose for intraperitoneal tumour (MIRD formalism) was 13 mGy/MBq. This indicates that the radioiodinated B72.3 antibody can be used for in vivo targeting and therapeutic applications of intraperitoneal pseudomyxoma. PMID- 10851514 TI - Establishment of a rat colonic carcinoma model for study of immunoreagents against the human tumor-associated TAG72 antigen. AB - TAG72 originally defined by the mouse B72.3 antibody is a mucin-like, human tumor associated antigen present in more than 85% of human colonic adenocarcinomas. Establishment of a tumor model expressing the TAG72 antigen in immunocompetent animals would be of great benefit in evaluating the therapeutic efficacy and studying anti-tumor immune mechanisms of anti-TAG72 immunoreagents. In this study, we screened 6 animal tumor cell lines including 3 derived from mouse colonic adenocarcinomas (MCR-26, MCR-38-LD and CA-51), 1 from mouse ovarian adenocarcinoma (MOT), 1 from rat colonic adenocarcinoma LMCR, and 1 from rat mammalial adenocarcinoma (R3230AB) for TAG72 expression by using the B72.3 antibody. Immunohistochemistry disclosed significant amounts of TAG72-expression in the dimethylhydrazine-induced BDIX rat colonic adenocarcinoma LMCR. The rat TAG72 antigen purified from rat LMCR tumors showed strong immunoreactivity for the B72.3 antibody in ELISA analysis and displayed a smear band of high molecular weight in Western blotting, which is similar to the human TAG72 antigen purified from human LS174T colonic adenocarcinoma. In addition, the iodinated B72.3 antibody was able to localize LMCR tumor in BDIX rats. Therefore, this rat LMCR model should be useful in studying human colonic cancer, especially in evaluating the therapeutic efficacy of anti-TAG72 immunoreagents such as the recombinant fusion proteins possessing the anti-TAG72 antibody fragment and the cytokine moiety. PMID- 10851515 TI - FDA update: 1996. PMID- 10851516 TI - Granulocyte colony stimulating factor (G-CSF): biology and clinical status. PMID- 10851517 TI - Biological modifiers (etretinate (changed from etetrinate) and alfa 2a) in the treatment of refractory cutaneous T-cell lymphoma. AB - To assess the efficacy and toxicity of biological modifiers in combination etetrinate, 0.8 mg/kg/day, po and interferon alfa 2a 9.0 MU, three times at week) in the treatment of refractory cutaneous T-cell lymphoma (CTLC) we began a clinical study on 12 heavily treated patients. After 1 year on treatment 10/12 patients (83%) achieved complete response. Two patients were considered failures with disease progression. After a median follow-up of 3 years, seven patients (56%) remained in complete remission. Toxicity was mild. All patients received 93% of the planned dose of etetrinate and interferon. We feel that biological modifiers, as etetrinate and interferons, are agents with limited hematological toxicity even in higher doses. The combination of two agents, with different mechanisms of action, could improve the outcome in patients with refractory CTCL. Controlled trials are necessary to define the roles of this type of therapy as first line of treatment. PMID- 10851518 TI - The 10th International Conference on Monoclonal Antibody Immunoconjugates for Cancer: Radiodosimetry and Radiopharmaceuticals for Therapy. PMID- 10851519 TI - Criteria for the selection of radionuclides for targeting nuclear antigens for cancer radioimmunotherapy. AB - The potential of utilizing immunoconjugates to selectively deliver radionuclides for the destruction of tumors has stimulated much research activity. From dosimetric and other considerations, the choice of radiolabel is an important factor that needs to be optimized for maximum effectiveness of radioimmunotherapy (RIT). This paper reviews and assesses a number of present and future radionuclides that are particularly suitable for RIT based on the various physical, chemical, and biological considerations. Although intermediate to high energy beta emitters (with and without gamma photons in their emission) possess a number of advantages for most RIT, the use of alpha, Auger, and short range conversion electron emitters could be attractive for targeting nuclear antigens when the radioimmunoconjugate is internalized into tumor cells. Factors relating to the production and availability of candidate radionuclides as well as their stable chemical attachment to monoclonal antibodies are discussed. PMID- 10851520 TI - Oldenlandia diffusa and Scutellaria barbata augment macrophage oxidative burst and inhibit tumor growth. AB - Oldenlandia diffusa (OD) and Scutellaria barbata (SB) have been used in traditional Chinese medicine for treating liver, lung and rectal tumors. In this study we determined the effects of these phytochemicals on oxidative burst as an indicator of phagocytic function in a murine macrophage cell line J774 using an automated micro-fluorometric assay. A dose-dependent augmentation of oxidative burst was observed with OD as well as SB. The effect of OD and SB on the growth of a murine renal cell carcinoma (Renca) was also determined. Balb/c mice were transplanted subcutaneously with 1 x 10(5) Renca cells and were randomized into groups of 10 to receive oral feeding of OD (4 mg/day), SB (4 mg/day), or saline as control. Oral feeding with either OD or SB significantly inhibited the growth of Renca in mice. The data of this study show that OD and SB are capable of enhancing macrophage function in vitro and inhibiting tumor growth in vivo. PMID- 10851521 TI - Putative effects of mitogenic lectin therapy corroborated by alloactivation data. AB - A firm theoretical case for PHA and other plant mitogens as superior immunomodulators has previously been presented, but direct confirmatory evidences have been compromised by experimental studies involving excessive dosages of erythroagglutinating PHA that often compromised the circulation in smaller animals, while inadequate amounts were applied in humans because this mitogen's availability in nonagglutinating form was restricted. The resulting underestimation of efficacy has failed to inspire the production of industrial quantities of mitogens required for reliable clinical trials. As a means of circumventing this dilemma, past favorable results from the immuno-stimulating activities of allocativation have been extrapolated to forecast the effects to be anticipated from mitogenic modulation. Such an extrapolation would underrate the latter's impact, which would neither be confined to stimulation nor dependent on the uncertainties of engraftment. This review cites clear examples showing that all recognized immune system pathways have been stimulated by alloactivation except perhaps the ADCC, and an example of this pathway's activation has been shown to have occurred with PHA therapy itself. Of the mitogenic lectins currently available, PHA, Con A, and PWM have each shown rare instances of hypersensitization that hopefully might be eliminated by exclusion of contaminants with recombinant DNA methods of production. PMID- 10851522 TI - Inhibition of cancer cell growth by internalized immuno-histone conjugates. AB - MAb NS 88 directed against breast cancer cells, which is internalized and translocated to the cell nucleus, was conjugated with histone and labeled with 125I. 125I-MAb-histone complexes (M(r) 250,000) were internalized by breast and cervical cancer cells and localized in the cytoplasm and chromatin. Electrophoretic analysis of the cells extracted from the conjugates revealed the same molecular weights of the cytoplasmic and chromatin complexes as those of the native conjugate. Nicotine (0.1%), which suppresses lysosomal degradation, stabilized the conjugates within the cell and prolonged the presence of nondegraded complexes inside the cytoplasm and chromatin from 1 day to at least 3 days. MAb-histone complexes, but not MAb alone, inhibited RNA synthesis and proliferation of cervical and breast cancer cells. A new application of internalized MAbs as the vehicles for protein inhibitors of transcription or replication is discussed. PMID- 10851523 TI - Immunochemical study of human immunoglobulin G Fc region. AB - Fifteen murine monoclonal antibodies (mAb) were produced that reacted with conformational epitopes present on the Fc portion of all human IgG subclasses (PAN-Fc). Inhibition and sandwich ELISA were used to elucidate overlapping and distinct epitopes. The epitopes were aligned to form a continuous "chain" of overlapping determinants from the CH2-CH3 interface to the direction of hinge. The five more hinge-proximal epitopes demonstrated high lability both in competitive and sandwich assays, being completely or partially destroyed when any of these 15 other mAb bound the IgG first. Heat-inactivation of IgG caused full disruption of these epitopes. In contrast, epitopes situated at the opposite distal of the "chain" were more stable and mAb binding could only be affected by occupying an overlapping epitope. Under heat-inactivation these epitopes were affected, but not completely destroyed. Human IgG class anti-DNA autoantibodies were bound to insolubilized dsDNA and their reaction with PAN-Fc mAb was studied. mAb titration plots on IgG and dsDNA-IgG were compared. Five epitopes proved to be altered by antigen (dsDNA) binding. Two of these were the labile hinge proximal epitopes and the other three were situated near the CH2-CH3 interface. Cross-reactivity of mAb with xenogeneic IgG was also studied. An "epitope map" of the crystallographic model of human IgG Fc portion drawn was based on these experimental data and printed matter, concerning the location of subclass specific amino acids and homology regions of human and animal IgG. PMID- 10851524 TI - Magic bullets: "deja vu all over again". PMID- 10851525 TI - Why event-free survival is better than tumor response or other measures of survival as an endpoint in cancer trials. PMID- 10851526 TI - Synthetic thymic fraction 5: effects of high dose administration on circulating lymphocytes in patients. AB - The synthetic pentapeptide (Arg-Lys-Asp-Val-Tyr; TP-5) corresponding to the active site of the hormone thymopoietin, was given at the dose of 300 mg/m2/day (1 day), higher than the usually administered, to a group of 27 immunodepressed patients in order to determine the tolerability and the immunomodulatory activity. The examination of a series of hematological parameters including counts of differential clustering of lymphocytes by cytofluorimetric analysis was performed 24 hr and 48 hr after treatment, and repeated at different intervals up to 14 days after treatment. TP-5 caused a significant increase of circulating lymphocytes and particularly of CD3+CD4+ and CD3+CD8+ subtypes, peaking at 48 hr and maintaining the increased values up to the last examination on day 14 from treatment. A faster increase (zenith at 24 hr) was observed for CD4+ cells, in comparison with CD8+ cells (zenith at 48 hr). The number of patients that increased total lymphocytes or lymphocyte subset after treatment ranged between 52.6 (CD4+ cells) and 69.2% (NK cells), whereas about 7.7% (NK cells) to 36.9% (CD4+ cells) remained unchanged and a smaller amount of 10.5% (CD4+ and CD8+ cells) or 23.1% (NK cells) showed a decrease greater than 10% of their respective basal value. No significant relationship between responders and non-responders can be found on the basis of previous treatments, cancer type, sex or age. PMID- 10851527 TI - Immunological directives for biotherapy improvement in the treatment of colorectal cancer. AB - A major cause of failure in biotherapy in cancer may be the non-existence of predictive indices to individualize the substances which might be helpful to switch the patients' immune response to tumour from an non-productive to a productive one. The defect in the patients' immune system needs to be identified and so the evaluation of their responses to biotherapeutic agents, in correlation to the disease progression is essential. We have addressed this problem in colorectal cancer at systemic level by examining the proliferative response of peripheral blood mononuclear cells (PBMC) to interleukins (IL) IL-2, IL-4, antiCD3 monoclonal antibody (antiCD3), IL-2+CD3, IL-2+IL-4, IL-4+CD3, IL-2+CD3+IL 4; the PBMC expression of phenotypic antigens CD3, CD4, CD8, CD25, DR, CD16, CD56, CD57 and CD19 in the patients and healthy subjects as control group. Analysing our data, it seems that as the disease progresses in these patients the peripheral blood cells change their ability to respond to activation agents which appears to be due to a phenotypic modification of their subsets. Our overall results might give a possible explanation of the variable responses to biotherapy in colorectal cancer patients. PMID- 10851528 TI - Treatment of stage D2 hormone refractory carcinoma of the prostate with 5 fluorouracil and Roferon-A: a Southwest Oncology Group study. AB - Based upon prior data suggesting that alpha-interferon possesses chemomodulatory activity, the Southwest Oncology Group conducted a study in which patients with hormone refractory, metastatic (stage D2) adenocarcinoma of the prostate were treated with 5-fluorouracil (5-FU) and Roferon-A. All patients had bidimensionally measurable disease. Treatment consisted of 5-FU 750 mg/m2/day by continuous i.v. infusion for 5 days with Roferon-A 9 million units subcutaneously ono days 1, 3 and 5. Roferon-A was continued three times weekly throughout treatment. Following a one week hiatus from 5-FU (week 2), 5-FU was continued at a dose of 750 mg/m2 i.v. bolus weekly. Nineteen patients were evaluable for toxicity. The most common toxicities were gastrointestinal and mucosal, hematologic and a flu-like syndrome. There were no deaths related to treatment. Among the 14 patients evaluable for response, the response rate was 0% (95% confidence interval, 0-18%). Thirteen of the 19 evaluable patients have died with a median survival of 9 months. The combination of 5-FU and Roferon-A does not have sufficient activity against advanced, hormone refractory prostate cancer to warrant further investigation. PMID- 10851529 TI - Radioimmunoscintigraphy of gastric adenocarcinomas with 99mTc-chimeric ccM4 antibody. AB - Fourteen patients with gastric lesions were studied for radioimmunoscintigraphy using the 99mTc-ccM4 chimeric antibody reactive with the human tumor-associated TAG72 antigen. These include 10 gastric adenocarcinomas, 3 gastric ulcers and 1 gastric lymphoma. Each patient received an infusion of 30-40 mCi 99mTc-ccM4 antibody. Gamma camera scans were performed at 12-h postadministration. All patients underwent operation after radioimmunoscintigraphy. All surgical specimens including gastric adenocarcinomas, lymphoma and ulcers were confirmed by histopathological studies. The 99mTc-ccM4 antibody was able to detect 6 out of 10 gastric adenocarcinomas (60%) by scans. Its accuracy was even 5 out of 6 (83%) in a group of patients with well/moderately-differentiated adenocarcinomas. In addition to the primary adenocarcinomatous lesions, the 99mTc-ccM4 antibody was also able to detect metastatic lesions in liver and lymph nodes. The smallest adenocarcinomatous lesion detected by scans was about 1 x 1 x 2 cm3. All four patients with gastric ulcers or lymphoma showed negative scan results. Therefore, the chimeric ccM4 antibody may be useful in radioimmunoscintigraphy of gastric adenocarcinomas, especially the well/moderately-differentiated adenocarcinomas. PMID- 10851530 TI - 186Re-labeled antibodies to p185HER2 as HER2-targeted radioimmunopharmaceutical agents: comparison of physical and biological characteristics with 125I and 131I labeled counterparts. AB - Overexpression of the HER2/neu protooncogene has been shown to correlate with poor clinical prognosis. A murine monoclonal antibody (4D5) directed against the extracellular domain (ECD) of p185HER2 has been shown to inhibit in vitro and in vivo growth of carcinomas overexpressing HER2 and has been humanized (rhuMAb HER2). The objective of the study was the identification of an agent which might be useful for in vitro studies, tumor imaging and/or radioimmunotherapy by linking beta-emitting radionuclides to these HER2-targeted antibodies. Murine 4D5 and humanized rhuMAb HER2 were radiolabeled with 125I, 131I or 186Re. Physical characteristics (TCA precipitability, SDS-PAGE, size exclusion chromatography), binding affinities to the HER2 ECD (in an ELISA and on SK-BR-3 cells) and antiproliferative activities of the radiolabeled antibodies were determined. Although 131I-4D5 and 131I-rhuMAb HER2 usually retained > 85% ECD binding, they exhibited increased aggregation and fragment content, drastically reduced antiproliferative activities and poor stability upon storage at 4 degrees C. For these antibody preparations, conservation of binding did not necessarily correlate with preservation of bioactivity indicating the importance of bioactivity determinations in radiolabeled antibody studies. Conversely, 4D5 and rhuMAb HER2 labeled with 125I or 186Re maintained physical properties, ECD binding, antiproliferative activities and were stable upon storage at 4 degrees C for at least 8 days. The superior retention of physical and biological characteristics of 186Re-labeled 4D5 and rhuMAb HER2 compared with their 131I labeled counterparts suggests the potential for their use as radioimaging and radioimmunotherapeutic agents in the treatment of HER2 overexpressing tumors. PMID- 10851531 TI - Attenuated Salmonella typhimurium containing interleukin-2 decreases MC-38 hepatic metastases: a novel anti-tumor agent. AB - Currently, there is no long-term effective treatment for unresectable hepatic malignancies. Salmonella sp. are known to naturally track to the liver during active infection. To develop a biological vector for delivery of Interleukin-2 (IL-2) to the liver for anti-tumor purposes, the avirulent and highly immunogenic chi 4550 strain of Salmonella typhimurium was used as a vector for IL-2. The gene for human IL-2 was cloned into plasmid pYA292 (renamed pIL-2) and inserted into the attenuated Salmonella typhimurium and renamed [chi 4550 (pIL-2)]. This transformant was found to produced biologically active IL-2 demonstrated by NK cell activation in a 4 hour chromium release cytotoxicity assay. To determine anti-tumor potential, MCA-38 murine adenocarcinoma cells were injected intrasplenically into C57BL/6 mice to produce hepatic metastases and metastases were subsequently enumerated after 12 days. Statistical significance was determined by ANOVA with Fisher's test for significance. Hepatic metastases enumerated by blinded observers revealed that the mean number of metastases was 106.4 in control mice, 103.7 in mice gavage fed attenuated salmonella without IL 2 [chi 4550(pYA292)], and 44.3 in mice fed the chi 4550(pIL2); (ANOVA: p < 0.01). Culture of livers and spleens in mice administered a single gavage dose of salmonella demonstrated persistent colonization for up to 4 weeks. No observable toxicity was seen to either IL-2 or salmonella. These studies demonstrate that the chi 4550(pIL2) is a novel form of in vivo biotherapy which produces biologically active IL-2 and employs the oral route of administration to stimulate an immune response against malignancy in the liver. PMID- 10851532 TI - Interleukin-2 secretion by transduced and unselected BDL-2 lymphoma results in increased survival in mice with previously established disseminated disease. AB - The transfer and expression of cytokine genes into malignant cells to provide a more effective tumor response has shown promise. The majority of murine models in which tumor vaccination strategies have been tested have utilized selected and expanded clones of tumor cells, which is impractical clinically. In a model of murine B lineage lymphoma (BDL-2), we compared the effectiveness of tumor vaccines composed of a) a BDL-2 clone established by G-418 resistance following transduction with the LIL2SN retrovirus and screened for maximal IL-2 secretion, b) a syngeneic fibroblast line transduced with LIL2SN and screened for G-418 resistance and IL-2 expression, which was co-injected with the parental line, and c) a heterogeneous (unselected) population of BDL-2 cells transduced with the MFG/IL2 virus, reported to provide enhanced expression of cytokine genes and minimize the need for selection. Testing of splenocytes derived from vaccinated animals reveals that injections of BDL-2 expressing IL-2 results in an increased capacity of splenocytes to kill BDL-2 in vitro, compared to vaccination of BDL-2 alone or in combination with IL-2 secreting fibroblasts. We show that a vaccine composed of MFG/IL2 transduced, unselected BDL-2 cells is equivalent or superior to a clone derived from LIL2SN transduction in prolonging survival of animals with previously established tumor. These studies provide evidence that transduction of tumor with MFG based vectors without in vitro selection leads to expression of high levels of IL-2 and can impact the survival of animals with disseminated tumor. PMID- 10851533 TI - Government oversight or what? PMID- 10851534 TI - Role of cysteine endopeptidases in cancerogenesis. AB - Role of cysteine endopeptidases in cancerogenesis steps: neoplastic transformation, invasion and metastasis is reviewed and discussed. Positive correlation between tumor invasiveness, as well as its metastatic potential and secretion of cysteine endopeptidase (particularly cathepsins B and L) has been documented well in literature. Based on our recent results we postulate that serum endopeptidase-like activity could be used as a marker of cancer aggressiveness in diagnostic procedures in oncology. We also propose that the cysteine endopeptidase inhibitor levels (total, active and latent) could be useful factors for recognising the activation of the organism self-defence mechanisms against cancer. In addition, our idea of use of urinary cysteine peptidase inhibitors (UCPI) as potential anticancer agents is presented and discussed. PMID- 10851535 TI - Upregulation of DF3, in association with ICAM-1 and MHC class II by IFN-gamma in short-term human mammary carcinoma cell cultures. AB - This study was designed to determine whether in vitro exposure of isolated short term human primary and metastatic breast tumor cell cultures to interferon-gamma (IFN-gamma) could enhance expression of the breast tumor associated DF3 antigen in association with the intercellular adhesion molecule 1 (ICAM-1) and MHC class II molecules. Cell cultures were established from primary solid tumors and metastatic cells as previously described (Sgagias et al., 1995). Data show that recombinant human IFN-gamma treatment, in vitro, dramatically increased the breast tumor associated DF3 antigen, in association with ICAM-1, and MHC class II antigens in primary breast cancer cell cultures. All primary breast tumor cell cultures constitutively expressed high levels of HLA-class I antigen. Metastatic breast cancer cell cultures expressed high levels of DF3 and recombinant human IFN-gamma treatment, in vitro, upregulated ICAM-1 and MHC class II antigens before and after passage of the metastatic cells through the nude mouse. Metastatic breast cancer cells similar to primary breast cancer cells constitutively expressed high levels of MHC class I antigens. In addition, three LAK cell lines significantly lysed the primary and the metastatic breast tumor cell cultures to the same degree before and after passage of the metastatic cancer cells through the nude mouse. These data indicate the upregulation of the breast tumor associated DF3 antigen in vitro after IFN-gamma treatment and its persistence in vivo, after passage of the metastatic breast cancer cells through the nude mouse. The ability of IFN-gamma to upregulate the breast tumor associated DF3 antigen in association with the ICAM-1 and HLA class II antigens may play an important role in eliciting an immune response which may contribute to the immunodiagnosis, and immunotherapy of breast cancer. PMID- 10851536 TI - Weekly 5-fluorouracil and interferon-alfa-2b in metastatic cancer. AB - Twenty-four patients with advanced cancer were treated with 5-Fluorouracil 600 mg/m2 intravenously and interferon-alfa-2b 3 MU subcutaneously both given weekly for 6 weeks followed by a 2-week hiatus. The median age of patients treated on this study was 66 years. Mild to moderate leukopenia, nausea/emesis, and anemia were the most common toxicities. There were no treatment-related deaths and only 1 episode of grade IV toxicity (leukopenia in a patient who was receiving concurrent radiation therapy). Two complete responses, 7 partial responses and 1 prolonged minor response (pancreas cancer--12+ months) have been seen. Weekly 5 FU and interferon is well-tolerated and shows activity in selected patients with advanced cancer. PMID- 10851537 TI - Flavonoid-mediated tumor growth suppression demonstrated by in vivo study. AB - Many of flavonoids as well as bioflavonoids extracted from higher plants, which were earlier revealed to have tumor cell growth suppression activity in vitro, were examined for their effect on tumors in vivo. Balb/c mice were inoculated i.p. with syngeneic tumor cells, meth/a, and then provided with flavonoids dissolved in their drinking water during the course of their survival time. Many flavonoids were effective in prolonging the survival period. Furthermore, flavonoids that did not show suppressive activities in the in vitro experiments were effective in the in vivo assay. The data suggested that sugar bonded to the A ring, which suppresses tumor growth inhibition in vitro, plays an important role. Many aglycones that were effective in the in vitro assay on the tumor growth suppression were not effective in the in vivo assay. The reason for this seems to be that most aglycones are unstable and thus break down in vivo. No acute nor chronic toxicity of flavonoids was observed in the mice. PMID- 10851538 TI - Cationic chlorophyl derivatives with SOD mimicking activity suppress the proliferation of human ovarian cancer cells. AB - Derivatives of chlorophyl, e.g. Fe-chlorin e6-Na, alpha, beta, gamma, delta Tetraphenylporphine-tetrasulfonic acid disulfonic acid salt tetrahydrate (Fe TPPTS) and alpha, beta, gamma, delta-Tetrakis (4-N-trimethylaminophenyl) porphine, tetra (p-toluensulfonate (Fe-TTMAPP), express SOD mimicking activity. Examination was made of suppressive effects of human cancer cell lines by derivatives of chlorophyl. Fe-TPPTS and Fe-TTMAPP suppressed proliferation of the human ovarian cancer cell lines but Fe-chlorin e6-Na failed to suppress the proliferation. Lipid peroxide was increased by application of Fe-TPPTS and Fe TTMAPP, but decreased by application of Fe-chlorin e6-Na. SOD activity of the cancer cells did not change by application of these drugs. TPPTS and TTMAPP have a cationic charge but Fe-chlorin e6-Na has an anionic charge. It is suggested that charge of these drugs relates to the suppressive effects of the cancer cell proliferation. PMID- 10851539 TI - Improved tumor localization and radioimaging with chemically modified monoclonal antibodies. AB - A method for the chemical modification of monoclonal antibodies using the heterobifunctional crosslinker succinimidyl 3-(2-pyridyldithio)propionate (SPDP), has been developed which dramatically alters the physiochemical properties of antibody reagents. For these studies, three murine monoclonal antibodies, B72.3, Lym-1, and TNT-1 were used to demonstrate the effects of chemical modification on clearance and biodistribution in tumor-bearing nude mice. In vitro, all three antibodies, modified to the same degree with SPDP, showed equal immunoreactivities and lower non-specific binding. Modified antibodies also were found to have lower isoelectric points compared to unmodified controls. In vivo, modified antibodies unexpectedly were found to have 2-6 times faster clearance in tumor-bearing nude mice similar to rates obtained with their F(ab')2 fragments. Paired-label in vivo biodistribution and external imaging experiments with intact antibodies and F(ab')2 fragments demonstrated that chemically modified antibodies gave 1.5-3 fold higher tumor uptake and retained less activity in normal organs thus markedly increasing the tumor to normal organ ratios. Because of these results, chemically modified antibodies produced clearer images at earlier time points by external scintigraphy. As "stealth" molecules, chemically modified monoclonal antibodies appear to have significantly improved uptake in tumors and faster clearance times compared to native molecules. These results suggest that alteration of the physicochemical properties of monoclonal antibodies may generate improved reagents for in vivo use. PMID- 10851540 TI - Optimization of radioiodination and biotinylation of monoclonal antibody chimeric BR96: an indirect labeling using N-succinimidyl-3-(tri-n-butylstannyl)benzoate conjugate. AB - Chimeric BR96 (chiBR96) is an oxidant sensitive, highly tumor-reactive and internalizing monoclonal antibody. Radioiodination of chiBR96 with direct labeling methods has a high risk of damaging the antibody's immunoreactivity. Combination of iodination and biotinylation of chiBR96 has been particularly difficult. In present studies, indirect radioiodine labeling by use of a conjugate of N-succinimidyl 3-(tri-n-butylstannyl)benzoate (ATE) was introduced for chiBR96 radioiodination and for combination of iodination and biotinylation of chiBR96. ATE was synthesised, radioiodinated with a modified and simplified method by omitting one step of separation. A same or slightly higher labeling efficiency was obtained comparing to earlier reports. Simultaneous biotinylation of the chiBR96 was performed by use of biotin reagent of N-hydroxysuccinimido biotin. In a comparative study of biotinylated and unbiotinylated 125I-chiBR96 iodinated with ATE conjugate, it was found that 125I-chiBR96 with or without biotinylation had the same stability, immunoreactivity and biodistribution with a high tumor targeting capacity. Hence, the ATE conjugate radioiodination method enables the combination of iodination and simultaneous biotinylation of chiBR96. PMID- 10851541 TI - FDA reform-real or just more politics? PMID- 10851542 TI - Alpha interferon-2b, leucovorin, and 5-fluorouracil (ALF) in non-small cell lung cancer. AB - Eighteen patients with non-small cell lung cancer (NSCLC) who had previously received no systemic chemotherapy, were treated on this phase II study with interferon-alfa-2b 8 MU tiw, leucovorin 500 mg/m2 IVPB over 2 hours and 5 Fluorouracil 500 mg/m2 i.v. for 6 weeks followed by a 2-week rest. There were was no rest period for interferon. Median age of patients on this study was 63 years. Fatigue, nausea, and diarrhea were the most common toxicities. There were no grade IV toxicities and no treatment-related deaths. Seven partial responses (39%) with median duration of 4.5 months were seen. An additional patient has had stable disease/minor response for 12+ months. Median survival is 10 months. ALF has activity in NSCLC. PMID- 10851543 TI - Positron emission tomography using [18F]fluorotamoxifen to evaluate therapeutic responses in patients with breast cancer: preliminary study. AB - Positron emission tomography (PET) was used to assess the biodistribution and clinical usefulness of [18F]fluorotamoxifen (FTX) in 10 patients with estrogen receptor(ER)-positive breast tumors. Ten patients with ER-positive breast cancer were prospectively studied, and the consecutive PET imagings (each takes 15 or 20 min) were obtained for 60 or 80 min after the injection of 88.8-392.2 MBq (2.4 10.6 mCi) of [18F]FTX. Twenty three suspected primary or metastatic lesions in 10 patients were evaluated and the tumor uptakes of [18F]FTX in nineteen tumor lesions were correlated to the response of tamoxifen therapy. Three lesions in three patients were considered to be truly negative for breast cancer on the bases of biopsy specimens and/or clinical course. Five (71.4%) of seven patients and 16 (80.0%) of 20 lesions were interpreted to be truly positive for breast cancer. The mean standardized uptake value (SUV) of the radiotracer in tumor was 3.0 on delayed images. There was no significant correlation between the standardized uptake values of [18F]FTX and the ER concentrations in primary lesions. Nineteen tumor lesions in six patients were evaluable to compare the [18F]FTX uptake with responses to tamoxifen therapy after the PET study. Three patients who had a good response to tamoxifen therapy showed positive lesions on PET images, whereas two of three patients who had a poor response showed negative lesions and one showed mixed results. There was no significant difference of [18F]FTX uptake in bone lesions between good and poor responders. However, when bone lesions were excluded, [18F]FTX uptakes in tumors with good responses were significantly higher than those with poor responses (mean and standard deviation of SUV: 2.46 +/- 0.62 vs 1.37 +/- 0.59, P < 0.05). PET imaging using [18F]FTX provides useful information in predicting the effect of tamoxifen therapy in patients with ER-positive breast cancer. Further study is warranted to confirm the clinical utility of PET using [18F]FTX in breast cancer patients. PMID- 10851544 TI - Influence of OH group and sugar bonded to flavonoids on flavonoid-mediated suppression of tumor growth in vitro. AB - Many flavonoids extracted from higher plants, synthesized or purified flavonoids, and their aglycones showed anti-tumor activity. Using purified or synthesized flavonoids, we studied the effect of the chemical structure of 2-benzoic flavone on the growth of human tumor cells (HCT-15 in vitro. The type of sugar combined with the A phenol of flavonoids played an important role in the tumor suppression; i.e., glucose attachment at the A phenol caused suppression of tumor cell growth, but other sugars such as rhamnose and lutinoside at that position did not suppress the growth of the cells. OH groups bonded to the B phenol also had a great effect on the growth. Flavonoids with OH groups conjugated to the 3', 4', and 5' of B phenol were stronger in anti-tumor effect than those with the OH groups attached at the 3' and 4' or 4' only, although anthocyanins were generally more effective than the other flavonoids. PMID- 10851545 TI - Tumor necrosis factor-alpha suppresses the regrowth of fractionated irradiated endothelial cells in vitro. AB - Cytokines from two sources may affect endothelial cells (ECs) in tumor therapy: endogenously from cells in tumors and exogenously from therapeutic applications. These cytokines could modulate the influence of other therapy on tumor ECs. We use the colorimetric MTT method to assess the growth of irradiated ECs isolated from bovine pulmonary artery (CPAEC) and human umbilical cord vein (HUVEC) treated by cytokines. CPAECs given a single radiation dose of 2.5 to 15 Gy showed a small reduction in viable cells 2 to 3 days post-treatment. Neither tumor necrosis factor-alpha (TNF), interleukin-1 alpha (IL-1), nor interferon-gamma (INF) altered the growth of CPAECs treated by single radiation doses. HUVECs irradiated by a single dose of 12 Gy showed continuous reduction in viable cell numbers while those treated by 3 fractions of 4 Gy in 3 days or 6 fractions of 2 Gy in 3 days began to regrow 7 to 8 days after irradiation. Addition of TNF during the fractionated irradiation period limits the regrowth of HUVECs. Addition of IL-1 does not have the same effect. We have also tested the combined effect of another EC active agent flavone acetic acid (FAA), which has also been shown to stimulate the expression of TNF, with radiation, FAA (200 micrograms/ml) has a greater inhibitory effect on the growth and regrowth of fractionatedly irradiated HUVECs than TNF. These data suggest that TNF or FAA should be explored along with radiotherapy for their anti-tumor effect. PMID- 10851546 TI - Adsorptive or by pit formation endocytosis of immunoglobulins without loss of function as potential biotherapeutical application. AB - We here describe an alternative way to microinjection by which cellular transport of immunoglobulins through surface membranes can be achieved after binding to specific surface receptors either induced or constutively present, or via Fc receptors (Ig-mediated). In this report, the internalisation of two antibodies in two different cellular systems is analysed: the anti-p21ras monoclonal antibody (MoAb) after surface Ig binding on murine placental cells and anti-cdc2 MoAb that binds directly to its surface receptor expressed on the human promyelocytic leukemia cell line HL-60. In both cases, binding and internalisation is followed by Electron Microscopy (EM) and function is assessed by different assays. The first involves abrogation of class II antigen expression induced by Interferon gamma (IFN-gamma) and 5-Azacytidine (5-AzaC) known to be mediated by activation of the ras pathway. The second involves growth cessation of HL-60 cells after antibody adsorption when a G1-S-specific culture supernatant containing anti-cdc2 activity is employed, whereas no growth hindrance is observed when a G2-M specific anti-cdc2 MoAb is used. Thus, the antibodies do not follow the lysosyme pathway and do not lose their functional activity. This method may be applied in the future in order to achieve biological or clinical therapies. PMID- 10851547 TI - Protective effect of ginseng on radiation-induced DNA double strand breaks and repair in murine lymphocytes. AB - We have examined the effects of ginseng on the induction and repair of gamma-ray induced DNA double strand breaks (dsb) using neutral filter elution technique at pH 9.6 in cultured murine spleen lymphocytes. Ginseng water extract 500 micrograms/ml was added to the culture medium either for 48 hours prior to irradiation. Ginseng extract showed protective effect against the formation of dsb when it was treated for 48 hours before 100 Gy gamma-ray-irradiation. While repair was almost completed until 220.2 minutes after irradiation, DNA repair of irradiated cells in the presence of ginseng extract was did not return to the corresponding control levels even after 621.8 minutes. From these data, it could be calculated that ginseng reduced the relative strand scission factor (RSSF) by about 2. Therefore, it could be concluded that ginseng has radioprotective effect against gamma-ray induced DNA dsb and repair in cultured mouse lymphocytes. PMID- 10851548 TI - Antitumor effect of hydrolyzed anthocyanin from grape rinds and red rice. AB - When Balb/C mice that were fed with red glutinous rice, white ordinary rice, or commercially available standard food were inoculated with syngeneic Meth/A lymphoma cells i.p., the group fed the red rice survived longer than the other two groups. In order to determine if the anthocyanins contained in red-coloured seeds and fruit rinds were responsible for the tumor suppressive effect, we added anthocyanins extracted from grape rinds and glutinous red rice to petri dishes that had been seeded with HCT-15 cells. After 4 days of culture, cell counts were made. These anthocyanins were not effective in suppressing the tumor growth. However, anthocyanidins, which were generated by keeping anthocyanins in 5 to 6% HC1 methanol for 5 to 6 hours, were effective in the suppression of tumor growth. Flowcytometric histograms were made after 2 days of culture with these bioflavonoids. The histogram of cells treated with crude anthocyanin was similar to that of the control. Hydrolyzed anthocyanidins gave the elevation of S phase, suggesting a block in the step from S-phase to G2-phase. It seemed that the anthocyanidins contained in the grape rinds and red rice were effective on the suppression of cell growth as observed previously for anthocyanins extracted from the petals of higher plants. PMID- 10851549 TI - [Antiretroviral therapy and immune reconstitution]. AB - The course of the HIV infection has been dramatically modified since the introduction of highly active antiretroviral therapy (HAART) combining inhibitors of the HIV-1 reverse transcriptase and protease. Despite some controversies about the extent to which the immune system can normalize, it is generally admitted nowadays that a numerical and functional CD4 cell profile more akin to asymptomatic HIV-infected individuals can be restored in AIDS patients and can confer host protection against opportunistic events. The best hallmark of such immune restoration is the massive decline in the mortality and morbidity related to AIDS that have been registered in all industrialized countries. These changes involve a recirculation of mature peripheral T cells, a regeneration of naive T cells from thymic origin and a restoration of memory CD4 T cell reactivities. Although these recent advances warrant increased optimism, HAART by reducing the virus burden is unable to restore immunity against HIV itself, except when introduced at the very early stage after virus inoculation. The single condition required for immune reconstitution is an efficient and durable inhibition of virus replication. These positive effects can be obtained at late stages of the disease even when the patients have been heavily pre-treated. They also demonstrate that HIV does not definitively alter the lymphoid tissues nor the immune defenses, even aller years of infection and severe immune suppression, except for HIV-specific CD4 T helper cells. PMID- 10851550 TI - [Deficiency of the CD3-TCR signal pathway in three patients with idiopathic CD4+ lymphocytopenia]. AB - Idiopathic CD4+ lymphocytopenia (ICL) is a rare syndrome affecting adults and defined by a stable loss of CD4+ T cells in the absence of any known cause of immune deficiency. Defective T-cell proliferations to mitogens and antigens have been described in some of such patients displaying clinical signs of immune deficiency such as opportunistic infections. We investigated here the hypothesis that T-cell depletion and dysfunction could be due to biochemical defects of the CD3-TCR pathway in CD4+ and/or CD8+ subsets from three patients with severe stable ICL (below 150 CD4+ T cells/microliter) and opportunistic infections. Patient 1 had a general T lymphocytopenia, whereas patients 2 and 3 displayed a selective loss of CD4+ T cells. We observed in all patients a major reduction of the proliferative response to CD3-TCR stimulation that affected only the depleted T-cell subpopulation. Moreover, in two cases, impaired early biochemical events of the CD3-TCR pathway were detected. In patient 1 and 3, we found a defect (of distinct intensity) of CD3-induced protein tyrosine phosphorylation in CD4+ cells compared to control cells, whereas this process was normally induced in CD4+ T cells from patient 2. Taken together, this study reveals that the heterogeneity of the ICL syndrome was situated at the cellular level, and involved in two cases abnormalities of transducing molecules of the CD3-TCR pathway. PMID- 10851551 TI - [The synaptonemal complex: a structure necessary for pairing, recombination or organization of the meiotic chromosome?]. AB - The synaptonemal complex (SC) is a prominent and evolutionaly well conserved structure which is strictly meiotic. Several evidences from mutant phenotypes support the hypothesis that recombination and SC formation are mutually interdependent processes. Moreover, the SC recombination nodules correspond in number and location to the crossing-over events. However, recent data confirm that SC formation does not require initiation of recombination, and several observations indicate that full synapsis is not required for recombination. The potential roles played by the SC will be discussed in the following framework: First, although not required for homology recognition, the SC could promote interhomolog interactions in situations where the normal processes have failed (interlocking, heterologous pairing, etc.); Second, polymerization of the SC components might permit the recombination process to progress by modulating the number and localisation of reciprocal versus nonreciprocal exchanges (i.e. interference) and; Third, the SC may play an important role in meiotic chromosome structure and especially in inter-sister interactions. PMID- 10851552 TI - [Mechanisms and control of meiotic recombination in the yeast Saccharomyces cerevisiae]. AB - Recent studies in Saccharomyces cerevisiae have provided new insights in our understanding of the molecular mechanisms of meiotic recombination. Meiosis specific DNA double-strand breaks have been detected and have been shown to be the lesions that initiate recombination events. These are located mostly in promoter regions where the chromatin is in an open configuration, and cluster in domains along the chromosome. They are likely to be made by a topoisomerase II like protein encoded by the SPO11 gene. Several DNA intermediates in the meiotic double strand-break repair pathway have been characterised and several multi protein complexes have been identified and shown to be involved at different steps in the repair pathway. The conservation of these protein complexes in higher eukaryotes suggests that the meiotic recombination mechanism could be conserved. With the application of the well characterised genetical, molecular, cytological and biochemical techniques and the recently developed technology for genomic studies (biochips), we can expect a rapid increase in our comprehension of the meiotic recombination process. PMID- 10851553 TI - [DNA repeats and homologous recombination: a probable role for DNA methylation in genome stability of eukaryotic cells]. AB - Homologous recombination between DNA repeats directly threatens the intact transmission of repeat-rich eukaryotic genomes through mitotic and meiotic cell divisions. Besides several other factors already known, DNA methylation might contribute, in some eukaryotes, to the limitation of crossover events between repeats. A strong inhibitory effect of DNA methylation has now been directly demonstrated, in the filamentous fungus Ascobolus. This therefore reinforces the question of the biological impact of this DNA modification on the recombinational stability of repeat-rich genomes, such as those of mammals. PMID- 10851554 TI - [Structure and evolution of human sub-telomeric regions]. AB - Recent progress in the field of human genome analysis has led to the development of new concepts in the definition of subtelomeric domains. Analysis of DNA sequences from human and yeast chromosome ends have shown that short stretches of degenerate TTAGGG are found at a distance from the telomeric repeats. These stretches define a boundary between two structurally different regions. The distal domain is characterised by numerous, short segments of interrupted homology to many other human telomeric regions and to a number of ESTs. The proximal domain shows much longer uninterrupted homology to a few chromosome ends. This domain evolved quickly within primates at least, as demonstrated by the detailed study of locus DNF92 which spread very recently in humans from 17 qter to at least ten other chromosome ends. At the different sites, presence absence polymorphisms are observed within humans. The region remained single locus at the paralogous site in higher primates. Conversely, a human and orangutan single locus telomeric domain occupies multiple chromosome ends in chimpanzee. Balanced translocation is the likely mechanism through which the spreading occurred. Some members of the olfactory receptor gene family show a similar behaviour: multiple telomeric locations, and presence-absence polymorphism. Strikingly, the set of chromosome ends occupied by the two regions is identical, except for the two ancestral sites. Moreover, the relative frequency of detection of the region at the different sites indicates some kind of competition between the two regions. Consequently, these two regions represent major new tools to investigate recent human genome evolution and human genome diversity in different populations. PMID- 10851555 TI - [Fibrocytes and activated fibrocytes in the healing of an open skin wound]. AB - The microscopic, electron microscopic and immunohistochemical observation of biopsy specimens taken at an early stage, at close and regular intervals (every 4 hours), from open skin wounds created in the pig and the monkey, together with quantitative analysis of the various cell types in the granulation tissue, supports the conception that the activated fibrocyte (fibroblast) originates from the fibrocyte of the wound edges and thus completes some earlier experimental studies. We describe here the various stages of the differentiation of the wound edge fibrocyte into an activated fibrocyte and its proliferation and migration from the edges to the site of the wound. This does not exclude the possibility that local mesenchymal cells take part in the formation of activated fibrocytes. The activated fibrocyte build the collagen of the granulation tissue and then remodel and ensure wound contraction by becoming fibroclasts and myofibroblasts. This article defines the signification of the terms fibrocyte, activated fibrocyte, fibroblast and activated fibroblast. PMID- 10851556 TI - [Pharmacologic markers predictive of neutropenia chemically induced by cisplatin (CDDP)]. AB - The aim of this study was to demonstrate the value of the level of platinum in lymphocytes, plasma and saliva, in order to predict neutropenia during the same cycle after cisplatin chemotherapy. Plasma and lymphocytes samples were obtained from 14 patients receiving 100 mg/m2 cisplatin in different combination. Saliva samples were obtained from 3 other patients. We found that platinum plasma concentration at the Cmax and 1 hour after the end of the infusion were significantly higher in the grade 4 neutropenia cycles (respectively 2.60 vs 2.05 and 2.55 vs 2.00 mg/l p < 0.05). Platinum DNA-adduct were not detectable by ICPMS. Maximum concentrations in saliva ranged between 0.05 to 0.08 mg/l at the end of the infusion. The ratio of platinum levels in plasma and saliva varied between 2 and 3%. Saliva seems to be useful for therapeutic drug monitoring of cisplatin because it can be obtained by non invasive and patient-friendly-means. However, new studies are required to demonstrate the relationship between these two biological fluids. PMID- 10851557 TI - [Involvement of P2X and P2U receptors in the constrictor effect of ATP on the pancreatic vascular bed]. AB - The effects of ATP on the pancreatic vascular bed were studied on the isolated rat pancreas perfused at a constant pressure so as any change in the vascular tone induces a modification in the flow rate. This study was performed in two different experimental conditions: 1) In the presence of indomethacin, inhibiting the cyclo-oxygenase and prostacyclin (PGI2) synthesis, ATP (which acts on vasodilatator P2Y receptors and vasoconstrictor P2X and P2U receptors) was used at a concentration (165 microM) which did not modify per se the vascular flow rate. With indomethacin, ATP induced a slight but significant and long lasting decrease in the flow rate. This effect is different from that induced by the stimulation of P2X receptors; it is comparable to that induced by the activation of P2U receptors. 2) In the presence of 2,2'pyridylisatogen tosilate (PIT) used at two different concentrations, the first (5 microM) inhibiting the P2Y effects on insulin secreting B cells and pancreatic vessels, the second (25 microM) inhibiting the P2X effects on pancreatic vessels. The effects of ATP are different according to the concentration of PIT. In both cases, ATP induced only a vasoconstriction. However, the kinetics of the flow rate is totally different: in the presence of 5 microM PIT, an immediate and drastic vasoconstriction was observed, followed by a long lasting vasoconstriction of lesser magnitude, which can be ascribed to P2X and P2U receptor activation, respectively. This hypothesis was confirmed by the results in the presence of PIT at 25 microM. At this concentration this compound completely suppressed the drastic and transient vasoconstriction, so that only a progressive and long lasting vasoconstriction of the P2U type could be observed. From these results, it can be concluded that: 1) PGI2 plays a part in the vasodilatator effects of ATP. 2) At the concentrations used, PIT does not block the vasoconstriction induced by P2U receptors. 3) The effects of ATP on pancreatic vessels is dependent on the balance between its vasodilator effect due to the activation of P2Y receptors and its vasoconstrictor effect which involves two types of receptors: P2X and P2U. PMID- 10851558 TI - [Molecular mechanism of action of the fibrates]. AB - Fibrates are old hypolipidemic drugs with pleitropic effects on lipid metabolism. Until, recently their intimate molecular mechanisms of action were mysterious. In the late 5 years, we have shown that the pharmacological effects of fibrates depend on their binding to "Peroxisome Proliferator Activated Receptor alpha" (PPAR alpha). The binding of fibrates to PPAR alpha induces the activation or the inhibition of multiple genes involved in lipid metabolism through the binding of the activated PPAR alpha to "Peroxisome Proliferator Response Element" (PPRE) located in the gene promoters. Fibrates reduce plasma triglyceride levels by altering the expression of numerous genes coding for proteins involved in fatty acid metabolism (fatty acid transport protein, acyl-CoA synthetase, etc.) and also by increasing the lipoprotein lipase synthesis and decreasing the apolipoprotein C-III synthesis. Fibrates increase HDL cholesterol levels by increasing apolipoprotein A-I and apolipoprotein A-II synthesis. Furthermore, we recently demonstrated that fibrates are potent anti-inflammatory molecules through an indirect modulation of the nuclear-factor-kappa B activity. Therefore, we suggest that fibrates inhibit atherosclerosis development not only by improving the plasma lipid profile but also by reducing inflammation in the vascular wall. PMID- 10851559 TI - [Cell receptors for human adenoviruses]. AB - During the early stage of the adenovirus infection, the virion binds to a "primary receptor" on the host cell plasma membrane via the fibre projection jetting out of the penton base capsomers located at the twelve apices of the icosahedral capsid. The second step consists of a receptor-mediated endocytosis which involves membrane integrin molecules (the "secondary receptors") and the RGD and/or LDV motifs of penton base. The latter step is inhibited at low temperature, whereas virus attachment to its primary receptor is temperature independent. Two different primary receptors with a high affinity for the Adenovirus have been recently identified. One is common to Coxsackievirus B3 and adenovirus (CAR), the other one corresponds to a conserved region of the alpha-2 domain of the heavy chain of the major histocompatibility complex class I molecules (MHC-I-alpha 2), overlapping tryptophane-167. The receptor usage by the virus is governed by both cellular and viral parameters. On the cellular side, the relative abundance of one versus the other type of primary receptors would theoretically determine the virus choice: CAR receptor has been mainly found in tissues from mesodermic origin, whereas MHC-I-alpha 2 is ubiquitous. On the virus side, the molecular determinants of the receptor usage have been mapped to the terminal knob of the fiber projection, and have been found to be different for CAR and MHC-I-alpha 2. CAR recognizes linear motifs in fiber knobs in a subgroup dependent manner, as it binds to all Adenovirus serotypes except for the subgroup B members. MHC-I-alpha 2 however recognizes conformational epitopes carried by fiber knobs from all serotypes tested including subgroup B members. These results should have significant implications in the cell targeting of adenoviral vectors used in gene therapy. PMID- 10851560 TI - [MUC genes: a superfamily of genes? Towards a functional classification of human apomucins]. AB - The MUC genes encode epithelial mucins. Eight different human genes have been well characterized, and two others identified more recently. Among them, a family of four genes, expressed in the respiratory and digestive tracts, is clustered to chromosome 11p15.5; and these genes encode gel-forming mucins which are structurally related to the superfamily of cystine-knot growth factors. A second group is composed of three independent genes encoding various isoforms of mucins including membrane-bound mucins associated to carcinomas. In this second group, MUC3 and MUC4 encode large apomucins containing EGF-like domains. PMID- 10851561 TI - ["Glyco-deglyco" processes during the biosynthesis of glycoproteins]. AB - For the past fifteen years, it has appeared increasingly evident that the N glycosylation process was accompanied by the release of oligomannoside type oligosaccharides. This material is constituted of oligosaccharide-phosphates and of neutral oligosaccharides possessing one GlcNAc (OS-Gn1) or two GlcNAc (OS-Gn2) at the reducing end. It has been demonstrated that oligosaccharide-phosphates originated from the cleavage, by a specific pyrophosphatase, of non-glucosylated cytosolic faced oligosaccharide-PP-Dol and chiefly the Man5GlcNAc2-PP-Dol. The Man5GlcNAc2-P, as the main product, is recovered in the cytosolic compartment and is further degraded to Man5GlcNAc1 by not-yet depicted enzymes. In contrast, OS Gn2 produced from hydrolysis of oligosaccharide-PP-Dol (presumably as a transfer reaction onto water) when the amount of protein acceptor is limiting, are generated into the lumen of rough endoplasmic reticulum (rough ER). They are further submitted to processing a-glucosidases and rough ER mannosidase and are (mainly as Man8GlcNAc2) exported into the cytosolic compartment. This material is further degraded into a single component, the Man5GlcNAc1, by the sequential action of a cytosolic neutral chitobiase followed by cytosolic mannosidase(s). Furthermore, OS-Gn1 could have a dual origin: in one hand, they originate from OS Gn2 by the cytosolic degradation pathway indicated above, on the other hand, we will discuss a possible origin from the degradation or remodelling of newly synthesized glycoproteins. Considered first as a minor phenomenon, these observations have lead to the concept of intracellular oligomannoside trafficking, a process which results from more fundamental phenomena such as the control of the dolichol cycle, and the so-called quality-control of glycoprotein. In this review, we would like to describe the evolution of ideas on the origin, intracellular trafficking and putative roles of these oligomannosides released during during the N-glycosylation process. We propose that these early stage "glyco-deglyco" processes represent a way of control of N-glycosylation and of the fate of N-glycoproteins. This review is dedicated to Pr Paul Boulanger who has spent a large part of his career to determine the structure of proteins in order to understand their functions. If it is well established that many biological functions are born by proteins, it appears more and more evident that co- or post translational modifications are of importance in the modulation of these functions. Among them, the glycosylation appears as a major event which intervene in the 3D structure of the protein, which control his biological time life and which may act in many recognition processes. PMID- 10851562 TI - Edmund Klein, M.D. (1921-1999). PMID- 10851563 TI - Thromboembolic disease susceptibility related to red cell membrane fluidity in patients with polycythemia vera and effect of phlebotomies. AB - Thromboembolic disorders are frequent complications in polycythemia vera. In addition to thrombocytosis with hyperaggregability, leukocytosis, and high hematocrit, hyperviscosity syndrome, a new component, is described in the pathophysiology of this phenomenon. There is decreased red cell membrane fluidity with decreased deformability which increases the susceptibility to microvascular occlusion and also increases the chance of disseminated intravascular coagulation (DIC). Periodic phlebotomies improved the hematologic picture in these patients and results in the removal of the "stiff" red cells with an increased production of young red cells, greater membrane fluidity, deformability and less chance of microvascular occlusion. PMID- 10851564 TI - The effects of a low-dose regimen of fosinopril on elevated urinary albumin excretion in normotensive type 1 diabetic patients. AB - Normotensive type 1 diabetics with persistent albuminuria were randomized between 12 months' treatment with low-dose fosinopril (10 mg/d) and placebo in a 24 month, double-blind, crossover trial. Metabolic and cardiovascular variables were measured every three months and two-dimensional and M-mode echocardiography was performed before and after each treatment phase. Low-dose fosinopril led to 70% reduction in urinary albumin excretion (UAE) (220 +/- 199 vs 82 +/- 67 mg/24-h, p < 0.05), but UAE did not change during placebo treatment; the maximal change became apparent after 3 months and was reversible once treatment was stopped. Even though blood pressure (BP) did not change during fosinopril treatment, there was a significant correlation between baseline UAE and diastolic BP (r = 0.77, p = 0.027) and between change in UAE and change in systolic BP (r = 0.84, p = 0.01). The change in UAE approached borderline significance in correlation with the change in triglyceride concentration (r = 0.66, p = 0.07). Low-dose fosinopril favorably affects the "cardiorenal syndrome" seen in type 1 diabetics with persistent microalbuminuria. PMID- 10851565 TI - Double filtration plasma exchange and immunoadsorption therapy in a case of stiff man syndrome with negative anti-GAD antibody. AB - We report the effects of double filtration plasma exchange and immunoadsorption therapy which were performed for a case of stiff-man syndrome even though the patient was negative for anti-glutamic acid decarboxylase (GAD) antibody. The patient underwent a course of four double filtration plasma exchanges, which resulted in marked clinical improvement. Painful muscle cramps disappeared and muscle stiffness reduced within a day after the first plasma exchange. The patient's improvement continued, but his condition declined again about ten months after plasma exchange. Immunoadsorption therapy was then performed, and this treatment was also effective. PMID- 10851566 TI - N-acetyl-beta-D-glucosaminidase is not a predictor, but an indicator of kidney injury in patients with cardiac surgery. AB - The urinary excretion of N-acetyl-beta-D-glucosaminidase (NAG) was evaluated in terms of its ability to predict renal failure following open-heart surgery. Twenty-two patients were divided into three groups; 6 patients with post operative renal failure (RF group), 9 without renal failure (non-RF group) and 7 who underwent non-cardiac surgery (non-ECC group). NAG levels during operation were significantly higher in the non-RF group than in the non-ECC group (2.28 +/- 0.50 U vs. 0.39 +/- 0.06 U, p < 0.01). NAG levels in the RF group were not significantly different from those in the non-ECC group (1.12 +/- 0.28 U/hr vs. 0.39 +/- 0.06 U/hr, p > 0.1), and in the non-RF group (1.12 +/- 0.28 U/hr vs. 2.28 +/- 0.50 U/hr, p > 0.1). Post-operative NAG levels were not significantly different between the RF and non-RF groups, whereas post-operative creatinine clearance values were significantly different (on 2, 6 and 7 post-operative day, p < 0.05). Our data suggest that the kidney is predisposed to renal failure by cardiopulmonary bypass and the chance of renal failure depends in some part on post-operative patient management. PMID- 10851567 TI - Characterization of psoriasis in vivo by reflectance confocal microscopy. AB - Among available non-invasive imaging tools, confocal reflectance microscopy (CM) provides the highest-resolution optical sectioning of skin in vivo, to a controlled depth of 200-350 microns, the level of the upper reticular dermis. In this study, CM was used to view the histological features of psoriasis in vivo in lesional and non-lesional skin of five patients with stable psoriasis vulgaris. Stereohistological analysis of non-invasive confocal sections, and the correlation with transverse (en face) hematoxylin- and eosin-stained sections from biopsies, was also performed. In psoriatic lesions, nucleated corneocytes and collections of infiltrating inflammatory cells were clearly seen. Morphometric parameters such as epidermal height, length of papillary dermis, and the count of dermal papillae were also easily quantified. In the upper dermis, dilated capillary loops were always present. Since CM sections are en face, the presence or absence of the granular layer could not be visualized in single frames, but could be monitored in a sequence of real-time videotaped images. In summary, CM provides a new technique for histologically evaluating psoriasis in vivo. PMID- 10851568 TI - An acquired factor V inhibitor: clinical and laboratory features. AB - A sixty-five year old white male presented with an acquired Factor V inhibitor after an episode of cholecystitis and cefotaxime therapy. Plasma Factor V activity was less than 1%. He developed lower gastrointestinal bleeding a week after onset of coagulopathy, and was treated with plasmapheresis, fresh frozen plasma, oral cyclophosphamide, and prednisone. The coagulopathy resolved within four days of treatment, and within two weeks of presentation. Laboratory studies revealed an IgG inhibitor to Factor V that closely mimicked the more commonly encountered lupus anticoagulant. We would like to alert clinicians to this entity because, in contrast to a lupus anticoagulant, the acquired Factor V inhibitor can be associated with clinical bleeding as in our patient, and requires therapy prior to any surgical procedures. PMID- 10851569 TI - Bone mineral density (BMD) in obesity effect of weight loss. AB - It is generally believed that bone mineral density (BMD) is increased in obese subjects, but the effect of weight loss on BMD has not been well studied. Therefore, we evaluated BMD among 11 obese women (mean age 45.5 +/- 14.2 years) before and after weight loss achieved by ingesting an 800 calorie diet for 12 weeks. BMD measurements were made at baseline, 6 months and 1 year intervals. Urinary hydroxyproline:creatinine (H:Cr), calcium:creatinine (Ca:Cr) ratios were measured as indices of bone turnover. Mean weight at baseline was 103.8 +/- 15.8 kg and decreased to 83.2 +/- 12.2 at six months and was 85.8 +/- 14.2 kg at one year. Total body, hip and lumbar spine BMD were 1.12 +/- 0.07, .87 +/- 0.11, and 1.02 +/- 0.12 gm/cm2, respectively. Total body BMD was significantly lower at 12 months compared to baseline. No significant change was observed in BMD of the lumbar spine. There was also a significant decrease in hip BMD at six months and 12 months compared to baseline. H:Cr and Ca:Cr ratios did not change over time. We conclude that weight loss achieved by VLCD is accompanied by a statistically significant change in BMD, but the BMD remained in the normal range. PMID- 10851570 TI - Menstrual bleeding, hormones, and the menopausal transition. AB - The perimenopause represents a time of great variability in reproductive hormone dynamics and menstrual cycle characteristics, but age-related changes begin prior to this. These changes include a gradual increase in follicle stimulating hormone (FSH) levels, a gradual shortening of mean cycle length, and a decline in the number of ovarian follicles. The onset of perimenopause is thought to occur with the first break in menstrual cycle regularity. With the onset of cycle irregularities, hormone concentrations exhibit large increases in variability and unpredictability, rather than following a gradual trend with the approach of menopause, the final menstrual period. Abrupt spikes in gonadotropins and considerable fluctuations in estradiol and inhibin levels have been observed. Variability is the norm in the perimenopause, with hormonal fluctuations contributing to the visible signs of menstrual cycle and bleeding irregularities. To date there is no single endocrine indicator to serve as an adequate marker of menopausal status. This paper provides a review of research to date on patterns of reproductive hormones and menstrual bleeding during the menopausal transition. An understanding of such patterns can contribute to a better ability to distinguish "normal" transitional events from more serious pathology. PMID- 10851571 TI - Age-related changes in growth hormone secretion: should the somatopause be treated? AB - Growth hormone (GH) secretion declines progressively with aging, and many age related changes resemble those of the adult GH deficiency (GHD) syndrome, including a decrease in lean body mass; an increase in body fat, especially in the visceral/abdominal compartment; adverse changes in lipoproteins; and a reduction in aerobic capacity. The increase in central obesity can further inhibit GH secretion. GH replacement is effective in reversing many of these changes in adult GHD, and GH is now FDA approved for treatment of adults with documented GHD or hypopituitarism, although there is still only limited experience with its long-term benefits, side effects, and risks. This early experience with GHD has led to speculation that replacing GH or stimulating its secretion may also be beneficial in normal aging, and to widespread off-label use of GH in this context; however, there are still very few well controlled studies of the effects and side effects of GH or GH secretagogues in aging. All published studies are of 6 months or shorter treatment periods. From this limited experience there is a consensus that GH has effects on body composition, but reports disagree on effects on psychological or physical functional performance. Older adults are much more susceptible to the dose-related side effects of GH, including peripheral edema, carpal tunnel syndrome, and a variable decrease in insulin sensitivity; and it is not known whether chronic GH treatment affects the risk of malignancy or has other long-term risks. Thus while short-term results are somewhat encouraging, the evidence on risks and clinically pertinent benefits is still lacking to support the use of GH in normal aging outside of clinical studies. In evaluating patients with clinical features suggesting GHD, which can be quite nonspecific, it is important to assess the presence or absence of true GH deficiency by the context (pituitary disease or its treatment, childhood GHD) and by appropriate GH stimulation tests before considering GH replacement. PMID- 10851572 TI - Changes in adrenocortical function with aging and therapeutic implications. AB - Throughout life, the adrenal cortex exhibits dramatic morphogenic and steroidogenic changes. While there is subtle senescent decline in aldosterone, and a similarly subtle increase in cortisol, the adrenal androgens dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEAS) decline with age in a situation similar to menopause, and this decline is considered by some to aggravate some age-related diseases. This decline is associated with an almost complete loss of the inner zone of the adrenal cortex, known as the zona reticularis. This review addresses these adrenal cortical changes, and explores their clinical significance. In particular, the clinical data on DHEA replacement in aging is addressed. PMID- 10851573 TI - Reproductive hormones, aging, and sleep. AB - Insomnia, disturbed sleep, and fatigue are among the most frequent health complaints of perimenopausal women. Estrogen replacement therapy (ERT) usually improves sleep, most likely by alleviating vasomotor symptoms. However, sleep difficulties are not restricted to the perimenopausal period. Older postmenopausal women typically experience longer latencies to sleep onset, increased nocturnal waking, increased fragmentation of sleep, and less slow wave (deep) sleep. These sleep changes in older women may be partially related to the postmenopausal profile of sex steroid hormones. Estrogen has powerful effects on several biological factors that directly influence sleep, including body temperature regulation, circadian rhythms, and stress reactivity. The link between sleep disturbance in older women and these CNS effects of estrogen is largely speculative at present. This article reviews what is known, what remains to be addressed, and some clinical implications. PMID- 10851574 TI - Aromatase in aging women. AB - Cessation of ovarian estrogen secretion is the key event during the climacteric. An enzyme termed aromatase in a number of human tissues and cells, including ovarian granulosa cells, the placental syncytiotrophoblast, adipose and skin fibroblasts, bone, and the brain, catalyzes the conversion of C19 steroids to estrogens. Aromatase expression in adipose tissue and possibly the skin primarily accounts for the extraglandular (peripheral) formation of estrogen and increases as a function of body weight and advancing age. Sufficient circulating levels of the biologically active estrogen, estradiol, can be produced as a result of extraglandular aromatization of androstenedione to estrone, which is subsequently reduced to estradiol in peripheral tissues, to cause uterine bleeding and endometrial hyperplasia and cancer in obese anovulatory or postmenopausal women. Extraglandular aromatase expression in adipose tissue and skin (via increasing circulating levels of estradiol) and bone (via increasing local estrogen concentrations) is of paramount importance in slowing the rate of postmenopausal bone loss. Moreover, excessive or inappropriate aromatase expression was demonstrated in adipose fibroblasts surrounding a breast carcinoma, endometriosis derived stromal cells, and stromal cells in endometrial cancer and gave rise to increased local estrogen concentrations in these tissues. Whether systemically delivered or locally produced, elevated estrogen levels promote the growth of these steroid-responsive tissues. Finally, local estrogen biosynthesis by aromatase activity in the brain may be important in the regulation of various cognitive and hypothalamic functions. The regulation of aromatase expression in human cells via alternatively used promoters, which can be activated or inhibited by various hormones, increases the complexity of estrogen biosynthesis in the human body. Aromatase expression is under the control of the classically located proximal promoter II in the ovary and a far distal promoter I.1 (40 kb upstream of the translation initiation site) in the placenta. In adipose tissue, two other promoters (I.4 and I.3) located between I.1 and II are used in addition to the ovarian-type promoter II. To add a further twist, promoter use in adipose fibroblasts switches between promoters II/I.3 and I.4 upon treatment of these cells with prostaglandin E2 (PGE2) versus glucocorticoids plus cytokines. Moreover, the presence of a carcinoma in breast adipose tissue causes a switch of promoter use from I.4 to II/I.3. Molecular and cellular mechanisms responsible for estrogen formation and their physiologic and clinical relevance will be reviewed in this article. PMID- 10851575 TI - Endocrine cancer risks for women during the perimenopause and beyond. AB - Cancer and its link to reproductive hormones is an area of intense concern for our patients and has been the subject of much speculation. But if estrogen causes breast cancer, for example, most women would eventually develop the disease. We know this is not the case! Actually, estrogen and progesterone have been linked to a decrease as well as an increase in cancer, depending upon the type of tumor under investigation. The purpose of this manuscript is to review the data supporting those relationships. PMID- 10851576 TI - [Surgical treatment of anorectal fistulas. A 17-year experience at the Instituto Nacional de la Nutricion "Salvador Zubiran"]. AB - INTRODUCTION: Surgical treatment for anorectal fistula may be difficult because of the risk of recurrence, prolonged healing or anal incontinence following the operation. OBJECTIVE: To analyze the experience with the surgical management of ano-rectal fistula during a period of 17 years. PATIENTS AND METHODS: The medical records of 105 patients with anorectal fistulas were reviewed retrospectively, with analysis of demographic and clinical data, operative treatment, and results. RESULTS: There were 73% men and 27% women. Mean age was 45 years. 86% had an underlying chronic disease, most frequently diabetes mellitus (21%) and obesity (14%). No anatomic classification of the fistulous tract was done in 86% of cases, and inter-sphincteric tracts were the most frequent type in the classified cases. In 90% of cases, treatment was fistulectomy. Complications occurred in 13% of cases, mainly delayed healing (6.5%). Recurrent disease was documented in 11 cases (10%), and the majority were treated with a new fistulectomy. There were no cases with anal incontinence following the operation. CONCLUSIONS: The necessity of performing the anatomic classification of ano-rectal fistula should be emphasized. Fistulectomy was the most frequent surgical procedure. PMID- 10851578 TI - [Fistulotomy vs fistulectomy. Ultrasonographic evaluation of lesion of the anal sphincter function]. AB - OBJECTIVE: To determine the extension of the lesion implicated on the mechanism of the anal sphincter with endoanal ultrasound in patients with simple fistulae, managed with fistulotomy versus fistulectomy. SITE: Central Military Hospital. Colon and Rectum Service. DESIGN OF STUDY: A prospective, comparative, descriptive and longitudinal study was performed. METHODS: A total of 40 patients with anal simple fistula were studied from march 1997 to march 1998. They were divided in two randomized groups: group A (n = 20) patients treated with fistulectomy, and group B (n = 20) patients managed with fistulotomy. Endoanal ultrasound was practice at the time of the diagnosis and six weeks later to identify integrity of both internal and external anal sphincter, and to register them in separate form. RESULTS: There were no significant differences in sex and age distribution, nor in type of fistula. The average of internal anal sphincter lesion in inter-sphincteric fistulae treated with fistulotomy was 8.5 mm versus 9.08 with fistulectomy (p > 0.05). The average of internal and external anal sphincter lesion in trans-sphincteric fistulae managed with fistulotomy was 9.25 mm versus 11.38 with fistulectomy (p < 0.05). The global analysis showed that the average of the lesion in the sphincter, mechanism was larger in the fistulectomy versus fistulotomy (p < 0.05). CONCLUSION: The major muscular injury made to the sphincter mechanism is caused mainly by the fistulectomy in comparison with the conventional fistulotomy. PMID- 10851577 TI - [Dose-effect of the administration of ferrous fumarate in aged persons with iron deficiency]. AB - BACKGROUND: Anemia is the most prevalent hematological problem in elderly persons, affecting 14% of the males and 6% of the females of the population over 60 years of age in Mexico City. OBJECTIVE: To determine the effect produced by the prolonged administration of ferrous fumarate in elderly persons with iron deficiency. METHOD: In a population of 178 subjects, aged between 65 to 100 years, iron deficiency was diagnosed in 51 (28.6%), who had serum iron concentrations below 80 micrograms/dL for men and 60 micrograms/dL for women, but only 21 patients (11.8%), accepted to participated in the study. The response to a 6 months oral administration of ferrous fumarate were studied with a daily oral dose of 5 mg/kg of elemental iron. The patients were classified in 3 groups according to the abnormal parameters of iron metabolism (group 1 = 10.9% anemia, group 2 = 28.0% and group 3 = 63.0% anemia). RESULTS: The efficacy of treatment was evaluated by quantification of the changes occurred in serum iron concentrations, hemoglobin, ferritin and transferrin saturation index, at 0, 30, 90 and 180 days of treatment. This study showed that the treatment of oral ferrous fumarate in elderly patients with iron deficiency, produces a quantifiable improvement in measures of iron parameters within 6 months. CONCLUSIONS: The results of this study suggest the usefulness of prolonged treatment with ferrous fumarate in elderly patients with iron deficiency, to avoid therapeutic failure as a consequence of non-compliance as is common in elderly patients. PMID- 10851579 TI - [Experience with 14 cases of pancreatic pseudocyst]. AB - INTRODUCTION: Pseudocyst of the pancreas (PP) develops in 2 to 5% the cases of acute pancreatitis (AP). Most cases of PP regress spontaneously. PP has been misdiagnosed as a malignant pancreatic cyst neoplasm, reason why the patients are referred to specialized institutions. OBJECTIVE: To describe the cases of PP treated in a 15 year-period. MATERIAL AND METHODS: Review of clinical records of 14 cases treated from 1975 to 1989. RESULTS: There were 5 men (36%) and 9 women (64%) Mean age was 41 years (range 18 to 77). In 50% of the cases the patient had a history of severe alcoholic abuse, cholelithiasis in 28% and abdominal trauma in 15%. Five patients (35%) had AP. The period between AP symptoms and the diagnosis of PP was a mean of 5.7 months. Abdominal pain and abdominal mass were present in all of the cases, in all cases, ultrasonography and CT scan made the diagnosis of PP. Thirteen cases were treated by surgery, 12 with internal drainage, one by resection and one by external drainage. An enterocutaneous fistula (7%) was recorded in one case. There were no operative mortality. The mean follow-up time was of 10 years and 4 months. CONCLUSIONS: PP is a uncommon pathology in oncologic centers. Internal drainage was the most frequent treatment. The diagnosis of cystic neoplasms of the pancreas should be ruled out. PMID- 10851580 TI - [Long-term usefulness and late complications of percutaneous endoscopic gastrostomy]. AB - OBJECTIVE: To investigate the usefulness of endoscopic gastrostomy and long-term complications. BACKGROUND DATA: Endoscopic gastrostomy is well established as the procedure of choice for long-term feeding, given the low morbidity-mortality and ease of placement. METHOD: We evaluated retrospectively one hundred endoscopically placed gastrostomy feeding tubes and complications occurring more than 30 days after placement were recorded. RESULTS: Gastrostomy feeding tubes remained in place for a mean of 92 days (range 30-547 days). Fifteen percent developed evident gastroesophageal reflux, two patients developed aspiration pneumonia and one presented with infection at the site of gastrostomy. Our long term complications rate thus was 3.0% and 0% mortality. CONCLUSIONS: Our experience suggests that endoscopic gastrostomy is a relatively simple procedure, associated with very low morbidity and mortality. It is the procedure of choice in patients requiring long-term enteral nutrition. PMID- 10851582 TI - [Sodium diphenylhydantoin as a probable cause of pancreatitis]. AB - INTRODUCTION: Drug-induced pancreatitis is much more common in children than in adults. Many drug have been implicated in its pathogenesis. Among the neuropsychiatric drugs only valproic acid, carbamazepine, clozapine and ergotamine have been reported. Phenytoin is commonly used for the treatment of epilepsy. It has been associated to pancreatitis only in two previous reports. CASE REPORT: Male adolescent who initiated with cerebellar hemorrhage due to an arteriovenous malformation. During his evolution he presented the following complications: pneumonia, two urinary tract infection, gastrointestinal bleeding and arterial hypertension. Eighteen days after admission he developed seizures and was treated with phenytoin. The next day he presented pancreatic symptoms and pancreatitis was confirmed by elevated enzymes and a CAT scan with pancreatic edema. Other etiologies were discarded. Pancreatic enzymes persisted high until phenytoin was stopped and have been within normal values after 18 months of follow-up. CONCLUSIONS: In this case three of the four Miller criteria have been fulfilled. We consider that the antiepileptic treatment was the direct cause of the pancreatitis because there was a clear temporal association of the symptoms with the initiation and suspension of the drug. PMID- 10851581 TI - [Rectorrhagia as complication of Klippel-Trenaunay syndrome]. AB - OBJECTIVE: To describe the clinical presentation and treatment of two patients with the Klippel-Trenaunay syndrome referred to our hospital because of rectal bleeding and to review the literature concerning the diagnosis and treatment of this complication. CASE 1: Fifteen year old male with the Klippel-Trenaunay syndrome and chronic anemia who presented with severe rectal bleeding. CASE 2: Nineteen year old female with the same syndrome and a two year history of intermittent rectal bleeding, anemia and thrombocytopenia. In both cases the study protocol revealed varicose lesions in the colon as the cause of bleeding and other vascular malformations related to their syndrome. TREATMENT: The first patient was treated with partial colectomy and colorectal anastomosis. Four years after surgery he presented with new episodes of bleeding and was treated with sclerosis of the residual rectal varices using formaldehyde. The second patient was treated with partial colectomy and colostomy. She has received to sessions of sclerosis with absolute alcohol of the residual varices in the rectal stump. Colostomy closure is soon to be performed. CONCLUSION: Klippel-Trenaunay syndrome is a rare clinical entity with vascular alterations at different levels. A small percentage of cases may present rectal bleeding due to colonic varices and can lead to chronic anemia or severe hemorrhage with hemodynamic implications. Treatment of this complication involves resection of the affected colonic segment combined with a secondary procedure to control bleeding of the residual rectal varices. PMID- 10851583 TI - [Cutaneous metastasis of gastric cancer]. PMID- 10851584 TI - Statistical dynamical theory of X-ray diffraction in the Bragg case: application to triple-crystal diffractometry AB - The statistical dynamical theory of X-ray diffraction is developed for a crystal containing statistically distributed microdefects. Fourier-component equations for coherent and diffuse (incoherent) scattered waves have been obtained in the case of so-called triple-crystal diffractometry. New correlation lengths and areas are introduced for characterization of the scattered volume. PMID- 10851585 TI - Reconstruction of the projected crystal potential in transmission electron microscopy by means of a maximum-likelihood refinement algorithm AB - The projected crystal potential is reconstructed from a nonperiodic high resolution transmission electron microscopy exit wave function using a maximum likelihood refinement algorithm. The convergence and the accuracy of the algorithm are investigated using simulated exit wave functions of SiGe, a Shockley partial dislocation in Ge and an area containing randomly distributed Ge columns at different specimen thicknesses. The performance of two different start models for the projected crystal potential is investigated: the weak-phase-object model and a model based on the electron-channelling approximation. The reconstruction is successful even under the strongly nonlinear dynamical diffraction conditions at larger specimen thicknesses, relevant for high resolution work, and on specimen areas large enough to cover defects in crystalline materials. PMID- 10851586 TI - Discrete Fourier transform in arbitrary dimensions by a generalized Beevers Lipson algorithm AB - The Beevers-Lipson procedure was developed as an economical evaluation of Fourier maps in two- and three-dimensional space. Straightforward generalization of this procedure towards a transformation in n-dimensional space would lead to n nested loops over the n coordinates, respectively, and different computer code is required for each dimension. An algorithm is proposed based on the generalization of the Beevers-Lipson procedure towards transforms in n-dimensional space that contains the dimension as a variable and that results in a single piece of computer code for arbitrary dimensions. The computational complexity is found to scale as N log(N), where N is the number of pixels in the map, and it is independent of the dimension of the transform. This procedure will find applications in the evaluation of Fourier maps of quasicrystals and other aperiodic crystals, and in the maximum-entropy method for aperiodic crystals. PMID- 10851587 TI - Evaluation of net atomic charges and atomic and molecular electrostatic moments through topological analysis of the experimental charge density AB - The atoms in molecules (AIM) theory may be used to derive atomic charges, atomic volumes and molecular dipole moments from the charge density. The theory is applied to theoretical periodic Hartree-Fock (PHF), density-functional (DFT) and experimental X-ray densities of p-nitroaniline using the program TOPOND and a newly developed program, TOPXD, for topological analysis of densities described by the Coppens-Hansen multipole formalism. Results show that, like dipole moments derived directly from the multipole refinement, AIM-derived atomic and molecular moments are dependent on the multipole model used. As expected, large differences are found between charges derived from the monopole parameters and those from AIM analysis of the experimental model density. Differences between the k'-restricted multipole model (KRMM) and the unrestricted multipole model (UMM) results are preserved in the AIM analysis. The enhancement of the molecular dipole moment of p-nitroaniline in the solid state is confirmed by both experiment and theory but the experimental dipole moment is in much better agreement with theoretical periodic Hartree-Fock and, especially, periodic DFT (PDFT) data when KRMM is used in the refinement. The AIM analysis allows a rigorous definition of the charges of the atoms in molecules and provides a realistic basis for comparison between molecules and between experiment and theory. PMID- 10851588 TI - A finite group that derives all the 14 Bravais lattices as its subgroups AB - A finite group was discovered that includes all the types of Bravais lattice as its subgroups. It is based on a new representation of affine transformation of a primitive cell. Its elements are represented by 6 x 6 matrices whose components are complex in general. The order of the group is 2,799,360. PMID- 10851589 TI - Enantiomorph determination using inverse reference-beam diffraction images. AB - It is shown that enantiomorph structures of a noncentrosymmetric crystal can be determined, in the absence of anomalous diffraction signals, by measuring two series of reference-beam oscillation diffraction patterns related by an inverse beam geometry. The corresponding intensities of the Friedel pairs recorded on the two sets of images exhibit the characteristic three-beam interference effects that provide the unambiguous phase information. The experimental arrangement and the data-analysis procedure are demonstrated through an experimental example on tetragonal lysozyme. PMID- 10851590 TI - Triplet-phase measurements using reference-beam X-ray diffraction. AB - Reference-beam diffraction (RBD) is a recently developed phase-sensitive X-ray diffraction technique that incorporates the principle of multiple-beam diffraction into the standard oscillating-crystal data-collection method [Shen (1998). Phys. Rev. Lett. 80, 3268-3271]. Using this technique, a large number of multiple-beam interference profiles can be recorded simultaneously on an area detector, from which a large number of triplet phases of Bragg reflections can be determined in a crystallography experiment. In this article, both the theoretical developments and the experimental procedures of the RBD technique are described in detail. Approximate theoretical approaches for RBD are outlined and simple analytical expressions are obtained that provide the basis for an automated data analysis procedure that can be used to extract triplet phases from a large number of measured reference-beam diffraction profiles. Experimental examples are given for a variety of crystals including GaAs, tetragonal lysozyme and AlPdMn quasicrystal, using both image plates and a charge-coupled device (CCD) as the area detector. Possible uses of the measured phases for crystal structure determination are discussed as well as future prospects of the RBD technique. PMID- 10851591 TI - Shake-and-Bake applications using simulated reference-beam data for crambin. AB - The Shake-and-Bake method, as implemented in the computer program SnB, has been applied to simulated reference-beam data for the small protein crambin at several resolutions in the range 1.5-3.0 A. Sets of triplet invariants were generated having simulated mean triplet-phase errors from 0 to 60 degrees. Provided that these errors were no larger than 40 degrees, it was possible (at all resolutions tested) to find trial sets of individual Bragg phases with mean errors of 40-45 degrees. At 1.5 A, this could be achieved using only a single reference-beam data set. Peak picking provided useful phase constraints even at the lowest resolution tested. These results suggest that direct methods may be useful in conjunction with reference-beam data at resolutions lower than 1.2 A. PMID- 10851592 TI - On the extrapolation of the magnitudes magnitude of E of the normalized structure factors E. AB - For each fixed reciprocal-lattice vector H, the correlation coefficient, rho H, of the pair (magnitude of EK, magnitude of EH-K), where the magnitudes magnitude of EK and magnitude of EH-K are presumed to be known, is itself positively correlated with magnitude of EH. Thus, the correlation coefficients rho H serve to mediate the transfer of information from one region of reciprocal space to another. In particular, the calculated values of the correlation coefficients rho H lead to estimates of the unknown magnitudes magnitude of EH, even when the latter lie outside the sphere of the experimentally observed magnitude of E s (extrapolation). The applications show that the procedure described here to carry out this extrapolation is superior to existing methods. PMID- 10851593 TI - The NaCl- to CsCl-type phase transition discussed on the basis of the cP to cI deformation with the symmetry Cmcm 4(c) m2m AB - A structure forming a cubic primitive lattice cP may be deformed into a structure forming a cubic body-centred lattice cI in the space group Cmcm at position 4(c) m2m 0,y,1/4. If in related structures the sites are alternately occupied by unlike atoms, the NaCl and the CsCl types occur, respectively. The corresponding phase transition can be described as a deformation of a heterogeneous sphere packing in the subgroup Pmmn (a,-c,b) of Cmcm. All sphere configurations with symmetry Cmcm 4(c) m2m were derived. On the basis of this information, further possibilities for phase transitions that also correspond to sphere-packing deformations were found with this symmetry. Two of them possibly may take place in metals. The first one leads from a primitive hexagonal lattice to a hexagonal close packing, the other from a cubic body-centred lattice also to a hexagonal close packing. PMID- 10851594 TI - Systematic intensity errors and model imperfection as the consequence of spectral truncation AB - The wavelength dispersion delta lambda/lambda in a graphite (002) monochromated Mo K alpha beam was analyzed. A wavelength window was found with 0.68 < lambda < 0.79 A, i.e. delta lambda/lambda = 0.14. The very large dispersion leads to systematic errors in Iobserved(H) caused by scan-angle-induced spectral truncation. A limit on the scan angle during data collection is unavoidable, in order that an omega/2 theta measurement should not encompass neighboring reflections. The systematic intensity errors increase with the Bragg angle. Therefore they influence the refined X-ray structure by adding a truncational component to the temperature factor: B(X-ray) = B(true) + B(truncation). For an Mo tube at 50 kV, we find B(truncation) = 0.05 A2, whereas a value of 0.22 A2 applies to the same tube but operated at 25 kV. The values of B(truncation) are temperature independent. The model bias was verified via a series of experimental data collections on spherical crystals of nickel sulfate hexahydrate and ammonium hydrogen tartrate. Monochromatic reference structures were obtained via a synchrotron experiment and via a 'balanced' tube experiment. PMID- 10851595 TI - X-ray dynamical diffraction: the concept of a locally plane wave AB - The long distance between the source and the experiment and the small source size, now available at third-generation synchrotron sources, leads to new optical characteristics for X-ray diffraction. It is shown that, under certain conditions, the intensity received by a point located on the exit surface of a crystal is described by the diffraction of a locally plane wave. Each point along the surface is influenced by a plane wave with a varying departure from Bragg angle. In such a case, it is possible to visualize the rocking curve of the crystal as a function of the position along the exit surface. This represents a topographic method to obtain the reflectivity curve of the crystal instead of the usual goniometric method. Applications will be described in a forthcoming paper. PMID- 10851596 TI - Na2Co(H2PO4)4.4H2O PMID- 10851598 TI - Tetraammine(selenito-O,O')cobaltate(III) nitrate monohydrate PMID- 10851597 TI - KNi3(AsO4)(As2O7) PMID- 10851599 TI - Alkaline positions in CsTiOAsO4 PMID- 10851600 TI - Rerefinement of K2 PMID- 10851601 TI - A binuclear manganese(II) complex, decaaqua(mu-1,2,4,5-benzenetetracarboxylato O1:O4)dimanganese(II) hydrate PMID- 10851602 TI - The dimeric mixed-ligand titanacarborane sandwich PMID- 10851603 TI - Three complexes of bis(N-methylbenzohydroxamato-O,O')copper(II) PMID- 10851604 TI - trans-(4-amino-2,2,6,6-tetramethylpiperidine-N4)bis(pentane-2,4-dionato-O, O')(triphenylphosphine-P)cobalt(III) hexafluorophosphate dichloromethane solvate PMID- 10851605 TI - Sodium dihydrogen tris(cyclopropanecarboxylate) PMID- 10851606 TI - 1,2:1,3-bis(mu-p-toluato-O:O')-1-(triphenylphosphine-P)-1-ruthena- closo undecaborane PMID- 10851607 TI - (Z)-bismu- PMID- 10851608 TI - Tris(2,2'-bipyridyl-N,N')nickel(II) thiosulfate heptahydrate PMID- 10851609 TI - Calcium(II) meso-2,3-diphenylsuccinate heptahydrate PMID- 10851610 TI - A binuclear rhodium(I) complex with two tetraphosphine ligands at 100 K PMID- 10851611 TI - Bis(4-hydroxy-3-methoxybenzaldehyde thiosemicarbazonato-N1,S)nickel(II) tetraethanol solvate. PMID- 10851612 TI - Bimetallic assemblies, PMID- 10851613 TI - Chlorobis PMID- 10851615 TI - [eta 5,sigma-3,3-dimethyl-1-] PMID- 10851614 TI - Anhydrous polymeric zinc(II) octanoate PMID- 10851616 TI - Dichloro(tetrahydrofuran-O)- PMID- 10851617 TI - A 1:1 cocrystal of di-mu-chloro-bis- PMID- 10851618 TI - [Ferrocene-1,1'-diylbis] PMID- 10851619 TI - [1a(1a alpha,5 beta,9a alpha)]-1,1a,4,5,7,8,9,9a-Octahydro-3-hydroxy-1,1,2,5- tetramethyl-7-methylene-6H-cyclopropa-[3,4]cyclohept[1,2-e] inden-6-one. PMID- 10851620 TI - [In Process Citation] PMID- 10851621 TI - 13H-dibenzo PMID- 10851623 TI - 2,5-bis(methylthio)-1,4-benzoquinone and 2-methyl-3-(methylsulfonyl)-benzo PMID- 10851622 TI - Tris(2-cyanoethyl) isocyanurate PMID- 10851624 TI - Strong intramolecular O--H...O hydrogen bonds in quinaldic acid N-oxide and picolinic acid N-oxide PMID- 10851625 TI - 2',3'-Didehydro-3'-deoxythymidine N-methyl-2-pyrrolidone solvate (D4T.NMPO). PMID- 10851626 TI - 1,3(R):4,6(R)-di-O-benzylidene-D-mannitol PMID- 10851627 TI - Bis(diethoxyselenophosphinoyl) triselenide and bis(diisopropoxyselenophosphinoyl) diselenide PMID- 10851628 TI - Two new conformationally restricted 4,5-dihydroxynorvaline analogues with a norbornane skeleton. PMID- 10851629 TI - 3-[4-(3,4-Dimethylphenyl)-1,3-thiazol-2-yl]-2-(2-hydroxyphenyl)-1,2,3,4- tetrahydroquinazolin-4-one. PMID- 10851630 TI - Gallic acid monohydrate. PMID- 10851631 TI - 1,3:16,18-bis(xylyl)-30-crown-8 PMID- 10851632 TI - 2,2,5,5,8,8-hexamethyl-2,3,5,6,7,8-hexahydroimidazo PMID- 10851633 TI - Polyhedral azaborane chemistry, 6-(C5H5N)-arachno-4-NB8H11 PMID- 10851634 TI - 2,4,4-trimethyl-2-phenyl-3-pentanone oxime PMID- 10851635 TI - 5-Nitro-N-phenyl-2-thiofuramide. PMID- 10851636 TI - cis-bis(2,2'-bipyridyl-N-N')dichlorosilicon diiodide PMID- 10851637 TI - Complexes of mixed silicon halides with 4-picoline PMID- 10851638 TI - trans-dichlorotetrakis(4-methylpyridine)silicon bis(triiodide) chloroform solvate PMID- 10851639 TI - Asperuloside monohydrate. PMID- 10851640 TI - O-H...O, C-H...O and C-H...pi(arene) Intermolecular interactions in (2S)-2-(1-oxo 1H-2,3-dihydroisoindol-2-yl)pentanoic acid and (2S)-3-methyl-2-(1-oxo-1H-2,3 dihydroisoindol-2-yl)butanoic acid. PMID- 10851641 TI - [Chlamydia. The molecular organization of the cell and the pathogenetic characteristics of infections]. PMID- 10851642 TI - [The hypolipidemic action of antibiotic 86/88 (enniatin B) in a hepatoblastoma G2 cell culture]. AB - A cyclodepsipeptide antibiotic 86/88 (enniatin B) with strong hypolipidemic action was isolated from the culture liquid of the fungus INA F-86/88 identified as Fusarium lateritium Nees var. stilboides (Wr.) Bilai. In the Hep G2 cell culture the antibiotic suppressed 14C-acetate incorporation into cholesterol (IC50 1.75 microM), cholesterol ethers (IC50 1 microM), triglycerides (IC50 1.3 microM) and free fatty acids (IC50 2.2 microM). The most pronounced effect of the drugs, i.e. the suppression of the cholesterol ethers synthesis is likely due not only to the ACAT inhibition but also to the inhibition of the triglyceride synthesis and the diminishing of the free fatty acids pool in the cells. PMID- 10851643 TI - [The effect of protoplast formation on the antibiotic activity and composition of the actinomycin complex in a strain of Streptomyces galbus (F) subsp. achromogenes 695 and in its active variants]. AB - Protoplasting and regeneration promoted variation by the antibiotic production property in Streptomyces galbus (F) subsp. achromogenes 695 and its active variants 695-3-2 and 695-3-2-206. Variant 695-P24 with the potency 2 times higher than that of the initial strain 695 revertants was selected. No variants lacking the capacity for biosynthesis of the main components of antibiotic A-695 were detected among the revertants still, protoplasting of strains 695-3-2-206 and 695 P24 resulted in formation of variants synthesizing new components of the actinomycin complex. PMID- 10851644 TI - [The efficacy and safety of ciprofloxacin in treating children with mucoviscidosis]. AB - Ciprofloxacin clinical and bacteriological efficacies, as well as tolerability mainly with respect to chondrotoxicity were evaluated in the treatment of children with mucoviscidosis. The drug was shown to have high clinical and moderate bacteriological efficacies. As for its tolerability, adverse reactions chiefly associated with affection of the gastrointestinal tract, i.e. nausea, stomach pain, diarrhea, increased transaminase levels were recorded. The arthrotoxicity episode was single and transitory. The use of ciprofloxacin had no negative effect on the children growth. No chondrotoxic effect of ciprofloxacin in the treatment of children was observed which is explained in the paper. It is concluded that ciprofloxacin is in general an efficient and safe antibiotic useful for the treatment of children. PMID- 10851645 TI - [A comparative evaluation of the pharmacokinetics of different forms of benzathine benzylpenicillin]. AB - Comparative randomized opened pharmacokinetic evaluation of benzathine benzylpenicillin in three dosage forms was performed. Benzathine benzylpenicillin was used as extencilline (2.4 million U or 1.2 million U, "Rhone-Poulenc Rorer", France) and as bicillin-5 (1.5 million U, "Synthesis" Russia). 33 patients were included in investigation (23 women and 10 men aged 16-60 years). 25 persons had verified rheumatism without blood circulation failure signs, 4--had chronic tonsillitis and 4 were healthy volunteers. Benzylpenicillin concentration was estimated by microbiology test in blood samples taken at 1, 3, 24 hours and 7, 14 and 21 days after intramuscular drug injection. After 2.4 million U extencilline injection (12 patients) its concentration, was at the inhibition level for beta hemolytic streptococcus group A (25 ng/ml) for 3 weeks-period in 83.3 per cent of patients. After 1.2 million U extencilline injection (10 patients) or 1.5 million U bicillin-5 injection (12 patients) the above mentioned concentration was achieved on the 21st day in 30 and 0 per cent of patients respectively. Thus the treatment with benzathine benzylpenicillin at the 1.2 million U dose in the form of extencilline or bicillin-5 doesn't provide adequate antistreptococcal concentration in blood in prolonged period and is not suitable for correct rheumatism prophylaxis in adult patients. PMID- 10851646 TI - [Acute rheumatic fever and Streptococcus group A tonsillitis: the current status of the problem and the questions of antibiotic therapy]. PMID- 10851647 TI - [Enterobacterial sensitivity to beta-lactam antibiotics]. AB - The susceptibility to betalactams of 868 enteric bacteria isolated from the patients at the hospital was studied. The isolated pathogens included: E. coli (549), Klebsiella sp. (195), Serratia sp. (124). Ampicillin and cefazoline demonstrated the lowest activity. Cefotaxime, ceftazidime and imipenem were active against 90 per cent of isolates. Among E. coli isolates the susceptibility to the above mentioned drugs was the following: 95.1, 96.9, 99.3 per cent, among Klebsiella sp.--89.7, 88.7, 97.9 per cent, among Proteus sp.--89.5, 90.3, 91.9 per cent respectively. Thus cefotaxime may be used in antibacterial empiric therapy if Pseudomonas aeruginosa infection is excluded. PMID- 10851648 TI - [The antibacterial therapy of acute cystitis and pyelonephritis in adults]. PMID- 10851649 TI - [The pharmacoepidemiological approach in treating patients with acute sinusitis]. PMID- 10851650 TI - [The effect of the pharmacodynamics of different classes of antibacterial preparations on their dosage regimens]. PMID- 10851651 TI - Physiologic monitoring systems. AB - Physiologic monitoring systems monitor vital physiologic parameters so that clinicians can be informed of changes in a patient's condition. They typically consist of several distinct components, including a central station, bedside monitors, and ambulatory telemetry transmitters and receivers. We previously evaluated physiologic monitoring systems in our January-February 1999 issue (Health Devices 28[1-2]). For this update, we have tested systems from two additional suppliers, using the same criteria and test methods that we used for the eight systems in the original study. As in the earlier Evaluation, we have examined how each system functions as a whole, rather than focusing on the performance of individual components. We judged the systems primarily on adaptability, alarm implementation, and human factors design. In the Conclusions, we compare all 10 units evaluated to date. We also provide updated product information for the systems evaluated in our original study; in some cases, the changes that have been made to those systems have caused us to revise our ratings. We rated all the systems based on their capabilities for each of six applications: (1) critical care unit, (2) emergency department, (3) intermediate care unit and general medical/surgical floor, (4) operating room, (5) postanesthesia care unit, and (6) transport. As in our earlier study, we have determined that most of the evaluated systems have drawbacks for one or more of the evaluated applications. In our January-February 1999 Evaluation of physiologic monitoring systems (Health Devices 28[1-2]), we discussed the issues surrounding the purchase and use of these systems. In this introduction, we recap the most significant points from that discussion, provide some new information about ambulatory telemetry and wireless networks, and briefly describe the components of this Update Evaluation. PMID- 10851652 TI - New FDA regulation on reprocessing single-use devices. How it may affect U.S. hospitals. PMID- 10851654 TI - Two-piece Luer lock may not be locked. PMID- 10851653 TI - Injuries--one fatal--highlight the need for pre-use testing of disposable breathing circuits. PMID- 10851655 TI - Don't place blankets--or anything else--on infant warmers. PMID- 10851656 TI - Understanding fuse problems. PMID- 10851657 TI - Unexpected shutoff of Agilent/Hewlett-Packard CodeMaster 100 defibrillators when used with external DC power module (M2478A). PMID- 10851658 TI - Anesthesiology and the academic medical center: place and promise at the start of the new millennium. PMID- 10851659 TI - Effects of vagal perineural capsaicin treatment on vagal efferent and airway neurogenic responses in anesthetized rats. AB - The acute effect of vagal perineural capsaicin treatment (VPCT) on parasympathetic bradycardia and tracheal neurogenic protein extravasation was examined. In nine anesthetized male Wistar rats the effect of VPCT on the bradycardia induced by electrical stimulation of the vagus was examined. In 24 anesthetized male Wistar rats the effect of VPCT on the tracheal protein extravasation induced by the inhalation of capsaicin aerosols was also studied. VPCT did not alter the bradycardia induced by vagal stimulation or the tracheal protein extravasation induced by the inhalation of capsaicin aerosol. The results of these studies further demonstrate the selectivity of perineural capsaicin treatment on vagal sensory nonmyelinated fibers in the rat and indicate that it is a useful tool for examining the role of sensory vagal C-fibers in pulmonary and cardiovascular reflexes and in isolating C-fiber-mediated reflex responses from those mediated by the release of neuropeptides. PMID- 10851660 TI - The effect of cadmium and experimental diabetes on EEG spectral data. AB - Fifty-two healthy male Swiss albino rats, aged three months, were used in this study. They were divided into four groups: control (C), diabetic (D), cadmium (Cd) and diabetic + Cd (D + Cd). Diabetes was induced in the D and D + Cd groups by administration of alloxane (5 mg/100 g). After this treatment, the Cd and D + Cd groups were injected with CdCl2 i.p. (2 mg/kg/week). At the end of the two month experimental period, EEGs of the four groups were recorded and amplitude spectral analysis was computed by Fast Fourier Transform (FFT) algorithm. Significant amplitude (dB) increment was observed in 16-30 Hz of Cd group compared with C and D groups. Significant amplitude decrement was found in the 2 4 Hz frequency band of the D + Cd group compared with C, Cd and D groups. PMID- 10851661 TI - Effects of the combination of halothane and serotonin uptake blockers on synaptic transmission in the rat dentate gyrus in vitro. AB - Although the exact basis of their action remains unknown, volatile agents affect noradrenergic and serotoninergic systems. Imipramine and fluoxetine have documented effects on these neurotransmitter transmission systems. Given the common sites of action of these antidepressants and halothane, we examined their individual and combined effects on tonic excitatory post-synaptic potentials (EPSPs) and frequency dependent blockade in the rat dentate gyrus in vitro. Extracellular recordings of field EPSPs were maintained from the dentate gyrus, in the presence of picrotoxin (100 microM). Stimulation at 30 Hz (200 ms) allowed investigation of frequency dependent blockade. Once a stable equilibrium was established, halothane, imipramine and fluoxetine were administered via the perfusate and recordings were made. Halothane produced a dose dependent reduction in EPSP amplitude (EC50 0.28 mM; n = 12). Imipramine (1-10 microM) potentiated the EPSP amplitude (148.2 +/- 8.2%; imipramine 1 microM; n = 6). Fluoxetine (0.5 10 microM) reduced EPSP amplitude to 83.7 +/- 22.1% of control (n = 6). In the presence of halothane 0.2 mM, imipramine reduced the EPSP amplitude to 56.5 +/- 9.9% of control (imipramine 10 microM; n = 6; p < 0.05 compared with imipramine alone). Halothane (0.2 mM) demonstrated frequency dependent blockade. However, neither imipramine nor fluoxetine showed use dependent inhibition at the doses investigated. When combined with halothane 0.2 mM, fluoxetine 10 microM demonstrated frequency dependent blockade at the sixth pulse in the train compared with controls (13.8 +/- 4.7% vs 38.1 +/- 8.3%; n = 6; p < 0.05). The halothane-imipramine combination did not exhibit use dependent blockade greater than controls. The reversal of imipramine-induced EPSP potentiation by the preapplication of halothane has not been previously reported. It may be due to modulation of noradrenergic transmission by halothane. The frequency dependent blockade produced by the combination of fluoxetine 10 microM and halothane may be mediated by a nonspecific membrane effect on 5-HT uptake. These differing effects underline the broad action of volatile agents on synaptic mechanisms. PMID- 10851662 TI - Effects of high nitrate intake in rats. AB - The purpose of this study was to determine the effects of administration of high dose nitrate in drinking water on weight gain, hematological parameters and osmotic fragility in rats. We compared these parameters in 40 rats divided into four groups (one control and three treatment groups). Control animals drank filtered tap water containing a maximum of 10 mg/l nitrate while the treatment groups drank 100 mg/l, 200 mg/l and 400 mg/l nitrate-containing water ad libitum for 60 days. Animals in the treatment groups gained less weight than the control group and the differences between the control and treatment groups were statistically significant (p < 0.05). At the concentration of 100 mg/l nitrate, platelet counts and hemoglobin levels were significantly increased compared with the control group (p < 0.05). At the concentration of 200 mg/l nitrate, erythrocyte counts, hemoglobin and hematocrit levels were significantly increased compared with the control group (p < 0.05). At the concentration of 400 mg/l nitrate, platelet counts were decreased significantly when compared with the first two treatment groups (p < 0.05). There were statistically significant differences in osmotic fragility ratios between treatment groups and the control group (p < 0.05). We concluded that high nitrate intake in drinking water decreases weight gain, affects hematological parameters by inducing bone marrow activity at low doses and inhibiting it at high doses, and increases erythrocyte osmotic fragility. PMID- 10851663 TI - 4-Aminopyridine can induce release of calcium from the sarcoplasmic reticulum of frog heart. AB - 4-Aminopyridine (4-AP), a K(+)-channel blocker, has been associated with cellular Ca2+ movements in various excitable tissues but its interaction with intracellular Ca2+ pools has not been described. Spontaneously beating ventricular strips of frog hearts responded to 4-AP (1.6 x 10(-2) M) by contracture that was susceptible to tolerance development when 4-AP was applied in a repetitive manner after washout periods. Under full mechanical inhibition by Ca(2+)-free Ringer or high-K+ Ringer or a combination of both, 4-AP could still produce contracture, a finding indicating that the source of Ca2+ for such an effect should be intracellular Ca2+ pools, namely the sarcoplasmic reticulum. It was concluded that 4-AP could induce release of Ca2+ from internal stores that might be of importance in its cardiac actions. PMID- 10851664 TI - Correlation of two levels of space proton flux with monthly distribution of deaths from cardiovascular disease and suicide. AB - In our previous studies /1-3/ we described some significant links between monthly number of deaths due to cardiovascular disease and suicide and space proton flux > 90 MeV. The aims of the present study were to compare the relationship of some solar and geomagnetic parameters with space proton fluxes of > 60 and > 90 MeV; to examine the monthly correlation of these two proton groups with the monthly death distribution in two countries, Israel and Lithuania. Physical data were obtained from the National Geophysical Data Center and the SESC in Boulder, CO; NSSDC in Goddard Space Flight Center, USA, and the Izmiran Institute of the Academy of Sciences in Russia. Pearson correlation coefficients and probabilities were compared for 56-180 consecutive months. Proton flux of > 60 MeV significantly correlated with three of the four studied monthly geomagnetic activity indices (Ap, Am, Dst), but not with such solar activity markers as sunspot number and solar flux (2800 MGH, 10.6 cm). There was no significant relationship between proton flux of > 60 MeV and monthly number of deaths from cardiovascular diseases and suicide, in contrast to the results for > 90 MeV. From the data available during the 36 months (1986-1988), there was no correlation between monthly levels of > 60 to > 90 MeV. In conclusion, a monthly space proton flux of > 60 MeV is not significantly correlated with the monthly death distribution from cardiovascular disease and suicide and some solar activity indices, such as proton flux of > 90 MeV. It is possible that the 60-90 MeV fraction in the > 60 MeV proton flux "blunts" the cosmobiological relationship between proton flux of > 90 MeV and monthly death number. PMID- 10851665 TI - Erythrocyte glutathione, glutathione peroxidase, superoxide dismutase and serum lipid peroxidation in epileptic children with valproate and carbamazepine monotherapy. AB - This prospective study was carried out to determine changes in the antioxidant system in epileptic children receiving long term antiepileptic drugs (AEDs). Levels of erythrocyte glutathione (GSH), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and serum lipid peroxidation were determined in 25 healthy children and 30 epileptic children who had not yet received AEDs. Sixteen patients were treated with valproic acid (VPA) and 14 with carbamazepine (CBZ); 13 months later these parameters were retested. The results showed that SOD and lipid peroxidation levels were increased but the GSH-Px levels were decreased in epileptic children on VPA therapy compared with the control group and the results before treatment. No significant differences of these parameters were found in epileptic children on CBZ therapy compared with the control group and the results before treatment, except that lipid peroxidation level was slightly higher in epileptic patients before treatment. We conclude that antioxidant systems in epileptic children on CBZ therapy are better regulated in comparison with epileptic children on VPA therapy. PMID- 10851666 TI - Reconstruction of the severely resorbed (Class VI) jaws: routine or exception? AB - After disappointing results with conventional preprosthetic procedures endosseous implants in combination with alveolar ridge augmentation opened up new prospects in reconstructive surgery. A total of 64 patients who underwent three-dimensional reconstruction of the alveolar ridge and insertion of endosseous implants for severe resorption were evaluated retrospectively. Despite the postoperative infection rate of 20.3% (13 patients), only 4.1% of the 266 inserted implants were lost in the long term. This indicates that augmentation using free autogenous iliac bone grafts and implants have a success rate of approximately 96% despite difficult initial situations. This success was mainly related to the soft tissue condition covering the graft. A technique for soft tissue dissection, especially in the maxilla, is presented. PMID- 10851667 TI - A new distraction device to compare continuous and discontinuous bone distraction in mini-pigs: a preliminary report. AB - Callus-distraction has become an accepted procedure to lengthen hypoplastic mandibles in humans. Extra- and intraoral devices have been applied successfully. Systematic studies have proven the importance of direction, stability, rate and frequency in callus-distraction. In an experimental animal study a newly developed intraoral microhydraulic osteodistractor was tested. Initially the pressures necessary to distract the mandible in mini-pigs were recorded in discontinuous callus-distraction. These results were used to perform continuous bone distraction. Besides testing the new distractor and evaluating the distraction pressures, the aim of the study was to prove that direct bone growth occurred without preceding cartilage formation. Clinical and microscopic results are presented. PMID- 10851668 TI - Effect of electrical stimulation on mandibular distraction osteogenesis. AB - This study was designed to examine whether the use of electrical stimulation during mandibular lengthening accelerates new bone formation. Twenty adult female rabbits weighing between 2800 g and 3200 g underwent left mandibular body osteotomy. After a 3 day latency period, an external fixation device was activated at a rate of 0.7 mm per day for 10 days. Direct current electrical stimulation of 10 microA was applied to 10 rabbit mandibles. Two of the screws were used as electrodes during the distraction phase. The other 10 rabbits (control group) were not stimulated. The device was then stabilized for periods of 10, 20, 30 and 60 days in both groups. The distraction segment was evaluated radiographically by assessing the proportion of bone mineral density using a dichroma scan. The amount of new bone formation was studied histologically with an image analyzer to evaluate the bone formation in the distraction gap. Histological examination showed that the new bone formation 10 and 20 days after distraction was greater in the electrical stimulation group than in the control group. Ten and 20 days after distraction, image analysis and analysis of bone mineral density in areas of newly formed bone indicated that there was a greater amount of new bone formation in the stimulation group than in the control group. The radiographic evaluation, however, did not demonstrate significantly different images between the stimulation group and the control group. Thirty and 60 days after distraction, no difference in the amount of new bone formation was noted in either the experimental or the control groups. These results indicate that electrical stimulation during gradual distraction promotes new bone formation in the early retention period in a rabbit model. PMID- 10851669 TI - Vertical distraction of a free vascularized fibula flap in a reconstructed hemimandible: case report. AB - The authors report a case of vertical distraction osteogenesis of a free revascularized fibula flap used to reconstruct an hemimandible lost as a result of a gunshot injury. The reconstruction procedure and the distraction protocol are described; clinical and radiological results are presented. The vertical discrepancy between the fibula and the native right hemimandible was corrected. PMID- 10851670 TI - Soft tissue profile changes after Le Fort I osteotomy in UCLP patients. AB - The changes in soft tissue profile after Le Fort I osteotomy were evaluated cephalometrically in 38 consecutive UCLP patients (25 males, 13 females) operated on between 1987 and 1995. Mean age at operation was 23.5 years. The one-piece Le Fort I osteotomy was fixed with titanium plates and the osteotomy site was bone grafted. Neither intermaxillary fixation nor occlusal splints were used postoperatively. Soft tissue changes were analyzed both horizontally and vertically by cephalograms taken shortly before surgery, 6 months and 1 year postoperatively. The mean maxillary skeletal advancement (point A) during surgery was 3.8 mm and mean vertical lengthening 4.4 mm. One year postoperatively the horizontal change in the upper lip profile (point a) was 80% of the skeletal change. Vertically, the soft tissue change in the upper lip was smaller 40%, but increased significantly (to 58%) if V-Y plasty was used. The V-Y plasty also increased the anteroposterior thickness of the upper lip. No significant soft tissue changes were observed between 6 months and 1 year postoperatively. PMID- 10851671 TI - Iliac crest donor site morbidity following open and closed methods of bone harvest for alveolar cleft osteoplasty. AB - Donor site morbidity after bone harvesting still remains a crucial problem in alveolar cleft osteoplasty. This study focuses on ilium donor site morbidity comparing two different techniques. A series of 52 consecutive patients was divided in half. All had anterior iliac crest bone grafts. In the study group the harvesting was performed with a closed osteotomy using a cylindrical Shepard osteotome. The control group underwent the traditional open osteotomy. In the open osteotomy group the short-term morbidity at the donor site was slightly greater than in the closed harvesting group. The low short-term morbidity in the closed harvesting group was reflected in the analgesic consumption which was three times higher in the open osteotomy group (p < 0.008). The most striking difference occurred in the appearance of the mature scar: a length of 24.2 mm (mean) in the closed harvesting group against 60.3 mm in the open osteotomy group (p < 0.0001), and a width of 4.9 mm (mean) versus 7.7 mm, respectively (p < 0.003). The long-term morbidity was negligible in both groups. Based on these findings we suggest that bone harvesting from the anterior iliac crest remains the preferred method, provided that closed harvesting is undertaken. PMID- 10851672 TI - Improvement of pain and function after arthroscopy and arthrocentesis of the temporomandibular joint: a comparative study. AB - In 1993 and 1994, 720 patients with pain in the temporo-mandibular joint area were examined and treated. The authors describe their therapeutic protocol. Sixty two patients were not relieved by conservative nonsurgical therapy and were treated by arthroscopy or arthrocentesis. In both groups the differences in functional result and in pain control were analysed. Results show that both arthroscopy and lavage are useful in improving function and diminishing pain. Arthroscopy shows better results for functional treatment whereas arthrocentesis and arthroscopy show similar results in pain control. PMID- 10851673 TI - Does expression of inducible nitric oxide synthase correlate with severity of oral epithelial dysplasia? AB - The small molecule nitric oxide (NO) has generated an exponential amount of research since its discovery as a biological messenger in 1987. It has a vast number of actions, many of which are poorly understood. It has been studied in a variety of human cancers and has been implicated both in tumour promotion and inhibition. Although NO is produced by three distinct isoforms of the enzyme nitric oxide synthase (NOS), most cancer research is directed towards the calcium independent form, iNOS which following induction, produces much higher quantities of NO than the other two. In this study the expression of iNOS is assessed by immunohistochemistry in 26 cases of oral epithelial dysplasia ranging in severity from mild to severe. iNOS staining was found in all 26 cases of dysplasia with the degree of staining correlating to the severity of dysplasia (p < 0.001). There was no iNOS staining seen in adjacent normal epithelium. The possible role of iNOS in the complex transformation from dysplasia to invasive oral cancer and the clinical applications are discussed. PMID- 10851674 TI - Survival evaluation of treatment modality in squamous cell carcinoma of the oral cavity and oropharynx. AB - The treatment of squamous cell carcinoma of the mouth and oropharynx continues to change. In this primary report, we compared the results obtained by combined surgery and radiation therapy, or either modality alone. Other methods such as brachytherapy, or hyperfractionated radiotherapy, were not included in our protocols. A statistical analysis of the 3- and 5-year survival rates in relation to location and size of the primary tumour, stage at initial presentation, treatment modality and recurrence, was carried out in 88 patients with squamous cell carcinoma of the oral cavity or oropharynx. The overall survival rate was 73.8% at 3 years and 66.3% at 5 years. Size of tumour and stage at presentation were significant when P value was adjusted by site. Survival was significantly associated with type of treatment (combined approach obtained superior results), location of primary tumour, and recurrence. The type of neck dissection did not show any effect. Therapeutic modality used, stage, and location of primary tumour significantly influenced survival. A more selective combined initial treatment according to site and stage (distribution) is recommended. PMID- 10851675 TI - Salivary gland tumours treated in the stomatological clinics in Bratislava. AB - During the period of 1951-1996 a group of 1021 patients (484 men and 537 women, mean age 53 years, range 2-87 years) with salivary gland tumours were analysed retrospectively. The mean follow-up period was 8 years (range 10 months-25 years). The frequency of benign tumours was 74% (n = 755) and malignant tumours 26% (n = 266). Lesions were sited in the parotid gland 83% (n = 847), in the submandibular gland 10.8% (n = 110), in the sublingual gland 3.2% (n = 33) and in the minor salivary glands 3% (n = 31). The most common benign tumours were pleomorphic adenoma in 53.9% (n = 550) and Warthin's tumour 9.7% (n = 99). Of the malignancies, the adenoid cystic carcinoma was most common (6.4% of cases, n = 65) and mucoepidermoid carcinoma occurred in 5.2% (n = 53). The predominant therapy was surgery alone or in combination with postoperative radiotherapy in 93.7% (957 cases), radiotherapy alone after fine needle aspiration biopsy for 4.7%, and 19 patients remained untreated. PMID- 10851676 TI - Competence in language at 24 months: relations with attachment security and home stimulation. AB - This study explored the interrelations among attachment, home stimulation, and language development in 58 toddlers (36 medically high risk and 22 low risk) at 24 months of age. The results indicated that there were additive effects of attachment and home stimulation on language competence, especially on receptive abilities. Mothers who had established secure relationships and provided stimulating home environments had children with the highest language scores. PMID- 10851677 TI - Intimacy and psychosocial adjustment in Hong Kong Chinese adolescents. AB - This study examined the association between 8 aspects of friend intimacy and 3 measures of psychosocial adjustment (self-esteem, deviant behavior, and purpose in life) among Hong Kong Chinese adolescents. The respondents were 289 students between 16 and 19 years of age from a cross-sectional study in Hong Kong. Bivariate correlation and multiple regression analyses revealed (a) friend intimacy was positively associated with self-esteem and purpose in life, and (b) friend intimacy was negatively correlated with deviant behavior. Results indicated that friend intimacy is an important variable that affects some aspects of psychosocial adjustment. Moreover, results of multiple regression analyses indicated that different measures of friend intimacy are related to deviant behavior and purpose in life. Therefore, it is crucial to study the relationship between different aspects of friend intimacy and different aspects of psychosocial adjustment among adolescents. PMID- 10851678 TI - Racial differences in adolescent coping and self-esteem. AB - Racial differences in coping strategies and self-esteem were examined for 361 male and female adolescents in Grades 7-12. Coping strategies were assessed with the Adolescent Coping Orientation for Problem Experiences (J. M. Patterson & H. I. McCubbin, 1986). Self-esteem was assessed by the Coopersmith Self-Esteem Inventory (S. Coopersmith, 1987). Multivariate analysis revealed racial differences in adolescent coping strategies of ventilating feelings, seeking diversions, developing self-reliance, avoiding problems, seeking spiritual support, investing in close friends, engaging in demanding activities, solving family problems, and relaxing. In particular, African American adolescents reported using diversions, self-reliance, spiritual support, close friends, demanding activities, family problems, and relaxation more frequently than Caucasian adolescents did. Implications for professionals and recommendations for future research are discussed. PMID- 10851679 TI - Emotional autonomy and depression among Chinese adolescents. AB - Depression is quite common among young people in Hong Kong Chinese society. This study examined the association between emotional autonomy and depressive symptomatology among Chinese young people in Hong Kong. The respondents were 512 young people between 16 and 18 years of age from a cross-sectional study in Hong Kong. Significant bivariate relationships were found between depressive symptomatology and three dimensions of emotional autonomy (individuation, nondependency on parents, and deidealization of parents). Using multiple regression models, the author found that depressive symptomatology was associated with two aspects of emotional autonomy: individuation and deidealization of parents. Results indicate that the relationships between depressive symptomatology and these three aspects of emotional autonomy are similar in both individualistic and collectivistic societies. PMID- 10851680 TI - Quality of mother-child interaction and the intergenerational transmission of sexual values: a panel study. AB - The role of mothers as socializing agents was examined in a longitudinal study. It was assumed that mothers can transmit their own values to their children, especially if the quality of their interaction is good. Whether children influence their mothers' values was also examined. The hypotheses were tested using analysis of variance and structural equation modeling, drawing on data from 253 English adolescent-mother pairs. The results provided little support for the hypotheses. There was some evidence that mothers influenced their children's sexual attitudes, but this effect was found for the families characterized by a low quality of parent-child interaction, rather than for the families characterized by a high quality of interaction. No evidence was found for the assumption that children influence their mothers' sexual attitudes and beliefs. Implications of the study for future research are discussed. PMID- 10851681 TI - Responses of preschoolers in two frustrating episodes: emergence of complex strategies for emotion regulation. AB - Although research on young children's abilities to organize emotional states has increased in recent years, little is known about the emergence of complex strategies for emotion regulation in preschoolers. In the present study, emotion regulation strategies used by 52 normally developing 3- and 4-year-olds were examined. Children and their primary caregivers (50 mothers, 2 fathers) participated in 2 controlled frustration episodes that were videotaped. Four types of strategies were coded: comforting behaviors, instrumental behaviors, distraction behaviors, and cognitive reappraisals. Results indicated that 3-year olds used proportionately more instrumental strategies than 4-year-olds, and parents of 3-year-olds showed the same pattern, whereas parents of 4-year-olds did not. Moreover, 3-year-olds used a variety of strategies when frustrated, including cognitive reappraisals. Significant positive correlations were found between the types of strategies used by the children and by the parents to help their children. It is suggested that children may be using strategies to organize their emotional states before they are able to accurately report on them. PMID- 10851682 TI - Effectiveness of primiparae and multiparae at soothing their newborn infants. AB - To examine the effectiveness of new parents at soothing their infants, the authors filmed primiparae (20 mothers, 20 fathers) and, for comparison, multiparae (25 mothers, 25 fathers) during an interaction with their crying 2- to 3-day-old infants. Data were derived from loudness ratings of the infants' distress signals and by measuring the time it took parents to quiet their infants. In addition, specific parental behaviors were coded by microanalysis. From these data, measures of soothing effectiveness and behavioral profiles were derived. Data analyses showed that most primiparae were effective at soothing their infants' cries and that there were no parity effects on measures of soothing effectiveness. However, mothers, regardless of parity, were more effective at quieting their infants than were fathers, and there were significant differences in mothers' and fathers' caregiving behaviors. These data suggest that primiparae are effective at quickly soothing their newborns and that their skill, as measured, does not depend on parenting experience. Moreover, the data point to significant differences in mothers' and fathers' competence at quieting their newborns whether or not they are experienced at parenting. PMID- 10851683 TI - The development of children's knowledge of attention and resource allocation in single and dual tasks. AB - Developmental changes in kindergarten, 1st-, and 4th-grade children's knowledge about the variables that affect attention sharing and resource allocation were examined. Findings from the 2 experiments showed that kindergartners understood that person and strategy variables affect performance in attention-sharing tasks. However, knowledge of how task variables affect performance was not evident to them and was inconsistent for 1st and 4th graders. Children's knowledge about resource allocation revealed a different pattern and varied according to the dissimilarity of task demands in the attention-sharing task. In Experiment 1, in which the dual attention tasks were similar (i.e., visual detection), kindergarten and 1st-grade children did not differentiate performance in single and dual tasks. Fourth graders demonstrated knowledge that performance on a single task would be better than performance on the dual tasks for only 2 of the variables examined. In Experiment 2, in which the dual attention tasks were dissimilar (i.e., visual and auditory detection), kindergarten and 1st-grade children demonstrated knowledge that performance in the single task would be better than in the dual tasks for 1 of the task variables examined. However, 4th grade children consistently gave higher ratings for performance on the single than on the dual attention tasks for all variables examined. These findings (a) underscore that children's meta-attention is not unitary and (b) demonstrate that children's knowledge about variables affecting attention sharing and resource allocation have different developmental pathways. Results show that knowledge about attention sharing and about the factors that influence the control of attention develops slowly and undergoes reorganization in middle childhood. PMID- 10851684 TI - Predictors of children's emotional distress in a mother-absent situation: implications for caregiving research. AB - Although many factors have been found to be associated with caregiving behavior among preschool-age siblings, few studies have considered the older child's emotional responses to the distress of a younger sibling in the mother's absence. In this study, the authors considered both individual and family factors that predict older and younger sibling distress in a mother-absent situation designed to elicit caregiving behavior. For both older and younger siblings, the strongest predictor of distress following the mother's departure was distress in the other sibling, suggesting the importance of situational factors. Individual and family factors, however, were also found to predict child distress. Among older siblings, shy and sociable temperaments were associated with greater distress during the mother's absence. Higher levels of parenting stress were found to be associated with less distress among both older and younger siblings in the mother absent situation. Results are discussed in terms of individual and family environment influences on the sibling relationship. PMID- 10851685 TI - Comparison of two tests of executive functioning. PMID- 10851686 TI - [Fracture care in advanced age--an increasing but manageable problem]. PMID- 10851687 TI - [Fracture management in the aged]. AB - Fracture care in a geriatric population should lead to a mobilisation with full weight bearing as soon as possible after the accident. Osteoporosis and associated preexisting disease influences the kind of fracture care as well as the social situation. A retrospective analysis of 888 consecutive patients from 1994 to 1998 showed that the fracture risk for a second independent bony lesion was 5.2% in this population with a mean age of 83.4 years (75 y-97.2 y). 946 fractures were treated operatively with a overall reoperation rate of 10%. 3.1% were soft tissue revisions due to infection, 2.0% were because of implant failure in connection with a deep infection and 5.2% of the reoperation were indicated because of implant failure alone. Distribution of complications showed the advantage of closed, indirect reduction and intramedullary devices and primary hemiarthroplasty in femoral and humeral head fractures. PMID- 10851688 TI - [Preoperative patient education. What is especially important?]. AB - Any medical procedure must be legally authorized by the patient, giving informed consent. This prerequisite is based on the right to self determination. Otherwise a medical procedure is considered as physical injury. Thus, the way, how a specific procedure is performed, predominantly depends on the patient's will. The attending physician's duty is to find out this will, which is especially important when old patients and patients incapable of giving valid consent are involved. Before the procedure the physician has to have a confidential talk with the patient. He has to point out the best treatment possible considering the individual circumstances of the patient's life. The physician has to find out, weather the patient is competent to refuse or consent to treatment. Furthermore the consultation is an opportunity to diminish the patient's fear and to increase his confidence. The main problem treating old patients is that they often lack the mental ability for appreciation of the nature of the situation. One has to differentiate between the physiological mental decay and dementia or psychiatric diseases. In these cases a third party consent is necessary. PMID- 10851689 TI - [Femoral neck fracture. Osteosynthesis or which endoprosthesis is indicated?]. AB - Femoral neck fractures are frequent fractures of the elderly. They can be treated by the use of dynamic hip screws, lag screws, bipolar hemi-protheses or total hip replacement. The results are markedly influenced by the timing of the operation and the choice of the implant. Using the Garden classification of femoral neck fracture we demonstrate a different therapeutic approach according to Garden stadium I-IV. PMID- 10851690 TI - [Therapeutic possibilities in trochanteric fractures. Safe--fast--stable]. AB - Trochanteric fractures frequently occur in elder patients with severe osteopenia. These fractures are highly unstable. Conservative treatment is not indicated because of the long period of immobilization until fracture healing is achieved. Simple fractures (A1) can be successfully fixed by either plate-hip screw systems like Dynamic Hip Screw (DHS) or nail-hip screw systems (Gamma-Nail, Proximal Femur Nail, Classic Nail). As a result of biomechanical and clinical trials the intramedullary implants should be preferred for fixation of more comminuted fractures (Type A2.2, A3.3) and subtrochanteric fractures. For ipsilateral femoral shaft and trochanteric fractures long nail-hip screw-systems are recommended like the long PFN or the long Gamma nail. In about 90% of the patients with trochanteric fractures postoperative weight bearing can be achieved by internal fixation if gliding systems are applied. Intraoperative and postoperative complications however occur more frequently after the implantation of Gamma-Nail than after application of a DHS. Other implants like Ender Nails, 130 degrees angled plate or twisted plate are not further recommended. PMID- 10851691 TI - [Spinal fractures in the elderly and their treatment]. AB - Spinal injuries referring to banal falls are common injuries in elderly persons. Up to 55 years of age spinal fractures are more frequent in men while its opposite in older persons. There are two typical lesions: odontoid fracture in the upper cervical spine and compression fracture in thoracolumbar spine. Odontoid fracture is the most frequent lesion in cervical spine in the elderly. This is caused by trabecular structure of dens axis and spine's stiffness in old age. Instable fractures in extension type II are most common. In many cases the instability of the lesion is hidden and can be first seen in functional examination controlled by x-ray. Conservative treatment is efficient in stable lesions type III while there is a high risk for pseudarthrosis in treatment of instable lesions in elderly with Halo Fixateur e.g. We think that the best operation for these fractures is the anterior screw fixation. If this is not sufficient because of low bone mass or early dislocation of the screws a dorsal fusion C1/C2 with trans-articular screwing e.g. should be done. Because of rare symptoms thoracolumbar fractures in osteoporosis are hidden frequently. In many cases it's not easy to distinguish acute lesions from healed fractures or tumor lesions. Neurological deficit is rare. Therefore there are just few compression fractures (A type) that have to be treated operatively. Instable lesions type B and C must be operated as well in the elderly as in the young. Because of low bone mass in elderly the dorsal instrumentation should include more than just the two injured segments. An even more adequate stabilisation is done with an additional vertebral replacement. PMID- 10851692 TI - [Fractures of the distal forearm. Which therapy is indicated when?]. AB - Every 15th case of a bone fracture in patients aged more than 65 years concerns the distal radius in Germany. This means the second rank of all geriatric fractures following fractures of the hip. According to the approved and increased apply of operative stabilisation there are arising more and more reports upon poor results of nonsurgical treatment. Especially in older patients the main reason for the discontented outcome of conservative management is osteoporosis, which is an affirmative circumstance for the genesis of fracture but also for secondary mal-aligment of comminuted thin cortical walls and crushed porotic cancellous bone. The rational of this perception is either filling artificial bonelike tissue--avoiding the need of harvest cancellous bone graft from a second surgical site--into the resultant cavity following reduction, or supplementary trans-styloidal or intrafocal K-wiring until remodeling is obtained within an average of 10 weeks. Both arrangements should be secured in addition with a trans articular external fixation. According to a literature review and our own experiences of 92 follow up cases of distal radius fractures in patients who were older than 65 years this procedure seems to be superior at present for A-2, A-3 and most cases of type-C fractures of the distal radius, despite the disadvantage of joint immobilisation for about 5 weeks. Type B-fractures, however, should be provided better with an internal fixation. Sudeck's algodystrophia is the mostly serious complication of the distal radius fracture and its treatment in older patients. Recognising punctually neurovegetative stimulated patients, treat them cautiously and coming in on their special situation is usually the best way to reduce this risk. To pay attention to the topography of the nerves during the application of the pins and to act at the first signs of complications immediately is also very important. We examined 92 patients who were older than 65 years with a fracture of the distal end of the radius in a follow up study. In this group we treated 62 distal radius fractures of the type A and C of the AO classification with an trans-articular external fixateur and with supplementary measures if necessary. Additionally we filled the bone cavity with an artificial bone graft (Endobon) following closed reduction in 32 cases, 12 times a complementary K-wiring was added and in 8 cases the external fixateur was combined with both. Sudeck's disease appeared in 1.1% of all cases. In 5 cases we recorded complications with an obligation to treatment as well. Lesion of the superficial branch of the radial nerve were noted in 2 cases (2.1%), as far as we extended the surgical approach for the pin application. Technical problems from the site of the external fixateur appeared in 3 cases, two of them could be attributed to an infirmity of the fixateurs ball joints which is now eliminated by the producer. 4 patients with a type-B fracture was provided by plating, additionally 4 patients with a type-C fracture because of non-compliance. The external fixateur is an essential part of a differentiated treatment with reference to the several types of distal radius fractures in older patients. PMID- 10851693 TI - [Proximal and distal humerus fractures in advanced age]. AB - In contrast to distal humeral fractures, humeral head fractures represent a common injury to the old patient. For both fracture localizations osteoporosis and multi-morbidity are of significant importance. The classification for humeral head fractures in one-, two-, three- and four-part fractures is generally accepted. Valgus impacted head fractures as well as head-splitting fractures are considered a separate entity. In none or minimally displaced fractures good functional results are achieved by conservative means. Although numerous therapeutical procedure are offered in the treatment of displaced fractures of the proximal humerus the result are often disappointing. Generally, minimal invasive surgical procedures should be preferred. However, in dislocated multi part fractures a primary humeral head replacement is often the treatment of choice. In patients with multifarious morbidity a conservative treatment is always to be included into the therapeutical considerations. Distal fractures of the humerus are classified into extraarticular, intraarticular uni-condylar and intraarticular bi-condylar fractures. The therapeutical recommendations, also in the elderly, is relatively homogeneous: primary open reduction and internal fixation (ORIF) should be carried out aiming for an early postoperative functional treatment. Depending from soft tissue conditions and accompanying injuries the functional results are often good or moderate and are generally comparable to those of younger patients. Following a correct indication for surgical intervention the main priority for both distal and proximal humeral fractures is an early definitive surgical treatment. PMID- 10851694 TI - [Metaphyseal comminuted fracture. A therapeutic challenge]. AB - The most frequent localization of fractures in elderly patients are the metaphysis of the distal radius, the metaphysis of the proximal humerus and the metaphysis of the proximal femur. Displaced fractures of the distal radius should be reduced anatomically with a broad indication for surgery. A unilateral frame is the method of choice in multi-fragmentary and articular fractures although this procedure carries some disadvantages. Displaced proximal fractures of the head of the humerus should be reduced and fixed operatively. Minimal invasive procedures like tension band fixation lead to better results than fixation with plates. Non-reducible four-part fractures require primary prosthetic replacement. If standard procedures in the treatment of distal fractures of the humerus fail because of severe osteoporosis, tension band fixation may allow functional post operative treatment. Fractures of the trochanteric region of the femur can be stabilized by several dynamic standard implants that permit early weight bearing. If fixations should fail, corrective osteotomies or changing of implants--in rare cases combined with bone cement--may be necessary. Each case requires an individualized procedure in order to ensure optimal restoration of function and mobility in elderly patients. PMID- 10851695 TI - [Signals from the back. Biomechanics of the Berlin Oskar-Helene Home win the renowned 2000 Volvo Prize for Research for telemetric studies of the spine]. PMID- 10851696 TI - [Osteoporosis. New research results on bone formation]. PMID- 10851697 TI - [Endogenous and exogenous modification of tissues and their healing properties]. PMID- 10851698 TI - Repression and "repressive style". PMID- 10851699 TI - "Ruthlessness gives way to Ruth": mothering and mourning in Marilynne Robinson's Housekeeping. PMID- 10851700 TI - The wisdom of Ecclesiastes and its meaning for psychoanalysis. PMID- 10851701 TI - On healing Eve's grief. PMID- 10851702 TI - Children watching movies. PMID- 10851703 TI - [Evidence-based medicine and the elderly]. PMID- 10851704 TI - [Evidence-based medicine: an introduction for clinical practitioners]. AB - In Germany, the term "evidence-based medicine" still leads to confusion. To our continental understanding "evidence" refers to the self-evident, what is obvious and unequivocally clear without any methodological mediation. In English speaking countries, "evidence" is defined as available and disputable facts indicating whether or not a proposition is valid. In clinical medicine both types of evidence are indispensable. However at present "external (i.e. anglosaxon) evidence" from sound-evaluative clinical research is actually needed to define and justify clinical indication rules. The rationale of the concept is obviously consequentialistic, it primarily considers the clinical and community effectiveness of any medical intervention. The paper finally discusses some of the ethical problems involved in evidence-based medicine. PMID- 10851705 TI - [Guidelines and evidence-based medicine in Germany]. AB - Clinical Practice Guidelines (CPGs) are increasingly common in the German health care system. The German physicians' self-governmental body's position regarding CPGs as a tool of Evidence-based medicine (EBM) was described in a joint policy paper concerning quality in health care in late 1998. The German Medical Association (GMA) and the National Association for Statutory Health Insurance Physicians (NASHIP) stated that "the principles of EBM should be implemented into the German Health Care System" by the following means: to assess systematically and appraise critically the evidence in health care to develop evidence-based consensus CPGs for priority problems in health care to implement CPGs using graduate, post-graduate, and continuing medical education, as well as audits and CPG-based information management to evaluate quality in health care against the background of CPGs. The following paper will discuss the aims and scopes and the limits of this concept. PMID- 10851706 TI - [Evaluating geriatric rehabilitation from the perspective of medical service of the national health insurance administration]. AB - There is evidence for the community effectiveness of preventive geriatric intervention programs prior to the need for help or long-term care from a couple of prospective randomized controlled trials (RCTs). For geriatric rehabilitation programs tailored to older people with imminent or manifest need for long-term care analogous--evidence is still lacking. The nationwide leading boards of the FRG's Sickness Insurance Administration in conjunction with its Central Medical Service have formulated guidelines for the formation of a nationwide ambulatory geriatric rehabilitation service to realize the postulate of the long-term care insurance legislation "rehabilitation prior to long-term care". These guidelines must be proven empirically. To prove the effectiveness and efficiency of the foreseen ambulatory geriatric rehabilitation service, the RCT design would be highly desirable. Unfortunately, the prerequisites in the field of methodology are poor since valid, reproducible and feasible criteria for the selection of suitable patients and measurement criteria which meet the requirement of proven medium-term sensitivity to change do not yet exist. Nevertheless, there is a great and urgent need, for the first time, to investigate the cost-effectiveness ratio for this ambulatory geriatric rehabilitation service to be established in the FRG, leaving aside the methodologic desiderata of randomized control-groups. PMID- 10851707 TI - [Effectiveness of special stroke units in treatment of acute stroke]. AB - BACKGROUND: In Germany the implementation of specialized wards for the care of stroke patients is proposed. However, which type of organized inpatient stroke unit care is most effective and which group of patients will benefit most remains unclear. METHODS: Based on the analyses of the Stroke Unit Trialists' Collaboration this paper reports results of randomized and quasi-randomized trials that compared organized inpatient (stroke unit) care with contemporary conventional care. The primary analyses examined death, dependency and institutionalization. Secondary outcome measures included patient quality of life, patient and carer satisfaction and length of stay in hospital and/or institution. RESULTS: The analysis of twenty trails with 3864 patients showed a reduction in the rate of deaths in the stroke unit group as compared with the control group (OR 0.83, 95% CI 0.71-0.97). The odds of death or institutionalized care were lower (OR 0.76, 95% CI 0.65-0.90) as were death or dependency (OR 0.75, 95% CI 0.65-0.87). The results were independent of patient age, sex, stroke severity, and type of stroke unit organization. CONCLUSION: Organized care in stroke units resulted in benefits for stroke patients with regard to survival, independence, and probability of living at home. However, these results refer exclusively to Anglo-American and Scandinavian trials. German stroke unit services are organized in a different way. No data about the effectiveness of the German model is yet available. PMID- 10851708 TI - [Evidence-based medicine--problems and application to geriatrics]. AB - The use of evidence-based medicine (EBM) in the field of geriatrics faces some specific problems. Deficits in representation of older people in medical trials and the attitude of the majority of patients and their physicians considering disease as normal aging counteract successful implementation. EBM is also feasible in older patients after some adjustments without dropping the principles of the method or the preferences of patients, thus, resulting in improvement in diagnostic and therapeutic decisions. The methods are exemplified in the use of prophylactic antibiotics in endoscopic gastrostomy, a relevant geriatric problem with practical consequences. PMID- 10851709 TI - [Personality, cognitive ability and health in East and West Germany: results of an interdisciplinary longitudinal study of aging]. AB - The goal of the Interdisciplinary Longitudinal Study of Aging (ILSE) is to assess healthy and satisfying aging in middle adulthood and later life. In addition, this study compares adults from eastern and western Germany. The data for this study were collected between late 1993 and early 1996 as part of the first measurement point of the ILSE. Five-hundred and one participants from the Leipzig region were compared with 500 participants from the Heidelberg area on four specific dimensions: personality, cognitive functioning, subjective well-being, and health. Age differences were obtained for neuroticism, extraversion, openness, and agreeableness. Differences were also obtained in cognitive functioning and health. Regional differences between eastern and western Germany were obtained in the personality dimensions neuroticism, openness and conscientiousness, as well as in subjective well-being. Significant age by region interactions were obtained for openness and some of the cognitive measures, as well as for the physicians' health ratings of the participants, suggesting selective historical effects. The results are discussed with respect to functional abilities and levels of satisfaction in midlife, as well as with respect to regional differences that might be due to prior political system differences in the east and west. PMID- 10851710 TI - [Retirement as crisis or good fortune? Results of a quantitative-qualitative longitudinal study]. AB - Results of a longitudinal study about the transition to retirement are presented. 329 persons were interviewed three times (quantitative study) and 20 persons conducted a semi-structured diary (qualitative study), covering in both groups a two-year period (from half a year before retirement to one and a half years after retirement). They were asked about their subjective well-being (happiness, joy, satisfaction, stresses), their life situation and their personal, social and material resources. The results show that the transition to retirement brings with it gains (leisure time) and losses (financial, social status), but overall maintains a high level of well-being. Specific analyses point out inter individual differences, risk factors and resources, like a high level of trait well-being, health, social networks and an optimistic outlook. The diary analyses lead to specifications, more detailed process analyses and hypotheses about cohort effects. Consequences for an adequate preparation for retirement are discussed. PMID- 10851711 TI - [Coping styles, treatment satisfaction and adjustment of geriatric inpatients]. AB - 82 geriatric and 48 younger patients were investigated in two medical departments at the end of their hospital stay concerning stressors, coping styles, social support and psychic adaptation. Overall the geriatric patients seemed to be well adapted although there were many stressors. Probably this resulted from an adaptive constellation of coping styles and from high contentment with treatment. There were no large differences between the geriatric and the younger patients, nor essential changes in the three months after dismission from the hospital for these patients, who took part in the catamnestic interview. The "holistic" way of treatment, which was realized in one of the two departments, seemed to help geriatric patients cope successfully with illness. PMID- 10851712 TI - Structured treatment interruption. PMID- 10851713 TI - Understanding HIV test results. PMID- 10851714 TI - Get ready for the guide! PMID- 10851715 TI - HIV, managed care, and outcomes. AB - While it is not intended or cannot be encyclopedic, this article should provide a template for those who are starting a quality measurement program and resources where more information can be obtained. Key resources include the HIV/AIDS Bureau of the HHS, the Johns Hopkins AIDS Program, the Forum for Collaborative HIV Research, the HIV Cost and Services Utilization Study, and the growing initiative of the Infectious Diseases Society of America (IDSA). The IDSA also plans to address quality on a more global level, beyond the individual program. In a discussion of its HIV Quality Care Network, the IDSA reminds us that there is also a rationale beyond volume, practice, and outcome. As the IDSA continues its efforts at measuring outcomes, it will also develop data on cost and outcomes in different delivery systems to assist with appropriate public policy concerning the care of HIV-infected patients. It is hoped that this will allow all of us to influence public policy to improve the quality of care for HIV/AIDS patients and to achieve fair compensation for providers. Quality and outcomes measurement are important tools for managed care survival. Quality measurement also provides a link among providers, health care administrators, and payers. These data provide valuable information on program activities and effectiveness and are good for all of us, especially for patients. PMID- 10851716 TI - HE2000 corrects immune system dysregulation in HIV-positive patients. PMID- 10851717 TI - Peptide derived from cobra venom inhibits HIV infection. PMID- 10851718 TI - United Nations invests $6 million to fight the spread of AIDS. PMID- 10851719 TI - Shedding light on correctional HIV care. PMID- 10851720 TI - HIV infection among incarcerated women: epidemic behind bars. AB - Approximately 1 in 109 adult women was under the care, custody, or control of adult criminal justice authorities on any given day in 1998, the year for which the most recent statistics on women in correctional institutions are available. Of the 84,400 women who were in prison in 1998, a large percentage--37% in state facilities and 72% in federal prisons--were charged with drug-related offenses. Besides drug use, an additional determinant of HIV infection among incarcerated women may be prior exposure to physical and sexual abuse. Linkages among histories of childhood sexual abuse, physical abuse, drug use, and sex work are believed to explain the disproportionately high prevalence of HIV infection among incarcerated women. Historically, HIV services have had to compete with other demands on correctional budgets for funding and personnel time, even though the correctional health care unit is a unique and highly cost-effective access point for providing HIV prevention and care for high-risk populations of women. Coalition building between correctional staff and medical staff (and, in some cases, departments of public health) has enabled some correctional institutions for women to establish outstanding programs for HIV-infected women. By diagnosing HIV and instituting a plan for treatment, correctional facilities for women can play a critically important role in the reduction of morbidity and mortality among HIV-infected women in high-risk populations. PMID- 10851721 TI - Evaluation of simplified protease inhibitor dosing regimens for the treatment of HIV infection. AB - Complicated dosing regimens can unfavorably affect patient adherence and drug efficacy. In an effort to increase adherence while maximizing the benefits of the protease inhibitor (PI) component of HAART, the efficacy and safety of less frequent PI dosing regimens have recently been under scrutiny. Limiting the number of drugs required in antiretroviral therapy regimens may also minimize toxicity and drug-drug interactions. This article reviews the current movement toward twice-daily regimens and examines the efficacy and safety data available on twice-daily dosing of amprenavir, indinavir, nelfinavir, saquinavir soft-gel capsules, and ritonavir. Future trends in dosing are also discussed. PMID- 10851722 TI - Gynecomastia and HAART. PMID- 10851723 TI - [Periurethral sphincter hyperactivity and sacral neuromodulation]. PMID- 10851724 TI - [Abnormal glycosylation of soluble ABH antigens in tumors of the urinary tract]. AB - OBJECTIVE: To determine the secretor expression in patients with bladder cancer, adenoma of the prostate and normal subjects. METHODS: The secretor character was determined in saliva of normal subjects (n = 40), patients with bladder cancer (n = 61) and adenoma of the prostate (n = 44) by the technique of hemoagglutination inhibition. RESULTS: 80% of the normal subjects were found to be secretors, which is in agreement with the data reported in the literature. Only 23 (37.71%) of the patients with bladder cancer were secretors and 24 (54.54%) of the patients with prostate adenoma expressed the secretor gene. CONCLUSIONS: The expression of soluble antigens decreased in patients with bladder cancer or prostate adenoma in comparison to the normal subjects. Deletion of ABH antigens in the membrane of tumor cells has been reported in other studies. This lack of expression results from a genetic alteration in the clones involved in tumor pathology. The decrease in soluble antigens in the patient groups analyzed might be due to the same mechanism of genetic alteration that could involve non tumor tissues. Most of the cancers in humans originate in epithelial cells and the changes in blood group antigens constitute an important aspect in tumor immunology. PMID- 10851725 TI - [Artificial urinary sphincter]. AB - OBJECTIVE: To review the literature on artificial urinary sphincters, to describe their function, technique of implantation, indications, results and complications, and to analyze the possible utility of the prosthesis in the treatment of stress incontinence. METHODS: Medline (Index Medicus Online) and Embase (Excerpta Medica Online) were accessed to review the literature on artificial urinary sphincters published from 1974 (when the first artificial urinary sphincter was described by Foley) to October 1999. Of 322 articles identified, only those that described the patient selection criteria, type of prosthesis utilized, results and complications, and mean follow-up were considered. Articles describing historical aspects and new prototypes were also reviewed. Papers by the authors of the present article were excluded to avoid the bias of author preference. The bias of the language barrier, which occurs when articles published in Spanish, English and French are reviewed, was minimum. RESULTS: The AMS-800 is the only model available today. The results achieved are excellent if the indication is correct and perioperative management is careful and exact. The ideal candidate is one with genuine stress urinary incontinence and normal bladder function, although hyper or hyporeflexia is not an absolute contraindication if corrected before, during or after insertion of the prosthesis. The surgical technique is relatively simple and the only difficulty consists in the choice of the appropriate cuff and reservoir. The complications include urethral atrophy, erosion, infection and bladder instability, and are less frequent in women with stress urinary incontinence type III and in men incontinent after prostate surgery, and more frequent in patients with incontinence following pelvic trauma, incontinence due to congenital malformation and those with a neurogenic bladder. The mechanical failures of the prosthesis have diminished with its improved design. New hydraulic and non-hydraulic prototypes have been designed to reduce the complications, but the results are as yet unavailable. CONCLUSIONS: Today, patients with stress urinary incontinence have more possibilities to recover continence. If incontinence persists after all the available medical and surgical options have been attempted, one possibility still remains: the artificial urinary sphincter. PMID- 10851726 TI - [Impact of prostatic symptoms in patients with prostatic benign hyperplasia]. AB - OBJECTIVE: To analyze the impact of prostatic symptoms and to identify the factors associated with the problems caused by these symptoms. METHODS: A descriptive, transverse study was conducted on 133 patients. The problems arising from prostatism were analyzed by means of a self-administered questionnaire, using the Symptomatic Prostatic Index (SPI), which was compared with the International Prostate Symptom Score (I-PSS), uroflowmetry, morbidity, medication required and sociodemographic variables. The reliability and consistency of the SPI scale were analyzed and the variables associated with a greater impact of prostatism were determined by linear regression analysis. RESULTS: Patient mean age was 68.8 years. Mean scores were 20.1 and 4.75 for the I-PSS and IQL item, respectively. Mean and maximum urinary flow were below the 50th percentile in 95.2%. A high correlation was found between the items of the SPI questionnaire and between the items and the total score. The SPI scale showed a high discriminating power (delta = 0.95) and internal consistency (alpha = 0.82), and factorial analysis showed only one factor accounted for the 49.05% total variance. The SPI questionnaire score was 15.5 and involved the irritative symptoms, basically nocturia, proportionally more than the obstructive symptoms (p < 0.0001). A direct correlation was found between the higher SPI score and the severity of the prostatic symptoms (p < 0.0001) and, consequently, a worse quality of life (p < 0.0001). Younger patients tolerated the symptoms poorly (p = 0.002). Linear regression analysis confirmed that tolerance was worse in the younger patients with more severe symptoms and no other disease (r2 = 0.43, p < 0.0001). CONCLUSIONS: The impact of prostatism increases according to its severity, particularly for the irritative symptoms, basically nocturia. Psychological factors may probably affect the variability of patient tolerance, indicating that the decision for treatment of BPH might be based on the problems it may cause and the impact on the quality of life more than on the severity of the symptoms. PMID- 10851727 TI - [Clinical significance of prostatic intraepithelial neoplasia]. AB - OBJECTIVE: To determine the value of high grade prostatic intraepithelial neoplasia (with or without the influence of certain risk factors) in predicting prostate cancer in subsequent biopsies. METHODS: The study comprised 41 patients from a prostate cancer screening program with high grade prostatic intraepithelial neoplasia. Subsequent biopsies were reviewed and the probability of detecting prostate cancer was calculated. We analyzed the influence of age, DRE and transrectal US findings, PSA levels and PSA density on the results of the repeat biopsies. RESULTS: The patients were aged 50 to 83 years (mean 62.8 +/- 1.6 SD, median 61). Only 27 of the 41 patients with high grade prostatic intraepithelial neoplasia accepted a repeat biopsy. Of these, prostate cancer was demonstrated in 11 (40.7%; all cases were clinically localized at the time of diagnosis) and 16 showed no changes (59.3%) on repeat biopsy. By univariate and multivariate analysis, patient age, DRE and transrectal US findings, PSA levels and PSA density were not found to be predictors of cancer in the subsequent biopsies. CONCLUSIONS: The finding of high grade prostatic intraepithelial neoplasia in the prostate biopsy carries a high probability of detecting cancer in subsequent biopsies. We therefore advocate performing a repeat biopsy in these patients. PMID- 10851728 TI - [Telomerase activity as marker of superficial tumor of the bladder]. AB - OBJECTIVE: To determine the telomerase activity in urine samples of patients with superficial bladder cancer, its sensitivity and specificity as a tumor marker, and possible implications for prognosis, diagnosis and therapeutic efficacy. METHODS: Urine samples of 50 patients with superficial bladder cancer were analyzed. Telomerase activity was determined by TRAP (telomeric repeat amplification protocol). Standard urinary cytology, urine culture and urinary sediment analysis were performed, as well as a pathological analysis of the surgical specimen. RESULTS: 84% of the patients were positive for telomerase activity and only 52% had a positive cytology. Telomerase activity showed a sensitivity and specificity of 73.6% and 92.7%, respectively, versus 53.2% and 81.8% for urinary cytology. The degree of cell differentiation and, to a lesser extent, bladder wall infiltration showed differences in comparison with conventional urinary cytology. CONCLUSIONS: Telomerase activity can be determined in voided urine samples of patients with superficial bladder cancer. It has a higher sensitivity and specificity than conventional urinary cytology and is a good marker for diagnosis and follow-up of these patients. PMID- 10851729 TI - [P185 (Neu) oncoprotein in the prognosis of bladder carcinoma. Experience of 5 years]. AB - OBJECTIVE: To determine the utility of p185 oncogene in the biological characterization of transitional cell carcinoma and in the prediction of recurrence, and to analyze survival at 5 years mean follow-up. METHODS: A prospective clinical cohort study was conducted on 81 patients. Tissue specimens were obtained between November 1992 and November 1993. The study comprised two groups: nontumoral bladder tissue specimens from 20 patients (group I) and tissue specimens from 61 patients with bladder carcinoma (group II). p185 expression was determined by enzyme immunoanalysis (EIA). A statistical analysis of the results was performed. RESULTS: p185 oncoprotein levels were higher in patients with recurrence (1098.97 HNU/mg protein vs. 924.54 HNU/mg). Although higher levels of p185 were found in the patients that had died vs those who are alive, the differences were not statistically significant for overall survival or stratification by tumor grade or infiltration (p = 0.556; ns). CONCLUSIONS: Determination of p185 oncoprotein was found to be useful in the prediction of tumor recurrence at 5 years mean follow-up. PMID- 10851730 TI - [Metachronous bilateral Wilms' tumor]. AB - OBJECTIVE: To report on a female patient with metachronous bilateral Wilms' tumor with a long survival. METHODS: An 11-year-old girl underwent total nephrectomy when she was 4 years old for Wilms' tumor of the right kidney. She received cobalt therapy and chemotherapy, and the evolution was good for three years, at which time a control US scan detected a tumor in the left kidney that was diagnosed as Wilms' tumor. She underwent a partial nephrectomy at the Hospital Pediatrico de Centro Habana (Cuba) and received chemotherapy. RESULTS/CONCLUSION: A recent US scan and a descending pyelogram showed completely normal findings and growth of the remaining renal parenchyma. The patient is alive and well 7 years and 8 months later. PMID- 10851731 TI - [Acute lobar nephronia. Report of a pediatric case]. AB - OBJECTIVE: A case of lobar nephronia in a child is presented. METHODS: Herein we describe a case of acute lobar nephronia in a 10-year-old boy. The clinical, diagnostic and therapeutic aspects are discussed. RESULTS/CONCLUSIONS: Acute lobar nephronia or acute focal bacterial nephritis is an uncommon form of pyelonephritis that can affect both adults and children, although few cases have been reported in children. Imaging techniques are necessary for diagnosis and to distinguish it from other conditions, such as abscess or renal masses that require a different treatment. Ultrasound is the imaging technique of choice in the diagnosis and follow-up of lobar nephronia. PMID- 10851732 TI - [Treatment of urethral stenosis with thermo-expandable prosthesis "Memotherm". Our experience]. AB - OBJECTIVE: To report our experience in the treatment of recurrent urethral stricture in the male with the Memotherm heat-expansible stent. METHODS: From December 1995 to March 1999, the Memotherm heat-expansible stent was utilized in 4 patients with urethral stricture that had undergone urethrotomy procedures, periodic urethral dilatation and in those cases with post-traumatic stricture, open surgery for urethroplasty and urethral reattachment. All patients had multiple recurrences of the urethral stricture that was not amenable to the treatments utilized, therefore the intraurethral stent was inserted. RESULTS: All patients had a good postoperative course with unhampered voiding and ample stream. The patients were followed in the outpatient setting by a 6-monthly assessment of micturition and a yearly endoscopic control evaluation to detect hypertrophic growth of the urethral mucosa and/or intraluminal calcification. All the stents were completely enveloped in the urethral wall 12 months after insertion. The only side effects observed was limited postvoid leaking during the first few months following insertion of the prosthesis and one case of transient hemospermia. CONCLUSIONS: The Memotherm intraurethral heat-expansible stent is a valid treatment option for selected cases of recurrent urethral stricture. Although this approach can be utilized before performing urethroplasty, which is often a complicated technique with uncertain results, currently it is a valid treatment option in case of failure. PMID- 10851733 TI - [Extra-adrenal pheochromocytoma. Report of a case]. AB - OBJECTIVE: To report a case of extraadrenal pheochromocytoma with special reference to the diagnostic and therapeutic aspects. METHODS: A case of extraadrenal pheochromocytoma in a 20-year-old female with severe hypertension is presented. The clinical and biochemical aspects are reviewed with special reference to the diagnostic imaging methods. RESULTS: Plasma noradrenaline and urinary normetanephrine levels were elevated. CT and MRI showed a well-defined mass, 6 cm in diameter, adjacent to the left kidney, abdominal aorta and psoas muscle. Angiography demonstrated a high vascularization in the area of the tumor. MIBG scintigraphy revealed a well-defined mass, but no other distant lesions. Surgical treatment was performed with preoperative alpha and beta adrenergic blockade. Currently the patient has a normal blood pressure and catecholamine levels, with no evidence of lesions on the MIBG scintiscan. CONCLUSIONS: Plasma catecholamine and urinary normetanephrine levels levels confirmed the presumptive diagnosis of pheochromocytoma. MIBG is the technique of choice for the localization of the mass and suspected metastases. CT, MRI and angiography demonstrated the anatomic relationships of the tumor. The best results are achieved with complete resection and preoperative adrenergic blockade. PMID- 10851734 TI - [Ileovesical fistula in Crohn's disease]. AB - OBJECTIVE: To discuss the clinical presentation, complementary evaluation procedures and treatment of ileovesical fistula, an uncommon complication of Crohn's disease. METHODS/RESULTS: After the clinical presentation, complementary evaluation procedures were performed to confirm the diagnosis in all cases. Treatment was based on the patient's general condition; surgery was performed in two cases and one case was carefully followed. CONCLUSIONS: Ileovesical fistula is an uncommon complication of Crohn's disease. Occasionally, the urological symptoms may precede the digestive symptoms, therefore this condition should be suspected particularly if the complementary evaluation procedures are not very sensitive. Surgery is the treatment of choice. PMID- 10851735 TI - [Primary retroperitoneal hydatid cyst]. AB - OBJECTIVE: To review the pathogenesis, clinical features and treatment of a rarely observed site of presentation of hydatid cyst. METHODS/RESULTS: A case of an isolated retroperitoneal hydatid cyst is presented. Diagnosis was based on the immunodiagnostic test and radiological examination. Treatment was by surgery. CONCLUSIONS: Isolated retroperitoneal hydatic cyst is rare. The abdominal mass is the most frequent sign, and less frequently, concomitant compressive syndrome. Diagnosis is based on the biological examinations, percutaneous puncture and radiological findings. Treatment is by surgery. Azoles are useful for intraoperative ruptured cysts and to reduce the risk of recurrence. PMID- 10851736 TI - [Epididymal ectopic adrenal tumor]. AB - OBJECTIVE: A case of a nonfunctioning ectopic adrenal tissue tumor in the epipidymis is described. METHODS/RESULTS: A case of a nonfunctioning ectopic adrenal tissue tumor in the epipidymis is presented. A left testicular mass had been incidentally detected in this patient. CONCLUSIONS: Tumors of this type localized in the juxtafuniculogonadal region are usually benign. However, resection and histological analysis are always indicated in order to detect metastasis or confirm the histiological type is normal. PMID- 10851737 TI - [Epididymal carcinoma. Bibliographic review in reference to a case]. AB - OBJECTIVE: To describe a case of epithelial cell paratesticular carcinoma of the epididymis and briefly review the literature on this tumor type. METHODS/RESULTS: A 69-year-old male consulted for a testicular mass and intrascrotal pain, together with irritative bladder symptoms. The patient underwent orchidectomy, but consulted again shortly thereafter for persistent irritative bladder symptoms. A TUR biopsy of the bladder wall demonstrated undifferentiated carcinoma arising from the epididymis. The patient did not respond to chemotherapy. He developed systemic metastasis and died 4 months after the diagnosis. CONCLUSIONS: Carcinoma of the epididymis is a rare malignant paratesticular tumor arising from the epithelial cells with a very poor prognosis. Its clinical features are unspecific and this tumor type should be taken into account when making differential diagnosis from intrascrotal masses arsing from other causes. Due to the rarity of this disease, it has not been possible to identify treatments that might achieve better results. PMID- 10851738 TI - [Fournier's gangrene after vasectomy]. AB - OBJECTIVE: An uncommon case of Fournier's gangrene following vasectomy is described. METHODS/RESULTS: A 35-year-old male with no remarkable previous history, who underwent vasectomy in another hospital, developed a clinical picture compatible with Fournier's gangrene 7-8 days later. The patient required wide, aggressive surgical debridement on several occasions with broad spectrum antibiotic coverage. After a long stay at the hospital, the patient was finally discharged and referred to another hospital for plastic surgery. CONCLUSIONS: Fournier's gangrene is a polymicrobial infection of the perineoscrotal region that manifests as a rapidly progressive necrotizing fasciitis. Most of the cases have a predisposing and/or triggering factor. Fournier's gangrene following vasectomy is uncommon. The morbidity and mortality in this severe complication depend on early diagnosis and aggressive surgical management. PMID- 10851739 TI - Traumatic rupture of angiomyolipoma: a case report. AB - OBJECTIVE: To describe a case of traumatic rupture of renal angiomyolipoma (AML). METHODS: The images and clinical data of the present case are presented. RESULTS: A rare case with exuberant clinical presentation of a perirenal hematoma resulting from traumatic rupture of renal AML is presented with a brief review of the role of ultrasound (US) and body-CT in the diagnosis of this pathology and its complications. CONCLUSIONS: Whenever there is a collection detected by US in the various anatomic renal spaces, in a patient with flank pain and low hemoglobin shortly after abdominal trauma, it is advisable to perform abdominal CT and search for a hematoma. Small amounts of fat, detected by US and body-CT, may lead to the diagnosis of an underlying AML that can rupture, even in the case minor forces are applied to the kidney. PMID- 10851740 TI - Environmental radon daughters reveal pathognomonic changes in the brain proteins and lipids in patients with Alzheimer's disease and Parkinson's disease, and cigarette smokers. AB - This paper presents an investigation of the retention of environmental radon daughters, 210Po (alpha particle emitting radio-nuclide) and 210Bi (beta particle emitting radio-nuclide), in lipid and protein fractions of the cortical grey and subcortical white matter from the frontal and temporal brain lobes of patients who had suffered from Alzheimer's disease or Parkinson's disease, of cigarette smokers, and of control subjects. 210Po and 210Bi radioactivity increased tenfold in the cortical grey and subcortical white protein fraction in patients with Alzheimer's disease and smokers, and tenfold in the cortical grey and subcortical white lipid fraction in patients with Parkinson's disease. Free radicals generated by radon daughters may add to the severity of the radio-chemical injury to the brain astrocytes. The pathognomonic distribution of radon daughters to lipids in patients with Parkinson's disease and to proteins in patients with Alzheimer's disease was attributed to high chlorine affinity of radon daughters. The changes in the membrane protein pores, channels, and gates in patients with Alzheimer's disease and in the lipid bilayer in patients with Parkinson's disease are at the core of what the authors think are two systemic brain diseases. PMID- 10851741 TI - Occupational physical demands and hip osteoarthritis. AB - The authors investigated the influence of physical strain at work on radiological signs of hip osteoarthritis. The study included 295 men and 298 women aged over 45 from an urban area who were classified in four groups according to physical demands of their occupation. The evaluation included clinical and radiological signs of hip osteoarthritis. The association between hip osteoarthritis and occupation was analysed using logistic regression. Though not significantly, radiological signs of hip osteoarthritis were common in subjects who worked in a standing position (odds 1.45 for men, 1.50 for women). Clinical signs of osteoarthritis in women were significantly associated with performance in a standing position (odds 3.00), whereas in men the association was more significant for jobs with high physical strain (odds 2.19). There was a sustained trend toward an increase in health risk with years of work in all job categories. Occupation did not appear to influence the development of radiological hiposteoarthritis, but the authors did establish association between clinical signs of hip osteoarthritis and work. PMID- 10851742 TI - Short-term neurobehavioural effects in anaesthetists with low exposure to nitrous oxide. AB - The aim of this study was to assess whether a sample of 37 anaesthetists occupationally exposed only to N2O showed any deterioration in vigilance and/or mood. The anaesthetists were examined with three neurobehavioural tests (Simple Reaction Time and Colour Word Vigilance to measure the vigilance and Mood Rating Scale to evaluate the level of stress and arousal) and underwent N2O biological monitoring (to correlate the test results with the N2O exposure) on the first and on the last day of the work week, before and after work in the operating room. No significant relationship was found between the biological monitoring and the test results. The only significant statistical difference was found between the beginning and the end of each workday in the arousal level, regardless of the result of the biological monitoring. PMID- 10851743 TI - Evaluation of the impingement of the pronator muscle in occupational carpal tunnel syndrome by electromyographic and ultrasonographic techniques. AB - Normal flexion of the fingers involves an involuntary contraction of the pronator and lumbrical muscle. In individuals whose profession involves constant flexing of the fingers those muscles become hypertrophied, impinging on the carpal tunnel. The narrowing of the carpal tunnel yields well to ultrasonography. The objective of this investigation was to find an ultrasonographic index of occupational carpal tunnel syndrome. Thus "Index M" denotes the variation obtained in the "M Space" before and after flexion-extension of the fingers. The study included 45 subjects performing tasks which involved the risk of cumulative trauma disorders. The subjects were tested using the electromyography and ultrasonography. The method was based on relation between the decrease in conduction of the median nerve measured by electromyography and the ultrasonographically measured variation of "M Space" in terms of sensitivity and specificity. The sensitivity of ultrasonography was 85%, as it confirmed the pathologic findings determined by electromyography ("M Index" positive) in twenty two out of 26 hands, but the specificity was not statistically significant. Ultrasonography seems to have found very important application as a screening technique in occupational medicine. It is non-invasive, sensitive, easily repeated, and costs little. PMID- 10851744 TI - [Shiftwork and quality of life]. AB - The aim of this study was to compare the quality of life of shiftworkers and non shiftworkers. Satisfaction with various aspects of life and the overall satisfaction were examined by means of the Quality of Life Scale. A total number of 107 chemical industry workers participated in the study, of whom 56 worked shifts and 51 worked regular hours. The results revealed that the average satisfaction with the present job and financial status was lower in shiftworkers than in the non-shiftworkers (P < 0.05). The differences in the two predictors of life quality did not affect the overall satisfaction with life in either group. PMID- 10851745 TI - Studies of psychophysiological and temporal conditions of work. AB - The main areas studied until the 70s were fatigue and rest. Many tests for fatigue were examined, and studies failed to find their practical use. Fatigue grows according to the positive acceleration curve while recovery grows according to the negative acceleration curve. Different mechanisms are involved in intelligence test performance when subjects are fatigued and when rested. Contrary to the pharmacological, psychological stimulants increase performance without adverse effects on the organism. The differences in absenteeism between male and female workers are related to family duties. After the 70s, research was focused on shiftwork. Shiftwork is associated with imposed change in a worker's normal activity pattern. It brings about fatigue, negative moods, and impaired health, sleep, safety, and working capacity. The shiftwork tolerance is connected with introversion, neuroticism, morningness, control of behavioural arousal, and parameters of circadian rhythms. The most important predictors of shiftwork tolerance are dimensions of control of behavioural arousal and morningness, while the most important criterion is sleep quality. PMID- 10851746 TI - Bibliographic output of the Institute for Medical Research and Occupational Health in Zagreb between 1994 and 1998. AB - This paper brings a classification of the bibliographic output of the Institute for Medical Research and Occupational Health over the period 1994-1998 into fourteen main categories according to the type of publication and its coverage in different bibliographic databases. The academic staff was classified according to scientific fields in which they received the bachelor's degree and in which they were appointed into a scientific grade. The authors compared the Institute's scientific performance in the last five years to previous periods and with achievements of similar institutions in Croatia. Regardless of a large decrease in the Institute's personnel, the number of scientists with a Ph.D. degree remained unaltered. The ratio between published papers covered by Current Contents and the number of scientists holding a Ph.D. degree slightly dropped, while the ratio between the publication of conference abstracts and Ph.D.s doubled. PMID- 10851747 TI - The destiny of manuscripts submitted for publication in the Archives of Industrial Hygiene and Toxicology over the past 21 years. PMID- 10851748 TI - [Rural medicine: a view to the future]. PMID- 10851749 TI - [Self-perception of health and mortality in elderly from a rural community]. AB - OBJECTIVE: To assess the relationship between self-perception of their health and mortality in a representative sample of persons over 65 in a rural community. DESIGN: Survival study of population with three years observance. SETTING: A non coastal and rural borough in Galicia. PARTICIPANTS: 408 people over 65, chosen by random sampling. MEASUREMENTS AND MAIN RESULTS: The demographic, social and health (objective and subjective) parameters were determined through an initial survey, with subsequent follow-up to find the date and cause of death should this have occurred. Of the 404 elderly people observed, 67 (16%) died. The main causes of death were diseases of the circulatory apparatus (48%) followed by tumours. Mortality was higher in people who were older, unmarried, didn't drink alcohol, were seriously ill, had restricted mobility, consumed more medicines and had a perception of their health as poor. We found an association between self perceived health and mortality, after adjustment for age, sex, marital status, educational level and consumption of medicines, only when we looked at the elderly without restricted mobility (RR = 2.3; 95% CI = 1.0-5.3). CONCLUSIONS: We think that self-perception of health status can be an overall indicator of health, linked to mortality, and that in the elderly with good mobility, this association can be independent of age, sex, educational level, marital status and consumption of medicines. PMID- 10851750 TI - [Relationship between quality and cost of the drug prescription in primary care]. AB - OBJECTIVES: With a system of qualitative indicators, to analyse the pharmaceutical prescription of general practitioners (GPs), and to evaluate the relationship of these indicators to the overall pharmaceutical prescription expenditure per inhabitant. DESIGN: Retrospective descriptive study. SETTING: Primary care. MEASUREMENTS AND MAIN RESULTS: The drugs prescription of 285 GPs from 32 primary care teams was evaluated, with the individual prescription of each doctor as the unit of analysis. The prescription was classified in 3 categories according to its intrinsic value (IV): low (< or = 75%), medium (76 79%) and high (> or = 80%). Selected as tracers of over-prescription were: daily dose per inhabitant (DDI) of antibiotics (AB), DDI of non-steroid anti inflammatory drugs (NSAID), and DDI of ulcer drugs (ULC). Selected as tracers of selection were: % DDI third-generation cephalosporins/DDI total cephalosporins; % DDI broad-spectre quinolones/DDI total quinolones; and % DDI NSAID/DDI NSAID plus analgesics. Quantitative indicators studied were: total expenditure per allocated population, cost per drugs prescription of doubtful efficacy, and cost per daily dose of AB, NSAID and ULC. Variance analysis, including the Scheffe test for multiple comparisons and Pearson's linear correlation, was applied. 26% of the prescriptions had an IV below 75%, and 34% had an IV above 80%. The means of DDI of AB among the categories of IV were different (p < 0.0001), as were those of DDI of NSAID (p < 0.0001) and of ULC (p = 0.007). Lower consumption of AB, NSAID and ULC was found in prescriptions with the highest IV %. The third-generation cephalosporins and the NSAID + analgesics showed significant differences in the three IV categories (p < 0.0001 and p = 0.041), unlike broad-spectrum quinolones (p = 0.18). The total expenditure per allocated population was less for GPs whose prescriptions had the highest IV %; whereas the cost per prescription and cost per daily dose showed no significant differences for IV categories. CONCLUSIONS: The doctors with the best qualitative profile on these indicators had less expenditure per inhabitant. However, no differences were found in the cost per prescription or cost per treatment between doctors. Therefore, interventions must prioritize improving drug prescription quality rather than just promoting changes to lower-cost drugs. PMID- 10851751 TI - [Type 2 diabetes mellitus: incidence and diagnosis at a primary care center]. AB - OBJECTIVE: To find the annual incidence and reasons for type-2 Diabetes Mellitus (DM2), and the methods used to diagnose it, on the basis of the validation of a computerised record for 1991-1995. DESIGN: Retrospective, longitudinal study. SETTING: Primary care centre. MEASUREMENTS AND MAIN RESULTS: Of the 387 diabetics registered as new cases in the 1991-1995 period out of 17031 people over 14 who were seen, 21 were not diabetics, 60 were cases of late diagnosis or late recording, 75 came from another centre and 9 were type-1. All these were excluded. The mean age of the 222 (57.4%) real new cases was 59 (ED 11.4). 53% were women. The most common causes of diagnosis were the existence of previous with diagnostic hiperglycemia (50.9%) and the application of protocols for other cardiovascular risk factors (19.8%). The diagnostic methods were two basal glucaemias > or = 140 mg/dl (70.7%), 1 glucaemia > or = 200 mg/dl with typical clinical picture (6.7%) and oral overload of glucose (23%). 97% of cases were diagnosed at the centre itself. The density of annual incidence was 30.1 per 10,000 inhabitants. Prevalence at start and end of the study was 4.4 and 4.9%. CONCLUSIONS: The incidence and prevalence described are greater than described in other studies. The most common reasons for diagnosis were the existence of previous nondiagnostic hyperglycemia and the application of protocols for other risk factors. PMID- 10851752 TI - [Assessment of accessibility to health services facilities]. AB - OBJECTIVE: To assess health care access integrating the availability of resources, medical institution and the patient point of view. SETTING: Nuevo Leon, Mexico. MEASUREMENTS AND MAIN RESULTS: A random sample of patients were interviewed about their perceptions on different barriers, which also were assessed for the institution utilizing the corresponding indicator. Availability of resources were also measured for every primary and secondary medical care unit of the greatest Mexican health care system in Nuevo Leon. It was observed a 70% access; 70% for primary care and 73% for secondary care. Availability of human resources was an important factor but barriers as observed by the institution were the most important (waiting time and traveling cost). Barriers were rated different by the institution and the patient. CONCLUSIONS: The combination of institutional barriers, patient barriers and resources for assessing health care access is discussed. PMID- 10851753 TI - [Hysteria epidemics, epidemic conversion disorder or epidemic somatotrophic disorders?: a new case of a fact for the 21st century]. AB - OBJECTIVES: To describe a new outbreak of "mass hysteria" or "epidemic conversion disorder" occurring in Barcelona in 1997. Based on this outbreak's features differentiating it from other similar outbreaks described in the literature or treated by the same team, to propose a change in the theoretical framework for these group somatomorphic phenomena. DESIGN: A study describing clinical epidemiological research and the interventions performed. SETTING: Urban health centre. Epidemiological research and data analysis performed by the Municipal Health Institute (MHI) of Barcelona. INTERVENTIONS AND MEASUREMENTS: The following were performed: a) Diagnostic screening of population. b) The usual epidemiological surveys of the MHI. c) Semi-structured interviews with the quarter's care professionals. d) Two group sessions with the quarter's health professionals. MHI experts analysed statistically the data provided by measurements b) and c). RESULTS: 276 people (42% of the population of the quarter) were studied. The attack rate was 10%. Data analysis noted that communication of the fantasy of the outbreak of scabies facilitated self diagnosis and even diagnosis by health professionals. CONCLUSIONS: Given the frequency and the social and health implications of this kind of somatomorphic disorder, both in the developed countries and those "on the road to development", clinical researchers and epidemiologists should not wait to pose the possibility of hysteria until after the "organic" aetiology of an epidemic outbreak has been completely discounted. The second series of conclusions points towards the incorrectness of defining all these outbreaks as mass hysterical disorders. In reality, we should be thinking, more openly and in a more scientifically modern way, of epidemic somatomorphic disorders of various kinds. The outbreak described in the current study could be referred to as the hypochondriform variety of outbreak, whereas those described elsewhere by the same team could be understood as the conversion variety. PMID- 10851754 TI - [Intervention for alcoholism control among chronic drinkers in primary care]. AB - OBJECTIVE: To evaluate the evolution of alcohol consumption in chronic drinkers after a primary care alcohol intervention over two years. DESIGN: Prospective intervention study. SETTING: Urban primary care centre. PATIENTS: Males between 20 and 60 years old who consumed 100 or more grams of alcohol per day for at least the previous two years. MEASUREMENTS AND MAIN RESULTS: Detoxification (out patient or hospital according to the degree of dependency) and habit-breaking, which consisted of psychological support (techniques of brief counselling, brief motivating interview) and/or referral to the care and observance centre (COC) for individual and/or group psychotherapy, took place. A minimum of eight visits were programmed and consumption was assessed in gr/day at the start and at one (1 m), three (3 m), six (6 m), twelve (12 m), eighteen (18 m) and twenty-four months (24 m). 64 out of 129 (49.5%) responded to the appointment. Control visits: nil observance (0 visits) 6 patients (9%), and excellent observance (> or = 9 visits) 21 patients (33%). INTERVENTIONS: Psychotherapy treatment: psychological support 55 patients (86%), referral to the COC 4 (6%), group psychotherapy 2 (3%), and non-treatment 6 (9%). Data on mean consumption: start 131 g/day (SD = 52), 12 m 31 (SD = 41), and 24 m 38 (SD = 42). Mean reduction of consumption at 24 months according to the observance: insufficient -19%, acceptable -71.1%, excellent 83.9% (p = 0.001). CONCLUSIONS: Low response to appointments. Evolution of alcohol consumption was similar to that in other studies. Clear relationship between number of visits and consumption at the end of the study. Given the positive findings, we think a primary care intervention on alcohol is essential. PMID- 10851755 TI - [Comparative study of pneumotonometer and Goldmann tonometer for screening high intraocular pressure in primary care]. AB - OBJECTIVE: Evaluation of reliability of measurements of IOP obtained with air puff noncontact tonometer respect the obtained with the conventional Goldmann tonometer. As well, analysis of the possibility of a higher difference between the measurements by both methods respect the fact of being myope or hypermetrope. DESIGN: Comparative study of two measurement methods. SETTING: General ophthalmology clinic and primary care clinic of our sanitary area. PATIENTS: Aleatory sample among the patients who went to the general ophthalmology clinic for any cause from the first of May to the thirtieth of June of 1996. The patients with predisposing processes for glaucoma were excluded, as well as those who suffered ocular surgery or those who took drugs which influenced the IOP. INTERVENTIONS: The ophthalmologist made three measurements of IOP in each eye and the sight of all the patients were tested. The arithmetic average was made among every three measurements. The physician of primary care made three measurements of IOP in each eye with the air-puff noncontact tonometer, and the arithmetic average was made. RESULTS: 81 patients were included, from whom 7 presented high IOP. The sensibility of the air-puff noncontact tonometer, compared with Goldmann tonometer, was 86% (95% CI, 18.20-99.63%), and the specificity 84% (95% CI, 89.66 78.08%). The air-puff noncontact tonometer obtained measurements between 1.116 and 2.008 mmHg higher than the Goldman tonometer. This difference, worthless from the clinic point of view, didn't find a relationship with the fact of being myope or hypermetrope. It was found a positive lineal association between the measurements made by both methods, with a correlation coefficient of 0.8086 (p < 0.001, 95% CI, 0.7476-0.8560). CONCLUSIONS: The results obtained are similar to the ones of other published series. A tendency of higher measurements with the air-puff noncontact tonometer is observed. It hasn't been observed a higher difference between both methods in myopes. The air-puff tonometer is a valid reliable technique to be used in primary care, it is easy to use, it doesn't transmit infectious illnesses, and it isn't necessary to use anaesthetic or staining eyedrops. PMID- 10851756 TI - [Knowledge of care activity: another professional factor associated with utilization?]. AB - OBJECTIVES: To analyse the doctors' level of understanding of the use of resources in primary care and the relationship of their understanding to the proper use of resources. SETTING: Primary care in the Autonomous Community of Murcia. DESIGN: Descriptive, crossover study. RESULTS: There was little concordance between the real data contributed by the unit of information and registration of primary care management and data calculated by the doctors themselves. The best-known parameters are frequency of attendance (kappa index = 0.11) and patient pressure (kappa = 0.39). The profile of the doctors did not in general relate to this degree of understanding; it was only observed in a tendency in older doctors (p < 0.001) to overvalue frequency of attendance. The doctors with less understanding used radiology resources more with 164 requests per 1000 inhabitants per year, made more specialist referrals (826 per thousand inhabitants per year) and prescribed more medicines (21 prescriptions per inhabitant per year). Likewise, doctors who used resources most tended to underestimate their use of resources (p < 0.001). CONCLUSIONS: The doctor's understanding of his/her health-care activity and of the resources it involves is scant and independent of the doctor's professional characteristics. This is a factor, which we could call the professional factor, clearly related to resource use. PMID- 10851757 TI - [Tailored guidelines on publishing in scientific journal for qualitative research]. PMID- 10851758 TI - [Bioethics and family medicine (and V). Working Group of the semFYC]. PMID- 10851759 TI - [Factors explaining drug expenditure in primary care]. PMID- 10851760 TI - [Educational intervention targeted to caregivers of elderly dependents]. PMID- 10851761 TI - [Arterial hypertension and anti-influenza agents]. PMID- 10851762 TI - [Digestive system endoscopy and primary care: experience with direct demand of endoscopy by the family physician]. PMID- 10851763 TI - [Hypersensitivity reaction to enoxaparine]. PMID- 10851764 TI - Hantavirus pulmonary syndrome in Canada, 1989-1999. PMID- 10851765 TI - Outbreak of mumps, Montreal, October 1998 to March 1999--with a particular focus on a school. PMID- 10851766 TI - Use of sunscreen in health care professionals. The health belief model. AB - The purpose of this study was to examine the relationship among perceived susceptibility, demographic variables, and use of sunscreen in health care providers, using the Health Belief Model Questionnaire. The sample consisted of 90 participants from the Southeast, ages 24 to 60 years. This sample included nurses, pharmacists, psychologists, nurse practitioners, and physicians. Demographics revealed that 63% had postgraduate or professional degrees, 91% were white, 88% were women, 4% had a family history of skin cancer, and 4% had a personal history of skin cancer. Participants at actual high risk and those at actual low risk reported appropriate perceived susceptibility scores. However, those whose actual risk was average, perceived their risk to be low. There was no significant relationship found between actual risk of developing skin cancer and use of sunscreen. There was a significant relationship between age and perceived susceptibility to skin cancer. The older the subject, the higher the perceived susceptibility. Health care providers have the ability to influence individuals to use sunscreen. Accurately perceived susceptibility to skin cancer will increase the likelihood of health care providers recommending sunscreen to their patients. PMID- 10851767 TI - Knowledge, barriers, and motivators related to cervical cancer screening among Korean-American women. A focus group approach. AB - Cervical cancer is a significant health problem for Korean-American women. It currently is the number one female cancer diagnosed among women in South Korea. Despite this fact, Korean-American women have very low rates of cervical cancer screening. The purpose of this research were to gain an understanding of Korean women's knowledge about cervical cancer, and to identify major barriers to early screening for cervical cancer and the motivators for prevention and early detection. It is hoped that the findings will guide the development of community based cervical cancer education and screening programs for adult Korean-American women. The health belief model (HBM) provided the theoretical basis for the study. A qualitative study with eight focus groups (n = 102) was conducted using 11 questions derived from the HBM. Focus group discussions revealed that there was misinformation and a lack of knowledge about cervical cancer. The women therefore were confused about the causative factors and preventive strategies related to cervical cancer. The findings showed that major structural barriers were economic and time factors along with language problems. Many participants were recent immigrants with no medical insurance and long work hours. The main psychosocial barriers were fear/fatalism, denial, and Confucian thinking. Participants stated that medical advice and education would influence them most to undergo a Pap test. Recommendations were made to reduce certain barriers and to increase knowledge and motivations. PMID- 10851768 TI - Information needs regarding menopause. Results from a survey of women receiving cancer prevention and detection services. AB - Women often have questions related to menopause and hormone replacement therapy (HRT). A brochure entitled Understanding Menopause and Beyond was developed to address these issues. The purpose of this study was (a) to formally evaluate the relevance and utility of a brochure in understanding menopause and related health concerns and (b) to describe women's information needs at menopause. This descriptive study was conducted using a self-administered survey with a convenience sample of 200 pre-, peri-, and postmenopausal women attending a cancer screening center. On the basis of survey results, the topics most likely to be discussed with a health care provider were the risks and benefits of HRT and bone mineral density testing. The topics most frequently cited in the brochure that women previously did not know or understand were the questions to ask and the information to share with a health care provider and the risk factors for osteoporosis. The most important and informative sections of the brochure were those describing the risk factors for osteoporosis, the questions to ask and the information to share with a health care provider, and the risks and benefits of HRT. It was concluded that women have information needs regarding menopause not only related to the potential cancer risks, but also about related health issues such as osteoporosis, cardiovascular health, and emotional health. Furthermore, a targeted brochure can be effective in addressing these information needs. Oncology nurses who provide cancer screening and education services frequently receive questions about the safety and efficacy of HRT and other questions related to menopause. They therefore are uniquely qualified to address these concerns with their patients. PMID- 10851769 TI - Needs analysis of a cancer education program in south western Sydney. AB - Before establishing a cancer education program in South Western Sydney (SWS), a study was conducted to investigate the level of interest and presentation format preferred by patients with cancer and their relatives/friends regarding five types of cancer-related information: medical, psychological, and physical care issues as well as support services and available resources. A questionnaire using a Likert scale and open- and close-ended questions was distributed to the local cancer population and their families comprising a total sample size of 141. The results showed a uniformly high level of interest in learning about all five cancer-related topics with a preferred program format that consisted of a mixed media presentation of 1 to 6 weeks duration, consisting of 1- to 2-hour weekly sessions. A cancer education program based on the needs assessment findings was developed and implemented at Liverpool Hospital, which was positively evaluated by participants. PMID- 10851770 TI - Caregiver quality of life after autologous bone marrow transplantation. AB - Bone marrow transplantation (BMT) is a unique cancer therapy characterized by its novelty, intensity, and toxicity. Although families have been identified as having a critical influence on patient adaptation during the acute phase of BMT, minimal attention has been paid to their experiences during extended survivorship. This article reviews findings from a descriptive study on quality of life in primary caregivers of adult autologous bone marrow transplantation (AuBMT) survivors after acute hospitalization. Caregiver perceptions of their survival are delineated in an effort to characterize the dynamics of family recovery after BMT. Specifically, caregivers of AuBMT survivors require ongoing assistance to maintain their primary support role after BMT. PMID- 10851771 TI - Empowerment of Chinese patients with cancer through self-help groups in Hong Kong. AB - This study was conducted to identify the process and outcomes of empowerment as experienced by Hong Kong Chinese patients with cancer through participation in cancer self-help groups. The study involved in-depth individual interviews of self-help group members (n = 12) and participant observation of the group meetings over a period of 6 months. The empowered outcomes at a personal level included interconnectedness, confidence and hope, support and affirmation, and a feeling of usefulness. At a social level, expanded social network and opportunities to participate in more activities were reported. Collective efficacy also was demonstrated, although this happened only occasionally through participation in the group. One main theme that runs through the process of participation is empowerment among the members. On the basis of the efficacy demonstrated by self-help groups in this study, nurses should strongly consider referral of patients to such groups. Self-help groups serve as an important resource for patients with cancer in the Hong Kong Chinese community. PMID- 10851772 TI - Hope in newly diagnosed patients with cancer. AB - In this study, 131 Norwegian patients with recently diagnosed cancer completed the Nowotny Hope Scale (NHS). The NHS is composed of six subscales, and both global and subscale scores were assessed. Most of the patients were found to be hopeful or moderately hopeful. The variable with the single most positive contribution to hope was whether the patient lived alone. Younger people, in particular, experienced less hope when living alone. Gender, the time elapsed since diagnosis, and treatments had no observable effect on the global hope score. However, age, education level, and type of cancer was associated with particular domains of hope. PMID- 10851773 TI - The effects of hydrocolloid dressing and gentian violet on radiation-induced moist desquamation wound healing. AB - The aim of the study was to compare the effect of a gentian violet topical application with that of a moist dressing (hydrocolloid) on the rate and efficacy of radiotherapy-induced moist desquamation wound healing and the patients' satisfaction level with each method. This prospective randomized clinical trial used a stratified sampling design. A sample of 39 patients with 60 wounds had their wounds assessed on alternate days in terms of several wound-healing parameters including wound size, wound pain, incidence of infection, and time required for healing. Patient satisfaction with each treatment was evaluated at the completion of the study. Gentian violet significantly decreased wound size and reduced wound pain. However, this treatment received significantly lower ratings for dressing comfort and dressing aesthetic acceptance. Nevertheless, the time required for healing was not statistically different in the two groups. These findings suggest that the lower score of dressing satisfaction level in the gentian violet group may result from the skin discoloration and drying effects of the treatment, which renders patients unable to move or stretch their skin. Although the aim is to have complete wound healing, this may not be realistic for many lesions such as radiotherapy-induced moist desquamation wounds. The best evidence on which to make decisions about individual care can now be based on patients' own perception of quality. PMID- 10851774 TI - Intimacy and sexuality for the woman with breast cancer. AB - Human sexuality is more than sexual function. It is an ever-changing lived experience affecting the manner in which we view ourselves and our bodies. Most health professionals fail to address sexuality in the clinical setting and feel more comfortable focusing on treatment outcomes, such as the management of treatment side effects, than in addressing issues related to sexual behavior. Perhaps this is because many health professionals are uncomfortable about initiating a topic regarding a person's sexuality, or because they are unsure of their knowledge relating to changes in a person's sexuality after the management of cancer. Cultural issues in our society, such as the myth that older women with breast cancer are no longer interested in sexuality and intimacy, and the presumption that issues of survival overshadow sexuality, provide barriers to open communication about sexuality in women with breast cancer. Sexuality in the patient with breast cancer needs to be addressed by the nurse irrespective of the woman's age, partnership, and disease status. Knowledge related to changes in a woman's sexuality and intimacy after the management of breast cancer are explored, and strategies are provided for the nurse to use in communicating openly about sexuality in the clinical setting. PMID- 10851775 TI - Foot massage. A nursing intervention to modify the distressing symptoms of pain and nausea in patients hospitalized with cancer. AB - This article describes the findings of an empirical study on the use of foot massage as a nursing intervention in patients hospitalized with cancer. The study was developed from the earlier work of Ferrell-Torry and Glick (1992). In a sample of 87 subjects, a 10-minute foot massage (5 minutes per foot) was found to have a significant immediate effect on the perceptions of pain, nausea, and relaxation when measured with a visual analog scale. The use of foot massage as a complementary method is recommended as a relatively simple nursing intervention for patients experiencing nausea or pain related to the cancer experience. Further research into its effectiveness in the management of these symptoms by the family at home is warranted. PMID- 10851776 TI - The making of a leukocyte receptor: origin, genes and regulation of human CD38 and related molecules. PMID- 10851777 TI - Retinoid-mediated signaling in CD38 antigen expression. PMID- 10851778 TI - Enzymatic functions and structures of CD38 and homologs. PMID- 10851779 TI - Enzymic and signal transduction properties of CD38/NADase and PC 1/phosphodiesterase. PMID- 10851780 TI - Topology of CD38. PMID- 10851781 TI - CD38/CD31, a receptor/ligand system ruling adhesion and signaling in human leukocytes. PMID- 10851782 TI - Physiological and pathological significance of the CD38-cyclic ADP-ribose signaling system. PMID- 10851783 TI - CD38 in T- and B-cell functions. PMID- 10851784 TI - CD38 in hematopoiesis. PMID- 10851785 TI - CD38 in hematopoietic malignancies. PMID- 10851786 TI - CD38+CD8+ T cells as a marker of poor response to therapy in HIV-infected individuals. PMID- 10851787 TI - CD38 in health and disease. PMID- 10851788 TI - BST-1/CD157 regulates the humoral immune responses in vivo. PMID- 10851789 TI - Schematic portrait of human CD38 and related molecules. AB - Finally, a summary of the main characteristics of the CDB8 molecule are outlined in figure 7. PMID- 10851790 TI - Individual characteristics and peer relations of psychiatrically hospitalized aggressive youths: implications for treatment. AB - OBJECTIVE: This study identified individual and peer-relations problems of inpatient youths who are aggressive, and whether youths who are aggressive in two settings have greater treatment needs than youths who are aggressive in one setting only. METHODS: 85 youths aged 10 to 16 years who were consecutively admitted to a psychiatric facility served as participants. Based on ratings by parents and hospital staff, youths were identified as aggressive in the community only, aggressive in the hospital only, aggressive in both settings, or nonaggressive. Dependent measures consisted of youth self-reports and ratings by parents and hospital staff. RESULTS: Youths who demonstrate aggressive behavior in two settings have more nonaggressive behavior problems, more disturbed peer relations, and more hostile thinking than do nonaggressive youths, and some youths, although they may behave aggressively during hospitalization, have similar treatment needs as nonaggressive youths. CONCLUSIONS: Mental health professionals who work with youths in psychiatric settings need to develop treatment plans that directly address the more severe externalizing problems, hostile thinking, and peer problems of aggressive youths over and above that of nonaggressive youths, and should be aware that youths who behave aggressively during hospitalization may not have problems more severe than those of nonaggressive youths. PMID- 10851791 TI - Depression and school functioning in non-referred adolescents: a pilot study. AB - Self-image and self-perceived competencies have been considered to be related to depression in childhood and adolescence. Data from literature points to school functioning as one of the most important factors in self-esteem and self-worth during adolescence. Academic self image, defined as the way adolescents represent themselves as students, directly affects the global self-image; for this reason it has important psychopathological implications. The major aim of this preliminary report is to specifically analyze the relationship between academic self-image (assessed with a specific questionnaire), and self-reported depressive symptoms (assessed with the Children's Depression Inventory) in a school sample 150 adolescents. Our data indicate that the emotional beliefs about schooling and learning were significantly related to depressive symptomatology. Females scored higher in CDI and school anxiety. A real school failure did not affect the academic self image. These data seem to suggest that different components of the academic self-image can be differently associated with depressive feelings. PMID- 10851792 TI - Mothers' and children's perceptions of medication for children with attention deficit hyperactivity disorder. AB - This study assessed the perceptions about medication held by mothers and children with ADHD. Thirty-one mother-child dyads completed questionnaires assessing their knowledge and perceptions of stimulant medication and their level of perceived side effects associated with taking medication for ADHD. Mothers tended to view their children's medication as more beneficial than did their children. Mothers also demonstrated significantly more knowledge about their children's medication. Implications are drawn for research and clinical education of children and parents about medication for ADHD. PMID- 10851793 TI - Right frontal EEG asymmetry and lack of empathy in preschool children of depressed mothers. AB - EEG activity, empathic reactions to emotion-inducing stimuli, and the ability to complete a teaching task were examined in preschool children of depressed and non depressed mothers. EEG activity from frontal and parietal regions was recorded. Repeated measures MANOVAs indicated that the children of depressed mothers had greater relative right frontal EEG asymmetry, a pattern that typically accompanies greater negative affect and showed less empathic responses to a crying infant as well as to their own mothers' simulated distress. Children of depressed mothers were slower in completing the teaching task (involving mutual cooperation with their mother) and they spent more time asking for help than children of non-depressed mothers. Further, the depressed mothers stated their approval less often and spent less time helping their child complete the task. PMID- 10851794 TI - Depressive comorbidity in children and adolescents with generalized anxiety disorder. AB - Aim of this study is to examine the effect of depressive comorbidity in 108 children and adolescents with Generalized Anxiety Disorder (GAD). Fifty-five patients with GAD and depression were compared with 53 patients with GAD without depression. Age, gender and socioeconomic status did not differentiate the groups. Patients with comorbid depression had significantly more anxiety symptoms than patients without depression. Clinical presentation of GAD and pattern of comorbidity was similar in the two groups. Subjects with comorbid depression showed a more severe functional impairment, assessed with C-GAS. Data are discussed in the light of conceptualizations about the relationship between anxiety and depression. PMID- 10851795 TI - Pneumococcal disease in Australia. AB - The proceedings of the Pneumococcal Disease in Australia Workshop, held on 26-27 March 1999 are presented in this report. The world-wide epidemiology of the pneumococcus, with its predilection for the very young and the very old, differs between the developing and the developed world, and between indigenous and non indigenous populations. Sources of data on pneumococcal disease in each of the Australian States, clinical aspects of invasive and non-invasive disease, and the role of the public health laboratory in surveillance of serotypes and antimicrobial sensitivity, both nationally and over time, were discussed at the Workshop. Polysaccharide pneumococcal vaccines are recommended for those over 65 years of age and for at-risk groups, but are supplied free of charge only in Victoria and for indigenous Australians over 50 years of age. Children will require conjugate vaccines, which are likely to be licensed in the United States of America early in 2000. In Australia indigenous children, especially in rural areas, will be the priority group for conjugate vaccines. PMID- 10851796 TI - Surveillance of pneumococcal disease in Australian states and territories. AB - Information on pneumococcal disease, including immunisation programs, and optimum future surveillance in each Australian State and Territory were discussed at the Pneumococcal Disease in Australia Workshop on 26-27 March 1999. Workshop participants further expanded on the surveillance aspects of the Workshop in this report. Most participants favoured notification by laboratories of pneumococcal isolates from sterile sites, to provide baseline surveillance data before immunisation programs are fully implemented. It was also thought that trends in antimicrobial resistance should be notified. PMID- 10851797 TI - Communicable diseases surveillance. PMID- 10851798 TI - Fetal electrocardiogram extraction by blind source subspace separation. AB - In this paper, we propose the emerging technique of independent component analysis, also known as blind source separation, as an interesting tool for the extraction of the antepartum fetal electrocardiogram from multilead cutaneous potential recordings. The technique is illustrated by means of a real-life example. PMID- 10851799 TI - Blind signal separation from optical imaging recordings with extended spatial decorrelation. AB - Optical imaging is the video recording of two-dimensional patterns of changes in light reflectance from cortical tissue evoked by stimulation. We derived a method, extended spatial decorrelation (ESD), that uses second-order statistics in space for separating the intrinsic signals into the stimulus related components and the nonspecific variations. The performance of ESD on model data is compared to independent component analysis algorithms using statistics of fourth and higher order. Robustness against sensor noise is scored. When applied to optical images, ESD separates the stimulus specific signal well from biological noise and artifacts. PMID- 10851800 TI - Observer of autonomic cardiac outflow based on blind source separation of ECG parameters. AB - We present a novel method which provides an observer of the autonomic cardiac outflow using heartbeat intervals (RR) and QT intervals. The model of the observer is inferred from qualitative physiological knowledge. It consists in a problem of blind source separation of noisy mixtures which is resolved by a simple and robust algorithm. The robustness of the algorithm has been assessed by numerical simulations in adverse noisy environments. In clinical applications, we have validated the observer on subjects exposed to experimental conditions known to elicit sympathetic or parasympathetic response. PMID- 10851801 TI - Extraction of event-related signals from multichannel bioelectrical measurements. AB - Independent component analysis (ICA) is a powerful tool for separating signals from their mixtures. In this field, many algorithms were proposed, but they poorly use a priori information in order to find the desired signal. Here, we propose a fixed point algorithm which uses a priori information to find the signal of interest out of a number of sensors. We particularly applied the algorithm to cancel cardiac artifacts from a magnetoencephalogram. PMID- 10851802 TI - Independent component approach to the analysis of EEG and MEG recordings. AB - Multichannel recordings of the electromagnetic fields emerging from neural currents in the brain generate large amounts of data. Suitable feature extraction methods are, therefore, useful to facilitate the representation and interpretation of the data. Recently developed independent component analysis (ICA) has been shown to be an efficient tool for artifact identification and extraction from electroencephalographic (EEG) and magnetoencephalographic (MEG) recordings. In addition, ICA has been applied to the analysis of brain signals evoked by sensory stimuli. This paper reviews our recent results in this field. PMID- 10851803 TI - Independent component analysis of noninvasively recorded cortical magnetic DC fields in humans. AB - We apply a recently developed multivariate statistical data analysis technique- so called blind source separation (BSS) by independent component analysis--to process magnetoencephalogram recordings of near-dc fields. The extraction of near dc fields from MEG recordings has great relevance for medical applications since slowly varying dc-phenomena have been found, e.g., in cerebral anoxia and spreading depression in animals. Comparing several BSS approaches, it turns out that an algorithm based on temporal decorrelation successfully extracted a dc component which was induced in the auditory cortex by presentation of music. The task is challenging because of the limited amount of available data and the corruption by outliers, which makes it an interesting real-world testbed for studying the robustness of ICA methods. PMID- 10851804 TI - Magnetic coil design considerations for functional magnetic stimulation. AB - Our studies have demonstrated effective stimulation of the bladder, bowel, and expiratory muscles in patients with spinal cord injury using functional magnetic stimulation. However, one limitation of the magnetic coils (MC) is related to their inability to specifically stimulate the target tissue without activation of surrounding tissue. The primary goal of this study was to determine the governing parameters in the MC design, such as coil configuration, diameter, and number of turns in one loop of the coil. By varying these parameters, our approach was to design, construct, and evaluate the induced electric field distributions of two sets of novel MC's. Based on the slinky coil design, the first set of coils was constructed to compare their abilities in generating induced electric fields for focal nerve excitation. The second set of coils was built to determine the effect that changes in two parameters, coil diameter and number of turns in one loop, had on field penetration. The results showed that the slinky coil design produced more focalized stimulation when compared to the planar round coils. The primary to-secondary peak ratios of the induced electric field from slinky 1 to 5 were 1.00, 2.20, 2.85, 2.62, and 3.54. We also determined that coils with larger diameters had better penetration than those with smaller diameters. Coils with less number of turns in one loop had higher initial field strengths; when compared to coils that had more turns per loop, initial field strengths remained higher as distance from the coil increased. In our attempt to customize MC design according to each functional magnetic stimulation application and patients of different sizes, the parameters of MC explored in this study may facilitate designing an optimal MC for a certain clinical application. PMID- 10851805 TI - Insulin release at the molecular level: metabolic-electrophysiological modeling of the pancreatic beta-cells. AB - The role of pancreatic beta-cells is fundamental in the control endocrine system, maintaining the blood glucose homeostais in a physiological regime, via the glucose-induced release of insulin. An increasing amount of detailed experimental evidences at the cellular and molecular biology levels have been collected on the key factors determining the insulin release by the pancreatic beta-cells. The direct transposition of such experimental data into accurate mathematical descriptions might contribute to considerably clarify the impact of each cellular component on the global glucose metabolism. Under these perspectives, we model and computer-simulate the stimulus-secretion coupling in beta-cells by describing four interacting cellular subsystems, consisting in the glucose transport and metabolism, the excitable electrophysiological behavior, the dynamics of the intracellular calcium ions, and the exocytosis of granules containing insulin. We explicit the molecular nature of each subsystem, expressing the mutual relationships and the feedbacks that determine the metabolic-electrophysiological behavior of an isolated beta-cell. Finally, we discuss the simulation results of the behavior of isolated beta-cells as well as of population of electrically coupled beta-cells in Langerhans islets, under physiological and pathological conditions, including noninsulin-dependent diabetes mellitus (NIDDM) and hyperinsulinemic hypoglycaemia (PHHI). PMID- 10851806 TI - Monte Carlo modeling for implantable fluorescent analyte sensors. AB - A Monte Carlo simulation of photon propagation through human skin and interaction with a subcutaneous fluorescent sensing layer is presented. The algorithm will facilitate design of an optical probe for an implantable fluorescent sensor, which holds potential for monitoring many parameters of biomedical interest. Results are analyzed with respect to output light intensity as a function of radial distance from source, angle of exit for escaping photons, and sensor fluorescence (SF) relative to tissue autofluorescence (AF). A sensitivity study was performed to elucidate the effects on the output due to changes in optical properties, thickness of tissue layers, thickness of the sensor layer, and both tissue and sensor quantum yields. The optical properties as well as the thickness of the stratum corneum, epidermis, (tissue layers through which photons must pass to reach the sensor) and the papillary dermis (tissue distal to sensor) are highly influential. The spatial emission profile of the SF is broad compared that of the tissue fluorescence and the ratio of sensor to tissue fluorescence increases with distance from the source. The angular distribution of escaping photons is more concentrated around the normal for SF than for tissue AF. The information gained from these simulations will be helpful in designing appropriate optics for collection of the signal of interest. PMID- 10851807 TI - Microwave thermochemotherapy in the treatment of the bladder carcinoma- electromagnetic and dielectric studies--clinical protocol. AB - Microwave thermotherapy is currently used in clinical routines for benign prostatic hyperplasia treatments. The temperature increase is obtained using an endocavitary microwave applicator placed in the prostatic urethra. This urethral applicator after a technical modification can be placed inside the bladder in order to potentiate the effects of the treatment by chemotherapy of vesical carcinoma. This paper deals with electromagnetic studies of this new endocavitary applicator. First of all, the experimental determination of the dielectric permittivities for the propagation domain characterization is achieved in order to be used in the electromagnetic model. Compared to experimental results, these simulations obtained by the finite-difference time-domain formalism allow us to determine the electromagnetic performance of this applicator. Finally, the in vivo study realized on anesthetized dogs to determine the therapeutic protocol associating chemotherapy and thermotherapy in the treatment of the bladder cancer is presented. PMID- 10851808 TI - Estimating neural sources from each time-frequency component of magnetoencephalographic data. AB - We have developed a method that incorporates the time-frequency characteristics of neural sources into magnetoencephalographic (MEG) source estimation. This method, referred to as the time-frequency multiple-signal-classification algorithm, allows the locations of neural sources to be estimated from any time frequency region of interest. In this paper, we formulate the method based on the most general form of the quadratic time-frequency representations. We then apply it to two kinds of nonstationary MEG data: gamma-band (frequency range between 30 100 Hz) auditory activity data and spontaneous MEG data. Our method successfully detected the gamma-band source slightly medial to the N1m source location. The method was able to selectively localize sources for alpha-rhythm bursts at different locations. It also detected the mu-rhythm source from the alpha-rhythm dominant MEG data that was measured with the subject's eyes closed. The results of these applications validate the effectiveness of the time-frequency MUSIC algorithm for selectively localizing sources having different time-frequency signatures. PMID- 10851809 TI - A transform domain SVD filter for suppression of muscle noise artefacts in exercise ECG's. AB - The proposed filter assumes the noisy electrocardiography (ECG) to be modeled as a signal of deterministic nature, corrupted by additive muscle noise artefact. The muscle noise component is treated to be stationary with known second-order characteristics. Since noise-free ECG is shown to possess a narrow-band structure in discrete cosine transform (DCT) domain and the second-order statistical properties of the additive noise component is preserved due to the orthogonality property of DCT, noise abatement is easily accomplished via subspace decomposition in the transform domain. The subspace decomposition is performed using singular value decomposition (SVD). The order of the transform domain SVD filter required to achieve the desired degree of noise abatement is compared to that of a suboptimal Wiener filter using DCT. Since the Wiener filter assumes both the signal and noise structures to be statistical, with a priori known second-order characteristics, it yields a biased estimate of the ECG beat as compared to the SVD filter for a given value of mean-square error (mse). The filter order required for performing the subspace smoothing is shown to exceed a certain minimal value for which the mse profile of the SVD filter follows the minimum-mean-quare error (mmse) performance warranted by the suboptimal Wiener filter. The effective filter order required for reproducing clinically significant features in the noisy ECG is then set by an upper bound derived by means of a finite precision linear perturbation model. A significant advantage resulting from the application of the proposed SVD filter lies in its ability to perform noise suppression independently on a single lead ECG record with only a limited number of data samples. PMID- 10851810 TI - Optimal digital filtering for tremor suppression. AB - Remote manually operated tasks such as those found in teleoperation, virtual reality, or joystick-based computer access, require the generation of an intermediate electrical signal which is transmitted to the controlled subsystem (robot arm, virtual environment, or a cursor in a computer screen). When human movements are distorted, for instance, by tremor, performance can be improved by digitally filtering the intermediate signal before it reaches the controlled device. This paper introduces a novel tremor filtering framework in which digital equalizers are optimally designed through pursuit tracking task experiments. Due to inherent properties of the man-machine system, the design of tremor suppression equalizers presents two serious problems: 1) performance criteria leading to optimizations that minimize mean-squared error are not efficient for tremor elimination and 2) movement signals show ill-conditioned autocorrelation matrices, which often result in useless or unstable solutions. To address these problems, a new performance indicator in the context of tremor is introduced, and the optimal equalizer according to this new criterion is developed. Ill conditioning of the autocorrelation matrix is overcome using a novel method which we call pulled-optimization. Experiments performed with artificially induced vibrations and a subject with Parkinson's disease show significant improvement in performance. Additional results, along with MATLAB source code of the algorithms, and a customizable demo for PC joysticks, are available on the Internet at http:?tremor-suppression.com. PMID- 10851811 TI - Study of myographic signals from sternomastoid muscle in patients with chronic obstructive pulmonary disease. AB - Analysis of the respiratory muscle activity is a promising technique for diagnosis of respiratory diseases, such as chronic obstructive pulmonary disease (COPD). The sternomastoid muscle (SMM) was selected to study the activity of respiratory muscles due to its accessibility in order to define a noninvasive analysis. The aims of this work are two: analyze the relationship between the SMM function and pulmonary obstruction, and study the influence of spectral estimator on frequency parameters related with the muscle activity. For the first goal, we propose the analysis of vibromyographic and electromyographic signals from the SMM to study the muscle function during two ventilatory tests. Activity of SMM was found by means of several indexes: root-mean-square (rms) values, mean and median frequencies, and ratio between high and low-frequency components. For the second goal, spectral analysis was performed by means of nonparametric methods: Correlogram and Welch periodogram, and parametric methods: autoregressive (AR), moving average (MA), and ARMA models. It is deduced that these indexes show muscle activity and certain fatigue of the SMM, whose muscle function depends on the level of pulmonary obstruction, and they depend a lot of spectral estimator being the more suitable an AR model with high order. PMID- 10851812 TI - Continuous localization of cardiac activation sites using a database of multichannel ECG recordings. AB - Monomorphic ventricular tachycardia and ventricular extrasystoles have a specific exit site that can be localized using the multichannel surface electrocardiogram (ECG) and a database of paced ECG recordings. An algorithm is presented that improves on previous methods by providing a continuous estimate of the coordinates of the exit site instead of selecting one out of 25 predetermined segments. The accuracy improvement is greatest, and most useful, when adjacent pacing sites in individual patients are localized relative to each other. Important advantages of the new method are the objectivity and reproducibility of the localization results. PMID- 10851813 TI - Image restoration in chirp-pulse microwave CT (CP-MCT). AB - Chirp-pulse microwave computed tomography (CP-MCT) is a technique for imaging the distribution of temperature variations inside biological tissues. Even if resolution and contrast are adequate to this purpose, a further improvement of image quality is desirable. In this paper, we discuss the blur of CP-MCT images and we propose a method for estimating the corresponding point spread function (PSF). To this purpose we use both a measured and a computed projection of a cylindrical phantom. We find a good agreement between the two cases. Finally the estimated PSF is used for deconvolving data corresponding to various kinds of cylindrical phantoms. We use an iterative nonlinear deconvolution method which assures nonnegative solutions and we demonstrate the improvement of image quality which can be obtained in such a way. PMID- 10851814 TI - Cardiopulmonary resuscitation in India: ethical issues. PMID- 10851815 TI - Long-term stability of bronchial reactivity in guinea pigs. AB - Measurement of lung function and bronchial reactivity are widely used as outcome parameters to assess the efficacy of therapeutic interventions. In order to interpret the results correctly, it is necessary that the outcome parameters are themselves stable over time so that any significant changes measured may be attributed to the interventions. Specific airway conductance (SGaw) and airway reactivity to histamine are two commonly used parameters in animal models such as guinea pigs. Although short-term variability of these parameters has been investigated, there has been no study of long-term stability. In the present paper, SGaw and bronchial reactivity to histamine were measured in 111 conscious guinea pigs using a non-invasive, whole body plethysmograph. Baseline values of SGaw and ED35 histamine were measured and followed for eight weeks at weekly intervals. At baseline, mean SGaw in guinea pigs was 0.17 +/- 0.055 sec-1 cm H2O 1 and ED35 histamine ranged from 0.064 to more than 10 mg/ml. The distribution of ED35 histamine values was gaussian. We observed that the changes in SGaw and ED35 histamine recorded using this technique are highly reproducible over eight weeks. The reactivity varied by less than a doubling dose of histamine over any two consecutive weeks. Thus, the technique described in this paper is quick, easily learned, reproducible, independent of temperature-humidity artifact and highly suitable for studies of repeated measurements as in the study of dietary interventions and evaluation of effect of drugs. PMID- 10851816 TI - Traumatic diaphragmatic hernia: an Asir region (Saudi Arabia) experience. AB - Ten patients (nine males, one female), seen at the Asir Central Hospital of South Western Saudi Arabia with proven traumatic diaphragmatic hernia between 1987 and 1997, were reviewed retrospectively. The mean age was 29.6 years, range 5 to 50 years. Chest pain and vomiting were the commonest symptoms. Blunt trauma (road traffic accident--5, fall from height--1, (accounted for 60% of the cases) while gunshot wound and stab wounds were the causes in two patients each. The chest radiograph suggested the diagnosis in all the cases. Barium meal (in two patients) and barium enema (in two patients) complemented the diagnosis. Computed tomography (CT) scan was done in only one patient. Thoracotomy (in 2 patients), laparotomy (in 5 patients) and thoraco-laparotomy (in 3 patients) were the surgical approaches to management. Common herniated organs were liver, stomach, spleen and large bowel. The injuries were on the left side in seven patients and on the right side in three cases. Immediate surgical repair was done in four patients while it was done two days to four years later in others. Complications were minimal and there was only one death. PMID- 10851817 TI - Diurnal variation in peak expiratory flow in healthy young adults. AB - Diurnal variability in peak expiratory flow (PEF) was studied for three days in 152 healthy volunteers aged 20-40 years, and expressed as amplitude percent mean (A%M) and standard deviation percent mean (SD%M). The commonest pattern (74.6%) observed was that of a nadir at waking in morning, a progressive rise upto late afternoon and a plateau or a small decrease at bedtime. Mean diurnal variation in PEF on the third day was 7.23 +/- 3.48 (A%M) and 2.98 +/- 1.31 (SD%M). Both the diurnal variation and the calculated upper limits of normality were found to be lower than those reported previously for Western subjects. This knowledge of diurnal variation is important for any meaningful interpretation of PEF recordings used to monitor patients with asthma. PMID- 10851818 TI - Revised National Tuberculosis Control Programme: Indian perspective. AB - The global tuberculosis programme has promoted the revision of National Tuberculosis Programme by strengthening the focus on Directly Observed Treatment Short-course (DOTS). National Tuberculosis Control Programme (NTCP) which was established in 1962 had less than 30 per cent treatment completion. Based on an in-depth review of the programme by a high level committee in 1992, a Revised National Tuberculosis Control Programme (RNTCP) was envisaged with a view to achieve a cure rate of at least 85 per cent amongst newly detected sputum positive cases under DOTS. By December 1999, 130 million of population had been covered in the country under DOTS. However, there are many challenges that are required to be met before RNTCP can become a success story in our country. PMID- 10851819 TI - Chronic anaerobic pneumonitis presenting as a pseudohilar opacity. PMID- 10851820 TI - Pleural involvement by Salmonella senftenberg: a report of two cases. AB - Non-typhoidal serovars of salmonella are an unusual cause of pleuropulmonary infections. We report two patients with empyema caused by Salmonella senftenberg. One patient had associated diabetes and gall bladder carcinoma, and infection was acquired in hospital. Both patients responded well to parenteral antibiotics. PMID- 10851821 TI - Recurrent respiratory papillomatosis. AB - Recurrent respiratory papillomatosis is characterized by the appearance of benign laryngeal squamous papillomas in childhood. Lung involvement is rare. We report a case of childhood laryngeal papillomas who developed tracheobronchial papillomas and a nodule in the lung after a period of 21 years. Frequent sampling of pulmonary lesions to detect malignant transformation is suggested as prognosis of lung lesions are worse in comparison to laryngeal papillomas. PMID- 10851822 TI - ECG of the month. Appearances are deceiving. Trigeminal rhythm. PMID- 10851823 TI - The otologic manifestations of barotrauma. AB - Barotrauma is defined as an injury due to pressure differences between atmospheric and intratympanic pressures. Human beings are well suited to operate within an environment involving small alterations in atmospheric pressure. Man's persistence in operating outside this environment leads to exposure to large pressure differentials with resulting trauma. External, middle, and inner ear structures can all be injured due to alterations in pressure. The increase in popularity of sport diving and aviation travel has led to an increase in the number of otologic injuries caused by barotrauma. The physics, pathophysiology, symptoms, and treatment of barotrauma are presented. PMID- 10851824 TI - Radiology case of the month. Abdominal mass. Bilateral ovarian cystic teratoma. PMID- 10851825 TI - The journal 150 & 100 years ago. March 1850 and 1900. PMID- 10851826 TI - Partial colectomy required for resection of renal cell carcinoma: a case report and review of treatment options for locally advanced disease. AB - Because it is more commonly discovered as a result of an incidental finding on radiologic studies, renal cell carcinoma is being diagnosed at earlier stages. Patients still, however, present occasionally with locally advanced disease. Such a case is presented in a patient who required a partial colectomy at the time of radical nephrectomy to remove all of his disease. Also reviewed is the current state of treatment options available for renal cell carcinoma, including chemotherapy, radiation therapy, immunotherapy, and surgery. Despite advances in some of these areas, the mainstay of treatment for locally advanced renal cell carcinoma remains surgery. PMID- 10851827 TI - Diagnosis and management of a painful thyroid nodule in a patient with systemic sarcoidosis. AB - Painful thyroid nodules caused by sarcoid are exceedingly rare. Painless involvement of the thyroid by sarcoid in patients with systemic sarcoidosis is not. Several autoimmune thyroid illnesses are closely linked to sarcoid. These illnesses may form thyroid nodules which may or may not be painful. We present only the second reported case of a painful thyroid nodule caused by direct sarcoid involvement. Fine needle aspiration may not provide a definitive diagnosis in patients whose appropriate therapy would vary greatly depending on this diagnosis. When an open surgical procedure is indicated, total unilateral thyroid lobectomy should be considered. Multi-centric involvement of a lobe with postoperative recurrence in remaining ipsi-lateral thyroid tissue would be very likely if the entire lobe is not removed. PMID- 10851828 TI - Testicular tumors. Multi-disciplinary approach and follow up. PMID- 10851829 TI - Morphological pattern of testicular tumors. AB - OBJECTIVE: To find out the mode of presentation, age distribution and the prevalence of various histological subtypes of testicular tumors. METHOD: All consecutive cases of testicular tumors diagnosed in the department of pathology, the Aga Khan University Hospital, Karachi, during the period of eight years (1991 98) were included in this study. Relevant clinical details such as age, clinical presentation and side of involvement of the testis were also recorded, where available. RESULTS: During the span of eight years (1991-98), 170 cases of testicular tumors were diagnosed at the Aga Khan University Hospital, Karachi. Most of the tumors were diagnosed in the third and fourth decade of life. Scrotal mass or swelling was the predominant mode of presentation. There was a slight predominance of right-sided testicular tumors. Germ cell tumors constituted 83.5% of all malignant testicular neoplasms. Amongst these seminoma was the most common (36.5%) tumor followed by mixed germ cell tumors (28.82%). Yolk-sac tumor was the commonest testicular neoplasm in children while lymphoma was the predominant neoplasm in the elderly population. CONCLUSION: The overall relative frequency of testicular malignancy in this series correlated with that reported in the international literature. PMID- 10851830 TI - Role of cytokines in giardiasis. AB - OBJECTIVES: The purpose of this study was to determine the relationship of inteleukins with G. lamblia infection. METHODS: Serum interleukins were estimated in 42 patients suffering from giardiasis and 42 apparently healthy controls. Tumor Necrosis Factor (TNF) was also studied in 14 patients and 14 controls. Interleukins (IL-2, IL-4, IL-10) and TNF alpha were determined by ELISA. RESULTS: IL-4 was present in 14 (33.3%) patients (mean value 220 pg/ml) and 8 (19%) controls (mean value 93 pg/ml). IL-10 was present in 5 (12%) patients (mean value 57 pg/ml) and 6 (14%) controls (mean value 79 pg/ml). IL-2 was present in one patient only but absent in controls. TNF alpha was not detected in patients but was present in 2 (14.2%) controls (mean value 75 pg/ml). CONCLUSION: Results indicate that IL-4 being an inflammatory regulator appears to have some relationship with giardiasis, while TNF alpha was not detected in patients probably because G. lamblia is a non invasive parasite. PMID- 10851831 TI - Multivariate analysis of risk factors associated with genital ulcer disease among incarcerated males in Sindh. AB - OBJECTIVE: To evaluate the potential risk behaviors associated with the lifetime risk of self reported genital ulcer disease (GUD) among prison inmates. SETTING: Prison inmates from 14 prisons of Sindh Province. METHODS: A cross-sectional study was conducted on 3395 prison inmates during July to December, 1994. A questionnaire was used to assess the lifetime risk of self-reported GUD (whether or not the subject was ever affected with GUD up to present age) and to investigate demographic markers and risk behaviors for their possible association with lifetime risk of GUD using logistic regression analysis. RESULTS: The reported lifetime risk of GUD in the study sample was 11.4% (386/3395). In final multivariate logistic regression model the sexual behaviors which were independently associated with GUD were having sexual intercourse with female (adjusted OR = 1.7; 95% CI: 1.3-2.3, P = 0.0002), sexual intercourse with a prostitute (adjusted OR = 1.5; 95% CI: 1.2-2.0, P = 0.0008), sexual intercourse with man (adjusted OR = 2.2; 95% CI: 1.7-2.7, P = < 0.001) and sexual intercourse with man during current incarceration (adjusted OR = 1.9; 95% CI: 1.2-2.9, P = 0.0071). CONCLUSION: Health education needs to re-enforce monogamous relationship for high risk groups such as in our study. Although infrequent condom use was not a risk factor for GUD in this study, yet based on the results of previous studies, promotion of condom use should be the component of health education program. PMID- 10851832 TI - Distribution of birthweights of hospital born Pakistani infants. AB - OBJECTIVE: To determine the distribution of birthweights among newborns and its relationship to specific sociodemographic and medical factors. METHODS: All babies born after 24 weeks of gestation between November, 1994, and June, 1996 were included irrespective of the fact whether they were live or still borns. Infants having birth weights 2.5 to 4 kg were termed as normally weighed babies, under 2.5 kg as low birth weight and above 4 kg as macrosomics. Data gathered included sociodemographic and medical variables. Birthweights of the newborns were measured without clothes to the nearest 10 gm on an infants beam balance within 15-30 minutes of birth. The baby scale was calibrated daily for accuracy. RESULTS: Mean birth weight of the newborns was 2.91 kg. Weight of 78% babies ranged from 2.5 to 4 kg, 19% had low birth weight and 3% of neonates weighed above 4 kg. Of 1156 low birth weight babies 70% were preterm, 16% were growth retarded and 14% were both premature and growth retarded. Macrosomic babies were commonly born to the mothers who were either 35 years age or more or were para > 5, whereas 59% cases of low birth weight was associated with primipartiy and grandmultiparity. Causes of low birth weight included APH, twin pregnancy, PROM and severe preeclampsia and eclampsia. Risk factors for fetal macrosomia were advanced maternal age and parity, postdatism and diabetes mellitus. CONCLUSION: In this study the relative impact of some of the factors related to birthweight in reference to our population was highlighted. Further explorations preferably in population based studies are required as birthweight data is essential for monitoring and evaluating the progress towards achieving national goals for lowering neonatal and infant morbidities and mortalities. PMID- 10851833 TI - What is the effect of riskshaw noise on its driver? AB - OBJECTIVE: Occupational hearing loss is common in the industrialized world. Road noise is a major contributor to perceived environmental noise. The objective of this study was to assess hearing loss in rickshaw drivers due to rickshaw noise. METHODS: Hearing loss in rickshaw drivers and taxi drivers of Karachi who were 50 years of age or younger was estimated, with a Smith Hearing Screening (SHS) questionnaire that was modified, translated into the national language, Urdu and field tested prior to administration. RESULTS: Interviews for 91 rickshaw drivers and 94 taxi drivers were completed. All subjects were male; mean ages were 34 and 33 years for rickshaw and taxi drivers respectively. None of the rickshaws were fitted with silencers. Rickshaw drivers were about thrice as likely to be screened as hearing impaired by the SHS questionnaire (RR 2.9, 95% confidence interval 1.6, 5.0), twice as likely to report tinnitus (RR 2.2, 95% confidence interval, 1.1, 3.3) and two and a half times as likely to have difficulty in following telephonic conversations (RR 2.4, 95% confidence interval 1.2, 4.8). CONCLUSION: There is loss of hearing and tinnitus among rickshaw drivers that could be attributed to their trade. Use of silencers by rickshaw drivers could result in less hearing loss among rickshaw drivers and less noise in the environment for the other 11 million residents in the city. PMID- 10851834 TI - Reversibility of acute demyelinating lesions in relapsing-remitting multiple sclerosis. PMID- 10851835 TI - Sexual harassment in medical profession--perspective from Pakistan. AB - OBJECTIVE: To assess the extent of Sexual harassment of female nurses by male physicians, patients or patient's relatives. SETTING: A general hospital in Islamabad. METHOD: A cross sectional written study through a self administered brief questionnaire. RESULTS: Male physicians were identified as the major perpetrators of sexual harassment, followed by the patients and their relatives. CONCLUSION: The nurses and hospital administration need to work together for fostering work environment conducive to healthy environment for effective health care delivery. PMID- 10851836 TI - Uterine atony at a tertiary care hospital in Pakistan: a risk factor analysis. AB - OBJECTIVES: To identify risk factors for uterine atony following assisted or unassisted vaginal delivery. DESIGN: This hospital based case control study was done at The Aga Khan University Karachi, Pakistan. Cases were defined as all women with uterine atony within 24 hours of an assisted or unassisted vaginal delivery. Controls were based on women with normal assisted or unassisted vaginal delivery without uterine atony. Data abstracted form the medical records; adjusted odds ratios were estimated by multiple logistic regression. RESULTS: Factors having a significant association with uterine atony were gestational diabetes mellitus (odds ratio 7.6, 95% CI 6.9-9.0, p = 0.003) and a prolonged second stage of labour in multiparas (odds ratio 4.0, 95% CI 3.1-5.0, p = 0.002). No associations were found with high parity, age, preeclampsia, augmentation of labour, antenatal anemia and a history of poor maternal or perinatal outcomes. CONCLUSIONS: Among previously documented risk factors for uterine atony, only a prolonged second stage of labour in multiparas was found to be significant in this study. Gestational diabetes mellitus, a previously undocumented factor, has also been identified as an independent risk factor. Multiparity and age were not found to be significant risk factors. The study underlines the importance of confirming these findings for better prevention and management of uterine atony. PMID- 10851837 TI - Vitamin A and infant mortality: beyond intention-to-treat in a randomized trial. AB - This paper investigates the effect of one dose of vitamin A on subsequent 4 month mortality in children under 6 months of age in a randomized, double-blind placebo controlled community trial in Nepal. An earlier published intention-to-treat analysis showed no benefit, but ignored the information on actual receipt of treatment. Structural failure time models (Robins and Tsiatis, '91) use randomization based inference and incorporate compliance information which is possibly selective. The data presented here offer some new challenges for this approach: ward-based randomization induces correlation between survival outcomes; and the actual receipt of vitamin A dose is not always recorded. To tackle the problem of the clustered survival data we consider a robust version of the structural parameter vector estimator. A sensitivity analysis captures boundaries for the estimated structural parameters reflecting a range of potential values of children whose true receipt of treatment is unknown. The analysis suggests that the effect of vitamin A was beneficial in the beginning of the trial but towards the end of the trial there was a reversal of this effect. PMID- 10851838 TI - Assessing the impact of managed-care on the distribution of length-of-stay using Bayesian hierarchical models. AB - Hierarchical models provide a useful framework for the complexities encountered in policy-relevant research in which the impact of social programs is being assessed. Such complexities include multi-site data, censored data and over dispersion. In this paper, Bayesian inference through Markov Chain Monte Carlo methods is used for the analysis of a complex hierarchical log-normal model that shows the impact of a managed care strategy aimed at limiting length of hospital stays. Parameters in this model allow for variability in baseline length-of-stay as well as the program effect across hospitals. The authors demonstrate elicitation and sensitivity analysis with respect to prior distributions. All calculations for the posterior and predictive distributions were obtained using the software BUGS. PMID- 10851839 TI - A two-stage estimator of the dependence parameter for the Clayton-Oakes model. AB - This paper describes the properties of a two-stage estimator of the dependence parameter in the Clayton-Oakes multivariate failure time model. The parameter is estimated from a likelihood function in which the marginal hazard functions are replaced by estimates. The method extends the approach of Shih and Louis (1995) and Genest, Ghoudi and Rivest (1995) to allow the marginal hazard for failure times to follow a stratified Cox (1972) model. The method is computationally simple and under mild regularity conditions produces a consistent, asymptotically normal estimator. PMID- 10851840 TI - Alternative time scales and failure time models. AB - In many reliability applications, there may not be a unique plausible scale in which to measure time to failure or assess performance. This is especially the case when several measures of usage are available on each unit. For example, the age, the total number of flight hours, and the number of landings are usage measures that are often considered important in aircraft reliability. Similarly, in medical or biological applications of survival analysis there are often alternative scales (e.g., Oakes, 1995). This paper considers the definition of a "good" time scale, along with methods of determining a time scale. PMID- 10851841 TI - [Detection and evaluation of adverse drug effects]. AB - Adverse drug reactions (ADRs) occur in about 5% of drug-treated patients. Hospital admissions are caused by ADRs in 5% of patients and roughly 2% of hospitalized patients will experience an ADR. The economic burden of ADRs can only be estimated. Type A reactions can be explained by the pharmacological action of the drugs, and are preventable in many cases. However, Type B reactions involving the immune system and/or idiosyncratic reactions occur rarely and most of them are not fully understood. Genotyping represents an elegant method to explain the presence of abnormal enzyme activities and allows prediction of adverse drug effects in individual cases. Typical time frames have been identified for the occurrence of hypersensitivity reactions, although definite causality assessment is often impeded due to the absence or unavailability of specific laboratory tests and the impossibility of rechallenge. Diagnosis of an ADR is often difficult due to comorbidity and polypharmacy, thus causality assessment is often divergent even between specialists. In Germany, ADRs are reported preferably to the manufacturer of the suspicious drug and then collected and evaluated at the Bundesinstitut fur Arzneimittel und Medizinprodukte, BfArM. However, total number and quality of reported ADRs could be improved. PMID- 10851842 TI - [Dose-response relationship: relevance for medical practice]. AB - Dose-finding studies are performed routinely in patients and--if appropriate surrogate models exist--also in healthy volunteers. Such studies aim at establishing the optimal dose range for further clinical studies on the efficacy and the risk-benefit ratio of a new drug. The dose-response relationship of a drug is most often described by a sigmoidal curve. Its parameters include the mean effective dose, the maximal effect and the steepness. Interpretation of such curves should be done in the context of the intended clinical indications of the drug. The risk-benefit ratio of a drug can be assessed by overlapping the dose response curve of wanted and unwanted clinical effects, again, any overlapping (which can be described e.g. by the therapeutic index) should be seen in the context of the indication and available therapeutic alternatives. PMID- 10851843 TI - [Responsibilities of ethics committees]. AB - Increasing numbers of clinical research projects are submitted to ethical committees (institutional review boards) for approval. New therapeutic developments have to be evaluated by these committees to protect patients/volunteers. Thus, the responsibility of ethical committees is increasing. The "Nurnberger Kodex" and the "Declaration of Helsinki" are the background for these evaluations. According to the German drug law the physician is obligated by law to submit the protocol to such a committee. In addition, local state physician authorities require such a procedure. Important considerations during the review process besides ethical aspects are the informed consent, which should be written in an understandable form, and the obligations of the insurance. PMID- 10851844 TI - [Clinically significant" new drug interactions]. AB - The concomitant intake of drugs with the potential to cause drug interactions is frequent. In contrast, adverse effects due to drug interactions account for only a small fraction of all adverse effects. A reproducible evaluation of the clinical relevance of drug interactions is lacking. We now can accurately define the potential of a drug to cause interactions, primarily by comparative investigations within a drug class. Whether or not the selection of the drug based on this information is useful for the patient is unknown. Therefore, usually it is to be recommended to abandon therapeutically reasonable drug combinations with a risk for interactions only if equivalent therapeutic options are available. Several actual examples on interactions with selective serotonin re-uptake inhibitors, HMG-CoA reductase inhibitors, mibefradil, sildenafil, protease inhibitors and with grapefruit juice are discussed. PMID- 10851845 TI - [Placebo effects and adverse effects in clinical trials]. AB - BACKGROUND: It is well known that placebos evoke a variety of effects in the human body. It is less known that placebo medication can produce adverse drug reactions (ADRs). METHOD: The efficacy and, as the main topic, the tolerability were investigated from an international clinical data pool on placebo-controlled randomized multicenter studies of 5 different groups of indications. The therapeutic areas analyzed were neuropsychiatry (nimodipine, ipsapirone), cardiology (nisoldipine), metabolism (acarbose) and gastroenterology (hydrotalcit). RESULTS: The placebo efficacy was evident and varied not only in comparison to 5 indication groups analyzed, but also within them. Placebo showed for example hardly an effect on the symptomatology of stroke patients with a severe neurological deficit in comparison to verum while there was a 50% improvement in symptoms following placebo application in patients with a mild stroke. In angina patients exercise tolerance increased under placebo by 10% (time to onset of ST segment depression and symptoms of angina) while verum showed a 22% improvement. In diabetes placebo had no effect as compared to baseline and to a verum on blood glucose and HbA1C. ADRs could be observed under placebo treatment. The frequency and kind of placebo-induced ADRs varied also between the indication groups, i.e. typical CV side-effects were observed in CV controls. The placebo ADR profile was similar to the reference drug also. Some ADRs were reported even more frequently under placebo than under verum, like "dry mouth" in patients with general anxiety status. CONCLUSIONS: Placebo therapy is frequently effective and is not a "non-therapy". Placebo effects can be shown only by direct comparison with a "non-therapy". A placebo therapy is frequently accompanied by adverse drug reactions (ADRs) just as a verum therapy. Placebo ADRs are frequently illness- and verum-specific. Effects and ADRs of placebo therapy must be known, to judge the effect of verum medication in controlled clinical trials. The mechanisms of placebo effects are manifold (e.g. endorphine release, conditioned learning). Since placebo therapy outside of evidence-based medicine (use of drugs without proven efficacy = pseudoplacebos) may potentially also cause serious side effects, the use may not only be useless but also harmful. PMID- 10851846 TI - [Responsibilities of clinical pharmacology in the early phase of drug development]. AB - The path of a new drug from the idea to the product may be divided into 2 phases, namely drug discovery and drug development. Due to the scientific progress new and simple methods could be developed to determine the biological efficacy of a large number of compounds. During the first part of drug development necessary requirements for the first use in man are met by performing preclinical pharmacological, toxicological and pharmacokinetic investigations in the animal and in in-vitro testing. After a first clinical-pharmacological profile of the new substance has been established during phase I on the basis of which a decision for the continuation of the clinical trial is made, the aim of phases II and III is now to answer the important questions of the therapeutic efficacy and tolerability in a large number of patients with the target indication. Due to the continuously increasing time and costs of drug development, drug development should be streamlined combining preclinical and early clinical phases as an exploratory stage and later clinical development as a confirmatory stage. The development and appropriate use of surrogates and models may be helpful to determine drug actions in human and to assist in dose selection as the main requirement for a successful large clinical trial in the confirmatory stage. Identifying the genes responsible for the huge variations in how different patients respond to a drug, in terms of both the product's effectiveness and its side effects, and genotyping patients before including in large clinical trials may prevent selecting the wrong patient population and avoid expensive repetition of these studies. Taking responsibility as the link between research and development gives clinical pharmacology a major opportunity to assume a pivotal role in drug development. To reach this goal, clinical pharmacology must be fully integrated in the whole process of drug development from the candidate selection until the approval. PMID- 10851847 TI - [Testing, estimating, "significance"--concept and language of medical biometry]. AB - The justification for the application of medical treatments and diagnostic methods has gained importance during the last decades. Together with scientific argumentations also the concepts of medical statistics have found a widespread use and are nowadays indispensible tools for clinical research. In publications of clinical trials, epidemiological investigations and animal experiments p values, statistical tests and confidence intervals help the reader to evaluate the significance and relevance of the provided results. A prerequisite is a clear understanding of the concepts underlying these methodological terms. This paper presents the basic concepts of medical statistics and demonstrates the conclusions that can be drawn with the aid of these concepts. PMID- 10851848 TI - [Basic principles of clinical therapeutic studies--what, how and why?]. AB - Treatment evaluation is one of the most important tasks in medical research. Detailed standards have been developed during the last decades. The efficacy/effectiveness of treatments can only be assessed in comparison to control groups. To guarantee the internal validity of these comparisons, the groups have to be comparable at the beginning of the study. This can be achieved by randomized allocation of patients to treatment. Furthermore, as far as possible patient and physician should be blinded to treatment in order to avoid subjective influences on treatment results. Groups should still be comparable when analysing the trial, thus an analysis according to the principle of intention-to-treat ("as randomized") should be performed. These indispensable principles for design, conduct and analysis of clinical trials are widely accepted and contribute to reliable and credible results. PMID- 10851849 TI - [Methodology of diagnostic validation studies. Errors in study planning and evaluation]. AB - The evaluation of diagnostic tests play an important role in clinical research. Though detailed standards have been developed for treatment evaluation, widely accepted methodological standards in diagnostic test research are still lacking. Following recent methodological research German biostatisticians usually distinguish between 4 types of diagnostic validation studies (Phase I to IV). The diagnostic value of a test can be described by indexes like sensitivity, specificity and predictive values. Sensitivity and specificity are so-called nosological probabilities whereas predictive values describe the probabilities of the disease given a positive or negative test result, respectively. These posterior probabilities depend on the prevalence of the disease in question. The methodological problems that arise in the planning of diagnostic studies differ from those encountered in treatment evaluation. This contribution presents an annotated checklist of the most important faults and shortcomings in the planning and the analysis of diagnostic validation studies. Avoiding these deficiencies may help that the study results are accepted by biostatisticians, clinicians and cost units. PMID- 10851850 TI - [Statistical models and evaluation of observation studies]. AB - Although randomized clinical trials can be regarded as the gold standard in many fields of clinical research, non-randomized (so called observational) studies are indispensable. Methodologic quality of such studies is at least as important as that of randomized trials but has been much less standardized. The analysis and interpretation of the results of observational studies is a very demanding job which needs a high level of knowledge and experience--medical as well as statistical. Basis for these analyses are complex statistical models the grounds and application of which are discussed in the following article. PMID- 10851851 TI - [How can "significance" and "relevance" be combined?]. AB - Clinical trials are aimed at providing results which enable improvements in patient care. It is widely criticized, however, that the characterization of results as "significant" or "non-significant" does not allow any assessment of their clinical relevance. To counter this criticism 2 biostatistical concepts are available: the use of confidence intervals and the application of statistical tests with shifted null-hypotheses. Possibilities and limitations of these concepts are discussed in this contribution. PMID- 10851852 TI - [Monitoring clinical studies. Development, measures and consequences]. AB - BACKGROUND: Clinical trials play an important role in developing and establishing new therapeutic and diagnostic procedures. In the planning and execution of these trials procedures and measures which allow for continual observation, description and evaluation of the study progress are to be taken into account. This is due to ethical, scientific and economic considerations. Together these procedures and measures are termed "monitoring". Repeated evaluation of the main study question is one particular monitoring measure. Results from this sequential procedure may lead to an early termination of patient recruitment. In the last 3 decades methods for interim analyses were developed which take into account the increased chance of errors when evaluating repeatedly the same question. By adjustment they guarantee a prespecified level of significance in the end result of the trial. Even though statistical significance may be evident in an interim analysis, this has not always to result in early termination of the trial. The decision to end a trial early must include other considerations than the mere evaluation of the main study question. In particular, consequences of the decision such as credibility and transferability of the trial result to subsequent therapeutic routine application are to be discussed. PMID- 10851853 TI - [Sense and nonsense in post-authorization surveillance]. AB - Post-authorization surveillance studies analyze the intended use of medications in accordance with their authorized labeling. They are characterized by the principle of non-intervention, i.e. they do not take influence on the decision whether or how to treat and to survey treatment in the individual patient. These aspects define the difference between surveillance studies and phase-IV clinical trials. In consequence, surveillance studies--in contrast to phase-IV trials--are not subject to the regulations of sections 40, 41 of the German Drug Law and national and international Recommendations on Good Clinical Practice (GCP). In essence, they are pharmaco-epidemiological investigations that make use of personified patient data. Within the context of the European regulatory policies they are part of the overall pharmacovigilance effort. In Germany, surveillance studies may have an additional regulatory impact. Indeed, they may be accepted as a relevant extension to the documentation on both safety and efficacy and in some cases even as an alternative to the tedious experimental investigation thereof, provided they meet newly defined quality criteria (that are discussed here more extensively). Although not developed to this purpose and although these recommendations are mainly formal with some relevant shortcomings in defining content quality, they are an important source of reference for the individual physician to guide his decision about the value and acceptability of any surveillance study, including those not primarily intended to be used in a regulatory context. PMID- 10851854 TI - [Meta-analysis as a tool for gaining knowledge]. AB - In these days, more than one clinical trial is mostly performed to evaluate a new treatment or therapeutic intervention. This necessitates a combined evaluation of their results. An integration of evidence from several trials is also helpful to determine the actual knowledge. These are the main goals of meta-analyses. Since the end of the 80s meta-analyses are widely used in clinical research. At the beginning of a meta-analysis, a protocol has to be developed. Similar to a protocol of a clinical trial, the inclusion and exclusion criteria for trials, the hypotheses and the planned analyses have to be fixed. After a careful localization of trials, a combined statistical analysis is performed. An investigation of heterogeneity, i.e., differences between study results, is indispensable. During the last years, the tool meta-analysis has been criticized. The criticism mainly results from poorly conducted meta-analyses which generated results without prespecifying hypotheses or which merely combined study results. Well-planned meta-analyses, on the contrary, have an increasing influence in clinical research. PMID- 10851855 TI - [CONSORT--what a proper scientific publication should supply]. AB - The publication of the CONSORT Statement was aimed at the improvement of the quality of research reports of randomized controlled clinical trials. It contains the most important standards of high quality research. Although originally meant for randomized studies only most aspects of this statement can be applied to other kinds of clinical studies. The following--and last--article of this series on medical biostatistics presents and comments the most important criteria. PMID- 10851856 TI - [Recruitment strategies for a randomized clinical study]. AB - BACKGROUND: To value the level of acceptance of the Memphis Study, (a random clinical study suggested verifying the efficacy of a diet enriched with phytoestrogens to prevent menopausal problems), and underlining the reasons which induce the acceptance or refusal of the study. STUDY DESIGN: Prospective study. METHODS: Meeting groups have been held with 82 women needing Day Hospital treatment for Menopause at III Department of Obstetrics and Gynecology of the University of Bari. The features of women and the reasons that induce these++ acceptance or refusal of the study were valued by an anonymous questionnaire with a precoded reply. RESULTS: 92.9% of the women accepted participation in the study. The main reasons for acceptance were: 1) believing that the study was drawn up for women; 2) it was done by expert physicians; 3) it was not done for financial gain. The random standard was the main reason for refusal. CONCLUSIONS: With this method we had a very high acceptance, talking over the goals and problems that study aimed to address and to resolve. The chance offered to judge the reliability and competence of physicians is important. PMID- 10851857 TI - [Lifestyle and drinking habits during pregnancy]. AB - BACKGROUND: It has been reported that a high consumption of alcohol during pregnancy is correlated to fetal alcohol syndrome and other perinatal problems. Less is known about the effects linked to moderate and limited drinking during pregnancy. METHODS: A study was carried out in 140 pregnant women: the mean age was 33.05 +/- 5.22, whereas the gestational age was 28.56 +/- 12.1 weeks (range 5 42). Each woman was asked to compile a form giving personal details and details of psychosocial behaviour, a lifestyle questionnaire (CAGE) and a form entitled "How much do you drink" based on SQQ. RESULTS: 22.2% of women reported that they had drunk alcohol during the month prior to the interview. Of these, 15% had not altered their habits, 0.8% had increased their alcohol consumption and 6.4% had reduced it. Wine was the form of alcohol most frequently consumed. Only one woman was positive to the SQQ test, and only one pregnant woman was positive to CAGE. CONCLUSIONS: This study shows that dietary behaviour was correct in the majority of cases. PMID- 10851858 TI - [Territorial research on the availability of phytoestrogen rich foods]. AB - BACKGROUND: To verify the availability of foods containing phytoestrogens, the quality of available products, the degree of knowledge of these foods by dealers, the willingness of dealers to give cooperation in preparation of the MENFIS study, a study assessing the efficacy of a phytoestrogen rich diet on the long term effects of menopause. METHODS: Perspective research by questionnaires carried out in the herbalist's shops of Bari. RESULTS: For the aims of the study the herbalist's shops resulted in being the most suitable, because they were the most supplied with these products and with natural foods, with a high degree of knowledge of these foods and a helpfulness towards the users, perhaps because of the strong motivation towards this type of diet. CONCLUSIONS: The easy availability of these products, the good preparation of managers of the herbalist's shops and their good experience is a sound basis for those intending to start dietary programs with phytoestrogen rich foods. PMID- 10851859 TI - [Obstetric conduct in IUGR]. AB - Purpose of the study is to identify the correct attitude that the obstetrician must engage in the management of pregnancy and birth in case of IUGR. Different methods of diagnosis and therapy of IUGR and the formalities of assistance to the birth have been examined and compared. Accurate clinical examinations of the mother, the study of fetal kariotype and ultrasonography, are essential for the diagnosis of IUGR. The genetic study could be performed by collecting chorionic villi, amniocentesis, cordocentesis or placenta biopsy. Ultrasonography identifies the cases of IUGR, and distinguishes early IUGR from late IUGR. Color Doppler identifies the pathology of the flow in the umbilical artery, in the abdominal aorta and in the middle cerebral artery. After the 26th week, the follow-up of the fetus with IUGR is done with cardiotocography with or without acoustic stimulation or oxytocin. The amelioration of maternal conditions is obtained by avoiding the cigarette smoking, preferring to rest in bed and a balanced feeding; the hyperoxygenation doesn't find unanimous consent. The treatment off IUGR can consist of abdominal decompression, intra-abdominal infusion of amniotic liquid, or use of aspirin. The birth is carried out in the hospital, when the fetus has reached a sufficient maturity. The management of IUGR requires an accurate follow-up and an adequate antepartum therapy. The goal is a birth with less risk. PMID- 10851860 TI - [Treatment of maternal hyperthyroidism and fetal goiter]. AB - A fetal goitre is a potentially dangerous phenomenon because of mechanical obstruction and possible fetal thyroid function disorders. During pregnancy women with a history of Graves' disease under treatment with propylthiouracil (PTU) have an increased risk for fetal goitre. In this report a patient with Graves' disease diagnosed in early pregnancy and treated with PTU which resulted in a fetal goitre is described. The fetal thyroid status, investigated by percutaneous fetal umbilical cord blood sampling, was normal and the reduction of PTU dosage was sufficient to decrease goitre volume. PMID- 10851861 TI - Effective treatment of beta-thalassemia intermedia during pregnancy with rHuEpo. A case report. AB - The anemia of pregnancy in presence of beta-thalassemia intermedia usually aggravates pregnancy procedure. In the present study we investigated whether the administration of recombinant human erythropoietin (rHuEpo) recombinant human erythropoietin in combination with iron and folic acid may ameliorate blood indices as an alternative choice to blood transfusion. PMID- 10851862 TI - [Epidemiological study of Chlamydia trachomatis infection in a sample of women requesting voluntary termination of pregnancy, treated with rokitamycin]. AB - BACKGROUND: Infection with Chlamydia trachomatis usually involves the cervix uteri, causing no symptoms, and may easily be unrecognised and untreated until troublesome symptoms arise, such as pelvic inflammatory disease, which can affect fertility and reproductive life. Therapies include the macrolide antibiotics, and in this class rokitamycin offers marked lipophilia, excellent intracellular penetration, and bactericidal activity at concentrations close to the MIC. The present study was therefore designed to establish the frequency of intracervical infection with Chlamydia trachomatis in women applying for termination of pregnancy, and to assess the efficacy and safety of this drug in this indication. METHODS: Women aged 18-40 years were admitted for termination of pregnancy, with a positive cervical swab for Chlamydia trachomatis. The study was conducted in accordance with the Declaration of Helsinki and amendments. Patients were given one oral tablet of 400 mg rokitamycin in the morning, and one in the evening, for two weeks. Treatment started ten days before the termination, within 48 h of taking the swab. Partners were to receive the same treatment. RESULTS: 292 women requiring termination of pregnancy, on average at the 9th week of pregnancy, were assessed. Of these, 24 (8.2%), mean (+/- SD) age 27.1 +/- 6.1 years, range 18-39, with a positive cervical swab for Chlamydia trachomatis, were treated with rokitamycin; 22 of their partners were treated too. Forty days after the start of treatment 22 patients (92%) gave negative results; these were all the cases whose partners had received treatment. No adverse events were reported and the acceptability of the treatment was considered good or excellent in 91% and fair in 9% of the cases. CONCLUSIONS: The findings confirm that rokitamycin is one of the most useful and effective macrolides for the treatment of infections caused by intracellular microorganisms; it is extremely well tolerated and has marked microbiological efficacy. PMID- 10851863 TI - [Motives for a revision of the physiology of labor and an updating of obstetric terminology]. AB - During the second half of our century, revolutionary progress has been made in obstetrics in terms of care, prevention and technology, but curiously a number of doctrinal questions have been left unresolved which are more important in cultural and practical terms than is commonly thought. We refer in particular to the physiology of the mechanics of labour whose inveterate lacunae of interpretation and major conceptual inaccuracies would be easily resolved if tackled with sufficient interest. These are easy questions to resolve because the missing or incorrect aspects of the traditional model used to interpret the mechanics of labour are for the most part only caused by clashes with the basic principles of physical science. If the mechanics of labour are re-examined with a view to anchoring them rigorously and satisfactorily to the principles of physics, the traditional model of interpretation, which is currently in vogue, is completely invalidated and replaced by another based on a series of hydrodynamic phenomena of considerable obstetric interest. The advantages of these hydrodynamic phenomena are represented by the onset of the so-called "pelvic press", doubling the motor forces of labour during the release of the fetus, the constant maintenance of a balance of forces in the fetal environment, the rotary movements caused autonomously by a hydraulic and muscular system of fixing the womb during the release of the fetus, and above all by the equal interaction of the fetus in the mechanics of its own birth. Clearly, this new interpretative model calls for the use of partly updated and partly innovative obstetric terminology. PMID- 10851864 TI - [Indications for specialized quenching. Superficial burn in the intestines]. PMID- 10851865 TI - [New enemy for our hearts. What do we know today about the role of Chlamydia?]. PMID- 10851866 TI - [Risk stratification in acute coronary syndrome. Practical procedure depends on severity]. PMID- 10851867 TI - [Complementary medicine--popular and controversial. Are there "solid data" for general practice?]. PMID- 10851868 TI - [Red card for chelate therapy. Complementary medicine, 1]. PMID- 10851869 TI - [Increasing depression in children and adolescents. Only early intervention can prevent dire outcome]. AB - Numerous recent epidemiological studies indicate that depressive disorders in children and adolescents are quite common. Roughly 15% of adolescents admit to having suffered from such a disorder at some time or other. Depressive disorders increase with age with a preponderance of girls over boys. Risk factors that have been found to be associated with depressive disorders include parental psychopathology, familial dysfunction, and negative life events. Depression frequently occurs together with other disorders, and shows a tendency to become chronic. Finally the article discusses the implications of the latest findings for preventive and interventional strategies. PMID- 10851870 TI - [Insulin and oral antidiabetic drugs. How to combine them in type 2 diabetes?]. PMID- 10851871 TI - [New morphine preparations with extended effect duration. 2 treatment observations]. PMID- 10851872 TI - [Compression instead of support. Thrombosis ABC, 16: Compression in venous diseases]. PMID- 10851873 TI - [Exanthematous infectious diseases in childhood. 3: Viral and bacterial toxin induced exanthema]. PMID- 10851874 TI - [Diagnostic quiz. Small hole with big sequelae. Spontaneous pneumothorax with mediastinal shift (minor tension pneumothorax)]. PMID- 10851875 TI - [Hamburg methadone contract expired. Is drug substitution at an end?]. PMID- 10851876 TI - [Have you allowed yourself to be talked into life insurance? There's a better way to prepare for old age]. PMID- 10851877 TI - [MEDI GmbH Medical Society. Does membership revoke individual rights?. Interview by Dr. med. Annette Paul]. PMID- 10851878 TI - [Report of experiences. A grip on migraine, "sleeping off" tension headache]. PMID- 10851879 TI - [Smoking cessation. Power act with patches, spray and chewing gum]. PMID- 10851880 TI - [Recent developments in therapy of condylar fractures of the mandible]. AB - Due to advances in surgical methods, such as miniplate and lag screw osteosynthesis, there is current discussion about the adequate therapy of condylar fractures. Regarding the insufficient outcome of conservative treatment confirmed by refined methods of functional diagnosis, there is a trend towards surgical therapy, at least in cases of condylar displacement out of the fossa. This trend was also prompted when surgeons learned that the condyle determines local growth but does not serve as a growth center of the mandible. Along with increased interest in temporomandibular function and surgical therapy of internal disorders, surgical therapy of traumatic soft tissue lesions was emphasized. This consequently led to surgical restoration of intraarticular fractures. Conservative treatment will nonetheless be sufficient in cases of minor displacement due to a higher ability of remodeling, even more so in children. PMID- 10851881 TI - [Primary thinning and de-epithelialization of microsurgical transplants from the lateral thigh]. AB - To expand the indicational spectrum of the myocutaneous vastus lateralis flap, which is often too voluminous for intraoral application, we performed extreme, primary thinning of the fat and muscle component of this microsurgical transplant in 14 patients. After subfascial localization of the 0.5- to 1.0-mm-thick perforating vessel, it is exposed through the fascia and muscles up to its exit from the descending branch of the lateral circumflex femoral artery. After isolating the perforating vessel, it is no longer necessary to include parts of the vastus lateralis muscle in the flap. The fatty tissue of the remaining epifascial fat component is completely removed except for a ca. 1- to 2-cm-wide cuff of fatty tissue and fascia around the perforating vessel. When performing this primary radical removal of the subcutaneous fatty tissue, care should be taken not to injure the deep subdermal vascular plexus. In addition to the thinning procedure, de-epithelialization of the skin was performed using scalpel blade dissection (five patients) or carbon dioxide laser (6 W, five patients). This thinning technique was used for covering ten intraoral and four extraoral defects and enabled the raising of skin flaps with a thickness of 3-5 mm even in obese patients. The vessel pedicle length of thinned flaps was between 12 and 16 cm; flap size varied between 4 x 5 and 9 x 15 cm, and the donor sites were directly closed. In one case, there was a partial necrosis (20%), but the remaining flaps healed without complications. On the intraoral flaps, a thin, smooth and pliable surface developed after re-epithelialization within 3-6 weeks. The described method expands the application possibilities of the myocutaneous vastus lateralis flap for a large number of intraoral and flat defects with minimal donor-site morbidity. PMID- 10851882 TI - Velopharyngoplasty according to Sanvenero Rosselli. Historical review and contemporary judgement. AB - The history of the pharyngeal flap and the contributions of Schoenborn, Sanvenero Rosselli, Padgett, Sercer and Rosenthal are described. The summary Divisione palatina e sua cura chirurgica in the Italian/French language is translated into English. PMID- 10851883 TI - [Incidence, risk factors and follow-up of sensation disorders after surgical wisdom tooth removal. Study of 1,106 cases]. AB - A study was carried out to determine the risk of dysesthesia of the inferior alveolar and of the lingual nerve after molar surgery. A total of 1103 impacted lower wisdom teeth and 3 impacted lower second molars were removed in 687 patients, all of whom with unaltered sensibility preoperatively. Clinical, radiological, and surgical factors of each case were recorded. Postoperative disturbances in the sensibility of the lip and tongue were evaluated by neurological examination. Follow-up was carried out for a maximum of 35 weeks. Dysesthesia of the inferior alveolar nerve occurred with an incidence of 3.57%. The lingual nerve was injured in 2.1% of patients. Most of the initially reported alterations in sensation resolved within the follow-up period. Dysesthesia of the inferior alveolar nerve persisted in 0.91%, and of the lingual nerve in 0.37%. However, the extent of the prolonged impairment was slight in general. The effect of the documented factors on the incidence of dysesthesia was analyzed. For the inferior alveolar nerve, analysis revealed significant effects in older patients, for completely developed roots, for deeply impacted teeth, in the radiological relationship of the roots and the inferior alveolar canal, for difficult surgery, and for intraoperative exposure of the nerve. The surgeon and the anesthesia had a significant influence on lingual dysesthesia. PMID- 10851884 TI - [(18F)-2-fluorodeoxyglucose PET. Prospects for secondary prevention of mouth cavity carcinoma]. AB - AIMS: The predominant cause of death due to oral cancer is the failure to control local tumor due to regional tumor recurrence. The sequelae of surgical resection and high-dose irradiation cause substantial changes in head and neck anatomy, leading to considerable problems in the early morphological detection of recurrent disease. Therefore, this study evaluates the verification of cancer recurrence by means of its pathologic glucose metabolism. MATERIALS AND METHODS: We reviewed a total of 50 [18F]-2-fluordeoxyglucose positron emission tomography (18FDG-PET) investigations performed in 44 patients who had undergone surgical resection of oral cancer. In 23 cases, re-staging (group A) was indicated due to suspicion of recurrent or secondary tumor manifestation. In 27 cases, PET served as a screening procedure (group B). Statistic evaluation included sensitivity, specificity, positive/negative predictive value and accuracy of 18FDG-PET for the detection of tumor manifestation. RESULTS: 18FDG-PET correctly identified 23 of 26 tumor sites (88%) in the re-staging group and 9 of 10 tumor sites (90%) in the screening group. We encountered a total number of 16 false-positive foci with an increased 18FDG-uptake. In six patients, 18FDG-PET detected tumor recurrence several months before a morphological correlative could be identified. In 5 of these 6 patients, the PET findings for the latter tumor sites determined the patient's fate. Specificity was 63% for local recurrence, 97% for secondary lymph node involvement and 90% for distant metastasis. CONCLUSION: According to these data, 18FDG-PET is the most effective diagnostic tool in the follow-up of oral cancer patients to date. Due to the high prevalence of recurrent disease in the follow-up of oral cancer, either the detection of early recurrence or the identification of additional, incurable tumors may add substantially to a rational therapeutic management. We therefore recommend 18FDG-PET for screening and re-staging of recurrent oral cancer. PMID- 10851885 TI - [Position and mobility of the articular disk after surgical management of diacapitular and high condylar dislocation fractures of the temporomandibular joint]. AB - Magnetic resonance imaging (MRI) assessment of traumatized temporomandibular joints (TMJ) usually focuses on disc position, defining regular joint function by normal, excentric or displaced disc position. So far, there are only few reports regarding disc position after open reduction of diacapitular or high condylar fractures of the TMJ with dislocation. The aim of the present study was to evaluate the role of the disc as regards postoperative functional outcome by electronic axiographic recordings of condylar movements and MRI, displacement of the disc and lesions of TMJ soft tissues being frequent in this type of mandibular fractures. A total of 30 subjects with 37 condylar fractures in whom osteosynthesis was performed using a preauricular approach were imaged postoperatively (mean 24 months) with a 1.5-Tesla MRI system to determine, (a) the position of the disc, (b) the range of mobility of the disc and (c) condylar mobility in closed and open mouth position, comparing fractured sides (FS) vs nonfractured sides (NFS). Linear movements between the two jaw positions in the sagittal plane were measured by superimposing transparencies. The results indicate: (1) more than 70% of the discs (FS) were found to be in normal position; there was no disc displacement without reduction. However, these data stood in contrast to severe limitations of the axiographic tracings as presented by almost 30% of the subjects. (2) Significant correlations were found between fixed (alpha = 0.05) or highly immobilized (alpha = 0.01) discs and axiographic limitations, suggesting disc mobility to be a valuable parameter for assessment of the postoperative functional outcome. PMID- 10851886 TI - [Therapeutic concept in combined orthodontic-surgical treatment of class II malocclusion with short face syndrome. New aspects for surgical lengthening of the lower face]. AB - The main aesthetic concern of patients with Angle class II deformities with skeletal deep bite--short face syndrome--is the short lower face. It is the orthodontist's task to correct this deformity insofar as possible. Depending on the extent, even orthodontists at the beginning of their training will recognize this. Since presurgical orthodontic treatment determines the kind and extent of the surgical procedure, the orthodontist has to plan the treatment in for individual case. It is the purpose of this article to demonstrate a concept of treatment for patients with class II deformities, skeletal deep bite, and a short lower face. Presurgical orthodontic treatment and the surgical procedure to correct the deformity are discussed. The treatment results show that it is necessary to leave or to create a certain curve of Speed, depending on the extent of the deformity, to achieve a satisfactory result in terms of to function, aesthetics, and stability. It can be concluded that it is only possible to reach the preset treatment goals with an exact diagnosis and the knowledge of the necessary preparation in combination with the surgical procedure. PMID- 10851887 TI - [Dose response relations of collagen IV expression in irradiated submandibular gland of the rat]. AB - The extent of radiogenic salivary gland damage depends on the radiation dose, the fractionation, and the localization of the gland in the radiation field. Because extracellular matrix proteins, for example collagen IV, belong to the main components of basement membranes (BM), which are considered to posses cell- and structure-regulating functions, this study set out to describe radiogenic BM changes. The staining profile of collagen IV was studied in 110 female rat mandibular glands depending on age (1 year vs. 1 1/2 years), dosage (2 Gy/d; total dosages 20, 40, or 60 Gy), the radiation field (inside vs. outside), and on the latency period (1/2 year vs. 1 year), using immunohistochemical methods. The stainings were assessed semiquantitatively and differences were evaluated using multivariate analysis. The staining pattern of the polyclonal antibody in rat tissues did not differ from the pattern found in human salivary glands. Collagen IV was detected at variable levels in the glandular and nerve tissue and in vascular walls (negative: adventitia). Irradiated tissues were stained more strongly than non-irradiated, and differences were significant in salivary glands exposed to more than 20 Gy. Age and the latency period had no significant effect on staining. The BM constituent collagen IV showed dose-dependent increasing expression analogous to the salivary gland damage, which could contribute to disturbed cell-matrix interactions following salivary gland radiation exposure. Several tissue structures may be more sensitive to possible scattered radiation. Information on the pretreatment status is mandatory in histopathological studies on the BM of salivary glands. PMID- 10851888 TI - [Prevention in pulmonology]. AB - In the fight against respiratory diseases the well established most effective strategy is their prevention. Mainly the field of screening-caring--so called the second and third levels in prevention--belongs to the competency of the pulmonology specialty, while the key-issue of primary prevention consisting of the reduction of environmental harm and health education is not--or at least not only--the responsibility of the health care system. In the light of national epidemiological data BCG vaccination of the new-born is necessary, however, the system of revaccination of school-age children needs revision. The system of chest screening by rtg nowadays is adapted to the local prevalence of tuberculosis; it is needed to concentrate to high-risk subjects. The effectivity of radiological screening now is recognised internationally and this perspective partly based on our national results. In the care of patients with chronic obstructive pulmonary diseases (COPD) the key-issue is the identification and regular follow-up, care of high-risk subjects with rapid decrease in lung function by effective screening. The most important issue in this work is smoking cessation. Of course, if we could prevent smoking addiction, we could significantly reduce the number of several other widespread diseases, not only lung cancer. PMID- 10851889 TI - [Snake bite injuries]. AB - Authors treated five patients who suffered venomous snake-bite injury. Although these snakes are not native in Hungary, this kind of injury is estimated to be more frequent, because of the increasing number of the private collections and illegal import of these reptiles. The local and general symptoms, the therapeutic steps are summarised in this study considering the literature as well. Two patients did not show any systemic or local symptoms at the level of injury, they needed only short observation, and woundcare. The other three patients had serious transient systematic symptoms (vasolability, hypotension/shock, coagulopathy, confusion). Two of them were given specific antivenom. As the third patient did not agree with the serum therapy, plasmapheresis was the choice to treat him, and it seemed to be effective. Few hours later the patients needed surgery because of serious compartment syndrome of their affected upper extremity. Surgical decompression of all the compartments and different possibilities of the secondary skin closure technique are demonstrated. Two patient healed completely, but the right thumb of the third was lost. Authors summarise the effects of the poisons, the symptoms, and the basic therapeutic steps during the first aid and in the primary hospital phase, respectively. They point out the indications of the serum therapy and the correct surgical decompression of the injured extremity. PMID- 10851890 TI - [Initial experiences with 123-Iodine-IBZM neuroreceptor scintigraphy in extrapyramidal disorders]. AB - The receptor scintigraphy of the dopaminergic system of the brain is of interest in the evaluation of movement disorders. The 123I-IBZM is a radiopharmaceutical with affinity predominantly to postsynaptic D2 receptors. The aim of this study was to evaluate the role of IBZM SPECT investigations in the differentiation of disorders with Parkinson's syndrome. Eight patients with idiopathic Parkinson's syndrome and 8 patients Parkinson's syndrome with other etiology were investigated with 123I-IBZM SPECT (6 females, 10 males, mean age +/- SD: 59 +/- 9). The patients according to the clinical signs and symptoms, results of CT/MRI and rCBF SPECT investigation were categorized. The reconstructed SPECT slices were evaluated visually and quantitatively. The visual interpretation of the images were performed by two observer and scored the radiopharmaceutical uptake (from 1-3) of the cortex and the striatum separately. For quantification striatum/frontal cortex activity ratio were calculated with ROI technique. The differences between the patient groups were statistically analyzed by two tailed t-test. The IBZM uptake were different in the two group of patients. The striatal IBZM accumulation was higher in the idiopathic Parkinson's syndrome patients compared to the other parkinsonians. The striatum/frontal lobe activity ratio was 1.69 +/- 0.9 (mean +/- SD) in the right, 1.67 +/- 0.04 (mean +/- SD) in the left hemisphere of the patients with idiopathic Parkinson's syndrome. The corresponding data in the nonidiopathic parkinsonian group were 1.53 +/- 0.06 (mean +/- SD), 1.52 +/- 0.04 (mean +/- SD) respectively (p < 0.01). The quantitative data correlated with the results of the visual evaluation. According to the data presented IBZM-SPECT is an effective tool in the differentiation of disorders with Parkinson syndrome. PMID- 10851891 TI - [Lipomatous meningioma: report of two cases and review of the literature]. AB - Lipomatous meningioma is a benign tumor characterized either by an admixture of mature adipocytes and meningioma or the production of triglycerides by neoplastic meningothelial cells assuming a lipoblast-like appearance. The authors report on two instances of this exceedingly rare lesion occurring in the left middle cranial fossa and over the right frontal convexity of two female patients aged 79 years and 60 years, respectively. In the former, the tumor was an incidental autopsy finding, while the latter underwent surgery for symptoms of intracranial space occupation. Light microscopy showed interwoven islands of fatty tissue and transitional meningioma in the first case; whereas a monomorphous signet-ring cell phenotype prevailed in the second. Oil-Red-O staining confirmed the presence of neutral fat in both specimens. Immunohistochemical coexpression of epithelial membrane antigen, vimentin, and S100 protein supported the meningothelial origin of tumor cells. On the other hand, the CD 68 macrophage antigen was not detected. Cytoplasmic lipid droplets along with hallmarks of meningothelial differentiation were visualized ultrastructurally in part of the meningioma component of the first case and throughout the second. These findings are consistent with a metaplastic origin of the adipocytic element. Whatever its histogenesis, lipomatous meningioma may, on occasion, represent a major challenge with therapeutic implications for both preoperative imaging and histological diagnosis. PMID- 10851892 TI - [Physicians in developing Hungarian veterinary medicine and veterinarians' education]. PMID- 10851893 TI - [Professor Geza Illyes (1870-1951)]. PMID- 10851894 TI - [The accessibility of prophylactic care for the people comprising a risk group]. PMID- 10851895 TI - [The social hygiene characteristics of the health status of the population under the current conditions]. AB - Time course, tendencies, and sociohygienic characteristics of population morbidity and mortality with regard to the major diseases have been evaluated by investigation of population morbidity carried out in association with censuses and analysis of official statistical data and interviews of physicians and populations in 1991-1998. Differences in the health status of individual population groups, quality and level of their life are analyzed. PMID- 10851896 TI - [The role of prevention in decreasing endocrine pathology]. AB - Time course of changes in the incidence of thyroid diseases and their association with other noncontagious chronic diseases (NCCD) have been evaluated among therapeutic patients hospitalized at Research Center for Clinical and Experimental Medicine of Siberian Division of Russian Academy of Medical Sciences in 1983-1995. The incidence of thyroid diseases was low in 1983, when prevention of iodine-deficiency diseases was a state program. In 1989, when iodine deficiency prevention was reduced, the incidence of cases with enlarged gland and its dysfunction has drastically increased. In 1995, when the system of population protection from endemic goiter virtually ceased to function, the incidence of thyroid diseases increased, including such serious diseases as diffuse toxic goiter, nodular goiter, primary hypothyrosis, and autoimmune thyroiditis; the number of patients in whom thyroid diseases were concomitant with other NCCD increased. These results demonstrate the principal significance of the preventive trend in public health. PMID- 10851897 TI - [Ownership in public health]. AB - The authors attempt to bind the known theoretical concepts on the property to their interpretation and realization in actual public health. They specify and offer examples of their perception from a modern viewpoint and analyze their significance for further reforms in public health. PMID- 10851898 TI - [The ethical-legal problems of physician and patient relationships]. AB - The paternalistic pattern of doctor-patient relationships will be evidently retained in Russian medicine in the nearest future. Therefore the society should be steadily step-by-step prepared to perception of the material values most fully ensuring the patient's rights in the course of therapeutic cooperation. PMID- 10851899 TI - [The development of organizational technologies in the optimization of the medical care for the population of a region]. AB - This paper presents a model of a new type of outpatient medical care: specialized course of therapy on an outpatient basis. Indications for rendering such care mainly to patients with chronic diseases are formulated. Treatment of such patients in an outpatient setting by intermittent courses with possible short term hospitalization and temporary invalidity conforms to the standards of hospital care. Data on social and medical efficiency of such treatment organization are presented. PMID- 10851900 TI - [The problems of medical support for the population under the current conditions (based on the data from special sociological research)]. PMID- 10851901 TI - [The social hygiene aspects of monitoring and assessing the efficacy of the activities of treatment and prevention institutions]. AB - The proposed system for evaluating the quality and efficiency of medical care, including universal parameters with quantitative expression, allows the comparison, evaluation, and mathematical processing of these parameters. The method helps detect violations of medical technologies, irrational utilization of resources, directly compare the results and use them for preliminary testing of validity of the managing decisions. PMID- 10851902 TI - [The outpatient forms of the organization of social medical care for older persons in Russia]. AB - Presents data on the non-hospital forms of medicosocial care of elderly and senile subjects in the Russian Federation, which proved to be the most effective as regards the economy and staff under conditions of the population aging. PMID- 10851903 TI - [A municipal order for the delivery of medical care to the population of a specific area as a variant in onging planning (the estimation of the critical services volume)]. PMID- 10851904 TI - [The organization of orthopedic-traumatological care for the patients in a large administrative center]. PMID- 10851905 TI - [The teaching of pathological anatomy at Moscow University. 4. Pathological anatomy in the General Statutes of Imperial Russian Universities]. PMID- 10851906 TI - [The closing of the Moscow Medical Surgical Academy]. PMID- 10851907 TI - [Prof. F. F. Mering and the internal disease clinic in Russia in the 2nd half of the 19th century (the history of the Faculty Therapeutic Clinic of Saint Vladimir University). 4]. PMID- 10851908 TI - [Someone has to put his foot down]. PMID- 10851909 TI - [Pathologic diagnosis--now and in the future]. PMID- 10851910 TI - [Risk of HIV transmission]. PMID- 10851911 TI - [Gastroesophageal reflux in children]. PMID- 10851912 TI - [Reflux disease and 24-hour esophageal pH monitoring in children]. AB - Gastrooesophageal reflux disease has a variety of symptoms in children. 24-hour pH monitoring in the lower oesophagus is the gold standard for documenting gastrooesophageal reflux. We present our experience with 24-hour pH monitoring in children. 150 pH recordings in 120 children were performed. Clinical background and results from pH monitoring were recorded, in addition to supplementary examinations and treatment. No complications were recorded, but ten recordings (8.3%) were unsuccessful. Mean age was 3.5 years (median 13 months; range one month to 15 years). 44% had a pathological reflux index. Indications for pH monitoring were dominated by regurgitation/vomiting (63%), failure to thrive (45%) and respiratory symptoms (32%). Of the supplementary examinations performed, upper gastrointestinal contrast series provided no additional information (34 children), while endoscopy (20 children) showed oesophagitis in 11. Medical treatment was prescribed in 66% of the cases based on the pH monitoring results and clinical evaluation. Five patients were given anti-reflux surgery, and ten received gastrostomy. Our experience with this recording technique is good. pH monitoring should be available in paediatric departments, as a large number of the recordings had clinical consequences for the patient. PMID- 10851913 TI - [Esophageal stricture as a complication of gastroesophageal reflux in children]. AB - General practitioners and paediatricians often get in contact with children who present with regurgitation and vomiting. To some extent, regurgitation is a normal situation in infants during their first months of life. Abnormal gastrooesophageal reflux, however, should be diagnosed and treated adequately, as there is a risk of complications. At the Department of Paediatrics, Haukeland Hospital, Bergen we have diagnosed oesophageal strictures due to undiagnosed gastrooesophageal reflux in six children. One patient had previously been treated for gastrooesophageal reflux during infancy. The main reasons for admitting the children were dysphagia and weight loss. When the correct diagnosis had been established, the patients were treated with cisapride and omeprazole orally. Several dilatations of the strictures were performed by upper endoscopy. A Nissen's fundoplication has been performed in two patients and is probably necessary to undertake in the remaining four. Clinical awareness of gastrooesophageal reflux is important, since undiagnosed reflux carries a risk of oesophagitis and stricture formation. PMID- 10851914 TI - [Sex differences concerning the habit patterns and health among intravenous heroin addicts in Oslo]. AB - The issue addressed below concerns sex differences in consumption patterns and ways of financing the drug habit among intravenous heroin users. The findings are linked to recorded differences in various health indicators. The article is based on data collected through 1,840 interviews with heroin abusers who contacted the "Needle-Exchange Bus" in Oslo during the years 1993-97. Analysis indicates the existence of substantial differences between women and men along a number of variables. Women report injecting more frequently than men and using on average more heroin per injection. Women also finance their drug habit differently, as more than half of them report some income from prostitution. The finding may seem paradoxical as women also appear to have a lower risk of dying from an overdose/poisoning and a lower risk of contracting hepatitis A and B than male injecting misusers. The risk of contracting HIV and other sexually transmitted diseases, on the other hand, appears to be greater among female injectors. PMID- 10851915 TI - [Use of methadone in the treatment of psychotic patients with heroin dependence]. AB - Comorbidity of psychosis and substance abuse has gained increased attention for some time. However, pharmacological treatment for opioid dependence among psychotic patients is seldom described. We present the treatment of four opioid dependent psychotic patients with methadone and antipsychotic medication. Three of the four patients had initially a period of increased psychotic symptoms, but all four patients have shown marked improvement following the introduction of methadone. Compliance with treatment has increased, they have had enduring non psychotic periods and a markedly reduced use of illegal substances. The results should obviously be interpreted with caution, given the small number of subjects. Further research is therefore important. PMID- 10851916 TI - [Bad shots--skin and soft tissue infections following intravenous drug abuse]. AB - Infection at the injection site following parenteral drug abuse is a well known complication. In Oslo, Norway's capital city with a population of 500,000, most of these infections are treated on an out-patient basis in the surgical department at Oslo Legevakt, a publicly funded primary health care facility. During the four last months of 1998, 179 patients were admitted with skin and soft tissue infections at the injection site compared to only 46 in the same period in 1993. This suggests that the problem is increasing. In this retrospective study these populations were analysed according to their age, sex, clinical appearance, and the treatment given. In 1998, 36 patients were admitted to hospital, the rest treated on an out-patient basis. A total of 112 patients were treated with simple incision and drainage, 63 of whom were given antibiotics. 37 patients were treated with antibiotics only. There were few complications; two patients with deep venous thrombosis and one in need of skin transplantation. We saw no development of life threatening infections among our patients. The article also gives suggestions for treatment. PMID- 10851917 TI - [What do health professionals ask RELIS Vest service and how satisfied are they with the answers?]. AB - RELIS Vest provides drug information by answering questions from mainly physicians and pharmacists. In the present paper we evaluated this service. All questions to RELIS Vest during the 1995-98 period were systematically categorized and analysed. The quality of the service was assessed by evaluation forms. Of a total of 875 queries, 393 came from physicians and 370 from pharmacists. 42% were about psychoactive drugs; the most frequent types of queries concerned documentation and adverse effects. Analysis of 180 evaluation forms showed that 79% of the physicians and 56% of the pharmacists found the answers fast enough, relevant, adequately comprehensive and with valuable references while 16% of the physicians and 29% of the pharmacists found that the answers satisfied three of these four criteria. We conclude that problem-oriented drug information based on actual cases of prescribing is highly appreciated. PMID- 10851918 TI - [Children and HIV/AIDS--prevention, follow up and treatment]. AB - At the global level, the HIV/AIDS epidemic has reached dramatic proportions. According to WHO, more than 1 million children are currently living with HIV infection. In Norway (population 4.4 million), a cumulative total of only 33 children with HIV infection have been reported, making HIV/AIDS a rare disease. In view of the limited experience with HIV/AIDS among Norwegian children, strategies for the clinical management of HIV infection are discussed in light of recent guidelines from the United States and Europe. Procedures for prophylactic treatment and follow-up of children born to HIV infected mothers, and procedures and follow-up of antiretroviral treatment of HIV infected children are proposed. PMID- 10851919 TI - [Children and adolescents selling sex]. AB - Our knowledge about prostitution is shaped by female street prostitutes, many of whom are drug addicts. Less is known about children and adolescents from the normal population who sell sex. Total cohorts of pupils in all Oslo's schools (N 10,812) in the age groups 14-17 filled in a questionnaire at school. 1.4% of the adolescents had sold sex at least one time, more than three times as many boys as girls. Mean age for first episode was 12.6 years in boys, and 14.1 years in girls, and seventy per cent had sold sex more than three times. There were no associations to indicators of parental social class or residential area in Oslo. The adolescents who had sold sex were more lonely and more often reported symptoms of depression and anxiety than others. A multivariate analysis showed that associations to conduct problems, alcohol problems and use of drugs were most important. The majority reported no contact with public services aiming at helping adolescents with psychosocial problems. A group of children and adolescents in Oslo sell sex. Boys are involved more often than girls, their customers are assumed to be bisexual or homosexual men. These behaviours fall into a pattern of conduct and substance problems, and some of these adolescents could have high long-term risk of antisocial problems and drug abuse. More knowledge is needed, and preventive work should be intensified. PMID- 10851920 TI - [HIV screening of pregnant women in Norway]. AB - The prevalence of HIV infection among pregnant women is low in Norway (4.5/100,000). HIV screening has been offered on a routine basis in antenatal care since 1987. 96% of all pregnant women are screened for HIV. After the introduction of the screening programme, effective treatment (zidovudin) has become available to pregnant women. This treatment reduces mother-to-child transmission of HIV by two thirds. By screening all pregnant women during a period of two years (95% confidence interval 1-6 years), transmission can be reduced to one child if the HIV positive women are offered and accept treatment. According to a review presented here, doctors and midwives involved in antenatal care do not seem to have good enough routines for giving women information about the screening test and for offering informed choices. PMID- 10851921 TI - [New diagnostic methods in non-Hodgkin lymphoma]. AB - We describe five patients with non-Hodgkin's lymphoma (NHL) in which immunohistochemical investigation (IH) and polymerase chain reaction (PCR) were important for diagnosis and choice of treatment. One patient got the diagnosis follicular NHL after PCR showed t(14;18) in a fine needle biopsy from a tumour in the abdomen. Two small and partly traumatized biopsies from two patients were diagnosed as low grade NHL of B-cell type after PCR showed monoclonality for B lymphocytes and IH showed B-cell phenotype and low proliferation rate. Biopsy from another patient showed positive immunohistochemical reaction for cyclin D1 and t(11;14) by PCR compatible with mantel cell lymphoma. One patient was diagnosed with high grade NHL of T-cell type after IH showed positive reaction for T-cell markers and high proliferation rate, and PCR showed monoclonality for T-lymphocytes. These cases demonstrate that IH and PCR can often benefit patients with NHL. The new methods contribute to enhanced diagnostic security and precision in subclassification and make possible the use of small biopsies which save the patients from major surgery for diagnostic purposes. PMID- 10851922 TI - [Immunohistochemical diagnosis by means of tyramide signal amplification]. AB - Immunohistochemistry is used for in situ detection of proteins in histological slides and is now an important diagnostic tool. Due to several methodological and biological factors, conventional immunohistochemical procedures may sometimes have too low sensitivity, especially for tracing the histogenesis of malignant tumours. A few years ago, an amplification technique was introduced which greatly increased the sensitivity of some of the commonly used immunohistochemical methods. This technique permits the use of primary antibodies in significantly lower concentrations compared with the conventional methods. Alternatively, one can keep the antibody concentration unchanged and use the enhanced sensitivity to detect scarce proteins, which are not visualised by traditional immunohistochemical procedures. We present a brief description of the technique and show some examples of its use in the diagnosis of neuroendocrine carcinomas. Tyramide signal amplification might become an important supplement for the diagnosis and classification of malignant tumours. PMID- 10851923 TI - [Puncture cytology]. AB - Limited public medical resources make it important to consider any diagnostic test which is inexpensive and reliable and without major side effects. The presented data are based on retrieval from the medical databases Medline and Cancerline, as well as the authors' clinical experience. The role of fine needle aspiration cytology for the diagnosis of solid and non-solid tumours is well established in the literature. The technique is simple and cost-effective and without major side effects. The sensitivity and specificity concurs that of histology in different tumours and may reduce the need for histopathology. In this review we discuss the combined use of fine needle aspiration cytology and modern immunocytochemistry and molecular techniques which may enhance diagnostic precision. We present the role of ancillary techniques which may prevent unnecessary extensive examinations and influence the therapeutic management of the patient. PMID- 10851924 TI - [How to achieve an optimal intake of vitamin D in children]. PMID- 10851925 TI - [Induced abortion--simple answers to difficult questions?]. PMID- 10851926 TI - [Benefits for national economy of mammography screening in Norway]. PMID- 10851927 TI - [Should research and health administration substitute parts of the basic anesthesiology education?]. PMID- 10851928 TI - [Optometrists want diagnostic eye drops]. PMID- 10851929 TI - [Norway no longer among the medical elite?]. PMID- 10851930 TI - [COX-2-inhibitors]. PMID- 10851931 TI - [Modern technology in the diagnosis of breast diseases--magnetic resonance tomography]. PMID- 10851932 TI - [Postpartum period in patient hotel]. PMID- 10851933 TI - [Vertigo]. PMID- 10851934 TI - [Patient information on tape]. AB - Giving patients access at home to tape recordings of their hospital discharge information was thought to improve the patient information. There could be an effect on their knowledge of their own disease and possibly also on clinical events. 50 patients with first time myocardial infarction were included; 26 received tape recordings and tape players. 24 patients were included as controls. The equipment was returned after one week, and the patients were followed up with a questionnaire after one, eight and 52 weeks. No difference in medical knowledge was observed. However, fewer patients in the tape group had long term sick leaves, and readmissions or need for emergency visits were less common. Baseline characteristics were almost the same in the two groups. Since acquired knowledge about their own disease did not appear to differ, an effect of the tape information upon the patient's family is considered. PMID- 10851935 TI - [Postnatal care--sleep, rest and satisfaction]. AB - BACKGROUND: During the last decade most hospitals in Norway have introduced breastfeeding according to the baby's needs during night-time as well. The aim of this study was to examine whether women had sufficient sleep and rest in the maternity unit, and the factors influencing insufficient sleep and rest. The degree of satisfaction with the stay in the maternity unit and factors associated with satisfaction were also studied. MATERIAL AND METHODS: From April to November 1998, 160 postnatal women in two Norwegian communities were included in a questionnaire-based study, representing 89% of all women eligible for the study. RESULTS: 47% (75/160) of the women reported lack of sleep and rest in the maternity unit. The factor most strongly associated with lack of sleep and rest was not having a single room (adjusted odds ratio 11.0; 95% confidence interval: 1.7-69.1). 56% (88/158) of the women reported to be very satisfied, 39% (68/158) were moderately satisfied, and 5% (8/158) dissatisfied with the stay at the maternity unit. Not being very satisfied was associated with the hospital of delivery (adjusted OR 18.0; 95% CI: 2.2-149.1), and with insufficient sleep and rest (OR 3.3: 1.3-8.1). INTERPRETATION: Our results suggest that women do not get sufficient sleep and rest under existing circumstances in maternity units. PMID- 10851936 TI - [Aftercare of newborn infants in a patient hotel]. AB - INTRODUCTION: Postnatal care of the healthy term newborn and the mother has, in modern times, taken place in the hospital setting. As a result of tightened hospital budgets as well as maternal preferences the duration of hospital stay has successively been shortened. Most women in Scandinavia today leave the hospital within four days after delivery. Postnatal care in a hotel like setting has emerged as an alternative to the well-baby nursery unless medical conditions makes this option inappropriate. MATERIAL AND METHODS: To evaluate the safety of postnatal care a study was undertaken to investigate whether correct criteria were being used for referral of the newborn to the hotel. We also wanted to document the duration of stay, unexpected medical complications, and weight development of the infants. Data from 865 infants were used for analysis. RESULTS: Ten (1.1%) newborns had to be readmitted to the hospital due to medical complications. 488 (56%) of the mothers went home within 96 hours, and only 23 (2.6%) stayed more than 120 hours. The weight of the infants reached a nadir on the fourth day post partum (-5.2% of birth weight). INTERPRETATION: The patient hotel is a medically safe alternative to the traditional well-baby nursery, provided that appropriate criteria for referral are used. PMID- 10851937 TI - [Argon plasma coagulation--a new method in therapeutic endoscopy]. AB - BACKGROUND: By argon plasma coagulation (APC), a current is applied to tissues as ionised gas. Special probes have recently been developed for applying the gas through flexible endoscopes. In the field of therapeutic endoscopy, this method is promising for several diseases in the gastrointestinal tract. MATERIAL AND METHODS: At Ostfold Hospital in Fredrikstad, Norway, 122 treatments in 80 patients were performed during the years 1997-99. RESULTS: The new method was useful for endoscopic treatment of haemorrhages, tumour debulking and tumour ingrowth and overgrowth in oesophageal stents. Abdominal pain was related to insufflation of air and gas. Complications related to the method were not observed. INTERPRETATION: Our experience with this new method was very positive. The method was effective, had a very low complication rate, and the equipment was easy to use. The application of APC in premalignant conditions is discussed. PMID- 10851938 TI - [Bacterial tracheitis in children]. AB - BACKGROUND: Bacterial tracheitis is an uncommon, but serious cause of acute respiratory distress in children. The incidence is not known. MATERIAL AND METHODS: The medical records of four children with bacterial tracheitis treated in our hospital are presented, and the literature reviewed to describe symptoms, diagnosis and treatment. A questionnaire was sent to all pediatric departments in Norway to assess the incidence of bacterial tracheitis and epiglotitis during the 1994-98 period. RESULTS: The yearly incidence of bacterial tracheitis was estimated to 4 per 1,000,000 for children aged 0-15, and 8 per 1,000,000 for children aged 0-5. The incidence of epiglotitis was 1.0 per 1,000,000 for children 0-15 years. INTERPRETATION: Bacterial tracheitis is now more common than epiglotitis, and the diagnosis has to be considered in children presenting with acute illness and upper airway respiratory distress. The disease is characterised by marked purulent exudate and formation of pseudomembranes in the trachea. Staphylococcus aureus and Haemophilus influenzae type b are the predominant causes of bacterial tracheitis. Most patients require endotracheal intubation, with the highest frequency in the youngest children. Reported complications include cardiopulmonary arrest, toxic shock syndrome and pulmonary oedema. Appropriate treatment with antibiotics is essential. PMID- 10851939 TI - [Acute porphyria]. PMID- 10851940 TI - [Pathophysiology and clinical manifestations in pre-eclampsia]. AB - BACKGROUND: Preeclampsia is a progressive, multisystem disorder characterised by hypertension and proteinuria. A body of evidence suggest a genetic basis; it is generally accepted that the underlying pathological processes are in the placenta. MATERIAL AND METHODS: This article is a review of the pathophysiology of preeclampsia based on literature mainly obtained through PubMed and Medline searches. RESULTS: A poorly perfused placenta, secondary to defective placental invasion of the spiral arteries, may lead to hypoxia and insufficient perfusion and cause release of cytokines which damage endothelial cells and cause dysfunction. Women with preeclampsia have markedly elevated concentrations of triglyceride-rich lipoproteins. Lipid peroxidation also causes endothelial dysfunction and thus contributes to preeclampsia. Placenta is one source of the lipid peroxides. Antioxidant deficiency is also a predisposing factor. Hyperhomocysteinaemia, protein S and protein C deficiency, and activated protein C resistance appear to be involved in the pathophysiology of severe preeclampsia and early onset preeclampsia. INTERPRETATION: The new information about mechanisms for development of preeclampsia gives a basis for new treatment modalities. PMID- 10851941 TI - [HELLP syndrome]. AB - BACKGROUND: The HELLP syndrome (H = hemolysis, EL = elevated liver enzymes, LP = low platelets) is a pregnancy complication which affects 10-20% of cases of severe preeclampsia. MATERIAL AND METHODS: The article is a review of the literature. RESULTS: Approximately 70% of HELLP syndrome cases occur before delivery, 15% as early as in the second trimester, the remainder after delivery. The classical HELLP syndrome is characterised by abdominal pain, pathological liver tests and low platelets. However, some cases are atypical; hypertension and abdominal pain may both be absent. Genetic as well as immunologic factors are involved in the pathogenesis. There is an imbalance in the coagulation process in the placenta. Activated leukocytes and macrophages induce production of cytokines that may reach the general circulation and cause endothelial dysfunction. In the HELLP syndrome fibrin deposits are also found in the vessels and in the liver sinusoides. INTERPRETATION: A mother with a classic HELLP syndrome should be delivered after stabilisation of the clinical condition. A partial HELLP syndrome can be observed. Treatment with corticosteroids is beneficial. PMID- 10851942 TI - [Pre-eclampsia--a review]. AB - BACKGROUND: Preeclampsia is characterized by hypertension and proteinuria with or without oedema. MATERIAL AND METHODS: The authors highlight some aspects of preeclampsia: epidemiology, classification, clinical evaluation and treatment. RESULTS: The condition may be classified as light or severe. Preeclampsia can induce damage to the placenta, liver, kidneys and brain, in addition to complications like the HELLP syndrome, placental abruption and eclampsia. Thrombocyte activation may cause activation of the coagulation system and thrombocytopenia. Early onset preeclampsia (< 34 weeks) is often associated with placental infarcts and reduced fetal growth. INTERPRETATION: We focus on early signs and close clinical surveillance. The diastolic blood pressure should be estimated with Korotkoffs' phase V. Patients with early onset preeclampsia should be hospitalized, as should women with hypertension and newly developed proteinuria. Antihypertensive treatment is discussed. Cases with reduced fetal growth and those with severe preeclampsia should in most cases be delivered preterm. Vaginal delivery is preferable. Labour may be induced by oxtocin, following cervical prostaglandin stimulation as indicated. In such cases cardiotocography surveillance during labour should be performed. Caesarean section may be performed in selected cases. Patients with mild preeclampsia can await spontaneous vaginal delivery at term, but delivery should be induced if they proceed past term. PMID- 10851943 TI - [Screening for ovarian cancer]. AB - BACKGROUND: Ovarian cancer is a common malignant neoplasm in affluent societies. Although the prognosis has continuously improved since the 1950s, it is still poor, with five-year relative survival rates of about 40%. MATERIAL AND METHODS: The paper discusses the potential use of screening for early ovarian cancer detection. RESULTS: The disease is a suitable candidate for screening because the prognosis is strongly dependent on the stage of disease at the time of diagnosis. Ovarian cancer does not, however, meet all the prerequisites for successful screening; there is no preclinical stage that can be detected, and the tests are not sensitive and specific enough to be effective. Furthermore, there is so far no direct evidence that screening would decrease mortality from ovarian cancer, though recent data suggest that screening may increase survival. INTERPRETATION: Screening women at higher risk of developing ovarian cancer might be beneficial. PMID- 10851944 TI - [Diagnosis of breast diseases with magnetic resonance tomography]. AB - Magnetic resonance imaging (MRI) of the breast is currently used for evaluation of both parenchymal disease and silicone gel implants. MRI is widely recognised as the most accurate imaging method for evaluation of breast implant integrity. Knowledge of the MRI appearance of the different types of implants and the possible complications of their use is important for diagnosis in these patients. The use of contrast-enhanced MRI in breast cancer diagnosis has been investigated thoroughly during the last decade. It is a sensitive imaging method to detect breast pathology, and may have an important diagnostic impact in carefully selected patient groups. The development of new MR techniques may improve the utility of MR in breast cancer diagnosis, treatment and research. PMID- 10851945 TI - [Use of magnetic resonance tomography in the diagnosis of gastrointestinal diseases]. AB - BACKGROUND: Magnetic resonance imaging (MRI) is a promising method for examination of the gastrointestinal tract. In this article we present the present status and potential of MRI. MATERIAL AND METHODS: The review is based on personal experience and selected published, international papers. RESULTS: Magnetic resonance cholangio-pancreatography (MRCP) and MRI of the liver are well documented examinations which are widely performed. These examinations partly replace existing modalities such as CT and ERCP and represent additional possibilities for examining this anatomic area. MRI is not as yet accepted as the standard examination of the pancreas, but it is regarded as comparable to CT. The adrenals may be examined even more accurately with MRI than with CT. MRI examinations of the oesophagus and gastric ventricle seem promising, but it should be said that they are in an early and not well documented phase. MRI of the small and large bowels is not common. With the exception of preoperative staging of rectal cancers, it is still experimental. MR angiography, functional MRI studies and perfusion studies are not yet in common abdominal diseases. We may, however, believe that they will become important diagnostic tools. INTERPRETATION: MRI is rapidly increasing its share of gastrointestinal imaging examinations. This is mainly due to the increased speed of newer machines. The diagnostic quality has improved, and will improve more. It is thus likely that MRI will, in a large amount of abdominal imaging, replace CT, and to some extent other diagnostic modalities. It is, however, not possible to replace CT for emergencies, and we will probably look forward to more and better imaging with more diagnostic modalities in the future. PMID- 10851946 TI - [Ketamine--a trendy doping agent in the mass media?]. PMID- 10851947 TI - [Optometrists and use of diagnostic agents]. PMID- 10851948 TI - [Can good routines in acute myocardial infarction be improved?]. PMID- 10851949 TI - [Is thrombolytic therapy efficient in stroke?]. PMID- 10851950 TI - [Post-traumatic carpal collapse--follow-up and therapeutic concept]. PMID- 10851951 TI - [Post-traumatic carpal collapse. Follow-up and therapeutic concept]. AB - Significant progress has been made in the understanding of carpal kinematics and posttraumatic disorders of the wrist. The importance of stabilization of the scaphoid is well known. More and more ligament injuries of the proximal carpal row have been diagnosed in cases of severe arthrotic changes. Long-standing scaphoid nounion or scapholunate ligament injuries can lead to progressive carpal collapse due to a break of the continuity of the proximal carpal row. SLAC-wrist (scapholunate advanced collapse) and SNAC-wrist (scaphoid nonunion advanced collapse) after missed fusion of scaphoid fractures should be differentiated. Severity of degenerative changes is classified into three stages. Salvage procedures preserving wrist mobility, like midcarpal fusion, are preferable to total wrist fusion because of the functional benefit. With complete excision of the scaphoid and fusion of the midcarpal joint, all arthritic joint surfaces are eliminated and motion is preserved in the radiolunate joint which is usually spared of degenerative changes. PMID- 10851952 TI - [Mechanism of pelvic girdle injuries in street traffic. Medical-technical accident analysis]. AB - During 1985 and 1993, 7,410 persons were injured in traffic accidents in the area of Hanover. Of these, 306 (4.1%) sustained a pelvic girdle injury. In 139 cases (45%), the pelvic girdle injuries were further classified (Pennal and Tile) and a technical reconstruction of the accident situation was performed. 52% were type A, 27% type B and 21% type C injuries. Some 47% of the casualties were vehicle occupants, 31% pedestrians, 12% motorcyclists and 10% cyclists. In restrained vehicle occupants pelvic girdle injuries occurred mostly in accidents with a delta-v of more than 30 km/h, whereas in unrestrained vehicle occupants, pedestrians and cyclists they also occurred with lower delta-v or collision speed. The percentage of type B and C injuries increased with higher velocities. In addition to further improvements in passive safety, lower collision speed or delta-v is necessary to reduce or prevent pelvic girdle injuries. The reconstruction of pelvic girdle injury mechanism in traffic accidents is possible, when both technical and medical parameters are considered. PMID- 10851953 TI - [Compartment syndrome of the thigh with sciatic nerve paralysis]. AB - Acute compartment syndrome of the thigh has been infrequently reported in the literature. Closed femoral fractures and blunt soft tissue trauma are the main causes of this injury. The multiple injured patient in this case report developed a compartment syndrome of the thigh after intramedullary nailing of a comminuted fracture of the femur. Fasciotomy was performed two days after surgery because of extense swelling of the thigh in the ventilated and sedated patient. Sciatic and femoral nerve palsy was recognized after extubation of the patient nine days after the injury. During the following weeks the paresis of the femoral nerve recovered but neither motor nor sensory function of the sciatic nerve could be demonstrated. Therefore an operative revision of the sciatic nerve was performed eighteen weeks after trauma. No direct nerve injury could be detected but there were adhesions around the nerve as a sign of compression neuropathy caused by the compartment syndrome. The tibial component of the sciatic nerve showed a complete recovery within the next months but there was a persisting peroneal nerve palsy. CONCLUSION: Early clinical symptoms of a compartment syndrome like pain, paresthesia and paresis can not be ascertained in a ventilated and sedated patient. Tense swelling of the muscles is often the only detectable sign. Frequent measurements of compartment pressure should be done in these patients. We suggest early decompressive fasciotomy because the morbidity caused by fasciotomy in a borderline compartment syndrome is far outweighed by the morbidity that accompanies an undiagnosed untreated compartment syndrome with possible nerve palsy. PMID- 10851954 TI - [Surgical vs. conservative treatment of fractures of the thoracolumbar transition]. AB - A total of 86 patients suffering from fractures of the thoracolumbar spine were followed up after an average time period of 57 months (12-98). Of these patients, 56 were treated operatively and 30 conservatively. According to the AO/ASIF classification, 66% of the operated group were fractures of type A, 29% of type B, and 5% of type C. All patients were operated on by means of dorsal locking instrumentation with pedicular fixation and, apart from six patients, with transpedicular cancellous bone grafting. The conservative group was treated according to the guidelines of Bohler with closed reduction, plaster cast, and rehabilitation program. All fractures in the conservative group were of type A. At follow-up of all operated cases, the local gibbus angle had improved by a reduction of on average 18.6 degrees and was followed by a loss of correction of 12.5 degrees ending in a final gain of 6.1 degrees at follow-up. At follow-up of the conservatively treated cases, the local gibbus angle showed an improvement of 11.1 degrees at reduction and a loss of correction of 14.9 degrees after reduction. The remaining result was -3.6 degrees, that means an increase of kyphoses compared to the x-ray at admission. In order to be able to compare two homogeneous groups only fractures of type A were used. Comparison of the two groups showed an improvement of the vertebral body angle of 70% (11.3 degrees) after reduction in the surgical group and 46% (6.1 degrees) in the conservatively treated group. The subsequent loss of correction was 19% (3 degrees) in the surgical and 34% (4.5 degrees) in the conservatively treated group. The remaining gain at follow-up was 51% (8.3 degrees) in the surgical and only 12% (1.6 degrees) in the conservative group. The local gibbus angle had improved on average by 17.1 degrees after reduction in the surgical and by 11.1 degrees in the conservatively treated group. Loss of correction was 71% (12.2 degrees) and 132% (14.9 degrees), respectively. The final result at follow-up showed a decrease of kyphosis of 4.9 degrees in the surgical and an increase of kyphosis of 3.7 degrees in the conservatively treated group. The difference was significant. Within the surgical group, 75% of the loss of correction was caused by the discs and 25% by the vertebral body. In the conservatively treated group it was 69% and 31%, respectively. Concerning loss of correction, no difference was seen between patients with and without intercorporal bone grafting. There was no relationship between radiological and clinical outcome. Whereas 15% of the patients of the surgical group were not satisfied or moderately satisfied with the result, all patients in the conservatively treated group were satisfied or very satisfied. Based on the good clinical results of the conservative treatment we can conclude that in stable fractures without severe deformity, and in patients who are in bad general condition, conservative treatment can considered as an alternative to surgical treatment. PMID- 10851955 TI - [Prognostically relevant factors in treatment of the post-traumatic unstable shoulder joint]. AB - The purpose of this prospective study was to determine concomittant lesions of glenohumeral dislocation and their correlation with the outcome, to determine the influence of various stabilizing operation techniques on the outcome. 106 shoulders [104 patients, 81 men, 23 women, mean age 36 (16-83) years] operated because of shoulder instability after at minimum one shoulder luxation between 1988 to 1995 were evaluated 3 years (0.8-8 years) postoperatively using Constant score. Preoperative arthroscopy was performed to document intraarticular lesions. There were 7 reluxations (5 because of a new accident), mostly after Putti-Platt procedures. The average modified Constant Score was 66 (10-85) points (maximum 85). 29 patients (27%) had less than 60 points, mostly because of pain and restriction of function, and were evaluated as poor. Lesions of rotator cuff, long biceps tendon and subscapularis tendon had a bad prognosis. The severity of arthroscopically found lesions correlated with the outcome. Operations with shortening of subscapularis tendon or using a bone graft gave bad results. Anatomic reconstruction of the labrum-ligament complex in combination with a plasty of the extended capsule gave best results. Preoperative arthroscopy can determine the extent of different lesions after shoulder luxation which influence the outcome and is useful to select the optimal operation technique. PMID- 10851956 TI - [Difficulties in evaluating follow-up outcome in calcaneus fracture managed with plate osteosynthesis. Is there a reliable score?]. AB - A huge amount of publications is existing dealing with topics of operative or nonoperative treatment, results and follow-up of fractures of the os calcis. Comparing the different studies one can see that there are a lot of different scores applicated for follow-up and outcome. Therefore it's difficult or impossible to compare the results of the different publications. The problems are discussed while evaluating own results of operative treated fractures of the os calcis. Between 1993 and 1996 we treated 27 fractures with ORIF (open reduction and internal fixation). 25 of the patients could be followed-up after a mean of 22.1 months. For follow-up three recently published scores, the AOFAS Ankle Hindfoot Scale (AOFAS), the Calcaneal Fractures Scoring System (CFSS) according to Kerr and the Functional Outcome Assessment-Score (FOA) according to Thordarson were used. The mean AOFAS-Score was 76.52 +/- 14.22, the mean CFSS was 77.40 +/- 15.22 and the mean FOA was 70.08 +/- 18.53 points. The differences between the AOFAS and the FOA-Score were significant (p = 0.017). The differences between the CFSS and the FOA-Score were significant (p = 0.004), too. These different values elucidate the need of uniform structured follow-ups of fractures of the os calcis. To compare different national or international studies uniform scores should be used. In anglo-american literature the AOFAS Ankle-Hindfoot Scale is well established. This scoring-system proved to be quite useful in our own analysis. We suggest that the AOFAS Ankle-Hindfoot Scale also should be taken into account in german literature, especially in follow-up of calcaneal fractures. PMID- 10851957 TI - [Fractures of the facial skull]. PMID- 10851958 TI - [Intrathoracic displacement of the transverse colon as a late complication after abdominal knife stab wound. A case report]. AB - The treatment of stab wounds of the abdomen is discussed controversely. The management of these wounds depends on a high degree on the clinical situation of the patient. Repeated physical examination, peritoneal lavage and laparotomy are recommended. The identification of an isolated diaphragmatic injury is worrisome because its subtle clinical presentation is often not alleviated by adjunctive tests. Delayed recognition of an incarcerated diaphragmatic hernia has a high mortality rate. We report about an 23 year old African male, who was injured by an intraabdominal stab wound. Initially he did not have any clinical problems, but after 6 months he was admitted to the hospital due to severe pulmonary complications. A diaphragmatic hernia with dislocation of the transverse colon into the left thorax was identified on x-rays. A thoracotomy as well as a laparotomy were performed to restore the anatomic conditions and to dose the hernia. After two weeks, the patient could leave the hospital without pulmonary complications. Under consideration of the literature we recommend exploratory laparotomy for stab wounds that penetrate the left side of the chest below the fourth intercostal space anteriorly, the sixth intercostal space laterally and the tip of the scapula posteriorly. The high mortality rate after a delayed recognition appears to justify an early operative approach. PMID- 10851959 TI - [Inferior epigastric artery. An atypical source of bleeding in pelvic fracture]. AB - The most common mechanism of pelvic fractures after blunt trauma is lateral compression of the pelvis. Most of these fractures are of slight severity but it is possible, that a life-threatening hemorrhage can appear. The inferior epigastric artery is an atypical bleeding site but it has to be considered in the search of the origin of the hemorrhage. Diagnostic tools are ultrasound, computed tomography and angiography. After external fixation of the pelvis and persistent haemodynamic active bleeding is the percutaneous transcatheter embolization (PTE) in our opinion the treatment of choice. The introduced case is confirm with this statement. PMID- 10851960 TI - [Tuberculous spondylitis of the cervical spine]. AB - WHO estimates that worldwide more than 4 million people are infected with tuberculosis (TB), 95% of them living in third world countries. TB is once again the most frequent infectious disease [1, 2, 7]. Extrapulmonary forms (EPTB) frequently appear with HIV-associated TB [3, 4, 6]. We present the case of a 23 year old patient with abscess forming TB of the right apical lung with infiltration and partial osteolytical destruction of the 7th cervical- and the first thoracal vertebral body. Progressive spinal compression lead to partial sensomotorical deficits. Surgical revision of the abscess, an atypical resection of the right apical lung and right wing hemilaminectomy of Th1/2 for spinal decompression became necessary. The spinal stabilisation was achieved by conservative treatment for 6 weeks only by a Minerva cast and for another 8 weeks by a cervical stiffneck. Following mobilisation was without problems and the neurological deficits subsided within 3 months. Clinical examination and functional X-rays analysis proved vertebral stability and good function. The treatment of TB is basically conservative. Surgery is only recommended in case of functional and infectious complications. PMID- 10851961 TI - [Pigmented villonodular synovitis. A rare differential diagnosis of popliteal space-occupying lesion]. AB - Pigmented villonodular synovitis (PVNS) is a disease that involves the lining of joints, bursae and tendon sheaths. The incidence is low and estimated to be 1.8 patients per million population. The cause of PVNS is unclear and discussed to be either inflammatory or neoplastic. PVNS has been described in 2 forms different for prognosis and treatment (nodular and diffuse). The articular form almost appears in the knee joint as we describe below in a 14 year-old patient. PVNS was first defined in 1941 by Jaffe e.a. [7]. Because of the uncommon occurrence of the disease it is difficult to amass patient series to allow confirmed statements on therapy and outcome. Larger patient series raise out of long periods of time. Differences in outcome and recurrence rates exist for the nodular and diffuse form. Clinical findings are moderate pain and swelling of joints due to effusion and synovial proliferation. Magnetic resonance imaging shows typical findings. Surgical procedures are recommended as open or arthroscopic synovectomy for the diffuse form of PVNS, local excision for the nodular form and arthrodesis or prosthetic replacement for joint destruction. PMID- 10851962 TI - [Industry sponsorship in the hospital. Legal stumbling blocks for the physician]. PMID- 10851963 TI - [True medicine? Economic analysis and perspectives for medical service]. PMID- 10851964 TI - A "natural" approach to treating osteoarthritis. PMID- 10851965 TI - Aspirin to prevent strokes: fact or fiction? PMID- 10851966 TI - Drug recalls underscore safety concerns. PMID- 10851967 TI - Room for improvement in stroke treatment. PMID- 10851968 TI - Antioxidant overhaul. PMID- 10851969 TI - Problematic painkillers. PMID- 10851970 TI - Antioxidant supplements and dementia. PMID- 10851971 TI - PSA "bounce" need not cause alarm. PMID- 10851972 TI - HRT and your heart. PMID- 10851974 TI - Diet and cataracts. PMID- 10851973 TI - Lower your LDL with nuts? PMID- 10851975 TI - Olive oil lowers blood pressure. PMID- 10851976 TI - Bottled water vs tap: which is better? PMID- 10851977 TI - Is there a difference between GERD and chronic heartburn? How should they be treated? PMID- 10851978 TI - What causes Bell's palsy? I have had two episodes and am concerned about another recurrence. PMID- 10851979 TI - [The seroresistance of enterobacteria isolated from different sources]. AB - 395 representatives of the family Enterobacteriaceae, isolated from household and drinking water, from healthy and sick persons, were used as an example to demonstrate that their seroresistance was greatly connected with the type of the source from which these bacteria had been isolated. Experiments showed the phenotypic conversion of Escherichia coli towards an increase in their seroresistance on contact with human blood serum. The study revealed the pleiotropic effect produced by the action of serum on a number of other properties, responsible for survival in a macroorganism (anticomplement and antilysozyme activities, adhesive capacity and hydrophobic properties of the outer surface). PMID- 10851980 TI - [The development of a process for culturing Bordetella pertussis immobilized on a polyurethane carrier]. AB - The processes of the cultivation of Bordetella pertussis, immobilized on polyurethane carrier in a fermenter, were carried out and studied. Acellular pertussis preparations were produced from the culture fluid obtained in the batch and multi-cycle cultivation processes with immobilized cells, as well as in the process with interrupted fermentation (for confirming the possibility of the preservation of cell viability). The content of protein and B. pertussis toxin in these preparations, as well as their leukocytes-stimulating and hemagglutinating activity, did not differ from similar characteristics of preparations obtained from culture fluid in homogeneous cultivation. PMID- 10851981 TI - [The comparative characteristics of the pathogenic action of Vibrio cholerae of the non-O1 group O139 serovar and of the O1 group on the intestinal APUD system of suckling rabbits]. AB - The comparative study of the enteropathogenic action of V. cholerae strains of group non-O1, serovar O139, and group O1 with different virulence on the APUD system of the intestine of suckling rabbits after intraenteral infection revealed that V. cholerae of group non-O1 induced inflammatory changes in the intestine and the pronounced toxic lesion of parenchymal organs. This was accompanied by a decrease in the number of apudocytes and an increase in the functional tension of the APUD system. After the infection of the animals with V. cholerae of group O1 changes in the APUD system and internal organs directly depended on the virulence of the microbes and the infective dose. PMID- 10851982 TI - [The characteristics of the action of the capsular antigen of Yersinia pestis on cells from plague-susceptible and -insusceptible rodents]. AB - The state of the functional activity of peritoneal leukocytes from different species of gerbils (Mongolian gerbils, Libyan jirds, as well as gerbils inhabiting the right and left banks of the Volga), exposed to the action of Y. pestis capsular antigen, has been found to correlate with the susceptibility of these animals to plague. PMID- 10851983 TI - [The effect of the cultivation conditions on the growth and pigmentation of Serratia marcescens]. AB - The influence of cultivation conditions on the growth and pigmentation of S. marcescens was studied. The cultures under study grew in media containing glycerin, glucose or acetate and organic or mineral nitrogen. Pigment formation occurred in a medium with organic nitrogen and glycerin or acetate, but not glucose. Sodium chloride inhibited the growth of cultures, but at a concentration not exceeding 4-5% increased the accumulation of prodigiosin. Prodigiosin was accumulation by the culture growing at 28 degrees C, while at 37 degrees C no accumulation of the pigment occurred. The illumination of the growing culture with visible light decreased the intensity of its pigmentation. Prodigiosin apparently plays an important role in the metabolism of S. marcescens and is linked with the energy exchange of the cell. PMID- 10851984 TI - [The use of computer technologies in the operational analysis of outbreaks of hospital infections]. AB - The description of the mathematical model for the study of outbreaks of hospital infections caused by Klebsiella pneumoniae in intensive care units and the example of its use are presented. Calculations in connection with the evaluation and prognostication of the number of sick and deceased children on account of Klebsiella hospital infection were carried out with the use of the proposed computer program. The importance creating an effective hospital information and analytical system, suitable for the treatment and analysis of data on the epidemiological monitoring of hospital infections, with the elements permitting the registration and prognostication of the cases of the appearance and development of hospital infections, the evaluation of the morbidity level, as well as the effectiveness of using different prophylactic measures. PMID- 10851985 TI - [The spread of rotavirus gastroenteritis in the Republic of Tajikistan]. AB - The article deals with the results of the investigation which has demonstrated the spread of rotavirus infection on the territory of the Republic of Tajikistan. This infection has gained an important place in the regional pathology and requires increased epidemiological surveillance on its spread. As established in the course of this investigation, in patients with the clinical manifestations of pronounced gastroenteritis rotavirus plays the role of the etiological agent in 25.8% of cases in children aged up to 14 years and in 11.0% of cases in adults. Synchronism in the seasonal dynamics of morbidity in acute enteric infections of unclear etiology and rotavirus gastroenteritis among children aged 0-2 years, annually reaching morbidity peak in October, has been revealed. Rotavirus gastroenteritis requires close attention as regards its prophylaxis as one of the main nosological forms of acute enteric infections in the Republic. PMID- 10851986 TI - [Hepatitis C antibodies in groups at epidemic risk]. AB - Blood sera from patients with acute virus hepatitis, sexually transmitted diseases, psychiatric and oncological diseases and pulmonary tuberculosis were studied for the presence of antibodies to hepatitis C virus (anti-HCV) and HBsAg with the use of solid-phase enzyme immunoassay. Anti-HCV were detected, respectively, in 9.1% and 67.8% of patients with acute virus hepatitis, in 5.7% and 6.1% of patients with sexually transmitted diseases, in 7.5% and 9.2% of psychiatric patients, in 7.5% and 13% with pulmonary tuberculosis and malignant tumors. PMID- 10851987 TI - [The work of the State Epidemiological Health Service of Russia on the prevention of the most wide-spread diseases in 1996-1997]. PMID- 10851988 TI - [The effectiveness of using the Vaxigrip vaccine from the firm of Pasteur Merieux Connaught for influenza prevention in organized groups of adults]. AB - The results of the study of the effectiveness of using vaccine Vaxigrip for the prophylaxis of influenza in organized groups of adults are presented. The vaccine was found to have high epidemiological effectiveness (the epidemiological index was 2.6), moderate reactogenicity and pronounced immunological activity (the protection level was 89.0-100.0%). The vaccine may be recommended for the prophylaxis of influenza among adults. PMID- 10851989 TI - [The antidiphtheria activity of the pre-EGF fragment]. AB - The DNA fragment, coding a part of the protein molecule--the precursor of the epidermal growth factor (pre-EGF76-208)--and containing the sequence of 133 N-end amino acid residues, was obtained with the use of gene engineering and molecular biological techniques. For this purpose a fraction of poly(A+) = RNA was isolated from the kidney of a newborn infant; on this fraction the "library" of cDNA fragments whose coding capacity corresponded to the required sequence of mRNA in the pre-EGF gene was obtained in the reaction of reverse transcription, conjugated with the polymerase chain reaction (PCR). After the determination of the nucleotide sequences in 11 fragments only 1 fragment which contained practically no mutations appearing in PCR as the result of amplifications was chosen. This fragment was used for the construction of a hybrid plasmid controlling the synthesis of hybrid fusion protein with the sequence of pre-EGF76 208. Fusion protein was synthesized with very low effectiveness, but the use of metallo-affinity chromatography permitted to isolate and purify it, its purity reaching 98%. Then its antitoxic activity was determined in the skin test on guinea pigs and found to be not less then 10(6) I.U./mg of protein. PMID- 10851990 TI - [The bactericidal action of human neutrophils on meningococci in vitro]. AB - 32 Russian patients with late complement component deficiency (LCCD) were immunize with tetravalent meningococcal polysaccharide vaccine (A + C + W135 + Y). Their immune response and infectious morbidity rate were followed for 6 years and the partial protective efficacy of vaccination was demonstrated. As the antibody-mediated complement-induced bactericidal activity of plasma was completely absent in persons with LCCD, the bactericidal action of human neutrophils on meningococci of groups A, W135 and B was studied under the conditions of incubation with serum samples collected from persons with LCCD before and after vaccination. In LCCD serum alone the exponential growth of meningococci was observed, while the addition of human neutrophils resulted in the essential inhibition of the growth of meningococci (up to their complete elimination). The proportion of serum samples stimulating the elimination of group A and W 135 meningococci by neutrophils was almost 40% of the serum samples collected before vaccination and significantly increased among the serum samples collected after vaccination (up to 84%) or revaccination (up to 90%). At the same time the capacity of an individual serum sample to promote the bactericidal effect of neutrophils against meningococci correlated with its content of specific anti-polysaccharide IgG and IgM antibodies, as well as antibodies to the inner core of lipopolysaccharide. The interaction of neutrophils with meningococci was significantly inhibited after incubation in heat-inactivated serum, suggesting that this interaction was partly mediated along the following path: the binding of IgM and IgG antibodies with bacteria--the activation of complement and the deposition of C3 complement on bacteria--the binding of meningococci with CR3 receptors of neutrophils. PMID- 10851991 TI - [The outlook for the development of live vaccines for the prevention of melioidosis]. AB - The effectiveness of immunization with Burkholderia pseudomallei attenuated strains (Pur and Ts), heterologous vaccines and the recombinant culture of Francisella tularensis RM2 carrying a plasmid with fragments of B. pseudomallei chromosome was studied in four species of experimental animals, essentially differing in their sensitivity to melioidosis. The most immunogenic B. pseudomallei mutants, introduced subcutaneously, created a statistically significant level of protection in animals, moderately sensitive to melioidosis, but proved to be ineffective in highly sensitive animal models when tested under the same conditions. In aerogenic infection the effectiveness of the experimental vaccines under study in all species of the animals was on the same level. The study showed good prospects of using tularemia vaccine for inducing heterologous immunity to melioidosis, as well as the possibility of its use as the basis of a bivalent gene-engineering vaccine. PMID- 10851992 TI - [Liposome-incorporated interferon. The design of liposomal interferon]. AB - To develop bound interferon, purified phospholipids (PL), obtained from waste products of soy-bean oil production, were chosen as lipid carries. The method for the development of the liposomal interferon complex was proposed. As shown in this study, with regard to the composition of the most significant PL fractions the carrier obtained from waste products of soy-bean processing proved to be more acceptable for humans than egg-yolk PL, traditionally used in practical liposomology. The advantage of the purified concentrate of waste products of soy bean oil production was its greater affinity with the PL fraction profile of human internal organs (the liver, the kidneys) than the composition of PL of animal origin. The dispersion and stability of interferon-containing liposomes was evaluated. The interferon incorporation coefficient of soy-bean PL proved to exceed that of egg-yolk PL. PMID- 10851993 TI - [The nature of the respiratory tract microflora and the immunity indices in a severe form of bronchial asthma in children]. AB - 65 children with the severe course of bronchial asthma (BA) were examined. The character of respiratory tract microflora, humoral immunity characteristics, mediators, of inflammation and pathomorphological features of the bronchial tree during the severe course of BA in children at the period of remission were studied. The specific features of the colonization of the bronchial tree by microflora in the severe form of BA were established. The severe course of BA in children at the period of remission was characterized by a higher content of tumor necrosis factor and an increase in the lysozyme activity of bronchoalveolar fluid. Disturbances in the proportion of cAMP and cGMP in smooth muscles and an increase in the number of apudocytes containing adrenalin and serotonin may probably be one of the mechanisms of bronchial hyperreactivity. PMID- 10851994 TI - [The complement system and circulating immune complexes in burn patients]. AB - The results of the study of humoral immunity in 80 burn patients are presented. A typical feature of all burn patients was a decrease in the total activity of the alternative path of the activation of complement and its factors B and D, beginning from the first day after the trauma. The character of changes in the functional activity of components C1-C5 of the classical path depended on the area of damages with their activation if burn area was less than 20% of the skin surface and deactivation if the burn area exceeded 20%. PMID- 10851995 TI - [The intensity of cell receptor catabolism in intestinal yersiniosis and pseudotuberculosis]. AB - The intensity of the catabolism of cell receptors in yersiniosis was evaluated by the level of R-proteins in the blood sera of 95 patients. The study revealed that the degree of an increase in the level of R-protein depended on the severity and the course of the pathological process was the same in both nosological forms of the disease. The high and stable level of R-proteins was indicative of the possibility of the prolonged and relapsing course of yersiniosis. PMID- 10851996 TI - [Bacterial endotoxinemia in children with intestinal dysbacteriosis]. AB - 34 children with gastrointestinal diseases of infectious, allergic and mixed etiology were examined. The state of normal microflora in the large intestine as indicated by fecal bacterial charts and the level of secretory immunoglobulin A (sIgA) in the contents of the intestine as indicated by the results of radial immunodiffusion were studied. In addition, the content of endotoxin in the children's plasma was determined with the use of the Limulus (LAL) test. The presence of endotoxin in the plasma of children with intestinal dysbiosis was determined in 71.1% of cases. The frequency of the detection of antigenemia was found to be related to the severity of manifestations of dysbiotic changes in the intestine and to the level of sIgA in fecal supernatants. The inclusion of the probiotic preparation Bifidumbacterin forte containing live bifidobacteria adsorbed on activated charcoal into the complex therapy of digestive tract diseases ensured a decrease in the detection rate of endotoxinemia, which correlated with the tendency towards the normalization of defective intestinal microflora. PMID- 10851997 TI - [The characteristics of detecting the tick-borne encephalitis virus antigen in the ELISA and indirect hemagglutination reaction by means of scanning electron microscopy]. AB - The surface of polystyrene plates was studied at different stages of the enzyme immunoassay (EIA) and the passive hemagglutination (PHA) test by the method of scanning electron microscopy in the detection of tick-borne encephalitis (TBE) virus antigen. The study revealed that in the process of EIA larger antigens were washed away from the plate surface. The objects detected on the polystyrene surface were identified as conglomerations of the virions of TBE virus, but whole virions were shown to play no decisive role in EIA. The conclusion was made that, due to some specific features of this method, EIA was more sensitive in reaction with small antigens (individual glycoproteids, their small complexes). And, respectively, the PHA test was more sensitive in reaction with large antigenic complexes (whole virions, their conglomerations, immune complexes). PMID- 10851998 TI - [The rapid identification of bacteria by agglutination on a slide of antibody sensitized erythrocytes]. AB - The method for the identification of Brucella (genus) and shigellae (species) by using slide agglutination of antibody erythrocytic immune reagents is substantiated. Such reagents, obtained from polyclonal nonadsorbed immune sera, ensure high specificity of taxon identification. The use of the proposed method may greatly accelerate bacteriological analysis. PMID- 10851999 TI - [The intrastrain heterogeneity of the causative agent of anthrax]. PMID- 10852000 TI - [The etiological structure of leptospirosis in agricultural animals in Colombia]. PMID- 10852001 TI - [The sensitivity of clinical isolates of Helicobacter pylori and associated microflora to antimicrobial and anti-ulcer preparations]. PMID- 10852002 TI - [The antibiotic sensitivity of the causative agents of suppurative-inflammatory diseases in the clinics of the city of Kharkiv in 1990-1997]. PMID- 10852003 TI - [Drug-resistant strains of Mycobacterium tuberculosis and their epidemic significance]. PMID- 10852004 TI - [The hemo-cytolysin of Vibrio cholerae: its mechanisms of action in vitro and in vivo]. PMID- 10852005 TI - [The ecology of Salmonella typhi and its relation to the manifestations of typhoid infection]. PMID- 10852006 TI - [Superantigens and their role in pathology]. PMID- 10852007 TI - [The immunostimulating activity of muramyl dipeptide and its derivatives]. PMID- 10852008 TI - [The results and outlook for vaccinal prophylaxis in the Republic of Bashkortostan]. PMID- 10852009 TI - [Why is additional gene testing of donor blood and its preparations, as well as of the blood of newborns from HIV-infected mothers using PCR necessary]. PMID- 10852010 TI - [In Process Citation] PMID- 10852011 TI - [A lingering outbreak of viral hepatitis A in a children's collective]. PMID- 10852012 TI - [A case report: a small outbreak of Flexner dysentery in a kindergarten]. PMID- 10852013 TI - [The organization of the instruction of foreign students in a department of microbiology]. PMID- 10852014 TI - [Vaccinal prophylaxis. The results of the 20th century and the outlook for the coming one]. PMID- 10852015 TI - [Viral hepatitis morbidity in Russia and the measures for its decrease]. PMID- 10852016 TI - [The real place of infectious pathology in overall population morbidity]. AB - The statistical decrease of the proportion of infections in the structure of morbidity of the population reflects the existing classification of diseases when only acute diseases are classified with the group "infectious and parasitic diseases". The proportion of diseases caused by infective agents remains constantly high. According to WHO data, such diseases make up one-third of all diseases in the world. In Moscow the proportion of infectious diseases in all diseases registered among the inhabitants of this big city fluctuated within 36.1% and 49.7% during the period of 1926-1997. PMID- 10852017 TI - [The current trends in the development of new immunobiological preparations]. AB - In this article information on the main immunobiological preparations, including vaccines and toxoids, phages, eubiotics, preparations obtained on the basis of antibodies, immunomodulators, new adaptogens of different origin, is presented. The problem of the development of new diagnostic preparations and biosensors whose action is based on specific immunological reactions is discussed. The expected use of more effective, more natural, as well as free from many drawbacks, modern immunobiological preparations in the medical practice of the XXI century is emphasized. PMID- 10852018 TI - [The current status and outlook for molecular genetic methods in solving the tasks of medical microbiology]. AB - The article deals with modern methods, viz. PCR, molecular display and genotherapy, which permit the new approach to the solution of problems connected with the identification of infective agents, the study of the mechanisms of the pathogenesis of infectious diseases and their treatment. In this article concrete examples, clearly demonstrating how each of the above-mentioned technologies makes it possible to broaden the circle of problems solved in infectious pathology of man, are presented. PMID- 10852019 TI - [Natural-focus infections: the key questions and new vantage points]. AB - A short history of the concept of natural focal infections is presented: the idea put forward by D. K. Zabolotnyi, E. I. Pavlovskii's teaching, 3 stages of its development. A number of fundamental questions and the modern content of the concept are considered. The natural foci of infections are a combination of surface, soil and/or water ecosystems, including the population of the causative agent of infection. In contrast to surface ecosystems, in soil and water ecosystems the hosts of the causative agents of sapronotic infections ae soil invertebrates and hydrobios, in which these agents may circulate in biocenotic trophic chains. The circulation of the causative agents in natural foci is a discrete process; the mechanisms and forms of the existence of pathogenic bacteria during seasonal and prolonged periods between epidemics is considered. Special attention is given to latent (nonculturable) forms of bacteria. The complex character of the status of the causative agents of natural focal infections is discussed. PMID- 10852020 TI - [The factors of bacterial pathogenicity and their role in the development of the infectious process]. AB - The latest data concerning the characterization of the pathogenicity factors of bacteria and the evaluation of their role in the realization of definite phases of the development of the infectious process are presented. The infectious process is regarded as the result of the complicated simultaneous interaction of microorganisms and different cells and tissues of the host body. The problems of the polydeterminant character of pathogenicity factors, tho possibility of the joint action of different factors at one and the same stage of the development of the infectious process and, vice versa, the action of the same factors at different stages of the interaction of the infective agent and the susceptible host are discussed. Modern data on the genetic control of pathogenicity factors, on the localization of their genetic determinants on the chromosome and the virulence plasmids, information of pathogenicity "islets" which jointly determine the pathogenic potential of the infective agent are given. The emphasis is made on fact that the general principle of the genetic control of bacterial pathogenicity is complicated relationship between chromosomal and nonchromosomal determinants; some of them form a part of genetic pathogenicity "islets", simultaneously regulating and expressing the pathogenicity factors of the infective agent. PMID- 10852021 TI - [Supertoxic complexes of botulinum toxins]. AB - The super-toxic super-complexes (SSC) of botulinic neurotoxins, types A and B, have been isolated. The preparation of type A SSC (SSC/A) consist of the proteolyzed form of type A botulinic neurotoxin (BoNT/A), 50 and 90 kD, nontoxic nonhemagglutinating protein of 140 kD (NTNH140) in the nonproteolyzed form, hemagglutinin of 17 kD (Ha17), hemagglutinin of 34 kD (Ha34) and the proteolyzed form of hemagglutinin of 70 kD (Ha70) (20 and 50 kD). The preparations of type B SSC (SSC/B) consist of the nonproteolyzed form of type B botulinic neurotoxin (BoNT/B) of 150 kD, the proteolyzed form of BoNT/B of 150 kD (45 and 105 kD), the nonproteolyzed form of NTNH140, Ha17, Ha34 and two nonidentified proteins (32 and 40 kD). As shown in this study, toxic complexes both in native toxins and in the preparations of SSC do not dissociate for several weeks at pH 8.0 and for 18 hours in 3% SDS, as well as after treatment with RNAase or 1 M NaCl. Some part of SSC/A (neurotoxin and Ha70) has been found to dissociate in 3% SDS after 1-hour incubation at 22 degrees C after the addition of 2-ME. The preparations of SSC contain nucleic acids (A260 nm/A280 nm = 2.0), supposedly ensuring the stability of the complexes. In contrast to the L-forms of Clostridium botulinum toxins, the preparations of SSC/A and SSC/B have been found to possess increased toxicity. The specific toxicity of SSC/A has proved to be 1-2 x 10(9) DLM per 1 OD280 nm and that of SSC/B, from 5 x 10(8) to 1 x 10(9) DLM per OD280 nm. One minimal lethal oral dose of these SSC preparations for mice was less than 10 DLM, introduces intraperitoneally. PMID- 10852022 TI - [The apoptogenic mechanisms for the occurrence of immunodeficient diseases]. AB - Suggestion has been made that the development of immunodeficient states should be considered as the result of the acceleration of the apoptosis of immune cells. The pathogenetic basis of the apoptogenic mechanism of the development of immunodeficient morbidity is considered. PMID- 10852023 TI - [The capsular polysaccharide of serogroup-B meningococci: its immunobiological properties]. PMID- 10852024 TI - [Bifidogenic factors as drug preparations]. AB - The review of new data on the study of bifidobacterial factors of different origin and the probable mechanisms of their favorable action on the microflora of the intestinal tract if presented. The main emphasis is made on the analysis of data on the use of oligosaccharides, including fructo-oligosaccharides, as compounds stimulating the growth and development of bifidobacteria both in pure cultures and in intestinal microflora. Methods for the treatment of natural compounds with a view to enhancing their bifidogenic effect are presented. The possibilities and/or advantages of using bifidogenic factors in vivo and in vitro as medicinal preparations either alone or incorporated in probiotic compositions are evaluated. Suggestion has been made that the choice of the method for using bifidogenic factors may depend on the kind and severity of disturbances in indigenous microflora. PMID- 10852025 TI - [The antimicrobial activity of nitric oxide and its role in the infectious process]. AB - In this article information on an important role of nitric oxide (NO) in inhibiting the growth of a number of pathogenic microorganisms, including intracellular parasites, and their elimination from the host body is presented. Differences between the mechanisms of the production of NO and free-radical compounds having antimicrobial action are given. The regulation of the activity of constitutive NO-synthase and inducible NO-synthase and the relationship between the latter and the phagocytic activity and production of anti inflammatory cytokines are described. An important role of NO in the development of the nonspecific resistance of the body is mentioned. PMID- 10852026 TI - [The theory of the etiological selectivity of the main (primary) routes of transmission and their inequality in different nosological forms of intestinal infections]. AB - The article deals with the main scientific propositions of an original epidemiological theory, viz. the theory of the etiological selectivity of the main (primary) routes of transmission and their differences in various nosological forms of enteric infections (the theory of conformity). As proved on the basis of scientific investigations carried out for the period of several years, the role of various routes forming the fecal-oral mechanism of transmission (alimentary, aqueous, household) is highly different. In some enteric infections one of these routes becomes the primary (main) route, while other routes are secondary (additional). The primary route determines the wide spread of the corresponding nosological forms of enteric infections (the theory of conformity) and the preservation of their causative agents in nature as species. The secondary routes are only complementary to the activity of the main route of transmission in some degree. In controlling enteric infections, the measures taken with a view to suppressing the activity of the primary route of transmission are, from the epidemiological standpoint in the close and remote prospects, the cardinal ones, while the measures directed against the secondary routes are only palliative. PMID- 10852027 TI - [The antagonistic activity of bifidobacteria in vitro and in vivo studied by using gnotobiological technology]. AB - The antagonistic activity of 4 strains of bifidobacteria (B. adolescentis 2 F1, B. longum Z4, B. breve R2 and B. bifidum G1), isolated from the vagina of healthy females of the reproductive age, with respect to Escherichia coli, Klebsiella ozaenae, Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa and Gardnerella vaginalis were studied in vitro and in vivo. The in vitro experiments revealed that all above-mentioned bifidobacteria were capable of inhibiting the growth of all indicator bacterial strains. Still of all the bifidobacteria under study had different levels of activity. B. adolescentis strain 2 F1 exhibited the highest inhibiting activity in vitro. In contrast to in vitro experiments, in vivo experiments with B. breve R2 demonstrated its high antagonistic activity with respect to E. coli. The data thus obtained indicate that in the study of antagonistic activity the use of the in vivo model as also expedient, for it is mainly in vivo that probiotic preparations show their activity. PMID- 10852028 TI - [The use of batumin-containing disks for the rapid identification of staphylococci]. AB - The study of 467 microbial strains obtained from collections and from clinical sources revealed that microorganisms of the genus Staphylococcus were highly sensitive to batumin, a new antibiotic obtained from bacteria of the genus Pseudomonas. 378 strains of 15 Staphylococcus species proved to be highly sensitive to the diagnostic preparation "Diastaph", developed on the basis of batumin (antibiotic-impregnated discs); After 18-hour incubation the diameter of the growth inhibition zones on agar-containing culture media was 18-38 mm. Strains belonging to the genera Micrococcus, Dermacoccus, Kocuria and Kytococcus, as well as the tested representatives of other taxa (Planococcus, Streptococcus, Corynebacterium, Acinetobacter, Pseudomonas, Neisseria, the representatives of all tested genera of the family Enterobacteriaceae, fungi of the genus Candida) were insensitive to the diagnosticum. "Diastaph" permits not only the rapid identification of staphylococci pure cultures, but also the determination of their presence in association with other microbial species directly in pathological material, which makes it possible to recommend this diagnostic preparation for use in medical, veterinary and sanitary microbiology. PMID- 10852029 TI - [The genetic control of the F1-specific components of the capsular antigen in Yersinia pestis]. AB - The preparation of Y. pestis capsular antigen F1, isolated from Y. pestis Fra positive strain by Baker's method, has been shown to have a composition and contain three components: protein, glycoprotein and a lipid-containing component. Each of them equally reacts with antibodies to F1 and diagnostic preparations based on these antibodies. The synthesis of protein and glycoprotein is temperature-dependent and controlled by pYT (Caf). The synthesis of glycoprotein is constitutive and determined by chromosomal genes. Protein and glycoprotein have almost identical mol. wt., 18 and 17 kD respectively, but their localization is different: protein is secreted on the surface of bacterial cells and into the environment, while glycoprotein can be found inside the cells and is similar to intracellular glycoprotein of different enterobacteria, described in our earlier works and exhibiting F1 specificity. Different biological role of these F1 specific components is suggested. PMID- 10852030 TI - [The status of the morbidity and the main trends in the improvement of the epidemiological surveillance of sexually transmitted infections in the city of Moscow]. AB - The present situation in sexually transmitted diseases (STD) is regarded as unfavorable. Differing tendencies in morbidity levels for various nosological forms have been established. The continuing growth of syphilis morbidity among children is considered to be specially alarming, and an increase in the number of cases of congenital syphilis is noted. The reasons of the growth of STD morbidity have not only medical, but mainly socio-economical, psychological, ethical and moral character. The main trends in the improvement of epidemiological surveillance on STD have been determined. PMID- 10852031 TI - [The etiological structure of the morbidity from influenza and other ARDs on the territory of Russia in the season of 1997-1998]. AB - The antigenic properties of 51 strains of influenza virus A(H1N1), isolated in different cities of Russia during the epidemic of 1998, were studied. Most of these strains (49) proved to be similar to virus A/Bern/07/95 in the antigenic structure of hemagglutinin, but 2 strains isolated in Ulan-Ude were found to be closely related to new antigenic variants of this virus: A/Beijing/262/95 and A/Fukuoka/c7/98. The analysis of the antigenic structure of influenza-like diseases (ILD) in different cities of Russia revealed that adenoviruses causing up to 10.9-14.6% of all acute respiratory virus infections dominated at the pre- and post-epidemic periods. RS-viruses, parainfluenza viruses of types 2 and 3 circulated during the whole season (their proportion was 5.1-6.6%). The intensity of the circulation of influenza viruses A(H1N1) and A(H3N2) increased, starting from January, and continued till April 1998; its peak was observed in February March in most of the cities of Russia (up to 37.5-41.6% according to the results of immunofluorescent diagnostics and 53-73% of ILD according to the results of the hemagglutination inhibition test). The occurrence of influenza B during this season was very low. PMID- 10852032 TI - [Variants in the immune response to revaccination with diphtheria anatoxin in adults]. AB - The immunological effectiveness of the revaccination (made in two injections) of 488 adults aged 18-67 years with diphtheria-tetanus toxoid is discussed; the parallel study of the results of this revaccination was carried out in the diphtheria toxin neutralization test on Vero cells and in the passive hemagglutination (PHA) test. The specific features of the dynamics of the increase of diphtheria antitoxic antibodies, depending on the initial immunity level, the age and the sex of revaccinated persons, were determined. Among persons with the low level of circulating antibodies before revaccination four variants of immune response to the injection of diphtheria toxoid were registered: variant 1--rapid reaction like in secondary immune response (53.6%); variant 2--delayed but effective reaction like in primary immune response (27.3%); variant 3--slow weak response (6.5%); and variant 4--the absence of effective immune response to immunization made in 2-3 injections (12.6%). The immunological and neutralizing properties of diphtheria antitoxic antibodies in the process of immunization made in 2 injections were evaluated. Persons with abnormal immune response (variants 3 and 4) produced defective antibodies, displaying immunological activity in the PHA test, but in most cases unable to neutralize diphtheria toxin in vitro when tested on Vero cells. PMID- 10852033 TI - [Monoclonal antibodies in the diagnosis of infections caused by the herpes simplex virus]. AB - A series of hybridomas producing monoclonal antibodies (McAb), specifically interacting with Herpes simplex virus (HSV) proteins, types 1 and 2, has been obtained. McAb 7c4 and 4f6 have been shown to be highly active in the solid-phase enzyme immunoassay (EIA) and to produce no reaction with HSV antigen in the indirect immunofluorescent assay (IIFA). McAb 2b6, 3e5, 4A, 2C effectively detect McAb in IIFA, but not EIA, while McAb 3d 10 exhibit activity in both biochemical assays. Moreover, as established in this investigation, McAb 4A are active against the protein of HSV capsid, McAb 3d10 and 2b6 detect two individual epitopes on the molecule of ribonucleohydreductase, McAb 2C are specific with respect to surface glycoprotein gB, McAb 7c4 and 416 recognize one or two overlapping epitopes of protein gD. McAb 2C are capable of completely neutralizing the infectious activity of HSV in the in vitro cell system. As determined by IIFA, McAb 4A and 4e5 stain specific inclusions in the nucleus of HSV-infected cells, while McAb 2C stain HSV protein, localized in the cytoplasm. All above-mentioned McAb are active against two common antigenic determinants of HSV 1 and HSV 2. The data obtained in this investigation suggest that the series of McAb under study may serve as the basis for the development of diagnostic test systems for the detection of HSV, types 1 and 2, by EIA and IIFA techniques. PMID- 10852034 TI - [The adaptational properties and immunoregulatory action of a preparation of proteolytic enzymes in experimental stress]. AB - The use of the experimental model of the development of acute secondary immunodeficiency, accompanied by the redistribution of immunoglobulins from plasma to blood cells, for evaluating the effect of immunomodulation is substantiated. The preparation of proteolytic enzymes Wobenzym [correction of "Vobenzyme"] has been shown to produce significant and dose-dependent decrease the sorption of immunoglobulins on mouse cells in the process of swimming. PMID- 10852035 TI - [The modulation of phagocyte oxygen metabolism by recombinant cytokines and a complex of natural cytokines]. AB - The data on the modulating function of cytokines on the oxygen-producing function of peritoneal exudate cells of rats are presented. As priming agents, recombinant cytokines IL1 beta and TFR beta 1, as well as the natural complex of cytokines, were used. The priming action of cytokines was studied by changing in the production of active forms of oxygen by peritoneal exudate cells of rats, stimulated with opsonized zymosan, by the method of luminol-dependent chemiluminescence. The study revealed that IL1 beta and the natural complex of cytokines primed peritoneal exudate cells for the production of active forms of oxygen. The maximum value of the prestimulation index was 1.9 +/- 0.1 and 2.95 +/ 0.27 respectively. The preincubation of peritoneal exudate cells of rats with TFR beta 1 led to the pronounced inhibition of the intensity of the chemiluminescent response of cells. The prestimulation index did not exceed 1.06 +/- 0.1. Moreover, as revealed with the use of the probe Fura-2/AM, in the process the prestimulation of phagocytes with the natural complex of cytokines the intracellular concentration of calcium increased from 0.86 +/- 0.15 to 1.86 +/- 0.2 microM/ml. The mechanism of the prestimulation of peritoneal exudate cells of rats cytokines seems to be calcium-dependent. PMID- 10852036 TI - [The determination of lipopolysaccharide admixtures in protein preparations]. AB - A new method for the determination of lipopolysaccharide (LPS) admixtures in protein solutions has been developed. The method includes the periodate oxidation of LPS, biotinylation with biotin hydraside, immobilization on a nitrocellulose membrane and the development of biotinylated LPS in the streptavidin--alkaline phosphatase system. Proteins are previously removed from the solution by treatment with hot phenol. Development with the use of 5-bromoinodyl phosphate and nitrotetrazolium blue makes it possible to detect about 30 pg of LPS immobilized on the nitrocellulose membrane. PMID- 10852038 TI - [The possible effect of the components of the alternative activation pathway of the complement system on the outcome of HIV infection]. PMID- 10852037 TI - [The development and testing of test systems for the determination of herpes simplex virus and cytomegalovirus antigens by lanthanide immunofluorescence analysis]. AB - On the basis of the lanthanide immunofluorescent assay (LIFA) test systems for the determination of herpes simplex virus (HSV-LIFA) and cytomegalovirus (CMV LIFA) antigens have been developed. The test system HSV-LIFA includes the sandwich of monoclonal antibodies (McAb) HSV A.3.3. or B-11 to type 2 HSV, strain BH; the test system CMV-LIFA includes the sandwich of rabbit McAb to CMV, strain AD 169. The approbation of the test systems has revealed that they insure the specific detection of HSV and CMV antigens in clinical specimens (urine, blood, liquor, saliva), the LIFA results well correlating with the data on the isolation of viruses in cell cultures, with the results obtained by other diagnostic methods and with clinical manifestations of diseases. LIFA has been shown to be more sensitive than the enzyme immunoassay. PMID- 10852039 TI - [The reciprocal effect of the causative agents in a mixed infection in burn injury]. AB - In vitro experiments on the joint cultivation of Pseudomonas aeruginosa, Escherichia coli, staphylococci and in vivo experiments carried out on mice with the experimental mixed infection of a burn injury revealed the pronounced antagonistic action of P. aeruginosa with respect to staphylococci and E. coli. Under the same conditions, in the joint cultivation of P. aeruginosa and fungi of the genus Candida and in the mixed infection of a burn wound caused by the same microorganisms the mutual stimulation of multiplication and a considerable aggravation of the clinical course of a burn wound were observed. The mutual influence of associates in the mixed infection of a burn injury is manifested by an increase in the number of an antibiotic-resistance microorganisms in their populations and, so far as the macroorganism is concerned, in the aggravation of the course of the infectious process and the formation of the pronounced state of immunodeficiency. PMID- 10852040 TI - [The subcellular mechanisms of the action of disinfectants. The effect of iodine and chloramine-B on bacterial ribosomes]. AB - The composition of ribosomal material in bacterial cells of 5 families, subjected to the minimal bactericidal action of iodine and chloramine-B, was studied with the use of electrophoresis in polyacrylamide gel. The study demonstrated that under these conditions protein-synthesizing organelles of vibrios and bifidobacteria did not undergo perceptible changes, while Escherichia, Pseudomonas and bacilli essential ribosomal degradation was found to occur, which was manifested by a decrease in the intensity of bands 23S and 16S of ribosomal RNA. The conclusion was made that the in vivo destruction of ribosomes was not the direct result of the action of disinfectants, but mediated by some cellular mechanisms. PMID- 10852041 TI - [The microbiological characteristics of soft-tissue surgical infection]. AB - The dynamic study of microflora in the foci of surgical infection of soft tissues was made. In 45 patients the course of the infectious process was characterized by the change of causative agents in the course of the disease. As established in this study the frequency of changes in the species composition of microflora in the foci of surgical infection in the course of the disease directly correlated with the occurrence of purulent septic complications requiring repeated surgical intervention. Changes in the species composition of causative agents should be regarded as their adaptation to the anti-infectious protection of the body, which is one of the regulatory mechanisms of the parasitic system. PMID- 10852042 TI - [Morbidity from zooanthroponotic and natural-focus infections and the measures for their prevention]. PMID- 10852043 TI - [Experience in using correlation analysis for evaluating the synchronism of the fluctuations in the level of infectious morbidity in individual areas]. AB - The possibility of using correlation analysis for evaluating the synchronism of changes in infectious morbidity rate on different territories was studied. The study demonstrated that the intensity of correlative relationships was mainly influenced by the degree of manifestation and the direction of prolonged tendencies in the changes of infectious morbidity on the territories under study. The presence of prolonged pronounced tendencies towards growth or decrease in the compared curves characterizing prolonged morbidity dynamics was found to limit the use of correlation analysis for the evaluation of synchronism of changes in morbidity rate. PMID- 10852044 TI - [Diphtheria in the Maritime Territory]. AB - The results of the analysis of the present epidemiological situation in diphtheria in the Maritime Territory are presented. The data on absolute and intensive characteristics, death rate due to this infection, the vaccination status of diphtheria patients, as well as the results of the study of the level and intensity of antidiphtheria immunity in the healthy population of the territory. The study revealed that the defects of collective antidiphtheria immunity correlated with morbidity rate among adults and children. Adults aged 40 50 years and children aged 6-7 years were regarded as risk groups; among them the highest proportion of nonimmune persons and the highest diphtheria morbidity rate were registered. Stabilization achieved by the present moment was the result of the mass immunization of the adult population, ensuring the necessary coverage (92%) of vaccination in 1995. PMID- 10852045 TI - [Viral hepatitis E: its epidemiological characteristics in the Republic of Tajikistan]. AB - The characteristic feature of the Republic of Tajikistan, as well as other republics of Central Asia, is the wide spread of virus hepatitis E. The epidemiology of this grave disease, recently known as virus hepatitis non A, non B with the fecal-oral mechanism of transmission of this infection, has been yet insufficiently studied. The article points out to the specific character of this infection which essentially differs, both epidemiologically and clinically, from other enteric hepatitis (hepatitis A), also hyperendemic for the republic. The results of the study of the immunostructure of the population with respect to both hepatitis E and hepatitis A are presented. PMID- 10852046 TI - [The vaccination of children with severe somatic pathology]. AB - 1,696 children were vaccinated; of these, 1,487 children had different kinds of somatic pathology, including 1,181 children with CNS lesions, 29 children with malignant tumors, 45 children with congenital defects, 82 children with allergic diseases, etc. The group of relatively healthy vaccinees consisted of 209 children. The following vaccines were used for immunization: Tetracoq 05, D.T.Vax, Rudivax, Imovax Polio, Vaxigrip (Pasteur Merieux Connaught, France); HBVax, MMRII (Merck Sharp & Dohme, USA); as well as vaccines against hepatitis B produced by Smith Kline Beecham (UK) and Combiotech (Russia). In no case severe vaccine-associated complications were observed. The frequency and manifestation of reactions in children with somatic pathology did nor essentially differ from those in relatively healthy children. The increase of the number of vaccine components did not lead to the increase of the number of side effects of the severity of their manifestation. These investigations demonstrated the safety of vaccination for children with somatic pathology. PMID- 10852047 TI - [The immunological efficacy of one, two and three inoculations against diphtheria in adults]. AB - The outbreak of diphtheria in the Republic of Belarus developed in the presence of a sharp deficiency of immunity among the population aged 35 years and over. In connection with a higher initial immunity level resulting from vaccination made in one injection the minimum antibody level (> or = 0.01 I.U./ml) was found in 79.7% of the examinees among the population under 35 years. For the age group of 35-60 years vaccination in one and two injections proved to be insufficiently effective (the minimum protective antibody titer was found, respectively, in 64.3% and 63.3% of the vaccinees). To achieve the full protection of this group of the population, vaccination in three injections was necessary; after that the minimum protective antibody titer was found in 90.7%, the protective antibody titer (0.1 I.U./ml) was found in 53.5% and the highly protective antibody titer (1.0 I.U./ml) was found in 11.6% of the examinees. The low level of highly protected persons among vaccinees receiving three injections was due to a low dose of diphtheria toxoid (2 Lf) in adsorbed DT-m vaccine with reduced antigen content. PMID- 10852048 TI - [The evaluation of the reactogenicity and immunogenic activity of a new concentrated inactivated leptospirosis vaccine]. AB - In the controlled field trial the reactogenicity, safety and antigenic activity of a new concentrated inactivated leptospirosis vaccine after its administration in one and two injections of 0.5 ml were studied in comparison with those of the existing commercial vaccine, introduced in two injections in doses of 2.0 and 2.5 ml. The new experimental vaccine exhibited low reactogenicity and was found to be safe and highly immunogenic when introduced in a single injection of 0.5 ml. As shown in this trial, the immunogenic characteristics of immunization made in a single injection were not inferior than those obtained as the result of immunization made in two injections, yielding high percentage of seroconversions (89.8% to 98.3%) with respect to 4 Leptospira serogroups and leading to the production of the protective titers of corresponding antibodies. The existing commercial vaccine was inferior to the experimental one in antigenic activity (the frequency of seroconversions, antibody titers). The results of the trial make it possible to recommend the experimental concentrated leptospirosis vaccine for use in medical practice in a dose of 0.5 ml introduced in a single injection. PMID- 10852049 TI - [The evaluation of the efficacy of a first course of immunization against diphtheria and tetanus in children with kidney diseases]. AB - The results of the determination of the level of antitoxic antibodies (Ab) to diphtheria and tetanus toxoids in children with glomerulonephritis and pyelonephritis are presented. The influence of the form of the disease and the kind of medicinal therapy on level of specific Ab has been studied. As shown in this study, the injection of adsorbed DT toxoid with reduced antigen content leads to the formation of the protective titers of antidiphtheria and antitetanus Ab, but a lower level than after the injection of adsorbed DPT vaccine. PMID- 10852050 TI - [The revaccination against diphtheria and tetanus with adsorbed DT-m anatoxin of children having solid tumors in their case histories]. AB - Data on the safety and effect of the vaccination of children with solid tumors are presented. As revealed in this study, the injection of adsorbed DT-m toxoid with reduced antigen content at the period of remission does not induce the relapse of the disease and leads to the production of specific antitoxic antibodies on the protective level. The study has shown that complex antitumor treatment does not essentially affect the formation of specific immune response, though prolonged changes in cell-mediated immunity can be observed. PMID- 10852051 TI - [The immunomodulating and antitoxic properties of preparations of lipopolysaccharides from representatives of the genus Francisella]. AB - The study of the biological properties of lipopolysaccharides (LPS) of bacteria of the genus Francisella (F. tularensis, F. novicida, F. novicida-like, F. philomiragia) revealed that the preparation of Francisella LPS possessed immunomodulating and antitoxic properties in the absence of toxicity. At the same time the structure of LPS (S or R) was found to produce an essential effect of the immunobiological activity of this molecule. Thus, the S-forms of LPS proved to be more effective as immunomodulators and the R-forms of LPS, as antagonists of classical endotoxins. PMID- 10852052 TI - [The effect of platelet-activating factor and its antagonist--BN 52021--on immune reactions in vivo in mice and in vitro]. AB - The effect produced by the injection of platelet activation factor (PAF) and its antagonist BN 52021 on the intensity of humoral immune response in (CBA x C57BL)F1 mice was studied. PAF was found to stimulate the formation of antibodies to sheep red blood cells. In addition PAF stimulated the phagocytic activity of mouse peritoneal macrophages. The stimulation of immune response under the action of PAF may be attributed to an increase in the phagocytic activity of macrophages. The stimulating effect of PAF on immune response in vivo was abolished by the injection of BN 52021, the antagonist of PAF. At the same time the dose-dependent decrease of immune response was observed after the injection of BN 52021. Indomethacin, an inhibitor of prostaglandin synthesis, when administered to mice treated with BN 52021, abolished the BN 52021-induced suppression of humoral immune response. Mouse peritoneal macrophages, treated in vitro with BN 52021, were found to produce significantly more prostaglandin E than control macrophages. Thus, BN 52021 induced the suppression of humoral immune response in vivo; this suppression was probably due to the action of prostaglandin E2, a messenger of the second order. Besides, the PAF antagonist BN 52021 significantly decreased leukotriene B4 production by macrophages in vitro. BN 52021 may be supposed to switch over the synthesis and/or secretion of arachidonic acid from the lipoxygenase pathway to the cycloxygenase one. PMID- 10852053 TI - [Immuno- and erythropoiesis in mice with graft vs host disease against a background of immunodeficiency]. AB - In BALB/c mice immunodeficiency was induced by the transfer of lymphocytes immune to alloantigen. This model is one of experimental models of AIDS. The work was aimed at the study of disturbances in the immuno--and erythropoiesis in immunodeficient mice. The state of erythropoiesis was evaluated by the level of level of hemoglobin, hematocrit and the content of reticulocytes in peripheral blood, by the number of erythroid bursitis-forming units and the percentage of erythrokaryocytes in the marrow, as well as by the number of colony-forming units in the spleen by days 5 and 8. The study revealed that in BALB/c mice hypoplastic anemia, accompanied by the decreased phagocytic activity of macrophages and the reduced production of interleukin 1 and tumor necrosis factor, developed on months 5-6 of the disease. Macrophagal dysfunction was supposed to be one of the causes of hypoplastic anemia in immunodeficient mice. PMID- 10852054 TI - [The effect of Mycoplasma arthritidis superantigen on immunogenesis in transplacental infection]. AB - The immune responsiveness of the progeny of BALB/c mice, responsive to M. arthritidis superantigen (MAS), and C57BL/6 mice, nonresponsive to MAS, infected with M. arthritidis during the second half of pregnancy was studied. The investigation revealed that in responsive animals the proliferative response of spleen cells to MAS was suppressed with the level of response to concanavalin A remaining unchanged. The spleen cells of the test mice reacted to syngenic intact cells as to xenogenic ones and suppressed reaction to MAS and the production of interleukin 1 in the culture of spleen cells taken from the intact syngenic animals. The data obtained in this study suggest that after the infection of pregnant BALB/c mice with M. arthritidis immune tolerance to MAS developed in their progeny, which was accompanied by the induction of suppressor cells inhibiting the production of interleukin 1. PMID- 10852055 TI - [The characteristics of the vaginal microflora in intrauterine contraception]. AB - The results of the study of vaginal microbiocenosis in women using intrauterine contraceptives are presented. Inflammatory complications in intrauterine contraception were shown to be linked with vaginal dysbiosis. Disturbances in vaginal microbiocenosis were characterized by the deficiency of lactoflora and the presence of enterobacteria, Staphylococcus aureus, Gardnerella of fungi of the genus Candida. The problem of the possibility of complications of microbial etiology in women using intrauterine contraceptives are discussed. PMID- 10852056 TI - [The microbiological characteristics of intestinal dysbacteriosis in children and adults in the city of Moscow]. AB - The bacteriological analysis of 302 children has revealed that dysbacteriosis with the increased content of hemolytic Escherichia in the intestine develops more frequently in the presence of the deficiency of bifido- and lactobacteria. The development of other kinds of dysbacteriosis with the increased content of different opportunistic enterobacteria, staphylococci, enterococci and fungi, as well as dysbacteriosis with the decreased content of Escherichia, does not practically depend on the deficiency of bifido- and lactobacteria. In patients with the increased content of Escherichia an increase in the content of opportunistic enterobacteria, staphylococci and fungi is observed more frequently than in patients with the low content of Escherichia. PMID- 10852057 TI - [The diagnosis of intestinal dysbacteriosis by the spectrum of fecal amino acids]. AB - As reveals in the analysis of 45 fecal specimens taken from patients, the occurrence of amino acids of indigenous origin increases with the increase of the imbalance in the intestinal microflora (31.2% in dysbacteriosis of the I degree and 75% in cases of the III degree). The presence, or absence, of beta aspartyllysine, beta-aspartylglycine, gamma-aminobutyric acid, 5-aminovalerianic acid, lactosolysine, fructosolysine, ornithine is the sign of the development of dysbacteriosis. PMID- 10852058 TI - [The immuno-microbiological characteristics of the small intestine and the translocation of the enteral microflora in acute intestinal obstruction]. AB - The complex examination of 72 patients with acute ileus (AI) of nontumor nature with different severity of endotoxicosis was carried out. The study revealed that AI was accompanied by deep suppression of the immunosecretory and motor evacuatory function of the small intestine, as well as by its pronounced bacterial contamination, mainly due to the significant quantitative prevalence of Gram-negative microflora. The combination of these factors played the key role in the increase of the permeability of the enteric barrier for symbiotic microflora and its massive translocation from the intestinal tract to the internal organs of the body (peritoneal exudate, portal bed), which directly correlated with the severity of endotoxicosis in AI patients. The deficiency of the barrier function of the liver was accompanied by the penetration of infective agents into the general blood stream, thus causing the development of endotoxic shock in AI patients. The analysis of the results thus obtained made it possible to determine the main ways for the elimination of intestinogenic intoxication in AI; they should be aimed at the bacterial decontamination of the small intestine, the restoration of its motor evacuatory and protective barrier functions, the liquidation of portal and systemic bacteremia, the correction of the functional deficiency of the liver. PMID- 10852059 TI - [The vaginal Bifidobacterium flora in women of reproductive age]. AB - The composition of vaginal bifidoflora in 56 clinically healthy women of reproductive age was studied. The study revealed that four species of bifidobacteria, viz. Bifidobacterium bifidum, B. breve, B. adolescentis 2 and B. longum, dominated in the composition of this bifidobacterial population. Nine out of 11 isolated strains were found to be capable of inhibiting indicator microorganisms Staphylococcus aureus and Enterococcus faecalis when tested in vitro; in addition, strains B. adolescentis 2 F1, B. bifidum G1, B. breve P2 and B. longum Z4 inhibited Klebsiella ozaenae, Pseudomonas aeruginosa, Escherichia coli and were also active acid producers. Three of these 4 bifidobacterial strains were capable of adhesion to vaginal epitheliocytes, while B. bifidum G1 was practically incapable of adherence to these cells, similarly to B. bifidum strain 791 of intestinal origin. In addition, the spectra of antibiotic susceptibility varied from strain to strain, but all bifidobacterial strains were susceptible to benzylpenicillin and resistant to lomefloxacin, most of them being also resistant to cyprofloxacin and gentamicin. Thus the data presented in this work are indicative of the possibility and advantages of using bifidobacterial strains belonging to this ecological niche as probiotics for the correction of the microflora of the urogenital tract in females. PMID- 10852060 TI - [The use of internal controls of different lengths in the detection of Chlamydia trachomatis DNA by the polymerase chain reaction method]. AB - To detect C. trachomatis DNA, the polymerase chain reaction (PCR) with the use of primers corresponding to variable sites of rRNA gene 16S was carried out. As the positive control of the reaction, the amplification fragment of gene 16S of rRNA, cloned in the plasmid vector and having the length of 530 nucleotide pairs (n.p.), was used. On its basis 2 kinds of the internal control of the reaction were obtained with the deletion of 110 n.p. (pMOS-Chl420) and the insertion of 930 n.p. (pMOS-Chl1460) within the cloned amplification fragment. The study revealed that the addition of the DNA of pMOS-Chl420 or pMOS-Chl1460 into the reaction mixture did not affect the sensitivity of PCR (0.02 pg of bacterial DNA in the sample) in the detection of C. trachomatis DNA isolated both from the culture of bacterial cells and from clinical samples. But in some cases of the amplification of the DNA of internal control pMOS-Chl420, but not pMOS-Chl1460, was observed in the presence of DNA obtained from clinical samples. It was supposedly linked with a higher sensitivity of Taq DNA-polymerase to the action of inhibitors in the synthesis of high-molecular DNA fragments. The observed high frequency of the inhibition (17%) of PCR makes it expedient to carry out this reaction with the use of the internal control. PMID- 10852061 TI - [The use of the polymerase chain reaction in the diagnosis of pulmonary tuberculosis]. AB - The data on the use of the polymerase chain reaction (PCR) with primers INS1 and INS2 for the diagnosis of pulmonary tuberculosis are presented. All stages of PCR are described: from the treatment of biological material to the conditions necessary for carrying PCR and the registration of the results. Simultaneously with this reaction, PCR in a Cobas Amplicor apparatus was carried out and Mycobacterium tuberculosis culture was grown in a liquid medium in an MB/BacT apparatus. The study revealed that the method of PCR in pure M. tuberculosis culture made it possible to detect even the DNA of those cells which formed no colonies on Lowenstein--Jensen medium. The detection of M. tuberculosis in clinical samples (sputum, pleural exudate) taken from 31 patients with different pulmonary pathology showed that in 87.1% of cases diagnostics with the use of PCR carried out in a Cobas Amplicor apparatus and with primers INS1 and INS2 yielded similar results. In patients with pulmonary tuberculosis the results of PCR were positive, while the results of the analysis made with use of an MB/BacT apparatus were negative. The proposed primers INS1 and INS2, the conditions of amplification and detection make up the test system for the detection of mycobacteria of the tuberculosis complex. PMID- 10852062 TI - [The detection of house dust mites in the apartments of children with allergic diseases]. AB - The acarological study of house dust in 239 apartments of children with different allergic diseases (bronchial asthma, rhinosinusitis, atopic dermatitis) developing as the result of indoor sensitization has been carried out. The significant difference between the occurrence of house-dust mites in apartments inhabited by healthy and sick children has been established. The occurrence of mites and their number have been found to affect the symptoms of an allergic disease and its exacerbation. PMID- 10852063 TI - [The diagnostic value of determining the Klebsiella persistence factors in dysbacteriosis]. PMID- 10852064 TI - [The effect of the glucocorticoid immunodepressant kenalog on the resistance of mice to the influenza virus]. PMID- 10852065 TI - [The detection and characteristics of the Yersinia pestis antigen exhibiting the properties of S-layer proteins]. PMID- 10852066 TI - [The capacity of the causative agent of anthrax to reduce methylene blue]. PMID- 10852067 TI - [The use of dot analysis and immunogold markers for the rapid diagnosis of acute intestinal infections]. PMID- 10852068 TI - [Diphtheria in Pskov Province over the course of 43 years]. PMID- 10852069 TI - [The treatment of bacterial keratitis with infrasound and its effect on reparative process in the cornea]. PMID- 10852070 TI - [The role of herpesviruses, mycoplasmas, toxoplasmas and the rubella virus in the etiology of manifest intrauterine infections in the Republic of Belarus]. PMID- 10852071 TI - [A comparative evaluation of different diagnostic agents in the PHA in brucellosis]. PMID- 10852072 TI - [Bacterial vaginosis]. PMID- 10852073 TI - [The current status of the problem of Vibrio cholerae variability]. PMID- 10852074 TI - [The basic properties of the causative agent of chlamydiosis and its role in the development of urogenital tract infections]. PMID- 10852075 TI - [Veterinary public health and its importance in human infectious pathology]. PMID- 10852076 TI - [The prognosis of the HIV infection epidemic in Russia]. PMID- 10852077 TI - [The epidemiological and epizootiological situations on rabies in the Jewish Autonomous Province]. PMID- 10852078 TI - [An outbreak of salmonellosis in a city hospital due to Salmonella typhimurium (1)]. PMID- 10852079 TI - [An outbreak of salmonellosis in one of the wholesale markets of Moscow due to Salmonella enteritidis]. PMID- 10852080 TI - [An outbreak of salmonellosis in a city hospital due to Salmonella typhimurium (2, with the authors' comments)]. PMID- 10852081 TI - Optimization of inpatient warfarin therapy: impact of daily consultation by a pharmacist-managed anticoagulation service. AB - OBJECTIVE: To determine the effect of daily consultation by a team of hospital pharmacists on the accuracy and rapidity of optimizing warfarin therapy. DESIGN: Comparison of a historical control cohort with a prospective cohort matched for treatment indication. SETTING: A 400-bed university teaching hospital. PATIENTS: Sixty consecutive patients hospitalized in 1992 and starting warfarin for the first time, with anticoagulation therapy managed by physicians, were compared with 60 patients matched for warfarin indication hospitalized in 1995, but with anticoagulation therapy managed with pharmacy consultation. RESULTS: Pharmacist management of initial warfarin therapy resulted in a significant reduction in the length of hospitalization compared with physician dosing, from 9.5 +/- 5.6 days to 6.8 +/- 4.4 days (p = 0.009). The number of patients and patient-days with international normalized ratio (INR) values >3.5 were reduced by pharmacist dosing from 37 patients and 142 days to 16 patients and 29 days, respectively (p < 0.001). Similarly, the number of patients and patient-days with INR >6.0 were reduced from 20 patients and 50 days to two patients and six days, respectively (p < 0.001). There were six documented bleeding complications in 1992 compared with one in 1995 (p = 0.11). The mean INR at discharge was significantly lower in the pharmacy surveillance group, 2.6 +/- 0.58, compared with the physician cohort, 3.3 +/- 2.1 (p = 0.07). Readmissions after discharge due to bleeding or recurrent thrombosis were reduced from five (at 1 mo) and 10 (at 3 mo) to two and five readmissions, respectively, by pharmacist intervention (p = 0.43). The number of patients with concurrently prescribed drugs known to significantly interact with warfarin was significantly lower (6 vs. 13; p = 0.02) in the pharmacy surveillance group. CONCLUSIONS: Among patients starting warfarin for the first time, daily consultation by a pharmacist significantly decreased the length of hospital stay and the number of patients who received excessive anticoagulation therapy. These findings translate into improved quality of care and potentially significant cost savings. PMID- 10852082 TI - Anthropometric and pharmacotherapeutic variables on acute emesis induced by cisplatin-containing chemotherapy. AB - OBJECTIVE: To characterize the effects of anthropometric and pharmacotherapeutic variables on acute emesis induced by cisplatin-containing regimens with dosages > or =50 mg x m(-2). METHODS: A prospective, cross-sectional, noncontrolled study was performed to analyze acute vomiting during the first 24 hours in patients treated in a Spanish hospital. The patients received an intravenous combination of drugs (2 doses of metoclopramide 3 mg/kg, dexamethasone 20 mg) as first-choice antiemetic therapy. Intravenous ondansetron 8 mg and dexamethasone 20 mg served as an alternative regimen in patients <30 years old with a history of extrapyramidal manifestations or emesis in previous cycles. Therapeutic failure was used as a dependent variable, defined as three or more vomiting episodes documented by the patients. Other variables were the chemotherapeutic regimen; antiemetic regimen; patient gender, age, weight, and height; and cycle number. The reference logistic model and two reduced-models derived from the latter were designed. The logistic models were subsequently validated by means of receiving operating characteristic curves. RESULTS: A total of 319 cycles involving 106 patients were studied. The metoclopramide regimen was administered in 66% of the cycles. The therapeutic failure rate was 21% for the metoclopramide regimen and 32% for the ondansetron treatment. The logistic model developed identified the type of chemotherapeutic regimen provided as the most significant prognostic variable (p < 0.0001). Patient weight (odds ratio 1.64) and height (odds ratio 1.28) were identified as prognostic factors related with therapeutic failure. CONCLUSIONS: The type of chemotherapeutic regimen administered and the anthropometric characteristics of the patients exert a clear conditioning effect on risks associated with therapeutic failure against acute emesis following high dose cisplatin therapy. Such anthropometric parameters have not been previously identified as prognostic factors. PMID- 10852083 TI - Hyperglycemia associated with protease inhibitors in an urban HIV-infected minority patient population. AB - BACKGROUND: Hyperglycemia and new-onset diabetes mellitus have been reported to occur in HIV-infected patients treated with protease inhibitors. OBJECTIVE: To determine the effect of protease inhibitor therapy on serum glucose in a predominantly minority patient population. DESIGN: Retrospective record review. SETTING: Clinical HIV program of an urban Veterans Affairs medical center. PATIENTS: All HIV-infected patients receiving a protease inhibitor over a one year period from September 1996 through August 1997. RESULTS: One hundred seventeen patients not previously known to be diabetic received protease inhibitors; seven (6%) developed symptomatic diabetes mellitus. Eight other patients had one or more serum glucose values >150 mg/dL. Mean random glucose values for patients who did not develop diabetes were higher during therapy than prior to initiation of protease inhibitors. CONCLUSIONS: Urban minority HIV infected patients receiving combination antiretroviral therapy including a protease inhibitor may be at increased risk for the development of hyperglycemia and diabetes mellitus. Risk factors for diabetes mellitus should be identified and blood glucose monitored in all patients receiving protease inhibitors. PMID- 10852084 TI - Impact of the dial access drug information service on patient outcome. AB - OBJECTIVE: To determine the impact of a drug information service on patient outcomes. DESIGN: Prospective evaluation of patient-specific drug information requests. SETTING: Healthcare professional and consumer drug information service located at a college of pharmacy. PARTICIPANTS: Consumers and healthcare professionals of the province. INTERVENTION: Patient-specific questions received by the drug information service were reviewed and evaluated for actual patient outcome, inquirers' opinion of impact of the service with respect to patient outcome, and for objectivity and timeliness of the response. An expert panel determined whether the responses and recommendations given by the service were appropriate, determined what impact the service had on the patient, and assessed the seriousness of the inquiry. MAIN OUTCOME MEASURE: Classification of patient outcome by objective and subjective data based on predetermined desired outcomes. RESULTS: Ninety-eight and 68 patient-specific requests were received from healthcare professionals and consumers, respectively. The panel concluded that 94.9% of the healthcare requests and 98.5% of the consumer requests were answered appropriately and that the majority of the requests involved potentially serious drug-related problems. The panel also determined that 46.8% of the recommendations to healthcare professionals and 41.0% of the recommendations to consumers resulted in positive patient outcomes. The majority of the positive outcomes involved the prevention of a disease or its symptoms (professional section) and the reduction or elimination of symptoms (consumer section). CONCLUSIONS: The drug information service not only met its objectives of providing drug information in an accurate, objective, and timely manner, but was also able to provide positive patient outcomes. PMID- 10852085 TI - Nephrology pharmaceutical care preceptorship: a programmatic and clinical outcomes assessment. AB - OBJECTIVE: The University of Pittsburgh Nephrology Pharmaceutical Care Preceptorship (NPCP) program was conceived to acquaint health system pharmacists with the pharmacotherapeutic management of dialysis patients, enhance the delivery of pharmaceutical care, and improve clinical outcomes through the development of specialized professional skills. A survey designed to determine the impact of the NPCP program was sent to all 145 participants of the program. METHODS: The survey, designed to collect demographic information and data about the participants' practice sites, professional activities prior to and after the completion of the program, and markers of disease status, was mailed to all participants in September 1997. The 96 respondents (66.2%) were involved in a wide variety of clinical practices; inpatient management of peritoneal dialysis, hemodialysis, or renal transplant patients were most commonly reported. RESULTS: More than 80% of the participants believed that the educational content of the NPCP program was sufficient to allow them to establish a specialized service for the management of dialysis patients. However, two-thirds would have preferred to have more contact time (an additional 1-2 d) with the preceptorship faculty. The percentage of the pharmacists' time devoted to the provision of pharmaceutical care for dialysis patients almost doubled, from 13.1% to 25.2% (p < 0.001). The components of pharmaceutical care performed by these pharmacists also changed as a result of their completion of the NPCP program. Time devoted to clinical services and the provision of educational programs (inservices) increased significantly, while the time allocated to distributive activities decreased from a mean of 32.4% to 26.4% (almost 20% from baseline). The number of pharmacists who provided some component of pharmaceutical care for ambulatory dialysis patients increased significantly, from 10 to 33, after completion of the program. In the survey given after the preceptorship, almost 70% of these 33 pharmacists self-reported that the mean hematocrit of their ambulatory dialysis patients increased; 45% reported that the epoetin dose was lower. Parenteral iron use was also reported to have increased in 78.8% of the dialysis units, and an increase in serum ferritin and transferrin saturation was observed in 54.5% and 60.6% of the units. respectively. Although far fewer pharmacists (n = 15) initiated a renal osteodystrophy management program, 73.3% of those who did so reported an increase in their patients' compliance with phosphate binder therapy, which was reflected in a drop in serum phosphorous in 40% of the units. CONCLUSIONS: The NPCP program resulted in changes in the professional activities of the participants: fewer distributive activities and increased clinical and educational activities. These significant changes were noted in all areas of outpatient care. Participation in the NPCP program enhanced the delivery of pharmaceutical care to dialysis patients and improved the markers of disease status. PMID- 10852086 TI - Stability of suspension formulations of lansoprazole and omeprazole stored in amber-colored plastic oral syringes. AB - OBJECTIVE: To determine the stability of lansoprazole and omeprazole suspensions at ambient and refrigerated temperatures using HPLC. DESIGN: The contents of lansoprazole and omeprazole capsules were suspended in separate flasks containing sodium bicarbonate 8.4% to concentrations of 3 and 2 mg/mL, respectively. The contents of each flask were drawn into six amber-colored oral syringes, with one half of the syringes stored at 22 degrees C (ambient) and the other half at 4 degrees C. Lansoprazole and omeprazole concentrations were determined by a stability-indicating HPLC assay at baseline and at 4, 8, 12, and 24 hours, and on days 4, 7, 14, 21, 30, 45, and 60 after mixing. Both omeprazole and lansoprazole were considered stable if they retained > or =90% of the baseline drug concentration. RESULTS: Omeprazole was stable for up to 14 days at 22 degrees C and 45 days at 4 degrees C. Lansoprazole was stable for eight hours at 22 degrees C and for 14 days at 4 degrees C. CONCLUSIONS: When compared with ambient or refrigerated storage conditions, omeprazole was stable for a longer duration than lansoprazole. Pharmacists may use these results to guide compounding and storage of proton-pump inhibitor suspensions. PMID- 10852087 TI - Fludrocortisone for the treatment of heparin-induced hyperkalemia. AB - OBJECTIVE: To report the use of fludrocortisone for heparin-induced hyperkalemia and to briefly review the available literature relating to heparin-induced hyperkalemia. CASE SUMMARY: A 34-year-old African-American man was admitted to the hospital for pneumococcal pneumonia and sepsis. His hospital course was complicated by the development of acute respiratory distress syndrome, severe sepsis, acute renal failure, placement of a tracheostomy, and recurrent nasopharyngeal bleeding. The patient also developed a subclavian vein thrombosis with extension to the cephalic and basilic veins secondary to placement of a pulmonary artery catheter; anticoagulation with heparin was required. On day 9 of heparin therapy, the patient developed symptomatic hyperkalemia refractory to conventional therapies. Oral fludrocortisone 0.1 mg/d was initiated with resolution of the hyperkalemia within 24 hours despite the continued administration of heparin. DATA SOURCES: A MEDLINE (1966-October 1999) search was performed to identify case reports and clinical trials discussing heparin-induced hyperkalemia or the use of fludrocortisone for hyperkalemia. DISCUSSION: Heparin has the potential to induce hyperkalemia by several mechanisms, including decreased aldosterone synthesis, reduction in number and affinity of aldosterone II receptors, and atrophy of the renal zona glomerulosa. Fludrocortisone promotes potassium excretion by its direct actions on the renal distal tubules. In this patient, fludrocortisone resulted in a significant and rapid decrease in serum potassium even with continued heparin administration and acute renal failure. CONCLUSIONS: This case suggests that fludrocortisone is a reasonable alternative therapy for patients with hyperkalemia secondary to heparin therapy when the continued administration of heparin is necessary. PMID- 10852088 TI - Hypertensive crisis and myocardial infarction following massive clonidine overdose. AB - OBJECTIVE: To describe a patient who experienced a hypertensive crisis and myocardial infarction following a massive dose of parenteral clonidine. CASE SUMMARY: A 62-year-old white woman with stage 3 breast cancer metastatic to the spine and a history of hypertension received a combined injection of hydromorphone 48.3 mg and clonidine 12.24 mg subcutaneously in an attempt to refill an implanted epidural infusion pump. She promptly developed mental deterioration, blurred vision, worsening respiration, tachycardia, and hypertension. She was immediately treated with naloxone, but subsequently experienced hypertensive urgency, a short-duration tonic-clonic seizure, and an anteroseptal myocardial infarction. Cardiac catheterization showed no arteriolar narrowing or blockage, but an anterior infarct was confirmed. DISCUSSION: Clonidine is a commonly used alpha-adrenergic agonist. At usual oral doses of 0.2 2 mg/d, it acts centrally to produce hypotensive effects; at doses >7 mg/d, it acts peripherally to stimulate alpha1- and alpha2-adrenergic receptors, leading to vasoconstriction and increased blood pressure. These effects are not easy to control by standard medical therapies and can cause significant morbidity. CONCLUSIONS: Clonidine, although a safe medication with usual dosages, must be used with caution when given in injectable form. An overdose of this alpha adrenoreceptor agonist can produce significant vasospasm and hypertensive emergency. Drugs used to treat overdose, such as naloxone, can potentiate clonidine's adverse effects, leading to further morbidity. PMID- 10852089 TI - Clozapine-associated extrapyramidal reaction. AB - OBJECTIVE: To report a case of extrapyramidal reaction associated with a dosage increase of clozapine. CASE SUMMARY: A 44-year-old white man with a 20-year history of chronic paranoid schizophrenia was admitted to an inpatient psychiatric facility. His prior medications restarted on admission were clozapine 650 mg at bedtime, haloperidol 10 mg at bedtime, clonazepam 2 mg/d, and aspirin 325 mg/d. Two days after admission (hospital day 3), clozapine and clonazepam were discontinued, and he was prescribed haloperidol 5 mg every morning and 10 mg every evening. Stabilization occurred over the following 24 days, with progressively lower dosages of haloperidol and increasing dosages of clozapine. Haloperidol was discontinued on day 24. On day 47, the patient was agitated and making bizarre statements; thus, the morning dose of clozapine was increased by 50 mg (total 450 mg/d). On day 48 at 2200, a dystonic reaction was diagnosed; he received intramuscular diphenhydramine 50 mg, which caused the reaction to subside. At the time of the adverse reaction, he was prescribed clozapine 450 mg/d, vitamin E 400 IU three times daily, aspirin 325 mg/d, and acetaminophen, milk of magnesia, and Maalox as needed. DISCUSSION: Although the risk of extrapyramidal symptoms (EPS) is significantly lower with clozapine than with conventional agents, elevated clozapine blood concentrations have been reported to cause EPS; other reports have cited severe dystonias and dyskinesias on abrupt clozapine withdrawal. Considering the medications prescribed at the time and the discontinuation of haloperidol 24 days before the event, clozapine was the most likely cause of the extrapyramidal reaction. CONCLUSIONS: Regardless of anticipated safety associated with novel antipsychotics such as clozapine, reports of dystonic reactions must be taken into account and patients monitored appropriately. PMID- 10852090 TI - Mania with bupropion: a dose-related phenomenon? AB - OBJECTIVE: To report a case in which bipolar depression was resistant to usual therapies, requiring dosages of bupropion >450 mg/d and to review the literature on mania associated with bupropion and propose a potential theory of a dose related threshold associated with bupropion and mania. CASE SUMMARY: A 44-year old white man with a 25-year history of bipolar affective disorder presented with depression resistant to usual therapies. Bupropion therapy was initiated and the dosage was titrated to 600 mg/d. After exceeding the maximum recommended daily dose (450 mg/d), he experienced a manic episode attributed to high-dose bupropion. DISCUSSION: Due to increased risk of seizures, current prescribing guidelines state that the total daily dose of bupropion is not to exceed 450 mg/d. Since bupropion is the agent least likely to cause a manic switch in bipolar disorder, this agent seemed a logical choice to treat the patient's depression. Due to a lack of response, the bupropion dosage was titrated to a maximum of 600 mg/d. Since the patient did not switch into mania until the dosage exceeded 450 mg/d, we speculate that this adverse reaction is a dose-related phenomenon. Scientific literature supports this theory. CONCLUSIONS: A switch into mania is a potential risk associated with antidepressant drug use in bipolar affective disorder. Bupropion is believed to be associated with a decreased risk compared with other antidepressant therapies. However, our case report as well as others support the theory that this decreased risk may be due to dosages not exceeding the recommended daily dose (450 mg/d). Doses of bupropion >450 mg/d should be used with caution in depressed patients with bipolar affective disorder. PMID- 10852091 TI - The use of calcium salts in the prevention and management of verapamil-induced hypotension. AB - OBJECTIVE: To review the available literature on the use of intravenous calcium salts for the prevention of hypotension associated with intravenous verapamil. METHODS: A MEDLINE search (1966-June 1999) identified pertinent articles; references from these articles were identified to serve as additional resources. DISCUSSION: Verapamil is effective in inhibiting atrioventricular nodal conduction, thereby controlling ventricular rate in patients with atrial fibrillation/flutter and terminating paroxysmal supraventricular tachycardia. However, hypotension may be caused by the negative inotropic and vasodilating effects of verapamil. In vitro and animal data suggest that calcium pretreatment may minimize the effects of verapamil on cardiac output and blood pressure. Case reports suggest that intravenous calcium may be useful for both prevention and reversal of the hemodynamic effects of verapamil. A number of small clinical trials have been performed, suggesting that calcium administered prior to intravenous verapamil results in a decreased incidence of hypotension. The most common adverse effect of intravenous calcium is flushing. CONCLUSIONS: Calcium pretreatment prior to intravenous calcium-channel blocker administration should be considered in patients in whom further reductions in blood pressure may precipitate hypoperfusion or worsen underlying cardiovascular status. A dose of calcium gluconate 1 g (ionized calcium 90 mg) administered over three minutes is recommended for preventing or lessening the hypotensive effect of verapamil without affecting the antiarrhythmic effects of verapamil. PMID- 10852092 TI - The role of carnitine supplementation during valproic acid therapy. AB - OBJECTIVE: To review the pathophysiology and significance of valproic acid induced carnitine deficiency; to present and evaluate the literature pertaining to carnitine supplementation in pediatric patients receiving valproic acid; and to present the consensus guidelines for carnitine supplementation during valproic acid therapy. DATA SOURCES: A MEDLINE search (1966-December 1998) restricted to English-language literature, using MeSH headings of carnitine and valproic acid, was conducted to identify clinically relevant articles. Selected articles and references focusing on the pediatric population were included for review. DATA EXTRACTION: Study design, patient population, methods, and clinical outcomes were evaluated. DATA SYNTHESIS: Valproic acid, a widely used antiepileptic agent in the pediatric population, is limited by a 1/800 incidence of fatal hepatotoxicity in children under the age of two years. Carnitine is an essential amino acid necessary in beta-oxidation of fatty acids and energy production in cellular mitochondria. It has been hypothesized that valproic acid may induce a carnitine deficiency in children and cause nonspecific symptoms of deficiency, hepatotoxicity, and hyperammonemia. Relevant published case reports and trials studying this relationship are evaluated, and a consensus statement by the Pediatric Neurology Advisory Committee is reviewed. CONCLUSIONS: Despite the lack of prospective, randomized clinical trials documenting efficacy of carnitine supplementation in preventing valproic acid-induced hepatotoxicity, the few limited studies available have shown carnitine supplementation to result in subjective and objective improvements along with increases in carnitine serum concentrations in patients receiving valproic acid. The Pediatric Neurology Advisory Committee in 1996 provided more concrete indications on the role of carnitine in valproic acid therapy, such as valproic acid overdose and valproic acid-induced hepatotoxicity. Carnitine was strongly recommended for children at risk of developing a carnitine deficiency. Although carnitine has been well tolerated, future studies are needed to evaluate the efficacy of prophylactic carnitine supplementation for the prevention of hepatotoxicity. PMID- 10852093 TI - Pharmacogenetic screening for susceptibility to fetal malformations in women. AB - OBJECTIVE: To present a review of the literature and research on the pharmacogenetics of congenital defects, with a focus on the need for predictive maternal genotype assays. DATA SOURCE: MEDLINE searches (January 1985-January 1999), past reference reviews, and unpublished research. STUDY SELECTION: Review of relevant human, animal, and basic science studies. DATA EXTRACTION: Data on research on polymorphisms, genotyping, cytochrome P450 enzyme systems, epoxide hydrolase, folate metabolism, metabolism of anticonvulsant medications, molecular genetics of neural tube defects, variations in drug metabolism, and environmental exposures were evaluated. DATA SYNTHESIS: Data synthesis includes not only a review of the literature but suggests ways such data might be used to facilitate the development of maternal genotype assays, with the goal of preventing birth defects. CONCLUSIONS: Individuals vary in how they metabolize drugs and handle toxic environmental exposures. In an ideal pregnancy, there is no or limited exposure to medications and environmental agents. However, in women with chronic medical conditions such as heart disease and seizures, this is often not possible. Unfortunately, no techniques have been available to identify those at risk in this population. Gene polymorphisms for a specific enzyme may result in an absence or reduction in the level of enzyme activity or in no change at all, with little effect on the structure/function of the gene product(s); they are not associated with clinical phenotypes in either the mother or the fetus. Other polymorphisms may be only markers. Thus, developing genotyping assays for women that are predictive of phenotype expression in the fetus is the key to screening for polymorphisms. As more mutations are identified and clinical, pharmacologic, biologic, and pharmacokinetic relationships are established, using these polymorphisms to develop a genotyping assay for women may become a clinical reality, possibly leading to preventive prepregnancy or prenatal treatment that may play an increasingly effective role in maternal care. PMID- 10852094 TI - Urinary incontinence. AB - BACKGROUND: One of the more prevalent conditions associated with aging is urinary incontinence (UI), which may affect up to 55% of women and 34% of men older than 65 years. As a result of increasing longevity in developed nations, the proportion of UI-susceptible individuals continues to grow, presenting clinical and economic challenges to healthcare providers. OBJECTIVE: To assist the clinician in making informed decisions regarding UI, provide information on the wider ramifications of the disease, and provide a comprehensive overview of the condition. DATA SOURCES: MEDLINE (1966-December 1998) was searched for relevant publications using the following search terms: UI, UI in the elderly, treatment of UI, oxybutynin, flavoxate, vasopressin, quality of life in UI, and economic impact of UI. DATA SYNTHESIS: Key articles relating to the etiology, diagnosis, classification, economic burden, quality of life, and treatment of UI were retrieved, and this information formed the basis of the review. CONCLUSIONS: Although UI can be controlled relatively well with existing therapies, only about 50% of affected patients may actually seek care. There are a variety of therapeutic options available for the treatment of UI, although pharmacologic intervention is presently a relatively minor component of overall care; this suggests that effective drug therapy might play a more significant role in the future. The economic burden associated with the care of the incontinent patient is substantial, and in the US the direct medical cost of the disease was estimated at $25.5 billion in 1995. The disease also has a large impact on the individual UI patient, negatively affecting many parameters normally associated with a tolerable health-related quality of life. PMID- 10852095 TI - Treatment of listeriosis. AB - OBJECTIVE: To review the most currently accepted treatment options for the treatment of listeriosis. DATA SOURCES: Clinical literature was accessed through MEDLINE (1966-October 1999). Key search terms included Listeria monocytogenes, food-borne illness, penicillins, fluoroquinolones, cephalosporins, and vancomycin. DATA SYNTHESIS: Listeriosis is mainly a food-borne illness caused by L. monocytogenes; people most prone to the disease are pregnant women, newborns, elderly, and those with HIV or other diseases compromising immunity. Listeria infections are associated with a high mortality rate, and thus effective antibiotic treatment is essential. Although a variety of antibiotics have activity against the organism, ampicillin alone or in combination with gentamicin remains the treatment of choice. Some patients may require alternative therapies due to allergies or certain disease states. Second-line agents for these cases include trimethoprim/sulfamethoxazole, erythromycin, vancomycin, and the fluoroquinolones. Cephalosporins are not active against Listeria. CONCLUSIONS: Ampicillin is currently the drug of choice for treating L. monocytogenes infections. Many antibiotics have been shown to be effective and are used as second-line agents. However, further study is required for some of the most recently introduced antibiotics, such as the fluoroquinolones, to determine their place in the treatment of Listeria infections. PMID- 10852096 TI - Clozapine-induced hypersalivation. AB - OBJECTIVE: To review underlying pathophysiology and possible treatments for clozapine-induced hypersalivation. DATA SOURCES: Primary literature was accessed through MEDLINE (1966-May 1999). Key search terms included clozapine, hypersalivation, sialorrhea, and treatment. DATA SYNTHESIS: Hypersalivation occurs in up to 54% of patients receiving clozapine. An evaluation of studies and case reports focusing on management of clozapine-induced hypersalivation was conducted. CONCLUSIONS: It is unclear whether clozapine increases salivation through its muscarinic M4 receptor activation and/or blockade of alpha2 adrenoceptors, or by causing a distortion in swallowing reflex. Treatment options include chewing gum, reducing the dosage of clozapine, or prescribing pharmacologic agents such as anticholinergics or alpha2-adrenoceptor agonists. PMID- 10852097 TI - Potential interaction between celecoxib and warfarin. PMID- 10852098 TI - Possible neuroleptic malignant syndrome associated with olanzapine. PMID- 10852099 TI - Relapsing vivax malaria, chloroquine toxicity, anxiety, or alcohol misuse? PMID- 10852100 TI - Relationship between structure and drug-induced parkinsonism. PMID- 10852101 TI - Molecular mechanisms of innate immunity. AB - All species require a rapid, systemic reply to pathogens in their environment. This response is known as the innate immune response and is characterized by de novo synthesis of mediators that directly or indirectly through phagocytosis remove and kill the pathogen. Innate immune responses have been preserved throughout evolution and have been studied in detail in organisms from the fruit fly Drosophila melanogaster to humans. In my laboratory, studies performed during the past 25 yr have focused on defining the molecular basis of innate immune responses to microbial pathogens. Specifically, we have used bacterial endotoxin (lipopolysaccharide) as a model stimulus to define how the innate immune system recognizes products of microbial pathogens and initiates responses to remove and/or kill such organisms. Such studies also serve as models to understand more fully the mechanisms underlying a serious human disease known as septic shock. This article discusses septic shock and its relationship to innate immunity. PMID- 10852102 TI - Elucidating tumor necrosis factor signaling pathway using a functional gene identification approach. AB - Functional identification of genes is an efficient way to study many biological processes in lower eukaryotes. However, an effective approach in mammalian cells is still under development. We designed a functional gene identification procedure and applied it in a study of tumor necrosis factor (TNF)-induced cell killing. This procedure employed a specially designed retroviral vector that allows random truncation of genes, efficiently selecting clones in which a gene was disrupted and quickly identifying disrupted genes. We have identified several novel genes by a preliminary test of this approach and confirmed by reconstitution that the genes we identified are required for TNF cytotoxicity in L929 cells. Because of the efficient identification of these components in TNF induced cell killing, we have already been able to outline the killing pathway of TNF in L929 cells. Application of this method could be widespread because it can be used in studying any cellular responses if a specific selection assay can be set up. PMID- 10852103 TI - A role for epithelial gammadelta T cells in tissue repair. AB - My colleagues and I have a long-term interest in interactions between intraepithelial gammadelta T cells and neighboring epithelial cells. We have focused our studies on interactions in the thymus, skin, and intestine, and are investigating the development, specificity, and function of these gammadelta T cells. Our results have defined unique properties of these cells and support a specialized role for epithelial gammadelta T cells in immune surveillance, wound repair, inflammation, and protection from malignancy. PMID- 10852104 TI - Regulation of T cell development in the thymus. AB - My colleagues and I are studying the regulation of T cell differentiation and lineage commitment in the thymus. Most recently, we have focused on the role of the mitogen-activated protein kinase (MAPK) signaling pathway in these processes and, in particular, the temporal pattern of activation of this pathway and its effect on downstream gene targets. Our data suggest that thymocyte differentiation to either the CD4 or CD8 lineages requires sustained low-level signaling via MAPK, although the latter requires a weaker signal. We have proposed that both the amplitude and kinetics of MAPK signaling may be one aspect of the link between T cell receptor affinity and cell fate. In addition, we have shown that the Egr family of transcription factors is induced as a consequence of MAPK activation during positive selection in the thymus, and we are taking several approaches to identify other genes that are involved in regulating this developmental process. PMID- 10852105 TI - Molecular signaling mechanisms of cell migration and invasion. AB - The ability of immune cells to migrate and invade the extracellular matrix (ECM) is a central process involved in immunologic surveillance as well as inflammation. We have shown that interaction of cells with adhesive proteins or growth factors (chemokines) present in the ECM control cell migration/invasion through activation of mitogen-activated protein kinases ERK1 and ERK2 and molecular coupling of the adapter proteins p130CAS and c-CrkII. During cell migration, ERK and CAS/Crk coupling operate as distinct signaling pathways that facilitate actin-myosin motor assembly and actin membrane ruffles, respectively. Furthermore, activation of these signaling pathways protects cells from apoptosis during invasion of the ECM, which is necessary as migratory cells colonize foreign sites in the body. PMID- 10852106 TI - Relative roles of somatic and Darwinian evolution in shaping the antibody response. AB - The need for a highly specific system of recognition in immunity has resulted in the evolution of several somatic mechanisms such as V(D)J recombination, to diversify the repertoire of B cells. Therefore, repertoire diversification is the driving force for the cells that constitute the bulk of the response to unpredictable pathogens, the B2 naive B cells. Predictability of antigen, on the other hand, has played a major role in shaping the neonatal repertoire, in which evolution to recognize commonly encountered pathogens has driven the germline sequence of several VH segments that are used frequently in the neonatal repertoire. A third population, the memory B cell population, is generated to respond to a known pathogen, but predictability of the pathogen is not acquired until after a first exposure. Therefore, it is somatic evolution in germinal centers that drives the generation of high-affinity memory B cells. PMID- 10852107 TI - Rho GTPase signaling in inflammation and transformation. AB - Intracellular Rho GTPases provide an important regulatory mechanism to connect cell-surface-generated signals with the nucleus. By cycling between the active (guanosine 5'-triphosphate [GTP]) and inactive (guanosine 5'-diphosphate) state, these GTP-binding proteins control cellular functions ranging from dynamic actin remodeling and activation of transcription factors to cell-cycle progression and cellular transformation. Their contribution to these very diverse processes makes them an essential part of cell movement, growth, and apoptosis. Upstream regulatory mechanisms, as well as a variety of downstream effector molecules, enable Rho GTPases to act in a specific, orchestrated manner, dictating cellular responses. In this article, I review my laboratory's work centering on the goal of determining how specificity in intracellular signaling is achieved and identifying molecular mechanisms of Rho GTPase-mediated processes in innate immune and transformed cells. PMID- 10852108 TI - Genetic studies in systemic autoimmunity and aging. AB - The etiopathogenesis of systemic lupus erythematosus remains an enigma that will probably not be solved until the genetic basis for susceptibility is defined. Through genomewide searches, we have provided a foundation for this by identifying and characterizing loci predisposing to specific disease traits in four major lupus-susceptible mouse strains. Further ongoing work that includes the study of interval-specific congenic lines and precise mapping of loci should lead to identification of the corresponding genes and elucidation of processes critical for disease pathogenesis. Another important area of investigation is the study of cell-cycle and apoptosis genes in systemic autoimmunity and aging. Based on earlier work, we proposed that the characteristic overexpansion of memory phenotype cells in these conditions may be owing to replicative senescence. Understanding the molecular mechanisms that regulate the generation of these cells may permit selective manipulations to control this process. Other areas of investigation that we are actively engaged in are the role of T cell receptor repertoire in disease and the definition of cellular genes affected by infection with human immunodeficiency virus. PMID- 10852109 TI - Function of intestinal gammadelta T cells. AB - Mucosal tissues including the intestine, lung, reproductive tract, and skin form the major interfaces between the outside and internal milieus. Facing the outside is an epithelial cell layer, the epithelium, built on a vascular connective surface. In addition to performing specialized functions, mucosal tissues are sites where immune, epithelial, and neuronal cell types act in concert to maintain tissue integrity and fight invading pathogens. This article presents the latest findings from my laboratory describing a novel protective function for the intestinal intraepithelial gammadelta T cells (gammadelta intraepithelial lymphocytes). PMID- 10852110 TI - Interleukin-8 and its receptor CXCR2 in atherosclerosis. AB - The participation of inflammatory cells in atherosclerosis is a well-known process that involves numerous molecules including chemotactic cytokines (chemokines) for their entry into the vessel wall. Although the C-C chemokine monocyte chemoattractant protein-1 and its receptor, CCR2, have been implicated in atherosclerosis, the role of the classic C-X-C chemokine, interleukin-8 (KC/growth-related oncogene alpha in mice) and its receptor CXCR2 has not been studied in the pathogenesis of atherosclerosis. Our research has shown that CXCR2 is strongly expressed on macrophages (Mphi) in atherosclerotic lesion. This CXCR2 expression is proatherogenic in that CXCR2 deficiency significantly reduces the progression of advanced atherosclerosis in mice. Although the mechanism still needs to be worked out, it appears that CXCR2 expression on lesion Mphi is essential for these cells to be retained in the lesion. PMID- 10852111 TI - Regulation of cell function by Rho family GTPases. AB - Rho GTPases act as molecular switches to control many basic cellular activities that are also critical to the specialized functions of phagocytic leukocytes. Our laboratory has studied the regulation of Rho GTPase function, how these GTPases interact with specific effectors to modulate cell function, and how these events are coordinated in the stimulated cell. Areas of major interest include NADPH oxidase regulation by Rac2, Rac- and Cdc42-mediated control of the actin-myosin cytoskeleton via p21-activated kinase (PAK), and modulation of the apoptotic program by Rho GTPases and PAK. PMID- 10852112 TI - Regulation of development and function of memory CD4 subsets. AB - Immunologic memory refers to the dramatic response to previously encountered antigen (Ag) that is largely controlled by CD4 T cells. Understanding how CD4 memory is regulated is essential for exploiting the immune system to protect against disease and to dampen immunopathology in allergic responses and autoimmunity. Using defined adoptive-transfer models, we are studying parameters that affect differentiation of memory CD4 cells in vivo and have found that a complex interplay of T cell receptor signaling, costimulation, and cytokines can determine the extent of memory development and the balance of Th1 and Th2 memory subsets. On challenge, memory CD4 cells localize in sites of Ag exposure and develop into effectors that regulate memory responses. We are investigating the roles of adhesion molecules, cytokines, and chemokines in the selective recruitment of CD4 memory subsets to address mechanisms by which memory T cells provide long-lasting immunity and, in our recent studies, to determine how memory CD4 cells contribute to the development of autoimmune diabetes. PMID- 10852113 TI - Focal inflammation in the brain: role in Alzheimer's disease. AB - We hypothesize that amyloid (Abeta) peptide-containing neuritic plaques in the brains of patients with Alzheimer's disease represent chronic inflammatory foci mediated by the actions of the complement system and proinflammatory cytokines. In support of this, in vitro studies show that the (Abeta) peptide is a potent complement activator and that such complement activation leads to the formation of covalent (Abeta)-C3 activation fragment complexes, the generation of the chemokine-like C5a complement activation peptide, and the formation of the proinflammatory C5b-9 complex in functionally active form able to insert into neuronal cell membranes. Other studies show that C5a, together with (Abeta), synergistically augments the release of proinflammatory cytokines from human monocytes. These studies, which provide in vitro support for the hypothesis, are being pursued in an animal model of Alzheimer's disease. PMID- 10852114 TI - Participation of innate and acquired immunity in atherosclerosis. AB - Coronary artery disease, the major manifestation of atherosclerosis, is the leading cause of death in the Western world. However, the pathogenesis of atherosclerosis is still poorly understood. Controversy exists regarding the participation of innate immunity involving macrophages and natural killer (NK) cells vs antigen-specific acquired immunity involving lymphocytes. Macrophages predominate in atherosclerotic lesions. NK cells, although smaller in number, are present as well. Furthermore, T lymphocytes that participate in acquired immunity are frequently observed in lesions and can modulate lesion progression. By using mouse models of hyperlipidemia, our laboratory is addressing in vivo the participation of both innate inflammatory responses and acquired immune responses in atherosclerosis. PMID- 10852115 TI - Allograft rejection results from a failed attempt by the immune system to protect foreign tissue. AB - The focus of our research is to understand the immune response to foreign tissue. We believe that a dichotomy exists within the immune response to an allograft such that part of the response is dedicated to the protection of the graft. Nevertheless, in a dominantly graft-aggressive environment, rejection typically ensues. In this article, we describe models that have been set up to test directly the ability of potentially protective aspects of the immune response to prevent allograft rejection. We discuss our data in the context of a growing body of exciting and often controversial literature. PMID- 10852116 TI - Human antibodies in cancer and autoimmune disease. AB - In recent years, a number of novel human autoantigens and tumor-associated antigens have been identified using patient sera. Several of these antigens have been used as diagnostic markers, but defining their role in disease pathogenesis has been hampered by the lack of cloned human antibodies and antigens. Focusing on the solid cancers of the breast and colon and on autoimmune hematologic diseases, we are studying the role of human antibodies in disease pathogenesis. We have generated several human monoclonal autoimmune and cancer-associated antibodies, using antibody phage display technology, and have identified, cloned, and expressed their corresponding (novel) antigens. Using the monoclonal human antibodies as probes, we are elucidating the processes that lead to the generation of these antibodies and their possible pathogenic or protective effect. These studies may lead to the development of reagents for diagnosis and therapeutic intervention of these important diseases. PMID- 10852117 TI - Factors that influence formation of B cell repertoire. AB - V, D, and J gene segments rearrange at different frequencies in vivo. In my laboratory, we are interested in determining the reasons for this unequal rearrangement of V genes during B cell development, and also in gaining insights into the mechanisms that control recombination. Every V, D, and J gene segment is flanked on its recombining side(s) by a recombination signal sequence (RSS), which is composed of a conserved heptamer and nonamer, separated by a spacer of conserved length. In this article, we summarize data showing that in many cases the RSS can account for differences in recombination frequencies observed in vivo. The approach that we use is to determine the frequency of initial rearrangement of the V genes in vivo. The RSSs of two V genes are then placed into a competition recombination substrate to determine the relative frequency with which the two RSSs recombine. In one example, we have shown that a single base pair polymorphism in the RSS of a Vkappa gene may play a major role in susceptibility to Haemophilus influenzae type b infection. PMID- 10852118 TI - Role of chemokines in inflammation and immunoregulation. AB - Chemokines are first noted for their ability to attract and activate leukocytes, as well as their potential role as mediators of inflammation. However, emerging data have shown that various chemokines may exert other biologic effects both inside and outside the immune system. Inducible chemokines participate primarily in inflammatory responses and comprise the bulk of the chemokine family. Constitutive chemokines are expressed primarily in secondary lymphoid organs and some nonlymphoid organs, where they play a major role in lymphocyte homing. Studies expanding to areas beyond inflammatory leukocyte recruitment will likely give us a more complete picture of chemokine function, its regulation in lymphoid and nonlymphoid tissues, and ways of utilizing endogenous chemokines to intervene with immune and inflammatory reactions. PMID- 10852119 TI - Human immunodeficiency virus type 1 hijacks host cyclophilin A for its attachment to target cells. AB - Our laboratory has identified a new facet of human immunodeficiency virus type 1 (HIV-1) entry. We demonstrated that the incorporation of host cyclophilin A (CypA) into nascent viruses is absolutely required for HIV-1 attachment to target cells. Although CypA is initially incorporated into the interior of the virus, we found that during maturation CypA relocates to the viral surface. Our work indicates that exposed CypA mediates HIV-1 attachment to target cells via heparans. We believe that this interaction between CypA and heparan represents the initial step in HIV-1 entry. PMID- 10852120 TI - Principles of homeostasis in governing virus activation and latency. AB - The goal of our work is to understand, from the molecular to the organismal level, the principles that drive and sustain lifelong infection by viruses. These infectious agents live in a dynamic equilibrium (homeostasis) with their hosts in which both immune and nonimmune pathways contribute to viral homeostasis. Disruption of these pathways can have dramatic consequences on pathogenesis. Immune responses to infection provide a vital countermeasure by the host but are nonsterilizing. They effect an essential and primary control mechanism for viral growth. Essential nonimmune pathways for effecting control of a viral life cycle relate to the obligate dependency of the virus on its host. For these reasons, we view infections as a highly dynamic interplay that takes place between the pathogen and host. This, in many cases, leads to the establishment of an incurable lifelong infection that remains benign but can become life threatening once key homeostatic pathways are disrupted. We discuss these issues in the context of our studies using cytomegalovirus as a clinically relevant pathogen. PMID- 10852121 TI - T cell receptor binding kinetics and special role of Valpha in T cell development and activation. AB - The kinetics of the interaction between T cell receptor (TCR) and major histocompatibility complex (MHC) has an important role in determining thymocyte positive and -negative selection in the thymus, as well as in T cell activation. The alpha chain of the TCR is the major player in determining how the TCR fits onto the MHC ligand, and thus has a major role in determining whether a T cell develops as class I or class II restricted. In this article, we summarize recent data from our laboratory and others on the role of polymorphism in the Valpha combining site in determining MHC class restriction, and on kinetic parameters in thymocyte selection. PMID- 10852122 TI - Role of BMK1 in regulation of growth factor-induced cellular responses. AB - Big mitogen-activated protein kinase (MAPK) 1 (BMK1), also known as ERK5, is a recently identified member of the mammalian MAPK family. Cellular stimulation of BMK1 is induced in response to growth factors, oxidative stress, and hyperosmolar conditions. Specific members of the myocyte enhancer factor 2 family of transcription factors that regulate growth factor-induced early gene expression have been identified as direct downstream targets of BMK1 activity. Recent studies have shown that growth factors of the epidermal growth factor family mediate the sequential activation of a kinase cascade consisting of MAPK kinase kinase 3, MAPK kinase 5, and BMK1. Most importantly, the activation of this signal transduction pathway has been shown to be required for growth factor mediated cell proliferation and cell-cycle progression. Collectively, these studies establish BMK1 as an important regulator of growth factor-induced cellular responses. PMID- 10852123 TI - Immune regulation: susceptibility and resistance to autoimmunity. AB - Susceptibility and resistance to autoimmune diabetes appear to be regulated at several levels. Whereas the effector functions of autoimmunity are dependent on the antigen-specific responses of autoreactive T cells, the development of tissue specific autoimmunity appears to be additionally dependent on tissue-specific factors. Thus, the recruitment of T lymphocytes into the vicinity of islet tissue is driven by the local production of chemokines with a pattern unique to islet tissue. The accumulation of lymphocytes triggers events that lead to the local development of new lymphoid tissue, enhancing the presentation of islet-specific antigens to the immune system. In the presence of additional genetic factors allowing persistent immune responses, autoreactivity is permitted to progress until end-stage autoimmune diabetes develops. My laboratory is studying the influence of tissue-specific factors and the genetic regulation of lymphocyte reactivity. An integrated understanding of these effects should lead to a more physiologic approach to prevention and treatment of autoimmune diabetes. PMID- 10852124 TI - Lipopolysaccharide induction of gene expression in human monocytic cells. AB - Monocytes become activated at sites of inflammation and contribute to the pathology of many diseases, including septic shock. In these cells, induction of genes expressing various inflammatory mediators, such as cytokines, chemokines, and growth factors, is regulated by nuclear factor-kappaB (NF-kappaB)/Rel transcription factors. Recent studies have identified components of the signal transduction pathways leading to the activation of NF-kappaB/Rel proteins. Inhibition of these signaling pathways provides a novel therapeutic approach to prevent inducible gene expression in monocytes. PMID- 10852125 TI - Virus and target cell evolution in human immunodeficiency virus type 1 infection. AB - Human immunodeficiency virus (HIV) infection leads to a prolonged struggle between a rapidly evolving viral population and a potent immune response. In the vast majority of infected individuals, the virus wins this struggle. In my laboratory, we focus on understanding both the viral and immune factors that contribute to this outcome. The results of our studies and those of many others indicate that HIV can escape a potent immune response by a combination of mechanisms including rapid mutation, shedding of decoy antigens, modulation of host major histocompatibility complex, and destruction of cytotoxic T lymphocytes. The target cells for viral infection change as the virus evolves to use different chemokine coreceptors for entry. The initial targets are activated and resting memory T cells that express both CD4 and CCR5, but both naive and memory CD4 T cells are targeted by viruses capable of using CXCR4 for entry, and macrophages become the primary target cells when most CD4 T cells are depleted. Compelling evidence is emerging that the availability of target cells for infection is as limiting for the spread of virus as the immune response. PMID- 10852126 TI - Role of B cell antigen receptor in regulation of V(D)J recombination and cell survival. AB - B lymphocytes learn through the interaction of the B cell receptor with antigens in the context of B cell developmental stage and environmental cues. B cells can respond by proliferation and antibody secretion, programmed cell death, or modification of the antibody genes themselves through secondary immunoglobulin gene rearrangements or somatic point mutation. A critical learning process is that of self/nonself-discrimination. We have shown that one potent mechanism for immune self-tolerance in B cells is ongoing antibody light chain gene rearrangements, which can result in "receptor editing" that changes antigen receptor specificity. This process appears to be developmentally regulated, because it is confined to cells at an immature stage of development. Cells at later stages of development can be tolerized by apoptosis, but probably not by receptor editing. PMID- 10852127 TI - Antibodies in human infectious disease. AB - Investigation of human antibody responses to viral pathogens at the molecular level is revealing novel aspects of the interplay of viruses with the humoral immune system. In viral infection, at least two types of human antibody responses exist: a response to mature envelope on virions that is neutralizing and a response to immature forms of envelope (viral debris) that is not. Many pathogens have, to varying degrees, evolved envelopes to minimize antibody responses against epitopes exposed on the virion. In this article, we review recent studies on human immunodeficiency virus type 1, Ebola virus, and respiratory syncytial virus. Prion diseases are diseases of protein conformation. We have generated a large panel of antibodies recognizing the cellular prion protein (PrP(c)), some of which also react with the abnormally folded infectious prion protein (PrP(Sc)). These antibodies are being used to gain insight into both the molecular events leading to the formation of infectious PrP and the physiologic role played by PrP in normal and prion-infected cells. PMID- 10852128 TI - Targeted cytokines for cancer immunotherapy. AB - Targeting of cytokines into the tumor microenvironment using antibody-cytokine fusion proteins, called immunocytokines, represents a novel approach in cancer immunotherapy. This article summarizes therapeutic efficacy and immune mechanisms involved in targeting interleukin-2 (IL-2) to neuroectodermal tumors using ganglioside GD2-specific antibody-IL-2 fusion protein (ch14.18-IL-2). Treatment of established melanoma metastases with ch14.18-IL-2 resulted in eradication of disease followed by a vaccination effect protecting mice from lethal challenges with wild-type tumor calls. In a syngeneic neuroblastoma model, targeted IL-2 was effective in the amplification of a weak memory immune response previously induced by IL-12 gene therapy using an engineered linear version of this heterodimeric cytokine. These findings show that targeted IL-2 may provide an effective tool in cancer immunotherapy and establish the missing link between T cell-mediated vaccination and objective clinical responses. PMID- 10852129 TI - Diverse functions of protease receptor tissue factor in inflammation and metastasis. AB - Accumulating evidence suggests that protease receptors and their cognate protease ligands play important roles in cell-signaling events that regulate cell adhesion and migration in inflammation as well as tumor invasion and metastasis. Tissue factor (TF), the cell surface receptor for the serine protease VIIa and the initiator of the coagulation pathways, supports metastatic implantation by activating extracellular, protease-dependent signaling pathways and by intracellular links to the actin cytoskeleton. The adhesion of TF-expressing tumor cells can be mediated by interactions of the receptor-protease complex with specific matrix-associated inhibitors, suggesting a novel bridging mechanism by which proteases participate in migratory functions of cells. PMID- 10852130 TI - Focal adhesion kinase functions as a receptor-proximal signaling component required for directed cell migration. AB - In performing host-defense functions, cells of the immune system become activated by soluble chemokine signals and must migrate through endothelial cell or solid tissue barriers to reach sites of inflammation or infection. Regulated adhesive interactions of immune cells with endothelium, extracellular matrix components, and cells of solid organs are critical control points of the overall immune response. Both the soluble chemokine and cell adhesion receptor-mediated migration signals must converge on common intracellular targets to engage the cell migration machinery. In this article, we focus on the role of focal adhesion kinase (FAK) and its homolog Pyk2 as cytoplasmic mediators of motility events in multiple cell types. We introduce the overall domain structure of the FAK and Pyk2 nonreceptor protein tyrosine kinases (PTKs), highlight some of the signals that activate these PTKs, and detail the molecules that functionally interact and signal transduction pathways that may mediate cell migration responses. Emphasis is placed on the knowledge gained from studies using FAK-null cells as a model system to decipher the role of this PTK in promoting cell motility. PMID- 10852131 TI - Self-tolerance and the composition of T cell repertoire. AB - T cell recognition of self-major histocompatibility complex-peptide complexes dictates the composition of the T cell receptor repertoire. Research projects in our laboratory deal with the mechanisms that regulate the composition of the repertoire specific for self-antigens and the defects that can result in autoimmunity. Two different types of disease models are under investigation: juvenile (type I) diabetes and cancer. Both of these diseases are impacted by the presence of anti-self CD8 cells, yet in opposite ways. By understanding the mechanisms of peripheral tolerance and the reasons they fail in autoimmunity, we may learn how to prevent undesirable autoimmunity and how to encourage an autoimmune response when it is needed to eliminate tumor cells. PMID- 10852132 TI - The thymus and negative selection. AB - Self-tolerance induction is largely a reflection of negative selection (deletion) of autoreactive T cells in the thymus. Evidence is presented that negative selection occurs at a relatively late stage of thymocyte differentiation and affects a population of semimature HSA(hi) CD4+8- cells found in the medulla. Negative selection involves a number of cell-surface molecules on T cells, including Fas, CD28, CD5, and CD43. These molecules appear to act in consort, thereby ensuring that negative selection is highly efficient. PMID- 10852133 TI - Function and dysfunction of T cell receptor: structural studies. AB - The engagement of the T cell receptor (TCR) to its ligand, the major histocompatibility complex (MHC)-peptide complex, leads to T cell activation. The molecular mechanisms leading to this activation are still unknown. Dimerization or substantial conformational changes following TCR ligation have not been observed by classical biochemical methods or by X-ray crystallography of the TCR/MHC complex. However, most of these experiments have used reductionist approaches in which only MHC and TCR molecules were taken into account. In fact, the TCR is only one of many molecules forming the TCR complex (TCRC), and the interplay among the components of this larger complex have not been studied in depth. The reconstitution of a complete TCRC using recombinant molecules is our goal and will be the first step to new structural and functional studies. PMID- 10852134 TI - Role of self-major histocompatibility complex/peptide ligands in selection and maintenance of a diverse T cell repertoire. AB - Positive selection has long been thought to be a devise for producing a repertoire of T cells that can efficiently recognize foreign peptides in the context of self-major histocompatibility complex (MHC) molecules. However, in the light of recent evidence that long-term survival of mature T cells requires continuous contact with self-MHC molecules, the possibility for an additional role for positive selection has emerged: to generate a repertoire of T cells that can be maintained in the periphery through contact with self-MHC/peptide ligands. In support of this idea, our recent work suggests that positive selection is highly peptide specific and, more important, that mature T cells require extrathymic contact with the same MHC/peptide ligands that initially induced positive selection in the thymus in order for prolonged survival and to undergo homeostatic proliferation in response to T cell deficiency. PMID- 10852135 TI - Innate immune system recognition of microbial pathogens. AB - The long-term goal of our laboratory is to understand vertebrate host recognition of microbial pathogens. Although our work is primarily curiosity driven, it is certainly true that understanding how a host recognizes microbial pathogens should have some medical application. Probably more than 50,000 people die each year in the United States of septic shock or the systemic inflammatory response syndrome, and there is no good therapy for this problem. Understanding the molecular basis of its origin should suggest novel therapeutic approaches. PMID- 10852136 TI - Controlling mature CD4+ T cell responses. AB - Immune responses are by necessity highly regulated to achieve the appropriate balance of aggression and restraint. Among the many factors involved in maintaining this balance are the interactions between accessory molecule receptors expressed on T cells and their ligands on antigen-presenting cells. Our studies during the past several years have focused on defining how particular accessory molecule interactions influence the activation of naive CD4+ T cells and the subsequent development of effector function. In this article, we discuss our findings on the effects of distinct accessory molecules with particular attention to the unique roles of LFA-1 and CD28 during different phases of the naive CD4+ cell response. PMID- 10852137 TI - Etiology of autoimmunity. AB - Autoimmunity reflects the destruction of host tissue by the immune system. In my laboratory we are focused on understanding autoimmunity and the complex regulation of the autoimmune process. We believe that host defense mechanisms, which are normally activated to fight microorganisms, can malfunction and attack self. The role of cytokines and other host-elicited factors are a major focus of our group. PMID- 10852138 TI - Mechanotransduction and arterial smooth muscle cells: new insight into hypertension and atherosclerosis. AB - Vascular cells depend on multiple stimuli to maintain a biomechanically and biologically stable environment. Mechanical stresses contribute significantly to multiple cellular processes that regulate vascular structure and function. For example, fluid shear stresses control endothelial cell molecular responses. Less attention has focused on responses of the smooth muscle cell, the 'other' major vascular cell, to mechanical stimuli, in part because of the experimental difficulties in applying precisely controlled deformation. With the advent of new bioengineered devices, combined with modern technologies for studying molecular expression, we are beginning to understand how the smooth muscle cell responds to and controls the biomechanical environment. These studies will help us to understand vascular diseases where vascular mechanics plays a prominent role, such as hypertension, aneurysm formation and atherosclerotic plaque rupture. PMID- 10852139 TI - Awareness of time during sleep. AB - One cognitive ability that has been demonstrated in many people when asleep is the capacity to keep track of time sufficiently so that they can self-awaken when desired. Possible practical, physiological and evolutionary sources of this skill are outlined. Such findings help to establish the likely presence of significant cognitive ability when asleep. PMID- 10852140 TI - Effectiveness of individual lifestyle interventions in reducing cardiovascular disease and risk factors. AB - In order to assess the effectiveness of lifestyle interventions in reducing cardiovascular disease risk factors, morbidity and mortality among working-age adults, we undertook a systematic review of randomized controlled trials of various lifestyle interventions (diet, exercise, smoking cessation, alcohol intake reduction) in adults followed for 1 year or longer. Twenty-one single factor and 21 multifactorial interventions were analysed by outcome. Changes in cardiovascular morbidity and mortality and total mortality were considered as main outcomes. Changes in weight, total cholesterol, blood pressure, sodium excretion, smoking and alcohol consumption were also analysed, and numbers needed to treat were calculated for smoking, morbidity and mortality. In secondary prevention, both single and multifactorial lifestyle interventions were shown to reduce morbidity and mortality, and multifactorial approaches reduced cholesterol levels. Primary prevention was found to reduce risk factors efficiently, especially when the intervention is multifactorial. Effect sizes were heterogeneous with wide confidence intervals. Standardized ways of describing interventions, measuring their effects and reporting outcomes systematically would facilitate effect-size evaluations. Interventions should optimally be multifactorial and targeted at high-risk patients with multiple risk factors for cardiovascular disease. PMID- 10852141 TI - Poor prognosis of depression in elderly people: causes and actions. AB - Depression is the most common mental health problem of older people. It is a serious disorder which can lead to persistent suffering, increased mortality, from both suicide and general medical causes, and poorer overall health. Although presenting symptoms are similar in all age groups there are different aetiological pathways. In older people the waning effect of genetic predisposition to affective disorder may be replaced by subcortical brain abnormalities of presumed vascular aetiology. These may influence prognosis. Depression in later life is often under-diagnosed and under-treated; these two factors are the main hurdles to an improved prognosis. Antidepressant treatment should be tailored to the patient and works best when combined with psychological therapy, but the latter treatment modality is woefully neglected in later life psychiatry. Improvements in prognosis are unlikely to come from new revolutionary treatments but from vigorous treatment in the acute phase, continuation after recovery for at least 12-18 months and long-term maintenance treatment for those at high risk of recurrence. PMID- 10852142 TI - Prevention of delirium in hospitalized older patients: risk factors and targeted intervention strategies. AB - Delirium is a common, costly, and potentially devastating condition for hospitalized older patients. Delirium is a multifactorial syndrome, involving the inter-relationship between patient vulnerability, or predisposing factors at admission, and noxious insults or precipitating factors during hospitalization. Through a series of studies, we first identified significant predisposing factors for delirium, including vision impairment, severe illness, cognitive impairment, and dehydration. Subsequently, significant precipitating factors were identified, including physical restraint use, malnutrition, adding more than three drugs, bladder catheter use, and any iatrogenic event. Through targeting preventive strategies towards six identified risk factors in a controlled clinical trial, we were successful in the primary prevention of delirium. In 852 subjects, the incidence of delirium was significantly reduced in the intervention group compared with usual care (9.9% vs 15.0%, matched odds ratio: 0.60; 95% confidence interval: 0.39-0.92). The total number of days and episodes of delirium were also significantly reduced in the intervention group. Based on this work, evidence based recommendations for delirium prevention are proposed. While not all cases of delirium will be preventable with this approach, unifying medical and epidemiological approaches to delirium represents a key advance essential to reducing the high morbidity and mortality associated with delirium in the older population. PMID- 10852143 TI - Impact of transposable elements on the human genome. AB - Presence of transposable elements (TEs) in the human genome has profound effects on genome function, structure and evolution. TE mobility and inter-TE recombination are the origin of a large spectrum of mutations and genome reorganization leading to diseases. From the data provided by the Human Genome Project and from information on the detection and dynamics of TEs within and between species acquired during the last two decades, we now know that these elements are not only involved in mutagenesis but can also participate in many cellular functions including recombination, gene regulation, protein-coding RNA messages and, possibly, cellular stress response and centromere function. TEs also promote a general genome shuffling process that has been important for the evolution of several gene families and for the development of new regulatory pathways. PMID- 10852144 TI - Function of C-reactive protein. AB - C-reactive protein (CRP) is an ancient highly conserved molecule and a member of the pentraxin family of proteins. CRP is secreted by the liver in response to a variety of inflammatory cytokines. Levels of CRP increase very rapidly in response to trauma, inflammation, and infection and decrease just as rapidly with the resolution of the condition. Thus, the measurement of CRP is widely used to monitor various inflammatory states. CRP binds to damaged tissue, to nuclear antigens and to certain pathogenic organisms in a calcium-dependent manner. The function of CRP is felt to be related to its role in the innate immune system. Similar to immunoglobulin (Ig)G, it activates complement, binds to Fc receptors and acts as an opsonin for various pathogens. Interaction of CRP with Fc receptors leads to the generation of proinflammatory cytokines that enhance the inflammatory response. Unlike IgG, which specifically recognizes distinct antigenic epitopes, CRP recognizes altered self and foreign molecules based on pattern recognition. Thus, CRP is though to act as a surveillance molecule for altered self and certain pathogens. This recognition provides early defense and leads to a proinflammatory signal and activation of the humoural, adaptive immune system. PMID- 10852145 TI - Ligase chain reaction assay is clinically useful in the discrimination of smear positive pulmonary tuberculosis from atypical mycobacterioses. AB - We evaluated the usefulness of the ligase chain reaction (LCR) (Abbott LCx Mycobacterium tuberculosis assay) during the initial diagnosis of tuberculosis. LCx was carried out in parallel with conventional methods for the analysis of clinical samples. Out of 86 patients who were examined clinically, 53 were suspected of having pulmonary tuberculosis, eight had residual X-ray scars from previous tuberculosis and 25 served as asymptomatic controls. Ten bronchoscopy samples and 237 sputum samples were analysed by direct microscopy, culture and LCx. All 11 smear-positive and two of three smear-negative tuberculosis patients had at least one LCx-positive specimen. All samples that were both LCx- and smear positive were culture-positive for M. tuberculosis. The smear-positive samples from the five patients with atypical mycobacteriosis were LCx-negative. There were three false-positive results: one in a smear-negative sample from a patient with M. malmoense infection and two from two pneumonia patients. All samples from controls and patients with previous tuberculosis were LCx-negative. The sensitivity, specificity and the positive and negative predictive values of LCx in patient analysis were 92.9%, 95.8%, 81.3% and 98.6%, respectively. LCx assay of M. tuberculosis is useful in rapid confirmation of tuberculosis or atypical mycobacteriosis from a smear-positive sample and may aid in diagnosing smear negative tuberculosis. PMID- 10852146 TI - Introduction: Understanding and treating burnout in a changing culture. PMID- 10852147 TI - Burnout in teachers: shattered dreams of impeccable professional performance. AB - Burnout usually is conceptualized as a work-related syndrome stemming from the individual's perception of a significant gap between expectations of successful professional performance and an observed, far less satisfying reality. The article examines this perception as a discrepancy between expected and observed levels of the individual's professional self-efficacy. The teaching profession and its service providers--teachers--serve as a model to illustrate and support this examination. Self-reports of novice teachers' experiences in their first year of teaching are given, reflecting a world of shattered dreams of idealistic performance. Finally, a number of suggestions for programs and activities that have proven helpful in alleviating stress and burnout among teachers are described. PMID- 10852148 TI - Burnout and diabetes: reflections from working with educators and patients. AB - Patients with Type-2 diabetes present with a range of psychosocial symptoms that, in combination with social and organizational pressures, often serve to exacerbate the stress of diabetes educators and contribute to burnout. Some of the more salient sources of both patient and educator stress are elaborated upon, and the nature of parallel processes between these two groups is noted. A case illustration with a burned-out diabetes educator demonstrates how enhancing self understanding and achieving a greater sense of balance can reduce symptoms of burnout, depression, and anxiety. This article highlights the need for educators, and more broadly all health professionals, to develop self-management skills. PMID- 10852149 TI - Clergy burnout: an integrative approach. AB - Understanding how clergy, who begin their careers with high idealism, optimism, and compassion, burn out is difficult. One body of research suggests that clergy, among others, burn out because of the systems in which they work. From this perspective, burnout is the result of external systemic factors such as bureaucracy, poor administrative support, and difficult work conditions. The other body of research suggests that burnout is the result of intrapersonal factors such as high idealism, Type-A personality, narcissism, and perfectionism. It is our position that these two bodies of research are compatible, and that by integrating the Self psychology of Kohut with the general systems theory of Bowen, it becomes easier to understand burnout. Further, by integrating these two theories, principles for treatment become clearer. PMID- 10852150 TI - Treating career burnout: a psychodynamic existential perspective. AB - This article presents an approach for treating career burnout based on a psychodynamic existential perspective. Psychodynamic theory contributes the idea that people choose an occupation that enables them to replicate significant childhood experiences. Existential theory contributes the idea that people attempt to find existential significance through their work. It is suggested that when treating career burnout it is essential to address three questions: Why, psychodynamically, did this person choose this particular career, and how was it expected to provide existential significance? Why does this individual feel a sense of failure in the existential quest, and how is the sense of failure related to burnout? What changes need to take place for this individual to derive a sense of existential significance from work? A case illustration is presented that demonstrates the application of this approach. PMID- 10852151 TI - Preventing burnout in professionals and paraprofessionals who work with child abuse and neglect cases: a cognitive behavioral approach to supervision. AB - Professionals and paraprofessionals who treat children and families where child maltreatment has occurred are subject to many strains. This article focuses on the potential for burnout in such work. It discusses strategies in supervision to combat early manifestations of burnout and to prevent its full-blown occurrence. A cognitive-behavioral framework is used to help supervisors identify the sources of strain, the maladaptive, and inflexible assumptions regarding their own capacities as professionals and their own views of families that these strains may violate, and ways to work with supervisees to reduce the impact these violations have. It also addresses supervisors' own reactions to the high level of needs such families and children present and the strain on the supervisory relationship they produce. Institutionally based and systemic issues are highlighted. PMID- 10852152 TI - Prisoners of liberation: a psychoanalytic perspective on disenchantment and burnout among career women lawyers. AB - Using a psychoanalytic perspective, this article addresses the roots and treatment of disillusionment and incipient burnout in female corporate lawyers. It suggests that one of the primary issues that needs to be addressed in therapy with this group is the tendency to be self-punishing, a characteristic that often may be traced back to insensitive parenting. This formulation, combined with the penchant of law firms to regard as "good and expected" the inclination of their workers (especially women) to work without respite and with little regard for their own needs, places individuals at high risk for burnout. A case example is used to illustrate this phenomenon. It is concluded that whereas psychoanalytic treatment greatly may help overstressed professionals, society at large must address the values that foster the attitude of high career achievement at any cost. PMID- 10852153 TI - Treatment strategies for different types of teacher burnout. AB - Using teachers as a prototype, this article suggests that there are three types of burnout: "wearout," wherein an individual gives up, feeling depleted in confronting stress; "classic" burnout, wherein an individual works increasingly hard in the face of stress; and an "underchallenged" type, wherein an individual is faced not with excessive degrees of stress per se (e.g., overload), but rather with monotonous and unstimulating work conditions. The major arguments put forward are that: a) clinicians should avoid treating teacher burnout as if it were a single phenomenon, and instead tailor their treatment to the specific type of burnout manifested by their client; and b) these treatments, while embodying different elements, should be essentially integrative in nature. Psychoanalytic insight, cognitive restructuring, empathic concern, and stress-reduction techniques may all be necessary, albeit in different combinations, to treat successfully burnout of each type. PMID- 10852154 TI - CREST syndrome; a changing clinical significance. PMID- 10852155 TI - Arterial stenting and balloon angioplasty in renal artery stenosis. PMID- 10852156 TI - Phosphate as another major regulator of parathyroid function. PMID- 10852157 TI - Severe myopathy in patients with thyrotoxicosis. PMID- 10852158 TI - Rare complication of myositis in chronic graft-versus-host disease. PMID- 10852159 TI - Cardiac valve diseases and antiphospholipid syndrome. PMID- 10852160 TI - Vasculopathy in dermatomyositis. PMID- 10852161 TI - Antineutrophil cytoplasmic antibody-related vasculitis and liver diseases. PMID- 10852162 TI - Clinical, serological and genetic study in patients with CREST syndrome. AB - OBJECTIVE: To assess the clinical, serological and genetic features of Japanese patients with CREST syndrome. PATIENTS AND METHODS: Clinical features, autoantibodies and human histocompatibility leukocyte antigen (HLA) typing were studied in thirty patients with CREST syndrome, including 29 females and one male, with a mean age of 59.0 years (ranging from 40 to 76 years). RESULTS: Interstitial pneumonia on chest X-ray and renal involvement were rare. Mitral regurgitation and tricuspid regurgitation were present in 56.7% and 76.7%, respectively. Sjoren's syndrome (SS) and primary biliary cirrhosis (PBC) were highly associated, however the positivity of the marker antibodies to those syndromes, such as anti-SSA, anti-SSB, anti-mitochondrial (AMA) and anti-smooth muscle autoantibodies were less frequent than that of primary SS and PBC without the other autoimmune diseases. The histological findings of PBC were all early stages in Scheuer's classification. HLA-Cw6 were associated with CREST-PBC overlap syndrome (p<0.05). However the HLA antigen was not correlated with CREST syndrome, and the frequency of HLA-DR2 between CREST syndrome with or without PBC was significantly different (p<0.01). CONCLUSION: It was suggested that there was a genetic difference between CREST syndrome alone and CREST-PBC overlap syndrome and there were differences (the positivity of AMA and the severity of bile duct lesion) between PBC and CREST-PBC overlap syndrome. PMID- 10852163 TI - Reevaluation of laboratory parameters in relation to histological findings in primary and secondary Sjogren's syndrome. AB - OBJECTIVE: We reevaluated the diagnostic value of laboratory parameters in relation to histopathological findings in Sjogren's syndrome (SS) to clarify whether autoantibodies are useful diagnostic criteria for SS, and whether any laboratory data are useful in estimating the degree of salivary gland change. PATIENTS AND METHODS: Laboratory parameters and histopathological findings were analyzed in 96 patients examined by labial biopsy. RESULTS: The percentage of cases with positive assays of rheumatoid factor and anti-SS-A/Ro antibodies was significantly higher in Definite SS. Patients with dense mononuclear cell infiltration of salivary tissues also had higher titers of rheumatoid factor. No useful laboratory parameters were found for the diagnosis of secondary SS. CONCLUSION: Rheumatoid factor and anti-SS-A/Ro antibodies are useful for the diagnosis of primary SS, and rheumatoid factor is also an indicator of the severity of salivary glandular damage. PMID- 10852164 TI - Treatment of renovascular hypertension using stent implantation in an elderly patient with NIDDM. AB - A 70-year-old man with NIDDM was diagnosed as having renovascular hypertension (RVH), based on a stenosis of the ostial portion of the left renal artery with markedly elevated plasma renin activity (PRA) in both the left renal vein and the peripheral blood, and positive captopril tests. After percutaneous transluminal renal angioplasty (PTRA), his blood pressure (BP) and PRA normalized. However, since restenosis occurred three months later, stent therapy was applied, and consequently BP and PRA normalized immediately after this procedure. During the one-year follow-up, side effects have not been noted. We propose that stent therapy may be feasible for ostial renal artery stenosis in elderly diabetic patients. PMID- 10852165 TI - Tertiary hyperparathyroidism in X-linked hypophosphatemic rickets. AB - We report a case of tertiary hyperparathyroidism in an X-linked familial hypophosphatemic rickets (XLH) patient under regular calcitriol and self-adjusted large doses of oral phosphate salt (2.4-3.6 g/day in 4-5 divided doses) according to his serum phosphate level. Tertiary hyperparathyroidism is an unusual complication of XLH patients during treatment. As there is growing evidence that a high phosphate diet may induce hyperplasia of the parathyroid glands, it is important to avoid the stimulation of the parathyroid glands by high doses of phosphate administration in XLH patients. Serum calcium, phosphate, alkaline phosphatase, and also parathyroid hormone should be measured regularly in order to facilitate an early diagnosis of secondary hyperparathyroidism during the treatment of XLH patients, since this stage is reversible with calcitriol and reduced doses of phosphate salt. PMID- 10852166 TI - Hyperthyroidism presenting as dysphagia. AB - A 65-year-old man presented with hyperthyroidism associated with thyrotoxic dysphagia. Treatment with thiamazole improved his symptoms promptly. Although dysphagia is a rare manifestation of thyrotoxicosis, it should be emphasized that the possibility of hyperthyroidism must be discussed in unexplained dysphagia because it is readily treatable. PMID- 10852167 TI - Malignant insulinoma which expressed a unique creatine kinase isoenzyme: clinical value of arterial embolization as a palliative therapy. AB - A 76-year-old man with hypoglycemic coma was diagnosed as malignant insulinoma with multiple hepatic metastases. Embolization was done for two-thirds of the hepatic mass and it rapidly lowered the serum immunoreactive insulin. He was discharged without medication and has been free from hypoglycemia. After the embolization, the serum creatine kinase (CK) level increased transiently although there was no evidence of myocardial infarction. On electrophoresis, the CK activity showed an abnormal peak, suggesting mitochondrial CK. CK release after embolization has been reported in only a few cases with endocrine tumors, which might indicate some relationship between active energy metabolism and mitochondrial CK. PMID- 10852168 TI - Response to combined modality treatment in a five-year survivor of extensive small cell lung cancer with severe complications. AB - We present a rare case of a five-year survivor of small cell lung cancer with severe complications who responded to combined modality treatment. Prior to initial chemotherapy, he experienced severe complications including sepsis, pneumonia, ileus, and a performance status of 4. He was treated with an ileus tube and IVH, and was managed by mechanical ventilation for four days. After his general condition improved, he received combination chemotherapy of carboplatin, with the target area under the plasma concentration versus the time curve (AUC) of 5 mg x min/ml day 1, and etoposide (80 mg/m2) on days 1, 2, 3 for four courses, and complete remission (CR) was obtained. Six months later, systemic relapse occurred, but he achieved complete remission again with nine courses of CODE (cisplatin, vincristine, adriamycin, and etoposide) chemotherapy and sequential chest radiotherapy. Five years after the initial chemotherapy, the patient is alive and disease free. PMID- 10852169 TI - Myositis as a manifestation of chronic graft-versus-host disease. AB - We report a 22-year-old man who had myositis in the course of chronic graft versus-host disease after bone marrow transplantation for acute monocytic leukemia. The distribution of muscular involvement was different from idiopathic polymyositis. Muscular atrophy and weakness were noted in the distal muscles as well as in the proximal muscles of the upper extremities but there was little weakness in the proximal muscles of the lower extremities. However, histological and immunohistochemical study of the biceps brachii muscle showed findings similar to those of idiopathic polymyositis. It was suggested that myositis can be a manifestation of chronic GVHD caused by a cellular immune reaction by donor T cells. PMID- 10852170 TI - Acute rhombencephalitis: neuroimaging evidence. AB - Following a high fever, a healthy woman became comatose within a few days. Severe cerebellar symptoms appeared when she regained consciousness. The brain MRIs revealed abnormal signal intensity of the cerebellar cortex and brainstem gray matter, however, no abnormalities were revealed in the cerebral hemispheres. Acute inflammation due to direct viral or autoimmune involvement of the cerebellar and brainstem gray matter was a likely explanation and thus acute cerebellitis may in fact be a rhombencephalitis. Among the previous reports of acute cerebellar ataxia, this is perhaps one of the most profoundly affected cases and appears important for the understanding of the target of this particular form of encephalitis. PMID- 10852171 TI - Myasthenia gravis with membranous nephropathy, successfully treated with extended total thymectomy. AB - A 46-year-old woman showed proteinuria and hematuria after left blepharoptosis, and revealed a histopathology of membranous nephropathy (MN) at renal biopsy. She was diagnosed as having myasthenia gravis (MG) because of a positive edrophonium test and anti-acetylcholine receptor (AchR) antibodies in serum. We found a decrease in anti-AchR antibodies after extended total thymectomy, in parallel with an improvement in both urinary findings and myasthenic symptoms. In this case, MG preceded MN and the thymectomy was effective for both diseases, suggesting that the thymus might play an important role in the pathogenesis of MN. PMID- 10852172 TI - Diabetic ketoacidosis associated with Guillain-Barre syndrome with autonomic dysfunction. AB - A 37-year-old woman was admitted in a comatose state, after exhibiting fever and diarrhea. Diabetic ketoacidosis was diagnosed due to an increased blood glucose level (672 mg/dl), metabolic acidosis, and positive urinary ketone bodies. On the fifth hospital day, despite recovery from the critical state of ketoacidosis, the patient suffered from dysphagia, hypesthesia and motor weakness, followed by respiratory failure. Cerebrospinal fluid analysis was suggestive of Guillain Barre syndrome (GBS). Autonomic dysfunction was manifested as tachycardia and mild hypertension in the acute stage. Marked orthostatic hypotension persisted long after paresis was improved, indicating an atypical clinical course of GBS. PMID- 10852173 TI - An adult case of mumps brainstem encephalitis. AB - We present an adult case of mumps brainstem encephalitis. He was successfully treated with steroid pulse therapy and recovered completely except for persistent dysuria. He had not been vaccinated and had no history of acute mumps infection. We consider that encephalitis in this case was caused by a reversible autoimmune process triggered by mumps infection. We emphasize the usefulness of pulse therapy for the treatment of some cases of mumps brainstem encephalitis in addition to the importance of mumps vaccination to prevent such a severe complication as encephalitis. PMID- 10852174 TI - Guillain-Barre syndrome following hand-foot-and-mouth disease. AB - We describe a patient who developed Guillain-Barre syndrome (GBS) following hand foot-and-mouth disease (HFMD) which is known to be caused by enterovirus infection. A 35-year-old man developed acute paraparesis and dysesthesia in the four limbs following typical skin eruption of HFMD. Electrophysiologic studies showed peripheral nerve demyelination predominant in the distal terminals. HFMD is a rare cause of meningitis, encephalitis, and polio-like myelitis, but GBS following HFMD has never been described. PMID- 10852175 TI - Overt congestive heart failure with mitral and aortic regurgitation due to antiphospholipid syndrome in a patient with systemic lupus erythematosus. AB - A 51-year-old woman with overt congestive heart failure with pleural and pericardial effusion was treated with furosemide and nifedipine, leading to improvement in her condition and a decrease in effusions. An echocardiography demonstrated mitral and aortic regurgitation with mitral valve prolapse, which caused the congestive heart failure. Since leukocytopenia and lymphocytopenia with arthralgia could be observed, serological investigations were performed. She was diagnosed as having systemic lupus erythematosus (SLE) with antiphospholipid syndrome, and started on a treatment of prednisolone and aspirin. Based on the treatment, the pleural and pericardial effusion went into complete remission, indicating that the serositis related to SLE had overlapped the heart failure. Since there was no evidence of any other diseases that could be responsible for the valvular lesions, we concluded that they were due to antiphospholipid syndrome. The administration of prednisolone had no significant effect on valvular morphology or function as demonstrated by echocardiography. When patients with valvular disease are seen, a valvulopathy related to antiphospholipid syndrome should be considered as part of the differential diagnosis. PMID- 10852176 TI - Dermatomyositis with splenic and renal infarctions during corticosteroid therapy. AB - A 60-year-old woman was admitted to our hospital with complaints of muscle weakness and erythema on her extremities. Gottron's sign, heliotrope rash, elevation of serum myogenic enzymes, electromyography and magnetic resonance imaging findings established a diagnosis of dermatomyositis (DM). She was treated with 60 mg of daily prednisolone. One week later, she suddenly developed splenic and renal infarctions, which were considered to have resulted from vasculopathy associated with DM. Cyclophosphamide and anticoagulants along with increasing the dosage of corticosteroid were effective. This is the first report describing splenic and renal infarctions in a patient with adult-onset DM. PMID- 10852177 TI - Microscopic polyangiitis that presented liver dysfunction prior to noted renal manifestations. AB - In microscopic polyangiitis (MPA), renal manifestations are very common as first symptoms. Here, we report a case of MPA which presented liver dysfunction prior to noted renal manifestations. A 58-year-old woman was hospitalized because of a fever for 8 weeks. A laboratory examination revealed marked elevation of alkaline phosphatase and gamma-glutamyl transpeptidase, while blood urea nitrogen and creatinine levels remained normal. Although apparent renal dysfunction developed in this case soon after hospitalization, physicians should be aware of the variety of clinical manifestations in MPA. Moreover, antineutrophil cytoplasmic autoantibodies were found to be helpful for diagnosing MPA. PMID- 10852178 TI - Humanizing neonatal care. PMID- 10852179 TI - Clinical audit in the management of children with epilepsy. PMID- 10852180 TI - Brain development, head circumference and medication. PMID- 10852181 TI - Body mass index and child obesity: are we nearing a definition? PMID- 10852182 TI - Randomized trial comparing natural and synthetic surfactant: increased infection rate after natural surfactant? PMID- 10852183 TI - Bone turnover in children with vitamin D deficiency rickets before and during treatment. AB - Biochemical markers of bone formation [alkaline phosphatase, osteocalcin, and carboxyterminal propeptide of type I procollagen (PICP)] and bone resorption [cross-linked carboxyterminal telopeptide of type I collagen (ICTP) and cross linked N-telopeptides of type I collagen (NTX)] were measured in 14 children aged 8.5-10.5 mo with vitamin D deficiency rickets before and longitudinally during vitamin D treatment (3000-4000 IU/daily). Forty-four healthy children aged 8-10.5 mo were enrolled as sex- and age-matched controls. Before treatment, serum levels of alkaline phosphatase, PICP, and ICTP, and urinary excretion values of NTX were significantly higher, and serum osteocalcin levels significantly lower than controls (31.4 +/- 3.5 microkat/L and 9.8 +/- 2.9 microkat/L, p < 0.001; 1025 +/- 89 microg/L and 952 +/- 97.4 microg/L, p < 0.02; 15.6 +/- 2.6 microg/L and 14.2 +/- 1.3 microg/L, p < 0.01; 370.7 +/- 109.4 nmol BCE and 201.8 +/- 69.2 nmol BCE, p < 0.001: 17.6 +/- 9.1 microg/L and 22.5 +/- 7.6 microg/L, p < 0.05, respectively). During treatment, serum alkaline phosphatase levels progressively declined in association with the radiographic healing of the skeletal lesions. Serum levels of osteocalcin, PICP, and ICTP, and urinary excretion values of NTX showed a transient but significant (p < 0.05 to p < 0.001) increase in comparison with baseline values during the first 2-4 wk of treatment, and decreased slowly thereafter. They were within the mean +/- 2 SD of controls before the recovery of the skeletal lesions. CONCLUSIONS: These findings suggest that children with vitamin D deficiency rickets have increased bone turnover before and during the first weeks of treatment. Alkaline phosphatase is a more reliable marker than osteocalcin, PICP, ICTP and NTX for diagnosing and monitoring these patients. PMID- 10852184 TI - Leukocyte adhesiveness/aggregation test (LAAT) to discriminate between viral and bacterial infections in children. AB - OBJECTIVES: We previously noted that white blood cells (WBC) have increased adhesive properties during bacterial infections. Here, we aim to explore the possibility of using the different adhesive properties of WBC as a means of differentiating between viral and bacterial infections, a common problem in paediatrics. METHODS: The adhesive properties of WBC in the peripheral blood of 25 children with documented bacterial infections, 15 with documented viral infections and 36 with probable viral infections, were studied by means of a leukocyte adhesiveness/aggregation slide test (LAAT). The results of the LAAT were compared with those of the other acute phase reactants, namely WBC, differential count and erythrocyte sedimentation rate (ESR), which were taken in the same blood sample in each patient. RESULTS: The sensitivity, specificity and positive predictive value were 92%, 96%, and 92%, respectively for the LAAT; 83%, 87% and 80% for the ESR; 56%, 78% and 56% for the white blood cell count; and 54%, 74% and 50% for the differential count. CONCLUSIONS: The presence of bacterial infections in children can be tested using a simple slide test to reveal the increased state of leukocyte adhesiveness/aggregation in the peripheral blood. The LAAT is a reliable, rapid and inexpensive test, and it can be a useful laboratory tool for the paediatrician treating a child with acute febrile illness. PMID- 10852185 TI - Development of atopic sensitization and allergic diseases in early childhood. AB - Atopic diseases and atopic sensitization were studied from birth up to 2 y in a population-based prospective study. Physical examinations were done at 6, 12 and 24 mo, including skin-prick tests and blood samples, for the determination of serum IgE level and circulating IgE antibodies to food and inhalant allergens. In addition, questionnaire surveys were done about clinical symptoms of allergy, infections and the home environment. At 2 y the prevalences of definite and probable atopic diseases were 17% and 13%, respectively. The cumulative incidence of allergy was 25%. Atopic dermatitis was the main manifestation of atopy, with a peak prevalence of 15% at 2 y. The prevalence of positive skin-prick tests was constant at all ages, i.e. 7% at 6, 12 and 24 mo. Positive skin-prick tests against inhaled allergens were more than twice as common at 24 than at 12 mo, while sensitivity to food decreased. In contrast, circulating IgE antibodies to egg became more common with age, reaching 20% at 2 y. CONCLUSIONS: The present study indicates that manifestations of allergic diseases and incidence of sensitization to foods and inhalants are equally common in Estonian and Scandinavian children during the first 2 y of life, despite a much lower prevalence among older children and adults in Estonia. PMID- 10852186 TI - Growth and neurodevelopmental outcome of very low birthweight infants with necrotizing enterocolitis. AB - The aim of the study was to assess the effect of necrotizing enterocolitis (NEC) on neurodevelopmental outcome and growth. Neurodevelopmental outcome of 20 out of 22 suriviving very low birthweight infants (VLBW) diagnosed with NEC between 1992 and 1996 was compared with 40 control infants matched for gestational age and year of admission. Follow-up studies were performed at 12 and 20 mo of corrected age. The German revision of the Griffiths' scales was used for development assessment. Neurodevelopment was significantly delayed in infants with NEC at 12 mo (median general developmental quotient: 90.0 vs 97.8; p = 0.04) and 20 mo (86.4 vs 97.7; p = 0.004) of age. Somatic growth did not differ between infants with and without NEC. Fifty-five percent of infants suffering from NEC but only 22.5% of the infants without NEC were severely retarded (developmental quotient < -2 SD of a control group of healthy newborns) at 20 mo of corrected age. CONCLUSION: Preterm infants developing NEC are at risk for neurodevelopmental impairment and need close neurodevelopmental follow-up for the first years of life. PMID- 10852187 TI - Association of prenatal phenobarbital and phenytoin exposure with small head size at birth and with learning problems. AB - Small head size has been observed in prenatally anticonvulsant-exposed neonates. In infancy, cognitive impairments were revealed. It is presently unknown whether these impairments are permanent or disappear after puberty. We studied the link between the prenatal influence of anticonvulsants on brain development and cognitive functioning in adulthood: a retrospective study on head size and a follow-up assessing cognitive capacities among adults who had been included in the retrospective study. The retrospective study comprised 172 exposed and 168 control neonates, matched with respect to age, sex and their mothers' age. Prenatally phenobarbital + phenytoin-exposed neonates had a significantly smaller occipitofrontal circumference (OFC) than prenatally phenobarbital-monotherapy exposed and control neonates (mean difference of 0.7 cm). In the follow-up, no difference in cognitive functioning was found between the exposed and the control groups. Most of the prenatally anticonvulsant-exposed subjects had normal intellectual capacity. However, 12% of the exposed subjects versus 1% of the controls had persistent learning problems. In addition, more of the exposed subjects were mentally retarded. There was no clear relationship between learning problems and small OFC, maternal epilepsy or unfavourable family climate. CONCLUSIONS: We conclude that the combination of phenobarbital + phenytoin affects the fetal OFC. The smaller OFC does not seem to be related to cognitive functioning in adulthood, but learning problems and mental retardation proved to be more prevalent among exposed subjects. Phenobarbital and phenytoin may therefore affect cognitive capacity but only in infants who are susceptible to this particular influence of the drugs. PMID- 10852188 TI - Long-term prospective study on the natural history of Wolff-Parkinson-White syndrome detected during a heart screening program at school. AB - In the period 1985 to 1993, a total of 802 school-aged children (284 first graders and 518 seventh-graders) were referred to our hospital for further evaluation of electrocardiographic abnormalities. Among them, 57 (male 24 and female 33) children were confirmed as having Wolff-Parkinson-White (WPW) syndrome based on the findings of 12-lead surface electrocardiograms (ECG). According to Lindsay's criteria, the locations of the accessory pathways were as follows: Left lateral in 10 (18%), left-posterior in 2 (4%), right-free-wall in 28 (49%), anterior-septum in 13 (23%) and posterior-septum in 3 (5%). One 12-y-old girl had multiple accessory pathways. Six patients had associated diseases: Ebstein's anomaly in 4, epilepsy in 1 and mental retardation with scoliosis in 1. Follow-up periods ranged from 2.0 to 13.0 y (mean +/- SD: 8.0 +/- 3.3 y) for 23 first graders with WPW syndrome, and from 2.0 to 13.0y (mean +/- SD: 7.3 +/- 4.2y) for 34 seventh-graders, respectively. Initially, 5 children had at least one episode of supraventricular tachycardia (SVT) by history and 6 children developed SVT during the follow-up. One girl with multiple accessory pathways and recurrent SVT required long-term drug therapy. CONCLUSIONS: The outcome of children with WPW syndrome detected by a heart screening program at school was favorable. Our 8 y follow-up of 57 children with WPW syndrome will serve as additional information concerning the indication of radio-frequency catheter ablation therapy for WPW syndrome in children. PMID- 10852189 TI - Morphometry of the head of the caudate nucleus in patients with velocardiofacial syndrome (del 22q11.2). AB - Velocardiofacial syndrome (VCFS) is a chromosomal anomaly syndrome characterized by multiple congenital malformations, including cleft palate and cardiac anomalies. Many patients have attention-deficit (hyperactivity) disorders (AD[H]D) in childhood and schizophrenia in adulthood. We reviewed cranial magnetic resonance imaging (MRI) scans with particular attention to the head of the caudate nucleus and found that in control subjects, the head of the caudate was larger on the left than on the right, whereas VCFS patients showed a reversed right > left pattern or no significant asymmetry. A similar right > left asymmetry or a lack of this asymmetry has been reported in patients with AD(H)D. CONCLUSIONS: These results suggest a common pathophysiological mechanism of behavioural and cognitive problems in patients with AD(H)D and those with VCFS. PMID- 10852190 TI - Exclusively breastfed, low birthweight term infants do not need supplemental water. AB - Breast milk intake, urine volume and urine-specific gravity (USG) of exclusively breastfed, low birthweight (LBW) term male infants in Honduras were measured during 8-h periods at 2 (n = 59) and 8 (n = 68) wk of age. Ambient temperature was 22-36 degrees C and relative humidity was 37-86%. Maximum USG ranged from 1.001 to 1.012, all within normal limits. CONCLUSIONS: We conclude that supplemental water is not required for exclusively breastfed, LBW term infants, even in hot conditions. PMID- 10852191 TI - Evaluation of protein S-100 serum concentrations in healthy newborns and seven newborns with perinatal acidosis. AB - We measured Protein S-100 serum levels in 66 healthy newborns during the first week of life and in 7 newborns with perinatal acidosis. Normal values (n = 66) constantly ranged between 0.66 and 3.33 microg/l (2.5 and 97.5 percentiles) during the evaluation period. CONCLUSIONS: Newborns with signs of hypoxic ischaemic encephalopathy (HIE) after perinatal acidosis showed elevated Protein S 100 serum levels, whereas newborns without these signs had normal concentrations. S-100 might thus be a marker of central nervous system damage in newborns. Key words: Asphyxia, brain damage, neonate PMID- 10852192 TI - Randomized trial comparing natural and synthetic surfactant: increased infection rate after natural surfactant? AB - The efficacy of a natural porcine surfactant and a synthetic surfactant were compared in a randomized trial. In three neonatal intensive care units, 228 neonates with respiratory distress and a ratio of arterial to alveolar partial pressure of oxygen <0.22 were randomly assigned to receive either Curosurf 100 mgkg-1 or Exosurf Neonatal 5 ml.kg-1. After Curosurf, the fraction of inspired oxygen was lower from 15 min (0.45 +/- 0.22 vs 0.70 +/- 0.22, p = 0.0001) to 6 h (0.48 +/- 0.26 vs 0.64 +/- 0.23, p = 0.0001) and the mean airway pressure was lower at 1 h (8.3 +/- 3.2 mm H20 vs 9.4 +/- 3.1 mm H20, p = 0.01). Thereafter the respiratory parameters were similar. The duration of mechanical ventilation (median 6 vs 5 d) and the duration of oxygen supplementation (median 5 vs 4 d) were similar for Curosurf and Exosurf. After Curosurf, C-reactive protein value over 40 mg l-1 occurred in 45% (vs 12%; RR 3.62, 95%CI 2.12-6.17, p = 0.001), leukopenia in 52% (vs 28%; RR 1.85, 95% CI 1.31-2.61, p = 0.001) and bacteraemia in 11% (vs 4%; RR 3.17, 95% CI 1.05-9.52, p < 0.05). We conclude that when given as rescue therapy Curosurf had no advantage compared with Exosurf in addition to the more effective initial response. Curosurf may increase the risk of infection. PMID- 10852193 TI - Dexamethasone treatment and fluid balance in preterm infants at risk for chronic lung disease. AB - The influence of dexamethasone on diuresis in preterm infants has not been well studied. We examined 15 preterm infants at risk for chronic lung disease with gestational ages ranging from 26 to 29 wk (median 27.6 wk) and birthweights ranging from 700 to 1485 g (median 965 g). Urine output, blood glucose, serum urea, serum creatinine, serum sodium and serum potassium, as well as systolic, diastolic and mean arterial pressure were measured on the day before, and on 4 consecutive days after starting treatment with dexamethasone (0.25 mg kg-1 i.v., twice daily). We found an increase of diuresis of 30 ml kg -1 d-1, 48-96 h after starting dexamethasone treatment. This coincided with a gradual but significant increase of serum urea levels and arterial pressure. During the study period, fluid and protein intake remained constant. Blood glucose and serum creatinine levels did not change. Our findings suggest that the increased urine output following dexamethasone treatment might be caused by two factors: (1) pressure diuresis induced by the increase of arterial pressure and (2) an increase of the osmolar load to the kidney due to an increase of serum urea. CONCLUSIONS: This study demonstrates that a significant increase of diuresis occurs in preterm infants, 48-96 h after starting dexamethasone. A careful monitoring of fluid balance during the first days of dexamethasone treatment is recommended. PMID- 10852194 TI - Infants of women with severe early pre-eclampsia: the effect of absent end diastolic umbilical artery doppler flow velocities on neurodevelopmental outcome. AB - Umbilical artery Doppler flow velocity waveform studies were performed over a period of 4 y on 242 women with severe pre-eclampsia before 34 wk gestation. Sixty-eight (28%) had absent end-diastolic umbilical artery Doppler flow velocities. One hundred and ninety-three infants survived to hospital discharge and were followed at 6-monthly intervals until 48 mo of age. The mean corrected developmental quotient was 94 +/- 8 at 24 mo of age and 87 +/- 9 at 48 mo. Ninety two percent of the infants had a developmental quotient of >80 at 24 mo and 72% at 48 mo of age. This decline is thought to be due to the impact of social circumstances. There were no differences between the developmental quotients of the infants with normal and those with absent end-diastolic umbilical artery Doppler flow velocities at either 24 or 48 mo of age. At 24 mo of age, infants with absent end-diastolic umbilical artery Doppler flow velocities scored lower in the Performance subscale test (p = 0.03). The developmental quotients of infants from poorer socioeconomic backgrounds were significantly lower than those living in more privileged circumstances. At 48 mo, 153 (97%) of the children presented with normal gross motor development. Four infants had cerebral palsy. No differences were noted in the motor outcomes between the infants of women with normal umbilical artery waveforms and those with absent end-diastolic umbilical artery Doppler flow velocities. PMID- 10852195 TI - False alarms in very low birthweight infants: comparison between three intensive care monitoring systems. AB - Monitor alarms are a major burden on both patients and staff in intensive care units. We compared alarm rates from three different monitor systems (Hewlett Packard (HP), Kontron Instruments (KI), Marquette-Hellige (MH)) in a tertiary neonatal intensive care unit. Monitors were used in random order on three consecutive days over 8 h each in 16 preterm infants (median gestational age at birth 29 wk (range 24-34), age at study 18 d (8-53), weight at study 1,160g (595 1,430)). Alarms were classified as true or false using flow sheets based on continuous observation of both the patient and related parameters. There was one alarm every 9 min of monitoring. The median number of true alarms did not differ significantly between systems, being 28 per 8 h (range 9-87) for HP, 26 (3-81) for KI, and 30 (5-135) for MH. The median number of false alarms differed widely, with the HP system generating 32 (7-77) such alarms per 8 h, compared to 8 (0-19) for KI and 15 (2-32) for MH (p < 0.01 HP vs KI and MH, p < 0.05 KI vs MH). These differences between systems were mainly due to differences in pulse oximeter and transcutaneous PO2 monitor alarm rates. CONCLUSIONS: In conclusion, this study shows marked differences between both parameters and manufacturers in the frequency with which false alarms occur. It may provide a basis from which reductions in alarm rates can be sought. PMID- 10852196 TI - Altering the NICU and measuring infants' responses. AB - The aim of the study was to measure the impact of a designated Quiet period on the NICU environment and its influence on the infants' physiological and movement responses. The study group comprised 10 preterm infants on assisted ventilation (mean gestational age 28.7 wk (range 24-32 wk), mean birthweight 1,322 g (range 600-2,060 g), mean age 5.2 d). The environment in which the infants were nursed was altered in terms of reduced light, noise, staff activity and infant handling. The infants' heart rate, blood pressure, oxygen saturation and movement responses were recorded during this Quiet period and compared with a period of Normal activity. When the Quiet period was compared with the Normal period (median values), the NICU environment had significantly altered in terms of Light: Quiet period 3.0 Lux, Normal period 254.5 Lux (p < 0.01); Noise: Quiet period 54.0 dB, Normal period 58.0 dB (p < 0.01); Alarm events: Quiet period 491.5 sec, Normal period 1,180.5 sec (p < 0.01); Staff conversation: Quiet period 16.0 occasions per hour, Normal period 60.0 occasions per hour (p < 0.01); Staff activity: Quiet period 25.5 occasions per hour, Normal period 59.0 occasions per hour (p <0.01); Infant handling: Quiet period 0.0 events per hour, Normal period 4.5 events per hour (p < 0.01). Infants' diastolic blood pressure and mean arterial pressure: median reduction of 2 mmHg for both during the Quiet period (p < 0.05). Infants' movements: Quiet period 14.5 movements per hour, Normal period 84.0 movements per hour (p < 0.05). DISCUSSION: This study demonstrates that Quiet periods are feasible for infants undergoing neonatal intensive care. The NICU environment was altered significantly for light, noise, infant handling and staff activity for a specified time period. These changes were associated with a reduced median diastolic blood pressure and mean arterial pressure and a decrease in infant movements. PMID- 10852197 TI - Population-based body mass index reference values from Goteborg, sweden: birth to 18 years of age. AB - The body mass index or BMI (weight/height2) is a somewhat crude estimate of nutritional status. However, due to its simplicity and high correlation with total body fat, it has been the method of choice in both paediatric clinics and research over the years. Since BMI is not an equivalent measure of the percentage of body fat in different ethnic groups and in the two sexes, population-specific BMI reference data is needed. Several BMI reference values have been published for French, American, British and Hong Kong children in recent years. In Sweden, weight-for-age and height-for-age reference values, which were published in 1976, are still used as the current national growth reference values. Updated growth reference values are needed for assessing nutritional status due to the secular trend toward and increasing prevalence of childhood obesity. The aim of this study was to produce BMI reference values for Swedish children of paediatric age. The series came from a large Swedish population-based longitudinal growth study of 3650 full-term babies followed from birth to 18 y of age. The children in this data set were born in the early 1970s. The pattern and level of 50th centile BMI values presented here are quite similar to those of the Swedish cohort study in the 1950s. In comparison with the US BMI reference values, the Swedish values are much lower, especially for the higher centile values. CONCLUSION: The new Swedish BMI chart from our study may provide a useful tool for paediatricians to assess body fat, and consequently nutritional status, in Swedish children. PMID- 10852198 TI - Parental perception, worries and needs in children with epilepsy. AB - Despite advancement in medical care, prejudice and misunderstanding of epilepsy still exist. In this study, we investigate the problems faced by epileptic children, at home and in school, and make suggestions for improvement. Questionnaires were randomly distributed to parents of epileptic children attending normal and special classes (groups A and B, respectively). Return of questionnaires was anonymous. Ninety-one percent responded. Of the responders, 56 children were in Group A and 30 in Group B. Chronic and intractable epilepsy was more frequently observed in Group B than in Group A patients (47% vs 14%, p < 0.05). Main family concerns were seizures, school performance and side effects of medication. Half of the parents complained that their children were more restless and short-tempered. Only 43% of parents were aware that seizures were caused by abnormal brain discharges. Twenty percent thought swimming should be prohibited even if seizures could be controlled. Schools were informed of the disease by 84% of the families. Only 29% of parents knew the name and dose of the current medication. Information was considered adequate in 27% of patients. Drug compliance was better in epileptic children with associated handicaps than in those without handicaps. Half of the parents requested more information about epilepsy and closer communication between teachers and physicians. CONCLUSIONS: To establish comprehensive care that satisfies the needs of epileptic children and their families, further training of medical specialists in epilepsy and enhancement of networks among relevant organizations are needed. PMID- 10852199 TI - Bodily pain, sleep problems and mental distress in schoolchildren. AB - It has been suggested that childhood pain could be the beginning of a career with chronic disabling pain. Bodily pain is frequent in children. We examined the association between self-reported bodily pain, mental distress and sleep problems in schoolchildren to test the following hypotheses: (i) that self-reported bodily pain is associated with mental distress and sleep problems, (ii) that the association is dependent on the localization of pain, and (iii) that the association increases with number of painful areas. Eighty-six percent of the pupils (569) in the 4th form (mean age 10.5 y), 7th form (mean age 13.5 y) and 9th form (mean age 15.5 y) from all the schools in a local community answered a questionnaire about self-esteem, body-image, physical activity and bodily pain. We found a strong association between the reporting of pain, mental distress and sleep problems. Pain in the knees was the only problem reported more frequently by boys than by girls, and knee pain did not show the same association with mental distress and sleep problems as pain from other regions. CONCLUSIONS: A possible cause-effect relationship between pain, mental distress and sleep problems is discussed, and the possibility that all the complaints are the simultaneous signs of a multisymptom syndrome is introduced. PMID- 10852200 TI - Education of staff--a key factor for a safe environment in day care. AB - In order to create a safe environment in day-care settings, an understanding of factors within the organization of day care, factors which influence safety, is essential. Day-care directors in 83 daycare centres completed a mail-in survey that contained questions about professional experience, the day-care centre's organization of child safety measures and a battery of questions designed to evaluate the directors' perceptions and beliefs about child safety. The day-care directors also carried out a safety inspection at their centre. The results were analysed using the multivariate logistic regression technique. The existence of a continuing plan for continued staff education in child safety was shown to be the strongest predictor of few safety hazards in day-care centres. The day-care directors' perceptions and beliefs about injury prevention were of less importance. This study indicates that in order to promote safety in day-care settings, an on-going plan for continued staff education in child safety should be a matter of routine. The introduction of such a plan should be the concern of the individual day-care directors, policy-makers and managers at the local and national level, and health professionals working in this field. PMID- 10852201 TI - Relationship between acute respiratory infection and malnutrition in children under 5 years of age. AB - Data from a population-based survey, conducted in the northeast of Brazil, were used to examine the association between acute respiratory infection (ARI) and malnutrition in children under 5 y of age. Current and past malnutrition were associated with acute lower respiratory infection (ALRI), even after adjusting for potential confounders (odds ratio: 2.03; 95% confidence interval: 1.202.43). Decreasing malnutrition along with timely and proper treatment of ARI may improve children's health in developing countries. PMID- 10852202 TI - Structural hair shaft abnormalities in hypomelanosis of ito and other ectodermal dysplasias. AB - Hair samples from patients with different ectodermal dysplasias; hypohidrotic ectodermal dysplasia, pachyonychia congenita, tricho-dento-osseous syndrome, tricho-rhino-phalangeal syndrome, and hypomelanosis of Ito were investigated using a scanning electron microscope. The hairs of the patients showed different structural abnormalities; twisted hairs, longitudinal grooves, trichorrhexis nodosa as well as variations in the hair caliber. Hair shaft abnormalities, as in our patients with tricho-dento-osseous syndrome, and hypomelanosis of Ito have so far not been described. PMID- 10852203 TI - Rotavirus gastroenteritis possibly causing reye syndrome. PMID- 10852204 TI - A case of salmonella enterocolitis with paradoxical hypertension. PMID- 10852205 TI - Platelet-specific hemophagocytosis in a patient with juvenile dermatomyositis. PMID- 10852206 TI - Haemorrhagic shock and encephalopathy syndrome in four Turkish children. PMID- 10852207 TI - Gender differences and critical medical illness. PMID- 10852208 TI - Mismatch negativity (MMN) reveals sound grouping in the human brain. AB - To investigate a part of the structure of the memory trace, auditory event related potentials (ERPs) were recorded from reading subjects while they were presented with two different stimulus-series simultaneously. A clear mismatch negativity (MMN) was obtained from each series, when the stimulus sequence consisted of a high-frequency series and a low-frequency series. Moreover, the MMN showed independent elicitation within each series. However, if the frequency range of one series overlapped with that of the other series, the amplitude of the MMN was prominently reduced, suggesting that the two processing functions indexed by MMN coexisted simultaneously in the preattentive acoustic system and were produced by the respective grouping of high-frequency tones and low frequency tones. PMID- 10852209 TI - Increase of GAP-43 expression following kainic acid injection into whisker barrel cortex. AB - Plasticity after microinjection of kainic acid (KA) into the adult rat whisker barrel cortex was investigated with immunohistochemical staining of phosphorylated growth-associated protein (GAP)-43. After mapping the barrel cortex with the technique of intrinsic signal optical imaging, a small volume of KA was injected into one barrel. Rats were sacrificed at 2 days, 3 days, 1 week, and 6 weeks after lesioning. GAP-43 staining demonstrated intense immunoreactivity (IR) at the injected barrel which spread to the inter-barrel septa and the surrounding barrels. Elevated IR of GAP-43 was visible 2 days after KA injection, and increased gradually at least 6 weeks following the lesion. This model has the possibility of offering a simple and reliable tool for studying cortical plasticity. PMID- 10852210 TI - A new member of acid-sensing ion channels from pituitary gland. AB - Acid-sensing ion channels (ASICs) constitute a branch of the super-gene family of amiloride-sensitive sodium channels. So far five different ASICs have been cloned from mammalian tissues. They are activated by a drop of extracellular pH but differ with respect to effective agonist concentration, desensitization and mRNA expression pattern. Here we report cloning of ASIC4, a new protein showing about 45% identity to other ASICs. ASIC4 is 97% identical between rat and human and shows strongest expression in pituitary gland. Moreover, we detected expression throughout the brain, in spinal cord, and inner ear. ASIC4 cannot be activated by a drop of extracellular pH in Xenopus oocytes, suggesting association with other subunits or activation by a ligand different from protons. Our results suggest a role for ASICs also in endocrine glands. PMID- 10852211 TI - Differences in working memory involvement in analytical and creative tasks: an ERP study. AB - If, as suggested, creative (insight) problem solving is less systematic and employs less planning than analytical problem solving, the former requires substantially less working memory (WM) than the latter. Subjects simultaneously solved problems and counted auditory stimuli (concurrent WM task), in response to which ERPs were recorded. Counting disrupted analytical, but not creative performance. Peak and time-window average P300 were more frontal during analytical problem solving as compared to insight or counting tones only (control). A PCA extracted two factors in the P3 range, one frontal and one broad left-lateralized, which distinguished analytical from creative problem solving. The findings indicate distinct processing pathways for the two types of tasks with more WM involvement in analytical tasks. PMID- 10852212 TI - Neuromodulation of the prefrontal cortex during sleep: a microdialysis study in rats. AB - To test the hypothesis that biogenic amines of the prefrontal cortex are involved in state-dependent cortical and behavioural activation, changes in extracellular levels of serotonin (5-HT), dopamine (DA), and noradrenaline (NA) were determined during the sleep-wake cycle in freely moving rats using microdialysis probes with parallel EEG recording. Serotonin gradually increased up to 450% during wakefulness (W) as compared to slow wave sleep (SWS), before decreasing toward stable levels during the next episode of SWS. Dopamine and its metabolite homovanillic acid (HVA) were reduced during W as compared to SWS. Although contradictory with the generally admitted enhancement of DA activity related to vigilance, this may be due to the particular role of DA neurons in the prefrontal cortex. However, DA and HVA showed dramatic changes announcing the transition between SWS and W. During paradoxical sleep (PS), DA and 5-HT showed complex changes, the direction of which depended on whether PS was followed by SWS or W. Biogenic amines of the prefrontal cortex are probably involved in cortical and behavioural activation. PMID- 10852213 TI - Phosphorylation of MAP kinase and CREB in mouse cortex and amygdala during taste aversion learning. AB - Mice were subjected to a taste aversion conditioning procedure in which they drank water, familiar saccharin, or novel saccharin, followed by injection of either NaCl or the emetic agent LiCl. Immunohistochemistry was used to localize phosphorylated MAP kinase (ppERK) and phosphoCREB (pCREB) in the brain shortly after conditioning. An examination of insular cortex and amygdala revealed specific phosphorylation of MAP kinase by novel saccharin and LiCl, and CREB by LiCl. Pairing of novel saccharin with LiCl was the only stimulus sufficient to increase ppERK and pCREB immunoreactivity in the lateral amygdala, suggesting this as a site of CS-US convergence. ppERK immunoreactivity was localized to both nuclear and non-nuclear compartments, suggesting multiple functional roles of MAP kinase during learning. PMID- 10852214 TI - Evidence that DHPG-induced nociception depends on glutamate release from primary afferent C-fibres. AB - We examined whether enhanced glutamate release contributes to the expression of persistent spontaneous nociceptive behaviours (SNBs) in rats induced by intrathecal (i.t.) administration of the selective group I mGluR agonist, (RS) 3,5-dihydroxyphenylglycine ((RS)-DHPG). Pretreatment with drugs that have been shown to inhibit glutamate release, including a group II metabotropic glutamate receptor (mGluR) agonist (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate ((2R,4R) APDC), a group III mGluR agonist L-2-amino-4-phosphonobutyrate (L-AP4), or the use-dependent sodium channel blockers 3,5-diamino-6-(2,3-diclorophenyl)-1,2,4 triazine (lamotrigine) and 2-amino-6-trifluoromethoxybenzothiazole (riluzole), produced dose-dependent reductions in (RS)-DHPG-induced SNBs. We have also shown that incubation of rat lumbar spinal cord slices with (RS)-DHPG potentiates 4 aminopyridine-evoked (4-AP) release of glutamate. Furthermore, we found that destruction of unmyelinated primary afferent C-fibres by neonatal capsaicin treatment significantly reduced (RS)-DHPG-induced SNBs in adult rats. Together, these results suggest that (RS)-DHPG-induced nociception is dependent on spinal glutamate release, probably from primary afferent C-fibres. PMID- 10852215 TI - Functional dissociation of the early and late portions of human K-complexes. AB - We analyzed K-complexes (KCs) evoked during sleep stage II by the subject's own name and by other names. KCs were composed either of four consecutive waves (full KCs, N2-P3-N3-P4) or of the two first components only (N2-P3). The amplitude of the late phase of KCs (N3-P4) was identical to all stimuli; conversely, own names enhanced selectively the N2-P3 waves, whether they were or not part of a full KC. Two independent phenomena appear to coexist during a full KC, one being connected to the physical characteristics of the stimulus (N3-P4) and the other to its intrinsic significance. This latter may appear either within a full KC or in isolation, and in this case it is reminiscent of the N200-P300 complex observed in wakefulness. PMID- 10852216 TI - The effects of electromagnetic field emitted by GSM phones on working memory. AB - The influence of pulsed radiofrequency (RF) electromagnetic fields of digital GSM mobile phones on working memory in healthy subjects were studied. Memory load was varied from 0 to 3 items in an n-back task. Each subject was tested twice within a single session, with and without the RF exposure (902MHz, 217Hz). The RF field speeded up response times when the memory load was three items but no effects of RF were observed with lower loads. The results suggest that RF fields have a measurable effect on human cognitive performance and encourage further studies on the interactions of RF fields with brain function. PMID- 10852217 TI - Dynamic size-change of hand peripersonal space following tool use. AB - Humans and monkeys share similar sensory integrated processing of tactile and peri-hand visual inputs for coding peripersonal space surrounding the hand. In monkeys, tool use is known to induce a transient elongation of hand-centred peripersonal space along the tool axis. Here we report evidence that, also in humans, the use of a tool can increase the spatial extent of the representation of peri-hand visual space to incorporate the tool. We investigated this phenomenon in patients with tactile extinction, by using a cross-modal paradigm well suited to reveal visual-tactile integration near patients' hand. In the present study cross-modal extinction was assessed far from patients' ipsilesional hand, at the distal edge of a hand-held rake. We found that cross-modal extinction was more severe after patients used the rake to retrieve distant objects with respect to a condition in which the rake was not used. This evidence of an expansion of peri-hand space lasted only a few minutes after tool use. By contrast, peri-hand space expansion was not observed when motor actions towards distant objects did not involve the tool. These findings show that visual peri hand space has important dynamic properties in humans; it can be expanded and contracted depending upon tool use. PMID- 10852218 TI - Expression of glial cell line-derived neurotrophic factor family of growth factors in peripheral nerve injury in rats. AB - The glial cell line-derived neurotrophic factor (GDNF) family of growth factors may be involved in the regenerative support of neurons in the peripheral nervous system. In order to study the role of these growth factors and their receptors following rat peripheral nerve injury we examined the changes in their mRNA levels in the spinal cord, the dorsal root ganglia and the peripheral nerve trunk. Following transaction of the sciatic nerve GDNF mRNA was up-regulated rapidly in the denervated nerve distal to the cut along with the mRNA for one of its receptors, GFRalpha-1. GFRalpha-1 mRNA was also increased in the DRG ipsilateral to the nerve injury suggesting that GDNF may be involved in the trophic support of DRG sensory neurons. In contrast there were no analogous changes in the mRNA levels of neurturin, persephin and artemin following injury. PMID- 10852219 TI - Size distortion in spatial neglect. AB - One of the suggested theories to explain some aspects of neglect in patients with right parietal lesions is the spatial distortion hypothesis. To determine whether a distorted representation of space can account for the performance of neglect patients in visuospatial tasks we asked 27 neglect patients to compare the width of two horizontally aligned bars. The bars were presented symmetric to the body midline or either on the left or right side of egocentric space. The size comparison data are in accordance with the hypothesis of a distorted egocentric representation based on a dynamic remapping of space. The results support the idea that the abnormalities observed in the size comparison tasks are due to a distorted internal representation of the outside world. There are some hints that this distortion could be based on a dynamic mapping of space determined by the distribution of visuospatial attention. PMID- 10852220 TI - Neural discharge coupled to saccade offset in the cat visual cortex. AB - The increase in neural activity in cat visual cortex associated with eye movements has been thought to reflect a replica of the motor command signal. We examined the timing of the saccade-related increase in neural activity in cat areas 17 and 18 in relation to saccade onset and offset. The increase in activity was temporally coupled to saccade offset rather than onset both for visually guided saccades and for spontaneous saccades in the dark. Overall, it occurred 63 ms after saccade offset, and the peak was higher and sharper for data aligned at saccade offset than for onset. These results are inconsistent with the idea that saccade-related activity in cat visual cortex reflects a copy of the motor command signal. PMID- 10852221 TI - Increased survival of embryonic nigral neurons when grafted to hypothermic rats. AB - Hypothermia can reduce neuronal death caused by ischemia and traumatic brain injury. We therefore investigated whether mild hypothermia in rats receiving a transplant of embryonic mesencephalic rat tissue increases survival of the implanted dopaminergic neurons. Mild hypothermia (32-33 degrees C) during graft implantation and for the following 90 min significantly increased the survival of transplanted dopaminergic neurons to 171% of control values in normothermic (37 degrees C) rats. This demonstrates that treatment of the graft recipient for a relatively short period during and after surgery has a favorable effect on the survival of grafted dopaminergic neurons. These findings may be of importance for clinical neural transplantation trials which are in need of procedures that improve transplant survival. PMID- 10852222 TI - ATP P2X receptors play little role in the maintenance of neuropathic hyperalgesia. AB - There is good evidence that ATP receptors play a role in nociception in the periphery. We sought evidence that they contribute to neuropathic hyperalgesia. We carried out a partial ligation of the sciatic nerve in rats to induce nerve injury and neuropathic hyperalgesia. Intrathecal injection of suramin (a P2 purinoceptor antagonist) provided minor alleviation of thermal hyperalgesia, while PPADS (a selective P2X receptor antagonist) had no effect. Both suramin and PPADS caused abnormal behavior including aggressiveness and subsequent hyporeactivity and immobility. P2X receptors in the spinal cord do not appear to play a significant role in the maintenance of thermal hyperalgesia. However, P2X receptors may play an important role in the control of behaviour. PMID- 10852223 TI - fMRI activations of SI and SII cortices during tactile stimulation depend on attention. AB - Somatosensory cortical areas are widely studied with new functional imaging techniques, but usually with poor control over the attentional state of subjects. We applied fMRI to determine possible changes in activations of these areas due to modulations of attention. Attention induced large regional changes, mostly enlargements of activated areas, and more of those at SII than at SI. The number of instances where activation was only seen in attend conditions was also larger at SII than at SI. These results show the importance of controlling the attentional state of the subjects in imaging studies, and give evidence of the different roles of SI and SII cortices in the somatosensory system. PMID- 10852224 TI - Differential expression of Bax and Bcl-2 in the brains of hamsters infected with 263K scrapie agent. AB - To study the mechanism(s) of neuronal cell death during scrapie infection, we investigated the expression of Bax and Bcl-2 in brains of hamsters infected with 263K scrapie agent. The expression of Bcl-2 mRNA was significantly decreased in the brains of 263K scrapie-infected hamsters compared with controls, whereas the expression levels of Bax mRNA were significantly increased in scrapie-infected brain. The levels of Bax and Bcl-2 proteins in brains of scrapie and control animals reflected the difference in mRNA levels. Immunoreactivity for Bax and Bcl 2 were found predominantly within neurons. In scrapie-infected brains, the number of neuronal cells positive for Bcl-2 was significantly lower in the hippocampal CA3 region and was decreased in the cerebral cortex, whereas the number of neuronal cells positive for Bax was significantly increased in both regions. The possibility that differential regulation of Bax and Bcl-2 expression may play an important role in neuronal cell death induced by scrapie infection is discussed. PMID- 10852225 TI - ATP evokes different currents in TTX-sensitive and TTX-resistant cells of dorsal root ganglia. AB - The relationship between the level of expression of tetrodotoxin-sensitive (TTX S) and tetrodotoxin-resistant (TTX-R) sodium currents and the occurrence of two kinetically different ATP-induced currents in rat dorsal root ganglion neurons was studied. ATP evoked two distinct types of currents, one with fast activation and desensitization (I-fast) and the other with slow and persistent development (I-slow). In all TTX-S cells which expressed solely TTX-S sodium currents ATP evoked I-fast. The other cells expressed a variable proportion of TTX-S and TTX-R sodium currents. Only 15% of these TTX-R+S cells responded to ATP with I-fast. I slow was evoked in both cell types but the magnitude of response was much greater in TTX-R+S cells. This result suggests that a different array of ion channels is equipped in different types of sensory neurons. PMID- 10852226 TI - Cerebral correlates of working memory for temporal information. AB - Studies of neural correlates of working memory functions in the auditory-verbal, visuo-spatial and visuo-object domain suggest a category-specific organization of working memory processes in prefrontal cortex. Here, we used fMRI to explore brain areas that underlie different working memory operations directed to the temporal domain, which so far has been widely neglected. Significant activations related to memory updating and comparison processes were found right-accentuated in prefrontal and lateral premotor cortices. Furthermore, both subvocal rhythm encoding and maintenance enhanced left-lateralized activity in Broca's and supplementary motor area as well as in the sensorimotor cortex. Hemispheric lateralization effects of brain activity during temporal processing tasks may depend on the presence or absence of subvocal rehearsal strategies. PMID- 10852227 TI - Screening of AP endonuclease as a candidate gene for amyotrophic lateral sclerosis (ALS). AB - DNA extracted from CNS tissue of 84 patients was screened by single-stranded conformation polymorphism (SSCP) and heteroduplex analysis for mutations in the apurinic/apyrimidinic endonuclease (APE) gene. One mutation was identified and characterized as a 4bp deletion in the 3'UTR. A rare polymorphism was identified in exon 3 and a common polymorphism in the coding region of exon 5. These results suggest that APE mutations do not account for a large number of ALS cases. PMID- 10852228 TI - Central infusion of interleukin-1 receptor antagonist fails to alter feeding and weight gain. AB - Interleukin-1 (IL-1) administered either i.p. or i.c.v. provokes sickness behaviors, including suppression of feeding. As well, the possibility exists that IL-1 contributes to the cascade of factors that regulate feeding under basal conditions. The current study assessed the contribution of IL-1 in the control of food intake and body weight under physiological conditions in male rats. Pretreatment with an IL- I receptor antagonist (IL-1ra, 16 mg/kg, i.p.) completely blocked the suppression of food intake produced by injection of IL 1beta (4 microg/kg, i.p.). However, neither daily injections of IL-1ra (16 mg/kg, i.p.) for 4 consecutive days nor infusion of IL-1ra (500 microg/day, i.c.v.) for 7 days altered daily food intake and the rate of body weight gain. These findings suggest while IL-1 may play a role in anorexia associated with sickness, this cytokine likely does not play a physiological role in the regulation of daily food intake and long-term energy balance. PMID- 10852229 TI - Spike doublets in neurons of the lateral amygdala: mechanisms and contribution to rhythmic activity. AB - A majority of projection neurons in the lateral amygdala generate oscillatory spike firing in the theta-frequency range, largely due to intrinsic membrane properties. Here we report on the occurrence of spike doublets in about 70% of these cells. Spike doublets consisted of a fast initial and a second slower component, which were mediated by sodium- and calcium-dependent mechanisms, respectively. With increased level of depolarization, there was a gradual transition of fast action potentials, regular alternation of fast action potentials and spike doublets, regular spike doublets, and high-threshold oscillations. Fast Fourier transforms demonstrated the rhythmic nature of spike doublets at around 3 Hz with an intra-doublet frequency of 25-80 Hz. Spike doublets may thus contribute to the overall rhythmicity in the membrane potential patterns of projection cells and support the integration of synaptic input signals. PMID- 10852230 TI - Lung inflation inhibits rapidly adapting receptor relay neurons in the rat. AB - Pulmonary slowly adapting receptors (SARs) and rapidly adapting receptors (RARs) are important components of various respiratory reflexes. In anesthetized, paralyzed and artificially ventilated rats, we found an inhibitory linkage from the former to the latter system at the level of their second-order relay neurons (P cells and RAR cells, respectively). Lung inflation which activates RARs as well as SARs suppressed RAR cell activity evoked by electrical stimulation of the vagus nerve. Intracellular recordings from RAR cells showed IPSPs locked to electrical stimulation of the ipsilateral and contralateral vagus nerves at an intensity just above the threshold for P cell activation. Activation of P cells with glutamate suppressed RAR cell firing. Since P cells project to the area of RAR cells, taken together, these results strongly suggest that P cells synaptically inhibit RAR cells. PMID- 10852231 TI - Task-relevant selective modulation of somatosensory afferent paths from the lower limb. AB - Leg movement attenuates initial somatosensory evoked potentials (SEPS) from both cutaneous and muscle afferent origin. To date, as different sensory inputs become relevant for task performance, selective facilitation from such movement-related gating influences has not been shown. We hypothesized that initial SEP amplitudes from cutaneous (sural nerve, SN) and muscle afferent (tibial nerve, TN) sources are dependent on the relevance of the specific afferent information to task performance. SEPs were obtained at rest and during three movement conditions. In each movement condition, the left foot was passively moved episodically and additional cutaneous 'codes' of sensory information were applied to the dorsum of the left foot. Subjects were instructed to: simply relax (passive), or to make a response following the cessation of movement, dependent either on the cutaneous code (cutaneous task), or the passive movement trajectory of the left foot (position task). Passive movement, with no required subsequent response, attenuated initial TN and SN SEPs to approximately 40% of that at rest (p < 0.05). Versus passive movement, when cutaneous inputs provided the relevant cue for the task, mean SN SEPs significantly increased (p < 0.05), and when the proprioceptive inputs provided the relevant cue for the task, mean TN SEPs significantly increased (p < 0.05). We conclude that specific relevancy of sensory information selectively facilitates somatosensory paths from movement related attenuation. PMID- 10852232 TI - Activation of NMDA receptors drives action potentials in superficial dorsal horn from neonatal rats. AB - We have investigated the role of NMDA receptors in mediating synaptic transmission in spinal cord lamina II over the first 2 weeks of postnatal development. High intensity root stimulation evoked D-APV-sensitive slow synaptic activity in lamina II neurons that drove action potential firing. This NMDA receptor-mediated activity was enhanced when bicuculline and strychnine were used to block synaptic inhibition. When activated by repetitive focal stimulation, synaptic activity mediated by NMDA receptors alone drove action potential firing. NMDA receptors were also able to drive action potential firing at synapses where AMPA receptors were present but blocked. Our data show that in lamina II of the dorsal horn, NMDA receptors significantly affect neuronal excitability even in the absence of co-activation of AMPA receptors. PMID- 10852233 TI - Silent speechreading in the absence of scanner noise: an event-related fMRI study. AB - In a previous study we used functional magnetic resonance imaging (fMRI) to demonstrate activation in auditory cortex during silent speechreading. Since image acquisition during fMRI generates acoustic noise, this pattern of activation could have reflected an interaction between background scanner noise and the visual lip-read stimuli. In this study we employed an event-related fMRI design which allowed us to measure activation during speechreading in the absence of acoustic scanner noise. In the experimental condition, hearing subjects were required to speechread random numbers from a silent speaker. In the control condition subjects watched a static image of the same speaker with mouth closed and were required to subvocally count an intermittent visual cue. A single volume of images was collected to coincide with the estimated peak of the blood oxygen level dependent (BOLD) response to these stimuli across multiple baseline and experimental trials. Silent speechreading led to greater activation in lateral temporal cortex relative to the control condition. This indicates that activation of auditory areas during silent speechreading is not a function of acoustic scanner noise and confirms that silent speechreading engages similar regions of auditory cortex as listening to speech. PMID- 10852234 TI - Estrogen receptors and insulin-like growth factor-I receptors mediate estrogen dependent synaptic plasticity. AB - Previous studies have shown that estradiol induces a transient disconnection of axo-somatic inhibitory synapses in the hypothalamic arcuate nucleus of adult ovariectomized rats. The synaptic disconnection is accompanied by an increase in the levels of insulin-like growth factor-I (IGF-I) in the arcuate nucleus, suggesting that IGF-I signaling may be involved in the estrogen-induced synaptic plasticity. The role of estrogen receptors and IGF-I receptors in the synaptic changes has been studied by assessing the number of axo-somatic synapses in ovariectomized rats treated with intracerebroventricular administration of the estrogen receptor antagonist ICI 182,780 and the IGF-I receptor antagonist JBI to ovariectomized rats. Estradiol administration resulted in a significant decrease in the number of axo-somatic synapses on arcuate neurons in control ovariectomized rats. Both the estrogen receptor antagonist and the IGF-I receptor antagonist blocked the estrogen-induced synaptic decrease. This finding suggest that estrogen-induced synaptic plasticity in the arcuate nucleus is dependent on the activation of both estrogen receptors and IGF-I receptors. PMID- 10852235 TI - Selective attention to emotional stimuli in a verbal go/no-go task: an fMRI study. AB - Tasks requiring subjects to attend emotional attributes of words have been used to study mood-congruent information processing biases in anxiety and affective disorders. In this study we adapted an emotional go/no-go task, for use with fMRI to assess the neural substrates of focusing on emotional attributes of words in normal subjects. The key findings were that responding to targets defined on the basis of meaning of words compared to targets defined on the basis of perceptual features was associated with response in inferior frontal gyrus and dorsal anterior cingulate. Further, selecting emotional targets, whether happy or sad, was associated with enhanced response in the subgenual cingulate, while happy targets elicited enhanced neural response in ventral anterior cingulate. These findings reaffirm the importance of medial prefrontal regions in normal emotional processing. PMID- 10852236 TI - Differential effects of vestibular stimulation on walking and running. AB - Prompted by our recent observation that an acute vestibular tone imbalance causes less deviation from the intended path when running than when slowly walking, we examined 10 healthy subjects when walking or running at different step frequencies during galvanic vestibular stimulation. Blindfolded subjects were asked to walk (1 Hz step frequency) or run (3 Hz step frequency) straight ahead toward a previously seen target. The mean gait deviation after 10 s was 6.0 +/- 2.4 degrees at 1 Hz and 2.8 +/- 1.8 degrees at 3 Hz step frequency (n = 10; p < 0.001, paired t-test). In a second experiment walking and running in place were investigated. There was no significant difference in body displacement. Walking and running are highly automated processes based on spinal locomotor generators that are under supraspinal control. We conclude that vestibular input is differentially regulated depending on the locomotion speed and pattern used. PMID- 10852237 TI - Greater song complexity is associated with augmented song system anatomy in zebra finches. AB - We revisited the relationship between brain anatomy and song behavior in zebra finches. Consistent with previous studies in other songbirds, we find that differences in volume of the telencephalic song control nucleus HVc is predictive of differences in repertoire size and phrase duration in zebra finches. We extend the study of brain and behavior correlations in song birds by showing that repertoire size in zebra finches can be predicted by variance in several brain nuclei, providing the first demonstration that volumetric differences across multiple components of a neural network are predictive of song behavior. PMID- 10852238 TI - Hypocretin/orexin depolarizes and decreases potassium conductance in locus coeruleus neurons. AB - Recent studies demonstrated that noradrenergic locus coeruleus (LC) neurons are a particularly strong target of the novel neuropeptide, hypocretin (orexin). The present study sought to elucidate the action of hypocretin-B (HCRT) on LC neurons recorded intracellularly in rat brain slices. Bath (1.0 microM) or local puff application (50-100 microM in pipette) of HCRT depolarized LC neurons in rat brain slices and increased their spontaneous discharge rate. Depolarization evoked by HCRT was persistent in the presence of tetrodotoxin (TTX, 1 microM) and Co2+ (1 mM), indicating that HCRT directly activated LC neurons, and that its effect on the postsynaptic cell was not due to activation of TTX-sensitive sodium channels or Co2+-sensitive calcium channels. The apparent input resistance was significantly increased in the majority of LC neurons during the HCRT-evoked depolarization. Moreover, the HCRT-evoked depolarization was decreased in amplitude with hyperpolarization of membrane. The present results indicate that decreased potassium conductance is involved in the effect of HCRT on LC neurons. PMID- 10852239 TI - LPS-induced neuroinflammatory effects do not recover with time. AB - Inflammatory processes develop in the vicinity of the neuropathological hallmarks associated with Alzheimer's disease (AD) and may play a role in the progression of the disease and its clinical expression. We have previously reported that chronic infusion of LPS into the fourth ventricle of rat brains reproduced many of the inflammatory and pathological changes seen in the brain of AD patients. In the current study, we used the same animal model to investigate the effects of longer infusion of LPS and whether these effects could recover over time. The results show that doubling the time of LPS infusion did not increase the inflammatory reaction and did not produce a significantly greater behavioral impairment. Waiting for 37 days after the cessation of the LPS infusion did not decrease the density of activated microglia and did not improve performances in the Morris water maze task. The results suggest that inflammation may contribute to the pathogenic mechanisms that underlie the clinical expression of AD. PMID- 10852240 TI - Lipid peroxidation inhibition in spinal cord injury: cyclosporin-A vs methylprednisolone. AB - To compare the effectiveness of cyclosporin-A (CsA) with methylprednisolone (MP) or a combination of both upon inhibition of lipid peroxidation (LP) after spinal cord (SC) injury, rats were treated with either CsA, MP, CSA+MP or vehicle starting 1 h after SC contusion at T9 level. LP was assessed 24h after injury by the lipid fluorescent product formation method. The survival rate was also evaluated in other series of rats by the Kaplan-Meier curves. Lipid peroxidation was similarly inhibited in rats treated with CsA, MP, or CSA+MP (p>0.05). Animals receiving MP (alone or combined with CsA) showed the poorest surviving rate. LP was inhibited by CsA to the same extent as by MP but without the lethal effect of the latter. PMID- 10852241 TI - Exogenous administration of L-arginine protects cholinergic neurons from colchicine neurotoxicity. AB - The effect of L-arginine administration on susceptibility/resistance of NO producing neurons was investigated in non-ischemic neurodegenerative conditions. ChAT mRNA level was measured by in situ hybridization in the basal forebrain after i.c.v. colchicine administration. Colchicine induced down-regulation of ChAT mRNA level followed by degeneration of cholinergic neurons. L-Arginine treatment increased ChAT mRNA level and prevented/delayed ChAT mRNA decrease by colchicine. These data suggest that L-arginine treatment may protect cholinergic neurons from colchicine-induced degeneration. PMID- 10852242 TI - Identification of PLC beta and PKC in pheromone receptor neurons of Antheraea polyphemus. AB - Two proteins of the IP3 transduction pathway were identified by Western blots in homogenates of isolated pheromone-sensitive sensilla of the silkmoth Antheraea polyphemus. A 110 kDa protein was recognized by an antiserum raised against the Drosophila phospholipase C beta (PLC beta p121) and a 80kDa protein was labelled by an antiserum against a synthetic peptide of a conserved region of protein kinase C (PKC). Incubation of homogenized sensory hairs with the main sex pheromone component, (E,Z) 6-11 hexadecadienyl acetate, resulted in a 6-fold increase in the activity of PKC compared to controls without pheromone. In contrast, incubation with pheromone did not affect the activity of protein kinase A (PKA). Activation of PKC by the membrane permeable dioctanoylglycerol led to excitation of the pheromone-sensitive receptor neurons. These data support the current concept that pheromone perception of moths is mediated by the IP3 transduction pathway. PMID- 10852243 TI - Excitatory amino acids modulate epileptogenesis in the brain stem. AB - Activation of cholinergic mechanisms in the pontine reticular formation by local microinjections of carbachol was shown to induce pontine electrographic seizures and clonic convulsions. In this study we found that glutamate microinjections into the pons induced similar electrographic seizures and clonic convulsions. Microinjections into the PRF of glutamate in subconvulsive doses prior to carbachol potentiated the epileptogenic effect of carbachol. The duration of the seizure activity increased and the convulsions became more severe. The NMDA receptor antagonist MK-801 and the non-NMDA receptor antagonists DNQX significantly reduced the potentiating effect of glutamate. These results indicate a possible role of EAA receptors in the generation of epilepsy in the pons. They also suggest the importance of studying the role of synergistic interactions between EAA mechanisms and cholinergic mechanisms in the various pontine functions. PMID- 10852244 TI - GDNF but not BDNF is increased in cerebrospinal fluid in amyotrophic lateral sclerosis. AB - Cerebrospinal fluid from 15 patients with ALS and 11 controls without neurological disease were analysed for levels of the neurotrophic factors BDNF and GDNF. Analyses were performed using a sensitive sandwich immunoassay (ELISA). There was no significant difference in BDNF levels between the ALS patients and the control subjects studied. Measurable levels of GDNF were found in 12 out of 15 ALS samples. GDNF was not detected in CSF from any of the control subjects. The finding of increased CSF levels of GDNF in ALS compared to controls, together with earlier findings of increased expression of GDNF mRNA in muscle in ALS, indicates that the capacity to synthesize GDNF is enhanced in this disorder. PMID- 10852245 TI - Inhibition of rat nucleus tractus solitarius neurones by activation of 5-HT2C receptors. AB - In vivo, nucleus tractus solitarius (NTS) neurones receiving monosynaptic vagal input and inactive intermediate neurones were inhibited by both DOI and a selective 5-HT2C receptor agonist, MK-212. Cells receiving a more polysynaptic input were excited by DOI and although MK-212 also excited a few of these cells, the majority of cells in these groups were unaffected by MK-212. The inhibitory, but not the excitatory actions of both MK-212 and DOI were prevented by a selective 5-HT2C receptor antagonist, RS-102221. In contrast, most dorsal vagal preganglionic neurones were unaffected by application of either DOI or MK-212, the few remaining cells being excited by both agonists. These data demonstrate that DOI-evoked inhibition of NTS cells activated by vagal afferent input and DOI evoked excitation of vagal preganglionic neurones is mediated by 5-HT2C receptors. PMID- 10852246 TI - Neuroprotective effect of retro-inverso Prosaptide D5 on focal cerebral ischemia in rat. AB - Prosaposin (the precursor of saposins A-D) has been identified as a neurotrophic factor in vitro and in vivo. In this study, a novel 11-mer retro-inverso peptidomimetic, Prosaptide D5, was injected i.m. to assess its effectiveness in a rat ischemic model produced by reversible total occlusion of the left middle cerebral artery (MCA). Prosaptide (300 microg/kg, i.m.) injected 3 h after reversible occlusion reduced brain infarct area by 56% compared with a saline group (p < 0.01) at 21 h of reperfusion. A similar injection of D5 6h after occlusion produced a 32% decrease. PMID- 10852247 TI - Amino acids in spinal dorsal horn of patients during surgery for neuropathic pain or spasticity. AB - A specific microdialysis probe combined with high performance liquid chromatography analysis was used to compare the basal levels of extracellular amino acids (AAs) in the dorsal horn (DH) between two groups of patients undergoing spinal surgery for the treatment of peripheral neuropathic pain (n = 5) or disabling spasticity (n = 5). A stabilized concentration was reached in the dialysates 45 min after probe implantation for excitatory AAs (glutamate and aspartate) and inhibitory AAs (GABA and glycine). A significant increase in the ratios aspartate/GABA and aspartate/glycine was found in the group of patients suffering from neuropathic pain. This study shows the feasibility of a microdialysis investigation in the DH of patients during a neurosurgical operation and supports in humans the hypothesis of an imbalance between excitatory AAs and inhibitory AAs within the DH in neuropathic pain states, as suggested by previous animal studies. PMID- 10852248 TI - Adhesive capsulitis--research advances frozen in time? PMID- 10852249 TI - Things do not get better by being left alone. The physician and complementary medicine. PMID- 10852250 TI - Colchicine use in cyclosporine treated transplant recipients: how little is too much? PMID- 10852251 TI - Analysis of the effect of COX-2 specific inhibitors and recommendations for their use in clinical practice. PMID- 10852252 TI - Enhanced neutrophilic granulopoiesis in rheumatoid arthritis. Involvement of neutrophils in disease progression. AB - OBJECTIVE: To investigate enhanced granulopoiesis in bone marrow of patients with rheumatoid arthritis (RA), and the role of neutrophils in RA pathogenesis. METHODS: Aspirated bone marrow cells and peripheral blood leukocytes from patients with RA and non-RA patient controls were analyzed morphologically and by 2 color flow cytometry. Thirteen iliac bones (8 RA, 5 non-RA) were examined by light and transmission electron microscope (TEM). RESULTS: The percentage of CD15+CD16- cells (immature neutrophils) in RA bone marrow (64.3+/-13.4%, mean +/- SD) increased significantly compared to that of non-RA controls (43.2+/-14.3%), whereas the fraction of CD15+CD]6+ cells (mature neutrophils)was greatly decreased (RA 21.8+/-10.1%; non-RA 38.1+/-8.9%). The absolute number of CD15+CD16 cells also increased markedly in RA bone marrow. The ratio of immature cells to the total granulocytes (% CD15+CD16- to % CD15+) correlated with the Lansbury Index score (R = 0.76, p<0.0001). TEM observations revealed that abundant immature neutrophils adhered closely to the trabeculae of the iliac bone. Margins of trabeculae were mostly irregular, especially in severe RA, and collagenous fibers frequently disappeared in those trabeculae with ragged margins. CONCLUSION: In RA bone marrow, immature neutrophils (CD15+CD16-) were markedly increased in number; by contrast, no changes were found for mature cells. Augmented production of immature neutrophils (at the promyelocyte-to-myelocyte stage) might lead to the destruction of collagenous fibers in RA bone trabeculae, as revealed by TEM. Generalized bone destruction in RA might, at least in part, be caused by enhanced production of immature neutrophils. PMID- 10852253 TI - Interleukin-1beta induces elevation of spermidine/spermine N1-acetyltransferase activity and an increase in the amount of putrescine in synovial adherent cells from patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate polyamine metabolism in rheumatoid synovial adherent cells stimulated by interleukin- 1beta (IL-1beta). METHODS: Synovial adherent cells obtained from patients with rheumatoid arthritis (RA) were cultured and incubated in the presence or absence of human recombinant IL-1beta at a concentration of 10 ng/ml for 24 h. The cellular contents of polyamines as well as the activities of spermidine/spermine N1-acetyltransferase (SAT) and ornithine decarboxylase (ODC) were measured. RESULTS: Polyamines in synovial adherent cells decreased significantly after 24 h incubation in the absence of IL-1beta. However, in the presence of IL-Ibeta, putrescine and N'-acetylspermidine increased significantly. No significant difference was observed between the amount of spermidine in synovial adherent cells incubated with and without IL 1beta. Spermine and N8-acetylspermidine in synovial adherent cells incubated with IL-1beta decreased significantly more than in synovial adherent cells incubated without. SAT activity reached a peak 12 h after the addition of IL-1beta and then decreased, while the ODC activity did not increase. SAT activity was elevated by the addition of IL-1beta in a dose dependent manner. CONCLUSION: An increase in the putrescine level in rheumatoid synovial adherent cells as a result of the elevation of SAT activity induced by IL-1beta may play a role in RA. PMID- 10852254 TI - In vitro and in vivo inhibition of activation induced T cell apoptosis by bucillamine. AB - OBJECTIVE: To investigate the mechanism of autoimmune phenomena, occasionally seen in patients with rheumatoid arthritis treated with bucillamine (BUC) and D penicillamine (D-Pen), by evaluating their effects on apoptosis of T cells induced by T cell receptor activation or dexamethasone. METHODS: In vitro apoptosis was induced in a T cell hybridoma (SSP3.7) and a B cell line (WEHI 231) by activation of respective receptors or dexamethasone, in the presence or absence of BUC or D-Pen. In vivo apoptosis was induced in BALB/c mice by staphylococcal enterotoxin B (SEB), with or without BUC or D-Pen, and thymocytes were examined for it by FACS. RESULTS: Stimulation with anti-CD3 and dexamethasone induced apoptosis in 72% and 71% of SSP3.7 cells, respectively. However, only 16% of SSP3.7 cells became apoptotic by anti-CD3 when BUC was added to the culture media. By contrast, 80% of SSP3.7 cells became apoptotic when stimulated by dexamethasone, even in the presence of BUC. BUC did not affect apoptosis of WEHI 231 cells induced by anti-IgM. Although SA981 (a metabolite of BUC) inhibited apoptosis of SSP3.7 cells induced by anti-CD3, D-Pen did not. BUC, SA981, or D-Pen did not significantly influence the level of interleukin 2 secretion stimulated by anti-CD3. In contrast, both BUC and D-Pen inhibited apoptosis of Vbeta8+ thymocytes induced in vivo by SEB superantigen. Neither BUC nor D-Pen significantly changed the number of CD4+CD8+ thymocytes in BALB/c mice injected with dexamethasone. CONCLUSION: BUC decreased, while D-Pen did not, the apoptosis of T cells stimulated by anti-CD3 in vitro, although they both inhibited the deletion of immature thymocytes reactive with SEB in vivo. This may explain autoimmune phenomena sometimes seen during the treatment of rheumatic patients with these drugs. PMID- 10852255 TI - Randomized double blind trial of an ayurvedic plant derived formulation for treatment of rheumatoid arthritis. AB - OBJECTIVE: To evaluate RA-1, a standardized plant extract formulation, traditionally considered a safe, effective antiarthritic in the Asian-Indian Ayurvedic medicinal system. METHODS: One hundred eighty-two patients with active on-chronic rheumatoid arthritis (RA) participated in a 16 week randomized, double blind, placebo controlled, parallel efficacy clinical drug trial in Pune, India. Tenderness, pain, swelling, and several other efficacy measures were assessed by (1) ACR core set 20% and 50% improvement; (2) ACR 20% improvement response. An intent-to-treat analysis was performed; p<0.05 considered significant. RESULTS: Seventeen patients withdrew (active = 9; placebo = 8); none withdrew due to drug toxicity. An unprecedented placebo response (often p<0.001 in within-group change) was observed. The active RA-1 group remained numerically superior at all evaluation timepoints. RA-1 demonstrated few significant differences: (1) increased proportion with 50% reduction in swollen joint count (95% CI approximately 1.52, 29.90) and swollen joint score (95% CI approximately 0.91, 28.73); (2) reduced rheumatoid factor (95% CI approximately -303.7, -2.72); 39% in the RA-1 group versus 30% placebo showed ACR 20% improvement (95% CI approximately -5.48, 24.59). Only minor side effects were seen, with no significant differences by treatment group. CONCLUSION: In a trial with sufficient power, RA-1 revealed efficacy that was not significantly superior to the strong placebo response, except for improvement in joint swelling. Further, the effect on RF and good safety profile led to an open label phase. PMID- 10852256 TI - Gastrointestinal symptoms and health related quality of life in patients with arthritis. AB - OBJECTIVE: To evaluate the relationship between gastrointestinal (GI) symptoms and health related quality of life (QOL) in patients with osteoarthritis (OA) and rheumatoid arthritis (RA). METHODS: A total of 1773 patients with arthritis participating in a longterm outcome study (OA of the hip or knee = 648, RA = 1125) completed mailed surveys that included assessments of GI symptoms and overall GI symptom severity, Short Form-36, the visual analog scale (VAS) for the EuroQol (Health QOL), a VAS global disease severity scale, and measures of disease and psychological status. The overall response rate exceeded 85%. RESULTS: Dyspepsia (heartburn, bloating, or belching) and upper abdominal/epigastric pain were identified as the most important GI contributors to reduction in QOL, and the simultaneous presence of both these symptoms was associated with lower QOL (54.5) compared to those without symptoms (70.9) on the 0-100 Health QOL scale. Similarly, those in the upper tertile of the global GI severity scale had Health QOL scores of 55.7 compared to 76.4 for those in the lower tertile. These differences in GI symptoms and GI severity, however, were reduced substantially when the effects of functional disability, pain, and depression were adjusted for: 62.3 to 68.6 (p = 0.003) and 63.7 to 70.3 (p<0.001) for the GI symptoms and GI severity scales, respectively. CONCLUSION: QOL is significantly impaired among unselected arthritis patients with GI symptoms compared to those without these symptoms. Dyspepsia and upper abdominal/epigastric pain are more strongly related to QOL measures than other GI symptoms, and are common among arthritis patients. It is possible to construct a simple scale of these 2 symptoms or to use the VAS GI severity scale and get a clinically useful idea of the current level of GI distress and alteration of QOL by GI problems. Two components of impairment can be identified, one that is smaller and unrelated to disease or psychological factors, and a second that is larger and includes these factors. Because GI symptoms can alter function, pain, and psychological status, it is likely that the true effect of GI symptoms on QOL is somewhere between the unadjusted and adjusted values cited above. PMID- 10852257 TI - Immunoglobulin G glycosylation and clinical outcome in rheumatoid arthritis during pregnancy. AB - OBJECTIVE: To determine whether clinical outcome during pregnancy in rheumatoid arthritis (RA) is associated with changes in the levels of exposed immunoglobulin G (IgG) terminal sugars. METHODS: Serum IgG glycosylation from 23 pregnant patients with RA was analyzed during the prenatal, antenatal, and post-partum periods. Patients were randomly selected on the basis of whether they achieved spontaneous remission (n = 11) or did not remit (n = 12); of the latter group 6 patients experienced a relapse in disease activity. Levels of exposed terminal IgG sugars, galactose (Gal), N-acetylglucosamine (GlcNAc), and sialic acid (SA), were estimated in a lectin binding assay using Ricinis (communis, Bandeiraea simplicifolia II, and Sambucus nigra, respectively. RESULTS: Exposed Gal levels increased (p<0.02) and GlcNAc levels decreased (p<0.05) in the antenatal period, and returned to preconception levels during post-partum. GlcNAc rebound was instantaneous (p<0.005), whereas Gal remained high for a further 10 weeks. SA did not undergo any major changes. Remission was associated with an earlier and significantly greater antenatal reduction in GlcNAc (2nd and 3rd trimester; p<0.02) in comparison to the groups that did not experience a decrease in disease activity. Analysis of individual IgG samples during the first trimester revealed a significant negative correlation between Gal and GlcNAc in the remission group (r = -0.80; p<0.05), which was opposite to that found in the relapse group (r = +0.87; p<0.03). There was no significant difference between the groups with regard to the timing and/or incidence of a post-partum flare of disease. CONCLUSION: Temporal changes in the levels of IgG terminal sugars, in particular exposed GlcNAc, are integrally associated with the clinical manifestation of RA in pregnancy. Generation of IgG sugar micro-heterogeneity is complex and understanding it may help unravel pathogenic features associated with RA. PMID- 10852258 TI - Incidence and prevalence of rheumatoid arthritis in the county of Troms, northern Norway. AB - OBJECTIVE: To investigate the population incidence and prevalence of rheumatoid arthritis (RA) in persons above the age of 20 in the county of Troms, northern Norway, during the period 1987-1996. METHODS: All records of patients with RA registered at the Department of Rheumatology at the University Hospital of Tromso during the years 1987 to 1996 were reviewed. The diagnosis of RA was set in accordance with the 1987 American Rheumatism Association criteria, and the population data were based on the 1989 and 1994 census. Total and age-specific incidence rates were calculated as number of new cases per 100,000 inhabitants and year. Age adjusted incidence rates were obtained by the direct method. Prevalence rates of RA for January 1, 1989, and January 1, 1994, were estimated. RESULTS: The total annual incidence rate for the period 1987-1996 was 28.7/100,000 per year (36.0/100,000 in women and 21.4/100,000 in men). No significant difference in incidence rates was found between the periods 1987-1991 and 1992-1996. Total prevalence of RA was 0.39% in 1989 and 0.47% in 1994. The corresponding data for women and men were 0.54% and 0.24% in 1989, and 0.63% and 0.30% in 1994, respectively. CONCLUSION: We found a rather low incidence and prevalence of RA in the county of Troms in northern Norway. Females contracted RA significantly more often than males. There was no change in incidence rates during the observation period, lending no support to suggestions of a continuously decreasing occurrence of RA. PMID- 10852259 TI - The accuracy of self-report of physician diagnosed rheumatoid arthritis in moderately to severely disabled older women. Women's Health and Aging Collaborative Research Group. AB - OBJECTIVE: To determine the accuracy of self-report of physician diagnosed rheumatoid arthritis (RA) in moderately to severely disabled older women. METHODS: A total of 1002 participants in the Women's Health and Aging Study were included. These women were > or =65 years old, had an average of 4 chronic illnesses, and represented the one-third most disabled women living in the community. Self-report of a physician's diagnosis of RA was compared to cases of "definite" RA that were adjudicated using an algorithm modeled on the American College of Rheumatology criteria for RA. RESULTS: The sensitivity of self-report of physician diagnosed RA was 77%, with 90.6% specificity and 99% negative predictive value, kappa = 0.46. The positive predictive value was 34% and likely reflected the low prevalence of RA in this sample (3.1%). Five of the 6 women who did not correctly report RA were under the care of a rheumatologist. CONCLUSION: The accuracy of self-report of a physician's diagnosis of RA in this sample of disabled women with multiple chronic illnesses matched that observed in the general adult population of previous studies. Accuracy was enhanced by including report of receiving care by a rheumatologist. PMID- 10852260 TI - Patient outcomes following Swanson silastic metacarpophalangeal joint arthroplasty in the rheumatoid hand: a systematic overview. AB - OBJECTIVE: To investigate the effectiveness of Swanson silastic metacarpophalangeal joint arthroplasty (SMPA) in improving hand function for patients with rheumatoid arthritis. METHODS: A systematic overview of all published series in the literature on SMPA from 1966 to 1999. RESULTS: Research design deficiencies were quite prevalent in the literature on SMPA. However, SMPA was effective in correcting ulnar drift and in improving the arc of motion of the fingers. Health related quality of life was improved in the domains of hand function, pain, activities of daily living, aesthetics, and satisfaction. CONCLUSION: SMPA appeared to be an effective procedure in correcting rheumatoid hand deformities. Future research must establish objective, quantifiable measures of hand function improvement by using standardized hand function tests and validated hand outcome questionnaires. PMID- 10852261 TI - Clinical significance of anticentromere antibodies in patients with systemic lupus erythematosus. AB - OBJECTIVE: To clarify the clinical significance of anticentromere antibodies (ACA) in patients with systemic lupus erythematosus (SLE). METHODS: Two hundred sixteen patients with SLE who were treated in our department were surveyed cross sectionally for the presence of ACA using indirect immunofluorescence on HEp-2 cell lines. ACA were identified by their discrete speckled pattern. Antibodies to the major centromere protein, CENP-B, were also studied with ELISA. Serial determinations of anti-CENP-B were carried out using stored serum samples, if available. RESULTS: ACA were recognized in 12 (5.6%) patients with SLE. All patients were receiving steroid therapy, with a mean dose of prednisolone of 14.4 mg/day. These patients also tested positive for anti-CENP-B with high titers despite the low serological disease activity in most. Three or more CREST features were observed in 2 patients and 2 others had no such features. Both patients without CREST features had a relatively short disease duration. The age at onset of SLE was significantly higher and Raynaud's phenomenon was more frequent in patients with ACA than in patients without ACA. In 8 of 10 patients tested, retrospective analysis using stored sera revealed no consistent change in anti-CENP-B titers over time. CONCLUSION: The presence of ACA in patients with SLE is apparently more frequent than previously believed. Patients with SLE with ACA may be a distinct subgroup. A longterm followup is warranted to fully determine the clinical significance of ACA in patients with SLE. PMID- 10852262 TI - Development and testing of a systemic lupus-specific risk adjustment index for in hospital mortality. AB - OBJECTIVE: Valid comparison of patient outcomes among hospitals requires adjustment for differences in the severity of patients' illness. Disease-specific indexes of severity of illness may permit more accurate risk adjustment than generic indexes. The objective of this study was to develop a systemic lupus specific risk adjustment index for in-hospital mortality, and to compare its performance to that of the generic Charlson index. METHODS: A systemic lupus specific risk adjustment index was developed using discharge abstract data from a 50% random sample (n = 4994) of patients with systemic lupus erythematosus (SLE) hospitalized on an emergent or urgent basis in California from 1991 to 1994 (n = 9989). The index was tested on the remaining members of the sample. Candidate variables for the index were the diagnoses included in the original Charlson index, and nephritis, chronic renal failure, pericarditis, pleuritis, psychosis, seizures, hemolytic anemia, and thrombocytopenia. Multivariate logistic regression analysis was used to identify the set of variables that best differentiated those patients who died in the hospital from those who survived, and to provide the weights for construction of the index. RESULTS: In the derivation set of patients, the SLE-specific index accurately predicted in hospital mortality (area under the receiver operating characteristic curve c = 0.79). In the test set, the SLE-specific index (c = 0.72) was similar to the original Charlson index (c = 0.74) in its ability to predict in-hospital mortality (p = 0.32). However, the SLE-specific index accounted for substantially more variation in the risk of mortality among patients (R2 = 0.069) than did the Charlson index (R2 = 0.036). Use of the SLE-specific index rather than the Charlson index for risk adjustment did not alter the association between hospital experience and the probability of in-hospital mortality. Results were similar in the subgroups of patients with emergent hospitalizations and those with emergent hospitalizations due to SLE. CONCLUSION: The SLE-specific risk adjustment index developed from diagnoses recorded in administrative discharge abstracts performed similarly to the generic Charlson index in correctly classifying mortality outcomes, but the SLE-specific index stratified patients by their level of risk of mortality better than the Charlson index. Adjustment for SLE-specific risks of mortality did not alter the association between hospital experience and the risk of in-hospital mortality. PMID- 10852263 TI - A prospective study of factors affecting quality of life in systemic lupus erythematosus. AB - OBJECTIVE: To prospectively identify factors influencing quality of life (QOL) over 6 months in patients with systemic lupus erythematosus (SLE). METHODS: Ninety ethnically diverse patients with SLE completed questionnaires administered 6 months apart assessing QOL (using the Medical Outcomes Study Short Form-36) and demographic, socioeconomic, psychosocial, and behavioral factors. Disease activity, damage, and treatment were recorded at both evaluations. Multiple linear regression (adjusting for baseline health status) was used to identify factors influencing mental and physical health. RESULTS: Improved physical health after 6 months was associated with reductions in learned helplessness (p = 0.034), improved mental health (p<0.001), longer disease duration (p = 0.009), and better physical health at baseline (p = 0.027). Improved mental health after 6 months was associated with better family support (p = 0.002), improvements in physical health (p<0.001), disease activity, and prednisolone dose (interaction term p = 0.019), less disease related damage (p<0.001), non-use of cytotoxic drugs (p = 0.02), and older age at diagnosis (p = 0.007). CONCLUSION: Potentially modifiable psychosocial, disease, and therapy related factors influence QOL in patients with SLE. PMID- 10852264 TI - Do lupus disease activity measures detect clinically important change? AB - OBJECTIVE: New scales for the clinical assessment of patients with systemic lupus erythematosus (SLE) are valid and reliable, and quantitate disease activity. We assessed the responsiveness to change of 2 widely used standardized multi-item lupus activity measures, the revised Systemic Lupus Activity Measure (SLAM-R) and the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI), and their ability to detect clinically relevant changes. METHODS: Ninety-six (96) patients with definite SLE participated in this study. The group mean age was 45.0 (13.7) years, 91% were female, and the mean disease duration was 14.9 (7.5) years. Sociodemographic information, lupus activity (SLAM-R, SLEDAI), and damage were recorded at baseline. At each of the 5 monthly followup visits, the activity measures were repeated and a transition scale asked the physician if their patient's lupus activity had changed. Five different methods were used to compare the responsiveness of the activity measures studied: 1. the effect size; 2. the standardized response mean; 3. the control standardized response mean; 4. the area under the curve of a receiver operating characteristic (ROC) curve; and 5. a new multiple response modeling approach. RESULTS: Both SLAM-R and SLEDAI are responsive. SLAM-R is consistently, although moderately, more responsive than SLEDAI. All 5 methods of evaluating responsiveness yielded a consistent ranking of disease activity measures. CONCLUSION: SLAM-R and SLEDAI are responsive measures of lupus activity. SLAM-R appears to be more responsive than SLEDAI. PMID- 10852265 TI - Thalidomide in the treatment of cutaneous lupus refractory to conventional therapy. AB - OBJECTIVE: We describe a prospective treatment study of thalidomide in a series of 22 patients with cutaneous lupus refractory to other treatments. METHODS: From 1992 to 1998, 22 patients with cutaneous lupus (9 with discoid lupus erythematosus, 7 with subacute cutaneous lupus, 4 with profundus lupus, 2 with nonspecific rash) were treated with thalidomide. Initial treatment was started at 100 mg daily. If the cutaneous lesions vanished, the dose was lowered to 50-25 mg daily as a maintenance therapy and it was considered a complete remission. If the lesions improved but remained, this was considered a partial response and treatment was continued until the lesions were not further modified. Periodically, adverse effects were evaluated. RESULTS: Three patients discontinued treatment because of side effects such as vertigo, persistent drowsiness, or paresthesia. Rash improved in 16/19 patients (84%). Complete remission occurred in 12/16 (75%). In 9 (65%) the rash resolved, but recurred 4 16 weeks after withdrawal of thalidomide; when it was used again, they improved. Partial response was achieved in 4/16 (25%) patients. No response occurred in 3/19 (16%). Many patients noted improvement within 2 weeks after starting thalidomide and maximum benefit was achieved within 3 months. Five of the 14 women had amenorrhea during the treatment with thalidomide. CONCLUSION: Thalidomide is effective in the treatment of cutaneous lupus refractory to other treatments. However, only some patients had a remission; the remainder relapsed when treatment was withdrawn, or required low doses of thalidomide to preserve inactive lesions. Amenorrhea was observed as a new secondary effect of thalidomide. PMID- 10852266 TI - Connective tissue disease and other related rheumatic conditions among patients with finger and hand and temporomandibular joint prostheses in Denmark. AB - OBJECTIVE: To determine if finger and hand joint prostheses or temporomandibular joint (TMJ) implants may be involved in the initiation of specific connective tissue diseases (CTD), a nationwide cohort in Denmark was followed prospectively to evaluate rates of CTD after receiving these implants. METHODS: Danish patients with finger and hand joint implants (n = 562) or TMJ implants (n = 351) were identified and followed for subsequent hospitalizations. Observed numbers of hospitalizations due to CTD were compared with expected numbers based on national CTD hospitalization rates. To avoid confounding by indication, patients with a hospital discharge diagnosis of a CTD prior to prosthetic surgery were excluded from the cohort. RESULTS: After 4142 person-years of followup in the finger and hand joint cohort, 9 hospitalizations due to CTD were found [standard hospitalization rate (SHR) = 1.5; 95% CI 0.7-2.9]. The TMJ cohort had 1500 person years of followup and 2 hospitalizations due to CTD (SHR = 1.3; 95% CI 0.2-4.5). CONCLUSION: This is the first cohort study to examine the relations between these implants and CTD. Although the number of events was small, this systematic national study revealed no significant or large increase in risk of CTD after finger and hand joint implants or TMJ implants. PMID- 10852267 TI - IgG anti-beta2-glycoprotein I antibodies in adult patients with systemic lupus erythematosus: prevalence and diagnostic value for the antiphospholipid syndrome. AB - OBJECTIVE: To investigate the prevalence of serum anti-beta2-glycoprotein I (anti beta2-GPI) antibodies and other antiphospholipid antibodies (aPL) in patients with systemic lupus erythematosus (SLE). To study their diagnostic value for the antiphospholipid syndrome (APS). METHODS: Anti-beta2-GPI and IgG anticardiolipin (aCL) were determined in sera from 102 consecutive patients with SLE using ELISA. Serum and plasma tests were also done for lupus anticoagulant (LAC), syphilis, and antibodies to dsDNA. Clinical and laboratory features of APS were observed. RESULTS: Prevalences were 23.5% for aCL and 18.6% for anti-beta2-GPI. Correlations between the presence of aCL and anti-beta2-GPI and between their titers were statistically significant (p<0.0001). No associations were found between anti-beta2-GPI and disease activity criteria (SLEDAI, ECLAM, dsDNA). Anti beta2-GPI were significantly associated with LAC (p = 0.005), APS (p = 0.005), and a high aCL titer (aCL > 5 SD; p< or =0.001). LAC was the best diagnostic criterion for APS. CONCLUSION: These data suggest that determination of anti beta2-GPI in addition to aCL and LAC is unlikely to improve the diagnosis of APS in patients with SLE. PMID- 10852268 TI - Serum levels of soluble CD44 in primary Sjogren's syndrome. AB - OBJECTIVE: To determine whether elevated soluble CD44 (sCD44) levels serve as a marker of inflammation and lymphoproliferation in primary Sjogren's syndrome (SS). METHODS: We measured sCD44 levels by ELISA in serum samples from a cross section of healthy individuals and patients seen in a rheumatology clinic for evaluation of possible primary SS. RESULTS: Median serum levels of sCD44 were significantly higher in 48 healthy men compared to 52 healthy women (16 vs. 12 nmol/l; p = 0.0034). There was no relationship between serum levels of sCD44 and age or ethnic background. Slightly higher median levels of sCD44 were found in the serum of 37 women with primary SS compared to healthy women (14 vs. 12 nmol/l; p = 0.0402). However, these levels were comparable to those of 33 female patients without primary SS who were seen in the same clinic (p = 0.1233). CONCLUSION: Serum levels of sCD44 were slightly higher in female patients with primary SS compared to healthy women, but they are not likely to discriminate between patients with and without primary SS in a rheumatology practice. PMID- 10852269 TI - The frequency and significance of anticardiolipin antibodies in scleroderma. AB - OBJECTIVE: To determine the frequency and significance of anticardiolipin antibodies (aCL) in scleroderma. METHODS: We serially tested for aCL in a standardized fashion in patients with scleroderma who were outpatients and gave consent to enter this study. RESULTS: Sixty-three patients participated in this study. Thirty-six had diffuse scleroderma and 27 had limited scleroderma. Three (4.8%) were positive for aCL, of whom 2 had limited scleroderma. In only one patient aCL may have been clinically significant. This woman had limited scleroderma for years and had medium vessel occlusive disease with gangrene eventually requiring midfoot amputations and chronic warfarin treatment. She did not have features of systemic lupus erythematosus or other overlap conditions. The other 2 patients had no manifestations of the antiphospholipid antibody syndrome. CONCLUSION: aCL in scleroderma rarely manifest clinically. The range of frequency of positive aCL in the literature is from 0 to 63%. From our study and the literature, we cannot ascertain if the prevalence is different in limited and diffuse scleroderma. PMID- 10852270 TI - Cellular immune response to Klebsiella pneumoniae antigens in patients with HLA B27+ ankylosing spondylitis. AB - OBJECTIVE: To study the reactivity of peripheral blood mononuclear cells (PBMC) of patients with ankylosing spondylitis (AS) and rheumatoid arthritis (RA) and healthy controls to Klebsiella pneumoniae antigens and to the GroEL-like proteins from K. pneumoniae (HSP60Kp) and Mycobacterium leprae recombinant heat shock protein 65 (rHSP65Ml). METHODS: PBMC of 13 patients with AS and 9 with RA and 10 controls were stimulated in vitro by heat shock induced K. pneumoniae antigens in a cell blot assay, by insolubilized HSP60Kp, by cytosolic proteins (CP) from K. pneumoniae cultivated at 37 degrees C or 45 degrees C, by soluble HSP60Kp, or by rHSP65Ml. RESULTS: In the cell blot assay 7/13 AS and 3/9 RA patients responded to fraction 4, which contains mainly HSP60Kp, and no controls responded (AS vs. controls: p = 0.007). The response to the insolubilized HSP60Kp was positive in 6/13 AS patients but negative in RA patients and controls (p = 0.004). The response to CP45 degrees C was positive in 7/13 AS, in 2/9 RA, and no controls (AS vs controls: p<0.015). Response to the soluble HSP60Kp was found in 7/13 AS and 5/9 RA patients, but no controls (AS vs. controls: p = 0.0075). Response to rHSP65Ml was positive in 3/13 AS, 7/9 RA patients, and 1/10 controls (AS vs RA: p = 0.027; RA vs. controls: p = 0.005; AS vs. controls: nonsignificant). CONCLUSION: In PBMC of the majority of patients with AS and in some with RA, but not in healthy controls, there are cells that proliferate in the presence of HSP60 of K. pneumoniae. PMID- 10852271 TI - Interleukin 6 gene promoter polymorphism is not associated with ankylosing spondylitis. AB - OBJECTIVE: To investigate the possible association between the recently described interleukin 6 (IL-6) promoter polymorphisms at position -174, and susceptibility to ankylosing spondylitis (AS). METHODS: Ninety-two patients with AS, 157 healthy controls, and an additional group of 52 HLA-B27 positive unrelated individuals were included in this study. The -174 polymorphic site in the promoter region of IL-6 gene was typed by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: No statistically significant differences were observed when IL-6 promoter genotype and allele distribution between patients with AS and healthy controls were compared. CONCLUSION: Our results suggest the -174 IL-6 polymorphism does not play an important role in susceptibility to AS. Larger studies are needed to provide more conclusive evidence on the role of -174 IL-6 polymorphism in AS. PMID- 10852272 TI - Double blind randomized clinical trial examining the efficacy of bupivacaine suprascapular nerve blocks in frozen shoulder. AB - OBJECTIVE: To determine whether the pain, contracture, and disability associated with idiopathic frozen shoulder are diminished by a series of 3 indirect bupivacaine suprascapular nerve blocks delivered in an ambulatory care clinic. METHODS: A double blind randomized controlled trial of patients referred by primary care and specialty clinics in Montreal to an ambulatory tertiary care academic facility. Patients and controls underwent a series of 3 indirect suprascapular nerve blocks at 7 day intervals using either 10 c.c. bupivacaine 0.5 (Marcaine) in the treatment group or 10 c.c. of physiological saline in controls. Subjects in both groups were taught a program of shoulder range of motion exercises to be done at home. The primary outcome measure was the McGill Melzack Pain Questionnaire (MPQ) short form at 1 month post-randomization (2 weeks after last injection). The secondary outcome measures were disability measured by the simple shoulder test and glenohumeral joint contracture measured by shoulder range of motion measurements. RESULTS: Thirty-four subjects were randomized from 58 screened. Average age of subjects was 52 years. Mean duration of pain prior to randomization was one year. Dropout rate was 11% in the treatment group, 30% in the placebo group. A 64% reduction in pain in the treatment group versus 13% in the placebo group was observed at one month as measured by the MPQ multidimensional pain descriptors score (p = 0.03). A nonsignificant 15.8% improvement in shoulder function in the treatment group versus 4% in the placebo group (p = 0.24) was also noted. No improvement in shoulder range of movement was noted. No side effects other than transient vagal symptoms and local tenderness at the injection site were reported. CONCLUSION: The use of bupivacaine suprascapular nerve blocks was effective in reducing the pain of frozen shoulder at one month. Clinical studies with a larger number of subjects and a longer study period will help determine the duration and nature of the effect of bupivacaine suprascapular nerve blocks in treating the pain, disability, and glenohumeral joint contracture of frozen shoulder. PMID- 10852273 TI - Corticosteroid injections in polymyalgia rheumatica: a double-blind, prospective, randomized, placebo controlled study. AB - OBJECTIVE: To determine the efficacy and safety of shoulder corticosteroid injections in polymyalgia rheumatica (PMR). METHODS: Twenty consecutive patients with active PMR were randomized into a 7 month, double blind, placebo controlled study. Patients received either bilateral shoulder injections of 40 mg of 6 methylprednisolone acetate or placebo (1 ml saline solution). Responders were treated weekly with the same regimen for a total of 4 bilateral injections and then followed for 6 months. Response was defined as a 70% reduction in visual analog scale (VAS) score for pain and for patient and physician global assessment, and duration of morning stiffness. Bilateral shoulder magnetic resonance imaging (MRI) was performed at different times to evaluate the response of lesions to therapy. RESULTS: All 10 corticosteroid treated patients responded to the first injection with a significant reduction in duration of morning stiffness, VAS pain scale, patient and physician global assessment, erythrocyte sedimentation rate, and C-reactive protein. Interleukin 6 serum levels were significantly reduced after the 2nd injection. In 5 patients, the response persisted throughout the followup period. The other 5 withdrew within 4 weeks after the 4th injection due to recurrence of symptoms. None of the 10 patients of the placebo group responded to the first injection. The difference between the 2 groups was significant (p = 0.03). No side effects were recorded. MRI showed marked improvement of shoulder lesions one week after first injection and an almost complete resolution one week after last injection in the responders. CONCLUSION: Shoulder corticosteroid injections seem to be an effective and safe therapy for PMR. PMID- 10852274 TI - Intravenous clodronate in the treatment of reflex sympathetic dystrophy syndrome. A randomized, double blind, placebo controlled study. AB - OBJECTIVE: To evaluate the efficacy of intravenous (i.v.) clodronate in patients with reflex sympathetic dystrophy syndrome (RSDS) and to assess the urinary excretion of type I collagen crosslinked N-telopeptide (NTx) before and after the treatment. METHODS: Thirty-two patients with RSDS were randomized to receive either i.v. clodronate 300 mg daily for 10 consecutive days or placebo. Forty days later, the placebo treated patients received the clodronate treatment. Outcome measures included as a primary endpoint the visual analog scale of pain (VAS, range 0-100); secondary endpoints were a clinical global assessment (CGA, range 0-3) and an efficacy verbal score (EVS, range 0-3). Clinical and biochemical assessments were performed before the treatment, 40 (T40), 90 (T90), and 180 (T180) days later. RESULTS: At T40 the 15 patients randomized to clodronate treatment showed significant decreases of VAS and CGA (p = 0.002, p = 0.001, respectively). Compared with the placebo group (17 patients), significant differences were found in all clinical variables (VAS: p = 0.001; CGA: p = 0.001; EVS: p<0.0001). A further clinical improvement was observed throughout the study. Pooling the results of all 32 patients after clodronate treatment, at T180 the overall percentage decrease of VAS was 93.2+/-15.6%, with 30 patients significantly improved or asymptomatic. Significant inverse correlations between baseline NTx values and decreases of VAS were found at T90 (p = 0.03) and T180 (p = 0.01). No adverse events related to treatment occurred. CONCLUSION: A 10 day i.v. clodronate course is better than placebo and effective in the treatment of RSDS. NTx seems to be a predictive factor for clodronate efficacy. PMID- 10852275 TI - A randomized, multicenter, controlled trial using intravenous pulses of methylprednisolone in the initial treatment of simple forms of giant cell arteritis: a one year followup study of 164 patients. AB - OBJECTIVE: (1) To evaluate the corticosteroid sparing effect of an initial intravenous (i.v.) pulse of methylprednisolone (MP) in the treatment of simple forms of giant cell arteritis (GCA). (2) To analyze corticosteroid response, steroid related side effects, and GCA complications. METHODS: Patients received a 240 mg i.v. pulse of MP followed by 0.7 mg/kg/day oral prednisone (Group 1) or 0.7 mg/kg/day prednisone without an i.v. pulse (Group 2, controls), or a 240 mg i.v. pulse of MP followed by 0.5 mg/kg/day prednisone (Group 3). Corticosteroid dosage was reduced after normalization of 2 biological inflammatory variables to obtain half-dosage after 4 weeks in Groups 1 and 2 and 20 mg/day after 2 weeks in Group 3. Tapering was systematically attempted from the 6th month of treatment. RESULTS: One hundred sixty-four patients were included in the trial (1992-96). Cumulative doses of corticosteroids after one year were identical for all groups (p = 0.39). No significant differences were observed in the time required for normalization of C-reactive protein, corticosteroid resistance (13.5%), and corticosteroid related side effects (39% of patients; p = 0.37). Corticosteroid resistant patients received larger doses and showed a high risk of GCA related complications (p = 0.02). CONCLUSION: MP pulses have no significant longterm, corticosteroid sparing effects in the treatment of simple forms of GCA and should be limited to complicated forms. Moreover, corticosteroid resistance is a real risk factor for GCA complications. PMID- 10852276 TI - Familial Mediterranean fever: high gene frequency and heterogeneous disease among an Israeli-Arab population. AB - OBJECTIVE: Familial Mediterranean fever (FMF) is an autosomal recessive disease that primarily affects non-Ashkenazi Jews, Armenians, Arabs, and Turks. The FMF (MEFV) gene responsible for the disease has been recently identified. Four missense mutations in exon 10 of the FMF gene seem to account for 86% of the DNA variations identified in patients with FMF. We conducted a phenotype/genotype correlation study in a homogenous population of Israeli-Moslem Arab patients with FMF and performed a mutational screening analysis on DNA samples from healthy individuals of this ethnic group. METHODS: Sixty-five patients clinically diagnosed as having FMF underwent molecular genetic studies using polymerase chain reaction and restriction endonuclease digestion methods to detect the presence of the 4 mutations (M694V, V726A, M680I, M694I). We then correlated the presence of each mutation with age of onset, clinical manifestations, and disease severity; patients whose allelic combination included M694V were then excluded from further statistical analysis, since the association of severe disease with the M694V allele has already been shown. In addition, we screened for FMF mutations the DNA samples from 318 healthy Moslem Arab individuals for the presence of these mutations. RESULTS: Among the 65 patients who were clinically diagnosed as having FMF, 78.5% had one or 2 mutation-bearing chromosomes. The most prevalent mutation was V726A, followed by M680I, M694V, and M6941. No significant difference in phenotypic characteristics was found between the patients with the diverse mutations. The total carrier frequency for the 4 mutations was 10.4% (95% confidence interval 0.07 to 0.137). CONCLUSION: A high FMF gene frequency was found among an Israeli-Moslem Arab population. Among the FMF patients from this ethnic group, several mutations were detected, none of which was found to correlate with a severe course of the disease. PMID- 10852277 TI - Patterns of medication use before and after bone densitometry: factors associated with appropriate treatment. AB - OBJECTIVE: We examined the medications used by women before and after bone densitometry to determine whether patient or physician factors were associated with appropriate osteoporosis therapy. METHODS: Appropriate osteoporosis treatment was defined as alendronate, etidronate, calcitonin, or hormone replacement therapy (HRT) for women with any bone mineral density (BMD) t score < -2.5 or no osteoporosis therapy, except HRT, for women with t scores > -1.0. We observed a cohort of women who underwent bone densitometry at one outpatient osteoporosis clinic. Medical history, medication use, and demographic data were collected at the time of bone densitometry. A followup questionnaire assessed the medication use patterns since bone densitometry and attitudes about osteoporosis therapy. RESULTS: We recruited 553 women who underwent bone densitometry in 1996. Their mean age was 62 years and 95% were postmenopausal. Prior to bone density scans, 27% of patients used HRT, 15% used bisphosphonates, and 6% used calcitonin. Scan results and surveys revealed that 40% of patients had BMD below a t score of -2.5 at any site. Of women with osteoporosis 78% reported taking an appropriate medication after their scans. Patients most likely to receive appropriate treatment were those who understood their bone densitometry results (odds ratio, OR, 2.5; 95% confidence interval, CI, 1.3 to 4.8) and patients who were taking an osteoporosis medication (OR 1.9; 95% CI 1.0 to 3.6). Neither the specialty of the referring physician nor patients' medical history was associated with use of appropriate osteoporosis therapy. CONCLUSION: Of women with osteoporosis who underwent bone densitometry 78% received appropriate therapy after this test. Patient factors were associated with the likelihood that they received appropriate therapy, suggesting that strategies aimed at educating patients may improve the use of osteoporosis medications. PMID- 10852278 TI - The interaction between uric acid level and other risk factors on the development of gout among asymptomatic hyperuricemic men in a prospective study. AB - OBJECTIVE: To investigate the incidence of gout and the interaction between uric acid level and other risk factors in the development of gout. METHODS: Two hundred twenty-three asymptomatic hyperuricemic men initially studied in 1991-92 were reassessed in 1996-97. Gout was clinically diagnosed by a senior rheumatologist based on history and physical according to the clinical criteria of Wallace. Basic demographic and lifestyle variables as well as biochemical data were collected in both baseline and followup periods. Both the stability analysis and the analysis of repeated relationships were applied. RESULTS: The 5-year cumulative incidence of gout was 18.83% (42/223). The risk factors for gout based on the analysis of repeated relationships were uric acid level, alcohol consumption, use of diuretics, and obesity. The only predictor of gout at baseline was uric acid level. After adjusting for baseline uric acid level, followup uric acid increase, persistent alcohol consumption, use of diuretics in the followup period, and body mass index increase were independent predictors for gout among asymptomatic hyperuricemic men. Excessive alcohol consumption, particularly if occasional, was the most important factor in the development of gout, even when the concentration of uric acid level was below 8 mg/dl. CONCLUSION: Uric acid level is the key factor for prevention of gout and needs constant monitoring. Other contributing or possible etiologic factors such as alcohol consumption, diuretics use, and excess weight gain carry an increased risk of gout attack among patients with hyperuricemia. PMID- 10852279 TI - A Canadian survey on the management of corticosteroid induced osteoporosis by rheumatologists. AB - OBJECTIVE: To survey the practice pattern of Canadian rheumatologists (CR) on their management of corticosteroid induced osteoporosis in their premenopausal (PrM) and postmenopausal (PoM) female patients. METHODS: The practice pattern was surveyed using a 17 item questionnaire probing the diagnosis, prevention, treatment, and monitoring of osteoporosis in PrM and PoM women receiving longterm oral systemic corticosteroid therapy. RESULTS: Most CR investigated and treated osteoporosis themselves, 13% referred to other specialists for investigation, and 22% referred for treatment. Eighty-two percent of CR used dual energy x-ray absorptiometry (DEXA) to confirm a diagnosis of osteoporosis. Most CR initiated investigation for osteoporosis at the start or within the first year of starting longterm systemic corticosteroid therapy: PrM 87% and PoM 93%. The most frequently used initial strategy for the prevention of osteoporosis was as follows. PrM: calcium and vitamin D3 (53%); PoM: hormone replacement therapy (HRT) and calcium (29%). The most common initial choice for treatment of established osteoporosis was as follows: PrM: etidronate (53%); PoM: bisphosphonates +/- HRT (53%). Ninety-six percent of CR used only bone mineral density (BMD) measurement to monitor therapy for corticosteroid induced osteoporosis. Most CR monitored BMD every 12 to 24 months for PrM (81%) and PoM (84%). The BMD parameter(s) (T and Z scores as measured by DEXA) used to initiate therapy for corticosteroid induced osteoporosis was variable. CONCLUSION: It appears that, while certain trends are evident, there is still considerable variability in the management of corticosteroid induced osteoporosis. PMID- 10852280 TI - Analysis of the discordance between radiographic changes and knee pain in osteoarthritis of the knee. AB - OBJECTIVE: To analyze cross sectional data from the National Health and Nutrition Examination Survey (NHANES I) concerning 3 indicators of osteoarthritis (OA) of the knee: radiographic evidence of structural damage, self-reported knee pain, and self-report of a diagnosis of arthritis at any joint by a physician. METHODS: Analysis of NHANES I data for 6880 persons ages 25-74 in the United States. RESULTS: Radiographic stage 2-4 knee OA was found in 319 subjects (3.7%); only 47% of these individuals reported knee pain, and only 61% reported that a physician had told them that they had arthritis. Knee pain was reported by 1004 subjects (14.6%), only 15% of whom had radiographic stage 2-4 changes of OA, and 59% of whom reported having a diagnosis of arthritis by a physician. A report of arthritis diagnosed by a physician was given by 1762 subjects (25.6%), of whom only 11% had stage 2-4 radiographic knee OA and 34% reported knee pain. CONCLUSION: Substantial discordance exists in this population based study between radiographic OA of the knee versus knee pain, versus a diagnosis of arthritis by a physician. These phenomena may be important in the design of clinical research studies, as well as in criteria for OA. PMID- 10852281 TI - Association of HLA-DR7 with rheumatic fever in the Brazilian population. AB - OBJECTIVE: Rheumatic fever (RF) is a multisystem inflammatory disease that develops as a sequel of untreated throat infection by the group A beta-hemolytic streptococcus. As HLA antigens are known to be important in controlling immunological responsiveness, studies have investigated HLA antigen association with RF. Studies with Caucasians, Black Americans, and Indians showed associations with HLA-DR4, DR2, and DR3, respectively. One study on a Brazilian population suggested an association with HLA-DR7 and HLA-DR53. We investigated the association between RF and antigens HLA-DR7 and DR53 in the white Brazilian population. METHODS: Thirty-five patients and 209 healthy individuals living in the northern region of the state of Parana, Brazil, were used as test and control groups, respectively. Classical statistical methods were used to compare HLA frequencies between these groups. Results. Data confirmed positive association with HLA-DR7 (46.7 vs. 25.7%; p = 0.015), but not with HLA-DR53 (54.3 vs. 44.5%; p = 0.28). The relative risk and etiologic fractions were 2.4 and 0.27%, respectively. CONCLUSION: Positive association between HLA-DR7 specificity and RF was observed in the white Brazilian population by 2 independent studies, supporting the hypothesis of the involvement of genetic factors in susceptibility of rheumatic fever. PMID- 10852282 TI - The prevalence of pain complaints in a general population in Israel and its implications for utilization of health services. AB - OBJECTIVE: To determine the prevalence of pain complaints, specifically of chronic widespread pain, in the general population; and to explore the utilization of health services by various pain groups. METHODS: Cross sectional population survey of 2210 adults in the southern part of Israel, who were classified into 5 pain groups: no pain, transient pain, chronic regional pain, chronic widespread pain, and other. Participants were interviewed about pain patterns and utilization of health services. RESULTS: Forty-four percent reported pain on the day of the interview. The prevalence of chronic widespread pain in the study population was 9.9%, 14% in women and 3% in men (p<0.01). The prevalence in the Israeli adult population was estimated after adjusting for sex and age as 10.2%. The prevalence of any chronic pain (regional or widespread) increased with age. The prevalence of chronic widespread pain was significantly higher in women than in men across all age groups (p<0.01). Persons with chronic widespread pain reported most frequent visits to their physicians (10.8 visits/year) and most frequent use of antiinflammatory and analgesic drugs. They were more frequently referred to specialists and tended to be hospitalized slightly more often. CONCLUSION: In the general population in Israel, widespread pain is common, and its prevalence is comparable with reports from USA, UK, and Canada. PMID- 10852283 TI - Follicular phase hypothalamic-pituitary-gonadal axis function in women with fibromyalgia and chronic fatigue syndrome. AB - OBJECTIVE: Fibromyalgia (FM) and chronic fatigue syndrome (CFS) are clinically overlapping stress associated disorders. Neuroendocrine perturbations have been noted in both syndromes, and they are more common in women, suggesting abnormalities of gonadal steroid hormones. We tested the hypothesis that women with FM and CFS manifest abnormalities of the hypothalamic-pituitary-gonadal (HPG) hormonal axis. METHODS: We examined the secretory characteristics of estradiol, progesterone, follicle stimulating hormone (FSH), and luteinizing hormone (LH), including a detailed analysis of LH in premenopausal women with FM (n = 9) or CFS (n = 8) during the follicular phase of the menstrual cycle compared to matched healthy controls. Blood was collected from an indwelling intravenous catheter every 10 min. over a 12 h period. LH was assayed from every sample; pulses of LH were identified by a pulse-detection program. FSH and progesterone were assayed from a pool of hourly samples for the 12 h period and estradiol from samples pooled over four 3 h time periods. RESULTS: There were no significant differences in FSH, progesterone, or estradiol levels in patients versus controls. There were no significant differences in pulsatile secretion of LH. CONCLUSION: There is no indication of abnormal gonadotropin secretion or gonadal steroid levels in this small, but systematic, study of HPG axis function in patients with FM and CFS. PMID- 10852284 TI - Musculoskeletal examination teaching in rheumatoid arthritis education: trained patient educators compared to nonspecialist doctors. AB - OBJECTIVE: To assess whether students taught by trained patients acquire the same levels of competence in musculoskeletal examination skills as students taught by nonspecialist doctors. METHODS: Year 1 Graduate Medical Program (GMP) students (N = 25) at Hospital A were randomized to 8 tutorial groups, each comprising 3-4 students. Groups were taught hand and wrist examination skills by patient educators trained by the Searle Patient Partners in Arthritis program (patient partners). Students at Hospitals B and C (N = 12) remained in their normal tutorial groups, each comprising 3-4 students. Groups at Hospitals B and C were taught hand and wrist examination skills by doctors who had no specialized training in musculoskeletal medicine or orthopedics, with an untrained patient present. RESULTS: Students' mean self-ratings of examination skills before and students' patient partners, and consultants' mean ratings of students' examination skills after the tutorial were summed. Before the tutorial there were no significant differences in level of skill between students at Hospitals A, B, and C as measured by students' self-ratings. After the tutorial all students showed clear gains in levels of skill. Students taught by patient partners had higher levels of skill than those taught by doctors for 3 (p<0.01) and 4 (p<0.05) out of 14 examination skills and all 4 communication skills (p<0.05), as measured by both patient partners' and consultants' ratings. Students taught by doctors showed higher levels of skill for 2 observation skills, but these differences were not significant. CONCLUSION: Patient partners are either equal or superior to doctors not specifically trained in musculoskeletal medicine or orthopedics, in the teaching of musculoskeletal examination techniques and basic communication skills. PMID- 10852285 TI - Evaluation of patient partners in the teaching of the musculoskeletal examination. AB - OBJECTIVE: To evaluate student preferences and examination outcomes of 2 different methods of teaching musculoskeletal (MSK) medicine examination techniques. METHODS: Year 2 students in a 4 year graduate medical school were randomized to 3 teaching groups: students in Group 1 were taught by rheumatology fellows, Group 2 by patient partners, while group 3 were randomly allocated to either patient partner (3A) or rheumatology trainee (3B) teaching. All students were debriefed at the end of each teaching block of 4 weeks for feedback on their teaching experience using a standardized questionnaire. In addition, at the end of the academic year, all students sat an objective structured clinical examination (OSCE) in clinical skills that contained a rheumatology station. The effect of method of teaching on students' performance in the rheumatology station was analyzed. RESULTS: Eighty medical students participated in the study. Overall, there was no difference in student ratings of either mode of teaching, although the students reported that patient partner teaching gave more opportunity to practice MSK examination skills and also provided greater feedback to the student. Students reported a preference for rheumatology trainee teaching because they believed the teaching would be more relevant to the content of the examination. There was no statistically significant difference in the performance of the students in the OSCE rheumatology station in regard to the mode of teaching they had received prior to the examination. CONCLUSION: Patient partner teaching is as effective a method of teaching clinical skills in MSK medicine as a traditional resident based form of teaching, with student benefits from patient feedback and greater "hands-on" opportunities. The assessment process should incorporate patient partners to assess the unique aspects of patient educator based teaching of MSK examination techniques. PMID- 10852286 TI - Evidence of subclinical intestinal inflammation by 99m technetium leukocyte scintigraphy in patients with HLA-B27 positive juvenile onset active spondyloarthropathy. AB - OBJECTIVE: The concept that gut inflammation is implicated in the pathogenesis of spondyloarthropathies (SpA) has long been considered. Subclinical intestinal inflammation has been reported in adult patients with SpA by histological examination of intestinal biopsies. We assessed the presence of gut inflammation by abdominal 99mTc-hexamethyl propylene amine oxime (99mTc-HMPAO) labeled leukocyte scintigraphy in a group of children and adolescents with HLA-B27 positive SpA without gastrointestinal (GI) symptoms, and correlated the scintigraphic results to disease activity. METHODS: Abdominal scintigraphy with 99mTc-HMPAO labeled leukocytes was performed in 27 HLA-B27 positive children and adolescents with SpA without GI symptoms. Patients were divided into 2 groups according to the presence of active or inactive joint disease: Group A, 17 patients with active disease, and Group B, 10 patients with inactive disease. Patients with positive abdominal scintigraphy underwent complete bowel investigation by means of small bowel barium follow-through, abdominal ultrasound scan, and ileocolonoscopy with mucosal biopsies. RESULTS: Thirteen of 27 patients (48%) had scintigraphy indicating the presence of bowel inflammation. All patients with abnormal scan had active joint disease, whereas no patient with inactive disease had a positive intestinal uptake of labeled leukocytes. Bowel investigation revealed the presence of aspecific mucosal inflammatory changes in the majority of patients with positive scintigraphy. CONCLUSION: The presence of intestinal leukocyte uptake only in patients with active joint disease, even if intestinal inflammatory changes were minimal and clinical gut manifestations were absent, supports the role of gut inflammation in the pathogenesis of joint disease in HLA-B27 positive patients with SpA. PMID- 10852287 TI - Mycophenolate mofetil in the treatment of severe skin manifestations of dermatomyositis: a series of 4 cases. AB - Four cases with classic skin manifestations and histologic evidence of dermatomyositis are presented. No occult malignancies were found. After conventional therapy with corticosteroids, hydroxychloroquine and/or methotrexate proved to be limited by side effects or an inadequate clinical response, a therapeutic trial of mycophenolate mofetil was instituted. This relatively new drug, which inhibits lymphocyte proliferation, was effective [with a mean duration of treatment of 13 months (range 6-20)] at controlling cutaneous disease activity, resulting in a decrease of the steroid dose. PMID- 10852288 TI - Primary cutaneous B cell lymphoma during methotrexate therapy for rheumatoid arthritis. AB - A patient with rheumatoid arthritis developed a reversible, primary cutaneous, large B cell lymphoma during prolonged methotrexate (MTX) treatment. Regression of the skin lesions after discontinuation of the drug suggested a close relationship to MTX. Increased clinical awareness, discontinuation of MTX, and close observation are important in the initial management of this rare lymphoproliferative disorder. PMID- 10852289 TI - "Coated aorta": a new sign of Erdheim-Chester disease. AB - Erdheim-Chester disease is a rare, non-Langerhans cell form of histiocytosis characterized by osteosclerosis of the metaphyseal regions of long bones, diabetes insipidus, proptosis, and retroperitoneal fibrosis. The latter usually involves the perirenal area and leads to hydronephrosis. Periaortic fibrosis is less frequent. We describe 3 unusual cases of Erdheim-Chester disease with periaortic fibrosis involving the whole aorta and leading to a "coated aorta" appearance on computed tomography scans. Faced with such a singular "coated aorta," bone scintigraphy can be very helpful when searching for Erdheim-Chester disease. PMID- 10852290 TI - Splenic infarction in Wegener's granulomatosis. AB - We describe a case of severe Wegener's granulomatosis associated with asymptomatic splenic infarction. The 4 previous case reports are reviewed and the implication of this finding for preventive strategies is highlighted. PMID- 10852291 TI - Skeletal muscle pathology in 2 siblings infected with Toxoplasma gondii. AB - Skeletal muscle can be the site of inflammatory diseases that lead to muscle weakness, pain, and increased myogenic serum enzymes. Most of these inflammatory myopathies are idiopathic. In some cases inflammatory myopathies are due to infectious agents. We describe the pathological aspects of muscle biopsies of 2 Brazilian siblings who acquired toxoplasmosis at the same time and in similar conditions. One developed a tetraplegia that was confirmed to be due to inflammatory myositis due to toxoplasma. The other developed myocarditis, with heart failure, without skeletal muscle weakness. In both cases many toxoplasma organisms were observed in the muscle biopsies, but in case 1 only was there an inflammatory myopathy with myofiber necrosis; the inflammatory cells were predominantly macrophages with some CD4+ cells and rare CD20+ cells. In case 1, expression of CD54 was observed in many inflammatory cells as well in endothelial cells, but only in endothelial cells in case 2. After treatment with clindamycin and corticosteroids both cases had only partial improvement, case 1 with a residual muscle weakness and case 2 with residual cardiac insufficiency (requiring digoxin). These cases show that the presence of the parasite in myofibers is not enough to induce an inflammatory myositis with muscle cell necrosis. This suggests that immunological disturbances may contribute to the development of inflammatory myositis due to toxoplasma. PMID- 10852292 TI - Intraarticular osteoid osteoma. PMID- 10852293 TI - Primary hypertrophic osteoarthropathy. PMID- 10852294 TI - Ultrasonography and psoriatic arthritis. PMID- 10852295 TI - Reducing serologic tests for rheumatic diseases. PMID- 10852296 TI - Cost effectiveness analysis--assessing the assumptions behind the assumptions. PMID- 10852297 TI - Minocycline induced lupus and autoimmune hepatitis. PMID- 10852298 TI - Revision of the proposed classification criteria for juvenile idiopathic arthritis: Durban, 1997. PMID- 10852299 TI - High prevalence of Helicobacter pylori type I virulent strains in patients with systemic sclerosis. PMID- 10852300 TI - Bacillus Calmette-Guerin induced reactive arthritis. PMID- 10852301 TI - Inhibition of collagenase by a bisphosphonate-group drug in patients with rheumatoid arthritis. PMID- 10852302 TI - Adverse reaction to Hylan GF-20. PMID- 10852303 TI - Remission of rheumatoid arthritis with taxol in a patient with breast carcinoma. PMID- 10852304 TI - Use of alternative drug therapies by patients with rheumatoid arthritis in Korea. PMID- 10852305 TI - Nailfold videocapillaroscopy abnormalities in patients with antiphospholipid antibodies. PMID- 10852306 TI - Relevance of the chlorophyll phytyl chain on lamellar phase formation and organisation. AB - A series of modified chlorophylls (chlorophyll a, pyrochlorophyll a, Zn pheophytin a and Zn-pheophorbide a) have been inserted into lamellar phases of sodium bis-(2-ethylhexyl)-sulfosuccinate (AOT). The role played by the different functional groups in affecting the bilayer formation and organisation has been investigated by means of the NMR quadrupolar splitting technique. Evidence is reported for the first time on the capacity of the phytyl chain of the chlorophylls to anchor the tetrapyrroles into the bilayer, favouring at the same time the regular formation of the lamellae. PMID- 10852307 TI - Interaction between cell shape and contraction pattern in the Physarum plasmodium. AB - The relationship between cell shape and rhythmic contractile activity in the large amoeboid organism Physarum polycephalum was studied. The organism develops intricate networks of veins in which protoplasmic sol moved to and fro very regularly. When migrating on plain agar, the plasmodium extends like a sheet and develops dendritic veins toward the rear. After a particular stimulation, the vein organization changes into veinless or vein-network structures. In both structures, the mixing rate of the protoplasm, which is related to communication among contraction oscillators, decreased compared with that of the dendritic one. Accompanying these changes in vein structure, the spatio-temporal pattern of the rhythmic contraction changed into a small-structured pattern from a synchronized one. In the above process, cell shape affects the contraction pattern, but, conversely, the contraction pattern effects the cell shape. To demonstrate this, a phase difference in the rhythmic contraction was induced artificially by entraining the intrinsic rhythm to external temperature oscillations. New veins then formed along the direction parallel to the phase difference of the rhythm. Consequently, the vein organization of the cell interacts with the contractile activity to form a feedback loop in a mechanism of contraction pattern formation. PMID- 10852308 TI - Formation and structural determinants of multi-stranded guanine-rich DNA complexes. AB - We have investigated the complexes formed by oligonucleotides with the general sequence d(T15,Gn), where n = 4-15. Two distinct classes of structures are formed, namely, the four-stranded tetraplex and frayed wires. Frayed wires differ from four-stranded tetraplexes in both strand association stoichiometry and the ability of dimethyl sulfate to methylate the N7 position of guanine. Thus, it appears that these two guanine-rich multistranded assemblies are stabilised by different guanine-guanine interactions. The number of contiguous guanine residues determines which of the complexes is favoured. Based on the stoichiometry of the associated species and the accessibility of the N7 position of guanine to methylation we have found that oligonucleotides with smaller number of contiguous guanines; n = 5-8, form primarily four-stranded tetraplex. Oligonucleotides with larger numbers of contiguous guanines adapt primarily the frayed wire structure. The stability of the complexes formed by this series of oligonucleotides is determined by the number and arrangement of the guanines within the sequences. We propose that the formation of the two types of complex proceed by a parallel reaction pathways that may share common intermediates. PMID- 10852309 TI - Interpretation of thermodynamic non-ideality in sedimentation equilibrium experiments on proteins. AB - This investigation re-examines theoretical aspects of the allowance for effects of thermodynamic non-ideality on the sedimentation equilibrium distribution for a single macromolecular solute, and thereby resolves the question of the constraints that pertain to the definition of the activity coefficient term in the basic sedimentation equilibrium expression. Sedimentation equilibrium results for ovalbumin are then presented to illustrate a simple procedure for evaluating the net charge (valence) of a protein from the magnitude of the second virial coefficient in situations where the effective radius of the protein can be assigned. Finally, published sedimentation equilibrium results on lysozyme are reanalysed to demonstrate the feasibility of employing the dependence of the second virial coefficient upon ionic strength to evaluate both the valence and the effective radius of the non-interacting solute. PMID- 10852310 TI - Interaction of alkaline-earth metal ions with calf thymus DNA. Volume and compressibility effects in diluted aqueous solutions. AB - The binding of Mg2+, Ca2+, Sr2+ and Ba2+ ions to calf thymus DNA in solutions has been investigated by ultrasonic and densimetric techniques. The obtained parameters, the apparent molar volume, phiV, and the apparent molar adiabatic compressibility, phiK(S), are very sensitive to hydration of investigated molecules. The interaction between the cations and DNA is accompanied by overlapping their hydration shells and consequently releasing the water molecules from hydration shells to bulk state. The change in the hydration is reflected in the measured parameters, phiV and phiK(S). The magnitude of these hydration changes is determined by the position of the cation relative to DNA atomic groups involved in the binding, and thus can characterize the structure of cation-DNA complexes. The values of the dehydration effects of the binding, deltaphiV and deltaphiK(S), correspond to two direct or higher number of indirect contacts between calf thymus DNA and the cations. PMID- 10852311 TI - A study on the enthalpy-entropy compensation in protein unfolding. AB - A large number of thermodynamic data including the free energy, enthalpy, entropy, and heat capacity changes were collected for the denaturation of various proteins. Regression indicated that remarkable enthalpy-entropy compensation occurred in protein unfolding, which meant that the change in enthalpy was almost compensated by a corresponding change in entropy resulting in a smaller net free energy change. This behavior was proposed to result from the water molecule reorganization, which contributed significantly to the enthalpy and entropy changes but little to the free energy change in protein unfolding. It turned out that the enthalpy-entropy compensation could provide novel insights into the problem of enthalpy and entropy convergence in protein unfolding. PMID- 10852312 TI - 2,3-DPG-Hb complex: a hypothesis for an asymmetric binding. AB - This study was undertaken to test the symmetry of 2,3-diphosphoglycerate (2,3 DPG) binding site in hemoglobin (Hb). From Arnone's study [A. Arnone, Nature (London) 237 (1972) 146] the 2,3-DPG binding site is located at the top of the cavity, that runs through the center of the deoxy-Hb molecule. However, it is possible that this symmetry reported by Arnone, for crystals of 2,3-DPG-Hb complex, might not be conserved in solution. In this paper, we report the 31P nuclear magnetic resonances of the 2,3-DPG interaction with Hb. The 2,3-DPG chemical shifts of the P2 and P3 resonance are both pH- and hemoglobin-dependent [protein from man, polar bear (Ursus maritimus), Arctic fox (Alopex lagopus) and bovine]. 2,3-DPG binds tightly to deoxyhemoglobin and weakly, nevertheless significantly, to oxyhemoglobin. In particular, our results suggest similar spatial position of the binding site of 2,3-DPG in both forms of Hb in solutions. However, the most unexpected result was the apparent loss of symmetry in the binding site, which might correlate with the ability of the hemoglobin to modulate its functional behavior. The different interactions of the phosphate groups indicate small differences in the quaternary structure of the different deoxy forms of hemoglobin. Given the above structural perturbation an asymmetric binding in the complex could justify, at least in part, different physiological properties of Hb. Regardless, functionally relevant effects of 2,3-DPG seem to be measured and best elucidated through solution studies. PMID- 10852313 TI - Reconstitution of the Golgi reassembly process in semi-intact MDCK cells. AB - The Golgi apparatus, which consists of stacks of cisternae during interphase, is fragmented or dispersed throughout the cytoplasm at the onset of mitosis. A sea sponge metabolite, ilimaquinone (IQ), causes Golgi membranes to vesiculate. And after its removal, the vesiculated membranes reassemble into stacks of cisternae in the perinuclear region. To study the mechanism of Golgi membrane dynamics during mitosis, we have reconstituted the reassembly process of IQ-induced vesiculated Golgi membranes in streptolysin O-permeabilized Mardin-Darby canine kidney (MDCK) cells. Monitoring the dynamics of Golgi membranes labeled with a green fluorescence protein (GFP)-tagged protein, we dissected the process into two elementary components: the reassembly of vesiculated Golgi membranes into punctate structures; and the subsequent reformation of these structures into stacks of cisternae near the nucleus. Using morphometric analysis, we studied the kinetics and biochemical requirements for the process, and revealed that an NEM sensitive factor, cytoplasmic dynein, and GTP binding protein were involved in the Golgi reassembly. PMID- 10852314 TI - Compared effects of cholesterol and 7-dehydrocholesterol on sphingomyelin glycerophospholipid bilayers studied by ESR. AB - The ESR of 7- and 16-doxylstearic spin-labeled fatty acids (7NS and 16NS, respectively) reveal the distinct influence of cholesterol or cholesterol precursor analogue, delta7-dehydrocholesterol, on the molecular ordering and the fluidity of lipid mixtures containing sphingomyelin (SM). The phase-separation of sphingomyelin domains mixed within fluid glycerophospholipids (phosphatidylethanolamine and phosphatidylserine) can be followed by ESR as a function of the temperature and in the presence of sterols [cholesterol (CHOL) or 7-dehydrocholesterol (DHCHOL)]. The time scale of spin-label exchange among phases is appropriate to follow the occurrence of the specific sphingomyelin/sterol association forming liquid ordered (Lo) microdomains which separate from the fluid surrounding phase Lalpha. Sphingomyelin embedded within the fluid bilayer associates with both sterols below 36 degrees C to give a phase Lo traceable by ESR in the form of a highly anisotropic component. Above 36 degrees C, the contribution in the ESR spectrum, of the Lo phase formed by 7 dehydrocholesterol with sphingomyelin is reduced by contrast with cholesterol forming a temperature-stable liquid ordered phase up to 42 degrees C. The consequences of this destabilization of the SM/sterol microdomains are envisioned in the biosynthesis defect where the precursor 7-dehydrocholesterol substitutes, for a significant part, the embryonic cell cholesterol. PMID- 10852315 TI - Long-term registration has improved the quality of hip replacement: a review of the Swedish THR Register comparing 160,000 cases. AB - The Swedish Hip Register has defined the epidemiology of total hip replacement in Sweden. Most hip implants are fully cemented. Serious complications and rates of revision have declined significantly despite an increasing number of patients at risk. During the past 5 years, only 8-9% of hip replacements are revisions. Aseptic loosening with or without osteolysis is the major problem and constitutes 71% of the revisions, but the incidence had decreased three times during the past 15 years to less than 3% at 10 years. The effectiveness of the surgical technique is the most important factor for reducing the risk of revision because of aseptic loosening, but choice of implant is also important. In practice, total hip replacement in Sweden has improved, as judged by information from this Register about individualized patient risks, implant safety, and the greater efficacy of surgical and cementing techniques. PMID- 10852316 TI - A systematic survey of 13 randomized trials of non-steroidal anti-inflammatory drugs for the prevention of heterotopic bone formation after major hip surgery. AB - We performed a systematic survey of randomized trials to determine the effects of perioperative NSAIDs on the occurrence of heterotopic bone formation, gastrointestinal side-effects and long-term clinical outcomes after major hip surgery. 13 trials involving 4,129 individuals were identified. Overall, in 12 small trials of medium-to-high-dose regimens, there was a 57% reduction (95% confidence interval 51%-63%) in the risk of heterotopic bone formation. The results of one large trial of low-dose aspirin differed markedly (2% reduction (95% CI 12% reduction to 15% increase)). The NSAID regimens studied had no definite effect on gastrointestinal complications, and data about the effects of NSAIDs on pain and function were too few, and too incompletely reported, to draw conclusions about their effects on these outcomes. Routine prophylaxis against heterotopic bone formation with NSAIDs may be a useful adjuvant therapy for patients undergoing major hip surgery, but the overall balance of risks and benefits requires assessment in a large-scale randomized trial. PMID- 10852317 TI - No effect of low-dose aspirin for the prevention of heterotopic bone formation after total hip replacement: a randomized trial of 2,649 patients. AB - 2,649 patients scheduled for elective total hip replacement were recruited to the Heterotopic Bone Formation Sub-study of the Pulmonary Embolism Prevention Trial. Heterotopic bone formation was determined by radiographic examination and associated late postoperative outcomes were assessed by telephone interview. Heterotopic bone formation was observed in 627 (31%) of 2,048 radiographic examinations. There was no detectable effect of low-dose aspirin on the risks of heterotopic bone formation (RR 0.98; 95% CI 0.85-1.12), late postoperative pain (RR 1.10; 95%CI 0.91-1.35) or late postoperative impaired function (RR 1.03; 95% CI 0.94-1.12). The balance of benefits and risks of low-dose aspirin is determined by its effects on vascular events and bleeding, since it has no major effects on heterotopic bone formation or associated clinical outcomes. PMID- 10852318 TI - Impaction bone-grafting of severely defective femora in revision total hip surgery: 21 hips followed for 41-85 months. AB - We performed a prospective study to evaluate the application of impacted allograft bone particles at revision surgery of severely defective femora. According to the Endo-Klinik classification, 21 hips had grade III or IV femoral defects, of which 4 had a preoperative fracture. No femoral re-revisions had been necessary after a mean follow-up of 60 (41-85) months. The mean Harris Hip Score improved by 39 points to 78 points. 2 fractures occurred postoperatively, 1 of which needed reoperation with osteosynthesis. 1 patient needed a closed reduction after dislocation. 4 stems showed significant subsidence (> 10 mm) in the follow up radiographs. In total hip revision surgery, even severely damaged femora can be successfully treated by impaction allografting. PMID- 10852319 TI - Impacted morsellized bone grafting and cemented primary total hip arthroplasty for acetabular protrusion in patients with rheumatoid arthritis: an 8- to 18-year follow-up study of 36 hips. AB - Between 1979 and 1989, we performed 36 primary total hip replacements in 31 rheumatoid arthritis patients with protrusio acetabuli. The deficient acetabulum was reconstructed with autologous morsellized bone grafts from the femoral head. 3 patients were lost to follow-up. 12 patients (13 hips) died within 8 years postoperatively, none had a revision. 16 patients (20 hips) were reviewed at an average follow-up of 12 (8-18) years. In 2 hips, a revision was performed for aseptic loosening of the acetabular component, 65 and 8 years after primary surgery, which means a 90% (95% CI: 77%-100%) survival rate at 12 years (Kaplan Meier analysis). This technique is a good option in cases with protrusio acetabuli due to rheumatoid arthritis. PMID- 10852320 TI - The incidence of acute cardiorespiratory and vascular dysfunction following intramedullary nail fixation of femoral metastasis. AB - Intramedullary nail fixation is a common treatment for metastatic tumors of the femur with overt or impending femoral fracture. This procedure sometimes causes severe cardiorespiratory and vascular dysfunction. The clinical relevance of this is not dear. We reviewed 45 operations in 43 patients, where intramedullary nail fixation was used to treat metastatic femoral fractures and impending fractures. We studied the incidence of intraoperative oxygen desaturation and hypotension associated with intramedullary manipulation as markers of cardiorespiratory and vascular dysfunction. Acute oxygen desaturation and hypotension occurred in 11 of our 45 patients. Of these, 3 died, 2 required intensive care postoperatively and 6 made uneventful recoveries. We hope to highlight a serious complication in this patient group. PMID- 10852321 TI - Proprioception after total knee arthroplasty: a comparison with clinical outcome. AB - We determined proprioception in replaced and unreplaced arthrotic knees by measuring threshold levels for the perception of passive knee motion. In addition, results of these proprioception measurements were compared with the clinical outcomes in patients with a total knee arthroplasty. Threshold detection levels were significantly higher in the replaced than in the unreplaced knees. Moreover, detection-failure rates were significantly higher in the replaced knees as well. In contrast to this diminished movement sense in the replaced knees, clinical examination of these knees showed good or excellent outcome in all cases. A correlation between the clinical outcome and the ability to perceive passive motion in either patient group could not be found. We hypothesize that our findings may be due to the operative removal of intraarticular receptor-rich tissue that is affected by arthrosis. This would not only contribute to marked clinical improvements but also to a significant decrease in proprioception. PMID- 10852322 TI - Displaced tibial shaft fractures: a prospective randomized study of closed intramedullary nailing versus cast treatment in 53 patients. AB - Of 53 patients with unilateral, displaced and closed or grade 1 open tibial shaft fractures, 27 patients (group I) were randomized to treatment with an intramedullary nail and 26 patients (group II) to treatment with a plaster cast. 12 fractures in the latter group were considered stable enough for treatment with only a cast (group IIa), while 14 fractures in group II showed redisplacement during reduction under anesthesia or at 1 week follow-up. Therefore, these fractures were stabilized with cerclage or screws (group IIb), which was a prerequisite for continuing cast treatment. The mean time-to-union was 19 weeks for group I, and 25 weeks for group II. 6 patients in group I and 16 in group II had delayed union. The Nottingham Health Profile index scores on physical mobility, social isolation, work ability, and sexual life were significantly better in group I than in group II at 3 months after injury. Delayed union, malunion, and restricted range of motion at the ankle joint were common complications when these fractures were treated with a cast. We recommend intramedullary nailing for these fractures. PMID- 10852323 TI - Blood supply of the peroneal tendons: injection and immunohistochemical studies of cadaver tendons. AB - We studied the vascular pattern of human peroneal tendons with injection techniques and immunohistochemically by using antibodies against laminin The main blood supply arises from the peroneal artery. The distal part of the peroneus longus tendon is supplied by branches of the medial tarsal artery. Blood vessels enter the peritenon of both tendons via a mesotenon from the posterior aspect. From the peritenon, the blood vessels penetrate the peroneus brevis and peroneus longus tendons and anastomose with a longitudinally-oriented intratendinous network. The amount of vessels in the tendon substance is consistently less than in the surrounding peritenon. The distribution of blood vessels in the peroneal tendons is not homogeneous. In the region where the peroneus brevis tendon passes through the fibular groove, the longitudinally-oriented intratendinous vascular network is interrupted and the tendon is almost avascular. In this region, the tendon is squeezed between the peroneus longus tendon and the bony slide bearing of the lateral malleolus. The peroneus longus tendon has two avascular zones. In the region where the peroneus longus tendon curves around the lateral malleolus and the peroneal trochlea of the calcaneus, the anterior part of the tendon which is directed towards the pulley is avascular. A second avascular zone is located more distally in the region where the tendon changes direction and wraps around the cuboid. PMID- 10852324 TI - Accidental falls and related fractures in 65-74 year olds: a retrospective study of 332 patients. AB - We investigated, by studying medical records, background factors and consequences of accidental falls of patients 65-74 years who attended the Department of Orthopedics' emergency clinic in Lund. We also assessed possible prevention measures. Fractures occurred in three quarters of the registered falls. Women were more prone to sustain fractures than men. Forearm fractures were commonest among women while hip fractures were commonest among men. One third of the patients were admitted to an orthopedic ward because of the fall. The patients who were less healthy had sustained fractures oftener and also needed more hospital care. Information regarding risk factors for falls and fractures were often missing in the patients' medical records. Impaired walking and balance, and medication increased the risk of falls. Such patients constitute a high risk group for future falls and fractures. A newly developed instrument is suggested as a routine in the emergency department to increase the awareness of risk factors for falls in the elderly. Satisfactory documentation is a prerequisite for further treatment and referrals to prevent falls and fractures. PMID- 10852325 TI - Outcome after cup hemiarthroplasty in the rheumatoid shoulder: a retrospective evaluation of 39 patients followed for 2-6 years. AB - 33 rheumatoid patients, treated with hemispherical cup resurfacing hemiarthroplasty of the shoulder without medullary fixation (6 bilaterally), were reviewed after mean 4.4 (2-6) years. The median Constant score was 30 (15-79), mean proximal migration of the humerus 55 (SD 5.2) mm and mean glenoid erosion 2.6 (SD 1.7) mm. Proximal migration and glenoid erosion did not correlate with shoulder function or pain. Radiographic signs of loosening (changes in cup inclination combined with changes in cup distance above the greater tuberosity) occurred in one quarter of the shoulders. At follow-up, 26 patients were satisfied with the procedure, despite poor shoulder function and radiographic deterioration. PMID- 10852326 TI - Debridement arthroplasty for osteoarthrosis of the elbow: 50 patients followed mean 5 years. AB - The characteristics and surgical outcome of debridement arthroplasty were investigated in athletes and manual laborers with osteoarthrosis of the elbow. There were 26 elbows in athletes, and 24 elbows in laborers. The mean age was 32 years in athletes and 50 years in laborers. The osteoarthrosis was mainly mild in athletes, but moderate or severe in laborers. Debridement arthroplasty, consisting of resection, osteophytes and removal of loose bodies, was performed in all cases. The medial approach was most frequently employed. Surgery relieved pain and improved range of motion at an average follow-up of 59.5 months. Evaluation of the long-term outcomes at more than 5 years showed recurrence of mild osteoarthrosis with minimal symptoms. Debridement arthroplasty is an effective treatment in athletes and manual laborers with osteoarthrosis of the elbow. PMID- 10852327 TI - Function after early radial head resection for fracture: a retrospective evaluation of 15 patients followed for 3-18 years. AB - We reviewed the results of early radial head resection in 15 patients after fracture of the radial head or neck: 4 Mason's type II fractures and 11 Mason's type III fractures. The average age at operation was 41 years and the patients were reexamined after mean 10 (3-18) years. Only 5 patients had no pain and all patients had reduced elbow power, in several of them a substantial loss. PMID- 10852328 TI - Clodronate increases mineralization of callus after Colles' fracture: a randomized, double-blind, placebo-controlled, prospective trial in 32 patients. AB - In a randomized study of 32 postmenopausal women with a Colles' fracture, we studied whether 8 weeks of treatment with clodronate, a bisphosphonate, could prevent posttraumatic osteopenia. The patients were treated with a plaster splint for 4 weeks. The bone mineral density (BMD) of the forearm bones was measured at 2 levels with dual-energy x-ray absorptiometry (DEXA) 2, 6 and 12 months after the fracture. At 2 months, in the clodronate group, there was a median 53% higher BMD in the fracture region of the radius than in the uninjured radius. In the placebo group, we found a 33% higher BMD in the fractured radius at that level than in the uninjured radius. This increase in BMD of the fractured radius, caused by clodronate, was statistically significant. At 12 months, the BMD of the fracture side had been reduced by 17% and 12%, respectively, at that time it was still significantly increased in the clodronate group alone. In the ulna at the same level, we found no significant changes in BMD in either group on either side at any time. At 2 months, at the level between the distal and middle thirds, in the fractured radius, the median BMD was 7% lower in the clodronate group and 6% lower in the placebo group than in the uninjured radius. Although the reduction in BMD at that level was significant, there was no difference between the two treatment groups. At this level, the ulna on the fractured side showed a similar pattern, with a 5% lower BMD in the clodronate group and a 4% lower BMD in the placebo group. This osteopenia showed a small but significant progression on the fractured side after 6 and 12 months. PMID- 10852329 TI - "Under-the-plate" reduction band technique. PMID- 10852330 TI - Modified lateral approach for knee arthroplasty in a fixed valgus knee--the medial quadriceps snip. PMID- 10852331 TI - Fracture of the acetabulum complicated by a tear of the femoral vein--a case report after 5 years. PMID- 10852332 TI - Urinary frequency caused by a misplaced acetabular reinforcement ring--a case report. PMID- 10852333 TI - Avulsion of the ischial tuberosity simulating neoplasm--a report of 2 cases. PMID- 10852334 TI - Low-molecular weight heparin as prophylaxis against thromboembolism after total hip replacement--is it worth the price? PMID- 10852335 TI - Bioethics as methodological case resolution: specification, specified principlism and casuistry. AB - Bioethical decision-making depends on presuppositions about the function and goal of bioethics. The authors in this issue of The Journal of Medicine and Philosophy share the assumption that bioethics is about resolving cases, not about moral theory, and that the best method of bioethical decision-making is that which produces useful answers. Because we have no universally agreed upon background moral theory which can serve as the basis for bioethical decision-making, they try to move bioethics away from theory. For them, a good method of bioethical decision-making is one which resolves cases in ways that are justifiable to the parties involved, not necessarily in ways that bring us "close" to the right and the true. The authors consider how the move away from theory and toward actual cases is best accomplished. In particular, the debate in this issue is about specification, specified principlism, and casuistry. PMID- 10852336 TI - Specifying, balancing, and interpreting bioethical principles. AB - The notion that it is useful to specify norms progressively in order to resolve doubts about what to do, which I developed initially in a 1990 article, has been only partly assimilated by the bioethics literature. The thought is not just that it is helpful to work with relatively specific norms. It is more than that: specification can replace deductive subsumption and balancing. Here I argue against two versions of reliance on balancing that are prominent in recent bioethical discussions. Without meaning to address the substance or the overall merits of either view I criticize, I attack Gert, Culver and Clouser's implicit reliance on some overall dimension of balancing as a basis of resolving conflicts among norms and Beauchamp and Childress's residual acceptance of 'justified balancing'. The former authors' description of resolving conflicts depends upon a type of value commensurability that (as they otherwise seem to admit) does not obtain, while the latter authors' role for justified balancing would be better served by continued specification. PMID- 10852337 TI - Specified principlism: what is it, and does it really resolve cases better than casuistry? AB - Principlism has been advocated as an approach to resolving concrete cases and issues in bioethics, but critics have pointed out that a main problem for principlism is its lack of a method for assigning priorities to conflicting ethical principles. A version of principlism referred to as 'specified principlism' has been put forward in an attempt to overcome this problem. However, none of the advocates of specified principlism have attempted to demonstrate that the method actually works in resolving detailed clinical cases. This paper shows that when one tries to use it, specified principlism fails to provide practical assistance in deciding how to resolve concrete cases. Proponents of specified principlism have attempted to defend it by arguing that it is superior to casuistry, but it can be shown that their arguments are faulty. Because of these reasons, specified principlism should not be considered a leading contender in the search for methods of making justifiable decisions in clinical cases. PMID- 10852338 TI - Strong on specification. PMID- 10852339 TI - Enhanced cellular uptake and transport of polyclonal immunoglobulin G and fab after their cationization. AB - Antibodies are poorly transported across cell membranes and biological barriers in vivo. Cationization of antibody molecules by the derivatization of surface carboxyl groups generating primary amino groups could represent a strategy for intracellular antibody delivery. Before cationization of polyclonal colchicine specific IgG and Fab, using hexamethylenediamine the isoelectric point (pl) of native IgG and Fab (nIgG and nFab) was in the range of 5.9 9.0 and 8.7-9.3, respectively. The pI of cationized IgG and Fab (cIgG and cFab) were both higher at 8.7, 10.3 and 9.5 -11, respectively. The affinity and specificity of both IgG and Fab were not modified by cationization. When HL 60 cells were incubated with the native or cationized 125I-BSA. -IgG and -Fab, the maximal cellular uptake of clgG and cFab was 3.2 and 2.4 times higher than that of nIgG and nFab at an extracellular concentration of 500 ng/ml. Results also indicated that the uptake was dose- and temperature-dependent suggesting absorptive-mediated endocytosis of cationized antibodies by HL 60 cells. Confocal microscopy analysis indicated that the cationized antibodies were present in the plasma membranes and cytoplasm of HL 60 cells. Finally, a study with bovine arterial endothelial monolayer cells showed that the transport of cIgG and cFab through the monolayer cells was 3.3- and 4.3-fold higher for 125I-cIgG and 125I-cFab than those of the corresponding native forms. PMID- 10852340 TI - Similar efficiency of DNA-liposome complexes and retrovirus-producing cells for HSV-tk suicide gene therapy of peritoneal carcinomatosis. AB - Several experimental approaches have been tested for suicide gene delivery into tumor cells, including viral and non-viral vectors. In this study, we compared the efficiency of Herpes Simplex Virus type 1 thymidine kinase gene (HSV-tk) delivery by retrovirus-producing cells and DNA/liposome complexes for the treatment of peritoneal carcinomatosis induced in syngeneic rats by DHD/K12 colorectal adenocarcinoma cells. After in vitro determination of the best transduction conditions, rats were treated with multiple intraperitoneal injections of plasmid DNA containing one or two copies of CMV-driven HSV-tk gene (pCMV-TK and p(CMV-TK)2, respectively) associated with LipofectAMINE, each injection being followed by a Ganciclovir (GCV) course. Animals treated by DNA/liposome complexes and GCV or with retrovirus-producing cells and GCV showed a similar increase of survival as compared to the control group. After DNA/ liposome injections, expression of the tk transgene was detected in tumor nodes (epiploon) and also in liver, lung, spleen, bowels and brain. The expression was not homogeneous throughout the different organs and most likely reflected the transfection of only a limited number of cells. PMID- 10852341 TI - Block and graft copolymers and NanoGel copolymer networks for DNA delivery into cell. AB - Self-assembling complexes from nucleic acids and synthetic polymers are evaluated for plasmid and oligonucleotide (oligo) delivery. Polycations having linear, branched, dendritic. block- or graft copolymer architectures are used in these studies. All these molecules bind to nucleic acids due to formation of cooperative systems of salt bonds between the cationic groups of the polycation and phosphate groups of the DNA. To improve solubility of the DNA/polycation complexes, cationic block and graft copolymers containing segments from polycations and non-ionic soluble polymers, for example, poly(ethylene oxide) (PEO) were developed. Binding of these copolymers with short DNA chains, such as oligos, results in formation of species containing hydrophobic sites from neutralized DNA polycation complex and hydrophilic sites from PEO. These species spontaneously associate into polyion complex micelles with a hydrophobic core from neutralized polyions and a hydrophilic shell from PEO. Such complexes are very small (10-40 nm) and stable in solution despite complete neutralization of charge. They reveal significant activity with oligos in vitro and in vivo. Binding of cationic copolymers to plasmid DNA forms larger (70-200 nm) complexes. which are practically inactive in cell transfection studies. It is likely that PEO prevents binding of these complexes with the cell membranes ("stealth effect"). However attaching specific ligands to the PEO-corona can produce complexes, which are both stable in solution and bind to target cells. The most efficient complexes were obtained when PEO in the cationic copolymer was replaced with membrane-active PEO-b-poly(propylene oxide)-b-PEO molecules (Pluronic 123). Such complexes exhibited elevated levels of transgene expression in liver following systemic administration in mice. To increase stability of the complexes, NanoGel carriers were developed that represent small hydrogel particles synthesized by cross-linking of PEI with double end activated PEO using an emulsification/solvent evaporation technique. Oligos are immobilized by mixing with NanoGel suspension, which results in the formation of small particles (80 nm). Oligos incorporated in NanoGel are able to reach targets within the cell and suppress gene expression in a sequence-specific fashion. Further. loaded NanoGel particles cross-polarized monolayers of intestinal cells (Caco-2) suggesting potential usefulness of these systems for oral administration of oligos. In conclusion the approaches using polycations for gene delivery for the design of gene transfer complexes that exhibit a very broad range of physicochemical and biological properties, which is essential for design of a new generation of more effective non-viral gene delivery systems. PMID- 10852342 TI - Schizophrenia and drug delivery systems. AB - Schizophrenia is a severe non-curable illness of the brain with serious consequences if not properly treated and kept under control. Antipsychotic drugs have revolutionised the therapy and management of schizophrenia. However, patient compliance rates are notoriously poor due to the nature of the disease and troublesome side-effects, and are major causes of symptom recurrence. Although some new antipsychotic agents have been marketed to offer broader efficacy with much reduced side-effect profiles, the drug delivery systems for antipsychotics are still in the stage of conventional dosage forms, such as tablets, capsules and solutions, and need to be dosed at the frequency of 2-4 times daily. Doubtless. novel drug delivery systems, such as sustained and controlled release systems, will be useful for antipsychotics. They should reduce the frequency of dosing. enhance drug bioavailability and improve patient compliance. In this article, the specificity and characterisation of schizophrenia and pathophysiology. drug therapy. and the development and future prospects of neuroleptic drug delivery systems are reviewed. PMID- 10852343 TI - Folic acid targeting of protein conjugates into ascites tumour cells from ovarian cancer patients. AB - Despite a wealth of in vitro data describing the use of folic acid for drug and DNA delivery into ovarian cancer cell lines, there have been no reports describing the targeting of such compounds to freshly isolated tumour cells. We have carried out a study to determine the usefulness of folic acid as a targeting ligand for ovarian cancer by measuring the uptake of folic acid-BSA-FITC in tumour cells isolated from the ascitic fluid of ovarian cancer patients. In 7 out of 7 patients we have found folic acid mediated uptake of the fluorescently labelled albumin, with the accumulation (average cell fluorescence) and differential uptake (ratio between receptor mediated and fluid phase uptake) varying between patients. Accumulation of folic acid-albumin FITC occurs in ascites tumour cells expressing the epithelial cell marker EMA, with a significant proportion of EMA negative cells also accumulating the conjugate. There is no correlation between cell cycle and uptake of folic acid-BSA-FITC. These results suggest that folic acid-targeting of therapeutics is a promising approach for the treatment of ovarian cancer. PMID- 10852344 TI - Optimization of cationic lipid/DNA complexes for systemic gene transfer to tumor lesions. AB - Intravenous (i.v.) administration of cationic lipid N-[( 1-(2-3 dioleyloxy)propyl)]-N-N-N-trimethylammonium chloride (DOTMA)-based transfection complexes in mice with subcutaneous squamous cell tumors yielded plasmid delivery and expression in tumor lesions. The efficiency of gene transfer in tumors was significantly lower than in the lung. This was consistent with low plasmid levels associated with the tumor, suggesting that plasmid delivery to the tumor site was a limiting factor. Lowering the lipid/DNA charge ratio from 5:1 to 0.8:1 (+/-) did not change DNA levels in tumor but significantly reduced DNA levels in lung. However, expression levels were significantly reduced in both tissues at lower lipid/DNA charge ratios. Complexes prepared from small unilamellar liposomes gave significantly lower expression levels in the lungs but similar expression levels in tumors when compared to complexes prepared from larger unilamellar liposomes. The small liposome complexes were better tolerated than large liposome complexes. Varying the cationic lipid to colipid (cholesterol or DOPE) molar ratio from 4: 1 to 1: 1 significantly reduced expression levels in both tumor and lung. Cationic lipid substitution, using a cholesterol cationic lipid, diethyldiamino-carbamyl cholesterol instead of DOTMA, produced reduced expression in all other tissues except tumor. Incorporation of PEG into preformed transfection complexes reduced DNA delivery to lung, increased circulation half-life, and enhanced DNA delivery to tumor. In a lung metastatic mouse tumor model, where the accessibility of the i.v. administered transfection complexes to tumor lesions should be less challenging, DOTMA: CHOL complexes (4: 1 lipid to colipid molar ratio, 3: 1 +/- lipid to plasmid charge ratio) were preferentially localized in tumor lesions. These data demonstrate that systemic gene transfer to distal tumor sites by lipid/ DNA complexes may be limited by low plasmid delivery. Modifying the chemical surface properties of transfection complexes enhanced both DNA delivery and expression in tumor and is one approach that may overcome limitations. PMID- 10852345 TI - Characterization of a human cell line derived from liposarcoma tissue: is the beta subunit of prolyl 4-hydroxylase specific for a fibroblastic phenotype in culture cells? PMID- 10852346 TI - Umbilical cord endothelial cells expressing large T antigen: comparison with primary cultures and effect of cell age. AB - A number of human endothelial cell lines from umbilical cord cells (HUVECs) have been generated by transfection with SV40 large T and small t antigen sequences. Comparison of these lines with primary cultures of HUVECs has been carried out by monitoring the expression of a number of endothelial cell markers with specific regard to cell age. The secreted levels of the protein plasminogen activator inhibitor (PAI) was found to be significantly reduced in SV40-transfected cells when compared to untransfected controls. Tissue plasminogen activator (tPA) and urokinase (uPA) levels were unchanged. As cells entered crisis, there was a rapid and significant increase in the levels of tPA, uPA, and PAl and this was observed for all clones screened. The endothelial cell marker von Willebrand Factor (vWF) was found intracellularly and was also secreted into the medium. The levels were not altered between transfected and untransfected cells. Angiotensin converting enzyme (ACE) activity was maintained in cell lines at levels found in nonimmortalized HUVECs. Both isoforms (alpha and beta) of IL-1 (interleukin-1) increased as cells approached crisis, and the presence of these cytokines may be responsible for the increased levels of tPA, PAI, and uPA. With one exception, the ability of the transfected cells to produce prostacyclin (PGI2) was lost by all clones. PMID- 10852347 TI - Characterization of JOK-1, a human gastric epithelial cell line. AB - Human gastric epithelial cells were isolated from samples of human gastric lining and immortalized with simian virus 40 (SV40) to generate the stable human gastric epithelial cell line "JOK-l." These cells express conventional epithelial markers (vimentin, cytokeratin-18, occludin, N- and E-cadherins, beta-catenin, ZO-1, ZO 2, mucin, epithelial specific antigen) as well as SV40 large T-antigen. These cells rapidly externalized E-cadherin in response to acidic medium, and exhibited epithelial-like barrier properties that are also regulated by media pH. In contrast, the kidney epithelial cell line "MDCK" also expresses several epithelial markers (vimentin, cytokeratin-18, occludin, N- and E-cadherin, beta catenin, ZO-1, ZO-2, epithelial specific antigen), but does not express mucin, or large T-antigen. However, MDCK rapidly internalize their E-cadherin from the cell surface and increase the solute flux in an acidic medium. These data suggest that the JOK-1 cell line is a potentially useful cell line for developing models of gastric epithelial function, development, and disease. PMID- 10852348 TI - Efficient isolation and long-term viability of bovine small preantral follicles in vitro. AB - A comparison of isolation techniques for small preantral follicles (30-70 microm) from bovine ovaries using a mechanical method with a grating device or collagenase treatment was performed. The mean number (157.0) of intact follicles per ovary isolated by the mechanical method was significantly greater (P < 0.05) than that (26.0) of follicles isolated by the enzymatic method. Isolated morphologically normal follicles (MNF) were cultured for up to 30 d either in control cultures (non-coculture) or in coculture with bovine ovary mesenchymal cells (BOM), fetal bovine skin fibroblasts (FBF), and/or bovine granulosa cells (BGC). In control cultures, most of the follicles degenerated and only a few MNF (1.2%) were present after 30 d in culture. In contrast, the cocultures with BOM, FBF, and BGC resulted in 50.7, 46.6, and 21.4% viable MNF, respectively. Trypan blue and Hoechst 33258 staining were used for a quick and sensitive assessment of oocyte and granulosa cell viability during follicle isolation and culture in vitro. After 30 d, percentages of viable follicles in coculture with BOM (18.6%) and FBF (17.1%) were significantly greater than those of follicles in the control cultures (0%) or in coculture with BGC (10.0%). There was a gradual increase in the average diameter of the MNF during culture. The mean diameter of the follicles increased by 15.4 and 30.0% in coculture with BOM and FBF, respectively, by day 30. In conclusion, small bovine preantral follicles were efficiently isolated using a mechanical method that utilizes a grating device, and could be maintained for up to 30 d in the presence of mesenchymal cell cocultures such as BOM and FBF. This in vitro culture system that supports long term survival of bovine preantral follicles should be beneficial for studying follicle growth and development. PMID- 10852349 TI - Isolation of a spontaneously fusing BC3H1 muscle cell line: fusion alters the response to serum stimulation. AB - Differentiation of skeletal muscle cells involves two distinct events: exit from the cell cycle and expression of muscle-specific contractile genes and formation of multinucleated myocytes. Although many studies have shown that growth factors regulate the initial step of differentiation, little is known about regulation of fusion. BC3H1 cells are a skeletal muscle cell line characterized by a nonfusing phenotype and an ability to dedifferentiate. When subjected to serum or growth factors, differentiated BC3H1 cells lose muscle-specific gene expression and re enter the cell cycle. In this study, we describe a spontaneously fusing clone of BC3H1 cells. We demonstrate that this fusion capability is not due to altered muscle regulatory factor or adhesion molecule expression. Furthermore, we show that fusion inhibits dedifferentiation. Multinucleated BC3H1 cells do not lose myosin expression, nor do they re-enter the cell cycle. Fused BC3HI cells react to serum stimulation with a hypertrophic response. Our results suggest that the state of differentiation, mono- or multi-nucleated, is essential to how myocytes react to growth stimulation and may provide a mechanism for how differentiation, fusion, and hypertrophy are regulated in vivo. PMID- 10852350 TI - Retinoic acid increases amount of phosphorylated RAF; ectopic expression of cFMS reveals that retinoic acid-induced differentiation is more strongly dependent on ERK2 signaling than induced GO arrest is. AB - Retinoic acid is known to cause the myeloid differentiation and G1/0 cell cycle arrest of HL-60 cells in a process that requires mitogen-activated protein/extracellular signal regulated kinase (MEK)-dependent extracellular signal regulated kinase (ERK)2 activation. It has also been shown that ectopic expression of cFMS, a platelet-derived growth factor (PDGF)-family transmembrane tyrosine kinase receptor, enhances retinoic acid-induced differentiation and G1/0 arrest. The mechanism of how the retinoic acid and cFMS signaling pathways intersect is not known. The present data show that the ectopic expression of cFMS results in the differential loss of sensitivity of retinoic acid-induced differentiation or G1/0 arrest to inhibition of ERK2 activation. PD98059 was used to inhibit MEK and consequently ERK2. In wildtype HL-60 cells, PD98059 blocked retinoic acid-induced differentiation; but in cFMS stable transfectants, PD98059 only attenuated the induced differentiation, with the resulting response resembling that of retinoic acid-treated wild-type HL-60. In wild-type HL-60, PD98059 greatly attenuated the retinoic acid-induced G1/0 arrest allied with retinoblastoma (RB) hypophosphorylation; but in cFMS stable transfectants, PD98059 had no inhibitory effect on RB hypophosphorylation and G1/0 arrest. This differential sensitivity to PD98059 and uncoupling of retinoic acid-induced differentiation and G1/0 arrest in cFMS transfectants is associated with changes in mitogen-activated protein kinase signaling molecules. The cFMS transfectants had more activated ERK2 than did the wild-type cells, which surprisingly was not attributable to enhanced mitogen-activated protein-kinase-kinase-kinase (RAF) phosphorylation. Retinoic acid increased the amount of activated ERK2 and phosphorylated RAF in both cell lines. But PD98059 eliminated detectable ERK2 activation, as well as inhibited RAF phosphorylation, in untreated and retinoic acid-treated wild-type HL-60 and cFMS transfectants, consistent with MEK or ERK feedback-regulation of RAF, in all four cases. Since PD98059 blocks the cFMS conferred enhancement of the retinoic acid-induced differentiation, but not growth arrest, the data indicate that cFMS-enhanced differentiation acts primarily through MEK and ERK2, but cFMS-enhanced G1/0 arrest allied with RB hypophosphorylation depends on another cFMS signal route, which by itself can effect G1/0 arrest without activated ERK2. Ectopic expression of cFMS and differential sensitivity to ERK2 inhibition thus reveal that retinoic acid induced HL-60 cell differentiation and G1/0 arrest are differentially dependent on ERK2 and can be uncoupled. A significant unanticipated finding was that retinoic acid caused a MEK-dependent increase in the amount of phosphorylated RAF. This increase may help sustain prolonged ERK2 activation. PMID- 10852351 TI - Inhibitory effects of human serum on human fetal skin fibroblast migration: migration-inhibitory activity and substances in serum, and its age-related changes. AB - In order to clarify the environmental factors modulating cell migration, we investigated the effects of human serum on cell migration, and found that serum from adult donors strongly (by 48%) suppressed the migration of human fetal skin fibroblasts into a denuded area in a cell monolayer. Human serum from old donors inhibited cell migration more strongly than that from adult donors. Next, we investigated the properties of migration-inhibitory activity of human serum and serum proteins in order to identify migration-inhibitory substances. Human serum from adult donors strongly suppressed the migration of human fetal skin fibroblasts, although it stimulated cell proliferation more strongly than fetal bovine serum (FBS), indicating that the inhibitory effects of human serum on cell migration was not due to its toxic effects. The inhibition of cell migration by human serum was concentration dependent. It was demonstrated that the inhibition did not depend on the inhibitory effects of human serum on collagen synthesis. The migration-inhibitory activity was seen in fractions over 100 kDa, as determined by an ultrafiltration membrane, and no inhibitory activity was observed in fractions under 100 kDa. On the other hand, it was not detected either in fractions over 100 kDa or under 100 kDa in FBS. Among the over 100 kDa human serum proteins examined, gamma-globulin, alpha2-macroglobulin, and low density lipoprotein (LDL) suppressed fibroblast migration in a concentration dependent manner. However, among the three, cell migration-inhibiting activity of gamma-globulin almost disappeared when cell migration was conducted in 10% FBS supplemented medium. These results indicated that alplha2-macroglobulin and LDL were candidate substances for cell migration-inhibiting activity in human serum. PMID- 10852352 TI - Cell viability improves following inhibition of cryopreservation-induced apoptosis. AB - A new concept in cryopreservation solution design was developed that focuses on the use of an intracellular-type, hypothermic maintenance medium coupled with additives that inhibit cryopreservation-induced apoptosis. HypoThermosol' (HTS), a hypothermic (4 degrees C) maintenance medium utilized in the long-term storage of cell, tissue, and organ systems, was tested for cryoprotective capability on a renal cell line (Madin-Darby Canine Kidney cells). HTS and HTS derivatives were tested against conventional cell culture medium (Dulbecco's Minimal Essential medium, DME) as the cryoprotectant carrier solution because (1) cells are exposed to an extended state of hypothermia during the freeze-thaw process, and (2) HTS is designed to protect cells exposed to a hypothermic state. Cells separately cryopreserved in either HTS or DME + 5% dimethyl sulfoxide (DMSO) yielded equivalent 24-h postthaw survival (approximately 30%) and 5-d recovery (approximately 90%). Cells cryopreserved in CryoStor CS 5, a HTS derivative containing 5% DMSO, yielded approximately 75% 24-h postthaw survival and recovery to 100% within 3 d. DNA gel electrophoresis was performed to determine the mechanisms of cell death contributing to cryopreservation failure. Cells preserved in DME (DMSO-free) died primarily through necrosis, whereas cells preserved in either DME + 5% DMSO, HTS, or CryoStor CS 5 died through a combination of apoptosis and necrosis. This observation led to the inclusion of an apoptotic inhibitor designed to improve cryopreservation outcome. MDCK cells cryopreserved in CryoStor CS 5 supplemented with an apoptotic inhibitor (Caspase I Inhibitor V), hereafter termed CryoStor CS 5N, resulted in a 24-h postthaw survival and recovery rate exceeding that of any other cryoprotective solution tested (85%). We conclude that: (1) the use of HTS (a dextran-based, intracellular-type solution) without DMSO can yield postthaw viability equivalent to that of standard DMSO-based cryopreservation methods, (2) postthaw viability can be significantly increased through the use of an intracellular-type solution in conjunction with DMSO, (3) the use of HTS allows for cryopreservation to be accomplished with reduced levels of cryoprotectants, and (4) the regulation of apoptosis is essential for the improvement of cryopreservation outcome. PMID- 10852353 TI - In vitro host range studies with a new baculovirus isolate from the diamondback moth Plutella xylostella (L.) (Plutellidae: Lepidoptera). AB - The in vitro host range of a newly isolated baculovirus from the diamondback moth Plutella xylostella was tested against six lepidopteran cell liness. Two baculoviruses with wide host ranges from the alfalfa looper Autographa californica (A. californica multiple nucleopolvhedrovirus, AcMNPV) and the celery looper Anagrapha falcifera (AfMNPV) were also included in this study for comparative purposes. PxMNPV replicated in all six cell lines and produced occlusion bodies, with HV-AM1 and TN-CL1 cells producing the highest viral titers and greatest number of occlusion bodies. There was no significant replication of AcMNPV and AfMNPV in the HZ-FB33 cell line and thus no production of occlusion bodies. The restriction endonuclease profiles of the three baculoviruses showed similarities but could be readily distinguished from each other. Either HV-AM1 or TN-CL1 would be suitable cell lines for the in vitro production of PxMNPV. PMID- 10852354 TI - Anger management group treatment for cocaine dependence: preliminary outcomes. AB - Cocaine abusers who fail to manage anger appropriately may have greater difficulty achieving and maintaining abstinence. We conducted a pilot study to examine an anger management group treatment in a sample of 59 men and 32 women with a diagnosis of cocaine dependence. Participants attended a 12-week anger management group treatment and background substance abuse treatment. Levels of anger, negative affect, and anger control were measured at baseline, weekly during treatment, and at 3-month posttreatment follow-up. Levels of anger decreased and anger control increased between baseline and the end of treatment. End-of-treatment changes were maintained at follow-up. These findings were not moderated by gender, age, or psychiatric medication use. In the absence of a randomized control group, we cannot make conclusive statements regarding the effectiveness of the anger management group treatment. However, these preliminary findings demonstrate the need for a randomized clinical trial to test the efficacy of the anger management group treatment. PMID- 10852355 TI - The relationship between anxiety levels and outcome of cocaine abuse treatment. AB - Although a number of studies have examined the comorbidity of anxiety disorders and substance use disorders, much less is known about the impact of anxiety symptoms on substance use and on substance abuse treatment outcome. In the current study, we examined how self-reported anxiety levels, as measured by the Spielberger State-Trait Anxiety Inventory, were related to cocaine use variables and patterns following substance abuse treatment. There were 108 patients in substance abuse treatment who met DSM-III-R diagnostic criteria for cocaine abuse or dependence who completed an assessment battery at pretreatment, posttreatment, and 3-month follow-up. State anxiety scores significantly declined from pre- to posttreatment and remained stable into the 3-month follow-up period regardless of relapse status. Trait anxiety was correlated positively with negative consequences due to cocaine use and negatively correlated with days in treatment. State and trait anxiety both were correlated positively with the Alcohol Composite Index of the Addiction Severity Index (ASI). These findings suggest that elevated anxiety scores at pretreatment subside with time, do not require clinical management of associated anxiety symptoms, and may be a temporary by product of experiencing negative consequences due to recent cocaine use. PMID- 10852356 TI - Adherence to antiretroviral therapy among HIV-infected methadone patients: effect of ongoing illicit drug use. AB - Methadone maintenance patients infected with human immunodeficiency virus (HIV) currently receiving antiretroviral therapy had HIV RNA testing and were surveyed regarding their adherence to their treatment regimens. Adherence was measured using self-report on four questions relating to medication use in the last day and last month and whether the patient took "drug holidays." Of the patients (N = 42), 52% were receiving two-drug antiretroviral therapy and 48% were receiving triple therapy that included a protease inhibitor. Persons on triple therapy reported higher rates of adherence on all measures and were more likely to have undetectable HIV RNA levels than persons on dual therapy (60% vs. 50%). Ongoing illicit drug injection was the only factor significantly associated (p < .05) with multiple measure nonadherence; however, it was not associated with undetectable HIV RNA level. Levels of nonadherence were comparable to estimates from other chronic diseases, but this finding has important implications for patients receiving highly active antiretroviral therapy. PMID- 10852357 TI - Predictors of treatment outcomes in men and women admitted to a therapeutic community. AB - This study compared factors that predict treatment outcomes in men and women randomly assigned to two therapeutic communities differing primarily in length of inpatient and outpatient treatment. Based on the prior research literature comparing treatment outcomes of men and women, we formulated the following research question: Do factors that predict drug use at follow-up, postdischarge arrest, and employment at follow-up differ for men and women? Self-reports and objective measures of criminal activity and substance abuse were collected at pre and posttreatment interviews. Separate regression analyses were conducted for men and women for each of the three outcome variables. The results showed that the predictors of outcome for men and women were similar. Clients who completed the 12-month treatment programs had better outcomes regardless of gender. Men and women who completed treatment were characterized at follow-up by substantial reductions in drug use and arrests and by increased employment. Results further suggested that the longer residential treatment program had a particularly beneficial impact on women. Number of prior arrests was also associated with postdischarge outcomes for women. Women with more arrests at admission were more likely to have a postdischarge arrest and less likely to be employed at follow up. This finding provides invaluable information about which women may be at greater risk for relapse and in need of additional services. We conclude that completion of treatment is the key predictor of treatment outcomes for both men and women. PMID- 10852358 TI - Drugs and prisoners: treatment needs on entering prison. AB - An interview study was conducted among a group of incoming prisoners in a county jail in Ohio during the summers of 1997 and 1998 to assess their current drug treatment needs. "Incoming prisoners" refers to individuals who were being transferred from this county jail to the state prison system. Marijuana and cocaine were the most commonly tried illegal drugs among the subjects as well as the drugs of choice during the month prior to imprisonment. The Diagnostic Interview Schedule, combined with questions employed in the Drug Use Forecasting (DUF) project, was used to construct the questionnaire for this study. Based on the criteria of DSM-IV diagnoses, 57.5% of those interviewed had exhibited drug dependency at some point in their lives, and 51% were currently dependent on some substance. Thus, more than half of the incoming prisoners were in need of treatment for use of at least one substance. Cocaine dependence was the greatest problem facing this group of inmates, with an especially notable problem among the older females. Younger males were more likely to have current marijuana dependence. The study found that individuals currently dependent on cocaine or opiates perceived that they had a need for drug treatment, while those currently dependent on marijuana did not share this perception. PMID- 10852359 TI - Factors predicting entry of injecting drug users into substance abuse treatment. AB - A prospective study of 823 injecting drug users (IDUs) was made to identify baseline variables differentiating those who entered treatment during the study from those who did not enter. Variables independently associated with entering treatment in a multiple logistic regression model included (a) expressed desire for treatment, (b) being eligible for methadone maintenance, (c) two or more previous treatment admissions, (d) frequency of injection, (e) heroin use in the past 30 days, (e) being human immunodeficiency virus (HIV) positive, (f) giving money or drugs for sex, and (g) level of injection-related risk for HIV infection. PMID- 10852360 TI - Needle sharing: a longitudinal study of female injection drug users. AB - The objective of this study was to examine the psychosocial risk and protective factors related to needle-sharing behavior among female intravenous drug users (IDUs) positive (N = 96) and negative (N = 128) for human immunodeficiency virus (HIV). Participants in this longitudinal study were interviewed individually at two points in time, with a 6-month interval between interviews. The interviewers used a structured questionnaire, which included psychosocial measures and questions about drug and sexual risk behaviors. Data were analyzed using Pearson correlations and hierarchical regression analyses. The findings supported a developmental model in which the psychosocial domains and HIV status predicted T1 (initial) needle-sharing behavior, which in turn was related to T2 (follow-up) needle-sharing behavior. In addition, the relationship between personality and peer risk factors and T2 needle sharing was buffered by family-related protective factors. While HIV-positive status had a direct effect on T1 needle sharing with strangers, its effect was mediated by all of the psychosocial variables in its relation to T1 needle sharing with familiar people. Comparisons of these results were made with a companion study of male IDUs. The results suggest several intervention and treatment approaches that can be implemented at different points in the developmental pathways leading to risky needle-sharing practices among female IDUs. PMID- 10852361 TI - Homelessness and gender in out-of-treatment drug users. AB - This study examines 5225 out-of-treatment crack users and drug injectors drawn from five different geographic areas to examine selected factors associated with homelessness. Of these crack users, 27% considered themselves undomiciled, and 60% had previously entered some type of drug treatment. Logistic regression found that substance abusers who were married, female, and persons of color were less likely to be without a home when other variables were controlled. Trading sex for money and perceived chance of getting acquired immunodeficiency syndrome (AIDS) were associated positively with homelessness, while participating in methadone detoxification and methadone maintenance programs seemed to offer some protection from homelessness. PMID- 10852362 TI - Changes in Millon Clinical Multiaxial Inventory scores among opiate addicts as a function of retention in methadone maintenance treatment and recent drug use. AB - The Millon Clinical Multiaxial Inventory (MCMI) was administered to 144 men and 86 women within 1 month of admission to methadone maintenance treatment and was readministered 18 months following admission. Based on prior research, we hypothesized there would be significant decreases on scales measuring affective disturbance, anxiety, and social isolation and little change in scales measuring antisocial and narcissistic traits. In addition, it was hypothesized that changes on the MCMI would be related to retention in treatment and illicit drug use during the interim between initial assessment and follow-up. Data were analyzed using a multivariate analysis of variance (MANOVA) for repeated measures. There was an overall decrease in MCMI scores, indicating less psychopathology between initial assessment and follow-up. MCMI scales did not change as a function of retention status, but decreases in MCMI scale scores were greater for subjects who were light drug users in the 6 months prior to the follow-up compared to heavy users. Inspection of individual MCMI scales supported our hypothesis; there were decreases on scales measuring affective disturbance, anxiety, and social isolation, but not on scales measuring antisocial and narcissistic traits. PMID- 10852363 TI - Trends in combinational use of alcohol and illicit drugs among minority adolescents, 1983-1994. AB - Combinational use of substances refers to taking two or more substances together so that they affect the person at the same time. This pattern of substance use presents unique health and safety risks. Trends in combinational use of alcohol and marijuana or alcohol and cocaine were determined using data from three large comparable samples of students in grades 7-12 in New York State, from surveys conducted in 1983, 1990, and 1994. Each of the three samples was demographically diverse, permitting detailed analysis of trends in various adolescent subgroups according to gender, grade level (age), and race/ethnicity. These two forms of adolescent combinational use of alcohol and illicit drugs dropped sharply from 1983 to 1990, but increased or remained stable from 1990 to 1994. Use of alcohol and marijuana together increased sharply from 1990 to 1994, much more for blacks and Hispanics than for whites, while use of alcohol and crack or cocaine together remained stable at a low level in the 1990s. Both forms of combinational use increased in the 1990s more among younger adolescents than among older ones. Analyses controlling for rates of use also suggest that these forms of combinational use are incidental to the use of the individual substances, rather than uniquely sought "highs." Prevention programs should include warnings about the dangers of combinational use, especially for younger adolescents. PMID- 10852364 TI - Card Perseveration Task performance and post-task feeling states: relationship to drug use in adolescents. AB - This study examined whether performance on the Card Perseveration Task (Card Task) and self-report of feeling state after the task are related to self-report of drug use. The evaluation was of 64 adolescents from an adolescent psychiatric outpatient clinic (40 males, aged 15.5 years, SD = 1.6; 24 females aged 16.9 years, SD = 1.5). Drug use histories were obtained using a substance dependence symptom checklist based on DSM-III-R. The Card Task was administered, and after completion, a Post-Task Self-Report (PTSR) was administered. A factor analysis with varimax rotation grouped the 28 items of the PTSR into Distress, Happy, Satisfied, and Wanting to Win subscales. Correlations of drug use with performance on the Card Task and the PTSR subscales were obtained. Cards Played on the Card Task were correlated with alcohol (cc = .31, p < or = .01); marijuana (cc = .35, p < or = .01) and polydrug (cc = .26, p < or = .05) dependence symptoms. Money Won on the Card Task was correlated negatively with nicotine (cc = -.26, p < or = .05) and marijuana (cc = -.27, p < or = .05) dependence symptoms. The PTSR Distress subscale correlated with nicotine (cc = .49, p < or = .001), alcohol (cc = .37, p < or = .01), marijuana (cc = .39, p < or = .01), and polydrug (cc = .49, p < or = .001) dependence symptoms. These findings provide evidence that both the Card Task and feeling states associated with task performance are related to self-reports of drug use. PMID- 10852365 TI - Drug use among welfare recipients in the United States. AB - This study examined the prevalence of drug use in a nationally representative sample of 1989 recipients and 6840 nonrecipients of four welfare programs. Data from the 1995 National Household Survey on Drug Abuse (NHSDA) were analyzed using the conditional form of multiple logistic regression with matching of respondents on neighborhood of residence. Weighted proportions and variances accounting for the complex sample design of the NHSDA survey were estimated using the Taylor series linearization method. The results indicate that drug use is 50% more common in households with welfare recipients than in nonwelfare households. Programs making welfare eligibility contingent on the recipient working toward a drug-free lifestyle are worth examining, although a vigilant eye must be kept on the potential unintended consequences. PMID- 10852366 TI - Recent advances in the molecular basis of inherited photoreceptor degeneration. AB - To date, 118 loci have been associated with photoreceptor degenerative disease. In this review, we will discuss recent advances in the identification of genes that cause progressive photoreceptor cell death when mutated. We will focus on 12 genes isolated within the last two years that have been shown to be photoreceptor specific, or that have provided insight into photoreceptor biology and the mechanisms of photoreceptor cell death. To aid in understanding the biologic basis for these diseases, we also briefly review photoreceptor biology. Finally, we report on recent advances towards the treatment of these disorders. PMID- 10852367 TI - The molecular basis for developmental disorders of the pituitary gland in man. AB - The development of the anterior pituitary gland is dependent upon a cascade of signalling molecules and developmental genes that function as transcription factors. Many of these genes are homeobox genes which contain a DNA-binding region or homeobox. Animal models have given a valuable insight into human pituitary disease. For example, Pit-1 and Prop1 mutants are known to have deficiencies of growth hormone, prolactin and thyroid-stimulating hormone. Human phenotypes arising as a result of mutations in these genes are similar to the mouse mutants. Mutations in the novel homeobox gene Hesx1/HESX1 are associated with the highly variable phenotype of septo-optic dysplasia in mouse and man. The unravelling of this complex developmental cascade is just commencing. PMID- 10852368 TI - Genetic defects as recorded in the pottery of the Moche culture of Peru. PMID- 10852369 TI - Somatic segregation errors predominantly contribute to the gain or loss of a paternal chromosome leading to uniparental disomy for chromosome 15. AB - Paternal uniparental disomy (UPD) for chromosome 15 (UPD15), which is found in approximately 2% of Angelman syndrome (AS) patients, is much less frequent than maternal UPD15, which is found in 25% of Prader-Willi syndrome patients. Such a difference cannot be easily accounted for if 'gamete complementation' is the main mechanism leading to UPD. If we assume that non-disjunction of chromosome 15 in male meiosis is relatively rare, then the gain or loss of the paternal chromosome involved in paternal and maternal UPD15, respectively, may be more likely to result from a post-zygotic rather than a meiotic event. To test this hypothesis, the origin of the extra chromosome 15 was determined in 21 AS patients with paternal UPD15 with a paternal origin of the trisomy. Only 4 of 21 paternal UPD15 cases could be clearly attributed to a meiotic error. Furthermore, significant non-random X-chromosome inactivation (XCI) observed in maternal UPD15 patients (p < 0.001) provides indirect evidence that a post-zygotic error is also typically involved in loss of the paternal chromosome. The mean maternal and paternal ages of 33.4 and 39.4 years, respectively, for paternal UPD15 cases are increased as compared with normal controls. This may be simply the consequence of an age association with maternal non-disjunction leading to nullisomy for chromosome 15 in the oocyte, although the higher paternal age in paternal UPD15 as compared with maternal UPD15 cases is suggestive that paternal age may also play a role in the origin of paternal UPD15. PMID- 10852370 TI - Aberrant SP-B mRNA in lung tissue of patients with congenital alveolar proteinosis (CAP). AB - Mutations in the surfactant protein (SP)-B gene are responsible for SP-B deficiency in congenital alveolar proteinosis (CAP) (Nogee et al. J Clin Invest 1994: 93: 1860-1883; Lin et al. Mol Genet Metab 1998: 64: 25-35; Klein et al. Pediatrics 1998: 132: 244-248; Ballard et al. Pediatrics 1995: 96: 1046-1052). The multigenerational consanguineous pedigree under study does not carry any of the known mutations, although this pedigree had 14 infant deaths following respiratory distress at birth. Immunostaining of the lungs from three such infants revealed decreased or absent SP-B. By sequencing of SP-B exons, exon intron junctions, and the 5' and 3' flanking regions, nine polymorphisms were found in this pedigree, but none of them could explain the observed SP-B deficiency. Further analysis of SP-B mRNA by reverse transcription-polymerase chain reaction from paraffin-embedded lung tissue of CAP patients showed that SP B mRNA is not intact. Although the sequence of mRNA from exon 1-exon 7 and from exon 8-exon 10 could be amplified, the region between exons 7 and 8 could not. From fluorescence in situ hybridization of the short arm of chromosome 2p, only 2 signals were identified, eliminating the possibility of translocation as the cause of the SP-B mRNA aberrance. Although the nature of the genetic basis of SP B deficiency in this family is currently unknown, the existence of aberrant SP-B mRNA may, at least in part, be responsible for the SP-B deficiency in this pedigree. PMID- 10852371 TI - Genetic testing, ethical concerns, and the role of patent law. AB - This article examines the changing debate over gene patenting and the possible connection between patent law and the ethical and policy concerns associated with the use of genetic testing technologies (e.g. the premature implementation and inappropriate marketing of genetic tests). Arguably, patent law helps to form the market forces that lead to these concerns. It is suggested that existing safeguards fail to control these concerns because of, for example, a lack of provider knowledge and an absence of an adequate regulatory framework. While patent law can be associated with a number of ethical and policy concerns, the article also suggests that patent law may have a positive role in reducing them. Patent law provides policy makers and the public with a focal point - the patent holder - upon which to attach accountability for ethical and legal conduct. The article concludes by inviting policy makers to consider the ways in which patent law could be modified in order to optimize its constructive influence. PMID- 10852372 TI - Perceptions of Ashkenazi Jewish breast cancer patients on genetic testing for mutations in BRCA1 and BRCA2. AB - The perceived benefits and risks of genetic testing may vary between groups of individuals with different cultural, demographic, and family history features. This multicentre study examined the factors that influenced the decision to undergo genetic testing for BRCA1 and BRCA2 in Canadian Jewish women with breast cancer. A self-administered questionnaire was developed and distributed to 134 individuals enrolled in a research-based testing program for Ashkenazi women. The questionnaire assessed demographic, social, and family history parameters, and the influence of medical, family, social, psychological, and cultural/religious factors on decision making about genetic testing. Seventy-six percent of women completed the questionnaire. Forty-one percent of study participants had no family history of breast or ovarian cancer. The most important factors influencing the decision to undergo testing were a desire to contribute to research, potential benefit to other family members, curiosity, and the potential for relief if not found to be a carrier (endorsed by 87, 78, 70, and 60% of participants, respectively). The main perceived risks of undergoing genetic testing related to insurance discrimination, confidentiality, accuracy and interpretability of results, potential impact on marriage prospects for family members, and focus on the Jewish community (endorsed by 28, 24, 30, 17, and 14% of participants, respectively). This study provides novel information on the motivating factors for BRCA1 and BRCA2 mutation testing in Canadian women of Ashkenazi Jewish descent. The focus on altruistic factors and those related to perceived psychological benefits of testing is notable. PMID- 10852373 TI - Clinical presentation and mutation identification in the NBS1 gene in a boy with Nijmegen breakage syndrome. AB - Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder which belongs to the group of inherited chromosomal instability syndromes. The clinical characteristics include severe microcephaly, a dysmorphic facies, and immunodeficiency with predisposition to malignancies. While the cellular characteristics of ataxia teleangiectasia (AT) and NBS are similar, the clinical findings are quite distinct. NBS patients show characteristic microcephaly, which is rare in association with AT and they do not develop ataxia and teleangiectasia. Recently, the gene mutated in NBS has been identified. Here we report a 5-year-old Bosnian boy with severe microcephaly. Because of multiple structural aberrations involving chromosomes 7 and 14 typical for AT (MIM 208900) and NBS (MIM 251260), AT was diagnosed. We suggested the diagnosis of NBS because of the boy's remarkable microcephaly, his facial appearance, and the absence of ataxia and teleangiectasia. DNA analysis was performed and revealed that the boy is homozygous for the major mutation (657de15) in the NBS1 gene. This finding confirms the diagnosis of NBS in our patient and offers the possibility to perform a most reliable prenatal diagnosis in a further pregnancy. PMID- 10852374 TI - Holoprosencephaly, sacral anomalies, and situs ambiguus in an infant with partial monosomy 7q/trisomy 2p and SHH and HLXB9 haploinsufficiency.. AB - We report an infant with holoprosencephaly (HPE), sacral anomalies, and situs ambiguus with a 46,XY,der(7)t(2;7)(p23.2;q36.1) karyotype as a result of an adjacent-1 segregation of a t(2;7)pat. The chromosomal abnormality was diagnosed prenatally after sonographic detection of HPE in the fetus. The baby was born at 37 weeks gestation, and died in the newborn period; he had dysmorphic features consistent with HPE sequence. Postmortem internal evaluation showed semilobar HPE, abdominal situs ambiguus, multiple segments of bowel atresia, dilatation of the ureters, and bony sacral anomalies. Molecular analysis confirmed hemizygosity for the SHH and HLXB9 genes, which are likely to be responsible for the HPE and sacral phenotypes, respectively. Immunohistochemical studies showed intact dopaminergic pathways in the mesencephalon, suggesting that midbrain dopamine neuron induction appears to require only one functioning SHH allele. PMID- 10852375 TI - Germline mutation screening of the STK11/LKB1 gene in familial breast cancer with LOH on 19p. AB - The recently cloned STK11/LKB1 on chromosome 19p has been shown to be a new tumor suppressor gene. Mutations in the LKB1/STK11 gene on chromosome 19p account for most cases of Peutz-Jeghers syndrome (PJS), in which intestinal hamartomas are associated with elevated risks of several cancer types, including breast cancer. A previous study revealed that familial breast cancer is associated with loss of heterozygosity (LOH) on 19p. To establish whether germline mutations of STK11/LKB1 account for familial breast cancer, 22 patients from 14 breast cancer families with LOH on 19p and one PJS family were selected for screening for germline mutations of LKB1/STK11. A combination of polymerase chain reaction (PCR)-heteroduplex, single-strand conformational polymorphism (SSCP) analyses, Southern blot analysis and direct sequencing were used for mutation detection. No mutations were identified. Germline mutations of LKB1/STK11 did not contribute to breast cancer in these families. PMID- 10852376 TI - A novel mutation in the HEXA gene specific to Tay-Sachs disease carriers of Jewish Iraqi origin. AB - An increased frequency of carriers of 1:140, as defined by reduced hexosaminidase A (HexA) activity, was observed among Iraqi Jews participating in the Tay-Sachs disease (TSD) carrier detection program. Prior to this finding, TSD among Jews had been restricted to those of Eastern European (Ashkenazi) and Moroccan descent with carrier frequencies of 1:29 and 1:110 for Jews of Ashkenazi and Moroccan extraction, respectively. A general, pan-ethnic frequency of approximately 1:280 has been observed among other Jewish Israeli populations. Analysis of 48 DNA samples from Iraqi Jews suspected, by enzymatic assay, to be carriers revealed a total of five mutations, one of which was novel. In nine carriers (19%), a known mutation typical to either Ashkenazi or Moroccan Jews was identified. DeltaF304/ 305 was detected in four individuals, and + 1278TATC in three. G269S and R170Q each appeared in a single person. The new mutation, G749T, resulting in a substitution of glycine to valine at position 250 has been found in 19 of the DNA samples (40%). This mutation was not detected among 100 non-carrier, Iraqi Jews and 65 Ashkenazi enzymatically determined carriers. Aside from Ashkenazi and Moroccan Jews, a specific mutation in the HEXA gene has now also been identified in Jews of Iraqi descent. PMID- 10852377 TI - Mild phenotype in two siblings with distal monosomy 12p13.31-->pter. AB - We report two sibs with trisomy for the region 2p25.1--> pter and monosomy for the region 12p13.31--> pter, due to adjacent-1 segregation of a maternal balanced reciprocal translocation, 46,XX,t(2;12)(p25.1;p13.31). These sibs presented with a mild phenotype, but nevertheless showed features of each of the contributing aneusomies. Monosomy 12p has previously been considered to have a variable and indistinct phenotype. Comparison of these patients with previous reports showed that many features, including microcephaly, facial dysmorphia, developmental and growth delay and dental and digital anomalies are frequently associated with monosomy for 12p. Many of these features are common to other aneusomies, thereby mitigating against a distinct 12p monosomy syndrome at this time. However, the combination of digital and dental anomalies may suggest the presence of this particular monosomy. The proband and his sister had some of the more non-specific features of 2p trisomy syndrome, and comparison with previous reports suggested that the characteristic 2p trisomy syndrome is more usually associated with larger or more proximal trisomies of 2p. PMID- 10852378 TI - A patient with maternal chromosome 14 UPD presenting with a mild phenotype and MODY. PMID- 10852379 TI - Limited plasticity in the recognition of peptide epitope variants by an alloreactive CTL clone correlates directly with conservation of critical residues and inversely with peptide length. AB - Although self-restricted T cells are peptide-specific and can distinguish among closely related ligands, they have some flexibility in the recognition of sequence variants of their natural peptide epitopes. Alloreactive cytotoxic T lymphocytes (CTL) can recognize specific peptides bound to the allo-major histocompatibility complex (MHC) molecule, but their plasticity in the recognition of related peptide variants has not been properly defined. The anti B*2705 alloreactive CTL 27S69 specifically recognizes a natural octamer ligand of HLA-B*2705. In this study, we tested the recognition of a nested set of epitope variants by this CTL clone. Although none of these peptides was recognized equally as the natural epitope, two of the peptide variants were recognized with only slightly decreased efficiency. Peptide sensitization assays showed that CTL recognition of epitope variants correlated directly with conservation of two non anchor residues that were critical for recognition of the natural epitope, and inversely with peptide length. Molecular modeling of the peptide variants complexed with B*2705 provided a rational explanation for their differential recognition. Location of the two critical peptide residues at the right three dimensional space favored efficient recognition by CTL 27S69. The negative effect of increasing peptide length on recognition was due to the bigger bulging surface between the two critical residues, which precluded for optimal interaction with the specific T-cell receptors (TCR). Our results demonstrate that an alloreactive CTL has a degree of plasticity in the recognition of peptide epitope variants that is comparable to that of peptide-specific self-restricted CTL, and define the structural features determining crossreaction among related peptides. PMID- 10852380 TI - Analysis of three HLA-A*3303 binding peptide anchors using an HLA-A*3303 stabilization assay. AB - The affinity of 232 8- to 11-mer peptides carrying HLA-A*3303 anchor residues at position 2 (P2) (Ala, Ile, Leu, Val, Phe or Tyr) and the C-terminus (Arg) was analysed by a stabilization assay using RMA-S transfectants expressing HLA-A*3303 and human beta2-microglobulin. One hundred and nineteen of these peptides (51.3%) bound to HLA-A*3303, confirming that these residues are anchors for HLA-A*3303. Evaluation of P2 residues demonstrated that binding of peptides with Phe or Tyr at P2 is stronger than that of peptides with aliphatic hydrophobic residues at P2. This was confirmed by analysis of a panel of peptides mutated at P2. Analysis of the C-terminal mutant peptides showed that substitution of Lys for Arg had minimal influence on binding to HLA-A*3303. This implies that peptides carrying HLA-A*1101 anchor residues (Val, Ile, Phe or Tyr at P2 and Lys at the C-terminus) can bind to HLA-A*3303. However, such peptides showed lower binding for HLA A*3303 than for HLA-A*1101. Thus, Arg at the C-terminus is much stronger anchor for HLA-A*3303 than Lys. The preference for Arg and Lys at the C-terminus by HLA A*1101 and HLA-A*3303 respectively may be due to sequences of three residues (70, 97 and 114) forming the F-pocket of these HLA class I molecules. Statistical analysis of 232 peptides further showed a positive effect of negatively charged residues at P1 for peptide binding to HLA-A*3303. Thus, residues at P1, P2 and the C-terminus play an important role in peptide binding to HLA-A*3303. PMID- 10852381 TI - HLA-DQ/human CD4-restricted immune response to cockroach allergens in transgenic mice. AB - We investigated the immune response to the German cockroach (Blattella germanica), and one of its major antigens, Blattella germanica group 5 (Bla g 5), in a double-transgenic, double-knockout mouse expressing human HLA-DQ8, HLA-DQ6 and CD4 molecules in the absence of mouse class II and mouse CD4. Transgenic mice were primed and challenged with CR extract or individual synthetic peptides representing Bla g 5. Strong T-cell responses to CR extract were detected in both HLA-DQ/hCD4+ transgenic mice. The responses were two times lower in mice expressing HLA-DQ molecule in the context of mouse CD4. Under similar treatment, no responses were found in the double-knockout Abetadegrees/mCD4degrees mice and in mice expressing human CD4 molecule alone. HLA-DQ/hCD4+ mice produced primarily interleukin (IL)-5, IL-10, and IL-13. Minimal amounts of IL-4 were detected only in HLA-DQ6/ hCD4+ mice. Interferon (IFN)-gamma production was low in both transgenic mouse, suggesting a predominantly T-helper 2 (Th2)-type response. Cockroach allergen extract immunized HLA-DQ8/hCD4+ mice recognized only one of the 20 peptides of Bla g 5 while HLA-DQ6/hCD4+ mice responded primarily to three peptides. Primed with individual peptides, both HLA-DQ/hCD4+ mice responded maximally to peptides 10 (residues 91-110) and 17 (residues 161-180). In addition, HLA-DQ6/hCD4+ mice responded to peptide 16 (residues 151-170). Thus, peptides 10 and 17 contained the major HLA-DQ-restricted hCD4+ T-cell epitopes and could be recognized by both HLA-DQ8 and HLA-DQ6 transgenic mice. Transgenic mice represent a new tool for investigating the immune responses to cockroach allergen. Our results suggest that therapeutic strategies aimed at developing antagonist peptides might be a useful treatment (immunotherapy) for allergic asthma. PMID- 10852382 TI - HLA DRB1*1501 and intrathecal inflammation in multiple sclerosis. AB - CD4 T cells are considered to be pivotal in the pathogenesis of multiple sclerosis (MS), and the human leukocyte antigen (HLA) haplotype associated with DRB1*1501 confers susceptibility to MS in patients of Northern European descent. Some previous studies have suggested an association of DRB1*1501 with T- and B cell reactivity to specific myelin protein peptides, other studies suggested an association with enhanced cytokine production or intrathecal immunoglobulin (Ig) synthesis. In order to further assess the role of DRB1*1501 in the pathogenesis of MS, we studied intrathecal inflammation and T-cell phenotypes in patients with possible onset symptoms or clinically definite MS. Presence of DRB1*1501 was associated with higher levels of cerebrospinal fluid (CSF) inflammation as assessed by IgG synthesis levels and higher levels of matrix metalloproteinase-9 activity. DRB1*1501-positive patients also had a lower percentage of T cells in CSF expressing HLA-DR without co-expressing CD25. These findings suggest that enhanced intrathecal inflammation and an altered T-cell activation status may be of importance in conferring the DRB1*1501-associated susceptibility to MS. PMID- 10852383 TI - Different gene loci within the HLA-DR and TNF regions are independently associated with susceptibility and severity in Spanish rheumatoid arthritis patients. AB - The aim of this study was to investigate whether polymorphisms in the tumor necrosis factor (TNF) and HLA-DRB1 gene regions are independently associated with rheumatoid arthritis (RA) in a population from Lugo region of northwestern Spain. RA patients (n=179) attending hospital outpatient clinics in Lugo, northwestern Spain and matched controls (n=145) were recruited. RA susceptibility in this population was predominantly associated with DRB1*0401, while erosive disease was associated with HLA-DRB1*0101 and DRB1*04. The increase in DRB1*04 was accounted for by an increase in DRB1*0404 and *0405 but not *0401 frequencies. In contrast, *0401 frequency was significantly increased in seropositive patients. The rheumatoid arthritis shared epitope (SE) was associated with increased risk for seropositive and erosive disease and this appeared to operate in a dose-dependent manner. Logistic regression analyses revealed that the TNF microsatellite markers TNFc1 and b3 were associated with RA independently of DRB1*04 and the SE. Carriage of a TNF c1 allele provided an increased risk of RA in SE-negative and SE-heterozygous individuals. TNFc1 and TNFb3 were not associated with erosive or seropositive disease. In contrast, TNF a2 was significantly associated with erosive disease which was independent of DRB1*04 and the SE. Further studies will be needed to establish why (TNFc1) polymorphism seemingly associated with low TNFalpha production, is a risk factor for RA. PMID- 10852384 TI - HLA class I antigen downregulation by interleukin (IL)-10 is predominantly governed by NK-kappaB in the short term and by TAP1+2 in the long term. AB - The present study was designed to determine the molecular mechanisms by which interleukin (IL)-10 prevents the HLA class I antigen expression at the cell surface. In this context, the potential role of transporter associated with antigen presentation 1+2 (TAP1+2) molecules and NF-kappaB transcription factors was addressed. The IL-10 effect was investigated in a human lymphoblastoid cell system defective for TAP1+2 genes (T2 cell line) and in the related TAP1+2 transfectants (T3 cell line). In this experimental system, after 48 h of incubation in the presence of IL-10, the HLA class I antigen downmodulation was observed in the T3 but not in the T2 cell line, suggesting a potential role of TAP1+2 molecules. In the same experimental conditions, the NF-kappaB activity was unaffected. Instead, after 3 h of exposure to IL-10, the HLA downmodulation was observed in both cell lines, the NF-kappaB factors activity being strongly reduced. In addition, the transfection of the inhibitor of NF-kappaB, IkappaBalpha, prevented the IL-10 effect on HLA class I antigen expression in the T3 cell line. This phenomenon was observed after 3 h but not 48 h of IL-10 incubation. These evidences indicate a time dependent involvement of TAP1+2 antigens and of NF-kappabeta activity in the IL-10-induced major histocompatibility complex (MHC) class I downmodulation. PMID- 10852385 TI - Increased beta2-microglobulin-free HLA class I heavy chain serum levels in the course of immune responses to viral antigens and to mismatched HLA antigens. AB - Besides being present in serum in association with beta2-mu, HLA class I heavy chains are also present in serum as beta2-micro-free moieties. The increase in serum levels of beta2-micro-associated HLA class I heavy chains in conditions associated with an activation of the immune system have prompted us to measure the serum levels of beta2-mu-free HLA class I heavy chains in the course of immune responses to viral antigens and to mismatched histocompatibility antigens. The serum level of beta2-mu-free HLA class I heavy chains, like that of beta2-mu associated HLA class I heavy chains was significantly increased in patients affected by advanced HIV-1 infection or by chronic hepatitis C (CHC). In the latter group of patients an association was found between a reduction in the beta2-mu-free HLA class I heavy chain serum level and response to therapy with interferon alpha and ribavirin. Moreover, the beta2-mu-free HLA class I heavy chain serum level was increased more than that of beta2-mu-associated HLA class I heavy chains during episodes of liver ischemia following liver transplantation and in the course of acute graft rejection and of acute graft-versus-host-disease (GVHD) after allogeneic bone marrow transplantation (BMT). These results suggest that the serum levels of beta2-mu-free and beta2-mu-associated HLA class I heavy chains are independently regulated. Furthermore, beta2-mu-free HLA class I heavy chain serum level may be a useful marker to monitor response to therapy in CHC patients and the clinical course of liver and bone marrow grafts. PMID- 10852386 TI - A porcine cell surface receptor identified by monoclonal antibodies to SWC3 is a member of the signal regulatory protein family and associates with protein tyrosine phosphatase SHP-1. AB - SWC3 was defined at the First International Swine CD Workshop as a specific myelomonocytic antigen of 230 kDa with mAbs 74-22-15, 6F3 and DH59B. In this report, we describe two new mAbs (BL1H7 and BA1C11) that react selectively with granulocytes, monocytes and macrophages. These monoclonal antibodies (mAbs) recognize a molecule in the range of 90-115 kDa in immunoprecipitation and/or Western blotting analyses. Two-colour FACS analyses showed that the distribution of BL1H7 and BA1C11 antigens was identical to that of SWC3. Moreover, in this assay, mAb 74-22-15 appeared to partially block the binding of mAbs BL1H7 and BA1C11, suggesting that all these mAbs reacted with the same or spatially close epitopes. Cross-blocking analyses indicated that it was the case with mAbs 74-22 15 and BL1H7. Immunoprecipitation experiments with mAbs 74-22-15, BL1H7 and BA1C11, followed by immunoblotting with mAb BL1H7 confirmed that all three mAbs recognize the same molecule. Analysis of the N-terminal sequence carried out on the affinity purified protein revealed homology with members of signal regulatory protein (SIRP) family. Like other members of this family, after treatment with sodium pervanadate, SWC3 became phosphorylated in tyrosines, and associated with the protein-tyrosine phosphatase SHP-1. PMID- 10852387 TI - Large-scale DNA-based typing of HLA-A and HLA-B at low resolution is highly accurate specific and reliable. AB - DNA-based typing of HLA class I alleles of the HLA-A and HLA-B loci using sequence-specific oligonucleotide primers and/or probes has been used for the large-scale typing of individuals for the National Marrow Donor Program unrelated donor registry. Typing was performed by 16 laboratories at a low level of resolution (e.g. A*01, B*07). The results of blinded quality control analysis for the first 12 months of the project show the typing to be highly accurate, specific and reliable. The total error rate based on 11,545 HLA-A and 11,428 HLA B assignments was 1.1% for HLA-A and 1.9% for HLA-B. This level of accuracy is particularly remarkable because the quality control samples could not be distinguished from 64,180 donor samples tested at the same time by the laboratories. PMID- 10852388 TI - Tumor necrosis factor genomic polymorphism in Spanish IGA deficiency patients. AB - Selective IgA deficiency (IgAD) is the most common form of primary immunodeficiency. Its association with genes within the major histocompatibility complex (MHC) has been repeatedly reported. Recently the susceptibility gene has been located in the class III region, around the tumor necrosis factor (TNF) cluster. In this study we have examined IgAD association with TNF-alpha gene promoter polymorphisms and TNFa and b microsatellites. No significant association was found with the former polymorphisms and the observed associations with TNFa2 allele and haplotypes TNFa2b1 and TNFa2b3 were proven to be secondary to their occurrence on the B14-DR1 and B8-DR3 haplotypes, previously reported to be associated with susceptibility to IgAD. However, a primary negative (protective) association was found between the TNFa10 allele and IgAD. PMID- 10852389 TI - Sequence-based typing of HLA class II DQB1. AB - Due to the expanding number of known HLA class II DQB1 alleles, high-resolution oligotyping is becoming ineffective, therefore a sequence-based typing (SBT) strategy was developed to provide rapid and definitive typing of HLA-DQB1. HLA DQB1*02, *03, *04, *05, and *06 alleles were individually amplified by polymerase chain reaction (PCR) using exon 2 group-specific primers. Forward and reverse PCR primers were tailed with M13 universal and M13 reverse sequences, respectively. Subsequent bi-directional cycle-sequencing was carried out using Cy5.5-labeled M13 universal primer and Cy5.0-labeled M13 reverse primer. Automated sequencing was performed in 30 min using a Visible Genetics, Inc. (VGI) MicroGene Clipper Sequencer. Full concordance was observed between this SBT method and oligotyping among 151 individuals. PMID- 10852390 TI - Seventeen more novel HLA-A locus alleles. AB - This paper describes seventeen novel HLA-A locus alleles: A*0106, *0235, *0236, *0237, *1105, *2302, *2303, *24032, *2422, *2424, *2503, *2613, *3007, *3203, *3204, *6809 and *6810. All alleles were identified due to unexpected probe reaction patterns during routine SSOP typing. Exons 2 and 3 of these alleles were subsequently characterized by DNA sequencing. The alleles represent a shuffling of sequence motifs, likely by interallelic conversion, expanding the diversity of the HLA system. PMID- 10852391 TI - A novel HLA-B*40 allele and novel exon 1 sequences of two B*40 alleles identified in potential marrow donors. AB - Sequence-specific oligonucleotide probe hybridization and sequence-specific primer polymerase chain reaction (PCR) typing suggested the presence of variants of HLA-B*40 in three individuals. Two were part of 3,500 potential marrow donors being screened for the National Marrow Donor Program, while the third was a clinical specimen. PCR products encompassing HLA-B locus exons 1 through 3 were prepared and subcloned. In one individual, a native of the Pacific Islands, sequencing revealed a novel HLA-B*40 allele (B*4023). In two other individuals, a previously unknown exon 1 sequence was determined for HLA-B*4016 (ethnicity unknown) and B*4020 (Hispanic). These findings further illustrate the substantial genetic variation present at the HLA-B locus within human populations. PMID- 10852392 TI - A novel HLA-B null allele (B*4022N) generated by a nonsense codon in the alpha1 domain. AB - We describe in this work a novel HLA-B null allele designated B*4022N. This new variant was found in a Caucasian individual who was serologically typed for one HLA-B allele as a B-blank, Bw-blank. Retrospective DNA typing by polymerase chain reaction using sequence-specific primers (PCR-SSP) has established the correspondence of this blank allele with the classical HLA-B*4001 allele. Nucleotide sequence analysis of exon 2 and 3 has revealed the presence of two adjacent point mutations at position 170 and 171 of exon 2 (GG to TT). While the first difference is silent, the second leads to the creation of a nonsense codon at position 58 of the alpha1 domain, providing the most likely mechanism underlying the observed null phenotype. PMID- 10852393 TI - Nomenclature for factors of the HLA system, update December 1999. WHO Nomenclature Committee for Factors of the HLA System. PMID- 10852394 TI - The evaluation of the potential of botulinum C3 enzyme as an exogenous differentiation inducing factor to neurons. AB - Botulinum C3 enzyme produced by Clostridium botulinum type C and D strains modifies Rho proteins. In a previous study, we observed that the LDH isozyme pattern of neurons treated with C3 enzyme was different from that induced with endogenous growth factor of neurons such as NGF [21]. This type of change is considered to have an advantage in the medical use of C3 enzyme for neural disorder. To determine the functional similarity of C3-treated neurons to control and NGF-treated neurons, we examined the responses of C3-treated neurons to various drugs, including some neurotransmitters, by measuring the rise of intracellular Ca ions into the neurons. The time course of the rise of intracellular Ca ions induced by high concentration of potassium in the C3 treated neurons was similar to that in the NGF-treated neurons. The C3-treated neurons responded to glutamic acid, aspartic acid, kainic acid, gamma aminobutylic acid, muscarine and ACh with similar time courses and magnitudes as the control neurons. These results suggest that the C3 enzyme induces the functional differentiation of neurons, and that C3 enzyme has the potential for the medical use as an exogenous differentiation-inducing factor of neurons. PMID- 10852395 TI - Antibacterial effect of chloramphenicol, thiamphenicol and florfenicol against aquatic animal bacteria. AB - The minimum inhibitory concentration (MIC) was measured to evaluate the antibacterial activities of chloramphenicol (CP), thiamphenicol (TP) and florfenicol (FFC) against the aquatic bacterial isolates from soft-shell turtles, fish and shellfish. Amoxicillin (AMPC), oxytetracycline (OTC) and oxolinic acid (OA) were included to compare with above protein synthesis inhibitors. The results showed that the order of MIC range of the isolates from soft-shell turtles for tested drugs was OA>FFC, CP>TP> AMPC, OTC. The percentage of the resistant strains indicated that OA was the lowest (7.14%) and OTC was the highest (85.07%). The order of antibacterial activity against the isolates from fish was OA>FFC>CP>AMPC>OTC>TP. The percentage of the resistant strains revealed that OA (13.64%) and OTC (80.91%) were the lowest and the highest, respectively. For the isolates from shellfish, the order of antimicrobial activity was OA>CP, FFC>AMPC, OTC, TP. TP showed the greatest percentage of the resistant strains (58.7%), but that of OA was the lowest (4.35%). The most common resistant patterns of the isolates from turtles, fish and shellfish were AMPC-OTC, CP-TP AMPC-OTC, and FFC-CP-TP-AMPC-OTC, respectively. There were partially-complete resistance of the resistant isolates among CP, TP and FFC. The findings indicated that previous treatment might affect the choice of drug to use for aquatic bacterial diseases. PMID- 10852396 TI - Infectivity to experimental rodents of Cryptosporidium parvum oocysts from Siberian chipmunks (Tamias sibiricus) originated in the People's Republic of China. AB - We isolated Cryptosporidium parvum-type oocysts from naturally infected siberian chipmunks which originated in the People's Republic of China and examined the infectivity to rodents as experimental animals. The naturally infected chipmunks did not show any clinical symptoms. The oocysts were 4.8 x 4.2 microm on average in size. They were ovoid and morphologically similar to the C. parvum oocysts isolated from human and cattle. Experimental rodents were inoculated with 1.6 x 10(6) original oocysts each. SCID mice began to shed oocysts on day 7 and the OPG value was 10(5) from 50 days. The oocysts were found from ICR mice on days 13 and 16 by only sugar flotation method, however, any oocysts were not detected from the rats, guinea pigs and rabbits until 30 days. Two infected SCID mice were necropsied on days 100 and 102 and examined for coccidian organisms. Merozoites and oocysts were found in the low part of jejunum and ileum, however, no parasites were detected in the stomach. Consequently, it was considered that the present species was C. parvum and was probably genotype 2 from result of infectivity to rodents. PMID- 10852397 TI - D0870, an antifungal agent, induces reverse use-dependent QT prolongation in dogs. AB - We previously reported that D0870 induced QT prolongation and sudden death due to torsades de pointes (TdP) in dogs and that catecholamines played an important part in the development of the sudden death. In the present study, we analyzed in detail the ambulatory electrocardiographic recordings obtained from the just mentioned study to elucidate the mechanism of the onset of TdPs and conducted an in vitro study using isolated canine Purkinje fibers to assess the effect of D0870 on repolarization. The hearts with TdPs observed before the sudden death showed a higher sinus rate for 5 and 10 sec before the onset, a shorter coupling interval, and a higher ventricular tachycardia rate compared with those having the non-sustained TdPs. These findings suggest that D0870-induced fatal TdPs may be provoked by a triggered activity developed from delayed after depolarizations. In contrast, as the pause-dependent, non-sustained TdPs in bradycardia showed a typical "short-long-short" sequence, they may be developed from early afterdepolarization . Moreover, the results of the in vitro study supported our contention that D0870 induced QT prolongation in a reverse use-dependent manner in vivo and suggested that it may inhibit not only rapidly activating delayed rectifier potassium current (Ik(r)) but also L-type Ca current (I(ca-L)). PMID- 10852398 TI - 8-Difluoromethoxy-4-quinolone derivatives as anti-feline immunodeficiency virus (FIV) agents: important structural features for inhibitory activity of FIV replication. AB - The inhibitory activities of various 8-difluoromethoxy-4-quinolone derivatives against feline immunodeficiency virus (FIV) replication in the chronically infected cell line P-CrFK were investigated. Certain derivatives were found to inhibit FIV production from P-CrFK cells in a dose-dependent manner without exhibiting cytotoxic effects at inhibitory concentrations. Based on this study, the structures important for anti-FIV activity are suggested to be (i) a carboxyl group at position C-3, and (ii) an aromatic modification at position 4 of the C-7 piperazinyl moiety. PMID- 10852399 TI - Effects of TAK-044, a nonselective endothelin receptor antagonist, on the spontaneous and indomethacin- or methylene blue-induced constriction of the ductus arteriosus in rats. AB - We studied the effects of TAK-044, a nonselective endothelin (ET) receptor antagonist, on the indomethacin- or methylene blue-induced constriction of the ductus arteriosus (DA) in rats and compared them with the effects on spontaneous DA constriction. Injection of TAK-044 into 21-day-old fetuses in utero was performed through the uterine wall of laparotomized mother rats under light ether anesthesia. The fetuses were autopsied 3 hr after treatment with TAK-044 (10 mg/kg) in utero and simultaneous administration to the laparotomized mother rats of indomethacin (3 mg/kg, p.o.) or methylene blue (100 mg/kg, i.p.). In the second experiment, pregnant rats were decapitated on day 21 of gestation to obtain newborn rats by cesarean delivery. Newborn rats which were given TAK-044 (2, 10 mg/kg) immediately after or 1 hr before cesarean delivery were autopsied at various times after birth. In both experiments, pups were rapidly frozen in an acetone-dry ice mixture at autopsy to evaluate the DA constriction by the whole body freezing and shaving method. TAK-044 injection into the fetus 3 hr before autopsy completely inhibited the DA constriction induced by maternal treatment with indomethacin or methylene blue. TAK-044 caused dose-dependent inhibition of the spontaneous closure of the DA after birth. The inhibitory effect was more pronounced in pups which were given TAK-044 in utero 1 hr before birth; however, the inhibitory effect was incomplete in newborn pups. These results, together with the previous finding that BQ-123, an ETA-specific receptor antagonist, inhibits the ductal constriction induced by oxygen in vitro [Coceani et al., 1992], indicate that the ETA receptor plays a significant role in the indomethacin- or methylene blue-induced DA constriction as well as in the spontaneous DA constriction after birth, and also indicate that the inhibition of ETA receptor by TAK-044 was more easily achieved in fetuses than in neonates. PMID- 10852400 TI - The developmental study on lamination of the optic tectum in relation to the retinotectal projection in chicks and chick embryos. AB - The tectal lamination was investigated in the central part of the chick embryonic tectum. Two and 5 layers were observed above the neuroepithelium (NE) on embryonic day 6 (E6) and E8, respectively. Optic fibers extended on the surface of the tectum by E8. On E10-11, the outer tectum was composed of 2 layers, that is, a fibrous layer forming the optic fiber layer on the tectal surface and a cellular layer showing the gradient of cell density. In the inner tectum, the lamination was almost completed. On E12-13, the outer tectal layers, which showed the gradient of cell density, was divided into dark and light cellular layers. The dark cellular layer was divided into 2 layers on E14-15 and further into 4 layers (layer C-F in chick) on E18. On the other hand, the light cellular layer did not change until E18, but finally, it was divided into 2 layers (layer A and B in chick) by E20. Optic fibers reached the bottom of the outer tectum by E14 showing different densities of terminals. Stratification by optic fibers was going to step into the final stage on E18. On E20, laminations according to cytoarchitectural features and the optic fiber terminals were substantially completed. In the tectum affected by destruction of the contralateral embryonic eye (E4), some cellular layers were incompletely discriminated by differences of cell density. PMID- 10852401 TI - Osteometrical skull character in the four species of tree shrew. AB - The skull size and shape were osteometrically examined in the four species of the tree shrews (Tupaia tana, T. javanica, T. minor and T. dorsalis). We suggest that the skull characters were affected by the species specific behavior and terrestrial, arbo-terrestrial and arboreal life, among the genus Tupaia. The neurocranium was laterally narrower in the braincase area, and the splanchnocranium was longer only on dorsal side in T. tana, and these characters were opposite to T. minor. The principal component analysis confirmed the obviously separated clusters among T. tana, T. javanica and T. minor, affected by the adaptation for each behavior. T. dorsalis was considered as terrestrial species from the results of proportion analysis and the principal component analysis. PMID- 10852402 TI - Hump attachment structure of the two-humped camel (Camelus bactrianus). AB - The hump attachment structure was morphologically examined in the two-humped camel (Camelus bactrianus). The cranial hump is fixed by the trapezius and rhomboid muscles in the thoracic region. The strong collagen sheet in the basement of the hump is attached to the segmented bellies of the trapezius muscle, and the thoracic rhomboid muscle and the nuchal-supraspinous ligament support the attachment function of the trapezius muscle. The basement sheet possesses the line structure of collagen fibers, which are fitted to the segmented bundles of the trapezius muscle, and we observed that the muscle cells of the trapezius muscle are intermingled with the collagen fibers around the attachment line structure. In contrast, the caudal hump is directly attached to the subcutaneous tissue in the superficial region of the lumbar longissimus and lumbar iliocostal muscles. These findings demonstrated that the caudal hump of the two-humped camel is consistent with the hump of the one-humped camel in the attachment structure. PMID- 10852403 TI - Neospora caninum infected the alimentary tract of nude mice and was transmitted to other mice by intraperitoneal inoculation with the intestinal contents. AB - Neospora caninum (BT-2 strain) that originated from the brain of a Holstein calf was serially passaged through 10 generations of BALB/c nude mice by intraperitoneal inoculation. Histological examination of the mice revealed that numerous clusters of tachyzoites appeared in the pancreas, stomach and small intestine as well as in the central nervous system (CNS) and skeletal muscles. Intestinal contents of the infected mice were inoculated intraperitoneally into uninfected nude mice and 3 of the 17 inoculated mice showed clinical signs at post inoculation days 3 to 10. The present experiments demonstrated a proliferation of N. caninum tachyzoites in the mucosa of the alimentary tract and pancreas of the nude mice and the intestinal contents of the mice were infective to other nude mice. PMID- 10852404 TI - Azoospermia of dogs with apoptotic germ cells and Leydig cells. AB - Apoptotic cell death in the testes of 4 dogs with azoospermia was examined. Blood plasma luteinizing hormone (LH), testosterone (T), and estradiol-17beta (E2) concentrations, and testicular transferrin (Tf) concentration as a marker of Sertoli cell function were measured in the 4 azoospermic dogs and in 5 normal dogs. The spermatids in 2 of the 4 azoospermic dogs and the Leydig cells in 3 of them exhibited apoptotic cell death. Mean LH, E2, and Tf concentrations in the 4 azoospermic dogs were significantly higher than in the normal dogs (P<0.01). These findings suggested that the azoospermia in all 4 dogs might has been caused by abnormal functions of Sertoli cells as well as Leydig cells. PMID- 10852405 TI - Effects of orvus ES paste on canine spermatozoal longevity after freezing and thawing. AB - Because of weak resistance of canine sperm to freezing, an applicable method of preparing canine frozen semen has not yet been established. We added various concentrations of Orvus ES Paste (OEP) to egg yolk Tris-fructose citrate, and investigated its effectiveness on survival of spermatozoa. Addition of 0.5-1.0% OEP to the extender for freezing canine semen was effective in prolonging post thaw survival of spermatozoa. PMID- 10852406 TI - Effect of addition of Orvus ES paste to frozen canine semen extender on sperm acrosomes. AB - Using the triple-stain technique, we investigated whether sperm acrosomes in frozen canine semen were protected during freezing and thawing by addition of a surfactant, Orvus ES Paste (OEP), to the extender. Acrosomes were clearly shown to be protected by the addition of OEP to the entender when compared with those in sperm frozen without OEP addition (p<0.05). PMID- 10852407 TI - Malignant fibrous histiocytoma in a Djungarian hamster. AB - A subcutaneous malignant fibrous histiocytoma (MFH) was observed in the region between the right posterior trunk and right hind limb of a 2-year-old male Djungarian hamster weighing 45 g. Histologically, the tumor consisted of bizarre multinucleated giant cells, histiocytic cells, and fibroblastic cells with a storiform pattern, and was considered to be of the storiform-pleomorphic type of MFH. Severe nuclear atypia with prominent nucleoli and many mitotic figures was also observed. Electron microscopy demonstrated fibroblastic cells and histiocytic cells. The fibroblastic cells were spindle-shaped, and sometimes had an invaginated nucleus. The histiocytic cells were polygonal with an oval or kidney-shaped nucleus. The cytoplasm of both cells contained numerous free ribosomes, small amounts of rough endoplasmic reticulum, and round mitochondria. Tumor cells were immunohistochemically positive for vimentin, and were thought to be of undifferentiated mesenchymal cell origin. This is the first report of spontaneous MFH in a hamster. PMID- 10852408 TI - Lectin-binding capacity of glycoconjugates in Escherichia coli 09:K103:NM, 987P+ST+-infected porcine lower small intestines. AB - Composition of glycoconjugates were investigated in Escherichia coli 09:K103:NM, 987P+ST+-infected lower small intestines of 1-week-old pigs by the use of twenty one biotinylated-labelled lectins with avidin-biotin-peroxidase complex method. Piglets with experimental group were inoculated by feeding 5 ml of culture inoculum (5 x 10(9) colony-forming units/ml) with 15 ml of milk replacer. At the onset of diarrhea, experimental piglets and time-matched control piglets were euthanatized using electrocution, necropsied, and tested by lectin histochemistry. As compared with control, staining intensity of seven lectins altered in ileal villus brush border and goblet cells of pigs inoculated with the pathogen. PMID- 10852409 TI - Effects of a canine Elizabethan collar on ambulatory electrocardiogram recorded by a Holter recording system and spontaneous activities measured continuously by an accelerometer in Beagle dogs. AB - Ambulatory electrocardiogram (ECG) has been recorded in dogs wearing a jacket to protect a Holter recording system, but the jacket was often damaged by dogs. We compared ECG recorded by a Holter recording system and spontaneous activity measured by an accelerometer in Beagle dogs with or without an Elizabethan collar. There were few significant differences in mean values (per hr) of the heart rate and the amount of spontaneous activity between dogs with or without the Elizabethan collar. Mean values (per 23 hr) of them had no significant difference between them. We concluded that the Elizabethan collar did not have any effect on ambulatory ECG and canine movements and was effective to protect the recording apparatus. PMID- 10852410 TI - Spontaneous Aleutian disease in a ferret. AB - A 3-year-old female ferret died five days after admission to a veterinary clinic for treatment of acute dyspnea and posterior paresis. Blood chemistry showed no hypergammaglobulinemia. Histopathological examination revealed mild to severe inflammatory infiltrates, composed mostly of plasma cells, in multiple organs. Lesions were especially severe in the kidneys, where focal segmental membranous glomerulopathy was also present. In the liver, in addition to lymphocytic and plasmacytic infiltration in periportal areas, dilatation and proliferation of the bile ducts were seen. On analysis of PCR products, using primers directed against the gene encoding Aleutian disease (AD) viral capsid and formalin-fixed kidney samples, we detected a single band of about 400 bp, specific to the AD virus. PMID- 10852411 TI - Risk factors in causing outbreaks of food-borne illness originating in schoollunch facilities in Japan. AB - We reviewed records of all outbreaks of food-borne illnesses due to schoollunch in Japan from 1987 through 1996 to determine the risk factors causing these outbreaks. Major hazards in 269 outbreaks were Salmonella spp., Campylobacter jejuni, Escherichia coli and Staphylococcus aureus. Foods including uncooked or partially cooked items, salad or egg products presented a high risk in 62 outbreaks with confirmed food sources. Contaminated food items were involved in 29 incidents (46.8%); storage of foods for an extended period before serving in 29 incidents (46.8%), inadequate cooking and cross contamination in 21 incidents (33.9%) each; infected employees in nine incidents (14.5%). PMID- 10852412 TI - Earl Sutherland--Nobel Prize for Hormone Research. PMID- 10852413 TI - The necessity of cooperation in medicine. PMID- 10852414 TI - The persistent importance of autopsies. PMID- 10852415 TI - The Women's Health Initiative: a heart-to-HRT conversation. PMID- 10852416 TI - Comparison of premortem clinical diagnoses in critically iII patients and subsequent autopsy findings. AB - OBJECTIVE: To determine whether our practice of requesting an autopsy for patients who die in the medical intensive care unit (MICU) continues to be a valid approach to obtain clinically and educationally relevant findings. METHODS: In this retrospective study conducted in an adult MICU population of a university hospital, the clinical diagnoses and postmortem major diagnoses of 100 patients who died in 1996 (autopsy rate of 93%) were compared. RESULTS: Eighty-one percent of the clinical diagnoses were confirmed at autopsy. In 16%, autopsy findings revealed a major diagnosis that, if known before death, might have led to a change in therapy and prolonged survival (class I missed major diagnoses). The most frequent class I missed major diagnoses were fungal infection, cardiac tamponade, abdominal hemorrhage, and myocardial infarction. Another 10% of autopsies revealed a diagnosis that, if known before death, would probably not have led to a change in therapy (class II error). CONCLUSIONS: Autopsy remains an important tool for education and quality control. In contrast with historical series of 1 to 2 decades ago, there is a clear shift in the type of class I missed major diagnoses toward opportunistic infections. Bedside-applicable techniques such as electrocardiography with supplemental posterior leads, echocardiography, and meticulous abdominal ultrasonography might improve the outcome in selected MICU patients. PMID- 10852417 TI - Biodistribution of radiolabeled adenosylcobalamin in patients diagnosed with various malignancies. AB - OBJECTIVE: To study the biodistribution of a vitamin B12 analog, indium In 111 labeled diethylenetriaminepentaacetate adenosylcobalamin (In 111 DAC), in patients recently diagnosed as having primary or recurrent malignancy. PATIENTS AND METHODS: Thirty patients (14 women and 16 men) with radiographically or clinically diagnosed breast, lung, colon, sarcomatous, thyroid, or central nervous system malignancies were studied prior to definitive surgery or biopsy. A maximum of 650 microCi (2.2 microg) of In 111 DAC was administered intravenously. Vitamin B12 and folate levels were determined prior to injection. Serum clearance and urinary and stool excretion of the tracer were measured. Images were routinely obtained at 0.5, 3 to 5, and 20 to 24 hours after injection. Biodistribution of In 111 DAC was determined by computer analysis of regions of interest. RESULTS: Serum T1/2 clearance was 7 minutes. Average urinary and stool excretion of the injected dose over 24 hours was 26.1% and 0.4%, respectively. The greatest focal uptake of In 111 DAC occurred in the liver and spleen, followed by the nasal cavity and salivary and lacrimal glands. The average tumor uptake of the injected dose was 2% at 30 minutes and 1.5% at 24 hours. High-grade primary and metastatic breast, lung, colon, thyroid, and sarcomatous malignancies were all imaged at 3 to 5 hours after injection. Central nervous system tumors and advanced metastatic prostate cancer were best identified at 24 hours. Mammographically occult, palpable, and nonpalpable breast cancers were delineated by In 111 DAC. Low-grade malignancies as well as early skeletal metastatic disease were not effectively imaged by the vitamin B12 tracer. Patients with elevated baseline vitamin B12 or those concurrently taking corticosteroids appeared to have optimal visualization of their malignancies. CONCLUSION: Vitamin B12 may be a useful vehicle for delivering diagnostic and therapeutic agents to various malignancies. Further evaluation of cobalamin analogs and their interaction with transport proteins and cellular receptors within malignant tissue and infection is warranted. PMID- 10852418 TI - Renal cell carcinoma metastatic to the pancreas: clinical and radiological features. AB - OBJECTIVE: To review the clinical features, computed tomographic (CT) appearance, and treatment outcomes in a case series of patients with renal cell carcinoma (RCC) metastatic to the pancreas. PATIENTS AND METHODS: We retrospectively reviewed the records of 23 patients (15 men and 8 women) with RCC metastatic to the pancreas, detected by CT examination between 1986 and 1996. All patients had undergone a previous nephrectomy for RCC. RESULTS: Isolated mild elevation in liver function test results (in 5 patients) or in serum amylase level (in 8 patients) was observed. New-onset diabetes was detected in 3 patients. The CT characteristics of the pancreatic metastases generally resembled those of primary RCC with well-defined margins and greater enhancement than normal pancreas with a central area of low attenuation. The mean interval between resection of the primary RCC and detection of the pancreatic metastases was 116 months (range, 1 295 months). In 18 patients (78%), the pancreatic metastases were diagnosed more than 5 years after nephrectomy. The pancreas was the initial metastatic site in 12 patients (52%). Survival was shortened with higher tumor grade (mean survival time of 41 months and 10 months in patients with grade 2 and 3, respectively). Surgical resection was carried out in 11 patients (7 distal and 3 total pancreatectomies and 1 distal pancreatectomy followed 4 years later by total pancreatectomy), with 8 patients alive at a mean follow-up of 4 years, 6 of whom remained free of recurrence. Overall, 12 patients (52%) were alive at a mean of 42 months after diagnosis of metastatic disease. CONCLUSIONS: The appearance of metastatic RCC lesions in the pancreas closely resembles the appearance of primary RCC on CT images. Pancreatic metastases from RCC are frequently detected many years after nephrectomy. Patient survival correlates with tumor grade. Histologic analysis of pancreatic masses in patients with a history of resected primary RCC is important since the prognosis for RCC metastatic to the pancreas is much better than that for primary pancreatic adenocarcinoma. PMID- 10852419 TI - Peripheral blood eosinophilia in association with sarcoidosis. AB - OBJECTIVES: To review retrospectively our experience with peripheral blood eosinophilia (PBE) in sarcoidosis and to analyze histologically lung biopsy specimens for the presence of lung tissue eosinophils. PATIENTS AND METHODS: We reviewed 140 cases of sarcoidosis diagnosed between May 1975 and January 1998. Ninety-five patients (66.3% women; 70.5% African American; mean age, 35.9 years) met the inclusion criteria. Transbronchial biopsy specimens from 82 patients were divided into 4 morphologic compartments: parenchyma, bronchial wall, parenchymal granulomas, and bronchial wall granulomas. Within compartments, up to 10 high power fields were scored semiquantitatively for eosinophils, from 0 (none) to 4+ (numerous). RESULTS: Thirty-nine patients (41%) had PBE. Four had PBE greater than 10%. The highest eosinophil count (21%) occurred in 1 patient. Sixty-five (79%) of 82 patients had no or few (1+) eosinophils in lung tissue; 17 patients had eosinophils scored as 2+ or higher. There was no correlation between peripheral blood eosinophil count and presence of eosinophils in transbronchial biopsy specimens. Eosinophils were least conspicuous in parenchyma but evenly distributed in bronchial wall and parenchymal and bronchial wall granulomas. CONCLUSIONS: Peripheral blood eosinophilia occurs frequently in sarcoidosis. However, there appears to be no association between peripheral blood eosinophil count and presence of lung tissue eosinophils. Whether eosinophils participate in the pathogenesis of sarcoidosis requires further study. PMID- 10852420 TI - High frequency of pulmonary lymphangioleiomyomatosis in women with tuberous sclerosis complex. AB - OBJECTIVE: To determine the frequency of pulmonary lymphangioleiomyomatosis (LAM), a rare cystic lung disorder that occurs almost exclusively in women of reproductive age, in women with tuberous sclerosis complex (TSC), an inheritable multiorgan hamartomatosis. PATIENTS AND METHODS: In this retrospective cohort study, the medical records of 78 women with definite TSC were reviewed, and pertinent information was recorded, including the demographic data, clinical manifestations, results of lung biopsies and autopsies, and findings on imaging studies of the chest and abdomen. All available computed tomographic (CT) scans of the chest and abdomen were reviewed. RESULTS: Of 78 women with definite TSC seen from 1977 to 1998, 20 (26%) had evidence of LAM. Surgical lung biopsy or autopsy in 7 patients confirmed the diagnosis of their lung disease. Characteristic CT findings of LAM were noted in 13 additional patients. Twelve of these 20 patients with TSC-associated LAM had respiratory symptoms, including exertional shortness of breath and spontaneous pneumothorax, that eventually led to their pulmonary diagnosis. CONCLUSIONS: The frequency of lung involvement (LAM) in women with TSC is substantially higher than previously suspected and may be even higher than reported in this retrospective study. These findings support the recommendation for a screening CT of the chest for all women with TSC. PMID- 10852421 TI - Prothrombin G20210A polymorphism and thrombophilia. AB - Recently, a single mutation in the 3'-untranslated region of the prothrombin gene was reported, resulting in a G-to-A substitution. This finding added to the growing list of genetic disorders thought to be responsible for familial thrombophilia. Although most studies generally agree about the increased risk of venous thrombosis in individuals carrying this mutation, its role in the first event of venous thromboembolism and in recurrent events is unclear. Even less clear is the role this mutation has in the formation of arterial thrombosis (including coronary artery disease and cerebral ischemia) due to contradicting results of studies. This mutation has important clinical implications since it is a common cause of genetic thrombophilia, second only to the factor V Leiden mutation. However, the mutation by itself may not be enough to trigger disease because thromboembolic disease is now generally accepted as a multifactorial disorder. Careful evaluation of this mutation will augment the clinician's ability to stratify systematically an individual's risk of developing spontaneous thrombosis. PMID- 10852422 TI - From Inuit to implementation: omega-3 fatty acids come of age. AB - During the past 25 years, the cardiovascular effects of marine omega-3 (omega-3) fatty acids have been the subject of increasing investigation. In the late 1970s, epidemiological studies revealed that Greenland Inuits had substantially reduced rates of acute myocardial infarction compared with Western control subjects. These observations generated more than 4,500 studies to explore this and other effects of omega-3 fatty acids on human metabolism and health. From epidemiology to cell culture and animal studies to randomized controlled trials, the cardioprotective effects of omega-3 fatty acids are becoming recognized. These fatty acids, when incorporated into the diet at levels of about 1 g/d, seem to be able to stabilize myocardial membranes electrically, resulting in reduced susceptibility to ventricular dysrhythmias, thereby reducing the risk of sudden death. The recent GISSI (Gruppo Italiano per lo Studio della Sopravvivenza nell'Infarto miocardico)-Prevention study of 11,324 patients showed a 45% decrease in risk of sudden cardiac death and a 20% reduction in all-cause mortality in the group taking 850 mg/d of omega-3 fatty acids. These fatty acids have potent anti-inflammatory effects and may also be antiatherogenic. Higher doses of omega-3 fatty acids can lower elevated serum triglyceride levels; 3 to 5 g/ d can reduce triglyceride levels by 30% to 50%, minimizing the risk of both coronary heart disease and acute pancreatitis. This review summarizes the emerging evidence of the use of omega-3 fatty acids in the prevention of coronary heart disease. PMID- 10852423 TI - Subtraction ictal SPECT coregistered to MRI for seizure focus localization in partial epilepsy. AB - Peri-ictal single-photon emission computed tomography (SPECT) of the brain is increasingly used in localizing the seizure focus in presurgical evaluation of patients with partial epilepsy. However, traditional side-by-side visual interpretation of ictal and interictal SPECT films is hampered by differences in slice location and tracer activity. Precise correlation of the seizure focus with a high-quality image of the underlying brain anatomy can improve the physician's understanding of seizure neurophysiology and assist in surgical planning. Computer-based methods have been developed for aligning, normalizing, and subtracting digital ictal and interictal SPECT images of the patient's brain to produce a map of the blood flow changes occurring between the seizure and resting states. These maps are then aligned with a high-resolution magnetic resonance image (MRI) of the patient's brain anatomy and fused to identify anatomical regions involved in the seizure. The purpose of this article is to review the technical components and clinical implementation of subtraction ictal SPECT, as well as to discuss recent technological advances that could extend and improve the diagnostic and localizing capacity of this method. PMID- 10852424 TI - Pulmonary hypertension: diagnostics and therapeutics. AB - Pulmonary hypertension (PH) may develop because of a spectrum of insults to the lungs; in some patients, there seems to be no cause. Noninvasive tests, such as standard chest radiography, electrocardiography, and transthoracic Doppler echocardiography, provide effective screening if PH is suspected. This synopsis focuses on these screening studies and the more common clinical problems, including primary cardiac abnormalities, obstructive sleep apnea, chronic pulmonary embolism, pulmonary parenchymal problems, connective tissue disorders, cirrhosis with portal hypertension, and use of appetite suppressants, that should be considered when PH exists. Treatment options for PH, including intravenous prostacyclin (epoprostenol), and investigational agents such as subcutaneous or oral prostacyclin analogues and oral endothelin receptor antagonists are described. PMID- 10852425 TI - Churg-Strauss syndrome complicated by eosinophilic endomyocarditis. AB - A 34-year-old woman with asthma had increasing dyspnea on exertion for 9 months and new-onset mononeuritis multiplex. An examination demonstrated sinus tachycardia, elevated jugular venous pressure, and a tender nonpulsatile liver. The leukocyte count was 15.8 x 10(9)/L, with 23% eosinophils. Echocardiography revealed a laminated thrombus obliterating much of the right ventricular cavity, with encasement of the tricuspid valve. Ultrafast computed tomography showed no evidence of pulmonary emboli. Biopsy specimens of skin nodules revealed extravascular palisading granulomas. The thrombus was refractory to corticosteroids, and right ventricular thrombectomy was performed. To our knowledge, this is the third reported case of Churg-Strauss syndrome with thrombotic complications from coexistent eosinophilic endomyocarditis. In an asthmatic patient with chronic dyspnea, eosinophilic tissue infiltration, and neuropathy, Churg-Strauss syndrome should be considered; evaluation for cardiac involvement may be warranted. PMID- 10852426 TI - Primary aldosteronism in a patient with familial adenomatous polyposis. AB - Patients with familial adenomatous polyposis (FAP) frequently have extracolonic manifestations of their disease. Prior reports have indicated an increased prevalence of adrenal lesions in patients with FAP. Although most of the adrenal lesions represent nonfunctioning adenomas, some patients have had hypercortisolism due to adrenocortical carcinoma or bilateral nodular hyperplasia. We present a case of a patient with FAP who had mineralocorticoid excess due to an aldosterone-producing adrenocortical adenoma. PMID- 10852427 TI - Ruptured pulmonary infarction: a rare, fatal complication of thromboembolic disease. AB - We describe 2 men, ages 69 and 49 years, who experienced fatal rupture of pulmonary infarcts. Both patients had documented prior thromboembolic events and subsequently had abrupt deterioration in cardiorespiratory function. Autopsies showed massive unilateral hemothorax in both patients. Rupture of a pulmonary infarct may occur spontaneously or iatrogenically due to aggressive anticoagulation. This may be difficult to distinguish from secondary hemothorax with an intact pleura, but rupture typically has a considerably more rapid clinical evolution. Treatment should include immediate withdrawal of thrombolytic or anticoagulant medications and evacuation of the pleural space. Surgical intervention can be considered, although the utility of that approach must await prospective trials. PMID- 10852428 TI - Recurrent panniculitis in a man with asthma receiving treatment with leukotriene modifying agents. AB - Leukotriene-modifying drugs are novel agents introduced recently to treat asthma. Both 5-lipoxygenase inhibitors, such as zileuton, and leukotriene receptor antagonists, such as zafirlukast and montelukast, have proved effective in the treatment of asthma. To our knowledge, there have been no detailed reports regarding dermatologic manifestations of this class of drugs. This article describes an unusual case of erythema nodosum in a 46-year-old asthmatic man who received 2 different leukotriene modifiers. PMID- 10852429 TI - Fournier gangrene associated with Crohn disease. AB - A 17-year-old boy presented with Fournier gangrene associated with previously undiagnosed Crohn ileocolitis. Fournier gangrene was managed by debridement, broad-spectrum antibiotics, and hyperbaric oxygen. A diverting ileostomy was performed before skin grafting and scrotal reconstruction. Microscopy of a full layer surgical sample from the terminal ileum revealed granulomas with multinucleated histiocytes, consistent with Crohn disease. Crohn disease was treated with mesalamine, metronidazole, 6-mercaptopurine, and infliximab. The patient was discharged on hospital day 32. At 6-month follow-up, reconstruction of his scrotum had completely healed. Ostomy output was normal. PMID- 10852430 TI - 35-year-old man with fever, hemoptysis, and lymphadenopathy. PMID- 10852431 TI - Difficult-to-control hypertension. PMID- 10852432 TI - Staphylococcus lugdunensis endocarditis after angiography. PMID- 10852433 TI - Adding preventive medicine training to medical school curricula. PMID- 10852434 TI - Duplex ultrasonography and carotid artery dissection. PMID- 10852435 TI - Hyperlipidemia: diagnostic and therapeutic perspectives. PMID- 10852436 TI - Insulin resistance is not necessarily an essential component of type 2 diabetes. PMID- 10852437 TI - Hashimoto's thyroiditis with heterogeneous antithyrotropin receptor antibodies: unique epitopes may contribute to the regulation of thyroid function by the antibodies. AB - Blocking-type TSH-binding inhibitor Igs (TBIIs) are known to cause hypothyroidism and an atrophic thyroid gland in patients with primary myxedema. They can block the activity of thyroid-stimulating antibodies (TSAbs) in Graves' patients as well as the activity of TSH. The majority of the epitopes for these blocking-type TBIIs have been, and are shown herein, to be present on the C-terminal region of the extracellular domain of the human TSH receptor (TSHR), whereas those for Graves' TSAbs are on the N-terminus. We report on a patient with Hashimoto's thyroiditis who suffered from mild hypothyroidism and a moderately sized goiter. Her serum had a potent blocking-type TBII and a weak TSAb in human and porcine TSHR systems. Using human TSHR/lutropin-CG receptor chimeras, we determined that the functional epitope of her blocking-type TBII was uniquely present on the N terminal, rather than the C-terminal, region of the extracellular domain of the TSHR, unlike the case for blocking-type TBIIs in primary myxedema patients. The epitope of her TSAb was also unusual. Although the functional epitopes of most TSAbs are known to involve the N-terminal region of the receptor, her TSAb epitope did not seem to be present solely on the N- or C-terminus of the extracellular domain of the receptor. Blocking-type TBIIs from patients with primary myxedema blocked her TSAb activity as well as stimulation by TSH; her blocking-type TBII was able to only partially block her TSAb. In contrast, her blocking-type TBII almost completely blocked TSAbs from Graves' patients. Thus, we suggest that the unique epitopes of this patient's heterogeneous population of TSH receptor antibodies, at least in part, contribute to regulation of her thyroid function. PMID- 10852438 TI - Osteoporosis: an unusual presentation of childhood Crohn's disease. AB - Osteoporosis is known to be associated with Crohn's disease. We report a 12-yr old boy without a history of steroid use, in whom severe osteoporosis and multiple collapsed vertebrae were the presenting manifestations of Crohn's disease. After treatment of the Crohn's disease, he resumed normal growth and progressed through puberty. Concomitantly, he demonstrated a substantial recovery of vertebral bone mineral density and structure. Possible pathophysiological mechanisms underlying the osteoporosis and the subsequent improvement in bone density are discussed. PMID- 10852439 TI - Sequential parathyroid hormone/alendronate therapy for osteoporosis--robbing Peter to pay Paul? PMID- 10852440 TI - Enhancement of bone mass in osteoporotic women with parathyroid hormone followed by alendronate. AB - Treatment of osteoporosis with PTH causes a marked increase in vertebral bone mineral density (BMD). However, this effect is rapidly reversed when the treatment is stopped. The purpose of the present study was to determine whether the bisphosphonate alendronate could preserve or enhance bone density in patients previously treated with PTH. Sixty-six postmenopausal osteoporotic women were treated for 1 yr with 50, 75, or 100 microg recombinant human PTH-(1-84) or placebo, and then were given 10 mg alendronate daily for an additional year. BMD was measured in the femoral neck, lumbar spine, and whole body. Markers of bone turnover included skeletal alkaline phosphatase, osteocalcin, and N-telopeptide. During the first year, changes in BMD (mean +/- SD) in women receiving PTH (all doses combined) were 7.1 +/- 5.6% (spine), 0.3 +/- 6.2% (femoral neck), and -2.3 +/- 3.3% (total body). After switching to alendronate for 1 yr in women who previously had received PTH, mean changes in BMD were 13.4 +/- 6.4% (spine), 4.4 +/- 7.2% (femoral neck), and 2.6 +/- 3.1% (whole body). In the subgroup of patients who had received the highest dose of PTH, the mean increase in vertebral BMD was 14.6 +/- 7.9%. All markers of bone turnover increased during treatment with PTH and decreased to below baseline after 1 yr of alendronate. In conclusion, sequential treatment of osteoporosis with PTH and alendronate results in an increase in vertebral bone density that is considerably more than has been reported with alendronate or estrogens alone. This combination of drugs may be a useful approach to maximizing bone density in women with vertebral osteoporosis. PMID- 10852441 TI - Age-related decreases in melatonin secretion--clinical consequences. PMID- 10852442 TI - Nocturnal melatonin patterns in children. AB - Time patterns in nocturnal concentrations of circulating melatonin of children are quantified in 8 girls and 8 boys, 8.7-16.8 yr of age, classified by Tanner pubertal stage. Between 1900 and 0700 h, each provided blood samples at 30-min intervals for melatonin RIA. Associations with gender, body mass index, and chronological and pubertal age determined by multiple linear regression and ANOVA reveal that the area under the curve of 12-h melatonin concentrations was affected by pubertal rather than chronological age, an effect to which data collected during darkness contributed the most. Each data series was also analyzed by a least squares spectrum at frequencies of 1-20 cycles/day. Ultradian changes with periods of 3.4 and 1.5 h, putatively associated with rapid eye movement sleep cycles, characterize nocturnal melatonin in boys and girls. PMID- 10852443 TI - Are gender differences in the responses to hypoglycemia relevant to iatrogenic hypoglycemia in type 1 diabetes? PMID- 10852444 TI - Gender-related differences in counterregulatory responses to antecedent hypoglycemia in normal humans. AB - Compared to men, inherent counterregulatory responses are reduced in healthy and type 1 diabetic women. Despite this, the prevalence of hypoglycemia in patients with type 1 diabetes (type 1 DM) is gender neutral. The aim of this study was to determine the in vivo mechanism(s) responsible for this apparent clinical paradox. The central importance of antecedent hypoglycemia in causing subsequent counterregulatory failure is now established. We, therefore, hypothesized that a gender-related difference to the blunting effects of prior hypoglycemia may exist, and this could explain why type 1 DM women do not have an increased prevalence of hypoglycemia despite reduced counterregulatory responses. Fifteen healthy male and female individuals (eight men and seven women) underwent four separate 2-day experimental protocols in a randomized fashion. Day 1 involved identical morning and afternoon 2-h hyperinsulinemic (9 pmol/kg x min) glucose clamp studies with 5.1 +/- 0.1, 3.9 +/- 0.1, 3.3 +/- 0.1, or 2.9 +/- 0.1 mmol/L. Day 2 consisted of a single 2-h hypoglycemic clamp of 2.9 +/- 0.1 mmol/L. Insulin levels were similar on both days of each protocol in men and women. After day 1 euglycemia (5.1 +/- 0.1 mmol/L), day 2 counterregulatory responses were significantly increased (P < 0.01) in men relative to women. In women, counterregulatory responses were resistant to the effects of day 1 hypoglycemia. Antecedent hypoglycemia of 3.9, 3.3, and 2.9 +/- 0.1 mmol/L produced 3 +/- 2%, 5 +/- 2%, and 25 +/- 4% aggregate reductions in day 2 neuroendocrine, muscle sympathetic nerve activity, and metabolic counterregulatory responses. In marked contrast, identical day 1 hypoglycemia of 3.9, 3.3, and 2.9 +/- 0.1 mmol/L in men produced significantly greater reductions in day 2 counterregulatory responses of 30 +/- 6%, 39 +/- 6%, and 52 +/- 6%, respectively. The net effect of the differential gender effects of antecedent hypoglycemia was to overcome the usually increased (50%) sympathetic nervous system (SNS) counterregulatory responses to hypoglycemia found in men. We conclude that 1) antecedent hypoglycemia produces less blunting of counterregulatory responses to subsequent hypoglycemia in women relative to men; 2) two episodes of antecedent hypoglycemia can overcome the greater SNS response to hypoglycemia usually found in men; and 3) the reduced susceptibility of women to the blunting effects of antecedent hypoglycemia may be the mechanism explaining why, despite inherently reduced SNS counterregulatory responses, female type 1 DM patients have a similar prevalence of hypoglycemia compared to men. PMID- 10852445 TI - Sex and the single gene--FH-1. PMID- 10852446 TI - Severity of hypertension in familial hyperaldosteronism type I: relationship to gender and degree of biochemical disturbance. AB - In familial hyperaldosteronism type I (FH-I), inheritance of a hybrid 11beta hydroxylase/aldosterone synthase gene causes ACTH-regulated aldosterone overproduction. In an attempt to understand the marked variability in hypertension severity in FH-I, we compared clinical and biochemical characteristics of 9 affected individuals with mild hypertension (normotensive or onset of hypertension after 15 yr, blood pressure never >160/100 mm Hg, < or = 1 medication required to control hypertension, no history of stroke, age >18 yr when studied) with those of 17 subjects with severe hypertension (onset before 15 yr, or systolic blood pressure >180 mm Hg or diastolic blood pressure >120 mm Hg at least once, or > or = 2 medications, or history of stroke). Severe hypertension was more frequent in males (11 of 13 males vs. 6 of 13 females; P < 0.05). All 4 subjects still normotensive after age 18 yr were females. Of 10 other affected, deceased individuals (7 males and 3 females) from a single family, all six who died before 60 yr of age (4 by stroke) were males. Biochemical studies were conducted in 6 mild and 16 severe subjects. The 2 groups were similar in terms of urinary sodium excretion. Mild subjects tended, although not significantly, to have lower urinary 18-oxo-cortisol (mean +/- SD, 27.4 +/- 9.0 vs. 35.2 +/- 12.9 nmol/mmol creatinine x day), higher plasma potassium (4.0 +/- 0.3 vs. 3.6 +/- 0.4 mmol/L), and lower recumbent (0800 h after overnight recumbency) plasma aldosterone levels (498 +/- 279 vs. 744 +/- 290 pmol/L). Upright (midmorning after 2-3 h of upright posture) plasma aldosterone levels were similar (mild, 485 +/- 150; severe, 474 +/- 188 pmol/L). In 1 normotensive female, upright PRA was much higher, and the upright aldosterone/PRA ratio was much lower than that in the other subjects. The remaining mild subjects had similar upright PRA levels (mild, 2.8 +/- 1.4; severe, 3.7 +/- 3.2 pmol/ L x min) and aldosterone/PRA ratios (mild, 199.5 +/- 133.4; severe, 200.6 +/- 150.9) as severe subjects. During angiotensin II (AII) infusion studies (n = 6 mild and 10 severe), performed during recumbency, aldosterone levels were lower in the mild group both basally (404 +/- 144 vs. 843 +/- 498 pmol/L; P < 0.05) and after 60 min AII (2 ng/kg x min; 261 +/- 130 vs. 520 +/- 330 pmol/L; P < 0.05). Aldosterone was unresponsive (rose by <50%) to AII in all subjects. Day curve studies (blood collected every 2 h for 24 h; n = 2 mild and 7 severe) demonstrated abnormal regulation of aldosterone by ACTH rather than by AII in both groups. In conclusion, in this series of patients with FH-I, males had more severe hypertension, and the degree of hybrid gene-induced aldosterone overproduction may have contributed to the severity of hypertension. PMID- 10852447 TI - Exercise and fatigue--is neuroendocrinology an important factor? PMID- 10852448 TI - Effects of caffeine on muscle glycogen utilization and the neuroendocrine axis during exercise. AB - To examine the effect of caffeine ingestion on muscle glycogen utilization and the neuroendocrine axis during exercise, we studied 20 muscle glycogen-loaded subjects who were given placebo or caffeine (6 mg/kg) in a double blinded fashion 90 min before cycling for 2 h at 65% of their maximal oxygen consumption. Exercise-induced glycogen depletion in the thigh muscle was noninvasively measured by means of 13C nuclear magnetic resonance spectroscopy (NMR) spectroscopy, and plasma concentrations of substrates and neuroendocrine hormones, including beta-endorphins, were also assessed. Muscle glycogen content was increased 140% above normal values on the caffeine trial day (P < 0.001). After cycling for 2 h, caffeine ingestion was associated with a greater increase in plasma lactate (caffeine: +1.0 +/- 0.2 mmol/L; placebo, +0.1 +/- 0.2 mmol/L; P < 0.005), epinephrine (caffeine, +223 +/- 82 pg/mL; placebo, +56 +/- 26 pg/mL; P < 0.05), and cortisol (caffeine, +12 +/- 3 mg/mL; placebo, +2 +/- 2 mg/mL; P < 0.001) levels. However, plasma free fatty acid concentrations increased (caffeine, +814 +/- 133 mmol/L; placebo, +785 +/- 85 mmol/L; P = NS), and muscle glycogen content decreased (caffeine, -57 +/- 6 mmol/L muscle; placebo, -53 +/- 5 mmol/L muscle; P = NS) to the same extent in both groups. At the same time, plasma beta-endorphin levels almost doubled (from 30 +/- 5 to 53 +/- 13 pg/mL; P < 0.05) in the caffeine-treated group, whereas no change occurred in the placebo group. We conclude that caffeine ingestion 90 min before prolonged exercise does not exert a muscle glycogen-sparing effect in athletes with high muscle glycogen content. However, these data suggest that caffeine lowers the threshold for exercise-induced beta-endorphin and cortisol release, which may contribute to the reported benefits of caffeine on exercise endurance. PMID- 10852449 TI - The I27L amino acid polymorphism of hepatic nuclear factor-1alpha is associated with insulin resistance. AB - Mutations of the hepatic nuclear factor-1alpha (HNF-1alpha) gene have been found in patients with maturity-onset diabetes of the young. We examined the relation between the I27L polymorphism of HNF-1alpha and insulin sensitivity and beta-cell function assessed by a hyperglycemic clamp. This study included 52 healthy glucose-tolerant and normotensive subjects (age, 19-40 yr; body mass index, 17.58 35.61 kg/m2; waist/hip ratio, 0.65-1.03). We identified 19 LL subjects, 24 IL, and 9 II subjects. No difference was noted in the demographic features among the three genotypes. The LL group had the highest postchallenge insulin levels at 30 and 90 min (P = 0.038 and P = 0.015, respectively) and also the highest insulin area under curve (P = 0.009) among the three genotypes. The LL group was more insulin resistant than the IL and II groups (P = 0.042 for insulin sensitivity index). After adjusting for age, gender, obesity, and ethnicity, the I27L polymorphism was an independent determinant of the insulin sensitivity index (P = 0.001). However, it had no impact on either the first or second phase insulin response. Therefore, we conclude that the I27L polymorphism is associated with insulin resistance, but not beta-cell function. The mechanism of this association is unclear, but HNF-1alpha may play a role in regulating hepatic glucose metabolism. PMID- 10852450 TI - Stimulatory effects of stress on gonadotropin secretion in estrogen-treated women. AB - Although stress is known to inhibit the hypothalamic-pituitary-gonadal axis, recent studies in the monkey show that, under certain conditions, in the presence of estrogen, stress may actually stimulate LH release. We investigated the effects of a mild inflammatory stress (2.0-3.0 ng/kg endotoxin) on LH release in five postmenopausal women with and without transdermal estradiol (E2, 0.1 mg) replacement. In another five E2-treated women, LH release was studied when the adrenal was stimulated directly by a 3-h ACTH infusion (Cortrosyn, 50 microg/h). Mean E2 levels were less than 12 pg/mL in the unreplaced subjects and were 86 +/- 10 pg/mL and 102 +/- 18 pg/mL in the two groups of E2-replaced subjects. Blood was sampled every 15-20 min for 2 h before and for 7 h after endotoxin or ACTH injection. Mean cortisol and progesterone levels increased in all three groups over time (P < 0.001). In the women without E2 replacement, basal LH was 26.8 +/- 5.3 mIU/mL and did not change significantly, over time, after endotoxin (P = 0.58). In the same women on E2, however, a significant increase in LH occurred after endotoxin (P = 0.02), from a mean hourly baseline of 15.3 +/- 5.4 mIU/mL to a peak of 50.0 +/- 25.2 mIU/mL. During the ACTH infusion, there was a significant stimulation of LH release in the E2-replaced subjects (P < 0.001), from a mean hourly baseline of 13.3 +/- 3.0 mIU/mL to a peak of 44.1 +/- 11.7 mIU/mL. In both groups, this increase occurred 2-4 h after the initial rise in progesterone and persisted to the end. We conclude that, in the presence of sufficient estrogen, activation of the hypothalamic-pituitary-adrenal axis leads to a stimulation of LH release. This is likely related to a rise in adrenal progesterone and its known stimulatory effect on LH release in the presence of E2. These studies provide a potential mechanism in the human by which an acute stress during the follicular phase of the menstrual cycle might lead to a premature LH surge and thereby interfere with follicular maturation and ovulation. PMID- 10852451 TI - Absence of detectable melatonin and preservation of cortisol and thyrotropin rhythms in tetraplegia. AB - The human circadian timing system regulates the temporal organization of several endocrine functions, including the production of melatonin (via a neural pathway that includes the spinal cord), TSH, and cortisol. In traumatic spinal cord injury, afferent and efferent circuits that influence the basal production of these hormones may be disrupted. We studied five subjects with chronic spinal cord injury (three tetraplegic and two paraplegic, all neurologically complete injuries) under stringent conditions in which the underlying circadian rhythmicity of these hormones could be examined. Melatonin production was absent in the three tetraplegic subjects with injury to their lower cervical spinal cord and was of normal amplitude and timing in the two paraplegic subjects with injury to their upper thoracic spinal cord. The amplitude and the timing of TSH and cortisol rhythms were robust in the paraplegics and in the tetraplegics. Our results indicate that neurologically complete cervical spinal injury results in the complete loss of pineal melatonin production and that neither the loss of melatonin nor the loss of spinal afferent information disrupts the rhythmicity of cortisol or TSH secretion. PMID- 10852452 TI - A comparison of the effects of raloxifene and estrogen on bone in postmenopausal women. AB - Raloxifene HCl, a selective estrogen receptor modulator, has been shown to increase bone mineral density (BMD) and decrease biochemical markers of bone turnover in postmenopausal women without stimulatory effects on the breast and uterus. However, it is not known whether the changes in BMD and bone turnover are associated with changes at the tissue level, nor how changes with raloxifene compare with estrogen. In this randomized, double blind study, we evaluated the effects of raloxifene (Evista, 60 mg/day) or conjugated equine estrogens (CEE; Premarin, 0.625 mg/day) on bone architecture, bone turnover, and BMD. Iliac crest bone biopsies were obtained at baseline and at the end of the study after double tetracycline labeling and were analyzed for standard histomorphometric indexes. Serum and urinary biochemical markers of bone turnover were measured at baseline and at 4, 10, 18, and 24 weeks of treatment. Total body, lumbar spine, and hip BMD were measured at baseline and at the end of the study by dual energy x-ray absorptiometry. Activation frequency and bone formation rate/bone volume were significantly decreased from baseline in the CEE, but not in the raloxifene, group. Bone mineralization did not change in either group. Most markers of bone resorption and formation decreased in both groups, but to a greater degree in the CEE group (P < .05). Total body and lumbar spine BMD increased from baseline in both groups, with a greater increase in the CEE group (P < 0.05). Hip BMD significantly increased from baseline in the raloxifene group, but the change was not different from that in the CEE group. These results suggest that raloxifene reduces bone turnover and increases bone density, although to a lesser extent than CEE. Thus, raloxifene is an alternative to CEE for the prevention and treatment of osteoporosis in postmenopausal women. PMID- 10852453 TI - Impact of age on cortisol secretory dynamics basally and as driven by nutrient withdrawal stress. AB - The present study tests the clinical hypothesis that aging impairs homeostatic adaptations of cortisol secretion to stress. To this end, we implemented a short term 3.5-day fast as an ethically acceptable metabolic stressor in eight young (ages 18-35 yr) and eight older (ages 60-72 yr) healthy men. Volunteers were studied in randomly ordered fed vs. fasting sessions. To capture the more complex dynamics of cortisol's feedback control, blood was sampled every 10 min for 24 h for later RIA of serum cortisol concentrations and quantitation of the pulsatile, entropic, and 24-h rhythmic modes of cortisol release using deconvolution analysis, the approximate entropy statistic, and cosine regression, respectively. The stress of fasting elevated the mean (24-h) serum cortisol concentration equivalently in the two age cohorts [i.e. from 7.2 +/- 0.35 to 11.6 +/- 0.71 microg/dL in young men and from 7.7 +/- 0.39 to 12.6 +/- 0.59 microg/dL in older individuals (P < 10(-7))]. The rise in integrated cortisol output was driven mechanistically by selective augmentation of cortisol secretory burst mass (P = 0.002). The resultant daily (pulsatile) cortisol secretion rate increased significantly but equally in young (from 94 +/- 6.3 to 151 +/- 15 microg/dL x day) and older (from 85 +/- 5.4 to 145 +/- 7.3 microg/dL x day) volunteers (P < 10(-4)). Nutrient restriction also prompted a marked reduction in the quantifiable regularity of (univariate) cortisol release patterns in both cohorts (P < 10(-4)). However, older men showed loss of joint synchrony of cortisol and LH secretion even in the fed state, which failed to change with metabolic stress (P < 10(-6)). In addition, older individuals maintained a premature (early-day) cortisol elevation in the fed state and unexpectedly evolved an anomalous further cortisol phase advance of 99 +/- 16 min during fasting (P < 10(-5)). Caloric deprivation in aging men also disproportionately elevated the mesor of 24-h rhythmic cortisol release (P = 10(-7)) and elicited a greater increment in the mean day-night variation in cortisol secretory-burst mass (P < 0.01 vs. young controls). Lastly, short-term caloric depletion in older subjects paradoxically normalized their age-associated suppression of the 24-h rhythm in cortisol interburst intervals. In summary, acute metabolic stress in healthy aging men (compared with young individuals) unmasks distinct, albeit complex, disruption of cortisol homeostasis. These dynamic anomalies impact the feedback-dependent and time-sensitive coupling of pulsatile and 24-h rhythmic cortisol secretion. Nutrient-withdrawal stress in the older male heightens the cortisol phase disparity already evident in fed elderly individuals. Conversely, the stress of fasting in young men paradoxically reproduces selected features of the aging unstressed (fed) cortisol axis; viz., abrogation of joint cortisol-LH synchrony and suppression of the normal diurnal variation in cortisol burst frequency. Whether fasting would unveil analogous disruption of feedback-dependent control of the corticotropic axis in healthy aging women is not yet known. PMID- 10852454 TI - Endocrine consequences of long-term intrathecal administration of opioids. AB - Intrathecal administration of opioids is a very efficient tool in the long-term control of intractable nonmalignant pain. However, despite the well known role of opioids in endocrine regulation, few data are available about possible effects on hypothalamic-pituitary function during this treatment. Seventy-three patients (29 men and 44 women; mean age, 49.2 +/- 11.7 yr) receiving opioids intrathecally for nonmalignant pain were enrolled for extensive endocrine investigation. At the time of hormonal determination, the mean duration of opioid treatment was 26.6 +/ 16.3 months; the mean daily dose of morphine was 4.8 +/- 3.2 mg. The control group consisted of 20 patients (11 men and 9 women; mean age, 54.2 +/- 14.0 yr) with a comparable pain syndrome but not treated with opioids. Decreased libido or impotency was present in 23 of 24 men receiving opioids. The serum testosterone level was below 9 nmol/L in 25 of 29 men and was significantly lower than that in the control group (P < 0.001). The free androgen index was below normal in 18 of 29 men and was significantly lower than that in the control group (P < 0.001). The serum LH level was less than 2 U/L in 20 of 29 men and was significantly lower than that in the control group (P < 0.001). Serum FSH was comparable in both groups. Decreased libido was present in 22 of 32 women receiving opioids. All 21 premenopausal females developed either amenorrhea or an irregular menstrual cycle, with ovulation in only 1. Serum LH, estradiol, and progesterone levels were lower in the opioid group. In all 18 postmenopausal females significantly decreased serum LH (P < 0.001) and FSH (P = 0.012) levels were found. The 24-h urinary free cortisol excretion was below 20 microg/day in 14 of 71 opioid patients and was significantly lower than that in the control group (P = 0.003). The peak cortisol response to insulin-induced hypoglycemia was below 180 microg/L in 9 of 61 opioid patients and was significantly lower than that in the nonopioid group (P = 0.002). The insulin-like growth factor I SD score was below -2 SD in 12 of 73 opioid patients and was significantly lower than that in the control group (P = 0.002). The peak GH response to hypoglycemia was below 3 microg/L in 9 of 62 subjects and was significantly lower than that in the control group (P = 0.010). Thyroid function tests and PRL levels were considered normal. No metabolic disturbances were recorded, apart from significantly decreased high density lipoprotein cholesterol levels (P = 0.041) and elevated total/high density lipoprotein cholesterol ratio (P = 0.008) in the opioid group compared to the control group. Supplementation with gonadal steroids improved sexual function in most patients. In conclusion, of all patients receiving intrathecal opioids, the large majority of men and all women developed hypogonadotropic hypogonadism, about 15% developed central hypocorticism, and about 15% developed GH deficiency. These findings suggest that further investigations are required to determine the need for systematic endocrine work-up in these patients and the necessity for substitutive therapy. PMID- 10852455 TI - Fuel homeostasis during physical inactivity induced by bed rest. AB - The consequences of physical inactivity on fuel homeostasis were evaluated during 7 days of head-down bed rest (HDBR), a model mimicking weightlessness. Eight men (32.4 +/- 1.9 yr; body mass index, 23.9 +/- 0.7 kg/m2) and eight women (27.9 +/- 0.9 yr; body mass index, 20.9 +/- 0.6 kg/m2) underwent an oral glucose tolerance test (OGTT; 1 g/kg) before and after HDBR. The glucose load was labeled with 13C and associated with D-[6,6-2H2] glucose infusion, indirect calorimetry, breath tests, and plasma measurements to determine the glucose turnover and biodisponibility, substrate oxidation, and endocrine responses. Body composition was assessed using H2(18)O dilution. In addition, hormones were measured in daily blood and 24-h urine samples. No change in body composition was noted. Daily fasting insulin increased during HDBR (men, 34%; women, 26%), as did the insulin to glucose ratio (men, 30%; women, 25%). The normetanephrine level dropped (men, 30%; women, 16%), but metanephrine was unchanged. During OGTTs, the insulin response was increased after HDBR (men, 47%; women, 67%), whereas plasma glucose levels were similar. Nonesterified fatty acids and beta-hydroxybutyrate levels were lower. Endogenous glucose production dropped (28%), and exogenous glucose oxidation increased (28%) only in men. Resting energy expenditure was unchanged, but nonproteic respiratory quotient increased (men, 10%; women, 14%). Basal levels of lipid oxidation dropped in both sexes (approximately 90%), but those of carbohydrate oxidation increased in men (40%); as did lipogenesis in women (570%). In response to OGTTs, lipid oxidation was 80% reduced in both sexes after HDBR, but carbohydrate oxidation increased (25%) in men. Lipogenesis occurred in men (304%) and women (74%), but the latter had higher absolute levels. Therefore, 7 days of HDBR resulted in 1) reduced sympathetic activity, 2) insulin resistance suggested at the muscle level in men and at both the muscle and liver levels in women, 3) no changes in glucose biodisponibility, suggesting no alterations in the gastrointestinal function, and 4) a shift toward carbohydrate oxidation in men and a net lipogenesis in women. Such results suggest gender differences in response to sedentary life style and warrant further analysis. PMID- 10852456 TI - Low-dose monitored mitotane treatment achieves the therapeutic range with manageable side effects in patients with adrenocortical cancer. AB - Eight patients with adrenocortical cancer were treated with low doses of mitotane (2-3 g daily) while monitoring drug plasma levels. When the mitotane concentrations reached the therapeutic range (defined as mitotane plasma levels between 14-20 microg/mL), a dose reduction was performed to avoid toxicity. Thereafter, the mitotane dose was tailored according to plasma levels. A progressive increase in plasma mitotane concentrations was observed during treatment, and a highly significant linear correlation was found between plasma drug levels and the total mitotane dose. The therapeutic threshold was reached in all patients after 3-5 months and a total mitotane dose of 283-387 g/days (median, 363). The duration of treatment was 8-40 months (median, 9). Toxicity was manageable in all but one patient, who discontinued treatment. It is therefore possible to design a standard low dose schedule, e.g. 3 g/daily for about 3-4 months with following dose adjustments guided by the monitoring of plasma mitotane levels. This approach is able to provide therapeutic mitotane concentrations and limit the unwanted effects. The present data provide a rationale to change the approach to mitotane treatment in patients with adrenocortical carcinoma from high dose to low dose regimens. PMID- 10852457 TI - Pseudohypoparathyroidism 1b: exclusion of parathyroid hormone and its receptors as candidate disease genes. AB - Pseudohypoparathyroidism 1b (PHP 1b) is characterized by specific resistance of target tissues to PTH, but no mutations in the PTH/PTH-related peptide (PTHrP) receptor gene have been identified. To investigate the basis for defective PTH signaling, we used polymorphic markers in or near the genes encoding PTH and its receptors to perform linkage analysis between these loci and PHP 1b. Two multiplex PHP 1b families (families M and K) were informative for an intragenic polymorphism in exon 13 of the PTH/PTHrP receptor gene detected by PCR amplification and resolved by denaturing gradient gel electrophoresis. Linkage analysis revealed discordance of the PTH/PTHrP receptor with PHP1b. One PHP 1b kindred (family M) was informative for a intragenic polymorphism in exon 3 of the PTH gene detected by PCR amplification and resolved by denaturing gradient gel electrophoresis. The PTH gene polymorphism segregation was discordant with PHP 1b. Probands from each family had normal PTH genes by direct sequence analysis. In three PHP 1b kindreds, we analyzed simple sequence polymorphisms in three microsatellite markers flanking the PTH type 2 receptor locus located at 2q33. Linkage analysis demonstrated no linkage. In conclusion, neither the PTH gene nor the PTH receptor genes (type 1 and 2) are linked to PHP 1b. PMID- 10852458 TI - Macroprolactinoma shrinkage during cabergoline treatment is greater in naive patients than in patients pretreated with other dopamine agonists: a prospective study in 110 patients. AB - To investigate whether previous treatment with bromocriptine (BRC) or quinagolide (CV) impairs a subsequent response to long-term cabergoline (CAB) treatment, we prospectively studied 110 patients with macroprolactinoma. Four groups of patients were considered: 1) naive: 26 untreated patients with a mean serum PRL levels of 1013.4 +/- 277.7 microg/L (+/- SEM; range, 185.5-5611 microg/L); 2) intolerant: 19 patients previously shown to be intolerant of BRC treatment with a mean serum PRL level of 539.4 +/- 172.2 microg/L (range, 174-3564 microg/L); 3) resistant: 37 patients shown to be resistant/hyporesponsive to BRC, CV, or both, with a mean serum PRL level of 602.6 +/- 136.8 microg/L (range, 148-3511 microg/L); and 4) responsive: 28 patients previously treated with BRC or CV for 1 5 yr, achieving normoprolactinemia and restoration of gonadal function, but no longer treated with BRC or CV because of poor compliance or because the drug was not available. After a 15- to 30-day washout period, the serum PRL level was 397 +/- 43.1 microg/L (140-978 microg/L). CAB treatment was given at doses ranging 0.25-3.5 mg weekly for 1 yr to 110 patients, for 2 yr to 104 patients, and for 3 yr to 81 patients. Magnetic resonance imaging was performed before and after 12, 24, and 36 months of CAB treatment to evaluate significant tumor shrinkage (>80% reduction of pretreatment tumor volume). Among the 26 naive patients, normoprolactinemia was achieved in 21 (80.8%) after 1-6 months at 0.25-2 mg/week and in 5 patients after 24 months at 0.5-3 mg/week. Tumor volume was reduced from 1431.5 +/- 310.3 to 47.2 +/- 21.5 mm3 (P < 0.0001); average tumor shrinkage was 92.1 +/- 2.9%; significant tumor shrinkage was observed in 92.3% of patients, and tumor mass completely disappeared in 16 patients (61.5%). Among the 19 intolerant patients, normoprolactinemia was achieved in 18 (94.7%) after 1-6 months of CAB treatment at 0.25-1 mg/week. One patient remained mildly hyperprolactinemic. Tumor volume was reduced from 1925 +/- 423.1 to 842.0 +/- 330.7 mm3 (P < 0.001); average tumor shrinkage was 66.2 +/- 6.4%; significant tumor shrinkage was obtained in 42.1% of patients, and tumor mass completely disappeared in 4 patients (21%). Among the 37 resistant patients, normoprolactinemia was achieved in 19 (51.3%) after 6-12 months at 1-2 mg/week and in the remaining 18 patients after 18-24 months at 3-3.5 mg/week. Tumor volume was reduced from 1208.0 +/- 173.7 to 471.2 +/- 87.3 mm3 (P < 0.005); average tumor shrinkage was 58.4 +/- 4.9%; significant tumor shrinkage was obtained in 10 of 33 patients (30.3%), and in no patient did tumor mass completely disappear. Among the 28 responsive patients, normoprolactinemia was achieved in 23 (82.1%) after 1-6 months at 1-2 mg/week and in 5 patients after 12 months at 3 mg/week. Tumor volume was reduced from 1351.3 +/- 181.5 to 757.1 +/- 193.6 mm3 (P < 0.01); average tumor shrinkage was 59.2 +/- 6.2%; significant tumor shrinkage was obtained in 10 of 26 patients (38.4%), and tumor mass completely disappeared in 4 patients (15.4%). Nadir PRL levels and percent tumor shrinkage during CAB treatment in naive patients were significantly lower (P < 0.001) and higher (P < 0.001), respectively, than those in the remaining three groups, and the average weekly dose of CAB in resistant patients was significantly higher (P < 0.001) than that in the remaining three groups. A significant association was found between tumor shrinkage and previous treatments (chi2 = 27.1; P < 0.0001). At the multistep correlation analysis, nadir PRL levels were the strongest predictors of tumor shrinkage (r2 = 0.556; P < 0.0001), followed by CAB dose (r2 = 0.577; P < 0.0001). The tolerability was excellent in 105 patients (95.4%). In conclusion, the prevalence of macroprolactinoma shrinkage after CAB treatment at standard doses for 1-3 yr was higher in naive patients (92.3%) than in intolerant (42.1%), resistant (30.3%), and responsive patients (38.4%). Thus, C PMID- 10852459 TI - Preserved male fertility despite decreased androgen sensitivity caused by a mutation in the ligand-binding domain of the androgen receptor gene. AB - Mutations in the androgen receptor gene are considered as incompatible with preservation of fertility and have been suggested as a cause of male infertility. Two adult brothers, referred because of gynecomastia and hormonal levels in serum indicating androgen insensitivity (high sex hormone-binding globulin, and LH levels, despite extremely high testosterone concentration), turned out to be relatives to a third young man, referred independently of the two others and exhibiting identical clinical and hormonal stigmata. In all three men, we found a C-->A substitution at position 2470 (exon 7) in the androgen receptor gene, leading to a Gln824Lys mutation in the ligand-binding domain of the receptor. Exploring the family history revealed that their grandfathers, on their mothers' side, were brothers; and the Gln824Lys mutation was also found in the one of them who was still alive. Binding studies with the mutant receptor in transfected COS 7 cells, with mibolerone as ligand, exhibited equal Kd (0.7 vs. 1.0 nmol/ L), IC50 (0.8 vs. 1.1 nmol/L), and maximum binding (7.1 vs. 8.9 fmol/ 10(6) cells), as compared with the wild-type (WT) receptor. In a chloramphenicol acetyl transferase trans-activation assay, the activity of the mutant receptor was identical to that of the WT, when the synthetic androgen R1881 was'used as a ligand; but with dihydrotestosterone, in concentrations up to 10 nmol/L, the activity of Gln824Lys mutated receptor was 10-62% of the WT variant. Thus, Gln824Lys mutation was found, both in vivo and in vitro, to cause slight impairment of receptor function but was compatible with preservation of male fertility. The patients inherited the mutation from their grandfathers through their mothers, and one of the young men possessing the mutation has fathered a daughter. PMID- 10852460 TI - Blood micronutrient and thyroid hormone concentrations in the oldest-old. AB - Several micronutrients are involved in thyroid hormone metabolism, but it is unclear whether their marginal deficits may contribute to the alterations in thyroid function observed in extreme aging. The relationships among blood concentrations of thyroid hormones and selenium, zinc, retinol, and alpha tocopherol were studied in 44 healthy Northern Italian oldest-old subjects (age range, 90-107 yr), selected by the criteria of the SENIEUR protocol. Control groups included 44 healthy adult (age range, 20-65 yr) and 44 SENIEUR elderly (age range, 65-89 yr) subjects. Oldest-old subjects had higher TSH (P < 0.01) and lower free T3 (FT3)/freeT4 (FT4) ratio, zinc, and selenium serum values (P < 0.001) than adult and elderly control subjects. No significant difference was found for plasma retinol and a-tocopherol values. The associations between micronutrients and thyroid hormones were evaluated by multivariate analysis. In oldest-old subjects, plasma retinol was negatively associated with FT4 (P = 0.019) and TSH serum levels (P = 0.040), whereas serum zinc was positively associated with serum FT3 (P = 0.010) and FT3/FT4 ratio (P = 0.011). In younger subjects, no significant association was found among thyroid variables and micronutrients. In conclusion, blood levels of specific micronutrients are associated with serum iodothyronine levels in extreme aging. PMID- 10852461 TI - Leucine 7 to proline 7 polymorphism in the neuropeptide Y gene is associated with enhanced carotid atherosclerosis in elderly patients with type 2 diabetes and control subjects. AB - We have recently demonstrated that subjects having Pro7 in the signal peptide ofneuropeptide Y (NPY) have higher serum cholesterol and apolipoprotein B levels than individuals with wild-type (Leu7Leu7) signal peptide sequence. We investigated the association of Leu7Pro polymorphism with common carotid intima media thickness (IMT) assessed by ultrasonograph in patients with type 2 diabetes (n = 81; 41 men and 40 women; mean age, 67.1 yr) and nondiabetic subjects (n = 105; 48 men and 57 women; mean age, 65.5 yr) and genotyped for the Leu7Pro polymorphism in prepro-NPY. The frequency of Pro7 in prepro-NPY was 9.9% (8 of 81) in diabetic patients and 14.3% (15 of 105) in control subjects (P = 0.360). The mean common carotid IMT was 1.04 +/- 0.02 mm in nondiabetic subjects without the Leu7Pro polymorphism and 1.14 +/- 0.04 mm in nondiabetic subjects with in (P = 0.156) and 1:18 +/- 0.03 and 1.58 +/- 0.21mm in diabetic patients without and with the Leu7Pro polymorphism (P = 0.004), respectively. In the analysis of covariance of the entire group, the mean common carotid IMT was independently associated with the Leu7Pro polymorphism (F = 5.165; P = 0.024) after adjustment for known risk factors. Thus, the presence of the Pro7 substitution in the prepro NPY associates with increased carotid atherosclerosis. PMID- 10852462 TI - Activating thyrotropin receptor mutations are present in nonadenomatous hyperfunctioning nodules of toxic or autonomous multinodular goiter. AB - Toxic multinodular goiter, a heterogeneous disease producing hyperthyroidism, is frequently found in iodine-deficient areas. The pathogenesis of this common clinical entity is still unclear. The aim of the present study was to search for activating TSH receptor (TSHr) or Gs alpha mutations in areas of toxic or functionally autonomous multinodular goiters that appeared hyperfunctioning at thyroid scintiscan but did not clearly correspond to definite nodules at physical or ultrasonographic examination. Surgical tissue specimens from nine patients were carefully dissected, matching thyroid scintiscan and thyroid ultrasonography, to isolate hyperfunctioning and nonfunctioning areas even if they did not correspond to well-defined nodules. TSHr and Gs alpha mutations were searched for by direct sequencing after PCR amplification of genomic DNA. Only 2 adenomas were identified at microscopic examination, whereas the remaining 18 hyperfunctioning areas corresponded to hyperplastic nodules containing multiple aggregates of micromacrofollicules not surrounded by a capsule. Activating TSHr mutations were detected in 14 of these 20 hyperfunctioning areas, whereas no mutation was identified in nonfunctioning nodules or areas contained in the same gland. No Gs alpha mutation was found. In conclusion, activating TSHr mutations are present in the majority of nonadenomatous hyperfunctioning nodules scattered throughout the gland in patients with toxic or functionally autonomous multinodular goiter. PMID- 10852463 TI - Reduced pancreatic B cell compensation to the insulin resistance of aging: impact on proinsulin and insulin levels. AB - Type 2 diabetes mellitus is associated with insulin resistance, reduced B cell function, and an increase in the proinsulin (PI) to immunoreactive insulin (IRI) ratio (PI/IRI); the latter is thought to be an indication of B cell dysfunction. Normal aging is associated with insulin resistance and reduced B cell function, but it is not known whether changes in PI and the PI/IRI ratio are also a feature of the aging-associated B cell dysfunction. Therefore, we tested whether the aging-associated changes in insulin sensitivity and B cell function result in changes in PI and IRI levels that are proportionate or whether they are disproportionate as in type 2 diabetes. Twenty-six healthy older (mean +/- SEM age, 67 +/- 1 yr) and 22 younger (28 +/- 1 yr) subjects with similar body mass indexes (27.9 +/- 0.6 vs. 26.3 +/- 1.0 kg/m2) were studied. PI was measured by a RIA recognizing both intact PI and its conversion intermediates. The insulin sensitivity index (SI) was quantified using the minimal model, and B cell function was measured as fasting insulin levels, the acute insulin response to glucose (AIRglucose), and as the acute insulin response to arginine at maximal glycemic potentiation (AIRmax). B cell function was also adjusted for SI based on the known hyperbolic relationship between these two variables. Older and younger subjects had similar fasting glucose (5.3 +/- 0.1 vs. 5.2 +/- 0.1 mmol/L), IRI (83 +/- 8 vs. 76 +/- 9 pmol/L), PI (8.9 +/- 0.8 vs. 10.6 +/- 2.0 pmol/L), and PI/IRI ratio (12.3 +/- 1.3% vs. 13.9 +/- 1.6%; all P = NS) despite a 50% reduction of insulin sensitivity (SI, 1.94 +/- 0.21 vs. 3.88 +/- 0.38 x 10(-5) min(-1)/pmol x L; P < 0.001) and in B cell function [SI x fasting IRI, 139 +/- 18 vs. 244 +/- 24 x 10(-5)(P < 0.001); SI x AIRglucose, 0.75 +/- 0.13 vs. 1.70 +/- 0.15 x 10(-2) min(-1) (P < 0.001); SI x AIRmax, 3.63 +/- 0.53 vs. 6.81 +/- 0.70 x 10(-2) min(-1) (P < 0.001)] in the older subjects. These findings suggest that the B cell dysfunction in older subjects is not associated with disproportionate proinsulinemia. However, in older subjects the B cell response to the insulin resistance of aging is reduced whether measured as fasting levels of PI or IRI or as the acute response to secretagogues. Thus, when examined in terms of the degree of insulin sensitivity, the lower fasting IRI levels in older subjects suggest that the utility of fasting insulin levels as a surrogate measure of insulin resistance in older individuals may be limited. PMID- 10852464 TI - Male hypogonadism caused by homozygous deletion of exon 10 of the luteinizing hormone (LH) receptor: differential action of human chorionic gonadotropin and LH. AB - We report the unique case of a patient with Leydig cell hypoplasia (LCH) type II caused by a genomic deletion resulting in the complete absence of exon 10 of the LH receptor (LHR). The patient presented at the age of 18 yr with retarded pubertal development, small testicles, and delayed bone maturation. LH was highly elevated, with very low serum testosterone levels. Genetic analysis revealed a homozygous deletion of approximately 5 kbp encompassing exon 10 of the LHR gene. Screening of family members demonstrated heterozygosity for the deletion, indicating autosomal recessive inheritance. At the time of examination, the patient displayed nearly normal male phenotype, but lacked pubertal development and was hypogonadal. Obviously, fetal male development sustained by hCG was normal, whereas LH action, important for pubertal development, was impaired. A hCG stimulation test induced testosterone biosynthesis and secretion within the normal range. Subsequently, hCG treatment was continued, resulting in an increase in testicular volume and the appearance of spermatozoa in the ejaculate after 16 weeks of treatment (5.3 million/mL). Despite highly elevated endogenous LH serum levels, the response to hCG indicates a possible dual mechanism of hormone binding and signal transduction for hCG and LH on a LHR that lacks exon 10. Furthermore, this patient represents the clinical counterpart of the normal male marmoset monkey (Callithrix jacchus), in which the expressed LHR lacks exon 10 in toto. This case provides important clinical insights about the possible role of exon 10 of the LHR in discriminating between LH and hCG actions. PMID- 10852465 TI - Identification of a new missense mutation (Gly95Glu) in a highly conserved codon within the high-mobility group box of the sex-determining region Y gene: report on a 46,XY female with gonadal dysgenesis and yolk-sac tumor. AB - Leydig cells and Sertoli cells of the testes produce hormones that cause male differentiation, if receptors are present. The Y chromosomal SRY gene (sex determining Region Y gene) acts as TDF and is required for regular male sex determination. SRY represents a transcription factor belonging to the superfamily of genes sharing the HMG-box motif(high-mobility group-box), which acts as DNA binding region. Here, we describe a nonmosaic XY sex-reversed female with pure gonadal dysgenesis (46,XY karyotype, completely female external genitalia, normal Mullerian ducts, absence of Wolffian ducts, streak gonads) who harbored a yolk sac tumor and was referred for the assessment of primary amenorrhea. Using genomic PCR analysis, a 423-bp PCR product, encompassing the HMG-box of the SRY gene, was amplified from the proposita, her father, and her three brothers, whereas no band was visible in the patient's mother and her three sisters. The PCR products were sequenced for mutations subsequently. A new de novo missense mutation within the HMG-box of the SRY gene was discovered in the proposita. A G is replaced by an A in codon 95 at position +284, resulting in the replacement of the nonpolar aminoacid glycine by the polar amino acid glutamate. The glycine at codon 95 is highly conserved between the family of HMG-box proteins and between species. This point mutation has not been described earlier and brings the total number of SRY mutations described so far to 36, each mutation being unique. This mutation was not detected in the patient's father and her male siblings. The present data provide further evidence to support the functional importance of the putative DNA binding activity of the SRY HMG-box domain. PMID- 10852466 TI - Measurement of plasma free luteinizing hormone beta-subunit in women. AB - Little is known about the physiological secretion of the free beta-subunit of LH (LHbeta). The aim of this study was to compare in women the secretion of LHbeta, using sensitive and specific two-site immunoassays, with dimeric LH and the free common alpha-subunit (FAS). The LHbeta assay does not recognize the dimeric LH and cross-reacts only with free hCG beta-subunit (CGbeta). Thus, all of the plasma samples were also tested with a highly specific immunoradiometric assay for free CGbeta. Molar concentrations (i.e. picomoles per L) were used to compare the plasma levels of LH and its free subunits. Plasma LH, LHbeta, FAS, and CGbeta levels were measured in five normally cycling women during the early follicular phase and the ovulatory peak of LH. The pulsatile profiles of LH, LHbeta, FAS, and CGbeta were studied in five postmenopausal women before and 21 days after injection of a depot preparation of the GnRH agonist D-Trp6 (3.75 mg, im) and in five women with functional hypothalamic amenorrhea (FHA), i.e. low plasma LH levels, during pulsatile GnRH administration (20 microg/pulse, 90 min, sc). Afterward, one of the patients with FHA received a single sc injection of 1350 U recombinant human LH, and plasma LH, LHbeta, FAS, and CGbeta levels were measured and compared with the high plasma levels of one postmenopausal woman. In cycling women, basal plasma LHbeta and CGbeta levels were below the detection limit of the assays (1.34 and 0.65 pmol/L, respectively), and plasma FAS levels were 13.60 +/- 0.13 pmol/L. During the LH surge, there was a parallel increase in LH, LHbeta, and FAS. Plasma CGbeta levels remained undetectable. In normal postmenopausal women, basal plasma dimeric LH, LHbeta, and FAS levels were increased in parallel, and their pulsatile profiles were similar, without measurable plasma CGbeta levels. After D-Trp6 administration, plasma LH and LHbeta levels were completely suppressed, whereas plasma FAS levels increased, and plasma CGbeta remained below 0.65 pmol/L. In FHA women, basal plasma levels of LH and FAS were low, without detectable LHbeta and CGbeta levels. During pulsatile GnRH administration, LHbeta became detectable, and pulses were synchronous with those of LH and FAS. The secretion of LH and LHbeta was almost equimolar. Plasma CGbeta levels remained undetectable. In the patient with FHA, administration of recombinant human LH increased only plasma LH levels, whereas plasma LHbeta and FAS levels remained very low. In conclusion, when the production of dimeric LH increases, a concomitant, parallel, and almost equimolar hypersecretion of uncombined and biologically inactive LHbeta occurs. Like the alpha-subunit, LHbeta may be secreted in the dissociated free form. This can lead to pitfalls during clinical investigations if assays of free CGbeta display some cross-reaction with free LHbeta. PMID- 10852467 TI - Quantitative assessment of pituitary resistance to thyroid hormone from plots of the logarithm of thyrotropin versus serum free thyroxine index. AB - Previous studies have shown that, in patients with primary alterations in thyroid hormone secretion, the level of the natural logarithm of serum TSH (lnTSH) is negatively related to the level of free T4. Because such patients can generally be assumed to exhibit normal tissue responsivity to thyroid hormone, we were interested in determining whether the lnTSH/free T4 index (FTI) relationship in patients with established thyroid hormone resistance (THR) exhibit a lower slope than patients with normal tissue sensitivity to thyroid hormone. We have therefore analyzed the relationship between the lnTSH and the FTI in members of three families with documented THR. In these patients, a given dose of T4 was maintained for a 1- to 2-month period, to achieve hormonal equilibration. Two of the families, though not related, exhibited the same mutation, E460K. The third was identified as A317T. As anticipated, the slope of the lnTSH/FTI ratio was significantly lower in the patients with THR than in T4-treated patients who were presumed to have normal sensitivity to thyroid hormone. The slope of the lnTSH/FTI relationship seemed to be characteristic of the specific mutation involved in the three genotypes (wild-type and two mutations) examined. Further, the in vivo slope of the lnTSH/FTI relationship seemed to be linearly related to the T3 association constant of the in vitro translated receptor. These findings support the potential usefulness of measuring the slope of lnTSH, as a function of the FTI, in quantitating pituitary THR. PMID- 10852468 TI - Differential activity of the cytochrome P450 17alpha-hydroxylase and steroidogenic acute regulatory protein gene promoters in normal and polycystic ovary syndrome theca cells. AB - 17alpha-Hydroxylase (CYP17) expression in propagated theca cells isolated from the ovaries of women with polycystic ovary syndrome (PCOS) is persistently elevated, compared with theca cells isolated from normal ovaries. To investigate the mechanism for increased CYP17 messenger RNA accumulation in PCOS theca cells, we examined CYP17 and steroidogenic acute regulatory protein (StAR) promoter activities in normal and PCOS theca cells. Conditions were established to transiently transfect human theca cells with reporter gene constructs containing 5' truncations of the human CYP17 and StAR promoters. Cotransfection of a steroidogenic factor-1 expression plasmid was found to be required for detection of basal and forskolin-stimulated CYP17 promoter activity but not for StAR promoter activity. However, cotransfection with a steroidogenic factor-1 expression plasmid augmented both basal and forskolin-stimulated StAR promoter activity. CYP17 reporter activity was compared in theca cells isolated from normal and PCOS patients. Basal and forskolin-stimulated CYP17 promoter activity was 4-fold greater in PCOS cells than in theca cells isolated from normal ovaries. In contrast, StAR promoter activity, and the activity of a reporter construct containing three copies of a cAMP response element (3xCRE), were similar in normal and PCOS theca cells. The results of these studies document: 1) that basal and cAMP-dependent CYP17 gene transcription is increased in PCOS theca cells; 2) that there is differential regulation of promoters of genes required for steroidogenesis in PCOS theca cells; and 3) that passaged normal and PCOS theca cells provide a model system for studying tissue-specific regulation of genes encoding steroidogenic enzymes and identifying the molecular mechanisms involved in increased androgen production in PCOS. PMID- 10852469 TI - Physiological relationships of uncoupling protein-2 gene expression in human adipose tissue in vivo. AB - The physiological significance of changes in uncoupling protein-2 (UCP-2) gene expression is controversial. In this study we investigated the biochemical and functional correlates of UCP-2 gene expression in sc abdominal adipose tissue in humans in vivo. UCP-2 messenger ribonucleic acid expression was quantified by nuclease protection in adipose tissue from lean and obese humans in both the fasting and postprandial states. Plasma fatty acids, insulin, and leptin were all determined in paired samples from the superficial epigastric vein and radial artery, and local production rates were calculated from 133Xe washout. In the fasting state UCP-2 expression correlated inversely with body mass index (r = 0.45; P = 0.026), percent body fat (r = -0.41; P = 0.05), plasma insulin (r = 0.47; P = 0.02), epigastric venous fatty acids (r = -0.45; P = 0.04), and leptin (r = -0.50; P = 0.018). UCP-2 expression remained inversely related with plasma leptin after controlling for percent body (r = -0.45; P = 0.038). At 2 or 4 h postprandially, there were no significant relationships between UCP-2 expression and biochemical parameters. In conclusion, 1) UCP-2 messenger ribonucleic acid expression in sc adipose tissue is inversely related to adiposity and independently linked to local plasma leptin levels; and 2) UCP-2 expression is not acutely regulated by food intake, insulin, or fatty acids. Reduced UCP-2 expression may be a maladaptive response to sustained energy surplus and could contribute to the pathogenesis and maintenance of obesity. PMID- 10852470 TI - Expression and relationship between endothelin-1 messenger ribonucleic acid (mRNA) and inducible/endothelial nitric oxide synthase mRNA isoforms from normal and preeclamptic placentas. AB - Preeclampsia is a mainly vascular disease of pregnancy, probably caused by an imbalance between vasodilator and vasoconstrictor agents that results in generalized vasospasm and poor perfusion in many organs. Among these factors, endothelin-1 (ET-1), a potent vasoconstrictor, is highly increased in preeclamptic women, while nitric oxide (NO), a vasodilator of human utero placental arteries, is reduced in the same patients. The present study was designed to investigate the interactions between ET-1 and the NO system in the feto-placental unit; to this purpose we also examined the messenger ribonucleic acid (mRNA) expression of ET-1, inducible NO synthase (iNOS), and endothelial NOS (eNOS) in human cultured placental trophoblastic cells obtained from preeclamptic (PE) and normotensive (NT) pregnancies. We also studied whether exogenous ET-1 may affect the expression of iNOS and eNOS in human placental trophoblastic cells. Interestingly, by Northern blot analysis we observed an increased ET-1 mRNA expression level in PE trophoblastic cells compared to NT trophoblastic cells. Furthermore, exogenous ET-1 (10(-7) mol/L) was able to up-regulate its own mRNA expression in both NT and PE trophoblastic cells. iNOS and eNOS mRNA expression was then detected, by semiquantitative PCR, in both NT and PE trophoblastic cells. PE trophoblastic cells expressed lower iNOS mRNA levels compared with NT pregnancies. On the contrary, eNOS mRNA expression was higher in PE trophoblastic cells than in NT cells. Moreover, in the presence of ET-1 we observed a decrease in iNOS and an increase in eNOS mRNA expression levels in both NT and PE trophoblastic cells compared with the respective untreated cells. In conclusion, we demonstrate that ET-1 expression is increased in PE cells, whereas iNOS, which represents the main source of NO synthesis, is decreased; conversely, eNOS expression is increased. Finally, ET-1 is able to influence its own as well as NOS isoform expression in normal and PE trophoblastic cultured cells. These findings suggest the existence of a functional relationships between ET(s) and NOS isoforms that could constitute the biological mechanism leading to the reduced placental blood flow and increased resistance to flow in the feto maternal circulation, which are characteristic of the pathophysiology of preeclampsia. PMID- 10852471 TI - 3beta-hydroxysteroid dehydrogenase autoantibodies are rare in premature ovarian failure. AB - Premature ovarian failure (POF) is a disorder of heterogeneous etiology, and autoimmunity has been suspected as one cause of POF. The steroidogenic enzyme, 3beta-hydroxysteroid dehydrogenase (3betaHSD), has been characterized as a potential autoantigen in POF as well as in insulin-dependent diabetes mellitus (type 1 diabetes). Here we studied the presence of steroid cell antibodies (SCA), autoantibodies to 3betaHSD and to two other known autoantigens in ovarian failure, steroidogenic enzymes 17alpha-hydroxylase (P450c17), and side-chain cleavage enzyme (P450scc) in POF patients and patient groups with autoimmune polyendocrinopathy syndromes type 1 and 2 (APS1 and -2), isolated Addison's disease, type 1 diabetes, and healthy controls. The SCA were found in 2 of 48 POF, 11 of 15 APS1, and 1 of 9 APS2, and autoantibodies to in vitro translated 3betaHSD protein were detected in 1 POF serum associated with Addison's disease and 3 APS1 sera. All 3betaHSD precipitating sera were also positive for SCA. However, no SCA or 3betaHSD autoantibodies were found in 38 Addison's disease, 28 type 1 diabetes, and 71 healthy control sera. In analysis of autoantibodies to P450c17 and P450scc, antibodies to these enzymes were not found in POF sera, but were found in 10 and 12 APS1 patient sera, respectively, and 1 APS2 patient serum contained anti-P450c17 antibodies. Our results show that autoantibodies to 3betaHSD in POF patients are rare and are also found in patients with APS1. PMID- 10852472 TI - Immunoassay of insulin-like growth factor-binding protein-3 (IGFBP-3): new means to quantifying IGFBP-3 proteolysis. AB - Posttranslational modifications, particularly proteolysis, may play a significant role in the regulation of insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) physiology, and thus, measurement of modified variants of IGFBP-3 and/or their combination ratios may have important research and diagnostic relevance. Based on evaluation of a panel of monoclonal and polyclonal IGFBP-3 antibodies, we constructed three new enzyme-linked immunosorbent assays (ELISAs) using a common capture and polyclonal (ELISA-3) or monoclonal (ELISA-1 and -2) detection antibodies and evaluated them in a two-step colorimetric procedure. Evaluation of ELISA-1-3 demonstrated detection limit, dynamic range, overall precision, and recovery of the added IGFBP-3 to be generally less than 0.04 microg/L, 2-100 microg/L, less than 10%, and 91-113%, respectively. IGF-I and II, and IGFBP-1, -2, -4, -5, and -6 did not interfere. In normal adult sera (n = 26), seminal plasma (n = 14), pregnancy sera (n = 30), and amniotic fluid (n = 30), ELISA-1-3 detected significantly different IGFBP-3 levels (by up to 6-fold, on the average), whereas levels in seminal plasma determined by ELISA-1 were undetectable. Comparison of the values obtained vs. corresponding levels by an established method (Diagnostic Systems Laboratories, Inc., active IGFBP-3 ELISA) were similarly sample dependent and, on the average, varied by up to 19-fold. Only ELISA-3 compared well with the Diagnostic Systems Laboratories, Inc., IGFBP 3 ELISA when samples from normal adults were analyzed. The observed variability could not be totally explained by 50% lower reactivity of ELISA-1-3 for glycosylated IGFBP-3 vs. the nonglycosylated form, and changes in phosphorylation had no effect on immunoreactivity. Evaluation of IGFBP-3 after proteolysis by seminal plasma, plasmin, or thrombin suggested recognition of intact IGFBP-3 by ELISA-1, whereas ELISA-3 appeared to measure intact and proteolyzed IGFBP-3 (total IGFBP-3) with similar potency. In contrast, levels determined by ELISA-2 increased severalfold, indicating preferential recognition of IGFBP-3 fragments. We propose that immunoassay capable of differential determination of IGFBP-3 variants may help better define the physiological importance and potential clinical value of IGFBP-3 measurements. PMID- 10852473 TI - Changes in endometrial PTEN expression throughout the human menstrual cycle. AB - Frequent mutation of the PTEN tumor suppressor gene in endometrial adenocarcinoma has led to the prediction that its product, a phosphatase that regulates the cell cycle, apoptosis, and possibly cell adhesion, is functionally active within normal endometrial tissues. We examined PTEN expression in normal human endometrium during response to changing physiological levels of steroid hormones. PTEN ribonucleic acid levels, assessed by RT-PCR, increase severalfold in secretory compared to proliferative endometrium. This suggested that progesterone, a known antineoplastic factor for endometrial adenocarcinoma, increases PTEN levels. Immunohistochemistry with an anti-PTEN monoclonal antibody displayed a complex pattern of coordinate stromal and epithelial expression. Early in the menstrual cycle under the dominant influence of estrogens, the proliferative endometrium shows ubiquitous cytoplasmic and nuclear PTEN expression. After 3-4 days of progesterone exposure, glandular epithelium of early secretory endometrium maintains cytoplasmic PTEN protein in an apical distribution offset by expanding PTEN-free basal secretory vacuoles. By the midsecretory phase, epithelial PTEN is exhausted, but increases dramatically in the cytoplasm of stromal cells undergoing decidual change. We conclude that stromal and epithelial compartments contribute to the hormone-driven changes in endometrial PTEN expression and infer that abnormal hormonal conditions may, in turn, disrupt normal patterns of PTEN expression in this tissue. PMID- 10852474 TI - Inhibition of early luteal angiogenesis by gonadotropin-releasing hormone antagonist treatment in the primate. AB - Angiogenesis during luteal development is essential for normal lutein cell function, but the control of this process and the relationships between the steroidogenic and endothelial cells have still to be elucidated. The aim of this study was to: 1) quantify endothelial cell proliferation throughout the luteal phase of the marmoset ovulatory cycle; 2) determine the effect of gonadotropin withdrawal using GnRH antagonist treatment on the early luteal phase angiogenesis peak; and 3) describe the resultant morphological changes in the corpus luteum (CL). Ovaries were collected during the early, mid-, and late luteal phase, and changes in angiogenic activity were determined by quantification of bromodeoxyuridine incorporation. Animals were treated with a GnRH antagonist, on luteal days 1 and 2, and ovaries were collected on day 3. A proliferation index was obtained by counting the number of bromodeoxyuridine immunopositive cells in luteal sections. Cell proliferation was maximal in the early luteal phase and fell significantly in the mid- and late CL. GnRH antagonist treatment reduced the early luteal phase proliferation peak by 90%, suppressed plasma progesterone, and severely disrupted lutein cell morphology. These results demonstrate that the intense angiogenesis in the early primate CL is dependent on gonadotropin stimulation of lutein cells. PMID- 10852475 TI - Expression of the heme oxygenase-carbon monoxide signalling system in human placenta. AB - Heme oxygenase (HO) catalyses the formation of endogenous carbon monoxide and bilirubin from heme. CO, a potent vasodilator, and bilirubin, an antioxidant may have local actions in the fetal-placental vasculature of the placenta. We sought evidence of expression of the two known isoforms of HO in normal human term placenta using reverse transcription polymerase chain reaction (RT-PCR), western blotting and immunohistochemistry. RT-PCR demonstrated the presence of messenger ribonucleic acid for HO-1 and HO-2. Immunoreactive proteins of appropriate size for each HO enzyme were identified in placental cell membrane preparations. Immunohistochemistry showed a wide distribution of HO immunoreactivity, including the syncytiotrophoblast layer of placental villi, the endothelium and smooth muscle cells of umbilical-placental blood vessels, and in all layers of the fetal membranes. These data demonstrate the expression of the two known isoforms of HO in human placenta and suggest that endogenous CO and bilirubin may have important roles in the control of placental vascular function. PMID- 10852476 TI - The Aspergillus nidulans creC gene involved in carbon catabolite repression encodes a WD40 repeat protein. AB - Expression of many microbial genes required for the utilisation of less favoured carbon sources is carbon catabolite repressed in the presence of a preferred carbon source such as D-glucose. In Aspergillus nidulans, creC mutants show derepression in the presence of D-glucose of some, but not all, systems normally subject to carbon catabolite repression. These mutants also fail to grow on some carbon sources, and show minor morphological impairment and altered sensitivity to toxic compounds including molybdate and acriflavin. The pleiotropic nature of the phenotype suggests a role for the creC gene product in the carbon regulatory cascade. The creC gene was cloned and found to encode a protein which contains five WD40 motifs. The sequence changes in three mutant alleles were found to lead to production of truncated proteins which lack one or more of the WD40 repeats. The similarity of the phenotypes conferred by these alleles implies that these alleles represent loss of function alleles. Deletion analysis also showed that at least the most C-terminal WD40 motif is required for function. The CreC protein is highly conserved relative to the Schizosaccharomyces pombe protein Yde3--whose function is unknown--and human and mouse DMR-N9, which may be associated with myotonic dystrophy. PMID- 10852477 TI - A novel secreted ribonuclease from Bacillus intermedius: gene structure and regulatory control. AB - A second secreted ribonuclease, designated binase II, has been detected in Bacillus intermedius 7P, and its structural gene was cloned and sequenced. Unlike the well-known binase I, a 109-amino acid guanyl-specific enzyme, the 292-residue binase II is closely related to the B. subtilis nuclease Bsn, in structure and in its enzymatic properties. Binase II is also insensitive to inactivation by barstar, an inhibitor protein that is specific for guanyl-specific ribonucleases. While both B. intermedius enzymes are induced upon phosphate starvation, only the gene for binase I belongs to the pho regulon system and carries pho-box elements adjacent to its promoter sequence. The gene for binase II is similar to that for Bsn in lacking such elements. The birB gene coding for binase II appears to be located next to the 3'-end of a ferric ion transport operon, with which it convergently overlaps. This would allow attenuator control over binase II expression under conditions of starvation for ferric ions. PMID- 10852478 TI - Complete nucleotide sequence of the Oenothera elata plastid chromosome, representing plastome I of the five distinguishable euoenothera plastomes. AB - We describe the 159,443-bp [corrected] sequence of the plastid chromosome of Oenothera elata (evening primrose). The Oe. elata plastid chromosome represents type I of the five genetically distinguishable basic plastomes found in the subsection Euoenothera. The genus Oenothera provides an ideal system in which to address fundamental questions regarding the functional integration of the compartmentalised genetic system characteristic of the eukaryotic cell. Its highly developed taxonomy and genetics, together with a favourable combination of features in its genetic structure (interspecific fertility, stable heterozygous progeny, biparental transmission of organelles, and the phenomenon of complex heterozygosity), allow facile exchanges of nuclei, plastids and mitochondria, as well as individual chromosome pairs, between species. The resulting hybrids or cybrids are usually viable and fertile, but can display various forms of developmental disturbance. PMID- 10852479 TI - Ribosomal DNAs: an exception to the conservation of gene order in rice genomes. AB - rDNA (18S-5.8S-25S rDNA) and 5S rDNA loci were visualized on the chromosomes of six species of the genus Oryza by fluorescence in situ hybridization (FISH) and the labeled rice chromosomes were identified based on their condensation patterns. As a result, the chromosomes harboring rDNA and/or 5S rDNA loci were determined in the complement for all the known rice genomes. Variation in the location of the rDNA loci indicated the transpositional nature of the rDNAs in the genus Oryza, as also suggested in Triticeae and Allium. Comparative analysis of the locations of rDNA loci among rice, maize and wheat revealed that variability in the physical location of the rDNA loci was characteristic of the genus Oryza and also of the genera of Gramineae. This variability in the location of the rDNA loci between evolutionarily related species is in sharp contrast to the conservation of the general order of genes in their genomes. PMID- 10852480 TI - The bacteriophage D108 Ner repressor binds a conformationally distinct operator. AB - The Ner protein encoded by the transposable coliphage D108, an 8.6 kDa lambda Cro like repressor, binds to an operator spanning 50 bp of DNA. The distinguishing features of this operator are two perfect 11-bp inverted repeats (5'-CCGTGAGCTAC 3') that are separated by an 8-bp AT-rich spacer. Hyperreactivity of the ner operator to potassium permanganate and the hydroxyl radical indicate that the AT rich spacer assumes a variant conformation consistent with a bend. Using an electrophoretic mobility shift assay, we demonstrated that Ner does not display significant affinity for a single 11-bp site. Furthermore, DNase I protection analysis and circular-permutation binding assays reveal that alterations in the length and sequence of the AT-rich spacer that separates the 11-bp inverted repeats significantly alter Ner-operator interactions, and demonstrate that the intrinsically bent ner operator is conformationally altered upon protein binding. PMID- 10852481 TI - Regulation of expression of the Aspergillus niger benzoate para-hydroxylase cytochrome P450 system. AB - Cytochrome P450 enzyme systems are found throughout nature and are involved in many different, often complex, bioconversions. In the endoplasmic reticulum of the filamentous fungus Aspergillus niger a cytochrome P450 enzyme system is present that is capable of the para-hydroxylation of benzoate. The expression of the two genes encoding the components of this system, the cytochrome P450 gene encoding benzoate para-hydroxylase (bphA) and the gene encoding cytochrome P450 reductase (cprA), is inducible by benzoate. The BPH system was used as a model system to study the mechanisms that result in co-regulation of both components of an eukaryote cytochrome P450 enzyme system. Deletion analysis of the transcription control regions of cprA and bphA resulted in the identification of a region that was involved in benzoate induction of gene expression. The functional competence of the cprA Benzoate Responsive Region thus defined was demonstrated directly by cloning this fragment upstream of a constitutively expressed mini-promoter and analysing expression of the hybrid transcription control region in a lacZ reporter system. Further analysis of cprA gene expression revealed a clear quantitative discrepancy between induction at the protein level (approximately 4-fold) and at the transcription level (> 20-fold). The majority of the transcripts observed after benzoate induction (cprAbeta) were larger then the constitutively expressed cprAalpha transcript. The difference in size between the cprAalpha and cprAbeta transcript is caused by differential promoter use. As the longer cprAbeta transcript carries a small uORF we propose that post-transcriptional regulation of CPR expression underlies the discrepancy in the degree of induction at the protein and transcriptional level. Our results show that regulation of CPR expression is particularly complex, involving regulatory promoter elements, differential promoter use and regulation at the post-transcriptional level. PMID- 10852482 TI - Disruption of the petB-petD intergenic region in tobacco chloroplasts affects petD RNA accumulation and translation. AB - Translation initiation in chloroplasts is a complex process involving a variety of cis-elements and trans-acting factors. Many chloroplast mRNAs are processed products of polycistronic primary transcripts, but the functional requirement for processing is mostly enigmatic. In tobacco, the petB and petD genes, which encode subunits of the cytochrome b6/f complex, are transcribed from the psbB operon, whose primary transcript is processed into products including di- or tricistronic, but not monocistronic, petB and petD mRNAs. To begin to identify elements important for petB and/or petD translation, we generated tobacco chloroplast transformants by inserting selectable aadA marker cassette in the petB-petD intergenic region. The resulting plants required sucrose for growth, and their phenotypes depended on the orientation of the aadA cassette. When aadA was inserted in the same transcriptional orientation as the psbB operon, petB and petD mRNAs were abundantly produced but aberrant in size, and only 25% of the wild-type amount of the cytochrome b6/f complex accumulated. With the aadA cassette in the opposing orientation, however, very little petD mRNA accumulated, and the cytochrome b6/f complex was undetectable. Polysome analysis suggested that petD mRNAs in both transformants were poorly translated, indicating that the intergenic region contains essential translational elements. PMID- 10852483 TI - Core-binding specificity of bacteriophage integrases. AB - The site-specific recombination systems of bacteriophages lambda and HK022 share the same mechanism and their integrase proteins show strong homology. Nevertheless the integrase protein of each phage can only catalyze recombination between its own att sites. Previous work has shown that the specificity determinants in the att sites are located within the sequences that bind the integrase to the core of att. DNA fragments that carry attL and attR sites of each phage were challenged with each of the two integrases and the DNA-protein complexes were examined by the gel-retardation technique. The results show that each integrase can form higher-order DNA-protein complexes only with its cognate att sites, suggesting that differences in the mode of binding to the core are responsible for the specificity difference between the two integrases. PMID- 10852484 TI - REAL, an LTR retrotransposon from the plant pathogenic fungus Alternaria alternata. AB - A retrotransposon was isolated and characterized from strain 15A of the Japanese pear pathotype of Alternaria alternata, which causes black spot disease in certain cultivars of Japanese pear by producing a host-specific toxin known as AK toxin. The element, which we have named REAL (Retrotransposon of Alternaria alternata), is 6046 bp in size and contains direct long terminal repeats (LTRs) of 218 bp. Target-site duplication of 5 bp was found. REAL contains two long overlapping ORFs. The first ORF shows homology to retroviral gag genes. The second ORF has homology to protease, reverse transcriptase, RNase H and integrase domains of the retroelement pol genes, in that order. Phylogenetic comparison of reverse transcriptase domains from retrotransposons placed REAL in the Ty3/gypsy group of LTR retrotransposons, most closely related to grasshopper from Magnaporthe grisea. Northern analysis detected REAL transcripts of about 2.0 and 6.0 kb. The 6.0-kb species corresponds to a full-length transcript of the element. The element was found by Southern analysis in 12 out of 13 strains of the Japanese pear pathotype, and the banding patterns, copy numbers and signal intensities in these strains were variable. REAL-related elements were also found in some, but not all, of the other strains tested, including nonpathogenic A. alternata and other pathotypes, which cause diseases on different plant species by producing distinct hostspecific toxins. These results suggest that the distribution of REAL in A. alternata is not pathotype specific. PMID- 10852485 TI - Direct isolation of differentially expressed genes from a specific chromosome region of common wheat: application of the amplified fragment length polymorphism based mRNA fingerprinting (AMF) method in combination with a deletion line of wheat. AB - The amplified restriction fragment length polymorphism (AFLP)-based mRNA fingerprinting (AMF) method makes it possible systematically and conveniently to identify differentially expressed cDNAs with high reproducibility. We have applied the AMF method to the cloning of the Q gene of common wheat, which is located on the long arm of chromosome 5A and pleiotropically controls the spike morphology and the threshing character of seeds. Using the AMF method, we compared the fingerprints of mRNA samples extracted from the young spikes of Triticum aestivum cv. Chinese Spring (CS) carrying the Q gene to those of a chromosome deletion line of CS, namely, q5, which lacks 15% of 5AL including the Q gene. Approximately 12,200 fragments were produced after PCR with 256 primer combinations. Of these, 92 fragments were differentially expressed between CS and q5. Northern and Southern analyses showed that 16 fragments gave specific or relatively stronger transcript signals in CS, and these clones were present in single copy or in low copy numbers in the wheat genome. Four clones were genetically mapped to the region deleted in q5. Subsequently, one clone, pTaQ22, was mapped at the same locus as the Q gene, indicating that pTaQ22 corresponds to the Q gene or is tightly linked to it. DNA sequence data showed that pTaQ22 had no homology to any known genes, thus suggesting a novel function for this gene in flower morphogenesis. This AMF method might provide a straightforward method for isolating genes in the hexaploid background of common wheat. PMID- 10852486 TI - Extra-ribosomal function(s) of the plastid ribosomal protein L4 in the expression of ribosomal components in spinach. AB - We have previously characterised the cDNA corresponding to the nucleus-encoded, plastid ribosomal protein L4 from spinach. The L4 protein belongs to the group of ribosomal proteins for which extra-ribosomal functions have been demonstrated in prokaryotes. In general, these functions are concerned with the expression of ribosomal components. In order to analyse whether the plastid L4 protein might also have (an) extra-ribosomal function(s) we have produced the plastid L4 protein as a thioredoxin fusion protein and analysed its role in both prokaryotic (E. coli) and plastid systems. We found that the plastid L4 protein can replace the E. coli L4 protein in the NusA-dependent attenuation control of the E. coli S10 operon by stabilising stalled transcription complexes in a NusA-dependent reaction. In plastids, the L4 protein inhibits transcription of the rrn operon. Our results thus suggest extra-ribosomal function(s) for the plastid L4 protein in the expression of ribosomal components. PMID- 10852487 TI - Transformation of Arabidopsis with a Brassica SLG/SRK region and ARC1 gene is not sufficient to transfer the self-incompatibility phenotype. AB - Self-incompatibility (SI) promotes outbreeding in flowering plants, and in Brassica SI is genetically controlled by the S locus. Self-incompatible Brassica and self-fertile Arabidopsis belong to the same crucifer family. In addition, a comparative analysis reveals a high degree of microsynteny between the B. campestris S locus and its homologous region in Arabidopsis--with the notable exception that the Brassica SI genes, SLG and SRK, are missing. Brassica ARC1 encodes a component of the SRK signal transduction pathway leading to self-pollen rejection, and no closely related ARC1 homolog has been identified in Arabidopsis. The purpose of the research reported here was to introduce Brassica SI components into Arabidopsis in an attempt to compensate for the missing genes and to investigate whether the SI phenotype can be transferred. Inserts of approximately 40 kb from the fosmid clones F20 and F22, which span the B. napus W1 SLG-SRK region, were cloned into the plant transformation vector pBIBAC2. Transgenic plants were generated that expressed the Brassica SI genes in the flower buds. In addition, the endogenous, SLG-like, gene AtS1 was not co suppressed by the Brassica SLG transgene. No SI phenotype was observed among the T1 BIBAC2-F20 and BIBAC2-F22 transgenic plants. When the ARC1 gene was transformed into BIBAC2-F20 or BIBAC2-F22 plants, the resulting BIBAC2-F20-ARC1 and BIBAC2-F22-ARC1 plants still set seeds normally, and no rejection response was observed when self-incompatible B. napus W1 pollen was placed on BIBAC2-F20 ARC1 or BIBAC2-F22-ARC1 Arabidopsis stigmas. Taken together, our results suggest that complementing Arabidopsis genome with Brassica SLG, SRK and ARC1 genes is unlikely to be sufficient to transfer the SI phenotype. PMID- 10852488 TI - A novel nucleic acid-binding protein in the cyanobacterium Synechococcus sp. PCC6301: a soluble 33-kDa polypeptide with high sequence similarity to ribosomal protein S1. AB - Cyanobacteria are prokaryotes that carry out plant-type photosynthesis and contain several eukaryotic-type RNA-binding proteins. Using a single-stranded DNA column, a 33-kDa protein was isolated and characterized from Synechococcus sp. PCC6301. This protein of 293 amino acids is similar in overall structure to the ribosomal protein S1 found in the same species, and contains three repeated units that are highly similar to the S1 motif originally found in the ribosomal protein S1 of Escherichia coli. However, the 33-kDa protein was found not to be associated with ribosomes and its nucleic acid binding specificity is distinct from that of the ribosomal protein S1. As this protein has high affinity for both single- and double-stranded DNA, as well as for poly(G) and poly(A), we tentatively named it nucleic acid-binding protein 1 (Nbp1). PMID- 10852489 TI - Cloning and expression of the S-adenosylmethionine decarboxylase gene of Neurospora crassa and processing of its product. AB - S-adenosylmethionine decarboxylase (AdoMetDC) catalyzes the formation of decarboxylated AdoMetDC, a precursor of the polyamines spermidine and spermine. The enzyme is derived from a proenzyme by autocatalytic cleavage. We report the cloning and regulation of the gene for AdoMetDC in Neurospora crassa, spe-2, and the effect of putrescine on enzyme maturation and activity. The gene was cloned from a genomic library by complementation of a spe-2 mutant. Like other AdoMetDCs, that of Neurospora is derived by cleavage of a proenzyme. The deduced sequence of the Neurospora proenzyme (503 codons) is over 100 codons longer than any other AdoMetDC sequence available in genomic databases. The additional amino acids are found only in the AdoMetDC of another fungus, Aspergillus nidulans, a cDNA for which we also sequenced. Despite the conserved processing site and four acidic residues required for putrescine stimulation of human proenzyme processing, putrescine has no effect on the rate (t0.5 approximately 10 min) of processing of the Neurospora gene product. However, putrescine is absolutely required for activity of the Neurospora enzyme (K0.5 approximately 100 microM). The abundance of spe-2 mRNA and enzyme activity is regulated 2- to 4-fold by spermidine. PMID- 10852490 TI - RFLP-based analysis of three RbcS subfamilies in diploid and polyploid species of wheat. AB - The RbcS multigene family of hexaploid (bread) wheat, Triticum aestivum (genome BBAADD), which encodes the small subunit of Rubisco, comprises at least 22 genes. Based on their 3' non-coding sequences, these genes have been classified into four subfamilies (SFs), of which three (SF-2, SF-3 and SF-4) are located on chromosomes of homoeologous group 2 and one (SF-1) on homoeologous group 5. In the present study we hybridized three RbcS subfamily-specific probes (for SF-1, SF-2 and SF-3) to total DNA digested with four restriction enzymes and analyzed the RFLP patterns of these subfamilies in eight diploid species of Aegilops and Triticum, and in two tetraploid and one hexaploid species of wheat (the diploid species are the putative progenitors of the polyploid wheats). The three subfamilies varied in their level of polymorphism, with SF-2 being the most polymorphic in all species. In the diploids, the order of polymorphism was SF-2 > SF-3 > SF-1, and in the polyploids SF-2 > SF-1 > SF-3. The RbcS genes of the conserved SF-1 were previously reported to have the highest expression levels in all the wheat tissues studied, indicating a negative correlation between polymorphism and gene expression. Among the diploids, the species with the D and the S genomes were the most polymorphic and the A-genome species were the least polymorphic. The polyploids were less polymorphic than the diploids. Within the polyploids, the A genome was somewhat more polymorphic than the B genome, while the D genome was the most conserved. Among the diploid species with the A genome, the RFLP pattern of T. urartu was closer to that of the A genome of the common wheat cultivar Chinese Spring (CS) than to that of T. monococcum. The pattern in Ae. tauschii was similar to that of the D genome of CS. Only partial resemblance was found between the RFLP patterns of the species with the S genome and the B genome of CS. PMID- 10852491 TI - A high-resolution linkage map of the vicinity of the rice submergence tolerance locus Sub1. AB - Resistance to submergence stress is an important breeding objective in areas where rice cultivars are subjected to complete inundation for a week or more. The present study was conducted to develop a high-resolution map of the region surrounding the submergence tolerance gene Sub1 in rice, which derives from the Indian cultivar FR13A. Submergence screening of 8-day-old plants of F3 families kept for 14 days submerged in 60 cm of water allowed an accurate classification of Sub1 phenotypes. Bulked segregant analysis was used to identify AFLP markers linked to Sub1. A population of 2950 F2 plants segregating for Sub1 was screened with two RFLP markers flanking the Sub1 locus, 2.4 and 4.9 cM away. Submergence tolerance was measured in the recombinant plants, and AFLP markers closely linked to Sub1 were mapped. Two AFLP markers cosegregated with Sub1 in this large population, and other markers were localized within 0.2 cM of Sub1. The high resolution map should serve as the basis for map-based cloning of this important locus, as it will permit the identification of BAC clones spanning the region. PMID- 10852492 TI - The Drosophila dominant wing mutation Dichaete results from ectopic expression of a Sox-domain gene. AB - The dominant Drosophila wing mutation Dichaete is characterised by the deletion of proximal wing structures. By analysing a number of new Dichaete alleles, phenotypic revertants and enhancer piracy lines, we show that the wing phenotype results from ectopic expression of the Sox-domain gene Dichaete. Ectopic expression of the Sox gene results in an increase in cell death in the proximal region of the wing imaginal disc and leads to alterations in the normal expression of wingless. Since ectopic expression of wingless in the proximal region of the wing disc can rescue aspects of the Dichaete phenotype, it is likely that Dichaete specifically interferes with the establishment or maintenance of a critical domain of wingless expression in the wing disc. PMID- 10852493 TI - Tagging of genes involved in multidrug resistance in Aspergillus nidulans. AB - We have used a plasmid containing the Neurospora crassa pyr4 gene to transform an Aspergillus nidulans pyrG89 mutant strain in the presence of Bam-HI, and isolated multidrug-sensitive mutants among the transformants. Using this approach, we hoped to identify genes whose products are important for drug resistance by analyzing gene disruptions that alter the drug sensitivity of the cell. About 1300 transformants isolated following transformation were screened for sensitivity to drugs or various stress agents with different and/or the same mechanism of action. Seventy-seven of these transformants showed sensitivity to at least one drug, while fourteen transformants showed a complex phenotype of sensitivity to different drugs. The pyr4 marker was shown to be tightly linked to the mutant phenotype in only 36% of the pleiotropic mutants analyzed in sexual crosses. Genetic crosses between our multidrug-sensitive transformants and cycloheximide-sensitive and imazalil-resistant mutants of A nidulans were performed to determine whether mutations were present at the same loci. We have shown that the gene imaD that confers resistance to imazalil may also be involved in cycloheximide and hygromycin sensitivity, since this mutation is allelic to scyB (mutant scy290). In addition, the cross between the transformant R223 and the imazalil-resistant mutant ima535 showed that both mutations are in the same complementation group, suggesting that the gene imaG could also be involved in cycloheximide and itraconazole sensitivity. PMID- 10852494 TI - Differential complementation of a Neurospora crassa Galpha(i) mutation using mammalian Galpha protein genes. AB - Heterotrimeric (alphabetagamma) G proteins interact with sensory receptors to transduce signals to downstream effectors in eukaryotes. We previously reported that GNA-1 from Neurospora crassa is a microbial member of the Galphai family found in higher organisms. Deletion of gna-1 leads to female sterility, slower growth rates on normal and hyperosmotic solid medium, and increased resistance to heat and oxidative stress. In this study we compare mammalian genes for proteins of the Galphai sub-family (Galphai, Galphao, Galphat and Galphaz), and Galphas (which is not a member of the Galphai family) with the N. crassa gna-1 gene with respect to their ability to complement deltagna-1 phenotypes. Northern analysis detected full-length transcripts of all these genes, except that for Galphai, in N. crassa transformants. Measurements of pertussis toxin-catalyzed ADP ribosylation and Western analysis showed that the GNA-1, Galphaz, Galphao and Galphas proteins were present in the respective transformed strains. Strains in which the mammalian Galpha protein could be detected were subjected to phenotypic testing. During the vegetative cycle, none of the mammalian Galpha genes complemented the thermotolerance phenotype of deltagna-1. However, the three expressed mammalian Galpha genes achieved at least partial complementation of the defects in vegetative apical extension rate. cAMP levels did not correlate with restoration of vegetative growth rate by the mammalian genes. During the sexual cycle, Galphao was the only mammalian Galpha gene that rescued the defect in female fertility characteristic of deltagna-1 strains. Alignment of GNA-1, Galphaz, Galphao and Galphas protein sequences revealed correlations between the observed complementation pattern and the degree of identity to GNA-1 in various functional motifs. The finding that Galphac gave the best restoration of vegetative growth but could not restore normal female fertility implies that GNA 1 regulates different pathways that are important for vegetative and sexual growth in N. crassa. PMID- 10852495 TI - Homologous recombination and transposition generate chromosome I neopolymorphism during meiosis in Saccharomyces cerevisiae. AB - We have studied the meiotic segregation of a chromosome length polymorphism (CLP) in the yeast Saccharomyces cerevisiae. The neopolymorphism frequently observed within the smallest chromosomes (I, VI, III and IX) is not completely understood. We focused on the analysis of the structure of chromosome I in 88 segregants from a cross between YNN295 and FL100trp. Strain FL100trp is known to carry a reciprocal translocation between the left arm of chromosome III and the right arm of chromosome I. PCR and Southern hybridization analyses were performed and a method for the rapid detection of chromosome I rearrangements was developed. Seven chromosome I types were identified among the 88 segregants. We detected 22 recombination events between homologous chromosomes I and seven ectopic recombination events between FL100trp chromosome III and YNN295 chromosome I. These recombination events occurred in 20 of the 22 tetrads studied (91%). Nine tetrads (41%) showed two recombination events. This showed that homologous recombination involving polymorphic homologues or heterologous chromosomes is the main source of neopolymorphism. Only one of the seven chromosome I variants resulted from a transposition event rather than a recombination event. We demonstrated that a Tyl element had transposed within the translocated region of chromosome I, generating mutations in the 3' LTR, at the border between U5 and PBS. PMID- 10852496 TI - Transfer of the mitochondrial rps10 gene to the nucleus in rice: acquisition of the 5' untranslated region followed by gene duplication. AB - Mitochondrial ribosomal protein S10 (rps10) is encoded by the mitochondrial genome in potato and pea. Here we show that the rps10 gene is absent from the mitochondrial genome of rice and has been transferred to the nucleus. Cloning and transcriptional analysis show that there are two rps10 genes in the rice nuclear genome and that their transcripts differ in abundance. Western analysis detected the RPS10 protein in the soluble fraction of rice mitochondria, although neither RPS10 has any obvious N-terminal presequence for targeting to mitochondria. This result suggests that targeting information is present in the internal region of rice RPS10. Genomic sequence analysis indicated that each rps10 gene has an intron in the 5' untranslated region (5' UTR) and that these intron sequences are homologous to each other. This result strongly suggests that a duplication event occurred after transfer of the rps10 gene to the nucleus. The duplicated rps10 genes have since been translocated to different chromosomes, because the two rps10 genes were mapped on chromosomes 6 and 12 by RFLP analysis. Interestingly, the 5' UTR and the intron of the rice rps10 genes are homologous to sequences found in several rice genes with various functions, such as osk4, EF-1beta2 and RAG1, suggesting a common origin and a functional role for the 5' UTR. Acquisition of the 5' flanking region might have accelerated the activation of the mitochondrial rps10 gene which was transferred to the nuclear genome. PMID- 10852497 TI - Renal toxicity and carcinogenicity of trichloroethylene: key results, mechanisms, and controversies. AB - The discussion on renal carcinogenicity of trichloroethylene addresses epidemiological, mechanistic, and metabolic aspects. After trichloroethylene exposure of rats, renal cell tumors were found increased in males, and an increased incidence of interstitial cell tumors of the testes was reported. Studies on the metabolism of trichloroethylene in rodents and in humans support the role of bioactivation reactions for the development of tumors following exposure to trichloroethylene. Epidemiological cohort studies addressing the carcinogenicity of trichloroethylene with respect to the renal or urothelial target sites have been conducted, and no clear evidence for an elevated renal or urinary tract cancer risk in trichloroethylene-exposed groups was visible in exposed populations. However, a cohort study of 169 male workers having been exposed to unusually high levels of trichloroethylene in Germany within the period between 1956 and 1975 supported a nephrocarcinogenic effect of trichloroethylene in humans. The results of this study were discussed in the literature with considerable reserve; criticism was based mainly on the choice of the study group, which had been recruited from personnel of a company in which a cluster of four renal tumors was observed previously. Hence, a further case control study was conducted in the same region. This study confirmed the results of the previous cohort study, supporting the concept of involvement of prolonged and high-dose trichloroethylene exposures in the development of renal cell cancer. Further investigations on patients with renal cell carcinoma and with histories of high trichloroethylene exposures, on the basis of excretion of marker proteins in the urine, pointed to toxic damage to the proximal renal tubules by trichloroethylene. The hypothesis of implication of a glutathione transferase-dependent bioactivating pathway of trichloroethylene, established in experimental animals, seems at least also plausible for humans. Apparently, the occurrence of renal cell carcinomas in man follows high-dose exposures to trichloroethylene that are also accompanied by damage to tubular renal cells. Development of renal cell carcinomas has been related to mutations in the vonHippel-Lindau (VHL) tumor suppressor gene. Renal cell carcinoma tissues of persons with histories of prolonged high-dose exposure to trichloroethylene were investigated for the occurrence of mutations of the vonHippel-Lindau (VHL) tumor suppressor gene. VHL gene mutations were found in the majority of renal cell tumors associated with high-level exposure to trichloroethylene. A specific mutational hot spot at the VHL nucleotide 454 was addressed as a unique mutation pattern of the VHL tumor suppressor gene. A synopsis of all experimental, clinical, and epidemiological data suggests that reactive metabolites of trichloroethylene, with likely involvement of dichlorovinyl-cysteine (DCVC), exert a genotoxic effect on the proximal tubule of the human kidney. This constitutes a tumor-initiating process of genotoxic nature, the initial genotoxic effect apparently being linked with mutational changes in the VHL tumor suppressor gene. However, there is compelling evidence that the full development of a malignant tumor requires continued promotional stimuli. Repetitive episodes of high peak exposures to trichloroethylene over a prolonged period of time apparently led to nephrotoxicity, visualized by the excretion of tubular marker proteins in the urine. This critical process of development of tubular damage by trichloroethylene must follow a "conventional" dose-dependence, implying a practical threshold. This view is much corroborated by the fact that the occurrence of human renal cell cancer is obviously confined to cases of unusually high trichloroethylene exposures in the past, with special characteristics of very high and repetitive peak exposures. Current instruments of regulation should be adjusted to allow adequate consideration of su PMID- 10852498 TI - Heterologous expression of xenobiotic mammalian-metabolizing enzymes in mutagenicity tester bacteria: an update and practical considerations. AB - There is an increasing need for metabolic competent cell systems for the mechanistic studies of biotransformation of xenobiotics in toxicology in general and in genotoxicology in particular. These cell systems combine the heterologous expression of a particular mammalian biotransformation enzyme with a specific target/ end point by which a functional analysis of the expressed gene product in the (geno)toxicity of chemicals can be performed. cDNAs of an increasing number of mammalian biotransformation enzymes is being cloned. The construction of specific expression vectors permits their heterologous expression in laboratory bacteria, such as Escherichia coli strains. This development does not only allow biochemical and enzymatic studies of (pure) enzyme preparations but also facilitates the engineering of metabolically competent mutagenicity tester bacteria, thereby providing new tools for genotoxicity testing and for studying of the roles of biotransformation in chemical carcinogenesis. In this review, we describe an update as well as an evaluation of enzymes expressed in mutagenicity tester bacteria. Four types of biotransformation enzymes are now expressed in these bacteria, namely, GSTs, CYPs, NATs, and STs. The expression of these enzymes in the tester bacteria and their subsequent application in mutagenicity assays demonstrates that heterologous expression in this type of bacteria has a number implications for the functionality of the biotransformation enzymes as well as for the functioning of the tester bacteria in mutagenicity detection. We also describe here a number of practical considerations in this regard. PMID- 10852500 TI - Current status of topical nasal antimicrobial agents. AB - The nasal cavity and paranasal sinuses are probably one of the last frontiers in the head and neck region where the use of topical antimicrobial agents is not yet established. Although the anatomy of the nasal cavity and the paranasal sinuses can theoretically be exploited for the administration of antimicrobials in rhinosinusitis, very few studies have been conducted to test the feasibility of this mode of therapy. We review the anatomical and physiological factors that should be considered in the use of topical nasal antimicrobial agents and the current status of topical nasal antimicrobial usage, and we make recommendations for the administration of topical nasal antimicrobial agents. PMID- 10852499 TI - Toxicity review of ethylene glycol monomethyl ether and its acetate ester. AB - Ethylene glycol monomethyl ether (EGME) and its acetate ester (EGMEA) are highly flammable, colorless, moderately volatile liquids with very good solubility properties. They are used in paints, lacquers, stains, inks and surface coatings, silk-screen printing, photographic and photo lithographic processes, for example, in the semiconductor industry, textile and leather finishing, production of food contact plastics, and as an antiicing additive in hydraulic fluids and jet fuel. EGME and EGMEA are efficiently absorbed by inhalation as well as via dermal penetration. Dermal absorption may contribute substantially to the total uptake following skin contact with liquids or vapours containing EGME or EGMEA. EGMEA is rapidly converted to EGME in the body and the two substances are equally toxic in animals. Therefore, the two substances should be considered as equally hazardous to man. Effects on peripheral blood, testes, and sperm have been reported at occupational exposure levels ranging between 0.4 and 10 ppm EGME in air, and with additional, possibly substantial, dermal exposure. Severe malformations and disturbed hematopoiesis have been linked with exposure to EGME and EGMEA at unknown, probably high, levels. Embryonic deaths in monkeys and impaired spermatogenesis in rabbits have been reported after daily oral doses of 12 and 25 mg per kg body weight, respectively. In several studies, increased frequency of spontaneous abortions, disturbed menstrual cycle, and subfertility have been demonstrated in women working in the semiconductor industry. The contribution of EGME in relation to other exposure factors in the semiconductor industry is unclear. PMID- 10852501 TI - Universal newborn hearing screening in an inner-city, managed care environment. AB - OBJECTIVES/HYPOTHESIS: Universal neonatal hearing screening (UNHS) programs aim to identify and treat educationally significant hearing loss in the first months of life. Several states have mandated UNHS for all newborns. Such programs have been successful in small, homogeneous populations. As larger states attempt to implement such programs, important obstacles have arisen, particularly in sparsely populated rural environments and in the inner city, where poverty, unstable living situations, and inadequate access to health care make follow-up of infants failing initial testing difficult. STUDY DESIGN: We performed a prospective longitudinal study examining the effects of increasingly complex and expensive interventions designed to ensure that children failing initial hearing screening returned for complete evaluation and habilitation. METHODS: A UNHS program based on transient evoked otoacoustic emissions testing was implemented at Temple University Hospital, with 2,000 births per year. At 6 months into the program, efficacy was assessed and modifications in follow-up methodology were made in an attempt to improved rate of return of infants failing newborn screening. The effect of these interventions was reassessed 6 months later. RESULTS: In its first 12 months, the Temple University Infant and Young Child Hearing Intervention Initiative successfully screened 95% (2,031) of all newborns using transient evoked otoacoustic emissions. Collecting a complete database profile for each newborn, establishing rapport with the family, and offering immediate follow-up appointments yielded a 61% return rate after discharge. The addition of a dedicated project secretary, free day-care for siblings, and cab vouchers for transportation and the elimination of a requirement for health maintenance organization referrals increased follow-up yield to 75%. CONCLUSION: Given adequate resources and planning, UNHS can be successful, even in economically depressed environments. PMID- 10852502 TI - Quality of life and recurrence concern in survivors of head and neck cancer. AB - OBJECTIVES/HYPOTHESIS: A cohort of 3-year survivors of head and neck cancer was evaluated for persistent quality of life (QOL) concerns and long-term treatment effects. STUDY DESIGN: Mailed questionnaire. METHODS: The questionnaire with the University of Washington Quality of Life (UWQOL) scale, the Performance Status Scale for Head and Neck Cancer (PSS-HN), the Functional Assessment of Cancer Therapy (FACT) scale, and the Functional Assessment of Cancer Therapy Head and Neck (FACT-HN) scale and locally prepared questions was sent to 111 3-year disease-free survivors. Analysis was performed to statistically evaluate the effect of stage, site, treatment type, surgery, and cancer concern on QOL. Current smoking information was gathered. RESULTS: Seventy-two survivors completed the questionnaire. Advanced stage was correlated with lower QOL scores in the domains of disfigurement, chewing ability, speech, and eating in public. QOL scores did not vary by initial tumor site. Patients treated with irradiation alone had statistically better QOL scores than those treated with combined surgery/radiation therapy in the pain, disfigurement, chewing, and speech domains. Laryngectomy and composite resection survivors reported lower QOL scores than patients treated with irradiation alone. A low level of cancer concern persisted in about half of the long-term survivors. Cancer concern was associated with continued pain, disfigurement, and limitations on eating in public. Three quarters of the tobacco users had quit by the time of the questionnaire. Nevertheless, the patients were not thoroughly convinced that tobacco had caused their cancer. CONCLUSIONS: Long-term survivors of head and neck cancer experience QOL effects well after completion of treatment. Effects are most pronounced in survivors who required combined surgery/radiation therapy. Continuing low levels of cancer concern persist in about half of the survivors. Many cancer survivors successfully quit smoking. PMID- 10852503 TI - Polymer chemotherapy for head and neck cancer. AB - OBJECTIVES: To study a new method of delivery of chemotherapy for the treatment of squamous cell carcinomas (SCCs) of the head and neck, to evaluate the pharmacokinetics of four anticancer agents (cisplatin, fluorouracil [5-FU], methotrexate [MTX], and paclitaxel) loaded into the biodegradable polymer, polyanhydride polymer poly(FAD:SA), and to evaluate the effectiveness and toxicity of the drug-polymer combination against human SCCs, both in vitro and in vivo. STUDY DESIGN: Poly(FAD:SA) was loaded with different chemotherapeutic drugs and its in vitro and in vivo drug release and tissue penetration characteristics were studied. The biocompatibility and toxicity of the polymer-drug combination were determined. The effectiveness of the drug-polymer was evaluated against three different human SCCs (larynx O11, pharynx FADU, and floor of mouth UM- SCC1) cultured in vitro and in nude mice carrying human SCC xenografts. METHODS: The in vitro drug release pharmacokinetics of the drugs were performed using atomic absorption spectrometry for cisplatin and high-pressure liquid chromatography for the 5-FU, MTX, and paclitaxel studies. In vitro tumor cytotoxicity was assessed by growth assay. In vivo cytotoxicity was assessed by growth rate inhibition in a nude mouse model. RESULTS: All four chemotherapy drugs demonstrated a continuous release that followed first-order kinetics from the polymer. More than 95% of the MTX and 5-FU, 70% of the cisplatin, and 20% of the paclitaxel was released within the 10 days of the assay. Tumor cytotoxicity revealed that the polymer was very effective against the human SCCs O11, FADU, and UM- SCC1 in vitro. When a small amount of polymer (1-2 g) was added to the cell culture and left for 7 days, 96.6% of the UM-SCC1 cells, 86.9% of the FADU cells, and 94.6% of the O11 cells were killed. When the culture medium was then changed every 2 days to remove the effect of nutrient depletion or chemicals released by the degrading polymer, 74% of the UM-SCC1 cells, 94.5% of the FADU cells, and 66.1% of the O11 cells were killed at 7 days. The tumor animal model was the nude mouse carrying human floor of mouth SCC xenografts. Different amounts of cisplatin were incorporated into the polymers (5% and 7% drug/polymer at a weight/weight [wt/wt] load). Thirty-five days after implantation of the polymer in nude mice, the mean treated tumor size was 65.5% of controls in the 5% group and 31.8% in the 7% group. Seventy days after implantation the mean treated tumor size was 41.4% of controls in the 5% group and 38.1% in the 7% group, indicating a statistically significant delay of tumor growth compared with controls or with intraperitoneally injected cisplatin. The blank polymer was well tolerated by the mouse and had no effect on tumor growth. CONCLUSIONS: The study results indicate that polymer chemotherapy is effective against a variety of SCCs of the head and neck, both in vitro and in vivo, and may become a useful therapeutic option for head and neck cancer. PMID- 10852504 TI - Surgical management of localized amyloidosis. AB - OBJECTIVE: To demonstrate the role of two-dimensional reconstruction images on computed tomography (CT) in the treatment planning for laryngeal amyloidosis. To discuss the treatment for isolated laryngeal amyloidosis and compare the role of endoscopic versus an open surgical approach to management. STUDY DESIGN: Retrospective review. METHODS: The medical records from 1984 to the present with the diagnosis of localized respiratory tract amyloidosis at Geisinger Medical Center were reviewed. RESULTS: Five previously unpublished cases of localized laryngeal amyloidosis were identified with the supraglottic region the major site of involvement. Hoarseness and airway compromise were the presenting symptoms. CT two-dimensional reconstruction imaging was used to evaluate two cases with extensive laryngeal involvement that required an external surgical approach to relieve symptoms. CONCLUSIONS: Localized laryngeal amyloidosis is a rare disease that requires surgical management when symptomatic. CT two-dimensional reconstruction can be helpful in detailing the extent of disease and planning surgery. A lateral external supraglottic approach has been found to be successful in treating patients with large supraglottic masses. PMID- 10852505 TI - Partial parotidectomies: morbidity and benign tumor recurrence rates in a series of 94 cases. AB - OBJECTIVES: Review the methods available for parotidectomy. Describe the technique of partial parotidectomy assisted by evoked electromyographic nerve location and the expected morbidity and benign and low-grade cancer tumor recurrence rates from this modified procedure. STUDY DESIGN: From 1983 to 1999 the author performed or assisted in 94 parotidectomies (79 partial), all done by a single specialty group and all using evoked electromyographic nerve location. The cases were surveyed by reviewing all the hospital and office records on these cases and tabulating the type and extent of surgery, pathology, postoperative problems recorded, and long-term follow-up. METHODS: Partial parotidectomy was elected in those cases of benign and low-grade malignant disease in which adequate tumor removal required a less than complete lobectomy or total parotidectomy. Heavy reliance was placed on proactive nerve location by an evoked electromyographic device with dissecting/stimulating hemostat. A retrospective review focusing on these cases was performed based on the patient charts and their continued documentation by the practice. RESULTS: In 79 partial parotidectomies there were no documented facial nerve injuries and one incidence of recurrence of a benign mixed tumor and an acinic cell carcinoma, respectively. CONCLUSION: Partial parotidectomy has the advantages of reduced risk to the facial nerve, reduced operating time, possible outpatient surgery, and no apparent risk of increased recurrence of benign tumors. PMID- 10852506 TI - Antisense RNA to eIF4E suppresses oncogenic properties of a head and neck squamous cell carcinoma cell line. AB - OBJECTIVE: The translation initiation factor eIF4E is elevated in all head and neck squamous cell cancers (HNSCCs) and appears to be essential in the progression of solid tumors. Overexpression of eIF4E results in preferential upregulation of two angiogenic factors, vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF-2). We wanted to determine whether reducing eIF4E in a HNSCC cell line could suppress its oncogenic properties and in turn decrease expression of VEGF and FGF-2. METHODS: Levels of eIF4E protein expression were determined in a panel of HNSCC cell lines. An episomal vector containing antisense RNA to eIF4E was used to reduce the eIF4E level in one of these cell lines, FaDu. After a stable transfection, Western blot analysis was performed to determine the level of eIF4E and FGF-2 reduction, while an enzyme linked immunosorbent assay (ELISA) was used to determine the level of VEGF reduction. In vitro and in vivo experiments were performed to determine whether there was a reversion in the tumorigenic properties of the FaDu cells. RESULTS: All six cell lines had elevated levels of eIF4E compared with Detroit 551, a normal cell line. Reducing eIF4E expression via antisense RNA suppressed both the tumorigenic and angiogenic properties of the FaDu cells, as demonstrated by loss of capacity to grow in soft agar, reduced expression of angiogenic factors, and loss of tumorigenicity in nude mice. CONCLUSIONS: Antisense RNA therapy to eIF4E can potentially be used as adjuvant therapy for head and neck cancers, particularly in cases in which elevated eIF4E is found in the surgical margins. PMID- 10852507 TI - Allograft dermal implant (AlloDerm) in a previously irradiated field. AB - OBJECTIVE: To evaluate the integration of AlloDerm (LifeCell Corp., The Woodlands, TX) in a field exposed to external-beam radiation (EBR) by analyzing graft thickness, fibroblast recellularization, and neovascularization. STUDY DESIGN: Randomized control. METHODS: Thirty-six male Sprague-Dawley rats (n = 36) were randomly assigned to four groups. One hind leg of each rat was exposed to 20 Gy of EBR; the other limb served as the nonirradiated control. Two weeks after irradiation, AlloDerm was implanted into both hind legs. Grafts were harvested at 3, 4, 6, and 14 weeks after implantation and underwent histological analyses. RESULTS: There was no statistically significant difference in graft thickness, fibroblast count, or neovascularization between the grafts placed in the irradiated bed and the controls (n = 33, P = .332, P = .336, and P = .057, respectively). However, at week 3, fibroblast counts in the graft placed in the field exposed to EBR were significantly lower than those of controls (P = .019), although at week 14 the counts in the experimental limb were higher than those of the controls (P = .002). Graft thickness (P = .001) and fibroblast count (P < .004) were lower at week 14 than at earlier time periods for both the experimental and control grafts. CONCLUSIONS: In the rat model, graft thickness and neovascularization of the AlloDerm dermal implant do not appear to be adversely affected by a field that has received EBR. Fibroblast ingrowth may be hindered in the early postimplantation period but appears to normalize in the long term. Furthermore, overall graft thickness and fibroblast counts decrease over time, regardless of irradiation status. PMID- 10852508 TI - A novel approach to laryngeal suspension after partial laryngectomy. AB - OBJECTIVES: Supraglottic laryngectomy is a well-established surgical therapy for selected carcinomas of the larynx and hypopharynx. Most compromised by this procedure and its variations is the laryngeal mechanism that protects the lower respiratory tract from aspiration. Laryngeal suspension serves to compensate for the loss of the resected laryngeal elevator muscles by pulling the larynx upward and forward beneath the tongue base. In this study we describe a method of laryngeal suspension in supraglottic laryngectomy using a cartilage-anchored suture carrier device. STUDY DESIGN: Report of this novel approach to laryngeal suspension using seven suture anchors in two patients undergoing supraglottic laryngectomy. METHODS: Seven Mitek Micro anchors (Mitek, Westwood, MA) were used to perform laryngeal suspension in two patients undergoing supraglottic laryngectomy. Our technique is compared with traditional methods. Operative data as well as postoperative functional results are reviewed. RESULTS: Laryngeal suspension using suture anchors was successful, with failure of only one anchor. Oral alimentation was quickly reestablished in both patients. There were no perioperative or postoperative complications. CONCLUSIONS: We describe a novel approach to laryngeal suspension that overcomes some of the technical challenges inherent in traditional suturing techniques. This novel approach is technically easier and more efficient than traditional methods and accomplishes distribution of stress forces on the thyroid cartilage remnant. PMID- 10852509 TI - Effect of photodynamic therapy on revascularization of fasciocutaneous flaps. AB - OBJECTIVE: Photodynamic therapy (PDT) may be used as an adjuvant intraoperative treatment to improve locoregional control. PDT has been shown both to delay wound healing and to have a deleterious effect on flap survival after a primary ischemic insult. This delay in wound healing may make the flap dependent on its pedicled blood supply for a prolonged period. Long-term flap loss may be experienced. The effect of PDT on flap revascularization, with subsequent dependence on its vascular pedicle, is evaluated. STUDY DESIGN: Randomized controlled trial using a rodent model. METHODS: A rat fasciocutaneous flap was used. Study groups were as follows: group I received no treatment; group II received treatment with 630-nm light; groups IH and IV were given Photofrin (in group III, loupes without a fiberoptic light source were used for flap elevation, and in group IV, light source was employed); and group V was given Photofrin and 630-nm light. Primary ischemic times of 2 or 4 hours were used. Vascular pedicles were ligated on postoperative day (POD) 5, 6, or 7, and percentage of flap survival was evaluated 7 days later. RESULTS: With 2 hours of ischemia, revascularization was decreased in the PDT group on POD 6 (P < .05) and on day 7 (P < .005) when compared with the other groups. With 4 hours of ischemia, revascularization was decreased in the PDT group on PODs 5 (P < .001), 6 (P < .01), and 7 (P < .005). CONCLUSION: Intraoperative PDT decreases revascularization of a rat fasciocutaneous flap. PMID- 10852510 TI - Neural cell adhesion molecule expression in adenoid cystic carcinoma of the head and neck. AB - OBJECTIVE: To investigate whether there is a correlation between neural cell adhesion molecule (NCAM) expression and perineural spread in patients with adenoid cystic carcinoma of the head and neck (ACCHN). STUDY DESIGN: Retrospective review of medical records and immunohistochemical staining of specimens from 37 patients treated at the University of Arkansas in Little Rock from 1987 to 1997. METHODS: Sections from paraffin-embedded specimens were examined for the presence of NCAM using monoclonal anti-NCAM antibody by avidin biotin-peroxidase immunohistochemical staining. NCAM staining was scored in each specimen and correlated with the data obtained from patient charts. RESULTS: Twenty-five of 37 specimens (68%) showed histopathological evidence of perineural spread. All 37 specimens (100%) stained positive for NCAM, regardless of perineural spread status. CONCLUSION: Our results suggested that the use of NCAM expression as a predictor of perineural spread is highly unlikely. PMID- 10852511 TI - The Becker technique for otoplasty: modified and revisited with long-term outcomes. AB - OBJECTIVES: To demonstrate a modification of the Becker technique for otoplasty and to evaluate the long-term results. STUDY DESIGN: Case series with follow-up survey assessment. METHODS: A sample of 16 patients treated by a single surgeon at an academic pediatric referral center who met the inclusion criteria was reviewed for surgical results and patient/parental satisfaction. RESULTS: A total of 30 ears underwent repair. Patients ranged from 4 to 17 years (mean age, 8.2 y) with an average follow-up of 4.6 years. One patient had an immediate postoperative hematoma from blunt trauma that was treated with good long-term results. No cases required revision surgery. Preservation of the antihelix with good to excellent ear symmetry was obtained in all patients at follow-up. All patients were happy or very happy with the surgical procedure. All patients had at least five of the six criteria for surgical success as defined by the survey. CONCLUSIONS: This modification of the Becker technique of otoplasty is efficacious for correction of protruding ears with excellent long-term results. PMID- 10852512 TI - Interlocking calvarial bone grafts: a solution for the short, depressed nose. AB - OBJECTIVES/HYPOTHESIS: In patients severely affected with collapse of the nose, deprojection and upward rotation of the nasal tip are commonly seen. Traditional maneuvers to derotate and project the tip may be insufficient, because of the natural tendency of the nasal skin/soft tissue envelope to pull the tip in a cephalic and posterior direction. If the forces of scar contracture can be resisted, the tip and dorsum should remain adequately positioned. STUDY DESIGN: Retrospective chart review of 20 cases. METHODS: Using an open rhinoplasty approach, two strips of calvarial bone are fitted together in a tongue-in-groove fashion, and esthetics are analyzed. Rotation and projection are altered as indicated. A screw inserted at the indicated level along the caudal bone graft acts to prevent retrodisplacement of either the dorsal or caudal strut as scarring occurs. RESULTS: The procedure has been used in 20 patients. Two patients had displacement of the dorsal bone graft. Two patients have been lost to follow-up. Follow-up in the remainder has ranged from 6 weeks to 4 and a half years. All have maintained adequate tip and dorsal projection without excess upward tip rotation. Bone grafts have undergone minimal resorption. CONCLUSION: The interlocking calvarial bone graft technique stabilizes the nasal tip and dorsum in such a way that resists the forces of contracture and provides improved esthetics and function. PMID- 10852513 TI - Pneumococcal antibiotic resistance and rates of meningitis in children. AB - OBJECTIVES/HYPOTHESIS: Recent studies have shown alarmingly high rates of antibiotic resistance in Streptococcus pneumoniae isolates from patients with otitis media. A recent study has implicated resistant S pneumoniae for rising rates of acute mastoiditis. The purpose of this study was to determine whether S pneumoniae antibiotic resistance has similarly affected the rate of pediatric community-acquired meningitis, the most common intracranial complication of otitis media. STUDY DESIGN: Retrospective chart review. METHODS: All cases of pediatric community-acquired meningitis treated at an academic tertiary care hospital during a 10-year period were reviewed, and meningitis rates were calculated as a proportion of yearly admissions. RESULTS: The overall rate of meningitis decreased linearly during the study period (P = .001). This was largely because of a drop in the rate of Haemophilus influenzae meningitis (P = .001), corresponding with the introduction of H influenzae type B vaccine. Annual rates of S pneumoniae meningitis did not change. Only one case of S pneumoniae meningitis was due to a highly penicillin-resistant strain and isolates from four cases had intermediate sensitivity. Twenty-four of 83 cases were associated with antecedent acute otitis media and 63% of these had been treated with antibiotics before admission. Otitis media, as a cause of meningitis, did not increase during the study period. CONCLUSION: S pneumoniae is responsible for a greater proportion of cases of pediatric community-acquired meningitis. However, this is because of a decline in the rate of H influenzae cases, not the rise in S pneumoniae antibiotic resistance. PMID- 10852514 TI - Development of a staging system for inverted papilloma. AB - OBJECTIVES: Inverted papillomas of the nose and sinuses are uncommon neoplasms. In the past decade there has been a trend toward the use of endoscopic surgical techniques in the management of these tumors, in contrast to the extensive open procedures recommended previously. This trend has not been without controversy, given the association of inverted papillomas with malignancy. It has been difficult to compare surgical approaches to these neoplasms, because of the absence of a uniformly applied staging system representing the extent of disease. It was the purpose of this study to develop such a system that could be easily applied in outcomes research. STUDY DESIGN: This study involved an integrated literature review and a synthesis of findings from a number of studies. METHODS: Previous and current clinical studies examining the treatment of inverted papilloma were reviewed. Findings were organized, and a staging system was framed based on this review. RESULTS: A simple, easily applied staging system was developed based on the extent of tumor involvement noted on endoscopic examination of the nasal cavity and computed tomography (CT) scan evaluation. CONCLUSIONS: Stage I disease is limited to the nasal cavity alone. Stage II disease is limited to the ethmoid sinuses and medial and superior portions of the maxillary sinuses. Stage III disease involves the lateral or inferior aspects of the maxillary sinuses or extension into the frontal or sphenoid sinuses. Stage IV disease involves tumor spread outside the confines of the nose and sinuses, as well as any malignancy. PMID- 10852515 TI - Bacteriology of nontraumatic maxillary sinus mucoceles versus chronic sinusitis. AB - OBJECTIVE: To compare the bacteriology of maxillary sinus mucoceles to chronic sinusitis and understand the pathogenesis of nontraumatic maxillary sinus mucoceles (NTMSM). STUDY DESIGN: Retrospective review. METHODS: Review of intraoperative bacteriology culture results obtained in patients with NTMSM. Patients with history of facial trauma or previous paranasal sinus surgery were not included in the study. The results were compared to intraoperative cultures obtained from patients with chronic sinusitis (CS). RESULTS: The study groups consisted of 16 patients with NTMSM (9 male and 7 female patients) and 211 patients with CS (86 male and 125 female patients). Cultures in the NTMSM group were positive in 7 of 16 patients (44%) (four cultures had more than one isolate). There was no growth in cultures of 9 patients (56%). On the other hand, cultures in 176 patients with CS (83%) grew organisms (42 cultures had more than one isolate); there was no growth in 35 of 211 patients (17%) (P = .0007). The cultures grew aerobic bacteria in 7 of 16 (44%) and 160 of 211 (76%) patients of the NTMSM and CS groups, respectively. Anaerobic bacteria were detected in cultures of 2 of 16 patients (12.5%) with NTMSM compared with 13 of 211 patients (6.2%) in the CS group (P = .286). The most common pathogenic aerobe in the NTMSM group was alpha-hemolytic Streptococcus, while Staphylococcus aureus was the most common in the CS group. CONCLUSION: The bacteriology of maxillary sinus mucoceles is different from that of CS. The majority of patients with mucoceles have sterile intraoperative cultures. The data do not support infection as the main origin of NTMSM. PMID- 10852516 TI - Physician experience with an optical image guidance system for sinus surgery. AB - OBJECTIVES/HYPOTHESIS: Intraoperative guidance systems have been developed which use infrared tracking technology to assist with anatomical localization during sinus surgery. Although the introduction of this technology is intended to increase the safety and efficacy of sinus surgery, little is known about its actual impact in the clinical setting. The objective of this report was to study the application and utilization of an image guidance system shared by multiple sinus surgeons in a specialty hospital. STUDY DESIGN: Combined prospective case study and retrospective analysis of physician surveys. METHODS: An optical-based image guidance system (LandmarX, Xomed, Inc., Jacksonville, FL) was used by 34 physicians to perform 754 sinonasal surgeries over a 2.5-year period at Massachusetts Eye and Ear Infirmary. In 19 cases, system registration was repeated during surgery to measure the effect of fiducial placement on system accuracy. RESULTS: The measured accuracy of anatomical localization at the start of surgery (mean value, 1.69 +/- 0.38 mm) was comparable to the perceived accuracy of 1 to 3 mm that was reported by 79% of surgeons surveyed. Operating room time (mean period, 130.6 +/- 41.1 min) correlated with the surgical procedure performed (P < .05), but not with the disease stage or revision rate. According to a majority of surgeons, use of the image guidance equipment increased operating room time by 15 to 30 minutes during initial cases and by 5 to 15 minutes once experience with the equipment had been acquired. More than 90% of surgeons anticipated their continued use of the image guidance equipment for sinus surgery at a similar or greater level in the future. CONCLUSION: An optical based image guidance system can be successfully integrated into a multisurgeon operating room environment. Use of the system provides accurate anatomical localization during sinus surgery and results in a relatively high level of physician satisfaction. PMID- 10852517 TI - A retrospective analysis of temporomandibular joint reconstruction with free fibula microvascular flap. AB - OBJECTIVES: The temporomandibular joint is occasionally encountered in extirpative surgery of the head and neck. It presents a difficult management issue. Little has been reported on functional outcomes after resection and reconstruction of the temporomandibular joint. DESIGN: A retrospective analysis consisting of chart reviews and phone interviews was performed on 17 patients who underwent fibular free flap reconstruction of the temporomandibular joint from 1993 to 1998. RESULTS: Mean follow-up in surviving patients (10) was 41.3 months. Mean age of the group was 62; male-to-female ratio was 11:6. Average hospital stay was 11.6 days. Four patients had no radiation therapy, 2 had preoperative and 11 had postoperative treatment. Five patients had one osteotomy, seven had two, one had three, and one had four. Ten patients could chew, one could not, and none were recorded for the remaining. Diet consisted of regular food for two patients, soft food for seven, full liquids for four, and tube feeds for four. Cosmesis was judged as excellent by eight patients, acceptable by two, and unacceptable by two. Five patients did not describe cosmesis. Most patients stated that bony contour was excellent, but that the soft tissue defect was noticeable. Speech was judged as intelligible by seven and moderately understandable by one. Nine patients did not describe speech. Two patients had postoperative displacement of the fibular head out of the fossa. CONCLUSION: Primary reconstruction of the temporomandibular joint with microvascular fibular flaps is a viable and effective means of restoring function. The majority of patients are able to resume oral feeds, obtain excellent or pleasing cosmetic results, and maintain intelligible speech. PMID- 10852518 TI - Morbidly obese patients with severe obstructive sleep apnea: is airway reconstructive surgery a viable treatment option? AB - OBJECTIVE: To evaluate the outcomes of airway reconstructive surgery for the treatment of severe obstructive sleep apnea in the morbidly obese patient. METHODS: Retrospective review of consecutively treated patients. Variables examined include age, sex, body mass index (BMI), respiratory disturbance index (RDI), lowest oxygen saturation (LSAT), cephalometric data, and complications. RESULTS: Twenty-one patients (13 men) with a mean age of 42.6 +/- 7.9 years and mean BMI of 45 +/- 5.4 kg/m2 were identified. The mean RDI improved from 83 +/- 30.1 to 10.6 +/- 10.8 events per hour with an improved mean apnea index from 38.4 +/- 31.3 to 1.2 +/- 1.8 events per hour. The mean LSAT improved from 63.9 +/- 17.7% to 86 +/- 7.9%. The mean BMI at the 6-month postoperative polysomnographic recording was 43 +/- 4.3 kg/m2 (P < .001). Seventeen patients (81%) were successfully treated (RDI < 20 and with minimal desaturation < 90%). The mean follow-up was 21.8 +/- 15.4 months (range, 7-66 mo). Coexisting obesity hypoventilation syndrome was related to treatment failure in two patients. One patient noted recurrence of daytime fatigue after significant weight gain 4 years after surgery and the polysomnographic recordings demonstrated the recurrence of obstructive sleep apnea. CONCLUSION: Airway reconstruction is an effective treatment for severe obstructive sleep apnea in the morbidly obese patient. Careful patient selection and identifying potential coexisting obesity hypoventilation syndrome, as well as counseling on weight reduction and avoiding continual weight gain will improve treatment outcomes. Key Words: Obstructive sleep apnea, sleep-disordered breathing, obstructive sleep apnea surgery, obesity, maxillomandibular advancement. PMID- 10852519 TI - Long-term experience with endoscopic diagnosis and treatment of salivary gland inflammatory diseases. AB - OBJECTIVES: To assess the efficacy of the sialoendoscopic technique for treatment of inflammatory salivary gland diseases. This report documents the authors' long term experience with sialoendoscopy and discusses the long-term results of the procedure, technical issues, and varieties that they have utilized, as well as the advantages and limitations of this modality. STUDY DESIGN: Retrospective clinicopathologic study of 236 patients who were endoscopically treated from 1994 to 1999 for suspected salivary gland obstructive disease. METHODS: Endoscope employed was the third generation sialoendoscope (Nahlieli Sialoendoscope, Karl Storz, Tuttlingen, Germany). RESULTS: Ten sialoendoscopies were immediate failures as a result of technical problems. In the remaining 226 glands, 170 had obstructions and 56 had sialadenitis without evidence of obstructions. The success rate was 83%. Multiple endoscopic findings were encountered. No severe complications were noted. CONCLUSION: This report demonstrates the efficacy and safety of sialoendoscopy as a promising new method for use in the diagnosis, removal, and postoperative management of sialolithiasis, sialadenitis, and other obstructive salivary gland diseases. PMID- 10852520 TI - Microbiology of healthy and diseased adenoids. AB - OBJECTIVE: To determine the qualitative and quantitative microbiology of core adenoid tissue obtained from four groups of 15 children each, with recurrent otitis media (ROM), recurrent adenotonsillitis (RAT), obstructive adenoid hypertrophy (OAH), and occlusion or speech abnormalities (controls). METHODS: Core cultures of surgically removed diseased adenoids and of healthy controls were cultured for aerobic and anaerobic bacteria. RESULTS: Polymicrobial aerobic anaerobic flora were present in all instances. Ninety-four organisms were isolated from control specimens, and 148 from ROM, 142 from RAT, and 149 from OAH specimens. The predominant aerobes in all groups were alpha-hemolytic and gamma hemolytic streptococci, Haemophilus influenzae, Staphylococcus aureus, group A beta-hemolytic streptococci, and Moraxella catarrhalis. The prominent anaerobes were Peptostreptococcus, Prevotella, and Fusobacterium species. The number, concentration and distribution of types of most organisms did not vary among the three groups of diseased adenoids. However, the number of those that are potential pathogens and those that produced beta-lactamase was lower in the control than the diseased adenoids (P < .001). CONCLUSION: The study highlights the importance of the bacterial load in the adenoids in contributing to the etiology of ROM, RAT, and OAH. PMID- 10852521 TI - Treatment of laryngeal carcinomas by laser endoscopic microsurgery. AB - OBJECTIVES: To determine if laser endoscopic microsurgery is a reliable and appropriate approach in the treatment of laryngeal cancers. STUDY DESIGN: Retrospective study of 160 patients treated from 1988 to 1996 at Liege. Analysis of indication, technique, and oncologic results. METHODS: Glottic tumors were treated with either type I, type II, or type III cordectomy, with or without conservation of an inferior muscular band, and extended if necessary to all or part of the contralateral cord. For supraglottic cancers, an excision limited to a part of the vestibule, a trans-preepiglottic resection, or a radical supraglottic resection was carried out. RESULTS: Our corrected actuarial survival at 5 years was 97% for the 98 infiltrative glottic tumors and 100% for the 18 infiltrative supraglottic and 27 in situ carcinomas. No local recurrences were noted, in either the group of 118 infiltrating cancers (in whom two precancerous lesions were treated with a further laser excision), or in the 27 in situ carcinomas. Local control was thus 100%. One patient died of his cancer, with lung metastases after neck recurrence. CONCLUSIONS: Like Steiner and Rudert, this series demonstrates the oncologic validity of this surgical approach to the treatment of unadvanced glottic tumors. Unlike these authors' study, however, strict case selection, as in cases with significant involvement of the anterior commissure, has allowed us to avoid local recurrences and consequently to avoid salvage total laryngectomies. Our experience with supraglottic cancers is too small to confirm the oncologic validity of this type of surgery but seems promising. PMID- 10852522 TI - High incidence of laryngopharyngeal reflux in patients with head and neck cancer. AB - OBJECTIVES: Laryngopharyngeal reflux may play a role in the etiology of squamous cell cancer of the head and neck and contribute to complications in head and neck cancer patients after surgery or during radiotherapy. STUDY DESIGN: Prospective study. METHODS: To investigate the incidence of laryngopharyngeal and gastroesophageal reflux in patients with head and neck cancer, ambulatory 24-hour double-probe pH monitoring was performed in 24 untreated patients with laryngeal or pharyngeal squamous cell carcinoma. In addition, 10 patients who had been irradiated in the head and neck area were analyzed for reflux to study the effect of radiotherapy on reflux. RESULTS: Only 4 of the 24 head and neck cancer patients (17%) had neither pathological laryngopharyngeal nor gastroesophageal reflux. Esophageal acid exposure was abnormal in five patients and acid exposure at the level of the upper esophageal sphincter was abnormal in four patients. Eleven patients had pathological reflux in both areas. Irradiated patients did not differ from the untreated patients considering the incidence of pathological laryngopharyngeal or gastroesophageal reflux. CONCLUSIONS: The data obtained in this study indicate that reflux is a common event in head and neck cancer patients. PMID- 10852523 TI - Ki-67 (MIB1), p53, and Lewis-X (LeuM1) as prognostic factors of recurrence in T1 and T2 laryngeal carcinoma. AB - OBJECTIVES: Recently published data suggest a prognostic value of immunohistochemical proliferation markers for limited laryngeal carcinoma. Previous studies have reported contrasting findings on this issue. In this context, different treatment modalities may be responsible for contradictory findings. To study the relationship between proliferative activity--expressed by the immunohistochemical labeling index of proliferation-associated markers Ki-67 (MIB1), Lewis-X (LeuM1), and proliferating cell nuclear antigen (PCNA) and by p53 status--and treatment failure in a matched-pair study on recurrent and nonrecurrent T1 and T2 glottic carcinoma having received primary transoral laser surgery. METHODS: Twenty-one patients with tumor recurrence were randomly selected and matched with 26 patients with nonrecurrent disease regarding histopathological grading and age. MIB1 staining was used to determine the Ki-67 labeling index, and LeuM1 staining for detecting the Lewis-X antigen; immunohistochemistry determined the p53 status and PCNA labeling index. RESULTS: The Ki-67 labeling index was significantly (P = .001) higher in tumors from patients who had treatment failure (mean = 20.02%) than in patients who did not fail treatment ("nonfailures") (mean = 9.95%). Carcinoma with a Ki-67 (MIB1) labeling index above the median (15%) of the general study population showed a mean time to relapse of 23 months (n = 21), compared with 50 months for cases (n = 26) below the median (P = .016). PCNA labeling index correlated less impressively with tumor recurrence (mean = 28.59% for treatment failures, mean = 21.75% for nonfailures, P = .022). Positive detection of the Lewis-X antigen was significantly associated with recurrence (P = .015) and time to relapse (P = .006). Status of p53 was not a significant prognostic factor. CONCLUSION: The Ki 67 (MIB1) labeling index may be associated with early relapse of limited laryngeal carcinoma treated with transoral laser surgery. Since the prognostic relevance of Ki-67 seems to be different for radiological and surgical concepts of treatment, Ki-67 might become useful as criterion of therapy selection. The Lewis-X antigen, for the first time used on laryngeal carcinoma, seems to be a strong prognostic marker deserving further investigations. PMID- 10852524 TI - Aging effects on motor units in the human thyroarytenoid muscle. AB - OBJECTIVES: To determine whether age differences are present in the human laryngeal thyroarytenoid muscle that would indicate that different normative values would be needed for identifying motor unit abnormalities. STUDY DESIGN: Twenty-six consecutively recruited healthy subjects between the ages of 21 and 72 years participated in a laryngeal electromyography study. METHODS: Bipolar needle electrodes were used to record motor unit action potentials from several locations in the right and left thyroarytenoid muscles of each subject. The duration of a motor unit was measured when at least 10 firings of the same motor unit could be identified. On the average, four units were measured per muscle. RESULTS: In the subjects less than 60 years of age, motor unit duration did not increase significantly with age. However, motor units from subjects greater than 60 years of age had longer durations than those from subjects less than 60 years of age (P < .00005), and 25% of the units measured in subjects greater than 60 years of age had longer durations than any of the units measured in subjects less than 60 years of age. Further, the older subjects differed from each other in their mean unit durations (P < .0001). In subjects less than 60 years of age, significantly longer durations were found for units innervated by the longer, left-side recurrent laryngeal nerve in comparison with the right-side nerve (P = .005). CONCLUSIONS: Different mean and SD values should be used for patients less than and greater than 60 years of age and for the right and left sides, when evaluating motor units in the thyroarytenoid muscles. PMID- 10852525 TI - Fat augmentation for glottic insufficiency. AB - OBJECTIVES: Fat lipoinjection augmentation for glottic insufficiency has been used in patients with vocal fold paralysis. Relatively little information is available on the effectiveness of fat injection in patients with vocal atrophy, intubation trauma, and post-hemilaryngectomy defects. STUDY DESIGN: This paper retrospectively compares the efficiency of fat injection in patients with vocal cord paralysis (n = 9), vocal scar (n = 13), and vocal atrophy (n = 11). METHODS: The perceptual acoustic, phonatory function, and videolaryngostroboscopic data were evaluated before and after fat augmentation in 33 patients. RESULTS: Mean follow-up time was 9.7 months. Nineteen patients had excellent results. Three patients had no change. Five patients had late failure. Six patients were lost to follow-up. Phonatory function showed significant improvement in jitter, shimmer, noise-to-harmonic ratio, maximal phonation time, grade, asthenia, and breathiness (P < .05). Videolaryngostroboscopic rating showed significant improvement in right linearity of the vocal fold edge, amplitude of vocal fold vibration, excursion of the mucosal wave, vibratory behavior, and phase symmetry (P < .05). Anterior defects did better than posterior defects. Small vocal fold defects did better than large defects. CONCLUSIONS: Fat injection is a good autogenous implant and may be considered as an option in management of patients with vocal fold scar, defect, or atrophy. Reabsorption of fat is a problem, but the procedure may be repeated. PMID- 10852526 TI - Wedge turbinectomy: a new combined photocoagulative Nd:YAG laser technique. AB - OBJECTIVES: To demonstrate that Nd:YAG laser photocoagulation using a combination of interstitial and contact approaches in the surgical treatment of inferior turbinate hypertrophy yields improved results in terms of postoperative nasal patency, complications, and relapse. STUDY DESIGN: A group of 121 patients with symptoms of nasal obstruction due to hypertrophied inferior turbinates were treated between January 1994 and December 1997 at the Otolaryngology-Head and Neck Surgery Unit of the Main Military Hospital of Rome using the wedge turbinectomy, a new endonasal laser technique. METHODS: This is a combined photocoagulative procedure performed under local anesthesia. In the first surgical step we perform interstitial photocoagulation using Nd:YAG laser and in the second step we use a contact approach, making two strips of photocoagulated mucosa running side by side from the tail to the head of turbinate. RESULTS: At 1 year follow-up, the complication rate in all our patients treated with this laser technique was very low and we achieved a steady improvement in nasal patency in 104 patients (85.9%). The relapse rate was approximately 14%, but we observed that 65% of the patients who experienced long-term failure were affected by allergic rhinitis. CONCLUSIONS: An accurate preoperative evaluation of the cause of the turbinate hypertrophy is fundamental to achieving better results after laser turbinectomy and reducing the risk of relapse. PMID- 10852527 TI - Osteoplastic frontal sinus surgery with fat obliteration: technique and long-term results using magnetic resonance imaging in 82 operations. AB - OBJECTIVE: To evaluate the intraoperative and late complications of osteoplastic sinus surgery with fat obliteration with long-term magnetic resonance imaging (MRI) follow-up. METHODS: The operative records of all patients who underwent osteoplastic frontal sinus surgery with fat obliteration between January 1, 1986 and December 31, 1997 were reviewed and the postoperative clinical course and magnetic resonance imaging (MRI) scans were analyzed if available. MRI analyses revealed that changes in the distribution of fatty and fibrous tissue, the development of necrosis or oil cysts, recurrences, inflammatory complications, and mucoceles were time-dependent occurrences. RESULTS: Eighty-two operative records were evaluated and 59 patients were followed 1 to 12 years after surgery. Eighty-six MRI scans in 51 patients were available for analysis. The most frequent intraoperative complications were exposure of orbital fat (19.5%), unintentional fracture of the anterior wall (19.5%), incorrect placement of the anterior wall (17%), and dural injury (9.8%). Persistent changes of the frontal contour (embossment, depression) occurred in 10.2% and the esthetic result was unfavorable in 5.1% of the cases. Mucoceles could be detected in 5 of 51 cases (9.8%). The amount of adipose tissue detectable in the last scan was less than 20% in the majority of cases (53%), and more than 60% in only 18% of the cases. The amount of adipose tissue decreased significantly with time (the median half life was 15.4 mo). CONCLUSIONS: Osteoplastic frontal sinus surgery with fat obliteration is very useful and successful in patients in whom the frontal sinus is not accessible via an endonasal approach or the natural drainage cannot be reestablished. MRI is currently the most valuable diagnostic tool to evaluate the frontal sinus after obliteration with adipose tissue. The method has some limitations with regard to detection of small recurrent mucoceles and differentiating vital adipose tissue from fat necroses in the form of oil cysts. In these difficult cases, long-term MRI follow-up is necessary. PMID- 10852528 TI - Endoscopic terminal dacryocystorhinostomy. AB - OBJECTIVE: Traditional endoscopic dacryocystorhinostomy (DCR) involves creation of a lacrimal sac side hole that may close down in the presence of a partially patent lower lacrimal drainage system. Endoscopic terminal DCR is a modified DCR procedure designed to prevent this problem. METHODS: From September 1996 to June 1999, 16 patients (17 sides) had endoscopic terminal DCR that involved resection of the lacrimal sac-duct junction to achieve total separation of the lacrimal sac from the nasolarcrimal duct and creation of a terminal DCR opening. RESULTS: Ten of the 17 DCRs were primary and 7 were revisions. The follow-up ranged from 3 to 36 months. There were two failures. One failure was due to canalicular obstruction and the other was due to prolapse of orbital fat hindering proper fashioning of the mucosal flap. The overall success rate was 88%. CONCLUSIONS: Instead of creating a side hole in the lacrimal sac as proposed by most endoscopic DCR studies, we advocate total diversion of tear flow by performing a terminal DCR opening that would further improve the success rate of endoscopic DCR, particularly in cases of idiopathic and partial obstruction. PMID- 10852529 TI - Sinus tissue pharmacokinetics after oral administration of amoxicillin/clavulanic acid. AB - OBJECTIVES: The in vitro synergy of the amoxicillin/clavulanic acid combination has not always translated in vivo into clinical superiority compared with amoxicillin alone. Specifically, conflicting reports have disputed the superiority of the combination in the treatment of both acute otitis media and acute sinusitis. One possible reason for this may have to do with inadequate target tissue pharmacokinetics. To explore this possibility in the sinuses, we undertook the present investigation. STUDY DESIGN: A randomized, open, single dose, sinus tissue pharmacokinetic study with oral amoxicillin/clavulanic acid. METHODS: Twenty-three adult patients with chronic rhinosinusitis who had been selected for surgery were randomly allocated to receive a tablet of 875/125 mg amoxicillin/clavulanate 2 to 4 hours before surgery began. During the operation tissue samples were collected at specific sinonasal sites for determination of both amoxicillin and clavulanic acid concentration levels. RESULTS: Amoxicillin displayed adequate tissue levels throughout the sinuses, high enough to cover common susceptible pathogens. However, the presence of clavulanate was detected in only half of the sinonasal tissue samples. CONCLUSIONS: The kinetics of oral clavulanic acid apparently fails to provide a widespread anti-beta-lactamase activity capable of enhancing the activity of amoxicillin in all parts of the sinuses. Despite this, amoxicillin/clavulanic acid maintains a central role in the treatment of acute rhinosinusitis, because amoxicillin is still the most effective oral beta-lactam against Streptococcus pneumoniae, a particularly virulent and increasingly resistant upper respiratory tract pathogen. Also, as our data show, a concomitant anti-beta-lactamase activity can be expected to occur, although in an unpredictable fashion. PMID- 10852530 TI - Nasal interleukin-5, immunoglobulin E, eosinophilic cationic protein, and soluble intercellular adhesion molecule-1 in chronic sinusitis, allergic rhinitis, and nasal polyposis. AB - OBJECTIVE: To compare concentrations of interleukin-5 (IL-5), immunoglobulin E (IgE), eosinophilic cationic protein (ECP), and soluble intercellular adhesion molecule-1 (sICAM-1) in nasal secretion and serum of patients with chronic nonallergic sinusitis, allergic rhinitis, and nonallergic nasal polyposis to obtain information about the pathogenesis of these diseases. METHODS: Nasal secretion and serum were analyzed by routine enzyme-linked immunosorbent assay techniques. Nineteen patients with chronic nonallergic sinusitis, 24 patients with seasonal allergic rhinitis, and 18 patients with nonallergic nasal polyposis were included in the study. Eight healthy, nonallergic probands served as control subjects. RESULTS: Significantly elevated concentrations of IL-5 (5-fold, P < .05) and IgE (15-fold, P < .01) were detected in nasal secretion of patients with allergic rhinitis (IL-5, 51.8 +/- 13.2 pg/mL; IgE, 41.9 +/- 20.9 kU/L) or nonallergic nasal polyposis (IL-5, 57.9 +/- 36.9 pg(mL; IgE, 40.5 +/- 20.2 kU/L) compared with controls (IL-5, 10.6 +/- 7.8 pg/mL; IgE, 2.8 +/- 0.5 kU/L) or with patients with chronic nonallergic sinusitis (IL-5, 16.5 +/- 13.2 pg/mL; IgE, 5.4 +/- 3.1 kU/L). There were no significant differences between patients with allergic rhinitis and those with nonallergic nasal polyposis. Concentrations of ECP were significantly elevated (sixfold, P < .01) in patients with allergic rhinitis (297.8 ng/mL +/- 173.1) compared with controls (52.4 +/- 28.0 ng/mL) or patients with chronic nonallergic sinusitis (44.8 +/- 40.1 ng/mL), whereas twofold higher concentrations (not significant) of ECP were observed in patients with nonallergic nasal polyposis (107.1 +/- 26.6 ng/mL). Significantly elevated concentrations of sICAM-1 in nasal secretion (threefold, P < .05) were detected only in patients with chronic nonallergic sinusitis (79.4 +/- 45.6 ng/mL). The elevated sICAM-1 nasal secretion values in this group correlated significantly (P < .05) to the serum values. CONCLUSIONS: Equally elevated concentrations of IL-5 and IgE in patients with allergic rhinitis and nonallergic nasal polyposis implicated similar pathogenic processes in both diseases. Whereas the pathogenesis of allergic rhinitis is IgE-specific, the pathogenesis of nasal polyps is not as clear. IL-5 was suggested to play a pivotal role in tissue eosinophilia, which was confirmed by data in the present study. Elevated concentrations of ECP were suggested to result from tissue eosinophilia--a characteristic of both diseases. Elevated concentrations of sICAM-1 in patients with chronic nonallergic sinusitis pointed to its key role in the recruitment of neutrophils into the inflamed tissue, whereas an important role in eosinophil recruitment was ruled out. PMID- 10852531 TI - Outpatient retrograde placement of the indwelling voice prosthesis. PMID- 10852532 TI - Microvascular reconstruction of composite defects of the mandible and lip: aesthetic and functional considerations. PMID- 10852533 TI - Nitinol tracheobronchial stents: a word of caution. PMID- 10852534 TI - Understanding multiple sclerosis: lessons from pathology. PMID- 10852535 TI - The role of B cells and autoantibodies in multiple sclerosis. AB - A variety of cellular and humoral immunological abnormalities have been observed in multiple sclerosis (MS). In the past few years, several lines of evidence converged to imply an important role of autoreactive antibodies and B cells in the pathogenesis of MS. Recent data suggest that autoantibodies may be harmful in lesion formation but also potentially beneficial in repair. This review surveys recent advances in the concepts of generation and nature of pathogenetic autoantibodies, their potential modes of action, mechanisms of their long-term persistence, and the role of the inflamed brain tissue as a B-cell-supporting microenvironment in MS. Based on the presence of specific autoantibodies, it seems possible to define distinct MS subgroups in the near future. The therapeutic relevance of these new findings is presented. PMID- 10852536 TI - Heterogeneity of multiple sclerosis lesions: implications for the pathogenesis of demyelination. AB - Multiple sclerosis (MS) is a disease with profound heterogeneity in clinical course, neuroradiological appearance of the lesions, involvement of susceptibility gene loci, and response to therapy. These features are supported by experimental evidence, which demonstrates that fundamentally different processes, such as autoimmunity or virus infection, may induce MS-like inflammatory demyelinating plaques and suggest that MS may be a disease with heterogeneous pathogenetic mechanisms. From a large pathology sample of MS, collected in three international centers, we selected 51 biopsies and 32 autopsies that contained actively demyelinating lesions defined by stringent criteria. The pathology of the lesions was analyzed using a broad spectrum of immunological and neurobiological markers. Four fundamentally different patterns of demyelination were found, defined on the basis of myelin protein loss, the geography and extension of plaques, the patterns of oligodendrocyte destruction, and the immunopathological evidence of complement activation. Two patterns (I and II) showed close similarities to T-cell-mediated or T-cell plus antibody-mediated autoimmune encephalomyelitis, respectively. The other patterns (III and IV) were highly suggestive of a primary oligodendrocyte dystrophy, reminiscent of virus- or toxin-induced demyelination rather than autoimmunity. At a given time point of the disease--as reflected in autopsy cases--the patterns of demyelination were heterogeneous between patients, but were homogenous within multiple active lesions from the same patient. This pathogenetic heterogeneity of plaques from different MS patients may have fundamental implications for the diagnosis and therapy of this disease. PMID- 10852537 TI - Regional brain atrophy in progressive supranuclear palsy and Lewy body disease. AB - There have been no previous three-dimensional volumetric studies of regional brain atrophy in patients with pathologically confirmed progressive supranuclear palsy (PSP). Postmortem cortical and subcortical volumes were compared with neuropathology in 9 patients with PSP, 15 patients with Parkinson's disease, 10 patients with dementia with Lewy bodies, and 23 controls. Cases with the neuritic pathology of Alzheimer's disease were excluded. The topography of brain atrophy differed according to clinicopathological phenotype. Patients with Parkinson's disease had atrophy confined to the amygdala. Atrophy of the frontal lobe was found in both PSP and dementia with Lewy bodies and correlated with increasing neurofibrillary tangle or Lewy body densities, respectively. Patients with PSP could be differentiated by their marked atrophy of the internal globus pallidus. Further analysis of variance revealed that trends for greater frontal lobe atrophy correlated with clinical dementia in PSP, whereas both greater frontal and hippocampal atrophy and higher densities of Lewy bodies and Lewy neurites correlated with clinical dementia in cases with Lewy bodies. The present study provides evidence for selective regional atrophy that correlates with the underlying pathology of PSP and Lewy body disease. PMID- 10852538 TI - Neonatal seizures induced persistent changes in intrinsic properties of CA1 rat hippocampal cells. AB - We investigated the effects of repeated early-life seizures induced by flurothyl inhalation on intrinsic membrane properties of hippocampal pyramidal neurons from young rats (postnatal day 15-20). Intracellular recordings of CA1 and CA3 pyramidal neurons from flurothyl-treated and control rats revealed no significant differences in resting membrane potential, input resistance, membrane time constant, and action potential characteristics. In CA1 pyramidal cells from flurothyl-treated rats, the spike frequency adaptation and afterhyperpolarizing potential following a spike train were markedly reduced when compared with controls. In contrast, no significant alterations in the firing properties of CA3 pyramidal neurons were found. It is concluded that neonatal seizures lead to persistent changes in intrinsic membrane properties of CA1 pyramidal neurons. These alterations are consistent with an increase in neuronal excitability and may contribute to the behavioral deficit and epileptogenic predisposition observed in rats that experienced repeated neonatal seizures. PMID- 10852539 TI - Apolipoprotein E facilitates neuritic and cerebrovascular plaque formation in an Alzheimer's disease model. AB - The epsilon4 allele of apolipoprotein E (ApoE) is an important genetic risk factor for Alzheimer's disease (AD). Increasing evidence suggests that this association may be linked to the ability of ApoE to interact with the amyloid beta (Abeta) peptide and influence its concentration and structure. To determine the effect of ApoE on Abeta and other AD pathology in vivo, we used APPsw transgenic mice and ApoE knockout (-/-) mice to generate APPsw animals that carried two (ApoE +/+), one (ApoE +/-), or no copies (ApoE -/-) of the normal mouse ApoE gene. At 12 months of age, Abeta deposition was present in the cortex and hippocampus and was also prominent within leptomeningeal and cortical blood vessels of all APPsw ApoE +/+ mice. Importantly, although Abeta deposition still occurred in APPsw ApoE -/- mice, no fibrillar Abeta deposits were detected in the brain parenchyma or cerebrovasculature. There was also no neuritic degeneration associated with Abeta deposition in the absence of ApoE. These data demonstrate that ApoE facilitates the formation of both neuritic and cerebrovascular plaques, which are pathological hallmarks of AD and cerebral amyloid angiopathy. PMID- 10852540 TI - Hemorrhagic complications in vein of Galen malformations. AB - The authors report on a series of spontaneous intracranial hemorrhages associated with vein of Galen aneurysmal malformations (VGAMs). Thirty-four children with VGAMs have been treated at this institution since 1986. Eight children (24%) harbored the mural-type malformation, and 26 (76%) had the choroidal-type lesion. Two children (25%) with mural lesions and 1 (4%) with a choroidal lesion suffered hemorrhagic complications. Two presented with acute intracranial hemorrhage. A third child developed acute intracranial hemorrhage due to delayed dural sinus thrombosis after endovascular treatment of his choroidal-type VGAM. The subjects ranged in age from 13 days to 17 months at the time of presentation. Each patient underwent rapid radiological evaluation and treatment with endovascular surgery. Post-procedural arteriography demonstrated complete occlusion of the malformation in each patient. For the 3 patients with hemorrhage, follow-up has taken place over 49-, 107-, and 43-month intervals, respectively. Vein of Galen aneurysmal malformations can present with acute intracranial hemorrhage or develop delayed intracranial hemorrhage but respond to treatment using standard endovascular techniques. The presence of hemorrhage does not de facto portend a poor prognosis. PMID- 10852541 TI - Fukutin protein is expressed in neurons of the normal developing human brain but is reduced in Fukuyama-type congenital muscular dystrophy brain. AB - Fukuyama-type congenital muscular dystrophy (FCMD) results from a mutation in a gene on chromosome 9q31, fukutin, and is characterized pathologically by micropolygyria of the cerebral and cerebellar cortices. To elucidate the physiological function of fukutin as well as its pathological role in FCMD, we raised antisera against fukutin protein and observed its expression in developing human brains with or without FCMD. Western blotting using these antibodies demonstrated a 60-kd band in the fetal but not in postnatal cerebral cortex of the controls. This band appeared negligible in the brains of FCMD fetuses. Immunohistochemistry revealed the localization of fukutin in Cajal-Retzius cells, the subpial granular layer, the neuropil of the marginal zone, the cortical plate neurons, and the ventricular neuroepithelium of the fetal cerebrum. In the fetal cerebellum, fukutin immunoreactivity was localized to the external granule cell layer, molecular layer, Purkinje cells, and some internal granular cells. The immunoreactivity in these structures was reduced markedly in postnatal normal brains, as well as in an FCMD cerebrum at 23 gestational weeks. The spatial and temporal pattern of fukutin expression is compatible with its predicted role: the regulation of neuronal migration in the fetal cerebrum and cerebellum. PMID- 10852542 TI - Passive transfer of demyelination by serum or IgG from chronic inflammatory demyelinating polyneuropathy patients. AB - Chronic inflammatory demyelinating polyneuropathy (CIDP) is regarded as an autoimmune disorder, but no clearly defined autoimmune mechanism has been described. Although most patients respond to plasma exchange, no convincing role for autoantibodies has yet been demonstrated. In this study, we have successfully passively transferred disease using sera and purified IgG from 4 of 12 patients responsive to plasma exchange by bypassing the blood-nerve barrier by intraneural injection or opening it by activated T cells. The sera from CIDP patients or purified IgG produced marked conduction block and demyelination, but normal sera or IgG or that from patients with multiple sclerosis or other neuropathies did not. These observations strongly support an important role for anti myelin/Schwann cell autoantibodies in the pathogenesis of CIDP at least in some patients. PMID- 10852543 TI - Clinical and biochemical characteristics of congenital disorder of glycosylation type Ic, the first recognized endoplasmic reticulum defect in N-glycan synthesis. AB - We report on 8 patients with a recently described novel subtype of congenital disorder of glycosylation type Ic (CDG-Ic). Their clinical presentation was mainly neurological with developmental retardation, muscular hypotonia, and epilepsy. Several symptoms commonly seen in CDG-Ia such as inverted nipples, abnormal fat distribution, and cerebellar hypoplasia were not observed. The clinical course is milder overall, with a better neurological outcome, than in CDG-Ia. The isoelectric focusing pattern of serum transferrin in CDG-Ia and CDG Ic is indistinguishable. Interestingly, beta-trace protein in cerebrospinal fluid derived from immunoblot analysis of the brain showed a less pronounced hypoglycosylation pattern in CDG-Ic patients than in CDG-Ia patients. Analysis of lipid-linked oligosaccharides revealed an accumulation of Man9GlcNAc2 intermediates due to dolichol pyrophosphate-Man9GlcNAc2 alpha-1,3 glucosyltransferase deficiency. All patients were homozygous for an A333V mutation. PMID- 10852544 TI - Mice overexpressing rat heat shock protein 70 are protected against cerebral infarction. AB - Increased expression of heat shock protein 70 (HSP70) in the brain has been extensively documented in association with a variety of insults, including ischemia, and is suggested to play a role in cell survival and recovery after ischemic injury. To more directly assess the protective role of HSP70 during ischemic brain damage, we used transgenic mice overexpressing the rat HSP70 (HSP70tg mice). In contrast to wild-type (wt) littermates, high levels of HSP70 messenger RNA and protein were detected in brains of HSP70tg mice under normal conditions, and immunohistochemical analysis revealed primarily neuronal expression of HSP70. Heterozygous HSP70tg mice and their wt littermates were subjected to permanent focal cerebral ischemia by intraluminal blockade of the middle cerebral artery. Cerebral infarction after 6 hours of ischemia, as evaluated by Nissl staining, was significantly less in HSP70tg mice compared with wt mice. This reduction in infarction volume in HSP70tg mice was not attributable to an altered cardiovascular anatomy or to initial differences in body temperature or hemodynamic parameters. The HSP70tg mice were still protected against cerebral infarction 24 hours after permanent focal ischemia. The data suggest that HSP70 can markedly protect the brain against ischemic damage and that approaches aimed at inducing HSP70 may lead to new therapeutic interventions in cerebrovascular injuries. PMID- 10852545 TI - Mitochondrial neurogastrointestinal encephalomyopathy: an autosomal recessive disorder due to thymidine phosphorylase mutations. AB - Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is an autosomal recessive disorder defined clinically by severe gastrointestinal dysmotility; cachexia; ptosis, ophthalmoparesis, or both; peripheral neuropathy; leukoencephalopathy; and mitochondrial abnormalities. The disease is caused by mutations in the thymidine phosphorylase (TP) gene. TP protein catalyzes phosphorolysis of thymidine to thymine and deoxyribose 1-phosphate. We identified 21 probands (35 patients) who fulfilled our clinical criteria for MNGIE. MNGIE has clinically homogeneous features but varies in age at onset and rate of progression. Gastrointestinal dysmotility is the most prominent manifestation, with recurrent diarrhea, borborygmi, and intestinal pseudo-obstruction. Patients usually die in early adulthood (mean, 37.6 years; range, 26-58 years). Cerebral leukodystrophy is characteristic. Mitochondrial DNA (mtDNA) has depletion, multiple deletions, or both. We have identified 16 TP mutations. Homozygous or compound heterozygous mutations were present in all patients tested. Leukocyte TP activity was reduced drastically in all patients tested, 0.009 +/- 0.021 micromol/hr/mg (mean +/- SD; n = 16), compared with controls, 0.67 +/- 0.21 micromol/hr/mg (n = 19). MNGIE is a recognizable clinical syndrome caused by mutations in thymidine phosphorylase. Severe reduction of TP activity in leukocytes is diagnostic. Altered mitochondrial nucleoside and nucleotide pools may impair mtDNA replication, repair, or both. PMID- 10852546 TI - Decreased cerebrospinal fluid levels of beta-phenylethylamine in patients with Rett syndrome. AB - To clarify the mechanism of brain impairment in Rett syndrome, we measured the cerebrospinal fluid levels of beta-phenylethylamine (PEA) in 17 patients with Rett syndrome. Findings were compared with those obtained in age-matched controls and diseased controls. The cerebrospinal fluid level of PEA was significantly lower in patients with Rett syndrome than in the controls (31% of control values). The alteration in the cerebrospinal fluid level of PEA may reflect dopamine system impairment in Rett syndrome. PMID- 10852547 TI - Progression in Parkinson's disease: a positron emission tomography study with a dopamine transporter ligand [18F]CFT. AB - We studied the rate of progression of striatal dopamine transporter function in Parkinson's disease (PD). Eight patients with early PD without antiparkinsonian medication and 7 healthy volunteers were investigated with [18F]CFT positron emission tomography (PET). The PET scan was carried out twice at an approximate 2 year interval. The uptake of [18F]CFT was calculated as a region cerebellum:cerebellum ratio at 180 to 210 minutes after injection. At the first PET scan, the [18F]CFT uptake in PD patients in the putamen was 1.45 +/- 0.45 (mean +/- SD) (42% of the control mean) and 2.43 +/- 0.59 in the caudate nucleus (76% of the control mean). The ratios declined by the time of the second PET scan, and the rate of annual decline of the baseline mean in PD patients was 13.1% in the putamen and 12.5% in the caudate nucleus. In controls, the corresponding figures were 2.1% for the putamen and 2.9% for the caudate nucleus. The decline in [18F]CFT uptake was significantly higher in PD patients than in controls. Thus, dopamine transporter ligands such as [18F]CFT seem to be sensitive markers for the rate of progression in PD. PMID- 10852548 TI - Myelin widenings and MGUS-IgA: an immunoelectron microscopic study. AB - A few studies have reported a variety of nonspecific histological lesions in patients with IgA monoclonal gammopathies and polyneuropathy. In our case, using electron microscopy, we observed widenings of the myelin lamellae identical to those commonly described in IgM neuropathies with anti-myelin-associated glycoprotein activity. Using immunoelectron microscopy, we demonstrated a direct involvement of IgA in myelin lesions. The search for a direct link between monoclonal dysglobulinemia, regardless of type, and polyneuropathy is important and may influence treatment. PMID- 10852549 TI - Partial laminin alpha2 chain deficiency in a patient with myopathy resembling inclusion body myositis. AB - It is becoming evident that clinical phenotypes associated with partial laminin alpha2 chain deficiency are variable. We recently observed a 29-year-old man with leukoencephalopathy and vacuolar myopathy resembling inclusion body myositis. Laminin alpha2 immunohistochemical analysis showed reduction of the protein on muscle fiber surfaces. Molecular analysis revealed two novel compound heterozygous mutations in the LAMA2 gene. This is the first report linking a mutation in the LaMA2 gene with leukoencephalopathy and inclusion body-like myositis. PMID- 10852550 TI - Rapid clearance of human immunodeficiency virus type 1 from ventricular cerebrospinal fluid during antiretroviral treatment. AB - To understand the pathogenesis of human immunodeficiency virus-induced neuropathology, it is critical to know the dynamics of viral replication in the central nervous system. Viral decay kinetics were mathematically analyzed from multiple serial specimens of ventricular cerebrospinal fluid and plasma during antiretroviral therapy in a patient with asymptomatic human immunodeficiency virus infection and an external ventricular catheter for hydrocephalus. A rapid exponential decay of virus with an elimination half-life of 4.2 days in ventricular cerebrospinal fluid and 2.3 days in plasma was found. Sequencing the V3 loop-encoding envelope gene of virus in both compartments revealed high sequence homology. The combined data suggest that virus in ventricular cerebrospinal fluid is at least partly contributed by rapidly replicating virus producing cells recruited from the circulation. PMID- 10852551 TI - A common mechanism for the control of eye and head movements in humans. AB - How the human brain controls the subtle coupling between eye and head movements is still debated. The brain could either coordinate two separate (eye and head) networks or use a single system involved in gaze (eye + head) control. In a recent report, a total transfer from eye to head movements was observed in a patient with congenital ophthalmoplegia. This led the authors to hypothesize that such transfer resulted from a long-term adaptation between oculomotor and head movement systems. We report on a patient in whom a similar transfer was observed but at the acute stage of an acquired ophthalmoplegia. This case demonstrates that the transfer between head and eye movements does not necessarily require long-term adaptation and supports the hypothesis of a common unique gaze motor command in which eye and head movements would be rapidly exchangeable. PMID- 10852552 TI - A novel mutation of KCNQ3 (c.925T-->C) in a Japanese family with benign familial neonatal convulsions. AB - At present, only one mutation of KCNQ3, a KCNQ potassium channel gene, has been identified as a cause of benign familial neonatal convulsions type 2 (BFNC2). We found a T to C substitution (c.925T-C) on one allele of affected individuals in a Japanese family with BFNC but not on 200 alleles from healthy subjects. c.925T- >C replaced Trp309, a conserved residue within the P-loop of the KCNQ potassium channel family that holds the channel pore open, with an Arg (W309R). We report c.925T-->C as the second mutation of KCNQ3 responsible for BFNC2. PMID- 10852553 TI - Changes of copper-transporting proteins and ceruloplasmin in the lentiform nuclei in primary adult-onset dystonia. AB - A recent study reported an increase of brain tissue copper content in the lentiform nuclei of patients with primary adult-onset dystonia. In this study we analyze copper-metabolizing proteins (Menkes protein, Wilson protein, ceruloplasmin) by Western blot analysis in frozen brain tissue (lentiform nuclei) of 3 patients with primary dystonia. Menkes protein was reduced in all patients, while Wilson protein and ceruloplasmin were increased in the 2 patients with focal dystonia and reduced in the patient with generalized dystonia. Our data provides further evidence for a disturbance of copper metabolism in primary dystonia. PMID- 10852554 TI - Diagnostic criteria for primary progressive multiple sclerosis: a position paper. AB - The unique clinical characteristics of primary progressive multiple sclerosis (PPMS) pose particular diagnostic difficulties, both in excluding other causes of progressive syndromes and in confirming the diagnosis of MS, which is not adequately addressed by current diagnostic criteria. This article presents new diagnostic criteria developed by a group of investigators on the basis of a review of their considerable experience with PPMS. (We conclude that at least 1 year of clinical progression must be documented before a diagnosis of PPMS is made.) Three levels of diagnostic certainty have been defined-definite, probable, and possible--based on clinical findings, abnormal cerebrospinal fluid, abnormalities on magnetic resonance imaging (MRI) of the brain and spinal cord, and evoked potentials. In definite PPMS, evidence of intrathecal synthesis of immunoglobulin G together with one of the following three MRI criteria is required: (1) nine brain lesions, (2) two spinal cord lesions, or (3) four to eight brain lesions and one spinal cord lesion. Preliminary testing of these criteria was carried out on a cohort of 156 patients participating in a European natural history study of PPMS: 64% fulfilled the criteria for definite PPMS, 35% for probable PPMS, and only 1% for possible PPMS. These criteria now require prospective validation in a cohort of newly diagnosed patients and by postmortem examination. PMID- 10852555 TI - Spanish families with cerebral cavernous angioma do not bear 742C-->T Hispanic American mutation of the KRIT1 gene. PMID- 10852556 TI - Rapid diagnosis of peroxisome biogenesis disorders through immunofluorescence staining of buccal smears. PMID- 10852557 TI - Topiramate and essential tremor. PMID- 10852558 TI - Magnetic resonance spectroscopy of episodic ataxia type 2 and migraine. PMID- 10852559 TI - Clinical heterogeneity in pedigrees with 2q-linked febrile seizures. PMID- 10852560 TI - Locus for febrile seizures. PMID- 10852561 TI - Reduction of Escherichia coli O157:H7 counts on whole fresh apples by treatment with sanitizers. AB - The objectives of this study were to determine if washing of whole apples with solutions of three different sanitizers (peroxyacetic acid, chlorine dioxide, or a chlorine-phosphate buffer solution) could reduce a contaminating nonpathogenic Escherichia coli O157:H7 population by 5 logs and at what sanitizer concentration and wash time such a reduction could be achieved. Sanitizers were tested at 1, 2, 4, 8, and 16 times the manufacturer's recommended concentration at wash times of 5, 10, and 15 min. Whole, sound Braeburn apples were inoculated with approximately 1 x 108 or 7 x 106 CFU per apple, stored for 24 h, then washed with sterile water (control) or with sanitizers for the prescribed time. Recovered bacteria were enumerated on trypticase soy agar. Washing with water alone reduced the recoverable population by almost 2 logs from the starting population; this can be attributed to physical removal of organisms from the apple surface. No sanitizer, when used at the recommended concentration, reduced the recovered E. coli population by 5 logs under the test conditions. The most effective sanitizer, peroxyacetic acid, achieved a 5-log reduction when used at 2.1 to 14 times its recommended concentration, depending on the length of the wash time. The chlorine-phosphate buffer solution reduced the population by 5 logs when used at 3 to 15 times its recommended concentration, depending on wash time. At no concentration or wash time tested did chlorine dioxide achieve the 5-log reduction. PMID- 10852562 TI - Effects of antibiotic regimens on the fecal shedding patterns of pigs infected with salmonella typhimurium. AB - An experiment was conducted to determine (i) the effects of antibiotic regimens on the shedding patterns of pigs infected with Salmonella Typhimurium and (ii) whether antibiotic resistance increases the incidence of pathogen shedding. The experiment involved 48 50-day-old pigs challenged with Salmonella Typhimurium and receiving one of four antibiotic regimens including (i) intramuscular injection of ceftiofur sodium followed by inclusion of oxytetracycline in the feed; (ii) apramycin in the feed for 14 days followed by oxytetracycline; (iii) carbadox in the feed until pigs reached 35 kg followed by oxytetracycline; (iv) no antibiotics (control). Fecal samples were collected preinoculation, 2 and 4 days postinoculation (DPI) and at weekly and biweekly intervals thereafter to determine shedding patterns. Salmonella Typhimurium isolates from 2, 4, 7, 21, 42, and 70 DPI were analyzed for antibiotic resistance. A time effect (P < 0.05) was observed, indicating that the proportion of isolates resistant to at least one antibiotic varied over time. Overall resistance was determined to be 46% at 2 DPI and increased significantly (P < .05) thereafter. Treatment x time and antibiotic x time interactions were also observed (P < 0.05) as the percentage of isolates resistant to each test antibiotic increased over time. In no case did the development of antibiotic resistance result in an increased incidence of shedding of the original inoculate. The incidence of shedding was reduced in pigs receiving the apramycin-oxytetracycline treatment, when compared to control pigs; however, no differences were observed between antibiotic treatments. PMID- 10852563 TI - Survival of listeria innocua in salmon following cold-smoke application. AB - The ability of Listeria innocua to survive on salmon fillets during cold smoking in a commercial processing plant was investigated using a central composite rotatable response surface design to examine smoking temperatures in the range of 18 to 30 degrees C and a smoke time from 2 to 14 h. Smoke temperature did not significantly (P < 0.05) reduce counts of L. innocua on the salmon. However, the smoking time had a significant effect on L. innocua. The smoking time was directly related to the reduction in count (R2 = 0.831), and a 3-log cycle reduction in count was observed when the smoking time was 12 h. The reduction in L. innocua levels on the fish was unaffected by the pH, water activity, and salt concentration of the fillet. PMID- 10852564 TI - Application of enterocins as biopreservatives against Listeria innocua in meat products. AB - The antilisterial effect of enterocins A and B in meat and meat products (cooked ham, minced pork meat, deboned chicken breasts, pate, and slightly fermented sausages [espetec]) have been shown. An infective dose of 5 to 10 most probable numbers (MPN)/g to simulate the counts of Listeria generally found in meat products was used. Enterocins at 4,800 AU/g reduced the numbers of Listeria innocua by 7.98 log cycles in cooked ham and by 9 log cycles in pate when stored at 7 degrees C for 37 days. In deboned chicken breasts stored at 70 degrees C for 7 days, 4,800 AU/cm2 of enterocins diminished the L. innocua counts in 5.26 log cycles when compared to the control batch. In minced pork meat held at 7 degrees C for up to 6 days, 1,600 AU/g kept L. innocua counts under 3 MPN/g, while the control batch reached 50 CFU/g. In espetec sausages, 648 AU/g diminished the number of L. innocua under 50 CFU/g from the fifth day until the end of the process (12 days) while the control batch kept the initial counts (3 x 104 CFU/g). This is the first report on enterocins showing an antilisterial effect in different types of meat products. PMID- 10852565 TI - Antibacterial mechanism of allyl isothiocyanate. AB - Allyl isothiocyanate (AITC), a natural compound in plants belonging to the family Cruciferae, has been shown to have strong antimicrobial activity in liquid media as well as in its vapor form. To understand its antimicrobial mechanism, AITC was tested for bactericidal activities to Salmonella Montevideo, Escherichia coli O157:H7, and Listeria monocytogenes Scott A at different stages of growth and was compared with streptomycin, penicillin G, and polymyxin B, each of known antibacterial mechanisms. Bactericidal activities were determined by measuring bacterial viability and leakage of metabolites. To determine its effects on membrane permeability, beta-galactosidase activity was examined after exposure of E. coli K-12 strain 3.300 to the three antibiotics and to AITC. The two gram negative bacteria, Salmonella Montevideo and E. coli O157:H7, were more sensitive to AITC and to polymyxin B than the gram-positive L. monocytogenes. AITC and polymyxin B were effective bactericidal agents to bacteria at all growth stages, whereas penicillin G and streptomycin did not exhibit bactericidal activity to stationary cells. High A260 and A280 values of cellular filtrate and beta galactosidase activity were obtained after treatments of AITC and polymyxin B. These data indicated that AITC was most similar to polymyxin B with respect to its antibacterial effect on cell membranes and on leakage of cellular metabolites. Gaseous AITC caused metabolite leakages, measurable increases in 3 galactosidase activity, and reduction of viable bacteria. The effectiveness of AITC in inhibiting bacteria at all growth stages and its strong activity in vapor phase support its application in food preservation. PMID- 10852566 TI - Combined effect of hop resins and sodium hexametaphosphate against certain strains of Escherichia coli. AB - The combined antimicrobial effects of hop resins with sodium hexametaphosphate, glycerol monocaprate, and lysozyme were investigated aiming to make an effective agent against Escherichia coli. When they are used separately, the antimicrobial activity against E. coli was minimal. However, the combination of hop resins with sodium hexametaphosphate exhibited strong antimicrobial activity against E. coli, but no effect was found in combinations of hop resins with the other agents. The activity was strongest when the combination was added at the beginning of growth of the bacteria, resulting in a prolonged lag phase. However, when the antimicrobials were added during the log phase, growth was depressed considerably. By addition of these materials, cell components with absorbance near 260 nm were leaked out. This possibly may have resulted from damage to the cell membranes of the bacteria. The combined effect was also detected in model food systems such as mashed potatos. The use of hop resins and sodium hexametaphosphate in combination may thus be useful for controlling E. coli. PMID- 10852567 TI - Combined effect of nisin and pulsed electric fields on the inactivation of Escherichia coli. AB - The Doehlert design was applied in order to investigate the combined effect of nisin and high voltage pulsed electric fields (PEF) on the inactivation of Escherichia coli in simulated milk ultrafiltrate media. Nisin alone was totally inactivated by PEF, but in the presence of bacterial cells a protective effect was observed. However, the effectiveness of nisin was still decreased when bacterial cells were subjected to the combined treatment. In spite of this phenomenon, an almost additive response emerged as a consequence of the combined treatment. A 4-log cycle reduction may be accomplished with around 1,000 IU/ml (7.15 microM) of nisin and three pulses of 11.25 kV/cm or 500 IU/ml for five pulses of the same intensity. The observed efficacy arising from the combination of both treatments suggests the possibility of using PEF for improving the action spectrum of natural antimicrobials. PMID- 10852568 TI - Antibacterial activity of a chitooligosaccharide mixture prepared by cellulase digestion of shrimp chitosan and its application to milk preservation. AB - The antibacterial activity of a chitooligosaccharide mixture prepared by digestion of shrimp chitosan with cellulase at 50 degrees C for 14 h was evaluated. Sugars with 1 to 8 degrees of polymer (DP) were found in this chitooligosaccharide mixture, and the weight percentage of sugars with DP > or = 6 was 44.3%. Minimal lethal concentrations of this mixture against Aeromonas hydrophila, Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella Typhimurium, Shigella dysenteriae, Staphylococcus aureus, Vibrio cholerae, and Vibrio parahaemolyticus in nutrient broth were 5 to 29 ppm, which were much lower than those of the chitosan reactant (50 to 1,000 ppm). The antibacterial activity of this mixture in the sterilized milk against E. coli O157, L. monocytogenes, Salmonella Typhimurium, and S. aureus was much stronger at 4 degrees C than at 37 degrees C. When raw milk was supplemented with either 0.24% or 0.48% (wt/vol) of this oligosaccharide mixture and stored at 4 degrees C for 12 days, its mesophilic and psychrotrophic counts were reduced by at least 3 log cycles, and there was very little change in pH. In addition, this mixture retarded the growth of Salmonella species and caused quicker reduction of Staphylococcus species in raw milk. Accordingly, the shelf life of raw milk at 4 degrees C was extended by at least 4 days. PMID- 10852569 TI - Inactivation of Bacillus subtilis spores on aluminum and polyethylene preformed cartons by UV-excimer laser irradiation. AB - The efficacy of UV KrF-excimer laser light (at 248 nm) to inactivate Bacillus subtilis spores loaded onto preformed cartons was found to be dependent on the interior carton coating and scheme by which the irradiation was applied. When the carton was held static during UV laser treatment, the majority of the dose was delivered to the base of the carton and to a lesser extent to the upper part of the pack. In this arrangement no irradiation of the interior sides of the carton was observed. A more even distribution of dose was achieved, however, by moving the carton within the laser beam during irradiation treatment. The distribution of UV was also found to be dependent on the type of carton interior coating. With aluminum cartons the dose measured was found to be significantly greater (P < 0.01) and more evenly distributed across the interior compared to when polyethylene packs were tested. Under optimized conditions no spore survivors were detected on aluminum cartons preloaded with 9.5 x l0 B. subtilis spores by applying a UV laser output dose of 160 J. In comparison, the same conditions only achieved a significantly lower (P < 0.01) reduction in spore numbers (log count reduction 4.2) when polyethylene cartons were used. This difference in lethality and UV distribution of laser light was associated with the higher internal reflection of photons with aluminum cartons. The suitability of UV-excimer lasers for sterilizing preformed cartons over traditional germicidal lamp-based methods is discussed. PMID- 10852570 TI - Effect of carbon dioxide under high pressure on the survival of cheese starter cultures. AB - A new processing method that rapidly forms curds and whey from milk has the potential to improve cheesemaking procedures if cheese starter cultures can tolerate the processing conditions. The survival of Lactobacillus delbrueckii ssp. bulgaricus, Lactococcus lactis ssp. lactis, or Streptococcus thermophilus through this new process was evaluated. Inoculated milk containing 0, 1, or 3.25% fat or Lactobacillus MRS broth or tryptone yeast lactose broth (depending on microorganism used) was sparged with CO2 to a pressure of 5.52 MPa and held for 5 min at 38 degrees C. Broth contained 7.93 to 8.78 log CFU/ ml before processing and 7.84 to 8.66 log CFU/ml afterward. Before processing, milk inoculated with L bulgaricus, L. lactis, or S. thermophilus contained 6.81, 7.35, or 6.75 log CFU/ml, respectively. After processing, the curds contained 5.68, 7.32, or 6.50 log CFU/g, and the whey had 5.05, 6.43, or 6.14 log CFU/ml, respectively. After processing, the pHs of control samples were lower by 0.41 units in broth, 0.53 units in whey, and 0.89 units in curd. The pH of the processed inoculated samples decreased by 0.3 to 0.53 units in broth, 0.32 to 0.37 units in whey, and 0.93 to 0.98 units in the curd. Storing curds containing L. lactis at 30 degrees C or control curds and curds with L. bulgaricus or S. thermophilus at 37 degrees C for an additional 48 h resulted in pHs of 5.22, 5.41, 4.53, or 4.99, respectively. This study showed that milk inoculated with cheese starter cultures and treated with CO2 under high pressure to precipitate casein-produced curds that contained sufficient numbers of viable starter culture to produce lactic acid, thereby decreasing the pH. PMID- 10852571 TI - Chlordane and toxaphene residues following cooking of treated channel catfish fillets. AB - The reduction in residues of chlordane and toxaphene following cooking (frying, baking, and smoking) of fillets obtained from treated Channel catfish (Ictalurus punctatus) was determined. On average, cooking reduced moisture content by 17% and increased fat content by 28 to 274%. Frying reduced chlordane residues by 56 to 86% on a dry basis (db) or 84 to 92% on a percent fat basis (fb) when raw fillets were compared to cooked fillets. Baking and smoking reduced chlordane significantly less (P < 0.05) than frying with reductions in residues of 12% and 9% (db) or 30% and 33% (fb), respectively. Frying reduced toxaphene residues by 40 to 49% (db) or 65 to 77% (fb), while baking and smoking reduced toxaphene by 35% and 24% (db) or 51% and 59% (fb), respectively. PMID- 10852572 TI - Cost effectiveness of vaccinating food service workers against hepatitis A infection. AB - Foodborne transmission is an important means of hepatitis A infection that may be reduced through vaccination of food service workers (FSWs). Several states are considering actions to encourage or mandate FSW vaccination, but the cost effectiveness of such policies has not been assessed. We estimated the clinical and economic consequences of vaccinating FSWs from the 10 states with the highest reported rates of hepatitis A. A decision analytic model was used to predict the effects of vaccinating FSWs at age 20 years. It was assumed all FSWs would receive one dose of inactivated hepatitis A vaccine, and 50% would receive the second recommended dose. Parameter estimates were obtained from published reports and Centers for Disease Control and Prevention databases. The primary endpoint was cost per year of life saved (YOLS). Secondary endpoints were symptomatic infections, days of illness, deaths, and costs of hepatitis A treatment, public health intervention, and work loss. Each endpoint was considered separately for FSWs and patrons. We estimate vaccination of 100,000 FSWs would cost $8.1 million but reduce the costs of hepatitis A treatment, public health intervention, and work loss by $3.0 million, $2.3 million, and $3.1 million, respectively. Vaccination would prevent approximately 2,500 symptomatic infections, 93,000 days of illness, and 8 deaths. A vaccination policy would reduce societal costs while costing the health system $13,969 per YOLS, a ratio that exceeds generally accepted standards of cost effectiveness. PMID- 10852573 TI - Isolation and enumeration of Giardia cysts, cryptosporidium oocysts, and Ascaris eggs from fruits and vegetables. AB - Published techniques for recovering parasites from fruit and vegetables are generally inadequate, with low and variable recovery efficiencies. Here we describe an improved methodology for analyzing fruit and vegetables for Giardia cysts, Cryptosporidium oocysts, and Ascaris eggs. The method includes washing procedures, sonication, and, for Giardia and Cryptosporidium, immunomagnetic separation. Identification is by immunofluorescence (Giardia and Cryptosporidium) or brightfield microscopy (Ascaris). Recovery efficiencies from lettuce, Chinese leaves, and strawberries were found to be approximately 67% for Giardia, 42% for Cryptosporidium, and 72% for Ascaris. Recovery efficiencies from bean sprouts tended to be more variable and lower. This could be due to material removed with the parasites during the washing procedures, which, in turn, appeared related to the age of the bean sprouts. It is therefore recommended that fruit and vegetables should be as fresh as possible when analyzed for parasites. PMID- 10852574 TI - Review of studies on the thermal resistance of Salmonellae. AB - Heat resistance data for different serotypes of Salmonella enterica in different food products and laboratory media are reviewed. From all D-values reported, the highest heat resistance of Salmonella was in liquid eggs and liquid egg yolks. The equation from a line drawn through the highest D-values, and above all values reported, was log D-value = 11.7 - 0.188T degrees C. From this equation, the calculated z-value was 5.3 degrees C (9.5 degrees F), and a process at 71degrees C (160 degrees F) will require 1.2 s to inactivate 1 log of Salmonella cells. This calculation did not include data that evaluated the heat resistance after stress conditions or data for Salmonella Senftenberg. The heat resistance of Salmonella is highly influenced by the strain tested, the type of experiment (log reduction versus end-point), culture conditions prior to the experiment, heating menstruum, and recovery conditions. Heat resistance data for Salmonella are still nonexistent or scarce in chicken meat, fruit juices, and aquacultured fish. PMID- 10852575 TI - Review of centralized packaging systems for distribution of retail-ready meat. AB - There is growing interest in centralized preparation of retail-ready meat cuts for distribution to widely dispersed retail stores due to the convenience of having high-quality ready-to-go products that are consistently provided to consumers at lower cost. Various centralized packaging techniques are described. Of all packaging techniques, master packaging is the most economical and shows promise for commercial application. Nevertheless, the master-packaging technique must be integrated with strict temperature control in a narrow range just above freezing (- 1.5 +/- 0.5 degrees C), good processing hygiene, and maintenance of a completely anoxic atmosphere in the package headspace throughout the distribution period to maximize storage life. Packaging using the CAPTECH process reduces the residual O2 present in the headspace to 300 ppm. Oxygen scavengers must be incorporated in the package to absorb the residual O2 and preserve the metmyoglobin reducing activity of meat tissues. Integration of all these technologies can provide a storage life of retail-ready meat up to 10 weeks in the master package followed by 3 days of retail display life. This extension of storage life is sufficient for transporting meat to distant markets. PMID- 10852576 TI - Incidence of foodborne illnesses reported by the foodborne diseases active surveillance network (FoodNet)-1997. FoodNet Working Group. AB - In 1997, the Foodborne Diseases Active Surveillance Program (FoodNet) conducted active surveillance for culture-confirmed cases of Campylobacter, Escherichia coli O157, Listeria, Salmonella, Shigella, Vibrio, Yersinia, Cyclospora, and Cryptosporidium in five Emerging Infections Program sites. FoodNet is a collaborative effort of the Centers for Disease Control and Prevention's National Center for Infectious Diseases, the United States Department of Agriculture's Food Safety and Inspection Service, the Food and Drug Administration's Center for Food Safety and Applied Nutrition, and state health departments in California, Connecticut, Georgia, Minnesota, and Oregon. The population under active surveillance for foodborne infections was approximately 16.1 million persons or roughly 6% of the United States Population. Through weekly or monthly contact with all clinical laboratories in these sites, 8,576 total isolations were recorded: 2,205 cases of salmonellosis, 1,273 cases of shigellosis, 468 cases of cryptosporidiosis, 340 of E. coli O157:H7 infections, 139 of yersiniosis, 77 of listeriosis, 51 of Vibrio infections, and 49 of cyclosporiasis. Results from 1997 demonstrate that while there are regional and seasonal differences in reported incidence rates of certain bacterial and parasitic diseases, and that some pathogens showed a change in incidence from 1996, the overall incidence of illness caused by pathogens under surveillance was stable. More data over more years are needed to assess if observed variations in incidence reflect yearly fluctuations or true changes in the burden of foodborne illness. PMID- 10852577 TI - Use of microbial data for hazard analysis and critical control point verification -Food and Drug Administration perspective. AB - This paper examines the role that the microbiologist and microbiological testing play in implementing hazard analysis and critical control point (HACCP) programs. HACCP offers a more comprehensive and science-based alternative for controlling food safety hazards compared with traditional sanitation programs based upon good manufacturing practices. Controlling hazards under an HACCP program requires a systematic assemblage of reliable data relating to the occurrence, elimination, prevention, and reduction of hazards. These data need to be developed in a transparent environment that will ensure that the best scientific methodologies have been employed in developing the needed data. The two mechanisms used in HACCP to assess the adequacy of the database are validation studies and the verification assessments. Microbiological testing is an important mechanism for collecting data used in developing and implementing an HACCP plan. Microbial sample data can help establish standard operating procedures (SOPs) for sanitation, assess the likelihood of the occurrence of hazards, establish critical limits, and assess the validity of the HACCP plan. The use of a performance standard to assess whether microbiological hazards have been reduced to an acceptable level creates an especially important use for microbial analysis. Microbial testing is also useful in implementing an HACCP plan by helping to monitor the effectiveness of sanitation SOPs, the compliance of incoming ingredients with safety criteria, the safety of product being held for corrective action, and the safety of the finished product. The verification audits demonstrate that all control measures have been applied as designed in the HACCP plan. Although auditing HACCP records is the primary means of verification, microbial sampling can play an important role as well. PMID- 10852578 TI - Industry perspectives on the use of microbial data for hazard analysis and critical control point validation and verification. AB - Microbial testing is an essential element in validation of critical limits identified within a hazard analysis critical control point (HACCP) plan. Without appropriate validation there is no assurance that the plan will control the hazards of concern. Once critical control points have been validated to effectively prevent, reduce, or eliminate hazards, application of routine testing for pathogens in finished product becomes an ineffective means to assure process control and therefore safety of the product. The occurrence of a pathogen in a product produced under an effective HACCP plan is so rare that sampling protocols are not capable of finding the needle in the haystack. Quantitative indicators can provide a much more effective tool for verifying that HACCP is properly implemented. Choice of appropriate indicators is product and process specific. In certain applications, finished product testing for even indicator organisms provides no meaningful data for verification of HACCP (e.g., canned products). Tests chosen should provide meaningful information that directs resources toward prevention and improvement of the system. PMID- 10852579 TI - How does Escherichia coli O157:H7 testing in meat compare with what we are seeing clinically? AB - Escherichia coli O157:H7 is but one of a group of Shiga toxin-producing E. coli (STEC) that cause both intestinal disease such as bloody and nonbloody diarrhea and serious complications like hemolytic uremic syndrome (HUS). While E. coli O157: H7 is the most renowned STEC, over 200 different types of STEC have been documented in meat and animals, at least 60 of which have been linked with human disease. A number of studies have suggested that non-O157 STEC are associated with clinical disease, and non-O157 STEC are present in the food supply. Non-O157 STEC, such as O111 have caused large outbreaks and HUS in the United States and other countries. The current policy in the United States is to examine ground beef for O157:H7 only, but restricting the focus to O157 will miss other important human STEC pathogens. PMID- 10852580 TI - Clinical pathology of foodborne diseases: notes on the patient with foodborne gastrointestinal illness. AB - The symptoms and signs in persons with food- or waterborne infections provide clues to the nature of the infecting microbe. Proper treatment of the affected individual, and protection of those exposed to the same source, is dependent on time-honored methods of diagnosis: exposure history, and physical examination. Laboratory testing may help to identify the responsible agent. Spontaneous recovery is the most likely outcome once supportive measures such as fluid and electrolyte replacement are addressed. Antibiotics are often unnecessary and may prolong fecal excretion of certain microorganisms. In immunosuppressed persons or those weakened by marginal nutrition, foodborne infection can be more severe, mandating more specific therapy. Management requires knowing the level of tissue invasion and organ infected by each of the commonly encountered microbes. Some of the most life-threatening infections (cholera, for example) are associated with no visible tissue injury, yet they have a profound impact on gut function. In contrast, salmonellosis and shigellosis can cause severe gut injury, and when foodborne infections extend beyond the confines of the gut, skilled care is essential. Examples are hemolytic uremic syndrome of Escherichia coli infections, or listeriosis, both of which require urgent attention. Long-term consequences of gut infections such as the paralytic Guillain-Barre syndrome following Campylobacter infections illustrates the long-term problems sometimes encountered. Because it is unlikely that all infectious agents will ever be removed from food and water in any country, sound medical intervention tailored to the extent of illness will be the mainstay of handling such illnesses. PMID- 10852581 TI - Epidemiology, microbiology, and risk assessment of waterborne pathogens including Cryptosporidium. AB - Cryptosporidium is one of a suite of relatively recently emerging pathogens of concern in drinking water. Based on human dose-response tests, guidelines for exposure yielding defined levels of endemic risk have been developed. This risk assessment procedure is grounded in the process used for chemical risk assessment. From outbreak data, critical concentrations in water that may lead to epidemic levels have been postulated. Development of these levels will be discussed. Validation of the information using outbreak reports from the 1993 Milwaukee incident can be made. Use of this approach must be tempered by the existence of substantial waterborne cases in the absence of detectable oocyst levels as in the Las Vegas outbreak, and (apparent) high levels of oocysts without (apparent) significant health effects as in the case of the (at the time of this writing) ongoing incident in Sydney, Australia. PMID- 10852582 TI - Acquisition of microbiological data to enhance food safety. AB - The routine acquisition and archiving of microbiological data is undertaken for two reasons. The first is the development of historical microbiological profiles of foods, ingredients, or processes in order to determine or verify that microorganisms of concern are being controlled to the level desired. The second reason is data concerning the pathogenicity or virulence of foodborne pathogens and their behavior in foods in order to develop strategies and criteria for assuring microbiological safety. Both types of microbiological data are essential to effective food safety programs. A firm understanding of the uses and limitations of both is essential to correct acquisition, interpretation, and use of such data. PMID- 10852583 TI - Gene therapy and transplantation. AB - Advances in molecular biology and in techniques of gene transfer have resulted in the development of practical approaches to human gene therapy. Many applications are of relevance to manipulation of the immune system and have potential in organ and cell transplantation. For example, gene therapy approaches may facilitate the induction of immunological tolerance to a donor organ or protect it locally against the host's immune response. Based on a comprehensive review of the world literature, examples of current research efforts in both allogeneic and xenogeneic transplantation are presented and discussed. PMID- 10852584 TI - Renal transplantation: single or dual for donors aging > or =60 years? PMID- 10852585 TI - Preconditioning versus brain death in clinical transplantation. PMID- 10852586 TI - The only good T cell is a dead T cell? PMID- 10852587 TI - Exploring the potential for graft vs. tumor response in a combine bone marrow and liver transplantation for large hepatocellular carcinoma. PMID- 10852588 TI - Effects of humanized monoclonal antibody to rhesus CD11a in rhesus monkey cardiac allograft recipients. AB - INTRODUCTION: Leukocyte function-associated antigen-1 (LFA-1, CD11a) monoclonal antibody (mAb) affects many leukocyte functions without cell depletion. We hypothesized that the use of a humanized, anti-rhesus modified LFA-1 mAb (H2C12) in rhesus monkeys would cause: (1) prolonged heart allograft survival, (2) inhibition of primary but not secondary antibody responses, and (3) minimal drug toxicity. METHODS AND RESULTS: Control (n=5) and H2C12-treated (n=7) (8-20 mg/kg i.v. on day -1 followed by 10 mg/kg/day) adult male rhesus recipients were inoculated with GP120 protein antigen on day -28 and -1 and grafted with heterotopic abdominal hearts (day 0). Donor-recipient pairs were equally MLR mismatched (4329.8+/-1124.1 CPM controls vs. 7289.0+/-1926.5 treated, P=NS). Mean heart allograft survival as evaluated by daily abdominal palpation was significantly prolonged in high dose recipients (23.0+/-2.6, n=4) vesus controls (8.2+/-1.3, n=5, P<0.02, Mann-Whitney U test). H2C12 treatment did not produce signs of cytokine release or toxicity, was nondepleting, but down-modulated PBL CD11a expression to 43.4+/-3.6% (n=4) of control levels (n=5) at day 7 as demonstrated by flow cytometry. It had no effect on postoperative Con A or MLR and did not prevent mAb clearance due to the rhesus-antihuman antibody response. The addition of mycophenolate mofitil prevented rhesus-antihuman antibody response with therapeutic H2C12 levels seen for >35 days. CONCLUSIONS: The use of this mAb to block CD11a had the benefit of being a well tolerated, highly targeted therapy. These are the first results showing that monotherapy with anti leukocyte function-associated antigen-1 mAb prolonged survival of MLR mismatched allogenic cardiac grafts in primates. PMID- 10852589 TI - Acute brain death abolishes the cardioprotective effects of ischemic preconditioning in the rabbit. AB - BACKGROUND: Myocardial preconditioning with brief coronary artery occlusions before a sustained ischemic period is reported to reduce infarct size. We wished to evaluate whether ischemic preconditioning (IP) is efficient in an experimental brain death (BD) model in the rabbit. METHODS: Rabbits were randomized into four experimental groups of eight animals each. In the control group (CTRL), anaesthetized rabbits were subjected to 30 min of left coronary marginal branch occlusion and 90 min of reperfusion without any pretreatment. In the CTRL+IP group, anaesthetized rabbits were preconditioned with a 3-min ischemia and 3-min reperfusion sequence before coronary occlusion. In the BD group, rabbits were subjected to 90 min of BD before 30 min of coronary occlusion and 90 min of reperfusion. In the BD+IP group, BD rabbits were preconditioned as in the CTRL+IP group before coronary occlusion. BD was induced by rapid inflation of an intracranial balloon and was validated by clinical and electroencephalographic examinations. At the termination of the experiment, left ventricular volume (LVV), myocardial volume at risk (VAR) and infarct volume (IV) were determined with methylene blue and tetrazolium staining and were measured using planimetry. RESULTS: LW was not significantly different among the four experimental groups (CTRL, 6.54+/-0.90 cm3; CTRL+IP, 5.92+/-0.60 cm3; BD, 5.87+/-0.81 cm3; BD+IP, 6.16+/-0.95 cm3; P=ns). Furthermore, myocardial VAR, expressed as a percentage of LVV, was not significantly different between groups (CTRL, 20.0+/-4.2%; CTRL+IP, 22.32+/-2.25%; BD, 21.38+/-3.36%; BD+IP, 21.64+/-3.39%; P=NS). IV, expressed as a percentage of VAR, was significantly reduced in the CTRL+IP group compared with the CTRL group (15.76+/-8.47% vs. 49.95+/-1.51%; P<0.0001). In contrast, there was no significant difference in IV, expressed as a percentage of VAR, between the BD and the BD+IP groups (50.0+/-1.52% vs. 49.72+/-1.58%; P=NS). CONCLUSION: The data indicate that the infarct-limiting effect of IP is lost in BD rabbits. Thus, the clinical potential of IP in the context of organ transplantation seems to be severely compromised. PMID- 10852590 TI - The effects of inhaled nitric oxide, gabexate mesilate, and retrograde flush in the lung graft from non-heart beating minipig donors. AB - BACKGROUND: The use of lung grafts from non-heart-beating donors (NHBD) is one way of solving the donor organ shortage problem. In this experiment, we studied the effect of retrograde flush (RF) from the left atrium before harvest, inhaled nitric oxide (NO), and gabexate mesilate (FOY), a protease inhibitor, in the lung grafts from NHBD. METHODS: Forty-eight Lee-Sung, small-ear, miniature pigs (15-20 kg) were divided into 24 pairs (donor and recipient) and four groups. The donor lungs were flushed and harvested 90 min after cardiac arrest. No i.v. heparin was administered until the time before flush and harvest. Left single lung transplantation was undertaken, and the recipients were observed for 18 hr. The grafts warm and cold ischemia times were 90 (controlled) and 183+/-23.4 min. Group 1 (untreated control, UC, n=6) had core perfusion through a Swan-Ganz catheter followed by a single, antegrade flush with modified Euro-Collin's solution containing heparin, urokinase, and PGE1. Group 2 (RF group, n=6) had the same as group 1, except that one additive retrograde flush through the left atrium was administered. Group 3 (NO group, n=6) had the same as group 1, except that 20 parts per million (ppm) inhaled NO was administered for the cadaver donors before the graft harvest, and for the recipients after the grafts reperfusion. Group 4 (FOY group, n=6) had the same as group 1, except that the recipients received FOY i.v. infusion from the beginning of the recipient's operation and continuously throughout the experiments. RESULTS: Compared with the group 1 (control), group 2 (RF) had significantly (P<0.05) lower mean pulmonary artery pressure, pulmonary vascular resistance (PVR), lung wet/dry ratio, histological lung injury score, and higher PaO2/FiO2 and pulmonary dynamic compliance. Group 3 (NO) had significantly lower mean pulmonary arterial pressure, PVR, lung injury score, degree of tissue neutrophils infiltration (histological and myeloperoxidase assay), bronchoalveolar lavage fluid protein content and neutrophils (PMNs) percentage, and higher PaO2/FiO2 and pulmonary dynamic compliance. Group 4 (FOY) had significantly lower PMNs infiltration, lung injury score, wet/dry ratio, bronchoalveolar lavage fluid protein and PMNs percentage, and higher PaO2/FiO2. Group 2 (RF) revealed better gas exchange (PaO2/FiO2) than the control (group 1) at earlier reperfusion periods (1st and 5th hr). On the contrary, group 4 (FOY) had higher PaO2/FiO2 than group 1 only at later period (18th hr). Pathologically, retrograde flush (group 2, RF) inhibited the intravascular thrombi formation more effectively than the NO or FOY treatment. However, the NO or FOY treatment inhibited the neutrophil infiltration more effectively than did the retrograde flush. CONCLUSION: The retrograde flush, inhaled NO and FOY infusion are beneficial to the protection of the NHBD lung grafts at an early reperfusion period, through different mechanisms. The use of these treatments in combination might help us to find a better way to protect the NHBD grafts against the preservation and reperfusion injury. PMID- 10852591 TI - Warm ischemia and reperfusion injury in diet-induced canine fatty livers. AB - BACKGROUND: Fatty liver is associated with primary nonfunction after liver transplantation, contributing a shortage of suitable liver grafts. Because extensive investigation of mechanisms underlying such nonfunction has been limited largely to rodents, we made a new fatty liver model in dogs and studied primary nonfunction after warm ischemia. METHODS: We developed a diet rich in fat but deficient in choline to induce fatty change in canine liver and investigated effects of 60 min of warm ischemia and reperfusion in dogs with such fatty livers. RESULTS: Microscopically evident steatosis increased with duration of dietary manipulation (up to 12 weeks), as did hepatic total lipid and triglyceride levels. No dog with >30% of steatotic hepatocytes, >445 mg/g hepatic total lipid or >145 mg/g hepatic triglyceride survived after 60 min of warm ischemia. Arterial ketone body ratios decreased and blood endotoxin increased after reperfusion in nonsurvivors. The main histologic finding in livers of nonsurvivors was marked sinusoidal congestion. CONCLUSIONS: Damage to hepatocytes and nonparenchymal cells after warm ischemia and reperfusion was thought to be closely related to sinusoidal microcirculatory disturbances in fatty livers. The canine fatty liver model reported here may be useful in studying the pathology of primary nonfunction and in establishing criteria for allowable degrees of fatty change in potential liver grafts. PMID- 10852592 TI - Nutritional immunomodulation leads to enhanced allograft survival in combination with cyclosporine A and rapamycin, but not FK506. AB - BACKGROUND: Recently, specific immunonutrients were found to increase experimental allograft survival when combined with cyclosporine A (CsA). This study compared the effect on rat cardiac allograft survival when nutritional immunomodulation was used with CsA, rapamycin (Rapa), or tacrolimus (FK506). METHODS: Intra-abdominal ACI to Lewis cardiac allografts were performed and assessed daily by palpation. Study groups included untreated controls and those receiving CsA, Rapa, or FK506. Rats were fed ad libitum with Impact diet (fortified with fish oil, arginine, and RNA) or standard rat food. Further study groups were transplanted that received a donor-specific transfusion in addition to immunosuppression and diet. RESULTS: Allograft survival was extended by combining Impact with CsA (45.3+/-19 days) and Rapa (165.3+/-52 days), but not FK506 (12.4+/-3.2 days). Mean graft survival in the Rapa/Impact group met criteria for functional tolerance. The addition of a donor-specific transfusion did not lead to graft survival advantages over similar groups not receiving a donor-specific transfusion. CONCLUSIONS: The use of immunonutrients improves transplant outcome in animals treated with short courses of CsA and Rapa, but not FK506. These findings highlight the potential differences in the effects of nutritional immunomodulation with different immunosuppressive drugs in the treatment of transplant patients. PMID- 10852593 TI - Chronic inhibitory effect of insulin on plasma lipid concentrations in rats with transplanted pancreas. AB - BACKGROUND: Hyperinsulinemia, which is usually related to insulin resistance, is considered to be an important risk factor for coronary artery disease. Our study examines the influence of portal delivery of endogenous insulin after pancreas transplantation on plasma lipid metabolism, as compared with systemic delivery of insulin. METHOD: Pancreas was transplanted heterotopically in normal rats by anastomosis of the donor portal vein to the recipient portal vein (PPTx) or inferior vena cava (CPTx) as an in vivo model of endogenous hyperinsulinemia. RESULTS: The mean value of plasma insulin concentration of CPTx and PPTx rats was 149 and 165% that of control rats, whereas the plasma glucose concentration of CPTx and PPTx rats did not differ significantly from that of control rats. Plasma triglyceride (TG) concentrations were significantly lower in both PPTx and CPTx rats than control rats. During the intravenous glucose tolerance test, the area under the insulin concentration curves of CPTx and PPTx rats was 204 and 215% that of control rats, and they were correlated negatively with plasma TG concentrations. Plasma cholesterol and TG concentrations were significantly lower in PPTx rats than in control and CPTx rats. CONCLUSIONS: Chronic hyperinsulinemia has a dose-dependent inhibitory effect on the regulation of plasma TG concentrations in rats with a transplanted pancreas. Significant lower lipid levels in PPTx rats than in CPTx rats suggest that portal delivery of insulin from the transplanted pancreas is relatively preventive for the atherosclerotic process as compared with systemic delivery of insulin. PMID- 10852594 TI - Transplantation of autologous and allogeneic bone marrow with liver from a cadaveric donor for primary liver cancer. AB - BACKGROUND: In histocompatibility mismatched experimental animals, a combination of T-cell-depleted autologous and allogeneic marrow may induce mixed chimerism and tolerance. Patients with large primary liver tumors have a poor outcome. We investigated whether it were possible to induce mixed chimerism and obtain an antitumor effect in a patient with a large primary liver cancer after combined liver and bone marrow transplantation (BMT). METHODS: A 46-year-old female with a primary non resectable liver cancer received a liver transplant from a cadaveric donor. Subsequently, she was conditioned with 4x2 Gy of total lymphoid irradiation, 120 mg/kg cyclophosphamide, and 7.5 Gy total body irradiation. Twelve days after liver transplantation, she received T-cell-depleted autologous:cadaveric 5/6 antigen HLA-mismatched marrow in a proportion of CD34+ cells of 0.5:3.0x10(6)/kg. Chimerism status was determined with polymerase chain reaction amplification of variable number tandem repeats from DNA obtained from CD3+, CD19+, and CD45+ magnetic-bead-separated cells. RESULTS: The early posttransplant period was uneventful; liver function was normal and the hematopoietic engraftment of donor and recipient origin was prompt. Alpha fetoprotein levels dropped from 440 to 35 microg/l. One month after marrow transplantation, donor T-cells decreased markedly. Monoclonal antibody OKT-3 and 10(5)/kg donor T-cells were given. One month later, the patient developed diarrhea and abdominal pain. A colonoscopy showed moderate gastrointestinal acute graft-versus-host disease and a Cryptosporidium infection. Three months after BMT, she became a complete donor chimera. Chimera cells showed little, if any, reactivity in mixed lymphocyte cultures to recipient and donor cells, but reacted to third party. Five months after BMT, she developed progressive Aspergillus fumigatus pneumonia and died. No tumor was found at the autopsy. CONCLUSION: We obtained mixed donor-recipient hematopoietic chimerism without severe acute graft versus-host-disease, after combined T-cell depleted autologous and allogeneic BMT and a transplantation of a liver from an HLA-mismatched cadaveric donor. Additional donor T-cells enhanced donor bone marrow engraftment, but rejected the autograft. On the basis of this first attempt, further clinical studies are warranted. PMID- 10852595 TI - Immunologic evaluation during the first year of life of infants born to cyclosporine-treated female kidney transplant recipients: analysis of lymphocyte subpopulations and immunoglobulin serum levels. AB - BACKGROUND: In rodents, CsA has been shown to affect T-cell development, giving rise to an abnormal production of mature T cells and the absence of many T-cell subsets as well as to autoimmunity. Surprisingly, only a few studies investigated the effect of the immunosuppressive drug on the immune system of the human fetus. METHODS: We examined six infants born to female kidney transplant recipients who had received cyclosporine and methylprednisolone throughout their pregnancies. Peripheral blood was obtained 1 day and 2, 4, 6, and 12 months after birth, and two-color flow cytometric immunophenotyping of lymphocytes was performed. RESULTS: Total T cells, as well as CD4+ and CD8+ T cells, were low at birth, but normalized thereafter. Among T-cell activation markers, the expression of CD25, the alpha chain of the interleukin-2 receptor, was below the normal range or low range throughout the study period, and HLA-DR expression was extremely low at birth and failed to increase up to 12 months. The number of total B cells was lower than normal at birth, but steeply increased over time. In contrast, B-cell subset bearing CD5 antigen was severely depleted throughout the first year of life. Total IgG concentration was significantly lower than in controls at 2 months, mainly because of subnormal levels of IgG1 and IgG3 subclasses, which remained in the low range up to 6 months. Finally, infants showed normal numbers of true natural killer (NK) cells (CD3-CD16+CD56+), whereas the expression of CD57 antigen, defining non-MHC-restricted cytotoxic lymphocytes, was barely detectable at birth and failed to increase over time, in both CD8+ and CD8- subsets. Of note, none of the infants had clinical evidence of an immunodeficient state. CONCLUSIONS: continuous exposure to CsA in utero seemingly impairs T-, B-, and NK-cell development and/or maturation, and most of its effects are still apparent at 1 year, which might suggest that conventional vaccinations should be delayed in these infants. PMID- 10852596 TI - Kidney function in cyclosporine-treated paediatric pulmonary transplant recipients. AB - BACKGROUND: Lung or heart-lung transplantation is a useful therapy in life threatening pulmonary disorders during childhood. Cyclosporine A is a major immunosuppressive treatment but has a number of adverse effects including nephrotoxicity. There have been no reports on the long-term evolution of renal function in a large series of paediatric pulmonary transplantation recipients. METHODS: We examined 19 patients followed up for at least 3 years after pulmonary transplantation. The mean time of follow-up was 5.36 years. Kidney function was evaluated by calculation of glomerular filtration rate (GFR) according the Schwartz formula. RESULTS: The GFR was normal before transplantation in all patients. The short-term evolution of GFR was marked by a significant drop during the first and until the 6th month. Then, regardless of the level reached at the end of the 6th month, the GFR remained stable in all patients except one until the end of follow-up. At the end of follow-up, 31% had normal GFR, 57% had mild chronic renal failure, and 5% had advanced renal failure. Hypertension was frequent and associated with renal failure. CONCLUSIONS: Paediatric pulmonary recipients showed evidence of long-term cyclosporine A-associated nephrotoxicity. Most of this toxicity occurred during the first 6 months. PMID- 10852597 TI - Double versus single renal allografts from aged donors. AB - BACKGROUND: The age limit of the cadaver kidney donors is increasing in response to the growing demand for renal transplantation. Simultaneous double kidney transplantation (SDKT) with kidneys obtained from elderly adults has been proposed to increase the transplantation number and improve its results. However, if SDKT is performed when there are no clear indications, a negative effect could be produced on the total number of transplanted patients as both kidneys would be used for only one recipient. MATERIAL AND METHODS: In December 1996 we designed a transplantation protocol to be able to extend the selection of cadaver kidney donors with normal serum creatinine levels without establishing any age limit. A pregraft renal biopsy was always performed to analyze the glomerulosclerosis (GE) percentage whenever the donors were 60 years of age or older. A SDKT was performed in a single recipient when the donor age was 75 years or older or when the donors between 60 and 74 years old had a GE rate of more than 15%. On the contrary, a single kidney transplantation was performed in two different recipients for kidneys from donors between 60 and 74 years of age with a GE rate of less than 15%. Kidneys having GE rates of more than 50% were discarded for transplantation. Donor kidneys from subjects younger than 60 years of age were always used for a single kidney transplantation. RESULTS: Based on the above mentioned protocol, from December 1996 to May 1998, 181 patients received a kidney transplantation in our hospital. These patients were divided into three groups: group I which included the SDKT recipients (n=21), group II or single kidney recipients from 60- to 74-year-old donors (n=40), and group III or recipients from <60-year-old donors (n=120). The mean follow-up time was 15+/-5 months (range 6-24). Mean donor age was 75+/-7 years in group I, this was significantly higher than in group II (67+/-4, P<0.001) and group III (37+/-15, P<0.001). The primary nonfunction rate was low in the three groups, there being no statistically significant differences (5, 5, and 4%, respectively). A significantly greater percentage of patients from group I (76%) presented immediate renal graft function as compared with group II (43%, P<0.01) and III (50%, P<0.05). The acute rejections rate was very low in all three groups (9.5, 7.5, and 22%, respectively) with significant differences between groups II and III (P<0.05). No significant differences between the different groups were observed for one year actuarial patient survival (100, 95, and 98%, respectively) or graft survival rates (95, 90, and 93%, respectively). The 6-month serum creatinine levels were excellent in the three groups, although there were significant differences between groups I and II (1.6+/-0.3 vs. 1.9+/-0.6 mg/dl, P<0.05), II and III (1.9+/-0.6 vs. 1.4+/-0.4 mg/dl, P<0.001), and I and III (P<0.05). CONCLUSIONS: Simultaneous double kidney transplantations make it possible to use kidneys from extremely elderly donors (>75 years) or those whose GE>15%. In addition, kidneys from donor 60-74 years old in which the GE<15% can be used for single kidney transplantations in two different recipients with excellent results. PMID- 10852598 TI - Kidney donors don't regret: follow-up of 370 donors in Stockholm since 1964. AB - BACKGROUND: The aim of the study was to present the views of our kidney donors since 1964, at the time of donation, as well as later on--and to assess their current subjective health. METHODS: A total of 451 living-donor nephrectomies were performed on Swedish residents in Stockholm from April 1964 until the end of 1995. A questionnaire with 11 questions about the donation and a standardized health form (SF-36) were sent to all donors alive in 1997 (n=403). RESULTS: The mean age (+/-SD) of the donors was 61+/-14 years at follow-up and the time-since donation was 12.5+/-7.7 years. The response rate was very good (92%). Current health, as assessed by form SF-36, was satisfactory. Donors scored somewhat better than those reported in a random sample of the Swedish population. The decision to donate had been easy: 86% made the decision themselves, without being pushed. Twenty-three percent thought that the nephrectomy had been troublesome. A higher percentage of young donors had felt that the postoperative period was difficult. Most donors (56%) stated that it had taken more than 2 months before they returned to a "normal" life, and 5% felt that they never completely recovered. Less than 1% of the donors regretted the donation. The commonest current medical prescription was antihypertensives (15%). The actual mean serum creatinine was 103+/-22 (range 48-219) micromol/L. CONCLUSIONS: The results indicate that the degree of health is at least as high as in the general population. The decision to donate was easy for most of the donors, but surgery and the recovery period were troublesome and lasted longer than expected. Kidney function was acceptable. PMID- 10852599 TI - Regulation of glucose tolerance in patients after liver transplantation: impact of cyclosporin versus tacrolimus therapy. AB - BACKGROUND: We investigated the factors regulating glucose homeostasis in 10 healthy (control) subjects, as well as in stable, long-term, liver-grafted patients receiving monotherapy in the form of either cyclosporin A (n=10) or tacrolimus (n=10). METHODS: We measured insulin sensitivity, first- and second phase insulin secretion, with a minimal modeling technique based on the analysis of glucose, insulin, and C-peptide profiles during frequently sampled intravenous glucose tolerance tests (FSIGTT). Proinsulin levels, as a marker of beta-cell dysfunction, were measured in the fasting state and during FSIGTT. RESULTS: Glucose and insulin concentrations before and after glucose loading did not differ in liver transplant patients and in control subjects. Fasting C-peptide levels in both liver-grafted groups were higher than in healthy subjects and remained elevated during FSIGTT (P<0.05). Intravenous glucose tolerance [(K(G)), i.e. the slope of the regression of logarithm of the blood glucose concentrations vs. time], insulin sensitivity, and first-phase insulin secretion did not differ in liver-grafted groups and healthy subjects. Second-phase insulin secretion was about 56% higher in liver-grafted patients than in controls (P<0.05). Body mass index was the overall determinant of insulin sensitivity in all groups. CONCLUSIONS: Long-term monotherapy with cyclosporin A or tacrolimus has no deleterious effects on insulin sensitivity, first-phase insulin secretion, and insulin synthesis in liver transplant patients. Normal insulin sensitivity (posthepatic insulin effect) and enhanced second-phase insulin secretion (prehepatic insulin) point to an accelerated hepatic insulin clearance rate in liver transplant patients. Increased hepatic insulin clearance is compensated by enhanced insulin secretion, indicating that insulin clearance is the major determinant of pancreatic function in liver-grafted patients. PMID- 10852600 TI - Parenchymal liver injury in orthotopic liver transplantation. AB - BACKGROUND: A 35-year period of clinical development resulted in orthotopic liver transplantation (OLT) becoming a standardized surgical procedure. Despite this progress, the rate of technical complications is still high. Although the main problem in most analyses is vascular or bile duct failure, we observed a remarkable number of parenchymal liver injuries that led to intraoperative problems. Our aim, therefore, is to present an overall report on the incidence, treatment, and clinical course of parenchymal liver injuries in OLT. METHODS: Five hundred seventy-two consecutive OLT procedures performed between 1988 and 1998 were analyzed in a retrospective study. Parenchymal liver injury was diagnosed by means of examination of the surgical reports. Donor- and recipient related data followed the medical report. The lesions were classified according to the Organ Injury Scale. RESULTS: Parenchymal liver injury was diagnosed in 23 patients (4%). The lesions were classified as grade Ia (13.1%), grade Ib (13.1%), grade IIb (52.1%), grade IIIa (17.1%), and grade IIIb (4.3%). In 19 patients (82.6%), the lesion was detected during OLT, and in four patients (17.4%), during relaparotomy. The latter group showed significantly higher-grade injuries. Treatment was suture or fibringlue alone, 17.4%; fibringlue and hemostyptics, 26.1%, mesh wrapping 30.4%, and mesh packing 26.1%. Seven patients (30.4%) underwent relaparotomy. Further active bleeding was not found in any of them. Statistical analysis found a correlation between injury grade and relaparotomy rate. No patients died as a result of parenchymal liver injury. CONCLUSIONS: Parenchymal liver injuries can be treated well, with no adverse effect on patient or graft survival. An early decision concerning the surgical procedure for controlling hemorrhage is required. A basically aggressive therapeutic approach might avoid further complications relating to reperfusion edema. PMID- 10852601 TI - Sirolimus-induced thrombocytopenia and leukopenia in renal transplant recipients: risk factors, incidence, progression, and management. AB - BACKGROUND: Our study assessed the factors that predispose renal transplant recipients to the occurrence of thrombocytopenia and leukopenia, as well as the severity and the time- and concentration-dependence of these side-effects, after administration of sirolimus (SRL) in combination with a cyclosporine (CsA) and prednisone (Pred) regimen. METHODS: The clinical courses of two cohorts of renal transplant recipients were compared over 1 year: 119 patients received SRL in addition to CsA and Pred, and 65 demographically similar, concurrent patients received only CsA and Pred. Using an analysis of variance, pretransplant laboratory values and SRL trough concentrations (C0) were correlated with the occurrence, severity, and persistence of drug-induced thrombocytopenia (platelet count <150x10(3) cell/mm3) and/or leukopenia (white blood cell count <5,000/mm3). RESULTS: Neither the ethnic background nor the pretransplant cytomegalovirus serological status was associated with the occurrence of hematological complications. Thrombocytopenia was usually observed during the first 4 weeks of treatment (P=0.004). The occurrence, but not the severity or the persistence, of both thrombocytopenia and leukopenia correlated significantly with SRL trough concentrations > or =16 ng/ml (P=0.001 and 0.0001, respectively). A significant correlation is evident between the occurrence of the two adverse effects (P=0.001). In 89% of patients, the first episode of either type of cytopenia resolved spontaneously. Among the remaining 11%, 7% responded to SRL dose reduction, and 4% to temporary suspension. No patient required permanent cessation of SRL therapy. Most patients experienced repeated, but self-limited, episodes of toxicity. CONCLUSION: Thrombocytopenia and leukopenia are not infrequent occurrences with SRL treatment, and they generally resolve spontaneously. PMID- 10852602 TI - HBV genotypic resistance to lamivudine in kidney recipients and hemodialyzed patients. AB - BACKGROUND: Lamivudine is a potent inhibitor of human immunodeficiency virus reverse transcriptase and hepatitis B virus (HBV) DNA polymerase. Its overall efficiency is clearly hampered by relapse at discontinuation and by risk of genotypic resistance. We describe herein the first cases of HBV resistance to lamivudine in kidney recipients and hemodialyzed patients. METHODS: We analyzed 26 HBV-infected kidney recipients and five hemodialyzed patients treated with lamivudine who became serum HBV DNA-negative (by Digene test). The biological and virological follow-up identified breakthrough as defined by the reappearance of serum HBV DNA. In two cases of breakthrough, HBV DNA was amplified and sequenced through the polymerase domain, including the YMDD motif, before the beginning of treatment and at time of breakthrough to determine genotypic mutations. RESULTS: Ten breakthroughs (reappearance of serum HBV DNA) were observed after a median follow-up of 11 months in eight kidney recipients and two hemodialyzed patients after a median duration of treatment of 16.5 (from 4 to 31) months of treatment. Previous HBe/anti-HBe seroconversion was not observed in the patients who escaped. In two kidney recipients, the comparison of HBV-DNA sequences before the treatment and after the breakthrough identified in one case a mutation of the highly conserved YMDD motif (YVDD), whereas in the second case, no genotypic mutation was observed in the sequenced region. CONCLUSION: We report the first cases of HBV genotypic resistance to lamivudine in kidney recipients and hemodialysis patients. Genotypic resistance is observed after 4-31 months of therapy. The YMDD mutation does not account for all cases of virological escape. PMID- 10852603 TI - Non-heart-beating organ donors: a potential source of islets for transplantation? AB - BACKGROUND: The shortage of organ donors relative to the number of patients on transplant waiting lists has led to a renewed interest in the use of non-heart beating (NHB) organ donors in many centers. The lack of donors is also a problem for islet transplantation. The disparity between donor organs and potential recipients is further exacerbated by the requirement to transplant a large number of islets to increase the chance of success and the high level of variability in islet isolation yield. Non-heart-beating (NHB) donors have not previously been assessed as a source of islets for transplantation, and it is unknown what affects the additional factor of warm ischemic injury associated with NHB organs may have on the success of islet isolation. METHODS: This study assesses the yield and function of islets from NHB donors and compares the results with islets obtained from heart-beating brain-dead (HB) donors. RESULTS: There were no differences in the yield of islets per gram of pancreas, 1788 (0-4620) NHB vs. 1580 (26-2544) HB (median, range). The secretory function was also similar in both groups, with stimulation indices of 0.71-3.49 for NHB vs. 0.30-3.57 for HB (overall range). There was no correlation between islet yield and warm ischemia time in the NHB donor group. CONCLUSIONS: In conclusion, the study has demonstrated that it is possible to isolate large numbers of islets from NHB donor pancreata and that, where NHB donor programs exist, these could provide a significant addition to the number of potentially transplantable islets. PMID- 10852604 TI - Allograft loss in renal transplant recipients with Fabry's disease and activated protein C resistance. AB - INTRODUCTION: Fabry's disease is associated with an increased incidence of thrombotic events and rejection. Spontaneous thrombosis of a functioning cadaveric renal allograft in a recipient with Fabry's disease prompted prospective evaluation of all transplant candidates with Fabry's disease for hypercoagulability. MATERIALS AND METHODS: Transplant candidates with Fabry's disease were tested for hypercoagulability, analyzed for HLA-type and ABO group, and comorbid conditions suggestive of hypercoagulability. RESULTS: A unique association of Fabry's disease with activated protein C Resistance was documented in a cohort of Caucasian male renal transplant recipients with Fabry's disease. Four of five patients were blood group A and had no significant comorbid conditions suggestive of hypercoagulability. The resistance to activation of protein C (APCR)(+) patients shared HLA loci-B8 and Dr3, although the APCR(-) patients shared HLA loci-B27 and -B38. CONCLUSIONS: Due to the observed increase in the incidence of APCR in our Fabry's cohort, we suggest screening all patients with Fabry's disease for APCR. Because factor V and factor Va receptors are found on vascular endothelium and peripheral blood monocytes, APCR in the presence of Fabry's disease may be a nonimmunological stimulus for rejection. Analysis of HLA typing in patients with Fabry's disease may further elucidate HLA-based association of Fabry's disease and resistance to activated protein C with the risk of thrombosis and rejection. PMID- 10852605 TI - Increased expression of the lymphocyte early activation marker CD69 in peripheral blood correlates with histologic evidence of cardiac allograft rejection. AB - BACKGROUND: The human leukocyte membrane protein CD69 is an early activation marker induced in T lymphocytes, B cells, and natural killer cells in response to inflammatory stimuli. Cardiac catheterization and endomyocardial biopsy remain the "gold standard" for diagnosis of rejection after transplantation, and noninvasive methods of rejection surveillance have long been sought. We studied CD69 membrane protein expression in peripheral blood T lymphocytes obtained from pediatric cardiac transplant recipients at the time of biopsy and correlated the results with histologic rejection scores. METHODS: Heparinized whole blood samples were obtained from pediatric cardiac transplant recipients at the time of cardiac biopsy, as well as from control subjects. Lymphocytes were labeled with antibodies for CD3, CD4, CD8, and CD69 and analysis performed using flow cytometric methods. RESULTS: Resting CD69 expression (measured as a percentage of gated events) was significantly increased in patients with concurrent histologic evidence of rejection (International Society for Heart and Lung Transplantation grade > or =3A) when compared to those with minimal or no rejection and controls. Although statistically significant for both lymphocyte subsets, this relationship was more pronounced for CD8+ T cells (P<0.001) than for CD4+ T cells (P=0.001). When data were analyzed by rejection score, a percentage activation of the CD8+ subset (CD69+/CD8+ cells as a percentage of total gated events) exceeding 15% correlated with significant rejection. CONCLUSIONS: Measurement of the expression of the early activation marker CD69 in peripheral blood lymphocytes by flow cytometry may provide a noninvasive means of assessing immune activation and possible rejection in cardiac transplant recipients. PMID- 10852606 TI - Segmental degradation of left ventricular wall motion after persistent coronary fistula in a posttransplantation patient: a case report and short review of literature. AB - A 50-year-old man received an orthotopic heart transplant because of severe coronary heart disease and congestive heart failure. Two years after the transplantation, a continuous murmur occurred at the left sternal edge after repeated endomyocardial biopsies. Echocardiography and coronary angiography revealed a dilated left anterior descending artery with a fistula to the right ventricle. The circumflex was large with an equally postero-lateral branch, and the right coronary artery was rather small with collaterals to the distal part of the left anterior descending branch. The patient had refused any intervention to close the fistula. The left ventricular levogram was normal. Two years later, in a follow-up angiogram, the left ventricular ejection fraction had decreased as a result of hypo- and akinesis of the apex and posterior wall. We suggest that this local wall motion disturbance derives from a steal phenomenon rather than being a sequela of rejection. The decrease in left ventricular ejection fraction was associated with shortness of breath upon moderate exercise. Standard heart failure medication relieved the patient's symptoms. The observation of local wall motion disturbances in this case, as well as conflicting views in the literature, raises the question whether postbiopsy coronary fistulas in transplant patients should be closed. PMID- 10852607 TI - Cardiac transplantation in survivors of lymphoma: a multi-institutional survey. AB - BACKGROUND: Cardiac transplantation has been successfully performed in patients with a history of presumably cured Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL). Though the risk of recurrence is a major concern, the long-term influence of prior cancer and cancer therapy on posttransplant outcome has not been previously investigated. METHODS: Questionnaires were sent to 130 cardiac transplant centers in the United States registered with the United Network for Organ Sharing. Data collected included patient demographics; type, stage, and timing of HD/NHL; treatment for HD/NHL; posttransplant immunosuppressive regimen, rejection history, and outcomes; and Epstein-Barr virus status. RESULTS: Thirty four cardiac transplant recipients with a previous history of HD (n=16) or NHL (n=18) were identified. HD patients averaged 41+/-15 years of age, with a mean disease-free interval of 15+/-9 years at the time of transplantation. NHL patients averaged 42+/-17 years of age with a mean disease-free interval of 10+/ 9 years at the time of transplantation. The mean follow-up for the entire group was 50 months (range, 2 days to 136 months), and mean follow-up for the survivors was 67 months (range, 23-136 months). The 1-, 3-, 5-, 7-, and 10-year actuarial survival estimates for the entire group are 77%, 64%, 64%, 64%, and 50%, respectively. Actuarial survival was lower in HD patients (P=0.04) and in patients who had previously undergone splenectomy (P=0.008). Cox regression analysis identified only prior splenectomy (P=0.02) as an independent risk factor for mortality after cardiac transplantation with an adjusted relative risk of 6.2 (1.7-21.9, 95% confidence intervals). CONCLUSIONS: Although the numbers are small, these data strongly suggest that there is an increased mortality risk for cardiac transplant recipients with prior HD who have undergone splenectomy. PMID- 10852608 TI - Improved fibrinolytic capacity after withdrawal of steroid immunosuppression in renal transplant recipients. AB - BACKGROUND: Long-term steroid immunosuppression has been associated with the prothrombotic state observed in renal transplant (RT) patients, in whom both hypercoagulability due to an increase of von Willebrand factor/factor VIII complex, and impaired fibrinolysis due to PAI-1 excess have been demonstrated. Our aim was to investigate the effect of steroid withdrawal on fibrinolytic capacity in a group of RT patients. METHODS: The fibrinolytic study was performed in 28 RT patients under stable immunosuppression therapy with cyclosporine, azathioprine, and methylprednisolone; only 12 of these patients could repeat the study 6 months after steroid withdrawal. Euglobulin lysis time (ELT), tissue plasminogen activator activity (t-PA:act) and antigen (t-PA:Ag), PAI-1 activity (PAI-1:act), and antigen (PAI-1:Ag) were assayed on blood samples drawn before and 20 min after the venous occlusion test (VO). RESULTS: An hypofibrinolytic state due to a significant increase in PAI-1 levels was confirmed in RT patients receiving triple immunosuppression therapy. RT patient who stayed off steroids showed a significant shortening of ELT both before (P=0.01) and 20' after VO (P=0.005) at the 6-month control. Moreover, after steroid withdrawal, PAI-1:Ag levels decreased significantly (P=0.002) and normalized; in a similar manner PAI 1:act levels also showed a significant decrease both before (P=0.001), and after VO (P=0.0001). The prevalence of RT patients with impaired fibrinolytic capacity was as high as 83.3% during steroid treatment, and dropped to 16.7% after steroid withdrawal. CONCLUSIONS: Our findings confirm that steroid withdrawal may normalize impaired fibrinolytic capacity in RT patients; this improvement may further contribute to reduce the thrombotic risk associated with renal transplantation. PMID- 10852609 TI - High prevalence of erectile dysfunction after renal transplantation. AB - BACKGROUND AND METHODS: A cross-sectional study of multifaceted male sexual function in 323 consecutive kidney transplant recipients was conducted by mail by means of the validated International Index of Erectile Function (IIEF). All five IIEF domains (IIEF-5), i.e., erectile function, orgasmic function, sexual desire, intercourse satisfaction, and overall satisfaction, were scored for each responder. IIEF-5 scoring that conformed to the National Institutes of Health definition of erectile dysfunction (ED) was computed for all patients sexually active within the past 4 weeks. RESULTS: Two hundred and seventy-one patients replied. Compared to the controls used for IIEF psychometric validation, kidney transplant recipients gave lower erectile function (P<0.01) and intercourse satisfaction (P<0.05) scores, despite their being younger. ED, according to the IIEF-5 method, was demonstrated in 55.7% of the sexually active patients (n=212). Age, time on dialysis, and iterative transplants were significantly and negatively related to erectile dysfunction. CONCLUSION: IIEF proved to be a valuable means of unveiling highly prevalent erectile dysfunction in male kidney transplant recipients. The negative impact of the time on dialysis was emphasized in the results. PMID- 10852610 TI - Peri-operative blood lactate levels in recipients of living-related liver transplantation. AB - BACKGROUND: The role of changes in peri-operative blood lactate levels in recipients of living-related liver transplants has not yet been clarified. METHODS: Forty-three recipients were included in this study. Blood lactate, plasma total bilirubin, aminotransferase, body temperature, and gastric mucosal PCO2 levels were measured at six time points during surgery: just before the initiation of surgery, just after dissection of the hepatic vasculature, at the end of the anhepatic phase, and 30, 60, and 120 min after reperfusion. We calculated the rate of lactate accumulation during the pre-anhepatic and anhepatic phases and the elimination rate during reperfusion (neohepatic phase), and examined the correlation between these results and the clinical findings. RESULTS: The rate of lactate elimination during the neohepatic phase was correlated with the ratio of graft weight to standard liver volume (P<0.0001). There was also a significant correlation between the rate of lactate accumulation during the pre-anhepatic phase and the preoperative total bilirubin levels (P=0.0008). CONCLUSIONS: Each pre-anhepatic, anhepatic, and neohepatic phase had a characteristic blood lactate profile. The graft size strongly affected lactate levels during the early neohepatic phase. PMID- 10852611 TI - Treatment of mild hyperhomocysteinemia in renal transplant recipients versus hemodialysis patients. AB - BACKGROUND: Mild hyperhomocysteinemia is common among maintenance hemodialysis (HD) patients and renal transplant recipients (RTR) and may contribute to the excess incidence of arteriosclerotic outcomes experienced by both patient groups. Relative to their RTR counterparts, the hyperhomocysteinemia of HD patients seems to be considerably more refractory to treatment with high-dose folic acid (FA) based B-vitamin supplementation regimens, although controlled comparison data are lacking. METHODS: We compared the relative responsiveness of (n=10) RTR and (n=39) HD patients with equivalent baseline total homocysteine (tHcy) levels (i.e., RTR range=14.2-23.6 micromol/L; HD range=14.4-24.9 micromol/L) to 12 weeks of tHcy-lowering treatment. The RTR received 2.4 mg/day of FA, 50.0 mg/day of vitamin B6, and 0.4 mg/day of vitamin B12, while the HD patients received 15 mg/day of FA or an equimolar amount (17 mg/day) of the reduced folate, L-5 methyltetrahydrofolate, in addition to 50.0 mg/day of vitamin B6, and 1.0 mg/day of vitamin B12. RESULTS: The mean percent (%) reductions (+/-95% confidence interval) in tHcy were: RTR=28.1% (16.2-40.0%); HD=12.1% (6.6-17.7%), P=0.027 for comparison of between-groups differences by analysis of covariance adjusted for baseline tHcy levels. Moreover, (50.0%) of 10 of the RTR versus only (5.1%) of 39 of the HD patients had final on-treatment tHcy levels <12 micromol/L; P=0.002 for comparison of between-groups differences by Fisher's exact test. CONCLUSION: Relative to RTR with comparable baseline tHcy levels, the mild hyperhomocysteinemia of maintenance HD patients is much more refractory to tHcy lowering B-vitamin treatment regimens featuring supraphysiological amounts of FA or the reduced folate, L-5-methyltetrahydrofolate. Accordingly, RTR are a preferable target population for controlled clinical trials testing the hypothesis that tHcy-lowering B-vitamin intervention may reduce arteriosclerotic cardiovascular disease event rates in patients with chronic renal disease. PMID- 10852612 TI - Docosahexaenoic acid decreases IRF-1 mRNA and thus inhibits activation of both the IRF-E and NFkappa d response elements of the iNOS promoter. AB - BACKGROUND: Nitric oxide produced by inducible nitric oxide synthase (iNOS) may be cytotoxic during cardiac, hepatic, and renal acute allograft rejection. Because the incidence of rejection is decreased by fish oils that contain docosahexaenoic acid (DHA), we investigated the effects of DHA on iNOS. Using nuclear run-on assays and iNOS-promoter constructs, we previously showed that docosahexaenoic acid (DHA) inhibits activation of the iNOS gene by murine macrophages that had been stimulated in vitro by IFNgamma plus lipopolysaccharide. METHODS: In our current investigation, our purpose has been to determine how DHA inhibits iNOS gene activation in murine macrophages, by using gel retardation and Northern blotting techniques. We studied the effects of DHA on the formation nuclear protein complexes that interact with the critical iNOS promoter's response elements for IRF-1 (IRF-E -923 to -913 bp) and NF-kappaB (NFkappa d -85 to -75 bp). RESULTS: We now show that DHA inhibited increases of IRF-1 mRNA abundance in response to IFNgamma plus lipopolysaccharide. As expected, we found that this prevented formation of the nuclear protein complex that binds to the IRF-E DNA response element. We also found that inhibition of IRF-1 inhibited formation of the nuclear protein complex that binds to the NFkappa d DNA response element. CONCLUSIONS: DHA decreases the abundance of IRF-1 mRNA in stimulated cells. That, in turn, results in the decreased nuclear protein binding to the major iNOS promoter response elements (IRF-E and NF-kappaB). We found that this occurred because IRF-1 is a component of both the nuclear protein complex that binds to IRF-E and the nuclear protein complex that binds to NFkappa d. PMID- 10852613 TI - Altered intragraft immune responses and improved renal function in MHC class II deficient mouse kidney allografts. AB - BACKGROUND: During renal allograft rejection, expression of MHC class II antigens is up-regulated on the parenchymal cells of the kidney. This up-regulation of MHC class II proteins may stimulate the intragraft alloimmune response by promoting their recognition by recipient CD4+ T cells. In previous studies, absence of donor MHC class II antigens did not affect skin graft survival, but resulted in prolonged survival of cardiac allografts. METHODS: To further explore the role of MHC class II antigens in kidney graft rejection, we performed vascularized kidney transplants using donor kidneys from A(beta)b-deficient mice that lack MHC class II expression. RESULTS: At 4 weeks after transplant, GFR was substantially depressed in control allografts (2.18+/-0.46 ml/min/kg) compared to nonrejecting isografts (7.98+/-1.62 ml/min/kg; P<0.01), but significantly higher in class II- allografts (4.38+/-0.60 ml/min/kg; P<0.05). Despite the improvement in renal function, class II- allograft demonstrated histologic features of acute rejection, not unlike control allografts. However, morphometric analysis at 1 week after transplantation demonstrated significantly fewer CD4+ T cells infiltrating class II- allografts (12.8+/-1.2 cells/mm2) compared to controls (25.5+/-2.6 cells/mm2; P=0.0007). Finally, the intragraft profile of cytokines was altered in class II- allografts, with significantly reduced expression of Th2 cytokine mRNA compared to controls. CONCLUSIONS: These results support a role of MHC class II antigens in the kidney regulating immune cells within the graft. Further, effector pathways triggered by class II antigens promote renal injury during rejection. PMID- 10852614 TI - Anergic T cells generated in vitro suppress rejection response to islet allografts. AB - BACKGROUND: Induction of antigen-specific unresponsiveness to grafts is the ultimate goal for organ transplantation. It has been shown that anergic T cells generated in vivo can be transferred as suppressor cells. Anergic cells generated in vitro have never been successfully used to prevent allograft rejection in vivo. We examined whether anergic cells generated in vitro by blocking CD28/B7 costimulatory pathway can suppress allograft rejection in vivo. METHODS: Anergic T cells were generated in vitro by the addition of anti-B7-1 and anti-B7-2 monoclonal antibodies (mAbs) to primary mixed lymphocyte reaction (MLR) consisting of C57BL/6 (B6) splenocytes as responder and irradiated BALB/c splenocytes as stimulator. We tested the ability of these cells to respond to various stimuli and to suppress alloreactive T-cell responses in vitro. For in vivo studies, 4x10(7) anergic cells were injected intravenously immediately after transplantation of BALB/c islets under the renal subcapsular space of streptozotocin-induced diabetic and 2.5-Gy X-irradiated B6 mice. RESULTS: Anergic cells treated with both mAbs in the primary MLR did not proliferate in secondary MLR against BALB/c and third-party C3H/He stimulators. The cells also failed to respond to immobilized anti-CD3 mAb, although they proliferated in response to concanavalin A or phorbol myristate acetate + ionomycin. The anergic state was reversed by the addition of exogenous IL-2. Furthermore, these cells suppressed the proliferation of naive B6 T cells against either the same (BALB/c) or third party (C3H/He) stimulator cells. In in vivo studies, irradiated B6 mice rejected BALB/c islet allografts acutely with a mean survival time of 27.0+/-8.3 days, whereas two of six animals injected with the anergic cells accepted the allografts indefinitely (>100 days) with a mean survival time of 52.0+/-38.2 days. CONCLUSIONS: Anergic cells generated in vitro by blocking CD28/B7 costimulatory pathway suppress islet allograft rejection after adoptive transfer. This procedure might be clinically useful for promoting allograft survival. PMID- 10852615 TI - The role of renal sympathetic nerves in experimental chronic cyclosporine nephropathy. AB - BACKGROUND: Sympathetic nervous system hyperactivity has been postulated to play a major role in the intense intrarenal vasospasm and hypertension provoked by cyclosporine. It has been argued that the denervated renal allograft may be partially protected from the tubulointerstitial fibrosis associated with chronic cyclosporine administration compared with innervated kidneys in extrarenal transplantation. METHODS: Utilizing a model of chronic cyclosporine nephropathy in which striped fibrosis develops in the uninephrectomized salt-depleted rat, the effect of renal denervation on renal structure and function was examined. Sprague-Dawley rats maintained on a low-salt diet underwent uninephrectomy and contralateral renal denervation or sham denervation, followed by cyclosporine 15 mg/kg daily by injection. RESULTS: After 21 days, glomerular filtration was markedly depressed and linear zones of tubular atrophy and interstitial fibrosis had developed compared with vehicle-treated control animals (P<0.001). However, there was no significant difference in either renal function or structure between denervated and sham-operated animals treated with cyclosporine. CONCLUSION: We conclude that renal sympathetic neural hyperactivity is not important in the development of chronic cyclosporine nephropathy. PMID- 10852616 TI - Prolonged allograft survival through conditional and specific ablation of alloreactive T cells expressing a suicide gene. AB - BACKGROUND: Control of antidonor activated T cells involved in allograft rejection while preserving immunocompetence is a challenging goal in transplantation. Engineered T cells expressing a viral thymidine kinase (TK) suicide gene metabolize the nontoxic prodrug ganciclovir (GCV) into a metabolite toxic only to dividing cells. We evaluated this suicide gene strategy for inducing transplantation tolerance in mice. METHODS: Transgenic mice expressing TK in mature T cells were analyzed for (i) specific T-cell depletion under GCV treatment upon various stimulations; (ii) outcome of allogeneic nonvascularized skin or heart allografts under a short 14-day GCV treatment initiated at the time of transplantation; and (iii) the capacities of T cells from such allotransplanted mice to proliferate in mixed lymphocyte reactions and to induce graft-versus-host disease in irradiated recipients with the genetic background of the donor allograft. RESULTS: Upon in vitro or in vivo GCV treatment, only activated dividing TK T cells but not B cells were efficiently depleted. Acute rejection of allogeneic grafts was prevented and a significant prolongation of graft survival was obtained, although associated with signs of chronic rejection. Prolonged skin graft survival correlated with decreased in vitro and in vivo T cell reactivities against donor alloantigens, whereas overall immunocompetence was preserved. CONCLUSIONS: Efficient and specific depletion of alloreactive TK T cells can be achieved by administrating GCV. These results open new perspectives for the control of allogeneic graft rejection using suicide gene therapy. PMID- 10852617 TI - Ingested interferon-alpha prevents allograft islet transplant rejection. AB - BACKGROUND: Ingested interferon (IFN)-alpha is a biological response modifier in experimental autoimmune encephalomyelitis and multiple sclerosis, and prevents type 1 diabetes in nonobese diabetic mice. Islet transplantation possesses significant potential advantages over whole-gland transplantation because it is simple, may achieve insulin independence, and has clear advantages over exogenous insulin therapy. Therefore, we examined whether ingested IFN-alpha, administered to islet allograft recipients, could prevent islet allograft rejection. METHODS: Recipient C3H mice (H2k) were made diabetic and either untreated or treated with 10-1000 international units (IU) of ingested murine IFN-alpha daily from day -7 through day +14 after transplantation for a total of 21 days. Seven days after diabetes induction, recipients received allograft islets isolated from C57BL.10 donors (H2b) under the kidney capsule and were followed for overt diabetes via elevated blood glucose. RESULTS: Control recipients and recipients fed 1000 IU all became diabetic by day 13, whereas mice ingesting IFN-alpha had delayed rejection for up to 27 (10 IU) to 29 days (100 IU) after islet transplantation. Treatment of recipients of islet allografts with ingested IFN-alpha doubles the time period before rejection compared with control mice. The feeding period with daily IFN-alpha was doubled from 21 days to 42 days in total, 7 days before transplantation and 35 days after transplantation. CONCLUSION: Treatment of recipients of islet allografts with prolonged ingested IFN-alpha prevents rejection in a subset of recipients. Ingested IFN-alpha may prevent rejection if given continuously after transplantation. PMID- 10852618 TI - A polymerase chain reaction-based protocol for the detection of transmission of pig endogenous retroviruses in pig to human xenotransplantation. AB - BACKGROUND: Xenotransplantation of pig organs and tissues to humans bears the risk of infection of immunosuppressed recipients by porcine endogenous retrovirus (PERV) released from the transplanted tissue. However, when diagnosing potential PERV transmission, it is essential to exclude microchimerism, i.e., persisting pig cells in analyzed bioptic material of xenotransplanted patients, which give rise to false positive PERV signals. Polymerase chain reaction (PCR) is so far the only suitable method to diagnose a cross-species transfer of PERV, but the exclusion of microchimerism might be a serious problem because most of the presently employed primer pairs detect PERV sequences with higher sensitivity than primers used for the detection of contaminating pig sequences. METHODS: We designed and evaluated a novel and improved primer set for detection of pig sequences as well as complementing positive control primers on the basis of mitochondrial cytochrome B, an approved marker for phylogenetic studies. We further established primer pairs derived from the long terminal repeat/leader region of PERV isolated from a Duroc German Landrace cross-bred pig and tested their sensitivity in comparison with known PERV- and pig-specific PCR markers. RESULTS: In standard PCR assays, the new cytochrome B-derived primers are at least 10 times more sensitive than the presently used PERV retroviral polymerase gene and mammalian beta-actin primers. When tested in a tissue culture infection model, PERV transmission to human 293 cells can be unambiguously demonstrated, even in the presence of up to 10% pig cells. One of the primer combinations derived from the PERV DuxDL3791 long terminal repeat/leader region amplifies with even lower sensitivity than primers detecting porcine beta-globin, thus permitting the exclusion of microchimerism also via chromosomal loci. CONCLUSIONS: The availability of the new PCR markers allows the proposal of a rigorous setup for the routine detection of PERV transmission after xenotransplantation. PMID- 10852619 TI - The relationship between acute rejection and chronic rejection is highly dependent on specific MHC matching: a multi-strain rat heterotopic heart transplant study. AB - BACKGROUND: The impact of acute rejection, immunosuppression, and infection, specifically cytomegalovirus infection, on the development of chronic rejection in the cardiac allograft, has been the subject of a large number of investigations. One of the difficulties in finding associations has been the marked immunologic heterogeneity of the patient population coupled with the lack of the ability to HLA match. Furthermore, the ideal animal model, which duplicates as well as controls for this immunologic heterogeneity, is lacking. METHODS: To try to simulate differences in HLA matching, immunosuppression regiments and cytomegalovirus infection, heterotopic heart transplantation was performed in two separate, complete MHC mismatch, rat strain combinations (WF LEW, BN-LEW) requiring chronic immunosuppression and employing four separate immunosuppression/infection protocols. Animals were followed for 6 months, killed, and rejection and vascular changes were scored blinded to the group. RESULTS: The mean vascular and acute rejection scores were not significantly different between treatment regiments for either specific strain combination. There was a trend for the subtherapeutic groups to have higher vascular scores. Overall, there were no significant differences in vascular scores between the WF LEW and BN-LEW groups (1.25+/-0.18 vs. 1.13+/-0.20, P=NS). Similar numbers of WF LEW and BN-LEW exhibited cellular infiltration and necrosis of the allograft, but the intensity of the response (rejection score) was more severe in the WF-LEW combination (4.54+/-0.22 vs. 3.92+/-0.21, P=0.052) when limiting the analysis to those with myocyte necrosis. There was no significant correlation between acute rejection and vascular lesion severity in the WF-LEW combination (r=0.22, P=NS) but a high correlation between these parameters in the BN-LEW combination (r=0.74, P<0.0001). CONCLUSIONS: These data suggest that, although acute rejection and chronic rejection are related, MHC matching may influence their interdependence. These data also may explain why the clinical association between acute and chronic rejection is difficult to demonstrate. PMID- 10852620 TI - The influence of conventional and cross-reactive group HLA matching on cardiac transplant outcome: an analysis from the United Network of Organ Sharing Scientific Registry. AB - BACKGROUND: The short tolerable cold ischemia time and the importance of other risk factors have generally superseded the role of HLA matching in the allocation of donor hearts. Recent advances in the accuracy and time required to perform HLA typing and crossmatching, however, have led us to re-examine the United Network of Organ Sharing Transplant Registry for the effects of the HLA incompatibility on outcome in relation to other possible risk factors. METHODS: These include conventional HLA-A, -B, and cross-reactive group (CREG) mismatching (mm), HLA-DR mm, pretransplantation panel-reactive antibody (PRA), recipient and donor race and donor age, cold ischemia time, and the pretransplantation use of either a left ventricular assist device or an intra-aortic balloon pump. RESULTS: Three year survival was clearly inferior in non-white (0.6921) as compared with white (0.7632) recipients, but this difference could not be accounted for by the degree of donor-recipient HLA mm that had occurred by chance. Nevertheless, the degree of mm that did occur seemed to have an impact on survival. The importance of HLA DR mm was confirmed, and it ranked only behind the use of an assist device and recipient race in the multivariate analysis. HLA-A and B mm exerted an additional effect, but this was only true in white recipients. Of these, HLA-A achieved statistical significance as an independent risk factor. In general, CREG mm was not a significant variable. However, more than twice as many 0-1 or 0-2 CREG, 0 DR mm as compared with 0-1 or 0-2 A,B, 0 DR mm transplants enjoyed approximately equal and very good 1- and 3-year survival. Assuming no change is cold ischemia time, the potential number of 0 CREG, 0 DR mm, ABO-compatible transplants that could be achieved when an Organ Procurement Organization had 50-100 patients on their waiting list was calculated. The surprisingly high frequency of approximately 24-36% suggests that this favorable match could be considered along with other important factors in the local allocation process. When pretransplantation PRA was analyzed as a continuous variable from 0 to 100%, it was a highly significant risk factor, but this effect was more strikingly evident when the PRA was analyzed in 20% increments above zero. Recently, left ventricular assist device usage has become increasingly common, and it has been associated with strikingly increased pretransplantation PRA levels. When they occur together, the data indicates that these patients are at a very high risk for graft failure. CONCLUSIONS: We believe that newer typing and crossmatching techniques make it possible to add HLA criteria to the allocation protocol of donor cardiac organs and would lead to improved long-term survival. PMID- 10852621 TI - Increased urinary excretion of endothelin during hypothermic perfusion preservation in kidneys subjected to preretrieval warm ischemic injury. AB - The purpose of the study was two-fold: 1) to determine whether endothelin (ET) levels could be detected in the ureteral effluent during hypothermic perfusion preservation (HPP) and; 2) to determine whether preretrieval warm ischemic (WI) injury is associated with increased ureteral excretion of ET. In situ pre-WI injury was induced in Lewis rats (n=10) by a 30-min extrinsic occlusion of the suprarenal aorta. The left kidney underwent 16 hr of HPP, and ureteral effluent (UE) from ischemic and control kidneys (n=10) was collected over 16 hr of HPP. The UE ET concentration and total ET excretion over 16 hr of HPP were significantly higher in kidneys subjected to pre-WI injury compared with nonischemic controls. Kidneys subjected to pre-WI injury can be distinguished from nonischemic control kidneys during HPP by a significantly higher concentration of ET in the UE and a higher overall excretion of ET during HPP. PMID- 10852622 TI - Apoptosis of thymocytes during acute graft-versus-host disease is independent of glucocorticoids. AB - BACKGROUND: Elimination of immature thymocytes resulting in thymic atrophy is characteristic of acute graft-versus-host disease (aGVHD). Because aGVHD has been associated with elevated glucocorticoid (GC) production, and CD4,CD8 double positive thymocytes undergo rapid apoptosis in response to GCs, we hypothesized that administration of the GC receptor antagonist RU486 (mifepristone) should alter aGVHD-mediated thymocyte apoptosis. METHODS: Thymic development in the presence of aGVHD was studied in a haploidentical nonirradiated murine transplantation model (C57BL/6-->B6D2F1). Recipients were treated with RU486 or vehicle alone. Thymic development was analyzed by flow cytometry at different times post transplant. RESULTS: Acute thymic GVHD was characterized (1) by infiltration of mature donor-derived T cells and (2) by increased apoptosis of immature CD4+CD8+ thymocytes between 1 and 2 weeks after allogeneic transplantation. Contrary to expectations, administration of RU486 had no effect on these two parameters. CONCLUSIONS: Our data suggest that thymic pathology during aGVHD is mediated via a glucocorticoid-independent mechanism of apoptosis as blockade of glucocorticoid receptors did not alter the GVHD-induced thymic phenotype. PMID- 10852623 TI - Interaction between nelfinavir and tacrolimus after orthoptic liver transplantation in a patient coinfected with HIV and hepatitis C virus (HCV). AB - A 49-year old male patient with severe hemophilia A, coinfected with HIV and HCV, who underwent orthoptic liver transplantation because of hepatitis C cirrhosis is presented. We describe a strong interaction between nelfinavir and tacrolimus postoperatively, that caused a reduction of the dose of tacrolimus by a factor 70 compared with normal, to achieve therapeutic blood concentrations and to avoid toxic side effects. We suggest that nelfinavir inhibits the metabolism of tacrolimus because both compounds are well-known substrates for the cytochrome P450 isoenzyme CYP 3A4. The nelfinavir serum concentrations were not affected by the institution of tacrolimus. Although the interaction dramatically changed the tacrolimus dose-concentration relationship, the situation was manageable by frequent monitoring of blood concentrations of tacrolimus. PMID- 10852624 TI - Percutaneous portal vein thrombolysis and endovascular stent for management of posttransplant portal venous conduit thrombosis. AB - BACKGROUND: Thrombosis of a portal vein conduit after liver transplant is an uncommon clinical situation. Percutaneous thrombolytic therapy for this condition has not been widely described. METHODS: We describe a case of thrombosis of a portal vein (PV) conduit subsequent to orthotopic liver transplantation that was successfully treated by percutaneous portal vein thrombolysis by using tissue plasminogen activator, angioplasty, and endovascular stent placement. RESULTS: A satisfactory outcome was achieved with a patent portal vein, on ultrasound, at 8 month follow-up. CONCLUSION: A percutaneous transhepatic approach to treatment of thrombosis of a portal vein conduit appears to be a promising technique to use to avoid surgery, with good medium-term results. PMID- 10852625 TI - Parvovirus B19 in kidney transplant patients. AB - Renal transplant patients were screened for the presence of parvovirus B19, before transplantation and monthly for 4 months after transplantation, by means of a sensitive nested PCR assay. Upon screening plasma from 110 patients, we found that two asymptomatic patients were B19 DNA positive. One of these patients was PCR positive in the plasma sample taken 2 months after transplantation; the plasma contained anti-B19 IgG antibodies before transplant and throughout the follow-up period, with an increase in the IgG level in the second posttransplant sample coinciding with the detection of B19 DNA. IgM antibodies to B19 were not detected in this patient. Because, for this patient, the donor's spleen DNA was also B19 DNA positive, we suspect B19 transmission from the donor and limited B19 replication, inasmuch as this patient already had a primed immune response to B19. The other patient was PCR positive in the pretransplant and in the plasma sample taken 1 month after transplant and contained a strong anti-B19 IgG response in the pretransplant sample and throughout the follow-up period-and anti B19 IgM antibodies were not detected before or after transplantation. By testing samples taken from this patient at 2 weeks, 2 months, and 3 months before transplantation, we were able to determine that the infection occurred shortly before transplantation. Unexpectedly, this graft failed and was removed 2 days after transplantation despite a negative cross-match. A pathological examination of the kidney indicated acute vascular rejection, suggesting a possible role for B19 in this complication. PMID- 10852626 TI - Emergency adult to adult living donor liver transplantation for fulminant hepatic failure. AB - BACKGROUND: The high mortality rate associated with fulminant hepatic failure combined with the limited availability of cadaveric organs requires consideration of alternatives to conventional cadaveric transplantation. Use of the donor right lobe in adult-to-adult living donor transplantation holds promise in a variety of circumstances, including high-acuity situations. METHODS: A 28-year-old male with fulminant hepatic failure secondary to hepatitis B was referred to our institution. He rapidly progressed to grade IV encephalopathy, and laboratory values were indicative of a poor prognosis without transplantation. He was listed for transplantation as UNOS status I. Three siblings were simultaneously evaluated for living liver donation. Following established protocols, we completed donor evaluation in less than 24 hr, and donor right lobectomy and living donor transplantation were performed within 36 hr of the recipient's admission to our center. RESULTS: The donor surgery was uncomplicated, and the patient was discharged on postoperative day 4. The recipient experienced full recovery and was discharged home on postoperative day 14. Of note, the first offer for a cadaveric liver came more than 60 hr after living donor transplantation. CONCLUSIONS: Thorough donor workup can be completed in less than 24 hr without inappropriate abbreviation of the evaluation. Simultaneous workup of willing individuals prevents unnecessary delay. Living donor transplantation should be considered for patients with fulminant hepatic failure who are appropriate transplant candidates. PMID- 10852627 TI - Effect of liver transplantation on hepatic glucose metabolism in a patient with type I glycogen storage disease. AB - BACKGROUND: In type I glycogenosis, mutation of the glucose-6-phosphatase gene results in absent glucose-6-phosphatase activity in liver cells leading to fasting hypoglycemia. Liver transplantation is expected to normalize glucose homeostasis. METHODS: Endogenous glucose production (6,6 2H2 glucose) was measured after an overnight fast and during exogenous 13C-labeled glycerol infusion in a patient with glycogenosis type I 24 months after liver transplantation and in a group of healthy subjects. RESULTS: Compared with healthy subjects, the glycogenosis patient had normal fasting glucose production and glucose and insulin concentrations after liver transplantation, but mildly elevated plasma glucagon concentrations. Gluconeogenesis from exogenous glycerol (13C glucose synthesis) was similar and did not lead to enhancement of glucose production in both healthy controls and the patient. CONCLUSIONS: Liver glucoregulatory function is restored by orthotopic liver transplantation in type I glycogenosis. PMID- 10852628 TI - Sialyl Lewis(x) (CD15s) monitoring as a means to select antirejection therapy in patients with rejection after renal transplantation: CD15s monitoring for treatment and diagnosis in patients with acute rejection after renal transplantation. AB - BACKGROUND: Sialyl Lewis(x) (CD15s), which is known to serve as a ligand for the cell adhesion molecules E-selectin and P-selectin, is expressed on peripheral lymphocytes in renal transplant patients with rejection. In this study, we examined whether CD15s monitoring could differentiate the patients with rejection in whom steroids were effective from those in whom steroids were not effective. METHODS: To investigate CD15s expression on peripheral lymphocytes, flow cytometry was performed before and after steroid pulse therapy in 20 recipients with rejection after renal transplantation, including 5 recipients resistant to steroid therapy. We also compared CD15s expression with pathological findings before and after steroid pulse therapy. RESULTS: CD15s expression was stronger before steroid pulse therapy in the 5 patients resistant to steroids than in the 15 patients responsive to steroid treatment. In addition, CD15s expression remained high without any pathological improvement in the 5 patients resistant to steroids after steroid treatment, although CD15s recovered to normal levels with remarkable improvement of pathological findings in the other 15 patients. Five patients in whom steroids were not effective, had full recovery of serum creatinine levels as well as CD15s expression after muromonab (OKT3) therapy. CONCLUSIONS: CD15s monitoring might help clinicians to select antirejection therapy. PMID- 10852629 TI - Laparoscopic donor nephrectomy increases the supply of living donor kidneys: a center-specific microeconomic analysis. AB - BACKGROUND: A tenet of microeconomics is that new technology will shift the supply curve to the right. Laparoscopic donor nephrectomy (LDN) is a new technique for removal of living donor kidneys. Centers performing this procedure have noted an increased number of patients presenting for donor evaluation. This has not been previously studied. METHODS: The records of all LDN performed from May 1998 to February 1999 were reviewed. The following variables were examined: sex, age, related vs. unrelated donation, estimated blood loss, i.v. analgesia, length of stay, and time out of work. Donors undergoing traditional open donor nephrectomy during January 1997 to May 1998 served as the control group. A composite cost index was constructed. RESULTS: LDN significantly decreased length of stay, pain, and time out of work; the supply function shifted to the right. Telephone interviews revealed that 47% donated solely because of the LDN procedure. CONCLUSIONS: LDN increases the supply of living donor kidneys. PMID- 10852630 TI - Spontaneous rupture of the liver upon revascularization during transplantation. AB - Spontaneous rupture of the liver has been described in association with many benign and malignant conditions. We report, to our knowledge, the first case of spontaneous rupture of the liver upon revascularization, requiring total hepatectomy and portocaval shunt, followed by successful retransplantation. Routine pathological examination of the explanted liver failed to reveal the etiology of the rupture. However, electron microscopy demonstrated abnormal collagen in the hepatic arterial wall compatible with a collagen disorder such as Ehlers-Danlos type IV disease. We conclude that the donor liver had a previously undiagnosed collagen disorder. Review of the literature does not preclude the use of livers from donors with a history of connective tissue disorders. Based on our experience one should exercise caution when using livers from such donors. With a history of connective tissue disorder in an immediate family member, further tests should be performed in the donor to rule out a subclinical connective tissue disorder. In addition, a review of all patients reported thus far to have undergone total hepatectomy and portocaval shunt, followed by liver transplantation as a two-stage procedure is presented. PMID- 10852631 TI - Emergency portacaval shunt for control of hemorrhage from a parenchymal fracture after adult-to-adult living donor liver transplantation. AB - As more adults undergo transplantation with partial liver grafts, the unique features of these segments and their clinical significance will become apparent. A patient presented with life-threatening hemorrhage from an iatrogenic laceration to a right lobe graft 11 days after transplantation. The creation of a portacaval shunt effectively controlled the bleeding, allowing more elective replacement of the organ with another right lobe graft. The regeneration process combined with increased portal blood flow and relative outflow limitation may have set the stage for this complication. Any disruption of the liver parenchyma during transplantation should be securely repaired and followed cautiously. Portacaval shunting is an option for controlling hemorrhage from the liver in transplant recipients. The timely availability of a second organ was likely the ultimate determinant of survival for this patient. PMID- 10852632 TI - Cholesterol embolization in renal allografts. AB - Renal cholesterol embolization (RCE) in native kidneys has a dismal outcome and frequently leads to irreversible renal failure. RCE may rarely occur in renal allografts as well, particularly if the recipient or the donor has prominent atherosclerosis. The natural history of RCE in renal transplants is unknown. We have reviewed the surgical pathology files of The Johns Hopkins Hospital in the 14-year period between 1984 and early 1999 and found 7 RCE cases among 1500 renal transplant biopsies (0.47%). One of the seven cases had three biopsies showing cholesterol emboli, the first of which was a postreperfusion (immediate posttransplant) biopsy. The probable source of the cholesterol emboli was the recipient in six cases and the donor in one case. Five donors were cadaveric and two were living donors. Six biopsies were taken within the first 4 months posttransplant (four were postreperfusion biopsies). One recent patient had the inciting event of arteriography and stent placement 2 years posttransplant and is currently doing well. One kidney failed due to posttransplant lymphoproliferative disorder (PTLD), another kidney failed with complicating opportunistic infections, and the other five were functioning 2 to 6 years posttransplant. A literature review revealed additional 14 RCE cases in renal transplants. Combining our cases with those in the literature (21 cases), reveals that the origin of the RCE was probably the recipient in 11 cases (seven cadaveric, two living-related, and two unknown), and the donor in 10 cases (eight cadaveric and two unknown). Graft failure occurred in two of the 11 cases, where RCE was of probable recipient origin. Seven of the 10 kidneys, where the RCE was probably of donor origin, failed due to allograft dysfunction; one of them also developed superimposed rejection and cytomegalovirus infection. We conclude that if RCE is originating in the recipient, graft survival is usually good. In contrast, if RCE is of donor origin, graft dysfunction and subsequent graft loss are common. The reason for this difference may be the more extensive RCE developing in an atherosclerotic cadaveric donor during organ procurement or severe trauma leading to death. PMID- 10852633 TI - Evaluation of a multivariate model predicting noncompliance with medication regimens among renal transplant patients. AB - BACKGROUND: Because noncompliance with medication regimens is a major cause of renal allograft failure, we evaluated the stability over time of two logistic regression models (sets of variables) that predict noncompliance with immunosuppressive regimens. METHODS: Models were based on questionnaire data from 1402 patients (all over 18, receiving cyclosporine or a cyclosporine-like replacement drug, and with a functioning renal graft). The same questionnaire was completed by a subset of 548 (39.1%) patients approximately 18 months later. The goodness of fit of each model to the new data set was tested. RESULTS: The noncompliance logistic regression model including patient beliefs as well as patient and transplant characteristics was an excellent fit to the second data set. A noncompliance model composed of only patient and transplant characteristics fit the new data set less well. CONCLUSIONS: Clinicians and educators need to take explicit account of renal transplant patients' attitudes when evaluating risks of noncompliance and when developing interventions and educational programs to minimize noncompliance. PMID- 10852634 TI - Profound drop of cyclosporin A whole blood trough levels caused by St. John's wort (Hypericum perforatum) PMID- 10852635 TI - Iatrogenic Kaposi's sarcoma: HHV8 positivity persists but the tumors regress almost completely without immunosuppressive therapy. PMID- 10852636 TI - Contralateral inflow occlusion to optimize graft volume and to reduce blood loss during parenchymal dissection in living-related liver transplantation. PMID- 10852637 TI - The genomics of cardiovascular disorders: therapeutic implications. AB - Cardiovascular disease (CVD) is a complicated series of disorders that result from the interaction between genetic predisposing mechanisms and environmental factors. Over the last few years substantial progress has been made in defining the molecular basis of several genetically transmitted non-atherosclerotic CVD such as hypertrophic and dilated cardiomyopathies, long-QT syndrome and essential hypertension. This review represents a summary of the current knowledge about the major gene polymorphisms found to be associated with these CVDs. Moreover, we will discuss how the discovery of disease-associated genes will greatly enhance the ability to formulate advanced diagnoses, to define prophylactic therapeutic strategies to prevent or reduce the progression of the disease and, finally, to proceed to the development of new drugs tailored for the specific cellular or molecular functions altered as consequence of the predisposing genes. PMID- 10852638 TI - Clinical potential of matrix metalloprotease inhibitors in cancer therapy. AB - Matrix metalloproteases (MMP) are a family of enzymes that contribute to the degradation of the extracellular matrix. The destruction of the extracellular matrix eventually leads to tumour invasion, metastasis and angiogenesis. Realising this mechanism of action, there is tremendous potential for inhibitors of MMP in cancer therapy. Extensive preclinical data have shown that administration of matrix metalloprotease inhibitors (MMPI) to different animal models results in a reduction in primary tumour growth as well as in the number and size of metastatic lesions. Based on promising preclinical studies, synthetic MMPI have been developed and taken into clinical trials. These include marimastat, BAY- 129566, CGS-27023A, prinomastat (AG-3340), BMS-275291 and metastat (COL-3). These drugs are all in different stages of clinical development, ranging from phase I to III. In general, musculoskeletal problems, such as joint stiffness and pain in hands, arms and shoulders seem to affect most patients in varying degrees, depending on the dose and type of compound administered. In addition to single agent therapy, several MMPI have entered trials of combination therapy. The objective of combining chemotherapy with an MMPI is to potentiate tumour cytotoxicity as well as to reduce the size and number of metastatic lesions. Several compounds have entered phase III combination therapy trials, but it is still too early to report any data. There is ongoing research in correlating biological endpoints, such as levels of MMP and markers of angiogenesis with clinical response. As the field of MMP and their inhibitors continues to mature, its role in cancer therapeutics will be better defined. PMID- 10852639 TI - Pharmacological management of intermittent claudication: a meta-analysis of randomised trials. AB - Intermittent claudication, a symptom of atherosclerosis in the large vessels of the lower limbs, greatly affects patient mobility and quality of life. Medical therapy for a moderate form of this condition includes vasodilators, antiplatelet agents and alternative treatments such as ginkgo biloba.A meta-analysis of results from 52 trials (including 5088 patients) was conducted for all current medical therapies for intermittent claudication. After 24 weeks, some of the medical therapies were found to be more effective than placebo for the primary end-points of either pain-free walking distance or maximum walking distance. Vasodilators presented the best results in walking distance. Pentoxifylline offered better results than naftidrofuryl, although the treatment benefit, measured in additional metres walked with treatment than without, was modest. Antiplatelets, ginkgo biloba and levocarnitine were slightly more effective than placebo, although the treatment benefit was of limited clinical importance. On average, patients walked 60m further with therapy than without, and only about half of that added distance was pain-free. Very little consistent information was available for other clinical end-points, such as overall mortality and adverse effects. These data suggest that some of the medical therapy, pentoxifylline in particular, can only modestly increase functional status in patients with moderate intermittent claudication. There is a need for uniformity in research design and reporting of trials. A future trial comparing medical therapy with physical therapy is indicated. PMID- 10852640 TI - Pancreatic cancer: a review of emerging therapies. AB - The incidence of adenocarcinoma of the pancreas has risen steadily over the past 4 decades. Since pancreatic cancer is diagnosed at an advanced stage, and because of the lack of effective therapies, the prognosis of such patients is extremely poor. Despite advances in our understanding of the molecular biology of pancreatic cancer, the systemic treatment of this disease remains unsatisfactory. Conventional chemotherapy has not produced dramatic improvements in response rates or patient survival. New treatment strategies are clearly needed. This paper reviews emerging therapies for pancreatic carcinoma. A more profound understanding of the molecular biology of cell growth and proliferation, as well as of neoplastic cell transformation, has led to advances in several areas, including the use of somatostatin analogues and antiandrogens as adjuvant therapy; inhibition of tumour growth and metastasis by inhibitors of matrix metalloproteinases and angiogenesis, and by small molecules such as retinoids, which interfere with progression through the cell cycle; immunotherapy with monoclonal antibodies; disruption of intracellular signal transduction with farnesyltransferase inhibitors; and finally gene therapy with specifically designed vaccines. PMID- 10852641 TI - Pharmacological prophylaxis of post-traumatic epilepsy. AB - Early and late epileptic seizures are a frequent complication of severe head traumas. The administration of anticonvulsant drugs immediately after head injury is commonly implemented as a prophylactic measure; however, there is a lack of consensus on the usefulness of prophylaxis with anticonvulsants for the prevention of late post-traumatic epilepsy (PTE). The inconsistent evidence accumulated so far from clinical studies, most nonrandomised and uncontrolled in design, and the limited knowledge of the processes underlying post-traumatic epileptogenesis, do not warrant empirical pharmacological prophylaxis with long term administration of conventional anticonvulsants. Phenytoin and phenobarbital (phenobarbitone) are used to a large extent in this indication. As a general rule, a benefit/risk analysis in individual patients should drive prophylactic drug prescription in PTE as it can have potential detrimental effects on a patient's recovery. New compounds, such as free-radical scavengers and antiperoxidants, show encouraging experimental results, but their clinical use is still very limited. PMID- 10852642 TI - Diabetic dyslipidaemia: current treatment recommendations. AB - Insulin deficiency and hyperglycaemia in type 1 (insulin-dependent) diabetes mellitus produce lipid abnormalities, which can be corrected by appropriate insulin therapy. Diabetic nephropathy, which is the main risk factor for coronary heart disease (CHD) in type 1 diabetes, causes pro-atherosclerotic changes in lipid metabolism. Detection and treatment of elevated cholesterol levels is likely to be of benefit in these patients. Type 2 (noninsulin-dependent) diabetes mellitus is associated with abnormal lipid metabolism, even when glycaemic control is good and nephropathy absent. Elevated triglyceride levels, reduced high density lipoprotein (HDL) cholesterol and a preponderance of small, dense low density lipoprotein (LDL) particles are the key abnormalities that constitute diabetic dyslipidaemia. The prevalence of hypercholesterolaemia is the same as for the nondiabetic population, but the relative risk of CHD is greatly increased at every level of cholesterol. Based on effectiveness, tolerability and clinical trial results, treatment with HMG-CoA reductase inhibitors to lower LDL cholesterol is recommended as primary therapy. These agents are also moderately effective at reducing triglyceride and increasing HDL cholesterol levels. If hypertriglyceridaemia predominates, treatment with fibric acid derivatives is appropriate, although there is currently only limited clinical trial evidence that the risk of CHD will be reduced. In type 1 diabetes, but particularly in type 2 diabetes, lipid disorders are likely to contribute significantly to the increased risk of macrovascular complications. especially CHD. Management of the disordered lipid metabolism should be given a high priority in the clinical care of all patients with diabetes. PMID- 10852644 TI - Lipid-lowering treatment in coronary artery disease: how low should cholesterol go? AB - The randomised clinical trial data, which supports preventing coronary heart disease (CHD) events by lowering low density lipoprotein cholesterol (LDL-C) levels, is substantial, consistent and highly significant. HMG-CoA reductase inhibitors (statins), which are the preferred medications for lowering LDL-C levels, are well tolerated, with greater efficacy than other lipid-altering medications. In 1993, the National Cholesterol Education Program (NCEP) guidelines recommended LDL-C target levels to be achieved with therapy in high risk individuals. In particular, the LDL-C goal of therapy in patients with CHD was < or = 100 mg/dl (2.6 mmol/L), with no specific guidance as to the lower limit or whether additional clinical benefit could be expected. Because little clinical trial data existed at that time to offer support, and because some epidemiological data raised concern about the potential detriments associated with very low total cholesterol and LDL-C levels, the NCEP Adult Treatment Panel remained appropriately vague on the 'how low should you go' question. In the last few years, several additional clinical trials have provided sufficient efficacy and safety data to re-examine that question. Analyses of on-treatment LDL-C levels and subsequent CHD events from three landmark trials with HMG-CoA reductase inhibitors suggest that progressively lower LDL-C levels are associated with lower CHD events in a curvilinear fashion. The Post Coronary Artery Bypass Graft (Post-CABG) trial and Atorvastatin Versus Revascularisation Trial (AVERT) examined a more intensive versus less intensive drug regimen for LDL-C reduction, and concluded that the more aggressively treated patients had better angiographic and end-point outcomes. Most importantly, there did not appear to be any change in noncardiovascular end-points associated with lower LDL-C levels. In several ongoing clinical trials, patients with CHD have been randomised to receive HMG CoA reductase inhibitors with targets for LDL-C levels of 100 mg/dl versus 75 mg/dl (1.94 mmol/L). These trials have sufficient patient numbers and power to definitely determine if reducing LDL-C levels to approximately 75 mg/dl can provide an acceptable benefit-to-risk-ratio. PMID- 10852643 TI - Treatment of postherpetic neuralgia: an update. AB - Postherpetic neuralgia (PHN) is a chronic pain syndrome that is often refractory to treatment and can last for years, causing physical and social disability, psychological distress, and increased use of the healthcare system. In this paper we provide an update on recent developments in the treatment of PHN. We emphasise the results of recent studies that provide an evidence-based approach for treating PHN that was not available until very recently. In randomised, controlled clinical trials, the topical lidocaine patch, gabapentin, and controlled release oxycodone have been shown to provide superior pain relief in patients with PHN when compared with placebo. It has also recently been demonstrated that the tricyclic antidepressant nortriptyline provides equivalent analgesic benefit when compared with amitriptyline, but is better tolerated. Based on these results, nortriptyline can now be considered the preferred antidepressant for the treatment of PHN, although desipramine may be used if the patient experiences unacceptable sedation from nortriptyline. The topical lidocaine patch, gabapentin and controlled release oxycodone all appear to be as effective as tricyclic antidepressants in the treatment of patients with PHN, and the results of these recent studies suggest that each of these treatments should be considered early in the course of treatment. Additional controlled trials are needed to compare the efficacy and tolerability of these 4 treatments- tricyclic antidepressants, gabapentin, the topical lidocaine patch and controlled release opioid analgesics--used singly and in various combinations in the treatment of patients with PHN. PMID- 10852645 TI - Gemifloxacin. AB - Gemifloxacin is a fluoroquinolone antibacterial agent which has an enhanced affinity for topoisomerase i.v.. It has potent activity against most Gram positive bacteria, particularly Streptococcus pneumoniae. Gemifloxacin is over 30 fold more active than ciprofloxacin and 4- to 8-fold more active than moxifloxacin against this pathogen. Gemifloxacin has excellent activity against Haemophilus influenzae and Moraxella catarrhalis, and is unaffected by beta lactamase production. It is generally 2-fold less active than ciprofloxacin against most Enterobacteriaceae. Atypical respiratory pathogens (Legionella, Mycoplasma and Chlamydia spp.) are highly susceptible to gemifloxacin. Preliminary results from phase II trials show that oral gemifloxacin 320 mg/day produced bacteriological responses of 94.7% in patients with acute exacerbations of chronic bronchitis and 95% of patients with uncomplicated urinary tract infections. Adverse events included nausea, abdominal pain, headache and mild rash in patients and healthy volunteers treated with gemifloxacin 320 mg/day. Gemifloxacin has a low potential for mild phototoxicity (comparable to that of ciprofloxacin). PMID- 10852646 TI - Lisinopril: a review of its use in congestive heart failure. AB - The ACE inhibitor lisinopril is a lysine derivative of enalaprilat, the active metabolite of enalapril. In patients with heart failure, maximum pharmacodynamic effects are produced 6 to 8 hours after administration of the drug and persist for 12 to 24 hours. High doses (32.5 to 35mg, administered once daily) of lisinopril in the Assessment of Treatment with Lisinopril and Survival (ATLAS) study demonstrated clinically important advantages over low doses (2.5 to 5mg, administered once daily) of the drug in the treatment of congestive heart failure. High doses of lisinopril were more effective than low doses in reducing the risk of major clinical events in patients with heart failure treated for 39 to 58 months. Compared with recipients of low doses, those receiving high doses of lisinopril had an 8% lower risk of all-cause mortality (p = 0.128), a 12% lower risk of death or hospitalisation for any reason (p = 0.002) and 24% fewer hospitalisations for heart failure (p = 0.002). These benefits were associated with significant cost savings. In short term (generally 12 weeks' duration) randomised, double-blind, parallel-group, multicentre clinical trials, lisinopril was significantly more effective than placebo and was at least as effective as captopril, enalapril, digoxin and irbesartan at improving symptomatic end-points and clinical status in patients with heart failure. Lisinopril is generally well tolerated by patients with heart failure. In controlled clinical trials, the most common adverse events occurring in recipients of the drug were dizziness, headache, hypotension and diarrhoea. Overall adverse event profiles for patients treated with high or low doses of lisinopril in the ATLAS study were similar. However, high doses of lisinopril used in the ATLAS study were associated with a higher incidence of adverse events, importantly hypotension and worsening renal function; nevertheless, these events were generally well managed by altering the dose of lisinopril or concomitant medications. Furthermore, despite the higher incidence of some adverse events with high doses of lisinopril, the frequency of treatment discontinuations because of adverse events was the same in the high and low dose groups. CONCLUSIONS: Lisinopril (when added to diuretics and/or digoxin) provides symptomatic benefits in patients with congestive heart failure. The ATLAS study demonstrated that high doses of lisinopril significantly reduced the risk of the combined end-point of morbidity and mortality compared with low doses of the drug. Importantly, there was no clinically significant decrease in the tolerability of the drug with use of a high dose. Lisinopril is at least as effective and as well tolerated as other members of the ACE inhibitor class for the treatment of congestive heart failure. PMID- 10852647 TI - Iomeprol: a review of its use as a contrast medium. AB - Iomeprol is a nonionic, monomeric iodinated contrast medium. Unlike the older ionic agents, iomeprol has low chemotoxicity, osmolality and viscosity and high water solubility. Compared with other nonionic contrast media, the osmolality and viscosity are lower and the water solubility is reported to be higher with iomeprol. Most radiographs (about 67 to 100%) obtained with iomeprol (containing 150 to 400 mg/ml of iodine) were of good or excellent quality in noncomparative and comparative trials recruiting 40 to 6127 patients undergoing various radiographic procedures. As expected, the diagnostic efficacy of iomeprol did not differ significantly from that of other nonionic agents (iopamidol, iopromide, iohexol and iotrolan). Iomeprol (containing 150 to 400 mg/ml of iodine) was well tolerated in clinical trials. Most adverse events were transient and of mild to moderate intensity and were similar to those observed with other contrast media. The overall incidence of adverse events ranged from 3 to 49.7% and mainly included localised pain (< or =6%) and heat sensations (8 to 45%), taste disturbances (3 to 27%) and various pseudoallergic reactions (< or =20% for each type of event). The incidence of heat or pain and taste disturbances with iomeprol was similar to that observed with iopromide and iopamidol. Pain (but not heat sensations) was reported significantly less frequently and taste disturbances reported significantly more frequently with iomeprol than with iohexol in a comparative trial. Pseudoallergic reactions (such as nausea, vomiting, skin reactions, dizziness, headache) were significantly less common with iomeprol than with ioxaglate and occurred at a similar frequency to that with iopromide and iopamidol. Cardiovascular events were rarely observed with iomeprol. Currently available iomeprol solutions contain a range of iodine concentrations (150 to 400 mg/ml) and are approved for a wide variety of diagnostic procedures. Iomeprol solutions are chemically stable which negates the need for chelating agents. Formulations of this agent are therefore the first not to contain edetic acid (EDTA). CONCLUSIONS: Iomeprol shows equivalent diagnostic efficacy, and a similar adverse event profile, to that of other nonionic contrast media. The availability of a range of iodine concentrations enables iomeprol to be used in a variety of diagnostic procedures. Iomeprol, like others in its class, is suitable for use in diagnostic imaging. PMID- 10852648 TI - Irbesartan: an updated review of its use in cardiovascular disorders. AB - Irbesartan interrupts the renin-angiotensin system via selective blockade of the angiotensin II subtype 1 receptor; the latter being responsible for the pressor related effects of angiotensin II. As treatment for mild to moderate hypertension, irbesartan 150 mg/day controlled diastolic BP in 56% of patients according to pooled data from several phase III studies and 77% of patients in a large phase IV study. in comparative trials, irbesartan was significantly more effective than losartan and valsartan as treatment for mild to moderate essential hypertension and as effective as enalapril or atenolol. Results from many studies show an additive antihypertensive effect when hydrochlorothiazide is added to irbesartan monotherapy. The drug also induces statistically significant regression of left ventricular mass in patients with hypertension and left ventricular hypertrophy, and preliminary evidence suggests it has beneficial haemodynamic effects in patients with heart failure. Irbesartan is very well tolerated, exhibiting an adverse event profile similar to that seen with placebo in comparative trials. In conclusion, although the role of irbesartan as a treatment for heart failure is little clearer than it was 2 years ago, the place of the drug in the management of hypertension is now better established. There is evidence to suggest the drug may have a role as initial therapy for hypertension, although formal recommendation in management guidelines will almost certainly not occur until long term morbidity and mortality benefits are established. PMID- 10852649 TI - Fibrous scar in the infrapatellar fat pad after arthroscopy: MR imaging. AB - PURPOSE: We describe the MR appearance of fibrous scars in the infrapatellar fat pad after arthroscopy. MATERIALS AND METHODS: The subjects were 96 patients who underwent arthroscope-assisted anterior cruciate ligament (ACL) reconstruction and were examined by oblique sagittal MR imaging at different follow-up intervals. Two observers evaluated the characteristics of the fibrous scars in the infrapatellar fat pad. RESULTS: All fibrous scars with low signal intensity were accentuated at the portal and coursed horizontally through the infrapatellar fat pad. The fibrous scar within the fat pad occurred and peaked within 6 months after arthroscopy. It then subsided gradually and had disappeared by one year later in nearly half of the patients. CONCLUSION: Identifying MR imaging characteristics of fibrous scars in the fat pad after arthroscopy may be clinically helpful to differentiate these scars from other abnormalities that involve the infrapatellar fat pad. PMID- 10852650 TI - Calculation of T2 relaxation time from ultrafast single shot sequences for differentiation of liver tumors: comparison of echo-planar, HASTE, and spin-echo sequences. AB - PURPOSE: The purpose of this study was to evaluate the accuracy of T2 calculation from single shot imaging sequences such as echo-planar imaging (EPI) and half Fourier single shot turbo spin-echo (HASTE) imaging. MATERIALS AND METHODS: For the phantom study, we prepared vials containing different concentrations of agarose, copper sulfate, and nickel chloride. The temperature of the phantom was kept at 22 degrees C. MR images were obtained with a 1.5-Tesla superconductive magnet. Spin-echo (SE) -type EPI and HASTE sequences with different TEs were obtained for T2 calculation, and the T2 values were compared with those obtained from the Carr-Purcell-Meiborm-Gill (CPMG) sequence. The clinical study group consisted of 30 consecutive patients referred for MR imaging to characterize focal liver lesions. A total of 40 focal liver lesions were evaluated, including 25 primary or metastatic solid masses and 15 non-solid lesions. Single shot SE type EPI and HASTE were both performed with TEs of 64 and 90 msec. RESULTS: In the phantom study, the T2 values obtained from both single shot sequences showed significant correlations with those from the CPMG sequence (T2 on EPI vs. T2 on CPMG: r=0.98, p<0.01; T2 on HASTE vs. T2 on CPMG: r=0.99, p<0.01). In the clinical study, mean T2 values for liver calculated from EPI (42 msec) were significantly shorter than those calculated from the HASTE sequence (58 msec) (p<0.001). Mean T2 values for solid tumors were 95 msec with HASTE and 72 msec with EPI, and mean T2 values for non-solid lesions were 128 msec with HASTE and 159 msec with EPI. Although mean T2 values between solid and non-solid lesions were significantly different for both EPI and HASTE sequences (p=0.01 for HASTE, p<0.001 for EPI), the overlap of solid and non-solid lesions was less frequent in EPI than in HASTE. CONCLUSION: With single shot sequences, it is possible to obtain the T2 values that show excellent correlation with the CPMG sequence. Although both HASTE and EPI are useful to calculate T2 values, EPI appears to be more accurate in characterizing focal liver lesions. PMID- 10852651 TI - Radiation therapy for metastatic spinal tumors. AB - PURPOSE: The results of radiation therapy for metastatic spinal tumors were evaluated in terms of pain relief, improvement of neurological impairment, and survival. MATERIALS AND METHODS: Between 1986 and 1995, 52 symptomatic patients with metastatic spinal tumors treated with radiation therapy were evaluated. The patients all received irradiation of megavoltage energy. Therapeutic efficacy was evaluated in terms of pain relief and improvement of neurological impairment. RESULTS: Pain relief was observed in 29 (61.7%) of 47 patients with pain. Therapy was effective for 17 (70.8%) of 24 patients without neurological impairment, and efficacy was detected in 12 (52.2%) of 23 patients with neurological impairment. Improvement of neurological symptoms was obtained in seven (25.0%) of 28 patients with neurological impairment. CONCLUSION: Radiation therapy was effective for pain relief in patients with metastatic spinal tumors. In patients with neurological impairment, less pain relief was observed than in those without impairment. Improvement of neurological impairment was restricted, but radiation therapy was thought to be effective in some cases in the early stage of neurological deterioration. Radiation therapy for metastatic spinal tumors contraindicated for surgery was considered effective for improvement of patients' activities of daily living. PMID- 10852652 TI - Detection of minute pleural fluid in the pleural retracted space associated with peripheral lung cancer: evaluation with MR imaging. AB - PURPOSE: To evaluate the efficacy of magnetic resonance imaging (MRI) in detecting minute pleural fluid in the pleural retracted space (PRS) associated with peripheral lung cancer. MATERIALS AND METHODS: Our subjects were 20 patients with peripheral lung cancer in whom thin-section CT in the lung window setting demonstrated lesions adjacent to the pleural surface, and who were referred for MR imaging. The imaging findings were retrospectively evaluated and correlated with the histopathologic specimens. Pleural fluid was aspirated for cytology under ultrasound (US) guidance in six patients. RESULTS: STIR MR images revealed water SI areas beneath the chest wall associated with the lung cancer, whereas, on CT images, lung cancer and minute pleural fluid in the PRS showed similar soft tissue density without enabling easy differentiation. Two of the six patients who underwent aspiration cytology showed malignancy. All histopathologic specimens obtained from 18 patients who underwent surgery showed pleural retraction corresponding to the water SI areas on STIR images. Histopathological study revealed that the fibrotic focus of the tumor tended to occur more intensively when the shape of pleural retraction was thinner and deeper. CONCLUSION: Water SI areas on STIR images were thought to suggest pleural fluid retention in the PRS. MRI was sensitive in detecting minute pleural fluid in the PRS and may help to avoid overdiagnosis of chest wall invasion induced from peripheral lung cancers. PMID- 10852653 TI - A preliminary study of discrimination among the components of small pulmonary nodules by MR imaging: correlation between MR images and histologic appearance. AB - PURPOSE: To investigate whether magnetic resonance (MR) imaging depicts the internal characteristics of small pulmonary nodules. METHODS: We reviewed MR images of 39 surgically resected pulmonary nodules 3 cm or less and compared the components within the nodules. In 22 malignant nodules, eight histologic components were characterized by signal and enhancement patterns on MR images. RESULTS: MR images obtained from any single sequence discriminated all components in 26 (67%) nodules, whereas the combination of images from various sequences allowed discrimination in 35 (90%). Fourteen of 16 components of aggregated tumor cells showed marked early enhancement. Although fibrotic and necrotic components showed no or slight early enhancement, nine of 10 fibrotic components showed hypointensity and six of seven necrotic components showed hyperintensity on T2 weighted images. Component characterization in eight histologies by MR imaging was possible in 71-100%. CONCLUSION: Our study demonstrated that MR imaging offers the possibility of high tissue-contrast resolution in small pulmonary nodules. PMID- 10852654 TI - Delayed adverse reactions to iodinated contrast media and their risk factors. AB - PURPOSE: The present prospective survey was performed to obtain information on delayed adverse reactions (DARs) to five types of low-osmolar iodinated contrast media, including their frequency, common manifestations, and the patient's history of allergy. METHODS: We investigated data from 15,890 consecutive patients who underwent contrast-enhanced computed tomography (CT) during a 15 month period. All patients were given a questionnaire asking about the occurrence of DARs, their symptoms and duration, and asked to consult a dermatologist if they had a skin reaction. RESULTS: Of 11,121 patients who returned the questionnaire (response rate, 70.0%), DARs were observed in 1,058 patients (9.5%). DARs tended to occur with higher incidence in patients with no previous history of examinations using contrast media, with past adverse reactions caused by contrast media, with a history of allergy, or with a serum creatinine level greater than 2.0 mg/dl. Among the 331 patients who reported skin reactions, 41 patients consulted a dermatologist. Skin reactions were observed significantly more frequently in patients for whom iotrolan was used, and 60% of these reactions were severe or moderate. CONCLUSIONS: Four risk factors for DARs were identified in the present investigation. PMID- 10852655 TI - Helical CT imaging of gastric cancer: normal wall appearance and the potential for staging. AB - PURPOSE: To evaluate the CT appearance of the normal gastric wall and the effectiveness of contrast enhanced helical CT for T-staging of gastric cancer. METHODS: For the basic experiment, two resected stomachs with gastric cancer were filled with water and examined by helical CT imaging. For the clinical study, 59 consecutive patients with gastric cancer who had received preoperative helical CT examination and had also been operated on were entered in this study. Helical CT images were evaluated independently by three radiologists without knowledge of histological staging results. RESULTS: The basic examination of a histopathological correlation with CT images revealed that the inner layer with high attenuation corresponded to the mucosa and the muscular layer of the mucosa, the middle layer with low attenuation to the submucosal layer consisting of coarse tissues and containing fatty tissues, and the outer layer with slightly high attenuation to the proper muscle with serosa. The clinical study revealed that the rate of correct diagnosis through consensus reading was 66.1%. CONCLUSION: The entire stomach with a well-stained mucosa can be visualized by contrast enhanced helical CT. However, T-staging of gastric cancer by helical CT did not appear to improve the accuracy of staging. PMID- 10852656 TI - A case of esophageal carcinoma surgically treated after discontinuance of the simultaneous application of radiotherapy and chemotherapy with low doses of CDDP and 5-FU. AB - The patient, a 69-year-old man with esophageal cancer, had a type 2 tumor in the Mt region, accompanied with an ulcer measuring 12 cm in the major axis. In addition, lymph node metastasis, approximately 5x4 cm, was observed in the lesser curvature of the stomach. The clinical stage of the lesion was T3N1M0 (stage III), and simultaneous therapy combining radiotherapy (2 Gy/day) with chemotherapy employing CDDP (6 mg/day) and 5-FU (300 mg/day) was started on October 21, 1996. During treatment, tumor invasion into the gastric walls from lymph node metastasis was observed on endoscopy, and radiotherapy was discontinued at a total dose of 40 Gy to avoid the possibility of bleeding. Surgery was performed on January 8, 1997. Although tumor invasion from lymph node metastasis in the lesser curvature of the stomach was observed in the pancreas, no remaining cancer cells were noted in the primary nest and metastasized lymph node, suggesting the usefulness of the simultaneous combined therapy. PMID- 10852657 TI - Radiation pneumonitis following multi-field radiation therapy. AB - The mechanisms of radiation pneumonitis have not been established. In a study on multi-field radiation therapy for lung cancer, one patient developed severe radiation pneumonitis even though the target volume was small. Radiation therapy was performed at a dose of 75 Gy in 50 fractions over five weeks. High-density areas conforming to the radiation field were observed by high-resolution CT. They were observed in the irradiated volume at doses under 20 Gy in the contralateral lung as well as in the ipsilateral lung. PMID- 10852658 TI - A case of jugular foramen chordoma with extension to the neck: CT and MR findings. AB - We describe a case of jugular foramen chordoma with extracranial extension into the carotid, parapharyngeal, and perivertebral spaces through the jugular foramen. Although this type of extension is unusual, the differential diagnosis of head and neck tumors includes unusual extension of chordoma as well as extension of meningioma and neurinoma from the posterior skull base. PMID- 10852659 TI - MR imaging of mesenteric hemangioma: a case report. AB - A 62-year-old woman presented with a mobile abdominal palpable mass. She underwent MR examination twice. Because of the mobility of the mass, it was out of the field of view on the first MR examination. The second MR examination detected the mass, which showed heterogeneous signal intensity including low and high intensity on T2-weighted spin echo images. The mass, which was cavernous hemangioma with old hemorrhage, was difficult to differentiate from fibroma or thecoma of the ovary or subserosal leiomyoma of the uterus. PMID- 10852660 TI - Real-time interactive MR imaging system: sequence optimization, and basic and clinical evaluations. AB - A real-time interactive MR imaging system (real-time MRI) is an MR scanner which has a fast image updating cycle and the ability to freely change slice orientation, just like an ultrasound imaging system. Recently, such a system has been developed and installed on a clinical 1.5-Tesla system. The purpose of this study was to optimize the pulse sequences for clinical use and to evaluate the clinical usefulness and basic functionality of real-time MRI. For T1-weighted imaging, FLASH (fast low angle shot) can be selected, and up to 5 frames per second can be acquired depending on the matrix size. For T2-weighted imaging, true FISP (fast imaging with steady-state precession) can be selected, and up to 4 frames per second can be acquired. Maximum C/N between liver and spleen was obtained at a flip angle of 20 degrees on FLASH. Maximum C/N between cardiac cavity and wall was obtained at a flip angle of 60 degrees on true FISP. Localization of the right and left coronary arteries could be performed within 30 seconds in three volunteers. Although the present real-time MRI system has drawbacks such as low spatial resolution and relatively low contrast resolution, we expect real-time MRI to be one of the most important tools for future clinical MRI. PMID- 10852661 TI - Physicians' recommendations for patients who undergo noncardiac surgery. AB - OBJECTIVE: To investigate how consulting physicians attempt to modify perioperative cardiac risk for patients who undergo noncardiac surgery by comparing the preoperative cardiac recommendations of consulting physicians in 2 university centres. DESIGN: Retrospective cross-sectional analysis. SETTING: Five hospitals affiliated with 2 Canadian universities. PATIENTS: Three hundred and eight preoperative consultations were evaluated in 297 patients who were 40 years of age or older and scheduled for noncardiac surgery. OUTCOME MEASURES: Cardiac drug recommendations at the preoperative consultation [corrected]; overall recommendations and practice variation between the 2 centres. RESULTS: The greatest changes in drug management suggested by consultants were the initiation of nitrates in 13% of the patients and a decrease in acetylsalicylic acid administration from 27% to 17%. Centre A physicians recommended adding an angiotensin-converting enzyme inhibitor 11% of the time, whereas centre B physicians recommended such an inhibitor in only 1% of the patients (p = 0.001). In patients taking acetylsalicylic acid at the preoperative consultation, Centres A and B physicians recommended withholding the drug 47% and 22% of the time, respectively (p = 0.03). These differences persisted between the 2 centres after controlling for physician estimates of risk. CONCLUSIONS: Consultants frequently recommended perioperative changes in the use of cardiac medications, and there were differences in practice patterns between the 2 centres. These differences may be affecting patient outcomes and highlight the need for randomized clinical trials to determine the impact of perioperative drug administration on bleeding, myocardial infarction and death. PMID- 10852662 TI - Chelation therapy in the JCR:LA-cp rat: experimental assessment of a putative antiatherosclerotic treatment. AB - OBJECTIVE: To test the efficacy of chelation therapy, an alternative medical treatment, as an antiatherosclerotic procedure, using an animal model of insulin resistance and vascular disease. DESIGN: A prospective animal experiment with procedures modelled on human chelation treatments. SUBJECTS: The JCR:LA-cp rat, a strain that, if homozygous for the autosomal recessive cp gene, becomes obese and insulin resistant, with marked hyperinsulinemia and hypertriglyceridemia, and is unique in the spontaneous development of atherosclerosis and ischemic myocardial lesions. EXPERIMENTAL PROTOCOL: Eight-month-old, obese, male JCR:LA-cp rats were fitted with indwelling venous cannulae and infused over 4 weeks with ethylenediaminetetraacetic acid (EDTA) 5 days a week at a daily dose of 40 mg/kg body weight. At the end of the treatment period, samples were taken for assay of blood parameters and for mineral content of bone. The rats were sacrificed and perfusion-fixed for scanning electron microscopy of the aortic arch. RESULTS: Plasma cholesterol concentrations were not changed by the EDTA treatment. In contrast, plasma triglyceride concentrations were raised significantly (74%, p < 0.05). Lean control rats showed minimal abnormality of the aortic arch, whereas the obese control rats had raised intimal lesions, frequent adherent macrophages and endothelial damage. The frequency of these vascular abnormalities in the EDTA treated rats was not different from that seen in the obese controls. The bone contents of calcium and magnesium were not significantly reduced. CONCLUSIONS: Chelation therapy using intravenous EDTA has no beneficial effects on the arterial lesions in the atherosclerotic JCR:LA-cp rat. The increase in plasma triglyceride concentrations would be grounds for concern in human patients. PMID- 10852663 TI - A 2-year prospective cohort study of cardiac resuscitation in a major Canadian hospital. AB - OBJECTIVES: To determine the outcome of cardiopulmonary resuscitation (CPR) for in-hospital cardiac arrest and to identify risk factors associated with survival to the time of hospital discharge. DESIGN: A 2-year prospective cohort study. SETTING: Foothills Medical Centre, a 700-bed tertiary, academic and regional referral centre for Calgary and southern Alberta. PATIENTS: Adult inpatients, excluding those who had cardiac arrest in the Emergency Department or operating room. INTERVENTION: Cardiac resuscitation. MAIN OUTCOME MEASURES: Spontaneous return of the pulse with a minimum systolic blood pressure of 80 mm Hg and survival defined as survival to the time of hospital discharge. RESULTS: In 334 patients there were 390 cardiac arrests, of which 200 were primary cardiac arrests and 39 cardiac arrests that occurred while the resuscitation team was in attendance. Of 239 resuscitated patients, 51 (21.3%) survived. Fifteen variables were identified as being associated with survival. This association could be explained, through multivariate analysis, by the effect of the following 3 variables (odds ratio [OR], 95% confidence interval [CI]): initial observed rhythm other than pulseless electrical activity or asystole (OR 17.34, 95% CI 8.2 to 36.8); a patient who was ambulatory and able to provide self-care (OR 3.8, 95% CI 1.9 to 7.5); and a spontaneous return of circulation with resuscitation in less than 20 minutes (OR 12.9, 95% CI 4.8 to 20.7). CONCLUSIONS: Survival to hospital discharge after cardiac arrest remains static. Initial cardiac rhythm and duration of resuscitation before spontaneous return of circulation were the most important risk factors for survival. These factors and the patient's functional status are relevant when discussing cardiac resuscitation with patients or when considering whether to discontinue resuscitation efforts. PMID- 10852664 TI - Long-term use of angiotensin-converting enzyme inhibitors to modify endothelial dysfunction: a review of clinical investigations. AB - OBJECTIVES: Endothelial dysfunction can be modified by angiotensin-converting enzyme (ACE) inhibitors. The purpose of this paper is to review clinical studies assessing the effect of long-term, oral ACE inhibition on endothelial dysfunction in specific disease syndromes and to identify areas requiring further research. DATA SOURCES: A computer search of the entire MEDLINE database and Current Contents complemented by detailed analysis of references in the papers identified. STUDY SELECTION: Analysis of patients treated on a long-term basis with orally administered ACE inhibitors to modify endothelial function. DATA SYNTHESIS: Studies were identified of patients with hypertension, diabetes, congestive heart failure, coronary artery disease, dyslipidemias and immunoglobulin A nephropathy (IgAN). These studies used diverse endothelium mediated end-points, which included dilatory responses in conduit or resistance vessels, measures of coagulant and fibrinolytic factors, soluble adhesion molecules, endothelin-1, systemic and glomerular barrier functions and renal blood flow. Few trials enrolled large numbers of patients or used randomized, double-blind, placebo-controlled designs. However, consistent and positive effects were noted in patients with coronary artery disease, dyslipidemia or IgAN. In hypertensive patients, conduit artery and renal endothelium-mediated responses could be improved earlier and more easily than resistance vessel function, which appears to require prolonged therapy before improvement is seen. Highly disparate results were found in patients with congestive heart failure or diabetes. CONCLUSIONS: ACE inhibitors appear to improve endothelial dysfunction in patients with coronary artery disease, dyslipidemia, hypertension and IgAN. Conflicting evidence exists in studies of patients with congestive heart failure and diabetes. Further trials are required to clarify and define the prevalence of endothelial dysfunction and the predictors of response in all these conditions. PMID- 10852665 TI - Attaining gender and ethnic diversity in health intervention research: cultural responsiveness versus resource provision. AB - Despite the National Institutes of Health (NIH) mandate to include women and diverse ethnic groups in all NIH-funded research projects, these groups are still excluded as participants in health intervention research. This exclusion has denied them access to state-of-the-art treatments and prevention strategies. making them vulnerable to increased morbidity and mortality and decreased longevity. This article compares two conceptual approaches to inclusion: cultural responsiveness and resource provision. Several issues are raised as to why women and ethnic people of color are not involved in health intervention research. For each of these issues, an appraisal is made as to whether cultural responsiveness or resource provision would more successfully address the problem. It is concluded that cultural responsiveness facilitates participation in research but is not sufficient. An equally important, if not more important, approach may be the provision of resources to empower participants to address problems of access and burden. PMID- 10852666 TI - Othering: toward an understanding of difference. AB - This article proposes a theoretical framework for analyzing how we engage with Others, those perceived as different from self. This engagement, termed Othering, is presented as two particular processes: Exclusionary and Inclusionary. A theoretical framework is developed from a review of the literature and interpretations of completed research exploring the teaching practices of doctorally prepared Latina nursing faculty. Conceptualizing Othering as both exclusive and inclusive processes expands the boundaries for understanding and interacting with those perceived as different. Exclusionary Othering often utilizes the power within relationships for domination and subordination, whereas Inclusionary Othering attempts to utilize power within relationships for transformation and coalition building. The implications of this framework for nursing practice are addressed. PMID- 10852667 TI - Sexual harassment: everyday violence in the lives of girls and women. AB - Sexual harassment is one of the most insidious, yet pervasive, forms of violence that affects all girls, not merely those traditionally thought to be vulnerable or at risk. Although harassment in the workplace has been the focus of considerable attention during the last decade, there is a growing recognition that girls experience varied forms of sexual harassment, and that this behavior begins at a surprisingly early age. This article examines the plight of the "girl child" and presents findings from the first phase of a national action research project currently being conducted by the Canadian Alliance of Five Research Centres on Violence. A major objective of this project is to examine how violence becomes "normalized" in the lives of girls and young women. Implications for nurses, including strategies aimed at encouraging resistance among this population, are addressed. PMID- 10852668 TI - Caring on the ragged edge: nursing persons who are disenfranchised. AB - The narratives of seven nurses who work with clients estranged from mainstream society are interpreted using a phenomenological strategy to reveal the metaphors of going around the wall of fear separating clients and community. Nurses explain four dimensions of meaning experienced through their fearless caring. Ten themes characterize Caring on the Edge; they are organized as three meta-themes: the Human Connection, the Community Connection, and Making Self-Care Possible. PMID- 10852669 TI - Recovering: a process of empowerment. AB - Persons with borderline personality disorder constitute a vulnerable population not only because of the natural history of the disorder, but also because they are frequently stigmatized by persons entrusted with altering the course of the disorder. In this article, the predominant forms of treatment that have been available to persons with borderline personality disorder are reviewed, and it is concluded that they are, albeit unintentionally, oppressive. It is demonstrated how a critical examination of the concept of recovery opens the possibility for delivering care in a fundamentally different way. Grounded in the underlying assumptions of recovery and interpretive phenomenological research, a proposal that moves beyond recovery toward developing and disseminating a practical theory of recovering is presented. PMID- 10852670 TI - Development of a positive professional identity: liberating oneself from the oppressor within. AB - Although oppressed group behavior has been discussed as important for empowering nurses, little has been written about the process of liberation from oppression. One of the major factors that keeps the oppressed from becoming empowered is poor self- and group esteem and identity. This article explores models of positive identity development from other oppressed groups and explores their relevance for nursing. A model is created, based on the other models, which proposes a process for nurses as they begin to understand their oppression and develop more positive images of themselves and other nurses. PMID- 10852671 TI - "Is there a need for blood substitutes in the new millennium and what should we expect in the way of safety and efficacy?". PMID- 10852672 TI - A rapid interferent-free assay for the determination of LD-1 in biological samples. AB - A highly specific enzyme linked immunosorbent assay (ELISA) for the detection of lactate dehydrogenase-1 (LD-1) has been developed. A competition assay is described with polyclonal antibody to LD-1 passively adsorbed on an ELISA 96-well plate, with non-labelled and labelled LD-1 antigen competing. The LD-1 antigen is conjugated to alkaline phosphatase (ALP) using the one step glutaraldehyde method. A linear range of 10-4000 U/L was obtained, and cross-reactivity was only observed with LD-2. It was possible to eliminate this cross-reactivity by carrying out the final incubation step at 65 degrees C. The developed assay was applied to the analysis of spiked serum and plasma samples and the recoveries obtained were acceptable. PMID- 10852673 TI - Factors affecting hepatocyte viability and CYPIA1 activity during encapsulation. AB - Hepatocytes encapsulated in alginate-poly-1-lysine-alginate (APA) are used in transplantation studies and in bioartificial liver support systems. Loss of cell viability in the process of APA encapsulation is usually 20-30% while the effect on cytochrome CYP450 activity is rarely reported. This work investigates the negative influences on hepatocyte viability and CYPIA1 activity during APA encapsulation, and reports methods to alleviate these influences by incorporating certain reagents into the encapsulation solution. The results show that loss of hepatocyte viability and CYPIA1 activity was caused almost entirely by extracellular calcium toxicity rather than by mechanical damage (p < 0.05). Use of 10 mM instead of 100 mM calcium chloride (CaCl2) in the encapsulation process improved CYPIA1 activity (p < 0.05), but did not improve hepatocyte viability (p > 0.05) or result in satisfactory microcapsules. Hepatocyte viability was 25% higher (p < 0.05) in CaCl2 than in calcium lactate (CaLa) when the cells were gelled by contact with these calcium solutions at room temperature (RT). Hepatocyte viability showed little improvement by processing at 4 degrees C than at RT in CaCl2 (p > 0.05) but was 23% higher at 4 degrees C than at RT in CaLa (p < 0.05). Calcium used in the process of encapsulation caused cell necrosis rather than apoptosis. Addition of Dulbecco's modified Eagle's medium (containing 10% foetal bovine serum) or 20 mM fructose to the calcium solution did not improve cell survival. However, nifedipine at a final concentration of 25 mM modestly improved hepatocyte survival in solution containing 100 mM CaCl2 (p = 0.003). Glutathione and taurine in certain concentrations showed protective effects against loss of CYPIA1 activity (p < 0.05 and <0.01 respectively). In conclusion, to optimise the use of calcium during the process of encapsulation, CaCl2 is preferred to CaLa and inclusion of nifedipine, glutathione or taurine in 100 mM CaCl2 solution is recommended. PMID- 10852674 TI - Effect of reuse on surface characteristics of balloon angioplasty catheters. AB - Surfaces of reused angioplasty catheter balloons were characterized by scanning electron microscopy (SEM), image analysis, Fourier transform infrared (FT-IR) spectroscopy and X-ray photoelectron spectroscopy (XPS). The surfaces were found to have longitudinal ridges, cracks, wrinkled regions and particulates. Most of the particles were carbon-based, and contained traces of Cl and Si. The average number of particles ranged between 11 and 38 per mm2, while the average particle size ranged from 14 micron to 28 micron. About 70%-90% of the particles were larger then 10 micron which is approximately the diameter of the fine blood capillaries. The particles appeared to be firmly attached to the outer surface of the balloon. FT-IR and XPS analysis revealed the presence of Si, the absence of proteins, and suggested that the surface of the reused balloon catheters was oxidized. The study highlights the large number of particles created and released during angioplasty. PMID- 10852675 TI - Covalently bonded heparin to alter the pericardial calcification. AB - Calcification is the leading cause of failure of a wide spectrum of cardiovascular and non-cardiovascular medical devices. In this study our aim was to immobilize polyethylene glycol (PEG) and heparin on multiple crosslinked bovine pericardium with Glutaraldehyde (GA) and carbodiimide. Grafting of PEG and heparin through an intermediate tissue bound substrate containing aldehyde and imide functional groups showed reduction in calcification. In this experimental protocol, we used Golomb and Wagner's in vitro model for studying pericardial calcification and a diffusion cell with two compartments for evaluating the diffusion of calcium across the BP. The results showed that heparin immobilization on the surface reduces calcification independent of its concentration in the incubating medium. It is conceivable that inactivation of unpaired aldehydic moieties present in pericardium after exposure to GA act as potential site for PEG grafting and imide functionalities of EDC can covalently bind heparin, would be the key step in the prevention of calcification. It is well-established fact that heparin has a potent antithrombotic effect. But the exact role of heparin in the anticalcification process of bioprostheses still remain elusive. PMID- 10852676 TI - pH-responsive swelling behavior of collagen complex materials. AB - This article deals with an natural collagen blended with chitosan, poly(vinyl alcohol)(PVA). We investigated the swelling properties of collagen and collagen complex in the solution with different pH, and the swelling/deswelling behavior of collagen and collagen-chitosan complex when changing the pH of the medium from 5.4 to 1.8 and in the reverse. The results show that the swelling degree of collagen and collagen complex are dependent to pH of the solution, and the swelling behavior of collagen and collagen-chitosan complex are pH-responsive. PMID- 10852677 TI - Encapsulation of Urease and PEG-Urease in erythrocyte. AB - Erythrocytes can be used to entrap drugs, enzymes or other molecules with active properties, with various encapsulation procedures. The carrier is nonimmunogenic, biodegradable, and circulates freely throughout the body. Urease was covalently immobilized on activated methoxypolyethyleneglycol-5000 (PEG-5000) (1:3 molar ratio). Urease and PEG-Urease were encapsulated in erythrocyte (1/1) (v/v) by using slow dialysis methods. To optimize the loading of erythrocyte, the above base procedure was varied to test the effect of some parameters. Dialysis time, dialysis temperature, storage condition for erythrocyte conjugate, Urease and PEG Urease concentration were investigated. PMID- 10852678 TI - Workshop discusses priorities in veterinary helminthology for South Africa. AB - A workshop was held at Onderstepoort to set priorities in veterinary helminthology for South Africa. Representatives from 19 organisations attended. The workshop achieved 2 of the 3 aims set, namely to identify the problems within the field and to develop strategies to address these challenges. The 7 strategies proposed are: motivation, education, therapeutic, worm resistance, animal tolerance, biological control and diagnostic strategies. A follow-up session is being planned to formulate action plans for each sphere. PMID- 10852679 TI - The application of a selenium fertiliser for the correction of marginal deficiencies in grazing sheep. AB - A commercial fertiliser, consisting of a poorly soluble barium selenate core with a coating of highly soluble sodium selenite, was evaluated in 2 trials for the provision of selenium (Se) to grazing sheep. The fertiliser was administered at a level of 1 kg per hectare to 3 of 6 kikuyu paddocks during 1995 and 1996 in Trial 1, while the other paddocks were left untreated. The Se status of SA mutton merino ram lambs, as reflected by whole blood, liver and kidney Se concentrations, was elevated (P < 0.01) for at least 5 months after application of the fertiliser. Whole blood and liver Se concentrations of animals grazing unfertilised control paddocks were indicative of a subclinical Se deficiency at times (<100 ng Se/ml whole blood and <300 microg Se/kg liver dry matter). In Trial 2, 4 of 7 paddocks on which an oat fodder crop was established were treated with the Se fertiliser during 1995 and 1997. The remaining 3 paddocks were left unfertilised as controls. Groups of 10-15 pregnant SA mutton merino ewes were introduced to these paddocks within 2 weeks of parturition. These ewes and their progeny utilised these paddocks for a mean (+/- SD) period of 41 +/- 8 days after parturition. The whole blood Se concentrations of these ewes and their offspring were elevated (P < 0.01) relative to their contemporaries utilising control paddocks. No suggestion of a subclinical Se deficiency was discernible in animals grazing control paddocks, although whole blood Se levels approached 100 ng Se/ml during 1997. The application of Se fertiliser did not result in improvements in ewe reproduction or lamb growth. There was a suggestion of an improvement (P = 0.21) in mean (+/- SE) lamb survival on paddocks receiving Se fertiliser compared to control paddocks (71.5 +/- 4.6% vs 62.2 +/- 5.3% respectively). PMID- 10852680 TI - The sedative and analgesic effects of detomidine-butorphanol and detomidine alone in donkeys. AB - Butorphanol and detomidine constitute an effective combination for sedation and analgesia in horses. This trial was undertaken to assess the effectiveness of this combination in donkeys. The detomidine and butorphanol were given intravenously one after the other. A dose of 10 microg/kg of detomidine and 25 microg/kg of butorphanol was used. Sedation is easily extended by additional doses of butorphanol. The average dose of detomidine was 11.24 microg/kg and that of butorphanol was 28.0 microg/kg. Four donkeys in the detomidine group required additional sedation and analgesia. Detomidine alone did not totally eliminate coronary band pain. Heart rates dropped significantly in the first minute after the injection of the combination. One donkey developed an atrioventricular block, while another developed a sino-atrial block. Four donkeys developed a Cheyne Stokes respiratory pattern. The combination of detomidine and butorphanol is an effective combination for sedation and analgesia of donkeys for standing procedures. PMID- 10852681 TI - A survey on environmental contamination of suburban parks and playgrounds in Harare, Zimbabwe, with canine helminths of zoonotic significance. AB - In an attempt to assess the possible risk to humans of soil-transmitted canine helminths of of zoonotic significance, 161 faecal samples and 81 soil samples were collected from 6 public parks and playgrounds in Harare between March and June 1998 and examined for nematode ova. Of the 161 faecal samples collected, 17.4% were positive for Ancylostoma sp. ova and 5.6% were positive for T. canis ova. No other nematode species ova were found. Over 50% of the faecal samples positive for Ancylostoma sp. ova were 'moist', and this suggests that the moisture content of faeces contributes to the development and survival of this parasite in the environment. Only 3 of the 81 soil samples collected were positive for T. canis. The low levels of contamination of public playgrounds and parks with T. canis ova suggests that environmental contamination might not be important in the aetiology of human toxocarosis in Harare. PMID- 10852682 TI - Observations on some cardiopulmonary effects of midazolam, xylazine and a midazolam/ketamine combination in the goat. AB - Xylazine, midazolam and a midazolam/ketamine combination were administered to 6 goats in a randomised 3-way block design. All goats received all treatments with at least a 7-day interval between treatments. Statistically significant (P < 0.05) changes were observed in some of the measured cardiopulmonary variables for xylazine and midazolam/ ketamine. Xylazine administration resulted in statistically significant decreases in minute volume, arterial partial pressure of oxygen, heart rate and mean arterial blood pressure. The increase in arterial partial pressure of carbon dioxide was not statistically significant. For the midazolam/ketamine combination, the decrease in tidal volume was statistically significant, but not the decrease in minute volume and increase in arterial partial pressure of carbon dioxide. The decrease in the arterial partial pressure of oxygen was also statistically significant. The mean arterial blood pressure for the combination was statistically significantly higher compared to xylazine. The changes in cardiopulmonary variables after midazolam administration were not statistically significant, such as tidal and minute volume, arterial partial pressure of oxygen and carbon dioxide. However, clinically significant effects such as hypoventilation and hypoxia were observed after its administration. The change in mean arterial blood pressure was minimal. PMID- 10852683 TI - Hysterotomy by a colpotomy approach for treatment of foetal mummification in a cow. AB - A 7-year-old Brahman cow was diagnosed as suffering from chronic foetal mummification of unknown aetiology, concurrent cystic ovarian disease, prolapse of the 2nd cervical ring and chronic cervicitis. Repeated treatment with prostaglandin F2alpha and oestrogen failed to resolve the mummification. A hysterotomy was performed via an incision in the dorsolateral vaginal wall. Good exposure of the uterine horn was achieved and mild post-operative complications were observed. Colpotomy can be regarded as an alternative surgical approach to the moderately enlarged bovine uterus. PMID- 10852684 TI - Trans-vaginal oocyte retrieval and subsequent in vitro production of embryos from a cow involuntarily culled. AB - A Holstein cow of high genetic merit, in late lactation (205 days) and diagnosed with salpingitis (after 4 infertile services and veterinary consultation), was subjected to 1 trans-vaginal oocyte collection attempt, prior to slaughter. Of an estimated 10 follicles punctured, a total of 4 cumulus-oocyte complexes were retrieved. These were matured in vitro in a maturation medium for 24 hours. After 24 hours maturation, the oocytes were fertilised in vitro with Percoll-processed frozen/thawed imported semen, of the owner's choice. Fertilisation was achieved in a modified Tyrode's medium. At 18 hours post-insemination, the presumptive zygotes were transferred into culture in vitro in Charles Rosenkran's amino-acid medium and supplemented on Day 4 post-insemination with 10% foetal calf serum. All in vitro procedures were conducted in 50 microl medium droplets, under oil, in a humidified incubator at 38.5 degrees C in 5% CO2 in air. Three of the potential zygotes cleaved and, by Day 7 of culture, these had developed to the morula stage. The embryos were frozen in 1.5 M ethylene glycol and later transferred non-surgically to synchronised Holstein recipient heifers. One morula resulted in the only pregnancy and subsequent birth of a healthy heifer calf. An independent commercial company confirmed parentage through standard blood-typing assays. The genetic salvage of oocytes, for in vitro production of embryos, has potential benefits to the producer. PMID- 10852685 TI - Feline herpesvirus infection in a group of semi-captive cheetahs. AB - Clinical disease caused by feline herpesvirus type-1 in wild felid species is similar to that in domestic cats. Herpesviruses are endemic in free-ranging lions in South Africa but actual clinical disease due to them has not been reported in free-ranging felids. The first reports of feline herpesvirus infection associated with clinical disease in wild felids came from Australia and the USA in 1970. Subsequent reports of clinical disease in cheetahs and other wild felid species were limited to captive animals. This report deals with clinical disease in a group of semi-captive cheetahs in which 18 animals were affected, and included 12 adult males, 4 adult females and 2 subadults. No mortalities occurred in this group, the most common clinical signs being sneezing, nasal discharge and loss of appetite. PMID- 10852686 TI - Feline transfusion practice in South Africa: current status and practical solutions. AB - Blood transfusion therapy is often under-utilised in feline practice in South Africa. However, it is a technique that can be safely and effectively introduced in practice. Cats have naturally occurring allo-antibodies against the blood type that they lack, which makes blood typing, or alternatively cross-matching, essential before transfusions. Feline blood donors must be carefully selected, be disease free and should be sedated before blood collection. The preferred anticoagulant for feline blood collection is citrate-phosphate-dextrose-adenine. Blood can either be administered intravenously or into the medullary cavity, with the transfusion rate depending on the cat's hydration status and cardiac function. Transfusion reactions can be immediate or delayed and they are classified as immunological or non-immunological. Indications, methods and techniques to do feline blood transfusions in a safe and economical way are highlighted. PMID- 10852687 TI - Competency and the practice of nephrology nursing. PMID- 10852688 TI - Patient and family adjustment to kidney transplantation with and without an interim period of dialysis. AB - This exploratory study was designed to examine the process of patient and family adjustment to kidney transplantation and to compare differences between those who had and those who did not have dialysis pretransplant. To capitalize on benefits of longitudinal and cross-sectional designs, a mixed-method approach described by Aaronson and Kingry (1988) was used. Participants were recruited from two major transplant centers in Western Canada. Twenty patients with ESRD and their partners were studied longitudinally at three points: during the period before the kidney transplant assessment phase and then at 3 and 6 months posttransplant. Forty-seven ESRD patients and their partners comprised the cross-sectional sample. Participants comprising the cross-sectional sample provided responses from at least one of the three time points. Patients and/or their partners completed a series of questionnaires focusing on family functioning (Family Inventory of Life Events and Change [FILE], Feetham Family Functioning Survey [FFFS]); support and resources (Family Inventory of Resources for Management [FIRM]); as well as uncertainty (Uncertainty Stress Scale [USS]); sickness impact (Sickness Impact Profile [SIP]); and sense of coherence (Sense of Coherence Scale [SOC]). The findings indicated that patients and their partners were under considerable stress as they underwent kidney transplantation. Illness and family care strains as well as financial strains were significant. The participants expressed low to moderate uncertainty that decreased over time from the pretransplant to the posttransplant period. Patients on dialysis prior to transplant were more physically and psychologically affected than patients pre empted to transplant. Those who were pre-empted to transplant reported minimal impact pretransplant but considerable improvement in their psychological and physical state posttransplant. In conclusion, transplantation without prior dialysis resulted in less physical and psychological impact for patients and their spouses. These findings point to a need for interdisciplinary education and support programs at both the pretransplant and posttransplant phase to help patients and their partners adjust to living with a kidney transplant. Given the beneficial effects of pre-emptive transplantation, emphasis must be placed on increasing the organ donor pool so that more pre-emptive transplants can be conducted. PMID- 10852689 TI - The safety of intravenous iron dextran (Dexferrum) during hemodialysis in patients with end stage renal disease. AB - The National Kidney Foundation recently published guidelines stating that regular use of intravenous iron therapy will prevent iron deficiency and promote better erythropoiesis than oral iron therapy in patients with end stage renal disease (ESRD) who are undergoing hemodialysis. Although intravenous iron dextran has been shown to be clinically effective in maintaining iron stores in such patients, some clinicians are concerned about the incidence of adverse events associated with this mode of iron supplementation. We conducted a retrospective review of adverse events associated with the use of Dexferrum (American Regent Laboratories, Inc., Shirley, NY) in ESRD patients at an outpatient dialysis clinic. During the 6-month study period, only 1 patient out of 62 (1.6%) experienced adverse events (hypotension, chest pain) related to treatment with Dexferrum. No patients developed anaphylactoid reactions. PMID- 10852690 TI - Attitude, self-image, and quality of life of living kidney donors. AB - The purpose of this research was to assess the attitude, self-image, and quality of life of living kidney donors. This research employed an exploratory design. Instruments included Simmons and colleagues' (1977, 1987) measures on donor attitude and self-image, Ferrans and Powers (1992) scale on quality of life, and Cantril's (1965) ladder of life. Social desirability was also measured. Fifty five living kidney donors from one transplant program participated in the research. Donations had been made recently or as long as 25 years ago. The research determined that men were significantly more ambivalent about donating than women. Significantly higher levels of predicted self-esteem and independence were found in African-American donors, those with higher levels of education, and those who had recently donated a kidney. Scores on quality of life were high for all donors, and they expected that their quality of life would improve in the next 5 years. Social desirability scores were high for 65% of the donors. The quality of life of donors is high and similar to other healthy persons from reported research. The findings in the difference in self-esteem and independence between those who donated before and after 1990 as well as the social desirability scores are reasons to conduct further research on living donors. PMID- 10852691 TI - Nursing considerations in support of a patient pursuing his dream. AB - A 30-year-old man with end stage renal disease (ESRD) undergoing thrice weekly hemodialysis set out to achieve a personal goal to ride his bicycle across the United States within 30 days. To support patients in achieving such a goal, nephrology nurses must consider not only nutritional components, but must be prepared to deal with the physical challenges that face patients in the daily struggles struggle to reach their goal. It became a priority and a challenge for the nephrology team to treat this young man as an athlete who also happened to have ESRD, rather than as a dialysis patient who was a cyclist. PMID- 10852692 TI - Tools for analyzing trends in anemia management case study of the anemic patient. AB - An increasing number of dialysis facilities are empowering nurses to coordinate, monitor, and manage most aspects of anemia-related care. Careful analysis of the trends in laboratory values can provide nurses with a valuable assessment tool to guide therapeutic decisions. This article reviews several proven methods for analyzing laboratory trends and uses case studies to illustrate how to interpret these trends to identify potential etiologies that can cause hyporesponse to Epoetin alfa therapy. PMID- 10852693 TI - Benefits of early utilization of intravenous iron. AB - Better anemia management has dramatically improved the lives of many patients with end stage renal disease (ESRD). Nephrology professionals frequently use two tools--erythropoietin and supplemental iron--to manage anemia. The National Kidney Foundation Dialysis Outcomes Quality Initiative (NKF-DOQI) suggests that most ESRD patients will need intravenous (i.v.) iron to optimize their response to erythropoietin. In this report, the author reviews published studies showing that i.v. iron reduces erythropoietin dose requirements, resulting in cost savings. She presents data from her center illustrating that i.v. administration of the newly approved Ferrlecit (sodium ferric gluconate) also improves anemia management and reduces erythropoietin dose requirements. The author reviews studies showing the efficacy of i.v. iron as monotherapy for anemia in ESRD patients. These data support the importance of i.v. iron as an agent to be used alone or in conjunction with erythropoietin in the management of anemia in patients with ESRD. PMID- 10852694 TI - Intravenous iron products. PMID- 10852695 TI - Hectorol: a new vitamin D prohormone. PMID- 10852696 TI - Subclavian vein stenosis revisited. PMID- 10852697 TI - Prophylactic antibiotic lock therapy for hemodialysis catheters. PMID- 10852698 TI - Effective meetings--to meet or not to meet: that is the question. PMID- 10852699 TI - Organ donation: increasing awareness. PMID- 10852700 TI - Does the structure of the water-oxidizing photosystem II-manganese complex at room temperature differ from its low-temperature structure? A comparative X-ray absorption study. AB - Detailed information on room-temperature structure and oxidation state of the Photosystem II (PS II) manganese complex is needed to put mechanistic considerations on solid grounds. Because previously this information had not been available, the tetranuclear manganese complex was investigated by X-ray absorption spectroscopy (XAS) on PS II membrane particles at 290 K. Due to methodical progress (collection of XAS spectra within 10 s or less), significant X-ray radiation damage can be avoided; room-temperature XAS investigations on the PS II in its native membrane environment become feasible. Thus, the ambiguity with respect to the mechanistic relevance of low-temperature XAS results is avoidable. At 290 K as well as at 18 K, the manganese complex in its dark-stable state (S(1)-state) seemingly is a Mn(III)(2)Mn(IV)(2) complex comprising two di mu(2)-oxo bridged binuclear manganese units characterized by the same Mn-Mn distance of 2.71-2.72 A at both temperatures. Most likely, manganese oxidation states and the protonation state of the bridging oxides are fully temperature independent. Remarkably, at room-temperature manganese-ligand distances of 3.10 and 3.65 A are clearly discernible in the EXAFS spectra. The type of bridging assumed to result in Mn-Mn or Mn-Ca distances around 3.1 A is, possibly, temperature-dependent as suggested by distance lengthening upon cooling by 0.13 A. However, mechanistic proposals on photosynthetic water oxidation, which involve the dimer-of-dimers model [Yachandra, V. K., et al. (1993) Science 260, 675-679] are not invalidated by the presented results. PMID- 10852701 TI - Transcriptional properties of genomic transgene integration sites marked by electroporation or retroviral infection. AB - As a possible consequence of their survival strategy, proviruses are predominantly found in transcription-promoting genomic sites. For certain applications, these findings have led to the preferential use of retroviral vectors for the stable integration of transgenes. This study demonstrates that transcription levels of single-copy proviruses, which have been established either by infection or by single-copy transfection (electroporation), are rather comparable. Therefore, electroporation is suggested as an alternative gene transfer route in cases where the use of infectious retroviral vehicles is to be avoided due to safety considerations. A difference between clones derived from these two gene transfer routes concerns the inactivation pattern which, for electroporated clones, is an exclusive property of the low expressers. This difference may be due to the nature of the illegitimate recombination event which is thought to be less invasive if catalyzed by the retroviral integrase. Substantial differences between infection and Ca phosphate-mediated transfection that have been reported earlier are explained by the respective transfection parameters. PMID- 10852702 TI - Synthesis and biophysical properties of arabinonucleic acids (ANA): circular dichroic spectra, melting temperatures, and ribonuclease H susceptibility of ANA.RNA hybrid duplexes. AB - Arabinonucleic acid (ANA), the 2'-epimer of RNA, was synthesized from arabinonucleoside building blocks by conventional solid-phase phosphoramidite synthesis. In addition, the biochemical and physicochemical properties of ANA strands of mixed base composition were evaluated for the first time. ANA exhibit certain characteristics desirable for use as antisense agents. They form duplexes with complementary RNA, direct RNase H degradation of target RNA molecules, and display resistance to 3'-exonucleases. Since RNA does not elicit RNase H activity, our findings establish that the stereochemistry at C2' (ANA versus RNA) is a key determinant in the activation of the enzyme RNase H. Inversion of stereochemistry at C2' is most likely accompanied by a conformational change in the furanose sugar pucker from C3'-endo (RNA) to C2'-endo ("DNA-like") pucker (ANA) [Noronha and Damha (1998) Nucleic Acids Res. 26, 2665-2671; Venkateswarlu and Ferguson (1999) J. Am. Chem. Soc. 121, 5609-5610]. This produces ANA/RNA hybrids whose CD spectra (i.e., helical conformation) are more similar to the native DNA/RNA substrates than to those of the pure RNA/RNA duplex. These features, combined with the fact that ara-2'OH groups project into the major groove of the helix (where they should not interfere with RNase H binding), help to explain the RNase H activity of ANA/RNA hybrids. PMID- 10852703 TI - NMR structure of free RGS4 reveals an induced conformational change upon binding Galpha. AB - Heterotrimeric guanine nucleotide-binding proteins (G-proteins) are transducers in many cellular transmembrane signaling systems where regulators of G-protein signaling (RGS) act as attenuators of the G-protein signal cascade by binding to the Galpha subunit of G-proteins (G(i)(alpha)(1)) and increasing the rate of GTP hydrolysis. The high-resolution solution structure of free RGS4 has been determined using two-dimensional and three-dimensional heteronuclear NMR spectroscopy. A total of 30 structures were calculated by means of hybrid distance geometry-simulated annealing using a total of 2871 experimental NMR restraints. The atomic rms distribution about the mean coordinate positions for residues 5-134 for the 30 structures is 0.47 +/- 0.05 A for the backbone atoms, 0. 86 +/- 0.05 A for all atoms, and 0.56 +/- 0.04 A for all atoms excluding disordered side chains. The NMR solution structure of free RGS4 suggests a significant conformational change upon binding G(i)(alpha)(1) as evident by the backbone atomic rms difference of 1. 94 A between the free and bound forms of RGS4. The underlying cause of this structural change is a perturbation in the secondary structure elements in the vicinity of the G(i)(alpha)(1) binding site. A kink in the helix between residues K116-Y119 is more pronounced in the RGS4 G(i)(alpha)(1) X-ray structure relative to the free RGS4 NMR structure, resulting in a reorganization of the packing of the N-terminal and C-terminal helices. The presence of the helical disruption in the RGS4-G(i)(alpha)(1) X-ray structure allows for the formation of a hydrogen-bonding network within the binding pocket for G(i)(alpha)(1) on RGS4, where RGS4 residues D117, S118, and R121 interact with residue T182 from G(i)(alpha)(1). The binding pocket for G(i)(alpha)(1) on RGS4 is larger and more accessible in the free RGS4 NMR structure and does not present the preformed binding site observed in the RGS4-G(i)(alpha)(1) X-ray structure. This observation implies that the successful complex formation between RGS4 and G(i)(alpha)(1) is dependent on both the formation of the bound RGS4 conformation and the proper orientation of T182 from G(i)(alpha)(1). The observed changes for the free RGS4 NMR structure suggest a mechanism for its selectivity for the Galpha-GTP-Mg(2+) complex and a means to facilitate the GTPase cycle. PMID- 10852704 TI - Solution structure of an RNA duplex including a C-U base pair. AB - The formation of the C-U base pair in a duplex was observed in solution by means of the temperature profile of (15)N chemical shifts, and the precise geometry of the C-U base pair was also determined by NOE-based structure calculation. From the solution structure of the RNA oligomer, r[CGACUCAGG].r[CCUGCGUCG], it was found that a single C-U mismatch preferred being stacked in the duplex rather than being flipped-out even in solution. Moreover, it adopts an irregular geometry, where the amino nitrogen (N4) of the cytidine and keto-oxygen (O4) of the uridine are within hydrogen-bonding distance, as seen in crystals. To further prove the presence of a hydrogen bond in the C-U pair, we employed a point labeled cytidine at the exocyclic amino nitrogen of the cytidine in the C-U pair. The temperature profile of its (15)N chemical shift showed a sigmoidal transition curve, indicating the presence of a hydrogen bond in the C-U pair in the duplex. PMID- 10852705 TI - Simple modifications of the serpin reactive site loop convert SCCA2 into a cysteine proteinase inhibitor: a critical role for the P3' proline in facilitating RSL cleavage. AB - The human squamous cell carcinoma antigens (SCCA) 1 and 2 are members of the serpin family that are 92% identical in their amino acid sequence. Despite this similarity, they inhibit distinct classes of proteinases. SCCA1 neutralizes the papain-like cysteine proteinases, cathepsins (cat) S, L, and K; and SCCA2 inhibits the chymotrypsin-like serine proteinases, catG and human mast cell chymase. SCCA2 also can inhibit catS, as well as other papain-like cysteine proteinases, albeit at a rate 50-fold less than that of SCCA1. Analysis of the mechanism of inhibition by SCCA1 revealed that the reactive site loop (RSL) is important for cysteine proteinase inhibition. The inhibition of catS by a mutant SCCA2 containing the RSL of SCCA1 is comparable to that of wild-type SCCA1. This finding suggested that there were no motifs outside and only eight residues within the RSL that were directing catS-specific inhibition. The purpose of this study was to determine which of these residues might account for the marked difference in the ability of SCCA1 and SCCA2 to inhibit papain-like cysteine proteinases. SCCA2 molecules containing different RSL mutations showed that no single amino acid substitution could convert SCCA2 into a more potent cysteine proteinase inhibitor. Rather, different combinations of mutations led to incremental increases in catS inhibitory activity with residues in four positions (P1, P3', P4', and P11') accounting for 80% of the difference in activity between SCCA1 and SCCA2. Interestingly, the RSL cleavage site differed between wild-type SCCA2 and this mutant. Moreover, these data established the importance of a Pro residue in the P3' position for efficient inhibition of catS by both wild-type SCCA1 and mutated SCCA2. Molecular modeling studies suggested that this residue might facilitate positioning of the RSL within the active site of the cysteine proteinase. PMID- 10852706 TI - Inhibiting protein-protein interactions: a model for antagonist design. AB - Protein-protein interactions (PPI) are a ubiquitous mode of transmitting signals in cells and tissues. We are testing a stepwise, generic, structure-driven approach for finding low molecular weight inhibitors of protein-protein interactions. The approach requires development of a high-affinity, single chain antibody directed specifically against the interaction surface of one of the proteins to obtain structural information on the interface. To this end, we developed a single chain antibody (sc1E3) against hIL-1beta that exhibited the equivalent affinity of the soluble IL-1 receptor type I (sIL-1R) for hIL-1beta and competitively blocked the sIL-1R from binding to the cytokine. The antibody proved to be more specific for hIL-1beta than the sIL-1R in that it failed to bind to either murine IL-1beta or human/murine IL-1alpha proteins. Additionally, failure of sc1E3 to bind to several hIL-1beta mutant proteins, altered at receptor site B, indicated that the antibody interacted preferentially with this site. This, coupled with other surface plasmon resonance and isothermal titration calorimetry measurements, shows that sc1E3 can achieve comparable affinity of binding hIL-1beta as the receptor through interactions at a smaller interface. This stable single chain antibody based heterodimer has simplified the complexity of the IL-1/IL-1R PPI system and will facilitate the design of the low molecular weight inhibitors of this interaction. PMID- 10852707 TI - Effects of Rett syndrome mutations of the methyl-CpG binding domain of the transcriptional repressor MeCP2 on selectivity for association with methylated DNA. AB - We have investigated the properties of mutant forms of the methyl-CpG binding transcriptional repressor MeCP2 associated with Rett syndrome, a childhood neurodevelopmental disorder. We find that four Rett syndrome mutations at known sites within the methyl-CpG binding domain (MBD) impair binding to methylated DNA, but have little effect on nonspecific interactions with unmethylated DNA. Three of these mutations (R106W, R133C, and F155S) have their binding affinities for methylated DNA reduced more than 100-fold; this is consistent with the hypothesis that impaired selectivity for methylated DNA of mutant MeCP2 contributes to Rett syndrome. However, a fourth mutant, T158M, has its binding affinity for methylated DNA reduced only 2-fold, indicative either of additional distinct regulatory functions associated with the MBD or of an exquisite sensitivity of developing neurons to the selective association of MeCP2 with methylated DNA. PMID- 10852708 TI - Human topoisomerase I poisoning by protoberberines: potential roles for both drug DNA and drug-enzyme interactions. AB - Protoberberines represent a structural class of organic cations that induce topoisomerase I-mediated DNA cleavage, a behavior termed topoisomerase I poisoning. We have employed a broad range of biophysical, biochemical, and computer modeling techniques to characterize and cross-correlate the DNA-binding and topoisomerase poisoning properties of four protoberberine analogues that differ with respect to the substituents on their A- and/or D-rings. Our data reveal the following significant features: (i) The binding of the four protoberberines unwinds duplex DNA by approximately 11 degrees, an observation consistent with an intercalative mode of interaction. (ii) Enthalpically favorable interactions, such as stacking interactions between the intercalated ligand and the neighboring base pairs, provide <50% of the thermodynamic driving force for the complexation of the protoberberines to duplex DNA. Computer modeling studies on protoberberine-DNA complexes suggest that only rings C and D intercalate into the host DNA helix, while rings A and B protrude out of the helix interior into the minor groove. (iii) All four protoberberine analogues are topoisomerase I-specific poisons, exhibiting little or no topoisomerase II poisoning activity. (iv) Modifications of the D-ring influence both DNA binding and topoisomerase I poisoning properties. Specifically, transference of a methoxy substituent from the 11- to the 9-position diminishes both DNA binding affinity and topoisomerase I poisoning activity, an observation suggesting that DNA binding is important in the poisoning of topoisomerase I by protoberberines. (v) Modifications of the A-ring have a negligible impact on DNA binding affinity, while exerting a profound influence on topoisomerase I poisoning activity. Specifically, protoberberine analogues containing either 2,3-dimethoxy; 3,4 dimethoxy; or 3, 4-methylenedioxy substituents all bind DNA with a similar affinity. By contrast, these analogues exhibit markedly different topoisomerase I poisoning activities, with these activities following the hierarchy: 3,4 methylenedioxy > 2,3-dimethoxy >> 3, 4-dimethoxy. These differences in topoisomerase I poisoning activity may reflect the differing abilities of the analogues to interact with specific functionalities on the enzyme, thereby stabilizing the enzyme in its cleavable state. In the aggregate, our results are consistent with a mechanistic model in which both ligand-DNA and ligand-enzyme interactions are important for the poisoning of topoisomerase I by protoberberines, with the DNA-directed interactions involving ring D and the enzyme-directed interactions involving ring A. It is reasonable to suggest that the poisoning of topoisomerase I by a broad range of other naturally occurring and synthetic ligands may entail a similar mechanism. PMID- 10852709 TI - Monitoring mobility in the early steps of unfolding: the case of oxidized cytochrome b(5) in the presence of 2 M guanidinium chloride. AB - A Model-Free analysis of the (15)N relaxation properties of oxidized cytochrome b(5), a heme-containing electron-transfer protein, has been performed in 2 M guanidinium chloride (GdmCl), i.e., just before the heme is released by the action of denaturant. This analysis provides information on the mobility in the nano- to picosecond time range. A parallel study on the motions in the milli- to microsecond time scale has also been performed by analyzing rotating-frame (15)N relaxation rates. The protein contains a 60:40 ratio of two conformers (A and B) differing for the rotation of the heme group around the alpha-gamma meso axis. The effect of denaturant has been followed for both species, and the mobility properties have been compared with the analogous information in the absence of denaturant. To complete the picture, we also performed (15)N relaxation measurements and the Model-Free analysis of the native B form, whereas data on the A form [Dangi, B., Sarma, S., Yan, C., Banville, D. L., Guiles, R. D. (1998) J. Phys. Chem. B 102, 8201-8208], as well as rotating-frame measurements for both native forms [Banci, L., Bertini, I., Cavazza, C., Felli, I. C., Koulougliotis, D. (1998) Biochemistry 37, 12320-12330; Arnesano, F., Banci, L., Bertini, I., Felli, I. C., Koulougliotis, D. (1999) Eur. J. Biochem. 260, 347-354], are already available in the literature. It is found that GdmCl tends to increase the internal mobility, although some residues are rigidified on both time scales. In the milli- to microsecond time scale, the tendency to increased mobility is reflected in a decrease in the tau(ex) values rather than in the number of residues experiencing conformational equilibria. In the nano- to picosecond time scale, the tendency to increased mobility is indicated by an overall decrease in the S(2) values. Color pictures are reported to visually show these effects. On the fast time scale, the B form is more mobile than the A form, reflecting the different stability with respect to unfolding. The increase in mobility upon addition of denaturant largely occurs around the heme pocket, which facilitates the release of the heme. The relevance of the internal motions with respect to the early steps of the unfolding process is also analyzed and discussed. PMID- 10852710 TI - Multiple replacements of glutamine 143 in human manganese superoxide dismutase: effects on structure, stability, and catalysis. AB - Glutamine 143 in human manganese superoxide dismutase (MnSOD) forms a hydrogen bond with the manganese-bound solvent molecule and is investigated by replacement using site-specific mutagenesis. Crystal structures showed that the replacement of Gln 143 with Ala made no significant change in the overall structure of the mutant enzyme. Two new water molecules in Q143A MnSOD were situated in positions nearly identical with the Oepsilon1 and Nepsilon2 of the replaced Gln 143 side chain and maintained a hydrogen-bonded network connecting the manganese-bound solvent molecule to other residues in the active site. However, their presence could not sustain the stability and activity of the enzyme; the main unfolding transition of Q143A was decreased 16 degrees C and its catalysis decreased 250 fold to k(cat)/K(m) = 3 x 10(6) M(-)(1) s(-)(1), as determined by stopped-flow spectrophotometry and pulse radiolysis. The mutant Q143A MnSOD and other mutants at position 143 showed very low levels of product inhibition and favored Mn(II)SOD in the resting state, whereas the wild type showed strong product inhibition and favored Mn(III)SOD. However, these differences did not affect the rate constant for dissociation of the product-inhibited complex in Q143A MnSOD which was determined from a characteristic absorbance at 420 nm and was comparable in magnitude ( approximately 100 s(-)(1)) to that of the wild-type enzyme. Hence, Gln 143, which is necessary for maximal activity in superoxide dismutation, appears to have no role in stabilization and dissociation of the product-inhibited complex. PMID- 10852711 TI - Interaction with heparin and matrix metalloproteinase 2 cleavage expose a cryptic anti-adhesive site of fibronectin. AB - We recently found that fibronectin (FN) had a functional site [YTIYVIAL sequence in the heparin-binding domain 2 (Hep 2)] that was capable of suppressing the integrin-mediated cell adhesion to extracellular matrix. However, our results also indicated that this anti-adhesive site seemed to be usually buried within the Hep 2 domain structure because of its hydrophobic nature, raising a question as to the physiological significance of the cryptic anti-adhesive activity of FN. The present study demonstrates that the cryptic anti-adhesive activity can be exposed through the physiological processes. A 30-kDa chymotryptic FN fragment derived from Hep 2 domain (Hep 2 fragment), which had no effect on adhesion of MSV-transformed nonproducer 3T3 cell line (KN(7)8) to FN, expressed the anti adhesive activity after treatment with 6 M urea. Light scattering and circular dichroism measurements showed that the urea treatment induced the conformational change of the Hep 2 fragment from a more compact form to an unfolded one. Incubation of the Hep 2 fragment with heparin also induced similar conformational changes and expression of anti-adhesive activity. Additionally, both the urea and heparin treatments made the Hep 2 fragment and intact FN much more accessible to the polyclonal antibody (alphaIII14A), with a recognition site near the anti adhesive site of FN. Specific cleavage of either the Hep 2 fragment or intact FN by matrix metalloproteinase 2 (MMP-2) released a 10-kDa fragment with the anti adhesive activity, which was shown to have the exposed anti-adhesive site on the amino-terminal region. Thus, the cryptic anti-adhesive activity of FN can be expressed upon conformational change and proteolytic cleavage of Hep 2 domain. PMID- 10852712 TI - Near-ultraviolet magnetic circular dichroism spectroscopy of protein conformational states: correlation of tryptophan band position and intensity with hemoglobin allostery. AB - The near-UV magnetic circular dichroism spectroscopy of the aromatic amino acid bands of hemoglobin was investigated as a potential probe of structural changes at the alpha(1)beta(2) interface during the allosteric transition. Allosteric effectors were used to direct carp and chemically modified human hemoglobins into the R (relaxed) or T (tense) state in order to determine the heme-ligation independent spectral characteristics of the quaternary states. The tryptophan magnetic circular dichroism (MCD) peak observed at 293 nm in the R state of N ethylsuccinimide- (NES-) des-Arg-modified human hemoglobin (Hb) was shifted to a slightly longer wavelength in the T state, consistent with the shift expected for tryptophan acting as a proton donor in a T-state hydrogen bond. Moreover, the increase observed in the T-state MCD intensity of this band relative to the R state intensity was consistent with the effect expected for proton donation by tryptophan on the basis of the Michl perimeter model of aromatic MCD. The peak-to trough magnitude of the R - T MCD difference spectrum is equal to 30% of the total R-state peak intensity contributed by all six tryptophans present in the human tetramer; the relative magnitude specific to the two beta37 tryptophans undergoing conformational change is estimated accordingly to be 3 times larger. The Trp-beta37 spectral shift, about 200 cm(-)(1), is in good agreement with the shifts observed in other H-bonded proton donors and provides corroborating spectral evidence for the formation in solution of a T-state Trp beta37-Asp alpha94 hydrogen bond observed in X-ray diffraction studies of deoxyHb crystals. PMID- 10852713 TI - Thermodynamic characterization of mutants of human fibroblast growth factor 1 with an increased physiological half-life. AB - Human acidic fibroblast growth factor (FGF-1) is a potent mitogen and angiogenic factor, with reportedly poor thermal stability and a relatively short in vivo half-life. However, certain mutants of FGF-1 have been described that exhibit a significant increase in half-life in tissue culture-based assays. FGF-1 contains three cysteine residues, two of which are highly conserved and buried within the protein core. Mutant forms of FGF-1 that substitute a serine residue at these cysteine positions have been reported to increase the protein's half-life and specific activity as well as decrease the dependence upon heparin for full activity. However, the underlying physical basis for this increase in half-life has not been determined. Possible effects include stabilization of protein structure and elimination of sulfhydryl chemistry at these positions. Here we have used differential scanning calorimetry and isothermal equilibrium denaturation to characterize thermodynamic parameters of unfolding for individual, and combination, cysteine to serine mutations in human FGF-1. The results show that substitution by serine is destabilizing at each cysteine position in wild-type FGF-1. Thus, the increased half-life previously reported for these mutations does not correlate with thermal stability and is most likely due to elimination of sulfhydryl chemistry. The results also suggest a method by which protein half-life may be modulated by rational design. PMID- 10852714 TI - Design of salt-insensitive glycine-rich antimicrobial peptides with cyclic tricystine structures. AB - Cyclic peptide backbone and cystine constraints were used to develop a broadly active salt-insensitive antimicrobial peptide [Gly(6)]ccTP 1a with eight Gly residues in an 18-residue sequence. The importance of rigidity and amphipathicity imparted by the cyclic and cystine constraints was examined in two peptide series based on tachyplesin, a known beta-stranded antimicrobial peptide. The first series, which retained the charge and hydrophobic amino acids of tachyplesin, but contained zero to four covalent constraints, included a cyclic tricystine tachyplesin (ccTP 1). Corresponding [Gly(6)] analogues were prepared in a parallel series with all six bulky hydrophobic amino acids in their sequences replaced with Gly. Circular dichroism measurements showed that ccTP 1 and [Gly(6)]ccTP 1a exhibited well-ordered beta-sheet structures, while the less constrained [Gly(6)] analogues were disordered. Except for linear peptides assayed under high-salt conditions, peptides with increased or decreased conformational constraints retained broad activity spectra with small variations in potency of 2-10-fold compared to that of tachyplesin. In contrast, Gly replacement analogues resulted in large variations in activity spectra and significant decreases in potency that roughly correlated with the decreases in conformational constraints. Except against Escherichia coli, the Gly-rich analogues with two or fewer covalent constraints were largely inactive under high salt conditions. Remarkably, the most constrained [Gly(6)]ccTP 1a retained a broad activity spectrum against all 10 test microbes in both low- and high-salt assays. Collectively, our results show that [Gly(6)]ccTP 1acould serve as a template for further analogue study to improve potency and specificity through single or multiple replacements of hydrophobic or unnatural amino acids. PMID- 10852715 TI - A single substitution of the insulin receptor kinase inhibits serine autophosphorylation in vitro: evidence for an interaction between the C-terminus and the activation loop. AB - We examined the effects of mutations of tyrosine and serine autophosphorylation sites on the dual specificity of the insulin receptor kinase (IRKD) in vitro using autophosphorylation and substrate phosphorylation and phosphopeptide mapping. For comparable studies, the recombinant kinases were overexpressed in the baculovirus system, purified, and analyzed. The phosphate incorporation into the enzymes was in the range of 3-4.5 mol/mol, and initial velocities of autophosphorylation were reduced up to 2-fold. However, the mutation Y1151F in the activation loop inhibited phosphate incorporation in the C-terminal serine residues 1275 and 1309, due to a 10-fold decrease of the initial velocity of serine autophosphorylation. Although the K(M) and V(MAX) values of this mutant were only slightly altered in substrate phosphorylation reactions using a recombinant C-terminal insulin receptor peptide (K(M): Y1151F, 9.9 +/- 0.4 microM; IRKD, 6.1 +/- 0.2 microM; V(MAX): Y1151F, 72 +/- 4 nmol min(-)(1) mg( )(1); IRKD, 117 +/- 6 nmol min(-)(1) mg(-)(1)), diminished phosphate incorporation into serine residues of the peptide was observed. In contrast, the phosphorylation of a recombinant IRS-1 fragment, which was shown to be phosphorylated markedly on serine residues by IRKD, was not affected by any kinase mutation. These results underline that IRKD is a kinase with dual specificity. The substrate specificity toward C-terminal serine phosphorylation sites can be modified by a single amino acid substitution in the activation loop, whereas the specificity toward IRS-1 is not affected, suggesting that the C terminus and the activation loop interact. PMID- 10852716 TI - Cysteine 981 of the human insulin receptor is required for covalent cross-linking between beta-subunit and a thiol-reactive membrane-associated protein. AB - The cytoplasmic domain of the insulin receptor (IR) beta-subunit contains cysteine (Cys) residues whose reactivity and function remain uncertain. In this study, we examined the ability of the bifunctional cross-linking reagent 1,6 bismaleimidohexane (BMH) to covalently link IR with interacting proteins that possess reactive thiols. Transfected Chinese hamster ovary cells expressing either the wild-type human IR, C-terminally truncated receptors, or mutant receptors with Cys --> Ala substitutions and mouse 3T3-L1 adipocytes were used to compare the BMH effect. The results showed the formation of a large complex between the wild-type human receptor beta-subunit and molecule X, a thiol reactive membrane-associated protein, in both intact and semipermeabilized cells in response to BMH. Prior cell stimulation with insulin had only a modest effect in this process. Western blot analysis revealed that the receptor alpha-subunit was not present in the beta-X complex. The BMH cross-linking did not inhibit in vitro tyrosine phosphorylation of the receptor complexed with molecule X. Both the human IR Cys981Ala mutant and murine IR, that lacks the equivalent of human Cys(981), failed to react with BMH. Finally, no covalent association between IR beta-subunit and IRS-1, the protein tyrosine phosphatase LAR or SHP-2 was observed in BMH-treated cells expressing the wild-type human IR. These results demonstrate a striking difference in reactivity among the cytoplasmic IR beta subunit thiols and clearly show that Cys(981) of human IR beta-subunit is in close proximity to a thiol-reactive membrane-associated protein under basal and insulin-stimulated conditions. PMID- 10852717 TI - The mechanism of GTP hydrolysis by dynamin II: a transient kinetic study. AB - Dynamin II is a 98 kDa protein (870 amino acids) required for the late stages of clathrin-mediated endocytosis. The GTPase activity of dynamin is required for its function in the budding stages of receptor-mediated endocytosis and synaptic vesicle recycling. This activity is stimulated when dynamin self-associates on multivalent binding surfaces, such as microtubules and anionic liposomes. We first investigated the oligomeric state of dynamin II by analytical ultracentrifuge sedimentation equilibrium measurements at high ionic strength and found that it was best described by a monomer-tetramer equilibrium. We then studied the intrinsic dynamin GTPase mechanism by using a combination of fluorescence stopped-flow and HPLC methods using the fluorescent analogue of GTP, mantdGTP (2'-deoxy-3'-O-(N-methylanthraniloyl) guanosine-5'-triphosphate), under the same ionic strength conditions. The results are interpreted as showing that mantdGTP binds to dynamin in a two-step mechanism. The dissociation constant of mantdGTP binding to dynamin, calculated from the ratio of the off-rate to the on rate (k(off)/k(on)), was 0.5 microM. Cleavage of mantdGTP then occurs to mantdGDP and P(i) followed by the rapid release of mantdGDP and P(i). No evidence of reversibility of hydrolysis was observed. The cleavage step itself is the rate limiting step in the mechanism. This mechanism more closely resembles that of the Ras family of proteins involved in cell signaling than the myosin ATPase involved in cellular motility. PMID- 10852718 TI - Fatty acid binding proteins from different tissues show distinct patterns of fatty acid interactions. AB - Fatty acid binding proteins (FABP) form a family of proteins displaying tissue specific expression. These proteins are involved in fatty acid (FA) transport and metabolism by mechanisms that also appear to be tissue-specific. Cellular retinoid binding proteins are related proteins with unknown roles in FA transport and metabolism. To better understand the origin of these tissue-specific differences we report new measurements, using the acrylodated intestinal fatty acid binding protein (ADIFAB) method, of the binding of fatty acids (FA) to human fatty acid binding proteins (FABP) from brain, heart, intestine, liver, and myelin. We also measured binding of FA to a retinoic acid (CRABP-I) and a retinol (CRBP-II) binding protein and we have extended to 19 different FA our characterization of the FA-ADIFAB and FA-rat intestinal FABP interactions. These studies extend our previous analyses of human FABP from adipocyte and rat FABPs from heart, intestine, and liver. Binding affinities varied according to the order brain approximately myelin approximately heart > liver > intestine > CRABP > CRBP. In contrast to previous studies, no protein revealed a high degree of selectivity for particular FA. The results indicate that FA solubility (hydrophobicity) plays a major role in governing binding affinities; affinities tend to increase with increasing hydrophobicity (decreasing solubility) of the FA. However, our results also reveal that, with the exception of the intestinal protein, FABPs exhibit an additional attractive interaction for unsaturated FA that partially compensates for their trend toward lower affinities due to their higher aqueous solubilities. Thermodynamic potentials were determined for oleate and arachidonate binding to a subset of the FABP and retinoid binding proteins. FA binding to all FABPs was enthalpically driven. The DeltaH degrees values for paralogous FABPs, proteins from the same species but different tissues, reveal an exceptionally wide range of values, from -22 kcal/mol (myelin) to -7 kcal/mol (adipocyte). For orthologous FABPs from the same tissue but different species, DeltaH degrees values were similar. In contrast to the enthalpic dominance of FA binding to FABP, binding of FA to CRABP-I was entropically driven. This is consistent with the notion that FA specificity for FABP is determined by the enthalpy of binding. Proteins from different tissues also revealed considerable heterogeneity in heat capacity changes upon FA binding, DeltaC(p) values ranged between 0 and -1.3 kcal mol(-1) K(-1). The results demonstrate that thermodynamic parameters are quite different for paralogous but are quite similar for orthologous FABP, suggesting tissue-specific differences in FABP function that may be conserved across species. PMID- 10852719 TI - Characterization of a new dihemic c(4)-type cytochrome isolated from Thiobacillus ferrooxidans. AB - A new soluble c-type cytochrome has been purified to homogeneity from the acidophilic proteobacterium Thiobacillus ferrooxidans BRGM. It is characterized by an alpha-peak wavelength of 552 nm, a molecular mass of 26 567 Da (as determined by mass spectroscopy) and a pI value of 8. Optical redox titrations at pH 4.0 revealed the presence of two distinguishable redox species with an E(m) of 510 mV and an E(m) of 430 +/- 20 mV. EPR spectra recorded for this heme protein demonstrated the presence of stoichiometric amounts of two low-spin hemes with a g(z)() of 3.08 (510 mV species) and a g(z)() of 3.22 (430 mV species). Modifications of the physicochemical properties of the cytochrome were observed on complex formation with the blue copper protein rusticyanin, another soluble electron carrier in the genus Thiobacillus. N-Terminal sequencing yielded the polypeptide sequence up to the 50th residue. The determined sequence was found to be present (at 100% amino acid identity) in the (unfinished) genome of T. ferrooxidans ATCC 23270, and the corresponding full-length protein turned out to be surprisingly similar (34.5% amino acid identity) to another c(4)-type diheme protein from T. ferrooxidans BRGM [Cavazza, C., et al. (1996) Eur. J. Biochem. 242, 308-314], the gene of which is also present (at 97% amino acid identity) in the T. ferrooxidans ATCC 23270 genome. The physicochemical properties and sequence characteristics of both c(4) cytochromes present in the same bacteria are compared, and the functional role of this new diheme protein in the iron(II) oxidizing electron transport chain in the genus Thiobacillus is discussed. PMID- 10852720 TI - Electron transfer in reaction center core complexes from the green sulfur bacteria Prosthecochloris aestuarii and Chlorobium tepidum. AB - Electron transfer in reaction center core (RCC) complexes from the green sulfur bacteria Prosthecochloris aestuarii and Chlorobium tepidum was studied by measuring flash-induced absorbance changes. The first preparation contained approximately three iron-sulfur centers, indicating that the three putative electron acceptors F(X), F(A), and F(B) were present; the Chl. tepidum complex contained on the average only one. In the RCC complex of Ptc. aestuarii at 277 K essentially all of the oxidized primary donor (P840(+)) created by a flash was rereduced in several seconds by N-methylphenazonium methosulfate. In RCC complexes of Chl. tepidum two decay components, one of 0.7 ms and a smaller one of about 2 s, with identical absorbance difference spectra were observed. The fast component might be due to a back reaction of P840(+) with a reduced electron acceptor, in agreement with the notion that the terminal electron acceptors, F(A) and F(B), were lost in most of the Chl. tepidum complexes. In both complexes the terminal electron acceptor (F(A) or F(B)) could be reduced by dithionite, yielding a back reaction of 170 ms with P840(+). At 10 K in the RCC complexes of both species P840(+) was rereduced in 40 ms, presumably by a back reaction with F(X)(-). In addition, a 350 micros component occurred that can be ascribed to decay of the triplet of P840, formed in part of the complexes. For P840(+) rereduction a pronounced temperature dependence was observed, indicating that electron transfer is blocked after F(X) at temperatures below 200 K. PMID- 10852721 TI - Construction and characterization of a heterodimeric iron protein: defining roles for adenosine triphosphate in nitrogenase catalysis. AB - One molecule of MgATP binds to each subunit of the homodimeric Fe protein component of nitrogenase. Both MgATP molecules are hydrolyzed to MgADP and P(i) in reactions coupled to the transfer of one electron into the MoFe protein component. As an approach to assess the contributions of individual ATP binding sites, a heterodimeric Fe protein was produced that has an Asn substituted for residue 39 in the ATP binding domain in one subunit, while the normal Asp(39) residue within the other subunit remains unchanged. Separation of the heterodimeric Fe protein from a mixed population with homodimeric Fe proteins contained in crude extracts was accomplished by construction of a seven His tag on one subunit and a differential immobilized-metal-affinity chromatography technique. Three forms of the Fe protein (wild-type homodimeric Fe protein [Asp(39)/Asp(39)], altered homodimeric Fe protein [Asn(39)/Asn(39)], and heterodimeric Fe protein [Asp(39)/Asn(39)]) were compared on the basis of the biochemical and biophysical changes elicited by nucleotide binding. Among those features examined were the MgATP- and MgADP-induced protein conformational changes that are manifested by the susceptibility of the [4Fe-4S] cluster to chelation and by alterations in the electron paramagnetic resonance, circular dichroism, and midpoint potential of the [4Fe-4S] cluster. The results indicate that changes in the [4Fe-4S] cluster caused by nucleotide binding are the result of additive conformational changes contributed by the individual subunits. The [Asp(39)/Asn(39)] Fe protein did not support substrate reduction activity but did hydrolyze MgATP and showed MgATP-dependent primary electron transfer to the MoFe protein. These results support a model where each MgATP site contributes to the rate acceleration of primary electron transfer, but both MgATP sites must be functioning properly for substrate reduction. Like the altered homodimeric [Asn(39)/Asn(39)] Fe protein, the heterodimeric [Asp(39)/Asn(39)] Fe protein was found to form a high affinity complex with the MoFe protein, revealing that alteration on one subunit is sufficient to create a tight complex. PMID- 10852722 TI - Resolution of the membrane domain of bovine complex I into subcomplexes: implications for the structural organization of the enzyme. AB - Complex I (NADH:ubiquinone oxidoreductase) purified from bovine heart mitochondria was treated with the detergent N, N-dimethyldodecylamine N-oxide (LDAO). The enzyme dissociated into two known subcomplexes, Ialpha and Ibeta, containing mostly hydrophilic and hydrophobic subunits, and a previously undetected fragment referred to as Igamma. Subcomplex Igamma contains the hydrophobic subunits ND1, ND2, ND3, and ND4L which are encoded in the mitochondrial genome, and the nuclear-encoded subunit KFYI. During size-exclusion chromatography in the presence of LDAO, subcomplex Ialpha lost several subunits and formed another characterized subcomplex known as Ilambda. Similarly, subcomplex Ibeta dissociated into two smaller subcomplexes, one of which contains the hydrophobic subunits ND4 and ND5; subcomplex Igamma released a fragment containing ND1 and ND2. These results suggest that in the intact complex subunits ND1 and ND2 are likely to be in a different region of the membrane domain than subunits ND4 and ND5. The compositions of the various subcomplexes and fragments of complex I provide an organization of the subunits of the enzyme in the framework of the known low resolution structure of the enzyme. PMID- 10852723 TI - Yeast ribosomal protein L24 affects the kinetics of protein synthesis and ribosomal protein L39 improves translational accuracy, while mutants lacking both remain viable. AB - Four mutant strains from Saccharomyces cerevisiae were used to study ribosome structure and function. They included a strain carrying deletions of the two genes encoding ribosomal protein L24, a strain carrying a mutation spb2 in the gene for ribosomal protein L39, a strain carrying a deletion of the gene for L39, and a mutant lacking both L24 and L39. The mutant lacking only L24 showed just 25% of the normal polyphenylalanine-synthesizing activity followed by a decrease in P-site binding, suggesting the possibility that protein L24 is involved in the kinetics of translation. Each of the two L39 mutants displayed a 4-fold increase of their error frequencies over the wild type. This was accompanied by a substantial increase in A-site binding, typical of error-prone mutants. The absence of L39 also increased sensitivity to paromomycin, decreased the ribosomal subunit ratio, and caused a cold-sensitive phenotype. Mutant cells lacking both ribosomal proteins remained viable. Their ribosomes showed reduced initial rates caused by the absence of L24 but a normal extent of polyphenylalanine synthesis and a substantial in vivo reduction in the amount of 80S ribosomes compared to wild type. Moreover, this mutant displayed decreased translational accuracy, hypersensitivity to the antibiotic paromomycin, and a cold-sensitive phenotype, all caused mainly by the deletion of L39. Protein L39 is the first protein of the 60S ribosomal subunit implicated in translational accuracy. PMID- 10852724 TI - Evidence that DNA polymerase delta isolated by immunoaffinity chromatography exhibits high-molecular weight characteristics and is associated with the KIAA0039 protein and RPA. AB - DNA polymerase delta, the key enzyme for eukaryotic chromosomal replication, has been well characterized as consisting of a core enzyme of a 125 kDa catalytic subunit and a smaller 50 kDa subunit. However, less is known about the other proteins that may comprise additional subunits or participate in the macromolecular protein complex that is involved in chromosomal DNA replication. In this study, the properties of calf thymus pol delta preparations isolated by immunoaffinity chromatography were investigated. It is demonstrated for the first time using highly purified preparations that the pol delta heterodimer is associated with other polypeptides in high-molecular weight species that range from 260000 to >500000 in size, as determined by FPLC gel filtration. These preparations are associated with polypeptides of ca. 68-70, 34, 32, and 25 kDa. Similar findings were revealed with glycerol gradient ultracentrifugation. The p68 polypeptide was shown to be a PCNA binding protein by overlay methods with biotinylated PCNA. Protein sequencing of the p68, p34, and p25 polypeptide bands revealed sequences that correspond to the hypothetical protein KIAA0039. KIAA0039 displays a small but significant degree of homology to Schizosaccharomyces pombe Cdc27, which, like Saccharomyces cerevisiae Pol32p, has been described as the third subunit of yeast pol delta. These studies provide evidence that p68 is a subunit of pol delta. In addition, the p68-70 and p32 polypeptides were found to be derived from the 70 and 32 kDa subunits of RPA, respectively. PMID- 10852725 TI - RNA aptamers to S-adenosylhomocysteine: kinetic properties, divalent cation dependency, and comparison with anti-S-adenosylhomocysteine antibody. AB - To explore the potential of RNA aptamers as small-molecule discriminating devices, we have characterized the properties of aptamers selected from a library of approximately 10(14) variants through their interaction with S adenosylhomocysteine (SAH, AdoHcy). Competition studies with SAH and azaSAM analogues revealed that the Hoogsteen face of adenine is the main contributor to binding, whereas specificity for SAH is conferred by a secondary contact point at or near the sulfur/thioether of homocysteine (Hcy). Binding specificities were determined by both affinity chromatography and a novel method designed for the biosensor. The kinetic properties of individual aptamers, including the "classic" ATP aptamer that also emerged in our selection, were studied by biosensor analysis. Association rates were slow, but the complexes were stable, suggesting micro- to submicromolar affinities. A solution affinity of approximately 0.1 microM was found for the strongest binding variant under the conditions used for selection (5 mM Mg(2+)). Systematic studies of the effect of Mg(2+) and Mn(2+) on binding, however, revealed that the affinity of the aptamers could be substantially improved, and at optimized conditions of Mn(2+) the affinity of one of the aptamers approached that of an anti-SAH antibody with similar/identical binding specificity. Comparisons with the MAb suggest that the on rate is the limiting factor for high-affinity binding by these aptamers, and comparison with a truncated aptamer shows that shortening of RNA constructs may alter binding kinetics as well as sensitivity to ions. PMID- 10852726 TI - 3-Hydroxykynurenine and 3-hydroxyanthranilic acid generate hydrogen peroxide and promote alpha-crystallin cross-linking by metal ion reduction. AB - The kynurenine pathway catabolite 3-hydroxykynurenine (3HK) and redox-active metals such as copper and iron are implicated in cataractogenesis. Here we investigate the reaction of kynurenine pathway catabolites with copper and iron, as well as interactions with the major lenticular structural proteins, the alpha crystallins. The o-aminophenol kynurenine catabolites 3HK and 3 hydroxyanthranilic acid (3HAA) reduced Cu(II)>Fe(III) to Cu(I) and Fe(II), respectively, whereas quinolinic acid and the nonphenolic kynurenine catabolites kynurenine and anthranilic acid did not reduce either metal. Both 3HK and 3HAA generated superoxide and hydrogen peroxide in a copper-dependent manner. In addition, 3HK and 3HAA fostered copper-dependent alpha-crystallin cross-linking. 3HK- or 3HAA-modifed alpha-crystallin showed enhanced redox activity in comparison to unmodified alpha-crystallin or ascorbate-modified alpha-crystallin. These data support the possibility that 3HK and 3HAA may be cofactors in the oxidative damage of proteins, such as alpha-crystallin, through interactions with redox-active metals and especially copper. These findings may have relevance for understanding cataractogenesis and other degenerative conditions in which the kynurenine pathway is activated. PMID- 10852727 TI - Mutational evidence of transition state stabilization by serine 88 in Escherichia coli type I signal peptidase. AB - Type I signal peptidase (SPase I) catalyzes the hydrolytic cleavage of the N terminal signal peptide from translocated preproteins. SPase I belongs to a novel class of Ser proteases that utilize a Ser/Lys dyad catalytic mechanism instead of the classical Ser/His/Asp triad found in most Ser proteases. Recent X-ray crystallographic studies indicate that the backbone amide nitrogen of the catalytic Ser 90 and the hydroxyl side chain of Ser 88 might participate as H bond donors in the transition-state oxyanion hole. In this work, contribution of the side-chain Ser 88 in Escherichia coli SPase I to the stabilization of the transition state was investigated through in vivo and in vitro characterizations of Ala-, Cys-, and Thr-substituted mutants. The S88T mutant maintains near-wild type activity with the substrate pro-OmpA nuclease A. In contrast, substitution with Cys at position 88 results in more than a 740-fold reduction in activity (k(cat)) whereas S88A retains much less activity (>2440-fold decrease). Measurements of the kinetic constants of the individual mutant enzymes indicate that these decreases in activity are attributed mainly to decreases in k(cat) while effects on K(m) are minimal. Thermal inactivation and CD spectroscopic analyses indicate no global conformational perturbations of the Ser 88 mutants relative to the wild-type E. coli SPase I enzyme. These results provide strong evidence for the stabilization by Ser 88 of the oxyanion intermediate during catalysis by E. coli SPase I. PMID- 10852728 TI - Calmodulin-peptide interactions: apocalmodulin binding to the myosin light chain kinase target-site. AB - Noncovalent binding of the synthetic peptide RS20 to calmodulin in the presence of calcium was confirmed by electrospray ionization coupled with Fourier transform ion cyclotron resonance mass spectrometry to form a complex with a 1:1:4 calmodulin/RS20/calcium stoichiometry. There was no evidence for formation of a calmodulin-RS20-Ca(2) species. The absence of calmodulin-RS20-Ca(2) would be consistent with models in which the two globular domains are coupled functionally. There was evidence that calmodulin, RS20-calmodulin without associated calcium, and calmodulin-RS20-Ca(4) existed together in solution, whereas calmodulin-calcium complexes were absent. It is proposed that calcium binding to form the calmodulin-RS20-Ca(4) complex occurs after an initial RS20 calmodulin binding event, and serves to secure the target within the calmodulin structure. The binding of more than one RS20 molecule to calmodulin was observed to induce unfolding of calmodulin. PMID- 10852729 TI - Calcium-sensitive interaction between calmodulin and modified forms of rat brain neurogranin/RC3. AB - Neurogranin (NG) binding of calmodulin (CaM) at its IQ domain is sensitive to Ca(2+) concentration and to modifications by protein kinase C (PKC) and oxidants. The PKC phosphorylation site of NG is within the IQ domain whereas the four oxidant-sensitive Cys residues are outside this region. These Cys residues were oxidized forming two pairs of intramolecular disulfides, and could also be glutathiolated by S-nitrosoglutathione resulting in the incorporation of four glutathiones per NG. Circular dichroism (CD) showed that modification of NG by phosphorylation, oxidation forming intramolecular disulfides, or glutathiolation did not affect the alpha-helical content of this protein. Mutation of the four Cys residues [Cys(-)-NG] to Gly and Ser did not affect the alpha-helical content either. Interaction of CaM with the reduced (red)-, glutathiolated (GS)-, or Cys( )-NG in the Ca(2+)-free solution resulted in an increase in the alpha-helicity determined by their CD spectra, but relatively little change was seen with the oxidized NG (ox-NG) or phosphorylated NG (PO(4)-NG). The binding affinities between the various modified forms of NG and CaM were determined by CD spectrometry and sedimentation equilibrium: their affinities were Cys(-)-NG > red NG, GS-NG > ox-NG > PO(4)-NG. Unlike Cys(-)-, red-, and GS-NG, neither ox- nor PO(4)-NG bound to a CaM-affinity column. Thus, both oxidation of NG to form intramolecular disulfides and phosphorylation of NG by PKC are effective in modulating the intracellular level of CaM. These results indicate that modification of NG to form intramolecular disulfides outside the IQ domain provides an alternative mechanism for regulation of its binding affinity to CaM. PMID- 10852730 TI - Intersubunit association induces unique allosteric dependence of the T127L CRP mutant on pH. AB - The allosteric activation of the T127-->L mutant of 3',5'-cyclic adenosine monophosphate (cAMP) receptor protein (CRP) by cAMP changes from an exothermic, independent two-site binding mechanism at pH 7.0 to an endothermic, interacting two-site binding mechanism at pH 5.2, similar to that observed for CRP at pH 7.0 and 5.2. Since the T127-->L mutation at the subunit interface of the CRP dimer creates a more perfect leucine-zipper motif, it is believed to increase the intersubunit association and the stability of the CRP, as is observed by the higher thermal stability of the T127L mutant relative to that of CRP in differential scanning calorimetry (DSC) measurements. The DSC scans also exhibit a single thermal denaturation transition for CRP and a S128A mutant from pH 5.2 to 7. 0, while the broader transition peak of the T127L mutant becomes resolvable into two transitions below pH < or =5.2. Circular dichroism measurements on T127L and CRP at pH 7.0 and 5.2 show changes in the tertiary structure of both proteins with the exception of the tertiary structure around the two tryptophan residues in the amino-terminal domain. Although gel electrophoresis of the proteolysis (pH 5.2) products of T127L, CRP, and their cAMP- and cGMP-ligated complexes shows the subunit band and an amino-terminal domain fragment band, the fully allosterically activated complexes of T127L and CRP show the amino-terminal domain fragment band but not the subunit band. The results are interpreted in terms of the allosteric activation of CRP by cAMP by a conformational change from an "open" to a "closed" form of CRP, which involves changes in the tertiary structure of the carboxyl terminal domains that are partially induced by an increase in the intersubunit association in T127L. While T127L retains its intersubunit association from pH 5.2 to 7.0, changes occur in the carboxyl-terminal domain so that the endothermic, allosteric activation mechanism of CRP by cAMP is restored in T127L at pH 5.2. PMID- 10852731 TI - 15N isotope effects in glutamine hydrolysis catalyzed by carbamyl phosphate synthetase: evidence for a tetrahedral intermediate in the mechanism. AB - 15N isotope effects have been measured on the hydrolysis of glutamine catalyzed by carbamyl phosphate synthetase of Escherichia coli. The isotope effect in the amide nitrogen of glutamine is 1. 0217 at 37 degrees C with the wild-type enzyme in the presence of MgATP and HCO(3)(-) (overall reaction taking place). This V/K isotope effect indicates that breakdown of the tetrahedral intermediate formed with Cys 269 to release ammonia is the rate-limiting step in the hydrolysis. A full isotope effect of 1. 0215 is also seen in the partial reaction catalyzed by an E841K mutant enzyme, whose rate of glutamine hydrolysis is not affected by MgATP and HCO(3)(-). With wild-type enzyme in the absence of MgATP and HCO(3)(-), however, the (15)N isotope effect is reduced to 1. 0157. These isotope effects are interpreted in terms of partitioning of the tetrahedral intermediate whose rate of formation is dependent upon a conformation change which closes the active site after glutamine binding and prepares the enzyme for catalysis. An Ordered Uni Bi mechanism for glutamine hydrolysis that is consistent with the isotope effects and with the catalytic properties of the enzyme is proposed. PMID- 10852732 TI - Role of an interdomain Gly-Gly sequence at the regulatory-substrate domain interface in the regulation of Escherichia coli. D-3-phosphoglycerate dehydrogenase. AB - The regulatory and substrate binding domains of D-3-phosphoglycerate dehydrogenase (PGDH, EC 1.1.1.95) from Escherichia coli are connected by a single polypeptide strand that contains a Gly-Gly sequence approximately midway between the domains. The potential flexibility of this sequence and its strategic location between major domain structures suggests that it may function in the conformational change leading from effector binding to inhibition of the active site. Site-directed mutagenesis of this region (Gly-336-Gly-337) supports this hypothesis. When bulky side chains were substituted for the glycines at these positions, substantial changes in the ability of serine to inhibit the enzyme were seen with little effect on the activity of the enzyme. The effect of these substitutions could be alleviated by placing a new glycine residue at position 335, immediately flanking the original glycine pair. On the other hand, substituting a glycine at position 338 revealed a critical role for the side chain of Arg-338. This residue may function in stabilizing the conformation about the Gly-Gly turn, resulting in a specific orientation of the adjacent domains relative to each other. Rotation about the phi or psi bonds of either Gly-336 or Gly-337 would have a profound effect on this orientation. The data are consistent with this as a role for the Gly-Gly sequence between the regulatory and substrate binding domains of PGDH. PMID- 10852733 TI - Spatial orientation and dynamics of the U1A proteins in the U1A-UTR complex. AB - The human U1A protein contains three distinct domains: the N-terminal RBD1 (amino acids 1-101), the C-terminal RBD2 (amino acids 195-282), and the linker region (amino acids 102-194). The RBD1 domains of two U1A proteins bind specifically to two internal loops in the 3' untranslated region (3' UTR) of its own pre-mRNA. Tryptophan fluorescence and fluorescence resonance energy transfer data show that the two RBD2 domains do not interact with any regions of the UTR complex and display an overall tumbling that is uncorrelated from the core of the complex (formed by RBD1-UTR), indicating that the linker regions of the two U1A proteins remain flexible. The two RBD2 domains are separated by an apparent distance greater than 74 A in the UTR complex. The linker region adjacent to the RBD1 domain (103-ERDRKREKRKPKSQETP-119) is supposedly involved in protein-protein interactions (12). A single cysteine, introduced at position 101 or 121 of the U1A protein, was used as a specific attachment site for the fluorophore pair IAEDANS [N'-iodoacetyl-N'-(1-sulfo-5-n-naphthyl)ethylenediamine]/DABMI [4 (dimethylamino)-phenylazophenyl-4'-maleimide]. In the U1A-UTR complex (2:1), the dyes at the 101 position are separated by = approximately 51 A, while the dyes at the 121 position are at an apparent distance = approximately 58 A. The 101-121 crossed distance on adjacent U1A proteins averages to = 55 A. These results suggest that the amino acid sequence 101-121 of the two U1A proteins in the complex are held in proximity to each other in a compact conformation. PMID- 10852734 TI - Three-dimensional structure of escherichia coli asparagine synthetase B: A short journey from substrate to product PMID- 10852735 TI - Zygomycosis in the 1990s in a tertiary-care cancer center. AB - Twenty-four patients with cancer met predetermined criteria for a diagnosis of zygomycosis over a 10-year period at our institution. All had hematologic malignancy, and most had either neutropenia or steroid use as a risk factor. Pulmonary involvement mimicking invasive aspergillosis was the most common presentation, and dissemination was seen in 58% of patients on whom autopsies were performed. Three-fourths of the patients with pulmonary zygomycosis had pathogenic microorganisms other than zygomycetes isolated from respiratory specimens. The sensitivity of cultures in detecting zygomycetes from respiratory specimens was low. A culture positive for zygomycetes was typically a preterminal finding in the fatal, acute cases. Two-thirds of the patients died. Favorable outcome seemed to correlate with lack of pulmonary involvement, surgical debridement, neutrophil recovery, and a cumulative total amphotericin B dose of 2000 mg. Therapy with high-dose amphotericin B, combined with aggressive surgery and immune reconstitution, offers the best chance for survival of cancer patients with zygomycosis. PMID- 10852736 TI - Antimicrobial resistance and clinical outcome of Bacteroides bacteremia: findings of a multicenter prospective observational trial. AB - There is debate regarding the correlation between in vitro susceptibility testing and clinical response to therapy for Bacteroides bacteremia. We conducted a prospective multicenter observational study of 128 patients with bacteroides bacteremia. Outcome was correlated with results of in vitro susceptibility testing of Bacteroides isolates recovered from blood and/or nonblood sites, determined with use of 3 end points: mortality at 30 days, clinical response (cure vs. failure), and microbiological response (eradication vs. persistence). The mortality rate among patients who received inactive therapy (45%) was higher than among patients who received active therapy (16%; P=.04). Clinical failure (82%) and microbiological persistence (42%) were higher for patients who received inactive therapy than for patients who received active therapy (22% and 12%, respectively; P=.0002 and.06, respectively). In vitro activity of agents directed at Bacteroides species reliably predicts outcome: the specificity was 97%, and positive predictive value was 82%. Antimicrobial susceptibility testing may be indicated for patients whose blood specimens yield Bacteroides species. PMID- 10852737 TI - Immunization of pediatric solid organ transplant candidates and recipients. AB - Organ transplantation has evolved from an experimental procedure to an accepted treatment for otherwise irreversible or congenital disorders. The immunosuppression necessary to prevent rejection enhances the severity of many infectious diseases and may potentially attenuate the response to vaccines designed to prevent disease. In spite of the frequency and severity of infectious diseases in organ transplant recipients, many children are not fully vaccinated before transplantation. The safety and efficacy of many of the currently available vaccines for solid organ transplant recipients have not been evaluated. We review the currently available data on immunization safety and efficacy, discuss experimental vaccines, and outline strategies to avoid vaccine preventable diseases in pediatric organ transplant recipients. PMID- 10852738 TI - Type 1 cytokines and the pathogenesis of tuberculosis. PMID- 10852739 TI - The efficacy of nasal continuous positive airway pressure in the treatment of obstructive sleep apnea syndrome is proven. PMID- 10852740 TI - The efficacy of nasal continuous positive airway pressure in the treatment of obstructive sleep apnea syndrome is not proven. PMID- 10852741 TI - REBUTTAL FROM DRS. DAVIES AND STRADLING. PMID- 10852742 TI - REBUTTAL FROM DRS. WRIGHT AND SHELDON. PMID- 10852743 TI - How flow met volume in three-dimensional space. PMID- 10852744 TI - Effects of nitric oxide in septic shock. AB - Nitric oxide (NO) is believed to play a key role in the pathogenesis of septic shock, although many aspects of NO's involvement remain poorly defined. Recent years have seen advances in our understanding of the production and effects of NO, but much of the work has been done in animal models and may not be directly relevant to the clinical situation. Differences between species and models can account for many of the apparently conflicting results obtained. Nevertheless, NO directed strategies have been developed and tested clinically. However, NO can have both beneficial and detrimental effects on many organ systems in sepsis and attempts to nonselectively block all its actions may therefore not yield positive results on outcome. Further exploration and precision of the role of NO and development of techniques to assess the NO balance in individual patients is necessary before further progress can be made in this field. PMID- 10852745 TI - Increased levels of circulating interleukin-18 in patients with advanced tuberculosis. AB - Interleukin-18 (IL-18) has recently been identified as an interferon- gamma inducing factor and it plays an important role in the Th1 response. We measured serum levels of IL-18 and interferon-gamma (IFN-gamma) in 43 patients with pulmonary tuberculosis and 25 healthy control subjects. Significantly increased levels of circulating IL-18 and IFN-gamma were found in pulmonary tuberculosis as compared with those in healthy control subjects. Circulating IL-18 and IFN-gamma correlated with the extent of disease in pulmonary tuberculosis. We found significantly increased levels of circulating IL-18 and IFN-gamma in the patients with high-grade fever. Circulating IL-18 significantly correlated with circulating IFN-gamma. IL-18 may play an important role in immune response to human infection with Mycobacterium tuberculosis. PMID- 10852746 TI - Assessment of the Th1/Th2 paradigm in whole blood in atopy and asthma. Increased IFN-gamma-producing CD8(+) T cells in asthma. AB - Atopy is characterized by an immune system that is biased to T helper cell, type 2 (Th2) activation. This condition predisposes to asthma, a disease in which a Th2 activation was found in blood and lungs. However, most blood studies have considered purified cells, which might give an incomplete view of immune reactions. In this study, we assessed in whole blood cultures the Th1/Th2 paradigm in atopy and asthma. Sixty-nine subjects (31 atopic asthmatics, six nonatopic asthmatics, 13 atopic nonasthmatics, and 19 control subjects) were included in this study. Interleukin-4 (IL-4), interferon gamma (IFN-gamma), and IL-12 were assayed in stimulated whole blood culture supernatants by using a flow cytometer microsphere-based assay. Intracellular IL-4 and IFN-gamma were detected in T cells and CD8(+) T cells by flow cytometry. Atopy was characterized by a higher production of IL-4, which was correlated to total IgE levels, and by an impairment of the T-cell capacity to produce IFN-gamma. This impairment was correlated to the number of positive skin tests. In asthma, the overproduction of IL-4 was still found if atopy was present. Unexpectedly, an overproduction of IFN gamma was found, which was related to an increased capacity of CD8(+) T cells to produce IFN-gamma. The number of IFN-gamma-producing CD8(+) T cells was related to asthma severity, to bronchial hyperresponsiveness, and to blood eosinophilia. In addition, this number was correlated to IL-12 production. These results show that in addition to the well-known Th2 inflammation in asthma, there are IFN gamma-producing CD8(+) T cells in the blood, possibly controlled by IL-12. PMID- 10852747 TI - Biologic variability in mechanical ventilation rate and tidal volume does not improve oxygenation or lung mechanics in canine oleic acid lung injury. AB - Mechanical ventilation in patients with acute respiratory distress syndrome and acute lung injury (ALI) remains a difficult challenge because of the conflict between maintaining adequate gas exchange and furthering lung injury via overdistention. In a recent study, Lefevre and colleagues (Am. J. Respir. Crit. Care Med. 1996;154: 1567-1572) suggested that mechanical ventilation with natural biologic variability (BV) in breath-to-breath respiratory frequency (f) and VT could reduce lung injury and improve gas exchange without increases in mean airway pressure (Paw) or peak inspiratory pressure (PIP). However, significant differences in cardiac output (CO), Pa(CO(2)), pH, and delivered VT between the treatment groups in their study could have influenced these results. Because of the potential implications of these findings for patient care, we attempted to confirm these findings by Lefevre and colleagues in a canine model of oleic acid induced lung injury. Eighteen mongrel dogs were anesthetized in the supine position, paralyzed, and mechanically ventilated with 50% O(2) at f = 15 breaths/min, and VT was adjusted to achieve an end-tidal CO(2) of 30 to 35 mm Hg. Lung injury was produced by infusion of 0.06 ml/kg oleic acid solution into the right atrium over a 30-min period. Animals were then randomized to either conventional ventilation at the baseline settings (n = 9) or to BV at the same mean VT and f (n = 9). Both groups received comparable degrees of injury, and hemodynamic and ventilatory parameters were closely matched, with no differences in mean VT, PIP, mean Paw, Pa(CO(2)), pH, CO, pulmonary artery occlusion pressure, or arterial pressure (Pa). However, no differences between the two groups were found in Pa(O(2)), shunt, or static compliance over a 4-h period. When hemodynamic and ventilatory parameters were well matched in a canine model of ALI, BV showed no advantage over conventional ventilation at constant VT and f. PMID- 10852748 TI - Lung function response to cold air challenge in asthmatic and healthy children of 2-5 years of age. AB - The aim of the study was to assess feasibility, sensitivity, specificity, predictive value, and repeatability of cold, dry air challenge (CACh) as a diagnostic test for asthma in young children 2 to 5 yr of age. Response to a 4 min single-step isocapnic CACh was measured in 38 asthmatics and 29 control subjects. Specific airway resistance (sRaw) by whole body plethysmography was the primary outcome. In addition, lung function was measured as respiratory resistance by the interrupter technique (Rint) and respiratory resistance and reactance at 5 Hz (Rrs5, Xrs5) by the impulse oscillation technique. At baseline, lung function measures differed significantly between asthmatics and healthy control subjects. CACh was readily performed in young children. Response was expressed as change from baseline in numbers of within-subject standard deviation (SDw). Hyperresponsiveness defined as change in lung function of more than 3 SDw was detected by sRaw in 26 of 38 asthmatics versus 2 of 29 control subjects, by Rint in 12 of 38 asthmatics versus 1 of 29 control subjects, by Xrs5 in 9 of 38 asthmatics versus zero of 29 control subjects and by Rrs5 in 7 of 38 asthmatics versus 1 of 29 control subjects. Thus sRaw had the highest sensitivity (68%). Specificity ranged from 93 to 100%. The correlation coefficient between sRaw responses to CACh repeated within 8 wk was 96%. In conclusion, CACh is feasible in young children age 2 to 5 yr. Whole body plethysmography (sRaw) was superior in separating asthmatics from healthy control subjects. Change in sRaw in response to CACh may be used as a diagnostic test for asthma in young children. PMID- 10852749 TI - A parental history of asthma is a risk factor for wheezing and nonwheezing respiratory illnesses in infants younger than 18 months of age. AB - The relationship between respiratory infection and allergy as risk factors for the development of wheezing illnesses in infants has been in dispute. We hypothesized that a parental history of allergic diseases would be associated with an increased rate of respiratory infections as well as an increased rate of wheezing during infectious episodes. We prospectively evaluated 1,193 infants from birth to 18 mo of age, using bi-weekly telephone surveillance to document all respiratory events. The overall rate of respiratory illness (all RI) increased to a maximum of 10.6 illnesses/infant/year in the 7- to 9-mo age group and then leveled off in the older infants. Multivariable models adjusting for demographic variables, breast feeding, month of illness, number of siblings, and attendance at day care showed an increase in the rate of all RI in infants older than 7 mo of age who had a parental history of asthma (OR = 1.24, CI = 1.09 to 1.41) or a parental history of atopy (OR = 1.14, CI = 1.03 to 1.26). The rate of lower respiratory illnesses accompanied by wheezing was related only to a parental history of asthma (OR = 2.06, CI = 1.36 to 3.11). We conclude that all RI, most of which represent viral infections, are increased in otherwise normal infants with a parental history of asthma or atopy, whereas wheezing is related only to a parental history of asthma. PMID- 10852750 TI - A simple "new" method to accelerate clearance of carbon monoxide. AB - The currently recommended prehospital treatment for carbon monoxide (CO) poisoning is administration of 100% O(2). We have shown in dogs that normocapnic hyperpnea with O(2) further accelerates CO elimination. The purpose of this study was to examine the relation between minute ventilation (V E) and the rate of elimination of CO in humans. Seven healthy male volunteers were exposed to CO (400 to 1,000 ppm) in air until their carboxyhemoglobin (COHb) levels reached 10 to 12%. They then breathed either 100% O(2) at resting V E (4.3 to 9.0 L min) for 60 min or O(2) containing 4.5 to 4.8% CO(2) (to maintain normocapnia) at two to six times resting V E for 90 min. The half-time of the decrease in COHb fell from 78 +/- 24 min (mean +/- SD) during resting V E with 100% O(2) to 31 +/- 6 min (p < 0. 001) during normocapnic hyperpnea with O(2). The relation between V E and the half-time of COHb reduction approximated a rectangular hyperbola. Because both the method and circuit are simple, this approach may enhance the first-aid treatment of CO poisoning. PMID- 10852751 TI - Lung function growth and its relation to airway hyperresponsiveness and recent wheeze. Results from a longitudinal population study. AB - To evaluate the association between growth in height and growth in lung function, and to identify the potential temporal relationships between airway hyperresponsiveness (AHR), respiratory symptoms, and lung function growth during adolescence and young adulthood, we analyzed data collected from the Belmont cohort. Among the 718 schoolchildren initially studied at 1982 (aged 8-10 yr), 557 were studied between two times and six times at 2-yr intervals until 1992. Baseline lung function, AHR by histamine inhalation test, and recent wheeze by questionnaires, were measured at each visit. We found that between 17 and 19 yr of age, when growth in height had stopped, growth in FEV(1) was approximately 200 ml/yr in boys and 100 ml/yr in girls. Peak growth velocity of height occurred at age 13 both in boys and in girls, whereas peak growth velocity of FEV(1) occurred at the same age only in girls and 1 yr later in boys. Having AHR and recent wheeze at the previous study time were both associated with lower subsequent growth in FEV(1), but not with subsequent growth in FVC. We conclude that lung function continues to grow after the cessation of height growth and that growth in FEV(1) is reduced in subjects with AHR and/or recent wheeze. PMID- 10852752 TI - Alterations in airway wall properties in infants with a history of wheezing disorders. AB - Airway diameter and airway wall mechanics (compliance) are important determinants of flow limitation and wheezing. We have previously used the high-speed interrupter technique (HIT) to measure input impedance (Zin) in infants at frequencies up to 900 Hz, including antiresonance phenomena, which are known to be related to wave propagation velocity, and have shown that the frequency at which the first antiresonance occurs (f(ar,1)) is a function of airway wall compliance. We aimed to determine whether f(ar,1) (and thus airway wall compliance) was different in infants with a history of wheezing disorders. We compared 23 asymptomatic infants (aged 36 to 81 wk) with a history of wheezing with an age-matched group of 19 healthy control infants. We found that f(ar,1) was significantly lower in infants with wheezing disorders than in the control group (p < 0. 005), implying differences in airway wall compliance, even when they were clinically asymptomatic. Developmental differences in airway wall mechanics may be important in the pathogenesis of wheezing disorders or, alternatively, alterations in airway wall mechanics might be a consequence of postinflammatory remodeling. PMID- 10852753 TI - Respiratory-related evoked potentials in children with life-threatening asthma. AB - Respiratory-related evoked potentials (RREPs) have been elicited by inspiratory occlusion and recorded over the somatosensory cortex. The first positive peak (P(1)) amplitude has been correlated with the magnitude of inspiratory loads. Since children with life-threatening asthma (LTA) have a decreased perceptual sensitivity of inspiratory loads, we hypothesized that a subpopulation of patients with LTA have an impaired ability to sense mechanical loads, and that these patients would have an abnormal RREP. The RREP was recorded from C(Z) -C(3) and C(Z) -C(4) in three groups: LTA asthmatic, control asthmatic, and nonasthmatic children. Two inspiratory-interruption occlusions trials and a control trial were recorded. All the evoked potentials were analyzed after the averaged control trial was subtracted from the averaged occlusion trials. The RREP P(1) peak was observed in all 14 nonasthmatic children and in 14 of 15 control asthmatic children. The RREP was absent in 6 of 11 patients with LTA. When present, there were no between-group significant differences in P(1) peak latency or amplitude. These results demonstrate that the RREP elicited by inspiratory occlusion is present bilaterally in nonasthmatic and asthmatic children. There is a subpopulation of LTA children in which inspiratory occlusion fails to elicit the P(1) peak of the RREP, suggesting an altered neural processing of inspiratory load information. PMID- 10852754 TI - Independent inheritance of serum immunoglobulin E concentrations and airway responsiveness. AB - Elevated serum Immunoglobulin E (IgE) levels and increased airway responsiveness (AR) are correlated traits that are characteristic of asthma. It is not known to what extent these traits arise from distinct or shared genetic determinants. We investigated the genetic and environmental components of variance of serum total and specific IgE levels and AR in an Australian population-based sample of 232 Caucasian nuclear families. The inter-relationships of the genetic determinants of these traits were also investigated. Log(e) total serum IgE levels had a narrow-sense heritability (h(2)(N)) of 47.3% (SE = 10.0%). Specific serum IgE levels against house dust mite and timothy grass, measured as a RAST Index, ad a h(2)(N) of 33.8% (SE = 7.3%). AR, quantified by the log(e) dose-response slope to methacholine (DRS), had a h(2)(N) of 30.0% (SE = 12.3%). Extended modeling demonstrated an approximate 70% overlap in the genetic determinants of total and specific serum IgE levels. The genetic determinants of serum IgE levels and AR exhibited less than 30% sharing. These data are consistent with the existence of multiple genetic determinants of the pathophysiologic traits associated with asthma, and suggest that AR is genetically distinct from atopy. These results have implications for gene discovery programs. PMID- 10852755 TI - Effect of eotaxin and platelet-activating factor on airway inflammation and hyperresponsiveness in guinea pigs in vivo. AB - Although eotaxin causes selective infiltration of eosinophils into the lung, its role in airway hyperresponsiveness remains unclear. We studied the effects of local administration of eotaxin on airway inflammation and hyperresponsiveness in guinea pigs in vivo. Airway responsiveness to inhaled histamine and differential cell counts in bronchoalveolar lavage fluid (BALF) were evaluated 12 h, 24 h, 3 d, and 7 d after intratracheal instillation of eotaxin. Significant eosinophilia in BALF was observed between 6 h and 7 d after eotaxin administration. Histologically, eosinophil accumulation was observed in the airways but not in the alveoli. In contrast, eotaxin did not affect airway responsiveness between 12 h and 7 d after its administration. We then studied the effects on airway responsiveness of subthreshold doses of interleukin 5, leukotriene D(4) (LTD(4)), and platelet-activating factor (PAF) combined with eotaxin. Neither interleukin 5 nor LTD(4) affected airway responsiveness. After eotaxin treatment, PAF significantly enhanced airway responsiveness without further increases in eosinophil counts. Eotaxin plus PAF significantly increased in eosinophil peroxidase activity in BALF compared with control and with eotaxin alone. These data indicate that eotaxin alone causes eosinophil accumulation in the airways but not hyperresponsiveness, and that additional factors such as PAF are needed to activate eosinophils for the development of airway hyperresponsiveness. PMID- 10852756 TI - Heparin inhibits eicosanoid metabolism and hyperventilation-induced bronchoconstriction in dogs. AB - Inhalation of heparin, an anticoagulant, attenuates exercise- induced asthma (EIA) in human subjects. The purpose of this study was to determine if heparin inhibits hyperventilation-induced bronchoconstriction (HIB) in a canine model of EIA, and if its mode of action involves the inhibition of eicosanoid mediator production and release. We used a wedged bronchoscope technique to measure baseline peripheral airway resistance (Rp). We then performed either a 2-min or 5 min dry air challenge (DAC) by temporarily increasing from 200 to 2,000 ml/min the flow of 5% CO(2) in air used to ventilate a wedged sublobar segment. We compared HIB before and 60 min after aerosol treatment with either bacteriostatic water (BW) or heparin. We found that (1) heparin had no effect on baseline Rp, (2) BW did not alter the response to DAC, and (3) heparin reduced HIB by approximately 50-60%. On the basis of bronchoalveolar lavage fluid (BALF) cell analysis, heparin and BW caused acute infiltration of macrophages and eosinophils, and heparin increased the number of erythrocytes recovered immediately after DAC. Despite these acute inflammatory effects initiated prior to DAC, BALF mediator analyses revealed that pretreatment with heparin either attenuated or abolished hyperventilation-induced leukotriene, prostaglandin, and thromboxane release. Thus, our data provide direct evidence that inhaled heparin inhibits eicosanoid mediator production and release caused by hyperventilation with dry air, and significantly attenuates HIB. PMID- 10852757 TI - The effect of repeated ozone exposures on inflammatory markers in bronchoalveolar lavage fluid and mucosal biopsies. AB - The aim of this study was to investigate the cellular and biochemical events associated with repeated exposures to ozone. Twenty-three healthy subjects underwent single exposures to 200 ppb ozone and to filtered air (FA), as well as repeated exposures to 200 ppb ozone on 4 consecutive days, each for 4 h of intermittent exercise. Bronchoalveolar lavage was performed and mucosal biopsies were taken 20 h after the single or the last of the repeated exposures. As compared with FA, the single exposure to ozone caused a decrease in FEV(1), an increase in the percentages of neutrophils and lymphocytes, the concentrations of total protein, IL-6, IL-8, reduced glutathione, urate, and ortho-tyrosine in BAL fluid (BALF), but no changes in the cellular composition of biopsy. After the repeated exposure, the effect on lung function was abolished and differential cell counts in BALF were not significantly different from those after FA. However, the concentrations of total protein, IL-6, IL-8, reduced glutathione, and ortho-tyrosine were still increased. IL-10 could only be detected in BALF after repeated ozone exposures. Furthermore, macroscopic scores for bronchitis, erythema, and hypervulnerability of airway mucosa were increased, as well as numbers of neutrophils in bronchial mucosal biopsies. Our data demonstrate that airway inflammation persists after repeated ozone exposure, despite attenuation of some inflammatory markers in BALF and adaptation of lung function. PMID- 10852758 TI - First-line therapy for adult patients with acute asthma receiving a multiple-dose protocol of ipratropium bromide plus albuterol in the emergency department. AB - We designed a larger, double-blind, randomized, prospective trial to test our hypothesis that patients with acute asthma given combination high dose therapy with ipratropium bromide (IB) and beta(2)-agonists will have greater improvement in pulmonary function and fewer hospital admissions than those given beta(2) agonists alone. One hundred eighty patients (mean age +/- SD, 34.3 +/- 10.5 yr) who presented to an emergency department (ED) for treatment of an exacerbation of asthma (baseline FEV(1) < 50% of predicted) were assigned in a randomized, double blind fashion to receive albuterol and placebo (n = 92) or albuterol and IB (n = 88). Both drugs were administered through a metered-dose inhaler and spacer at 10 min intervals for 3 h (24 puffs or 2,880 microg of albuterol and 504 microg of IB each hour). Primary outcome measures were improvement in pulmonary function (FEV(1) or peak expiratory flow [PEF]), and hospital admission rates. In both groups, pulmonary function improved significantly over baseline values (p < 0.01). Subjects who received IB had an overall 20.5% (95% CI: 2.6 to 38.4%) (p = 0.02) greater improvement in PEF and a 48.1% (95% CI: 19.8 to 76.4%) (p = 0.001) greater improvement in FEV(1) from the control group. At the end of protocol (3 h), 39% (n = 36) of patients in the control group and 20% (n = 18) in the IB group were admitted (p = 0.01). The use of high doses of IB reduced the risk of hospital admission 49% (relative risk = 0.51, 95% CI: 0.31 to 0.83). Five (95% CI: 3 to 17) patients would need to be treated with high doses of IB to prevent a single admission. Kaplan-Meier-estimated curves of the proportion of patients who reached the discharge threshold during the 3 h of treatment, showed a significant difference in favor of the IB group (log-rank test = 0.005). A subgroup analysis showed that patients most likely to benefit from the addition of high doses of IB were those with more severe obstruction (FEV(1) /= 24 h). On the contrary, previous use of inhaled beta(2)-agonists did not modify the admission rate and the pulmonary function response to IB. In conclusion, our data support a substantial therapeutic benefit from the addition of IB to albuterol administered in high doses through MDI plus spacer, particularly in patients with FEV(1) less than 30%, and with long duration of symptoms before the ED presentation (>/= 24 h). PMID- 10852759 TI - High-flow transtracheal insufflation treats obstructive sleep apnea. A pilot study. AB - To determine the effect of transtracheal insufflation (TTI) on obstructive sleep apnea (OSA), we examined breathing patterns in five tracheostomized patients with OSA at varying TTI flow rates when breathing with a closed tracheostomy. The breathing patterns and polysomnographic responses to air insufflation were studied as TTI was increased from 0 to 15 L/min for brief periods of non-rapid eye movement (NREM) sleep (Experiment 1). The frequency of sleep-disordered breathing episodes remained high at 0 and 5 L/min (87.0 +/- 33.7 and 79.4 +/- 24.4 episodes per hour NREM) and decreased significantly to 41.3 +/- 31.5 and 43.4 +/- 31.4 episodes/h NREM sleep at rates of 10 and 15 L/min, respectively (p = 0.003). At high levels of TTI (10 and 15 L/min), obstructive apneas and hypopneas decreased but periodic laryngeal obstructions were induced during stage 1 NREM sleep. To prevent laryngeal obstructions, a servo-control system was used to briefly interrupt TTI during these events. When this system was implemented for more prolonged periods of sleep (Experiment 2, total sleep time 176.6 +/- 12.5 min), high-flow TTI (hf-TTI, 15 L/min) led to an overall reduction in the combined frequency of obstructive apneas and laryngeal obstructions from 63.8 +/- 21.8 to 10.7 +/- 9.1 (p < 0.03) and was associated with a marked reduction in arousal frequency from 60.0 +/- 26.0 to 8. 3 +/- 5.4/h in NREM sleep, and from 67.5 +/- 3.5 to 0 +/- 0/h in rapid eye movement (REM) sleep. Our findings demonstrate that hf-TTI stabilized breathing patterns in apneic patients, and was safe and efficacious for prolonged periods of sleep. PMID- 10852760 TI - Serum vascular endothelial growth factor is elevated in cystic fibrosis and decreases with treatment of acute pulmonary exacerbation. AB - Chronic bacterial infection and neutrophilic inflammation characterize cystic fibrosis (CF) pulmonary disease. In many disorders, inflammation and angiogenesis are codependent phenomena. We previously noted excessive angiogenesis in CF tissues and elevated vascular endothelial growth factor (VEGF) in random serum samples from subjects with CF. To further explore this finding, we measured serum VEGF in 38 subjects with stable CF and in 25 subjects with other pulmonary diseases. Mean VEGF was elevated in both groups compared with reference values, but it was higher in CF: 403 +/- 280 versus 255 +/- 169 pg/ml, p = 0.02. VEGF was negatively correlated with FEV(1) in CF, r = -0.51, p = 0.007. To assess the effect of airway infection on VEGF, 10 subjects with CF were studied before and after intravenous antibiotic therapy for pulmonary exacerbation. VEGF levels decreased with antibiotic therapy, from 537 +/- 220 to 259 +/- 176 pg/ml, p = 0.001. We conclude that circulating VEGF is increased in subjects with CF and other inflammatory pulmonary disorders. In CF, VEGF elevation is related to airway infection. We speculate that increased circulating VEGF is related to chronic inflammation, which is robust in CF. Elevated circulating VEGF may result in tissue angiogenesis, furthering the progression of pulmonary disease. PMID- 10852761 TI - Leukotriene receptor antagonists and synthesis inhibitors reverse survival in eosinophils of asthmatic individuals. AB - Eosinophilia is a feature of airway inflammation associated with asthma. Leukotriene antagonists provide therapeutic benefit in asthma, but their potential antiinflammatory actions have not been fully explored. We have examined the role of eosinophil-derived cysteinyl leukotrienes in the maintenance of eosinophil survival, and the involvement of leukotrienes in the paracrine stimulation of eosinophil survival by mast cells and lymphocytes. We obtained eosinophils and autologous lymphocytes from peripheral blood of asthmatic subjects. Leukotriene (LT)-B(4), LTC(4) and LTD(4), granulocyte-macrophage colony stimulating factor (GM-CSF), and fibronectin promoted eosinophil survival. LTD(4) (10(-)(6) M) was as effective as GM-CSF (5 ng/ml) and fibronectin (400 ng/ml) in promoting survival. Lymphocytes and conditioned medium from a human mast cell line (HMC-1) induced eosinophil survival. Blockade of cysteinyl leukotriene receptors with SKF 104353 (pobilukast, 3 nM), and inhibition of 5-lipoxygenase (5 LO) with BW A4C (1 microM) and of 5-LO activating protein with MK 886 (1 microM), all increased basal rates of eosinophil apoptosis and reversed GM-CSF-induced eosinophil survival. Fifty percent reversal of GM-CSF- induced survival was achieved with SKF 104353 at 0.3 nM. The potency of SKF 104353 was two orders of magnitude greater than that of the LTB(4) receptor antagonist SB 201146. Mast cell- and lymphocyte-induced eosinophil survival were completely reversed by SB 201146, SKF 104353, BW A4C, and MK 886. These findings provide evidence for the involvement of an autocrine cysteinyl leukotriene pathway that supports eosinophil survival in response to a range of survival stimuli. They also suggest that LTB(4) could act as a paracrine stimulus of eosinophil survival. PMID- 10852762 TI - Peak expiratory flow profiles delivered by pump systems. Limitations due to wave action. AB - Pump systems are currently used to test the performance of both spirometers and peak expiratory flow (PEF) meters, but for certain flow profiles the input signal (i.e., requested profile) and the output profile can differ. We developed a mathematical model of wave action within a pump and compared the recorded flow profiles with both the input profiles and the output predicted by the model. Three American Thoracic Society (ATS) flow profiles and four artificial flow versus-time profiles were delivered by a pump, first to a pneumotachograph (PT) on its own, then to the PT with a 32-cm upstream extension tube (which would favor wave action), and lastly with the PT in series with and immediately downstream to a mini-Wright peak flow meter. With the PT on its own, recorded flow for the seven profiles was 2.4 +/- 1.9% (mean +/- SD) higher than the pump's input flow, and similarly was 2.3 +/- 2.3% higher than the pump's output flow as predicted by the model. With the extension tube in place, the recorded flow was 6.6 +/- 6.4% higher than the input flow (range: 0.1 to 18.4%), but was only 1.2 +/- 2.5% higher than the output flow predicted by the model (range: -0.8 to 5.2%). With the mini-Wright meter in series, the flow recorded by the PT was on average 6.1 +/- 9.1% below the input flow (range: -23.8 to 2. 5%), but was only 0.6 +/- 3.3% above the pump's output flow predicted by the model (range: -5.5 to 3.9%). The mini-Wright meter's reading (corrected for its nonlinearity) was on average 1.3 +/- 3.6% below the model's predicted output flow (range: -9.0 to 1. 5%). The mini-Wright meter would be deemed outside ATS limits for accuracy for three of the seven profiles when compared with the pump's input PEF, but this would be true for only one profile when compared with the pump's output PEF as predicted by the model. Our study shows that the output flow from pump systems can differ from the input waveform depending on the operating configuration. This effect can be predicted with reasonable accuracy using a model based on nonsteady flow analysis that takes account of pressure wave reflections within pump systems. PMID- 10852763 TI - The effects of oxitropium bromide on exercise performance in patients with stable chronic obstructive pulmonary disease. A comparison of three different exercise tests. AB - The purpose of the present study was to compare the characteristics of three different exercise tests in evaluating the effects of oxitropium bromide on exercise performance. Thirty-eight males with stable chronic obstructive pulmonary disease (COPD) (FEV(1) = 40.8 +/- 16.5% predicted; mean +/- SD) completed randomized, double-blind, placebo-controlled, crossover studies for each exercise test. The exercise tests were performed 60 min after the inhalation of either oxitropium bromide 400 microg or placebo. The patients performed 6-min walking tests (6MWT) on Days 1 and 2, progressive cycle ergometry (PCE) on Days 3 and 4, and cycle endurance tests at 80% of the maximal workload of PCE on Days 5 and 6. Spirometry was conducted before and at 45 and 90 min after the inhalation. Oxitropium bromide significantly increased FEV(1) as compared with placebo. Oxitropium bromide increased the endurance time significantly, by 19% (p < 0.001), and caused a small but significant increase in the 6-min walking distance by 1% (p < 0.05), but induced no significant increase in maximal oxygen consumption (V O(2)max) in PCE. The responses in these three exercise tests were different, and we conclude that the endurance test was the most sensitive in detecting the effects of inhaled anticholinergic agents on exercise performance in patients with stable COPD. An endurance procedure may be performed to detect clinical changes in evaluating the effects of oxitropium bromide on exercise performance. PMID- 10852764 TI - Recurrent exacerbations in severe asthma are associated with enhanced airway closure during stable episodes. AB - Excessive airway narrowing is a cardinal feature of asthma, and results in closure of airways. Therefore, asthmatic patients in whom airway closure occurs relatively early during expiration might be prone to severe asthma attacks. To test this hypothesis, we compared closing volume (CV) and closing capacity (CC) in a group of asthmatic patients with recurrent exacerbations (more than two exacerbations in the previous year; difficult-to-control asthma), consisting of 11 males and two females, aged 20 to 51 yr, with those in a group of equally severely asthmatic controls without recurrent exacerbations (stable asthma) consisting of 13 males and two females aged 18 to 52 yr. Both groups used equivalent doses of inhaled corticosteroids and were matched for sex, age, atopy, postbronchodilator FEV(1), and provocative concentration of methacholine causing a 20% decrease in FEV(1). They were studied during a clinically stable period of their disease. The patients inhaled 400 microg salbutamol via a spacer device, after which TLC and RV were measured by multibreath helium equilibration, together with the slope of Phase 3 (dN(2)), CV, and CC, by single-breath nitrogen washout. CV and CC were expressed as ratios of VC and TLC, respectively, and all data are presented as % predicted (mean +/- SEM). There was no difference in TLC in patients with difficult-to-control asthma and those with stable asthma (106.7 +/- 4.0% predicted versus 101.7 +/- 4.3% predicted, p = 0.40), RV (113.1 +/- 7.8% predicted versus 100.9 +/- 7.1% predicted, p = 0.26), or dN(2) (142.7 +/- 16.3% predicted versus 116.0 +/- 20.2% predicted, p = 0.23). In contrast, CV and CC were increased in the patients with difficult-to-control asthma as compared with the group with stable asthma (CV: 159.5 +/- 26.8% predicted versus 98.8 +/- 12.5% predicted, p = 0.024; CC: 114.0 +/- 6.4% predicted versus 99.9 +/- 3. 6% predicted, p = 0.030). These findings show that asthmatic individuals with recurrent exacerbations have increased CV and CC as compared with equally severely asthmatic but stable controls, even after bronchodilation during well controlled episodes. The findings imply that airway closure at relatively high lung volumes under clinically stable conditions might be a risk factor for severe exacerbations in asthmatic patients. PMID- 10852765 TI - The effect of glutathione and N-acetylcysteine on lipoperoxidative damage in patients with early septic shock. AB - Both the hyperproduction of oxygen free radicals (OFR) and the weakening of natural scavenging mechanisms have been implicated as contributors to multiple organ failure in septic shock. This study examined whether the antioxidants glutathione (GSH) and N-acetyl-L-cysteine (NAC) play a protective role against damage by OFR in early septic shock. We randomly entered 30 patients with septic shock into one of three groups within 24 h of diagnosis. All of the patients received septic shock therapy, including parenteral nutrition, antibiotics, and volume-expanding and inotropic agents. One group (Group B) also received 70 mg/kg/d of intravenous GSH, and a second group (Group C), 70 mg/kg/d of intravenous GSH and 75 mg/kg/d of intravenous NAC. The protection against OFR damage was evaluated by measuring expired ethane, plasma malondialdehyde, erythrocyte deformability, complement activation, and clinical scores at admission and on Days 3 and 5 of treatment. A significant decrease in peroxidative indexes was observed at Day 5 in Group B as compared with both the control group and basal values. The decrease in peroxidative indexes was even more marked in Group C. Clinical scores in this group were also significantly improved. In conclusion, the administration of high doses of NAC added to GSH significantly decreased the peroxidative stress of patients with septic shock. PMID- 10852766 TI - Effect of unplanned extubation on outcome of mechanical ventilation. AB - Unplanned extubation is a major complication of translaryngeal intubation, but its impact on mortality, duration of mechanical ventilation (MV), length of intensive care unit (ICU) and hospital stay, and need for ongoing hospital care has not been adequately defined. We performed a case-control study in a tertiary care medical ICU, comparing 75 patients with unplanned extubation and 150 controls matched for Acute Physiology and Chronic Health Evaluation II score, presence of comorbid conditions, age, indication for MV, and sex. Forty-two (56%) patients required reintubation after unplanned extubation (74% immediately, 86% within 12 h). Thirty-three (44%) unplanned extubations occurred during weaning trials, and 30% of these patients needed reintubation (failed unplanned extubation). In contrast, 76% of patients with unplanned extubation occurring during ventilatory support required reintubation. Although mortality was similar to that of controls (failed unplanned extubation 40%, versus control 31%, p > 0.2), patients with failed unplanned extubation had a significantly longer duration of MV (19 versus 11 d, p < 0.01), longer stay in the ICU (21 versus 14 d, p < 0.05), and longer hospital stay (30 versus 21 d, p < 0.01), and survivors were more likely to require chronic care (64% versus 24%, p < 0.001). Successfully tolerated unplanned extubation was associated with a reduction in time from beginning of weaning to extubation (0.9 versus 2.0 d, p = 0.06), but with no difference in overall duration of MV, mortality, discharge location, ICU, or hospital stay as compared with these measures for controls. We conclude that unplanned extubation is not associated with increased mortality when compared with that of matched controls, although it does result in prolonged MV, longer ICU and hospital stay, and increased need for chronic care. These effects are due exclusively to patients who fail to tolerate unplanned extubation. Although successfully tolerated unplanned extubation decreased the duration of weaning trials, it had no other measurable beneficial impact on outcome. PMID- 10852767 TI - Synthesis of arachidonic acid-derived lipoxygenase and cytochrome P450 products in the intact human lung vasculature. AB - Lipoxygenase (LO) and cytochrome P450 monooxygenase products of arachidonic acid (AA) have been implicated in a large number of vasoregulatory processes. In intact, blood-free, perfused and ventilated human lungs (n = 8), isolated during surgery for bronchial carcinoma, we analyzed leukotrienes (LTs), hydroxyeicosatetraenoic acids (HETEs), and epoxyeicosatrienoic acids (EETs) by sequential sampling of the recirculating buffer fluid. For the analysis we used multistep, solid-phase extraction, isocratic reversed-phase high-performance liquid chromatography, with elution of all metabolites within one run and photodiode array detection to obtain full UV spectra of eluting compounds. We detected no LT release in a 15-min baseline period, but the admixture of the calcium ionophore A23187 with the buffer fluid provoked the rapid appearance of all LTs. Some baseline release of 15-HETE was observed, and in response to A23187, maximum buffer concentrations were noted for 5-HETE, with 8-HETE, 9-HETE, 11-HETE, and 12-HETE being detected at lower levels. Marked baseline liberation of 11,12-EET and 8,9-EET was observed. In response to A23187, high oxirane buffer concentrations were registered, which far surpassed those of LTs and HETEs. The eicosanoid release was paralleled by a limited pulmonary artery pressor response and progressive vascular leakage. We conclude that ex-vivo-perfused human lungs release EETs > LTs > HETEs into the vascular compartment in response to inflammatory challenge. The marked oxirane synthesis in the lung vasculature may have major impact on lung vasoregulation when considering the possible function of these AA epoxides as endothelium-derived hyperpolarizing factors. PMID- 10852768 TI - Early detection of airway involvement in obliterative bronchiolitis after lung transplantation. Functional and bronchoalveolar lavage cell findings. AB - As defined by the International Society for Heart and Lung Transplantation, the diagnosis of posttransplant obliterative bronchiolitis (OB) is based on histopathologic features and/or spirometric staging criteria, using FEV(1) to determine the extent of disease. However, this last parameter reflects an advanced bronchiolar process. The present study investigated whether physiologic parameters reflecting smaller airways dysfunction on one hand, and neutrophils in bronchoalveolar lavage fluid (BALF) on the other hand, could be useful for the earlier detection of bronchiolitis obliterans syndrome (BOS). We analyzed data obtained both from 765 pulmonary function test results and from 467 BALF specimens from 45 patients who survived at least 1 yr after surgery (n = 47, including two retransplantations). Of the transplant procedures, 22 were associated with BOS and 25 were not. The mean delay from transplantation to the diagnosis of BOS was 578 d (range: 122 to 2,619 d). The threshold values of the following parameters were studied: decline in the forced expiratory flow rate at 25% to 75% of FVC (FEF(25-75)) to 3%, and alveolar neutrophilia >/= 20% of the total BALF cell count. Agreement on the diagnosis of BOS (using the decline in FEV(1)) was equally good for each of the four markers (kappa coefficient > 0.65, p < 10(-)(5)). In the OB group, mean delays after the threshold was reached for each of these parameters were 110 d (p = 0.09), 173 d (p = 0.03), 150 d (p = 0.003), and 131 d (p = 0.1), respectively, before the FEV(1) criteria were fulfilled. At the chosen threshold values, the decline in FEF(25-75), increase in DeltaN(2), and development of a substantial alveolar neutrophilia all occurred significantly before a decline in FEV(1) in posttransplant OB. PMID- 10852769 TI - Decline of ambient air pollution and respiratory symptoms in children. AB - Several regional cross-sectional studies have shown a consistently higher prevalence of respiratory disorders in children with exposure to total suspended particulates (TSP) than in children living in less polluted areas. The aim of the present study was to investigate the temporal changes in the prevalence of nonasthmatic respiratory symptoms and diseases in children living in three areas of East Germany during a phase of strong improvement in ambient air quality. Groups of 2,470 and 2,814 school children between 5 and 14 yr, respectively, participated in two regional cross-sectional studies in 1992-1993 and 1995-1996. In the three areas (Hettstedt, Bitterfeld, and Zerbst) examined in the study, the annual mean TSP decreased from 65, 48, and 44 microg/m(3), respectively, in 1993 to 43, 39, and 36 microg/m(3) in 1995. In the same time interval, the crude prevalence of bronchitis in the three respective areas decreased from 62%, 52%, and 50% to 47%, 40%, and 39%. During the 3-yr period between the two regional studies, prevalence decreased significantly for bronchitis (odds ratio [OR]: 0.55; confidence interval [CI]: 0.49 to 0.62), for otitis media (OR: 0.83; CI: 0.73 to 0.96), for frequent colds (OR: 0.74; CI: 0.64 to 0.86), and for febrile infections (OR: 0.76; CI: 0.66 to 0.88) after adjustment for several potential predictors. In conclusion, we found that the prevalence of nonasthmatic respiratory symptoms decreased from the first period to the second period in all three study areas. PMID- 10852770 TI - Bronchiolitis obliterans syndrome after lung transplantation and health-related quality of life. AB - The present study was undertaken to assess the relationship between health related quality of life (HRQOL) and bronchiolitis obliterans syndrome (BOS), as both represent important parameters of outcome after lung transplantation. HRQOL was measured both cross-sectionally and longitudinally by standardized patient self-administered questionnaires, including the Nottingham Health Profile, the State-trait Anxiety Inventory, the Zung Self-Rating Depression Scale, and the Index of Well-Being. Data were collected at 4 and 7 mo, and every 6 mo afterwards for as long as 49 mo post-transplantation. The number of patients who completed the questionnaires varied from 72 at 4 mo, to 27 at 49 mo after transplantation. Cross-sectionally, the patients with BOS reported persistently statistically significantly more restrictions on the dimensions energy and physical mobility of the Nottingham Health Profile compared with patients without BOS. Other domains, i.e., pain, sleep, social interaction, and emotional reactions, were not affected. Additionally, patients with BOS reported statistically significantly more depressive symptoms and anxiety 1 and 2 yr after transplantation. Results from the longitudinal analysis support these findings, although no change in depressive symptoms could be found after onset of BOS. This study suggests that all lung transplant recipients improve in HRQOL. The development of BOS, however, is associated with a significantly reduced HRQOL. PMID- 10852771 TI - Multicenter prospective study of ventilator-associated pneumonia during acute respiratory distress syndrome. Incidence, prognosis, and risk factors. ARDS Study Group. AB - We investigated the incidence, risk factors for, and outcome of ventilator associated pneumonia (VAP) in patients with acute respiratory distress syndrome (ARDS). We compared 134 patients with ARDS with 744 patients without ARDS on mechanical ventilation. Fiberoptic bronchoscopic examination and quantitative bacterial cultures (protected brush or catheter sampling [threshold: 10(3) cfu/ml], or bronchoalveolar lavage [threshold: 10(4) cfu/ml]) were used to diagnose pneumonia. VAP occurred in 49 patients (36.5%). The incidence of pneumonia was 23% (173 of 744 patients) among patients without ARDS (p < 0.002). Nonfermenting gram-negative rods caused significantly more pneumonia in ARDS patients. Mortality rates were identical in ARDS patients with (28 of 49 patients, 57%) and without (50 of 85 patients, 59%) pulmonary infection (p = 0.8). VAP resulted in a considerable increase in attributable time on mechanical ventilation of both the overall population of ARDS patients and of survivors. Both the use of sucralfate (adjusted odds ratio [OR]: 4. 42; 95% confidence interval [CI]: 2.01 to 9.7, p = 0.0002) and the duration of exposure to sucralfate (adjusted OR: 1.206; 95% CI: 1. 095 to 1.328, p = 0.0002) were associated with an increased risk of VAP during ARDS. VAP considerably prolongs the time on mechanical ventilation without affecting survival. Patients given sucralfate may be at greater risk of developing pulmonary infection during ARDS. PMID- 10852772 TI - Oxygenation response to a recruitment maneuver during supine and prone positions in an oleic acid-induced lung injury model. AB - Prone position and recruitment maneuvers (RM) are proposed as adjuncts to mechanical ventilation to open up the lung and keep it open. We studied the oxygenation response to a RM (composed of a 30-s sustained inflation at 60 cm H(2)O airway pressure) performed in prone and supine positions in dogs after oleic acid- induced lung injury using an inspired O(2) fraction of 0.60. In one group (n = 6) first supine then prone positions were examined after a RM at 8 cm H(2)O and 15 cm H(2)O of positive end-expiratory pressure (PEEP). In the second group (n = 6) the sequence of positions was reversed. Prone positioning after supine position always improved oxygenation, whereas the decrement in Pa(O(2)) was relatively small when dogs were returned to the supine position. Oxygenation improved in both groups after a RM, and the improvement was sustained (after 15 min) in the prone position at 8 cm H(2)O of PEEP, but 15 cm H(2)O of PEEP was required in supine position. Our results suggest that a RM improves oxygenation more effectively with a decreased PEEP requirement for the preservation of the oxygenation response in prone compared with supine position. PMID- 10852773 TI - Closing volume influences the postural effect on oxygenation in unilateral lung disease. AB - In normal adults, both blood flow and ventilation are distributed preferentially to the dependent lung zones. In adults with unilateral lung disease, arterial oxygenation improves when they are positioned with their good lung down because of improved matching of ventilation and perfusion. When the closing volume is increased, dependent airways are closed during tidal breathing, so that reduced ventilation-perfusion ratio and hypoxia develops and ventilation is preferentially distributed to the upper lung zones. We undertook an observational study on the effects of lateral recumbency on arterial oxygenation in adult patients with unilateral lung disease and tested the hypothesis that oxygenation in lateral recumbency might be influenced by an increase in closing volume. Arterial blood gases were analyzed in the supine, right and left lateral decubitus positions and the AaPO(2) was calculated in 44 randomly selected patients 49.9 +/- 18.7 yr of age with unilateral pneumonia (23 cases) or pulmonary tuberculosis (21 cases). In 26 patients, individual Pa(O(2)) with the normal lung in the dependent position was higher than that with the diseased lung; the opposite was true for 18 patients. The difference in Pa(O(2)) and AaPO(2) between the two positions was statistically significant in both groups. In 16 patients (10 men and six women 49.2 +/- 18.2 yr of age), we measured closing volume and determined the fractional ventilation to each lung by (133)Xe lung scan in the three positions. In these 16 patients, the difference in Pa(O(2)) between the normal and the diseased lung in the dependent position was related significantly to the difference in the fractional ventilation going to the normal lung between the dependent and the supine position (r = 0.642, p = 0. 007). The latter was related significantly to the % predicted closing volume (CV/VC) (r = -0.597, p = 0.015). This study has shown that closing volume, as well as posture, might be involved in determining oxygenation in lateral recumbency in patients with unilateral lung disease. PMID- 10852774 TI - Inhaled furosemide greatly alleviates the sensation of experimentally induced dyspnea. AB - Furosemide is known to influence the activity of vagally mediated mechanoreceptors in the airways. Because vagal afferent fibers may play an important role in modulation of the sensation of dyspnea, it is possible that inhaled furosemide may modify the sensation of dyspnea. In a double-blind, randomized, crossover study, we compared the effect of inhaled furosemide on dyspneic sensation with that of placebo. Severe dyspneic sensation was induced in 12 healthy subjects in two ways: (1) breathholding and (2) loaded breathing with a combination of inspiratory resistive load (240 cm H(2)O/L/s) and hypercapnia induced by extra mechanical dead space (0.26 L). Subjects were asked to rate their sensation of respiratory discomfort using a visual analogue scale (dyspneic VAS). Breathholding times and changes in dyspneic VAS score during a 5-min period of loaded breathing were measured after inhalation of placebo and furosemide (40 mg). Total breathholding time after inhalation of furosemide (median, 93 [interquartile range, 78 to 112]s) was prolonged compared with the total breathholding time after placebo inhalation (67 [47-74]s). We also found that respiratory discomfort during loaded breathing after inhalation of furosemide develops more slowly and is less than that observed after inhalation of placebo. Our findings indicate that inhaled furosemide greatly alleviates the sensation of dyspnea induced experimentally by breathholding and by a combination of resistive loading and hypercapnia. PMID- 10852775 TI - Pulmonary glutathione levels in acute episodes of Farmer's lung. AB - Acute episodes of farmer's lung (FL) are associated with activation and migration of neutrophils into the lungs, causing oxidative stress. We conducted a study to evaluate the effect of episodes of FL on antioxidant defense of the lung by glutathione (GSH). A total of 15 patients with symptomatic FL (one female and 14 males, age 42 +/- 1 yr [mean +/- SEM]) underwent a standardized hay exposure test for 1 h and were then monitored through lung function measurements for 6 h, after which bronchoalveolar lavage (BAL) was performed. As a control, 10 asymptomatic farmers (AF) (two males and eight females, age 43 +/- 1 yr) underwent the same diagnostic procedures. At 3 to 6 h after antigen exposure, the lung function of FL patients was significantly impaired (VC: -31 +/- 4%; single-breath diffusing capacity of carbon monoxide [DL(CO)]: -17 +/- 3%; and Pa(O(2)): -14 +/- 2%, all versus baseline, whereas in AF, only minor changes occurred VC: -4 +/- 5%; DL(CO): -9 +/- 3%, and Pa(O(2)): -5 +/- 2%, all versus baseline). The number of neutrophils in bronchoalveolar lavage fluid was increased in FL patients as compared with AF (29 +/- 7 x 10(4)/ml versus 10 +/- 7 x 10(4)/ml, p < 0.05). The concentrations of total and reduced glutathione (GSH(T) and GSH, respectively) in epithelial lining fluid were decreased in FL patients and increased in AF (GSH(T): 292.5 +/- 27.5 microM versus 1, 185.0 +/- 189.9 microM, respectively, p < 0.001; GSH: 256.8 +/- 22.1 microM versus 1,054.5 +/- 172.9 microM, respectively, p < 0.001). These findings suggest that the individual ability to upregulate GSH in the alveolar space in response to an inflammatory stimulus may have implications for the development of symptomatic FL. We conclude that intrapulmonary GSH levels are distinctly different in patients with FL and AF, and that the regulation of GSH may play an important role in the pathogenesis of FL. PMID- 10852776 TI - The degree of branching of the glycans of alpha(1)-acid glycoprotein in asthma. A correlation with lung function and inflammatory parameters. AB - Alpha(1)-acid glycoprotein (AGP) is a plasma protein belonging to the group of acute-phase proteins. It contains five N-linked glycans which, depending on pathophysiologic state, differ in their degree of branching (i.e., in the relative proportions of di-, tri-, and tetraantennary glycans). Changes in the degree of branching of these glycans have been shown to affect various immunomodulatory properties of AGP. We wanted to investigate whether changes occur in the branching of AGP glycans in plasma and in bronchoalveolar lavage fluid (BALF) in asthma. For this purpose, we selected three groups of patients for study: patients with atopic asthma (AA), atopic nonasthmatic patients, and a group of patients with various interstitial lung diseases (ILDs). The plasma AGP concentration was normal in both atopic study groups, but was increased in ILD patients. In contrast, the branching of glycans of AGP was altered in subjects with AA, whereas it was normal in the other study groups. The presence of asthma symptoms correlated with the increased glycan branching of AGP in both plasma and BALF. Additionally, the degree of branching of AGP in BALF was related to FEV(1), to the provocative dose of histamine causing a 20% decrease in FEV (PD(20)), and to the number of eosinophils. In conclusion, asthma is accompanied by changes in the branching of AGP glycans that indicate an inflammatory reaction that differs markedly from a normal acute-phase response, in which decreased branching of AGP occurs. PMID- 10852777 TI - A model of obstructive sleep apnea in normal humans. Role of the upper airway. AB - We determined whether upper airway obstruction in normal individuals with intact reflexes could produce the syndrome of obstructive sleep apnea. Upper airway obstruction was produced in 12 normal individuals by lowering nasal pressure to 10 cm H(2)O during sleep. Full night polysomnography was performed during two consecutive nights of sleep with subatmospheric nasal pressure and compared with control nights before and after the negative pressure nights. We found that the application of negative pressure was associated with the development of recurrent obstructive apneas (non-REM-disordered breathing rate, 32.6 +/- 34.8 and 37.8 +/- 29.1 events/h during each of two negative pressure nights; p < 0.001) that were associated with oxyhemoglobin desaturation, arousals from sleep, and alterations in sleep stage distribution. Moreover, the median daytime sleep latency after two nights of sleep with subatmospheric pressure fell from 6.9 +/- 1.1 to 3.4 +/- 0.6 min, and rose significantly again to 8.1 +/- 1.5 min (p < 0.03) after the control night following subatmospheric pressure nights. Our findings suggest that a decrease in the pharyngeal transmural pressure alone is a sufficient condition for the production of the sleep apnea syndrome in normal individuals. PMID- 10852778 TI - Nerve growth factor-induced phenotypic switch in guinea pig airway sensory neurons. AB - Immunohistochemistry was combined with retrograde tracing techniques to characterize the effect of nerve growth factor (NGF) on substance P (SP) producing vagal neurons innervating the guinea pig trachea. Fast blue dye instilled into the trachea retrogradely labeled nerve cell bodies located in the nodose and jugular ganglia. In untreated guinea pigs > 99% of the SP-containing neurons labeled with fast blue were located in the jugular ganglia. The SP positive neurons were small in diameter (23 +/- 1 microm) and were negative for neurofilament immunoreactivity. The fast-blue-positive neurons in the nodose ganglia, by contrast, were large in diameter (40 +/- 3 microm) and were negative for SP immunoreactivity and positive for neurofilament immunoreactivity. After NGF-beta injections into the tracheal wall, approximately 10% of the large diameter nodose neurofilament-positive neurons projecting fibers to the trachea became SP-positive (p < 0.05). We previously demonstrated that nodose nerve endings supplying the trachea are exquisitely mechanically sensitive, but capsaicin- and bradykinin-insensitive. These results suggest that NGF not only increases SP expression in airway neurons, but changes the neuronal phenotype such that large, capsaicin-insensitive nodose neurons with fast-conducting "Adelta" fibers provide a component of the tachykinergic innervation. PMID- 10852779 TI - Insulinlike growth factor-1 in lung transplants with obliterative bronchiolitis. AB - Bronchiolitis obliterans syndrome (BOS) is the major complication limiting survival of lung transplant recipients (Tx patients). The mechanisms underlying this fibrotic process are not known. We assessed IGF-1 and IGFBP-3 expression, critical mediators in different models of pulmonary fibrosis, in nine Tx patients. Three of them developed a BOS at 8, 14, and 17 mo postgraft, respectively. Two of the remaining six displayed a recurrent cytomegalovirus (CMV) infection, and four are in stable condition. IGF-1 mRNA expression was quantitated by RT-PCR in cells from four to six BAL per patient performed during the first 6 mo postgraft. Contrasting with a constantly low expression of IGF-1 mRNA in BAL cells from the six patients without BOS, the three patients with BOS presented marked peaks of IGF-1 on two to five occasions during the study period. These peaks, 3- to 13-fold increased compared with values from the former patients, preceded the diagnosis of BOS by 7, 13, and 17 mo, respectively. On the other hand, IGFBP-3 was highly and exclusively expressed in the three patients with BOS, the mRNA as well as the gene product as demonstrated by Western blotting. Our data strongly argue for a role of IGF-1 and IGFBP-3 in the fibrotic process underlying BOS, and for their possible value as an early marker of this complication. PMID- 10852780 TI - Angiotensin II is mitogenic for human lung fibroblasts via activation of the type 1 receptor. AB - The expression of renin-angiotensin system components and the elevation of angiotensin-converting enzyme (ACE) in a number of fibrotic lung diseases suggests angiotensin II (AII) could play a role in the pathogenesis of pulmonary fibrosis. However, the effect of AII on lung fibroblasts has not previously been assessed and the mechanisms by which AII induces cell proliferation in mesenchymal cells are not fully understood. We have examined the ability of AII to stimulate fetal and adult human lung fibroblast proliferation in vitro. In particular, we have assessed the receptor subtypes involved and the possible autocrine role of transforming growth factor beta (TGF-beta) and platelet-derived growth factor (PDGF), two recognized fibroblast mitogens. Angiotensin type 1 (AT1), but not type 2, receptors were identified on fetal and adult human lung fibroblasts by immunocytochemistry. AII (1 microM) increased DNA synthesis (determined by [(3)H]thymidine incorporation) in fetal and adult cells by 211 +/- 18% and 150 +/- 14%, respectively (p < 0.01), and was inhibited by a specific AT1 receptor antagonist, Losartan (74 +/- 14%). A proliferative response to AII was confirmed by direct cell counts. Subsequently, fibroblasts were incubated with neutralizing antibodies to TGF-beta and PDGF. Anti-TGF-beta antibodies inhibited AII-induced DNA synthesis by 73 +/- 13%. However, no effect was seen with anti PDGF antibodies. In conclusion, we have shown that angiotensin II induces human lung fibroblast proliferation in vitro via activation of the AT1 receptor and involves the autocrine action of TGF-beta. PMID- 10852781 TI - Role of the heart in the loss of aeration characterizing lower lobes in acute respiratory distress syndrome. CT Scan ARDS Study Group. AB - In the acute respiratory distress syndrome (ARDS), lower lobes appear essentially non-aerated in contrast to upper lobes whose aeration can be preserved in some patients. The aim of this study was to assess the mechanical compression exerted by the heart on lower lobes in patients with ARDS. Fourteen healthy volunteers and 38 patients with ARDS free of left ventricular failure were studied. Cardiorespiratory parameters were recorded and the cardiac dimensions, the pressure exerted by the heart on subjacent lower lobes, and the gas tissue ratio of lower lobes in the supine position were measured using computer tomography and Lungview, a specifically designed software. In patients with ARDS, the heart was larger and heavier than in healthy volunteers. The enlargement of the heart was mainly related to a left cardiac protrusion and the pressure exerted by the left heart on the lower lobes was higher in patients with ARDS than in healthy volunteers (8 +/- 3 g. cm(-)(2) versus 6 +/- 1 g. cm(-)(2), p < 0.01). As a consequence, the faction of gas represented 62% of the left lower lobes in healthy volunteers and 12% only in patients with ARDS. The present study demonstrates that apart from the already known anteroposterior and cephalocaudal gradients of pressure depending on the lung weight and abdominal pressure, the heart plays an important role in the dramatic loss of aeration characterizing lower lobes of patients with ARDS lying in the supine position. PMID- 10852782 TI - Delayed neutrophil elastase inhibition prevents subsequent progression of acute lung injury induced by endotoxin inhalation in hamsters. AB - To define the role of neutrophil elastase (NE) in the progression of acute lung injury (ALI), we examined the effects of post-treatment with a specific NE inhibitor, sivelestat sodium hydrate (sivelestat), on ALI induced by endotoxin (ET) inhalation in hamsters. Inhalation of ET (300 microg/ml, 30 min) in conscious hamsters increased inflammatory cell count, protein concentration, and hemorrhage in bronchoalveolar lavage fluid (BALF) that peaked 24 h after ET inhalation. These changes were significant 2 h after ET inhalation and paralleled the increase in NE activity in BALF. When intravenously infused from 2 to 24 h post-ET inhalation, sivelestat (0.03 to 3 mg/kg/h) dose-dependently attenuated changes in these BALF parameters at 24 h post-ET inhalation in a manner dependent on the inhibition of NE activity in BALF. Histopathological analysis also indicated that sivelestat prevented the progression of lung inflammation such as alveolar neutrophil infiltration and hemorrhage. In contrast, dexamethasone (3 mg/kg, intravenously) was not effective in this model when administered 2 h after ET inhalation, although it was highly effective when applied before ET. We conclude that delayed inhibition of NE activity with sivelestat prevents subsequent progression of ALI in hamsters after ET inhalation. Thus NE may play an important role in the progression of ALI. PMID- 10852783 TI - Pulmonary angiotensin-converting enzyme (ACE) binding and inhibition in humans. A positron emission tomography study. AB - Angiotensin-converting enzyme (ACE) inhibition attenuates pulmonary hypertension and delays the development of pulmonary vascular remodeling in animal models. Thus, ACE inhibition might be a useful treatment for primary pulmonary hypertension (PPH). To determine the dose of ACE inhibitor required to specifically block pulmonary ACE in humans, we measured the combined forward rate constant (CFRC) for [(18)F]-fluorocaptopril, which is proportional to the mass of ACE in the lung, using positron emission tomography (PET). In five normal subjects, CFRC was measured twice, 1 wk apart, to assess measurement reproducibility. The CFRC was 0.151 +/- 0.067 for the first measurement and 0.140 +/- 0.060 for the second measurement (p = not significant [NS]). In five normals, CFRC decreased on average 84%, from 0.177 +/- 0.053/s to 0.028 +/- 0.017/s (p < 0.05), after 1 wk ingestion of 5 mg enalapril orally once a day (the scans were performed 24 h after the last medication). Similarly, in five patients with PPH, CFRC decreased on average 76%, from 0.052 +/- 0. 020/s to 0.012 +/- 0.003 (p < 0.01), after 1 wk enalapril, despite much lower baseline values. We conclude that the total mass of pulmonary ACE appears to be significantly reduced in PPH and that only low doses of ACE inhibitors may be needed to block the effects of ACE on vascular remodeling in PPH. PMID- 10852784 TI - Mast cell basic fibroblast growth factor in silicosis. AB - To investigate the role of mast cells (MC) and their fibrogenic growth factors in silicosis, we performed quantitative immunohistochemistry for MC tryptase and for basic fibroblast growth factor (bFGF) in lung tissue from silicotic and control subjects. Anti-bFGF antibody was bound to lung MC, basement membrane, endothelial cells, and smooth-muscle cells. Morphometric analysis revealed that the volume density (V(v)) of MC was increased in silicotic lung and that the V(v) of bFGF positive (bFGF(+)) cells was significantly higher than normal in silicotic lung. Most MC contained bFGF (rho = 0.88, p < 0.001). The V(v) of collagen/reticulin fibers was increased in silicosis and correlated with the V(v) of bFGF(+) cells (rho = 0.81, p < 0.001). Immature silicotic nodules contained bFGF(+) MC throughout the loose array of collagen/reticulin fibers. In large, mature nodules, the density of collagen/reticulin fibers was higher, and bFGF(+) MC were found only in the nodule periphery. Because of this circumferential MC alignment in silicotic nodules, we observed a negative correlation between the V(v) of bFGF(+) MC and the density of collagen/reticulin fibers in silicotic nodules (rho = -0.80, p < 0.001) and between the V(v) of all other nodule-associated cells and the density of collagen/reticulin fibers in the hypocellular nodule centers (rho = -0.84, p < 0.001). We conclude that MC that produce bFGF may play an important role in the development of silicosis. PMID- 10852785 TI - Protein nitration, metabolites of reactive nitrogen species, and inflammation in lung allografts. AB - This study investigated nitration and chlorination of epithelial lining fluid (ELF) proteins in patients (n = 29) who had undergone lung allotransplantation. We assayed lung lavage nitrotyrosine (NT) and chlorotyrosine (CT) by HPLC. We measured NT, nitrate (NO(3)(-)), and nitrate (NO(2)(-)) in bronchoalveolar lavage fluid (BALF) and total nitrite (NO(2)(-) + NO(3)(-)) in serum of another group of lung transplant patients (n = 82). In the first group (n = 29), percent nitration of tyrosines (Tyr) (NT/total Tyr x 100) in BALF proteins was: patients, 0.01 (0.00-0.12)%; median (25th-75th% confidence interval), and control subjects 0.01 (0.00-0.02)%. CT (CT/ total Tyr x 100) occurred only in the patients' BALF: 0.01 (0. 00- 0.02)%. In the second group (n = 82), nitrotyrosine (NT) was detected by ELISA in the BALF of patients: 9 (0-41) pmol/mg pro and control subjects: 28 (26 33). Total nitrite (NO(2)(-) + NO(3)(-)) in BALF of the patients: 3.3 (1.9-5.1) microM significantly exceeded that in control subjects: 1.3 (0.8-1.3) microM; p = 0.0133. Serum nitrite also was significantly higher in patients: 37 (26-55) microM than control subjects: 19 (17-20) microM; p = 0.0037. Airway inflammation in transbronchial biopsies (B score) correlated with NT in BALF (p = 0.0369). Lung transplants have increased airway concentrations of reactive nitrogen species (RNS) metabolites. NT, a marker of peroxynitrite (ONOO(-)), is related to the degree of airway inflammation in lung transplants. PMID- 10852786 TI - Differential patterns of apoptosis in resolving and nonresolving bacterial pneumonia. AB - Infection with either Streptococcus sanguis or Streptococcus pneumoniae type 25 causes acute pneumonitis in rats. Pneumonia caused by S. sanguis resolves over the course of 8 d, whereas pneumonia caused by S. pneumoniae type 25 progresses to fibrosis. To examine the role of apoptosis in these models, we performed assays with the terminal deoxynucleotidyltransferase-uridine nucleotide end labeling technique on tissue sections from rat lungs at various times, and quantified the results with image analysis. Apoptosis was a feature of both the acute and resolving stages of pneumonia. The pattern and extent of apoptosis were similar in both models during the acute stage, and the number of apoptotic nuclei increased in both models through 4 d after infection. Although there were differences in the cellular pattern of apoptosis after 2 d and 4 d of infection, the extent of apoptosis was the same in both models. After 8 d, major differences were observed. In the resolving model, apoptosis was limited primarily to an abscess in the base of the lung. In the nonresolving model, apoptosis was persistent. We also found that cyclin-dependent kinase-5 expression is upregulated during apoptosis induced by bacterial infection. These data indicate that the location and timing of apoptosis may determine whether pneumonia resolves or progresses to fibrosis. PMID- 10852787 TI - Segmental bronchial provocation induces nasal inflammation in allergic rhinitis patients. AB - Allergic rhinitis and asthma often coexist and share a genetic background. Pathophysiologic connections between the nose and lungs are still not entirely understood. This study was undertaken to compare allergic inflammation and clinical findings in the upper and lower airways after segmental bronchial provocation (SBP) in nonasthmatic allergic rhinitis patients. Eight nonasthmatic, grass pollen-sensitive patients with allergic rhinitis and eight healthy controls were included. Bronchial biopsies and blood samples were taken before (T(0)) and 24 h (T(24)) after SBP. Nasal biopsies were obtained at T(0), 1 h after SBP (T(1)), and T(24). Immunohistochemical staining was performed for eosinophils (BMK13), interleukin (IL)-5, and eotaxin. The number of eosinophils increased in the challenged and unchallenged bronchial mucosa (p < 0.05) and in the blood (p = 0.03) of atopic subjects at T(24). We detected an increase of BMK13-positive and eotaxin-positive cells in the nasal lamina propria and enhanced expression of IL 5 in the nasal epithelium of atopic subjects only at T(24) (p < 0.05). SBP induced nasal and bronchial symptoms as well as reductions in pulmonary and nasal function in the allergic group. No significant changes could be observed in healthy controls. The study shows that SBP in nonasthmatic allergic rhinitis patients results in peripheral blood eosinophilia, and that SBP can induce allergic inflammation in the nose. PMID- 10852788 TI - Beneficial effect of lung preservation is related to ultrastructural integrity of tubular myelin after experimental ischemia and reperfusion. AB - Ischemia/reperfusion (I/R) injury results in the impairment of surfactant activity. The hypothesis that the differences in lung preservation quality obtained by EuroCollins (EC) and Celsior (CE) solutions were related to surfactant alterations was tested. To avoid extensive structural damage and edema formation, which can secondarily affect the surfactant system, lungs were stored for a short ischemic period (2 h at 10 degrees C) and reperfused (50 min) in an isolated perfused rat lung model after preservation with either potassium-reduced (40 mmol) EC40 or with CE. Using a modified stereological approach ultrastructure, total amount and distribution of phospholipid membranes composing tubular myelin (tm) and small (s) and large (l) unilameliar vesicles (ul) were investigated in the organ in lungs fixed by vascular perfusion either in situ (controls) or after I/R (n = 5 per group). The total amount of intraalveolar surfactant was increased after I/R. However, a significant amount (p = 0.008) of tm was displaced into the alveolar lumen and showed wider meshes of the tm lattices than did the controls (p = 0.023) where almost all tm was epithelial. In lungs preserved with EC40, epithelial tm was significantly reduced (p = 0.018), resulting in a higher ratio (p = 0.034) of surface-inactive small ul (0.05 to 0.3 microm) to surface-active epithelial tm. In the CE group approximately 50% of the total tm pool was epithelial. This was accompanied by higher parenchymal air space and improved functional parameters. Epithelial and endothelial cell specific immunostaining did not reveal any gross damage of the blood-gas barrier. In summary, improved lung function during reperfusion was associated with beneficial effects of lung preservation on tm integrity after I/R. These observations suggest that preservation solutions ameliorate events leading to surfactant disturbance even before extensive lung injury is manifested. PMID- 10852789 TI - Alterations in large and small proteoglycans in bleomycin-induced pulmonary fibrosis in rats. AB - In bleomycin (BM)-induced lung fibrosis, alterations have been shown in the expression and deposition of small proteoglycans (PGs). Less, however, is known about changes in large PGs. We investigated changes in large aggregating (versican [VS]), basement membrane (heparan sulfate proteoglycan [HSPG]), and small (biglycan and fibromodulin) PGs during the development of BM-induced pulmonary fibrosis. BM (1.5 U) was instilled intratracheally into male Sprague Dawley rats. Control rats received saline. At 7, 14, and 28 d after administration of BM, lungs were excised; one lung was fixed in formalin and 5 microm sections were cut and stained with hematoxylin-eosin. The other lung was used for PG extraction. PGs were extracted with guanidine and were separated through composite gel polyacrylamide gel electrophoresis (PAGE) (large PGs) and sodium dodecylsulfate-PAGE (small PGs). Gels were either stained or electrophoretically transferred and probed with antibodies to VS, HSPG, biglycan, and fibromodulin. Histologic samples showed prominent inflammation, with abundant proteinaceous material, most evident in the samples obtained at 7 and 14 d after administration of BM. By 28 d after BM, much of the inflammatory response had resolved, and heterogeneous distribution of fibrosis was observed. Immunoblots showed a relative abundance of VS at 7 and 14 d. Control lungs stained minimally for VS. Levels of HSPG, biglycan, and fibromodulin were increased maximally at 14 d after administration of BM. Immunocytochemistry showed intense immunostaining of biglycan and fibromodulin in the areas of injured lung tissue from rats 14 and 28 d after BM administration. Control lungs revealed minimal staining for small PGs. Our findings indicate that changes in all subclasses of PGs occur during the development of BM-induced pulmonary fibrosis. PMID- 10852790 TI - New aspects of degranulation and fates of airway mucosal eosinophils. PMID- 10852791 TI - Evidence of airway inflammation and remodeling in ski athletes with and without bronchial hyperresponsiveness to methacholine. AB - Asthma-like symptoms, methacholine hyperresponsiveness, and use of asthma medication are prevalent in elite cross-country skiers. We quantitated mucosal inflammatory cell infiltration and tenascin expression in the subepithelial basement membrane in endobronchial biopsy specimens of the proximal airways from 40 elite, competitive skiers (mean: 17.5; range: 16 to 20 yr) without a diagnosis of asthma, in 12 subjects with mild asthma, and in 12 healthy controls, through immunohistochemistry and indirect immunofluorescence, respectively. All of the subjects were nonsmokers. T-lymphocyte, macrophage, and eosinophil counts were, respectively, greater by 43-fold (p < 0.001), 26-fold (p < 0.001), and twofold (p < 0.001) in skiers, and by 70-fold (p < 0.001), 63-fold (p < 0.001), and eightfold (p < 0.001) in asthmatic subjects than in controls. In skiers, neutrophil counts were more than twofold greater than in asthmatic subjects, and mast cell counts were not significantly different than in controls. Tenascin expression (as measured through the thickness of the tenascin-specific immunoreactivity band in the basement membrane) was increased in skiers (median: 6.7 microm; interquartile range [IQR]: 5.3 to 8.5 microm, p < 0.001) and asthmatic subjects (mean: 8.8 microm; IQR: 7.2 to 10.8 microm, p < 0. 001) compared with controls (mean: 0.8 microm; IQR: 0 to 3.1 microm) and did not correlate with inflammatory cell counts. Inflammatory changes were present irrespective of asthmalike symptoms, hyperresponsiveness, or atopy. Prolonged repeated exposure of the airways to inadequately conditioned air may induce inflammation and remodeling in competitive skiers. PMID- 10852792 TI - Predictors of late asthmatic response. Logistic regression and classification tree analyses. AB - To identify predictors of the late asthmatic response (LAR), we reviewed data from 60 asthmatic subjects who had undergone allergen challenge over the past 5 yr (33 females, age 31.4 +/- 6.7 yr [mean +/- SD], FEV(1) 90% +/- 14% predicted). Variables considered likely predictors of LAR included baseline FEV(1), PC(20) methacholine (PC(20)), sputum eosinophil percent, and the decrease in FEV(1) within 20 min of allergen challenge. A LAR (FEV(1) >/= 15% fall between 3 and 7 h after challenge) was documented in 57% of subjects. A variety of logistic regression methods revealed a significant inverse association between LAR and PC(20) (odds ratio [OR] = 0.14 [95% CI = 0.03-0.66]) and a positive association between LAR and the decrease in FEV(1) at 20 min (OR = 1.18 [1.04 -1.33]). Classification tree analysis revealed that a threshold of 0.25 mg/ml for PC(20) was most predictive of LAR; LAR developed in 87% of those with PC(20) 0.25 mg/ml (n = 37). Notably, in subjects with PC(20) > 0.25 mg/ml, the incidence of LAR increased from 38% to 57% if the allergen-induced decline in FEV(1) at 20 min was >/= 27%. Surprisingly, baseline FEV(1) and percent eosinophils in induced sputum were not significantly associated with LAR. We conclude that a threshold value of 0.25 mg/ml for PC(20) methacholine is a good predictor of LAR. Measuring the PC(20) methacholine may be useful as a screening method to improve the efficiency of identifying asthmatic subjects with a LAR. PMID- 10852793 TI - Nedocromil sodium inhibits responsiveness to inhaled mannitol in asthmatic subjects. AB - Nedocromil sodium inhibits the response to exercise-induced asthma (EIA). Mannitol given as a powder by inhalation is an osmotic stimulus that identifies EIA. We studied the acute effect of nedocromil on airway responsiveness to mannitol in 24 asthmatic subjects. After a control day, nedocromil (8 mg) or its placebo was administered randomized, double blind, 10 min before a challenge with progressively increasing doses of mannitol. Nedocromil inhibited the response to mannitol and there was a significant increase in the dose of mannitol required to cause a 15% reduction in FEV(1) (PD(15)) after nedocromil 409 (316,503) mg compared with placebo 156 (106,229) mg (p < 0.001). In the presence of nedocromil 12 subjects no longer recorded a 15% decrease in FEV(1) in response to mannitol. The remaining 12 required a significantly greater dose of mannitol to achieve a 15% decrease in FEV(1) after nedocromil. Following nedocromil, a plateau in responsiveness to mannitol was observed in 14 subjects. Nedocromil significantly inhibits the responsiveness to inhaled mannitol in asthmatic subjects. PMID- 10852794 TI - Respiratory control and respiratory sensation in a patient with a ganglioglioma within the dorsocaudal brain stem. AB - We encountered a young woman with severe central sleep apnea caused by a medullary glioma located slightly dorsal to and to the right of the midline, a region not generally associated with CO(2) chemosensitivity. The patient had normal spirometric readings, lung volumes, diffusing capacity, maximal inspiratory pressure, and alveolar-arterial oxygen difference. While awake, she displayed marked irregularity in her breathing pattern; her end-tidal CO(2) (FET(CO(2))) ranged from 5.3 to 10.9%. During voluntary hyperpnea, she could quickly reduce her FET(CO(2)) to 4.2%, but her PCO(2) did not change after administration of acetazolamide or progesterone. Like patients with congenital central hypoventilation syndrome (CCHS), our patient had a relatively intact ventilatory response to exercise; her PCO(2) was high at the start of exercise and increased slightly thereafter. In contrast to CCHS patients, however, our patient had an intact hypoxic ventilatory response (DeltaVE/ DeltaSa(O(2)) = 0.37 L/min/Sa(O(2))). In further contrast to CCHS patients, our patient had a very short breathholding time and described a sensation of air hunger as the factor limiting her breathholding ability. Her heart rate and blood pressure responses to the Valsalva maneuver were normal. PMID- 10852795 TI - Unusual respiratory response to oxygen in an infant with repetitive cyanotic episodes. AB - High inspired oxygen concentrations have recently been recommended to control Cheyne-Stokes respiration in adults, with the intention of averting periodic apnea and its attendant arterial desaturation. We report a case study on an infant presenting with recurrent apnea and cyanosis in which oxygen treatment led to a gross form of respiratory instability we call episodic breathing, in which a breathing phase of 60 to 90 s alternated with an apnea lasting up to 60 s. When oxygen was discontinued, a profound arterial desaturation developed before breathing recommenced and restored oxygen levels. We propose that episodic breathing is an unusual respiratory pattern that involves the central chemoreceptors and results from the ventilatory threshold (the central PCO(2) at which breathing starts) lying considerably above the apneic threshold (the central PCO(2 )at which breathing stops). This feature predisposes to lengthy periods of hyperpnea alternating with lengthy periods of apnea. We suggest that when the case infant returned to air during episodic breathing, termination of apnea was entirely dependent upon carotid body activity, which reached a sufficient level to restart breathing only when arterial desaturation was severe. We conclude that oxygen therapy involves potential risks when employed to treat respiratory disorders involving unstable breathing patterns in the infant. PMID- 10852796 TI - Thermally induced asthma and airway drying. PMID- 10852797 TI - Exhaled nitric oxide does not provide a marker of vascular endothelial function in healthy humans. PMID- 10852798 TI - Systemic effects of inhaled fluticasone propionate and budesonide in adult patients with asthma. PMID- 10852799 TI - Immunolocalization of the Toxin Latrunculin B within the Red Sea Sponge Negombata magnifica (Demospongiae, Latrunculiidae). AB - The location of latrunculin B, the major toxin of the Red Sea sponge Negombata magnifica, was revealed using specific antibodies. Antibodies from rabbits immunized with a conjugate of latrunculin B with keyhole limpet hemocyanin (KLH) were purified over a latrunculin B-Sepharose affinity column. Analysis of immunohistochemical and immunogold-stained sponge sections, using light and transmission electron microscopy, revealed latrunculin B labeling mostly beneath the sponge cortex at the border between the external (ectosome) and internal (endosome) layers (ectosome-endosome border). The endosome was less labeled than the border. Immunogold localization revealed latrunculin B in the sponge cells but not in its prokaryotic symbionts. Archeocytes and choanocytes were significantly more labeled than other cells. The antibodies primarily labeled membrane-limited vacuoles within archeocytes and choanocytes that are perhaps latrunculin B secretory or storage vesicles. Peripheral latrunculin B may have a role in defense against external epibionts, predators, and competitors. PMID- 10852800 TI - Analysis of initial fouling process in coastal environment: effects of settlement, attachment, and metamorphosis promoters. AB - Effects of lumichrome, L-tryptophan, and curcumin on fouling organisms were examined on panels immersed in a near-shore aquatic environment. These products showed effective action in aquariums in preliminary screening tests for promoting different steps of fouling. Lumichrome showed metamorphosis-inducing activity for ascidian larvae (Halocynthia), L-tryptophan was a settlement-inducer for larvae of barnacles (Balanus), and curcumin showed attachment-promoting activity on the blue mussel (Mytilus). In order to establish that these tests are helpful in screening actual fouling or antifouling compounds, we examined the action of these three compounds in a coastal environment by following the first steps of biofouling, that is, by studying the quantity of chlorophyll, the number of bacterial cells, and the larvae settled. These experiments on the seashore indicated that these compounds did not act as promoters for the target organisms; however, they did show promoting effects on some nontarget organisms. PMID- 10852801 TI - Identification of Hox Gene Sequences in the Sea Cucumber Holothuria glaberrima Selenka (Holothuroidea: Echinodermata). AB - The Echinodermata is a unique animal group forming an early branch in the deuterostomes phylogenetic tree. In echinoids and asteroids a single Hox cluster with nine cognates of the vertebrate Hox paralogous groups has been reported, but no data are available from other echinoderm classes. We report here nine Hox-type sequences from the sea cucumber Holothuria glaberrima, a member of the class Holothuroidea. Partial homeodomain sequences were amplified by polymerase chain reaction from genomic DNA and from a regenerating gastrointestinal tract complementary DNA library. Sequence analyses suggest that the holothuroid cluster has at least three genes of the anterior, one of the medial, and five of the posterior groups. This is the first evidence of five posterior sequences in echinoderms. PMID- 10852802 TI - Molecular Cloning and Novel Repeated Sequences of a C-type Lysozyme Gene in Japanese Flounder (Paralichthys olivaceus). AB - A clone of a DNA fragment of approximately 10 kb that included a c-type lysozyme gene of Japanese flounder (Paralichthys olivaceus) was isolated from a Japanese flounder genomic DNA library. This clone was subcloned, and the nucleotide sequence of the c-type lysozyme gene was determined. The flounder c-type lysozyme gene, which consisted of 3,617 bp, was found to be composed of four exons and three introns. The transcription start site was determined to be 44 bp upstream of the ATG codon by primer extension analysis. The number of exons and introns and the positions of the catalytic residues of the flounder gene were the same as those of the human and chicken c-type lysozyme genes. However, introns 1 and 3 of the flounder gene were shorter, and intron 2 was longer, than those in the other reported vertebrates genes. There were no Alu repeated sequences in the flounder lysozyme gene, as determined for the human lysozyme gene, but 45 repeated sequences were found in the second intron of the flounder lysozyme gene. We suggest that the length of intron 2 in the Japanese flounder c-type lysozyme gene depends on the presence of this repeated sequence. Furthermore, Southern blot analysis revealed that the c-type lysozyme gene was probably a single copy in the genomic DNA of homo-cloned Japanese flounder. PMID- 10852803 TI - Interspecific Utility of Microsatellites in Fish: A Case Study of (CT)(n) and (GT)(n) Markers in the Shanny Lipophrys pholis (Pisces: Blenniidae) and Their Use in Other Blennioidei. AB - We report the development of new microsatellite markers that can be used for population analyses in the shanny Lipophrys pholis. The procedure involved the construction of a microsatellite-enriched genomic bank. Five (GT)(n) and (CT)(n) microsatellites have been characterized, four of which are polymorphic. The analysis of one population allowed us to verify their usefulness as markers in population studies. Moreover, interspecific amplifications have been performed using primers defined in other species to amplify Lipophrys pholis, or using the primers defined in Lipophrys pholis to amplify other species. We use these results to discuss the hypothesis that microsatellites are highly conserved in fish. PMID- 10852804 TI - Stable Expression of a Foreign Gene, Delivered by Gene Gun, in the Muscle of Rainbow Trout Oncorhynchus mykiss. AB - We report the efficient delivery of a foreign gene into muscle of rainbow trout Oncorhynchus mykiss with a gene gun. The foreign gene was a reporter gene, chloramphenicol acetyltransferase (CAT). Two CAT-containing plasmids were used: pCMV-CAT, which contains cytomegalovirus immediate early promoter, and pSV2-CAT, which contains the simian virus 40 early promoter. All plasmids were introduced by particle bombardment using a gene gun. During the 90-day sampling period following bombardment, CAT was strongly and stably expressed in the muscle of all the fish bombarded with pCMV-CAT and pSV2-CAT. No CAT expression was detected in the blood samples until 90 days after introduction, when it was found in only one fish from the pCMV-CAT group and one from the pSV2-CAT group. The stable and long term expression of plasmid DNA in muscle makes muscle an attractive target tissue for the introduction of viral DNA for the purpose of DNA vaccination. PMID- 10852805 TI - Purification and Some Properties of Two Carboxypeptidases from the Hepatopancreas of the Crab Paralithodes camtschatica. AB - High activity of carboxypeptidases was detected in the hepatopancreas of the crab Paralithodes camtschatica, while aminopeptidase activity in this tissue was low. Two crab carboxypeptidases were purified by chromatography on DEAE-cellulose, phenyl-Sepharose, and Sephadex G-75 to homogeneity. The molecular weight values of carboxypeptidases I and II were 40,000 and 37,000, respectively. The isoelectric point value for both carboxypeptidases was 4.5. The crab carboxypeptidases were activated by NaCl, with maximal activity of carboxypeptidases I and II at 1.0 M and 0.6 M NaCl, respectively. Using 19 N blocked dipeptides with the general structures Bz-Gly-X and Z-Gly-X, broad substrate specificity of the purified enzymes was demonstrated. Under optimal conditions the values of K(m) and k(cat) for the hydrolysis of Bz-Gly-l-Phe, Bz Gly-l-Arg, and Bz-Gly-l-Lys by crab carboxypeptidases were determined. Inhibition data led to classification of the crab enzymes as metallopeptidases. Both carboxypeptidases were stable under neutral and mildly alkaline conditions. In addition, the presence of 1 M NaCl decreased the thermostability of the crab carboxypeptidases. PMID- 10852806 TI - Molecular analysis of ribosomal RNA gene of red tide algae obtained from the seto inland sea. AB - Eleven clones from five species of the planktonic microalgae, (Chattonella antiqua, Chattonella marina, Heterosigma akashiwo, Alexandrium catenella, and Scrippsiella trochoidea), which were collected from the Seto Inland Sea in Japan and from Thailand, were subjected to nucleotide sequence analysis of the D1/D2 domain of the large subunit (LSU) of their ribosomal RNA genes. After amplification by polymerase chain reaction using degenerated primers, whole nucleotide sequences for the D1/D2 domains of the LSU rRNA gene of 11 microalgae were analyzed. Phylogenic tree analysis using these nucleotide sequences showed each species located in a cluster corresponding to its morphological classification. The nucleotide sequence data for Chattonella spp. suggest that multiple clones of both Chattonella antiqua and Chattonella marina are present in the Seto Inland Sea and that red tide blooms of Chattonella spp. in different years may have contained different clones. PMID- 10852807 TI - Mitochondrial DNA variation, phylogenetic relationships, and evolution of four mediterranean genera of soles (soleidae, pleuronectiformes). AB - To increase knowledge about the systematics and evolution of Mediterranean soles, we assessed mitochondrial DNA variation, molecular phylogeny, and evolution in eight species from the genera Solea, Microchirus, Monochirus, and Buglossidium by large ribosomal subunit (16S) and cytochrome b (cytb) sequence analysis. Relevant molecular features are the great variation of base composition among species at the third codon in cytb and the heterogeneity of the nucleotide substitution rate. Phylogenies recovered using 16S nucleotide and cytb amino acid sequences agree with those based on morphology in assessing monophyly of Solea species and ancestry of Buglossidium luteum, but they are against the intergeneric differentiation of Microchirus and Monochirus. Conversely, phylogenetic trees based on cytb nucleotide sequences yielded relationships among taxa regardless of their evolutionary histories. The incongruities between morphological and molecular issues suggest the need for reassessing the systematic value of some morphological characters. Approximate estimates of the divergence time of Mediterranean soleid lineages range from 40 to 13 Mya (Oligocene-Miocene), indicating an ancient origin for the group. PMID- 10852808 TI - Interaction of Cytotoxic Bicyclic Peptides, Theonellamides A and F, with Glutamate Dehydrogenase and 17beta-Hydroxysteroid Dehydrogenase IV. AB - Theonellamide A, a bicyclic peptide isolated from a Theonella sponge, was fixed on hydrazide-containing gel beads and screened for its binding proteins from rabbit liver tissues. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that two major proteins of 80 kDa and 55 kDa interacted with theonellamide A. The interaction between theonellamide A and two proteins was confirmed by competition experiments in which these two proteins failed to bind to theonellamide A-conjugated gel beads in the presence of theonellamide A or F. Amino-terminal amino acid sequence analysis of peptide fragments derived from the binding proteins by lysylendopeptidase digestion demonstrated that the 80-kDa and 55-kDa proteins were 17beta-hydroxysteroid dehydrogenase IV and glutamate dehydrogenase, respectively. In an in vitro assay system, amination of alpha-ketoglutarate by glutamate dehydrogenase was activated with theonellamide F, although this effect was weaker than that with adenosine diphosphate, a well known activator. PMID- 10852809 TI - Gene-Gun-Mediated Transfer of Reporter Genes to Somatic Zebrafish (Danio rerio) Tissues. AB - We describe the use of gene-gun-mediated transfer of luciferase and green fluorescent protein (GFP) reporter genes in zebrafish (Danio rerio). Optimization of DNA transfer parameters indicated highest overall luciferase expression in epidermis and dermis using 1-MUm microcarriers and 1 MUg of pCMVL plasmid DNA at a delivery pressure of 200 psi. Time course studies revealed luciferase activity peaking at 18 hours and decreasing to 30% of the maximum at day 8 after DNA transfer. Onset of reporter gene (GFP) expression was detected at 13 minutes after DNA delivery, and by 65 minutes approximately 100% of the cells in the target area exhibited GFP expression. No germline association or integration events were detected in a screen of approximately 250,000 zebrafish sperm cells by fluorescence in situ hybridization at 15 or 30 days after delivery of 1 MUg of pCMVL DNA, suggesting incidental male germline integration should not be considered as a risk factor when using the biolistic DNA delivery parameters and target tissues described. PMID- 10852810 TI - AFLP Analysis Confirms Exclusive Maternal Genomic Contribution of Meiogynogenetic Sea Bass (Dicentrarchus labrax L.). AB - Meiogynogenesis was induced in the European sea bass Dicentrarchus labrax L. by fertilizing eggs with UV-irradiated sperm followed by inhibition of the second meiotic division by a cold shock. Putative gynogenetic progeny derived from three groups of breeders were analyzed for maternal inheritance using amplified fragment length polymorphism (AFLP) markers. Discrimination of fingerprints was based on male-specific bands, which were absent in females. Four of 64 MseI/EcoRI primer pairs used to analyze parental polymerase chain reaction products were selected to screen progeny for paternal AFLP markers in each group. Four to 11 diagnostic bands per fish confirmed the gynogenetic origin of the progeny. AFLP analysis determined that 89.5%, 100%, and 100% of the sea bass from groups 1, 2, and 3, respectively, were gynogenetic. Our results show that AFLP analysis is suitable for verification of gynogenesis in fish. PMID- 10852811 TI - Cases of cat-associated human plague in the Western US, 1977-1998. AB - Exposure to cats infected with Yersinia pestis is a recently recognized risk for human plague in the US. Twenty-three cases of cat-associated human plague (5 of which were fatal) occurred in 8 western states from 1977 through 1998, which represent 7.7% of the total 297 cases reported in that period. Bites, scratches, or other contact with infectious materials while handling infected cats resulted in 17 cases of bubonic plague, 1 case of primary septicemic plague, and 5 cases of primary pneumonic plague. The 5 fatal cases were associated with misdiagnosis or delays in seeking treatment, which resulted in overwhelming infection and various manifestations of the systemic inflammatory response syndrome. Unlike infections acquired by flea bites, the occurrence of cat-associated human plague did not increase significantly during summer months. Plague epizootics in rodents also were observed less frequently at exposure sites for cases of cat-associated human plague than at exposure sites for other cases. The risk of cat-associated human plague is likely to increase as residential development continues in areas where plague foci exist in the western US. Enhanced awareness is needed for prompt diagnosis and treatment. PMID- 10852812 TI - Influence of the normal menstrual cycle on vaginal tissue, discharge, and microflora. AB - The objective of this study was to examine genital tissue, vaginal fluid, and vaginal microbial flora at 3 phases of the menstrual cycle in asymptomatic women. Vaginal examinations were performed 3 times in 74 women: at the menstrual phase (days 1-5), the preovulatory phase (days 7-12), and the postovulatory phase (days 19-24). Flora of 50 women without bacterial vaginosis (BV) was analyzed separately from flora of 24 women with BV. The volume of vaginal discharge increased and the amount of cervical mucus decreased over the menstrual cycle. Among subjects without BV, the rate of recovery of any Lactobacillus changed little (range, 82% to 98%; P = .2); however, a small increase occurred in the rate of recovery of heavy (3+ to 4+ semiquantitative) growth of Lactobacillus over the menstrual cycle (P = .04). A linear decrease occurred in the rate of recovery of heavy growth of any non-Lactobacillus species, from 72% at days 1-5 to 40% at days 19-24 (P = .002). A linear decrease also occurred in the rate of recovery of Prevotella species, from 56% on days 1-5 to 28% on days 19-24 (P =. 007), while a small linear increase occurred in the rate of recovery of Bacteroides fragilis (P=.05). Among subjects with BV, the only significant change was an increase in the rate of recovery of Lactobacillus, from 33% at days 1-5 to 54% at days 19-24 (P = .008). Among all subjects, the rate of recovery of heavy growth of Lactobacillus increased over the menstrual cycle and, in contrast, the concentration of non-Lactobacillus species tended to be higher at menses, which is evidence that the vaginal flora becomes less stable at this time. PMID- 10852813 TI - Spectrin tethers and mesh in the biosynthetic pathway. AB - The paradox of how the Golgi and other organelles can sort a continuous flux of protein and lipid but maintain temporal and morphological stability remains unresolved. Recent discoveries highlight a role for the cytoskeleton in guiding the structure and dynamics of organelles. Perhaps one of the more striking, albeit less expected, of these discoveries is the recognition that a spectrin skeleton associates with many organelles and contributes to the maintenance of Golgi structure and the efficiency of protein trafficking in the early secretory pathway. Spectrin interacts directly with phosphoinositides and with membrane proteins. The small GTPase ARF, a key player in Golgi dynamics, regulates the assembly of the Golgi spectrin skeleton through its ability to control phosphoinositide levels in Golgi membranes, whereas adapter molecules such as ankyrin link spectrin to other membrane proteins. Direct interactions of spectrin with actin and centractin (ARP1) provide a link to dynein, myosin and presumably other motors involved with intracellular transport. Building on the recognized ability of spectrin to organize macromolecular complexes of membrane and cytosolic proteins into a multifaceted scaffold linked to filamentous structural elements (termed linked mosaics), recent evidence supports a similar role for spectrin in organelle function and the secretory pathway. Two working models accommodate much of the available data: the Golgi mesh hypothesis and the spectrin ankyrin adapter protein tethering system (SAATS) hypothesis. PMID- 10852814 TI - Emerging issues in receptor protein tyrosine phosphatase function: lifting fog or simply shifting? AB - Transmembrane (receptor) tyrosine phosphatases are intimately involved in responses to cell-cell and cell-matrix contact. Several important issues regarding the targets and regulation of this protein family are now emerging. For example, these phosphatases exhibit complex interactions with signaling pathways involving SRC family kinases, which result from their ability to control phosphorylation of both activating and inhibitory sites in these kinases and possibly also their substrates. Similarly, integrin signaling illustrates how phosphorylation of a single protein, or the activity of a pathway, can be controlled by multiple tyrosine phosphatases, attesting to the intricate integration of these enzymes in cellular regulation. Lastly, we are starting to appreciate the roles of intracellular topology, tyrosine phosphorylation and oligomerization among the many mechanisms regulating tyrosine phosphatase activity. PMID- 10852815 TI - Dietary fat and reduced levels of TGFbeta1 act synergistically to promote activation of the vascular endothelium and formation of lipid lesions. AB - Transforming growth factor-(beta) (TGF(beta)) has a wide range of activities on vascular cells and inflammatory cells, suggesting it may have different functions during various stages of atherogenesis. We report that mice heterozygous for the deletion of the tgfb1 gene (tgfb1(+/-) mice) have reduced levels of TGF(beta)1 in the artery wall until at least 8 weeks of age. On a normal mouse chow diet, the vascular endothelium of tgfb1(+/-) mice is indistinguishable from wild-type littermates, assessed by morphology and intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression. In contrast, levels of the smooth muscle isoforms of actin and myosin in medial smooth muscle cells of tgfb1(+/-) mice are significantly reduced. Following feeding a cholesterol-enriched diet for 12 weeks, high levels of ICAM-1 and VCAM-1 were detected in the vascular endothelial cells of tgfb1(+/-) mice, but not wild-type mice. Furthermore, marked deposition of lipid into the artery wall was only observed in the tgfb1(+/-) mice on the cholesterol-enriched diet. These vascular lipid lesions were accompanied by local invasion of macrophages. We conclude that deletion of a single allele of the tgfb1 gene results in a reduced level of TGFbeta1 antigen in the aorta together with reduced smooth muscle cell differentiation, whereas the addition of a high fat dietary challenge is required to activate the vascular endothelium and to promote the formation of fatty streaks resembling early atherosclerosis in humans. PMID- 10852816 TI - Human junction adhesion molecule regulates tight junction resealing in epithelia. AB - Epithelial cells form a highly selective barrier and line many organs. The epithelial barrier is maintained by closely apposed cell-cell contacts containing tight junctions, the regulation of which is incompletely understood. Here we report the cloning, tissue localization and evidence for a role in epithelial barrier regulation of an immunoglobulin superfamily member that likely represents the human homolog of murine junction adhesion molecule (JAM). Analysis of the primary structure of human JAM, cloned from T84 epithelial cells, predicts a transmembrane protein with an extracellular domain that contains two IgV loops. Monoclonal antibodies generated against the putative extracellular domain were reactive with a 35-39 kDa protein from both T84 epithelial cells and human neutrophils. By immunofluorescence, JAM mAbs labeled epithelial cells from intestine, lung, and kidney, prominently in the region of tight junctions (co localization with occludin) and also along lateral cell membranes below the tight junctions. Flow cytometric studies confirmed predominant JAM expression in epithelial cells but also revealed expression on endothelial and hematopoietic cells of all lineages. Functional studies demonstrated that JAM specific mAbs markedly inhibited transepithelial resistance recovery of T84 monolayers after disruption of intercellular junctions (including tight junctions) by transient calcium depletion. Morphologic analysis revealed that, after disassembly of cell cell junctions, anti-JAM inhibition of barrier function recovery correlated with a loss of both occludin and JAM, but not ZO-1, in reassembling tight junction structure. Reassembly of the major adherens junction component E-cadherin was not affected by JAM specific mAbs. Our findings suggest that JAM plays an important role in the regulation of tight junction assembly in epithelia. Furthermore, these JAM-mediated effects may occur by either direct, or indirect interactions with occludin. PMID- 10852817 TI - Contraction of collagen matrices mediated by alpha2beta1A and alpha(v)beta3 integrins. AB - The (beta)1-null fibroblastic cell line GD25 and its derivatives were studied to gain an understanding of the roles of (beta)1 and (beta)3 integrins in the initial (1-hour) contraction of collagen gels. Stable transfectants of GD25 cells expressing the (beta)1A splice variant of (beta)1 ((beta)1A-GD25) did not express (alpha)2(beta)1A and did not adhere to collagen. After transfection of (alpha)2 into (beta)1A-GD25 cells, the (alpha)2(beta)1A-GD25 transfectants contracted collagen gels in the presence of serum, whereas (beta)1A-GD25 cells did not. The GD25 parental cells, however, also contracted collagen gels. Collagen gel contraction by GD25 cells was blocked by antibodies to (alpha)v(beta)3 or a RGD containing peptide, indicating that (alpha)v(beta)3 is the integrin responsible for mediation of contraction by GD25 cells. Collagen gel contraction by (alpha)2(beta)1A-GD25 cells was not inhibited by antibodies to (alpha)v(beta)3 or RGD-containing peptide, but was inhibited by anti-(alpha)2 antibody. Flow cytometry demonstrated negligible expression of (alpha)v(beta)3 by (beta)1A-GD25 and (alpha)2(beta)1A-GD25 cells when compared to GD25 cells. Platelet derived growth factor (PDGF) and sphingosine-1-phosphate (S1P) enabled gel contraction by (alpha)2(beta)1A-GD25 and GD25 cells, respectively, in the absence of serum. PDGF stimulated contraction by (alpha)2(beta)1A-GD25 cells was attenuated in the presence of inhibitors of phosphatidylinositol-3-kinase whereas such inhibitors had no effect on S1P-stimulated contraction by GD25 cells. These experiments using the (beta)1-null GD25 cells and (beta)1A and (alpha)2(beta)1A transfectants demonstrate that (alpha)2(beta)1A and (alpha)v(beta)3 independently mediate collagen gel contraction and are regulated by different serum factors and signaling pathways. PMID- 10852818 TI - Galectin-8 binding to integrins inhibits cell adhesion and induces apoptosis. AB - The interaction of cells with the extracellular matrix regulates cell adhesion, motility, growth, survival and differentiation through integrin-mediated signal transduction. Here we demonstrate that galectin-8, a secreted mammalian (beta) galactoside binding protein, inhibits adhesion of human carcinoma (1299) cells to plates coated with integrin ligands, and induces cell apoptosis. Pretreatment of the cells with Mn(2+), which increases the affinity of integrins for their ligands, abolished the inhibitory effects of galectin-8. The inhibitory effects of galectin-8 were specific and were not mimicked by plant lectins or other galectins (galectin-1 and galectin-3). In accordance with its anti-adhesive effects, transfection of galectin-8 cDNA into 1299 cells significantly reduced (by 75%) colony formation, when compared to the number of colonies formed by cells transfected with an empty vector. Affinity chromatography over immobilized galectin-8 indicated that few membrane proteins interacted with galectin-8 in a sugar-dependent manner. Microsequencing and western immunoblotting revealed that (alpha)(3)(beta)(1 )integrin derived from 1299 as well as other cells (e.g. HeLa and human endothelial cells) is a major galectin-8 binding-protein. Furthermore, immunoprecipitation and immunohistochemical studies suggested that endogenous galectin-8, secreted from 1299 cells, forms complexes with (alpha)(3)(beta)(1) integrins expressed on the surface of 1299 cells. Galectin-8 also interacts with other members of the integrin family, like (alpha)(6)(beta)(1 )integrins. In contrast, galectin-8 only minimally interacts with (alpha)(4 )or (beta)(3 )integrins. We propose that galectin-8 is an integrin binding-protein that interacts to a different extent with several, but not all members of the integrin family. Binding of galectin-8 modulates integrin interactions with the extracellular matrix and thus regulates cell adhesion and cell survival. PMID- 10852819 TI - In mouse myoblasts nuclear prosomes are associated with the nuclear matrix and accumulate preferentially in the perinucleolar areas. AB - Prosomes are the core of 26S proteasomes, although they were originally observed as 20S particles associated with cytoplasmic mRNPs. Here we show for the first time that prosomes are also genuine constituents of the nuclear matrix, chromatin and the nuclear RNP networks. Using mouse myoblasts we tested three monoclonal antibodies recognising the prosomal subunits p23K, p27K and p30K, and found that the corresponding prosome subclasses are characterised by a variable distribution pattern within the nuclei. Their presence on the nuclear matrix, and most abundantly in the perinucleolar area, is of particular importance. When myoblasts fuse into myotubes, the distribution pattern of certain types of prosomes on the nuclear matrix changes drastically. Surprisingly, DNA strongly interferes with the detection of prosomal antigens by immunofluorescence methods, whereas RNA, histones and other proteins soluble in 2 M NaCl have no such effect. This 'masking' of prosomes can be completely overcome by extensive or even mild digestion with DNase I or restriction enzymes. Many nuclear prosomes can be solubilized by combined treatment with 0.5% Triton X-100 and 2 M NaCl, and others can be released by digestion of DNA and/or RNA, and about 10-20% of nuclear prosomes remain tightly bound to the protein-based nuclear matrix. PMID- 10852820 TI - Toxoplasma gondii catalase: are there peroxisomes in toxoplasma? AB - The intracellular protozoan parasite Toxoplasma gondii, like all members of the phylum Apicomplexa, is known to possess many organelles: in addition to mitochondria and the compartments of the secretory pathway, there is a reduced chloroplast (the apicoplast) and the phylum-specific components of the apical complex: dense granules, micronemes and rhoptries. Conspicuously missing so far are microbodies, organelles that can be found in nearly all eukaryotic organisms. Microbodies show a large variation with regard to their size, number and contents, depending on the organism and cell type. One marker enzyme of this single membrane-bound organelle is catalase, which is responsible for the degradation of hydrogen peroxide to water and oxygen. The EST project in T. gondii revealed the existence of two overlapping clones which showed similarity with catalase, and these were used to clone the corresponding gene. The predicted sequence of T. gondii catalase has -AKM at the C terminus, which falls within the consensus of the PTS1 peroxisomal targeting signal. Southern blot analysis confirmed the presence of a single copy gene. Northern and western blot analyses showed that the catalase gene is transcribed and translated. Immunofluorescence assays using an antibody raised against a catalase peptide identified a distinct structure towards the apical end, but other catalase-specific antibodies failed to confirm this localisation. Cell fractionations indicated that the majority of the enzyme was in the cytosol. The fusion of the C-terminal twelve amino acids, including AKM, or the canonical peroxisomal targeting signal, -SKL, to GFP resulted in predominantly cytosolic localization in T. gondii. There was therefore no evidence for membrane-bound peroxisomes in Toxoplasma. PMID- 10852821 TI - Fission yeast Rng3p: an UCS-domain protein that mediates myosin II assembly during cytokinesis. AB - Cell division in many eukaryotes, including the fission yeast Schizosaccharomyces pombe, utilizes a contractile actomyosin ring. In S. pombe, the actomyosin ring is assembled at the medial cortex upon entry into mitosis and constricts at the end of anaphase to guide the centripetal deposition of the septum. Despite identification of several structural components essential for actomyosin ring assembly, the interdependencies between these gene-products in the process of ring assembly are unknown. This study investigates the role of Rng3p, a member of the UCS-domain containing protein family (Unc-45p, Cro1p, She4p), in actomyosin ring assembly. Null mutants in rng3 resemble deletion mutants in the type II myosin heavy chain (myo2) and rng3(ts) mutants show strong negative interactions with the myo2-E1 mutant, suggesting that Rng3p is involved in modulating aspects of type II myosin function. Interestingly, a green fluorescent protein (GFP) tagged Rng3p fusion is detected at the division site in the myo2-E1 mutant, but not in other myo2-alleles, wild-type cells or in 18 other cytokinesis mutants. Assembly and maintenance of Rng3p at the division site in the myo2-E1 mutant requires F-actin. Rng3p is also required for the proper assembly of Myo2p and F actin into a functional actomyosin ring but is not necessary for their accumulation at the division site. We conclude that Rng3p is a novel component of the F-actin cytoskeleton essential for a late step in actomyosin ring assembly and that it might monitor some aspect of type II myosin assembly during actomyosin ring construction. PMID- 10852822 TI - Misdirected vimentin messenger RNA alters cell morphology and motility. AB - Localized messenger RNAs were first observed as embryonic determinants that altered development when mislocalized. In recent years localized mRNAs have been found for several cytoskeletal proteins, including actin, vimentin and several microtubule associated proteins. We sought to determine whether redirecting mRNA for a cytoskeletal protein to an inappropriate address would alter cellular phenotypes. To do so we generated vimentin mRNAs with a myc epitope tag and the (beta)-actin 3' untranslated region (3' UTR) as a localization signal. When misdirected vimentin mRNAs are expressed in either fibroblasts or SW13 cells, cells develop numerous, extremely long processes; these cells also move more slowly to enter a wound of the monolayer. In situ hybridization revealed that the misdirected mRNA was often localized in the processes, in contrast to endogenous vimentin mRNA. The processes usually contained actin distal to the transgenic vimentin and microtubules proximal to it. SW13 cells lacking vimentin produced fewer and shorter processes, suggesting a dominant negative effect that involves recruitment of endogenous vimentin. Control experiments that transfected in constructs expressing tagged, correctly localized vimentin, or (beta) galactosidase that localized through the (beta)-actin 3' UTR, indicate that neither the shape nor the motility changes are solely due to the level of vimentin expression in the cell. This is direct evidence that the site of expression for at least one cytoskeletal mRNA alters the phenotype of the cell in which it is expressed. Messenger RNA localization is proving to be as essential for the normal maintenance of somatic cell phenotypes as embryonic determinants are for embryogenesis. PMID- 10852823 TI - A role for TGFbeta(1) in osteoclast differentiation and survival. AB - Recently, tumour necrosis factor-related activation-induced cytokine (TRANCE) was shown to be necessary for osteoclast formation. We now report that TGF(beta), a cytokine enriched in bone matrix, is also required. TGF(beta) not only powerfully synergized with TRANCE for induction of osteoclast-like cells (OCL) from bone marrow precursors and monocytes, but OCL formation was abolished by recombinant soluble TGF(beta) receptor II (TGF(beta)sRII). Preincubation in TGF(beta) was as effective as simultaneous incubation with TRANCE. TGF(beta)-preincubation enhanced OCL formation at least partly by preventing the development of resistance to OCL-induction that otherwise occurs when precursors are incubated in M-CSF. OCL formed in TRANCE also showed more rapid apoptosis than OCL in TRANCE plus TGF(beta). Like TGF(beta), incubation on bone matrix prolonged and enhanced the sensitivity of precursors to OCL-induction by TRANCE, and this was reversed by TGF(beta)sRII. Taken together, this data is compelling evidence for a model in which TGF(beta) in matrix or released from bone-lining or other cells maintains and enhances the osteoclast-forming potential of precursors as they migrate towards sites of cell-bound TRANCE. Thus, the specific circumstances necessary for osteoclast formation and survival are TRANCE expression on osteoblastic cells and TGF(beta) in bone. PMID- 10852824 TI - Impaired wound healing in embryonic and adult mice lacking vimentin. AB - It is generally assumed that the vimentin intermediate filament network present in most mesenchymally-derived cells is in part responsible for the strength and integrity of these cells, and necessary for any tissue movements that require the generation of significant tractional forces. Surprisingly, we have shown that transgenic KO mice deficient for vimentin are apparently able to undergo embryonic development absolutely normally and go onto develop into adulthood and breed without showing any obvious phenotype. However, fibroblasts derived from these mice are mechanically weak and severely disabled in their capacity to migrate and to contract a 3-D collagen network. To assess whether these functions are necessary for more challenging tissue movements such as those driving in vivo tissue repair processes, we have analysed wound healing ability in wild-type versus vimentin-deficient embryos and adult mice. Wounds in vimentin-deficient adult animals showed delayed migration of fibroblasts into the wound site and subsequently retarded contraction that correlated with a delayed appearance of myofibroblasts at the wound site. Wounds made to vimentin-deficient embryos also failed to heal during the 24 hour culture period it takes for wild-type embryos to fully heal an equivalent wound. By DiI marking the wound mesenchyme and following its fate during the healing process we showed that this impaired healing is almost entirely due to a failure of mesenchymal contraction at the embryonic wound site. These observations reveal an in vivo phenotype for the vimentin-deficient mouse, and challenge the dogma that key morphogenetic events occurring during development require generation of significant tractional forces by mesenchymal cells. PMID- 10852825 TI - Selective recruitment of arrestin-3 to clathrin coated pits upon stimulation of G protein-coupled receptors. AB - Non-visual arrestins (arrestin-2 and arrestin-3) play critical roles in the desensitization and internalization of many G protein-coupled receptors. In vitro experiments have shown that both non-visual arrestins bind with high and approximately comparable affinities to activated, phosphorylated forms of receptors. They also exhibit high affinity binding, again of comparable magnitude, to clathrin. Further, agonist-promoted internalization of many receptors has been found to be stimulated by exogenous over-expression of either arrestin2 or arrestin3. The existence of multiple arrestins raises the question whether stimulated receptors are selective for a specific endogenous arrestin under more physiological conditions. Here we address this question in RBL-2H3 cells, a cell line that expresses comparable levels of endogenous arrestin-2 and arrestin-3. When (beta)(2)-adrenergic receptors are stably expressed in these cells the receptors internalize efficiently following agonist stimulation. However, by immunofluorescence microscopy we determine that only arrestin-3, but not arrestin-2, is rapidly recruited to clathrin coated pits upon receptor stimulation. Similarly, in RBL-2H3 cells that stably express physiological levels of m1AChR, the addition of carbachol selectively induces the localization of arrestin-3, but not arrestin-2, to coated pits. Thus, this work demonstrates coupling of G protein-coupled receptors to a specific non-visual arrestin in an in vivo setting. PMID- 10852826 TI - Interaction of plakophilins with desmoplakin and intermediate filament proteins: an in vitro analysis. AB - Plakophilin 1 and 2 (PKP1, PKP2) are members of the arm-repeat protein family. They are both constitutively expressed in most vertebrate cells, in two splice forms named a and b, and display a remarkable dual location: they occur in the nuclei of cells and, in epithelial cells, at the plasma membrane within the desmosomal plaques. We have shown by solid phase-binding assays that both PKP1a and PKP2a bind to intermediate filament (IF) proteins, in particular to cytokeratins (CKs) from epidermal as well as simple epithelial cells and, to some extent, to vimentin. In line with this we show that recombinant PKP1a binds strongly to IFs assembled in vitro from CKs 8/18, 5/14, vimentin or desmin and integrates them into thick (up to 120 nm in diameter) IF bundles extending for several microm. The basic amino-terminal, non-arm-repeat domain of PKP1a is necessary and sufficient for this specific interaction as shown by blot overlay and centrifugation experiments. In particular, the binding of PKP1a to IF proteins is saturable at an approximately equimolar ratio. In extracts from HaCaT cells, distinct soluble complexes containing PKP1a and desmoplakin I (DPI) have been identified by co-immunoprecipitation and sucrose density fractionation. The significance of these interactions of PKP1a with IF proteins on the one hand and desmoplakin on the other is discussed in relation to the fact that PKP1a is not bound - and does not bind - to extended IFs in vivo. We postulate that (1) effective cellular regulatory mechanisms exist that prevent plakophilins from unscheduled IF-binding, and (2) specific desmoplakin interactions with either PKP1, PKP2 or PKP3, or combinations thereof, are involved in the selective recruitment of plakophilins to the desmosomal plaques. PMID- 10852827 TI - A constitutive region is responsible for nuclear targeting of 4.1R: modulation by alternative sequences results in differential intracellular localization. AB - Red blood cell protein 4.1, 4.1R, is an extreme variation on the theme of isoform multiplicity. The diverse 4.1R isoforms, mainly generated by alternative pre-mRNA splicing, are localized at different intracellular sites, including the nucleus. To characterize nonerythroid 4.1 proteins lacking the most upstream translation initiation site, analyze their intracellular localization and define specific domains involved in differential intracellular targeting of 4.1R, we cloned 4.1 cDNAs lacking that translation initiation site. Seven different 4.1R cDNAs were isolated. Four of these encoded 4.1R proteins localized predominantly to the nucleus and the other three localized to the cytoplasm. Three of the nuclear 4.1R isoforms did not contain the nuclear localization signal previously identified in the alternative exon 16. A comparative analysis of the exon composition of the naturally occurring 4.1R cDNAs cloned and of appropriate composite cDNA constructs, with the subcellular distribution of their respective products, demonstrated that a region encoded by constitutive exons, which is therefore common to all 4.1R isoforms and has been termed 'core region', had the capacity of localizing to the nucleus. This region was able to confer nuclear targeting to a cytosolic reporter. In protein 4.1R isoforms, the nuclear targeting of the core region is modulated by the expression of alternative exons. Thus, exon 5-encoded sequences eclipsed nuclear entry of the core region, resulting in 4.1R isoforms that predominantly distributed to the cytoplasm. Exon 5 was also able to confer cytoplasmic localization to a nuclear reporter. In protein 4.1R isoforms, when exons 5 and 16 were both expressed the nuclear targeting effect of exon 16 was dominant to the inhibitory effect observed by the expression of exon 5, yielding proteins that predominantly localized to the nucleus. Taken together, these results indicate that all 4.1R molecules contain a conserved region that is sufficient to target the protein to the nucleus, but that specific exon-encoded sequences modulate this capacity by acting in a hierarchical order. PMID- 10852828 TI - XCS-1, a maternally expressed gene product involved in regulating mitosis in Xenopus. AB - The regulation of the cell cycle during early development is an important and complex biological process. We have cloned a cDNA, XCS-1, that may play an important role in regulating mitosis during early embryogenesis in Xenopus laevis. XCS-1 is a maternally expressed gene product that is the Xenopus homologue of the human cleavage signal protein (CS-1). XCS-1 transcripts were detected in oocytes with the titer decreasing just prior to the MBT. During development the XCS-1 protein was detected on the membrane and in the nucleus of blastomeres. It was also detected on the mitotic spindle in mitotic cells and on the centrosomes in interphase cells. Overexpression of myc-XCS-1 in Xenopus embryos resulted in abnormal mitoses with increased numbers of centrosomes, multipolar spindles, and abnormal distribution of chromosomes. Also, we observed incomplete cytokinesis resulting in multiple nuclei residing in the same cytoplasm with the daughter nuclei in different phases of the cell cycle. The phenotype depended on the presence of the N terminus of XCS-1 (aa 1-73) and a consensus NIMA kinase phosphorylation site (aa159-167). Mutations in this site affected the ability of the overexpressed XCS-1 protein to produce the phenotype. These results suggest that XCS-1 is a maternal factor playing an important role in the regulation of the cell cycle during early embryogenesis and that its function depends on its state of phosphorylation. PMID- 10852829 TI - Coupled transport of p24 family members. AB - Recent studies show that small trans-membrane proteins of approximately 22-24 kDa (the p24 family), which are grouped into 4 sub-families by sequence homology (p23, p24, p25 and p26), are involved in the early secretory pathway. In this study, we have investigated the mutual requirements of ectopically expressed members of the p24 family for targeting to their proper cellular destination. We find that coexpression of p23 and p24 is both necessary and sufficient for each protein to be transported to the cis-Golgi network/Golgi complex. Proteins from other subfamilies did not substitute for either p23 or p24, even after multiple coexpression. However, trafficking of the p23/p24 couple was facilitated by coexpression of proteins from other sub-families. In addition, we find that the sequence resembling an endoplasmic reticulum retrieval signal present in the cytoplasmic domain of p23 (but not p24) is dispensable. In contrast, the conserved coiled-coil region in the lumenal domain is absolutely required in both p23 and p24 for proper targeting of the p23/p24 couple. These data demonstrate that p23 and p24 must interact with each other to reach their destination, but that this strict requirement is combined with a mutual dependence amongst p24 proteins. We speculate that p24 proteins can form different oligomeric complexes, which contribute to confer specialized sorting/trafficking properties to membranes of the early secretory pathway, perhaps serving as membrane organizers. PMID- 10852830 TI - The developing brain and the environment: an introduction. AB - mental retardation: timing and thresholds; (italic)b(/italic)) endocrine dysfunction and developmental disabilities: dose and target implications; (italic)c(/italic)) attention-deficit disorder-ADHD and learning disabilities; and (italic)d(/italic)) new horizons: extending the boundaries. Support for the Rochester conference came from both public and private sources. The National Institute of Environmental Health Sciences (NIEHS), the National Institute of Child Health and Human Development, and the EPA represented the federal government. The conference also received grants from several foundations: the Jennifer Altman Foundation, the Heinz Family Foundation, the National Alliance for Autism Research, the Violence Research Foundation, the Wacker Foundation, and the Winslow Foundation. The second of these conferences helped launch a new Center for Children's Health and the Environment at the Mount Sinai School of Medicine. It was held in New York City on 24-25 May 1999, and was convened specifically to consider the intersection between neurodevelopmental impairment, environmental chemicals, and prevention. Over 300 health scientists, pediatricians, and public health professionals examined the growing body of evidence linking environmental toxins to neurobehavioral disorders. The conference title was Environmental Influences on Children: Brain, Development, and Behavior. The conference began by reviewing well-known examples of deleterious effects of environmental chemicals, including lead and PCBs, on children's brains. The conferees then considered the potential impact of environmental chemicals on neurological disorders with particular focus on ADHD, autism, and Parkinson's disease. The inclusion of Parkinson's disease was intended to signal the notion that exposures in early life may have an influence on the evolution of neurological disease in later life. Support for the Mount Sinai conference came from the Superfund Basic Research Program (NIEHS); The Pew Charitable Trusts; the Institute for Health and the Environment at the University of Albany School of Public Health; the Agency for Toxic Substances and Disease Research (ATSDR); the Ambulatory Pediatric Association; Myron A. Mehlman, PhD; the National Center for Environmental Assessment (EPA); the National Center for Environmental Health (CDC); the National Institute of Child Health and Human Development; the Office of Children's Health Protection (EPA); Physicians for Social Responsibility; The New York Academy of Medicine; The New York Community Trust; and the Wallace Genetic Foundation. The impact of environmental toxins on children's health has become a topic of major concern in the federal government. Eight new research centers in children's environmental health have been established in the past 2 years with joint funding from EPA and NIEHS. Clinical units that specialize in the treatment of children with environmentally induced illness have been developed across the nation with grant support from ATSDR. The American Academy of Pediatrics has just published its (italic)Handbook of Pediatric Environmental Health (/italic)((italic)17(/italic)), the "Green Book," which is available to pediatricians throughout the Americas. Children's environmental health has climbed to a critical position as we launch the new millennium. This monograph marks a significant milestone in the evolution of this emerging discipline. PMID- 10852831 TI - Vulnerability of children and the developing brain to neurotoxic hazards. AB - For much of the history of toxicology, the sensitivity of the developing organism to chemical perturbation attracted limited attention. Several tragic episodes and new insights finally taught us that the course of early brain development incurs unique risks. Although the process is exquisitely controlled, its lability renders it highly susceptible to damage from environmental chemicals. Such disturbances, as recognized by current testing protocols and legislation such as the Food Quality Protection Act, can result in outcomes ranging from death to malformations to functional impairment. The latter are the most difficult to determine. First, they require a variety of measures to assay their extent. Second, adult responses may prove an inadequate guide to the response of the developing brain, which is part of the reason for proposing additional safety factors for children. Third, neuropsychological tests are deployed in complex circumstances in which many factors, including economic status, combine to produce a particular effect such as lowered intelligence quotient score. Fourth, the magnitude of the effect, for most environmental exposure levels, may be relatively small but extremely significant for public health. Fifth, changes in brain function occur throughout life, and some consequences of early damage may not even emerge until advanced age. Such factors need to be addressed in estimating the influence of a particular agent or group of agents on brain development and its functional expression. It is especially important to consider ways of dealing with multiple risks and their combinations in addition to the prevailing practice of estimating risks in isolation. PMID- 10852832 TI - Environmental agents that have the potential to trigger massive apoptotic neurodegeneration in the developing brain. AB - We review recent findings pertaining to several environmental agents (ethanol, phencyclidine, ketamine, nitrous oxide, barbiturates, benzodiazepines, halothane, isoflurane, and propofol) that have the potential to delete large numbers of neurons from the developing brain by a newly discovered mechanism involving interference in the action of neurotransmitters [glutamate and gamma-amino butyric acid (GABA) at (italic)N(/italic)-methyl-d-aspartate (NMDA)] and GABA(subscript)A(/subscript) receptors during the synaptogenesis period, also known as the brain growth-spurt period. Transient interference (lasting >= 4 hr) in the activity of these transmitters during the synaptogenesis period (the last trimester of pregnancy and the first several years after birth in humans) causes millions of developing neurons to commit suicide (die by apoptosis). Many of these agents are drugs of abuse (ethanol is a prime example) to which the human fetal brain may be exposed during the third trimester by drug-abusing mothers. Ethanol triggers massive apoptotic neurodegeneration in the developing brain by interfering with both the NMDA and GABA(subscript)A(/subscript) receptor systems, and this can explain the reduced brain mass and lifelong neurobehavioral disturbances associated with intrauterine exposure of the human fetus to ethanol (fetal alcohol syndrome). Exposure of the immature brain in a medical treatment context is also of concern because many of these agents are drugs used frequently as sedatives, tranquilizers, anticonvulsants, or anesthetics in pediatric and/or obstetrical medicine. Because this is a newly discovered mechanism, further research will be required to fully ascertain the nature and degree of risk posed by exposure of the developing human brain to environmental agents that act by this mechanism. PMID- 10852833 TI - Examining childhood development in contaminated urban settings. AB - Normal childhood development and growth is affected by such factors as genetics, nutrition, and multiple familial and social factors. In large urban settings, children are constantly exposed to varying amounts of assorted toxic chemicals both inside and outside the home. Many of these contaminants are suspected to be associated with developmental alterations. The heterogeneity of risk factors in urban populations poses a challenging situation for research. Change must be made in the manner in which developmental toxicological research is undertaken. Plans should be made for immediate data collection after a large-scale exposure to prevent the loss of valuable information. Retrospective studies would benefit from applying rapid assessment techniques to identify high- and low-risk children. In all cases, the development of research design and investigative format needs to reflect the strengths of both social factors and scientific facts. Cross-disciplinary approaches, using physicians and physical and social scientists and incorporating community knowledge, are required for the evaluation of children in urban settings, with each discipline contributing to theory and methodology. PMID- 10852834 TI - Mental retardation and developmental disabilities influenced by environmental neurotoxic insults. AB - This paper sets a framework for the discussion of neurotoxicity as a potentially major contributor to the etiology of many types of mental retardation and developmental disabilities. In the past the literatures on developmental neurotoxicology and on mental retardation have evolved independently, yet we know that the developing brain is a target for neurotoxicity in the developing central nervous system through many stages of pregnancy as well as during infancy and early childhood. Our definitions and theories of mental retardation and developmental disabilities affect the models of neurotoxicity we espouse. For instance, models of developmental risk in neurotoxicology have guided environmental regulation to reduce the likelihood of neurotoxic effects. On the other hand, models of developmental risk for mental retardation aim not only at primary prevention,but also at secondary and tertiary prevention through early intervention. In the future, dynamic models of neuroplasticity based on the study of gene-brain-behavior relationships are likely to guide our views of developmental neurotoxicology and prevention of mental retardation and other disabilities. PMID- 10852835 TI - The environment as an etiologic factor in autism: a new direction for research. AB - Autism is one of a group of developmental disorders that have devastating lifelong effects on its victims. Despite the severity of the disease and the fact that it is relatively common (15 in 10,000), there is still little understanding of its etiology. Although believed to be highly genetic, no abnormal genes have been found. Recent findings in autism and in related disorders point to the possibility that the disease is caused by a gene-environment interaction. Epidemiologic studies indicate that the number of cases of autism is increasing dramatically each year. It is not clear whether this is due to a real increase in the disease or whether this is an artifact of ascertainment. A new theory regarding the etiology of autism suggests that it may be a disease of very early fetal development (approximately day 20-24 of gestation). This theory has initiated new lines of investigation into developmental genes. Environmental exposures during pregnancy could cause or contribute to autism based on the neurobiology of these genes. PMID- 10852836 TI - Parallels between attention deficit hyperactivity disorder and behavioral deficits produced by neurotoxic exposure in monkeys. AB - Attention deficit hyperactivity disorder (ADHD) is a disability that affects between 3 and 7% of children, with a significant number of individuals continuing to be affected into adolescence and adulthood. ADHD is characterized in part by an inability to organize complex sequences of behavior, to persist in the face of distracting stimuli, and to respond appropriately to the consequences of past behavior. There are some parallels between the features of ADHD and the behavior of monkeys exposed developmentally to lead or polychlorinated biphenyls (PCBs), as evidenced by research from our laboratory. Both lead and PCB exposure produce deficits on discrimination reversal and spatial delayed alternation performance; treated monkeys exhibit deficits in their ability to change an already established response strategy and inhibit inappropriate responses. Monkeys exposed developmentally to lead or PCBs also perform differently from control monkeys on a fixed interval schedule of reinforcement, which requires the temporal organization of behavior using only internal cues. Whereas the etiology of ADHD is multifactorial, the possibility that neurotoxic agents in the environment contribute to the incidence of ADHD warrants attention. PMID- 10852837 TI - Dopamine function in Lesch-Nyhan disease. AB - Lesch-Nyhan disease is a disorder of purine metabolism resulting from mutations in the gene for hypoxanthine guanine phosphoribosyl transferase on the X chromosome. It is characterized by hyperuricemia and all of its consequences, as in gout; but in addition, patients have impressive disease of the central nervous system. This includes spasticity, involuntary movements, and retardation of motor development. The behavioral phenotype is best remembered by self-injurious biting behavior with attendant destruction of tissue. The connection between aberrant metabolism of purines and these neurologic and behavioral features of the disease is not clear. Increasing evidence points to imbalance of neurotransmitters. There is increased excretion of the serotonin metabolite 5-hydroxyindoleacetic acid in the urine. There are decreased quantities and activities of a number of dopaminergic functions. Positron emission tomography scanning has indicated deficiency in the dopamine transporter. PMID- 10852838 TI - Does methylmercury have a role in causing developmental disabilities in children? AB - Methylmercury (MeHg) is a potent neurotoxin that in high exposures can cause mental retardation, cerebral palsy, and seizures. The developing brain appears particularly sensitive to MeHg. Exposure levels in pregnant experimental animals that do not result in detectable signs or symptoms in the mother can adversely affect the offspring's development. Studies of human poisonings suggest this may also occur in humans. Human exposure to MeHg is primarily dietary through the consumption of fish: MeHg is present in all fresh and saltwater fish. Populations that depend on fish as a major source of dietary protein may achieve MeHg exposure levels hypothesized to adversely affect brain development. Increasing mercury levels in the environment have heightened concerns about dietary exposure and a possible role for MeHg in developmental disabilities. Follow-up studies of an outbreak of MeHg poisoning in Iraq revealed a dose-response relationship for prenatal MeHg exposure. That relationship suggested that prenatal exposure as low as 10 ppm (measured in maternal hair growing during pregnancy) could adversely affect fetal brain development. However, using the same end points as were used in the Iraq study, no associations have been reported in fish-eating populations. Using a more extensive range of developmental end points, some studies of populations consuming seafood have reported associations with prenatal MeHg exposure, whereas others have found none. This paper reviews the data presently available associating MeHg exposure with development and poses some of the unanswered questions in this field. PMID- 10852839 TI - On categorizations in analyses of alcohol teratogenesis. AB - In biomedical scientific investigations, expositions of findings are conceptually simplest when they comprise comparisons of discrete groups of individuals or involve discrete features or characteristics of individuals. But the descriptive benefits of categorization become outweighed by their limitations in studies involving dose-response relationships, as in many teratogenic and environmental exposure studies. This article addresses a pair of categorization issues concerning the effects of prenatal alcohol exposure that have important public health consequences: the labeling of individuals as fetal alcohol syndrome (FAS) versus fetal alcohol effects (FAE) or alcohol-related neurodevelopmental disorder (ARND), and the categorization of prenatal exposure dose by thresholds. We present data showing that patients with FAS and others with FAE do not have meaningfully different behavioral performance, standardized scores of IQ, arithmetic and adaptive behavior, or secondary disabilities. Similarly overlapping distributions on measures of executive functioning offer a basis for identifying alcohol-affected individuals in a manner that does not simply reflect IQ deficits. At the other end of the teratological continuum, we turn to the reporting of threshold effects in dose-response relationships. Here we illustrate the importance of multivariate analyses using data from the Seattle, Washington, longitudinal prospective study on alcohol and pregnancy. Relationships between many neurobehavioral outcomes and measures of prenatal alcohol exposure are monotone without threshold down to the lowest nonzero levels of exposure, a finding consistent with reports from animal studies. In sum, alcohol effects on the developing human brain appear to be a continuum without threshold when dose and behavioral effects are quantified appropriately. PMID- 10852840 TI - Manganese: brain transport and emerging research needs. AB - Idiopathic Parkinson's disease (IPD) represents a common neurodegenerative disorder. An estimated 2% of the U.S. population, age 65 and older, develops IPD. The number of IPD patients will certainly increase over the next several decades as the baby-boomers gradually step into this high-risk age group, concomitant with the increase in the average life expectancy. While many studies have suggested that industrial chemicals and pesticides may underlie IPD, its etiology remains elusive. Among the toxic metals, the relationship between manganese intoxication and IPD has long been recognized. The neurological signs of manganism have received close attention because they resemble several clinical disorders collectively described as extrapyramidal motor system dysfunction, and in particular, IPD and dystonia. However, distinct dissimilarities between IPD and manganism are well established, and it remains to be determined whether Mn plays an etiologic role in IPD. It is particularly noteworthy that as a result of a recent court decision, methylcyclopentadienyl Mn tricarbonyl (MMT) is presently available in the United States and Canada for use in fuel, replacing lead as an antiknock additive. The impact of potential long-term exposure to low levels of MMT combustion products that may be present in emissions from automobiles has yet to be fully evaluated. Nevertheless, it should be pointed out that recent studies with various environmental modeling approaches in the Montreal metropolitan (where MMT has been used for more than 10 years) suggest that airborne Mn levels were quite similar to those in areas where MMT was not used. These studies also show that Mn is emitted from the tail pipe of motor vehicles primarily as a mixture of manganese phosphate and manganese sulfate. This brief review characterizes the Mn speciation in the blood and the transport kinetics of Mn into the central nervous system, a critical step in the accumulation of Mn within the brain, outlines the potential susceptibility of selected populations (e.g., iron-deficient) to Mn exposure, and addresses future research needs for Mn. PMID- 10852842 TI - New horizons: future directions in neurotoxicology. AB - Neurotoxicology is a relatively young discipline that has undergone significant growth during the last 25 years. During the late 1970s and 1980s, numerous national and international conferences and meetings were devoted to the topic of neurotoxicology, the formation of societies or specialty sections related to neurotoxicology, and the establishment of two independent peer-reviewed journals devoted to neurotoxicology. This decade was also associated with a rapid increase in our knowledge of chemical effects on the structure and function of the nervous system. During the 1990s, regulatory agencies such as the U.S. Environmental Protection Agency accepted neurotoxicology as a crucial end point and neurotoxicity testing and risk assessment guidelines were published. Neurotoxicology has also been accepted at the international level as evidenced by environmental criteria documents published by the International Programme on Chemical Safety and testing guidelines by the Organization of Economic Cooperation and Development. In recent years, there has been increased concern that the etiology of some neurodegenerative diseases may be associated with exposure to neurotoxic agents and that subpopulations of humans such as children and the elderly may be differentially sensitive to neurotoxic exposure. In the future, mechanistic information derived from basic research will be used in the identification and characterization of chemicals with neurotoxic potential. PMID- 10852841 TI - Thyroidal dysfunction and environmental chemicals--potential impact on brain development. AB - Certain polyhalogenated aromatic hydrocarbons such as polychlorinated biphenyls (PCBs) and dibenzo-p-dioxins (dioxins, 2,3,7, 8-tetrachlorodibenzo-p-dioxin) have been shown to have neurotoxic effects and to alter thyroid function during critical periods of thyroid hormone-dependent brain development. This has led to the suggestion that some of the neurotoxic effects of these compounds could be mediated through the thyroid system. Thyroid hormones are essential for normal brain development during a critical period beginning in utero and extending through the first 2 years postpartum. They regulate neuronal proliferation, migration, and differentiation in discrete regions of the brain during definitive time periods. Even transient disruption of this normal pattern can impair brain development. Thyroid hormones are necessary for normal cytoskeletal assembly and stability and the cytoskeletal system is essential for migration and neuronal outgrowth. In addition, they regulate development of cholinergic and dopaminergic systems serving the cerebral cortex and hippocampus. Animals perinatally exposed to certain environmental organohalogens such as many of the PCBs and dioxins have abnormal thyroid function and neurologic impairment. Although there are both species and congener variabilities, most reports show exposure results in thyroid enlargement and reduced serum T(4) levels with normal T(3) levels. Initial research concentrated on studying the direct actions of xenobiotics on the thyroid; however, some of these compounds bear a structural resemblance to the natural thyroid hormones and have high affinity with thyroid hormone-binding proteins such as transthyretin. These compounds could act as agonists or antagonists for receptors of the thyroid/steroid/retinoic acid superfamily. These structurally similar organohalogens could act at multiple points to alter thyroid hormone action. The similarity of the neurologic impairment seen in thyroid disorders to that seen following PCB or dioxin exposure suggests that one mechanism of neurotoxicity of these compounds could involve interaction with the thyroid system. PMID- 10852843 TI - Chemicals in the environment and developmental toxicity to children: a public health and policy perspective. AB - There are numerous pesticides and toxic chemicals in the environment that have yet to be evaluated for potential to cause developmental neurotoxicity. Recent legislation and testing initiatives provide an impetus to generating more information about potential hazards to children. In the United States, the 1996 Food Quality Protection Act (FQPA) required the U.S. Environmental Protection Agency (U.S. EPA) to make a finding that a pesticide food use is safe for children. In addition, the law requires U.S. EPA to incorporate an additional 10 fold factor in risk assessments for pesticide residue tolerances to take into account the special sensitivities of infants and children as well as incomplete data with respect to toxicity and exposures. The potential of chemicals in food and drinking water to cause endocrine disruption will also be examined via the Endocrine Disruptor Screening and Testing Program required by the FQPA and the 1996 Safe Drinking Water Act. In addition, a new voluntary chemical information program will provide screening-level information for the some 2,800 high-volume chemicals in commerce in the United States. These initiatives will need to be accompanied by research focused on developmental toxicity for children, including developmental disabilities. Developmental disabilities exact a large toll on children's health in the United States. Three major developmental disabilities- autism, cerebral palsy, and severe mental retardation--each affect substantial numbers of children. We know very little about the etiology of these conditions. A number of priority areas for research are suggested, including a large environmental prospective study of developmental neurotoxicity. PMID- 10852844 TI - Identifying critical windows of exposure for children's health. AB - Several authors have considered the importance of exposure timing and how this affects the outcomes observed, but no one has systematically compiled preconceptional, prenatal, and postnatal developmental exposures and subsequent outcomes. Efforts were undertaken to examine the information available and to evaluate implications for risk assessment for several areas: a) respiratory and immune systems, b) reproductive system, c) nervous system, d) cardiovascular system, endocrine system, and general growth, and e) cancer. Major conclusions from a workshop on "Critical Windows of Exposure for Children's Health" included a) broad windows of sensitivity can be identified for many systems but detailed information is limited; b) cross-species comparisons of dose to target tissue and better data on the exposure-dose-outcome continuum are needed; c) increased interaction among scientific disciplines can further understanding by using laboratory animal results in designing epidemiological studies and human data to suggest specific laboratory studies on mechanisms and agent-target interactions; and d) thus far, only limited attention has been given to peripubertal/adolescent exposures, adult consequences of developmental exposures, and genome-environment interactions. More specific information on developmental windows will improve risk assessment by identifying the most sensitive window(s) for evaluation of dose-response relationships and exposure, evaluation of biological plausibility of research findings in humans, and comparison of data across species. In public health and risk management, information on critical windows may help identify especially susceptible subgroups for specific interventions. PMID- 10852845 TI - The mammalian respiratory system and critical windows of exposure for children's health. AB - The respiratory system is a complex organ system composed of multiple cell types involved in a variety of functions. The development of the respiratory system occurs from embryogenesis to adult life, passing through several distinct stages of maturation and growth. We review embryonic, fetal, and postnatal phases of lung development. We also discuss branching morphogenesis and cellular differentiation of the respiratory system, as well as the postnatal development of xenobiotic metabolizing systems within the lungs. Exposure of the respiratory system to a wide range of chemicals and environmental toxicants during perinatal life has the potential to significantly affect the maturation, growth, and function of this organ system. Although the potential targets for exposure to toxic factors are currently not known, they are likely to affect critical molecular signals expressed during distinct stages of lung development. The effects of exposure to environmental tobacco smoke during critical windows of perinatal growth are provided as an example leading to altered cellular and physiological function of the lungs. An understanding of critical windows of exposure of the respiratory system on children's health requires consideration that lung development is a multistep process and cannot be based on studies in adults. PMID- 10852847 TI - Development of atopy and asthma: candidate environmental influences and important periods of exposure. AB - Atopy is a major risk factor for the development of asthma. Immune processes that lead to the development of antigen-specific IgE are essential to the development of atopy. This review examines the immune processes that are candidate targets for modulation by environmental agents; environmental and lifestyle factors that have been suggested as modulators of the development of atopy; and the impact of known environmental agents on atopic processes in the airway. The most important periods of immune development with regard to expression of atopy are likely during gestation and early childhood. A better understanding of which environmental agents are important, as well as the period of life during which these agents may exert an important effect, is essential to devising rational environmental avoidance strategies for at-risk populations. PMID- 10852846 TI - Development of the murine and human immune system: differential effects of immunotoxicants depend on time of exposure. AB - Fetal and early postnatal life represent critical periods in vertebrate immune system development. Disruption of such development by perinatal immunotoxic chemical exposure has been widely described in experimental animal models. The resultant inhibited postnatal immune responses in such animals are often more dramatic and persistent than those after exposure during adult life. Further, recent reports suggest that prenatal exposure to immunotoxicants may exacerbate postnatal aberrant immune responses (e.g., hypersensitivity disorders and autoimmune disease) in genetically predisposed rodents. Limited information is available regarding the possibility of inhibited postnatal immune capacity in humans as a result of developmental immunotoxicant exposure. The multifactorial nature of hypersensitivity and autoimmune responses will further complicate the elucidation of possible relationships between chemical exposure during ontogeny of the human immune system and immune-mediated disease later in life. Taken together, however, the available animal data suggest the potential for altered postnatal immune function in humans exposed to immunotoxicants (e.g., environmental chemicals and therapeutic agents) during fetal and/or early postnatal life. PMID- 10852848 TI - Workshop to identify critical windows of exposure for children's health: immune and respiratory systems work group summary. AB - Fetuses, infants, and juveniles (preadults) should not be considered simply "small adults" when it comes to toxicological risk. We present specific examples of developmental toxicants that are more toxic to children than to adults, focusing on effects on the immune and respiratory systems. We describe differences in both the pharmacokinetics of the developing immune and respiratory systems as well as changes in target organ sensitivities to toxicants. Differential windows of vulnerability during development are identified in the context of available animal models. We provide specific approaches to directly investigate differential windows of vulnerability. These approaches are based on fundamental developmental biology and the existence of discrete developmental processes within the immune and respiratory systems. The processes are likely to influence differential developmental susceptibility to toxicants, resulting in lifelong toxicological changes. We also provide a template for comparative research. Finally, we discuss the application of these data to risk assessment. PMID- 10852849 TI - Critical windows of exposure for children's health: the reproductive system in animals and humans. AB - Drugs and environmental chemicals can adversely affect the reproductive system. Currently, available data indicate that the consequences of exposure depend on the nature of the chemical, its target, and the timing of exposure relative to critical windows in development of the reproductive system. The reproductive system is designed to produce gametes in far greater excess than would seem to be necessary for the survival of species. Ten to hundreds of millions of spermatozoa are generated daily by most adult male mammals, yet very few of these germ cells succeed in transmitting their genetic material to the next generation. Although the number of oocytes produced in mammalian females is more limited, and their production occurs only during fetal life, most ovaries contain several orders of magnitude more oocytes than ever will be fertilized. Toxicant exposures may affect critical events in the development of the reproductive system, ranging from early primordial germ cell determination to gonadal differentiation, gametogenesis, external genitalia, or signaling events regulating sexual behavior. Although there are differences between the human reproductive system and that of the usual animal models, such models have been extremely useful in assessing risks for key human reproductive and developmental processes. The objectives for future studies should include the elucidation of the specific cellular and molecular targets of known toxicants; the design of a systematic approach to the identification of reproductive toxicants; and the development of sensitive, specific, and predictive animal models, minimally invasive surrogate markers, or in vitro tests to assess reproductive system function during embryonic, postnatal, and adult life. PMID- 10852850 TI - Workshop to identify critical windows of exposure for children's health: reproductive health in children and adolescents work group summary. AB - This work group report addresses the central question: What are the critical windows during development (preconception through puberty) when exposure to xenobiotics may have the greatest adverse impact on subsequent reproductive health? The reproductive system develops in stages, with sex-specific organogenesis occurring prenatally and further maturational events occurring in the perinatal period and at puberty. Complex endocrine signals as well as other regulatory factors (genetics, growth factors) are involved at all stages. Evidence from animal models and human studies indicates that many specific events can be perturbed by a variety of toxicants, with endocrine-mediated mechanisms being the more widely studied. Prioritized research needs include basic studies on the cellular-molecular and endocrine regulation of sexual differentiation and development; increased efforts regarding potential adverse effects on development in females, including breast development; expanded animal studies on different classes of chemicals, comparing responses during development (prenatal and postnatal) with responses in adults; and, more extensive explorations regarding the reproductive biology and toxicology of puberty in humans. PMID- 10852852 TI - Workshop to identify critical windows of exposure for children's health: neurobehavioral work group summary. AB - This paper summarizes the deliberations of a work group charged with addressing specific questions relevant to risk estimation in developmental neurotoxicology. We focused on eight questions. a) Does it make sense to think about discrete windows of vulnerability in the development of the nervous system? If it does, which time periods are of greatest importance? b) Are there cascades of developmental disorders in the nervous system? For example, are there critical points that determine the course of development that can lead to differences in vulnerabilities at later times? c) Can information on critical windows suggest the most susceptible subgroups of children (i.e., age groups, socioeconomic status, geographic areas, race, etc.)? d) What are the gaps in existing data for the nervous system or end points of exposure to it? e) What are the best ways to examine exposure-response relationships and estimate exposures in vulnerable life stages? f) What other exposures that affect development at certain ages may interact with exposures of concern? g) How well do laboratory animal data predict human response? h) How can all of this information be used to improve risk assessment and public health (risk management)? In addressing these questions, we provide a brief overview of brain development from conception through adolescence and emphasize vulnerability to toxic insult throughout this period. Methodological issues focus on major variables that influence exposure or its detection through disruptions of behavior, neuroanatomy, or neurochemical end points. Supportive evidence from studies of major neurotoxicants is provided. PMID- 10852853 TI - Fetal, infant, and childhood growth are predictors of coronary heart disease, diabetes, and hypertension in adult men and women. AB - Many human fetuses have to adapt to a limited supply of nutrients. In doing so they permanently change their structure and metabolism. These programmed changes may be the origins of a number of diseases in later life, including coronary heart disease, hypertension, and noninsulin- dependent diabetes. We review epidemiologic studies in which the incidence of these diseases has been related to the recorded, early growth of individuals, while considering factors in the adult lifestyle, such as obesity and socioeconomic status. We discuss possible mechanisms. For hypertension these mechanisms include placentation, maternal blood pressure, fetal undernutrition; childhood growth, activation of the renin angiotensin system, renal structure, programming of the hypothalamic-pituitary adrenal axis, vascular structure, and sympathetic nervous activity. For noninsulin-dependent diabetes we discuss mechanisms concerning both insulin resistance and insulin deficiency. We include a review of evidence for the programming of serum cholesterol and clotting factor concentrations. We address the timing of critical windows for coronary heart disease, reviewing studies that allow assessment of the relative importance of fetal, infant, and childhood growth. We argue for a research strategy that combines clinical, animal, and epidemiological studies. PMID- 10852854 TI - Susceptible periods during embryogenesis of the heart and endocrine glands. AB - One of the original principles of teratology states that, "Susceptibility to teratogenesis varies with the developmental stage at the time of exposure to an adverse influence" [Wilson JG. Environment and Birth Defects. New York:Academic Press, 1973]. The time of greatest sensitivity encompasses the period of organ formation during weeks 3-8 following fertilization in human gestation. At this time, stem cell populations for each organ's morphogenesis are established and inductive events for the initiation of differentiation occur. Structural defects of the heart and endocrine system are no exception to this axiom and have their origins during this time frame. Although the function and maturation of these organs may be affected at later stages, structural defects and loss of cell types usually occur during these early phases of development. Thus, to determine critical windows for studying mechanisms of teratogenesis, it is essential to understand the developmental processes that establish these organs. PMID- 10852851 TI - Critical periods of vulnerability for the developing nervous system: evidence from humans and animal models. AB - Vulnerable periods during the development of the nervous system are sensitive to environmental insults because they are dependent on the temporal and regional emergence of critical developmental processes (i.e., proliferation, migration, differentiation, synaptogenesis, myelination, and apoptosis). Evidence from numerous sources demonstrates that neural development extends from the embryonic period through adolescence. In general, the sequence of events is comparable among species, although the time scales are considerably different. Developmental exposure of animals or humans to numerous agents (e.g., X-ray irradiation, methylazoxymethanol, ethanol, lead, methyl mercury, or chlorpyrifos) demonstrates that interference with one or more of these developmental processes can lead to developmental neurotoxicity. Different behavioral domains (e.g., sensory, motor, and various cognitive functions) are subserved by different brain areas. Although there are important differences between the rodent and human brain, analogous structures can be identified. Moreover, the ontogeny of specific behaviors can be used to draw inferences regarding the maturation of specific brain structures or neural circuits in rodents and primates, including humans. Furthermore, various clinical disorders in humans (e.g., schizophrenia, dyslexia, epilepsy, and autism) may also be the result of interference with normal ontogeny of developmental processes in the nervous system. Of critical concern is the possibility that developmental exposure to neurotoxicants may result in an acceleration of age-related decline in function. This concern is compounded by the fact that developmental neurotoxicity that results in small effects can have a profound societal impact when amortized across the entire population and across the life span of humans. PMID- 10852855 TI - Influences of pre- and postnatal nutritional exposures on vascular/endocrine systems in animals. AB - Human epidemiological and animal studies have revealed the long-term effects of malnutrition during gestation and early life on the health of the offspring. The aim of the current review is to survey the different means of achieving fetal malnutrition and its consequences, mainly in animals, and to identify key areas in which to direct future research. We address the impact of various models of a maternal protein-restricted diet and global maternal caloric restriction (either through the reduction of nutrient supply or through mechanic devices), the influence of maternal diabetes, and other maternal causes of fetal damage (maternal infections and toxic food components). More specifically, we enumerate data on how the different insults at different prenatal and early postnatal periods affect and program the development and the function of organs involved in diabetes, hypertension, and cardiovascular disease. Particular emphasis is given to the endocrine pancreas, but insulin-sensitive tissues, kidneys, and vasculature are also analyzed. Where available, the protective effects of maternal food supplementation for fetal organ development and function are discussed. Specific attention is paid to the amino acids profile, and the preventive role of taurine is discussed. Tentative indications about critical time windows for fetal development under different deleterious conditions are presented whenever possible. We also discuss future research and intervention. PMID- 10852856 TI - Workshop to identify critical windows of exposure for children's health: cardiovascular and endocrine work group summary. AB - The work group on cardiovascular and endocrine effects was asked to review the current state of knowledge about children's windows of vulnerability to developmental toxicants and to recommend how that information may be used to improve risk assessment and public health. We considered differences between structural defects, where periods of vulnerability are rather well defined, and functional defects, where periods of vulnerability are quite elusive. PMID- 10852858 TI - Workshop to identify critical windows of exposure for children's health: cancer work group summary. AB - We considered whether there are discrete windows of vulnerability in the development of cancer and which time periods may be of the greatest importance. Cancer was considered broadly, including cancers in childhood as well as adult cancers that may have an in utero or childhood origin. We concluded that there was evidence from animal and epidemiologic studies for causal relationships for preconceptional, in utero, and childhood exposures and cancer occurrence in children and adults. However, the evidence is incomplete and all relevant critical windows may not have been identified. The comprehensive evaluation of the relative importance of specific time windows of exposure is limited. Improvements in the design of epidemiologic studies and additional animal studies of mechanisms are warranted. PMID- 10852859 TI - Living in stools is not as dumb as you think. PMID- 10852857 TI - Critical windows of exposure for children's health: cancer in human epidemiological studies and neoplasms in experimental animal models. AB - In humans, cancer may be caused by genetics and environmental exposures; however, in the majority of instances the identification of the critical time window of exposure is problematic. The evidence for exposures occurring during the preconceptional period that have an association with childhood or adulthood cancers is equivocal. Agents definitely related to cancer in children, and adulthood if exposure occurs in utero, include: maternal exposure to ionizing radiation during pregnancy and childhood leukemia and certain other cancers, and maternal use of diethylstilbestrol during pregnancy and clear-cell adenocarcinoma of the vagina of their daughters. The list of environmental exposures that occur during the perinatal/postnatal period with potential to increase the risk of cancer is lengthening, but evidence available to date is inconsistent and inconclusive. In animal models, preconceptional carcinogenesis has been demonstrated for a variety of types of radiation and chemicals, with demonstrated sensitivity for all stages from fetal gonocytes to postmeiotic germ cells. Transplacental and neonatal carcinogenesis show marked ontogenetic stage specificity in some cases. Mechanistic factors include the number of cells at risk, the rate of cell division, the development of differentiated characteristics including the ability to activate and detoxify carcinogens, the presence of stem cells, and possibly others. Usefulness for human risk estimation would be strengthened by the study of these factors in more than one species, and by a focus on specific human risk issues. PMID- 10852860 TI - Point mutations in a peptidoglycan biosynthesis gene cause competence induction in Haemophilus influenzae. AB - We have identified three new Haemophilus influenzae mutations causing cells to exhibit extreme hypercompetence at all stages of growth. The mutations are in murE, which encodes the meso-diaminopimelate-adding enzyme of peptidoglycan synthesis. All are point mutations causing nonconservative amino acid substitutions, two at a poorly conserved residue (G(435)-->R and G(435)-->W) and the third at a highly conserved leucine (L(361)-->S). The mutant strains have very similar phenotypes and do not exhibit any defects in cell growth, permeability, or sensitivity to peptidoglycan antibiotics. Cells retain the normal specificity of DNA uptake for the H. influenzae uptake signal sequence. The mutations do not bypass genes known to be needed for competence induction but do dramatically increase expression of genes required for the normal pathway of DNA uptake. We conclude that the mutations do not act by increasing cell permeability but by causing induction of the normal competence pathway via a previously unsuspected signal. PMID- 10852861 TI - Genetic antagonism and hypermutability in Mycobacterium smegmatis. AB - Multidrug-resistant strains of Mycobacterium tuberculosis are a serious and continuing human health problem. Such strains may contain as many as four or five different mutations, and M. tuberculosis strains that are resistant to both streptomycin and rifampin contain mutations in the rpsL and rpoB genes, respectively. Coexisting mutations of this kind in Escherichia coli have been shown to interact negatively (S. L. Chakrabarti and L. Gorini, Proc. Natl. Acad. Sci. USA 72:2084-2087, 1975; S. L. Chakrabarti and L. Gorini, Proc. Natl. Acad. Sci. USA 74:1157-1161, 1977). We investigated this possibility in Mycobacterium smegmatis by analyzing the frequency and nature of spontaneous mutants that are resistant to either streptomycin or rifampin or to both antibiotics. Mutants resistant to streptomycin were isolated from characterized rifampin-resistant mutants of M. smegmatis under selection either for one or for both antibiotics. Similarly, mutants resistant to rifampin were isolated from streptomycin resistant strains. The second antibiotic resistance mutation occurred at a lower frequency in both cases. Surprisingly, in both cases a very high rate of reversion of the initial antibiotic resistance allele was detected when single antibiotic selection was used; the majority of strains resistant to only one antibiotic were isolated by this process. Determinations of rates of mutation to antibiotic resistance in M. smegmatis showed that the frequencies were enhanced up to 10(4)-fold during stationary phase. If such behavior is also typical of slow-growing pathogenic mycobacteria, these studies suggest that the generation of multiply drug-resistant strains by successive mutations may be a more complex genetic phenomenon than suspected. PMID- 10852862 TI - Cyanobacterial sulfide-quinone reductase: cloning and heterologous expression. AB - The gene encoding sulfide-quinone reductase (SQR; E.C.1.8.5.'), the enzyme catalyzing the first step of anoxygenic photosynthesis in the filamentous cyanobacterium Oscillatoria limnetica, was cloned by use of amino acid sequences of tryptic peptides as well as sequences conserved in the Rhodobacter capsulatus SQR and in an open reading frame found in the genome of Aquifex aeolicus. SQR activity was also detected in the unicellular cyanobacterium Aphanothece halophytica following sulfide induction, with a V(max) of 180 micromol of plastoquinone-1 (PQ-1) reduced/mg of chlorophyll/h and apparent K(m) values of 20 and 40 microM for sulfide and quinone, respectively. Based on the conserved sequences, the gene encoding A. halophytica SQR was also cloned. The SQR polypeptides deduced from the two cyanobacterial genes consist of 436 amino acids for O. limnetica SQR and 437 amino acids for A. halophytica SQR and show 58% identity and 74% similarity. The calculated molecular mass is about 48 kDa for both proteins; the theoretical isoelectric points are 7.7 and 5.6 and the net charges at a neutral pH are 0 and -14 for O. limnetica SQR and A. halophytica SQR, respectively. A search of databases showed SQR homologs in the genomes of the cyanobacterium Anabaena PCC7120 as well as the chemolithotrophic bacteria Shewanella putrefaciens and Thiobacillus ferrooxidans. All SQR enzymes contain characteristic flavin adenine dinucleotide binding fingerprints. The cyanobacterial proteins were expressed in Escherichia coli under the control of the T7 promoter. Membranes isolated from E. coli cells expressing A. halophytica SQR performed sulfide-dependent PQ-1 reduction that was sensitive to the quinone analog inhibitor 2n-nonyl-4-hydroxyquinoline-N-oxide. The wide distribution of SQR genes emphasizes the important role of SQR in the sulfur cycle in nature. PMID- 10852863 TI - An IS257-derived hybrid promoter directs transcription of a tetA(K) tetracycline resistance gene in the Staphylococcus aureus chromosomal mec region. AB - Transcription of the tetA(K) tetracycline resistance determinant encoded by an IS257-flanked cointegrated copy of a pT181-like plasmid, located within the chromosomal mec region of a methicillin-resistant Staphylococcus aureus isolate, has been investigated. The results demonstrated that transcription of tetA(K) in this strain is directed by both an IS257-derived hybrid promoter, which is stronger than the native tetA(K) promoter in the autonomous form of pT181, and a complete outwardly directed promoter identified within one end of IS257. Despite lower gene dosage, the chromosomal configuration was shown to afford a higher level of resistance than that mediated by pT181 in an autonomous multicopy state. Furthermore, competition studies revealed that a strain carrying the chromosomal tetA(K) determinant exhibited a higher level of fitness in the presence of tetracycline but not in its absence. This finding suggests that tetracycline has been a selective factor in the emergence of strains carrying a cointegrated pT181 like plasmid in their chromosomes. The results highlight the potential of IS257 to influence the expression of neighboring genes, a property likely to enhance its capacity to mediate advantageous genetic rearrangements. PMID- 10852864 TI - Hyperrecombination in Streptococcus pneumoniae depends on an atypical mutY homologue. AB - The unusual behavior of the mutation ami36, which generates hyperrecombination in two point crosses, was previously attributed to a localized conversion process changing A/G mispairs into CG pairs. Although the mechanism was found to be dependent on the DNA polymerase I, the specific function responsible for this correction was still unknown. Analysis of the pneumococcal genome sequence has revealed the presence of an open reading frame homologous to the gene mutY of Escherichia coli. The gene mutY encodes an adenine glycosylase active on A/G and A/7,8-dihydro-8-oxoguanine (8-OxoG) mismatches, inducing their repair to CG and C/8-OxoG, respectively. Here we report that disrupting the pneumococcal mutY homologue abolishes the hyperrecombination induced by ami36 and leads to a mutator phenotype specifically enhancing AT-to-CG transversions. The deduced amino acid sequence of the pneumococcal MutY protein reveals the absence of four cysteines, highly conserved in the endonuclease III/MutY glycosylase family, which ligate a [4Fe-4S](2+) cluster. The actual function of this cluster is still intriguing, inasmuch as we show that the pneumococcal gene complements a mutY strain of E. coli. PMID- 10852865 TI - Hypersensitivity of Escherichia coli Delta(uvrB-bio) mutants to 6 hydroxylaminopurine and other base analogs is due to a defect in molybdenum cofactor biosynthesis. AB - We have shown previously that Escherichia coli and Salmonella enterica serovar Typhimurium strains carrying a deletion of the uvrB-bio region are hypersensitive to the mutagenic and toxic action of 6-hydroxylaminopurine (HAP) and related base analogs. This sensitivity is not due to the uvrB excision repair defect associated with this deletion because a uvrB point mutation or a uvrA deficiency does not cause hypersensitivity. In the present work, we have investigated which gene(s) within the deleted region may be responsible for this effect. Using independent approaches, we isolated both a point mutation and a transposon insertion in the moeA gene, which is located in the region covered by the deletion, that conferred HAP sensitivity equal to that conferred by the uvrB-bio deletion. The moeAB operon provides one of a large number of genes responsible for biosynthesis of the molybdenum cofactor. Defects in other genes in the same pathway, such as moa or mod, also lead to the same HAP-hypersensitive phenotype. We propose that the molybdenum cofactor is required as a cofactor for an as yet unidentified enzyme (or enzymes) that acts to inactivate HAP and other related compounds. PMID- 10852866 TI - Inactivation of gltB abolishes expression of the assimilatory nitrate reductase gene (nasB) in Pseudomonas putida KT2442. AB - By using mini-Tn5 transposon mutagenesis, random transcriptional fusions of promoterless bacterial luciferase, luxAB, to genes of Pseudomonas putida KT2442 were generated. Insertion mutants that responded to ammonium deficiency by induction of bioluminescence were selected. The mutant that responded most strongly was genetically analyzed and is demonstrated to bear the transposon within the assimilatory nitrate reductase gene (nasB) of P. putida KT2442. Genetic evidence as well as sequence analyses of the DNA regions flanking nasB suggest that the genes required for nitrate assimilation are not clustered. We isolated three second-site mutants in which induction of nasB expression was completely abolished under nitrogen-limiting conditions. Nucleotide sequence analysis of the chromosomal junctions revealed that in all three mutants the secondary transposon had inserted at different sites in the gltB gene of P. putida KT2442 encoding the major subunit of the glutamate synthase. A detailed physiological characterization of the gltB mutants revealed that they are unable to utilize a number of potential nitrogen sources, are defective in the ability to express nitrogen starvation proteins, display an aberrant cell morphology under nitrogen-limiting conditions, and are impaired in the capacity to survive prolonged nitrogen starvation periods. PMID- 10852867 TI - A mutation in secY that causes enhanced SecA insertion and impaired late functions in protein translocation. AB - A cold-sensitive secY mutant (secY125) with an amino acid substitution in the first periplasmic domain causes in vivo retardation of protein export. Inverted membrane vesicles prepared from this mutant were as active as the wild-type membrane vesicles in translocation of a minute amount of radioactive preprotein. The mutant membrane also allowed enhanced insertion of SecA, and this SecA insertion was dependent on the SecD and SecF functions. These and other observations suggested that the early events in translocation, such as SecA dependent insertion of the signal sequence region, is actually enhanced by the SecY125 alteration. In contrast, since the mutant membrane vesicles had decreased capacity to translocate chemical quantity of pro-OmpA and since they were readily inactivated by pretreatment of the vesicles under the conditions in which a pro OmpA translocation intermediate once accumulated, the late translocation functions appear to be impaired. We conclude that this periplasmic secY mutation causes unbalanced early and late functions in translocation, compromising the translocase's ability to catalyze multiple rounds of reactions. PMID- 10852868 TI - Salmonella enterica serovar typhimurium peptidase B is a leucyl aminopeptidase with specificity for acidic amino acids. AB - Peptidase B (PepB) of Salmonella enterica serovar Typhimurium is one of three broad-specificity aminopeptidases found in this organism. We have sequenced the pepB gene and found that it encodes a 427-amino-acid (46.36-kDa) protein, which can be unambiguously assigned to the leucyl aminopeptidase (LAP) structural family. PepB has been overexpressed and purified. The active enzyme shows many similarities to other members of the LAP family: it is a heat-stable (70 degrees C; 20 min) hexameric ( approximately 270-kDa) metallopeptidase with a pH optimum of 8.5 to 9.5. A detailed study of the substrate specificity of the purified protein shows that it differs from other members of the family in its ability to hydrolyze peptides with N-terminal acidic residues. The preferred substrates for PepB are peptides with N-terminal Asp or Glu residues. Comparison of the amino acid sequence of PepB with those of other LAPs leads to the conclusion that PepB is the prototype of a new LAP subfamily with representatives in several other eubacterial species and to the prediction that the members of this family share the ability to hydrolyze peptides with N-terminal acidic residues. Site-directed mutagenesis has been used to show that this specificity appears to be determined by a single Lys residue present in a sequence motif conserved in all members of the subfamily. PMID- 10852869 TI - A point mutation in the mma3 gene is responsible for impaired methoxymycolic acid production in Mycobacterium bovis BCG strains obtained after 1927. AB - BCG vaccines are substrains of Mycobacterium bovis derived by attenuation in vitro. After the original attenuation (1908 to 1921), BCG strains were maintained by serial propagation in different BCG laboratories (1921 to 1961). As a result, various BCG substrains developed which are now known to differ in a number of genetic and phenotypic properties. However, to date, none of these differences has permitted a direct phenotype-genotype link. Since BCG strains differ in their abilities to synthesize methoxymycolic acids and since recent work has shown that the mma3 gene is responsible for O-methylation of hydroxymycolate precursors to form methoxymycolic acids, we analyzed methoxymycolate production and mma3 gene sequences for a genetically defined collection of BCG strains. We found that BCG strains obtained from the Pasteur Institute in 1927 and earlier produced methoxymycolates in vitro but that those obtained from the Pasteur Institute in 1931 and later all failed to synthesize methoxymycolates, and furthermore, the mma3 sequence of the latter strains differs from that of Mycobacterium tuberculosis H37Rv by a point mutation at bp 293. Site-specific introduction of this guanine-to-adenine mutation into wild-type mma3 (resulting in the replacement of glycine 98 with aspartic acid) eliminated the ability of this enzyme to produce O-methylated mycolic acids when the mutant was cloned in tandem with mma4 into Mycobacterium smegmatis. These findings indicate that a point mutation in mma3 occurred between 1927 and 1931, and that this mutant population became the dominant clone of BCG at the Pasteur Institute. PMID- 10852870 TI - Identification and characterization of two chemotactic transducers for inorganic phosphate in Pseudomonas aeruginosa. AB - Two chemotactic transducers for inorganic phosphate (P(i)), designated CtpH and CtpL, have been identified in Pseudomonas aeruginosa. The corresponding genes (ctpH and ctpL) were inactivated by inserting kanamycin and tetracycline resistance gene cassettes into the wild-type genes in the P. aeruginosa PAO1 genome. Computer-assisted capillary assays showed that the ctpH single mutant failed to exhibit P(i) taxis when the concentration of P(i) in the capillary was higher than 5 mM. Conversely, the ctpL single mutant could not respond to P(i) at the concentration of 0.01 mM. The ctpH ctpL double mutant was defective in P(i) taxis at any concentration ranging from 0.01 to 10 mM. To investigate regulation of P(i) taxis, the ctpH and ctpL genes were also disrupted individually in the P. aeruginosa phoU and phoB single mutants. The ctpH phoU and ctpH phoB double mutants were defective in P(i) taxis, regardless of whether the cells were starved for P(i). The ctpL phoU double mutant was constitutive for P(i) taxis, whereas the ctpL phoB double mutant was induced by P(i) limitation for P(i) taxis. The region upstream of ctpL, but not ctpH, contained a putative pho box sequence. Expression of ctpL::lacZ was induced by P(i) limitation in PAO1, while it was constitutive in the phoU mutant. In contrast, the phoB mutant showed only background levels of ctpL::lacZ expression. These results showed that ctpL is involved in the pho regulon genes in P. aeruginosa. The ctpH phoU mutant, which failed to exhibit P(i) taxis, was constitutive for ctpL::lacZ expression, suggesting that the P(i) detection by CtpL requires PhoU. Like PAO1, the phoB and phoU single mutants were constitutive for expression of ctpH::lacZ. Thus, the evidence that the ctpL phoU mutant, but not the ctpL phoB mutant and PAO1, was constitutive for P(i) taxis raised the possibility that PhoU exerts a negative control on P(i) detection by CtpH at the posttranscriptional level. PMID- 10852871 TI - Vibrio vulnificus has the transmembrane transcription activator ToxRS stimulating the expression of the hemolysin gene vvhA. AB - In an attempt to dissect the virulence regulatory mechanism in Vibrio vulnificus, we tried to identify the V. cholerae transmembrane virulence regulator toxRS (toxRS(Vc)) homologs in V. vulnificus. By comparing the sequences of toxRS of V. cholerae and V. parahaemolyticus (toxRS(Vp)), we designed a degenerate primer set targeting well-conserved sequences. Using the PCR product as an authentic probe for Southern blot hybridization, a 1.6-kb BglII-HindIII fragment and a 1.2-kb HindIII fragment containing two complete open reading frames and one partial open reading frame attributable to toxR(Vv), toxS(Vv), and htpG(Vv) were cloned. ToxR(Vv) shared 55.0 and 63.0% sequence homology with ToxR(Vc) and ToxR(Vp), respectively. ToxS(Vv) was 71.5 and 65.7% homologous to ToxS(Vc) and ToxS(Vp), respectively. The amino acid sequences of ToxRS(Vv) showed transmembrane and activity domains similar to those observed in ToxRS(Vc) and ToxRS(Vp). Western blot analysis proved the expression of ToxR(Vv) in V. vulnificus. ToxRS(Vv) enhanced, in an Escherichia coli background, the expression of the V. vulnificus hemolysin gene (vvhA) fivefold. ToxRS(Vv) also activated the ToxR(Vc)-regulated ctx promoter incorporated into an E. coli chromosome. A toxR(Vv) null mutation decreased hemolysin production. The defect in hemolysin production could be complemented by a plasmid harboring the wild-type gene. The toxR(Vv) mutation also showed a reversed outer membrane protein expression profile in comparison to the isogenic wild-type strain. These results demonstrate that ToxR(Vv) may regulate the virulence expression of V. vulnificus. PMID- 10852872 TI - In Lactobacillus plantarum, carbamoyl phosphate is synthesized by two carbamoyl phosphate synthetases (CPS): carbon dioxide differentiates the arginine-repressed from the pyrimidine-regulated CPS. AB - Carbamoyl phosphate (CP) is an intermediate in pyrimidine and arginine biosynthesis. Carbamoyl-phosphate synthetase (CPS) contains a small amidotransferase subunit (GLN) that hydrolyzes glutamine and transfers ammonia to the large synthetase subunit (SYN), where CP biosynthesis occurs in the presence of ATP and CO(2). Lactobacillus plantarum, a lactic acid bacterium, harbors a pyrimidine-inhibited CPS (CPS-P; Elagoz et al., Gene 182:37-43, 1996) and an arginine-repressed CPS (CPS-A). Sequencing has shown that CPS-A is encoded by carA (GLN) and carB (SYN). Transcriptional studies have demonstrated that carB is transcribed both monocistronically and in the carAB arginine-repressed operon. CP biosynthesis in L. plantarum was studied with three mutants (DeltaCPS-P, DeltaCPS A, and double deletion). In the absence of both CPSs, auxotrophy for pyrimidines and arginine was observed. CPS-P produced enough CP for both pathways. In CO(2) enriched air but not in ordinary air, CPS-A provided CP only for arginine biosynthesis. Therefore, the uracil sensitivity observed in prototrophic wild type L. plantarum without CO(2) enrichment may be due to the low affinity of CPS A for its substrate CO(2) or to regulation of the CP pool by the cellular CO(2)/bicarbonate level. PMID- 10852873 TI - Purification and characterization of a membrane-bound hydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus. AB - Highly washed membrane preparations from cells of the hyperthermophilic archaeon Pyrococcus furiosus contain high hydrogenase activity (9.4 micromol of H(2) evolved/mg at 80 degrees C) using reduced methyl viologen as the electron donor. The enzyme was solubilized with n-dodecyl-beta-D-maltoside and purified by multistep chromatography in the presence of Triton X-100. The purified preparation contained two major proteins (alpha and beta) in an approximate 1:1 ratio with a minimum molecular mass near 65 kDa and contained approximately 1 Ni and 4 Fe atoms/mol. The reduced enzyme gave rise to an electron paramagnetic resonance signal typical of the so-called Ni-C center of mesophilic NiFe hydrogenases. Neither highly washed membranes nor the purified enzyme used NAD(P)(H) or P. furiosus ferredoxin as an electron carrier, nor did either catalyze the reduction of elemental sulfur with H(2) as the electron donor. Using N-terminal amino acid sequence information, the genes proposed to encode the alpha and beta subunits were located in the genome database within a putative 14 gene operon (termed mbh). The deduced sequences of the two subunits (Mbh 11 and 12) were distinctly different from those of the four subunits that comprise each of the two cytoplasmic NiFe-hydrogenases of P. furiosus and show that the alpha subunit contains the NiFe-catalytic site. Six of the open reading frames (ORFs) in the operon, including those encoding the alpha and beta subunits, show high sequence similarity (>30% identity) with proteins associated with the membrane bound NiFe-hydrogenase complexes from Methanosarcina barkeri, Escherichia coli, and Rhodospirillum rubrum. The remaining eight ORFs encode small (<19-kDa) hypothetical proteins. These data suggest that P. furiosus, which was thought to be solely a fermentative organism, may contain a previously unrecognized respiratory system in which H(2) metabolism is coupled to energy conservation. PMID- 10852874 TI - The hydrogenase cytochrome b heme ligands of Azotobacter vinelandii are required for full H(2) oxidation capability. AB - The hydrogenase in Azotobacter vinelandii, like other membrane-bound [NiFe] hydrogenases, consists of a catalytic heterodimer and an integral membrane cytochrome b. The histidines ligating the hemes in this cytochrome b were identified by H(2) oxidation properties of altered proteins produced by site directed mutagenesis. Four fully conserved and four partially conserved histidines in HoxZ were substituted with alanine or tyrosine. The roles of these histidines in HoxZ heme binding and hydrogenase were characterized by O(2) dependent H(2) oxidation and H(2)-dependent methylene blue reduction in vivo. Mutants H33A/Y (H33 replaced by A or Y), H74A/Y, H194A, H208A/Y, and H194,208A lost O(2)-dependent H(2) oxidation activity, H194Y and H136A had partial activity, and H97Y,H98A and H191A had full activity. These results suggest that the fully conserved histidines 33, 74, 194, and 208 are ligands to the hemes, tyrosine can serve as an alternate ligand in position 194, and H136 plays a role in H(2) oxidation. In mutant H194A/Y, imidazole (Imd) rescued H(2) oxidation activity in intact cells, which suggests that Imd acts as an exogenous ligand. The heterodimer activity, quantitatively determined as H(2)-dependent methylene blue reduction, indicated that the heterodimers of all mutants were catalytically active. H33A/Y had wild-type levels of methylene blue reduction, but the other HoxZ ligand mutants had significantly less than wild-type levels. Imd reconstituted full methylene blue reduction activity in mutants H194A/Y and H208A/Y and partial activity in H194,208A. These results indicate that structural and functional integrity of HoxZ is required for physiologically relevant H(2) oxidation, and structural integrity of HoxZ is necessary for full heterodimer catalyzed H(2) oxidation. PMID- 10852875 TI - The N- and C-terminal portions of the Agrobacterium VirB1 protein independently enhance tumorigenesis. AB - Genetic transformation of plants by Agrobacterium tumefaciens is mediated by a virulence (vir)-specific type IV secretion apparatus assembled from 11 VirB proteins and VirD4. VirB1, targeted to the periplasm by an N-terminal signal peptide, is processed to yield VirB1*, comprising the C-terminal 73 amino acids. The N-terminal segment, which shares homology with chicken egg white lysozyme as well as lytic transglycosylases, may provide local lysis of the peptidoglycan cell wall to create channels for transporter assembly. Synthesis of VirB1* followed by its secretion to the exterior of the cell suggests that VirB1* may also have a role in virulence. In the present study, we provide evidence for the dual roles of VirB1 in tumorigenesis as well as the requirements for processing and secretion of VirB1*. Complementation of a virB1 deletion strain with constructs expressing either the N-terminal lysozyme-homologous region or VirB1* results in tumors intermediate in size between those induced by a wild-type strain and a virB1 deletion strain, suggesting that each domain has a unique role in tumorigenesis. The secretion of VirB1* translationally fused to the signal peptide indicates that processing and secretion are not coupled. When expressed independently of all other vir genes, VirB1 was processed and VirB1* was secreted. When restricted to the cytoplasm by deletion of the signal peptide, VirB1 was neither processed nor secreted and did not restore virulence to the virB1 deletion strain. Thus, factors that mediate processing of VirB1 and secretion of VirB1* are localized in the periplasm or outer membrane and are not subject to vir regulation. PMID- 10852876 TI - Control of sporulation gene expression in Bacillus subtilis by the chromosome partitioning proteins Soj (ParA) and Spo0J (ParB). AB - Two chromosome partitioning proteins, Soj (ParA) and Spo0J (ParB), regulate the initiation of sporulation in Bacillus subtilis. In a spo0J null mutant, sporulation is inhibited by the action of Soj. Soj negatively regulates expression of several sporulation genes by binding to the promoter regions and inhibiting transcription. All of the genes known to be inhibited by Soj are also activated by the phosphorylated form of the transcription factor Spo0A (Spo0A approximately P). We found that, in a spo0J null mutant, Soj affected sporulation, in part, by decreasing the level of Spo0A protein. Soj negatively regulated transcription of spo0A and associated with the spo0A promoter region in vivo. Expression of spo0A from a heterologous promoter in a spo0J null mutant restored Spo0A levels and partly bypassed the sporulation and gene expression defects. Soj did not appear to significantly affect phosphorylation of Spo0A. Thus, in the absence of Spo0J, Soj inhibits sporulation and sporulation gene expression by inhibiting accumulation of the activator protein Spo0A and by acting downstream of Spo0A to inhibit gene expression directly. PMID- 10852877 TI - The archaeon Sulfolobus solfataricus contains a membrane-associated protein kinase activity that preferentially phosphorylates threonine residues in vitro. AB - The extreme acidothermophilic archaeon Sulfolobus solfataricus harbors a membrane associated protein kinase activity. Its solubilization and stabilization required detergents, suggesting that this activity resides within an integral membrane protein. The archaeal protein kinase utilized purine nucleotides as phosphoryl donors in vitro. A noticeable preference for nucleotide triphosphates over nucleotide diphosphates and for adenyl nucleotides over the corresponding guanyl ones was observed. The molecular mass of the solubilized, partially purified enzyme was estimated to be approximately 125 kDa by gel filtration chromatography. Catalytic activity resided in a polypeptide with an apparent molecular mass of approximately 67 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Challenges with several exogenous substrates revealed the protein kinase to be relatively selective. Only casein, histone H4, reduced carboxyamidomethylated and maleylated lysozyme, and a peptide modeled after myosin light chains (KKRAARATSNVFA) were phosphorylated to appreciable levels in vitro. All of the aforementioned substrates were phosphorylated on threonine residues, while histone H4 was phosphorylated on serine as well. Substitution of serine for the phosphoacceptor threonine in the myosin light chain peptide produced a noticeably inferior substrate. The protein kinase underwent autophosphorylation on threonine and was relatively insensitive to a set of known inhibitors of "eukaryotic" protein kinases. PMID- 10852878 TI - Analysis of guanine nucleotide binding and exchange kinetics of the Escherichia coli GTPase Era. AB - Era is an essential Escherichia coli guanine nucleotide binding protein that appears to play a number of cellular roles. Although the kinetics of Era guanine nucleotide binding and hydrolysis have been described, guanine nucleotide exchange rates have never been reported. Here we describe a kinetic analysis of guanine nucleotide binding, exchange, and hydrolysis by Era using the fluorescent mant (N-methyl-3'-O-anthraniloyl) guanine nucleotide analogs. The equilibrium binding constants (K(D)) for mGDP and mGTP (0.61 +/- 0. 12 microgM and 3.6 +/- 0.80 microM, respectively) are similar to those of the unmodified nucleotides. The single turnover rates for mGTP hydrolysis by Era were 3.1 +/- 0.2 mmol of mGTP hydrolyzed/min/mol in the presence of 5 mM MgCl(2) and 5.6 +/- 0.3 mmol of mGTP hydrolyzed/min/mol in the presence of 0.2 mM MgCl(2). Moreover, Era associates with and exchanges guanine nucleotide rapidly (on the order of seconds) in both the presence and absence of Mg(2+). We suggest that models of Era function should reflect the rapid exchange of nucleotides in addition to the GTPase activity inherent to Era. PMID- 10852879 TI - Differential expression of over 60 chromosomal genes in Escherichia coli by constitutive expression of MarA. AB - In Escherichia coli, the MarA protein controls expression of multiple chromosomal genes affecting resistance to antibiotics and other environmental hazards. For a more-complete characterization of the mar regulon, duplicate macroarrays containing 4,290 open reading frames of the E. coli genome were hybridized to radiolabeled cDNA populations derived from mar-deleted and mar-expressing E. coli. Strains constitutively expressing MarA showed altered expression of more than 60 chromosomal genes: 76% showed increased expression and 24% showed decreased expression. Although some of the genes were already known to be MarA regulated, the majority were newly determined and belonged to a variety of functional groups. Some of the genes identified have been associated with iron transport and metabolism; other genes were previously known to be part of the soxRS regulon. Northern blot analysis of selected genes confirmed the results obtained with the macroarrays. The findings reveal that the mar locus mediates a global stress response involving one of the largest networks of genes described. PMID- 10852880 TI - Genetic and phenotypic analyses of the rdx locus of Rhodobacter sphaeroides 2.4.1. AB - Previously, we reported that rdxB, encoding a likely membrane-bound two [4Fe-4S] containing center, is involved in the aerobic regulation of photosystem gene expression in Rhodobacter sphaeroides 2.4.1. To further investigate the role of rdxB as well as other genes of the rdxBHIS operon on photosystem gene expression, we constructed a series of nonpolar, in-frame deletion mutations in each of the rdx genes. Using both puc and puf operon lacZ fusions to monitor photosystem gene expression, under aerobic conditions, in each of the mutant strains revealed significant increased photosynthesis gene expression. In the case of mutations in either rdxH, rdxI, or rdxS, the aerobic induction of photosystem gene expression is believed to be indirect by virtue of a posttranscriptional effect on cbb(3) cytochrome oxidase structure and integrity. For RdxB, we suggest that this redox protein has a more direct effect on photosystem gene expression by virtue of its interaction with the cbb(3) oxidase. An associated phenotype, involving the enhanced conversion of the carotenoid spheroidene to spheroidenone, is also observed in the RdxB, -H, -I, and -S mutant strains. This phenotype is also suggested to be the result of the role of the rdxBHIS locus in cbb(3) oxidase activity and/or structure. RdxI is suggested to be a new class of metal transporter of the CPx-type ATPases. PMID- 10852881 TI - The Brucella abortus CcrM DNA methyltransferase is essential for viability, and its overexpression attenuates intracellular replication in murine macrophages. AB - The CcrM DNA methyltransferase of the alpha-proteobacteria catalyzes the methylation of the adenine in the sequence GAnTC. Like Dam in the enterobacteria, CcrM plays a regulatory role in Caulobacter crescentus and Rhizobium meliloti. CcrM is essential for viability in both of these organisms, and we show here that it is also essential in Brucella abortus. Further, increased copy number of the ccrM gene results in striking changes in B. abortus morphology, DNA replication, and growth in murine macrophages. We generated strains that carry ccrM either on a low-copy-number plasmid (strain GR131) or on a moderate-copy-number plasmid (strain GR132). Strain GR131 has wild-type morphology and chromosome number, as assessed by flow cytometry. In contrast, strain GR132 has abnormal branched morphology, suggesting aberrant cell division, and increased chromosome number. Although these strains exhibit different morphologies and DNA content, the replication of both strains in macrophages is attenuated. These data imply that the reduction in survival in host cells is not due solely to a cell division defect but is due to additional functions of CcrM. Because CcrM is essential in B. abortus and increased ccrM copy number attenuates survival in host cells, we propose that CcrM is an appropriate target for new antibiotics. PMID- 10852882 TI - Analysis of the SOS response in Salmonella enterica serovar typhimurium using RNA fingerprinting by arbitrarily primed PCR. AB - We report an analysis of a sample of the SOS response of Salmonella enterica serovar Typhimurium using the differential display of RNA fingerprinting gels of arbitrarily primed PCR products. The SOS response was induced by the addition of mitomycin C to an exponentially growing culture of serovar Typhimurium, and the RNA population was sampled during the following 2 h. These experiments revealed 21 differentially expressed PCR fragments representing mRNA transcripts. These 21 fragments correspond to 20 distinct genes. All of these transcripts were positively regulated, with the observed induction starting 10 to 120 min after addition of mitomycin C. Fifteen of the 21 transcripts have no homologue in the public sequence data banks and are therefore classified as novel. The remaining six transcripts corresponded to the recE, stpA, sulA, and umuC genes, and to a gene encoding a hypothetical protein in the Escherichia coli lysU-cadA intergenic region; the recE gene was represented twice by nonoverlapping fragments. In order to determine if the induction of these 20 transcripts constitutes part of a classical SOS regulon, we assessed the induction of these genes in a recA mutant. With one exception, the increased expression of these genes in response to mitomycin C was dependent on the presence of a functional recA allele. The exception was fivefold induced in the absence of a functional RecA protein, suggesting another layer of regulation in response to mitomycin C, in addition to the RecA-LexA pathway of SOS induction. Our data reveal several genes belonging to operons known to be directly involved in pathogenesis. In addition, we have found several phage-like sequences, some of which may be landmarks of pathogenicity determinants. On the basis of these observations, we propose that the general use of DNA-damaging agents coupled with differential gene expression analysis may be a useful and easy method for identifying pathogenicity determinants in diverse organisms. PMID- 10852883 TI - Virulence of the phytopathogen Pseudomonas syringae pv. maculicola is rpoN dependent. AB - We cloned the rpoN (ntrA and glnF) gene encoding sigma(54) from the phytopathogen Pseudomonas syringae pv. maculicola strain ES4326. The P. syringae ES4326 rpoN gene complemented Pseudomonas aeruginosa, Escherichia coli, and Klebsiella aerogenes rpoN mutants for a variety of rpoN mutant phenotypes, including the inability to utilize nitrate as sole nitrogen source. DNA sequence analysis of the P. syringae ES4326 rpoN gene revealed that the deduced amino acid sequence was most similar (86% identity; 95% similarity) to the sigma(54) protein encoded by the Pseudomonas putida rpoN gene. A marker exchange protocol was used to construct an ES4326 rpoN insertional mutation, rpoN::Km(r). In contrast to wild type ES4326, ES4326 rpoN::Km(r) was nonmotile and could not utilize nitrate, urea, C(4)-dicarboxylic acids, several amino acids, or concentrations of ammonia below 2 mM as nitrogen sources. rpoN was essential for production of the phytotoxin coronatine and for expression of the structural genes encoding coronamic acid. In addition, ES4326 rpoN::Km(r) did not multiply or elicit disease symptoms when infiltrated into Arabidopsis thaliana leaves, did not elicit the accumulation of several Arabidopsis defense-related mRNAs, and did not elicit a hypersensitive response (HR) when infiltrated into tobacco (Nicotiana tabacum) leaves. Furthermore, whereas P. syringae ES4326 carrying the avirulence gene avrRpt2 elicited an HR when infiltrated into Arabidopsis ecotype Columbia leaves, ES4326 rpoN::Km(r) carrying avrRpt2 elicited no response. Constitutive expression of ES4326 hrpL in ES4326 rpoN::Km(r) partially restored defense related mRNA accumulation, showing a direct role for the hrp cluster in host defense gene induction in a compatible host-pathogen interaction. However, constitutive expression of hrpL in ES4326 rpoN::Km(r) did not restore coronatine production, showing that coronatine biosynthesis requires factors other than hrpL. PMID- 10852884 TI - The alternative sigma factor RpoN is required for hrp activity in Pseudomonas syringae pv. maculicola and acts at the level of hrpL transcription. AB - beta-Glucuronidase (uidA) reporter gene fusions were constructed for the hrpZ, hrpL, and hrpS genes from the phytopathogen Pseudomonas syringae pv. maculicola strain ES4326. These reporters, as well as an avrRpt2-uidA fusion, were used to measure transcriptional activity in ES4326 and a ES4326 rpoN mutant. rpoN was required for the expression of avrRpt2, hrpZ, and hrpL in vitro in minimal media and in vivo when infiltrated into Arabidopsis thaliana leaves. In contrast, the expression of hrpS was essentially the same in wild-type and rpoN mutant strains. Constitutive expression of hrpL in an rpoN mutant restored hrpZ transcription to wild-type levels, restored the hypersensitive response when infiltrated into tobacco (Nicotiana tobacum), and partially restored the elicitation of virulence related symptoms but not growth when infiltrated into Arabidopsis leaves. These data indicate that rpoN-mediated control of hrp gene expression acts at the level of hrpL and that in planta growth of P. syringae is not required for the elicitation of disease symptoms. PMID- 10852885 TI - The dual-specificity protein phosphatase Yvh1p regulates sporulation, growth, and glycogen accumulation independently of catalytic activity in Saccharomyces cerevisiae via the cyclic AMP-dependent protein kinase cascade. AB - Yvh1p, a dual-specific protein phosphatase induced specifically by nitrogen starvation, regulates cell growth as well as initiation and completion of sporulation. We demonstrate that yvh1 disruption mutants are also unable to accumulate glycogen in stationary phase. A catalytically inactive variant of yvh1 (C117S) and a DNA fragment encoding only the Yvh1p C-terminal 159 amino acids (which completely lacks the phosphatase domain) complement all three phenotypes as well as the wild-type allele; no complementation occurs with a fragment encoding only the C-terminal 74 amino acids. These observations argue that phosphatase activity is not required for the Yvh1p functions we measured. Mutations which decrease endogenous cyclic AMP (cAMP) levels partially suppress the sporulation and glycogen accumulation defects. In addition, reporter gene expression supported by a DRR2 promoter fragment, containing two stress response elements known to respond to cAMP-protein kinase A, decreases in a yvh1 disruption mutant. Therefore, our results identify three cellular processes that both require Yvh1p and respond to alterations in cAMP, and they lead us to suggest that Yvh1p may be a participant in and/or a contributor to regulation of the cAMP-dependent protein kinase cascade. The fact that decreasing the levels of cAMP alleviates the need for Yvh1p function supports this suggestion. PMID- 10852886 TI - Roles of cyclic AMP receptor protein and the carboxyl-terminal domain of the alpha subunit in transcription activation of the Escherichia coli rhaBAD operon. AB - The Escherichia coli rhaBAD operon encodes the enzymes for catabolism of the sugar L-rhamnose. Full rhaBAD activation requires the AraC family activator RhaS (bound to a site that overlaps the -35 region of the promoter) and the cyclic AMP receptor protein (CRP; bound immediately upstream of RhaS at -92.5). We tested alanine substitutions in activating regions (AR) 1 and 2 of CRP for their effect on rhaBAD activation. Some, but not all, of the substitutions in both AR1 and AR2 resulted in approximately twofold defects in expression from rhaBAD promoter fusions. We also expressed a derivative of the alpha subunit of RNA polymerase deleted for the entire C-terminal domain (alpha-Delta235) and assayed expression from rhaBAD promoter fusions. The greatest defect (54-fold) occurred at a truncated promoter where RhaS was the only activator, while the defect at the full-length promoter (RhaS plus CRP) was smaller (13-fold). Analysis of a plasmid library expressing alanine substitutions at every residue in the carboxyl terminal domain of the alpha subunit (alpha-CTD) identified 15 residues (mostly in the DNA-binding determinant) that were important at both the full-length and truncated promoters. Only one substitution was defective at the full-length but not the truncated promoter, and this residue was located in the DNA-binding determinant. Six substitutions were defective only at the promoter activated by RhaS alone, and these may define a protein-contacting determinant on alpha-CTD. Overall, our results suggest that CRP interaction with alpha-CTD may not be required for rhaBAD activation; however, alpha-CTD does contribute to full activation, probably through interactions with DNA and possibly RhaS. PMID- 10852887 TI - Identification of iron-responsive, differential gene expression in the cyanobacterium Synechocystis sp. strain PCC 6803 with a customized amplification library. AB - We describe the use of a method called differential expression using customized amplification library (DECAL) to study the global changes in gene expression in iron-deficient versus iron-reconstituting cells of Synechocystis sp. strain PCC 6803. We identified a number of genes, such as isiA, idiA, psbA, cpcG, and slr0374, whose expression either increased or decreased in response to iron availability. Further analysis led to the identification of additional genes related to those identified by DECAL (e.g., psbC, psbO, psaA, apcABC, cpcBAC1C2D, and nblA) that were differentially regulated by iron availability. Expression of cpcG, psbC, psbO, psaA, apcABC, and cpcBAC1C2D increased, whereas that of isiA, idiA, nblA, psbA, and slr0374 decreased, in iron-reconstituting cells. S1 nuclease protection studies showed that increased transcript levels of psbA in iron-deficient cells was due to the increased expression of both psbA2 and psbA3 genes, although the steady-state level of psbA2 remained higher than that of psbA3. PMID- 10852888 TI - Isolation and characterization of nonchemotactic CheZ mutants of Escherichia coli. AB - The Escherichia coli CheZ protein stimulates dephosphorylation of CheY, a response regulator in the chemotaxis signal transduction pathway, by an unknown mechanism. Genetic analysis of CheZ has lagged behind biochemical and biophysical characterization. To identify putative regions of functional importance in CheZ, we subjected cheZ to random mutagenesis and isolated 107 nonchemotactic CheZ mutants. Missense mutations clustered in six regions of cheZ, whereas nonsense and frameshift mutations were scattered reasonably uniformly across the gene. Intragenic complementation experiments showed restoration of swarming activity when compatible plasmids containing genes for the truncated CheZ(1-189) peptide and either CheZA65V, CheZL90S, or CheZD143G were both present, implying the existence of at least two independent functional domains in each chain of the CheZ dimer. Six mutant CheZ proteins, one from each cluster of loss-of-function missense mutations, were purified and characterized biochemically. All of the tested mutant proteins were defective in their ability to dephosphorylate CheY-P, with activities ranging from 0.45 to 16% of that of wild-type CheZ. There was good correlation between the phosphatase activity of CheZ and the ability to form large chemically cross-linked complexes with CheY in the presence of the CheY phosphodonor acetyl phosphate. In consideration of both the genetic and biochemical data, the most severe functional impairments in this set of CheZ mutants seemed to be concentrated in regions which are located in a proposed large N-terminal domain of the CheZ protein. PMID- 10852889 TI - A common step for changing cell shape in fruiting body and starvation-independent sporulation of Myxococcus xanthus. AB - Myxococcus xanthus can sporulate in either of two ways: at the end of the program of fruiting body development or after exposure of growing cells to certain reagents such as concentrated glycerol. Fruiting body sporulation requires starvation, while glycerol sporulation requires rapid growth, and since the two types of spores are structurally somewhat different, it has generally been assumed that the two processes are different. However, a Tn5 Lac insertion mutation, Omega7536, has been isolated which simultaneously blocks the development of fruiting body spores as well as glycerol-induced spores. Both sporulation pathways are blocked in the mutant within the process that converts a rod-shaped cell into a spherical spore. The Omega7536 locus is expressed at the time of cell shape change appropriate to each process, early after glycerol induction and late after starvation induction. On the C-signal response pathway, it is possible to identify positions for the normal function of the Omega7536 locus and for the inducing stimulus from glycerol that are unique and consistent with the observations. Although the two sporulation pathways differ in certain respects, it is shown that they share at least one step for changing a rod-shaped cell into a spherical spore. PMID- 10852891 TI - Characterization of devH, a gene encoding a putative DNA binding protein required for heterocyst function in Anabaena sp. strain PCC 7120. AB - The devH gene was identified in a screen for Anabaena sp. strain PCC 7120 sequences whose transcripts increase in abundance during a heterocyst development time course. The product of devH contains a helix-turn-helix motif similar to the DNA binding domain of members of the cyclic AMP receptor protein family, and the protein is most closely related to the cyanobacterial transcriptional activator NtcA. devH transcripts are barely detectable in vegetative cells and are induced approximately fivefold after nitrogen starvation. This induction is absent in the two developmental mutants hetR and ntcA. The gene is expressed as monocistronic transcripts with multiple 5' termini, and the approximately 500-bp region 5' to devH was shown to have promoter activity in vivo. The devH gene was insertionally inactivated by the integration of plasmid sequences within the open reading frame. Nitrogen starvation of the devH mutant induces heterocysts of wild-type morphology, but the mutant is inviable in the absence of fixed nitrogen and unable to reduce acetylene aerobically. PMID- 10852890 TI - Characterization of a Bacteroides mobilizable transposon, NBU2, which carries a functional lincomycin resistance gene. AB - The mobilizable Bacteroides element NBU2 (11 kbp) was found originally in two Bacteroides clinical isolates, Bacteroides fragilis ERL and B. thetaiotaomicron DOT. At first, NBU2 appeared to be very similar to another mobilizable Bacteroides element, NBU1, in a 2.5-kbp internal region, but further examination of the full DNA sequence of NBU2 now reveals that the region of near identity between NBU1 and NBU2 is limited to this small region and that, outside this region, there is little sequence similarity between the two elements. The integrase gene of NBU2, intN2, was located at one end of the element. This gene was necessary and sufficient for the integration of NBU2. The integrase of NBU2 has the conserved amino acids (R-H-R-Y) in the C-terminal end that are found in members of the lambda family of site-specific integrases. This was also the only region in which the NBU1 and NBU2 integrases shared any similarity (28% amino acid sequence identity and 49% sequence similarity). Integration of NBU2 was site specific in Bacteroides species. Integration occurred in two primary sites in B. thetaiotaomicron. Both of these sites were located in the 3' end of a serine-tRNA gene NBU2 also integrated in Escherichia coli, but integration was much less site specific than in B. thetaiotaomicron. Analysis of the sequence of NBU2 revealed two potential antibiotic resistance genes. The amino acid sequences of the putative proteins encoded by these genes had similarity to resistances found in gram-positive bacteria. Only one of these genes was expressed in B. thetaiotaomicron, the homolog of linA, a lincomycin resistance gene from Staphylococcus aureus. To determine how widespread elements related to NBU1 and NBU2 are in Bacteroides species, we screened 291 Bacteroides strains. Elements with some sequence similarity to NBU2 and NBU1 were widespread in Bacteroides strains, and the presence of linA(N) in Bacteroides strains was highly correlated with the presence of NBU2, suggesting that NBU2 has been responsible for the spread of this gene among Bacteroides strains. Our results suggest that the NBU related elements form a large and heterogeneous family, whose members have similar integration mechanisms but have different target sites and differ in whether they carry resistance genes. PMID- 10852892 TI - Megaplasmid pRme2011a of Sinorhizobium meliloti is not required for viability. AB - We report the curing of the 1,360-kb megaplasmid pRme2011a from Sinorhizobium meliloti strain Rm2011. With a positive selection strategy that utilized Tn5B12-S containing the sacB gene, we were able to cure this replicon by successive rounds of selecting for deletion formation in vivo. Subsequent Southern blot, Eckhardt gel, and pulsed-field gel electrophoresis analyses were consistent with the hypothesis that the resultant strain was indeed missing pRme2011a. The cured derivative grew as well as the wild-type strain in both complex and defined media but was unable to use a number of substrates as a sole source of carbon on defined media. PMID- 10852893 TI - The length of the combined 3' untranslated region and poly(A) tail does not control rates of glyceraldehyde-3-phosphate dehydrogenase mRNA translation in three species of parasitic protists. AB - Experimental observations suggested that the length of the glyceraldehyde-3 phosphate dehydrogenase (GAPDH) mRNA 3' end has a role in regulating rates of translation in the parasitic protists Trypanosoma brucei, Leishmania donovani, and Trichomonas vaginalis. Using a PCR assay for poly(A) tail length, we measured the size of the RNA 3' end under different growth conditions in all three species. Our results showed that the combined 3' untranslated region and poly(A) tail of GAPDH mRNA do not vary with different rates of translation. PMID- 10852894 TI - In vivo splicing and functional characterization of Mycobacterium leprae RecA. AB - The RecA proteins from Mycobacterium tuberculosis and Mycobacterium leprae contain inteins. In contrast to the M. tuberculosis RecA, the M. leprae RecA is not spliced in Escherichia coli. We demonstrate here that M. leprae RecA is functionally spliced in Mycobacterium smegmatis and produces resistance toward DNA-damaging agents and homologous recombination. PMID- 10852895 TI - Exopolysaccharide production is required for development of Escherichia coli K-12 biofilm architecture. AB - Although exopolysaccharides (EPSs) are a large component of bacterial biofilms, their contribution to biofilm structure and function has been examined for only a few organisms. In each of these cases EPS has been shown to be required for cellular attachment to abiotic surfaces. Here, we undertook a genetic approach to examine the potential role of colanic acid, an EPS of Escherichia coli K-12, in biofilm formation. Strains either proficient or deficient in colanic acid production were grown and allowed to adhere to abiotic surfaces and were then examined both macroscopically and microscopically. Surprisingly, we found that colanic acid production is not required for surface attachment. Rather, colanic acid is critical for the formation of the complex three-dimensional structure and depth of E. coli biofilms. PMID- 10852896 TI - Cryptic and exposed invariable regions of VlsE, the variable surface antigen of Borrelia burgdorferi sl. AB - Borrelia burgdorferi, the Lyme disease spirochete, possesses a surface protein, VlsE, which undergoes antigenic variation. VlsE contains two invariable domains and a variable one that includes six variable and six invariable regions (IRs). Five of the IRs are conserved among strains and genospecies of B. burgdorferi sensu lato. IR(6) is conserved, immunodominant, and exposed at the VlsE surface but not at the spirochete surface, as assessed in vitro. In the present study, the remaining conserved IRs (IR(2) to IR(5)) were investigated. Antisera to synthetic peptides based on each of the IR(2) to IR(5) sequences were produced in rabbits. Antipeptide antibody titers were similarly high in all antisera. Native VlsE was immunoprecipitable with antibodies to IR(2), IR(4), and IR(5) but not to IR(3), indicating that the first three sequences were exposed at the VlsE surface. However, negative surface immunofluorescence and in vitro antibody mediated killing results indicated that none of the IRs were accessible to antibody at the spirochetal surface in vitro. PMID- 10852897 TI - First evidence for existence of an uphill electron transfer through the bc(1) and NADH-Q oxidoreductase complexes of the acidophilic obligate chemolithotrophic ferrous ion-oxidizing bacterium Thiobacillus ferrooxidans. AB - The energy-dependent electron transfer pathway involved in the reduction of pyridine nucleotides which is required for CO(2) fixation to occur in the acidophilic chemolithotrophic organism Thiobacillus ferrooxidans was investigated using ferrocytochrome c as the electron donor. The experimental results show that this uphill pathway involves a bc(1) and an NADH-Q oxidoreductase complex functioning in reverse, using an electrochemical proton gradient generated by ATP hydrolysis. Based on these results, a model is presented to explain the balance of the reducing equivalent from ferrocytochrome c between the exergonic and endergonic electron transfer pathways. PMID- 10852898 TI - Septal localization of the membrane-bound division proteins of Bacillus subtilis DivIB and DivIC is codependent only at high temperatures and requires FtsZ. AB - Using immunofluorescence microscopy, we have examined the dependency of localization among three Bacillus subtilis division proteins, FtsZ, DivIB, and DivIC, to the division site. DivIC is required for DivIB localization. However, DivIC localization is dependent on DivIB only at high growth temperatures, at which DivIB is essential for division. FtsZ localization is required for septal recruitment of DivIB and DivIC, but FtsZ can be recruited independently of DivIB. These localization studies suggest a more specific role for DivIB in division, involving interaction with DivIC. PMID- 10852899 TI - A novel anti-tumor cytokine contains an RNA binding motif present in aminoacyl tRNA synthetases. AB - Endothelial monocyte-activating polypeptide II (EMAP II) is a novel pro-apoptotic cytokine that shares sequence homology with the C-terminal regions of several tRNA synthetases. Pro-EMAP II, the precursor of EMAP II, is associated with the multi-tRNA synthetase complex and facilitates aminoacylation activity. The structure of human EMAP II, solved at 1.8 A resolution, revealed the oligomer binding fold for binding different tRNAs and a domain that is structurally homologous to other chemokines. The similar structures to the RNA binding motif of EMAP II was previously observed in the anticodon binding domain of yeast Asp tRNA synthetase (AspRSSC) and the B2 domain of Thermus thermophilus Phe-tRNA synthetase. The RNA binding pattern of EMAP II is likely to be nonspecific, in contrast to the AspRSSC. The peptide sequence that is responsible for cytokine activity is located, for the most part, in the beta1 strand. It is divided into two regions by a neighboring loop. PMID- 10852900 TI - Natural animal coloration can Be determined by a nonfluorescent green fluorescent protein homolog. AB - It is generally accepted that the colors displayed by living organisms are determined by low molecular weight pigments or chromoproteins that require a prosthetic group. The exception to this rule is green fluorescent protein (GFP) from Aequorea victoria that forms a fluorophore by self-catalyzed protein backbone modification. Here we found a naturally nonfluorescent homolog of GFP to determine strong purple coloration of tentacles in the sea anemone Anemonia sulcata. Under certain conditions, this novel chromoprotein produces a trace amount of red fluorescence (emission lambda(max) = 595 nm). The fluorescence demonstrates unique behavior: its intensity increases in the presence of green light but is inhibited by blue light. The quantum yield of fluorescence can be enhanced dramatically by single amino acid replacement, which probably restores the ancestral fluorescent state of the protein. Other fluorescent variants of the novel protein have emission peaks that are red-shifted up to 610 nm. They demonstrate that long wavelength fluorescence is attainable in GFP-like fluorescent proteins. PMID- 10852901 TI - Defective heparan sulfate biosynthesis and neonatal lethality in mice lacking N deacetylase/N-sulfotransferase-1. AB - Heparan sulfate is a sulfated polysaccharide present on most cell surfaces and in the extracellular matrix. In vivo functions of heparan sulfate can be studied in mouse strains lacking enzymes involved in the biosynthesis of heparan sulfate. Glucosaminyl N-deacetylase/N-sulfotransferase (NDST) catalyzes the first modifying step in the biosynthesis of the polysaccharide. This bifunctional enzyme occurs in several isoforms. We here report that targeted gene disruption of NDST-1 in the mouse results in a structural alteration of heparan sulfate in most basement membranes as revealed by immunohistochemical staining of fetal tissue sections using antibodies raised against heparan sulfate. Biochemical analysis of heparan sulfate purified from fibroblast cultures, lung, and liver of NDST-1-deficient embryos demonstrated a dramatic reduction in N-sulfate content. Most NDST-1-deficient embryos survive until birth; however, they turn out to be cyanotic and die neonatally in a condition resembling respiratory distress syndrome. In addition, a minor proportion of NDST-1-deficient embryos die during the embryonic period. The cause of the embryonic lethality is still obscure, but incompletely penetrant defects of the skull and the eyes have been observed. PMID- 10852902 TI - The stabilities of mammalian apomyoglobins vary over a 600-fold range and can be enhanced by comparative mutagenesis. AB - Apomyoglobins from 13 different mammals were examined for resistance to denaturation by guanidinium chloride. Unfolding was followed by circular dichroism and tryptophan fluorescence and analyzed globally using the two-step, three-state mechanism first described by Barrick and Baldwin (Barrick, D., and Baldwin, R. L. (1993) Biochemistry 32, 3790-3796). With one exception, the rise and fall of Trp fluorescence intensity correlates quantitatively with the native to intermediate to unfolded steps seen in the CD curves. Although the O(2) binding properties of the holoproteins are nearly identical, the unfolding transitions of the apomyoglobins show 600-fold differences in resistance to guanidinium chloride denaturation. Apomyoglobins from diving mammals, particularly from sperm whales, are the most stable, whereas the apoproteins from pig, horse, and sheep are the least stable, indicating selective pressure for resistance to denaturation in the whale proteins. Sequence comparisons suggest that the key stabilizing residues in whale globins are Ala(5), His(12), Ile(28), Thr(51), Ala(53), Ala(74), Lys(87), Lys(140), and Ile(142). Combinations of these residues were substituted into pig myoglobin. The resultant multiple mutants showed stabilities approaching that of recombinant sperm whale apomyoglobin. Thus, comparative mutagenesis can be used to increase heme protein stability and improve expression yields in bacteria without compromising function. PMID- 10852903 TI - Autocrine action and its underlying mechanism of nitric oxide on intracellular Ca2+ homeostasis in vascular endothelial cells. AB - The rise in cytosolic Ca(2+) concentration (Ca(2+)(i)) in vascular endothelial cells (ECs) activates the production and release of nitric oxide (NO). NO modifies Ca(2+)(i) homeostasis in many types of nonendothelial cells. However, its effect on endothelial Ca(2+)(i) homeostasis at basal and excited states remains unclear. In the present study, to elucidate the effect of NO on basal Ca(2+)(i), inositol 1,4,5-trisphosphate-induced Ca(2+)(i) release (IICR) was blocked by expressing an antisense against type-1 inositol 1,4,5-trisphosphate receptors or by microinjecting heparin to individual ECs, and the effects of NO that was released by and diffused from adjacent IICR-intact ECs were recorded. After ATP or bradykinin stimulation, IICR-inhibited ECs showed a marked reduction of basal Ca(2+)(i), which was abolished by N(G)-monomethyl-l-arginine monoacetate pretreatment. The reduction disappeared in sparsely seeded ECs. Exogenous NO gas mimicked the effect of ATP or bradykinin to reduce basal Ca(2+)(i). Blocking plasma membrane Ca(2+)-ATPase (PMCA), but not Na(+)-Ca(2+) exchange or sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase, suppressed the reduction, indicating that the reduction resulted from a NO-dependent potentiation of PMCA. To elucidate the effect of NO on elevated Ca(2+)(i), ATP-, bradykinin-, or thapsigargin-evoked Ca(2+)(i) response in the presence and absence of NO production was compared in adjacent IICR-intact ECs. NO was found to potentiate PMCA, which, in turn, greatly attenuated agonist-evoked Ca(2+)(i) elevation. NO also potentiated Ca(2+) influx, which markedly increased the sustained phase of Ca(2+)(i) elevation and possibly NO production. NO did not affect other Ca(2+)(i) elevating and Ca(2+)(i)-sequestrating components. Thus, NO-dependent potentiation of PMCA is crucial for Ca(2+)(i) homeostasis over a wide Ca(2+)(i) range. PMID- 10852904 TI - Retention of the human Rad9 checkpoint complex in extraction-resistant nuclear complexes after DNA damage. AB - Studies in yeasts and mammals have identified many genes important for DNA damage induced checkpoint activation, including Rad9, Hus1, and Rad1; however, the functions of these gene products are unknown. In this study we show by immunolocalization that human Rad9 (hRad9) is localized exclusively in the nucleus. However, hRad9 was readily released from the nucleus into the soluble extract upon biochemical fractionation of un-irradiated cells. In contrast, DNA damage promptly converted hRad9 to an extraction-resistant form that was retained at discrete sites within the nucleus. Conversion of hRad9 to the extraction resistant nuclear form occurred in response to diverse DNA-damaging agents and the replication inhibitor hydroxyurea but not other cytotoxic stimuli. Additionally, extraction-resistant hRad9 interacted with its binding partners, hHus1 and an inducibly phosphorylated form of hRad1. Thus, these studies demonstrate that hRad9 is a nuclear protein that becomes more firmly anchored to nuclear components after DNA damage, consistent with a proximal function in DNA damage-activated checkpoint signaling pathways. PMID- 10852905 TI - Lectins from tropical sponges. Purification and characterization of lectins from genus Aplysina. AB - Only a few animal phyla have been screened for the presence and distribution of lectins. Probably the most intensively studied group is the mollusk. In this investigation, 22 species from 12 families of tropical sponges collected in Los Roques National Park (Venezuela) were screened for the presence of lectins. Nine saline extracts exhibited strong hemagglutinating activity against pronase treated hamster red blood cells; five of these reacted against rabbit red blood cells, four with trypsin-treated bovine red blood cells, and five with human red blood cells regardless of the blood group type. Extracts from the three species studied from genus Aplysina (archeri, lawnosa, and cauliformis) were highly reactive and panagglutinating against the panel of red blood cells tested. The lectins from A. archeri and A. lawnosa were purified to homogeneity by ammonium sulfate fractionation, affinity chromatography on p-aminobenzyl-beta-1 thiogalactopyranoside-agarose, and gel filtration chromatography. Both lectins exhibited a native molecular mass of 63 kDa and by SDS-polyacrylamide gel electrophoresis under reducing conditions have an apparent molecular mass of 16 kDa, thus suggesting they occur as homotetramers. The purified lectins contain 3 4 mol of divalent cation per molecule, which are essential for their biological activity. Hapten inhibition of hemagglutination was carried out to define the sugar binding specificity of the purified A. archeri lectin. The results indicate a preference of the lectin for nonreducing beta-linked d-Gal residues being the best inhibitors of red blood cells binding methyl-beta-d-Gal and thiodigalactoside (Gal beta 1-4-thiogalactopyranoside). The behavior of several glycans on immobilized lectin affinity chromatography confirmed and extended the specificity data obtained by hapten inhibition. PMID- 10852906 TI - Adenylyl cyclase regulates signal onset via the inhibitory GTP-binding protein, Gi. AB - Adenylyl cyclase, the enzyme that converts ATP to cAMP, is regulated by its stimulatory and inhibitory GTP-binding proteins, G(s) and G(i), respectively. Recently, we demonstrated that besides catalyzing the synthesis of cAMP, type V adenylyl cyclase (ACV) can act as a GTPase-activating protein for Galpha(s) and also enhance the ability of activated receptors to stimulate GTP-GDP exchange on heterotrimeric G(s) (Scholich, K., Mullenix, J. B., Wittpoth, C., Poppleton, H. M., Pierre, S. C., Lindorfer, M. A., Garrison, J. C., and Patel, T. B. (1999) Science 283, 1328-1331). This latter action of ACV would facilitate the rapid onset of signaling via G(s). Because the C1 region of ACV interacts with the inhibitory GTP-binding protein Galpha(i), we investigated whether the receptor mediated activation of heterotrimeric G(i) was also regulated by ACV and its subdomains. Our data show that ACV and its C1 domain increased the ability of a muscarinic receptor mimetic peptide (MIII-4) to enhance activation of heterotrimeric G(i) such that the amount of peptide required to stimulate G(i) in steady-state GTPase activity assays was 3-4 orders of magnitude less than without the C1 domain. Additionally, the MIII-4-mediated binding of guanosine 5'-(gamma thio)triphosphate (GTPgammaS) to G(i) was also markedly increased in the presence of ACV or its C1 domain. In contrast, the C2 domain of ACV was not able to alter either the GTPase activity or the GTPgammaS binding to G(i) in the presence of MIII-4. Furthermore, in adenylyl cyclase assays employing S49 cyc(-) cell membranes, the C1 (but not the C2) domain of ACV enhanced the ability of peptide MIII-4 as well as endogenous somatostatin receptors to activate endogenous G(i) and to inhibit adenylyl cyclase activity. These data demonstrate that adenylyl cyclase and its C1 domain facilitate receptor-mediated activation of G(i). PMID- 10852907 TI - Inhibition of myogenesis by depletion of the glycogen-associated regulatory subunit of protein phosphatase-1 in rat skeletal muscle cells. AB - In this study, we examined the role of the glycogen-associated regulatory subunit of protein phosphatase-1 (PP-1(G)) in L6 rat skeletal muscle cell myogenesis. The level of PP-1(G) was depleted by transfection with an inducible antisense oriented PP-1(G) gene. Western blot analysis of the PP-1(G)-depleted cell line revealed a >90% depletion of PP-1(G) protein and a 45% reduction in cellular PP-1 activity and abolished the ability of L6 myoblasts to differentiate into multinucleated myotubes. PP-1(G)-depleted cells also exhibited a marked reduction in the expression of the differentiation marker myogenin as well as creatine kinase. After 7 days in culture, PP-1(G)-depleted cells sustained myoblast levels of inhibitor of differentiation-2, whereas control L6 cells had a severely lower inhibitor of differentiation-2 level and progressed into myotubes. Myoblasts were unable to exit the cell cycle, as measured by the impaired induction of p27 cyclin-dependent kinase inhibitor, a >2-fold increase in DNA synthesis, and elevated levels of phosphorylated retinoblastoma protein (pRb). Replacement of the PP-1(G) gene restored PP-1(G) protein expression, PP-1 enzymatic activity, and the ability to differentiate into myotubes. We conclude that PP-1(G) plays a definite role in L6 myogenesis via its regulation of PP-1 catalytic activity. PMID- 10852908 TI - Characterization of the Vibrio parahaemolyticus Na+/Glucose cotransporter. A bacterial member of the sodium/glucose transporter (SGLT) family. AB - The Vibrio parahaemolyticus sodium/glucose transporter (vSGLT) is a bacterial member of the SGLT gene family. Wild-type and mutant vSGLT proteins were expressed in Escherichia coli, and transport activity was measured in intact cells and plasma membrane vesicles. Two cysteine-less vSGLT proteins exhibited sugar transport rates comparable with that of the wild-type protein. Six residues in two regions of vSGLT known to be of functional importance in SGLT1 were replaced individually with cysteine in the cysteine-less protein. Characterization of these single cysteine-substituted vSGLTs showed that two residues (Gly-151 and Gln-428) are essential for transport function, whereas the other four residues (Leu-147, Leu-149, Ala-423, and Gln-425) are not. 2 Aminoethylmethanethiosulfonate (MTSEA) blocked Na(+)/glucose transport by only the transporter bearing a cysteine at position 425 (Q425C). MTSEA inhibition was reversed by dithiothreitol and blocked by the presence of both Na(+) and d glucose, indicating that conformational changes of the vSGLT protein are involved in Na(+)/glucose transport. A split version of vSGLT was generated by co expression of the N-terminal (N(7)) and C-terminal (C(7)) halves of the transporter. The split vSGLT maintained Na(+)-dependent glucose transport activity. Chemical cross-linking of split vSGLT, with a cysteine in each N(7) and C(7) fragment, suggested that hydrophilic loops between helices 4 and 5 and between helices 10 and 11 are within 8 A of each other. We conclude that the mechanism of Na(+)/glucose transport by vSGLT is similar to mammalian SGLTs and that further studies on vSGLT will provide novel insight to the structure and function of this class of cotransporters. PMID- 10852909 TI - Mapping the domain of troponin T responsible for the activation of actomyosin ATPase activity. Identification of residues involved in binding to actin. AB - The in vitro Ca(2+) regulation of the actomyosin Mg(2+)-ATPase at physiological ratios of actin, tropomyosin, and troponin occurs only in the presence of troponin T. We have previously demonstrated that a polypeptide corresponding to the first 191 amino acids of troponin T (TnT-(1-191)) activates the actomyosin Mg(2+)-ATPase in the presence of tropomyosin. In order to further characterize this activation domain, we constructed troponin T fragments corresponding to residues 1-157 (TnT-(1-157)), 1-76 (TnT-(1-76)), 77-157 (TnT-(77-157)), 77-191 (TnT-(77-191)), and 158-191 (TnT-(158-191)). Assays using these fragments demonstrated the following: (a) residues 1-76 do not bind to tropomyosin or actin; (b) residues 158-191 bind to actin cooperatively but not to tropomyosin; (c) the sequence 77-157 is necessary for troponin interaction with residue 263 of tropomyosin; (d) TnT-(77-191) on its own activates the actomyosin ATPase activity as described previously for TnT-(1-191). TnT-(1-157), TnT-(1-76), TnT-(77-157), TnT-(158-191), and combinations of TnT-(158-191) with TnT-(1-157) or TnT-(77-157) showed no effect on the ATPase activity. We conclude that the activation of actomyosin ATPase activity is mediated by a direct interaction between amino acids 77 and 191 of troponin T, tropomyosin, and actin. PMID- 10852910 TI - Characterization of p26olf, a novel calcium-binding protein in the frog olfactory epithelium. AB - We have previously shown that p26olf is a novel S100-like Ca(2+)-binding protein in the frog olfactory epithelium. In this paper, we characterized the Ca(2+) binding property of p26olf, examined the precise localization in the frog olfactory epithelium, and searched for the possible target proteins of p26olf. By flow dialysis experiments using (45)Ca, p26olf was suggested to bind approximately 4 Ca(2+). Circular dichroism measurement showed that binding of Ca(2+) to p26olf induces an increase in the apparent content of both alpha-helix and beta-sheet with an apparent K(d) value of 2.4 micrometer. Electron microscopic observation disclosed p26olf immunoreactivity in the cilia, dendritic knob, and dendrite of olfactory receptor cells. Blot overlay analysis and affinity purification of p26olf-binding proteins showed that p26olf binds to a frog beta-adrenergic receptor kinase-like protein in a Ca(2+)-dependent manner. These results suggested that p26olf has some roles in the olfactory transduction or adaptation. PMID- 10852911 TI - Muscarinic acetylcholine receptors induce the expression of the immediate early growth regulatory gene CYR61. AB - In brain, muscarinic acetylcholine receptors (mAChRs) modulate neuronal functions including long term potentiation and synaptic plasticity in neuronal circuits that are involved in learning and memory formation. To identify mAChR-inducible genes, we used a differential display approach and found that mAChRs rapidly induced transcription of the immediate early gene CYR61 in HEK 293 cells with a maximum expression after 1 h of receptor stimulation. CYR61 is a member of the emerging CCN gene family that includes CYR61/CEF10, CTGF/FISP-12, and NOV; these encode secretory growth regulatory proteins with distinct functions in cell proliferation, migration, adhesion, and survival. We found that CYR61, CTGF, and NOV were expressed throughout the human central nervous system. Stimulation of mAChRs induced CYR61 expression in primary neurons and rat brain where CYR61 mRNA was detected in cortical layers V and VI and in thalamic nuclei. In contrast, CTGF and NOV expression was not altered by mAChRs neither in neuronal tissue culture nor rat brain. Receptor subtype analyses demonstrated that m1 and m3 mAChR subtypes strongly induced CYR61 expression, whereas m2 and m4 mAChRs had only subtle effects. Increased CYR61 expression was coupled to mAChRs by both protein kinase C and elevations of intracellular Ca(2+). Our results establish that CYR61 expression in mammalian brain is under the control of cholinergic neurotransmission; it may thus be involved in cholinergic regulation of synaptic plasticity. PMID- 10852912 TI - Oxidative stress promotes specific protein damage in Saccharomyces cerevisiae. AB - We have analyzed the proteins that are oxidatively damaged when Saccharomyces cerevisiae cells are exposed to stressing conditions. Carbonyl groups generated by hydrogen peroxide or menadione on proteins of aerobically respiring cells were detected by Western blotting, purified, and identified. Mitochondrial proteins such as E2 subunits of both pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase, aconitase, heat-shock protein 60, and the cytosolic fatty acid synthase (alpha subunit) and glyceraldehyde-3-phosphate dehydrogenase were the major targets. In addition we also report the in vivo modification of lipoamide present in the above-mentioned E2 subunits under the stressing conditions tested and that this also occurs with the homologous enzymes present in Escherichia coli cells that were used for comparative analysis. Under fermentative conditions, the main protein targets in S. cerevisiae cells treated with hydrogen peroxide or menadione were pyruvate decarboxylase, enolase, fatty acid synthase, and glyceraldehyde-3-phosphate dehydrogenase. Under the stress conditions tested, fermenting cells exhibit a lower viability than aerobically respiring cells and, consistently, increased peroxide generation as well as higher content of protein carbonyls and lipid peroxides. Our results strongly suggest that the oxidative stress in prokaryotic and eukaryotic cells shares common features. PMID- 10852913 TI - Characterization of human RhCG and mouse Rhcg as novel nonerythroid Rh glycoprotein homologues predominantly expressed in kidney and testis. AB - In mammals, the Rh family includes the variable Rh polypeptides and invariant RhAG glycoprotein. These polytopic proteins are confined to the erythroid lineage and are assembled into a multisubunit complex essential for Rh antigen expression and plasma membrane integrity. Here, we report the characterization of RhCG and Rhcg, a pair of novel Rh homologues present in human and mouse nonerythroid tissues. Despite sharing a notable similarity to the erythroid forms, including the 12-transmembrane topological fold, the RHCG/Rhcg pair is distinct in chromosome location, genomic organization, promoter structure, and tissue specific expression. RHCG and Rhcg map at 15q25 of human chromosome 15 and the long arm of mouse chromosome 7, respectively, each having 11 exons and a CpG-rich promoter. Northern blots detected kidney and testis as the major organs of RHCG or Rhcg expression. In situ hybridization revealed strong expression of Rhcg in the kidney collecting tubules and testis seminiferous tubules. Confocal imaging of transiently expressed green fluorescence protein fusion proteins localized RhCG exclusively to the plasma membrane, a distribution confirmed by cellular fractionation and Western blot analysis. In vitro translation and ex vivo expression showed that RhCG carries a complex N-glycan, probably at the (48)NLS(50) sequon of exoloop 1. These results pinpoint RhCG and Rhcg as novel polytopic membrane glycoproteins that may function as epithelial transporters maintaining normal homeostatic conditions in kidney and testis. PMID- 10852914 TI - Gamma-interferon-inducible lysosomal thiol reductase (GILT). Maturation, activity, and mechanism of action. AB - We recently identified a gamma-interferon-inducible lysosomal thiol reductase (GILT), constitutively expressed in antigen-presenting cells, that catalyzes disulfide bond reduction both in vitro and in vivo and is optimally active at acidic pH. GILT is synthesized as a 35-kDa precursor, and following delivery to major histocompatibility complex (MHC) class II-containing compartments (MIICs), is processed to the mature 30-kDa form via cleavage of N- and C-terminal propeptides. The generation of MHC class II epitopes requires both protein denaturation and reduction of intra- and inter-chain disulfide bonds prior to proteolysis. GILT may be important in disulfide bond reduction of proteins delivered to MIICs and consequently in antigen processing. In this report we show that, like its mature form, precursor GILT reduces disulfide bonds with an acidic pH optimum, suggesting that it may also be involved in disulfide bond reduction in the endocytic pathway. We also show that processing of precursor GILT can be mediated by multiple lysosomal proteases and provide evidence that the mechanism of action of GILT resembles that of other thiol oxidoreductases. PMID- 10852915 TI - Farnesyl transferase inhibitors block the farnesylation of CENP-E and CENP-F and alter the association of CENP-E with the microtubules. AB - Human tumor cell lines that are sensitive to the effects of farnesyl transferase inhibitors accumulate in G(2) --> M (except for cells with an activated Ha-ras that accumulate in G(1)). A search for CAAX box proteins from Swiss-Prot revealed more than 300 peptides. Of these, the centromeric proteins CENP-E and CENP-F are preferentially expressed during mitosis and are implicated as mediators of the G(2) --> M checkpoint. Experiments performed here show that peptides from the COOH-terminal CAAX box of CENP-E and CENP-F are substrates for farnesyl transferase but not geranylgeranyl transferase-I. Although both proteins are prenylated in the human tumor cell line DLD-1, their prenylation is completely inhibited by the farnesyl transferase inhibitor, SCH 66336. Immunohistochemical data with the lung carcinoma cell line, A549, showed that preventing the farnesylation of CENP-E and CENP-F by treatment with the farnesyl transferase inhibitor SCH 66336 does not affect their localization to the kinetochores. However, the presence of farnesyl transferase inhibitors alters the association between CENP-E and the microtubules. Our results imply that the inhibition of CENP-E farnesylation results in the alteration of the microtubule-centromere interaction during mitosis and results in the accumulation of cells prior to metaphase. PMID- 10852916 TI - Synucleins are a novel class of substrates for G protein-coupled receptor kinases. AB - G protein-coupled receptor kinases (GRKs) specifically recognize and phosphorylate the agonist-occupied form of numerous G protein-coupled receptors (GPCRs), ultimately resulting in desensitization of receptor signaling. Until recently, GPCRs were considered to be the only natural substrates for GRKs. However, the recent discovery that GRKs also phosphorylate tubulin raised the possibility that additional GRK substrates exist and that the cellular role of GRKs may be much broader than just GPCR regulation. Here we report that synucleins are a novel class of GRK substrates. Synucleins (alpha, beta, gamma, and synoretin) are 14-kDa proteins that are highly expressed in brain but also found in numerous other tissues. alpha-Synuclein has been linked to the development of Alzheimer's and Parkinson's diseases. We found that all synucleins are GRK substrates, with GRK2 preferentially phosphorylating the alpha and beta isoforms, whereas GRK5 prefers alpha-synuclein as a substrate. GRK-mediated phosphorylation of synuclein is activated by factors that stimulate receptor phosphorylation, such as lipids (all GRKs) and Gbetagamma subunits (GRK2/3), suggesting that GPCR activation may regulate synuclein phosphorylation. GRKs phosphorylate synucleins at a single serine residue within the C-terminal domain. Although the function of synucleins remains largely unknown, recent studies have demonstrated that these proteins can interact with phospholipids and are potent inhibitors of phospholipase D2 (PLD2) in vitro. PLD2 regulates the breakdown of phosphatidylcholine and has been implicated in vesicular trafficking. We found that GRK-mediated phosphorylation inhibits synuclein's interaction with both phospholipids and PLD2. These findings suggest that GPCRs may be able to indirectly stimulate PLD2 activity via their ability to regulate GRK-promoted phosphorylation of synuclein. PMID- 10852917 TI - Purification and kinetic analysis of eIF2B from Saccharomyces cerevisiae. AB - Eukaryotic translation initiation factor 2B (eIF2B) is the heteropentameric guanine nucleotide exchange factor for translation initiation factor 2 (eIF2). Recent studies in the yeast Saccharomyces cerevisiae have served to characterize genetically the exchange factor. However, enzyme kinetic studies of the yeast enzyme have been hindered by the lack of sufficient quantities of protein suitable for biochemical analysis. We have purified yeast eIF2B and characterized its catalytic properties in vitro. Values for K(m) and V(max) were determined to be 12.2 nm and 250.7 fmol/min, respectively, at 0 degrees C. The calculated turnover number (K(cat)) of 43.2 pmol of GDP released per min/pmol of eIF2B at 30 degrees C is approximately 1 order of magnitude lower than values previously reported for the mammalian factor. Reciprocal plots at varying fixed concentrations of the second substrate were linear and intersected to the left of the y axis. This is consistent with a sequential catalytic mechanism and argues against a ping-pong mechanism similar to that proposed for EF-Tu/EF-Ts. In support of this model, our yeast eIF2B preparations bind guanine nucleotides, with an apparent dissociation constant for GTP in the low micromolar range. PMID- 10852918 TI - Activation of Ca2+/calmodulin-dependent protein kinase II within post-synaptic dendritic spines of cultured hippocampal neurons. AB - To understand the cell signaling of protein kinases, it is essential to monitor their activity in each of the subcellular compartments. Here we developed a method to visualize the activities of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) in the cytoplasm, plasma membrane, and nucleus, separately, by utilizing targeted phosphorylation motifs and phosphorylation-specific antibodies. This approach was used to monitor the activities of post-synaptic CaMKII in cultured hippocampal neurons. Strong stimulation of the neurons by N methyl-d-aspartate led to global activations of CaMKII in the cell bodies and dendrites. On the other hand, weak stimulation by removal of Mg(2+) block of N methyl-d-aspartate receptors induced CaMKII signaling localized within single dendritic spines. Post-synaptic CaMKII is thought to modify synaptic efficiency. The present data for the first time demonstrate the activation of CaMKII localized within single dendritic spines and are consistent with the notion that synaptic efficiency is modified by CaMKII in single or multiple spine level depending on the strength of receptor activation. PMID- 10852919 TI - Protein kinase C evokes quantal catecholamine release from PC12 cells via activation of L-type Ca2+ channels. AB - Application of the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) to PC12 cells under resting conditions evoked quantal catecholamine secretion, as detected amperometrically. This effect was not mimicked by 4alpha-phorbol-12,13 didecanoate, another phorbol ester, which is inactive with respect to protein kinase C activation, and was prevented by the protein kinase C inhibitor bisindolylmaleimide. TPA also caused a rise of [Ca(2+)](i) in Fura-2-loaded PC12 cells, and again this was not mimicked by 4alpha-phorbol-12,13-didecanoate and could be blocked by bisindolylmaleimide. TPA-evoked secretion was entirely dependent on extracellular Ca(2+) and was fully abolished by nifedipine, as were TPA-induced rises of [Ca(2+)](i). Resting membrane potential, monitored using perforated patch recordings, was unaffected by TPA. However, a small (6-8 mV) hyperpolarizing shift in the voltage dependence of Ca(2+) channel currents (determined using whole-cell patch clamp recordings) was induced by TPA, and this could be fully prevented by nifedipine. In contrast to results with depolarizing stimuli, which evoke exocytosis because of Ca(2+) influx through N-type channels in these cells, the present results indicate that protein kinase C activation leads directly to quantal catecholamine secretion in the absence of depolarizing stimuli via a selective shift in the activation of L-type Ca(2+) channels. PMID- 10852920 TI - Schwann cells synthesize type V collagen that contains a novel alpha 4 chain. Molecular cloning, biochemical characterization, and high affinity heparin binding of alpha 4(V) collagen. AB - Previously, we reported the isolation of a heparan sulfate-binding collagenous protein, p200, that is expressed by Schwann cells in developing peripheral nerves ((1996) J. Biol. Chem. 271, 13844-13853; (1999) J. Neurosci. Res. 56, 284-294). Here, we report the cloning of p200 cDNA from a Schwann cell cDNA library. The deduced amino acid sequence identifies p200 as a novel member of the collagen type V gene family. This polypeptide, which we have named alpha4 type V (alpha4(V)) collagen, contains an uninterrupted Gly-X-X collagen domain of 1011 amino acids that shows 82% sequence identity to human alpha3(V) collagen and 71% identity to rat alpha1(V) collagen. alpha4(V) is secreted by Schwann cells as a collagen heterotrimer that also contains alpha1(V) chains. alpha4(V)-containing collagen molecules synthesized by Schwann cells retain their amino-terminal non collagenous domains. alpha4(V) mRNA was detected by reverse transcriptase-linked polymerase chain reaction amplification in neonatal and adult brain and neonatal peripheral nerve. alpha4(V) mRNA and protein were not detected in most other tissues, including the placenta and heart, which are known to contain alpha3(V). This pattern of alpha4(V) expression contrasted with that of alpha1(V) mRNA and protein, which were ubiquitously expressed. The isolated alpha4(V) chain demonstrated an unusually high affinity for heparin. The restricted expression and unusual properties of alpha4(V)-containing collagen type V molecules suggest a unique and important role for these molecules in peripheral nerve development. PMID- 10852921 TI - Calmodulin directly gates gap junction channels. AB - Cytosolic changes control gap junction channel gating via poorly understood mechanisms. In the past two decades calmodulin participation in gating has been suggested, but compelling evidence for it has been lacking. Here we show that calmodulin indeed is associated with gap junctions and plays a direct role in chemical gating. Expression of a calmodulin mutant with the N-terminal EF hand pair replaced by a copy of the C-terminal pair dramatically increases the chemical gating sensitivity of gap junction channels composed of connexin 32 and decreases their sensitivity to transjunctional voltage. The increased chemical gating sensitivity, most likely because of the higher overall Ca(2+) binding affinity of this mutant as compared with native calmodulin, and the decreased voltage sensitivity are only observed when the mutant is expressed before connexin 32. This indicates that the mutant, and by extension native calmodulin, must interact with connexin 32 before gap junctions are formed. Immunofluorescence data suggest further that this interaction leads to incorporation of native or mutant calmodulin into the connexon as an integral regulatory subunit. PMID- 10852922 TI - Kinesin has three nucleotide-dependent conformations. Implications for strain dependent release. AB - Although crystallographic information is available on several nucleotide-induced states in myosin, little is known about the corresponding structural changes in kinesin, since a crystallographic model is only available for the kinesin:ADP complex. This makes it difficult to characterize at a molecular level the structural changes that occur in this motor through the course of its ATPase cycle. In this study, we report on the production of a series of single tryptophan mutants of a monomeric human kinesin motor domain, which demonstrate nucleotide-dependent changes in microtubule affinity that are similar to wild type. We have used these mutations to measure intramolecular distances in both strong and weak binding states, using fluorescence resonance energy transfer. This work provides direct evidence that movement of the switch II loop and helix are essential to mediate communication between the catalytic and microtubule binding sites, evidence that is supported as well by molecular modeling. Kinetic studies of fluorescent nucleotide binding to these mutants are consistent with these distance changes, and demonstrate as well that binding of ADP produces two structural transitions, neither of which are identical to that produced by the binding of ATP. This study provides a basis for understanding current structural models of the kinesin mechanochemical cycle. PMID- 10852923 TI - Regulation of the liver fatty acid-binding protein gene by hepatocyte nuclear factor 1alpha (HNF1alpha). Alterations in fatty acid homeostasis in HNF1alpha deficient mice. AB - Hepatocyte nuclear factor 1alpha (HNF1alpha)-null mice have enlarged fatty livers and alterations in the expression of genes encoding enzymes involved in the synthesis, catabolism, and transport of fatty acids. Elevations in the expression of genes encoding fatty acid synthetic enzymes (fatty acid synthase and acyl-CoA carboxylase) and peroxisomal beta-oxidation enzymes (CYP4A3, bifunctional enzyme, and thiolase) were observed in the livers of HNF1alpha-null mice, whereas hepatic mitochondrial beta-oxidation gene (medium and short chain acyl-CoA dehydrogenase) expression levels remain unchanged relative to HNF1alpha-heterozygous controls. An elevation in the levels of fatty acid transporter gene expression was also observed. In contrast, there was a marked reduction of liver fatty acid-binding protein (l-FABP) gene expression in the livers of HNF1alpha-null mice. Isolation and sequence analysis of the 5'-flanking region of the mouse l-FABP gene revealed the presence of two HNF1alpha regulatory elements. The results of transient transfection studies indicate that HNF1alpha is required to trans-activate the expression of the l-FABP promoter. Taken together, these data define a critical role for HNF1alpha in the pathogenesis of a phenotype marked by fatty infiltration of the liver and in the regulation of the l-FABP gene, the expression of which may have a direct impact on the maintenance of fatty acid homeostasis. PMID- 10852925 TI - The PDZ-interacting domain of cystic fibrosis transmembrane conductance regulator is required for functional expression in the apical plasma membrane. AB - Polarization of cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-activated chloride channel to the apical plasma membrane in epithelial cells is critical for vectorial chloride transport. Previously, we reported that the C terminus of CFTR constitutes a PDZ-interacting domain that is required for CFTR polarization to the apical plasma membrane and interaction with the PDZ domain containing protein EBP50 (NHERF). PDZ-interacting domains are typically composed of the C-terminal three to five amino acids, which in CFTR are QDTRL. Our goal was to identify the key amino acid(s) in the PDZ-interacting domain of CFTR with regard to its apical polarization, interaction with EBP50, and ability to mediate transepithelial chloride secretion. Point substitution of the C-terminal leucine (Leu at position 0) with alanine abrogated apical polarization of CFTR, interaction between CFTR and EBP50, efficient expression of CFTR in the apical membrane, and chloride secretion. Point substitution of the threonine (Thr at position -2) with alanine or valine had no effect on the apical polarization of CFTR, but reduced interaction between CFTR and EBP50, efficient expression of CFTR in the apical membrane as well as chloride secretion. By contrast, individual point substitution of the other C-terminal amino acids (Gln at position -4, Asp at position -3 and Arg at position -1) with alanine had no effect on measured parameters. We conclude that the PDZ-interacting domain, in particular the leucine (position 0) and threonine (position -2) residues, are required for the efficient, polarized expression of CFTR in the apical plasma membrane, interaction of CFTR with EBP50, and for the ability of CFTR to mediate chloride secretion. Mutations that delete the C terminus of CFTR may cause cystic fibrosis because CFTR is not polarized, complexed with EBP50, or efficiently expressed in the apical membrane of epithelial cells. PMID- 10852926 TI - Therostasin, a novel clotting factor Xa inhibitor from the rhynchobdellid leech, Theromyzon tessulatum. AB - Therostasin is a potent naturally occurring tight-binding inhibitor of mammalian Factor Xa (K(i), 34 pm), isolated from the rhynchobdellid leech Theromyzon tessulatum. Therostasin is a cysteine-rich protein (8991 Da) consisting of 82 amino acid residues with 16 cysteine residues. Its amino acid sequence has been determined by a combination of techniques, including Edman degradation, enzymatic cleavage, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) on the native and s-beta-pyridylethylated compound. Sequence analysis reveals that it shares no significant homology with other Factor Xa inhibitors except for the putative reactive site. Moreover, it contains a signature pattern for proteins of the endothelin family, potent vasoconstrictors isolated in mammal and snake venom. Therostasin cDNA (825 bp) codes for a polypeptide of 82 amino acid residues preceded by 19 residues, representing a signal peptide sequence. As for the other known inhibitors of Factor Xa, therostasin is expressed and stored in the cells of the leech salivary glands. PMID- 10852927 TI - Molecular determinants by which a long chain toxin from snake venom interacts with the neuronal alpha 7-nicotinic acetylcholine receptor. AB - Long chain curarimimetic toxins from snake venom bind with high affinities to both muscular type nicotinic acetylcholine receptors (AChRs) (K(d) in the pm range) and neuronal alpha 7-AChRs (K(d) in the nm range). To understand the molecular basis of this dual function, we submitted alpha-cobratoxin (alpha Cbtx), a typical long chain curarimimetic toxin, to an extensive mutational analysis. By exploring 36 toxin mutants, we found that Trp-25, Asp-27, Phe-29, Arg-33, Arg-36, and Phe-65 are involved in binding to both neuronal and Torpedo (Antil, S., Servent, D., and Menez, A. (1999) J. Biol. Chem. 274, 34851-34858) AChRs and that some of them (Trp-25, Asp-27, and Arg-33) have similar binding energy contributions for the two receptors. In contrast, Ala-28, Lys-35, and Cys 26-Cys-30 selectively bind to the alpha 7-AChR, whereas Lys-23 and Lys-49 bind solely to the Torpedo AChR. Therefore, alpha-Cbtx binds to two AChR subtypes using both common and specific residues. Double mutant cycle analyses suggested that Arg-33 in alpha-Cbtx is close to Tyr-187 and Pro-193 in the alpha 7 receptor. Since Arg-33 of another curarimimetic toxin is close to the homologous alpha Tyr-190 of the muscular receptor (Ackermann, E. J., Ang, E. T. H., Kanter, J. R., Tsigelny, I., and Taylor, P. (1998) J. Biol. Chem. 273, 10958-10964), toxin binding probably occurs in homologous regions of neuronal and muscular AChRs. However, no coupling was seen between alpha-Cbtx Arg-33 and alpha 7 receptor Trp-54, Leu-118, and Asp-163, in contrast to what was observed in a homologous situation involving another toxin and a muscular receptor (Osaka, H., Malany, S., Molles, B. E., Sine, S. M., and Taylor, P. (2000) J. Biol. Chem. 275, 5478-5484). Therefore, although occurring in homologous regions, the detailed modes of toxin binding to alpha 7 and muscular receptors are likely to be different. These data offer a molecular basis for the design of toxins with predetermined specificities for various members of the AChR family. PMID- 10852928 TI - Cartilage oligomeric matrix protein is a calcium-binding protein, and a mutation in its type 3 repeats causes conformational changes. AB - Mutations in residues in the type 3 calcium-binding repeats and COOH-terminal globular region of cartilage oligomeric matrix protein (COMP) lead to two skeletal dysplasias, pseudoachondroplasia and multiple epiphyseal dysplasia. It has been hypothesized that these mutations cause COMP to misfold and to be retained in the endoplasmic reticulum. However, this hypothesis is not supported by previous reports that COMP, when purified in the presence of EDTA, shows no obvious difference in electron microscopic appearance in the presence or absence of calcium ions. Since this discrepancy may be due to the removal of calcium during purification, we have expressed wild-type COMP and the most common mutant form found in pseudoachondroplasia, MUT3, using a mammalian expression system and have purified both proteins in the presence of calcium. Both proteins are expressed as pentamers. Direct calcium binding experiments demonstrate that wild type COMP, when purified in the presence of calcium, is a calcium-binding protein. Rotary shadowing electron microscopy and limited trypsin digestion at various calcium concentrations show that there are conformational changes associated with calcium binding to COMP. Whereas COMP exists in a more compact conformation in the presence of calcium, it shows a more extended conformation when calcium is removed. MUT3, with a single aspartic acid deletion in the type 3 repeats, binds less calcium and presents an intermediate conformation between the calcium-replete and calcium-depleted forms of COMP. In conclusion, we show that a single mutation in the type 3 repeats of COMP causes the mutant protein to misfold. Our data demonstrate the importance of calcium binding to the structure of COMP and provide a plausible explanation for the observation that mutations in the type 3 repeats and COOH-terminal globular region lead to pseudoachondroplasia. PMID- 10852929 TI - Reconstitution of dimethylamine:coenzyme M methyl transfer with a discrete corrinoid protein and two methyltransferases purified from Methanosarcina barkeri. AB - Methyl transfer from dimethylamine to coenzyme M was reconstituted in vitro for the first time using only highly purified proteins. These proteins isolated from Methanosarcina barkeri included the previously unidentified corrinoid protein MtbC, which copurified with MtbA, the methylcorrinoid:Coenzyme M methyltransferase specific for methanogenesis from methylamines. MtbC binds 1.0 mol of corrinoid cofactor/mol of 24-kDa polypeptide and stimulated dimethylamine:coenzyme M methyl transfer 3.4-fold in a cell extract. Purified MtbC and MtbA were used to assay and purify a dimethylamine:corrinoid methyltransferase, MtbB1. MtbB1 is a 230-kDa protein composed of 51-kDa subunits that do not possess a corrinoid prosthetic group. Purified MtbB1, MtbC, and MtbA were the sole protein requirements for in vitro dimethylamine:coenzyme M methyl transfer. An MtbB1:MtbC ratio of 1 was optimal for coenzyme M methylation with dimethylamine. MtbB1 methylated either corrinoid bound to MtbC or free cob(I)alamin with dimethylamine, indicating MtbB1 carries an active site for dimethylamine demethylation and corrinoid methylation. Experiments in which different proteins of the resolved monomethylamine:coenzyme M methyl transfer reaction replaced proteins involved in dimethylamine:coenzyme M methyl transfer indicated high specificity of MtbB1 and MtbC in dimethylamine:coenzyme M methyl transfer activity. These results indicate MtbB1 demethylates dimethylamine and specifically methylates the corrinoid prosthetic group of MtbC, which is subsequently demethylated by MtbA to methylate coenzyme M during methanogenesis from dimethylamine. PMID- 10852930 TI - Maize genetics 2000-and beyond. PMID- 10852932 TI - A shaker-like K(+) channel with weak rectification is expressed in both source and sink phloem tissues of Arabidopsis. AB - RNA gel blot and reverse transcription-polymerase chain reaction experiments were used to identify a single K(+) channel gene in Arabidopsis as expressed throughout the plant. Use of the beta-glucuronidase reporter gene revealed expression of this gene, AKT2/AKT3, in both source and sink phloem tissues. The AKT2/AKT3 gene corresponds to two previously identified cDNAs, AKT2 (reconstructed at its 5' end) and AKT3, the open reading frame of the latter being shorter at its 5' end than that of the former. Rapid amplification of cDNA ends with polymerase chain reaction and site-directed mutagenesis was performed to identify the initiation codon for AKT2 translation. All of the data are consistent with the hypothesis that the encoded polypeptide corresponds to the longest open reading frame previously identified (AKT2). Electrophysiological characterization (macroscopic and single-channel currents) of AKT2 in both Xenopus oocytes and COS cells revealed a unique gating mode and sensitivity to pH (weak inward rectification, inhibition, and increased rectification upon internal or external acidification), suggesting that AKT2 has enough functional plasticity to perform different functions in phloem tissue of source and sink organs. The plant stress hormone abscisic acid was shown to increase the amount of AKT2 transcript, suggesting a role for the AKT2 in the plant response to drought. PMID- 10852933 TI - Sterol methyltransferase 1 controls the level of cholesterol in plants. AB - The side chain in plant sterols can have either a methyl or ethyl addition at carbon 24 that is absent in cholesterol. The ethyl addition is the product of two sequential methyl additions. Arabidopsis contains three genes-sterol methyltransferase 1 (SMT1), SMT2, and SMT3-homologous to yeast ERG6, which is known to encode an S-adenosylmethionine-dependent C-24 SMT that catalyzes a single methyl addition. The SMT1 polypeptide is the most similar of these Arabidopsis homologs to yeast Erg6p. Moreover, expression of Arabidopsis SMT1 in erg6 restores SMT activity to the yeast mutant. The smt1 plants have pleiotropic defects: poor growth and fertility, sensitivity of the root to calcium, and a loss of proper embryo morphogenesis. smt1 has an altered sterol content: it accumulates cholesterol and has less C-24 alkylated sterols content. Escherichia coli extracts, obtained from a strain expressing the Arabidopsis SMT1 protein, can perform both the methyl and ethyl additions to appropriate sterol substrates, although with different kinetics. The fact that smt1 null mutants still produce alkylated sterols and that SMT1 can catalyze both alkylation steps shows that there is considerable overlap in the substrate specificity of enzymes in sterol biosynthesis. The availability of the SMT1 gene and mutant should permit the manipulation of phytosterol composition, which will help elucidate the role of sterols in animal nutrition. PMID- 10852934 TI - leafy hull sterile1 is a homeotic mutation in a rice MADS box gene affecting rice flower development. AB - Rice contains several MADS box genes. It has been demonstrated previously that one of these genes, OsMADS1 (for Oryza sativa MADS box gene1), is expressed preferentially in flowers and causes early flowering when ectopically expressed in tobacco plants. In this study, we demonstrated that ectopic expression of OsMADS1 in rice also results in early flowering. To further investigate the role of OsMADS1 during rice flower development, we generated transgenic rice plants expressing altered OsMADS1 genes that contain missense mutations in the MADS domain. There was no visible alteration in the transgenic plants during the vegetative stage. However, transgenic panicles typically exhibited phenotypic alterations, including spikelets consisting of elongated leafy paleae and lemmas that exhibit a feature of open hull, two pairs of leafy palea-like and lemma-like lodicules, a decrease in stamen number, and an increase in the number of carpels. In addition, some spikelets generated an additional floret from the same rachilla. These characteristics are very similar to those of leafy hull sterile1 (lhs1). The map position of OsMADS1 is closely linked to that of lhs1 on chromosome 3. Examination of lhs1 revealed that it contains two missense mutations in the OsMADS1 MADS domain. A genetic complementation experiment showed that the 11.9-kb genomic DNA fragment containing the wild-type OsMADS1 gene rescued the mutant phenotypes. In addition, ectopic expression of the OsMADS1 gene isolated from the lhs1 line resulted in lhs1-conferred phenotypes. These lines of evidence demonstrate that OsMADS1 is the lhs1 gene. PMID- 10852935 TI - Mutagenesis of plants overexpressing CONSTANS demonstrates novel interactions among Arabidopsis flowering-time genes. AB - CONSTANS (CO) promotes flowering of Arabidopsis in response to long photoperiods. Transgenic plants carrying CO fused with the cauliflower mosaic virus 35S promoter (35S::CO) flowered earlier than did the wild type and were almost completely insensitive to length of day. Genes required for CO to promote flowering were identified by screening for mutations that suppress the effect of 35S::CO. Four mutations were identified that partially suppressed the early flowering phenotype caused by 35S::CO. One of these mutations, suppressor of overexpression of CO 1 (soc1), defines a new locus, demonstrating that the mutagenesis approach is effective in identifying novel flowering-time mutations. The other three suppressor mutations are allelic with previously described mutations that cause late flowering. Two of them are alleles of ft, indicating that FT is required for CO to promote early flowering and most likely acts after CO in the hierarchy of flowering-time genes. The fourth suppressor mutation is an allele of fwa, and fwa soc1 35S::CO plants flowered at approximately the same time as co mutants, suggesting that a combination of fwa and soc1 abolishes the promotion of flowering by CO. Besides delaying flowering, fwa acted synergistically with 35S::CO to repress floral development after bolting. The latter phenotype was not shown by any of the progenitors and was most probably caused by a reduction in the function of LEAFY. These genetic interactions suggest models for how CO, FWA, FT, and SOC1 interact during the transition to flowering. PMID- 10852936 TI - Repression of shoot growth, a bZIP transcriptional activator, regulates cell elongation by controlling the level of gibberellins. AB - Cell expansion, a developmental process regulated by both endogenous programs and environmental stimuli, is critically important for plant growth. Here, we report the isolation and characterization of RSG (for repression of shoot growth), a transcriptional activator with a basic leucine zipper (bZIP) domain. To examine the role of RSG in plant development, we generated transgenic tobacco plants expressing a dominant-negative form of RSG, which repressed the activity of full length RSG. In transgenic plants, this expression severely inhibited stem internode growth, specifically cell elongation. These plants also had less endogenous amounts of the major active gibberellin (GA) in tobacco, GA(1). Applying GAs restored the dwarf phenotypes of transgenic tobacco plants that expressed the dominant-negative form of RSG. To investigate the function of RSG in the regulation of the endogenous amounts of GAs, we identified a target for RSG. RSG bound and activated the promoter of Arabidopsis GA3, one of the genes encoding enzymes involved in GA biosynthesis. Moreover, the dominant-negative form of RSG decreased expression of the GA3 homolog in transgenic tobacco plants. Our results show that RSG, a bZIP transcriptional activator, regulates the morphology of plants by controlling the endogenous amounts of GAs. PMID- 10852937 TI - Reduced levels of chloroplast FtsH protein in tobacco mosaic virus-infected tobacco leaves accelerate the hypersensitive reaction. AB - In tobacco cultivars resistant to tobacco mosaic virus (TMV), infection results in the death of the infected cells accompanying the formation of necrotic lesions. To identify the genes involved in this hypersensitive reaction, we isolated the cDNA of tobacco DS9, the transcript of which decreases before the appearance of necrotic lesions. The DS9 gene encodes a chloroplastic homolog of bacterial FtsH protein, which serves to maintain quality control of some cytoplasmic and membrane proteins. A large quantity of DS9 protein was found in healthy leaves, whereas the quantity of DS9 protein in infected leaves decreased before the lesions appeared. In transgenic tobacco plants containing less and more DS9 protein than wild-type plants, the necrotic lesions induced by TMV were smaller and larger, respectively, than those on wild-type plants. These results suggest that a decrease in the level of DS9 protein in TMV-infected cells, resulting in a subsequent loss of function of the chloroplasts, accelerates the hypersensitive reaction. PMID- 10852938 TI - Syncytial-type cell plates: a novel kind of cell plate involved in endosperm cellularization of Arabidopsis. AB - Cell wall formation in the syncytial endosperm of Arabidopsis was studied by using high-pressure-frozen/freeze-substituted developing seeds and immunocytochemical techniques. The endosperm cellularization process begins at the late globular embryo stage with the synchronous organization of small clusters of oppositely oriented microtubules ( approximately 10 microtubules in each set) into phragmoplast-like structures termed mini-phragmoplasts between both sister and nonsister nuclei. These mini-phragmoplasts produce a novel kind of cell plate, the syncytial-type cell plate, from Golgi-derived vesicles approximately 63 nm in diameter, which fuse by way of hourglass-shaped intermediates into wide ( approximately 45 nm in diameter) tubules. These wide tubules quickly become coated and surrounded by a ribosome-excluding matrix; as they grow, they branch and fuse with each other to form wide tubular networks. The mini-phragmoplasts formed between a given pair of nuclei produce aligned tubular networks that grow centrifugally until they merge into a coherent wide tubular network with the mini-phragmoplasts positioned along the network margins. The individual wide tubular networks expand laterally until they meet and eventually fuse with each other at the sites of the future cell corners. Transformation of the wide tubular networks into noncoated, thin ( approximately 27 nm in diameter) tubular networks begins at multiple sites and coincides with the appearance of clathrin-coated budding structures. After fusion with the syncytial cell wall, the thin tubular networks are converted into fenestrated sheets and cell walls. Immunolabeling experiments show that the cell plates and cell walls of the endosperm differ from those of the embryo and maternal tissue in two features: their xyloglucans lack terminal fucose residues on the side chain, and callose persists in the cell walls after the cell plates fuse with the parental plasma membrane. The lack of terminal fucose residues on xyloglucans suggests that these cell wall matrix molecules serve both structural and storage functions. PMID- 10852939 TI - Developmental regulation of methyl benzoate biosynthesis and emission in snapdragon flowers. AB - In snapdragon flowers, the volatile ester methyl benzoate is the most abundant scent compound. It is synthesized by and emitted from only the upper and lower lobes of petals, where pollinators (bumblebees) come in contact with the flower. Emission of methyl benzoate occurs in a rhythmic manner, with maximum emission during the day, which correlates with pollinator activity. A novel S-adenosyl-l methionine:benzoic acid carboxyl methyl transferase (BAMT), the final enzyme in the biosynthesis of methyl benzoate, and its corresponding cDNA have been isolated and characterized. The complete amino acid sequence of the BAMT protein has only low levels of sequence similarity to other previously characterized proteins, including plant O-methyl transferases. During the life span of the flower, the levels of methyl benzoate emission, BAMT activity, BAMT gene expression, and the amounts of BAMT protein and benzoic acid are developmentally and differentially regulated. Linear regression analysis revealed that production of methyl benzoate is regulated by the amount of benzoic acid and the amount of BAMT protein, which in turn is regulated at the transcriptional level. PMID- 10852940 TI - cDNA-AFLP reveals a striking overlap in race-specific resistance and wound response gene expression profiles. AB - The tomato Cf-9 gene confers resistance to races of the fungal pathogen Cladosporium fulvum expressing the Avr9 gene. cDNA amplified fragment length polymorphism analysis was used to display transcripts whose expression is rapidly altered during the Avr9- and Cf-9-mediated defense response in tobacco cell cultures. Diphenyleneiodonium was used to abolish the production of active oxygen species during gene induction. Of 30,000 fragments inspected, 290 showed altered abundance, of which 263 were induced independently of active oxygen species. cDNA clones were obtained for 13 ACRE (for Avr9/Cf-9 rapidly elicited) genes. ACRE gene induction occurred in the presence of cycloheximide. Avr9 induced ACRE gene expression in leaves. Surprisingly, ACRE genes were also rapidly but transiently induced in leaves in response to other stresses. The amino acid sequences of some ACRE proteins are homologous to sequences of known proteins such as ethylene response element binding protein transcription factors, the N resistance protein, a calcium binding protein, 13-lipoxygenase, and a RING-H2 zinc finger protein. Rapid induction of ACRE genes suggests that they play a pivotal role during plant defense responses. PMID- 10852941 TI - The kinesin-like calmodulin binding protein is differentially involved in cell division. AB - The kinesin-like calmodulin (CaM) binding protein (KCBP), a minus end-directed microtubule motor protein unique to plants, has been implicated in cell division. KCBP is negatively regulated by Ca(2)+ and CaM, and antibodies raised against the CaM binding region inhibit CaM binding to KCBP in vitro; therefore, these antibodies can be used to activate KCBP constitutively. Injection of these antibodies into Tradescantia virginiana stamen hair cells during late prophase induces breakdown of the nuclear envelope within 2 to 10 min and leads the cell into prometaphase. However, mitosis is arrested, and the cell does not progress into anaphase. Injection of antibodies later during cell division has no effect on anaphase transition but causes aberrant phragmoplast formation and delays the completion of cytokinesis by approximately 15 min. These effects are achieved without any apparent degradation of the microtubule cytoskeleton. We propose that during nuclear envelope breakdown and anaphase, activated KCBP promotes the formation of a converging bipolar spindle by sliding and bundling microtubules. During metaphase and telophase, we suggest that its activity is downregulated. PMID- 10852942 TI - Arabidopsis MutS homologs-AtMSH2, AtMSH3, AtMSH6, and a novel AtMSH7-form three distinct protein heterodimers with different specificities for mismatched DNA. AB - Arabidopsis mismatch repair genes predict MutS-like proteins remarkably similar to eukaryotic MutS homologs-MSH2, MSH3, and MSH6. A novel feature in Arabidopsis is the presence of two MSH6-like proteins, designated AtMSH6 and AtMSH7. Combinations of Arabidopsis AtMSH2 with AtMSH3, AtMSH6, or AtMSH7 proteins products of in vitro transcription and translation-were analyzed for interactions by analytical gel filtration chromatography. The AtMSH2 protein formed heterodimers with AtMSH3, AtMSH6, and AtMSH7, but no single proteins formed homodimers. The abilities of the various heterodimers to bind to mismatched 51 mer duplexes were measured by electrophoretic mobility-shift assays. Similar to the behavior of the corresponding human proteins, AtMSH2*AtMSH3 heterodimers bound "insertion-deletion" DNA with three nucleotides (+AAG) or one nucleotide (+T) looped out much better than they bound DNA with a base/base mispair (T/G), whereas AtMSH2*AtMSH6 bound the (+T) substrate strongly, (T/G) well, and (+AAG) no better than it did a (T/A) homoduplex. However, AtMSH2*AtMSH7 showed a different specificity: moderate affinity for a (T/G) substrate and weak binding of (+T). Thus, AtMSH2*AtMSH7 may be specialized for lesions/base mispairs not tested or for (T/G) mispairs in special contexts. PMID- 10852943 TI - A ligand-reversible dimerization system for controlling protein-protein interactions. AB - Chemically induced dimerization provides a general way to gain control over intracellular processes. Typically, FK506-binding protein (FKBP) domains are fused to a signaling domain of interest, allowing crosslinking to be initiated by addition of a bivalent FKBP ligand. In the course of protein engineering studies on human FKBP, we discovered that a single point mutation in the ligand-binding site (Phe-36 --> Met) converts the normally monomeric protein into a ligand reversible dimer. Two-hybrid, gel filtration, analytical ultracentrifugation, and x-ray crystallographic studies show that the mutant (F(M)) forms discrete homodimers with micromolar affinity that can be completely dissociated within minutes by addition of monomeric synthetic ligands. These unexpected properties form the basis for a "reverse dimerization" regulatory system involving F(M) fusion proteins, in which association is the ground state and addition of ligand abolishes interactions. We have used this strategy to rapidly and reversibly aggregate fusion proteins in different cellular compartments, and to provide an off switch for transcription. Reiterated F(M) domains should be generally useful as conditional aggregation domains (CADs) to control intracellular events where rapid, reversible dissolution of interactions is required. Our results also suggest that dimerization is a latent property of the FKBP fold: the crystal structure reveals a remarkably complementary interaction between the monomer binding sites, with only subtle changes in side-chain disposition accounting for the dramatic change in quaternary structure. PMID- 10852944 TI - Stochastic focusing: fluctuation-enhanced sensitivity of intracellular regulation. AB - Many regulatory molecules are present in low copy numbers per cell so that significant random fluctuations emerge spontaneously. Because cell viability depends on precise regulation of key events, such signal noise has been thought to impose a threat that cells must carefully eliminate. However, the precision of control is also greatly affected by the regulatory mechanisms' capacity for sensitivity amplification. Here we show that even if signal noise reduces the capacity for sensitivity amplification of threshold mechanisms, the effect on realistic regulatory kinetics can be the opposite: stochastic focusing (SF). SF particularly exploits tails of probability distributions and can be formulated as conventional multistep sensitivity amplification where signal noise provides the degrees of freedom. When signal fluctuations are sufficiently rapid, effects of time correlations in signal-dependent rates are negligible and SF works just like conventional sensitivity amplification. This means that, quite counterintuitively, signal noise can reduce the uncertainty in regulated processes. SF is exemplified by standard hyperbolic inhibition, and all probability distributions for signal noise are first derived from underlying chemical master equations. The negative binomial is suggested as a paradigmatic distribution for intracellular kinetics, applicable to stochastic gene expression as well as simple systems with Michaelis-Menten degradation or positive feedback. SF resembles stochastic resonance in that noise facilitates signal detection in nonlinear systems, but stochastic resonance is related to how noise in threshold systems allows for detection of subthreshold signals and SF describes how fluctuations can make a gradual response mechanism work more like a threshold mechanism. PMID- 10852945 TI - The role of spatiotemporal edges in visibility and visual masking. AB - What parts of a visual stimulus produce the greatest neural signal? Previous studies have explored this question and found that the onset of a stimulus's edge is what excites early visual neurons most strongly. The role of inhibition at the edges of stimuli has remained less clear, however, and the importance of neural responses associated with the termination of stimuli has only recently been examined. Understanding all of these spatiotemporal parameters (the excitation and inhibition evoked by the stimulus's onset and termination, as well as its spatial edges) is crucial if we are to develop a general principle concerning the relationship between neural signals and the parts of the stimulus that generate them. Here, we use visual masking illusions to explore this issue, in combination with human psychophysics, awake behaving primate neurophysiology in the lateral geniculate nucleus of the thalamus, and optical recording in the primary visual cortex of anesthetized monkeys. The edges of the stimulus, rather than its interior, generate the strongest excitatory and inhibitory responses both perceptually and physiologically. These edges can be imaged directly by using optical recording techniques. Excitation and inhibition are moreover most powerful when the stimulus turns both on and off (what might be thought of as the stimulus's temporal edges). We thus conclude that there is a general principle that relates the generation of neural signals (excitatory and inhibitory) to the spatiotemporal edges of stimuli in the early visual system. PMID- 10852946 TI - Systematic identification of mutations that constitutively activate the angiotensin II type 1A receptor by screening a randomly mutated cDNA library with an original pharmacological bioassay. AB - The constitutive activation of G-protein-coupled receptors is a major new approach to investigating their physiopathology and pharmacology. A large number of spontaneous and site-directed mutations resulting in constitutive activity have been identified, but systematic mapping of the amino acids involved for a given receptor would be extremely useful for complete elucidation of the molecular mechanisms underlying its activation. We carried out such mapping for the angiotensin II type 1A (AT(1A)) receptor by screening a randomly mutated cDNA library after expressing the mutated clones in eukaryotic cells. To test the AT(1A) mutants generated, we developed an original, specific, and highly sensitive assay based on the properties of CGP42112A. This classical AT(2) agonist is a weak partial agonist of the wild-type AT(1A) receptor and becomes a full agonist for constitutively active AT(1A) mutants, as shown experimentally and in allostery-based theoretical models. Activation of the mutated receptors by CGP42112A was monitored by using the bioluminescent protein aequorin, a very sensitive and specific sensor of intracellular calcium mobilization. The screening of 4,800 clones, providing an exhaustive coverage of all of the mutations generated, led to the identification of 16 mutations in sequences encoding the transmembrane domains that were responsible for high sensitivity to CGP42112A. The constitutive activity was confirmed by agonist-independent production of inositol phosphates, which showed that at least half of the clones had significantly increased basal activity. These data demonstrate that this new type of approach is very efficient for the systematic identification of constitutively active mutants of G-protein-coupled receptors. PMID- 10852947 TI - A bacterial two-hybrid selection system for studying protein-DNA and protein protein interactions. AB - We have developed a bacterial "two-hybrid" system that readily allows selection from libraries larger than 10(8) in size. Our bacterial system may be used to study either protein-DNA or protein-protein interactions, and it offers a number of potentially significant advantages over existing yeast-based one-hybrid and two-hybrid methods. We tested our system by selecting zinc finger variants (from a large randomized library) that bind tightly and specifically to desired DNA target sites. Our method allows sequence-specific zinc fingers to be isolated in a single selection step, and thus it should be more rapid than phage display strategies that typically require multiple enrichment/amplification cycles. Given the large library sizes our bacterial-based selection system can handle, this method should provide a powerful tool for identifying and optimizing protein-DNA and protein-protein interactions. PMID- 10852948 TI - A confederacy of bunches: fundamentals and applications of a self-associating protein. PMID- 10852949 TI - Minimal exclusion of plasma membrane proteins during retroviral envelope formation. AB - The retrovirus forms its envelope by budding at the plasma membrane (PM). This process is primarily driven by its cytoplasmic core-precursor protein, Gag, as shown by the efficient formation of virus-like Gag particles in the absence of its envelope protein, Env. Most interestingly, several studies have demonstrated incorporation of various PM proteins into retrovirus, but the underlying mechanism of this phenomenon has remained elusive. We have purified Moloney murine leukemia virus Gag particles by sedimentation in an iodixanol gradient and donor PMs by flotation in a sucrose gradient and compared their protein compositions at equal lipid basis. We found that most PM proteins are present at similar density in both membranes. The inclusion of PM proteins was unaffected by incorporation of Env protein into the envelope of the Gag particles and whether these were produced at high or low level in the cells. These findings indicate that most PM proteins become incorporated into the retrovirus envelope without significant sorting. This feature of retrovirus assembly should be considered when studying retrovirus functions and developing retrovirus vectors. PMID- 10852950 TI - A role of transcriptional activators as antirepressors for the autoinhibitory activity of TATA box binding of transcription factor IID. AB - The TATA box-binding activity of transcription factor IID (TFIID) is autoinhibited by the N-terminal domain of the Drosophila TATA box-binding protein (TBP) associated factor 230/yeast TBP-associated factor 145 subunit, which binds to the TATA box-binding domain of TBP by mimicking the TATA box structure. Here, we propose a mechanism of transcriptional activation that involves antirepression of this autoinhibitory activity by transcriptional activators. Like the autoinhibitory domain of TFIID, various acidic activators interact with the TATA box-binding domain of TBP. Moreover, the autoinhibitory domain of TFIID, which is known to interact with only the TATA box-binding domain of TBP, acts as an activation domain when fused to the GAL4 DNA-binding domain, indicating that interaction with the TATA-binding domain of TBP is crucial for activation of transcription. In a reciprocal fashion, the acidic activation domains can function as the autoinhibitory domain when the latter is replaced by the former within TFIID. These results indicate that activation domains and the autoinhibitory domain of TFIID are interchangeable, supporting a role for transcriptional activators as antirepressors of the autoinhibitory activity of the TATA box binding of TFIID. PMID- 10852951 TI - Size exclusion chromatography does not require pores. AB - Separation of macromolecules on the basis of their molecular weight by size exclusion chromatography has long been considered to be caused by the geometry dependent partition of macromolecules between a continuous phase and the porous interior of a gel or cross-linked bead. The volume of a pore accessible to a solute is limited by its relative dimensions, so larger molecules will have access to a smaller volume and will remain in a bead for a shorter time than smaller solutes. Our recent alternate picture proposes that the partition coefficient can be calculated from a thermodynamic model for the free energy of mixing of the solute with the gel phase. Size-dependent exclusion caused by the unfavorable entropy of mixing associated with the partition is predicted; the magnitude of the effect is modified by enthalpic interactions between the solute and the gel phase. This concept is extended here to describe the partition of macromolecules into a layer of terminally attached polymer chains grafted onto a solid bead. Both simple mean field and self-consistent field theory calculations predict size-dependent entropic exclusion. Experimental results obtained with neutral polymer chains grafted onto solid polystyrene latex beads confirm the predictions. PMID- 10852952 TI - Transferred DNA (T-DNA)-associated proteins of Agrobacterium tumefaciens are exported independently of virB. AB - The transfer of T-DNA from Agrobacterium to plant cells is mediated by a system which involves the virB operon of the Ti plasmid. We report that VirE2 and VirD2, two T-DNA-associated proteins, as well as VirF, a protein known to be secreted into plant cells, are present in the periplasm and supernatant fractions of growing cells of Agrobacterium as are VirJ and ChvE, two known periplasmic proteins. Two cytoplasmic proteins, Ros and chloramphenicol acetyl transferase, and a VirE2green fluorescent protein construct were not detected in the above fraction. Export of VirE2 into the culture supernatant did not require any Ti plasmid genes, except for VirE1, a specific chaperone for VirE2. The levels of the VirE2 and VirD2 proteins in the supernatant increased significantly when cells were grown at 19 degrees C as compared with 28 degrees C. When Agrobacterium expressed the oncogenic suppressive activity protein (Osa), VirE2 and VirF proteins could not be detected in the supernatant or the periplasm and the level of VirD2 was greatly reduced. However, oncogenic suppressive activity protein did not block the accumulation of VirJ and ChvE in the periplasm. Our data suggest that VirD2, VirE2, and VirF are transported across the cytoplasmic membrane by a specific pathway, independent of virB. Thus, transfer of the T complex of Agrobacterium may take place in two steps, the first mediated by an unidentified pathway and the second by the virB system. PMID- 10852953 TI - Gamma and beta frequency oscillations in response to novel auditory stimuli: A comparison of human electroencephalogram (EEG) data with in vitro models. AB - Investigations using hippocampal slices maintained in vitro have demonstrated that bursts of oscillatory field potentials in the gamma frequency range (30-80 Hz) are followed by a slower oscillation in the beta 1 range (12-20 Hz). In this study, we demonstrate that a comparable gamma-to-beta transition is seen in the human electroencephalogram (EEG) in response to novel auditory stimuli. Correlations between gamma and beta 1 activity revealed a high degree of interdependence of synchronized oscillations in these bands in the human EEG. Evoked (stimulus-locked) gamma oscillations preceded beta 1 oscillations in response to novel stimuli, suggesting that this may be analogous to the gamma-to beta shift observed in vitro. Beta 1 oscillations were the earliest discriminatory responses to show enhancement to novel stimuli, preceding changes in the broad-band event-related potential (mismatch negativity). Later peaks of induced beta activity over the parietal cortex were always accompanied by an underlying gamma frequency oscillation as seen in vitro. A further analogy between in vitro and human recordings was that both gamma and beta oscillations habituated markedly after the initial novel stimulus presentation. PMID- 10852955 TI - Neanderthal diet at Vindija and Neanderthal predation: the evidence from stable isotopes. AB - Archeological analysis of faunal remains and of lithic and bone tools has suggested that hunting of medium to large mammals was a major element of Neanderthal subsistence. Plant foods are almost invisible in the archeological record, and it is impossible to estimate accurately their dietary importance. However, stable isotope (delta(13)C and delta(15)N) analysis of mammal bone collagen provides a direct measure of diet and has been applied to two Neanderthals and various faunal species from Vindija Cave, Croatia. The isotope evidence overwhelmingly points to the Neanderthals behaving as top-level carnivores, obtaining almost all of their dietary protein from animal sources. Earlier Neanderthals in France and Belgium have yielded similar results, and a pattern of European Neanderthal adaptation as carnivores is emerging. These data reinforce current taphonomic assessments of associated faunal elements and make it unlikely that the Neanderthals were acquiring animal protein principally through scavenging. Instead, these findings portray them as effective predators. PMID- 10852954 TI - Regulation of protein kinase CbetaI by two protein-tyrosine kinases, Btk and Syk. AB - Two protein-tyrosine kinases, Bruton's tyrosine kinase (Btk) and Syk, and members of the protein kinase C (PKC) subfamily of serine/threonine kinases play crucial roles in signal transduction through antigen receptors in B lymphocytes and high affinity IgE receptors (FcepsilonRI) in mast cells. The present study provides genetic, biochemical, and pharmacological evidence that, on FcepsilonRI stimulation, Syk regulates Btk, and Btk selectively regulates the membrane translocation and enzymatic activity of PKCbetaI among the conventional PKC isoforms (alpha, betaI, and betaII) expressed in mast cells. Syk/Btk-mediated PKCbetaI regulation is involved in transcriptional activation of the IL-2 and tumor necrosis factor alpha genes through the JNK pathway induced by FcepsilonRI stimulation. Accordingly, FcepsilonRI-induced production of these cytokines is inhibited by specific inhibitors of Btk and Syk, as well as broad-specificity inhibitors of PKC and a selective inhibitor of PKCbeta. Specific regulation of PKCbetaI by Btk is consistent with the selective association of Btk with PKCbetaI. Components of this signaling pathway may represent an attractive set of potential targets of pharmaceutical interference for the treatment of allergic and other immunologic diseases. PMID- 10852956 TI - A macrophage receptor for apolipoprotein B48: cloning, expression, and atherosclerosis. AB - We have cloned a human macrophage receptor that binds to apolipoprotein (apo)B48 of dietary triglyceride (TG)-rich lipoproteins. TG-rich lipoprotein uptake by the apoB48R rapidly converts macrophages and apoB48R-transfected Chinese hamster ovary cells in vitro into lipid-filled foam cells, as seen in atherosclerotic lesions. The apoB48R cDNA (3,744 bp) encodes a protein with no known homologs. Its approximately 3.8-kb mRNA is expressed primarily by reticuloendothelial cells: monocytes, macrophages, and endothelial cells. Immunohistochemistry shows the apoB48R is in human atherosclerotic lesion foam cells. Normally, the apoB48R may provide essential lipids to reticuloendothelial cells. If overwhelmed, foam cell formation, endothelial dysfunction, and atherothrombogenesis may ensue, a mechanism for cardiovascular disease risk of elevated TG. PMID- 10852957 TI - Adenosylcobalamin inhibits ribosome binding to btuB RNA. AB - Expression of the btuB gene encoding the outer membrane cobalamin transporter in Escherichia coli is strongly reduced on growth with cobalamins. Previous studies have shown that this regulation occurs in response to adenosylcobalamin (Ado-Cbl) and operates primarily at the translational level. Changes in the level and stability of btuB RNA are consequences of the modulated translation initiation. To examine how Ado-Cbl affects translation, the binding of E. coli 30S ribosomal subunits to btuB RNA was investigated by using a primer extension inhibition assay. Ribosome binding to btuB RNA was much less efficient than to other RNAs and was preferentially lost when the ribosomes were subjected to a high-salt wash. Ribosome binding to btuB RNA was inhibited by Ado-Cbl but not by cyanocobalamin, with half-maximal inhibition around 0.3 microM Ado-Cbl. Ribosome binding activity was increased or decreased by mutations in the btuB leader region, which affected two predicted RNA hairpins and altered expression of btuB lacZ reporters. Finally, the presence of Ado-Cbl elicited formation of a single primer extension-inhibition product with the same specificity and Cbl concentration dependence as the inhibition of ribosome binding. These results indicate that btuB expression is controlled by the specific binding of Ado-Cbl to btuB RNA, which then affects access to its ribosome-binding sequence. PMID- 10852958 TI - Regulation of the homeodomain CCAAT displacement/cut protein function by histone acetyltransferases p300/CREB-binding protein (CBP)-associated factor and CBP. AB - The CCAAT displacement protein/cut homologue (CDP/cut) is a divergent homeodomain protein that is highly conserved through evolution and has properties of a potent transcriptional repressor. CDP/cut contains three conserved cut-repeat domains and a conserved homeobox, each involved in directing binding specificity to unique nucleotide sequence elements. Furthermore, CDP/cut may play a role as a structural component of chromatin through its direct interaction with nucleosomal DNA and association with nuclear matrix attachment regions. CDP/cut is cell-cycle regulated through interactions with Rb, p107, specific kinases and phosphatases directing the transcriptional activity of CDP/cut on such genes encoding p21(WAF1,CIP1), c-myc, thymidine kinase, and histones. Our previous studies indicate that CDP/cut is associated with histone deacetylase activity and is associated with a corepressor complex through interactions with histone deacetylases. Here, we report the interaction of CDP/cut with CBP and p300/CREB binding protein-associated factor (PCAF) along with the modification of CDP/cut by the histone acetyltransferase PCAF. Acetylation of CDP/cut by PCAF is directed at conserved lysine residues near the homeodomain region and regulates CDP/cut function. These observations are consistent with the ability of CDP/cut to regulate genes as a transcriptional repressor, suggesting acetylation as a mechanism that regulates CDP/cut function. PMID- 10852959 TI - A M(r) 34,000 proinflammatory outer membrane protein (oipA) of Helicobacter pylori. AB - The complete genome sequence revealed a family of 32 outer membrane proteins (OMPs) in Helicobacter pylori. We examined the effect of four OMPs (HP0638, HP0796, HP1501, and babA2) on the production of the proinflammatory cytokine, IL 8. Mutants of the four OMPs, as well as cagE and galE from H. pylori from the U.S. and Japan, were constructed by inserting a chloramphenicol-resistant cassette into the gene. Twenty-two pairs of parental and mutant H. pylori strains, as well as 160 clinical isolates (80 from Japanese and 80 from U.S.), were cocultured with gastric cancer cell lines. IL-8 production in the supernatant and adhesion was assayed by ELISA. HP0796, HP1501, babA2, and galE gene knockouts had no significant effect on IL-8 production. Knockout of the HP0638 gene in 81% of cag-positive strains reduced IL-8 production approximately 50%. The three cag-positive strains in which IL-8 levels were unchanged by HP0638 knockout had five or seven CT dinucleotide repeats in the 5' region, resulting in a frame shift and truncation. Strains with naturally inactive HP0638 gene were all from the U.S.; Japanese strains were always "on" and thus, on average, may be more virulent. Although cag-negative isolates produced a limited IL-8 response, cag-negative strains that contained a functional HP0638 gene produced more than 3 fold greater IL-8 than cag-negative nonfunctional HP0638 strains. We hypothesize that functional HP0638 gene may be an important virulence factor in relation to the risk of clinically significant outcomes of H. pylori infection. We denote HP0638 gene as outer inflammatory protein (oipA). PMID- 10852960 TI - Biosynthesis of a major lipofuscin fluorophore in mice and humans with ABCR mediated retinal and macular degeneration. AB - Increased accumulation of lipofuscin in cells of the retinal pigment epithelium (RPE) is seen in several forms of macular degeneration, a common cause of blindness in humans. A major fluorophore of lipofuscin is the toxic bis-retinoid, N-retinylidene-N-retinylethanolamine (A2E). Previously, we generated mice with a knockout mutation in the abcr gene. This gene encodes rim protein (RmP), an ATP binding cassette transporter in rod outer segments. Mice lacking RmP accumulate A2E in RPE cells at a greatly increased rate over controls. Here, we identify three precursors of A2E in ocular tissues from abcr-/- mice and humans with ABCR mediated recessive macular degenerations. Our results corroborate the scheme proposed by C. A. Parish, M. Hashimoto, K. Nakanishi, J. Dillon & J. Sparrow [Proc. Natl. Acad. Sci. USA (1998) 95, 14609-14613], for the biosynthesis of A2E: (i) condensation of all-trans-retinaldehyde (all-trans-RAL) with phosphatidylethanolamine to form a Schiff base; (ii) condensation of the amine product with a second all-trans-RAL to form a bis-retinoid; (iii) oxidation to yield a pyridinium salt; and (iv) hydrolysis of the phosphate ester to yield A2E. The latter two reactions probably occur within RPE phagolysosomes. As predicted by this model, formation of A2E was completely inhibited when abcr-/- mice were raised in total darkness. Also, once formed, A2E was not eliminated by the RPE. These data suggest that humans with retinal or macular degeneration caused by loss of RmP function may slow progression of their disease by limiting exposure to light. The precursors of A2E identified in this study may represent pharmacological targets for the treatment of ABCR-mediated macular degeneration. PMID- 10852961 TI - St. John's wort induces hepatic drug metabolism through activation of the pregnane X receptor. AB - St. John's wort (Hypericum perforatum) is an herbal remedy used widely for the treatment of depression. Recent clinical studies demonstrate that hypericum extracts increase the metabolism of various drugs, including combined oral contraceptives, cyclosporin, and indinavir. In this report, we show that hyperforin, a constituent of St. John's wort with antidepressant activity, is a potent ligand (K(i) = 27 nM) for the pregnane X receptor, an orphan nuclear receptor that regulates expression of the cytochrome P450 (CYP) 3A4 monooxygenase. Treatment of primary human hepatocytes with hypericum extracts or hyperforin results in a marked induction of CYP3A4 expression. Because CYP3A4 is involved in the oxidative metabolism of >50% of all drugs, our findings provide a molecular mechanism for the interaction of St. John's wort with drugs and suggest that hypericum extracts are likely to interact with many more drugs than previously had been realized. PMID- 10852962 TI - Unarmed, tumor-specific monoclonal antibody effectively treats brain tumors. AB - The epidermal growth factor receptor (EGFR) is often amplified and rearranged structurally in tumors of the brain, breast, lung, and ovary. The most common mutation, EGFRvIII, is characterized by an in-frame deletion of 801 base pairs, resulting in the generation of a novel tumor-specific epitope at the fusion junction. A murine homologue of the human EGFRvIII mutation was created, and an IgG2a murine mAb, Y10, was generated that recognizes the human and murine equivalents of this tumor-specific antigen. In vitro, Y10 was found to inhibit DNA synthesis and cellular proliferation and to induce autonomous, complement mediated, and antibody-dependent cell-mediated cytotoxicity. Systemic treatment with i.p. Y10 of s.c. B16 melanomas transfected to express stably the murine EGFRvIII led to long-term survival in all mice treated (n = 20; P < 0.001). Similar therapy with i.p. Y10 failed to increase median survival of mice with EGFRvIII-expressing B16 melanomas in the brain; however, treatment with a single intratumoral injection of Y10 increased median survival by an average 286%, with 26% long-term survivors (n = 117; P < 0.001). The mechanism of action of Y10 in vivo was shown to be independent of complement, granulocytes, natural killer cells, and T lymphocytes through in vivo complement and cell subset depletions. Treatment with Y10 in Fc receptor knockout mice demonstrated the mechanism of Y10 to be Fc receptor-dependent. These data indicate that an unarmed, tumor-specific mAb may be an effective immunotherapy against human tumors and potentially other pathologic processes in the "immunologically privileged" central nervous system. PMID- 10852963 TI - MEK kinase 1 gene disruption alters cell migration and c-Jun NH2-terminal kinase regulation but does not cause a measurable defect in NF-kappa B activation. AB - MEK kinase 1 (MEKK1) is a 196-kDa mitogen-activated protein kinase (MAPK) kinase kinase that, in addition to regulating the c-Jun NH(2)-terminal kinase (JNK) pathway, is involved in the control of cell motility. MEKK1(-/-) mice are defective in eyelid closure, a TGFalpha-directed process involving the migration of epithelial cells. MEKK1 expression in epithelial cells stimulates lamellipodia formation, a process required for cell movement. In addition, mouse embryo fibroblasts derived from MEKK1(-/-) mice are inhibited in their migration relative to MEKK1(+/+) fibroblasts. MEKK1 is required for JNK but not NF-kappaB activation in response to virus infection, microtubule disruption, and stimulation of embryonic stem cells with lysophosphatidic acid. MEKK1 is not required for TNFalpha or IL-1 regulation of JNK or NF-kappaB activation in macrophages or fibroblasts. Thus, MEKK1 senses microtubule integrity, contributes to the regulation of fibroblast and epithelial cell migration, and is required for activation of JNK but not NF-kappaB in response to selected stress stimuli. PMID- 10852964 TI - Dissociating the role of the medial and lateral anterior prefrontal cortex in human planning. AB - The anterior prefrontal cortex is known to subserve higher cognitive functions such as task management and planning. Less is known, however, about the functional specialization of this cortical region in humans. Using functional MRI, we report a double dissociation: the medial anterior prefrontal cortex, in association with the ventral striatum, was engaged preferentially when subjects executed tasks in sequences that were expected, whereas the polar prefrontal cortex, in association with the dorsolateral striatum, was involved preferentially when subjects performed tasks in sequences that were contingent on unpredictable events. These results parallel the functional segregation previously described between the medial and lateral premotor cortex underlying planned and contingent motor control and extend this division to the anterior prefrontal cortex, when task management and planning are required. Thus, our findings support the assumption that common frontal organizational principles underlie motor and higher executive functions in humans. PMID- 10852965 TI - A substance P-opioid chimeric peptide as a unique nontolerance-forming analgesic. AB - To elucidate mechanisms of acute and chronic pain, it is important to understand how spinal excitatory systems influence opioid analgesia. The tachykinin substance P (SP) represents the prototypic spinal excitatory peptide neurotransmitter/neuromodulator, acting in concert with endogenous opioid systems to regulate analgesic responses to nociceptive stimuli. We have synthesized and pharmacologically characterized a chimeric peptide containing overlapping NH(2)- and COOH-terminal functional domains of the endogenous opioid endomorphin-2 (EM 2) and the tachykinin SP, respectively. Repeated administration of the chimeric molecule YPFFGLM-NH(2), designated ESP7, into the rat spinal cord produces opioid dependent analgesia without loss of potency over 5 days. In contrast, repeated administration of ESP7 with concurrent SP receptor (SPR) blockade results in a progressive loss of analgesic potency, consistent with the development of tolerance. Furthermore, tolerant animals completely regain opioid sensitivity after post hoc administration of ESP7 alone, suggesting that coactivation of SPRs is essential to maintaining opioid responsiveness. Radioligand binding and signaling assays, using recombinant receptors, confirm that ESP7 can coactivate mu-opioid receptors (MOR) and SPRs in vitro. We hypothesize that coincidental activation of the MOR- and SPR-expressing systems in the spinal cord mimics an ongoing state of reciprocal excitation and inhibition, which is normally encountered in nociceptive processing. Due to the ability of ESP7 to interact with both MOR and SPRs, it represents a unique prototypic, anti-tolerance-forming analgesic with future therapeutic potential. PMID- 10852966 TI - The X-linked lymphoproliferative syndrome gene product SH2D1A associates with p62dok (Dok1) and activates NF-kappa B. AB - The X-linked lymphoproliferative syndrome (XLP) is a genetic disorder in which affected males have a morbid or fatal response to Epstein-Barr virus infection. The XLP deficiency has been mapped to a gene encoding a 128-residue protein, SH2D1A, which is comprised principally of a Src homology 2 (SH2) domain. We now report that SH2D1A associates with Dok1, a protein that interacts with Ras-GAP, Csk, and Nck. An SH2D1A SH2 domain mutant that has been identified in XLP does not associate with Dok1, in accord with the hypothesis that this interaction is linked to XLP. The association of SH2D1A with Dok1 also depends on phosphorylation of Dok1 Y(449) in the sequence ALYSQVQK. Further, overexpression of SH2D1A is found to activate NF-kappaB in 293T cells. NF-kappaB activation by SH2D1A does not depend on the wild-type SH2 domain and is inhibited by a dominant negative IkappaB kinase beta. Thus, SH2D1A can affect multiple intracellular signaling pathways that are potentially important in the normal effective host response to Epstein-Barr virus infection. PMID- 10852967 TI - Precise gene localization by phenotypic assay of radiation hybrid cells. AB - A high resolution map of the human genome previously has been constructed by using the G3 panel of human/hamster radiation hybrid cell lines and >15,000 unique human genetic markers. By determining whether human DNA sequences are present or absent in each of the hybrids, localization of single genes may routinely be achieved at approximately 250-kb resolution. In this paper we have tested whether similarly precise localization might be achieved by phenotypic screening of the hybrids to facilitate positional cloning of unknown genes. We assayed the susceptibility of each of the hybrid cell lines to transduction by retroviral vectors bearing different retroviral envelope proteins that recognize receptors present on human but not on hamster cells. The results for each of the retroviral vectors were informative and allowed precise localization of the receptor genes for the RD114 cat endogenous retrovirus, xenotropic murine leukemia virus, and type C feline leukemia virus. After cloning of the receptors for these retroviruses, we found that standard genotypic mapping by PCR gave results that were nearly identical to those from phenotypic mapping. These experiments show that precise gene localization by phenotypic assay of radiation hybrids is practical and was not appreciably impacted by the known instability of such hybrid cells. This technique should be applicable to many other human genes having discernible phenotypes in hamster cells and, with completion of the human genome project, will allow rapid identification of unknown genes on the basis of phenotype. PMID- 10852968 TI - Antiapoptotic role of the p38 mitogen-activated protein kinase-myocyte enhancer factor 2 transcription factor pathway during neuronal differentiation. AB - Myocyte enhancer factor 2 (MEF2) is in the MADS (MCM1agamous-deficiens-serum response factor) family of transcription factors. Although MEF2 is known as a myogenic factor, the expression pattern of the MEF2 family of genes (MEF2A-D) in developing brain also suggests a role in neurogenesis. Here we show that transfection with MEF2C, the predominant form in mammalian cerebral cortex, induces a mixed neuronal/myogenic phenotype in undifferentiated P19 precursor cells. During retinoic acid-induced neurogenesis of these cells, a dominant negative form of MEF2 enhances apoptosis but does not affect cell division. The mitogen-activated protein kinase p38alpha activates MEF2C. Dominant negative p38alpha also enhances apoptotic death of differentiating neurons, but these cells can be rescued from apoptosis by coexpression of constitutively active MEF2C. These findings suggest that the p38alpha/MEF2 pathway prevents cell death during neuronal differentiation. PMID- 10852969 TI - Pectin engineering: modification of potato pectin by in vivo expression of an endo-1,4-beta-D-galactanase. AB - Potato tuber pectin is rich in galactan (oligomer of beta-1,4-linked galactosyl residues). We have expressed a fungal endo-galactanase cDNA in potato under control of the granule bound starch synthase promoter to obtain expression of the enzyme in tubers during growth. The transgenic plants displayed no altered phenotype compared with the wild type. Fungal endo-galactanase activity was quantified in the transgenic tubers, and its expression was verified by Western blot analysis. The effect of the endo-galactanase activity on potato tuber pectin was studied by Fourier transform infrared microspectroscopy, immuno-gold labeling, and sugar analysis. All analyses revealed alterations in pectin composition. Monosaccharide composition of total cell walls and isolated rhamnogalacturonan I fragments showed a reduction in galactosyl content to 30% in the transformants compared with the wild type. Increased solubility of pectin from transgenic cell walls by endo-polygalacturonase/pectin methylesterase digestion points to other changes in wall architecture. PMID- 10852970 TI - The C-terminal domain-phosphorylated IIO form of RNA polymerase II is associated with the transcription repressor NC2 (Dr1/DRAP1) and is required for transcription activation in human nuclear extracts. AB - Activation of class II gene transcription may involve alleviation of transcription repression as well as stimulation of the assembly and function of the general RNA polymerase (RNAP) II transcription machinery. Here, we investigated whether activator-reversible transcription repression by NC2 (Dr1/DRAP1) contributes to maximum induction levels in unfractionated HeLa nuclear extracts. Surprisingly, we found that depletion of NC2 does not significantly affect basal transcription, but dramatically reduces activated transcription. Immunoblot analyses revealed that the loss of activator function coincides with selective removal of the C-terminal domain (CTD) hyperphosphorylated RNAP IIO along with NC2. Coimmunoprecipitation experiments with purified factors confirmed that NC2 interacts with RNAP IIO, but not with the unphosphorylated or hypophosphorylated RNAP IIA or CTD-less RNAP IIB forms. Finally, we demonstrate that, in contrast to previously published observations in cell-free systems reconstituted with purified factors, only the CTD phosphorylated form of RNAP II can mediate activator function in the context of unfractionated HeLa nuclear extracts. These findings reveal an unexpected link between NC2 and transcription activation and suggest that regulation of RNAP II transcription through reversible CTD phosphorylation might be more complex than previously proposed. PMID- 10852972 TI - Peaks and troughs in linkage mapping for the rheumatic diseases. PMID- 10852971 TI - v-Crk activates the phosphoinositide 3-kinase/AKT pathway in transformation. AB - v-Crk induces cellular tyrosine phosphorylation and transformation of chicken embryo fibroblasts (CEF). We studied the molecular mechanism of the v-Crk-induced transformation. Experiments with Src homology (SH)2 and SH3 domain mutants revealed that the induction of tyrosine phosphorylation of cellular proteins requires only the SH2 domain, but both the SH2 and SH3 domains are required for complete transformation. Analysis of three well defined signaling pathways, the mitogen-activated protein kinase (MAPK) pathway, the Jun N-terminal kinase (JNK) pathway, and the phosphoinositide 3-kinase (PI3K)/AKT pathway, demonstrated that only the PI3K/AKT pathway is constitutively activated in v-Crk-transformed CEF. Both the SH2 and SH3 domains are required for this activation of the PI3K/AKT pathway in CEF. We also found that the colony formation of CEF is strongly induced by a constitutively active PI3K mutant, and that a PI3K inhibitor, LY294002, suppresses the v-Crk-induced transformation. These results strongly suggest that constitutive activation of the PI3K/AKT pathway plays an essential role in v-Crk-induced transformation of CEF. PMID- 10852973 TI - Shared care--the way forward for patient-orientated follow-up care in rheumatoid arthritis. PMID- 10852974 TI - Tracking T cells in arthritis. PMID- 10852975 TI - Nuclear medicine in vasculitis. PMID- 10852976 TI - Fas-associated death domain protein is a Fas-mediated apoptosis modulator in synoviocytes. AB - OBJECTIVE: To understand the intracellular regulatory mechanisms in Fas-mediated apoptosis of synoviocytes, we examined the involvement of caspases [caspase-1/ICE (interleukin-1beta converting enzyme), caspase-3/CPP32, and caspase-8/FLICE] and Fas-associated death domain protein (FADD) forming a death-inducing signalling complex (DISC) in Fas-mediated apoptosis of synoviocytes. METHODS: Synoviocytes were obtained from rheumatoid arthritis (RA) and osteoarthritis (OA) patients. The number of dead cells was counted after treatment with anti-Fas monoclonal antibody in the presence of caspase-1-, -3-, or -8-specific inhibitors. The involvement of caspases and FADD in Fas-mediated apoptosis of RA synoviocytes was examined by immunoblot and immunoprecipitation analyses. RESULTS: RA synoviocytes expressed high levels of caspase-3, caspase-8, and FADD compared with OA synoviocytes. Interestingly, Fas ligation activated caspase-8 and caspase-3 with the cleavage of poly(ADP-ribose) polymerase (PARP), corresponding to apoptosis of RA synoviocytes. Furthermore, specific inhibitors for caspase-3 and caspase-8 but not caspase-1 suppressed Fas-induced apoptosis of RA synoviocytes in a dose- and time-dependent manner. Caspase-8-specific inhibitor suppressed the activation of caspase-3 after Fas ligation on RA synoviocytes. Importantly, FADD was selectively recruited to the Fas death domain during Fas-mediated apoptosis of RA synoviocytes, consistent with sensitivity to the Fas-mediated apoptosis. CONCLUSION: Our findings suggest that Fas-mediated apoptosis in synoviocytes may be regulated at the level of recruitment of FADD to the DISC, subsequently leading to the activation of the FADD/caspase-8/caspase-3 signalling pathway. PMID- 10852977 TI - Prospective evaluation of the frequency and clinical significance of antineutrophil cytoplasmic and anticardiolipin antibodies in community cases of patients with rheumatoid arthritis. AB - OBJECTIVES: To evaluate the frequencies of antineutrophil cytoplasmic (ANCA), anticardiolipin (aCLA) and anti-beta(2)-glycoprotein 1 antibodies (abeta(2)-GP1A) in rheumatoid arthritis (RA) of limited duration in patients recruited primarily from private practitioners (80%), and to attempt to correlate the presence of these antibodies with certain clinical and/or biological criteria. Patients and methods. Patients (n = 102) with RA evolving for <5 yr (mean 2.2 yr) were recruited. A home evaluation collected clinical data [Ritchie articular index, Health Assessment Questionnaire (HAQ) index, extra-articular manifestations] and blood for biological analyses [C-reactive protein (CRP), rheumatoid factor, ANCA, aCLA, abeta(2)-GP1A]. ANCA were detected by indirect immunofluorescence on neutrophils and their specificity was determined by enzyme-linked immunosorbent assay (ELISA) and confirmed by immunoblotting; aCLA and abeta(2)-GP1A were detected by ELISA. RESULTS: Patients had mild RA (Ritchie = 11/78 +/- 9.6; HAQ = 0.79/3 +/- 0.7), probably due to the recruitment procedure. ANCA, aCLA and abeta(2)-GP1A frequencies were 18.5, 7 and 0%, respectively. Titres of ANCA and aCLA were low. A perinuclear ANCA staining pattern was exclusively observed and lactoferrin was shown to be the major antigen recognized. No relationship was found between ANCA and aCLA and/or rheumatoid factor, or any clinical manifestations. ANCA were more common in RA of longer duration (cut-off: 4 yr; P = 0.05) and aCLA were correlated with the CRP level (P = 0.05). CONCLUSIONS: In RA of recent onset, ANCA and aCLA were detected at low titres and frequencies, and were not associated with any clinical manifestations. A longitudinal study is needed to determine whether their early appearance is predictive of subsequent disease severity. PMID- 10852978 TI - Factors associated with functional impairment in symptomatic knee osteoarthritis. AB - OBJECTIVES: Knee osteoarthritis (OA) is a major cause of disability, particularly in the elderly. The factors determining disability remain unclear. The aim of this study was to assess the impact of clinical and psychosocial variables on function in knee OA and to develop models to account for observed variance in self-reported disability. METHODS: The subjects (n = 69) were hospital out patients. Self-reported disability was measured by the Western Ontario and McMaster Universities (WOMAC) OA index. Pain was measured by the WOMAC and the McGill pain questionnaire. Depression, anxiety, helplessness, self-efficacy, fatigue and quality of life were measured by standard instruments. A detailed knee examination, including pain threshold by dolorimetry, was performed. Radiographs were scored for individual features. RESULTS: Pain severity, obesity and helplessness were the most important determinants of disability: a model including these variables accounted for 59.9% variance in WOMAC disability. Anxiety remained associated with disability in some models. Disability was unrelated to radiographic change. CONCLUSIONS: Function in symptomatic knee OA is determined more by pain and obesity than by structural change, at least as seen on plain X-ray. Our study provides further support for interventions targeting anxiety and helplessness in knee OA. PMID- 10852979 TI - Do patients with ankylosing spondylitis have poorer balance than normal subjects? AB - OBJECTIVES: To investigate whether patients with ankylosing spondylitis have poorer balance than normal subjects, and to study the relationship between balance and posture. METHODS: Balance was studied in 30 ankylosing spondylitis subjects using sway magnetometry, making quantitative measurements of movement at the hips with eyes open and eyes closed. The results were compared with data from 58 normal subjects. Balance was also compared with quantitative measurements of posture. RESULTS: The numbers of patients with poor balance, above the 95th percentile for normal, were significantly greater than expected; 18% for eyes open (P = 0. 03) and 23% for eyes closed (P = 0.004). No significant relationships between balance and any of the quantitative descriptions of posture were demonstrated. CONCLUSION: A significant proportion of ankylosing spondylitis patients have poorer balance than normal subjects. PMID- 10852980 TI - An observer-blinded comparison of supervised and unsupervised aerobic exercise regimens in fibromyalgia. AB - OBJECTIVE: To compare a supervised 12-week aerobic exercise class with unsupervised home aerobic exercises in the treatment of patients with fibromyalgia. METHODS: This was a 48-week randomized single (observer) blind study in a teaching hospital rheumatology and physiotherapy department. The subjects were 74 patients who fulfilled the American College of Rheumatology criteria for fibromyalgia. Results and conclusions. A 12-week exercise class programme with home exercises demonstrated no benefit over a single physiotherapy session with home exercises in the treatment of pain in patients with fibromyalgia. Neither group (nor the groups combined) showed an improvement in pain compared with baseline. There was some significant benefit in psychological well-being in the exercise class group and perhaps a slowing of functional deterioration in this group. PMID- 10852981 TI - Quantitation of microcirculatory abnormalities in patients with primary Raynaud's phenomenon and systemic sclerosis by video capillaroscopy. AB - OBJECTIVE: : To assess nailfold capillary density and dimensions in patients with primary Raynaud's phenomenon (PRP), limited cutaneous systemic sclerosis (LSSc) and diffuse cutaneous SSc (DSSc), and healthy control subjects. METHODS: : Using the technique of nailfold video capillaroscopy, capillary density and dimensions were averaged from all visible capillaries in a 3 mm length of the nailfold from right and left ring fingers of each subject. Twenty healthy control subjects, 15 patients with PRP, 13 patients with DSSc and 21 patients with LSSc were examined. Intra-observer and inter-observer variability were calculated in 18 and 23 patients, respectively. RESULTS: : There were significant trends for capillary density to fall and for all dimensions to rise across the four groups (P < 0. 0001 for density and all dimensions, order healthy controls, PRP, DSSc and LSSc). Intra- and inter-observer reproducibility studies showed that although there was good correlation between and within observers, the limits of agreement were between +/-25-50% indicating lack of reproducibility. CONCLUSIONS: : Microcirculatory abnormalities can be quantified using the technique of video capillaroscopy and were most marked in patients with LSSc. PMID- 10852982 TI - Changes in the Th1 or Th2 cytokine dominance in the synovium of rheumatoid arthritis (RA): a kinetic study of the Th subsets in one unusual RA patient. AB - OBJECTIVE: To perform a kinetic study of the Th1/Th2 balance in the rheumatoid arthritis (RA) synovium. METHODS: Three different synovial tissue (ST) samples were obtained from one patient with erosive RA. The characterization of Th1 and Th2 responses was performed by interferon-gamma and interleukin-4 measurements and by expression of the chemokine receptors CCR5 and CCR3. Measurements of secreted and surface immunoglobulin determined the types of B cells. RESULTS: The first ST sample yielded 31 CD4+ T cell clones which showed an unusual Th2 dominant pattern in the inflamed synovium. The Th2 response was associated with predominantly synovial IgG B cells, and a predominantly Th1 profile in the peripheral blood. In contrast, ST samples obtained 2 and 2.5 yr later displayed first a Th0 and thereafter a Th1 profile, and the synovial B cell response was predominantly of IgM type. The T cell lines from the Th1/Th0 tissues expressed the Th1 marker CCR5 but not CCR3, while the T cells from the Th2 tissue expressed the Th2 marker CCR3 and no CCR5. CONCLUSION: These results demonstrate that a predominantly Th2 response can be associated with active erosive RA. However, the Th2 profile was not permanent and changed into a Th0 and thereafter a Th1 profile. PMID- 10852983 TI - Acute effects of etidronate on glucocorticoid-induced bone degradation. AB - OBJECTIVES: To study the acute short-term effects on the biochemical parameters of calcium and bone homeostasis in post-menopausal women treated with a high dose of prednisone alone or with additional etidronate, before and during 5 days of treatment. METHODS: Serum calcium, phosphorus, creatinine, alkaline phosphatase activity, osteocalcin, carboxy-terminal propeptide of type I procollagen (PICP), cross-linked carboxy-terminal telopeptide of type I collagen (ICTP), parathyroid hormone (PTH), 25-hydroxyvitamin D and urinary excretion of calcium over 24 h were measured before and during 5 days of treatment in 14 post-menopausal women treated with a high dose of prednisone (60 mg/day) alone (group A) or combined with cyclical etidronate (group B). RESULTS: Significant differences from baseline were found in osteocalcin and urinary excretion of calcium in both groups and for ICTP in group B. Significant differences between groups were calculated at day 5 of the study for osteocalcin, ICTP and 24 h urine calcium excretion (P < 0.01). Urinary excretion of calcium over 24 h increased in group A (+14.7%; P < 0.05) and decreased in group B (-22.1%; P < 0.01). Osteocalcin levels decreased in group A (- 38.1%) and increased in group B (+27.4%; both P < 0.01). ICTP decreased only in group B (-19.4%; P < 0.01). CONCLUSIONS: The results are consistent with the fact that etidronate is acutely able to prevent bone resorption due to corticosteroids. The increase in osteocalcin in the etidronate-treated group is a new feature. A direct or indirect (PTH, 1,25 vitamin D?) stimulatory effect of etidronate on the osteoblast cannot be excluded. PMID- 10852984 TI - Polymorphic CAG repeats of the androgen receptor gene in Japanese male patients with ankylosing spondylitis. AB - OBJECTIVE: In view of a possible role of androgens in the pathogenesis of ankylosing spondylitis (AS), we investigated the association between Japanese male patients with AS and CAG microsatellites of the androgen receptor (AR) gene which related to the AR transactivation function. METHODS: Peripheral blood was collected from 39 men with AS and 305 male control subjects. The number of CAG repeats in exon 1 of the AR gene was determined. RESULTS: CAG repeat lengths in AS patients were significantly shorter than those in the controls (median value 22 vs 23; P = 0.03). However, there was no significant difference in CAG repeats between HLA-B27-positive and -negative patients (median value 22 vs 22; P = 0.78). CONCLUSIONS: Shorter CAG repeats of the AR gene, presenting high levels of transactivation activity, may play a role in male AS. PMID- 10852985 TI - Methotrexate as a preferential cyclooxygenase 2 inhibitor in whole blood of patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate the regulation of whole-blood cyclooxygenase-1 and -2 (COX-2 and COX-1) activities by methotrexate (MTX) in rheumatoid arthritis (RA) patients. METHODS: Whole blood was withdrawn from nine healthy volunteers, 12 RA patients treated with MTX (RA/MTX) and six RA patients treated with chloroquine (RA/CQ). COX-1 activity was quantified as platelet thromboxane B(2) production in unstimulated blood and COX-2 activity was measured as prostaglandin E(2) (PGE(2)) production in whole blood stimulated with LPS. Thromboxane B(2) and PGE(2) were measured by radioimmunoassay. We studied the drug effect in vitro by direct incubation of MTX with blood obtained from normal donors. Ex vivo assays were performed with blood collected from RA/MTX and RA/CQ patients. The influence of serum factors on enzyme activities was analysed in blood collected from normal donors and incubated with RA/MTX, autologous or heterologous serum. RESULTS: In vitro assays showed no direct action of MTX on the activity of either enzyme. Assays performed with blood from RA/MTX patients showed preferential inhibition of COX-2 activity (PGE(2) = 10.11 +/- 2.42 ng/ml) when compared with blood of normal donors (PGE(2) = 37.7 +/- 4.36 ng/ml; P = 0.001). Inhibition of COX-2 activity was also observed when blood of normal donors was co-incubated with RA/MTX serum. CONCLUSION: Our results clearly show that the anti-inflammatory action of low-dose MTX is partly mediated by a serum factor induced by MTX or a MTX metabolite that preferentially inhibits the activity of COX-2. PMID- 10852986 TI - Geographic clustering of an outer surface protein A mutant of Borrelia burgdorferi. Possible implications of multiple variants for Lyme disease persistence. AB - DNA sequences encoding full-length outer surface protein (Osp) A were amplified from four joint fluid samples over 4.5 months from a patient with chronic Lyme arthritis, with a variant from wild type only found in sample 3. Rather than a mutation in vivo, these findings suggested a mixed infection in which BORRELIA: containing the wild-type and mutant ospA were waxing and waning in the patient's joint. If so, we reasoned that the mutant should be present in the community. We therefore took the novel epitope resulting from the mutation, expressed as a fusion protein in Escherichia coli, and performed Western blots on 80 high-titred stored sera; however, all except that of our index patient were negative. We then collected 36 stored sera from patients with Lyme disease residing within 10 miles of where the index patient had lived. An additional two sera from this circumscribed area were positive (P = 0.038). These findings show that results from single samples can be misleading, and suggest that the OspAs expressed in force late in Lyme arthritis are the same ones introduced initially into the host. Moreover, they allow a speculative mechanism for disease persistence not previously considered, in which antigenically distinct B. burgdorferi variant proteins present themselves serially to the immune system. PMID- 10852987 TI - Clinical quality management in rheumatoid arthritis: putting theory into practice. Swiss Clinical Quality Management in Rheumatoid Arthritis. AB - Clinical quality management (CQM) in rheumatoid arthritis (RA) aims to reduce inflammatory activity and pain in the short term, and damage, and consequently disability, in the long term. Within CQM as used in Switzerland rheumatologists are provided with a measurement feedback system with which they can regularly follow their patients. Inflammatory activity is measured with the Disease Activity Score (DAS28) and the Rheumatoid Arthritis Disease Activity Index questionnaire (RADAI), damage with an X-ray score and disability with the Stanford Health Assessment Questionnaire (HAQ). Feedback is used to optimize therapy, which in the short term allows the activity of the inflammatory process to be adjusted or 'titrated'. In the long term, the therapy result for the individual patient is monitored by the course of disability and damage. In this paper we present a series of cases to illustrate the usefulness of the CQM system in the management of individual RA patients. CQM in RA may be helpful when making decisions about adjustment of treatment, and to document and communicate these decisions based on quantitative data. PMID- 10852988 TI - Oral health and juvenile idiopathic arthritis: a review. AB - Juvenile idiopathic arthritis (JIA) results in significant morbidity that includes an adverse impact on oral health that is generally not well recognized. This review describes current literature which demonstrates poor oral health in children with JIA. The impact of JIA on oral health is probably multifactorial and these factors are discussed. This review emphasizes the role of paediatric dentistry in the multidisciplinary management of JIA and highlights the need for further research. PMID- 10852989 TI - Favourable outcome in 135 children with juvenile systemic sclerosis: results of a multi-national survey. AB - OBJECTIVE: To increase the current knowledge of the outcome of juvenile systemic sclerosis (jSSc), which is currently limited. METHODS: In order to investigate the patient outcome and prognostic factors, starting October 1994, we distributed questionnaires to 324 paediatric rheumatology centres. RESULTS: Until 15 May 1998 responses from 46 centres were received, 34 of which returned completed questionnaires on a total of 135 patients. One hundred and twenty-two of the 135 patients were Caucasian, 100 were female. The mean age at disease onset was 8.8 yr (S.D. +/- 3.3 yr). The mean disease duration at the last follow-up was 5 yr(S.D. +/- 3.3 yr). At the last follow-up the disease was still active and required medication in 82 patients, 36 had inactive disease on medication, and 16 were in remission. Ninety per cent of the living patients were fully active in daily life at the last follow-up. Eight of the 135 patients had died. These patients had a median age at onset of the disease of 10.5 yr (range 6.7-15.8 yr). The median disease duration until death was 2 yr (range 1-8 yr). The causes of death were heart failure (five), renal failure (one), sepsis (one) and in one case the cause was not defined. The 1 yr survival rate was 99%, the 2 yr was 97% and the 4 yr was 95%. CONCLUSIONS: At a mean follow-up of 5 yr, the current results show a favourable outcome in most patients with childhood onset jSSc and a significantly better survival than in the adult SSc patients. PMID- 10852990 TI - How to supervise a thesis--best practice. Research and Training Committee of the British Society for Rheumatology. PMID- 10852991 TI - Autologous stem cell transplantation in a lymphoma patient with a long history of ankylosing spondylitis. PMID- 10852992 TI - Antiphospholipid antibody-associated haemophagocytic syndrome. PMID- 10852993 TI - Antiphospholipid antibody syndrome and polymyalgia rheumatica/giant cell arteritis. PMID- 10852994 TI - Use of pamidronate for acute pain relief following osteoporotic vertebral fractures. PMID- 10852995 TI - Anticardiolipin antibodies and antibodies to beta(2)-glycoprotein I in patients with Wegener's granulomatosis. PMID- 10852996 TI - Radiation synovectomy for warfarin-induced haemarthroses. PMID- 10852997 TI - Robin goodfellow PMID- 10852998 TI - A comparison of colonoscopy and double-contrast barium enema for surveillance after polypectomy. National Polyp Study Work Group. AB - BACKGROUND: After patients have undergone colonoscopic polypectomy, it is uncertain whether colonoscopic examination or a barium enema is the better method of surveillance. METHODS: As part of the National Polyp Study, we offered colonoscopic examination and double-contrast barium enema for surveillance to patients with newly diagnosed adenomatous polyps. Although barium enema was performed first, the endoscopist did not know the results. RESULTS: A total of 973 patients underwent one or more colonoscopic examinations for surveillance. In the case of 580 of these patients, we performed 862 paired colonoscopic examinations and barium-enema examinations that met the requirements of the protocol. The findings on barium enema were positive in 222 (26 percent) of the paired examinations, including 139 of the 392 colonoscopic examinations in which one or more polyps were detected (rate of detection, 35 percent; 95 percent confidence interval, 31 to 40 percent). The proportion of examinations in which adenomatous polyps were detected by barium enema colonoscopy was significantly related to the size of the adenomas (P=0.009); the rate was 32 percent for colonoscopic examinations in which the largest adenomas detected were 0.5 cm or less, 53 percent for those in which the largest adenomas detected were 0.6 to 1.0 cm, and 48 percent for those in which the largest adenomas detected exceeded 1.0 cm. Among the 139 paired examinations with positive results on barium enema and negative results on colonoscopic examination in the same location, 19 additional polyps, 12 of which were adenomas, were detected on colonoscopic reexamination. CONCLUSIONS: In patients who have undergone colonoscopic polypectomy, colonoscopic examination is a more effective method of surveillance than double contrast barium enema. PMID- 10852999 TI - Thrombotic thrombocytopenic purpura associated with clopidogrel. AB - BACKGROUND: The antiplatelet drug clopidogrel is a new thienopyridine derivative whose mechanism of action and chemical structure are similar to those of ticlopidine. The estimated incidence of ticlopidine-associated thrombotic thrombocytopenic purpura is 1 per 1600 to 5000 patients treated, whereas no clopidogrel-associated cases were observed among 20,000 closely monitored patients treated in phase 3 clinical trials and cohort studies. Because of the association between ticlopidine use and thrombotic thrombocytopenic purpura and other adverse effects, clopidogrel has largely replaced ticlopidine in clinical practice. More than 3 million patients have received clopidogrel. We report the clinical and laboratory findings in 11 patients in whom thrombotic thrombocytopenic purpura developed during or soon after treatment with clopidogrel. METHODS: The 11 patients were identified by active surveillance by the medical directors of blood banks (3 patients), hematologists (6), and the manufacturer of clopidogrel (2). RESULTS: Ten of the 11 patients received clopidogrel for 14 days or less before the onset of thrombotic thrombocytopenic purpura. Although 10 of the 11 patients had a response to plasma exchange, 2 required 20 or more exchanges before clinical improvement occurred, and 2 had relapses while not receiving clopidogrel. One patient died despite undergoing plasma exchange soon after diagnosis. CONCLUSIONS: Thrombotic thrombocytopenic purpura can occur after the initiation of clopidogrel therapy, often within the first two weeks of treatment. Physicians should be aware of the possibility of this syndrome when initiating clopidogrel treatment. PMID- 10853000 TI - Magnitude of left ventricular hypertrophy and risk of sudden death in hypertrophic cardiomyopathy. AB - BACKGROUND: Sudden death is known to be a possible consequence of hypertrophic cardiomyopathy. Quantification of the risk of sudden death, however, remains imprecise for most patients with this disease. METHODS: We assessed the relation between the magnitude of left ventricular hypertrophy and mortality in 480 consecutive patients with hypertrophic cardiomyopathy. The patients were categorized into five subgroups according to maximal wall thickness: 15 mm or less, 16 to 19 mm, 20 to 24 mm, 25 to 29 mm, and 30 mm or more. Their ages ranged from 1 to 89 years (median, 47). RESULTS: Over a mean follow-up period of 6.5 years, 65 of the 480 patients (14 percent) died: 23 suddenly, 15 of heart failure, and 27 of noncardiac causes or stroke. The risk of sudden death increased progressively and in direct relation to wall thickness (P=0.001), ranging from 0 per 1000 person-years (95 percent confidence interval, 0 to 14.4) for a wall thickness of 15 mm or less to 18.2 per 1000 person-years (95 percent confidence interval, 7.3 to 37.6) for a wall thickness of 30 mm or more and almost doubling from each wall-thickness subgroup to the next. The cumulative risk 20 years after the initial evaluation was close to zero for patients with a wall thickness of 19 mm or less but almost 40 percent for wall thicknesses of 30 mm or more. As compared with the other subgroups, patients with extreme hypertrophy were the youngest (mean age, 31 years), and most (41 of 43) had mild symptoms or no symptoms; of the 12 patients who were less than 18 years old at the initial evaluation, 5 died suddenly. CONCLUSIONS: In hypertrophic cardiomyopathy, the magnitude of hypertrophy is directly related to the risk of sudden death and is a strong and independent predictor of prognosis. Young patients with extreme hypertrophy, even those with few or no symptoms, appear to be at substantial long-term risk and deserve consideration for interventions to prevent sudden death. The majority of patients with mild hypertrophy are at low risk and can be reassured regarding their prognosis. PMID- 10853001 TI - Genetic and environmental factors in age-related nuclear cataracts in monozygotic and dizygotic twins. AB - BACKGROUND: Age-related cataracts are a major public health problem. The relative importance of genes and environment in the causation of nuclear cataracts, the most common form of age-related cataracts, is not known. METHODS: We studied 506 pairs of female twins (226 monozygotic and 280 dizygotic) who were 50 to 79 years old (mean, 62). The amount of nuclear cataract in the right and left eyes was determined objectively by analysis of Scheimpflug lens photographs (yielding three measures) and subjectively with use of the Oxford Clinical Cataract Classification and Grading System (yielding one measure). All eight measures (four in each eye) were subsequently combined in one summary measure of nuclear cataract for each woman. A univariate maximum-likelihood model was used to estimate the variance of the genetic and environmental contributions to each of the measures. RESULTS: The different measures of cataract formation were highly correlated (correlation coefficients, 0.71 to 0.94). The mean scores were similar for the right and left eyes and for monozygotic and dizygotic twins. Quantitative genetic modeling of each of the nuclear-cataract scores invariably resulted in a best-fitting model that involved additive genetic effects, unique environmental effects, and age. The common environmental and dominant genetic effects could be removed from the models without significant loss of fit. The overall heritability in the combined nuclear-cataract score (the proportion of the variance explained by genetic factors) was 48 percent (95 percent confidence interval, 42 to 54 percent); age accounted for 38 percent of the variance (95 percent confidence interval, 31 to 44 percent) and unique environmental effects for 14 percent (95 percent confidence interval, 12 to 18 percent). CONCLUSIONS: Genetic effects are important even in such a clearly age-related disease as nuclear cataract, explaining almost 50 percent of the variation in the severity of this disease. PMID- 10853002 TI - Images in clinical medicine. Elemental mercury embolism to the lung. PMID- 10853003 TI - Plasminogen-activator inhibitor type 1 and coronary artery disease. PMID- 10853004 TI - Erectile dysfunction. PMID- 10853005 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 18-2000. A 45-year-old woman with a thoracic mass and Pancoast's syndrome. PMID- 10853006 TI - The end of barium enemas? PMID- 10853007 TI - Thrombotic thrombocytopenic purpura and clopidogrel--a need for new approaches to drug safety. PMID- 10853008 TI - The Icelandic Healthcare Database and informed consent. PMID- 10853010 TI - Correction: Ischemic Retinopathy Caused by Severe Megaloblastic Anemia. PMID- 10853009 TI - Rules for research on human genetic variation--lessons from Iceland. PMID- 10853011 TI - Fas and Fas ligand interactions in malignant disease. AB - Apoptosis has been implicated in tumor development and progression. Fas (CD95) and Fas ligand (FasL) are an interacting receptor ligand pair that elicits apoptosis in many cell types. Although originally described as proteins regulating peripheral immune tolerance, accumulating evidence suggests that Fas/FasL may play an important role in carcinogenesis, tumor outgrowth, and metastasis. This review summarizes our current knowledge about the regulation of Fas and FasL expression, Fas signaling, soluble Fas production, the role(s) of Fas and FasL in hematopoietic and non-hematopoietic tumorigenesis and progression, and the potential application of Fas-induced apoptosis in cancer therapy. PMID- 10853012 TI - Twenty years of experience with the steroid receptor external quality assessment program - the paradigm for tumour biomarker EQA studies. On behalf of the EROTC Receptor and Biomarker Study Group. AB - Estrogen receptor (ER) assays have clinical relevance in selecting women who would benefit from endocrine intervention. As the degree of benefit from endocrine therapy is directly related to the quantity of receptor present in the tumour, the quality of the steroid receptor assays is important. Moreover, since patients entered in multi-centre trials often include stratification based on the receptor status, receptor assays should be comparable between different institutes. ER- and progesterone receptor (PgR)-assays have been evaluated in quality assessment studies for almost 20 years by the EORTC Receptor and Biomarker Study Group. This study analyses our findings over these years and concludes with a recommended minimum structure on which QA schemes for any biomarker could be based. PMID- 10853013 TI - Elevated phosphorylation of AKT and Stat3 in prostate, breast, and cervical cancer cells. AB - We examined whether the persistent activation of AKT or Stat3 oncogene product is present in prostate, breast, and cervical cancer cells. We found that some prostate and breast cancer cell lines express high levels of phosphorylated AKT. Interestingly, the cancer cells, which only express low levels of phosphorylated AKT express high levels of phosphorylated Stat3. AKT or Stat3 is also highly phosphorylated in human papilloma virus-negative cervical cancer cells. Therefore, these results indicate that AKT and Stat3 are highly phosphorylated in some breast, prostate and cervical cancer cells, which may play a role in tumorigenesis of these cancers. PMID- 10853014 TI - Expression of the SART1 tumor-rejection antigen in human osteosarcomas. AB - We recently reported a tumor-rejection antigen, SART1259, possessing tumor epitopes capable of inducing cytotoxic T lymphocytes (CTLs). This study investigated the expression of SART1259 antigen in osteosarcoma and other skeletal malignant tumors to explore for a potential molecule for use in specific immunotherapy. The SART1259 antigen was detected in the cytosol fraction of 13 of 21 (62%) osteosarcoma cell lines and 3 of 8 (38%) osteosarcoma tissues, and 3 of 10 (30%) malignant fibrous histiocytoma (MFH) tissues. The HLA-A24+ and SART1259+ osteosarcoma cells were recognized by the HLA-A24 restricted and SART1 specific CTLs. These results raise a possibility that the SART1259 would be an appropriate molecule for use in specific immunotherapy of approximately one-third of HLA-A24+ patients with osteosarcoma and MFH. PMID- 10853015 TI - Expression levels of thymidine phosphorylase and dihydropyrimidine dehydrogenase in various human tumor tissues. AB - Thymidine phosphorylase (dThdPase) is the rate-limiting enzyme that metabolizes 5'-deoxy-5-fluorouridine (5'-dFUrd, doxifluridine), an intermediate metabolite of capecitabine, to the active drug 5-fluorouracil (5-FUra), while dihydropyrimidine dehydrogenase (DPD) catabolizes 5-FUra to an inactive molecule. The susceptibility of tumors to fluoropyrimidines is reported to correlate with tumor levels of these enzymes. To obtain some insight into the tumor types susceptible to fluoropyrimidine therapy, we measured expression levels of these two enzymes in various types of human cancer tissues (241 tissue samples) by the ELISA methods. DPD exists in all the cancer types studied, such as bladder, breast, cervical, colorectal, esophageal, gastric, hepatic, pancreatic, prostate, and renal cancers. Among them, the cervical, hepatic, pancreatic, esophageal, and breast cancer tissues expressed high levels of DPD (median >70 U/mg protein), while high concentrations of the dThdPase were expressed in esophageal, cervical, breast, and pancreatic cancers and hepatoma (median >150 U/mg protein). The dThdPase/DPD ratio, which was reported to correlate with the susceptibility of human cancer xenografts to capecitabine, was high in esophageal, renal, breast, colorectal, and gastric cancers (median ratio of >1.5). In any of these three parameters, the inter-patient DPD variability for each cancer type was much larger than the DPD variability among cancer types; highest/lowest ratios for dThdPase, DPD, and dThdPase/DPD were 10-321, 7-513, and 2-293, respectively. These results indicate that measurements of the three parameters, DPD, dThdPase and dThdPase/DPD, would be useful criteria for selecting cancer patients suitable for fluoropyrimidine therapy rather than for selecting cancer types. PMID- 10853016 TI - Establishment of a human colon cancer cell line (PMF-ko14) displaying highly metastatic activity. AB - A new human colon cancer cell line (PMF-ko14) derived from a peritoneal disseminated tumor has been established and maintained for over 25 months. In tissue culture, the cells grew in a mainly monolayered sheet with a population doubling time of about 27 h. Chromosome counts at the 60th passage ranged from 79 to 84 with a modal number of 83. Flow cytometry of the cell surface antigen expression indicated that CD49b, CD29, carcinoembryonic antigen (CEA), sialyl Lewis a (sLea), and CD49c were positive in more than 70% of the cells. The nude mouse xenograft models indicated are: subcutaneous or intraperitoneal injection model, spleen injection-liver metastasis model, and orthotopic implantation spontaneous metastasis model. As PMF-ko14 has highly metastatic activity it should prove to be a useful tool for research in biological behavior of metastatic colon cancer. PMID- 10853017 TI - Intratumoral expression of thymidylate synthase and dihydropyrimidine dehydrogenase in non-small cell lung cancer patients treated with 5-FU-based chemotherapy. AB - 5-fluorouracil (5-FU) has been used widely, including in gastrointestinal cancer, breast cancer, and non-small cell lung cancer (NSCLC), and its efficacy has been reported to be associated with intratumoral expression of thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD). In order to clarify the clinical value of their expression in NSCLC patients treated with 5-FU, we investigated intratumoral expression of TS and DPD immunohistochemically. Of the 68 tumors studied, 40 carcinomas (58. 8%) had high TS expression, and 33 carcinomas (48.5%) had high DPD expression. The 5-year survival rate of the patients with high-TS tumors was significantly lower than that of the patients with low-TS tumors (P=0.0267), and the 5-year survival rate of the patients with high-DPD tumors was significantly lower than that of the patients with low-DPD tumors (P=0.0268). The Cox regression analysis of prognostic variables for NSCLC patients treated with 5 FU demonstrated that the simultaneous evaluation for both TS and DPD expression was found to be a significant indicator of a poor prognosis (P=0.0043). Our results demonstrated that the evaluation of intratumoral TS and DPD activity can be used to accurately predict responsiveness to 5-FU-based chemotherapy in NSCLC patients. PMID- 10853018 TI - Stromal sialyl Le(a) expression is correlated with vascular invasion of human gallbladder adenocarcinoma. AB - Sialyl Le(a) antigen (CA19-9), a member of a family of high molecular weight glycoproteins, was originally described as a gastrointestinal- and pancreatic specific tumor marker. Recent studies have demonstrated that sialyl Lea is a ligand for E-selectin and may play an important role in tumor metastasis. However, expression patterns of sialyl Le(a) have not yet been established in human gallbladder carcinomas. In this study, we examined sialyl Le(a) expression in human gallbladder adenocarcinoma and its clinicopathological significance. Sialyl Le(a) immunoreactivity was detected not only in cancer cells (cytoplasmic type; 68.5%, 37/54) but also in cancer stroma (stromal type; 46.3%, 24/54). According to TNM classification, stromal sialyl Le(a) expression was detected in 60. 0% (24/40) and 7.1% (1/14) of the T2-4 and T1 cancers, respectively (p<0.01). Stromal sialyl Le(a)-positive gallbladder cancers frequently showed lymphatic invasion, venous invasion and lymph node metastasis (62.9%, 62.5% and 70.0%, respectively) (p<0.01). These observations suggested that sialyl Le(a) expression plays important roles in vascular invasion and metastasis of human gallbladder adenocarcinomas. PMID- 10853019 TI - A novel role for the urokinase-type plasminogen activator receptor in apoptosis of malignant gliomas. AB - Glioblastomas express more urokinase-type plasminogen activator receptor (uPAR) than do low-grade gliomas and normal brain tissue. We previously showed that downregulation of uPAR through the transfection of SNB19 cells with an antisense cDNA construct corresponding to 300 bp of the 5' end of the human uPAR gene inhibited tumor cell invasion in vitro and tumor formation in vivo. Here we sought to determine whether uPAR is necessary for cell survival and whether the inhibition of tumor formation in nude mice is due to apoptosis of intracerebrally injected SNB19 cells. Apoptosis measured by DNA fragmentation were higher in the brains of animals injected with the antisense stable transfectants than in those injected with the parental cells. Moreover, the increase in apoptotic cell death in vitro was associated with increased expression of apoptotic protein BAX in antisense clones compared to controls. To our knowledge, this is the first report of uPAR playing a novel role in cell survival in human gliomas. PMID- 10853020 TI - Gene amplification at chromosome 1pter-p33 including the genes PAX7 and ENO1 in squamous cell lung carcinoma. AB - Gene amplification is a frequent event in lung cancer, specifically in squamous cell lung carcinoma. Recently, we reported amplifications on chromosomal bands 3q26.1-q26.3 with the genes BCHE and SLC2A2 amplified in 40% of squamous cell lung carcinomas. Here, we identified an amplified domain within chromosomal bands 1pter-p33 in squamous cell lung carcinoma using reverse chromosome painting. A panel of nine genes which have previously been assigned to region 1pter-p33 was tested for amplification using comparative PCR. The ENO1 gene that encodes enolase and the PAX7 gene that encodes a transcription factor were most frequently amplified. Specifically, the gene ENO1 was amplified in six and the gene PAX7 in five out of 37 cases which included both biopsies and paraffin embedded tissues of squamous cell lung carcinomas. In total, we identified amplifications of at least one gene at bands 1pter-p33 in 10 out of 37 tumors (27%). Together, our data indicate that a novel and frequent amplification unit is present in squamous cell lung carcinoma with the center of the amplified domain in the vicinity of the genes PAX7 and ENO1. PMID- 10853021 TI - Distinct chemotactic and angiogenic activities of peptides derived from Kaposi's sarcoma virus encoded chemokines. AB - The vMIPs are chemokine-like proteins expressed by the Kaposi's sarcoma associated herpesvirus (KSHV/HHV8) during the lytic phase of viral infection. vMIP-I activates CCR8, a chemokine receptor expressed by Th2 lymphocytes and cultured monocytes. vMIP-II is an agonist for CCR3, a receptor expressed by eosinophils, and an antagonist for several other chemokine receptors. Both are highly angiogenic in the chick chorio-allantoic membrane. We designed and tested three 26-mer peptides, derived from vMIP-I (pK-I), from vMIP-II (pK-II) and from the control MIP-1alpha (pM), spanning key residues of chemokines. pK-I, pK-II and pM all were able to activate a strong chemotactic response in monocytes, higher than parental vMIP-I and II. This corresponded to induction of calcium fluxes in these cells, typical of chemokines. Interestingly, pK-II and pM were also active on PMN neutrophils. In vivo studies (matrigel sponge and rabbit cornea models) showed that pK-I retains the strong angiogenic potential exerted by vMIP-I, while pK-II and pM induced an inflammatory response, probably mediated by PMN recruitment. Our observations indicate that chemokine-derived peptides can show biological activity at pharmacological concentrations. pK-I, in particular, displays the angiogenic activity of full-length vMIP-I, while all peptides appear to have acquired additional properties, stimulating new cellular targets. PMID- 10853022 TI - Expression of interferon alpha/beta receptor in human hepatocellular carcinoma. AB - Interferon-alpha (IFNalpha) plays a crucial role in the antiproliferation and immunoregulatory activity through the specific cell surface receptor, interferon alpha/beta receptor (IFNalpha/betaR). We examined the immunohistochemical expression of IFNalpha/betaR in 91 hepatocellular carcinoma (HCC), HCV-related chronic hepatitis (n=38) and cirrhosis (n=53), dysplastic nodules (n=5), and normal liver (n=9). The level of IFNalpha/betaR increased in chronic hepatitis and cirrhosis compared with normal liver. All the dysplastic nodules showed moderate or high expression. In HCCs, 26% (24/91) of patients showed high IFNalpha/betaR expression while the remaining 38% (35/91) showed moderate, and 35% (32/91) no or faint expression. Clinicopathological survey demonstrated a significant correlation between IFNalpha/betaR expression and differentiation of carcinoma (P=0.0008) although there was no correlation between IFNalpha/betaR expression in HCC and survival or disease-free survival. Thus, IFNalpha/betaR was expressed not only in chronic hepatitis or liver cirrhosis but in HCC and its expression was significantly correlated with tissue differentiation of carcinoma. PMID- 10853023 TI - Oncogenic transformation increases the sensitivity for apoptosis induction by inhibitors of macromolecular synthesis. AB - Inhibition of RNA or protein synthesis causes apoptosis in fibroblasts. This points to the constitutive expression of a long-lived apoptosis machinery which is controlled by shortlived negative regulatory proteins, termed endogenous survival factors. The length of time between addition of the inhibitor of macromolecular synthesis and the onset of apoptosis can be used as an estimation of the effective survival factor concentration. Transformation of rat fibroblasts by a constitutively expressed src oncogene or an inducible ras oncogene increases the sensitivity for apoptosis induction by inhibitors of macromolecular synthesis, indicating that their endogenous survival factor pool has been decreased. PMID- 10853024 TI - Reversible and oxidative inactivation of endogenous survival factors after removal of exogenous survival factors. AB - Serum contains exogenous survival factors for fibroblasts. Removal of these exogenous factors causes inactivation of endogenous survival factors, subsequent release of a constitutively expressed apoptosis machinery from negative control and apoptosis. Inactivation of endogenous survival factors after serum withdrawal is mediated by reactive oxygen species and can be rapidly reversed through re addition of serum. Oxidative inactivation of endogenous survival factors after depletion of exogenous survival factors as well as the process of reversion to the active state do not require protein synthesis, indicating the existence of a constitutively expressed regulatory system that controls the interaction of exogenous and endogenous survival factors. PMID- 10853025 TI - Detection of c-erbB-2 and FGF-3 (INT-2) gene amplification in epithelial ovarian cancer. AB - We examined gene amplifications of the c-erbB-2 and FGF-3 gene in 48 epithelial ovarian cancers by differential PCR and addressed their association with clinico pathological features including clinical outcome. Overall, 25.0 and 20.8% of ovarian cancers displayed amplified c-erbB-2 or FGF-3 gene, respectively. Amplification of the c-erbB-2 gene was significantly associated with particular histological cell types, higher histological grade, and low levels of serous CA125. However, there was no correlation between c-erbB-2 gene amplification and patient outcome. No correlation was observed between FGF-3 gene amplification and any clinico-pathological features including overall survival. These findings suggested that c-erbB-2 or FGF-3 gene amplification might be one of the oncogenic events implicated in the development of ovarian cancers, yet is not a useful prognostic marker. PMID- 10853026 TI - Overexpression of manganese superoxide dismutase mRNA may correlate with aggressiveness in gastric and colorectal adenocarcinomas. AB - The expression or activity of manganese superoxide dismutase (Mn-SOD) is reduced in a variety of malignant tumors and Mn-SOD may act as a new type of tumor suppressor gene. On the other hand, increased expression of Mn-SOD can diminish the cytotoxic effects of several anticancer modalities, including tumor necrosis factor alpha, ionizing radiation, certain chemotherapeutic agents and hyperthermia. Although Mn-SOD expression and its role in various cancers are intensely studied, little is known about its function in gastrointestinal carcinomas. To examine the expression level and significance of Mn-SOD in gastrointestinal carcinomas, Mn-SOD mRNA expression was examined in 53 gastric carcinoma and 38 colorectal carcinoma by reverse transcription-polymerase chain reaction and was compared with those in the corresponding normal mucosal tissues. The tumor/normal (T/N) ratio was calculated and the data were clinicopathologically analyzed. The average T/N ratios of Mn-SOD mRNA expression in gastric and colorectal carcinomas were 2.19 and 3. 72, respectively. Clinicopathologic analyses revealed positive correlation between the Mn-SOD expression level and venous invasion in both gastric and colorectal carcinomas (p<0.05 and p<0.05, respectively). Furthermore, the colorectal carcinoma with lymph node metastasis showed significantly higher Mn-SOD expression than those without it (p<0.05). Our results suggest that Mn-SOD mRNA overexpression can occur in gastric and colorectal carcinomas and may be related to increased aggressiveness. PMID- 10853027 TI - Correlation of non-random chromosomal aberrations in lymphocytes of prostate cancer patients with specific clinical parameters. AB - The purpose of this research was to correlate non-random chromosomal aberrations in the peripheral blood lymphocytes (PBLs) of prostate cancer patients with specific clinical parameters. Peripheral blood samples were analyzed from 59 informative prostate cancer patients. Non-random chromosomal alterations detected in the PBLs and their correlation with any specific clinical parameters were analyzed statistically. A comparison was made between specific chromosomal abnormalities in the patients having an early (<65 years) or late (> or =65 years) age at disease onset, low-grade (Gleason grade <7) or high-grade (Gleason grade > or =7) tumors, a low (<10 ng/ml) or high (> or =10 ng/ml) prostate specific antigen (PSA) level, and androgen-sensitive or -insensitive disease. In examining the specific chromosomal breakpoints, the regions 1p13, 2q21, 3p21, 4q13, 5q31, 6p21, 7p15, 7p13, 7q32, 10p11, 10q26, 11p15, 11p11, 14q12, and 16q12 showed breaks in at least four cases. Chromosome 15 (P=0. 045) was significantly altered in patients having a PSA value greater than or equal to 10, while it (P=0.017) and chromosome 19 (P=0.036) were significantly altered in patients having a PSA value greater than or equal to 20. In addition, chromosomes 5 (P=0.032), 8 (P=0.020), 16 (P=0.009), and 20 (P=0.047) were significantly altered in patients having a Gleason grade greater than 7. Also, chromosomes 2 (P=0.020) and 3 (P=0.044) were significantly altered in patients who had early disease onset. Additionally, chromosome 10 (P=0.041) was significantly altered in patients having metastasis, and chromosomes 4 (P=0.006) and 7 (P=0.028) were significantly altered in patients having androgen-insensitive disease. In spite of the small subset of patients, chromosome 8 (p=0.003) was significantly altered in patients having small cell carcinoma of the prostate. From these results we conclude that non-random chromosomal aberrations present in PBLs of prostate cancer patients can be correlated with specific clinical parameters. These correlations can be used to identify a prostate cancer patient's risk response to therapy. PMID- 10853028 TI - A new human pleomorphic rhabdomyosarcoma cell-line, HS-RMS-1, exhibiting MyoD1 and myogenin. AB - A number of human cell lines derived from alveolar or embryonal rhabdomyosarcoma (RMS) have been described. To our knowledge, however, no cell line established from pleomorphic RMS has been reported. We describe here the establishment and characterization of a new human cell line, HS-RMS-1, which originated from a typical pleomorphic RMS arising in the gluteal muscle of a 26-year-old man. HS RMS-1 cells had pseudotetraploid complex karyotypes with no specific abnormalities. Both in vitro and in vivo the cells on light microscopic examination exhibited pleomorphic features with immunopositive reaction for myogenic antigens including MyoD1 and myogenin, although no Z band-like structures were detected electron-microscopically. RT-PCR demonstrated the expression of MyoD1 and myogenin in HS-RMS-1 cells at the mRNA level, and direct sequencing analysis revealed cDNAs of MyoD1 and myogenin identical to those previously reported. This cell line, HS-RMS-1, established from pleomophic RMS will be useful for further studies including the molecular aspects of human RMS. PMID- 10853029 TI - Relation between food habits and p53 mutational spectrum in gastric cancer patients. AB - p53 tumour suppressor gene mutations were analysed in gastric cancer in relation to food habits and social class in 56 patients from a high risk region of Italy. Exons 5-8 were analysed with DGGE method on amplified DNA from formalin-fixed paraffin-embedded samples. All p53 mutations were observed in patients belonging to low social class and the majority of mutations were found in intestinal type cancers. A positive association was also found with low raw vegetables, fresh, dried and preserved fruits, and ascorbic acid intake. Moreover, specific types of mutations were significantly associated with particular factors, thus suggesting the presence of specific molecular etiologic process in stomach carcinogenesis. PMID- 10853030 TI - Effect of p16INK4a on chemosensitivity in nasopharyngeal carcinoma cells. AB - The p16INK4a tumor suppressor gene is frequently inactivated in nasopharyngeal carcinoma (NPC) and hence it may play an important role in the suppression of this tumor. In order to study the effect of p16INK4a restoration in NPC cells, full-length human p16INK4a gene was transfected into a NPC cell line, CNE1. Four individual clones with differential levels of p16INK4a protein expression, were selected for further studies. The introduction of p16INK4a into CNE1 cells induced growth suppression through G0/G1 cell cycle arrest; however, the cell growth rate was not correlated to the levels of p16INK4a protein expression. To study whether transfection of p16INK4a could protect NPC cells from radiation, cisplatin and 5-fluorouracil (5FU), the cellular sensitivity of p16INK4a transfectants and vector control were investigated. An increase in sensitivity to 5FU was observed (2-fold compared to IC50) in all 4 clones compared to vector transfected control. P16INK4a transfection also resulted in increased sensitivity to cisplatin (1.5-1.8-fold) in 3 out of 4 cell lines. However, no difference in radiosensitivity was found between the p16INK4a transfectants and the control. These findings indicate that p16INK4a suppresses NPC cell growth through G0/G1 arrest and modulating cellular response to chemotherapeutic drugs in NPC cells. Therefore, restoration of p16INK4a may have a therapeutic purpose in the treatment of NPC. PMID- 10853031 TI - Biological properties and expression of mucins in 5-fluorouracil resistant HT29 human colon cancer cells. AB - We have previously reported that HT29 human colon cancer cells selected by adaptation to 5-fluorouracil (5FU) (HT29-5FU cells) express increased levels of a major intestinal mucin MUC2 mRNA compared with parental HT29 cells. In this study, we examined in detail the changes in synthesis and secretion of mucin that occur in these cells and accompanying changes in the expression of cancer associated mucin related carbohydrate antigens and cell lineage associated biochemical markers. We further investigated their relationship to biological properties of cells. Northern blot analysis revealed a markedly increased level of MUC2 mRNA but no significant change in the mRNA levels of other mucins in HT29 5FU cells compared with parental HT29 cells. Labeling with radiolabeled precursors demonstrated increased synthesis and secretion of mucin glycoproteins by HT29-5FU cells. Immunoblot analysis showed a higher expression of mucin associated carbohydrate antigens such as T, Tn, sialyl Tn, sialyl Lea, sialyl Lex and non-O-acetylated sialic acid concomitant with significant increases in the expression of goblet cell lineage marker, MUC2 apomucin and a panepithelial cell marker, carcinoembryonic antigen. HT29-5FU cells showed significantly higher adhesion to E-selectin and to matrigel and in vitro invasive properties and significantly increased liver colonization capacity in nude mice following splenic vein injection. Nude mouse xenograft tumors produced by HT29-5FU cells showed a greater degree of differentiation, consisting of mucin secreting glands than those produced by parental HT29 cells. These results indicate that predominantly colonic type mucin, MUC2, has been selectively induced in HT29-5FU cells and that altered regulation of mucin genes associated with altered synthesis and secretion of mucin glycoproteins and the degree of differentiation in cancer cells may be responsible for the altered biological properties of these cells. PMID- 10853032 TI - Clinical usefulness of serum and plasma vascular endothelial growth factor in cancer patients: which is the optimal specimen? AB - Vascular endothelial growth factor (VEGF) is secreted by various human cancer cells and plays a key role in cancer angiogenesis and metastasis. Recently, evidence of VEGF storage in blood cells including platelets has been reported. The serum VEGF levels were reported to increase during clotting as a result of its release from platelets, and plasma sample instead of serum was recommended for measuring the circulating VEGF more accurately. However, platelets have been implicated in tumor metastasis since circulating tumor cells forming aggregates with platelets were observed. The purpose of this study was to clarify which is an optimal specimen to measure VEGF in cancer patients, serum or plasma. We measured serum and plasma VEGF levels and platelet counts in 173 cancer patients and 42 healthy people, and found that serum VEGF levels were significantly higher than matched plasma VEGF and the VEGF difference (serum VEGF - plasma VEGF) correlated with platelet counts (r=0.624, p<0.05) in both cancer patients and healthy controls. We selected cancer patients with normal platelet counts (130 400x103/microl, Plt-normal cancer group). Interestingly, serum VEGF levels were higher in Plt-normal cancer group than in healthy controls. The theoretical platelet-derived VEGF in serum, calculated based on actual blood platelet counts (pg per 106 platelets), was also significantly higher in Plt-normal cancer group than in normal controls. It is, therefore, suggested that, although the serum VEGF levels are affected by blood platelets, platelet-derived VEGF also reflect biology of cancer cells, and that serum would be the more useful specimen for measurement of circulating VEGF in cancer patients for prognosis. PMID- 10853033 TI - Application of the enrichment approach to identify putative markers of response to 5-fluorouracil therapy in advanced colorectal carcinomas. AB - A wide range of tumor response is seen amongst patients with the same stage of colorectal cancer, even with the use of uniform chemotherapy. The significant economic and personal impact of chemotherapy provides the impetus for the identification of markers of response for use in guiding patient treatment. However, practical constraints prevent evaluation of all putative markers in a definitive manner. In this study, the enrichment approach was evaluated by examining the expression of a panel of putative response markers in selected patient populations with advanced colorectal cancer (i.e., those demonstrating the best and the poorest clinical response to a standardized 5 fluorouracil/folinic acid chemotherapy regimen). Patients showing a good response had a significantly increased survival when compared with poor responders (P=0.0013). Markers were then ranked for clinical importance based on differences in expression between the two groups. This allows for the relatively rapid and inexpensive investigation of multiple markers, using defined patient groups. Bcl 2 overexpression in primary colorectal tumor specimens was found to correlate with clinical response of metastatic deposits to chemotherapy (P=0.044), as did the site of the primary tumor (P=0.011). However, no clear association was observed between response status and the other examined factors (p53, PCNA, TP, MMPs 1, 2 or 9, TIMPs 1 or 2, TS, Dukes' stage at initial diagnosis, histological grade, sex or age). This approach has allowed prioritization of markers of clinical response on which larger, statistically definitive studies will be performed. PMID- 10853034 TI - Alteration of tumor phenotypes of B16 melanoma after genetic remodeling of the ganglioside profile. AB - Gangliosides have been thought to be tumor markers of various neuroectoderm derived cancers. However, their roles in malignant phenotypes of cancer cells are not well understood. We have performed the remodeling of ganglioside profiles of mouse melanoma B16 (B78 subline) by introducing GM2/GD2 synthase cDNA, and analyzed the phenotypic changes such as cell morphology, growth, adhesion and c fos gene expression. Although newly expressed GM2 was clearly detected, GM2 positive transfectant cells showed rather reduced growth rates in vivo and in vitro, lower expression levels of c-fos gene and increased levels of cell adhesion to extracellular matrices compared to control cells. These results suggested that GM2 is involved in the regulation of cell-cell or cell extracellular matrix interaction, and negatively control cell proliferation. PMID- 10853035 TI - Expression of Lewis carbohydrate antigens and chromogranin A in human prostatic cancer. AB - The prostate specific antigen (PSA) content, the neuroendocrine differentiation and the Lewis(y) and the expression of related carbohydrate antigens in pathological prostatic tissues were determined. These included 13 cancers and 11 benign hyperplasias. PSA is expressed strongly in hyperplastic and poorly in neoplastic tissues. The neuroendocrine differentiation detected by a monoclonal antibody directed against chromogranin A (CgA) is a frequent event in carcinomas and rare in hyperplastic prostate. The Lewis(y) and related carbohydrate antigens, evaluated by the reactivity of the tissues to two monoclonal antibodies MAbB3 and MAbB1, are expressed in a considerable percentage in malignant tissues of prostate and only occasionally in benign lesions. Our results suggest that immunoblotting with antibodies against CgA, B3 and B1 on the tissues, obtained after surgery, may be useful to obtain more information on the neoplastic transformation of human prostate. Furthermore, the expression of Lewis(y) and related carbohydrate antigens on the surface of prostate cancer suggest that, following a clinical trial, an immunotoxin combination of MAbB3 or MAbB1 and Pseudomonas exotoxin, may be used in the treatment of prostate cancer. PMID- 10853036 TI - Chemoprevention of colon cancer carcinogenesis by balsalazide: inhibition of azoxymethane-induced aberrant crypt formation in the rat colon and intestinal tumor formation in the B6-Min/+ mouse. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) including aspirin have been shown to suppress colon carcinogenesis and in some cases reduce the size of colorectal polyps. Balsalazide disodium (BSZ) is a colon-specific prodrug of the salicylate, 5-aminosalicylic acid. The aim of the present study was to test the chemopreventive activity of BSZ in two established animal models of colon tumorigenesis, azoxymethane-induced aberrant crypt formation in the rat and intestinal tumor formation in the B6-Min/+ mouse. Aberrant crypt foci (ACF) were induced in Fischer 344 rats via 2 subcutaneous injections of azoxymethane (20 mg/kg). BSZ was supplied in the drinking water for 8 weeks and ACF quantitated. B6-Min/+ mice were treated from 55 days of age for 90 days and intestinal tumors scored for number, size and location. BSZ treatment of AOM-injected rats reduced ACF formation in a dose-dependent manner by 60% with the greatest effect observed on ACF with 4 or more crypts. In B6-Min/+ mice a dose-dependent reduction of intestinal tumor number was observed which reached 80% in the distal small intestine and colon. A preliminary mechanistic study in cultured human colon cancer cells showed that both BSZ and 5-ASA inhibited colon cancer cell proliferation in vitro. However, 5-ASA but not BSZ produced changes consistent with the induction of apoptosis. BSZ produces a dose-dependent chemopreventive effect on colon carcinogenesis. A possible mechanism is consistent with the inhibition of cellular proliferation and the induction of apoptosis. PMID- 10853037 TI - Adrenalectomy for isolated adrenal metastases from non-adrenal cancer. AB - Adrenalectomy for metastatic cancer is rarely performed. The survival benefit for patients undergoing resection of isolated adrenal metastases is not clear. The goal of this study was to compile a series of such cases from national and international sources and examine patient survival. The patient series was derived from published series and case reports, plus eight new cases from an international registry of patients. We found 77 patients. We examined the effect of primary tumor site, metastasis size, and disease-free interval on postoperative survival duration, including only cases where complete resection with negative margins was achieved. We compared these patients with a large series from Memorial Sloan-Kettering Cancer Center (N=37). The median survival time after adrenalectomy was 23 months, with an operative mortality rate of 3.9%. There was a significant difference in survival duration depending on primary tumor site. A longer disease-free interval from time of primary cancer therapy to adrenal metastasis was associated with a longer postoperative survival after adrenalectomy. Metastasis size did not affect survival. Survival times for USA and non-USA patients were similar. Survival duration of the 77 analytical patients was similar to that of the 37 non-analytical patients from Memorial Sloan-Kettering Cancer Center. Selected patients, particularly those with long disease-free intervals and favorable tumor biology, should be offered resection for isolated adrenal metastases. PMID- 10853038 TI - Binding of the growth suppressor p53 protein to the cell cycle regulator phosphatase cdc25C. AB - Human p53 is a growth suppressor which not only functions in mammalian cells but also in fission yeast. It was previously shown that the cell cycle regulating phosphatase cdc25C suppresses the p53 induced growth arrest in fission yeast. In the present study we analysed the mechanism of this suppression. We found that cdc25C directly interacts with p53. By using different deletion mutants the binding region was narrowed down on the polypeptide chain of p53 to amino acids 287-340. To test the functional significance we analysed the effect of this interaction on the DNA binding activity of p53. As shown by band shift experiments binding of cdc25C to p53 does not modify the DNA binding activity of p53. Our data suggest that the observed suppression of the p53 induced growth arrest by cdc25C might be achieved by direct binding of cdc25C to the C-terminus of p53. PMID- 10853039 TI - Medical services of Croat people in Bosnia and Herzegovina during 1992-1995 war: losses, adaptation, organization, and transformation. AB - During the 1992-1995 war in Bosnia and Herzegovina (BH), Croatian people in BH had 19,600 (2.6%) killed and 135,000 (17.6%) displaced persons, and 222,500 (28.9%) refugees. They lost around two thirds of both physicians and other health personnel, and were left with 8. 5% of prewar patient beds. Fortunately, the organized defence against Serbs was initiated in time and Croats defended the territories where they formed majority. The first defense unit established was the Medical Corps Headquarters (MCH), caring for soldiers and civilians alike. The MCH was soon incorporated in the Croatian Defense Council (CDC, armed forces of Croatian people in BH). The MCH had two chains of command. One went through the district commanders of medical services and their subordinated physicians to paramedics in military units, and the other directly to the commanders of 14 war hospitals. After its formation in 1993, the Ministry of Health took the jurisdiction over the civilian medical services and after the Washington Peace Agreement (April 1994) over the war hospitals, too, whereas the medical services within military units remained under control of the Ministry of Defense. Dayton Peace Agreement divided BH into the Federation of BH and Republic Srpska, each with their own army. The Federation of BH Army is composed of the CDC and Bosniac controlled Army of BH, with overall numerical ratio 1:2.3 for Bosniacs, and organized in accordance with NATO standards. Military medical services are provided by the Logistics Sector of both Ministry of Defense and Military Corps Headquarters (Joint Command). PMID- 10853040 TI - From service to commodity: corporization, competition, commodification, and customer culture transforms health care. AB - Corporate medical practice, a market economy, and a consumer culture are transforming health care. The service relationships of doctors with patients are now commodities. The doctor, directed by disease management protocols (to improve outcomes, reduce costs, and standardize care), is, in effect, providing programmed service commodities. In addition, medical-surgical specialties, now "packaged" for the care of body parts and conditions (as Breast, Stroke, Obesity, Aneurysm Centers), are also made service commodities, marketed by newspaper advertisements, TV, radio, and Internet to patient-customers in search of a healthy body. In sum, the promise of corporate practice in a competitive market economy is greater efficiency and productivity to reduce the costs of care that are a burden on industries and the state. Viewed from office encounters with patients, such transformation of services to commodities changes the doctor patient relationship and the moral mission of care. PMID- 10853041 TI - Decisions at the end of life. AB - This paper presents a system for making decisions at the end of life. It emphasizes the role of patient autonomy and the importance of patient and family participation with the physician in decision-making. Definitions are presented for the terms: terminal illness, withholding and withdrawing life sustaining treatment, physician assisted suicide and euthanasia. Three cases are briefly described to illustrate the application of the decision-making system. A detailed discussion is then presented of the divergent views expressed by different authors about the moral differences or similarities between foregoing life sustaining treatment and physician assistance in dying. It is concluded that the view that these two actions are fundamentally different, as supported by the United States Supreme Court, in 1997, is the correct one. Physician assisted suicide (PAS) remains a controversial issue. Physicians and societies in individual countries must work out their own approaches to PAS. However, foregoing invasive or intensive life support in terminally ill patients consistent with their wishes is considered appropriate. PMID- 10853042 TI - Small countries and the dioxin scandal: how to control imported food? AB - The aspiration of the countries in transition to join the developed European countries resulted in opening their borders and several-fold increase in import, especially of food products. The imported foods are less expensive than domestic foods, but their quality is often highly questionable. In analyzing the safety of these products for human health, small countries encounter at least two sets of problems. One is related to legal provisions on the parameters to be analyzed, whenever new requirements emerge in practice, like the latest one on dioxin. The other, even more difficult set of problems, is related to the expensive equipment needed for the monitoring of foodstuff safety, the procurement of which exceeds the financial possibilities of these countries. For example, from June 11 until July 31, 1999, during the so-called European dioxin crisis, a total of 58 foodstuffs produced in Belgium, Netherlands, and France between January 19 and March 3, 1999, were referred to the Department of Health Ecology, Zagreb Institute of Public Health, the only laboratory authorized for identification of polychlorinated dibenzodioxins and dibenzofurans in the Republic of Croatia. In 40 samples, the level of dioxin was below the detection limit of 0.5 ng - international toxic equivalents per kg fat (ng-I-TEQ/kg fat), whereas in 18 positive samples the level of dioxin did not exceed the limit of 5 ng-I-TEQ/kg fat for the foodstuff commercial usability. Although highly contaminated products have not yet appeared on the Croatian market, recent developments in Europe have pointed out that establishing an authorized laboratory for dioxins in the Republic of Croatia or in the region is needed. PMID- 10853044 TI - Histopathological features of gastritis before and after treatment for Helicobacter pylori. AB - AIM: To assess five main histological features of gastritis in gastric mucosa colonized with Helicobacter pylori before and after the treatment. METHODS: Histologic assessment of H. pylori-associated gastritis was performed according to the Sydney classification before and after the treatment in 97 patients. Two additional parameters - the presence of lymphocytic aggregates and coccoid forms of bacteria - were also analyzed. Helical and coccoid forms of H. pylori were detected by immunohistochemistry in biopsies after the treatment. RESULTS: Whereas acute epithelial damage was quickly repaired, some of the local responses to bacteria, e.g., lymphoid aggregates and intestinal metaplasia, persisted after treatment. Higher H. pylori and cocci density was found before and after treatment in patients with intestinal metaplasia (p=0.020). Correlation between H. pylori and mucosal atrophy was found only after treatment (p=0.009). Immunohistochemical staining was more sensitive in detecting of H. pylori than Giemsa staining (p=0.007) in cases where, using only Giemsa staining, it was not possible to distinguish coccoid forms of H. pylori from other cocci. CONCLUSION: After treatment, H. pylori-associated gastritis showed reduction of acute and chronic inflammation, but lymphoid aggregates and intestinal metaplasia persisted. Immunohistochemistry of different forms of H. pylori may be a valuable technique in monitoring the success of the treatment. PMID- 10853043 TI - Prognostic significance of DNA ploidy pattern and nucleolar organizer regions (AgNOR) in colorectal carcinoma. AB - AIM: To investigate the prognostic significance of DNA ploidy and silver stained nucleolar organizer regions (AgNOR), as well as their relation to the histological grade and Dukes' stages of colorectal carcinomas, and the relation of tumor cells proportion in the S-phase and Dukes' stage, histologic grade, DNA ploidy, or AgNOR count. METHODS: DNA flow cytometric analysis and AgNOR were performed on 94 surgically removed colorectal carcinomas. The mean AgNOR count was calculated in 200 tumor cells for each case. Survival rates and tests for significance were evaluated using the log-rank test and Cox regression model. RESULTS: There were no significant correlations between the ploidy pattern, histological grade, and Dukes' stage. Diploid tumors had a significantly lower AgNOR count (median 2.5, range 2.1-7.7) than aneuploid (median 6.2, range 2.0 7.9). Dukes' C stage tumors exhibited higher AgNOR count than Dukes' A or B stages. The proportion of tumor cells in S-phase did not correlate with any other parameter. Each of these parameters failed to show any correlation with survival. After dividing the tumors into those with high (>5) and low AgNOR count (<5), no correlation was found in the latter group between AgNOR and any other studied parameters, whereas in the group with high AgNOR count correlations to Dukes' stage, DNA ploidy, and histological grade were established.Conclusions. The difference in survival between well, moderately, and poorly differentiated tumors were significant in the group with high AgNOR counts. Dukes' C stage and aneuploid tumors had the worst prognosis. PMID- 10853045 TI - Comparison between critical pathway guidelines and management of deep-vein thrombosis: retrospective cohort study. AB - AIM: To compare the key steps of standard deep-vein thrombosis management with the critical pathway practice guidelines, and to assess the outcome of the treatment after 6 months. METHOD: This retrospective cohort study (from January 1, 1997 to December 31, 1998) included 172 patients with uncomplicated deep-vein thrombosis of lower extremities, consecutively admitted via emergency room. The data were collected from the entry register in emergency room and medical charts. The outcome of therapy was assessed 6 months after the acute event. RESULTS: A bolus dose of heparin was administered to 81 (46%) patients. The recommended initial heparin infusion rate at 1250 U/h was employed in only 26 (15%) patients. Time to activated partial thromboplastin time >60 s was met in 29 (17%) patients. All patients but one received heparin therapy longer than 96 h. The recommended time to a therapeutic international normalized ratio of less than 120 h was achieved in 134 (78%) patients, but the average length of a stay in the hospital exceeded the recommended 5. 5 days by 86%. Six months later, compressive ultrasonography revealed 44 (28.9%) cases of complete vein obstruction, 67 (44.1%) cases of partial recanalization and 41 (27%) cases with a normal finding. Recurrent thrombosis developed in 16 patients (10.5%) and acute pulmonary embolism in 4 (2.6%) patients. CONCLUSION: Our results considerably differ from the critical pathway guidelines due to the lower initial heparin doses and longer diagnostic assessment of thrombosis etiology. Our approach to deep-vein thrombosis treatment was a combination of the critical pathway guidelines and the conventional regimen. The clinical outcome in our series did not differ significantly from the outcome after the conventional way of treatment. PMID- 10853046 TI - Survival of cementless and cemented porous-coated anatomic knee replacements: retrospective cohort study. AB - AIM: To evaluate the effect of cement use in porous-coated anatomic (PCA) total knee prosthesis on its survival. METHODS: The study was a retrospective analysis of 142 PCA total condylar arthroplasties performed in 124 patients from 1985 to 1991. Uncemented prostheses were used in 87 knees, the prostheses were cemented in 44 knees, and hybrid prosthesis components were used in 11 knees. The average follow-up time was 88 months (range 66-140). The survival of prosthesis was assessed using the Kaplan-Meier's method. The Baltimore score was evaluated as a measure of clinical performance in 115 replacements. RESULTS: The overall cumulative survival rate of a PCA total knee prosthesis was 77% over the average follow-up period of 88 months. No significant differences in survival rates among cementless, cemented, or hybrid fixations could be demonstrated. The survival rate of the prostheses in patients with rheumatoid arthritis (82.5%) was significantly higher than in patients with osteoarthritis (73.8%). Revision was necessary in 29 (20.4%) replacements. CONCLUSION: The survival of PCA endoprosthesis, regardless of the components used for implantation, is not satisfactory. PMID- 10853047 TI - Comorbidity of posttraumatic stress disorder and alcohol dependence in displaced persons. AB - AIM: To investigate in displaced persons a) the prevalence rate of current posttraumatic stress disorder (PTSD) and alcohol dependence; b) the relationship of alcohol dependence and current PTSD; and c) trauma exposure in relation to alcohol dependence comorbid to PTSD. METHODS: A group of displaced persons (157 men and 211 women) was interviewed using structured clinical interview based on DSM-III-R criteria for diagnosing PTSD and alcohol dependence, Watson's PTSD Questionnaire, and CAGE Questionnaire. RESULTS: Men showed higher prevalence rate of a current PTSD (50.3% of men vs. 36.5% of women, p=0.011), alcohol dependence (60.5% of men vs. 8.1% of women, p<0. 001), and alcohol dependence comorbid with PTSD (69.6% of men vs. 11. 7% of women, p<0.001). The rate of alcohol dependence increased in relation to current PTSD in men but not in women. Comorbidity of alcohol dependence and PTSD in women was influenced by alcohol-related problems before the war, whereas in men it was not influenced by any of the pre-war variables. The highest number of traumas was experienced by the displaced persons with a current PTSD only, followed by those with PTSD and alcohol dependence. The lowest number of war traumas was experienced by displaced persons with alcohol dependence, but without current PTSD. CONCLUSION: War traumas may have a role in the development of alcohol dependence in displaced men with current PTSD. The number of war traumas had a strong effect on the development of PTSD. Sex is a relevant factor in studying comorbidity of current PTSD and alcohol dependence. PMID- 10853048 TI - Psychotic symptoms and comorbid psychiatric disorders in Croatian combat-related posttraumatic stress disorder patients. AB - AIM: To investigate the prevalence rate of post-traumatic stress disorder (PTSD) comorbid psychiatric disorders and to explore psychotic symptoms in patients with combat-related current PTSD. METHOD: The sample included Croatian war veterans (N=41) who were hospitalized at the University Department of Psychiatry of the Vrapee Psychiatric Hospital during the 1995-1996 period and fulfilled the DSM-IV criteria for the current and chronic PTSD. The Schedule for Affective Disorder and Schizophrenia (SADS-L) was applied for the assessment of current and lifetime psychiatric disorders. Only three subjects had a prewar Axis I psychiatric disorder. One third of the patients met the criteria for personality disorder. RESULTS: After severe combat trauma, the majority of PTSD patients (33/41) had at least one comorbid psychiatric diagnosis on Axis I. In those with personality disorders the most frequent was alcohol dependence, whereas in those without personality disorders it was major depressive disorder. Psychotic symptoms occurred in 8 out of 41 PTSD patients. None of them had a primary psychotic disorder or a personality disorder. In all the patients, psychotic symptoms were different from flashbacks. They were symbolically related to the trauma and resistant to antipsychotic treatment. Psychotic symptoms were associated with depression in 5 out of 8 patients with psychotic symptoms. CONCLUSION: Severe and prolonged combat trauma may be followed by the co-occurrence of PTSD and psychotic symptoms, forming the atypical clinical picture of PTSD. PMID- 10853049 TI - Brachial biceps tendon injuries in young female high-level tennis players. AB - AIM: To evaluate brachial biceps tendon lesions in four young female tennis players who complained about anterior shoulder pain on their dominant side. METHODS: Medical and sport's activity history, palpation of the painful zone, Ghilchrist (palm-up) test, and brachial biceps contraction against resistance were performed. RESULTS: The two girls who suffered from mild tenderness in the bicipital groove and over the anterior aspect of the upper arm and the shoulder joint, had tendinitis of the long biceps head. The two girls who suffered from severe tenderness just under the groove, had a partial tear in the long head of the biceps. Ghilchrist test was positive in all girls. CONCLUSION: Tennis players can have shoulder pain without clear history of trauma. Pain occurred probably as a result of technical errors or use of inadequate equipment. PMID- 10853050 TI - Influence of 1991-1995 war on breast-feeding in Croatia: questionnaire study. AB - AIM: To investigate the influence of 1991-1995 war on the prevalence, duration, and practice of breast-feeding Croatian children up to 5 years of age. METHOD: In 1996, interviews were conducted in households with children up to 2 years of age (757 children) and 2-5 years of age (1,180 children). Data for war-free areas, war-affected areas, and areas liberated after several years of occupation were analyzed separately. RESULTS: In 1996, 94.6% of mothers started breast-feeding, which lasted for an average of 3.4+/-2.9 months. The proportion of mothers who started breast-feeding did not vary with respect to either war-related or geographic areas of the country. Breast-feeding was significantly longer in war free than in war-affected areas (3.7+/-3.1 vs. 2.7+/-2.1 months, respectively; p=0.015). The duration of breast-feeding in Croatia's geographic regions, Istria, Hrvatsko Primorje, and Gorski Kotar, was significantly longer than in Slavonia (3.9+/-3.4 vs. 3.4+/-3.0, respectively; p=0.037). On the country level, 49.4% of babies were fed on demand and 43.3% according to a daily schedule. The percent of children who were not breast-fed was significantly higher (p=0. 002) in the older age group (2-5 years of age, 9.3%) than in the younger age group (up to 2 years of age, 5.4%). CONCLUSIONS: The war decreased the prevalence and duration of breast-feeding, which might be related to regular humanitarian donations of infant food and mother's milk substitutes, especially in the war-affected areas. UNICEF breast-feeding campaign, which started in 1993, appeared to be effective. PMID- 10853051 TI - Decreasing risk of viral transfusion-transmitted diseases in Croatia. AB - AIM: To assess the risk of viral transfusion-transmitted infections in Croatia. METHODS: The following parameters were analyzed: frequency of blood donations repeatedly reactive for HBsAg and anti-HCV (1993-1999); blood donations confirmed positive for HBsAg and anti-HCV (1997-1999), anti-HIV1/2, and syphilis reactivity (1993-1999); number of registered patients with hepatitis B and C; transfusion associated hepatitis B and hepatitis C; and frequency of HBV, HCV and HIV markers in patients with congenital bleeding disorders (1993-1998). RESULTS: The frequency of repeatedly reactive HBsAg and anti HCV markers and confirmed positive HBsAg, anti-HCV, and syphilis markers in donors blood decreased during the study, whereas the frequency of anti-HIV1/2 positivity did not change. The frequency of confirmed positive donors in 1999 was 0.068% for HBsAg, 0.035% for anti HCV, 0.002% for anti HIV1/2, and 0.0056% for syphilis. The number of patients with hepatitis B, hepatitis C, and transfusion-associated hepatitis B and C steadily decreased during the 1993-1998 period. The number of transfusion associated hepatitis patients leveled off in 1997. From the beginning of the follow-up of AIDS patients in 1987, only 7 (2%) of hemophiliacs have been HIV infected, all before 1990 and due to non-inactivated coagulation factor concentrates. There were no cases of transfusion-associated HIV2 infection in patients with congenital bleeding disorders or transfusion-associated HIV1 infection through transfusion of labile blood components. CONCLUSION: The safety of transfusion therapy in Croatia has improved, and the present risks of viral transfusion transmitted diseases are very low. PMID- 10853052 TI - Cyclic patterns of incidence variations for stomach cancer in the North-Western region of England. AB - AIM: To analyze temporal dynamics and model trends and variations of the annual incidence rates of stomach cancer in the North-Western Region of England. METHODS: The data consisted of 23,465 new cases of stomach cancer as provided by the population-based registry of the Centre for Cancer Epidemiology (Manchester, England, UK). The parameter studied was the annual incidence rate of stomach cancer per 100,000 persons as age-adjusted to the world standard population and presented as time-series over the interval from 1971 to 1990. The hypotheses to be tested, regarding the annual incidence rates, were: 1) existence of specific temporal characteristics; 2) appearance of cyclic patterns of variability; and 3) usefulness of cyclicity in predictive modeling. RESULTS: The decreasing tendency of annual incidence rates of stomach cancer for both men and women was best fitted by a quadratic trend (y=a+b?t2) out of 13 available linear/nonlinear regression models. This trend explained only about 49% of variability. Cyclic patterns of variability in incidence rates were established (short-term cycle of 8.9 years; long-term hypercycle of about 22-23 years). The best fit to the real incidence rates was achieved by bi-cyclic regression model. The summary cosine sine model contained both cycles of 8.875 and 22-23 years; it showed the least variance of regression (men - Sy2=1.09; women - Sy2=0.65) and best prognostic index (PI=1.47 for men, and PI=1.29 for women). This trigonometric model explained about 83-86% of variability of incidence rates of stomach cancer. CONCLUSION: Cyclic patterns of variability of the annual incidence rates of stomach cancer have been established. Such cyclicity might not only find likely implications in predictive modeling and forecasting of incidence rates, but it could also be considered useful in research on risk/prevention factors for stomach cancer. PMID- 10853053 TI - Effects of renal transplantation on hearing and ocular changes in a monozygotic twin with Alport's syndrome: comparison with other twin on hemodialysis. AB - AIM: To present a unique case of Alport's syndrome in monozygotic twins with two different treatment modalities - renal transplantation and hemodialysis, and to evaluate the effects of therapy on hearing and ophthalmological findings. METHODS: Pure-tone audiogram and ophthalmologic examinations were performed in both twins at the age of 30. At the age of 46, 4 years after renal transplantation in the first twin and after 6 years of hemodialysis in the second twin, both twins underwent control audiometric and ophthalmologic examinations. RESULTS: Control audiometric measurements showed the progression of bilateral sensorineural hearing loss in the high-frequency range (>2,000 Hz) in both twins. The hearing threshold progressed from initial 50 dB in both twins at the time of the diagnosis to 55 dB in the twin on hemodialysis, and 85 dB in the twin with a transplanted kidney. Retinal blurry hyperpigmentations disappeared in the twin with a transplanted kidney. CONCLUSION: In comparison with hemodialysis, renal transplantation in Alport's syndrome may have deleterious effect on hearing, when associated with plasma hyperviscosity and hyperlipidemia, but may lead to regression of retinal hyperpigmentation. PMID- 10853054 TI - Recurrent posterior dislocation following primary posterior-stabilized total knee arthroplasty. AB - In our series of 136 patients with primary total knee arthroplasty using posterior-stabilized prosthesis, a female patient with Parkinson disease developed posterior dislocation of the knee 9 months after surgery. Eventually, the dislocation became recurrent, occurring several times a day. The patient made the reposition always by herself. Two months after the first dislocation, we performed the revision of the polyethylene tibial insert and found wearing of the tibial insert's cam as an hitherto unreported cause of the mechanical instability of the total knee prosthesis. PMID- 10853055 TI - Cerebrospinal fluid seepage through polyglactin 910 dura substitute manifested as spinal extradural collection of fluid. AB - Following excision of pilocytic astrocytoma, a 12-year-old girl underwent posterior cranial fossa synthetic duraplasty with polyglactin 910 mesh. On the 8th postoperative day, unusual extradural collection was diagnosed by spinal magnetic resonance imaging. On the 14th postoperative day, cerebrospinal fluid leakage in the upper part of the postoperative wound was noticed. Unusual extradural collection detected by spinal magnetic resonance imaging was assumed to be the consequence of cerebrospinal fluid seepage and a warning sign of cerebrospinal fluid leakage following synthetic posterior fossa duraplasty. This case shows that polyglactin 910 mesh may be ineffective when used for posterior cranial fossa duraplasty in children, although it is considered as valuable as autologous tissue. PMID- 10853056 TI - History of the American Society for Stereotactic and Functional Neurosurgery. AB - The field of human stereotactic neurosurgery has just passed the half-century mark. Soon after its inception, the pioneers in the field began to meet to exchange information and ideas, which led to an international forum for stereotactic surgery. In 1973, the organization was expanded to form both the World Society for Stereotactic and Functional Neurosurgery, as well as the European and American Societies for Stereotactic and Functional Neurosurgery. Reviewing the programs of the meetings of those Societies permits one to review the nature of the information that was exchanged through the years, and, in doing so, to monitor the pulse of the field as it has developed. The first independent meeting of the American Society for Stereotactic and Functional Neurosurgery took place in Houston in 1980, at which there were 27 papers, 40% of which were on the newly emerging field of image-guided neurosurgery and the rest on classical functional neurosurgery. The five meetings since, occurring at approximately 4 year intervals, have documented the progress in epilepsy surgery, the reemergence of stereotactic surgery for movement disorders, the growth of stereotactic radiosurgery, and the genesis of frameless stereotactic techniques which have now become widespread. PMID- 10853057 TI - Stereotaxy, navigation and the temporal concatenation. AB - Nautical and cerebral navigation share similar elements of functional need and similar developmental pathways. The need for orientation necessitates the development of appropriate concepts, and such concepts are dependent on technology for practical realization. Occasionally, a concept precedes technology in time and requires periods of delay for appropriate development. A temporal concatenation exists where time allows the additive as need, concept and technology ultimately provide an endpoint of elegant solution. Nautical navigation has proceeded through periods of dead reckoning and celestial navigation to satellite orientation with associated refinements of instrumentation and charts for guidance. Cerebral navigation has progressed from craniometric orientation and burr hole mounted guidance systems to simple rectolinear and arc-centered devices based on radiographs to guidance by complex anatomical and functional maps provided as an amalgam of modern imaging modes. These maps are now augmented by complex frame and frameless systems which allow not only precise orientation, but also point and volumetric action. These complex technical modalities required and developed in part from elements of maritime navigation that have been translated to cerebral navigation in a temporal concatenation. PMID- 10853058 TI - Concordance between functional magnetic resonance imaging and intraoperative language mapping. AB - Although the correspondence between functional-magnetic resonance imaging (fMRI) representations of the sensorimotor cortex and intraoperative electrophysiology (including somatosensory evoked potential, SSEP, recordings and direct cortical stimulation) has been reported, a similar correspondence between fMRI and intraoperative localization of the language-sensitive cortex is not as well established. The aim of the present study was to evaluate the concordance between fMRI and intraoperative electrophysiology with respect to the localization of the language-sensitive and sensorimotor cortices. We present the results of 21 patients who underwent language and sensorimotor mapping by fMRI and intraoperative electrophysiology including SSEP recordings (n = 21), direct cortical stimulation of motor cortex (n = 15) and direct cortical stimulation of Broca's and Wernicke's area (n = 5). When responses were obtained with both methods, localization of function concurred in all cases. These observations suggest that fMRI represents a reliable preoperative tool for the identification of language-sensitive areas. PMID- 10853059 TI - Updated neuroimaging using intraoperative brain modeling and sparse data. AB - A strategy to update preoperative imaging for image-guided surgery using readily available intraoperative information has been developed and implemented. A patient-specific three-dimensional finite element model of the brain is generated from preoperative MRI and used to simulate deformation resulting from multiple surgical processes. Intraoperatively obtained sparse imaging data, such as from digital cameras or ultrasonography, is then used to prescribe the displacement of selected points within the model. Interpolation to the resolution of preoperative imaging may then be performed based upon the model. The algorithms for generation of the finite element model and for its subsequent deformation have been successfully validated using a pig brain model, and preliminary clinical application in the operating room has demonstrated feasibility. PMID- 10853060 TI - A system for microscope-assisted guided interventions. AB - We present a system for surgical navigation using stereo overlays in the operating microscope aligned to the operative scene. This augmented reality system provides 3D information about nearby structures and offers a significant advancement over pointer-based guidance, which provides only the location of one point and requires the surgeon to look away from the operative scene. With a previous version of this system, we demonstrated feasibility, but it became clear that to achieve convincing guidance through the magnified microscope view, a very high alignment accuracy was required. We have made progress with several aspects of the system, including automated calibration, error simulation, bone-implanted fiducials and a dental attachment for tracking. We have performed experiments to establish the visual display parameters required to perceive overlaid structures beneath the operative surface. Easy perception of real and virtual structures with the correct transparency has been demonstrated in a laboratory and through the microscope. The result is a system with a predicted accuracy of 0.9 mm and phantom errors of 0.5 mm. In clinical practice errors are 0.5-1.5 mm, rising to 2 4 mm when brain deformation occurs. PMID- 10853061 TI - Interactive image-guided surgery system with high-performance computing capabilities on low-cost workstations: a prototype. AB - We present a new frameless stereotatic system prototype that has been initially validated in functional neurosurgery operations and that makes use of an optical position tracker for image-guided neurosurgery. Several devices for tracking different surgical instruments have been designed and manufactured. These devices include an array of infrared light-emitting diodes that are tracked by three charge-coupled device cameras. The system presents several new approaches for surgery planning. For high-quality 3D images of the patient's anatomy, we have developed a parallel version of a volume-rendering algorithm, thus enabling real time 3D anatomy manipulation on low-cost PC workstations. In order to test the accuracy of the system, the localization of the target by means of a stereotatic frame has been compared with frameless techniques, obtaining a difference of about 1 +/- 1 mm. PMID- 10853062 TI - Accuracy and availability of the computed assisted neurosurgery navigation system during epilepsy surgery. AB - The magnetic-force-based Computed Assisted Neurosurgery System was used for epilepsy surgery to localize targets accurately in the operative field. The correlation between X components of target coordinates in the axial plane and the coronal plane for the same target was strong in all cases. Concerning Y components, there were statistically significant differences in 2 cases. There was a case that showed statistically significant differences only in the Z dimension. The interdisk distance by data sets of coordinates obtained from neuronavigation was calculated to quantify localization error, and the measuring error ranged from -5 to 13.3 mm (1.3 +/- 3.2 mm). The magnitude of the application errors in this study tended to be large in the frontal and vertex regions. PMID- 10853063 TI - Frameless stereotactic neurosurgery: two steps towards the Holy Grail of surgical navigation. AB - The holy grail of surgical navigation is to provide precise continuous feedback during surgery about the target and its surrounding structures. The first step was the ability of hardware and software technology to allow patient-to-image registration using a multi-potentiometer position-sensing articulated arm system. We used such a system (OAS; Radionics, Burlington, Mass., USA) in 169 consecutive patients with common intracranial lesions. We achieved a mean application accuracy of 2.5 mm, which was sufficiently reliable for most neurosurgical procedures. However, to get the feedback information, the surgeon has to look away from the operative field to the workstation monitor. As psychological studies of manual workers including surgeons indicated that performance is better when the worker is looking in a downward gaze at his hands, the natural progression was to project feedback information between the eyes and the hands. Therefore, the second step was to link tracking technology to the surgical microscope with head-up display. We used such a system (SMN-Zeiss, Germany) in 65 consecutive patients with a mean application accuracy of 1.4 mm. This was again sufficiently reliable for neuronavigation. The head-up display provided continuous feedback to the surgeon about the target, risk zones and areas of interest without the need to interrupt the procedure to get such information. Furthermore, the use of the focal length of SMN with autofocus to perform the registration improved the application accuracy of this technology. The ability of the software to process all MRI sequences (T(1), T(2), MPR and CISS) allowed us to use a variety of image sequences to delineate the lesion more exquisitely. PMID- 10853064 TI - Interventional MRI-guided stereotactic aspiration of acute/subacute intracerebral hematomas. AB - Surgical interventions for hypertensive intracerebral hematomas are still controversial. Many believe only hyperacute intervention is of any real utility. The majority of present interventions require a formal craniotomy with standard neurosurgical techniques. There are, however, a few reports on CT-guided stereotactic aspiration of these hematomas with favorable results. We report 10 patients treated with frameless fiduciless stereotactic means using an intraoperative MRI scanner (GE 0.5 T Signa SP). These patients were initially diagnosed as having hypertensive intracerebral hematoma and operated on within 1 34 days after hemorrhage. The actual operating time averaged less than 120 min, including intraoperative imaging. Clot volumes ranged from 2.5 to 75 cm(3) with a mean of 31 cm(3). There were 2 thalamic hematomas and 8 basal gangliar hematomas. Three patients had intraventricular hematoma extension and all 3, as well as an additional patient, required extraventricular drainage. However, no patients required permanent posthemorrhage ventriculoperitoneal shunting. Aspiration was successful in all cases to 70-90% of clot removal. Two cases utilized intrahematoma t-PA infusion with subsequent 80-90% clot removal. There were no complications or rehemorrhages. All patients showed some form of improvement that included either improved blood pressure control, speech or cognitive abilities. We conclude that using an intraoperative MRI scanner to perform frameless, fiduciless stereotactic aspiration of acute/subacute intracerebral hematoma is a safe and potentially effective means of treating intracerebral hematomas. PMID- 10853065 TI - The relationship of imaging techniques to the accuracy of frameless stereotaxy. AB - OBJECTIVE: We analyzed the accuracy of a frameless stereotactic system using computed tomographic (CT) and magnetic resonance imaging (MRI) scans of different slice thickness and T(1) versus T(2) weighting of MRI. METHODS: An open skull with graphite pegs fixed to its base was used for all scans. CT scans were done with slice thicknesses of 1, 2 and 3 mm. MRI-visible markers were placed on top of pegs for T(1)-weighted and T(2)-weighted MRI scans, which were acquired at thicknesses of 1.5, 3 and 5 mm. For each scan, 3 separate registrations of a probe were performed; the distance between the actual probe location and that displayed on the registered image was noted. Each measurements was repeated 3 times for each registration. RESULTS: Greatest accuracy was achieved with 3-mm slice CT scans; this was not improved by using thinner slices. T(1)-weighted 1.5 mm MRI scans were 23% less accurate and T(2)-weighted 3-mm scans 37% less accurate. CONCLUSIONS: Frameless stereotaxy should be done using CT scans when the greatest possible accuracy is desired. There appears to be no advantage to using slice thicknesses less than 3 mm. For most craniotomy applications, T(1) weighted MRI using 3-mm slices provides sufficient accuracy. Lesions imaged only on T(2)-weighted MRI also can be approached with adequate precision using 3-mm scans. PMID- 10853066 TI - Image registrations using points and surfaces simultaneously PMID- 10853067 TI - AcouStick: An optically tracked A-mode ultrasonography system for registration in image-guided neurosurgery PMID- 10853068 TI - Frameless, fiduciless stereotactic neurosurgery using interventional MRI PMID- 10853069 TI - Placement of deep brain stimulators into the subthalamic nucleus. AB - We present our technique for deep brain stimulation (DBS) of the subthalamic nucleus (STN) and include information which may be helpful in general DBS. With the patient in a stereotactic head frame, the anterior and posterior commissures are identified on SPGR-sequence magnetic resonance imaging (MRI). STN coordinates are based on a stereotactic brain atlas at 12 mm lateral, 2 mm posterior and 5 mm caudal to the midcommissural point. Surgical navigation software allows for planning of the trajectory. Electromyography is used to quantitatively measure tremor responses to macrostimulation. Permanent lead placement is confirmed with intraoperative fluoroscopy and postoperative MRI. PMID- 10853070 TI - Targeting for thalamic deep brain stimulator implantation without computer guidance: assessment of targeting accuracy. AB - The authors assess the accuracy of targeting nucleus ventralis intermedius (Vim) with fast spin echo inversion recovery (FSE/IR) magnetic resonance imaging (MRI) in 18 successful deep brain stimulator (DBS) implants for medically refractory tremor. FSE/IR-MRI-derived coordinates are compared to the final coordinates employed for DBS lead placement, selected with intraoperative neurophysiology. The authors conclude that FSE/IR MRI is sufficiently reliable to serve as the sole means of anatomically targeting Vim for DBS lead placement. An independent computer workstation is not required for accurate targeting; however, intraoperative neurophysiology remains essential. PMID- 10853071 TI - Audit of neurophysiological recording during movement disorder surgery. AB - Sixty-two cases of thalamic pallidal and subthalamic surgery in Dundee were audited to assess the influence of physiological localisation on the procedure. Methods included microelectrode recording, evoked potential and stimulation techniques. Although anatomical localisation is improving with modern techniques, the physiological information is still modifying the surgery in 67% of cases. PMID- 10853072 TI - Do microelectrode techniques increase accuracy or decrease risks in pallidotomy and deep brain stimulation? A critical review of the literature. AB - Several recent publications have stated that the use of microelectrode recording (MER) during pallidotomy or deep brain stimulation (DBS) contributes to decreasing risks and side effects of surgery, and that such a technique is a prerequisite for minimizing lesion size and for accurate placement of the stereotactic lesion or the DBS electrode. To evaluate the consistency of these statements, we reviewed hundreds of papers and congress reports on MER- and non MER-guided procedures published since 1992. This review showed that MER groups published more often than non-MER groups. While side effects of surgery were not uncommon in both groups, the rate of severe complications, such as hematoma, and mortality appeared to be higher when microelectrodes were used, both in ablative surgery and in DBS procedures. Besides, the nonaccurate placement of lesions or DBS electrodes, as assessed on published MRI figures, was not uncommon in MER publications. Lesion volume was, when reported, not different in both techniques. The electrical parameters of stimulation of implanted electrodes in the thalamic ventral intermediate (Vim) nucleus for treatment of tremor were higher in MER guided surgery. The available literature suggests that MER techniques may increase the risks of surgery without enhancing its accuracy, compared to MRI based macrostimulation techniques. To date, there is no randomized trial by one and the same group on the use of micro- versus macroelectrodes in surgery for movement disorders. A prerequisite for such a trial in the future must imply that the investigators have an equal nonprejudiced attitude towards, and equal confidence and experience in, either technique. Since such a prerequisite does not exist so far in the functional stereotactic community, a critical and comparative study of the available literature remains the only way to evaluate the pros and cons of either technique, in terms of targeting accuracy and surgical complications. PMID- 10853073 TI - Subthalamic stimulation in Parkinson's disease. Preliminary results. AB - OBJECTIVES: We wanted to evaluate chronic subthalamic nucleus (STN) stimulation as an alternative to pallidotomy for severe Parkinson's disease symptomatology. METHODS: Nine patients met clinical criteria for unilateral standard pallidotomy. All had severe medically refractory drug-induced dyskinesia and had reached maximal daily levodopa therapy. Pre- and postoperative videos, neuropsychometric testings and clinical stagings were administered. Three patients were selected to undergo stereotactic implantation of a deep brain stimulator (DBS) after Institutional Review Board approval and informed consent. These were performed using digitized microrecordings. The other group received unilateral pallidotomy. RESULTS: At a mean follow-up of 6 months, our results support recent findings of significant major improvement in motor scores, activity of daily living and decrease in amount of daily levodopa intake by close to 50% after 3 months of stimulation. CONCLUSIONS: Chronic stimulation of the STN appears to provide significant motor improvement in patients with severe Parkinson's disease and is more beneficial than pallidotomy. PMID- 10853074 TI - Stereotactic ventral intermedial thalamotomy for the treatment of essential tremor: results of a series of 37 patients. AB - We have analyzed 43 ventral intermediate thalamotomies performed in our center for treatment of medically intractable essential tremor (ET) in 37 patients. The mean age of patients was 70.9 years (range 42-84), duration of symptoms 33.3 years (1-65). The surgery in all cases was performed with stereotactic technique using MRI or CT localization. Intraoperative neurophysiological confirmation of the target location was obtained using a macrostimulation technique. All patients experienced either complete abolition of the contralateral tremor or significant improvement in tremor intensity immediately after the surgery. At follow-up examination 1-13 months after the operation, 60.5% of patients had no tremor, and 13.9% had mild residual tremor without interference with daily life. Tremor recurrence was observed in 5 patients, all of whom underwent repeat ventral intermedial (VIM) thalamotomy with excellent results. Transient problems with speech and motor functions were observed after 15 thalamotomies, permanent hemiparesis and speech difficulties were seen in 6 patients. We conclude that VIM thalamotomy is a highly effective procedure for medically intractable ET and may be performed with no mortality and low morbidity rate. PMID- 10853075 TI - A comparison of surgical approaches for the management of tremor: radiofrequency thalamotomy, gamma knife thalamotomy and thalamic stimulation. AB - OBJECTIVE: Between April 1994 and January 1999, 39 stereotactic procedures for patients with intractable tremor were performed at the University of Pittsburgh Medical Center. A retrospective analysis of results of radiosurgical thalamotomy (n = 15), MR-guided stereotactic radiofrequency thalamotomy (n = 13), and deep brain thalamic stimulation (DBS; n = 11) was performed to study relative advantages and risks of these procedures. METHODS: All options were discussed with the patients, but radiosurgery usually was performed in elderly patients with concurrent medical problems. Stereotactic thalamotomy and DBS was performed with MR guidance and macrostimulation. For radiosurgery, a median dose of 140 Gy (range 130-150 Gy) was delivered using a single 4-mm collimator. RESULTS: Of the 13 patients who underwent radiofrequency thalamotomy, 5 had immediate complete arrest of tremor, 6 had a significant reduction and 2 had partial reduction. All 11 patients who underwent DBS had excellent control of tremor immediately after the procedure, and in longer-term follow-up 10/11 maintained excellent tremor control. Of the 12 evaluable radiosurgery patients, 10 noted excellent relief and 2 had partial relief. CONCLUSION: Stereotactic procedures for tremor control are safe and effective. Each procedure has specific advantages and disadvantages that are important for patient selection. PMID- 10853076 TI - Electrophysiological recordings in pallidotomy localized to 3D stereoscopic imaging. AB - A quantitative on-line analysis of electrical activity in the pallidum of Parkinsonian patients has been developed to determine the focal point of lesioning. A 3D volume image system has been developed to display basal ganglia anatomy and coregister the electrophysiological data within the globus pallidus. Thirty patients undergoing 41 pallidotomies are presented. Neuronal activity from the pallidum is recorded using a semi-microelectrode. Based on this activity, lesioning is performed. Postlesion recordings are made to determine the necessity of additional lesioning. A stereoscopic 3D volume MR image system has been developed that along with on-line signal processing allows visualization of high neural activity in the pallidum and postlesion residual activity. PMID- 10853077 TI - Comparison of three techniques for calculation of target coordinates in functional stereotactic procedures. AB - We compared three techniques of target coordinate determination for various functional stereotactic procedures. All procedures were based on preoperative MRI with contiguous 3-mm cuts. The first technique involved determination of anatomical landmarks and fiducial markers of the stereotactic frame on the monitor screen of an MRI scanner and calculation of the target point using a series of formulas; the second technique used a Leksell tabletop localizer, and the third technique is a part of 'Stealth' stereotactic navigation software. Final coordinates for the procedure were derived from all three techniques and subsequently adjusted using intraoperative electrical macrostimulation. We found that difference between techniques was on average 0.9 +/- 0.4 mm in each of three directions, and 1.8 +/- 0.9 mm in absolute distance. There were 7 cases in which one of the techniques had a discrepancy of more than 3 mm (more than 1 MRI slice thickness) compared with the other two, indicating a potential error in coordinate determination. This difference could potentially result in inappropriate placement of the electrode, thus affecting the procedure outcome. In 6 cases, such an error apparently occurred with the first or second technique of calculation. The average number of mapping trajectories decreased from 1.8 to 1.4 since this stereotactic software became a part of operative planning. We conclude that use of computerized planning software increases the precision of target coordinate calculation and improves the accuracy of functional stereotactic procedures. PMID- 10853078 TI - Thalamic stimulation in patients with multiple sclerosis. AB - OBJECTIVE: To assess tremor control and side effects in patients with multiple sclerosis (MS) treated with chronic thalamic stimulation for relief of upper extremity tremor. METHODS: Five patients were studied before and after thalamic placement of a deep brain stimulation (DBS) system. Preoperative and postoperative evaluation included magnetic resonance imaging, Extended Disability Status Scale (EDSS), the Bain-Finchley visual analog scale for tremor, video recording and neuropsychological testing. Stereotactic targeting of the Vim nucleus was done using computed tomography; intraoperative testing was done under local anesthesia before permanent implantation. RESULTS: Functionally useful tremor suppression was obtained in 3/5 patients. Neuropsychological deficits of higher cortical function, memory and visuospatial coordination were observed in all patients before surgery; in 1 patient with improved postoperative visuospatial coordination, worsened memory was found. New brainstem plaque formation was seen several weeks after surgery in 1 patient who had an acute worsening of MS which improved after high-dose intravenous steroids. CONCLUSIONS: Chronic thalamic stimulation may help selected patients with MS-induced tremor. Given the complexity of their underlying illness, patients must be selected carefully, and long-term follow-up is vital to evaluate the true utility of DBS. PMID- 10853079 TI - Electrical inhibition of basal ganglia nuclei in Parkinson's disease: long-term results. AB - Since 1995 a group of 57 patients with familial tremor and Parkinson's disease have been operated in two main sets: the first, with 11 cases, with predominant tremor, where the ventro-oralis posterior (Vop) thalamic nucleus was stimulated, and the second, with 44 cases, with mainly rigidity and akinesia, where the globus pallidus internus (GPi) was unilaterally (10 cases) or bilaterally (34 cases) stimulated. In 2 cases, a combined thalamic-pallidal stimulation was achieved. In the last cases, an image fusion software with 3D correlation of MR and CT images and volumetric image matching has been used to correct MRI distortion. In all cases, an Itrell II stimulation system (Medtronic) was implanted. Result analysis shows that Vop stimulation is mostly effective for tremor, and GPi stimulation is better for rigidity and akinesia. Transcortical magnetic stimulation tests suggest that GPi pallidal electrical inhibition increases cortical excitability, as opposed to Vop thalamic stimulation, which implies a different mechanism of action. PMID- 10853080 TI - Tolerance and tremor rebound following long-term chronic thalamic stimulation for Parkinsonian and essential tremor. AB - Fifty-eight patients, 36 with essential tremor (ET) and 22 with Parkinson's disease (PD), received deep brain stimulation (DBS) in the thalamic ventral intermediate (Vim) nucleus. The mean follow-up was 17 months for ET and 21 months for PD patients. Stimulation parameters were adjusted as needed, at various intervals after surgery. Results were assessed using routine clinical evaluation and established outcome scales. All patients needed incremental increase in stimulation parameters at various intervals during the first 6-12 months after surgery. The mean voltage 1 week postoperatively was 1. 45 V in PD patients, and 1.37 V in ET patients. Twelve months later, the figures were 2.14 V in PD and 2.25 V in ET patients. At 1 year, the Essential Tremor Rating Scale (ETRS) improved from 54 to 28 (p < 0.0001). The motor part of the Unified Parkinson's Disease Rating Scale (UPDRS) improved from 37 to 26 (p < 0.01). Tremor items of the UPDRS improved more markedly (p < 0.0001). One week postoperatively 90% of PD, and 89% of ET patients were tremor free. One year later, 70% of PD and 60% of ET patients remained mostly tremor free. Upon switching off stimulation, there was a clear tendency for tremor rebound (p = 0.07) in the PD group, requiring continuous 24-hour stimulation in some patients. Permanent non-adjustable ataxia was induced by stimulation in 2 PD patients. PMID- 10853081 TI - Outcomes and complications of fetal tissue transplantation in Parkinson's disease. AB - This study was undertaken to investigate the outcomes, complication rates and risk factors of stereotactic intrastriatal neurotransplantation for Parkinson's disease (PD). Bilateral stereotactic neurotransplantation was performed (as previously described) in 60 patients with idiopathic PD. Clinical outcome was evaluated using the Unified Parkinson's Disease Rating Scale (UPDRS). The incidence of complication was evaluated by retrospective analysis of the clinical outcomes of the transplanted patients. Patients demonstrated significant improvement in UPDRS scores 12 months after transplantation. Nine patients experienced adverse effects after neurotransplantation, 3 requiring surgical intervention. Patients showed a significant overall improvement and no greater incidence of risk than that of other intracranial procedures. PMID- 10853082 TI - Correlation between MRI-based stereotactic thalamic deep brain stimulation electrode placement, macroelectrode stimulation and clinical response to tremor control. AB - In this study we compared the position of the electronically active contact of the thalamic (Vim) deep brain stimulation (DBS) electrode to the stereotactic location of its tip. Fifteen patients with either Parkinson's disease (PD) or essential tremor (ET) underwent stereotactic, MRI-based placement of the Medtronic quadripolar DBS electrode. An overall improvement of 69% was achieved in the tremor scores during a period of 1-13 months after implantation of the DBS electrode. Eleven patients with ET showed 70% clinical improvement of tremor, compared to a 58% response observed in the 4 patients with PD. The electrode tip center was 11.2 +/- 1.54 mm lateral to the third ventricular wall, 5.38 +/- 1.02 mm anterior to the posterior commissure and 2.9 +/- 3.57 mm inferior to the level of AC-PC line. The most significant deviation from the planned stereotactic target was observed in the Z-coordinate. In our group of patients, stimulation settings favored the contacts closer to the AC-PC line, correcting the electrode tip position to 0.80 +/- 2.84 mm (p < 0.001) inferior to the level of the AC-PC line. In our experience, thalamic DBS offers a reversible and adjustable 'lesion' to compensate for the anatomic variabilities encountered in the positioning of the DBS electrode tip. PMID- 10853083 TI - Stereotactic posterioventral pallidotomy improves balance control as assessed by computerized posturography. AB - Postural instability is arguably the most debilitating symptom of Parkinson's disease (PD). Recently, posterioventral pallidotomy/pallidoansotomy (PVP) has been advocated to improve a multitude of symptoms associated with PD. Dyskinesias, rigidity and bradykinesia are the most talked about improved symptoms, but posture and gait are also affected after PVP. To analyze the effect of PVP on postural control, 14 patients with PD were prospectively studied using a computerized dynamic posturography machine. Seven males and 7 females underwent a total of 18 procedures, 6 left PVP, 6 right PVP, 2 bilateral and 2 had Vim thalamotomies in addition to PVP. Data were collected pre- and postoperatively after a 12-hour drug-free interval ('off' period) and 1-2 h after medications ('on' period). Postoperative analyses were performed between 1 and 3 months postoperatively. As a group, patients' balance, in the off period, improved after surgery in a dynamic setting. Prior to surgery, patients' anterior-posterior sway exceeded their stability limits (patient fell) on 31% of the trials. After surgery, the fall rate decreased to 23%. Anterior-posterior sway decreased significantly (p < 0.05) postoperatively when the platform was sway referenced. In comparing the effect of surgery in decreasing sway with that of medication preoperatively, improvement after surgery (off period) was better than the preoperative on period (p < 0.05). Patients also improved in ostoperative off state when compared to preoperative off state with the platform sway referenced (p < 0.05), controlling for improvement in dyskinesia-induced imbalance. In conclusion, PVP improves standing balance performance better than that achieved with medications preoperatively. Since central input parameters were improved, the mechanism of PVP may be centralized. PMID- 10853084 TI - Vocal tremor reduction with deep brain stimulation. AB - We present a case illustration of the significant effect that deep brain stimulation (DBS) of the thalamus can have on vocal tremor. A 72-year-old female with a history of essential tremor was noted preoperatively to have a moderate vocal tremor (3 on a scale of 1-5). Following bilateral DBS of the thalamus, the vocal tremor rating improved to 1. Acoustic analysis demonstrated her vocal tremor to be affecting the amplitude of her voice at 5.58 Hz preoperatively, at 1. 93 Hz postoperatively with both leads on and at 1.54 Hz with only the left lead on. A videotaped endoscopic view of the patient's vocal cords (presented at the 1999 ASSFN meeting) clearly illustrated the dramatic changes apparent in the vocal tremor when the stimulators were turned on and off. PMID- 10853085 TI - Non-microelectrode guided stereotactic pallidotomy for Parkinson's disease: surgical technique and results PMID- 10853086 TI - Human spinal microelectrode recordings PMID- 10853087 TI - Placement of deep brain stimulators into the subthalamic nucleus: technical approach PMID- 10853088 TI - Unilateral pallidal stimulation in cervical dystonia. AB - Cervical dystonia (spasmodic torticollis) is a focal dystonia of the cervical region. Various treatment modalities have been performed with variable success rates. We present a 42-year-old woman complaining of involuntary head rotation for the last 3 years. Different medical treatments had been used for 3 years. Botulinum toxin injections resulted in temporary and moderate improvement for periods of 3-4 months. Pallidal stimulation was performed using a quadripolar electrode and a battery-operated programmable pulse generator. We conclude that a unilateral pallidal lesion or stimulation is an effective method of treatment in focal dystonia. The target must be the pallidum contralateral to the contracted sternocloidomastoid muscle. Deep brain stimulation is superior to lesioning because of the capability of manipulating the stimulation parameters which can modify the pallidotomy effect. PMID- 10853089 TI - Multimodality stereotactic brain tissue identification: the NASA smart probe project. AB - Real-time tissue identification can benefit procedures such as stereotactic brain biopsy, functional neurosurgery and brain tumor excision. Optical scattering spectroscopy has been shown to be effective at discriminating cancer from noncancerous conditions in the colon, bladder and breast. The NASA Smart Probe extends the concept of 'optical biopsy' by using neural network techniques to combine the output from 3 microsensors contained within a cannula 2. 7 mm in diameter (i.e. the diameter of a stereotactic brain biopsy needle). Experimental data from 5 rats show the clear differentiation between tissues such as brain, nerve, fat, artery and muscle that can be achieved with optical scattering spectroscopy alone. These data and previous findings with other modalities such as (1) analysis of the image from a fiberoptic neuroendoscope and (2) the output from a microstrain gauge suggest the Smart Probe multiple microsensor technique shows promise for real-time tissue identification in neurosurgical procedures. PMID- 10853090 TI - Distinguishing recurrent tumor and radiation necrosis with positron emission tomography versus stereotactic biopsy. AB - With the recent approval of reimbursement for positron emission tomography (PET), it has become important to clarify the utility of this diagnostic study. We evaluated the utility of PET to distinguish radiation necrosis from recurrent tumor in a retrospective review of patients with primary glial neoplasms. Fifteen patients had preoperative contrast-enhanced MRI and PET images followed by stereotactic biopsy or craniotomy and histological confirmation. The sensitivity of PET was 43% (6/14) and the specificity was 100% (1/1). We examined the sensitivity of PET as a function of volumetric contrast enhancement on MRI. Eighty percent of true-positive PET studies occurred with volume enhancement greater than 10 cm(3). Seventy-five percent of false negatives occurred with volume enhancement less than 6 cm(3). Given the clinical significance of distinguishing tumor progression from radiation necrosis, we believe that PET is insufficient to resolve radiation necrosis versus tumor progression. PMID- 10853091 TI - MKM-guided resection of diffuse brainstem neoplasms. AB - OBJECTIVES: Some primary brainstem tumors, when extensive, are considered inoperable. We wanted to assess the value of robotic image-guided microscopic surgery in the resection of these tumors and to improve survival and quality of life for these patients. METHODS: Two patients with extensive brainstem tumors were evaluated at our center. They previously underwent several biopsies, attempted partial resections, radiotherapy and shunting. They presented with progressive neurological deterioration, 'coma vigil' for several months, and required life-supporting measures prior to surgery. Both patients underwent frameless stereotactic craniotomy using a MKM robotic microscope, intraoperative neurophysiological monitoring, and extensive resection of their recurrent brainstem tumors. RESULTS: In the immediate weeks after surgery, both patients became interactive and regained major motor and cranial nerve deficits present prior to surgery. Nine months after surgery, 1 patient succumbed to pneumonia. At 2 years after the operation, 1 patient has maintained his neurological status and showed no recurrence on imaging studies. CONCLUSIONS: Image-guided surgery with an MKM microscope allows surgical outlines to be injected in the microscope viewer and facilitates resection of extensive brainstem tumors previously considered inoperable. PMID- 10853092 TI - Analysis of brain tumours suitable for curability via gene therapy in North East Malaysia. AB - Two hundred primary brain tumours in both adults and children from the year 1990 to 1998 presenting for treatment to the Neurosurgical Division of the Hospital of the University of Sciences Malaysia were studied retrospectively. Volumes of tumours were taken from CT scans with contrast using two formulas and divided into 4 groups: (1) less than 20 cm(3), (2) 20-50 cm(3), (3) 50-100 cm(3) (4) larger than 100 cm(3). The majority of the brain tumours were in the volume range of 50-100 cm(3), and are thus potentially curable with retroviral gene therapy. PMID- 10853093 TI - Adjuvant stereotactic radiosurgery for anaplastic ependymoma. AB - OBJECT: The purpose of this retrospective study is to evaluate the role of stereotactic radiosurgery using the Gamma Knife as an adjuvant to other modalities used in the treatment of malignant ependymomas of both children and adults and to assess its efficacy in terms of tumor control and overall survival. METHOD: Between 1987 and 1998, 22 patients in the age range of 1.5-65 years (mean age 22. 3) with progressive anaplastic ependymoma were treated by stereotactic radiosurgery using the 201 source Co-60 Leksell Gamma Knife at the University of Pittsburgh. The irradiated tumor volume varied from 0.84 to 36.8 cm(3) (mean 13.7). The median dose delivered to the tumor margin was 16.1 Gy (range 10-20), and the mean maximal dose was 32.2 Gy (range 20-40). The disease-free survival, the tumor control rate and the overall survival were recorded to evaluate the efficacy of radiosurgery. The median follow-up from radiosurgery was 21 months (range 4-84). RESULTS: Median survival after radiosurgery was 2.2 years (46.6 +/- 12.1% 5-year actuarial). Median survival from the initial diagnosis was 10. 1 years (50.3 +/- 12.5% at 5 years, 37.7 +/- 14.4% at 10 years). Reduction or stabilization of the treated tumor was seen in 16 out of 22 (68%) patients. Forty one percent of the patients eventually developed delayed distant cerebral recurrence outside the treated volume. The 5-year actuarial rates for local control and cranial control at any location were 62.3 +/- 13.6% and 32.4 +/- 10.8%, respectively. No complication occurred as a side effect of radiosurgery. CONCLUSION: For patients with locally recurrent or progressive anaplastic ependymomas, Gamma Knife stereotactic radiosurgery proved to be safe and effective as a salvage adjuvant therapy to achieve local tumor control and improve survival. PMID- 10853094 TI - Functional magnetic resonance image-guided surgery of tumors in or near the primary visual cortex. AB - OBJECTIVE: To assess the accuracy of functional magnetic resonance imaging (fMRI) of the primary visual cortex in patients undergoing surgery for tumors in the occipital lobe. METHODS: Two patients with nondominant occipital lobe tumors were studied, one with a solitary lung metastasis and another with radiation necrosis after radiosurgery for a low-grade astrocytoma. At surgery, visual evoked potentials (VEPs) were stimulated using Light-emitting-diode goggles and recorded using cortical grids placed immediately after brain exposure. The location of the peak VEP was compared to that predicted by the registered functional scan. RESULTS: In each case, the epicenter of visual activation as represented on the registered fMRI corresponded to the site of peak VEP recording. Prediction error for the visual cortex, measured in patient 1, was 1.0 mm. Visual confirmation showed the registration in the second patient to be accurate as well. CONCLUSION: As previously demonstrated for sensorimotor fMRI, visual fMRI accurately predicts the location of the primary visual cortex. Additional confirmation is expected with more clinical experience. PMID- 10853095 TI - Stereotactic interstitial radiosurgery for malignant brain tumors PMID- 10853096 TI - Functional magnetic resonance imaging and radiosurgical dose planning. AB - OBJECTIVE: To assess the effect of functional magnetic resonance imaging (fMRI) on stereotactic radiosurgical (SRS) dose planning. METHODS: Patients included those undergoing SRS whose lesions were in or near areas that could be identified with fMRI. After processing, an fMR scan was registered to the anatomic scan, and this dataset was registered to a stereotactic CT scan. The imaged functional areas were contoured along with standard anatomical targets. Dose planning was done at first with the functional volumes rendered invisible; the plans were then adjusted as needed using the functional targets. Doses were measured using a dose volume histogram tool. RESULTS: SRS was performed in 12 patients, 1 of whom also underwent SRT. Functional volumes studied included motor cortex in 8 patients, visual in 6 and language in 3; a total of 33 functional targets were imaged. Prescription doses ranged from 12 to 22.5 Gy (mean 19.5 Gy), and the maximum dose to functional volumes from 8 cGy to 18.5 Gy (mean 2.9 Gy). In 6 patients, arc adjustment using functional targets yielded a >50% reduction in dose to at least one functional volume; in all patients, the dose reduction to 50 and 75% of functional volumes averaged 4% (12 cGy) and 13% (30 cGy), respectively, while the reduction of maximal dose averaged 24% (50 cGy). CONCLUSIONS: fMRI can be used in SRS to reduce irradiation of eloquent brain using standard prescription doses. Appropriate arc adjustment may allow for escalation of the dose to the targeted lesion. PMID- 10853097 TI - Fractionated stereotactic radiosurgery for treatment of acoustic neuromas. AB - BACKGROUND: Microsurgery and single-fraction radiosurgery for acoustic neuromas are associated with high rates of control, but can result in facial palsy and trigeminal neuropathy. To reduce the morbidity of treatment for acoustic neuromas while maintaining efficacy, we explored fractionated stereotactic radiosurgery (FSR). METHODS: We reviewed data for 31 acoustic neuromas in 30 patients treated with 25 Gy (linear accelerator) given in 5 consecutive daily fractions. The minimum follow-up was 6 months (6-44 months). The mean tumor volume was 1.1 cm(3) (0.1-8.74 cm(3)). RESULTS: All tumors remain controlled (9 smaller, 22 unchanged). No patient has experienced post-radiosurgery facial motor dysfunction. Two patients developed new trigeminal neuropathy; 2 patients with preexisting trigeminal nerve symptoms had improvement after FSR. Balance improved in 3 patients, was unchanged in 20 and worsened in 7 patients. Of the 12 patients with useful hearing (PTA < or = 50 dB) prior to treatment, 9 patients retained useful hearing following FSR. Subjectively, of 25 patients with any hearing prior to treatment, 2 had improvement, 10 remained unchanged and 13 had worsening. CONCLUSIONS: Short course FSR for acoustic neuromas results in acceptable toxicity and may provide high control of tumors. Longer follow-up is needed to assess outcomes. PMID- 10853098 TI - Stereotactic radiosurgery of cerebral arteriovenous malformations: appearance of perinidal T(2) hyperintensity signal as a predictor of favorable treatment response. AB - The purpose of this study was to analyze the significance of perinidal T(2) hyperintensity appearance after radiosurgery of arteriovenous malformations (AVMs), as a predictor of treatment response. Our initial experience with linear accelerator (LINAC) radiosurgery at University of California, Los Angeles, between 1990 and 1997 involved treatment of 129 patients affected by cerebral AVMs. Based upon availability of neuroimaging follow-up, 48 patients with 50 AVMs were selected for review. Forty (80%) of the AVMs underwent complete obliteration or significant reduction on follow-up MRI, on average 20 months after radiosurgery. Thirteen (72%) of 18 obliterated AVMs were associated with perinidal T(2) hyperintensity signal, on average 18 months (6-49) after radiosurgery. Ten (20%) of 50 AVMs (average volume 23.1 cm(3), ranging 7.5-46.5) were unchanged. Furthermore, only 3 AVMs in this group showed reversible T(2) signal changes. In patients with complete nidal obliteration, appearance of T(2) hyperintensity signal achieves 72% sensitivity in predicting successful treatment response. PMID- 10853099 TI - Stereotactic radiosurgery in the treatment of metastatic disease to the brain. AB - We review 190 consecutive patients with 434 metastatic tumors treated by gamma knife stereotactic radiosurgery, from August 1994 to February 1999. Median actuarial survival for all patients was 34 weeks. Factors correlated with significantly improved survival included controlled systemic disease and nonmelanoma histology. We found that no significant survival benefit could be discerned from adjuvant whole brain radiotherapy in this patient group. Survival was not statistically different for patients initially presenting with 1-4 metastases at initial treatment. PMID- 10853100 TI - Treatment of nasopharyngeal carcinoma: stereotactic radiosurgical boost following fractionated radiotherapy. AB - Treatment of patients with nasopharyngeal carcinoma (NPC) using external beam radiation therapy (XRT) alone results in significant local recurrence. To improve local control, stereotactic radiosurgery (SRS) was used to boost radiation to the primary tumor site following XRT in 23 patients with NPC. SRS was delivered utilizing a frame-based linear accelerator as a boost (range 7-15 Gy, median 12 Gy) following XRT (range 64.8- 70 Gy, median 66 Gy). In all 23 patients (100%) receiving SRS following XRT local control was achieved at a mean follow-up of 21 months (range 2-64 months). There have been no complications of treatment caused by SRS. However, 8 patients (35%) have subsequently developed regional or distant metastases. SRS boost following XRT provides excellent local control in NPC and should be considered for patients with skull base involvement. PMID- 10853101 TI - Gamma knife radiosurgery for arteriovenous malformations: lessons learned from japan PMID- 10853102 TI - Frame based stereotactic spinal radiosurgery: experience from the first 19 patients treated PMID- 10853104 TI - Gamma knife stereotactic radiosurgical treatment for refractory trigeminal neuralgia PMID- 10853103 TI - Respiration induced target drift in spinal stereotactic radiosurgery: evaluation of skeletal fixation in a porcine model PMID- 10853105 TI - Dose-volume effect in gamma knife radiosurgery of meningiomas. AB - OBJECTIVE: To study the relationship between dose-volume effect and complication in gamma knife radiosurgery of intracranial meningiomas. METHOD: By using an integrated logistic formula to establish mathematical models, the authors analyzed the relationship between treatment volume and logistic probability for 15 Gy, and the relationship between prescribed dose and treatment volume in 3% logistic probability. Furthermore, 37 meningiomas treated by gamma knife radiosurgery were analyzed retrospectively to investigate how the 3% isoeffect curve impacted on complication. RESULT: There was a linear relation between treatment volume and logistic probability for 15 Gy. The regression formula was f(x) = 0.0042x + 0.0007. The 3% isoeffect curve indicated that the prescribed dose was inversely proportional to the treatment volume. During 16.3 months follow-up on average, 18 cases below the 3% isodose curve were without complication, but in 19 cases above the 3% isodose curve, 5 cases suffered complications, the difference being statistically significant. CONCLUSION: Dose selection in gamma knife radiosurgery is volume dependent. The 3% isodose curve may possibly be the guideline in gamma knife radiosurgery of meningiomas. PMID- 10853106 TI - Long-term seizure outcome following corpus callosotomy in children. AB - INTRODUCTION: The long-term outcome of pediatric patients undergoing corpus callosotomy (CC) for palliative control of medically intractable seizures is presented. METHODS: During a 27-year period, 43 patients, 20 years of age or younger, underwent CC for seizure palliation and had a minimum of 1 year follow up. Seizure reduction and stability of that outcome for individual seizure types and for most disabling seizure were reviewed. RESULTS: Overall, 63% of the seizures documented showed a good response. For the most disabling seizure, 56% of the patients had good outcomes. Changes in outcome status occurred within the first 6 months, and outcome was largely maintained after that point. CONCLUSION: Callosotomy achieves the goal of seizure palliation in more than half of the patients, with stable, good outcomes being maintained in the majority of patients. PMID- 10853107 TI - MRI assessment of the anatomy of optic radiations after temporal lobe epilepsy surgery. AB - OBJECTIVE: The aim of this study was to determine the course of the temporal optic radiations. MATERIAL AND METHODS: Eighteen patients were included in this prospective study. All of them underwent a temporal lobectomy for epilepsy, including the mesial temporal structures and a variable extent of lateral neocortex (from 2 to 7 cm behind the temporal tip). An MRI was performed 2 months postoperatively, allowing assessment of the extent of lateral resection. Postoperative visual fields were determined by automatic static perimetry (ASP). RESULTS: (1) No patient complained of a disabling visual field deficit. (2) ASP, a highly sensitive technique, however, detected postoperative visual field deficits in 83% of patients, confined to the superior homonymous field contralateral to the resection. (3) A strong correlation was found between the presence of a visual field deficit and the extent of laterotemporal resection. (4) The smallest anteroposterior resection resulting in a field defect was limited to 20 mm from the temporal tip. CONCLUSION: (1) This study confirms a strong correlation between postoperative visual field deficits and the extent of lateral neocortical temporal resection. (2) The anterior limit of Meyer's loop is likely to be located more rostrally than previously believed. (3) Despite this, lateral resection remains useful in some cases for seizure control. PMID- 10853108 TI - Surgical management of intractable epilepsy in children with hemophilia. AB - Intracranial hemorrhage occurs in 2-8% of patients with hemophilia and can result in neurologic sequelae including seizure disorders. There is a paucity of data concerning the surgical treatment of epilepsy in children with hemophilia. We review our experience with 2 children who developed medically refractory seizure disorders. Both children underwent hemispherotomy, at 18 months (case 1) and 13 years of age (case 2), respectively. Perioperative management included continuous factor replacement. Both children tolerated surgical intervention without complications or increased neurologic deficit. Case 2 showed a 90% reduction in seizure frequency, and case 1 is seizure free. Surgical management of intractable epilepsy in children with hemophilia is safe and effective. PMID- 10853109 TI - Usefulness of the dipole tracing method with a scalp-skull-brain head model: relationship between the epileptic focus and equivalent current dipole locations. AB - PURPOSE: We examined whether the dipole tracing (DT) method with a realistic three-shell head model and inhomogeneous electric conductivity is useful to estimate the epileptic focus from interictal spikes. METHOD: This study included 17 temporal lobe epilepsy (TLE) cases, classified into three types as type A (unilateral), type B (intermediate) and type C (bilateral) and 5 extratemporal epilepsy (XTLE) cases. The epileptic areas were determined by noninvasive and/or invasive examinations. Selected interictal spikes were analyzed and the calculated equivalent current dipoles (ECDs) of dipolarity greater than 0.98 were superimposed over the realistic head model in each patient. We evaluated the ECD concentration within and around the epileptic area. RESULTS: In TLE cases, types A showed better ECD concentration (87%) within the epileptic area than other types (type B: 68%; type C: 74%). XTLE exhibited variable ECD distribution within and around the epileptic area. CONCLUSION: The DT method with a realistic head model and inhomogeneous electric conductivity can be useful to estimate the epileptic area from interictal spikes, especially in unilateral TLE cases. PMID- 10853110 TI - A 13-year experience with epilepsy surgery. AB - Between 1985 and 1997, 563 therapeutic craniotomies were performed: 311 anterior temporal (ATL) and 158 extramesial temporal (XMT) resections, 67 callosotomies, 20 hemispherectomies and 7 multiple subpial transections. Sixty-seven percent of nonlesional ATL cases were seizure free (SF), and 76% of nonlesional ATL cases < or =18 years old were SF. Seventy-eight percent of lesional ATL cases with complete resection were SF. Seventy-three percent of lesional cases < or =18 were SF. Thirty-seven percent of nonlesional XMT cases were SF. Seventy percent of XMT lesional cases with complete resection were SF, and 82% of lesional XMT cases < or =18 were SF. Of the anterior callosotomy cases, there was a > or =90% decrease in generalized tonic-clonic seizures in 50% of patients, and in tonic seizures, drop attacks, absence and myoclonic seizures in approximately 60-70% of patients. Of 20 hemispherectomies, 65% were SF. Of 7 multiple subpial transections, 29% were SF. PMID- 10853111 TI - Long-term multicenter experience with vagus nerve stimulation for intractable partial seizures: results of the XE5 trial. AB - OBJECTIVE: Intermittent stimulation of the left cervical vagus nerve trunk (VNS) with the NeuroCybernetic Prosthesis (NCP) is emerging as a novel adjunct in the management of medically refractory epilepsy. We review the safety and efficacy of VNS 1 year after completion of the E05 study, the largest controlled clinical trial of VNS to date. METHODS: One hundred and ninety-nine patients with intractable epilepsy and at least 6 complex partial or secondarily generalized seizures per month enrolled in a randomized, double-blinded, partial crossover trial of high versus low parameters of stimulation (E05). After 3 months, all patients received high stimulation during an open-label, nonblinded extension trial (XE5). Seizure frequency, adverse events and multiple physiologic variables were monitored at regular intervals. RESULTS: At 3 months, the mean reduction in seizure frequency among patients receiving high stimulation during E05 was 28%. Of the 199 subjects participating in this acute-phase trial, 195 continued in the long-term protocol. Among the latter patients, 21 subsequently exited the study due to lack of efficacy, and 2 others died from causes unrelated to VNS. Complete data were obtained for 164 of the remaining subjects. Using a declining N analysis, the mean and median reduction in seizure frequency at 15 months was 37 and 45%, respectively. A last visit carried forward analysis, which controls for dropouts and incomplete follow-up, yielded comparable results (34 and 45%, respectively), indicating little potential for selection bias. At 15 months, 39% of the subjects had a greater than 50% reduction in seizures, including 21% who had a greater than 75% reduction, and 2% have remained seizure free. Few serious adverse events, physiological perturbation or device failures were reported. CONCLUSIONS: The long-term multicenter safety, efficacy, feasibility and tolerability of VNS, as well as the durability of the NCP device have been confirmed. Unlike chronic therapy with antiepileptic medication, the efficacy of VNS is maintained during prolonged stimulation, and overall seizure control continues to improve with time. PMID- 10853112 TI - The relationship of magnetic source imaging to ictal electrocorticography in a neuronavigational workspace. AB - Magnetic source imaging (MSI) registers magnetoencephalographic (MEG) activity to a three-dimensional MRI volume. State-of-the-art MSI allows concurrent whole head coverage, but is practically restricted to interictal recording. However, the purpose of the presurgical evaluation of epileptic patients, in which MSI is playing an increasing role, is the elucidation of the ictal epileptogenic focus. The manner in which interictal MSI activity relates to the ictal focus has not yet been adequately examined. To facilitate this analysis, we are developing techniques to precisely coregister MSI to the ictal onset zone as defined by extraoperative intracranial grid/strip monitoring. The neuronavigational workspace is a convenient area in which to precisely coregister these (and other) imaging and physiological data sets. PMID- 10853113 TI - The behavioral and electroencephalographic effects of stereotactic radiosurgery for the treatment of epilepsy evaluated in the rat kainic acid model PMID- 10853114 TI - Current source density analysis of synaptic generators of human interictal spike PMID- 10853115 TI - Neurophysiological navigation in the trigeminal nerve: use of masticatory responses and facial motor responses evoked by electrical stimulationof the trigeminal rootlets for RF-thermorhizotomy guidance. AB - RF-thermorhizotomy of the trigeminal nerve is an effective and safe treatment of trigeminal neuralgia, provided lesioning of the sensory fibers is performed precisely. To control the accurate placement of the electrode tip, electrical stimulation testing, prior to thermal lesioning, is of prime importance. The clinical observation of direct masticatory responses (DMR) and facial evoked motor responses (EMR) produced by stimulation of the trigeminal rootlets (at 5 Hz) helps to place the electrode tip in the optimal location. The best location is the one where the threshold for eliciting DMR is high and the threshold for evoking facial EMR in the area corresponding to the trigger zone is low. EMR in orbicularis oculi indicates location in V1, levator labii EMR signifies V2 and orbicular oris EMR corresponds to V3. PMID- 10853116 TI - Precentral cortex stimulation for the treatment of central neuropathic pain: results of a prospective study in a 20-patient series. AB - The authors report a series of 23 patients with central neuropathic pain who were treated with the recently developed technique of precentral cortex stimulation (PCS). Of the 20 patients with a follow-up of more than 1 year (mean of 23 months) 25% had an excellent, 35% a good and 15% a fair relief of pain. In 25% the method failed. On the basis of these findings and the literature data (127 reported cases), the authors advocate PCS in patients with severe and medically refractory poststroke pain. PMID- 10853118 TI - Gamma knife radiosurgery for trigeminal neuralgia: experience at the Barrow Neurological Institute. AB - Forty-three patients with trigeminal neuralgia (TN) unresponsive to pharmacologic treatment and/or prior invasive procedures underwent stereotactic radiosurgery with the Gamma Knife (GK). Outcome was evaluated by a standardized questionnaire mailed to each patient. The mean follow-up was 9 months. Fifteen patients (35%) reported no trigeminal pain and were no longer taking medication. Three patients (7%) experienced occasional pain, but were no longer taking medication. In 15 patients (35%), pain improved and was adequately controlled by medication, often in lower dosages than preoperatively. Pain was reduced in 9 patients (21%), but their symptoms were still inadequately controlled by drug therapy, and 1 patient (2%) reported no pain relief after treatment. Three patients (7%) described new facial numbness, but in none was this bothersome. GK radiosurgery for TN appears to have minimal morbidity, although the success rate may be slightly lower than that of other operative procedures. More patients and longer follow-up are needed before drawing final conclusions regarding efficacy and complications. PMID- 10853117 TI - Efficacy of transverse tripolar stimulation for relief of chronic low back pain: results of a single center. AB - The goal of this study was to evaluate the efficacy of the transverse tripolar spinal cord stimulation system (TTS) in providing relief of low back pain in patients with chronic non-malignant pain. Transverse tripolar electrodes were implanted in the lower thoracic region (T(8-9) to T(12)-L(1)) in 10 patients with chronic neuropathic pain, all of whom reported a significant component of low back pain in combination with unilateral or bilateral leg pain. One patient reported inadequate pain relief during the trial and was not implanted with a permanent generator. A visual analogue scale of low back pain showed a nonsignificant decrease from 64 +/- 19 to 47 +/- 30 (p = 0.25; paired t test) after 1 month of stimulation. Similarly, functional disability evaluated using Oswestry Low Back Pain Questionnaire was not improved (p = 0. 46; paired t test). We conclude that chronic low back pain is not particularly responsive to the transverse stimulation provided by the TTS system. PMID- 10853119 TI - Spinal cord stimulation electrode design: A prospective randomized, controlled trial comparing percutaneous and laminectomy electrodes PMID- 10853120 TI - Image guidance: the Foundation for the Future Design of Neurosurgical Procedural Facilities. AB - Today's image guidance systems are the foundation of minimally invasive diagnosis and therapy and are the basis for the rational design of the neurosurgical operating room of the future. The building blocks of this procedural arena will be (1) high-resolution MR/CT anatomic imaging supplemented with (2) functional imaging using MR and magnetoencephalography, (3) real-time image monitoring using ultrasound probes, open MR and portable CT adapted to image guidance systems and (4) conformal radiosurgery systems. Although the initial investment in such facilities may be high, eventual cost saving through added precision and safety will result in shorter inpatient stays, lower morbidity and more complete realization of treatment goals. PMID- 10853121 TI - Staged functional neurosurgery using image fusion: electronic atlas and microelectrode recording at Dundee. AB - The unforgiving nature of the thalamus, the globus pallidus and the subthalamic nucleus necessitates precise localization of functional targets. This requires the total attention of both the patient and the surgeon. To maximize the concentration of the patient and provide the most accurate localization, we performed staged stereotactic functional procedures. The first stage was performed under general anesthesia to abolish any head movement. We fused CT and MRI images and correlated the fused images with a digitized Talairach brain atlas. We calculated the target coordinates and fixed a modified Bennett Sphere to the skull with the central hole defining the trajectory to the target. The surrounding 12 holes gave parallel trajectories to targets surrounding the anatomical target at 2-mm intervals. The second stage was performed at least a week later under local anesthesia. Microelectrode recording using three simultaneous channels was used to refine the target. Once the microelectrode recordings and macrostimulation confirmed the desired target, a lesion was created or an Activa neurostimulator was inserted. Our early results using this technique in 28 procedures (in 19 patients) indicate a good outcome in 86% and a technical failure in 1 patient. PMID- 10853122 TI - Coregistered ultrasound as a neurosurgical guide. AB - INTRODUCTION: The dynamic nature and three dimensionality of ultrasound data can be utilized to enhance the capabilities of image guidance systems. METHODS: Coregistration of ultrasound data was done using an electromagnetic digitizer, and subsequent ultrasound images were correlated with preoperative MRI studies. Thirty-two patients undergoing craniotomy were investigated in this manner. RESULTS: Phantom testing done with a rigid stylus and 3D ultrasound tracker demonstrated an accuracy of 1.36 +/- 1.67 mm in determining the location of a point. Thirty-two clinical cases were coregistered without difficulty. CONCLUSION: Coregistered ultrasound is a useful methodology that can aid in neuronavigation. PMID- 10853123 TI - Comparison of stereotactic brain biopsy to interventional magnetic-resonance imaging-guided brain biopsy. AB - Lesions within the brain are commonly sampled using stereotactic techniques. The advent of interventional magnetic resonance (MR) imaging now allows neurosurgeons to interactively investigate specific regions with exquisite visualization. We compared the safety and efficacy of this new surgical approach with stereotaxis. From February 1991 to June 1998, 134 stereotactic and 35 interventional MR-guided brain biopsies were performed. Stereotactic biopsies utilized preoperative scanning. Interactive scanning was used to confirm accurate positioning of the biopsy needle within the region of interest. Intraoperative pathologic examination of biopsy material was performed to verify the presence of diagnostic tissue in both biopsy groups. Intra- and postoperative MR imaging was obtained to exclude the presence of intraoperative hemorrhage. Recently, MR spectroscopic targeting has been utilized in 6 patients. In the stereotactic group, 129/134 (96%) biopsies were diagnostic. One patient had a transient hemiparesis after a brain stem biopsy and another suffered a fatal hemorrhage for a morbidity rate of 0.7% and a mortality rate of 0.7%. In reviewing 7,471 stereotactic biopsies, the morbidity was 3.5%, mortality 0.7% and diagnostic yield 91%. All 35 MR-guided brain biopsies were diagnostic (100%). MR spectroscopy was accurate in all cases in distinguishing recurrent tumor (5 cases) from radiation necrosis (1 case). One patient (3%) suffered a transient hemiparesis following a pontine biopsy and another patient (3%) developed a postoperative scalp cellulitis. No patient sustained a clinically or radiologically significant hemorrhage as determined by the immediate postbiopsy, intraoperative MR imaging. Interventional MR-guided brain biopsy is a safe and effective technique for evaluating lesions of the brain with morbidity and mortality rates comparable to those of stereotaxis. MR guided biopsy appears to have a higher diagnostic yield than stereotaxis, which may reflect the ability to perform interactive, intraoperative scanning with that technique. PMID- 10853124 TI - Intraoperative image guidance using the brain Lab/Vector vision system. indications and costs in 166 consecutive cases PMID- 10853133 TI - Local tumor control and survival: clinical evidence and tumor biologic basis. AB - Clinical and tumor biologic evidence indicates that local tumor recurrence is associated with a higher rate of distant metastases and adverse effects on patient survival. The findings discussed in this article are based on results from breast, prostate, and lung carcinoma, all tumors with high metastatic potential for which local tumor control was thought to be relatively unimportant. The fact that local tumor recurrence is associated with tumor progression and significant increases in distant metastases has important consequences in the relative emphasis on and the sequencing of locoregional and systemic therapies in the future. PMID- 10853134 TI - Conformal radiotherapy: what is it and why does it matter? AB - Three-dimensional conformal radiotherapy is quickly becoming a standard radiation oncology practice at many academic and community-based departments. Three dimensional conformal radiotherapy allows clinicians to deliver safer and more accurate treatments to patients. It is also being used to increase tumor doses for situations in which traditional radiation techniques have been unsuccessful. Current and future clinical trials of dose-escalated three-dimensional conformal radiotherapy will examine its ultimate clinical effectiveness. PMID- 10853135 TI - Three-dimensional conformal radiotherapy for early stage prostatic cancer: techniques, outcomes, and possible pitfalls. AB - Advances in radiation therapy have led to the development of multiple methods to deliver radiotherapy accurately to a defined three-dimensional target. This technology is most appropriate in the management of early stage prostatic cancer. Precise delivery of radiotherapy can control prostate cancer while minimizing normal tissue (e.g., bladder and rectum) complications. PMID- 10853136 TI - Three-dimensional conformal radiation therapy: what are the costs and benefits? AB - Increased attention is being focused on the cost of various medical treatments as limited societal resources are recognized. Three-dimensional conformal radiation therapy is a sophisticated technique that allows high doses of radiation to be focused safely on a target. This technique is more expensive to implement and deliver compared with conventional radiation techniques. A consensus, however, is emerging after reviewing the data that shows three-dimensional conformal radiation therapy to be cost-effective when the clinical benefit is most apparent. More data, sophisticated analyses, and follow-up are necessary before more definitive conclusions can be made. PMID- 10853137 TI - Stereotactic radiosurgery: techniques and clinical applications. AB - Radiation is a common treatment modality for cancer. Although commonly used, the treatment techniques of radiation delivery have changed substantially. One of the most important changes in implementation is the widespread application of stereotactic techniques and their acceptance into the mainstream of radiotherapeutic delivery. The distinguishing characteristics of stereotactic radiosurgery and its current and future application are important for all physicians to understand. This article discusses these treatment techniques and applications from the perspective of a surgical oncologist. PMID- 10853138 TI - New frontiers in brachytherapy. AB - Brachytherapy gives a high dose to a tumor (better cell kill) because the sources are close to or within the tumor. Because of the location of the source (usually within target volume), the surrounding normal tissues are spared, resulting in lower morbidity. In the past, radiation exposure hazard to caregivers was a major disadvantage of brachytherapy; however, this concern has been reduced with the use of low energy isotopes and remote controlled afterloading techniques. The need for surgical exposure and its associated trauma has been overcome by using percutaneous, image-guided brachytherapy techniques. Use of brachytherapy to treat prostate and intravascular sites can be expected to increase. PMID- 10853139 TI - The contemporary role of the use of radiation therapy in the management of sarcoma. AB - In this article the current role of radiation therapy in the management of soft tissue sarcoma is evaluated. The importance of a multidisciplinary approach in the management of these patients is emphasized. The available literature supporting the possibility of limb salvage is reviewed. Finally, the newest developments in radiotherapy technology are also addressed. PMID- 10853140 TI - Is there a role for radiotherapy in the management of upper gastrointestinal malignancies? AB - Radiation therapy has several established roles in the management of upper gastrointestinal malignancy. These roles parallel those for which radiation is used in other anatomic regions (e.g., single and multimodality management with curative intent, organ preservation, and palliation). This article provides a brief summary of the use of radiotherapy in the management of upper gastrointestinal malignancies. It includes applications for radiotherapy used with curative and palliative intent. PMID- 10853141 TI - The contemporary role of radiation therapy in the management of lung cancer. AB - This article reviews the current role of radiotherapy in the management of non small-cell lung cancer. This modality is used extensively in lung cancer patients. In a variety of different contexts, the specific indications, results, controversies, and emerging areas are highlighted, with an emphasis on the author's personal techniques and a review of the pertinent literature. PMID- 10853142 TI - Advances in radiotherapy for carcinoma of the head and neck. AB - Increasingly, progress in the radiotherapy of carcinomas of the head and neck is being driven by principles of radiobiology. This article discusses some of the major advances in head and neck radiotherapy, including altered fractionation, concomitant chemotherapy, and intensity-modulated radiotherapy, in the context of radiobiologic rationale, potential impact on tumor control, and normal tissue complications. PMID- 10853143 TI - Vascular radiation therapy to reduce coronary artery restenosis. AB - Restenosis of the dilated segment following percutaneous transluminal angioplasty is a commonly occurring problem. Several techniques such as pharmacologic agents, directional atherectomy, and endovascular stenting have been attempted with little success. This article gives an outline on the scope of the problem of restenosis, its pathophysiology, attempted treatments, and the promise of coronary vascular radiation therapy. PMID- 10853144 TI - Contemporary issues in the use of radiation therapy for early invasive breast cancer. AB - Breast conservation therapy consisting of tumor excision followed by whole breast irradiation is an accepted alternative to mastectomy for many women with early invasive breast cancer. Ongoing research questions include defining the role of the tumor bed irradiation boost, the identification of patients who are at sufficiently low risk of breast cancer recurrence to be treated with excision only, exploring tumor bed brachytherapy as an alternative to whole breast irradiation, and biologic considerations in the future local management of breast cancer. PMID- 10853145 TI - Locally advanced breast cancer and postmastectomy radiotherapy. AB - Most patients with locally advanced breast cancer should be treated with a combination of chemotherapy, mastectomy (with immediate reconstruction if the patient so desires), and radiotherapy. Nonetheless, it seems reasonable to offer breast-conserving treatment to selected individuals who respond well to neoadjuvant therapy. Postmastectomy radiotherapy clearly reduces the risk of local-regional and distant failure for patients with earlier-stage invasive breast cancer with involved axillary lymph nodes. Certain subgroups of patients, however, may have such low local-regional failure rates that patients will not routinely find the benefits of radiotherapy sufficient to undergo such treatment. Further investigation of this issue is needed. PMID- 10853146 TI - The contemporary use of radiation therapy in the management of lymphoma. AB - In this article the classification and the treatment regimens for malignant lymphomas are discussed in a stage-oriented approach. The clinical trials that have redefined the role of staging laparotomy are reviewed. Intensity-modulated radiation therapy allows for highly conformal target definition. This novel delivery system is discussed through a case presentation. PMID- 10853147 TI - The surgical treatment of patients with skeletal injuries. PMID- 10853148 TI - Castless ambulatory method of treating fractures. 1942. PMID- 10853149 TI - The evolution of indirect reduction techniques for the treatment of fractures. AB - During the last decade, classic AO/ASIF techniques for internal fixation shifted from direct reduction and rigid fixation to biologic internal fixation using indirect reduction techniques. Biologic internal fixation is characterized by the preservation of bone and soft tissue vascularity and relative rather than absolute mechanical stability. Reduction is achieved by using soft tissue traction while obtaining axial and rotational alignment and the correct length. Stabilization is performed when possible by compression plating for load sharing or by bridge plating in comminuted fractures. Advancements of these techniques and the development of newer implants that minimize vascular damage have contributed to the development of biologic internal fixation. By using indirect reduction, by using longer plates to improve the mechanical leverage, and by applying fewer screws to avoid unnecessary damage to the bone, fracture union rates were high. There also was a decreased need for supplemental bone grafting. All of these factors provided stable fixation and allowed early motion. PMID- 10853150 TI - Percutaneous fixation of pelvic ring disruptions. AB - Percutaneous pelvic fixation is possible because intraoperative fluoroscopic imaging and other technologies have been refined. Anterior and posterior unstable pelvic ring disruptions are amenable to percutaneous fixation after closed manipulation or open reduction. Stable and safe fixation is achieved only after an accurate reduction. Anterior pelvic external fixation remains the most common form of percutaneous pelvic fixation; however, percutaneously inserted medullary pubic ramus, transiliac, and iliosacral screws stabilize pelvic disruptions directly while diminishing operative blood loss and operative time. These percutaneous techniques do not decompress the pelvic hematoma allowing early definitive fixation without the risk of additional hemorrhage. Complications associated with open posterior pelvic surgical procedures are similarly avoided by using percutaneous techniques. A thorough knowledge of pelvic osseous anatomy, injury patterns, deformities, and their fluoroscopic correlations are mandatory for percutaneous pelvic fixation to be effective. PMID- 10853151 TI - Percutaneous treatment of peritrochanteric fractures using the Gamma nail. AB - The goal of the current study was to analyze patients treated with the Gamma nail, and to describe techniques that prevent commonly reported complications. One hundred patients with closed peritrochanteric femur fractures were treated by one surgeon using the Gamma nail. Ninety patients met the minimum 6 month followup requirement. Eighty-eight of the 90 fractures (98%) healed after the index procedure. There were nine (10%) complications (nine patients) with four patients (4%) requiring surgical intervention. One patient required total hip arthroplasty because of nonunion, and one patient required cerclage wiring of a postoperative femoral fracture. The third patient had an infection develop which resolved after debridement and a course of antibiotics. The fourth patient complained of thigh pain, which resulted in hardware removal. In the five remaining patients with complications, one patient had multi-infarct dementia and the family refused additional treatment, two patients with intraoperative femur fractures did not require treatment and two patients with thigh pain did not require additional treatment. Surgical time averaged 53 minutes and blood loss averaged 104 cc. Percutaneous fixation using the Gamma nail is effective in treating patients with peritrochanteric fractures. Surgical time and blood loss were minimized, early weightbearing was initiated, and previously reported complications were decreased. PMID- 10853152 TI - Treatment of floating knee injuries through a single percutaneous approach. AB - The current study is a review of 20 patients treated by percutaneous stabilization for a floating knee. All patients were treated with a retrograde femoral intramedullary nail and a small diameter tibial intramedullary nail through a 4-cm medial parapatellar tendon incision. The average Injury Severity Score was 19. Two patients died in the early postoperative period and one patient was lost to followup. The average time to union for the 17 remaining patients with femoral shaft fractures was 14.7 weeks. One patient required dynamization. Four of the 17 patients with tibia fractures required an exchange nailing procedure, one with bone graft, to achieve union. One patient required dynamization and one patient with bone loss required only a bone graft. The average time to union for the tibia fractures was 23 weeks. One patient achieved 115 degrees knee flexion and the remaining 15 patients had full knee motion by 12 weeks, which they were able to maintain. No patient had signs or symptoms of knee pain. This demanding surgical technique using a small incision has yielded good clinical results. Although it is an excellent treatment option for patients with ipsilateral femoral and tibial shaft fractures, the mortality and tibial fracture complication rates remain high. PMID- 10853153 TI - Supracondylar femur fractures treated percutaneously. AB - One hundred twenty-five supracondylar fractures in 118 patients treated with the Green-Seligson-Henry supracondylar intramedullary nail were evaluated. One hundred four patients (111 fractures) were followed up to fracture union. The percutaneous technique was compared with open reduction and internal fixation using the same device. The mean operative time was greater for the open reduction technique when compared with the percutaneous technique (176 minutes versus 76.6 minutes, respectively), as was the mean estimated blood loss (229 cc versus 96.2 cc). The incidence of delayed union was approximately the same for patients who were treated with both techniques. However, the nonunion rate was significantly higher in the patients treated with open reduction and internal fixation than the patients treated with the percutaneous technique (5.6% versus 2.6%). Twenty-nine (39%) patients who were treated with open reduction and internal fixation required bone grafting versus only three (7%) patients who were treated with the percutaneous technique. Additionally, the percutaneous technique did not produce a higher incidence of malalignment and resulted in a greater postoperative range of motion than the open technique. The current study shows that percutaneous treatment of supracondylar femur fractures is possible and can decrease operative times, blood loss, the need for bone grafting, increase rates of union, and improve functional outcomes. PMID- 10853154 TI - Percutaneous methods of tibial plateau fixation. AB - Various methods of percutaneous fixation of tibial plateau fractures are available. The optimal method of fixation is dictated by soft tissue injury, fracture characteristics, and functional demands of the patient. Unicondylar fractures are amenable to percutaneous stabilization with screws or plates although some fractures are best approached with open techniques. Hybrid and ring external fixators are most appropriate for patients with bicondylar injuries who have severe soft tissue trauma. Use of intramedullary nails to align ipsilateral shaft fractures adjacent to percutaneously fixed plateau injuries remains controversial but may be indicated for some patients with bicondylar lesions and combined plateau and shaft fractures. PMID- 10853155 TI - Treatment of complex tibial periarticular fractures using percutaneous techniques. AB - Open reduction and internal fixation typically is reserved for the treatment of patients with articular or periarticular tibia fractures, or other tibial injuries that are treated inadequately with intramedullary nailing. This approach can result in extensive dissection and tissue devitalization. By modifying the method of fixation, the plating of tibial fractures has been expanded using a percutaneous technique. Using this approach, the fracture is reduced indirectly and plates are placed through subcutaneous or submuscular tunnels through limited incisions. Between 1992 and 1998, 17 patients with tibial shaft fractures and associated severe soft tissue injury, were treated using a percutaneous plating technique. Followup was available in 14 patients. Six patients required bone grafting procedures for delayed union or nonunion, although four of these patients had significant bone loss related to their injury. There were no malunions. Three patients had superficial infections related to external fixator pin sites and one patient had osteomyelitis develop. Percutaneous plating of the tibia offers an alternative method for stabilizing complex fractures with severely compromised soft tissues, especially those injuries with periarticular extension. This technique is thought to cause no increase in the risk of infection or soft tissue damage and permits rapid mobilization of the limb and patient. When using this treatment for patients with significant bone loss, bone grafting should be considered. PMID- 10853156 TI - Pilon fractures. Treatment protocol based on severity of soft tissue injury. AB - One hundred seven pilon fractures in 107 patients were treated according to a staged prospective protocol. All pilon fractures were stabilized immediately by the application of calcaneal traction. Open fractures or fractures in patients with multiple injuries were stabilized with traveling traction that was applied in the operating room. A distraction computed tomography scan was obtained before definitive treatment. Treatment groups were based on the degree of soft tissue compromise. Forty-one patients with Tscherne Grade 0 or Grade I injuries underwent open reduction and internal fixation (open plating) using contemporary techniques and low-profile implants. Sixty-four patients with Tscherne Grade II and Grade III closed injuries and all patients with open fractures underwent definitive treatment with limited open reduction and stabilization using small wire circular external fixators. Clinical and radiographic evaluations were performed at an average 4.9 years after injury. For all fracture types (AO classification), 81% of the patients who were treated with external fixation and 75% of the patients who were treated with open plating had good or excellent results. For severe fracture patterns (Type C), patients in both groups had significantly poorer results than patients with Types A and B fractures. The patients in the open plating group had a significantly higher rate of nonunion, malunion, and severe wound complications compared with the patients who received external fixation for Type C fracture patterns. Because of the increased incidence of bony and soft tissue complications when treating open or closed Type C fractures, use of limited exposures and stabilization with small wire circular external fixators is recommended. PMID- 10853157 TI - Percutaneous treatment of calcaneal fractures. AB - Percutaneous fixation of calcaneal fractures has limited indications. It is most useful for tongue-type fractures in which the displaced portion of posterior facet remains intact to the tuberosity. This allows the tuberosity to be used as a reduction tool for the posterior facet. The technique has been used successfully in 41 patients. In the current study, the indications and technique are reviewed in detail. PMID- 10853158 TI - Percutaneous fixation of proximal humeral fractures. AB - The purpose of the current study is to evaluate the technique of closed reduction and percutaneous pinning of proximal humeral fractures and to determine whether this technique provides enough stability to permit early active range of motion and subsequent fracture healing. Fractures were classified according to Neer et al and were included if the surgical or anatomic neck were angulated greater than 45 degrees, separation between fragments was greater than 1 cm, or the greater tuberosity was displaced more than 0.5 cm. There were 21 Type II, 16 Type III, and four Type IV fractures. Fractures were pinned using distally threaded Dynamic Hip Screw guide pins, 2-mm Kirschner wires, or 2.5-mm distally threaded Schantz pins. Patients were evaluated for union rates and motion. Assessment was made using the Modified American Shoulder and Elbow Surgeons Form. Thirty-six patients with 37 fractures were available for review with followup averaging 40 months (range, 12-68 months). All patients with Neer Type IV fractures did not respond to fixation and three had avascular necrosis develop, irrespective of the type of pin used. In the remaining 33 patients with Neer Type II and Type III fractures, a union rate of 94% was observed at an average of 2.6 months. All patients had good functional results. In the current series, there were no failures using Schantz pins. There was a 20% failure rate with Dynamic Hip Screw pins (2% if the patients with Type IV fractures were excluded) and a 100% failure rate with Kirschner wires. Stable fixation with early motion and subsequently good results can be obtained using percutaneous fixation in patients with Type II and Type III fractures; however, terminally threaded pins must be used and smooth Kirschner wires must be avoided. Percutaneous fixation cannot be recommended in patients with Type IV fractures. PMID- 10853159 TI - Percutaneous and limited open fixation of fractures of the distal radius. AB - Percutaneous and limited open fixation of fractures of the distal radius is an important method of treatment for many unstable fractures such as unstable dorsal bending fractures, shearing fractures of the radial styloid and lunate facet, and simple articular fractures. The quality of the reduction is monitored with image intensification and the tactic of the reduction is based on manipulation of the fracture fragments by longitudinal traction, percutaneous manipulation, and in some instances by direct manipulation through small incisions. The role of arthroscopy remains uncertain and may represent an alternative to open exposure of the articular surface in some patients although bone grafting may be necessary in patients with fractures with significant metaphyseal defects. PMID- 10853160 TI - Percutaneous treatment of carpal, metacarpal, and phalangeal injuries. AB - Percutaneous fixation of hand fractures is a common technique that takes advantage of the subcutaneous nature of hand bones, their small size, and their limited loading potential for stress placed on hardware. Percutaneous wire fixation supplements cast fixation when plaster cannot hold particular reductions, and allow surgical fixation with limited postoperative swelling. In the first part of the current study, the types of wires that are used for hand fixation, fluoroscopy, helpful instruments, and the basic techniques used for this type of surgery are discussed. In the second part of the study, specific fixation techniques for different fractures of the carpals, metacarpals, and phalanges are outlined. PMID- 10853161 TI - The effects, risks, and guidelines for radiation use in orthopaedic surgery. AB - Radiation is used during orthopaedic surgery in more than 15 million studies performed yearly. The biologic effects of radiation have been shown to inhibit mitosis by producing irrepairable deoxyribonucleic acid double strand breaks or create structural changes by damaging the nucleus, thereby producing potential genetic transmissions. Although human cells are thought to be resistant to malignant change and no studies have shown toxic effects resulting from long-term exposure to low-dose radiation, risks still are assumed. To decrease all risks, radiographic units should undergo periodic calibration, surgeons should wear protective devices, increase their working distance from the x-ray beam, and limit their duration of radiation exposure by making certain that they follow the guidelines set forth by the National Council for Radiation Protection and Measurement. PMID- 10853162 TI - Deformity correction and reconstructive procedures using percutaneous techniques. AB - Corrections of deformities in adults traditionally have been performed with open exposure of the lesions, precise bone cuts, rigid plate fixation, and delayed weightbearing. By obtaining a good preoperative evaluation, and meticulously planning each step of the corrective sequence, good technical results and improved functional capabilities can be attained using percutaneous techniques. PMID- 10853163 TI - Arthroscopic treatment for limitation of motion of the elbow. AB - This study describes the long-term clinical results and serial changes of postoperative range of motion after arthroscopic treatment for limitation of motion of the elbow. Sixty-three patients with limitation of motion of the elbow were treated with arthroscopic procedures. The total range of motion was 79 degrees before surgery. The range of motion showed a progressive increase until 1 year after surgery (mean, 121 degrees). However, after 1 year, the range of motion showed little additional increase. The range of motion acquired during surgery (mean, 122 degrees) usually was the same range that patients achieved during the rehabilitation period (mean, 122 degrees at an average 42.5 months of followup). Extension improved an average of 21 degrees, and flexion increased an average of 23 degrees. The range of motion showed more improvement in patients whose duration of symptoms was less than 1 year (49 degrees) than in those whose duration of symptoms was longer than 1 year (30 degrees). Patients with posttraumatic stiffness had more marked limitation of extension and decreased total range of motion (73 degrees) than did those with degenerative stiffness (86 degrees) before surgery. However, no significant difference existed in the postoperative total range of motion (posttraumatic stiffness, 123 degrees; and degenerative stiffness, 121 degrees). Based on the authors' experience, 92% of patients obtained significant improvement in range of motion after arthroscopic procedures. The minimally invasive nature of elbow arthroscopy is a reproducible and effective procedure for limitation of motion of the elbow with minimal morbidity. PMID- 10853164 TI - Epidural corticosteroid injection in the conservative management of sciatica. AB - In this prospective randomized clinical trial, the results of epidural corticosteroid injections were evaluated in patients with lumbosciatic pain caused by herniated nucleus pulposus. Thirty-six patients with radicular lumbosciatic pain and positive straight leg raising test because of confirmed prolapsed intervertebral lumbar discs were randomized into two groups with (17 patients) and without (19 patients) epidural corticosteroid injection. Members of the treatment groups received three injections of 100 mg methylprednisolone in 10 mL bupivacaine 0.25% each. Additional therapy was standardized and identical in both groups. Followup examinations were performed at 2 weeks, 6 weeks, and 6 months. The examinations included pain level (visual analogue scale), straight leg raising test, and functional status (Hannover Functional Ability Questionnaire). At 2 weeks, patients receiving methylprednisolone injection showed a significant improvement in straight leg raising test results compared with patients in the control group. Results were better in the methylprednisolone group, although not statistically significant for pain relief and mobility. At 6 weeks and 6 months, pain relief, improvement of straight leg raising, and improvement of functional status showed no statistical significance. Epidural corticosteroid injections can be recommended as additional therapy only in the acute phase of the conservative management of lumbosciatic pain. PMID- 10853165 TI - Survival of hip replacements. A comparison of a randomized trial and a registry. AB - At the authors' hospital, 410 primary total hip replacements were performed on 372 patients between September 1, 1985, and May 31, 1989. All hips were assigned randomly to receive a Charnley prosthesis with an ogee flanged cup or a Spectron prosthesis with a metal backed cup. Eleven-year survivor analysis, using revision as the end point definition of failure, revealed a survival rate of 93.2% +/- 5.8% for the Charnley replacement and 95.9% +/- 3.0% for the Spectron. If each component of the systems was analyzed (concerning aseptic loosening), the ogee cup and the Spectron stem had 100% survival. The survivorship for all 410 hips was 94.5% +/- 3.4%. If the end point definition of failure was expanded to include patient dissatisfaction, the survival rate decreased to 86.3% +/- 4.9%. These survival rates were compared with the rates obtained by the Swedish National Hip Registry. The national cohort included all patients in Sweden who were treated surgically with a Charnley (14,053 patients) or Spectron (metal backed cup) prosthesis (726 patients) between September 1, 1985, and May 31, 1989. Eleven-year survivor analysis revealed a national survival rate of 92.1% +/ 0.7% for the Charnley replacement and 88.6% +/- 6.1% for the Spectron. The analyses from the Swedish Registry are based on more than 160,000 primary operations and 11,500 revisions. Despite the enormous amount of data, there are drawbacks, and registries never can replace the prospective, randomized trial. One reason is the Swedish National Registry is unable to discriminate between the individual cup and stem components when analyzing the cause of revision, and no clinical or radiographic information is collected. A potential drawback for the randomized trial is performance bias because surgeons from specialized centers might perform better than the general orthopaedic surgeon. PMID- 10853166 TI - Deep venous thrombosis after total hip or total knee arthroplasty in patients in Japan. AB - A single center, prospective, epidemiologic study was conducted to estimate the incidence of deep venous thrombosis detected by venography in patients in Japan undergoing total hip arthroplasty or total knee arthroplasty without prophylactic anticoagulant therapy. Venograms of 164 patients who had total hip arthroplasty and 138 patients who had total knee arthroplasty were evaluated. The incidences of deep venous thrombosis were 22.6% in patients who had total hip arthroplasty and 48.6% in those who had total knee arthroplasty. The incidences of proximal deep venous thrombosis were 9.8% in patients who had total hip arthroplasty and 14.5% in those who had total knee arthroplasty. Statistical analysis revealed that the type of operation influenced the development of deep venous thrombosis. Patients who had total knee arthroplasty were 3.2 times more likely to have deep venous thrombosis develop than were patients who had total hip arthroplasty. Body mass index and age were identified as statistically significant risk factors. PMID- 10853167 TI - Total knee arthroplasty after high tibial osteotomy. AB - Between 1980 and 1995, 95 consecutive total knee replacements were performed at an average of 10 years 4 months after high tibial osteotomy. The average age of the 82 patients was 66 years, with a preoperative diagnosis of osteoarthritis in 94 knees. One patient died 6 months after surgery. The followup of the remaining 81 patients (94 knees) averaged 8.6 years (range, 2-17 years). Knee Society knee score at final followup improved to an average of 87.6 points from a preoperative average of 38.1 points. No pain was present in 86.2% of knees, and 12.8% of knees had only mild or occasional pain. Tibial radiolucencies were identified in 12 (12.8%) knees at final followup, and in only four knees were radiolucent lines found about the lateral zones. Only one tibial component required revision 3 years after surgery. Although no preoperative factor was identified that predisposed to an inferior knee score, function score, or pain score, the severity of the preoperative flexion contracture and the number of previous surgeries did relate to diminished postoperative motion. However, an increased number of patellar radiolucencies were seen in the knees in which the lateral joint line was raised (referenced from the fibular head) a greater degree. The clinical results of total knee replacement after high tibial osteotomy appeared similar to those of primary total knee replacement. The previous high tibial osteotomy had no adverse effect on the eventual results of a cemented posterior cruciate retaining total knee replacement. PMID- 10853168 TI - Effects of lateral-wedged insoles on kinetics at the knee. AB - Lateral-wedged insoles have been shown to help clinically alleviate pain associated with medial compartment osteoarthritis. This study analyzed the effects of lateral-wedged insoles on the gait and medial knee compartment load of 17 healthy subjects. Three-dimensional gait analysis was performed for each subject with and without wearing a 5 degrees lateral-wedged insole. Subjects walked at a constant velocity for both conditions. A motion analysis system and force plate were used to calculate temporal and spatial parameters, joint angles, moments, and powers. An analytical model was developed to estimate medial compartment loads at the knee for each subject during both conditions. Results were compared with a Student's paired t test. There were no significant differences in temporal and spatial parameters, joint angles at the hip, knee, and ankle, or kinetics at the hip and ankle. However, the external varus moment and estimated medial compartment load at the knee were reduced significantly with the addition of the lateral-wedged insole. These results suggest that the pain relief and improvement in function reported by patients with osteoarthritis while using lateral-wedged insoles may be achieved by a reduction in external varus moment and medial compartment load. PMID- 10853169 TI - Peroneal latency in normal and injured ankles at varying angles of perturbation. AB - The aim of this study was to determine whether there was a difference in latency of the peroneus longus muscle at varying amplitudes of ankle inversion perturbation and between individuals with and without a history of ankle injury. Thirty-four male athletes from different football codes (soccer, rugby) received four random tilts to their left ankles at 5 degrees, 10 degrees, and 15 degrees in the frontal plane on a dual platform trap door. Peroneal latency was defined as the time difference between onset of the trap door movement, as detected by an accelerometer, and the onset of muscle activation above a resting baseline, as recorded using surface electromyography. Latency was determined using an algorithm. A series of repeated measures analyses of variance indicated that the latency was reliable between trials. There was no statistical evidence that history of injury or subjective ankle instability influenced the latency; however, there was a systematic difference between dominant and nondominant legs (dominant, 6.3 ms faster), and there was a small systematic effect (3 ms) for the angle of inversion perturbation. Muscle latency responses in male football players are thought to be influenced more by dominance than by history of injury or amplitude of perturbation. PMID- 10853170 TI - Lower extremity deformities associated with thrombocytopenia and absent radius syndrome. AB - The cases of 11 patients with the syndrome of thrombocytopenia and absent radius (TAR syndrome) who presented at two institutions between 1970 and 1996 were reviewed. Knee deformities have been well documented in thrombocytopenia and absent radius syndrome, but an inordinate frequency of other lower extremity problems needing orthopaedic attention was seen. Five of the 11 patients with thrombocytopenia and absent radius syndrome studied had knee deformities, but seven of the 11 had 11 other lower extremity deformities. This article documents all of the lower extremity problems seen in the seven patients with thrombocytopenia and absent radius syndrome with anomalies other than those of the knee. PMID- 10853171 TI - Extraabdominal desmoid tumor. A study of 83 cases. AB - One hundred three patients with extraabdominal desmoid tumor were treated between 1970 and 1996 at the authors' institution. Among these, 15 patients were lost to followup and five were excluded because they had less than 1-year followup. The remaining 83 patients were followed up for a mean of 11.2 years. Thirty-seven (44.6%) patients experienced local recurrence on average 1.8 years after treatment. There was no difference in the incidence of recurrence between the two groups treated with surgery only (45.3%) or with adjuvant radiation therapy administered after inadequate surgical resection of the tumor margins (41.2%). Recurrence was not related to age, gender, and site. None of the 83 patients died of the disease. For recurrent but stable lesions, clinical observation alone may be considered. PMID- 10853172 TI - Complications of irradiated allografts in orthopaedic tumor surgery. AB - Massive structural allografts used for replacement of bone defects after removal of bone tumors have several complications, including fracture, infection, and nonunion. To decrease the rate of infection, irradiation of selected allografts before their implantation was performed. This study evaluated the complications in patients with these irradiated grafts. Twenty-four patients were identified who had received allografts from 1987 through 1991 that were irradiated before implantation. The dosage of radiation was between 10 kGy and 30 kGy. The mean length of followup of the patients was 5 years (range, 2-9 years). These grafts were compared with a control group of grafts that were not irradiated but were implanted during the same time and used for similar diagnostic problems with defects of similar size. The outcomes of the groups differed significantly only in the incidence of allograft fracture. These findings indicate that high-dose irradiation to bone allografts is associated with a higher rate of fracture than are similar reconstructions using nonirradiated allografts. PMID- 10853173 TI - Reconstruction of hip stability after proximal and total femur resections. AB - Dislocation is the most common complication after proximal and total femur endoprosthetic reconstruction. The current study describes a surgical technique of acetabular preservation and reconstruction of the joint capsule and abductor mechanism that recreates joint stability and avoids dislocation. Between 1980 and 1996, 57 patients underwent proximal or total femur resection with endoprosthetic reconstruction. Forty-six patients had primary sarcoma of bone, nine had other bone tumors, and two had metabolic bone disease. The acetabulum was spared and not resurfaced in all patients. Bipolar hemiarthroplasty was performed in 49 patients, and fixed unipolar hemiarthroplasty was performed in eight. Soft tissue reconstruction included Dacron tape capsulorrhaphy over the prosthetic neck, reattachment of the abductor mechanism to the prosthesis, and extracortical bone fixation. The average followup period was 6.5 years (range, 2-18.2 years). Dislocation occurred in only one (1.7%) patient, and aseptic prosthetic loosening occurred in three (5.3%) patients. Four patients with primary bone sarcoma had local recurrence, of whom one required amputation of the limb. The limb salvage rate was 98%. Eighty-one percent of the patients had a good to excellent functional outcome. Acetabular preservation, capsulorrhaphy, and reconstruction of the abductor mechanism recreate hip stability and avoid dislocation after proximal and total femur endoprosthetic reconstruction. PMID- 10853174 TI - Tibial plateau fractures. A new classification scheme. AB - Fractures of the tibial plateaus are common injuries. Various classification schemes have been used to describe these injuries. Although each system has its own purpose, the simpler systems do not allow comparison with more complex divisions. The problem is compounded by the variable use of adjectives that describe these fractures. A comprehensive classification of tibial plateau fractures should group fractures that are similar in topography, morphology, and pathogenesis, requiring similar treatment, and having a similar prognosis. Fracture dislocations and standard tibial plateau fractures should be incorporated into a single classification to avoid the use of two complementary classifications. Any such classification should not be difficult to remember or to use. Keeping in mind these requirements, the authors devised a simple yet comprehensive classification. The authors studied 80 cases of tibial plateau fractures from January 1988 to September 1997, and used contemporary classifications of tibial plateau fractures as a database to formulate the new classification. A new fracture, subcondylar bicondylar with coronal split, has been classified for the first time. An alphanumeric system has been developed that has made nomenclature easy to remember and use. An effort has been made to address the profoundly confusing issue of variable adjectives that describe these injuries. A review of the literature shows that fractures in the authors' classification have been grouped according to similar pathomechanics, treatment, and functional results. PMID- 10853175 TI - Influenza A induced rhabdomyolysis resulting in extensive compartment syndrome. AB - This is a case of influenza A induced rhabdomyolysis resulting in extensive compartment syndrome and acute renal failure in a 10-year-old child. The patient required fasciotomies in all four extremities. Even after fasciotomies were performed, the muscle tissue continued to swell, suggesting a primary myositis. This case emphasizes the importance of considering the diagnosis of compartment syndrome in patients with influenza infection and severe myalgia. PMID- 10853176 TI - Mechanics of the deltoid muscle. A new approach. AB - The Inman concept of the mechanics of the deltoid describes a vertical upward oriented traction exerted on the upper end of the humerus at the beginning of arm elevation. However, Duchenne de Boulogne showed that the middle deltoid pushes the head downward. When the arm is at rest, the trajectory of the middle deltoid fibers changes by more than 90 degrees so that the humeral head is enveloped by the muscle; this suggests that the deltoid acts on the humeral head like a cable on a pulley. The authors studied the area of contact between the deltoid and the humeral head in three-dimensional reconstructed shoulders. A new model, which includes the pulley effect, was designed to explore the resultant total force applied by the deltoid onto the humerus. In most cases the resultant vertical force was oriented downward. Thus, the conventional model is not complete. The current model indicates that the deltoid prevents upward migration of the humeral head and compresses it against the glenoid. This explains why many shoulders function well despite a massive cuff tear. This also implies that reeducation of the deltoid is a major aspect of the rehabilitation for patients with a rotator cuff tear. PMID- 10853177 TI - Mechanical and histologic evaluation of Collagraft in an ovine lumbar fusion model. AB - The purpose of this study was to determine the effectiveness of a composite material composed of Type I bovine dermal collagen, 65% hydroxyapatite, and 35% tricalcium phosphate ceramic (Collagraft Bone Graft Matrix Strip NeuColl Incorporated, Palo Alto, CA) as a bone graft substitute for spinal fusion with and without the use of autologous bone marrow in an ovine lumbar spine model with pedicle screw fixation. Twenty-four adult sheep underwent a single level posterolateral (intertransverse process) L3-L4 lumbar fusion with one of three graft materials combined with rigid pedicle screw fixation. The three graft materials were Collagraft, Collagraft with marrow, and autogenous corticocancellous bone graft. Animals were euthanized 6 months after surgery and evaluated using dual energy x-ray absorptiometry, radiographs, histologic analysis, and mechanical testing. Dual energy xray absorptiometry between the transverse processes revealed that the mineral densities for the two Collagraft groups were significantly higher than the autogenous bone graft group. Histologic analysis confirmed that Collagraft was highly compatible and was well incorporated into the fusion mass. Both Collagraft groups had thick trabeculae and a mixture of lamellar and plexiform bone. The autogenous bone graft group had a smaller fusion complex, composed primarily of lamellar bone with thinner and fewer trabeculae. All three groups had similar mechanical properties. These results support the use of Collagraft in spinal fusion with pedicle screw fixation. PMID- 10853178 TI - Effect of suture size on locking and grasping flexor tendon repair techniques. AB - An experimental study was performed using human cadaver flexor tendons to investigate the effect of locking and grasping loop techniques on the tensile properties of repaired flexor tendons, which closely resemble the clinical model. Statistically significant improvement was observed only with the locking loop technique for ultimate and gap strength values using 2-0 core suture and ultimate strength values using 3-0 core suture. There was no statistically significant increase in tensile strength values using 4-0 core suture material. A heavier core suture used with the locking loop technique provided greater ultimate and gap strength of a repaired tendon than when used with the grasping loop technique. PMID- 10853179 TI - The effects of processing and low dose irradiation on cortical bone grafts. AB - The authors studied the effects of standard processing and preprocessing low dose gamma irradiation (1.5 Mrad) on the strength and incorporation of syngeneic and allogeneic cortical bone grafts. Bilateral femoral middiaphyseal 8-mm segmental defects in 120 male Fisher rats were stabilized with internal fixation. Each defect received one of six types of grafts: fresh syngeneic, processed syngeneic, irradiated processed syngeneic, fresh allogeneic, processed allogeneic, and irradiated processed allogeneic grafts. Graft processing included soaking in 70% ethanol and deep freezing for preservation. Irradiation was performed by 60Co source immediately before processing. Grafts were evaluated by histologic analysis, histomorphometric analysis, and biomechanical testing at 4 and 6 months after surgery. Graft treatment, either processing or irradiation processing, did not affect consistently or significantly the incorporation of syngeneic or allogeneic grafts. Graft allogenicity was the major determinant of the revascularization and the histologic pattern of graft incorporation. Processed and irradiated processed allogeneic grafts gained compressive strength with time and were as strong as syngeneic grafts at 6 months. Biomechanical and histologic data from this study suggest that standard processing and preprocessing low dose irradiation do not compromise the natural course of allogeneic cortical bone graft incorporation. PMID- 10853180 TI - Comparative effects of laser and radiofrequency energy on joint capsule. AB - The study compared the effects of laser and monopolar radiofrequency energy on thermal and architectural properties of joint capsular tissue in an in vitro ovine model. Sheep glenohumeral joint capsular specimens were treated with laser (5, 10, 15 W) or radiofrequency energy (55 degrees, 65 degrees, 75 degrees C) (n = six per group). Energy application caused significant tissue shrinkage and decreased surface area in all laser and radiofrequency treatment groups. Tissue thickness significantly increased in all treatment groups except for radiofrequency 55 degrees C. Tissue shrinkage, surface area, and thickness each correlated significantly with the delivered laser energy per tissue area or mean radiofrequency probe temperature. There were no significant differences among laser 10 W, laser 15 W, and radiofrequency 75 degrees C treatment groups for these three architectural parameters. Tissue temperature was elevated significantly in the laser 10 W, laser 15 W, radiofrequency 65 degrees C, and radiofrequency 75 degrees C groups when compared with the control. Tissue temperature changes between the laser 10 W and radiofrequency 75 degrees C groups were similar; however, laser treatment produced a steeper temperature increase accompanying its peak temperature. Despite different mechanisms, laser and radiofrequency energy can achieve similar and predictable tissue modification, which is temperature dependent. Additional in vivo studies must be performed to evaluate the applicability of these techniques to clinical use. PMID- 10853181 TI - Osteogenesis by recombinant human bone morphogenetic protein-2 at skeletal sites. AB - Osteogenesis was evaluated in the mandibular bone by combinations of various dosages of recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite (four groups: Group I, 2 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Group II, 10 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Group III, 50 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Control Group, only atelopeptide Type I collagen and porous hydroxyapatite). The prepared materials were implanted in the mandibular bone hole (7 mm in diameter, 2 mm deep). Three weeks later, the alkaline phosphatase activity in the implanted region was determined, and the histologic features of the excised tissue were examined. There were significant differences in histologic and biochemical findings among the four groups. In the recombinant human bone morphogenetic protein-2 implanted groups, osteogenesis increased with the dosage of recombinant human bone morphogenetic protein-2, as assessed by alkaline phosphatase activity and histologic findings. The results suggest that atelopeptide Type I collagen is an effective carrier for recombinant human bone morphogenetic protein-2 and that porous hydroxyapatite would be advantageous for clinical application as a material to maintain its original form after implantation. PMID- 10853182 TI - The effects of conformity and load in total knee replacement. AB - The effects of different conformity ratios and loads on the ultrahigh molecular weight polyethylene stress levels acting on knee implants were examined using a nonlinear, finite element analysis. The contact condition between a rigid cylinder with a radius of 30 mm and a polyethylene plate was modeled. Nonlinear behavior of polyethylene was assumed. The polyethylene plate was constructed with varying radii, with a minimal thickness of 6 mm and with a width of 40 mm. The ratio of the cylinder radius to the radius of the polyethylene plate was defined as the conformity ratio; a conformity ratio of 0 represented a flat tibial inlay, whereas the highest ratio modeled of 0.99 was nearly conforming. The conformity ratios modeled were 0, 0.2, 0.4, 0.6, 0.7, 0.8, 0.9, 0.95, and 0.99. The loads applied were 1000 N, 2000 N, 3000 N, 4000 N, 5000 N, and 6000 N. The effects of different conformity ratios and loads on the contact area (mm2), the compressive surface stress (MPa), the shear stress (MPa), and the von Mises stress (MPa) were investigated. It was found that all of these parameters were affected by changes to the conformity ratio and to a lesser extent by load changes. That is, increasing the load from 3000 N to 6000 N resulted in a surface and shear stress increase lower than the increase in stress caused by the small change of the conformity ratio from 0.99 to 0.95. The effect of an increasing conformity ratio on the reduction in stress was more pronounced for conformity ratios above 0.8. In addition, the effect of a load increase for a flat tibial inlay was two times greater than for one with near full conformity. PMID- 10853183 TI - Effects of a cane on floor reaction force and center of force during gait. AB - In comparative studies of various gait patterns of 20 healthy subjects who used a cane, the vertical reaction forces and the center of force on the foot were measured and recorded by a force recording and analyzing device. The results indicated that when a cane was used in the ipsilateral hand, the center of force did not shift significantly compared with normal gait. When a cane was used in the contralateral hand, the center of force shifted medially compared with normal gait. In analysis of the vertical floor reaction force acting on the foot, the most efficient way to use a cane was to control the pacing so that the tip of the cane and the foot touched the ground simultaneously. By doing so, the cane could share as much as 34.3% of force at heel strike, 25.3% at midstance, and 29.7% at toeoff of the stance phase of the gait cycle. When prescribing use of a cane for a patient with varus gonarthritis, the patient should be instructed to use the cane in the ipsilateral hand so as not to shift the center of force medially; for a patient with valgus gonarthritis, the cane should be used in the contralateral hand to shift the center of force medially. Patients should be taught to control pacing so that the tip of the cane and the foot touch the ground simultaneously. PMID- 10853184 TI - Multiple masses in a 2-year 5-month old girl. PMID- 10853185 TI - The Marshall R. Urist Young Investigator Award. Orthopaedic applications of gene therapy. From concept to clinic. AB - Gene therapy offers new possibilities for the clinical management of orthopaedic conditions that are difficult to treat by traditional surgical or medical means. To bring the potential of this novel technology into the clinic, a research program was initiated that aimed to identify orthopaedically useful genes and develop methods for delivering them to suitable sites under conditions in which gene expression remains at therapeutic levels for the appropriate periods of time; this program is now 10 years old. Rheumatoid arthritis was selected as the lead disease. Preclinical studies evaluating the local and systemic delivery of numerous different genes by in vivo and ex vivo methods in murine and lapin models led to the development of a human gene therapy protocol for arthritis. In this protocol, a gene encoding the human interleukin-1 receptor antagonist protein is transferred to the metacarpophalangeal joints of female patients with rheumatoid arthritis. The first patient was treated this way in July 1996. This is not only the first orthopaedic application of human gene therapy, but also the first use of gene therapy approved for a nonlethal disease. In addition to providing additional therapeutic options for the treatment of rheumatoid arthritis, the experimental data from this study suggest that gene transfer approaches may improve the treatment of osteoarthritis, the repair of cartilage, ligaments, tendons, menisci, intervertebral discs and bone, and the management of disorders such as osteoporosis and osteogenesis imperfecta. They also show promise as a means for developing novel and improved animal models of orthopaedic diseases. If the current rate of progress continues, wide clinical application of gene therapy in various orthopaedic indications should occur within the next 5 to 10 years. PMID- 10853186 TI - Three-dimensional collagen gel culture promotes osteoblastic phenotype in bone marrow derived cells. AB - When calvarial cells or osteogenic cell lines were cultured in type I collagen gel, calcification was observed early and diffusely compared to monolayer culture. Bone marrow derived cells were cultured in three-dimensional type I collagen gel to investigate whether the cells can differentiate into osteogenic cells. In terms of efficient induction of osteogenic differentiation, we compared collagen gel culture to type I collagen coated dish culture and collagen-free plastic dish culture by morphology, alkaline phosphatase activity, and mRNA expression for type I collagen and osteopontin. Bone marrow derived primary cells formed colonies consisting of fibroblastic cells positively expressing alkaline phosphatase activity. Mineral deposition was observed in both primary and the 3rd passaged cells cultured in collagen gel, whereas the 3rd passaged cells on plastic dishes failed to be mineralized. Cells in collagen gel showed higher alkaline phosphatase activity than those in the other two methods suggesting that three-dimensional collagen network stimulated osteoblastic differentiation effectively. The expression level for type I collagen mRNA of the cells in collagen gel was three times higher in the 3rd passaged cells, and was slightly decreased in primary cells compared to the other two methods. The osteopontin mRNA expression of the cells in collagen gel was four times higher in the 3rd passaged cell culture but lower in primary cell cultures. These results suggested that collagen gel culture might be a useful environment for osteogenic induction of passaged cells derived from bone marrow. PMID- 10853187 TI - Meningioma presented as subarachnoid haemorrhage: case report. AB - A case of parasagittal meningioma causing subarachnoidal haemorrhage (SAH) is reported. Computed tomography (CT) was found negative in the patient with acute severe headache and haemorrhage was observed on cerebrospinal fluid (CSF) examination. Digital subtraction angiography (DSA) showed an avascular space over the convexity and Magnetic resonance imaging (MRI) revealed the tumour. The importance of MRI for the detection of underlying pathology in SAH with unknown aetiology is emphasised. PMID- 10853188 TI - Leptin stimulates basal and GHRH-induced GH release from cultured rat anterior pituitary cells in vitro. AB - Leptin, the product of ob gene, is secreted from adipocytes and appears to regulate food intake and energy expenditure through its receptor in the hypothalamus. In addition, leptin is reported to modulate pituitary hormones including LH, FSH and ACTH, probably via the hypothalamus. In obesity, growth hormone (GH) secretion is impaired, while serum leptin levels are elevated. To investigate the possibility that leptin serves as a metabolic signal that influences GH secretion from the pituitary, we studied the effect of leptin on GH secretion from primary monolayer cultures of rat anterior pituitary cells. Twenty micrograms/ml leptin increased GH secretion, but did not increase GH release at a physiological concentration up to 200 ng/ml. However, 200 ng/ml leptin stimulated GH release in the presence of GH-releasing hormone (GHRH) ranging from 10(-8) to 10(-7) M. Twenty ng/ml leptin tended to increase 10(-7) M GHRH-induced GH release, whereas 20 ng/ml leptin did not increase either 10(-9) or 10(-8) M GHRH induced GH release. This result suggests that leptin has a direct effect on the pituitary to enhance GHRH-induced GH secretion. PMID- 10853189 TI - Experimental study on paratumoral injection of cisplatin-loaded microspheres for gastric cancer. AB - Microspheres from glycolide-L-lactide copolymers incorporating cisplatin (CDDP MS) were prepared to evaluate the sustained release and anticancer effect by paratumoral injection on the gastric cancer with regional lymphnode metastases induced by VX2 tumor in rabbits. In the first set of experiment, the rabbits were divided into three groups subjected to treatment and compared the tissue cisplatin distribution. In the first group (CDDP-MS pt group), 1 mg/kg of cisplatin was administered by the method of paratumoral injection in the form of CDDP-MS. In the second group (CDDP solution pt group), the same dose was given in the form of CDDP aqueous solution in the same way and in the third group (CDDP solution i.v. group), the same dose was intravenously administered. In the second set of experiment, after twice of each therapy the anticancer effects were compared between CDDP-MS pt and CDDP solution i.v. groups. In results, the platinum concentrations of the tumor and regional lymphnodes were 3.14 +/- 6.22, 0.65 +/- 0.79 micrograms/g in the first group, 0.43 +/- 0.39, 0.16 +/- 0.16 microgram/g in the second group and 0.03 +/- 0.01, 0.07 +/- 0.05 microgram/g in the third group, respectively. PMID- 10853190 TI - [Nephrology in Spain in the 21st century]. PMID- 10853191 TI - [Hypertensive crises in hemodialysis: and now, what should we do?]. PMID- 10853192 TI - [The nephrology white book in Spain (I). Spanish Society of Nephrology]. PMID- 10853193 TI - [Heterozygosity loss and somatic mutations in type I and II dominant autosomal renal polycystic kidney disease: evidence of a recessive mechanism at a cell level in cystogenesis]. AB - Autosomal dominant polycystic kidney disease (ADPKD) is a systemic disorder mainly characterized by renal cyst formation. Cysts in ADPKD are focal in nature, since only a small fraction of nephrons become cystic. The hypothesis that a second hit may be required for cyst formation has been proposed. This hypothesis suggests that inactivation of the inherited wild-type allele by a somatic mutation triggers cyst formation. In some cases, this second hit eliminates the normal allele and the affected cells remain with a single allele, which is the inherited mutated copy, and we only visualize one allele after the amplification by polymerase chain reaction; this is called loss of heterozygosity (LOH). In this study we have analysed the DNA isolated from epitehlial cells from 164 cysts of 8 kidneys affected by ADPKD type I and 30 cysts form a kidney affected by ADPKD type II. We have demonstrated the presence of LOH in 20.1% of PKD1 cysts and in 10% of PKD2 cysts. We have also found eight other different mutations in PKD2 cysts without LOH; so the percentage of somatic mutations in the PKD2 kidney reaches 36.6% of cysts. In conclusion, our data suggest that a recessive mechanism at the cellular level is implicated in cyst formation in the PKD1 and the PKD2 disease. The loss of both copies of the gene triggers the proliferation of a single cell, resulting in the cyst formation. PMID- 10853194 TI - [Prevalence of genetic prothrombotic factors (factor V Leiden and II20210 prothrombin mutation) in glomerular nephropathies with or without thrombosis]. AB - The presence of genetic prothrombotic factors (factor V Leiden and the prothrombin II20210 mutation) was investigated in 38 patients with glomerulonephritis with or without a history of thrombotic events and/or nephrotic syndrome. We found an increased prevalence (36%) of heterozygous factor V Leiden in those patients with a history of thrombotic events. This is ten times the prevalence in the normal Spanish population. Carrier status for this mutation may be a determining factor in the development of thrombotic events along with the acquired disorders of coagulation to which these patients are prone. We found only one patient who was a carrier of the G-A II20210 mutation of the prothrombin gene; this patient had no history of venous thrombosis or embolism. Our findings suggest the need to measure activated protein C resistance and to look for the most frequent genotype causing it, Factor V Leiden, in patients with glomerulonephritis to identify those at risk who may benefit from prophylaxis against thrombosis. PMID- 10853195 TI - [Relationship between ionic dialysance and urea clearance]. AB - INTRODUCTION: Ionic dialysance is a method of continuous on-line monitoring of delivered dialysis without blood sampling. To compare the results obtained by ionic dialysance and those obtained by the traditional measurements of the dialysis dose, it is necessary to know the relationship between the ionic dialysance and urea clearance. MATERIAL AND METHODS: Ionic dialysance and the urea clearance were determined in 18 patients (13 dialyzed with cuprophan and 5 patients with AN69). Urea clearance was measured by 6 different methods: urea clearance in whole blood calculated with the arteriovenous difference in the urea concentration rates and the arterial flow measured by the rolling pump (KBAVb) or by ultrasounds (KBAVu); urea clearance in whole blood measured by the urea concentration in the dialysate (KBD); urea blood water clearance measured by the arteriovenous difference in the concentration rates using the arterial flow measured by the roller pump (KwBAVb) or by ultrasounds (KwBAVu) and urea blood water clearance measured by the urea concentration in dialysate (KwBD). RESULTS: The mean arterial flow measured by the roller pump was 314.4 +/- 16.2 ml/min and 275.1 +/- 13.8 ml/min when measured by ultrasounds (p < 0.001). The data of ionic dialysance and urea clearances were as follow (ml/min): ionic dialysance 185.6 +/ 11.7; KBAVb 245.7 +/- 15.7; KBAVu 215.4 +/- 13.2; KBD 231.6 +/- 13.1; KwBAVb 218.1 +/- 14; KwBAVu 191.2 +/- 11.8; KwBD 183.1 +/- 11.7. The absolute difference of ionic dialysance with the KwBAVu was 8.4 +/- 6 ml/min (range between -17.8 and 11.5 ml) and with the KwBD was 7.6 +/- 5.4 ml (range between -12.9 and 21.4 ml). CONCLUSIONS: There was a relationship between ionic dialysance and urea blood water clearance. The best concordance was obtained when the clearance was calculated with the urea concentration of dialysate, or with the arteriovenous difference of the urea concentration rates and the arterial blood flow measured by ultrasounds. PMID- 10853196 TI - [Familial juvenile nephronophthisis (report on 16 families with shared family tree)]. AB - This is a study of a group of 23 patients from 16 families with a shared family tree, developing chronic renal insufficiency (CRI). Out of the 23 patients, 18 were female and five male with an average renal death age of 18.4 years old, showing fevo clinical manifestations. The main reason for consultation was the significant level of anemia. 17 patients had normal arterial tension, 1 patient manifested severe artery hypertension (AHT), 3 manifested mild AHT, and 2 manifested slight AHT. All the patients entered the final stage of CRI with a low level of hemoglobin overaging 6.5 g%. The urinalysis revealed an average SG of 1,010, without proteinuria or with slight proteinuria, lower than 500 mg in 24 hours. Three patients had microhematuria and the remainder had normal urinary sediment. A renal ultrasound in 18 cases revealed a bilateral reduction in the kidney size, loss of the cortcomedullar relation, an increase in the echogenety of the renal parenchyma, scattered in all cases, and the presence of corticomedullar cysts in 5 cases. The histopathological study performed in 8 cases revealed some findings which were compatible with chronic interstitial nephritis with corticomedullar cysts. The findings resemble those described in the literature in cases of familial juvenile nephronophthisis (FJN). An important aspect to be pointed out is the presence of an interstitial infiltrate with mononuclear cells, an even more significant feature than any previously reported. We can conclude that the members of these familial groups are carriers of FJN of recessive autosomic transmission, which, in view of some differences in the clinical presentation, age of onset of, CRI some biochemical and morphological findings, and the absence of genetic alterations as described in type 1 FJN, is a variant of this disease. PMID- 10853197 TI - [Effects of dietary phosphorus restriction on the production of 1,25(OH)2D3 (calcitriol) in patients with moderated renal failure]. AB - Calcitriol deficiency and phosphate retention are two main factors in the pathogenesis of renal hyperparathyroidism. In spite of normal serum levels, phosphate may have an important role even in moderate RI. The aim of this study was to evaluate the effect of dietary phosphorus restriction on serum levels of calcitriol in patients with moderate RI. We studied 21 patients (7 F/14 M); mean age 61.7 +/- 15 years old; corrected creatinine clearance 51.4 +/- 14 ml/m. Serum PTH, calcitriol 25(OH)D3, calcium, phosphorus and urinary excretion of calcium and phosphorus were measured before and after 30 days on phosphorus restricted diet (700 mg/day). RESULTS: [table: see text] CONCLUSIONS: Our patients with moderate RI have elevated serum levels of PTH while calcitriol was in the lower normal range. Dietary phosphorus restriction resulted in a significant decrease in PTH levels and a significant increase in serum calcitriol concentrations. The levels of 25(OH)D3 did not change in this study. PMID- 10853198 TI - [Study of various factors that could have an impact on the treatment with erythropoietin of hemodialysis anemia]. AB - Several factors influence the efficacy of the action of human recombinant erythropoietin during treatment of anaemia in haemodialysis patients. We carried out a six-month prospective study of 23 stable patients who had been on haemodialysis for at least one year to attempt to evaluate those factors modifying the dose of the hormone to attain a similar haematocrit, such as use or not of angiotensin converting enzyme inhibitors, hepatitis C virus positive or negative, age older or younger than 60 years, acquired cystic kidney disease or not, and sex. The patients were treated with subcutaneaous erythropoietin for over a year to attain a haematocrit of 35%, intravenous iron to reach plasma ferritin levels > 250 ng/ml and a transferrin saturation index > 20%, folic acid and group B vitamins. Parameters studied included age, time and duration of haemodialysis, Kt/V, albumin, haematocrit, erythropoietin in U/kg/week, intact PTH, hepatitis C virus, PCR of the hepatitis C virus, transaminases, ferritin, transferrin saturation index, folic acid, vitamin B12, and aluminium. No statistically significant differences were seen between the patients with and without hepatitis or in age or acquired cystic kidney disease and sex in the hormone dose given to achieve similar levels of haematocrit. Higher doses of erythropoietin were necessary in those patients treated with antihypertensive agents (71 +/- 25 vs 44 +/- 25 U/kg/week; p < 0.05). There were no differences between groups in factors known to cause resistance to the action of the hormone. The most important conclusions from this study concern the cost-benefit relation of treating hypertensive patients on haemodialysis with angiotensin converting enzyme inhibitors and erythropoietin. PMID- 10853199 TI - [Factors associated with health related quality of life in patients undergoing renal replacement therapy]. AB - The aim of this study was to investigate the sociodemographic and clinical variables which influence health-relate quality of life (HRQOL) of patients on renal replacement therapy (RRT). A cross-sectional study was carried out with a sample including all patients on hemodialysis (n = 170) and transplant patients (n = 210) of our region. The HRQOL assessment instruments used in this study were: the Spanish versions of the sickness impact profile (SIP) and the SF-36 health survey (SF-36). Sociodemographic and clinical data (including age at start of RRT, age at the interview, gender, hospital, socioeconomic level, educational level, living conditions, inclusion in transplant waiting list, renal disease diagnosis, time in any RRT, hemoglobin, hematocrit, serum urea, creatinine, proteins and albumin, hospital admissions and length of hospital stay during last year), a comorbidity index and the Karnofsky performance scale score step. To investigate which studied variables had independent influence over the HRQOL measures, logistic regression method was employed in the case of the SF-36, and multiple regression, in the case of the SIP. A model was adjusted step by step in each RRT method (hemodialysis and transplantation) for each dimension of the PCE (physical dimension, psychosocial dimension and total score), and for each component summary score of the SF-36 (physical and mental component summary). In patients on hemodialysis, variables associated with better HRQOL were: higher age, female gender, higher educational level, and better functional status; and variables associated with worse HRQOL were: higher number of hospital admissions, and higher comorbidity index. In transplant patients, variables associated with better HRQOL were: higher age and higher functional status; and variables associated with worse HRQOL were: longer time on dialysis before transplant, longer time with functioning transplant, and higher comorbidity index. Despite the independent influence on the HRQOL demonstrated for some of the studied variables, it seems that HRQOL assessment instruments scores may mainly depend on other non-studied variables, and it may be that these instruments evaluate other aspects of the patients which have not been taken into account until now. PMID- 10853200 TI - [Prognosis factors in the clinical course of segmental and focal glomerulosclerosis]. PMID- 10853201 TI - [Heart failure after second vascular port for hemodialysis]. PMID- 10853202 TI - [Use of intestine and preparation of the lower urinary tract for kidney transplantation]. PMID- 10853203 TI - [Renal arteriography during heart catheterization: 2 explorations with the price of one]. PMID- 10853204 TI - Early blood transfusion equipment. PMID- 10853205 TI - An investigation of the early effects of manual lung hyperinflation in critically ill patients. AB - This prospective within-group multicentre study was designed to assess the safety and short-term effectiveness of manual lung hyperinflation in mechanically ventilated patients. Eighteen patients from the intensive care units of two tertiary institutions were included and acted as their own control. Manual lung hyperinflation treatment involved patient positioning (side-lying), suctioning and manual lung hyperinflation. Side-lying treatment involved patient positioning and suctioning alone. Patients received both treatments on the day of data collection. Results demonstrated significant improvement for static respiratory system compliance (P = 0.001) with manual lung hyperinflation treatment compared to side-lying treatment. Manual lung hyperinflation treatment also cleared a significantly greater wet weight of sputum (P = 0.039). There were no differences between manual lung hyperinflation and side-lying treatment for gas exchange (PaO2/FIO2 and PaCO2), mean arterial pressure or heart rate. In conclusion, total static respiratory system compliance and sputum clearance were improved by the addition of manual hyperinflation to a physiotherapy treatment of positioning and suctioning in mechanically ventilated patients without compromise to cardiovascular stability or gas exchange. PMID- 10853206 TI - Subhypnotic dose of propofol for the prevention of nausea and vomiting during spinal anaesthesia for caesarean section. AB - This study was undertaken to evaluate the efficacy of subhypnotic doses of propofol for the prevention of nausea and vomiting in parturients undergoing caesarean section under spinal anaesthesia. In a randomized, double-blinded, placebo-controlled manner, 60 patients received intravenously lignocaine 0.1 mg/kg (for injection pain relief) followed by either placebo (Intralipid) or propofol at subhypnotic dose (1.0 mg/kg/h) (n = 30 of each) immediately after clamping of the fetal umbilical cord. Emetic episodes and safety assessments were performed during spinal anesthesia for caesarean section. The incidence of patients who were emesis-free in the intraoperative, post-delivery period was 37% with placebo and 77% with propofol, respectively (P = 0.001). No clinically important adverse events were observed in either group. In conclusion, a subhypnotic dose (1.0 mg/kg/h) of propofol is effective for preventing nausea and vomiting in parturients undergoing caesarean section under spinal anaesthesia. PMID- 10853207 TI - Granisetron/dexamethasone combination for the prevention of postoperative nausea and vomiting after thyroidectomy. AB - This study was undertaken to evaluate the efficacy of granisetron plus dexamethasone for preventing postoperative nausea and vomiting (PONV) after thyroidectomy. In a prospective, randomized, double-blind study, 130 female patients received either granisetron 40 micrograms/kg or granisetron 40 micrograms/kg plus dexamethasone 8 mg intravenously immediately before the induction of anaesthesia (n = 65 in each group). A standard general anaesthetic technique was used. A complete response (no PONV, no rescue) during the first three hours after anaesthesia occurred in 85% of patients with granisetron alone and 98% with granisetron plus dexamethasone; the corresponding incidence for the period 3 to 24 hours after anaesthesia was 86% and 98% (P < 0.05; Fisher's exact probability test). No clinically serious adverse events were observed in any of the groups. In conclusion, prophylactic use of granisetron/dexamethasone combination is more effective than granisetron alone for preventing PONV in women undergoing thyroidectomy. PMID- 10853208 TI - The effect of nitrous oxide on hearing. AB - There is a belief that the administration of nitrous oxide (N2O) increases hearing acuity. This increase can be interpreted as hearing low intensity sounds more loudly. This study examined auditory threshold levels, acoustic impedance, acoustic perception, memory, and discrimination to determine if hearing was altered by end-tidal 10% or 20% nitrous oxide. Subjects also qualitatively interpreted three intensity levels using a subjective intensity test. Observations were made in the breathing mediums of room air in the control stages (Stages 1 and 4), compared to N2O and supplemental oxygen (Stages 2 and 3), in 16 human subjects in a quiet room. Breathing end-tidal 10% and 20% N2O significantly increased middle ear pressure, and produced a significant effect on the subjective interpretation of a sound's intensity. There was no significant effect on auditory threshold at a range of frequencies. It appears that 10% or 20% N2O inhalation does not lead to the commonly held view of increased hearing acuity as measured in terms of auditory threshold. Rather, at these levels of N2O inhalation, subjects experience a state similar to a pre-sleep stage, whereby the hearing diminishes but remains active for loud intensity sounds. PMID- 10853209 TI - Measurement of quality of recovery in 5672 patients after anaesthesia and surgery. AB - Quality of recovery after an operation is an important dimension of the patient's experience and may be related to the quality of anaesthesia care. Satisfaction with anaesthesia is a vital component of quality care but difficult to measure. We examined our database of 5672 adult patients to determine if quality of recovery is associated with satisfaction with anaesthesia and to identify the perioperative factors that might influence both these outcome measures. We found that a nine-item quality of recovery score ("QoR Score") was related to satisfaction with anaesthesia (P < 0.0005): the overall level of satisfaction was high (97.2%; median QoR Score 16); 106 patients (2.1%; median QoR Score 14) were "somewhat dissatisfied" and 32 patients (0.6%; median QoR Score 13) were "dissatisfied" with their anaesthesia care. Patients who experienced any of a number of perioperative complications had lower QoR Scores (P < 0.0005). We have further demonstrated the validity and clinical utility of the QoR Score, and in particular, its relationship to patient satisfaction in adult surgical patients. PMID- 10853210 TI - Tracheal intubation with the Macintosh laryngoscope versus intubating laryngeal mask airway in adults with normal airways. AB - We tested the hypothesis that haemodynamic changes to intubation and postoperative pharyngolaryngeal morbidity are similar for blind intubating laryngeal mask (ILM)-guided compared with laryngoscope-guided tracheal intubation in adults with normal airways. We also compared intubation success rates and airway complications. One-hundred and fifty paralysed, anaesthetized adult patients undergoing elective surgery were randomly assigned to one of three equal sized groups: 1. blind intubation via the ILM using a straight, silicone tube; 2. intubation with a Macintosh laryngoscope using a straight silicone tube and 3. intubation with a Macintosh laryngoscope using a polyvinyl chloride tube (controls). A standard sequence of adjusting manoeuvres was followed if intubation was difficult. The number of adjusting manoeuvres and intubation attempts, time to intubation, intubation success rate (first attempt and within 3 min), haemodynamic changes (pre-induction, post-induction, post-intubation), oesophageal intubation, mucosal trauma (blood detected), hypoxia (SpO2 < 95%) and postoperative pharyngolaryngeal morbidity (double-blinded) were documented. Time to successful intubation was longer (57 vs 35 s), and more intubation attempts were required in the ILM group (P < 0.0001). The intubation success rate was 100% (all first attempt) for the laryngoscope groups and 94% (56% first attempt) for the ILM group. There were no significant differences in heart rate or blood pressure among groups. Oesophageal intubation (26 v 0%) and mucosal trauma (19 v 2%) were more common in the ILM group. Hypoxia and postoperative pharyngolaryngeal morbidity were similar among groups. Blind intubation through the ILM offers no advantages over the Macintosh laryngoscope for adult patients requiring intubation for elective surgery with normal airways, but it is a feasible alternative. PMID- 10853211 TI - The effect of PVC packaging on the acidity of 0.9% saline. AB - Intravenous fluids in polyvinyl chloride (PVC) packaging are known to be acidic. We proposed to determine the effect of PVC packaging on the pH of 0.9% saline solutions by comparing the predicted and measured pH of 0.9% saline equilibrated with atmospheric carbon dioxide and the measured pH of commercial solutions of 0.9% saline in PVC and polypropylene packaging. Calculation of pH was made from available physical chemistry constants and data. Measurement was made of the pH of 12 samples of prepared 0.9% saline equilibrated with atmospheric carbon dioxide. Comparison with the pH of seven commercial samples of saline in PVC packaging for intravenous use was undertaken. Further comparison was made between commercial samples of 0.9% saline in PVC or polypropylene packaging. The calculated pH of 0.9% saline was 5.61 at 20 degrees C. The median pH of the prepared samples was statistically significantly less acidic than the median pH of the PVC packaged samples for intravenous use: 5.47 vs 4.60, P < 0.05. The median pH of the PVC packaged saline was also statistically significantly more acidic than the pH of the polypropylene packaged saline: 4.62 vs 5.71, P < 0.05. The acidity of the intravenous solutions of 0.9% saline packaged in PVC was much greater than expected and is only partially explained by dissolved carbon dioxide. This acidity could be a result of packaging in PVC. PMID- 10853212 TI - The development of a performance indicator to objectively monitor the quality of care provided by an acute pain team. AB - Quality assurance procedures are essential in the maintenance of clinical standards in medicine. Conventional analysis techniques have difficulty in detecting gradual changes over time. Cumulative sum techniques monitor the frequency with which an event occurs and can detect changes in its frequency as soon as they become statistically significant. This study explores the use of cumulative sum techniques to monitor the performance of an acute pain team in a teaching hospital. It shows that periods of suboptimal performance can be readily identified. The prospective use of these techniques in clinical audit may allow the earlier identification and correction of technical or organisational problems. These should lead to improvements in patient care and satisfaction. PMID- 10853213 TI - Anaesthesia and fatigue: an analysis of the first 10 years of the Australian Incident Monitoring Study 1987-1997. AB - The Australian Incident Monitoring Study (AIMS) database of the Australian Patient Safety Foundation (APSF) was reviewed from its inception in April 1987 to October 1997. A total of 5600 AIMS reports were lodged in that period. Reports in which fatigue was listed as a Factor Contributing to Incident were examined. This occurred in 152 reports, or 2.7% of all reports. Confidence interval analysis suggested that fatigue was associated with various concurrently reported factors. These included pharmacological incidents (especially syringe swaps) and time of day. Other factors significantly associated with fatigue reports were haste, distraction, inattention and failure to check equipment. Relieving anaesthetists and healthy patients were reported more often as factors minimizing incidents. Anaesthetists reporting fatigue more often reported incidents during induction. These data suggest that fatigue alleviation strategies and equipment checking routines, improved workplace design (including drug ampoule and syringe labelling protocols) and regulation of working hours will facilitate minimization of fatigue-related incidents. Definitive prospective studies might be most usefully targeted at these and related interventions. PMID- 10853214 TI - Acyclovir induced coma in the intensive care unit. AB - A 73-year-old man with multiorgan failure requiring mechanical ventilation and haemodialysis developed herpes labialis infection during his stay in the ICU. This was treated with enteral acyclovir. He developed persistent neurologic impairment soon after acyclovir administration, which, over the course of seven days, progressed to coma, the aetiology of which was unclear. The computed tomograph (CT) of the brain and the cerebrospinal fluid (CSF) examination was normal. The electroencephalogram (EEG) showed generalized slowing. The possibility of acyclovir neurotoxicity was considered and the drug was discontinued. Haemodialysis was instituted and the patient made a complete neurological recovery. We believe that this is the first reported case of coma due to enteral acyclovir. PMID- 10853215 TI - Postoperative confusion preceded by decreased frontal lobe haemoglobin oxygen saturation. AB - We describe a 58-year-old male patient with confusion and prolonged recovery after liver transplantation. A cause was not apparent for the confusion, but during surgery, monitoring of the frontal lobe cerebral haemoglobin oxygen saturation by near-infrared spectrophotometry showed cerebral hypo-oxygenation despite optimization of conventional cardiovascular parameters. It is possible that intraoperative cerebral ischaemia is the cause of postsurgical confusion and with near-infrared spectrophotometry this hypothesis may be tested clinically. PMID- 10853216 TI - Pethidine reverses morphine-induced delirium. AB - A young patient had unexpected and prolonged postoperative delirium apparently associated with morphine-induced biliary colic. Naloxone had no therapeutic effect, but a small dose of pethidine produced a dramatic return to lucidity. Unrecognized biliary spasm should be considered as a cause of agitation in the recovery room in postoperative patients who have received morphine. PMID- 10853217 TI - Vasopressin effective in reversing catecholamine-resistant vasodilatory shock. AB - A patient with perforated appendicitis developed progressive vasodilatory shock which was complicated by perioperative acute myocardial infarction. Cardiovascular support included dopamine infusion, and later, intra-aortic balloon counterpulsation balloon pump and noradrenaline and dobutamine infusion. Vasopressin was introduced as a final attempt to reverse the refractory shock and was associated with recovery. The experience with this case suggests that vasopressin may be a valuable adjunct to the treatment of catecholamine-resistant vasodilatory shock. PMID- 10853218 TI - Target-controlled intravenous anaesthesia with bispectral index monitoring for thoracotomy in a patient with severely impaired left ventricular function. AB - The anaesthetic management of an elderly patient with severely impaired left ventricular function undergoing thoracotomy and lobectomy is described. Total intravenous anaesthesia (TIVA) with remifentanil and target-controlled infusion of propofol titrated according to the bispectral index (BIS) was used, with thoracic epidural anaesthesia commenced at the end of surgery providing postoperative analgesia. Avoidance of intraoperative epidural local anaesthetics and careful titration and dose reduction of propofol using the BIS was associated with excellent haemodynamic stability. The rapid offset of action of remifentanil and low-dose propofol facilitated early recovery and tracheal extubation. The BIS was a valuable monitor in optimal titration of TIVA. PMID- 10853219 TI - Acute heart failure in the parturient--do not forget phaeochromocytoma. AB - Phaeochromocytoma is a rare condition and extremely rare in pregnancy. Diagnosis is notoriously difficult, as phaeochromocytoma may present a broad spectrum of clinical manifestations. The key to a successful outcome is a high index of suspicion of its existence and its early diagnosis. PMID- 10853220 TI - Oesophageal rupture in a patient with postoperative nausea and vomiting. AB - Rupture of the oesophagus (Boerhaave's syndrome) is a rare complication of forceful or suppressed vomiting. Postoperative nausea and vomiting is common but does not usually lead to life-threatening complications. A case of oesophageal rupture in a man who experienced postoperative nausea and vomiting after an uncomplicated procedure is described in this report. Delayed diagnosis mandated conservative treatment. The clinical presentation, diagnosis and management of oesophageal rupture is discussed. PMID- 10853221 TI - Mortality from peripartum meningitis. AB - A young primigravid parturient had an uneventful labour under epidural analgesia and delivered a healthy male infant. She returned 48 hours later with fever, vomiting and severe headache, but was misdiagnosed as having endometritis. Further signs of meningitis appeared six hours later, however she succumbed to the infection and died four weeks later despite intensive care and high-dose antibiotic management. Causes of meningitis in the peripartum period are discussed. The possibility of a causal association between the patient's epidural analgesia and her infection are considered and preventive measures discussed. PMID- 10853222 TI - Re-wiring an arterial line. A novel technique. AB - In cases where the loss of a small volume of blood is critical to the patient's welfare, the technique of re-wiring intra-arterial and intravenous catheters that minimize the amount of blood lost is important. A simple technique is described using the needleless systems which are now widely employed throughout hospitals. PMID- 10853223 TI - Bain circuit adapter fault. PMID- 10853224 TI - Leaking catheter mount swivels. PMID- 10853225 TI - Diathermy burn to a visitor in the operating theatre. PMID- 10853226 TI - BIPP aspiration. PMID- 10853227 TI - Ephedrine may predispose to arrhythmias in obstetric anaesthesia. PMID- 10853228 TI - Reducing the risk of drug errors. PMID- 10853229 TI - Phantom pain and regional anaesthesia. PMID- 10853230 TI - Rickettsial diseases and their serological diagnosis. AB - Rickettsial diseases (typhus and spotted fever group rickettsioses, scrub typhus and Q fever) may pose a serious public health problem, namely when they are non diagnosed or misdiagnosed. Although rickettsiae can be isolated from or detected in clinical specimens, serological tests still remain an indispensable tool in the diagnosis of rickettsial diseases. The complement fixation test widely used in the past is being replaced by other tests which make differentiation of immunoglobulin classes possible. Of these tests microimmunofluorescence is considered the test of choice followed by the latex agglutination, indirect hemagglutination, immunoperoxidase assay, and enzyme-linked immunosorbent assay. The last one is also suitable for seroepidemiological studies. Immunoblot analysis can be used to confirm the results of other tests. The use of the low specific and low-sensitive Weil-Felix test should be reserved only for situations in which other serologic tests are not available. PMID- 10853231 TI - Prevalence of hepatitis G in patients on chronic hemodialysis. AB - Hepatitis G virus (HGV) is a newly described RNA virus from the family of flaviviridae. It is closely related to the hepatitis C Virus (HCV) but is more common than HCV among healthy blood donors. The pathogenicity of HGV in immunosuppressed patients such as those undergoing hemodialysis is unclear. We measured the incidence of HGV in 105 patients undergoing hemodialysis in a chronic outpatient hemodialysis facility. HGV-RNA was detected using a RT-PCR method with primers directed against the 5' non-coding region and the NS5a gene of HGV. Nine (8.6%) patients were HGV RNA positive, eleven (10.5%) were anti-HCV positive, three (2.9%) were positive for hepatitis B surface antigen. Four patients were positive for both HGV and HCV; three of them had normal liver enzymes while one showed elevated ALT levels but no other signs of exacerbation of preexisting hepatitis. The prevalence of HGV among dialysis patients is comparable to that of HCV. The transmission route for HCV is nosocomial transmission during dialysis, whereas HGV shows both ways of transmission: blood transfusion mediated by a high prevalence of HGV among healthy blood donors and nosocomial transmission. HGV appears to play a minor role in acute hepatitis, even in immunosuppressed patients. PMID- 10853232 TI - Hepatitis C virus and oral lichen planus/lichenoid reactions: lack of evidence for an association. AB - Some studies have suggested an association between the mucocutaneous disorder lichen planus and chronic infection with hepatitis C virus. Most of these studies have been based purely on serological markers. The present study sought to detect hepatitis C virus RNA in both peripheral blood and in biopsy material collected from oral mucosal lesions. Twenty-seven patients were studied, six with classical lichen planus and 21 with oral lichenoid reactions. The diagnoses were confirmed by histopathological examination. Reverse transcription PCR was employed to detect hepatitis C virus RNA in the blood specimens. The same method was used to detect hepatitis C virus RNA in lesional tissue, following RNA extraction from sections of the biopsies. The virus was not detected in any of the paired blood and tissue specimens examined. It is concluded that hepatitis C virus is not commonly associated with oral lichen planus or lichenoid reactions in Scotland. PMID- 10853233 TI - A closer look at the insertion within helix 54 of the 23S-rRNA of the genus Corynebacterium (and related taxa). AB - Direct sequencing of the insertion in helix 54 of the 23S-rRNA of different strains of 8 species of the genus Corynebacterium was performed in order to determine the extent of variations of these strains. The results demonstrated that there is considerable variation within different strains of a single species. While this clearly jeopardizes the usefulness of the insertion with regard to species-specific diagnostic probes, the variations were found to concentrate within two clearly defined regions comprising the ascending and descending parts of the distal helix of a putative stemlike secondary structure. The remainder of the insertion sequences seem to be fairly constant within a single species while displaying considerable differences toward sequences of other species of the same genus. PMID- 10853234 TI - Influence of urea on HbA1c-determinations by Menarini HA-8140 and on the difference between immunoturbidimetric and HPLC-HbA1c-results. AB - A secondary peak (#C) included in the HbA1c-calculation by the HA-8140 HPLC (Menarini) shows a fairly good correlation with serum urea. The correlation with HbA1c and with serum glucose is at first glance significant but reveals at a closer look being biased by some incorrect assumptions. The difference between immunoturbidimetric determination (Tinaquant HbA1c II Roche) and HPLC shows a similar behaviour to urea as does the #C-peak of the chromatographic separation. This peak as well as the difference between both determinations of HbA1c could be attributed to carbamylated haemoglobin. The definitive identity has to be proven. This peak could be a monitoring tool for long-time urea. The integration of this peak into total HbA1c by the HA-8140 (Menarini) can lead to a false diagnosis of diabetes in non-diabetic patients with elevated urea. PMID- 10853235 TI - Community acquired diarrhea--the incidence of Astrovirus infections in Germany. AB - Astroviruses are increasingly recognized as a cause of human gastroenteritis. Electron microscopy (EM) has been considered the "gold standard" method for diagnosis, but this approach is limited to a few laboratories. We evaluated a commercial enzyme immunoassay (EIA) (IDEIA Astrovirus, DAKO Diagnostika, Hamburg, Germany) for the direct detection of antigen in fecal samples. In comparison to EM, the assay scored 100% in sensitivity and specificity (n = 213; 26 positive samples) and reacted with strains representing all known serotypes. Over an 11 month period 4,211 stool samples from unselected German patients suffering from acute gastroenteritis were examined. Etiologically responsible microorganisms were found in 13.0% of cases, with astrovirus the third most common pathogen (1.2%) behind Salmonella spp. (2.9%) and Rotavirus (2.5%), representing 13.5% of all positive specimens. Norwalk-like viruses (NLV), fungi, and protozoa were not tested. In infants of < 2 years of age (n = 458) the incidence of astrovirus infection was significantly higher (2.8%) compared to children of 2-7 years of age (n = 578; 1.7%) and those of > 7 years of age (n = 3,175; 0.9%). The frequency revealed a peak in winter (mean November-February: 2.0% versus other months: 0.8%). PMID- 10853236 TI - Comparison of immunoassays for the detection of anti-GAD65 autoantibodies in patients with diabetes mellitus. AB - Recent studies have demonstrated that a combination of GAD-antibody assays and IA 2 autoantibody assays show a high diagnostic specificity for Type 1 diabetes. For this reason there is increasing interest in the use of GAD-antibody measurement for Type 1 risk assessment. Since a number of different assays have been published and documented in the literature, the aim of this study was to evaluate four different anti-GAD test systems that are commercially available in Germany. We tested the anti-GAD prevalences in five patient groups with the different immunoassays and compared them with the values obtained by an immunoprecipitation test (IP-Test). All assays correlated well with the IP-test and showed high sensitivity and specificity in the group of patients with recent onset Type 1 diabetes and the control group. The groups tested consisted of 20 subjects with recent onset Type 1 diabetes (< 6 weeks) (sensitivity 70-90%), nine subjects with a Type 1 duration of more than 2 years (sensitivity 11-33%), 21 patients with pluriglandular insufficiency (sensitivity 28.5-47.5%), 10 patients with Type 2 (specificity: 90-100%), and 14 healthy control subjects (specificity: 93-100%). Our data show a high level of sensitivity and specificity of the tested, commercially available, assays. Since almost every laboratory should be able to establish one of these assays, this may facilitate the possibility of further large scale population studies with the aim of investigating GAD-antibody prevalences in screening for Type 1 diabetes. Increased measurement of the diabetes-associated antibodies will be helpful in the differential diagnosis of gestational diabetes mellitus (GDM) and latent autoimmune diabetes of the adult (LADA). PMID- 10853237 TI - Chemiluminescence immunometric assay for measuring osteocalcin in healthy children. AB - Osteocalcin (OC) was measured in serum samples from 92 children and adolescents (57 females, 35 males) by a two-site chemiluminescence immunometric assay (Nichols Institute Diagnostics, USA), which recognizes the 1-19 region of the whole molecule as well as the large N-terminal midregion fragment representing the main part of OC in serum. The highest OC levels are measured between the age of 10-15 years. The linear correlations between OC and alkaline phosphatase were 0.65 (p < .01) for total alkaline phosphatase (TAP) and 0.61 (p < 0.1) for bone alkaline phosphatase (BAP). PMID- 10853238 TI - Bone resorption marker in pre- and postmenopausal females. PMID- 10853239 TI - Effect of 6-hour exposure to 20 degrees C on the ATP content and other biochemical measures of CPDA-1 packed red cells. AB - Blood donations for clinical use are routinely stored at 2 degrees C to 6 degrees C for 35 to 42 days. It is common practice for RBCs exposed to temperatures above 10 degrees C to be destroyed, although the American Association of Blood Banks Technical Manual states "Blood exposed to temperatures above 10 degrees C is not necessarily unsuitable for transfusion". To clarify this issue we investigated the effect of 6-hour storage at 20 degrees C on the content of ATP and other biochemical measures of CPDA-1 packed red cells. CPDA-1 packed RBC units were exposed once at day 5 (group 1), day 15 (group 2) or day 30 (group 3) of their shelf life to 20 degrees C for 6 hours. Control groups were continuously refrigerated. Under all conditions of storage, the ATP concentrations decreased with time. Initial ATP levels of five-day old CPDA-1 packed RBCs were 3.94 mumol/g Hb in the test group and 3.73 mumol/g Hb in the control group. At day 30 after warming (day 35 of the shelf-life) the ATP concentrations declined to 2.78 mumol/g Hb (test group) and to 3.55 mumol/gHb (controls). In the test series which were warmed at day 15 and day 30 of shelf-life the ATP levels declined to 3.16 mumol ATP/g Hb and 2.92 mumol ATP/g Hb at day 35 of shelf-life. There was no significant difference between test and control group with respect to the lactate levels, whole-blood glucose, sodium and potassium. The percentage of hemolysis was lower than 0.5% under all conditions of storage. Our data show that a shorter period of moderate warming (6h, 20 degrees C) does not lead to a critical decline of ATP and glucose concentrations in CPDA-1 packed RBCs. The survival of RBCs stored in CPDA-1 is most highly correlated with maintaining ATP concentrations above a value of about 2 mumol per g of Hb [3]. The ATP levels in our study were well above this threshold. PMID- 10853240 TI - Elecsys parathyroid hormone (PTH) not detecting the large PTH fragment hPTH (7 84)? PMID- 10853241 TI - Additional comments on pre-analytical aspects of lipoprotein measurement. PMID- 10853242 TI - [The role of leukocytic factors in the preparation for and induction of labor]. AB - Preparation for labour and its induction are controlled by an integrative but not single process. The primary stimulus that comprises this integrative hormonal prostaglandin-leukocytic mechanism is a suppression of local progesterone action with further inhibition of perivascular prostaglandine dehydrogenase and sharp increase of E2 and F2 prostaglandine content in uteroplacentary mechanism bed. Vasodilatory effect of prostaglandines along with synergication of interleukine 8 leads to leukocyte emigration and formation of infiltrates consisting of neutrophilic granulocytes and certain number of macrophages and mast cells. Hydrolytic enzymes released by neutrophilic granulocytes cause degradation of collagenic tissue in uterine cervix promoting its ripening. The factors mentioned also influence loosening of choriodecidual interface. Further amplification of effect of prostaglandine including those produced by neutrophilic granulocytes along with the other uteroconstrictors increase myometrium contractions which leads to labour induction. PMID- 10853243 TI - [The developmental characteristics of speech-motor areas 44 and 45 in the left and right hemispheres of the human brain in early postnatal development]. AB - Cytoarchitecture of speech areas 44 and 45 of human brain was studied on frontal series of sections of brain of a newborn, 6 months, 1 year and 2 years old child. Sections were stained with cresyl violet after Nissl. Density of neurons and gliocytes location as well as fraction of satellite gliocytes and surrounded by the latter, glia index were determined by quantitative methods. General principles of cortical areas 44 and 45 were distinguished as well as peculiarities of their formation if left and right hemispheres of human brain in postnatal ontogenesis. Heterochronia of postnatal development of areas 44 and 45 was demonstrated. A hypothesis on different activity of right and left hemispheres speech areas in different periods of speech development of the child was discussed. PMID- 10853244 TI - [The individual characteristics of the cytoarchitectonics of the human motor cortex (based on computer analysis data)]. AB - Age and individual peculiarities of cytoarchitecture of layer III field 4p, periods of intensive and slow growth of the area of profile fields (APF) of pyramid neurons were established by method of computer analysis of histological preparations of human motor cortex from birth up to 20 yrs. It was shown that the greatest changes of APF of pyramid neurons including their individual parameters occur in field 4 of motor cortex during first three years. PMID- 10853245 TI - [The neuronal-glial correlations in areas of the frontal area of the brain in children at different stages of life]. AB - Density of location of all the pyramid neurons (N), satellite glia (SG) and neurons surrounded by it (NS) in brain cortex fields 8 and 47 was studied by cytoarchitectonic and qualitative methods using MBI-3 microscope, Continuous growth of number of NS, SG and NS/N, the important parameters of neuronal activity, was established. This process is irregular. It lowers and intensifies at 2 "crucial" periods of ontogenesis, where difference in neuron activity between fields is most distinct. In puberty these difference are more manifested: the above mentioned parameters are sharply increased in field 8 and decrease in field 47. PMID- 10853246 TI - [A morphological analysis of the cluster organization of the cortico-cortical neurons in area 17 of the cat visual cortex]. AB - Spatial organisation and quantitative features of cortico-cortical neurons in area 17 that project to area 21a of cat visual cortex were investigated. We used the HRP technique and two-dimensional reconstruction from serial sections of brain. Pattern of distribution of labelled cells in some regions of areas 17 and 18, on the lateral and medial surface of the brain were analysed. The data about cluster organisation of cortico-cortical neurones, localisation, layer distribution and density of neurons were obtained. PMID- 10853247 TI - [A comparative analysis of the descending projections of areas 5 and 7 of the parietal cortex to the cat brain stem]. AB - Relative quantity of descending fibres of parietal fields 5 and 7 of cat brain stem was studied. Maximal projections of these field were determined, maximal projections of fields 5 and 7 to red nucleus reticular nucleus of midbrain and nucleus proprius of pons were shown. Topographic and quantitative overlapping of projections of fields 5 and 7 in red nucleus and reticular nucleus of midbrain was noted. Field 5 is maximally presented in ventral nucleus while field 7 has maximal number of projections in lateral nucleus. Projection of fields 5 an 7 to superior colliculus of tectum of the midbrain was determined that was significant although less pronounced than in the above mentioned nuclei. The number of fibres projecting onto the superior colliculus of tectum of the midbrain (its caudal part) from field 7 exceeds that from field 5. There are less fibres in inferior colliculus of tectum of the midbrain than in superior colliculus. The number of fibres projecting from field 5 is greater than those from field 7 in inferior colliculus. In caudal reticular nucleus of pons the presence of fields 5 and 7 was not significant. Comparison of the authors' data with that available in literature allows to suggest the possible inhibitory, i.e. modulating influence of parietal cortex on nuclei of the pons. PMID- 10853248 TI - [The neuronal organization of the periamygdaloid cortex in the rat brain]. AB - Cytoarchitectonic peculiarities and neuronal organization of periamygdaloid cortex (PAC) of rat brain was studied at three levels: rostral and caudal regions of central part and in posterior part of amygdala using Nissl and Golgi methods. Neuronal composition characteristic for each level and interrelations between neurons of different layers were determined. Different levels of PAC vary in architecture of neurons. Periamygdaloid cortex at the rostral level of central region is characterized with more simple organization. PMID- 10853249 TI - [The structural organization of the normal rat area postrema and under conditions of chronic exposure to gravitational loads]. AB - Topography and structure of rat area postrema were specified using light optic and electron microscope. Disturbance of liquor-encephalic transport (hydrops of ependymocytes), sharp changes of cytoplasmic organelles in neurons, signs of intercellular desintergration (synapse degeneration after "clear" type), death of part of neurons, disorder in secretory processes in neuroendocrine cells were shown to occur following chronic effect of gravitation overloads, in its structural elements. These changes take place on the background of hemomicrocirculatory disturbances, appearance of connective tissue fibres in perivascular space of Virchow-Robin and are probably conditioned by a developing circulatory hypoxia. PMID- 10853250 TI - [The descending cortical pathways of the frontal lobe to the nuclei of the hypothalamic mamillary bodies in human craniocerebral trauma]. AB - Structural organization of conducting ways of the frontal lobe cortex to hypothalamic nuclei was studied in sections of brain from 5 patients who died in short terms after craniocerebral trauma and during the experiment in 5 macaque rhesus monkeys with unilateral destruction of different cortical fields of frontal area. Series of brain sections were processed using silver nitrate impregnation after Bielschowsky [correction of Bilshovsky] with counterstaining of nuclear structures after Kavamura with cresyl violet. The presence of direct corticohypothalamic ways from the cortex of the orbital surface of inferior (field 47 and its subfields) and superior (field 11) frontal gyri and frontal pole (field 10) to nuclei of mamillary complex and lateral hypothalamus was established. Direct frontomamillary ways from these cortical areas terminate basically in medial mamillary nucleus. In monkey direct frontomamillary ways were distinguished in the cortex of field 47. In man widening of the area where corticohypothalamic ways arise is connected with progressive development of the telencephalic cortex and phylogenetically new formation of hypothalamus- mamillary bodies. PMID- 10853251 TI - [The prenatal development of the tectum mesencephali and substantia nigra in man]. AB - The development of tectum of the midbrain was studied in situ in human 6-11 wks embryos. Using electron microscopy and immunocytochemistry processes that occur in the anlage of tectum of the midbrain at early stages of formation of dopaminergic populations of cells were followed up. Growth of pallium zone due to migration of cells from ventricular zone and their differentiation is proportional to growth of the terms of embryonal development. Proliferating cells were located not only in ventricular zone. Expression of tyrosine hydroxylase was found in 6.5 wks embryos in the fraction of cells that migrated from ventricular zone and was observed only during neuroblast migration. Cells migrate along the radial glia fibres in dorsoventral direction. Microgliocytes with short processes near to vessels located in the vicinity of ventricular zone were found in 7 wks embryos for the first time. Peculiarities of morphogenetic processes participating in anlage of tectum of the midbrain and possible role of microgliocytes in development of embryonal nerve tissue are discussed. PMID- 10853252 TI - [The structure of the developing blood vessels of the neocortical anlage of the human embryo]. AB - Using light and electron microscopy the structure of blood vessels of neocortical anlage of human 7-12 embryos was studied. It was shown that at the early stage of formation of intraorgan vascular network the wall of blood vessels of ventricular zone successively differentiate, which is characterized by the appearance of second layer of cells (pericytes), accumulation of basement membrane components, widening of the zone of contacts between endotheliocytes and establishment of the contacts with bipolar cells of neocortex anlage. The morphological data obtained assist in comprehension of physiological aspects of formation of blood brain barrier and regulation of blood flow in human embryonal neocortex. PMID- 10853253 TI - [The structure of the optic nerve in experimental glaucoma and in the means for its treatment]. AB - Using light and electron microscopy disorders of structure of the optic nerve were studied in rabbits with adrenaline induced glaucoma and after the action of antioxidants (Emoxipine, Erisode) used for preventive and medical purposes. Changes in glaucoma and after administration of drugs in axons and their myelinated sheath were similar but differently pronounced in right and left eye of the same animal. Positive effect of Erisode used for preventive and medical purposes was noted. Emoxipine enhanced the disturbances in optic nerve structure. PMID- 10853254 TI - [The 3-dimensional organization of the sensory epithelium of the cochlea in birds]. AB - Using authors' original approach to studying histoarchitecture of cellular layers three-dimensional structure of bird helix sensory epithelium was studied theoretically and experimentally. The composition of its elementary morphofunctional unit, AB3 (one sensory cell per three supporting cells) was established. Three dimensional model of this unit was worked out that described shape and arrangement of cells at apical, basal and intermediate levels of the layer. The presence of its translation symmetry was demonstrated. Comparison of theoretical model and results of morphological research of sensory epithelium allowed to conclude that the model corresponds with the real tissue. The model assists in three-dimensional reconstruction of sensory epithelium. And allows to deal with minimum number of sections and also gives an opportunity to forecast its developmental changes. PMID- 10853255 TI - [The ultrastructural changes to the ovaries and oviducts in experimental inflammation under the influence of eplir phonophoresis]. AB - Ultrastructural changes of blood vessels of ovaries and oviducts, destruction of the part of follicular epitheliocytes and disruption of other structures of blood follicular barrier occur at the early period of experimental inflammations of appendage uterus on the background of traditional antibiotic therapy. All changes lead to atresia of growing follicules. Fibrotic-sclerotic changes develop on the 30th day and during later period. Course of eplir phonophoresis decreases ultrastructural disruptions of vessels, follicular apparatus of ovaries and oviducts epithelium. Effect of physiotherapy is manifested though activity of fibroclasts and macrophages, that regulate collagen production/resorption ratio thus preventing the development of fibrotic changes of ovary stroma and oviduct wall. PMID- 10853256 TI - [The structural manifestations of the adaptive mechanisms occurring in the villous placenta during gestosis]. AB - Using morphometry, optic and structural analysis and immune histochemistry structural manifestations of adaptive mechanisms in placenta of hemo- and desmochorial type during gestation were studied. Signs of similarity and diversity of realization of adaptive mechanisms at tissue and cellular level were found. Reduction in placental barrier thickness and number of surface capillaries and also activation of trophoblastic elements genome relate to similar signs. Formation of terminal villi is a divergent adaptive mechanism in hemochorial placenta, while intensification of migration of main functionally active cells from trophoblast lining to mother tissue is those in desmochorial placenta. PMID- 10853257 TI - [The importance of the supporting and duplicating parameters of the midface for determining the dimensions of the external nose]. PMID- 10853258 TI - [Asymmetry of the parameters is the basis of the structure of the space-time organization of functions]. PMID- 10853259 TI - [The development and heterogeneity of mast cells]. PMID- 10853260 TI - [A comparative analysis of different designs of test tasks during their use in a department of histology of a medical college]. PMID- 10853261 TI - [The creative legacy of Prof. A. A. Maksimov (from the Chicago period of his scientific activities)]. PMID- 10853262 TI - [The Russian Academy of Medical Sciences on the brink of the 21st century]. PMID- 10853263 TI - [The resolution of the doctoral and candidate dissertations in specialty 14.00.02, Human Anatomy, defended and approved by the Higher Certification Commission of the Russian Federation in 1998]. PMID- 10853264 TI - Expression of murine adenosine deaminase (ADA) in transgenic maize. AB - A murine adenosine deaminase (ADA) gene, driven by the maize ubi-1 promoter and intron region, was transformed into embryogenic maize callus, along with a bar and gusA gene-containing plasmid, using microparticle bombardment. Selection in the presence of either the herbicide Basta or the adenosine analogue 2' deoxyadenosine resulted in transgenic cultures that expressed GUS and accumulated a 41-kD protein that immunoprecipated with an ADA-specific polyclonal antibody. ADA enzyme activity was observed in extracts from transgenic callus as well as regenerated plants and progeny. Culltures expressing ADA grew in the presence of 200 mg/l 2'-deoxyadenosine, a concentration which completely inhibited the growth of non-transgenic cultures. ADA activity appeared to segregate in progency of regenerated plants as a single, dominant Mendelian trait. These results suggest that ADA, in combination with adenosine analogue selection, represents a potentially viable selectable marker system for transgenic maize production. PMID- 10853265 TI - Linear transgene constructs lacking vector backbone sequences generate low-copy number transgenic plants with simple integration patterns. AB - Whole plasmids are used in both Agrobacterium-mediated transformation and direct DNA transfer, generally leading to the integration of vector backbone sequences into the host genome along with the transgene(s). This is undesirable, as vector backbone sequences often have negative effects on transgene or endogenous gene expression, and can promote transgene rearrangements. We, therefore, bombarded rice tissue with two constructs: a plasmid containing the bar gene, and a linear DNA fragment isolated from the same plasmid, corresponding to the minimal bar gene expression cassette (promoter, open reading frame and terminator). We recovered phosphinothricin-resistant plants from both experiments, showing that the selectable marker was efficiently expressed. Transformation with such constructs resulted in predominantly 'simple' integration events (one or two bands on Southern blots), producing low-copy-number transgenic plants with a low frequency of transgene rearrangements. Conversely, transformation with supercoiled or linearized whole plasmids generated plants with 'complex' integration patterns, that is, higher copy numbers and frequent transgene rearrangements. We monitored transgenic lines through to the R4 generation and observed no silencing in plants carrying minimal constructs. We also carried out experiments in which rice tissue was simultaneously bombarded with minimal linear hpt and gusA cassettes. We observed robust GUS activity in hygromycin-resistant plants, confirming co-expression of the selectable and nonselectable markers. Furthermore, the efficiency of cotransformation using minimal constructs was the same as that using supercoiled plasmid cointegrate vectors. PMID- 10853266 TI - Transformation of barley by microinjection into isolated zygote protoplasts. AB - Barley zygote protoplasts were mechanically isolated, embedded in agarose droplets, and microinjected with a rice actin promoter Act1-gusA-nos gene construct. On average 62% of the cells survived the injection and of these 55% continued development into embryo-like structures and eventually to plants. PCR screening for the presence of a 307-bp fragment in the middle of the gusA gene showed that on average 21% of the derived structures contained this fragment. However, among the hundreds of injected zygotes, derived structures and regenerants we only found significant GUS expression in two cases (embryo-like structures nine days after injection). Two lines of green plants, derived from zygotes microinjected with linearized plasmid (line A147-1) or an isolated Act1 gusA-nos gene cassette (line A166-h) proved to be transgenic. Line A147-1 appeared to contain a single and intact copy of the expression cassette but a PCR based progeny analysis indicated the presence of additional shorter fragments of the cassette. Line A166-h appeared to contain a single fragment of the gusA gene that was transferred to the progeny as a single Mendelian trait. One additional fragment of the gusA gene was identified in this line. The present data show that transformation of barley by microinjection of DNA into isolated zygotes is feasible but also that gene expression rarely is achieved, possibly due to degradation of the introduced DNA. PMID- 10853267 TI - Promoter strength influences polyamine metabolism and morphogenic capacity in transgenic rice tissues expressing the oat adc cDNA constitutively. AB - We analyzed molecularly and biochemically a series of transgenic rice lines expressing the oat adc (arginine decarboxylase) cDNA under the control of the constitutive maize ubiquitin 1 promoter. We established baseline biochemical parameters to elucidate the role of polyamines (PAs) during morphogenesis. We measured mRNA levels, ADC enzyme activity and cellular PAs in dedifferentiated callus. Polyamine levels were also quantified in two subsequent developmental stages--regenerating tissue and differentiated shoots. We observed significant (P < 0.05) differences in the levels of individual PAs at the three developmental stages. The amounts of putrescine (Put) and spermidine (Spd) in dedifferentiated transgenic callus were lower than those in the wild type or in hpt (hygromycin resistant)-controls, whereas the amount of spermine (Spm) was increased up to two fold. In regenerating tissue, this trend was reversed, with significantly higher levels of Put and Spd (P < 0.05), and lower levels of Spm (P < 0.05) compared to non-transformed or hpt-control tissues at the same developmental stage. In differentiated shoots, there was a general increase in PA levels, with significant increases in Put, Spd, and Spm (P < 0.05); on occasion reaching six times the level observed in wild type and hpt-control tissues. These results contrast those we reported previously using the weaker CaMV 35S promoter driving adc expression. mRNA measurements and ADC enzyme activity were consistently higher (P < 0.01) in all tissues expressing pUbiadcs compared to equivalent tissues engineered with 35Sadc. Our findings are consistent with a threshold model which postulates that high adc expression leading to production of Put above a basal level is necessary to generate a big enough metabolic pool to trigger PA flux through the pathway leading to an increase in the concentration of Spd and Spm. This can be best accomplished by a strong constitutive promoter driving adc. We discuss our results in the context of flux through the PA pathway and its impact on morphogenesis. PMID- 10853268 TI - Expression of Acidothermus cellulolyticus endoglucanase E1 in transgenic tobacco: biochemical characteristics and physiological effects. AB - The expression of the Acidothermus cellulolyticus endoglucanase E1 gene in transgenic tobacco (Nicotiana tabacum) was examined in this study, where E1 coding sequence was transcribed under the control of a leaf specific Rubisco small subunit promoter (tomato RbcS-3C). Targeting the E1 protein to the chloroplast was established using a chloroplast transit peptide of Rubisco small subunit protein (tomato RbcS-2A) and confirmed by immunocytochemistry. The E1 produced in transgenic tobacco plants was found to be biologically active, and to accumulate in leaves at levels of up to 1.35% of total soluble protein. Optimum temperature and pH for E1 enzyme activity in leaf extracts were 81 degrees C and 5.25, respectively. E1 activity remained constant on a gram fresh leaf weight basis, but dramatically increased on a total leaf soluble protein basis as leaves aged, or when leaf discs were dehydrated. E1 protein in old leaves, or after 5 h dehydration, was partially degraded although E1 activity remained constant. Transgenic plants exhibited normal growth and developmental characteristics with photosynthetic rates similar to those of untransformed SR1 tobacco plants. Results from these biochemical and physiological analyses suggest that the chloroplast is a suitable cellular compartment for accumulation of the hydrolytic E1 enzyme. PMID- 10853269 TI - Enhancing the efficiency of introducing precise mutations into the mouse genome by hit and run gene targeting. AB - The creation of precise clinical mutations by targeting is important in elucidating disease pathogenesis using mouse models. 'Hit and run' gene targeting is an elegant method to achieve this goal. This uses first a positive selection to introduce the targeting vector carrying the required mutation and then a negative selection to identify clones which have removed vector and wild-type sequences by intrachromosomal recombination. However, this approach has only been successfully used in a handful of cases. We used this procedure to introduce precise clinical mutations into the exon 10 region of the cystic fibrosis transmembrane conductance regulator (Cftr) gene. Using a CMV promoter driven hygromycin/thymidine kinase (hyg/tk) fusion gene as both our dominant and negative selectable marker, we targeted the Cftr locus very efficiently but only identified false runs after the negative selection step. This defect in thymidine kinase induced toxicity to gancyclovir correlated with methylation of the transgene. Consequently we devised a stringent screening procedure to select only true 'run' clones. Unfortunately these 'run' clones had lost the mutation so we altered the vector design to bias the run step to retain the mutation and used a different tk selection cassette with a HSVtk promoter sequence. This new vector design allowed both efficient 'hit and run' for two cystic fibrosis (CF) mutations with no false positives and successful germline transmission of the novel G480C missense mutation. PMID- 10853270 TI - Use of a cryptic splice donor site in the chloramphenicol acetyltransferase (CAT) SV40 small-t antigen cassette generates alternative transcripts in transgenic rats. AB - The bacterial gene chloramphenicol acetyltransferase (CAT) is a widely used reporter in both in-vitro and in-vivo studies of genetic regulation. We have recently generated novel rat transgenic lines carrying an arylalkylamine N acetyltransferase (AA-NAT) promoter-reporter construct in which CAT (with associated SV40 small-t antigen sequence) is the reporter. In addition to the predicted transgene transcript (1.9 kb), we identified an abundant 1.5 kb transcript which derives from an alternative splicing event that utilises a cryptic splice donor site located within the CAT gene. The native CAT open reading frame (ORF) is lost in the 1.5 kb transcript, and a western analysis has shown that protein deriving from an aberrant open reading frame is not expressed at detectable levels. PMID- 10853271 TI - Expression of full-length bioactive antimicrobial human lactoferrin in potato plants. AB - A cDNA fragment encoding human lactoferrin (hLF) linked to a plant microsomal retention signal peptide (SEK-DEL) was stably integrated into the Solanum tuberosum genome by Agrobacterium tumefaciens-mediated leaf disk transformation methods. The lactoferrin gene was expressed under control of both the auxin inducible manopine synthase (mas) P2 promoter and the cauliflower mosaic virus (CaMV) 35S tandem promoter. The presence of the hLF cDNA in the genome of regenerated transformed potato plants was detected by polymerase chain reaction amplification methods. Full-length hLF protein was identified by immunoblot analysis in tuber tissue extracts from the transformed plants by immunoblot analysis. The hLF produced in transgenic plant tissues migrated during polyacrylamide gel electrophoresis as a single band with an approximate molecular mass equal to hLF. Auxin activation of the mas P2 promoter increased lactoferrin expression levels in transformed tuber and leaf tissues to approximately 0.1% of total soluble plant protein. Antimicrobial activity against four different human pathogenic bacterial strains was detected in extracts of lactoferrin-containing potato tuber tissues. This is the first report of synthesis of full length, biologically active hLF in edible plants. PMID- 10853272 TI - Preoperative evaluation and management of the emergency surgical small animal patient. AB - The initial management and stabilization of the emergency surgical patient is challenging. A consistent systematic evaluation of the four major body systems, determination of the underlying cause, and aggressive stabilization before surgery can optimize the outcome. PMID- 10853273 TI - Emergency vascular access. AB - Establishing and maintaining vascular access is often vital to the preoperative, intraoperative, and postoperative management of the small animal emergency surgical patient. Vascular access may be used for the delivery of crystalloids, colloids, blood components, medications, and anesthetic or sedative agents. It can also facilitate venous and arterial blood sampling and allow direct measurement of arterial and venous pressures. The small animal emergency and critical care veterinarian should have a thorough knowledge of vascular access techniques, including peripheral and central venous catheterization, intraosseous, and arterial access. Competence in percutaneous, percutaneous facilitative, and surgical cutdown approaches should ensure that vascular access can always be established in the critically ill patient. PMID- 10853274 TI - Anesthesia for the emergency small animal patient. AB - Anesthesia for the debilitated or emergency patient requires a thorough knowledge of the pharmacologic and physiologic effects of the anesthetic agents available. Preoperative evaluation and preparation of the patient, intraoperative monitoring and prompt attention to potential crises, and postoperative monitoring and pain management are all critical to the success of dealing with these cases. Special attention is devoted to cases with cardiovascular, respiratory, or neurologic instability. PMID- 10853275 TI - Surgical emergencies of the respiratory system. AB - Management of the patient in respiratory distress requires an efficient and accurate diagnostic and therapeutic strategy. This article describes the approach to patients with respiratory compromise, including the indications and techniques for performing emergency surgical procedures. The clinical features of upper airway obstruction, thoracic wall trauma, and pleural space disease are discussed. PMID- 10853276 TI - Gastrointestinal emergencies. AB - The animal with a surgical gastrointestinal emergency usually requires a rapid, thorough physical examination with concurrent resuscitation. As the diagnosis is being made, the animal must be made as stable as possible before undergoing general anesthesia. During surgery, there must be a critical evaluation of gastrointestinal viability and the use of precise technical skills to achieve the best outcome. Adept postoperative management, including careful monitoring and an index of suspicion for potential complications, is vital. PMID- 10853277 TI - Surgical emergencies of the urinary tract. AB - True emergencies of the urinary tract center on three major issues, including uncontrolled renal hemorrhage, accumulation of urine within the peritoneal cavity or retroperitoneal space, and obstruction to urine outflow. Successful management of urinary tract emergencies in small animal patients is based not only on the severity of the injury or obstruction but on the condition of the patient at the time of diagnosis and the patient's response to medical stabilization. When most urinary tract emergencies are initially recognized, patients are metabolically and hemodynamically unstable. Therefore, urinary tract emergencies are first regarded as medical emergencies, and emergency surgical procedures are aimed at patient stabilization and lifesaving measures. PMID- 10853278 TI - Diagnosis and management of peritonitis. AB - Peritonitis is a clinical syndrome rather than a definitive diagnosis and, as such, the underlying cause must be identified. Many potential causes exist, and peritonitis may be classified as primary or secondary, local or diffuse, acute or chronic, or according to the causative agent. Peritonitis is one of the most common initiators of the systemic inflammatory response syndrome faced by the small animal practitioner. Because systemic inflammatory response syndrome is such a devastating disease process, aggressive stabilization, rapid definitive diagnosis, and prompt surgical and medical treatment are vital for optimizing outcome. PMID- 10853279 TI - Neurosurgical emergencies. AB - The neurologic patient is considered a neurosurgical emergency when delay of treatment may influence the patient's outcome. Diseases of the spinal cord, brain, and peripheral nerves are presented in this article. Diagnostic tools (i.e., advanced imaging and electrophysiologic tests), differential diagnoses, treatment options (conventional and controversial), whether the patient requires surgery, and the optimal time for surgical intervention are discussed. PMID- 10853280 TI - Emergency management of open fractures and luxations. AB - This article discusses the emergency management of open fractures and traumatic joint luxations. The emergency management of open fractures can be challenging. The proper initial evaluation and treatment of open fractures can have a positive impact on their successful outcome. The clinical signs and diagnosis of traumatic joint luxations are reviewed. Emphasis is placed on the management of these injuries in the emergency situation. PMID- 10853281 TI - A practical approach to hemoperitoneum in the dog and cat. AB - Hemoperitoneum can occur in animals of any age and can result from many different disease processes. Neoplastic and traumatic conditions are the most common causes. Many of these patients present with hemodynamic instability but stabilize with rational intravenous fluid therapy and abdominal counterpressure. Surgical exploration of the abdomen is indicated in many situations. Surgical therapy is aimed at resection or control of the bleeding focus, removal of any devitalized tissue, and biopsy of additional sites of suspicion. Optimal treatment for all patients with hemoperitoneum frequently requires advanced critical care, anesthesia, and surgical techniques. Finally, the treatment outcome is variable and dependent on the underlying cause and its severity. PMID- 10853282 TI - Bite wounds in dogs and cats. AB - Bite wounds are a frequent injury in dogs and cats. Some bites are severe, causing crushing, avulsion, and devitalization of tissues beneath the skin. Bites can also crush the airway or penetrate the abdominal or thoracic cavities, resulting in life-threatening injuries. This article reviews the local mechanisms of trauma and systemic mediators involved in severe bite wound injuries and provides a plan for stabilization, injury assessment, and definitive bite would management. PMID- 10853283 TI - Postoperative management of the emergency surgery small animal patient. AB - Hypovolemia, hypothermia, and hypotension are common postoperative findings that predispose the critically ill patient to secondary complications. This patient population is especially vulnerable to sepsis, hypoxia, and immune dysfunction. Careful monitoring is essential for early recognition of potentially life threatening physiologic derangements. Early and aggressive intervention may help minimize systemic insult before it progresses to acute respiratory distress syndrome, acute renal failure, disseminated intravascular coagulation, or multiple organ failure. PMID- 10853284 TI - Using compartmentalized inhalation physiologically-based pharmacokinetic modeling to calculate occupational and environmental risks: a case study involving toluene. PMID- 10853285 TI - Controlling cleaning-solvent vapors at small printers. National Institute for Occupational Safety and Health. PMID- 10853286 TI - Virtual reality for mine safety training. AB - Mining has long remained one of America's most hazardous occupations. Researchers believe that by developing realistic, affordable VR training software, miners will be able to receive accurate training in hazard recognition and avoidance. In addition, the VR software will allow miners to follow mine evacuation routes and safe procedures without exposing themselves to danger. This VR software may ultimately be tailored to provide training in other industries, such as the construction, agricultural, and petroleum industries. PMID- 10853287 TI - Comparison of sampling methods for monomer and polyisocyanates of 1,6 hexamethylene diisocyanate during spray finishing operations. AB - A comparison study of isocyanate sampling methods for 1,6-hexamethylene diisocyanate (HDI) monomer and HDI-based polyisocyanates was conducted in spray painting environments. This study compared the performance of the Iso-chek sampler against existing and proposed National Institute of Occupational Safety and Health (NIOSH) and Occupational Safety and Health Administration (OSHA) monitoring methods for HDI-based isocyanates. Six methods for monitoring HDI monomer and polyisocyanate levels were compared. Fifty-eight sampling sets were collected during spray painting of aircraft and aircraft parts at four U.S. Air Force bases. Impinger and cassette samplers were mounted side-by-side on a mannequin located in paint overspray areas. For HDI monomer sampling results, there were no significant differences between NIOSH 5521, NIOSH 5522, OSHA 42, MAP (the proposed NIOSH method), and the Iso-Chek. For HDI-based polyisocyanates, NIOSH 5522, NIOSH 5521, Iso-Chek, and the Total Aerosol Mass Method (TAMM) were significantly different from one another. There was no significant difference between MAP and the NIOSH 5522 polyisocyanate sampling results. This study suggests the Iso-Chek and MAP sampling methods compare favorably with established methods for monitoring in HDI spray painting environments and the Total Aerosol Mass Method provides a reasonable upper boundary for estimating HDI polyisocyanate concentrations. The results also reemphasize aerosol sampling physics and sampler geometries must be carefully considered and appropriate samplers used when measuring exposures in spray paint environments where particulates are of the inhalable size. PMID- 10853288 TI - Capability of respirator wearers to detect aerosolized qualitative fit test agents (sweetener and Bitrex) with known fixed leaks. AB - This study was designed to evaluate and compare the ability of respirator wearers to detect qualitative respirator fit test agents (saccharin and Bitrex) when the respirators were modified to include fixed size leaks. In recent years the number of persons who require fit testing has increased, partly in response to the needs of health care workers with potential exposure to infectious bio-aerosols. Many health care providers have chosen qualitative respirator fit testing using saccharin and/or Bitrex for a variety of reasons, including (but not limited to) low initial equipment cost. Respirators were modified to include a mid-line sampling probe between the nose and mouth for quantitative fit testing with a TSI PortaCount. A second modification included the introduction of a shortened 14 gauge intravenous catheter at the bridge of the nose. The fixed leak was designed to produce fit factors < 100 when unplugged, with an average fit factor of 67 among 26 respirator wearers. A complete fit test was not performed, because one purpose of this study was to determine the ability of respirator wearers to detect a known fixed leak during a single normal breathing exercise, without introducing unknown and potentially variable size leaks. Sensitivity threshold screening included a placebo and requirement to correctly characterize the taste of the agent used. Quantitative fit factors without leaks ranged from 96 to > 20,000 and 22 to 160 with the leak present. Twenty four of 26 subjects had fit factors < 100 (92%) when fixed leaks were induced. All subjects correctly detected Bitrex with fixed leaks (sensitivity = 100%). Nine of 26 subjects (35%) were unable to detect saccharin in the presence of a known fixed leak even though the average fit factor for these subjects was 77. When the two subjects with fit factors > 100 were excluded, only 16 of 24 respirator wearers were able to detect saccharin with fixed leaks (sensitivity = 67%). There were several important aspects of our study design worth noting, including the introduction of a placebo during sensitivity threshold testing, limiting the subject response time to a single maneuver, using a higher concentration of Bitrex than commercially available, and requiring the subjects to correctly characterize the taste of the qualitative test agent. In conclusion, leak detection was correctly identified with Bitrex, but not saccharin. PMID- 10853289 TI - Exposures to jet fuel and benzene during aircraft fuel tank repair in the U.S. Air Force. AB - Jet fuel and benzene vapor exposures were measured during aircraft fuel tank entry and repair at twelve U.S. Air Force bases. Breathing zone samples were collected on the fuel workers who performed the repair. In addition, instantaneous samples were taken at various points during the procedures with SUMMA canisters and subsequent analysis by mass spectrometry. The highest eight hour time-weighted average (TWA) fuel exposure found was 1304 mg/m3; the highest 15-minute short-term exposure was 10,295 mg/m3. The results indicate workers who repair fuel tanks containing explosion suppression foam have a significantly higher exposure to jet fuel as compared to workers who repair tanks without foam (p < 0.001). It is assumed these elevations result from the tendency for fuel, absorbed by the foam, to volatilize during the foam removal process. Fuel tanks that allow flow-through ventilation during repair resulted in lower exposures compared to those tanks that have only one access port and, as a result, cannot be ventilated efficiently. The instantaneous sampling results confirm that benzene exposures occur during fuel tank repair; levels up to 49.1 mg/m3 were found inside the tanks during the repairs. As with jet fuel, these elevated benzene concentrations were more likely to occur in foamed tanks. The high temperatures associated with fuel tank repair, along with the requirement to wear vapor-permeable cotton coveralls for fire reasons, could result in an increase in the benzene body burden of tank entrants. PMID- 10853290 TI - Characterization of metalworking fluid exposure indices for a study of acute respiratory effects. AB - Although metalworking fluids have been widely used throughout industry for decades, occupational exposures to metalworking fluid aerosols and their constituents have not been well characterized. This article describes an exposure assessment for a study of metalworking fluid aerosols and acute respiratory effects. This exposure assessment was unique in its inclusion of multiple exposure measures relevant to a complex environment, and extensive personal sampling for bacteria and endotoxin. The specific objectives were to: (1) obtain indices of personal exposure to metalworking fluid aerosols in an automotive transmission plant, either directly (by sampling) or indirectly (by estimation), and (2) identify and adjust for sources of error in exposure/dose measures, where possible. No prior studies have characterized personal exposures to metalworking fluid aerosols so extensively. Exposure data were obtained during a pilot phase and three principal rounds of data collection over a 15-month period in conjunction with spirometric testing. Subjects worked in one of two machining departments, Case and Valve Body, or in a comparison department, Final Assembly. The primary exposure measures for this study were thoracic fraction particulate, thoracic fraction bacteria (viable plus non-viable), and total endotoxin. Mean personal air concentrations of thoracic particulate across all study rounds were 0.13 mg/m3 in Final Assembly, 0.32 mg/m3 in Valve Body, and 0.56 mg/m3 in Case. Average personal exposures to thoracic fraction bacteria were 0.38 bacteria/cc in Final Assembly, 0.87 bacteria/cc in Valve Body, and 2.66 bacteria/cc in Case. Average personal endotoxin measurements, collected in Round 3 of the study, were 16.4 endotoxin units (EU)/m3 in Assembly, 34.7 EU/m3 in Valve Body, and 234 EU/m3 in Case. Sump fluid contained on the order of 10(8) bacteria/ml, and 10(4)-10(5) EU/ml. Air concentrations of thoracic particulate, thoracic bacteria, and total endotoxin were highly correlated in metalworking operations. Thus, reducing airborne particulate levels should also reduce ambient bacteria and endotoxin, which are suspect agents of respiratory impairment. The elevated endotoxin levels in Final Assembly were unexpected, and suggest an independent source of endotoxin contamination in this department. PMID- 10853291 TI - Optimized portable cordless vacuum method for sampling dry, hard surfaces for dusts. AB - Surface sampling in industrial/environmental hygiene is a growing field that needs validated standardized methods. There are few standard methods, one being the American Society for Testing and Materials (ASTM) method involving a portable, cordless air sampling pump. In the present work, the ASTM technique was modified to increase efficiency and versatility. A soil sample was first dried and sieved. Known weights of different sieved sizes (125 microns-180 microns, 90 microns-125 microns, and 63 microns-90 microns) were then sampled at an average flow rate of 4.0 +/- 0.2 L/min from a template of inner dimensions 10 cm by 10 cm on two different surfaces (rough and smooth). Five consecutive sampling passes were performed. For the smooth surface, the first pass efficiency for the largest particles were 45% +/- 45% (CV = 100%), and 75% +/- 20% (CV = 27%) for the smallest particles. After three passes, the efficiency independent of particle size exceeded 83 percent with a CV better than 11 percent. After five passes, the efficiency exceeded at least 85 percent with about the same precision as for three passes. The rough surface allowed higher efficiencies for the first two sampling passes. Three to five passes are recommended to achieve acceptable efficiencies for the surface loose dust/soil range 100 micrograms/cm2 to 1,500 micrograms/cm2. PMID- 10853292 TI - The determination of the ability of polyurethane foam to release toluene diisocyanate into air. AB - A study was conducted to assess the ability of polyurethane foam to release toluene diisocyanate (TDI) into the air. The study was completed in two phases. In the first phase, three-day post-production foam samples were "extracted" using 37 degrees C, 30 percent relative humidity air for a total of three days. In the second phase, foam samples were "loaded" with TDI (approximately 1 ppm (w/w) in the foam) by passing air containing a controlled level of TDI vapor through the foam. The "loaded" foam was then aged for three days (to simulate minimum possible expected time between production and consumer contact) before being extracted for three days using 37 degrees C, 30 percent relative humidity air. In both phases of the study, the extracted TDI was quantified by trapping it from the air using glass-fiber filters coated with 1,(2-pyridyl)piperazine (PP) derivatizing agent (modified OSHA Method 42) then analyzing the TDI derivative formed using high-performance liquid chromatography with ultraviolet detection. Validation of the test system was conducted using diffusion-tube-generated TDI atmospheres. Results of the Phase 1 portion of the study showed no quantifiable TDI being extracted from the three-day post-production commercial foam samples at a limit of quantitation of approximately 0.1 ppb TDI in air. Results of the Phase 2 portion of the study showed no quantifiable TDI being extracted from the "loaded" foams at a limit of quantitation of 0.12 microgram TDI (less than 0.03% of the level of TDI loaded into the foams). In conclusion, the results from this study indicate that it is not likely that TDI would be released from three-day post-production polyurethane foams in amounts likely to produce air concentrations of concern. PMID- 10853293 TI - [Mechanism of itch transmission]. PMID- 10853294 TI - [Useful indices for asthma management]. PMID- 10853295 TI - [Quantification of dust mite allergens in indoor environment and its application]. PMID- 10853296 TI - [Long-term prognosis of asthmatic patients treated with low-dose beclomethasone dipropionate]. AB - To clarify the prognosis of asthmatics treated with low-dose of inhaled beclomethasone dipropionate (BDP), we retrospectively assessed 43 patients treated with initial dose of 200 or 400 micrograms/day for 5 years, and obtained the following results. 1) 15 patients achieved step-down therapy (group A), 17 patients maintained initial dose of BDP (group B), and 11 patients required step up therapy of BDP or daily use of oral prednisolone (group C). 2) There was no significant difference in age, sex, duration of disease, severity of disease, peripheral eosinophil counts, %FEV1 and histamine PC20 before BDP treatment among three groups. The percentage of atopic asthmatics was significantly higher in group C than in group A. 3) There was no significant difference in symptom and histamine PC20 between after 1 year state and after 5 years state in three groups. 4) After 1 year from the start of BDP treatment, only 18% patients got symptom free and neither patients exceeded 20,000 micrograms/ml of histamine PC20 in group C. Long-term treatment of low-dose BDP inhalation was effective on mild/moderate asthmatics. Patients requiring step-up therapy had not got sufficient improvement in bronchial hyperresponsiveness after one-year treatment. PMID- 10853297 TI - [L-selectin expressed on peripheral monocytes and soluble L-selectin in sera in atopic diseases]. AB - For the purpose of assessing the involvement of L-selectin in atopic diseases, we measured L-selectin expressed on peripheral monocytes and serum soluble L selectin (sL-selectin). We enrolled 7 patients with atopic bronchial asthma(BA), 6 patients with atopic dermatitis, 12 patients with Japanese cedar pollinosis(P), and 9 healthy controls(C). L-selectin expressed on peripheral monocytes was detected and analyzed by using flow cytometry. Serum sL-selectin was measured by enzyme-linked immunosorbent assay. Compared with the group C (81.7 on average), mean fluorescence intensity of CD 62 L on monocytes in the group BA was significantly higher (110.4 on average, p < 0.05). The group AD also showed higher tendency (101.3 on average, p < 0.08). No significant difference was found in serum sL-selectin levels among the four groups(BA: 1086 ng/mL, AD: 1226 ng/mL, P: 945 ng/mL, C: 1052 ng/mL, on average). There was a negative correlation (r = 0.69) between serum sL-selectin and age in all subjects (BA + AD + P + C). The results suggest that L-selectin is involved in the extravasation of monocytes toward local inflammatory sites in atopic diseases. Serum sL-selectin seems to stay at normal level in mild and stable atopic diseases, while the trend of high serum sL-selectin level in children may participate in the pathogenesis. PMID- 10853298 TI - [Measurement of formaldehyde-specific IgE antibodies in adult asthmatics]. AB - Formaldehyde (FA) is known to be a potent contact sensitizer eliciting dermatitis or bronchial asthma in occupational settings. However, several pieces of recent evidence indicate that FA is a participant of indoor air pollution even at home and that it may be one of the causative agents for the sick-house syndrome. In the present study, we examined whether production of FA-specific IgE antibody (FA IgE) occurred in adult chronic asthmatics or not. We randomly selected 80 adult asthmatics, and obtained blood sampling under the condition where the symptoms were well controlled. We measured FA-IgE, eosinophil count and serum concentration of eosinophil cationic protein (sECP), and also measured lung functions (FEV1%, %VC, V25/HT). We found that FA-IgE was positive in two patients (33-year-old female and 56-year-old male), while negative (less than 0.35 UA/ml) in the others. State of asthma was severe in the female patient, while mild intermittent in the male patient. In the female patient, the total IgE titer and the FA-IgE titer were 181 IU/ml and 0.81 UA/ml, respectively, and there was no occupational environments concerning FA. In the male patient, both the total IgE and the FA-IgE titer were quite high (8400 IU/ml and 2.99 UA/ml, respectively). In this case, there was an occupational environment where the adhesive smell was strong. sECP was 44.2 micrograms/l in the female patient, while 4.7 micrograms/l in the male patient. In the present study, IgE-mediated allergy for FA was rare in adult chronic asthmatics, and the FA-IgE titer did not seem parallel to severity or pathophysiological condition in bronchial asthma. But, further development of precise methodology for clinically clarifying FA-mediated mechanisms in allergic disorders, might be still important. PMID- 10853299 TI - [Chlamydia pneumoniae infection in patients with acute bronchitis and bronchial asthma]. AB - In this study, a total of 60 patients with acute bronchitis, 71 patients with bronchial asthma and 20 healthy volunteers were serologically and bacteriologically analyzed to investigate whether Chlamydia pneumoniae infection is associated with the onset and the exacerbation with acute bronchitis and bronchial asthma. Antibody titers to Chlamydia pneumoniae were also measured and compared by ELISA method. The antibody-positive rate in the patients with acute bronchitis (88.4%) was significantly higher than that in the patients with bronchial asthma (73.3%) or that in the healthy volunteers (60%). And the levels of the IgA antibody in the patients with acute bronchitis were significantly higher than those in the patients with bronchial asthma or those in the healthy volunteers. The rate of acute C. pneumoniae infection in the patients with acute bronchitis (20%) did not show significantly differences compared with that in the patients with bronchial asthma (15.5%) or that in the healthy volunteers (10%). The cases of acute C. pneumoniae infection had both as a single etiologic agent and as a mixed infection, most often with Streptococcus pneumoniae. Therefore, we demonstrated that the acute C. pneumoniae infection may be associated with the onset and the exacerbation in acute bronchitis and bronchial asthma. PMID- 10853300 TI - [Study of how to continue peak flow monitoring for patients with bronchial asthma: questionnaire to continuation and interruption patients]. AB - Since 1993, our hospital has actively pursuing introduction of peak flow monitoring for patients with bronchial asthma. However, a number of patients are found to discontinue PEF monitoring. In order to determine the causes, a comparative analysis of recognition between patients who continued PEF monitoring (continuation group, hereafter) and those who discontinued PEF monitoring (interruption group, hereafter) was conducted through a questionnaire survey. The number of patients with bronchial asthma who responded to the questionnaire was 120; among them, the number of cases who continued and interrupted PEF monitoring were 68 and 52, respectively. The percentage of patients whose symptoms improved after the introduction of PEF monitoring was higher in the continuation group than in the interruption group. Approximately half of the patients in the interruption group commented that PEF monitoring was valid. The percentage of patients who believed that PEF monitoring is valid was higher in the group of patients who experienced reduction in the severity of symptom after introduction of PEF monitoring compared with those who did not. From these results, to keep high compliance of PEF monitoring, we propose that introduction of PEF monitoring and education of self-management should be positively stated for patients in unstable state. Furthermore, we found that some of the patients in the interruption group expressed interest in restarting PEF monitoring, indicating that each patient should be dealt with on a case by case basis. Appeals from doctors, patients education and advice of relatively few measurements, were found to be especially important in motivating patients to use PEF monitoring. PMID- 10853301 TI - [30 cases of occupational latex allergy]. AB - We experienced 30 patients with occupational latex allergy (LA) in a 4-year period between 1995 and 1998 in Fujita Health University Hospital. All of them were medical personnel. We studied clinical symptoms and the clinical relevance of latex specific IgE and skin test (prick test and use test) in 30 cases. As a result, skin test result was most related of the diagnosis and the severity of LA. The average of the duration to decide LA diagnosis was 21.2 months. The reasons of the delayed diagnosis were that they were left and treated as an unexplained urticaria or asthma. Therefore, we have to warn about LA to the medical personnel who are frequently exposed to latex in Japan. PMID- 10853302 TI - Topical anesthesia without intracameral lidocaine in cataract surgery. AB - PURPOSE: To evaluate the efficacy of topical anesthesia with oxybuprocaine 0.4% without intracameral lidocaine as an alternative to peribulbar or retrobulbar anesthesia in cataract surgery. METHODS: Fifty-eight patients (eighty-two eyes) were included in this study. All patients received topical anesthesia with oxybuprocaine 0.4%. No intracameral lidocaine was used at the start of the intervention. Seventy-five per cent of patients received oral sedation with lorazepam 2.5 mg. All surgery was done using a superior corneal incision and phacoemulsification followed by a foldable IOL implantation. Subjective pain was assessed at 4 intervals during surgery using a 4-point pain scale. All patients were evaluated for intraoperative eye motility and blepharospasm. Patient and surgeon satisfaction was measured with a 4-point satisfaction scale. RESULTS: 15% of patients experienced mild pain during phaco and 43% had mild pain during corneal suturing. No patient had severe pain during the operation. In 4% of patients, intracameral lidocaine was used to relieve pain. The surgeon and patient satisfaction was high. No eye movements or blepharospasm were recorded in 75% and 62% of cases respectively. No serious complications occurred. CONCLUSION: Topical anesthesia is a safe and effective alternative to peribulbar and retrobulbar anesthesia in corneal cataract surgery for the experienced surgeon. PMID- 10853303 TI - The sensitivity and specificity of TOP, FDP and GDX in screening for early glaucoma. AB - The purpose of this study was to evaluate the efficacy of three screening tests in detecting glaucoma in its early stage: the Tendency Oriented Perimetry (TOP) and Frequency Doubling Perimetry (FDP) visual field tests, and the Glaucoma Diagnostic (GDx) nerve fibre layer analyser. Eighteen patients with glaucoma who showed an early defect on HFA c 24-2 and twenty normals underwent the three tests. TOP showed a sensitivity of 94.4% and a specificity of 75%, FDP showed a sensitivity of 72.2% and a specificity of 100%, and GDx a sensitivity of 77.7% and a specificity of 60%. PMID- 10853304 TI - Ocular findings in children with homozygous sickle cell disease in the Democratic Republic of Congo. AB - OBJECTIVE: The purpose of the study is to determine the frequency of ocular manifestations in Congolese children with homozygous sickle cell disease. METHODS: Sixty-six children with homozygous sickle cell disease were examined for ocular abnormalities between March 1 and August 31, 1998. The ages of children ranged from two to 18 years. Routine ophthalmic examination included measurement of visual acuity, inspection of the adnexa and cornea, refraction, silt-lamp examination and dilated ophthalmoscopy. RESULTS: Ocular abnormalities were found in 47 (71%) children of 66 subjects. Ophthalmologic abnormalities included conjunctival signs (32%), retinal vascular tortuosity (29%) and dilatation (26%), AV crossing (29%). CONCLUSIONS: Ocular findings in this study were similar to those previously published in Africa, which showed a low frequency of retinal changes. PMID- 10853305 TI - Outer orbital distance, inner canthal distance and interpupillary distance, proptosis in children with homozygous sickle cell disease. AB - BACKGROUND: On basis of clinical observation paediatricians in Kinshasa had the impression that children with homozygous sickle cell disease have a special face characterised by hypertelorism. OBJECTIVE: The purpose of the study is to determine outer orbital, inner canthal and interpupillary distances as well as proptosis in children with sickle cell disease. METHODS: These measurements were performed on 66 Congolese children with homozygous sickle cell disease, aged from 2 to 18 years. The measurements were performed with the Hertel exophthalmometer for the proptosis and the outer orbital distance, with the pupillometer model PD 2 meter for the interpupillary distance and with a ruler for the inner canthal distance. The results were compared with those of 95 healthy children of similar age. RESULTS: All measurements were age related. In every age group the values for inner canthal distance were identical to those of healthy children, but the interpupillary, the outer orbital distances and the proptosis were significantly smaller. CONCLUSIONS: A slow growth of orbital tissue in children with homozygous sickle cell was suggested to explain the difference with healthy children. Other biometric studies comparing the orbital measurements with the rest of the body are needed to confirm or refute this hypothesis. PMID- 10853306 TI - [Solar retinopathy acquired after gazing at the sun during prayers]. AB - The authors present the results of a study based on six persons who developed retinopathy after gazing at the sun during prayers. Risk factors were studied and the prognosis evaluated. The mean follow-up time was 82 days. Risk factors were present in all patients. The visual acuity was reduced in all patients at first presentation and total improvement was noted in four of them. Of all six patients, only one had his ocular fundus normalized. Amsler's grid testing revealed in all patients bilateral central scotomas which persisted in four of them. Fluorescein retinal angiography, which was found to be abnormal in all cases, became normal in only one case. However, all patients continued to complain of visual disability because of the persistence of photophobia, meta morphopsia and scotomas. In conclusion, the prognosis of solar retinopathy is variable and the recovery of visual acuity does not necessarily imply the improvement of vision. Appropriate eye protection such as solar filters must be used when gazing at the sun to protect from retinal damage. PMID- 10853307 TI - [Report of a case of Acanthamoeba keratitis]. AB - CLINICAL REPORT: A clinical report of a contact lenses wearer with Acanthamoeba keratitis pointed out the diagnosis problem. The medical treatment is needed previously to any surgery. Finally the patient underwent enucleation. DISCUSSION: The authors are considering the microbiological aspects and laboratory techniques are described. CONCLUSION: For this very severe but hopefully rare pathology, the sooner the treatment the best. A therapeutic approach is described. PMID- 10853308 TI - [Surgery for macular hole without positioning]. AB - Although the macular hole surgery with gas tamponade obtains good results, the face-down positioning for minimum 5 days remains constraining and is often a cause of refusal or abstention concerning the operation. We describe 7 patients who underwent the macular hole surgery by pars plana vitrectomy with internal limiting membrane peeling and fluid-silicone exchange without position restrictions. The silicone oil has been removed 2 to 3 months later. All the holes have been closed except in one case where the internal limiting membrane had not been peeled with certainty. A second surgery with peeling of this membrane has permitted the hole closure. We don't report any complications induced by the silicone oil. Silicone oil tamponade for macular hole surgery with internal limiting membrane peeling shows a good efficiency without particular positioning and without additional complications. PMID- 10853309 TI - [Refractive power evolution following cataract surgery]. AB - A retrospective study was carried out to evaluate and quantify the refractive power evolution following cataract procedure. Three different intraocular lenses were compared: Corneal ACR6D (acrylic), Kelios 600 (silicone), Storz HM60 (hydrogel). Visual acuity and post-operative refraction were compared at one week and after two months. The spherical equivalent change after those follow-ups shows the refractive evolution. Refraction is stable with time for the Storz and Kelios lenses. However, post-operative hyperopia was found for monobloc Corneal implant. PMID- 10853310 TI - Optic nerve sheath meningiomas: clinical features, functional prognosis and controversial treatment. AB - Optic nerve sheath meningiomas (ONSM) are rare benign neoplastic lesions arising from meningothelial cells of the meninges. As clinical features are highly variable, the diagnosis is often delayed. From 1995 to 1999, 6 patients were diagnosed with ONSM in our department. We compared our series with the literature data. Visual prognosis is usually poor. Despite a large literature, the treatment guidelines are still highly controversial. PMID- 10853311 TI - OCT study of fellow eyes of macular holes. AB - The aim of this study was to examine the vitreomacular interface in symptom-free fellow eyes of macular holes using optical coherence tomography (OCT) to add information to the pathogenesis of macular holes and to refine prognostic factors for bilateral involvement. Sixty-six patients with a full thickness macular hole in one eye and a symptom-free fellow eye were included in the study between 01/98 05/99. The finding on OCT that a perifoveal vitreous detachment can result in a foveal cyst and subsequently a macular hole confirms the theory of Gass of vitreous traction. Symptom-free fellow eyes with a foveal cyst on OCT represent an elevated risk (55%) for macular hole development. Vitreofoveal separation is probably a good prognostic sign. PMID- 10853312 TI - [Surgery for idiopathic macular hole: use of internal limiting membrane peeling]. AB - In order to evaluate the hypothesis that macular internal limiting membrane peeling is useful for closing stages III and IV, but not stages II idiopathic macular holes, anatomic and functional surgical results are compared in a consecutive series of 54 operated eyes including 3 groups: 17 eyes with a stage II macular hole and 18 eyes with a stage III-IV macular hole without additional internal limiting membrane peeling, and 19 eyes with stage III-IV macular hole with additional internal limiting membrane peeling. PMID- 10853313 TI - [Protective efficacy of BCG Tokyo 172 in the guinea pig model of pulmonary tuberculosis]. AB - BCG Tokyo 172 strain was examined for its protective efficacy against pulmonary tuberculosis in a guinea pig model. Guinea pigs were vaccinated with an intradermal injection of 10(3) CFU of BCG Tokyo 172 strain. BCG Copenhagen 1331 was employed as a control strain. Eight weeks after the vaccination, the animals were infected with about 10 CFU of M. tuberculosis H37Rv by a respiratory route in an aerosol chamber. Five weeks after infection, the animals were euthanized and their spleens, lungs and livers were obtained for enumeration of M. tuberculosis H37Rv and histopathological examinations. The mean log10 CFU of M. tuberculosis H37Rv recovered from right lower lung lobes of guinea pigs vaccinated with BCG Tokyo 172 (frozen), BCG Tokyo 172 (freeze-dried), BCG Copenhagen 1331 (freeze-dried) and placebo were 4.72, 4.23, 4.35 and 5.76, respectively. The mean log10 CFU of the bacteria recovered from spleens were 2.11, 1.51, 1.37 and 5.90, respectively. There was a significant difference in bacterial recovery from both lung and spleen between the vaccinated and the non vaccinated groups. No significant difference was seen among the groups vaccinated with different strains of BCG in any organ. The lungs exhibited just small granulomatous nodules and the spleens showed no granulomatous nodules in the BCG vaccinated guinea pigs. On the other hand, the lungs and spleens from non vaccinated guinea pigs showed much larger granulomatous nodules with central necrosis. These histopathological difference between the vaccinated and the non vaccinated guinea pigs was consistent with the difference of bacterial growth between them. The results of this study have clearly indicated that BCG Tokyo 172 strain possesses a significant protective efficacy against M. tuberculosis as well as BCG Copenhagen 1331 strain. These results have also shown that the respiratory infection model in guinea pigs is very useful to evaluate efficacy of vaccines against pulmonary tuberculosis. PMID- 10853314 TI - [An outbreak of pulmonary tuberculosis in the dormitory of construction labors suspected to have been due to exogenous reinfection]. AB - We report an outbreak of pulmonary tuberculosis (TB) in a dormitory of construction labors, and this outbreak is suspected to have been caused by exogenous reinfection, based on the restriction fragment length polymorphism (RFLP) analysis and other findings. After a patient entered our hospital with active TB, 12 new other patients were discovered by contacts examination. These patients lived together in the same dormitory. They were all male and single, and were aged from 43 to 63 years old. Except one patient (No. 3) previously treated for TB for three months about 2 years ago and was suspected to be the index case of this outbreak, 12 other patients did not have a medical history of TB. The bacilli cultured from 11 patients (No. 1-11) were tested by RFLP analysis, three patterns were identified, and the fingerprints from 9 patients (No. 1-9) were identical, and the patterns of incomplete resistance of some antituberculous drugs were quite similar between No. 1-9 and No. 12 and between No. 10 and No. 13, respectively. The locations of the main lesions of TB on chest X-ray pictures were the apico-posterior segments of bilateral upper lobes. No signs suspected to indicate primary tuberculosis were detected. Considering the rate of tuberculous infection in Japan among the middle age and above as well as the identical RFLP results, most of patients in this outbreak except the index case No. 3 were suspected to have TB due to the exogenous reinfection. PMID- 10853315 TI - [Acute respiratory failure caused by tuberculosis requiring mechanical ventilation]. AB - The patients with active tuberculosis in whom respiratory failure requiring mechanical ventilation developed were studied retrospectively. Nine patients (M 8, F 1) were identified at the National Tokyo Hospital during 5 years from January, 1993 to December, 1997. Seven of 9 patients were single men, and the duration of symptoms before admission was over 1 month in all patients, while the time from first visit to diagnosis was less than 7 days. All patients were identified as malnourished, and 7 patients suffered from another underlying diseases. The patients were classified into two groups. Six of 9 patients had pulmonary tuberculosis and the other three had miliary disease. The proportion of cases requiring mechanical ventilation was 0.3% and 8.6%, respectively, in pulmonary tuberculosis and miliary tuberculosis. At the start of mechanical ventilation, PaO2/FIO2 was lower than 200 in all 9 patients, and 6 patients were probably ARDS. Steroids (methylprednisolone 250-1000 mg/day) were used in all 9 patients. Despite the use of mechanical ventilation and antituberculous therapy, 8 out of 9 patients died. Only one patient with miliary tuberculosis survived. The establishment of the therapy for acute respiratory failure is needed so as to improve prognosis of such cases. At the same time, the delay in consulting a doctor led to acute respiratory failure in most cases, so it is also important to encourage tuberculosis patients to visit a doctor as soon as possible, after the appearance of symptoms. PMID- 10853316 TI - [A case of primary tuberculosis, with subpleural infiltrative shadow, hilar lymphadenopathy perforated into bronchus, causing tuberculous pneumonia]. AB - A 47-year-old asymptomatic female was referred to our outpatient department with pulmonary infiltration in the left S6. PPD skin test revealed 12 x 11 mm redness. The infiltration disappeared without treatment. Hilar lymphadenopathy appeared 2 months after the first visit. In 6 months after the first visit, the hilar lymphadenopathy disappeared without treatment, but infiltration appeared in the left S6 (different site to that of the first visit). A repeated PPD revealed 47 x 40 mm redness and 20 x 15 mm induration. Bronchoscopy revealed a polipoid lesion at the orifice of left B6, MTD (Mycobacterium tuberculosis direct test) was positive from the bronchial washing of left B6. She was diagnosed as tuberculosis based on the courses and radiographic appearances. After antituberculous chemotherapy, the pulmonary infiltration and the polipoid lesion at the orifice of left B6 improved. PMID- 10853317 TI - Peripheral nerve repair and grafting techniques: a review. AB - In this review, various conventional nerve repair techniques including direct epineurial repair, grouped fascicular repair, fascicular repair, and nerve grafting are described. The indications for use, as well as the relative advantage and disadvantage, of each technique are discussed. The experimental and clinical evidence from a review of the pertinent literature does not demonstrate a significant difference in outcome of one method over the others. Surgical decisions should be made by a thorough evaluation of all aspects of the nerve injury and surgical methods. All nerve injuries cannot be repaired using only one type of nerve repair method. The surgeon should be familiar with all the techniques described and be prepared to use them under appropriate circumstances. PMID- 10853318 TI - Multivariate analysis of patients with head injury using quantification theory type II--with special reference to prediction of patient outcome. AB - Quantification theory type II was used to predict the outcome of 63 patients with head injury. Age, sex, two factors based on neurological examination, and seven factors based on findings of skull radiography and computed tomography of the head were selected as predictors. Patient outcome was evaluated 6 months after injury and assigned to good recovery, severe disability, or death. Discriminant analysis of patient outcomes was performed using the 11 factors. Two category scores were obtained for each category, since the highest correlation ratios were 0.869 and 0.252, and others were less than 10(-15). For each patient, a pair of sample scores was then obtained by simple summation of the 11 category scores. Pairs of sample scores were plotted, and the three groups of patient outcomes were clearly distinguishable without exception. These findings show that the outcome of patients with head injury can be accurately discriminated by quantitative analysis of qualitative data. PMID- 10853319 TI - Trajectory of the hypoglossal nerve in the hypoglossal canal: significance for the transcondylar approach. AB - A microanatomical study of the hypoglossal canal and its surrounding area was carried out using dry skulls and cadaveric heads to determine the course of the hypoglossal nerve in the hypoglossal canal, especially the significance for the transcondylar approach. The hypoglossal nerve enters the superomedial part of the hypoglossal canal as two bundles, which then change course abruptly to an anterosuperior direction, and unite as one trunk before exiting the canal. The hypoglossal nerve has an oblique course in the canal rather than being located in the center, and exits through the inferolateral part of the canal. A venous plexus surrounds the entire length of the nerve bundles in the canal. The present results suggest that during drilling the occipital condyle toward the hypoglossal canal from behind, the surgeon does not need to be overly concerned even if some bleeding occurs from the posterolateral edge of the hypoglossal canal. PMID- 10853320 TI - Spontaneous vertebral arteriovenous fistula--case report. AB - A 57-year-old male presented with a rare case of spontaneous vertebral arteriovenous fistula manifesting as radiculopathy of the right arm, subsequently associated with pulsating tinnitus and vascular bruit in the nape. He had a past history of chiropractic-induced vertebrobasilar infarction. Angiography showed a simple and direct fistula between the third segment of the right vertebral artery and the epidural veins at the C-1 level, where the artery runs backward above the arch of the C-1 just proximal to the penetration of the dura. The fistula was successfully obliterated by coil embolization, resulting in rapid improvement of the signs and symptoms. Mechanical compression to the nerve roots by the engorged epidural veins with arterial pressure was considered to be the major cause of radiculopathy. Vertebral artery dissection induced by chiropractic manipulation is most likely responsible for the development of the fistula. PMID- 10853321 TI - Unilateral middle cerebral artery stenosis in an adult with Down's syndrome--case report. AB - A 29-year-old male with Down's syndrome presented with severe headache and vomiting. Computed tomography demonstrated subarachnoid hemorrhage. Left carotid angiography showed severe stenosis of the middle cerebral artery 2 cm distal to its origin, as well as abnormal hyper-vascularization near the stenosis site similar to that seen in moyamoya disease. Right carotid angiography showed no abnormalities. However, slight stenosis of the distal part of the bilateral vertebral arteries was noted. There was no aneurysm. We judged that the subarachnoid hemorrhage had been caused by rupture of the moyamoya-like vessel. Some patients with Down's syndrome have anatomical vascular abnormality and vascular fragility. The cerebral vascular abnormality found in this case may be part of the systemic vascular abnormalities associated with Down's syndrome. The vascular changes in some adult patients with Down's syndrome may be a sign of premature aging, and long-term studies with periodic vascular examination of patients with Down's syndrome need to be performed. PMID- 10853322 TI - Radiation-induced cerebrovasculopathy of the distal middle cerebral artery and distal posterior cerebral artery--case report. AB - A 15-year-old girl underwent partial removal of a pituitary adenoma followed by local irradiation of the brain with a total of 70 Gy through two lateral opposing ports. Twenty years later, she experienced frequent transient ischemic attacks with left sensory disturbance. Cerebral angiography revealed stenoses of the right distal middle cerebral artery (MCA) and the right distal posterior cerebral artery without net-like vessels. There was a severe decrease of vasoreactivity in the right hemisphere. Right superficial temporal artery (STA)-MCA anastomosis was performed. Her neurological deficits were resolved and perfusion reserve capacity had markedly improved 6 months later. We recommend STA-MCA anastomosis in such cases. PMID- 10853323 TI - Clinical cure of glioblastoma--two case reports. AB - We report two cases of "decade survivor" of glioblastoma. Case 1 is a 34-year-old female who developed a tumor in the left frontal pole, and Case 2 is a 46-year old male with a left frontal tumor. Both tumors were surgically excised and the patients received postoperative chemo-radiotherapy. Histological re-evaluation of the resected tumor tissue confirmed the most malignant type of glioma glioblastoma. We speculate that the entire extent of tumors had been extirpated by surgery in both cases. PMID- 10853324 TI - Acoustic schwannoma and arachnoid cyst colocated in the cerebellopontine angle- case report. AB - A 50-year-old female presented with a right acoustic schwannoma colocated with a cerebellopontine angle arachnoid cyst. The arachnoid cyst was distinct from the arachnoid cap surrounding the acoustic schwannoma. Initial excision of the arachnoid cyst created the space required to excise the schwannoma. The acoustic schwannoma had surprisingly dense adhesions to the brainstem, probably due to the constant pressure exerted by the cyst displacing the tumor towards the brainstem. The acoustic schwannoma was excised by meticulous dissection. Such a coexisting lesion should be suspected when a large cystic collection surrounds an acoustic schwannoma. Initial excision of the arachnoid cyst will prevent excessive cerebellar retraction. PMID- 10853325 TI - Calcification of the cervical ligamentum flavum--case report. AB - A 52-year-old male presented with calcification of the cervical ligamentum flavum manifesting as hypesthesia of the bilateral middle, ring, and little fingers and ulnar halves of both forearms, as well as motor weakness in the bilateral upper extremities and gait disturbance. Cervical x-ray tomography detected a round calcified mass on the posterior wall of the cervical canal at the C-5 level. Computed tomography showed the round, nodular calcified mass more clearly. Magnetic resonance imaging showed an epidural low intensity mass compressing and distorting the cervical cord at the C-5 level on both T1- and T2-weighted images. Administration of gadolinium-diethylenetriaminepenta-acetic acid caused marginal enhancement of the mass. The lesion was eventually removed by posterior laminectomy. The mass was composed of a very hard crystal-like calcified deposition in the ligamentum flavum. X-ray diffraction analysis of the histological specimen showed calcium pyrophosphate dihydrate (CPPD) and hydroxyapatite in the crystal-like substance, confirming that CPPD is responsible for calcification of the cervical ligamentum flavum. PMID- 10853326 TI - Idiopathic hypertrophic cranial pachymeningitis with perifocal brain edema--case report. AB - A 51-year-old female presented with an extremely rare case of idiopathic hypertrophic cranial pachymeningitis manifesting as markedly thickened frontotemporal meninges with expanding perifocal edema. Magnetic resonance imaging with gadolinium revealed enhancement of the thickened dura mater protruding into the brain parenchyma accompanied by focal edema causing a mass effect. Histological examination of a biopsy specimen revealed thickened dura with infiltrating lymphocytes. Serological and immunological tests were normal. No inflammatory response or evidence of malignant tumors was observed. The patient was treated with predonisolone, resulting in marked improvement of the mass effect. High-dose steroid therapy appears to be effective for intracranial pachymeningitis associated with expanding perifocal brain edema. PMID- 10853327 TI - [Clinics and molecular biology in colorectal diseases]. PMID- 10853328 TI - [Current status of clinical approach for breast cancer in the collaborating institutes of the surgical departments of Kyoto University--cancer conference of the collaborating institutes of the surgical departments of Kyoto University]. PMID- 10853329 TI - Two cases of appendicitis in Kawasaki disease. AB - Two cases of appendicitis in Kawasaki disease (KD) are presented. Appendicitis in KD is very rare condition, and only a few cases have been previously reported. This report suggests the need to expand the differential diagnosis of abdominal pain in this group of patients. PMID- 10853330 TI - [Two cases of traumatic-diaphragmatic hernia]. AB - We report two cases of traumatic-diaphragmatic hernia caused by blunt injury. In case 1, abdominal findings were discovered on the 2nd day after the patient fell on the bottom from a horse, and in case 2, an emergency laparotomy was carried out due to the respiratory dysfunction which appeared immediately after a labor injury. The diaphragmatic ruptures at the left side observed in both cases, were closed directly. The prolapsed organs which shifted into the thoracic cavity were the stomach and colon in case 1, and the spleen and the tail part of the pancreas in case 2. In both cases, the organs were easily repositioned into the abdominal cavity without a hernial sac, but a splenectomy had to be performed because of splenic lacerations. In case 2, right rib fractures and lacerations of the left adrenal gland with retroperitoneal hematoma were observed. In the case of thoraco abdominal blunt injuries, it is important to perform an early diagnosis and to consider the possibility of diaphragmatic hernia. PMID- 10853331 TI - [A study of the clinical effect of estradiol transdermal therapeutic system alone on pollakisuria and urinary incontinence in postmenopausal woman]. AB - PURPOSE: Our purpose of this study was to evaluate the effect of a transdermal estradiol delivery system in postmenopausal women with confirmed pollakisuria and urinary incontinence. PATIENTS AND METHODS: We investigated 10 postmenopausal women, age 54-83 years, with pollakisuria and urinary incontinence but did not show distinct urological and/or neurological abnormalities. In this study, estradiol transdermal therapeutic system (Estraderm TTS 2 mg) alone were administrated for total of 8-week and this is observational, not randomized, blinded or controlled. A clinical evaluation were performed two times at before and after administration. RESULTS: In seven eligible cases, the severity of urinary incontinence was graded down in almost of them and the therapeutic effect on urinary incontinence was evaluated as "very effective" in 3 cases, "improved" in 2, "slightly improved" in 1 and "no change" in 1, respectively. In three eligible cases, the severity of pollakisuria was no change in all of them. CONCLUSION: Thus, the estrogen supplement therapy was considered effective for postmenopausal urinary incontinence. PMID- 10853332 TI - [Color Doppler ultrasonography in the diagnosis of the acute scrotum]. AB - PURPOSE: To determine the value of color Doppler ultrasonography (CDUS) in the diagnosis of acute scrotum. MATERIALS AND METHODS: 10 patients referred to our hospital with acute scrotal pain were included in this study. All patients were evaluated with CDUS after the initial clinical examinations. Blood flow of the involved testis was compared semiquantitatively to that of the opposite testis. Patients with a diagnosis of testicular torsion by CDUS underwent surgical exploration. Patients with CDUS diagnosis of epididymitis were treated with intravenous antibiotics. RESULTS: Of the 10 patients evaluated, CDUS diagnosed 5 patients with testicular torsion and 5 patients with epididymitis. All cases of torsion were confirmed intraoperatively. 2 cases with no intratesticular blood flow on CDUS had necrotic testes and underwent orchiectomy with orchiopexy of the contralateral testes. A case with absent flow and 2 cases with decreased flow had bilateral orchiopexy. CDUS findings of normal or increased flow were present in all patients with epididymitis. No cases of testicular atrophy were encountered on long-term follow up in patients with epididymitis. CONCLUSION: CDUS is helpful in detecting the perfusion of the testis as well as in getting anatomical information. CDUS is a very useful device which causes a minimal burden to the patient with acute scrotum. In most cases it will rapidly provides us the correct information although it may not be regarded as the definitive adjunct. PMID- 10853333 TI - [An investigation of serum soluble interferon receptor levels in patients with renal cell carcinoma]. AB - OBJECTIVE: Serum soluble interferon alpha/beta receptor (s-IFN-receptor) levels were determined in renal cell carcinoma (RCC) patients to study the clinical significance of the measurement. SUBJECTS AND METHODS: S-IFN-receptor levels were measured in RCC patients (n = 27) and healthy volunteers (n = 22) by enzyme immunoassay technique. RESULTS: Significantly higher serum s-IFN-receptor levels were observed in RCC patients compared with the healthy volunteers (p < 0.003). The high s-IFN receptor levels in the patients suggested seriousness and mal prognosis of this disease. The 4-years survival rate of the higher level group (with the mean value of 2.7 +/- 1.7 ng/ml or more) was 53.3%, while the lower level group's rate was 78.7% (Statistical analysis result by Logrank (Mantel-Cox) test; p = 0.4289). CONCLUSION: Further study in more subjects is required to determine the feasibility of the s-IFN receptor levels as a prognosis marker, since correlation between the prognosis and s-IFN receptor level was not clarified by this study result. PMID- 10853334 TI - [Clinical review of 39 cases with ectopic ureteroceles]. AB - BACKGROUND: There are many controversies surrounding the management of ectopic ureteroceles (EUC). The aim of this study is to review our cases with EUCs and to show our policy of choice of treatments of EUCs. METHODS: The medical records of 39 patients with EUCs treated at Kobe. Children's Hospital from 1978 to 1998 were reviewed retrospectively. Patients' age, affected site, presentation, treatment, and postoperative course were recorded. RESULTS: The age at presentation ranged from 0 month to 13 years (mean; 6 years). The left EUCs were found in 15 patients, the right in 17 patients, and the bilateral in 7 patients. The EUCs with duplicated system of the kidney were involved in 35 cases (42 kidneys) and single system in 4 cases (4 kidneys). The most common mode of presentation was urinary tract infection (n = 24) followed by abdominal distention (n = 6) and fetal ultrasonography (n = 6). One patient presented with incontinence and in two patients EUCs were discovered incidentaly. Thirty-five cases (42 kidneys) were followed up over six months. In these cases diversion including nephrostomy and ureterostomy was performed in 5 kidneys, heminephrectomy and/or excision of the EUC and ureteral reimplantation in 8 kidneys, nephrectomy in 3 kidneys, pyeloureterostomy in 2 kidneys, excision of the EUC and ureteral reimplantation in 10 kidneys, and transurethral incision (TUI) of the EUC in 14 kidneys. After these treatments the second surgery was totally required in 15 kidneys(36%) including 7 kidneys in which TUI was performed. Furthermore, in two kidneys the third operation was performed. CONCLUSIONS: Reoperation was required in about one third of patients with ectopic ureteroceles. It is easy to perform TUI, however the rate to reoperation is high. PMID- 10853335 TI - [Metachronous bilateral primary malignant lymphoma of the testis: a case report]. AB - A 72-year-old man was referred to our department with the chief complaint of painless swelling of the left scrotum in May 1997. Left high orchiectomy was performed under the diagnosis of primary testicular tumor. Histological findings revealed non-Hodgkin's lymphoma (NHL) of diffuse, mixed type, B cells. No evidence of tumors in any other site was detected by further examinations. About 3 years and a month earlier, he had undergone right high orchiectomy and postoperative radiotherapy (inverted Y irradiation) and chemotherapy (CHOP 5 cycles) for a right testicular tumor whose histological findings were NHL of diffuse, large cell type, B cells. Metachronous bilateral primary malignant lymphoma of the testis is very rare and we discussed each tumor origin by using IgH gene (IgJHDNA) rearrangement as a tumor specific marker of B cell lineage malignant lymphoma. We discussed the clonality of IgJHDNA rearrangement using polymerase chain reaction (PCR) in each paraffin fragment diagnosed pathologically as NHL of B cell origin. PMID- 10853336 TI - [An autopsy case of prostatic neuroendocrine cell carcinoma and adenocarcinoma initially found by brain and abdominal wall metastasis]. AB - A 62 year-old man had been suffered from headache and left shoulder pain since March 1997. In November 1997, he visited our hospital complaining of work incapability, slow-moving and somnolence. Multiple nodular lesions were found in his brain and abdominal wall. Biopsy of the abdominal wall mass revealed small cell carcinoma/neuroendocrine cell carcinoma. Radiation therapy on brain and abdominal wall was done and these tumor nodules became decreased. However, recurrence and metastasis occurred later and died at March 1998. The autopsy revealed the origin of these tumors was the prostate. The prostatic tumors revealed neuroendocrine cell carcinoma mainly, combining a portion of adenocarcinoma. Most parts of the metastatic tumors were neuroendocrine cell carcinoma. Only the seventh thoracic vertebral metastasis was bone-sclerosing metastasis of adenocarcinoma. PMID- 10853337 TI - [Study of matrix metalloproteinase-2 and -9 in thyroid papillary cancer]. AB - Metastasis of cancer starts with the penetration of cancer cells through the membrane surrounding the cancer focus into the stroma (extracellular matrix). The focal membrane consists of mainly type-IV collagen. An immunochemical study of 28 patients with benign thyroid nodular diseases and 27 patients with papillary carcinoma revealed the fragmentation of type-IV collagen in 4 patients with papillary carcinoma. Matrix metalloprotease (MMP)-2 and MMP-9 are the major enzymes which decompose type-IV collagen, and they have been suggested to be related to cancer metastasis. Therefore, we conducted biochemical and immunohistochemical studies to determine the relationship between these MMPs and the degree of malignancy in thyroid diseases. The concentration of MMP-2 in the serum of patients with papillary carcinoma and patients with benign nodules was 526.0 +/- 96.6 and 522.7 +/- 114.6 ng/ml, respectively, and that of MMP-9 was 53.8 +/- 40.3 and 39.9 +/- 36.0 respectively. There was no significant difference between the two groups in the concentration of either enzyme. The concentration of TIMP-2 in the serum was below the detectable level. On the other hand, the concentration of MMP-2 in the tissue of papillary carcinoma, benign nodules and normal tissue was 12.1 +/- 8.1, 5.7 +/- 4.3, and 0.6 +/- 0.5 ng/mg tissue protein, respectively, and that of MMP-9 was 4.2 +/- 4.1, 2.1 +/- 1.7, and 0.4 +/ 0.3 ng/mg tissue protein, respectively. Concentrations of both enzymes were significantly higher in the papillary carcinoma tissue. Immunohistochemical studies revealed a diffuse granular distribution of MMP-2 in the cytoplasm of the tumor cells. These findings imply that MMP-2 and MMP-9 are related to the degree of malignancy of cancer, especially metastasis. PMID- 10853338 TI - [Treatment of 522 patients with sudden deafness performed oxygenation at high pressure]. AB - INTRODUCTION: Oxygenation at high pressure (OHP) is thought to be useful, even though regional blood flow is decreased, because increasing dissolved oxygen prevents the death of nerve tissue. In this report, we retrospectively investigated the effect of OHP on sudden deafness. OBJECT AND METHOD: We reviewed 522 patients treated with OHP at Kagawa Rosai Hospital over a ten-year period (January 1989 to December 1998). We discussed some prognostic factors: comparison between cases which had been treated with OHP previously and those which had not, number of days between onset and beginning of the treatment which included OHP, age, initial averaged five-frequency hearing level, vertigo, tinnitus, complications of OHP, cases of relapse and the time of the onset, which is about season, month and week. OHP was administered at a pressure of 2.5 atmospheres for 80 minutes a day from 10 to 15 times. All patients also received a course of intravenous administration of steroid, vitamin B12, Prostaglandin E1, ATP, and low-molecular dextran. RESULTS: Overall, complete recovery occurred in 19.7% of the patients, definite improvement in 34.9% (complete recovery included), and slight improvement in 58.1% (definite improvement included). Most of the patients (78.0%) were referred by other hospitals, because our hospital was the only one in the Sikoku area which had a big equipment of OHP. All 161 patients had already been treated in other hospitals over 8 days, but they had shown little improvement after the initial therapy. Of this group, complete recovery after the second course of treatment occurred in 13.0% of the patients, definite improvement in 19.3%, and slight improvement in 39.1%. OHP was thus effective for about 40% of patients who had been unresponsive to the initial therapy. Delay in treatment usually produces poor hearing recovery. There was a significant difference between those patients treated within 14 days and those treated 15 days or more after onset. The improvement rate also decreased with age. The prognosis of patients with vertigo was worse than those without vertigo. Tinnitus had no influence on the prognosis. There were no severe complications during the course of OHP, but otitis media with effusion occurred in 90 patients, and paracentesis was performed for 53 patients. CONCLUSION: The treatment of sudden deafness with OHP has been discussed in this report. Important prognostic factors were time between onset and beginning of the treatment which included OHP, age, vertigo, and the initial averaged five-frequency hearing level. We conclude that OHP should be performed within 14 days from onset, and that OHP was able to achieve hearing improvement in many cases unresponsive to the initial therapy if it was performed very early. PMID- 10853339 TI - [Endoscopic sinus surgery in flowing water]. AB - A balloon has been developed that completely fills the choana, preventing water from leaking into the pharynx even when the water is entering into the nasal cavity at a rate of 1000 ml per minute. The balloon enables endoscopic sinus surgery (ESS) to be safely performed in "flowing water". This surgical technique is similar to that used in transurethral resections of the prostate because the tip of the endoscope is kept clean, and blood, debris and resected tissues are continuously removed by the water flow. In addition, the water pressure helps to suppress bleeding. This technique enables ESS to be performed with greater ease and efficiency. We have performed ESS in flowing water on 38 patients with chronic sinusitis under local anaesthesia. No complications, such as water leakage into the pharynx, were encountered, and only a few patients felt discomfort from the insertion of the balloon. Even if the balloon had burst, an emergency could have been easily prevented by withdrawing the endoscope from the nasal cavity and stopping the flow of water. Ultrasonography (USG) was used to examine the water-filled nasal cavity during surgery (SSD-2000 and Micro Tip Radial (ASU-101); Aloka, Ltd., Japan). Using USG, the middle turbinate, the inferior turbinate and the nasal septum could be visualized in a single coronal image. When the sensor was in the posterior ethmoid sinus, the orbit and its optic nerve could also be visualized. Since this surgery is performed under local anesthesia, eye movements can rapidly alter the position of the optic nerve. Thus, visualization of the optic nerve's exact position is extremely important. Unfortunately, USG is not very useful for localizing structures and guiding the surgeon to distant tumors or cysts located behind thick bones, since ultrasound can not penetrate hard masses or bones. In this situation, navigation systems are more reliable than USG. Nevertheless, USG is often useful for depicting surgical sites, especially during a crisis, if the medial wall of the orbit is thin or if the skull base has been broken, exposing the dura. USG can also provide early warning of an impending complication. USG also has several practical advantages over navigation systems: the cost of USG is much lower, preparation for surgery is unnecessary, visual information can be obtained in real time, and measurement accuracy (estimated to be about 2 mm for navigation systems) is not a consideration. Thus, USG can be easily used to avoid complications in most surgeries for chronic sinusitis. Flowing water also allows the nasal eavity to be completely washed and sterilized at the end of the surgery. This not only prevents post-operative infection, but enables sinus function to be more quickly recovered. In addition, the pressure from the balloon also prevents post operative nasal hemorrhaging. This allows patients to be safely discharged from the hospital at an earlier time. The balloon can also be used for non-surgical purposes. For example, emergent epistaxis can be easily stopped by the insertion of this balloon, even if the doctor is not an otorhinolaryngologist. In addition, the balloon's soft pressure allows it to be left in the nose for long periods without any complications. We conclude that this simple balloon, which we have named the "Noda Balloon", is extremely useful for nasal treatments. PMID- 10853340 TI - [Cervical lymph node metastasis from an unknown primary tumor]. AB - Between June/1986 and June/1999, we treated 18 cases of previously untreated metastatic carcinoma in cervical lymph nodes from an unknown primary tumor. Among them, 15 cases were squamous cell carcinoma, 2 were adenocarcinoma, and one was undifferentiated carcinoma. The dose of radiotherapy, which was planned only for the cervical lymph nodes, was 60-70 Gy at the metastatic sites. Preventive radiotherapy at possible primary sites was performed only in one case. After these treatments, a primary lesion appeared in only two cases: in one case a tumor was found in the hypopharynx, and the other, which had been diagnosed as undifferentiated carcinoma, proved to be malignant lymphoma. Outcomes were analyzed except for these two cases. The five-year survival rate was 31%. The loco-regional control rate was 57% after radiation treatment limited to the affected cervical lymph nodes. Distant metastasis appeared more often in open biopsy cases than in those receiving fine-needle aspirations (FNA). According to these results, treatment of possible primary sites may not be necessary, and for initial pathological diagnosis, FNA is recommended, while open biopsy should be avoided, if possible. PMID- 10853341 TI - [Clinical study of time-shift evoked potentials]. AB - Quick change of the interaural time difference (ITD) generates moving sound stimuli. Specific biphasic event-related potentials called "time-shift evoked potentials (TSEPs)" can be recorded when this moving sound is given. The results of TSEPs in various types of hearing loss were analyzed in comparison with auditory brainstem response (ABR) and slow vertical response (SVR) in order to evaluate clinial applicability of TSEPs. Firstly, the detection threshold of TSEPs was established in groups of patients with low tone sensorineural hearing loss and with steep high tone sensorineural hearing loss. The usefulness of TSEPs was then evaluated in patients with retrocochlear hearing loss and with functional deafness. The patients with retrocochlear hearing loss were divided into 2 subgroups, one with auditory nerve disorder and the other with cortical disorder. It was found that TSEPs participate in the transference of auditory time-factors. They reflect the function of not only the auditory nerve and brainstem which form major components of ABR, but also the central nervous system superior to the inferior colliculus. TSEPs could be recorded in most patients with functional deafness and are more useful for its diagnosis than using the conventional directional hearing test. It is concluded that TSEPs is useful as a clinical test for detection of cortical disorder and functional deafness. PMID- 10853342 TI - [Fatty degeneration of the parotid gland after ovariectomy]. AB - We investigated the influence of ovariectomy on the salivary glands. Thirty-six rats received ovariectomies at the age of 15 weeks, and removal of parotid, submandibular and sublingual glands at the ages of 18, 21, 25, 27, 33, and 39 weeks. The weight of each gland and the total body were measured. Histological examinations noted the point of fatty degeneration. Twelve female rats were used as controls, each gland being observed at the age of 19, 28, and 41 weeks. Body weights of the ovariectomized rats were greater than those of the controls, but neither the weights of the parotid gland nor the submandiblar-sublingual gland were different. Fatty degeneration was observed only in the parotid glands and it increased gradually. There was no change in the parotid gland of the control group. The ratio of the area of fatty tissue showed a statistically significant increase over time. These results suggest that the parotid gland requires ovarian hormone to maintain its tissue. PMID- 10853343 TI - [Relationship between intensity of annoyance due to throat discomfort and psychological test results]. AB - For 87 male and 100 female patients (average age of 58.5 years) with throat discomfort, we investigated the relationship between the intensity of annoyance caused by throat discomfort and psychological distress. We asked patients to assess their throat discomfort using a 100-mm horizontal visual analogue scale (VAS) which defined the left end (0) as no annoyance and the right end (100) as overwhelming annoyance attributed to throat discomfort. The extent of their psychological distress was evaluated by 3 tests: Cornell Medical Index-Health Questionnaire (CMI), Self-rating Depression Scale (SDS) and Self-rating Questionnaire for Depression (SRQ-D). Our results indicate that the increase in intensity of annoyance caused by throat discomfort was significantly related to the increase in somatic symptoms (Spearman's rho = 0.164, p = 0.025), symptoms connected to autonomic nerve dysfunction (rho = 0.203, p = 0.006) as measured by CMI and neurotic condition as classified by Fukamachi's diagnostic method for CMI (p = 0.049 by Kruskal-Wallis test). The increase in intensity of annoyance caused by throat discomfort was significantly related to the increase in depressive condition as measured using SRQ-D (rho = 0.263, p < 0.001), but not to that measured using SDS (rho = 0.097, p = 0.185). The results obtained from the present study indicate that VAS is a useful tool for measuring annoyance caused by throat discomfort, yielding valuable information about the intensity of patients' somatic and psychological complaints. PMID- 10853344 TI - [Molecular analysis of pathogens of upper respiratory tract infections in children--a study of nasopharyngeal S. pneumoniae and PBP genes in acute otitis media]. AB - We have recently been confronted with refractory upper respiratory infections with an increasing prevalence of penicillin (Pc)-resistant S. pneumoniae. There has been a broad consensus that acute otitis media (AOM) is caused by migration of pathogens from nasopharynx and proliferation in the middle ear space, and thus it is, very important to study the bacterial environment in the nasopharynx as the source of middle ear infections. Eighty pneumococcal isolates from the nasopharynx of children with acute otitis media were evaluated by polymerase chain reaction (PCR) for mutation of Pc-binding protein (PBP) genes. The results showed mutation of all three PBP genes, pbp 1a, pbp 2x, and pbp 2b, in 30% of the isolates, while 74% were found to possess various PBP gene mutations, mostly in one-year-old children. Of the 46 isolates whose minimum inhibitory concentration (MIC) of Pc was < or = 0.06 microgram/mL, 43% were found to possess a pbp 2x mutaion, which affects cefem resistance. We genotyped each pneumococcal isolate from the nasopharynx of children with recurrent AOM by pulsed-field gel electrophoresis (PFGE). In 9 of 11 pairs (82%) of consecutive AOM episodes, the nasopharyngeal isolate in the second episode was different. In addition, discrimination of each isolate based upon the mutation profile of its PBP genes in 8 pairs (72%) of consecutive AOM episodes showed that the isolates were different, and there was little difference between the results of PBP gene mutation and PFGE analysis. These findings suggest that most nasopharyngeal isolates from children with AOM possess PBP mutations and that children with increased numbers of drug-resistant bacteria in their nasopharynx during AOM has been colonized or recolonized by different strains during each episode. We therefore emphasize that clinicians should assess the antibiotic susceptibility of nasopharyngeal isolates from children during each episode. PBP gene mutation analysis of S. pneumoniae is useful not only in providing valuable information on the antibiotic susceptibility of each strain but for assessing changes in causative strains in the sequential episodes of pneumococcal infection. PMID- 10853345 TI - [Pharmacologic therapy of Crohn's disease and ulcerative colitis]. AB - Important progress has been made in recent years in the understanding of pathogenesis of Crohn's disease and ulcerative colitis, but the cause of IBD remains obscure, so curative therapy is still lacking. Current treatment strategies as sulphasalazine, mesalasine, glucocorticosteroids are mainly anti inflammatory. In the past years the greatest advances have been characterised by the more widespread use of topically acting steroids, immunosuppressants and by the introduction of immunomodulatory agents as cytokines and anticytokines. The author summarises the standard therapy and new possibilities of medical treatment for IBD and suggests some algorythms for clinical practice. PMID- 10853346 TI - [Genetic counseling and prenatal care after medications during the first trimester]. AB - Analysis of the outcome of 127 pregnancies with first trimester medication (8.4% of the total number of the patients seeking genetic advice in 1997 at the Institute of Medical Genetics in Szeged) was carried out. The gestational age at the time of the medication and genetic counselling, the indications of the treatment, the drugs, the estimated fetal risk, and results of genetic ultrasound examinations and pregnancy outcome were evaluated. The majority of pregnant woman (78%) asked for genetic counselling before the 12. gestational week. The main indications the treatment were: infections, psychiatric-neurologic (depression, anxiety, epilepsy), endocrine (diabetes, hyperthyreoidism), and cardiovascular diseases and gastrointestinal problems. The main groups of the drugs were: antibiotics, antipyretic-, antidepressive-, antidiabetic- and antihypertensive drugs. When the multiple medication was conducted by simultaneous administration of two or more drugs, a complex risk calculation was performed. The fetal risk was higher than 10% in 31 cases (24%). The ultrasound examinations performed by qualified sonographer contributed to a correct evaluation and to reliable follow up of pregnancies. No suspicious ultrasound finding was reported in the first trimester. However, a severe fatal brain malformation was found in a second trimester pregnancy, which was terminated by the couple's request in the 18th gestational week. A complete follow-up was obtained in 70.9% (90) of the cases. Out of 64 pregnancies intended to continue to term 4 fetal malformations were found. Of them three malformations (patent ductus arteriosus, Robin sequence and a ventricular septal defect) were explored at birth or in the newborn period. The actual 6.3% of fetal malformations was higher compared to the rate expected at birth, but almost equal to the rate of congenital malformation found up to the end of the first year of age in Hungary. PMID- 10853347 TI - [Long-term results of tube insertion in treating otitis media with effusion]. AB - The authors present the long-term results of 50 patients, who were treated with tube insertion because of chronic serous otitis. During the 7 years observation period 60% of the patients were healed, in 40% of the patients at the end of the observation period negative pressure with effusion or without effusion were found or tube was inserted. In 5% of their patients even repeated tube insertion could not prevent the developing of adhesive otitis or cholesteatoma. On the basis of patients material it was found, that the period of the existence of serous otitis is a major factor in the development of irreversible damage. However the usage of the pressure equilibration tube possibly decrease the irreversible damage, it will be a future to establish new treatment methods. PMID- 10853349 TI - [History of the foundation and location of the military hospital in Buda in 1789]. PMID- 10853348 TI - [Cognitive enhancement effect of piracetam in patients with mild cognitive impairment and dementia]. AB - Effectiveness and tolerability of two piracetam-containing drugs were compared in the frame of an open, multicentre, Phase-IV, prospective study with group and self control carried out in 9 Hungarian centres in 1998. Patients with cognitive decline from Alzheimer's disease and/or cerebrovascular origin have received the drug, in the first 4 weeks in 4800, later 2400 mg daily doses. One hundred four patients finished the study. No relevant difference with statistically significant degree was registered between the two groups. Based on this fact in this study data of the 104 patients were examined together. Authors examined two problems. The first: on which cognitive function is more effective the drug. Five factors of the modified Mini-Mental State Examination were separated and compared. Nearly all of them significantly increased especially the factors of memory, and concentration-psychomotor speed. The second examined field: are there certain subgroups with prognostic value about the effectiveness of piracetam treatment. Neither the duration of the illness, nor etiologic diagnosis, severity of cognitive decline, or former treatment with piracetam did influence significantly the efficacy. In case of depressive symptoms such connection was established: the more pronounced the severity of these symptoms the higher improvement can be expected in cognitive functions. This was also stressed by the logistic regression analysis. Authors described an original evaluation method of the trail-making test, which could widen the application of this popular test in psychopharmacologic studies. Final conclusions: the cognitive enhancer effect of piracetam appeared in a few weeks. This treatment could be effective even in Alzheimer's disease, in case of more pronounced cognitive decline, longer duration of the illness, and in case of former piracetam treatment as well. The degree of cognitive improvement was most pronounced in patients with comorbid depressive symptoms. PMID- 10853350 TI - [Vincenz Priessnitz (1799-1851)]. PMID- 10853351 TI - [Periodic albuminuria. 1900]. PMID- 10853352 TI - [What do the dead teach us?]. PMID- 10853353 TI - [Neurotic symptoms frequency]. AB - The analysis of symptom checklists completed by 3196 patients before treatment- 1970 females and 1226 males starting the therapy due to neurotic disorders- revealed that 12 symptoms occur im more than 80% of patients; 9 symptoms in 70 80%, 18 dysfunctions in 70-60% and 17 other dysfunctions in 60-50% of examined population. This means that a large number of symptoms including tension, mood depression an anxiety as well as difficulties in concentration, lack of self confidence, sense of tiredness, loss of energy, persistent thoughts and images, constant fear, absent-mindedness, motor tensions, pessimism, thought flood, the sense of difficulty in thinking and tachycardia occur in almost all patients suffering from neurotic disorders, mo matter what these disorders are. Tle largest group consisted of patients with the diagnosis of anxiety disorders like phobias and the like (ca. 25%). Results of the study suggest the necessity of verification of the present views on the picture of dysfunctions combining into neurotic disorders. A similar occurrence rate of ca. 30% of the 95 analysed dysfunctions in the subgroups of females and males as well as a higher occurrence rate of 10% of the dysfunctions in the male group also make it necessary to verify the prevalent convictions about the sex-dependent differences in the picture and course of neurotic disorders. PMID- 10853354 TI - [Frame of mind among Polish population in research carried out by the Central Statistical Office: initial analysis]. AB - The authors are going to estimate the prevalence poor frame of mind and neurosis among Polish adult people, and try to appoint the relationships between psychiatric disorders and gender, age, civil status, education and maintenance. The questionnaire contains questions about quality of sleep, possibility to fix one's attention on acting, inner tension, self-confidence. Almost 1/4 of women and 18% of men have poor frame of mind. We find very strong and important relationships between neurosis and the poor frame of mind. There were no differences in mental state between people living in towns and villages. The unemployed and the cripples have worse psychological condition than working men. People who are divorced and widowed have statistically more often poor frame of mind and neurosis than the married. We also found a major correlation between poor frame of mind and neurosis and education. Low education is connected with poor psychological condition. PMID- 10853355 TI - [The issue of cults in psychiatry]. AB - The cult indoctrination, mechanisms operating within sects, manipulation techniques and the role of the leader are presented in the article. Authors discuss personality features and situational factors leading to cult-entering and psychopathological effects on cult members. There are also the basics of brainwashing and its efficiency included, followed by the psychiatrists and families position in diagnosing and therapy of cultists. PMID- 10853357 TI - [An estimate of family doctor's knowledge of diagnosis and treatment of depression and anxiety disorders]. AB - Depression is more widespread than hypertension. Nevertheless, physicians specialized in family medicine did not have knowledge necessary to diagnose and treat depression and anxiety disorders. Only six persons out of fifty five who filled in the questionnaire could recognize depression correctly (ICD-10 criteria). Doctors knew antidepressant drugs, but they did not know therapeutic doses. We can try to explain this situation through doctor's pre and postgraduate education. PMID- 10853356 TI - [Depressive disorders in general medical practice, particularly in basic health care]. AB - Early, correct treatment of depression as well as introduction of prophylactic procedures highly improves the prognosis in affective disorders. Periods of relapse become shorter, the risk of their occurrence decreases, the risk of suicidal attempts is much lower and the too high death rate in the younger age groups goes down. Epidemiological studies and analyses show that a large proportion of patients with depression does not receive any help of the health service, some receive psychiatric help and some contact general practitioners. The latest group constitutes 10-20% of patients coming to various institutions of basic health service. Only in 25% of this group depressive disorders are recognised, even less patients receive effective help. Physicians employed in basic health service often treat depression inappropriately (an erroneous selection of medicines, inappropriate, usually too low dosage of the drug, too short the period ot treatment). In order to improve the effectiveness of medical help provided to the patients with depressions, various educational actions for physicians employed in the basic health service are organised. Their aim is to improve the physicians' competence in diagnosing and treatment of depression. As the hitherto experience of execution of educational programs shows, the necessary condition of improving the efficiency of first contact physicians in treatment of patients with depression consists in providing them with opportunities to receive systematic supervision and consultation of psychiatrists. PMID- 10853358 TI - [Depressive disturbance and quality of life in patients with coronary artery disease]. AB - The aim the study was to evaluate the impact of mood disturbances on the subjective quality of life in patients with coronary artery disease after an effective angioplasty. The study covered 100 patients with the optimum result of PTCA. Their condition was evaluated one day before and four weeks after angioplasty. Significant differences in the subjective quality of life assessment were detected depending on the occurrence and dynamics of depressive symptoms. The authors postulate evaluation of psychological state and introduction of anti depressive therapy in patients with coronary artery disease subjected to revascularisation. PMID- 10853359 TI - [Development and application of cognitive therapy in affective disorders]. AB - Cognitive psychotherapy was originally created for out-patient treatment of mild and moderate, non-psychotic, unipolar depressive disorder. Further development of the therapy resulted in its use in various mental disorders. Cognitive therapy has also been used in wide spectrum of affective disorders, including severe, endogenous depression, chronic depression, bipolar disorder and suicidality. Therapeutic programs involve individual, group, family and marital cognitive psychotherapy. Effectiveness and clear conceptualization encourages to wide use of this kind of therapy. PMID- 10853360 TI - [Oedipus complex in the case of sexual abuse]. AB - The paper describes a specific form of Oedipus complex that may occur as a result of sexual abuse. The author emphasises that formation of the image of one parent due to actual sexual abuse and the activity of the child's (patient's) own iscestual impulses causes that the second parent is experienced as a terrorizing and revenging rival. As a result, the child in his/her psyche, must face not only his/her own incestual desires, but also the parental objects, which do not provide any support, sense of safety or stability of existence. The author illustrates his thesis with a summary of three sessions of analytic psychotherapy of the patient with whom he worked in Cassel Hospital in London. In the described case, it can be seen how the patient's involvement in the therapy and her good will connected with it stimulated her incestual desires addressed at the analytician, what, in turn, activised the patient's destructive, punishing behaviour. PMID- 10853361 TI - [The influence of genes on the development of personality]. AB - Antisocial behavior and personality disorders are both heterogeneous and the product of interacting genetic and environmental factors acting at different levels of causation. Heritability studies show that individual differences in predisposition to antisocial behavior are transmitted by genetic mechanisms in families. Direct gene analysis and genetic linkage analysis have identified structural variants in genes involved in neurotransmitter function, and some progress has been made towards relating these genetic variants to antisocial personality and other behaviors. The monoamine oxidase-A variant leads to aggressive behavior in one family. Direct gene analyses have revealed amino acid substitutions and structural variants at DRD2, DRD3 and DRD4 dopamine receptors and 5-HT2A, 5-HT2C serotonin receptors, serotonin transporter gene, and genes for enzymes, metabolizing biogenic amines MAO-A, MAO-B. The stage is set to identify the phenotypic significance of these as well as genetic variants at other loci which may be relevant as candidate genes for antisocial behavior and related behavioral differences. PMID- 10853362 TI - [The behavioral effects of the transcranial magnetic brain stimulation in rat: the comparison with electroshock]. AB - The few experimental studies suggest that repetitive rapid-rate transcranial magnetic stimulation (TMS) evokes in the brain functional and structural changes similar to those evoked by electroconvulsive therapy (ECT). The aim of the present work was to compare the influence of the repetitive TMS (B = 1.6 T; f = 20 and 30 Hz; t = 5 and 5.5 minutes; N = 9 and 18 days) and that of ECT (I = 150 mA; f = 50 Hz; t = 0.5 s; N = 9 days) on rats' behaviour in the tests of free field, tail flick, motor hyperactivity after administration of apomorphine and in forced swimming. None of the rats subjected to TMS suffered from convulsive attack, which followed every electroconvulsive shock. In the free field test it was detected that neither TMS nor ECT applied individually or repetitively disturbed general motor activity of rats. Repetitive electroconvulsive shocks caused analgesia, prolonging the latency of tail flick by 46% (p < 0.001). Moreover, the tail flick test revealed hyperalgesia in the rats subjected to TMS (24 and 21% of control values respectively; p = 0.05). Motor hyperactivity of rats stimulated with administration of apomorphine was intensified both by TMS (by 58% at maximum in the 30th minute of the experiment; p = 0.001) and, all the more, by ECT (by 92% at maximum at the end of the test; p = 0.01). In the forced swimming test, the greatest decrease of inertia period was observed ECT--up to 50% of control values (p = 0.001). TMS had weaker effects--the decrease amounted to 29% of control values (p = 0.01). The shortening effect depended on the parameters of TSM. The obtained results seem to confirm that TMS, like ECT, evokes in rats certain reactions suggesting its antidepressive action, but causes less undesirable effects. PMID- 10853363 TI - [Function of NF-E2-related transcription factors]. PMID- 10853364 TI - [Roles of cell surface carbohydrates in cell adhesion; in particular those for sialyl Lewis x and polysialic acid]. PMID- 10853365 TI - [Mechanism of interaction of ras with its effectors]. PMID- 10853366 TI - [FGF10, a key factor for limb and lung formation]. PMID- 10853367 TI - [Small Maf proteins in bZip transcription factor net-work]. PMID- 10853368 TI - [Molecular biology of histidine decarboxylase]. PMID- 10853369 TI - [A sexual reproduction and proteases in tunicates: the role of TRAMP inproliferation and dedifferentiation of multipotent cells]. PMID- 10853370 TI - [Regulation of ionotropic glutamate receptors by their phosphorylation]. PMID- 10853371 TI - [Medical malpractice (II). Details and examples]. PMID- 10853372 TI - [Cost developments in private health insurance]. PMID- 10853373 TI - [Value of specialized pain treatment]. AB - The paper describes new forms of specifically organized headache prevention and headache therapy in Germany with the aid of two examples. It provides an overview of the clinical and economic efficiency of facilities with new forms of organization for specialized pain therapy. Their effectiveness can be seen both in a marked reduction in individual suffering and in low-cost provision of services for the institutions bearing the costs and for society. This effectiveness can be seen not only from subjective parameters by interviewing the patients, but also from objective target parameters such as resumption of work, reduction in intake of medication, cost reductions due to reduced demands on the healthcare system, avoidance of premature pensioning. By contrast, patients who do not receive specially organized treatment despite the fact that they need it display either constant suffering or even a worsening of their symptoms and an increased need for financial compensation. In view of this clear situation, specialized headache and pain therapy with a special focus on catering for the needs of chronic headache suffering is called for, both on economic and ethical grounds. PMID- 10853374 TI - [Aspects of occupational disability in psychosomatic disorders]. AB - In 1997, 30% of the persons going into early retirement because of occupational disability and received pensions were psychosomatically ill. An additional large number of retirees suffered from untreatable pain such as chronic low back pain, some of them might as well have a chronic somatoform pain disorder. The article describes frequent psychosomatic diseases like somatization disorder, fibromyalgia and chronic fatigue syndrome with respect to their pathophysiology and psychological aspects as well as therapeutic advancements. It is postulated that an interdisciplinary access to these patients early in the course of their illness involving both somatic medical and psychiatric competence is the most promising means to tackle this enormous medical and health protection problem. PMID- 10853375 TI - [Is presentation of symptoms enough for detection of an illness? On the correlation between symptoms, findings, diagnosis and disability]. AB - In 1999, the IV. civil division of the German Federal Supreme Court has issued a verdict which states that for some diseases the medical proof of a disease may be established solely on grounds of the individual complaints supporting the respective diagnosis. In the eyes of the psychiatrist, this statement is wrong and results from a confusion of diagnosis and disease. It also reveals a curious misconception of how psychiatric diagnoses, as instances of non-physical disorders, are established. Responsible for the confusion are the modern diagnostic manuals, which are misunderstood and thus misused as containing a list of diseases. However, the diagnostic manuals only intend to classify characteristic complexes of complaints and signs, without establishing them as diseases. The present paper explores the relationship between diagnosis and disease and explains how psychiatric diagnoses are made. PMID- 10853376 TI - [Physical treatment in pain therapy]. AB - The application of physical therapy for the treatment of chronic and recurrent pain, above all in the locomotor system, is indeed common, but its importance for pain therapy is often underestimated. The use of physical measures frequently does not follow scientific aspects. Their application without a concept and without regard to the functional and structural alteration as typical of pain in skin, connective tissue and muscles is to be criticized. A survey of the essential physical measures and their therapeutic effects is given and the principles of prescription are discussed. PMID- 10853377 TI - [Inguinal hernia: accident sequela?]. AB - The evaluation of an inguinal hernia as a result of an accident has to be newly discussed because of new radiologic tools like US, CT and MRI. Therefore an injury of the abdominal wall after an adequate trauma has to be proved. This can also be achieved by radiology, in which signs of injury, as contusion and hematoma, can be detected. Moreover the histological signs of injury and bleeding, i.e. fibrinexudation, capillary granulation tissue, round cells, siderophages, fibrocytes and fibroblast proliferation must be proved. PMID- 10853378 TI - [Legal aspects in early defibrillation by trained lay respondents]. AB - Ventricular fibrillation is the main reason for cardiac arrest. The probability for survival decreases by 10% every minute, therefore immediate resuscitation is necessary. Cardiopulmonary Resuscitation (CPR) by trained first responders is already established, when a doctor is not available. Today automated external defibrillators (AED) are available to first responders for an effective therapy of ventricular fibrillation. Thanks to the high reliability of today's automated external defibrillators they can be used by trained first responders without any legal reservations. If a defibrillator is available, a trained first responder is obliged to use it in an emergency. PMID- 10853380 TI - [In Process Citation] PMID- 10853379 TI - [Sequelae of genome analysis for insurance management]. PMID- 10853381 TI - [Comment on J. Fritze: Specialized departments for natural healing in the hospital--medically necessary?]. PMID- 10853382 TI - [Comment on J. Fritze: Specialized departments for natural healing in the hospital--medically necessary?]. PMID- 10853383 TI - [Comment on J. Fritze: Specialized departments for natural healing in the hospital--medically necessary?]. PMID- 10853384 TI - WHO Expert Committee on Biological Standardization. AB - This report presents the recommendations of a WHO Expert Committee commissioned to coordinate activities leading to the adoption of international requirements for the production and control of vaccines and other biologicals and the establishment of international biological reference materials. The report starts with a discussion of general issues brought to the Committee's attention and provides information on the status and development of reference materials for various antibodies, antibiotics, antigens, blood products and related substances, cytokines and growth factors and other substances for which the Committee has discerned a need for international reference materials. The second part of the report, of particular relevance of manufacturers and national control authorities, contains guidelines for the production and control of synthetic peptide vaccines, requirements for tick-borne encephalitis vaccine (inactivated), guidelines for thromboplastins and plasma used to control oral anticoagulant therapy, an amendment to the requirements for hepatitis B vaccine made by recombinant DNA techniques and a report on the standardization and calibration of cytokine immunoassays. PMID- 10853385 TI - Diamorphine, a pitfall for South African doctors practising in the UK. PMID- 10853386 TI - Doctor migration--the editor under fire again. PMID- 10853387 TI - Neonatal intensive care--the people have spoken. PMID- 10853388 TI - Abortion--ethical obligations. PMID- 10853389 TI - Calls to balance skewed health care provision. PMID- 10853390 TI - Manto's latest prescription--take 500 interns a year. PMID- 10853391 TI - How to stay sane in the crazy world of medicine. PMID- 10853392 TI - Legalities of the MSA (Part 2). PMID- 10853393 TI - Principles of treating HIV. PMID- 10853394 TI - The importance of high-density lipoprotein cholesterol in the management of cardiovascular risk. PMID- 10853395 TI - Health care at police stations. PMID- 10853396 TI - TB--are we losing the battle? PMID- 10853397 TI - Betting the whole arm. PMID- 10853398 TI - Serious consequences to misuse of propofol anaesthetic. PMID- 10853399 TI - Trauma, alcohol and other substances. PMID- 10853400 TI - The impact of service provision on contraceptive usage and immunisation coverage. PMID- 10853401 TI - Confirmation of transmission of the microsporidian parasite Enterocytozoon bieneusi in South Africa. PMID- 10853402 TI - Substance abuse and trauma in Cape Town. AB - OBJECTIVE: To obtain baseline data on the incidence of acute alcohol intoxication, chronic alcoholism and illicit drug usage among a cohort of injured patients. DESIGN: A prospective, descriptive study of 254 injured patients presenting at the trauma unit of Groote Schuur Hospital over an idealized week in 1997. Alcohol consumption was assessed by means of the Lion SD2 alcolmeter. Chronic alcoholism was assessed using the CAGE questionnaire. Each patient's urine was analysed for four drugs (cannabis, morphine, opiates and methaqualone) using conventional 'wet' analysis. Sweat was tested for cannabis using a Drugwipe. MAIN OUTCOME MEASURES: Socio-demographics, cause of injury, injury severity, acute alcohol intoxication, chronic alcohol usage and illicit drug involvement. RESULTS: Patients were predominantly male, coloured and an average of 31.3 years old. The majority had been injured as a result of interpersonal violence. Self-reported alcohol consumption was reliable but this was not so for self-reported drug usage. Sixty per cent of patients had positive alcohol levels on breath analysis. More than one-quarter of all the patients could be classified as chronic alcoholics on the CAGE questionnaire. On urine analysis, 40% of patients were found to have used at least one illicit drug in the recent past. The most commonly abused drugs were cannabis or a combination of cannabis and Mandrax, locally called a 'white pipe'. Use of the white pipe was confined almost exclusively to patients injured as a result of interpersonal violence. CONCLUSIONS: Alcohol remains the most commonly abused substance among trauma patients, but there are growing numbers of patients who simultaneously abuse illicit drugs. This study will be conducted annually to detect trends and identify emerging problems. PMID- 10853403 TI - Randomised controlled trial of the efficacy of misoprostol used as a cervical ripening agent prior to termination of pregnancy in the first trimester. AB - BACKGROUND: Misoprostol is being used increasingly in clinical practice for cervical ripening in first-trimester abortions, but because of lack of good evidence of its effectiveness, administration consensus has not been reached on dosage, route of administration, time of administration pre-operatively and gestational age group. In this study we tested the hypothesis that self administration of 600 micrograms vaginal misoprostol is feasible and when used 2 4 hours pre-operatively results in sufficient cervical dilatation to make suction curettage easier. METHODS: A double-blind, randomised, placebo-controlled trial was undertaken. Two hundred and seventy-eight women scheduled for termination of pregnancy of up to 12 weeks' duration by manual vacuum aspiration were assigned to receive either 600 micrograms misoprostol pre-operatively, or placebo. The achievement of 'satisfactory' (> or = 7 mm) baseline cervical dilatation after 2 4 hours was evaluated as the primary outcome. Secondary outcome measurements included ease and duration of the procedure. Side-effects such as pre-operative bleeding, gastro-intestinal complaints and pain as well as adverse events were noted in all cases. FINDINGS: Self-administration of vaginal misoprostol was successful in all women and 273 women were evaluated for main end-points. A significantly larger proportion of patients in the treatment group reached cervical dilatation of > or = 7 mm (67.3% v. 30.9%, P < 0.0001). The side-effects were minimal and comparable in the two groups. In the treatment group the mean procedure duration was significantly shorter (220 seconds v. 321 seconds, P = 0.0013) and the procedure was more likely to be rated by the operator as 'easy' (81.8% v. 63.3%, P = 0.0082). This resulted in a significant reduction in treatment failure in the < 70-day gestation group (5.0% v. 14.7%, P = 0.005). CONCLUSION: It is feasible, safe and effective for 600 micrograms misoprostol to be self-administered vaginally 2-4 hours pre-operatively for cervical priming prior to manual vacuum aspiration. Further research is needed to establish optimal use in the first trimester and to determine patient acceptance. PMID- 10853404 TI - Epilepsy in rural South African children--prevalence, associated disability and management. AB - OBJECTIVE: To determine the prevalence of epilepsy and its associated disabilities in rural South African children aged 2-9 years. SETTING: Eight villages in the district of Bushbuckridge, Northern Province, South Africa. DESIGN: A two-phase design was used. The first phase involved screening children on a house-to-house basis by interviewing mothers or caregivers using an internationally validated questionnaire for detecting childhood disability in developing countries. The second phase consisted of a paediatric/neurodevelopmental assessment of the children who screened positive. RESULTS: A total of 6,692 children were screened; 722 (10.8%) had a paediatric evaluation and 49 (0.73%) had epilepsy. The lifetime and active prevalences of epilepsy in these children were 7.3/1,000 and 6.7/1,000 respectively. Associated developmental disability was recorded in 35 affected children (71.4%), including 8 (16.3%) in whom this was moderate to severe. More than a half of the children with epilepsy (57.1%) did not receive anticonvulsant medication. CONCLUSION: The prevalence of epilepsy in the rural childhood population investigated is higher than that recorded in most similar studies from sub-Saharan Africa, and the poor utilisation of appropriate anticonvulsant treatment is cause for concern. This study highlights the paucity of relevant information on the epidemiology of epilepsy in South Africa and that the system available for its management, especially in rural areas, appears to have functional deficiencies. Appropriate research is needed to identify the problems in service delivery and to enable the planning and implementation of an appropriate primary health care-based system for the diagnosis and management of epilepsy in children. PMID- 10853405 TI - Epidemics of the Roman Empire, 27 BC-AD 476. PMID- 10853406 TI - Stroke therapy clinical guideline. South African Medical Association-- Neurological Association of South Africa Stroke Working Group. AB - OBJECTIVE: To describe the prevention, management and rehabilitation of stroke in South Africa as provided by a range of caregivers. OPTIONS: Emphasis should predominantly be on finding the cause, preventing, and treating stroke in the transient and mild stroke group. Moderate stroke patients require maximal rehabilitation and secondary prevention. Severe disabling stroke patients require home and community care. OUTCOMES: The most effective use of resources (personnel and facilities) in relation to the different types of stroke. These should be used to decrease stroke incidence, increase stroke awareness, improve acute stroke therapy, and improve access to rehabilitation. EVIDENCE: Based on international reports and research. Meta-analyses were used when the topic warranted such specific literature review. Stroke consensus has been achieved internationally and widely reported. A wide range of South African reviewers from various health fields, including representatives of local and national health groups, were consulted. Evidence-based documents were considered and incorporated. VALUES: As far as possible, cultural and economic preferences were given major emphasis. The aim of the guideline is to optimise stroke care from the point of view of individual patients, health practitioners, reimbursing agencies and society as a whole. Where possible, best-practice stroke care of international standard has been used in this document. However, at no stage has the Working Group deviated from the unique set of South African stroke problems, namely differing risk and preventive factors, multi-cultural, genetic and traditional practices and the problems of cost saving from best-practice funding for stroke care. BENEFITS, HARMS, COSTS: It is difficult to assess the local situation without prospective research. However, stroke causes the following in South Africa: between 8% and 10% of all reported deaths 7.5% of deaths in the workforce (25-64 years of age) age-standardised mortality rate of 125 175/100,000. More can be done to improve the cost-effectiveness of available resources by increasing education of health care professionals and patients. RECOMMENDATIONS: Assess and treat all cerebral vascular events within 6 hours if possible; categorize the stoke into mild, moderate or severe; transient ischaemic attack (TIA), minor or reversible stroke should be intensively investigated to find the cause and this should be treated vigorously; all risk factors should be treated; moderate strokes require active rehabilitation; secondary prevention is required to prevent another stroke; when vegetative symptoms persist after 7 days, therapy should be supportive. VALIDATION: This guideline is similar to others produced in other countries but with reference to South Africa, and has been developed in collaboration with the Neurological Association of South Africa. The initial document was developed in 1997 with a multidisciplinary group with special interest in the management of stroke. In July 1998 a nationally representative stroke consensus meeting was held and the document was widely scrutinised. The guideline is endorsed by the South African Medical Association. PMID- 10853407 TI - [Role of the p53 and bcl-2 genes in apoptosis and drug resistance of tumors]. PMID- 10853408 TI - [Prognosis of leukopenia at the early stages of radio- and chemotherapy for Hodgkin's disease]. PMID- 10853409 TI - [Melatonin inhibits large bowel carcinogenesis induced by 1,2-dimethylhydrazine in rats: effects and possible mechanisms]. PMID- 10853410 TI - [Establishing criteria for survival of oncology patients on a regional level]. PMID- 10853411 TI - [Radioimmunoassay of serum pepsinogen I in chronic gastritis and stomach cancer]. AB - Blood serum in stomach cancer and chronic gastritis has been compared. A sharp decrease in pepsinogen 1 level both in cancer and gastritis patients was found as compared with healthy subjects. Pepsinogen 1 level in poorly-differentiated tumor (37.4 ng/ml) was lower than in well-differentiated one (58.2 ng/ml). PMID- 10853412 TI - [MALT-lymphomas with primary gastric focus]. AB - The peculiar features of the clinical course and treatment of primary gastric MALT-lymphoma in 7 patients are discussed. Diagnosis was established by morphological examination of resected material. Six patients had low-grade tumor while one showed signs of blast-transformation. Stage IEB disease was identified in 4 cases, IIEB--3. Complete remission duration after combined surgical and chemoradiation treatment ranged 3.5-80 months (mean duration--35.1 months), survival--5-84 months (mean duration--43.1 months). Relapse was registered in one case 3.5 months after operation. PMID- 10853413 TI - [Adjuvant chemotherapy with anthracyclines in comparison with the standard combination of CMF in breast cancer with high risk of relapse]. AB - The effectiveness of adjuvant therapy with adriablastin and doxorubicin for breast cancer has been compared to that of standard CMF. During 1985-1990, the study included 349 patients with T1-2N2M0 and T3N0-2M0 tumors; mean age--46 yrs; mean follow-up--96.7 months. Overall survival rate in the doxorubicin group was 73%, CMF--62%; relapse-free survival--62.1 and 55%, respectively. The absolute difference in overall survival rates (11%) proved barely significant (p = 0.056). However, the difference in overall survival (p < 0.05) after anthracyclines and CMF in patients with tumors T1-2N2M0 and T3N1M0 was significant and in favor of the former. As far as frequency and degree of side-effects is concerned, their patterns were practically identical in both groups, except for the significantly higher frequency of cardiotoxity and complete alopecia in doxorubicin therapy. Cardiotoxic complication rate was significantly reduced from 13.8 to 3.9% by cardioxane treatment. PMID- 10853414 TI - [Radiotherapy for recurrent breast cancer]. AB - The study is concerned with the clinico-radiobiological characteristics of radiotherapy for relapsed breast cancer. Adequate choice of tissue mass to be exposed appeared much more important than any change in focal dose within 50-80 Gy, to achieve higher frequency of locoregional therapeutic effect. No significant noticeable relationship was established between efficacy of recurrence treatment, on the one hand, and such factors as cluster-like lesion, ulceration of tumor and additional excision of a relapsed node, on the other. However, recurrent tumors larger than 3 cm showed lower radiosensitivity involving a sharp rise in the likelihood of dissemination. Radiotherapy for primary tumor did not affect the radiosensitivity of recurrent malignancies but slowed down the rate of ies growth. Also, it might speed up tumor dissemination. PMID- 10853415 TI - [Hormonal status of patients with mastopathy]. AB - The investigation compared blood levels of certain hormones produced both by the pituitary and peripheral endocrine glands in cases of mastopathy and healthy subjects. No significant differences for either parameter under study were found. Changes in the ratios of certain hormones suggested disturbed regulation of the endocrine system in patients with mastopathy. PMID- 10853416 TI - [Papillomavirus infection in healthy women from St.Petersburg]. AB - The study was aimed to evaluate the features of human papillomavirus (HPV) infection in healthy female inhabitants of St. Petersburg. 309 attendants of gynecological clinics were recruited for the screening. All participants were of reproductive age and were lacking the clinical or morphological symptoms of HPV related diseases. PCR revealed HPV-specific sequences in 90 (29%) women. HPV type distribution was similar to the one in USA and Europe. HPV-positivity did not correlate with age of females, age at sexual debut, or total life-long duration of sexual activity. However, HPV were overrepresented among women reporting excessive number of contraceptive abortions. The data imply that an establishment of large-scale HPV-testing in Russia could be a desirable component of preventive anti-cancer programs. PMID- 10853417 TI - [Epidermal growth factor receptors and their ligands in endometrial carcinoma: correlation with clinico-morphologic factors and steroid receptors]. AB - Epidermal growth factor receptors (EGFR) were identified in the membrane fraction of tumor in 21 out of 58 (36%) endometrial cancer patients. EGFR ligands--EGF like peptides--were found in tumor cytosols in 36% of patients. However, EGFR and EGF-like peptides were observed in 7% of tumors only; there were no receptors or ligands in 39% of patients. Both EGFR and steroid receptors were expressed in 20 tumor patients. The frequency of EGFR expression in endometrial carcinomas was practically identical to that of normal endometrium at the proliferative stage of menstrual cycle and half that of hyperplastic endometrium. Postmenopausal patients tended to show a decrease in hormonal sensitivity of carcinoma and EGFR expression frequency rate and a lower degree of endometrial carcinoma cell differentiation. Also, a significant rise in the EGFR expression frequency and mean concentration was found in conjunction with a deeper invasion of tumor into the myometrium. PMID- 10853418 TI - [Investigation of nucleolar organizer region activity in epithelial cells of hyperplastic endometrial carcinoma tissue]. AB - Such characteristics of nucleolar organizer regions (NORs) as nucleolar and intra nucleolar grain counts, NOR mean area, mean total NOR area in nucleus, nuclear area and nucleolus/nucleus ratio were identified using vaginal aspirate-smears from 34 patients with histologically verified diagnosis (adenocarcinoma--10, atypical hyperplasia (ATH)--10, glandular endometrial hyperplasia (GEH)--4); silver-staining and micro-imaging analyzer techniques were employed. Atypical hyperplasia and nucleolar count in nucleus progressively and significantly increased in the following sequence: proliferative phase--GEH--adenocarcinoma. Mean and total area of NORs showed a significant rise in ATH and adenocarcinoma. Those indices of endometrial cell proliferation and GEH were identical. Nuclear area and nucleolus/nucleus ratio offered most information for differentiating GEH, ATH and carcinoma. A highly reliable correlation was established between intra-nucleolar grain count and microscopically-assayed silver concentration, on the one hand, and NOR area, on the other. PMID- 10853419 TI - [The nature of hyperinsulinemia (insulin resistance) in endometrial carcinoma: of plasma levels of insulin and c-peptide]. AB - Standard glucose-tolerance test (SGTT) was carried out in 73 patients with endometrial tumors. Elevated concentrations of plasma insulin and C-peptide were established in endometrial carcinoma patients (irrespective of age and reproductive status) after night fast and 120 min after SGTT start, as compared to healthy subjects and breast cancer patients. Obese (BMI index 28 kg/m2) reproductive endometrial carcinoma patients showed pronounced hyperinsulinemia and resistance to insulin. Menopausal patients with endometrial tumors (BMI index < = 28) were characterized by a much faster metabolic clearance of insulin, as compared with all other patients. Therefore, degree of insulin resistance in endometrial carcinoma is determined by both enhanced secretion of insulin and lowered metabolic clearance of this hormone which in turn is associated with obesity. PMID- 10853420 TI - [Distribution of Cs-137 in the human body with malignant neoplasms of various localizations]. AB - 137Cs-concentrations and distribution in internal organs and malignant tumor tissues from cadavers and patients operated on for cancer were identified by gamma spectrometry in clinics of Vitebsk Region. Non-uniform distribution patterns in different organs were registered, irrespective of the presence or absence of malignancy. PMID- 10853421 TI - [The MDR gene and cellular sensitivity to various effects]. AB - mdr-Transfected K-562 cells revealed a relatively high resistance to cytotoxic monokines and ionizing radiation as compared to parental cells. Taken together with what is known about the resistance of mdr-expressing cells to multiple cytotoxic drugs, our results point to malignant cells having universal mechanisms of chemo-, bio- and radioresistance. PMID- 10853422 TI - [The radiosensitizer AK-2123 enhances sensitivity of MDR-tumors to Mitomycin C]. AB - It was demonstrated that radiosensitizer AK-2123 of the triazole group significantly enhanced the sensitivity of MDR-strains of P388 murine leukemia (reported by the authors earlier) to mitomycin C (MMC). There was a direct correlation between the modulating effect of AK-2123 and dose increase from 1 to 10 mg/kg. The effect depended on the initial sensitivity of the MMC-resistant strain which in turn correlated with the absence or presence of sorcin (cytosole low-molecular Ca(2+)-binding protein) gene coamplification in the mdr-amplicon. Since AK-2123 was reported earlier by us to disrupt active Ca(2+)-transport, it is suggested that the modulating effect of the radiosensitizer was at least partially due to said disruption. AK-2123 exerted no antitumor action of its own whatsoever. It could neither modify the sensitivity of parent strain P388 to MMC, nor overcome the cross resistance of one of the MDR-tumor strains under study to such drugs as etoposide and adriablastin. PMID- 10853423 TI - [First experience with hypothermia in radiotherapy for locally advanced skin cancer of the face]. AB - An original method of cryotherapy is presented. After each exposure during standard fractionation regimen tumor was cooled to 0-5 deg C in 20 patients with locoregional skin cancer of the face. Complete remission was registered in 18 cases of good cosmetic effect. Recurrence was reported in 2 cases during 9 month- 10 year follow-up. PMID- 10853424 TI - [Intraoperative radiotherapy during plastic surgery for lung cancer]. AB - The results of 28 bronchoplastic operations for non-small cell lung cancer and 3 8 year follow-up have been evaluated. Intraoperative radiotherapy (IORT) (10 Gy) was conducted in 6 patients. IORT was used as a component of combined treatment with good results and without postoperative complications. PMID- 10853425 TI - [Role of transthoracic fine-needle aspiration in the diagnosis of peripheral lung cancer]. AB - The efficacy of transthoracic fine-needle aspiration guided by roentgenotelevision (86 cases) and computed tomography (106 cases) for peripheral lung cancer was studied. Special needles Nos. 18-20G (dia. 1-1.2 mm) and Nos. 22 25G (dia. 0.5-0.7 mm) were used depending on size and site of tumor. Roentgenotelevision imaging proved more informative for small-size tumors (less than 2 cm). Computed tomography offered more advantage in imaging medium- and large-size tumors because it allowed to avoid repeated aspiration and, therefore, reduce the risk of complications. PMID- 10853426 TI - [Differential diagnosis of chronic ulcer and ulcerated cancer of the stomach]. PMID- 10853427 TI - [Transcatheter treatment of hepatic metastasis from stomach cancer]. AB - The data on arterial chemo-infusion (14) and chemoembolization (10) were compared in the treatment of 24 patients with unresectable metastases of gastric cancer into the liver. Decrease in or stabilization of tumor growth were found in 80 and 50%, respectively. Mean survival after chemo-infusion was 15.8 months, chemoembolization--9.6 months (p = 0.06); one-year survival--71 and 44%, respectively. These results match the literature data on resection of the liver. Therapy should start with chemo-infusion and should not be followed by chemoembolization unless the former fails. PMID- 10853428 TI - [Effectiveness of combined resection and exenteration of the pelvic organs as part of the comprehensive treatment of extensive malignancies of the rectum and female genitals]. AB - The results of the surgical treatment of 865 patients with extended malignancies of the rectum and female genitals are presented. Combined resection and exenteration of organs of the small pelvis (both radical and palliative to ensure cytoreduction of tumor) were carried out in 695 cases (palliative surgery for symptoms--170). The number of resections and sphincter-saving operations has increased while the lethality rates have dropped in recent years. Three-year survival after combined radical surgery for rectal cancer was 59.1%; five-year survival--49% (palliation with removal of distant metastases--26.0 and 14.8%; without removal--24.2 and 0%, respectively; palliative surgery for symptoms--2.3 and 0%, respectively). In cases of palliative surgery for cytoreduction of tumor of the female genitals, 3- and 5-year survival after removal of all distant foci was 66.2 and 54.1%; partial cytoreduction--42.2 and 28.8%, and surgery for symptoms--13.8 and 13.8%, respectively. Cytoreduction improved both the chances and efficacy of adjuvant radio- and chemotherapy. It is suggested that surgery be included as a component of complex treatment of malignancies; combined cytoreduction is fully justified even if its effect is merely palliative. PMID- 10853429 TI - [The use of Oxadol in tablet form for the treatment of chronic pain syndrome in late-stage cancer patients]. AB - Chronic pain management with tablets of oxadol, a non-narcotic central-action analgetic, was studied in 30 patients, focusing on combinations with "basic" drugs of the opiate and opioid group. Combined administration of tramal and oxadol for moderate pain relief raised the analgesic effect and improved certain parameters of quality of life. Possible side-effects and indications in cases of well-advanced tumor were established. PMID- 10853430 TI - [Results of combined therapy for lymphogranulomatosis in children]. PMID- 10853431 TI - [Non-Hodgkin's mantle-cell lymphoma]. PMID- 10853432 TI - [Photodynamic therapy as a new radical therapeutic method in patients with relapsing tumors at "inaccessible" sites]. PMID- 10853433 TI - [The use of computer technology in the cytologic diagnosis of neoplasms]. PMID- 10853434 TI - Adnexal torsion. A clinicopathologic review of 31 cases. AB - OBJECTIVE: To evaluate the correlation between clinical parameters, intraoperative assessment of tissue viability and histopathology in adnexal torsion. STUDY DESIGN: A retrospective study of 31 patients undergoing surgical intervention for adnexal torsion was performed, and clinical and pathologic parameters were compared. RESULTS: Intraoperative evaluation of viability was confirmed in 10/12 cases. Impression of necrosis was confirmed in 18/19 cases. Fever was seen in 2/12 patients with viable adnexa and in 7/19 patients with necrosis. CONCLUSION: Intraoperative impression of tissue viability correlated well with histopathology. Fever was significantly associated with tissue necrosis. PMID- 10853435 TI - Evaluating vaginal pH. Accuracy of two commercial pH papers in comparison to a hand-held digital pH meter. AB - OBJECTIVE: To determine the accuracy, in a clinical setting, of two commercial pH papers compared to a hand-held digital pH meter. STUDY DESIGN: Vaginal specimens from 30 women, ages 17-40, both asymptomatic and symptomatic, pregnant and nonpregnant, were evaluated for vaginal pH using pHydrion paper, ColorpHast paper and a reference hand-held, battery-operated pH meter. Pearson product-moment correlation analysis and concordance correlation analysis were performed comparing each of the pH papers to the pH meter. RESULTS: Pearson product correlation coefficients suggested a strong correlation between the pH papers compared to the commercial pH meter; however, concordance correlation coefficients were fair (< 97%). If a pH cutoff of 4.5 had been used as one of the diagnostic tools for the evaluation of bacterial vaginosis (i.e., pH > 4.5), the ColorpHast paper would have resulted in a theoretical false negative rate of 21%, and the pHydrion paper would have resulted in a false negative rate of 24%. There were no false positives. Using a single pH readout of 5.0 could have resulted in a correct value, ranging from 3.85-6.15 pH units with pHydrion paper and a range of 4.32-5.68 using the ColorpHast pH paper. The accuracy of ColorpHast paper was better than that of pHydrion paper. A 1 SD range for the mean pH difference for pHydrion paper was 1.054-0.854 and for ColorpHast was 0.619-0.501. The correlation coefficient for the pHydrion paper was .87, and the correlation coefficient for ColorpHast paper was .88. CONCLUSION: Two pH papers had questionable accuracy in a clinical setting as compared to the hand-held, battery powered pH meter. There was a theoretical 24% false negative rate if a pH cutoff of 4.5 had been used for pHydrion paper and a 21% false negative rate for ColorpHast paper. Although correlation coefficients were 88%, concordance correlations were inadequate for both papers. PMID- 10853436 TI - Prognostic significance of diagnostic laparoscopy for spontaneous fertility. AB - OBJECTIVE: To determine the prognostic significance of laparoscopy results for fertility outcome. STUDY DESIGN: Consecutive patients undergoing hysterosalpingography and laparoscopy for subfertility in our department between May 1985 and November 1987 were identified from medical records. The impact of tubal occlusion, hydrosalpinx and adhesions as detected at laparoscopy was studied. Kaplan-Meier curves for the occurrence of spontaneous intrauterine pregnancy were constructed for patients without tubal pathology, with mild tubal pathology (unilateral pathology or adhesions) and with severe tubal pathology (bilateral pathology). Fecundity rate ratios (FRR) were calculated to express the association between findings at laparoscopy and the occurrence of spontaneous intrauterine pregnancy. RESULTS: Of the 200 cases that could be analyzed, 129 (65%) showed no tubal occlusion on laparoscopy, 40 (20%) had unilateral tubal occlusion, and 31 (15%) had bilateral tubal occlusion. Unilateral hydrosalpinx was present in 13 (7%) patients, whereas 19 (10%) patients had bilateral hydrosalpinx. Adjusted FRRs were 0.65 and 0.20 for unilateral and bilateral tubal occlusion, and 0.46 and 0.32 for unilateral and bilateral hydrosalpinx. Peritubal adhesions were detected in 43% of patients and seemed to have no prognostic significance. CONCLUSION: Severe tubal pathology detected at laparoscopy affects fertility prospects strongly. However, since spontaneous intrauterine pregnancies occurred even in patients with bilateral tubal occlusion at laparoscopy, this technique should not be considered the gold standard in the diagnosis of tubal infertility. PMID- 10853437 TI - Human sperm cryobanking. Use of modified liquid nitrogen vapor. AB - OBJECTIVE: To study a modified liquid nitrogen vapor cryopreservation technique for human spermatozoa. STUDY DESIGN: The freezing rate of the modified liquid nitrogen vapor method was controlled using thermocouple probes to simultaneously measure the temperature of both the liquid nitrogen vapor and semen specimen. Sperm cryosurvival and postthaw velocity were compared using a programmable biologic freezer and our modified vapor freezing procedure. RESULTS: There were no significant differences in postthaw sperm viability between the two methods. CONCLUSION: This modified vapor freezing technique permits controlled cooling that is efficient, rapid and inexpensive. PMID- 10853438 TI - Follicular fluid levels of vascular endothelial growth factor. Are they predictive markers for ovarian hyperstimulation syndrome? AB - OBJECTIVE: To determine the possible predictive role of vascular endothelial growth factor (VEGF) levels in the follicular fluid (FF) at the time of oocyte retrieval in the development of ovarian hyperstimulation syndrome (OHSS) and its possible origin. STUDY DESIGN: FF was obtained from 174 high-responder patients at the time of oocyte retrieval. The study group comprised 16 high-responder patients who developed early, severe OHSS and from whom serum and peritoneal fluid (PF) were obtained during the active phase of the syndrome. These women were compared to 16 high-responder patients who did not develop OHSS. An additional control group comprised 16 low-responder patients who also did not develop OHSS. The FF, serum and PF samples were tested for VEGF by enzymelined immunosorbent assay. RESULTS: No differences in the FF VEGF levels were found among the OHSS group (1,742.3 +/- 522.4 pg/mL), the high-responder group that did not develop OHSS (1,802.0 +/- 584.3 pg/mL) and the low-responder group (1,686.7 +/- 374.2 pg/mL). In the OHSS group, no differences were found between the serum and PF VEGF levels (247.3 +/- 31.4 and 642.9 +/- 328.3 pg/mL, respectively). No correlation was found between the FF concentrations of VEGF and the mean serum 17 beta estradiol levels or number of oocytes retrieved. CONCLUSION: We conclude that preovulatory FF levels should not serve as a possible predictive factor for development of OHSS. The increased capillary permeability found in OHSS may be due to its systemic effect. PMID- 10853439 TI - Laparoscopic management of adnexal masses in pregnant women. AB - OBJECTIVE: To report on 14 cases of adnexal masses in the second trimester of pregnancy that were managed with laparoscopic surgery. STUDY DESIGN: A retrospective study. During the period between January 1994 and January 1998, 14 women presented with adnexal masses in pregnancy and were surgically managed with laparoscopy. A retrospective chart review of these patients was used to determine factors, including gestational age, operating time, length of hospital stay, pathology results, pregnancy outcomes and complications. RESULTS: Fourteen patients had laparoscopic removal of adnexal masses in their second trimester of pregnancy. Average gestational age was 16 weeks (range, 11.5-21), average operating time was 84 minutes (range, 32-145), and average hospital stay was 2.0 days (range, 1-5). Pathology revealed 4 serous cystadenomas, 3 mucinous cystadenomas, 3 mature teratomas, 3 functional cysts and 1 bilateral endometrioma. There were no postoperative complications except for one case of mild peritonitis, which resolved with expectant management. There were no cases of preterm labor associated with the surgery. Ten pregnancies continued to term without complications and delivered average-sized infants. Three pregnancies were in the third trimester without complications at this writing. There was one intrauterine fetal death at 31 weeks; it was found to be secondary to an acute cord accident on autopsy remote from surgery. CONCLUSION: Significant ovarian masses are diagnosed relatively frequently in the pregnant woman. The risk of malignancy is low, but complications resulting from distention, rupture and/or torsion of the adnexa can be a significant concern. As laparoscopic procedures improve and our experience with laparoscopy in the pregnant woman increases, most of these patients can forego laparotomy and be managed safely by laparoscopic removal of the mass. This series outlines laparoscopic technique and outcomes after removal of significant adnexal masses in pregnancy. PMID- 10853440 TI - Predicting style-of-care preferences of obstetric patients. Medical vs. midwifery model. AB - OBJECTIVE: To identify patient populations preferring more medically oriented vs. more midwifery-oriented medical care in order to provide an institutional resourcing model. STUDY DESIGN: Questionnaires were distributed to pregnant women regarding possible concerns related to style of care and length of hospitalization for obstetric delivery. Responses from the 135 usable surveys were analyzed by cluster analysis to search for common demographic or ideologic concerns that might distinguish two or more groups of patients according to style of-care preferences. RESULTS: Four clusters were identified and categorized based on the initial hypothesis as midwifery preference, medical preference, passive and wanting both. Cluster membership could not be predicted by any demographic or historical variable studied, and the most significant style-of-care discriminator was related to preferred length of postpartum hospital stay, with 80% of the midwifery preference cluster desiring a short (< 24 hour) stay as compared to a single patient (2%) in the medical preference cluster. CONCLUSION: While obstetric care is often offered in distinctly stylized packages according to the training of the provider, our study suggests that patient preferences are much more complex and may contain many elements of both midwifery and medical styles of care. Broadening access to obstetric care will involve moving from our own preconceived notions of appropriate packaging into a patient-based and multi option setting for delivery of these services. PMID- 10853441 TI - p53 tumor suppressor gene expression in the mouse ovary during an artificially induced ovulatory cycle. AB - OBJECTIVE: To evaluate the expression of p53 in the mouse ovary during an artificially induced ovulatory cycle. STUDY DESIGN: Ovulation induction was performed using pregnant mares' serum gonadotropin/human chorionic gonadotropin (PMSG/hCG). First, a p53 promoter-chloramphenicol acetyl transferase (CAT) transgenic mouse model was used. Protein samples from ovaries of transgenic mice were assayed for CAT activity as evidence of p53 promoter activation. Next, RNA extracted from CD-1 mouse ovaries was used for reverse transcription/polymerase chain reaction (PCR) and northern blot analysis using a p53-specific probe. RESULTS: Increased CAT activity was noted in transgenic mice treated with PMSG/hCG as compared with controls. PCR studies on transgenic mice using primers for CAT and on CD-1 mice using primers for wild type p53 substantiated this observation. Furthermore, CAT assay and northern analysis, performed on samples obtained at serial time intervals from induction, indicated that maximal p53 expression occurs around the time of ovulation, beginning 48 hours after PMSG and peaking 6-12 hours after hCG administration. CONCLUSION: The temporal expression of p53 in the ovary during a PMSG/hCG artificially induced ovulatory cycle may indicate a role for p53 in processes of differentiation of granulosa cells into luteal cells. PMID- 10853442 TI - Effects of endothelin-1 and calcium channel blockers on contractions in human myometrium. A study on myometrial strips from normal and diabetic pregnant women. AB - OBJECTIVE: We studied the effect of endothelin (ET)-1 on spontaneous contractions and the effects of nimodipine and isradipine on ET-1-induced contractions in myometrial tissue from normal and diabetic pregnant women. STUDY DESIGN: We isolated myometrial strips from seven normal pregnant and seven gestational diabetic women undergoing elective cesarean section at term. Three sets of experimental studies were performed with three myometrial strips obtained from every woman and mounted in organ baths for recording of isometric tension. In the first set, the effect of increasing concentrations of ET-1 (10(-11)-10(-8) M) on spontaneous contractions was recorded. In the second and third sets, effects of increasing concentration of nimodipine (10(-6)-3.10(-5) M) and isradipine (10(-5) 3.10(-4) M), respectively, on contractions following pretreatment with 10(-8) M ET-1 were recorded. RESULTS: ET-1, beginning from 10(-9) M and 10(-10) M, significantly increased the amplitude of contractions in normal and diabetic strips, respectively. ET-1, beginning from 10(-9) M, also increased the frequency of contractions in normal and diabetic strips. The amplitude of contractions was significantly higher in diabetic strips as compared with normal strips at 10(-9) and 10(-8) M. There was no significant difference in the frequency of contractions between normal and diabetic strips. ET-1 at 10(-8) M also increased the basal tone of all normal and diabetic strips. Nimodipine, beginning from 10( 6) M and 3.10(-6) M, and isradipine, beginning from 10(-5) M and 3.10(-5) M, significantly decreased the amplitude of contractions in normal and diabetic strips, respectively. Nimodipine at 10(-5) M and 3.10(-5) M and isradipine at 3.10(-4) M significantly decreased the frequency of contractions in normal strips. Nimodipine, except at 3.10(-5) M, and isradipine did not significantly decrease the frequency of contractions in diabetic strips. CONCLUSION: Gestational diabetes increases ET-1-induced contractile response in human myometrium. The contractile effect of ET-1 in normal and diabetic myometrium is mediated partly by dihydropyridine-sensitive calcium channels since it is significantly reduced by nimodipine and isradipine. The promising data from this study warrant clinical studies on the definitive place of nimodipine and isradipine in the treatment of preterm labor, especially in a diabetic woman, to avoid metabolic complications of beta-mimetic tocolytics. PMID- 10853443 TI - Postpartum hemorrhage and intrauterine balloon tamponade. A report of three cases. AB - BACKGROUND: Postpartum hemorrhage can become rapidly catastrophic. If medical management fails, then, according to recent reports, the use of an intrauterine inflated Foley catheter balloon for tamponade gives excellent results and can help avoid invasive procedures. CASE: We present one case of profuse hemorrhage following evacuation of the fetus after intrauterine fetal death at 17 weeks' gestation controlled with intrauterine balloon tamponade and two cases of severe postpartum hemorrhage (one immediate and one late) following normal vaginal deliveries, both controlled with Foley catheters. In either case the patient required no blood transfusions, and major surgery was avoided. CONCLUSION: Intrauterine balloon tamponade is highly effective. The catheter is readily available, is not expensive, does not require special training for insertion and, extremely important, can avoid major surgery. PMID- 10853444 TI - Pregnancy complicated by traumatic diaphragmatic rupture. A case report. AB - BACKGROUND: Diaphragmatic rupture may be easily overlooked at the time of multiple trauma. Occult diaphragmatic rupture may first manifest during pregnancy as severe dyspnea. CASE: A parous woman who had sustained multiple traumatic injuries prior to pregnancy presented in midtrimester with abdominal pain and dyspnea. Chest roentgenography and computed tomography revealed bowel in the left hemithorax, compatible with a left-sided diaphragmatic rupture. Surgical correction was indicated secondary to the symptomatic nature of the presentation. CONCLUSION: Diaphragmatic rupture may be occult and may first present during a subsequent pregnancy. Surgical therapy is the cornerstone of management when a diaphragmatic defect is symptomatic. The route of delivery may be individualized for patients with diaphragmatic repairs in whom there has been sufficient time for healing. PMID- 10853445 TI - Metastatic complete hydatidiform mole with a surviving coexistent twin. A case report. AB - BACKGROUND: Complete hydatidiform molar pregnancy with a coexistent live fetus is a rare clinical entity with poor fetal survival and an uncertain maternal risk. CASE: A twin gestation consisting of a complete molar pregnancy as one twin and a normal second "twin" was identified in the second trimester. The patient was managed expectantly until 28 weeks' gestation, at which time evidence of metastatic trophoblastic disease was identified. She delivered a healthy infant by cesarean section and received postoperative single-agent chemotherapy. She was disease free six months after treatment. CONCLUSION: Patients with documented complete molar pregnancies and coexistent live fetuses present several complex clinical issues. Though successful in some instances, expectant management should be used with caution in these patients. PMID- 10853446 TI - Persistent mullerian duct syndrome. A case report. AB - BACKGROUND: Persistent mullerian duct syndrome is a rare form of male pseudohermaphroditism in which well-developed mullerian structures are present in an otherwise normal male with XY chromosomes. The syndrome was first described by Nilson in 1939, and almost 100 cases have been reported. CASE: A 22-year-old man presented with mild, right-sided inguinal pain and heaviness in his scrotum. He underwent right-sided inguinal exploration because of having a palpable right sided, irreducible inguinal hernia. Two testicles with surface nodularity and a bicornuate uterus with rudimentary fallopian tubes were detected and removed as one specimen, and the hernia was repaired. CONCLUSION: Management of this syndrome is difficult because of the limited number of cases. If the diagnosis can be made before surgery, karyotyping can be useful to decide on orchiopexy or orchioectomy. In suspected cases, laparoscopy and ultrasonographic evaluation of all crytorchidic cases may be helpful for diagnosing this condition before surgery. All patients with this syndrome have a male phenotype; therefore, it is essential to preserve secondary sex characteristics. Androgen replacement therapy should be given to patients who have undergone gonadectomy and to those with low levels of testosterone. PMID- 10853447 TI - Induction of recombination by the bacterial RecA protein depends on the stability of the RecA-DNA complex. PMID- 10853448 TI - Accumulation of proteins immunochemically related to dehydrins in mitochondria of plants exposed to low temperature. PMID- 10853449 TI - Relationship between cold resistance of plants and the lipid content of their chloroplast membranes. PMID- 10853450 TI - The polymorphic element of the Epstein-Barr virus EBNA-3C gene: a preliminary characteristic and the use for distinguishing between mono- and polyclonal infected cells. PMID- 10853451 TI - Acetylcholine content and the localization of acetylcholine esterase activity in different tissues of magnetic-orientation types of radish. PMID- 10853452 TI - Detection and characterization of extended deletions in cosmid clones of ribosomal DNA of human chromosome 13. PMID- 10853453 TI - The structure-property correlation: estimates of the half-clearance time of perfluororganic compounds. PMID- 10853454 TI - Overexpression of bacterial RecA protein in somatic mammalian cells increases the frequency of gene targeting. PMID- 10853455 TI - How cancer centers manage quality of care. PMID- 10853456 TI - New strategies for managing metastatic breast cancer. AB - In 1999, metastatic breast cancer claimed the lives of almost 45,000 women. For the vast majority of patients, metastatic breast cancer is an incurable disease with a median survival of only 2 to 3 years after diagnosis. The major goal of therapy is palliation. New endocrine agents developed during the last several years provide a greater opportunity for response in receptor-positive patients. New chemotherapeutic options have led to the reemergence of single-agent therapy as an effective palliative approach. Clinical trials remain the mainstay of cutting-edge therapy for metastatic breast cancer and should always be considered, if available. This review will focus on major issues in the treatment of metastatic breast cancer, including new endocrine and chemotherapeutic agents and a suggested strategy for patient management. PMID- 10853457 TI - FDA approves irinotecan as first-line therapy for colorectal cancer. PMID- 10853458 TI - New indication sought for bicalutamide. PMID- 10853459 TI - Management of infections in patients with acute leukemia. AB - Several recent studies have addressed the management of infectious problems in patients with acute leukemia. Although those studies have served to emphasize the fundamental management principles formulated and proven almost 30 years ago, they have also contributed important new insights. This article describes recent developments in the management of infectious illnesses in patients who are neutropenic due to leukemia or its treatment. The discussion will focus on the increasing armamentarium of antimicrobial drugs and adjunctive agents. These expanding therapeutic options must be viewed in the context of newly emerging resistant organisms and special problems, such as the increased use of indwelling venous catheters. PMID- 10853460 TI - Clinical trials referral resource. Current clinical trials in epithelial ovarian cancer. PMID- 10853461 TI - An alternative algorithm for dosing transdermal fentanyl for cancer-related pain. AB - Many cancer patients are undermedicated and inappropriately managed for pain, leading to a diminished quality of life. Patients with moderate to severe pain often require opioid analgesics. Recently published guidelines emphasize individualization of opioid treatment to provide the drug and route of administration that meet the needs of the particular patient. Intolerable side effects, ineffective pain relief, or a change in the patient's clinical status can dictate the need for a new pain management regimen. Physicians must be able to readily quantify relative analgesic potency when converting from one opioid to another or from one route of administration to another. Transdermal fentanyl (Duragesic) is an opioid agonist that has been shown to be safe and effective for the treatment of cancer pain. However, clinicians should realize that the manufacturer's recommendations for equianalgesic dosing of transdermal fentanyl may result in initial doses that are too low in some patients, and in a titration period that is too long. Under these circumstances, the patient is likely to experience unrelieved pain. An alternative dosing algorithm that considers both a review of the literature and our combined clinical experience with transdermal fentanyl should help clinicians individualize the treatment of pain. PMID- 10853462 TI - Intravesical therapy for superficial bladder cancer. AB - Approximately 54,400 new cases of transitional cell carcinoma of the bladder were reported in the United States in 1999, with an estimated 12,500 deaths attributable to this cancer. Close to 75% of all bladder tumors are confined to the urothelium (stage Ta, or carcinoma in situ), and nearly 30% of papillary tumors invade the lamina propria (stage T1). The majority of superficial tumors are low grade with low rates of progression. Transurethral resection is the standard initial treatment for transitional cell carcinoma. Intravesical therapy is an important adjunct to transurethral resection in patients with superficial bladder cancer, many of whom are at risk for disease recurrence and progression. Cytotoxic and immunomodulating agents and, more recently, photosensitizers have demonstrated utility against superficial transitional cell carcinoma. Many studies have assessed and continue to examine the efficacy of various agents at different doses and in different combinations and schedules. Recently, valrubicin (Valstar) won Food and Drug Administration (FDA) approval only for the treatment of refractory carcinoma in situ. However, bacillus Calmette-Guerin (BCG) and mitomycin (Mutamycin) remain the most commonly used, most effective agents available for prophylaxis against recurrence and subsequent progression of superficial bladder cancer. This article reviews traditional and alternative intravesical agents useful in the therapy and prophylaxis of superficial transitional cell carcinoma of the bladder. PMID- 10853463 TI - Biological basis of radiation sensitivity. Part 2: Cellular and molecular determinants of radiosensitivity. AB - Recent studies have elucidated some of the molecular and cellular mechanisms that determine the sensitivity or resistance to ionizing radiation. These findings ultimately may be useful in devising new strategies to improve the therapeutic ratio in cancer treatment. Despite the rapid advances in knowledge of cellular functions that affect radiosensitivity, we still cannot account for most of the clinically observed heterogeneity of normal tissue and tumor responses to radiotherapy, nor can we accurately predict which individual tumors will be controlled locally and which patients will develop more severe normal tissue damage after radiotherapy. However, several candidate genes for which deletion or loss of function mutations may be associated with altered cellular radiosensitivity (e.g., ATM, p53, BRCA1, BRCA2, DNA-PK) have been identified. Some of the differences in normal tissue sensitivity to radiation may stem from mutations with milder effects, heterozygosity, or polymorphisms of these genes. Finally, molecular mechanisms linking genetic instability, radiosensitivity, and predisposition to cancer are being unraveled. PMID- 10853464 TI - Exisulind shows positive results in the prevention of precancerous colon polyps. PMID- 10853465 TI - [Pulmonary tuberculosis as a re-emergent infection]. PMID- 10853466 TI - [Changes and problems related to tuberculosis epidemics]. PMID- 10853467 TI - [Prevention and treatment of tuberculosis]. PMID- 10853468 TI - [Immunological status of tuberculosis]. PMID- 10853469 TI - [Interpretation of the results of tests for identification of Mycobacterium tuberculosis]. PMID- 10853471 TI - [Tuberculin reaction]. PMID- 10853470 TI - [Imaging diagnosis of tuberculosis and the key points related to the procedures]. PMID- 10853472 TI - [Diagnostic significance of pleural effusion and pleural biopsy]. PMID- 10853473 TI - [Pulmonary tuberculosis in primary care]. PMID- 10853474 TI - [Extra-pulmonary tuberculosis]. PMID- 10853475 TI - [Tuberculous infection and non-periodic screening]. PMID- 10853476 TI - [Treatment of tuberculosis and cooperation between hospitals and public health clinics]. PMID- 10853477 TI - [Guideline for chemotherapy of tuberculosis]. PMID- 10853478 TI - [DOTS (directly observed treatment, short-course)]. PMID- 10853479 TI - [Significance of BCG vaccination]. PMID- 10853480 TI - [Treatment of multidrug-resistant tuberculosis]. PMID- 10853481 TI - [Treatment of tuberculosis of the aged]. PMID- 10853482 TI - [Tuberculosis prevention]. PMID- 10853483 TI - [Current status and management of mass outbreaks and hospital infections]. PMID- 10853484 TI - [Health occupations and tuberculosis-related education]. PMID- 10853485 TI - [Modern concept of tuberculosis management (discussion]. PMID- 10853486 TI - [Pyogenic vertebral osteomyelitis caused by Gemella haemolysans]. PMID- 10853487 TI - [Isolated ACTH deficiency with severe muscle atrophy]. PMID- 10853488 TI - [Identical female twins diagnosed with type Ia glycogen storage disease in adulthood]. PMID- 10853489 TI - [Protein-losing enteropathy caused by ileal strongyloidiasis]. PMID- 10853490 TI - [CD8(+)-T-cell-rich B-cell lymphoma presenting in skin and bone]. PMID- 10853491 TI - [Current topics on inhalation therapy for obstructive pulmonary diseases]. PMID- 10853492 TI - [Primary biliary cirrhosis (PBC) and related diseases]. PMID- 10853494 TI - [Non-surgical approach to neurogenic bladder]. PMID- 10853493 TI - [Current status and overview on cord blood transplantation]. PMID- 10853495 TI - [Flu and emergencies in the year 2000]. PMID- 10853496 TI - [Diagnostic consistency between primary care and specialized care following emergency consultation]. AB - OBJECTIVE: To evaluate the diagnostic concordance between primary health-care level and hospital health-care level after emergency visits. DESIGN: Cross sectional study. SETTING: Health-care area 7 in Madrid. PARTICIPANTS: Any patient studied in the primary health-care level and further sent by referral request to the emergencies of the health-care area reference hospital. MEASUREMENTS AND RESULTS: The sample size was estimated according to a confidence level of 95%, a precision level of 5%, a concordance level of 50% and a 30% of referral requests without diagnosis. All diagnosis were codified by the ICD-9 CM. Single kappa index for each diagnosis and global kappa index were calculated. 559 patients were studied. 447 (80%) of the patients were referred with diagnosis and 112 (20%) without it. Kappa index was very high (> or = 0.8) for the following diagnosis: angina pectoris, and urticaria. Kappa index was high (0.6 > or = k < 0.8) for stroke, and deep venous thrombosis. It was moderate (0.4 > or = k < 0.6) for pneumonia, heart failure and heart attack. Kappa index was low (0.2 > or = k < 0.4) for appendicitis, and arthritis, and it was very low (< 0.2) for meningitis, and cellulitis. Global kappa index was 0.65 (95% CI, 0.58-0.72). CONCLUSIONS: Global concordance was high. The highest concordance was obtained for diseases with clinical diagnosis. Most of diseases with low and very low concordance are diseases that need specialized clinical tests. PMID- 10853497 TI - [Home care of patients with chronic disease: full physical and cognitive assessment and falls over 3 years of follow-up]. AB - OBJECTIVE: To assess the physical and cognitive capacity of chronically ill homebound patients, and the falls they suffered during three years of monitoring. DESIGN: Descriptive, longitudinal study. SETTING: "Raval Nord" Health District, Barcelona. PATIENTS AND OTHER PARTICIPANTS: All the 243 homebound chronic patients registered in the home care programme in May 1996 (67% women, average age 84). MEASUREMENTS AND MAIN RESULTS: After three years 16% had gone into an old people's home, 9% had moved house and 38% had died. The probability of not continuing in the programme after three years monitoring was related to less autonomy, presence of comorbidity, and worse cognitive capacity (p < 0.05). Of the 90 patients (37%) who remained active in May 1999, 41% showed disorders on the Short Portable Mental Status Questionnaire (SPMSQ), with a significant relationship to greater age, less autonomy and the presence of comorbidity. Numerous alterations in analysis (21.6%) and linked illnesses (18.9%) were found in the patients with cognitive deterioration. 42% of the patients active in May 99 had fallen during the monitoring period. 10% of the falls involved fractures. The number of falls was higher when there was visual-auditory loss, consumption of psychiatric drugs or absence of use of orthopaedic aids. There was also a greater probability of falls in patients who only had a part-time carer (p < 0.05). CONCLUSIONS: It is important to assess the autonomy, cognitive capacity and comorbidity of home-bound chronic patients when monitoring them. Likewise, cognitive disorders and falls must be properly weighed, as they are common in this class of patient. PMID- 10853498 TI - [Quality of antihypertensive drug prescription in a health area]. AB - OBJECTIVE: To find the compliance with previously established criteria on the quality of prescription of medication for hypertension. DESIGN: Retrospective and concurrent evaluation study of scientific and technical quality, with processing data, using as data source the clinical history. SETTING: Primary care teams in a Madrid Health Area. PARTICIPANTS: 873 clinical histories of hyper-intense patients in treatment with diuretics, beta-blockers, ACE inhibitors and/or calcium antagonists were chosen through systematic probabilistic sampling with a randomised start. MEASUREMENTS AND MAIN RESULTS: Data on age, sex, recording of treatment, linked pathologies and situations conditioning the choice of medicine were gathered. Information on the defined use criteria of the various pharmacological groups was also collected. 1145 drugs were used on 873 patients. Most common were the thiazide diuretics (36%), followed by ACE inhibitors (34.4%), calcium antagonists (21%) and beta-blockers (8.6%). 72% of the patients were undergoing one single therapy. 89.7% of the cases (95% CI, 87.43-91.59) had the treatment correctly recorded in the clinical record. Of the 721 hyperintense patients over 59 years old, 70.3% (95% CI, 66.81-73.60) fitted the defined criterion for use of diuretics. 48.7% fitted the ACE inhibitor criteria defined (CI, 43.71-53.78); 85.7% the beta-blocker criteria (CI, 76.85-91.69); and 58.7% the calcium antagonist criteria (95% CI, 52.17-64.9). CONCLUSIONS: The fit of the use of diuretics with the defined quality criterion is acceptable, while in the cases of ACE inhibitors and calcium antagonists the quality of prescription could be improved, while the use of beta-blockers is minimal. PMID- 10853499 TI - [Analysis of a program of early breast cancer detection in a rural area]. AB - OBJECTIVE: To find the results of a breast cancer early-detection programme run in a rural area, in terms of the activities corresponding to primary care. DESIGN: Cross-sectional, observational study. SETTING: Rural district in Primary Care Area 11, Madrid. The programme was run in this area between 1st February and 31st March 1999. PATIENTS: Women from 50 to 64, both inclusive, who had been called for a mammography. INTERVENTIONS: Inclusion in the breast cancer early detection programme and open telephone survey of those who did not take part to find the reasons for not taking part in the randomised sample. MEASUREMENTS AND MAIN RESULTS: In the period mentioned, 3902 women were called for screening (60.07% of all women between 50 and 64. The rest will be called in a second round in the year 2000). 2099 women attended for mammography (participation index 53.79%), with the following results: 5 (0.24%) highly likely to be malignant; 6 (0.29%) probably malignant; 172 (8.19%) probably benign; 1393 (66.36%) found to be benign; 438 (20.87%) negative; and 85 (4.05%) who needed further evaluation. The main reasons why 46.21% of the women called did not attend for mammography were: they had had one recently (29.9%); could not attend (22.7%); did not want to have one (17.5%); had not received the notification (17.5%). CONCLUSIONS: The participation rate was acceptable, with fewer malignant cases found than in other programmes. It would be important to call again those women who could not attend. PMID- 10853500 TI - [Anorexic behavior in a population of high-school students of a health area]. AB - OBJECTIVES: To calculate the prevalence of eating attitudes which determine eating disorders and their relationship to social, personal and cultural variables. DESIGN: Prevalence study. PARTICIPANTS: Centres of secondary education. Gijon Health Area (Asturias). Secondary school students (n = 17,000) selected by multi-stage stratified sampling. MEASUREMENTS AND MAIN RESULTS: The self-filled questionnaire included: social and personal variables, and Eating Attitudes Test 26 (anorexic conduct defined as scores = 20). Women also filled in the "Questionnaire on Influences on the Aesthetic Model of the Body" (CIMEC-26). There were 860 valid questionnaires, with 50% women. There was 12.8% prevalence of anorexic attitudes among women (95% CI, 9-16.5), and 1.8% among men (CI, 0.8 2.8). In the group with anorexic attitudes, 87.3% were women, with mean age 16.4; 88.3% lived in a city; 84% were in middle and middle-to-low social classes; 92% were studying their bachillerato; 28.5% attended private schools; 27% undertook activities related to having a thin body; 18.3% had separated parents; 27% had mothers working outside the home; 39.7% saw themselves as fat; and 81% wished to slim. The following variables showed statistically significant differences with the normal population: sex (OR = 7.7; 95% CI, 4.5-13.4), separated parents (OR = 1.9; CI, 1.4-2.8), undertaking activities relating to having a thin body (OR = 3.7; CI, 2.7-5.2); thinking oneself fat (OR = 4.7; CI, 3.1-7.1) and wishing to slim (OR = 7.2; CI, 4.6-11.2). 94.5% of women with disordered conduct showed a disorder on the CIMEC-26. CONCLUSIONS: There is a high prevalence of eating habits similar to those of patients with anorexia nervosa, which are related to the following variables: being a woman, having separated parents, seeing oneself as fat, desiring to slim and undertaking activities related to having a thin body. We observed no significant differences with the normal population in other social and personal variables. The socially imposed aesthetic model of the body determines anorexic conduct in women. PMID- 10853501 TI - [Improper use of medications with alcohol at a rural office]. AB - OBJECTIVES: To determine the prevalence of the joint use of alcohol and medication with the risk of interaction with alcohol, the social and personal features of patients most likely to use the two combined, and the extent of anti alcohol medical counselling of these patients. DESIGN: Prevalence study. SETTING: Primary care. Rural local clinic at Trigueros (Huelva). PARTICIPANTS: 581 histories of patients over 14, started between October 1993 and December 1996. MEASUREMENTS AND MAIN RESULTS: From the clinical histories we obtained the social and personal features, toxic habits, type and number of medicines prescribed, and whether or not there was medical counselling on alcohol consumption. Potential interactions were looked for through the list provided by Mengual Sandra and Gila Azanedo. 10.15% (95% CI, 7.8-12.9) of the subjects consumed at the same time medication with risk of interaction and alcohol. Males (OR = 7.1), over-65s (OR = 4.9), married people (OR = 3.1) and smokers (OR = 2.3) were more likely to combine alcohol and at-risk medication. 6.8% of the patients with potential risk of interaction were counselled against alcohol. CONCLUSIONS: The simultaneous use of alcohol (as drinks or a component of medicine) and at-risk drugs is a common reality in primary care. Men, the over-65s, smokers and married people are at greater risk. Medical counselling of patients at potential risk of interaction is quite rare, although it is greater than of drinkers who do not take at-risk medicines. PMID- 10853502 TI - [Job satisfaction in primary health care nursing in the counties of Alt and Baix Emporda]. AB - OBJECTIVES: To determine the labour satisfaction of primary care nursing professionals and find the related associated factors. DESIGN: Cross-sectional, descriptive, observation study. SETTING: Primary care in the counties of the Alt and Baix Emporda. PARTICIPANTS: All those with nursing diplomas who work in the primary care centres of this geographical area (131 professionals). MEASUREMENTS AND MAIN RESULTS: A self-administered questionnaire had a 79.3% response rate. The instrument had 9 dimensions on work satisfaction. The statistical analysis employed univariate and bivariate descriptive techniques (p = 0.05). The general lack of satisfaction was medium (2.18 +/- 0.45). The dimension with the highest score was "professional competence" (1.72 +/- 0.44), and with the lowest was "professional promotion" (2.80 +/- 0.89). The greatest number of significant differences occurred between the two classes of primary care, reformed and unreformed. CONCLUSIONS: The conditions in which professional nurses work in non reformed primary care causes lack of satisfaction in their work post, in its content and in their professional promotion. PMID- 10853503 TI - [Assessment of corticoid local injections at a health care center]. AB - OBJECTIVE: To evaluate the effect of corticosteroid infiltrations in osteoarticular and tendinous pathology in Primary Care. DESIGN: Observational and descriptive study. SETTING: La Union Health Center, a town of 14,000 inhabitants in Murcia. MEASUREMENTS AND MAIN RESULTS: All patients older than 14 years subjected to one or more infiltrations from 1-1-1997 to 1-5-1998 were included. The following variables were analysed using information obtained from the patient's medical records: age, sex, condition treated, evolution time, previous treatment, number of infiltrations, results and complications. Descriptive statistics including frequency and distribution tables were calculated. The 70.3% of the 138 cases are women, the mean age is 57.7 years. Most infiltrations were performed for shoulder conditions (60.1%). The mean time of evolution in these cases was 90.9 +/- 13.9 days. Only 10.9% of patients hadn't received any previous treatment. The mean number of infiltrations is 1.5 +/- 0.7 and the middle is 1. Symptoms improved in 82.62% of the cases, just completely (40.6%) or partially (42%). Only in 17.4% cases failed the intervention. Only a complication was registered, one case of residual pigmentation. CONCLUSIONS: The local corticosteroids infiltration is a useful technique in primary care because of a high effectivity, easy management, low cost and few complications. PMID- 10853504 TI - [Acute precordial pain: 100 cases in 3 years]. AB - OBJECTIVE: To find the effectiveness of diagnoses of acute precordial pain seen as an emergency at our centre. DESIGN: Observational, descriptive and retrospective study. SETTING: Urban primary care centre. PATIENTS: The 100 most recent patients who attended as an emergency with their first episode of acute precordial pain were included. STUDY PERIOD: December 1994 to March 1998. Home visits, patients without medical records and those seen on repeated attendance for precordialgia were excluded. MEASUREMENTS AND MAIN RESULTS: The emergency diagnosis and the diagnosis recorded afterwards in the clinical history of 100 people with acute precordialgia, aged 54.9 (16.7 years; 56% [n = 56] women), were gathered. Ischaemic cardiopathy (41%, n = 41) and mechanical precordialgia (36%, n = 36) were the most common initial diagnoses. We found 66.6% sensitivity and 81.4% specificity in the detection of ischaemic cardiopathy. The proportion of diagnostic errors was not linked to the pathological history of anxiety, ischaemic cardiopathy or oesophageal disease. CONCLUSIONS: 41% of precordialgias are diagnosed as presumably ischaemic and are potentially serious, although only 50% of them are confirmed as such. Our sensitivity in their diagnosis is comparable to that of other studies. PMID- 10853505 TI - [The registered population and its characteristics as adjustment element for individualized pharmacy budget allocation]. AB - OBJECTIVE: To find an equation that can calculate quality reference standards on the sum expected per doctor, adjusted for the size of his/her list, his/her characteristics and patients included in chronically ill services. This objective is posed within a strategy of individualizing information and evaluation as a tool to promote improvement. DESIGN: Crossover, descriptive study. SETTING: Autonomous community of the Canary Islands. Tenerife Health Area, primary care management. PARTICIPANTS: The study was performed with 51 doctors belonging to 10 primary care teams, for whom there was quality information available due to their having used the CRONOS computer programme for a sufficient period of time. The period analyzed ran from January to November 1997. MEASUREMENTS AND MAIN RESULTS: Multiple regression analysis through automatic algorithm of step-by-step selection of variables. There was a close relationship between the variability of the pharmacy sum per doctor (adjusted r2: 0.81; F, 66.05; p < 0.000) and the size of the list; also between the former and the characteristics of the users on the list. The most relevant variables were the percentage of women over 55 and the mean drug consumption per consultation with patients during the study period. It was seen, in relation to this last variable, that the percentage of pensioners and the number of hyper-intense patients included were important. CONCLUSION: Given the high variability explained, we propose broadening the study to all the doctors, so as to give the results greater consistency and offer quality reference information, in line with the above-mentioned improvement plan. PMID- 10853506 TI - [Qualitative research in primary care. Experience with open interviews]. PMID- 10853507 TI - [In Process Citation] PMID- 10853508 TI - [What do patients think of our health centers? Would they change anything? The situation at the beginning of the 21st century]. PMID- 10853509 TI - [Pneumology diagnostic consistency in the Bierzo Health Area]. PMID- 10853510 TI - [Lymphocytic colitis as a cause of chronic diarrhea: possible association with carbamazepine]. PMID- 10853511 TI - [All that glitters is not gold]. PMID- 10853512 TI - [Without the day-after island]. PMID- 10853513 TI - Introduction: why study age integration? PMID- 10853514 TI - Age integration: conceptual and historical background. PMID- 10853515 TI - Cultural and economic transfers between generations: one aspect of age integration. PMID- 10853516 TI - Age integration or age conflict as society ages? PMID- 10853517 TI - Integration of old and young. PMID- 10853518 TI - Age integration through interest mediation: political parties and unions. PMID- 10853519 TI - Paradox of opportunity: education, work, and age integration in the United States and Germany. PMID- 10853520 TI - The future of age integration in employment. PMID- 10853521 TI - Age integration as a solution to work-family conflict. PMID- 10853522 TI - Age integration in Europe: increasing or decreasing? PMID- 10853523 TI - Conflicting trends in The Netherlands. PMID- 10853524 TI - Public policy and the construction of old age in Europe. PMID- 10853525 TI - Predicting turnover and retention in nursing home administrators: management and policy implications. AB - Administrator turnover and its impact on the quality of patient care are important concerns in the nursing home industry. This study evaluates a model to determine which factors, attitudes, and personal characteristics can predict tenure. Responses to a survey from 290 nursing home administrators (NHAs) who furnished data on their previous positions were analyzed using logistic regression methods. The extracted model correlates tenure with the administrator's past patterns of stability, community attachment, organizational commitment, and facility performance. The model is particularly effective (85% accuracy) in flagging NHAs who are likely to depart within their first 3 years of employment. Implications of these findings for recruitment, retention, and licensure policy are discussed. PMID- 10853526 TI - Negative consequences of hearing impairment in old age: a longitudinal analysis. AB - To determine whether functional and psychosocial outcomes associated with hearing impairment are a direct result or stem from prevalent comorbidity, we analyzed the impact of two levels of reported hearing impairment on health and psychosocial functioning one year later with adjustments for baseline chronic conditions. Physical functioning, mental health, and social functioning decreased in a dose-response pattern for those with progressive levels of hearing impairment compared with those reporting no impairment. Our results demonstrate an independent impact of hearing impairment on functional outcomes, reveal increasing problems with higher levels of impairment, and support the importance of preventing and treating this highly prevalent condition. PMID- 10853527 TI - The use of preexisting and novel coping strategies in adapting to age-related vision loss. AB - Research has proposed that when faced with a stressor, individuals test novel coping strategies when preexisting strategies fail to reduce perceived threat. However, the utilization of novel coping strategies has received scant empirical attention. This study presents data in the form of spontaneous comments or responses to open-ended questions from three previous quantitative studies of adaptation to age-related vision loss (N = 155, 95, and 343 participants). Self reported coping strategies were identified using a "Grounded Theory" approach, and then examined for evidence of whether the strategy was recently utilized (novel) or whether it had been used prior to vision loss (preexisting). Results supported the utilization of novel coping strategies in the process of adaptation to a chronic impairment among older adults. Overall, the use of novel coping strategies was found to be associated with better adaptational outcomes, emphasizing the importance of novel coping in response to stressful life circumstances. PMID- 10853528 TI - The psychosocial preferences of older adults: a pilot examination of content and structure. AB - Individualizing care for older persons depends on knowing about a care recipient's psychosocial preferences. Currently, however, no comprehensive, empirically derived instruments exist to assess these preferences. As part of an effort to develop such an instrument, this pilot study examined the content and structure of psychosocial preferences in older adults using the statistical technique known as concept mapping. Results suggest two underlying dimensions to psychosocial preferences (Enrichment-Self-Maintenance and Extrapersonal Intrapersonal) and six distinct content domains (Social Contact, Growth Activities, Leisure Activities, Self-Dominion, Support Aids, and Caregivers and Care). Both the dimensions and the content domains provide valuable information for the construction of psychosocial preference instruments. They also might assist formal and informal caregivers in tailoring their interventions to provide individualized care that enhances quality of life for older adults. PMID- 10853529 TI - Preliminary explorations of the harmful interactive effects of widowhood and marital harmony on health, health service use, and health care costs. AB - This study examined effects of widowhood and marital harmony on health, health service use, and health care costs. The Americans Changing Lives data set contains 694 subjects who remained married and 61 subjects who became widowed between 1986 and 1989. Estimated annual mean 1989 health costs, adjusting for 1986 costs, age, sex, socioeconomic status, mental/physical health, 1989 health insurance, and selection biases are: $2,384 for widowed, $1,498 for married subjects. Adjusted annual 1989 estimates are: $2,766 for those widowed after harmonious marriages; $2,100 for those widowed after discordant marriages; $1,790 for spouses in discordant marriages; $1,228 for spouses in harmonious marriages. Harmonious marriages appear protective until widowhood, after which higher health costs result. PMID- 10853530 TI - Reducing turnover and improving health care in nursing homes: the potential effects of self-managed work teams. AB - This article describes the use of self-managed work teams (SMWTs) in a nursing home, their potential impacts on the provision of health care and employee satisfaction and turnover, and the factors reported to be important to SMWT effectiveness. Three SMWTs in a midsized nursing home in Wisconsin provide examples. Steps for implementing SMWTs are described. PMID- 10853531 TI - Professional development in law, health care, and aging: a model fellowship program. AB - There is a growing need for a strong core of professionals with the knowledge, skills, and sensitivities necessary to integrate the fields of law, health care, and gerontology. This article describes a unique professional development program that attempted to address this need by making it possible for a small number of recently graduated attorneys to observe, experience, and question on a firsthand level the provision of various forms of health care to older patients. PMID- 10853532 TI - Governance of pharmacy, 1852-1902. PMID- 10853533 TI - With resurgence in use of herbal remedies, unanswered questions take on greater urgency. PMID- 10853534 TI - On the pharmacist's role in pain-related quality-of-life measures. PMID- 10853535 TI - Use of herbal remedies by patients in a health maintenance organization. AB - OBJECTIVE: To examine the use of and experiences with herbal remedies among a group of patients enrolled in a health maintenance organization (HMO). DESIGN: Self-administered questionnaire. SETTING: Central Texas city. PARTICIPANTS: 135 HMO patients. INTERVENTION(S): Not applicable. MAIN OUTCOME MEASURE: Patients' self-reported use of herbal remedies. RESULTS: Almost 40% of patients indicated they had used herbal remedies. The majority had used herbal remedies to treat or prevent a health condition (e.g., common cold). The remedies most frequently used were garlic, aloe gel, cranberry, and echinacea. Most respondents gathered their information on herbal remedies from the popular media, and most based their use decisions primarily on the recommendations of friends and/or relatives. Although most were unsure of the quality of the products, they felt they were safe and somewhat effective, and few had experienced any direct side effects they attributed to the herbal remedies. Most patients used the products without the knowledge of their physician or pharmacist. Herbal remedies were most often used in place of prescription or over-the-counter (OTC) medications and most frequently purchased in health food stores and mass merchandizer/grocery stores. Herbal remedies were sometimes used along with prescription or OTC medications. CONCLUSION: Given that patients are using herbal remedies for a variety of health conditions without medical supervision, pharmacists need to actively and consistently obtain information about herbal remedy use to effectively advise patients and monitor outcomes. More research is needed on herbal remedy use among patient populations and on outcomes in patients who use herbal remedies to treat primary health conditions. PMID- 10853536 TI - The influence of ethnicity on use of herbal remedies in elderly Hispanics and non Hispanic whites. AB - OBJECTIVE: To compare the use of herbal remedies between elderly, self-identified Hispanics and non-Hispanic whites (NHW). DESIGN: Cross-sectional, interviewer administered survey. PATIENTS/SETTING: 186 patients, 65 years and older, at a university-based, ambulatory, senior health center. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Differences in herbal use patterns between Hispanic and NHW participants. Main outcome measures were participants' use of herbal remedies, types of remedies used, medical reason for use, age, sex, Hispanic or NHW ethnicity, income, and education. RESULTS: Overall, 61% of patients had used an herbal remedy at some time in their lives. A larger proportion of Hispanic subjects used herbal remedies than did NHW subjects (77% versus 47%, respectively). Hispanic subjects preferred to use the raw herb in a tea, whereas NHW subjects preferred processed herbs in a capsule or tablet form. Significantly more Hispanic subjects grew or gathered their own herbs and received their information about herbs from a family member than did NHW subjects. Few subjects in either ethnic group received their information about herbal remedies from an allopathic provider. For both groups, the herbs most often used were yerba buena, manzanilla, poleo, osha, and alhucema. The top perceived medical problems that herbs were used for were health care maintenance, dyspepsia, upper respiratory infection, skin problems, and anxiety/nerves/insomnia. CONCLUSION: Ethnicity was related to the frequency of herbal use, the choice and preferred form of herb, and the source of knowledge of herbal remedies. Hispanic culture may account for the observed differences. PMID- 10853537 TI - An evaluation of smoking cessation-related activities by pharmacists. AB - OBJECTIVES: To (1) describe the types of smoking cessation intervention activities performed by community pharmacists and (2) assess the perceived barriers to this type of intervention. DESIGN: Confidential mail questionnaire. SETTING AND PARTICIPANTS: 541 community pharmacists in North Carolina and 946 community pharmacists in Texas. RESULTS: North Carolina and Texas differ with respect to the sale of cigarettes at the practice site, with North Carolina pharmacies being more likely to sell tobacco products. Overall, 555 (92.5%) respondents reported that they do not routinely ask new patients if they smoke or use tobacco products. Pharmacists described themselves as knowledgeable about smoking cessation therapies, and 42% of respondents had attended an educational program on smoking cessation. A total of 230 (39.5%) reported consistently counseling individual patients about smoking cessation treatment strategies on at least a weekly basis. Exploratory factor analysis identified four dimensions of barriers that inhibit pharmacists from engaging in smoking cessation-related activities: (1) pharmacist interpersonal characteristics, (2) practice site considerations, (3) patient characteristics, and (4) financial concerns. CONCLUSION: Pharmacists have an opportunity to identify health risks and counsel patients about disease-preventing lifestyle changes. These findings suggest that although pharmacists believe they are qualified to perform smoking cessation interventions, they do not routinely identify smokers and they perceive several barriers to participating in such activities. Pharmacists should investigate increased involvement in smoking cessation activities for the benefit of their patients and for the potential professional and economic rewards. PMID- 10853538 TI - Pharmacists' attitudes toward diabetes. AB - OBJECTIVES: To assess pharmacists' attitudes toward diabetes, to evaluate the measurement properties of the Diabetes Attitude Scale (DAS) in a sample of pharmacists, and to estimate the number and attitudes of pharmacists certified as diabetes educators (CDE). DESIGN: Mail survey. SETTING/PARTICIPANTS: A systematic sample of 522 hospital, community, and other pharmacists registered with the professional pharmacy association in Alberta, Canada. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: The survey contained the 31 items of the DAS with 4 additional items that addressed attitudes toward the role of the pharmacist. RESULTS: Response rate was 65%. Pharmacists' mean attitude scores were positive to strongly positive toward the Special Training, Control/Complications, Team Care, and Pharmacist's Role factors and neutral toward the Compliance, NIDDM, Difficult to Treat, and Outpatient Education factors. Pharmacists' attitudes varied based on practice setting, year of graduation, and whether they had diabetes or a CDE designation. Most factor scores had good reliability. CONCLUSION: Pharmacists agree that they should be part of the health care team for managing diabetes, should be required to have specialized training to provide primary diabetes care, and that they have the skills to become diabetes educators. PMID- 10853539 TI - Consumer perceptions of risk and required cost savings for generic prescription drugs. AB - OBJECTIVE: To examine consumer risk perceptions for generic prescription drugs used to treat different types of medical conditions, and to explore the relationship between risk perceptions and the amount of cost savings required before consumers would purchase the generic version of a prescription drug. DESIGN: Cross-sectional mail survey. SETTING: Metropolitan area in central Wisconsin. PARTICIPANTS: Random sample of 500 consumers age 18 and older. INTERVENTIONS: None. MAIN OUTCOME MEASURE: Responses to 16 items on 8-page questionnaire. None. RESULTS: The response rate was 71.4%. The percentage of respondents who perceived that generic prescription drugs were riskier than brand name products varied from 14.2% to 53.8%, depending on the medical condition being treated. Significantly larger cost savings were required for consumers to purchase generic prescription drugs with higher perceived risk. CONCLUSION: Financial incentives to use generic prescription drugs may be successful, even for consumers who perceive generic drugs to be riskier than brand name prescription drugs. As the perceived level of risk increases, larger cost savings are required. PMID- 10853540 TI - An exploratory study of community pharmacy practice change. AB - OBJECTIVES: (1) To compare the resources and practitioner activities in community pharmacies that have changed practice with those in pharmacies that have not changed; and (2) in pharmacies that have changed practice, to identify factors affecting a pharmacy's ability to support pharmacy practice change. DESIGN: Multiple-case design, in which six pharmacies were studied in depth. SETTING: Six independent community pharmacies in Iowa. PARTICIPANTS: Pharmacy owners. INTERVENTION: A mail survey and an on-site personal interview were used to collect information about the pharmacy's practice changes and influences on the practice change process. MAIN OUTCOME MEASURES: 14 criteria of pharmacy practice change and 5 types of influences on change (environmental variables, organizational variables, owner/manager characteristics, strategy-making features, attributes of change). RESULTS: Three pharmacies had made considerable change, one had made some change, and two had made little or no change. After a broad set of initial changes, subsequent changes tended to be made incrementally. A variety of factors were identified that support pharmacy practice change. Most of the factors were associated with improving resources, such as upgraded staff skills, involvement in demonstration projects, regular environmental scanning, and regular interaction with advocates for pharmacy practice change (e.g., college of pharmacy, pharmacy associations, innovative practitioners). Also, experienced owners who looked to the future and actively addressed constraints were associated with making pharmacy practice change. CONCLUSION: Practitioners and other interested parties should consider a broad array of activities when trying to facilitate pharmacy practice change. Researchers can use these findings to develop studies that will provide stronger scientific evidence that can contribute to a model of pharmacy practice change. The continued study of pharmacy practice change can assist pharmacists working to translate a philosophy of pharmaceutical care into daily practice. PMID- 10853541 TI - Current strategies for prevention, detection, and treatment of ovarian cancer. AB - OBJECTIVE: To review the prevalence, etiology, risk factors, diagnosis, and treatment of ovarian cancer. DATA SOURCES: English-language journal articles retrieved from a MEDLINE search from 1990 to the present, and selected references retrieved from the bibliographies of those articles. STUDY SELECTION: Clinical trials and pertinent review articles that discussed the detection, prevention, and clinical management of ovarian cancer. DATA SYNTHESIS: Although relatively uncommon, ovarian cancer is the leading cause of mortality from gynecologic cancer. Because most ovarian cancers are not detected until the disease has metastasized beyond the ovary, the 5-year survival rate for all cases is only 50%. Pharmacists can educate women about strategies that can reduce ovarian cancer risk, especially the use of oral contraceptives. To aid in earlier detection, pharmacists also should be aware of the nonspecific symptoms that can be associated with the disease, and refer women with suggestive symptoms to their physicians for further evaluation. Treatment usually consists of hysterectomy with debulking surgery to remove as much tumor as possible, followed by chemotherapy, for which the current gold standard is cisplatin and paclitaxel. CONCLUSION: Pharmacist-provided education can help women reduce their risk of ovarian cancer. As integral members of the health care team, pharmacists also can optimize the efficacy and tolerability of chemotherapeutic regimens; assist with palliative care for nausea, vomiting, and pain; and serve as a resource for patient information and support. PMID- 10853542 TI - Quality-of-life assessment in acute, chronic, and cancer pain: a pharmacist's guide. AB - OBJECTIVE: To describe instrumentation, or measures, available for use in assessing the impact of pain on the quality of life (QOL) of patients, and methods to evaluate the appropriateness of these QOL measures. DATA SOURCES: MEDLINE, PSYCHLit, and CANCERLit were searched from 1980 through 1997 to identify QOL instruments that included a pain subscale or pain-related items. DATA SYNTHESIS: Given the high prevalence of chronic diseases or conditions that include pain as a primary or secondary symptom, pharmacists should understand how pain affects the QOL of patients. Over the past two decades, emphasis has increased on developing instruments that assess health-related QOL concerns, including pain. Scores of measures--including utility measures--are available to measure general QOL in patients with conditions involving pain. Condition specific instruments have also been developed to measure the impact of specific conditions, such as arthritis, on QOL. Guidelines are presented for evaluating QOL instrumentation, and existing measures used to evaluate the QOL of patients with acute and chronic pain are described. Pharmacists can use these guidelines to evaluate the usefulness of existing instruments for assessing the QOL of patients with pain. CONCLUSION: Using QOL measures in everyday practice may assist pharmacists in gaining insight into the effects of pain on their patients' QOL. This information may be useful in developing treatment programs that minimize pain and its associated side effects while maximizing patients' well being. PMID- 10853543 TI - Evaluating prescriptions for the elderly: drug/age criteria as a tool to help community pharmacists. AB - OBJECTIVES: To use drug/age criteria to determine (1) the prevalence of dispensing of drugs potentially inappropriate for use in elderly patients; (2) the dispensing rate of individual drugs considered potentially inappropriate for use in elderly patients; (3) the association between selected patient characteristics and the prevalence of potentially inappropriate drug dispensing. DESIGN, SETTING, PARTICIPANTS: A secondary database of 6,380 new prescription orders dispensed to patients of all ages in ambulatory pharmacies in a mid western state was used retrospectively for the analysis. A total of 1,530 (23.9%) of the new prescription orders were dispensed to 1,185 elderly patients. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Name, strength, and daily dose of each drug dispensed were compared with drug/age criteria to determine whether a dispensed drug was inappropriate for use in elderly patients. The association of the rate of dispensing of potentially inappropriate drugs with characteristics of the elderly patients, including age, sex, race, number of comorbidities, and prescription drug insurance coverage type, was determined. RESULTS: A total of 170 patients (14.3%) were dispensed potentially inappropriate medications. The three most common medications were propoxyphene and propoxyphene combinations, prescription and nonprescription antihistamines, and digoxin at doses > 0.125 mg/day. There was no statistically significant association between inappropriate drug dispensing and patient age, sex, race, number of comorbidities, and prescription drug insurance coverage type. CONCLUSION: Pharmacists can use drug/age criteria as a tool for an initial check to assess the appropriateness of drugs used by the elderly. Elderly patients appear equally at risk of using potentially inappropriate medications regardless of demographic, disease, or insurance characteristics. PMID- 10853544 TI - Pharmacists' attitudes toward competency exams. PMID- 10853545 TI - The dietary supplement dilemma: practice advice for pharmacists. PMID- 10853546 TI - Recent biotechnology approvals. PMID- 10853547 TI - [Management of the patient with hypertensive renal insufficiency]. PMID- 10853548 TI - [The Lariboisiere Hospital, its past and its present]. PMID- 10853549 TI - [Histology and elementary pathology of the peripheral nerve]. AB - The lesions which give rise to peripheral neuropathy are induced by a variety of mechanisms. Their etiology can often be established from the results of electrophysiological examination and (or) study of nerve biopsy; knowledge of the etiology will guide the treatment strategy. In this chapter, we discuss typical lesional mechanisms including: axonal involvement (Wallerian degeneration, axonopathies) and demyelinating lesions (whether segmental, indicative of an acquired inflammatory process, or of a more extensive nature, indicative of a genetic origin). The indication for nerve biopsy, which enables identification of the lesions, should be considered on a case by case basis. PMID- 10853550 TI - [Clinical and paraclinical investigations of chronic polyneuropathies]. AB - Given the composition of the nerve fibre, the signs and symptoms of chronic polyneuropathy are not uniform. Clinically, they include sensory, motor and reflex signs and symptoms with, according to the case, association of sensory, either painful or not, motor and sensory-motor forms, with a reflex component. Electrophysiologic tests can usually distinguish initially axonal lesions from those due to initial myelin alteration. Nerve biopsy, although not a routine examination, brings precise information concerning etiology. PMID- 10853551 TI - [Acquired demyelinating neuropathies]. AB - The acquired demyelinating polyneuropathies with either acute or chronic clinical presentation are considered autoimmune disorders. The Guillain-Barre syndrome is viewed as an acutely reactive and self-limited autoimmune disease, triggered by preceding bacterial or viral infections. There is a particularly strong association with the gastroenteric pathogen, Campylobacter jejuni, and with Cytomegalovirus and Epstein-Barr virus. It is likely that immune response directed towards the infecting organisms are involved in the pathogenesis of Guillain-Barre syndrome by cross-reaction with neural tissues. In the susceptible individual, the infecting organism induces humoral and cellular immune responses that, because of the sharing of homologous epitopes (molecular mimicry), cross react with ganglioside surface components of peripheral nerve. Immune reactions against target epitopes in the Schwann cell surface membrane or myelin result in acute inflammatory demyelinating neuropathy (90% of cases); reactions against epitopes contained in the axonal membrane cause the acute axonal forms of Guillain-Barre syndrome. Immunomodulation with infusions of IgG or plasma exchange treatments effectively foreshorten the disease course. The immunopathogenesis of the chronic disease forms, chronic inflammatory demyelinating peripheral neuropathy and multifocal motor neuropathy are less well known. Immunomodulatory treatments with corticosteroid or cytotoxic drug treatments, infusion of Ig or therapeutic plasma exchanges are variably effective. The article outlines the principles and practices of an individualized approach to therapy. PMID- 10853552 TI - [Metabolic and nutritional neuropathies]. AB - The two main causes of metabolic neuropathies are successively diabetes and chronic renal insufficiency. Diabetic neuropathies include both diffuse polyneuropathies and focal neuropathies. Sensori(motor) polyneuropathy is the most frequent form and different therapeutic trials have been initiated on the ground of the vascular and metabolic factors implicated in its pathogenesis. Autonomic neuropathy is the major cause of morbidity and mortality. In patients with chronic renal failure, the polyneuropathy is improved by renal transplantation. The carpal tunnel syndrome is frequent in hemodialysis patients, and surgery gives the opportunity to look for beta-2-microglobulin amyloid deposits. Among the less frequent causes of peripheral neuropathies in which metabolic factors have been considered, we review hypoglycemia, chronic respiratory insufficiency due to chronic obstructive pulmonary disease, chronic liver diseases, and the polyneuropathy occurring in the critically ill patients with nutritional or metabolic failures. In chronic excessive drinkers peripheral neuropathy is classically associated with thiamine deficiency, but the direct effect of alcohol itself has been discussed. Various vitaminic deficiencies have been responsible for the development of peripheral neuropathies. The clinical forms often associate peripheral neuropathy with myelopathy, and serum vitamin E concentrations should be measured in patients with spinocerebellar disorders. Usually nutritional deficiencies need multivitamins supplementation. PMID- 10853553 TI - [Hereditary neuropathies]. AB - Hereditary neuropathies are the most frequent genetically determined disease in neurology. The peripheral motor and sensory form is the Charcot-Marie-Tooth disease. The underlying genetic defects are now known for many of the demyelinating form. Several studies have identified the mutations on genes coding proteins of peripheral myelin. This has important implications for both diagnosis and genetic counselling in this group of conditions. The genetic defect in most cases of familial amyloid polyneuropathy is better known and should be considered especially in late-onset forms. PMID- 10853554 TI - [Infectious neuropathies]. AB - Infective neuropathies constitute a leading cause of neuropathies in the world. The number of patients with nerve lesions related to leprosy remains high despite decreasing number of new cases requiring multidrug regimens. Peripheral neuropathies associated with HIV infection may be found in up to 50% of patients. Neuropathies may be related to the inflammatory reaction against viral antigens, immunodepression, opportunistic infections, and iatrogenic complications of anti viral drugs. Hepatitis C virus infection has been found in cryoglobulinemic neuropathies. This virus should be screened for exploration of all neuropathies. Rare causes of neuropathy, such as poliomyelitis and diphtheria, and treatable neuropathies such as Lyme disease, should not be forgotten. PMID- 10853555 TI - [Vascular neuropathies]. AB - Vasculitic neuropathy is an important cause of peripheral neuropathy because of its potentially severe prognosis and an early recommended therapy. Diagnosis is easy when the clinical presentation is an acute, painful, multiple mononeuropathy but more difficult when presenting as a polyneuropathy. Electrophysiologic studies play an important role in the diagnosis. Muscle and nerve biopsies confirm the vasculitic process and demonstrate ischemic neuropathy lesions. Polyarteritis nodosa, rheumatoid arthritis and non-systematic vasculitic neuropathy are the main conditions of vasculitic neuropathy. Usual treatment includes a combination of corticosteroids and cyclophosphamide. PMID- 10853556 TI - [Neuropathies associated with cancers and dysglobulinemias]. AB - Peripheral neuropathies associated to cancers encompass several causes: entrapment or direct involvement of peripheral nerves, paraneoplastic and non tumoral causes, mainly infectious, metabolic or carential. Paraneoplastic neuropathy is mainly the so-called sensory ganglionopathy. A purely motor neuronopathy may be observed in Hodgkin's disease and "terminal" polyneuropathy may complicate may known cancer, in a context of severe weight loss. Polyneuropathies associated with paraproteinaemia include those associated with a defined lymphoproliferative disorder (myelomas, solitary plasmocytoma, Waldenstrom's disease, lymphoma), and associated with monoclonal gammopathy of undetermined significance. PMID- 10853557 TI - [Learning the critical lecture: choice morsels]. PMID- 10853558 TI - [Urinary lithiasis. Etiology, physiopathology, diagnosis, development, treatment]. PMID- 10853559 TI - [Herpetic encephalitis. Diagnosis and treatment]. PMID- 10853560 TI - [Prescription of a volume-expanding solution]. PMID- 10853561 TI - [Laryngeal dyspnea in children. Diagnostic orientation and management of emergency cases]. PMID- 10853562 TI - [Polyuria-polydipsia syndrome. Diagnostic approach]. PMID- 10853563 TI - [Chronic diarrhea in the adult. Diagnostic approach]. PMID- 10853564 TI - A comparison of certain practice characteristics of dental anesthesiologists in Canada and the United States. AB - An existing database was used to compare aspects of dental anesthesiology practice of dental anesthesiologists in Canada (n = 32) and the United States (n = 123). Data focusing on percutaneous injuries were obtained through a mailed questionnaire that was returned anonymously. Respondents provided information on the treatment of patients under deep sedation or general anesthesia only. Eighty one percent of Canadians and 61% of Americans returned the questionnaire. The vast majority (84%) of injuries reported were due to sharps associated with general dentistry compared with those associated with anesthesiology. Canadians were more likely to be operator-anesthetists (P < .01) and to experience a percutaneous injury (P < .01) than US practitioners. American practitioners were more likely to have a greater proportion of the caseload under the age of 20 (P < .02). No other significant differences were observed. These results illustrate a number of unique attributes of the practice of dental anesthesiology in these 2 countries. PMID- 10853565 TI - Evaluation of prilocaine for the reduction of pain associated with transmucosal anesthetic administration. AB - This investigation evaluated the use and efficacy of prilocaine HCl (4% plain Citanest) for minimizing pain associated with the intraoral administration of local anesthesia. Clinical anecdotes support the hypothesis that prilocaine without a vasoconstrictor reduces pain during injection. To determine relative injection discomfort, use of 4% plain prilocaine was compared with use of 2% lidocaine with 1:100,000 epinephrine and 2% mepivacaine with 1:20,000 levonordefrin. Prior to routine endodontic procedures, 150 adult patients received 0.3 to 1.8 mL of local anesthetic via the same gauge needle without the use of a topical local anesthetic. Injection methods included buccal infiltration, labial infiltration, palatal infiltration, and inferior alveolar nerve block. Following each injection, patients were asked to describe the level of discomfort by scoring on a visual analog scale of 1 to 10, where 1 = painless and 10 = severe pain. Analyses via 2-way analysis of variance revealed no interaction between anesthetic and site of injection. However, there were statistically significant differences among the injection sites. Post hoc analysis revealed that prilocaine was associated with significantly less pain perception when compared to mepivacaine and lidocaine. These results suggest that differences in initial pain perception during transmucosal injection may be a function of the local anesthetic use, and prilocaine can produce less discomfort than the others tested. PMID- 10853566 TI - Cardiovascular effects of bupivacaine and the role of this agent in preemptive dental analgesia. AB - Inappropriately high blood concentrations of bupivacaine have been reported to cause toxicity and even death. The potential for cardiovascular toxicity and the difficulty with which this may be reversed has made the dental practitioners reluctant to use this agent. Nevertheless, cardiovascular toxicity from its use in and around the mouth is exceedingly rare. This study was undertaken to assess bupivacaine's cardiotoxic potential in the practice of oral and maxillofacial surgery. Results showed a dose-dependent decrease in systolic blood pressure, but no other statistically significant cardiovascular change was noted. Preemptive treatment of postsurgical pain has been the subject of numerous trials. Bupivacaine administered preoperatively has been suggested to prevent central nervous system "conditioning," thus decreasing the perceived postoperative pain. However, there was no statistical support for any reduction in the perceived postoperative pain in the treatment groups in this study. PMID- 10853567 TI - The occupational risk to dental anesthesiologists of acquiring 3 bloodborne pathogens. AB - OBJECTIVE: To estimate the occupational risk to dental anesthesiologists of contracting 3 bloodborne pathogens: hepatitis B (HBV), hepatitis C (HCV), and human immunodeficiency virus (HIV). METHODS: Through an anonymously returned, mailed questionnaire, dental anesthesiologists in Canada and the United States provided information regarding percutaneous and mucocutaneous contacts with contaminated fluid during the treatment of patients under deep sedation and general anesthesia as well as other general practice information. A mathematical model was applied to determine the occupational risk. RESULTS: Of the 101 (65%) returned questionnaires, 98 reported having treated patients within the previous 6 months. Of these, 41 (42%) had at least one percutaneous accident (89 accidents in total), and the projected mean annual injury rate for dental anesthesiologists overall was 1.82. The most common causes of injury were burs, intraoral needles, and dental instruments. Operator error during use was associated with 31% of reported accidents. Significantly more injuries were reported by those who also reported a mucocutaneous contact and by those working more than 25 hours per week. The projected mean annual number of mucocutaneous exposures was 0.88 for dental anesthesiologists overall. CONCLUSIONS: The calculated annual risk to the average dental anesthesiologist of acquiring HBV (if not immune), HCV, and HIV following percutaneous injury was very low for all infections (HBV the most; HIV the least). The risk of contracting HIV following mucocutaneous contact was extremely low. PMID- 10853568 TI - Infiltrated lidocaine 2% with epinephrine 1:80,000 causes more postoperative pain than lidocaine 2% after oral soft tissue surgery. AB - A controlled, randomized, double-blind, within-patient, crossover study was made with 50 patients (28 women and 22 men) of mean age 47 years (range, 32-69 years) who were subjected to identical bilateral gingivectomies. On one occasion, lidocaine 2% was infiltrated as the local anesthetic. On the other occasion, lidocaine 2% with epinephrine 1:80,000 was given. Postoperative pain intensity was recorded by the patients on a 100-mm visual analogue scale every hour during an 11-hour observation period. The mean pain intensity was numerically higher after lidocaine 2% at 0 hours and 1 hour postoperatively. Then the mean pain intensity after lidocaine 2% was lower than that after lidocaine 2% with epinephrine 1:80,000 throughout the remaining observation period. The difference in pain intensity was statistically significant (P < .05) at 2, 4, 5, 6, and 7 hours after surgery. Mean sum (SEM) pain intensity over the 11-hour observation period was lower (P = .03) after lidocaine 2%, 66.5 (13.4) mm than after lidocaine 2% with epinephrine 1:80,000, 92.6 (15.4) mm. The study shows that high epinephrine concentration (1:80,000) increases the postoperative pain after dental soft tissue surgery with mild pain. PMID- 10853569 TI - Implant dentistry measurements using the Dexis digital radiography system by provision dental systems. PMID- 10853570 TI - Finding, using appropriate materials is an ongoing challenge. PMID- 10853571 TI - Use segmented hex-top implants to maximize restorative flexibility for individual patient needs. PMID- 10853572 TI - Go the extra step to develop a relationship between the specialist and the restorative doctor. PMID- 10853573 TI - Surgical reconstruction of the interdental papillae. PMID- 10853574 TI - Soft-tissue procedures related to dental implant surgery. PMID- 10853575 TI - Use of the OsteoHarvester for bone harvesting, collection, and placement. PMID- 10853576 TI - Immediate esthetic reconstruction for anterior single-tooth application using a hydroxyapatite-coated cylinder root-form implant. PMID- 10853577 TI - Implant selection considerations and criteria. PMID- 10853578 TI - Use of human collagen allograft material impregnated with demineralized freeze dried bone granules. PMID- 10853579 TI - Aesthetic restoration of endodontically treated teeth. AB - The utilization of all-ceramic anterior restorations has been widely advocated by dental clinicians for the creation of a natural appearance, even in teeth that have undergone endodontic treatment. A suitable aesthetic level is generally more difficult to achieve, however, when a post and core system is required in the absence of adequate dental structure. In particular, conventional cast-metallic posts negate light transmission, which has long been considered a vital component in the creation of optimal aesthetics. This article provides an overview of the materials, fabrication methods, and procedures involved in anterior reconstruction using aesthetic alternative systems and solutions. PMID- 10853580 TI - The tipped mandibular molar as a bridge abutment: Part 2. PMID- 10853581 TI - Three-dimensional shade analysis: perspectives of color--Part I. AB - Although color is evident in daily function, it is often misunderstood and obscured by misconceptions. While the literature focuses on mathematical formulas and specific facets of color, little research exists on the fundamental aspects of color realization. This article demonstrates the principles of color, which include definitions, measurements, sensations, and perceptions in conjunction with its practical application in restorative dentistry. The second segment of the article discusses shade selection and the use of contemporary shade systems for the fabrication of dental prostheses. PMID- 10853582 TI - Aesthetic posterior restoration utilizing condensable composite resin. PMID- 10853583 TI - Biological basis for clinical success: pulp protection and the tooth-restoration interface. AB - This article provides biological and technological information that strengthens clinicians' understanding of cohesive hybridization and pulp therapy in order to support their routine use of bonding and resin systems. Utilizing cohesive systems, clinicians should experience several advantages over traditional water soluble base and liner systems. When properly applied, cohesive hybridization of vital dentin prevents immediate postoperative hypersensitivity under all restorations and completely seals the entire tooth-restoration interface, which provides a reduction in recurrent caries. PMID- 10853584 TI - Air abrasion in the aesthetic restorative practice. PMID- 10853585 TI - Anterior single-tooth replacement: clinical examination and treatment planning. AB - The replacement of a single anterior tooth is a complex, challenging procedure that can be accomplished with implant-supported restorations as well as conventional porcelain-fused-to-metal and resin-bonded fixed partial dentures. A comprehensive diagnostic form may be beneficial in determining the most effective means of rendering treatment for each patient. This article demonstrates the use of this form, diagnostic models, and radiographs to diagnose and restore three patients who presented for the replacement of a single tooth in the anterior maxilla. PMID- 10853586 TI - The cessation of tobacco use: a clinical perspective. PMID- 10853587 TI - Transitional custom abutments: optimizing aesthetic treatment in implant supported restorations. AB - As the discipline of implant prosthodontics continues to evolve, additional emphasis has been focused on the role of adequate emergence profiles and peri implant soft tissue contours. Despite the development of anatomically shaped healing abutments, currently advocated techniques exhibit clinical limitations. This article introduces the transitional custom abutment technique as a method of manipulating the supra-implant soft tissue contours so that optimal emergence profiles and increased restorative flexibility can be achieved in the treatment of patients with compromised fixture angulation. PMID- 10853588 TI - Metal-free restorations for fractured teeth using a new glass ceramic. PMID- 10853589 TI - Carpal tunnel syndrome: a growing epidemic among dental professionals? PMID- 10853590 TI - Selection and ideal tridimensional implant position for soft tissue aesthetics. AB - While single-tooth replacement can be accomplished with predictability using implant therapy, this procedure is challenging in the anterior region where numerous criteria must be evaluated by the restorative team. The available height of bone, soft tissue volume, and three-dimensional position of the anticipated implant restoration are among the numerous concerns that must be addressed prior to the initiation of treatment. This article provides a comprehensive review for the selection and placement of implants in the aesthetic region and illustrates these principles with a case presentation. PMID- 10853591 TI - Utilization of an aesthetic post system for restoration of an endodontically treated tooth. PMID- 10853592 TI - Computerized occlusal management of a fixed/detachable implant prosthesis. AB - The long-term success of an implant-supported prosthesis is related to the management of occlusal forces upon the restoration. Since implants are not anchored in bone with periodontal ligament proprioceptors, subjective confirmation by the patient is not always an accurate means of determining occlusion. A computerized occlusal analysis system has been developed to determine the timing and nature of the occlusal forces on tooth contacts. This article details the implementation of this system as an aid in the occlusal adjustment procedure of an implant-supported fixed/detachable prosthesis. PMID- 10853593 TI - Root canal preparation using engine-driven nickel-titanium rotary instruments. PMID- 10853594 TI - The importance of laboratory communication in modern dental practice: stone models without faces. AB - The development of advanced communication technology has enabled clinicians and ceramists to deliver improved dental care to their patients. This article highlights the importance of communication between the clinician, laboratory technician, and patient for the fabrication and delivery of aesthetic restorations. The use of stone models, photography, shade prescriptions, and occlusal relationships is demonstrated as a means to achieve this objective. These principles are illustrated in the restoration of a patient who presented for aesthetic enhancement of the anterior maxilla. PMID- 10853595 TI - Contemporary materials for removable prosthodontics. PMID- 10853596 TI - Indirect resin inlay and onlay restorations: a comprehensive clinical overview. AB - Advances in adhesive technology and aesthetic dental materials have permitted clinicians to perform conservative preparation of the dentition for inlay/onlay restorations. The use of adhesive indirect procedures provides numerous advantages (e.g., aesthetics, reinforcement, adequate seal) to conventional restorative techniques. This article describes the indications/contraindications and material properties of aesthetic indirect restorations. It also highlights the preparation, bonding, and finishing procedures utilized to achieve an aesthetic indirect resin restoration and demonstrates the techniques with a series of case presentations. PMID- 10853597 TI - Manipulating light with the refractive index of an all-ceramic material. PMID- 10853598 TI - Helicobacter pylori and the oral environment. PMID- 10853599 TI - The periodontal-restorative interface: enhancement through magnification. AB - One critical factor associated with aesthetics, periodontal health, and the longevity of restorations is the precision of the margins at the periodontal restorative interface. Improper margins can cause overhangs and overcontouring that may ultimately result in caries, periodontal inflammation and breakdown, and compromised aesthetics. In order to prevent pathology at the restorative-tooth interface, each phase of the aesthetic treatment must be performed with precision and care. This article presents the utilization of an operating microscope to improve the marginal adaptation of all-porcelain restorations. PMID- 10853600 TI - Aesthetic rehabilitation of discolored dentition with metal-ceramic restorations. PMID- 10853601 TI - Root canal instrumentation with a patency technique. PMID- 10853602 TI - Adhesive protocol for the utilization of aluminum oxide crown restorations. AB - The ultimate objective of anterior tooth restoration is to achieve an aesthetic appearance that is in harmony with the natural dentition. It is often challenging to replicate the optical characteristics (i.e., transparency and translucency) of a natural tooth without utilizing all-ceramic crown restorations, which may provide enhanced aesthetics and biocompatibility in comparison to metal-based treatment. This article demonstrates the use of all-ceramic technology with a contemporary adhesive procedure as a means to achieve improved aesthetic results in the restoration of the anterior dentition. PMID- 10853603 TI - Reconstruction of the maxillary dentition utilizing a non-orthodontic technique. PMID- 10853604 TI - Glass-infiltrated zirconia/alumina-based ceramic for crowns and fixed partial dentures. AB - The increased demand for metal-free restorative alternatives has resulted in the proliferation of all-ceramic systems. While these materials can predictably achieve aesthetic results in the anterior, they have traditionally been contraindicated for posterior applications due to the greater stresses present in the region. This article discusses a zirconia/alumina-based ceramic system that has been developed to expand the alternatives for the aesthetic restoration of the dentition. Material properties and considerations for its use in crown restorations, fixed partial dentures, and custom implant abutments are similarly addressed. PMID- 10853605 TI - Orthodontic and surgical management of a partially erupted mandibular first molar. PMID- 10853606 TI - Public health dentistry in Virginia: past, present, and future. PMID- 10853607 TI - MRI of the inner ear: comparison of axial T2-weighted, three-dimensional turbo spin-echo images, maximum-intensity projections, and volume rendering. AB - RATIONALE AND OBJECTIVES: To compare the ability of axial T2-weighted, three dimensional, turbo spin-echo (3D TSE) images, targeted maximum-intensity projections (MIPs), and 3D volume reconstructions to depict anatomic details of the labyrinth. METHODS: In 24 volunteers, 3D TSE images were obtained. MIPs and 3D volume reconstructions were performed from the acquired data. All images were evaluated by three radiologists independently regarding the visualization of the different anatomic structures. RESULTS: In the axial slices, most anatomic details were visible in comparison with observations by the other modalities. The 2.5 windings of the cochlea were best depicted on the MIPs. Volume reconstructions rendered excellent spatial information regarding the vestibule and semicircular canals and were the only technique that demonstrated all three ampullae in all cases. CONCLUSIONS: Axial TSE images, MIPs, and 3D volume reconstructions are complementary modalities that provide different information. Our results suggest that improved diagnostic information can be obtained by applying these volume visualization reconstruction techniques. PMID- 10853608 TI - Magnetic resonance monitoring of stent deployment: in vitro evaluation of different stent designs and stent delivery systems. AB - RATIONALE AND OBJECTIVES: To evaluate MR imaging features of commercially available stents before, during, and after in vitro deployment as a step toward MR-guided stent deployment. METHODS: Fourteen stents were deployed in a phantom under MR monitoring at 1.5 T by using a gradient-echo sequence. Device visibility was rated on a four-point scale (excellent, fair, poor, not visible). RESULTS: The Memotherm stent and the rolling membrane (RM) Wallstent showed excellent stent visibility and at least fair scores for artifact-induced narrowing of the stent lumen. Three stents (Palmaz, AVE, Easy Wallstent) showed excellent visibility of the stent but no visible lumen. Five stents (Strecker, Accuflex, Hemobahn, Passager, Sinus) displayed fair visibility. The delivery catheters of four stent systems (Smart, Vascucoil, Symphony, ZA) displayed severe black hole artifacts. CONCLUSIONS: The imaging features of several stent systems might be suitable for MR-guided intervention. The Memotherm and the Wallstent RM combine good visibility of the stent and the lumen. PMID- 10853609 TI - Percutaneous venous thrombectomy with the use of a balloon sheath: first in vitro investigations of a new low-tech concept. AB - RATIONALE AND OBJECTIVES: To test mechanical thrombectomy of extensive iliofemoral and iliocaval thrombi in an in vitro flow model with the use of 12F and 18F balloon sheaths. METHODS: Newly developed 12F and 18F sheaths were evaluated in four vessel models (simulation of femoral, iliofemoral, iliocaval, and caval thrombi by clotted bovine blood in a flow model). After retrograde insertion of the sheath and blocking of the vessel proximal to the thrombus by inflating the balloon, mechanical fragmentation was performed coaxially through the sheath lumen by using a 7F pigtail rotation device. With an occlusion balloon catheter, residual thrombi were withdrawn to the orifice of the sheath and aspirated. Twelve silicone tubes occluded by thrombi were recanalized in each setting. In the latex model, seven recanalizations were performed. RESULTS: All clots were removed completely within a treatment duration of 2 to 14 minutes. Fluid loss during the procedure was 29.6 to 129.3 mL for the femoral flow model, 61.9 to 137.2 mL for the iliofemoral model, 74.5 to 163.4 mL for the iliocaval model, and 102.7 to 236.7 mL for the caval model. No fragments were washed downstream. In four settings, small residual thrombi were attached to the balloon after deflation of the sheath. CONCLUSIONS: Clot amounts up to 171 g were removed quickly and completely by using these large-caliber balloon sheaths. Fluid loss from aspiration was negligible. Balloon occlusion prevented embolization of thrombus fragments proximal to the sheath. Further studies are needed to prove the efficacy of this technique in vivo. PMID- 10853610 TI - Evaluation of fluid collection in the pericardial sinuses and recesses: noncontrast-enhanced electron beam tomography. AB - RATIONALE AND OBJECTIVES: To evaluate the attenuation, size, and volume of the pericardial sinuses and recesses by using electrocardiographically triggered, noncontrast-enhanced electron beam tomography (EBT) and to consider its relation with sex, age, and heart volume. METHODS: Findings in 213 consecutive patients without known pericardial disease were studied. The patients underwent EBT scanning of the heart to evaluate coronary artery calcification. Incremental electrocardiographically triggered noncontrast images were obtained with a 100-ms exposure time and a 3-mm slice thickness. The appearance, density, and volume of the pericardial sinuses and recesses were calculated. RESULTS: Among the 213 patients, 97.2% had at least one of the sinuses or recesses visible on EBT. The sinuses or recesses were seen with the following frequency: transverse sinus (93.9%), oblique sinus (71.8%), and superior aortic recess (51.2%). The mean attenuation and volume were 9.9 +/- 7.3 Hounsfield units (HU), 12.6 +/- 8.1 HU, and 12.6 +/- 8.7 HU, and 1.9 +/- 1.3 mL, 1.3 +/- 1.0 mL, and 0.8 +/- 0.8 mL, respectively. The total volume of the pericardial sinuses (3.3 +/- 2.2 mL) had no significant relation with the total heart volume. CONCLUSIONS: Pericardial sinuses and recesses were frequently and well depicted on noncontrast EBT images. In patients without obvious pericardial effusion, physiological fluid collections were observed in the transverse and oblique sinuses or other recesses. Location, attenuation, and volume were helpful in the differentiation of normal pericardial sinuses from pericardial effusions and mediastinal lymph nodes. PMID- 10853611 TI - Quantitative characterization of mass lesions on digitized mammograms for computer-assisted diagnosis. AB - RATIONALE AND OBJECTIVES: To investigate features for discriminating benign from malignant mammographic findings by using computer-aided diagnosis (CAD) and to test the accuracy of CAD interpretations of mass lesions. METHODS: Fifty-five sequential, mammographically detected mass lesions, referred for biopsy, were digitized for computerized reevaluation with a CAD system. Quantitative features that characterize spiculation were automatically extracted by the CAD system. Data generated by 271 known retrospective cases were used to set reference values indicating the range for malignant and benign lesions. After conventional interpretation of the 55 prospective cases, they were evaluated a second time by the radiologist using the extracted features and the reference ranges. In addition, a pattern-recognition scheme based on the extracted features was used to classify the prospective cases. Accuracy of interpretation with and without the CAD system was evaluated using receiver operating characteristic (ROC) curve analysis. RESULTS: Sensitivity of the CAD diagnosis for the prospective cases improved from 92% to 100%. Specificity improved significantly from 26.7% to 66.7%. This was accompanied by a significant increase in the accuracy of diagnosis from 56.4% to 81.8% and in the positive predictive value from 51.1% to 71.4%. The Az for the CAD ROC curve significantly increased from 0.73 to 0.90. The performance of the classification scheme was slightly lower than that of the radiologists' interpretation with the CAD system. CONCLUSIONS: Use of the CAD system significantly improved the accuracy of diagnosis. The findings suggest that the classification scheme may improve the radiologist's ability to differentiate benign from malignant mass lesions in the interpretation of mammograms. PMID- 10853612 TI - In vitro model of the human liver parenchyma to study hepatotoxic side effects by Dy-EOB-DTPA. AB - RATIONALE AND OBJECTIVES: In vivo studies have shown species-specific toxicity after application of the liver-specific contrast agent Dy-ethoxybenzyl (EOB) DTPA. To predict species differences in the laboratory, an in vitro model of the liver was used to examine the divergent results. METHODS: Rat, canine, porcine, and human hepatocytes were isolated and embedded between layers of collagen. During and after 48 hours of incubation with different concentrations of Dy-EOB DTPA (maximum concentration 50 mmol/L), morphological changes and enzyme leakage were determined. RESULTS: The response to the contrast agent varied for hepatocytes from different species. For canine cells, morphological changes and cell death were evident with as little as 5 mmol/L Dy-EOB-DTPA. Rat hepatocytes tolerated up to 50 mmol/L Dy-EOB-DTPA, and enzyme leakage was transient. Only after incubation with 50 mmol/L Dy-EOB-DTPA was the formation of intracellular vacuoles evident. In contrast, even the highest concentration of Dy-EOB-DTPA did not cause an enzyme leakage of porcine or human hepatocytes, although similar vacuoles were seen. CONCLUSIONS: These data demonstrate a species-dependent toxicity for Dy-EOB-DTPA in vitro, with similar responses in porcine and human hepatocytes. PMID- 10853613 TI - Reduction of motion artifacts in magnetic resonance imaging of the neck and cervical spine by long-term averaging. AB - RATIONALE AND OBJECTIVES: We performed a prospective comparison of T1-weighted turbo spin-echo (TSE) imaging with standard averaging and with the long-term averaging method (LOTA), comparing the effects on signal-to-artifact noise ratio (S/aN) and motion artifacts. METHODS: In 30 consecutive patients undergoing imaging of the neck or cervical spine, a transverse T1-weighted TSE sequence was applied with and without LOTA by using identical sequence parameters. Quantitative image analysis was done by calculating S/Ns in the phase-encoding direction (S/aN). Visual image analysis was performed by four independent, masked readers using a standardized score sheet for anatomic and pathological findings. RESULTS: The LOTA sequence yielded significantly superior S/aN values compared with the standard averaging sequence. In the subjective evaluation, the LOTA sequence showed significantly fewer motion artifacts and better visualization of normal anatomy of the neck, cervical spine, and spinal cord, as well as of the pathological findings. CONCLUSIONS: LOTA is a valuable method for increasing S/aN in magnetic resonance imaging of the neck and cervical spine. It reduces motion artifacts and increases the conspicuity of pathology without increasing acquisition time. No additional hardware is needed, and this technique can be combined with other artifact-reducing methods. PMID- 10853614 TI - Mechanical thrombectomy of acute thrombosis in transjugular intrahepatic portosystemic shunts. AB - RATIONALE AND OBJECTIVES: To evaluate the feasibility of mechanical thrombectomy with the use of the Amplatz thrombectomy device (ATD) in restoring patency to acutely thrombosed stent-shunts after transjugular intrahepatic portosystemic shunt (TIPS) placement. METHODS: Mechanical thrombectomy with the ATD was performed in 10 consecutive patients with angiographically documented complete thrombosis of the stent-shunt (mean +/- SD, 6.6 +/- 3.4 cm), which occurred an average of 2.8 months (range, 0-11 months) after the TIPS procedure. RESULTS: In all patients, immediate restoration of patency of the stent-shunt was achieved after thrombectomy alone (n = 1), thrombectomy plus percutaneous transluminal angioplasty (PTA; n = 4), and thrombectomy, PTA, and stenting (n = 5). The mean activation time of the ATD was 253 +/- 43 seconds. The pressure gradient for portal decompression decreased from 23 +/- 6 mmHg before to 11 +/- 3 mmHg after the procedure. The primary patency rate was 80% at 3 months and 60% at 11 months. CONCLUSIONS: Mechanical thrombectomy with the ATD in acutely thrombosed TIPS is technically feasible. Mechanical thrombectomy is a potential alternative to thrombolysis. PMID- 10853615 TI - Clinical pharmacy services, pharmacy staffing, and the total cost of care in United States hospitals. AB - This study evaluated direct relationships and associations among clinical pharmacy services, pharmacist staffing, and total cost of care in United States hospitals. A database was constructed from the 1992 American Hospital Association's Abridged Guide to the Health Care Field and the 1992 National Clinical Pharmacy Services Database. A multiple regression analysis, controlling for severity of illness, was employed to determine the relationships and associations. The study population consisted of 1016 hospitals. Six clinical pharmacy services were associated with lower total cost of care: drug use evaluation (p=0.001), drug information (p=0.003), adverse drug reaction monitoring (p=0.008), drug protocol management (p=0.001), medical rounds participation (p=0.0001), and admission drug histories (p=0.017). Two services were associated with higher total cost of care: total parenteral nutrition (TPN) team participation (p=0.001) and clinical research (p=0.0001). Total costs of care/hospital/year were lower when any of six clinical pharmacy services were present: drug use evaluation $1,119,810.18 (total $1,005,589,541.64 for the 898 hospitals offering the service), drug information $5,226,128.22 (total $1,212,461,747.04 for the 232 hospitals offering the service), adverse drug reporting monitoring $1,610,841.02 (total $1,101,815, 257.68 for the 684 hospitals offering the service), drug protocol management $1,729,608.41 (total $614,010,985.55 for the 355 hospitals offering the service), medical rounds participation $7,979,720.45 (total $1,212,917,508.41 for the 152 hospitals offering the service), and admission drug histories $6,964,145.17 (total $208,924,355.10 for the 30 hospitals offering the service). Clinical research $9,558,788.01 (total $1,013,231,529.06 for the 106 hospitals offering the service) and TPN team participation $3,211,355.12 (total $1,027,633,638.43 for the 320 hospitals offering the service) were associated with higher total costs of care. As staffing increased for hospital pharmacy administrators (p=0.0001) and clinical pharmacists (p=0.007), total cost of care decreased. As staffing increased for dispensing pharmacists, total cost of care increased (p=0.006). Based on this total cost of care model, optimal hospital pharmacy administrator staffing was 2.01/100 occupied beds. Staffing for dispensing pharmacists should be as low as possible, and definitely fewer than 5.11/100 occupied beds. Staffing for clinical pharmacists should be as high as possible, but definitely more than 1.11/100 occupied beds. The results of this study suggest that increased staffing levels of clinical pharmacists and pharmacy administrators, as well as some clinical pharmacy services, were associated with reduced total cost of care in United States hospitals. PMID- 10853616 TI - Gender differences in labetalol kinetics: importance of determining stereoisomer kinetics for racemic drugs. AB - STUDY OBJECTIVE: To evaluate the impact of gender on labetalol kinetics. DESIGN: Part of a randomized, crossover study. SETTING: Academic medical center. PATIENTS: Nineteen hypertensive patients (14 men, 5 women; 6 blacks, 13 whites). INTERVENTIONS: Participants had labetalol dosages titrated to a specific antihypertensive response, then underwent ambulatory blood pressure monitoring (ABPM) and a pharmacokinetic study. Labetalol plasma concentrations were measured by high-performance liquid chromatography (HPLC) and labetalol stereoisomer ratios were determined in a single plasma sample by chiral HPLC, both with fluorescence detection. MEASUREMENTS AND MAIN RESULTS: Labetalol concentrations were 80% higher in women (area under the concentration-time curve [AUC]/dose x 1000: 6.79 +/- 2.11 in women vs 3.82 +/- 1.37 hr/L in men, p<0.05), yet both genders had a similar antihypertensive response by 24-hour ABPM. Dose-corrected AUC (AUC/dose x 1000) for labetalol's stereoisomers in women and men, respectively, were S,R-labetalol 7.55 +/- 1.47 and 4.83 +/- 1.54 hr/L (p<0.05), S,S-labetalol 8.23 +/- 2.93 and 4.65 +/- 1.78 hr/L (p<0.05), R,S-labetalol 6.99 +/- 3.30 and 4.25 +/- 2.35 hr/L (p=0.11), and R,R-labetalol 3.91 +/- 2.57 and 3.55 +/- 3.08 hr/L (NS). CONCLUSION: The higher labetalol concentration in women than in men was explained largely by differences in inactive and alpha1-blocking stereoisomers. However, concentrations were similar between genders for the beta blocking stereoisomer (R,R-labetalol), possibly explaining the similarity in antihypertensive response to the drug. This study highlights the importance of determining stereoisomer kinetics for agents administered as racemates, particularly when relating concentrations to pharmacologic response. PMID- 10853617 TI - Incidence of first-time idiopathic seizures in users of tramadol. AB - STUDY OBJECTIVE: To assess the risk of idiopathic incident seizures among patients who ever took tramadol. DESIGN: Nested case-control design. SETTING: General Practice Research Database from November 1996-August 1998. PATIENTS: Eleven thousand three hundred eighty-three patients. INTERVENTION: Comparison of risks of idiopathic incident seizures during exposed and unexposed times among patients who had ever taken tramadol and other analgesics with 90-day follow-up. MEASUREMENTS AND MAIN RESULTS: Among the 11,383 subjects we identified 21 cases of idiopathic seizures, 10 of which were categorized as definite cases and 11 categorized as possible cases. Three patients were exposed to tramadol alone in the previous 90 days, 10 to opiates, three to both tramadol and opiates, one to other analgesics, and four to no analgesics. CONCLUSION: The risk of idiopathic seizures was similarly elevated in each analgesic exposure category compared with nonusers, suggesting that the risk for patients taking tramadol was not increased compared with other analgesics. PMID- 10853618 TI - Determinants of ceftriaxone clearance by continuous venovenous hemofiltration and hemodialysis. AB - STUDY OBJECTIVE: To guide individual ceftriaxone dosages in patients receiving continuous renal replacement therapy. DESIGN: Prospective, outpatient study. SETTING: University-affiliated general clinical research center. PATIENTS: Eight patients receiving hemodialysis. INTERVENTION: We performed controlled clearance studies with three hemofilters: an acrylonitrile copolymer 0.6 m2 (AN69), polymethylmethacrylate 2.1 m2 (PMMA), and polysulfone 0.65 m2 (PS). MEASUREMENTS AND MAIN RESULTS: Subjects received ceftriaxone 1000 mg intravenously before the start of a clearance study. The concentration of ceftriaxone in multiple plasma and dialysate-ultrafiltrate samples was determined by high-performance liquid chromatography. The diffusional clearances (Cl(diffusion)) and sieving coefficients (SC) of ceftriaxone, urea, and creatinine were compared by a mixed model repeated-measures analysis of variance with filter and blood, dialysate inflow, or ultrafiltration rate as the main effect and patient as a random effect. The fraction of ceftriaxone bound to plasma proteins was 43 +/- 15% (range 13-92%). Concentration dependence was evident in all three groups. The fraction unbound to plasma proteins (f(up)) at the time that SCs were determined was significantly lower in the PS group (0.16 +/- 0.07) than the AN69 group (0.30 +/- 0.17, p<0.01), but similar to that in the PMMA group (0.27 +/- 0.12). Despite the higher f(up), the SC of unbound ceftriaxone with the AN69 filter (0.48 +/- 0.13) was significantly lower than values for the PMMA (0.86 +/- 0.33) and PS (0.82 +/- 0.22) groups (p<0.05). Continuous venovenous hemofiltration clearance of urea and unbound ceftriaxone increased significantly only for the PMMA (p=0.006) and PS (p=0.015) filters when the ultrafiltration rate was increased. Significant linear relationships (p<0.0001) were observed between Cl(diffusion) of unbound ceftriaxone and clearance of urea for all three filters: AN69 slope = 0.57, PMMA slope = 0.90, and PS slope = 1.02. The slope of this relationship for the AN69 filter was significantly lower than for the other two filters. CONCLUSION: Ceftriaxone clearance was significantly increased and membrane dependent during continuous venovenous hemofiltration and continuous venovenous hemodialysis. Thus individual ceftriaxone dosages for patients receiving continuous renal replacement therapies should incorporate extracorporeal clearance. PMID- 10853619 TI - Caffeine citrate for the treatment of apnea of prematurity: a double-blind, placebo-controlled study. AB - STUDY OBJECTIVE: To evaluate the efficacy and safety of caffeine citrate for treatment of apnea of prematurity. DESIGN: Multicenter, parallel, randomized, double-blind, placebo-controlled trial with open-label rescue. SETTING: Nine neonatal intensive care units. PATIENTS: Eighty-five infants, 28-32 weeks postconception and 24 hours or more after birth who had six or more apnea episodes within 24 hours. INTERVENTION: Caffeine citrate 10 mg/kg (as caffeine base) administered intravenously, followed by 2.5 mg/kg/day orally or intravenously, or placebo, for up to 10 days. Infants failing double-blind therapy could receive open-label rescue. MEASUREMENTS AND MAIN RESULTS: Success was defined as 50% or greater reduction in apnea episodes and elimination of apnea. Caffeine citrate was significantly more effective than placebo in reducing apnea episodes by at least 50% in 6 days (p<0.05), and approached statistical significance (p<0.10) in 3 days. It was significantly better than placebo in eliminating apnea in 5 days (p<0.05), and approached significance (p<0.10) in 2 days. The number of infants with an aggregate of 7-10 days of at least a 50% reduction in apnea events or elimination of apnea was significantly higher in the caffeine citrate than in the placebo group. Adverse events did not differ significantly between groups. No correlations were found between success and mean daily plasma concentrations or baseline characteristics. Volume of distribution and clearance increased with weight, supporting weight-adjusted dosing of caffeine citrate. CONCLUSION: Caffeine citrate 10 mg/kg caffeine base (equivalent to 20 mg/kg caffeine citrate) intravenously followed by 2.5 mg/kg/day caffeine base (equivalent to 5 mg/kg/day caffeine citrate) either intravenously or orally for 10 days is safe and effective for treating apnea of prematurity in infants 28 32 weeks postconception. PMID- 10853620 TI - Vancomycin assay performance in patients with end-stage renal disease receiving hemodialysis. AB - STUDY OBJECTIVE: To compare the performance of polyclonal fluorescence polarization immunoassay (pFPIA) with that of enzyme-multiplied immunoassay technique (EMIT) in patients receiving vancomycin and hemodialysis. SETTING: Outpatient hemodialysis center. PATIENTS: Seven subjects with end-stage renal disease treated with hemodialysis 3 times/week with a cellulose triacetate hemodialyzer. INTERVENTION: Subjects received vancomycin 1000 mg intradialytically during the first study session and 750 mg every other hemodialysis session thereafter for 4 weeks. MEASUREMENTS AND MAIN RESULTS: Blood samples were obtained throughout the study, and vancomycin serum concentrations were determined by pFPIA and EMIT. The mean +/- SD difference (estimate of bias) between assays was -1.10 +/- 1.35 mg/L. The limits of agreement (mean difference +/- 1.96 x SD) between them were -3.80-1.60 mg/L. CONCLUSION: Our data suggest that the manufacturer's changes in the vancomycin pFPIA eliminated overestimation of drug concentrations in patients undergoing high-permeability hemodialysis. PMID- 10853621 TI - Comparison of the serum and intracellular pharmacokinetics of azithromycin in healthy and diabetic volunteers. AB - STUDY OBJECTIVE: To compare serum and intracellular pharmacokinetics of azithromycin in healthy volunteers and patients with diabetes. DESIGN: Open label, parallel study. SETTING: Clinical research center. SUBJECTS: Twelve patients with diabetes and 12 healthy volunteers. INTERVENTIONS: Subjects were given a single 500-mg dose of azithromycin followed by 250 mg/day for 2 days. Blood samples were obtained just before and after the third dose for up to 24 hours for serum and 168 hours for intracellular measurement of azithromycin. MEASUREMENTS AND MAIN RESULTS: Pharmacokinetic parameters were calculated by noncompartmental methods and compared with a t test. The groups did not differ in maximum concentration, time to maximum concentration, or area under the concentration-time curve in serum or polymorphonuclear cells (PMNs). Differences in the PMN:serum ratio were observed at the 24-hour time point (healthy 1209 +/- 432, diabetic 859 +/- 286, p=0.051). CONCLUSION: In general, the pharmacokinetics of azithromycin are comparable in diabetics and healthy volunteers. Accumulation of drug in macrophages was slightly lower in patients. PMID- 10853622 TI - A prospective evaluation of empiric versus protocol-based sedation and analgesia. AB - STUDY OBJECTIVE: To compare empiric and protocol-based therapies of sedation and analgesia in terms of pharmacologic cost, effects on mechanical ventilation and intensive care unit (ICU) stay, and quality of sedation and analgesia. DESIGN: Prospective study. SETTING: A 24-bed medical-surgical-neurologic ICU. PATIENTS: Seventy-two patients evaluated during empiric therapy and 86 during protocol based therapy. INTERVENTION: Assessment of data collected for 4 months before and 5 months after an evidence-based sedation and analgesia protocol was implemented. MEASUREMENTS AND MAIN RESULTS: Protocol adherence rate was 83.7%. The hourly cost (Canadian dollars) of sedation was less with protocol-based therapy ($5.68 +/- 4.27 vs $7.69 +/- 5.29, p<0.01) likely due to increased lorazepam use. Pharmacologic cost savings may be negated since sedation duration tended to be longer (122.7 +/- 142.8 vs 88.0 +/- 94.8 hrs, p<0.1) and extubation may have been delayed (61.6 +/- 97.4 vs 39.1 +/- 54.7 hrs, p=0.13) with protocol use. Duration of ICU stay after sedation was discontinued was not significantly different before and after protocol implementation. With the protocol, however, the percentage of modified Ramsay sedation scores representing discomfort decreased from 22.4 to 11% (p<0.001) and the percentage at a score of 4 increased from 17.2% to 29.6% (p<0.01). The percentage of modified visual analog measurements representing pain decreased from 9.6 to 5.9% (p<0.05) with the protocol. When data were stratified according to duration of sedation, the benefits and delayed extubation associated with protocol-based therapy were limited to patients requiring long-term sedation. CONCLUSION: Compliance with this protocol reduced drug costs and enhanced the quality of sedation and analgesia for patients requiring long-term sedation. Protocol-based therapy with lorazepam may have delayed extubation but did not delay ICU discharge. PMID- 10853623 TI - Comparative in vitro activity of vancomycin and levofloxacin in combination with rifampin against planktonic versus sessile cells of Staphylococcus epidermidis. AB - STUDY OBJECTIVE: To evaluate the activity of vancomycin and levofloxacin alone and combined with rifampin against planktonic and sessile cells. INTERVENTION: Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of the three drugs were determined against a clinical isolate of methicillin-resistant Staphylococcus epidermidis (MRSE 23) and a reference strain of MRSE (ATCC 35984). MEASUREMENTS AND MAIN RESULTS: The MICs/MBCs of vancomycin, levofloxacin, and rifampin against MRSE 23 were 0.78/0.78 microg/ml, 0.19/0.19 microg/ml or below, and 0.19/0.19 microg/ml or below, and against ATCC 35984 were 0.78/1.56 microg/ml, 0.19/0.19 microg/ml or below, and 0.19/0.19 microg/ml or below, respectively. A 99.9% killing activity was achieved with vancomycin, levofloxacin, and vancomycin-levofloxacin against planktonic cells of MRSE 23 (18.9, 21.3, and 17.5 hrs, respectively) and only with levofloxacin against ATCC 35984 (21.5 hrs). No regimen achieved 99.9% killing activity against sessile cells. CONCLUSION: Adding rifampin was antagonistic against planktonic cells and had an additive effect against sessile cells. Activity typically reported using nutrient-rich, planktonic cells may not be applicable to sessile cells under environmental and growth restrictions. PMID- 10853624 TI - The effect of beta-blockers on health-related quality of life in patients with heart failure. AB - Beta-blockers reduce the risk of death in patients with heart failure and are recommended in those with stable class II or III disease despite optimal standard therapy. Health-related quality of life (HRQOL) is an increasingly important end point in clinical trials. We reviewed all studies that determined the effect of beta-blockers on HRQOL in patients with heart failure. In these trials, HRQOL was assessed by the Quality of Life Questionnaire in Severe Heart Failure and the Minnesota Living with Heart Failure Questionnaire. Three of the 10 studies that used either of these instruments reported significant improvements in scores. When HRQOL was determined by a single-question global assessment, substantial improvements were observed by patients and physicians in five of the seven studies that used the instrument. Possible reasons for the lack of consistent effect on HRQOL include lack of responsiveness of currently available instruments, incomplete data collection, and true lack of effect of beta-blockers on HRQOL in these patients. PMID- 10853625 TI - Evaluation of echinacea for treatment of the common cold. AB - Considered to have immunostimulating activity, echinacea is a widely used phytomedicinal for treatment of the common cold and upper respiratory tract infections (URTIs). We reviewed the literature from the MEDLINE database (January 1966-July 1999), International Pharmaceutical Abstracts (IPA) online database, Cambridge Scientific Abstracts Biological Sciences online database, Alt-Health Watch online database, EMBase CD-ROM database, and references from published articles, reviews, and letters to evaluate evidence from clinical trials of echinacea's purported efficacy for treating or preventing URTIs. Twelve clinical studies published from 1961-1997 concluded that echinacea was efficacious for treating the common cold, but the results are unclear due to inherent flaws in study design. Five trials were published since 1997; two showed that echinacea lacked efficacy for treating and preventing URTI symptoms, and three concluded that it was effective in reducing the frequency, duration, and severity of common cold symptoms. Again, these results are unclear because of methodologic uncertainties, such as small populations and use of noncommercially available, nonstandardized dosage forms. Although evidence for echinacea's efficacy is inconclusive, it appears to be safe. Patients without contraindications to it may not be dissuaded from using an appropriate preparation to treat the common cold. PMID- 10853626 TI - Update on the interaction between aspirin and angiotensin-converting enzyme inhibitors. AB - We summarized recent published literature regarding the significance of an interaction between aspirin and angiotensin-converting enzyme (ACE) inhibitors in patients with various cardiovascular diseases. A MEDLINE search (January 1998 July 1999) was performed and abstracts from the 1999 American College of Cardiology and 1998 American Heart Association annual scientific sessions were reviewed to identify pertinent studies. Material for discussion was identified through a MEDLINE search from January 1996-July 1999 and through cited references. The results of several studies added to our understanding of the clinical ramifications of an aspirin-ACE inhibitor interaction, but also introduced questions. These studies are largely contradictory, but do reiterate the possibility of an interaction, if only in certain subsets of patients. Low dosages (< or = 100 mg/day) of aspirin appear to be safer in this regard than higher dosages. The frequency and severity of the interaction and possible predisposing factors await future research. PMID- 10853627 TI - The pharmacist's role in promoting optimal antimicrobial use. AB - Optimal use of antimicrobials is essential in the face of escalating antibiotic resistance, and requires cooperation from all sectors of the health care system. Although antibiotic-restriction policies in the hospital setting are important in altering microbial susceptibility patterns, an overall reduction in antibiotic prescriptions in the outpatient setting is more likely to significantly impact antibiotic resistance. Education of providers, application of clinical practice guidelines, audit and feedback activities, and multifaceted interventions all have had an effect in altering antibiotic prescribing in a research setting. Clinicians must alter antibiotic prescribing for the treatment of infectious diseases, and patients must change their perception of the need for these drugs. Pharmacists can play a major role through clinician education and focused clinical services. With cooperation of health care teams, the effectiveness of available antibiotics may be sustained and the threat of resistance minimized. PMID- 10853628 TI - Non-ergot dopamine agonist-induced sleep attacks. AB - Two patients with Parkinson's disease received treatment with ropinirole and/or pramipexole, during which both experienced sleep attacks. These attacks may be a class effect of non-ergot dopamine agonists. Health care professionals should be aware of the potential of these agents to cause sleep attacks and caution patients about this potentially life-threatening adverse effect. PMID- 10853629 TI - Use of fenofibrate in the management of protease inhibitor-associated lipid abnormalities. AB - Human immunodeficiency virus (HIV) protease inhibitors are associated with several metabolic abnormalities including hypercholesterolemia and hypertriglyceridemia. Fenofibrate is a new lipid-lowering agent for adults with very high triglyceride levels that was administered to two HIV-positive patients who were taking protease inhibitors and developed hypertriglyceridemia. Starting dosages were 134 and 201 mg/day, and were increased to 268 mg/day in both patients. Triglyceride levels decreased from 1450 to 337 mg/dl (76.8%) and from 1985 to 322 mg/dl (83.8%), respectively, after 10 months of therapy. High-density lipoprotein levels increased in both patients. PMID- 10853630 TI - Benzocaine-induced methemoglobinemia during an outpatient procedure. AB - Outpatient transesophageal echocardiography was performed in a 69-year-old man with a history of aortic valve repair. During the procedure the patient became markedly cyanotic and hypotensive. Oxygen saturation measured by pulse oximetry decreased from 97% to the mid-80s. The man's condition continued to deteriorate. On transfer to a critical care unit, blood analysis by co-oximetry showed methemoglobin saturation of 67.8%. The patient's condition improved significantly after intravenous administration of methylene blue 1 mg/kg. With increasing numbers of outpatient procedures performed under topical anesthesia, measures should be in place to deal with a potential life-threatening adverse event such as methemoglobinemia. PMID- 10853631 TI - Bias in the evaluation of low-magnitude associations: an empirical perspective. PMID- 10853632 TI - Invited commentary: shifting the burden of proof regarding biases and low magnitude associations. PMID- 10853633 TI - Antidepressant medication use and breast cancer risk. AB - Experimental and epidemiologic studies suggest that antidepressant medication use may be associated with breast cancer risk. This hypothesis was investigated using a population-based case-control study; cases diagnosed in 1995-1996 were identified using the Ontario Cancer Registry, and controls were randomly sampled from an Ontario Ministry of Finance database. Data were collected using a self administered questionnaire, and multivariate logistic regression was used to estimate odds ratios and 95% confidence intervals. Adjusted odds ratio estimates ranged from 0.7 to 0.8 and were not statistically significant for "ever" use of antidepressants, tricyclics, and selective serotonin reuptake inhibitors. Compared with no antidepressant use, use of tricyclic antidepressants for greater than 2 years' duration was associated with an elevated risk of breast cancer (odds ratio (OR) = 2.1, 95% confidence interval (CI): 0.9, 5.0). Of the six most commonly reported antidepressant medications, only paroxetine use was associated with an increase in breast cancer risk (OR = 7.2, 95% CI: 0.9, 58.3). Results from this study do not support the hypothesis that "ever" use of any antidepressant medications is associated with breast cancer risk. Use of tricyclic medications for greater than 2 years, however, may be associated with a twofold elevation, and use of paroxetine may be associated with a substantial increase in breast cancer risk. PMID- 10853634 TI - Prospective study of bowel movement, laxative use, and risk of colorectal cancer among women. AB - The authors prospectively examined the association between bowel movement frequency, laxative use, and the risk of colorectal cancer in 84,577 women of the Nurses' Health Study living in the United States, 36-61 years of age and free of cancer in 1982. Between 1984 and 1996, 611 incident cases of colorectal cancer were documented. After controlling for age, body mass index, fiber intake, postmenopausal status and hormone use, physical activity, and use of laxatives, the relative risks associated with having bowel movements every third day or less, compared with those with bowel movements once daily, were 0.94 (95% confidence interval (CI): 0.69, 1.28) for colorectal cancer, 0.88 (95% CI: 0.62, 1.26) for colon cancer, and 1.18 (95% CI: 0.63, 2.20) for rectal cancer. Compared with women who never used laxatives, the multivariate relative risks associated with weekly to daily laxative use were 1.00 (95% CI: 0.72, 1.40) for colorectal cancer, 1.09 (95% CI: 0.76, 1.57) for colon cancer, and 0.68 (95% CI: 0.29, 1.57) for rectal cancer. These findings do not support an association between infrequent bowel movement, laxative use, and risk of colorectal cancer and indicate that simple questions directed at bowel movement frequency are unlikely to enhance our ability to predict colorectal cancer risk. PMID- 10853635 TI - Methods to quantify the relation between disease progression in paired eyes. AB - The authors compared, in the context of diabetic retinopathy, alternative methods of quantifying the extent to which disease progression in one eye increases the risk of subsequent progression in the other eye. Data were gathered on 478 US patients with insulin-dependent diabetes mellitus who participated in the 1983 1988 Sorbinil Retinopathy Trial and were followed up for a median of 41 months. During that time, diabetic retinopathy progressed in 93 right eyes and 77 left eyes. Crude incidence rates of progression for right eyes were 7.7 times higher after the left eye had progressed and, for left eyes, were 4.4 times higher after the right eye had progressed. In eye-specific proportional hazards models that adjusted for increasing rates of progression over time and for baseline risk factors, the comparable relative risks associated with progression in the other eye were 2.6 (95% confidence interval (CI): 1.5, 4.7) for right eyes and 1.4 (95% CI: 0.72, 2.9) for left eyes. Two alternative proportional hazards models that included data on both eyes and accounted for their correlation produced estimated relative risks of 1.9 (95% CI: 1.2, 2.9) and 2.7 (95% CI: 1.8, 3.5), respectively. The more complex models for joint survival integrate information on both eyes and provide more stable estimates than do separate analyses of right or left eyes. PMID- 10853636 TI - Antioxidant nutrients and pulmonary function: the Third National Health and Nutrition Examination Survey (NHANES III). AB - Recent studies of chronic obstructive pulmonary disease have raised interest in its relation to nutrition. Several dietary antioxidants have been positively associated with lung function in healthy, general population samples. This study considered the separate and joint effects of vitamin C, vitamin E, beta-carotene, and selenium intake and used both dietary assessment and serum biomarkers of antioxidant status. The authors used data from the Third National Health and Nutrition Examination Survey comprising a sample representative of the US population in 1988-1994 (n = 18,162 subjects aged > or =17 years). Multiple linear regression analysis examined the separate and joint effects of the antioxidants on the ratio of forced expiratory volume in the first second (FEV1)/height2 adjusted for covariates. Each of the dietary and serum antioxidant nutrients was significantly associated with FEV1. When they were considered simultaneously (dietary and serum variables considered in separate models), independent associations were observed for most nutrients. Serum beta-carotene was less positively associated with FEV1 in smokers than nonsmokers, while serum selenium had a stronger positive association with FEV1 in smokers. The authors found that higher levels of antioxidant nutrients are associated with better lung function. The finding that the antioxidants differ in both their overall association with lung function and in whether this association varies by smoking status has implications for further research. PMID- 10853637 TI - Characterization of dust exposure for the study of chronic occupational lung disease: a comparison of different exposure assessment strategies. AB - Various exposure assessment strategies were compared in the study of the relation between dust exposure and 11-year lung function change in 1,172 miners with 36,824 concurrently measured personal dust samples available from the 1969-1981 US National Study of Coal Workers' Pneumoconiosis. A miner's average exposure was assessed by calculating average exposures based on dust samples taken from each individual and by using different job exposure matrices (JEMs) with different underlying exposure categorizations, based on occupational categories, job title, mine, and time, to obtain average exposure estimates. For each grouping procedure, intragroup and intergroup variances and the pooled standard error of the mean were calculated to assess relative efficiency. The results show that considerable variation in slopes of exposure-response relations was found using different exposure assessment strategies. Standard errors of the slopes of the exposure-response relations with exposure on an individual basis compared with JEMs. Exposure assessment on an individual basis was extremely sensitive to the number of exposure measurements per individual. The study demonstrates the advantages and disadvantages of different exposure assessment strategies and shows the need for explicit publication of exposure assessment strategies for epidemiologic studies. Careful assessment of the influence of misclassification error in the exposure assessment on exposure-response modeling is warranted. PMID- 10853638 TI - Investigation of design and bias issues in case-control studies of cancer screening using microsimulation. AB - Using a microsimulation approach, the authors examined design and bias issues in case-control studies of cancer screening. Specifically, they looked at the impact on the odds ratio of the way in which exposure to screening is defined, the type of age matching, the time scale used, and the criteria used to determine control eligibility. The results showed that defining exposure as "ever/never" screened produced, as expected, a serious bias in favor of screening. Defining exposure as being screened no later than the time the case's cancer is diagnosed has a serious bias against screening. An alternative exposure definition--screening can occur no later than the time the case would have been clinically diagnosed- eliminates the bias against screening. Further, the results showed that the type of age matching and the time scale used can produce a bias against screening and that this bias can be quite strong when case-control studies are performed in populations with a periodic screening program that is the only source of screening. Finally, control eligibility criteria had little effect. PMID- 10853639 TI - Fish consumption and coronary heart disease mortality in Finland, Italy, and The Netherlands. AB - Fish consumption seems to protect against death from coronary heart disease (CHD). If this association is due to n-3 polyunsaturated fatty acids, especially fatty fish may be responsible for this protective effect. The association between total, lean, and fatty fish consumption and the risk of CHD mortality was examined in 1,088 Finnish, 1,097 Italian, and 553 Dutch men participants in the Seven Countries Study who were aged 50-69 years and free of CHD around 1970. After 20 years of follow-up, 242 (22.2%) men in Finland, 116 (10.6%) men in Italy, and 105 (19.0%) men in the Netherlands had died of CHD. Cox proportional hazards analysis showed no association between total fish consumption and CHD mortality. After adjustments were made for age, body mass index, smoking, energy intake, and relevant dietary variables, the pooled relative risk for the highest quartile of total fish compared with no fish consumption in the three countries was 1.08 (95% confidence interval: 0.76, 1.53). Lean fish consumption also was not associated with CHD mortality in any country. Fatty fish compared with non fatty-fish consumption was associated with lower CHD mortality; the adjusted, pooled relative risk for fatty fish consumers was 0.66 (95% confidence interval: 0.49, 0.90). These data suggest that especially fatty fish is protective against CHD mortality. PMID- 10853640 TI - Blood pressure differences between blacks and whites in relation to body size among US children and adolescents. AB - No large national studies of ethnic differences in blood pressure among children accounting for body size differences have been published, to the authors' knowledge. This report details the similarities and differences in systolic and diastolic blood pressures between Black children and White children in the United States and examines the effects of age, sex, and body size on ethnic differences in blood pressure levels. Standardized measurements of seated systolic and diastolic pressures from eight large epidemiologic studies published between 1978 and 1991 that included measurements of 47,196 children on 68,556 occasions for systolic pressure and for 38,184 children on 52,053 occasions for diastolic pressure were used; 51 percent (24,048 children) were boys and 37 percent (17,466 children) were Black. Overall, there appear to be few substantive ethnic differences in either systolic or diastolic pressure during childhood and adolescence. The differences that were observed were small, inconsistent, and often explained by differences in body size. There was an ethnic group-body mass index (BMI) interaction that resulted in these findings that at lower levels of BMI Blacks have higher blood pressure and more hypertension than do Whites, but that at the highest levels of BMI, Whites have more hypertension (systolic or diastolic pressure) than do Blacks. PMID- 10853641 TI - Duration of human immunodeficiency virus infection and likelihood of giving birth in a Medicaid population in Maryland. AB - The objective of this study was to examine the effect of duration of human immunodeficiency virus (HIV) infection on a woman's likelihood of giving birth. Using longitudinal data from the Maryland state Human Immunodeficiency Virus Information System and a retrospective cohort design, the authors compared 1,642 women with acquired immunodeficiency syndrome (AIDS) to 8,443 uninfected women enrolled in the Medicaid program between 1985 and 1995. The decade before AIDS diagnosis was divided into four 2.5-year periods. Proximity to AIDS diagnosis served as a proxy for duration of infection. An extension of the Cox model was used to estimate the relative risk for giving birth, with adjustment for covariates and repeated outcomes. The average number of births per 100 person years was 6.0 for HIV-infected women and 11.1 for uninfected women (adjusted relative risk = 0.63; 95% confidence interval (CI): 0.57, 0.68). Accounting for duration of infection, the adjusted relative risks for birth among HIV-infected women, as compared with uninfected women, were 0.85 (95% CI: 0.71, 1.03), 0.74 (95% CI: 0.63, 0.86), 0.55 (95% CI: 0.47, 0.64), and 0.45 (95% CI: 0.38, 0.55) for successive 2.5-year periods before AIDS diagnosis. Demographic characteristics, contraception, abortion, fetal loss, or drug use could not fully explain the reductions. These results suggest that HIV-infected women experience a progressive reduction in births years before the onset of AIDS. This may compromise estimation of HIV prevalence and interpretation of time trends from serosurveillance of pregnant women. PMID- 10853642 TI - Predictors of correct treatment of children with fever seen at outpatient health facilities in the Central African Republic. AB - To identify factors associated with improved performance of health care workers who treat ill children in developing countries, the authors analyzed a sample of consultations of children with malaria (defined as any fever) from a national health facility survey conducted in the Central African Republic from December 1995 to January 1996. Twenty-eight health care workers and 204 children were studied. A univariate analysis revealed the following significant predictors of correct treatment, as defined by the Central African malaria control program: high fever (odds ratio (OR) = 3.25, 95% confidence interval (CI): 1.47, 7.17); correct health care worker diagnosis (OR = 2.59, 95% CI: 1.39, 4.85); and the caregiver's reporting the child's fever to the health care worker (OR = 2.18, 95% CI: 1.32, 3.62). There was an unexpected inverse association between the presence of a fever treatment chart and correct treatment (OR = 0.19, 95% CI: 0.04, 0.91). Correct treatment was marginally associated with a longer consultation time (p value for trend = 0.058). Neither in-service training in the treatment of fever nor supervision was significantly associated with correct treatment. For child health programs to improve, targeted studies are needed to understand which factors, alone or in combination, improve health care worker performance. PMID- 10853643 TI - Re: "invited commentary: circular epidemiology". PMID- 10853644 TI - SAR studies of 2-o-substituted-benzoyl- and 2-alkanoyl-cyclohexane-1,3-diones as inhibitors of 4-hydroxyphenylpyruvate dioxygenase. AB - Inhibition studies of 4-hydroxyphenylpyruvate dioxygenase (HPPD) with various synthesized 2-o-substituted-benzoyl- and 2-alkanoyl-cyclohexane-1,3-diones suggest that the presence of a strongly electronegative group at the ortho position and the conformation of the benzene ring moiety on the benzoylcyclohexane-1,3-dione inhibitors are crucial for potent HPPD inhibition. PMID- 10853645 TI - The synthesis and evaluation of 2-substituted-7-(alkylidene)cephalosporin sulfones as beta-lactamase inhibitors. AB - A series of 2-substituted-7-(alkylidene)cephalosporin sulfones were prepared and evaluated as beta-lactamase inhibitors. Compound 11c showed excellent activity as an inhibitor of the class C beta-lactamase derived from Enterobacter cloacae, strain P99. PMID- 10853646 TI - The synthesis and evaluation of 3-substituted-7-(alkylidene)cephalosporin sulfones as beta-lactamase inhibitors. AB - A series of 3-substituted-7-(alkylidene)cephaloporin sulfones were prepared and evaluated as inhibitors of representative class A and class C serine beta lactamase. Appropriate substituents resulted in a 1000-fold improvement in the inhibition of the class A enzymes and a simultaneous 20-fold improvement in the inhibition of class C. These new compounds have achieved the goal of creating broad scale inhibitors in the cephalosporin series. PMID- 10853647 TI - Three-dimensional quantitative structure-activity relationship analyses of RGD mimetics as fibrinogen receptor antagonists. AB - In order to better understand the structural requirements of fibrinogen receptor antagonists, variations in the platelet aggregation inhibitory activity of a series of RGD mimetics were examined using techniques for the analysis of three dimensional quantitative structure activity relationship, such as CoMFA. PMID- 10853648 TI - Synthesis of NF-kappaB activation inhibitors derived from epoxyquinomicin C. AB - In order to develop new inhibitors of NF-kappaB activation, we designed and synthesized dehydroxymethyl derivatives of epoxyquinomicin C, namely, DHM2EQ and its regioisomer DHM3EQ. These derivatives were synthesized from 2,5 dimethoxyaniline in 5 steps. Since DHM2EQ was more active and less toxic than DHM3EQ, its stereochemical configuration was determined by X-ray crystallographic analysis. Each enantiomer of the protected DHM2EQ was separated by a chiral column and deprotected. DHM2EQ inhibited TNF-alpha-induced activation of NF kappaB in human T cell leukemia cells, and also inhibited collagen-induced arthritis in a rheumatoid model in mice. PMID- 10853649 TI - Quinonic derivatives active against a virulent strain of Toxoplasma gondii. Synthesis of 2-methylfuro[2,3-g]- and [3,2-g]isoquinolinetriones. AB - 2-methylfuro[2,3-g]isoquinoline-4,7,9-trione (4) and 2-methylfuro[3,2 g]isoquinoline-4,6,9-trione (5) were prepared regiospecifically from 2-azadiene 9 and bromobenzofuran-4,7-diones 1 or 11. The activity of these two compounds and some other quinonic derivatives was evaluated in vitro against a virulent strain of Toxoplasma gondii. Compounds 4 and 7 were found to be as active as pyrimethamine. PMID- 10853650 TI - Novel phenylpiperazine derivatives as dual cytokine regulators with TNF-alpha suppressing and IL-10 augmenting activity. AB - Phenylpiperazine derivatives were synthesized as dual cytokine regulators with TNF-alpha suppressing and IL-10 augmenting activity. Lead optimization led to compound 5k having the potent regulatory activity and demonstrating remarkable protective effects against the lethal challenge of LPS in mice. suggesting that 5k would be a promising drug candidate for the treatment of TNF-alpha associated diseases including septic shock. PMID- 10853651 TI - Novel DMARDs on the basis of a new concept of dual cytokine regulation, TNF-alpha suppression and IL-10 augmentation. AB - A series of arylpiperazine derivatives was synthesized to obtain agents showing apparent therapeutic effects in a chronic inflammatory animal model, starting from a lead possessing potent dual cytokine regulatory activity in vivo. We found a pyrimidylpiperazine derivative 17c showing the dual regulatory activity and an excellent therapeutic effect in an adjuvant-induced arthritis model. PMID- 10853652 TI - Thiol-dependent DNA cleavage by 3H-1,2-benzodithiol-3-one 1,1-dioxide. AB - Hydrodisulfides (RSSH) have previously been implicated as key intermediates in thiol-triggered oxidative DNA damage by the antitumor agent leinamycin. In an effort to better understand DNA damage by RSSH and to expand on the number and type of chemical systems that produce this reactive intermediate, the ability of 3H-1,2-benzodithiol-3-one 1,1-dioxide (11) to serve as a thiol-dependent DNA cleaving agent has been investigated. The findings reported here indicate that reaction of 11 with thiols results in release of RSSH and subsequent oxidative DNA strand cleavage. PMID- 10853653 TI - Synthesis and evaluation of vitamin D-based cationic lipids for gene delivery in vitro. AB - A new panel of steroidal cationic lipids has been synthesized for gene delivery. Using commercially available vitamin D2 (calciferol) or vitamin D3 (cholecalciferol) as hydrophobic motifs and a variety of cationic head groups as binding sites for negatively charged phosphate groups in DNA, we demonstrated that the transfection activity of the synthetic vitamin D-based cationic lipids 1d, 2d formulated with dioleoylphosphatidylethanolamine (DOPE) as a co-lipid is comparable to that of 3-(-[N-N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC Chol). These synthetic lipids are effective in transfecting a variety of cell lines. These results suggest that vitamin D-based cationic lipids are useful transfection reagents for in vitro gene transfer studies. PMID- 10853654 TI - Synthesis and endothelin receptor binding activity of synthetic analogues of RES 1149-2. AB - A direct synthesis of analogues 6 and 7 is described. The key transformations are the addition of dibromomethyl lithium to a ketone and the subsequent mild hydrolysis to a hemiacetal. PMID- 10853655 TI - Structural analogues of some highly active non-competitive AMPA antagonists. AB - Some 5-methyl analogues (14a-e) of the non-competitive AMPA antagonists 3 acylated 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-4,5-dihydro-3H-2,3-benzodi azepines (2,3) have been synthesized. Generally they show diminished or low biological activity but two derivatives (14a,b) reveal effects comparable to those of GYKI 52466 (1), the prototype non competitive AMPA antagonist. PMID- 10853656 TI - 5-Thienyltryptamine derivatives as serotonin 5-HT1B/1D receptor agonists: potential treatments for migraine. AB - A series of 5-(2- or 3-thienyl)tryptamine derivatives (9) has been synthesized and shown to be potent and selective 5-HT1D versus 5-HT1B receptor agonists and, therefore, potential treatments for migraine. PMID- 10853657 TI - Incorporation of 4-thiothymidine into DNA by the Klenow fragment and HIV-1 reverse transcriptase. AB - The 5'-triphosphate of 4-thiothymidine (4S-TTP) is an excellent substrate for the Klenow fragment of Escherichia coli DNA polymerase 1 and HIV-1 reverse transcriptase with values of k(cat)/Km within a factor of approximately 3 of those for TTP. A large UV change (deltaepsilon= -9770 M(-1)cm(-1) at 340 nm) associated with incorporation of 4S-TMP into nucleic acid duplexes makes possible a rapid, continuous spectrophotometric assay of the reaction progress. PMID- 10853658 TI - Non-electrostatic complexes with DNA: towards novel synthetic gene delivery systems. AB - We have developed new DNA complexing amphiphile based on Hoechst 33258 interaction with DNA grooves. The synthesis and physicochemical characterisation of lipid/DNA complexes, as compared to that of classical lipopolyamine for gene delivery, are described and discussed. PMID- 10853659 TI - A one-pot multistep approach to alpha-azido-phosphonate and phosphonothioate diesters: key intermediates in the synthesis of haptens for the generation of antibody ligases. AB - A four-step, one-pot synthesis of mixed alpha-azido-phosphonates and phosphonothioates 12a-d is described. This chemistry has provided a facile route to haptens 6a b and 7 that have been employed for the elicitation of antibody ligases. Five hapten-specific antibodies have been identified as modest catalysts of a model peptide ligation reaction between thioester 1b and thiol 2 to give the amide product 5. PMID- 10853660 TI - Pyrrolo[3,2,1-ij]quinoline derivatives, a 5-HT2c receptor agonist with selectivity over the 5-HT2a receptor: potential therapeutic applications for epilepsy and obesity. AB - A series of pyrrolo[3,2,1-ij]quinoline derivatives was synthesized, evaluated for their activity against the 5-HT2c and 5-HT2a, receptors and found to be agonists at 5-HT2c with selectivity over 5-HT2a. PMID- 10853661 TI - Examination of novel non-phosphorus-containing phosphotyrosyl mimetics against protein-tyrosine phosphatase-1B and demonstration of differential affinities toward Grb2 SH2 domains. AB - Inhibitory potencies were compared of several mono- and dicarboxy-based pTyr mimetics in Grb2 SH2 domain versus PTP1B assays. Although in both systems pTyr residues provide critical binding elements, significant differences in the manner of recognition exist between the two. This is reflected in the current study, where marked variation in relative potencies was observed between the two systems. Of particular note was the poor potency of all monocarboxy-based pTyr mimetics against PTP1B when incorporated into a hexapeptide platform. The recently reported high PTP1B inhibitory potency of similar phenylphosphate mimicking moieties displayed in small molecule, non-peptide structures, raises questions on the limitations of using peptides as platforms for pTyr mimetics in the discovery of small molecule inhibitors. PMID- 10853662 TI - Synthesis and hybridization properties of polyamide based nucleic acid analogues incorporating pyrrolidine-derived nucleoamino acids. AB - Ndelta-Fmoc protected nucleoamino acids of type I (Base = T, C, A) have been synthesized and employed as building blocks for the construction of novel polyamide based nucleic acid analogues. Homopyrimidine oligomer A binds to complementary RNA with significant affinity and in a sequence-specific fashion, while no binding was observed to complementary DNA. PMID- 10853663 TI - New quinolinic derivatives as centrally active antioxidants. AB - A series of new 1,2-dihydro and 1,2,3,4-tetrahydroquinolines, synthesized from the corresponding propargylaniline intermediates, have been developed as antioxidants for the potential treatment of pathologies implicating central oxidative stress. PMID- 10853664 TI - Inhibition of neuraminidase with neuraminic acid C-glycosides. AB - Neuraminic (sialic) acid based alpha-C-glycosides have been synthesized and their inhibitory activity towards bacterial neuraminidase (sialidase) was examined. While some C-glycosides were found to be potent inhibitors (Ki 15-30 microM) of this neuraminidase, others afforded no measurable activity. The structure activity relationship of these C-glycosides is discussed in the context of other previously reported sialidase inhibitors. PMID- 10853665 TI - Substituted 5,7-diphenyl-pyrrolo[2,3d]pyrimidines: potent inhibitors of the tyrosine kinase c-Src. AB - 5,7-Diphenyl-pyrrolo[2,3d]pyrimidines represent a new class of highly potent inhibitors of the tyrosine kinase c-Src (IC50 < 50 nM) with specificity against a panel of different tyrosine kinases. The substitution pattern on the two phenyl rings determines potency and specificity and provides a means to modulate cellular activity. PMID- 10853666 TI - A simple, solid-phase binding assay for the nuclear import receptor karyopherin alpha. Part 1: direct binding. AB - The nuclear import receptor karyopherin alpha recognizes nuclear localization signals (NLSs), peptides that direct the transport of proteins into the nucleus. A simple, colorimetric assay has been developed to facilitate the identification and comparison of karyopherin ligands by direct and competitive binding using NLSs immobilized on the solid phase (TentaGel resin). PMID- 10853667 TI - A simple, solid-phase binding assay for the nuclear import receptor karyopherin alpha. Part 2: competitive binding. AB - A qualitative assay for the evaluation of soluble ligands of the nuclear import receptor karyopherin alpha has been developed. The assay relies on competition with an immobilized ligand, the nuclear localization signal (NLS) from nucleoplasmin, for binding to the receptor, which is detected by an enzyme-linked colorimetric method. PMID- 10853668 TI - Design, synthesis, and in vitro biological activity of indole-based factor Xa inhibitors. AB - A series of indole and carbazole based inhibitors of factor Xa (FXa) has been investigated. The most potent compound inhibits FXa with a Ki of 0.2 nM and has 900- and 750-fold selectivity over thrombin and trypsin, respectively. PMID- 10853669 TI - Design, synthesis, and in vitro biological activity of benzimidazole based factor Xa inhibitors. AB - Inhibitors based on the benzimidazole scaffold showed subnanomolar potency against Factor Xa with 500-1000-fold selectivity against thrombin and 50-100-fold selectivity against trypsin. The 2-substituent on the benzimidazole ring had a strong impact on the FXa inhibitory activity. Crystallography studies suggest that the 2-substituent may have a conformational effect favoring the extended binding conformation. PMID- 10853670 TI - New fluorogenic substrate for the first continuous steroid sulfatase assay. AB - The screening for new inhibitors of steroid sulfatase requires an efficient test system. To overcome the shortcomings of the available discontinuous fluorimetric assay, several coumarin-type compounds were investigated as potential new substrates. 3,4-Benzocoumarin 7-O-sulfate was found to have appropriate substrate properties for the establishment of the first direct continuous assay of steroid sulfatase. PMID- 10853671 TI - Design, synthesis, and biological activity of a cyclic peptide: an inhibitor of HIV-1 tat-TAR interactions in human cells. AB - Replication of human immunodeficiency virus type 1 (HIV-1) requires specific interactions of Tat protein with the transactivation responsive region (TAR) RNA, a 59-base stem-loop structure located at the 5'-end of all HIV mRNAs. A number of cyclic peptides are known to possess antibiotic activity and increased biological stability. Here we report the design, synthesis, and biological activity of a cyclic peptide (2), which inhibits transcriptional activation by Tat protein in human cells with an IC50 of approximately 40 nM. Cyclic peptides that can target specific RNA structures provide a new class of small molecules that can be used to control cellular processes involving RNA-protein interactions in vivo. PMID- 10853672 TI - Stereo-random synthesis of highly functionalized proline analogues by azomethine cycloaddition. AB - Highly substituted proline analogues were synthesized on Wang-resin bearing bisprotected histidine as starting material. The proline analogues (1,5 diazabicyclo[3.3.0]octane-2-carboxylic acid) were generated by 1,3-dipolar cycloaddition of azomethine ylides with maleimides, thus creating a library of maximum stereochemical diversity. Every compound set with the same empirical formula can theoretically consist of four diastereomers and can be tested in biological assays as mixture. Additionally different methods for the acylation of the proline nitrogen were evaluated. PMID- 10853673 TI - Beta-carbonyl substituted glutathione conjugates as inhibitors Of O. volvulus GST2. AB - A series of beta-carbonyl substituted glutathione conjugates were prepared and evaluated as inhibitors of OvGST2. Their specificity for the parasite derived protein was assessed through comparison with their inhibition of human piGST. Inhibition of OvGST2 has been demonstrated at low micromolar concentrations for these conjugates and selectivity for OvGST2 over human pi-GST of greater than 10 fold has been achieved. PMID- 10853674 TI - Synthesis and biological activity of peptidyl aldehyde urokinase inhibitors. AB - Solid- and solution-phase synthesis of peptidomimetic inhibitors of urokinase type plasminogen activator based on the sequence dSerAlaArg-al are described. The biological activities of these unique inhibitors are reported herein. Carbonate prodrugs were prepared and tested as potential drug delivery systems. PMID- 10853675 TI - Copper-promoted overall transformation of 4-tert-butylphenol to its para hydroxyquinonic derivative, 2-hydroxy-5-tert-butyl-1,4-benzoquinone. Biomimetic studies on the generation of topaquinone in copper amine oxidases. AB - Topaquinone (TPQ) is a cofactor present at the active site of copper amine oxidases, derived from a Tyr residue inserted in the polypeptide chain through a copper-dependent but otherwise largely unknown mechanism. A simple model system was developed that permits to obtain the overall transformation of 4-tert butylphenol, chosen as a model for Tyr, into a TPQ-like, para-hydroxyquinonic structure in the presence of Cu(II)-imidazole mononuclear complexes. PMID- 10853676 TI - Discovery and evaluation of potent, cysteine-based alpha4beta1 integrin antagonists. AB - Acyclic, disulphide derivatives of cysteine have been identified as moderately potent antagonists of alpha4beta1-mediated leukocyte cell adhesion to VCAM. This communication describes how they were discovered from a simple L-cystine derivative and using the structure-activity data of C*DThioPC* related cyclic peptides. PMID- 10853677 TI - Discovery and evaluation of potent, tyrosine-based alpha4beta1 integrin antagonists. AB - Using disulphide cysteine-based inhibitors as lead structures, this communication describes our strategy for identifying more stable, potent antagonists of the alpha4beta1 integrin. These studies ultimately discovered potent, low molecular weight inhibitors based on D-thioproline-L-tyrosine. PMID- 10853678 TI - Preoperative imaging of the carotid bifurcation. Current trends. AB - BACKGROUND: There is disagreement about the most appropriate imaging examination necessary for the preoperative assessment of the carotid bifurcation. Our objective was to find out the preferences of clinicians on this issue in one large hospital. And to determine whether and how these preferences have changed over time. METHODS: STUDY DESIGN: Observational, retrospective study. SETTING: Large metropolitan and university affiliated hospital. Numbers of patients per year who underwent carotid endarterectomy during 1990-1998. Numbers of patients per year who had conventional catheter carotid angiography during the same period. Assessment of the nature of preoperative carotid imaging in a sample of 400 patients (100 each in 1990, 1993, 1996 and 1998). Analysis of the types of imaging examinations by year and determination of underlying trends. MEASURES: Number of carotid angiograms expressed as a percentage of carotid endarterectomies performed each year. Types of preoperative imaging examinations of the carotids and changes over time. RESULTS: The number of preoperative conventional catheter angiograms decreased over time. In 1990 angiography was performed in 86% of carotid endarterectomies. In 1998 the proportion decreased to 16% (p<0.05). Conversely, the proportion of endarterectomies carried out based solely on ultrasonography increased from 6% in 1990 to 56% in 1998 (p<0.05). The proportion of endarterectomies performed based on the combined findings of ultrasound and magnetic resonance angiography increased from 3% in 1990 to 56% in 1996 and to 26% in 1998. Computed tomoangiography has not become popular. The observed reduction in the number of preoperative conventional carotid angiograms was independent of the presence/absence of symptoms, the level of serum creatinine, the subspecialty of the surgeon (vascular surgery vs neurosurgery) and the individual surgeon involved. CONCLUSIONS: In one large university affiliated hospital the trend in the preoperative imaging of the carotid arteries is moving away from conventional catheter angiography. There is increasing application of ultrasound combined with magnetic resonance angiography and a more pronounced trend towards the performance of carotid endarterectomy based only on ultrasonography. PMID- 10853679 TI - Exercise training in intermittent claudication. AB - BACKGROUND: Peripheral arterial occlusive disease (PAOD) at II stage results in a moderate to severe impairment in walking ability. Aim of this study, controlled and randomized, was to evaluate the efficacy of an intensive 4 weeks exercise training in PAOD followed by a six-month period and to analyse the risk factors for atherosclerosis and the site of the lesion for possible predictors of result outcome. METHODS: Patients with PAOD were included in the study (ankle/arm ratio < or = 0.7 and < or = 0.5 after exercise) with initial claudication distance (ICD) < or = 200 m and absolute claudication distance (ACD) < or = 500 m evaluated on a constant-load treadmill test (3 km/hr, 0% slope). Forty patients were randomized (all with antiplatelet therapy): 20 to a supervised walking exercise (mean ICD 121.8 m, ACD 289.7 m) and 20 to a non exercising control group (ICD 111.6 m, ACD 230.1 m). Both groups were tested at 4 weeks (T1) and 6 months (T2). Training group was enrolled in a 4-week supervised training program. RESULTS: In the training group 10% of patients became asymptomatic (>1000 m). At T1 ICD increased 141% (p<0.001) and ACD was with low-pain-claudication >1000 m in 50%, at T2 ICD was 200% (p<0.05) with 70% of asymptomatic for ICD and ACD. Control group has a no significant increase of ICD and ACD at T1 and T2. Only arterial hypertension and intermittent claudication severity emerged as negative predictive factors for the results of training. CONCLUSIONS: Vascular training produces a significant and lasting improvement in walking distance in PAOD. PMID- 10853680 TI - Treadmill exercise-induced release of endothelin-1 in patients with peripheral arterial occlusive disease at Fontaine stage IIb. AB - BACKGROUND: Endothelin-1 (ET-1) is an endothelial vasoconstrictor mitogenic peptide which is thought to be a marker of endothelial damage and a potential participant in the pathophysiological processes of the development of atherosclerotic lesions and disease states associated with vasoconstriction and vasospasm. METHODS: To investigate the endothelin-1 release in response to dynamic exercise in patients with peripheral arterial occlusive disease (PAOD), plasma concentrations were determined by radioimmunoassay in 16 patients (14 men, 2 women, mean age 56.2 +/- 8.1 years) with peripheral arterial occlusive disease at Fontaine stage IIb and in 10 control subjects (8 men, 2 women, mean age 58.1 +/- 7.2 years) in normal health during treadmill testing (slope 5%, speed 3 km/hr). Blood samples were collected at rest from an antecubital vein, at the onset of claudication pain, and 10 minutes after exercise. RESULTS: Mean plasma endothelin-concentrations during the stress test increased significantly in the patients with arterial disease, rising from basal values of 4.4 +/- 0.6 pmol/L to values of 8.9 +/- 0.7 pmol/L at the end of the test (p < 0.0001), whereas it did not change significantly in control subjects (rising from 2.6 +/- 0.4 pmol/L to 2.7 +/- 0.5 pmol/L). Further, plasma endothelin- in the patients with arterial disease was at all times higher than in the control subjects (p < 0.0001). CONCLUSIONS: In conclusion, this study shows that in patients with peripheral arterial occlusive disease, plasma endothelin-1 increases after treadmill exercise performed until claudication pain supervenes. Raised endothelin-1 could be a marker of ischaemic acute endothelial damage and/or could contribute to increase the vascular resistance in ischaemic limbs of these patients during dynamic exercise by promoting arterial/arteriolar vasoconstriction or vasospasm. PMID- 10853681 TI - Prospective study of transcutaneous oxygen tension (TcPO2) measurement in the testing period of spinal cord stimulation in diabetic patients with critical lower limb ischaemia. AB - BACKGROUND: Spinal cord stimulation improves microcirculatory blood flow, relieves diabetic neuropathic and ischaemic pain and reduces the amputation rate in patients with severe peripheral arterial occlusive disease. AIM: To evaluate whether transcutaneous oxygen tension (TcPO2) measurements can be used as a specific prognostic parameter in the assessment of suitability for permanent device implantation in a prospective controlled study on diabetic patients with peripheral arterial occlusive disease. METHODS: Sixty patients (39 men, 21 women; mean age: 60 years; range: 46-75) were submitted to implantation of a spinal cord electrical generator for severe peripheral vascular disease, after failed conservative or surgical treatment. The clinical status was classified as Fontaine's stage III and IV and the main pathology was diabetic vascular disease. Pedal TcPO2 was assessed on the dorsum of the foot and ankle and toe pressure Doppler measurements were performed before, two weeks and four weeks after implantation. RESULTS: Pain relief of over 75% and limb salvage were achieved in 35 diabetic patients, while in 12 a partial success with pain relief over 50% and limb salvage for at least 6 months was obtained. In 13 patients the method failed and the affected limbs were amputated. Clinical improvement and spinal cord stimulation success were associated with increases of TcPO2, within the first two weeks after implantation (temporary period). Limb salvage was achieved with significant increase of TcPO2 within the first two weeks of the testing period (from 21.4 to 31.5 mmHg in rest pain patients, p=0.030, from 15.1 to 22.0 mmHg, p=0.030 in patients with trophic lesions under 3 cm2 in size and in those with trophic lesions over 3 cm2, from 12.1 to 17.9 mmHg, p=0.025) unrelated to the stage of the disease and the initial TcPO2 value. TcPO2 changes were related to the presence of adequate paraesthesias and warmth in the painful area during the trial period. The systolic ankle/brachial blood pressure index and toe pressure did not change under stimulation. CONCLUSIONS: A two-week testing period should be performed in all diabetic patients treated with spinal cord stimulation for peripheral arterial occlusive disease to identify the candidates for permanent implantation. Only diabetic patients with significant increases of TcPO2 and clinical improvement, during the test period, should be considered for permanent implantation and not merely all patients with pain relief. TcPO2 changes could be used as a predictive index of the therapy success and should be considered in terms of cost effectiveness before the final decision to permanent implantation. PMID- 10853682 TI - Vascular endothelial growth factor- and platelet-derived growth factor angiogenesis depressed but fetal bovine serum-angiogenesis enhanced choroidal tissue cultures of streptozotocin-diabetic Wistar and GK rats. AB - BACKGROUND: Diabetic state-induced alterations of angiogenic activity of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) were compared with that of fetal bovine serum (FBS) in the cultured choroidal explants of streptozotocin (STZ)-diabetic Wistar and diabetic GK rats. METHODS: Choroidal explants (0.04-1.0 mm2) were isolated from rat eyeballs and cultured in fibrin gels with FBS-Dulbecco's modified Eagle's medium (0.5 mL) containing antibiotics and 300 microg/mL epsilon-amino caproic acid in the presence of recombinant mouse vascular endothelial growth factor and recombinant human platelet-derived growth factor BB at 37 degrees C under 5% CO2 and 95% air. Microvessels newly budded from these choroidal explants were photographed. The number and length of all microvessels per choroidal explant were counted and measured as indices of angiogenesis in vitro. RESULTS: Fetal bovine serum (5-10%) enhanced both angiogenic indices in the explants of STZ-diabetic Wistar and GK rats. The actions of the serum on both angiogenic indices in both diabetic rats were greater than those in age-matched normal rats. Vascular endothelial growth factor (3-30 ng/mL) with 1% fetal bovine serum increased the angiogenic indices in diabetic choroids, but was less pronounced than in normal choroids. The action of the growth factor (2.5 ng/mL) on angiogenesis was also less in diabetic choroids. CONCLUSIONS: Results suggest that the diabetic state may down-regulate the receptors for vascular endothelial and platelet-derived growth factors and/or desensitize their post-receptor signaling in the vascular endothelial cells of choroids, being inexplicable for the enhanced actions of fetal bovine serum on angiogenesis in diabetic choroids. PMID- 10853683 TI - Effects of bunazosin hydrochloride sustained-release formulation on cerebral circulation. AB - BACKGROUND: We investigated the cerebral blood flow in mild to moderately hypertensive patients with chronic cerebral infarction before and after the administration of bunazosin hydrochloride sustained-release formulation, a selective sympathetic alpha1 receptor blocker. METHODS: Eleven mild to moderately hypertensive patients (mean age 65.6 years) with chronic cerebral infarction were studied. INTERVENTIONS: The patients were on enalapril maleate, an angiotensin converting enzyme inhibitor, for one week and then enalapril maleate was switched to bunazosin hydrochloride sustained-release formulation. MEASURES: The cerebral blood flow study was performed before and 8 weeks after starting the administration of bunazosin hydrochloride sustained-release formulation. Cerebral blood flow was measured using the stable xenon CT method. The picture analysis was performed using AZ-7000. The regional cerebral blood flow was measured by placing the region of interest on the CT images. The regional cerebral blood flows were measured before and 20 minutes after intravenous injection of 17 mg/kg acetazolamide. RESULTS: The blood flows in the parietal cortex and caudate nucleus 8 weeks after starting the administration of bunazosin hydrochloride sustained-release formulation were significantly greater than those before. The cerebrovascular acetazolamide reactivity in the occipital cortex and caudate nucleus was significantly lower after switching to bunazosin hydrochloride sustained-release formulation than before. CONCLUSIONS: Considering the reports that angiotensin converting enzyme inhibitors show little influence on cerebral blood flow, the present study suggests that bunazosin hydrochloride sustained release formulation may show a good influence on cerebral blood flow in mild to moderately hypertensive patients with chronic cerebral infarction. PMID- 10853684 TI - Direct and indirect effects of pulsatile shear stress on the smooth muscle cell. AB - BACKGROUND: Anastomotic intimal hyperplasia is still an unsolved problem after small caliber prosthetic bypass grafting. Oscillatory turbulent flow occurs at the end to side anastomosis, and produces various effects on smooth muscle cells (SMCs) and endothelial cells (ECs), which compose intimal hyperplasia. We examined the influences of pulsatile oscillating shear stress on smooth muscle cells mitogenic activity induced by sheared endothelial cells. METHODS: 1) Smooth muscle cells were cultured under three different pulsatile shear conditions (mean: 0, 6, and 60 dyne/cm2). 2) Endothelial cells were cultured under both static and sheared condition (mean: 60 dyne/cm2). Using the conditioned media from each well, SMCs were cultured under static and sheared conditions (60 dyne/cm2). Four groups of SMCs were devised by combining the two types of media and the two culture conditions. SMC colony spreading distances were measured as an index of combined migration and proliferation activity. An MTT assay and a cell counting assay were used to determine the proliferation activities of SMCs. RESULTS: 1) SMC spreading activity was suppressed by shear stress. SMC proliferative activity was stimulated by pulsatile turbulent shear stress. 2) SMC spreading activity was stimulated by mitogens derived from ECs under shear stress. However, this augmented SMC spreading activity was attenuated under sheared conditions. The mitogens derived from ECs under pulsatile shear stress had no effects on SMC proliferation activity. CONCLUSIONS: Pulsatile oscillating shear stress attenuates SMC migration activity induced by EC-denve mitogens and stimulates SMC proliferative activity. PMID- 10853685 TI - Risk factor assessment in the management of patients with suspected deep venous thrombosis. AB - BACKGROUND: To evaluate the prevalence of thrombosis risk factors in a group of patients undergoing venous duplex scanning (VDS) and to design a risk factor stratification model with the ability to improve the diagnostic yield of VDS. METHODS: Risk factor assessment and VDS were performed on 1,000 patients with clinically suspected lower extremity deep vein thrombosis (DVT) and patients were divided into two groups based upon the outcome of their scan: those with and those without confirmed DVT. Univariate and multivariate logistic regression analyses were performed in order to determine the significance of each risk factor in relation to having a confirmed DVT. RESULTS: There were 181 patients (18.1%) with confirmed DVT. A prior history of DVT/pulmonary embolism, malignancy, prior immobilization, and age over 70 were the most important risk factors associated with having a DVT confirmed on VDS. A novel risk factor stratification model was created utilizing the odds ratios of those factors found to be significant and the prevalence of DVT was found to be 92.4% in the high risk category, 11.5% in the moderate risk category, and 3.2% in the low risk category using this model. CONCLUSIONS: Venous duplex scanning is established as the screening test of choice when one suspects the diagnosis of DVT despite the significant cost of performing and interpreting the test. We suggest that a better clinical model utilizing risk factor assessment may be the key to increasing the yield rate and cost-effectiveness of VDS by excluding low-risk patients from undergoing unnecessary testing. PMID- 10853686 TI - Decreased binding sites of angiotensin II in rat LY-80 and AH109A tumour and human gastric cancer using quantitative in vitro autoradiography. AB - BACKGROUND: Under systemic hypertension induced by angiotensin II (AII) infusion, an attenuated vasoconstrictive response to the infusion in tumours was observed and a marked increase in tumour blood flow was observed in comparison with that in normal tissues. The results show a parallel circuit that connects the vascular bed of the pre-existing tissue to that of the tumour. The phenomenon was absent when hypertension was provoked by other vasoconstrictive agents such as norepinephrine or endothelin-1. However, the biological basis for this attenuated vasoconstrictive response to angiotensin II observed in tumours has not been fully elucidated. METHODS: We assessed this response to characterise the angiotensin II receptor density and affinity in normal and tumour tissues. AH109A and LY80 tumour cell lines were transplanted to the skin in nude rats. Four weeks later, the rats were sacrificed. 125I-[Sar1, Ile8] angiotensin II was used to map its receptors in rat tissues using in vitro computerised autoradiography. Operated human gastric cancer tissues from a 49-year-old and a 66-year-old male patients were also investigated. RESULTS: The numbers of angiotensin II receptors were markedly reduced in tumour tissues without a change of affinity. The numbers in AII-R in tumours were shown to be mainly AT1 by the marked reduction in radioligand binding achieved by losartan but not by PD123177. The same results were observed in human gastric cancer. CONCLUSIONS: These results suggest that the decrease in angiotensin II receptors in tumours may explain the haemodynamic effect of angiotensin II-induced hypertension on tumour blood flow. This condition for drug delivery to tumour tissue may play a major role in enhancing the therapeutic effects of chemotherapy. PMID- 10853687 TI - Wall lesions of abdominal aortic aneurysms threatening an impending rupture: prognostic evaluations. AB - BACKGROUND: Final events preceding aneurysm rupture are not completely known. The current study relates to incomplete aortic aneurysm wall lesions (i.e. malacia, dark staining and blebs or blisters) as possible sites of aneurysm rupture. METHODS: 162 abdominal aortic aneurysms, resected between 1988 and 1996, have been reviewed and 27 cases of aneurysms presenting wall thickness lesions are reported. The lesions were scheduled by operative reports and compared to ultrasound and CT scans. RESULTS: The authors classify aortic aneurysms into three phases, depending on the degree of wall degeneration viz 1. Flawless wall aneurysms. 2. (a-b-c) Aneurysms with intraparietal lesions. 3. Ruptured aneurysms. CONCLUSIONS: It is concluded that stage 2 aortic aneurysms must be urgently operated on. They carry a high surgical risk and, consequently, higher morbidity and mortality compared with stage 1 aneurysms. PMID- 10853688 TI - Plasma and tissue levels of collagen types I and III markers in patients with abdominal aortic aneurysms. AB - BACKGROUND: To study the levels of the aminoterminal propeptide of type III(PIIINP) and carboxyterminal propeptide of type I procollagen (PICP) in plasma and in the wall of abdominal aortic aneurysms in relation to their size and symptomatology. PIIINP serves as a marker of turnover and PICP as a marker of the synthesis of the corresponding collagens. METHODS: EXPERIMENTAL DESIGN: A prospective non-randomised study. SETTING: University Hospital, Plzen, Czech Republic. PATIENTS: Eighty-six patients who underwent resection of abdominal aortic aneurysms, average age 70.1 years (range 45 to 91 years), men to women ratio 5:1. The indication for resection was its symptomatology without relation to its diameter, and diameter over 5 cm in asymptomatic patients. Twenty patients (with similar age and gender distribution) scheduled for hernia repair or laparoscopic cholecystectomy were examined as a control group. MAIN OUTCOME MEASURES: The plasma and tissue PICP and PIIINP concentrations were evaluated using radioimmunoassay methods. The plasma samples were taken from the cubital vein without the use of a tourniquet. Full-thickness sections of the anterior abdominal aortic aneurysm wall at the site of the largest aneurysm diameter were taken at the time of operation. RESULTS: A significant difference between plasma PIIINP levels in patients with abdominal aortic aneurysms and the control group was observed (p<0.01). No correlation of PICP, PIIINP plasma levels with diameter and symptomatology of abdominal aortic aneurysms was found. The increase in PHIIINP tissue concentration was significant in patients with increasing diameter and positive symptomatology (p<0.01). No statistically significant correlation between plasma and tissue PICP and PIIINP concentrations was observed. CONCLUSIONS: The metabolism of type III collagen is increased in patients with abdominal aortic aneurysm, in contrast to type I collagen. The result is a degradation of collagen in the aneurysmal wall. The turnover of type III collagen increases with the enlargement of the aneurysm diameter and with the positive symptomatology. Degradation of type III collagen in the aneurysmal wall has therefore a fundamental significance for abdominal aortic aneurysm rupture. Because no correlation between plasma and tissue levels of PIIINP was found, the plasma levels of PIIINP cannot be used as the plasma markers of this process. PMID- 10853689 TI - Sutureless vascular reconstruction with titanium clips. AB - Attempts to find alternatives to sutured vascular reconstruction techniques has continued for decades and include various forms of rings, tubes, endoluminal stents as well as gluing or welding techniques of large vessel anastomoses. One recently introduced technique uses nonpenetrating titanium clips for everted vessel approximation and closure. Experimental work on their use in various types of large vessel repairs and reconstructions has shown that the clips are easily applied with a short learning curve, create good conditions for vessel wall healing without causing excessive inflammation or fibrosis, and are considerably faster to apply when compared to standard suture techniques. Although there are some clinical reports of defective clipped closures causing postoperative bleeding complications, they are rare and most probably related to technical errors in applying the clips. The main disadvantages of the clips include the limited experience of their applicability in atherosclerotic vessels, lack of long term follow-up and cost. Potentially, the clips could be useful in the repair of multiple vascular injuries, in vessel repair or ligation performed in confined spaces, and in vascular procedures requiring the shortest possible cross clamping time. Future applications could include endoscopic procedures as well as the use of a one-shot device which simultaneously applies up to a dozen clips to symmetrically everted and approximated vessel edges. PMID- 10853690 TI - Free-floating femoral vein thrombus in a patient with aspergillosis. AB - The management of a free-floating thrombus in the femoropopliteal or iliocaval veins is controversial. Such patients may have an increased risk of pulmonary embolism. The differential diagnosis of intraluminal venous malignancy or septic thrombosis must also be considered, especially in immunocompromised patients. This report reviews the management of a 56-year-old woman with bronchopulmonary aspergillosis who was found to have a free-floating thrombus in the femoral vein. Appropriate preoperative evaluation, emphasizing non-invasive studies and duplex exam, are discussed. In addition, the differential diagnosis, surgical options and perioperative care are considered. This patients represents a complex case of venous thrombosis in an immunocompromised patient and, therefore, the optimal care to minimize complications, such as pulmonary embolism, and prevent recurrence or post-thrombotic changes, is necessary. PMID- 10853691 TI - Recurrent peripheral arterial embolism from pulmonary cancer. Case report and review of the literature. AB - Peripheral arterial embolism arising from a malignant tumour is an infrequent manifestation of neoplastic disease and also a rare cause of acute arterial occlusion. A case of recurrent arterial embolism of the lower extremities due to a primary lung cancer is reported and the literature on this topic is reviewed. PMID- 10853692 TI - E. Kondoleon: the man behind the procedure. AB - Historical events and their in-depth analysis have much to teach us about medicine today. Moreover, the lives of those who have made their mark in medical history can be a source of inspiration to future generations of medical scientists. A case in point is the career of E. Kondoleon, originator of the surgical procedure that bears his name. METHODS: Retrieval in the archives of the "Areteion" and "Hippocration" hospitals, where E. Kondoleon had worked and passed away, together with information obtained from the Chair of History of Medicine of the University of Athens, as well as information taken from the Hellenic Surgical Society relative to his life and the motives which led him to the Kondoleon surgical procedure namely: Wide excision of the fascia and concomitant partial excision of the hypertrophic tissue in the treatment of lymphoedema. RESULTS: The Kondoleon operative procedure was not a simple applicable idea of a surgical technique but the result of a long-term systematical study of the anatomy and pathophysiology of the lymphovascular system as well as the continuation of their experimental application. Thus in the beginning of our century, Kondoleon took the lead of carrying out his own technique on the human which for more than 50 years was the object of discussion and application in the treatment of lymphoedema. CONCLUSIONS: Although "the microworld" of the electronic microscope and the conventional microscope of surgery have illuminated many sections of pathophysiology and has led to newer evolutionary tactics (vascular anastomoses, transplantation of lymph-vessels) in the management of lymphoedema, the Kondoleon operative procedure has given the necessary stimuli for these developments. As with other prominent, celebrated men in the history of investigation, Kondoleon had a dramatic end. PMID- 10853693 TI - Long-term effects of different physical activity levels on coronary heart disease risk factors in middle-aged men. AB - In order to define the amount of physical activity appropriate in primary prevention of coronary heart disease (CHD), we have compared the effects during 5 years of physical activity in four groups of middle-aged men with different but stable approximate metabolic costs of leisure time sports activities (AMCSA): sedentary (n = 40; 0 kcal per week), low activity (n = 31; 1-999 kcal per week), moderate activity (n = 56; 1,000-1,999 kcal per week), and high activity (n = 71; > or = 2,000 kcal per week). Time related increase of body mass and BMI was more pronounced in lower activity groups. Changes in HDL cholesterol were more favourable in the high activity group as compared to sedentary and low activity groups. The increase of diastolic blood pressure (DBP) in the sedentary group was statistically significantly different from the decrease of DBP observed in both moderate and high activity groups. We conclude that favourable long-term stabilization of most coronary risk factors is achievable with physical activity energy expenditure above 1,000 kcal per week. Physical activity-related energy expenditure > or = 2,000 kcal per week is associated with some additional benefits, especially with a favourable modification of HDL cholesterol level. PMID- 10853694 TI - Comparison of laboratory and "on-court" endurance testing in tennis. AB - Treadmill testing (TT) commonly used in endurance testing is often not sport specific. Therefore a field test (FT) for tennis players was developed. The purpose was 1) to compare metabolic and cardiorespiratory response between TT and FT and 2) to assess tennis stroke ratings during FT. In both tests ventilatory variables (VO2, VE, VT, Bf, VE x VO2(-1)), heart rate (HR), and lactate (LA) were measured. For both tests an "individual anaerobic threshold" (IAT) was calculated. The comparison of TT and FT yielded significant differences in cardiorespiratory and metabolic response. LA and VE were significantly higher in TT compared to FT at VO2 of 35, 40, and 45 ml x kg(-1) x min(-1). There were statistical differences between IAT resulting from both tests (TT vs. FT): HR (165+/-16, 175+/-11, p<0.001), VO2 (44.4+/-4.3, 47.8+/-4.8, p<0.05), LA (3.1+/ 0.5, 2.5+/-0.4, p < 0.001), VE (97.0+/-15.6, 89.1+/-14.9, p < 0.05), VT (2.66+/ 0.34, 2.34+/-0.47, p<0.05), VE/VO2 (27.9+/-3.9, 23.9+/-2.9, p<0.01). High correlation was found between stroke ratings and the national ranking of the players. We concluded that 1) metabolic, ventilatory, and cardiorespiratory demands of TT vs. FT were (semi)sport-specific and significantly different and 2) that the stroke rating in our study was a good predictor for tournament performance (r = 0.94). This type of stroke rating can be implemented in a FT. PMID- 10853695 TI - Performance level and cardiopulmonary responses during a cycle-run trial. AB - To determine the effect of triathlete performance level on the cardiorespiratory responses elicited by the cycle-run succession, eight regionally and nationally ranked (Competitive) and five internationally-ranked (Elite) male triathletes underwent four successive laboratory trials: 1) an incremental treadmill test, 2) an incremental cycle test, 3) 30 min of cycling followed by 20 min of running (C R), and 4) a 20-min control run (R) at the same speed as the run in C-R. Before and 10 min after the third and fourth trials the triathletes underwent lung function testing: spirometry and diffusing capacity testing for carbon monoxide (DL(CO)). During the C-R trial blood samples were drawn to measure venous lactate concentration. During all trials ventilatory data were collected every minute using an automated breath-by-breath system. The results showed that 1) the oxygen uptake (VO2) of post-cycling running versus running alone was similar for both groups; 2) the ventilatory responses (VE, VE/VO2, VE/VC02 and f) of C-R running versus R were significantly higher (P < 0.005) for the Competitive group; and 3) a significant decrease (P< 0.05) in DL(CO) was also noted after the C-R trial in the Competitive group but not in the Elite group. We concluded that 1) the ventilatory responses during a run subsequent to cycling may be related to the triathlete performance level, and 2) the C-R trial induced specific alterations in pulmonary function that may be associated with respiratory muscle alteration and exercise-induced hypoxemia in the Competitive triathletes. PMID- 10853696 TI - Interactions between homocyst(e)ine and nitric oxide during acute submaximal exercise in adult males. AB - Experimental studies investigating the effects of exercise on plasma total homocyst(e)ine (H[e]) levels in humans are almost non-existent. H(e) has been demonstrated to represent an independent risk factor for cardiovascular disease. The exact mechanism through which H(e) exerts its effects on the arteries is unknown but it is thought to involve nitric oxide (NO). The present study was designed to assess the effects of acute submaximal exercise on H(e) while levels of NO inhalation were manipulated using an air-filter mask. The study was completed by seven male volunteers, aged 21.6+/-1.3 yr (X+/-SD), VO2max: 48.6+/ 7.6 mL x kg(-1) x min(-1). During two separate occasions the subjects performed a 1-hour bout of submaximal exercise on a stationary cycle ergometer at 60% of their VO2max. The two trials were completed in random order (with and without mask). Data were collected before (PRE) and after (POST) the acute exercise bouts. Plasma H(e) was directly measured by HPLC and NO by quantifying the enzymatic oxidation to nitrite (NO2-) & nitrate (NO3-). Mean H(e) concentrations were 10.89+/-2.05 nmol/mL (PRE) & 11.21+/-1.81 nmol/mL (POST) and were not significantly altered by submaximal exercise. When wearing a mask, the correlation of the PRE/POST H(e) differences with the PRE/ POST differences in NO3- were 0.77 (P=0.07). No correlation was found between either H(e) and NO2- or between NO2- and NO3-. However, a significant correlation (r= - 0.86, P= 0.03) was also observed between H(e) and NO2- but only for the post-exercise values when wearing a mask. The results suggest that: (1) plasma H(e) levels are not affected by acute submaximal exercise; (2) there is insufficient evidence to support the view that plasma H(e) levels are being mediated by NO during either rest or exercise. PMID- 10853697 TI - Oxygen cost for cycling as related to leg mass in males and females, aged 20 to 80. AB - In order to evaluate the determinants of the metabolic cost for cycle ergometry, we analyzed the relationship between VO2 and leg mass (LM) and total body mass (TBM) in 71 randomly-selected sedentary subjects (34 men), aged 20 to 80. Participants performed constant work rate (WR) tests at 0, 25, and 50 W (at 60 rpm) for 6 minutes in a randomized sequence: gross VO2, gross efficiency, and work efficiency were related to TBM and LM as assessed by dual energy x-ray absorptiometry. We found that gross VO2 and gross efficiency were more strongly related to LM than TBM but work efficiency values were independent of both (P>0.05). Significantly higher values of VO2TBM were found in subjects with large LM/TBM ratios and vice-versa; VO2/LM, however, did not change with anthropometric characteristics. Gross VO2 (mL/min) was predicted by a LM-based equation (10.6 [WR, W] + 16.8 [LM, kg] +75) with a mean error below 5%: this equation predicted the cost more accurately than previous TBM-based formulations (P<0.01). We conclude that leg mass actually provides the preferred frame of reference for predicting the oxygen cost for cycle ergometry at 60 rpm in sedentary subjects, independent of age or gender. PMID- 10853698 TI - Exercise in the heat: effects of an adenosine antagonist. AB - The purpose of this experiment was to examine the effects of an adenosine antagonist on cardiovascular, thermoregulatory, and exercise performance in the heat. Two doses (1 mg/kg and 10 mg/kg) of a selective adenosine A1 antagonist (1,3-di-n-propyl-8-[4-hydroxyphenyll]xanthine) (DPHPX) were tested in a rat model of exercise exhaustion, treadmill 11 m/min, 6 degrees incline, in the heat 30 degrees C. Pretreatment with the experimental adenosine antagonist caused a slight improvement p > 0.05 in run time (41+/-4 vs. 44+/-3 mm) at a low dose but reduced performance (41+/-4 vs. 29+/-3 mm) at a high dose despite elevated plasma lactate (6.41+/-0.82 vs. 9.91+/-1.0 and 12.42+/-1.1 micromole/L) levels in both dosage groups. At the low dose the antagonist provided a clear benefit in thermoregulation as evidenced by reduced heating rates (0.079+/-0.005 vs. 0.050+/ 0.009 degrees C/min). Heart rate and blood pressure tended to be preserved in the low dose group also. Blood gases remained closer to normal with either dosage of drug with arterial PO2 being remarkably preserved after exercise whereas venous PO2 was not different suggesting increased oxygen delivery and extraction. The results of this investigation indicate that antagonizing the effects of adenosine at a low dose with this agent did improve cardiovascular and thermoregulatory responses but did not provide a substantial overall benefit in exercise performance in the heat. PMID- 10853699 TI - The effects of a prolonged running exercise on strength characteristics. AB - The aim of this study was to examine concentric, isometric, and eccentric strength reductions in the quadriceps muscle following a prolonged running exercise. Before and after a 2 h run (28.4+/-1.4 km) peak torque (PT) of the knee extensors at angular velocities of -120, -90, -60, 0, 60, 120, 180, 240 degrees x s(-1) using an isokinetic dynamometer, electromyographic (EMG) activity of the vastus lateralis (VL) and vastus medialis (VM) muscles and height of a counter movement jump were recorded in twelve well-trained triathletes. Counter movement jump performances decreased by 10% and PT values were all significantly lower (p < 0.01) at each angular velocity following the run. The torque loss was significantly (p < 0.01) greater under eccentric contractions (from 18 to 21%) than under concentric ones (from 11 to 14%). EMG activity (RMS) was lower in both VL and VM muscles after the 2 h run but no difference existed in RMS losses between concentric and eccentric contractions. The present results demonstrate that 1) a prolonged running exercise more greatly affects eccentric force production in the quadriceps muscle, and 2) this specificity seems to be due to an impairment of the muscular contractile mechanism rather than a modification to the neural input. PMID- 10853700 TI - Re-examination of training effects by electrostimulation in the human elbow musculoskeletal system. AB - This study examines the effects of a 7 weeks sub-maximal training period of electrostimulation on the maximal isometric, concentric, eccentric voluntary torque and muscle contractile properties of the elbow flexor muscles of nine subjects. The daily program consisted of five series of six 6-s isometric actions (60 to 70% of maximal isometric voluntary action) at an elbow angle of 90 . After training the maximal voluntary isometric flexion torque increased significantly whereas the maximal voluntary isometric extension torque decreased significantly. Increases in isometric flexion torque were linked to an increase of the myoelectrical activity of the biceps brachii muscle. Under dynamic conditions flexion torque was significantly increased throughout the whole spectrum of angular velocities. These changes were accompanied by an increase in the myoelectrical activity of the agonist muscle under eccentric conditions and at two fast concentric angular velocities, without modifications of the myoelectrical activity of the antagonist muscle. The analysis of the electrical and mechanical twitches indicated that modifications of the muscle membrane electrical activity were also present at the muscle level. These results indicate that torque gains were attributed to neural adaptations and/or to a modification of the relative part of agonist and antagonist muscles in elbow flexion torque production. PMID- 10853701 TI - Influence of an acute exercise on neutrophil and platelet adhesion, nitric oxide plasma metabolites in inactive and active subjects. AB - In this work we studied the influence of an acute exercise either on nitrite/nitrate plasma levels or on neutrophil and platelet adhesion in inactive and active subjects. Twelve healthy subjects (6 inactives and 6 actives) exercised on a racing cycle ergometer performing stepwise increases in intensity until reaching, within 5 min, a heart rate of 150 beats x min(-1) which represents an oxygen consumption of about 75 % of the individual maximum rate of oxygen uptake. From peripheral venous blood samples (drawn from all subjects before, immediately after the end of exercise, and 1 hour later) neutrophils and platelets were isolated to test plate adhesion, and nitrite/nitrate concentrations were measured in the plasma. Immediately after the acute exercise, in active subjects we observed a significant decrease in the percentage of neutrophil adhesion (7.96+/-2.38 vs. 14.10+/-3.14), associated with an increase in nitrite/nitrate plasma levels (81.38+/-10.76 vs. 41.08+/-8.13 micromol x l( 1)), restored by a 40 min pre-incubation with NG-nitro-L-arginine methyl ester (L NAME). In unstimulated platelets we observed a significant lower percentage of platelet adhesion in active subjects compared to inactives after exercise. With thrombin or adenosine 5'-diphosphate as agonists platelet adhesion did not result significantly different in active subjects compared to inactives. In conclusion, our data show that physical exercise can induce changes in some cell activities, even if transient, and favour the generation of nitric oxide. The lower adhesion of neutrophils and platelets induced by regular exercise could be an important goal in the prevention of vascular and inflammatory diseases. PMID- 10853702 TI - Immune modulation following aerobic exercise in children with cystic fibrosis. AB - Previous studies have demonstrated altered immune response following exercise in healthy adults and children. As data are lacking in children with cystic fibrosis, we evaluated the immune response following acute exercise and hypothesized that acute increases in cellular changes would be seen but would be blunted in subjects with CF. Leukocytes, lymphocytes, and their subsets as well as natural killer cell number and activity were determined before, immediately after, and one hour post exhaustive exercise in 15 children with cystic fibrosis (8-21 yrs, FEV1 69.5+/-18.0%, colonized with P aeruginosa) and 15 healthy controls (8-18 yrs, FEV1 107.5+/-10.7%). At baseline the cystic fibrosis group had greater leukocytes (9.25+/-2.83 vs. 5.17+/-0.96 x 10(9) cells/liter). Immediately post exercise, the cystic fibrosis group demonstrated increases in cell counts for leukocytes (32.4%), lymphocytes (61.8%), granulocytes (36.4%), monocytes (76.2%), and natural killer cells (315%). Similar percentage increases were seen in cell counts for the controls (leukocytes: 39.5%, lymphocytes: 78.5%, granulocytes: 32.0%, monocytes: 75.9%, and NK cells: 442%). Natural killer cell activity also increased by 57.9% in the group with cystic fibrosis and by 43.6% in the healthy controls. Except for elevated leukocyte and granulocyte counts, values returned to baseline at one hour post-exercise. In conclusion, the cellular immune response to acute exercise in children with mild to moderate cystic fibrosis appears normal. PMID- 10853703 TI - Immune status and respiratory illness for elite swimmers during a 12-week training cycle. AB - The impact of a 12-week training program by elite swimmers on systemic and mucosal immunity was studied prospectively to examine the relationship between changes in immune parameters and the incidence of respiratory illness. Saliva was collected before and after selected training sessions at 2 weekly intervals. There were significant decreases in salivary IgA (p=0.05) and salivary IgM (p < 0.0001) concentrations after individual training sessions, but no significant changes in salivary IgG or albumin concentrations. Over the 12-week training program there were small but statistically significant increases in pre-exercise concentrations of salivary IgA (p<0.001), IgM (p=0.015) and IgG (p=0.003) and post-exercise salivary IgA (p <0.001). There were no significant trends over the 12 weeks for any class of serum immunoglobulins but a significant fall in NK-cell numbers (p<0.001). There were no associations between serum or salivary immunoglobulin levels or NK-cell numbers and upper respiratory tract illness (URTI) during the 12-week program. The data indicated that despite changes in some immune parameters during this final training program prior to competition there were no associations detected with URTI for this cohort of elite swimmers. PMID- 10853704 TI - Acute ulnar nerve compression syndrome in a powerlifter with triceps tendon rupture--a case report. AB - We report on the case of a bodybuilder and powerlifter who suffered from triceps tendon rupture complicated by acute ulnar nerve compression syndrome. The diagnosis was made clinically, radiologically, and sonographically. Ultrasound was helpful to demonstrate a large hematoma at the site of the injury. Early surgical intervention confirmed the presence of the hematoma compressing the ulnar nerve and led to a complete restoration of ulnar nerve and triceps muscle function. Few reports on distal triceps rupture have been published but its complication by acute ulnar nerve compression has not been reported on yet despite the close anatomical relationship of both structures. PMID- 10853705 TI - Morphology of FRTL-5 cell colonies in a semi-solid medium. AB - For the first time, 3, 7 and 10 days growth of normal thyroid follicular FRTL-5 cell colonies in a semi-solid medium of 0.6% methocel over 1% agar base was morphometrically analyzed. It was found that the areas of FRTL-5 colonies as well as their perimeters and maximum diameters increased, while according to their form factors the FRTL-5 colonies were regular in shape. After 10 days in a semi solid medium, 83% of the FRTL-5 colonies had maximum diameters greater than 50 microm. It is obvious that after long culturing of FRTL-5 cells under the influence of the pituitary thyroid-stimulating hormone (TSH) these cells are not uniform anymore and that they grow in a semi-solid medium. PMID- 10853706 TI - Role of insulin-like growth factor-I in primary osteoporosis: a correlative study. AB - Osteoporosis is characterized by impairment of bone mass and deterioration of bone microscopic structure, resulting in increased bone fragility and susceptibility to fracture. Recent reports have indicated that reduced plasma levels of IGF-I are associated with osteoporosis in both males and females. Moreover, there is accumulating clinical evidence that treatment with GH or IGF-I has beneficial effects on bone mass and bone remodeling in men with idiopathic osteoporosis, in the elderly and in hypopituitary patients. As correlative studies on IGF-I, IGF-BP3 and bone mass in the elderly are lacking, we studied the relationships between serum IGF-I, IGF-BP3, bone mineral density (BMD), body mass index (BMI), calciotropic hormones and age in 102 premenopausal and postmenopausal women. Our study indicates that the reduction of the anabolic processes mediated by IGF-I may account for the slow and progressive loss of bone mass that take place after the age of 40-50 years. In addition, nutritional caloric or proteic deficit may add to the effects of GH, age and other factors in decreasing IGF-I synthesis and therefore further contribute to the development of primary osteoporosis. PMID- 10853707 TI - Erythrocyte oxidant/antioxidant status of diabetic patients. AB - Present study aims to establish erythrocyte oxidant/antioxidant status in diabetic patients with and without atherosclerotic complications. Fasting blood samples were obtained from 23 diabetic and 12 control subjects. Thirteen patients had no disease other than diabetes mellitus and 10 patients had also atherosclerosis in addition to diabetes mellitus. Erythrocyte antioxidant potential (AOP) and thiobarbituric acid reagent substances (TBARS) levels were measured in these patients and results were compared with those of controls, who were chosen among healthy subjects. Results suggest that although there is an oxidant stress in the erythrocytes of diabetics, this is not due to reduced erythrocyte antioxidant defence potential but, rather, increased free radical production possibly due to hyperglycemia. PMID- 10853708 TI - Activin A stimulates insulin secretion in cultured human pancreatic islets. AB - Activin A is a dimeric glycoprotein showing a high sequence homology with transforming growth factor-beta (TGF-beta) and playing autocrine/paracrine actions in reproductive tissues. However, since the synthesis of activin is ubiquitous it may have a role in regulating cell growth and differentiation in several tissues. Previous studies showed that activin A is expressed by insulin positive B cells of human pancreatic islets, and women with gestational diabetes have higher serum activin A levels than healthy pregnant women at the same gestational age. The present study aimed to evaluate the effect of activin A on insulin secretion from cultured human pancreatic islets. With this purpose human pancreatic islets were incubated with varying concentrations of activin A (0.1 to 10.0 nM). In absence of glucose, activin A did not modify insulin secretion at the different concentrations used. In absence of activin A, 8.3 mM and 16.7 mM glucose significantly increased insulin secretion, with a dose-dependent pattern. In presence of a non stimulatory concentration of glucose (3.3 mM), activin A significantly increased insulin secretion starting from low concentration (0.1 nM). Furthermore, the addition of activin A to 8.3 mM and 16.7 mM glucose induced an additional effect of the dose-dependent glucose-mediated insulin secretion (p<0.001). The present data could support a role for activin A in human endocrine pancreas in modulating insulin response to different glucose concentrations. PMID- 10853709 TI - Low dose (1 microg) adrenocorticotropin stimulation test in the evaluation of hypothalamo-pituitary-adrenal axis in patients with active pulmonary tuberculosis. AB - Adrenocortical function in patients with active pulmonary tuberculosis is a debate of matter. Previous studies related to adrenocortical function in patients with active pulmonary tuberculosis demonstrated a high rate of suboptimal cortisol response to standard dose ACTH (250 microg) stimulation test. The aim of this study was to assess the hypothalamo-pituitary-adrenal (HPA) axis in low dose (1 microg) and standard dose ACTH (250 microg) stimulation tests in the patients with active pulmonary tuberculosis. Twenty-seven patients and 21 healthy subjects were included in the study. Cortisol levels were measured before, 30 and 60 min after ACTH (1 microg or 250 microg iv) injection. Cortisol responses to 1 microg ACTH at 30 and 60 min were significantly higher in the patient group than in the control group (p<0.05). Peak cortisol levels were significantly higher in the patient group than in the control group after both 1 microg and 250 microg ACTH administration (p<0.05). Cortisol responses to 250 microg ACTH at 30 and (at 30 and 60) 60 min were significantly higher in the patient group than in the control group (p<0.05). Peak cortisol levels obtained after 250 microg ACTH and after 1 microg ACTH were similar in the patient group (p>0.05). This study shows that 1 microg ACTH iv gives an equivalent peak cortisol value to 250 microg ACTH in patients with activated HPA axis. The cortisol levels obtained at 08:00, 11:00, 17:00 and 24:00 h were significantly higher in the patients than in the controls. This study clearly shows that HPA axis is activated in active pulmonary tuberculosis rather than underactivated. PMID- 10853710 TI - Endothelin-1 and endothelin-3 stimulate insulin release by isolated rat pancreatic islets. AB - Endothelins (ETs) are potent vasoconstrictive peptides released from the endothelium and other tissues, which act on target cells by receptorial calcium mediated mechanisms. ET-1 levels are increased in diabetes, and observations suggest the involvement of ETs in the pathogenesis of diabetic angiopathy. However, it is not possible to exclude that ETs might also influence insulin secretion or function. In vivo infusion of ET-1 in rats induces hypoglycaemia and hyperinsulinemia and in vitro incubation with ET-1 stimulates insulin release by mouse islets. Therefore, ETs might be involved in a circulus vitiosus, resulting in hyperinsulinemia and diabetic angiopathy. The purpose of our study was to verify the effect of ET-1 on rat islets, in both the presence and absence of physiological glucose concentration. Moreover, we tested the effect of another isoform of endothelins, ET-3, and verified the involvement of extracellular calcium in such events. Islets were incubated with increasing ET-1 or ET-3, with or without glucose 5.6 mM. Other samples were prepared using calcium-free medium. Incubation in medium containing ET-1 and ET-3, in the presence of glucose and calcium, induced an increase in insulin release. When ET-1 and ET-3 were incubated without glucose and calcium, insulin release was not modified. Our studies demonstrate that: 1) ET-3, like ET-1, stimulates insulin release by rat isolated islets; 2) direct insulin stimulating effect on islets of both ET-1 and ET-3 is evident with physiological glucose concentrations and is calcium mediated. These results support the hypothesis of ET involvement in the regulation of insulin secretion. PMID- 10853711 TI - Testosterone modulates serum leptin concentrations in a male patient with hypothalamic hypogonadism. AB - Serial measurements of body mass index (BMI), serum concentrations of testosterone (T), estradiol (E) and leptin (L) were performed before and after gonadotropin (Gn) therapy in an 18-year-old male subject (BMI 25.4 kg/m2) with idiopathic hypothalamic hypogonadism (IHH). We also measured the BMI and serum concentrations of L in 99 age-matched healthy subjects. Serum L correlated significantly with BMI in control subjects (r=0.84, p<0.0001). Baseline serum concentrations of L in our case were markedly high and both T and E were very low, but Gn therapy resulted in a gradual decrease in L and improvement in T and E, finally reaching the control levels of BMI-matched subjects. Our results demonstrate that T is a powerful negative modulator of serum L independent of BMI in conditions associated with low T levels, such as IHH. PMID- 10853712 TI - Incidentally discovered papillary carcinoma of the thyroid: value of ultrasonographic follow-up. A case-report. AB - Incidentalomas of the thyroid are common small nodules found occasionally during imaging procedures. Their pathological nature is generally benign, but about 4% may harbour malignant tissue. Most current studies only suggest clinical follow up, but there are no data about the natural history of malignant incidentalomas. The authors describe a patient with multiple incidentaloma of the thyroid submitted to fine-needle aspiration biopsy because his larger nodule grew 50% in 3 months at ultrasonographic follow-up. The cytological examination suggested thyroid malignancy in this nodule and surgical pathology showed multicentric papillary carcinoma. This case suggests that the larger diameter of a malignant incidentaloma may change rapidly. If more time had been spent before repeating the ultrasonography, the volume could have changed even more and the prognosis could have been changed as indicated by most prognostic score indexes. PMID- 10853713 TI - Cutaneous spreading of parathyroid carcinoma after fine needle aspiration cytology. AB - BACKGROUND: Ultrasound-guided fine needle aspiration cytology (FNAC) of suspect parathyroid adenomas is sometimes used for the diagnosis of primary hyperparathyroidism (PHPT). FNAC complications are rare or mild. We describe the first case in literature of cutaneous spread of parathyroid carcinoma after FNAC. CASE: A woman underwent a neck ultrasound which revealed a solid hypoechogenic nodule of 1.5 cm at the level of the inferior pole of the right thyroid. In the same time a FNAC of the nodule was performed. Cytology showed no atypical cells. Successively PHPT was diagnosed and a few weeks later the patient had a subcutaneous lump in the same area of FNAC. The patient underwent surgery and histology of the specimen showed a differentiated parathyroid carcinoma. The postoperative course was regular and calcium and parathormone resulted normal. CONCLUSION: The use of FNAC should be carefully assessed in the presence of suspect parathyroid carcinoma, because this could cause a possible diffusion of a parathyroid carcinoma along the needle tract. PMID- 10853714 TI - Lack of reduction in body fat after treatment with insulin-like growth factor-I in two children with growth hormone gene deletions. AB - Two patients with growth hormone (GH) gene deletions were treated with recombinant insulin-like growth factor-I (IGF-I) (80-240 (microg/kg/day) and the effects on bone mass and body composition were compared to administration of GH (0.075 U/kg/day) to 8 patients with idiopathic GH deficiency. Bone mass and body composition were measured by dual photon X-ray absorptiometry (DEXA ) before and 3 and 6 months after treatment with GH or IGF-I. Similar increases in growth velocities were observed after GH and IGF-I treatment. Treatment with GH resulted in prompt and significant reduction in body fat percentage (basal, 3 and 6 months: 22+/-10, 17+/-9, and 16+/-9%) whereas body fat percentage remained unchanged after IGF-I therapy (basal, 3 and 6 months: 49, 52 and 48% in patient 1 and 45, 42 and 43% in patient 2, respectively). Fat percentage remained elevated after 18 months of IGF-I treatment in patients 1 (51%) and 2 (44%), respectively. Lean mass and bone mineral content increased with GH and IGF-I therapies. We conclude that reduction of body fat measured by DEXA, observed after administration of GH but not after IGF-I treatment in these children with GH deficiency, suggests that the GH effect on body fat mass is not mediated by circulating IGF-I. PMID- 10853715 TI - Development of tertiary hyperparathyroidism after phosphate supplementation in oncogenic osteomalacia. AB - Oncogenic osteomalacia is a rare paraneoplastic syndrome. It is characterized by bone pain, muscle weakness, gait disturbance, fractures and skeletal deformities. Hypophosphatemia, diminished renal phosphate reabsorption, decreased 1,25 dihydroxy Vitamin D and elevated alkaline phosphatase are the biochemical hallmarks of this disorder. Most tumors are of mesenchymal origin. We report the case of a 39-year-old woman with oncogenic osteomalacia caused by osteosarcoma of the right scapula which was unrecognized for several years. She subsequently developed tertiary hyperparathyroidism after treatment with oral phosphate and Vitamin D. This case illustrates that oncogenic osteomalacia may persist for many years before the tumor is discovered. This is because the tumors are frequently very small and are in obscure locations. The uniqueness of this case is the coexistence of hyperparathyroidism and oncogenic osteomalacia. Five other cases have been reported up to date. All patients had received phosphate supplement, ranging from 10 to 14 years prior to their diagnosis. Interestingly, our patient was on the treatment for only 2 years. The proposed mechanism is that exogenous phosphate stimulates parathyroid activity through sequestration of calcium. PMID- 10853716 TI - "The old inhabitant of the Procida island." Lorenzo Mosca. Napels, Italy (?? 1789). PMID- 10853717 TI - Ethical issues in rehabilitation medicine. AB - It is only relatively recently that we have begun to examine ethical issues as they relate specifically to the speciality of Physical Medicine and Rehabilitation. Prior to this, most ethicists were more concerned with acute care medical issues involving life and death decisions. However, with the ageing population and the emphasis society now places on returning patients to the maximum possible level of function, greater consideration is being given to ethical dilemmas that are relevant to rehabilitation medicine. This paper examines the major ethical principles of autonomy, beneficence and justice. The issues of resource allocation and patient selection, the ethics of team care and ethical issues in goal setting, as they relate specifically to rehabilitation medicine, are examined in some detail. PMID- 10853718 TI - Asymmetric vestibular function in the elderly might be a significant contributor to hip fractures. AB - The aim of this study was to assess postural control, vestibular symmetry and health status in otherwise healthy hip fracture subjects and compare these factors with controls. The fracture subjects were recruited from 113 consecutive patients operated 12-33 months earlier. Nineteen of those were otherwise healthy and fulfilled the inclusion criteria. They were assessed and compared with 28 age and sex-matched controls. Nystagmus after head shake was checked for by video nystagmoscopy (charged couple device cameras). Vibration sensation was tested with a tuning fork, medical history and posturography of vibration-induced sway were studied. The subjects had a significantly higher frequency of head shake nystagmus (p = 0.03), indicating a vestibular asymmetry and a history of previous fractures (p = 0.002). Nine out of 12 subjects had fallen and sustained the hip fracture towards the slow phase of the nystagmus, which is expected in a vestibular related fall. Losing balance during testing was more frequent among the subjects than among the controls (p = 0.002). The subjects with head shake nystagmus swayed more than those without, especially in the sagittal plane during neck vibration with eyes closed (p < 0.001). Vibration perception was significantly poorer in the operated legs than in the healthy legs (p = 0.021) and in the legs of the controls (p = 0.001). The findings suggest that vestibular asymmetries may contribute to falls and fractures in elderly people. As such asymmetries can be compensated to a certain degree by specific training programs, these might be advisable for elderly people, especially those with a history of falls or fractures or where a vestibular asymmetry is suspected. PMID- 10853719 TI - Reliability of torque measurements during passive isokinetic knee movements in healthy subjects. AB - In the literature, few data are available about the reliability of torque measured during passive isokinetic knee movements. This study investigated the consistency of torque measurements during passive knee movements at 60, 180 and 300 degrees/second in 30 healthy subjects. Intraclass correlation values ranged between 0.78 and 0.92 when the results of two consecutive tests were compared. When retests were performed after repositioning the subjects, intraclass correlation values ranged between 0.43 and 0.87. These findings indicate the necessity for meticulous standardization of the test situation. Series of 10 consecutive movements, specifically repetitions of knee flexion at 180 and 300 degrees/second, indicated that torque measurements during the first two movements were less stable than those following. A concurrent change in electromyographic activity in the rectus femoris muscle suggested that these torque variations resulted from habituation of the stretch reflex. PMID- 10853720 TI - Effect of immobilization on ankle dorsiflexion strength. AB - This study was performed in order to determine the loss of strength of the dorsiflexors in healthy persons after immobilization of the ankle, and the ability of these muscles to regain strength. First, isometric ankle dorsiflexion strength was measured in 33 healthy male and 39 female subjects in age categories 20-40 and 40-80 years, in order to obtain reference data and to determine the reproducibility of the measurement protocol. Gender, age and ankle position had a significant influence on the ankle dorsiflexion torque. Secondly, torque was measured in 15 patients after 4-6 weeks' immobilization of the ankle due to a fracture. A 28% decrease in dorsiflexion torque was seen. Strength reduction in neutral position and in 30 degrees plantar flexion was not significantly different. Without specific therapy restoration of torque was almost complete 6 weeks after cast removal. PMID- 10853721 TI - Functional electrical stimulation-assisted walking for persons with incomplete spinal injuries: changes in the kinematics and physiological cost of overground walking. AB - This study was conducted to investigate the change in the kinematics and physiological cost of walking that occurs during training with functional electrical stimulation (FES)-assisted walking in persons with incomplete injuries. The main effect of FES-assisted walking was to change hip excursion and ankle dorsiflexion during swing and at foot contact, whereas training with FES assisted walking changed the spatio-temporal parameters of walking (walking speed, cycle length and frequency as well as time in stance). The use of FES assisted walking does not change the walking speed achieved during a 5-minute trial nor the physiological cost of walking but when combined with walking training, eight of the nine participants improved either their physiological cost index or their walking speed. It is concluded that FES-assisted walking changes the joint angular kinematic pattern of walking, but training is necessary to integrate these changes into functional gains. PMID- 10853722 TI - A randomized controlled trial of rehabilitation at home after stroke in Southwest Stockholm: outcome at six months. AB - A 6-month follow-up of a single-blind, randomized, controlled trial in Southwest Stockholm was performed in order to evaluate the effect of early supported discharge and continued rehabilitation at home after stroke. Eighty-three stroke patients with moderate neurological impairments, continent, independent in feeding, and mental function within normal limits one week after onset were included in the study. The patients were allocated 1:1 to early supported discharge and continued rehabilitation at home by a specialized team, versus routine rehabilitation. Patient outcomes measured were motor capacity, dysphasia, activities of daily living, social activities, perceived dysfunction, mortality and reported falls. Data on length of stay in hospital; initial and recurrent during 6 months were compared. The 6-month follow-up of 78 patients showed no statistically significant differences in patient outcome. The results of multivariate logistic regression analysis suggest a positive effect of home rehabilitation on activities of daily living. At 3-6 months the frequency of significant improvements was higher in the intervention group. Death or dependency in activities of daily living was 24% in the intervention group compared with 44% in the control group. The mean initial hospitalization was 29 days in routine rehabilitation group versus 14 days in the home rehabilitation group. We conclude that for moderately disabled stroke patients with mental function within normal limits, early supported discharge and continued rehabilitation at home had no less a beneficial effect on patient outcome than routine rehabilitation, reduced initial hospitalization significantly and had no adverse effects on mortality and number of falls. PMID- 10853723 TI - Adaptation of the modified Barthel Index for use in physical medicine and rehabilitation in Turkey. AB - The aim of this study was to adapt the modified Barthel Index for Turkey and to determine its reliability and validity. After the translation procedure, 50 stroke patients and 50 spinal cord injury patients, undergoing inpatient rehabilitation were assessed by the newly adapted index at admission and discharge. Reliability was tested using internal consistency, inter-rater reliability and the intra-class correlation coefficient. Construct validity was assessed by association with impairments (Brunnstrom motor stages in stroke, American Spinal Injury Association motor/sensory scores and impairment scale in spinal cord injury) and by Rasch analysis. Internal consistency was good at 0.93 for stroke, and 0.88 for spinal cord injury. The level of agreement between two raters was sufficient with Kappa levels of above 0.5 for spinal cord injury and above 0.6 for stroke. Intra-class correlation coefficients were 0.99 and 0.77 for stroke and spinal cord injury, respectively. The newly adapted index showed expected associations with the impairment scales, confirming its construct validity. However, Rasch analysis showed that bladder and bowel items compromise unidimensionality. In conclusion, adaptation of the modified Barthel Index has been successful and it can be used in Turkey as long as its limitations are recognized. PMID- 10853724 TI - How do stroke patients fare when discharged straight to their homes? A controlled study on the significance of hospital follow-up after one month. AB - In our experience, stroke patients discharged straight to their homes sometimes showed marked deterioration. We investigated whether this negative course of events could be prevented by means of follow-up visits entailing extensive testing and resultant measures one month after discharge. The patients in our study included a selection of mild cases with a short length of hospital stay. Forty-six patients returned to the stroke unit on a follow-up visit, and 49 patients made up the control group. The groups were compared after 3 months, by means of questionnaires. The results did not show any definite difference between the groups. However, after 3 months we detected depressions in 13 patients in the study group and in 11 patients in the control group, most of them untreated. The study points to a need for follow-up aimed specifically at detecting depression. PMID- 10853725 TI - How does the brain sustain a visual percept? AB - Perception involves the processing of sensory stimuli and their translation into conscious experience. A novel percept can, once synthesized, be maintained or discarded from awareness. We used event-related functional magnetic resonance imaging to separate the neural responses associated with the maintenance of a percept, produced by single-image, random-dot stereograms, from the response evoked at the onset of the percept. The latter was associated with distributed bilateral activation in the posterior thalamus and regions in the occipito temporal, parietal and frontal cortices. In contrast, sustained perception was associated with activation of the pre-frontal cortex and hippocampus. This observation suggests that sustaining a visual percept involves neuroanatomical systems which are implicated in memory function and which are distinct from those engaged during perceptual synthesis. PMID- 10853726 TI - A model for colour pattern formation in the butterfly wing of Papilio dardanus. AB - The butterfly Papilio dardanus is well known for the spectacular phenotypic polymorphism in the female of the species. We show that numerical simulations of a reaction diffusion model on a geometrically accurate wing domain produce spatial patterns that are consistent with many of those observed on the butterfly. Our results suggest that the wing coloration is due to a simple underlying stripe-like pattern of some pigment-inducing morphogen. We focus on the effect of key factors such as parameter values for mode selection, threshold values which determine colour, wing shape and boundary conditions. The generality of our approach should allow us to investigate other butterfly species. The relationship between these key factors and gene activities is discussed in the context of recent biological advances. PMID- 10853727 TI - Does the presence of ant nests matter for oviposition to a specialized myrmecophilous Maculinea butterfly? AB - More than 50% of the lycaenid butterflies have an ant-associated lifestyle (myrmecophily) which may vary from coexistence to specific mutualistic or even parasitic interactions. Ant-related host-plant selection and oviposition has been observed in some myrmecophilous lycaenids. Therefore, it is remarkable that there is no evidence for this behaviour in the highly specialized, obligate myrmecophilous butterflies of the genus Maculinea. In contrast with previous findings, our results provide evidence for ant-related oviposition patterns in Maculinea alcon in relation to the distribution of specific host-ant nests (i.e. Myrmica ruginodis) based on repeated egg counts during the flight period in two populations. We also show that ant-related oviposition can be counterbalanced by intraspecific competition and oviposition deterrency when host plants already carry several eggs. Therefore, the absence of a correlation between egg load and the presence of host-ant nests at the end of the flight period should be interpreted carefully Whether ovipositional cues are obtained either directly (from ants or their nests) or indirectly (from vegetation structure), and whether alternative explanations based on the phenology and growth form of host plants are possible, is discussed. PMID- 10853728 TI - Male killing can select for male mate choice: a novel solution to the paradox of the lek. AB - In lekking species, intense directional selection is applied to aspects of the male genotype by female choice. Under conventional quantitative genetics theory, the expectation is that this will lead to a rapid loss in additive genetic variance for the trait in question. However, despite female choice, male variation is maintained and hence it pays females to continue choosing. This has been termed the 'paradox of the lek'. Here we present a theoretical analysis of a putative sex-role-reversed lek in the butterfly Acraea encedon. Sex-role reversal appears to have come about because of infection with a male-killing Wolbachia. The bacterium is highly prevalent in some populations, such that there is a dearth of males. Receptive females form dense aggregations, and it has been suggested that males preferentially select females uninfected with the bacterium. As with more conventional systems, this presents a theoretical problem exactly analogous to the lek paradox, namely what maintains female variation and hence why do males continue to choose? We model the evolution of a male choice gene that allows discrimination between infected and uninfected females, and show that the stable maintenance of both female variation and male choice is likely, so long as males make mistakes when discriminating between females. Furthermore, our model allows the maintenance, in a panmictic population, of a male killer that is perfectly transmitted. This is the first model to allow this result, and may explain the long-term persistence of a male killer in Hypolimnas bolina. PMID- 10853729 TI - A comparative study of ejaculate traits in three endangered ungulates with different levels of inbreeding: fluctuating asymmetry as an indicator of reproductive and genetic stress. AB - We studied three closely related species of endangered gazelles (Gazella dorcas, Gazella dama and Gazella cuvieri) with different levels of inbreeding in order to determine at which intensities inbreeding influences ejaculate traits. We also examined whether fluctuating asymmetry (FA) is a reliable indicator of genetic as well as reproductive stress. Our results show that, within each population, the individual coefficient of inbreeding is inversely related to ejaculate quality only in the species with the highest levels of inbreeding (G. cuvieri). In addition, FA is a reliable indicator of individual levels of inbreeding in both the species with the highest levels of inbreeding (G. cuvieri) and the species with intermediate levels of inbreeding (G. dama). Thus, FA appears in individuals whose levels of inbreeding are still not high enough to affect male reproductive potential and should therefore be considered a sensitive indicator of genetic stress. Finally, FA is also a reliable indicator of male reproductive stress since it is related to individual semen quality in all the species studied. PMID- 10853730 TI - Testosterone treatment is immunosuppressive in superb fairy-wrens, yet free living males with high testosterone are more immunocompetent. AB - The immunocompetence handicap hypothesis proposes that the immunosuppressive effect of testosterone enforces honesty of sexual signalling via a physiological trade-off between signal intensity and immunocompetence. However, evidence that testosterone is immunosuppressive is scant, particularly in birds. I studied the correlation between immunocompetence and testosterone in superb fairy-wrens (Malurus cyaneus), a species with intense intersexual selection. Males are seasonally dichromatic and testosterone increases during the moult from dull brown eclipse plumage into bright nuptial plumage. I determined the primary antibody response to immunization with sheep red blood cells (SRBCs) in (i) control and testosterone-implanted males in captivity, and (ii) a cross-section of free-living males with basal and elevated testosterone (in eclipse plumage, moulting and in nuptial plumage). Experimental treatment with testosterone decreased the likelihood of an antibody response to SRBCs in captive birds. In contrast, free-living males which had acquired the nuptial plumage and had naturally elevated testosterone were more likely to respond to SRBCs than males in eclipse plumage with basal testosterone levels. The association between higher immunocompetence and higher immunosuppressive testosterone could arise if both are positively correlated with male phenotypic quality In addition, the association could result if males compensate for potential immunosuppression by enhancing their humoral immune responses, particularly since high testosterone is linked to other demanding activities such as moulting and courtship displays. PMID- 10853731 TI - The T-cell-mediated immune response and return rate of fledgling American kestrels are positively correlated with parental clutch size. AB - Life-history theory predicts that parents face a trade-off between the number and viability of the progeny they produce. We found evidence for an apparent trade off in a free-living population of American kestrels (Falco sparverius), as larger clutches produced more but lighter fledglings. However, while the body mass of fledglings has traditionally been used as a measure of survival prospect, offspring immunocompetence should also play an important role. We thus measured the T-cell-mediated immune response of fledgling kestrels in relation to brood traits and nest-rearing conditions through a cross-fostering experiment. The immune response was positively correlated with the body condition of fledglings, but was also higher in those hatched from five-egg than four-egg clutches. These results were not influenced by other brood traits, nor by current exposure to stressors and infectious agents, as measured by serological variables. Such ability to resist pathogens may account for why the probability of offspring returning to the study area in subsequent years, when controlling for brood size, was higher for five-egg than four-egg clutches. These results suggest an optimal clutch size through maternal effects on offspring immunocompetence rather than a trade-off between the number and quality of the offspring. PMID- 10853732 TI - Using presence-absence data to establish reserve selection procedures that are robust to temporal species turnover. AB - Previous studies suggest that a network of nature reserves with maximum efficiency (obtained by selecting the minimum area such that each species is represented once) is likely to be insufficient to maintain species in the network over time. Here, we test the performance of three selection strategies which require presence-absence data, two of them previously proposed (multiple representations and selecting an increasing percentage of each species' range) and a novel one based on selecting the site where each species has exhibited a higher permanence rate in the past. Multiple representations appear to be a safer strategy than selecting a percentage of range because the former gives priority to rarer species while the latter favours the most widespread. The most effective strategy was the one based on the permanence rate, indicating that the robustness of reserve networks can be improved by adopting reserve selection procedures that integrate information about the relative value of sites. This strategy was also very efficient, suggesting that the investment made in the monitoring schemes may be compensated for by a lower cost in reserve acquisition. PMID- 10853733 TI - Behavioural mimicry of honeybees (Apis mellifera) by droneflies (Diptera: Syrphidae: Eristalis spp.). AB - Droneflies (Syrphidae: Eristalis spp. resemble honeybees (Apis mellifera) in appearance and have often been considered to be Batesian mimics. This study used a focal watch technique in order to compare the foraging behaviour of droneflies Eristalis tenax, Eristalis pertinax, Eristalis arbustorum and Eristalis nemorum) whilst they were feeding on patches of flowers with the behaviour of honeybees and other hymenopterans and dipterans. It was found that, on a range of plant species, the time droneflies spent on individual flowers and the time spent flying between them was more similar to that of honeybees than to the times of other hymenopterans and dipterans. These results suggest that dronefly behaviour has evolved to become more similar to that of honeybees and they support the hypothesis that droneflies are Batesian mimics. PMID- 10853734 TI - Repercussions of El Nino: drought causes extinction and the breakdown of mutualism in Borneo. AB - Figs (Ficus spp.) and their species-specific pollinators, the fig wasps (Agaonidae), have coevolved one of the most intricate interactions found in nature, in which the fig wasps, in return for pollination services, raise their offspring in the fig inflorescence. Fig wasps, however, have very short adult lives and hence are dependent on the near-continuous production of inflorescences to maintain their populations. From January to March 1998 northern Borneo suffered a very severe drought linked to the El Nino-Southern Oscillation event of 1997-1998. This caused a substantial break in the production of inflorescences on dioecious figs and led to the local extinction of their pollinators at Lambir Hills National Park, Sarawak, Malaysia. Most pollinators had not recolonized six months after the drought and, given the high level of endemism and wide extent of the drought, some species may be totally extinct. Cascading effects on vertebrate seed dispersers, for which figs are often regarded as keystone resources, and the tree species dependent on their services are also likely. This has considerable implications for the maintenance of biodiversity under a scenario of climate change and greater climatic extremes. PMID- 10853735 TI - Body postures and patterns as amplifiers of physical condition. AB - The question of why receivers accept a selfish signaller's message as reliable or 'honest' has fuelled ample controversy in discussions of communication. The handicap mechanism is now widely accepted as a potent constraint on cheating. Handicap signals are deemed reliable by their costs: signallers must choose between investing in the signal or in other aspects of fitness. Accordingly, resources allocated to the signal come to reflect the signaller's fitness budget and, on average, cheating is uneconomic. However, that signals may also be deemed reliable by their design, regardless of costs, is not widely appreciated. Here we briefly describe indices and amplifiers, reliable signals that may be essentially cost free. Indices are reliable because they bear a direct association with the signalled quality rather than costs. Amplifiers do not directly provide information about signaller quality, but they facilitate assessment by increasing the apparency of pre-existing cues and signals that are associated with quality. We present results of experiments involving a jumping spider (Plexippus paykulli) to illustrate how amplifiers can facilitate assessment of cues associated with physical condition without invoking the costs required for handicap signalling. PMID- 10853736 TI - Food-related bray calls in wild bottlenose dolphins (Tursiops truncatus). AB - Because cetaceans are difficult to study in the wild, little is known about how they use their sounds in their natural environment. Only the recent development of passive acoustic localization systems has enabled observations of the communication behaviour of individuals for correlation with their surface behaviour. Using such a system, I show that bottlenose dolphins in the Moray Firth, Scotland, produce low-frequency bray calls which are clearly correlated with feeding on salmonids. The production of these calls is followed by fast approaches by conspecifics in the area. In animals which use sound as a foraging tool, it is difficult to distinguish between food calls which have evolved because of their role in attracting conspecifics, and food manipulation or searching calls which may attract conspecifics as a by-product. However, the low frequency structure of the bottlenose dolphin bray suggests that it evolved because of a role in manipulating prey rather than in attracting conspecifics. This conclusion suggests that dolphins exploit the perceptual systems of their prey to facilitate capture. PMID- 10853737 TI - Fragmentation of sea bass populations in the western and eastern Mediterranean as revealed by microsatellite polymorphism. AB - We studied the genetic structure at six microsatellite loci of the Mediterranean sea bass (Dicentrarchus labrax) on 19 samples collected from different localities in the western and eastern Mediterranean basins. Significant divergence was found between the two basins. The distance tree showed two separate clusters of populations which matched well with geography, with the noticeable exception of one Egyptian sample which grouped within the western clade, a fact attributable to the introduction of aquaculture broodstock. No heterogeneity was observed within the western basin (theta = 0.0014 and n.s.). However, a significant level of differentiation was found among samples of the eastern Mediterranean (theta = 0.026 and p < 0.001). These results match with water currents but probably not with the dispersal abilities of this fish species. We thus hypothesize that selective forces are at play which limit long-range dispersal, a fact to be taken into account in the debate about speciation processes in the marine environment. PMID- 10853738 TI - Female barn owls (Tyto alba) advertise good genes. AB - The good genes hypothesis of sexual selection postulates that ornamentation signals superior genetic quality to potential mates. Support for this hypothesis comes from studies on male ornamentation only, while it remains to be shown that female ornamentation may signal genetic quality as well. Female barn owls (Tyto alba) display more black spots on their plumage than males. The expression of this plumage trait has a genetic basis and it has been suggested that males prefer to mate with females displaying more black spots. Given the role of parasites in the evolution of sexually selected traits and of the immune system in parasite resistance, we hypothesize that the extent of female plumage 'spottiness' reflects immunological defence. We assessed the genetic variation in specific antibody production against a non-pathogenic antigen among cross fostered nestlings and studied its covariation with the plumage spottiness of genetic parents. The magnitude of the antibody response was positively correlated with the plumage spottiness of the genetic mother but not of the genetic father. Our study thereby provides the first experimental support, to our knowledge, for the hypothesis that female ornamentation signals genetic quality. PMID- 10853739 TI - Step-parents and infanticide: new data contradict evolutionary predictions. AB - Evolutionary psychologists have hypothesized, inspired by evolutionary biology, that parents should care less for children with whom they are not genetically related since these young do not contribute to the genetic fitness of the parents. Based on this, evolutionary psychologists have predicted that there will be an overrepresentation of step-parents as offenders in family-related killings of children. Data on child homicide, particularly from Canada, have supported this prediction in that the frequency of children killed was relatively high in families where one of the two parents was a step-parent. Here we present a survey of all child homicide that occurred in Sweden between 1975 and 1995. In contrast to the Canadian data, children in Sweden living in families with a step-parent were not at an increased risk compared with children living together with two parents to whom they were genetically related. In addition, there were no other indications that step-parents are overrepresented as offenders. PMID- 10853740 TI - The size distribution of conspecific populations: the peoples of New Guinea. AB - The size distribution of the language populations in New Guinea, which represent over 15% of the world's languages, is analysed using models analogous to the resource division models of species abundance distribution in ecological communities. A model distribution of resource segments reflecting population size is created by repeated selection of an existing resource segment and its division into two. We found that any dependency of the selection probability on the size of the segment generated negatively skewed abundance distributions after log transformation. Asymmetric segment division further exacerbated the negative skewness. Size-independent selection produced lognormal abundance distributions, irrespective of the segment division method. Size-dependent selection and asymmetric division were deemed reasonable assumptions since large language populations are more likely to generate isolates, which develop into new populations, than small ones, and these isolates are likely to be small relative to the progenitor population. A negatively skewed distribution of the log transformed population sizes was therefore expected. However, the observed distributions were lognormal, scale invariant for areas containing between 100 and over 1000 language populations. The dynamics of language differentiation, as reflected by the models, may therefore be unimportant relative to the effect of variable growth rates among populations. All lognormal distributions from resource division models had a higher variance than the observed one, where half of the 1053 populations had between 350 and 3000 individuals. The possible mechanisms maintaining such a low variance around a modal population size of 1000 are discussed. PMID- 10853741 TI - Referring patients abroad for proton therapy. PMID- 10853742 TI - Obtaining funding for proton therapy at Orsay, France. PMID- 10853743 TI - Proton therapy for base of skull chordoma: a report for the Royal College of Radiologists. The Proton Therapy Working Party. AB - Chordomas of the base of the skull are rare. They are locally infiltrative and frequently arise close to radiosensitive structures, which limits the ability to deliver a high dose of radiotherapy. Complete surgical excision is not usually possible. Conventional postoperative radiotherapy can result in approximately 50% 5-year survival and effective palliation, but long-term local control and cure are rare. The well-defined Bragg peak of protons allows planning with a sharp cut off outside the target volume. This permits a higher dose of radiotherapy to be delivered to the tumour while avoiding excessive irradiation to radiosensitive structures. Outcome after proton irradiation is superior to that reported for conventional photon irradiation. Radiotherapy schedules involving a mixed schedule of protons and photons have achieved an approximately 60% local control rate at 5 years. Some of this improvement may have resulted from better surgical techniques. Proton irradiation is also effective for base of skull chondrosarcomas. Protons of sufficient energy to treat base of skull tumours are not available in the UK. Patients have been referred to the proton facilities at the Harvard cyclotron, and at Orsay, France. They will continue to require referral abroad for proton therapy for base of skull tumours. Proton therapy has become standard treatment for ocular melanoma and this is available at the Douglas cyclotron at Clatterbridge Hospital. Proton therapy has the potential for improved dose distribution compared with conformal photon radiotherapy. This may be exploited effectively to irradiate target volumes close to radiosensitive structures such as the spinal cord. There is a need for further clinical research to evaluate proton therapy for tumours such as spinal and paraspinal sarcomas, and paediatric brain tumours. PMID- 10853744 TI - Management issues in chordoma: a case series. AB - This study presents a combined case series of chordomas from two Canadian institutes. Twenty-seven patients were identified for the period 1954-1998. Management issues with regard to diagnostic pitfalls, selection of charged particle treatment and retreatment of recurrences are discussed. The diagnosis of early stage chordoma requires a high index of suspicion. One patient in the series presented with hoarseness and is described in detail. The diagnosis was made coincidentally by a computed tomographic scan of the head, performed after a motor vehicle accident. The planning of both surgery and radiotherapy was optimized by using magnetic resonance imaging. A review of the literature supports the use of a combined surgical and radiotherapeutic approach. PMID- 10853745 TI - What is the role of high-dose therapy in the management of lymphoma? PMID- 10853746 TI - The road to Hodgkin's disease and on to the millennium. PMID- 10853747 TI - Lhermitte's sign following head and neck radiotherapy. AB - Lhermitte's sign is an uncommon sequel of radiotherapy to the cervical spinal cord. Although the exact mechanism underlying its occurrence remains unclear; it is felt to be the result of a temporary interference with the turnover and synthesis of myelin, leading to focal demyelination. We have undertaken a detailed analysis of the radiation delivered to four patients who developed the sign after irradiation for malignancies of the head and neck. Our data support the view that radiation dose is crucial to its development, but calculations using the linear-quadratic radiobiological model raise interesting questions regarding the dose-response relationship. In particular, we find that calculations of biologically effective doses are predictive of a late rather than an early normal tissue response. The onset of symptoms after irradiation was apparent in all four patients within 4 months, with resolution in all being complete within a further 6 months. The recognition of this benign transient form of radiation-induced paraesthesia and its differentiation from the later onset, progressive and unremitting symptoms associated with radiation myelopathy is essential in reassuring patients undergoing head and neck irradiation. PMID- 10853748 TI - Radiotherapy outpatient review: a nurse-led clinic. AB - The aim of this project was to improve the quality of the delivery of care to patients undergoing outpatient fractionated radiotherapy, in terms of effectiveness and efficiency. This article provides a descriptive account of the activities that took place within a traditional doctor-held radiotherapy review (floor) clinic (71 individual clinic episodes from 71 patients) and compares them with a nurse-led clinic (299 clinic episodes from 141 patients). The outcome measures encompassed the number and type of activities in both clinics, including the number of interventions that occurred, consultation time, waiting time, degree of involvement with other support services, reasons for doctor contact, and perceptions of patients, doctors, nurses and therapy radiographers. Nurse consultations lasted longer than doctor consultations. Waiting times were reduced. Of 299 nurse-led clinic episodes, only 21 contacts were made with the doctor. More interactions and activities occurred during nurse consultations. A greater number of referrals and liaisons with other support services occurred, resulting in the establishment of continuity of care. The results from this small sequential observational study require prospective confirmation, but they suggest that specialist nursing staff, given appropriate medical support, may provide more effective care for patients who are undergoing outpatient radiotherapy. PMID- 10853749 TI - Oncologist's nephropathy. AB - A 46-year-old, previously fit man underwent standard primary chemotherapy and then further standard chemotherapy, followed by high-dose chemotherapy (without total body irradiation) and an autologous marrow transplant for relapsed non Hodgkin's lymphoma. He also received antibiotics, antifungals and antivirals during this time. He developed episodic renal impairment with remissions, but his renal function never returned to baseline. Renal biopsy demonstrated the lesion to be due to thrombotic microangiopathy; he was treated by an angiotension II receptor antagonist, low-dose aspirin and warfarin. Seven years later the renal function remains mildly impaired but stable. This unusual iatrogenic nephropathy is discussed. PMID- 10853750 TI - Secondary acute myeloblastic leukaemia (AML) (expressing 11q23 mutation) occurring 11 months after chemotherapy/radiotherapy for paediatric non-Hodgkin lymphoma (NHL). AB - The occurrence of 11q23 cytogenetic abnormalities in drug-induced acute myeloid leukaemia (AML) is now well recognized. They are most frequently associated with topoisomerase II inhibitor administration. We here describe the case history of a 15-year-old child who presented with Stage III B-cell non-Hodgkin's lymphoma (NHL) and was treated with anthracyclines, alkylating agents and low-dose mediastinal radiotherapy. She developed an 11q23 mutation-related secondary AML at 11 months after therapy (15 months after her first exposure to drugs; 12 months after the first radiotherapy exposure), possibly the earliest yet reported in a paediatric patient. We discuss this newly recognized early form of refractory, secondary AML and its relationship to chemoradiotherapy. PMID- 10853751 TI - Opportunistic infection with Rhodotorula in cancer patients treated by chemotherapy: two case reports. AB - Rhodotorula species are commensal yeasts of variable pathogenicity. The authors report the case histories of two patients presenting with febrile neutropenia. The first was a 3-year-old girl who had been treated with combination chemotherapy for a tumour of the posterior fossa. The second was a 46-year-old man who had received chemotherapy for lymphoplasmocytic lymphoma, followed by consolidation treatment with autologous bone marrow transplantation. Investigation revealed infection caused by Rhodotorula. The outcome was favourable after removal of the catheter in both patients. Rhodotorula species have been isolated during a variety of infectious complications. Almost all published cases of fungaemia concern patients with central venous catheters that have been in place over long periods, who have also been treated with broad spectrum antibiotics. Neoplasia represents the most frequent underlying disease. The pathogenicity of Rhodotorula species appears to be moderate in most cases; fungal therapy or the removal of infected catheters is generally effective. Nevertheless, Rhodotorula has been reported to provoke fatal endocarditis or meningitis and can probably cause septic shock. PMID- 10853752 TI - Gas in the cranium: an unusual case of delayed pneumocephalus following craniotomy. AB - We present the case history of a 23-year-old man who underwent frontal craniotomy followed by radiotherapy for a Grade III anaplastic glioma. Magnetic resonance imaging (MRI) at the 3-month follow-up showed significant tumour response. He became unwell some weeks after the MRI with an upper respiratory tract infection, severe headache and mild right-sided weakness. A computed tomographic (CT) scan showed a very large volume of intracranial gas, thought to have entered via a defect in the frontal air sinus after craniotomy and brought to light by blowing his nose. Intracranial air is frequently present after craniotomy, but it is normally absorbed within 34 weeks. The presence of pneumocephalus on a delayed postoperative CT scan should raise the possibility of a cerebrospinal fluid (CSF) fistula, or infection with a gas-forming organism. Many CSF fistulae require surgical closure in order to prevent potentially life-threatening central nervous system infection and tension pneumocephalitis. Immediate neurosurgical review is advisable. PMID- 10853753 TI - Exceptional results in neuroendocrine-metastases-caused paraplegia treated with [90Y-DOTA]-D-Phe1-Tyr3-octreotide (90Y-DOTATOC), a radiolabelled somatostatin analogue. AB - The case history is presented of a patient with paraplegia caused by progressive spinal cord compression due to bone metastases of a neuroendocrine pulmonary tumour. After failed external radiotherapy, the patient received targeted internal radiotherapy administered as a fractionated treatment with intravenous injections of a total of 7400 MBq/m2 of [90YDOTA]-D-Phe1-Tyr3-octreotide (90Y DOTATOC), a radiolabelled somatostatin analogue. This case history highlights the value of 90Y-DOTATOC in the treatment of neuroendocrine tumours and the importance and possibility of good palliation of neuroendocrine bone metastases. PMID- 10853754 TI - Cerebral haemangiopericytoma treated with conservative surgery and radiotherapy. AB - Cerebral haemangiopericytomas are rare tumours that resemble meningiomas but behave more aggressively, with a tendency to metastasize. We report two patients with haemangiopericytoma who had limited surgical resections owing to perioperative blood loss but who had massive tumour shrinkage after a course of radical radiotherapy. We suggest a more conservative surgical approach to the management of these tumours. PMID- 10853755 TI - Position emission tomography scanning and early breast cancer. PMID- 10853756 TI - Lung cancer with synchronous brain and bone metastasis. PMID- 10853757 TI - Introduction: basic concepts and definitions in mechanics. AB - This introduction aims to expound the basic ideas of mechanics of materials to clinicians. What happens when a bone (mechanically a beam) is submitted to standard loads: Centric axial load, bending, eccentric axial load, torque? How does the bone deform? The basic ideas of mechanical engineering are presented using an eraser under load as an illustrative object, and trying to maintain the mathematical formulae as far as possible. PMID- 10853758 TI - Stiffness--an unknown world of mechanical science? AB - "Stiffness" is a term used to describe the force needed to achieve a certain deformation of a structure. In the biomechanical world, several different definitions of stiffness are used, but not all of them are explained adequately to those readers who are less familiar with biomechanical terminology. This paper gives examples for specific definitions which are based on the basic definition of stiffness of a loaded structure PMID- 10853759 TI - Graphical statics a forgotten tool for solving plane mechanical problems. AB - Graphical statics is an almost forgotten, intuitive drawing method for solving plane mechanical problems. It was already in use in the 19th century for biomechanical problems. It was still a standard method employed by civil engineers in the 1940s. Superceded by modern analytical methods, graphical statics disappeared almost completely. The method is restricted to plane static problems, but still remains a useful tool for visualizing, understanding and checking the actions of force groups occurring in modern biomechanical problems. After defining the basic mechanical terminology (body, motion, forces), the paper is written mainly as a teaching tool for immediate application. Many illustrative examples (sporting activities, functional forces in joints) help to clarify the difficult biomechanical content. For application, it must be assumed that the bodies investigated behave as rigid bodies under the action of the forces, but this does not prevent application of the method to deformable living bodies if specific static configurations of the bodies are considered. The application of the method requires a good anatomical knowledge and experience with the function of the musculoskeletal apparatus of living bodies. If reliable models are used, the method delivers quantitative results of sufficient accuracy. The paper may also help provide a better understanding of publications containing graphical solutions to bio-static problems. PMID- 10853760 TI - Functional load of plates in fracture fixation in vivo and its correlate in bone healing. AB - In clinical practice efforts are made to apply a fixation plate on the side opposite the strongest muscle pull. This achieves an optimal distribution of compression between the fragment ends (principle of tension band plating). This is however frequently impossible for anatomical or surgical reasons. In an 'in vivo' study lasting 8 weeks a standardized oblique osteotomy was performed on the tibia of 16 sheep in four different models of tension band plating (a contoured and an overbent plate with or without an interfragmentary lag screw) were assessed. Tension on the plate surface was recorded by strain gauges for different gait speeds on the treadmill. These measurements were performed throughout the experiment. Radiographs were taken at regular intervals in order to assess stability and polychrome sequential labelling and microradiographs served to investigate the healing process. Possible relationships and/or interactions between plate tension and bone healing were investigated. Implant loading under bending strain was reduced the most for the combination of plate overbending with a lag screw. The insertion of a lag screw reduces the surface strain on the plate whether it is contoured or overbent. The bending and torsional forces are greatest if a straight plate is used alone and the principle of tension band plating is not applied. Direct bone healing was only observed in the group with contoured plate and lag screw. Overbending combined with a lag screw provided only a relatively unstable fixation. A residual gap immediately beneath the plate permits "dynamic compression" since the screws slide towards the osteotomy when loaded producing bone resorption under the plate and signs of screw loosening. The models with contoured and overbent plates without a lag screw were histologically assessed as very unstable with signs of secondary fragment displacement, obvious callus formation, resorption at the fragment ends and under the plate, delayed and diminished Haversian remodelling and corrosion sites at the screw heads and at the adjacent site on the plate hole. In all groups, stripping of the periosteum under the plate was associated with porosis of the corresponding cortex as a sign of temporarily impaired blood supply. A relationship between implant loading and/or unloading (stress shielding) could not be demonstrated. Callus formation, measured quantitatively on the radiographs, is directly related to the strain on the plate. Direct bone healing is rapid and is seen histologically three weeks postoperatively, particularly for fixations with contoured plate and lag screw. The early appearance of fixation callus in the presence of an intact blood supply indicates a primary instability of the osteosynthesis. Later, it may be an indication of secondary instability. The time at which osteons appear, their number and location provides information on the stability of the osteosynthesis. At a time when indirect fracture reduction and stabilization using minimally invasive techniques and implants is being propagated, additional ways and means must be sought to assess clinically the load on the implants and the risk of implant failure. PMID- 10853761 TI - Long-term effects of plate osteosynthesis: comparison of four different plates. AB - Various phenomena have been observed subsequent to plate osteosynthesis, for example, refracture after plate removal. Experimental research has shown that changes in the cortex occur within the first three months after plating and again several months later. These changes are independent of the fracture and take the form of porosis under the plate and excessive bone growth around the plate. Porosis under the plate was regarded until recently as being due to unloading of the bone by the plate, also known as stress shielding. Investigations of the relationship between bone porosis and the changes in periosteal blood supply due to its compression by the plate, however, have been neglected. In this study, the effect of plate properties such as structural stiffness ('unloading'), implant material, and plate contact surface (altered periosteal blood supply) on bone after osteosynthesis were investigated. This was done by comparative histomorphometry of the altered bone in sheep after application of four plates differing in the above-mentioned properties. After plating the sheep tibia with a trapezoid plate with narrow contact surface, significantly larger bone cross sections were observed one year after the operation and considerable bone growth around the plate. The area of early temporary porosis in the cortex under the plate as observed in the first nine weeks and after one year was not significantly different for any particular plate, all of which were applied subperiosteally. The trapezoid plates were easier to remove, thus causing less damage to the bone lamellae along the sides of the plate. The marked increase in bone cross section after one year and the larger areas of bone growth around the trapezoid plates with smaller contact surface can be attributed to the larger grooves cut along the sides of the plates. Cortical porosis was mainly the result of impaired periosteal blood supply which was of equal size in all groups as a result of careful periosteal stripping and subperiosteal plate insertion. It was assumed that applying the plate onto the periosteum would be associated with effects on periosteal blood supply directly related to the plate and consequently cortical porosis. Plate related stress shielding and the implant material had no significant effect on the extent of cortical porosis. PMID- 10853762 TI - Prevention of malunions in the rotation of complex fractures of the distal femur treated using the Dynamic Condylar Screw (DCS): an anatomical graphic analysis using computed tomography on cadaveric specimens. AB - The AO ASIF dynamic condylar screw is a popular method for the treatment of severe distal femoral fractures. Two methods of application have been presented by the AO group, one in the AO manual and the other in the teaching video tapes. The small difference in the techniques suggested that one of these methods might present a risk of rotational malunion in cases of severe comminuted fractures. The question of determining which method was less appropriate was investigated on human cadaveric specimens using a graphic analysis of computed tomograms. We found disadvantages for both methods and propose a third method combining the advantages of the two original methods. PMID- 10853763 TI - A screen of yeast respiratory mutants for sensitivity against the mycotoxin citrinin identifies the vacuolar ATPase as an essential factor for the toxicity mechanism. AB - In countries with a hot climate the mycotoxin citrinin represents a serious problem in fungal food-poisoning. In humans the renal system is affected the most and the mitochondrial respiratory chain was identified as a possible sensitive target for this toxin. In addition, citrinin has an antifungal activity that also inhibits the growth of the yeast Saccharomyces cerevisiae. So far the precise mode of action and the subcellular targets for citrinin have not been identified. Therefore, we decided to use the model organism yeast for a genetic approach to identify genes that play a role in the sensitivity against this mycotoxin. A large collection of conditional respiratory deficient yeast mutants was screened for sensitivity against citrinin. One special pet-ts mutant was identified that exhibited a higher sensitivity against citrinin. The genetic system of yeast allowed the isolation of the respective wild-type gene. This yeast gene encodes the Vph2p subunit that is essential for the correct assembly of the vacuolar ATPase. Isolation of the mutated gene and gene-disruption experiments of VPH2 and the partially overlapping small YKL118W gene verified this finding. The wild-type VPH2 gene restores all defects of the mutants. In contrast to this, YKL118W gave no complementation and the null mutant showed no phenotype. Thereby the yeast vacuolar ATPase was found to be important for the toxic effect of citrinin in yeast cells. The consequences of this finding for the molecular mechanism of citrinin action and its relation to the mitochondrial respiratory chain are discussed. PMID- 10853764 TI - Recessive mutations in SUP35 and SUP45 genes coding for translation release factors affect chromosome stability in Saccharomyces cerevisiae. AB - Chromosome stability in suppressor mutants for SUP35 and SUP45 genes coding for translation release factors was studied. We obtained spontaneous and UV-induced sup35 or sup45 mutants in a haploid strain disomic for chromosome III and tested the stability of an extra copy of this chromosome. The majority of the mutants showed increased chromosome instability. This phenotype was correlated with an increased sensitivity to the microtubule-poisoning drug benomyl which affects chromosome segregation at anaphase. Our data suggest that termination-translation factors eRF3 and eRF1 control chromosome transmission at mitotic anaphase in Saccharomyces cerevisiae. PMID- 10853765 TI - Influence of homology size and polymorphism on plasmid integration in the yeast CYC1 DNA region. AB - We studied the influence of homology size and polymorphism on the integration of circular plasmids into the yeast CYC1 region. The plasmids used also contained the URA3 gene, and the proportion of Ura+ transformants resulting from plasmid integration into the CYC1 region was determined by Southern-blot analysis. A size dependent decrease in integration into the CYC1 region was observed from 858 bp to 363 bp of homology. However, with a homology size of 321, 259 or 107 bp, about 2% of the transformants still contained plasmid molecules integrated in the CYC1 region. A single point mutation in the 858-bp fragment decreased the proportion of integrations to the CYC1 gene, but the presence of additional mutations did not have a cumulative effect. For plasmids isolated in a single-stranded (ss) form, the presence of two or six point mutations did not influence integration. These results were compared with those obtained in other assays designed to study substrate requirements for homologous recombination. PMID- 10853766 TI - ARS sequences in homologous and heterologous ADE2 loci are capable of promoting autonomous replication of plasmids in Schwanniomyces occidentalis. AB - We have analyzed ARS elements linked to homologous and heterologous ADE2 loci functioning in Schwanniomyces occidentalis. We have identified a region of the ADE2 locus of S. occidentalis which promotes autonomous replication of plasmids in S. occidentalis cells. This region is within 385 bp preceding the ATG codon of the S. occidentalis ADE2 gene. It contains sequences similar to ARS core consensus sequences, ARS boxes, and a potential transcription activator binding site characterized in Saccharomyces cerevisiae. The ADE2 gene of S. cerevisiae was found to complement the ade2 mutation in S. occidentalis cells and the 5' UTR region of this gene is capable of supporting autonomous replication of plasmids in S. occidentalis. Furthermore, we confirmed that the origin of replication of the 2 microm plasmid and the ARS1 sequence of S. cerevisiae are also functional in S. occidentalis cells. Plasmids carrying either ARS, the SwARSA element of S. occidentalis, the ARS linked to the ADE2 gene of S. cerevisiae, and the ARS1 sequence or the 2 microm ori, were found to be maintained in S. occidentalis cells as episomal monomers or oligomers. However, their stability was low as already reported for the ARS in S. occidentalis. PMID- 10853767 TI - Phage T4-like intermediates of DNA replication and recombination in the mitochondria of the higher plant Chenopodium album (L.). AB - We have studied intermediates of the recombination and replication of chromosomal mitochondrial (mt) DNA prepared from suspension cultured cells of Chenopodium album (L.) by electron microscopy during the whole growth cycle. We identified several types of potential recombination and replication intermediates including rosette-like structures, as well as other branched and sigma-like molecules. The absolute and relative amounts of these structures changed dramatically during the growth cycle, indicating high dynamics in the structural organization of the mt genome. The rosette-like molecules had sizes of 2-5 genome units and were found to contain putative replication forks and 'Holliday'-junctions known from recombination intermediates. The high number of rosettes during the first days of culture, and their drastic reduction in the stationary growth stage, were found to be inversely related to changes in the quantity of linear molecules of 40-200 kb. This observation suggests that linear molecules participate in the formation of giant branched rosette-like structures. Most linear molecules were previously found to have at least one single-stranded end, which may allow recombinative invasion of other double-stranded molecules. Thus, recombination events may lead to the formation of more complex molecules and initiate replication similar to phage T4. We propose the coexistence of a recombination-dependent mode of replication with a presumably recombination-independent rolling-circle mode of replication in the mitochondria of C. album. PMID- 10853768 TI - The tobacco apocytochrome b gene predicts sensitivity to the respiratory inhibitors antimycin A and myxothiazol. AB - The potential for use of the cytochrome-pathway electron-transfer inhibitors antimycin A and myxothiazol in the selection of plant mitochondrial genome transformants was investigated. The net growth of Nicotiana tabacum L. (tobacco) suspension-culture cells was reduced by these inhibitors, but complete repression of cell growth occurred only in the presence of both cytochrome and alternative electron-transfer-pathway inhibitors. Antimycin A and myxothiazol bind to and block electron transfer through different sites in the cytochrome b (COB) subunit of the mitochondrial bc1 respiratory complex (complex III). The nucleotide sequence of the tobacco cob gene was determined and found to predict highly conserved glycine and phenylalanine residues that are associated with sensitivity to antimycin A and myxothiazol, respectively. These residues are altered by mutations that confer resistance to antimycin A or myxothiazol in diverse organisms. Tobacco cob cDNA clones were constructed and sequenced, revealing eight full and 11 partial RNA-editing sites. RNA editing did not, however, alter codons for the conserved glycine and phenylalanine residues associated with sensitivity to the respiratory inhibitors. Antimycin A or myxothiazol, in conjunction with a modified cob gene, may therefore be useful in the selection of tobacco cells carrying a genetically transformed mitochondrial genome. PMID- 10853769 TI - Comprehensive cloning and expression analysis of glycolytic genes from the filamentous fungus, Aspergillus oryzae. AB - We cloned all the glycolytic genes from Aspergillus oryzae and analyzed their transcriptional regulation by the carbon source in the medium. The deduced amino acid sequences of the glycolytic genes showed high identity (approximately 41 93%) to those from other lower eukaryotes. Genomic Southern hybridization indicated that all the genes existed as a single copy in the genome. Comparison of mRNA levels between mycelia grown on glucose and on pyruvate showed that most of the A. oryzae glycolytic genes were induced by glucose in the medium. The overall expression profiles of the A. oryzae glycolytic genes resembled those of Saccharomyces cerevisiae. The expression of one of the phosphofructokinase genes (pfkB), however, was repressed by glucose while both PFK1 and PFK2 were induced in S. cerevisiae. These findings indicate that further analysis of the transcriptional regulation of the A. oryzae glycolytic genes will be useful for investigating the evolutionary change of transcription regulation in lower eukaryotes and to construct promoters for industrial applications. PMID- 10853770 TI - Mutation of a putative AMPK phosphorylation site abolishes the repressor activity but not the nuclear targeting of the fungal glucose regulator CRE1. AB - In filamentous ascomycetes, glucose repression is mediated by CRE1, a zinc-finger protein related to Miglp from yeast. Five putative AMPK phosphorylation motifs identified in the glucose repressor from the phytopathogenic fungus Sclerotinia sclerotiorum were mutated in a GFP::CRE1 translational fusion. Complementation experiments in Aspergillus nidulans and fluorescence microscopy analyses showed that mutation of one site (Ser266) abolishes the repressor activity of the fusion protein but not its nuclear targeting, suggesting that an AMPK protein kinase may be involved in the function of the fungal glucose repressor. PMID- 10853771 TI - Cloning and heterologous expression of Solorina crocea pyrG. AB - A pyrG gene, encoding orotidine 5'-monophosphate decarboxylase, was cloned from a phage library derived from the lichen Solorina crocea. Phylogenetic analysis and a survey of geographically well-separated specimens were used to verify that the gene represented the fungal component of the lichen. Both coding and upstream sequences of S. crocea pyrG exhibited features typical of fungal genes. A 132-bp intron interrupting the coding region between nucleotides 157 and 288 was confirmed by RT-PCR and sequencing. Transformation of Aspergillus nidulans with S. crocea pyrG, controlled by either its native promoter or the A. nidulans trpC promoter, resulted in uridine-independent strains that exhibited appreciable growth only at 24 degrees C. Southern analysis indicated multiple integrations of S. crocea pyrG. These results demonstrate that heterologous expression may be used to investigate genes from lichens. PMID- 10853772 TI - Genome organization in Fusarium oxysporum: clusters of class II transposons. AB - Several families of transposable elements (TEs) are present in the genome of the phytopathogenic fungus Fusarium oxysporum. They are present in copy numbers ranging from just a few elements to tens or hundreds per genome. Sequence analysis of contiguous stretches of genomic DNA surrounding insertion sites of one family revealed that they are packed with repeated sequences. We have carried out a detailed study of the composition and arrangement of these repeats in three chromosomal regions. We found that they are essentially mixtures of several types of TEs, most of them being DNA transposons, different from those previously characterized. Some repeats are frequently reiterated and many of them are inserted into other elements. Parts of these regions are also duplications. These regions appear prone to rearrangement and transposition and are subject to rapid reorganization. PMID- 10853773 TI - Skin allografts--lymph veiled (dendritic) cells are responsible for initiation of rejection in canine skin/SCID mouse chimera model. AB - Skin allografts are acutely rejected despite of intensive immunosuppressive therapy. Resistance of skin dendritic cells to immunosuppressive drugs and irradiation may be responsible for this phenomenon. Skin allograft is a site of interaction between the dendritic cells and lymphocytes of the donor and host origin and "direct" and "indirect" pathway of antigen presentation. Increasing evidence supports the significant role for the "indirect" allorecognition in graft rejection. To investigate a critical role of skin dendritic cells in the "indirect" allorecognition and graft destruction we have used a canine skin to severe-combined-immunodeficient (SCID)-mice transplantation model. At the time the skin grafts were deprived of own dendritic (Langerhans) cells, SCID mice were reconstituted with allogeneic canine whole lymph leukocytes, lymph lymphocytes, lymph veiled (dendritic) cells or peripheral blood mononuclear cells, and an early phase of skin rejection was evaluated in histopathological studies. We demonstrated that circulating canine allogeneic veiled cells facilitate recruitment of T lymphocytes into skin graft and promote an extensive graft destruction, compared to the less expressed effect of allogeneic peripheral lymph lymphocytes or blood mononuclear cells. These drug and radiation-resistant dendritic cells may be responsible for initiation of the difficult to control rejection process. PMID- 10853774 TI - Hepatocyte transplantation--in vitro cytotoxic reaction of autologous granulocytes and mononuclears to isolated hepatocytes. AB - Hepatocyte (HC) transplantation (tx) may be useful for bridging patients to whole organ transplantation and for providing metabolic support during liver failure and for replacing whole organ transplantation in certain liver metabolic diseases. In specific situations where the death rate of host hepatocytes is high, the transplanted cells can repopulate the native liver. Successful transplantation of hepatocytes is hampered by lack of proper cellular (stromal) and humoral (cytokines) environment at the site of implantation. We have found that another factor responsible for low in vivo survival rate of transplanted HC is their rapid destroying by host granulocytes and monocytes. AIM. In this study we investigated the in vitro process of destruction of HC by granulocytes and mononuclear cells, the phenotypes of effector mononuclears and the tempo of HC lysis. METHODS: In vitro cell-mediated cytotoxicity, HC-PMN and HC-PBM rosette formation rate and HC lysis, as well as phenotypes of HC-adhering cells were investigated. RESULTS: Granulocytes formed rosettes with HC almost immediately after the beginning of incubation and were found highly cytotoxic to HC. The cellular mechanism of lysis was not mediated by serum natural antibodies. Also the in vitro mixed HC-granulocyte 51Cr test showed high granulocyte cytolytic activity. Monoclonal antibodies to class I and II antigens, CD11/18 and 54 did not block the granulocyte cytotoxicity. Blood mononuclear cells also formed rosettes with HC and were cytotoxic to them, but the level of cytotoxicity was lower than of granulocytes. ED1+ monocytes revealed highest cytolytic activity toward HC. Hepatocyte contained only trace levels of endotoxin and no chemotactic activity of granulocytes and monocytes toward HC could be observed. CONCLUSIONS: A random physical contact of blood leukocytes seems necessary for adhesion to isolated HC. Taken together, granulocytes and monocytes recognize intercellular surface molecules on HC "exposed during isolation" from the hepatic trabeculae as "non-self" and lyse HC by direct contact. PMID- 10853775 TI - Use of allografts in spinal surgery. AB - This is a review of 118 patients operated between January 1989 and February 1999, 78 for posterior fusions and 40 for anterior spinal reconstructions. For posterior fusion, meticulous preparation of the fusion bed was essential. The author strongly advocated using autografts from spinous processes for achieving facet joint fusion. Deep frozen allografts were used as a 50% allograft-autograft mixture. Good results were achieved -- no infection in all cases and implant failure with clinical non-union in only 5 cases. Massive anterior spinal reconstruction following corpectomy was performed in 34 cases. Allografts used included 15 femoral cortical rings, 11 tibial rings and 3 humeral cortical rings. The author advocated the use of an allograft-autograft composite by packing the medullary canal of the allograft with autografts. Deep-frozen cortical rings have been used (25 cases). Recently, more freeze-dried cortical rings made available were used (9 cases). The results of massive reconstructions were good. There was no infection and no implant failure in all cases. Both deep-frozen and freeze dried cortical allografts were strong enough to withstand loads taken by the reconstructed spine. PMID- 10853776 TI - Behaviour of bone allografts in the advancement of the tibial tuberosity. AB - In order to determine the behaviour of bone allografts in the advancement of the tibial tuberosity, we studied retrospectively 134 knees belonging to 119 recipients of frozen bone allograft for the treatment of a symptomatic patellofemoral osteoarthrosis. All patients had a 1.2-1.5 cm tibial tuberosity advancement with a release of the lateral patellar retinacula and no other additional surgery. Total incorporation of grafts took place in 116 cases (86.6%); graft resorption appeared in 16 patients (11.9%)(total resorption 3, and partial resorption 13). No disease transmission has been detected. To avoid donor site morbidity associated with harvesting iliac crest, the use of frozen bone allograft is a good alternative in the advancement of the tibial tuberosity. PMID- 10853777 TI - Use of bovine xenograft in reconstruction of traumatic anterior cranial fossa bone defects involving the frontal sinus. AB - Four patients underwent reconstruction of anterior cranial fossa of skull defects between November 1997 and January 1998. All of them had traumatic anterior cranial fossa defect and were reconstructed with lyophilised bovine cortical bone graft. There was no cerebrospinal fluid leakage, meningitis, extradural abscess or other infections. This study demonstrates that the use of bovine bone graft in the reconstruction of anterior base defects is safe. PMID- 10853778 TI - Application of frozen and radiation-sterilised bone allografts for treatment of bone cysts. AB - Between 1981 and 1998 frozen and radiation sterilized bone allografts were transplanted into 1376 patients at the Institute of Traumatology, Orthopaedic and Neurosurgery of the Military School of Medicine in Warsaw. Of these 179 (13%) required treatment due to benign tumours. Incidence of solitary cysts was highest (127 cases, 71%), mainly occurring in children (84%). During surgery bone cysts were excised and filled by bone grafts. Results were evaluated several years after surgery (2-10 yrs) using x-rays and clinical examination. Rebuilding of allografts was one of the most important prognostic factors. Within the period of observation time 83% of transplanted allografts were rebuilt and substituted by own patients bone. PMID- 10853779 TI - Frozen and radiation-sterilized bone allografts in the treatment of post traumatic malformation of bones. AB - Post-traumatical malformations of bone are often reconstructed with the use of preserved bone allografts. At the Institute of Traumatology, Orthopaedics and Neurosurgery 495 patients were treated with use of preserved (frozen and radiation-sterilized) bone allografts, following trauma, between 1981 and 1995. Non union of bone and osteomyelitis were main reasons for allotransplantation of bone. Remodelling of bone allografts has been observed during 2-8 years after surgery. Substitution of allografts and good result of treatment were found in 80% of all cases. In paper the analysis of results of treatment is presented. PMID- 10853780 TI - Fusion of spine in children scoliosis with frozen & radiation--sterilized bone allograft. AB - 102 children have been treated at the Institute of Traumatology, Orthopaedics & Neurosurgery a result of scoliosis. In all of these multi-step treatment has been applied. Initially a telescopic rod was implanted into the spine. Allografts were introduced around the rod hook, after 6 or 8 months, when angle of scoliosis increased, the rod was exchanged for a longer one. The final step was performed when conditions permitted and the scoliosis was markedly corrected and the fusion of the spine with the solid allograft was accomplished. The result of treatment was evaluated 1 to 7 years after surgery. Bone allografts were rebuilt within 6 month. Correction of scoliosis of 50% to 70% was achieved in all cases. PMID- 10853781 TI - Fractures of thoracic and lumbar spine; treatment and follow up. AB - The number of posttraumatic spinal fractures is constantly growing. About 15-20% of patients injury need surgery after injury, as result of spinal instability or canal stenosis often leading to neurological complications. In 1987-97 56 patients (6 female and 50 male) were treated following serious spine trauma with fractures. The age of the patients was 19-61 years. Car accidents were the most common cause of trauma affecting 30 patients (53%), falls from heights affecting 26 patients (47%). Multiple injuries were diagnosed in 24 patients (43%). More than one vertebra was fractured in 15 patients (27%). In 9 cases (16%) surgery was performed within 24 hours of the accident, in 22 (39%) within 72 hours and in 25 patients (45%) after 72 hours, in some cases even several weeks after the trauma. 20 patients (36%) required spinal decompression after the accident. The fracture in one case was located in the thoracic section, in 17 patients in Th11 L2 and in 2 patients in L3-L5 section. In 4 cases reconstruction of the meninx was performed. 48 patients were treated by stabilisation using Harrington rods; in 5 patients Kluger stabilisers were used, in the remaining 3 patients stabilization was carried out by wire loop. In all cases frozen and radiation sterilized bone allografts were applied. Results were evaluated 2-8 years after surgery using X-rays, CT-pictures and direct examination. In 25 patients (45%) results were found to be good, in 23 patients (41%) mediocre and in 8 patients (14%) results were unsatisfactory. PMID- 10853782 TI - Frozen allogenic spongy bone grafts in filling the defects caused by fractures of proximal tibia. AB - The authors present the way of using the allogenic, frozen, radio-sterilized, spongy bone grafts in operational filling of defects after infra-articular fracture of proximal tibia. Fifteen patients (11 men and 4 women) classified between 30 and 66 years old were evaluated. These patients were operated from 1996 to 1998. The operational treatment with using frozen spongy bone grafts was performed on patients with comminuted split fracture of lateral condyle and depression of tibial plateau. In each case after elevating depressed fragment of tibial plateau bone grafts and screws were used to create a stable joint surface. In the same time other injured structures of the joint (ligaments, meniscus) were being reconstructed. To evaluate the stage of grafts incorporation, radiological examinations were made in; 4 weeks, 8 weeks, 6 months, 1 year from the date of the operation. We observed the process of calcification and forming trabeculac in bone grafts. The mechanical abilities of the reconstructed joint were evaluated according to the IKDS and Lysholm score. In all cases the spongy bone grafts in the joint were reconstructed and rebuild successfully. Thirteen patients had good and very good results in further clinical examinations. We claim that frozen, allogenic grafts of the spongy bone are very useful in filing detects of that bone caused by complicated injuries. This method of treatment with its good results is recommended. PMID- 10853783 TI - The application of allogenic bone grafts for replantation of a hip prosthesis; an attempt at radiological evaluation of graft rebuilding. AB - An attempt at radiological estimation of remodelling of allogenic bone grafts used in reconstruction of bone stock in revision of hip arthroplasty. THE METHOD: x-ray pictures of the treated hips were taken immediately and at 3, 6 and 12 months after surgery. Changes of bone structure around the prosthesis were estimated. THE RESULTS: symptoms of remodelling (presence of newly formed bone trabecules between host bone and the grafts and in the surrounding area) were encountered at 3 to 6 months after reconstruction surgery. CONCLUSIONS: radiological estimation of bone remodelling remains subjective. Nevertheless it can help to determine bone graft rebuilding in revision surgery of the hip. PMID- 10853784 TI - The usage of frozen allografts of the spongy bone in filling the loss of bone after loosening of the hip prosthesis. AB - The authors present the way of using allogenic, frozen, radio-sterilizated, spongy bone grafts in operational reconstructing the defects of bone stock caused by loosening of the hip prosthesis. The bone grafts were sterelizated by radiation, formed in cubes and it had the fat removed. The operational reconstruction of the spongy bone was performed according to the method described by E. Gee, R. Ling and T.J.J.H. Sioof. Thirty five operations of the revision hip arthroplasty with using the bone grafts were made since 1995. Just before transplantation the grafts were morselised to pieces of size 3 x 3 x 3 mm and then stuck in the place of the defect. Rebuilding of the morselised grafts of the spongy bone starts from activating osteoblasts, while the same process in the compact bone starts with osteolysis by activating osteoclasts. The mechanical durability of these grafts is also very high. We claim that the biological connection between the host bone and the graft forms in a couple of weeks, but the forming of a solid, fully functional bone takes about few months. Only radiological examination shows the stage of allografts incorporation into host's skeleton. Between these 35 operations 2 patients needed to be operated again, because of the grafts resorption. In conclusion, operative revision hip arthroplasty to require the special operating tools large quantities spongy bone allografts. PMID- 10853785 TI - Replacement of the anterior cruciate ligament of the knee with deep frozen bone tendon-bone allografts. AB - Surgical treatment of the torn anterior cruciate ligament (ACL) and consequent knee instability showed great development over the last decade. Arthroscopic techniques and the use of different allogenic tissues became a routine. Between 1995 and 1998, 31 knees in 30 patients underwent ACL reconstruction of the knee with fresh-frozen allografts at the Department of Orthopedics, Medical University of Pecs, Hungary. The operations were performed with arthroscopic technique. This paper retrospectively assesses the outcomes with an average follow up of 28 months, which showed good results in most of the cases. The authors reviewed the literature emphasizing advantages and disadvantages of the method with special interest to possible complications resulting from the use of allografts: graft rejection, graft re-rupture, transmission of infection and synovitis evoked by immune response. PMID- 10853786 TI - Clinical application of amniotic membranes on a patient with epidermolysis bullosa. AB - The case of a patient with dystrophic epidermolysis bullosa treated with radiosterilised amniotic membranes is presented. The disorder is a congenital disease characterised by a poor desmosomal junction in the keratinocyte membrane. After proper donor screening, amnios were collected at Hospital Central Sur de Alta Especialidad (HCSAE), PEMEX and microbiological analysis was performed at Universidad Nacional Autonoma de Mexico, FQUNAM, (Biology Dept. of the Chemistry Faculty, National Autonomous University of Mexico), before and after radiation sterilisation. Processing, packaging and sterilisation were performed at Instituto Nacional de Investigaciones Nucleares, ININ, (National Nuclear Research Institute). The patient, a ten-year-old boy with severe malnutrition, extensive loss of skin and pseudomonad infection in the whole body, was treated with gentle debridement in a Hubbard bath. Later amnion application was performed with sterilised amnios by using two different processes, in one of which the amnion was sterilised with paracetic acid, preserved in glycerol, kindly donated by the German Institute for Tissue and Cell Replacement and applied by Dr. Johannes C. Bruck, IAEA visiting expert, and the other amnion was processed at ININ: air dried and sterilised by gamma radiation at dose of 30 kGy. After spontaneous epithelisation was successfully promoted for seven days, the pain was alleviated and mobility was improved in a few hours and the patient's general condition was so improved that in a month he was discharged. Unfortunately, because this disease is revertive and has malignant degeneration, the prognosis is not good. PMID- 10853788 TI - The clinical application of a fetal membrane on a diabetic patient's injury. AB - A description of the successful use of a fetal membrane in the recovery of a injured patient is presented. A diabetic female patient, 63 years old, was accidentally injured with a winnowing fork, which opened the skin of the front side of her left foot. For first two weeks, she was submitted to a treatment involving antibiotics, analgesics and antiinflammatories, but the infection did not disappear and the pain was almost unbearable. At the end of the third week, the injury underwent surgical washing with epidural blockage; suitable insulin dose and oral antidiabetics controlled the high-glucose concentration. However, due to the intensity of the pain, a risky epidural blockage was administered every four hours for 12 days. Finally, a radiosterilised human fetal membrane, which was collected at HCSAE and processed at ININ, was used as a biological wound dressing. Biological control of the tissue was performed at Facultad de Quimica de la Universidad Nacional Autonoma de Mexico (The Chemistry Faculty of the National Autonomous University of Mexico) (UNAM). As a result, the pain first diminished and then stopped. The healing process started in a few days and led to a complete recovery in 2 1/2 months. At present, the patient is in good condition, living a normal life. PMID- 10853787 TI - Application of acellular dermis and autograft on burns and scars. AB - The cases of two patients with burns treated with dermis allograft and of one patient for lip reconstructive aesthetic filling treated with less than one mm3 of radiosterilised acellular dermis are presented. This paper emphasizes the treatment with radiosterilised dermal grafts with a permanent character so far. Hospitals, therefore, can satisfy the demand for this kind of tissue in the case of disaster and patients with serious injuries. In the cases cited, histocompatibility analysis was not required, thus having the advantage of long time storage of the radiosterilised dermis used on these patients. Neither inflammatory reaction nor acute phase re-absorption were observed. Moreover, shrink (contract) healing was diminished. After two years, the results are still satisfactory. PMID- 10853789 TI - Application of the amniotic membrane in ocular surface pathology. AB - The amnion is a fine semi-transparent membrane that has been used in clinical practice to encourage epithelization in burns, in skin ulcers, or as a skin graft. Application in ocular surface disorders first took place in 1940. We carried out the membrane amniotic implantation on 11 patients with different pathologies: three cases of limbal stem cell deficiency (caustication with failure of prior keratoplasty, congenital aniridia and post-radiotherapy keratopathy), one case with persistent neurotrophic corneal ulcer after prior keratoplasty, four cases with epithelial defect of long evolution, one case of extensive Salzmann's degeneration of the cornea, and two cases after the resection of conjunctival tumour. The follow-up period varied between 2 and 6.5 months (mean = 4 months). Amniotic membrane was obtained by elective Caesarean, and it was preserved at -80 degrees C. In all transplanted patients the reabsorption of the amniotic membrane took place between the third and the fifth week. In the cases of resection of conjunctival tumour the epithelialization was completed between the first and the second post-operative week, with minimal residual scarring. In the other cases, with affliction of the corneal epithelium, the complete epithelialization, together with a marked reduction in the inflammatory response, occurred in all except 2 cases. In conclusion, the implantation of preserved human amniotic membrane can favour the recovery of a normal ocular surface in different pathologies, both in corneal and conjunctival lesions. PMID- 10853790 TI - Amnion allografts prepared in the Central Tissue Bank in Warsaw. AB - Applications of allogenic amnion grafts range from wound dressing of severe burns, dermabrasions and lower extremity ulcer treatments to plastic surgery, laryngology and ophthalmology. The aim of the present study was to elaborate the method of processing, preservation and sterilization of human amnion allografts prepared as wound dressing used mainly for burned patients. During the amniotic sac processing (after separation of chorion) special attention was paid to ensure that the epithelial side of amnion is placed directly on polyester net used as a support. After application on the wound, the epithelial side with the basement membrane is facing outwards; this will promote migration, attachment and spreading of the host cells encouraging epithelialization. Human amnion allografts were preserved by lyophilization or deep-freezing and subsequently radiation-sterilized with a dose of 35 kGy. It has been observed, however, that lyophilized irradiated allografts are resorbed within a few days, while frozen irradiated ones better adhere to wound and persist even 3 weeks after grafting, therefore, it has been decided to preserve amnion by deep-freezing. Since the beginning of 1998 over 400 preserved radiation-sterilized amnion allografts (with a total surface area over 40,000 cm2) have been prepared at the Central Tissue Bank in Warsaw and distributed to clinics and hospitals throughout the country. PMID- 10853791 TI - Inflammatory reactions associated with a calcium sulfate bone substitute. AB - OBJECTIVES: Loss of bone substance is a major source of disability that often requires grafting. Recently developed synthetic bone grafts have generated a lot of enthusiasm due to the lack of immunological reactions and infectious disease transmission risk. The current work describes some peculiar complications related to the use of calcium sulfate granules. METHODS: 15 implantations of calcium sulfate pellets Osteoset (Wright Medical Technology) were performed following resection of bone tumors at our service during 1999. Clinical or computerized tomography scans were available in all patients. RESULTS: 3 cases were encountered in which a severe inflammatory reaction developed. In one case serous drainage and an allergic reaction obligated graft removal. In another case, inflammation resolved two months following implantation. In the last case, wound breakdown occurred. CONCLUSIONS: A sterile inflammatory response has previously hindered the use of absorbable poly-lactic and poly-glycolic acid rods. Apparently due to rapid graft resorption, the resulting calcium-rich fluid incites inflammation. The single case of an allergic reaction is interesting. An allergy to plaster of Paris is rare and related to minor additives. These were not present in the bone substitute used. Inflammatory complications should be considered when assessing the risk-benefit ratio of using different types of bone replacement materials, and comparing allogeneic grafts to synthetic ones. PMID- 10853792 TI - Human osteoblast in contact with various biomaterials in vitro. AB - OBJECTIVES: The aim of the study was to investigate changes in osteoblast number and viability in contact with biomaterials and to compare both tests. METHODS: Human primary culture osteoblasts were seeded on the polished surfaces of hydroxyapatite, alumina, titanium and surgical steel. After 4, 9, 14, 24 and 48h cultures were subjected to XTT viability assay. Subsequently Hoechst staining of nuclei was performed. Number of cells on each sample was counted. RESULTS: There were no differences in cell viability measured by means of XTT assay between osteoblasts cultured on hydroxyapatite and alumina during 48 hours of experiment. However on titanium as well as on surgical steel cell viability was significantly lower than on bioceramics. The lowest viability was noticed on surgical steel. Cell number on titanium was significantly higher than on steel. There were no differences between cell numbers on hydroxyapatite and alumina as well as between investigated bioceramics and metals. Nucleus number and the results of viability assay were compared. There was no correlation found between number and viability of cells. CONCLUSION: the results of a single test may not provide sufficient information on the interaction between cells and implant. Application of a battery of tests is necessary in material biocompatibility investigation in vitro. PMID- 10853793 TI - Carbon fiber scaffolds in the surgical treatment of cartilage lesions. AB - We present a new method of treatment of chondral defects of the knee with the usage of carbon fibers. The prospective study describes experience with carbon fibers used as scaffolds in the drilled lesions to enhance ingrowth of regenerative tissue. We treated 35 patients for chondral defects with that method from December 93 to June 97. Average age was 46 years (range, 19-68 years) and average follow-up was 48 months (range, 24-55 months). The results were assessed by HSS knee scale, the Wallgren-Tegner activity score, VAS (visual analogue scale to measure pain) and patients assessment of the surgery. Twenty-five (71%) of 35 patients were rated good or excellent. The most striking result was good pain relief. Early follow-up results are good, but these must be confirmed in long term observations. PMID- 10853794 TI - In vitro proliferation, differentiation and immuno-magnetic bead purification of human myoblasts. AB - OBJECTIVES: In vitro culture of myoblasts and subsequent grafting into injured myocardium represents a new therapeutic approach for the treatment of myocardial infarct. A major limitation to developing enough myoblasts to engrafting purpose is the isolation and purification. In the present work we purified myoblast from primary culture using an immunomagnetic bead technique. METHODS: Primary culture was obtained by trypsin-EDTA digestion of human muscle biopsies. Cells were cultured in DMEM growth medium containing 10% FBS, 2 mM L-glutamine and antibiotics. Immunotechniques using both monoclonal anti-myosin heavy chain (skeletal fast) and 5.1.H11 antibody combining with flow cytometry did identification of myoblasts. Positive selection was on myoblasts bound to 5.1.H11 incubating with human antimouse IgG coated magnetic beads (Dynabead) and subsequent isolation by magnet, releasing cells from beads with DNAse. RESULTS: More than 59% of primary cell culture are positive to 5.1.H11 and decreasing with passage. The coating of culture dish surface increased specific growth rate of myoblast clones twice. Positive selection allows to increasing concentration of myoblasts from 8.4% in mixed culture to more than 90% without affecting neither viability nor platting efficiency. CONCLUSION: Purification procedure reported here is easy, efficient and requires small amount of sample, which will facilitate the purpose of autologous implant. PMID- 10853795 TI - Generalized monotonicity, integral conditions and partial survival. AB - An interesting paper of Vance and Coddington concerning the scalar equation x = xf(t, x) is improved by a drastic weakening of the assumptions. Surprising as it may seem, this weakening simplifies the analysis and leads to stronger conclusions. An extension to scalar inequalities of similar structure is applied to a large class of vector equations describing the evolution of n interacting species. Thus we obtain new criteria for extinction and for partial survival, in the sense that each of the separate species may come arbitrarily close to extinction but the population as a whole does not. PMID- 10853796 TI - Analytical solutions of a minimal model of species migration in a bounded domain. AB - A minimal model of species migration is presented which takes the form of a parabolic equation with boundary conditions and initial data. Solutions to the differential problem are obtained that can be used to describe the small- and large-time evolution of a species distribution within a bounded domain. These expressions are compared with the results of numerical simulations and are found to be satisfactory within appropriate temporal regimes. The solutions presented can be used to describe existing observations of nematode distributions, can be used as the basis for further work on nematode migration, and may also be interpreted more generally. PMID- 10853797 TI - Stochastic host-parasite interaction models. AB - We contribute to the discussion of causes and effects of aggregation (overdispersion) of macroparasite counts, focussing particularly upon the effects of clumped infections and parasite-induced host mortality. The simple nonlinear stochastic model for the evolution of the parasite load of a single host, investigated in Isham (1995), is extended to allow three parasite stages (larval, mature and offspring), and to allow durations of these stages to be non exponentially distributed. As in the earlier work, exact algebraic results are possible, providing insight into the aggregation mechanisms, as long as the only source of interaction between host and parasites is an excess host mortality linearly related to the parasite load. Results are obtained on the distribution of parasite load and on host survival. In particular, although parasite-induced host mortality is usually thought of as a process that reduces parasite aggregation (Anderson and Gordon 1982), it is shown that, for this model, parasite-induced host mortality cannot cause the index of dispersion to fall below unity. Host heterogeneity and disease control are also discussed. An approximation based on moment assumptions appropriate to a specially-constructed multivariate negative binomial distribution is proposed. This approximation, which is applicable to other processes, and an alternative based on the multivariate normal distribution are compared with exact results. PMID- 10853798 TI - An analysis of the hydraulic conductivity of the extracisternal space of the cochlear outer hair cell. AB - The cylindrically shaped cochlear outer hair cell (OHC) plays an important role in the transduction of acoustic energy into electrical energy in the cochlea. The extracisternal space (ECiS) of the lateral wall of the OHC is the fluid-filled space between the plasma membrane (PM) and the intracellular subsurface cisterna (SSC). In the ECiS, an array of cylindrical micropillars extends from the SSC to the PM. We obtain equations for the pressure, osmotic concentration and fluid velocity in the ECiS from the Brinkman-Stokes equations for steady incompressible flow in a plane channel that encloses an array of cylinders and whose upper wall, i.e. the plasma membrane, has a hydraulic conductivity of P(PM). From these equations we obtain an estimate for the hydraulic conductivity of the ECiS, P(ECiS). We show that the ECiS geometry accounts for P(ECiS) being several orders of magnitude larger than P(PM) and that P(ECiS) increases with the width of the ECiS and decreases with the length of the ECiS. PMID- 10853799 TI - The human sodium-iodine symporter (NIS) as a key for specific thymic iodine-131 uptake. PMID- 10853800 TI - Evaluation of [O-methyl-11C]RS-15385-197 as a positron emission tomography radioligand for central alpha2-adrenoceptors. AB - Carbon-11 labelled RS-15385-197 and its ethylsulphonyl analogue, RS-79948-197, were evaluated in rats as potential radioligands to image central alpha2 adrenoceptors in vivo. The biodistributions of both compounds were comparable with that obtained in an earlier study using tritiated RS-79948-197 and were consistent with the known localisation of alpha2-adrenoceptors. The maximal signals (total to non-specific binding) were, however, reduced, in the order [11C]RS-79948-197 < [11C]RS-15385-197 < [3H]RS-79948-197, primarily due to the difference in radiolabel position (O-methyl for carbon- 11 compared with S-ethyl for tritium). This resulted in the in-growth of radiolabelled metabolites in plasma, which, in turn, contributed to the non-specific component of brain radioactivity. Nonetheless, the signal ratio of approximately 5 for a receptor dense tissue compared with the receptor-sparse cerebellum, at 90-120 min after radioligand injection, encouraged the development of [O-methyl-11C]RS-15385-197 for human positron emission tomography (PET). Unfortunately, in two human PET scans (each of 90 min), brain extraction of the radioligand was minimal, with volumes of distribution more than an order of magnitude lower than that measured in rats. Following intravenous injection, radioactivity was retained in plasma and metabolism of the radiolabelled compound was very low. Retrospective measurements of in vitro plasma protein binding and in vivo brain uptake index (BUI) in rats demonstrated a higher protein binding of the radioligand in human compared with rat plasma and a lower BUI in the presence of human plasma. It is feasible that a higher affinity of RS-15385-197 for human plasma protein compared with receptor limited the transport of the radioligand. Although one of the PET scans showed a slight heterogeneity in biodistribution of radioactivity which was consistent with the known localisation of alpha2-adrenoceptors in human brain, it was concluded that [O-methyl-11C]RS-15385-197 showed little promise for routine quantification of alpha2-adrenoceptors in man. PMID- 10853801 TI - Iterative reconstruction: an improvement of technetium-99m MIBI SPET for the detection of parathyroid adenomas? AB - The purpose of this study was to assess the value of technetium-99m methoxyisobutylisonitrile (MIBI) single-photon emission tomography (SPET) and an iterative reconstruction algorithm for the preoperative localisation of parathyroid adenomas (PTAs). Seventy-two patients (26 male, 46 female, mean age 58+/-16 years) with known primary hyperparathyroidism were examined preoperatively. First, a thyroid examination was performed to detect possible MIBI-accumulating thyroid lesions. Planar scans were then acquired 15 and 120 min and tomographic images 120 min after intravenous injection of 740 MBq 99mTc-MIBI, using a triple-head gamma camera (Picker Prism 3000). Additionally, 99mTc-MIBI/ 99mTc-pertechnetate subtraction scintigraphy of the early planar images was performed. The SPET data were evaluated using an iterative reconstruction (multiplicative iterative SPET reconstruction: MISR) as well as a standard algorithm (FBP: filtered back-projection with application of a 3-D low-pass postfilter). The weight of the resected PTAs ranged from 110 mg to 5 g. Using planar MIBI scans, correct localisation of the side of the PTA was possible in 81% of cases (58% for PTAs weighing less than 500 mg). Sensitivity increased to 94% using SPET and FBP, while with MISR it rose further, to 97%. Patients with PTAs weighing less than 500 mg showed a sensitivity of 88% with MISR and 81% with FBP. Furthermore, there was a clear improvement in image quality using MISR. None of the normal parathyroid glands were visualised. This study indicates that, in comparison with planar scintigraphy, 99mTc-MIBI SPET is a more sensitive and specific tool for topographical localisation of PTAs, especially those that are small. There is a further improvement in sensitivity and image quality when iterative reconstruction is used instead of FBP. PMID- 10853802 TI - The value of fluorine-18 fluorodeoxyglucose PET in patients with medullary thyroid cancer. AB - The early detection of metastases from medullary thyroid cancer (MTC) is important because the only curative therapy consists in surgical removal of all tumour tissue. There is no single sensitive diagnostic imaging modality for the localization of all metastases in patients with MTC. Therefore, in many cases several imaging modalities (e.g. ultrasonography, magnetic resonance imaging, computerized tomography and scintigraphy using pentavalent technetium-99m dimercaptosuccinic acid, thallium-201 chloride, indium-111 pentetreotide, anti CEA antibodies or metaiodobenzylguanidine) must be performed consecutively in patients with elevated calcitonin levels until the tumour is localized. In this prospective study, we investigated the value of fluorine-18 fluorodeoxyglucose positron emission tomography ([18F]FDG PET) in the follow-up of patients with MTC. [18F]FDG PET examinations of the neck and the chest were performed in 20 patients with elevated calcitonin levels or sonographic abnormalities in the neck. Positive [18F]FDG findings were validated by histology, computerized tomography or selective venous catheterization. [18F]FDG PET detected tumour in 13/17 patients (nine cases were validated by histology, four by computerized tomography). Five patients showed completely negative PET scans (of these cases, one was true-negative and four false-negative). One patient with [18F]FDG accumulation in pulmonary lesions from silicosis and one patient with a neck lesion that was not subjected to histological validation had to be excluded. Considering all validated localizations, [18F]FDG PET detected 12/14 tumour manifestations in the neck, 6/7 mediastinal metastases, 2/2 pulmonary metastases and 2/2 bone metastases. In two patients with elevated calcitonin levels, no diagnostic modality was able to localize a tumour. The sensitivity of [18F]FDG PET in the follow-up of MTC was 76% (95% confidence interval 53%-94%); this is encouraging. [18F]FDG PET promises to be a valuable diagnostic method, especially for the detection of lymph node metastases, surgical resection of which can result in complete remission. PMID- 10853803 TI - The role of bone scanning in severe frostbite of the extremities: a retrospective study of 88 cases. AB - We performed a retrospective study of the results of two-phase technetium-99m hydroxymethylene diphosphonate bone scans in 88 patients with severe frostbite of the extremities. All patients were evaluated within 48 h after rewarming and all underwent a first bone scan (BS1) within 5 days after rewarming (median, day 2) (group 1). An excellent correlation was found between absence of tracer uptake in the phalanges and later amputation; this correlation was especially strong during the bone phase of the scans (specificity = 0.99, sensitivity = 0.96, positive predictive value = 0.92). Normal or high uptake in the phalanges was a reliable indicator of ultimate healing (negative predictive value = 0.99). The sensitivity of the examination was enhanced by performing a second scan (BS2) more than 5 days (median, day 8) after rewarming (group 2, n = 36). A comparative analysis of BS1 and BS2 demonstrated that some of the lesions continued to evolve between day 2 and day 8. This suggests that the lesions could still be modified during this period. On the basis of the findings it is recommended that bone scan be performed close to day 2 in all patients who present with lesions extending proximally to include the entirety of one or more phalanges. In the case of severe sepsis, the results of the first bone scan, BS1, can serve as an indication for emergency amputation. BS2 should be performed close to day 8 only if there is an area of low or absent uptake on BS1. It is concluded that scintigraphy is an excellent means of evaluating patients with severe frostbite of the extremities: as early as day 2 after the injury it can indicate whether amputation is necessary, and between days 2 and 8 it provides valuable information on the efficacy of treatment. PMID- 10853804 TI - Testicular function after radioiodine therapy for thyroid carcinoma. AB - Radiotherapy can cause infertility in both men and women. However, few data are available concerning the effects of radioiodine therapy for thyroid carcinoma on testicular function. We investigated 25 men (age 23-73 years) with differentiated thyroid carcinoma in a longitudinal prospective trial. Follicle-stimulating hormone (FSH), inhibin B, luteinising hormone (LH) and testosterone were measured before (n = 25) and 3 months (n = 11), 6 months (n = 18), 12 months (n = 22), and 18 months (n = 18) after radioiodine therapy [radioiodine dose (mean +/- SEM): 9.8+/-0.89 GBq]. Before therapy, FSH was 5.4+/-0.77 IU/l; it increased significantly (P<0.001) to 21.3+/-2.4 IU/l after 6 months and fell to 7.4+/-1.3 IU/l after 18 months (normal range: 1.8-9.2 IU/l). Inhibin B was significantly decreased (P<0.001) from 178+/-25.3 pg/ml before therapy to 22.2+/-5.5 pg/ml after 3 and 29.4+/-5.7 pg/ml after 6 months and rose to 154+/-23.3 pg/ml after 18 months (normal range 75-350 pg/ml). LH and testosterone were within the normal range during the whole study (1.6-9.2 IU/l and 10.4-34.7 nmol/l, respectively). LH was significantly increased (P<0.001) from 2.8+/-0.33 IU/l before therapy to 5.9+/-0.69 IU/l 6 months after therapy and then fell slowly to 4.0+/-0.45 IU/l after 18 months. Total testosterone was significantly increased (P<0.01) from 12.8+/-0.99 nmol/l at baseline to 19.8+/-1.7 nmol/l after 12 months and 19.6+/ 1.7 nmol/l after 18 months. The testosterone/LH ratio (normal range: 3.3-17.9 nmol/IU) fell from 5.8+/-0.66 nmol/IU to 3.0+/-0.36 nmol/IU after 3 months (P<0.01); it remained close to the latter value after 6 months (3.4+/-0.49 nmol/IU) and then rose to 5.5+/-0.6 nmol/IU after 18 months. In conclusion, 3 and 6 months after radioiodine therapy all patients showed elevated FSH and decreased inhibin B levels, reflecting severely impaired spermatogenesis. At the same time a compensated insufficiency of the Leydig cell function was observed. Eighteen months after the last radioiodine therapy, mean values of gonadal function had completely recovered. PMID- 10853805 TI - The effect of collateral flow and myocardial viability on the distribution of technetium-99m sestamibi in a closed-chest model of coronary occlusion and reperfusion. AB - Myocardial uptake of technetium-99m sestamibi at low coronary flow rates overestimates blood flow, but the relative impact of flow and viability on 99mTc sestamibi kinetics is unclear. The objective of this study was to determine the effect of myocardial viability and the degree of collateral blood flow on the uptake and retention of 99mTc-sestamibi by examining three animal models of coronary occlusion and reperfusion, each reflecting a different state of viability and collateral blood flow. Three closed-chest animal models were studied: canine (high collateral flow, preserved viability), porcine (low collateral flow, absent viability) and porcine with slowly occlusive coronary stents producing infarction and enhanced collateral blood flow (high collateral flow, absent viability). There were seven dogs, seven pigs and six pigs, respectively, in each animal model. Animals from all three models were subjected to a 40-min total left anterior descending artery (LAD) occlusion followed by 2 h of reperfusion. 99mTc-sestamibi and radiolabelled microspheres were injected during LAD occlusion 10 min prior to reperfusion. Animals were sacrificed after 2 h of reperfusion flow. Ex situ heart slice imaging to determine risk area was followed by viability staining to determine infarct size. Slices were subsequently sectioned into equally sized radial segments and placed in a gamma well counter. Risk area as determined by ex situ 99mTc-sestamibi imaging was not significantly different by model. Pathological infarct size differed significantly by model [canine = 1%+/-1% of the left ventricle (LV); porcine = 13%+/-8% LV; porcine with stent = 14%+/-7% LV; P = 0.002)]. Collateral blood flow by microspheres during occlusion tended to differ among models (overall P = 0.08), with the canine and porcine with stent models having relatively high flow rates compared with the acute porcine model. 99mTc-sestamibi activity correlated with microsphere blood flow in all three models, with r values for individual animals (n = 20) ranging from 0.86 to 0.96 (all P<0.0001). There was a significant difference in the regression line intercepts (P<0.0001) and slopes (P<0.01) among the three models comparing 99mTc-sestamibi uptake with myocardial blood flow. 99mTc-sestamibi uptake overestimated blood flow to a greater extent in the canine model (high flow with viability) than in the porcine model (low flow, absent viability). Despite enhanced collateral flow, there was significantly less overestimation of flow in the porcine stent model (high flow, absent viability). In conclusion, at low flow rates 99mTc-sestamibi activity overestimates myocardial blood flow. This effect is most pronounced in myocardium with significant collateral flow and preserved viability, consistent with over extraction or redistribution of the tracer. The effect is markedly decreased in non-viable myocardium regardless of blood flow. PMID- 10853806 TI - Use of ECG-gated SPET to assess the evolution of perfusion after acute myocardial infarction. AB - Serial improvement in myocardial perfusion images from the acute or subacute to the chronic stage of acute myocardial infarction (AMI) has been attributed to improved coronary microcirculation or cell function after acute ischaemia and reperfusion. However, conventionally used non-gated imaging cannot eliminate the effect of improved regional contraction. We studied the possibility that such scintigraphic improvement reflects the functional recovery by using ECG-gated myocardial perfusion imaging with technetium-99m sestamibi. Nineteen AMI patients who received acute reperfusion therapy underwent ECG-gated myocardial single photon emission tomography (SPET) in the subacute and chronic stages. Serial changes in regional image count distributions were analysed on the non-gated, end diastolic (ED) and end-systolic (ES) images by using segmental mean percent peak activity (MPA) and AMPA (MPA in chronic stage - MPA in subacute stage) on bull's eye polar maps. These changes were compared with those in regional wall motion on biplane left ventriculography (LVG) from the acute (just after reperfusion) to the chronic stage. During the follow-up, regional wall motion remained the same in 42 (group A) but improved in 17 (group B) of the 59 ischaemically compromised segments. MPA showed no improvement in group A but significant improvement in group B on the non-gated and ES images (P<0.0001 and P<0.001, respectively). However, MPA on the ED images showed no improvement in either group. In the follow-up study of AMI, the scintigraphic improvement documented on the non-gated myocardial images appears to be mainly related to the recovery of wall thickening and not to a real improvement in myocardial perfusion. Therefore, ECG-gated myocardial imaging, which enables simultaneous assessment of changes in perfusion and contraction, is preferable to conventional non-gated imaging for follow-up of AMI. PMID- 10853807 TI - Fluorine-18 fluorodeoxyglucose PET in infectious bone diseases: results of histologically confirmed cases. AB - The aim of this study was to evaluate the clinical use of fluorine-18 fluorodeoxyglucose positron emission tomography (FDG-PET) in acute and chronic osteomyelitis and inflammatory spondylitis. The study population comprised 21 patients suspected of having acute or chronic osteomyelitis or inflammatory spondylitis. Fifteen of these patients subsequently underwent surgery. FDG-PET results were correlated with histopathological findings. The remaining six patients, who underwent conservative therapy, were excluded from any further evaluation due to the lack of histopathological data. The histopathological findings revealed osteomyelitis or inflammatory spondylitis in all 15 patients: seven patients had acute osteomyelitis and eight patients had chronic osteomyelitis or inflammatory spondylitis. FDG-PET yielded 15 true-positive results. The tracer uptake correlated with the histopathological findings in each case. Bone scintigraphy performed in 11 patients yielded ten true-positive results and one false-negative result. Follow-up carried out on two patients revealed normal or clearly reduced tracer uptake, which correlated with a normalisation of clinical data. In early postoperative follow-up it was impossible to differentiate between postsurgical reactive changes and further infection using FDG-PET. It is concluded that acute and chronic osteomyelitis of the peripheral as well as the central skeleton can be detected using FDG-PET. Osteomyelitis can be differentiated from soft tissue infection surrounding the bone. Unlike computed tomography and magnetic resonance imaging, FDG-PET is not affected by metal implants used for fixing fractures. FDG-PET demonstrated promising initial results with respect to treatment monitoring. Nevertheless, in the early postoperative phase FDG-PET seems to be of limited value owing to unspecific tracer uptake. PMID- 10853808 TI - Change in regional pulmonary perfusion as a result of posture and lung volume assessed using technetium-99m macroaggregated albumin SPET. AB - The purpose of this study was to evaluate the effects of gravity and lung volume on regional pulmonary perfusion using technetium-99m macroaggregated albumin (99mTc MAA) single-photon emission tomography (SPET). Twenty-five subjects were classified into three groups according to their position during the injection of the tracer [11 subjects sitting, six supine and eight both supine and prone (S+P) positions]. All of these subjects were injected with the tracer during normal tidal breathing. In the S+P group, half of the tracer was injected while the subject was in each position. The remaining 11 subjects were classified into two groups according to their lung volume during the injection. Supine patients were instructed to hold their breath at residual volume (RV) (five subjects) or total lung capacity (TLC) (six subjects) while receiving the tracer injection. A region of interest with a ventrodorsal axis was defined in the centre of each lung. Profile curves were produced by plotting and normalizing the perfusion values as a percentage of the maximum value. The perfusion distributions for the sitting and S+P positions and at RV were relatively uniform. However, the distributions for the supine position and at TLC showed a gravitational influence [sitting vs. TLC: 87.8%+/-10.4% vs. 67.3%+/-8.7% for % maximum perfusion at +5 pixels from the midpoint of the upper lobe (P<0.00002)]. The gravity-related perfusion inhomogeneity was more prominent in the lower lobe than in the upper lobe. It is concluded that the physiological vertical gravitational gradient should be taken into consideration during the interpretation of lung SPET images. Preferably, patients should be injected with the tracer twice, once in the supine position and once in the prone position, while breathing normally. Alternatively, they may be injected with the tracer once while in the supine position and holding their breath at RV. Either of these protocols should ensure a uniform distribution of tracer. PMID- 10853809 TI - Evaluation of radiotherapeutic response in non-small cell lung cancer patients by technetium-99m MIBT and thallium-201 chloride SPET. AB - The purpose of this study was to investigate the relationship between technetium 99m hexakis-2-methoxyisobutylisonitrile (99mTc-MIBI) accumulation in tumours and response to radiotherapy in non-small cell lung cancer patients in comparison with the findings obtained using thallium-201 chloride (201Tl). Simultaneous dual single-photon emission tomography (SPET) imaging with 600 MBq 99mTc-MIBI and 111 MBq 201Tl was performed in 31 patients with biopsy- or sputum cytology-proven lung cancer. SPET images were acquired 15 min (early) and 2 h (delayed) after injection, and the early ratio, delayed ratio and retention index were measured. The tumours were classified into two groups on the basis of follow-up computed tomography (CT): responders (at least 50% reduction in tumour size) and non responders (little or no change in tumour size). The mean (+/-SD) values of early ratio, delayed ratio and retention index using 99mTc-MIBI SPET were 3.0+/-1.1, 2.7+/-1.0 and -9.5+/-12.7, respectively, in responders and 2.4+/-0.7, 2.0+/-0.5 and -18.4+/-9.0, respectively, in non-responders. The corresponding values using 201Tl chloride SPET were 3.7+/-1.0, 4.7+/-1.5 and 24.2+/-22.1 in responders and 3.3+/-1.2, 4.0+/-1.3 and 20.4+/-20.5 in nonresponders. Using 99mTc-MIBI, the delayed ratio and retention index in responders were significantly higher than those in non-responders (P<0.01 and P<0.05, respectively). The results of this study indicate that patients with higher delayed ratio and retention index values using 99mTc-MIBI SPET are likely to respond better to radiotherapy than those with lower values. 99mTc-MIBI SPET may give an indication of the short-term response to radiotherapy in patients with non-small cell lung cancer. PMID- 10853810 TI - O-(2-[18F]fluoroethyl)-L-tyrosine and L-[methyl-11C]methionine uptake in brain tumours: initial results of a comparative study. AB - O-(2-[18F]Fluoroethyl)-L-tyrosine (FET) is a recently described amino acid analogue that has shown high accumulation in animal tumours. The aim of this study was to compare the uptake of FET with that of L-[methyl-11C]methionine (MET) in patients with suspected primary or recurrent intracerebral tumours. Sixteen consecutive patients with intracerebral lesions were studied on the same day by positron emission tomography (PET) using MET and FET. Uptake of FET and MET was quantified by standardized uptake values. Tracer kinetics for normal brain and intracerebral lesions were compared. On the basis of the MET-PET studies, viable tumour tissue was found in 13 patients. All tumours showed rapid uptake of FET and were visualized with high contrast. Mean uptake of FET for normal grey matter, white matter and tumour tissue was 1.1+/-0.2, 0.8+/-0.2 and 2.7+/-0.8 SUV, respectively. In all three tissues, uptake of MET was slightly higher (1.4+/-0.2, 0.9+/-0.1 and 3.3+/-1.0 SUV; P<0.01). However, contrast between tumour and normal tissues was not significantly different between MET and FET. Uptake of FET in non-neoplastic lesions (1.0+/-0.1 SUV) was significantly lower than in tumour tissue (P = 0.007). For all lesions there was a close correlation (r = 0.98) between MET and FET uptake. In conclusion, in PET studies of human brain tumours, the uptake and image contrast of FET appear to be very similar to those of MET. The specificity of FET for tumour tissue is promising but has to be addressed in a larger series of patients with non-neoplastic lesions. PMID- 10853811 TI - Sequential scintigraphic strategy for the differentiation of brain tumours. AB - Both thallium-201 and iodine-123 alpha-methyltyrosine (123I-IMT) have been shown to be useful in the diagnostic evaluation of brain tumours. The aim of this study was to investigate the respective contributions of 201Tl and 123I-IMT single photon emission tomography (SPET) in the non-invasive evaluation of intracerebral tumours. We analysed 65 patients with the following brain tumours: 8 non neoplastic lesions, 4 meningiomas, 12 low-grade gliomas, 28 high-grade gliomas, 11 metastases and 2 high-grade lymphomas. 201Tl SPET and 123I-IMT SPET were performed [start of 201Tl SPET: 15 min p.i. (early) and 180 min p.i. (delayed); start of 123I-IMT SPET: 15 min p.i.]. The intensity of uptake was quantified as the ratio between tracer accumulation in the tumour and in the contralateral hemisphere. None of the non-neoplastic lesions or low-grade gliomas expressed marked 201Tl uptake. All malignant tumours except one small metastasis and all meningiomas except one small, cystic and degenerated lesion showed significant 201Tl accumulation [Tl(15')>2.0]; 123I-IMT uptake was either absent or intermediate in non-malignant lesions except in two low-grade gliomas; the highest levels were observed in high-grade gliomas followed by metastases and lymphomas (mean IMT: 2.7 vs. 2.1 vs. 1.8), with metastases showing a high variability in 123I-IMT uptake (range: 0.8-3.6). Using 201Tl to distinguish non neoplastic lesions from malignant tumours and meningiomas, 63 of 65 patients were characterised correctly. In the latter group, high-grade gliomas were correctly identified in 27 of 28 cases by their amino acid uptake. It is concluded that the combination of 201Tl and 123I-IMT surpasses the accuracy of each single test in the differentiation of space-occupying lesions of the brain. Based on these preliminary results, a sequential strategy is proposed involving an initial 201Tl SPET study and an additional 123I-IMT SPET study in the event of positive 201Tl uptake. PMID- 10853812 TI - Role of 123I-IMP SPET in the early diagnosis of borderline chronic hydrocephalus after aneurysmal subarachnoid haemorrhage. AB - Chronic hydrocephalus after aneurysmal sub-arachnoid haemorrhage (SAH) is easily diagnosed in most cases. However, the diagnosis is sometimes difficult in borderline cases, in which (a) pathognomonic clinical deterioration due to hydrocephalus is masked by the neurological deficits caused in the acute stage of SAH and (b) ventricular enlargement is not so marked on CT scan. The purpose of this study was to investigate whether or not iodine-123 labelled N-isopropyl-p iodo-amphetamine (123I-IMP) single-photon emission tomography (SPET) is of value for the early diagnosis of borderline chronic hydrocephalus after SAH. Fifteen patients who met the criteria of borderline chronic hydrocephalus were selected for the study, and underwent a shunt operation. The patients were divided into a shunt-effective group and a shunt-ineffective group according to neurological improvement after the shunt operation. 123I-IMP SPET was performed in the acute stage of SAH, within 1 week before the shunt operation, and 2 weeks after the shunt operation. Regional cerebral blood flow was estimated by the 123I-IMP autoradiographic method. Pre-shunting periventricular low-perfusion areas showed statistically significant differences between the two groups (P<0.05). In the shunt-effective group, peri-ventricular low-perfusion areas on pre-shunting SPET were significantly enlarged compared with those in the acute stage of SAH (P<0.05), and they were significantly reduced after the shunt operation (P<0.05). In the shunt-ineffective group, periventricular low-perfusion areas showed no significant changes during the course. These results suggest that periventricular low-perfusion areas enlarge in the early stage of chronic hydrocephalus after SAH, and that 123I-IMP SPET can be used for both the early diagnosis of borderline chronic hydrocephalus after SAH and the prediction of shunt effectiveness. PMID- 10853813 TI - Peripheral sympathetic dysfunction in patients with Parkinson's disease without autonomic failure is heart selective and disease specific. taki@med.kanazawa u.ac.jp. AB - The study was undertaken to investigate by means of iodine-123-labelled metaiodobenzylguanidine (MIBG) scintigraphy the peripheral sympathetic function in patients with Parkinson's disease (PD) without autonomic failure and in patients with related neurodegenerative diseases with parkinsonism. Seventy patients (33 men and 37 women, mean age 63+/-9.7 years) with parkinsonism and ten control subjects underwent MIBG scintigraphy. Of these 70 patients, 41 were diagnosed as having idiopathic PD, 9 multiple system atrophy (MSA), 6 progressive supranuclear palsy (PSP) and 2 corticobasal degeneration (CBD); the remaining 12 were diagnosed as having neurodegenerative disease with parkinsonism (P-nism) that did not meet the diagnostic criteria of any specific disease. Cardiac planar and tomographic imaging studies and subsequent whole-body imaging were performed 20 min and 3 h after the injection of 111 MBq MIBG. The early MIBG heart to mediastinum (H/M) ratio in PD (1.61+/-0.29) was significantly lower than that in the control group (2.24+/-0.14, P<0.01), P-nism (2.15+/-0.31, P<0.01), MSA (2.08+/-0.31, P<0.05) and PSP (2.30+/-0.24, P<0.01). The delayed H/M ratio in PD (1.47+/-0.34) was also significantly lower than that in the control group (2.37+/ 0.14, P<0.01), P-nism (2.13+/-0.38, P<0.01), PSP (2.36+/-0.36, P<0.01) and MSA (2.17+/-0.36, P<0.01). In patients with PD, early and delayed H/M ratios were significantly decreased in disease stages I, II and III (established using the Hoehn and Yahr criteria) as compared with control subjects, and there were no significant differences among the stages. Only PD showed a significantly higher washout rate (WR) than that in the control subjects (27%+/-8.0% vs. 11%+/-4.2%, P<0.01). Early and delayed uptake ratios of the lung, parotid gland, thyroid gland, liver and femoral muscles in each of the patient groups were not significantly different from those in control subjects. Only the early and delayed uptake ratios of the lower leg muscles in MSA were significantly lower than those in the control group (P<0.05). IN CONCLUSION: In patients with PD without autonomic failure, only cardiac MIBG uptake was severely reduced in the earliest phase of the disease (stage I). Parkinsonian syndromes other than PD did not demonstrate significant reduction in MIBG uptake in any organs except for the lower legs in MSA. In patients with PD without autonomic failure, reduction in MIBG uptake occurs selectively in the heart; this is considered to be a specific finding for PD and useful for the differential diagnosis of the parkinsonian syndromes. PMID- 10853814 TI - Alteration of myocardial metaiodobenzylguanidine uptake after treatment of phaeochromocytoma and neuroblastoma. AB - The relationships between changes in myocardial uptake of metaiodobenzylguanidine (MIBG) and those in circulating catecholamines and cardiac function after treatment of phaeochromocytoma and neuroblastoma were evaluated. Iodine-123 or iodine-131 MIBG scintigraphy was performed before and after surgical resection and/or chemotherapy for primary tumours in nine patients with phaeochromocytoma and 13 patients with neuroblastoma. Changes in myocardial MIBG uptake after treatment were estimated by the heart-to-upper mediastinum (H/M) uptake ratios on the images obtained 24 h after MIBG injection, which were compared with serum levels of noradrenaline (NA) and adrenaline (A). Cardiac function was assessed by echocardiography, with measurements of the left ventricular ejection fraction (LVEF). Before treatment, eight patients with phaeochromocytoma and three with neuroblastoma showed poor myocardial MIBG uptake, with highly elevated circulating NA and A. Echocardiography, however, did not show cardiac dysfunction in these patients with the exception of two patients with phaeochromocytoma. With normalization of NA and A levels after treatment, all of these patients except for the two with persistent cardiac dysfunction showed restoration of myocardial MIBG uptake. The H/M ratios increased significantly after treatment in both patient groups, i.e. with phaeochromocytoma and with neuroblastoma (P<0.0001 and P<0.05, respectively), and these ratios correlated inversely with circulating NA and A before and after treatment. By contrast, there was no significant correlation between H/M ratios and LVEF in these two groups. These results indicate that suppression of myocardial MIBG uptake usually may not be related to cardiac dysfunction and may be reversible following normalization of excess catecholamine levels after treatment in patients with neuroadrenergic tumours. However, the suppression may persist in the presence of catecholamine-induced cardiac dysfunction. The assessment of myocardial MIBG uptake can be a helpful adjunct in monitoring the normalization of circulating catecholamine levels and also in identifying the presence of cardiac dysfunction in treated patients with neuroadrenergic tumours. PMID- 10853815 TI - Radiation dose rates from patients undergoing PET: implications for technologists and waiting areas. AB - Increasingly hospitals are showing an interest in developing their imaging services to include positron emission tomography (PET). There is therefore a need to be aware of the radiation doses to critical groups. To assess the effective whole-body dose received by technologists within our dedicated PET centre, each staff member was issued with a dose rate meter, and was instructed to record the time spent in contact with any radioactive source, the dose received per working day and the daily injected activity. On average each technologist administered 831 MBq per day. The mean whole-body dose per MBq injected was 0.02 microSv/MBq( 1). The average time of close contact (<2.0 m) with a radioactive source per day was 32 min. The average effective dose per minute close contact was 0.5 microSv/min(-1), which resulted in a mean daily effective dose of 14.4 microSv. No technologist received greater than 60 microSv (the current UK limit for non classified workers) in any one day, and in general doses received were less than 24 microSv, the daily dose corresponding to the proposed new annual limit for non classified workers of 6.0 mSv per annum. However, we recognise that the layout of nuclear medicine departments will not mirror our own. We therefore measured the instantaneous dose rates at 0.1, 0.5, 1.0 and 2.0 m from the mid-thorax on 115 patients immediately after injection, to provide estimates of the likely effective doses that might be received by technologists operating dual-headed coincidence detection systems, and others coming into contact in the waiting room with patients who have been injected with fluorine-18 fluorodeoxyglucose. The mean (95th percentile) dose rates measured at the four aforementioned distances were 391.7 (549.5), 127.0 (199.8), 45.3 (70.0) and 17.1 (30.0) microSv/h(-1), respectively. A number of situations have been modelled showing that, with correct planning, FDG studies should not significantly increase the effective doses to technologists. However, one possible area of concern is that, depending on the number of patients in a waiting area at any one time, accompanying persons may approach the limits set by the new UK IRR 1999 regulations for members of the public. PMID- 10853816 TI - Fluorodeoxyglucose PET in the initial staging of germ cell tumours. AB - Testicular cancer is a rare tumour with the potential for cure at diagnosis. It is important, however, to identify those patients with metastases at presentation so as to ensure that the optimum treatment strategy is employed. Many criteria have been used to try to place patients into high- or low-risk groups, with variable success. Fluorine-18 fluorodeoxyglucose positron emission tomography (FDG-PET) has the potential to identify active disease and thereby influence further management. Here we report on a retrospective study of the use of FDG-PET in the detection of metastatic testicular carcinoma at diagnosis. Thirty-one patients [13 with seminoma and 18 with non-seminomatous germ cell tumours (13 teratomas, 5 mixed)] were staged by FDG-PET scanning. The imaging was performed using a Siemens ECAT 951 scanner. All results were assessed on the basis of histology or clinical follow-up. FDG-PET scan identified metastatic disease in ten and was negative in 16; there were no false-positives and five false negatives. There were six patients in whom FDG-PET was negative and computed tomography was regarded as suspicious but follow-up was inconclusive. The positive predictive value was 100%. The negative predictive value was 76% or 91%, depending on whether the aforementioned six cases were regarded as true-negatives or false-negatives. It may be concluded that FDG-PET is capable of detecting metastatic disease at diagnosis that is not identified by other imaging techniques. These preliminary results are sufficient to suggest that a large prospective study should be performed to evaluate the role of FDG-PET in primary staging of disease. PMID- 10853817 TI - Scintigraphic differentiation between two forms of primary dysautonomia early after onset of autonomic dysfunction: value of cardiac and pulmonary iodine-123 MIBG uptake. AB - Primary dysfunction of the autonomic nervous system can be observed in patients with Parkinson's disease and those with multiple system atrophy. However, the fate of the two diseases differs considerably and leads to different strategies for patient management. Differentiation of the two diseases currently requires a combination of several clinical and electrophysiological tests. First studies of myocardial innervation using iodine-123 metaiodobenzylguanidine (MIBG) indicated a possible role of scintigraphy for this purpose. An increase in the pulmonary uptake of 123I-MIBG has been reported in secondary dysautonomias. Whether sympathetic innervation of the lung is affected in primary dysautonomias is currently unknown. Therefore, cardiac and pulmonary uptake of 123I-MIBG was studied in 21 patients with Parkinson's disease, 7 patients with multiple system atrophy and 13 age- and sex-matched controls. Thoracic images were obtained in the anterior view 4 h after intravenous injection of 185 MBq 123I-MIBG, at which time the maximum neuronal uptake is reached. All patients with Parkinson's disease had significantly lower cardiac uptake of 123I-MIBG than patients with multiple system atrophy and controls. Sympathetic innervation of the lung was not affected in either disease. It is concluded that scintigraphy with 123I-MIBG appears to be a useful tool for differentiation between Parkinson's disease and multiple system atrophy early after onset of autonomic dysfunction. PMID- 10853818 TI - Role of nuclear medicine in the treatment of malignant gliomas: the locoregional radioimmunotherapy approach. AB - The high-grade malignant gliomas (anaplastic astrocytomas and glioblastoma) have a very bad prognosis since the available methods of treatment (surgery, radiotherapy and chemotherapy) are unable to control the progression of the disease for long. The use of specific monoclonal antibodies labelled with a suitable isotope (iodine-131 or yttrium-90) represents an effective approach to hamper tumour regrowth. Some authors have injected the antibodies intravenously, or have tried to increase the tumour/background ratio with the avidin/biotin system. In many cases the labelled monoclonal antibodies were injected directly into the tumoral bed after the operation. The authors' experiences concern a quite large locoregional radioimmunotherapy study which was performed by using antitenascin antibodies labelled initially with 131I and more recently with 90Y. The clinical results demonstrate the ability of this technique to control, for a long time, the growth of these tumours. The glioblastoma median survival was prolonged to 25 months (131I group) or 31 months (90Y group). The response rate (which comprises PR, CR and NED) was 47.1% (glioblastoma 131I group) or 40% ( glioblastoma 90Y group). In many cases a significant tumour shrinking effect was radiologically demonstrated. The use of 90Y proved more favourable in bulky lesions, and reduced the radioprotection problems. PMID- 10853819 TI - Correct unit notation in equations. PMID- 10853820 TI - 123I-MIBG scintigraphy of catecholamine systems: impediments to applications in clinical medicine. PMID- 10853821 TI - Thyroid carcinoma: administration of iodine-131 therapy early in the morning, please! PMID- 10853822 TI - Functional analysis of the TGFbeta receptor/Smad pathway through gene ablation in mice. AB - During recent years, our understanding of TGFbeta signalling through serine/threonine kinase receptors and Smads has increased enormously. Activation of R-Smads by receptor induced phosphorylation is followed by complex formation with co-Smads and translocation to the nucleus, where the transcription of specific genes is affected and ultimately results in changes in cell behaviour. Experimental analysis primarily of epithelial cells in culture has revealed that a number of members of the TGFbeta family are interchangeable in the effect they have on growth and differentiation. On the other hand, different ligands of the TGFbeta superfamily can result in different responses because of cell type specific expression of other components of the signalling pathway. The relative expression levels of receptors and Smads within the cell is an important determinant of TGFbeta induced responses. Functional analysis of genes in the TGFbeta superfamily signal transduction cascade in vivo in mice either lacking entire genes, or expressing dominant negative forms of particular proteins, are providing profound new insights into the signalling cascades, their interaction and their specificity (Table 3). For example, by phenotypical comparison and intercrossing different heterozygous mutants, it has become clear that nodal, until recently an orphan protein without receptor/signal complex, probably signals through the activin type II receptor, ALK-4 and Smad2 (Nomura and Li, 1998; Song et al., 1999). Many of the genes of this cascade that have been targeted in the mouse result in early embryonic lethal phenotypes, demonstrating an important function for the BMP and TGFbeta/activin-activated pathways in mesoderm formation and differentiation, but masking a possible role in later events. For example mutations in BMP2 and 4 are lethal at or soon after gastrulation so that their putative role in skeletogenesis cannot be studied in mice lacking these genes. The difference in severity of the phenotypes between ligand, receptor and Smad deficient mice suggest that other receptors and ligands may partially compensate for the loss of one protein. Chimeric analysis provides one tool for analysing later developmental functions. By rescuing the early defects it was demonstrated that TGFbeta family members have an important function in anterior development and left/right asymmetry. Temporal and spatial specific gene targeting will be a powerful tool for analysing the function of TGFbeta family members in for example, bone formation, angiogenesis and carcinogenesis. Isolation of cells from the different gene targeted mice provides a unique source of material to gain more insight in the biochemical mechanisms of specific pathways. For example, use of cells deficient in Smad2 for biochemical and cell biological assays could give a better view of the function of Smad3. Smad3 deficient mice already demonstrate that there is a clear difference between Smad2 and Smad3 during development. Full descriptions of the remaining gene ablation studies of this signal transduction cascade, namely those for ALK-5, BMPR-II and Smad1 and -7 are eagerly awaited to complete the puzzle. As more of these superfamily of ligands and their signalling pathways have been functionally dissected, it has become evident that this superfamily of growth factors plays a pivotal role in epiblast formation and gastrulation, signalling from both the epiblast as well as the extraembryonic tissues. Furthermore, it becomes clear that TGFbeta is indeed important for proper vessel formation and that it might use endoglin, as well as ALK-1, ALK-5 and Smad5 to mediate this function. Further analyses of these mice should provide a clearer understanding of the mechanism of TGFbeta action in vascular development and remodelling. PMID- 10853823 TI - Expression of the E2F family of transcription factors during murine development. AB - The E2F family of transcription factors plays a crucial role in the control of cell cycle progression and regulation of cellular proliferation, both processes fundamental to mammalian development. In the present study, we have examined the levels of expression of the six currently identified E2F proteins in murine embryos/fetuses as a function of gestational age, compared the expression of these six proteins in selected developing and adult tissues, and examined E2F expression in the embryonic murine palate, a tissue in which perturbation of proliferation is associated with induction of cleft palate. Our results indicate that: 1) multiple forms of individual E2F family members are present in embryonic, fetal and adult cells/tissues; 2) each of the six E2Fs is expressed in a tissue specific manner in both adult and embryonic/fetal organs; 3) certain forms of individual E2F family members are preferentially detected in adult tissues, whereas others are preferentially expressed in embryonic/fetal tissues; 4) expression of the various E2Fs and their isoforms follows distinct temporal patterns during murine gestation; and 5) individual E2F family members also exhibit differential patterns of temporal expression during murine palatogenesis. PMID- 10853824 TI - goosecoid expression represses Brachyury in embryonic stem cells and affects craniofacial development in chimeric mice. AB - The homeobox gene goosecoid, originally identified in Xenopus, is expressed in the organizer or its equivalent during gastrulation in the frog, chick, zebrafish and mouse. To investigate the role of goosecoid in mouse development, we have generated embryonic stem cells that stably overexpress the murine homolog of goosecoid. These cells show a repression of the gastrulation-associated gene Brachyury. Interestingly, repression of Brachyury is conserved between Xenopus and mouse despite the lack of conservation of the Brachyury promoter. Further characterization of the goosecoid-overexpressing ES cells revealed that they maintain the expression of stage-specific embryonic antigen-1, and teratomas derived from goosecoid-overexpressing cells show the presence of cell types derived from all three germ layers. Some highly chimeric mice derived from goosecoid-overexpressing cells displayed skull defects. These observations suggest that goosecoid may play a role in specification of anterior mesendodermal fates and specifically in mouse craniofacial development. PMID- 10853825 TI - Regeneration of halved embryonic lower first mouse molars: correlation with the distribution pattern of non dividing IDE cells, the putative organizers of morphogenetic units, the cusps. AB - Recently we demonstrated that non-cycling, cap-stage, mouse molar inner dental epithelial (IDE) cells corresponding to the primary enamel knot (EK) area underwent a coordinated temporo-spatial patterning leading to their patchy irregular segregation at the tips of the forming cusps. These non-cycling cells were suggested to perhaps represent the organizers of the morphogenetic units (OMU), the cusps. The present study has analyzed the regenerative capacity of halved cap-stage first lower mouse molars through three dimensional (3D) reconstructions. Partial regeneration of the anterior half and possible complete regeneration of the posterior half were documented. Using BrdU (5-bromo-2' deoxyuridine) labeling and 3D reconstructions of the IDE, we have correlated the patterns of cusp regeneration with the distribution of BrdU negative IDE cells. These data support a morphogenetic role for the non-cycling IDE cells. PMID- 10853826 TI - Snail is an immediate early target gene of parathyroid hormone related peptide signaling in parietal endoderm formation. AB - In mouse development, parietal endoderm (PE) is formed from both primitive endoderm (PrE) and visceral endoderm (VE). This process can be mimicked in vitro by using F9 embryonal carcinoma cells (EC) cells, differentiated to PrE or VE cells, and treating these with Parathyroid Hormone related Peptide (PTHrP). By means of differential display RT-PCR, we identified Snail (Sna) as a gene upregulated during the differentiation from F9 PrE to PE. We show that Sna is an immediate early target gene of PTHrP action in the formation of F9 PE cells. Using RT-PCR, we detected Sna transcripts in pre-implantation mouse embryos from the zygote-stage onwards. Sna was strongly upregulated in parallel with type 1 PTH/PTHrP Receptor (PTH(rP)-R1) mRNA in mouse blastocysts plated in culture, concomitant with detection of the PE-marker Follistatin and appearance of PE cells. By radioactive in situ hybridization on sections of mouse embryos, we found Sna expression in the earliest PE cells at E5.5. Sna remained expressed until at least E7.5. At this stage, we also observed clear expression in endoderm cells delaminating from the epithelial sheet of VE cells in the marginal zone. We conclude that PTH(rP)-R1 and Sna are expressed in endodermal cells that change from an epithelial to a mesenchymal phenotype. Since Sna expression has been described at other sites where epithelio-mesenchymal transitions (EMT) occur, such as the primitive streak at gastrulation and in pre-migratory neural crest cells, we hypothesize that Sna is instrumental in the action of PTHrP inducing PE formation, which we propose to be the first EMT in mouse development. PMID- 10853827 TI - Endochondral bone formation in toothless (osteopetrotic) rats: failures of chondrocyte patterning and type X collagen expression. AB - The pacemaker of endochondral bone growth is cell division and hypertrophy of chondrocytes. The developmental stages of chondrocytes, characterized by the expression of collagen types II and X, are arranged in arrays across the growth zone. Mutations in collagen II and X genes as well as the absence of their gene products lead to different, altered patterns of chondrocyte stages which remain aligned across the growth plate (GP). Here we analyze GP of rats bearing the mutation toothless (tl) which, apart from bone defects, develop a progressive, severe chondrodystrophy during postnatal weeks 3 to 6. Mutant GP exhibited disorganized, non-aligned chondrocytes and mineralized metaphyseal bone but without cartilage mineralization or cartilaginous extensions into the metaphysis. Expression of mRNA coding for collagen types II (Col II) and X (Col X) was examined in the tibial GP by in situ hybridization. Mutant rats at 2 weeks exhibited Col II RNA expression and some hypertrophied chondrocytes (HC) but no Col X RNA was detected. By 3rd week, HC had largely disappeared from the central part of the mutant GP and Col II RNA expression was present but weak and in 2 separate bands. Peripherally the GP contained HC but without Col X RNA expression. This abnormal pattern was exacerbated by the fourth week. Bone mineralized but cartilage in the GP did not. These data suggest that the tl mutation involves a regulatory function for chondrocyte maturation, including Col X RNA synthesis and mineralization, and that the GP abnormalities are related to the Col X deficiency. The differences in patterning in the tl rat GP compared to direct Col X mutations may be explained by compensatory effects. PMID- 10853828 TI - Characterization of desmosomal component expression during palatogenesis. AB - Adhesion of the opposing palatal shelves is a critical first step in the mechanism for palatal fusion. Formation of desmosomal junctions between the two medial edge epithelia provides a mechanism for palatal shelf adhesion. RT-PCR and immunohistochemistry were used to determine the pattern of expression of desmosomal components during palatogenesis. Desmosomal expression was specifically upregulated in the medial edge epithelia (MEE) at the early stages of palatal fusion as detected by both immunohistochemistry and electron microscopy. RT-PCR characterization of the desmosomal components detected all known elements, except desmocollin 1 (DSC1). Desmocollin 2 (DSC2) was expressed as both the DSC2a and DSC2b variants. The two variants are expressed at the same level. Western analysis of desmoglein expression paralleled the RT-PCR result. The temporal and spatial upregulation of desmosomal gene expression is evidence that the MEE induce new gene expression required to accomplish palatal shelf adhesion and initiate the first stage of palatal fusion. PMID- 10853829 TI - Germ cell-specific expression of green fluorescent protein in transgenic rainbow trout under control of the rainbow trout vasa-like gene promoter. AB - A technique to identify and isolate live fish primordial germ cells (PGCs) has not been established, in spite of the importance of purified germ cells for molecular and cellular studies. In rainbow trout, the distribution of vasa transcripts is restricted to the germ cell lineage, making this transcript a useful indicator of PGCs. Therefore, in this study, we cloned and characterized the rainbow trout vasa-like gene (RtVLG) regulatory regions and produced transgenic trout carrying the green fluorescent protein (GFP) gene driven by the RtVLG regulatory regions (p vasa-GFP) in order to identify live PGCs in vivo. In transgenic trout carrying the p vasa-GFP construct, cells showing green fluorescence were first observed at the mid-blastula stage; however, no cell-type specific expression was observed at this stage. At the eyed stage, about 30% of the transgenic embryos showed specific GFP expression in PGCs, and at the hatching stage, about 70% of the transgenic embryos did so. An immunohistochemical study of hatching stage embryos revealed that the GFP expressing cells are located in genital ridges. This transgenic trout, having visualizable PGCs, will make it possible to isolate live PGCs for in vitro studies and to study the ontogeny of PGCs including sex differentiation in live embryos. PMID- 10853830 TI - Expression patterns of follistatin and two follistatin-related proteins during mouse development. AB - We compared the expression patterns of follistatin and two follistatin-related proteins (FRP and m7365) during early mouse development. m7365 is expressed continuously during preimplantation development, in contrast to FRP and follistatin. At early postimplantation stages, follistatin and 7365 are expressed from E6.0, while FRP is detected from E7.5 onwards. Although there is some overlap between the expression of these genes in the primitive streak and somites, their overall expression patterns are distinct. PMID- 10853831 TI - Bulging medial edge epithelial cells and palatal fusion. AB - The surface of the medial edge epithelium of embryonic day 12, 13 and 14 mouse palatal shelves was observed utilising Environmental Scanning Electron Microscopy (ESEM). This technique offers the advantage of visualisation of biological samples after short fixation times in their natural hydrated state. Bulging epithelial cells were observed consistently on the medial edge epithelium prior to palatal shelf fusion. Additionally, we have used ESEM to compare the morphology and surface features of palatal shelves from embryonic day 13 to 16 mouse embryos that are homozygous null (TGF-beta3 -/-), heterozygous (TGF-beta3 +/-) or homozygous normal (TGF-beta3 +/+) for transforming growth factor beta-3 (TGF-beta3). At embryonic day 15 and 16 most TGF-beta3 +/- and +/+ embryos showed total palatal fusion, whilst all TGF-beta3 null mutants had cleft palate: the middle third of the palatal shelves had adhered, leaving an anterior and posterior cleft. From embryonic day 14 to 16 abundant cells were observed bulging on the medial edge epithelial surface of palates from the TGF-beta3 +/- and +/+ embryos. However, they were never seen in the TGF-beta3 null embryos, suggesting that these surface bulges might be important in palatal fusion and that their normal differentiation is induced by TGF-beta3. The expression pattern of E Cadherin, beta-catenin, chondroitin sulphate proteoglycan, beta-Actin and vinculin as assayed by immunocytochemistry in these cells shows specific variations that suggest their importance in palatal shelf adhesion. PMID- 10853832 TI - Differential expression of laminin-5 subunits during incisor and molar development in the mouse. AB - Rodent incisors are continuously growing teeth and enamel deposition is restricted to the labial side. In the present study, the expression of laminin-5 subunits (alpha3, beta3 and gamma2) has been analyzed by in situ hybridization in developing mouse lower incisors and compared to that reported in the molar. At the bud stage (E12), mRNAs for all subunits were detected in the whole epithelial thickening. At E14, when histogenesis had started, transcripts for alpha3 and gamma2 subunits were restricted to the outer dental epithelium (ODE), whereas the beta3 subunit was intensely expressed in the inner dental epithelium (IDE). A transient expression for alpha3 subunit was seen in the enamel knot area and disappeared at E15. Subsequently, all laminin-5 subunit genes were re-expressed in differentiating ameloblasts on the labial side. Similar patterns of transcription were observed in incisor and molar, suggesting that the differential expression of laminin-5 subunits in the IDE might be involved in the histogenesis of the IDE and ameloblast differentiation. At E16.5, cells of the IDE at the anterior extremity of the incisor and in the anterior part of the lingual IDE expressed transcripts for alpha3 and beta3 but not for gamma2 subunit. Similar expression patterns were observed in the enamel-free areas of the E18 molar. This specific expression might thus be related to cells that do not differentiate as functional ameloblasts. Throughout incisor development, intense expression for all laminin-5 subunits was restricted to the labial side of the cervical loop. The asymmetrical expression of laminin-5 might be related to incisor morphogenesis and to the differences in histogenesis and cytodifferentiation of the IDE that exist in the labial versus lingual aspect of the cervical loop. PMID- 10853833 TI - Study on measuring burden of disease. AB - DALY (Disability Adjusted Life Years) was recommended as a new indicator to measure burden of disease (BOD). Although BOD combines information from both morbidity and mortality, it only reflects the burden from the patients themselves because of their illness or death. As a common indicator, BOD should not only include the burden from the patients, but also the burden to the society around the patients, such as the input and support from the society to the ill person, and the losses from the related events. The aim of this study is to explore the scope and the magnitude of the burden to the society using stroke as an example. Results show that the burden due to time lost for caring for patients in hospitals accounts for 2.4% of total BOD (in a narrow sense), which indicates that BOD may be underestimated if the burden to the society is ignored. PMID- 10853834 TI - Cigarette smoking and urinary organic sulfides. AB - In order to observe how cigarette smoking influences levels of thio-thiazolidine 4-carboxylic acid (TTCA), high performance liquid chromatography (HPLC) was used to detect TTCA in urine from 18 healthy male volunteers. At the same time, the total amount of urinary organic sulfides was determined by the iodine azide test (IAT). Nine of the volunteers had smoking histories (5 to 10 cigarettes per day, as the smoking group), and the rest only occasionally smoke (1 to 2 cigarettes per month, as the control group). Samples were collected in the early morning (limosis) and 90 minutes after smoking a cigarette. Results showed that smoking a single cigarette could elevate the level of urinary organic sulfides both in the smoking and control groups, while a smoking habit appeared to have no significant influence on the urinary organic sulfide level. No significant cumulative effect of cigarette smoking on urinary organic sulfides was found. The influence of cigarette on urinary organic sulfides was temporary. The results suggest that cigarette smoking might be a confounding factor in biomonitoring the levels of carbon disulfide in exposed workers. PMID- 10853835 TI - Differential genotoxicity of the crude leaf extract of a medicinal plant, Casearia tomentosa. AB - The genotoxic potentiality of the crude leaf extract of Casearia tomentosa, a medicinal preparation, has been evaluated in Swiss albino mice. The extract significantly induced the division-disruptive chromosomal changes in bone marrow cells as well as in primary spermatocytes; the latter also exhibited marked increase in synaptic disruptions. A significant decrease in sperm count was noted. The incidence of structural damages in chromosomes, however, remained within the range of control level frequency. This herbal preparation, therefore, appears to be primarily spindle-poisoning in its action, but not clastogenic. The probable mechanism of this differential genotoxicity is discussed. PMID- 10853836 TI - Changes in tissue metals after cadmium intoxication and intervention with chlorpromazine in male rats. AB - Cadmium (Cd), one of the most dangerous heavy metals, has a very similar ionic radius to calcium (Ca). The interference of cadmium in calcium homeostasis may play an important role in cadmium toxicity. Recent reports indicate that calmodulin (CaM) inhibitors such as trifluoperazine and chlorpromazine (CPZ) could protect rodents against cadmium toxicity. It was also reported that pretreatment of mice with zinc (Zn) could reduce the adverse effects induced by cadmium. The aim of this study is to determine whether Cd changes the balance of other essential metals such as Zn and copper (Cu) in rat tissues, and whether CPZ can reverse these changes which are induced by cadmium intoxication. Adult male Sprague-Dawley (SD) rats were injected intraperitoneally (i.p.) with cadmium chloride (CdCl2) (0.2, 0.4, 0.8 mg Cd/kg body weight) alone and 0.4 mg Cd/kg in association with CPZ (5 mg/kg) daily for a week. The control animals were injected with normal saline only. The results showed that the cadmium content in the liver, kidney and testis increased significantly with a dose-response relationship. Cadmium treatment markedly increased the Zn and Ca content in some of the tissues. Hepatic and renal metallothionein (MT) increased significantly after cadmium intoxication. CPZ treatment, however, reduced cadmium content in liver, but not blood and kidney. CPZ seemed to decrease the content of MT in liver and significantly increase the amounts of MT in kidney. These data suggest that the intervention of cadmium with tissue essential metals may play a role in cadmium toxicity in rats, and calmodulin inhibitors to some extent can reduce the adverse effect of cadmium by decreasing the cadmium load in tissues and reversing the unbalance of essential metals. PMID- 10853837 TI - Effect of TCDD on maternal toxicity and chorionic gonadotropin--bioactivity in the immediate post-implantation period of macaque. AB - The purpose of this experiment was to observe the alterations in bioactivity of chorionic gonadotropin (CG) associated with early fetal loss (EFL), induced by the environmental toxin TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) in the cynomolgus macaque. Ten of twelve females administered single doses of 1, 2 or 4 microg/kg TCDD on gestational day (GD) 12 had EFL from ten to twenty days later. Seven control animals treated only with the vehicle had normal pregnancies. Blood samples were repeatedly collected for hormone evaluation, from two days before treatment to thirty-one days following treatment. Immunoreactive monkey chorionic gonadotropin (mCG) was measured in serum using ELISA, and bioactive mCG was measured using a luminescence LH/CG bioassay. No change in immunoreactive mCG levels was detected as a result of TCDD, treatment, but bioactive mCG levels were significantly lower in TCDD-treated animals compared to controls. This change in bioactivity of mCG was also reflected in the ratio of mCG bioactivity to mCG immunoreactivity (B/I ratio) which began to rise in normal pregnancies by GD 20, but did not rise in TCDD treated animals. These results demonstrate that normal pregnancy in the monkey, as in humans, is characterized by a post-implantation change in the B/I ratio of CG. These findings therefore suggest that changes in the production of bioactive CG may be used as a biomarker of environmental toxicant exposures which lead to EFL. PMID- 10853838 TI - Monitoring of human exposure to radiation with the binucleated lymphocyte micronucleus assay. AB - The micronuclei (MN) of peripheral blood lymphocytes from radiation-exposed people were monitored with the binucleated lymphocyte micronucleus assay (CBMN). MN rates in people with radiation-disease, radiation exposed and a control group were 12.57/1000, 4.20/1000 and 3.26/1000, respectively. The MN rate of patients with radiation-disease was significantly higher than those of other groups (P < 0.01). The difference between the radiation-exposed group and control group was not significant (P > 0.05). Meanwhile, chromosome aberrations (CA) of 3 groups were determined. The results were similar to those seen while the MN assay. CA rates were 2.06%, 0.93% and 0.69%, respectively. CA rate of the radiation-disease group was significantly higher than that of other groups (P < 0.05, P < 0.01). The difference between the radiation-exposed group and the control group was not significant (P > 0.05). The study indicates that the CBMN assay is a rapid, sensitive and accurate method which can be used to monitor a large population exposed to radiation. PMID- 10853839 TI - Integrated low-cost wastewater treatment for reuse in irrigation. AB - For sustainable wastewater management in developing countries, the implementation of low-cost, simple treatment systems should be encouraged. In this study, the performance of three treatment schemes was evaluated. The first step in all schemes was upflow anaerobic sludge blanket (UASB). The post treatment was either Algal Pond (AP). Lemna Pond (LP) or Rotating Biological Contactor (RBC). The results show that the performance of the UASB was satisfactory. Mean COD and BOD removal values were 78% and 85% respectively. The combination of UASB with an AP achieved significant improvement in the microbiological quality of the effluent. The geometric mean of fecal coliform in the effluent was 1.3 x 10(3) MPN/100 ml. Residual COD was 143 mg O2/L. This relatively high value was due to the presence of algae in the AP effluent. The use of the LP as a post treatment achieved better quality effluent. As indicated by the physico-chemical characteristics. However, fecal coliform removal was less by one log as compared to the AP effluent. When the RBC was used as a second stage. COD and BOD removal rates were 47% and 66% respectively. Also complete nitrification took place. Fecal coliform density declined by 5 logs. PMID- 10853840 TI - Effects of cigarette smoking and smoking cessation on plasma constituents and enzyme activities related to oxidative stress. AB - In order to study effects of cigarette smoking and smoking cessation on plasma constituents and enzyme activities related to oxidative stress, 1255 smokers and 524 healthy non-smokers were investigated in terms of plasma levels of lipoperoxides (LPO), nitric oxide (NO), vitamin C (VC), vitamin E (VE) and beta carotene (beta-CAR). Additionally, erythrocytes were examined to determine the level of LPO, the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px). The results showed that, when compared with the average values of the non-smoker group, the average plasma values of LPO, NO and the average erythrocyte value of LPO in the smoker group were significantly increased (P < 0. 001), while the average plasma values of VC, VE, beta-CAR, and the average erythrocyte activities of SOD, CAT, GSH-Px were significantly decreased (P < 0.001). A linear regression and correlation analysis for 65 male smokers who were all 40 years old showed that with longer smoking duration and greater daily smoking quantity, the plasma values of LPO, NO and the erythrocyte value of LPO were elevated, while the plasma values of VC, VE, beta-CAR and erythrocyte values of SOD, CAT, GSH-Px were decreased. In a group of 73 smokers who stopped smoking completely for six months, the average plasma values of LPO, NO and the average erythrocyte value of LPO decreased, although they were still significantly higher than those in the matched non-smoker group (P < 0.05). Additionally, the average plasma values of VC, VE, beta-CAR and the average erythrocyte values of SOD, CAT, GSH-Px increased, although they were still significantly lower than those in the matched non-smoker group (P < 0.05). However, after smoking cessation for one year the above average values were not significantly different from those in the matched non-smoker group (P > 0.05). This finding indicates that the markedly increased oxidative stress in smokers might gradually return to normal but only after a long period of smoking cessation. In conclusion, in the bodies of smokers a series of free radical chain reactions were gravely aggravated, the dynamic balance between oxidation and antioxidation was seriously disrupted, and oxidative stress was clearly exacerbated, which is closely related to many disorders or diseases in smokers. The present study underscored the need, urgency and importance of complete smoking cessation. PMID- 10853841 TI - Biological degradation of some organic compounds enrolled in paper industry--a pollution prevention approach. AB - Evaluation of the elimination and the "ultimate" biodegradation by aerobic microorganisms of some organic compounds commonly used in paper manufacturing technology was investigated. Biodegradation lines of nine organic compounds were determined as percentage removal of chemical oxygen demand (COD) over 7 days incubation. The results of the biodegradability test clearly revealed that some of the organic compounds under investigation are highly biodegradable, while others rank from fairly to even non-biodegradable. Significant biodegradation results were recorded for anti-coating ester (95.0%), basoplast 200D (85.3%) and basoplast PR 8050 (87.6%). A bleaching agent (formamidin-sulfinic acid), ukanol BSA and solidurit KM demonstrate moderate biodegradation results of 62.1%, 76.2% and 69.8%, respectively. Poor biodegradation results for Hedifix M/35 (12.7%), basazol orange (34.9%) and basazol brown (29.0%) were recorded. Accordingly, appropriate precaution should be taken into consideration when using these compounds for industrial applications. PMID- 10853842 TI - The problem of performing adequately sized randomized trials to demonstrate small survival benefits. PMID- 10853843 TI - Sequential re-analysis of a phase-III clinical trial in non-small cell lung cancer. AB - This paper presents a reanalysis of a randomized clinical trial conducted by the Cancer and Leukemia Group B (CALGB, Bethesda, MD, USA). This trial found a significant benefit of combination chemotherapy followed by irradiation (CTRT) in comparison to radiotherapy alone (RT) for the treatment of nonsmall cell lung cancer. The validity of the results obtained and the decision to terminate taken by the CALGB, were assessed using sequential methods. The reliability and efficiency of sequential methods were also assessed for this study. Two sequential designs were used: the triangular and the restricted procedure. Initial analyses were conducted with the data from patients actually recruited, adjusting for important prognostic variables at any interim analysis. As a confirmatory technique, a continuation of the trial was simulated, sampling extra patients under the assumption of no treatment difference, preserving the effect of the prognostic variables. Using the results from the 155 patients recruited by the CALGB (88 deaths at termination and 136 after follow-up), the sample path stayed within the continuation region of both sequential designs considered. An underpowered sequential analysis showed significant superiority of CTRT over RT (95% confidence interval (95% CI) 0.50-0.96, p=0.03 for the triangular; 95% CI 0.37-0.88, p=0.01 for the restricted procedure). Conventional analysis of the follow-up data also showed significant superiority of CTRT. The trial extended with simulated data ended at 60 months with 251 patients (178 deaths), showing significant superiority of CTRT under both designs (95% CI for hazard ratio 0.55 0.97). The two sequential procedures would have led to the same conclusion as that reached by the Cancer and Leukemia Group B, still achieving considerable savings in patients recruited and time over the conventional design. The data simulated under the rather conservative null hypothesis did not reverse the positive result claimed by the Cancer and Leukemia Group B. PMID- 10853844 TI - Exercise capacity and extent of resection as predictors of surgical risk in lung cancer. AB - Lung resection remains the most effective treatment for non-small cell lung cancer (NSCLC). However, there is no consensus about reliable operative risk assessment in these patients. The aim of this study was to identify predictors of postoperative complications and death after lung resection for NSCLC. In this prospective trial, 125 of 142 (88%) consecutive NSCLC patients from 1990 to August 1997 had complete data sets. All underwent functional assessment including spirometry and cardiopulmonary exercise tests and lung resection via thoracotomy. Complications occurred in 31 of 125 (25%) patients including 2 (1.6%) deaths. On logistic regression analysis, only maximal oxygen uptake (V'O2,max) x kg body weight(-1) expressed as a percentage of the predicted value (p<0.0001) and the estimated extent of lung tissue resection (p=0.02) were independent predictors of postoperative complications. Six of seven patients with a V'O2,max x kg body weight(-1) of <60% pred, but only eight of 65 with values >90% pred, exhibited postoperative complications. Maximal oxygen uptake and the estimated extent of lung tissue resection are independent predictors of postoperative complications. These simple parameters should be integrated into the preoperative decision analysis for operability in patients undergoing lung resection for lung cancer. PMID- 10853845 TI - Airway resistance and atopy in preschool children with wheeze and cough. AB - The extent to which the measurement of airways resistance by the interrupter technique (Rint) distinguishes preschool children with previous wheeze from those with no respiratory symptoms and helps to classify subjects with persistent cough, was investigated. Rint was measured before and after salbutamol treatment in 82 children with recurrent wheeze, 58 with isolated cough and 48 with no symptoms (control subjects). Their mean age (range) was 3.7 yrs (2-<5 yrs). Median baseline Rint was higher (p<0.0001) in wheezers than in either coughers or control subjects (1.16, 0.94 and 0.88 kPa x L(-1) x s(-1) respectively); coughers did not differ significantly from control subjects (p=0.14). The median ratios of baseline to post-salbutamol measurements (bronchodilator response (BDR)) in the groups differed significantly (1.40, 1.27 and 1.07, p< or =0.01 for all), suggesting that coughers occupy an intermediate position. A BDR ratio of >1.22 had a specificity and sensitivity for wheeze of 80% and 76% respectively. Twenty eight coughers had a BDR ratio >1.22. Wheezers' immunoglobulin E was inversely related to baseline Rint. It is concluded that measurements of airway resistance by the interrupter technique are useful for classifying preschool children with respiratory symptoms and could be used to monitor the effect of interventions. The relation between atopy and airways resistance suggests that they have separate roles in preschool wheezing. Coughers with a high bronchodilator response could represent "cough-variant" asthma in children who have baseline airway resistance by the interrupter technique measurements similar to control subjects. Whether these children develop classical asthma will only be known at follow-up later in childhood. PMID- 10853846 TI - Exhaled nitric oxide, serum ECP and airway responsiveness in mild asthmatic children. AB - The purpose of the present study was to assess the possible relationships between exhaled nitric oxide (ENO), a circulating marker of eosinophil activation, serum eosinophil cationic protein (SECP), level of airway responsiveness to methacholine and lung function in asthmatic children, as well as to compare these markers between children with and without inhaled steroid therapy. In a cross sectional study ENO, SECP and bronchial hyperresponsiveness to methacholine were evaluated in a group of 57 asthmatic children (21 without and 36 with regulator inhaled steroid therapy; aged 6-13 yrs). ENO was significantly lower in steroid treated children (p<0.01). No significant differences between steroid treated and untreated children were observed for the provocative concentration of methacholine causing a 20% fall in forced expiratory volume in one second (FEV1; PC20), SECP and FEV1. In the whole study population significant increase correlations were observed between PC20 and SECP (r=-0.329, p=0.013) and between ENO and FEV1% of predicted (r=-0.348, p<0.01). In the group not receiving inhaled steroids the inverse relationship between PC20 and SECP was more evident (r= 0.581, p<0.001). In the steroid-treated group a significant inverse relationship was observed between ENO and FEV1 (r=-0.426, p=0.0011). The level of exhaled nitric oxide and the relationships between lung function, bronchial reactivity and markers of inflammation are different between steroid-treated and untreated asthmatic children. This has implications for the monitoring of asthma in childhood. PMID- 10853847 TI - Asthma mortality in Danish children and young adults, 1973-1994: epidemiology and validity of death certificates. AB - Several reports indicate that asthma mortality has increased during the last few decades. International comparisons reveal some striking differences in the pattern of asthma mortality. The authors investigated the asthma mortality rate in the Danish child and youth population 1973-1994 and studied the validity of death certificates. The authors reviewed all death certificates coded as asthma death in the International Classification of Diseases (ICD 8-ICD 10 (1994)) and adjacent respiratory code numbers for the age group 1-19 yrs. Hospital records and autopsy reports were assessed to validate the cause of death. Age standardized and age-specific mortality rates were calculated. From 1973 to 1987 there was a significant upward trend in the mortality. On subdivision, this trend was limited to the age group 15-19 yrs. Generally the mortality rate decreased from 1988 to 1994. Four per cent coded as asthma were false positive. Twelve per cent were false negative asthma deaths, wrongly coded as due to other causes. Only 62% of all true positive death caused by asthma were appropriately coded. The number of false negative certifications increased with increasing autopsy frequency. Asthma mortality rates in Denmark increased in adolescents during 1973 1987 and decreased from 1988 to 1994. A possible explanation may be an increased awareness of asthma symptoms combined with a steadily improved treatment of asthma. Even in children and young adults under the age of 20 yrs, validity problems still make comparisons between countries difficult; even interpretation of national trends requires caution. PMID- 10853848 TI - Combined use of exhaled nitric oxide and airway hyperresponsiveness in characterizing asthma in a large population survey. AB - The aim of the present study was to see whether measurements of airway hyperresponsiveness (AHR) and nitric oxide (NO) in exhaled air (ENO) either separately or in combination, could differentiate between asthmatics and healthy control subjects in a population based survey. In central Norway 8,571 adolescents participated in a large-scale epidemiological survey (Young Helseundersokelsen i Nord-Trondelag (Health Survey in North-Trondelag; HUNT). Asthmatic symptoms when exposed to pollen, pets or house-dust were reported by 7.8% (suspected asthmatics), while 56% reported no asthmatic or allergic symptoms (control subjects). From these respective groups 151 and 213 adolescents were investigated with allergy screening, measurements of exhaled and nasal NO, and methacholine challenge test. AHR (provocative dose of methacholine causing a 20% fall in forced expiratory volume in one second (PD20) <2 mg) was confirmed in 75% of the suspected asthmatics versus 25% of the control subjects, whereas 52% versus 20% had elevated levels of ENO (> or =8 parts per billion (ppb)). ENO and dose response ratio to methacholine (DRR) were positively correlated (r=0.41, p<0.001). ENO was significantly elevated in atopic versus nonatopic suspected asthmatics (11.7 ppb and 5.6 ppb respectively, p<0.001). Suspected asthmatics with both AHR and atopy had the highest levels of ENO (14.2 ppb). It is concluded that measurements of nitric oxide in exhaled air alone are not a useful tool in diagnosing asthma in population surveys, but that the combination of airway hyperresponsiveness and elevated nitric oxide in exhaled air is a very specific finding for allergic asthma. The use of dose response ratio to methacholine did not provide any additional information to the provocative dose of methacholine causing a 20% fall in forced expiratory volume in one second in this study. PMID- 10853849 TI - Effects of azithromycin on ozone-induced airway neutrophilia and cytokine release. AB - Exposure of humans to ozone causes increased neutrophils and inflammatory cytokines in airway lining fluid. Recent research shows that macrolide antibiotics may reduce interleukin (IL)-8 production by bronchial epithelial cells and inhibit neutrophil chemotaxis. A double-blind, cross-over study was performed in which 12 healthy subjects underwent two separate 4-h exposures to 0.2 parts per million ozone while exercising intermittently. In the 73.5 h before exposure, subjects were pretreated with either 1,250 mg azithromycin or placebo. Sputum induction conducted 74 h pre- and 18 h post-exposure was used to measure total cells, per cent neutrophils, IL-6, and IL-8. There were significant (p<0.05) pre- to post-exposure increases in total cells, neutrophils, IL-6 and IL 8 in both the azithromycin and placebo arms. However, no significant differences were found between azithromycin and placebo conditions in the post- minus pre exposure value for these variables. The results suggest that in healthy subjects, in the design used, azithromycin, in usual clinical doses, does not have anti inflammatory effects on human airways as indicated in the measured variables. PMID- 10853850 TI - Oral and bronchial provocation tests with aspirin for diagnosis of aspirin induced asthma. AB - In 35 asthmatic patients with acetylsalicylic acid (aspirin; ASA) intolerance (AIA) and 15 asthmatics tolerating ASA well, the authors compared the diagnostic value of the placebo-controlled oral ASA versus inhaled L-lysine (L) ASA challenges. All AIA subjects gave a history of asthmatic attacks following ingestion of ASA and in all of them the intolerance was confirmed by oral challenge test over the past 10 yrs. Doses of ASA increasing in geometric progression were used in oral tests 10-312 mg (cumulative dose 500 mg); in bronchial tests 0.18-115 mg (cumulative dose 182 mg). Either challenge was considered as positive, if forced expiratory volume in one second (FEV1) dropped at least 20% from the baseline value and/or strong extrabronchial symptoms of intolerance occurred. Urinary leukotriene E4 excretion was determined at baseline and following the challenges. In 24 out of 35 patients the oral test was positive, based on a 20% decrease in FEV1. When including extrabronchial symptoms this was positive in 31 cases. Bronchial L-ASA challenge led to > or =20% fall FEV1 in 21 out of 35 cases, and in 27 cases when including extrabronchial symptoms. No correlation was observed between ASA provocative dose causing a 20% fall in FEV1, determined by the oral route compared to the inhalation route. Urinary LTE4 increased after both challenges the rise being higher following oral as compared to inhalation provocation (p=0.0001). It is concluded that both tests had similar specificity whilst the oral test showed a tendency to higher sensitivity for the clinical diagnosis of acetylsalicylic acid intolerance. The inclusion of extrabronchial symptoms into the criteria of test positivity enhanced the diagnostic value of both procedures. In both tests the highest leukotriene E4 increases were found in the presence of extrabronchial symptoms, suggesting the participation of tissues other than the lung in aspirin induced leukotriene E4 release to urine. PMID- 10853851 TI - Differential effects of nitric oxide synthase inhibitors in an in vivo allergic rat model. AB - The in vivo role of nitric oxide in inflammatory cell migration, vascular permeability and the development of hyperresponsiveness to methacholine (MCh) was studied in rats 24 h following ovalbumin (OVA) challenge. The NO synthase (NOS) inhibitors N(G)-mono-methyl-L-arginine (L-NMMA; nonselective), aminoguanidine (two-fold inducible NOS-selective), N(omega)-nitro-L-arginine methyl ester (L NAME; 2000-fold endothelial cell NOS-selective) or S-methyl-L-thiocitrulline (100 fold neuronal NOS-selective) were administered (100 mg x kg(-1) s.c.) to OVA sensitized Piebald-Virol-Glaxo rats on 3 consecutive days during which they were challenged with allergen (1% OVA). Responses to inhaled MCh were measured in anaesthetized animals 24 h after OVA challenge. Cellular inflammation and vascular permeability were assessed using bronchoalveolar lavage (BAL) fluid collected 30 min after administration of Evans blue (50 mg x kg(-1) i.v.). OVA challenge in sensitized animals induced hyperresponsiveness to MCh, inflammatory cell influx and increased leakage of Evans blue into the BAL fluid (n=9, p<0.001). Aminoguanidine was effective in inhibiting the allergen-induced cellular influx and microvascular leakage (n=9, p<0.001) without altering responses to MCh. This effect was reserved by L-arginine. L-NAME (n=5, p<0.01) and S-methyl-L-thiocitrulline (n=6, p<0.001) further potentiated the allergen induced hyperresponsiveness without altering cellular inflammation. L-NMMA attenuated both the OVA-induced cellular influx and Evans blue leakage (n=8, p<0.001) as well as further potentiating the hyperresponsiveness to MCh (p<0.05). From these studies, it is suggested that, in allergic Piebald-Virol-Glaxo rats, nitric oxide production by inducible nitric oxide synthase plays a role in the migration of inflammatory cells and increase in vascular permeability following allergen challenge, whereas nitric oxide produced by the constitutively expressed neuronal nitric oxide synthase limits hyperresponsiveness to methacholine. PMID- 10853852 TI - Long-term treatment of chronic obstructive pulmonary disease with salmeterol and the additive effect of ipratropium. AB - The efficacy and safety of salmeterol alone was compared with the combination of salmeterol plus ipratropium and with placebo during long-term treatment in patients with stable chronic obstructive pulmonary disease. In addition, the single-dose effect in response to the first dose of treatment was studied over 12 h. The patients (n=144; age 64+/-7 yrs, forced expiratory volume in one second (FEV1) 44+/-11% pred) participated in a three-centre double-blind double-placebo parallel group study and were randomized after a run-in period of 2 weeks to receive either salmeterol 50 microg b.i.d., salmeterol 5 microg b.i.d. plus ipratropium 40 microg q.i.d. or placebo for a period of 12 weeks. The single-dose study demonstrated that salmeterol produced a significant increase in FEV1 (peak of 7% pred) and specific airway conductance (sGaw) (maximum of 60% baseline) for > or =12 h. The combination of salmeterol plus ipratropium elicited a greater bronchodilator response (11% and 94% increases respectively) than salmeterol alone during the first 6 h after inhalation. During treatment there were significant improvements in daytime symptom scores and morning peak expiratory flow in both the salmeterol and the salmeterol plus ipratropium groups (p<0.001), with an associated decrease in the use of rescue salbutamol. Improvements in FEV1 and sGaw were greater in the salmeterol plus ipratropium group than in the patients receiving only salmeterol. Thirty-five patients had an exacerbation; 11 (23%) in the salmeterol group (versus placebo NS), six (13%) in the salmeterol plus ipratropium group (versus placebo p<0.01) and 18 (36%) in the placebo group. In conclusion, in patients with severe stable chronic obstructive pulmonary disease, long-term treatment with either salmeterol alone or salmeterol plus ipratropium is safe and effective. There was added benefit from the combination therapy in terms of improvement in airways obstruction, but not for improvement in symptom control or need for rescue salbutamol. PMID- 10853853 TI - Airways inflammation in chronic bronchitis: the effects of smoking and alpha1 antitrypsin deficiency. AB - Airways inflammation in chronic bronchitis is thought predominantly to be a direct consequence of neutrophil recruitment and release of elastase in response to factors such as cigarette smoke. The aims of this study were to assess the role of smoking and determine whether the serum elastase inhibitor alpha1 antitrypsin (alpha1AT) influenced the process. Airways inflammation was compared between patients with chronic obstructive bronchitis with (n=39) and without (n=42) severe alpha1AT deficiency. The authors assessed the sputum concentration of the neutrophil chemoattractants interleukin-8 (IL-8) and leukotriene (LT)B4, myeloperoxidase (MPO) as a marker of neutrophil influx, neutrophil elastase activity and its natural inhibitors, alpha1AT and secretory leukoprotease inhibitor (SLPI). Finally serum alpha1AT was measured to determine the degree of protein leakage (sputum sol serum alpha1AT ratio). Compared to current smokers, the exsmokers had a lower concentration of the chemoattractant IL-8 (p<0.05) and a lower MPO concentration, although this failed to reach conventional statistical significance (p=0.06). Patients with alpha1AT deficiency had greater inflammation in the larger airways with increased LTB4 (p<0.005), MPO (p<0.001), neutrophil elastase activity (p<0.01), protein leak (p<0.001), and were found to have a lower anti-proteinase screen with both reduced sputum alpha1AT (p<0.001) and SLPI concentrations (p<0.05). The reduction in sputum interleukin-8 levels in exsmokers may decrease neutrophil influx and thus explain the slower rate of neutrophil mediated progression of lung disease compared to subjects who continue to smoke. Patients with alpha1-antitrypsin deficiency had greater inflammation suggesting that alpha1-antitrypsin plays an important role in protecting the larger airways from the inflammatory effects of elastase activity and may explain their more rapid progression of disease. PMID- 10853854 TI - Gene polymorphism for microsomal epoxide hydrolase and susceptibility to emphysema in a Japanese population. AB - Recently, it was reported that gene polymorphism for microsomal epoxide hydrolase (mEPHX), an enzyme involved in the first-pass metabolism of epoxide intermediates, was associated with susceptibility to emphysema. This association was examined in a Japanese population, performing polymerase chain reaction (PCR) based direct sequencing and restriction fragment length polymorphism assays for variant forms of mEPHX. The subjects consisted of 79 smokers with moderate to severe emphysema diagnosed by lung computed tomography scans, 58 smokers without emphysema, with a comparable smoking history, and 114 consecutive subjects who undertook annual health checkups. The allele frequency of exon 3 Tyr113 to His113, which was reported to confer slow mEPHX activity, was substantially higher in the population control group compared with that of the Caucasian control subjects in a previous study. However, neither the genotype distribution of exon 3, nor that of exon 4 His139 to Arg139, was significantly different between the two groups of smokers. These data indicate that the gene polymorphism for mEPHX is not associated with susceptibility to emphysema in the Japanese population. The discrepancy between the two studies may be explained either by racial difference or by the selection bias of subjects in the previous study, which examined those who had only mild to moderate emphysema with lung cancer or those who were clinically diagnosed as having chronic obstructive pulmonary disease. PMID- 10853855 TI - G-CSF and IL-8 but not GM-CSF correlate with severity of pulmonary neutrophilia in acute respiratory distress syndrome. AB - Activated neutrophils play a major role in the pathogenesis of acute respiratory distress syndrome (ARDS), and persistence of pulmonary neutrophilia is related to poor survival. Interleukin (IL)-8 is implicated in recruiting neutrophils to the lungs but it has been postulated that granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF), which can promote the survival of neutrophils by delaying apoptosis, may prolong the inflammatory response. The aim of this study was to investigate the levels of GM CSF and G-CSF in the lungs of patients with ARDS and determine their relationship relative to IL-8 with levels of neutrophils and clinical outcome. The lungs of 31 patients with ARDS were sampled by means of bronchoalveolar lavage (BAL) and assays of the three cytokines were conducted via enzyme-linked immunosorbent assay. GM-CSF, G-CSF and IL-8 were all increased in the patients compared to healthy controls but concentrations of GM-CSF were much lower than those of G-CSF and IL-8 (GM-CSF75% pred (p<0.05). In conclusion, chronic smoking causes phenotypic changes in circulating polymorphonuclear leukocytes that are characteristic of chronic stimulation of the bone marrow and it is speculated that the increased number of immature polymorphonuclear leukocytes contributes to the chronic lung inflammation associated with cigarette smoking. PMID- 10853860 TI - Effect of serial-day exposure to nitrogen dioxide on airway and blood leukocytes and lymphocyte subsets. AB - Nitrogen dioxide (NO2) is a free radical-producing oxidant gas. Inhalation of NO2 could cause airway inflammation, and decrease immune function. This experiment tested the hypothesis that exposure to NO2 would: 1) increase leukocytes in bronchoalveolar lavage (BAL); and 2) change the distribution of lymphocyte subsets and activation in BAL and peripheral blood (PB). Using a counter balanced, repeated-measures design, 15 healthy volunteers were exposed to filtered air (FA) or 2.0 parts per million NO2 for 4 h x day(-1) (4 x 30 min of exercise), for three consecutive days. Bronchoscopy was performed 18 h following each exposure set, and PB was drawn pre-exposure and pre-bronchoscopy. Flow cytometry was used to enumerate lymphocyte subsets and activation makers in BAL and PB. In the bronchial fraction, there was an increase in the percentage of neutrophils following NO2 exposure compared to FA (median (interquartile range): 10.6 (4.8-17.2)% versus 5.3 (2.5-8.3)%; p=0.005). In the BAL, there was a decrease in the percentage of T-helper cells following NO2 exposure compared to FA (55.9 (40.8-62.7)% versus 61.6 (52.6-65.2)%; p=0.022). For PB, there were no between-condition differences in any leukocyte or lymphocyte subsets, or activation. In conclusion exposure to nitrogen dioxide results in bronchial inflammation and a minimal change in bronchoalveolar lavage T-helper cells, and no changes in peripheral blood cells. PMID- 10853861 TI - Nasal airflow dynamics: mechanisms and responses associated with an external nasal dilator strip. AB - The adhesive external nasal dilator strip (ENDS) is widely advocated for prevention of snoring and promotion of nasal breathing during exercise. In the present study, the effects of the ENDS on nasal airflow resistance (Rn) in normal subjects were examined and factors determining individual responses to the ENDS explored. Using posterior rhinomanometry, 20 healthy Caucasian adults (10 males, 10 females; age: 18-56 yrs) were studied during quiet tidal breathing and voluntary hyperpnoea with (ENDS) and without (control) ENDS and with a placebo strip (placebo) before and after application of a topical nasal decongestant (oxymetazoline hydrochloride). During tidal breathing, only nine subjects showed a significantly (p<0.05) decreased inspiratory and/or expiratory Rn with the ENDS ("responders"). During the control, inspiratory Rn (at 0.4 L x s(-1)) was higher in "responders" than "nonresponders" (3.28+/-0.16 versus 2.60+/-0.08 cmH2O x L( 1) x s; p=0.04). The effects of nasal decongestant and the ENDS were additive. During voluntary hyperpnoea, inspiratory Rn (at 1.0 L x s(-1)) and the hysteresis of the inspiratory transnasal pressure/flow curve were decreased with the ENDS in most subjects. It is concluded that the external nasal dilator strip influences nasal airflow dynamics by both dilation of the nasal valve and stabilization of the lateral nasal vestibule walls and may be more effective in subjects with a high resting nasal airflow resistance. PMID- 10853862 TI - Activation of deltaF508 CFTR in a cystic fibrosis respiratory epithelial cell line by 4-phenylbutyrate, genistein and CPX. AB - The cellular basis of cystic fibrosis (CF) is a defect in a cyclic adenosine monophosphate (cAMP)-activated chloride channel (CF transmembrane conductance regulator) in epithelial cells that leads to decreased chloride ion transport and impaired water transport across the cell membrane. This study investigated whether it was possible to activate the defective chloride channel in cystic fibrosis respiratory epithelial cells with 4-phenylbutyrate (4PBA), genistein and 8-cyclopentyl-1,3-dipropylxanthine (CPX). The CF bronchial epithelial cell line CFBE41o-, which expresses the deltaF508 mutation, was treated with these agents and loss of Cl-, indicating Cl- efflux, measured by X-ray microanalysis. 8-bromo cAMP alone did not induce Cl- efflux in CFBE41o- cells, but after incubation with 4PBA a significant efflux of Cl- occurred. Stimulation of cells with a combination of genistein and cAMP also induced Cl- efflux, whereas a combination of pretreatment with 4PBA and a combined stimulation with genistein and cAMP induced an even larger Cl- efflux. Cl- efflux could also be stimulated by CPX, but this effect was not enhanced by 4PBA pretreatment. The deltaF508 mutation leads to impaired processing of the cystic fibrosis transmembrane conductance regulator. The increased efflux of chloride after 4-phenylbutyrate treatment can be explained by the fact that 4-phenylbutyrate allows the deltaF508 cystic fibrosis transmembrane conductance regulator to escape degradation and to be transported to the cell surface. Genistein and 8-cyclopentyl-1,3-dipropylxanthine act by stimulating chloride ion efflux by increasing the probability of the cystic fibrosis transmembrane conductance regulator being open. The combination of 4-phenylbutyrate and genistein may be useful in a potential pharmacological therapy for cystic fibrosis patients with the deltaF508 mutation. PMID- 10853863 TI - Effect of 3 weeks' rehabilitation on neutrophil surface antigens and lung function in cystic fibrosis. AB - Neutrophil leukocytes have been shown to be the predominant cells in inflammatory airway infiltrates of cystic fibrosis (CF) patients. The aim of this study was to investigate the effect of rehabilitation on neutrophil surface antigen expression and lung function in healthy controls and stable CF patients with moderately severe disease. The absolute number of neutrophils and the level of surface marker expression on neutrophils were elevated in 12 CF patients compared with eight healthy controls. The level of neutrophil surface marker expression was similar in bronchoalveolar lavage fluid from CF patients who underwent bronchoscopy for diagnostic or therapeutic reasons. After 3 weeks' rehabilitation, there was a significant reduction in the expression of CD11b (complement receptor type 3), CD13 (aminopeptidase N), CD32 (low-affinity Fc gamma chain receptor II), and CD35 (complement receptor type 1) in only the CF patients. At the same time, lung function improved significantly. The increase in forced vital capacity correlated significantly with the decrease in CD32 level. These results demonstrate that rehabilitation in a specialized clinic can reduce the neutrophil-dominated inflammation and improve the lung function of stable CF patients with moderately severe disease even without changing any medications. PMID- 10853864 TI - A new simple method of staining exogenous surfactant in experimental research. AB - Two commonly used techniques in experimental lung research have helped to determine which variables influence surfactant distribution within the lung: radioactive labelling of surfactant components and admixture of coloured microspheres to surfactant. However, neither technique allows the description of surfactant distribution at the alveolar level. The aim of this study was to establish a new technique using histology colourants for admixture to exogenous surfactant to make exogenous surfactant visible by light microscopy. In a step by step approach the authors evaluated the properties of a variety of green colourants when added to a natural porcine surfactant preparation for their ability to homogeneously mix with surfactant, to bind to surfactant, to adhere to a glass slide, to not be "overstained" by standard haematoxylin-eosin and Elastica van Giesson staining, to not influence in vitro surface tension properties of surfactant using a Wilhelmy balance, to not influence oxygenation and ventilation in a lung-lavage rat model and to preserve their colour and adherence to exogenous surfactant on lung specimens visualized by light microscopy. Only one of the tested green histology colourants (Green Dye) fulfilled all requirements and showed a brilliant green colour in a distribution pattern typical of surfactant at the alveolar level. It is concluded that the authors have established a new, simple and inexpensive method of staining exogenous surfactant for evaluation of its distribution by light microscopy at the alveolar level. PMID- 10853865 TI - Measurement of inflammatory markers in the breath condensate of children with cystic fibrosis. AB - Identifying noninvasive markers of pulmonary inflammation would be useful in assessing new therapies in children. Breath condensate is a simple and potentially acceptable sample medium even in small children. The technique has previously been used in adults, but not children with cystic fibrosis. The technique was assessed in 36 children with cystic fibrosis (mean age 10.4 yrs) and 17 control subjects, analysing samples for nitrite, interleukin(IL)-8 and salivary and nasal contamination. Correlations were made between levels of the inflammatory markers and forced expiratory volume in one second/forced vital capacity, chest radiograph score and use of inhaled steroids. On samples without significant contamination (<10 u x L(-1) amylase) nitrite was detected in 93% of samples at a median concentration of 3.0 microM compared with 50% of control samples at a median of 0.5 microM. Condensate amylase levels did not correlate with the nitrite value obtained (r=0.31). IL-8 was detected in 33% of CF samples. Breath condensate is an acceptable method of sample collection in children. Nitrite was raised in breath condensate from patients with cystic fibrosis when compared with control subjects. PMID- 10853866 TI - Marcel Proust (1871-1922): reassessment of his asthma and other maladies. AB - Marcel Proust endured severe allergies and bronchial asthma from early childhood. Those who suffer from the frightening and recurrent pangs of asthma often become dependent on their parents particularly mother; Proust was no exception. In his time asthma was poorly understood by physicians who considered the illness to be a type of hysteria. Decades later, we now understand that the severe, poorly controlled, suffocating episodes of asthma were responsible for the complex persona that Marcel Proust had assumed. PMID- 10853867 TI - Airway smooth muscle cells: contributing to and regulating airway mucosal inflammation? AB - In addition to its contractile properties, airway smooth muscle may contribute to the pathogenesis of asthma by increased proliferation, and by the expression and secretion of pro-inflammatory cytokines and mediators. Studies of airway smooth muscle cells in culture have shown that many mitogenic mediators can induce proliferation, and that these may therefore, contribute to the increase in airway smooth muscle mass observed in asthma. Other mechanisms for airway smooth muscle proliferation include the interaction with inflammatory cells such as T-cells and eosinophils. Airway smooth muscle cells may also be a source of inflammatory mediators and cytokines, in particular chemokines, thus implicating airway smooth muscle cells as contributors to the inflammatory mechanisms of asthma. The pro activating signals for converting airway smooth muscle cells into a proliferative and secretory cell in asthma are unknown, but may include viruses and immunoglobulin E. Airway smooth muscle contractility may also be altered in response to inflammation. Airway smooth muscle cells may play an important interactive role with inflammatory and other structural cells, contributing to inflammation, injury and repair of the airways. Such a recognition makes it imperative to consider the airway smooth muscle as a target of therapeutic drugs for suppressing not only the contractile but also the proliferative and secretory effects of asthma. PMID- 10853868 TI - Breathing techniques--adjunctive treatment modalities for asthma? A systematic review. AB - Breathing techniques are used by a large proportion of asthma sufferers. This systematic review was aimed at determining whether or not these interventions are effective. Four independent literature searches identified six randomized controlled trials. The results of these studies are not uniform. Collectively the data imply that physiotherapeutic breathing techniques may have some potential in benefiting patients with asthma. The safety issue has so far not been addressed satisfactorily. It is concluded that too few studies have been carried out to warrant firm judgements. Further rigorous trials should be carried out in order to redress this situation. PMID- 10853869 TI - Airway neutrophils and interleukin-17. AB - It is well known that exacerbations of obstructive airways disease such as asthma and chronic obstructive pulmonary disease are associated with an increased number of neutrophils in the airways. However, the mechanisms behind this phenomenon are poorly understood. There is in vivo experimental evidence that the number of airway neutrophils is controlled by certain T-lymphocytes, but the mediators responsible for this lymphocyte-related neutrophilia have not yet been identified. In this review, novel evidence that the T-lymphocyte-related cytokine interleukin (IL)-17 can link the activation of certain T-lymphocytes to the recruitment and activation of airway neutrophils is described. The IL-17-induced neutrophil recruitment is mediated via induced CXC chemokine release through steroid-sensitive mechanisms and is modulated by release of endogenous tachykinins. These effects of IL-17 are potentiated by other pro-inflammatory cytokines such as (IL-1beta) and tumour necrosis factor-alpha. Clinical studies are needed to evaluate whether or not targeting these mechanisms can provide a useful pharmacotherapeutical approach against exaggerated mobilization of neutrophils in obstructive airways disease. PMID- 10853870 TI - Inhaled corticosteroids and Churg-Strauss syndrome: a report of five cases. AB - Churg-Strauss syndrome is an eosinophil-associated, small vessel granulomatous vasculitis, characterized by late onset asthma, upper airways disease, eosinophilia, and clinical manifestations of systemic vasculitis. Several cases of Churg-Strauss syndrome have been recognized in patients treated with cysteinyl leukotriene-receptor antagonists and weaned off systemic corticosteroids. These cases have led to a general warning on the possible development of Churg-Strauss syndrome after taking cysteinyl leukotriene-receptor antagonists. The authors report five cases of Churg-Strauss syndrome in severe steroid dependent asthmatics in whom inhaled corticosteroids allowed systemic corticosteroid withdrawal. It is concluded that physicians should monitor patients carefully when severe asthma is controlled with any substance allowing withdrawal from (or even avoidance) of systemic corticosteroids. Case-control studies should identify more precisely the risk factors of Churg-Strauss syndrome. PMID- 10853871 TI - Prediction of metabolic and cardiopulmonary responses to maximum cycle ergometry. PMID- 10853872 TI - QT-interval prolongation by non-cardiac drugs: lessons to be learned from recent experience. AB - BACKGROUND: Evidence has accrued that several non-cardiac drugs may prolong cardiac repolarisation (hence, the QT interval of the surface electrocardiogram) to such a degree that potentially life-threatening ventricular arrhythmias (e.g. torsades de pointes) may occur, especially in case of overdosage or pharmacokinetic interactions. DISCUSSION: This has fostered discussion on the molecular mechanisms underlying the class-III antiarrhythmic effect shared by apparently disparate classes of drugs, on the clinical relevance of this side effect and on possible guidelines to be followed by drug companies, ethics committees and regulatory agencies in the risk-benefit assessment of new and licensed drugs. This review provides an update on the different classes of non cardiac drugs reported to prolong the QT interval (e.g. histamine H1-receptor antagonists, antipsychotics, antidepressants and macrolides), on the possible underlying molecular mechanisms and on the clinical relevance of the QT prolonging effect. Identification and widespread knowledge of risk factors that may precipitate prolongation of the QT interval into life-threatening arrhythmias becomes an important issue. Risk factors include congenital long QT syndrome, clinically significant bradycardia or heart disease, electrolyte imbalance (especially hypokalaemia, hypomagnesaemia), impaired hepatic/renal function and concomitant treatment with other drugs with known potential for pharmacokinetic/ pharmacodynamic interactions (e.g. azole antifungals, macrolide antibacterials and class-I or -III antiarrhythmic agents). Future perspectives for drug research and development are also briefly outlined. PMID- 10853873 TI - The use of fixed-mixture insulins in clinical practice. AB - BACKGROUND: Pre-mixed insulins currently represent about 40% of the world market in human insulin. Despite the fact that many patients are being treated with these insulin formulations, there are surprisingly few data about fixed mixtures and comparisons with other administration schedules. The main advantages of using a pre-mixed preparation over a self-mixed insulin are increased accuracy of dosage, efficacy and patient convenience. These benefits may lead to increased compliance resulting in better long-term control of the disease. Insulin mixtures are used by a wide variety of patients with type-1 and type-2 diabetes. Pre-mixed insulins also appear to be useful in elderly and adolescent patients with diabetes, although there are few published data on patients in these groups. CONCLUSIONS: It is likely that, in the future, fixed mixtures containing rapid acting insulin analogues, such as insulin lispro, and possibly newly formulated intermediate-acting insulin analogues, such as neutral protamine lispro, may allow further improvements in both metabolic control and patient convenience. PMID- 10853874 TI - Evidence of clinical efficacy of homeopathy. A meta-analysis of clinical trials. HMRAG. Homeopathic Medicines Research Advisory Group. AB - OBJECTIVE: To establish, using a systematic review and meta-analysis, whether there is any evidence from randomised controlled clinical trials of the efficacy of homeopathic treatment in patients with any disease. DATA SOURCES: Published and unpublished reports of controlled clinical trials available up to June 1998, identified by searching bibliographic databases (Medline, Embase, Biosis, PsychInfo, Cinahl, British Library Stock Alert Service, SIGLE, Amed), references lists of selected papers, hand searching homeopathic journals and conference abstracts, and contacting pharmaceutical companies. TRIALS SELECTION: Trials were selected using an unblinded process by two reviewers. The selection criteria were randomised, controlled trials in which the efficacy of homeopathic treatment was assessed relative to placebo in patients using clinical or surrogate endpoints. Prevention trials or those evaluating only biological effects were excluded. One hundred and eighteen randomised trials were identified and evaluated for inclusion. Sixteen trials, representing 17 comparisons and including a total of 2,617 evaluated patients, fulfilled the inclusion criteria. DATA EXTRACTION: Data were extracted by two reviewers independently, using a summary form. Disagreements were resolved by a third person. DATA SYNTHESIS: The evidence was synthesised by combining the significance levels (P values) for the primary outcomes from the individual trials. The combined P value for the 17 comparisons was highly significant P = 0.000036. However, sensitivity analysis showed that the P value tended towards a non-significant value (P = 0.08) as trials were excluded in a stepwise manner based on their level of quality. CONCLUSIONS: There is some evidence that homeopathic treatments are more effective than placebo; however, the strength of this evidence is low because of the low methodological quality of the trials. Studies of high methodological quality were more likely to be negative than the lower quality studies. Further high quality studies are needed to confirm these results. PMID- 10853875 TI - Nilvadipine protects low-density lipoprotein cholesterol from in vivo oxidation in hypertensive patients with risk factors for atherosclerosis. AB - OBJECTIVE: Nilvadipine, a calcium antagonist, has been shown to have antioxidant activity in vitro, but its effect on in vivo oxidation has not been assessed. The aim of this study was to investigate the antioxidant effect of this agent in vivo. The ratios of 7-keto cholestadien to cholesterol are believed to be an available marker of lipid peroxidation. Using these ratios, we have assessed the antioxidant effect of nilvadipine on low-density lipoprotein (LDL) in hypertensive patients with high risk of atherosclerosis. The risk factors of atherosclerosis may involve oxidation of LDL. METHODS: Fifteen healthy subjects (seven females and eight males aged 35-72 years, mean +/- SD = 55.3 +/-13.8 years) and fifteen hypertensive patients (seven females and eight males aged 45 80 years, mean +/- SD = 60.2 +/- 11.8 years) were recruited. Patients were treated orally with nilvadipine (4 mg b.i.d.) for 4 weeks. Cholesterol oxidation levels of LDL in patients before and after nilvadipine therapy and healthy subjects were studied. RESULTS: The ratios of 7-keto cholestadien to cholesterol in LDL of hypertensive patients before and 4 weeks after nilvadipine treatment and in healthy subjects were 6.5 +/- 1.6% (mean +/- SD), 3.8 +/- 1.2%, and 0.2 +/ 0.1%, respectively. There were significantly (P < 0.001) increased levels of cholesterol oxidation in LDL in patients with hypertension both before and after nilvadipine treatment compared with healthy subjects. However, there was a significantly (P < 0.001) decreased level of cholesterol oxidation in LDL in patients after nilvadipine treatment compared with patients before nilvadipine treatment. CONCLUSION: Our data showed that nilvadipine may protect LDL cholesterol from in vivo oxidation in hypertensive patients with high risk of atherosclerosis. PMID- 10853876 TI - Comparison of several approaches of therapeutic drug monitoring of cyclosporin A based on individual pharmacokinetics. AB - OBJECTIVE: The clinical outcome of patients after organ transplantation is correlated with cyclosporin A (CyA) exposure. It is generally accepted that the area under the concentration-time curve (AUC) provides a reliable means for drug exposure. However, in routine therapeutic drug monitoring (TDM) of CyA, trough levels are mostly used. Currently, a number of different new concepts of CyA-TDM, including approaches such as single, double or triple time-point and abbreviated AUC determinations, have been introduced. The purpose of this study was to compare the predictive value of the different strategies of TDM. METHODS: Calculations were based on 40 individual concentration time profiles after oral administration of CyA to patients who had been included into an ongoing prospective clinical trial. Non-compartmental analysis was used to calculate the AUC0-12h. Multiple linear regression was performed to describe the relationship between the different sets of blood concentrations and the respective AUC0-12h as well as to evaluate their predictive value regarding AUC. Predictive performance was assessed by prediction bias and prediction precision, which were estimated as the mean prediction error and root mean squared error, respectively. RESULTS: When comparing the various combinations of time points, it was found that one point approaches showed the strongest differences with regard to the predictive value; the associated r2 values differed from 0.203 to 0.792. The two and three time-point approaches showed lower differences - r2 0.802-0.972. The four-point and five-point approaches (r2 0.942-0.982) were the strongest predictors for CyA AUC0-12h. Relative bias ranged from -27.7% to 63.8% and changed significantly when multiple-point predictors were used. In those cases, the predictive performance improved. Considering the predictive performance as well as the smallest bias and highest prediction precision, C3, C1 + C3, C1 + C3 + C6 and C1 + C2 + C3 + C6 were the best predictors. CONCLUSION: The results of this study indicate that in kidney transplant patients a clinically sufficient precise estimation of the CyA AUC is possible using two or three concentration time points. PMID- 10853877 TI - Pharmacokinetics of intravenous ATP in cancer patients. AB - OBJECTIVE: To characterise the pharmacokinetics of adenosine 5'-triphosphate (ATP) in patients with lung cancer after i.v. administration of different ATP dosages. METHODS: Twenty-eight patients received a total of 176 i.v. ATP courses of 30 h. Fifty-two infusions were given as low-dose infusions of 25-40 microg kg( 1) min(-1), 47 as middle-dose infusions of 45-60 microg kg(-1) min(-1) and 77 as high-dose infusions of 65-75 microg kg(-1) min(-1) ATP. Kinetic data of ATP concentrations in erythrocytes were available from 124 ATP courses. Results are expressed as mean +/- SEM. RESULTS: Most ATP courses in cancer patients were without side effects (64%), and side effects occurring in the remaining courses were mild and transient, resolving within minutes after decreasing the infusion rate. Baseline ATP concentration in erythrocytes was 1,554 +/- 51 micromol l(-1). ATP plateau levels at 24 h were significantly increased by 53 +/- 3, 56 +/- 3 and 69 +/- 2% after low-dose, middle-dose and high-dose ATP infusions, respectively. At the same time, significant increases in plasma uric acid concentrations were observed: 0.06 +/- 0.01, 0.11 +/- 0.01 and 0.16 +/- 0.01 mmol l(-1), respectively. The mean half-time for disappearance of ATP from erythrocytes, measured in five patients, was 5.9 +/- 0.5 h. CONCLUSIONS: During constant i.v. infusion of ATP in lung cancer patients, ATP is taken up by erythrocytes and reaches dose-dependent plateau levels 50-70% above basal concentrations at approximately 24 h. PMID- 10853878 TI - The effect of itraconazole on the pharmacokinetics and pharmacodynamics of oral prednisolone. AB - OBJECTIVE: To examine the possible effect of itraconazole on the pharmacokinetics and pharmacodynamics of orally administered prednisolone. METHODS: In this double blind, randomised, two-phase cross-over study, ten healthy subjects received either 200 mg itraconazole or placebo orally once a day for 4 days. On day 4, 20 mg prednisolone was given orally. Plasma concentrations of prednisolone, cortisol, itraconazole, and hydroxyitraconazole were determined by means of high performance liquid chromatography up to 47 h. RESULTS: Itraconazole increased the total area under the plasma prednisolone concentration-time curve by 24% (P < 0.001) and the elimination half-life of prednisolone by 29% (P < 0.001) compared with placebo. The peak plasma concentration and time to the peak of prednisolone were not affected by itraconazole. The mean morning plasma cortisol concentration, measured 23 h after the ingestion of prednisolone, during the itraconazole phase was 73% of that during the placebo phase (P < 0.001). CONCLUSIONS: The observed minor interaction between itraconazole and oral prednisolone is probably of limited clinical significance. The susceptibility of prednisolone to interact with CYP3A4 inhibitors is considerably smaller than that of methylprednisolone, and itraconazole and probably also other inhibitors of CYP3A4 can be used concomitantly with prednisolone without marked changes in the effects of this corticosteroid. PMID- 10853879 TI - Multiple-dose pharmacokinetics of cefepime in long-term hemodialysis with high flux membranes. AB - OBJECTIVE: Among uremic patients on hemodialysis, infectious complications leading to a high incidence of morbidity and mortality are a well-documented problem. In this multi-dose study, the safety, tolerance, and pharmacokinetics of cefepime during high-flux hemodialysis were investigated and an improved dosing schedule is presented. METHODS: Six long-term hemodialysis patients received 2 g cefepime i.v. at the end of hemodialysis three times per week. RESULTS: Trough levels of cefepime were 23.3 +/- 7.3 mg/l and peak serum concentrations 165.6 +/- 48.7 mg/l. After 3.5 h of high-flux hemodialysis, 72.2 +/- 6.4% of cefepime was eliminated. The intradialytic half-life was 1.6 +/- 0.29 h and the interdialytic half-life 22.0 +/- 2.14 h. CONCLUSION: A dosage of 2 g cefepime after each hemodialysis session achieved drug levels well above the minimal inhibitory concentration (MIC)90 for most of the target pathogens. Thus, the described dosing schedule is an efficient and cost saving antmicrobial therapy for severe infections in long-term hemodialysis patients with no residual renal function. PMID- 10853880 TI - Lack of differences in diclofenac (a substrate for CYP2C9) pharmacokinetics in healthy volunteers with respect to the single CYP2C9*3 allele. AB - OBJECTIVES: Evidence exists to suggest that diclofenac is metabolised by CYP2C9. The present study was undertaken in order to evaluate the effect of the single CYP2C9*3 variant on drug metabolism using diclofenac as a probe drug. METHODS: A single dose of diclofenac was administered orally to 12 healthy subjects in whom the genotype of CYP2C9 had been determined previously. The disposition kinetics of diclofenac were compared between homozygotes for the wild type (CYP2C9*1/*1, n = 6) and heterozygotes for the Leu359 variant (CYP2C9*1/*3, n = 6). RESULTS: For diclofenac, the following kinetic parameters were observed in the CYP2C9*1/*1 and CYP2C9*1/*3 subjects, respectively (mean +/- SD): apparent oral clearance (ml/kg/h) 355.8 +/- 56.9 and 484.4 +/- 155.3; area under plasma concentration time curve (microg h/ml) 2.7 +/- 0.7 and 1.9 +/- 0.6. The formation clearance of 4'-hydroxydiclofenac (ml/kg/h) was 63.6 +/- 19.1 in the CYP2C9*1/*1 subjects compared with 75.9 +/- 27.6 in the CYP2C9*1/*3 subjects. There were no significant differences in any of the kinetic parameters for either diclofenac disposition or formation clearance of 4'-hydroxydiclofenac between the two genotype groups. CONCLUSION: Since the disposition kinetics of diclofenac does not change in subjects with the single CYP2C9*3 mutant allele, it is suggested that the effects of CYP2C9 polymorphisms on the drug metabolism tend to be substrate specific. PMID- 10853881 TI - Influence of renal function on the pharmacokinetics of cerivastatin in normocholesterolemic adults. AB - OBJECTIVE: The influence of impaired renal function on the pharmacokinetics of single and multiple doses of cerivastatin was evaluated in this nonrandomized, non-blinded, 7-day, multiple-dose study. METHODS: Thirty-five adults between the ages of 21 years and 75 years with normal renal function (CL(CR) >90 ml/min/1.73 m2, n = 9), or patients with either mild (CL(CR) 61 ml/min/1.73 m2 to < or =90 ml/min/1.73 m2, n = 9), moderate (CL(CR) 30 ml/min/1.73 m2 to < or =60 ml/ min/1.73 m2, n = 8), or severe (CL(CR) <30 ml/min/ 1.73 m2, but not on dialysis, n = 9) renal impairment were given cerivastatin 0.3 mg daily each evening for 7 days. The steady-state pharmacokinetics of cerivastatin, including the area under the concentration-time curve (AUC)0-24, peak plasma concentration (Cmax), time to reach Cmax (tmax) and elimination half-life (t1/2), were determined on day 1 and day 7. The logarithm of the pharmacokinetic variables was analyzed using analysis of variance (ANOVA). Safety assessments included physical examination, fundoscopy, vital signs, electrocardiogram (ECG), adverse events, and laboratory safety indices. RESULTS: The day-1 AUC in patients with mild renal impairment was similar to that of patients with normal function (19.6 microg/h/l vs 19.2 microg/h/l, respectively). However, the AUC for cerivastatin patients with moderate or severe renal impairment was 40-60% higher (30.8 microg/h/l and 29.0 microg/h/l, respectively). Cmax values for patients with normal, mild, moderate, and severe renal impairment were 3.3, 3.4, 4.6, and 5.2 microg/l, respectively. This modest increase in plasma cerivastatin levels is nearly equivalent to a 0.4 mg daily dose, which has been recently approved in the United States. The mean t1/2 of cerivastatin was less than 4.5 h in all patients, indicating that renal dysfunction did not promote cerivastatin accumulation. This observation was confirmed by the finding that the cerivastatin plasma levels on day 1 and day 7 were similar in all patient groups. Furthermore, the mean AUC and Cmax values for both demethylated and hydroxylated cerivastatin were similar in the patients with the most severe renal dysfunction to the corresponding values in healthy subjects. Cerivastatin was well tolerated in all patients irrespective of renal function. Adverse events were observed in 37% of the subjects; nearly all were mild and generally of short duration, and most resolved without intervention. Incidence of adverse events was similar across all three renal groups and the control group. There were no clinically significant laboratory changes other than those consistent with renal disease. CONCLUSION: This study demonstrates that dosage adjustment of the daily 0.3-mg cerivastatin dose in patients with significant renal impairment is likely unnecessary. PMID- 10853882 TI - Pharmacokinetics of propranolol and atenolol in patients after partial gastric resection: a comparative study. AB - OBJECTIVE: Partial gastric resection alters the anatomy and secretory activity of the gastrointestinal tract. It might be expected that the consequences of such changes should affect the pharmacokinetics, especially concerning the absorption of orally administered drugs. Propranolol and atenolol, as representatives of lipophilic and hydrophilic beta-adrenoreceptor antagonists, have been studied in order to define their pharmacokinetic characteristics in patients after partial gastrectomy. METHODS: The study was carried out in 29 patients after gastric resection with Billroth I (B1) anastomosis and in 18 healthy volunteers as controls. Pharmacokinetics of propranolol and atenolol was investigated after a single oral dose of 80 mg and 100 mg, respectively, following a cross-over schedule. Blood samples were collected ten times during the 24 h after the drug administration. Pharmacokinetic parameters of propranolol and atenolol were calculated using a one-compartment open model with first-order absorption. RESULTS: The average blood plasma concentrations of propranolol in gastrectomised patients were lower than those in controls, reaching significance between 1.5 h and 6.0 h of the observation period. Pharmacokinetic parameters of propranolol were different in subjects submitted to surgery compared with healthy persons. We observed a significant decrease in the area under the concentration-time curve (32%) and the peak plasma concentration (20%), and an increase in half-life (25%). Mean plasma concentrations and pharmacokinetic parameters of atenolol in patients following partial gastric resection were not significantly different from those in the controls. No relationship between time interval following partial gastrectomy and pharmacokinetic parameters of the investigated drugs was noted. CONCLUSION: Partial gastrectomy with B1 anastomosis affects the pharmacokinetics of propranolol (lipophilic drug) but not atenolol (hydrophilic drug). PMID- 10853883 TI - Inhibition of human liver phenol sulfotransferase by nonsteroidal anti inflammatory drugs. AB - OBJECTIVE: The aim of this investigation was to study the inhibition of 11 nonsteroidal anti-inflammatory drugs (NSAIDs) on the human liver phenol sulfotransferases (HL-PST) and catechol sulfotransferase (HL-CST). METHODS: The activities of HL-PST and HL-CST were measured with 4 microM 4-nitrophenol and 60 microM dopamine (the sulfate acceptors) and 0.4 microM 3'-phosphoadenosine-5' phosphosulfate [35S] (the sulfate donor). Samples of liver were obtained from five patients, aged 55-79 years, undergoing clinically indicated hepatectomy. The inhibition curves were constructed with at least five concentrations of the inhibitor. RESULTS: With the exception of piroxicam, NSAIDs inhibited HL-PST, and the estimates of the inhibitory concentration for 50% of responses (IC50; microM) were: 0.02 (mefenamic acid), 3.7 (diflunisal), 5.4 (nimesulide), 9.5 (diclofenac), 30 (salicylic acid), 41 (ketoprofen), 74 (indomethacin), 159 (ibuprofen), 245 (ketoralac) and 473 (naproxen). With 4-nitrophenol as the variable substrate, the inhibition of salicylic acid on HL-PST was non competitive and the Ki and Kies were 18 microM and 21 microM (n = 5; P = 0.548), respectively. HL-CST was less susceptible than HL-PST to inhibition by NSAIDs, with only five drugs inhibiting this enzyme. The IC50 estimates for these drugs (microM) were 76 (mefenamic acid), 79 (diflunisal), 103 (indomethacin), 609 (salicylic acid) and 753 (diclofenac). CONCLUSION: The comparison of the IC50 estimates of HL-PST with the therapeutic plasma concentrations of NSAIDs corrected for the plasma unbound fraction was consistent with the view that mefenamic acid and salicylic acid, when administered at therapeutic doses, should impair the hepatic sulfation of those compounds that are substrates of phenol sulfotransferase. PMID- 10853884 TI - Pharmacokinetics and pharmacodynamics of propofol 6% SAZN versus propofol 1% SAZN and Diprivan-10 for short-term sedation following coronary artery bypass surgery. AB - OBJECTIVE: A new formulation of propofol 6% in Lipofundin MCT/LCT 10% (propofol 6% SAZN) has been developed in order to reduce the fat, emulsifier and volume load that is given during prolonged infusions of propofol. The pharmacokinetics, pharmacodynamics and safety characteristics of propofol 6% SAZN were investigated during a short-term infusion and compared with the commercially available product propofol 1% in Intralipid 10% (Diprivan-10) and propofol 1% in Lipofundin MCT/LCT 10% (propofol 1% SAZN). METHODS: In a randomised double-blind study, 24 male patients received a 5-h infusion of propofol at the rate of 1 mg/kg/h for sedation in the immediate postoperative period following coronary artery bypass surgery. RESULTS: The average pharmacokinetic parameter estimates of clearance (Cl), volume of distribution at steady state (Vd,ss), elimination half-life (t1/2,beta) and distribution half-life (t1/2,alpha) observed in the three groups were 28 +/- 1.1 ml/kg/min, 1.8 +/- 0.12 l/kg, 94 +/- 4.1 min and 3.1 +/- 0.26 min, respectively (mean +/- SEM, n = 24) and no significant differences were noted between the three formulations (P > 0.05). In one patient receiving propofol 6% SAZN, in two patients receiving propofol 1% SAZN and in three patients receiving Diprivan-10, the level of sedation was inadequate and additional sedative medication had to be given. In all other 18 patients, the level of sedation was adequate. The mean propofol concentration in these six inadequately sedated patients was lower than the adequately sedated patients (P = 0.015). The serum triglyceride concentrations were not significantly different between the groups studied. No adverse events occurred in any of the patients. CONCLUSIONS: The pharmacokinetics, pharmacodynamics and safety characteristics of propofol 6% SAZN are in good agreement with those of the 1% formulations. Propofol 6% SAZN therefore provides a useful alternative to the commercially available 1% formulation for short-term sedation in the intensive care unit. Expected advantages in long-term sedation of the 6% over 1% formulation are the subject of an ongoing study. PMID- 10853885 TI - Antibiotic utilization at the university hospital after introducing an antibiotic policy. AB - OBJECTIVE: Antibiotic formulary restrictions are among the most popular methods to control antibiotic utilization in hospitals. The aim of the present survey was to investigate the influence of "reserve antibiotic" on antimicrobial utilization at the University Hospital Center (UHC) Rijeka. METHODS: At the UHC Rijeka, reserve antibiotic was implemented in July 1997. The antimicrobial drug consumption was monitored 6 months prior to and 6 months after the introduction of the method. Antimicrobial consumption was measured in defined daily doses (DDDs) among the major clinics. RESULTS: Reserve antibiotic has led to a decrease in total antibiotic consumption at the UHC Rijeka (45.9 DDDs/100 bed days vs 32.9 DDDs/100 bed days). Antibiotic utilization decreased in the second semester at most clinics: at the Clinic for Infectious Diseases 41%, at the Anesthesiology and Intensive Care Unit 30%, at the Clinic for Internal Medicine 18% and at the Surgical Clinic 12%. At the Clinic for Gynecology and Obstetrics, the antibiotic utilization remained the same, while at the Pediatric Clinic an increase of 28% in antibiotic utilization was noted. CONCLUSION: Our study indicates that restriction of usage of some antibacterial agents is a successful method to decrease antibiotic consumption and a way to bring cost savings and helps prevent emergence of resistant microorganisms in hospitals. To improve antimicrobial prescribing, additional methods such as education are required. PMID- 10853886 TI - Workplace drug testing (WDT) likely to increase in Europe. Report from the First European Symposium on WDT including selected abstracts. AB - Will it take a series of drug-related accidents that have already occurred in the USA before workplace drug testing (WDT) becomes accepted in Europe as a preventive measure? Currently, the development of WDT in most European countries lags some 10-15 years behind that in the USA. Labour authorities in Europe now ought to take initiatives to demand a mandatory programme for accrediting drug analytical laboratories for WDT. Companies should realise that illicit drug use is no longer only a problem at street corners, and that having a testing system in place is important, not just for public health, but also for their reputations as responsible societal actors. Improved networking among police and regulatory authorities is required to keep pace with the rapid appearance and dissemination of new substances of abuse. European research collaboration, including the newly formed European Workplace Drug Testing Group, is needed to assess the impact of drug-testing policies on accidents and other outcome variables, and thereby to convince the general public and politicians that drug testing is beneficial and necessary. A 1993-1994 survey of quality analysis in some 200 European laboratories reported from Institut Municipal d'Investigacio Medica (IMIM), Spain, showed good agreement between nominal and found concentrations but that only 10% of the laboratories could both screen, identify and quantify samples. Experiences from Italy show that proficiency testing schemes lead to improved accuracy of results. These were some major conclusions of the First European Symposium on Drug Testing held at Huddinge University Hospital in Stockholm, Sweden, 30 March to 1 April 1998, organised by Karolinska Institute, with participants from 22 countries. PMID- 10853887 TI - Evaluation of core decompression for early osteonecrosis of the femoral head. AB - Twenty-six hips (19 patients) with osteonecrosis of the femoral head with stage I or II of the disease, according to the Ficat and Arlet classification, underwent core decompression. Osteonecrosis was confirmed histologically in all 26 hips. Of 19 patients, 7 had prognostic factors traditionally associated with poor outcome including collagen vascular disease and continued use of steroids. The follow-up period averaged 7 years 10 months (range: 2 years 5 months-13 years 8 months) for 17 patients with 24 hips. Two patients died secondary to systemic illness. Seventeen hips (65.4%) had very good or good results using the Ficat criteria. Eight hips (30.8%) needed further operation [total hip arthroplasty (THA) for 7 hips, osteotomy for 1 hip]. Of the 12 hips in patients who had used steroids, 6 hips (50%) were converted to THA. Four hips in patients with systemic lupus erythematosus (SLE) needed THA (100%). We conclude that core decompression provides an effective treatment for steroid-associated osteonecrosis other than in cases with SLE, as well as providing effective treatment for non-steroid associated osteonecrosis in the early stages of the disease. PMID- 10853888 TI - Diagnostic and therapeutic management of lumbar and thoracic spondylodiscitis--an evaluation of 59 cases. AB - Fifty-nine patients with spondylodiscitis (SD) of the thoracic and/or lumbar spine were followed-up clinically and radiologically [X-ray, computed tomography (CT), magnetic resonance imaging (MRI)] over a mean time of 2.2 years (1-6.5 years). All patients without abscess formation (n = 35) were treated conservatively. Out of the group with abscess formation (n = 24) 6 patients were also treated conservatively, 11 were drained under CT control and 7 were operated. At time of diagnosis, "signs of florid inflammation" were seen in 60% of the roentgenograms, in 93% of the CTs and in all of the MRls. The sensitivity to differentiate between SD with and without abscess formation was 85% by MRI and 69% by CT. "Signs of regressive inflammation" and "signs of increasing osseous consolidation", essential facts for starting remobilization, could first be seen using CT 6 weeks after onset of therapy. Using MRI these signs were seen with a considerable delay at 12 weeks. Clinically, only 3 of the 59 analyzed patients developed recurrent SD. In conclusion, MRI is the radiological method of choice for establishing the diagnosis of SD, in particular with regard to differentiating between cases with and without abscess formations. In contrast, CT is superior for performing success control after treatment. Therapeutically, conservative, minimal-invasive and operative procedures are not rival but rather complementary. PMID- 10853889 TI - Therapeutic effect of transtrochanteric rotational osteotomy and hip arthroplasty on quality of life of patients with osteonecrosis. AB - We reviewed 37 patients with avascular necrosis of the femoral head (ANF). There were 23 men and 14 women with a mean age of 36 years at the time of the operation. The duration of follow-up was 9 years. Twenty patients had undergone transtrochanteric rotational osteotomy (TRO) and 17, hip arthroplasties. Assessment of their quality of life (QoL) was performed using the Rosser Index Matrix for disability and distress. Concerning TRO, the mean preoperative and postoperative QoL scores were 0.944 and 0.957, respectively. Twelve patients exhibited increases and 7 patients decreases in their scores. Regarding the arthroplasty, the mean preoperative and postoperative QoL scores were 0.949 and 0.998, respectively. All patients showed increases in QoL scores after arthroplastic surgery. Concerning heavy manual work, all five of those patients returned to their preoperative occupations. These findings suggest that hip arthroplasty has more reliable therapeutic effects than TRO on QoL improvement for patients with ANF. PMID- 10853890 TI - Rotational profile of the lower extremity and foot progression angle: computerized tomographic examination of 50 male adults. AB - Acetabular, femoral and tibial torsion of 50 normal adult male subjects were measured by computerized tomography and the relationship between these angles and foot-progression angle was examined. The mean acetabular anteversion was 15.6 degrees on the right and 15.8 degrees on the left, (range 3 degrees-30 degrees). The mean femoral torsion was 6.5 degrees on the right and 5.8 degrees on the left (range 14 degrees-28 degrees). The mean tibiofibular torsion was 30.9 degrees on the right and 29.1 degrees on the left (range 16 degrees-50 degrees). Although the normal range of torsional measurements of the lower extremity was very broad, subjects usually had out-toeing, with a mean foot-progression angle of 13.7 degrees on the right and 13.0 degrees on the left (range 6 degrees-21 degrees). No correlation was detected on the rotation between different levels of the lower limb. No difference was detected in the lower extremity rotational profile between right and left sides. PMID- 10853891 TI - Bipolar versus total hip endoprosthesis: functional results. AB - Some functional parameters of the hip-joint 3.3 years on average (range 2-8.6 years) after hip arthroplasty are compared with regard to two types of hip endoprostheses: the total (TEP) and the bipolar (BPEP). Flexion, extension, abduction, adduction, and internal and external rotation were measured for 75 patients with BPEP type Self-Locking, for 11 patients with BPEP type Vario-Cup, and for 43 with TEP type Lubinus. One-way analysis of variance with respect to co variances was used for statistical testing of the measured data. It was found that flexion, abduction and adduction were significantly higher in the BPEP endoprosthesis (P < 0.05). The results obtained are favourable for the bipolar hip endoprosthesis, and they can be related to the biomechanical differences between both types of hip endoprosthesis. PMID- 10853892 TI - Solitary osseous hemangioma outside the spinal and craniofacial bones. AB - Bone hemangioma is mainly seen in the skull and spine, and rarely occurs in other bones. We report herein four cases of osseous hemangioma arising in rare sites: In two cases, on a rib; a faintly painful mass in one case located on the scapula; and progressive pain in one case located on the ischium. The tumors presented clinically as incidental lesions on radiographs. All cases had an aggressive appearance, such as defect of the cortex, a soft-tissue mass, and a sunburst-like appearance. Markedly high signal intensity on T2-weighted magnetic resonance images was a characteristic finding. Open biopsy resulted in severe blood loss, but needle biopsy was performed safely under computed tomography guidance. It is important to note that bone hemangiomas may be misdiagnosed as malignant tumors. PMID- 10853893 TI - The influence of obesity on perioperative morbidity and mortality in revision total hip arthroplasty. AB - The significance of obesity as a risk factor for postoperative complications was determined in a consecutive series of 229 cases of revision total hip replacement. The body mass index (BMI) was used as an objective measure to classify the patients. The group-wise analysis of data included all medical and procedure-related complications, the number of fatal cases, operative time, requirement for analgesics, the number of transfusions and perioperative haemoglobin levels. The results of our study demonstrate a clear association between obesity and operative time, whereas no statistically significant relationships were observed between obesity and the other parameters. We conclude that obesity does not have any significant influence on perioperative morbidity and mortality but is clearly related to operation time and, therefore, to higher costs per operation. PMID- 10853894 TI - Biomechanical considerations in 'biological' femoral osteosynthesis: an experimental study of the 'bridging' and 'wave' plating techniques. AB - Although traditional compression plate fixation aims to abolish interfragmentary movement and achieve primary bone healing, the more recent 'biological' plate fixation methods such as the 'bridging' and wave' plate techniques aim to maintain fracture alignment without absolute stability and promote union by callus formation. Furthermore, some mechanical advantages have been attributed to the 'wave' plate fixation. Since no data have been published on the mechanical characteristics of the 'bridging' and 'wave' plate fixation methods, the aim of this biomechanical comparative study was to investigate the rigidity of those fixation methods in various types of femoral diaphyseal fractures. Using a composite femoral model, the rigidity characteristics of three fixation methods (short DCP, 'bridging' and 'wave' plates) were investigated. The results showed that when cortical contact between the main fragments is present, a 'bridging' plate can be equally rigid to the 'wave' plate in mediolateral bending by displaying a similar tension-band effect. Furthermore, in the absence of cortical contact, the axial fixation rigidity of the long 'bridging' plate is superior to that of the 'wave' plate. Both methods showed a significant 'stress-shielding' effect on the intact femur. In conclusion, this in vitro study failed to show any significant mechanical advantages of the 'wave' plate technique over the 'bridging' plating method. It appears that the 'bridging' plate fixation may be the mechanically optimal 'biological' plating method for the femoral diaphysis. PMID- 10853895 TI - Cemented Lubinus and Furlog total hip endoprosthesis: a 12-year follow-up study of 175 hips comparing the cementing technique. AB - We analyzed 175 total hip replacements with cemented Lubinus and Furlong arthroplasties in 164 patients with a median age of 65 (32-80) years and followed them for 12 years to evaluate and compare the efficacy of total hip prosthesis designs. Survival analysis was combined with an analysis of radiological findings and a study of functional outcome of the patients. The 12-year survival of Furlong arthroplasty in patients of 60 years of age and older was O.85 (95% CI 1.00-0.52). The survival of Lubinus arthroplasty in patients younger than 60 years of age was 0.70 (0.91-0.48), while the survival in older patients was 0.75 (0.89-0.61). The 12-year survival of well-cemented Lubinus prosthesis was 0.91 (1.00-0.79), indicating the importance of the cementing technique. The survival of the cups was marginally better than that of the stems. In the 12-year follow up study, the clinical state and function varied from hips ready for revision to hips where a continuously long survival could be predicted. Harris hip score did not differentiate between patients who had intact and loose components. We conclude that cemented arthroplasty affords a notable alternative with satisfactory long-term survival and function. The better survival of cemented cup than the stem may be utilized as a basis for "reverse" hybrid arthroplasty. Adequate long-term follow-up of all arthroplasties as a quality maintenance and to prevent difficult revisions is a major challenge. PMID- 10853896 TI - Functional results of braced humeral diaphyseal fractures: why do 38% lose external rotation of the shoulder? AB - A total of 67 humeral diaphyseal fractures treated with functional bracing was studied. The median follow-up was 30 weeks. Sixty-one fractures (91%) healed and 6 fractures (8.9%) progressed to non-unions. Fifty-four fractures could be functionally classified according to a modified Wasmer score. Pain, range of motion in the shoulder and elbow, and changes in activities of daily life were recorded. Loss of external rotation in the shoulder was most prominent, being present in 21 (38%) of the fractures. To evaluate the cause of loss of external rotation, 21 of the patients were selected for two groups to be studied with computed tomography (CT). Twelve patients had normal clinical findings without a loss of external rotation, while 9 patients had subnormal external rotation in the shoulder of the injured limb. Fracture consolidation in malrotation was seen frequently, and a linear correlation between the clinical loss of external rotation and CT findings was indicated, but no statistical agreement could be proved. The time between injury and brace application could possibly contribute to consolidation in malrotation. PMID- 10853897 TI - The changes occurring after the Putti-Platt procedure using magnetic resonance imaging. AB - The purpose of this study is to evaluate the magnetic resonance imaging (MRI) following Putti-Platt procedure for recurrent anterior dislocation of the shoulder. Six shoulders of six patients who had received Putti-Platt procedure were evaluated by the MRI before and after operation. After the Putti-Platt procedure the subscapularis tendon was thickened and an increased signal area on T2-weighted images were observed in four patients. The area of subscapularis tendons after operation was increased maximally 3.46-fold and the volume was increased on average 1.51-fold. The course of subscapularis muscle fiber before operation was described as a mild arc, but changed to a straight line after the procedure in five patients. The findings in this study suggest that the Putti Platt procedure leads to a remarkable increase in strength of subscapularis tendon and an improvement of laxity of subscapularis muscle. In conclusion, there is a good possibility that this procedure will increase the stability of the glenohumeral joint and be a successful treatment for recurrent anterior dislocation of the shoulder. PMID- 10853898 TI - Early recovery of isometric shoulder muscle strength after open acromioplasty in stage II impingement syndrome. AB - The recovery of shoulder muscle strength after open acromioplasty was evaluated in 48 patients (27 male, 21 female, mean age 44.3 years) who had undergone open acromioplasty because of stage II impingement syndrome. The isometric strengths of flexion, abduction and external rotation were measured before the operation and at 3, 6 and 12 months postoperatively. The mean preoperative flexion strength of the involved shoulder was 72.6% of that of the uninvolved shoulder, and this increased to 77.1% by 3 months post operation, to 88.3% at 6 and to 88.3% at 12 months. Corresponding abduction strengths were 68.4%, 80.4%, 88.7% and 91.0% and the external rotation strengths were 75.1%, 77.4%, 95.1% and 93.5%, respectively. These recoveries were markedly improved when the cases with poor subjective results at 1 year were not considered. The low preoperative strengths were more pronounced in women than in men, but recovery was better in women. It is concluded that shoulder muscle strengths recover to near normal in 1 year after open acromioplasty. PMID- 10853899 TI - Short- and medium-term results of the thrust plate prosthesis in patients with polyarthritis. AB - The thrust plate prosthesis is an implant with metaphyseal fixation to the proximal femur, which leaves the diaphyseal bone untouched. Therefore, this implant is preferred in younger patients. It is dependent on good bone quality in the proximal femur. Because the bone quality is reduced in patients with polyarthritis, this kind of endoprosthesis may have a higher failure rate than conventional stemmed endoprostheses in these patients. Therefore, in patients with polyarthritis, even short- and medium-term results of the thrust plate prosthesis should be analyzed. In all, 47 thrust plate prostheses were implanted in 42 patients with polyarthritis (29 with rheumatoid arthritis, 6 with juvenile chronic arthritis, and 7 with spondylarthritis) and followed prospectively. The average age at operation was 40.8+/-10.7 years. Each patient was clinically and radiologically examined preoperatively, 3 and 6 months after the operation, and at the end of each postoperative year. The mean follow-up was 26.1+/-10.7 months. The clinical findings were evaluated using the Harris hip score. Radiologically, 8 different zones of the thrust plate prosthesis were analyzed for radiolucencies. During the 1st year, the Harris hip score rose continuously from the preoperative average of 42.4+/-6.5 points to 78.8+/-10.3 points 3 months postoperatively, 82.3+/-9.8 points 6 months postoperatively, and 86.8+/-10.1 points 1 year after the operation. The subsequent examinations showed Harris hip score remained at the same level. Five patients (5 joints, 10.6%) had to undergo a revision of the thrust plate prosthesis due to aseptic loosening in 3 and septic loosening in 2. Six prostheses (12.6%) showed radiolucencies, mostly below the thrust plate in zones 1 and 2. Two of them were certainly radiologically loose, which raised the failure rate to 7 of 47 (14.8%). The thrust plate prosthesis improves function and alleviates pain in patients with polyarthritis to a satisfactory degree. Concerning the failure rate, this type seems to yield slightly worse results than cementless stemmed endoprostheses in the same patient group. Due to the preservation of the diaphyseal bone of the femur and the possibility of an unproblematic change to a stemmed endoprosthesis, the thrust plate prosthesis can be recommended for younger patients with polyarthritis. PMID- 10853900 TI - Is there still an indication for operative treatment of femoral neck fractures with a ceramic hemiprosthesis? Four-year results. AB - From 1985 to 1995, 417 patients with dislocated medial femoral neck fractures (Garden III-IV) were treated with hemiarthroplasty using a Biolox ceramic head. The average patient age at the time of operation was 81.5 years. A total of 140 survivors was available for follow-up examination with a mean prosthesis longevity of 55.8 months. The Harris hip score recorded a mean of 70.6 points. At the time of follow up, 5 patients had severe hip pain, and in 8 the roentgenographic examination revealed protrusio acetabuli. Five of these 8 patients underwent revision surgery for replacement of the cup, leaving the stem in situ. PMID- 10853901 TI - Extracorporal shock wave therapy in patients with tennis elbow and painful heel. AB - The aim of this study was to evaluate the effect of extracorporal shock wave therapy (ESWT) in tennis elbow and painful heel. Nineteen patients with tennis elbow and 44 patients with painful heel in which conservative treatment had failed underwent ESWT. Both groups received 3000 shock waves of 0.12 mJ/mm2 three times at weekly intervals. After a follow-up of 5 and 6 months respectively, pain measured on a visual analogue scale (VAS) decreased significantly in both groups. The success rate (excellent and good results) was 63% in tennis elbows and 70% in painful heels. ESWT seems to be a useful conservative alternative in the treatment of both conditions. PMID- 10853902 TI - Quality assurance in hip arthroplasty. AB - Documentation is key to quality assurance (QA): Data must be complete, plausible, and comparable, and then analyzed to implement corrective measures. Important factors are: qualification of care-providing staff, equipment and implants available (structural quality), effective scheduling of operations and therapy management (process quality), and patient status monitoring (outcome quality). The primary aim is to reveal deficits in process quality and develop and implement improvements in care. QA does not aim at exposing individual mistakes or flawed techniques; rather it is designed to analyze processes and treatments and implement specific solutions. An evaluation profile with the key quality indicators and a QA guideline is presented. A survey conducted in Germany, Austria, and Switzerland revealed: (1) up to 12-month waiting period for surgery in 6%, (2) only 40% written instructions, (3) data mostly written by hand, (4) differences in surgery planning and use of prosthesis passport, (6) inconsistent data analysis, (7) corrective measures rarely implemented. PMID- 10853903 TI - Can skin surface pressure under a cast reveal intracompartmental pressure? AB - Although monitoring intracompartmental pressure (IP) under a cast is very important, it is not possible to measure it in every patient undergoing cast treatment. This study aims to answer the question of whether skin surface pressure (SSP) under a cast can reveal IP. A plaster cast was applied to a sculpted inflatable forearm model with dorsal and volar compartments. SSP under the cast was measured at five different localizations from both dorsal and volar sides of the model and compared to the corresponding IP. In the second experiment, a posterior tibial compartment syndrome model was created in both limbs of five rabbits. Correlation analysis was performed between IP and SSP under the cast. All of the SSP measurements taken from the dorsal and volar side of the sculpted forearm model correlated with IP. Mean correlation coefficient of the measurements was 0.995 (P = 0.000) (SD 0.002, range 0.992-0.999). SSP and IP correlation analysis in the posterior tibial compartment syndrome model of 10 limbs in five rabbits revealed a high correlation. The mean correlation coefficient was 0.973 (P = 0.000) (SD 0.024, range 0.916-0.997). Measuring the pressure between the skin and cast can monitor IP. SSP monitoring can help the physician, patient or parents in the follow-up of patients undergoing cast treatment. PMID- 10853904 TI - The initial fixation of the press-fit acetabular shell--clinical observation and experimental study. AB - The initial solid fixation of an uncemented acetabular component affects the amount of bone ingrowth. We had several problems with broken screws in cases of acetabular revision. In recent years, the development of uncemented components without screws has attempted to improve these problems. We started to use "press fit"-type acetabular shells in November 1996. Our thirty cases undergoing 2 mm under-reaming show good initial stability. The aim of our biomechanical study was to assess the most suitable degree of under-reaming of the bony acetabulum for the implantation of an uncemented hemispherical porous coated component. PMID- 10853905 TI - Interspinous distance in congenital dislocation of the hip. AB - We measured radiographically the interspinous distance in 50 normal children and in 50 children with congenital dislocation of the hip. The value increased proportionally with age in normal children, but not in those with congenital dislocation of the hip. Our results indicate that a bony parameter, rather than the age of the child, is suitable for the indication of lower age limit for Salter's innominate osteotomy. PMID- 10853906 TI - Increased interleukin-8 (IL-8) expression is related to aseptic loosening of total hip replacement. AB - Aseptic loosening is an increasing problem in total hip replacement (THR). Chronic inflammatory reaction against implant wear particle results in collageno- and osteolysis, leading to loosening of the implant. Cytokines are known to play a major role in this particular inflammatory process. The aim of the present study was to examine interleukin-8 (IL-8) in the synovial-like interface membrane (SLIM) and pseudocapsular tissue of THRs and to compare it to normal knee synovial membrane. Eleven patients suffering from aseptically loosened THRs were included. All the SLIM and pseudocapsular tissue samples were obtained during revision operations. Ten control samples of normal synovium were collected per arthroscopy from the superior recessus of the knee. For immunohistochemical IL-8 detection, polyclonal mouse anti-human immunoglobulin (Ig)G1 IL-8-primary antibody was used with the alkaline phosphatase anti-alkaline phosphatase (APAAP) method. Results were quantitated using the Vidas image analysis system. The highest count levels (mean +/- SEM) were detected in SLIM tissue (386+/-82 cells/mm2). The difference was statistically significant compared with pseudocapsular tissue (193+/-36 cells/mm2) and control samples (18+/-5 cells/mm2). Count levels in control tissue were on average 5% of the SLIM tissues values. The present study determines for the first time the cellular origin of IL 8 in aseptically loosened THRs and also quantitates the IL-8-producing cells in the periprosthetic tissue. The results reveal a high rise in IL-8 concentration in SLIM and in synovial tissues. This finding moves us one step forward in solving the complex network of multiple factors affecting loosening of hip implants. PMID- 10853907 TI - Bone remodelling in humeral arthroplasty: follow-up using CT imaging and finite element modeling--an in vivo case study. AB - Little material is available in the literature about remodelling of the human humerus after implantation of a shoulder hemiarthroplasty. A 73-year-old patient was examined by CT 4 years after implantation of a right shoulder hemiarthroplasty, and the bone density as represented by Hounsfield values was compared with the contralateral side. Additionally, a three-dimensional finite element model was generated from the image data and analysed. Bone density was reduced around the prosthesis when compared with the contralateral side. The stresses were transmitted through the prosthesis, while low bone stresses were found surrounding the prosthesis. Distally from the prosthesis, high stresses were found. On the control side, a more homogeneous stress distribution was noted. The results could be explained by bone resorption around the prosthesis caused by stress shielding; this hypothesis has to be confirmed by future studies. PMID- 10853908 TI - The treatment of pulled elbow: a prospective randomized study. AB - To evaluate the effectiveness in decreasing recurrence of cast application after manual reduction of pulled elbow. Sixty-four children with pulled elbow were randomized into two treatment groups: Group A underwent manipulative reduction followed by splinting the elbow in a flexed and supinated position for 2 days; group B underwent manipulative reduction only. Both groups were examined 2, 5, and 10 days later. None of the 33 patients in group A had a pulled elbow at follow-up. Four (13%) of 31 patients in group B had a pulled elbow 2-5 days later. Immobilizing the elbow for 2 days after manipulative reduction improves the success of treatment of a pulled elbow. PMID- 10853909 TI - Results of arthroscopic joint debridement in different stages of chondromalacia of the knee joint. AB - A retrospective study was performed of 161 patients who had undergone arthroscopic operation for chondromalacia of the knee joint. After an average follow-up period of 40 (range 10-72) months, patients with severe articular cartilage lesions who had undergone articular lavage alone showed significantly poorer results (P < 0.001). With the same stage of chondromalacia and having undergone the same surgical procedure, younger patients showed better results than older patients. The more effective interruption of the circulus vitiosus during the development of degenerative joint diseases is the primary cause for better results achieved by mechanical debridement of the joint for patients suffering from grade 2 or higher. According to the literature, aggressive subchondral abrasion in severely degenerated knees does not show any benefits. Apparently, the success of the therapy depends to a great extent on the inferiority of the potential degenerative regenerate (lack of capacity of intrinsic regeneration of the hyaline cartilage) as well as on the grade and the progression of chondromalacia. Almost every second patient suffering from grade 4 chondromalacia complained of recurrent pain 1 year postoperatively. One of every 6 patients received a knee joint prosthesis within the 1st year. Therefore, the patients' preoperative expectations have to be clearly objectified. The surgical procedure as an operation with a low complication risk can also be justified as a temporary alternative to total knee arthroplasty in patients suffering from a high-grade degeneration of the joint. PMID- 10853910 TI - Human leukocyte antigen (HLA) phenotypes in siblings with osteosarcoma. AB - Osteosarcoma was detected in two siblings. Their human leukocyte antigen (HLA) phenotypes were completely identical, although they were different from those of osteosarcoma patients in previous reports. Despite an extensive search of family and past history, no significant background related to the induction of cancer could be found. These cases suggest that genetic similarity may influence the development of osteosarcoma. Ascertainment of the HLA phenotypes in siblings with osteosarcoma might be a useful strategy to facilitate the early diagnosis of this tumor. PMID- 10853911 TI - Ossification of the yellow ligament causing thoracic cord compression. AB - Ossification of the yellow ligament (OYL) is not infrequent in the cervical and lumbar regions but is very rare in the thoracic spine, with no more than 40 cases reported in the literature. We describe a 50-year-old male with progressive paraparesis and sensory dysfunction, secondary to OYL at T10-T11, studied by computed tomography (CT) and magnetic resonance imaging (MRI). Decompressive laminectomy and removal of the ligament resulted in marked clinical improvement. Patients with OYL may initially develop sensory dysfunction associated with leg weakness. This pathological entity can be well defined by CT and MRI, and surgery by decompressive laminectomy is advised for all cases. The OYL should be removed both posteriorly and laterally to the dural sac to obtain sufficient decompression of the spinal canal. PMID- 10853912 TI - Familial occurrence of glenoid dysplasia: report of two cases in two consecutive generations. AB - Glenoid dysplasia is a rare abnormality of the shoulder. We report glenoid dysplasia in two consecutive generations: a boy and his father. Both suffered recurrent shoulder dislocations, and radiological examination revealed bilateral glenoid dysplasia. Our cases confirm dominant inheritance of this osseous malformation. PMID- 10853913 TI - Late sciatic nerve palsy following avulsion of the biceps femoris muscle from the ischial tuberosity. AB - A case of late sciatic nerve palsy following avulsion of the biceps femoris muscle from the ischial tuberosity in a 27-year-old athlete is reported. PMID- 10853914 TI - Occult fractures of tibial plateau detected employing magnetic resonance imaging. AB - We describe two cases of spontaneous fracture at the tibial metaphysis not diagnosed by standard X-ray. In both cases, only magnetic resonance imaging supplied a precise diagnosis and allowed us to follow the evolution of the pathology. Scintigraphy is equally sensitive but unspecific. Osteoporosis was noted in all cases. Hypothetically, similar pathological situations could be present without being diagnosed since they are not always detected by standard X rays. PMID- 10853915 TI - Multiple occurrence of osteochondromas in dysplasia epiphysealis hemimelica. AB - Dysplasia epiphysealis hemimelica was defined by Trevor (1950) as a rare congenital growth disorder of the tarsus and of the epiphysis of the long bone. In this report, a rare case of dysplasia epiphysealis hemimelica associated with multiple extraskeletal osteochondromas is presented. Although different modes of expression of the same pathologic process have been suggested for dysplasia epiphysealis hemimelica and osteochondroma, the biological feature of cartilaginous overgrowth in the skeletal system still seems unclear. PMID- 10853916 TI - Simultaneous double interphalangeal dislocation in one finger. AB - Isolated dislocation of the proximal or distal interphalangeal joint of a finger is common, but simultaneous dislocation of both joints is rare. Three cases of simultaneous dislocations of both interphalangeal joints in the same finger are reported. Closed reduction was easily achieved in all cases. PMID- 10853917 TI - Bilateral sternoclavicular joint tuberculosis. AB - A unique case of bilateral sternoclavicular tuberculosis is presented, with discussion of the possible mechanism of infection. Early diagnosis is mandatory for good results, and with a world-wide resurgence of this disease, a high index of suspicion is mandatory (especially in immunocompromised patients and migrant populations). Computed tomography and magnetic resonance imaging are helpful for defining the exact extent of the disease. PMID- 10853918 TI - Fatigue failure of an AO spiral blade. AB - We report an unusual case of a femoral neck stress fracture leading to the fatigue failure of an AO spiral blade. An unreamed femoral nail with a spiral blade was inserted to treat an unstable subtrochanteric femoral fracture. which lead to fracture union at 5 months. Eight months post-operatively the patient started to complain of left hip pain. Serial radiographs revealed progressive osteoporosis of the proximal femur possibly due to the stress sharing effect of a stiff intramedullary device, which continued to bear a significant amount of the transmitted load. The cause of pain was a stress fracture of the femoral neck and the AO spiral blade, which only became radiologically visible 4 months after the start of the symptoms (1 year after the initial operation). The implant was removed and replaced by a cemented hemiarthroplasty. This case reaffirms the difficulty in diagnosing a stress fracture through a metallic implant. The delay in diagnosis may be shortened if stress fracture were included as an expected complication following an intramedullary nailing. PMID- 10853919 TI - Have some guidelines for the treatment of acute bipolar depression gone too far in the restriction of antidepressants? AB - This paper gives a critical review of recommendations concerning the drug treatment of acute bipolar depression. The suggestions of different guidelines and consensus papers, especially in US-American and Canadian psychiatry, have a strong tendency against antidepressants in bipolar depression; they prefer mono therapy with mood stabilizers and, in the case of co-medication with mood stabilizers and antidepressants in severe depression, to withdraw the antidepressant as early as possible. The intention of this restrictive use is to avoid the risk of mania and the risk of rapid cycling induced by antidepressants. However, apparently the risk of suicidal acts, which is as prominent in bipolar depression as in unipolar depression, has been totally neglected. Furthermore, the fact that none of the mood stabilizers have proven their antidepressive efficacy leads not only to the risk of depression-related suicidal behavior but also to the risk of chronicity of depressive symptoms due to undertreatment. Altogether the view expressed in some guidelines and consensus papers appears not well balanced. Furthermore, the fact that apparently the selective serotonin re uptake inhibitors and possibly some other modern antidepressants have only a low risk of inducing a switch to mania should stimulate a rewriting of the guidelines on drug treatment in acute bipolar depression in a less restrictive way concerning the use of antidepressants. PMID- 10853920 TI - A five-year longitudinal study of the regional cerebral metabolic changes of a schizophrenic patient from the first episode using Tc-99m HMPAO SPECT. AB - This is a naturalistic study of the relationship between cerebral metabolic activity, clinical symptoms and treatment response in a schizophrenic patient for 5 years from the onset of her illness. Serial technetium-99m-HMPAO brain SPECT was used to measure regional cerebral metabolism. The Cambridge Neurological Inventory and neuropsychological tests (WAIS-R verbal subscales, Wisconsin Card Sorting Test, semantic verbal fluency, logical memory in Weschler Memory Scale) were used for neurocognitive assessment. Under-activity of the left temporal area was observed in the course of patient illness despite remission of the psychotic symptoms. Bilateral prefrontal metabolic under-activity was noted at the emergence of negative symptoms, executive neurocognitive dysfunction and the treatment-resistant state. After response to clozapine, the right prefrontal activity returned to a normal level. Our findings suggested that persistent left temporal underactivity detected by SPECT despite clinical remission may indicate a vulnerability for further relapses and development of a treatment-resistant state. Treatment-resistant state, negative symptoms and executive neurocognitive deficit may involve abnormal prefrontal metabolic activity and can be alleviated in clozapine-responsive patients. PMID- 10853921 TI - Awareness of illness and outcome in schizophrenia. AB - The purpose of the present study was to investigate whether awareness of illness affects specific measures of outcome in schizophrenia. Patient awareness was evaluated using a shortened version of the Scale to Assess Unawareness of Mental Disorder (SUMD). Patient outcome was assessed by means of the Strauss-Carpenter scale. Our findings indicate that lack of awareness of "negative symptoms" has a considerable impact on outcome: in fact "Social Contacts" highly correlated with Blunt Affect, Anhedonia and Asociality items on the SUMD. Lack of awareness seems then to be a powerful predictor of poor outcome. PMID- 10853922 TI - Cavum veli interpositi and psychotic disorder in a monocygotic twin. AB - The case of monocygotic twins discordant for a psychotic disorder is presented. An anomaly of the septum pellucidum, a so-called cavum veli interpositi was found in the psychotic twin while his brother showed no such anomaly. Previous studies have shown a higher prevalence of septum pellucidum anomalies in schizophrenic patients. Abnormalities of the septum pellucidum may be associated with disturbed neuronal development in distinct limbic brain areas which cannot yet be visualized yet by brain imaging techniques. The finding of the cavum veli interpositi in the psychotic twin could be incidental; however, it may indicate a dysgenic process in early brain development and, thus, play a significant role in the etiology of psychosis. PMID- 10853923 TI - A study of visual and auditory verbal working memory in schizophrenic patients compared to healthy subjects. AB - Impaired working memory (WM) performance is considered as a central feature of schizophrenia. Divided into two components, verbal and spatial, WM has been shown to involve frontal and parietal regions. Verbal WM can be tested either visually or aurally. The present study aimed to test schizophrenic patients in both visual and auditory verbal WM in order to assess a possible distinct pattern of alteration of these two modalities. Twenty-four schizophrenic patients and 24 healthy controls were compared with 2-back continuous visual and auditory verbal WM testing. Both groups were also tested on a neuropsychological battery including Wisconsin Card Sorting Test (WCST). Schizophrenic patients were less efficient in both verbal WM tests. When taking age and educational level as covariates and both WM modalities as dependent variables, there was no differential effect of modalities across groups. In further exploratory analyses, partial correlations brought association between verbal WM and psychosocial adaptation, WCST and length of illness. These results suggest a similar pattern of alteration of both modalities of verbal WM in schizophrenic patients. The implications of this finding are discussed. PMID- 10853924 TI - Lowered serum dipeptidyl peptidase IV activity in patients with anorexia and bulimia nervosa. AB - The aim of this study was to examine whether anorexia nervosa and bulimia nervosa are accompanied by lower serum activity of dipeptidyl peptidase IV (DPP IV, EC 3.4.14.5), a membrane-bound serine protease that catalyses the cleavage of dipeptides from the amino-terminus of oligo- and polypeptides. Substrates of DPP IV are, amongst others, neuroactive eptides, such as substance P, growth hormone releasing hormone, neuropeptide Y, and peptide YY. DPP IV activity was measured in the serum of 21 women with anorexia nervosa, 21 women with bulimia nervosa and 18 normal women. Serum DPP IV activity was significantly lower in patients with anorexia nervosa and bulimia nervosa than in the normal controls. In the total study group, there were significant and inverse relationships between serum DPP IV activity and the total scores on the Bulimic Investigatory Test, Edinburgh, the Eating Disorder Inventory (EDI) and the Hamilton Depression Rating Scale. In the total study group no significant correlations between DPP IV and age, body weight or body mass index could be found. It is concluded that lowered serum DPP IV activity takes part in the pathophysiology of anorexia and bulimia nervosa. It is hypothesised that a combined dysregulation of DPP IV and neuroactive peptides, which are substrates of DPP IV, e.g. neuropeptide Y and peptide YY, could be an integral component of eating disorders. PMID- 10853925 TI - Five-factor model of schizophrenic psychopathology: how valid is it? AB - Aim of the study was to examine the consistency of the five-factor model of schizophrenic symptoms, assess its validity and evaluate its dimensional factor structure using confirmatory factor (CFA) analysis. A sample of 258 randomly assigned DSM-III R patients with schizophrenic disorders were studied by means of the structured clinical interview for the Greek validated Positive and Negative Syndrome Scale (PANSS) and were rated on its 30 items. Patients' scores were subjected to principal component analysis (PCA) with varimax rotation. Internal consistency for each of the components was determined by the use of Cronbach's alpha. External validity of the model derived was investigated by searching for possible relationships between the components and sociodemographic characteristics with the aid of canonical correlation analysis. Confirmatory factor analysis (CFA) was also performed. Using the scree plot criterion PCA revealed a five-factor model. These factors were interpreted as representing--in a decreasing order of relative importance--the following dimensions of schizophrenic psychopathology: negative, excitement, depression, positive and cognitive impairment. The model was comparable with six previous factor analytic studies. Internal consistency was quite satisfactory whereas external validity was found to be not so powerful. CFA did not show that the proposed model yields an adequate factor structure. PMID- 10853926 TI - Comorbidity in ADHD-children: effects of coexisting conduct disorder or tic disorder on event-related brain potentials in an auditory selective-attention task. AB - In children with attention-deficit hyperactivity disorder (ADHD) some deficits in auditory information processing seem to exist. Further, comorbidity of ADHD with conduct disorder (CD) and tic disorder (Tic) is quite common but not yet fully understood. Thus, we investigated the effects of these two disturbances, when combined with ADHD, on electrophysiological correlates of auditory information processing. An auditory selective-attention task was used, and temporal as well as frontal lobe sensitive event-related electrical brain activity indicators like mismatch negativity (MMN) and negative difference wave (Nd), as well as P300 were registered in four groups of children (healthy controls, ADHD-only, and combined ADHD + CD as well as ADHD + Tic; total number 42). Performance measures showed that ADHD + CD had a higher impact on errors and reaction times than ADHD + Tic. The MMN effect indicated that all ADHD groups showed lower MMN amplitudes compared to normals, but only the group with ADHD + CD suffered from a significant deficiency in automatic auditory information processing. Nd and P300 amplitudes showed no significant group differences. It may be assumed that neurodynamic sufficiency in ADHD-only and ADHD + Tic children seems to be similarly impaired while there might be a greater deficit in ADHD + CD. PMID- 10853927 TI - Stenotrophomonas maltophilia endophthalmitis after intraocular lens implantation. AB - BACKGROUND: Stenotrophomonas maltophilia is an opportunistic, gram-negative bacillus. Endophthalmitis induced by S. maltophilia has been described in only two cases after intraocular lens implantation. We report S. maltophilia endophthalmitis in two patients with diabetes mellitus after intraocular lens implantation and compare the characteristics of the S. maltophilia-induced endophthalmitis with two previous cases. METHODS: A 68-year-old woman and a 74 year-old man with diabetes mellitus developed S. maltophilia endophthalmitis within 5 days of intraocular lens implantation. We performed intraocular lens removal and vitrectomy, which resolved the inflammation. No recurrences were found. RESULTS: Cultures grew S. maltophilia in both cases, and one of the organisms was multi-resistant. The final visual acuity was counting fingers and 0.3. The first case revealed a tractional retinal detachment during vitrectomy. CONCLUSIONS: S. maltophilia is a potential opportunistic intraocular pathogen, and the incidence of multiresistant S. maltophilia is increasing. S. maltophilia causes acute endophthalmitis, and its prognosis may not be poor unless the eye has a history of serious disease before the cataract surgery. The combined procedure of intraocular lens removal and vitrectomy was useful in resolving the inflammation and preventing recurrences. PMID- 10853928 TI - Comparative acute effects of brimonidine 0.2% versus dorzolamide 2% combined with beta-blockers in glaucoma. AB - PURPOSE: To assess the acute intraocular hypotensive efficacy of brimonidine tartrate 0.2% (a highly selective alpha2-adrenergic agonist) compared with dorzolamide 2% (a topical carbonic anhydrase inhibitor) as adjunct therapy to topical beta-blockers in patients with primary open-angle glaucoma. METHODS: A randomized cross-over masked study was performed. We enrolled one eye of each of 28 patients who were on different beta-blocker therapy. We measured the intraocular pressure (IOP) 2 h after the beta-blocker instillation; we then randomly administered one of the two drugs and we compiled an IOP diurnal curve. One month later we repeated the same procedures with the second drug. Unpaired Mann-Whitney U-test was used to compare decreases in IOP between the two drugs (P<0.05). RESULTS: Both brimonidine 0.2% and dorzolamide 2% have good ocular hypotensive efficacy, significantly lowering IOP when compared to beta-blocker therapy alone, for the whole diurnal curve. Maximum mean percent IOP decrease from baseline was 22.0+/-15.7% (4.0+/-2.9 mmHg) for dorzolamide 2% 6 h after instillation and 35.5+/-16.4% (7.0+/-4.1 mmHg) for brimonidine 0.2% 8 h after administration of the drug. When we compared the two treatments, brimonidine 0.2% showed a higher hypotensive effect than 2% dorzolamide after 4 h (28.4+/-16.8% vs 17.6 +/-9.3%; P=0.04) and 8 h (35.5+/-16.4% vs 21.6 +/-10.8%; P=0.04). CONCLUSION: This study indicates that 0.2% brimonidine acutely associated with beta-blockers is an interesting new combination treatment useful in the management of glaucoma. PMID- 10853929 TI - Quantitative analysis of visual field and optic disk in glaucoma: retinal nerve fiber bundle-associated analysis. AB - BACKGROUND: A study was performed to evaluate whether visual field analysis using a perimetric nerve fiber bundle map gives information additional to global visual field indices and cumulative defect curves for early glaucoma diagnosis. METHODS: One hundred and four control subjects, 124 patients with ocular hypertension (OHT), 97 patients with high-tension glaucoma without visual field defects (preHTG) and 91 patients with open-angle glaucoma with visual field defects [30 low-tension glaucoma (LTG), 61 high-tension glaucoma (HTG)] were included in this study. Correlation analyses were performed between (a) global visual field indices and total neuroretinal rim (NRR) area; (b) local mean values of four visual field areas and the NRR area of the corresponding four optic disk sectors; and (c) local mean values of 10 perimetric nerve fiber bundles (PNFB 1-10) according to Weber and Ulrich (1991) and the four optic disk sectors. The correlations were adjusted for global mean defect and total NRR. RESULTS: There were no significant correlations between NRR area and visual field in control subjects or in patients with OHT or preHTG for all three analyses. Significant correlations were found between the global visual field indices and the total NRR area for LTG and HTG. Significant correlations between local mean defects and NRR area of corresponding optic disk sectors were found only in LTG for the superior and inferior visual field area and the PNFB covering these areas. CONCLUSION: The method used for visual field analysis and sectorization of the optic disk does not give additional information on visual field defects in patients with normal global visual field indices and a normal cumulative defect curve. The nerve fiber bundle-related visual field analysis allows the topographical determination and quantification of glaucomatous damage. PMID- 10853930 TI - Evoked cortical potentials after electrical stimulation of the inner retina in rabbits. AB - BACKGROUND: Electrical stimulation of the retina using implantable devices may be one possible approach in the treatment of blindness causing progressive degeneration of the outer retina. Stimulation of ganglion cells or fibers could be achieved by epiretinal positioning of a microelectrode array as one component of a retinal prosthesis. Experiments were performed in rabbits to determine whether it is possible to elicit cortical responses with current pulses delivered via an epiretinal placed microelectrode array. METHODS: Polyimide-based microelectrode arrays with platinum electrodes and silicon-based TiN electrode arrays were implanted and placed onto the retinal surface of healthy pigmented rabbits after vitrectomy. The devices were temporarily fixated under PFCL and connected to constant current sources delivering bi- or monophasic current pulses. Cortical evoked potentials were recorded using subcutaneously implanted EEG electrodes over the visual cortex. RESULTS: The surgical procedures for implantation and fixation could be performed without serious complications. The electrode array could be placed in the area of the visual streak. Cortical potentials could be recorded after pulse train stimulation either with biphasic or with monophasic pulses. At threshold, pulses of 10 microA/phase and 100 micros/phase were necessary for detection of an electrically evoked cortical potential (EEP). Charge delivery at threshold varied between 0.1 and 0.3 nC/phase and charge densities varied between 1 and 12 microC/cm2. The EEP amplitude increased with increasing stimulus strength. The cortical response could be completely blocked with a retrobulbar injection of 4 ml lidocaine 2%. CONCLUSION: Constant current pulses delivered via Retina Stimulator devices equipped either with platinum electrodes or with TiN electrodes placed onto the inner retinal surface were able to elicit cortical potentials in the rabbit visual system. At threshold the charge delivery seems to be in a safe range. PMID- 10853931 TI - Hematopoietically derived retinal perivascular microglia initiate uveoretinitis in experimental autoimmune uveitis. AB - BACKGROUND: Hematopoietically derived cells in the retina were studied for the expression of molecules associated with antigen presentation. METHODS: Bone marrow cells of (Lewis x Brown Norway) F1 rats (LBNF1) were transplanted to sublethally irradiated Brown Norway (BN) rats to construct chimeric rats (LBNF- >BN). Each of 21 established chimeras received an adoptive transfer of uveitogenic Lewis T lymphocytes. Three rats were killed on each of 7 consecutive days. The right eye of each rat was processed for flat-mount preparation of the retina; the left eye of each was frozen for cryostat sectioning. All tissues were then stained with one of the following antibodies: OX-3 (Lewis-specific MHC class II marker), anti-ICAM, anti-B7- 1, anti-TNF-alpha or anti-IL-1beta. RESULTS: Initial clinical signs of EAU appeared first on day 4; by day 6, full-blown EAU was noted. The flatmount preparations revealed the presence of OX-3+ cells in the retina, perivascularly exhibiting dendritic morphology on day 2. These cells were observed in the retinal nerve fiber layer (NFL). No B7-1+, ICAM-1+, TNF-alpha+ or IL-1beta + cells were detected. Cryostat sections revealed positive cell staining of perivascular microglia and astrocytes in the retinal NFL with anti-IL-1beta and anti-TNF-alpha antibodies. CONCLUSIONS: Since only perivascular bone marrow derived cells are seen to express MHC class II molecules prior to onset of EAU, and since these cells also generate the cytokines IL- 1beta and TNF-alpha, it appears that initial presentation of antigen in the retina could be by these cells. PMID- 10853932 TI - Natural history of choroidal neovascularization induced by vascular endothelial growth factor in the primate. AB - BACKGROUND: A new model of choroidal neovascularization (CNV) has been developed in the primate by implanting vascular endothelial growth factor (VEGF) impregnated microspheres in the subretinal space. METHODS: CNV was induced in Macaca mulatta monkeys by implanting VEGF-impregnated gelatin microspheres in the subretinal space. Progression of CNV was followed for 24 weeks after surgery using fluorescein angiography. Eyes were enucleated at various time points, and lesions were evaluated for evidence of CNV by light microscopy and by immunohistochemical staining. RESULTS: CNV developed in 12 (92%) of 13 eyes. Fluorescein leakage was first observed in the 2nd postoperative week and was apparent for the following 12 weeks. CD31 staining for endothelial cells was first observed at day 7 and was evident for the following 8 weeks. Glial fibrillary acidic protein staining revealed a glial adhesion between the proliferative membrane and the retina at 6 weeks after implantation. Smooth muscle actin-positive cells were found a +2 weeks and remained prominent for at least the next 6 weeks. Cytokeratin-positive retinal pigment epithelial (RPE) cells, first identified in the proliferative membrane at day 3, predominated throughout the growth of the membrane. Macrophages (RAM-II positive) were present at day 3 but were no longer observed after day 7. CONCLUSION: In monkeys, subretinal implantation of VEGF-impregnated gelatin microspheres leads to the development of CNV. Early, disciform and reparative stages of CNV were observed, similar to those seen in humans. This model will be useful for studying the pathogenesis of CNV and for evaluating potential treatment strategies. PMID- 10853933 TI - A submicron emulsion of HU-211, a synthetic cannabinoid, reduces intraocular pressure in rabbits. AB - PURPOSE: To study the ocular hypotensive effect of a nonpsychotropic cannabinoid, HU-211 (11 -hydroxy-delta8-tetra-hydrocannabinol, dimethylheptyl), an N-methyl-D aspartate (NMDA) agonist, in normotensive rabbits. METHODS: The cannabinoid HU 211, being lipophilic, was incorporated into a stable oil-in-water submicron sterile emulsion, consisting of 0.12% (w/w) HU-211. A single- dose, randomized and double-masked study was designed, using a Digilab 30R pneumotonometer to measure intraocular pressure (IOP) in normotensive rabbits. RESULTS: Application of a single dose of HU-211 ophthalmic preparation resulted in an IOP reduction of 5.3 mmHg (24% of baseline), first evident at 1.5 h post application and persisting for over 6 h. A small but significant lowering of pressure (12.5% of baseline) occurred in the contralateral eyes of HU-211 treated rabbits, lasting for 4 h post treatment. CONCLUSION: Our work demonstrated that HU-211, incorporated into submicron emulsion, caused a 6-h-long reduction in IOP in the treated eye, with a lesser reduction in the contralateral untreated eye. PMID- 10853934 TI - Thermal load of laser aperture masks in nonmechanical trephination for penetrating keratoplasty with the Er:YAG laser: comparison between stainless steel and ceramic masks. AB - PURPOSE: Thermal effects on the laser aperture mask may play a major role in the thermal loading of the cornea during nonmechanical trephination in penetrating keratoplasty. The purpose of this study was to assess the temperature increase on the laser mask using the 2.94-microm Er:YAG laser in order to find suitable parameters for avoidance of thermal damage to the cornea. METHODS: Thermal load measurements were performed on donor (7.5 mm trephination diameter, 0.7 mm thickness, central hole 3.0 mm) and recipient (7.5 mm trephination diameter, 0.7 mm thickness, outer diameter 13.0 mm) aperture masks. The masks were either mounted on a thermal isolator or fixed directly on porcine corneal samples. Temperature increase was measured either under static conditions in the ablation area (setup 1) and at the opposite side of the mask (setup 2) or in the ablation area under dynamic conditions, rotating the whole globe to simulate a constant trephination speed with the mask positioned directly on a porcine cornea (setup 3). We used the NWL Er:YAG solid-state laser in a 1.3-mm free-running spot mode focused on the trephination margin (half of the beam on the mask and half of it on the cornea) with a pulse energy of 200 or 400 mJ and 18CrNi10 stainless steel versus three different types of ceramic masks (silicium carbide, silicium nitrite, aluminum oxide). Temperature was assessed using an infrared pyrometer with automatic data acquisition software for a personal computer. RESULTS: Overall, the temperature rise ranged between 43.6 K (metal donor mask at the trephination area with 400 mJ pulse energy) and 3.3 K (silicium carbide recipient mask at the opposite side of the mask with 200 mJ pulse energy). With all setups and both energy levels, the heating of the metal mask was significantly higher (P<0.02) than the heating of the three types of ceramic masks. The silicium carbide masks revealed the lowest temperature rise. Comparing the three setups, the temperature rise was maximal under static conditions in the ablation area and minimal at the opposite side, with the dynamic setup ranging in between. Temperature rise was significantly greater (P<0.04) in donor masks than in recipient masks for each mask material and both energy levels. CONCLUSION: The physical characteristics of silicium carbide masks seem superior to those of metal masks with regard to minimizing the thermal load of the epithelium or superficial stroma during Er:YAG laser trephination of the cornea for penetrating keratoplasty. PMID- 10853935 TI - Experimental immune-mediated blepharoconjunctivitis in rats induced by immunization with ragweed pollen. AB - BACKGROUND: A study was performed to compare the effects of immunization with ragweed pollen (RW) in two different adjuvants on the characteristics of a previously described model of experimental immune-mediated blepharoconjunctivitis (EC) in rats. METHODS: Lewis or Brown Norway (BN) rats were immunized with 100 microg of RW in emulsion with aluminum hydroxide [Al(OH)3] or complete Freund's adjuvant (CFA). Three weeks later, the animals were challenged with eye drops containing RW in PBS. Twenty-four hours after topical challenge, eyes, blood, and lymph nodes were obtained for histology, measurement of antigen-specific antibodies, and proliferation or cytokine assays, respectively. In addition to active immunization, recipients of RW-primed lymph node cells were challenged and evaluated as above. RESULTS: RW in both adjuvants induced infiltration with predominantly mononuclear cells in Lewis rats and eosinophils in BN rats. As well as active immunization, eosinophils were detected only in BN rats by adoptive transfer of cells. Lymphocyte proliferative responses to RW were high in immunized Lewis rats when CFA was used as an adjuvant. In contrast, proliferative responses in BN rats were higher when Al(OH)3 was used. RW-specific IgE was detected only in BN rats. There were no significant differences in RW-specific IgG1/IgG2a ratio among the four groups. Lewis rats had higher level of RW specific interferon-gamma in the culture supernatant. CONCLUSIONS: The characteristics of EC are different in Lewis and BN rats, dependent on the genetic background of the rat strains. The response to RW was similar to other previously used antigens, such as ovalbumin. PMID- 10853936 TI - A model for xenogenic immune response. AB - PURPOSE: To develop a model for analyzing the immune response after xenogenic human fetal retinal pigment epithelium (HFRPE) transplantation. MATERIALS AND METHODS: Pure sheets of HFRPE cells were isolated and attached to poly (DL lactide-co-glycolide) polymer films and HFRPE spheroids were formed. The spheroids were transplanted into the subretinal space of New Zealand albino rabbits and were observed for 5 months. Bare polymer films were transplanted into the subretinal space of Dutch Belted pigmented rabbits, serving as control. RESULTS: The polymer film was biodegraded within 3 weeks in the subretinal space. No signs of inflammation in the retina or choroid were observed. The HFRPE spheroids were easily transplanted into the subretinal space. The immune response was followed with ophthalmoscopy. Light microscopy indicated a localized immune response in the transplanted area in which the retina and the choroid were infiltrated with immune cells. This infiltration was denser in the choroid. CONCLUSIONS: HFRPE spheroid transplantation may be utilized as a model for studying the xenogenic immune response after HFRPE transplantation. This model may also have applications in evaluating the role of immune suppressive agents in preventing rejection after HFRPE transplantation. PMID- 10853937 TI - Involvement of superoxide generated by polymorphonuclear leukocytes in endotoxin induced uveitis. AB - BACKGROUND: Although superoxide is thought to be involved in the development of endotoxin-induced uveitis (EIU), the role of superoxide generation by polymorphonuclear leukocytes (PMNs) has not been fully elucidated. The purpose of this study was to investigate the role of peripheral blood PMNs in the development of EIU. METHODS: EIU was induced in Lewis rats by injection of lipopolysaccharide (LPS) in one hind footpad. Superoxide generation was assayed by measuring the reduction of ferricytochrome c (cyt c). EIU severity was assessed by histological examination, and the relationship between the injected dose of LPS in vivo and the intensity of superoxide generation by peripheral PMNs or intraocular PMNs was studied. Twenty-four hours after the injection of LPS (2, 20, or 200 microg/rat), peripheral blood PMNs were collected and stimulated with phorbol 12-myristate 13-acetate (PMA). The time course of superoxide generation by PMNs after LPS injection (3, 6, 12, 24, 48, and 72 h) was also investigated. To test the possible inhibition of superoxide generation by protein kinase C (PKC) inhibitors, H-7 and staurosporine were added for the incubation. In addition to the measurement of cyt c reduction, western blotting was used to detect PKC activity. The direct effect of LPS on PMNs was tested by priming naive PMNs with LPS in vitro. RESULTS: The intensity of superoxide generation by PMNs and the severity of EIU were dependent on the dose of injected LPS. No apparent superoxide generation was detected from intraocular PMNs. The time course of superoxide generation was similar to that of EIU severity. H-7 or staurosporine inhibited superoxide generation dose dependently and suppressed phosphorylation of PKC. Priming with LPS in vitro prompted minimal superoxide generation by naive PMNs. CONCLUSION: Superoxide generation by peripheral blood PMNs but not by intraocular PMNs from rats with EIU was demonstrated, and it is suggested that superoxide generation by PKC cascade might be involved in the pathogenesis of EIU. PMID- 10853938 TI - Osseous metaplasia with functioning bone marrow in hydroxyapatite orbital implants. AB - BACKGROUND: Bone formation within the hydroxyapatite implant has been reported in explanted spheres in humans. Bone-specific differentiation was observed to occur earlier in the pores of spherical hydroxyapatite implants enhanced with osteogenin within the rabbit socket. We observed previously unreported bone marrow formation in a coralline hydroxyapatite implant placed into the rabbit orbit after evisceration. METHODS: One eye of each of 10 New Zealand white rabbits weighing between 2 and 3 kg was eviscerated and implanted with hydroxyapatite spheres. The explanted hydroxyapatite spheres 20 weeks after surgery were examined under the microscope. RESULTS: Histopathologic examination of the excised implants showed the presence of trabeculae of mature bone with fatty marrow and hematopoietic elements. Scattered throughout the fatty tissue were bone marrow elements consisting of precursors of the granulocytic and erythroid series and also megakaryocytes. CONCLUSION: The osseous metaplasia with functioning bone marrow was incidentally observed in the coralline hydroxyapatite implant without the addition of any osteogenesis-inducing agents. PMID- 10853939 TI - Correlation of clinical and neuroradiological findings in down-gaze palsy. AB - BACKGROUND: Isolated down-gaze palsy is the least common pathology of vertical gaze. Patients with low-gaze palsy may consult an ophthalmologist with difficulty in reading and this may be the only ocular finding of a central nervous system lesion. METHODS: A 43-year-old man with isolated down-gaze palsy was examined. The medical history of the patient revealed that he had had myocardial infarction. RESULT: Magnetic resonance imaging disclosed an ischemic area at the right thalamus. CONCLUSION: Down-gaze palsy may be an important sign for the diagnosis of thalamic infarctions due to embolic syndrome. PMID- 10853940 TI - Convergence patterns of the posterior semicircular canal and utricular inputs in single vestibular neurons in cats. AB - The convergence of the posterior semicircular canal (PC) and utricular (UT) inputs in single vestibular nuclei neurons was studied intracellularly in decerebrate cats. A total of 160 vestibular neurons were orthodromically activated by selective stimulation of the PC and the UT nerve and classified according to whether or not they were antidromically activated from the spinal cord and oculomotor nuclei into vestibulospinal (VS), vestibulooculospinal (VOS), vestibuloocular (VO), and unidentified vestibular neurons. Fifty-three (33%) of 160 vestibular neurons received convergent inputs from both the PC and UT nerves. Seventy-nine (49%) vestibular neurons responded to PC inputs alone, and 28 (18%) neurons received inputs only from the UT nerve. Of 53 convergent neurons, 8 (15%) were monosynaptically excited from both nerves. Thirty-five (66%) received monosynaptic excitatory inputs from the PC nerve and polysynaptic excitatory or inhibitory inputs from the UT nerve, or vice versa. Approximately one-third of VS and VOS neurons received convergent inputs. A majority of the VS neurons descended to the spinal cord through the lateral vestibulospinal tract, while almost all the VOS neurons descended to the spinal cord through the medial vestibulospinal tract. The convergent neurons were found in all vestibular nuclei but more in the lateral nucleus and descending nucleus. The VS neurons were more numerous than VO neurons or VOS neurons. PMID- 10853941 TI - Evidence for effector independent and dependent representations and their differential time course of acquisition during motor sequence learning. AB - To investigate the representation of motor sequence, we tested transfer effects in a motor sequence learning paradigm. We hypothesize that there are two sequence representations, effector independent and dependent. Further, we postulate that the effector independent representation is in visual/spatial coordinates, that the effector dependent representation is in motor coordinates, and that their time courses of acquisition during learning are different. Twelve subjects were tested in a modified 2x10 task. Subjects learned to press two keys (called a set) successively on a keypad in response to two lighted squares on a 3x3 display. The complete sequence to be learned was composed of ten such sets, called a hyperset. Training was given in the normal condition and sequence recall was assessed in the early, intermediate, and late stages in three conditions, normal, visual, and motor. In the visual condition, finger-keypad mapping was rotated 90 degrees while the keypad-display mapping was kept identical to normal. In the motor condition, the keypad-display mapping was also rotated 90 degrees, resulting in an identical finger-display mapping as in normal. Subjects formed two groups with each group using a different normal condition. One group learned the sequence in a standard keypad-hand setting and subsequently recalled the sequence using a rotated keypad-hand setting in the test conditions. The second group learned the sequence with a rotated keypad-hand setting and subsequently recalled the sequence with a standard keypad-hand setting in the test conditions. Response time (RT) and sequencing errors during recall were recorded. Although subjects committed more sequencing errors in both testing conditions, visual and motor, as compared to the normal condition, the errors were below chance level. Sequencing errors did not differ significantly between visual and motor conditions. Further, the sequence recall accuracy was over 70% even by the early stage when the subjects performed the sequence for the first time with the altered conditions, visual and motor. There were parallel improvements thereafter in all the conditions. These results of positive transfer of sequence knowledge across conditions that use dissimilar finger movements point to an effector independent sequence representation, possibly in visual/spatial coordinates. Initially the RTs were similar in the visual and the motor conditions, but with training RTs in the motor condition became significantly shorter than in the visual condition, as revealed by significant interaction for the testing stage and condition term in the repeated measures ANOVA. Moreover, using RTs for single key pressing in the three conditions as baseline indices, it was again observed that RTs in the visual and motor conditions were not significantly different in the early stage, but motor RTs became significantly shorter by the late testing stage. These results support the hypothesis that the motor condition benefits more than the visual because it uses identical effector movements to the normal condition. Further, these results argue for the existence of effector dependent sequence representation, in motor coordinates, which is acquired relatively slowly. The difference in the time course of learning of these two representations may account for the differential involvement of brain areas in early and late learning phases found in lesion and imaging studies. PMID- 10853942 TI - Lateralized EEG components with direction information for the preparation of saccades versus finger movements. AB - During preparation of horizontal saccades in humans, several lateralized (relative to saccade direction), event-related EEG components occur that have been interpreted as reflecting activity of frontal and parietal eye fields. We investigated to what degree these components are specific to saccade preparation. EEG lateralization was examined within the interval (1 s) between a first (S1) and a second (S2) stimulus, after which a response had to be made (look left or right, or press a button with the left or right index finger). The visual S1 indicated either the direction (left vs right) and/or the effector (eye vs finger), and S2 (visual/auditory in different blocks) added the information not given by S1. An occipital component (220 ms after S1) was effector-independent, probably reflecting processing of the direction code. The following parietotemporal component (320 ms after S1) was specific for direction information. This component seems more relevant for finger movements than for saccades and may reflect a link between visual perception to action. A later frontal component (480 ms after S1) was specific for direction information and may be related to the planning of a lateral movement. One component was entirely specific for the preparation of a finger movement (the lateralized readiness potential before S2). Thus, several different lateralized processes in the S1-S2 interval could be delineated, reflecting hand-specific preparation, processing of the direction code, and the coordination of perception and action, but no components were observed as being specific for saccade preparation. PMID- 10853943 TI - Direction-dependent distortions of retinocentric space in the visuomotor transformation for pointing. AB - The aim of this study was to: (1) quantify errors in open-loop pointing toward a spatially central (but retinally peripheral) visual target with gaze maintained in various eccentric horizontal, vertical, and oblique directions; and (2) determine the computational source of these errors. Eye and arm orientations were measured with the use of search coils while six head-fixed subjects looked and pointed toward remembered targets in complete darkness. On average, subjects made small exaggerations in both the vertical and horizontal components of retinal displacement (tending to overshoot the target relative to current gaze), but individual subjects showed considerable variations in this pattern. Moreover, pointing errors for oblique retinal targets were only partially predictable from errors for the cardinal directions, suggesting that most of these errors did not arise within independent vertical and horizontal coordinate channels. The remaining variance was related to nonhomogeneous, direction-dependent distortions in reading out the magnitudes and directions of retinal displacement. The largest and most consistent nonhomogeneities occurred as discontinuities between adjacent points across the vertical meridian of retinotopic space, perhaps related to the break between the representations of space in the left and right cortices. These findings are consistent with the hypothesis that at least some of these visuomotor distortions are due to miscalibrations in quasi-independent visuomotor readout mechanisms for "patches" of retinotopic space, with major discontinuities existing between patches at certain anatomic and/or physiological borders. PMID- 10853944 TI - Negative cortical d.c. shifts associated with coordination and control in a prehensile force task. AB - Movement-related cortical d.c. shifts accompanying the execution of four different prehensile tasks were investigated using six normal adult subjects. The goal was to identify patterns of brain electrical activity that differentiated a precision grip configuration (thumb and index finger or 2f) from a full precision grip configuration (thumb and all fingers or 5f) at different total force levels. As such, this was the first study to systematically manipulate both grip configuration and force level while also measuring movement-related potentials (MRP) during the control phase of an isometric prehensile task. This investigation focused on assessing the sustained, performance-related negativity (N-P) associated with the execution of particular grip configurations at different total force levels (percentage maximum voluntary force, MVF). The results from this study demonstrated significant interactions between grip configuration, force level and amplitude of the N-P. First, an overall increase in force output does not correspond to larger N-P amplitudes under these task conditions. Second, force level and grip configuration interact significantly in determining the peak N-P, especially in low-force conditions. Overall, the findings reveal a task-specific sensitivity of movement-related potentials associated with the control phase of a prehensile force task while humans execute different grip configurations and force levels. PMID- 10853945 TI - Pavlovian aversive and appetitive odor conditioning in humans: subjective, peripheral, and electrocortical changes. AB - Differential Pavlovian conditioning of aversive and appetitive odors was examined in 30 male healthy subjects. The appetitive conditioning group (n=15) received a pleasant odor (vanilla), the aversive conditioning group (n=15) an unpleasant odor (fermented yeast) as unconditioned stimulus. Slides of two different neutral faces that were easy to discriminate served as conditioned stimuli (CS). An EEG was recorded from nine electrodes. Electromyographic activity was measured bilaterally from the m. corrugator supercilii and m. zygomaticus. The startle response was obtained from the m. orbicularis oculi. Finally, heart rate and skin conductance response were assessed. The subjective data and the skin conductance response revealed successful differential aversive conditioning. By contrast, the pleasant odor failed to produce appetitive odor conditioning. The conditioned and unconditioned response of the corrugator muscles confirm previous reports on the m. corrugator being strongly involved in the expression of negative affect. Contrary to previous findings, magnitude of the startle reflex was not found to be modulated depending on the valence of the CS. Central psychophysiological parameters showed little change during differential conditioning. The presence of subjective-evaluative conditioning and contingency awareness without significant changes in cortical and cardiovascular correlates might be due to extremely localized cortical processing of conditioned olfactory cues or primarily subcortical processing. The latter interpretation is strengthened by the presence of differential conditioning in non-voluntary responses such as the corrugator muscles. PMID- 10853946 TI - Evidence that ventrolateral thalamotomy may eliminate the supraspinal component of both pathological and physiological tremors. AB - Ventrolateral (VL) thalamotomy produced a marked reduction of oscillations related to the supraspinal components of Parkinson's disease tremor (4-7 Hz) and physiological tremor (8-12 Hz). Finger tremor was examined in nine patients undergoing unilateral VL thalamotomy and in nine age-matched controls. In comparison to the preoperative state, the relative percentage of power within the 7.6-12.5 Hz band did not increase after the surgical procedure. Furthermore, the amount of absolute power within the 7.6-12.5 Hz band was much lower for post surgical patients in comparison to matched controls when periods of tremor having equal amplitudes were compared. These results suggest that VL thalamotomy interrupts a common circuit involved in the supraspinal component of both physiological and pathological tremors. We provide evidence that the thalamus may be involved in circuits generating physiological tremor in humans. PMID- 10853947 TI - Long-term potentiation and paired-pulse facilitation in the prelimbic cortex of the rat following stimulation in the contralateral hemisphere in vivo. AB - We demonstrate here for the first time that the afferent fibres to the prelimbic component of prefrontal cortex and/or their associated, recurrent collateral local-circuit axons are capable of expressing paired-pulse facilitation and long term potentiation after stimulation of the prelimbic component in the contralateral hemisphere. Long-term potentiation resulted from both high- and low frequency stimulation protocols. It was not possible to obtain either depotentiation of previously potentiated synapses or long-term depression with the protocol used. Input-output analyses revealed interactions between separate components of the evoked responses. Since neurophysiological and computational theories of the rodent navigational system include the prefrontal cortex, these findings add important information to theories of prefrontal function and spatial representation in the rodent. PMID- 10853948 TI - Kinematics of fast hemiparetic aiming movements toward stationary and moving targets. AB - The aim of the present study was to gain insight into the control that hemiparetic subjects have over fast, unimanual aiming movements. Twelve hemiparetic subjects with cerebral palsy and twelve healthy subjects were asked to hit, as quickly as possible, stationary and moving targets projected onto a frontoparallel screen. The task was performed with the nonpreferred (spastic/nondominant) and preferred (nonspastic/dominant) arm. Although the pattern of kinematics of hemiparetic subjects generally corresponded with that reported in earlier reaching and grasping studies, the commonly observed prolonged movement time of the nonpreferred arm as compared to the preferred arm was absent. The spatial variability of the lateral hand displacements toward stationary targets was highest in the spastic arm of the hemiparetic subjects, indicating diminished motion stability. Even though hemiparetic subjects were expected to be unable to adjust their movements flexibly to the position and the velocity of a moving target, they used an initial estimate of where moving targets would be hit in the same way as the healthy subjects did, i.e., they started aiming toward a position in front of the target. In both subject groups, this spatial estimate and the movement time (MT) varied as a function of target velocity, suggesting that the use of target-velocity information in hitting moving targets is unaffected in spastic hemiparetic subjects. The results are related to possible deficits in the regulation of cocontraction underlying movement stability. PMID- 10853949 TI - Functional adaptation of reactive saccades in humans: a PET study. AB - It is known that the saccadic system shows adaptive changes when the command sent to the extraocular muscles is inappropriate. Despite an abundance of supportive psychophysical investigations, the neurophysiological substrate of this process is still debated. The present study addresses this issue using H2(15)O positron emission tomography (PET). We contrasted three conditions in which healthy human subjects were required to perform saccadic eye movements toward peripheral visual targets. Two conditions involved a modification of the target location during the course of the initial saccade, when there is suppression of visual perception. In the RAND condition, intra-saccadic target displacement was random from trial-to trial, precluding any systematic modification of the primary saccade amplitude. In the ADAPT condition, intra-saccadic target displacement was uniform, causing adaptive modification of the primary saccade amplitude. In the third condition (stationary, STAT), the target remained at the same location during the entire trial. Difference images reflecting regional cerebral-blood-flow changes attributable to the process of saccadic adaptation (ADAPT minus RAND; ADAPT minus STAT) showed a selective activation in the oculomotor cerebellar vermis (OCV; lobules VI and VII). This finding is consistent with neurophysiological studies in monkeys. Additional analyses indicated that the cerebellar activation was not related to kinematic factors, and that the absence of significant activation within the frontal eye fields (FEF) or the superior colliculus (SC) did not represent a false negative inference. Besides the contribution of the OCV to saccadic adaptation, we also observed, in the RAND condition, that the saccade amplitude was significantly larger when the previous trial involved a forward jump than when the previous trial involved a backward jump. This observation indicates that saccade accuracy is constantly monitored on a trial-to-trial basis. Behavioral measurements and PET observations (RAND minus STAT) suggest that this single-trial control of saccade amplitude may be functionally distinct from the process of saccadic adaptation. PMID- 10853950 TI - Motor dynamics encoding in the rostral zone of the cat cerebellar flocculus during vertical optokinetic eye movements. AB - The complex spike (CS) and simple spike (SS) activities of Purkinje cells in the rostral zone of the cerebellar flocculus were recorded in alert cats during optokinetic responses (OKR) elicited by a stimulus sequence consisting of a constant-speed visual pattern movement in one direction for 1 s and then in the opposite direction for 1 s. The quick-phase-free trials were selected. Ninety eight cells were identified as rostral zone cells by the direction-selective CS activity that was modulated during vertical but not horizontal stimuli. In most of the majority population (88 cells), with an increasing CS firing rate during upward OKR and an increasing SS rate during downward OKR, the inverse dynamics approach was successful and the time course of the SS rate was reconstructed (mean coefficient of determination, 0.70 and 0.72 during upward and downward stimuli, respectively) by a linear weighted superposition of the eye acceleration, velocity, position, and constant terms, at a given time delay (mean 10 ms) from the unit response to the eye-movement response. Standard regression coefficient (SRC) analysis revealed that the contribution of the velocity term (mean SRC 0.98 for upward and 0.80 for downward) to regression was dominant over acceleration (mean SRC 0.018 and 0.058) and position (-0.14 and -0.12) terms. The velocity coefficient during upward stimuli (6.6 spikes/s per degree/s) was significantly (P<0.01) larger than that during downward stimuli (4.9 spikes/s per degree/s). In most of the minority population (10 cells), with both CS and SS firing rates increasing during upward OKR, the inverse dynamics approach was not successful. It is concluded that 1) in the cat rostral zone Purkinje cells, in which the preferred direction is upward for CS and downward for SS, eye velocity and acceleration information is encoded in SS firing to counteract the viscosity and inertia forces, respectively, on the eye during vertical OKR; 2) the eye position information encoded in SS firing is inappropriate for counteracting the elastic force; 3) encoding of eye velocity information during upward OKR is quantitatively different from that during downward OKR: SS firing modulation is larger for upward than for downward OKR of the same amplitude; and 4) encoding of motor dynamics is obscure in cells in which the preferred direction is upward for both CS and SS. PMID- 10853951 TI - Transneuronal retrograde degeneration of retinal ganglion cells following restricted lesions of striate cortex in the monkey. AB - Transneuronal retrograde degeneration of retinal ganglion cells follows extensive striate cortical removal in macaque monkeys. Its extent depends on the age of the monkey at operation, post-operative survival, species and retinal eccentricity. Some studies of human patients with occipital lobe injury have found no evidence for transneuronal retrograde degeneration, suggesting that either degeneration may not occur or, if present, it is caused directly by secondary damage impinging upon the underlying white matter or the blood supply to the dorsal lateral geniculate nucleus and optic tract. We therefore studied retinal ganglion cell degeneration in three macaques in which only the striate cortex corresponding to the macular retina had been removed, thereby sparing extrastriate cortex and precluding interruption of the vascular supply to the thalamus and optic tract. There was extensive loss of ganglion cells in the central retina, corresponding to the central 10 degrees of vision. As the cortical lesion was too small to affect the thalamus or optic tract directly, the retinal degeneration must be transneuronal. Quantitative analysis showed a 65-80% loss of ganglion cells in the corresponding perifoveal retinae along the horizontal meridian. The results confirm that the loss of retinal ganglion cells following striate cortical lesions is predominantly transneuronal. PMID- 10853952 TI - Effect of amitriptyline on the messenger RNA of thyroid hormone-responsive genes in rat cerebral tissue. AB - To determine the molecular mechanisms of the potentiating effect of thyroid hormones (TH) on the therapeutic efficacy of tricyclic antidepressants (TCA), the expression of two known TH-responsive mRNAs was measured in control rats and rats treated with triiodothyronine (T3, 10 microg/100 g for 10 days), amitriptyline (10 mg/kg for 10 days), or combined T3 and amitriptyline. Northern blot analysis was carried out to measure the cerebral tissue content of a novel translational repressor (NAT-1) and another thyroid hormone-responsive (THR) mRNA. Rats treated with the combination of T3 and amitriptyline had significantly higher NAT-1 expression (2691.1+/-134.1 arbitrary units) than rats treated with T3 only (1688.5+/-77.8) or with amitriptyline only (1452.5+/-87.5) or the untreated control rats (731.3+/-23.0), P<0.01. Amitriptyline treatment did not alter the expression of THR mRNA or THR protein in either control or T3-treated rats. It is concluded that alterations in the expression of selective T3 responsive genes in cerebral tissue could be a mechanism of the known T3 potentiation of the therapeutic efficacy of TCA. PMID- 10853953 TI - Parvalbumin in cortical epithelial cells of the pigeon thymus. AB - We examined the distribution of parvalbumin in the pigeon thymus by light and electron microscopic immunohistochemistry. Tissues were also examined by conventional electron microscopy to determine the ultrastructure of immunoreactive cells. Parvalbumin immunoreaction was located in epithelial cells of the cortex, which formed dense mesh-like structures. Parvalbumin-positive epithelial cells were classified into 2 types. The first comprised elongated cells. In these, the nucleus was spindle-shaped, oval, or triangular, with a slightly irregular contour and contained rich heterochromatin peripherally. The cytoplasm was pale and processes extended laterally or ramified among the surrounding thymocytes. This type of cell formed the majority of immunoreactive cells. The other cell type consisted of polygonal epithelial cells. The nucleus was oval with deep indentations. Euchromatin occupied a large part of the nucleus. The cytoplasm contained numerous cell organelles compared with the elongated type, in particular, electron-dense vacuoles of various sizes and often bundles of tonofilaments. Both types of epithelial cell were interconnected by desmosomes. No secretory granules were found in the cytoplasm of elongated or polygonal cells. These results indicate the presence of heterogeneous group of parvalbumin-immunoreactive epithelial cells and suggest the likelihood of different functional roles for parvalbumin in the pigeon thymus. PMID- 10853954 TI - Leucocyte phenotypes in involuting and fully involuted mammary glandular tissues and secretions of sheep. AB - Mammary glandular tissues and mammary secretions were obtained from sheep at 2-60 d after weaning to study the leucocyte phenotypes associated with mammary involution. From 2-4 d after weaning, neutrophils were the predominant leucocytes in the alveolar and ductal lumina. Lymphocytes were present in the alveolar and ductal epithelium, interalveolar and periductal areas. Most of the lymphocytes in the alveolar and ductal epithelium (IEL) were CD8+, some were CD45R+ and few were CD4+. In the periductal clusters and in the interalveolar areas most of the lymphocytes were CD4+. There was a significant increase (P < 0.05) in the percentages of CD45R+ granulated IEL from 2 to 7 d after weaning, and this paralleled the increase in the percentages of apoptotic cells in the glandular epithelium. By 7-60 d after weaning, most cells within the alveolar and ductal lumina were macrophages followed by predominantly CD8+ lymphocytes. CD8+ lymphocytes were still predominant in the alveolar and ductal epithelium while CD4+ cells were predominant in the interalveolar areas. Very few gammadelta+ T cells were observed at all the stages examined. The cells in the mammary secretions correlated with those observed in the alveolar and ductal lumina. At the early stages of involution, the neutrophils and macrophages were heavily laden with lipid droplets, casein and cellular debris. The most interesting feature was the presence of cells either with extensive cytoplasmic processes (LCA+MHC class II+) or cytoplasmic veils (LCA+MHC class II+CD1+), probably dendritic cells. It is concluded that the cellular constituents of the mammary gland at the latter part of involution may afford the mammary gland more resistance to infection than the lactating gland and the gland at early stages of involution. The CD45R+IEL may trigger apoptotic cell death in the mammary glandular epithelium during mammary involution. PMID- 10853955 TI - Ultrastructural localisation and size distribution of collagen fibrils in Glisson's sheath of rat liver: implications for mechanical environment and possible producing cells. AB - The ultrastructure and size distributions of collagen fibrils in Glisson's sheath were investigated in the rat liver to analyse the mechanical environment around the fibrils and their possible cells of origin. Glisson's sheath was found to contain 2 populations of collagen fibrils with different diameters and distinct localisations, namely fibroblast-associated and bile epithelium-associated. Fibroblast-associated collagen was composed of fibrils arranged in bundles and constituted the majority of the collagen in Glisson's sheath. Bile epithelium associated collagen was represented by small dispersed groups of fibrils just beneath the basement membrane of the bile duct. The basement membrane of the bile duct was frequently reduplicated into a few or as many as 10 layers of laminae densae, with scattered collagen fibrils between these laminae. The diameters of the fibrils of both groups of collagen increased in relation to the calibre of the bile duct, whereas at any given place in Glisson's sheath bile epithelium associated collagen fibrils had a smaller diameter compared with those of the fibroblast-associated fibrils. The increment in fibril diameter along the bile duct is considered to be correlated with the increase in mechanical stress acting on Glisson's sheath. The difference in diameter between the 2 populations as well as the incorporation of fibrils between the laminae densae of the basement membrane of the bile duct supports the view that the bile epithelium-associated collagen is produced by the epithelial cells of the bile duct, thus having a different origin from that of fibroblast-associated collagen. These findings provide the first evidence that the epithelial cells of the interlobular bile duct produce fibril-forming collagen. Furthermore, it is suggested that cholestasis stimulates the epithelial cells of interlobular bile duct to increased synthesis of fibril-forming collagen that is also produced by these cells under physiological conditions. PMID- 10853956 TI - Three-dimensional reconstruction of tetraploid<-->diploid chimaeric mouse blastocysts. AB - Studies of tetraploid<-->diploid (4n<-->2n) mouse chimaeras have demonstrated unequal contributions of 4n cells to different tissues of the midgestation conceptus. Such a pattern has also been reported in chimaeras as early as E3.5d, which show an enhanced contribution of 4n cells to the mural trophectoderm (Everett & West, 1996). In this study, sectioned 4n<-->2n and 2n<-->2n control chimaeric blastocysts were digitised and reconstructed in 3 dimensions (3-D). The 3-D images revealed only limited mixing of cells from the 2 contributing embryos of individual blastocysts in both chimaera groups. Consequently, the distribution pattern of the 2 cell types was dependent on the spatial relationship between the orientation of the blastocyst and the boundary between the 2 clusters of cells. The distribution patterns observed were not strikingly different for 4n<-->2n and 2n<-->2n chimaeras, each showing some transgenic positive cell contribution in all 3 identifiable developmental lineages. It was notable, however, that in all 4n<-->2n blastocysts at least some 4n cells were located adjacent to the blastocyst cavity. Such a consistent pattern was not evident in 2n<-->2n chimaeras. This study has demonstrated the value of 3-D reconstructions for the analysis of spatial relationships of 2 cell populations in chimaeric mouse blastocysts. PMID- 10853957 TI - Effects of chronic oestrogen treatment are not selective for uterine noradrenaline-containing sympathetic nerves: a transplantation study. AB - Previous studies have shown that chronic administration of oestrogen during postnatal rat development dramatically reduces the total content of noradrenaline in the uterine horn, abolishes myometrial noradrenergic innervation and reduces noradrenaline-fluorescence intensity of intrauterine perivascular nerve fibres. In the present study we analysed if this response is due to a direct and selective effect of oestrogen on the uterine noradrenaline-containing sympathetic nerves, using the in oculo transplantation method. Small pieces of myometrium from prepubertal rats were transplanted into the anterior eye chamber of adult ovariectomised host rats. The effect of systemic chronic oestrogen treatment on the reinnervation of the transplants by noradrenaline-containing sympathetic fibres from the superior cervical ganglion was analysed on cryostat tissue sections processed by the glyoxylic acid technique. In addition, the innervation of the host iris was assessed histochemically and biochemically. The histology of the transplants and irises was examined in toluidine blue-stained semithin sections. These studies showed that after 5 wk in oculo, the overall size of the oestrogen-treated transplants was substantially larger than controls, and histology showed that this change was related to an increase in the size and number of smooth muscle cells within the transplant. Chronic oestrogen treatment did not provoke trophic changes in the irideal muscle. Histochemistry showed that control transplants had a rich noradrenergic innervation, associated with both myometrium and blood vessels. Conversely, in oestrogen-treated transplants only occasional fibres were recognised, showing a reduced NA fluorescence intensity. No changes in the pattern and density of innervation or in the total content of noradrenaline of the host irises were detected after chronic exposure to oestrogen. We interpreted these results to indicate that the effects of oestrogen on uterine noradrenaline-containing sympathetic nerves are neither selective or direct, but result from an interaction between sympathetic nerve fibres with the oestradiol-primed uterine tissue. A potential effect of oestrogen on the neurotrophic capacity of the uterus is discussed. PMID- 10853958 TI - Characterisation of glycoconjugate sugar residues in the vomeronasal organ of the armadillo Chaetophractus villosus (Mammalia, Xenarthra). AB - Conventional carbohydrate histochemistry and the binding patterns of 21 lectins were analysed to characterise the glycoconjugate content in the components of the vomeronasal organ of the armadillo Chaetophractus villosus. The mucomicrovillous complex of the sensory epithelium bound most of the lectins studied. No reaction was observed with Con A, PSA, S-Con A and SBA, and the sustentacular cells were stained with UEA-I, DSL, LEL, STL and Con A. The vomeronasal receptor neurons were labelled with S-WGA, WGA, PNA, UEA-I, STL, Con A, S-Con A, ECL and RCA120. The basal cell layer reacted with S-WGA, WGA, LCA, UEA-I, DSL, LEL, STL, Con A, JAC and VVA. The nonsensory epithelium exhibited a differential staining in relation to the different components. The mucociliary complex stained with ECL, DBA, JAC, RCA120, STL, LCA, PHA-E, PHA-L, LEL, BSL-I and VVA. However, SJA and UEA-I stained the mucus complex lining a subpopulation of columnar cells. The cytoplasm and cell membranes of columnar cells was labelled with DBA, DSL and LCA. The apical region of these cells exhibited moderate reactivity with LEL and SJA. None of the lectins bound specifically to secretory granules of the nonsecretory cells. Basal cells of the nonsensory epithelium were labelled with DSL, LEL, LCA, BSL-I and STL. The vomeronasal glands showed a positive reaction with WGA, DSL, LEL, LCA, DBA, PNA, RCA120 and SBA. Subpopulations of acinar cells were observed with ECL, S-WGA, Con A, S-Con A and DBA. PNA and RCA120 stained the cells lining the glandular ducts. In comparison with previous results obtained in the olfactory mucosa of the same group of armadillos, the carbohydrate composition of the vomeronasal organ sensory epithelium differed from the olfactory sensory epithelium. This is probably related to the different nature of molecules involved in the perireceptor processes. PMID- 10853959 TI - Lectin binding characteristics of mouse placental cells. AB - The lectin binding characteristics of mouse placental cells were examined. Wax embedded tissue sections of placentae from d 14 pregnant mice were stained with 26 lectins, with a wide range of sugar specificities. Cell cultures prepared from d 14 mouse placentae and cultured for 24 h were stained with 7 of the lectins to determine if they could be used as markers for the different trophoblast cells in culture. In tissue sections all placental cell populations bound lectin but no lectin bound specifically to any single trophoblast population. All the lectins which bound to layer 1 cytotrophoblast lining the maternal blood spaces of the labyrinthine placenta also bound to the fetal endothelium of the labyrinthine placenta. Binding of lectin appeared strongest on the adluminal membrane of these cell populations suggesting a role for the carbohydrate moieties in nutrient transfer. Few lectins bound to junctional zone trophoblast. Overall, the binding of lectin to cultured cells did not correlate exactly with lectin binding to the cell populations in tissue sections. The value of lectins as markers for placental cells in culture was therefore found to be limited. Our findings indicate that carbohydrate expression by at least some placental cells may vary in culture from that expressed by the cells in vivo with obvious concerns for the validity of functional in vitro studies. PMID- 10853960 TI - Corticofugal axons from adjacent 'barrel' columns of rat somatosensory cortex: cortical and thalamic terminal patterns. AB - The cortical representations of the vibrissae of the rat form a matrix in which each whisker has its own area of cortex, called a 'barrel'. The afferent pathways from the periphery travel first to the trigeminal nuclei and thence via the ventroposteromedial thalamus (VPM) to the cortical barrels have been described in detail. We have studied the output from barrels by filling adjacent areas of the primary somatosensory cortex (SI) with either Phaseolus vulgaris leucoagglutinin (PHA-L) or biotinylated dextran amine (BDA) and demonstrating the course and terminations of the axons that arise within the barrel fields. The method not only dramatically illustrates the previously described corticothalamic pathway to VPM but also demonstrates a strict topography in the cortical afferents to the thalamic reticular nucleus (RT). Cells supplying the RT projection are found below the barrels in layer IV. Connections to the posterior thalamus, on the other hand, have no discernible topography and are derived from cortical areas surrounding the barrels. Thus the outputs of these 'septal' areas return to the region from which they receive thalamic input. The corticocortical connections are also visible in the same material. Contralateral cortical connections arise from the cells of the septa between barrels. The projections to secondary somatosensory area (SII) are mirror images of the barrel pattern in SI with rather more overlap but nonetheless a recognisable topography. PMID- 10853961 TI - Neonatal pulmonary hypertension prevents reorganisation of the pulmonary arterial smooth muscle cytoskeleton after birth. AB - The pulmonary arterial smooth muscle cell (SMC) cytoskeleton was studied in tissue from 36 piglets aged from within 5 min of birth to 21 d of age, and in 8 adults. An additional 16 piglets were made pulmonary hypertensive by exposure to hypobaric hypoxia (50.8 kPa) for 3 d. In conduit intrapulmonary elastic arteries alpha, beta and gamma actin, the 204, 200 and 196 kDa myosin heavy chain (MHC) isoforms and vinculin were localised by immunohistochemistry. The total actin content, the proportion of monomeric to filamentous alpha and gamma actin and changes in the proportions of the MHC isoforms were determined biochemically. Dividing SMCs were localised and quantified using Ki-67. We found a transient reduction in immunohistochemical expression of gamma actin, 204 kDa MHC isoform and vinculin at 3 and 6 d in the inner media, associated with a transient increase in Ki-67 labelling. The actin content also decreased at 3 and 6 d (P < 0.05), but there was a postnatal, permanent increase in monomeric actin, first the alpha then the gamma isoform. The relative proportions of the MHC isoforms did not change between birth and adulthood in elastic pulmonary arteries but in muscular arteries the 200 kDa isoform increased between 14 d and adulthood. Pulmonary hypertension prevented both the immunohistochemical changes and the postnatal burst of SMC replication and prevented the transient postnatal reduction in actin content. These findings suggest that rapid remodelling of the actin cytoskeleton is an essential prerequisite of a normal postnatal fall in pulmonary vascular resistance. PMID- 10853962 TI - X-ray microtomographic study of mineral distribution in enamel of mandibular rat incisors. AB - X-ray microtomography was used to study the mineral concentrations in sequential slices of enamel of 5 mandibular incisors which showed an increase from approximately 1.0 to approximately 2.7 g cm(-3) from the apex towards the incisal end. For points at the same distance from the apex, there were differences up to 0.6 g cm(-3) between the teeth. The change of mean concentrations in the slices with distance could be modelled as (different) saturating exponentials. Under the assumption of a uniform growth rate of a mandibular incisor of 0.6 mm per day and a common time origin for the start of maturation (taken as a mineral concentration of 1 g cm(-3)), the distances were transformed to a common time frame to give a pooled data set. A single saturating exponential could be fitted to this pooled transformed data; this was: Cm = 2.84-1.94exp (-0.18d) where Cm is the mean mineral concentration (g cm(-3)) and d the time (days) from the start of maturation. This gives an asymptotic concentration of 2.84 g cm(-3) towards the incisal end, with a time constant of 7.7 days. The mineral concentration distribution functions were found to be more positively skewed closer to the apex, but more negatively skewed towards the incisal end. The difference between the higher mineral concentration in the outer enamel and the enamel near the amelodentinal junction (ADJ) was approximately 3%. The direction of maximum increase in concentration from the outer enamel surface to the ADJ meets the boundary of the ADJ at approximately 80 degrees. Three dimensional surface rendering of isodensity contours showed that the previously described C-shaped pattern of mineralisation is not solely a surface phenomenon, but extends through the depth of the enamel. PMID- 10853963 TI - Normal growth and development of the lips: a 3-dimensional study from 6 years to adulthood using a geometric model. AB - A 3-dimensional computerised system with landmark representation of the soft tissue facial surface allows noninvasive and fast quantitative study of facial growth. The aims of the present investigation were (1) to provide reference data for selected dimensions of lips (linear distances and ratios, vermilion area, volume); (2) to quantify the relevant growth changes; and (3) to evaluate sex differences in growth patterns. The 3-dimensional coordinates of 6 soft-tissue landmarks on the lips were obtained by an optoelectronic instrument in a mixed longitudinal and cross-sectional study (2023 examinations in 1348 healthy subjects between 6 y of age and young adulthood). From the landmarks, several linear distances (mouth width, total vermilion height, total lip height, upper lip height), the vermilion height-to-mouth width ratio, some areas (vermilion of the upper lip, vermilion of the lower lip, total vermilion) and volumes (upper lip volume, lower lip volume, total lip volume) were calculated and averaged for age and sex. Male values were compared with female values by means of Student's t test. Within each age group all lip dimensions (distances, areas, volumes) were significantly larger in boys than in girls (P < 0.05), with some exceptions in the first age groups and coinciding with the earlier female growth spurt, whereas the vermilion height-to-mouth width ratio did not show a corresponding sexual dimorphism. Linear distances in girls had almost reached adult dimensions in the 13-14 y age group, while in boys a large increase was still to occur. The attainment of adult dimensions was faster in the upper than in the lower lip, especially in girls. The method used in the present investigation allowed the noninvasive evaluation of a large sample of nonpatient subjects, leading to the definition of 3-dimensional normative data. Data collected in the present study could represent a data base for the quantitative description of human lip morphology from childhood to young adulthood. PMID- 10853964 TI - The spleen of the one humped camel (Camelus dromedarius) has a unique histological structure. AB - The histology and structure of 38 spleens of the dromedary (aged 0.5-15 y) were studied in relation to age. The spleen was found to have a thick capsule (292+/ 106 mm) divided into an outer layer (113+/-39 mm) composed mainly of connective tissue and an inner layer (180+/-81 mm) consisting mainly of smooth muscle cells. Vascular and avascular trabeculae extend from the capsule, the former containing arteries and nerves but no trabecular veins, the latter being divided structurally into primary and secondary trabeculae. Subcapsular and peritrabecular blood sinuses around primary and vascular trabeculae are unique to the camel spleen. The central artery emerges from the periarterial lymphatic sheath and branches into up to 4 penicilli which extend as sheathed arterioles (42+/-8 microm). These are found near or surrounded by blood sinusoids of the red pulp. A wide marginal zone surrounds the white pulp and contains sheathed arteries but no marginal sinuses. The red pulp is characteristically divided into cords by secondary trabeculae and contains venous sinusoids of different sizes. The camel spleen is of a sinusal type that can store blood. The thick muscular capsule and trabeculae pump the stored blood according to the body's need. Both closed and open circulations are found. The venous return is unique as the blood flow is from the venous sinusoids of the red pulp to the peritrabecular sinuses to the subcapsular sinuses to the splenic vein. No significant structural differences related to age were found. PMID- 10853965 TI - Widespread expression of tartrate-resistant acid phosphatase (Acp 5) in the mouse embryo. AB - Tartrate-resistant acid phosphatase (TRAP, Acp 5) is considered to be a marker of the osteoclast and studies using 'knockout' mice have demonstrated that TRAP is critical for normal development of the skeleton. To investigate the distribution of TRAP in the mammalian embryo, cryostat sections of 18 d murine fetuses were examined by in situ hybridisation, immunohistochemistry and histochemical reactions in situ. Abundant expression of TRAP mRNA was observed in the skin and epithelial surfaces of the tongue, oropharynx and gastrointestinal tract including the colon, as well as the thymus, ossifying skeleton and dental papillae. TRAP protein was identified at the same sites, but the level of expression in the different tissues did not always correlate with apparent enzyme activity. The findings indicate that abundant TRAP expression is not confined to osteoclasts in bone, but occurs in diverse tissues harbouring cells of bone marrow origin, including dendritic cells and other cells belonging to the osteoclast/macrophage lineage. PMID- 10853966 TI - Ultrastructural study on the follicle-associated epithelium of nasal-associated lymphoid tissue in specific pathogen-free (SPF) and conventional environment adapted (SPF-CV) rats. AB - Membranous (M) cells in follicle-associated epithelium (FAE) play an important role in the mucosal immunity through transport of a variety of foreign antigens to the underlying mucosa-associated lymphoid tissue (MALT). We aimed to investigate the ultrastructure of M cells in the FAE covering nasal-associated lymphoid tissue (NALT) both in specific pathogen-free (SPF) rats and in conventional environment-adapted (SPF-CV) rats aged 8-38 wk. In NALT of both SPF and SPF-CV rats, FAE included the nonciliated microvillous cell, which appears to be an analogue of M cell previously described in other MALT. In SPF rats, M cells increased in number only slightly with age, and they maintained morphological uniformity irrespective of age. In SPF-CV rats, M cells selectively increased in number resulting in prominent expansion of FAE surface area in parallel with the duration of maintenance in a conventional environment. In addition, M cells in SPF-CV rats showed heterogeneity in their surface morphology such as the length and number of microvilli and cell surface area and outline. In addition, the FAE was stratified by various subtypes of M cells, which were characterised by several subcellular alterations including the presence of many keratin filaments, homogeneous dark bodies and extensive cytoplasmic interfoliation with wide intercellular spaces filled with amorphous proteinaceous material. These characteristics of M cells in SPF-CV rat were intimately related with a preferential influx of immunocompetent cells into the FAE, which was not seen or was very rare in SPF rats irrespective of age. The results suggest the possibility that NALT may effectively carry out the mucosal immune response against antigenic stimuli of different magnitude through the unique dynamics of M cells which seem to be influenced by the infiltration of immunocompetent cells. PMID- 10853967 TI - To what extent are the retinal capillaries ensheathed by Muller cells? A stereological study in the tree shrew Tupaia belangeri. AB - The cellular ensheathment of capillaries in the 3 outer capillary layers of the central retina of the adult tree shrew Tupaia belangeri was studied quantitatively by transmission electron microscopy. Using a stereological approach, the relative surface of capillary basal lamina ensheathed by Muller cells and by nonmacroglial cells (collectively termed non-Muller cells) was estimated in 5 animals. The participation of Muller cells was distinctly different in the 3 capillary layers studied. In the outermost capillary layer 1, the mean (standard deviation) percentage surface coverage by non-Muller cell processes was 46.8 (15.3)%. Much less of the capillary basal lamina was ensheathed by non-Muller cells in capillary layers 2 and 3 (3.0 (2.1)% and 0.3 (0.3)% respectively). The observed total variation of the stereological estimates for the surface fraction of Muller cells (expressed as the between-subject coefficient of variation) was significantly higher in capillary layer 1 (28.8%) compared with capillary layers 2 (2.2%) and 3 (0.3%). In capillary layer 1, the high observed total variation was due to a high biological variation among animals for the fractions of both Muller cell and non-Muller cell ensheathment. The rare occurrence of direct contacts between the capillary basal lamina and the perikarya of either microglial cells (capillary layer 3) or amacrine cells (capillary layer 2) corresponded well to the low stereological values obtained for the relative capillary surface ensheathed by non-Muller cells in these capillary layers. Previously, extensive and frequent contacts between the basal lamina of capillaries belonging to capillary layer 1 and horizontal cells had been observed in single sections. The present study quantitatively demonstrates a marked paucity of macroglial investment of capillaries located in capillary layer 1 of Tupaia. It can be concluded that horizontal cells ensheath most of the capillary surface not invested by Muller cells. PMID- 10853968 TI - Determination of age at death using combined morphology and histology of the femur. AB - Bone is characterised by age-related morphological and histological changes. We have previously established an automated method of recording bone morphometry and histology from entire transverse sections of cortical bone. Our aim was to determine whether data acquired using this automated system were useful in the prediction of age. Ninety-six specimens of human femoral middiaphysis were studied from subjects aged 21-92 y. Equations predicting specimen age were constructed using macroscopic data (total subperiosteal area (TSPA), periosteal perimeter (PP), endosteal perimeter (EP), cortical bone area (CA) and moments of area) and microscopic data (the number, size and diversity of pores and intracortical porosity) together with sex, height and weight. Both TSPA and PP were independent predictors of age but the number of pores was not a significant predictor of age in any equation. The age predicted by these equations was inaccurate by more than 8 y in over half the subjects. We conclude that we could not predict age at a clinically acceptable level using data from our automated system. This most likely reflects an insensitivity to regional age-related changes in bone histology because we recorded data from each entire cortex. Automated bone measurement according to cortical region might be more useful in the prediction of age. The inclusion of TSPA together with PP as independent predictors of age raises the possibility that a future measure of periosteal shape at the femoral diaphysis could also be helpful in the prediction of age. The accuracy reached with the relatively simple methods described here is sufficient to encourage the development of image-analysis systems for the automatic detection of more complex features. PMID- 10853969 TI - Human patellar articular proportions: recent and Pleistocene patterns. AB - The degrees of mediolateral asymmetry of the patellar articular facet, as well as the median and lateral articular angles of the facet, were compared across samples of recent humans and of Pleistocene archaic and modern fossil humans. All samples exhibit considerable variability in these patellar proportions. The articular angles are similar across the different samples, but there is a trend towards decreasing lateral angles with decreasing robusticity. The archaic humans exhibit significantly more symmetry of the medial and lateral facets than do any of the recent human samples. However, given the variability in medial versus lateral patellofemoral contact forces documented for extant humans and the roles of the distal oblique portions of vastus medialis and vastus lateralis in patellar stabilisation, it is unclear to what extent this variation in patellar articular proportions may affect knee kinesiology. The contrasts may be related to different levels of patellar stability and/or musculoskeletal hypertrophy, but they appear unlikely to have affected primary knee function. PMID- 10853970 TI - Bones in the heart skeleton of the otter (Lutra lutra). AB - In most mammalian species the cardiac skeleton is composed of coarse collagen fibres, fibrocartilage, and pieces of hyaline cartilage. Bone, the os cordis, is a regular constituent of the ruminant heart. The cardiac skeleton of the otter (Lutra lutra) has not previously been described. The skeleton in 30 otter hearts was studied by x-ray analysis and light microscopy. Serial sections were cut parallel to the atrioventricular plane and histochemical staining methods were performed to identify connective tissue fibres, glycosaminoglycans, mineral deposits, and bone. Age and sex of the animals under investigation were considered. The otter heart skeleton was composed of coarse collagen fibres with intercalated pieces of fibrous and/or hyaline cartilage, calcified cartilage, and lamellar bone with red or white marrow. Pieces of hyaline cartilage were not clearly defined: a perichondrial layer was missing and coarse connective tissue continuously transformed into fibrous and hyaline cartilage. In both sexes the amount of cartilage and bone were found to increase with age. Our results establish the presence of bony material in the heart skeleton of the otter, a small mammalian species. This finding indicates that differentiation of bone is not exclusively related to the size of the organ. Increasing amounts of calcified cartilage and bone correlated with increasing age. PMID- 10853971 TI - The steady-state pharmacokinetics of nevirapine during once daily and twice daily dosing in HIV-1-infected individuals. AB - OBJECTIVE: To investigate and to compare the steady-state plasma pharmacokinetics of nevirapine in a dosing regimen of 400 mg once daily versus 200 mg twice daily in HIV-1-infected individuals. DESIGN: Open-label, randomized, cross-over study. METHODS: Twenty HIV-1-infected individuals who already used nevirapine as part of their antiretroviral regimen were randomized to continue their current regimen (200 mg twice daily) or to switch to the alternate regimen (400 mg once daily). The steady-state plasma pharmacokinetics of nevirapine were assessed after 2 weeks during a 24-h period. Subsequently, patients were switched to the alternate regimen and the pharmacokinetics of nevirapine were assessed again after 2 weeks. Non-compartmental methods were used to calculate the area under the plasma concentration versus time curve (AUC[24h]), and the maximal (Cmax) and minimal plasma concentration (Cmin), the time to Cmax (t(max)), the plasma elimination half-life (t1/2), the apparent oral clearance (Cl/F) and the apparent volume of distribution (V/F). Differences in these pharmacokinetic parameters for the two dosing regimens were tested using ANOVA. RESULTS: The exposure to nevirapine, as measured by the AUC[24h], was not significantly different between the 400 mg once daily and 200 mg twice daily dosing regimen (P = 0.60). Furthermore, the values for t(max), t1/2 Cl/F and V/F were not significantly different between the two dosing regimens (P > or = 0.08). However, Cmax and Cmin were higher and lower, respectively, when nevirapine was used in the once daily regimen as compared with the twice daily regimen. The median values for Cmax and Cmin as measured for the once daily and twice daily regimens were 6.69 and 5.74 microg/ml, respectively (P = 0.03), and 2.88 and 3.73 microg/ml, respectively (P < 0.01). CONCLUSION: These data show that the daily exposure to nevirapine, as measured by the plasma AUC[24h], is not different between a 400 mg once daily and a 200 mg twice daily dosing regimen. However, Cmax and Cmin are higher and lower, respectively, for the once daily regimen as compared with the twice daily regimen. The clinical implications of these differences remain to be established. PMID- 10853972 TI - Clearance of human herpesvirus type 8 viraemia in HIV-1-positive patients with Kaposi's sarcoma treated with liposomal doxorubicin. Caelyx/KS Spanish Study Group. AB - OBJECTIVE: To assess the impact of liposomal doxorubicin on human herpesvirus type 8 (HHV-8) cell viraemia in HIV-infected patients with Kaposi's sarcoma. DESIGN: Prospective, non-controlled, multicenter study. METHODS: The presence of HHV-8 DNA was investigated by polymerase chain reaction in peripheral blood mononuclear cells from 46 HIV-positive patients with Kaposi's sarcoma. Samples were tested at baseline and every 3 months during treatment with liposomal doxorubicin. CD4 cell counts, plasma HIV RNA, and clinical outcome were recorded at baseline and at follow-up visits. RESULTS: HHV-8 sequences were detected in 32 (70%) patients at baseline. No significant differences were found between subjects with HHV-8 positive and negative results. The proportion of patients with positive HHV-8 viraemia decreased to 38% (10 of 26) after 3 months of treatment with liposomal doxorubicin (P < 0.01). Overall, 12 of 22 (57%) subjects with positive HHV-8 cell viraemia at baseline became negative during the treatment period. However, in one of them HHV-8 reappeared 8 months later despite being on therapy. On the other hand, six of eight subjects with negative HHV-8 at baseline remained negative thereafter. There were no significant changes in plasma HIV RNA, total lymphocyte, or CD4 cell counts during the treatment period. Clinical response of Kaposi's sarcoma to liposomal doxorubicin and clearance of HHV-8 viraemia did not correlate well. CONCLUSIONS: HHV-8 cell viraemia significantly decreased during treatment with liposomal doxorubicin in HIV infected patients with Kaposi's sarcoma, although the clinical response and HHV-8 clearance did not correlate well. PMID- 10853973 TI - HIV-1 alters T helper cytokines, interleukin-12 and interleukin-18 responses to the protozoan parasite Leishmania donovani. AB - OBJECTIVE: To investigate the in vitro and in vivo effect of HIV-1 on lymphoproliferative and T helper (Th) cytokine responses in leishmaniasis. METHODS: Th1 [interleukin (IL)-2 and interferon (IFN)-gamma] and Th2 (IL-4 and IL 10) as well as IFN-gamma-inducing cytokines (IL-12 and IL-18) were measured in antigen and mitogen-stimulated culture supernatants of peripheral blood mononuclear cells (PBMC) of healthy donors, HIV-infected and visceral leishmaniasis (VL) patients with or without HIV co-infection. RESULTS: Proliferative responses to phytohaemagglutinin (PHA) were significantly lower in PBMC from VL and asymptomatic HIV-infected persons compared with responses in healthy individuals. VL-HIV co-infected patients showed the lowest responses. Although there was no significant difference in the Leishmania-induced proliferative responses among the healthy group and those infected with HIV only, VL patients (with or without HIV) exhibited very low proliferation. When cultured with PHA or Leishmania, PBMC from healthy donors produced high levels of a Th1 cytokine (IFN-gamma) and low levels of Th2 cytokines (IL-4 and IL-10). In addition, co-culturing PBMC from healthy donors with a killed HIV preparation abrogated the production of IFN-gamma induced by Leishmania and augmented IL-4 and IL-10 production. Cells from HIV-infected patients produced low levels of IFN gamma, but high levels of IL-10. The addition of anti-IL-10 did not increase Leishmania-induced proliferative responses or IFN-gamma production. Both IL-12 and/or IL-18 responses were lower in VL patients, HIV-infected, or VL-HIV co infected patients as compared with those of healthy donors. CONCLUSION: The data suggest that the inhibitory effect of HIV and VL on proliferation and IFN-gamma production is not due to IL-10 alone, but that the defect induced by HIV and VL probably operates at the level of regulation of IFN-gamma-inducing factors, such as IL-12 and IL-18. PMID- 10853974 TI - The immunoglobulin superantigen-binding site of HIV-1 gp120 activates human basophils. AB - OBJECTIVE: To investigate the mechanism whereby HIV-1 envelope glycoprotein gp120 from four different isolates obtained in three different countries induces proinflammatory mediator release from normal human basophils. METHODS: Histamine, cysteinyl leukotriene C4 (LTC4) and interleukin 4 (IL-4) release into the supernatant was measured in gp120-stimulated peripheral blood basophils from HIV 1 and HIV-2 negative subjects. RESULTS: The HIV glycoprotein was a potent stimulus for release of these mediators in basophils purified from donors negative for HIV-1 and HIV-2. There was also a correlation (r = 0.58; P < 0.01) between the maximum IL-4 release from basophils induced by gp120 and by anti-IgE. Basophils from which IgE had been dissociated by brief exposure to lactic acid no longer released histamine in response to gp120 and anti-IgE. Anti-IgE specifically desensitized basophils to a subsequent challenge with anti-IgE and gp120. Human monoclonal IgM carrying the VH3 domain, but not that carrying the VH6 domain, inhibited gp120-induced secretion of histamine from basophils in a concentration-dependent manner. Synthetic peptides identical to regions distant from the N- and C-termini of gp120MN inhibited its activating capacity. CONCLUSIONS: gp120 acts as a viral superantigen interacting with the VH3 domain of IgE to induce the release of preformed and de novo synthesized mediators from human cells carrying the Fc fragment Fc epsilonRI receptor. PMID- 10853975 TI - Spontaneous and anti-Fas-induced apoptosis in lymphocytes from HIV-infected patients undergoing highly active anti-retroviral therapy. AB - OBJECTIVE: The aim of this study was to investigate susceptibility to spontaneous or anti-Fas-induced apoptosis in peripheral blood mononuclear cells (PBMC) from HIV-positive patients before and during highly active anti-retroviral therapy (HAART). DESIGN: A longitudinal study was performed on 12 evaluable patients on HAART. This cohort was analysed prior to and at week 2, 4, 8, 16 and 24 after beginning HAART. Variations in CD4 and CD8 cells, viral load, susceptibility to spontaneous or anti-Fas-induced apoptosis in the presence of IL-2, IL-4 or IL-12 were studied. Expression of Fas and Bcl-2 were also assessed. METHODS: Levels of HIV RNA were determined by a quantitative reverse transcription-PCR assay. Apoptosis was evaluated by staining isolated nuclei with propidium iodide followed by multiparameter flow cytometry analysis. RESULTS: Spontaneous apoptosis of PBMC was promptly inhibited after the start of treatment. Similarly, anti-Fas-induced apoptosis diminished greatly during treatment. Expression of Fas decreased significantly, while that of Bcl-2 remained statistically unchanged during the first 24 weeks of therapy. Levels of apoptosis correlated inversely to CD4 cell counts and directly to viral load in a highly significant way. Expression of Fas was directly correlated to apoptosis. Interleukin (IL)-2, but not IL-4 or IL-12, protected PBMC of HIV-positive individuals from spontaneous or anti-Fas-induced apoptosis before and during HAART. CONCLUSION: These results suggest that regulation of apoptosis and of Fas expression are involved in immunoreconstitution during HAART. PMID- 10853976 TI - 'Modeling' relationships among HIV-1 replication, immune activation and CD4+ T cell losses using adjusted correlative analyses. AB - OBJECTIVE: To model the relationships among HIV-1 replication, immune activation and CD4+ T-cell losses in HIV-1 infection. METHODS: Cross-sectional analysis of baseline data from the Viral Activation by Transfusion Study. Comparisons of unadjusted and adjusted correlative analyses to establish models for mechanisms of cell loss in AIDS. RESULTS: Using these analyses, significant correlations were found among plasma levels of tumor necrosis factor alpha (TNFalpha) and its type two receptor (TNFrII), interleukin-6 (IL-6), beta2-microglobulin, expression of CD38 and HLA-DR on CD8+ T lymphocytes and plasma levels of HIV-1 RNA. When correlations among these indices were adjusted for possible intermediary correlations, the relationship between HIV-1 RNA levels and all plasma markers of immune activation could be accounted for by the correlation between plasma HIV-1 RNA and plasma TNFrII levels. In addition, the negative correlations that both HIV-1 RNA levels and TNFrII levels had with CD4+ T-cell counts were partially accounted for by the correlations of HIV-1 RNA and TNFrII with CD38 expression on CD8+ T cells. In persons with advanced disease (CD4+ T cells < 50 x 10(6)/l) IL-6 levels were inversely correlated with CD4+ T-cell counts. CONCLUSIONS: This analysis is consistent with a model wherein HIV-1 replication induces TNFalpha expression that induces multiple other indices of immune activation. In this model, HIV-1 replication and TNFalpha expression induce CD4+ T-cell losses at least in part through mechanisms reflected in heightened CD38 expression. PMID- 10853977 TI - Long-term immunological response in HIV-1-infected subjects receiving potent antiretroviral therapy. AB - OBJECTIVE: To determine the long-term T-lymphocyte response to highly active antiretroviral therapy (HAART) and to define predictors of the immunological response. DESIGN: Cohort study, including 135 HIV-1-infected subjects at a city general practice who commenced HAART between 1996 and 1998. METHODS: Collection of plasma HIV-1 RNA, CD4+ and CD8+ T-lymphocyte data at 3-6 monthly time intervals over 2 years. RESULTS: Seventy-three subjects (54%) achieved suppression of plasma HIV-1 RNA to levels below 400 copies/ml during the observation period, 31 individuals (23%) had detectable plasma HIV-1 RNA below 10,000 copies/ml and 31 subjects (23%) had virological failures with viral loads above 10,000 copies/mL. Median CD4+ T lymphocytes increased from 246 to 463 x 10(6) cells/l, showing a median rise of 20 x 10(6) cells/l per month in the first 3 months and 7 x 10(6) cells/l per month thereafter. The proportion of individuals who reached CD4+ cell counts above 500 x 10(6) cells/l increased from 8% at baseline to 54% at 2 years. Treatment-naive individuals, subjects with a large reduction of HIV-1 RNA or a large early CD8+ increase had better early CD4+ responses. Long-term CD4+ T-cell increases were inversely correlated with mean plasma HIV-1 RNA levels. Baseline CD4+ T-cell count was the most important determinant of reaching CD4+ cell counts above 500 x 10(6) cells/l. Nineteen per cent of subjects had no further CD4+ T-cell increases in the second year of therapy despite undetectable viral load. CONCLUSIONS: Immune reconstitution is a slow process, showing a large individual variability. The virological response to HAART was the most important determinant of the immunological short- and long term response. PMID- 10853978 TI - Clinical progression of HIV-1 infection according to the viral response during the first year of antiretroviral treatment. Groupe d'Epidemiologie du SIDA en Aquitaine (GECSA). AB - OBJECTIVE: To compare HIV-disease progression according to changes of plasma HIV RNA observed in the year following initiation of a new antiretroviral treatment. DESIGN: Prospective cohort treated with two nucleoside analogues or a triple combination including a protease inhibitor. METHODS: A Cox model was used to estimate the effect of viral response during the first year after initiation of treatment on the subsequent occurrence of new AIDS-defining events or death. Viral response was fitted either as HIV RNA reduction during the initial 4-12 months of treatment or reduction during the first month. RESULTS: Among 773 patients (47% with triple drug combination) followed for a median period of 27 months, 62 patients experienced a clinical event. Poor viral responders (at least two measurements > 3.7 log10 copies/ml during 4-12 months of treatment) had a higher risk of disease progression than good responders (RNA < 2.7 log10 copies/ml) after adjustment [hazard ratio (HR), 2.24; 95% confidence interval (CI), 1.1 7-4.29]. Intermediate responders (2.7 < or = RNA < or = 3.7 log10 copies/ml) had a risk of progression comparable with that of good responders (HR, 1.43; 95% CI, 0.64-3.22). A large initial viral reduction was also a protective factor for clinical progression (HR, 0.51 for 1 log10 copies/ml increase of the reduction; 95% CI, 0.31-0.85) and was associated with the viral response during the subsequent 4-12 month period. No patient with a reduction < 0.5 log10 copies/ml in the first month was classified as a good responder in the subsequent 4-12 month period (P < 0.01). CONCLUSIONS: A sustained HIV RNA > 3.7 log10 copies/ml should suggest a prompt change of treatment. When the reduction in HIV RNA is < 0.5 log10 after 1 month of treatment, this action should be anticipated. A sustained HIV RNA level between 2.7 and 3.7 log10 copies/ml may permit the deferral of a change of drug regimen according to the patient's history and therapeutic options. PMID- 10853979 TI - Impact of protease inhibitor therapy on HIV-related oropharyngeal candidiasis. AB - OBJECTIVE: To determine the relationship between antiretroviral therapy and changes in prevalence and amount of oropharyngeal candidiasis (OPC) and skin test reactivity for delayed type hypersensitivity. DESIGN: Observational cohort. SETTING: University-based public hospital AIDS clinic. PATIENTS: Adults with advanced HIV infection who had been taking nucleoside transcriptase inhibitor drugs but had not taken a protease inhibitor and who started antiretroviral treatment with ritonavir. MAIN OUTCOME MEASURES: OPC lesions score, oral candidal colonization, oral candidal quantification, skin test reactivity for delayed type hypersensitivity (purified protein derivative, candidal and streptokinase antigens), plasma HIV RNA and CD4 cell count at weeks 8, 16 and 48 weeks. RESULTS: In the 99 patients who entered the study, there was a significant reduction in the HIV plasma RNA (mean log decrease from baseline at 48 weeks 0.88) and a significant increase in CD4 cell counts (mean CD4 cell increase from baseline at 48 weeks 128 x 10(6) cells/l). Only 17% of patients had < 200 copies/ml HIV RNA at 48 weeks. There were significant decreases in the prevalence of OPC lesions (31% at baseline to 1% at 48 weeks; P < 0.001), and in oral candidal loads [2226 to 811 colony-forming units (CFU)/ml; P = 0.0171]. The percentage of patients with at least one positive skin test increased significantly (6 to 28%; P < 0.05). Patients whose CD4 lymphocyte count was > 200 x 10(6) cells/l at 48 weeks had significantly lower oral candidal loads and were more likely to have a positive skin test than patients whose CD4 cell count was < 200 x 10(6) cells/l. CONCLUSION: In patients with advanced HIV infection, antiretroviral treatment including a protease inhibitor has a positive impact in the natural history of OPC. This positive impact appears to be correlated with a better immunological function and occurs despite continuous HIV replication. PMID- 10853980 TI - Long-term efficacy on Kaposi's sarcoma of highly active antiretroviral therapy in a cohort of HIV-positive patients. CISIH 92. Centre d'information et de soins de l'immunodeficience humaine. AB - OBJECTIVE: To assess the efficacy of highly active antiretroviral treatment (HAART) on AIDS-Kaposi's sarcoma (KS). DESIGN: Prospective cohort of patients followed for 24 months. SETTING: Four referral hospitals of the West Paris metropolitan area. PATIENTS/INTERVENTION: Thirty-nine AIDS-KS patients, 42 +/- 9 years old, who began HAART (HIV-protease inhibitor and two nucleoside analogues) between March and December 1996, were enrolled. One was lost to follow-up at month 12. MAIN OUTCOME MEASURES: KS response, using criteria of the AIDS clinical trials group (ACTG), CD4 cell counts, and plasma HIV-RNA, assessed every 6 months. ACTG TIS staging of KS. RESULTS: Eighteen patients had T1 KS and 21 T0 KS. One patient died from KS at month 6. KS improved progressively, with complete and partial response rates of 46% and 28% at month 24, respectively. Only six patients were still receiving systemic KS therapy at month 24. Complete response was observed in 10 of the 19 patients without systemic KS therapy at inclusion. Patients with complete response at month 24 had higher CD4 cell counts than others (465 +/- 343 versus 185 +/- 167 x 10(6)/l; P < 0.01), but the proportion of patients with HIV-1 RNA < 500 copies/ml was not significantly different. An increase in CD4 cell counts from inclusion to month 12 of > 150 x 10(6)/l [odds ratio (OR), 13.4; 95% confidence interval (CI), 2-82] and T0 KS at inclusion: [OR, 7; 95% CI, 1.1-42] were predictive of complete response at month 24. CONCLUSIONS: HAART appears to have prolonged efficacy on AIDS-KS, even without specific KS therapy, and this effect appears to be linked to the restoration of immune function. PMID- 10853981 TI - Kaposi's sarcoma and central nervous system disease: a real association or an artifact of the control group? Terry Beirn Community Programs for Clinical Research on AIDS. AB - OBJECTIVES: To test the hypothesis that Kaposi's sarcoma (KS) protects against four central nervous system (CNS) diseases in HIV-1-infected individuals. STUDY POPULATION AND DESIGN: The study population of 9404 subjects included participants in Terry Beirn Community Programs for Clinical Research on AIDS (CPCRA) protocols who were enrolled between September 1990 and September 1998. This was an observational study. METHODS: Proportional hazards regression was used to estimate adjusted relative risks for predictors of four central nervous system diseases. Covariates included occurrence of Kaposi's sarcoma, occurrence of other opportunistic infections or malignancies, baseline CD4+ count, and other baseline characteristics. RESULTS: Among the 5944 participants without progression to AIDS at entry, 451 developed a CNS disease. The adjusted relative risk of any CNS disease for those who developed Kaposi's sarcoma versus those who did not develop any AIDS-defining event was 1.41 [95% confidence interval (CI), 0.98-2.03; P = 0.06]. In contrast, the adjusted relative risk of any CNS disease for those with Kaposi's sarcoma versus those with some other non-Kaposi's sarcoma AIDS-defining event was 0.37 (95% CI, 0.24-0.57; P < 0.0001). Among the 3460 participants with progression to AIDS at entry, the adjusted relative risk of any CNS disease for those with Kaposi's sarcoma versus those with some other non Kaposi's sarcoma AIDS-defining event was 0.71 (95% CI, 0.40-1.25; P = 0.23). CONCLUSIONS: Our analyses indicate that the risk of CNS disease associated with Kaposi's sarcoma depends strongly on the reference or control group chosen. When compared to individuals with other non-Kaposi's sarcoma AIDS-defining diseases, Kaposi's sarcoma is associated with a lower risk of CNS disease in HIV-1 positive individuals. However, when compared to individuals with no AIDS-defining disease or with a similarly mild AIDS-defining disease such as invasive candidiasis, Kaposi's sarcoma is associated with an equivalent risk of CNS disease. PMID- 10853982 TI - Two decades of HIV infection in a cohort of haemophilic individuals: clinical outcomes and response to highly active antiretroviral therapy. AB - OBJECTIVES: Many haemophilic individuals infected with HIV died before receiving antiretroviral therapy (ART). Most who remain alive are chronically infected with hepatitis C virus (HCV), which has implications for their prognosis and choice of ART. The clinical status of a cohort of HIV-positive haemophilic men is reported together with their response to highly active antiretroviral therapy (HAART). DESIGN: Longitudinal cohort study. SETTING: A comprehensive care haemophilia centre. PATIENTS: A group of 111 haemophilic men who seroconverted to HIV in the period 1979 to 1985. RESULTS: The cohort has been followed since 1979. By 30 April 1999, 57 of the 111 men had developed AIDS and 65 had died: Kaplan-Meier rates of 57.0% [95% confidence interval (CI) 46.9-67.0) and 65.1% (95% CI 52.7 77.4) by 19.5 years, respectively. AIDS rates have declined since 1997 but death rates have remained high, largely owing to deaths from non-HIV-related causes. Thirty-five patients remain alive and under follow-up at the clinic. The 28 men who had received ART had lower CD4 cell counts than the seven patients who had not received ART, but the two groups were otherwise similar. In total, 21 patients are known to have started HAART while under care at the centre. By 10-12 months after starting HAART, viral loads dropped by 2.06 log10 copies/ml and CD4 cell counts increased by 60 x 10(6) cells/l. In 10 out of 18 patients with viral loads initially > 400 copies/ml, a viral load below this level was attained; four had changed therapy at the time. CONCLUSIONS: While the decision to initiate HAART in haemophilic men should be made carefully because of the possible adverse events, our results suggest that a good response rate was achieved in this group of men. PMID- 10853983 TI - Prevalence of genotypic and phenotypic resistance to anti-retroviral drugs in a cohort of therapy-naive HIV-1 infected US military personnel. AB - OBJECTIVE: While transmission of drug-resistant HIV-1 has been reported, estimates of prevalence of resistance in drug-naive populations are incomplete. We investigated the prevalence of genotypic mutations and phenotypic antiretroviral resistance in a cohort of HIV-1 infected U.S. military personnel prior to the institution of antiretroviral therapy. DESIGN: Cross-sectional cohort study. METHODS: Plasma was obtained from 114 recently HIV-1 infected subjects enrolled in an epidemiological study. Genotypic resistance was determined by consensus sequencing of a PCR product from the HIV-1 pol gene. Sequences were interpreted by a phenotypic-genotypic correlative database. Resistance phenotypes were determined by a recombinant virus cell culture assay. RESULTS: Genotypic mutations and phenotypic resistance were found at a higher than expected frequency. Resistance to non-nucleoside reverse transcriptase inhibitors was most common, with a prevalence of 15% of 95 subjects by genotype and 26% of 91 subjects by phenotype. Genotypic and phenotypic resistance respectively were found in 4% and 8% of subjects for nucleoside reverse transcriptase inhibitors and in 10% and 1% for protease inhibitors. One subject harbored virus with resistance to all three drug classes. CONCLUSIONS: A substantial frequency of resistance to antiretroviral drugs was identified in a therapy-naive U.S. cohort. In most cases, the genotypic and phenotypic assays yielded similar results, although the genotypic assay could detect some protease inhibitor resistance-associated mutations in the absence of phenotypic resistance. These data suggest the need for optimization of treatment guidelines based on current estimates of the prevalence of drug resistance in HIV-1 seroconverters. PMID- 10853984 TI - Preventing mother-to-child transmission of HIV-1 in Africa in the year 2000. AB - OBJECTIVES: Various approaches to preventing mother-to-child transmission (MTCT) of HIV have recently been, or are being, evaluated in developing countries, especially in Africa. New findings from these trials are now becoming available, the implications of which, for population-based intervention programmes, need urgent consideration. METHOD: A critical review of 18 randomized trials and other relevant studies from developing and industrialized countries. RESULTS: Most African results relate to trials of antiretroviral agents (ARV). They demonstrate efficacy in reducing transmission in the first 6 months of life with short regimens of zidovudine (ZDV), with or without lamivudine (3TC), and nevirapine (NVP) alone. Preliminary results suggest the long-term efficacy of zidovudine. Antiseptic and nutritional interventions have been shown to reduce maternal and infant mortality and morbidity but not MTCT of HIV. HIV confidential voluntary counselling and testing for pregnant women, a short regimen of peripartum ARV with alternatives to breastfeeding such as early weaning or breast milk substitutes from birth currently represent the best option to reduce MTCTof HIV in Africa. However, the prevention of postnatal transmission requires further research, particularly in view of the consequences of different feeding options and the possibility of post-perinatal exposure prophylaxis of newborns with ARV. Issues relating to the implementation of currently validated strategies are discussed. PMID- 10853985 TI - HIV-1 prevalence, HIV-1 subtypes and risk factors among fishermen in the Gulf of Thailand and the Andaman Sea. AB - OBJECTIVE: Mobile populations are thought to be at high risk for HIV-1 infection. This study aims to determine the prevalence of HIV-1 infection, HIV-1 subtypes and socio-demographic and behavioural risk factors among fishermen in the Gulf of Thailand and the Andaman Sea. METHODS: A cross-sectional survey, consisting of face-to-face interviews and the collection of oral fluid samples, was conducted in Samut Sakorn, Ranong, Songkhla and Traat Provinces, Thailand, between January and April 1998. Oral fluid samples were double-tested for HIV-1 antibody by IgG antibody capture enzyme-linked immunosorbent assay and enzyme immunoassay, and confirmed by Western blot. The presence of subtypes B' and E was assessed using a peptide enzyme immunoassay. RESULTS: Of the 818 fishermen (582 Thai, 137 Burmese, 99 Khmer) 15.5% were HIV-1 positive: 14.6% among Thai, 16.1% among Burmese and 20.2% among Khmer. Of the 119 HIV-1 positive samples available for subtyping, 72 (61%) were subtype E, 15 (13%) were subtype B'; the subtype could not be determined for 32 (27%) samples. Sixteen per cent of subjects had ever visited a commercial sex worker outside Thailand. This behaviour was more prevalent among Khmer (40%) than among Thai and Burmese (12%). In univariate logistic regression analysis, being 25 to 32 years of age, compared with being older or younger; working as a fisherman between 4 and 10 years, compared with working for a shorter or longer period; being unmarried; being a steersman or mechanic, compared with being a skipper or ship hand; greater number of visits to commercial sex workers; having visited a commercial sex worker outside Thailand; alcohol or drug use before or during sex; being tattooed; and having a history of sexually transmitted disease were significantly related to prevalent HIV-1 infection. Male-to-male sex and injection drug use were rarely reported in this population. In multivariate analysis, being 25 to 32 years of age, being unmarried, having a tattoo and a greater number of visits to commercial sex workers remained in the model to predict HIV-1 prevalence. A history of drug injection was predictive for infection with HIV-1 subtype B'. CONCLUSIONS: These findings indicate a high HIV-1 prevalence among fishermen in the Gulf of Thailand and the Andaman Sea. Risk factor analysis suggests that heterosexual intercourse is the major mode of transmission in this population. Increased efforts to reduce the spread of HIV-1 among this epidemiologically important group are urgently needed. PMID- 10853986 TI - Is offering post-exposure prevention for sexual exposures to HIV related to sexual risk behavior in gay men? AB - OBJECTIVE: To examine the impact of the availability of postexposure prevention (PEP) for sexual exposures to HIV on sexual risk behavior among gay men. METHODS: Two cross-sectional samples of 529 gay men in San Francisco (June 1998, January 1999) completed face-to-face street interviews assessing sexual risk behavior and whether they had heard of PEP in general as well as whether they knew that PEP was available in San Francisco. The second sample was collected after a community wide outreach campaign had been conducted to increase people's knowledge that PEP was available in San Francisco. RESULTS: Of those who had heard of PEP at Time 1, 24% had recently had unprotected anal intercourse, versus 26% of those who had not heard of PEP. At Time 2, 37% of those who had heard of PEP had recently engaged in unprotected anal intercourse versus 26% of those who had not heard of PEP (chi2, 4.06; P = 0.03). At both time points, however, men who actually knew that PEP was available in San Francisco did not report more risk behavior than men who did not know PEP was available in San Francisco. In addition, only a small percentage at both time points self-reported that PEP had the effect of increasing their sexual risk behavior. CONCLUSIONS: There is little evidence that the availability of PEP for sexual exposures may be related to increased sexual risk-taking among gay men in San Francisco. The potential impact of PEP on risk behavior must, however, still be considered as part of the larger context of HIV/AIDS treatment optimism and possibly escalating levels of risk behavior among gay men. PMID- 10853988 TI - Ethical challenges of HIV vaccine trials in less developed nations: conflict and consensus in the international arena. PMID- 10853987 TI - Plasma cytomegalovirus DNA, pp65 antigenaemia and a low CD4 cell count remain risk factors for cytomegalovirus disease in patients receiving highly active antiretroviral therapy. AB - OBJECTIVE: To study the natural history and the current risk factors for cytomegalovirus (CMV) disease in the context of highly active antiretroviral therapy (HAART). SETTING: Prospective multicentre cohort in 15 university hospitals in France. METHODS: A group of 198 patients with CD4 cell count < 100 x 10(6) cells/l (or < 200 x 10(6) cells/l under HAART for at least 2 months), no previous CMV disease and CMV-positive serology were followed every 4 months clinically and for virological testing including HIV RNA and CMV blood markers (culture, pp65 antigenaemia, plasma CMV DNA and CMV late mRNA by the polymerase chain reaction). RESULTS: At inclusion, median CD4 was 77 x 10(6) cells/l (0-308) and 85% of the patients received protease inhibitors. The percentage of patients receiving HAART reached 99% at 12 months. After a follow-up of 23.6 months, the incidence of CMV disease was 3.2/100 patient-years [95% confidence interval (CI) 1.3-5.0]. In univariate Cox models, all the CMV markers, a CD4 cell count remaining < 75 x 10(6) cells/l and an HIV viral load > 100,000 copies/ml were predictive for CMV disease. The hazard ratios for CMV disease were 11 for blood culture; 14 and 70 for pp65 antigenaemia of > or = 1 and > or = 100 nuclei/200,000 cells, respectively; 35 for plasma CMV DNA; 6 for CMV mRNA; 29 for CD4 < 75 x 10(6) cells/l; and 12 for HIV RNA > 100,000 copies/ml. In a stepwise multivariate analysis, only three covariates were independently associated with the occurrence of a disease: plasma CMV DNA, pp65 antigenaemia > or = 100 nuclei/200,000 cells and a CD4 count < 75 x 10(6) cells/l. CONCLUSION: CMV blood markers and CD4 count < 75 x 10(6) cells/l remain risk factors for CMV disease in patients receiving HAART. Analysis of plasma CMV DNA by the polymerase chain reaction is a reproducible and standardized tool that could be used as a decision marker for initiating CMV pre-emptive therapy. PMID- 10853989 TI - Extracellular HIV-1 Tat protein activates the transcriptional repressor inducible cAMP early repressor in both the Jurkat cell line and primary peripheral blood mononuclear cells. PMID- 10853990 TI - Pharmacokinetics of indinavir in HIV-positive pregnant women. PMID- 10853991 TI - Efavirenz dosing in patients receiving continuous ambulatory peritoneal dialysis. PMID- 10853992 TI - Detection of drug-selected mutations in HIV-1 subtype E reverse transcriptase sequence using the line probe assay. PMID- 10853993 TI - Clinical features of acute HIV-1 infection: zidovudine-resistant isolates compared with zidovudine-sensitive isolates. PMID- 10853994 TI - Programmed granulocyte neutrophil death in patients at different stages of HIV infection. PMID- 10853995 TI - Decline of infectious skin manifestations in the era of highly active antiretroviral therapy. PMID- 10853996 TI - Increased prevalence of subclinical hypothyroidism in HIV patients treated with highly active antiretroviral therapy. PMID- 10853997 TI - Intestinal Chagas' disease in patients with AIDS. PMID- 10853998 TI - Human herpesvirus 8 infection of the genital tract of HIV-seropositive and HIV seronegative women at risk of sexually transmitted diseases. Canadian Women's HIV Study Group. PMID- 10853999 TI - Topical cidofovir is effective in treating extensive penile condylomata acuminata. PMID- 10854000 TI - Medical care costs of children born to HIV-infected mothers, in Abidjan, Cote d'Ivoire 1996-1997. DITRAME Study Group (ANRS 049a clinical trial) PMID- 10854001 TI - Endothelial dysfunction--a challenge for hypertension research. PMID- 10854002 TI - Endothelium-dependent vasodilation in hypertension: a review. AB - Using both in vitro and in vivo techniques, it has repeatedly been shown that endothelium-dependent vasodilation (EDV) is impaired in different forms of experimental hypertension (SHR, Dahl salt-sensitive rat, DOCA-salt rat and renovascular hypertension). EDV has also been found to be impaired in primary, as well as in secondary forms of human hypertension. Although impaired EDV is a general finding in hypertension, the pathophysiological mechanisms might differ between different forms of hypertension and between different types of vessels and vascular beds. Impaired activity of nitric oxide synthase, increased release of endothelin-1, increased production of a prostanoid-derived contracting factor, decreased generation of endothelium-derived hyperpolarizing factor/s and impairment caused by superoxide ions have all been shown to contribute to the impairment of EDV during different conditions. While most antihypertensive treatments improve EDV in experimental hypertension, no uniform picture has been seen in human hypertension, possibly because different antihypertensive drugs have different direct actions on EDV. This review shows that while impaired EDV has been found to be a general feature of hypertension, the mechanisms involved and the therapeutic opportunities have still to be established. PMID- 10854003 TI - Vascular endothelial cell activation associated with increased plasma asymmetric dimethyl arginine in children and young adults with hypertension: a basis for atheroma? AB - The mechanism behind the development of vascular complications of hypertension in the young human remains unclear. To explore the role of vascular endothelium generated nitric oxide (a known mediator of leucocyte-platelet-endothelial interactions) in this context, we investigated markers of endothelial activation (soluble vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), P-selectin, E-selectin), and von Willebrand factor and the plasma level of the endogenous nitric oxide inhibitor asymmetric dimethyl arginine (ADMA) in a group of 31 (17 male, mean age 9.4 years) hypertensive and 9 (4 male, mean age 9.1 years) healthy, normotensive children and young adults. We found raised levels of ADMA (mean (SEM) 235 (32) n mol/l) and VCAM-1 (median (range) 1237 (675-2700) ng/ml) in the plasma of hypertensive subjects compared with those of normotensives (ADMA, 103 (7) n mol/l and VCAM-1, 1005 (425-1650) ng/ml, respectively). Furthermore, in hypertensive subjects, higher VCAM-1 concentrations (r = 0.66, p < 0.001) and vWF concentrations (r = 0.37, p = 0.04) were significantly associated with a higher plasma ADMA level. Therefore, an isolated increase in plasma VCAM-1 in hypertensives in association with raised ADMA may signify a selective "non-inflammatory" endothelial activation triggered by endothelial nitric oxide synthase inhibition. Since VCAM-1 is implicated in the origins of atherosclerosis, ADMA may be an important contributory factor in increasing the risk of atheroma formation in hypertensive children and young adults. PMID- 10854004 TI - Effects of hormone replacement therapy on ambulatory blood pressure and vascular responses in normotensive women. AB - The effects of chronic oestrogen replacement therapy (ERT) (conjugated equine oestrogen 0.625 mg/day) and combined oestrogen and progestogen replacement therapy (HRT) (ERT plus continuous medroxyprogesterone acetate 5 mg/day) on 24-h ambulatory blood pressure recordings, forearm vascular resistance (FVR) and FVR responses to noradrenaline, angiotensin II, acetylcholine and nitroprusside were studied in 17 normotensive postmenopausal women in a 3-month randomized, double blind, placebo-controlled crossover trial with 1 month of therapy in each treatment arm. During the last few days of each 1-month treatment period, the subjects underwent 24-h ambulatory blood pressure recordings and measurements of FVR responses. ERT and HRT reduced mean 24-h diastolic blood pressure by 4 and 5 mmHg, systolic blood pressure by 6 and 9 mmHg and mean 24-h heart rate by 5 and 3 beats/min, respectively for ERT and HRT (p < 0.05). Basal FVR was reduced by approximately 18% by ERT and HRT, but FVR responses to noradrenaline, angiotensin II, acetylcholine and nitroprusside were unaffected. ERT and HRT therapy for 1 month lowers blood pressure and basal FVR, but does not appear to influence FVR responses to acetylcholine, nitroprusside, noradrenaline and angiotensin II. PMID- 10854005 TI - Organ damage in treated middle-aged hypertensives compared to normotensives: results from a cross-sectional study in general practice. AB - BACKGROUND: High blood pressure contributes to organ damage. However, during the past two decades there have been great advances in the medical treatment of hypertension. Technical progress has also made it easier to visualize organ damage. Hence we found it of interest to examine heart, brain and retina in a group of middle-aged treated hypertensives, comparing the results with those from a group of middle-aged normotensives. METHODS: The subjects were 40 (20 men) treated hypertensives and 40 (20 men) normotensives, who had previously taken part in a study in which ambulatory blood pressure monitoring had been performed. The heart was examined by echocardiography, the retina by photography and the brain by magnetic resonance imaging. RESULTS: Office blood pressure and 24-h systolic/diastolic blood pressure (S/D) were 141/86 (13/7) mmHg and 128/81 (11/6) mmHg in the hypertensives and 125/78 (10/8) mmHg and 118/74 (8/5) mmHg in the normotensives, respectively. Left ventricular mass was 101 (27) g/m2 in the hypertensives and 85 (18) g/m2 in the normotensives (p = 0.0025). The corresponding figures for the left atrium were 21.1 (3.1) mm/m2 in the hypertensives and 19.5 (2.2) mm/m2 in the normotensives (p < 0.001). E/A wave quotient was 1.09 (0.26) in the hypertensives and 1.26 (0.26) in the normotensives (p = 0.0045), while left ventricular systolic function did not differ between the groups. Ten hypertensives and one normotensive subject had left ventricular mass above normal range. Narrow retinal arteries were found in 22 hypertensives and 8 normotensives (p < 0.001). Brain magnetic resonance changes (deep white matter and/or periventricular) were found in 19 hypertensives and 9 normotensives (p = 0.0431). CONCLUSIONS: The hypertensives differed significantly from the normotensives concerning left ventricular mass, left atrium, left ventricular diastolic function and retinal vessel changes. Deep white matter and periventricular changes in the brain were also significantly different in the two groups. We can only speculate as to whether earlier antihypertensive treatment or further blood pressure reduction could have affected these differences. PMID- 10854006 TI - Cerebral perfusion in hypertensives with carotid artery stenosis: a comparative study of lacidipine and hydrochlorothiazide. AB - Focal cerebral hypoperfusion is a common finding in uncomplicated hypertensives even in the absence of large vessel atherosclerosis, and neuropsychological deficits correlate with cerebral hypoperfusion in hypertensive patients with cerebral microangiopathy. We investigated the effects on cerebral perfusion of the dihydropiridine calcium antagonist lacidipine and of hydrochlorothiazide (HCTZ) in asymptomatic hypertensive patients with concomitant atherosclerosis of the carotid arteries. Fifteen essential hypertensives (including 13 males, aged 55-75 years) with at least one 30-60% stenosis of the internal carotid artery at echo-color Doppler examination were treated in a double-blind, randomized, parallel study with lacidipine (4-6 mg od) or HCTZ (25-50 mg od) for 3 months after a 4-week single-blind placebo period. Regional cerebral perfusion was assessed at baseline and at the end of the treatment period with HMPAO-SPECT. Relative perfusion of cortical and subcortical areas was calculated as the ratio between their tracer activity and that of the cerebellum. At baseline, mean relative perfusion (MRP) of the cortical and subcortical areas was similar in the stenotic and the contralateral side. Despite the fall in pressure, lacidipine increased MRP both in the cortical and in the subcortical areas, whereas HCTZ increased MRP only in the cortical areas. The mean change in local vascular resistance, adjusted for initial perfusion value, was -20 A.U. (arbitrary unit) with lacidipine and -12 A.U. with HCTZ (p < 0.001). These differential effects of antihypertensive drugs on subcortical perfusion may be of benefit in the long term prevention of vascular dementia in hypertensive patients. PMID- 10854007 TI - Angiotensin-converting enzyme: induction by hypertension-induced vessel distension. AB - Vascular angiotensin-converting enzyme (ACE) activity is increased in a variety of disorders representing both early and late stages of atherosclerosis. The factors governing the induction of vascular ACE are poorly understood. We hypothesized that vascular ACE activity might be increased by hypertension induced vessel distension. Hypertension was induced in rats by suprarenal coarctation of the aorta. Analyses were performed 5 days and 4 weeks post operation. ACE activity and ACE mRNA level were increased in thoracic aortae from coarctation hypertensive rats that had been exposed to elevated blood pressure, whereas they remained at normal level in abdominal aortae from those rats that had been exposed to normal blood pressure. The degree of aortic ACE induction correlated well with the degree of the trans-stenotic blood pressure gradient. An increase in ACE transcript level was also observed in carotid arteries from coarctation hypertensive rats that had been exposed to elevated blood pressure. In contrast, ACE activity and ACE mRNA expression were not altered in tissues that did not contain any large arteries from coarctation hypertensive rats, although these tissues had been exposed to elevated blood pressure. These results demonstrate an induction of ACE in large arteries that had been exposed to elevated blood pressure, and they imply that the induction of vascular ACE is due to hypertension-induced vessel distension. PMID- 10854008 TI - Lack of association of angiotensin-converting enzyme and angiotensinogen genes polymorphisms with left ventricular structure in young normotensive men. AB - Left ventricular (LV) hypertrophy is a strong predictive factor for cardiovascular morbidity and mortality. LV structure and function are influenced by many variables, including genetic background. The potential role of gene polymorphisms of different components of the renin angiotensin system remains controversial. The aim of this study was to determine the influence of deletion/insertion (D/I) polymorphism of the angiotensin-converting enzyme (ACE) gene and M235-->T polymorphism of the angiotensinogen (AG) gene on left ventricular morphology and function in young normotensive men. The study included 110 normotensive healthy males (mean age 24 +/- 4 years, age range 18 to 34 years) who were assessed by echocardiography for LV structure and function. In all subjects ACE D/I polymorphism was evaluated using a polymerase chain reaction (PCR) method. M235-->T polymorphism assessment was available in 98 individuals. Significant differences between groups according to ACE I/D or AG M235-->T polymorphisms were not found for parameters of LV morphology or for parameters of systolic and diastolic function. When subjects with DD or ID genotypes were grouped, their LV mass index was higher than that in subjects with II genotypes (86 +/- 14 vs 79 +/- 15, r = 0.033, p = 0.05). There were no significant differences among other variables. In a population of young normotensive men where the influence of confounding variables such as age, gender and associated pathological conditions is minimized, the gene polymorphisms of ACE I/D and AG M235-->T are not important determinants of LV structure and function. PMID- 10854010 TI - Climb every mountain, till you find for your dream. PMID- 10854009 TI - Impact of a history of hypertension on symptoms and quality of life prior to and at five years after coronary artery bypass grafting. AB - AIM: To describe symptoms and other aspects of health-related quality of life (QoL) prior to and 5 years after coronary artery bypass grafting (CABG) in relation to a history of hypertension. METHODS: Patients who underwent CABG in western Sweden were approached prior to surgery and 5 years after the operation. Health-related QoL was estimated with the Physical Activity Score, the Nottingham Health Profile and the Psychological General Well-Being Index. RESULTS: In patients with a history of hypertension (n = 740) the 5-year mortality was 16.9% versus 12.4% among patients with no history (n = 1257; p = 0.004). Of 1717 patients available for the survey, 876 (51%) responded both prior to and 5 years after CABG. Of these, 36% had a history of hypertension. Compared with the situation prior to surgery there was an improvement in both hypertensive and non hypertensive patients in terms of physical activity, symptoms of dyspnea and chest pain and other estimates of health-related QoL. However, physical activity and dyspnea improved less in hypertensive than in non-hypertensive patients. CONCLUSION: Five years after CABG, a marked and significant improvement in terms of symptoms and other aspects of health-related QoL was observed among both hypertensive and non-hypertensive patients. However, improvement in physical activity was less marked in patients with a history of hypertension. Overall, a history of hypertension seemed to have a minor impact on improved well-being 5 years after coronary surgery. However, because of the limited response rate the results may not be applicable in a non-selected CABG population. PMID- 10854011 TI - Cloning of local growth factors involved in the determination of muscle mass. PMID- 10854012 TI - Exercise for cancer patients: a new challenge in sports medicine. PMID- 10854013 TI - The immune system in sport: getting the balance right. PMID- 10854014 TI - Effects of physical training in asthma: a systematic review. AB - OBJECTIVES: To assess the evidence for the effects of physical training on pulmonary function, symptoms, cardiopulmonary fitness, and quality of life in subjects with asthma. METHODS: A search was conducted for randomised controlled trials of subjects with asthma undertaking physical training using the Cochrane Airways Group register of controlled clinical trials, Medline, Embase, Sportdiscus, Science citation index, and Current contents index. Studies were included in the review if the subjects had asthma, were 8 years of age or older, and had undertaken physical training for at least 20 minutes per session, twice a week, for a minimum of four weeks. The eligibility of trials for inclusion in the review and the quality of the trials were independently assessed by two reviewers. RESULTS: Eight studies with a total of 226 subjects met the inclusion criteria for this review. Physical training had no effect on resting lung function but led to an improvement in cardiopulmonary fitness as measured by an increase in maximum oxygen uptake of 5.6 ml/kg/min (95% confidence interval 3.9 to 7.2). None of the studies measured quality of life. CONCLUSIONS: Physical training improves cardiopulmonary fitness without changing lung function. It is not clear if the improvement in fitness translates into a reduction in symptoms or an improvement in the quality of life. There is a need for further randomised controlled trials of the effects of physical training in the management of asthma. PMID- 10854015 TI - Aerobic responses of prepubertal boys to two modes of training. AB - OBJECTIVE: To investigate the effects of two contrasting eight week training programmes on the aerobic performance of 39 prepubescent boys (mean age 10.1 years). METHODS: All boys were volunteer subjects from three city schools and the schools were matched by a health related behaviour questionnaire. All of the boys were assessed as Tanner stage one for genitalia and pubic hair development. Criterion laboratory tests included peak VO2 as assessed by an incremental discontinuous treadmill test to voluntary exhaustion. Submaximal measurements of heart rate, minute ventilation (VE) and VO2 were also recorded during the treadmill test. One of the schools provided the control group (n = 14), and boys from the other schools followed two contrasting training programmes. The first was a sprint interval running programme (n = 12) comprising 10 second and 30 second sprints, and the second a continuous cycle ergometer programme (n = 13) maintaining a heart rate in the range 80-85% of maximum for 20 minutes on a Monark cycle ergometer. After eight weeks training three times a week, the three groups were retested. RESULTS: There were no significant differences in peak VO2 (p>0.05) with training in either of the groups. Neither were there significant changes in any of the submaximal variables VO2, VE, or heart rate (p>0.05). CONCLUSION: The findings of this study indicate that neither eight week sprint interval running nor continuous cycle ergometer training programmes significantly improve maximal or submaximal indicators of the aerobic performance of prepubertal boys. PMID- 10854016 TI - Suprascapular neuropathy in volleyball players. AB - BACKGROUND: Suprascapular nerve entrapment with isolated paralysis of the infraspinatus muscle is uncommon. However, this pathology has been reported in volleyball players. Despite a lack of scientific evidence, excessive strain on the nerve is often cited as a possible cause of this syndrome. Previous research has shown a close association between shoulder range of motion and strain on the suprascapular nerve. No clinical studies have so far been designed to examine the association between excessive shoulder mobility and the presence of this pathology. AIM: To study the possible association between the range of motion of the shoulder joint and the presence of suprascapular neuropathy by clinically examining the Belgian male volleyball team with respect to several parameters. METHODS: An electromyographic investigation, a clinical shoulder examination, shoulder range of motion measurements, and an isokinetic concentric peak torque shoulder internal/external rotation strength test were performed in 16 professional players. RESULTS: The electrodiagnostic study showed a severe suprascapular neuropathy in four players which affected only the infraspinatus muscle. In each of these four players, suprascapular nerve entrapment was present on the dominant side. Except for the hypotrophy of the infraspinatus muscle, no significant differences between the affected and non-affected players were observed on clinical examination. Significant differences between the affected and non-affected players were found for range of motion measurements of external rotation, horizontal flexion and forward flexion, and for flexion of the shoulder girdle (protraction); all were found to be higher in the affected players than the non-affected players. CONCLUSIONS: This study suggests an association between increased range of motion of the shoulder joint and the presence of isolated paralysis of the infraspinatus muscle in volleyball players. However, the small number of patients in this study prevents definite conclusions from being drawn. PMID- 10854017 TI - Immune function in female elite rowers and non-athletes. AB - OBJECTIVE: To compare immune function in female rowers and controls in the resting state, and then correlate the results with a two month history of upper respiratory tract infection (URTI). METHODS: Subjects included 20 elite female rowers located at the ARCO Olympic Training Centre in Chula Vista, California, and 19 non-athletic female controls. These two groups were compared cross sectionally for immune function and infection rates. RESULTS: Granulocyte/monocyte phagocytosis, oxidative burst activity, and plasma cytokine concentrations (interleukin-6, tumour necrosis factor-alpha, and interleukin-1 receptor antagonist) did not differ significantly between groups. Phytohaemagglutinin induced lymphocyte proliferative response (adjusted whole blood method) was significantly higher (31% and 36% for optimal and suboptimal concentrations respectively) in rowers than in controls. Natural killer cell activity was substantially higher (1.6-fold for total lytic units) in the female rowers than in controls. Two month health logs disclosed 5.2 (1.2) and 3.3 (1.1) days with URTI symptoms for the rowers and controls respectively (p = 0.268). For all 39 subjects combined, and for the 20 rowers separately, none of the immune parameters correlated significantly with number of days with URTI symptoms. CONCLUSIONS: In this cross sectional comparison of elite female rowers and non athletes, a group difference was found for natural killer cell activity and phytohaemagglutinin induced proliferative response (whole blood technique), but not other measures of immune function. The number of days with URTI symptoms during the spring season did not differ between groups, and variations in blood measures of immunity were unrelated to URTI. PMID- 10854018 TI - Sport, age, and sex specific incidence of sports injuries in Western Australia. AB - OBJECTIVE: To describe the trends in recreational sports injury in Perth, Western Australia. DESIGN: A prospective cohort study of sports injuries during the 1997 winter season (May to September). SETTING: Sample of Australian football, field hockey, basketball, and netball players from the Perth metropolitan area, Western Australia. METHODS: A cohort of sports participants was followed over the five month winter sports season. Before the season, participants completed a baseline questionnaire and during the season were interviewed every four weeks by telephone. RESULTS: Overall, 92% of participants (n = 1391) who completed a baseline questionnaire completed at least one follow up telephone interview. About half (51%) of the cohort sustained one or more injuries during the winter season accounting for a total of 1034 injuries. Most injuries were of moderate (58%, n = 598) or minor (40%, n = 412) severity, with only 3% (n = 24) requiring emergency department treatment or a hospital stay. The injury incidence rate was greatest for football (20.3/1000 hours of participation), similar for field hockey and basketball (15.2/1000 hours and 15.1/1000 hours respectively), and lowest for netball (12.1/1000 hours). The incidence of injury was greatest in the first four weeks of the season, and participants aged between 26 and 30 years had about a 55% greater risk of injury than those aged less than 18 years. CONCLUSIONS: This is one of the first studies to show that recreational sports are safe. Although the likelihood of injury was greatest in the first month of the season, few injuries required admission to hospital or emergency department treatment. A greater emphasis on prevention in the early part of the season should help to reduce the elevated incidence of injury found at this time. PMID- 10854019 TI - Do high impact exercises produce higher tibial strains than running? AB - BACKGROUND: Bone must have sufficient strength to withstand both instantaneous forces and lower repetitive forces. Repetitive loading, especially when bone strain and/or strain rates are high, can create microdamage and result in stress fracture AIM: To measure in vivo strains and strain rates in human tibia during high impact and moderate impact exercises. METHODS: Three strain gauged bone staples were mounted percutaneously in a rosette pattern in the mid diaphysis of the medial tibia in six normal subjects, and in vivo tibial strains were measured during running at 17 km/h and drop jumping from heights of 26, 39, and 52 cm. RESULTS: Complete data for all three drop jumps were obtained for four of the six subjects. No statistically significant differences were found in compression, tension, or shear strains with increasing drop jump height, but, at the 52 cm height, shear strain rate was reduced by one third (p = 0.03). No relation was found between peak compression strain and calculated drop jump energy, indicating that subjects were able to dissipate part of the potential energy of successively higher drop jumps by increasing the range of motion of their knee and ankle joints and not transmitting the energy to their tibia. No statistically significant differences were found between the principal strains during running and drop jumping from 52 cm, but compression (p = 0.01) and tension (p = 0.004) strain rates were significantly higher during running. CONCLUSIONS: High impact exercises, as represented by drop jumping in this experiment, do not cause higher tibial strains and strain rates than running and therefore are unlikely to place an athlete who is accustomed to fast running at higher risk for bone fatigue. PMID- 10854020 TI - Red blood cell variables in highly trained pubescent athletes: a comparative analysis. AB - BACKGROUND: A suboptimal haematological status has often been recorded in athletes involved in intensive physical activity. There have even been reports of "sports anaemia" associated with intensive physical exercise. However, studies on the effect of different types of exercise practiced over a long period of time on the red blood cell variables in pubescent athletes are very few. AIM: To assess the basic red blood cell variables in highly trained pubescent athletes from different sports and to compare the results with those for a control untrained group. Sex related differences in these variables were also assessed. METHODS: 876 highly trained athletes (559 boys and 317 girls) were included in the study. Their mean (SEM) age, weight, and duration of training were: 14.01 (0.06) years, 56.24 (0.52) kg, and 3.52 (0.07) years respectively. The control group consisted of 357 untrained subjects (171 boys and 186 girls) with mean (SEM) age and weight of 14.58 (0.09) years and 57.75 (0.67) kg. The group of athletes was divided into seven subgroups according to the sport practiced: athletics (105), swimming (107), rowing (230), wrestling (225), weight lifting (47), various team sports (92), and other sports (67). Venous blood samples were drawn from the cubital vein, and the red blood cell count, packed cell volume, haemoglobin concentration, and mean corpuscular volume were measured. Statistical indices were computed for each group and for each variable, and analysis of variance factorial analysis was performed to evaluate the statistical significance of the differences detected. RESULTS: The highly trained group was found to have lower red blood cell count, packed cell volume, and haemoglobin concentration (p<0.001) than the control untrained group (4.61 (0.01) x 10(12)/l v 4.75 (0.02) x 10(12)/l, 0.389 (0.001) v 0.404 (0.002) l/l, and 133.01 (0.38) v 139.9 (0.62) g/l respectively). These variables were lower for the boys of the trained group than for the boys of the control group (p<0.001), and similarly for the girls (p<0.001). The lowest red blood cell count, packed cell volume, and haemoglobin concentration were measured in blood samples from the boys of the swimming subgroup (4.54 (0.06) x 10(12)/l, 0.386 (0.006) l/l, and 129.38 (1.80) g/l respectively) and the rowing subgroup (4.66 (0.03) x 10(12)/l, 0.400 (0.003) l/l, and 136.21 (0.94) respectively). The same distribution was found for the girls: lowest in the rowing subgroup (4.32 (0.04) x 10(12)/l, 0.314 (0.003) l/l, and 124.27 (0.93) g/l) and the swimming subgroup (4.40 (0.05) x 10(12)/l, 0.375 (0.005) l/l, and 125.90 (1.30) g/l). No differences were found in the mean corpuscular volume. CONCLUSIONS: Continuous (more than one year) high intensity sports training (twice a day/five days a week) results in a decrease in the basic red blood cell variables in pubescent boys and girls, this being most pronounced in the submaximal sports. PMID- 10854021 TI - Medical provision and usage for the 1999 Everest marathon. PMID- 10854022 TI - Continuous and intermittent exposure to the hypoxia of altitude: implications for glutamine metabolism and exercise performance. PMID- 10854023 TI - Sports medicine? The ultimate folly. PMID- 10854024 TI - Anthropometric characteristics of elite male junior rowers. AB - During the 1997 Federation Internationale des Societes d'Aviron World Junior Rowing Championships, anthropometric data on 383 male junior rowers were assessed. With 430 participating athletes, the sample represented 89% of the population. In addition to age, 27 dimensions were measured: body mass, six heights or lengths, four breadths, 10 girths, and six skinfolds. The elite male junior rowers were tall (187.4 (5.8) cm; mean (SD)) and heavy (82.2 (7.4) kg), with larger length, breadth, and girth dimensions than a nationally representative sample of Belgian boys of the same chronological age. A rowing specific anthropometric profile chart with norms was constructed. The stature of the junior rowers is similar to that of adult heavyweight elite rowers, except that the junior rowers are lighter. Compared with non-finalists, finalists are heavier (but still lighter than the adult heavyweight elite rower) and taller, with greater length, breadth (except for the bicristal diameter), and girth dimensions. PMID- 10854025 TI - Jerry Morris: pathfinder for health through an active and fit way of life. PMID- 10854026 TI - Traumatic duodenal rupture in a soccer player. AB - Traumatic duodenal rupture resulting from blunt trauma during soccer is an extremely rare occurrence. A case report of this unusual condition is presented together with a review of the literature. PMID- 10854027 TI - Considerations for the design and analysis of experimental studies in physical activity and exercise promotion: advantages of the randomised controlled trial. PMID- 10854028 TI - Advanced life support. PMID- 10854029 TI - Is the sacroiliac joint mobile and how should it be treated? PMID- 10854030 TI - Epidemiology of knee injuries: diagnosis and triage. PMID- 10854031 TI - The ethics of boxing. PMID- 10854032 TI - The catalytic subunit of telomerase is expressed in developing brain neurons and serves a cell survival-promoting function. AB - Telomerase, a specialized reverse transcriptase (RT) linked to cell immortalization and cancer, has been thought not to be expressed in postmitotic cells. We now report that telomerase activity and its essential catalytic subunit, telomerase reverse transcriptase (TERT), are expressed in neurons in the brains of rodents during embryonic and early postnatal development, and are subsequently downregulated. Suppression of TERT expression in cultured embryonic hippocampal neurons increases their vulnerability to apoptosis and excitotoxicity. Overexpression of TERT in PC12 cells suppresses apoptosis induced by trophic factor withdrawal. TERT exerts its anti-apoptotic action at an early stage of the cell death process prior to mitochondrial dysfunction and caspase activation. TERT may serve a neuron survival-promoting function in the developing brain, and downregulation of TERT in the adult brain may contribute to increased neuronal vulnerability in various age-related neurodegenerative disorders. PMID- 10854033 TI - Ultrastructural analysis of neurosecretory cells in the antennae of the mosquito, Culex salinarius (Diptera: Culicidae). AB - An antiserum raised against the peptide, culetachykinin II, immunocytochemically detected a group of neurosecretory cells in the first flagellar segment of the antennae of both males and females of the mosquito, Culex salinarius. This is the first insect species in which neurosecretory cells have been found in the antennae. The ultrastructure of these antennal neurosecretory cells (ANC) is described, as well as their relationship to other neurons in the antennae and antennal lobe of the mosquito. These tachykinin-reactive cells contain relatively small (140-220 nm) elementary neurosecretory granules. Not only do the ANC have axons that terminate on specific glomeruli of the deutocerebrum, but these neurons also have collaterals that form neurohemal terminals in the receptor lymph channels of the dendrites of the sensory neurons. Thus, the ANC not only influence higher centers of the brain that interpret signals from the antennal sensillae, but also modulate the response of the sensory receptors. To our knowledge, this is the first report of neurosecretory cells directly affecting the signal reception of sensory neurons. PMID- 10854034 TI - Expression of neurotrophins BDNF and NT-3, and their receptors in rat brain after administration of antipsychotic and psychotrophic agents. AB - We have investigated the potential role of neurotrophic factors in antipsychotic drug action by examining the effects of antipsychotic and psychotropic treatments on the mRNA expression of brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), and their receptors, trkB and trkC, respectively, in rat brain. Neither acute nor chronic clozapine treatment significantly affected the expression of these mRNAs in any brain area investigated, except for a decrease in trkB expression in the granule cells of the olfactory bulb. We then examined the effects of the psychotropic agent MK-801. MK-801 (5 mg/kg; 4 h) significantly increased BDNF mRNA in the entorhinal cortex, but did not influence NT-3, trkB, or trkC expression in any brain area except for the olfactory bulb. The induction of BDNF mRNA by MK-801 was attenuated by pre-treatment (1 h prior to MK-801 administration) with the antipsychotics, clozapine (25 mg/kg) and haloperidol (2 mg/kg), but not with the antidepressant desipramine (15 mg/kg). Finally, we confirmed that the effects of MK-801 on BDNF mRNA were reflected in the respective changes in BDNF protein levels: MK-801 significantly increased anti BDNF reactivity in the entorhinal cortex (126 +/- 7% of control) while concomitantly decreasing in the hippocampus (71 +/- 2% of control). These data do not support the hypothesis that neurotrophins play an important role in antipsychotic drug action, but rather suggest that induction of BDNF in the entorhinal cortex may play a significant role in the psychotropic action of MK 801. PMID- 10854035 TI - Molecular analysis of trkC in the cat visual cortex. AB - trkC belongs to the trk family of neurotrophin receptors. Several isoforms of trkC have been cloned to date; a full-length catalytic form containing a tyrosine kinase (TK) domain, three full-length isoforms with amino-acid insertions (14, 25, and 39 amino acids) in the TK domain, and five noncatalytic truncated forms that completely lack the TK domain. These isoforms have been studied in several mammalian species, including the pig, rat, mouse, monkey, and human. In this article we report the cloning and sequencing of five trkC isoforms isolated from 30-d postnatal cat visual cortex. The first isoform corresponded to the previously reported full-length trkC transcript containing the 14 amino-acid insert. To search for the presence of other inserts, reverse transcription polymerase chain reaction (RT-PCR) was performed on 30-d postnatal cat visual cortex mRNA using primers that flank the insertion site in the TK domain. Both the isoform containing the 14 amino-acid insert and the isoform lacking any insertion were present in abundant amounts, whereas the other two insert containing isoforms (TK25 and TK39) were much less abundant. The fifth isoform discovered corresponds to the previously reported truncated transcript. Overall, there is a high degree of identity (89-98%) and homology (97-99%) between the cat trkC nucleotide and amino-acid sequences among all mammals. The extracellular juxtamembrane domain was found to be highly divergent among all mammals that have been studied to date. This divergent region also included a proline deletion in the cat trkC sequence. This is the first report of the cloning, sequencing, and RT-PCR analysis of trkC in cat visual cortex, a system extensively studied using anatomical and physiological approaches. PMID- 10854036 TI - Changes in tissue-plasminogen activator mRNA expression following cortical ablation in the rat brain. AB - Tissue plasminogen activator (tPA) has been used to treat acute thrombotic lesions. Roles other than the activation of fibrinolytic pathways have been suggested for tPA in the mature brain. We used the in situ hybridization technique to investigate the changes in tPA mRNA expression within the brain after cortical ablation. We found that expression of tPA mRNA started to increase diffusely in the cortex ipsilateral to the injury 6 h after ablation. This increase had become prominent 24 h after ablation. On d 5, the expression of tPA mRNA had returned to that of the control animals except for the area near the injury. We also found that administration of MK-801 before injury suppressed the increase of tPA mRNA in the ipsilateral cortex. These results suggest that the increase in tPA mRNA is likely to be mediated via activation of NMDA receptors. PMID- 10854037 TI - A new concept in the pharmacology of neuroprotection. AB - Vasoactive intestinal peptide (VIP), originally discovered in the intestine as a peptide of 28 amino acids, was later found to be a major brain peptide having neuroprotective activities. To exert neuroprotective activity, VIP requires glial cells secreting neuroprotective proteins. Activity-dependent neurotrophic factor (ADNF) is a recently isolated factor secreted by glial cells under the action of VIP. This protein, isolated by sequential chromatographic methods, was named activity-dependent neurotrophic factor since it protected neurons from death associated with blockade of electrical activity. A fourteen-amino-acid fragment of ADNF (ADNF-14) and the more potent, nine-amino-acid derivative (ADNF-9), exhibit activity that surpasses that of the parent protein with regard to potency and a broader range of effective concentration. Furthermore, the peptides exhibit protective activity in Alzheimer's disease-related systems (e.g., beta-amyloid toxicity and apolipoprotein E deficiencies, genes that have been associated with Alzheimer's disease onset and progression). ADNP is another glial mediator of VIP associated neuroprotection. NAP, an eight-amino-acid peptide derived from ADNP (sharing structural and functional similarities with ADNF-9), was identified as the most potent neuroprotectant described to-date in an animal model of apolipoprotein E-deficiency (knock-out mice). These femtomolar-acting peptides form a basis for a new concept in pharmacology: femtomolar neuroprotection. PMID- 10854038 TI - NGF stimulation of erk phosphorylation is impaired by a point mutation in the transmembrane domain of trkA receptor. AB - The nerve growth factor (NGF) trkA receptor is a transmembrane glycoprotein composed of a large extracellular ligand-binding region connected to the cytoplasmic tyrosine kinase region by a single transmembrane domain (TMD). To explore the role of TMD in the process of receptor activation, we substituted the hydrophobic amino-acid residue valine 432 with the charged amino-acid glutamic acid (designated V432E mutant) by utilizing in vitro site-directed mutagenesis. NIH 3T3 cells lacking endogenous NGF receptors were stably transfected with a pRc/CMV vector carrying either wild-type (trkA) or mutated (V432E) receptors. Stable transfectants were shown, using 125I-NGF binding and Western-blot analysis, to express the trkA recombinant receptors. Scatchard analysis revealed similar affinity for NGF in wild-type and V432E receptors. Although the level of basal trkA receptor tyrosine phosphorylation was higher in the mutant than in the wild-type, NGF stimulation of WT 11 and V432E transfectants resulted in a rapid increase in receptor tyrosine phosphorylation and of its intracellular adaptor protein SHC. In contrast to WT 11, V432E mutants showed very low levels of NGF-, and moderate levels of FGF-induced erks phosphorylation, respectively. Collectively, these findings suggest that a single substitution (V432E) in the trkA TMD results in a selective impairment of trkA-mediated erks signaling pathway. PMID- 10854039 TI - Effects of citalopram on dopamine D2 receptor expression in the rat brain striatum. AB - Effects of citalopram on dopamine D2 receptor expression in the rat brain striatum were studied. Repeated administration of citalopram increased the amount of dopamine D2 receptors, the level of dopamine D2 receptor mRNA, and the transcription rate of the dopamine D2 receptor gene. Single administration of citalopram also increased the level of dopamine D2 receptor mRNA with a maximum effect in 2-4 h after the treatment, and the transcription rate of the dopamine D2 receptor gene. The administration of 5-hydroxytryptophan (5-HTP) also increased the level of dopamine D2 receptor mRNA. These results suggest that the increase in the dopamine D2 receptor expression induced by citalopram may be owing, at least partially, to the stimulation of the dopamine D2 receptor gene transcription, and that serotonin (5-HT) may mediate the effects of citalopram in the induction of dopamine D2 receptor expression. PMID- 10854040 TI - Elevated UTP and CTP content in cultured neurons from HPRT-deficient transgenic mice. AB - Hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8.; HPRT) catalyzes the salvage synthesis of inosine-5'-monophosphate (IMP) and guanosine-5' monophosphate (GMP) from the purine bases hypoxanthine and guanine, respectively. Complete deficiency of HPRT activity is associated with the Lesch-Nyhan syndrome (LNS), characterized by excessive purine production and severe neurological manifestations. The etiology of the metabolic consequences of HPRT deficiency is clarified, but that of the neurological manifestations is not yet understood. HPRT-deficient mice represent an experimental animal model of LNS. In search for a possible metabolic abnormality in LNS brains, connecting the neurological deficit to HPRT deficiency, the purine and pyrimidine nucleotide content of cultured neurons, prepared from HPRT-deficient transgenic mice, was now determined. The HPRT-deficient neuronal cultures exhibited a significantly elevated content of the pyrimidine nucleotides UTP (1.33-fold the normal level, p = 0.0002) and CTP (1.28-fold the normal level, p = 0.02), but normal content of the purine nucleotides ATP and GTP. This abnormality in neuronal pyrimidine nucleotide content may be associated with the pathophysiology of the neurological deficit in LNS. PMID- 10854041 TI - Synaptogenesis and myopathy under acetylcholinesterase overexpression. AB - Environmental, congenital, and acquired immunological insults perturbing neuromuscular junction (NMJ) activity may induce a variety of debilitating neuromuscular pathologies. However, the molecular elements linking NMJ dysfunction to long-term myopathies are unknown. Here, we report dramatically elevated levels of mRNA encoding c-Fos and the "readthrough" (R) variant of acetylcholinesterase (AChE) in muscles of transgenic mice overexpressing synaptic (S) AChE in motoneurons and in control mice treated with the irreversible cholinesterase inhibitor diisopropylfluorophosphonate (DFP). Tongue muscles from DFP-treated and AChE-S transgenic mice displayed exaggerated neurite branching and disorganized, wasting fibers. Moreover, diaphragm muscles from both transgenic and DFP-treated mice exhibited NMJ proliferation. 2'-O-methyl protected antisense oligonucleotides targeted to AChE mRNA suppressed feedback upregulation of AChE and ameliorated DFP-induced NMJ proliferation. Our findings demonstrate common transcriptional responses to cholinergic NMJ stress of diverse origin, and implicate deregulated AChE expression in excessive neurite outgrowth, uncontrolled synaptogenesis, and myopathology. PMID- 10854042 TI - Cloning and analysis of a murine PIAS family member, PIASgamma, in developing skin and neurons. AB - Signal transducer and activator of transcription (STAT) proteins are latent cytoplasmic transcription factors that become activated in response to stimulation by various cytokines. Recently a new family of five structurally related proteins, called PIAS (Protein Inhibitor of Activated STAT) has been identified as potentially important downregulators of this pathway. Members of the PIAS family of STAT inhibitors may play a prominent role in the downregulation of STAT-mediated signaling processes. In this article we describe the isolation of the cDNA and expression of the gene for the murine homologue of the human STAT inhibitor family member PIASgamma. The cDNA for mPIASgamma encodes a protein of 507 amino acids that is highly homologous to the human protein and is expressed in the mouse as early as d 7.5 of gestation. In situ hybridizations of staged mouse embryos localized the transcript for the PIASgamma gene to the limbs, neuroepithelium, and the inner root sheath of the hair follicle, suggesting a role in the development of these structures. Immunostaining studies with a polyclonal antibody (PAb) recognizing human PIASgamma localized the protein in the hair follicle of human scalp hair and in monkey neuronal cells. Thus PIASgamma exhibits a highly selective pattern of expression, suggesting that it modulates the response of cells to developmental cues. PMID- 10854043 TI - Introduction: the role of combination therapy in modern antihypertensive therapy. PMID- 10854044 TI - Do we effectively lower blood pressure? AB - Blood pressure control is needed in order to attain a normalization of the increased cardiovascular risk present in the hypertensive population. However, we are far from obtaining the most effective control of blood pressure. A more aggressive attitude, in particular in high-risk patients, is required, together with an adequate selection of antihypertensive pharmacological therapies. In this sense combined therapy has to be more widely used in patients presenting with the characteristics detailed in the newly published mega-trials like the Hypertension Optimal Treatment study. An adequate selection of the components of combination could also be of interest through a better protective capacity mediated by blood pressure-independent effects. PMID- 10854045 TI - The importance of sustained blood pressure control. AB - The case for antihypertensive drug regimens that produce consistent 24-h blood pressure control has largely been founded upon a series of epidemiological observations that were either cross-sectional or alternatively relatively small scale follow-up studies. More recent data have unequivocally demonstrated, in a prospective study in hypertensive subjects with left ventricular hypertrophy, that the reduction in left ventricular mass index during the course of one year's antihypertensive treatment, was predicted much more closely by treatment-induced changes in ambulatory blood pressure than by changes in clinic blood pressure. This provides definitive and confirmatory data to support the aim of achieving blood pressure control, which is based upon a smooth and consistent antihypertensive effect over a full 24-h dosage interval. Regimens which provide such control may also offer the advantage of a sustained duration of effect beyond 24 h. This characteristic is attractive because even the most compliant patient may inadvertently miss at least one dose of medication each week. Evidence from a number of studies which have sought to mimic this pattern of suboptimal compliance by deliberately inserting a placebo phase into a steady state treatment regimen, has clearly demonstrated the benefits of antihypertensive drugs with intrinsically long duration of action. Furthermore, there is evidence to suggest that following cessation of therapy there is a biphasic reversion of blood pressure towards baseline levels with a maintenance of a residual effect which is more pronounced with a long-acting agent when compared to a shorter-acting drug from the same therapeutic class. There is increasing evidence, albeit not derived from prospective outcome studies, that indicates that the benefits of antihypertensive therapy are likely to be maximized by treatment regimens which result in sustained blood pressure control. PMID- 10854046 TI - How can hypertensive patients be better treated? The contribution of combination therapy. AB - Recent studies demonstrated that target blood pressure (BP) in treated hypertensive patients should be below 140 mmHg for systolic blood pressure (SBP) and below 90 mmHg for diastolic blood pressure (DBP). However, population studies from several countries have demonstrated that in clinical practice the proportion of controlled hypertensive patients is less than 30%. In order to elucidate these questions in France we analysed a large population of 145,000 subjects examined at the Centre d'Investigations Preventives et Cliniques in Paris (IPC). Among those with high BP at the time of their IPC visit, only 20% received an antihypertensive treatment. Among those receiving an antihypertensive treatment, less than 27% (24% in men and 30% in women) presented with BP values less than 140 mmHg for SBP and less than 90 mmHg for DBP. This analysis also showed that 72% of hypertensive patients presented with at least one modifiable associated cardiovascular risk factor and that more than 30% of hypertensive men and more than 25% of hypertensive women presented with at least two associated risk factors. The use of combination therapies could help to increase the percentage of well-controlled hypertensive subjects. It has been shown that in order to reach this BP level, combination therapy should be used in more than two-thirds of the treated subjects. The trandolapril-verapamil combination is the first fixed combination of an angiotensin-converting enzyme inhibitor and a non dihydropyridine calcium-channel blocker. Administered once daily, this combination reduces BP more than a classic monotherapy. The effects of the trandolapril-verapamil combination on risk factors are either neutral (metabolic parameters), or even beneficial (reduction in heart rate). PMID- 10854047 TI - Cardiac effects of combination therapy in hypertension. AB - Control of hypertension and treatment of concomitant pathophysiologic conditions require use of multiple drugs in more than half of all patients. Unfortunately, most studies dealing with cardiovascular effects of antihypertensive drugs have focused on monotherapy. Thus, our knowledge of combination therapy in the treatment of hypertension is, to a great extent, extrapolation from monotherapy. Angiotensin-converting enzyme inhibitors in combination with calcium antagonists should be particularly efficacious in reducing left ventricular hypertrophy. Drug classes that either stimulate the renin-angiotensin system or the sympathetic nervous system are less likely to reduce left ventricular hypertrophy and should be avoided. In hypertensive patients with left ventricular dysfunction, beta blockers should be combined with angiotensin-converting enzyme inhibitor, whereas in the postmyocardial ischemia patient, verapamil and angiotensin-converting enzyme inhibitors may exert some additional beneficial effects with regard to reinfarction rates. Some of the undesirable metabolic side-effects of diuretics can be mitigated by the addition of an angiotensin-converting enzyme inhibitor or an angiotensin-receptor blocker. Given that two drugs when used separately are beneficial in a disorder does not necessarily mean that their combination is equally or even more beneficial. Sometimes combination therapy is used to counteract different limbs of pathophysiologic cascade operative in hypertensive patients with complications or comorbidities. PMID- 10854048 TI - How to improve adherence with prescribed treatment in hypertensive patients? AB - Low adherence of hypertensive patients to prescribed antihypertensive medications is a major cause of unsatisfactory blood pressure control. Several factors might have a negative influence on long-term adherence with treatment, for example a poor patient-doctor relationship and the presence of drug-induced side-effects. Various strategies are recommended in order to improve patient compliance, including educational programmes, self-measurement of blood pressure and monitoring of compliance. All methods may be helpful to encourage the patient to take the prescribed medication(s) regularly. It is also important to find a drug regimen which is at the same time simple, efficacious and well tolerated. Finally it should be pointed out that the motivation of the patient to follow the treatment requires the doctor to be equally motivated. PMID- 10854049 TI - Rab3B regulates ZO-1 targeting and actin organization in PC12 neuroendocrine cells. AB - Rab3B is a monomeric GTPase that modulates norepinephrine secretion when expressed in PC12 neuroendocrine cells. In the present study we determined whether rab3B also regulates the organization of intercellular junctions, since this GTPase localizes to regions of cell contact in multiple cell types. The stable expression of rab3B, but not the closely related rab3A, led to two morphological phenotypes in PC12 cells: (i) reorganization of F-actin into long filopodia and (ii) redistribution of the junction-associated protein ZO-1. ZO-1 localization was not appreciably affected by the expression of a GTP binding mutant of rab3B (N135I) that stimulates norepinephrine secretion by PC12 cells. The apparent diversity of these rab3B phenotypes implies that this GTPase is capable of influencing cell signaling pathways that in turn modulate the cytoskeleton and junction organization. In support of this hypothesis we observed that rab3B expression also altered the profile of proteins that interact with the signaling molecule, phosphatidylinositol 3-kinase (PI3-kinase). The effect of rab3B on protein interactions with PI3-kinase was reversed by inhibitors of this kinase. Furthermore, PI3-kinase inhibitors virtually abolished ZO-1 localization at the surfaces of cells that express rab3B, but not rab3A, whereas these inhibitors had no effect on rab3B-dependent norepinephrine secretion. Our results indicate that rab3B can influence junctional protein targeting and secretion by distinct mechanisms. PMID- 10854050 TI - Centrosomal and cytoplasmic Cdc2/cyclin B1 activation precedes nuclear mitotic events. AB - The activation of cdc2/cyclin B is the trigger for entry into mitosis. The mechanism of cdc2/cyclin B activation is complex, but the final step is the dephosphorylation of the Thr14 and Tyr15 residues on the cdc2 subunit, catalyzed by a member of the Cdc25 family of phosphatases. Cdc2/cyclin B1 accumulates at the centrosome in late G2 phase and has been implicated in the conversion of the centrosome from an interphase to a mitotic microtubule organizing center. Here we demonstrate biochemically that cdc2/cyclin B1 accumulates at the centrosome in late G2 as the inactive, phosphotyrosine 15 form and that the centrosomal cdc2/cyclin B1 can be activated in vitro by recombinant cdc25B. We provide evidence that a portion of the cdc2/cyclin B1 translocated into the nucleus in prophase is the inactive tyrosine-15-phosphorylated form. At this time the centrosomal and cytoplasmic cdc2/cyclin B1 is already active. This provides evidence that the activation of cdc2/cyclin B1 is initiated in the cytoplasm and that full activation of the translocated pool occurs in the nucleus. PMID- 10854051 TI - Cloning and characterization of DIP1, a novel protein that is related to the Id family of proteins. AB - Using human cyclin D1 as the "bait" in a yeast two-hybrid system, together with a HL60 cDNA library, we identified a novel human nuclear protein designated DIP1. This protein is expressed in a variety of cell types, and in fibroblasts its level remains constant throughout the cell cycle. However, the level of this protein increases severalfold during the differentiation of HL60 cells. The DIP1 protein can be phosphorylated in vitro by a cellular kinase and this activity reaches its maximum in extracts obtained from cells in the G1 phase of the cell cycle. DIP1 contains a helix-loop-helix motif but lacks an adjacent basic DNA binding domain, thus resembling the Id family of proteins. The dip1 gene is located on human chromosome 16p11.2-12, a locus that is amplified in several types of human cancer. These results suggest that DIP1 may be involved in the control of gene expression and differentiation, but its precise function remains to be determined. PMID- 10854052 TI - Late endocytic compartments are major sites of annexin VI localization in NRK fibroblasts and polarized WIF-B hepatoma cells. AB - Annexin VI is an abundant calcium- and phospholipid-binding protein whose intracellular distribution and function are still controversial. Using a highly specific antibody, we have studied the distribution of annexin VI in NRK fibroblasts and the polarized hepatic cell line WIF-B by confocal microscopy. In NRK cells, annexin VI was almost exclusively found associated with endocytic compartments, which were defined by their ability to receive fluid-phase marker internalized from the cell surface. However, extensive colocalization of annexin VI and the endocytic marker was only observed after about 45 min, indicating that annexin VI was primarily in late endocytic compartments or (pre)lysosomes. Consistent with this, annexin VI was predominantly seen on structures that contained the lysosomal protein lgp120, although not on dense core lysosomes by electron microscopy. Two major populations of annexin VI-containing structures were present in polarized WIF-B hepatocytes. One correlated to lgp120-positive (pre)lysosomes and was still observed after treatment with brefeldin A (BFA), while the other appeared to be partially associated with Golgi membranes and was BFA-sensitive. The striking association with prelysosomal compartments in NRK and WIF-B cells suggests that annexin VI could play a role in fusion events in the late endocytic pathway, possibly by acting as a tether between membranes. PMID- 10854053 TI - Ubiquitin-dependent protein processing controls radiation-induced apoptosis through the N-end rule pathway. AB - The ubiquitination of nuclear proteins activated in human lymphocytes undergoing radiation-induced apoptosis and the subsequent downstream proteasomal protein processing, shown to be involved in apoptotic death control, may be dependent on an amino-terminal sequence identity of ubiquitin target proteins, the "N-end rule" pathway. Here we report that this selective pathway controls radiation induced apoptosis and that it is involved in the initiation of this type of cell death. Dipeptide competitors of protein ubiquitination/processing dependent solely on the basic amino-terminal residues (type I) efficiently inhibited the radiation-induced apoptotic death phenotype, indicating that only the substrates of ubiquitination with basic NH2-terminal amino acids are involved in apoptotic death control. This selective inhibition was followed by an early, overall but also target-specific inhibition of ubiquitination and by an activation and stabilization of poly(ADP-ribose) polymerase (PARP) that occurs through inhibition of ubiquitination of its cleaved form (85 kDa). Interestingly, caspases-3 and -7 were not activated following irradiation, further suggesting that PARP cleavage may be regulated by an N-end rule pathway in a caspase independent manner. These results highly suggest involvement of this subset of the ubiquitin system in the apoptotic death control and in the specific regulation of PARP activity. PMID- 10854054 TI - Heat shock protein 70 inhibits caspase-dependent and -independent apoptosis in Jurkat T cells. AB - Heat shock protein 70 (hsp70) is a stress-inducible protein that prevents apoptosis induced by a wide range of cytotoxic agents by an as yet undefined mechanism. The caspase family of cysteine proteases have been attributed a central role in the execution of apoptosis. However, several cases of caspase independent apoptosis have been recently reported, suggesting that caspases may not be necessary for apoptosis in all cells. This study examines the protective role of hsp70 in both caspase-dependent and -independent apoptosis. Hydrogen peroxide (H2O2) used at low and high concentrations in Jurkat T cells induces caspase-dependent and -independent apoptosis, respectively. A hsp70-transfected Jurkat clone was used to observe the protection mediated by hsp70 during these two forms of apoptosis. Results reveal that hsp70 inhibits both caspase-dependent and -independent apoptosis. Furthermore, measurement of caspase-3 activity during caspase-dependent apoptosis revealed that caspase activation was inhibited in hsp70 transfectants. Early apoptotic events, such as mitochondrial depolarization, cytochrome c release, and increased intracellular calcium, were demonstrated to be common to both caspase-dependent and -independent H2O2-induced apoptosis. The inhibition of these events by hsp70 suggests that hsp70 may be an important anti-apoptotic regulator, functioning at a very early stage in the apoptotic pathway. PMID- 10854055 TI - High-molecular-weight serum protein complexes differentially promote cell migration and the focal adhesion localization of the urokinase receptor in human glioma cells. AB - The distribution of the urokinase-type plasminogen activator receptor (uPAR) on human glioma cells was examined as a function of culture conditions, using immunofluorescence and immunophotoelectron microscopy. Both uPAR colocalization with focal adhesion proteins and glioma cell motility were maximal in medium containing whole serum or a serum fraction retained by a 500,000 mol wt cutoff centrifugal concentration filter. High motility also took place in medium containing a serum fraction passed by the 500,000 cutoff filter but retained by a 100,000 cutoff filter and in minimal medium containing added vitronectin; however, under these conditions only a small percentage of the otherwise abundant focal adhesions contained colocalized uPAR. Glioma cells in minimal medium with added laminin migrated with a highly elongated morphology but without either classical focal adhesions or well-defined uPAR labeling. In contrast, glioma cells in minimal medium with no additions did not migrate, nor did they adhere well or display defined labeling patterns for focal adhesion proteins or uPAR. The results indicate that high-molecular-weight serum protein complexes promote both uPAR-focal adhesion colocalization and cell migration in glioma cells. However, conditions can be selected in which migration takes place with minimal uPAR-focal adhesion localization, as well as in the absence of apparent focal adhesions. PMID- 10854056 TI - Caspase-dependent and -independent events in apoptosis induced by hydrogen peroxide. AB - To define the role of caspase-3 in H2O2-induced apoptosis, we introduced caspase 3 cDNA into MCF-7 breast carcinoma cells that otherwise lack caspase-3 expression. H2O2 treatment induced DNA fragmentation and nuclear condensation in the caspase-3-expressing cells, but not in the caspase-3-deficient cells. This indicated that caspase-3 is essential for nuclear events. However, H2O2 induced an externalization of membrane phosphatidylserine (PS) and cell death regardless of caspase-3 expression. These events were not suppressed by Ac-DEVD-CHO and Z VAD-fmk, which inhibit DEVD-specific caspases and a broad spectrum of caspases, respectively. In Jurkat T cells, these inhibitors abolished H2O2-induced PS relocalization, but not cell death. Therefore, caspases appear to be dispensable for lethality by H2O2, but required for PS redistribution in a cell-type-specific manner. PMID- 10854057 TI - Activation of H-ras61L-specific signaling pathways does not require posttranslational processing of H-ras. AB - We have previously demonstrated that H-ras61L retained transforming activity when lacking C-terminal lipid modifications, provided that plasma membrane localization was restored by an N-terminal transmembrane domain. Since several ras-activated pathways contribute to the transformed phenotype, we utilized a novel set of transmembrane domain-anchored H-ras derivatives to examine if lipids are required for activation of any specific signaling pathways. We demonstrate here that H-ras61L-induced activation of the Raf/MEK/MAPK pathway, including recruitment of Raf to the plasma membrane and activation of Raf and MAPK, does not require C-terminal processing of H-ras61L. Biochemical fractionation experiments confirm the localization of TM-ras derivatives to the plasma membrane, as well as the ras-mediated recruitment of c-Raf-1. Changes in the actin cytoskeleton, controlled by H-ras61L-mediated activation of the Rac/ Rho pathway, as well as PI 3-kinase activation, can also occur in the absence of C terminal lipid modifications. Finally, downstream events, such as the induction of the immediate-early gene c-fos or neurite outgrowth in PC12 cells, are stimulated by the expression of plasma membrane-anchored, nonlipidated H-ras6lL. These results demonstrate that H-ras can be functionally targeted to the plasma membrane using a transmembrane domain sequence and that several signal transduction pathways downstream of H-ras can be activated without the presence of normal lipid modifications. PMID- 10854058 TI - Clusterin protects granulosa cells from apoptotic cell death during follicular atresia. AB - Clusterin expression is associated with programmed cell death (apoptosis) in many cell types but its exact role has not yet been defined. This study was carried out to determine the cellular localization of clusterin in the ovary and its functional role in the apoptotic cell death of ovarian follicles. A homogenous population of healthy and atretic follicles was obtained by treating immature rats with pregnant mare serum gonadotropin (PMSG). Apoptotic cell death was evaluated by TUNEL. Clusterin expression in the healthy and atretic follicles was examined by immunohistochemical and Western blot analyses, and gene expression was examined by Northern blot analysis. Clusterin protein and its mRNA are only expressed in granulosa cells of atretic follicles obtained from PMSG-treated rats on day 5 of the treatment. Healthy follicles from PMSG-treated rats on day 2 of the treatment do not express clusterin. Theca and stroma cells of both healthy and atretic follicles showed no signs of apoptosis and did not express clusterin. Withdrawal of trophic support from granulosa cells in cultures to induce apoptosis resulted in a dramatic increase in the levels of clusterin and its mRNA compared to cells cultured in serum-supplemented medium. In an attempt to establish the functional role of clusterin in the apoptotic cell death of ovarian follicles, the biosynthesis of clusterin in granulosa cells of healthy follicles was blocked by treatment of cells with antisense oligonucleotide to its cDNA. Treatment of granulosa cells with the antisense oligonucleotide resulted in an increase in the apoptotic cell death compared to the control. These findings indicate that depletion of clusterin can lead to the programmed cell death in ovary, suggesting a functional role for this protein in follicular atresia. PMID- 10854059 TI - Induction of apoptosis in rat cardiocytes by A3 adenosine receptor activation and its suppression by isoproterenol. AB - The purpose of the present study was to investigate the mechanisms involved in the induction of apoptosis in newborn cultured cardiomyocytes by activation of adenosine (ADO) A3 receptors and to examine the protective effects of beta adrenoceptors. The selective agonist for A3 ADO receptors Cl-IB-MECA (2-chloro-N6 iodobenzyl-5-N-methylcarboxamidoadenosine) and the antagonist MRS1523 (5-propyl-2 ethyl-4-propyl-3-(ethylsulfanylcarbonyl)-6-phenylpy rid ine-5-carboxylate) were used. High concentrations of the Cl-IB-MECA (> or = 10 microM) agonist induced morphological modifications of myogenic cells, such as rounding and retraction of cell body and dissolution of contractile filaments, followed by apoptotic death. In addition, Cl-IB-MECA caused a sustained and reversible increase in [Ca2+]i, which was prevented by the selective antagonist MRS1523. Furthermore, MRS1523 protected the cardiocytes if briefly exposed to Cl-IB-MECA and partially protected from prolonged (48 h) agonist exposure. Apoptosis induced by Cl-IB-MECA was not redox-dependent, since the mitochondrial membrane potential remained constant until the terminal stage of cell death. Cl-IB-MECA activated caspase-3 protease in a concentration-dependent manner after 7 h of treatment and more effectively after 18 h of exposure. Bcl-2 protein was readily detected in control cells, and its expression was significantly decreased after 24 and 48 h of treatment with Cl-IB-MECA. Beta-adrenergic stimulation antagonized the pro apoptotic effects of Cl-IB-MECA, probably through a cAMP/protein kinase A independent mechanism, since addition of dibutyryl-cAMP did not abolish the apoptosis induced by Cl-IB-MECA. Incubation of cultured myocytes with isoproterenol (5 microM) for 3 or 24 h almost completely abolished the increase in [Ca2+]i. Prolonged incubation of cardiomyocytes with isoproterenol and Cl-IB MECA did not induce apoptosis. Our data suggest that the apoptosis-inducing signal from activation of adenosine A3 receptors (or counteracting beta adrenergic signal) leads to the activation of the G-protein-coupled enzymes and downstream pathways to a self-amplifying cascade. Expression of different genes within this cascade is responsible for orchestrating either cardiomyocyte apoptosis or its protection. PMID- 10854060 TI - The human cyclin B1 protein modulates sensitivity of DNA mismatch repair deficient prostate cancer cell lines to alkylating agents. AB - DNA damage caused by alkylating agents results in a G2 checkpoint arrest. DNA mismatch repair (MMR) deficient cells are resistant to killing by alkylating agents and are unable to arrest the cell cycle in G2 phase after alkylation damage. We investigated the response of two MMR-deficient prostate cancer cell lines DU145 and LNCaP to the alkylating agent MNNG. Our studies reveal that DU145 cancer cells are more sensitive to killing by MNNG than LNCaP. Investigation of the underlying reasons for lower resistance revealed that the DU145 cells contain low endogenous levels of cyclin B1. We provide direct evidence that the endogenous level of cyclin B1 modulates the sensitivity of MMR-deficient prostate cancer cells to alkylating agents. PMID- 10854061 TI - Functional role of cyclin A on induction of fibroblast apoptosis due to ligation of CD44 matrix receptor by anti-CD44 antibody. AB - Ligation of cell surface matrix adhesion receptors such as integrins can increase expression of specific cell cycle regulatory proteins such as cyclin A, thereby regulating cell cycle progression. Disruption of cell surface matrix receptor interaction with the extracellular matrix can trigger apoptosis. Induction of apoptosis has been linked to unscheduled up-regulation of cyclin A and activation of cyclin-A-associated dependent kinase 2 activity due to cleavage of cyclin dependent kinase inhibitors by caspases. We have found that ligation of the cell surface matrix adhesion receptor CD44 by anti-CD44 antibody induces cell detachment and triggers apoptosis. In this report we show that ligation of CD44 by anti-CD44 antibody increases the expression of cyclin A protein prior to activation of caspase-3-like activity and morphological changes of apoptosis. Down-regulation of cyclin A protein levels by cyclin A antisense oligonucleotides dramatically decreased fibroblast apoptosis in response to anti-CD44 antibody. These data identify an important functional role of cyclin A in the induction of fibroblast apoptosis due to the ligation of the cell surface adhesion receptor CD44 by anti-CD44 antibody. PMID- 10854062 TI - Cdk7- and Cdc25A-independent dephosphorylation of Cdk2 during phorbol ester mediated cell cycle arrest in U937 cells. AB - The molecular mechanism underlying protein kinase C (PKC)-mediated cell cycle arrest is poorly understood. We undertook to characterize phorbol ester-activated PKC-mediated cell cycle arrest. Treatment with phorbol ester inhibited cell growth of human histiocytic lymphoma U937 cells with 83% of the cells arrested in G1 phase. Reduced activity of cdk2 correlated with cdk2 dephosphorylation and accumulation of cdk2 inhibitor p21Waf in phorbol ester-treated cells. Dephosphorylation of cdk2 was not associated with cdk7 and cdc25A activity in phorbol ester-treated cells. Protein phosphatase inhibitor assays suggest that the dephosphorylation of cdk2 results in the activation of a specific protein tyrosine phosphatase. Thus, dephosphorylation of cdk2 as well as accumulation of cdk2 inhibitor is likely to contribute to the G1 phase arrest in phorbol ester treated in U937 cells. PMID- 10854063 TI - Nuclear distribution of prothymosin alpha and parathymosin: evidence that prothymosin alpha is associated with RNA synthesis processing and parathymosin with early DNA replication. AB - Prothymosin alpha and parathymosin are two ubiquitous small acidic nuclear proteins that are thought to be involved in cell cycle progression, proliferation, and cell differentiation. In an effort to investigate the molecular function of the two proteins, we studied their spatial distribution by indirect immunofluorescence labeling and confocal scanning laser microscopy in relation to nuclear components involved in transcription, translation, and splicing. Results indicate that both proteins exhibit a punctuated nuclear distribution and are excluded by nucleoli. The distribution of prothymosin alpha in the nucleus is related to that of transcription sites, whereas the distribution of parathymosin correlates with early replication sites. This implies that prothymosin alpha and parathymosin are involved in transcription and replication, respectively. In addition to the punctate distribution, prothymosin alpha also is found concentrated in 1-6 nuclear domains per cell. These domains are found in more than 80% of randomly growing T24 human bladder carcinoma cells. They have a diameter of 0.2-2.5 microm, their size being inversely related to the number of domains per cell. The domains disappear during mitosis and the protein is excluded from the metaphase chromosomes. Double-labeling experiments associate these prothymosin alpha domains with PML and CstF64 containing nuclear bodies, but not with hnRNP-I containing domains or coiled bodies. PMID- 10854064 TI - Is beta-galactosidase staining a marker of senescence in vitro and in vivo? AB - Cytochemically detectable beta-galactosidase (beta-gal) at pH 6.0 has been reported to increase during the replicative senescence of fibroblast cultures and has been used widely as a marker of cellular senescence in vivo and in vitro. In this study, we have characterized changes in senescence-associated (SA) beta-gal staining in early and late passage cultures, cultures established from donors of different ages, virally immortalized cells, and tissue slices obtained from donors of different ages. The effects of different culture conditions were also examined. While we confirm the previous report that SA beta-gal staining increased in low-density cultures of proliferatively senescent cells, we were unable to demonstrate that it is a specific marker for aging in vitro. Cultures established from donors of different ages stained for SA beta-gal activity as a function of in vitro replicative age, not donor age. We also failed to observe any differences in SA beta-gal staining in skin cells in situ as a marker of aging in vivo. The level of cytochemically detectable SA beta-gal was elevated in confluent nontransformed fibroblast cultures, in immortal fibroblast cultures that had reached a high cell density, and in low-density, young, normal cultures oxidatively challenged by treatment with H2O2. Although we clearly demonstrate that SA beta-gal staining in cells is increased under a variety of different conditions, the interpretation of increased staining remains unclear, as does the question of whether the same mechanisms are responsible for the increased SA beta gal staining observed in senescent cells and changes observed in cells under other conditions. PMID- 10854065 TI - A novel rap1/B-Raf/14-3-3 theta protein complex is formed in vivo during the morphogenetic differentiation of postmeiotic male germ cells. AB - The 14-3-3 family of proteins is expressed in a broad range of organisms and tissues. Based on data essentially obtained with tissue culture cells and yeast, 14-3-3 proteins have been implicated as potential regulators of diverse signaling pathways, in particular those involving the activity of the Raf family protein kinases. The 14-3-3 theta mouse isoform is expressed almost exclusively in testis and brain. In an effort to understand the function of 14-3-3 theta in testis, we sought to identify endogenous proteins that interact with 14-3-3 theta in spermatogenic cells. A recombinant 14-3-3 theta fusion protein was used in Far Western and GST pulldown assays. Here we report that 14-3-3 theta interacts in vivo and in vitro with 93- to 95-kDa B-Raf, originally described as specific of neural tissues and never reported in male germ cells. Moreover, in mouse spermatids, i.e., the haploid cytodifferentiating cells, a so far unidentified protein complex formed by endogenous Rap1/B-Raf/14-3-3 theta can be coimmunoprecipitated. The intracellular localization of endogenous 14-3-3 theta, B-Raf, and Rap1 was analyzed in distinct spermatogenic cell types and a peculiar codistribution of the three proteins was immunorevealed in differentiating spermatids. Together, these data demonstrate that a protein complex formed by endogenous Rap1, 93- to 95-kDa B-Raf, and 14-3-3 theta exists in vivo and the finding that this has been detected in cytodifferentiating, not dividing cells, strengthens the hypothesis for a role of Rap1/B-Raf-mediated signaling in cell morphogenesis and differentiation. PMID- 10854066 TI - Transforming growth factor-beta1 promotes the morphological and functional differentiation of the myofibroblast. AB - The myofibroblast is responsible for the generation of contractile force associated with wound contraction and pathological contractures and is characterized by the presence of alpha-smooth muscle (alpha-sm) actin-containing stress fibers, vinculin-containing fibronexus adhesion complexes, and fibronectin fibrils containing the ED-A splice variant. Transforming growth factor-beta1 (TGF beta1) can promote the expression of alpha-sm actin in myofibroblasts, but the functional significance of this increased expression is unclear. In this study, we demonstrate, using the stress-relaxed collagen lattice contraction assay, that TGF-beta1 promoted a dose-dependent increase in the generation of contractile force in myofibroblasts and a concomitant increase in the expression of alpha-sm actin. We also demonstrate that TGF-beta1 enhanced the formation of the structural elements important in myofibroblast contractile force generation and transmission, including stress fibers, vinculin-containing fibronexus adhesion complexes, and fibronectin fibrils, and that this enhancement occurred prior to, and independent of, alpha-sm actin expression. This differentiated myofibroblast phenotype was not stable. Removal of TGF-beta1 resulted in reduced expression of alpha-sm actin as well as a decreased assembly of stress fibers and vinculin containing adhesion complexes; however, there was no reduction in fibronectin fibrils. We conclude that TGF-beta1 promotes the morphological and functional differentiation of the myofibroblast by first enhancing the formation of the structural elements characteristic of the myofibroblast followed by increased expression of alpha-sm actin and contractile force generation. PMID- 10854067 TI - Activation of ERK and p38 MAP kinases in human fibroblasts during collagen matrix contraction. AB - Studies were carried out to characterize changes in MAP kinase activation during contraction of collagen matrices by fibroblasts under isometric tension. We found that both ERK and p38 MAP kinases were activated during contraction, as determined by immunoblotting and in vitro kinase assays. ERK activation was biphasic, with peaks at 10 min and 2 h; whereas p38 activation was monophasic, with a single peak at 10 min. Activation of ERK, but not p38, appeared to depend at least in part on the Gi class of heterotrimeric G proteins. The results show that ERK and p38 cooperate in contraction-stimulated activation of c-fos transcription. PMID- 10854068 TI - Dissociation of bone morphogenetic protein-mediated growth arrest and apoptosis of mouse B cells by HPV-16 E6/E7. AB - We have previously found that bone morphogenetic protein-2 (BMP-2), a member of the transforming growth factor-beta family, induces cell-cycle arrest in the G1 phase and apoptotic cell death of HS-72 mouse hybridoma cells. In this study, we show that BMP-2 did not alter expression of cyclin D, cyclin E, cyclin-dependent kinase 2 (CDK2), CDK4, p27KIP1, p16INK4a, or p15INK4b, but enhanced expression of p21(CIP1/WAF1). Accumulation of p21(CIP1/WAF1) resulted in increased binding of p21(CIP1/WAF1) to CDK4 and concomitantly caused a profound decrease in the in vitro retinoblastoma protein (Rb) kinase activity of CDK4. Furthermore, the ectopic expression of human papilloma virus type-16 E7, an inhibitor of p21(CIP1/WAF1) and Rb, reverted G1 arrest induced by BMP-2. Expression of E6/E7, without increasing the p53 level, blocked inhibition of Rb phosphorylation and G1 arrest, but did not attenuate cell death in BMP-treated HS-72 cells. Taken together, these results suggest that inhibition of Rb phosphorylation by p21(CIP1/WAF1) is responsible for BMP-2-mediated G1 arrest and that BMP-2 induction of apoptosis might be independent of Rb hypophosphorylation. PMID- 10854069 TI - Study of the cytolethal distending toxin-induced cell cycle arrest in HeLa cells: involvement of the CDC25 phosphatase. AB - HeLa cells exposed to Escherichia coli cytolethal distending toxins (CDT) arrest their cell cycle at the G2/M transition. We have shown previously that in these cells the CDK1/cyclin B complex is inactive and can be reactivated in vitro using recombinant CDC25 phosphatase. Here we have investigated in vivo the effects of CDC25 on this cell cycle checkpoint. We report that overexpression of CDC25B or CDC25C overrides an established CDT-induced G2 cell cycle arrest and leads the cells to accumulate in an abnormal mitotic stage with condensed chromatin and high CDK1 activity. This effect can be counteracted by coexpression of the WEE1 kinase. In contrast, overexpression of CDC25B or C prior to CDT treatment prevents G2 arrest and allows most of the cells to progress through mitosis with only a low percentage of cells arrested in abnormal mitosis. The implications of these results on the biochemical nature of the CDT-induced cell cycle arrest are discussed. PMID- 10854070 TI - Involvement of Ets transcription factors and targets in osteoblast differentiation and matrix mineralization. AB - The osteoblast-like MC3T3-E1 cell line provides an excellent in vitro model of bone development. This system undergoes three orderly time-dependent phases characterized by proliferating preosteoblasts, matrix accumulation by postmitotic differentiating osteoblasts, and mineralization of the matrix, which results in the formation of multilayered bone nodules. The Ets family transcription factors regulate genetic programs that affect the proliferation and differentiation of osteoblasts. Of the eight Ets family transcription factors examined by our laboratory, only Etsl and Ets2 were found to be expressed at significant levels in this osteogenic system. Etsl is expressed in proliferating preosteoblastic cells, whereas Ets2, silent during this phase, is expressed by differentiating and mature osteoblasts. In addition, the expression of Etsl can be induced in MC3T3-E1 and fetal rat calvaria cells by retinoic acid (RA) which is known to exert profound effects on skeletal growth and development and bone turnover and induce specific cellular responses in bone cells. Thus, the multiple functions of RA in bone cells are likely to be mediated in part by Etsl. We show that the expression of Ets2 precedes and then parallels osteopontin expression and that the OPN promoter contains Ets binding sites and is a transcriptional target of Ets2. In order to identify other potential Ets target genes, we analyzed promoter regions of genes revealed by serial analysis of gene expression as present in the differentiation stage. The functional analysis of these genes has the potential to provide much needed information as to their function in osteogenesis and mineralization of the extracellular matrix and in bone-related diseases. PMID- 10854072 TI - Hypertension and cardiovascular health in Venezuela and Latin American countries. AB - Since 1950 all countries of the Latin-American subcontinent have experienced very important changes in several health indicators, in the demographic, epidemiological, socio-cultural and way of living profiles. The proportion of the population over 65 years old tend to be low in the Latin American countries in contrast to developed countries. Cardiovascular diseases are the main cause of death in most of the Latin American countries at a similar rate to that of the developed world. As infectious diseases are reduced, cardiovascular diseases takes their place as the main cause of death in Latin American countries. Prevalence of hypertension in different reports show variations from 40 to 8% in the adult population, but on average 20 to 23% of the adult population have elevated blood pressure. This prevalence is similar to reports in the developed world. However there is considerable variability in each country and its regions so it is important that local studies of prevalence and local factors in the development of hypertension are investigated. The degree of awareness, treatment and control of hypertension is lower than that reported in the developed world, and it is important to establish programmes to attend to this public health problem, from prevention to treatment, from primary care to higher levels of attention. PMID- 10854071 TI - Cripto-1-induced increase in vimentin expression is associated with enhanced migration of human Caski cervical carcinoma cells. AB - Cripto-1 (CR-1), a member of the EGF-CFC peptide family, plays an essential role during mesoderm formation in vertebrates as well as in cancer development. Using cDNA gene expression array, Western blot, and indirect immunofluorescence, an increase in vimentin expression was demonstrated in CR-1-transfected human Caski cervical carcinoma cells compared to control vector-transfected cells. In parental Caski cells, recombinant CR-1 induced a dose-dependent increase of vimentin protein expression within 24 h. Since vimentin expression has been demonstrated to correlate with a more aggressive phenotype in human cervical cancer, the migration capacity of CR-1-transfected or CR-1-treated Caski cells was studied in the Boyden chamber assay. Compared to the vector-transfected or untreated Caski cells, CR-1-transfected cells or cells treated with recombinant CR-1 exhibit enhanced migration, both through collagen- and through gelatin coated membranes. Additionally, CR-1 can function as a chemoattractant for Caski cells. These findings are of biological significance since CR-1 is overexpressed in several types of human carcinomas. The present data demonstrate that CR-1 can increase vimentin expression and modulate migration in human cervical carcinoma cells. PMID- 10854073 TI - Epidemiologic aspects of arterial hypertension in Maracaibo, Venezuela. AB - The purpose of this study was to determine the prevalence of arterial hypertension (HT) awareness and the influence of age, sex and body mass index on the degree of control of HT in the population of Maracaibo, State of Zulia, Venezuela. It included 7424 subjects, 3640 males (M) and 3784 females (F). Information was collected through domiciliary visits with a questionnaire designed for this purpose. Hypertension was defined as such when values were > or =140 mm Hg for systolic blood pressure (SBP) and > or =90 mm Hg for diastolic blood pressure (DBP). In the total sample, 36.9% were hypertensive. A higher prevalence in M (45.2%) than in F (28.9%), was observed. The percentage of HT increased with age in both genders. There was a high percentage of hypertensives with obesity (73.5%) which did not vary when discriminating for gender and age. Obese subjects were more prone to have HT until age 50. Those younger than 40 took less medication but were proportionally better controlled. Of the hypertensive population 54.3% were not aware of their condition, of 45.7% remaining, 22.8% did not have regular control visits, 18.4% inspite of medication were not controlled and only 4.5% were well controlled. Better control was observed in F (6.2%) than in M (3.3%), P < 0.001. It is concluded that HT is a serious public health problem because of its high prevalence and lack of control, and it is necessary to implement educational and medical programmes for the detection and control of this disease. PMID- 10854074 TI - Blood pressure studies in paediatric populations: metabolic syndrome in hypertensive children and adolescents. AB - The purpose of this paper is to outline our studies and observations regarding the prevalence of high blood pressure values in paediatric and juvenile populations, the early response of the left ventricle (left ventricular mass and septal and parietal wall thickness) to elevated blood pressure, the presence of a metabolic syndrome in children and adolescents with high blood pressure values, and the influence of this latter finding in the management and treatment of paediatric and juvenile hypertension. PMID- 10854075 TI - Role of prostaglandins in hypertension. AB - The role of prostaglandins (PGs) in hypertension (HT) is reviewed, emphasising their biochemical characteristics, physiological effects and functions, especially in the cardiovascular area, and the current evidence of their participation in the antihypertensive activity of a balanced mechanism to maintain normal blood pressure. Also, the clinical use of PGs and the future of such autacoids in the treatment of HT and other diseases or conditions is mentioned. PMID- 10854076 TI - Endothelial dysfunction in arterial hypertension. AB - Systemic arterial pressure is a dynamic and reactive physiological parameter depending on a great many factors. The endothelial cells of the vascular system are responsible for many biochemical reactions maintaining vascular homeostasis and therefore arterial pressure. Arterial hypertension, atherosclerosis and endothelial dysfunction constitute risk factors increasing morbidity and mortality of cardiovascular origin. These three elements are closely related and frequently act simultaneously damaging different organs. In this paper we review the physiology of the endothelium and the probable consequences of endothelial dysfunction on the pathophysiology of arterial pressure. PMID- 10854077 TI - Diabetes and hypertension physiopathology and therapeutics. AB - Diabetes mellitus by itself, is a frequent and increasing public health problem. The prevalence in most Western countries varies between 2 to 5% and it is rapidly increasing in Asiatic countries due to changes in dietary habits during the last years. The association between diabetes mellitus and hypertension has been described in 60 to 65% of diabetics. In hypertension we find insulin resistance mainly in skeletal muscle involving the conversion of glucose to glycogen independently of blood flow. The degree of resistance is related to the severity of hypertension and varies between races. States of hyperinsulinaemia and insulin resistance have been postulated as causes and/or consequences of hypertension. Regardless of the type of diabetes, hypertension is two to three times more common among diabetics compared with non-diabetics. In this paper we propose to review the essential physiopathological mechanisms involved in this association that causes high morbidity and mortality rates and increases disability among the population involved. PMID- 10854078 TI - Insulin and blood pressure responses to changes in salt intake. AB - In this study we evaluated the role of insulin in hypertension and on salt sensitivity. The study was conducted in 47 consecutive patients attending the Center for the Detection and Treatment of Cardiovascular and Metabolic Risk factors. The relationships between fasting and post-glucose load insulin levels and the blood pressure (BP) responses to changes in salt intake, were investigated. No correlation was observed between fasting or 2-h post-load insulin levels and mean BP (MBP), systolic BP (SBP) or diastolic BP (DBP). The plasma concentrations of insulin were not significantly related to body mass index (BMI) (r2 = 0.05; P = 0.135). Neither fasting nor 2-h post-load insulin predicted the BP response to changes in salt intake. A reduction in salt intake from 316 +/- 13 to 26 PM 3 mmoles/day, produced similar BP lowering in subjects with fasting insulin >15 microU/ml and in subjects with normal fasting insulin levels (<15 microU/ml). In addition, no relationship was observed between the magnitude of the BP responses to salt and the levels of insulin, either fasting (r2 = 0.007; P = 0.86) or 2-h after a glucose load (r2 = 0.01; P = 0.213). A very strong association was found between body weight or BMI and MBP (r2 = 0.443; P< 0.0001). In conclusion, our results are against the view of a cause-effect relationship between insulin and BP levels. In addition, the insulin status of a patient does not predict (nor determines) his (her) vascular reactivity to changes in salt intake. Finally, our findings further support the existence of a strong and direct association between body weight and hypertension, and speak against a major role of insulin in the pathogenesis of hypertension associated with obesity. PMID- 10854079 TI - Renin-angiotensin-aldosterone system in pregnancy-induced hypertension. AB - Angiotensin-converting enzyme (ACE) and aldosterone concentration were measured in the plasma of 32 pregnant women with normal blood pressure or with pregnancy induced hypertension (PIH). Mean arterial pressure (MAP) in PIH women was significantly higher than in normotensive pregnant women. Plasma ACE activity and aldosterone concentration in 17 PIH patients was significantly lower than their normotensive counterparts (15 cases). Mean arterial pressure was negatively correlated with the level of plasma ACE activity and with plasma aldosterone concentration in all pregnant women. Our results indicate that the circulating renin-angiotensin-aldosterone system is suppressed in pregnancy-induced hypertension. PMID- 10854080 TI - Role of bradykinins and nitric oxide in the AT2 receptor-mediated hypotension. AB - Footshocks increases mean arterial pressure and heart rate. Systemic or intracerebroventricular (IVT) administration of losartan, a specific angiotensin AT1 receptor antagonist, not only inhibited the pressor response to footshocks but resulted in vasodepression. Peripheral or IVT administration of PD 123319, a specific angiotensin AT2 receptor antagonist, did not alter the haemodynamic response to footshocks. However, simultaneous blockade of angiotensin AT1 and AT2 receptors by combined systemic or central administration of losartan and PD 12319, eliminated the vasodepressor response to footshocks unmasked in losartan pretreated rats. Our data suggest that activation of peripheral or brain angiotensin AT2 receptor mediated the vasodepressor response to footshocks in the presence of angiotensin AT1 receptor antagonist. We studied the role of kinins and nitric oxide in the vasodepressor response observed after footshocks. The decrease in mean arterial pressure observed after footshocks in losartan treated rats was blunted by systemic or IVT administration of icatibant (HOE 140) or N(G) nitro-L-arginine-methyl ester, indicating that peripheral or brain kinins and nitric oxide are involved in the hypotensor response to footshocks during angiotensin AT1 receptor blockade. Our results suggest a role for angiotensin AT2 receptor in the regulation of arterial blood pressure, possibly through the release of vasodilator autacoids such as bradykinins and nitric oxide. PMID- 10854081 TI - Dopamine: a role in the pathogenesis and treatment of hypertension. AB - The catecholamine dopamine (DA), activates two distinct classes of DA-specific receptors in the cardiovascular system and kidney--each capable of influencing systemic blood pressure. D1 receptors on vascular smooth muscle cells mediate vasodilation, while on renal tubular cells they modulate sodium excretion. D2 receptors on pre-synaptic nerve terminals influence noradrenaline release and, consequently, heart rate and vascular resistance. Activation of both, by low dose DA lowers blood pressure. While DA also binds to alpha- and beta-adrenoceptors, selective agonists at both DA receptor classes have been studied in the treatment of hypertension. An unfavourable side-effect profile (largely nausea and orthostasis) have precluded wide use of D2 agonists. In contrast, the D1 selective agonist fenoldopam has been licensed for the parenteral treatment of severe hypertension. Apart from inducing systemic vasodilation it induces a diuresis and natriuresis, enhanced renal blood flow, and a small increment in glomerular filtration rate. Evidence is emerging that abnormalities in DA production, or in signal transduction of the D1 receptor in renal proximal tubules, may result in salt retention and high blood pressure in some humans and in several animal models of hypertension. PMID- 10854082 TI - The modern treatment of hypertension. AB - Progress in the treatment of hypertension over the past 50 years is reviewed. While achievements have been considerable, they have sometimes been exaggerated by uncritical analyses and meta-analyses of trials. Data of the large but seriously flawed American HDFP study especially have distorted appreciation of therapeutic benefits. PMID- 10854083 TI - New approaches to the uses of beta blocking drugs in hypertension. AB - After a slow start, beta-blockers have become widely used as first-line agents in the treatment of hypertension, and recommended as such in recently published guidelines. There is evidence that the beta1-selective agents are more efficacious than non-selective blockers that inhibit both beta1 and beta2 receptors. Notwithstanding some earlier evidence to the contrary, it appears that beta1-selective drugs are equi-effective in young and elderly whites, younger, ie, under mid 60s, blacks. It is with the combination of age and being black that beta-blockers are usually less useful than some other groups of antihypertensive drugs, most notably calcium antagonists and diuretics. Primary prevention studies indicate beta-blockers reduce the incidence of cerebro-vascular disease and coronary heart disease in younger patients but they appear less effective than diuretics in the elderly. Beta-blockers are particularly indicated in patients who have experienced a myocardial infarction where they are often under used. There is some evidence that even in post-infarction patients with co-existent chronic obstructive airways disease, usually regarded as a contra-indication, experience an improved 2-year survival with the use of beta-blockers. Recently they have also been demonstrated to improve prognosis in heart failure patients, previously regarded as a contra-indication. Likewise, recent studies have shown that atenolol was at least as effective as captopril in improving the outlook in hypertensive patients with non-insulin dependent diabetes. While earlier comparisons with the non-selective lipid soluble propranolol indicated otherwise, comparisons with beta1-selective agents have indicated a similar effect on quality of life assessments with angiotensin-converting enzyme inhibitors. PMID- 10854084 TI - Angiotensin II receptor antagonists in arterial hypertension. AB - Angiotensin II receptor antagonists (AT-1) represent a new group of orally active antihypertensive agents. Activation on AT-1 receptor leads to vasoconstriction, stimulation of the release of catecholamines and antidiuretic hormone with production of thirst, and promote growth of vascular and cardiac muscle; these effects are blocked by AT-1 antagonist agents. The first chemically useful, orally active AT-1 receptor antagonist was losartan, followed by other agents currently in clinical use, such as: valsartan, eprosartan, irbesartan, telmisartan, candesartan, and many others under investigation. AT-1 receptor antagonists are effective in reducing high blood pressure in hypertensive patients. Monotherapy in mild to moderate hypertension controls blood pressure in 40 to 50% of these patients; when a low dose of a thiazide diuretic is added, 60 to 70% of patients are controlled. The efficacy is similar to angiotensin converting enzyme inhibitors, diuretics, calcium antagonists and beta-blocking agents. Tolerability has been reported to be very good. AT-1 receptor antagonists would be a drug of choice in otherwise well-controlled hypertensive patients treated with angiotensin-converting enzyme inhibitors who developed cough or angioedema. The final position in the antihypertensive therapy in this special population and other clinical situations, such as left ventricular hypertrophy, heart failure, diabetes mellitus and renal disease, has to be determined in large prospective clinical trials, some of which are now being conducted. PMID- 10854085 TI - Clinical pharmacokinetics of angiotensin II (AT1) receptor blockers in hypertension. AB - Angiotensin II receptor blockers (ARBs) represent a new class of effective and well tolerated orally active antihypertensive agents. Recent clinical trials have shown the added benefits of ARBs in hypertensive patients (reduction in left ventricular hypertrophy, improvement in diastolic function, decrease in ventricular arrhythmias, reduction in microalbuminuria, and improvement in renal function), and cardioprotective effect in patients with heart failure. Several large long-term studies are in progress to assess the beneficial effects of ARBs on cardiac hypertrophy, renal function, and cardiovascular and cerebrovascular morbidity and mortality in hypertensive patients with or without diabetes mellitus, and the value of these drugs in patients with heart disease and diabetic nephropathy. The ARBs specifically block the interaction of angiotensin II at the AT1 receptor, thereby relaxing smooth muscle, increasing salt and water excretion, reducing plasma volume, and decreasing cellular hypertrophy. These agents exert their blood pressure-lowering effect mainly by reducing peripheral vascular resistance usually without a rise in heart rate. Most of the commercially available ARBs control blood pressure for 24 h after once daily dosing. Sustained efficacy of blood pressure control, without any evidence of tachyphylaxis, has been demonstrated after long-term administration (3 years) of some of the ARBs. The efficacy of ARBs is similar to that of thiazide diuretics, beta-blockers, angiotensin-converting enzyme inhibitors or calcium channel blockers in patients with similar degree of hypertension. Higher daily doses, dietary salt restriction, and concomitant diuretic or ACE inhibitor administration amplify the antihypertensive effect of ARBs. The ARBs have a low incidence of adverse effects (headache, upper respiratory infection, back pain, muscle cramps, fatigue and dizziness), even in the elderly patients. After the approval of losartan, five other ARBs (candesartan cilexetil, eprosartan, irbesartan, telmisartan, and valsartan) and three combinations with hydrochlorothiazide (irbesartan, losartan and valsartan) have been approved as antihypertensive agents, and some 28 compounds are in various stages of development. The ARBs are non-peptide compounds with varied structures; some (candesartan, losartan, irbesartan, and valsartan) have a common tetrazolo biphenyl structure. Except for irbesartan, all active ARBs have a carboxylic acid group. Candesartan cilexetil is a prodrug, while losartan has a metabolite (EXP3174) which is more active than the parent drug. No other metabolites of ARBs contribute significantly to the antihypertensive effect. The variation in the molecular structure of the ARBs results in differences in the binding affinity to the receptor and pharmacokinetic profiles. The differences observed in lipid solubility, absorption/distribution, plasma protein binding, bioavailability, biotransformation, plasma half-life, and systemic elimination influence the time of onset, duration of action, and efficacy of the ARBs. On the basis of the daily mg dose, the antihypertensive potency of the ARBs follows the sequence: candesartan cilexetil > telmisartan approximately = losartan > irbesartan approximately = valsartan > eprosartan. After oral administration, the ARBs are rapidly absorbed (time for peak plasma levels = 0.5-4 h) but they have a wide range of bioavailability (from a low of 13% for eprosartan to a high of 60-80% for irbesartan); food does not influence the bioavailability, except for valsartan (a reduction of 40-50%) and eprosartan (increase). A limited dose-peak plasma levels/areas under the plasma level-time curve proportionality is observed for some of the ARBs. Most of these drugs have high plasma protein binding (95 100%); irbesartan has the lowest binding among the group (90%). The steady-state volumes of distribution vary from a low of 9 L (candesartan) to a high of 500 L (telmisartan). (ABSTRACT TRUNCATE PMID- 10854086 TI - Angiotensin II-receptor blockers: will they replace angiotensin-converting enzyme inhibitors in the treatment of hypertension? AB - In the past few years, angiotensin II-receptor blockers have become available and are being heavily marketed and increasingly used. In various ways they differ from angiotensin-converting enzyme inhibitors (ACEIs). Until outcome data are available, they should continue to be used primarily in patients who should receive an ACEI but cannot tolerate the drug because of cough. PMID- 10854087 TI - Effect of lacidipine and nifedipine GITS on platelet function in patients with essential hypertension. AB - With the aim of evaluating the effects on blood pressure, platelet function and insulin sensitivity of the dihydropiridines lacidipine and nifedipine GITS, a parallel double-blind study was carried out in a group of 20 patients with mild to moderate essential hypertension. They received a placebo for 4 weeks; then were divided at random into two groups of 10 patients each. Nifedipine GITS, 30 mg and lacidipine, 4 mg, were given during 16 weeks of active treatment. Blood pressure and heart rate were measured at the clinic in supine, sitting and standing positions, 24 +/- 1 h after the last dose. After the placebo and active phases were carried out, a platelet aggregation test was performed to determine platelet malondialdehyde production and a tolerance to 100 g of glucose by measuring glucaemia and plasma insulin. Both drugs reduced systolic and diastolic blood pressure at the same level, however there were observable differences in the rate of reduction. The nifedipine GITS reduced supine systolic blood pressure by 25 mm Hg in the first week, while the lacidipine did so by 11 mm Hg. At the end of the study period nifedipine reduced supine systolic blood pressure by 28 mm Hg and lacidipine by 20 mm Hg. Heart rate was increased slightly but significantly in the nifedipine GITS group only in the standing position. Both drugs reduced platelet aggregation ex vivo only marginally but they modified the malondialdehyde production, indicating an action on the arachidonic acid metabolic pathway. PMID- 10854088 TI - Calcium channel blockers: do they have pleiotropic effects on atherosclerosis? AB - Since 1980, calcium channel blockers (CCBs) have been used effectively to lower high blood pressure (HBP). Flekestein proposed that CCBs could be used to avoid calcium deposition on the arterial wall, one of the main components of atherosclerosis. This is also called ectopic calcium. This review briefly looks over the current role of CCBs in atherosclerosis and the future based on human trials still in progress. PMID- 10854089 TI - Nutritional factors in high blood pressure. AB - Metabolism of Na+, Ca+ and Zn+ cations is clearly disturbed and involved in the development and maintenance of a hypertensive condition. These changes are closely related to each other; therefore, when their effects on primary hypertension (PH) are studied, they should always be globally (and not separately) considered. These changes of the aforementioned electrolytes in PH maintain a close, but unclear, relationship with various hormonal systems, mainly with the renin-angiotensin-aldosterone system. Daily control in the intake of these electrolytes (especially Na+ and Ca++) remains a cornerstone in the adjuvant treatment of PH. Na+ dietary restriction is indicated in hypertensives showing higher sensitivity to salt; in most cases they have low (70%) plasma renin activity (PRA) and belong to one of the following five groups: black, elderly, obese or diabetic (type 2) patients, and mixed blood young people from our community with low levels of PRA and serum ionic calcium. For best results, this moderate Na+ restriction (4-6 g of NaCl) should always be accompanied by an oral calcium supplement, or at least the assurance that the subject takes an appropriate amount of Ca++ (>800 mg/day) in his/her diet. Hypertensives with low PRA exhibit obvious changes of their calcium metabolism. We do not know the role of Zn++ in the development of PH; however, older hypertensives with very low PRA have high urinary excretion of Zn++ with low serum levels, a factor that could contribute to Zn++ depletion in these hypertensive patients. The oral administration of calcium corrects the Zn++ changes by a still unclear mechanism. PMID- 10854090 TI - Narrowing the Duane syndrome critical region at chromosome 8q13 down to 40 kb. AB - Duane syndrome (MIM 126800) is an autosomal dominant disorder characterised by primary strabismus and other ocular anomalies, associated with variable deficiency of binocular sight. We have recently identified a < 3 cM smallest region of deletion overlap (SRO) by comparing interstitial deletions at band 8q13 in two patients (one described by Vincent et al, 1994, and the other by Calabrese et al, 1998). Here we report on another patient with Duane syndrome carrying a reciprocal translation t(6;8)(q26;q13). FISH and PCR analyses using a YAC contig spanning the SRO narrowed the Duane region to a < 1 cM interval between markers SHGC37325 and W14901. In addition, the identification and mapping of two PAC clones flanking the translocation breakpoint, allowed us to further narrow the critical region to about 40 kb. As part of these mapping studies, we have also refined the map position of AMYB, a putative candidate gene, to 8q13, centromeric to Duane locus. AMYB is expressed in brain cortex and genital crests and has been previously mapped to 8q22. PMID- 10854091 TI - Preserved speech variant is allelic of classic Rett syndrome. AB - Rett syndrome is a neurological disorder affecting predominantly females with regression loss of speech and purposeful hand use, after a few months of almost normal development. Postnatal microcephaly, hand dispraxia, stereotypic 'hand washing' activities, ataxia, and abnormal breathing are among its most characteristic features. Another aspect of this disorder is growth failure. The preserved speech variant (PSV) shares with Rett syndrome the same course and the stereotypic hand-washing activities but it differs in that patients typically recover some degree of speech and hand use and usually do not show growth failure. Progressive scoliosis, epilepsy and other minor handicaps, usually present in Rett syndrome, are rare in the preserved speech variant. Here we explore the spectrum of mutations affecting the MECP2 gene in a group of 25 classic Rett syndrome girls and in three patients with the preserved speech variant. Among the Rett syndrome group, two novel mutational hot spots (R270X and R294X), four novel mutations, two novel small deletions, as well as the previously reported 806delG, R168X and R255X mutations, were identified in 20/25 patients. Of note, among the preserved speech variants, two patients carry deletions of 41 bp and 44 bp each, which are strikingly similar to those observed in classic Rett syndrome. Our results confirm the presence of mutational hot spots in MECP2, broaden the spectrum of mutations, pinpoint additional mutational hot spots and establish that the preserved speech variant is indeed allelic of the classic form. Phenotype variability is only partially dependent on the kind of MECP2 mutation and other mechanisms such as skewed X-inactivation, and/or modifier gene effects should be investigated to explain the variable recovery in speech and hand use. PMID- 10854092 TI - Importance of searching for associated mitochondrial DNA alterations in patients with multiple deletions. AB - Multiple mitochondrial DNA (mtDNA) deletions have been reported in patients with autosomal dominant and recessive disorders. We studied several affected and one non-affected individuals belonging to a pedigree in which the inheritance of the pathological trait was compatible with an autosomique dominant transmission. Affected members had late-onset multisystem disorders with multiple mtDNA deletions in skeletal muscle. But this family presented a striking difference from previously described cases, because none of the patients had progressive external ophthalmoplegia (PEO). We also studied one young boy with a no contributary family history. He had a cerebellar ataxia with PEO and multiple mtDNA deletions in muscle. Molecular analysis revealed that in the first family, repeated sequences were present at the breakpoint junctions, whereas such motifs were not found in the young patient's case. In the first family, we evidenced mtDNA point mutations in clones containing breakpoint junctions and a 9-bp motif triplication in the intergenic COII/tRNA(Lys) region, whereas this sequence is repeated twice in the wild type mtDNA. Our results suggest that multiple deletions observed in the two pedigrees result from different molecular mechanisms and point out the role of repeated sequences in the first pedigree. No mtDNA repair system has been described in mammals so far, but the molecular abnormalities found in the first family suggest that a defect in an mtDNA repair system, homologous to the E. coli MutHLS pathway, could be responsible for such a phenotype. PMID- 10854093 TI - MtDNA and Y chromosome polymorphisms in Hungary: inferences from the palaeolithic, neolithic and Uralic influences on the modern Hungarian gene pool. AB - Magyars imposed their language on Hungarians but seem not to have affected their genetic structure. To better investigate this point, we analysed some mtDNA and Y chromosome polymorphisms in a sample of the Hungarian Paloc who, for historical reasons, could have retained genetic traces of Magyars more than other groups. In addition, we examined a mixed sample from Budapest. About 100 individuals were tested for the markers defining all the European and Asian mtDNA haplogroups and about 50 individuals for some Y chromosome markers, namely the 12f2 and 49a,f/TaqI RFLPs, the YAP insertion, the microsatellites YCAIIa, YCAIIb, DYS19 and the Asian 50f2/C deletion. In the mtDNA analysis only two subjects belonged to the Asian B and M haplogroups. The Y chromosome analyses showed that the Paloc differed from the Budapest sample by the absence of YAP+ allele and by the DYS19 allele distribution; that the proto-European 49a,f Ht 15 and the neolithic 12f2 8Kb were rather uncommon in both groups; that there is a high prevalence of the 49a,f Ht 11 and the YCAII a5-b1; and that the Asian 50f2/C deletion is absent. These results suggest that the influence of Magyars on the Hungarian gene pool has been very low through both females and males and the Hungarian language could be an example of cultural dominance. Alternative explanations are discussed. An expansion centred on YAP-, 49a,f Ht 11 is revealed by the median network based on compound haplotypes. 49a,f Ht 11 could represent either a paleolithic marker of eastern Europe which underwent expansion after the last glacial period, or a marker of the more recent spread of the Yamnaia culture from southern Ukraine. PMID- 10854094 TI - Cell type specificity in alternative splicing of the human mismatch repair gene hMSH2. AB - Human non-polyposis colorectal cancer is caused by germline mutations in the DNA mismatch repair genes hMSH2 and hMHL1. Several alternatively spliced mRNA species of these genes are present in peripheral blood lymphocytes of normal individuals, which can confound RT-PCR based techniques of mutation detection. Using RT-PCR, we compared the pattern of alternative splicing in whole peripheral blood lymphocytes (PBLs), separated T and B cells, lymphoblastoid cell lines (LCLs) from the same individuals, and a variety of tissues. Alternatively spliced forms of hMLH1 lacking exons 9/10, 10/11 and 9/10/11 were found to have similar patterns of expression in T cells, B cells, and LCLs. By contrast, a subset of hMSH2 transcripts, some of which were produced by utilisation of novel splicing motifs, were generally expressed in T but not in B cells. LCLs derived from the same blood samples showed no expression of any hMSH2 splicing variants. The hMSH2 delta ex13 transcript, while absent from LCLs, was expressed in whole PBLs and both T and B cell fractions. This transcript was furthermore largely undetectable in tissues other than mononuclear blood cells. These data provide evidence for tissue specificity in the regulation of alternative splicing in hMSH2. In particular we show that LCLs generally do not express alternatively spliced forms of hMSH2 mRNA and are thus suited for RT-PCR based mutation screening in that gene. PMID- 10854095 TI - Novel mutations in the duplicated region of PKD1 gene. AB - Autosomal dominant polycystic kidney disease (ADPKD) exhibits a genetically heterogeneous transmission involving at least three different genes. PKD1 gene linked mutations are responsible for the disease in about 85% of ADPKD cases. The search for mutations is a very important step in understanding the molecular mechanisms underlying ADPKD. We undertook this study using denaturing gradient gel electrophoresis (DGGE), after a stage of long range PCR, to scan for mutations in the duplicated region of the PKD1 gene in French ADPKD families. This allowed us to identify eight novel mutations and several polymorphisms: among the mutations, three are nonsense mutations, two are deletions in the coding sequence leading to frameshift mutations, one is a splice mutation and two are highly probable missense mutations. In this paper, we also provide a review of the mutations reported so far which are widespread throughout the gene. Although no clear hot spot for mutation is apparent, we will focus on some clustering observed. PMID- 10854096 TI - Genetic structure of north-west Africa revealed by STR analysis. AB - We have analysed a large set of autosomal short tandem repeat (STR) loci in several Arabic and Berber-speaking groups from north-west Africa (ie Moroccan Arabs, northern-central and southern Moroccan Berbers, Saharawis, and Mozabites). Two levels of analysis have been devised using two sets of 12STR loci, (D3S1358, vWA, FGA, THO1, TPOX, CSF1PO, D8S1179, D21S11, D18S51, D5S818, D13S317 and D7S820) and 21 (the former set plus D9S926, D11S2010, D13S767, D14S306, D18S848, D2S1328, D4S243, F13A1, and FES/FPS). For each set, data for a number of external reference populations were gathered from the literature. Several methods of analysis based on genetic distances (neighbour-joining trees, principal coordinate analysis, boundary detection), as well as AMOVA, showed that genetic differentiation among NW African populations was very low and devoid of any spatial pattern. When the NW African populations were grouped according to cultural or linguistic differences, the partition was not associated with genetic differentiation. Thus, it is likely that Arabisation was mainly a cultural process. A clear genetic difference was found between NW African populations and Iberians, which underscores the Gilbraltar Straits as a strong barrier to genetic exchange; nonetheless, some degree of gene flow into Southern Iberia may have existed. NW Africans were genetically closer to Iberians and to other Europeans than to African Americans. PMID- 10854097 TI - Lack of Hardy-Weinberg equilibrium for the most prevalent PMM2 mutation in CDG-Ia (congenital disorders of glycosylation type Ia). AB - The R141H mutation in the PMM2 gene is the most frequent mutation in type Ia of the congenital disorders of glycosylation (formerly carbohydrate-deficient glycoprotein syndromes)(CDG-Ia). However, it has never been observed in the homozygous state. Homozygosity for this mutation is probably incompatible with life. In this study, we determined the frequency of R141H in two normal populations: in neonates of Dutch origin 1/79 were carriers, whilst in the Danish population, a carrier frequency of 1/60 was found. These figures are clearly in disequilibrium with the frequency of CDG-Ia that has been estimated at 1/80,000 to 1/40,000 in these populations. Haplotype analysis of 43 patients with the R141H mutation of different geographic origins indicated that the R141H is an old mutation in the Caucasian population. Based on the new data, the disease frequency has been calculated at 1/20,000 in these populations. It is concluded that the disease is probably underdiagnosed. PMID- 10854098 TI - Clouston hidrotic ectodermal dysplasia (HED): genetic homogeneity, presence of a founder effect in the French Canadian population and fine genetic mapping. AB - HED is an autosomal dominant skin disorder that is particularly common in the French Canadian population of south-west Quebec. We previously mapped the HED gene to the pericentromeric region of chromosome 13q using linkage analysis in eight French Canadian families. In this study, we extend our genetic analysis to include a multiethnic group of 29 families with 10 polymorphic markers spanning 5.1 cM in the candidate region. Two-point linkage analysis strongly suggests absence of genetic heterogeneity in HED in four families of French, Spanish, African and Malaysian origins. Multipoint linkage analysis in all 29 families generated a peak lod score of 53.5 at D13S1835 with a 1 lod unit support interval spanning 1.8 cM. Recombination mapping placed the HED gene in a 2.4 cM region flanked by D13S1828 proximally and D13S1830 distally. We next show evidence for a strong founder effect in families of French Canadian origin thereby representing the first example of a founder disease in the south-west part of the province of Quebec. Significant association was found between HED in these families and all markers analysed (Fisher's exact test, P < 0.001). Complete allelic association was detected at D13S1828, D13S1827, D13S1835, D13S141 and D13S175 (P(excess) = 1) spanning 1.3 cM. A major haplotype including all 10 associated alleles was present on 65% of affected chromosomes. This haplotype most likely represents the founder haplotype that introduced the HED mutation into the French Canadian population. Luria-Delbruck equations and multipoint likelihood linkage disequilibrium analysis positioned the gene at the D13S1828 locus (likely range estimate: 1.75 cM) and 0.58 cM telomeric to this marker (support interval: 3.27 cM) respectively. PMID- 10854099 TI - Positional cloning and characterisation of the human DLGAP2 gene and its exclusion in progressive epilepsy with mental retardation. AB - In search of the gene for progressive epilepsy with mental retardation (EPMR) we identified DLGAP2, the human homolog of the gene encoding the rat PSD-95/SAP90 associated protein-2 (Dlgap2). We extended the transcript in both the 5' and 3' directions and characterised the genomic structure of the approximately 10 kb gene. Sequence comparisons of human DLGAP2 cDNA sequences obtained from human testis and brain cDNA libraries with homologous rat genes suggest alternative splicing in the 5' end of the gene. The 5' coding sequence of the testis cDNA is complete, whereas based on homology with the rat gene 103 bp of coding sequence may still be missing in the 5' end of the DLGAP2 brain transcript. DLGAP2 was excluded as the gene responsible for EPMR. PMID- 10854100 TI - No evidence for the involvement of CAG/CTG repeats from within 18q21.33-q23 in bipolar disorder. AB - We previously identified 18q21.33-q23 as a candidate region in one BP family and constructed a yeast artificial chromosome (YAC) contig map. Here, we mapped eight known CAG/CTG repeats relative to 18q21.33-q23. We also isolated four CAG/CTG repeats from within the region using CAG/CTG YAC fragmentation, one of which is located in the 5' untranslated region of the CAP2 gene coding for a brain expressed serine proteinase inhibitor. The triplet repeats located in the 18q21.33-q23 BP candidate region showed no expanded alleles in the linked BP family nor in a BP case-control sample. Moreover, only the CAP2 triplet repeat was polymorphic but no genetic association with BP disorder was observed. PMID- 10854101 TI - Haemochromatosis gene mutations and risk of coronary artery disease. AB - The identification of mutations in the haemochromatosis gene (HFE) (C282Y and H63D) provides the unique opportunity to test whether genetic variants that are associated with tissue iron accumulation may influence the risk of coronary atherosclerosis. To this aim the prevalence of C282Y and H63D mutations was determined in 174 patients with angiographically documented CAD (>50% stenosis) and history of MI, 187 healthy free-living individuals and 142 blood donors. C282Y and H63D mutations were not found to be more frequent in coronary patients as compared to controls. Moreover, these HFE variants were unrelated to the severity of coronary atherosclerosis. These findings did not provide evidence of an association between HFE mutations and the presence of coronary atherosclerosis or its major ischaemic complications, thus indicating that HFE mutations are poor genetic markers of coronary risk. PMID- 10854102 TI - Microdissection of chromosome 2--between-arm intrachromosomal insertion. AB - This report describes a mother with a balanced intrachromosomal insertion of band q22 on chromosome number 2 into band p24 on the same chromosome. She had had four spontaneous abortions and two induced abortions. One foetus had a suspected obstruction of the uretero-pelvic part of the urinary tract and monosomy of band 2q22, the other foetus had anencephaly and trisomy of band 2q22. By microdissection we have generated a painting probe from the mother's abnormal short arm of chromosome 2 (der2p probe). This family specific probe will be used in future pregnancies for precise diagnosis. PMID- 10854103 TI - Molecular analysis of the mature T cell proliferation-1 (MTCP-1) gene in Xq28 linked incontinentia pigmenti. PMID- 10854104 TI - Phenotype in patients with Angelman syndrome. PMID- 10854105 TI - Familial Mediterranean fever in the 'Chuetas' of Mallorca: a question of Jewish origin or genetic heterogeneity. AB - Familial Mediterranean fever (FMF) is a hereditary disease commonly found among Jews, Armenians, Turks and Arabs. Recently, FMF was found in the 'Chuetas', a unique community on the island of Mallorca (Spain). To address the question of their possible Jewish origin, we analysed markers known to be linked to the gene responsible for FMF in Jews (MEFV) in this population. We found that 1/3 of the 16p13.3 chromosomes of the 'Chuetas' FMF patients bore the major ancestral haplotypes (S,S2) and their corresponding M694V and E148Q mutations, displayed by Jews from North Africa. Furthermore, we also detected a novel mutation (L110P) in this community. Yet 2/3 of these patients bore S negative haplotypes and lack the mutations commonly known to cause FMF. These results confirm that at least some of the 'Chuetas' share a common origin with Jews. However, they also provide evidence for the possibility of genetic heterogeneity in this disorder. PMID- 10854106 TI - Reproductive and menstrual history of females with fragile X expansions. AB - FRAXA premutations have been associated with premature ovarian failure (POF) or menopause before the age of 40. We have studied women in families ascertained because of a mentally retarded full mutation relative and determined their age of menopause, serum hormone levels in premenopausal individuals and the outcome of any pregnancies. Survival analysis was used as a measure of menopause and demonstrated a significant decrease in age of menopause in premutation carriers compared with their full mutation carrier and normal relatives. Serum FSH was also raised in premutation carriers, although oestradiol, inhibin A and inhibin B were not significantly different. However, we did not find an excess of dizygous twins or pregnancy loss/trisomies, both of which are associated with aging ovaries. Thus premutation carriers as a group have an earlier menopause and raised serum FSH but do not appear to manifest other features of an aging ovary. PMID- 10854107 TI - Systematic analysis of X-inactivation in 19XLMR families: extremely skewed profiles in carriers in three families. AB - It has been demonstrated in several X-linked disorders, both with and without mental retardation, that the X-inactivation process plays a significant role in the expression of X-linked diseases in females. Moreover, in some disorders extremely skewed inactivation of the X chromosome is constant in carriers, and this is thought to result from a proliferation or a survival advantage for cells expressing the normal allele at this locus over cells expressing the mutated allele. X-linked mental retardation (XLMR) is heterogeneous, and cloning and characterization of the mutated genes are in progress. XLMR can be expressed in carrier females but often with milder manifestations. We report the systematic study of the X-inactivation profile of obligate carriers and other females in 19 multiplex XLMR pedigrees, using leucocyte-extracted DNA. Extremely skewed profiles were observed in carriers in three of 19 families. PMID- 10854108 TI - Identification of a novel 4.6-kb genomic deletion in presenilin-1 gene which results in exclusion of exon 9 in a Finnish early onset Alzheimer's disease family: an Alu core sequence-stimulated recombination? AB - Mutations in the presenilin-1 (PS-1) gene have been shown to cause early onset Alzheimer's disease (EOAD) in an autosomal dominant manner. We have identified a novel 4.6-kb genomic deletion in the PS-1 gene in a Finnish EOAD family, which leads to an inframe exclusion of exon9 (delta9) from the mRNA transcript. This germline mutation results in a similar alteration in mRNA level as previously described with the variant AD and the delta9 splice-site mutations. In this present EOAD family, the clinical and neuropathological phenotype of patients are those of the typical AD without indications of spastic paraparesis or 'cotton wool' plaques, which are the hallmarks of the variant AD. A sequence analysis of the deletion crossover site of the mutant and corresponding wild type regions revealed complete homology with the recombigenic 26 bp Alu core sequence at intron 8. In addition, a segment at the intron 9 breakpoint displayed homology with the core sequence, but comparison of the 5' and 3' breakpoint sequences did not reveal significant identity favouring involvement of Alu core sequence stimulated non-homologous recombination rather than Alu-mediated homologous pairing of the fragments. This study shows that large genomic rearrangements can affect the EOAD gene PS-1 through a mechanism, which may involve Alu core sequence-stimulated recombination. PMID- 10854109 TI - A gene for ataxic cerebral palsy maps to chromosome 9p12-q12. AB - Cerebral palsy (CP) has an incidence of approximately 1 in 750 births, although this varies between ethnic groups. Genetic forms of the disease account for about 2% of cases in most countries, but contribute a larger proportion in certain sub types of the condition and in populations with a large proportion of consanguineous marriages. Ataxic cerebral palsy accounts for 5-10% of all forms of CP and it is estimated that approximately 50% of ataxic cerebral palsy is inherited as an autosomal recessive trait. We have identified a complex consanguineous Asian pedigree with four children in two sibships affected with ataxic cerebral palsy and have used homozygosity mapping to map the disorder in this family. A genome-wide search was performed using 343 fluorescently labelled polymorphic markers and linkage to chromosome 9p12-q12 was demonstrated. A maximum Lod score of 3.4 was observed between the markers D9S50 and D9S167 using multipoint analysis, a region of approximately 23cM. We have identified a family that segregates both ataxic CP and ataxic diplegia and have mapped the genetic locus responsible in this family to chromosome 9p12-q12. The identification of gene(s) involved in the aetiology of CP will offer the possibility of prenatal/premarital testing to some families with children affected with the disorder and will greatly increase our understanding of the development of the control of motor function. PMID- 10854110 TI - Linkage of Marie-Unna hypotrichosis locus to chromosome 8p21 and exclusion of 10 genes including the hairless gene by mutation analysis. AB - Marie-Unna hypotrichosis (MU) is a rare autosomal dominant congenital alopecia characterised by progressive hair loss starting in early childhood, often aggravated at puberty and leading to scarring alopecia of variable severity. We have studied three multigeneration families of Belgian, British and French descent. The human genome was screened with microsatellite markers spaced at 10 cM intervals and significant evidence for linkage to the disease was observed on chromosome 8p21, with a maximum two-point lod score of 8.26 for D8S1786 at a recombination fraction of 0. Recombinants narrowed the region of interest to a genetic interval of about 12 cM flanked by markers D8S280 and D8S1839. This interval contains the hairless gene which is mutated in autosomal recessive congenital atrichia. Sequencing of the entire coding region and intronic splice sites of the hairless gene in these three families and in two unrelated familial cases revealed several polymorphic changes but failed to identify causative mutations. Nine other genes located within this region and expressed in skin were also excluded by mutation analysis. Together with a recent linkage study performed in a Dutch and a British family by van Steensel et al these results provide evidence for the presence of a gene distinct from hairless in chromosomal region 8p21 playing an important role in hair follicle biology. PMID- 10854111 TI - Small in-frame deletions and missense mutations in CADASIL: 3D models predict misfolding of Notch3 EGF-like repeat domains. AB - CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy) is a hereditary microangiopathic condition causing stroke in young adults. The responsible gene has recently been identified as the Notch3 gene. Notch3 encodes a large transmembrane receptor with 34 extracellularly localised epidermal growth factor-like (EGF) repeat domains. We screened 71 unrelated CADASIL families for mutations in two exons coding for the first five EGF-like repeats and found mutations in 70% of the families (n = 50). Two types of mutations were identified: 48 families (96%) had missense mutations and two families (4%) had small in-frame deletions. Seven mutations occurred multiple times. All of them are C to T transitions that affect CpG dinucleotides, suggesting that their multiple occurrence is due to the hypermutability of this sequence. All mutations, including the two deletions, result in the gain or loss of a cysteine residue, thus substantiating the pivotal role of an uneven number of cysteine residues within EGF-like repeat domains of Notch3 in the pathogenesis of CADASIL. To study the potential effects of these mutations 3D homology models of the first six EGF domains were generated on the basis of NMR data from human fibrillin-1. These models predict domain misfolding for a subset of mutations. PMID- 10854112 TI - Mutations in the VMD2 gene are associated with juvenile-onset vitelliform macular dystrophy (Best disease) and adult vitelliform macular dystrophy but not age related macular degeneration. AB - Recently, the VMD2 gene has been identified as the causative gene in juvenile onset vitelliform macular dystrophy (Best disease), a central retinopathy primarily characterised by an impaired function of the retinal pigment epithelium. In this study we have further characterised the spectrum of VMD2 mutations in a series of 41 unrelated Best disease patients. Furthermore we expanded our analysis to include 32 unrelated patients with adult vitelliform macular dystrophy (AVMD) and 200 patients with age-related macular degeneration (AMD). Both AVMD and AMD share some phenotypic features with Best disease such as abnormal subretinal accumulation of lipofuscin material, progressive geographic atrophy and choroidal neovascularisation, and may be the consequence of a common pathogenic mechanism. In total, we have identified 23 distinct disease-associated mutations in Best disease and four different mutations in AVMD. Two of the mutations found in the AVMD patients were also seen in Best disease suggesting a considerable overlap in the aetiology of these two disorders. There were no mutations found in the AMD group. In addition, four frequent intragenic polymorphisms did not reveal allelic association of the VMD2 locus with AMD. These data exclude a direct role of VMD2 in the predisposition to AMD. PMID- 10854113 TI - Identification of female carriers for Duchenne and Becker muscular dystrophies using a FISH-based approach. AB - Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are X linked recessive neuromuscular diseases caused by dystrophin gene mutations. Deletions, or more rarely duplications, of single or multiple exons within the dystrophin gene can be detected by current molecular methods in approximately 65% of DMD patients. Mothers of affected males have a two-thirds chance of carrying a dystrophin mutation, whilst approximately one-third of affected males have de novo mutations. Currently, Southern blot analysis and multiplex PCR directed against exons in deletion hot spots are used to determine female carrier status. However, both of these assays depend on dosage assessment to accurately identify carriers since, in females, the normal X chromosome is also present. To obviate quantitation of gene dosage, we have developed exon-specific probes from the dystrophin gene and applied them to a screen for potential carrier females using fluorescence in situ hybridization (FISH). Cosmid clones, representing 16 exons, were identified and used in FISH analysis of DMD/BMD families. Our preliminary work has identified multiple, informative probes for several families with dystrophin deletions and has shown that a FISH-based assay can be an effective and direct method for establishing the DMD/BMD carrier status of females. PMID- 10854114 TI - RNA-based mutation screening in German families with Sjogren-Larsson syndrome. AB - Sjogren-Larsson syndrome (SLS) is a rare autosomal recessively inherited disorder characterised by mental retardation, spasticity and ichthyosis. SLS patients have a profound deficiency in fatty aldehyde dehydrogenase (FALDH) activity. The human cDNA of FALDH has been shown to map to the SLS locus on chromosome 17p11.2. Here we describe a method based on reverse transcriptase-polymerase chain reaction (RT PCR) and protein truncation test to identify mutations in the FALDH gene in nine German SLS families. Using this detection system both disease-causing mutations were found in eight of the nine SLS families examined (17/18 chromosomes). Seven different mutations were identified: an exon 2 skipping due to exon 2 splice donor mutation; two different exon 3 splice donor mutations resulting in combined exon 2 and 3 skipping; a 906delT deletion in exon 6; a genomic deletion of about 6 kb including exon 9; a 1277T > G transversion resulting in a Leu426Ter nonsense mutation; and a 1297delGA deletion. Two of the mutations identified, the genomic exon 9 deletion and the 906delT in exon 6 affected five out of seven SLS patients from a small region of Northern Bavaria. Therefore these two mutations accounted for 71% (10/14 chromosomes) of Bavarian SLS alleles and so far have not been described in SLS families from other countries. Our findings do not support our 'historical' hypothesis, that a possible region clustering in Northern Bavaria could be due to the presence of Swedish soldiers during the 30 Years War (1618 1648), but suggest that two mutations causing SLS syndrome originated in Northern Bavaria. PMID- 10854115 TI - Higher than expected carrier rates for familial Mediterranean fever in various Jewish ethnic groups. AB - Familial Mediterranean fever (FMF) is an autosomal recessive disease characterised by recurrent attacks of inflammation of serosal membranes. Amyloidosis leading to renal failure is the most severe complication in untreated patients. In Israel FMF is most frequent among Jews of North African origin. Recently the causative gene (MEFV) has been found and the common mutations characterised. The aim of this study was to investigate the carrier rates of the common MEFV mutations among 400 healthy members of four different ethnic groups (100 in each group) in Israel, and to compare the distribution of the different mutations between FMF carriers and patients. We found a high frequency of carriers among Jews from the various ethnic groups. In North African Jews it was 22%, in Iraqi Jews 39%, in Ashkenazi Jews 21%, and in Iranian Jews 6%. The distribution of the four most common MEFV mutations among healthy individuals (M694V 29%, V726A 16%, M6801 2% and E148Q 53%) was significantly different (P < 0.003) from that found in patients (M694V 84.4%, V726A 9.0%, M6801 0% and E148Q 6.6%). Six healthy asymptomatic individuals were found to carry mutations in both alleles: two homozygotes for E148Q and four compound heterozygotes E148Q/other. These results demonstrate a very high carrier rate among all Jewish ethnic groups. They confirm that mutation E148Q is associated with a milder phenotype, which explains the lower prevalence of FMF among the Ashkenazi and Iraqi Jews. This study raises the question of the need for molecular screening for M694V homozygotes in the Israeli North African Jewish community. PMID- 10854116 TI - Deletion of a branch-point consensus sequence in the LMX1B gene causes exon skipping in a family with nail patella syndrome. AB - Nail patella syndrome (NPS) has been shown to result from loss of function mutations within the transcription factor LMX1B. In a large NPS family a 17 bp intronic deletion encompassing a consensus branchpoint sequence was observed to segregate with the NPS phenotype. RNA analysis demonstrated that deletion of the branchpoint sequence resulted in skipping of the downstream exon. A mechanism to explain this phenomenon is presented. PMID- 10854117 TI - Frequency of mitochondrial DNA point mutations among patients with familial sensorineural hearing impairment. AB - Several point mutations in mitochondrial DNA (mtDNA) have been shown to cause sensorineural hearing impairment (SNHI), but the frequency of these mutations among patients is not known. We identified 117 patients with possible matrilineal SNHI from population-based registers and found the 3243A > G mutation to be present in 4.3% and 1555A > G in 2.6%, while 7445T > C, 7472insC and 8344A > G were absent. Patients with 3243A > G and 1555A > G were clinically distinct. The prevalence of 1555A > G in the general adult population was estimated to be at least 4.7/100,000, but these and previous data suggest that the figure may vary between populations. Screening for mtDNA mutations is worthwhile in connection with the diagnosis of SNHI. PMID- 10854118 TI - Computer modeling of population exposure to a carcinogen: the example of asbestos and mesothelioma mortality in France. AB - The multistage theory of carcinogenesis allows models to be constructed that provide the individual probability of a diagnosed tumor at a given age as a function of the person's past exposure to carcinogenic agents (exposure level versus age), the time since exposure, and age. When exposure to a carcinogenic agent and its impact in terms of morbidity and mortality are modeled on the scale of the general population, individual exposures must often be estimated. If appropriate data do not already exist, this difficult task necessitates expensive and difficult investigations. We propose here a method that allows this global exposure to be modeled without needing to know the individual exposures. The method is used and illustrated in the context of modeling the asbestos exposure of the French male population and calculating its mortality rate from mesothelioma (a type of cancer for which asbestos is the only carcinogen and for which a risk function based on the multistage theory of carcinogenesis exists). This method assumes that the exposure functions (how exposure levels vary with age) for all individuals are the same, with the exception of one parameter. That is, it utilizes a hypothesis that we called the hypothesis of the Standard Exposure Window (SEW). We used two methods to calculate the probability of death from mesothelioma in a representative sample of the French male population for whom individual histories of asbestos exposure are known: we applied the risk function to all the individuals, and we applied the SEW hypothesis. The number of deaths obtained by the two methods are very close and fit the observed mortality data satisfactorily. PMID- 10854119 TI - The polysomnogram assay: a method to represent the overnight polysomnogram in a condensed format. AB - We present the polysomnogram assay (PSGA), a new representation format for the polysomnogram (PSG), designed to assist in the interpretation of overnight PSG studies. The technique condenses the PSG record by a factor of 30 while preserving the ability to portray PSG features of diagnostic relevance, including sleep architecture, arousals, movement, leg jerks, cyclic alternating pattern, and increased breathing effort. The PSGA patterns associated with these events are described and illustrated by examples. The new format considerably reduces the effort required to evaluate sleep quality and continuity, making it more practicable for the polysomnographer to interpret the entire overnight PSG study. The compressed time scale also facilitates analysis of relatively long PSG episodes and allows assessment of signal activity surrounding critical PSG events. The PSGA appears capable of improving identification of arousals, leg jerks, and upper airway resistance, and may be especially amenable for automatic analysis of PSG data. PMID- 10854120 TI - IMM/Scrub: a domain-specific tool for the deduplication of vaccination history records in childhood immunization registries. AB - IMM/Scrub is a pilot tool developed to assist in the deduplication of vaccination history records in childhood immunization registries. This problem is complicated by a number of factors including that fact that: (1) some doses are numbered and some are not, (2) doses may have different dose numbers, (3) doses may specify different preparations within a vaccine series, (4) one dose may indicate a combination vaccine and the other dose may specify one component of that combination, (5) two doses may have slightly different dates, and (6) combinations of any of these problems may occur together. IMM/Scrub is designed to help detect 10 different types of vaccination dose duplicates and also allows the user to specify flexibly the conditions in which a duplicate dose might be automatically eliminated. In addition, IMM/Scrub is linked to the IMM/Serve immunization forecasting program, which can provide additional assistance in the data cleaning process. The paper describes (1) the design of the current pilot implementation of IMM/Scrub, (2) the lessons learned during its implementation, and (3) our preliminary experience applying it to data from three immunization databases, from a state, a metropolitan area, and an academic medical center. PMID- 10854121 TI - Enhancing the precision of ECG baseline correction: selective filtering and removal of residual error. AB - Reemergence of the problem of baseline correction is related to recent advancements in the electrocardiographic (ECG) analysis of beat-to-beat repolarization changes which play an important role in risk assessment and the prediction of sudden cardiac death. These alterations often have an amplitude of a few microvolts and duration of several milliseconds and their detection requires special accuracy of baseline estimation. Using detailed analysis of various types of residual errors we designed a two-step procedure for selective filtering of ECG and removal of residual error with minimal distortion of cardiac complexes and tested this approach on 100 simulated and 210 real ECG signals. Application of this procedure provided a twofold reduction in the error of baseline estimation and T-wave amplitude measurements compared to high-pass filtering. Selective application of this approach to the segments with low baseline drift allowed analysis of low-amplitude, beat-to-beat changes in repolarization during more than 70% of the recording time. PMID- 10854122 TI - Safety evaluation and risk assessment of the herbicide Roundup and its active ingredient, glyphosate, for humans. AB - Reviews on the safety of glyphosate and Roundup herbicide that have been conducted by several regulatory agencies and scientific institutions worldwide have concluded that there is no indication of any human health concern. Nevertheless, questions regarding their safety are periodically raised. This review was undertaken to produce a current and comprehensive safety evaluation and risk assessment for humans. It includes assessments of glyphosate, its major breakdown product [aminomethylphosphonic acid (AMPA)], its Roundup formulations, and the predominant surfactant [polyethoxylated tallow amine (POEA)] used in Roundup formulations worldwide. The studies evaluated in this review included those performed for regulatory purposes as well as published research reports. The oral absorption of glyphosate and AMPA is low, and both materials are eliminated essentially unmetabolized. Dermal penetration studies with Roundup showed very low absorption. Experimental evidence has shown that neither glyphosate nor AMPA bioaccumulates in any animal tissue. No significant toxicity occurred in acute, subchronic, and chronic studies. Direct ocular exposure to the concentrated Roundup formulation can result in transient irritation, while normal spray dilutions cause, at most, only minimal effects. The genotoxicity data for glyphosate and Roundup were assessed using a weight-of-evidence approach and standard evaluation criteria. There was no convincing evidence for direct DNA damage in vitro or in vivo, and it was concluded that Roundup and its components do not pose a risk for the production of heritable/somatic mutations in humans. Multiple lifetime feeding studies have failed to demonstrate any tumorigenic potential for glyphosate. Accordingly, it was concluded that glyphosate is noncarcinogenic. Glyphosate, AMPA, and POEA were not teratogenic or developmentally toxic. There were no effects on fertility or reproductive parameters in two multigeneration reproduction studies with glyphosate. Likewise there were no adverse effects in reproductive tissues from animals treated with glyphosate, AMPA, or POEA in chronic and/or subchronic studies. Results from standard studies with these materials also failed to show any effects indicative of endocrine modulation. Therefore, it is concluded that the use of Roundup herbicide does not result in adverse effects on development, reproduction, or endocrine systems in humans and other mammals. For purposes of risk assessment, no-observed-adverse-effect levels (NOAELs) were identified for all subchronic, chronic, developmental, and reproduction studies with glyphosate, AMPA, and POEA. Margins-of-exposure for chronic risk were calculated for each compound by dividing the lowest applicable NOAEL by worst-case estimates of chronic exposure. Acute risks were assessed by comparison of oral LD50 values to estimated maximum acute human exposure. It was concluded that, under present and expected conditions of use, Roundup herbicide does not pose a health risk to humans. PMID- 10854123 TI - Criteria for development of a database for safety evaluation of fragrance ingredients. AB - Over 2000 different ingredients are used in the manufacture of fragrances. The majority of these ingredients have been used for many decades. Despite this long history of use, all of these ingredients need continued monitoring to ensure that each ingredient meets acceptable safety standards. As with other large databases of existing chemicals, fulfilling this need requires an organized approach to identify the most important potential hazards. One such approach, specifically considering the dermal route of exposure as the most relevant one for fragrance ingredients, has been developed. This approach provides a rational selection of materials for review and gives guidance for determining the test data that would normally be considered necessary for the elevation of safety under intended conditions of use. As a first step, the process takes into account the following criteria: quantity of use, consumer exposure, and chemical structure. These are then used for the orderly selection of materials for review with higher quantity, higher exposure, and the presence of defined structural alerts all contributing to a higher priority for review. These structural alerts along with certain exposure and volume limits are then used to develop guidelines for determining the quality and quantity of data considered necessary to support an adequate safety evaluation of the chosen materials, taking into account existing data on the substance itself as well as on closely related analogs. This approach can be considered an alternative to testing; therefore, it is designed to be conservative but not so much so as to require excessive effort when not justified. PMID- 10854124 TI - The high production volume chemical challenge program: the relevance of the in vivo micronucleus assay. AB - The in vivo rodent bone marrow micronucleus assay (Mnt) has assumed a pivotal role in screening strategies for the identification of substances potentially carcinogenic to humans. The analysis of the results of the current international 5-year effort to provide toxicological data for high production volume chemicals will play a crucial role in developing future strategies for identifying health hazards. As part of that program, consideration is being given to accepting either in vitro genotoxicity data or results of the Mnt. The present analyses indicate that for hazard identification purposes that, in fact, in vitro genotoxicity test results, such as those derived from the Salmonella mutagenicity assay, may be an acceptable alternative. PMID- 10854125 TI - Evaluation of biologically based dose-response modeling for developmental toxicity: a workshop report. AB - Biologically based dose-response (BBDR) modeling represents a novel approach for quantitative assessment of health risk by incorporating pharmacokinetic and pharmacodynamic characteristics of a chemical and by relating the immediate cellular responses to a cascade of aberrant biological actions that leads to detectable adverse outcomes. The quantitative relationship of each of the intervening events can be described in mathematical forms that are amenable for adjustment and extrapolation over a range of doses and across species. A team of investigators at the Reproductive Toxicology Division of the U.S. Environmental Protection Agency has explored the feasibility of BBDR modeling by examining the developmental toxicity of a known teratogen, 5-fluorouracil. A panel of researchers from academic and industrial laboratories, biomathematical modelers, and risk assessment scientists was convened in a workshop to evaluate the approaches undertaken by the EPA team and to discuss the future prospects of BBDR modeling. This report summarizes the lessons learned from one approach to BBDR modeling and comments from the panelists: while it is possible to incorporate mechanistic information into quantitative dose-response models for the assessment of health risks, the process is enormously data-intensive and costly; in addition, the confidence of the model is directly proportional to our current understanding of basic biology and can be enhanced only through the ongoing novel discoveries. More importantly, the extent of "uncertainty" (inherent with the default assumptions associated with the NOAEL or benchmark approach) reducible by BBDR modeling requires further scrutiny and comparison. PMID- 10854126 TI - Effects of subchronic exposure of rats to dichloramine and trichloramine in drinking water. AB - The subchronic toxicity of 0.2-200 ppm dichloramine and 0.2-90 ppm trichloramine in the drinking water of rats was investigated using biochemical, hematological, and histopathological parameters. Animals in the highest dose groups consumed 5 15% less fluid than controls with no significant decrease in body weight gain. No clinical signs of toxicity were observed in either case. Both males and females dosed with 90 ppm trichloramine had significantly increased relative kidney/body weights and the females had increased hepatic glutathione S-transferase and UPD glucuronosyltransferase activities. No significant changes were detected in other xenobiotic metabolizing enzymes or in serum biochemistry, urine biochemistry, or hematology. Both dichloramine and trichloramine induced minimal to mild adaptive histopathological changes in thyroids and kidneys of animals of both sexes. Dichloramine, but not trichloramine, was associated with histological changes in the gastric cardia characterized by epithelial hyperplasia at concentrations of 2 ppm and above in the males and 200 ppm in the females. This study indicates that dichloramine produced mild histological effects at drinking water concentrations of >0.2 ppm in males (0.019 mg/kg/day) and >2 ppm in females (0.26 mg/kg/day) while trichloramine produced biochemical and mild histological effects at levels of >2 ppm both in males (0.23 mg/kg/day) and in females (0.29 mg/kg/day). PMID- 10854127 TI - Hazard identification and dose response of inhaled nickel-soluble salts. AB - A substantial body of occupational epidemiology data has shown that exposure to mixed soluble and insoluble nickel causes the development of lung and nasal cancer. However, due to coexposure of these populations to soluble and insoluble forms of nickel, and limitations in exposure measurements, the contribution of soluble nickel is difficult to determine. Soluble nickel was negative in an NTP inhalation bioassay, while there was some evidence for tumorigenicity in rats for less soluble nickel oxide, and there was clear evidence for tumorigenicity of insoluble nickel subsulfide in rats. Results of parenteral assays follow a similar pattern, but provide evidence of weak carcinogenicity of soluble nickel. Kinetic factors also indicate that exposure to soluble nickel alone has a low carcinogenic potential. Overall, we conclude that the carcinogenic activity of insoluble nickel compounds should not be used to predict the carcinogenic potential of water-soluble nickel salts. The overall data suggest a nonlinear dose-response relationship for carcinogenicity, but the data are insufficient to determine the doses at which such nonlinearities occur. Under the U.S. EPA's 1996 proposed "Guidelines for Carcinogen Risk Assessment," inhaled soluble nickel compounds would be classified as "cannot be determined," because the existing evidence is composed of conflicting data. A reference concentration of 2 x 10(-4) mg Ni/cu x m was calculated, based on lung fibrosis in male rats observed in the NTP study. PMID- 10854128 TI - Hazard identification and dose response of ingested nickel-soluble salts. AB - People can ingest soluble nickel compounds as a normal constituent of food or as a contaminant in drinking water. This paper presents an assessment of the noncancer and cancer human health risks from ingestion of soluble nickel compounds. A reference dose (RfD) of 8 x 10(-3) mg Ni/kg/day in addition to the amount in food was calculated, based on albuminuria in female rats exposed to nickel sulfate in drinking water for 6 months (A. Vyskocil et al., 1994, Hum. Exp. Toxicol. 13, 689-693). This RfD is comparable to the current RfD based on decreased body weight in a chronic feeding study in rats (A. M. Ambrose et al., 1976, J. Food Sci. Technol. 13, 181-187). The potential for nickel-induced reproductive toxicity was also taken into account in the derivation of the RfD. There are a number of negative animal bioassays with soluble nickel salts, but all of them have deficiencies that preclude a definitive conclusion. According to EPA's 1996 draft cancer guidelines, the carcinogenic potential of oral exposure to soluble nickel "cannot be determined because there are inadequate data to perform an assessment." PMID- 10854129 TI - O6-benzylguanine-mediated enhancement of nitrosourea activity in Mer- central nervous system tumor xenografts--implications for clinical trials. AB - PURPOSE: To evaluate the role of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) plus O6-benzylguanine (O6-BG) in the treatment of both Mer+ and Mer- tumors. METHODS: The effect of pretreatment with O6-BG on the activity of BCNU against Mer- human central nervous tumor xenografts D-54 MG and D-245 MG was evaluated in athymic nude mice. RESULTS: BCNU (1.0 LD10; dose lethal to 10% of treated animals) produced growth delays of 8.9 days and 7.5 days and tumor regressions in six of ten and one of nine animals against D-54 MG, which was derived from a human malignant glioma xenograft. Dose reduction of BCNU to 0.38 LD10 eliminated antitumor activity. The combination of BCNU (0.38 LD10) plus O6-BG produced growth delays of 8.8 days and 7.9 days, with tumor regressions in four of ten and two of nine animals, respectively. BCNU (1.0 LD10) produced a growth delay of 49.8 days and ten of ten tumor regressions against D-245 MG, which was derived from a glioblastoma multiforme. BCNU (0.38 LD10) produced a growth delay of 19.4 days, with nine of ten tumor regressions. The combination of BCNU (0.38 LD10) plus O6-BG produced a growth delay of 65.7 days and seven of eight tumor regressions. CONCLUSION: These results suggest that the combination of BCNU plus O6-BG may be a rational intervention for both Mer+ as well as Mer- tumors. PMID- 10854130 TI - Induction of oocyte maturation by jun-N-terminal kinase (JNK) on the oncogenic ras-p21 pathway is dependent on the raf-MEK signal transduction pathway. AB - PURPOSE: We have previously found that microinjection of activated MEK (mitogen activated kinase kinase) and ERK (mitogen-activated protein; MAP kinase) fails to induce oocyte maturation, but that maturation, induced by oncogenic ras-p21 and insulin-activated cell ras-p21, is blocked by peptides from the ras-binding domain of raf. We also found that jun kinase (JNK), on the stress-activated protein (SAP) pathway, which is critical to the oncogenic ras-p21 signal transduction pathway, is a strong inducer of oocyte maturation. Our purpose in this study was to determine the role of the raf-MEK-MAP kinase pathway in oocyte maturation and how it interacts with JNK from the SAP pathway. METHODS: We microinjected raf dominant negative mutant mRNA (DN-raf) and the MEK-specific phosphatase, MKP-T4, either together with oncogenic p21 or c-raf mRNA, into oocytes or into oocytes incubated with insulin to determine the effects of these raf-MEK-MAP kinase pathway inhibitors. RESULTS: We found that oocyte maturation induced by both oncogenic and activated normal p21 is inhibited by both DN-raf and by MKP-T4. The latter more strongly blocks the oncogenic pathway. Also an mRNA encoding a constitutively activated MEK strongly induces oocyte maturation that is not inhibited by DN-raf or by MKP-T4. Surprisingly, we found that oocyte maturation induced by JNK is blocked both by DN-raf and MKP-T4. Furthermore, we discovered that c-raf induces oocyte maturation that is inhibited by glutathione S-transferase (GST), which we have found to be a potent and selective inhibitor of JNK. CONCLUSION: We conclude that there is a strong reciprocal interaction between the SAP pathway involving JNK and the raf-MEK-MAP kinase pathway and that oncogenic ras-p21 can be preferentially inhibited by MEK inhibitors. The results imply that blockade of both MEK and JNK-oncogenic ras-p21 interactions may constitute selective synergistic combination chemotherapy against oncogenic ras induced tumors. PMID- 10854131 TI - A comparison of the effects of nine folate analogs on early and late murine hematopoietic progenitor cells in vitro. AB - PURPOSE: Since the clinical introduction of the antifolates aminopterin (AMT) and methotrexate (MTX) many promising analogs have been developed. A common feature of these compounds is their ability to induce bone marrow suppression. However, few studies have been undertaken on the effect of the folic acid analogs on the cells comprising the hematopoietic system. METHODS: In this paper we describe the effects of the novel thymidylate synthase (TS) inhibitors raltitrexed (Tomudex, ZD1694), AG337 (nolatrexed, Thymitaq), and the two closely related analogs 5,8 dideazaisofolic acid (IAHQ2a) and 2-desamino-2-methyl 5,8-dideazaisofolic acid (IAHQ2c), the glycinamide-ribonucleosyl (GAR) transformylase inhibitor lometrexol (DDATHF), and the dihydrofolate reductase (DHFR) inhibitors MTX, AMT, trimetrexate (TMTX), and edatrexate (EDX) on purified populations of early and late murine hematopoietic progenitor cells. RESULTS/CONCLUSION: All the antifolates inhibited bone marrow proliferation in suspension cultures and all drugs except DDATHF inhibited colony formation by more mature progenitor cells (CFU-C) in clonogenic assays. The lipophilic agents TMTX and AG337 were most toxic, totally abolishing CFU-C colony formation at high concentrations. When IAHQ2c, raltitrexed, DDATHF, and MTX were investigated further for effects on the immature high proliferative potential colony-forming cells (HPP-CFCs) in semisolid and limiting dilution cultures, none of these agents were found to be toxic to the HPP-CFC, but induced a reversible developmental arrest in the progenitor cell population. PMID- 10854132 TI - Pharmacokinetics of intraperitoneal hyperthermic perfusion with mitoxantrone in ovarian cancer. AB - PURPOSE: Theoretical data and experimental assumptions indicate that intraperitoneal hyperthermic chemotherapy may play a role in the treatment of peritoneal carcinomatosis. The feasibility, tolerability and pharmacokinetics of intraperitoneal hyperthermic perfusion with mitoxantrone were studied in patients with pretreated ovarian cancer. METHODS: After cytoreductive surgery, 11 patients underwent intraperitoneal hyperthermic perfusion with mitoxantrone. A heated (42 43 degrees C) solution of the drug (28 mg/m2) was recycled through a perfusion apparatus into the abdominal cavity for 90 min. Treatment was repeated every month for two to four cycles. In six patients blood and peritoneal perfusate samples were collected at 0.5, 1, 1.5, 2, 4, 8, 16 and 24 h after drug administration and mitoxantrone was assayed by an HPLC method. RESULTS: Although treatment was generally well tolerated, all patients developed transient intestinal subocclusion. Maximal mitoxantrone plasma concentrations (Cmax), times to Cmax (Tpeak) and area under the curves (AUC) were highly variable between subjects (Cmax 14-337 ng/ml; Tpeak 0.5-8 h; AUC 222-4130 ng x ml(-1) x h). The plasma to peritoneal fluid AUC ratio was significantly higher during the second (0.177) than during the first cycle (0.066), suggesting a cycle-dependent increase in systemic bioavailability. Furthermore, when comparing present data with those reported previously, hyperthermic perfusion may have lowered the mitoxantrone levels in the peritoneal fluid without greatly influencing plasma levels. CONCLUSIONS: Intraperitoneal mitoxantrone administered under hyperthermia to advanced ovarian cancer patients is feasible and well tolerated. Mitoxantrone pharmacokinetics may be altered by repeated intraperitoneal administration (increased bioavailability) and by hyperthermic perfusion (possibly, increased peritoneal tissue uptake). PMID- 10854133 TI - The pharmacokinetics of a 1-h paclitaxel infusion. AB - PURPOSE: To characterize the disposition of paclitaxel (PAC) after a 1-h infusion in humans and define if possible a pharmacodynamic relationship between PAC disposition and the observed toxicity. PATIENTS AND METHODS: PAC pharmacokinetics were studied in 43 courses of therapy in 30 patients (30 first course, 13 PK third course). PAC was administered at 150, 175, 200, 225 and 250 mg/m2 by a 1-h infusion to patients with advanced cancer (lung, breast, ovarian, cervix, and head and neck). PAC was quantified by high-performance liquid chromatography (HPLC). Pharmacokinetic parameters were calculated by noncompartmental and model dependent methods. RESULTS: Increases in the area under the curve and the peak plasma concentration were not proportional to increases in the dose. However, the deviation from linearity is rather moderate. The dose-limiting toxicity was central neuropathy which was not associated with pharmacokinetic deviations. Owing to the absence of grade 3 or 4 myelotoxicity, no clear correlation between this toxicity and pharmacokinetic parameters could be established. CONCLUSION: Within the evaluated dose range of the 1-h infusion there was only a moderate nonlinear disposition of PAC in humans and therefore a dose of 225 mg/m2 is recommended as safe. The observation of central neuropathy could not be directly related to a pharmacokinetic parameter. The complexity of the formulation which included Cremophor EL and ethanol may offer an explanation for the observed central neurotoxicity. PMID- 10854134 TI - In vitro synergistic effects of vinflunine, a novel fluorinated vinca alkaloid, in combination with other anticancer drugs. AB - PURPOSE: Vinflunine (20'-20'-difluoro-3',4'-dihydrovinorelbine), a novel derivative of vinorelbine characterized by marked antitumour activity in vivo in a series of experimental murine and human tumours is currently undergoing phase I evaluation. To investigate its potential for inclusion in combination chemotherapy regimens, this preclinical study was undertaken. The in vitro cytotoxicity of vinflunine incubated simultaneously with one of the following drugs was investigated: camptothecin, cisplatin, doxorubicin, etoposide, 5 fluorouracil, gemcitabine, mitomycin C, paclitaxel or vinorelbine. METHODS: The combinations were first evaluated in vitro against the A549 human non-small-cell lung cancer cell line using median-effect analyses. RESULTS: The results revealed synergistic cytotoxicity when vinflunine was combined with cisplatin, mitomycin C, doxorubicin or 5-fluorouracil. Synergy was also observed when testing similar combinations against CCRF-CEM human leukaemia cells. Finally, these findings were comparable with those resulting from such combinations involving vinorelbine instead of vinflunine. CONCLUSION: Vinflunine appears a promising candidate for combining with other anticancer drugs. PMID- 10854135 TI - Dihydropyrimidine dehydrogenase activity in normal, inflammatory and tumour tissues of colon and liver in humans. AB - BACKGROUND/PURPOSE: Dihydropyridmidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in the catabolism of 5-fluorouracil (5FU). Although this catabolism is likely to occur in the liver in humans, there may be a local inactivation in tumours, modifying the efficacy of 5FU. The aim of this study was to examine the DPD activity in normal, inflammatory and malignant tissues from both the colon and the liver to assess the modifications of DPD activity in the process of tumourigenesis. METHODS: DPD activity was evaluated in 107 patients, corresponding to 194 samples (70 colorectal tumour and normal colon, nine metastases secondary to a colon cancer, ten inflammatory colon, 20 samples of normal liver, seven from primary liver cancer, and eight from inflammatory liver). DPD activity was determined using an enzymatic reaction followed by analysis of 5FU and its catabolite dihydro-5FU by high-performance liquid chromatograph. Results were expressed as pmol of 5FU catabolized/min x mg protein. RESULTS: DPD was highly variable in tumour and normal tissues, both from colon and liver. In colon, the correlation between DPD activity in tumour and normal mucosa was weak, even if it was statistically significant due to the higher number of samples. In inflammatory colon tissue (ulcerative colitis or Crohn's disease), DPD activity was significantly higher than in normal tissue (P = 0.006). In liver metastases from colon cancer, DPD activity was not significantly different from that observed in primary colon tumour (P = 0.32). In liver, DPD activity was significantly lower in primary liver tumour than in uninvolved liver specimens (P = 0.001). In inflammatory liver tissue (hepatitis), DPD activity ranged between normal and tumour tissues, and did not differ significantly either from normal tissue or primary liver cancer. CONCLUSIONS: DPD activity was modified in colon and in liver during a pathological process and the dysregulation of DPD increased from a benign to a malignant tissue. PMID- 10854136 TI - Investigation of bioavailability and pharmacokinetics of treosulfan capsules in patients with relapsed ovarian cancer. AB - PURPOSE: Treosulfan (L-threitol-1,4-bis-methanesulfonate, Ovastat) is a prodrug of a bifunctional alkylating agent with activity in ovarian carcinoma and other solid tumors. In a pharmacologic study of the bioavailability of treosulfan in a capsule formulation, patients with relapsed ovarian carcinoma were treated with alternating doses of oral and intravenous (i.v.) treosulfan of 1.5 or 2.0 g daily for 5 to 8 days. METHODS: A sensitive method for the determination of treosulfan in plasma and urine by reversed-phase high-performance liquid chromatography had previously been developed. Pharmacokinetic analyses of treosulfan were carried on plasma and urine samples from 20 i.v. courses and 20 courses of oral administration. RESULTS: The bioavailability ratio (f) of oral to i.v. administration was calculated as 0.97 +/- 0.18 (mean +/- SD) using the values AUC oral = 82.1 +/- 39.4 microg/ml h and AUC i.v. = 85.4 +/- 30.3 microg/ml h. The peak plasma concentration cmax (29 +/- 14 microg/ml vs 65 +/- 23 microg/ml) was significantly (P < 0.01) higher after i.v. administration and the tmax after oral administration was 1.5 +/- 0.34 h. The terminal half-life of treosulfan was about 1.8 h. The mean urinary excretion of the parent compound was about 15% of the administered total dose over 24 h (range 6-26%). CONCLUSIONS: The high and relatively constant bioavailability of treosulfan indicates that capsules provide a satisfactory noninvasive treatment alternative. A feasible and reliable oral treosulfan formulation could provide the basis for the development of long-term low-dose outpatient treatment of patients with malignant diseases. PMID- 10854137 TI - Development of a schedule-dependent population pharmacodynamic model for rhizoxin without quantitation of plasma concentrations. AB - In previous phase I reports of short bolus infusion of rhizoxin, problems in assay sensitivity prevented the description of pharmacokinetic-pharmacodynamic relationships, and a pharmacologically guided approach to dose escalation was deemed not feasible. In this report, we describe a mathematical model, which explains the schedule-dependent interpatient pharmacodynamic variability of rhizoxin administered on a continuous infusion schedule. Using patient demographic and toxicity data from 45 patients treated in a phase I dose and duration escalation study of rhizoxin, we sought to model the nadir neutrophil count. We hypothesized that a surrogate derived variable based on dose and duration would reflect a pharmacokinetic parameter that would be a significant covariate. Multiple linear regression analysis was carried out to determine the other significant covariates. Dose/m2 x Log(DUR/ALB) was significantly correlated with the LogANCnadir (Log10 neutrophil nadir; r = 0.56, P < 0.001). Other significant covariates included baseline performance status (PS), baseline serum bilirubin (BIL), and Log10 baseline neutrophil count (LogANCbaseline). Model bias and precision were assessed using the mean prediction error (MPE) and the root mean square error (RMSE) of the ANCnadir, respectively. We constructed 1-4 covariate models. The variability of ANCnadir was modeled with good precision and accuracy with a 4-covariate model (MPE and RMSE 0.113 +/- 0.182 x 10(3) cells/microl and 1.22 x 10(3) cells/microl, respectively). This model should be validated and improved on with further clinical data. We believe that such pharmacodynamic modeling should be explored further to determine its performance and clinical relevance compared with modeling using pharmacokinetic parameters. PMID- 10854138 TI - The pharmacokinetics of liposomal encapsulated daunorubicin are not modified by HAART in patients with HIV-associated Kaposi's sarcoma. AB - PURPOSE: To investigate the pharmacokinetics of liposomal daunorubicin (DaunoXome) administered alone or in combination with antiviral therapy including protease inhibitors (PI) to HIV-positive patients affected by Kaposi's sarcoma (KS). PATIENTS AND METHODS: A group of 18 patients with extensive or rapidly progressing AIDS-related KS received DaunoXome at a dose of 40 mg/m2 alone or in association with a triple combination therapy consisting of one PI plus two nucleoside reverse transcriptase inhibitors (NRTI). Daunorubicin pharmacokinetics were determined in a total of 23 cycles, 6 with DaunoXome alone, 9 in combination with indinavir, 6 with ritonavir and 2 with saquinavir. Plasma samples were obtained at different times during the 72 h after DaunoXome administration. Daunorubicin and daunorubicinol plasma levels were determined by high-performance liquid chromatography. RESULTS: After the DaunoXome infusion, daunorubicin was rapidly cleared from the body following, in most cases, a one-compartment open kinetic model. The daunorubicin peak concentrations, clearances and elimination half-lives were (means +/- SD): 16.3 +/- 2.8 microg/ml, 0.3 +/- 0.1 l/h per m2 and 5.6 +/- 2.6 h after DaunoXome alone; 15.1 +/- 4.9 microg/ml, 0.5 +/- 0.3 l/ h per m2 and 5.8 +/- 2.1 h after the combination with indinavir; and 14.5 +/- 2.8 microg/ml, 0.4 +/- 0.2 l/h per m2 and 6.5 +/- 3.9 h after the combination with ritonavir. In all groups, daunorubicinol plasma levels were approximately 25-30 times lower than those of the parent drug. CONCLUSION: Our data show that there are no significant differences in the pharmacokinetic parameters of daunorubicin in patients receiving DaunoXome in combination with indinavir and ritonavir compared with those in patients not receiving PIs. Therefore in patients affected by AIDS-related KS treated with Highly Active AntiRetroviral Therapy (HAART) there is no pharmacokinetic justification for reducing the doses of DaunoXome. PMID- 10854139 TI - New chemotherapeutics in malignant mesothelioma: effects on cell growth and IL-6 production. AB - PURPOSE: The benefits of chemotherapy can be assessed in terms of tumour shrinkage, prolongation of life or simply palliation of symptoms. In the study reported here, in vitro correlates of these parameters were sought as a rational guide to the choice of newer agents in the clinic. METHODS: The cytotoxicity and effects on IL-6 production of ten chemotherapy agents representing four different classes of drugs were tested against a panel of five mesothelioma cell lines. RESULTS: The mesothelioma cells were more sensitive to the action of irinotecan (and its active metabolite SN38) and gemcitabine than the control cell lines. Gemcitabine and to a lesser extent irinotecan inhibited the secretion of the proinflammatory cytokine IL-6 at concentrations of each drug that produced only small decreases in cell viability. This effect was not seen in cells treated with docetaxel or vindesine. Higher doses of gemcitabine and irinotecan caused a surge in IL-6 release and this was not due to release of intracellular stores of IL-6 through lysis of the cells. CONCLUSIONS: These results suggest that irinotecan and gemcitabine are not only more likely to be active against mesothelioma than other new chemotherapy agents but may also produce a palliative effect in nonresponders to these agents by decreasing the secretion of IL-6. PMID- 10854140 TI - A phase II study of temozolomide in hormone-refractory prostate cancer. AB - INTRODUCTION: Hormone-refractory disseminated prostate cancer is a major oncological problem. Preclinical studies with temozolomide, an oral alkylating agent, in prostate cancer have shown encouraging results. In phase I studies the safety of temozolomide in non-prostate cancer patients has been demonstrated. Based on these results, a phase II study of temozolomide in patients with metastatic disease who had developed progressive symptomatic disease while on antiandrogen therapy, was initiated. METHODS: A group of 18 patients started a 5 day temozolomide regimen, with a 28-day treatment cycle. Response parameters (prostate-specific antigen, bone scan, quality of life questionnaire) and toxicity (common toxicity criteria for international studies) were recorded at regular intervals. RESULTS: Of the 18 patients, 16 were evaluable by completing two or three cycles. All patients developed progressive disease within two cycles, except one who had progressive disease at the end of cycle 3. Of the 16 evaluable patients, 11 developed new bone metastases (bone scan), 1 developed lung metastases, 4 had progressive disease as reflected by a 25% increase in serum PSA together with subjective progression, and 7 and 5 had progressive disease as reflected by decreased quality of life and increased pain score, respectively. Toxicity was limited to nausea and vomiting, which was effectively treated with antiemetic medication, and anemia and thrombocytopenia, which returned to normal values within 1 week. DISCUSSION: Treatment with temozolomide was generally well tolerated, with occasionally moderate toxicity. As all patients developed progressive disease the results are rather discouraging. Temozolomide is ineffective for the treatment of patients with symptomatic progressive hormone-refractory prostate cancer. PMID- 10854141 TI - Ifosfamide and vinorelbine in advanced pretreated ovarian cancer: a phase II study. AB - PURPOSE: The response rate to salvage chemotherapy in advanced ovarian cancer (AOC) has been disappointing in patients who do not respond or relapse after platinum-containing regimens. Ifosfamide (IFO) showed an overall response rate of 20% and vinorelbine (VNR) 15.6%. Trials of the association of these two drugs for AOC have not yet been published. PATIENTS AND METHODS: Between April 1996 and August 1997, 17 patients with AOC were treated with intravenous IFO 2000 mg/m2 per day, days 1 to 3, with mesna uroprotection, and VNR 25 mg/m2 per day, days 1 and 8, every 3 weeks. All patients but one had been heavily pretreated. All patients had been treated with platinum compounds and 16/17 with taxanes. RESULTS: All 17 patients were evaluable for toxicity, and 16 for response (one lost to follow-up). One patient showed a partial response, 12 progressive disease and three stable disease. No complete responses were observed. The main toxicity was neutropenia (grade 3-4 in 82% of patients) with neutropenic fever in 17.6% of patients. In 70.5% of patients (19/59 of courses) VNR was not administered on day 8. In four patients (10/59 courses) the dose was reduced by 25% for persistent leukopenia grade 2-3. Other toxicities were not significant. CONCLUSIONS: This combination showed no activity in this set of patients. The poor outcome, as compared with the significant activity reported with the agents used singly, could be ascribed to the patients' characteristics, the low dose intensity of VNR administered and possible cross-resistance between the study drugs and previously used agents. PMID- 10854142 TI - Gene therapy in urologic oncology. PMID- 10854143 TI - Transcriptionally regulated adenoviruses for prostate-specific gene therapy. AB - Most virally based vectors for gene therapy contain viral promoters that are tissue-nonspecific. Consequently, unintended expression of toxic therapeutic genes in normal tissues may potentially occur. We have constructed adenoviruses that contain a bacterial beta-galactosidase (beta-gal) gene (lacZ) under the control of three different prostate-specific promoters: prostate-specific antigen (PSA), probasin, and the mouse mammary-tumor-virus long terminal repeat (MMTV; prostate-specific Ad-lacZ). In general, these prostate-specific Ad-lacZ can effectively transduce and express beta-gal in prostate cells and display weak, if any, expression of beta-gal in nonprostate cells in vitro. In vivo, these adenoviruses showed a high level of beta-gal expression in canine prostate but also disseminated to tissues other than prostate after intraprostatic (i.p.) injection. However, none of the prostate-specific Ad-lacZ expressed beta-gal in these nonprostate tissues. Furthermore, prostate-specific Ad-lacZ expressed beta gal only in xenograft tumors grown in nude mice, derived from human prostate cancer cells DU145 and PPC-1, but showed no beta-gal expression in tumors derived from human bladder-cancer cells RT4. These results indicate that adenoviruses containing prostate-specific promoters may express intended transgenes specifically in prostate in vivo. PMID- 10854144 TI - Osteocalcin-directed gene therapy for prostate-cancer bone metastasis. AB - Osteocalcin (OC) is a major noncollagenous bone protein whose expression is limited almost exclusively to osteotropic tumors and mature calcified tissue (differentiated osteoblasts). The function of OC, a highly conserved gamma carboxyglutamic acid-containing protein, relies in part on its ability to bind hydroxyapatite and act as a chemoattractant for bone-resorbing cells. Serum osteocalcin levels are used clinically as an index of active bone turnover. Research in our laboratory has revealed that OC is expressed in several solid tumors, including osteosarcoma and ovarian, lung, brain, and prostate cancers. Evidence arising from reverse-transcription polymerase chain reaction (RT-PCR; detection of OC mRNA), immunohistochemical staining (detection of OC protein), and transient transfection and reporter assay (detection of OC mRNA transcription) reveals that OC expression is up-regulated in numerous solid tumors, with its expression being further elevated in androgen-independent prostate cancers. A recombinant, replication-defective adenovirus, Ad-OC-TK (OC promoter-driven herpes-simplex-virus thymidine kinase) was constructed and, when combined with the appropriate prodrug, either ganciclovir (GCV) or acyclovir (ACV), was found to be effective at destroying prostate-cancer cell lines in vitro and prostate tumor xenografts in vivo in both subcutaneous and bone sites. Additionally, via use of the OC promoter the supporting bone stromal cells are cotargeted when the prostate cancer interdigitates with bone stroma at the metastatic skeletal sites. Thus, maximal tissue-specific cell toxicity is achieved by the interruption of cellular communication between the prostate cancer and the bone stroma. We describe herein the preclinical foundation as well as the design and implementation of an ongoing phase I clinical trial at the University of Virginia that targets androgen-independent metastatic prostate cancer using the Ad-OC-TK vector. PMID- 10854145 TI - Adenovirus p16 gene therapy for prostate cancer. AB - Surgery, radiation, or hormone deprivation alone does not adequately affect local control of clinical or pathologic stage T3 prostate cancer. Lack of local cancer control ultimately leads to a higher incidence of morbidity, distant metastasis, and decreased survival, with patients having disease-specific mortality exceeding 75%. Other novel therapies against this devastating and common disease are needed for the achievement of long-term local cancer control. For this purpose, therapeutic interventions should target prostate-cancer cells at the molecular and cellular level in ways not possible by current modalities of cancer treatment. Any strategy that can modify the biologic behavior of these cells may potentially have the most significant clinical impact. As prostate cancer represents an accumulation of genetic mutations that causes a prostate cell to lose the ability to control its growth, one new approach against prostate cancer may be gene therapy. Identification of key missing or mutated tumor-suppressor genes that, when replaced, may inhibit or destroy prostate-cancer cells may have the best chance of clinical success. One such gene appears to be tumor-suppressor gene p16 (also known as MTS1, INK4A, and CDKN2). Tumor-suppressor gene p16 is an important negative cell-cycle regulator whose functional loss may significantly contribute to malignant transformation and progression. Alterations in the p16 gene and its protein expression often occur in prostate cancer. An adenoviral vector containing wild-type p16 (Adp16) had a high transduction efficiency in prostate-cancer cells both in vitro and in vivo. Moreover, prostate tumors injected with Adp16 expressed p16 and the adenoviral vector expressed the transgene for up to 14 days. Wild-type p16 inhibited prostate-cancer proliferation in vitro and markedly suppressed tumors in vivo. Pathologic evaluation of the Adp16-treated tumors showed dose-dependent necrosis and fibrosis. Although the mechanism of p16 inhibition in cancer remains to be elucidated, senescence and apoptosis may both be important; however, the data suggest that p16-induced growth inhibition can function independently of the retinoblastoma gene product. PMID- 10854146 TI - Ad5CMVp53 gene therapy for locally advanced prostate cancer--where do we stand? AB - Despite the introduction of screening procedures and an increased public awareness of prostate cancer, a substantial number of patients present with locally advanced prostate cancer. Traditional therapies (such as radiation therapy or radical prostatectomy) applied either alone or in combination fail to control local disease in a large number of cases and have no effect on disseminated disease. Recent advances in molecular oncology and genetics have led to such novel therapies as p53 gene therapy, which we are currently evaluating in a clinical protocol in patients with locally advanced (nonmetastatic) prostatic cancer. Ad5CMVp53 (RPR/INGN 201) has previously shown promise in both patients with lung cancer and those with head and neck cancer. The traditional end points used to appraise prostate cancer preclude rapid evaluation of the patient's disease and prevent modification of the therapeutic strategy, and we suggest that the pathologic stage after therapy be evaluated as an intermediate end point. PMID- 10854147 TI - Suicide gene therapy for prostate cancer using a replication-deficient adenovirus containing the herpesvirus thymidine kinase gene. AB - Current therapies for localized prostate cancer include radical prostatectomy, local radiation therapy, and cryoablation and are associated with a high rate of cure and acceptable morbidity. However, for men who have failed primary curative attempts or have metastatic disease, no effective therapy associated with acceptable morbidity exists. "Suicide" gene therapy delivered alone or in combination with other forms of treatment could potentially provide simultaneous efficacy against localized and systemic disease via the generation of cytotoxic activity and/or systemic immunity to the cancer. In this article we discuss our preclinical and clinical experience with a herpes-simplex-virus thymidine kinase/ganciclovir gene-therapy protocol for prostate cancer. PMID- 10854148 TI - Prospects for herpes-simplex-virus thymidine-kinase and cytokine gene transduction as immunomodulatory gene therapy for prostate cancer. AB - In completed and ongoing clinical trials, adenovirus-mediated (Ad.) expression of herpes-simplex-virus thymidine-kinase (HSV-tk) gene transduction followed by ganciclovir (GCV) therapy has produced limited toxicity and evidence of antitumor activity following injection of the prostate. Furthermore, this system has been shown to direct systemic antitumor activity in several experimental cancer models, including that of prostate cancer, which may serve as the basis for in situ immunomodulatory gene therapy. In a mouse model of prostate cancer, natural killer (NK) cells have been identified as the mediator of antimetastatic activity following Ad.HSV-tk + GCV, resulting in the combination of Ad.HSV-tk and adenovirus-mediated expression of interleukin 12 (Ad.IL-12) to exploit this cytokine's ability to enhance NK proliferation and cytotoxicity. Combination therapy demonstrated superior local and systemic growth suppression over that obtained with either therapy alone. Importantly, when the metastatic tumor burden was increased to an extent that negated the growth-suppressive activity directed by Ad.HSV-tk + GCV or Ad.IL-12 alone, combination therapy continued to demonstrate significant growth suppression. Examination of tumor-infiltrating lymphocytes documented enhanced NK lytic activity following combination therapy. Therefore, it appears that the combination of Ad.HSV-tk and Ad.IL-12 should be validated in a clinical trial for the treatment of prostate cancer. PMID- 10854149 TI - Antitumor efficacy of tumor-antigen-encoding recombinant poxvirus immunization in Dunning rat prostate cancer: implications for clinical genetic vaccine development. AB - One potential use for prostate-cancer-associated genes discovered through ongoing genetics studies entails the construction of virus- or plasmid-based recombinant vector vaccines encoding these new tumor-associated antigens (TAA) to induce TAA specific immune responses for the prevention or therapy of prostate cancer. Clinical trials evaluating prototypes of such recombinant vaccines are under way. TAA-encoding recombinant vector vaccines, however, have not previously been evaluated in a prostate-cancer animal model. For assessment of the potential susceptibility of prostate cancer to genetic immunization strategies using TAA encoding recombinant vectors, the antitumor efficacy of a model recombinant viral vector encoding a TAA was evaluated in rat Dunning prostate cancer. Recombinant vaccinia was chosen as a prototype virus vector encoding a TAA for these studies, and beta-galactosidase was chosen as a model target TAA. Dunning AT-2 cells were transduced with a retroviral vector to express beta-galactosidase, and the susceptibility of tumorigenic AT-2-lacZ cells to immunization with vaccinia-lacZ was measured using protection studies in Copenhagen and nu/nu rats. Stably transduced AT-2-lacZ cells expressing beta-galactosidase as measured by enzymatic substrate-based assays were found to retain their tumorigenicity in vivo despite abundant expression of rat major histocompatibility complex (MHC) class I. Immunization with model TAA-encoding recombinant vaccinia-lacZ conferred significant protection against subsequent growth of AT-2-lacZ cells in vivo (P = 0.01); however, the efficacy of such immunization was markedly dependent on the volume of tumor challenge. The antitumor efficacy of TAA-encoding recombinant vaccinia immunization was abrogated in nu/nu rats, suggesting a T-cell-dependent mechanism of activity. These studies suggest that prostate cancer may be a suitable target for immunization strategies using TAA-encoding recombinant vectors. Such immunization strategies may be more effective in settings of minimal cancer burden. PMID- 10854150 TI - Gene therapy for prostate cancer at the University of California, Los Angeles: preliminary results and future directions. AB - The treatment options for patients with advanced prostate cancer are limited. Because of recent advances in the understanding of the molecular biology of prostate cancer, the accessibility of the prostate for injection, and the availability of gene promotors that allow tissue-specific expression of therapeutic gene products, gene therapy for prostate cancer has realized significant achievements in recent years. What once belonged to the realm of basic science is now entering the domain of phase II clinical trials. In this review, current results and future directions in prostate gene therapy at the University of California, Los Angeles, are discussed. PMID- 10854151 TI - Gene therapy for bladder cancer. AB - Tumor-suppressor genes can be transferred into tumor cells in vivo using a replication-defective adenoviral vector. P53 mutations are frequent in bladder cancer, and adenovirus-mediated p53 gene transfer is growth-inhibitory to bladder cancer cells in vitro. The vector Ad5CMV-P53, which contains human wild-type p53, is being administered intravesically to patients with bladder cancer in a phase I clinical trial. The results of this study will provide the basis for phase II and phase III trials in which gene therapy will be integrated with existing therapies for improved local control and opportunities for bladder preservation. PMID- 10854152 TI - Intratumoral interleukin 2 for renal-cell carcinoma by direct gene transfer of a plasmid DNA/DMRIE/DOPE lipid complex. AB - Metastatic renal-cell carcinoma (RCC) is not responsive to conventional cytotoxic chemotherapy, but a subset of patients achieve a durable remission with the use of interleukin-2 (IL-2). IL-2 is currently the only Food and Drug Administration (FDA)-approved treatment for metastatic RCC, and it benefits 10-20% of those who receive it. However, it is accompanied by significant, occasionally life threatening toxicity. Attempts to maintain the efficacy of IL-2 while minimizing systemic side effects have led to the development of IL-2 gene therapies. Leuvectin is a plasmid DNA/lipid complex composed of a plasmid DNA expression vector (VCL-1102, 30) encoding human IL-2 complexed in a 5:1 mass ratio with DMRIE/DOPE lipid (1,2-dimyristyloxypropyl-3-dimethylhydroxyethyl ammonium bromide/dioleoylphosphatidyl ethanolamine), which has been developed for the treatment of malignancy. DMRIE/DOPE is a cationic lipid that has been shown to facilitate in vitro transfection of plasmid DNA. It has been demonstrated that in vitro transfection with the IL-2 plasmid DNA/DMRIE/DOPE complex results in the expression of sustained levels of biologically active IL-2. Established human tumor cell lines and primary human tumor cells obtained from biopsies are readily transfected in vitro, resulting in the expression of IL-2. Following in vitro transfection, IL-2 expression has been found to persist for up to several weeks in primary tumor cells. In preclinical efficacy studies in a murine model of renal-cell carcinoma the direct intratumoral administration of an IL-2 plasmid DNA/DMRIE/DOPE complex resulted in complete tumor regression in the majority of mice. In preclinical animal-safety studies, repeated administration of Leuvectin was safe and well tolerated. Following these promising preclinical trials, Leuvectin has been taken into clinical trial. The results of two early studies indicate that Leuvectin is safe, is free of systemic toxicity, and has biologic activity. PMID- 10854153 TI - B7-1 gene-modified autologous tumor-cell vaccines for renal-cell carcinoma. AB - In recent years, interest in the development of immunologic approaches to malignancies has increased, and there is good evidence that the growth of renal cell carcinoma (RCC) can be modulated by the host's immune system. Indeed, use of the immunomodulatory cytokine interleukin-2 (IL-2) has been approved for the treatment for this disease. The efficacy of this approach remains low, and there is no other reasonable conventional therapy for patients with metastatic RCC. Therefore, there is a need for the development of novel treatment strategies. The development of autologous tumor-cell vaccines that have been genetically modified to become more immunogenic is an approach that is actively being studied. One of the genetic manipulations that is being employed by several groups is the induction of overexpression of B7-1 to provide costimulation to tumor-reactive T cells. The rationale for this strategy is that T-cells need two signals before they can mount a cytotoxic response: the binding of the T-cell receptor (TCR) to an antigenic peptide presented on major histocompatibility complex (MHC) molecules and the binding of CD28 to B7-1. Since B7-1 is not normally expressed by RCC cells, the expression forced by transfection of an exogenous B7-1 gene could make the tumor cells more immunogenic. This has been shown to be the case in mice, in which the injection of tumor cells transfected with B7-1 can result in the T-cell-mediated rejection of unmanipulated parental tumor cells. We have applied this approach to the treatment of patients with metastatic RCC. Patients enrolled on our phase I protocol are treated with autologous tumor cells modified to express B7-1, which functions as a tumor vaccine. Primary tumors or metastases are resected from the patients. The tumor cells are adapted to in vitro culture, infected with a recombinant adenoviral vector containing human B7-1 cDNA driven by the cytomegalovirus (CMV) promoter, radiated, and stored in liquid nitrogen. Aliquots of the B7-1 gene-modified tumor cells are given to the patients as a vaccine at varying intervals according to a dose-escalation scheme. The patients also receive systemic IL-2 for the dual purpose of providing accepted therapy for this disease as well as expanding the tumor-reactive T-cells activated by the vaccine. The immunogenicity and toxicity of the vaccine as well as the clinical response are being assessed in three to five patients at each of three dose levels. PMID- 10854154 TI - Interstitial laser coagulation technique. Executive summary. AB - In March 1999, it was again my privilege to moderate a meeting of distinguished colleagues organized by Indigo Medical, Inc. as their BPH Advisory Panel. This executive summary highlights the most important conclusions of our day-long session. Details can be found in the subsequent sections of the publication. Our discussion of ILC technique is based on several exciting new concepts in prostatic anatomy. These concepts are derived from anatomical studies using fresh cadavers that have revealed the structures of the prostate in greater detail than ever before. Such findings are important because the great majority of descriptions of this anatomy are incomplete or inaccurate. An overview of the current clinical anatomy is found in Section VI. PMID- 10854155 TI - Interstitial laser coagulation technique: ILC blueprint. PMID- 10854156 TI - Interstitial laser coagulation technique: clinical research updates. PMID- 10854157 TI - Interstitial laser coagulation technique: considerations for office -based ILC. PMID- 10854158 TI - Interstitial laser coagulation techniques: local anesthesia techniques. PMID- 10854159 TI - Interstitial laser coagulation technique. Anatomic review. PMID- 10854160 TI - Interstitial laser coagulation technique. Bibliography. PMID- 10854161 TI - Identification by mass spectroscopy of three major early proteins associated with virosomes in vaccinia virus-infected cells. AB - Virosomes are cytoplasmic sites of replication of vaccinia virus DNA and were prepared from virus-infected HeLa cells. The early virosomal proteins were 35S labelled and SDS polyacrylamide gel electrophoresis revealed the presence of three major early 35S-labelled proteins of 34, 24 and 45 kDa. The masses of molecules present in the 34 and 24 kDa proteins were measured by the convenient and sensitive MALDI TOF mass spectroscopy technique. Identification of the three virosomal proteins was carried out by MALDI mass spectroscopy of corresponding tryptic digests. For each protein at least 13 measured masses matched, within less than 0.1 Da, calculated tryptic peptides of the vaccinia virus proteins H5R (34 kDa), E3L (24 kDa) and E5R (45 kDa). In addition, virosomes contained several structural proteins from the infecting virus and a 45 kDa keratin-related protein. This work demonstrates directly that the abundant early vaccinia virus proteins H5R, E3L and E5R are associated with the virosomes. PMID- 10854162 TI - The live attenuated subgroup B respiratory syncytial virus vaccine candidate RSV 2B33F is attenuated and immunogenic in chimpanzees, but exhibits partial loss of the ts phenotype following replication in vivo. AB - The cold-adapted (ca), temperature-sensitive (ts) respiratory syncytial virus (RSV) subgroup B vaccine candidate, designated RSV 2B33F, was found previously to be restricted in replication, immunogenic, and protective against wild-type (wt) virus challenge in rodents and African green monkeys. We sought to investigate the level of attenuation, immunogenicity and genetic stability of this vaccine candidate in seronegative chimpanzees. The 2B33F vaccine candidate was attenuated in chimpanzees and manifested a ten- and 1000-fold restriction in replication in the upper and lower respiratory tracts respectively, compared with its wt RSV 2B parent virus. Despite this attenuation, chimpanzees immunized with RSV 2B33F were completely resistant to respiratory tract disease and virus replication upon challenge with wt virus. The ts phenotype of the RSV 2B33F mutant exhibited some alteration during replication in vivo in three of four chimpanzees tested. Virus present in nasopharyngeal swab or tracheal lavage secretions of these three chimpanzees was biologically cloned by plaque passage in Vero cells at permissive temperature. The plaque progeny retained the ts phenotype, but uniformly produced plaques at 39 and 40 degrees C to a level intermediate between that of the 2B33F input virus and the 2B wt parent virus, indicating that partial loss of the level of temperature sensitivity occurred following replication in vivo. The implications of these findings for RSV vaccine development are discussed. PMID- 10854163 TI - Identification of the US3 gene product of BHV-1 as a protein kinase and characterization of BHV-1 mutants of the US3 gene. AB - We have identified the product of the US3 gene of bovine herpes virus type 1 (BHV 1), which is homologous to the herpes simplex virus type 1 (HSV-1) US3 protein kinase (PK) gene. The antibodies against the BHV-1 US3 gene product reacted with a 58 kDa polypeptide in BHV-1 infected cells and the 58 kDa polypeptide purified by immuno-precipitation demonstrated PK activity. Recent reports indicating that the US3 gene of HSV-1 is involved in the blockage of apoptosis in virus infected cells. As to the apoptosis in BHV-1 infected cells, we found following: (1) no apoptosis was observed in cells infected with wild type BHV-1 and the US3 mutants (2) the apoptosis induced by the osmotic shock of sorbitol treatment was blocked when cells were infected by the wild type BHV-1 (3) the US3 mutants of BHV-1 blocked the apoptosis of sorbitol treated cell, but the suppressive effect was delayed relative to that of wild type BHV-1 (4) the other BHV-1 mutants, with the intact US3 gene but with some other non-essential gene (genes) deleted behaved similar way to the US3 mutant. It is concluded that the US3 gene of BHV-1 is not directly involved in the blockage of apoptosis in infected cells. PMID- 10854164 TI - Genetic variability of hepatitis E virus within and between three epidemics in India. AB - Hepatitis E virus (HEV) is an important cause of epidemic and sporadic acute viral hepatitis in many developing countries, including India. We evaluated the genetic variability within two regions (a 476-nt long ORF1 segment and a 304-nt long ORF2 segment) from specimens collected during three outbreaks in the cities of Karnal (1987), Yamunanagar (1989), and Meerut (1996), India, and from one patient, residing in Lucknow, India, who had a case of sporadic hepatitis (1996). Within an outbreak, sequences in the ORF1 and ORF2 regions were 99.3-100.0% identical. However, when strains were compared between outbreaks, identity in the ORF1 and ORF2 region was 97.1-99.2 and 96.4-100.0%, respectively. A comparison of these sequences to previously published Indian ORF1 and ORF2 sequences revealed even lower similarities, 95.2-98.5 and 95.1-98.7%, respectively. One patient in the Meerut outbreak had genomic sequences that differed substantially from the other patients affected during this outbreak and probably reflected a sporadic infection. The sporadic hepatitis E strain from Lucknow clustered with a previously described HEV strain from a patient with fulminant hepatic failure (FHF). Our data suggest that the ORF1 and ORF2 segments can be used to study the molecular epidemiology of HEV infection and indicate that much remains to be determined about the genetic variability of Indian HEV strains. PMID- 10854165 TI - Fine characterization of a V3-region neutralizing epitope in human immunodeficiency virus type 2. AB - We have previously identified two distinct antigenic sites in the third variable region (V3) of human immunodeficiency virus type 2 (HIV-2) corresponding to the principal neutralizing determinant (PND) of HIV-1, the conserved Phe-His-Ser-Gln and Trp-Cys-Arg motifs (positions 315-318 and 329-331), which possibly interact to form a discontinuous antigenic site. The aim of this study was to further identify and characterize the immunogenic sites in the V3-loop of HIV-2 that are important in the binding of neutralizing antibodies and to study in detail the importance of different configurations of peptides corresponding to this region. Peptides representing modifications of the V3-region of HIV-2(SBL6669-ISY) were used for immunization of guinea pigs. With one exception, both the Phe-His-Ser Gln and the Trp-Cys-Arg motifs were required in the peptide sequences to obtain neutralizing hyperimmune guinea pig sera, and the highest titers were obtained after immunization with 20-27 amino acids (aa) long peptides. Neither substitutions nor deletions of residues between the two motifs, nor the addition of peptide sequences representing a T-helper epitope improved the induction of neutralizing antibodies. Computer simulation modeling revealed that the Phe-315, His-316, Trp-329 and Cys-330 are likely to participate in the formation of a discontinuous epitope. Taken together, these data support the hypothesis that the well conserved motifs FHSQ (positions 315-318) and WCR (positions 329-331) of the HIV-2(SBL6669) V3 region are important targets for neutralizing antibodies, and this may have implications for the design of a future HIV-2 vaccine. PMID- 10854166 TI - Four genotypic variants of a Spodoptera exigua Nucleopolyhedrovirus (Se-SP2) are distinguishable by a hypervariable genomic region. AB - Four genotypes named SP2A, SP2B, SP2C and SP2D were obtained in vivo by infecting S. exigua larvae with limiting dilutions of the Spanish field isolate Spodoptera exigua Nucleopolyhedrovirus (Se-SP2) of SeMNPV. The cloning of variants SP2A, SP2B and SP2C took 1, 6, and 3 passages, respectively, before the DNA profiles showed all bands in equimolar concentrations, and they remained constant for at least six further passages indicating the stability of their genotypes. The SP2D variant isolation took over ten passages and it was genetically less stable. Physical maps of their genomes were constructed for the restriction enzymes BamHI, BglII, PstI, and XbaI. The region between 8-10 m.u. was highly variable and characteristic of each cloned genotype and, hence, can be used as RFLP markers for all four genotypic variants. This region, included in the PstI-MB fragment, was cloned and sequenced showing that all the Se-SP2 variants contained a homologous region (hr) with a variable number of 98 bp sequences tandemly repeated, which were used to distinguish genotypic variants from each other. The biological activity of the genotypic variants SP2A, SP2B, and SP2C when compared in terms of LD50 and LT50, were not significantly different. However, the SP2D genotypic variant was found to be significantly less infective (higher LD50). The emergence of new genotypes in the Se-SP2 field populations is discussed. PMID- 10854167 TI - Echovirus 5: infectious transcripts and complete nucleotide sequence from uncloned cDNA. AB - Echovirus 5 (EV5) may be isolated from various neurological and exanthematic diseases. To determine the relationship of EV5 to other enteroviruses and for studies of its interactions with the target cell, the complete nucleotide sequence of EV5 was determined. Three overlapping fragments, collectively representing the complete genome, were amplified with RT-PCR and sequenced. Analysis of the EV5 sequence revealed a typical enterovirus-like organization of the genome. To verify that the cDNA generated sequence was derived from infectious viruses, complete EV5 genomes were amplified in one amplicon by long distance PCR. Transfection of in vitro transcribed RNA from these amplicons into cell cultures resulted in replicating EV5. Comparison of the overall nucleotide and amino acid sequences demonstrates that EV5 can be regarded as a coxsackievirus B-like enterovirus. Variable sequences between EV5 and the well characterized coxsackievirus B3 (CVB3) are for the most part observed for amino acid residues that correspond to exposed sequences in the CVB3 capsid. This observation indicates that the reported EV5 strain recently diverged from group B coxsackieviruses. PMID- 10854168 TI - Genetic variability in envelope-associated protein genes of closely related group A strains of respiratory syncytial virus. AB - The genetic and antigenic diversity present in respiratory syncytial virus (RSV) strains may in part be explained by genetic drift similar to that which occurs with influenza virus B. To study drift in RSV strains, we sequenced the five membrane-associated genes, M, SH, G, F, and M2, from three sets of RSV isolates: one set of seven closely related isolates obtained over 5 years in St. Louis, MO, and two sets of four closely related RSV isolates from other communities. We found nucleotide-variable and conserved regions in all five genes, and the greatest diversity in the SH and G genes. We did not find clear evidence of genetic drift in the seven isolates from St. Louis for any of the five genes. Although the relationships between strains were usually maintained independent of the genes studied, for several isolates there was a dramatic shift in genetic relationships for one of the five genes. Our inability to demonstrate genetic drift and the dramatic shift in genetic relationships between some strains for some genes suggest that we need to better define the mechanisms and rate of change in this virus to accurately define phylogenetic relationships between strains. PMID- 10854169 TI - Molecular characterization of the Japanese encephalitis virus representative immunotype strain JaGAr 01. AB - We determined the full genomic sequence of the Japanese encephalitis virus JaGAr 01 strain and its predicted amino acid sequence. Nucleotide sequence comparison with ten fully sequenced JE strains shows a homology range from 89.62 to 99.49%. Amino acid sequence homologies range from 96.85 to 99.74%. Comparison of amino acid sequences shows a unique amino acid, arginine, for JaGAr 01 at position 123 of the E-protein, while the eight other strains contained serine. Secondary structure prediction by free energy minimization shows a unique structure for JaGAr 01 that includes an RNA segment that is conserved for all flaviviruses. Speculation is made about the role these results may play in the replication and antigenic characteristics of JaGAr 01. Phylogenetic analyses of the E-protein of JaGAr 01 together with 35 other JE strains showed diversity in amino acid characteristics between the prototype strains Nakayama, JaGAr 01 and Beijing-1. Phylogenetic trees computed by neighbor joining and Fitch Margoliash analysis of nucleic acid and protein sequences showed Nakayama and Beijing in one cluster different from JaGAr 01. PMID- 10854170 TI - Integrated viral genomes can be lost from adenovirus type 12-induced hamster tumor cells in a clone-specific, multistep process with retention of the oncogenic phenotype. AB - In adenovirus type 12 (Ad12)-induced tumor cells, in Ad12-transformed cells and in continuously passaged cell lines from these sources, the viral DNA is integrated in multiple copies, usually at a single chromosomal location. In different tumors or cell lines, the sites of integration of Ad12 DNA are all different. Rare exceptions exist. In most instances, the integrated viral DNA resides very stably in the host cell genomes. However, upon continuous serial passage of such cell lines, the integrated viral DNA can be destabilized and lost. In two instances, i.e. in the Ad12-induced hamster tumor cell lines H1111(1) and CLAC1, we have investigated the loss of integrated viral DNA in detail. After extended serial passage, these two cell lines seemed to be devoid of Ad12 DNA sequences, as detectable by Southern blot hybridization, but continued to induce tumors after reinjection into hamsters. Cells from these two cell lines were now recloned three times, and DNAs from cultures derived from several individual clones were reinvestigated for the presence of several parts of the viral genome by the polymerase chain reaction (PCR). Some of the clones still carried parts of the Ad12 genome. However, several clones were isolated that proved free of all parts of the viral genome, except for minute segments from the right terminus of the Ad12 genome. Apparently, the loss of integrated viral DNA from these cell lines proceeded as a continuous, gradual, multistep process whose pattern could differ from cell clone to cell clone, once destabilization had been initiated. The mechanism of destabilization is not understood. Cell populations of 2 x 10(6) to 3 x 10(7), and as low as 10(2), cells from the clones, that contained only minimal remnants from the right viral DNA terminus, were reinjected into newborn or 13-20 day-old weanling Syrian hamsters (Mesocricetus auratus). Tumors developed within 5-17 days after injection. Tumor cell clones also grew in soft agar. The injection of primary hamster skin fibroblasts never elicited tumor formation. The tumor cells induced by this reinjection proved repeatedly free of Ad12 DNA both by Southern blot hybridization and by PCR, except for those cell and tumor clones that contained small segments of the right terminal E4 region of the Ad12 genome. The tumor cells, however, retained their oncogenic phenotype. The results raise questions about the cell clone-specific excision patterns of integrated foreign DNA from the recipient genome and the possibility of a hit-and-run mechanism of adenoviral oncogenesis. PMID- 10854171 TI - The status of tumor immunology and cancer immunotherapy. Introduction. PMID- 10854172 TI - Identification of lung tumor antigens for cancer immunotherapy: immunological and molecular approaches. PMID- 10854173 TI - Approaches for immunotherapy of lymphomas. PMID- 10854174 TI - Down-regulation of MHC class I antigen presentation by HCMV; lessons for tumor immunology. PMID- 10854175 TI - Role of chaperones in antigen processing. PMID- 10854176 TI - MHC-binding peptides as immunotherapeutics for cancer. PMID- 10854177 TI - The role of CD40-CD154 interactions in the regulation of cell mediated immunity. AB - CD40 is expressed on a diverse array of cell types from the hematopoietic and non hematopoietic compartments. Within the hematopoietic compartment, CD40 is found constitutively expressed on B cells, dendritic cells (DC) and macrophages. The function of CD40 in B cells has been documented as being essential in the control of humoral immunity. In DCs and macrophages, CD40 has been shown to be important in the induction of antigen-presenting cell (APC) maturation and effector function. CD40 is also expressed on non-hematopoietic cells like keratinocytes, epithelial cells, and vascular endothelial cells and has been shown to be functionally important on these cell types. PMID- 10854178 TI - Amplification of T cell-mediated immune responses by antibody-cytokine fusion proteins. PMID- 10854179 TI - Dendritic cell/peptide cancer vaccines: clinical responsiveness and epitope spreading. PMID- 10854180 TI - Tumor vaccine strategies that employ dendritic cells and tumor lysates: experimental and clinical studies. PMID- 10854181 TI - Heat shock proteins and cancer immunotherapy. AB - Vaccination with heat shock proteins from tumor have been shown to elicit an anti tumor response. Current studies indicate that the immunogenicity of HSPs is derived from the antigenic peptides which they associate with. Mechanisms by which the HSP-peptide complexes induce an immune response and the possible role of HSPs in antigen presentation is discussed in this article. The use of HSP peptide complexes can be used as tumor vaccines for cancer immunotherapy is reviewed. PMID- 10854182 TI - Use of mild, whole body hyperthermia in cancer therapy. PMID- 10854183 TI - Cytokines and cancer immunotherapy. PMID- 10854184 TI - Immunologic monitoring of clinical trials in patients with cancer: technology versus common sense. AB - Monitoring of CTL or helper T -cell responses to tumor antigens in the course of clinical trials is an essential and necessary component of specific immunotherapy of cancer. While a variety of new immunologic or molecular technologies are now available for measuring these responses at a single-cell level, none has been standardized or validated to meet this need. The clinical investigator is thus faced with a selection of a method that is most likely to meet the desirable objective of establishing a correlation between clinical and immunologic responses to biotherapy. Selection of a relevant assay for this purpose is not a trivial task. Furthermore, few immunologic techniques can be easily adapted to reliable serial monitoring of patients on clinical protocols. The ELISPOT assay performs well in this setting, allowing for tracking of antigen specific T cells in the peripheral blood of cancer patients who receive tumor vaccines. In our hands, this assay is sensitive, accurate and cost effective. Its reliable performance in the context of clinical vaccination trials is highly likely to provide information necessary for optimizing vaccine administration to cancer patients, including timing, route of delivery, antigen concentration and use of DC or other adjuvants. PMID- 10854185 TI - Monitoring antigen-specific T cells using MHC-Ig dimers. AB - To summarize, two novel approaches are currently being examined that allow for identification of antigen-specific T cells. A biochemical approach to generating soluble multivalent MHC complexes has been to generate tetrameric MHC complexes linked to avidin. We have also generated a general approach for producing soluble divalent versions of class I and class II MHC molecules, using Ig as a molecular scaffold. The experimental system described here outlines a general approach of using multivalent high affinity ligands to study cell-cell interactions, driven by multivalent ligand-receptor interactions. Our work indicates that divalent chimeric molecules are high-avidity analogs of proteins useful in probing and selectively regulating cellular responses. PMID- 10854186 TI - Patient immune response to tumors monitored using SCID mouse models. PMID- 10854187 TI - Dendritic cells promote T-cell survival or death depending upon their maturation state and presentation of antigen. PMID- 10854188 TI - Trace minerals in human growth and development. AB - Trace mineral deficiencies may affect several biological functions in humans, including physical growth, psychomotor development and immunity. We have reviewed the mechanisms whereby several trace mineral deficiencies may affect these biological functions at different ages (fetal life, infancy, childhood and adolescence), as well as the evidence supporting this association. We describe the effects of zinc deficiency on the hormonal regulation of growth and sexual development in both humans and animal models. We provide data regarding the effects of iron deficiency on growth and psychomotor development. We mention the effects of copper, manganese, selenium and iodine deficiencies on growth and development. We conclude that iron deficiency may affect psychomotor development, but does not appear to affect growth. Zinc deficiency may cause growth retardation and psychomotor delay. PMID- 10854189 TI - Oral health in children and adolescents with IDDM--a review. AB - Children with insulin-dependent diabetes mellitus have a lower salivary flow rate, pH and buffer capacity, but a higher glucose content and peroxidase, IgA, magnesium and calcium concentration, in comparison with healthy children. Nevertheless the incidence of caries is lower than normal in diabetic children with good metabolic control. Periodontal disease usually starts at puberty as mild gingivitis with bleeding and gingival recession, and it may develop into severe periodontitis, especially in children with poor control of diabetes. Microangiopathy, impaired immune response, different bacterial microflora and collagen metabolism are involved in the pathogenesis of diabetic periodontal disease. The gingival flora is mostly composed of Gram-negative, anaerobic bacteria, while collagen has a lower solubility and is atrophic and inadequate to support the occlusion forces. For these reasons, prevention of periodontitis is important in diabetic children; they should receive oral hygiene instruction and visit a dentist at least twice a year. PMID- 10854204 TI - Genetic progression in the pancreatic ducts. PMID- 10854205 TI - Genetic imbalances in precursor lesions of endometrial cancer detected by comparative genomic hybridization. AB - Endometrial hyperplasia is regarded as a precursor lesion of endometrioid adenocarcinomas of the endometrium. The genetic events involved in the multistep process from normal endometrial glandular tissue to invasive endometrial carcinomas are primarily unknown. We chose endometrial hyperplasia as a model for identifying chromosomal aberrations occurring during carcinogenesis. Comparative genomic hybridization (CGH) was performed on 47 formalin-fixed, paraffin-embedded specimens of endometrial hyperplasia using the microdissection technique to increase the number of tumor cells in the samples and reduce contamination from normal cells. CGH analysis revealed that 24 out of 47 (51%) samples had detectable chromosomal imbalances, whereas 23 (49%) were in a genetically balanced state. The incidence of aberrant CGH profiles tended to parallel dysplasia grade, ranging from 22% aberrant profiles in simple hyperplasia to 67% in complex hyperplasia with atypia. The most frequent imbalances were 1p, 16p, and 20q underrepresentations and 4q overrepresentations. Copy number changes in 1p were more frequent in atypical complex hyperplasia than in complex lesions without atypical cells or simple lesions (42% versus 20% and 0%). Our results show that endometrial hyperplasia reveals recurrent chromosomal imbalances which tend to increase with the presence of atypical cells. The most frequent aberrations in endometrial cancer, 1q and 8q overrepresentations, are not present or are rare in its precursor lesions. This analysis provides evidence that tumorigenesis proceeds through the accumulation of a series of genetic alterations and suggests a stepwise mode of tumorigenesis. PMID- 10854206 TI - Association of active extracellular signal-regulated protein kinase with paired helical filaments of inclusion-body myositis muscle suggests its role in inclusion-body myositis tau phosphorylation. AB - The possible role of extracellular signal-regulated kinase (ERK) in the pathogenesis of inclusion-body myositis (IBM) was investigated by immunostaining the active phosphorylated form of ERK in muscle biopsies of six IBM and 14 control patients. Between 80% and 90% of IBM vacuolated muscle fibers contained well-defined ERK-immunoreactive inclusions, which were co-localized by light microscopy, with phosphorylated tau in 70 to 80% of those fibers. Immunoelectronmicroscopy colocalized ERK to small amorphous tufts adjacent to the muscle fiber paired-helical filaments. Strong ERK immunoreactivity was also present at the postsynaptic domain of all human neuromuscular junctions. Our study suggests 1) that ERK, a signal transducer, might play a role in IBM pathogenesis, including participation in the pathological phosphorylation of IBM tau; and 2) that signal transduction abnormalities may be a component of the IBM pathogenic cascade. Our novel immunolocalization of ERK at the postsynaptic domain of human neuromuscular junctions supports a role in transcription of junctional-protein genes. The ERK localized in nonjunctional regions of IBM fibers may underlie the known pathological up-regulation of junctional proteins there. PMID- 10854207 TI - Nuclear localization of catechol-O-methyltransferase in neoplastic and nonneoplastic mammary epithelial cells. AB - Catechol-O-methyltransferase (COMT) plays both a regulatory and protective role in catechol homeostasis. It contributes to the regulation of tissue levels of catecholamines and catecholestrogens (CEs) and, by blocking oxidative metabolism of catechols, prevents endogenous and exogenous catechols from becoming a source of potentially mutagenic electrophiles. Evidence implicating CEs in carcinogenesis, in particular in the hamster kidney model of estrogen-induced cancer, has focused attention on the protective role of COMT in estrogen target tissues. We have previously reported that treating hamsters with estrogens causes translocation of COMT to nuclei of epithelial cells in the renal cortex, the site of CE biosynthesis and where the cancers arise. This finding suggested that nuclear COMT may be a marker of a threat to the genome by catechols, including CEs. It is postulated that CEs play a role in the genesis of breast cancer by contributing to a state of chronic oxidative stress that is presumed to underlie the high incidence of this disease in the United States. Therefore, here we used immunocytochemistry to re-examine human breast parenchyma for nuclear COMT. In addition to confirming previous reports of cytoplasmic COMT in mammary epithelial cells, we identified nuclear COMT in foci of mammary epithelial cells in histologically normal breast tissue of virtually all control (macromastia) and cancer patients and in breast cancer cells. There was no correlation between tissue histology and the numbers of cells with nuclear COMT, the size of foci containing such cells, or intensity of nuclear COMT immunostaining. The focal nature of the phenomenon suggests that nuclear COMT does not serve a housekeeping function but that it reflects a protective response to an increased local catechol load, presumably of CEs and, as such, that it may be a characteristic of the population of women studied who share the same major risk factor for developing breast cancer, that of living in the industrialized West. PMID- 10854208 TI - Distribution and characterization of GFP(+) donor hematogenous cells in Twitcher mice after bone marrow transplantation. AB - The twitcher mouse is a murine model of globoid cell leukodystropy, a genetic demyelinating disease caused by a mutation of the galactosylceramidase gene. Demyelination of the central nervous system commences around 20 postnatal days. Using GFP-transgenic mice as donors, the distribution of hematogenous cells after bone marrow transplantation was investigated in the twitcher mice. Bone marrow transplantation was carried out at 8 postnatal days. In twitcher chimeric mice examined before 30 postnatal days, numerous GFP(+) cells were detected in spleen and peripheral nerve but only a few were detected in the liver, lung, and spinal white matter. In contrast, at 35 to 40 postnatal days when demyelination is evident, many GFP(+) cells with ameboid form were detected in the white matter of the spinal cord, brainstem, and cerebrum. Approximately half of these GFP(+) cells were co-labeled with Mac-1. In twitcher chimeric mice examined after 100 postnatal days, the majority of GFP/Mac-1 double-positive cells displayed the morphological features of ramified microglia with fine delicate processes and was distributed diffusely in both gray and white matter. These results suggest that a significant number of donor hematogenous cells are able to infiltrate into the brain parenchyma, repositioning themselves into areas previously occupied by microglia, and to ameliorate lethality. PMID- 10854209 TI - Detection of gene amplification in archival breast cancer specimens by laser assisted microdissection and quantitative real-time polymerase chain reaction. AB - Gene amplification is one of the most important mechanisms leading to deregulated gene expression in cancer. The exact quantitative detection of this frequent genomic alteration in solid tumors is often hampered by an admixture of nonneoplastic bystander and stroma cells. To overcome this obstacle and to develop an objective quantitative method we have combined laser-assisted microdissection of tumor cells with the novel 5'-exonuclease-based real-time polymerase chain reaction (PCR) assay. The latter method enables the highly reproducible exact quantification of minute amounts of nucleic acids. As a model system amplification of c-erbB2/Her-2/neu gene and the adjacent topoisomerase IIalpha gene was determined in paraffin-embedded breast cancer specimens (n = 23) after immunohistochemical labeling and laser-based microdissection of tumor cells. The high sensitivity of real-time PCR enabled the reliable and objective detection of low-level amplifications in as few as 50 cells from archival tissue sections. Low-level amplifications were shown to escape from detection unless tumor cells were isolated by microdissection. In selected cases intratumor heterogeneity was demonstrated using areas of approximately 50 to 100 cells. This novel approach combining immunohistochemistry, laser microdissection, and quantitative kinetic PCR allows morphology-guided studies in archival tissue specimens and will enable the exact quantification of gene copy numbers in even small and precancerous lesions. PMID- 10854210 TI - Disruption of the plasminogen gene in mice abolishes wound healing after myocardial infarction. AB - The plasminogen system plays an important role in the proteolytic degradation of extracellular matrices during wound healing. In the present study we investigated the impact of the plasminogen system on cardiac wound healing and function after myocardial infarction. Myocardial infarction was induced in plasminogen-deficient mice (Plg-/-) and in wild-type controls (Plg+/+). Structural analysis 1, 2, and 5 weeks after infarction revealed that infarct healing was virtually abolished in Plg-/- mice, indicating that the plasminogen system is required for the repair process of the heart after infarction. In the absence of plasminogen, inflammatory cells did not migrate into the infarcted myocardium. Necrotic cardiomyocytes were not removed and the formation of granulation tissue and fibrous tissue did not occur. In these non-healing infarcted hearts, LV dilatation was not altered. In addition, gelatinolytic activity of MMP-2 and MMP 9 was depressed in the Plg-/- infarcted hearts, suggesting that the plasmin effect on infarct healing may be mediated by MMPs. Surprisingly, cardiac function was only attenuated to a rather small extent in the Plg-/- infarcted mice when compared to the wild-types. This study provides direct prove that plasmin mediated proteolysis plays a central role in cardiac wound healing after myocardial infarction in mice. PMID- 10854211 TI - Increased expression and activation of stress-activated protein kinases/c-Jun NH(2)-terminal protein kinases in atherosclerotic lesions coincide with p53. AB - Hyperlipidemia alters gene expression of arterial endothelial and smooth muscle cells (SMCs) and induces atherosclerotic lesions, in which cell proliferation and apoptosis co-exist. The signal transduction pathways that mediate these responses in the vessel wall in vivo have yet to be identified. Stress-activated protein kinases (SAPKs) or c-Jun NH(2)-terminal protein kinases (JNKs) are thought to be crucial in transmitting transmembrane signals required for cell differentiation and apoptosis in vitro. In the present study, we investigated the localization and activity of SAPK/JNK in atherosclerotic lesions of cholesterol-fed rabbits. Immunofluorescence analysis revealed abundant and heterogeneous distribution of pan-SAPK/JNK and phosphorylated SAPK/JNK, which were mainly localized in cell nuclei of the lesional cap and basal regions. Double staining of the lesions demonstrated that a portion of alpha-actin(+) SMCs and RAM11(+) macrophages contained abundant phosphorylated SAPK/JNK proteins. SAPK/JNK protein levels in protein extracts from atherosclerotic lesions were two- to threefold higher than the vessels of chow-fed rabbits. SAPK/JNK activities were elevated three- to fivefold higher than the normal vessels. Interestingly, increased SAPK/JNK in lesions was co-localized or coincided with high levels of transcription factor p53 as identified by double labeling and immunoprecipitation. Abundant pro apoptotic protein BAX and BCL-X(S) were also observed. Furthermore, low-density lipoprotein (LDL) and oxidized LDL stimulated SAPK/JNK activation in cultured SMCs in a time- and dose-dependent manner. LDL also induced SAPK/JNK activation in vascular SMCs derived from LDL-receptor-deficient Watanabe rabbits, indicating a LDL-receptor-independent process. Thus, SAPK/JNK persistently hyperexpressed and activated in lesions may play a key role in mediating cell differentiation and apoptosis during the development of atherosclerosis via activation of transcription factor p53. PMID- 10854212 TI - Gene expression profiling in an in vitro model of angiogenesis. AB - In the present study we have used a novel, comprehensive mRNA profiling technique (GeneCalling) for determining differential gene expression profiles of human endothelial cells undergoing differentiation into tubelike structures. One hundred fifteen cDNA fragments were identified and shown to represent 90 distinct genes. Although some of the genes identified have previously been implicated in angiogenesis, potential roles for many new genes, including OX-40, white protein homolog, KIAA0188, a homolog of angiopoietin-2, ADAMTS-4 (aggrecanase-1), and stanniocalcin were revealed. Support for the biological significance was confirmed by the abrogation of the changes in the expression of angiogenesis inhibitors and in situ hybridization studies. This study has significantly extends the molecular fingerprint of the changes in gene expression that occur during endothelial differentiation and provides new insights into the potential role of a number of new molecules in angiogenesis. PMID- 10854213 TI - Glomerular expression of type IV collagen chains in normal and X-linked Alport syndrome kidneys. AB - Alport syndrome is an inherited nephropathy characterized by alterations of the glomerular basement membrane because of mutations in type IV collagen genes. COL4A5 mutations, causing X-linked Alport syndrome, frequently result in the loss of the alpha5 chains of type IV collagen in basement membranes. This is associated with the absence of the alpha3(IV) and alpha4(IV) chains and increased amounts of alpha1(IV) and alpha2(IV) in glomerular basement membranes. The mechanisms resulting in such a configuration are still controversial and are of fundamental importance for understanding the pathology of the disease and for considering gene therapy. In this article we studied, for the first time, type IV collagen expression in kidneys from X-linked Alport syndrome patients, using in situ hybridization and immunohistochemistry. We show that, independent of the type of mutation and of the level of COL4A5 transcription, both COL4A3 and COL4A4 genes are actively transcribed in podocytes. Moreover, using immunofluorescence amplification, we were able to demonstrate that the alpha3 chain of type IV collagen was present in the podocytes of all patients. Finally, the alpha1(IV) chain, which accumulates within glomerular basement membranes, was found to be synthesized by mesangial/endothelial cells. These results strongly suggest that, contrary to what has been found in dogs affected with X-linked Alport syndrome, there is no transcriptional co-regulation of COL4A3, COL4A4, and COL4A5 genes in humans, and that the absence of alpha3(IV) to alpha5(IV) in glomerular basement membranes in the patients results from events downstream of transcription, RNA processing, and protein synthesis. PMID- 10854214 TI - Apolipoprotein AI and transthyretin as components of amyloid fibrils in a kindred with apoAI Leu178His amyloidosis. AB - We found a new C-terminal amyloidogenic variant of apolipoprotein AI (apoAI), Leu178His in a French kindred, associated with cardiac and larynx amyloidosis and skin lesions with onset during the fourth decade. This single-point mutation in exon 4 of the apoAI gene was detected by DNA sequencing of polymerase chain reaction amplified material and restriction fragment length polymorphism analysis in two siblings. Blood, larynx, and skin biopsies were available from one sibling. Anti-apoAI immunoblotting of isoelectric focusing of plasma showed a +1 alteration in the charge of the protein. Extraction of fibrils from the skin biopsy revealed both full-length and N-terminal fragments of apoAI and transthyretin (TTR). ApoAI and TTR co-localized in amyloid deposits as demonstrated by immunohistochemistry. The present report, together with the first recently described C-terminal amyloidogenic variant of apoAI, Arg173Pro, shows that amyloidogenicity of apoAI is not a feature exclusive to N-terminal variants. The most striking characteristic of amyloid fibrils in Leu178His is that wild type TTR is co-localized with apoAI in the fibrils. We have previously determined that a fraction of plasma TTR circulates in plasma bound to high-density lipoprotein and that this interaction occurs through binding to apoAI. Therefore we hypothesize that nonmutated TTR might influence deposition of apoAI as amyloid. PMID- 10854215 TI - 4'-Iodo-4'-deoxydoxorubicin disrupts the fibrillar structure of transthyretin amyloid. AB - Transthyretin (TTR) is a tetrameric protein synthesized mainly by the liver and the choroid plexus, from where it is secreted into the plasma and the cerebrospinal fluid, respectively. Some forms of polyneuropathy, vitreopathy, and cardiomyopathy are caused by the deposition of normal and/or mutant TTR molecules in the form of amyloid fibrils. Familial amyloidotic polyneuropathy is the most common form of TTR amyloidosis related to the V30M variant. It is still unclear the process by which soluble proteins deposit as amyloid. The treatment of amyloid-related disorders might attempt the stabilization of the soluble protein precursor to retard or inhibit its deposition as amyloid; or aim at the resorption of the deposited amyloid. The anthracycline 4'-iodo-4' deoxydoxorubicin (I-DOX) has been shown to reduce the amyloid load in immunoglobulin light-chain amyloidosis. We investigated 1) whether I-DOX has affinity for TTR amyloid in tissues, 2) determined the I-DOX binding constants to TTR synthetic fibrils, and 3) determined the nature of the effect of I-DOX on TTR fibrils. We report that 1) I-DOX co-localizes with amyloid deposits in tissue sections of patients with familial amyloidotic polyneuropathy; 2) I-DOX strongly interacts with TTR amyloid fibrils and presents two binding sites with k(d) of 1.5 x 10(-11) mol/L and 5.6 x 10(-10) mol/L, respectively; and 3) I-DOX disrupts the fibrillar structure of TTR amyloid into amorphous material, as assessed by electron microscopy but does not solubilize the fibrils as confirmed by filter assays. These data support the hypothesis that I-DOX and less toxic derivatives can prove efficient in the treatment of TTR-related amyloidosis. PMID- 10854216 TI - Expression of the integrin alpha8beta1 during pulmonary and hepatic fibrosis. AB - The fibrotic response after diverse forms of injury is characterized by the accumulation of extracellular matrix proteins, proliferation of myofibroblast like cells, and organ contraction. Myofibroblasts are key effector cells in the development of the fibrotic response. They contribute to fibrosis through both increased cell number (proliferation) and enhanced matrix synthesis. Integrins, a class of cell adhesion molecules, are mediators of cell-extracellular matrix protein interactions that are important in the proliferative and migratory response of cells to matrix proteins. We have previously cloned the human integrin subunit alpha8, documented its high expression in lung tissue, and established it as a receptor for the matrix proteins fibronectin, vitronectin, and tenascin. We now demonstrate that alveolar interstitial cells are the primary cell type expressing alpha8beta1 in the lung parenchyma. Expression of alpha8beta1 is concentrated primarily along the thinned extensions of cells and at the tips of filopodia. Because of its unique distribution in alveolar interstitial cells, we hypothesized that it may play a role in the fibrotic response after injury. In bleomycin-induced pulmonary fibrosis, there is increased expression of alpha8beta1 by interstitial fibroblasts, the majority of which coexpress alpha smooth muscle actin, a marker of tissue myofibroblasts. To establish a more general role for alpha8beta1 during organ fibrosis, we further examined its expression in two rat models of liver fibrosis. During hepatic injury due to either carbon tetrachloride injury or bile duct ligation, we demonstrate de novo expression of alpha8beta1 in activated hepatic stellate cells, the myofibroblast equivalent in liver. Taken together, the data localize alpha8beta1 to myofibroblast-like cells during wound healing and suggest that signal transduction through the alpha8beta1 integrin may contribute to the fibrotic response of organs to injury. PMID- 10854217 TI - In vivo expression of the novel CXC chemokine BRAK in normal and cancerous human tissue. AB - Using differential display, we cloned a gene with reduced expression in short term explants of head and neck squamous cell carcinoma (HNSCC) tumors compared to cultured normal oral epithelial cells. The differentially expressed gene was identical to the recently cloned CXC chemokine BRAK, which is ubiquitously expressed in normal tissue extracts but is absent from many tumor cell lines in vitro. To define the cell populations expressing BRAK in vivo, in situ mRNA hybridization was performed on normal and cancerous tissues from six different histological sites. The predominant normal cell type constitutively expressing BRAK in vivo was squamous epithelium. Expression in tumors was heterogeneous, with the majority of HNSCCs and some cervical squamous cell carcinomas (SCCs) showing loss of BRAK mRNA. Although absent in unstimulated peripheral blood mononuclear cells, high levels of BRAK were consistently found in infiltrating inflammatory cells (with lymphocyte morphology) in nearly all cancers examined. Furthermore, BRAK expression was demonstrated in B cells and monocytes, after stimulation of peripheral blood mononuclear cells with lipopolysaccharide. This study demonstrates for the first time up-regulation of BRAK mRNA by inflammatory cells in the tumor microenvironment and lost expression from certain cancers in vivo. The data suggest that BRAK may have a role in host-tumor interactions. PMID- 10854218 TI - Contrasting effects of CCR5 and CCR2 deficiency in the pulmonary inflammatory response to influenza A virus. AB - The immune response to influenza A virus is characterized by an influx of both macrophages and T lymphocytes into the lungs of the infected host, accompanied by induced expression of a number of CC chemokines. CC chemokine receptors CCR5 and CCR2 are both expressed on activated macrophages and T cells. We examined how the absence of these chemokine receptors would affect pulmonary chemokine expression and induced leukocyte recruitment by infecting CCR5-deficient mice and CCR2 deficient mice with a mouse-adapted strain of influenza A virus. CCR5(-/-) mice displayed increased mortality rates associated with acute, severe pneumonitis, whereas CCR2(-/-) mice were protected from the early pathological manifestations of influenza because of defective macrophage recruitment. This delay in macrophage accumulation in CCR2(-/-) mice caused a subsequent delay in T cell migration, which correlated with high pulmonary viral titers at early time points. Infected CCR5(-/-) mice and CCR2(-/-) mice both exhibited increased expression of the gene for MCP-1, the major ligand for CCR2(-/-) and a key regulator of induced macrophage migration. These studies illustrate the very different roles that CCR5 and CCR2 play in the macrophage response to influenza infection and demonstrate how defects in macrophage recruitment affect the normal development of the cell-mediated immune response. PMID- 10854219 TI - Induction of HHV-8 lytic cycle replication by inflammatory cytokines produced by HIV-1-infected T cells. AB - Human herpesvirus 8 (HHV-8) is a gamma2-herpesvirus consistently identified in Kaposi's sarcoma (KS), primary effusion lymphoma, and multicentric Castleman's disease. Although HHV-8 infection appears to be necessary, it may not be sufficient for development of KS without the involvement of other cofactors. One potentially important cofactor is HIV-1. HIV-1-infected cells produce HIV-1 related proteins and cytokines, both of which have been shown to promote growth of KS cells in vitro. Though HIV-1 is not absolutely necessary for KS development, KS is the most frequent neoplasm in AIDS patients, and AIDS-KS is recognized as a particularly aggressive form of the disease. To determine whether HIV-1 could participate in the pathogenesis of KS by modulating HHV-8 replication (rather than by inducing immunodeficiency), HIV-1-infected T cells were cocultured with the HHV-8-infected cell line, BCBL-1. The results demonstrate soluble factors produced by or in response to HIV-1-infected T cells induced HHV 8 replication, as determined by production of lytic phase mRNA transcripts, viral proteins, and detection of progeny virions. By focusing on cytokines produced in the coculture system, several cytokines known to be important in growth and proliferation of KS cells in vitro, particularly Oncostatin M, hepatocyte growth factor/scatter factor, and interferon-gamma, were found to induce HHV-8 lytic replication when added individually to BCBL-1 cells. These results suggest specific cytokines can play an important role in the initiation and progression of KS through reactivation of HHV-8. Thus, HIV-1 may participate more directly than previously recognized in KS by promoting HHV-8 replication and, hence, increasing local HHV-8 viral load. PMID- 10854220 TI - Treatment-induced decline of human immunodeficiency virus-1 p24 and HIV-1 RNA in lymphoid tissue of patients with early human immunodeficiency virus-1 infection. AB - We report detailed quantitative analysis of human immunodeficiency virus-1 (HIV 1) p24 and HIV-1 RNA in tonsil biopsies from 13 patients with early, asymptomatic HIV infection before and during combination antiretroviral therapy. Using fluorescent microscopy in conjunction with reverse transcriptase-polymerase chain reaction of frozen tissue sections, we show that plasma and tissue viral loads decreased by approximately 3 logs during the 1-year treatment period, with good correlation between the HIV-1 p24 and HIV-1 RNA response in tissue. The decrease of tissue viral load was delayed compared to plasma viral load, possibly explained by the observation that the amount of follicular dendritic cell associated virus correlated best with the area under the curve of plasma HIV-1 RNA throughout the last 12 weeks. Before and during treatment, the relative proportions of HIV-1 on follicular dendritic cells and within mononuclear cells remained constant, suggesting similar decay characteristics in these two lymphoid tissue compartments. However, viral p24 or RNA remained almost always detectable in tissue despite full suppression of HIV-1 RNA in plasma, and increased even after short-term rebounds in plasma viral load. Thus, full and sustained suppression of viral replication was required to efficiently decrease viral load in lymphoid tissue, but complete abolition of residual viral replication was not achieved. PMID- 10854221 TI - INK4a/ARF locus alterations in human non-Hodgkin's lymphomas mainly occur in tumors with wild-type p53 gene. AB - INK4a/ARF locus codes for two different proteins, p16(INK4a) and p14(ARF), involved in cell cycle regulation. p14(ARF) is considered an upstream regulator of p53 function. To determine the role of these genes in the pathogenesis of human non-Hodgkin's lymphomas we have analyzed exon 1beta, 1alpha, and 2 of the INK4a/ARF locus and p53 gene aberrations in 97 tumors previously characterized for p16(INK4a) alterations. p53 alterations were detected in four of 51 (8%) indolent lymphomas but in 15 of 46 (33%) aggressive tumors. Inactivation of p14(ARF) was always associated with p16(INK4a) alterations. Exon 1beta was concomitantly deleted with exon 1alpha and 2 in eight tumors. One additional lymphoblastic lymphoma showed deletion of exon 1alpha and 2 but retained exon 1beta. No mutations were detected in exon 1alpha and 1beta in any case. Two of the three mutations detected in exon 2 caused a nonsense mutation in the p16(INK4a) reading frame and a missense mutation in the ARF reading frame involving the nucleolar transport domain of the protein. The third mutation was a missense mutation in the p16(INK4a) reading frame, but it was outside the coding region of p14(ARF). Aggressive lymphomas with p14(ARF) inactivation and p53 wild type showed a significantly lower p53 protein expression than tumors with no alteration in any of these genes. In this series of tumors, inactivation of the INK4a/ARF locus mainly occurred in tumors with a wild-type p53 gene because only two lymphomas showed simultaneous aberrations in these genes. Tumors with concomitant alterations of p16(INK4a) and p14(ARF)/p53 genes seem to exhibit a worse clinical behavior than lymphomas with no alterations or isolated inactivation of any of these genes. These findings indicate that p14(ARF) genetic alterations occur in a subset of aggressive NHLs, but they are always associated with p16(INK4a) aberrations. Concomitant disruption of p16(INK4a) and p14(ARF)/p53 regulatory pathways may have a cooperative effect in the progression of these tumors. PMID- 10854222 TI - Somatic mutations of the APC gene in primary breast cancers. AB - APC gene mutations play an important role in the initiation step of colorectal carcinogenesis in both familial adenomatous polyposis (FAP) patients and non-FAP patients. Although the APC gene is expressed in most tissues, including the lung, liver, kidney, and mammary gland, its somatic mutations have rarely been found in primary tumors affecting these organs. We have developed a sensitive yeast-based assay for screening almost the entire coding region of the APC gene. By this method, we have been able to detect somatic mutations of the APC gene in 57% of colorectal cancers and none in non-small cell lung cancers. Interestingly, the assay detected somatic APC gene mutations in 18% of breast cancers, in which APC gene mutation was previously considered rare. In the breast cancers, most of the APC mutations were distributed outside the mutation cluster region that has been advocated for colorectal cancers. We also noted a difference in the mutation pattern of the APC between colorectal and breast cancers. In colorectal cancers, all base substitutions were observed at C residues (5 of 5), whereas in breast cancers the majority of them were found at G residues (4 of 5). Furthermore, APC mutations were observed at a significantly high frequency in advanced stages of primary breast cancers (TNM classification, P < 0.05; T category, P < 0.01). Our data suggest that the etiology of the APC mutations and their biological role in carcinogenesis may differ between colorectal and breast cancers. PMID- 10854223 TI - Human gastric cancer kinase profile and prognostic significance of MKK4 kinase. AB - Alterations of protein tyrosine kinase are often associated with uncontrolled cell growth and tumor progression. Knowledge of the overall expression pattern of tyrosine kinases should prove beneficial in understanding the signaling pathways involved in gastric cancer oncogenesis and in providing possible biomarkers for gastric cancer progression. To establish a general tyrosine-kinase expression profile, degenerated polymerase chain reaction primers designed from the consensus catalytic kinase motifs were used to amplify protein tyrosine kinase molecules from gastric cancer tissues. We observed more than 50 tyrosine and serine/threonine kinases from matching pairs of gastric cancer tissue and normal mucosa. Based on this new kinase profile information, we selected the MKK4 gene for further immunohistochemical studies. Statistical analysis of MKK4 protein expression and clinicopathological features indicated that MKK4 kinase expression could serve as a significant prognostic factor for relapse-free survival and for overall survival. We demonstrated a simple and sensitive method for establishing protein tyrosine-kinase expression profiles of human gastric cancer tissues as well as for discovering novel and useful clinical biomarkers from such kinase expression profiles. PMID- 10854224 TI - Proliferation and differentiation of fetal liver epithelial progenitor cells after transplantation into adult rat liver. AB - To identify cells that have the ability to proliferate and differentiate into all epithelial components of the liver lobule, we isolated fetal liver epithelial cells (FLEC) from ED 14 Fischer (F) 344 rats and transplanted these cells in conjunction with two-thirds partial hepatectomy into the liver of normal and retrorsine (Rs) treated syngeneic dipeptidyl peptidase IV mutant (DPPIV(-)) F344 rats. Using dual label immunohistochemistry/in situ hybridization, three subpopulations of FLEC were identified: cells expressing both alpha-fetoprotein (AFP) and albumin, but not CK-19; cells expressing CK-19, but not AFP or albumin, and cells expressing AFP, albumin, and cytokeratins-19 (CK-19). Proliferation, differentiation, and expansion of transplanted FLEC differed significantly in the two models. In normal liver, 1 to 2 weeks after transplantation, mainly cells with a single phenotype, hepatocytic (expressing AFP and albumin) or bile ductular (expressing only CK-19), had proliferated. In Rs-treated rats, in which the proliferative capacity of endogenous hepatocytes is impaired, transplanted cells showed mainly a dual phenotype (expressing both AFP/albumin and CK-19). One month after transplantation, DPPIV(+) FLEC engrafted into the parenchyma exhibited an hepatocytic phenotype and generated new hepatic cord structures. FLEC, localized in the vicinity of bile ducts, exhibited a biliary epithelial phenotype and formed new bile duct structures or were incorporated into pre existing bile ducts. In the absence of a proliferative stimulus, ED 14 FLEC did not proliferate or differentiate. Our results demonstrate that 14-day fetal liver contains lineage committed (unipotential) and uncommitted (bipotential) progenitor cells exerting different repopulating capacities, which are affected by the proliferative status of the recipient liver and the host site within the liver where the transplanted cells become engrafted. These findings have important implications in future studies directed toward liver repopulation and ex vivo gene therapy. PMID- 10854225 TI - The in vitro differentiating capacity of nonparenchymal epithelial cells derived from adult porcine livers. AB - Specific nonparenchymal epithelial cell (NPEC) clusters derived from normal adult porcine livers demonstrate a characteristic developmental pattern in the presence of other types of nonparenchymal cells in vitro. This pattern includes scattering, colonial growth, and an emergence of duct-like structures (DLSs) in the colonies. It has been confirmed that 96% of the scattered cell clusters in these cultures develop into colonies containing DLSs. In this study, we examine the differentiation of NPEC clusters using the scattered formation as a marker of the DLS-emerged colonies. We report that the NPECs expressed albumin, alpha fetoprotein, transferrin, cytokeratin (CK) 18, CK7, and c-met, but not alpha-1 antitrypsin (AAT), at the scattering stage. In addition, at the same stage, NPECs expressed oval-cell-related markers such as OV6, but not biliary epithelial cell (BEC) markers such as gamma-glutamyltransferase, CK19, and CK14. At the DLS emerging stage, hepatocyte markers, including AAT, were detectable in the cells either at the periphery of colonies or in the cells surrounded by the DLSs. On the other hand, the cells constituting DLSs expressed BEC markers, suggesting a bile duct nature of the DLSs. Furthermore, the cells in the colonies possessed an ultrastructural appearance of differentiated hepatocytes and BECs. These results suggest that certain NPECs are bipotent, and that, in culture, they mimic hepatoblast development in vivo. PMID- 10854226 TI - Depletion of hepatic glutathione prevents death receptor-dependent apoptotic and necrotic liver injury in mice. AB - The activation of the death receptors, tumor necrosis factor-receptor-1 (TNF-R1) or CD95, is a hallmark of inflammatory or viral liver disease. In different murine in vivo models, we found that livers depleted of gamma-glutamyl-cysteinyl glycine (GSH) by endogenous enzymatic conjugation after phorone treatment were resistant against death receptor-elicited injury as assessed by transaminase release and histopathology. In apoptotic models initiated by engagement of CD95, or by injection of TNF or lipopolysaccharide into galactosamine-sensitized mice, hepatic caspase-3-like proteases were not activated in the GSH-depleted state. Under GSH depletion, also caspase-independent, TNF-R1-mediated injury (high-dose actinomycin D or alpha-amanitin), as well as necrotic hepatotoxicity (high-dose lipopolysaccharide) were entirely blocked. In the T-cell-dependent model of concanavalin A-induced hepatotoxicity, GSH depletion resulted in a suppression of interferon-gamma release, delay of systemic TNF release, hepatic nuclear factor kappaB activation, and an abrogation of sinusoidal endothelial cell detachment as assessed by electron microscopy. When GSH depletion was initiated 3 hours after concanavalin A injection, ie, after the peak of early pro-inflammatory cytokines, livers were still protected. We conclude that sufficient hepatic GSH levels are a prerequisite for the execution of death receptor-mediated hepatocyte demise. PMID- 10854227 TI - The endogenous adjuvant squalene can induce a chronic T-cell-mediated arthritis in rats. AB - Squalene is a cholesterol precursor, which stimulates the immune system nonspecifically. We demonstrate that one intradermal injection of this adjuvant lipid can induce joint-specific inflammation in arthritis-prone DA rats. Histopathological and immunohistochemical analyses revealed erosion of bone and cartilage, and that development of polyarthritis coincided with infiltration of alphabeta(+) T cells. Depletion of these cells with anti-alphabeta TcR monoclonal antibody (R73) resulted in complete recovery, whereas anti-CD8 and anti gammadelta TcR injections were ineffective. The apparent dependence on CD4(+) T cells suggested a role for genes within the major histocompatibility complex (MHC), and this was concluded from comparative studies of MHC congenic rat strains, in which DA.1H rats were less susceptible than DA rats. Furthermore, LEW.1AV1 and PVG.1AV1 rats with MHC identical to DA rats were arthritis resistant, demonstrating that non-MHC genes also determine susceptibility. Some of these genetic influences could be linked to previously described arthritis susceptibility loci in an F2 intercross between DA and LEW.1AV1 rats (ie, Cia3, Oia2 and Cia5). Interestingly, some F2 hybrid rats developed chronic arthritis, a phenotype not apparent in the parental inbred strains. Our demonstration that an autoadjuvant can trigger chronic, immune-mediated joint-specific inflammation may give clues to the pathogenesis of rheumatoid arthritis, and it raises new questions concerning the role of endogenous molecules with adjuvant properties in chronic inflammatory diseases. PMID- 10854228 TI - Asebia-2J (Scd1(ab2J)): a new allele and a model for scarring alopecia. AB - A spontaneous, autosomal, recessive mouse mutation exhibiting mild scaly skin, progressive scarring alopecia, slightly runted growth, and photophobia arose at The Jackson Laboratory in 1993 in the inbred mouse strain DBA/1LacJ. Because this mutant mouse showed genetic, anatomical, and laboratory similarities to the asebia mutation, crosses were done between the new mutant and mice carrying the asebia-J allele. Because the F1 offspring were affected, indicating the two mutants were allelic, the new mutation was named asebia-2J. Careful histological analysis of skin development of mice homozygous and heterozygous for either asebia-J or asebia-2J revealed that both types of mutant mice are very similar regardless of their background. Notable histopathological features of mice homozygous for either allele included extreme sebaceous gland hypoplasia, abnormally long anagen follicles, retained inner root sheath, hair fiber perforation of the anagen follicle base, and progressive follicular replacement by scarring. In this article we present a new pathogenetic hypothesis based on the importance of the sebaceous gland in hair fiber sheath dissociation: in the absence of a functional sebaceous gland the hair follicle is destroyed. The cutaneous pathology of this mutant mouse underscores the importance of the sebaceous gland to follicular biology and presents an animal model for studying the human scarring alopecias, which characteristically begin with sebaceous gland ablation. PMID- 10854229 TI - Hypoxia-inducible angiopoietin-2 expression is mimicked by iodonium compounds and occurs in the rat brain and skin in response to systemic hypoxia and tissue ischemia. AB - Angiopoietins are ligands for the endothelial cell tyrosine kinase receptor Tie 2. Ang-1, the major physiological activator of Tie-2, promotes blood vessel maturation and stability. Ang-2 counteracts this effect by competitively inhibiting the binding of Ang-1 to Tie-2. Using a combined RNase protection/semiquantitative reverse transcriptase-polymerase chain reaction approach, we demonstrate that hypoxia up-regulates Ang-2 mRNA levels by up to 3.3 fold in two human endothelial cell lines. In bovine microvascular endothelial (BME) cells, the flavoprotein oxidoreductase inhibitor diphenylene iodonium (DPI) and the related compound iodonium diphenyl mimic induction of Ang-2 but not vascular endothelial growth factor (VEGF) by hypoxia; in combination with hypoxia, DPI further increases Ang-2 expression but has no effect on the induction of VEGF by hypoxia. Neither Ang-2 or VEGF was increased by cyanide or rotenone, suggesting that failure in mitochondrial electron transport is not involved in the oxygen-sensing system that controls their expression. In ischemic rat dorsal skin flaps or in the brain of rats maintained for 12 hours under conditions of hypoxia, Ang-2 mRNA was up-regulated 7.5- or 17.6- fold, respectively. VEGF was concomitantly increased, whereas expression of Ang-1, Tie 2, and the related receptor Tie-1 was unaltered. In situ hybridization localized Ang-2 mRNA to endothelial cells in hypoxic skin. These findings 1) show that up regulation of Ang-2 by hypoxia occurs widely in endothelial cells in vitro and in vivo; 2) suggest that induction of Ang-2, but not VEGF, by hypoxia in BME cells is controlled by a flavoprotein oxidoreductase that is sensitive to iodonium compounds; and 3) point to Ang-2 and VEGF as independently regulated and selective effectors of hypoxia-induced vascular sprouting. PMID- 10854230 TI - Complement C5b-9-mediated arachidonic acid metabolism in glomerular epithelial cells : role of cyclooxygenase-1 and -2. AB - In the passive Heymann nephritis (PHN) model of membranous nephropathy, complement C5b-9 induces glomerular epithelial cell (GEC) injury and proteinuria, which is partially mediated by eicosanoids. This study addresses the role of cyclooxygenase (COX)-1 and -2 in C5b-9-mediated eicosanoid production in GEC. Unstimulated rat GEC in culture primarily express COX-1. When stimulated with sublytic C5b-9, COX-2 was significantly up-regulated, whereas COX-1 was not affected. Compared with control, complement-treated GEC produced 32% more prostaglandin (PG) E(2) in the presence of exogenous substrate, and the increase was abolished with the COX-2-selective inhibitor, NS-398. Release of arachidonic acid from GEC phospholipids via C5b-9-induced activation of cytosolic phospholipase A(2) was associated with a marked stimulation of PGE(2) production, which was inhibited by 60% with NS-398. The results in cultured GEC were extended to GEC injury in vivo by examining COX-1 and -2 expression in PHN. Glomeruli from rats with PHN expressed significantly more COX-1 and COX-2, as compared with normal rats. PGE(2) production in glomeruli of rats with PHN was about twofold greater than in control glomeruli, and the increase was partially inhibited with NS-398. Thus, in GEC in culture and in vivo, C5b-9-induced eicosanoid production is regulated by both isoforms of COX. The inducible COX-2 may be an important novel mediator of C5b-9-induced glomerular injury. PMID- 10854231 TI - Complement activation promotes muscle inflammation during modified muscle use. AB - Modified muscle use can result in muscle inflammation that is triggered by unidentified events. In the present investigation, we tested whether the activation of the complement system is a component of muscle inflammation that results from changes in muscle loading. Modified rat hindlimb muscle loading was achieved by removing weight-bearing from the hindlimbs for 10 days followed by reloading through normal ambulation. Experimental animals were injected with the recombinant, soluble complement receptor sCR1 to inhibit complement activation. Assays for complement C4 or factor B in sera showed that sCR1 produced large reductions in the capacity for activation of the complement system through both the classical and alternative pathways. Analysis of complement C4 concentration in serum in untreated animals showed that the classical pathway was activated during the first 2 hours of reloading. Analysis of factor B concentration in untreated animals showed activation of the alternative pathway at 6 hours of reloading. Administration of sCR1 significantly attenuated the invasion of neutrophils (-49%) and ED1(+) macrophages (-52%) that occurred in nontreated animals after 6 hours of reloading. The presence of sCR1 also reduced significantly the degree of edema by 22% as compared to untreated animals. Together, these data show that increased muscle loading activated the complement system which then briefly contributes to the early recruitment of inflammatory cells during modified muscle loading. PMID- 10854232 TI - Cytochrome c-dependent activation of caspase-3 by tumor necrosis factor requires induction of the mitochondrial permeability transition. AB - The killing of L929 mouse fibroblasts by tumor necrosis factor-alpha (TNF-alpha) in the presence of 0.5 microg/ml actinomycin D (Act D) is prevented by inhibition of the mitochondrial permeability transition (MPT) with cyclosporin A (CyA) in combination with the phospholipase A(2) inhibitor aristolochic acid (ArA). The MPT is accompanied by the release of cytochrome c from the mitochondria, caspase 8 and caspase-3 activation in the cytosol, cleavage of the nuclear enzyme poly(ADP-ribose)polymerase (PARP), and DNA fragmentation, all of which were inhibited by CyA plus ArA. The caspase-3 inhibitor z-Asp-Glu-Val-aspartic acid fluoromethyl-ketone (Z-DEVD-FMK) did not prevent the loss of viability or the redistribution of cytochrome c, but it did prevent caspase-3 activation, PARP cleavage, and DNA fragmentation. Inhibition of the MPT reduced the activation of caspase-8 to the level occurring with TNF-alpha alone (no ActD). The caspase-8 inhibitor z-Ile-Glu(OMe)-Thr-Asp(OMe) fluoromethylketone (Z-IETD-FMK) did not prevent the cell killing and decreased only slightly the translocation of Bid to the mitochondria. These data indicate that induction of the MTP by TNF-alpha causes a release of cytochrome c, caspase-3 activation with PARP cleavage and DNA fragmentation. The loss of viability is dependent on the MPT but independent of the activation of caspase-3. The activation of caspase-8 is not dependent on the MPT. There is no evidence linking this enzyme to the loss of viability. Thus, the killing of L929 fibroblasts by TNF-alpha can occur in the absence of either caspase-3 or caspase-8 activity. Alternatively, cell death can be prevented despite an activation of caspase-8. PMID- 10854233 TI - Genetic progression and divergence in pancreatic carcinoma. AB - Genetic alterations of pancreatic intraductal lesions adjacent to invasive ductal carcinoma were investigated. We submitted nine foci of ordinary epithelium, 12 foci of nonpapillary hyperplasia, 12 foci of papillary hyperplasia (pap HP), 66 foci of severe ductal dysplasia, and 27 invasive foci from a total of 10 pancreatic carcinomas for genetic analysis. All foci were individually microdissected and allelic losses of 3p, 4q, 5q, 6q, 8p, 9p, 10q, 11q, 13q, 16q, 17p, and 18q were studied. All invasive and severely dysplastic intraductal foci exhibited loss of heterozygosity (LOH) at more than one chromosomal locus. For each case, allelic loss was frequently observed on 9p (severe ductal dysplasia 90%, invasion 100%), 17p (severe ductal dysplasia 80%, invasion 80%), and 18q (severe ductal dysplasia 88%, invasion 88%). Ninety-four percent of severe ductal dysplasia and 96% of invasive foci had multiple LOH. Seventeen percent of nonpapillary hyperplasia and 33% of pap HP showed LOH. Only one focus of pap HP showed multiple LOH. The patterns of allelic loss identified in severe ductal dysplasia were generally conserved in synchronous infiltrating tumors, supporting the paradigm that infiltrating tumors are clonally derived from severe ductal dysplasia. In eight of 10 cases, however, we found frequent genetic heterogeneity in the intraductal lesion, suggestive of genetic progression or diversion. These findings indicate that invasive pancreatic carcinoma evolves through successive and divergent genetic changes with selection of aggressive subclones in the intraductal component. PMID- 10854234 TI - Involvement of cyclins in cell proliferation and their clinical implications in soft tissue smooth muscle tumors. AB - Expression of cyclins A and E and cyclin-dependent kinase 2 (cdk2) was examined immunohistochemically in 55 cases of soft tissue smooth muscle tumors, including vascular leiomyoma, and compared to expression of Ki-67 and proliferating cell nuclear antigen. Cyclin A was expressed in 70% of the leiomyoma cases, but with much lower labeling indexes than in leiomyosarcoma. Cyclin E was expressed exclusively in leiomyosarcoma. Although the differences of cyclin A- and cyclin E labeling indexes between leiomyoma and leiomyosarcoma were statistically significant, no significant differences were found in the other markers. Furthermore, cyclin A- and/or E-positivity predicted a poor prognosis in recurrence- or metastasis-free survivals and overall survival. Immunoblotting revealed that cyclins A and E were expressed, in complex with cdk2, exclusively in tumors. In addition, not only leiomyosarcoma, but also leiomyoma specimens that exhibited negligible levels of complex expression, manifested detectable cdk2 activity. These results suggest 1) up-regulation of active cyclin A/cdk2 expression and associated kinase activity is critical for unrestrained cell proliferation; 2) cyclin E/cdk2 complexes may play a crucial role in leiomyosarcoma; 3) immunohistochemical detection of cyclins can be a more reliable tool for differential diagnosis between leiomyoma versus leiomyosarcoma than that of Ki-67 or proliferating cell nuclear antigen, and be a possible prognostic indicator. PMID- 10854235 TI - Expression of stress-response and cell proliferation genes in renal cell carcinoma induced by oxidative stress. AB - Ferric nitrilotriacetate induces oxidative damage in renal proximal tubules that ultimately leads to a high incidence of renal cell carcinoma (RCC) in rats. In search of genes specifically involved in oxystress-induced carcinogenesis, we have applied a modified fluorescent differential display technique to the tumors and an established cell line as well as their non-neoplastic counterparts. We screened approximately 84,000 products. Reverse Northern blotting confirmed differential expression of 20 transcripts, which showed either significant increase, decrease or lack of expression in the RCCs. Five cDNA clones encoded novel products of unknown function. Fifteen cDNA clones were identified by homology search, which included annexin II, Y-box binding protein, ribosomal proteins, heat shock proteins, DNA polymerase, nonmuscle caldesmon (increased); protein tyrosine phosphatase (decreased); selenoprotein P, stromal cell-derived factor 1, intestinal trefoil protein, nicotinamide adenine dinucleotide, reduced form (NADH) dehydrogenase, and insulin-like growth factor binding protein 7 (deleted). Most of the identified genes were associated with stress-response or cellular proliferation. These results suggest that multiple, interactive genetic pathways are involved in carcinogenesis induced by oxidative stress. PMID- 10854236 TI - Perturbation of hyaluronan interactions by soluble CD44 inhibits growth of murine mammary carcinoma cells in ascites. AB - Hyaluronan accumulates in ascites during intraperitoneal proliferation of TA3/St murine mammary carcinoma cells and at sites of their invasion of the peritoneal wall. To determine whether hyaluronan is functionally involved in these events, ascites tumor formation was compared in mice injected intraperitoneally with stable transfectants of TA3/St cells that overexpress soluble CD44, a hyaluronan binding protein, versus in mice injected with transfectants expressing mutated soluble CD44 that does not bind hyaluronan. The soluble CD44 transfectants temporarily grew at a reduced rate within the peritoneal cavity, then went into G(1) arrest and were subsequently cleared from the peritoneum. However, transfectants overexpressing mutant soluble CD44 that does not bind hyaluronan exhibited similar ascites accumulation, growth rates, and cell-cycle profiles in vivo to wild-type and vector-transfected TA3/St cells, all of which continued to grow until the tumors became fatal. The soluble CD44-transfected TA3/St cells also failed to attach to and form tumors in the peritoneal wall. When grown in vitro in soft agar, the soluble CD44 transfectants exhibited a dramatic reduction in colony formation compared to wild-type, vector-transfected, and mutant soluble CD44-transfected TA3/St cells. Thus, perturbation of hyaluronan interactions by soluble CD44 has a direct effect on the growth characteristics of these tumor cells, leading to inhibition of anchorage-independent growth in vitro and ascites growth in vivo. PMID- 10854237 TI - Dextran sulfate sodium-induced colonic histopathology, but not altered epithelial ion transport, is reduced by inhibition of phosphodiesterase activity. AB - Inhibition of phosphodiesterase (PDE) activity is beneficial in models of arthritis and airway inflammation. Here we assessed the ability of PDE inhibitors to modulate colitis by exposing mice to 4% (w/v) dextran sulfate sodium (DSS) drinking water for 5 days with or without rolipram, an inhibitor of PDE type 4, or the nonselective PDE inhibitor, pentoxifylline (both at 5 mg/kg, i.p., twice daily). Controls received saline, vehicle, or drug only. Colonic histology, myeloperoxidase (MPO) and tumor necrosis factor-alpha (TNF-alpha) levels, and epithelial ion transport (baseline and stimulated by electrical nerve stimulation, carbachol, and forskolin) were examined. DSS-treated mice displayed a variable diarrhea, significant histopathology in the mid-distal colon, elevated MPO activity, and reduced (>50%) responses to all three pro-secretory stimuli. Treatment with rolipram, and to a lesser extent pentoxifylline, significantly reduced the severity of the colonic histopathology and MPO levels. Neither PDE inhibitor had any affect on the diminished ion transport events caused by DSS induced colitis. However, although stimulated ion transport events were still reduced 3 days after DSS treatment, colonic segments from DSS + rolipram-treated mice displayed enhanced recovery in their secretory responsiveness, particularly to carbachol. These findings indicate that specific PDE4 inhibition can significantly reduce the tissue damage that accompanies colitis and enhance recovery of normal colonic function. PMID- 10854238 TI - Expression of Tie1, Tie2, and angiopoietins 1, 2, and 4 in Kaposi's sarcoma and cutaneous angiosarcoma. AB - The angiopoietins are recently described growth factors for vascular endothelium. The Tie1 and Tie2 receptors are expressed by endothelium. Acquired immune deficiency syndrome (AIDS)-associated Kaposi's sarcoma (KS) and cutaneous angiosarcoma are malignancies of endothelial origin. KS involves primarily the skin and mucosal surfaces and is common in AIDS patients. In an effort to determine whether the angiopoietins and Tie receptors play a role in the pathobiology of angiosarcoma and KS, we studied the expression of angiopoietin-1, angiopoietin-2, angiopoietin-4, Tie1, and Tie2 mRNAs in biopsies of KS from 12 AIDS patients, in biopsies of cutaneous angiosarcoma from two patients, and in control biopsies of normal skin from three volunteers by in situ hybridization. Strong expression of angiopoietin-2, Tie1, and Tie2 mRNAs was detected in the tumor cells of KS and cutaneous angiosarcomas, in contrast to the focal low-level expression in normal skin biopsies. Focal low-level expression of angiopoietin-1 was seen in KS, cutaneous angiosarcomas, and in normal skin. Focal low-level expression of angiopoietin-4 was identified in a minority of KS lesions. These findings suggest that the angiopoietins and Tie receptors may play an important role in the pathobiology of KS and cutaneous angiosarcoma and identify additional potential targets for therapeutic intervention in these vascular malignancies. PMID- 10854239 TI - Angiopoietin-1 and angiopoietin-2 activate trophoblast Tie-2 to promote growth and migration during placental development. AB - Human placental development involves coordinated angiogenesis and trophoblast outgrowth that are compromised in intrauterine growth restriction (IUGR). As Tie 2((-/-)) mice exhibit growth retardation and vascular network malformation, the expression of Tie-2 and its ligands, angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2), were investigated in human placenta from normal pregnancies and those complicated by severe IUGR. Ribonucleotide protection assays showed no significant change in the expression of Ang-2 mRNA between gestationally matched normal and IUGR placentas; however, immunoblots revealed that Ang-2 protein was significantly decreased in IUGR, suggesting that this may contribute to the abnormal development of the villous vasculature. In situ hybridization studies showed that Ang-1 and Tie-2 were detected in the cyto/syncytiotrophoblast bilayer in first-trimester placenta, whereas Ang-2 mRNA was restricted to the cytotrophoblast, suggesting their role in trophoblast function. At term, Ang-1 mRNA and immunoreactive protein were restricted to the paravascular tissues of the primary stem villi, supporting its role in vessel maturation. In contrast, Ang-2 was expressed throughout the term villous core, perhaps to permit the developing placental vascular network to remain in a state of fluidity. As these studies also revealed that trophoblast, in addition to endothelial cells, expressed Tie-2 receptors, we investigated the potential role of Ang-1/Ang-2 on trophoblast proliferation, migration, and the release of NO. Using spontaneously transformed first-trimester trophoblast cell lines that exhibit cytotrophoblast like (ED(27)) and extravillous trophoblast-like (ED(77)) properties, we show that the addition of Ang-2 (250 ng/ml) stimulated DNA synthesis in ED(27) trophoblast cells and triggered the release of NO. Ang-1 stimulated trophoblast (ED(77)) migration in a dose-dependent manner that was inhibited by recombinant Tie-2-FC. These data thus imply, for the first time, a specific role for angiopoietins as regulators of trophoblast behavior in the development of the utero/fetoplacental circulation, an action independent of their well-established roles in vascular endothelium. PMID- 10854240 TI - A2-Pancortins (Pancortin-3 and -4) are the dominant pancortins during neocortical development. AB - We have identified a novel mouse gene named pancortin that is expressed dominantly in the mature cerebral cortex. This gene produces four different species of proteins, Pancortin-1-4, sharing a common region in the middle of their structure with two variations at the N-terminal (A1 or A2 part) and C terminal (C1 or C2 part) sides, respectively. In the present study, we showed that expression of mRNAs for A2-Pancortins (Pancortin species that contain the A2 part, i.e., Pancortin-3 and -4) is more dominant than that of mRNAs for A1 Pancortins (Pancortin species that contain the A1 part, i.e., Pancortin-1 and -2) in the prenatal mouse cerebral neocortex. Using western blot analysis, we found that substantial amounts of both A2-Pancortins were present in the prenatal cerebral neocortex and P19 cells after inducing neuronal differentiation. A2 Pancortins were still present in the cerebral neocortex of the adult, although their mRNAs were hardly detected. In contrast, the amount of A1-Pancortins did not increase after the third postnatal week in spite of their intense gene expression. Furthermore, we showed that recombinant Pancortin-3, one of the A2 Pancortins, was a secreted protein, in contrast to Pancortin-1 (one of the A1 Pancortins). These results suggest that A2-Pancortins are extracellular proteins essential for neuronal differentiation and that their molecular behavior is distinct from that of A1-Pancortins. PMID- 10854241 TI - A systemic administration of NMDA induces immediate early gene pip92 in the hippocampus. AB - In the mammalian CNS, aspartate and glutamate are major excitatory amino acids, and their receptors are believed to mediate a wide range of physiological and pathological processes, including neurotransmission, plasticity, excitotoxicity, and various forms of neurodegeneration. The immediate early gene pip92 has been identified in serum-stimulated BALB/c 3T3 fibroblasts, activated T lymphocytes treated with cycloheximide, and fibroblast growth factor-stimulated hippocampal cells during neuronal differentiation. In this study we have demonstrated that pip92 is expressed in the mouse brain after a single intraperitoneal injection of NMDA. The distribution of pip92 mRNA levels in the NMDA-treated mouse brain was investigated using in situ RT-PCR. The region-specific activation of pip92 in the CNS was observed 3 h after NMDA injection, and high levels of pip92 mRNA were detected in the hippocampal dentate gyrus and piriform cortex regions. In addition, the activation of pip92 by NMDA was mediated by activation of mitogen activated protein kinases (MAPKs), such as c-Jun N-terminal kinase (JNK) and p38 kinase, but not extracellular signal-regulated kinase (ERK) in the mouse hippocampus and immortalized rat hippocampal progenitor cells. This study suggests that pip92 is likely to play an important role in neuronal cell death induced by excitotoxic NMDA injury in the CNS. PMID- 10854242 TI - Neuronal and extraneuronal expression and regulation of the human alpha5 nicotinic receptor subunit gene. AB - The mRNA encoding the human alpha5 nicotinic subunit was detected in several structures of the nervous system but appeared to be mainly expressed in cerebellum, thalamus, and the autonomic ganglia. For the first time, the alpha5 transcript was also detected in several non-neuronal tissues, with maximal expressions being found throughout the gastrointestinal tract, thymus, and testis. Many other extraneuronal sites expressed alpha5, but there were also nonexpressing organs, such as the liver, spleen, and kidney. To understand the transcriptional mechanisms controlling such a diversified expression of alpha5 in neuronal and nonneuronal cells, we isolated the 5'-regulatory region of the human gene and characterized its properties. Here we identify the alpha5 core promoter and demonstrate that the DNA regions surrounding it contain elements (with positive or negative activities) that work in a tissue-specific fashion. In particular, the segment specifying the 5'-untranslated region in neuronal cells has most of the properties of an enhancer because it activates a heterologous promoter in a position- and orientation-independent fashion. We therefore conclude that the expression of alpha5 relies on a highly complex promoter that uses distinct regulatory elements to comply with the different functional and developmental requirements of the various tissues and organs. PMID- 10854243 TI - Disruption of the epilepsy KCNQ2 gene results in neural hyperexcitability. AB - Benign familial neonatal convulsion (BFNC) is a common idiopathic epilepsy with autosomal dominant inheritance. Recently, two novel voltage-dependent potassium channel genes, KCNQ2 and KCNQ3, were identified by positional cloning as being responsible for BFNC. Heterotetramers of the products of these genes form M channels and regulate the threshold of electrical excitability of neurons. We disrupted the mouse KCNQ2 gene via gene targeting to study the relationship between KCNQ2 and epilepsy. Homozygous pups (KCNQ2 -/-) died within a few hours after birth owing to pulmonary atelectasis that was not due to the status of epileptic seizures, although their development was morphologically normal. Heterozygous mice had decreased expression of KCNQ2 and showed hypersensitivity to pentylenetetrazole, an inducer of seizure. These data indicate that the decreased expression of KCNQ2 might cause a hyperexcitability of the CNS, which accounts for the mechanism of BFNC. PMID- 10854244 TI - Expression of retinoid receptors during the retinoic acid-induced neuronal differentiation of human embryonal carcinoma cells. AB - Retinoic acid (RA), a derivative of vitamin A, is essential for normal patterning and neurogenesis during development. Until recently, studies have been focused on the physiological roles of RA receptors (RARs), one of the two types of nuclear receptors, whereas the functions of the other nuclear receptors, retinoid X receptors (RXRs), have not been explored. Accumulating evidence now suggests that RXRalpha is a critical receptor component mediating the effects of RA during embryonic development. In this study, we have examined the expression profiles of RXRalpha and RARs during the RA-induced neuronal differentiation in a human embryonal carcinoma cell line, NT2. Distinct expression profiles of RXRalpha, RARalpha, RARbeta, and RARgamma were observed following treatment with RA. In particular, we found that RA treatment resulted in a biphasic up-regulation of RXRalpha expression in NT2 cells. The induced RXRalpha was found to bind specifically to the retinoid X response element based on gel mobility retardation assays. Furthermore, immunocytochemical analysis revealed that RXRalpha expression could be localized to the somatoaxonal regions of the NT2 neurons, including the tyrosine hydroxylase- and vasoactive intestinal peptide-positive neurons. Taken together, our findings provide the first demonstration of the cellular localization and regulation of RXRalpha expression in NT2 cells and suggest that RXRalpha might play a crucial role in the cellular functions of human CNS neurons. PMID- 10854245 TI - Alternative splicing of the neurotrophin-3 gene gives rise to different transcripts in a number of human and rat tissues. AB - The mouse neurotrophin-3 (NT-3) gene has been shown to contain two exons (exon 1A and exon 1B) upstream of the single coding exon (exon 2). These upstream exons are alternatively spliced to the coding exon, generating two different NT-3 transcripts. We investigated whether alternative splicing of two upstream exons also occurs in the human and rat NT-3 gene. It was found that the human and rat NT-3 gene also contains two exons upstream of the main coding exon and that alternative splicing of these upstream exons generates two different NT-3 transcripts : transcript 1A and transcript 1B (TR1B). These two transcripts were widely expressed in several human and rat tissues. Also, a third transcript, transcript 1A1C, derived from splicing of exon 1A to exon 1B before splicing to the coding exon was seen in a small number of rat tissues. Previous quantification of neurotrophin-3 mRNA has not been transcript-specific. Here we describe a transcript-specific semiquantitative RT-PCR method allowing the quantification of TR1B in human tissues. We show that this is the major NT-3 transcript and that expression of this transcript was much higher in the adult when compared with the corresponding fetal tissues. PMID- 10854246 TI - Ubiquitination and degradation of the zebrafish paired-like homeobox protein VSX 1. AB - Vsx-1 is a paired-like : CVC homeobox protein dynamically expressed during zebrafish development. Previous results indicate that Vsx-1 influences bipolar cell differentiation and maintenance of these cells in the adult retina. To understand the developmental regulation of this transcription factor, we investigated ubiquitination as a possible posttranslational mechanism. In vitro, Vsx-1 was conjugated with multiple ubiquitin moieties. Proteasome inhibitors and added ubiquitin increased the accumulation of Vsx-1-ubiquitin(n) complexes and stabilized unmodified Vsx-1. Also, in transiently transfected COS-7 cells, Vsx-1 is ubiquitinated, and pulse-chase experiments show that Vsx-1 proteolysis occurs. Vsx-1 proteins with C-terminal deletions retained the capacity for initial modification by ubiquitin but lost the capacity for efficient chain elongation. These results show that Vsx-1 is a substrate of the ubiquitin/proteasome pathway and suggest that C-terminal sequences of Vsx-1 are critical for ubiquitin chain elongation. In addition, our findings suggest that ubiquitin-dependent proteolysis regulates Vsx-1 during zebrafish retinal development. PMID- 10854247 TI - Glutamic acid decarboxylase-expressing astrocytes exhibit enhanced energetic metabolism and increase PC12 cell survival under glucose deprivation. AB - Astrocytes play a key role by catabolizing glutamate from extracellular space into glutamine and tricarboxylic acid components. We previously produced an astrocytic cell line that constitutively expressed glutamic acid decarboxylase (GAD67), which converts glutamate into GABA to increase the capacity of astrocytes to metabolize glutamate. In this study, GAD-expressing astrocytes in the presence of glutamate were shown to have increased energy metabolism, as determined by a moderate increase of 3-(4, 5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide reduction, by an increased ATP level, and by enhanced lactate release. These changes were due to GAD transgene expression because transient expression of a GAD antisense plasmid resulted in partial suppression of the ATP level increase. These astrocytes had an increased survival in response to glucose deprivation in the presence of glutamate compared with the parental astrocytes, and they were also able to enhance survival of a neuronal-like cell line (PC12) under glucose deprivation. This protection may be partially due to the increased lactate release by GAD-expressing astrocytes because PC12 cell survival was enhanced by lactate and pyruvate under glucose deprivation. These results suggest that the establishment of GAD expression in astrocytes enhancing glutamate catabolism could be an interesting strategy to increase neuronal survival under hypoglycemia conditions. PMID- 10854248 TI - The endogenous amine 1-methyl-1,2,3,4- tetrahydroisoquinoline prevents the inhibition of complex I of the respiratory chain produced by MPP(+). AB - The endogenous monoamine 1-methyl-1,2,3,4-tetrahydroisoquinoline has been shown to prevent the neurotoxic effect of MPP(+) and other endogenous neurotoxins, which produce a parkinsonian-like syndrome in humans. We have tested its potential protective effect in vivo by measuring the protection of 1-methyl 1,2,3,4-tetrahydroisoquinoline in the neurotoxicity elicited by MPP(+) in rat striatum by tyrosine hydroxylase immunocytochemistry. Because we know that cellular damage caused by MPP(+) is primarily the result of mitochondrial respiratory inhibition at the complex I level, we have extended the study further to understand this protective mechanism. We found that the inhibitory effect on the mitochondrial respiration rate induced by MPP(+) in isolated rat liver mitochondria and striatal synaptosomes was prevented by addition of 1-methyl 1,2,3,4-tetrahydroisoquinoline. This compound has no antioxidant capacity; therefore, this property is not involved in its protective effect. Thus, we postulate that the preventive effect that 1-methyl-1,2,3,4-tetrahydroisoquinoline has on mitochondrial inhibition for MPP(+) could be due to a "shielding effect," protecting the energetic machinery, thus preventing energetic failure. These results suggest that this endogenous amine may protect against the effect of several parkinsonism-inducing compounds that are associated with progressive impairment of the mitochondrial function. PMID- 10854249 TI - Nerve growth factor-induced differentiation does not alter the biochemical properties of a mutant prion protein expressed in PC12 cells. AB - Insertional and point mutations in the gene encoding the prion protein (PrP) are responsible for familial prion diseases. We have previously generated lines of Chinese hamster ovary cells that express PrP molecules carrying pathogenic mutations, and found that the mutant proteins display several biochemical properties reminiscent of PrP(Sc), the infectious isoform of PrP. To analyze the properties and effects of mutant PrP molecules expressed in cells with a neuronal phenotype, we have constructed stably transfected lines of PC12 cells that synthesize a PrP molecule carrying a nine-octapeptide insertion. We report here that this mutant PrP acquires scrapie-like properties, including detergent insolubility, protease resistance, and resistance to phospholipase cleavage of its glycolipid anchor. A detergent-insoluble and phospholipase-resistant form of the mutant protein is also released spontaneously into conditioned medium. These scrapie-like biochemical properties are quantitatively similar to those seen in Chinese hamster ovary cells and are not affected by differentiation of the PC12 cells into sympathetic neurons by nerve growth factor. Moreover, there is no detectable effect of mutant PrP expression on the morphology or viability of the cells in either the differentiated or undifferentiated state. These results indicate that conversion of mutant PrP into a PrP(Sc)-like form does not depend critically on the cellular context, and they suggest that mutant PrP expressed in cultured cells, even those having the phenotype of differentiated neurons, is not neurotoxic. PMID- 10854250 TI - Neuronal apoptosis induced by pharmacological concentrations of 3 hydroxykynurenine: characterization and protection by dantrolene and Bcl-2 overexpression. AB - We have studied neurotoxicity induced by pharmacological concentrations of 3 hydroxykynurenine (3-HK), an endogenous toxin implicated in certain neurodegenerative diseases, in cerebellar granule cells, PC12 pheochromocytoma cells, and GT1-7 hypothalamic neurosecretory cells. In all three cell types, the toxicity was induced in a dose-dependent manner by 3-HK at high micromolar concentrations and had features characteristic of apoptosis, including chromatin condensation and internucleosomal DNA cleavage. In cerebellar granule cells, the 3-HK neurotoxicity was unaffected by xanthine oxidase inhibitors but markedly potentiated by superoxide dismutase and its hemelike mimetic, MnTBAP [manganese(III) tetrakis(benzoic acid)porphyrin chloride]. Catalase blocked 3-HK neurotoxicity in the absence and presence of superoxide dismutase or MnTBAP. The formation of H(2)O(2) was demonstrated in PC12 and GT1-7 cells treated with 3-HK, by measuring the increase in the fluorescent product, 2',7'-dichlorofluorescein. In both PC12 and cerebellar granule cells, inhibitors of the neutral amino acid transporter that mediates the uptake of 3-HK failed to block 3-HK toxicity. However, their toxicity was slightly potentiated by the iron chelator, deferoxamine. Taken together, our results suggest that neurotoxicity induced by pharmacological concentrations of 3-HK in these cell types is mediated primarily by H(2)O(2), which is formed most likely by auto-oxidation of 3-HK in extracellular compartments. 3-HK-induced death of PC12 and GT1-7 cells was protected by dantrolene, an inhibitor of calcium release from the endoplasmic reticulum. The protection by dantrolene was associated with a marked increase in the protein level of Bcl-2, a prominent antiapoptotic gene product. Moreover, overexpression of Bcl-2 in GT1-7 cells elicited by gene transfection suppressed 3 HK toxicity. Thus, dantrolene may elicit its neuroprotective effects by mechanisms involving up-regulation of the level and function of Bcl-2 protein. PMID- 10854251 TI - The transcription factor E2F1 modulates apoptosis of neurons. AB - The transcription factor E2F1 is known to mediate apoptosis in isolated quiescent and postmitotic cardiac myocytes, and its absence decreases the size of brain infarction following cerebral ischemia. To demonstrate directly that E2F1 modulates neuronal apoptosis, we used cultured cortical neurons to show a temporal association of the transcription and expression of E2F1 in neurons with increased neuronal apoptosis. Cortical neurons lacking E2F1 expression (derived from E2F1 -/- mice) were resistant to staurosporine-induced apoptosis as evidenced by the significantly lower caspase 3-like activity and a lesser number of cells with apoptotic morphology in comparison with cortical cultures derived from wild-type mice. Furthermore, overexpressing E2F1 alone using replication deficient recombinant adenovirus was sufficient to cause neuronal cell death by apoptosis, as evidenced by the appearance of hallmarks of apoptosis, such as the threefold increase in caspase 3-like activity and increased laddered DNA fragmentation, in situ endlabeled DNA fragmentation, and numbers of neuronal cells with punctate nuclei. Taken together, we conclude that E2F1 plays a key role in modulating neuronal apoptosis. PMID- 10854252 TI - Nuclear factor-kappaB mediates the cell survival-promoting action of activity dependent neurotrophic factor peptide-9. AB - Activity-dependent neurotrophic factor (ADNF) is produced by astrocytes in response to neuronal depolarization and, in turn, promotes neuronal survival. A nineamino acid ADNF peptide (ADNF9) exhibits full neurotrophic activity and potently protects cultured embryonic rat hippocampal neurons from oxidative injury and apoptosis. Picomolar concentrations of ADNF9 induced an increase in nuclear factor-kappaB (NF-kappaB) DNA-binding activity within 1 h of exposure, with a maximum increase of approximately 10-fold by 6 h. Activation of NF-kappaB was correlated with increased resistance of neurons to apoptosis induced by exposure to Fe(2+). The antiapoptotic action of ADNF9 was abolished when NF kappaB activation was specifically blocked with kappaB decoy DNA. Oxidative stress was attenuated in neurons pretreated with ADNF9, and this effect of ADNF9 was blocked by kappaB decoy DNA, suggesting that ADNF9 suppresses apoptosis by reducing oxidative stress. ADNF9 also prevented neuronal apoptosis following trophic factor withdrawal via an NF-kappaB-mediated mechanism. Thus, NF-kappaB mediates the neuron survival-promoting effects of ADNF9 in experimental models relevant to developmental neuronal death and neurodegenerative disorders. PMID- 10854253 TI - Isolation and characterization of novel presenilin binding protein. AB - Approximately 50% of familial Alzheimer's disease (AD) cases are linked to the presenilin (PS) gene. This suggests that an altered function of mutated PSs accounts for a fundamental process leading to AD. Here we identify a new PS binding protein, PBP, which is highly expressed in cerebral cortex and hippocampus. immunohistochemical studies and cell fractionation analysis show that PBP redistributes from cytoplasm to membranes in the presence of PS. In addition, PBP is deficient in the soluble fraction of sporadic AD brains. PMID- 10854254 TI - The catalytic subunit of telomerase protects neurons against amyloid beta-peptide induced apoptosis. AB - The catalytic subunit of telomerase (TERT) is a specialized reverse transcriptase that has been associated with cell immortalization and cancer. It was reported recently that TERT is expressed in neurons throughout the brain in embryonic and early postnatal development, but is absent from neurons in the adult brain. We now report that suppression of TERT levels and function in embryonic mouse hippocampal neurons in culture using antisense technology and the telomerase inhibitor 3' -azido-2' 3' -dideoxythymidine significantly increases their vulnerability to cell death induced by amyloid beta-peptide, a neurotoxic protein believed to promote neuronal degeneration in Alzheimer's disease. Neurons in which TERT levels were reduced exhibited increased levels of oxidative stress and mitochondrial dysfunction following exposure to amyloid beta-peptide. Overexpression of TERT in pheochromocytoma cells resulted in decreased vulnerability to amyloid beta-peptide-induced apoptosis. Our findings demonstrate a neuroprotective function of TERT in an experimental model relevant to Alzheimer's disease, and suggest the possibility that restoration of TERT expression in neurons in the adult brain may protect against age-related neurodegeneration. PMID- 10854255 TI - Contribution of anion transporters to the acidosis-induced swelling and intracellular acidification of glial cells. AB - This study examines the contribution of anion transporters to the swelling and intracellular acidification of glial cells from an extracellular lactacidosis, a condition well-known to accompany cerebral ischemia and traumatic brain injury. Suspended C6 glioma cells were exposed to lactacidosis in physiological or anion depleted media, and different anion transport inhibitors were applied. Changes in cell volume and intracellular pH (pH(i)) were simultaneously quantified by flow cytometry. Extracellular lactacidosis (pH 6.2) led to an increase in cell volume to 125.1 +/- 2.5% of baseline within 60 min, whereas the pH(i) dropped from the physiological value of 7.13 +/- 0.05 to 6.32 +/- 0.03. Suspension in Cl(-)-free or HCO(3)(-)/CO(2)-free media or application of anion transport inhibitors [0.1 mM bumetanide or 0.5 mM 4, 4'-diisothio-cyanatostilbene-2,2'-disulfonic acid (DIDS)] did not affect cell volume during baseline conditions but significantly reduced cell swelling from lactacidosis. In addition, the Cl(-)-free or HCO(3)( )/CO(2)-free media and DIDS attenuated intracellular acidosis on extracellular acidification. From these findings it is concluded that besides the known activation of the Na(+)/H(+) exchanger, activation of the Na(+)-independent Cl( )/HCO(3)(-) exchanger and the Na(+)-K(+)-Cl(-) cotransporter contributes to acidosis-induced glial swelling and the intracellular acidification. Inhibition of these processes may be of interest for future strategies in the treatment of cytotoxic brain edema from cerebral ischemia or traumatic brain injury. PMID- 10854256 TI - Effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine on differentiating mouse N2a neuroblastoma cells. AB - The effect of the neurotoxin 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP) was investigated in mouse N2a neuroblastoma cells, induced to differentiate by serum withdrawal and addition of dibutyryl cyclic AMP, over a 24-h period. Addition of MPTP (10 microM) during differentiation caused a change in cell morphology characterised by an inhibition of axon outgrowth, in the absence of cell death. Biochemical characterisation by western blotting revealed that MPTP had no significant effects on the levels of actin, alpha-tubulin, or total heavy chain neurofilament (NF-H). However, NF-H phosphorylation appeared to increase following MPTP treatment when blots were probed with the phosphorylation state specific antibodies RMd09 and Ta51. In addition, indirect immunofluorescence analysis revealed an accumulation of phosphorylated NF-H in the cell perikaryon, suggesting that altered NF-H distribution was associated with the observed effects of MPTP on cell morphology. These changes may represent a useful in vitro marker of MPTP neurotoxicity within a simple differentiating neuronal cell model system. PMID- 10854257 TI - trans-resveratrol protects embryonic mesencephalic cells from tert-butyl hydroperoxide: electron paramagnetic resonance spin trapping evidence for a radical scavenging mechanism. AB - In recent years, the antioxidant and other pharmacological properties of resveratrol, a natural product present in grapes and wine, have attracted considerable interest from the biomedical research community. In an examination of the potential neuroprotective properties of the compound, we have investigated the ability of resveratrol to protect rat embryonic mesencephalic tissue, rich in dopaminergic neurones, from the prooxidant tert-butyl hydroperoxide. Using the electron paramagnetic resonance (EPR) spin-trapping technique, the main radicals detected in cell suspensions were the tert-butoxyl radical and the methyl radical, indicating the one-electron reduction of the peroxide followed by a beta scission reaction. The appearance of EPR signals from the trapped radicals preceded the onset of cytotoxicity, which was almost exclusively necrotic in nature. The inclusion of resveratrol in incubations resulted in the marked protection of cells from tert-butyl hydroperoxide. In parallel spin-trapping experiments, we were able to demonstrate the scavenging of radicals by resveratrol, which involved direct competition between resveratrol and the spin trap for reaction with the radicals. To our knowledge, this is the first example in which cytoprotection by resveratrol has been demonstrated by EPR spin-trapping competition kinetics to be due to its scavenging of the radicals responsible for the toxicity of a prooxidant. PMID- 10854258 TI - Endogenous and exogenous fibroblast growth factor 2 support survival of chick retinal neurons by control of neuronal neuronal bcl-x(L) and bcl-2 expression through a fibroblast berowth factor receptor 1- and ERK-dependent pathway. AB - Fibroblast growth factor (FGF) 2 is a survival factor for various cell types, including retinal neurons. However, little is understood about the molecular bases of the neuroprotective role of FGF2 in the retina. In this report, FGF2 survival activity was studied in chick retinal neurons subjected to apoptosis by serum deprivation. Exogenous FGF2 supported neuronal survival after serum deprivation and increased neuronal bcl-x(L) and bcl-2 expression, through binding to its receptor R1 (FGF-R1), and subsequent extracellular signal-regulated kinase (ERK) activation. Endogenous FGF2 was transiently overexpressed after serum deprivation. Its down-regulation by antisense oligonucleotides and blockade of its signaling pathway (binding to FGF-R1, tyrosine phosphorylation, and ERK inhibition) decreased bcl-x(L) and bcl-2 levels and and enhanced apoptosis, suggesting that endogenous FGF2 supported neuronal survival through a pathway similar to that of exogenous FGF2. This pathway may serve to up-regulate, or maintain, bcl-x(L) and bcl-2 levels that normally decrease during the onset of apoptosis. Indeed, long-term ERK activation and high bcl-x(L) levels are necessary for the survival activity of both exogenous and endogenous FGF2. Because FGF2 is upregulated following retinal injury in vivo, we suggest that an injury-stimulated autocrine/paracrine FGF2 loop may serve to maintain high levels of survival proteins, such as Bcl-x(L), through ERK activation in retinal neurons. PMID- 10854259 TI - Association of a lower molecular weight protein to the mu-opioid receptor demonstrated by (125)I-beta-endorphin cross-linking studies. AB - Cross-linking experiments using the (125)I-beta-endorphin revealed the presence of several receptor-related species in cell lines expressing endogenous opioid receptors, including a small molecular mass protein (approximately 22 kDa). Previous reports have suggested that this 22-kDa (125)I-beta-endorphin cross linked protein could be the degradative product from a higher molecular mass species, i.e., a fragment of the receptor. To determine if this protein is indeed a degraded receptor fragment, (125)I-beta-endorphin was cross-linked to the (His)(6) epitope-tagged mu-opioid receptor (His-mu) stably expressed in the murine neuroblastoma Neuro(2A) cells. Similar to earlier reports with cell lines expressing endogenous receptors, two major bands of 72- and 25-kDa proteins were specifically cross-linked. Initial cross-linking experiments indicated the absolute requirement of the high-affinity (125)I-beta-endorphin binding to the mu opioid receptor prior to the appearance of the low molecular weight species, suggesting that the 22-kDa protein could be a degraded fragment of the receptor. However, variations in the ratios of these protein bands being cross-linked by several homo- or heterobifunctional cross-linking agents were observed. Although neither the carboxyl terminus mu-opioid receptor-specific antibodies nor the antibodies against the epitope at the amino terminus of the receptor could recognize the 22-kDa protein, this (125)I-beta-endorphin cross-linked species could be coimmunoprecipitated with the receptor antibodies or could be isolated with a nickel resin affinity chromatography. The direct physical association of the 22-kDa protein with the receptor was demonstrated also by the observation that the 22-kDa protein could not bind to the nickel resin alone, but that its binding to the nickel resin was restored in the presence of the His-mu. Taken together, these results suggest that the 22-kDa protein cross-linked by (125)I beta-endorphin is not a degradative product, but a protein located within the proximity of the mu-opioid receptor, and that it is tightly associated with the receptor. PMID- 10854260 TI - Changes in NMDA receptor subunit expression in response to contusive spinal cord injury. AB - Differential assembly of N-methyl-D-aspartate (NMDA) receptor subunits determines their functional characteristics. Using in situ hybridization, we found a selective increase of the subunits NR1 and NR2A mRNA at 24 h in ventral motor neurons (VMN) caudal to a standardized spinal cord contusion injury (SCI). Other neuronal cell populations and VMN rostral to the injury site appeared unaffected. Significant up-regulation of NR2A mRNA also was seen 1 month after SCI in thoracic and lumbar VMN. The selective effects on VMN caudal to the injury site suggest that the loss of descending innervation leads to increased NMDA receptor subunit expression in these cells after SCI, which may alter their responses to glutamate. In contrast, protein levels determined by western blot analysis show decreased levels of NR2A 1 month after SCI in whole thoracic segments of spinal cord that included the injury sites. No effects of injury were seen on subunit levels in cervical or lumbar segments. Taken together with our previous study showing alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor subunit down-regulation after injury, our data suggest that glutamate receptor composition is significantly altered after SCI. These changes need to be taken into account to properly understand the function of, and potential pharmacotherapy for, the chronically injured spinal cord. PMID- 10854261 TI - Expression of mRNAs encoding for two different olfactory receptors in a subset of olfactory receptor neurons. AB - Evidence has accumulated to support a model for odorant detection in which individual olfactory receptor neurons (ORNs) express one of a large family of G protein-coupled receptor proteins that are activated by a small number of closely related volatile chemicals. However, the issue of whether an individual ORN expresses one or multiple types of receptor proteins has yet to be definitively addressed. Physiological data indicate that some individual ORNs can be activated by odorants differing substantially in structure and/or perceived quality, suggesting multiple receptors or one nonspecific receptor per cell. In contrast, molecular biological studies favor a scheme with a single, fairly selective receptor per cell. The present studies directly assessed whether individual rat ORNs can express multiple receptors using single-cell PCR techniques with degenerate primers designed to amplify a wide variety of receptor sequences. We found that whereas only a single OR sequence was obtained from most ORNs examined, one ORN produced two distinct receptor sequences that represented different receptor gene families. Double-label in situ hybridization studies indicated that a subset of ORNs co-express two distinct receptor mRNAs. A laminar segregation analysis of the cell nuclei of ORNs labeled with the two OR mRNA probes showed that for one probe, the histogram of the distribution of the cell nuclei along the depth of the epithelium was bimodal, with one peak overlapping the (unimodal) histogram for the other probe. These results are consistent with co-expression of two OR mRNAs in a population of single ORNs. PMID- 10854262 TI - HIV-1 coat glycoprotein gp120 induces apoptosis in rat brain neocortex by deranging the arachidonate cascade in favor of prostanoids. AB - Human immunodeficiency virus type-1 coat glycoprotein gp 120 causes delayed programmed cell death (apoptosis) in rat brain neocortex. Here, we investigated the possible role of the arachidonate cascade and membrane peroxidation in this process. It is shown that gp 120 causes a rapid increase in the activity and expression of the arachidonate-metabolizing enzyme prostaglandin H synthase, paralleled by increased prostaglandin E(2) levels. The selective inhibitor of prostaglandin H synthase indomethacin inhibited enzyme activity, reduced prostaglandin E(2) content, and partially protected neocortex against gp 120 induced apoptosis. Conversely, the activity and expression of the arachidonate metabolizing enzyme 5-lipoxygenase decreased upon gp 120 treatment, as well as the level of its product, leukotriene B(4). Treatment with gp 120 also reduced membrane lipid peroxidation, and this may be implicated in the execution of programmed cell death. These results suggest that early derangement of the arachidonate cascade in favor of prostanoids may be instrumental in the execution of delayed apoptosis in the brain neocortex of rats. PMID- 10854263 TI - The activation and inhibition of human nicotinic acetylcholine receptor by RJR 2403 indicate a selectivity for the alpha4beta2 receptor subtype. AB - Human nicotinic acetylcholine (ACh) receptor subtypes expressed in Xenopus oocytes were characterized in terms of their activation by the experimental agonist RJR-2403. Responses to RJR-2403 were compared with those evoked by ACh and nicotine. These agonists were also characterized in terms of whether application of the drugs had the effect of producing a residual inhibition that was manifest as a decrease in subsequent control responses to ACh measured 5 min after the washout of the drug. For the activation of alpha4beta2 receptors, RJR 2403 had an efficacy equivalent to that of ACh and was more potent than ACh. RJR 2403 was less efficacious than ACh for other human receptor subtypes, suggesting that it is a partial agonist for all these receptors. Nicotine activated peak currents in human alpha4beta2 and alpha3beta2 receptors that were 85 and 50% of the respective ACh maximum responses. Nicotine was an efficacious activator of human alpha7 receptors, with a potency similar to ACh, whereas RJR-2403 had very low potency and efficacy for these receptors. At concentrations of <1 mM, RJR 2403 did not produce any residual inhibition of subsequent ACh responses for any receptor subtype. In contrast, nicotine produced profound residual inhibition of human alpha4beta2, alpha3beta2, and alpha7 receptors with IC(50) values of 150, 200, and 150 microM, respectively. Co-expression of the human alpha5 subunit with alpha3 and beta2 subunits had the effect of producing protracted responses to ACh and increasing residual inhibition by ACh and nicotine but not RJR-2403. In conclusion, our results, presented in the context of the complex pharmacology of nicotine for both activating and inhibiting neuronal nicotinic receptor subtypes, suggest that RJR-2403 will be a potent and relatively selective activator of human alpha4beta2 receptors. PMID- 10854264 TI - Rod-type transducin alpha-subunit mediates a phototransduction pathway in the chicken pineal gland. AB - The chicken pineal gland is a photosensitive neuroendocrine organ producing melatonin in circadian clock-regulated and light-sensitive manners. To understand the relationship between the photoreceptive molecule pinopsin and the light dependent melatonin suppression that is sensitive to pertussis toxin treatment, we have searched for pertussis toxin-sensitive G protein alpha-subunits expressed in the chicken pineal gland. Here we report the cDNA cloning of the pineal transducin alpha-subunit (Gtalpha), which is highly homologous to human retinal rod cell-specific Gt(1)alpha. Concurrent cDNA cloning of chicken retinal Gt(1)alpha and Gt(2)alpha (rod and cone cell-specific alpha-subunits of transducin, respectively) revealed that the chicken pineal Gtalpha is identical to the retinal Gt(1)alpha. Double-immunostaining analysis of the chicken pineal sections localized Gt(1)alpha-immunoreactivity in the rudimentary outer segments of both follicular and parafollicular pinealocytes that were immunopositive to anti-pinopsin antibody. To examine whether pineal Gt(1)alpha is involved in the pineal phototransduction pathway, trypsin protection assay was applied for detecting the conversion of GDP-bound Gt(1)alpha into the guanosine 5'-O-(3 thiotriphosphate) (GTPgammaS)-bound form in the pineal membrane homogenate. It was clearly demonstrated that the pineal Gt(1)alpha is activated in a light dependent manner in the presence of GTPgammaS. These data together suggest strongly that pineal Gt(1)alpha mediates the phototransduction pathway triggered by pinopsin in the chicken pinealocytes. PMID- 10854265 TI - Stoichiometry of tyrosine hydroxylase phosphorylation in the nigrostriatal and mesolimbic systems in vivo: effects of acute haloperidol and related compounds. AB - Electrical stimulation of the medial forebrain bundle increases (32)P incorporation into striatal tyrosine hydroxylase (TH) at Ser (19), Ser(31), and Ser(40). In the present studies, the effects of acute haloperidol and related drugs on sitespecific TH phosphorylation stoichiometry (PS) in the nigrostriatal and mesolimbic systems were determined by quantitative blot immunolabeling using phosphorylation statespecific antibodies. The striatum (Str), substantia nigra (SN), nucleus accumbens (NAc), and ventral tegmental area (VTA) from Sprague Dawley rats were harvested 30-40 min after a single injection of either vehicle, haloperidol (2 mg/kg), raclopride (2 mg/kg), clozapine (30 mg/kg), or SCH23390 (0.5 mg/kg). In vehicle-injected control rats, Ser(19) PS was 1.5- to 2. 5-fold lower in Str and NAc than in SN and VTA, Ser(31) PS was two-to fourfold higher in Str and NAc than in SN and VTA, and Ser(40) PS was similar between the terminal field and cell body regions. After haloperidol, Ser(40) PS increased twofold in Str and NAc, whereas a smaller increase in SN and VTA was observed. The effects of haloperidol on Ser(19) PS were similar to those on Ser(40) in each region; however, haloperidol treatment increased Ser(31) PS at least 1.6-fold in all regions. The effects of raclopride on TH PS were comparable to those of haloperidol, whereas clozapine treatment increased TH PS at all sites in all regions. By contrast, the effects of SCH23390 on TH PS were relatively small and restricted to the NAc. The stoichiometries of site-specific TH phosphorylation in vivo are presented for the first time. The nigrostriatal and mesolimbic systems have common features of TH PS, distinguished by differences in TH PS between the terminal field and cell body regions and by dissimilar increases in TH PS in the terminal field and cell body regions after acute haloperidol. PMID- 10854266 TI - Involvement of betagamma subunits of G(q/11) in muscarinic M(1) receptor potentiation of corticotropin-releasing hormone-stimulated adenylyl cyclase activity in rat frontal cortex. AB - In the present study, we investigated the involvement of betagamma subunits of G(q/11) in the muscarinic M(1) receptor-induced potentiation of corticotropin releasing hormone (CRH)-stimulated adenylyl cyclase activity in membranes of rat frontal cortex. Tissue exposure to either one of two betagamma scavengers, the QEHA fragment type II adenylyl cyclase and the GDP-bound form of the alpha subunit of transducin, inhibited the muscarinic M(1) facilitatory effect. Moreover, like acetylcholine (ACh), exogenously added betagamma subunits of transducin potentiated the CRH-stimulated adenylyl cyclase activity, and this effect was not additive with that elicited by ACh. Western blot analysis indicated the expression in frontal cortex of both type II and type IV adenylyl cyclases, two isoforms stimulated by betagamma subunits in synergism with activated G(s). The M(1) receptor-induced enhancement of the adenylyl cyclase response to CRH was counteracted by the G(q/11) antagonist GpAnt-2A but not by GpAnt-2, a preferential G(i/o) antagonist. In addition, the muscarinic facilitatory effect was inhibited by membrane preincubation with antiserum directed against the C terminus of the alpha subunit of G(q/11), whereas the same treatment with antiserum against either G(i1/2) or G(o) was without effect. These data indicate that in membranes of rat frontal cortex, activation of muscarinic M(1) receptors potentiates CRH-stimulated adenylyl cyclase activity through betagamma subunits of G(q/11). PMID- 10854267 TI - Evidence for expression of heteromeric serotonin 5-HT(3) receptors in rodents. AB - The gene and cDNAs that encode a novel subunit of rodent serotonin 5-HT(3) receptors were isolated from mouse and rat tissues. Each of the new rodent subunits shares 40% amino acid identity with the rat 5-HT(3A) subunit and 73% identity with the human 5-HT(3B) subunit. Despite a relatively low level of structural conservation, sequence analysis and functional studies suggest that the new rodent subunits are orthologues of the human 5-HT(3B) subunit. In common with homologous human receptors, rat heteromeric 5-HT(3) receptors displayed a substantially larger single-channel conductance than homomeric 5-HT(3A) receptors. In addition, the rat heteromeric receptors were less sensitive to antagonism by tubocurarine. However, in contrast to human heteromeric receptors, those of the rat displayed pronounced inward rectification of both the whole-cell and single-channel current amplitudes. Transcripts of the mouse 5-HT(3A) and 5 HT(3B) subunits are coexpressed in several cell lines that possess endogenous 5 HT(3) receptors. In addition, treatment of rat PC12 cells with nerve growth factor induced expression of both subunit mRNAs, with a similar time course for accumulation of each transcript. The combination of functional data and expression patterns is consistent with the existence of heteromeric 5-HT(3) receptors in rodent neurons. PMID- 10854268 TI - Dopamine D(1) receptor-induced gene transcription is modulated by DARPP-32. AB - The role of the dopamine- and cyclic AMP-regulated phosphoprotein of M(r) 32,000 (DARPP-32) in dopaminergic regulation of gene transcription in striatum and globus pallidus was examined. Mice with targeted disruption of the gene encoding DARPP-32, its homologue, inhibitor-1, or both, were used. Pharmacological characterization showed that mutant mice had normal basal levels of dopamine D(1) and D(2) receptors and adenosine A(2A) receptors. Basal expression levels of the striatonigral-specific neuropeptides substance P and prodynorphin and the immediate early genes c-fos and NGFI-A were also unaltered in mutant mice. A full D(1) receptor agonist, SKF 82958, up-regulated the expression of these neuropeptides and immediate early genes significantly more in wild-type mice than in mice lacking DARPP-32. Moreover, the additive stimulation of SKF 82958 and quinelorane, a D(2) receptor agonist, on c-fos mRNA in globus pallidus was significantly decreased in DARPP-32 and DARPP-32/I-1 knockout mice. No changes in dopamine receptor-induced gene expression were found in I-1 knockout mice. These results demonstrate an important involvement of DARPP-32 in dopamine receptor mediated regulation of gene expression both in striatal neurons, which are enriched in DARPP-32, and in pallidal neurons, which do not contain DARPP-32. PMID- 10854269 TI - Region-specific enhancement of basal extracellular and cocaine-evoked dopamine levels following constitutive deletion of the Serotonin(1B) receptor. AB - The behavioral effects of cocaine are enhanced following constitutive deletion of the serotonin(1B) receptor. The neural substrates mediating the enhanced response to cocaine are unknown. The present studies determined whether basal dopamine dynamics or cocaine-evoked dopamine levels are altered in projection areas of mesostriatal or mesoaccumbens dopamine neurons following serotonin(1B) receptor deletion. Male wild-type and serotonin(1B) knockout mice were implanted with microdialysis guide cannulas aimed at the dorsal striatum or nucleus accumbens. The zero net flux method of quantitative microdialysis was used to quantify basal extracellular dopamine concentrations (DA(ext)) and the extraction fraction of dopamine (E(d)), which provides an index of dopamine uptake. Conventional microdialysis techniques were used to quantify cocaine (0, 5.0, and 20.0 mg/kg) evoked dopamine overflow. Basal DA(ext) and E(d) did not differ in striatum of wild-type and knockout mice. Similarly, cocaine-stimulated dopamine overflow did not differ between genotype. The basal E(d) did not differ in the nucleus accumbens of wild-type and knockout mice. However, DA(ext) was significantly elevated in the nucleus accumbens of knockout mice. Cocaine-evoked dopamine overflow (nM) was also enhanced in the nucleus accumbens of knockout mice. However, the cocaine-induced increase in dopamine levels, relative to basal values, did not differ between genotype. These data demonstrate that deletion of the serotonin(1B) receptor is associated with increases in basal DA(ext) in the nucleus accumbens. This increase is not associated with an alteration in E(d), suggesting increased basal dopamine release in these animals. It is hypothesized that these alterations in presynaptic neuronal activity are a compensatory response to constitutive deletion of the serotonin(1B) receptor and may contribute to the enhanced behavioral effects of psychostimulants observed in knockout mice. PMID- 10854270 TI - Metallothionein-III prevents glutamate and nitric oxide neurotoxicity in primary cultures of cerebellar neurons. AB - Metallothionein (MT)-III, a member of the MT family of metal-binding proteins, is mainly expressed in the CNS and is abundant in glutamatergic neurons. Results in genetically altered mice indicate that MT-III may play neuroprotective roles in the brain, but the mechanisms through which this protein functions have not been elucidated. The aim of this work was to assess whether MT-III is able to prevent glutamate neurotoxicity and to identify the step of the neurotoxic process interfered with by MT-III. Glutamate neurotoxicity in cerebellar neurons in culture is mediated by excessive activation of glutamate receptors, increased intracellular calcium, and increased nitric oxide. It is shown that MT-III prevented glutamate- and nitric oxide-induced neurotoxicity in a dose-dependent manner, with nearly complete protection at 0.3-1 microgram/ml. MT-III did not prevent the glutamate-induced rise of intracellular calcium level but reduced significantly the nitric oxide-induced formation of cyclic GMP. Circular dichroism analysis revealed that nitric oxide triggers the release of the metals coordinated to the cysteine residues of MT-III, indicative of the S(Cys) nitrosylation of the protein. Therefore, the present results indicate that MT-III can quench pathological levels of nitric oxide, thus preventing glutamate and nitric oxide neurotoxicity. PMID- 10854271 TI - Phospholipase C, protein kinase C, Ca(2+)/calmodulin-dependent protein kinase II, and tyrosine phosphorylation are involved in carbachol-induced phospholipase D activation in Chinese hamster ovary cells expressing muscarinic acetylcholine receptor of Caenorhabditis elegans. AB - Recently, we have isolated a cDNA encoding a muscarinic acetylcholine receptor (mAChR) from Caenorhabditis elegans. To investigate the regulation of phospholipase D (PLD) signaling via a muscarinic receptor, we generated stable transfected Chinese hamster ovary (CHO) cells that overexpress the mAChR of C. elegans (CHO-GAR-3). Carbachol (CCh) induced inositol phosphate formation and a significantly higher Ca(2+) elevation and stimulated PLD activity through the mAChR; this was insensitive to pertussis toxin, but its activity was abolished by the phospholipase C (PLC) inhibitor U73122. Western blot analysis revealed several apparent tyrosine-phosphorylated protein bands after CCh treatment. The CCh-induced PLD activation and tyrosine phosphorylation were significantly reduced by the protein kinase C (PKC) inhibitor calphostin C and down-regulation of PKC and the tyrosine kinase inhibitor genistein. Moreover, the Ca(2+) calmodulin-dependent protein kinase II (CaM kinase II) inhibitor KN62, in addition to chelation of extracellular or intracellular Ca(2+) by EGTA and BAPTA/AM, abolished CCh-induced PLD activation and protein tyrosine phosphorylation. Taken together, these results suggest that the PLC/PKC-PLD pathway and the CaM kinase II/tyrosine kinase-PLD pathway are involved in the activation of PLD through mAChRs of C. elegans. PMID- 10854272 TI - The postsynaptic density protein PSD-95 differentially regulates insulin- and Src mediated current modulation of mouse NMDA receptors expressed in Xenopus oocytes. AB - The NMDA subtype of glutamate receptor is physically associated with the postsynaptic density protein PSD-95 at glutamatergic synapses. The channel activity of NMDA receptors is regulated by different signaling molecules, including protein tyrosine kinases. Because previous results have suggested a role for protein kinase C (PKC) in insulin potentiation of NMDA currents in oocytes, the effects of coexpression of PSD-95 on insulin and PKC potentiation of NMDA currents from these receptors were compared. Another primary objective was to determine if PSD-95 could enable Src to potentiate currents from NR2A/NR1 and NR2B/NR1 receptors expressed in Xenopus oocytes. The results show opposite effects of PSD-95 coexpression on Src and insulin modulation of NR2A/NR1 receptor currents. Src potentiation of mouse NR2A/NR1 currents required PSD-95 coexpression. In contrast, PSD-95 coexpression eliminated insulin-mediated potentiation of NR2A/NR1 receptor currents. PSD-95 coexpression also eliminated PKC potentiation of NR2A/NR1 receptor currents. PSD-95 may therefore play a key role in controlling kinase modulation of NR2A/NR1 receptor currents at glutamatergic synapses. PMID- 10854273 TI - Ionotropic glutamate receptors trigger microvesicle-mediated exocytosis of L glutamate in rat pinealocytes. AB - Rat pinealocytes receive noradrenergic innervation that stimulates melatonin synthesis. Besides melatonin, we showed previously that pinealocytes accumulate L glutamate in microvesicles and secrete it through an exocytic mechanism. The secreted glutamate binds to the class II metabotropic glutamate receptor and inhibits norepinephrine-stimulated melatonin synthesis in neighboring pinealocytes through an inhibitory cyclic AMP cascade. In this study, it was found that, in addition to metabotropic receptors, pinealocytes express functional ionotropic receptors. RT-PCR and northern analyses indicated the expression of mRNA for GluR1, KA2, and NR2C in pineal gland. The presence of GluR1 protein was confirmed by immunological techniques, but neither KA2 nor NR2C was detected. Consistent with this observation, the presence of (RS)-alpha-amino 3-hydroxy-5-methyl-4-isoxazolepropionic acid or kainate, non-N-methyl-D-aspartate receptor agonists, transiently stimulated increased the intracellular Ca(2+) concentration of cultured pinealocytes, whereas N-methyl-D-aspartate did not. These responses were prevented by 6-cyano-7-nitroquinoxaline-2,3-dione, a selective antagonist for non-N-methyl-D-aspartate receptors, by L-type Ca(2+) channel blockers such as nifedipine, or by omitting Ca(2+) or Na(+) in the medium. In the presence of Ca(2+) and Na(+), (RS)-alpha-amino-3-hydroxy-5-methyl 4-isoxazolepropionic acid or kainate evoked glutamate secretion from the cultured cells, which was prevented by 6-cyano-7-nitroquinoxaline-2,3-dione, L-type Ca(2+) channel blockers, type E or B botulinum neurotoxin, or incubation at <20 degrees C. These results strongly suggest that GluR1 is functionally expressed in pinealocytes and triggers microvesicle-mediated exocytosis of L-glutamate via activation of L-type Ca(2+) channels. It is possible that GluR1 participates in a signaling cascade that enhances and expands the L-glutamate signal throughout the pineal gland. PMID- 10854274 TI - Epidermal growth factor induces oxidative neuronal injury in cortical culture. AB - Recently, we have demonstrated that certain neurotrophic factors can induce oxidative neuronal necrosis by acting at the cognate tyrosine kinase-linked receptors. Epidermal growth factor (EGF) has neurotrophic effects via the tyrosine kinase-linked EGF receptor (EGFR), but its neurotoxic potential has not been studied. Here, we examined this possibility in mouse cortical culture. Exposure of cortical cultures to 1-100 ng/ml EGF induced gradually developing neuronal death, which was complete in 48-72 h; no injury to astrocytes was noted. Electron microscopic findings of EGF-induced neuronal death were consistent with necrosis; severe mitochondrial swelling and disruption of cytoplasmic membrane occurred, whereas nuclei appeared relatively intact. The EGF-induced neuronal death was accompanied by increased free radical generation and blocked by the anti-oxidant Trolox. Suggesting mediation by the EGFR, an EGFR tyrosine kinase specific inhibitor, C56, attenuated EGF-induced neuronal death. In addition, inhibitors of extracellular signal-regulated protein kinase 1/2 (Erk-1/2) (PD98056), protein kinase A (H89), and protein kinase C (GF109203X) blocked EGF induced neuronal death. A p38 mitogen-activated protein kinase inhibitor (SB203580) or glutamate antagonists (MK-801 and 6-cyano-7-nitroquinoxaline-2,3 dione) showed no protective effect. The present results suggest that prolonged activation of the EGFR may trigger oxidative neuronal injury in central neurons. PMID- 10854275 TI - Neurons overexpressing heme oxygenase-1 resist oxidative stress-mediated cell death. AB - This is the first report on the protective effect of heme oxygenase-1 (HO-1) overexpression against oxidative stress-mediated neuronal cell death and demonstration of a decreased production of oxygen free radicals when HO-1 levels are increased. HO-1 is the heat shock/stress cognate of the heat shock protein 32 family of proteins. A known function of these proteins is alpha-meso bridge specific cleavage of the heme molecule. For the present study, we used cerebellar granular neurons (CGNs) isolated from homozygous transgenic (Tg) mice that overexpress HO-1 under neuron-specific enolase control and nontransgenic (Ntg) littermates. The Tg mouse CGNs were characterized by increased levels of HO-1 mRNA and protein, a lower resting intracellular calcium concentration, and a reduced HO-1 transcriptional response to glutamate-mediated oxidative stress. Compared with the Ntg neurons, when exposed to glutamate (30 microM or 3 mM), the magnitude of cell viability was increased and the number of cells exhibiting membrane permeability and chromatin condensation were significantly decreased in the Tg CGN cultures. The population of neurons surviving glutamate toxicity decreased when HO-1 activity was inhibited by a peptide inhibitor. The neuroprotective effect by HO-1 was extended to H(2)O(2)-induced cell death. The mechanism of protection may involve in part a reduced production of reactive oxygen species upon exposure to glutamate. We suggest that induction of HO-1 by pharmacological means may be a novel approach to amelioration of oxidative insults to neurons. PMID- 10854276 TI - Beneficial effects of dietary restriction on cerebral cortical synaptic terminals: preservation of glucose and glutamate transport and mitochondrial function after exposure to amyloid beta-peptide, iron, and 3-nitropropionic acid. AB - Recent studies have shown that rats and mice maintained on a dietary restriction (DR) regimen exhibit increased resistance of neurons to excitotoxic, oxidative, and metabolic insults in experimental models of Alzheimer's, Parkinson's, and Huntington's diseases and stroke. Because synaptic terminals are sites where the neurodegenerative process may begin in such neurodegenerative disorders, we determined the effects of DR on synaptic homeostasis and vulnerability to oxidative and metabolic insults. Basal levels of glucose uptake were similar in cerebral cortical synaptosomes from rats maintained on DR for 3 months compared with synaptosomes from rats fed ad libitum. Exposure of synaptosomes to oxidative insults (amyloid beta-peptide and Fe(2+)) and a metabolic insult (the mitochondrial toxin 3-nitropropionic acid) resulted in decreased levels of glucose uptake. Impairment of glucose uptake following oxidative and metabolic insults was significantly attenuated in synaptosomes from rats maintained on DR. DR was also effective in protecting synaptosomes against oxidative and metabolic impairment of glutamate uptake. Loss of mitochondrial function caused by oxidative and metabolic insults, as indicated by increased levels of reactive oxygen species and decreased transmembrane potential, was significantly attenuated in synaptosomes from rats maintained on DR. Levels of the stress proteins HSP-70 and GRP-78 were increased in synaptosomes from DR rats, consistent with previous data suggesting that the neuroprotective mechanism of DR involves a "preconditioning" effect. Collectively, our data provide the first evidence that DR can alter synaptic homeostasis in a manner that enhances the ability of synapses to withstand adversity. PMID- 10854277 TI - Mechanisms of 5-aminolevulinic acid uptake at the choroid plexus. AB - 5-Aminolevulinic acid (5-ALA) is a precursor of porphyrins and heme that has been implicated in the neuropsychiatric symptoms associated with porphyrias. It is also being used clinically to delineate malignant gliomas. The blood-CSF barrier may be an important interface for 5-ALA transport between blood and brain as in vivo studies have indicated 5-ALA is taken up by the choroid plexuses whereas the normal blood-brain barrier appears to be relatively impermeable. This study examines the mechanisms of 5-[(3)H]ALA uptake into isolated rat lateral ventricle choroid plexuses. Results suggest that there are two uptake mechanisms. The first was a Na(+)-independent uptake system that was pH dependent (being stimulated at low pH). Uptake was inhibited by the dipeptide Gly-Gly and by cefadroxil, an alpha-amino-containing cephalosporin. These properties are the same as the proton dependent peptide transporters PEPT1 and PEPT2, which have recently been shown to transport 5-ALA in frog oocyte expression experiments. Choroid plexus uptake was not inhibited by captopril, a PEPT1 inhibitor, suggesting PEPT2-mediated uptake. The presence of PEPT2 and absence of PEPT1 in the choroid plexus were confirmed by western blotting. The second potential mechanism was both Na(+) and HCO(3)(-) dependent and appears to be an organic anion transporter, although it is possible that removal of Na(+) and HCO(3)(-) may indirectly affect PEPT2 by affecting intracellular pH. The presence of PEPT2 and a putative Na(+)/HCO(3)(-)-dependent organic anion transporter is important not only for an understanding of 5-ALA movement between blood and brain but also because these transporters may affect the distribution of a number of drugs between blood and CSF. PMID- 10854278 TI - Protein binding of NADH on chemical preconditioning. AB - Chemical preconditioning, an emerging neuroprotective strategy described in recent years, results in preserved energy metabolism during hypoxia via yet unknown mechanisms. The hypoxic increase of NADH content is attenuated by preconditioning. The goal of the present study was to investigate whether attenuation of the hypoxic NADH increase is due to a shift between free and protein-bound NADH. NADH in solution has a fluorescence maximum at 469.2 nm. In untreated mouse hippocampal slices, lambda(control onset) is 456.2 +/- 5.3 nm in CA1 (mean +/- SD; p < 0.01 vs. solution) and 454.6 +/- 6.1 nm in CA3 [p < 0.01 vs. solution, not significant (NS) to lambda(control onset) in CA1]. In slices prepared from animals pretreated in vivo with 20 mg/kg 3-nitropropionate, lambda(preconditioning onset) is 439.2 +/- 5.0 nm (p < 0.001 vs. control) in CA1 and 434.2 +/- 6.4 nm in CA3 (p < 0.001 vs. control; NS to lambda(preconditioning onset) in CA1). In controls, the fluorescence maximum shifts to lambda(control hypoxia) 458.2 +/- 1.3 nm in CA1 (NS vs. onset) and 456.0 +/- 3.6 nm in CA3 (NS vs. onset). On preconditioning with 3-nitropropionate, lambda(preconditioning hypoxia) shifts to 446.4 +/- 4.3 nm in CA1 (p < 0.03 vs. onset) and 438.6 +/- 6.9 nm in CA3 (p < 0.03 vs. onset). Posthypoxic decay of free and protein-bound NADH is diminished after preconditioning. We conclude that the free NADH level is reduced on an increase of hypoxic tolerance by chemical preconditioning. Reduction of free NADH content is maintained during hypoxia after preconditioning. PMID- 10854279 TI - Trimerization of cell adhesion molecule L1 mimics clustered L1 expression on the cell surface: influence on L1-ligand interactions and on promotion of neurite outgrowth. AB - Several studies indicate that cell adhesion molecules have to be clustered on the cell surface to engage in adhesive functions. We investigated adhesive functions of clustered versus monomeric L1 extracellular parts in vitro to distinguish how clustering affects ligand binding and promotion of neurite outgrowth. Trimeric L1 was recombinantly expressed and covalently assembled by the cartilage matrix protein's coiled-coil domain. Trimeric L1 has an apparent molecular mass of approximately 380 kDa in the nonreduced form and approximately 130 kDa in the reduced form. Rotary shadowing electron micrographs of trimeric L1 revealed a rod like shape terminating in three globular domains. Monomeric L1 assumes a horseshoe shape of domains Ig I-IV followed by a rod-like structure consisting of Ig V and VI and fibronectin type III 1-5. Circular dichroism measurements showed that the secondary structure consists of beta-sheets. Trimeric L1 binds to itself, to monomeric L1, to laminin-1, and to alpha5beta1 integrin in a concentration-dependent manner. In contrast, binding of monomeric L1 could only be saturated with itself but not with laminin-1 and with alpha5beta1 integrin. Promotion of neurite outgrowth from PC12 cells cultured on adsorbed trimeric L1 was increased by 100%, whereas on monomeric L1 the increase was only 50% over the control value. Promotion of neurite outgrowth from PC12 cells was specifically inhibited in a concentration-dependent manner by a polyclonal antibody against L1. These findings show that clustering of only three extracellular domains increases considerably L1's binding affinity to different ligands and enhances neurite outgrowth, suggesting that adhesive functions of L1 on the cell surface depend on cluster formation. PMID- 10854280 TI - Tyrosine phosphorylation of PNS myelin P(0) occurs in the cytoplasmic domain and is maximal during early development. AB - P(0), the major protein of PNS myelin, is considered to play a critical role in the compaction and stabilization of myelin lamellae. The protein undergoes extensive posttranslational modifications, including phosphorylation at multiple serine moieties in the cytoplasmic region. Recently, we demonstrated that P(0) is phosphorylated on one or more tyrosine residues in rat nerve homogenates after incubation. In this study, we show that P(0) phosphorylated on tyrosine is also present in the intact animal. The proportion of P(0) molecules phosphorylated on tyrosine is highest during the first postnatal week, a period that coincides with the most rapid period of myelin deposition in the PNS. A peptide that constitutes the cytoplasmic domain was isolated from purified P(0) and shown by immunochemical and chemical means to be phosphorylated on the tyrosine corresponding to Y(191) in the intact protein. No evidence was obtained supporting the possibility that P(0) is phosphorylated on other tyrosine residues. The sequence of amino acids surrounding Y(191) resemble known substrate phosphorylation sites for some nonreceptor cytoplasmic tyrosine kinases, as well as tyrosine-based recognition signals associated with clathrin vesicle-mediated cndocytosis. PMID- 10854281 TI - Light-mediated activation of diacylglycerol kinase in rat and bovine rod outer segments. AB - The hydrolysis of phosphatidylinositol 4,5-bisphosphate is regulated by light in retinal rod outer segment (ROS) membranes. We recently reported that the activities of phosphatidylinositol synthetase and phosphatidylinositol 3-kinase are also higher in bleached (light-exposed) ROS (B-ROS). In this study, we investigated the effect of bleaching on diacylglycerol (DAG) kinase (DAG-kinase) activity in bovine and rat ROS membranes prepared from dark-adapted (D-ROS) or bleached (B-ROS) retinas. In bovine ROS, DAG-kinase activity toward endogenous DAG substrate was higher in B-ROS than in D-ROS. Quantification of DAG in both sets of membranes showed that the levels were the same, eliminating the possibility that the greater DAG-kinase activity was due to higher levels of endogenous substrate in B-ROS. DAG-kinase activity was also higher in B-ROS against an exogenous, water-soluable substrate (1, 2-didecanoyl-rac-glycerol), which competed with endogenous DAG substrate and saturated at approximately 2 mM. Immunoblot analysis with an anti-DAG-kinase gamma polyclonal antibody demonstrated that the gamma isoform was present in isolated bovine ROS. Immunocytochemistry of frozen bovine retinal sections confirmed the presence of DAG-kinase gamma immunoreactivity in ROS, as well as other retinal cells. Quantification of the immunoreactive products on western blots showed that more DAG-kinase gamma was present in B-ROS than in D-ROS. In an in vivo experiment, ROS prepared from rats exposed to 30 min of room light had greater DAG-kinase activity than ROS prepared from dark-adapted animals. Taken together, these data suggest that light exposure leads to the translocation of DAG-kinase from the cytosol to ROS membranes and that the greater DAG-kinase activity in B-ROS is due to the presence of more protein associated with ROS membranes. PMID- 10854282 TI - Heat shock transcription factors and the hsp70 induction response in brain and kidney of the hyperthermic rat during postnatal development. AB - Heat shock transcription factor (HSF) 1 levels increase in brain regions and decline in kidney during postnatal rat development. In both neonatal and adult rats, levels of HSF1 protein in brain and kidney are proportional to the levels of HSF DNA-binding activity and the magnitude of heat shock protein hsp70 induction after thermal stress. There appears to be more HSF1 protein in adult brain than is needed for induction of hsp70 after thermal stress, suggesting that HSF1 may have other functions in addition to its role as a stress-inducible activator of heat shock genes. HSF2 protein levels decline during postnatal rat development in brain regions and kidney. Gel mobility shift analysis shows that HSF2 is not in a DNA-binding form in the neonatal brain and kidney, suggesting that HSF2 may not be involved in the constitutive expression of hsps in early postnatal development. There is no apparent relationship between levels of HSF2 protein and basal levels of hsp90, hsp70, heat shock cognate protein hsc70, and hsp60. PMID- 10854283 TI - Site-specific phosphorylation of neurofilament-L is mediated by calcium/calmodulin-dependent protein kinase II in the apical dendrites during long-term potentiation. AB - Neurofilament-L (NF-L), one subunit of the neuronal intermediate filaments, is a major element of neuronal cytoskeletons. The dynamics of NF-L are regulated by phosphorylation of its head domain. The phosphorylation sites of the NF-L head domain by protein kinase A, protein kinase C, and Rho-associated kinase have been previously identified, and those by calcium/calmodulin-dependent protein kinase II (CaMKII) were identified in this study. A series of site- and phosphorylation state-specific antibodies against NF-L was prepared to investigate NF-L phosphorylation in neuronal systems. Long-term potentiation (LTP) is a cellular model of neuronal plasticity that is thought to involve the phosphorylation of various proteins. NF-L is considered a possible substrate for phosphorylation. During LTP stimulation of mouse hippocampal slices, the series of antibodies demonstrated the increase in the phosphorylation level of Ser(57) in NF-L and the visualization of the localized distribution of Ser(57) phosphorylation in a subpopulation of apical dendrites of the pyramidal neurons. Furthermore, Ser(57) phosphorylation during LTP is suggested to be mediated by CaMKII. Here we show that NF-L is phosphorylated by CaMKII in a subpopulation of apical dendrites during LTP, indicating that Ser(57) is a novel phosphorylation site of NF-L in vivo related to the neuronal signal transduction. PMID- 10854284 TI - Immunocytochemical characterization of the mitochondrially encoded ND1 subunit of complex I (NADH : ubiquinone oxidoreductase) in rat brain. AB - In Parkinson's disease, there is a selective defect in complex I of the electron transfer chain. To better understand complex I and its involvement in neurodegenerative disease, we raised an antibody against a conserved epitope of the human mitochondrially encoded subunit 1 of complex I (ND1). Antibodies were affinity purified and assessed by ELISA, immunoblotting, and immunocytochemistry. Immunoblots of brain homogenates from mouse, rat, and monkey brain showed a single 33-kDa band consistent with the predicted molecular mass of the protein. Subcellular fractionation showed the protein to be enriched in mitochondria. Immunocytochemistry in rat brain revealed punctate labeling in cell bodies and processes of neurons. Immunoreactively generally co-localized with subunit IV of complex IV. In striatum, ND1 immunoreactively was greatly enriched in large cholinergic neurons and neurons containing nitric oxide synthase, two cell populations that are resistant to excitotoxic and metabolic insults. In substantia nigra, many dopaminergic neurons had little ND1 immunoreactivity, which may help to explain their sensitivity to complex I inhibitors. In spinal cord, ND1 immunoreactively was enriched in motor neurons. We conclude that complex I is differentially distributed across brain regions, between neurons and glia, and between types of neurons. This antibody should provide a valuable tool for assessing complex I in normal and pathological conditions. PMID- 10854285 TI - Differential expression of the G protein beta(5) gene: analysis of mouse brain, peripheral tissues, and cultured cell lines. AB - A neurally expressed heterotrimeric G protein beta subunit, Gbeta(5), has been found to exhibit functional specialization with respect to its interactions with effector targets and Galpha subunits. A splice variant of Gbeta(5) that contains an N-terminal 42-residue extension, Gbeta(5)-long, has been described in the retina. To define better the potential range of its specialized interactions, analysis of Gbeta(5) gene transcript and protein expression in mouse brain and other tissues and cell lines was performed. Quantification by ribonuclease protection assay of Gbeta(5) transcript expression in the developing brain demonstrates a fivefold increase that occurs postnatally. Analysis of transcript expression by in situ hybridization and ribonuclease protection assay indicates that the Gbeta(5) gene is differentially expressed among multiple adult mouse brain regions, including the motor and occipital cortex, the olfactory bulb and associated rhinencephalic structures, hypothalamus, pontine cochlear nuclei, and Purkinje cells in the cerebellum. Gbeta(5) is also expressed in several cultured cell lines of neuroendocrine origin, including murine alphaT3-1 pituitary gonadotrophs and GT1-7 hypothalamic cells, and rat PC12 pheochromocytoma cells. Immunoblotting of tissue homogenates with antibodies to two peptides common to Gbeta(5) and Gbeta(5)-long confirmed expression of Gbeta(5) in multiple brain regions and in spinal cord and expression of Gbeta(5)-long in retina. Taken together, these results suggest that the specialized molecular properties of Gbeta(5) have been adapted to diverse neural functions in the adult brain. PMID- 10854286 TI - High-affinity anti-ganglioside IgG antibodies raised in complex ganglioside knockout mice: reexamination of GD1a immunolocalization. AB - Gangliosides, sialic acid-bearing glycosphingolipids, are highly enriched in the vertebrate nervous system. Anti-ganglioside antibodies are associated with various human neuropathies, although the pathogenicity of these antibodies remains unproven. Testing the pathogenic role of anti-ganglioside antibodies will be facilitated by developing high-affinity IgG-class complement-fixing monoclonal anti-bodies against major brain gangliosides, a goal that has been difficult to achieve. In this study, mice lacking complex gangliosides were used as immune naive hosts to raise anti-ganglioside antibodies. Wild-type mice and knockout mice with a disrupted gene for GM2/GD2 synthase (UDP-N-acetyl-D-galactosamine : GM3/GD3 N-acetyl-D-glactosaminyltransferase) were immunized with GD1a conjugated to keyhole limpet hemocyanin. The knockout mice produced a vigorous anti-GD1a IgG response, whereas wildtype littermates failed to do so. Fusion of spleen cells from an immunized knockout mouse with myeloma cells yielded numerous IgG anti GD1a antibody-producing colonies. Ganglioside binding studies revealed two specificity classes; one colony representing each class was cloned and characterized. High-affinity monoclonal antibody was produced by each hybridoma : an IgG1 that bound nearly exclusively to GD1a and an IgG2b that bound GD1a, GT1b, and GT1aalpha. Both antibodies readily readily detected gangliosides via ELISA, TLC immune overlay, immunohistochemistry, and immunocytochemistry. In contrast to prior reports using anti-GD1a and anti-GT1b IgM class monoclonal antibodies, the new antibodies bound avidly to granule neurons in brain tissue sections and cell cultures. Mice lacking complex gangliosides are improved hosts for raising high affinity, high-titer anti-ganglioside IgG antibodies for probing for the distribution and physiology of gangliosides and the pathophysiology of anti ganglioside antibodies. PMID- 10854287 TI - Behaviroal, pharmacological, and molecular characterization of an amphibian cannabinoid receptor. AB - Investigation of cannabinoid pharmacology in a vertebrate with a phylogenetic history distinct from that of mammals may allow better understanding of the physiological significance of cannabinoid neurochemistry. Taricha granulosa, the roughskin newt, was used here to characterize an amphibian cannabinoid receptor. Behavioral experiments demonstrated that the cannabinoid agonist levonantradol inhibits both newt spontaneous locomotor activity and courtship clasping behavior. Inhibition of clasping was dose-dependent and potent (IC(50) = 1.2 microgram per animal). Radioligand binding studies using [(3)H]CP-55940 allowed identification of a specific binding site (K(D) = 6.5 nM, B(max) = 1,853 fmol/mg of protein) in brain membranes. Rank order of affinity of several ligands was consistent with that reported for mammalian species (K(D), nM) : CP-55940 (3.8) > levonantradol (13.0) > WIN55212-2 (25.7) >> anandamide (1,665) approximately anandamide 100 microM phenylmethylsulfonyl fluoride (2,398). The cDNA encoding the newt CB1 cannabinoid receptor was cloned, and the corresponding mRNA of 5.9 kb was found to be highly expressed in brain. A nonclonal Chinese hamster ovary cell line stably expressing the newt CB1 cannabinoid receptor was prepared that allowed demonstration of cannabinoid-mediated inhibition of adenylate cyclase (EC 4.6.1.1) activity. This inhibition was dose-dependent and occurred at concentrations consistent with affinities determined through radioligand binding experiments. The behavioral, pharmacological, and molecular cloning results demonstrate that a CB1 cannabinoid receptor is expressed in the CNS of the roughskin newt. This amphibian CB1 is very similar in density, ligand binding affinity, ligand binding specificity, and amino acid sequence to mammalian CB1. The high degree of evolutionary conservation of cannabinoid signaling systems implies an important physiological role in vertebrate brain function. PMID- 10854288 TI - Assessment of stereoselectivity of trimethylphenylalanine analogues of delta opioid [D-Pen(2),D-Pen(5)]-enkephalin. AB - [D-Pen(2),D-Pen(5)]-Enkephalin (DPDPE) is an enzymatically stable delta-opioid receptor-selective peptide, which was modified by the trimethylation of the Phe(4) residue to give beta-methyl-2', 6'-dimethylphenylalanine (TMP), resulting in four conformations : (2R,3S)-beta-Phe-DPDPE, (2R,3R)-beta-Phe-DPDPE, (2R, 3S) beta-Phe-DPDPE, and (2S,3R)-beta-Phe-DPDPE. Synthesis was by solid-phase techniques using enantiomerically pure amino acids to give the four optically pure diastereoisomer peptides. The potency and selectivity (delta- versus mu opioid receptor) were evaluated by radioreceptor binding in rat brain, with a mu/delta ratio decrease for all TMP conformations, compared with the parent compound (DPDPE). Octanol/buffer distribution analysis showed enhanced lipophilicity of all TMP forms, with a sixfold enhancement associated with (2S,3S)-TMP. In situ vascular perfusion in anesthetized rats showed a 1.6-fold (p < 0.01) increase in the ratio of brain uptake for (2S,3S)-TMP and a 1.5-fold (p < 0.01) decrease in uptake for (2R,3R)-TMP. Saturability of (2S,3S)-TMP was shown (p < 0.01) against 100 microM unlabeled DPDPE, showing a shared nondiffusionary transport system. P-glycoprotein affinity was shown in situ for the parent and (2S,3S)-TMP (p < 0.01). Protein binding capacity of the TMP compounds in rat plasma and in situ mammalian bovine serum albumin-Ringer showed (2R,3S)-TMP and (2S,3R)-TMP with the lowest degree of protein binding (p < 0.01), and (2S,3S)-TMP and (2R,3R)-TMP with comparable affinities to DPDPE. Analgesia, via intravenous administration, showed significantly reduced (p < 0.01) end effect and time course for (2R,3R)-TMP, (2R,3S)-TMP, and (2S, 3R)-TMP as compared with DPDPE. These results demonstrate that topographical modification in a conformationally restricted peptide can significantly modulate potency and receptor selectivity, binding capacity, enzymatic stability, lipophilicity, P-glycoprotein affinity, and blood-brain barrier permeability, resulting in a change of bioavailability, and thereby provides insight for future peptide drug design. PMID- 10854289 TI - Impaired proteasome function in Alzheimer's disease. AB - Inhibition of proteasome activity is sufficient to induce neuron degeneration and death; however, altered proteasome activity in a neurodegenerative disorder has not been demonstrated. In the present study, we analyzed proteasome activity in short-postmortem-interval autopsied brains from 16 Alzheimer's disease (AD) and nine age- and sex-matched controls. A significant decrease in proteasome activity was observed in the hippocampus and parahippocampal gyrus (48%), superior and middle temporal gyri (38%), and inferior parietal lobule (28%) of AD patients compared with controls. In contrast, no significant decrease in proteasome activity was observed in either the occipital lobe or the cerebellum. The loss of proteasome activity was not associated with a decrease in proteasome expression, suggesting that the proteasome may become inhibited in AD by a posttranslational modification. Together, these data indicate a possible role for proteasome inhibition in the neurodegeneration associated with AD. PMID- 10854313 TI - Bullying: children hurting children. PMID- 10854314 TI - Infant feeding. PMID- 10854315 TI - Syncope. PMID- 10854317 TI - More Lessons for the Clinician. PMID- 10854316 TI - Index of suspicion. Case 1. Organophosphate intoxication. PMID- 10854318 TI - An infant who has head trauma. PMID- 10854319 TI - Earning CME Credit-Completing the PIR Quiz. PMID- 10854320 TI - Discrete interactions in cell adhesion measured by single-molecule force spectroscopy. AB - Cell-cell adhesion mediated by specific cell-surface molecules is essential for multicellular development. Here we quantify de-adhesion forces at the resolution of individual cell-adhesion molecules, by controlling the interactions between single cells and combining single-molecule force spectroscopy with genetic manipulation. Our measurements are focused on a glycoprotein, contact site A (csA), as a prototype of cell-adhesion proteins. csA is expressed in aggregating cells of Dictyostelium discoideum, which are engaged in development of a multicellular organism. Adhesion between two adjacent cell surfaces involves discrete interactions characterized by an unbinding force of 23 +/- 8 pN, measured at a rupture rate of 2.5 +/- 0.5 microm s-1. PMID- 10854321 TI - Changes in intramitochondrial and cytosolic pH: early events that modulate caspase activation during apoptosis. AB - Mitochondria trigger apoptosis by releasing caspase activators, including cytochrome c (cytC). Here we show, using a pH-sensitive green fluorescent protein (GFP), that mitochondria-dependent apoptotic stimuli (such as Bax, staurosporine and ultraviolet irradiation) induce rapid, Bcl-2-inhibitable mitochondrial alkalinization and cytosol acidification, followed by cytC release, caspase activation and mitochondrial swelling and depolarization. These events are not induced by mitochondria-independent apoptotic stimuli, such as Fas. Activation of cytosolic caspases by cytC in vitro is minimal at neutral pH, but maximal at acidic pH, indicating that mitochondria-induced acidification of the cytosol may be important for caspase activation; this finding is supported by results obtained from cells using protonophores. Cytosol acidification and cytC release are suppressed by oligomycin, a FoF1-ATPase/H +-pump inhibitor, but not by caspase inhibitors. Ectopic expression of Bax in wild-type, but not FoF1/H+-pump deficient, yeast cells similarly results in mitochondrial matrix alkalinization, cytosol acidification and cell death. These findings indicate that mitochondria mediated alteration of intracellular pH may be an early event that regulates caspase activation in the mitochondrial pathway for apoptosis. PMID- 10854322 TI - Dynamic interaction of BiP and ER stress transducers in the unfolded-protein response. AB - PERK and IRE1 are type-I transmembrane protein kinases that reside in the endoplasmic reticulum (ER) and transmit stress signals in response to perturbation of protein folding. Here we show that the lumenal domains of these two proteins are functionally interchangeable in mediating an ER stress response and that, in unstressed cells, both lumenal domains form a stable complex with the ER chaperone BiP. Perturbation of protein folding promotes reversible dissociation of BiP from the lumenal domains of PERK and IRE1. Loss of BiP correlates with the formation of high-molecular-mass complexes of activated PERK or IRE1, and overexpression of BiP attenuates their activation. These findings are consistent with a model in which BiP represses signalling through PERK and IRE1 and protein misfolding relieves this repression by effecting the release of BiP from the PERK and IRE1 lumenal domains. PMID- 10854323 TI - Cargo binding and regulatory sites in the tail of fungal conventional kinesin. AB - Here, using a quantitative in vivo assay, we map three regions in the carboxy terminus of conventional kinesin that are involved in cargo association, folding and regulation, respectively. Using C-terminal and internal deletions, point mutations, localization studies, and an engineered 'minimal' kinesin, we identify five heptads of a coiled-coil domain in the kinesin tail that are necessary and sufficient for cargo association. Mutational analysis and in vitro ATPase assays highlight a conserved motif in the globular tail that is involved in regulation of the motor domain; a region preceding this motif participates in folding. Although these sites are spatially and functionally distinct, they probably cooperate during activation of the motor for cargo transport. PMID- 10854324 TI - Nitric oxide upregulates expression of DNA-PKcs to protect cells from DNA damaging anti-tumour agents. AB - Nitric-oxide synthase (NOS) activity has been detected in many human tumours, although its function is unclear. Here we show that exposure of cells to nitric oxide (NO) results in a 4-5-fold increase in expression of the DNA-dependent protein-kinase catalytic subunit (DNA-PKcs), one of the key enzymes involved in repairing double-stranded DNA breaks. This NO-mediated increase in enzymatically active DNA-PK not only protects cells from the toxic effects of NO, but also provides crossprotection against clinically important DNA-damaging agents, such as X-ray radiation, adriamycin, bleomycin and cisplatin. The NO-mediated increase in DNA-PKcs described here demonstrates the presence of a new and highly effective NO-mediated mechanism for DNA repair. PMID- 10854325 TI - Tollip, a new component of the IL-1RI pathway, links IRAK to the IL-1 receptor. AB - Interleukin-1 (IL-1) is a proinflammatory cytokine that elicits its pleiotropic effects through activation of the transcription factors NF-kappaB and AP-1. Binding of IL-1 to its receptor results in rapid assembly of a membrane-proximal signalling complex that consists of two different receptor chains (IL-1Rs), IL 1RI and IL-1RAcP, the adaptor protein MyD88, the serine/threonine kinase IRAK and a new protein, which we have named Tollip. Here we show that, before IL-1beta treatment, Tollip is present in a complex with IRAK, and that recruitment of Tollip-IRAK complexes to the activated receptor complex occurs through association of Tollip with IL-1RAcP. Co-recruited MyD88 then triggers IRAK autophosphorylation, which in turn leads to rapid dissociation of IRAK from Tollip (and IL-1Rs). As overexpression of Tollip results in impaired NF-kappaB activation, we conclude that Tollip is an important constituent of the IL-1R signalling pathway. PMID- 10854326 TI - Immunostructural evidence for the template mechanism of microtubule nucleation. AB - Two opposing models have been proposed to explain how the gamma-tubulin ring complex (gammaTuRC) induces microtubule nucleation. In the 'protofilament' model, the gammaTuRC induces nucleation as a partially or completely straightened protofilament that is incorporated longitudinally into the wall of the nascent microtubule, whereas the 'template' model proposes that the gammaTuRC acts as a helical template that constitutes the base of the newly-formed polymer. Here we appraise these two models, using high-resolution structural and immunolocalization methods. We show that components of the gammaTuRC localize to a narrow zone at the extreme minus end of the microtubule and that these ends terminate in a pointed cap. Together, these results strongly favour the template model of microtubule nucleation. PMID- 10854327 TI - A new function for the gamma-tubulin ring complex as a microtubule minus-end cap. AB - Microtubule nucleation from centrosomes involves a lockwasher-shaped protein complex containing gamma-tubulin, named the gamma-tubulin ring complex (gammaTuRC). Here we investigate the mechanism by which the gammaTuRC nucleates microtubules, using a direct labelling method to visualize the behaviour of individual gammaTuRCs. A fluorescently-labelled version of the gammaTuRC binds to the minus ends of microtubules nucleated in vitro. Both gammaTuRC-mediated nucleation and binding of the gammaTuRC to preformed microtubules block further minus-end growth and prevent microtubule depolymerization. The gammaTuRC therefore acts as a minus-end-capping protein, as confirmed by electron microscopic examination of gold-labelled gammaTuRCs. These data support a nucleation model for gammaTuRC function that involves capping of microtubules. PMID- 10854328 TI - Structure of the gamma-tubulin ring complex: a template for microtubule nucleation. AB - The gamma-tubulin ring complex (gammaTuRC) is a protein complex of relative molecular mass approximately 2.2 x 10(6) that nucleates microtubules at the centrosome. Here we use electron-microscopic tomography and metal shadowing to examine the structure of isolated Drosophila gammaTuRCs and the ends of microtubules nucleated by gammaTuRCs and by centrosomes. We show that the gammaTuRC is a lockwasher-like structure made up of repeating subunits, topped asymmetrically with a cap. A similar capped ring is also visible at one end of microtubules grown from isolated gammaTuRCs and from centrosomes. Antibodies against gamma-tubulin label microtubule ends, but not walls, in centrosomes. These data are consistent with a template-mediated mechanism for microtubule nucleation by the gammaTuRC. PMID- 10854329 TI - Smooth-muscle contraction without smooth-muscle myosin. AB - Here we have used gene-targeting to eliminate expression of smooth-muscle myosin heavy chain. Elimination of this gene does not affect expression of non-muscle myosin heavy chain, and knockout individuals typically survive for three days. Prolonged activation, by KCl depolarisation, of intact bladder preparations from wild-type neonatal mice produces an initial transient state (phase 1) of high force generation and maximal shortening velocity, which is followed by a sustained state (phase 2) characterized by low force generation and maximal shortening velocity. Similar preparations from knockout neonatal mice do not undergo phase 1, but exhibit a normal phase 2. We propose that, in neonatal smooth muscle phase 1 is generated by recruitment of smooth-muscle myosin heavy chain, whereas phase 2 can be generated by activation of non-muscle myosin heavy chain. We conclude that phase 1 becomes indispensable for survival and normal growth soon after birth, particularly for functions such as homeostasis and circulation. PMID- 10854331 TI - Drowning by numbers. PMID- 10854330 TI - Latrunculin alters the actin-monomer subunit interface to prevent polymerization. PMID- 10854332 TI - Gamma-tubulin nucleation: template or protofilament? AB - Gamma-tubulin is known to nucleate microtubule assembly from alpha/beta-tubulin, but the molecular mechanism by which this process occurs is the subject of some controversy. Four recent papers have provided new structural and biochemical constraints on the models proposed for nucleation. These have refined, but not yet resolved, the debate. PMID- 10854333 TI - Convergence of DNA repair and cell-cycle checkpoint control. PMID- 10854334 TI - It's a kar9ochore to capture microtubules. AB - Microtubule orientation to cortical spatial cues is essential for the fidelity of asymmetric cellular processes. A cortical microtubule-capture site, composed of Bim1 and Kar9, has now been identified in yeast. Bim1 is the yeast homologue of EB1, a binding partner of the adenomatous polyposis coli (APC), indicating that important features of this complex may be highly conserved. PMID- 10854335 TI - Different site, different splice. AB - The MKK3/6-p38 pathway has been found to induce the relocalization of premessenger-RNA splicing factors from the nucleus to the cytoplasm. This represents the first physiological mechanism that alters the nuclear ratios of splicing factors and modulates alternative splice-site choice in vivo. PMID- 10854336 TI - Drosophila p53: meeting the Grim Reaper. AB - The recent discovery of a Drosophila orthologue of the p53 tumour suppressor promises new insights into the complex function, regulation and evolution of one of the most intensely studied human disease proteins. PMID- 10854337 TI - COP1 patrols the night beat. AB - Light regulates the behaviour of many organisms. New data indicate that the greening of plants is facilitated by light-dependent stabilization of a transcription factor that is rapidly degraded in darkness. Thus, photomorphogenesis joins cell division and circadian rhythm as another critical biological process that is governed by proteolysis. PMID- 10854338 TI - "Eat me" signals of apoptotic bodies. PMID- 10854339 TI - Giving protein traffic the green light. AB - Most proteins that are secreted or expressed on a cell surface are synthesized on membrane polysomes and enter the endoplasmic reticulum (ER) as unfolded polypeptide chains. A complex series of interactions with resident enzymes and molecular chaperones ensure that these proteins are folded and assembled to achieve their correct tertiary structures before being transported to the Golgi and along the secretory pathway. However, the mechanism by which properly folded molecules are sorted from incompletely or improperly folded proteins and from the resident proteins that guide this process remains unclear. PMID- 10854340 TI - To be or not to be in the nucleolus. AB - Compartmentalization has long been known to have a key role in regulation of cellular processes. By keeping enzymes and regulatory complexes in compartments where the delivery of substrate or exit of product is controlled, competing reactions can occur simultaneously in different parts of the cell. Moreover, spatial confinement facilitates the working of molecules participating in reaction chains and is crucial for coupling unfavourable with energetically favourable chemical reactions. Although in many cases intracellular compartmentalization relies on boundaries imposed by membranes, several non membrane-bounded compartments exist in eukaryotic cells. One of these, the nucleolus, has recently attracted much attention. The emerging view is that molecular confinement in the nucleolus actively contributes to the control of cellular survival and proliferation. PMID- 10854341 TI - Systemic drugs with antipruritic potency. AB - Despite the predominance of itch as a leading and distressing symptom in most of the dermatological and several systemic diseases, there is relatively little progress in understanding its pathophysiology. This is most likely the main reason for the limited number of satisfactory anti-itch treatments and the fact that even today various therapies have empirical but not evidence-based character. There are no specific antipruritic drugs on the market, but there are a high number of case reports and experimental investigations describing medications with antipruritic potency. It is therefore the aim of this article to briefly review the major systemic antipruritic drugs and give a short overview on the different types of pruritus and their possible systemic therapy. PMID- 10854342 TI - DPCP for the treatment of alopecia areata. AB - Topical immunotherapy with diphencyprone (DPCP) for the treatment of severe alopecia areata has been used since 1983 and is felt to be the treatment of choice by many dermatologists. Although there have been no major side effects reported since its initial use, there remain some unknowns regarding its safety. Because DPCP has at least a 40% success rate for cosmetically acceptable regrowth in extensive alopecia areata, its availability is an important matter for patients with alopecia areata. PMID- 10854343 TI - Introduction. Essential tremor. PMID- 10854344 TI - Diagnostic criteria for essential tremor and differential diagnosis. AB - Classic essential tremor (ET) is a condition in which the upper limbs (approximately 95% of patients) and, less commonly, the head (approximately 34%), face (approximately 5%), voice (approximately 12%), trunk (approximately 5%), and lower limbs (approximately 20%) exhibit a mixed postural and kinetic tremor without other neurologic abnormalities. Most patients with ET probably inherit the disease through an autosomal dominant gene, but the true ratio of hereditary versus sporadic ET is unknown. Isolated focal, position-specific, and task specific tremors are probably not ET in most patients and are often due to subtle dystonia. Unilateral tremor, gait disturbance, rigidity, bradykinesia, rest tremor, and rapid onset of symptoms are indications of other tremorogenic disorders. PMID- 10854345 TI - Criteria for the diagnosis of essential tremor. PMID- 10854346 TI - Epidemiology and genetics of essential tremor. AB - Essential tremor (ET) is probably the most common movement disorder and is a common cause of social, physical, and psychological handicaps. Its etiology and pathogenesis are unknown. Phenomenologically, ET overlaps and is associated with other disorders of movement, such as parkinsonism and dystonia. There is large variation in the stated prevalence of ET as well as limited availability of epidemiologic studies. Prevalence variations reflect differences in the definition of ET and the methodologies of investigation. The familial and sporadic forms of ET are generally assumed to be similar. The familial form appears to have a narrow phenotype. Wide variation in the reported percentage of patients with positive family history reflects ascertainment and classification differences. Linkage of ET to two different chromosome locations has been reported. PMID- 10854347 TI - The pathophysiology of essential tremor. AB - The pathophysiologic abnormalities that underlie essential tremor (ET) are difficult to decipher because autopsy studies reveal no gross or microscopic abnormalities. Electrophysiologic studies are consistent with a central source of tremorogenic oscillation. The inferior olive and cerebellum are implicated by PET studies. Harmaline tremor in animals shares many features with ET, and the inferior olive has been identified as the source of oscillation in this animal model. Therefore, a disturbance of olivocerebellar rhythmicity is at present the most popular hypothesis for the etiology of ET. Although electrophysiologic tests are available that are helpful in the diagnosis of ET, a gold-standard test or biologic marker for ET is still lacking. PMID- 10854348 TI - Essential tremor: clinical characteristics. AB - Essential tremor (ET) is the most common movement disorder. However, only a small percentage of people affected by this genetically transmitted neurologic disorder seek medical attention. Lack of consensus on the diagnostic criteria for ET is an impediment to accurate diagnosis and leads to difficulty in accessing accurate prevalence data. Although a positive family history, alcohol sensitivity, and propranolol responsiveness are characteristic of ET, these factors should not be considered necessary for the diagnosis of ET. ET can produce substantial physical and psychosocial disabilities. The occasional coexistence of ET and Parkinson's disease (PD) in the same individual may present a diagnostic challenge. PMID- 10854349 TI - Tremor assessment and quality of life measurements. AB - Essential tremor (ET) can be measured objectively by physiological techniques, simple tests of the tremor's impact on function, or subjective use of clinical rating scales. The methods of measuring ET and its influence on patients are reviewed. Multidimensional evaluations are recommended for the assessment of the severity of ET in clinical trials. The term "detractor" describes the relationships between ET and the disability and handicap that it produces. PMID- 10854350 TI - Pharmacologic treatment of essential tremor. AB - Essential tremor (ET) is a common movement disorder that often causes functional disability, potentially leading to physical and emotional difficulties. The paucity of data available regarding the underlying pathophysiologic mechanism of ET hinders the development of innovative approaches to pharmacotherapeutic treatments. Options for drug therapy include the use of primidone, beta adrenergic blockers, such as propranolol, alcohol, and other drugs, such as benzodiazepines, gabapentin, carbonic anhydrase inhibitors, clozapine, flunarizine, clonidine, and the methylxanthine derivative theophylline. Chemodenervation with botulinum toxin type A may be a therapeutic option for selected patients with ET. Each drug is classified as to the quality of evidence for efficacy and the suggested strength of therapeutic recommendation. In general clinical practice, primidone and propranolol have proven efficacy in ET. PMID- 10854351 TI - Surgical treatment of essential tremor. AB - Surgical treatment for essential tremor (ET) has been used since the early 1950s. Initially, different areas were targeted for tremor control. However, the optimal target was eventually determined to be the ventralis intermedius (VIM) nucleus of the thalamus. Thalamotomy improves contralateral tremor in more than 90% of patients. Long-term studies of thalamotomy indicate that the benefits continue in most patients. Persistent morbidity associated with thalamotomy, which occurs in less than 10% of patients, includes dysarthria, dysequilibrium, weakness, and cognitive impairment. Bilateral thalamotomy is associated with substantial morbidity and is usually avoided. Studies demonstrate that chronic stimulation of the VIM is safe and effective for tremor. Adverse effects of chronic stimulation include paresthesia, dysarthria, dysequilibrium, and localized pain. In many patients, bilateral thalamic stimulation is performed without a substantial increase in morbidity. ET patients with disabling medication-resistant tremor are reasonable candidates for these stereotactic procedures. PMID- 10854352 TI - Support organizations. WE MOVE and the International Tremor Foundation. Worldwide Education and Awareness for Movement Disorders. PMID- 10854353 TI - Frequency and impact of neurologic diseases in the elderly of Europe: A collaborative study of population-based cohorts. PMID- 10854354 TI - Prevalence of dementia and major subtypes in Europe: A collaborative study of population-based cohorts. Neurologic Diseases in the Elderly Research Group. AB - The last comparison of prevalence figures of dementia across European studies was 10 years ago. Using studies conducted in the 1990s, the authors compare the age- and sex-specific prevalence of dementia, AD, and vascular dementia (VaD) across European population-based studies of persons 65 years and older. Data from these studies were also pooled to obtain stable estimates of age- and sex-specific prevalence. A total of 2346 cases of mild to severe dementia were identified in 11 cohorts. Age-standardized prevalence was 6.4% for dementia (all causes), 4.4% for AD, and 1.6% for VaD. The prevalence of dementia increased continuously with age and was 0.8% in the group age 65 to 69 years and 28.5% at age 90 years and older. The corresponding figures for AD (53.7% of cases) were 0.6% and 22.2%, and for VaD (15.8% of cases), 0.3% and 5.2%. Variation of AD prevalence across studies was greatest for men. In the VaD subtype, a large variation across studies was observed, as well as a difference in prevalence between men and women that was age dependent. Dementia is more prevalent in women, and AD is the main contributor to the steep increase of prevalence with age. PMID- 10854355 TI - Incidence of dementia and major subtypes in Europe: A collaborative study of population-based cohorts. Neurologic Diseases in the Elderly Research Group. AB - The authors examined the association of incident dementia and subtypes with age, sex, and geographic area in Europe. Incidence data from eight population-based studies carried out in seven European countries were compared and pooled. The pooled data included 835 mild to severe dementia cases and 42,996 person-years of follow-up. In all studies a higher proportion of cases were diagnosed with AD (60 to 70% of all demented cases) than vascular dementia (VaD). The incidence of dementia and AD continued to increase with age up to age 85 years, after which rates increased in women but not men. There was a large variation in VaD incidence across studies. In the pooled analysis, the incidence rates increased with age without any substantial difference between men and women. Surprisingly, higher incidence rates of dementia and AD were found in the very old in northwest countries than in southern countries. This study confirms that AD is the most frequent dementing disorder in all ages, and that there is a higher incidence of dementia, specifically AD, in women than men among the very old. Finally, there may be regional differences in dementia incidence. PMID- 10854356 TI - Prognosis with dementia in Europe: A collaborative study of population-based cohorts. Neurologic Diseases in the Elderly Research Group. AB - The effect of dementia on time to death and institutionalization in elderly populations is of importance to resource planning, as well as to patients and their carers. The authors report a collaborative reanalysis of nine population based studies conducted in Europe to compare dementia cases and noncases in risk of and time to death and to institutionalization. Prevalent and incident cases were more likely than noncases to reside in an institution at baseline and were more likely to enter institutional care. Prevalent cases also had over twice the risk of death compared to noncases and survival for men with dementia was consistently lower than that for women with dementia of the same age group. PMID- 10854357 TI - Prevalence of Parkinson's disease in Europe: A collaborative study of population based cohorts. Neurologic Diseases in the Elderly Research Group. AB - The results of seven population-based studies were examined separately and pooled to obtain age- and sex-specific estimates of the prevalence of PD. An in-person screening instrument and diagnostic clinical examination were used to detect potential PD cases. The overall prevalence (per 100 population) in persons 65 years of age and older was 1.8, with an increase from 0.6 for those age 65 to 69 years to 2.6 for those 85 to 89 years. There were no sex differences in prevalence of PD. PMID- 10854358 TI - Prognosis with Parkinson's disease in europe: A collaborative study of population based cohorts. Neurologic Diseases in the Elderly Research Group. AB - Data are lacking on the prognosis (institutionalization and death) of PD cases identified in population-based studies. Data from five population-based European studies were compared and pooled. Each study used comparable two-step screening methods to identify cases and performed one or more follow-up examinations of their respective participants after defined periods of time. PD was classified on the basis of questionnaire and clinical data. The studies include 16,143 participants (252 with PD). The relative risk (RR) (95% CI) of death associated with PD was 2.3 (1.8 to 3.0). The risk for death in men with PD (RR 3.1 [2.1 to 4.4]) was higher than in women with PD (RR 1. 8 [1.2 to 5.1]). The rate of institutionalization varied across studies, increased with age, and was considerably higher in PD cases compared to noncases. Women with PD had a fivefold higher risk to live in a care facility than did men with PD. These data on mortality and rate of institutionalization reflect the high burden of PD in the population. PMID- 10854359 TI - Frequency of stroke in Europe: A collaborative study of population-based cohorts. ILSA Working Group and the Neurologic Diseases in the Elderly Research Group. Italian Longitudinal Study on Aging. AB - The authors estimated stroke frequency in Europe using data from six population based studies that were analyzed separately and also pooled. Overall, these surveys comprised 19,132 individuals age 55 years and older for the prevalence analyses, and 35,577 person-years for the incidence analyses. The overall prevalence of stroke, age- and sex-adjusted to the 1991 European population, was 4.84% (95% CI 4.47 to 5.21) in individuals age 65 to 84 years and 7.06% (95% CI 6.52 to 7.60) in those 75 years and older. Age- and sex-specific rates did not differ substantially across the studies. Incidence rates of first-ever stroke rose markedly with age, and the pooled results confirmed that incidence keeps increasing even at 90 years of age and older. The overall age- and sex standardized incidence rates were 8.72 per 1000 person-years (95% CI 7.47 to 10.06) for individuals age 65 to 84 years, and 17.31 per 1000 person-years (95% CI 14.79 to 20.02) for those age 75 years and over. PMID- 10854360 TI - Prognosis with stroke in Europe: A collaborative study of population-based cohorts. Neurologic Diseases in the Elderly Research Group. AB - The authors pooled four population-based European studies to determine the risk for institutionalization and death after stroke. Risk for institutionalization was assessed in 13,095 subjects. Survival was calculated for prevalent stroke in 7929 individuals and incident stroke in 8550 individuals. The adjusted risk of long-term care for cases compared to noncases was 2.8 (95% CI 2.6 to 4.5). At 5 years, the excess risk of death in prevalent cases was 1.6 (1.2 to 2.0), and the age- and sex-adjusted survival rate was 0.87 compared to 0.91 in noncases. The 5 year survival rate for incident cases was 0.84. PMID- 10854361 TI - Dementia, Parkinson's disease, and stroke in Europe: A commentary. PMID- 10854362 TI - Tuberculous radiculomyelitis complicating tuberculous meningitis: case report and review. AB - Tuberculous radiculomyelitis (TBRM) is a complication of tuberculous meningitis (TBM), which has been reported rarely in the modern medical literature. We describe a case of TBRM that developed in an human immunodeficiency virus (HIV) infected patient, despite prompt antituberculous treatment. To our knowledge, this is the second case of TBRM reported in an HIV-infected patient. We also review 74 previously reported cases of TBRM. TBRM develops at various periods after TBM, even in adequately treated patients after sterilization of the cerebrospinal fluid (CSF). The most common symptoms are subacute paraparesis, radicular pain, bladder disturbance, and subsequent paralysis. CSF evaluation usually shows an active inflammatory response with a very high protein level. MRI and CT scan are critical for diagnosis, revealing loculation and obliteration of the subarachnoid space along with linear intradural enhancement. As in other forms of paradoxical reactions to antituberculous treatment, there is evidence that steroid treatment might have a beneficial effect. PMID- 10854363 TI - Factors associated with severe manifestations of histoplasmosis in AIDS. AB - We report factors associated with severe manifestations of histoplasmosis (such as shock, respiratory failure, and death) in patients with AIDS during an outbreak. Severe disease was present in 28 of 155 patients (17.9%). The following factors were associated with severe disease: black race (odds ratio [OR], 2.8; 95% confidence interval [CI], 1.2-6.2); hemoglobin level <9.5 g/dL (OR, 2.7; 95% CI, 1.2-6.4), partial thromboplastin time >45 s (OR, 3.1; 95% CI, 1.1-9.3); alkaline phosphatase level >2.5 times normal (OR, 3.4; 95% CI, 1.3-8.7); aspartate aminotransferase level >2.5 times normal (OR, 4.2; 95% CI, 1.7-10.0); bilirubin level concentration >1.5 mg/dL (OR, 9.2; 95% CI, 2.5-34.3); creatinine concentration >2.1 mg/dL (OR, 8.3; 95% CI, 2.2-31.9); and albumin concentration <3.5 g/dL (OR, 4.6; 95% CI, 1.3-16.4). Zidovudine use was associated with decreased risk of severe disease (OR, 0.3; 95% CI, 0.1-0.7). Multivariate analysis showed that a creatinine value >2.1 mg/dL (OR, 9.5; 95% CI, 1.7-52) and an albumin value <3.5 g/dL (OR, 4.8; 95% CI, 1.0-22) were associated with an increased risk of severe disease, and zidovudine therapy remained associated with a decreased risk (OR, 0.2; 95% CI, 0.1-0.6). Findings associated with severe histoplasmosis should be recognized early and the cases managed aggressively. PMID- 10854364 TI - Management of protease inhibitor-associated diarrhea. AB - Diarrhea is a common and often inadequately treated complication in patients with human immunodeficiency virus infection. Diarrhea has a significant impact on quality of life (QOL) and can contribute to malnutrition, weight loss, immunosuppression, and mortality. In addition, diarrhea may have a significant impact on compliance with antiretroviral therapy; however, this impact has not been adequately assessed. Medications, including protease inhibitors (PIs), are recognized as a common cause of diarrhea. Treatment of PI-associated diarrhea is largely nonspecific; most of the available literature is published only in abstract form and is based primarily on retrospective and survey data. Agents for which some efficacy has been shown for treatment of PI-associated diarrhea include oat bran, psyllium, loperamide, calcium carbonate, SP-303, and pancrelipase. Practitioners and patients need to work together to determine which treatment modality is appropriate based on efficacy, cost, and lifestyle. Management of diarrhea is crucial to improving QOL, controlling weight loss, and enhancing overall efficacy of antiretroviral therapy. PMID- 10854365 TI - Let's talk about error. PMID- 10854366 TI - Accreditation's role in reducing medical errors. PMID- 10854368 TI - Festive weight gain PMID- 10854367 TI - Medical error: the second victim PMID- 10854369 TI - Unsupervised nurses may soon give anesthetics. PMID- 10854370 TI - FDA committee recommends approval for viagra rival PMID- 10854371 TI - George W bush proposes insurance help for the poor PMID- 10854372 TI - Bypass grafts protect diabetic patients better than angioplasty. PMID- 10854374 TI - A book to make you think PMID- 10854373 TI - The high price of aging. PMID- 10854375 TI - Better clinical management of anorexia nervosa in teens. PMID- 10854377 TI - Teen passengers are a hazard to the driver PMID- 10854376 TI - Itchy bumps on the back. PMID- 10854378 TI - How to submit your medical images for publication in Med.Pix: Do you have images (slides, photos, etc.) of compelling visual cases in clinical medicine? PMID- 10854379 TI - Prevalence of HIV infection, sexually transmitted diseases, and hepatitis and related risk behavior in young women living in low-income neighborhoods of northern California. AB - OBJECTIVE: To estimate the prevalence of human immunodeficiency virus (HIV) infection, sexually transmitted diseases, and hepatitis and the associated sexual and drug-using behavior among women residing in low-income neighborhoods in 5 northern California counties. METHODS: From April 4, 1996, to January 6, 1998, women aged 18 to 29 years were recruited door-to-door from randomly selected street blocks within 1990 census block groups below the 10th percentile for median household income for each county. RESULTS: Of 24,223 dwellings enumerated, contact was made with residents from 19,546 (80.7%). Within contacted dwellings, 3,560 eligible women were identified and 2,545 enrolled (71.5%). Weighted estimates for disease prevalence were HIV infection, 0.3% (95% confidence interval, 0.1%-0.4%); syphilis, 0.7% (0.3%-1.1%); gonorrhea, 0.8% (0.3%-1.3%); chlamydia, 3.3% (2.4%-4.8%); herpes simplex virus, type 1, 73.7% (71.6%-76.9%); herpes simplex virus, type 2, 34.4% (29.9%-39.0%); hepatitis A, 33.5% (28.3% 38.7%); chronic hepatitis B, 0.8% (0.3%-1.2%); and hepatitis C, 2.5% (1.4%-3. 6%). Condom use at last sexual intercourse with a new partner was reported by 44.0% (33.9%-54.1%). Injection drug use in the last 6 months was reported by 1.8% (1.0%-2.7%). CONCLUSIONS: The Young Women's Survey provided population-based estimates of the prevalence of 8 infectious diseases and related risk behavior within a population for whom data are often difficult to collect. Population based data are needed for appropriate targeting and planning of primary and secondary disease prevention. PMID- 10854380 TI - Poor young women need information and empowerment. PMID- 10854381 TI - Family reports of pain in dying hospitalized patients: a structured telephone survey. AB - OBJECTIVES: To see how often families in Oregon reported moderate to severe pain in dying patients in late 1998 compared with late 1997. DESIGN: A systematic random sample of death certificates was used to identify family members of decedents who died in a hospital setting between October 1 and December 31, 1998. A structured telephone interview was used to obtain data. PARTICIPANTS: Family members of 103 decedents (who died in hospitals 2 to 4 months before data collection) were identified with the use of death certificates. RESULTS: In late 1998, 56 family members (54%) reported that their loved one experienced moderate to severe pain in the last week of life. CONCLUSIONS: Family reports of moderate to severe pain in dying hospitalized Oregonians remain high. The influence of environmental factors on pain management may have implications for practice and policy nationwide. PMID- 10854383 TI - Netphiles PMID- 10854382 TI - Measuring quality of care at the end of life: who? when? where? and how? PMID- 10854384 TI - Use of preventive health behaviors by lesbian, bisexual, and heterosexual women: questionnaire survey. AB - OBJECTIVES: To determine whether lesbians and bisexual women are less likely than heterosexual women to use preventive health measures. DESIGN: Written, anonymous, self-administered questionnaire. SETTING: 33 physicians' offices and community clinics mainly in urban areas of 13 states. PARTICIPANTS: 524 lesbians, 143 bisexual women, and 637 heterosexual women. RESULTS: Bisexual women were less likely than heterosexual women to have had appropriate cholesterol screening (odds ratio 0.29, 95% confidence interval 0.11 to 0.73) or appropriate mammography (0.33, 0.13 to 0.84). Human immunodeficiency virus testing was more common in lesbians (2.38, 1. 51 to 3.74) and bisexual women (1.99, 1.17 to 3.38) than in heterosexual women. Illicit drug use was higher in lesbians (2.04, 1. 14 to 3.70) and bisexual women (1.96, 1.07 to 3.57) than in heterosexual women. Lesbians were more likely than heterosexual women to practice safer sex (2.60, 1.23 to 5.49) and less likely to have ever been infected with human papillomavirus (0.48, 0.25 to 0. 89). CONCLUSION: There were important differences in the preventive health measures taken by lesbians and bisexual women and those taken by heterosexual women. All patients should receive standard health tests, such as cholesterol screening and mammography, regardless of their sexual orientation. Lesbians and bisexual women who report illicit drug use should receive counseling, as appropriate. PMID- 10854385 TI - Funding for more research is urgently needed. PMID- 10854386 TI - Survey of directors of emergency departments in California on overcrowding. AB - OBJECTIVE: To survey the directors of emergency departments in California on their opinions of the extent and factors associated with overcrowding in emergency departments. METHODS: Surveys were mailed to a random sample of emergency department directors. Questions included estimated magnitude, frequency, causes, and effects of overcrowding. RESULTS: Of 160 directors surveyed, 113 (71%) responded, and 109 (96%) reported overcrowding as a problem. All (n = 21) university or county hospital directors and most (n = 88 [96%]) private or community hospital directors reported overcrowding. The 4 private or community hospital directors reporting no overcrowding serve smaller communities with populations less than 250,000. Thirty-two directors (28%) reported daily overcrowding. The most cited causes were increasing patient acuity and volume, hospital bed shortage, laboratory delays, and nursing shortage. These putative causes were similar between university or county and private or community hospital directors, except for consultant delays, which were more prevalent in university or county hospital emergency departments. CONCLUSIONS: Overcrowding is perceived to be a serious problem by emergency department directors. Many factors may contribute to overcrowding, and most are beyond the control of emergency departments. PMID- 10854388 TI - Practice point: keep looking for a reason PMID- 10854387 TI - Give emergency medicine true departmental control. PMID- 10854389 TI - Epidemiology of medical error PMID- 10854390 TI - Human error: models and management PMID- 10854391 TI - Sport medicine and the ethics of boxing PMID- 10854392 TI - Acute mountain sickness: the "poison of the pass" PMID- 10854393 TI - A book to make you think PMID- 10854394 TI - How to talk about sex with patients who are not heterosexual. PMID- 10854395 TI - Bottled water contains more bugs PMID- 10854396 TI - Health care problems of lesbian, gay, bisexual, and transgender patients. PMID- 10854397 TI - The challenge of preventive medicine in the year 2000 PMID- 10854399 TI - High fat diet may reduce seizure rate PMID- 10854398 TI - Infectious disease: diarrhea. PMID- 10854400 TI - At what age should a child begin regular continuous exercise at moderate or high intensity? PMID- 10854401 TI - Medical myth: Measuring white blood cells in the stools is useful in the management of acute diarrhea. PMID- 10854402 TI - Vietnamese family relationships: a lesson in cross-cultural care. PMID- 10854403 TI - Mission to Mindanao. PMID- 10854405 TI - Medicine for what ails you PMID- 10854404 TI - Sensing a purpose. PMID- 10854406 TI - Acquisition and metastability of centromere identity and function: sequence analysis of a human neocentromere. PMID- 10854407 TI - Tandem clusters of membrane proteins in complete genome sequences. AB - The distribution of genes coding for membrane proteins was investigated in 16 complete genomes: 4 archaea, 11 bacteria, and 1 eukaryote. Membrane proteins were identified by our new method of predicting transmembrane segments () after the removal of amino-terminal signal peptides. Interestingly, about half of the membrane protein genes in each genome were found to be located next to another, forming tandem clusters. Roughly 10%-30% of the tandem clusters were conserved among organisms, and most of the conserved tandem clusters belonged to one of the three functional groups, namely, transporters, the electron transport system, and cell motility. A tandem cluster sometimes contained paralogous membrane proteins, in which case the cluster size and the number of transmembrane segments could be related to a functional category, especially to transporters. In addition to the clustering of membrane proteins, the clustering of membrane proteins and ATP binding proteins in the complete genomes was also analyzed. Although this clustering was not statistically significant, it was useful to identify candidate membrane protein partners of isolated ATP-binding protein components in the ABC transporters. Possible implications of tandem cluster organization of membrane protein genes are discussed including the complex formation and other functional coupling of protein products and the mechanism of protein translocation to the cell membrane. PMID- 10854408 TI - Conservation of DNA regulatory motifs and discovery of new motifs in microbial genomes. AB - Regulatory motifs can be found by local multiple alignment of upstream regions from coregulated sets of genes, or regulons. We searched for regulatory motifs using the program AlignACE together with a set of filters that helped us choose the motifs most likely to be biologically relevant in 17 complete microbial genomes. We searched the upstream regions of potentially coregulated genes grouped by three methods: (1) genes that make up functional pathways; (2) genes homologous to regulons from a well-studied species (Escherichia coli); and (3) groups of genes derived from conserved operons. This last group is based on the observation that genes making up homologous regulons in different species are often assorted into coregulated operons in different combinations. This allows partial reconstruction of regulons by looking at operon structure across several species. Unlike other methods for predicting regulons, this method does not depend on the availability of experimental data other than the genome sequence and the locations of genes. New, statistically significant motifs were found in the genome sequence of each organism using each grouping method. The most significant new motif was found upstream of genes in the methane-metabolism functional group in Methanobacterium thermoautotrophicum. We found that at least 27% of the known E. coli DNA-regulatory motifs are conserved in one or more distantly related eubacteria. We also observed significant motifs that differed from the E. coli motif in other organisms upstream of sets of genes homologous to known E. coli regulons, including Crp, LexA, and ArcA in Bacillus subtilis; four anaerobic regulons in Archaeoglobus fulgidus (NarL, NarP, Fnr, and ModE); and the PhoB, PurR, RpoH, and FhlA regulons in other archaebacterial species. We also used motif conservation to aid in finding new motifs by grouping upstream regions from closely related bacteria, thus increasing the number of instances of the motif in the sequence to be aligned. For example, by grouping upstream sequences from three archaebacterial species, we found a conserved motif that may regulate ferrous ion transport that was not found in individual genomes. Discovery of conserved motifs becomes easier as the number of closely related genome sequences increases. PMID- 10854409 TI - Comparative genome sequence analysis of the Bpa/Str region in mouse and Man. AB - The progress of human and mouse genome sequencing programs presages the possibility of systematic cross-species comparison of the two genomes as a powerful tool for gene and regulatory element identification. As the opportunities to perform comparative sequence analysis emerge, it is important to develop parameters for such analyses and to examine the outcomes of cross-species comparison. Our analysis used gene prediction and a database search of 430 kb of genomic sequence covering the Bpa/Str region of the mouse X chromosome, and 745 kb of genomic sequence from the homologous human X chromosome region. We identified 11 genes in mouse and 13 genes and two pseudogenes in human. In addition, we compared the mouse and human sequences using pairwise alignment and searches for evolutionary conserved regions (ECRs) exceeding a defined threshold of sequence identity. This approach aided the identification of at least four further putative conserved genes in the region. Comparative sequencing revealed that this region is a mosaic in evolutionary terms, with considerably more rearrangement between the two species than realized previously from comparative mapping studies. Surprisingly, this region showed an extremely high LINE and low SINE content, low G+C content, and yet a relatively high gene density, in contrast to the low gene density usually associated with such regions. PMID- 10854410 TI - An EST-enriched comparative map of Brassica oleracea and Arabidopsis thaliana. AB - A detailed comparative map of Brassica oleracea and Arabidopsis thaliana has been established based largely on mapping of Arabidopsis ESTs in two Arabidopsis and four Brassica populations. Based on conservative criteria for inferring synteny, "one to one correspondence" between Brassica and Arabidopsis chromosomes accounted for 57% of comparative loci. Based on 186 corresponding loci detected in B. oleracea and A. thaliana, at least 19 chromosome structural rearrangements differentiate B. oleracea and A. thaliana orthologs. Chromosomal duplication in the B. oleracea genome was strongly suggested by parallel arrangements of duplicated loci on different chromosomes, which accounted for 41% of loci mapped in Brassica. Based on 367 loci mapped, at least 22 chromosomal rearrangements differentiate B. oleracea homologs from one another. Triplication of some Brassica chromatin and duplication of some Arabidopsis chromatin were suggested by data that could not be accounted for by the one-to-one and duplication models, respectively. Twenty-seven probes detected three or more loci in Brassica, which represent 25.3% of the 367 loci mapped in Brassica. Thirty-one probes detected two or more loci in Arabidopsis, which represent 23.7% of the 262 loci mapped in Arabidopsis. Application of an EST-based, cross-species genomic framework to isolation of alleles conferring phenotypes unique to Brassica, as well as the challenges and opportunities in extrapolating genetic information from Arabidopsis to Brassica and to more distantly related crops, are discussed. PMID- 10854411 TI - A high-throughput AFLP-based method for constructing integrated genetic and physical maps: progress toward a sorghum genome map. AB - Sorghum is an important target for plant genomic mapping because of its adaptation to harsh environments, diverse germplasm collection, and value for comparing the genomes of grass species such as corn and rice. The construction of an integrated genetic and physical map of the sorghum genome (750 Mbp) is a primary goal of our sorghum genome project. To help accomplish this task, we have developed a new high-throughput PCR-based method for building BAC contigs and locating BAC clones on the sorghum genetic map. This task involved pooling 24,576 sorghum BAC clones ( approximately 4x genome equivalents) in six different matrices to create 184 pools of BAC DNA. DNA fragments from each pool were amplified using amplified fragment length polymorphism (AFLP) technology, resolved on a LI-COR dual-dye DNA sequencing system, and analyzed using Bionumerics software. On average, each set of AFLP primers amplified 28 single copy DNA markers that were useful for identifying overlapping BAC clones. Data from 32 different AFLP primer combinations identified approximately 2400 BACs and ordered approximately 700 BAC contigs. Analysis of a sorghum RIL mapping population using the same primer pairs located approximately 200 of the BAC contigs on the sorghum genetic map. Restriction endonuclease fingerprinting of the entire collection of sorghum BAC clones was applied to test and extend the contigs constructed using this PCR-based methodology. Analysis of the fingerprint data allowed for the identification of 3366 contigs each containing an average of 5 BACs. BACs in approximately 65% of the contigs aligned by AFLP analysis had sufficient overlap to be confirmed by DNA fingerprint analysis. In addition, 30% of the overlapping BACs aligned by AFLP analysis provided information for merging contigs and singletons that could not be joined using fingerprint data alone. Thus, the combination of fingerprinting and AFLP-based contig assembly and mapping provides a reliable, high-throughput method for building an integrated genetic and physical map of the sorghum genome. PMID- 10854412 TI - Whole-genome trees based on the occurrence of folds and orthologs: implications for comparing genomes on different levels. AB - We built whole-genome trees based on the presence or absence of particular molecular features, either orthologs or folds, in the genomes of a number of recently sequenced microorganisms. To put these genomic trees into perspective, we compared them to the traditional ribosomal phylogeny and also to trees based on the sequence similarity of individual orthologous proteins. We found that our genomic trees based on the overall occurrence of orthologs did not agree well with the traditional tree. This discrepancy, however, vanished when one restricted the tree to proteins involved in transcription and translation, not including problematic proteins involved in metabolism. Protein folds unite superficially unrelated sequence families and represent a most fundamental molecular unit described by genomes. We found that our genomic occurrence tree based on folds agreed fairly well with the traditional ribosomal phylogeny. Surprisingly, despite this overall agreement, certain classes of folds, particularly all-beta ones, had a somewhat different phylogenetic distribution. We also compared our occurrence trees to whole-genome clusters based on the composition of amino acids and di-nucleotides. Finally, we analyzed some technical aspects of genomic trees-e.g., comparing parsimony versus distance based approaches and examining the effects of increasing numbers of organisms. Additional information (e.g. clickable trees) is available from http://bioinfo.mbb.yale.edu/genome/trees. PMID- 10854413 TI - The complete mitochondrial DNA sequence of Scenedesmus obliquus reflects an intermediate stage in the evolution of the green algal mitochondrial genome. AB - Two distinct mitochondrial genome types have been described among the green algal lineages investigated to date: a reduced-derived, Chlamydomonas-like type and an ancestral, Prototheca-like type. To determine if this unexpected dichotomy is real or is due to insufficient or biased sampling and to define trends in the evolution of the green algal mitochondrial genome, we sequenced and analyzed the mitochondrial DNA (mtDNA) of Scenedesmus obliquus. This genome is 42,919 bp in size and encodes 42 conserved genes (i.e., large and small subunit rRNA genes, 27 tRNA and 13 respiratory protein-coding genes), four additional free-standing open reading frames with no known homologs, and an intronic reading frame with endonuclease/maturase similarity. No 5S rRNA or ribosomal protein-coding genes have been identified in Scenedesmus mtDNA. The standard protein-coding genes feature a deviant genetic code characterized by the use of UAG (normally a stop codon) to specify leucine, and the unprecedented use of UCA (normally a serine codon) as a signal for termination of translation. The mitochondrial genome of Scenedesmus combines features of both green algal mitochondrial genome types: the presence of a more complex set of protein-coding and tRNA genes is shared with the ancestral type, whereas the lack of 5S rRNA and ribosomal protein-coding genes as well as the presence of fragmented and scrambled rRNA genes are shared with the reduced-derived type of mitochondrial genome organization. Furthermore, the gene content and the fragmentation pattern of the rRNA genes suggest that this genome represents an intermediate stage in the evolutionary process of mitochondrial genome streamlining in green algae. PMID- 10854414 TI - The 10q25 neocentromere and its inactive progenitor have identical primary nucleotide sequence: further evidence for epigenetic modification. AB - We have previously localized the core centromere protein-binding domain of a 10q25.2-derived neocentromere to an 80-kb genomic region. Detailed analysis has indicated that the 80-kb neocentromere (NC) DNA has a similar overall organization to the corresponding region on a normal chromosome 10 (HC) DNA, derived from a genetically unrelated CEPH individual. Here we report sequencing of the HC DNA and its comparison to the NC sequence. Single-base differences were observed at a maximum rate of 4.6 per kb; however, no deletions, insertions, or other structural rearrangements were detected. To investigate whether the observed changes, or subsets of these, might be de novo mutations involved in neocentromerization (i.e., in committing a region of a chromosome to neocentromere formation), the progenitor DNA (PnC) from which the NC DNA descended, was cloned and sequenced. Direct comparison of the PnC and NC sequences revealed 100% identity, suggesting that the differences between NC and HC DNA are single nucleotide polymorphisms (SNPs) and that formation of the 10q25.2 NC did not involve a change in DNA sequence in the core centromere protein-binding NC region. This is the first study in which a cloned NC DNA has been compared directly with its inactive progenitor DNA at the primary sequence level. The results form the basis for future sequence comparison outside the core protein-binding domain, and provide direct support for the involvement of an epigenetic mechanism in neocentromerization. PMID- 10854415 TI - The mosaic structure of human pericentromeric DNA: a strategy for characterizing complex regions of the human genome. AB - The pericentromeric regions of human chromosomes pose particular problems for both mapping and sequencing. These difficulties are due, in large part, to the presence of duplicated genomic segments that are distributed among multiple human chromosomes. To ensure contiguity of genomic sequence in these regions, we designed a sequence-based strategy to characterize different pericentromeric regions using a single (162 kb) 2p11 seed sequence as a point of reference. Molecular and cytogenetic techniques were first used to construct a paralogy map that delineated the interchromosomal distribution of duplicated segments throughout the human genome. Monochromosomal hybrid DNAs were PCR amplified by primer pairs designed to the 2p11 reference sequence. The PCR products were directly sequenced and used to develop a catalog of sequence tags for each duplicon for each chromosome. A total of 685 paralogous sequence variants were generated by sequencing 34.7 kb of paralogous pericentromeric sequence. Using PCR products as hybridization probes, we were able to identify 702 human BAC clones, of which a subset, 107 clones, were analyzed at the sequence level. We used diagnostic paralogous sequence variants to assign 65 of these BACs to at least 9 chromosomal pericentromeric regions: 1q12, 2p11, 9p11/q12, 10p11, 14q11, 15q11, 16p11, 17p11, and 22q11. Comparisons with existing sequence and physical maps for the human genome suggest that many of these BACs map to regions of the genome with sequence gaps. Our analysis indicates that large portions of pericentromeric DNA are virtually devoid of unique sequences. Instead, they consist of a mosaic of different genomic segments that have had different propensities for duplication. These biologic properties may be exploited for the rapid characterization of, not only pericentromeric DNA, but also other complex paralogous regions of the human genome. PMID- 10854416 TI - Parallel genotyping of human SNPs using generic high-density oligonucleotide tag arrays. AB - Large scale human genetic studies require technologies for generating millions of genotypes with relative ease but also at a reasonable cost and with high accuracy. We describe a highly parallel method for genotyping single nucleotide polymorphisms (SNPs), using generic high-density oligonucleotide arrays that contain thousands of preselected 20-mer oligonucleotide tags. First, marker specific primers are used in PCR amplifications of genomic regions containing SNPs. Second, the amplification products are used as templates in single base extension (SBE) reactions using chimeric primers with 3' complementarity to the specific SNP loci and 5' complementarity to specific probes, or tags, synthesized on the array. The SBE primers, terminating one base before the polymorphic site, are extended in the presence of labeled dideoxy NTPs, using a different label for each of the two SNP alleles, and hybridized to the tag array. Third, genotypes are deduced from the fluorescence intensity ratio of the two colors. This approach takes advantage of multiplexed sample preparation, hybridization, and analysis at each stage. We illustrate and test this method by genotyping 44 individuals for 142 human SNPs identified previously in 62 candidate hypertension genes. Because the hybridization results are quantitative, this method can also be used for allele-frequency estimation in pooled DNA samples. PMID- 10854417 TI - Differentially painting human chromosome arms with combined binary ratio-labeling fluorescence in situ hybridization. AB - Recently we developed a novel strategy for differentially painting all 24 human chromosomes. It is termed COBRA-FISH, short for combined binary ratio labeling fluorescence in situ hybridization. COBRA-FISH is distinct from the pure combinatorial approach in that only 4 instead of 5 fluorophores are needed to achieve color discrimination of 24 targets. Furthermore, multiplicity can be increased to 48 by introduction of a fifth fluorophore. Here we show that color identification by COBRA-FISH of all of the p and q arms of human chromosomes is feasible, and we apply the technique for detecting and elucidating intra- and interchromosomal rearrangements. Compared with 24-color whole chromosome painting FISH, PQ-COBRA-FISH considerably enhances the ability to determine the composition of rearranged chromosomes as demonstrated by the identification of pericentric inversions and isochromosomes as well as the elucidation of the arm identity of chromosomal material involved in complex translocations that occur in solid tumors. PMID- 10854418 TI - A gene-enriched BAC library for cloning large allele-specific fragments from maize: isolation of a 240-kb contig of the bronze region. AB - A generic bacterial artificial chromosome (BAC) library from a complex plant genome like maize may not be suitable for some types of genomic analysis, for example, for establishing correlations between the genetic and the physical organization of a given chromosome region. Previously, we carried out extensive genetic analysis of the bronze (Bz) region in Zea mays using a W22 inbred line carrying the Bz-McC allele; however, BAC libraries of that line are neither available nor under construction. Here, we report the isolation of large, adjacent BAC clones of this region from a partial BAC library of W22. We developed a BAC vector suitable for cloning NotI fragments and used it to clone size-fractionated genomic DNA that had been cut to completion with the methylation-sensitive, rare-cutting enzyme NotI. This strategy resulted in a very significant enrichment of large genic DNA. From a library of about 20,000 BACs, containing just two-thirds of a maize genome, we isolated 16 BAC clones of the 110-kb distal Bz fragment and 10 BAC clones of the 130-kb proximal Bz fragment. This recovery means that our strategy resulted in a 15- to 24-fold enrichment of specific sequences. The order of the BAC clones in the 240-kb contig, predetermined from an internal NotI site in the Bz-McC allele was confirmed by hybridization with sequences from sites previously mapped proximal and distal to Bz and by sequencing. To show the general utility of our approach and the value of our partial BAC library, we also isolated BAC clones of other sequences, such as tub4 and the complex R-r allele, contained in the same size fraction of DNA. This is the first report of the use of a BAC vector to clone allele-specific large DNA fragments from a plant with a large genome, circumventing the need to construct a complete BAC library. PMID- 10854419 TI - A human genomic library enriched in transcriptionally active sequences (aDNA library). AB - Core histone hyperacetylation, in particular of H4, is concentrated in the promoter-upstream regions of active genes and in certain cases is locuswide. Antibodies to hyperacetylated H4 were used to immunoprecipitate dinucleosomal chromatin derived from K562 human erythroleukemic cells by micrococcal nuclease digestion. The extracted DNA was made into a genomic library and was expected to contain sequences from genes active in K562 cells (an active, 'aDNA' library). Clones (180) were randomly selected from the library; 24 of 103 tested (23%) contained highly repeated sequences, as determined by their hybridization to total genomic DNA, and were not analyzed further. An additional 10 clones (6%) were shown to contain no insert DNA. The remaining 146 were sequenced and compared with the nucleic acid databases and in all six frames to the protein databases: Sixeen clones could be assigned to known genes, the majority of which (12) were tissue specific. All but 2 of these 16 corresponded to segments 5' of the coding sequences, as expected if H4 acetylation is concentrated at promoter regions. Thirty-three clones (23%) displayed high sequence identity to cDNAs in the expressed sequence tag database (dbEST). Northern blots and reverse transcription (RT)-PCR were used to determine the proportion of clones representing sequences expressed in K562 cells: Although only 1 of 34 tested clones showed a band in Northern hybridization, RT-PCR demonstrated that at least 12 of 40 tested clones (30%) were present in the mRNA population. Because a further 8 of these 40 clones were identified as gene fragments by database sequence comparisons, it follows that about half of this subset of 40 clones is derived from genes. The aDNA library is thus very gene rich and not skewed toward the most highly expressed sequences, as in mRNA libraries. The aDNA library is also rich in promoters and could be a valuable source of such sequences, particularly those that lack CpG islands or other features that allow their specific selection. PMID- 10854420 TI - Activation of malonyl-CoA decarboxylase in rat skeletal muscle by contraction and the AMP-activated protein kinase activator 5-aminoimidazole-4-carboxamide-1-beta D-ribofuranoside. AB - Alterations in the concentration of malonyl-CoA, an inhibitor of carnitine palmitoyltransferase I, have been linked to the regulation of fatty acid oxidation in skeletal muscle. During contraction decreases in muscle malonyl-CoA concentration have been related to activation of AMP-activated protein kinase (AMPK), which phosphorylates and inhibits acetyl-CoA carboxylase (ACC), the rate limiting enzyme in malonyl-CoA formation. We report here that the activity of malonyl-CoA decarboxylase (MCD) is increased in contracting muscle. Using either immunopurified enzyme or enzyme partially purified by (NH(4))(2)SO(4) precipitation, 2-3-fold increases in the V(max) of MCD and a 40% decrease in its K(m) for malonyl-CoA (190 versus 119 micrometer) were observed in rat gastrocnemius muscle after 5 min of contraction, induced by electrical stimulation of the sciatic nerve. The increase in MCD activity was markedly diminished when immunopurified enzyme was treated with protein phosphatase 2A or when phosphatase inhibitors were omitted from the homogenizing solution and assay mixture. Incubation of extensor digitorum longus muscle for 1 h with 2 mm 5 aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside, a cell-permeable activator of AMPK, increased MCD activity 2-fold. Here, too, addition of protein phosphatase 2A to the immunopellets reversed the increase of MCD activity. The results strongly suggest that activation of AMPK during muscle contraction leads to phosphorylation of MCD and an increase in its activity. They also suggest a dual control of malonyl-CoA concentration by ACC and MCD, via AMPK, during exercise. PMID- 10854421 TI - Interactions of the DNA ligase IV-XRCC4 complex with DNA ends and the DNA dependent protein kinase. AB - The DNA-dependent protein kinase (DNA-PK), consisting of Ku and the DNA-PK catalytic subunit (DNA-PKcs), and the DNA ligase IV-XRCC4 complex function together in the repair of DNA double-strand breaks by non-homologous end joining. These protein complexes are also required for the completion of V(D)J recombination events in immune cells. Here we demonstrate that the DNA ligase IV XRCC4 complex binds specifically to the ends of duplex DNA molecules and can act as a bridging factor, linking together duplex DNA molecules with complementary but non-ligatable ends. Although the DNA end-binding protein Ku inhibited DNA joining by DNA ligase IV-XRCC4, it did not prevent this complex from binding to DNA. Instead, DNA ligase IV-XRCC4 and Ku bound simultaneously to the ends of duplex DNA molecules. DNA ligase IV-XRCC4 and DNA-PKcs also formed complexes at the ends of DNA molecules, but DNA-PKcs did not inhibit ligation. Interestingly, DNA-PKcs stimulated intermolecular ligation by DNA ligase IV-XRCC4. In the presence of DNA-PK, the majority of the joining events catalyzed by DNA ligase IV XRCC4 were intermolecular because Ku inhibited intramolecular ligation, but DNA PKcs still stimulated intramolecular ligation. We suggest that DNA-PKcs containing complexes formed at DNA ends enhance the association of DNA ends via protein-protein interactions, thereby stimulating intermolecular ligation. PMID- 10854422 TI - Aspartic proteases from the nematode Caenorhabditis elegans. Structural organization and developmental and cell-specific expression of asp-1. AB - A Caenorhabditis elegans gene (asp-1) and cDNA that encode a homologue of cathepsin D aspartic protease were cloned and characterized. The asp-1 mRNA is transcribed from a single exon, and it begins with the SL1 trans-splice leader sequence. The protein (ASP-1) is expressed as a 396-amino acid, 42.7-kDa pre-pro peptide that is post-translationally processed into a approximately 40-kDa lysosomal protein. ASP-1 shares approximately 60% sequence identity with the aspartic protease precursor from the nematode Strongyloides stercoralis. The amino acid sequences adjacent to the two active site aspartic acid residues in ASP-1 are 100% identical to those in other eukaryotic aspartic proteases. In addition, ASP-1 contains conserved, potential disulfide bond-forming cysteine residues and N-glycosylation sites. The asp-1 gene is exclusively transcribed in the intestinal cells, with the highest levels of expression observed at late embryonic and early larval stages of development. asp-1 transcription is not observed in adult nematodes or mature larvae. Furthermore, transcription predominantly occurs in eight anterior cells of the intestine (int6-int8). Analyses of ASP-1 nucleotide and amino acid sequences revealed the presence of five additional C. elegans aspartic proteases. PMID- 10854423 TI - 3-Methyladenine-DNA glycosylase (MPG protein) interacts with human RAD23 proteins. AB - Human 3-methyladenine-DNA glycosylase (MPG protein) initiates base excision repair by severing the glycosylic bond of numerous damaged bases. In comparison, homologues of the Rad23 proteins (hHR23) and the hXPC protein are involved in the recognition of damaged bases in global genome repair, a subset of nucleotide excision repair. In this report, we show that the hHR23A and -B also interact with the MPG protein and can serve as accessory proteins for DNA damage recognition in base excision repair. Furthermore, the MPG.hHR23 protein complex elevates the rate of MPG protein-catalyzed excision from hypoxanthine-containing substrates. This increased excision rate is correlated with a greater binding affinity of the MPG protein-hHR23 protein complex for damaged DNA. These data suggest that the hHR23 proteins function as universal DNA damage recognition accessory proteins in both of these major excision repair pathways. PMID- 10854424 TI - Receptor subunit-specific action of oncostatin M in hepatic cells and its modulation by leukemia inhibitory factor. AB - The related cytokines, interleukin-6 (IL-6), oncostatin M (OSM), and leukemia inhibitory factor (LIF) direct the formation of specific heteromeric receptor complexes to achieve signaling. Each complex includes the common signal transducing subunit gp130. OSM and LIF also recruit the signaling competent, but structurally distinct OSMRbeta and LIFRalpha subunits, respectively. To test the hypothesis that the particularly prominent cell regulation by OSM is due to signals contributed by OSMRbeta, we introduced stable expression of human or mouse OSMRbeta in rat hepatoma cells which have endogenous receptors for IL-6 and LIF, but not OSM. Both mouse and human OSM engaged gp130 with their respective OSMRbeta subunits, but only human OSM also acted through LIFR. Signaling by OSMRbeta-containing receptors was characterized by highest activation of STAT5 and ERK, recruitment of the insulin receptor substrate and Jun-N-terminal kinase pathways, and induction of a characteristic pattern of acute phase proteins. Since LIF together with LIFRalpha appear to form a more stable complex with gp130 than OSM with gp130 and OSMRbeta, co-activation of LIFR and OSMR resulted in a predominant LIF-like response. These results suggest that signaling by IL-6 cytokines is not identical, and that a hierarchical order of cytokine receptor action exists in which LIFR ranks as dominant member. PMID- 10854425 TI - Translational induction of liver-enriched transcriptional inhibitory protein during acute phase response leads to repression of CCAAT/enhancer binding protein alpha mRNA. AB - Lipopolysacharide (LPS) induced acute phase response (APR) in mouse liver leads to elevation of the low molecular weight CCAAT/Enhancer binding protein (C/EBP) beta isoform, liver-enriched transcriptional inhibitory protein (LIP). In this paper, we investigate the pathway for LIP induction during APR and the role of LIP in regulation of the C/EBPalpha promoter. The 5' region of C/EBPbeta mRNA has been shown to be involved in the regulation of LIP translation. Our data demonstrate that binding of cytoplasmic proteins to the 5' region of C/EBPbeta mRNA is altered in response to LPS administration. One of the major changes is induced binding of a cytoplasmic protein that is immunologically identical to the previously characterized RNA-binding protein CUGBP1. Induction of CUGBP1 binding activity in liver cytoplasm during APR is accompanied by the elevation of CUGBP1 binding activity on polysomes. CUGBP1 immunoprecipitated from livers of LPS treated mice, but not from normal animals, is capable of inducing LIP translation in a cell-free translation system. The ability of CUGBP1 to induce LIP translation during APR depends on phosphorylation of CUGBP1. We show that elevation of LIP during APR and after partial hepatectomy leads to increased binding of LIP to the C/EBP consensus site found within the mouse C/EBPalpha promoter. This binding correlates with reduction of C/EBPalpha mRNA levels in both biological situations. Co-transfection experiments showed that full-length C/EBPbeta activates the C/EBPalpha promoter, while LIP blocks this activation. Our data suggest that the dominant negative isoform of C/EBPbeta, LIP, down regulates the C/EBPalpha promoter in liver and in cultured hepatocytes. Because full-length C/EBPalpha and C/EBPbeta proteins regulate liver proliferation, this function of LIP may be important in liver growth and differentiation. PMID- 10854426 TI - Biosynthesis of type 3 capsular polysaccharide in Streptococcus pneumoniae. Enzymatic chain release by an abortive translocation process. AB - The type 3 polysaccharide synthase from Streptococcus pneumoniae catalyzes sugar transfer from UDP-Glc and UDP-glucuronic acid (GlcUA) to a polymer with the repeating disaccharide unit of [3)-beta-d-GlcUA-(1-->4)-beta-d-Glc-(1-->]. Evidence is presented that release of the polysaccharide chains from S. pneumoniae membranes is time-, temperature-, and pH-dependent and saturable with respect to specific catalytic metabolites. In these studies, the membrane-bound synthase was shown to catalyze a rapid release of enzyme-bound polysaccharide when either UDP-Glc or UDP-GlcUA alone was present in the reaction. Only a slow release of polysaccharide occurred when both UDP sugars were present or when both UDP sugars were absent. Chain size was not a specific determinant in polymer release. The release reaction was saturable with increasing concentrations of UDP Glc or UDP-GlcUA, with respective apparent K(m) values of 880 and 0.004 micrometer. The apparent V(max) was 48-fold greater with UDP-Glc compared with UDP-GlcUA. The UDP-Glc-actuated reaction was inhibited by UDP-GlcUA with an approximate K(i) of 2 micrometer, and UDP-GlcUA-actuated release was inhibited by UDP-Glc with an approximate K(i) of 5 micrometer. In conjunction with kinetic data regarding the polymerization reaction, these data indicate that UDP-Glc and UDP-GlcUA bind to the same synthase sites in both the biosynthetic reaction and the chain release reaction and that polymer release is catalyzed when one binding site is filled and the concentration of the conjugate UDP-precursor is insufficient to fill the other binding site. The approximate energy of activation values of the biosynthetic and release reactions indicate that release of the polysaccharide occurs by an abortive translocation process. These results are the first to demonstrate a specific enzymatic mechanism for the termination and release of a polysaccharide. PMID- 10854427 TI - Expression cloning of a new member of the ABO blood group glycosyltransferases, iGb3 synthase, that directs the synthesis of isoglobo-glycosphingolipids. AB - The large array of different glycolipids described in mammalian tissues is a reflection, in part, of diverse glycosyltransferase expression. Herein, we describe the cloning of a UDP-galactose: beta-d-galactosyl-1,4-glucosylceramide alpha-1, 3-galactosyltransferase (iGb(3) synthase) from a rat placental cDNA expression library. iGb(3) synthase acts on lactosylceramide, LacCer (Galbeta1,4Glcbeta1Cer) to form iGb(3) (Galalpha1,3Galbeta1, 4Glcbeta1Cer) initiating the synthesis of the isoglobo-series of glycosphingolipids. The isolated cDNA encoded a predicted protein of 339 amino acids, which shows extensive homology (40-50% identity) to members of the ABO gene family that includes: murine alpha1, 3-galactosyltransferase, Forssman (Gb(5)) synthase, and the ABO glycosyltransferases. In contrast to the murine alpha1, 3 galactosyltransferase, iGb(3) synthase preferentially modifies glycolipids over glycoprotein substrates. Reverse transcriptase-polymerase chain reaction revealed a widespread tissue distribution of iGb(3) synthase RNA expression, with high levels observed in spleen, thymus, and skeletal muscle. As an indirect consequence of the expression cloning strategy used, we have been able to identify several potential glycolipid biosynthetic pathways where iGb(3) functions, including the globo- and isoglobo-series of glycolipids. PMID- 10854428 TI - Cloning of Gb3 synthase, the key enzyme in globo-series glycosphingolipid synthesis, predicts a family of alpha 1, 4-glycosyltransferases conserved in plants, insects, and mammals. AB - We have cloned Gb(3) synthase, the key alpha1, 4-galactosyltransferase in globo series glycosphingolipid (GSL) synthesis, via a phenotypic screen, which previously yielded iGb(3) synthase, the alpha1,3-galactosyltransferase required in isoglobo-series GSL (Keusch, J. J., Manzella, S. M., Nyame, K. A., Cummings, R. D., and Baenziger, J. U. (2000) J. Biol. Chem. 33). Both transferases act on lactosylceramide, Galbeta1,4Glcbeta1Cer (LacCer), to produce Gb(3) (Galalpha1,4LacCer) or iGb(3) (Galalpha1, 3LacCer), respectively. GalNAc can be added sequentially to either Gb(3) or iGb(3) yielding globoside and Forssman from Gb(3), and isogloboside and isoForssman from iGb(3). Gb(3) synthase is not homologous to iGb(3) synthase but shows 43% identity to a human alpha1,4GlcNAc transferase that transfers a UDP-sugar in an alpha1, 4-linkage to a beta-linked Gal found in mucin. Extensive homology (35% identity) is also present between Gb(3) synthase and genes in Drosophila melanogaster and Arabidopsis thaliana, supporting conserved expression of an alpha1,4-glycosyltransferase, possibly Gb(3) synthase, throughout evolution. The isolated Gb(3) synthase cDNA encodes a type II transmembrane glycosyltransferase of 360 amino acids. The highest tissue expression of Gb(3) synthase RNA is found in the kidney, mesenteric lymph node, spleen, and brain. Gb(3) glycolipid, also called P(k) antigen or CD77, is a known receptor for verotoxins. CHO cells that do not express Gb(3) and are resistant to verotoxin become susceptible to the toxin following transfection with Gb(3) synthase cDNA. PMID- 10854429 TI - Engagement of bone morphogenetic protein type IB receptor and Smad1 signaling by anti-Mullerian hormone and its type II receptor. AB - Anti-Mullerian hormone induces the regression of fetal Mullerian ducts and inhibits the transcription of gonadal steroidogenic enzymes. It belongs to the transforming growth factor-beta family whose members signal through a pair of serine/threonine kinase receptors and Smad effectors. Only the anti-Mullerian hormone type II receptor has been identified. Our goal was to determine whether anti-Mullerian hormone could share a type I receptor with another family member. Co-immunoprecipitation of known type I receptors with anti-Mullerian hormone type II receptor clearly showed that the bone morphogenetic protein type IB receptor was the only cloned type I receptor interacting in a ligand-dependent manner with this type II receptor. Anti-Mullerian hormone also activates the bone morphogenetic protein-specific Smad1 pathway and the XVent2 reporter gene, an anti-Mullerian hormone type II receptor-dependent effect abrogated by a dominant negative version of bone morphogenetic protein type IB receptor. Reverse amplification experiments showed that bone morphogenetic protein type IB receptor is co-expressed with anti-Mullerian hormone type II receptor in most anti Mullerian hormone target tissues. Our data support a model in which a ligand, anti-Mullerian hormone, gains access to a shared type I receptor and Smad1 system through a highly restricted type II receptor. PMID- 10854430 TI - Determinants of vitellogenin B1 promoter architecture. HNF3 and estrogen responsive transcription within chromatin. AB - The liver-specific vitellogenin B1 promoter is efficiently activated by estrogen within a nucleosomal environment after microinjection into Xenopus laevis oocytes, consistent with the hypothesis that significant nucleosome remodeling over this promoter is not a prerequisite for the activation by the estrogen receptor (ERalpha). This observation lead us to investigate determinants other than ERalpha of chromatin structure and transcriptional activation of the vitellogenin B1 promoter in this system and in vitro. We find that the liver enriched transcription factor HNF3 has an important organizational role for chromatin structure as demonstrated by DNase I-hypersensitive site mapping. Both HNF3 and the estrogen receptor activate transcription synergistically and are able to interact with chromatin reconstituted in vitro with three positioned nucleosomes. We propose that HNF3 is the cellular determinant which establishes a promoter environment favorable to a rapid transcriptional activation by the estrogen receptor. PMID- 10854431 TI - Stress-activated protein kinase/JNK activation and apoptotic induction by the macrophage P2X7 nucleotide receptor. AB - In human and rodent macrophages, activation of the P2X7 nucleotide receptor stimulates interleukin-1beta processing and release, apoptosis, and killing of intracellular Mycobacterium tuberculosis. Signaling pathways downstream of this ionotropic ATP receptor are poorly understood. Here we describe the rapid activation of the stress-activated protein kinase (SAPK)/JNK pathway in BAC1 murine macrophages stimulated by extracellular ATP. Brief exposure of the cells to ATP (10-30 min) was sufficient to trigger a rapid accumulation of activated SAPK that was then sustained for >120 min. Several observations indicated that the P2X7 receptor mediated this effect. 1) ATP and 3'-O-(4-benzoyl)benzoyl-ATP were the only agonistic nucleotides. 2) The effect was inhibited by oxidized ATP and the isoquinoline KN-62, two known P2X7 receptor antagonists. 3) ATP-induced SAPK activation could be recapitulated in P2X7 receptor-transfected HEK293 cells, but not in wild-type HEK293 cells. Because P2X7 receptor stimulation can rapidly activate caspase family proteases that have been implicated in the induction of the SAPK pathway, we investigated whether ATP-dependent SAPK activation involved such proteases. Brief exposure of BAC1 macrophages to extracellular ATP induced DNA fragmentation, alpha-fodrin breakdown, and elevated levels of caspase-3-type activity. Asp-Glu-Val-Asp-cho, a caspase-3 inhibitor, inhibited ATP-induced DNA fragmentation and alpha-fodrin proteolysis, but had no effect on ATP-induced SAPK activation. Tyr-Val-Ala-Asp-chloromethyl ketone, a caspase-1 inhibitor, prevented ATP-induced release of processed interleukin-1beta, but not ATP-dependent SAPK activity. We conclude that activation of ionotropic P2X7 nucleotide receptors triggers a strong activation of SAPK via a pathway independent of caspase-1- or caspase-3-like proteases. PMID- 10854432 TI - Alternatively spliced isoforms of TFII-I. Complex formation, nuclear translocation, and differential gene regulation. AB - TFII-I is a multifunctional phosphoprotein with roles in transcription and signal transduction. Here we report characterization of three additional alternatively spliced isoforms of TFII-I. Employing isoform-specific antibodies, we show that the isoforms form a stable complex in vivo preferentially in the nucleus compared with the cytoplasm. We further show that both homomeric and heteromeric interactions are possible and that the heteromeric interactions between a wild type and a nuclear localization-deficient mutant result in nuclear translocation of the complex, leading us to postulate that complex formation might aid in nuclear translocation. In functional assays all four isoforms individually bind to DNA and transactivate reporter genes to a similar extent. However, although co expression of different TFII-I isoforms leads to enhanced basal activity, it results in attenuated signal responsive activity. Thus, TFII-I might differentially regulate its target genes via complex or subcomplex formation. PMID- 10854433 TI - Crystal structure and thermodynamic analysis of human brain fatty acid-binding protein. AB - Expression of brain fatty acid-binding protein (B-FABP) is spatially and temporally correlated with neuronal differentiation during brain development. Isothermal titration calorimetry demonstrates that recombinant human B-FABP clearly exhibits high affinity for the polyunsaturated n-3 fatty acids alpha linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, and for monounsaturated n-9 oleic acid (K(d) from 28 to 53 nm) over polyunsaturated n-6 fatty acids, linoleic acid, and arachidonic acid (K(d) from 115 to 206 nm). B FABP has low binding affinity for saturated long chain fatty acids. The three dimensional structure of recombinant human B-FABP in complex with oleic acid shows that the oleic acid hydrocarbon tail assumes a "U-shaped" conformation, whereas in the complex with docosahexaenoic acid the hydrocarbon tail adopts a helical conformation. A comparison of the three-dimensional structures and binding properties of human B-FABP with other homologous FABPs, indicates that the binding specificity is in part the result of nonconserved amino acid Phe(104), which interacts with double bonds present in the lipid hydrocarbon tail. In this context, analysis of the primary and tertiary structures of human B FABP provides a rationale for its high affinity and specificity for polyunsaturated fatty acids. The expression of B-FABP in glial cells and its high affinity for docosahexaenoic acid, which is known to be an important component of neuronal membranes, points toward a role for B-FABP in supplying brain abundant fatty acids to the developing neuron. PMID- 10854436 TI - beta 2-adrenergic receptor internalization, endosomal sorting, and plasma membrane recycling are regulated by rab GTPases. AB - Rab GTPases are recognized as critical regulatory factors involved in vesicular membrane transport and endosomal fusion. For example, Rab5 directs the transport and fusion of endocytic vesicles to and with early endosomes, whereas Rab4 is thought to control protein trafficking from early endosomes back to the plasma membrane. In the present study, we investigated the role of Rab5 and Rab4 GTPases in regulating the endocytosis, intracellular sorting, and the plasma membrane recycling of the beta(2)AR. In cells expressing the dominant-negative Rab5-S34N mutant, beta(2)AR internalization was impaired, and beta(2)AR-bearing endocytic vesicles remained in either close juxtaposition or physically attached to the plasma membrane. In contrast, a constitutively active Rab5-Q79L mutant redirected internalized beta(2)AR to enlarged endosomes but did not prevent beta(2)AR dephosphorylation and recycling. The expression of either wild-type Rab4 or a Rab4-N121I mutant did not prevent beta(2)AR dephosphorylation. However, the dominant-negative Rab4-N121I mutant blocked beta(2)AR resensitization by blocking receptor recycling from endosomes back to the cell surface. Our data indicate that, in addition to regulating the intracellular trafficking and fusion of beta(2)AR-bearing endocytic vesicles, Rab5 also contributes to the formation and/or budding of clathrin-coated vesicles. Furthermore, beta(2)AR dephosphorylation occurs as the receptor transits between Rab5- and Rab4-positive compartments. PMID- 10854437 TI - Identification of Rho GTPase-dependent sites in the Dbl homology domain of oncogenic Dbl that are required for transformation. AB - The Dbl family guanine-nucleotide exchange factors (GEFs) for Rho GTPases share the structural array of a Dbl homology (DH) domain in tandem with a Pleckstrin homology (PH) domain. For oncogenic Dbl, the DH domain is responsible for the GEF activity, and the DH-PH module constitutes the minimum structural unit required for cellular transformation. To understand the structure-function relationship of the DH domain, we have investigated the role of specific residues of the DH domain of Dbl in interaction with Rho GTPases and in Dbl-induced transformation. Alanine substitution mutagenesis identified a panel of DH mutants made in the alpha1, alpha6, and alpha9 regions and the PH junction site that suffer complete or partial loss of GEF activity toward Cdc42 and RhoA. Kinetic and binding analysis of these mutants revealed that although most displayed decreased k(cat) values in the GEF reaction, the substrate binding activities of T506A and R634A were significantly reduced. E502A, Q633A, and N673A/D674A, on the other hand, retained the binding capability to the Rho GTPases but lost the GEF catalytic activity. In general, the in vitro GEF activity of the DH mutants correlated with the in vivo Cdc42- and RhoA-activating potential, and the GEF catalytic efficiency mirrored the transforming activity in NIH 3T3 cells. Moreover, the N673A/D674A mutant exhibited a potent dominant-negative effect on serum-induced cell growth and caused retraction of actin structures. These studies identify important sites of the DH domain involved in binding or catalysis of Rho proteins and demonstrate that maintaining a threshold of GEF catalytic activity, in addition to the Rho GTPase binding activity, is essential for efficient transformation by oncogenic Dbl. PMID- 10854438 TI - Disulfide bonds are generated by quinone reduction. AB - The chemistry of disulfide exchange in biological systems is well studied. However, very little information is available concerning the actual origin of disulfide bonds. Here we show that DsbB, a protein required for disulfide bond formation in vivo, uses the oxidizing power of quinones to generate disulfides de novo. This is a novel catalytic activity, which to our knowledge has not yet been described. This catalytic activity is apparently the major source of disulfides in vivo. We developed a new assay to characterize further this previously undescribed enzymatic activity, and we show that quinones get reduced during the course of the reaction. DsbB contains a single high affinity quinone-binding site. We reconstitute oxidative folding in vitro in the presence of the following components that are necessary in vivo: DsbA, DsbB, and quinone. We show that the oxidative refolding of ribonuclease A is catalyzed by this system in a quinone dependent manner. The disulfide isomerase DsbC is required to regain ribonuclease activity suggesting that the DsbA-DsbB system introduces at least some non-native disulfide bonds. We show that the oxidative and isomerase systems are kinetically isolated in vitro. This helps explain how the cell avoids oxidative inactivation of the disulfide isomerization pathway. PMID- 10854439 TI - Molecular determinants of tuberoinfundibular peptide of 39 residues (TIP39) selectivity for the parathyroid hormone-2 (PTH2) receptor. N-terminal truncation of TIP39 reverses PTH2 receptor/PTH1 receptor binding selectivity. AB - Tuberoinfundibular peptide of 39 residues (TIP39) and the parathyroid hormone-2 (PTH2) receptor form part of an extended family of related signaling molecules that includes the PTH1 receptor, which responds to PTH and PTH-related protein. TIP39 does not appreciably activate the PTH1 receptor, but in this study it is shown to bind the receptor with moderate affinity (59 nm). In this study, we investigated the molecular determinants of both ligand and receptor for the PTH2 receptor selectivity of TIP39 and quantitatively evaluated the role of molecular elements in the binding of TIP39 to the PTH2 and PTH1 receptors. A chimeric receptor composed of the N-terminal extracellular domain of the PTH1 receptor and the remainder (juxtamembrane domain) of the PTH2 receptor (P2-NP1) was fully activated by TIP39 (E(max) = 98% of the rPTH-(1-34), E(max), EC(50) = 2.0 nm). This receptor chimera bound TIP39 with an equivalent affinity to the wild-type PTH2 receptor (2. 3 and 2.0 nm, respectively). The reciprocal chimeric receptor (P1-NP2) was not activated by TIP39 and bound the ligand with an affinity equivalent to that of the PTH1 receptor. Thus, the juxtamembrane receptor domain specifies the signaling and binding selectivity of TIP39 for the PTH2 receptor over the PTH1 receptor. Removing six N-terminal residues of TIP39 eliminated activation of the PTH2 receptor and reduced binding affinity 70-fold. In contrast, this truncation increased affinity for the PTH1 receptor 10-fold, reversing the PTH2/PTH1 receptor binding selectivity and resulting in a high affinity interaction of TIP-(7-39) with the PTH1 receptor (6 nm). These findings can be explained by a strong interaction between the N-terminal region of TIP39 and the juxtamembrane domain of the PTH2 receptor, with the corresponding domain of the PTH1 receptor acting as a selectivity barrier against high affinity binding of TIP39. As a result, TIP-(7-39) is a highly potent, selective antagonist for the PTH1 receptor. PMID- 10854440 TI - A human SCO2 mutation helps define the role of Sco1p in the cytochrome oxidase assembly pathway. AB - Deficiencies in cytochrome oxidase, the terminal enzyme of the mitochondrial respiratory chain, are most often caused by an inability to complete assembly of the enzyme. Pathogenic mutations in SCO2, which encodes a cytochrome oxidase assembly factor, were recently described in several cases of fatal infantile cardioencephalomyopathy. To determine the molecular etiology of these disorders, we describe the generation and characterization of the parallel mutations in the homologous yeast SCO1 gene. We show that the E155K yeast sco1 mutant is respiration-competent, whereas the S240F mutant is not. Interestingly, the S240F mutation allows partial but incorrect assembly of cytochrome oxidase, as judged by an altered cytochrome aa(3) peak. Immunoblot analysis reveals a specific absence of subunit 2 from the cytochrome oxidase in this mutant. Taken together, our data suggest that Sco1p provides copper to the Cu(A) site on subunit 2 at a step occurring late in the assembly pathway. This is the first instance of a yeast cytochrome oxidase assembly mutant that is partially assembled. The S240F mutant also represents a powerful new tool with which to elucidate further steps in the cytochrome oxidase assembly pathway. PMID- 10854441 TI - Acute cadmium exposure inactivates thioltransferase (Glutaredoxin), inhibits intracellular reduction of protein-glutathionyl-mixed disulfides, and initiates apoptosis. AB - Oxidative stress broadly impacts cells, initiating regulatory pathways as well as apoptosis and necrosis. A key molecular event is protein S-glutathionylation, and thioltransferase (glutaredoxin) is a specific and efficient catalyst of protein SSG reduction. In this study 30-min exposure of H9 and Jurkat cells to cadmium inhibited intracellular protein-SSG reduction, and this correlated with inhibition of the thioltransferase system, consistent with thioltransferase being the primary intracellular catalyst of deglutathionylation. The thioredoxin system contributed very little to total deglutathionylase activity. Thioltransferase and GSSG reductase in situ displayed similar dose-response curves (50% inhibition near 10 micrometer cadmium in extracellular buffer). Acute cadmium exposure also initiated apoptosis, with H9 cells being more sensitive than Jurkat. Moreover, transfection with antisense thioltransferase cDNA was incompatible with cell survival. Collectively, these data suggest that thioltransferase has a vital role in sulfhydryl homeostasis and cell survival. In separate experiments, cadmium inhibited the isolated component enzymes of the thioltransferase and thioredoxin systems, consistent with the vicinal dithiol nature of their active sites: thioltransferase (IC(50) approximately 1 micrometer), GSSG reductase (IC(50) approximately 1 micrometer), thioredoxin (IC(50) approximately 8 micrometer), thioredoxin reductase (IC(50) approximately 0.2 micrometer). Disruption of the vicinal dithiol on thioltransferase (via oxidation to C22-SS-C25; or C25S mutation) protected against cadmium, consistent with a dithiol chelation mechanism of inactivation. PMID- 10854442 TI - A small molecule antagonist of chemokine receptors CCR1 and CCR3. Potent inhibition of eosinophil function and CCR3-mediated HIV-1 entry. AB - We describe a small molecule chemokine receptor antagonist, UCB35625 (the trans isomer J113863 published by Banyu Pharmaceutical Co., patent WO98/04554), which is a potent, selective inhibitor of CCR1 and CCR3. Nanomolar concentrations of UCB35625 were sufficient to inhibit eosinophil shape change responses to MIP 1alpha, MCP-4, and eotaxin, while greater concentrations could inhibit the chemokine-induced internalization of both CCR1 and CCR3. UCB35625 also inhibited the CCR3-mediated entry of the human immunodeficiency virus-1 primary isolate 89.6 into the glial cell line, NP-2 (IC(50) = 57 nm). Chemotaxis of transfected cells expressing either CCR1 or CCR3 was inhibited by nanomolar concentrations of the compound (IC(50) values of CCR1-MIP-1alpha = 9.6 nm, CCR3-eotaxin = 93.7 nm). However, competitive ligand binding assays on the same transfectants revealed that considerably larger concentrations of UCB35625 were needed for effective ligand displacement than were needed for the inhibition of receptor function. Thus, it appears that the compound may interact with a region present in both receptors that inhibits the conformational change necessary to initiate intracellular signaling. By virtue of its potency at the two major eosinophil chemokine receptors, UCB35625 is a prototypic therapy for the treatment of eosinophil-mediated inflammatory disorders, such as asthma and as an inhibitor of CCR3-mediated human immunodeficiency virus-1 entry. PMID- 10854443 TI - A novel tissue inhibitor of metalloproteinases-3 mutation reveals a common molecular phenotype in Sorsby's fundus dystrophy. AB - Sorsby's fundus dystrophy (SFD) is a dominantly inherited degenerative disease of the retina that leads to loss of vision in middle age. It has been shown to be caused by mutations in the gene for tissue inhibitor of metalloproteinases-3 (TIMP-3). Five different mutations have previously been identified, all introducing an extra cysteine residue into exon 5 (which forms part of the C terminal domain) of the TIMP-3 molecule; however, the significance of these mutations to the disease phenotype was unknown. In this report, we describe the expression of several of these mutated genes, together with a previously unreported novel TIMP-3 mutation from a family with SFD that results in truncation of most of the C-terminal domain of the molecule. Despite these differences, all of these molecules are expressed and exhibit characteristics of the normal protein, including inhibition of metalloproteinases and binding to the extracellular matrix. However, unlike wild-type TIMP-3, they all form dimers. These observations, together with the recent finding that expression of TIMP-3 is increased, rather than decreased, in eyes from patients with SFD, provides compelling evidence that dimerized TIMP-3 plays an active role in the disease process by accumulating in the eye. Increased expression of TIMP-3 is also observed in other degenerative retinal diseases, including the more severe forms of age-related macular degeneration, the most common cause of blindness in the elderly in developed countries. We hypothesize that overexpression of TIMP-3 may prove to be a critical step in the progression of a variety of degenerative retinopathies. PMID- 10854444 TI - Identification of a glutamic acid and an aspartic acid residue essential for catalytic activity of aspergillopepsin II, a non-pepsin type acid proteinase. AB - Aspergillopepsin II from Aspergillus niger var. macrosporus is a non-pepsin type or pepstatin-insensitive acid proteinase. To identify the catalytic residues of the enzyme, all acidic residues that are conserved in the homologous proteinases of family A4 were replaced with Asn, Gln, or Ala using site-directed mutagenesis. The wild-type and mutant pro-enzymes were heterologously expressed in Escherichia coli and refolded in vitro. The wild-type pro-enzyme was shown to be processed into a two-chain active enzyme under acidic conditions. Most of the recombinant mutant pro-enzymes showed significant activity under acidic conditions because of autocatalytic activation except for the D123N, D123A, E219Q, and E219A mutants. The D123A, E219Q, and E219A mutants showed neither enzymatic activity nor autoprocessing activity under acidic conditions. The circular dichroism spectra of the mutant pro- and mature enzymes were essentially the same as those of the wild-type pro- and mature enzyme, respectively, indicating that the mutant pro enzymes were correctly folded. In addition, two single and one double mutant pro enzyme, D123E, E219D, and D123E/E219D, did not show enzymatic activity under acidic conditions. Taken together, Glu-219 and Asp-123 are deduced to be the catalytic residues of aspergillopepsin II. PMID- 10854445 TI - Foreword PMID- 10854446 TI - Cancer Risk Communication: What We Know and What We Need to Learn. December 10 11, 1998. PMID- 10854447 TI - Workshop keynote address PMID- 10854448 TI - Introduction of section: challenges inherent in communicating cancer risk information PMID- 10854449 TI - Why (cancer) risk communication can be hard. AB - Effective risk communication uses audience members' time well by providing them with the information that they most need, in a form that they can easily comprehend. Accomplishing this task can be hard because of problems with both the transmitter and the receiver. The former must determine what is most worth saying. The latter must integrate that message with their often fragmentary mental models of the processes creating and controlling the risks. One strategy for improving communication is to use analytic methods for selecting the information to transmit, based on its criticality to recipients' decision making. A second strategy for improving communications is adapting the message to the cognitive processes of its recipients. Together, these strategies can reveal the limits to communication and how best to work within them. PMID- 10854450 TI - Improving cancer risk communication: a discussion of Fischhoff. PMID- 10854451 TI - What does it mean to understand a risk? Evaluating risk comprehension. AB - Risk communications are frequently intended to help people understand hazards they face, with the hope that this understanding will help them make better decisions about the need for action or help them choose among alternative actions. To evaluate the success of such communications, a definition of "understanding" is needed. This paper suggests that decisions about personal risks require, at a minimum, information about the nature and likelihood of potential ill effects, information about the risk factors that modify one's susceptibility, and information about the ease or difficulty of avoiding harm. Even if these attributes are accepted as essential criteria for understanding, research on risk perceptions suggests that assessing what people know or believe is sometimes quite difficult. The focus of the paper is on the several dimensions of risk comprehension. Examples of how each can be assessed are drawn from research on public perceptions of the risks from smoking. These examples demonstrate that the public has only a limited understanding of smoking risks. PMID- 10854452 TI - Introduction of section: overarching considerations in risk communications: romancing the message. PMID- 10854453 TI - Communicating health risk to ethnic groups: reaching Hispanics as a case study. PMID- 10854454 TI - Dealing with competing and conflicting risks in cancer communication. AB - Applied research on cancer risk communication is sparse, and even less is known about effective communication under conditions of multiple risks. This paper briefly describes the need and rationale for cancer risk communication, then describes what is known and needs to be known about communication addressing multiple risks. Its focus is on two specific communication issues: 1) comparing different risks and 2) prioritizing between multiple risks. There is considerable unmet need in cancer risk communication for new knowledge and recommendations for best practices. Those professionals choosing to pursue this work can make a significant contribution to the field. PMID- 10854455 TI - New directions for risk communication research: a discussion with additional suggestions. AB - The papers by Huerta and Macario and Kreuter share the theme of suggesting new directions for risk communication research in cancer prevention and control. Huerta and Macario remind us once again that sociocultural factors must be considered when conducting risk communication research on underserved populations. Of special note is their recommendation to target the family, which could introduce a compelling new chapter in risk communication research in cancer prevention and control. In contrast, Kreuter challenges us to consider multiple cancer risks and risk-reducing behaviors in our research and provides a provocative framework for achieving this goal. Given this common theme and the need to position specific recommendations within the larger context of other competing research questions, this paper also highlights several additional recommendations for future research. These recommendations include the following: more research on risk presentation; establishing guidelines for measuring risk; additional research testing strategies to de-bias optimistic and pessimistic perceptions of risk and evaluating risk communication as a strategy for behavior change; more research investigating the sociology of risk communication, with a special emphasis on the family as the unit of investigation; and, finally, more research that specifically targets underserved populations in diverse community settings. PMID- 10854456 TI - Introduction of section: enabling informed decisions about cancer risk. PMID- 10854457 TI - Treating people with information: an analysis and review of approaches to communicating health risk information. AB - The communication of risk information is a fundamental aspect of nearly all health promotion interventions. However, no consensus exists regarding the most effective way to provide people with risk information. We will review and evaluate the relative merits of two approaches to risk communication. One approach relies on the presentation of numerical information regarding the probability of a health problem occurring, whereas the other relies on the presentation of information about the antecedents and consequences of a health problem. Because people have considerable difficulty understanding and using quantitative information, the effectiveness of interventions that rely solely on numerical probability information has been limited. Interventions that provide people with a broader informational context in which to think about a health problem have had greater success systematically influencing perceptions of personal risk but have several important limitations. However, before any final conclusions can be drawn regarding the relative merits of different communication strategies, investigators must agree on the specific criteria that should be used to identify an effective intervention. PMID- 10854458 TI - Cancer screening decisions. AB - This review focuses on why people decide to obtain or to avoid screening for cancer. We discuss three topics: (a) physician prompts that may elicit compliant screening behavior, (b) the independent and joint effects of risk perceptions and worry, and (c) the costs and benefits of getting screened. Overall, the data suggest that each of these factors will influence screening. So, for example, people are more likely to seek screening if a physician recommends the behavior, if they feel personally vulnerable and worry a little about cancer, if insurance covers the screening, and if they believe that the test is an effective early detection procedure. Future research needs include studies comparing theories, longitudinal rather than cross-sectional studies, and true experiments. We also need to know more about why physicians are such powerful change agents and the trade-offs of increasing personal risk versus exacerbating worry. Practical recommendations for promoting cancer screening include encouraging physician interventions, explaining risk, and lowering the costs while emphasizing the benefits of screening. PMID- 10854459 TI - Risk communication in genetic testing for cancer susceptibility. AB - Risk communication is an integral part of genetic counseling and testing for cancer susceptibility. This paper reviews the emerging literature on this topic. Three relevant aspects of risk communication are addressed: communication of individual risk, communication of the risks inherent in genetic testing, and family communications related to risk. These studies suggest that (a) most individuals with some family history of cancer, including those at low to moderate risk, overestimate their personal cancer risk; (b) awareness of the risks of genetic testing is limited; (c) decision making about genetic testing is influenced strongly by exaggerated perceptions of personal cancer risk and less so by perceptions of the risks of genetic testing; (d) perceptions of personal risk of cancer are resistant to standard education and counseling approaches; (e) psychologic distress and coping processes influence the processing of risk information and subsequent decision making in genetic testing; and (f) family influences play an important role in risk awareness, genetic testing decisions, and outcomes. To study these issues further, new theoretical models and measures of risk perceptions need to be developed. Both observational and experimental methods should be used to examine both the content and process of risk communication in cancer genetic counseling and testing. Emotional, familial, and sociocultural influences on the risk communication process require special attention. PMID- 10854460 TI - Decision aids for patients considering options affecting cancer outcomes: evidence of efficacy and policy implications. AB - Some cancer screening and treatment decisions are not clear cut because outcomes are uncertain or options have different benefit/risk profiles. "Decision aids" have been developed as adjuncts to counseling so that patients can learn about benefits and risks, can consider their personal values, and can participate with their practitioner in decision making. The purpose of this paper is to review published evidence about the efficacy of decision aids focused on cancer outcomes and to outline research and dissemination issues. Studies evaluating cancer related decision aids demonstrate that they are acceptable to patients and help those who are uncertain at baseline to make choices. They also increase the likelihood that choices are based on better knowledge, realistic expectations of outcomes, and personal values. Decision aids reduce some dimensions of decisional conflict, and their effect on decisions is variable. Few studies examine the downstream effects of decision aids on long-term persistence with choices, regret, and quality of life. The differences between simpler and more intensive methods of decision support appear to be negligible in terms of knowledge and satisfaction as well as variable in terms of decisions and decisional conflict. However, more intensive methods are superior in terms of user acceptability and of the extent to which choices are based on realistic expectations and personal values. The clinical importance of these differences and the cost-effectiveness remain to be established. On the basis of this review, several recommendations for research are made, and dissemination issues are identified. PMID- 10854461 TI - Population risk, actual risk, perceived risk, and cancer control: a discussion. AB - Given the difficulty of converting population-based estimates of cancer risk into precise statements of individual risk, it is not surprising that (a) individual differences in risk perception are at best poorly correlated to the best available determination of "actual risk" and to behaviors to prevent and detect and treat cancer, and (b) success in bringing perceived risk into line with actual risk has been limited. These inconsistencies are of concern because individual perceptions of risk are thought to be important motivators of action for the prevention and early detection and treatment of cancer. Following the reviewer's suggestion that risk perceptions are readily influenced by contextual factors, we suggest examining risk perception in a self-regulatory framework in which both risk judgments and motivated action are products of underlying representations of cancer and the self. Self-assessments of risk may access only a part of the data necessary for motivation, whereas motivation to sustain action calls on a larger number of concrete features of the database (symptoms, time loss, consequences). Studies of cancer risk perception can make a major contribution to our understanding of processes involved in self-appraisals and self-management to maximize well-being and to avoid catastrophic disease. PMID- 10854462 TI - Matching strength of message to strength of evidence: a discussion. PMID- 10854463 TI - Introduction of section: persuasion for the purpose of cancer risk reduction: understanding responses to risk communications. AB - Risk behaviors and responses to risk communications are complex and multifaceted. Two target articles (1,2) conclude that little longitudinal evidence shows that risk perceptions predict precautionary behaviors. This paper focuses on several questions raised by these perplexing findings that have implications for future research on risk communications. A pressing need exists to understand how people process risk information over time and how such processing may differ as a function of risk status, individual differences, social context, and other factors. I will review evidence and methods from the study of persuasion and attitude change that suggest several ways to study message processing to understand what kinds of thoughts are brought to mind following a persuasive communication, as well as how such thoughts may be related to subsequent beliefs and behaviors as people encounter new information and make risk-relevant choices. PMID- 10854464 TI - The effect of risk communication on risk perceptions: the significance of individual differences. AB - The purpose of this paper is to address the literature on the relation between risk communication and the initiation of health behavior change. More specifically, we examine the evidence that providing risk information is an effective way to change risk perceptions, as well as the more limited evidence that altering risk perceptions influences risk behavior. The paper discusses significant developments in the research on these issues, describes specific studies that represent trends in this research, and discusses methodologic issues important to the development of the field. Although there are relatively few studies that demonstrate causal links between risk communication and behavior change, recent developments in the field point to the importance of tailoring risk communications to the individual characteristics of targets. Such tailoring has taken a variety of forms, including providing individualized feedback concerning risk status or genetic vulnerability and assessing readiness for behavior change. Future intervention efforts should combine individualized risk status feedback with assessment of individual differences in previous behavior and acceptance of personal vulnerability. PMID- 10854465 TI - Risk perception and risk communication for cancer screening behaviors: a review. AB - This review summarizes and synthesizes research findings on risk perception and risk communication related to cancer screening behaviors. The focus is on cancers for which there is evidence that screening reduces mortality, i.e., cervical, breast, and colorectal cancers. The following questions are addressed: 1) Is perceived risk associated with relevant cancer screening behaviors? 2) What factors are associated with perceived risk? 3) Is the relationship between perceived risk and cancer screening behaviors modified by other factors? 4) Have interventions to change perceived risk been effective in modifying risk perceptions? 5) Are these changes related to subsequent cancer screening behaviors? Methodologic issues are discussed, and future research needs are identified. There was consistent evidence that perceived risk was associated with mammography screening, but there were insufficient data on these associations for cervical or colorectal cancer screening behaviors. There was some evidence that perceived risk mediated the association between other variables and screening behaviors; however, because of the small number of studies, the findings are best viewed as hypothesis generating. Studies of interventions to modify risk perceptions provided some support for the view that they are modifiable, but there was conflicting evidence that these changes were related to subsequent cancer screening. Methodologic studies of how best to measure perceived risk are needed. Because most data on the correlates of perceived risk were cross sectional, it is difficult to determine whether perceived risk is a cause or an effect in relation to cancer screening. Longitudinal studies that measure perceived risk in defined populations with different cancer screening histories and that include follow-up for screening and repeated measurements of risk perception are needed to clarify this relationship. PMID- 10854466 TI - Persuasion for the purpose of cancer risk reduction: a discussion. AB - We comment on the preceding papers by Gerrard and Vernon concerning persuasion, perceived risk, and cancer-relevant behavior. Our purpose is to highlight several challenges for future investigators. First, relations between health cognition and health behavior (such as the link between perceived vulnerability and protective behaviors) are likely to be moderated by other variables, including individual differences and situational contexts. Second, we encourage health communication researchers to consider how persuasion is contextualized in social relationships and to employ mechanisms from the literature on social influence when promoting cancer prevention and early detection behaviors. Finally, we emphasize the importance of current feelings and anticipated emotions as motivators of salubrious actions. PMID- 10854467 TI - Introduction of section: implications for improving risk communication through various channels. PMID- 10854468 TI - Risk communication in clinical practice: putting cancer in context. AB - CONTEXT: Clinicians are increasingly urged-even mandated-to help patients make informed medical decisions by paying more attention to risk counseling. For many, the role of risk counseling is new and unfamiliar. This effort is made more difficult given the practical constraints created by 15-minute visits and competing demands (e.g., patient's chief complaint and institutional needs). OBJECTIVE: We detail a three-part approach for improving risk communication, acknowledging the role of clinicians, patients, and other communicators (i.e., media or public health agencies). PROPOSED APPROACH: Office-based tools to help clinicians do more. We suggest two ways to help make up-to-date estimates of disease risk and treatment benefit easily available during office visits. We propose the development of a comprehensive population database about disease risk and treatment benefit to be created and maintained by the federal government. Educating patients. We propose "Understanding Numbers in Health" a tutorial that reviews basic concepts of probability and their application to medical studies to help people become better critical readers of health information. Guidance for communicators. Finally, we propose a writer's guide to risk communication: a set of principles to help health communicators present data to the public clearly and objectively. CONCLUSION: In addition to tools to help clinicians better communicate risk information, serious efforts to improve risk communication must go beyond the clinic. Efforts that help the public to better interpret health risk information and guide communicators to better present such information are a place to start. PMID- 10854469 TI - Interactive multimedia and risk communication. AB - As our understanding of risk factors and their interaction with individual susceptibility to disease improves, general messages designed to communicate risk seem increasingly ineffective and often misleading. Risk messages communicated through the mass media cannot convey an individual's personal susceptibility to preventable diseases or the seriousness of these diseases. The advent of new media technologies allows us to better reach the public with programs tailored to the needs and interests of individual users. Although similar in outward appearance to mass media, programs delivered through the Internet, CD-ROM, and computer kiosks offer the potential for vastly improved efficacy in communicating risk. This paper outlines the potential uses of interactive multimedia within the traditional goals of risk communication. A significant research endeavor, coupled with stronger avenues for dissemination, is recommended to achieve the potential of new media in a timely manner. PMID- 10854470 TI - Is there a use for tailored print communications in cancer risk communication? AB - The manner of presentation of cancer risk information is critical to its understanding and acceptance by the individual recipient. Optimal communication of cancer risk information must effectively translate the technical meaning and subtleties of risk and its associated factors to a conceptual level understandable by the recipient. Tailored print communications (TPCs) may be an appropriate medium for cancer risk communication (CRC). TPCs are more refined than targeted communication materials. They are print materials created especially for an individual on the basis of knowledge about that person. The goal is to provide individually relevant and appropriate information. This review examines the nature of TPCs, assesses the use and potential of TPCs for the purpose of CRC, and highlights new directions in CRC. Articles dealing with TPCs were located by searching the MEDLINE(R) and PsychInfo(R) databases and seeking in-press articles. TPCs were identified for several areas of CRC, including dietary change, smoking cessation, mammography use, hormone replacement therapy, health risk appraisal, and genetic susceptibility to cancer. Although TPCs have been used in a number of different behavioral areas, they have not yet achieved their potential for CRC. The use of TPCs in the communication of cancer risk shows great promise, however, particularly as knowledge evolves regarding both the nature of risk and the most effective tailoring of health communication messages. PMID- 10854471 TI - The visual communication of risk. AB - This paper 1) provides reasons why graphics should be effective aids to communicate risk; 2) reviews the use of visuals, especially graphical displays, to communicate risk; 3) discusses issues to consider when designing graphs to communicate risk; and 4) provides suggestions for future research. Key articles and materials were obtained from MEDLINE(R) and PsychInfo(R) databases, from reference article citations, and from discussion with experts in risk communication. Research has been devoted primarily to communicating risk magnitudes. Among the various graphical displays, the risk ladder appears to be a promising tool for communicating absolute and relative risks. Preliminary evidence suggests that people understand risk information presented in histograms and pie charts. Areas that need further attention include 1) applying theoretical models to the visual communication of risk, 2) testing which graphical displays can be applied best to different risk communication tasks (e.g., which graphs best convey absolute or relative risks), 3) communicating risk uncertainty, and 4) testing whether the lay public's perceptions and understanding of risk varies by graphical format and whether the addition of graphical displays improves comprehension substantially beyond numerical or narrative translations of risk and, if so, by how much. There is a need to ascertain the extent to which graphics and other visuals enhance the public's understanding of disease risk to facilitate decision-making and behavioral change processes. Nine suggestions are provided to help achieve these ends. PMID- 10854472 TI - Risky business--communicating scientific findings to the public. PMID- 10854473 TI - Living can be hazardous to your health: how the news media cover cancer risks. AB - For more than two decades, the news media has bombarded the public with often conflicting information about health risks, contributing to an atmosphere of hype and hysteria about cancer and other diseases. Improvements in media reporting of health risks require greater efforts by both those who cover the news and those who create it. Guidelines for bringing more perspective and balance to media coverage of risk are provided. These include putting cancer in context with other diseases, explaining absolute and relative risks, differentiating between individual and population risks, stressing the degree of uncertainty of new research and how it fits with previous data, covering the process as well as end results of science, understanding different media constraints and needs, and taking into account the diverse backgrounds and needs of the target audience-the general public. PMID- 10854474 TI - Communicating cancer risk in print journalism. AB - The current barrage of information about real and potential cancer risks has created undue fears and misplaced concerns about cancer hazards faced by Americans. Most members of the general public are far more worried about minuscule, hypothetical risks presented by environmental contaminants than about the far greater well-established hazards that they inflict on themselves, for example, through smoking, dietary imbalance, and inactivity. It is the job of the print media to help set the record straight and to help place in perspective the myriad cancer risks that are aired almost weekly in 30-second radio and television broadcasts. PMID- 10854475 TI - Challenges to improving health risk communication in the 21st century: a discussion. PMID- 10854476 TI - Implications for improving risk communication through various channels: a discussion. AB - The news media need to improve their coverage of new and old cancer risks, avoiding sensational reporting of minor risks and underreporting of major ones. However, efforts to communicate about cancer risk to the general public need to go far beyond the traditional print and broadcast outlets into more innovative, personalized channels that can better educate individuals about the risks they face and what they can do about them. PMID- 10854477 TI - Introduction of section: breakout session reports. PMID- 10854478 TI - Cancer risk communication-what we need to learn. PMID- 10854479 TI - Cancer risk communication-what we know. PMID- 10854480 TI - Discovery of cancer susceptibility genes: study designs, analytic approaches, and trends in technology. AB - Determining the genetic causes of cancers has immense public health benefits, ranging from prevention to earlier detection and treatment of disease. Although a number of cancer susceptibility genes have been successfully identified, design and analytic issues remain that challenge the current paradigm of gene discovery. Some examples are the definition and measurement of cancer phenotype, the use of intermediate end points, the choice of sample (e.g., affected relative pairs versus large extended pedigrees), the choice of analytic method [e.g., parametric logarithm of the odds (LOD) score method versus model-free methods], and the influence of gene-environment interaction on linkage analysis. Furthermore, association methods, based on either the traditional case-control study design or family-based controls, are popular choices to evaluate candidate genes or screen for linkage disequilibrium. Finally, the study design and analytic methods for gene discovery are determined to some extent by what genomic technology is feasible within the laboratory. Many of the main issues related to gene discovery, as well as trends in genomic technology that will impact on gene discovery, are discussed from the perspective of their strengths and weaknesses, pointing to areas in need of further work. PMID- 10854481 TI - Design of gene characterization studies: an overview. AB - This collection of papers from the Gene Characterization Panel addresses design issues in studies aimed at assessing the population characteristics of cloned genes, such as their allele frequencies, penetrance, variation in these parameters across subpopulations, and gene-environment and gene-gene interactions. This paper provides an overview of the various designs that have been suggested, including cohort and case-control designs using independent and related individuals as well as optimal multistage sampling and hybrid designs. Various statistical (bias and efficiency) and practical considerations are suggested for evaluation of the alternative designs, with the aim of posing the question, "What is the optimal design for a particular situation"? The answer to this question clearly depends on such contextual issues as nature of the outcome variable, the gene frequency and genetic relative risk, and the importance of gene-environment and gene-gene interactions. Further methodologic work might be usefully directed toward assessment of the seriousness of the population stratification problem in general as well as methods of dealing with it, the utility of registries of high-risk families, and the merits of various hybrid designs for gene discovery and gene characterization. PMID- 10854482 TI - Case-control studies of common alleles and environmental factors. AB - It is clear from descriptive and migration studies that most cancer is environmental in origin. Descriptive, case-control and cohort studies have provided the foundation for our understanding of the environmental component of cancer etiology as well as most major causes of morbidity and mortality. We propose that the same epidemiologic methods that have provided fundamental insight into the etiology of cancer in the general population are optimally suited to study the impact of relatively common polymorphisms on chronic disease incidence. In this article, we describe the role of case-control studies in assessing the effects of genes in disease. Some of the advantages and disadvantages of the case-control design, particularly as an alternative to case control studies nested in a cohort in the context of the study of complex disease, are described. PMID- 10854483 TI - Family-based association studies. AB - We review case-control designs for studying gene associations in which relatives of case patients are used as control subjects. These designs have the advantage that they avoid the problem of population stratification that can lead to spurious associations with noncausal genes. We focus on designs that use sibling, cousin, or pseudosibling controls, the latter formed as the set of genotypes not transmitted to the case from his or her parents. We describe a common conditional likelihood framework for use in analyzing data from any of these designs and review what is known about the validity of the various design and analysis combinations for estimating the genetic relative risk. We also present comparisons of efficiency for each of the family-based designs relative to the standard population-control design in which unrelated controls are selected from the source population of cases. Because of overmatching on genotype, the use of sibling controls leads to estimates of genetic relative risk that are approximately half as efficient as those obtained with the use of population controls, while relative efficiency for cousin controls is approximately 90%. However, we find that, for a rare gene, the sibling-control design can lead to improved efficiency for estimating a G x E interaction effect. We also review some restricted designs that can substantially improve efficiency, e.g., restriction of the sample to case-sibling pairs with an affected parent. We conclude that family-based case-control studies are an attractive alternative to population-based case-control designs using unrelated control subjects. PMID- 10854484 TI - Cohort studies for characterizing measured genes. AB - We describe the advantages of using established cohort studies that have collected blood samples to investigate the role of genes in the etiology of cancer. These studies include the cost-efficiency and reliability of nested case control substudies from the cohort for exploration of gene-disease associations and gene-environment interactions as well as gene penetrance. Also, the cohort may serve as a well-defined "mini-population" from which to study population stratification and molecular markers of ethnicity. We conclude that cohort studies can play a significant role in assessing the role of genetic markers for common tumors or multiple cancer sites. PMID- 10854485 TI - Multistage sampling for disease family registries. AB - BACKGROUND: The objectives of a family-based disease registry range from characterizing measured genetic factors and gene-environment interaction effects to detecting novel susceptibility genes. Gathering complete information on exposure and disease status in all family members for a sample of affected subjects (probands) to address these diverse objectives would be prohibitively expensive. METHODS: Multistage sampling can be used to design an efficient family based disease registry. At each stage, the probands are classified on the basis of previously collected data, and a subsample is selected for more detailed observation. The design can be optimized to minimize the variance of any of the model parameter estimates, subject to a constraint on the total sample size. RESULTS: We describe the basic statistical theory and its application to a four stage sampling scheme proposed for the Cooperative Family Registry for Epidemiologic Studies of Colorectal Cancer at the University of Southern California. PMID- 10854486 TI - Detection of interaction involving identified genes: available study designs. AB - Advances in molecular genetic techniques have led to an increased ability to examine gene-environment interactions. Studies to detect gene-environment interactions are motivated by different situations, including 1) most identified cancer genes having associated lifetime risks less than 100% (i.e., incomplete penetrance), 2) hereditary factors that control the metabolism of carcinogens that may modulate risk of disease as hypothesized in pharmacogenetics, and 3) inconsistent associations across studies between a cancer and a suspected risk factor. The above situations and others have led to increased study of interaction between genetic and environmental factors. Less studied so far, but with increased potential for the future, is interaction between identified genes. Gene-gene interaction studies would also be motivated by the situations described above. Approaches to detect gene-environment and gene-gene interactions are reviewed. Available risk estimates, required types of subjects, and feasibility of the proposed study designs are discussed; efficiency and power for interaction assessment are summarized where available. In general, most designs allow for estimating risk associated with a genetic factor, environmental factor, and interaction effect. Although power and efficiency for detecting interactions have been assessed for specific situations in some of the methods, further investigations are needed to define the efficiency spectra of each design. PMID- 10854487 TI - Kin-cohort designs for gene characterization. AB - BACKGROUND: In the kin-cohort design, a volunteer with or without disease (the proband) agrees to be genotyped, and one obtains information on the history of a disease in first-degree relatives of the proband. From these data, one can estimate the penetrance of an autosomal dominant gene, and this technique has been used to estimate the probability that Ashkenazi Jewish women with specific mutations of BRCA1 or BRCA2 will develop breast cancer. METHODS: We review the advantages and disadvantages of the kin-cohort design and focus on dichotomous outcomes, although a few results on time-to-disease onset are presented. We also examine the effects of violations of assumptions on estimates of penetrance. We consider selection bias from preferential sampling of probands with heavily affected families, misclassification of the disease status of relatives, violation of Hardy-Weinberg equilibrium, violation of the assumption that family members' phenotypes are conditionally independent given their genotypes, and samples that are too small to ensure validity of asymptotic methods. RESULTS AND CONCLUSIONS: The kin-cohort design has several practical advantages, including comparatively rapid execution, modest reductions in required sample sizes compared with cohort or case-control designs, and the ability to study the effects of an autosomal dominant mutation on several disease outcomes. The design is, however, subject to several biases, including the following: selection bias that arises if a proband's tendency to participate depends on the disease status of relatives, information bias from inability of the proband to recall the disease histories of relatives accurately, and biases that arise in the analysis if the conditional independence assumption is invalid or if samples are too small to justify standard asymptotic approaches. PMID- 10854488 TI - Study design in genetic epidemiology: theoretical and practical considerations. AB - Recent advances in molecular genetics have created new opportunities and challenges for genetic epidemiologists. Here we review some of the issues that arise when designing a study involving the genetic epidemiology of chronic diseases of late onset, such as cancer. We discuss two considerations that influence the choice of design. The first consideration is the study's goals. We describe the goals of identifying new susceptibility genes for a disease, of estimating important characteristics of known genes, and of learning how to prevent the disease in the genetically susceptible. We indicate how these goals affect the choice of design and present some guidelines for choosing designs that effectively address them. The second consideration is the set of practical constraints to successfully conducting the research. These contraints include problems of potential selection bias, reduced response rates, problems particular to family registries, problems particular to the cultures of various ethnic groups, and ethical issues. We indicate how these constraints affect the choice of design and discuss ways to deal with them. PMID- 10854489 TI - Integrated designs for gene discovery and characterization. AB - Recent advances, including near completion of the human genome map, ever improving high-throughput technologies, and successes in discovering chronic disease-related genes, have stimulated the further development of genetic epidemiology. The primary mission of genetic epidemiology is to discover and characterize genes, whether independent of or interactive with environmental factors, that cause human diseases. To accomplish such a mission, genetic epidemiology needs to integrate both genetic and epidemiologic approaches. One of the challenges facing such an integrated approach is the identification of study designs that are efficient for both gene discovery and characterization. Because designs for gene discovery alone and designs for gene characterization alone have been elaborated in the other two panels, the focus of this paper is to describe those designs that may be useful for discovery and characterization jointly, including case-family and case-control-family designs. Examples of integrated designs are described, and studies of breast cancer conducted at the Fred Hutchinson Cancer Research Center are used for illustration. Finally, related analytic issues are also discussed. PMID- 10854490 TI - Confirmation of prostate cancer susceptibility genes using high-risk families. AB - Data from many types of studies support the hypothesis that strong familial components are involved in the etiology of prostate cancer. One way to access such genes is through the study of families with multiple affected family members and, in particular, families with individuals affected comparatively early in life. Several prostate cancer susceptibility loci have been described to date. Confirmation of the linkage and estimation of the proportion of families who are linked in large independent datasets is essential to understanding the significance of susceptibility genes. We explore the methodology used to perform such studies and the factors that can limit the ability to confirm linkage results. We report specifically the example of the HPC1 gene on 1q24-25. PMID- 10854491 TI - Study-design issues in the development of the University of Southern California Consortium's Colorectal Cancer Family Registry. AB - The University of Southern California Consortium is a participating center in the National Cancer Institute's Collaborative Family Registry for Colorectal Cancer Studies (CFRCCS). Because data collection takes time, money, and effort, all of which are in short supply, we first defined our research objectives and then attempted to design our registry to enable us to address these objectives in an efficient manner. We decided on a family-based design, and our objectives are to characterize cloned genes that are generally accepted causes of colorectal cancer, to assess putative candidate genes, to map new genes, and to conduct prevention trials in high-risk subjects. For the gene characterization objectives, our primary aim is to estimate gene frequency and penetrance, with a secondary aim to investigate factors that may affect penetrance (allele-specific effects plus gene-gene and gene-environment interactions). We describe a multiple stage design to select families into the registry. After a family is selected into the registry, we collect questionnaire data and blood samples on selected subjects only, and we tailor data collection decisions to each family (given who is affected and who is available) to optimize power per unit effort and cost. We also discuss practical decisions faced by our registry, including 1) defining a reference period for use in questionnaires; 2) deciding whether or not to establish cell lines and, if so, on whom; and 3) determining which cases should be tested for microsatellite instability. Finally, we address the appropriate use of data derived from high-risk clinics, within more broadly defined, population based research. PMID- 10854492 TI - Design and analysis issues in a population-based, case-control-family study of the genetic epidemiology of breast cancer and the Co-operative Family Registry for Breast Cancer Studies (CFRBCS). AB - BACKGROUND: Historically, studies of the "genetic epidemiology" of cancer have used nonsystematically sampled kindreds with numerous cases of cancer across multiple generations. From the epidemiologic viewpoint, it is difficult to extrapolate findings to the population because of the ad hoc ascertainment of these atypical, ill-defined families. Since 1992, we have been conducting a population-based, case-control-family study of breast cancer. METHODS: Families are identified through a single, population-sampled proband, who is either affected or unaffected, making adjustment for ascertainment straightforward. Administered questionnaires and blood samples are sought from cases, controls, and specified sets of relatives. From 1996 through 1999, a further 1200 case families have been recruited as part of the Co-operative Family Registry for Breast Cancer Studies (CFRBCS). Issues relevant to the study design and analysis are discussed. RESULTS: Epidemiologic and genetic findings published to date are summarized. In particular, this population-based study has shown that the so called "high-risk" families containing multiple cases of breast cancer are not typical of families in the general population in which BRCA1 or BRCA2 mutations are segregating. Most "hereditary" cancers are "sporadic." CONCLUSION: The collection of DNA, as well as data on disease status and risk factors, from population-sampled sets of relatives provides a powerful resource for addressing genetic and environmental determinants of cancer. A population-based multicenter, multidisciplinary enterprise, such as has been developed by the CFRBCS, may become a model for future research in cancer epidemiology, allowing genetic and environmental risk factors to be put into a proper population perspective. PMID- 10854493 TI - Combined influence of genetic and dietary factors on colorectal cancer incidence in Japanese Americans. AB - Colorectal cancer (CRC) rates for Japanese migrants to the United States increased rapidly to surpass the level of the host population. CRC rates for the Japanese in Hawaii and California are now the highest in the world. Rates for this disease have also increased in Japan, presumably as the result of the westernization of the diet. A series of population-based studies in Hawaii was undertaken to determine which dietary factors are responsible for this remarkable susceptibility of the Japanese to CRC and whether genetic factors are also involved. A first case-control study suggested that a high intake of red meat is a major risk factor for the disease in Hawaii Japanese men and that family history of CRC among first-degree relatives may strongly modify this association. A case-control family study is currently being completed to explore further the interaction between family history and the intake of red meat after adjustment for environmental covariates among family members. Also, a segregation analysis will guide gene discovery studies among high-risk Japanese families being recruited in the Cooperative Family Registry for Colorectal Cancer Studies. Retrospective and prospective studies are also ongoing to test associations of CRC with specific polymorphisms in genes controlling the metabolic activation or detoxification of the carcinogens associated with a diet high in red meat. Preliminary results suggest an association of the combined rapid NAT2 genotype and rapid CYP1A2 phenotype with CRC in individuals consuming well-done red meat. Populations in which dramatic changes in cancer incidence have occurred may offer opportunities to identify gene-environment interactions. PMID- 10854494 TI - Occupational risk factors for shoulder pain: a systematic review. AB - OBJECTIVES: To systematically evaluate the available evidence on occupational risk factors of shoulder pain. METHODS: Relevant reports were identified by a systematic search of Medline, Embase, Psychlit, Cinahl, and Current Contents. The quality of the methods of all selected publications was assessed by two independent reviewers using a standardised checklist. Details were extracted on the study population, exposures (physical load and psychosocial work environment), and results for the association between exposure variables and shoulder pain. RESULTS: 29 Studies were included in the review; three case control studies and 26 cross sectional designs. The median method score was 60% of the maximum attainable score. Potential risk factors related to physical load and included heavy work load, awkward postures, repetitive movements, vibration, and duration of employment. Consistent findings were found for repetitive movements, vibration, and duration of employment (odds ratio (OR) 1.4-46 in studies with method scores >/= 60%). Nearly all studies that assessed psychosocial risk factors reported at least one positive association with shoulder pain, but the results were not consistent across studies for either high psychological demands, poor control at work, poor social support, or job dissatisfaction. Studies with a method score >/=60% reported ORs between 1.3 and 4.0. Substantial heterogeneity across studies for methods used for exposure assessment and data analysis impeded statistical pooling of results. CONCLUSIONS: It seems likely that shoulder pain is the result of many factors, including physical load and the psychosocial work environment. The available evidence was not consistent across studies, however, and the associations were generally not strong. Future longitudinal research should evaluate the relative importance of each individual risk factor and the role of potential confounding variables-such as exposure during leisure time-to set priorities for the prevention of shoulder pain in occupational settings. PMID- 10854495 TI - Rheumatoid arthritis in workers exposed to silica in the pottery industry. AB - OBJECTIVE: To investigate the relation between rheumatoid arthritis and occupational exposure to silica in pottery and related industries. METHODS: Medical records of 8325 men and women born 1916-45 and employed in pottery, refractory material (aluminosilicate or silica), and sandstone industries were examined to identify cases of rheumatoid arthritis. Medical and employment histories were extracted for cases and matched referents. Indices of duration, cumulative exposure, and mean silica concentration were compiled. Conditional logistic regression was used to investigate the relation between rheumatoid arthritis and indices of exposure, having allowed for potential confounders of smoking, employment in the coal mining industry, and number of pregnancies. RESULTS: 58 Cases of rheumatoid arthritis (43 men, 15 women) were identified. Cases had significantly shorter duration of exposure than referents. There was no significant difference between cases and referents in mean silica concentration. Men who had worked in the coal mining industry were particularly at risk (odds ratio 5.36, 95% confidence interval 1.92 to 15.03). CONCLUSION: There was no evidence of increased risk of developing rheumatoid arthritis after occupational exposure to silica at mean exposures within the current United Kingdom exposure limits. PMID- 10854496 TI - Prevalence of Raynaud's phenomenon in Great Britain and its relation to hand transmitted vibration: a national postal survey. AB - OBJECTIVES: To assess the prevalence of Raynaud's phenomenon in the general population of Great Britain and to estimate the proportion and number of cases attributable to hand transmitted vibration (HTV). METHODS: A questionnaire was posted to a random sample of 22,194 adults of working age. Information was collected on the lifetime prevalence of finger blanching, smoking habits, and occupational and leisure time exposures to HTV. Associations with risk factors were explored by logistic regression, with odds ratios converted into prevalence ratios (PRs). RESULTS: Among the 12 907 respondents, 1835 (14.2%) reported finger blanching at some time, including 1529 (11. 8%) in whom symptoms were induced by cold, and 597 (4.6%) in whom the blanched area was also clearly demarcated. Prevalences were higher in women than men. Around one fifth of cases (2% of respondents) had consulted a doctor about their symptoms. By comparison with men who had never been exposed to HTV, the PR for cold induced blanching in those exposed only at work was 2.0 (95% CI 1.7 to 2.3), and in men exposed both at work and in leisure it was 2. 5 (95% CI 2.1 to 3.1). Higher risks were found in men who consulted a doctor about cold induced blanching, among whom 37.6% of cases were estimated to arise from exposure to HTV. The estimated number of cases attributable to HTV nationally was 222 000 in men who reported extensive blanching (blanching affecting at least eight of the digits or 15 phalanges). Similar patterns of risk were found in women, but the attributable proportion was much lower (5.3% in cases consulting a doctor). CONCLUSIONS: Raynaud's phenomenon is common in the general population. Many cases are attributable to HTV, especially in men, emphasising the public health importance of this common occupational hazard. PMID- 10854497 TI - Relation between immune variables and burnout in a sample of physicians. AB - OBJECTIVES: To evaluate in a group of physicians the relation between burnout, demographic or job characteristics, anxiety, and immune variables. METHODS: Seventy one physicians of all grades were recruited among different departments to a cross sectional survey. The Maslach burnout inventory, scales of emotional exhaustion, depersonalisation, and personal accomplishment, the trait scale of anxiety inventory (STAI-Y2), and a questionnaire on personal and professional characteristics were administered. The immune profile included quantitative (number (%) of lymphocytes and subsets) and functional (natural killer cytotoxicity) measures. RESULTS: With a model of stepwise multiple regression analysis, emotional exhaustion was significantly affected by both personal (marital, sex) and job characteristics (qualification, working activity), whereas only patient contact explained a portion of variation in depersonalisation. Furthermore, trait anxiety was found to predict the Maslach burnout inventory scores. After correction for potential confounders, physicians who scored high levels of personal accomplishment showed significantly higher numbers of total lymphocytes, T cells (CD3), T helper cells (CD4), and T suppressor cells (CD8) than those who scored low levels. No other correlation was found between burnout and immune variables. CONCLUSIONS: In our group of relatively young physicians a high degree of personal accomplishment was associated with an increase in the number of peripheral lymphocytes, particularly T subsets. The meaning of this is not clear, although it could be speculated that to evaluate oneself positively, particularly with regard to work with patients in the health services, might help to stimulate the immune system. By contrast, there is no evidence that to work hard, to feel tired from work, and to have a cynical reaction towards patient care is related to immunosuppression. PMID- 10854498 TI - Health complaints and immunological markers of exposure to bioaerosols among biowaste collectors and compost workers. AB - OBJECTIVES: In a cross sectional study, work related health complaints and diseases of 58 compost workers and 53 biowaste collectors were investigated and compared with 40 control subjects. Levels of specific IgG antibodies to moulds and bacteria were measured as immunological markers of exposure to bioaerosols. METHODS: With a standardised protocol, the participants of the study were interviewed for work related symptoms, conditions of exposure to bioaerosols at their workplaces, exposure to bioaerosols from other sources, atopic diseases, and smoking habits. They were clinically examined by physicians specialised in occupational medicine. Also, concentrations of specific IgG antibodies against antigens of moulds and actinomycetes occurring regularly at these workplaces were measured and compared with the health complaints of the workers. RESULTS: Compost workers had significantly more symptoms and diseases of the airways (p=0.003) and the skin (p=0.02) than the control subjects. Health complaints of biowaste collectors did not differ significantly from those of the control group. Subjects with atopic diseases were underrepresented in the compost workers (p=0.003). Significantly increased antibody concentrations against fungi and actinomycetes were measured in workers at composting plants. The concentrations in biowaste collectors did not differ significantly from those in the control subjects. A significant association between the diseases and increased antibody concentrations were found in the compost workers. CONCLUSION: The high exposure to bioaerosols of compost workers is significantly associated with a higher frequency of health complaints and diseases as well as higher concentrations of specific antibodies against moulds and actinomycetes. A healthy worker effect is indicated by the underrepresentation of atopic diseases among the compost workers compared with biowaste collectors and the control group. PMID- 10854499 TI - 109Cd K x ray fluorescence measurements of tibial lead content in young adults exposed to lead in early childhood. AB - OBJECTIVES: Tibia lead measurements were performed in a population of 19-29 year old people who had been highly exposed to lead in childhood to find whether lead had persisted in the bone matrix until adulthood. METHODS: (109)Cd K x ray fluorescence was used to measure the tibia lead concentrations of 262 exposed subjects and 268 age and sex matched controls. Questionnaire data allowed a years of residence index to be calculated for exposed subjects. A cumulative blood lead index was calculated from the time weighted integration of available data of blood lead. RESULTS: The mean (SEM) difference between exposed and control men was 4.51 (0.35) micrograms Pb/g bone mineral, and between exposed and control women was 3.94 (0. 61) micrograms Pb/g bone mineral. Grouped mean bone lead concentrations of exposed subjects were predicted best by age. When exposed and control subjects' data were combined, grouped mean bone lead concentrations were predicted best by cumulative blood lead index. The years of residence index was neither a good predictor of bone lead concentrations for exposed subjects nor for exposed and control subjects combined. Finally, exposed subjects had increased current blood lead concentrations that correlated significantly with bone lead values. CONCLUSION: Bone lead concentrations of exposed subjects were significantly increased compared with those of control subjects. Lead from exposure in early childhood had persisted in the bone matrix until adulthood. Exposed subjects had increased blood lead concentrations compared with controls. Some of this exposure could be related to ongoing exposure. However, some of the increase in blood lead concentration in adult exposed subjects seemed to be a result of endogenous exposure from increased bone lead stores. The endogenous exposure relation found for men was consistent with reported data, but the relation found for women was significantly lower. Further research is needed to find whether the observed differences are due to sex, or pregnancy and lactation. PMID- 10854500 TI - Personal exposures of children to nitrogen dioxide relative to concentrations in outdoor air. AB - OBJECTIVES: To investigate the relation between fluctuations in personal exposure to nitrogen dioxide (NO(2)) in school children and changes in outdoor NO(2) concentrations. METHODS: 114 Asthmatic school children aged 7-12 years were recruited from the Southampton area. Weekly average personal exposures to NO(2) were measured over a 13 month period with passive diffusion tubes. At the same time, outdoor NO(2) concentrations were monitored at a fixed site in the centre of Southampton. Correlations between weekly personal exposures and mean outdoor concentrations during the same periods were examined. RESULTS: Mean duration of follow up was 32 weeks. Measurements of weekly mean personal NO(2) exposures were generally low and ranged from 2.47 to 1751 [corrected] micrograms/m(3) with a geometric mean of 60 [corrected] micrograms/m(3). Substantial variation in personal exposures occurred between children and more especially within individual children from week to week. Daily outdoor concentrations of NO(2) ranged from 15.2 to 105.2 [corrected] micrograms/m(3), with a geometric mean of 43.4 [corrected] micrograms/m(3). There was no evidence of seasonal variation in outdoor concentrations. No significant correlation was found between each child's weekly mean personal exposures to NO(2) and mean outdoor concentrations for the corresponding periods. CONCLUSION: At low outdoor NO(2) concentrations, fluctuations in NO(2) in outdoor air as measured at a central monitoring station do not contribute importantly to variations in personal exposure when averaged over a week. PMID- 10854501 TI - Respiratory diseases in children and outdoor air pollution in Sao Paulo, Brazil: a time series analysis. AB - OBJECTIVES: To investigate the short term effects of air pollution on the respiratory morbidity of children living in Sao Paulo, Brazil, one of the largest cities in the developing world. METHODS: Daily counts of hospital admissions due to respiratory diseases along with daily levels of meteorological variables and air pollutants (PM(10), SO(2), NO(2), O(3), and CO) were analysed with Poisson regression. Final models were adjusted for the effects of time trends, seasonal patterns, weekdays, holidays, meteorological factors, and serial correlation. RESULTS: Daily admissions of children to hospital for total respiratory disease and pneumonia showed significant increases associated with O(3) (5-8%), NO(2) (9%), and with PM(10) (9%) (results are for an increase from the 10th to the 90th percentile of pollution measurements). Consistently, effects for pneumonia were greater than for all respiratory diseases combined. Also, effects on infants (children <1 year old) presented higher estimates. Similar associations were found for asthma admissions. Point estimates for most pollutants were higher for asthma than for other diagnosed admissions. However, these associations were not significant. CONCLUSIONS: These results agree with the limited publications on this subject but indicate a rather smaller magnitude of effects. Nevertheless, given the present concentrations of air pollution in Sao Paulo and the large population potentially exposed attention should be directed to minimise such effects. PMID- 10854502 TI - Mortality from cancer and chronic respiratory diseases among workers who manufacture carbon electrodes. AB - OBJECTIVES: To investigate the risk of cancer and non-neoplastic respiratory diseases among workers who manufacture carbon electrodes, as this industry entails exposure to mixtures of polycyclic aromatic hydrocarbons. METHODS: A historical cohort study was carried out of 1006 male workers employed for at least 1 year between 1945 and 1971 in a carbon (graphite) electrode production plant in central Italy, who were followed up for mortality between 1955 and 1996. The ratio of observed to expected deaths (standardised mortality ratios, SMRs) was computed from both national and (for the period 1964-96) regional age and period specific mortalities. A multivariate Poisson regression analysis was performed to investigate the relative risk (RR) of death according to duration of employment and time since first employment in the factory. RESULTS: A total of 424 workers had died, 538 were still alive, and 44 were lost to follow up. Mortalities from all causes, all cancers, and respiratory tract cancer were in line with the regional figure. An excess was found over the expected deaths from skin cancer including melanoma (SMR 3.16, 95% confidence interval (95% CI) 0.65 to 9.23) and from non-neoplastic respiratory diseases (SMR 1.58, 95% CI 1.16 to 2.11). Poisson regression analysis including age as a covariate showed an increased risk of dying from gastric cancer with increasing duration of employment, and an increase in the RR of dying from lung cancer and from non neoplastic respiratory diseases with increasing time since first employment, although the linear trend was not significant. CONCLUSION: This study supports previous findings that working in the carbon electrode manufacturing industry may not increase the risk of dying from respiratory cancer. However, a possible association with non-malignant respiratory diseases cannot be excluded. PMID- 10854503 TI - Future trends in mortality of French men from mesothelioma. AB - OBJECTIVES: Previous projections of mortality from mesothelioma among French men have used the age-generation method, based on the Poisson regression model. In this study an alternative method to model mortality from mesothelioma was used to predict its future trend: this method was based on the risk function that links this mortality to past exposure to asbestos, combined with population exposure data. METHOD: Data on past French asbestos imports were used to model the overall past exposure to asbestos in men and assess two extreme scenarios (optimistic and pessimistic) for its future trends. The number of male deaths occurring between the ages of 50 and 79, from 1997-2050, was then calculated with the risk function for mesothelioma. RESULTS: The results showed that mortality from mesothelioma among French men aged 50-79 will continue to increase, reaching a peak averaging between 1140 (optimistic scenario) and 1300 deaths (pessimistic scenario) annually around the years 2030 and 2040, respectively. No preventive measures applied now will affect this trend before then. These results are similar to those of two other predictions of mortality from mesothelioma among French men: a peak around 2030 of 800-1600 deaths annually among men aged 25-89 years, and a peak around 2020 of 1550 deaths annually among men aged 40-84. CONCLUSIONS: Our results indicate that between 1997 and 2050, the most optimistic and pessimistic trends of future exposure will lead to the deaths from mesothelioma of between 44 480 and 57 020 men, with a corresponding loss of from 877 200 to 1 171 500 person years of life. PMID- 10854504 TI - Lung and bladder cancer among workers in a Norwegian aluminium reduction plant. AB - OBJECTIVE: To investigate the relation between exposure to polycyclic aromatic hydrocarbons (PAHs) and the incidence of lung and bladder cancer among aluminium production workers. METHODS: The cohort comprised 1790 men employed for more than 5 years at a Norwegian aluminium plant contributing 36 587 person-years to the study. Historical exposure to PAHs was estimated by the use of industrial hygiene measurements and by a panel of three people familiar with the industry. Cancer incidence was investigated from 1953 to 1995. The observed cases of cancer among men were compared with expected numbers calculated from national rates for men, and dose-response relations were investigated by internal comparison by Poisson regression with age, period, smoking, and cumulative exposure included in the models. The effect of lagging exposure by 10, 20, and 30 years was also investigated. RESULT: The present study showed no increased risk of urinary bladder cancer or lung cancer with increasing cumulative exposure to PAHs. No significant changes in risk were found for different lag times. CONCLUSIONS: Due to the small size of this study, a minor increase in risk could not be excluded. PMID- 10854505 TI - Health law and policy: a survival guide, to medicolegal issues for practitioners PMID- 10854506 TI - Evaluation in occupational health practice, 1st edition PMID- 10854507 TI - Occupational health: recognising and preventing work related disease and injury, 4th edition PMID- 10854508 TI - Instant notes in immunology PMID- 10854509 TI - Environmental toxicants: human exposures and their health effects PMID- 10854510 TI - The 5-minute toxicology consult PMID- 10854511 TI - The dictionary of substances and their effects PMID- 10854512 TI - Inflammatory fibroid polyp of the duodenum. AB - Duodenal inflammatory fibroid polyps (IFP) are extemely rare lesions indistinguishable from submucosal tumors by endoscopic inspection alone. Like gastric inflammatory fibroid polyps, they can be managed by endoscopic polypectomy or mucosectomy. However, preoperative diagnosis of this benign lesion is difficult. Here we present a case of duodenal IFP causing gastrointestinal bleeding that was evaluated by endoscopic ultrasound before surgical removal. On endosonography, the duodenal IFP appeared as a coarsely heterogeneous isoechoic and hypoechoic mass circumscribed by a distinct margin and arising from the third layer of the duodenal wall. The endosonographic appearance of this lesion was in marked contrast to that previously reported for gastric IFPs, which have tended to appear as hypoechoic homogeneous lesions with indistinct margins. Endosonographic evaluation of suspected IFPs before endoscopic or surgical treatment is useful. However, the endosonographic appearances of duodenal and gastric IFPs may be significantly different, possibly because of differences in the makeup of the duodenal and gastric walls. PMID- 10854513 TI - Left thoracoscopic sympathectomy and stellate ganglionectomy for treatment of the long QT syndrome. AB - The long QT syndrome (LQTS) is a rare inherited cardiac disorder that may induce fatal cardiac arrhythmias. Patients diagnosed with this disorder generally have several treatment options, including beta-blockade, cardiac pacing, an implantable automatic defibrillator, or a high thoracic left sympathectomy. We report the case of a 6-year-old girl with the LQTS treated by left thoracoscopic sympathectomy and stellate ganglionectomy. The procedure was performed after an initial thorascopic attempt at another institution failed due to inadequate resection of the sympathetic chain. Operative time was 85 min and blood loss was minimal. There were no intraoperative or postoperative complications. The girl's QT interval decreased and she was discharged on the 4th postoperative day. After 9 months of follow-up, she remains asymptomatic. We conclude that the LQTS patients who fail medical treatment can be treated successfully with left thoracoscopic cervicothoracic sympathectomy. We recommend that the extent of sympathectomy for treating the LQTS be T1-T4 and either the entire stellate ganglion or at least the inferior one-third. PMID- 10854514 TI - A novel diagnosis of left paraduodenal hernia through laparoscopy. AB - A congenital intraperitoneal hernia, also known as a "paraduodenal hernia," is an extremely rare cause of intestinal obstruction. These hernias, which are caused by variations in intestinal rotation, present with symptoms ranging from intermittent abdominal pain to acute obstruction. Preoperative diagnosis is rare, and conventional treatment is usually by laparotomy. Laparoscopic diagnosis and repair has recently been reported in Japan. We present as case of a left paraduodenal hernia diagnosed and treated laparoscopically and a review of the literature. PMID- 10854515 TI - Laparoscopic management of enterocutaneous fistula. AB - Enterocutaneous fistulas develop in settings of prior abdominal surgery, inflammatory bowel disease, diverticulitis, radiation or malignancy. Traditional surgical management requires laparotomy with bowel resection and anastomosis and is associated with a high incidence of wound infection. Recent advances in instrumentation and accumulation of experience has allowed minimally invasive surgery to become an alternative and often preferred approach to handling complex surgical problems. We present a case of successful laparoscopic management of an enterocutaneous fistula that developed in the setting of prior colectomy and laparoscopic inguinal hernia repair with prosthetic mesh. Laparotomy and its attending complications were avoided facilitating recovery and return to work. PMID- 10854516 TI - Laparoscopic splenectomy for the treatment of gastric varices secondary to sinistral portal hypertension. AB - Portal hypertension presents significant challenges to the laparoscopic surgeon. Here we review the case of a successful laparoscopic splenectomy in a patient with sinistral portal hypertension. The value of preoperative splenic artery embolization is highlighted. PMID- 10854517 TI - Polyarteritis nodosa presenting as massive upper gastrointestinal hemorrhage. AB - We report the first case of massive upper gastrointestinal hemorrhage as the initial presentation of polyarteritis nodosa (PAN), which is an uncommon form of systemic necrotizing vasculitis that may involve many organ systems and could affect any age group. Abdominal pain is the most common sign of gastrointestinal involvement. Gastrointestinal bleeding occurs less frequently in approximately 6% of cases. Reported cases of gastrointestinal hemorrhage have been in the form of coffee ground emesis, melena, or hematochezia. Such bleeding complications have resulted from ischemic ulceration or perforation of the small or large intestine. However, we are unaware of previous reports showing massive hematemesis to be the initial presentation of PAN. PMID- 10854518 TI - Laparoscopic hepatic surgery guided by hookwire localization. AB - Due to recent improvements in radiographic technique, computed tomography (CT) occasionally delineates small hepatocellular carcinomas (HCCs) that are invisible with sonography. However, surgery has not been a viable option for these lesions because of the absence of tumor localization. We describe a new technique of preoperative tumor localization using a hookwire to guide laparoscopic surgery for such HCCs. A 68-year-old man with HCC had tumor recurrence after chemoembolization. Two recurrent lesions, 10 mm or less in diameter, located in segment III were demonstrated; not by sonography but by Lipiodol CT. We successfully placed a hookwire into the tumor through a 21-gauge needle under the guidance of CT. The hookwire instrument provided the only clue of tumor location at laparoscopy. The liver around the hookwire was thoroughly coagulated. The postoperative course was uneventful, and the tumor was completely ablated. Preoperative CT-guided hookwire placement is useful to localize and to laparoscopically treat small hepatic lesions. PMID- 10854519 TI - Fatal mediastinitis after routine laparoscopic cholecystectomy. AB - Laparoscopic cholecystectomy is now considered a routine operation with a low complication rate. In this case study, the authors present a laparoscopic cholecystectomy patient who died of masked mediastinitis and concomitant septicemia caused by an unrecognized esophageal perforation after difficult intubation. The authors call attention to the need for early detection of perforating mediastinitis to prevent a lethal outcome from this infrequent but life-threatening condition. PMID- 10854520 TI - Acute presentation of transverse colon injury following percutaneous endoscopic gastrostomy tube placement: case report and review of current management. AB - We describe a case of a patient who had a percutaneous endoscopic gastrostomy (PEG) tube placed for enteral access. The patient's medical history was remarkable for chronic malnutrition, coronary artery disease, coronary bypass surgery, and severe esophageal dysmotility. We discuss the patient&'s course through treatment and we review the management options for patients that sustain colonic injury related to PEG placement. We conclude that colonic injury can be difficult to diagnose in the acute setting and that diagnosis may be facilitated by abdominal computerized tomographic (CT) scanning. PMID- 10854521 TI - Laparoscopic resection of an adrenal neuroblastoma detected by mass screening that grew in size during the observation period. AB - Neuroblastomas (NB) identified by mass screening tests are characterized by benign features. Recently, laparoscopic resection has been applied to the treatment of patients with small adrenal NB (<2-3 cm). However, an increasing number of cases of small NB are followed without any treatment in Japan because many cases regress spontaneously. We describe a case of right adrenal NB detected by mass screening that increased in size during an observation period of 8 months. In this case, laparoscopic resection was performed successfully. The size of the tumor was 27 x 20 x 18 mm at diagnosis and 51 x 42 x 35 mm when it was excised. Small adrenal NB that do not regress during the observation period may require laparoscopic resection before they reach 5 cm in maximum diameter. PMID- 10854522 TI - Overcoming Wallstent malposition in the treatment of rectosigmoid obstruction. AB - In recent years, the use of transanal stenting of malignant colonic strictures for the palliation of obstructive symptoms has increased. Due to the rectosigmoid angle, stenting sigmoid tumors is more troublesome than rectal lesions, but the difficulty may be overcome by using a two-team approach. The radiologist assists the endoscopist with the use of fluoroscopy to ensure proper positioning of both the colonoscope and the stent. The most common complication is stent migration, but stent obstruction and colonic perforation may also occur. We treated a woman suffering from metastatic gastric cancer with peritoneal metastases by creating a 12-cm stricture in the sigmoid colon. Two adjoining Wallstents were required to bridge the obstruction. Following migration of the proximal stent, a third stent was introduced to bridge the previous two stents with satisfactory outcome. PMID- 10854523 TI - Gastric outlet obstruction by a gallstone (Bouveret's syndrome). AB - Gastric outlet obstruction caused by a gallstone in the duodenum or pylorus(Bouveret's syndrome) is a very rare complication of gallstone disease. Presenting symptoms include epigastric pain, nausea, and vomiting. Preoperative diagnosis is not easy. Oral endoscopy is one of the diagnostic procedures. We present a case in which the diagnosis was made by endoscopic examination. Multiple attempts at endoscopic extraction of the gallstone from the duodenum were unsuccessful. A one-stage surgical procedure consisting of the removal of the impacted stone, fistula repair, and cholecystectomy was performed in this case. The postoperative course was uneventful. PMID- 10854524 TI - Laparoscopic resection of a proximal splenic artery aneurysm. AB - The usual treatment for splenic artery aneurysm is resection under laparotomy. In recent years, the laparoscopic approach has consisted of ligation without resection. More recently,laparoscopic resection was reported by the Cleveland Clinic. In this paper, we describe the technique used in the laparoscopic resection of our first case of laparoscopic resection of splenic artery aneurysm (SAA). The patient was a young woman with a 12-mm SAA discovered on systematic abdominal ultrasound. The laparoscopic procedure was done successfully, and the aneurysm was resected using an ultrasonic dissector. The postoperative course was uneventful, and the patient was discharged on the 3rd postoperative day. Pathological examination revealed the atherosclerotic origin of the aneurysm. The patient is doing well 12 months after surgery, with normal splanchnic Doppler ultrasound. This procedure offers a one-step definitive cure via a minimally invasive surgical procedure. PMID- 10854525 TI - Is the loss of gallstones during laparoscopic cholecystectomy an underestimated complication? AB - Laparoscopic cholecystectomy entails an increased risk of gallbladder rupture and consequent loss of stones in the abdominal cavity. Herein we report the case of a 51-year-old male patient, who underwent laparoscopic cholecystectomy 2 years before presentation to our hospital. He had experienced tension sensation and epigastric pain since 4 months postoperatively. A well-defined epigastric mass, which was hard and painful on palpation, was detected and later confirmed by ultrasonography and CT scan. Explorative laparotomy revealed a mass in the area of the gastrocolic ligament,resulting from biliary gallstones in conjunction with a perimetral inflammatory reaction. A review of the literature showed that the incidence of gallbladder lesions during laparoscopy is 13-40%. In order to prevent this complication, meticulous isolation of the gallbladder, proper dissection of the cystic duct and artery, and careful extraction through the umbilical access are required. Ligation after the rupture or use of an endo-bag may be helpful. The loss of gallstones and their retention in the abdominal cavity should be noted in the description of the surgical procedure. PMID- 10854526 TI - Intraoperative Doppler color flow imaging combined with regulation of arterial inflow during surgery for intrahepatic arterioportal fistula. AB - Large fistulas associated with impaired liver function should be treated by direct obliteration or removal of the shunt orifice. In a large shunt with the portal branch lying on the arterial branch, identification of the exact site of the fistula can be a challenge. We report a case of impaired liver function due to a large intrahepatic arterioportal fistula. The site of the shunt orifice could not be located accurately by preoperative imaging. However, intraoperative color Doppler ultrasonography and the simple regulation of arterial inflow clearly demonstrated the shunt orifice. This original technique has allowed the precise definition of the problem and has optimized the surgical treatment for this critical condition. Consequently, it should be considered a new option for the definition and management of large intrahepatic arterioportal fistulas. PMID- 10854527 TI - Successful management of para-aortic lymphocyst with laparoscopic fenestration. AB - Para-aortic lymphocyst occasionally follows retroperitoneal para-aortic node dissection for neoplastic diseases. We present a case in which the leakage of chylous fluid and then a para-aortic lymphocyst followed right nephrectomy and para-aortic node dissection for kidney cancer. Our method of treatment utilized conservative management of chylous ascites and laparoscopic internal drainage of the retroperitoneal lymphocyst. PMID- 10854528 TI - Laparoscopic diagnosis of gallbladder agenesis. AB - In this article, we report two cases of gallbladder agenesis that were incorrectly diagnosed as cholelithiasis on preoperative sonography. In the first case, the diagnosis was made by laparoscopic surgery and confirmed by postoperative CT scan. The second case was confirmed by laparoscopic abdominal examination and by laparoscopic sonography. Both patients had undergone preoperative IV cholangiography. Preoperative cholangiography and laparoscopic exploration completed by laparoscopic sonography should be considered adequate modalities for the diagnosis of gallbladder agenesis, without the need for laparotomy and thorough postoperative workup. PMID- 10854529 TI - Radiation-induced apoptosis: predictive and therapeutic significance in radiotherapy of prostate cancer (review). AB - Current therapy for advanced prostate cancer is hampered by the propensity of the disease to progress from an androgen-dependent state to an androgen-independent state. Current treatment for advanced disease is palliative. Therefore, the therapeutic goal for prostate cancer treatment today is to arrest the disease at an early state when it is still localized to the gland. The standard treatment for clinically localized disease is radical prostatectomy or radiation therapy by way of external beam irradiation or local radioactive seed implants (brachytherapy). In advanced disease, the use of radiation therapy is limited to palliation of pain secondary to bone metastases and for spinal cord compression. Tracking residual disease and predicting outcome is limited to following the level of prostate specific antigen (PSA) production, evaluating for bone or solid organ metastasis, and analyzing their preoperative clinical stage, PSA and Gleason's score. Apoptosis as a molecular process of genetically regulated cell death has a critical endpoint that coincides with the goal of successful treatment of human malignancies. Since in cancer treatment the therapeutic goal is to trigger tumor-selective cell death, activation of the apoptotic pathway in prostatic tumor cells offers attractive and potentially effective therapeutic targets. As our understanding of the vital role of apoptosis in the development and growth of the prostate gland has expanded, numerous genes that encode apoptotic regulators have been identified that are severely impaired in prostate tumors. Human prostate cancer cells undergo apoptosis in response to androgen ablation, chemotherapeutic agents and ionizing irradiation. The expression of apoptotic modulators within individual prostate tumors appears to correlate with the cancer cell's sensitivity to traditional therapeutic modalities, including radiotherapy. No strict correlation between radiation-induced apoptosis and longevity of prostate cancer patients has emerged, possibly because the ability to achieve an initial remission alone does not adequately predict long-term outcome and patient survival. In this review we summarize the current understanding of the effects of radiation therapy on prostatic tumor cells within the context of the therapeutic significance of radiation-induced apoptosis in the effective elimination of androgen independent prostate cancer cells. As we enter a new millenium, identification of distinct molecular markers predictive of therapeutic response of prostatic tumors to radiation therapy may afford alternative prognostic indicators in optimizing our treatment protocols for advanced disease. PMID- 10854530 TI - HPV-16 E6 oncoprotein induces mutations via p53-dependent and -independent pathways. AB - The E6 oncoprotein of human papillomaviruses (HPV) promotes oncogenesis by inactivating tumor suppressor protein p53 i. e. it binds to and enhances the degradation of p53. To study whether inactivation of p53 is solely responsible for E6-induced oncogenesis, we constructed several plasmid vectors expressing wild-type (wt) or mutant (mt) E6 proteins. RKO cells that express wt p53 were stably transfected with these plasmids and challenged with DNA damaging agents. The level of p53 was significantly increased by DNA damaging agents in control cells and cells transfected with plasmids expressing mt E6 that do not bind to p53. As expected, p53 did not increase in cells transfected with plasmids expressing mt E6 that do bind to p53. To investigate the oncogenic effect of these various E6 proteins, we determined the mutation frequency of the hprt locus in control cells and cells expressing different E6 proteins. We found that cells expressing wt E6 and mt E6 (capable or incapable of binding to p53) showed notable increases in the mutation frequency at hprt locus compared with that of control cells. The elevation of mutation frequency in cells expressing mt E6 was similar to that in cells expressing wt E6. These data indicate that E6-induced mutagenicity is induced not only via p53 inactivation, but also via p53 independent pathways. PMID- 10854531 TI - Predictive value of cathepsin D and Ki-67 expression at the deepest penetration site for lymph node metastases in gastric cancer. AB - To search for reliable predictors for lymph node metastasis, we immunohistochemically analyzed surgically resected gastric cancer specimens that showed invasion of submucosa (sm) and muscularis propria (mp) of the tumor. The analysis investigated cathepsin D and Ki-67 expression in 136 specimens that were divided into an sm1/sm2 group and an sm3/mp group. In sm1/sm2 group, the incidence of lymph node metastases was significantly higher in tumors with high Ki-67 labeling index (LI) (44%) than in those with low Ki-67 LI (0%). In sm3/mp group, the incidence of lymph node metastases was significantly higher in cathepsin D-positive (56%) and high Ki-67 LI tumors (64%) than in cathepsin D negative (33%) and low Ki-67 LI (33%). Combined analysis of cathesin D expression and Ki-67 LI correlated strongly with lymph node metastases. No lesions with cathepsin D-negative expression and low Ki-67 LI had lymph node metastases in either group. Cathepsin D and Ki-67 expression may be useful predictors for lymph node metastases in gastric cancer with sm and mp invasion. As predictors, they can identify lesions without lymph node metastases and indicate lesions not needing additional treatment after endoscopic mucosal resection and laparoscopic gastrectomy. PMID- 10854532 TI - Percutaneous injection of a low-concentration alkaline solution targeting hepatocellular carcinoma. AB - We developed a percutaneous low-concentration alkali injection therapy (PLAIT) targeting hepatocellular carcinoma (HCC), and compared the necrotic areas in the livers of rats that had received PLAIT [an alkaline solution of sodium hydroxide (NaOH)] with those that had received percutaneous ethanol injection therapy (PEIT) and percutaneous acetic acid injection therapy (PAIT). The necrotic area increased with increasing concentrations of NaOH solution. The survival rate of rats was 100% up to a concentration of 4 N; however, the rate dropped below 80% with concentrations over 5 N. Also, at a concentration of 2 N, the necrotic area increased with increasing quantities from 0.01 ml to 0.1 ml. PLAIT using 0.05 ml of 2 N NaOH was 1.56 times more effective than PEIT using 0.05 ml of 99.5% ethanol, and 63.33% less effective than PAIT using 0.05 ml of 50% acetic acid. However, the survival rate after PAIT was 50%, while that after PLAIT was 100%. Histopathological observation of normal rat livers after injection of 2 N NaOH at a volume of 0.05 ml showed that the tissue necrosis spread radially from the site immediately after injection by PLAIT, but necroses were not found in other organs. We conclude that PLAIT has promise as a new form of local therapy for HCC. PMID- 10854533 TI - Expression and localization of vascular endothelial growth factor receptors in human hepatocellular carcinoma and non-HCC tissues. AB - Flt-1 (VEGF receptor-1) and KDR/Flk-1 (VEGF receptor-2) are the high-affinity receptors for the angiogenesis factor, vascular endothelial growth factor (VEGF). VEGF expression has been confirmed in human hepatocellular carcinoma (HCC), and VEGF is thought to be involved in the angiogenesis within HCC tissues. However, expressions of VEGF receptors in HCC have not been reported. We immunohistochemically examined expressions and localizations of Flt-1 and KDR in 28 surgically resected HCC tissues. In non-cancerous area, Flt-1 and KDR were mainly found in macrophages including Kupffer cells; both receptors were found in vascular endothelial cells in the portal veins and arteries within portal tracts; and KDR was also found in some sinusoidal endothelial cells. In cancerous area, Flt-1 and KDR were found in some macrophages, and also in the endothelial cells of intratumoral blood vessels. In 25 moderately and/or poorly differentiated HCCs, KDR expression in the blood space endothelial cells was clear and continuous in 20 cases, and focal in 5 cases. These results suggest that there would be an angiogenesis mechanism via VEGF/Flt-1 or VEGF/KDR in HCC, and the VEGF/KDR system would take a more important role. PMID- 10854534 TI - Correlation between metastatic potency and the down-regulation of E-cadherin in the mouse hepatoma cell lines G-1 and G-5. AB - Cell-cell adhesiveness, involving the adherens junction system including homophilic adhesion of cadherin and intracellular catenins, is a critical factor for tumor cell invasion and metastasis. We evaluated the levels of E-cadherin and beta-catenin in hepatoma cell sublines with high and low metastatic capacities. Stimulation of these cells with serum growth factors for more than 3 h after 24 h of starvation caused decreases in levels of E-cadherin and beta-catenin in the subline with high metastatic capacity, G-5. In contrast, no significant changes were observed in the subline with low metastatic capacity, G-1. Concomitantly with the decreases in E-cadherin and beta-catenin levels, G-5 cells were dissociated and detached from the culture dish, although G-1 cells again showed no morphological alterations. These in vitro results reflected the in vivo metastatic potencies of these hepatoma sublines, and further suggested the importance of the adherens junction system in determining metastatic potency of these parenchymal tumor cell lines as in epithelial/endothelial tumors. PMID- 10854535 TI - In vivo enhancement of chemosensitivity of human salivary gland cancer cells by overexpression of TGF-beta stimulated clone-22. AB - We have isolated transforming growth factor-beta-stimulated clone-22 (TSC-22) cDNA as an anti-cancer drug-inducible gene in a human salivary gland cancer cell line, TYS. We have previously reported that TSC-22 negatively regulates the growth of TYS cells, and that overexpression of TSC-22 protein in TYS cells enhanced the in vitro chemosensitivity of the cells. In this study, we examined the in vivo chemosensitivity of TSC-22-expressing TYS cells. TSC-22-expressing TYS cells formed tumors in nude mice, but tumors formed by TSC-22-expressing TYS cells were significantly smaller than tumors formed by control cells (p<0.001, one way ANOVA). Furthermore, intraperitoneal injection of 5-fluorouracil (5-FU) markedly inhibited the growth of the TSC-22-expressing TYS tumors, but did not affect the growth of control tumors. It was found by TUNEL assay that TSC-22 expressing TYS tumors were induced to undergo apoptosis by 5-FU treatment. These findings suggest that overexpression of TSC-22 protein in TYS cells enhances the in vivo chemosensitivity of the cells to 5-FU via induction of apoptosis. PMID- 10854536 TI - p53 overexpression and K-ras codon 12 mutations in submucosal invasive depressed type colorectal cancer. AB - We examined histological findings, p53 overexpressions and K-ras codon 12 mutations and the histology of submucosal invasive (sm) colorectal cancers. Sixty specimens of sporadic sm cancer were obtained by surgical resection or endoscopic polypectomy. p53 expression was examined by immunohistochemical staining using the streptavidin-biotin method. K-ras codon 12 mutations were detected by polymerase chain reaction (PCR) and dot blot hybridization. These tumors were classified as depressed-type or polypoid-type sm cancers. Among 60 sm cancer samples, depressed-type sm cancers were found in 9 (15%) and polypoid-type sm cancers were found in 51 (85%). The frequency of p53 expression was significantly higher in depressed-type [7/9 (78%)] as compared to polypoid-type [18/51 (35%)] sm cancers (p<0.05). The frequency of K-ras codon 12 point mutations was significantly lower in depressed-type [0/9 (0%)] as compared to polypoid-type [23/51 (45%)] sm cancers (p<0.05). We conclude that the development of depressed type cancers may involve a distinct genetic pathway. PMID- 10854537 TI - Detection of micrometastases in sentinel lymph nodes of the breast applying monoclonal antibodies AE1/AE3 to pancytokeratins. AB - Sentinel lymph node excision in breast cancer is a minimally invasive diagnostic procedure for accurate staging of the axilla and for avoiding unnecessary axillary dissection. In patients with palpable breast cancer we injected microcolloidal particles of human serum albumin labelled with technetium-99m the day before surgery. The sentinel node was detected intraoperatively with a handheld gammaprobe and then removed. Complete axillary dissection was performed and the nodes inspected by routine histological examination. The axillary lymph node status was correctly predicted by the sentinel node technique in 32 of 33 breast cancer patients. Two cases of micrometastases escaped routine histopathological detection but were identified by immunohistochemical analysis applying the antibody AE1/AE3 to pancytokeratins. Immunohistochemical examination of the sentinel node improves the diagnostic security of patients with breast carcinoma by detection of micrometastases. PMID- 10854538 TI - Antitumor properties of an anti-idiotypic monoclonal antibody in relation to N glycolyl-containing gangliosides. AB - We examined the antitumor effects of 1E10 monoclonal antibody, an anti-idiotypic IgG to an IgM monoclonal antibody, named P3, that reacts specifically with N glycolyl-containing gangliosides and also recognizes antigens in human breast and melanoma tumors. Two murine tumor cell lines positive for the P3 antibody, F3II mammary carcinoma (BALB/c) and B16 melanoma (C57BL/6), were employed. In BALB/c mice, vaccination with several i.p. doses at 14-day intervals of 50 microgram of 1E10 coupled to keyhole limpet hemocyanin in Freund's adjuvant, significantly reduced s.c. tumor growth of F3II carcinoma cells and the number of spontaneous lung metastases. Also, the effect of 1E10 as a biological response modifier on tumor lung colonization was evaluated in C57BL/6 mice injected i.v. with B16 melanoma cells. Interestingly, i.v. administration of 10 microgram of uncoupled 1E10 antibody, 10-14 days after inoculation of B16 cells, dramatically reduced the number of experimental metastases in comparison with lungs from mice treated with an irrelevant IgG. The present data suggest that this 'non-internal image' anti-idiotypic monoclonal antibody may activate more than one mechanism of antitumor response against melanoma and mammary tumor cells. PMID- 10854539 TI - Expression of growth hormone receptor in human liposarcomas and lipomas. AB - Our immunohistochemical results clearly demonstrated the occurrence of growth hormone receptors (GH-R) in the tumour cells of lipomas and liposarcomas. In liposarcomas staining intensity in the cytoplasm of tumour cells varied between weak and distinct but could not be correlated to the histological grade of the malignant tumours. These findings were corroborated to some extent by the RT-PCR results. RT-PCR analysis of human lipomas and liposarcomas revealed the amplified cDNA fragment of GH-R in 8 out of 12 lipomas but only in 3 out of 10 liposarcomas. The reduced number of GH-R positive tumours found with PCR may be explained by the extraction method of RNA from paraffin sections. An interesting finding was the distinct immunoreactivity of the endothelium of blood vessels in liposarcomas, which was especially pronounced in the newly forming capillaries. This points to an important role of GH-R in tumour angiogenesis which could significantly contribute to tumour growth in liposarcomas and may open the possibility for therapeutic intervention using antiangiogenic substances. PMID- 10854540 TI - Ocular metastases from breast carcinoma: A multicentric retrospective study. AB - Breast cancer may affect the eye and orbit by metastatic neoplastic infiltration, uvea being the most common site of presentation. Management of these cases with radiotherapy is usually gratifying with reported response rate of approximately 75%. A retrospective evaluation of cases treated in five Institutions participating in a collaborative radiation therapy group of north-Italy is reported. Fifty-four cases of metastases to the eye or orbit were referred for radiation therapy to the Departments participating in the survey in the period 1977-1995. There were 49 female patients aged between 28 and 75 years (median, 44 years) at presentation of orbital metastasis. Thirty-eight lesions (70%) were metastases to the choroid, 9 involved other parts of the eye, and 7 patients had orbital metastases. Five of the 49 patients had bilateral choroidal metastases. Radiotherapy was employed with megavoltage equipment. The median total dose delivered was 40 Gy (range, 16-60 Gy). All the patients were treated 5 times per week with fraction sizes ranging from 1.8 to 3.0 Gy (median, 2.0 Gy). Of the 43 evaluable eyes, 34 (79%) showed a definite improvement after radiotherapy. There was a stabilization of the process in 4 patients. The rest (11 lesions) were lost to detailed follow-up of the response of the eye metastases. Twelve patients experienced acute transient cheratoconjunctivitis and in a case a subconjunctival haemorrhage was observed; as late side effects, two cases of chataract were observed during a period of observation of 37 and 117 months. A median survival time of 17 months was observed. The goal of irradiation was to improve vision or at least prevent blindness and enucleation. The palliative effect of irradiation was confirmed with a response rate consistent with the data of the literature on this subject. PMID- 10854541 TI - The utility of tissue harmonic imaging in the liver: a comparison with conventional gray-scale sonography. AB - The purpose of this study was to determine if tissue harmonic imaging (THI) sonography produces higher quality images of the liver than conventional sonography. A prospective study was performed on 91 anatomic areas (60 liver tumors, 31 vascular structures) in 44 patients to compare the image quality of THI sonography with that of conventional sonography. THI sonography transmits at 2.0 MHz and receives at 4.0 MHz, while conventional sonography is used at 2.5 MHz or 4.0 MHz. All anatomic areas were studied by THI and by conventional ultrasonography at 2.5 MHz and at 4.0 MHz. Observers who were blinded to the sonographic technique ranked the quality of the obtained images. THI was the best technique for the evaluation of liver tumors, at an intermediate depth (p<0.0001) (4-8 cm from the body surface), but not for assessing superficial (4 cm or less) or deep (over 8 cm) tumors. THI was ranked as the best technique for examination of vascular structures regardless of depth. There was statistical difference at an intermediate depth (p<0.0001), but no significance was found for deep vascular structures (p=0.061). The number of superficial vascular structures was too small for statistical analysis. THI sonography is more useful than conventional ultrasonography for the study of vascular structures and intermediate-depth liver tumors, but is not always adequate for superficial or deep anatomic areas. PMID- 10854542 TI - Concurrent chemoradiotherapy combined with intraoperative radiotherapy for locally advanced pancreatic cancer: a feasibility study. AB - Initial clinical results of concurrent chemoradiotherapy combined with high-dose intraoperative radiotherapy (IOR) for locally advanced pancreatic cancer were analyzed. Between June 1996 and May 1999, 6 patients with locally advanced pancreatic cancer without distant metastasis were treated with preoperative concurrent chemoradiotherapy followed by IOR. Preoperative radiation therapy was given by the dynamic arc conformal technique with a daily fraction of 1.8 Gy to a total dose of 45 Gy in 5 weeks. Cisplatin (5 mg/day for 4 weeks) and 5 fluorouracil (250 mg/day for 5 weeks) were administered continuously during preoperative radiation therapy. IOR as a single dose of 28 or 30 Gy was given to the gross tumor volume using electron beams of 15- to 22-MeV. Concurrent chemoradiotherapy was well tolerated, although all of the patients complained of nausea and fatigue. Two patients developed grade III leukopenia. No other serious acute toxicity was noted. The median survival time of the 6 patients was 17.5 months, which was significantly longer than that of our historical control treated with external radiation therapy with IOR (8 months), although the difference in survival was borderline significant (p=0.068). Concurrent chemoradiotherapy followed by high-dose IOR was well tolerated in patients with locally advanced pancreatic cancer, and the initial clinical results appeared promising. PMID- 10854543 TI - Evaluation by dual energy X-ray absorptiometry of changed bone density in metastatic bone sites as a consequence of systemic treatment. AB - Fourteen cancer patients with bone metastases from various primary malignancies were submitted to repeated dual X-ray absorptiometry (DEXA) scan before and after systemic antineoplastic treatments. In the nine patients with lytic lesions the Bone Mineral Density (BMD) increased after chemotherapy + pamidronate in four (by +11.2%, +7.5%, +5.0% and +6.6%, respectively), decreased in four (by -19.9%, 8.1%, -7.5%, and -7.0%, respectively) and remained unchanged in one. BMD changes paralleled variations in painful symptomatology and biochemical markers. In patients with blastic metastases the BMD on target metastatic lesions did not change after hormone therapy or chemotherapy in one case but showed a significant increase in four. BMD increase was associated to bone pain improvement and PSA decrease in two cases, and with a worsening in skeletal pain and/or serum PSA in the remaining two. Our data suggest that BMD evaluation by DEXA instrument may be a reliable tool in assessing the response of bone metastases to treatment. PMID- 10854544 TI - Conditions of curability after endoscopic resection for colorectal carcinoma with submucosally massive invasion. AB - The deepest invasive portion of colorectal carcinoma (CRC) is considered to be the part, which ultimately will invade, spread locally and give metastasis. We have previously reported that histologic differentiation at the deepest invasive portion of CRC closely correlate with metastatic potential and is useful in understanding the curability of endoscopic mucosal resection (EMR). The aim of this study is to clarify the conditions of curative EMR for CRC with submucosally (sm) massive invasion. A total of 521 cases with sm invasive CRC (Group A, 470 surgically resected cases; Group B, 51 followed-up cases after EMR) were studied. The depth of sm invasion was defined as the practically measured distance from muscularis mucosae. Histologic subclassification was performed at the deepest invasive tumor margin as: well-differentiated (W), moderately differentiated (M) and poorly differentiated (Por). By assessing glandular configuration and cellular arrangement, M type was further subdivided into two different groups; moderately-well differentiated (Mw) and moderately-poorly differentiated (Mp). In group A, lymph node (LN) metastasis was detected in 45 (9.6%) of 470 cases. W or Mw lesions showed LN metastasis in 4.9% (19/388). Mp or Por lesions showed LN metastasis in 37.3% (25/67) (W/Mw vs Mp/Por; p<0.01). Of 45 cases with LN metastasis that could be measured the practical distance of sm invasion, W or Mw lesions showed no LN metastasis in cases within 1,500 micrometer invasion. However, Mp or Por lesions showed LN metastasis in cases within 1,500 micrometer invasion (5/15, 33.3%, minimum 400 micrometer invasion; so-called scanty invasion). In group B, none of 51 cases died of LN metastasis and showed no other metastasis, although 17 cases (33.3%) showed an sm invasion more than 1,500 micrometer. These results indicated that CRC even with sm massive invasion can be cured by complete EMR on conditions that the depth of sm invasion is within 1,500 micrometer and histologic grade at the deepest invasive portion is W or Mw, if there are no vessel involvement. However, cases with Mp or Por grade were not curative by EMR, even if they showed an sm scanty invasion. PMID- 10854545 TI - Mutational analysis of transforming growth factor beta receptor type II and DNA mismatch repair genes in sporadic endometrial carcinomas with microsatellite instability. AB - To clarify how microsatellite instability (MI) is involved in carcinogenesis of sporadic endometrial carcinoma, we examined mutations of the transforming growth factor beta receptor type II (TGF beta RII) gene in 32 patients with MI-positive sporadic endometrial carcinoma. Moreover, mutations of 4 DNA mismatch repair (MMR) genes (hPMS1, hPMS2, hMLH1, hMSH2), which are considered to cause MI, were investigated as well. With respect to the TGF beta RII gene, mutations in the 10 bp polyadenine repeat sequence were observed in 7 of 29 informative cases (24%). Concerning MMR genes, a T to C point mutation at the -6 intronic splice acceptor site of exon 13 of hMSH2 was detected in 43% (6/14). However, there was no mutation in any exon of these 4 MMR genes. These results suggest that there is a carcinogenic mechanism via mutation of the TGF beta RII gene in some cases of MI positive sporadic endometrial carcinoma. It seems unlikely that the unknown MMR genes are responsible for MI. The implication of the mutation at the intronic splice acceptor site in hMSH2 remains to be clarified. PMID- 10854546 TI - Protein kinase C activity in human gastric carcinoma. AB - Protein kinase C (PKC) is believed to play an important role in tumorigenesis, so we measured PKC activities of human gastric carcinomas and adjacent normal mucosae to elucidate its role for gastric carcinogenesis using PegTagtrade mark non-radioactive protein kinase C assay. The mean activities of microsomal PKC in carcinoma and normal mucosa were 449+/-179, 661+/-264 pmol/min/mg, respectively (p=0.001). Cancerous microsomal PKC activity decreased when the tumor was small, node-metastasis was negative, the depth of invasion was shallow and the histological type was differentiated. Our results suggested that PKC activity would be down-regulated in membrane of the early stage of gastric carcinoma. PMID- 10854547 TI - Neoadjuvant intra-arterial infusion chemotherapy combined with hormonal therapy for locally advanced breast cancer. AB - To determine the efficacy of combined neoadjuvant intra-arterial infusion chemotherapy and hormonal therapy for treating locally advanced breast cancer, we compared the outcomes of patients with or without this therapy, and also assessed histologic response. Ninety-four patients with locally advanced breast cancer (stage IIIa, 56; stage IIIb, 38). Nineteen stage IIIa and 17 stage IIIb patients received intra-arterial plus hormonal therapy while 37 stage IIIa and 21 stage IIIb patients with similar ages and follow-up durations did not. Treated patients received intra-arterial epirubicin plus oral medroxy-progesterone. Five-year disease-free survival rates were 77.5% for intra-arterially treated and 33.0% for other patients in stage IIIa, and 70.5% for intra-arterially treated and 38.1% for other patients in stage IIIb. Five-year overall survival rates were 94.4% for intra-arterially treated and 61.7% for other patients in stage IIIa, and 90.9% for intra-arterially treated and 56.3% for other patients in stage IIIb. Ten-year overall survival rates in stage IIIb were 90.9% for treated and 22.5% for other group patents. All differences were statistically significant (p<0.05). Good histologic response to intra-arterial therapy was seen in 75% of the primary tumors and 71% of involved lymph nodes. Neoadjuvant intra-arterial therapy with hormonal therapy yielded better survival rates than no intra-arterial therapy or our previous intra-arterial regimen. PMID- 10854548 TI - Estrogen-producing ovarian mucinous cystadenomas in postmenopausal women. AB - Cells from benign and borderline malignant mucinous cystadenomas surgically resected from three postmenopausal women with elevated serum estradiol levels were found to have different abilities to synthesize estrogen from [14C] cholesterol. Addition of gonadotropins to cultured cells with low levels of estrogen synthesis did not result in any significant increase in estrogen synthesis. Immunohistochemical studies of these mucinous cystadenomas showed that LDL receptors were distributed in the benign cystadenoma cells and in the stromal cells of borderline tumors, but not in the cystadenoma cells of borderline tumors. Sterol regulatory element binding protein was immunohistochemically shown to be distributed in the mucinous cystadenoma cells of borderline tumors, but not in the benign cystadenoma cells. PMID- 10854549 TI - Subtotal peritonectomy with chemohyperthermic peritoneal perfusion for peritonitis carcinomatosa in gastrointestinal cancer. AB - Treatment for peritonitis carcinomatosa in gastrointestinal cancer remains to be established though it is one of the commonest causes of cancer death. Subtotal peritonectomy (SP) with chemohyperthermic peritoneal perfusion (CHPP) was developed for the new therapeutic strategy for peritoneal dissemination in gastrointestinal cancer in our department. SP includes resection of stomach, colon, small bowel, spleen, gall bladder, and parietal peritoneum. CHPP was carried out by heated saline containing 25 mg/l cisplatin, 10 mg/l mitomycin C, and 20 mg/l etoposide. Intraperitoneal temperature was maintained at 42 degrees C for 60 min. Fifteen gastric cancer and three colon cancer patients with severe peritoneal dissemination underwent these procedures. The averages of operating time, intraoperative bleeding volume, and total perioperative transfused blood volume were 9 h, 4400 ml, and 5600 ml, respectively. The patients estimated as complete resection and residual disease by histopathological study numbered 11 and 7. There was no treatment-related deaths though bleeding occurred in 5 patients; perforation in 2 patients; and abscesses in 2 patients. The 1-year survival rate (1ysr) and the 2-year survival rate (2-ysr) of all the patients were 57% and 21%, respectively. The 1-ysr and the 2-ysr of the patients who underwent complete resection were 67% and 40% significantly greater than the 43% and 0% of the patients who had residual tumors (p=0.02). The combination therapy of SP and CHPP is feasible in spite of its morbidity and has great possibilities in complete resection of peritoneal dissemination and prolongation of patient's survival. PMID- 10854550 TI - A case-control study evaluating occult blood screening for colorectal cancer with hemoccult test and an immunochemical hemagglutination test. AB - A case control study to evaluate the occult blood screening for colorectal cancer was conducted in a town where colorectal cancer screening had been performed by Hemoccult test during the early years and subsequently by an immunochemical hemagglutination test. All residents aged >/=40 years had been offered the annual screening. Case series consisted of 51 subjects with fatal colorectal cancer. Three controls per case were selected from the list of residents who were alive at the time of diagnosis of the corresponding case and had been living in the town, matched by gender and by age. The odds ratio (OR) of dying of colorectal cancer for those having their most recent screening histories with Hemoccult test or the immunochemical test during the preceding 1 year and 1-2 year segment before case diagnosis were 0.20 [95% confidence interval (CI): 0.08-0.49] and 0. 17 (95% CI: 0.04-0.75), respectively. The OR increased towards 1.0 as the number of years since the most recent screening increased. The OR of dying of colorectal cancer was calculated to be 0.19 (95% CI: 0.05-0.70) for those screened with the immunochemical test alone during the preceding 1 year after adjustment for previous screening histories with the Hemoccult test. Corresponding OR was 0.36 (95% CI: 0.11-1.17) for those screened with Hemoccult test during the preceding 1 year. These results suggest that screening for colorectal cancer by fecal occult blood testings or immunochemical test alone would reduce mortality and that efficacy of the screening would be higher for the immunochemical test than for Hemoccult test. PMID- 10854551 TI - Raf-1 kinase activity predicts for paclitaxel resistance in TP53mut, but not TP53wt human ovarian cancer cells. AB - We have recently reported that there is a significant Raf-1 kinase dependency of paclitaxel resistance in human cervical tumor cell lines. In light of the possibility that Raf-1 kinase inhibitors could be used to enhance paclitaxel responsiveness in ovarian cancer, we have characterized the Raf-1 kinase dependency of paclitaxel resistance in ovarian cancer cells. The relationship between Raf-1 kinase activity and the sensitivity to clinically relevant paclitaxel concentrations was determined in four ovarian cancer cell lines (CA OV3, SK-OV3, 2780/WT and OAW42/WT). Furthermore, in recognition that such a drug combination would initially be used in patients whose tumors have recurred following cisplatin/paclitaxel treatment, we also determined the Raf-1 kinase dependency of paclitaxel cytotoxicity in cisplatin resistant variants of two of the ovarian cell lines (2780/CP and OAW42/CP). In the two cell lines (2780/WT and OAW42/WT) that possess a wild-type TP53 (TP53wt), the relationship between Raf-1 kinase activity and paclitaxel resistance was different from that observed in the cervical tumor cell lines. In these cell lines, paclitaxel-induced far more cell killing than would have been predicted from their Raf-1 kinase activity. However, in the ovarian cancer cell lines (CA-OV3, SK-OV3, 2780/CP and OAW42/CP) that have a mutant TP53 (TP53mut), the cytotoxicity induced by 60 nM paclitaxel exhibited the same relationship to Raf-1 kinase activity as previously observed in cervical tumor cell lines. These data suggest that the therapeutic efficacy of paclitaxel in ovarian cancer patient whose tumors have TP53mut might be increased if it is administered in combination with Raf-1 kinase inhibitors, e.g., ISIS 5132. PMID- 10854552 TI - Tumor marker and MR imaging criteria for evaluating the efficacy of cyclic balloon-occluded arterial infusion for advanced cancer of the uterine cervix. AB - We have already reported satisfactory therapeutic results of cisplatin-based cyclic balloon-occluded arterial infusion chemotherapy (BOAI) enabling treatment by simple total hysterectomy (STH) in patients with advanced cervical cancer of the uterus (cervical cancer), with 2- and 5-year survival rates of 88.9% and 66. 7%, respectively. Considering the fact that 3 to 4 months are needed to complete BOAI, the present study was a retrospective assessment of 18 cervical cancer patients to prepare criteria for the early determination of the efficacy of BOAI in patients who can be treated by STH. The results showed that STH could be performed after the first BOAI in patients with SCC value decreased to 34.3+/ 8.2% of the pretreatment value (p=0.0001) and whose tumor size had decreased by 45.1+/-7.5% (p=0.0002). PMID- 10854553 TI - Expressions of thymidine phosphorylase (dThdPase) and vascular endothelial growth factor on angiogenesis in intestinal-type gastric carcinoma. AB - We examined the clinical and pathological significance of thymidine phosphorylase (dThdPase) and vascular endothelial growth factor (VEGF) in human gastric carcinomas, in terms of intratumoral microvessel density (IMVD), P53 expression, and patient prognosis in a total of 128 patients. Mean IMVD was significantly higher in the carcinomas with dThdPase or VEGF expression than in carcinomas without the expression. The simultaneous expression of dThdPase and VEGF was correlated with increased IMVD of human gastric carcinomas. VEGF expression was associated with P53 expression and poor patient prognosis, but dThdPase expression was not. PMID- 10854554 TI - Effects of biopsy on lung metastasis. AB - In order to clarify whether biopsy promotes lung metastasis, open or needle aspiration biopsy was performed 10 or 21 days after S-SLM osteosarcoma cells were transplanted subcutaneously in Fischer rats. The lungs were excised after six weeks and the lung weight and the number of metastatic nodules were measured. The mean weight was more in open than needle biopsy, and the number of nodules was significantly higher in open biopsy after 10 days, compared to the control. From these results we concluded that open is more likely to promote lung metastases compared to needle biopsy under the specific experimental conditions of this study. PMID- 10854555 TI - Immunohistochemical expression of vascular endothelial growth factor can predict response to 5-fluorouracil and cisplatin in patients with gastric adenocarcinoma. AB - The combination of 5-fluorouracil (5-FU) and cisplatin is used most commonly for gastric carcinoma. Recent studies have indicated that vascular endothelial growth factor (VEGF) is related to drug delivery through angiogenesis and vascular permeability. In this study, we evaluated the efficacy and toxicity of continuous infusion of 5-FU and low dose cisplatin infusion as first-line treatment in patients with unresectable gastric adenocarcinoma. We also examined the relationship between chemotherapy response and immunohistochemical expression of VEGF in the biopsy samples of gastric primary. All 30 patients enrolled in this study were assessable for response, adverse reactions, and VEGF expression. The regimen consisted of 5-FU (350 mg/m2/day every day by continuous venous infusion) and low dose cisplatin (7 mg/m2/day by drip infusion over 1 h on days 1-5 every week). This treatment was repeated weekly for 3 consecutive weeks. Four weeks after the second cycle, mesurable lesions were estimated for response. An overall response rate was 46.7% (14/30). Patients with intestinal histologic type (10/12) and good performance status ([PS], 13/18) showed good response rate (83.3%, and 72.2%, respectively) compared to patients with diffuse histologic type (4/18) and poor PS [(1/12) 22.2%, and 8. 3%, respectively]. The response rate of VEGF positive cases and VEGF-negative cases was 75% (12/16), and 16.7% (2/14), respectively. Multivarite analysis revealed that VEGF-positive and good PS had a significant impact on chemotherapy response in this treatment. The most common garde 3 or higher toxicities were myelosuppression (30%) and diarrhea (13.3%). Continuous infusion of 5-FU and low dose cisplatin infusion is an effective treatment for patients with unresectable gastric carcinoma, and VEGF expression may be a useful predictor of chemotherapy response in this regimen. PMID- 10854556 TI - Inhibitory effect of resveratrol on the proliferation of human and rat hepatic derived cell lines. AB - Resveratrol is a polyphenolic compound especially produced by grapevine and consequently found in wine. Based on epidemiological studies resveratrol may act as a cancer chemopreventive compound. The ability of resveratrol to inhibit cell proliferation was studied in rat hepatoma Fao cell line and human hepatoblastoma HepG2 cell line. The results show that resveratrol strongly inhibits cell proliferation at the micromolar range in a time- and dose-dependent manner. Concentrations higher than 50 microM become toxic. Fao cells are more sensitive than HepG2 cells. Interestingly, the presence of ethanol lowers the threshold of resveratrol effect. Resveratrol appears to prevent or to delay the entry to mitosis since no inhibition of [3H]thymidine incorporation is observed, while there is an increase of cell number in S and G2/M phases. In conclusion, resveratrol shows a strong inhibition of hepatic cell proliferation where alcohol may act as an enhancing agent. PMID- 10854557 TI - Evaluation of argyrophilic nucleolar organizer regions (AgNORs) in differentiated thyroid carcinoma as an indicator for disease recurrence. AB - Differentiated thyroid carcinoma (DTC) is one of the least aggressive cancers of the human malignancies, however, recurrent disease is occasionally found and is difficult to determine the risk for recurrence only by clinicopathological features. In the present study, we investigated argyrophilic nucleolar organizer regions (AgNORs) score, a well-known indicator for proliferative potential of the cancer cells, in 89 cases of DTC. In tumors with capsular invasion, or with extended contra-lateral lymph node metastasis, AgNORs score was significantly higher than in tumors without them, while was not correlated with age or gender of the patients, nor the histological type of the tumor (papillary or follicular). Disease recurrence was observed in one-third of the patients in high AgNOR score group (scored more than mean value +/- standard deviation) when the patients were divided into four groups according to the AgNOR score, and a significantly higher risk for disease recurrence was demonstrated in those cases with high AgNOR score than cases with lower score. These results clearly indicated the usefulness of AgNOR score in selecting the patients at high risk for disease recurrence. All 5 recurrent cases displayed local recurrence in high AgNOR score group, while 5 of 7 cases with low AgNOR score did in the manner of hematological metastasis. In conclusion, this method was technically simple and accurate giving an exact value in individual cases. Thus, we believe AgNOR score might be clinically applicable as a useful indicator for disease recurrence in DTC. PMID- 10854558 TI - Establishment of new multidrug-resistant human osteosarcoma cell lines. AB - Multidrug-resistant clones of human osteosarcoma MNNG/HOS and MG63 cells were isolated by stepwise selection on exposure to increasing doses of doxorubicin (DXR). The final clones MNNG/HOS/DXR1000 and MG63/DXR1000, established after ethylmethane sulfonate mutagenesis, showed 96-fold and 121-fold higer resistance to DXR than their parental cell lines. They were also cross-resistant to vincristine, but not to cisplatinum or methotrexate. The levels of multidrug resistance-1 (MDR1) mRNA expression increased gradually according to the concentration of DXR in both cell lines. Although the parental MNNG/HOS cells expressed a low level of MDR1 mRNA, the parental MG63 cells showed no MDR1 expression. The IC50 values of MNNG/HOS and its resistant variant to DXR were higher than those of MG63 and its resistant clone. Multidrug-resistant associated protein (MRP) mRNA expression was detected in MNNG/HOS or MG63 parental cell lines, and in their resistant variants. MG63 and its resistant variants revealed stable expression of MRP, whereas the resistant phenotype of MNNG/HOS showed decreased MRP expression, compared to its parental cell line. No alteration in the levels of hepatocyte growth factor (HGF) or its receptor c-MET was recognized between parental lines and their resistant variants. The results indicate that our DXR-resistant variants of MNNG/HOS and MG63 reveal a classical MDR phenotype and can offer a model with which to investigate the mechanisms of multidrug resistance in osteosarcoma. PMID- 10854559 TI - Clinical significance of mucin staining in keratinizing-type squamous cell carcinoma of the uterine cervix. AB - Tumor tissue from 57 cases of keratinizing-type squamous cell carcinoma of the uterine cervix that had been stored in the archives of Osaka City University Hospital between May 1986 and October 1994 were stained with periodic acid-Schiff (PAS) reagent following amylase and alcian blue to identify the presence of intracellular mucin and to assess the value and significance of demonstrating the presence of mucin. Nineteen specimens (33.3%) stained positive for mucin with alcian blue or PAS following diastase. The results of mucin staining were not significant in terms of survival by Kaplan-Meier's analysis. PMID- 10854560 TI - Expression of tissue inhibitor of matrix metalloproteinase-1 in human breast carcinoma. AB - Tissue inhibitor of metalloproteinase-1 (TIMP-1) is an inhibitor of matrix metalloproteinase (MMP) in human carcinomas. TIMP-1 is thus considered to inhibit carcinoma invasion and metastasis. On the other hand, recent reports have disclosed that TIMP-1 also possesses a growth-promoting function. The clinical significance of TIMP-1 expression has not been fully determined in breast carcinoma. We thus examined the expression of TIMP-1 mRNA in tumor tissues of 100 breast carcinoma cases by a reverse transcriptase-polymerase chain reaction assay. The expression of TIMP-1 mRNA in each case was evaluated semi quantitatively with adjustment for the TIMP-1 expression in a control breast carcinoma cell line, MCF7. There was a significant inverse correlation between the TIMP-1 expression and lymph node metastasis (p<0.05). A multivariate analysis disclosed that TIMP-1 expression status was an independent determinant factor for lymph node metastasis. In addition, the tumors with positive estrogen or progesterone receptors showed a higher TIMP-1 mRNA expression than those without the receptors, but this did not reach statistical significance. The findings suggest that the breast tumors with high TIMP-1 expression might show less malignant potential than those with low TIMP-1 expression. PMID- 10854561 TI - Comparison of 5-FU and leucovorin to gemcitabine in the treatment of pancreatic cancer. AB - Gemcitabine has been recently added to the 5-fluorouracil (5-FU) protocol in the treatment of both new and 5-FU refractory patients with pancreatic cancer. We compared the efficacy of gemcitabine versus 5-FU and leucovorin (LCV) in 82 patients with advanced pancreatic cancer. Forty-seven patients received 5-FU and LCV and 35 patients received gemcitabine. Objective responses were documented in 4% on the 5-FU and LCV arm, compared to 9% in the gemcitabine arm. No change was observed in 48% on the 5-FU and LCV arm compared to 20% in the gemcitabine arm. Clinical benefit was observed in 19% on the 5-FU and LCV arm compared to 48% in the gemcitabine arm (p<0. 001). Toxicity was mild and well tolerated in both arms. One-year survival was 32% on the 5-FU and LCV arm and 23% in the gemcitabine arm. These results indicate that gemcitabine had a significantly higher clinical benefit than 5-FU and should be the standard treatment in advanced pancreatic cancer. PMID- 10854562 TI - Expression of soluble urokinase plasminogen activator receptor may be related to outcome in prostate cancer patients. AB - The urokinase-type plasminogen activator receptor (uPAR) exists as a GPI anchored glycoprotein (Mr=50-60 kDa) on the surface of various cell types. This receptor can be bound by or cleaved by urokinase. The cleaved receptor, soluble urokinase type plasminogen activator receptor (suPAR), with an Mr=35 kDa has no known physiological function and can be identified circulating in the blood of normal individuals. Although no function has been characterized, the soluble receptor has been reported to be of clinical significance. The objective of this study is to characterize novel serum markers that can be used for the early detection of prostate cancer and to predict patient prognosis. Thirty-nine patients at the University of Yaounde I, Yaounde, Cameroon, West Africa were examined for prostatic disorders. Of these, 46% were diagnosed with benign prostate hyperplasia (BPH), while 44% of the patients were diagnosed via biopsy with prostate cancer and graded accordingly. Here we show that serum from patients with BPH or prostate cancer contains elevated levels of suPAR. To examine the significance of suPAR as a diagnostic factor, we used a suPAR ELISA kit and compared these results with serum levels of prostate specific antigen (PSA), the current diagnostic marker for prostate cancer. PSA and serum suPAR levels in BPH and cancer patients were greatly elevated in the majority of patients, while others had undetectable levels of either. Serum levels of suPAR were high in cancer patients as well as, although to a lesser degree, in patients with BPH. Cancer patients who died during the follow-up period were found to have consistently higher serum suPAR levels than correlating serum PSA levels. These preliminary findings are the first evaluating serum suPAR levels as a possible diagnostic marker for the early detection of prostate cancer and for the prediction of patient prognosis. PMID- 10854563 TI - A phase II evaluation of bexarotene (Targretin) capsules in patients with metastatic melanoma. AB - A phase II study was undertaken to determine the efficacy of Bexarotene in melanoma. Between November 1997 and April 1998, 19 patients were given Bexarotene in single daily oral doses of 450 mg/m2 in capsule form continuously. Nineteen patients, four with choroidal metastatic melanoma, were treated. No responses were seen. Five patients had stable disease, two of the four with choroidal melanoma, had tumor progression. Myelosuppression was mild. Grade 3 myalgia, asthenia, diarrhea, cold hands/feet, and mood changes were seen in one patient each. Changes in serum triglyceride and thyroxine levels were common. Bexarotene, as used in this study, is not effective against melanoma. PMID- 10854564 TI - Left ventricular function in colon cancer patients receiving adjuvant fluoro folate chemotherapy: an echocardiographic study. AB - Cardiotoxicity has been reported with increasing frequency after 5-FU administration. Since 5-FU-based regimens are increasingly used as adjuvant treatment for resected colon cancer patients, this carries a potential increased risk of cardiotoxicity. In the present study, we evaluated if fluoro-folate chemotherapy has any effect on left ventricular (LV) diastolic function in resected colon cancer. Twenty-five resected colon cancer patients receiving adjuvant fluoro-folate chemotherapy, were prospectively studied. Both digitized M mode and Doppler echocardiography were performed, in order to investigate different aspects of LV diastolic function. None of the studied parameters was influenced by chemotherapy administration. Indeed, values recorded at the end of the 6-month treatment and 6 months later were not statistically different from baseline values. This study suggests that the use of fluoro-folate chemotherapy in otherwise normal subjects carries a very low risk of severe cardiotoxicity. PMID- 10854565 TI - Treatment of large cell lymphoma with the Fi2 regimen (doxorubicin, vincristine, cyclophosphamide, bleomycin and prednisone): 25-year experience. AB - The CHOP protocol is the reference treatment for large cell lymphomas, but several other schemes of different intensity have recently been studied with controversial clinical findings. We report here the results obtained at our institution with a CHOP-like regimen called Firenze 2 (Fi2), which is characterised by an original scheduling of chemotherapy administration. A total of 225 patients, who were diagnosed from 1974 to 1996, were included in this retrospective study. All patients received the Fi2 regimen as a first-line intervention. One-hundred and sixty-two (72%) achieved complete remission; the overall survival at 120 months was 51% with a disease-free survival of 67% (median follow-up = 78 months). The survival curve showed a stable plateau of 42% after 16 years, which remained stable for further 4 years. In a multivariate survival analysis, achievement of complete remission (p<0.001) and IPI index of 0 or 1 (p=0.05) were significantly associated with a better survival. Overall, the outcome of our patients was similar to that reported by others, but the distinguishing feature of our study is the very long follow-up of the patients. Our study confirms that first generation regimens are effective and can cure a substantial proportion of patients. The long-term results of our study are helpful to retrospectively identify high-risk patients whose prognosis is poor and who can be candidates for more aggressive schemes of chemotherapy. PMID- 10854566 TI - Test for fused-cell origin of tumors with Mus caroli carrying glucose-6-phosphate dehydrogenase cellular mosaicism. AB - A test system for examining the fused cell-origin of tumors was developed by application of Mus caroli carrying the X-chromosome inactivation cellular mosaicism for G6PD. A G6PD heterodimer pattern is expected if a tumor is initiated from a fused cell. Among tumors induced by subcutaneous injection of a high dose of MCA, three tumors exhibited a single G6PD phenotype and one tumor exhibited a multiple G6PD phenotype; however, the G6PD heterodimer pattern was not found. Although the results obtained were not conclusive, this system is thought to be useful for detecting a possible fused cell-origin of tumors. PMID- 10854567 TI - Is reoperation for recurrence of glioblastoma justified? AB - The purpose of the present study was to determine the effect of surgery on the time length and quality of survival in patients with recurrent glioblastoma multiforme. Two groups were compared; the first included 18 patients who underwent surgery at the time of tumour recurrence. The second group included 36 patients who did not undergo surgery at the time of tumour recurrence. Both groups were matched according to the following criteria: gender, age, Karnofsky Performance Scale (KPS) score, at the time of initial surgery and of tumour recurrence, extent of initial surgery, interval between initial surgery and tumour reccurence. Both groups received conventional treatment after initial surgery. There are no statistically significant differences between the two groups as regards to the previously mentioned criteria. After tumour recurrence, the median survival time was 5 months in the group of patients undergoing a second resection and 2 months in the group of patients not undergoing repeat surgery. The difference was statistically significant on univariate analysis. Moreover, the median length of time spent in an acceptable condition (KPS >/=60) from the time of tumour recurrence was found to be significantly longer in patient who underwent a second resection (4 months) compared with patients who did not undergo repeat surgery (1 month). Even in a relatively favorable subgroup of reoperated patients, the survival benefit although significant was only 3 months. It was impossible to completely match the two groups of patients suggesting that the difference might have been even less. Although symptomatic improvement is modestly achieved by repeat surgery, its transient nature necessitates clear discussion with patient and family on an individual basis. PMID- 10854568 TI - Detection of human papillomavirus types 16 and 18 in human neoplastic endometrium: lack of correlation with established prognostic factors. AB - HPV (types 16 and 18) DNA sequences are present in the majority of precancerous and cancerous lesions of the human uterine cervix. However, data concerning the involvement of HPVs infection in the pathogenesis of endometrial cancer are controversial. In the current study we investigated the frequency of the HPV types 16 and 18, detected by PCR amplification using the type 16- and 18-specific primers within the E7 Open Reading Frame (ORF) sequence, in 54 human endometrial carcinomas obtained from women of Polish origin. Moreover, we assessed the possible association of the HPV with the clinicopathological features of the cancer, patients' outcome as well as with the K-ras codon 12 gene point mutations. HPV type 16 was present in eleven out of 54 (20%) endometrial tumors, while HPV type 18 was detected only in three out of 54 (4%) neoplasms analyzed. HPV infection was not related either to the patients' age (r=0.11; p=0.428, Spearman correlation test) or to the clinicopathological parameters and patients' prognosis. A higher incidence of HPV 16/18 was detected in well (G1) differentiated than in moderately (G2) and poorly (G3) differentiated endometrial adenocarcinomas, but the difference was not statistically significant. Moreover, none of HPV-positive endometrial carcinomas harbored K-ras codon 12 gene point mutations. Our results suggest that some of the endometrial carcinomas are associated with HPV infection but the presence of the human papillamovirus types 16/18 is not related to the clinicopathological or prognostical features of the neoplasm. PMID- 10854569 TI - Presence of apolipoprotein E immunoreactivity in degenerating neurones of mice is dependent on the severity of kainic acid-induced lesion. AB - Apolipoprotein E (apoE) is a major apolipoprotein in the central nervous system (CNS) that may play a role in various CNS disorders. ApoE is primarily localised in astrocytes, but neuronal apoE mRNA expression has been demonstrated in normal and diseased human brain, as well as in ischaemic rat brain. To obtain further insight into the role of apoE in neuronal degeneration in the CNS and conditions of neuronal apoE localisation, we have investigated in mice the distribution of apoE following neuronal injury induced by kainic acid (n=35, 25 or 35 mg kainic acid/kg BW). Consecutive series of brain sections were immunostained for apoE and markers for astroglia (GFAP) and microglia/macrophage cells (CR3). Degenerating neurones were identified with a silver-degeneration staining technique. The intensity and cellular distribution of apoE-immunoreactivity (apoE-ir) was dependent on the severity of neuronal injury. Mice that developed mild neuronal degeneration, restricted to a subset of neurones in the hippocampus, showed increased apoE-ir in astrocytes concomitant with increased GFAP-ir and mild microgliosis. In these mice, no neuronal apoE-ir was detected. In contrast, mice developing severe neuronal injury in the hippocampus - frequently also showing degeneration in other brain regions including cortex, thalamus, striatum and amygdala - showed intense apoE-ir in degenerating neurones. Surrounding the lesion, apoE-ir was increased in neuropil recurrently whereas GFAP-ir astrocytes disappeared. Thus, in mice apoE accumulates in degenerating neurones in conditions of severe neuronal injury putatively in association with disruption of the glial network. PMID- 10854570 TI - Effects of xanthine derivatives on electroretinographic responsiveness. AB - In view of the use of synthetic propentofylline (PPF) as a protective agent in brain ischemia, its possible side effects on vision capacities have been explored by electroretinography in comparative experiments with theophylline. We used eyecup preparations of small-spotted dogfish sharks and of European eels, particularly suitable for long-lasting experiments. The drug exerted profound but reversible modifications of ERG records: (1) a dose-dependent increase of the amplitude and duration of the chemically isolated late receptor potential (LRP), (2) a partial unmasking of LRP, (3) a strong potentiation of the LRP-unmasking effect of low temperature, (4) a potentiation of light adaptation effects, and (5) a strong potentiation of the post-illumination hyperexcitability. The effects were explicable as due to a strong phosphodiesterase (PDE) inhibiting, cyclic guanosine monophosphate (cGMP) promoting, action of the drug. The effects were considerably stronger, or even of opposite sign, in comparison to those of the chemically related theophylline. PPF did not seriously affect the ERG c-wave originating in the pigment epithelium. The results suggested that the effects of PPF on vision may not seriously hamper the therapeutic use of the drug. They indicated, on the other hand, that PPF was a retinoactive drug of potential usefulness in the exploration of the complex biochemical events underlying visual transduction. PMID- 10854571 TI - 3,4-Dihydroxyphenylacetaldehyde potentiates the toxic effects of metabolic stress in PC12 cells. AB - 3,4-Dihydroxyphenylacetaldehyde (DOPAL) is a toxic metabolite formed by the oxidative deamination of dopamine. This aldehyde is mainly oxidized to 3,4 dihydroxyphenylacetic acid (DOPAC) by aldehyde dehydrogenase (ALDH), but is also partly reduced to 3, 4-dihydroxyphenylethanol (DOPET) by aldehyde or aldose reductase (ARs). In a previous study, we found that rotenone, a complex I inhibitor, induced a rapid accumulation of DOPAL and DOPET in the medium of cultured PC12 cells. Here, we examined the potential role of DOPAL in the toxicity induced by complex I inhibition in PC12 cells and compared the effects of rotenone on concentrations of DOPAL and DOPET to those of MPP(+). DOPAL and DOPET levels were increased by rotenone but decreased by MPP(+). Inhibition of ALDH by daidzein reduced the formation of DOPAC and increased the accumulation of DOPAL. Inhibition of ARs (with AL1576) diminished DOPET formation and elevated DOPAL concentrations. Combined inhibition of ALDH and ARs markedly elevated DOPAL concentrations while diminishing DOPET and DOPAC levels. The elevation of DOPAL levels induced by combined inhibition of ALDH and ARs had no effect on cell viability. However, combined inhibition of ALDH and ARs potentiated rotenone induced toxicity. Both the potentiation of toxicity and the increase in DOPAL levels were blocked by inhibition of monoamine oxidase with clorgyline indicating that accumulation of DOPAL was responsible for the potentiated rotenone-induced toxicity following combined inhibition of ALDH and ARs. Since complex I dysfunction is reported to be involved in the pathogenesis of Parkinson's disease, DOPAL potentiation of the deleterious effects of complex I inhibition may contribute to the specific vulnerability of dopaminergic neurons to injury. PMID- 10854572 TI - Role of bicarbonate ion in mediating decreased synaptic conductance in benzodiazepine tolerant hippocampal CA1 pyramidal neurons. AB - Chronic flurazepam treatment substantially impairs the function of GABAergic synapses on hippocampal CA1 pyramidal cells. Previous findings included a significant decrease in the synaptic and unitary conductance of CA1 pyramidal neuron GABA(A) receptor channels and the appearance of a GABA(A)-receptor mediated depolarizing potential. To investigate the ionic basis of the decreased conductance, whole-cell voltage-clamp techniques were used to record evoked, GABA(A) receptor-mediated IPSCs carried by HCO(3)(-)-Cl(-) or Cl(-) alone. Hippocampal slices were prepared from rats administered flurazepam orally for 1 week, 2 days after ending drug treatment. Slices were superfused with HCO(3)(-) aCSF or with HEPES-aCSF (without HCO(3)(-)) plus 50 microM APV and 10 microM DNQX. The micropipette contained 130 mM CsCl and 1 microM QX-314. GABA(A) receptors located on pyramidal cell somata or dendrites were activated monosynaptically by maximal stimulation of GABAergic terminals at the stratum oriens-pyramidale (SO-SP) or stratum lacunosum-molecular (S-L-M) border, respectively. In HCO(3)(-)-aCSF, there was a significant reduction in synaptic conductance in flurazepam-treated neurons following both SO-SP (control: 1058 pS, flurazepam: 226 pS, P<0.01) and S-L-M (control 998 pS, flurazepam: 179 pS, P<0.01) stimulation, as well as the total charge transfer, indicating a decreased HCO(3)(-)-Cl(-) flux. In HEPES-aCSF, the synaptic conductance and total charge transfer, and thus Cl(-) flux, was unchanged in flurazepam-treated neurons (SO SP: control 588 pS, flurazepam: 580 pS, P>0.05; S-L-M: control 595 pS, flurazepam: 527 pS, P>0.05). Taken together, these findings suggest that a reduction in HCO(3)(-) flux may play a prominent role in mediating the action of GABA and that a loss of HCO(3)(-) conductance may significantly contribute to impaired GABA(A) receptor function after chronic benzodiazepine treatment. PMID- 10854573 TI - Axotomy of the sciatic nerve differentially affects expression of metabotropic glutamate receptor mRNA in adult rat motoneurons. AB - Previous studies indicated that axotomy exposes motoneurons to glutamatergic excitotoxic stress and protection from glutamatergic overactivation might be crucial for survival. Depending on the experimental model and the subtype involved, activation of metabotropic glutamate receptors (mGluRs) may either enhance excitotoxicity or exert protective effects. To investigate a possible involvement of mGluRs in neuronal rescue mechanisms after axotomy we have monitored the distribution of mGluR mRNA with in situ hybridization in adult rat motoneurons 1, 2, 3, and 4 weeks after sciatic nerve transection. Motoneurons in sham-operated control animals expressed mGluR 1, 4, and 7 mRNA. The mGluR1 mRNA signal was reduced to 49.6+/-6.9% as compared to the contralateral side 2 weeks after axotomy and 31.2+/-8.3% after 4 weeks. The mGluR4 signal declined to 22.1+/ 5.1% after 1 week and 10.2+/-1.6% after 2 weeks, remaining stable thereafter. During the entire observation period the mRNA for mGluR7 was not significantly altered. Axotomy did not change the overall number of motoneurons on the ipsi- or contralateral side. The differential regulation of mGluR subtypes may be part of an adaptive cell program that helps to rescue adult motoneurons from excitotoxic cell death during the stress induced by peripheral denervation. PMID- 10854574 TI - Vinpocetine protects from aminoglycoside antibiotic-induced hearing loss in guinea pig in vivo. AB - The principal objective of this study is to explore the hypothesis that a blockade of Na(+) channels can prevent some of the mechanisms involved in ototoxicity. For this purpose, the potential action of the voltage sensitive Na(+) channel antagonist, vinpocetine, on the ototoxicity induced by the representative aminoglycoside antibiotic, amikacin, in guinea pigs was tested for almost half a year. Amikacin (450 mg/kg) administered daily (i.m.) for 5 days increases the thresholds of the auditory brainstem response (ABR) to the two frequencies tested (4 and 8 kHz). These threshold increases are permanent or at least long-lived, as after 40 days they are already established and are maintained until the end of the experiment (160 days after the antibiotic administration). Amikacin decreases the amplitude of ABR waves, particularly P1, and after 160 days increases the latency of ABR waves, particularly at the higher frequency tested (8 kHz). When the above amikacin regimen is followed by a daily (i.p.) vinpocetine (2 mg/kg) administration for 13 days the increase in ABR threshold and latency caused by amikacin alone is prevented. Moreover, the animals treated with amikacin alone show a decreased weight gain and a remarkable increased mortality in comparison with the group of animals post-treated with vinpocetine. We hope that the multiple beneficial effects exerted by the Na(+) channel blocker, vinpocetine, against aminoglycoside antibiotics-induced side effects could help to solve the serious limitations of the use of this type of antibiotic. PMID- 10854575 TI - Parathyroid hormone-related protein is expressed by transformed and fetal human astrocytes and inhibits cell proliferation. AB - Parathyroid hormone-related protein (PTHrP) and the PTH/PTHrP receptor are expressed in most normal tissues, including brain, where PTHrP is though to act locally in an autocrine or paracrine fashion. Previous in situ localization studies in adult rodents have documented CNS PTHrP expression in neurons but not in glial cells. However, a recent report describing immunoreactive PTHrP in human astrocytomas suggests that PTHrP expression may be a marker of dedifferentiation and/or malignant transformation in glial cells. To begin to test this hypothesis, constitutive and regulated PTHrP expression were examined in cultured fetal and transformed (U-373 MG) human astrocytes. PTHrP was expressed in untreated fetal astrocytes and U-373 MG cells, as determined by Northern analysis, immunocytochemical staining, and detection of PTHrP(1-84) protein in conditioned media. Epidermal growth factor and tumor necrosis factor, important growth factors in astrocyte development and malignant transformation, stimulated PTHrP expression in both cell types. Treatment of U-373 MG cells or fetal astrocytes with PTHrP(1-34) consistently inhibited cellular proliferation, as measured by [(3)H]-thymidine incorporation. These findings suggest that PTHrP, a peptide whose expression is induced by mitogens in both immature and transformed human astrocytes, may feedback inhibit cellular proliferation, an effect that may be of importance during malignant transformation as well as CNS development. Furthermore, when combined with previous evidence of PTHrP expression by PTH/PTHrP receptor-positive neurons, our demonstration of regulated PTHrP expression by receptor-positive astrocytes identifies PTHrP as a potential peptide mediator of cross-talk between glial cells and neurons. PMID- 10854576 TI - Changes in REM sleep occurrence due to rhythmical auditory stimulation in the rat. AB - The effects of the rhythmical delivery of an auditory stimulus (1000 Hz, from 50 to 100 dB, 20 ms, every 20 s) on the pattern of rapid eye movement (REM) sleep occurrence was studied in the rat. The stimulation was simultaneously carried out on pairs of rats over 5 consecutive days (10-h recording sessions), during which a tone of increasing intensity (50, 63, 75, 88, 100 dB) was used. In each experimental session, auditory stimulation was triggered by the REM sleep occurrence of one rat (REMS-selective stimulation) whilst the other rat simultaneously received the same stimuli, but during any stage of the wake-sleep cycle (REMS-unselective stimulation). The results showed that the total amount of REM sleep in the 10-h recording session was increased over the 5 days of stimulation in the REMS-unselective group. This effect was due to an increase in the mean duration of REM sleep episodes. However, no significant changes were observed in animals under REMS-selective stimulation, nor in a third group of animals in which the spontaneous evolution of REM sleep occurrence (REMS spontaneous) was studied. Since 86% of the stimuli under the REMS-unselective auditory stimulation fell outside REM sleep, the result would suggest that REM sleep occurrence is affected when the stimuli are delivered during a time period (i.e. during wakefulness or non-REM sleep) in which it is well known that physiological regulations are fully operant. PMID- 10854577 TI - Expression of the long form of leptin receptor (Ob-Rb) mRNA in the brain of mouse embryos and newborn mice. AB - The long form of the leptin receptor (Ob-Rb) has a cytoplasmic domain which activates the JAK-STAT signal transduction pathway. It is related to appetite and energy expenditure and is expressed in various parts of the brain in adults. In embryos, however, the detailed distribution of Ob-Rb expression sites and the function of the leptin-Ob-Rb system remain unclear, although leptin is detected in human cord plasma and leptin mRNA is detected in mouse embryos. In this study, we investigated the Ob-Rb mRNA expression pattern in the brains of mouse embryos and newborn mice by RT-PCR and in situ hybridization. At embryonic day 10.5 (E10.5), Ob-Rb mRNA was already detected in the brain by RT-PCR. By in situ hybridization, Ob-Rb mRNA was observed in the ventricular zone of the rhombencephalon at E11.5. At E12.5, it was also expressed in the ventricular zone of the telencephalon, mesencephalon and cerebellar primordium. From E14. 5 it was expressed in the cortical plate of the telencephalon and the ventricular zone of the thalamus. At E16.5, it was expressed in the premamillary hypothalamic nucleus, superficial gray matter of the superior colliculus, external germinal and Purkinje cell layers of the cerebellum, and facial nucleus. At E18.5, it was expressed in the arcuate nucleus and ventromedial hypothalamic nucleus. These results suggest that the leptin-Ob-Rb system is related to brain development. PMID- 10854578 TI - Theiler's murine encephalomyelitis virus induces rapid necrosis and delayed apoptosis in myelinated mouse cerebellar explant cultures. AB - Infection with the Daniel strain of Theiler's murine encephalomyelitis (TMEV-DA) virus induces persistent demyelinating lesions in mice and serves as a model for multiple sclerosis. During the acute phase of the disease, however, viral infection leads to cell death in vivo. Viral-induced death may result directly from viral infection of neural cells, or indirectly, by activation of the immune system. To examine the direct effects of TMEV infection on neural cells, myelinated explant cultures of the murine cerebellum were infected with 10(5) pfu of TMEV-DA for periods ranging from 1 to 72 h. Our results indicate that TMEV-DA replicates in cultured neural tissue. Initially, viral antigen is localized to a few isolated neural cells. However, within 72 h antigen was observed in multiple foci that included damaged cells and extracellular debris. Viral infection led to a rapid and cyclical induction of necrosis with a time period that was consistent with the lytic phase of the viral life-cycle. Simultaneously, we observed an increase in apoptosis 48 h post-infection. Electron micrographic analysis indicated that viral-infected cultures contained cells with fragmented nuclei and condensed cytoplasm, characteristic of apoptosis. The localization of apoptosis to the cerebellar granule cell layer, identified these cells as presumptive granule neurons. Viral infection, however, did not lead to myelin damage, though damaged axons were visible in TMEV-infected cultures. These results suggest that during the acute phase of infection, TMEV targets neural cells for apoptosis without directly disrupting myelin. Myelin damage may therefore result from the activation of the immune system. PMID- 10854579 TI - Opposite effects of glutamate antagonists and antiparkinsonian drugs on the activities of DOPA decarboxylase and 5-HTP decarboxylase in the rat brain. AB - This study measured the activities of L-DOPA and 5-HTP decarboxylase (DDC and 5 HTPDC) in the substantia nigra and corpus striatum of reserpine-treated rats. Acute injection of the NMDA receptor antagonists CGP 40116 (5 mg/kg) and HA 966 (5 mg/kg), and to a lesser extent eliprodil (10 mg/kg), greatly elevated DDC in both structures, whilst having no effect on (nigra) or inhibiting (striatum) 5 HTPDC. L-DOPA (25 mg/kg) on its own inhibited both enzymes in either brain region. The weak NMDA receptor-channel blockers (and antiparkinsonian drugs) budipine (10 mg/kg), memantine (40 mg/kg) and amantadine (40 mg/kg) strongly increased DDC, whilst not affecting or decreasing 5-HTPDC activity in nigra and striatum. The L-DOPA-induced suppression of DDC was mostly reversed by all three antiparkinsonian drugs, whilst L-DOPA-induced inhibition of 5-HTPDC was only reversed by CGP 40116 (striatum only). It is concluded that glutamate exerts a differential physiological influence on the biosynthesis of dopamine and 5-HT in the brain, by tonically suppressing DDC and tonically stimulating 5-HTPDC. The L DOPA-induced reduction in DDC may help to explain the eventual loss of efficacy of L-DOPA therapy in parkinsonian patients. It is suggested, however, that it may be possible to extend the lifetime of L-DOPA therapy with drugs which potentiate the activity of DDC, such as budipine and the 1-aminoadamantanes. PMID- 10854580 TI - Discrimination between changes in glucocorticoid receptor expression and activation in rat brain using western blot analysis. AB - These studies investigated autoregulation of glucocorticoid receptor (GR) protein expression and activation in rat brain using western blot methodology. By comparing GR immunoblot reactivity present in various tissue subcellular preparations we were able to discriminate between corticosterone-induced changes in GR activation or GR protein expression. Our cytosolic tissue preparation yielded a similar pattern of treatment effects on relative GR as measured by receptor binding assay or western blot. In both cases, short-term adrenalectomy (18 h) produced no change in cytosolic GR. On the other hand, long-term adrenalectomy (3-14 days) resulted in a large increase in cytosolic GR, whereas acute (1-2 h) treatment with high dose corticosterone produced a large decrease in cytosolic GR. Western blot measurement of GR levels in a nuclear extract or whole-cell extract from the same brains indicated that acute corticosterone treatment produced a large increase in nuclear GR and no change in whole-cell GR. Thus, all of the decrease in cytosolic GR observed after acute corticosterone treatment could be accounted for by receptor redistribution to the nuclear tissue fraction as opposed to rapid receptor protein downregulation. Long-term treatment of rats with adrenalectomy or high dose corticosterone produced a large increase and decrease, respectively, in whole-cell GR, indicating genuine changes in receptor protein expression. These studies indicate that in vivo regulation of GR protein expression in the rat brain can be studied using western blot analysis of a whole-cell tissue preparation. This procedure has an important advantage over receptor binding studies in that GR protein expression can be measured in adrenal intact rats. These studies also support the validity of using cytosolic receptor binding assays to estimate relative changes in GR occupation/activation when appropriate comparison groups are included. PMID- 10854581 TI - Decreased GABA and increased glutamate receptor-mediated activity on inferior colliculus neurons in vitro are associated with susceptibility to ethanol withdrawal seizures. AB - Cessation of ethanol administration in ethanol-dependent rats results in an ethanol withdrawal (ETX) syndrome, including audiogenic seizures (AGS). The inferior colliculus (IC) is the initiation site for AGS, and membrane properties of IC neurons exhibit hyperexcitability during ETX. Previous studies observed that ETX alters GABA and glutamate neurotransmission in certain brain sites. The present study evaluated synaptic properties and actions of GABA or glutamate antagonists during ETX in IC dorsal cortex (ICd) neurons in brain slices from rats treated with ethanol intragastrically 3 times daily for 4 days. A significant increase of spontaneous action potentials (APs) was observed during ETX. The width, area and rise time of excitatory postsynaptic potentials (EPSPs) evoked by stimulation in the commissure of IC were significantly elevated during ETX. A fast EPSP was sensitive to block by the non-NMDA receptor antagonist, 6 cyano-7-nitroquinoxaline-2,3-dione (CNQX), and a slow EPSP was sensitive to the NMDA receptor antagonist, 2-amino-5-phosphonovalerate (AP5). However, during ETX the concentration of CNQX or AP5 needed to block these EPSPs was elevated significantly. Inhibitory postsynaptic potentials (IPSPs) in ICd neurons evoked in both normal and ETX rats were blocked by the GABA(A) antagonist, bicuculline. However, IPSPs during ETX displayed a significantly greater sensitivity to bicuculline. These data indicate that decreased GABA(A)-mediated inhibition and increased glutamate-mediated excitability in IC may both be critical mechanisms of AGS initiation during ETX, which is similar to observations in a genetic form of AGS. The common changes in IC neurotransmission in these AGS forms may be general mechanisms subserving AGS and other forms of auditory system pathophysiology in which the IC is implicated. PMID- 10854582 TI - Development of NR1, NR2A and NR2B mRNA in NR1 immunoreactive cells of rat visual cortex. AB - In cells marked for N-methyl-D-aspartate receptors (NMDARs), we studied the relationship between the sensitive period for monocular deprivation and the expression of rat NMDAR subunits, NR2A and NR2B. In the rat the sensitive period ends sometime after postnatal day 50 (P50). Previous studies of the development of these subunit mRNAs focused on animals prior to the end of the sensitive period and did not examine the visual cortex specifically. We used a monoclonal antibody to the NR1 subunit of the receptor to identify cells containing NMDARs. We then used in situ hybridization to label the same sections for NR2A or NR2B mRNA. In an additional experiment we labeled sections for NR1 mRNA to see if the developmental profile was similar at both the mRNA and protein level. We used five animals at each of four ages: P22, P30, P45 and P90. Staining for NR2B mRNA, but not for NR2A mRNA, decreased dramatically from P22 to P45. Staining for NR1 mRNA declined dramatically between P22 and P45 even though most cells remained strongly immunopositive for the NR1 protein during this time. This discrepancy suggests that significant NR1 regulation occurs after gene transcription. Because most of the decrease in NR1 mRNA and NR2B mRNA occurs by P30, transcriptional regulation of these subunits does not easily explain the loss of sensitivity to monocular deprivation, which occurs around P50. The changes are, in fact, more closely synchronized with the beginning of experience-dependent plasticity than with its end. PMID- 10854583 TI - Neuroanatomical specificity of prolactin-induced hyperphagia in virgin female rats. AB - Intracerebroventricular (i.c.v.) administration of PRL increases food intake in virgin female rats but the brain site(s) at which PRL acts to promote feeding behavior is not known. The present studies investigated the role of the paraventricular nucleus (PVN), ventromedial nucleus (VMH), and medial preoptic nucleus (MPOA) in the hyperphagic actions of PRL. Ad-libitum-fed virgin female rats received twice daily site-specific injections of PRL (800 ng) over a period of 10 days. Only subjects demonstrating regular vaginal cyclicity were included in the study. Food intake, body weight, and vaginal cyclicity were measured daily. Results showed that PRL significantly increased food intake when injected into the PVN. A nonsignificant trend towards a hyperphagic response in the last 5 days of testing was observed in rats receiving intra-VMH injections of PRL, and the MPOA was not responsive to the feeding-stimulating properties of PRL. None of the manipulations affected body weight or vaginal cyclicity as demonstrated by vaginal smears. In sum, the present results reveal that one brain site at which PRL acts to increase food intake is the PVN, but these studies do not rule out the possibility that the effects of PRL on food intake may also involve other brain areas. PMID- 10854584 TI - Peripheral nerve regeneration across an 80-mm gap bridged by a polyglycolic acid (PGA)-collagen tube filled with laminin-coated collagen fibers: a histological and electrophysiological evaluation of regenerated nerves. AB - We evaluated peripheral nerve regeneration across an 80-mm gap using a novel artificial nerve conduit. The conduit was made of a polyglycolic acid (PGA) collagen tube filled with laminin-coated collagen fibers. Twelve beagle dogs underwent implantation of the nerve conduit across an 80-mm gap in the left peroneal nerve. In four other dogs used as negative controls, the nerve was resected and left unconnected. Histological observation showed that numerous unmyelinated and myelinated nerve fibers, all smaller in diameter and with a thinner myelin sheath than normal nerve fibers, regrew through and beyond the gap 12 months after implantation. The distribution of the regenerated axonal diameters was different from that of the normal axonal diameters. Compound muscle action potentials, motor evoked potentials, and somatosensory evoked potentials were recorded in most animals 3 months after implantation. Peak amplitudes and latencies recovered gradually, which indicating the functional establishment of the nerve connection with the target organs. In addition to the ordinary electrophysiological recoveries, potentials with distinct latencies originating from Aalpha, Adelta and C fibers became distinguishable at the 6th lumbar vertebra following stimulation of the peroneal nerve distal to the gap 12 months after implantation. The pattern of walking without load was restored to almost normal 10-12 months after implantation. Neither electrophysiological nor histological restoration was obtained in the controls. Our nerve conduit can guide peripheral nerve elongation and lead to favorable functional recovery across a wider nerve gap than previously reported artificial nerve conduits. PMID- 10854585 TI - Retention of concurrent taste aversion learning after electrolytic lesioning of the interpositus-dentate region of the cerebellum. AB - Lesions in the interpositus-dentate region of the cerebellum impair short-term, or concurrent, TAL. In this type of learning, animals must discriminate between two flavor stimuli presented at the same time, one of which is associated with an aversive product. The task is learned by the control animals, and within this group the animals that acquire it adequately enough (15/22, 70% criterion) retain the learned taste discrimination when they are subjected to it again after being lesioned in the interpositus-dentate region. These results suggest that the deep nuclei are essential in the concurrent TAL acquisition process, but not in its retention. PMID- 10854586 TI - Potassium depolarization-induced cAMP stimulates somatostatin mRNA levels in cultured diencephalic neurons. AB - In the nervous system, signals transmitted across synapses are known to regulate gene expression in the postsynaptic cells. This process often involves membrane depolarization and subsequent elevation of intracellular Ca(2+). We have previously demonstrated in fetal cerebrocortical cells, that somatostatin (SS) mRNA levels can be induced by depolarizing agents such as high potassium concentrations and veratridine (VTD), and that these effects are calcium dependent. SS expression is regulated by cAMP, and in the cerebral cortex adenylate cyclase activity is regulated through fluctuations in intracellular Ca(2+) concentrations. The present experiments were undertaken to determine the mechanism by which calcium upregulates the levels of SS mRNA. Cerebrocortical cells from 17-day-old fetuses were exposed to the different agents for 24 h and the levels of SS mRNA were measured by Northern blot. Incubation of cells with the calcium channel antagonist nifedipine (Nf), the calcium chelating agent EGTA, calcium free KRB and the calcium calmodulin inhibitors trifluoroperazine (TFP) and the napthelene sulfonamide, W7, resulted in the inhibition of K(+)-induced SS mRNA levels. K(+)-evoked depolarization increased the intracellular concentration of cAMP and this effect was antagonized by verapamil (VPM). Forskolin (Fk) provoked a higher increment in cAMP concentration than potassium, although the induction of SS mRNA was more evident following K(+) depolarization indicating a lack of correlation between levels of cAMP and induction of SS mRNA. The role of K(+)-induced cAMP on the increment of SS mRNA that occurred upon membrane depolarization was further explored with the inhibitor of protein kinase A (PKA), Rp cAMP whose presence significantly reduced depolarization-induced SS mRNA levels. This study confirms that Ca(2+) influx is required for K(+)depolarization induced stimulation of cAMP whereby the increment of SS mRNA is partly produced. PMID- 10854587 TI - Effects of acidosis on the post-hypoxic recovery of synaptic transmission in gerbil hippocampal slices. AB - We investigated the effects of acidosis on the hypoxic neuronal damage using gerbil hippocampal slices. Acidosis has delayed the onset of harmful hypoxic depolarization, resulting in a decrease in the total hypoxic period and the hypoxic depolarization. This effect has been considered to be protective. However, the synaptic recovery after reoxygenation was attenuated when acidosis (pH: 6.2-6.9) was sustained. Conversely, the synaptic recovery was potentiated when the acidosis was restored to the physiological milieu during the reoxygenation period. These results suggest that acidosis plays a protective effect against the hypoxic neuronal damage only when rapid appreciable pH recovery is achieved during reoxygenation. PMID- 10854588 TI - Co-localization of cart peptide immunoreactivity and nitric oxide synthase activity in rat hypothalamus. AB - Because of the reported presence of both CART peptide and NOS activity in the same hypothalamic nuclei, their colocalization was examined. Eighteen percent of the neurons in the supraoptic nuclei, and 16% of the neurons in the paraventricular nucleus contained both CART immunoreactivity and NOS activity. Many other neurons in these regions stained for only one marker although they were often close by. Thus, CART peptides and NO may interact in these regions. PMID- 10854589 TI - Upregulation of ciliary neurotrophic factor in reactive Muller cells in the rat retina following optic nerve transection. AB - We have investigated the expression and cellular localization of ciliary neurotrophic factor (CNTF) in the rat retina following optic nerve transection. In the normal retina, CNTF immunoreactivity was restricted to profiles in the ganglion cell layer. Following optic nerve transection, immunoreactivity appeared in Muller cell somata and processes and its intensity increased between three and seven days post-lesion. Quantitative evaluation by immunoblotting confirmed that CNTF expression increased continuously up to 7 days after optic nerve transection (to 430% of control levels), but decreased again to 250% of controls at 4 weeks post-lesion. Our findings suggest that CNTF supplied by Muller cells may play a protective role for axotomized ganglion cells in the rat retina. PMID- 10854590 TI - Estrogen effects on tyrosine hydroxylase-immunoreactive cells in the ventral mesencephalon of the female rat: further evidence for the two cell hypothesis of dopamine function. AB - The present study was undertaken to examine the differential effect of estrogen (E) on the expression of tyrosine hydroxylase (TH) in the substantia nigra compacta (SNc) and in two subdivisions of the ventral tegmental area in ovariectomized (ovx) and ovx plus estradiol benzoate (ovx+E)-treated female rats. Cell counting of TH-immunoreactive perikarya of the SNc, paranigral (PN) and interfascicular (IF) nucleus was performed and compared. Our findings demonstrate that E eliminated TH immunoreactivity from a number of midbrain neurons, while it seemingly did not affect it in others. This signifies a differential effect of E on ventral mesencephalic dopaminergic neurons. PMID- 10854591 TI - Leptin facilitates histamine release from the hypothalamus in rats. AB - We studied the effect of leptin on the release of histamine from the anterior hypothalamus using an in vivo microdialysis in anesthetized rats. Histamine release was significantly increased by leptin administration (1.3 mg/kg, i.p.) and kept at a high level for 4 h after the injection. The same dose of leptin significantly reduced food intake in another group of rats. This finding suggests that leptin activates the histaminergic system in the hypothalamus, which may contribute the expression of leptin-induced anorectic effect. PMID- 10854592 TI - Directed sampling for electrolyte analysis and water content of micro-punch samples shows large differences between normal and ischemic rat brain cortex. AB - Changes in sodium, potassium, and water content in brain tissue are important in the progression of pathology that follows ischemic stroke. Determining these parameters regionally in rodent models of experimental ischemia has been limited because typical tissue weights of more than 35 mg are too large. Identifying ischemic tissue to direct tissue sampling towards ischemic cortex is also represents a difficult generally unresolved area. We suggest that larger differences between normal and ischemic cortex of sodium, potassium, and water content than previously observed can be obtained from directed sampling of 2-mg brain tissue in a model of focal cerebral ischemia. In five rats, the middle cerebral artery and both common carotid arteries were occluded for 4.9+/-0.13 h (mean+/-SEM). Punch-sampling of 1-mm diameter tissue cores for water content (H(2)O%) by the wet-dry method, and [Na(+)] and [K(+)] by flame photometry, was guided by the observation of a subtle change in the surface reflectivity of ischemic cortex of quickly dried, 20-microm frozen brain sections, that was confirmed by MAP2 immunohistochemistry. The ratio of the lesion areas as determined by the reflective change and MAP2 immunoreactivity was 0.96+/-0.03 (n=5). In ischemic cortex H(2)O% was 79.9%+/-0.8%, [Na(+)] was 550+/-25 mEq/kg dry-weight, and [K(+)] 94.2+/-19.2 mEq/kg dry-weight (n=5), all significantly different from the values in border zone cortex, and in cortex contralateral to ischemic cortex and border zone (for all samples n=60, mean wet weight 2.037+/ 0.046 mg). Differences between ischemic and normal cortex were 5.4+/-1.1%, 317+/ 21 mEq/kg dry-weight, -304+/-27 mEq/kg dry-weight (n=5) for H(2)O%, [Na(+)], and [K(+)]. These differences between ischemic and normal cortex are 1.4-2.5, 1-3.11, and 1.4-3.5 times greater, respectively, than previous results obtained using samples weighing 35 mg or more. These results extend the association of sodium and potassium with ischemic brain edema in the rodent model, and show that these classical measurements can keep pace with the regionality of histochemical and morphological methods. PMID- 10854593 TI - Effects of sub-chronic combined treatment with pergolide and caffeine on contralateral rotational behavior in unilateral 6-hydroxydopamine-denervated rats. AB - We studied the synergistic effects of pergolide and bromocriptine with caffeine on turning behavior in 6-OHDA denervated rats. Both pergolide and bromocriptine were synergistic with caffeine, and prevented tolerance to caffeine-induced turning. When caffeine was removed, tolerance to bromocriptine effects was observed for 1 day only, while no tolerance was observed to pergolide. These results suggest that caffeine could be useful in the treatment of Parkinson's disease, preferentially as an adjuvant of mixed dopaminergic agonists like pergolide. PMID- 10854594 TI - Lack of correlation between cholinergic-induced changes in chemosensory activity and dopamine release from the cat carotid body in vitro. AB - We studied the effects of nicotine, acetylcholine (ACh) and dopamine (DA) on the frequency of chemosensory discharges (f(x)) and catecholamine (CA) efflux in the cat carotid body superfused in vitro. CA efflux was measured by changes in CA concentration (DeltaCA) determined by chronoamperometry with nafionated carbon fiber microelectrodes inserted in the carotid body, while f(x) was recorded simultaneously from the carotid (sinus) nerve. Nicotine (10-20 microg) and ACh (>100 microg) increased f(x) in all carotid bodies (n=16), but produced a delayed DeltaCA ( approximately 0.65 microM) in only half of them. Eserine potentiated ACh-evoked increases in f(x) and CA effluxes. Nicotine and ACh-induced DeltaCA were rapidly reduced upon repeated administration. While f(x) increases evoked by low doses of nicotine or ACh were reduced or abolished by prior administration of exogenous DA (>100 microg), CA effluxes were enhanced and hastened. Thus, cholinergic-induced changes in f(x) are dissociated from CA efflux. PMID- 10854595 TI - Analysis of EEG data from weak-field magnetic stimulation of mesial temporal lobe epilepsy patients. AB - Ten patients suffering from mesial temporal lobe epilepsy were exposed to weak, DC magnetic field stimulation following computer-controlled protocols established in previous studies. Poisson statistical analysis of the ten patients undergoing semi-invasive (foramen ovale) electrode monitoring reveals that for at least one experimental protocol, application of DC magnetic fields alters interictal epileptiform spike activity in five of ten patients. Similar results also have been observed in the analysis of both human and rat brains by employing weak, alternating magnetic field stimulation. Further study is necessary in order to optimize the magnetic field exposure protocol. PMID- 10854596 TI - Analysis of human self-reactive antibody repertoires by quantitative immunoblotting. AB - We review the use of a quantitative immunoblotting technique to characterize human self-reactive antibody repertoires in health and disease. The interactions of plasma IgM and IgG with tissue extracts as sources of self-antigens were analyzed by quantitative immunoblotting. Data were compared by means of multiparametric statistical analysis. The data summarized here demonstrate that natural self-reactive antibody repertoires of healthy individuals are restricted to a limited subset of immunodominant autoantigens that is selected early in development, and remains conserved between individuals through ageing. The selection of human natural self-reactive IgG antibody repertoires requires normal T-/B-cell interactions. The immunoblotting assay has the potential to distinguish between autoimmune diseases with organ-related oligoclonal expansion of self reactive clones and those characterized by broad alterations of immunoregulation. However, organ-specific autoimmune diseases may be characterized by altered patterns of antibody repertoires unrelated to the target organ. The assay also revealed an unexpected defect in the regulatory function of self-reactive IgM on the expression of self-reactive IgG repertoires in several systemic and organ specific autoimmune diseases. The results are discussed in the light of our current understanding of the processes of selection of self-reactive B-cells and the pathophysiology of autoimmunity. PMID- 10854597 TI - Detection of mRNAs in Peyer's patches of the developing mouse embryo. AB - We describe a method to identify cells expressing mRNA of interest in the developing digestive tract by whole mount in situ hybridization with digoxigenin labeled RNA probes. In preparing samples, serosal tissue surrounding the intestine was removed. Enzymatic reactions and probe concentrations were optimized. Furthermore, polyvinyl alcohol was included in the reaction mixture for the color development of alkaline phosphatase conjugated to the antibody against digoxigenin. These modifications improved the sensitivity and enabled us to identity cells that express mRNA in embryonic intestine. Using the antisense probe for VCAM-1, the protein product of which is an immunohistochemical marker of the Peyer's patch in the embryonic intestine, cells expressing mRNA were identified as spot-like clusters in Peyer's patches, confirming the validity of the method. With this method, mRNAs of both lymphotoxins alpha and beta, key molecules for peripheral lymphoid organ development, were found to be confined to the Peyer's patch in the developing intestine. Whole mount in situ hybridization analysis is a useful tool for exploring spatio-temporal expression profiles of mRNA in the developing immune organs. PMID- 10854598 TI - Digital video-imaging of leukocyte migration in the iris: intravital microscopy in a physiological model during the onset of endotoxin-induced uveitis. AB - The process of inflammation is accompanied by an alteration of leukocyte endothelial dynamics. Reciprocal changes in the endothelium and the white cell permit the leukocyte to relinquish its normal free-flowing state in order to roll, arrest, and emigrate through the endothelium. Although intravital microscopy is an established method to observe this process, the eye has been under-utilized for this purpose. Iris vasculature can be videophotographed without the artifact of trauma. We used rhodamine 6G in vivo staining of leukocytes from BALB/c mice in a model of inflammation induced by intravitreally injected endotoxin. Digital video technology was used to record observations at baseline, 2 h, and 4 h after the endotoxin injection. Off-line analysis of microhemodynamic parameters established that the percentage of venules exhibiting rolling increased significantly from 4% at baseline to 34% at 2 h and 82% at 4 h after endotoxin injection. We found a marked increase in leukocyte arrest within 4 h (601+/-119 cells per mm(2) vs. 2+/-1 cells per mm(2) in control animals). Although shear stress differs minimally between iris arterioles and venules, both rolling and arrest occurred preferentially in venules indicating that shear stress is not the dominant factor for determining cell adhesion. Compared to previous reports on intravital microscopy, our methodology includes refinements or advantages in visualizing cells that have transmigrated as well as the avoidance of surgical trauma. The resolution and quantifiable nature of this technique are such that the methodology can be applied to repetitive observation of leukocyte-endothelial dynamics during an immune response. PMID- 10854599 TI - A new method for histamine release from purified peripheral blood basophils using monoclonal antibody-coated magnetic beads. AB - A new method for evaluating histamine release from purified basophils was developed. Basophil-containing leukocytes were directly purified from a small amount of peripheral blood using monoclonal antibody BA312-coated magnetic beads. The purified basophils still rosetted to magnetic beads maintained a normal response to anti-IgE and to dust mite allergen in comparison with the conventional method using washed leukocytes. This methodology facilitates the purification of basophils, anti-IgE- and allergen-induced histamine release, and subsequent histamine determination within only 3 h. The released histamine was analyzed by an enzyme-linked immunosorbent assay (ELISA) with a characteristic detection profile. Since all steps were performed in 96-well microplates, many clinical samples could be analyzed at the same time, permitting easy applications in routine laboratories. PMID- 10854600 TI - Use of microbial transglutaminase for the enzymatic biotinylation of antibodies. AB - Nowadays many reagents are available for the biotinylation of proteins. As most of them bind to amino groups of the protein the degree of labelling differs from batch to batch and the possibility exists that the biological activity of the target protein may be affected by the labelling procedure. In the present study we have investigated an enzymatic approach to biotinylation using microbial transglutaminase (MTGase) from Streptoverticillium mobaraense. The proposed method is particularly suitable when only a few biotin molecules need to be attached to the target proteins. The enzyme catalyses the acyl transfer reaction between gamma-carboxyamide groups and various primary amines. This was exploited for biotinylation using two amino-modified biotin derivatives, biotinamido-5 pentylamin (BIAPA) and biotinoyl-1,8-diamino-3, 6-dioxaoctane (BIDADOO) as acyl acceptors and a monoclonal IgG against the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) as the acyl donor. Kinetic studies revealed that the MTGase-mediated reaction proceeds with low velocity and is almost complete after 34 h. Conjugation ratios ranging from 1.1 to 1.9 biotins per IgG were found by mass spectrometry. To investigate the influence of antibody conjugation on antigen binding a competitive ELISA for the determination of 2,4-D employing MTGase biotinoylated IgGs was developed. In this assay lower limits of detection of 0.3 and 1.0 microg/l of 2,4-D were achieved with BIDADOO- and BIAPA-modified antibodies, respectively. PMID- 10854601 TI - A method to study apoptosis in eosinophils by flow cytometry. AB - The aim of this study was to develop a simple flow cytometric procedure to study eosinophil apoptosis. Eosinophils were isolated from the peripheral blood of healthy, non-allergic individuals and then cultured in basal culture medium. The cells were examined after 24, 48 and 72 h for forward- and side scatter (FS-SSC) pattern, staining with FDA, PI, and anti-CD95, and light microscopic appearance. After culture for >24 h, two populations with different FS-SSC-patterns appeared, referred to as A and B. Population A consisted of living, FDA-positive eosinophils. The eosinophils in population B showed a lower FS scatter than those in population A and a staining pattern with PI indicating the presence of hypodiploid DNA. Anti-CD95 demonstrated a significant staining of the eosinophils in population B, which increased after 2 days in culture. The cells were sorted using a FACS-Scan cell sorter and by Annexin V-coated magnetic beads to permit separate analyses of PI-staining pattern, DNA electrophoresis, and light microscopic examination of the cells in population B. The present study suggest that it is possible to discriminate between apoptotic and living eosinophils using the FS-SSC pattern and the PI-staining pattern obtained by flow cytometry. PMID- 10854602 TI - Freezing of dendritic cells, generated from cryopreserved leukaphereses, does not influence their ability to induce antigen-specific immune responses or functionally react to maturation stimuli. AB - It is of practical clinical importance to be able to reinfuse into patients dendritic cells which have been previously frozen in aliquots. However, there are few studies comparing the function of fresh and frozen dendritic cells. We therefore decided to perform a systematic immunophenotypic and functional comparison of fresh and frozen dendritic cells. We chose to assess functional properties using proliferation tests and evaluating the preservation of specific antigens presentation in the context of MHC class II. Dendritic cells, generated from leukaphereses of normal volunteers, were loaded with proteins by a 2-h incubation at a protein concentration of 50 microg/ml, and were thereafter used fresh or after freeze-thawing in an IFNgamma Elispot assay. The IFNgamma release from antigen specific T cells was not affected by liquid nitrogen storage of pulsed immature dendritic cells. In the same way, the storage did not alter their stimulatory properties for antigen specific autologous T cells or for allogeneic CD8+ T lymphocytes in a proliferation assay. We also showed that freezing non pulsed immature dendritic cells did not alter their capacity to capture, process and generate antigen-specific reactions once thawed, nor did it impair their capacity to acquire fully mature characteristics using CD40L and IFNgamma, with respect to immunophenotype and bioactive IL-12 secretion. PMID- 10854603 TI - Measurement of the ADP:ATP ratio in human leukaemic cell lines can be used as an indicator of cell viability, necrosis and apoptosis. AB - In this study the relative levels of ADP and ATP have been measured in cells undergoing apoptosis. Using HL60, CEM7, Jurkat and U937 cell lines and cytotoxic agents known to induce apoptosis, there was a significant correlation (P<0.01 for all models) between the ADP:ATP ratio and the degree of apoptosis measured by TUNEL and estimation of the sub G(0) fraction by propidium iodide staining and flow cytometry. The ratio measured in viable proliferating cells was found to be less than 0.11 compared with ratios between 0.11 and 1.0 seen in cells undergoing apoptosis. The higher the percentage of hypodiploidy the greater the ratio. Necrosis induced by heat shock resulted in ADP:ATP ratios in excess of 15.0. When primary cultures of AML blast cells were used, there was again a significant correlation between the ADP:ATP ratio and the degree of hypodiploidy. Recent evidence suggests that apoptosis is accompanied by opening of the mitochondrial permeability pores, leading to disruption of the mitochondrial transmembrane potential (DeltaPsi(m)). This results in caspase activation due to the release of cytochrome c and apoptogenic factors into the cytosol. In five experiments using CEM7 and dexamethasone the mitochondrial transmembrane potential was assessed using the fluorescent cyanine dye JC-1 and flow cytometry. Functioning mitochondria concentrate the JC-1 to produce red fluorescence. Loss of mitochondrial transmembrane potential results in green fluorescence only. The percentage of cells exhibiting red fluorescence correlated positively with the ATP values and negatively with the ADP:ATP ratio. PMID- 10854604 TI - Fractionation of differentiating cells using density perturbation. AB - This paper describes the development of a new method for the fractionation of purified subpopulations of partially differentiated cells on continuous isopycnic gradients, using a density perturbation method based on the ability of cells to bind dense antibody-coated beads. Until now none of the available fractionation techniques, such as magnetic cell fractionation has been efficient for separating subpopulations of partially differentiated cells. The fractionation experiments described in this report used promyelocytic HL-60 and DMSO-induced granulocytic HL-60 cells as a model system. Populations of cells, modified by the binding of dense beads were fractionated on isotonic, isopycnic Optiprep gradients by centrifugation at 220xg for 90 min at 20 degrees C. Examination of the different gradient fractions showed that, as cells bind increasing numbers of beads, they are found in the denser regions of the isopycnic gradients. Indirect immunofluorescence was combined with flow cytometric techniques to characterise the fractionation of partially differentiated cells. Flow cytometric results confirmed that as antigenic determinants appear on the surface at higher levels of expression, the number of beads binding to each cell increased. Furthermore, after fractionation, when the bead-bound and non-bead-bound cells were cultured in the presence of DMSO, those cells that had bound more beads targeted to differentiated cells were found to achieve terminal differentiation faster than those cells that had not been associated with any beads. PMID- 10854605 TI - An improved procedure for the development of human mast cells from dispersed fetal liver cells in serum-free culture medium. AB - The in vitro development of human mast cells from fetal liver cells with recombinant human stem cell factor in serum-containing RPMI was compared to that in AIM-V media with and without serum. Compared to serum-containing media, AIM-V medium caused mast cells to develop earlier and in greater numbers. By 2 weeks, about 60% of cells in serum-free AIM-V medium were phenotypic mast cells, approximately 2 times the percentages in serum-containing media. By 6 weeks the percentages of mast cells were > or =80% under all conditions, but the number of mast cells was 3-4-fold greater in serum-free AIM-V medium than in serum supplemented media. Mast cells obtained in serum-free AIM-V medium exhibited rounded nuclei, like tissue-derived mast cells; mast cells obtained in serum supplemented media had segmented nuclei. By 10-12 weeks of culture about 40% of the AIM-V-derived cells showed strong chymase immunocytochemical staining, a pattern observed for only 14% of the cells in serum-containing media. AIM-V medium is a suitable medium for the development of human mast cells in vitro, and permits an earlier, more selective and greater expansion of mast cells than serum containing media. PMID- 10854606 TI - Production and characterisation of monoclonal antibodies against a very small hapten, 3-methylindole. AB - Monoclonal antibodies were produced against a very small (131.2 Da) hapten, 3 methylindole. Nine derivatives of 3-methylindole were synthesised with spacers ending in a carboxyl group, and coupled to immunogenic carriers and europium chelate labels. Almost all the antigens elicited an antihapten response, but the majority of the mAbs produced strongly recognised the spacer group and did not bind free 3-methylindole. However, specific antibodies were obtained with five immunogens. Specificity could be directed against the pyrrole ring by locating the bridging group to the aromatic moiety of the indole ring system. Any modification in the position 3 of the indole ring strongly hindered mAb binding to the compound, and the cross-reactivity of physiologically important compounds, such as tryptophan and tryptamine, was negligible for all of the mAbs. The developed hapten structures successfully focused antibody recognition to the important sub-determinants in the indole ring system. Similar constructs could also be useful in the development of antibodies against other indolic compounds. PMID- 10854607 TI - Human granulocyte CD11b expression as a pharmacodynamic biomarker of inflammation. AB - A method has been developed for the direct quantification of the CD11b integrin on granulocytes by flow cytometric analysis of whole blood specimens following either LTB(4) or lipopolysaccharide (LPS) stimulation. This method has utility in evaluating the pharmacodynamic action of either LTB(4) receptor antagonists or immune cell modulators in effecting CD11b integrin expression and granulocyte activation in human subjects administered such drugs. Previous studies using CD11b as a biomarker of granulocyte activation have faltered because of the difficulty in controlling the activation state of the granulocyte following removal of blood from subjects. The present study has made use of a newly validated method using either LTB(4) or LPS to stimulate CD11b expression on granulocytes and has been used, as one measure, in the evaluation of LPS activity when administered to normal human volunteers. PMID- 10854608 TI - A quantitative enzyme-linked immunoassay for the detection of 2, 6 dichlorobenzamide (BAM), a degradation product of the herbicide dichlobenil. AB - 2,6-Dichlorobenzamide (BAM) is the dominant degradation product in soil of the widely used herbicide dichlobenil. To detect BAM in water, a highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) was developed. As an alternative to conventional coating of ELISA plates, the assay is based on direct covalent immobilisation. We achieved a surface which requires a short time for the immobilisation of ligand, is stable under dry storage, and which permits assays with a low CV. The performance of the assay was demonstrated by an inter well CV that was generally less than 6%, a detection limit (DL(15)) of 0.02 microg/l and an IC(50) of 0.19 microg/l. Cross-reactivity was measured against nine analytes with structural homology to BAM. The highest degree of cross reactivity (10.8%) was seen with 2,6-dichlorothiobenzamide (Chlorthiamid). Considering an EU-limit of 0.1 microg/l as the permissible maximum for the presence of pesticides in drinking water, this ELISA-procedure is suitable for large-scale screening of water samples suspected of being contaminated with BAM. PMID- 10854609 TI - Granzyme B ELISPOT assay for ex vivo measurements of T cell immunity. AB - A major goal in immunodiagnostics has been the development of assay systems that can measure CD8(+) T cell immunity in humans, directly ex vivo, at high resolution, and with high throughput. We established granzyme B (grB) enzyme linked immunospot assay (ELISPOT) in conjunction with image analysis to this end. Using grB transfected and untransfected Chinese hamster ovary (CHO) cells and T cell lines, we show that the antibody pair utilized was grB-specific and that only activated T cells secrete grB. GrB release began within 4 h after antigen stimulation and stopped within 40 h. Side-by-side comparison showed grB ELISPOT assays to have a higher resolution than classic chromium-release assays in terms of signal-to-noise ratio. The linearity of the relation of the number of CD8(+) effector T cells plated to grB spots detected suggests that grB ELISPOT assays measure the frequencies of grB-secreting cells directly. Reactivity to HIV peptides was seen in grB ELISPOT assays of freshly isolated PBMC from HIV patients, consistent with the detection of peptide-specific memory cells. The higher resolution and lower labor and material investment should make grB ELISPOT assays an attractive alternative to chromium-release assays in monitoring the clonal sizes of specific CD8 memory cells in vivo. PMID- 10854610 TI - A surrogate method for assessment of beta(2)-integrin-dependent adhesion of human eosinophils to ICAM-1. AB - We have developed and validated an inexpensive and equivalent method for measuring eosinophil adhesion by beta(2)-integrin to endothelial ICAM-1 using bovine serum albumin (BSA) as a surrogate for the immunoglobulin supergene. The number of adherent eosinophils on BSA or ICAM-1 coated microplates was quantified by residual eosinophil peroxidase activity. Non-stimulated eosinophils did not adhere to either BSA or ICAM-1. However, after IL-5 stimulation, either BSA or ICAM-1 caused comparable and concentration-dependent adhesion of eosinophils. Eosinophil adhesion was rapid and occurred within 15 to 30 min of incubation for either BSA or ICAM-1. Preincubation of cells with CD11b or CD18 antibody specifically decreased adhesion to either BSA or ICAM-1. IL-5, PAF and fMLP all induced adhesion of eosinophils to either BSA or ICAM-1 in a concentration dependent manner, and the optimal IL-5, fMLP and PAF concentrations for adhesion to BSA were the same as for adhesion to ICAM-1. BSA-binding was specific for beta(2)-integrin; neither alpha-CD49d mAb directed against the alpha(4)-chain or alpha-CD29 directed against the common beta(1)-chain of VLA-4 blocked adhesion to BSA or ICAM-1 controls. The protein tyrosine kinase inhibitor, genistein, the phosphatidylinositol 3-kinase (PI-3 kinase) inhibitor, wortmanin, and mitogen activated protein kinase kinase (MEK) inhibitor, U0126, all inhibited IL-5 induced eosinophil adhesion to either BSA or ICAM-1 comparably. These results indicate that BSA is a reliable and economical surrogate ligand for ICAM-1 adhesion to beta(2)-integrin-dependent adhesion to ICAM-1. Ligation characteristics of BSA are identical to those for soluble ICAM-1, and the assay is suitable for assessment of signal transduction pathways mediating adhesion. PMID- 10854611 TI - A new peptide-affinity tag for the detection and affinity purification of recombinant proteins with a monoclonal antibody. AB - A monoclonal anti-peptide antibody (2E11) was raised against the synthetic peptide 38 (C-L-D-K-S-G-L-P-S-D-R-F-F-A) representing a part of the variable region of the Vbeta 6.2 T-cell receptor. This mAb (IgG(1), kappa light chain) bound very specifically to peptide 38 as shown by ELISA but did not recognize the corresponding native Vbeta 6.2 T-cell receptor on T-cells. For epitope analysis, overlapping peptides of 4-10 amino acids in length corresponding to the sequence of peptide 38 were synthesized and assayed by SPOT synthesis on cellulose sheets. The shortest peptide recognized was L-P-S-D-R. The specificity of mAb 2E11 was examined with 100 different peptides comprising other parts of the different variable Vbeta domains of the human T-cell receptor that do not include the epitope region L-P-S-D-R. None of these peptides were recognized. The chemical synthesis of a peptide with the sequence L-P-S-D-R on Sepharose beads allowed to efficiently purify the mAb 2E11 in a single step by affinity chromatography. An equilibrium binding constant of 4.9x10(6) l/mol was determined for mAb 2E11 by using rhodamine-green-labelled peptide 38 in fluorescence correlation spectroscopy. In order to demonstrate that peptide 38 can be used as an affinity tag, it was fused to the carboxyl-terminus of interferon regulatory factor-1 (IRF 1). It could be shown that in vitro translated peptide 38 tagged IRF-1 was immunoprecipitated by the mAb 2E11 and that the fusion protein could be purified by immunoaffinity chromatography. Additionally peptide 38 was fused to the amino terminus of the Taq polymerase. This recombinant protein was expressed in E. coli and specifically detected in a Dot blot and Western blot using mAb 2E11. PMID- 10854612 TI - Induction of immune responses and molecular cloning of the heavy chain antibody repertoire of Lama glama. AB - Functional heavy chain immunoglobulins have, so far, only been found in camels and llamas. Antigen-specific fragments of these heavy chain IgGs (V(HH)) are of great interest in biotechnology because they are very stable and can be produced at high level by the yeast Saccharomyces cerevisiae. The work described in this paper was conducted to determine whether llamas (Lama glama) are a practical source of antigen-specific V(HH) fragments. Llamas were immunised with various types of antigens and the antibody responses were examined during the course of immunisation. Both, conventional and heavy chain IgG antibodies were produced in response to each of the antigens. The heavy chain IgG repertoire displayed a recognition pattern different to that of conventional llama IgGs, resulting in the expansion of the accessible epitope repertoire. Llamas have a lower proportion of heavy chain IgG antibodies in their serum than have camels. To enable the specific and efficient isolation of V(HH) genes from peripheral blood B-cells, the long and short-hinge sequences of Lama glama heavy chain IgGs were determined, revealing the presence of a novel subclass of short-hinge heavy chain IgG. Long and short-hinge specific PCR primers were designed to be used in the construction of llama V(HH) libraries. We conclude that, using the techniques described, antigen-specific V(HH) antibody fragments are readily accessible from the llama, thus providing highly valuable binding molecules for a variety of applications. PMID- 10854613 TI - Dynamics of prey moving through a predator field: a model of migrating juvenile salmon. AB - The migration of a patch of prey through a field of relatively stationary predators is a situation that occurs frequently in nature. Making quantitative predictions concerning such phenomena may be difficult, however, because factors such as the number of the prey in the patch, the spatial length and velocity of the patch, and the feeding rate and satiation of the predators all interact in a complex way. However, such problems are of great practical importance in many management situations; e.g., calculating the mortality of juvenile salmon (smolts) swimming down a river or reservoir containing many predators. Salmon smolts often move downstream in patches short compared with the length of the reservoir. To take into account the spatial dependence of the interaction, we used a spatially-explicit, individual-based modeling approach. We found that the mortality of prey depends strongly on the number of prey in the patch, the downstream velocity of prey in the patch, and the dispersion or spread of the patch in size through time. Some counterintuitive phenomena are predicted, such as predators downstream capturing more prey per predator than those upstream, even though the number of prey may be greatly depleted by the time the prey patch reaches the downstream predators. Individual-based models may be necessary for complex spatial situations, such as salmonid migration, where processes such as schooling occur at fine scales and affect system predictions. We compare some results to predictions from other salmonid models. PMID- 10854614 TI - Early HIV infection in vivo: branching-process model for studying timing of immune responses and drug therapy. AB - We propose a stochastic, branching-process model of early events in vivo in human or simian immunodeficiency virus (HIV or SIV) infection and study the influence that the time of appearance of virus-specific antibodies or cytotoxic cells, or of administration of antiretroviral drugs, has on the probability of progression to a chronic infection. In some biological scenarios, our model predicts that a few days' delay in response or intervention would make little difference, while in others it would be highly deleterious. We show that prophylactic efficacy does not require perfect efficiency at neutralizing infectious virus. Data from a trial of PMPA, a potent antiretroviral drug, as post-exposure therapy for SIV infection in macaques, reported by C.-C. Tsai, P. Emau, K.E. Follis, T.W. Beck, R. E. Beneveniste, N. Bischofberger, J.D. Lifson, W.R. Morton (J. Virol. 72 (1998) 4265), provides a test of the model. We show that their observations are consistent with a branching-process without invoking supplementary viral- or host variability. Finally, most animal trials of antiviral drugs or vaccines use very high viral inoculums; our model demonstrates that in such experiments we risk greatly underestimating the efficacy of these agents. PMID- 10854615 TI - Stochastic models for aggregation processes. AB - Three models are presented, which describe the aggregation of objects into groups and the distributions of groups sizes and group numbers within habitats. The processes regarded are pure accumulation processes which involve only formation and invasion of groups. Invasion represents the special case of fusion when only single objects - and not groups - join a group of certain size. The basic model is derived by a single parameter, the formation probability q, which represents the probability of an object to form a new group. A novel, discrete and finite distribution that results for the group sizes is deduced from this aggregation process and it is shown that it converges to a geometric distribution if the number of objects tends to infinity. Two extensions of this model, which both converge to the Waring distribution, are added: the model can be extended either with a beta distributed formation probability or with the assumption that the invasion probability depends on the group size. Relationships between the limiting distributions involved are discussed. PMID- 10854616 TI - A probabilistic approach to the assessment of some life history pattern parameters in a Middle Pleistocene human population. AB - Parameters of a Middle Pleistocene human population such as the expected length of the female reproductive period (E(Y)), the expected interbirth interval (E(X)), the survival rate (tau) for females after the expected reproductive period, the rate (phi(2)) of women who, given that they reach first birth, do not survive to the end of the expected reproductive period, and the female infant plus juvenile mortality rate (phi(1)) have been assessed from a probabilistic standpoint provided that such a population were stationary. The hominid sample studied, the Sima de los Huesos (SH) cave site, Sierra de Atapuerca (Spain), is the most exhaustive human fossil sample currently available. Results suggest that the Atapuerca (SH) sample can derive from a stationary population. Further, in the case that the expected reproductive period ends between 37 and 40 yr of age, then 24 less, similarE(Y) less, similar27 yr, E(X)=3 yr, 0.224RC at high VL and RC>Ab at low VL, another the opposite pattern; the third was in between. We conclude that while different flautists use different strategies to control Pm, the results are similar. Independent control of V and Vel by Pm and Aem allow flautists to control I and F regardless of how Pm is generated. PMID- 10854622 TI - Serotonin reuptake inhibition does not enhance short term modulation of the exercise ventilatory response. AB - Increased respiratory dead space causes a serotonin (5-HT) dependent augmentation of the exercise ventilatory response known as short term modulation (STM). Contrary to predictions, 5-HT reuptake inhibition with fluoxetine failed to enhance, and even impaired STM with large dead space volumes (0.4-0.6 L). In this study, we tested the hypotheses that: (1) fluoxetine similarly impairs STM with smaller dead space volumes (0.2 L); whereas (2) acute 5-HT release and reuptake inhibition with fenfluramine would enhance STM. Ventilatory and blood gas measurements were made on five goats (37-58 kg) during rest and exercise, with the mask alone or with increased dead space (0.2 L). STM protocols were performed following chronic fluoxetine (>/=21 days, 1 mg/kg, SQ, SID) and acute fenfluramine administration (1 mg/kg, IV). Following fluoxetine, STM was partially impaired. Fenfluramine had no detectable effects on STM. The data suggest that: (1) chronic fluoxetine diminishes STM, possibly via down-regulation of relevant 5-HT receptors, and (2) drugs that release 5-HT acutely do not enhance STM. PMID- 10854623 TI - Apparent diffusion limitations for CO(2) excretion in rainbow trout are relieved by injections of carbonic anhydrase. AB - Experiments were performed in vivo to elucidate the underlying mechanism(s) of apparent diffusion limitations for CO(2) excretion in rainbow trout (Oncorhynchus mykiss). Ligation of two gill arches and the associated expected reduction in gill surface area of 30% caused pronounced respiratory acidosis as indicated by elevated arterial blood P(CO(2)) (Pa(CO(2))) and reduced arterial blood pH. Under conditions of normoxia, arterial blood P(O(2)) (Pa(O(2))) was not significantly (statistically) reduced. However, during hypoxia (water P(O(2))=70-80 mmHg), the apparent trend for reduced Pa(O(2)) values became statistically significant in fish with 15% surface area reduction. To determine whether the elevated Pa(CO(2)) in fish with reduced surface area (30%) reflected true diffusion limitations or chemical equilibrium limitations imposed by the relatively slow rate of red blood cell Cl(-)/HCO(3)(-) exchange, fish were injected with carbonic anhydrase (CA) to permit catalysis of HCO(3)(-) dehydration within the plasma. Injection of CA caused a lowering of Pa(CO(2)) by 0.87+/-0.32 mmHg within 120 min and thus essentially eliminated the increase in Pa(CO(2)) (1.04+/-0.33 mmHg) that was caused by the reduction in surface area. These results clearly demonstrate that the elevation in Pa(CO(2)) evoked by gill surface area reduction is a consequence of chemical equilibrium limitations rather than true diffusion limitations, per se. PMID- 10854624 TI - Hypercapnia-induced contraction in isolated pulmonary arteries is endothelium dependent. AB - It has been demonstrated previously that isohydric hypercapnia (IH) does not affect agonist-induced tension development in pulmonary arteries. The aim of the present study was to examine the effects of IH on depolarisation-induced, steady state tension in the isolated rat pulmonary artery. Rings were submaximally contracted with high KCl under control conditions (5% CO(2)-95% air). IH was achieved by switching to a modified PSS (isosmotic substitution of NaHCO(3) for NaCl), equilibrated with 10% CO(2) in air. On switching to IH, a significant increase in mean (+/-SEM) tension (25.3+/-6.3% Tmax) was observed in endothelium intact rings (n=6). Endothelial removal significantly reduced this response. Non specific inhibition of nitric oxide synthase (NOS) isoenzymes (L-NAME, 10(-3) M) abolished the IH-induced increase in tension while inhibition of neuronal NOS (TRIM, 10(-5) M) was without effect. The relaxant response to the nitric oxide donor sodium nitroprusside was similar in IH and control conditions. These results suggest that IH caused an endothelium-dependent increase in depolarisation-induced tension by reducing NO production. PMID- 10854625 TI - Gq protein level increases concurrently with antigen-induced airway hyperresponsiveness in rats. AB - In the present study, bronchial Gq protein level of the airway hyperresponsive rats was determined by using immunoblot analysis. In the airway hyperresponsive rats that were sensitized and repeatedly antigen challenged, the in vitro bronchial responsiveness to acetylcholine was significantly enhanced as compared with that in the sensitized control group. Moreover, the bronchial contraction induced by 10 microM AlF(4)(-) (generated by 10 microM AlCl(3) plus 10 mM NaF) was significantly elevated after repeated antigen challenge (0.44+/-0.13 and 1.09+/-0.09 g tension in the control and airway hyperresponsive groups, respectively; P<0.01). In both groups, immunoblotting with the antibody against G alpha q gave a single 42 kD band. The G alpha q protein levels in the airway hyperresponsive group (0.58+/-0.12) estimated by G alpha q/beta-actin ratio was significantly greater than those in the control group (0.30+/-0.10; P<0.05). These findings suggest that the increase in G alpha q protein level may be involved in the pathogenesis of antigen-induced airway hyperresponsiveness in rats. PMID- 10854626 TI - Distinct categories of immunologic changes in frail elderly. AB - Immune changes and their relationships in a frail elderly population (N=116, age 70-103, median 86 years) were defined in comparison to a healthy younger group. Previous immune studies in the elderly have generally focused on one or few parameters without correlation analyses. Furthermore, the study populations have been active elderly in relatively small numbers. A total of 33 immune parameters representing many aspects of the immune system were quantified. Most changes in the frail elderly were parallel to those reported in active elderly. A classification tree analysis revealed that increased plasma activation markers (neopterin and sTNF-R) and increased CD28 expression on CD8 T cells and proliferative response separated the aged and control populations. Statistical procedures utilizing principal components analyses, partial correlations and exploratory factor analyses all indicated that immunologic parameters in frail elderly are grouped in three major clusters of immunologic results. These involved (a) increased plasma levels of neopterin and sTNF receptor indicating elevated IFNgamma and TNF cytokine activity; (b) increased proportion of mature (CD45RO) versus naive (CD45RA) T cells; and (c) a diverse group of related changes including impaired proliferative response, reduced T cells, CD28 and CD25 expression, B cell percentage and lower CD4:CD8 ratios and increased HLA-DR expression. These findings emphasize that several different groups of immune parameters but not 33 independent immune changes, occurred in the aged population. PMID- 10854627 TI - Articular chondrocytes from aging rats respond poorly to insulin-like growth factor-1: an altered signaling pathway. AB - This study investigates the effect of insulin-like growth factor-1 (IGF-1) and phorbol 12-myrystate 13-acetate (PMA) on 3H-thymidine, 35SO(4) and 3H -glycine incorporations, adenosine 3':5'-cyclic monophosphate (cAMP) production and protein kinase C (PKC) activation in cultured rat articular chondrocyte monolayers (RACM) derived from animals of different ages. It was found that IGF-1 stimulates all these cellular functions in cultures derived from all age groups in a concentration dependent manner, although the cells from 14-month old animals responded poorly. IGF-1 also induces in cells from 1-month old rats an increase in the expression of mRNAs specific for aggrecan and type II collagen molecules as shown with RT-PCR. These effects are mediated via IGF-1 interaction with specific receptors because the monoclonal antibody against the receptor protein suppresses more than 60% of the ligand-induced DNA synthesis. PMA, a direct PKC activator, potentiated IGF-1-induced effects in all cells but much more strongly in cells from young than in cells from 14-month old animals. The age-related failure of RACM to respond adequately to IGF-1 was correlated with a decrease in IGF-1-induced cAMP production, and IGF-1-induced and PMA-induced PKC activations. These results show that IGF-1 regulates the synthesis of DNA, proteoglycans (PG) and collagen II at the level of transcription and suggest that the reduced response of cell monolayers derived from 14-month old rats to IGF-1 is probably due to a failure of old cells to adequately transduce IGF-1 receptor-generated downstream signaling. PMID- 10854628 TI - Electrophysiological analysis of NMDA receptor subunit changes in the aging mouse cortex. AB - NMDA receptors play an important role in memory processes and plasticity in the brain. We have previously demonstrated a significant decrease in NMDARepsilon2 subunit mRNA and protein with increasing age in the C57Bl/6 mouse frontal cortex. In the present study, two-electrode voltage clamp electrophysiology on Xenopus oocytes injected with total RNA harvested from the frontal cortex of young and old C57Bl mice was used to detect changes in receptor composition during aging. Ifenprodil concentration-response curves, magnesium current-voltage curves, and single channel conductances were determined for native receptors. In addition, ifenprodil and magnesium curves were generated for recombinant NMDA receptors of varying subunit ratios. Ifenprodil dose-response curves for all receptors were biphasic. The low affinity component of the curve increased slightly with age, while the high affinity population decreased, mimicking recombinant receptors with decreasing levels of epsilon2. A decrease in maximal current was also observed in aged animals with decreased levels of epsilon2, although single channel conductances were identical between young and old mice. In addition, an increase in sensitivity to magnesium was observed for receptors from older animals. Results are consistent with the interpretation that the epsilon2 subunit is reduced in older mouse frontal cortex. A change in NMDA receptor subunit composition could influence memory processes during aging. PMID- 10854629 TI - Influence of short-term repeated fasting on the longevity of female (NZB x NZW)F1 mice. AB - Caloric restriction in rodents is well known to retard the rate of aging, increase mean and maximum life-spans, and inhibit the occurrence of many age associated diseases. However, little is known about the influence of short-term repeated fasting on longevity. In this study, female (NZB x NZW)F1 mice were used to test the physiological effect of short-term repeated fasting (4 consecutive days, every 2 weeks). The results showed that fasting mice survived significantly longer than the full-fed mice, in spite of the fasting group having a heavier body weight than the control group. Mean survival times for fasting and control mice were 64.0+/-15.3 and 47.9+/-9.4 weeks, respectively. Short-term repeated fasting manipulation was also effective on the prolongation of life-span in autoimmune-prone mice. PMID- 10854630 TI - Kinetic studies of aflatoxin B1-glutathione conjugate formation in liver and kidneys of adult and weanling rats. AB - Aflatoxin B1(AFB1)-glutathione(GSH) conjugation is the major pathway for the detoxification of aflatoxin metabolites. This reaction is catalysed by GSH S transferase (GST) and play a major role in modulation of AFB1 adduct formation to nuclear DNA. Changes recorded in hepatic GST activity during development of rats can alter the balance between AFB1-GSH conjugation and AFB1-DNA adduct formation. Measurment of cytosolic GST using 1-chloro-2,4-dinitrobenzene (CDNB) as the substrate showed that the enzyme activity is initially lower in weanling tissues as compared to that of adults. But nevertheless hepatic and renal cytosolic GST activity is increased significantly in growing rats pretreated with AFB1. Kinetic studies of AFB1-GSH conjugate formation in kidneys and livers of the two-age groups of rats treated with a single i.p. dose of AFB1 (400 microg/kg b.w.) revealed that at the end of 24 h of AFB1 administration the rate of the conjugate formation in kidneys of immature rats was approximately twice of that measured in adults. Age-related differences in the GST activity as well as AFB1-GSH conjugation was more pronounced in kidneys. The conjugate formation in kidneys of growing rats during 6-24 h following AFB1 administration shows that urinary excretion of aflatoxin metabolites is relatively rapid in growing rats. The major portion of the AFB1-GSH is formed in liver but contribution of the renal tissue to the formation of detoxification metabolites can not be ruled out. These data demonstrate that aflatoxin metabolites are eliminated more efficiently from kidneys of a growing rat. AFB1-induced GST induction in renal tissues of growing animals during 24 h of the carcinogen administration could be considered as an important mechanism for GSH conjugate formation and aflatoxin detoxification. Therefore GST induction in response to hepatotoxic drugs can confer resistance to young animals being exposed for the first time to such drugs. It is also worthmentioning that the GST activity measured before AFB1 administration does not reflect the rate of AFB1 detoxification via GSH conjugation. PMID- 10854631 TI - Immunohistochemical analysis of ageing human B and T cell populations reveals an age-related decline of CD8 T cells in spleen but not gut-associated lymphoid tissue (GALT). AB - It is thought that senescence of the immune system is responsible, at least in part, for many health problems associated with ageing. Previous studies on changes in lymphocyte composition have used flow cytometry to study peripheral blood lymphocytes (PBL's), or cells isolated from rodent tissue, and have yielded conflicting results. We have used immunohistochemistry to determine whether the B and T cells in human tissue from spleen and gut are affected by age. Areas of germinal centre, mantle zone and marginal zone of B cell follicles were measured. In addition, CD4 and CD8 T cells in T cell areas and in B cell follicles were counted. We observed a striking age-related decrease in the proportion of CD8+ T cells in the T cell zones of the spleen. This decrease was not apparent in the T cell population that occupies splenic B cell areas, or in GALT. Further differences, in CD4+ cells, were seen between T cell populations in the T cell zones and those in B cell areas. These findings highlight differences between lymphocyte populations in different lymphoid tissues, and different compartments within each tissue, which may be of importance in future studies of the ageing immune system. PMID- 10854632 TI - Serum leptin response to endogenous hyperinsulinemia in aging rats. AB - To determine if aging is associated with altered serum leptin response to diet induced changes in endogenous hyperinsulinemia, male Fisher 344 (F344) rats at different age groups were studied while on regular rat chow and following 10 days of experimental diets consisting of 60% of the weight as fructose or glucose. The serum leptin concentration (ng/ml) gradually increased from basal levels of 2.5+/ 0.1 at age of 4 months to 3.7+/-0.1, 6.9+/-0.9, 9. 4+/-0.3 and 8.9+/-1.1 at 6, 12, 18 and 24 months of age, respectively (P<0.001). Hyperinsulinemia associated with 60% fructose diet was associated with increased serum leptin levels in 4, 12, and 24 month old rats to 5.1+/-0.8, 6.7+/-1.2, and 8.6+/-1.1, respectively (P<0.001). Feeding 60% glucose diet also was associated with increased serum leptin levels in 4, 12 and 24 month old rats to 7.6+/-0.6, 7.2+/-0.7, and 9.1+/ 1.1, respectively (P<0.001). Restricting dietary intake to 60% of the calories consumed by control rats for 10 days resulted in a decrease in serum leptin to 1.0+/-0.02 in 4 month old rats and 2.5+/-0.4 in 24 month old rats (P<0.01). It is concluded that aging in F344 rats is associated with increased serum leptin concentrations. However, diet-related hyperinsulinemic effect on leptin is blunted in aging rats although leptin response to caloric restriction is maintained. The inability of aging rats to mount hyperleptinemic response to dietary changes may contribute to the age-related increase in adiposity. PMID- 10854633 TI - Whole-body metabolic rate appears to determine the rate of DNA oxidative damage and glycation involved in aging. AB - While aging has been found to be a multifactorial process, it seems logical that different aging parameters which reflect the deleterious effects of normal basal metabolism should be directly related. Three such putative aging parameters were therefore measured in adult male Fischer 344 rats on three different long-term diets which have been shown to yield different lifespans. It was found that the daily caloric intake per unit organ weight, a measure of whole-body metabolic rate, was directly proportional to: (1) the level of 8-hydroxydeoxyguanosine in skin dermal cells, used as a measure of the rate of DNA oxidative damage; (2) the proportion of hemoglobin that was glycated, used as a measure of the rate of glycation. This appears to be the first evidence suggesting that whole-body metabolic rate plays a role in determining both the rate of DNA oxidative damage and the rate of glycation involved in aging, because whole-body metabolic rate was the only one of these three variables manipulated in the study. The study also found that there were no significant between-group differences in brain, kidney and liver 8-hydroxydeoxyguanosine, suggesting that DNA oxidative damage in non-mitotic and slow-dividing cells is not a reliable linear biomarker of aging. PMID- 10854634 TI - Effects of antidepressant treatments on polymorphonuclear elastase levels in patients with depression. AB - BACKGROUND: We have previously reported that severe depression is associated with immunological and inflammatory alterations and these alterations may be showed easily by polymorphonuclear elastase (PMNE) measurements. The purpose of the present study is to show how PMN elastase levels change before and after antidepressant treatment. METHODS: Fifty-five patients with depression (40 with major depression [MD], 15 with dysthymic disorder [DD]) were included in the study. Blood samples were drawn prior to drug treatment, and 3 months after the treatment. Severity of depression was measured by 24-item Hamilton depression rating scale (HDRS). RESULTS: There was a positive correlation between Delta PMNE levels and Delta HDRS in patients with MD, but not in patients with DD. Twenty eight patients were given moclobemide, and 27 patients were given imipramine. It was seen that PMN elastase levels were significantly reduced after 3-month antidepressant treatment period only in patients with MD. CONCLUSION: These findings suggest that PMNE activity is a state dependent parameter and improvement of depressive symptoms due to antidepressant treatment may lead to decrement of PMNE levels. CLINICAL IMPLICATION AND LIMITATIONS: PMN elastase measurements may be used as a sensitive biological marker to follow the time course of the disease activity in patients with major depression. PMID- 10854635 TI - Evaluating the parent-of-origin effect in bipolar affective disorder. Is a more penetrant subtype transmitted paternally? AB - BACKGROUND: Numerous genetic mechanisms and modes of transmission underlying bipolar affective disorder (BPAD) have been postulated. Recently, the discovery of genomic imprinting and mitochondrial transmission of illness in humans has stimulated study of parent-of-origin effects in the transmission of BPAD. METHODS: We examined a large sample of families from an associated linkage study to search for a possible parent-of-origin effect. Selecting for unilineal families with at least one offspring and/or parent diagnosed with BPAD after structured interview, we conducted three analyses: (1) the rates of illness among mothers and fathers of offspring affected with BPAD; (2) the observed frequency of transmission and rates of illness among maternal and paternal lineages; and (3) the rates of affective illness among offspring of parents affected with BPAD. RESULTS: Our results indicate no significant differences in the rates of illness among mothers and fathers of offspring affected with BPAD. Also, the frequency of transmission and rates of illness among maternal and paternal lineages did not differ significantly. However, the rate of BPAD among the offspring of fathers affected with BPAD was significantly higher than for mothers with the illness. LIMITATIONS: Substantially more women than men, and maternal than paternal relatives were studied - introducing possible gender biases. CONCLUSIONS: These results suggest a possible paternal parent-of-origin effect. PMID- 10854636 TI - Nonverbal interpersonal attunement and extravert personality predict outcome of light treatment in seasonal affective disorder. AB - We investigated whether personality and nonverbal interpersonal processes can predict the subsequent response to light treatment in seasonal affective disorder (SAD) patients. In 60 SAD patients, Neuroticism and Extraversion were assessed prior to light treatment (4 days with 30 min of 10.000 lux). From videotaped clinical interviews, the nonverbal interpersonal attunement (i.e. equalizing durations and frequencies of elements of behaviour between conversation partners) was registered for the patients' support seeking and the interviewers' support giving behaviour. The higher Extraversion and the more the patients and the interviewers got attuned over the interview, the more favourable the outcome of light treatment was. Hence, personality and nonverbal interpersonal processes may be involved in the response to light treatment in SAD. PMID- 10854637 TI - The relationship between quality of interpersonal relationships and major depressive disorder: findings from the National Comorbidity Survey. AB - BACKGROUND: The current study compared the quality of interpersonal relationships in individuals with major depressive disorder to individuals with dysthymia, comorbid depression, nonaffective disorders, and no psychiatric disorders. METHODS: Using data from the National Comorbidity Study, a series of logistic regressions, controlling for demographic variables, were conducted to examine the strength of the association between a major depressive disorder and interpersonal dysfunction (positive and negative interactions) in contrast to other psychiatric disorders. RESULTS: Respondents with current major depressive disorder reported significantly fewer positive interactions and more negative interactions with their spouse or live-in partner than those with nonaffective disorders, and than those with no psychiatric disorders. There were no significant differences in quality of interpersonal relationships between respondents with major depressive disorder and those with dysthymia. Among those with major depressive disorder, comorbidity or treatment-seeking behavior did not significantly contribute to degree of interpersonal difficulties. The strength of the association between interpersonal dysfunction and depression were, in general, comparable for men and women with major depressive disorder. LIMITATIONS: The cross-sectional design of this report precludes inferences regarding causality between quality of interpersonal relationship and current major depressive disorder. CONCLUSIONS: The results of this study indicate that, relative to psychiatric illness in general, poor intimate relationships are characteristic of a current major depressive disorder. PMID- 10854638 TI - The nature of bipolar depression: implications for the definition of melancholia. AB - AIM: To examine if melancholic depression is over-represented in those with 'bipolar depression' and, if confirmed, to use that phenomenon to assist the clinical definition of melancholia. METHODS: We contrast 83 bipolar and 904 unipolar depressed patients on three melancholic sub-typing systems (DSM, Clinical and CORE system) and compare representation of their clinical depressive features. RESULTS: By all three melancholic sub-typing systems, the bipolar patients were more likely to receive diagnoses of 'melancholia' and of psychotic depression. To the extent that this differential prevalence of depressive sub types was reflected in varying patterns of clinical features, we so indirectly identified a set of items defining 'melancholia'. By such a strategy, melancholia was most clearly distinguished by behaviourally-rated psychomotor disturbance. While a number of 'endogeneity symptoms' were significantly over-represented, logistic regression analyses refined the set to psychomotor disturbance (both as a symptom and as a sign) and pathological guilt. We also established a distinctly higher prevalence of bipolar depression in those where a refined diagnosis of melancholia was made. CONCLUSIONS: Bipolar depression appears to be more likely to be 'melancholic' in type, thus providing an indirect strategy for the clinical definition of melancholia. PMID- 10854639 TI - Venlafaxine monotherapy in women with bipolar II and unipolar major depression. AB - BACKGROUND: Women with bipolar (BP) disorder have more depressive episodes and drug-induced manic switches compared to men. Current guidelines suggest treating BP type I and type II major depressive episode (MDE) with both a mood-stabilizer and antidepressant. In a post hoc analysis, we examined the safety and efficacy of venlafaxine monotherapy in women with BP II MDE. METHODS: 15 women with BP II MDE (mean+/-SD age: 37+/-12 years) were compared to 17 women with unipolar (UP) MDE (41+/-12 years). Patients were randomized to double-blind treatment with once versus twice daily venlafaxine up to 225 mg for 6 weeks. Efficacy was measured using the HAM-D(21), MADRS and CGI scales. Drug-induced manic switch episodes characterized by agitation, irritability, euphoria or mood lability were assessed at each visit. RESULTS: No episodes of drug-induced hypomania or rapid cycling were observed during 6 weeks of venlafaxine monotherapy. Similar efficacy was observed in BP and UP depressed women (p=ns). LIMITATIONS: This study was retrospective in nature and limited in patient number. Only BP II women were included in this study, and it is possible that efficacy and the manic switch rate might have differed if BP I women were included. CONCLUSION: Short-term venlafaxine monotherapy may be a safe and effective antidepressant treatment in women with BP II MDE. PMID- 10854640 TI - Rorschach markers in offspring of manic-depressive patients. AB - BACKGROUND: Previously published data show large differences between euthymic Israeli adult bipolar patients and US normative data on several measures of psychological functioning as assessed with a sensitive projective measure (Rorschach Inkblot Test). The current study examines the Rorschach performance of healthy offspring of bipolar parents and compares them to matched normal controls. METHODS: 14 asymptomatic offspring of Israeli manic-depressive parents were matched for age, gender, and other demographic variables with 14 children of normal parents. All subjects were individually administered the Rorschach Inkblot Test, and protocols were scored blindly according to the Exner Comprehensive System. RESULTS: Offspring of bipolar parents, like bipolar patients themselves, show significantly increased incidence and severity of thought disorder (as defined by Exner), lower numbers of cognitively mediated affective responses, and fewer responses indicating conventional perceptions. LIMITATIONS AND CONCLUSIONS: Although the sample size is small, this study strengthens the possibility that these measures of psychological functioning may serve as markers for manic depressive illness. PMID- 10854641 TI - Coping and medication adherence in bipolar disorder. AB - BACKGROUND: Effective treatment of bipolar disorder depends on medication adherence, yet few correlates of adherence have been identified. The pleasure experienced during some manic episodes may render some individuals reluctant to adhere to medications that reduce these 'highs'. Clinical observers identify denial of the severity or existence of illness as common to both bipolar disorder and addiction. The Alcoholics Anonymous model promotes acceptance as a pathway to abstinence adherence. This report hypothesized that acceptance coping would correlate positively and denial coping would correlate inversely with adherence to mood-stabilizing medication among individuals with bipolar disorder. METHODS: Thirty-two participants diagnosed with bipolar I disorder were administered scales from the Brief COPE and an adherence self-report measure. RESULTS: Consistent with hypotheses, curvilinear relationships between acceptance and denial with adherence were detected, suggesting that low levels of acceptance and high levels of denial undermine medication adherence. LIMITATIONS: Given the cross-sectional, naturalistic design of the study, no causal inferences can be made. CONCLUSIONS: The results uncover links between coping styles and adherence in a psychiatric population. The link between acceptance-denial coping, and mature, self-supportive behavior may point the way towards more effective psychosocial interventions. PMID- 10854642 TI - Correlates of distress in children at risk for affective disorder: exploring predictors in the offspring of depressed and nondepressed mothers. AB - BACKGROUND: Efforts to understand the correlates of psychological distress in children frequently examine possible correlates in samples of children who are selected for high levels of distress. The propose of this study was to compare distress correlates in a sample with depressed mothers, and thus at high-risk for distress, to a low-risk sample. METHODS: Examining data from part of a larger project, the association of children's depressive symptoms and internalizing and externalizing problems to maternal depression level, life stress, verbal ability, and the experience of a traumatic event were examined in a series of regression equations. RESULTS: Results indicated that children's depressive symptoms, rather than internalizing and externalizing problems, tended to be most consistently related to maternal variables, and also suggested that any experience of maternal depressive symptoms was associated with child problems. It was also found that child depressive symptoms were correlated with life events, but only for nondepressed mothers, and that at-risk children with higher levels of verbal ability were significantly less likely to report experiencing depressive symptoms and internalizing problems than were those with lower levels of verbal ability. LIMITATIONS: Because these data are preliminary, further research examining a broader array of variables is important. CONCLUSIONS: These results suggest the need for different models of these processes in different populations of children. PMID- 10854643 TI - Pathogenic and regulatory cells in demyelinating diseases. Conference proceedings. Rome, Italy, 12-15 September 1999. PMID- 10854644 TI - Glia-T cell dialogue. AB - Interactions of CD4(+) T helper (Th) cells with microglia and astrocytes are likely to play an important role in regulating immune responses as well as tissue damage and repair during infectious and autoimmune central nervous system (CNS) diseases. T cells secreting Th1-type cytokines provide inducing signals for microglia to mature into functional antigen presenting cells (APC). The ability of microglia to act as efficient APC for the restimulation of Th1 cells suggests a role for these cells in the local amplification of pro-inflammatory immune responses. Conversely, the Th2-inducing capacity of microglia and astrocytes together with their ability to produce anti-inflammatory mediators could play a role in providing counter-regulatory signals limiting CNS inflammation. In this article, we review recent studies addressing the functional significance of T cell-CNS glia interactions and present new data on the expression of cyclooxygenase-2, the inducible enzyme involved in prostanoid biosynthesis, in microglia and astrocytes during the course of experimental allergic encephalomyelitis. PMID- 10854645 TI - Antigen and superantigen presentation in the human CNS. PMID- 10854646 TI - Evidence for a role of gammadelta T cells in demyelinating diseases as determined by activation states and responses to lipid antigens. AB - In this report we review current information on the phenotypic and functional properties of gammadelta T cells in demyelinating disorders. The results support the conclusion that although gammadelta T cells show evidence of activation in patients with either multiple sclerosis (MS) or Guillain Barre syndrome (GBS), differences exist in the phenotypic and functional properties of these cells between the two diseases. In particular, our data indicate that in patients with MS the Vdelta2 subset is activated and that these cells can be induced to secrete high levels of proinflammatory cytokines. In contrast, in patients with GBS, the Vdelta1 subset is expanded and can be induced to secrete cytokines more associated with a humoral response. PMID- 10854647 TI - T cell vaccination in secondary progressive multiple sclerosis. AB - Four secondary progressive MS patients were vaccinated with bovine myelin reactive irradiated T cell lines from their peripheral blood. Patients were followed for 30-39 months, and monitored for immunological responses toward the vaccine, and for their clinical characteristics. Two patients showed stable EDSS score over time, one patient showed improvement by one EDSS step, and in the remaining patient her EDSS advanced over time. After the second inoculation there was a progressive decline of circulating whole myelin-reactive T cells, MBP143 168, PLP104-117, and MOG43-55-peptide-reactive T cells. In contrast the frequency of tetanus toxoid-reactive T cells remained unchanged. T cell vaccination (TCV) was also associated with a decline of myelin-specific IL-2- and IFN-gamma secreting T cells. Twelve T cell lines (TCL) that recognize the inoculates were isolated from the peripheral blood of two patients. Ten of these TCL were CD8(+) and lysed the inoculates in a MHC Class I restricted manner. The remaining two TCL were CD4(+), and lysed the inoculates by MHC Class II restricted cytolytic activity. All T cell lines lysed not only myelin-reactive T cells, but also TCL specific for MBP143-168, PLP104-117 and MOG43-55 peptides. Control TCL specific for tetanus toxoid were not lysed. Neutralizing anti-Fas mAb did not influence the killing. Moreover, culture supernatants from two TCL which produce IL-10, were able to block the proliferation of myelin protein-specific TCL. This effect was abrogated using mAbs specific for IL-10. The data obtained indicated that TCV using autologous irradiated bovine myelin-reactive T cells promotes an effective depletion of T cells reactive against different myelin antigens. PMID- 10854648 TI - The role of matrix metalloproteinases in autoimmune damage to the central and peripheral nervous system. AB - Members of the family of matrix metalloproteinases (MMPs) have been implicated in the pathogenesis of inflammatory demyelination. MMPs apparently mediate important steps in the genesis of inflammatory demyelination, such as cell migration, blood brain/nerve barrier breakdown, demyelination, and cytokine activation. This review will highlight in vitro as well as in vivo findings, which support the importance of this group of proteases in the pathogenesis of inflammatory demyelinating disorders of the central and peripheral nervous system. PMID- 10854649 TI - Degeneracy in T-cell antigen recognition - implications for the pathogenesis of autoimmune diseases. AB - T-cells recognize by their T-cell receptor (TCR) short peptides presented by major histocompatibility complex (MHC) molecules. Based on functional and structural data, it has become widely accepted that this interaction is highly flexible thus allowing a specific TCR to interact with a broad range of different peptide ligands. Although cross-reactivity is essential for selection and maintenance of the T-cell repertoire, it also carries the danger of inducing autoreactivity following protective immune responses. This hypothesis has been supported by a large number of findings in vitro and in vivo experimental systems. Here we discuss recent findings on cross-recognition of T-cells and provide a new experimental approach to address specificity and cross-reactivity in autoimmune disorders. PMID- 10854650 TI - Obstacles to identifying viruses that cause autoimmune disease. AB - In addition to a clear genetic disposition, environmental factors and viral infections are thought to play a role in the pathogenesis of autoimmune diseases such as type I diabetes or multiple sclerosis (MS). This article will explore, by use of paradigms developed in a transgenic mouse model, why it has been so difficult to prove a causative association between viral infections and autoimmunity. Potential pathogenetic mechanisms and their impact on devising the best strategy to prove such an association will be discussed. These thoughts might also be important for applying new immune interventions in type I diabetes or MS. PMID- 10854651 TI - The neuroprotective effect of inflammation: implications for the therapy of multiple sclerosis. AB - Autoreactive T cells are a component of the normal immune system. It has been proposed that some of these autoreactive T cells even have a protective function. Recent studies support this notion by demonstrating that (a) myelin basic-protein (MBP-) specific T cells show neuroprotective effects in vivo, and (b) activated antigen-specific human T cells and other immune cells produce bioactive brain derived neurotrophic factor (BDNF) in vitro. Furthermore, BDNF is expressed in different types of inflammatory cells in brain lesions of patients with acute disseminated leukoencephalopathy or multiple sclerosis. We postulate that the neuroprotective effect of T cells and other immune cells observed in vivo is at least partially mediated by BDNF and other neurotrophic factors. The concept of neuroprotective autoimmunity has obvious implications for the therapy of multiple sclerosis and other neuroimmunological diseases. PMID- 10854652 TI - Extended observations on MS patients treated with IM interferon-beta1a (Avonex): implications for modern MS trials and therapeutics. AB - Extended observations of the pivotal phase III clinical trial of interferon beta1a (IFNbeta1a; Avonex, Biogen) in relapsing MS patients revealed that: (1) active treatment significantly slowed the accumulation of physical disability over time, reduced clinical exacerbations and MRI brain lesions; (2) clinical efficacy did not depend on disability endpoints; (3) active treatment benefited multiple MRI measures of brain lesions; (4) cerebral atrophy occurred over 2 years in relatively mildly disabled patients; and (5) Avonex could slow the development of atrophy after the first year of treatment. Data from this study were recently used to design a new outcome measure for MS clinical trials (the Multiple Sclerosis Functional Composite), and was also the basis for two ongoing studies of IFNbeta1a: one in patients with monosymptomatic onset of MS and the other in secondary progressive MS. PMID- 10854653 TI - Shaping and tuning of the chemokine system by regulation of receptor expression and signaling: dendritic cells as a paradigm. PMID- 10854654 TI - Restricted immune responses lead to CNS demyelination and axonal damage. AB - Although autoreactive T-cells have a pivotal role in initiating the inflammatory process in experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis (MS), recent evidence suggests a relevant role for autoantibodies specific for myelin proteins as well. To examine the role of B-cells in the cerebrospinal fluid of patients with MS, we analyzed the V(H) gene usage in ten MS patients by PCR technologies. Analysis of HCDR3 length revealed an oligoclonal accumulation of B-cells. Sequence analysis of the V(H)3 and V(H)4 gamma transcripts of two MS individuals demonstrated that this accumulation was related to the expansion and somatic diversification of a limited groups of B-cell clones. These findings are indicative of a chronic and intense antigenic stimulation occurring in the CNS. Animal models, such as EAE, are of particular importance in order to elucidate the pathogenetic effector mechanisms in autoimmune demyelination. In a non-human primate model of EAE, we describe that the immunodominant T-cell epitope is presented exclusively by a monomorphic DRB1 allele, suggesting that susceptibility to EAE may be linked to this unique restriction and, therefore, providing a possible mechanism for MHC linkage to diseases. Moreover, we report on the presence of inflammation, sharp demyelination and axonal damage in EAE induced with whole myelin as well as with recombinant myelin oligodendrocyte glycoprotein (MOG), but not with myelin basic protein alone. The presence of axonal pathology was supported by immunohistochemistry with anti-amyloid precursor protein and anti-non phosphorilated neurofilaments monoclonal antibodies within early active demyelinated plaques. These findings suggest that axonal damage may be an early event in the pathogenesis of autoimmune demyelinating diseases of the CNS and highlights the importance of animal models in which therapies targeting repair and axonal survival may be exploited. PMID- 10854655 TI - Cytokine therapy in immune-mediated demyelinating diseases of the central nervous system: a novel gene therapy approach. AB - Pro-inflammatory cytokines play a crucial role in the regulatory and effector phase of the immune-mediated mechanism sustaining multiple sclerosis pathogenesis (MS) thus supporting the use of anti-inflammatory cytokines as a therapeutic option. Systemic administration of cytokines shows, however, limited therapeutic efficacy and undesirable/unpredictable side-effects. We have developed a non toxic system to deliver cytokines within the central nervous system (CNS) based on the intrathecal (i.c.) administration of non-replicative herpes simplex (HSV) type-1-derived viral vectors engineered with heterologous cytokine genes. Compared to controls, mice affected by experimental autoimmune encephalomyelitis (EAE) and i.c. injected with an HSV-1-derived vector containing the gene of the anti-inflammatory cytokine IL-4 showed a significant amelioration of clinical and pathological EAE signs. A decreased mRNA expression of the monocyte chemoattractant protein-1 (MCP-1) by mononuclear CNS-infiltrating cells was also observed. Peripheral T cells from IL-4-treated mice were not affected both in their antigen-specific proliferative response and in the cytokine secretion pattern. Our results indicate that CNS cytokine delivery with HSV-1-derived vectors is a feasible therapeutic strategy and might represent an alternative approach for the treatment of immune-mediated demyelinating diseases. Advantages of this approach over systemic cytokine administration are the high cytokine level reached within the CNS and the absence of side-effects on the peripheral immune system. The short-lasting cytokine production in the CNS after a single vector administration (4 weeks) is the limiting factor of this novel technology which, although promising, has to be improved. PMID- 10854656 TI - Genetics of rat neuroinflammation. AB - The definition of genes regulating the pathogenetic pathways of autoimmune neuroinflammation, may provide targets for new therapeutic strategies. This is not easily accomplished in human disease. Such genetic dissection can more readily be done by the use of inbred rodent strains. With these, genetic heterogeneity is avoided and variation in the environmental influences is minimized. Close mimicking of the human disease characteristics is desirable in such endeavors. Chronic relapsing experimental autoimmune encephalomyelitis (EAE) with MS-like histopathology is achieved after immunization of certain rat strains with myelin oligodendrocyte glycoprotein (MOG) or spinal cord homogenate. The major histocompatibility complex (MHC) regulate the ease by which the environmental trigger in the form of immunisation induces disease. Use of intra MHC recombinant strains demonstrated major influences from the MHC class II genome region, but additional influences from both the MHC class I and III regions. These findings now provide a basis for studies of the mechanisms for MHC controlled autoimmune pathogenicity leading to MS-like disease. Gene mapping of F2 crosses between susceptible and resistant rat strains demonstrated nine genome regions outside the MHC which regulate different phenotypes of rat EAE. Many of these co-localize with genome regions regulating other organ-specific disease such experimental arthritis, suggesting a sharing of disease pathways. Further finemapping can lead to the exact identification of disease regulating genes. Interestingly, we have also demonstrated a non-MHC gene control of the inflammatory response, in the form of glial cell activation, and neuronal degeneration, subsequent to anterior nerve root avulsion in rats. The genetic dissection of these influences may unravel pathways controlling CNS vulnerability. PMID- 10854657 TI - Persisting viruses and autoimmunity. AB - Viral infections can be responsible for the onset and sustaining of autoimmune processes. We discuss how chronic inflammation associated with viral persistence is the prerequisite for initiation of a multi-step process leading to autoimmunity. Firstly, chronic inflammation may favor the priming of autoreactive T cells that have escaped thymic selection and are specific for self-mimicking viral peptides in the periphery. In addition, viral persistence and inflammation can act synergistically to induce and sustain autoimmunity either unveiling cryptic self-epitopes, or favoring determinant spreading, or activating dendritic cells, or promoting constant priming of new autoreactive T cells, or contributing to the efficient generation of effector cells, or, finally, restimulating memory T lymphocytes. PMID- 10854658 TI - The role of costimulation in autoimmune demyelination. AB - Experimental allergic encephalomyelitis (EAE) is a T cell-mediated, autoimmune disorder characterized by central nervous system (CNS) inflammation and demyelination, features reminiscent of the human disease, multiple sclerosis (MS). In addition to the signal the encephalitogenic T cell receives through the T cell receptor (TCR), a second signal, termed costimulation, is required for complete T cell activation. The B7 family of cell surface molecules expressed on antigen presenting cells (APC) is capable of providing this second signal to T cells via two receptors, CD28 and CTLA-4. Our studies have shown that costimulation provided by B7 molecules to its ligand CD28 is important in the initiation of the autoimmune response in EAE. Further, it appears the costimulation provided by B7-1 is important in disease development, while B7-2 may play an important regulatory role. We and others later showed that B7/CTLA-4 interaction plays a critical role in down-regulating the immune response. Previous work has shown that activated T cells and T cells of a memory phenotype are less dependent on costimulation than naive T cells. T cells reactive with myelin components that are involved in the pathogenesis of EAE and possibly MS would be expected to have been activated as part of the disease process. Building upon our prior work in the EAE model, we have tested the hypothesis that myelin reactive T cells, which are relevant to the pathogenesis of CNS inflammatory demyelination, can be distinguished from naive myelin-reactive T cells by a lack of dependence upon costimulation for activation and that the costimulatory requirements of these myelin-reactive T cells change during the course of disease. Our studies in the EAE model have also addressed the mechanisms of extrathymic (peripheral) T cell tolerance following intravenous (i.v. ) administration of high dose antigen. It is believed that TCR signaling in the absence of costimulation is a vital component of peripheral tolerance mechanisms. However, recent evidence suggests that peripheral tolerance of antigen-specific T cells induced in vivo may require CTLA-4 engagement of the tolerized T cells. We have begun to examine the molecular mechanisms of tolerance induction following intravenous and intraperitoneal administration of myelin antigens in the EAE model and test the hypothesis that tolerance induction is dependent on the B7:CD28/CTLA-4 pathway. The results from our studies will enhance our understanding of the role that myelin-reactive T cells may play in the pathogenesis of MS. We have determined that MBP-reactive T cells in MS patients are less dependent upon CD28 costimulation than in normal controls, suggesting that these T cells were previously primed in vivo. Characterization of these CD28 independent myelin-specific T cells will have broad implications for a variety of immunologically based therapies in diseases such as MS. PMID- 10854659 TI - Global immune disregulation in multiple sclerosis: from the adaptive response to the innate immunity. AB - Increasing evidences show a global immune disregulation in multiple sclerosis (MS). The possible involvement of myelin and non-myelin (auto-)antigens in the autoaggressive process as well as the disregulation of both adaptive and innate immunity challenge the concept of specific immunotherapy. T cells at the boundary between innate and adaptive immunity, whose immunoregulatory role is becoming increasingly clear, have recently been shown to bear relevance for MS pathogenesis. Global immune interventions (and type I interferons may be considered as such) aimed at interfering with both innate and acquired immune responses seem to be a most promising therapeutic option in MS. PMID- 10854660 TI - Involvement of natural cytotoxicity receptors in human natural killer cell mediated lysis of neuroblastoma and glioblastoma cell lines. AB - The surface receptors involved in natural killer (NK) cell triggering during the process of target cell lysis have been at least in part identified. These are members of a novel family of receptors that has been termed natural cytotoxicity receptors (NCR). The first three members of this emerging group of receptors are the NKp46, NKp44 and NKp30 molecules that all belong to the immunoglobulin superfamily. Blocking of these receptors inhibits NK-mediated cytotoxicity against a wide variety of tumor target cells. In the present study, we show that these NCR are also involved in NK-mediated killing of tumor cells of neural origin. Glioblastoma and neuroblastoma target cells were efficiently killed by all NK clones analyzed since little protection from NK lysis was mediated by HLA class I molecules. Blocking of one or another NCR inhibited cytotoxicity; however, optimal inhibition was only observed when the three receptors were blocked simultaneously. A sharp difference in cytotoxicity against neural tumors was demonstrated between NCR(bright) and NCR(dull) NK clones, further supporting the notion that NCR play a critical role in the induction of cytotoxicity against tumor target cells of different histotype. Finally, our data also indicate that CD16 does not function as a triggering receptor involved in lysis of neural tumors since no difference in cytotoxicity could be substantiated between CD16(+) and CD16(-) NK clones and no correlation could be detected between the NCR(bright)/NCR(dull) phenotype and CD16 expression. PMID- 10854661 TI - T cell design for therapy in autoimmune demyelinating disease. AB - It has become increasingly more evident that a considerable refinement of currently used reagents and conditions will be needed before an effective gene therapy strategy can be used in the treatment of human autoimmune diseases. Such refinements will focus on optimizing three basic requirements for effective gene therapy, viz.: (1) targeted delivery of the therapeutic gene and/or its gene product in a reliable, efficient manner; (2) long-term expression of the therapeutic gene; and (3) regulated expression of the therapeutic gene so that it is activated only when needed. Using an experimental autoimmune encephalomyelitis mouse model, we have examined the potential for using the T cell as a gene therapy vector for targeted, long-term, regulated delivery of therapeutic transgene factors to the autoimmune inflammatory milieu. Our data indicate that the autoreactive T cell may serve as a useful endogenous vector for antigen inducible, site-specific delivery of a variety of therapeutic transgene factors capable of mediating both inhibition of autoimmune inflammation and regeneration and/or protection of damaged tissue. PMID- 10854662 TI - Residual public repertoires to self. AB - The consensus view about the constitution of the T cell receptor repertoire has shifted greatly even during this decade. Although the discovery of autoimmunity in the fifties had clearly shown that a repertoire must exist directed against self antigens, the extent of this repertoire was not fully appreciated. In our work we have tried to elucidate the nature of the antigenic specificities against which this self-directed repertoire is directed. The non-tolerized (residual) self-directed repertoire is a direct consequence of the hierarchy of antigenic determinant display, and is the most important influence in the organism's choice of which T cells to delete. Certain determinants remain "silent" and are neither displayed in the thymus nor in the periphery: these are a heterogeneous group which are invisible to T cells for a variety of reasons. One reason relates to the processing and presentation of determinants, and a second derives from the nature of the T cell receptor (TcR) and the avidity of the T cell for its target specificity. PMID- 10854663 TI - Current research on influenza and other respiratory viruses: II international symposium. PMID- 10854664 TI - Targeted cleavage of HIV-1 coreceptor-CXCR-4 by RNA-cleaving DNA-enzyme: inhibition of coreceptor function. AB - HIV needs the chemokine receptors (HIV-1 coreceptors) to initiate infection and gain entry into a susceptible cell. CCR5 receptor is used by macrophage tropic viruses to establish infection, and CXCR-4 is used by T lymphocyte tropic virus which are usually found at the terminal stages of the disease. These chemokine receptors are, therefore, attractive targets to interfere with the entry as well as spread of HIV-1 in the host. As our antiviral approach, we have earlier assembled a DNA-enzyme-916 against CCR5 (Goila and Banerjea, 1998). We have now designed against the CXCR-4 gene a mono-DNA-enzyme, which showed sequence specific cleavage activity. When CXCR-4-DNA-enzyme was placed in tandem with CCR5 DNA-enzyme, specific cleavage of their respective target sites were observed using a 60 bases long synthetic target RNA which possessed the target sites for both the DNA-enzymes. The cleavage by the CXCR-4 DNA-enzyme was found to be significantly more efficient than by the CCR5-DNA-enzyme. Analyses of the cleaved fragments by mono- and di-DNA-enzyme indicated strongly that hybridization of the CCR5-DNA-enzyme with its cognate target RNA, actually facilitated the cleavage by the CXCR-4 DNA-enzyme. Furthermore, the di-DNA-enzyme was able to cleave the substrate RNA to completion. These DNA-enzymes, when introduced into a mammalian cell line expressing the appropriate chemokine receptor, interfered specifically with the HIV-1 coreceptor functions. Using this strategy, it may be possible to interfere with the infection and spread of R5 as well as X4 viruses. PMID- 10854665 TI - Topical effects of cidofovir on cutaneous rabbit warts: treatment regimen and inoculum dependence. AB - The present study examined topical effects of cidofovir on cutaneous rabbit warts. Based on an inoculum-dependency study, each New Zealand White rabbit was inoculated with a high and low titer of cottontail rabbit papillomavirus (CRPV) at four sites on each dorsolateral area. Inoculation with 50 ID(50) induced papillomas at 100% of the inoculation sites within 16+/-1 days, and the wart growth curve plateaued within approximately 7 weeks. With an inoculum of 5 ID(50), 80% of the inoculated sites developed papillomas within 21+/-1 days and their size plateaued at a later time. Cidofovir was applied topically twice daily on the inoculated sites at a concentration of 1% for 18 days, starting at three different time points. In the first experiment, treatment was initiated 7 days post-inoculation. One of the inoculated sides received cidofovir or the vehicle, PBS, while the other side was left untreated. With this treatment regimen, cidofovir significantly delayed the time of onset and the growth rate of papillomas induced with the high titer of inoculum. It completely prevented papilloma-induction on the sites inoculated with the low titer of CRPV. Reversible side-effects of cidofovir were observed on the directly treated area including erythema, necrosis, and flaking. Both therapeutic and side-effects were limited to the sites of direct exposure. In the second experiment, one of the two sides in each group of rabbits received cidofovir or vehicle starting on day 29 post-inoculation. With this treatment regimen, cidofovir significantly reduced wart growth against the low titer only. Topical treatment initiated on day 49 post-inoculation was not effective on warts initiated with either viral titer. These results demonstrated that topical cidofovir could be very effective against papillomavirus-induced wart growth if it is initiated early during the infection, especially against low titers of inoculum. PMID- 10854666 TI - The antiviral activity exerted by vaccinia virus on the growth of herpes simplex virus in BS-C-1 cells. AB - The growth of herpes simplex virus type 2 (HSV-2) in BS-C-1 cells, was inhibited following super-infection with vaccinia virus. This inhibition was efficiently induced by both the intracellular mature virus (IMV) form of vaccinia virus and the extracellular enveloped virus (EEV), containing an additional external viral membrane. Treatment of vaccinia IMV with the detergents NP-40, Brij-58 or n-octyl alpha-D-glucopyranoside, abolished its ability to inhibit the growth of HSV-2. Ultraviolet irradiation of vaccinia virus, that completely inactivated the infectivity of the virus, resulted in partial loss of the capability to inhibit the growth of HSV-2: 16-fold more irradiated virus was needed for the inhibition. Electron microscopy showed that the irradiated vaccinia virus adsorbed and penetrated into the HSV-infected cells but remained morphologically intact within the cells for at least 22 h. When the steps in the growth of HSV affected by the irradiated vaccinia virus were followed, it was found that while the synthesis of HSV DNA was partially decreased, the synthesis of HSV proteins was very strongly inhibited and virus particles were not formed. PMID- 10854667 TI - Genetic evolution of GB virus C/hepatitis G virus (GBV-C/HGV) under interferon pressure. AB - The epidemiology and clinical features of chronic GBV-C/HGV infection have largely been explored, but there is little information about the mechanisms enabling GBV-C/HGV to cause persistent infection. Since analysis of the genomic variation of GBV-C/HGV under interferon pressure might provide some insight into this issue, we analyzed the nucleotide sequence variation of the 5'NC and NS3 regions in GBV-C/HGV isolates obtained sequentially from seven patients co infected with HCV and treated with interferon. A reduction of GBV-C/HGV-RNA serum level below the detection limit of the RT-PCR assay was observed during treatment in all patients, but upon interferon withdrawal, viral RNA remained undetectable in only two patients. Among the five patients who did not clear GBV-C/HGV-RNA, viral strains emerging after treatment were identical to those present at baseline in three cases. In a further case, in whom GBV-C/HGV-RNA re-emerged during therapy (breakthrough episode), several mutations appeared in relapse samples. In the remaining patient, with a mixed infection before therapy, only one of the two GBV-C/HGV strains present at baseline was detected upon treatment withdrawal. These data raise the possibility that positive selection may act over GBV-C/HGV genome during interferon therapy, and contribute to persistence of infection with this virus. PMID- 10854668 TI - Effect of sediment-chemical contact time on availability of sediment-associated pyrene and benzo AB - Hydrophobic organic chemicals achieve equilibrium in sediments and soils by adsorption and desorption processes driven by diffusion into organic material of particle aggregates. Exposure of benthic animals to contaminated sediments results in bioaccumulation and, in theory, a steady state is established between sediment organic carbon and the organism lipids. The purpose of this study was to test the importance of sediment-chemical contact time in bioaccumulation of pyrene and benzo[a]pyrene to the freshwater oligochaete, Lumbriculus variegatus. Also, the applicability of semi-permeable membrane devices (SPMDs) in mimicking accumulation of sediment-associated chemicals by benthic invertebrates was evaluated. Feeding and nonfeeding animals were exposed to dual spiked lake sediment in five consecutive 144 h accumulation tests. SPMDs were exposed in five consecutive single point 12 h exposures to test the effect of aging of sediment on accumulation. SPMDs were also exposed in a 28 day accumulation test to determine uptake rate coefficients. Increase in sediment-chemical contact time decreased pyrene and benzo[a]pyrene uptake clearance coefficients of successive exposures for both feeding and nonfeeding animals. This decrease in bioavailability was strongest at the start of contact and slowed down with time. Ingestion of sediment considerably increased accumulation of both compounds indicating the importance of feeding behavior in bioaccumulation of sediment associated chemicals. The significance of ingested sediment as uptake route for pyrene varied between exposures. This was probably due to combined effect of variable ingestion rate and decreasing bioavailable fraction of chemical. Availability of PAHs decreased also for SPMDs with increasing sediment chemical contact time. SPMDs reached curvilinear portion of overall uptake curve in 28 days and calculated uptake clearance constants (k(s), g sediment/g SPMD per hour) corresponded to constants calculated for nonfeeding organisms. The results of this work emphasize the significance of sediment-chemical contact time in bioaccumulation and the importance of feeding behavior of deposit feeders in bioaccumulation of sediment-associated contaminants. Both should be taken into account when performing and modeling bioaccumulation of sedimented contaminants. Furthermore, uptake of sediment-associated PAHs by SPMDs appeared to mimic uptake by nonfeeding organisms. However, more research is needed to compare the uptake of artificial devices to sediment-dwelling species in sediment exposures. PMID- 10854669 TI - Pollutant-induced over-activation of phagocytes is concomitantly associated with peroxidative damage in fish tissues. AB - Pollutant-induced abnormal functioning of phagocytes and associated consequences were studied in freshwater catfish Heteropneustes fossilis (Bloch). Fish were exposed to effluent collected from bleached kraft type of paper mill at the concentration levels of 0.5, 1 and 2% for 15, 30, 60 and 90 days. Respiratory burst activity of peritoneal and head kidney phagocytes of exposed fish was measured by nitroblue tetrazolium reduction assay. Lipid peroxidation (LPO) was estimated in liver, gill and kidney of fish by measuring thiobarbituric acid reaction substances. It was observed that the phagocyte-activating xenobiotics of effluent induced an increase in the respiratory burst activity in phagocytes. The induction of respiratory burst activity was concomitantly associated with an increase in the peroxidative damage of tissues. The tissues most affected were kidney and gills. The change in LPO values in the gills of exposed fish was concentration- and time-dependent, showing significant increases (P<0.05 to <0.001) in all the exposed groups as compared with control fish. An almost similar pattern of LPO was observed in head kidney tissue (P<0.05 to <0.001). As regards liver, increase in LPO was not widespread, except at 0.5% for 90 days (P<0.05). In fact, reduced rates of LPO were observed in the livers of some groups. The results of respiratory burst corroborate with the phagocytic activation as well as with the extent of lipid peroxidation in the tissues, showing high population of circulatory phagocytes. Our results demonstrate that fish of polluted water are subjected to oxidative stress of multifarious dimensions. PMID- 10854670 TI - Optimization of a precision-cut trout liver tissue slice assay as a screen for vitellogenin induction: comparison of slice incubation techniques. AB - An in vitro male rainbow trout liver slice assay has been developed for long-term incubation of precision-cut slices for the detection of vitellogenin (VTG) protein induction in response to xenobiotic chemicals. The assay was optimized to allow 72 h of incubation of slices to maximize detection of VTG, while maintaining slice viability. Two methods of incubation frequently used with rat liver slices were compared: (1) slices were submerged in media (11 degrees C) and cultured in 12-well plates (PL) with continuous shaking; or (2) slices were floated onto titanium screens, placed into glass vials, and held under dynamic organ culture (DOC) conditions (11 degrees C). Slices (200 um) in modified L-15 media were exposed to 1.0 uM 17beta-estradiol (E2) or diethylstilbestrol (DES). Protein from media and slice was sampled for Western blot analysis, using a polyclonal antibody to detect appearance of VTG protein. Maximum VTG was seen at 72 h, with detectable protein at 24 and 48 h in slices and media following PL incubation. In contrast, slices incubated in DOC showed little detectable VTG above background levels after 72 h. This difference was not attributable to protein loss to vial or plate surfaces. Standard viability assays did not reveal any differences between slices incubated in PL or DOC. However, histopathological examination revealed earlier and more severe vacuolization in slices incubated in DOC. Significantly more E2 uptake and conversion to water-soluble metabolites was noted in PL, compared with DOC, as well as more production of VTG in response to DES and E2, correlated with less histologic change. The in vitro assay described allows tissue-level assessment of estrogenicity in aquatic organisms, and will be useful for assessing not only comparative species receptor binding and transactivation, but also the role of tissue-specific activation factors in the estrogenic response of fish. PMID- 10854671 TI - Enzymatic biomarker measurement and study of DNA adduct formation in benzo AB - The aim of this study was to improve the knowledge on the metabolic pathways involved in benzo[a]pyrene (B[a]P) activation and on the relationship between adduct levels and enzymatic biomarker activities. With this purpose, a model to assess pollutant exposure via food supply has been developed for the sentinel organism, Mytilus galloprovincialis. Mussels were fed for 4 weeks with B[a]P contaminated feed (50 mg/kg dry weight mussel). Bioaccumulation was studied by determination of B[a]P concentration in whole mussel by GC/MS analysis. Different biomarkers of pollutant exposure were measured to assess the metabolic state of the exposed organisms. CYP1A-like immunopositive protein titration and B[a]P hydroxylase (BPH) activity were assessed as indicators of phase I biotransformation. Glutathione-S-transferase (GST) activity was measured as an indicator of the conjugation activities. Catalase (CAT) and DT-diaphorase (DTD) activities were assessed as potential biomarkers of oxidative stress, whereas acetylthiocholine esterase (AChE) activity was measured as an indication of possible neurotoxicity of B[a]P exposure. DNA adduct levels were determined in digestive gland DNA by applying the 32P-postlabeling technique with nuclease P1 enhancement. For the developed conditions of exposure, B[a]P concentration reached in whole mussel tissues was very high (>500 mg/kg d.w. mussel) and significant B[a]P-induced changes were recorded for each enzymatic biomarkers. BPH and CAT activities were significantly increased by B[a]P exposure, whereas GST in the gills, DTD and AChE were significantly depressed. On the other hand, no change in CYP1A-like immunopositive protein content was observed. Induction and increase with time of bulky B[a]P-related DNA adducts were demonstrated in the digestive gland, although at low levels (0.269+/-0.082 adduct/10e8 dNps at maximum) by the 32P-postlabeling assay. DNA adduct level was significantly correlated with whole mussel tissue B[a]P concentration, so were all the enzymatic biomarkers measured except to GST activity in both gill and digestive gland tissues. BPH, DTD, CAT and AChE displayed a strong correlation with adduct levels. These results demonstrate the neurotoxicity and the genotoxicity of B[a]P exposure in the mussel. The induction of bulky DNA adducts in mussels demonstrates the existence of activation pathways already identified in vertebrates. It validates also the suitability of this model for further studies on B[a]P metabolism in mussels. Our results support the proposal of BPH, AChE, DTD and CAT activities as suitable biomarkers of PAH exposure for these sentinel species. PMID- 10854672 TI - In vivo and in vitro metabolism and organ distribution of nonylphenol in Atlantic salmon (Salmo salar). AB - In the environment, nonylphenol (NP) occurs predominantly as a degradation product of nonylphenol ethoxylate (NPE). They can be found in many types of products including detergents, plastics, emulsifiers, pesticides, and industrial and consumer cleaning products. As a consequence of their use in a variety of products, they are quite common in rivers and other aquatic environments that receive sewage discharges. Because of its enhanced resistance towards biodegradation, toxicity, estrogenic effects, and ability to bioaccumulate in aquatic organisms NP has been regarded as the most critical metabolite of APEs. We have studied the in vivo and in vitro metabolism and organ distribution of NP in juvenile salmon. Fish were exposed in vivo to waterborne [3H]-4-n-NP for a period up to 72 h or were administered a single oral dose of [3H]-4-n-NP. In vitro biotransformation of NP was studied by exposure of cultured salmon hepatocytes to [3H]-4-n-NP in the presence or absence of a CYP1A-inducer, beta naphthoflavone (betaNF). Our results show that 4-n-NP was mainly metabolized in vivo, to its corresponding glucuronide conjugates and hydroxylates. The major route of excretion was the bile. The half-life of residues in carcass and muscle was between 24 and 48 h in both waterborne and dietary exposure. In whole body autoradiography, intragastric administered [3H]-4-n-NP was mainly present in the gastrointestinal tract and bile. NP-derived radioactivity in fish exposed via water was more evenly distributed in the organs compared to intragastric exposure and were observed in the intestinal contents, liver, kidney, gills, skin, abdominal fat and brain. In vitro pretreatment of hepatocytes with betaNF had no effect on rates or patterns of NP biotransformation. The in vitro metabolic rate of NP were 118 pmol NP metabolized/h/0.5x10(6) cells without betaNF, and 98 pmol NP metabolized/h/0.5x10(6) cells when betaNF was added to the culture medium. PMID- 10854673 TI - Omapatrilat--the ups and downs of an exciting but complicated new drug. PMID- 10854674 TI - A suggestion for familial hypercholesterolemia (FH) heterozygosity clinical diagnosis based on epidemiological observations in a large Italian population. AB - We selected 247 subjects from 29 large familial hypercholesterolemia (FH) kindreds from 550 probable FH subjects in Emilia Romagna (Italy) on the basis of LDL-cholesterol plasmatic levels and family trees, in order to define the best diagnostic criteria for heterozygous patients. Familial hypercholesterolemia is a monogenic disease of cholesterol metabolism inherited as an autosomal dominant trait and characterised by early cardiovascular disease. A low xanthomas and xanthelasmas prevalence was found (8.6%); coronary heart disease (CHD) death occurs very frequently in heterozygous males (72% of all deaths; mean age at death 52 years), while in females the primary cause of death was thrombotic stroke (55%; mean age 69 years). Total cholesterol (TC) mean values were 389.8 (m) and 373.3 mg/dl (f) for FH trait carriers, and 223.3 (m) and 228.8 (f) for healthy relatives. No age-related change in TC was found in heterozygotes, while unaffected relatives of FH families showed mean TC and LDL-C values, and a TC frequency distribution and a TC age-related increasing trend similar to the expected values for the Italian population. The TC frequency distribution curve appeared bimodal, with a mid-point between heterozygous and homozygous FH modal values of 280 mg/dl. To identify the FH patients, the final FH heterozygosity risk was evaluated in an unselected free-living population (from 0.07 to 0.8%, respectively, for TC between 265-274 and 295-304 mg/dl) and in hypercholesterolemic families (31 to 83%, and the same TC classes). Our conclusion is that the clinical picture is rarely pathognomonic, while the FH heterozygosity final risk evaluation and the 280 mg/dl cut-off point can be used to guide the practical clinical diagnosis and to select the patients destined for B-E receptor activity evaluation. PMID- 10854675 TI - On the diagnosis of heterozygous familial hypercholesterolaemia HFH. PMID- 10854676 TI - Plasma endothelin-1 and thrombomodulin levels in coronary sinus during right atrial pacing and percutaneous transluminal coronary angioplasty. AB - Increases in the levels of plasma endothelins (ETs) have been reported after percutaneous transluminal coronary angioplasty (PTCA). To examine the mechanism of this increase, we measured plasma endothelin-1 (ET-1) and thrombomodulin (TM) levels in both the Valsalva sinus (VAL) and the great cardiac vein (GCV) together with oxygen saturation of the GCV (Sat.GCVO2) during right atrial pacing and PTCA. Thirty patients with stenoses in the left anterior descending coronary arteries were enrolled. A fiberoptic pulmonary artery catheter was placed in GCV for monitoring Sat.GCVO2, and blood sampling was repeated before and after each procedure. ET-1 did not increase during pacing, but after PTCA it significantly increased from basal levels to 24.4+/- 8.3 pg/ml in GCV (P<0.01) and 19.3 +/-7.4 in VAL (P<0.05). Basal TM levels in GCV and VAL were significantly higher in diabetic than in non-diabetic patients, but TM did not change during pacing and PTCA. Sat.GCVO2 significantly decreased from the basal level during pacing and PTCA. We speculate that direct endothelial cell damage is more responsible for the increase of ET-1 during PTCA than myocardial ischemia. Our data indicate that ET-1 may be a useful marker for acute endothelial damage, while TM reflects only chronic and general damage of endothelial cells. PMID- 10854677 TI - Radiofrequency thermal balloon angioplasty in an experimental model of peripheral arterial stenosis. AB - RATIONALE: To assess the effect of thermal balloon angioplasty on surgically created peripheral arterial stenoses. METHODS: Unilateral femoral arterial stenoses were created in 17 neonatal lambs (Dorset X Suffolk/Mule), using absorbable sutures. Six to 8 weeks later, the stenoses were dilated using either a standard (N=5), or thermal balloon angioplasty (N=9). Immediate angiography, flow and pressure gradient measurements were made to assess the acute result following either procedure. The survivors were followed up for a further period of 4 to 6 weeks, and a terminal angiographic study performed. The thermally treated segments of vessels were removed for qualitative histologic analysis. RESULTS: Thermal angioplasty appeared to be acutely successful in eight of nine animals, compared with one of five successful procedures following standard angioplasty. Higher therapeutic temperatures (> or = 80 degrees C) were associated with vascular complications. At the terminal study, stenoses had recurred in four of six survivors successfully treated with thermal angioplasty. Histologic studies demonstrated non-uniform effects of thermal dilation on the vessel wall, with variable changes ranging from partial or full-thickness tears, fibroblastic or myocyte proliferation, and disorganization of the vessel wall layers. CONCLUSION: Thermal angioplasty appears to be acutely more beneficial compared with standard angioplasty. However there is a significant recurrence of stenoses, and non-uniform changes in the vessel wall. PMID- 10854678 TI - The human paraoxonase Gln-Argl92 (Q/R) polymorphism in turkish patients with coronary artery disease. AB - It has been suggested that a Q/R (Glnl92Arg) polymorphism of paraoxonase (PON) might be associated with the predisposition to coronary artery disease (CAD). Therefore, we studied the human paraoxonase gene (PON1) polymorphism in Turkish patients with CAD by polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP). This polymorphism was determined in 96 CAD patients and in 105 control subjects. The frequencies of the QQ, QR, and RR genotypes were found as 36.5, 52.0, and 11.5% in CAD patients and 48.6, 41.0, and 10.4% in control subjects, respectively. The QR genotype was the most common in the patient group, whereas the QQ genotype was more frequent in individuals without CAD. Frequency of the R allele was higher among CAD patients compared to controls (38.5% versus 31%). However, neither the genotype nor the allele distribution of the Gln92Arg polymorphism of PON1 was statistically significantly different between the two groups (P>0.05). Although both systolic and diastolic blood pressure levels were slightly higher in patients with the QQ genotype, there was no differences in regard to age, sex, serum triglyceride, total cholesterol or high-density lipoprotein cholesterol among CAD patients with different PONI Gln192Arg genotypes. In summary, our results suggest that no association exists between the Gln192Arg polymorphism of paraoxonase and CAD in Turkish patients. PMID- 10854679 TI - Primary congenital anomalies of the coronary arteries: a coronary: arteriographic study. AB - Geographic variations in the incidence of different congenital coronary anomalies are well known, but infrequently studied in the Indian population. Among 4,100 adult patients who underwent diagnostic coronary arteriography, 39 (0.95%) patients (34 males, 5 females) had one or more anomalous coronary arteries. Their mean age was 46.4 +/- 8.2 years (range, 26-68 years). Thirty-five (89.74%) had anomalies of origin and distribution, while the remaining four (10.25%) had coronary artery fistulae. Right coronary artery was the commonest anomalous vessel, involved in 19 (48.74%) patients. It was originating from the left sinus of Valsalva in 15 and from the non-facing aortic sinus in four patients. Anomalous left circumflex artery was the second commonest anomaly, seen in 14 (35.89%) patients. Anomalous left anterior descending artery and anomalous left coronary artery from pulmonary artery were seen in one patient each. Among patients with coronary fistulae, two had fistulae between the left anterior descending artery and the main pulmonary artery, one between the conal artery and the right atrium, while the fourth patient had fistulae from the right coronary as well as from the left anterior descending artery to the left atrium. Atherosclerotic plaques in the anomalous arteries were seen in only 13 (33.33%) patients, much less than the overall incidence of coronary artery disease in patients with congenital coronary anomalies in this series (66.66%). In four (10.25%) patients, only the anomalous vessels were involved in coronary artery disease. Thus, in a small subgroup there does not appear to be an increased risk for development of atherosclerotic coronary artery disease in anomalous coronary arteries. Recognition of coronary anomalies is important in patients undergoing coronary arteriography, coronary interventions and cardiac surgery. Variations in the frequency of primary congenital coronary anomalies may possibly have a genetic background. PMID- 10854680 TI - Continuing controversy in the management of concomitant coronary and carotid disease: an overview. AB - OBJECTIVES: To perform an analytical overview of the risk factors, pathogenesis of stroke and the strategies for the management of concomitant coronary artery disease and carotid artery stenosis (CAS). Four strategies were analysed; CABG in the presence of CAS, combined (CE+CABG), reverse (CABG+CE<3 months) and prior staged (CE+CABG<3 months). METHODS: A literature search formed the basis of a reference database. Outcome was assessed by the 30-day permanent stroke and mortality rate for the different approaches. Accrued rates of permanent stroke and mortality rate were expressed in terms of mean stroke and mortality rate (MSR, MMR). Data was analysed comparatively and expressed in terms of P value, odds ratio and confidence limits. RESULTS: 33 different risk factors for stroke at CABG were identified. Significant factors included: ascending aortic atheroma, emergency procedures, impaired left ventricular function, cardioplegia and peripheral vascular disease. Risk of stroke at CABG increased with higher grade CAS (50 vs. 80%, P=0.009). Pathogenesis of stroke at CABG is multifactorial; the role of flow limiting CAS is controversial and other mechanisms are implicated. Analysis of the four strategies revealed that in the Prior Stage (n=573) the MSR was 1.5% and MMR 5.9%, in the Unprotected CABG+CAS series the MSR was 3.8% (n=840) and MMR (n=596) 4.4%, in the Reverse stage series (n=83) the MSR was 2.4%, and MMR 4.8%. For Combined procedures (n=3,295) the MSR was 3.9% and MMR 4.5%. Comparative analysis indicated a significant reduction in stroke for Prior vs. Combined (1.5 vs. 3.9%, P=0.007, odds 0.39, CI 0.2-0.77) with a higher mortality (5.9 vs. 4.5%, P=0.1, odds 1.41, Cl 0.96-2.06, NS). The stroke rate in the Prior stage also remained significantly lower compared to the Unprotected CABG group both mixed (P=0.015) and asymptomatic CAS (P=0.047). When total risks (MSR+MMR), were analysed, similar results were found between the groups; Prior 7.4%, Reverse stage 7.2%, Combined 8.4%, Unprotected CABG+ >50% CAS 11.5%. CONCLUSIONS: Stroke at CABG is due to multiple risk factors, one of which is high grade carotid stenosis. Pathophysiology of stroke, although multifactorial, supports embolism rather than flow limitation as the primary mechanism. Lack of randomised trials has made it impossible to draw firm conclusions regarding the best management strategy. There was no significant difference in the overall stroke and mortality risk between the various strategies, however, subgroup analysis suggests that, when carefully selected, patients do better by staging the operations. In our opinion patients without severe cardiac disease should be considered for Prior staging and the rest for Combined procedure. The role of reverse staging needs further evaluation. PMID- 10854681 TI - Positron emission tomography is a useful tool in differentiating idiopathic from ischemic dilated cardiomyopathy. AB - We evaluated the utility of positron emission tomography in differentiating patients with idiopathic dilated cardiomyopathy from those with ischemic cardiomyopathy. Twenty consecutive non-diabetic patients with dilatation (end diastolic volume > or = 120 cc/m2) and reduced systolic function (ejection fraction < or = 40%) of the left ventricle on cineangiography, underwent coronary angiography, F18 fluorodeoxyglucose (F18-FDG) (glucose load technique) and N13 ammonia (N13-NH3) positron emission tomography. A semiquantitative score based on the extension and the severity of the uptake defects was calculated. Endomyocardial biopsy was performed in patients with normal coronary arteries. Ten patients (group A) had normal coronary arteries and histologic features of the endomyocardium fitting with the diagnosis of idiopathic dilated cardiomyopathy. Cineangiography showed critical stenosis of at least one major coronary artery in the other 10 patients (group B). The two groups were similar in age. left ventricular end-diastolic volume and ejection fraction. Both N13 NH3, positron emission tomography and F18-FDG positron emission tomography scores were lower in group A than in group B: 0.1 +/- 0.3 vs. 10.6 +/- 5.1 (P<0.0001) and 2.4 +/- 4.4 vs. 9.9 +/- 4.1 (P<0.0001) respectively. but only N13-NH3 positron emission tomography allowed a complete separation of the two groups (score range 0-1 group A vs. 4-12 group B). The F18-FDG score value showed some overlapping between the two groups (score range 0-12 in the group A vs. 2-17 in the group B). All three idiopathic dilated cardiomyopathy patients with a F18-FDG score value >2 had left bundle branch block on standard ECG. Positron emission tomography imaging with N13-NH3 and F18-FDG provided a complete differentiation between idiopathic dilated cardiomyopathy and ischemic cardiomyopathy patients. However patients with left bundle branch block on ECG could present defects in FDG uptake even if affected by idiopathic dilated cardiomyopathy. PMID- 10854682 TI - Discovering the aetiology of heart failure, one link in the chain to improved survival. PMID- 10854683 TI - Effects of enalapril and isradipine alone and in combination on blood pressure, renal function and echocardiographic parameters in mild hypertension. AB - AIMS AND METHODS: A study was carried out to evaluate the influence of antihypertensive treatment with combined low doses of enalapril plus isradipine (5+5 mg daily) compared with those of either drug at a higher dose level (10 mg daily) by double-blind, three-way crossover study (balanced Latin square design) in 102 subjects (mean age 51.9 +/- 7.42 years) with essential hypertension. Left ventricular mass and function were evaluated by M-B mode echocardiography, renal function by glomerular filtration rate (GFR) and by serum and 24-h urinary Na+ and K+ during wash-out period and after 24 weeks of treatment. RESULTS: The supine blood pressure for subjects given placebo was 171/103 mmHg. After 24 weeks of treatment, systolic and diastolic supine blood pressure were significantly lower with 5 mg isradipine plus 5 mg enalapril (134/84 mmHg) than with 10 mg enalapril (137/84 mmHg) or with 10 mg isradipine (144/85 mmHg). Left ventricular posterior wall and septal thickness were significantly and similarly reduced in all groups. Left ventricular systolic and diastolic end diameters were not significantly changed. Left ventricular mass (LVM) was significantly reduced in E plus I group and enalapril group. GFR was not significantly altered. The 24-h urinary Na+ significantly increased with enalapril, more so than isradipine. The combination was tolerated better than either monotherapy. We observed no clinically significant changes in laboratory variables including blood lipoproteins. CONCLUSIONS: The combination of isradipine plus enalapril reduced blood pressure more effectively and was better tolerated than other drug alone. All three groups showed similar changes in echocardiographic indices and no change in renal function. PMID- 10854684 TI - Modulation of tight junctional permeability. PMID- 10854685 TI - The molecular structure of the tight junction. AB - The maintenance of a barrier with controlled permeability is an important characteristic for multi-cellular organisms. In mammalian cells, the tight junction functions in that role allowing compartments with different solute composition to be separate, but not absolutely unconnected. The permeability of this paracellular zone needs to be controlled by both internal and external factors allowing for modulation of the permeability under certain circumstances. The purpose of this chapter is to introduce the reader to the molecular components of the mammalian tight junction. Also provided, is a brief description of how these junctional components interact with other members of the tight junction plaque and components of both the cytoskelton and signaling cascade. PMID- 10854686 TI - Show me the pathway! Regulation of paracellular permeability by Na(+)-glucose cotransport. AB - The physiological impact of Na(+)-nutrient cotransport-dependent regulation of intestinal tight junction permeability has been controversial. Nonetheless, increased permeability of small intestinal mucosae and enterocyte tight junctions as a consequence of Na(+)-nutrient cotransport has been documented by a significant number of in vivo and in vitro studies. Some details of the intracellular signaling events that regulate this process have been described recently. The aims of this article are to: (i) review studies of tight junction regulation and paracellular nutrient absorption in mammalian intestine, (ii) identify potential applications of tight junction regulation, and (iii) summarize recent progress in defining molecular mechanisms that lead to altered tight junction permeability. PMID- 10854687 TI - Modification of tight junction function by protein kinase C isoforms. AB - The regulation of tight junction permeability by a variety of signal transduction pathways is summarized. An emphasis is placed on regulation of paracellular permeability by the protein kinase C family of isoforms, which involves the reporting of a large number of studies using the phorbol ester family of protein kinase C activators. The ability of protein kinase C activation to open epithelial barriers to a very wide range of solutes is emphasized, but then countered with discussion of the role of phorbol esters and protein kinase C activation in epithelial carcinogenesis. The ability of protein kinase C activation to enable growth factors to leak from luminal fluid compartments of epithelial tissues into lateral intercellular and interstitial fluid spaces may play a role in this carcinogenic action. An examination of protein kinase C effects on the phosphorylation states of tight junctional proteins suggests that downstream kinases and/or phosphatases mediate protein kinase C's effect on tight junction permeability. A role for protein kinase C in transepithelial drug delivery is questioned herein. The tight junctional leakiness associated with protein kinase C activation and apparently intrinsic to transformed epithelia suggests a potentially useful role for tight junction leakiness as a marker for early cancer diagnosis. PMID- 10854688 TI - Modulation of tight junction structure and function by cytokines. AB - Dynamic regulation of tight junction function is fundamental to many physiologic processes. Disruption of tight junction function drastically alters paracellular permeability and is a hallmark of many pathologic states. Recently, an increasing number of cytokines have been shown to influence tight junction function both in vitro and in vivo. Cytokine-induced effects on tight junction barrier function have also been correlated with effects on intrinsic tight junction proteins and the associated actin cytoskeleton. The aim of this article is to review studies relating to the effects of cytokines on tight junction function and structure. PMID- 10854689 TI - Modulation of epithelial and endothelial paracellular permeability by leukocytes. AB - The tight junction acts as a regulated barrier for diffusion of ions, larger solutes, and migration of leukocytes through the paracellular space. Barrier function varies with the tissue type, and positively correlates with the number and complexity of tight junction strand formation. Polarized epithelia and brain capillary endothelium display a high degree of barrier function and tight junction formation. Conversely, vascular endothelium in other tissues has fewer, loosely organized tight junctions strands and greater permeability. Through cytoskeletal association, the tight junction and the adherens junction form a functional unit termed the apical junction complex. The development and stabilization of the tight junction is dependent upon the adherens junction. It has become apparent that paracellular permeability is altered during the migration of leukocytes across the apical junction complex and that the apical junction complex plays an important role in the regulation of leukocyte transmigration (or extravasation) through the endothelium and epithelium. Evidence suggests that important cell-cell adhesive events between transmigrating leukocytes and the apical junction complex and subsequent signaling events result in the facilitation of the passage of cells through the paracellular space. Possible mechanisms for the regulation of barrier function pertaining to leukocyte transmigration are discussed. PMID- 10854690 TI - Modulation of tight junction function by G protein-coupled events. AB - Small G proteins or GTPases comprise a growing family of signal transduction molecules with inducible properties dependent upon reversible interactions with guanine nucleotides. Activation status of the proteins is characterized by preferential affinity for triphosphorylated guanine nucleotides, initiating signaling events that control fundamental processes involved in cell migration and contraction. Termination of small G protein signaling activity is in part achieved through intrinsic GTPase activity, which catalyzes the removal of GTP and its replacement with functionally inactive GDP. Recent investigations have implicated various small G proteins as messengers that control cell-cell contact between scaffold proteins and the actin cytoskeleton, suggesting an intrinsic mechanism for the regulation of paracellular permeability in polarized epithelial and endothelial cells. This review will examine current evidence for the control of tight junction permeability by small G proteins, and speculate upon future directions that may be of value in further exploring the biological importance of these key mediators. PMID- 10854692 TI - What is a cAMP response unit? AB - Phosphoenolpyruvate carboxykinase (PEPCK) is the rate-limiting enzyme of gluconeogenesis, and most, if not all, of the regulation of its activity is exerted at the level of gene expression, with transcriptional regulation being the most predominant. A number of hormones regulate transcription of this gene, but in a defined, tissue-specific fashion. For example, cAMP strongly induces PEPCK gene transcription in liver, but provides only a weak response in kidney. Results from a number of different studies indicate that cAMP responsiveness of this gene is mediated by a 'cAMP response unit' (CRU), consisting of five cis elements. All five sequences are required for maximal responsiveness and, potentially, four of these are binding sites for a CCAAT/enhancer binding protein (C/EBP). Since alpha- and beta-isoforms of C/EBP are liver-enriched, this may provide the molecular basis for the liver-specific responsiveness to cAMP. A curiosity of this promoter is that one of the cis-elements present in the CRU is a cAMP response element (CRE), which typically acts as a binding site for CRE binding protein (CREB). However, the non-consensus CRE in the PEPCK promoter also binds C/EBP proteins with high affinity, and C/EBPalpha can functionally substitute for CREB in this cAMP response unit while C/EBPbeta cannot. The available data suggest that the PEPCK promoter can exist in altered states of cAMP responsivity, depending on which transcription factors occupy specific cis elements in the CRU. PMID- 10854693 TI - Upregulation of estrogen receptors in the forebrain of aromatase knockout (ArKO) mice. AB - Estrogens have numerous reproductive and nonreproductive functions in brain. The actions of estrogens are mediated by estrogen receptors (ERs), and estrogens are believed to down-regulate their own receptors in many tissues. Assuming this to be true, if estrogens are removed there should be an upregulation of ERs. We have developed a mouse model in which estrogen synthesis is completely eliminated by homologous recombination to delete the gene encoding aromatase cytochrome P450 (P450(arom)). The P450(arom) enzyme catalyzes the synthesis of estrogens from androgens in the brain. The localization and density of ERs was studied in the brains of aromatase knockout (ArKO) and wild type male mice by using immunohistochemistry with a peptide antibody to ERalpha (ER-21) and computer imaging. In the wild-type animals a high density of ERalpha was found in a small number of hypothalamic cells; in the medial preoptic area, periventricular, arcuate, and ventromedial nuclei. A low and medium density of ERalpha was observed in cells of the lateral preoptic area, supraoptic, bed nucleus of the stria terminalis, and in central, medial and anterior cortical amygdaloid nuclei. The number of cells containing ERalpha-immunoreactivity was significantly increased (244%) in the medial preoptic area of the ArKO mice. In neither wild type nor ArKO animals was immunoreactivity observed in the cerebral cortex or striatum. There was intense ER-immunostaining in the nucleus of neurons in both wild type and ArKO mice. These data indicate that in the absence of estrogens there is as much as a 2-fold increase in the number of cells with ERalpha immunoreactivity in certain hypothalamic and limbic regions. Thus, estrogens can down-regulate ERalpha in brain. PMID- 10854694 TI - Proglucagon cDNAs from the leopard frog, Rana pipiens, encode two GLP-1-like peptides. AB - We have isolated and characterized proglucagon cDNAs from the intestine and pancreas of the leopard frog Rana pipiens. R. pipiens proglucagon encodes glucagon, glucagon-like peptides 2 (GLP-2), and two glucagon-like peptide 1 (GLP 1) like sequences. The pancreatic and intestinal cDNAs were of identical structure and sequence suggesting that, unlike many other non-mammalian vertebrates, there is little or no alternative splicing of the proglucagon mRNA in this species. A phylogenetic analysis of the GLP-1 encoding sequences implies that the exon encoding GLP-1 was triplicated early in frog evolution, more than 150 million years ago, before the divergence of modern frogs. PMID- 10854695 TI - Identification and cellular localisation of voltage-operated calcium channels in immature rat testis. AB - Sertoli cells regulate the spermatogenic process mainly through the secretion of a complex fluid into the lumen of the seminiferous tubules behind the blood testis barrier, containing many of the essential proteins necessary for maintenance and maturation of male germ cells. Thus, the study of Sertoli cell secretory processes is strictly correlated with the understanding of the regulatory mechanisms of spermatogenesis. In this work the authors have explored the voltage-sensitive calcium channel variety in the immature rat testis, their localisation and distribution within the seminiferous epithelium and peritubular and interstitial tissues as well as the possible role in the control of Sertoli cell secretion. The results reported in this paper, obtained by in situ hybridisation, immunohistology of rat testicular sections and Western blot analysis of Sertoli cell plasma membranes, show that mammalian Sertoli cells express mRNA encoding for several voltage-operated calcium channel subunits and express such proteins on their surface. Experiments performed on Sertoli cell monolayers cultured in the presence of specific toxins indicate that both N and P/Q-type Ca(2+) channels are involved in the regulation of protein secretion. PMID- 10854696 TI - Mouse receptor-activity-modifying proteins 1, -2 and -3: amino acid sequence, expression and function. AB - The calcitonin receptor-like receptor (CRLR) requires novel receptor-activity modifying proteins (RAMPs) for its function as an adrenomedullin (ADM) or a calcitonin (CT) gene-related peptide (CGRP) receptor. Here, mouse cDNA clones representing expressed sequence tags (ESTs) in the GenEMBL database have been identified. They encode for proteins with 70, 68 and 84% amino acid sequence identity with respect to human RAMP1, -2 and -3. On Northern blot analysis of polyA(+) RNA mouse RAMP1 (mRAMP1) encoding mRNA with an apparent size of 0.8 kb was predominantly observed in embryonic and adult brain and lung and in adult skeletal muscle. Mouse RAMP2 encoding 0.8 and 1.2 kb mRNA were recognized in all tissues analyzed with the highest levels in embryonic brain, lung and gut and in adult heart, lung, skeletal muscle and brain. A single 1.2 kb mRAMP3 encoding transcript was mainly expressed in embryonic and adult brain. In COS-7 cells co expressing rat CRLR (rCRLR) and mRAMP1, [125I]halphaCGRP binding was inhibited by ralphaCGRP(8-37), ralphaCGRP and rbetaCGRP with IC(50) of 1.4+/-0.5, 4.5+/-0.6 and 7+/-0.3 nM, respectively. CyclicAMP accumulation was maximally stimulated tenfold by rbetaCGRP and ralphaCGRP with EC(50) of 0. 65+/-0.67 and 0.86+/-0.6 nM. In the same cells co-expressing rCRLR and mRAMP2, binding of [125I]rADM was displaced by rADM and rADM(20-50) with IC(50) of 1.9+/-0.5 and 3.4+/-1.4 nM, respectively, and a maximal sevenfold stimulation of cAMP accumulation was observed with rADM with an EC(50) of 0.82+/-0.85 nM. In the cells co-expressing rCRLR and mRAMP3, [125I]halphaCGRP binding was inhibited by ralphaCGRP(8-37), rbetaCGRP, ralphaCGRP, rADM and rADM(20-50) with IC(50) between 4 and 22 nM. cAMP accumulation was stimulated by rADM with an EC(50) of 5.1+/-2.7 nM that was 12 fold and 11-fold lower than that of ralphaCGRP and rbetaCGRP. In conclusion, mouse RAMP1, -2 and -3 exhibit high amino acid sequence homology to the corresponding human RAMPs. Co-expression of rCRLR with mRAMP1, -2 or -3 in COS-7 cells revealed distinct CGRP-, ADM- or ADM/CGRP receptors. PMID- 10854697 TI - Molecular characterization of myocardial fibrosis during hypothyroidism: evidence for negative regulation of the pro-alpha1(I) collagen gene expression by thyroid hormone receptor. AB - The purpose of this study was to gain insights into the underlying mechanism of myocardial fibrosis during hypothyroidism. Treatment of cardiac fibroblasts with a medium lacking thyroid hormone led to a 47% increase in [3H]thymidine incorporation into the cell nuclei compared with that in untreated cells. Northern blot analysis of RNA from cardiac fibroblasts grown in a thyroid hormone depleted medium resulted in a 38% increase in the abundance of mRNA for pro alpha1(I) collagen. At the protein level, the amount of type I collagen, as determined by immunoprecipitation, was increased either in the cell lysate (46%) of cardiac fibroblasts grown in a thyroid hormone depleted medium or in the medium (44%). The chimeric plasmid, ColCAT 3.6, contains the 5'-flanking region of the rat pro-alphal(I) collagen gene (from bases -3520 to +115) fused to the chloramphenicol acetyltransferase (CAT) gene. The plasmid was cotransfected with thyroid hormone receptor (TR) expression plasmid into rat cardiac fibroblasts and COS-l cells (monkey mesangial cells). Cells transfected with the ColCAT plasmid in the presence of thyroid hormone (100 nM T(3)) had a significant decrease (39% in fibroblasts, P<0.01; 52% in COS-1 cells, P<0.001) in CAT activity when compared to cells not exposed to thyroid hormone. Transient co-transfection of TR with various pro-alphal(I) collagen/CAT deletion constructs showed that T(3) dependent repression was preserved with the deletion from 3520 bp of the flanking sequence to a 5' end point at position -224, indicating that a thyroid hormone response element (TRE) was localized at the region -224 to +115. The TR-DNA binding assays demonstrated binding of the human TRbeta1 to a fragment containing a proposed TRE located between position -35 and +115 in the 5'-flanking region of the rat pro-alphal(I) collagen gene. PMID- 10854698 TI - A PPARgamma mutant serves as a dominant negative inhibitor of PPAR signaling and is localized in the nucleus. AB - The peroxisomal proliferator-activated receptors (PPARs) are members of the nuclear receptor superfamily that act as ligand-activated transcription factors. PPARgamma plays a critical role in regulating adipocyte differentiation and lipid metabolism. Recently, thiazolidinedione (TZD) and select non-TZD antidiabetic agents have been identified as PPARgamma agonists. To further characterize this receptor subclass, a mutant hPPARgamma lacking five carboxyl-terminal amino acids was produced (hPPARgamma2Delta500). In COS-1 cells transfected with PPAR responsive reporter constructs, the mutant receptor could not be activated by a potent PPARgamma agonist. When cotransfected with hPPARgamma2 or hPPARalpha, hPPARgamma2Delta500 abrogated wild-type receptor activity in a dose-responsive manner. hPPARgamma2Delta500 was also impaired with respect to binding of a high affinity radioligand. In addition, its conformation was unaffected by normally saturating concentrations of PPARgamma agonist as determined by protease protection experiments. Electrophoretic mobility shift assays demonstrated that hPPARgamma2Delta500 and hPPARgamma2 both formed heterodimeric complexes with human retinoidxreceptor alpha (hRXRalpha) and could bind a peroxisome proliferator-responsive element (PPRE) with similar affinity. Therefore, hPPARgamma2Delta500 appears to repress PPAR activity by competing with wild type receptor to dimerize with RXR and bind the PPRE. In addition, the mutant receptor may titrate out factors required for PPAR-regulated transcriptional activation. Both hPPARgamma2 and hPPARgamma2Delta500 localized to the nucleus of transiently transfected COS-1 cells as determined by immunofluorescence using a PPARgamma specific antibody. Thus, nuclear localization of PPARgamma occurs independently of its activation state. The dominant negative mutant, hPPARgamma2Delta500, may prove useful in further studies to characterize PPAR functions both in vitro and in vivo PMID- 10854699 TI - Expression of alternatively spliced FGF-2 antisense RNA transcripts in the central nervous system: regulation of FGF-2 mRNA translation. AB - The fibroblast growth factor-2 (FGF-2) gene is bidirectionally transcribed to produce the FGF-2 mRNA and a 1.5 kb antisense (FGF-AS) transcript complementary to the 3' untranslated region of the FGF-2 transcript. The FGF-AS RNA has been postulated to play a role in the post-transcriptional regulation of FGF-2, but this function has not been conclusively demonstrated. We characterized FGF-AS cDNAs from rat brain and C6 glioma cells, and investigated their role in regulation of FGF-2 expression. Three FGF-AS cDNAs were isolated; the full-length FGF-AS mRNA and two alternative splice variants lacking exon 2 or exons 2 and 3 of the FGF-AS sequence. The alternatively spliced FGF-AS RNAs are widely expressed in the CNS, whereas liver predominantly expressed the full-length transcript. The full-length and first splice variant encode 35 and 28 kDa isoforms of GFG, a MutT-related nuclear protein, whereas the second splice variant was not translated. The effect of FGF-AS RNA on FGF-2 expression was evaluated in stable C6 transfectants over-expressing the full-length or alternatively spliced FGF-AS RNA forms. All three constructs suppressed cellular FGF-2 protein (but not FGF-2 mRNA) levels, and this effect correlated directly with the level of FGF-AS RNA. Cellular FGF receptor content was increased and cell proliferation inhibited compared to wild type or vector-transfected cells, indicating disruption of the FGF-2 autocrine pathway by FGF-AS RNA. These findings demonstrate for the first time that the FGF-AS RNA regulates FGF-2 expression in mammalian cells, and suggest that this effect is exerted predominantly at the level of translation. PMID- 10854700 TI - Expression of luteinising hormone and chorionic gonadotropin beta-subunit messenger-RNA and protein in human peripheral blood leukocytes. AB - Some pituitary hormones are expressed in leukocytes and are thought to play a role in the regulation of leukocyte function. We studied the expression of the mRNA for the beta-chains of luteinising hormone (LHbeta) and chorionic gonadotropin (CGbeta) and their translation into protein in various leukocyte subsets. Monocytes, granulocytes, B and T-cells from peripheral blood were separated. Lymphocytes were stimulated with various mitogens, prolactin and mixed lymphocyte culture. LHbeta and CGbeta mRNA expression was determined by reverse transcriptase polymerase chain reaction. LH, LHbeta, CG and CGbeta protein were determined in the culture medium by immunofluorometric assays. LHbeta mRNA expression was detected in all cell fractions and cultures and stimulation with prolactin induced LH protein in the culture medium. CGbeta mRNA expression appeared after culture of lymphocytes, but mitogens and prolactin had no clear stimulating effect. The LH expression in leukocytes shown here suggests an autocrine function of this hormone in blood cells. PMID- 10854701 TI - Tibolone inhibits leukocyte adhesion molecule expression in human endothelial cells. AB - Tibolone is a synthetic steroid with mixed estrogenic and progestogenic/androgenic activity used for post-menopausal hormone replacement therapy. Since its cardiovascular effects are still not clear, and no data have been published on possible direct actions on the vessel wall, we studied the effects of tibolone and its metabolites on lipopolysaccharide (LPS)-induced expression of leukocyte adhesion molecules on human endothelial cells. Tibolone and its two estrogenic 3alpha-OH and 3beta-OH metabolites, but not the progestogenic/androgenic Delta(4)-isomer, concentration-dependently decreased LPS induced vascular cell adhesion molecule-1 protein expression. This effect was estrogen receptor dependent, since it was completely blocked by the pure estrogen receptor antagonist ICI 182780. Furthermore, only tibolone, the 3alpha-OH and the 3beta-OH metabolites decreased endothelial expression of E-selectin, while none of the compounds changed the levels of intercellular adhesion molecule-1. These findings were associated with parallel changes in mRNA levels for the three adhesion molecules. Our data show that tibolone and its estrogenic metabolites exert direct actions on the vascular wall, decreasing the expression of endothelial-leukocyte adhesion molecules, thus producing potentially important direct anti-atherogenic effects. PMID- 10854702 TI - Molecular cloning and properties of the chicken leptin-receptor (CLEPR) gene. AB - The mammalian leptin receptor (LEPR) (formerly OB-R) mediates the weight regulatory effects of the circulating hormone leptin. The extreme obese phenotype of recessive mutations in the mouse leptin or LEPR genes (ob/ob and db/db mice, respectively) indicate the high potential of these genes for medical and agricultural research. In this paper, we report on the cloning of the full-length chicken leptin receptor (CLEPR) cDNA, which is the first non-mammalian cloning of a LEPR gene. The CLEPR gene shares a relatively low sequence similarity with its mammalian counterparts, with an average of 60% identical nucleotides. However, comparison between the predicted protein sequences has shown a tight conservation of most previously characterized LEPR motifs and essential tyrosine residues. Similarities between the chicken and the mammalian LEPR genes were also observed in the pattern of mRNA expression. The identification of the CLEPR gene should facilitate the study of the molecular mechanism involved in the regulation of body growth and composition in avian. PMID- 10854703 TI - Immunodetection of adenylyl cyclase protein in tissues. AB - Immunodetection of the adenylyl cyclase isoforms has been difficult in tissues because of its low quantity of protein expression. We have developed a one-step cellular protein purification method that enables a simple immunodetection of the adenylyl cyclase isoforms. The type I isoform was detected exclusively in the brain. The type III isoform was detected in the brain and lungs. Further, the protein expression of type III adenylyl cyclase in lungs changed ontogenically and was the lowest in neonates. Thus, the comparison of the amount of certain adenylyl cyclase isoforms protein in each tissue is now feasible using our method. PMID- 10854704 TI - Effect of deprival of LH on Leydig cell proliferation: involvement of PCNA, cyclin D3 and IGF-1. AB - The levels of proliferating cell nuclear antigen (PCNA) and cyclin D3 which are known markers of cellular proliferation were monitored by immunoblotting in progenitor Leydig cells (PLC), immature Leydig cells (ILC) and adult Leydig cells (ALC) isolated from 21, 35 and 90 day old rats, respectively which represent the Leydig cells at different stages of development. The levels of PCNA and cyclin D3 were highest in PLC, intermediate in ILC and lowest in ALC. Following administration of an antiserum to LH to deprive endogenous LH in 21 day old rats, a significant decrease in the levels of PCNA and Cyclin D3 were observed suggesting the involvement of Lutenizing hormone (LH) in PLC proliferation. In support of this observation, Bromodeoxyuridine (BrdU) incorporation was highest in PLC when compared with ILC and ALC, and administration of LH antiserum to 21 day old rats led to a total absence of BrdU incorporation by the isolated PLC. Also, there was a decrease in the level of IGF-1 and IGF-1 receptor mRNA levels by 55 and 35%, respectively as assessed by semi-quantitative RT-PCR. In addition, the PLC isolated from rats deprived of endogenous LH incorporated much less BrdU following addition of IGF-1. These results, which are obtained using an in vivo model system establish that LH has a very important role in Leydig cell proliferation in immature rats. PMID- 10854705 TI - GH induced lipolysis stimulation in 3T3-L1 adipocytes stably expressing hGHR: analysis on signaling pathway and activity of 20K hGH. AB - We have constructed a cell line of 3T3-L1 which can efficiently express human GHR (3T3-L1-hGHR) after differentiation to adipocytes. The expressed hGHR was detected as two bands with approximate molecular sizes of 120K by Western analysis using hGHR specific monoclonal antibody. Maximum lipolytic activity induced by hGH in the 3T3-L1-hGHR was enhanced 10-fold as compared to that in 3T3 L1, suggesting that expressed hGHR is functionally active. Comparative analysis using bGH and hGH revealed that 70% of lipolysis stimulation by 1-10 ng/ml hGH could be attributed to hGHR-mediated response. Analyses on inhibition and phosphorylation of signaling molecules suggested that GH-induced lipolysis stimulation is dependent on gene expression and not mediated through PKA-, PKC-, PLA-, PLC-, nor MAPK-pathway but possibly through JAK-STATs pathway. Duration of STAT5 activation by hGH continued up to 48 h. We also revealed that 22 K hGH isoform, 20K hGH which has been reported as a weaker agonist for GH-induced lipolysis stimulation, possesses equipotent activity and shows stronger action in the presence of hGHBP as compared to 22 K hGH. Taken together we conclude that the hGH-induced lipolysis was not mediated through MAP-, PKA-, PKC-, nor PLA pathway but might be mediated through STAT pathway and that 20K hGH might show higher lipolytic activity than 22 K hGH in adipose tissue that produces a large amount of GHBP. PMID- 10854706 TI - Development of a human stanniocalcin radioimmunoassay: serum and tissue hormone levels and pharmacokinetics in the rat. AB - Stanniocalcin (STC) is a polypeptide hormone that was first discovered in fish and recently identified in humans and other mammals. In fish STC is produced by one gland, circulates freely in the blood and plays an integral role in mineral homeostasis. In mammals, STC is produced in a number of different tissues and serves a variety of different functions. In kidney, STC regulates phosphate reabsorption by proximal tubule cells, whereas in ovary it appears to be involved in steroid hormone synthesis. However there is no information on circulating levels of STC in mammals or the regulation of its secretion. In this report we have developed a radioimmunoassay (RIA) for human STC. The RIA was validated for measuring tissue hormone levels. However human and other mammalian sera were completely devoid of immunoreactive STC (irSTC). To explore the possibility that mammalian STC might have a short half-life pharmacokinetic analysis was carried out in rats. STC pharmacokinetics were best described by a two compartment model where the distribution phase (t1/2(alpha)) equaled 1 min and the elimination phase (t1/2(beta)) was 60 min. However the STC in the elimination phase no longer crossreacted in the RIA indicating it had undergone substantial chemical modification, which could explain our inability to detect irSTC in mammalian sera. When we compared the pharmacokinetics of human and fish STC in mammalian and fish models the human hormone was always eliminated faster, indicating that human STC has unique structural properties. There also appears to be a unique clearance mechanism for STC in mammals. Hence there are major differences in the delivery and biology of mammalian STC. Unlike fishes, mammalian STC does not normally circulate in the blood and functions instead as a local mediator of cell function. Future studies will no doubt show that this has had important ramifications on function as well. PMID- 10854707 TI - Expression of leukemia inhibitory factor (LIF) and LIF receptor (LIF-R) in the human adrenal cortex: implications for steroidogenesis. AB - It is well established that steroidogenesis in the adrenal cortex is regulated by extraadrenal factors, such as ACTH and angiotensin II. However, over the last years, it has become increasingly clear that paracrine and autocrine mechanisms are also important for steroid synthesis in the adrenal gland. The current study was designed to analyze whether the pleiotropic cytokine leukemia inhibitory factor (LIF) and/or its receptor (LIF-R) are expressed in the normal human adrenal cortex, and whether they may play a role in regulating steroidogenesis. Using LIF- and LIF-R-specific primers, we show by RT-PCR that both mRNAs are expressed in this tissue, as well as in the NCI-H295 adrenal carcinoma cell line. The correct sequences of the PCR products were verified by restriction enzyme analysis and DNA sequencing. Immunohistochemistry, employing specific antibodies against LIF and LIF-R, reveals expression of both proteins in the normal human adrenal cortex. Finally, we show that LIF can significantly enhance basal and ACTH-induced production of cortisol and aldosterone in NCI-H295 cells. In summary, we show for the first time that LIF and its receptor are expressed in the normal human adrenal cortex. Our stimulation experiments indicate that the intraadrenal LIF/LIF-R system may participate in regulating adrenal steroidogenesis. PMID- 10854708 TI - Early induction of the orphan nuclear receptor NOR-1 during cell death of the human breast cancer cell line MCF-7. AB - The neuron-derived orphan receptor (NOR-1) is a member of the NGFI-B subfamily within the nuclear receptor superfamily. In T-cell apoptosis, where NGFI-B plays an essential role, a functional redundancy between NGFI-B and NOR-1 has been demonstrated. Here, we examined the regulation and expression of the NOR-1 gene during cell death induced by a calcium ionophore A23187 in the human breast cancer cell line MCF-7. A23187 caused a transient increase in NOR-1 mRNA levels within 6 h after treatment. To delineate the sequences required for the transitional response to A23187, a series of promoter deletion mutants were constructed. From the transient transfection experiments, the element responsive to A23187 was identified between -94 and -42 base pairs upstream from the transcription initiation site. This 53-base pairs region contains three copies of the cAMP response element (CRE). Furthermore, phosphorylation of the CRE-binding protein (CREB), which affects the transcription of the CRE dependent-genes, was detected 30 min after A23187 stimulation. Our findings are consistent with NOR-1 involvement in A23187-induced cell death via the CRE-CREB signaling pathway. PMID- 10854709 TI - Differential expression of human gonadotropin-releasing hormone receptor gene in pituitary and ovarian cells. AB - In terms of regulation of gene expression, gonadotropin-releasing hormone receptor (GnRHR) found in extrapituitary tissues has been suggested to be different from that in the pituitary. In the present study, we examined the molecular basis of this difference using the pituitary alphaT3-1 and ovarian carcinoma OVCAR-3 cells. As a first step, the different expression levels of GnRHR mRNA in the pituitary and ovarian cells were investigated using semi quantitative RT-PCR. Quantitative analysis showed that the expression level of hGnRHR is a nine-fold higher in primary pituitary tissues than the primary culture of ovarian carcinomas (PCO). In pituitary alphaT3-1 cells, the expression level of hGnRHR was ten-fold higher than ovarian carcinoma OVCAR-3 cells. The possibility of the differential use of various cell-specific promoters in different cells was addressed by transiently transfecting cells with 3'-deletion clones of human GnRHR promoter. Sequential deletion of the 5'-flanking region of the gene revealed the use of two putative promoters, designated PR1 (-771 to 557) and PR2 (-1351 to -1022), and a negative control region (-1022 to -771), in the pituitary and ovarian cells. The same promoters appeared to be utilized for driving the basal promoter activities in both alphaT3-1 and OVCAR-3 cells, suggesting that there is no cell-specific promoter usage for the human GnRHR gene. Alternatively, the involvement of different regulatory protein factors was investigated using electrophoretic gel mobility shift assays. When end-labeled PR1 was used as a probe, two unique shifted complexes were identified in OVCAR-3 cells when compared to alphaT3-1 cells. One unique protein-DNA complex was observed in alphaT3-1 cells compared to OVCAR-3 cells when incubated with end labeled PR2 as a probe. These DNA-protein complexes appeared to be specific, as they competed with excess amount of unlabelled competitor PR1 and PR2, respectively. In summary, it is unlikely that the use of a cell-specific promoter contributes to the different characteristics of ovarian GnRHR. Our study demonstrates that one mechanism by which cell-specific expression of human GnRHR is achieved is through the binding of distinct and/or combinations of cell specific regulatory factors to various promoter elements in the 5'-flanking region of the gene. PMID- 10854710 TI - Expression of an estrogen receptor alpha variant protein in cell lines and tumors. AB - Human estrogen receptor alpha (ER) mRNA is a mixture of wild type and alternatively spliced variants. Many studies have examined the potential of ER mRNA profiles to serve as diagnostic/prognostic cancer biomarkers, but only a few have attempted to correlate ER mRNA profiles with protein expression. Representative ER mRNA pools were reproduced from the cDNAs of MCF-7 cells, a human breast tumor and human uterus and translated in a protease-free environment by reticulocyte lysates to determine relative translation efficiencies between the various ER mRNA transcripts and to facilitate identification of translated proteins. Cell line and tumor extracts were then examined for expression of the ER variant proteins identified in reticulocyte lysate translations. Each of the ER mRNA pools were translated by reticulocyte lysates into two ER proteins with molecular weights of approximately 60 and 52 kD. Western immunoblotting with various C- and N-terminal-directed, anti-ER antibodies and comparison with expressed ER protein standards established that the 52 kD protein (ERDelta7P) was translated from the predominant splice variant mRNA in each pool, which is missing exon 7. The 60 kD protein contained wild type ER sequence minus 61 C terminal amino acids lost due to an intentional run off truncation. ERDelta7P expression was subsequently demonstrated in MCF-7 cells by Western immunoblotting with the site-directed antibodies. A protein corresponding to ERDelta7P was also detected in other ER positive breast tumor cell lines, and extracts of ER positive breast and uterine tumors. This widespread expression of ERDelta7P in vivo suggests that it may have some biological function. ERDelta7P may also affect immunohistochemical evaluation of ER positivity in tumors depending upon the level of its expression and the antibody used. PMID- 10854711 TI - Regulation of PKC delta expression by estrogen and rat placental lactogen-1 in luteinized rat ovarian granulosa cells. AB - Protein kinase C (PKC) delta is dramatically upregulated in the corpus luteum in the second half of pregnancy in the rat. To gain insight into the hormonal regulation of PKC delta expression, studies were undertaken to analyze the regulation of PKC delta expression in a luteinized rat granulosa cell model. PKC delta protein expression was evaluated in luteinized granulosa cells, isolated from human (h)CG-treated immature female rats 7 h after the injection of an ovulatory dose of hCG and cultured up to 12 days. Cytochrome P450 cholesterol side chain cleavage enzyme expression was observed throughout the culture period, and a majority of the cells expressed steroidogenic acute regulatory protein and responded to rat placental lactogen (rPL)-1 by exhibiting hypertrophy, consistent with maintenance of the luteal phenotype. Both PKC delta protein and mRNA expression increased 3.5-4-fold with time of culture, and PKC delta mRNA expression could be eliminated by treatment of cells with the PKC inhibitor GF109203X. E(2) caused a specific dose- and time-dependent increase in expression of PKC delta protein of twofold, whereas PKC delta mRNA was unaffected by E(2) over a 12-day culture period. Treatment of cells with 500 ng/ml rPL-1 for the final 4 days of a 12-day culture in the absence of E(2) had no effect on PKC delta protein or mRNA expression, while treatment with 500 or 3000 ng/ml rPL-1 in the presence of E(2) significantly enhanced both PKC delta protein and mRNA expression (up to threefold). These results show that two of the major regulators of luteal function in the second half of pregnancy in the rat, E(2) and rPL-1, cooperate to regulate PKC delta expression in luteinized granulosa cells. PMID- 10854712 TI - Glucocorticoid receptors in the euryhaline teleost Anguilla anguilla. AB - To determine the importance of glucocorticoids in the salt water adaptation of European yellow eel we have evaluated the concentration, affinity and physical properties of glucocorticoid receptors (GR) in gill from both sea water- (SW) and freshwater-adapted (FW) animals. Using ligand binding techniques we demonstrated that high affinity GR were present in both cytosolic and nuclear fractions obtained from whole gill. Isoelectric focusing (IEF) of branchial GR indicated the presence of two distinct species, with pI values of 6.1 and 6.7. The form at pI 6.7 sedimented with a Svedberg constant of 4S on glycerol density gradients while the pI 6. 1 sedimented in fractions corresponding to 9S. Treatment of the pI 6. 1 form with urea (4 M) resulted in generation of the form with pI 6. 7. The evidence thus suggested that the oligomeric urea-sensitive form (pI 6.1) contained a form of GR which, as a monomer, focused at pI 6.7. IEF revealed the same concentrations of the pI 6.7 form in both SW and FW. However, there was significantly more (3-fold) pI 6. 1 isoform in FW than in SW, and this form decreased gradually during the course of seawater transfer. A transient increase of the nuclear-bound GR was also observed during SW adaptation. The balance between these forms could represent a dynamic parameter with important implications regarding GR function and gill responses to cortisol in salt water adaptation in teleosts. PMID- 10854713 TI - Volume-sensitive amino acid efflux from a pancreatic beta-cell line. AB - Cell swelling, induced by a hyposmotic shock, increased the fractional release of taurine from INS-1 cells. Volume-sensitive taurine release was (a) dependent upon the extent of cell swelling; (b) fully reversible; and (c) temperature dependent. Volume-sensitive taurine efflux was independent from the trans-membrane Na(+) gradient. DIDS markedly inhibited volume-activated taurine efflux but not basal taurine release suggesting that the volume-sensitive pathway is quiescent under isosmotic conditions. Volume-activated taurine release inactivated in the continued presence of a hyposmotic shock. Cell-swelling also increased the fractional release of D-aspartate from INS-1 cells. Volume-activated D-aspartate efflux was inhibited by DIDS, albeit to a lesser extent than volume-sensitive taurine release. It is predicted that volume-sensitive amino acid efflux acts in parallel with other volume-activated transport mechanisms to regulate the volume of insulin-secreting cells. PMID- 10854714 TI - Ligand structure-dependent differences in activation of estrogen receptor alpha in human HepG2 liver and U2 osteogenic cancer cell lines. AB - Differences in ligand-activation of estrogen receptor alpha (ER(alpha)) were investigated in human HepG2 liver carcinoma and U2 osteogenic sarcoma cells transfected with wild-type ER (ER-wt) and variants expressing only activation function 1 (ERAF1) or AF2 (ER-AF2). The estrogen-responsive C3-luc construct containing the complement C3 gene promoter linked to a bacterial luciferase reporter gene was used to determine ligand-induced wild-type or variant ER activation. The quality pattern of ER-dependent responses was similar in both cell lines for a series of weakly estrogenic hydroxy and dihydroxyaromatic compounds including p-octylphenol, p-nonylphenol, 2',4',6'-trichloro-4 biphenylol, 2',3',4', 5'-tetrachloro-4-biphenylol, bisphenol A and 2, 2'-bis(p hydroxyphenyl)-1,1,1-trichloroethane. However, some significant quantitative differences in these compounds were also observed. The weakly estrogenic pesticide, kepone, and the phytoestrogens, resveratrol (a trihydroxystilbene) and naringen (a flavanone), induced distinctly different patterns of responses; induction by these compounds was not observed in either cell line cotransfected with ER-wt or ER-AF2. In contrast, naringen activated ER-AF1 in HepG2 cells and resveratrol activated ER-AF1 in U2 cells. In HepG2 cells cotreated with E2 plus the estrogenic compounds, only BPA and resveratrol exhibited ER(alpha) antagonist activity. Structure-dependent differences in ER(alpha) activation and inhibition are consistent with the increasingly complex patterns of ER action in various tissues and indicate that the estrogenic activity of an individual compound can only be determined by using an extensive testing protocol. PMID- 10854715 TI - Ability of the glucocorticoid modulatory element to modify glucocorticoid receptor transactivation indicates parallel pathways for the expression of glucocorticoid modulatory element and glucocorticoid response element activities. AB - The glucocorticoid modulatory element (GME) of the rat tyrosine aminotransferase gene is located at -3.6 kb and 1 kb upstream of the glucocorticoid response elements (GREs). The GME has the unique transcriptional properties of modulating both the dose-response curve of agonists bound to the glucocorticoid receptor (GR) and the residual agonist activity of GR-bound antisteroids. The expression of GME activity involves the binding of two novel proteins (GMEB-1 and GMEB-2) that we have recently cloned. However, the mechanistic details are limited. The DNA sequence requirements for GME activity (CGTC) also remain poorly defined, which restricts efforts to identify other GME modulated genes. To help understand the mechanism for the unusual activities of the GME and to identify permissive gene environments for GME activity, we compared the changes in GME activity and GRE action (i.e. the fold induction by GR) caused by modifying several parameters. Phasing between the GME and downstream tandem GREs was unimportant, in contrast to other cis-acting elements like the GRE, while GME activity decreased rapidly when placed at increasingly larger distances 3' to a tandem GRE. A minimal promoter was less effective in supporting GME than GRE activity. Although CREB binds to the GME, overexpression of CREB reduced GRE, but not GME, activity and a CRE was inactive when substituted for the GME. No effect of the GME was observed on the binding of GRs to a single GRE. However, the GME upstream of a single GRE was also unable to produce a left shift in the Dex dose-response curve under conditions where the GME was active with two GREs. In the absence of any GREs, the GME displayed intrinsic activity by elevating basal level expression. Collectively, these results indicate that an optimal position for a functional GME is within 250 bp upstream of a tandem GRE driving a complex promoter. Furthermore, as the changes in GME activity did not correlate with those for fold induction from the GRE, the mechanisms for expression of GME and GRE activities appear to utilize parallel, as opposed to common pathways. PMID- 10854717 TI - Olanzapine, quetiapine, and risperidone: long-term effects on monoamine transporters in rat forebrain. AB - Long-term effects of novel atypical antipsychotic drugs on monoamine transporters are unknown. We compared labeling of dopamine (DAT) and serotonin (SERT) transporter proteins in subregions of rat corpus striatum by quantitative autoradiography with [(3)H]2-beta-carbomethoxy-3-beta-[4'-iodophenyl]tropane ([(3)H]beta-CIT) and [(3)H]paroxetine after 28 days of continuous subcutaneous infusion of olanzapine, quetiapine, risperidone, or vehicle controls. Drug treatment did not significantly alter the abundance of either transporter type in caudate-putamen or nucleus accumbens, indicating that transporter proteins required to inactivate synaptically released dopamine and serotonin resist adaptations to long-term treatment with novel antipsychotics that affect neurotransmission by these amines. PMID- 10854718 TI - Denervation facilitates neuronal growth in the motor cortex of rats in the presence of behavioral demand. AB - This study tests the hypothesis that degeneration of a neocortical pathway may facilitate behaviorally-induced growth of neurons in a connected region of the cortex. Degeneration of trancallosal afferents to the motor cortex and changes in forelimb use were independently manipulated in adult rats. The combination of degeneration and behavioral change resulted in the growth of layer V pyramidal neuron dendrites which was not found as a result of either denervation or behavioral manipulation alone. These results indicate that mild degeneration in the adult brain can facilitate neuronal growth when accompanied by appropriate behavioral demand, a finding which has implications for rehabilitative therapy after brain damage. PMID- 10854719 TI - Effects of vagal cooling on esophageal cardiovascular reflex responses in the rat. AB - Distension of the distal esophagus in the anesthetized rat causes a vagally mediated arterial pressor and tachycardia response. To investigate the nature of viscerosensory fibers in the afferent limb of this reflex, the present study was carried out in urethane-anesthetized rats that were subjected to graded cooling of both cervical vagal trunks in situ. Distal esophageal distension was applied for 20 s by means of a water-filled high compliance balloon. Vagal cooling to 9 degrees C abolished pressor responses and unmasked a depressor component during maximal distension. Cooling to 7.5 degrees C blocked this inhibitory component, well above the temperature known to block C-fibers. We conclude that the cardiovascular response to esophageal distension is triggered via at least two subpopulations of A(delta) type vagal afferents that project to brain stem nuclei regulating central vasomotor tone. PMID- 10854720 TI - Functional coupling of human right and left cortical motor areas demonstrated with partial coherence analysis. AB - Although a linear correlation between oscillatory activities in the right and left motor cortices during movements has been shown in monkeys, there has been a debate whether scalp-recorded EEG coherence in human reflects a similar association. By applying partial coherence analysis, we demonstrated that interhemispheric coherence during movements cannot be explained by contamination from the occipital alpha rhythm or common reference signal. A significant increase of net interhemispheric communication in the beta1 band was shown during movements. We propose that the partial coherence method can be a useful tool to measure cortico-cortical functional coupling reliably. PMID- 10854721 TI - Suramin protects the murine motor nerves from the toxic effects of presynaptic Ca(2+) channel inhibitors. AB - The purpose of this study is to investigate whether suramin is capable of preventing the neurotoxic effects of Ca(2+) channel inhibitors at the presynaptic sites. Mouse diaphragm and triangularis sterni preparations were used for this study in order to measure the muscle tension and nerve terminal Ca(2+) current, respectively. Both omega-conotoxin MVIIC and omega-agatoxin IVA markedly inhibit the nerve-evoked muscle contractions as well as the nerve terminal Ca(2+) current respectively. Pretreatment with suramin (0.3 mM) significantly reduced the inhibitory effect of nerve-evoked muscle contractions and Ca(2+) current induced by either omega-conotoxin MVIIC or omega-agatoxin IVA but not that induced by the non-selective Ca(2+) channel blocker, Cd(2+). Neither suramin nor Ca(2+)-channel toxins significantly affect Na(+)- and K(+) currents of the nerve terminals. These findings indicate that suramin selectively interferes the action of presynaptic Ca(2+) channel neurotoxins and thus reduces their depressant effects on the muscle contractions. The implication of these findings is that suramin and its derivatives may potentially become useful agents in management of intoxication of Ca(2+) channel neurotoxins. PMID- 10854722 TI - Presence of unmyelinated axons in the lumbar ventral roots of the 129 mouse strain. AB - The 129 mouse strain has become of increasing interest to neurobiologists due to its importance in gene targeting studies. However it has been pointed out that 129 mice suffer from a number of neuroanatomical idiosyncrasies that may make them less attractive as animal models in neurobiology. Here we show that 129 mice also differ from other commonly used strains in possessing large numbers of unmyelinated axons in their lumbar motor roots. By contrast in all other strains of mice (C57BL/6, C3H, Swiss-Webster) that we studied the axons in the L5 roots are all myelinated. Additionally we show that 129 mice have smaller myelinated axons than other mouse strains and perform poorly in the rotorod test. These characteristics must be kept in mind in studies of mutant mice that are frequently performed on a mixed genetic background containing a129 contribution. PMID- 10854723 TI - Altered hippocampal expression of neuropeptide Y, somatostatin, and glutamate decarboxylase in Ihara's epileptic rats and spontaneously epileptic rats. AB - By in situ hybridization and immunocytochemistry, expression of neuropeptide Y (NPY), somatostatin and glutamate decarboxylase 65 (GAD65) was studied in the hippocampus of two different epileptic mutant rats, Ihara's epileptic rat (IER) and the spontaneously epileptic rat (SER). GAD65 mRNA expression was enhanced in interneurons of the hippocampus in young IER, that had not yet developed generalized seizures. In older IER and older SER that both showed spontaneous seizures, marked increases of NPY mRNA in hippocampal granule cells and interneurons were found, as well as elevated GAD65 mRNA levels in interneurons. NPY immunoreactivity was enhanced in hilar interneurons and the dentate gyrus of older IER. In addition, some older IER stained heavily for NPY in mossy fibers. These findings suggest that up-regulation of NPY and GAD65 synthesis may be important in epileptogenesis. PMID- 10854724 TI - Human post-mortem striatal alpha4beta2 nicotinic acetylcholine receptor density in schizophrenia and Parkinson's syndrome. AB - The density of nicotinic alpha4beta2 receptors, which are believed to largely mediate nicotine's effects, has been reported to be decreased in post-mortem hippocampus of patients with schizophrenia. In the present study, using [(3)H]cytisine as a radioligand, we observed a significant 30% decrease in post mortem striatum of patients with schizophrenia (n=12) as compared to controls (n=12). A 25% decrease of striatal alpha4beta2 receptor density in patients with Parkinson's syndrome (n=12) was not significant. As an upregulation of alpha4beta2 receptors has been observed due to nicotine consumption, the beneficial effects of nicotine described in patients with schizophrenia may be partly due to a compensation for a decrease in alpha4beta2 nicotinic acetylcholine receptors. PMID- 10854725 TI - Effects of naloxone on lactate, pyruvate metabolism and antioxidant enzyme activity in rat cerebral ischemia/reperfusion. AB - Whether naloxone may modulate energy metabolism and endogenous antioxidant enzyme activities in ischemic cortex was studied. Cerebral ischemia/reperfusion (I/R) was produced by occluding two common carotid arteries and the right middle cerebral artery for 90 min followed by reperfusion in anesthetized Sprague-Dawley rats. Both pre-treatment (0.03 or 0.3 mg) and post-treatment (0.3 mg) of naloxone by intracerebroventricular infusion significantly reduced cortical infarct volumes. Pre-treatment with 0.03 mg reduced ischemia-induced suppression of extracellular pyruvate level and enhancement of lactate/pyruvate ratio as well as cerebral I/R-induced increases of endogenous catalase, glutathione peroxidase, and manganese superoxide dismutase activities. In conclusion, neuroprotective effects of naloxone in terms of reducing brain infarction involve attenuation of the disturbance of cellular functions following cerebral I/R via restoration of mitochondrial activities or energy metabolism. PMID- 10854726 TI - Striatal glutamate release during novelty exposure-induced hyperactivity in olfactory bulbectomized rats. AB - Striatal glutamate release during novelty exposure-induced hyperactivity was studied by microdialysis in freely-moving olfactory bulbectomized (OBX) rats. After collecting three 10 min basal striatal dialysate samples, the animals were transferred to an open-field apparatus (novelty) and locomotor activity recorded for 60 min. OBX rats showed significantly more locomotor activity (1210+/-270 cm) than sham-operated rats (420+/-70 cm), but only in the first 10 min after exposure to the novel environment. During the same period, striatal glutamate levels increased to 163+/-21% of the basal value in OBX rats, while no changes were seen in the striatum of sham-operated controls. These findings suggest that olfactory bulbectomy results in an increased response of the striatal glutamatergic system to novelty stress, and may consequently cause hyperactivity. PMID- 10854727 TI - Microglial proliferation in the spinal cord of aged rats with a sciatic nerve injury. AB - Nerve injury may lead to chronic neuropathic pain syndromes. We determined whether the extent of central nervous system microglial activation that accompanies nerve injury is age dependent and correlated with behavioral manifestations of pain. We used the Bennett and Xie sciatic nerve chronic constriction injury model (Bennett, G.J., Xie, Y.-K., A peripheral mononeuropathy in rat that produces disorders of pain sensation like those seen in man, Pain, 33 (1998) 87-107) to induce neuropathic pain in three age cohorts of Fischer 344 FBNF1 hybrid rats (4-6, 14-16, and 24-26 months). Rats were assessed for thermal sensitivity (hyperalgesia) of their hind paws pre-injury (day 0) and up to 35 days post injury. On various days post injury, the L4-L5 levels of their spinal cords were reacted for localization of an antibody to OX-42, a marker for microlgia. OX-42 immunoreactivity (ir) was quantified by use of a Bioquant density analysis system. OX-42 ir was heavy in areas of sciatic nerve primary afferent terminations and in the motor columns of its neurons. Aging increases OX 42 ir in the absence of injury. After injury, OX-42 ir increased further, but the increases over control levels decreased with age. Ligation-induced analgesia and hyperalgesia were both correlated with the increases in OX-42 ir, regardless of age. PMID- 10854728 TI - Changes in the number of nitric oxide-synthesizing neurones on both sides of a chronic transection of the rat spinal cord. AB - Pain after chronic transection of the spinal cord is hypothesized to develop because of a hyperactivity in nociceptive neurones rostral to the lesion. One of the key substances in central nervous nociceptive processing is nitric oxide (NO). It has been demonstrated to tonically inhibit the background activity of dorsal horn neurones. Here, we show that in rats with chronic transection of the spinal cord there is a reduction of NO-synthesizing neurones on both sides of the lesion. This reduction is likely to be associated with a local lack of NO which could lead to an increased background activity of nociceptive dorsal horn neurones. The increased background activity of nociceptive neurones just rostral to the lesion might cause spontaneous pain that is perceived in segments close to the level of the lesion. PMID- 10854729 TI - M(2)/M(4)-muscarinic receptors mediate automodulation of acetylcholine outflow from mouse cortex. AB - Acetylcholine outflow can be modulated through inhibitory presynaptic muscarinic autoreceptors. This study was to identify which subtype is involved in mouse cortex. Five muscarinic antagonists and their ability to elevate stimulation induced (S-I) acetylcholine outflow were tested in the presence of neostigmine, which decreased S-I outflow. The potency of each antagonist was determined, expressed as a ratio of the potency of each other antagonist and compared with the potency ratios of the antagonists for each of the defined muscarinic receptors (M(1)-M(4)), as recorded in the literature. Linear regression analysis revealed that the data fitted the M(2) (r(2)>0.97) and M(4) (r(2)>0.85) subtypes best, with no correlation for the M(1) and M(3) subtypes. PMID- 10854730 TI - Neuropsychological evidence that somatic stimuli are spatially coded according to multiple frames of reference in a stroke patient with tactile extinction. AB - A right-brain damaged patient with pure tactile extinction was asked to report series of single or double light, brief touches delivered to both hands, the thumb or the pinkie of a single hand, the sides of a single index. The stimulated hand was positioned palm up or palm down, in front of or behind the patient, in anatomic or crossed position. In double touch conditions, stimuli coded as left sided were extinguished not only when delivered to both hands, but also when delivered on a single hand or a single finger. The findings suggest that tactile stimuli may be coded as left or right according to multiple body anchors that are dynamically scaled from the corporeal midline, to axes centered on a single hand or finger. PMID- 10854731 TI - A computer simulation of recurrent, excitatory networks of sensory neurons of the gut in guinea-pig. AB - Intrinsic sensory neurons of the intestine are connected together to form a recurrent network. They interact by slow excitatory post synaptic potentials (EPSPs), which have a complex dependence on the pattern of input. These networks are unstable and unable to give graded responses to sensory input without some form of inhibition, but inhibitory synaptic potentials are rare in this system. Intrinsic sensory neurons have a characteristic after-hyperpolarization (AHP), but this is depressed during slow EPSPs. To test whether AHPs can provide the inhibition necessary for stability, AHPs, slow EPSPs and their interactions were included in a computer simulation of realistic sensory neuron networks. Residual AHPs as small as 1% of control were found to lead to stable networks capable of giving graded responses. PMID- 10854732 TI - The alpha3 subtype of the nicotinic acetylcholine receptor is expressed in airway related neurons of the nucleus tractus solitarius, but is not essential for reflex bronchoconstriction in ferrets. AB - To assess the role of nicotinic acetylcholine receptors (nACh-R) in the transmission of afferent constricting inputs from bronchopulmonary receptors to the nucleus tractus solitarius (nTS) and in the mediation of reflex airway constriction, we performed a combined immunohistological and functional study. In ferrets, the expression of nAch-R on the nTS neurons activated by histamine stimulation of airway sensory receptors was studied using laser scanning confocal microscopy to co-immunolocalize c-fos encoded protein (cFos) and nACh-R alpha3 subunit. We observed that activation of airway sensory receptors by inhalation of aerosolized histamine, induced cFos expression in a subset of nTS neurons that also expressed the nAch-R alpha3 subtype. Furthermore, activation of nACh-R within the commissural subnucleus by nicotine, increased cholinergic outflow to the airways. These effects were diminished by prior administration of hexamethonium (nACh-R blocker) within the commissural subnucleus of the nTS. However, hexamethonium had no significant effects on airway reflex constrictions induced by lung deflation. These findings indicate that nACh-R are expressed by the nTS neurons receiving inputs from airway sensory receptors, activation of which by nicotine increases cholinergic outflow to the airways, but the nACh-R pathways are not required for reflex bronchoconstriction. PMID- 10854733 TI - Human rotation-mediated fetal mixed brain cell aggregate culture: characterization and N-methyl-D-aspartate toxicity. AB - One difficulty in generating in vitro models of neuropathogenesis lies in maintaining stable proportions of primary neurons within a mixed brain cell population. Rotation-mediated fetal brain aggregate culture has been modified to permit growth of human primary fetal brain cells containing 50 to 60% neurons. After 12 weeks cholinesterase, neuron specific enolase and microtubule-associated protein-2 were demonstrable by biochemical assay and immunocytochemical labelling of cryostat sections of human fetal brain aggregates. Upon exposure to the glutamate agonist; N-methyl-D-aspartate for 7 days at 35 days in vitro neuron specific enolase and cholinesterase decreased to 60 to 70% of untreated levels. Glial fibrillary acidic protein did not change significantly but swollen astrocytes were seen in labelled sections of treated aggregates. This method is useful to study human neurotoxicity and degeneration in mixed glial culture without astrocyte overgrowth. PMID- 10854734 TI - Long-term potentiation across rat cerebello-thalamic synapses in vitro. AB - We previously demonstrated that rat cerebello-thalamic synapses undergo an ultrastructural change, consistent with expression of long-term potentiation, in association with motor adaptation. The aim of these experiments was to determine if long-term potentiation can be expressed at these synapses physiologically. Excitatory post-synaptic potentials evoked by electrical stimulation of cut cerebello-thalamic afferents were recorded intracellularly from thalamic neurons in 17-24 day old rat brain slice preparations. The experimental protocol involved long periods of low frequency single-shock control stimuli interrupted by brief high frequency trains of conditioning stimuli. Conditioning at 100 Hz evoked long term potentiation in 5/6 cells and long-term depression in 1/6 cells. Conditioning at 200 Hz was unsuccessful in evoking long-term potentiation but did evoke long-term depression in 1/3 cells and no change in 2/3 cells. We conclude that long-term potentiation can be evoked across rat cerebello-thalamic synapses in vitro. PMID- 10854735 TI - A functional magnetic resonance imaging study of the role of left posterior superior temporal gyrus in speech production: implications for the explanation of conduction aphasia. AB - Conduction aphasia, characterized by good auditory comprehension and fluent but disordered speech production, is classically viewed as a disconnection syndrome. We review recent evidence which suggests that at least one form of conduction aphasia results from damage to cortical fields in the left posterior superior temporal gyrus which participate not only in speech perception, but also in phonemic aspects of speech production. As a test of this hypothesis, we carried out a 4T functional magnetic resonance imaging study in which subjects named visually presented objects sub-vocally. Group-based analyses showed that a majority of participants showed activation in two regions on the dorsal portion of the left posterior superior temporal gyrus. PMID- 10854736 TI - The norbornenyl moiety of cyclothiazide determines the preference for flip-flop variants of AMPA receptor subunits. AB - Cyclothiazide and two analogs in which the norbornenyl part was replaced with a cyclohexyl or a cyclohexenyl moiety were examined with regard to their preference for flop vs. flip splice variants of the (+/-)-alphaamino-3-hydroxy-5 methylisoxazole-4-propionic acid (AMPA) receptor subunits GluR2, 3 and 4. The studies were carried out by measuring the effects of the drugs on the binding of [(3)H]AMPA or [(3)H]fluorowillardiine to membranes from HEK293 cells that stably express the AMPA receptor subunits. Cyclothiazide had four to nine times lower EC(50) values at flip than at flop receptors, as previously reported. In contrast, the two analogs showed little discrimination for GluR3 or GluR4 splice variants and a clear preference for the flop variant in the case of GluR2. These results indicate that it is the norbornenyl component of cyclothiazide which confers the selectivity vis-a-vis flip-flop variants. PMID- 10854737 TI - Chemical ecology of host-plant selection by herbivorous arthropods: a multitrophic perspective. AB - Most herbivorous arthropods are specialists that feed on one or a few related plant species. To understand why this is so, both mechanistic and functional studies have been carried out, predominantly restricted to bitrophic aspects. Host-selection behaviour of herbivorous arthropods has been intensively studied and this has provided ample evidence for the role of secondary plant chemicals as source of information in behavioural decisions of herbivores. Many evolutionary studies have regarded co-evolution between plants and herbivores to explain the diversity of secondary plant chemicals and host specialisation of herbivores. However, many cases remain unexplained where herbivores select host plants that are suboptimal in terms of fitness returns. A stimulating paper by Bernays and Graham [(1988) Ecology 69, 886-892)] has initiated a discussion on the need of a multitrophic perspective to understand the evolution of host-plant specialisation by herbivorous arthropods. However, this has hardly resulted in ecological studies on host-selection behaviour that take a multitrophic perspective. Yet, evidence is accumulating that constitutive and induced infochemicals from natural enemies and competitors can affect herbivore behaviour. These cues may constitute important information on fitness prospects, just as plant cues can do. In this paper I selectively review how information from organisms at different trophic levels varies in space and time and how herbivores can integratively exploit this information during host selection. In doing so, research areas are identified that are likely to provide important new insights to explain several of the questions in herbivore host selection that remain unanswered so far. These research areas are at the interface of evolutionary ecology, behavioural ecology and chemical ecology. PMID- 10854738 TI - Phenolic glycosides and condensed tannins in Salix sericea, S. eriocephala and their F1 hybrids: not all hybrids are created equal. AB - The performance of hybrids depends upon the inheritance and expression of resistance traits. Secondary chemicals are one such resistance trait. In this study, we measured the concentrations of phenolic glycosides and condensed tannins in parental and F1 hybrid willows to examine the sources of chemical variation among hybrids. S. sericea produces phenolic glycosides, salicortin and 2'-cinnamoylsalicortin, and low concentrations of condensed tannin in its leaves. In contrast, S. eriocephala produces no phenolic glycosides but high concentrations of condensed tannins in its leaves. These traits are inherited quantitatively in hybrids. On average, F1 hybrids are intermediate for condensed tannins, suggesting predominantly additive inheritance or balanced ambidirectional dominance of this defensive chemical from the parental species. In contrast, the concentration of phenolic glycosides is lower than the parental midpoint, indicating directional dominance. However, there is extensive variation among F1 hybrids. The concentration of tannin and phenolic glycosides in F1 hybrid families is either (1) lower than the midpoint, (2) higher than the midpoint, or (3) indistinguishable from the midpoint of the two parental taxa. It appears that the production of the phenolic glycosides, especially 2' cinnamoylsalicortin, is controlled by one or more recessive alleles. We also observed a two-fold or greater difference in concentration between some hybrid families. We discuss how chemical variation may effect the relative susceptibility of hybrid willows to herbivores. PMID- 10854739 TI - Cyanotypic frequencies in adjacent and mixed populations of Trifolium occidentale Coombe and Trifolium repens L. are regulated by different mechanisms. AB - The cyanogenic polymorphism in Trifolium repens is caused by the variation in two genes, the interaction of which produces four distinct cyanotypes. Along the Atlantic coasts of Bretagne, T. repens is sometimes found in populations mixed with the related species Trifolium occidentale, although the latter species usually occurs only in a narrow fringe along the coast, whereas T. repens is a more inland species. No plants of T. occidentale have ever been reported to have linamarase activity. Indeed, of 763 T. occidentale plants studied, none contained linamarase activity. However, the variation in the proportion of cyanotypes in T. repens was enormous, even between sites less than 2km apart. Our results confirm the presumption that T. repens and T. occidentale are indeed separate species. Both the fact that T. occidentale plants never contain linamarase activity, and the difference in proportion of plants with cyanoglucosides in mixed stands show that gene flow between the species must be rare. These dissimilar distributions strongly indicate that cyanotypic frequencies in adjacent and mixed populations of the very closely related species T. occidentale and T. repens are regulated by different mechanisms PMID- 10854740 TI - Observations of high genetic variability in the endangered Australian terrestrial orchid Pterostylis gibbosa R. Br. (Orchidaceae). AB - The genetic variation in all known populations of an endangered Australian native terrestrial orchid Pterostylis gibbosa R.Br., was investigated with starch gel electrophoresis. A total of 16 isozyme loci were assayed. The percentage of polymorphic loci (P), the number of alleles per locus (A), observed and expected heterozygosity at population levels were 69%, 2.21, 0.210, 0.261, respectively. The G(st) value of 15% indicates that around 85% of variation resides within populations. Despite isolation by distance most alleles were distributed across most of the populations. High genetic variability along with low population divergence may be the result of recent population fragmentation or from extensive gene flow maintained by seed and pollen movement. To investigate whether poor seed viability contributed towards its rarity, an orchid seed viability test using Fluorescein diacetate revealed high seed viability (range 68-90%). Although endangered and restricted to only four geographical areas, P. gibbosa showed a higher level of genetic variation than other orchids with larger populations. PMID- 10854741 TI - Flavonol glycosides from Asplenium foreziense and its five related taxa and A. incisum. AB - The flavonoids of Asplenium foreziense, A. fontanum subsp. fontanum and subsp. pseudofontanum, A. obovatum subsp. obovatum var. obovatum and var. protobillotii, A. obovatum subsp. lanceolatum, and A. incisum were isolated and identified for chemotaxonomic survey. A major constituent of all taxa was kaempferol 3-O gentiobioside. As minor compounds, kaempferol 3,7-O-glycoside and/or kaempferol 3 O-glycoside were found in A. fontanum, A. obovatum and A. foreziense, and kaempferol 3-O-gentiobioside-4'-O-glucoside, kaempferol 3-O-glucoside and quercetin 3-O-diglucoside in A. incisum. It was suggested that A. foreziense, A. fontanum including subsp. pseudofontanum and A. obovatum including subsp. lanceolatum are not only morphologically but also chemotaxonomically related. The East Asian A. incisum was chemically and geographically different from these taxa. PMID- 10854742 TI - Flavonoid distribution in Pyracantha coccinea plants at different growth phases. AB - Flavonoid composition during the ontogenetic cycle was examined in Pyracantha coccinea. The flavonoid profiles of plants at different ages showed marked differences in aerial and hypogeal parts. In the vegetative phase there are flavonoids (flavanones, flavones, and flavonols) only in the aerial parts and they appear gradually during the plant life. These secondary metabolites are detectable in the roots exclusively in the reproductive phase. PMID- 10854743 TI - Geographical variation in the surface flavonoids of Pulicaria dysenterica. AB - Four chemical races were detected in Pulicaria dysenterica, when sampled within Europe, on the basis of the surface flavonoids present. One race uniquely contained quercetagetin 3,7-dimethyl ether and another 6-hydroxykaempferol 3,4' dimethyl ether. A third race was based on plants having 6-hydroxykaempferol 3,7 dimethyl ether together with quercetagetin 3,7,3'-trimethyl ether. The fourth race contained the above two compounds, as well as quercetagetin 3,7,3',4' tetramethyl ether and 6-hydroxykaempferol 3,7,4'-trimethyl ether. These lipophilic constituents were variously present on the surfaces of leaf, ray floret, disc floret and fruit. By contrast, the vacuolar flavonoid of all tissues and all races was uniformly quercetin 3-glucuronide. The kaempferol 3-glucoside previously reported in flowers was not detected. Of the lipophilic flavonoids newly reported from this plant, one 6-hydroxykaempferol 3,7,4'-trimethyl ether is new to nature. PMID- 10854744 TI - Sambunigrin and cyanogenic variability in populations of Sambucus canadensis L. (Caprifoliaceae). AB - The presence or absence of cyanogenic glycosides was determined for individuals from nine populations of Sambucus canadensis L. (elderberry) of east-central Illinois. In most of the populations tested, all or most of the individuals did not produce these compounds, in one, all were cyanogenic, whereas in another population this trait was highly variable. Addition of the enzyme emulsin to negative tests did not result in any further release of cyanide. The glycoside responsible is (S)-sambunigrin. PMID- 10854745 TI - Ratios of cis- and trans-Sabinene Hydrate in Origanum majorana L. and Origanum microphyllum (Bentham) Vogel. AB - (+)-cis-Sabinene hydrate and (+)-trans-sabinene hydrate are the main monoterpenes found in marjoram (Origanum majorana), but can also be found in other Origanum species as well, as in e.g. Melaleuca alternifolia. The synthesis of sabinene hydrate in marjoram (Origanum majorana) is performed by sabinene hydrate synthase. It is claimed, that both, (+)-cis- and (+)-trans-sabinene hydrate are produced by the same enzyme in an exact ratio of 10:1. To verify this in vitro result in vivo, we analysed single plants of 20 different genotypes of Origanum majorana and of three different populations of Origanum microphyllum and calculated the ratios of (+)-cis- to (+)-trans-sabinene hydrates. In Origanum majorana a constant ratio of 20:1 could be found, whereas in Origanum microphyllum the ratio did not prove to be constant. PMID- 10854746 TI - 2-Pyrrolidineacetic Acid and Pyrrolizidine Alkaloids from Melampodium divaricatum. PMID- 10854747 TI - Sesquiterpene lactones from Chrysolaena platensis. PMID- 10854748 TI - Isolation of Cneorubin X, an unusual diterpenoid from Ptaeroxylon obliquum (Ptaeroxylaceae). PMID- 10854749 TI - Flavonoids of Tagetes stenophylla Robinson (Asteraceae) as taxonomic markers. PMID- 10854750 TI - Expression of PSD-95/SAP90 is critical for N-methyl-D-aspartate receptor-mediated thermal hyperalgesia in the spinal cord. AB - PSD-95/SAP90, a molecular scaffold protein, attaches the N-methyl-D-aspartate receptor to cellular signaling pathways through PSD-95/DLG/Z0-1 domain interactions at neuronal synapses.(5,9) This suggests that PSD-95/SAP90 might be involved in many physiological and pathophysiological actions triggered via the N methyl-D-aspartate receptor in the central nervous system. Here, we present evidence that suppression of the expression of PSD-95/SAP90 in the spinal cord significantly attenuated facilitation of the tail-flick reflex triggered through N-methyl-D-aspartate receptor activation but not baseline tail-flick reflex latency. Moreover, PSD-95/SAP90's messenger RNA and protein were enriched in the spinal cord and selectively distributed in the superficial dorsal horn, where PSD 95/SAP90 overlapped with the N-methyl-D-aspartate receptor. In spinal cord neurons, PSD-95/SAP90 interacted with the N-methyl-D-aspartate receptor subunits 2A/2B. It is indicated that activation of the N-methyl-D-aspartate receptor in spinal hyperalgesia results in association of the N-methyl-D-aspartate receptor with PSD-95/SAP90 and that PSD-95/SAP90 is required for noxious thermal hyperalgesia triggered via the N-methyl-D-aspartate receptor at the spinal cord level. The present findings may provide novel insights into the mechanisms for persistent sensitization of the somatosensory system. PMID- 10854751 TI - Orientation biased extended surround of the receptive field of cat retinal ganglion cells. AB - Here we report that the extended surround outside the classical receptive center (hereafter called the extended surround) of most retinal ganglion cells in the cat exhibit significant orientation bias to grating stimuli, and that the center and the extended surround show different orientation biases at different spatial frequencies. As a result, some retinal ganglion cells possess a complex receptive field structure, which allows them to detect sophisticated image segmentation (e.g. texture segmentation) in addition to simple luminance edges. This property was previously thought to exist primarily in the visual cortex. Moreover, in about one quarter of 128 cells studied the center did not exhibit an orientation bias. Thus, these surrounds alone may determine the cells' orientation bias.In conclusion, the extended surround may play an important role in processing more complex pattern in natural scenes since the classical receptive field is too small to describe all the properties of a retinal ganglion cell. PMID- 10854752 TI - Acetylcholine induces neuritic outgrowth in rat primary olfactory bulb cultures. AB - The rat olfactory bulb is innervated by basal forebrain cholinergic neurons and is endowed with both nicotinic and muscarinic receptors. The development of this centrifugal cholinergic innervation occurs mainly in early postnatal stages. This developmental time-course and the demonstration that acetylcholine can modulate some aspects of neuronal proliferation, differentiation or death, suggests the possible involvement of cholinergic afferents in the morphogenesis and/or plasticity of the olfactory bulb. The purpose of the present work was to assess whether acetylcholine could modulate neuronal morphogenesis in the olfactory bulb. Toward this aim, we developed a primary culture model of rat olfactory bulbs. Three major cell types were identified on the basis of their morphological and immunocytochemical phenotype: neuronal-shaped cells expressing the neuronal markers neuron specific enolase, microtubule associated protein 2, neural cell adhesion molecule and beta-tubulin III; glial-like cells immunoreactive for glial fibrillary acidic protein and flattened cells immunolabelled with antibodies against beta-tubulin III and nestin, most likely neuronal precursors. After three to six days of treatment with 100-microM carbachol, a cholinergic agonist, significant increase in neuritic length was observed in cultured olfactory bulb neurons. The neurite outgrowth effect of carbachol was abolished by co-treatment with 1 microM alpha-bungarotoxin, an alpha 7 subunit nicotinic receptor antagonist, but was not affected by the addition of 10 microM atropine, a general muscarinic antagonist. The effect of carbachol was also mimicked by the nicotinic agonists, nicotine (100 microM) and epibatidine (10 microM). This pharmacological profile suggested the involvement of nicotinic receptors of the alpha 7-like subtype as confirmed using 125I-alpha-bungarotoxin receptor autoradiography.Taken together, these data argue for a role for nicotinic receptors in neuritic outgrowth in the rat olfactory bulb and provide a cellular support to the previously described effects of acetylcholine on olfactory bulb plasticity in vivo. PMID- 10854753 TI - A group of cortical interneurons expressing mu-opioid receptor-like immunoreactivity: a double immunofluorescence study in the rat cerebral cortex. AB - mu-Opioid receptor-expressing neurons in the rat cerebral neocortex were characterized by an immunolabeling method with an antibody to a carboxyl terminal portion of the receptor. They were small, bipolar, vertically elongated, non pyramidal neurons, and scattered mainly in layers II-IV. We examined chemical characteristics of mu-opioid receptor-expressing neocortical neurons by the double immunofluorescence method. Almost all neuronal cell bodies expressing mu opioid receptor-like immunoreactivity showed immunoreactivity for GABA, suggesting that they were cortical inhibitory interneurons. mu-Opioid receptor immunoreactive neurons were further studied by the double staining method with markers for the subgroups of cortical GABAergic neurons. Immunoreactivities for vasoactive intestinal polypeptide, corticotropin releasing factor, choline acetyltransferase, calretinin and cholecystokinin were found in 92, 79, 67, 35 and 35% of mu-opioid receptor-immunoreactive cortical neurons, respectively. In contrast, less than 10% of mu-opioid receptor-immunoreactive neurons showed immunoreactivity for parvalbumin, calbindin, somatostatin, neuropeptide Y or nitric oxide synthase. Moreover, mu-opioid receptor-immunoreactive neurons very frequently exhibited preproenkephalin immunoreactivity, but not preprodynorphin immunoreactivity. The present results indicate that mu-opioid receptor-expressing neurons belong to a distinct subgroup of neocortical GABAergic neurons, because vasoactive intestinal polypeptide, corticotropin releasing factor, choline acetyltransferase, calretinin and cholecystokinin have often been reported to coexist with one another in single neocortical neurons. Methionine-enkephalin, which is a major product of the preproenkephalin gene, is known to be one of the most potent endogenous ligands for mu-opioid receptor. Thus, the expression of mu opioid receptor in preproenkephalin-producing neurons suggested that mu-opioid receptor serves as an autoreceptor for the subpopulation of GABAergic interneurons at a single-neuron or population level. PMID- 10854754 TI - Role of agonist-dependent receptor internalization in the regulation of mu opioid receptors. AB - Organotypic cultures and ileal neuromuscular preparations were used to determine (i) whether endogenous release of opioids by electrical stimulation induces mu receptor endocytosis, and (ii) whether and under which conditions ligand-induced mu receptor endocytosis influences the responsiveness of neurons expressing native mu receptors. In longitudinal muscle-myenteric plexus preparations, electrical stimulation at 20 Hz induced a prominent endocytosis of mu receptors in enteric neurons, indicating endogenous release of opioids. A similar massive endocytosis was triggered by exogenous application of the mu receptor agonist, [D Ala(2),MePhe(4), Gly-ol(5)] enkephalin, whereas exogenous application of morphine was ineffective. [D-Ala(2),MePhe(4),Gly-ol(5)] enkephalin and morphine induced a concentration-dependent inhibition of neurogenic cholinergic twitch contractions to electrical stimulation at 0.1 Hz. beta-Chlornaltrexamine shifted to the right the inhibitory curve of both agonists with a concentration-dependent reduction of the maximum agonist response, which is consistent with the existence of spare mu opioid receptors. Under these conditions, the induction of mu receptor endocytosis by exogenously applied [D-Ala(2), MePhe(4),Gly-ol(5)] enkephalin diminished the inhibitory effect of this agonist on twitch contractions and tritiated acetylcholine release. In contrast, there was no reduction of the inhibitory effect of morphine, which failed to induce mu receptor endocytosis, on neurogenic cholinergic response. These results provide the first evidence for the occurrence of mu receptor endocytosis in neurons by endogenously released opioids and show that agonist-dependent mu receptor endocytosis could serve as a mechanism to regulate mu opioid receptor responsiveness to ligand stimulation when the opioid receptor reserve is reduced. PMID- 10854755 TI - Synergistic effects of unilateral immunolesions of the cholinergic projections from the basal forebrain and contralateral ablations of the inferotemporal cortex and hippocampus in monkeys. AB - Monkeys, with unilateral immunotoxic lesions of the basal nucleus of Meynert that remove cholinergic innervation of the ipsilesional neocortex, and ablations of the contralateral inferotemporal neocortex, were impaired on retention of visual discriminations learnt before surgery and on acquisition of new discriminations. This demonstrates that the cholinergic projection from the basal nucleus supports the functions of its cortical target area. Our previous studies have shown that the impairment on discrimination performance following bilateral lesions of the basal nucleus is transient and that bilateral lesions of the diagonal band of Broca, that remove cholinergic innervation of the hippocampus, are without effect on these tasks. However, the impairment resulting from bilateral lesions of the basal nucleus plus the diagonal band, or from bilateral inferotemporal cortex ablations, is severe and persistent. Bilateral inferotemporal ablations deprive the hippocampus of much of its visual input by producing a discontinuity in cortico-cortical transmission, whereas basal nucleus lesions may merely prevent the modification of visually-derived information in the inferotemporal cortex without depriving the hippocampus of visual input. In the monkeys with crossed unilateral basal nucleus plus inferotemporal cortex lesions, the addition of a diagonal band lesion to the basal nucleus lesion produced an impairment on retention of visual discriminations and sustained the acquisition impairment. This confirms the previous finding that the basal nucleus and diagonal band act synergistically in producing a severe and permanent impairment. Further addition of an excitotoxic hippocampal lesion to the hemisphere with the inferotemporal cortex ablation did not add to the learning impairment. This supports the suggestion that the inferotemporal cortex ablation has deprived the hippocampus of its visual input.Overall, these experiments demonstrate that the cholinergic projections from the basal nucleus and diagonal band participate in the learning and memory functions of the temporal lobes. PMID- 10854756 TI - Functional expression of cell surface cannabinoid CB(1) receptors on presynaptic inhibitory terminals in cultured rat hippocampal neurons. AB - At present, little is known about the mechanisms by which cannabinoids exert their effects on the central nervous system. In this study, fluorescence imaging and electrophysiological techniques were used to investigate the functional relationship between cell surface cannabinoid type 1 (CB(1)) receptors and GABAergic synaptic transmission in cultured hippocampal neurons. CB(1) receptors were labelled on living neurons using a polyclonal antibody directed against the N-terminal 77 amino acid residues of the rat cloned CB(1) receptor. Highly punctate CB(1) receptor labelling was observed on fine axons and at axonal growth cones, with little somatic labelling. The majority of these sites were associated with synaptic terminals, identified either with immunohistochemical markers or by using the styryl dye FM1-43 to label synaptic vesicles that had undergone active turnover. Dual labelling of neurons for CB(1) receptors with either the inhibitory neurotransmitter GABA or its synthesising enzyme glutamate decarboxylase, demonstrated a strong correspondence. The immunocytochemical data was supported by functional studies using whole-cell patch-clamp recordings of miniature inhibitory postsynaptic currents (mIPSCs). The cannabinoid agonist WIN55,212-2 (100nM) markedly inhibited (by 77+/-6.3%) the frequency of pharmacologically-isolated GABAergic mIPSCs. The effects of WIN55,212-2 were blocked in the presence of the selective CB(1) receptor antagonist SR141716A (100nM).In conclusion, the present data show that cell surface CB(1) receptors are expressed at presynaptic GABAergic terminals, where their activation inhibits GABA release. Their presence on growth cones could indicate a role in the targeting of inhibitory connections during development. PMID- 10854757 TI - Expression of D(3) receptor messenger RNA and binding sites in monkey striatum and substantia nigra after nigrostriatal degeneration: effect of levodopa treatment. AB - D(3) receptors are prominently localized in the primate caudate-putamen, and D(3) receptor agonist properties may offer an advantage in Parkinson's disease therapy. In the present experiments, we investigated the relationship between D(3) receptor mRNA, D(3) receptor sites and the dopamine transporter in monkey basal ganglia by comparing their distribution in the brain of control and 1 methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys (Samirai sciureus). In control monkeys, D(3) receptor mRNA appears to be widely expressed throughout the brain, with a distribution similar to that observed in both man and rodent. D(3) receptors are present in areas which express mRNA but also in some which do not, an observation which suggests they may be both pre- and postsynaptic in the monkey brain. Chronic MPTP administration, which selectively destroys the nigrostriatal system, resulted in a 70 to 99% depletion of the dopamine transporter in the basal ganglia. Autoradiographic analysis showed that after MPTP treatment there was a significant decline in D(3) receptors in the caudate, but not putamen, globus pallidus, substantia nigra or other dopaminergic regions. D(3) receptor mRNA expression was not changed in any region after nigrostriatal lesioning. Two weeks of L-3,4-dihydroxyphenylalanine (levodopa, L DOPA) treatment, which alleviated Parkinsonism but also induced dyskinesias, reversed the MPTP-induced decline in caudate D(3) receptors. These results show that there is a selective decline in D(3) receptors in the caudate after nigrostriatal degeneration, which is reversed by L-DOPA treatment. Since the majority of dopaminergic nerve terminals were destroyed after MPTP lesioning, the reversal in D(3) receptors after L-DOPA treatment may represent an increase in caudate postsynaptic receptors, which could conceivably contribute to an imbalance in striatal circuitry and the development of dyskinesias. PMID- 10854758 TI - Differential regulation of calmodulin content and calmodulin messenger RNA levels by acute and repeated, intermittent amphetamine in dopaminergic terminal and midbrain areas. AB - Repeated doses of psychoactive drugs often produce adaptive responses that differ from the initial drug application and additional adaptive processes occur following cessation of the drug. The relationship between alterations in calmodulin protein and messenger RNA produced by an initial versus a repeated dose of amphetamine was examined, as well as changes following drug cessation. Calmodulin protein and messenger RNA of the three individual calmodulin genes were measured in rat dopaminergic cell body and terminal areas following acute or repeated amphetamine. Rats were either injected once with 2.5mg/kg amphetamine or saline and decapitated after 3h, or given 10 injections of amphetamine three to four days apart and decapitated 3h after the final injection. Calmodulin messenger RNA and protein were also measured three and seven days after ceasing drug treatment. Acute amphetamine increased calmodulin 1.7-fold in the striatum and threefold in the ventral mesencephalon, with corresponding elevations in calmodulin messenger RNAs. In response to the 10th dose of amphetamine, however, the degree of increase in calmodulin was diminished in the striatum and ablated in the ventral mesencephalon. Correspondingly, select species of calmodulin messenger RNA were decreased from control levels. In the frontal cortex or nucleus accumbens, calmodulin levels were basically unaltered by the first or 10th doses of amphetamine, but both calmodulin and its messenger RNA were altered with time upon cessation of the drug. Three days later, both calmodulin protein and messenger RNA were decreased in select brain areas. By seven days after the 10th injection, calmodulin content was altered compared to saline controls in all areas, but the change in messenger RNA no longer paralleled the change in protein.Our findings demonstrate that both calmodulin protein and select species of calmodulin messenger RNA are altered by acute amphetamine, but this effect is attenuated after repeated, intermittent amphetamine. There are further time dependent changes after cessation of repeated amphetamine, which may reflect compensatory neuronal responses. The alterations in calmodulin content and synthesis could contribute to changes in patterns or duration of behaviors that occur upon cessation of repeated amphetamine. PMID- 10854759 TI - Changes in astrocytic basic fibroblast growth factor expression during and after prolonged exposure to escalating doses of amphetamine. AB - We have shown that brief exposure to amphetamine leads to sustained glutamate dependent increases in expression of the neurotrophic, neuroprotective factor, basic fibroblast growth factor, in astrocytes in dopaminergic cell body regions and that blockade of basic fibroblast growth factor in this region prevents the development of behavioral sensitization to amphetamine. Here we examine the effects of prolonged exposure to an escalating-dose regimen of amphetamine known to induce long-lasting sensitization to amphetamine and leading to increases in neuronal dendritic length and spine density in nucleus accumbens and prefrontal cortex and to decreases in spine density in occipital cortex. Astrocytic basic fibroblast growth factor immunoreactivity was increased in both dopaminergic cell body and terminal regions one week after termination of a two-week amphetamine treatment (1-4mg/kg). These effects were not evident one week after a five-week treatment (1-9mg/kg) and, in fact, one month later basic fibroblast growth factor levels in cell body regions were decreased. In the occipital cortex, basic fibroblast growth factor immunoreactivity was decreased one week after the two week amphetamine treatment, but was not different from that seen in saline treated animals after the five-week treatment. Increased astrocytic basic fibroblast growth factor expression appears to be an early, but relatively prolonged, response to amphetamine exposure and seems to parallel structural changes induced by repeated drug exposure.These findings suggest that basic fibroblast growth factor may participate in the development of structural changes brought about by amphetamine. The fact that the basic fibroblast growth factor response is not maintained after prolonged intense exposure to amphetamine suggests that the factors that initially induce basic fibroblast growth factor expression are self-regulating. PMID- 10854760 TI - Neurochemical and electrophysiological studies on the functional significance of burst firing in serotonergic neurons. AB - We have previously described a population of 5-hydroxytryptamine neurons which repetitively fires bursts of usually two (but occasionally three or four) action potentials, with a short (<20 ms) interspike interval within a regular low frequency firing pattern. Here we used a paradigm of electrical stimulation comprising twin pulses (with 7- or 10-ms inter-pulse intervals) to mimic this burst firing pattern, and compared the effects of single- and twin-pulse electrical stimulations in models of pre- and postsynaptic 5-hydroxytryptamine function. Firstly, we measured the effect of direct electrical stimulation (2 Hz for 2 min) of rat brain slices on efflux of preloaded [3H]5-hydroxytryptamine. In this in vitro model, twin-pulse stimulation increased the efflux of tritium by about twice as much as did single-pulse stimulation. This effect was evident in the medial prefrontal cortex (area under the curve: 2. 59+/-0.34 vs 1.28+/-0.22% relative fractional release), as well as in the caudate-putamen (3.93+/-0.65 vs 2.17+/-0.51%) and midbrain raphe nuclei (5.42+/-1.05 vs 2.51+/-0.75%). Secondly, we used in vivo microdialysis to monitor changes in endogenous extracellular 5 hydroxytryptamine in rat medial prefrontal cortex in response to electrical stimulation (3 Hz for 10 min) of the dorsal raphe nucleus. In this model, twin pulse stimulation of the dorsal raphe nucleus increased 5-hydroxytryptamine by approximately twice as much as did single-pulse stimulation at the same frequency (area under the curve: 50.4+/-9.0 vs 24.2+/-4.4 fmol). Finally, we used in vivo extracellular recording to follow the response of postsynaptic neurons in the rat medial prefrontal cortex to 5-hydroxytryptamine released by dorsal raphe stimulation. Electrical stimulation of the dorsal raphe nucleus (1 Hz) induced a clear-cut poststimulus inhibition in the majority of cortical neurons tested. In these experiments, the duration of poststimulus inhibition following twin-pulse stimulation was markedly longer than that induced by single-pulse stimulation (200+/-21 vs 77+/-18.5 ms). Taken together, the present in vitro and in vivo data suggest that in 5-hydroxytryptamine neurons, short bursts of action potentials will propagate along the axon to the nerve terminal and will enhance both the release of 5-hydroxytryptamine and its postsynaptic effect. PMID- 10854761 TI - Single-unit responses of serotonergic medullary and pontine raphe neurons to environmental cooling in freely moving cats. AB - Brain serotonin has long been implicated in the regulation of body temperature, although its precise role is not completely understood. The present study examined the effects of environmental cooling (4-8 degrees C for 2 or 4h) on the single-unit activity of serotonergic neurons recorded in the medullary raphe nuclei obscurus and pallidus and in the pontine dorsal raphe nucleus of freely moving cats. These neuronal groups have primarily descending projections to the spinal cord and ascending projections to the forebrain, respectively. Cold exposure induced shivering and piloerection, but no appreciable changes in core temperature. Of the medullary serotonergic cells studied (n=14), seven were activated and seven were unresponsive to cold exposure. For the responsive cells, the mean increase and peak effect in unit activity relative to baseline were 31% and 46%, respectively. Of the seven cold-responsive cells, the activity of four was monitored when the animals were transferred back to room temperature (23 degrees C). Within 15-30 min, the activity of these cells returned to baseline. In contrast, none of the dorsal raphe nucleus cells studied (n=14) displayed a significant change in neuronal activity during cold exposure, suggesting that these neurons do not receive afferent input from cold-sensitive cutaneous receptors or participate in thermoregulatory responses evoked by low ambient temperatures.Overall, these results suggest that a subset of medullary serotonergic neurons play a role in physiological mechanisms underlying cold defense (e.g. increases in motor output and/or autonomic outflow). On the other hand, the lack of responsiveness of serotonergic dorsal raphe nucleus neurons to cold exposure does not support a specific role for these cells in thermoregulation. PMID- 10854762 TI - Infusion of adenylyl cyclase inhibitor SQ22,536 into the medial pontine reticular formation of rats enhances rapid eye movement sleep. AB - Microinjection of cholinergic and adenosinergic agonists into the medial pontine reticular formation of rats produces long lasting increases in the time spent in rapid eye movement sleep. Several G-protein-coupled muscarinic and adenosinergic receptors share a common action of inhibition of adenylyl cyclase and inhibition of cyclic adenosine monophosphate. Inhibition of cyclic adenosine monophosphate has been implicated in the mechanism of rapid eye movement sleep induction in the cat. We sought to determine whether a direct inhibitor of adenylyl cyclase microinjected into the rat reticular formation at sites where muscarinic and adenosinergic agonists are effective in producing long lasting elevations in rapid eye movement sleep also result in similar effects on the sleep/wake cycle. The caudal, oral pontine reticular formation was unilaterally infused with 60 nl volumes of carbachol (0.1-1.1mM) and N(6)-cyclohexyladenosine (0.1mM) each within 1h of lights on. Sites effective for significantly elevating rapid eye movement sleep for the 8h following microinjection of both receptor agonists were additionally injected with the adenylyl cyclase inhibitor, SQ22,536 (0.1M). Pontine injections of SQ22,536 resulted in significant mean increases in rapid eye movement sleep time and rapid eye movement sleep period frequency at all of these sites. As with the receptor agonists, SQ22,536 did not alter latency to rapid eye movement sleep onset. Rapid eye movement sleep amounts were observed to be significantly elevated by SQ22,536 at two days, but not at four days, following a single microinjection. These data implicate inhibition of cyclic adenosine monophosphate in the pons of the rat as a mechanism involved in the long-term modulation of rapid eye movement sleep. This mechanism may underlie the homeostatic regulation exhibited by this sleep-state. PMID- 10854763 TI - Simultaneous activation of N-methyl-D-aspartate and neurokinin-1 receptors modulates c-Fos and Zif/268 expression in the rat trigeminal nucleus caudalis. AB - We examined the acute expression of c-Fos or Zif/268 by simultaneous activation of N-methyl-D-aspartate receptor and neurokinin-1 receptor of the trigeminal nucleus caudalis in anesthetized rats. A selective N-methyl-D-aspartate receptor agonist, N-methyl-D-aspartate, and/or a selective neurokinin-1 receptor agonist, substance P, was applied topically to the dorsal surface of the spinal trigeminal tract. Immunohistochemically stained nuclei for c-Fos and Zif/268 at laminae I and II of the trigeminal nucleus caudalis were counted. Ipsilateral c-Fos and Zif/268 were increased significantly dose-dependently by N-methyl-D-aspartate (at 136 and 340 microM, and at 68, 136 and 340 microM, respectively). On the contralateral side, only Zif/268 increased significantly (at 68, 136 or 340 microM). These increases were abolished by D-2-amino-5-phosphonovaleric acid (at 25 mM), a selective N-methyl-D-aspartate receptor antagonist. Substance P (at 3.7 or 7. 4 microM) significantly increased dose-dependently ipsilateral c-Fos and Zif/268. On the contralateral side, only c-Fos was significantly increased (at 3.7 and 7.4 microM). These increases were abolished by D-2-amino-5 phosphonovaleric acid (at 25 mM) and L-703,606 (at 10 microM), a selective neurokinin-1 receptor antagonist. The combined application of N-methyl-D aspartate 340 microM + substance P (at 0.74 or 3.7 microM) significantly increased ipsilateral c-Fos compared to either agent alone. Combined application of N-methyl-D-aspartate 340 microM + substance P at 0.74, 3.7 or 7.4 microM significantly increased ipsilateral Zif/268 expression compared to either drug alone. Other combinations did not increase c-Fos and Zif/268. Our results indicate that activation of N-methyl-D-aspartate or neurokinin-1 receptor of the trigeminal nucleus caudalis contributes to the acute induction of both c-Fos and Zif/268 on the ipsilateral superficial layer of this nucleus and simultaneous activation of both receptors by their agonists with specific concentrations produces a marked expression of these proteins. Simultaneous activation of N methyl-D-aspartate and neurokinin-1 receptors under some specific conditions may augment synaptic transmission, contributing to long-term neuronal change. PMID- 10854764 TI - GABA(A) receptor-mediated effects on expression of c-Fos in rat trigeminal nucleus following high- and low-intensity afferent stimulation. AB - We examined the effects of systemic administration of a GABA(A) receptor agonist, muscimol, or antagonist, bicuculline, on the expression of c-Fos protein induced 3h after electrical stimulation of the trigeminal ganglion at low (0.1 mA) or high intensities (1. 0 mA) in the urethane-anesthetized rat. In saline-treated rats, 10 min stimulation of the trigeminal ganglion induced c-Fos-immunopositive neurons throughout the full extent of the ipsilateral superficial layers of the trigeminal nucleus caudalis, and dorsal or dorsomedial part of the nuclei rostral to obex (trigeminal nucleus principalis, dorsomedial nucleus of trigeminal nucleus oralis, dorsomedial nucleus of trigeminal nucleus interpolaris). Animals stimulated at 1. 0 mA induced a significantly higher number of labeled neurons in all trigeminal sensory nucleus than animals stimulated at 0.1 mA. In rats treated with 1mg/kg i.p. muscimol and stimulated at 0.1 mA, the numbers of Fos-positive neurons in trigeminal nucleus caudalis, dorsomedial nucleus of trigeminal nucleus interpolaris, and dorsomedial nucleus of trigeminal nucleus oralis were significantly decreased. However, after stimulation at 1.0 mA, the numbers of Fos positive neurons in the superficial layers of trigeminal nucleus caudalis was increased and no changes occurred in the numbers of Fos-positive neurons in the magnocellular zone of trigeminal nucleus caudalis, the dorsomedial nucleus of trigeminal nucleus interpolaris, or dorsomedial nucleus of trigeminal nucleus oralis compared to saline-treated controls. In rats treated with 2mg/kg i.p. bicuculline and stimulated at 0.1 mA, the number of Fos-positive neurons increased in the superficial layers of trigeminal nucleus caudalis and trigeminal nucleus principalis. However, after stimulation at 1.0 mA, the number of Fos positive neurons was unchanged in superficial layers of trigeminal nucleus caudalis, but decreased in the magnocellular zone of trigeminal nucleus caudalis, dorsomedial nucleus of trigeminal nucleus interpolaris and dorsomedial nucleus of trigeminal nucleus oralis. There was a specific loss of Fos-positive neurons in the maxillary and ophthalmic divisions (ventrolateral half) of trigeminal nucleus caudalis. These results indicate that the expression of c-Fos in the trigeminal nucleus is differentially regulated through GABA(A) receptors in a manner that is dependent on the nucleus and the type of primary afferents that are activated by different stimulus intensities. PMID- 10854765 TI - Redox modulation of recombinant human GABA(A) receptors. AB - We previously reported that GABA-evoked currents of rat retinal ganglion cells were modulated by redox agents. In this study, we further characterized the effects of redox modulation on GABA receptors using recombinant human subunits in the Xenopus oocyte expression system with two-electrode voltage-clamp recording. GABA receptors composed of subunits alpha(1-3), beta(1-3), gamma(1), gamma(2S,) and rho(1) were expressed. The sulfhydryl reducing agent dithiothreitol reversibly potentiated the responses of various combinations of functional recombinant GABA(A) subunits, whether expressed as triplets (alpha(1)beta(1 3)gamma(1,2S)), pairs (alpha(1-3)beta(1-3); beta(1-3)gamma(1,2S)), or singly (beta(2)). These effects of dithiothreitol were rapidly reversible, and the oxidizing agent 5-5'-dithiobis-2-nitrobenzoic acid exerted the opposite effect. In contrast to these effects on GABA(A) receptors, dithiothreitol had no effect on the responses of homomeric GABA rho(1) (GABA(C)) receptors. The degree of dithiothreitol potentiation of GABA(A) receptor responses depended on subunit composition. Co-expression of gamma(2S) with alpha(1)beta(1-3) subunits resulted in markedly less dithiothreitol potentiation of GABA-evoked currents than that observed for alpha(1-3)beta(1-3) subunits in the absence of gamma(2S). None the less, the magnitude of dithiothreitol potentiation could be restored by using a combination of lower GABA concentrations (5-10 microM) and higher dithiothreitol concentrations (5-20mM). N,N,N', N'-tetrakis(2-pyridyl-methyl)ethylenediamine, a high-affinity Zn(2+) chelator, also potentiated GABA(A) receptor currents. However, the potentiation produced by 10mM dithiothreitol was larger than that produced by saturating concentrations of N,N,N', N'-tetrakis(2-pyridyl methyl)ethylenediamine (100 microM), implying that at least part of the effect of dithiothreitol was due to redox modulation rather than Zn(2+) chelation. Dithiothreitol also potentiated the spontaneous current of homomeric GABA(A) receptors composed of beta subunits. Mutation of a single cysteine residue in the M3 domain, yielding homomeric beta(3)(C313A) receptors, abrogated dithiothreitol potentiation of the spontaneous current. In summary, this study further characterizes the modulatory effects of redox agents on recombinant GABA(A) receptors. The degree of redox modulation of GABA(A) receptors depended on subunit composition. In contrast to their effect on GABA(A) receptors, redox agents were not found to modulate GABA(C) receptors composed of homomeric rho(1) subunits. Using site-directed mutagenesis, a cysteine residue was located in the beta(3) subunit which may comprise one of the redox-active sites that underlies the modulation of heteromeric GABA(A) receptors by reducing and oxidizing agents. PMID- 10854766 TI - The effect of intrathecal endomorphin-2 on the flexor reflex in normal, inflamed and axotomized rats: reduced effect in rats with autotomy. AB - Endomorphin-2, a newly discovered endogenous opioid peptide and agonist at the mu opioid receptor, was injected intrathecally in normal rats and animals with unilateral peripheral inflammation or sciatic nerve section and its effect on the nociceptive flexor reflex was analysed. In normal rats, intrathecal endomorphin-2 induced a strong and dose-dependent depression of the reflex, which was naloxone reversible. The effect of intrathecal endomorphin-2 was fairly brief, lasting for about 20-30 min at the highest dose, 4 microg. The effect of endomorphin-2 in inflamed rats was not significantly different from that in normals. After nerve section some rats developed autotomy behavior. In these rats endomorphin-2 had significantly reduced effect. However, the reflex depressive effect of intrathecal endomorphin-2 was unchanged in axotomized rats without autotomy. It is suggested that intrathecal endomorphin-2 has antinociceptive effect in the rat spinal cord under normal and inflammatory conditions. After peripheral nerve injury the sensitivity to endmorphin-2 may be reduced in rats that exhibit ongoing neuropathic pain-like behaviors. PMID- 10854767 TI - Deficits in visceral pain and hyperalgesia of mice with a disruption of the tachykinin NK1 receptor gene. AB - Studies in mice lacking genes encoding for substance P or its receptor (NK1), or with NK1 antagonists, have shown that this system contributes to nociception, but the data are complex. Here, we have further examined the role of NK1 receptors in pain and hyperalgesia by comparing nociceptive responses to mechanical and chemical stimulation of viscera and the resulting hyperalgesia and inflammation in NK1 knockout (-/-) and wild-type (+/+) mice. We concentrated on visceral nociception because substance P is expressed by a much greater proportion of visceral than cutaneous afferents. NK1 -/- mice showed normal responses to visceral mechanical stimuli, measured as behavioural responses to intraperitoneal acetylcholine or hypertonic saline or reflex responses to colon distension in anaesthetized mice, although -/- mice failed to encode the intensity of noxious colon distensions. In contrast, NK1 -/- mice showed profound deficits in spontaneous behavioural reactions to an acute visceral chemical stimulus (intracolonic capsaicin) and failed to develop referred hyperalgesia or tissue oedema. However, in an identical procedure, intracolonic mustard oil evoked normal spontaneous behaviour, referred hyperalgesia and oedema in -/- mice. The inflammatory effects of capsaicin were abolished by denervation of the extrinsic innervation of the colon in rats, whereas those of mustard oil were unchanged, showing that intracolonic capsaicin evokes neurogenic inflammation, but mustard oil does not. Tests of other neurogenic inflammatory stimuli in NK1 -/- mice revealed impaired behavioural responses to cyclophosphamide cystitis and no acute reflex responses or primary hyperalgesia to intracolonic acetic acid. We conclude that NK1 receptors have an essential role mediating central nociceptive and peripheral inflammatory responses to noxious stimuli that evoke neurogenic inflammation, and modulating responses to noxious mechanical stimuli. We propose that two separate hyperalgesia pathways exist, one of which is NK1 receptor dependent, whereas the other does not require intact substance P/NK1 signalling. PMID- 10854768 TI - Comparison of astrocytic and myocytic metabolic dysregulation in apolipoprotein E deficient and human apolipoprotein E transgenic mice. AB - The accumulation of tubular aggregates in type II skeletal muscle fibres and fibrillo-granular inclusions in hippocampal protoplasmic astrocytes are characteristic lesions of apolipoprotein E deficient mice. Moreover these inclusions reacted immunocytochemically with an antibody specific to fragment 17 24 of the published sequence of Alzheimer's amyloid peptide. In an effort to evaluate the role of apolipoprotein E in the formation of these abnormal structures, we examined the tibialis anterior muscle and the hippocampus of several groups of animals including: (i) apolipoprotein E "knockout" mice which had been whole body irradiated with 1200 rads and bone marrow replenished with apolipoprotein E sufficient marrow; and (ii) three transgenic murine strains that had been genetically engineered to express either human apolipoprotein E2, E3 or E4 protein on an apoE deficient background. The results of this study showed that the presence of murine apolipoprotein E (even in subnormal levels in the serum) in irradiated bone marrow replenished mice and in all three (E2, E3 or E4) human apoE transgenic strains was sufficient to prevent the aggregation of sarcoplasmic tubules in the tibialis anterior type II muscle fibres. Similarly apolipoprotein E "knockout" bone marrow replenished mice and all three transgenic strains expressing the different human apolipoprotein E alleles reduced the number of the astrocytic inclusions in the hippocampus to levels not significantly different to those observed in control C57Bl6J animals. The data obtained in this study indicate that neurological and neuromuscular abnormalities found in apoE deficient mice are reversed when apoE protein is replaced in the circulation, either by bone marrow transplantation of normal apoE sufficient marrow, or by gene therapy with the apoE gene, albeit of human origin and irrespective of the allele used. PMID- 10854769 TI - Morphometric analysis of the myelin-associated oligodendrocytic basic protein deficient mouse reveals a possible role for myelin-associated oligodendrocytic basic protein in regulating axonal diameter. AB - Myelin-associated oligodendrocytic basic protein is a member of the proteins constituting the central nervous system myelin. By morphometric analysis, we demonstrated that axons of myelin-associated oligodendrocytic basic protein deficient mice had larger diameters and more myelin lamellae as compared to those of wild-type mice at the same age. It is known that the number of myelin lamellae increases linearly with axonal diameter, and that the rate of radial axonal growth is the factor controlling the rate of myelin formation. In line with these observations, we found that the regression line for axonal diameter and the number of myelin lamellae in myelin-associated oligodendrocytic basic protein deficient mice appeared to be identical to that in wild-type mice, indicating that the increase in the number of myelin lamellae was the result of the increase in axonal diameter. Furthermore, we generated myelin basic protein/myelin associated oligodendrocytic basic protein-double-deficient mice through mating myelin-associated oligodendrocytic basic protein-deficient mice with shiverer mice, an autosomal recessive mutant characterized by a lack of all isoforms of myelin basic protein. With these knock-out mice, we showed that axons of the double-deficient mice had larger diameters and smaller form factor, an index of the deformation of the fiber contour, in ensheathed fibers than those of shiverer mice, although there was no difference in axonal diameter of unmyelinated fibers between them. Taken together, myelin-associated oligodendrocytic basic protein seemed to play a role in controlling axonal diameter and in keeping axons round. PMID- 10854770 TI - Manipulation of intracellular calcium has no effect on rate of migration of rat autonomic motor neurons in organotypic slice cultures. AB - Migration of neurons is a key step in the formation of the central nervous system, and an increase in internal Ca(2+) concentration has been shown to increase the rate of migration of granule cells along radial glial processes in slices of postnatal cerebellum. In embryonic spinal cord, the non-radial migration of autonomic motor neurons from the ventral horn dorsally into the region of the intermediolateral nucleus differs from that of granule cells, so it is possible that the role of Ca(2+) may also differ in the migration of these two types of neurons. To investigate this possibility, we made organotypic slice cultures of thoracic spinal cord from rat embryos. In control slices after about one day in vitro, diaphorase-positive autonomic motor neurons had migrated 100 microm at a rate of 3.6 microm/h. In experimental slice cultures, we added pharmacological reagents that are known to either increase or decrease internal Ca(2+) levels, including some reagents used successfully in the aforementioned granule cell studies. None of the nine reagents had a significant effect on migration speed of autonomic motor neurons in slice cultures. Our results suggest that autonomic motor neuron migration is not regulated by internal Ca(2+) levels, and hence this mechanism may not be used universally by all types of neurons. PMID- 10854771 TI - Efferent function of vestibular afferent endings? Similar localization of N-type calcium channels, synaptic vesicle and synaptic membrane-associated proteins. AB - We investigated the distribution of N-type voltage-dependent calcium channels that mediate Ca(2+) entry initiating transmitter release in the rat vestibular sensory epithelium. We used confocal microscopy to assess the in vitro labeling by fluorescent specific ligand binding, omega-conotoxin-GVIA and also the immunolabeling of presynaptic soluble N-ethylmaleimide-sensitive fusion factor attachment protein receptor (SNARE) proteins, syntaxin, 25,000 mol. wt synaptosome-associated protein and synaptotagmin: components of the neurotransmitter exocytosis machinery. We found that there was a close anatomical association between the voltage-gated calcium channels, the synaptic vesicle and synaptic membrane-associated proteins on the afferent nerve calyces and probably afferent boutons, which are postsynaptic compartments. Our data suggest that these peripheral afferent endings possess the presynaptic Ca(2+) channels and the components of the presynaptic SNARE proteins involved in synaptic vesicle docking and calcium-dependent exocytosis. They provide additional evidence for a secretory function and efferent role of these endings in hair cell neurotransmission. PMID- 10854772 TI - The role of some brain structures in the switching of the descending influences in operantly conditioned rats. AB - A hypothesis was proposed according to which the switching of descending influences by the corticospinal and corticorubrospinal systems was associated with rubro-olivary projection involvement depending on the context of movement [Kennedy P. R. (1990) Trends Neurosci. 13, 474-479]. Our results confirmed and extended this hypothesis. It was shown that a preliminary transection of the dorsolateral funiculus (containing the rubrospinal tract) accelerated the compensatory rehabilitation process following lesions of the red nucleus and the ventrolateral thalamic nucleus in albino rats with learned instrumental reflexes on equilibrium. A preliminary lesion of the ventrolateral thalamic nucleus considerably hampered the switching process; nevertheless, performance of the reflexes suggested that the switching of cerebellar ascending influences to the cerebral cortex could be completed through other cerebellocortical pathways as well. Comparison of the results of electrolytic and chemical lesions of the red nucleus suggested a similar conclusion. It was established that the conditioning and recovery of already learned instrumental reflexes were impossible after complete neurotoxic destruction of the inferior olive. The data obtained emphasize the role of the inferior olive, ventrolateral thalamic nucleus and red nucleus in the switching of descending influences in operantly motor conditioned rats. Motor deficit and the compensatory rehabilitation process depended on the severity of inferior olive destruction combined with a high transection of the dorsolateral funiculus and a destroyed red nucleus. Long-lasting training improved compensation of motor deficit and stabilized instrumental reflexes to some extent in rats with incomplete destruction of the inferior olive. It has been suggested that these modifications occur because of collateral sprouting in the olivocerebellar system. PMID- 10854773 TI - Abnormalities in cellular adhesion of neuroblastoma and fibroblast models of Lesch Nyhan syndrome. AB - Lesch Nyhan syndrome is a neurological paediatric condition characterized by mental retardation, choreathotosis and self-mutilation. Biochemically, this condition has been attributed to a deficiency in the purine enzyme, hypoxanthine guanine phosphoribosyltransferase, however, the way this affects the development of the nervous system is still unknown. Ma et al.(15) and Stacey et al.(25) found that hypoxanthine guanine phosphoribosyltransferase-deficient neuroblastoma, differentiated significantly more than cells with this enzyme. Here, we report that adhesion of hypoxanthine guanine phosphoribosyltransferase-deficient neuroblastoma as well as fibroblasts from patients with Lesch Nyhan syndrome, exhibited dramatically enhanced adhesion compared to control cells. This increase in adhesion was dependent upon the cell type, density of the cells and upon the substrate used. Development of the nervous system is dependent on adhesion, in particular in the processes of migration, nucleation, differentiation and fasciculation. Our results suggest that the increased adhesion of hypoxanthine guanine phosphoribosyltransferase-deficient neuroblastoma and fibroblasts in vitro underpins the neuropathology of Lesch Nyhan syndrome. PMID- 10854774 TI - The continued evolution of two-hybrid screening approaches in yeast: how to outwit different preys with different baits. AB - The original two-hybrid system, an experimental approach designed to detect protein interactions, exploited the modular nature of many transcription factors. It has provided the intellectual and technical seed for the evolution of an array of innovative approaches, the application of which broadens the scope of experimentally feasible questions to include the interaction of proteins with diverse binding partners. The available array of modified and alternative approaches facilitates the analysis of complex cellular machinery and signaling networks that rely on multiple protein interactions. Such advances have facilitated the functional analysis of proteins on the genome level, a feat considered untenable a decade ago. PMID- 10854775 TI - Genomic heterogeneity of nucleotide excision repair. AB - Nucleotide excision repair (NER) is one of the major cellular pathways that removes bulky DNA adducts and helix-distorting lesions. The biological consequences of defective NER in humans include UV-light-induced skin carcinogenesis and extensive neurodegeneration. Understanding the mechanism of the NER process is of great importance as the number of individuals diagnosed with skin cancer has increased considerably in recent years, particularly in the United States. Rapid progress made in the DNA repair field since the early 1980s has revealed the complexity of NER, which operates differently in different genomic regions. The genomic heterogeneity of repair seems to be governed by the functional compartmentalization of chromatin into transcriptionally active and inactive domains in the nucleus. Two sub-pathways of NER remove UV-induced photolesions: (I) Global Genome Repair (GGR) and (II) Transcription Coupled Repair (TCR). GGR is a random process that occurs slowly, while the TCR, which is tightly linked to RNA polymerase II transcription, is highly specific and efficient. The efficiency of these pathways is important in avoiding cancer and genomic instability. Studies with cell lines derived from Cockayne syndrome (CS) and Xeroderma pigmentosum (XP) group C patients, that are defective in the NER sub-pathways, have yielded valuable information regarding the genomic heterogeneity of DNA repair. This review deals with the complexity of repair heterogeneity, its mechanism and interacting molecular pathways as well as its relevance in the maintenance of genomic integrity. PMID- 10854776 TI - Isolation and characterization of a mouse betaine-homocysteine S methyltransferase gene and pseudogene. AB - Betaine-homocysteine S-methyltransferase (BHMT) is one of the enzymes involved in the branch point metabolism of homocysteine. Elevated levels of plasma homocysteine may be a risk factor for the development of vascular disease; however, whether BHMT has a significant role in the regulation of plasma levels of homocysteine remains to be determined. As a prelude to creating a mouse strain deficient in BHMT activity, we screened a lambda library containing mouse SvJ 129 genomic DNA for the mouse BHMT gene using random probes made from the human cDNA. One genomic isolate was completely sequenced and found to encode an intronless BHMT pseudogene (mBHMT-ps). mBHMT-ps was then used as a template for the generation of random probes that were used to screen a BAC library containing mouse 129 Sv/Ev genomic DNA. In order to discriminate between pseudogenes and the authentic BHMT gene, a secondary PCR-based screen was employed which used primers designed from the pseudogene sequence that would predictably amplify across introns. Using this strategy, we isolated six mouse genomic clones that tested positive for the presence of all seven introns characteristic of the human gene, and the BHMT gene of one clone was completely sequenced. Like the human BHMT gene, the mouse gene spans 21kb and is encoded by eight exons interrupted by seven introns. The structure of the mouse BHMT gene is described herein as well as the 5'-flanking region of the gene adjacent to exon 1, which we demonstrate is capable of conferring basal promoter activity in Chinese Hamster Ovary cells. PMID- 10854777 TI - Comparative analysis of the RED1 and RED2 A-to-I RNA editing genes from mammals, pufferfish and zebrafish. AB - One type of RNA editing involves the deamination of adenosine (A) residues to inosines (I) at specific sites in specific pre-mRNAs. These inosines are subsequently read as guanosines by the ribosome, with potentially significant consequences for protein sequence. In mammals, two such A-to-I RNA editases are RED1, which edits some serotonin and glutamate receptors, and RED2, with unidentified substrates. To study the evolutionary conservation among these editases, we have isolated homologous genes from the Japanese pufferfish, Fugu rubripes. Fugu has two genes homologous to Red1 that are similar in size and organization and that show a fivefold compaction relative to the human gene; they differ, however, in their base compositional features. The Fugu gene for RED2 is unusually large, spanning more than 50kb; within the largest intron, there is evidence for a novel gene on the opposite strand. Because of these unusual features, the partial genomic structure was determined for the mouse RED2 gene. A partial cDNA for RED1 was also isolated from zebrafish. Comparisons between fish and between fish and mammals of the protein sequences show that the catalytic domains are highly conserved for each gene, while the RNA-binding domains vary within a single protein in their levels of conservation. Different levels of conservation among domains of different functional roles may reflect differences in editase substrate specificity and/or substrate sequence conservation. PMID- 10854778 TI - Comparative analysis of the DRADA A-to-I RNA editing gene from mammals, pufferfish and zebrafish. AB - The DRADA gene in mammals encodes an A-to-I RNA editase, an adenosine deaminase that acts on pre-mRNAs to produce site specific inosines. DRADA has been shown to deaminate specific adenosine residues in a subset of glutamate and serotonin receptors, and this editing results in proteins of altered sequences and functional properties. DRADA thus plays a role in creating protein diversity. To study the evolutionary significance of this gene, we have characterized the genomic structure of DRADA from Fugu rubripes, and compared the protein sequences of DRADA from mammals, pufferfish and zebrafish. The DRADA gene from Fugu is three-fold compacted with respect to the human gene, and contains a novel intron within the large second coding exon. DRADA cDNAs were isolated from zebrafish and a second pufferfish, Tetraodon fluviatilis. Comparisons among fish, and between fish and mammals, of the protein sequences show that the catalytic domains are highly conserved for each gene, while the RNA binding domains vary within a single protein in their levels of conservation. Conservation within the Z DNA binding domain has also been assessed. Different levels of conservation among domains of different functional roles may reflect differences in editase substrate specificity and/or substrate sequence conservation. PMID- 10854779 TI - Primary structural features of the 20S proteasome subunits of rice (Oryza sativa). AB - The 20S proteasome is the proteolytic complex that is involved in removing abnormal proteins, and it also has other diverse biological functions. Its structure comprises 28 subunits arranged in four rings of seven subunits, and exists as a hollow cylinder. The two outer rings and two inner rings form an alpha7beta7beta7alpha7 structure, and each subunit, alpha and beta, exists as seven different types, thus giving 14 kinds of subunits. In this study, we report the primary structures of the 14 proteasomal subunit subfamilies in rice (Oryza sativa), representing the first set for all of the subunits from monocots. Amino acid sequence homology within the rice family (alpha-type: 28.9-42.1%; beta-type: 17.2-31. 9%) were lower than those between rice subunits and corresponding orthologs from Arabidopsis and yeast (alpha-type: 49.2-94.5%; beta-type: 34.8 87.7%). Structural features observed in eukaryotic proteasome subunits, i.e., alpha- or beta-type signature at the N-termini, Thr active sites in beta1, beta2 and beta5 subunits, and nuclear localization signal-like sequences in some alpha type subunits, were shown to be conserved in rice. PMID- 10854780 TI - Genomic structure and expression of parathyroid hormone-related protein gene (PTHrP) in a teleost, Fugu rubripes. AB - In this study we describe the isolation and characterisation of the parathyroid hormone-related protein (PTHrP) gene from the teleost Fugu rubripes. The gene has a relatively simple structure, compared with tetrapod PTHrP genes, composed of three exons and two introns, encompassing 2.25kb of genomic DNA. The gene encodes a protein of 163 amino acids, with a putative signal peptide of 37 amino acids and a mature peptide of 126 amino acids. The overall homology with known tetrapod PTHrP proteins is low (36%), with a novel sequence inserted between positions 38 and 65, the absence of the conserved pentapeptide (TRSAW) and shortened C terminal domain. The N-terminus shows greater conservation (62%), suggesting that it may have a hypercalcaemic function similar to that of tetrapod PTHrP. In situ localisation and RT-PCR have demonstrated the presence of PTHrP in a wide range of tissues with varying levels of expression. Sequence scanning of overlapping cosmids has identified three additional genes, TMPO, LDHB and KCNA1, which map to human chromosome 12, with the latter two mapping to 12p12-11.2. PTHrP in human also maps to this chromosome 12 sub-region, thus demonstrating conservation of synteny between human and Fugu. PMID- 10854781 TI - Drad21, a Drosophila rad21 homologue expressed in S-phase cells. AB - Cohesin is an evolutionarily conserved multiprotein complex required to establish and maintain sister chromatid cohesion. Here, we report the cloning and initial characterization of the Drosophila homologue of the fission yeast rad21 cohesin subunit, called Drad21. The Drad21 coding region was localized to centromeric heterochromatin and encodes a 715 amino acid (aa) protein with 42% aa identity to vertebrate Rad21p-homologues, 25% with Scc1p/Mcd1p (S. cerevisiae) and 28% with Rad21p (S. pombe). Sequences with similarity to the sites of proteolytic cleavage identified in Scc1p/Mcd1p are not evident in DRAD21. Northern blot and mRNA in situ studies show that Drad21 is developmentally regulated, with high levels of expression in early embryogenesis, in S-phase cells of proliferating imaginal tissues, and in the early endocycling cells of the embryonic gut. PMID- 10854782 TI - Identification of a 3.2 kb 5'-flanking region of the murine keratocan gene that directs beta-galactosidase expression in the adult corneal stroma of transgenic mice. AB - The mouse keratocan gene (Ktcn) expression tracks the corneal morphogenesis during eye development and becomes restricted to keratocytes of the adult, implicating a cornea-specific gene regulation of the mouse Ktcn [J. Biol. Chem., 273 (1998) 22584-22588]. To examine the functionality of the mouse Ktcn promoter, we have cloned and sequenced a 3.2kb genomic DNA fragment 5' of the mouse Ktcn gene, which was used to prepare a reporter gene construct that contained the 3.2kb 5' flanking sequence, exon 1 and 0.4kb of intron 1 of Ktcn, and beta-geo hybrid reporter gene. The beta-galactosidase (betaGal) activity was assayed in tissues of two of five transgenic mouse lines obtained via microinjection. In adult transgenic mice, betaGal activity was detected only in cornea, not in other tissues (e.g. lens, retina, sclera, lung, heart, liver, diaphragm, kidney, and brain). During ocular development, the spatial-temporal expression patterns of the betaGal recapitulated that of endogenous Ktcn in transgenic mice. Using XGal staining, strong betaGal activity was first detected in periocular tissues of E13.5 embryos, and restricted to corneal keratocytes at E14.5 and thereafter. Interestingly, in addition to cornea, betaGal activity was transiently found in some non-ocular tissues, i.e. ears, snout, and limbs of embryos of E13.5 and E14.5 but was no longer detected in those tissues of E16.5 embryos. The transient expression of endogenous keratocan in non-ocular tissues during embryonic development was confirmed by in situ hybridization. Taken together, our results suggest that the 3.2kb Ktcn promoter contains sufficient cis-regulatory elements to drive heterologous minigene expression in cells expressing keratocan. The identification of keratocyte-specific expression of betaGal reporter gene in the adult transgenic mice is an important first step in characterizing the Ktcn promoter in order to use it to drive a foreign gene expression in corneal stroma. PMID- 10854783 TI - Cloning and characterization of the S1 domain of four myosin isoforms from functionally divergent fiber types in adult Rana pipiens skeletal muscle. AB - The motor properties of myosin reside in the globular S1 region of the myosin heavy chain (MHC) subunit. All vertebrates express a family of MHC isoforms in skeletal muscle that have a major influence on the mechanical properties of the various fiber types. Differences in molecular composition of S1 among MHC isoforms within a species have not been studied to any great detail. Presently, we have isolated, cloned and sequenced the S1 subunit of four MHC isoforms from skeletal muscle in Rana pipiens that are specifically expressed in four mechanically divergent fiber types. Paired analysis showed that the overall amino acid identity was higher between the three S1 isoforms expressed in twitch fibers than between the twitch and tonic isoforms. Relatedness in amino acid composition was evaluated in regions reported to govern cross-bridge kinetics. Surface loops 1 and 2, thought to influence motor velocity and ATPase, respectively, were both highly divergent between isoforms. However, the divergence in the loops was roughly equal to that of the amino-terminal region, a domain considered less important for motor function. We tested the hypothesis that the loops are more conserved in pairs of isoforms with more similar kinetics. Comparisons including other vertebrate species showed no tendency for loops from pairs with similar kinetics to be more conserved. These data suggest that the overall structure of loops 1 and 2 is not critical in regulating the kinetic properties of R. pipiens S1 isoforms. Cloning of this family of frog S1 isoforms will facilitate future structure/function studies of the molecular basis of variability in myosin cross bridge kinetics. PMID- 10854784 TI - Recent amplification of miniature inverted-repeat transposable elements in the vector mosquito Culex pipiens: characterization of the Mimo family. AB - We describe a new family of repetitive elements, named Mimo, from the mosquito Culex pipiens. Structural characteristics of these elements fit well with those of miniature inverted-repeat transposable elements (MITEs), which are ubiquitous and highly abundant in plant genomes. The occurrence of Mimo in C. pipiens provides new evidence that MITEs are not restricted to plant genomes, but may be widespread in arthropods as well. The copy number of Mimo elements in C. pipiens ( approximately 1000 copies in a 540Mb genome) supports the hypothesis that there is a positive correlation between genome size and the magnitude of MITE proliferation. In contrast to most MITE families described so far, members of the Mimo family share a high sequence conservation, which may reflect a recent amplification history in this species. In addition, we found that Mimo elements are a frequent nest for other MITE-like elements, suggesting that multiple and successive MITE transposition events have occurred very recently in the C. pipiens genome. Despite evidence for recent mobility of these MITEs, no element has been found to encode a protein; therefore, we do not know how they have transposed and have spread in the genome. However, some sequence similarities in terminal inverted-repeats suggest a possible filiation of some of these mosquito MITEs with pogo-like DNA transposons. PMID- 10854785 TI - Identification of mammalian-like purple acid phosphatases in a wide range of plants. AB - Purple acid phosphatases (PAPs) comprise a family of binuclear metal-containing hydrolases, members of which have been isolated from plants, mammals and fungi. Polypeptide chains differ in size (animal approximately 35kDa, plant approximately 55kDa) and exhibit low sequence homology between kingdoms but all residues involved in co-ordination of the metal ions are invariant. A search of genomic databases was undertaken using a sequence pattern which includes the conserved residues. Several novel potential PAP sequences were detected, including the first known examples from bacterial sources. Ten plant ESTs were also identified which, although possessing the conserved sequence pattern, were not homologous throughout their sequences to previously known plant PAPs. Based on these EST sequences, novel cDNAs from sweet potato, soybean, red kidney bean and Arabidopsis thaliana were cloned and sequenced. These sequences are more closely related to mammalian PAP than to previously characterized plant enzymes. Their predicted secondary structure is similar to that of the mammalian enzyme. A model of the sweet potato enzyme was generated based on the coordinates of pig PAP. These observations strongly suggest that the cloned cDNA sequences represent a second group of plant PAPs with properties more similar to the mammalian enzymes than to the high molecular weight plant enzymes. PMID- 10854786 TI - Structures of six cDNAs expressed specifically at cypris larvae of barnacles, Balanus amphitrite. AB - We cloned six cDNAs by screening cDNA libraries of cypris larvae from barnacles, Balanus amphitrite, and studied their expression by Northern blot analysis. All of them are expressed in the cypris larvae at the settlement stage, but not in the earlier nauplii larvae nor in later adult barnacles. Therefore, we designated them as barnacle cypris larva-specific genes (bcs); bcs-1, bcs-2, bcs-3, bcs-4, bcs-5 and bcs-6. During the process of larval attachment and metamorphosis, the amounts of bcs-1 and bcs-2 mRNAs decreased, whereas the bcs-3, bcs-4, bcs-5 and bcs-6 mRNAs increased. A homology search showed that all cDNAs encode novel peptides containing characteristic amino acid sequences. This study strongly suggests that these bcs gene products are involved in the cypris larval attachment and metamorphosis of barnacles. PMID- 10854787 TI - Characterization of rat CD14 promoter and its regulation by transcription factors AP1 and Sp family proteins in hepatocytes. AB - CD14, a 55kDa glycoprotein, serves as a lipopolysaccharide (LPS) recognition molecule. CD14 is a monocyte differentiation antigen expressed by myeloid-derived cells, or other cells such as hepatocytes, as either a membrane-bound protein or a soluble serum protein. Increasing evidence indicates that soluble CD14 in plasma is an acute-phase protein derived, among other sources, from liver cells. Although information is available on the cellular expression of CD14, little is known about the cis- and trans-acting factors that regulate basal CD14 transcription in liver cells. We show here that liver cells have a relatively high basal CD14 transcription rate as determined by nuclear run-on assay. We cloned and sequenced an 883bp 5'-flanking region of the rat CD14 gene and demonstrated functional promoter activity in liver cells. Sequence analysis revealed that, like in the human and mouse CD14 genes, multiple Sp1 and AP1 binding elements exist in rat CD14. Site-directed mutagenesis and transient transfection assays demonstrated that an Sp1 element located at -836 and an AP1 element located at -270 are required for basal promoter activity in liver cells. Electrophoretic mobility shift assays indicate that both Sp1 and Sp3 nuclear factors interact with the -836 Sp1 element, while the AP1-related proteins Fra-2 and JunD bind to the AP1 motif. These data provide novel insights into the regulation of basal CD14 expression in liver cells. PMID- 10854788 TI - Expression pattern of Rhizobium etli ccmIEFH genes involved in c-type cytochrome maturation. AB - In different bacterial species, ccmIEFH genes have been suggested to code for subunits of a bacterial haem-lyase catalyzing the covalent attachment of haem to c-type apoproteins. In Rhizobium etli CE3 there are two copies of ccmIEFH: one in the chromosome and the other located in plasmid pf. However, the null phenotype of chromosomal ccmF mutant indicates that the gene locus of plasmid pf is not functional. Two ccmI chromosomal mutants, previously isolated, produced detectable levels of c-type cytochromes under certain culture conditions in contrast with the ccmF mutant, suggesting that ccmF could be transcribed independently. The transcriptional organization of ccmIEFH operon was established. Two promoters from the chromosomal locus were mapped by primer extension, one located upstream of ccmI and the second located upstream of ccmF. The regulation of the expression of both promoters was studied using appropriate lacZ gene fusions (ccmI-lacZ and ccmEF-lacZ). The ccmI-lacZ gene fusion was expressed in complex medium, during exponential growth, under microaerobic conditions and in a R. etli mutant that accumulates reducing power, conditions where a higher respiration rate could be limited by c-type cytochrome content. The ccmEF-lacZ fusion was also primarily expressed in complex medium and under microaerophilic conditions. The finding of two independent promoters in this gene locus could suggest that the step catalyzed by CcmFH could be a rate-limiting step for c-type cytochrome assembly under certain culture conditions. PMID- 10854789 TI - Characterization of CaGSP1, the Candida albicans RAN/GSP1 homologue. AB - Gsp1p is a small nuclear-located GTP binding protein from the yeast Saccharomyces cerevisiae. It is highly conserved among eucaryotic cells and is involved in numerous cellular processes, including nucleocytoplasmic trafficking of macromolecules. To learn more about the GSP1 structure/function, we have characterized its Candida albicans homologue. CaGsp1p is 214 amino acids long and displays 91% identity to the ScGsp1p. There is functional complementation in S. cerevisiae, and its mRNA is constitutively expressed in the diploid C. albicans grown under various physiological conditions. Disruption of both alleles was not possible, suggesting that it could be an essential gene, but heterozygous mutants exhibited genomic instability. PMID- 10854790 TI - Cloning and chromosomal localization of the human BARX2 homeobox protein gene. AB - The human BARX2 gene encodes a homeodomain-containing protein of 254 amino acids, which binds optimally to the DNA consensus sequence YYTAATGRTTTTY. BARX2 is highly expressed in adult salivary gland and is expressed at lower levels in other tissues, including mammary gland, kidney, and placenta. The BARX2 gene consists of four exons, and is located on human chromosome 11q25. This chromosomal location is within the minimal deletion region for Jacobsen syndrome, a syndrome including craniosynostosis and other developmental abnormalities. This chromosomal location, along with the reported expression of murine barx2 in craniofacial development, suggests that BARX2 may be causally involved in the craniofacial abnormalities in Jacobsen syndrome. PMID- 10854791 TI - Genomic characterisation and fine mapping of the human SOX13 gene. AB - SOX13 is the member of the SOX (Sry related HMG BOX) family of transcription factors which encodes the type-1 diabetes autoantigen, ICA12, and is expressed in a number of tissues including pancreatic islets and arterial walls. By fluorescence in situ hybridisation, radiation hybrid mapping and YAC analysis we determined that the human SOX13 gene maps to Chromosome 1q31.3-32.1 near the marker D1S504, a region associated with type-1 diabetes susceptibility and familial dilated cardiomyopathy. Mouse Sox13 maps to the syntenic region near the marker D1Mit57. The human SOX13 gene spans >15.5kb of genomic DNA and is composed of 14 exons with introns interrupting regions encoding the HMG DNA binding domain and the leucine zipper/glutamine-rich dimerisation domain. Comparison with the mouse Sox13 gene suggests the existence of long and short forms of the SOX13 protein which may arise by differential splicing during different stages in embryogenesis. The high sequence conservation between human SOX13 and mouse, Xenopus and trout orthologues implies a conserved function in vertebrates. SOX13 belongs to SOX Group D members which contain a leucine zipper/glutamine-rich region. Phylogenetic analyses of SOX proteins suggest that such domains were acquired after the initial divergence of groups A to G. PMID- 10854792 TI - DNA sequence and functional characterization of the human and rat epidermal growth factor promoter: regulation by cell growth. AB - Epidermal growth factor (EGF) regulates cell growth and differentiation through intracellular transduction networks activated by its tyrosine kinase receptor, EGFR. In this report we describe the structure and DNA sequence of transcriptional control regions from both human and Wistar-rat single copy EGF genes and their functional analysis in epithelial cell cultures. By sequence comparison we show these proximal gene regions have remained conserved in evolution to -640 (relative to the rodent mRNA initiation site), where similarity is interrupted by a rodent interspersed-repeat element (SINE). Transcript mapping reveals complexity in EGF initiation site selection: whereas a single rat liver initiation site (+1) appears 30bp 3' to the TTTAA element, an additional upstream site is detected in kidney RNA at -14. In contrast, in human RNA a single initiation is observed, which is displaced 12bp 3' to the rodent RNA terminus. Both promoters were defined in transient expression assays. Our results show the human promoter to be at least 20-fold more active than the equivalent rodent sequence, although both are activated during cell proliferation and negatively regulated in contact inhibited and quiescent cultures. The results indicate EGF gene expression and cell division are temporally linked, suggesting its promoter comprises a growth responsive regulatory domain. PMID- 10854793 TI - Isolation of a functional copy of the human BRCA1 gene by transformation associated recombination in yeast. AB - The BRCA1 gene, mutations of which contribute significantly to hereditary breast cancer, was not identified in the existing YAC and BAC libraries. The gene is now available only as a set of overlapping fragments that form a contig. In this work we describe direct isolation of a genomic copy of BRCA1 from human DNA by transformation-associated recombination (TAR) cloning. Despite the presence of multiple repeats, most of the primary BRCA1 YAC isolates did not contain detectable deletions and could be stably propagated in a host strain with conditional RAD52. Similar to other circular YACs, approximately 90kb BRCA1 YACs were efficiently and accurately retrofitted into bacterial artificial chromosomes (BACs) with the Neo(R) mammalian selectable marker and transferred as circular BAC/YACs in E. coli cells. The BRCA1 BAC/YAC DNAs were isolated from bacterial cells and were used to transfect mouse cells using the Neo(R) gene as selectable marker. Western blot analysis of transfectants showed that BRCA1 YACs isolated by a TAR cloning contained a functional gene. The advantage of this expression vector is that the expression of BRCA1 is generated from its own regulatory elements and does not require additional promoter elements that may result in overexpression of the protein. In contrast to the results with cDNA expression vectors, the level of BRCA1 expression from this TAR vector is stable, does not induce cell death, maintains serum regulation, and approximates the level of endogenously expressed BRCA1 in human cells. The entire isolation procedure of BRCA1 described in this paper can be accomplished in approximately 10 days and can be applied to isolation of gene from clinical material. We propose that the opportunity to directly isolate normal and mutant forms of BRCA1 will greatly facilitate analysis of the gene and its contribution to breast cancer. PMID- 10854794 TI - Isolation and characterization of medaka ribosomal protein S3a (fte-1) cDNA and gene. AB - Many ribosomal protein genes were cloned from different organisms. We describe here, for the first time, the isolation of the ribosomal protein S3a cDNA and gene from a teleost - the medaka (Oryzias latipes). The cDNA sequence is 863bp long and encodes an open reading frame of 266 amino acids. The gene is 2927bp long and contains six introns and five introns. The levels of the S3a mRNA are elevated during embryonal development. Transcription of the gene was also detected in different tissues of adult medaka. At the 5' untranslated region of the cDNA, the terminal pyrimidine tract, a common feature with all ribosomal protein genes, was found. snoRNA sequences were found in introns 3 and 5, similar to human and mouse U73b and U73a. PMID- 10854795 TI - Longitudinal follow-up of children born preterm: cognitive development at age 19. AB - In a long-term prospective study, 39 preterm children born before 35 completed weeks of gestation and 23 full-term children were followed up at 4, 9 and 19 years of age. Psychometric evaluation of the cognitive development at 4 years of age showed that the preterms fell within the normal range, although their performance was inferior to that of the full-terms. This difference between the groups was not found at 9 and 19 years of age. Within the preterm group there was no correlation between the test results and birthweight, gestational age, prenatal or perinatal optimality scores. Full-terms had better scholastic performance at the end of compulsory schooling, while there was no difference at 19 years of age. At 19 years of age, about 1/3 of the children in both groups rated themselves as having had attention deficits during their childhood and adolescence. In this group of moderately immature, low-risk children, preterm birth without major physical or mental disabilities poses a developmental risk that seems to have the greatest impact during the preschool years and then gradually attenuates. PMID- 10854796 TI - Abnormal cerebral neuronal migration in a rat model of intrauterine growth retardation induced by synthetic thromboxane A(2). AB - Many reports have associated intrauterine growth retardation (IUGR) with adverse neurological outcome, but the underlying pathology is imperfectly understood. We have developed a new rat model of IUGR using maternal administration of synthetic thromboxane A(2) (STA(2)). In the present study, the effect of this insult on neuronal migration in the rat cerebral cortex was examined. Bromodeoxyuridine (BrdU), a time-specific cell marker was administered intraperitoneally to the mothers on embryonic day (E) 19. At postnatal day (P) 3, P4, P5, and P6, pups were terminally anesthetized and brains were removed. BrdU-labeled cells were detected immunohistochemically and counted in cerebrum, which was divided into the cortical plate (CP), the intermediate zone, and the subventricular/ventricular zone (SVZ+VZ). Numbers of labeled cells in the three areas over time were compared between IUGR and control animals. Numbers of labeled cells in SVZ+VZ were significantly greater in IUGR than in controls at P3, 5, and 6 (P<0.05). In contrast, labeled cells in the CP were significantly less abundant in IUGR animals than in controls at P3, 4, and 6 (P<0.05). We concluded that neuronal migration was delayed in IUGR rats. PMID- 10854797 TI - Underexpression of neural cell adhesion molecule and neurotrophic factors in rat brain following thromboxane A(2)-induced intrauterine growth retardation. AB - Intrauterine growth retardation (IUGR) often results in clinical neurodevelopmental disorders. To clarify the influence of uteroplacental insufficiency on central nervous system development, we have created a model of IUGR in rats using maternal administration of synthetic thromboxane A(2). We investigated expression patterns of neural cell adhesion molecule (NCAM) and reelin in this model by semiquantitative competitive polymerase chain reactions. On postnatal day 2, NCAM expression was decreased in rat cerebral cortex, and reelin expression was decreased in hippocampus from levels in controls without maternal thromboxane exposure. No significant differences in NCAM expression were seen in hippocampus, nor did reelin expression differ in cerebral cortex between control and IUGR groups. We also examined expression of two neurotrophic factors, brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3). In cerebral cortex the IUGR group showed less BDNF and NT-3 expression than controls. Delay of neuronal migration and histological changes observed in our IUGR rats may be related to altered expression of these molecules. PMID- 10854798 TI - Bilateral basal ganglia-thalamic lesions subsequent to prolonged fetal bradycardia. AB - We report two infants with bilateral basal ganglia-thalamic lesions subsequent to prolonged fetal bradycardia. Cardiotocogram revealed severe bradycardia lasting for more than 20 min in both. They demonstrated a significant encephalopathy, abnormal muscle tones and signs of brainstem injury. Clinical or electrical seizures were not observed in either of them. CT during early neonatal period demonstrated decreased tissue attenuation in basal ganglia and thalami in the absence of extensive cortical changes. Both of them developed severe mental retardation and quadriplegia. MRI in late infancy demonstrated abnormal high intensity areas in bilateral basal ganglia, thalami and around central sulci on T2-weighted image. Close correlation between prolonged fetal bradycardia and basal ganglia-thalamic lesion was suggested. PMID- 10854799 TI - Vasoconstriction following spontaneous sighs and head-up tilts in infants sleeping prone and supine. AB - The cutaneous vasoconstrictor responses following a 60 degrees head-up tilt and a spontaneous sigh were measured in 36 infants at 1 and 3 months age to investigate the effects of age, sleep state and sleep position on these responses. The vasoconstrictor response was determined by a measure of cutaneous blood flow using a laser Doppler flowmeter. The mean reduction in blood flow (vasoconstriction) was 52% following the tilt, and 33% following the sigh. Prone positioning 1-month-old infants as compared to supine, reduced the degree of vasoconstriction following the tilt (P=0.027) and sigh (P=0.026). The supine to prone reduction was: tilt, -11% in quiet sleep (QS) (from 55.1 to 49.1% vasoconstriction) and -18% in active sleep (AS) (from 52.0 to 42.9%) and; sigh, 26% in QS (35-26%), and -15% in AS (31-26%). The degree of vasoconstriction following the sigh was significantly greater in 3- compared to 1-month-old infants (+26%, P=0.040). The mean response to the tilt in the older age group was 12% greater but this did not reach significance (P=0.069). Sleep state did not affect the degree of vasoconstriction but influenced transmission of the response so that latency to minimal vasoconstriction was 1 s shorter in AS than QS. This study provides data on two simple measures of sympathetic activity during sleep that have not previously been described in any detail in infant studies, and add more evidence that autonomic activity is reduced in the prone position compared to supine during sleep. PMID- 10854800 TI - Crying and behavior pattern in breast- and formula-fed infants. AB - Breast- and formula-fed infants were compared with regard to behavior patterns, especially crying behavior. A diary concerning seven behaviors was completed by 188 mothers of breast- and formula-fed infants at the well baby clinic. Breast fed infants were fed more frequently with longer feeding duration. They slept more, but their long bouts were shorter than those of formula-fed infants. The crying pattern of formula-fed infants was different from that of breast-fed infants in that they displayed an evening cluster and a 7-week peak of crying. Since the educational level of the formula group mothers was higher and there are more later born babies in that group, it is speculated that these mothers have a Westernized tendency in their caretaking style and, as a result, the crying pattern of formula-fed infants are similar to that of Western babies. In multivariate analysis, contact, play, sleep durations and mothers' education contributed significantly to cry duration. These results imply that contact with the mother and other caretaking practices are closely associated with infants' crying. PMID- 10854801 TI - Neurophysiological brainstem investigations in isolated Pierre Robin sequence. AB - Polysomnography, electromyography (EMG) of the face, tongue, and soft palate, blink reflexes (BRs), EMG during bottle-feeding, and brainstem auditory evoked responses (BAERs) were performed in 25 newborn babies with isolated Pierre Robin sequence (PRS) to aid in evaluation and management. Obstructive apneas were found in 23/24 patients (the 25th having undergone tracheotomy). Number and duration of central respiratory pauses were always normal, as well as electroencephalographic and clinical organization of sleep stages. EMG recruitment pattern in facial and lingual muscles, and BRs were normal in all cases. EMG recruitment pattern in muscles of the soft palate was normal in 14/25 patients, showed a reduced average amplitude with short-duration and low amplitude motor unit potentials in 10/25, and showed signs of denervation in 1/25. EMG during bottle-feeding showed sucking swallowing disorders in 20/25 patients. BAERs showed a bilateral conductive impairment with increased latencies and thresholds in 5/19 patients, but with normal and symmetric I-III and I-V interpeak latencies in 19/19. These neurophysiological findings suggest that in isolated PRS a dysfunction of the lingual and pharyngeal motor organization exists without any structural impairment in brainstem nuclei and pathways. PMID- 10854802 TI - The relationship between the response to external light stimulation and behavioral states in the human fetus: how it differs from vibroacoustic stimulation. AB - OBJECTIVE: To determine the effects of external light stimulation on fetal behavioral states and know the difference from those of vibroacoustic stimulation. METHODS: A flashlight and a vibroacoustic stimulator was applied directly on the maternal abdomen to determine the response of 56 normal fetuses at 36-40 weeks gestation. Fetal heart rate (FHR) and body movements were recorded using an actocardiograph, and fetal eye movements were observed using real-time ultrasonography. Using Nijhuis's criteria, the fetal behavioral states (1F-4F) were determined. FHR acceleration was considered a fetal response to the stimulations. RESULTS: The lag time between stimulation and fetal response was within 4 s. A positive response rate to the light stimulation was higher at behavioral states 2F (82%) and 3F (83%) than at state 1F (4%). Light stimulation changed the behavioral state of two of the six 3F fetuses (33%) from 3F to 4F. No change of state was observed in fetuses that were in states 1F and 2F. For vibroacoustic stimulation, fetal response was 100% positive and changes of states were observed frequently irrespective of the behavioral state before the stimulation. CONCLUSION: Fetal response to light stimulation is closely connected to fetal behavioral states and may reflect some distinct stages of the sleep wakefulness cycle. PMID- 10854803 TI - Criteria used when initiating antifungal therapy against Candida spp. in the intensive care unit. AB - Invasive candidiasis is a life threatening complication for intensive care unit (ICU) patients. The infection is difficult to recognise so that treatment may be delayed or even not given. Risk factors for candidiasis include the use of antimicrobial agents, central intravascular devices (mainly Hickmann catheters), recurrent gastrointestinal perforations, surgery for acute pancreatitis or splenectomy and renal dysfunction or haemodialysis. Therapy against Candida spp is recommended in ICU patients with endophthalmitis or chorioretinitis possibly caused by Candida spp., in symptomatic patients with risk factors for invasive candidiasis especially if two or more anatomical sites are colonised and for asymptomatic high-risk surgical patients (with recent abdominal surgery or recurrent gastrointestinal perforations or anastomotic leakages). The isolation of Candida from any site poses an increased risk but there are a few microbiological data that might help to establish the predictive value of a particular isolate. These include the site of isolation, the number of culture positive, noncontigous sites, the density of colonisation and the species isolated. Antifungals should be started when Candida spp. are recovered from blood cultures or from usually sterile body fluids, abscesses or wounds in burns patients. They should also be considered in patients with a colonisation index >0.5 or a corrected colonization index >0.4 or when the isolate is identified as Candida tropicalis. PMID- 10854804 TI - Toward multinational antimicrobial resistance surveillance systems in Europe. AB - While there is a growing concern about increasing antimicrobial resistance and international spread of resistant microorganisms, we are still lacking timely multinational, good-quality susceptibility data to guide our decisions on controlling such resistance. This review describes and compares current sources of multicentric antimicrobial susceptibility data, identifies problems responsible for the postponing of the implementation of epidemiological antimicrobial resistance surveillance systems and finally presents requirements for such systems. PMID- 10854805 TI - Macrolides and clindamycin suppress the release of Shiga-like toxins from Escherichia coli O157:H7 in vitro. AB - We investigated the effects of antimicrobial agents fosfomycin (FOS), cefdinir (CDIN), levofloxacin (LEVX), rokitamycin (ROK), roxithromycin (ROX), and clindamycin (CLI) on the release of Shiga-like toxins (SLTs) by enterohaemorrhagic Escherichia coli (EHEC). EHEC was cultured for 14 h in the presence of ROX, ROK or CLI at sub-minimum inhibitory concentrations (subMICs) of 1.56-6.25 mg/l, followed by assay of the level of SLTs in the supernatants using cytotoxicity assay and reversed passive latex agglutination method. Exposure to ROX, ROK or CLI reduced the amount of released SLTs compared with untreated control cultures (P<0.05). These agents however, did not decrease the number of viable EHEC, indicating the importance of bactericidal agents. When the bacteria was exposed to CDIN, FOS or LEVX, the level of SLTs in the culture supernatant increased with the destruction of bacterial cells in the order of CDIN, FOS, LEVX. When 0.5 mg/l of LEVX was added to cultures with or without pretreatment using ROX, ROK, or CLI, the release of SLTs was reduced by this pretreatment (P<0.05). These results may have clinical implications for the treatment of EHEC infection. PMID- 10854806 TI - In vitro efficacy of six cephalosporins tested against Enterobacteriaceae isolated at 38 North American medical centres participating in the SENTRY Antimicrobial Surveillance Program, 1997-1998. AB - The SENTRY Antimicrobial Surveillance Program is an ongoing international collaboration that monitors the predominant bacterial and fungal pathogens and antimicrobial susceptibility patterns associated with community-acquired and nosocomial infections. SENTRY data on the current cephalosporin susceptibility patterns (1997-98) of North American isolates of clinically important Enterobacteriaceae were analyzed. Susceptibility to a selection of cephalosporins was assessed at a central laboratory using reference broth microdilution methods and interpretive criteria specified by the National Committee for Clinical Laboratory Standards. The third- and fourth-generation cephalosporins tested demonstrated excellent activity against Escherichia coli and Klebsiella pneumoniae, whereas some of the older agents maintained good efficacy. Extended spectrum beta-lactamases were detected in all regions of the United States and Canada (1.8-10.7%). Cefepime was the most active agent tested against pathogens with the potential for enzyme-mediated resistance due to Amp C. The third generation agents maintained acceptable efficacy against Serratia marcescens, but were less effective against Citrobacter and Enterobacter species. The older cephalosporins were generally inadequate against these pathogens, in contrast to cefepime, which was the widest spectrum cephalosporin overall. Some significant regional variations in spectrum were detected. PMID- 10854807 TI - Efficacy of continuous infusion of ceftazidime for patients with neutropenic fever after high-dose chemotherapy and peripheral blood stem cell transplantation. AB - Neutropenia is an important complication of high-dose chemotherapy (HDCT). Neutropenic patients presenting with fever are routinely hospitalized for treatment with broad-spectrum antibiotics. Neutropenia up to 10 days is associated with a low-risk profile, and antimicrobial therapy administered on an outpatient basis might be an alternative to admission to hospital. This prospective study evaluates the safety of a continuous infusion of ceftazidime in neutropenic patients after HDCT and peripheral blood stem cell transplantation (PBSCT). From September 1995 to October 1999, 81 patients received a 2 g intravenous bolus of ceftazidime, followed by a 4 g continuous infusion per 24 h of ceftazidime using a portable infusion pump. If the fever persisted for 72 h, a glycopeptide antibiotic was added. The median patients' age was 44 years. Fifty two of 81 patients (64%) responded to the monotherapy with ceftazidime. After addition of a glycopeptide antibiotic, a further 17 patients (21%) became afebrile. The causes of fever were septicaemia in 11 patients, pneumonia in two and fever of unknown origin in 68 patients. Fifty-eight episodes (72%) were successfully managed by outpatient treatment alone. The reason for admission to hospital was the change to imipenem/cilastin, which had to be administered three times per day (12 patients), severe mucositis with parenteral nutrition (eight patients), or a Karnovsky index 4) was observed with any combination. Synergy by time kill curve (present in 21 combinations) was more often seen at 24 h than 2 or 5 h (P<0.001). On comparing the mean of the FIC and FBC indices, there were significant differences in only four chequerboards (P<0.05). The same checkerboard was repeated on 3 separate days to test reproducibility. There were no significant differences (P>0.05). All combinations showing synergism by FBC index were synergic by FIC index. Synergy by FIC index predicted synergy by FBC index in 67%. All combinations showing synergism by FIC index were synergic by time kill at 24 h but there was poor correlation between synergy at 2 or 5 h and synergy by FIC index or FBC index. In conclusion combining time kill and chequerboard tests gives reproducible results and good correlation between FIC and FBC indices. FIC indices showing synergy were also predictive of synergy in time kill studies. For bactericidal combinations unlikely to be antagonistic, calculation of FIC index may be a good indicator of synergic bactericidal activity. PMID- 10854809 TI - Evaluation of rational antibiotic use. AB - The emergence of antibiotic resistant bacteria is a major problem throughout the world and a rational use of antibiotics is therefore very important. This study was performed to estimate the appropriateness of antimicrobial drug use in Celal Bayar University Hospital in Manisa. The data of all inpatients (n=937) between October and December 1998 were collected according to the Kunin and Jones criteria. Of the patients, 16.6% (n=156) were receiving antibiotics, and in 63.5, 23.0 and 13.5% of these, a single, two and three agents were used, respectively. The purpose of antibiotic use was for prophylaxis in 23.9%, as an empiric decision in 71.4% and for therapeutic culture-based reasons in 4.7%. The rate of rational antibiotic use was 45.7% and it was statistically higher in those patients from whom specimens had been taken for culture than in patients receiving prophylactic or empiric antibiotics. On medical wards, rational antibiotic usage was 55.1%, while it was 26.3% in surgical wards (P<0.0001). The low rate of appropriate antibiotic use in our university hospital reflects the urgent need of rationalization. PMID- 10854810 TI - Surveillance of antimicrobial resistance in Escherichia coli strains isolated from pigs at Spanish slaughterhouses. AB - Antimicrobial resistance can make the efficient treatment of bacterial infections in humans and animals more difficult. Antimicrobial use in food animals may be one of the factors contributing to resistance. The Spanish surveillance network VAV has established a baseline of antimicrobial resistance in Escherichia coli strains from healthy pigs. Minimum inhibitory concentration and patterns of resistance to antimicrobials used in animals and humans were determined for 205 faecal strains isolated in a sampling frame of four slaughterhouses in Spain from 220 pigs in 1998. Higher levels of resistance were seen against antimicrobial agents authorised for use in food animals especially tetracycline, sulphonamides, trimethoprim and amoxycillin. All isolates were susceptible to antimicrobials employed mainly in humans such as ceftazidime, cefotaxime, imipenem, aztreonam and amikacin. PMID- 10854811 TI - Nosocomial bacterial and fungal meningitis in children; an eight year national survey reporting 101 cases. Pediatric Nosocomial Meningitis Study Group. AB - One hundred and one cases of nosocomial meningitis in children from a national survey over 8 years have been analyzed for risk factors and outcome. From 101 cases, 115 organisms were isolated. Seventy six were Gram-positive bacteria, 29 were Gram-negative and there were ten fungal isolates. Major risk factors for acquisition of nosocomial meningitis were neurosurgery (70.2%), ventriculoperitoneal shunt (42.9%), prior therapy with broad spectrum antibiotics (64.1%), central venous catheter (94.5%), premature neonates with very low birth weight (32.8%) and total parenteral nutrition (68.8%). Overall attributable mortality was 14. 9%; in bacterial infection it was 13.2% and in fungal nosocomial meningitis, 30.0%. Higher mortality was significantly related to perinatal pathology with CNS abnormality, prematurity polymicrobial infection with Enterobacteriaceae and concomitant bacteraemia. Prematurity in neonates, very low birth weight and infection with Enterobacteriaceae were significantly associated with a worse outcome. PMID- 10854812 TI - Microbiologic efficacy of moxifloxacin for the treatment of community-acquired pneumonia due to Chlamydia pneumoniae. AB - Nasopharyngeal specimens for culture of Chlamydia pneumoniae were obtained from patients participating in two pneumonia treatment studies: an open study of 400 mg moxifloxacin orally, qds for 10 days and a randomized, double-blind comparison of moxifloxacin, 400 mg orally, qds versus clarithromycin, 500 mg orally, bd, both for 10 days. C. pneumoniae was eradicated from the nasopharynx of seven of ten (70%) microbiologically evaluable patients who were treated with moxifloxacin and four of four who were treated with clarithromycin. Minimum inhibitory concentrations (MIC) and minimum bactericidal concentrations (MBC) of 21 isolates of C. pneumoniae from 18 patients obtained before and after therapy were performed against moxifloxacin and clarithromycin. The MIC(90)s and MBC(90)s for moxifloxacin and clarithromycin were 1 and 0.06 mg/l, respectively. The MICs and MBCs against moxifloxacin of six isolates from three persistently infected patients who were treated with the drug were the same at baseline and follow-up. The persistence of C. pneumoniae after treatment with moxifloxacin was probably not due to the emergence of resistance. PMID- 10854813 TI - Synergy between 6-amino-2-n-pentylthiobenzothiazole and ergosterol biosynthesis inhibiting antimycotics against Candida albicans in vitro. PMID- 10854814 TI - Extended spectrum beta-lactamases in salmonella strains isolated in Austria. PMID- 10854816 TI - Is the physician like the pilot?: Lessons learned from two professions demanding great responsibility. PMID- 10854815 TI - European organ transplantation: a real challenge(1). PMID- 10854817 TI - Novel approaches to therapy for systemic lupus erythematosus. AB - Current therapies for systemic lupus erythematosus (SLE) are targeted at immunosuppression and at reducing inflammation. The current therapies are broad spectrum and include steroids and cytotoxic agents that are counterbalanced by toxicity and side effects of the medications. Methotrexate can be utilized to reduce steroid requirements in mild to moderate SLE. Manipulation of the hormonal axis includes DHEA and bromocriptine. Mycophenolate mofetil is an immunosuppressive agent that is being investigated for SLE renal disease. Autologous stem cell transplantation or high-dose cyclophosphamide may be an option for severe refractory SLE. The aim of the future is to target therapies by altering specific known mechanisms of inflammation and autoimmunity. Although the inciting antigen is still unknown in SLE, it may be possible to alter the regulation of the immune response by targeted molecular therapy. Methods to do so would include manipulation of idiotypes, manipulation of second signal stimulation of the immune response, manipulation of cytokines, and the induction of tolerance by administration of blocking peptides. IVIg is an immunomodulator that has been successful in the treatment of SLE. Targeted molecular therapy is undergoing phase I trials with monoclonal anti-CD40L, a signaling inhibitor. Anti CTLA4Ig, another signaling blocker, is presently being investigated for psoriasis, but may be a potential therapy for SLE. Finally, therapies may include the administration of peptides to induce tolerance. PMID- 10854818 TI - Thrombotic microangiopathy. AB - The term 'thrombotic microangiopathy' (TMA) describes syndromes of microangiopathic hemolytic anemia, thrombocytopenia, and variable signs of organ impairment, due to platelet aggregation in the microcirculation. The term 'hemolytic uremic syndrome' (HUS) has entered clinical use to describe childhood cases of TMA dominated by renal impairment, while the term 'thrombotic thrombocytopenic purpura' (TTP) refers to adult cases of TMA with predominant neurological abnormalities. HUS and TTP show the same histological lesion characterized by widening of the subendothelial space and microvascular thrombosis and their different manifestations are secondary to the different distribution of the microvascular lesions. Available evidence orients towards endothelial injury as an important factor in the sequence of events leading to the microangiopathic process. Here we provide an overview of the pathophysiology, epidemiology, clinical manifestations, and management of TMA. PMID- 10854819 TI - The antihypertensive effects of doxazosin on the arterial system: changes in peripheral vascular resistance and wall tension. AB - Background: Hypertension is characterized by structural and functional abnormalities that affect the entire cardiovascular system, including the large arteries. The antihypertensive efficacy of doxazosin, a selective alpha(1) antagonist, and its effects on the arterial system were investigated. Method: In our double-blind, randomized, placebo-controlled study including 30 hypertensive patients (doxazosin group: nine males, 11 females; mean age 45+/-12 years; placebo group: four males, six females; mean age 47+/-9 years), the systolic, diastolic and mean blood pressure (BP), heart rate, diameter and area of the brachial artery, peak systolic velocity, end-diastolic velocity, pulsatility index (PI), resistance index (RI), S/D (systolic velocity/diastolic velocity), flow volume, local resistance, and wall tension were recorded before and 4 h after the administration of 2 mg doxazosin or placebo. The two groups were statistically compared. Results: In the doxazosin group, systolic, diastolic and mean pressures decreased significantly (P<0.001), while heart rate remained unchanged. Local resistance (P<0.001), RI (P<0.05), PI (P<0.05), and wall tension (P<0.001) all decreased significantly, while flow volume increased significantly (P<0.05). However, no significant changes were observed in arterial diameter, surface area, peak systolic velocity, end-diastolic velocity or S/D ratio. The placebo group did not show a significant difference in any of the parameters listed above. Conclusion: The antihypertensive effect of doxazosin is accompanied by a reduction in brachial arterial wall tension that occurs without any change in arterial diameter. The lack of change in the diameter of the artery leads us to suggest different effects on other vasomotor determinants. PMID- 10854820 TI - Extrapulmonary tuberculosis in the northeastern suburbs of Paris: 141 cases. AB - Background: Big cities were particularly affected by tuberculosis in the 1990s. Methods: We studied 141 cases of extrapulmonary tuberculosis in patients not infected by HIV in the northeastern suburbs of Paris. Results: A total of 84 men and 57 women were included in the study. Their average age at diagnosis was 42.2 years. Some 73.6% of the patients were foreign-born. A total of 182 sites were identified in 141 patients. There was an association with pulmonary tuberculosis in 38 cases. The sites were: lymph node (48.9%), pleural (25.5%), skeletal (22.7%), genitourinary (5.7%), and meninges (5%). Unfavorable social conditions were frequently observed. The average duration of treatment was 10 months. Twenty four adverse drug effects were noted. Sixty-eight strains of Mycobacterium tuberculosis were isolated. Five cases of primary resistance to at least one antituberculous drug and only one case of multidrug resistance were observed. Some 95.7% of the 93 patients who were not lost to follow up were cured. Conclusion: Independently of HIV infection, extrapulmonary tuberculosis is still present, particularly in the suburbs of big cities, where social conditions are poor. The significant number of patients lost to follow-up demands that measures be adapted for the therapeutic management of these patients. PMID- 10854821 TI - Effects of interferon treatment on the glucose metabolism of patients with chronic hepatitis C. AB - Background: The authors report on changes in carbohydrate metabolism observed in 32 patients undergoing therapy with interferon-alpha for chronic hepatitis C. Methods: Diabetes had been diagnosed in three patients and impaired glucose tolerance ascertained in one patient before interferon therapy was started. The remaining 28 patients were non-diabetic. Interferon-alpha was administered in 3 MU doses three times per week. Results: Glucose tolerance deteriorated in two of the three diabetics, and eventually these patients had to be switched from oral hypoglycemic agents to insulin. The patient with impaired glucose tolerance at baseline progressed gradually to overt diabetes. Nine of the 28 previously non diabetic patients developed impaired glucose tolerance during interferon therapy. Conclusion: The deleterious effects of interferon-alpha on carbohydrate metabolism proved to be reversible. Regular monitoring of the glucose level of patients during and after interferon therapy is mandatory. PMID- 10854822 TI - Increased pulmonary epithelial permeability in systemic sclerosis is associated with enhanced cutaneous nerve growth factor expression. AB - Background: Nerve growth factor (NGF), a neurotrophic factor that indirectly induces fibroblast proliferation and collagen production, has been found to be increased in the affected dermis of patients with systemic sclerosis (SSc). To investigate the possibility of a relationship between cutaneous NGF production and pulmonary damage in SSc, we studied seven non-smoking scleroderma patients. Methods: Abnormalities in lung structure were assessed by radiological lung examination, and pulmonary epithelial permeability (PEP) was determined by ventilation lung scintigraphy. All patients underwent skin punch biopsy with NGF immunohistological staining. Results: A statistically significant correlation was found between the PEP values and the cutaneous NGF staining scores, which were markedly increased in all of the patients examined, irrespective of the age, disease duration, or radiologically defined lung abnormalities. Conclusion: These results support the hypothesis that functional and anatomical changes in SSc target organs may be determined by a local tissue hyperproduction of NGF. PMID- 10854823 TI - Adult beta-thalassemic patients with chronic hepatitis C: long-term efficacy of alpha-IFN treatment. AB - Background: The purpose of this study was to assess the effectiveness of alpha IFN in adult beta-thalassemic patients with chronic hepatitis C. After a long term follow-up, we describe the special pattern of biochemical and virological response of thalassemics. Methods: Thirty-two anti-HCV-positive adult thalassemic patients (19 female and 13 male, mean age 23.4+/-5.5 years) with biopsy-proven chronic hepatitis were treated with IFN alpha2beta at a dose of 3 MU thrice weekly for 6-12 months. The patients were followed up until 45-62 months after the end of treatment. Results: A sustained response was obtained in eight patients (25%). Only two of the sustained responders (25%) normalized ALT during the first 3 months of treatment. Both early and late biochemical responders cleared HCV-RNA after 6 months of treatment. Eight patients (25%) responded with ALT normalization within 2 months of treatment but relapsed soon after stopping IFN. Sixteen patients (50%) did not respond to IFN. Conclusion: The response rate in multitransfused thalassemic patients with chronic hepatitis C treated with IFN is similar to that in non-thalassemics. The special feature of thalassemics is that early biochemical response does not predict a sustained response; on the contrary, patients who normalize ALT after 6 months of IFN treatment usually do not relapse. PMID- 10854824 TI - Multifactorial thrombotic-type microangiopathy with skin ulcers and hepatitis C infection. AB - The case of a patient with hepatitis C virus infection who presented with symmetric ulcers of the legs is reported. She was found to have type III mixed cryoglobulinemia, high titers of anticardiolipin antibodies, lupus anticoagulant, and a free protein S deficiency. To our knowledge, this is the first reported case of such an association. The role of these factors in the pathogenesis of skin lesions in the setting of hepatitis C virus infection is discussed. PMID- 10854825 TI - Cyclic Cushing's disease associated with primary empty sella. AB - A small number of cases of Cushing's disease (CD) associated with primary empty sella (ES) have been described in the literature. Pituitary microsurgery is the recommended treatment. An alternative is chronic treatment with ketoconazole, an inhibitor of adrenal cortisol synthesis. Cyclic Cushing's syndrome is characterized by episodic cortisol hypersecretion. CD is the most frequent etiology of cyclic Cushing's syndrome. To our knowledge, cyclic CD has not previously been reported in association with primary ES. We describe a patient with cyclic CD associated with primary ES who was initially treated with ketoconazole and subsequently cured by transsphenoidal surgery. PMID- 10854826 TI - Benign symmetric lipomatosis (Madelung's disease). AB - We report on a 50-year-old man who presented with benign, symmetric lipomatosis and chronic alcoholic liver disease. The characteristic clinical features and a brief review of the current literature are presented. PMID- 10854827 TI - The concept of general internal medicine of the Slovak Society of Internal Medicine. AB - The data demonstrated a trend toward increasing subspecialisation and away from generalism. The European Federation of Internal Medicine and American Society of Internal Medicine warn that such development could be disadvantageous for patients and society. Changes in age distribution of the population, polymorbid patients, limited financial sources, drug interactions, multisystemic disorders, urgent medicine and indication of surgical intervention are main reasons for the need to preserve General Internal Medicine. The ideas presented in this paper are trying to solve the problem. PMID- 10854828 TI - Poly(ADP-ribose) polymerase-independent potentiation of nitrosourea cytotoxicity by 3-aminobenzamide in human malignant glioma cells. AB - Poly(ADP-ribose) polymerase is a zinc-finger DNA-binding protein that detects specifically DNA strand breaks generated by genotoxic agents and is thought to be involved in DNA repair. Here, we examined the effects of 3-aminobenzamide, a poly(ADP-ribose) polymerase inhibitor, on the chemosensitivity of human malignant glioma cells. 3-Aminobenzamide selectively potentiated the cytotoxicity of the nitrosoureas, nimustine, carmustine and lomustine in 10 of 12 human malignant glioma cell lines. In contrast, 3-aminobenzamide did not modulate the cytotoxic effects of doxorubicine, teniposide, vincristine, camptothecin or cytarabine. The nitrosoureas did not induce poly(ADP-ribose) polymerase activity in the glioma cells. Ectopic expression of truncated poly(ADP-ribose) polymerase containing the poly(ADP-ribose) polymerase DNA-binding domain, which acts as a dominant-negative mutant, in LN-18 or LN-229 cells did not alter the 3-aminobenzamide effect on nitrosourea-mediated cytotoxicity. Thus, 3-aminobenzamide may target another nicotinamide adenine dinucleotide (NAD)-requiring enzyme, but not poly(ADP ribose) polymerase, when enhancing nitrosourea cytotoxicity in human malignant glioma cells. Carmustine cytotoxicity was associated with a G2/M arrest. Coexposure to carmustine and 3-aminobenzamide overcame this G2/M arrest in T98G cells, which are sensitized to carmustine by 3-aminobenzamide, but not in U251MG cells, which are refractory to 3-aminobenzamide-mediated sensitization to carmustine. Thus, 3-aminobenzamide-mediated sensitization to carmustine cytotoxicity may result from interference with the stable G2/M arrest response to carmustine in human glioma cells. PMID- 10854829 TI - Inverse agonism at alpha(2)-adrenoceptors in native tissue. AB - Several alpha(2)-adrenoceptor antagonists have inverse agonist properties in cell culture systems, usually expressing high levels or a constitutively active form of alpha(2)-adrenoceptors. In characterizing the binding of alpha(2)-adrenoceptor agonists to rat brain tissue sections, we found that conditions known to alter agonist affinity for these receptors, particularly the addition of 100 microM GTP, altered the binding of the alpha(2)-adrenoceptor antagonist, [3H](1,4 benzodioxan-2-methoxy-2-yl)-2-imidazoline hydrochloride (RX821002). In further studies, we found that under our conditions [3H]RX821002 demonstrates inverse agonist properties at alpha(2)-adrenoceptors. This is the first demonstration of inverse agonism at alpha(2)-adrenoceptors in native tissue. We found that the alpha(2)-adrenoceptor antagonist, (2S,12bS)1', 3' dimethylspiro(1,3,4,5',6,6',7,12b-octahydro-2H-benzo(b)fu ro(2, 3-a)quinazoline) 2,4'-pyrimidin-2'-one (MK-912), did not have clearly discernible inverse agonist properties and acted as a neutral antagonist in these studies. On the other hand, the antagonist rauwolscine actually displayed partial agonist properties in our studies. These findings indicate that the inverse agonist properties of alpha(2) adrenoceptor antagonists can be demonstrated in native tissue, as well as in tissue culture, and they strengthen the idea that inverse agonist properties may be of physiological and pharmacological importance. PMID- 10854830 TI - Inhibitory effect of anti-diabetic agents on rat organic anion transporter rOAT1. AB - The interactions of sulfonylureas and a novel anti-diabetic drug, nateglinide, with rat renal organic anion transporter (rOAT1) expressed in Xenopus laevis oocytes were studied. Uptake of p-aminohippurate via rOAT1 was markedly inhibited by glibenclamide and nateglinide, and moderately by chlorpropamide and tolbutamide. The inhibition constant values (K(i)) for chlorpropamide, glibenclamide, tolbutamide and nateglinide were 39.5, 1.6, 55.5 and 9.2 microM, respectively. Kinetic analysis showed that the inhibition of p-aminohippurate uptake by glibenclamide was competitive. Sulfonylureas examined and nateglinide did not show a trans-stimulation effect on [14C]p-aminohippurate efflux from rOAT1-expressing oocytes. There was no stimulation of [3H]glibenclamide uptake via rOAT1. These findings suggested that sulfonylureas and nateglinide interact with rOAT1, but these drugs are not translocated via the transporter. PMID- 10854832 TI - T-477, a novel Ca(2+)- and Na(+) channel blocker, prevents veratridine-induced neuronal injury. AB - To evaluate the effect of (R)-(+)-2-(4-chlorophenyl)-2, 3-dihydro-4-diethyl aminoacetyl-4H-1,4-benzothiazine hydrochloride (T-477), a novel Na(+)- and Ca(2+) channel blocker, on neuronal injury in vitro, we studied veratridine-induced injury in cultured rat hippocampal neurons. Neurons swelled extensively 10 min after the addition of veratridine, and returned to their initial size within 2 h. Intracellular Na(+) and Ca(2+) concentrations and amino acid release from the cells, in particular, that of glutamate, increased after the treatment with veratridine. Approximately 70% of neurons died within 24 h. T-477 inhibited both veratridine-induced swelling and death in a concentration-dependent manner. Moreover, T-477 concentration dependently reduced the increases in Na(+) and Ca(2+) influx and amino acid release. These results suggest that T-477 prevented the veratridine-induced influx of Na(+) and, thereby, reduced neuronal swelling. This, combined with the effects of T-477 on the inhibition of Ca(2+) influx and glutamate release, possibly by the blockade of Na(+) channels, may be the mechanism by which T-477 protects neurons from death induced by veratridine. PMID- 10854831 TI - Ca(2+) influx stimulated phospholipase C activity in bovine adrenal chromaffin cells: responses to K(+) depolarization and histamine. AB - The role of Ca(2+) influx in activating phospholipase C in bovine adrenal chromaffin cells has been investigated. Phospholipase C activity in response to K(+) depolarization (56 mM) was blocked by the L-type Ca(2+) channel antagonist nifedipine and partially inhibited by the omega-conotoxins GVIA and MVIIC. In contrast, phospholipase C activity in response to histamine receptor activation was unaffected by omega-conotoxin GVIA and partially inhibited by omega-conotoxin MVIIC or nifedipine. This response was however markedly inhibited by the non selective Ca(2+) channel antagonists La(3+) or 1-[beta-[3-(4 Methoxyphenyl)propoxy]-4-methoyphenethyl]-H-imidazol e (SKF-96365). Despite this Ca(2+) dependence phospholipase C activity was not increased during periods of "capacitative" Ca(2+) inflow generated by histamine-, caffeine- or thapsigargin mediated depletion of internal Ca(2+) stores. Thus, while Ca(2+) influx in response to K(+) depolarization or G-protein receptor activation can increase phospholipase C activity in these cells, in the latter case it appears to be ineffective unless there is concurrent agonist occupation of the receptor. PMID- 10854833 TI - Modulation of adenosine concentration by opioid receptor agonists in rat striatum. AB - There is evidence that adenosine and morphine interact in the striatum. However, little is known about the precise role of the opioid receptor subtypes implicated in the modulation of adenosine tissue concentration and in adenosine receptor expression and function. We sought to evaluate, in the absence of withdrawal symptoms, the effects of the short-term administration of selective mu-, delta- or kappa-opioid receptor agonists on adenosine concentration and on adenosine A(2A) receptor function in rat striatum. Adenosine A(2A) receptor was chosen because the neuronal sub-population expressing this receptor coexpresses enkephalin, suggesting that adenosine A(2A) receptor may be regulated by opioid receptor agonists. Oxymorphone hydrochloride mu-opioid receptor agonist, 6 mg/kg/day), +[-(5 alpha,7 alpha, 8 beta)-(-)-N-methyl-N(7-(1-pyrrolidinyl)1 oxaspiro (4.5)dec-8-yl) benzenacetamide] (U69593) (kappa-opioid receptor agonist, 0.75 mg/kg/day), and (+)-4[(alpha R)-alpha-((2S,5R)-4-allyl-2, 5-dimethyl-1 piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide) (SNC80) (delta-opioid receptor agonist, 9 mm/kg/day), or vehicle, were administered i.p 3 x daily during 5 days to groups of rats (n=6). We also investigated the effects of opioid receptor agonists on adenosine uptake by striatal cell extracts. We found that administration of mu- or delta-opioid receptor agonists significantly decreased adenosine uptake in striatal cell extracts and increased adenosine concentration (mean+24% and +45% for mu- and delta-opioid receptor agonist, respectively, relative to controls). None of the receptor agonists tested induced obvious modifications of adenosine A(2A) receptor function. However, the delta-opioid receptor agonist induced an increase in adenosine A(2A) mRNA expression (mean 44%). We conclude that mu and delta receptor agonists inhibit adenosine uptake by striatal cell extracts and increase adenosine concentrations in rat striatum. PMID- 10854834 TI - Intravenous and oral clozapine pharmacokinetics, pharmacodynamics, and concentration-effect relations: acute tolerance. AB - We examined the pharmacokinetics and pharmacodynamics of intravenous (1-5 mg/kg) and oral clozapine (2.5-10 mg/kg) in rats (terminal half-life=81.8 min; oral bioavailability=5.32%). Both dose- and concentration-effect relations of clozapine were characterized. Clozapine's effects were similar to those of benzodiazepines because of the similarity in effect-time profiles between the two classes of drugs. The IC(50) value increased as a function of dose; consequently, clozapine's relative potency decreased linearly with the logarithm of AUC((0 infinity)), or bioavailable dose regardless of route of administration. The IC(50) is an index for the sensitivity of behavioral performance to clozapine; relative potency provides an index for estimating the extent of acute tolerance. As IC(50) increases, relative potency decreases, and consequently, acute tolerance increases. Our results demonstrated that greater acute tolerance was observed for i.v. clozapine than for p.o. clozapine; however, clozapine exhibited a single concentration-effect relation across dose and route of administration after correcting for relative potencies. PMID- 10854835 TI - The effect of the acute administration of various selective 5-HT receptor antagonists on focal hippocampal seizures in freely-moving rats. AB - In this study, we assessed the effects of the acute administration of various 5 HT receptor antagonists on hippocampal partial seizures generated by low frequency electrical stimulation in male Wistar rats. The seizure threshold and severity were determined by measuring the pulse number threshold and primary and secondary afterdischarges, respectively, and the latency of secondary discharge was also determined. The administration of either the selective 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazineyl]ethyl]-N-(pyridinyl)-c yclohe xanecarboximimde 3 HCl (WAY 100635, 0.1-1 mg/kg i.p.), the selective 5-HT(3) receptor antagonist granisetron (0.3-3 mg/kg i.p.), the selective 5-HT(2A) receptor antagonist R-(+)-a-(2, 3-dimethoxyphenyl)-1-[2-(4-fluorophenyl) ethyl]-4 piperidine-methanol (MDL 100907, 0.3-3 mg/kg i.p.) or the 5-HT(2B,C) receptor antagonist antagonist N-(1-methyl-5-indolyl)-N'-(3-pyridyl) urea HCl (SKB 200646A, 5-50 mg/kg i.p.) did not alter the pulse number threshold compared to vehicle-treated animals. However, the acute administration of WAY 100635 (0.3 mg/kg) and M100907 (1 mg/kg) significantly increased, whereas granisetron (1 mg/kg) decreased, the primary afterdischarge duration compared to vehicle-treated animals. The latency of secondary after discharge was significantly decreased by WAY 100635 (1 mg/kg) and granisetron (3 mg/kg) compared to vehicle-treated animals. These results suggest that in this model, the antagonism of 5-HT(1A), 5 HT(2A), 5-HT(3) or 5-HT(2B,C) receptors do not lower or raise seizure threshold. However, the antagonism of 5-HT(1A) receptors may increase or augment seizure severity. PMID- 10854836 TI - Effects of ensaculin on dopamine metabolite levels and K(+)-induced glutamate release. AB - In vivo microdialysis with the new antidementia compound ensaculin was performed in freely moving rats to study the alterations in dopaminergic and glutamatergic neurotransmission. Ensaculin (0.1 and 1 mg/kg i.p.) significantly increased extracellular levels of the dopamine metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). Furthermore, ensaculin (1 mg/kg i.p.) showed a non-significant tendency to reduce the K(+)-induced glutamate release. The data suggest that ensaculin may have moderate D(2) antagonistic properties. Thus, besides its possible role as a cognitive enhancer, ensaculin may also have moderate antipsychotic properties. PMID- 10854837 TI - Effects of Delta 9-tetrahydrocannabinol, (R)-methanandamide, SR 141716,and d amphetamine before and during daily Delta 9-tetrahydrocannabinol dosing. AB - We examined the effects of Delta 9-tetrahydrocannabinol (Delta 9-THC), (R)-(+) arachidonyl-1'-hydroxy-2'-propylamide ((R)-methanandamide, AM 356), SR 141716, and d-amphetamine on fixed-ratio (FR) responding maintained by food in rats before and during daily dosing with Delta 9-THC. Rats responded under a FR 10 schedule of food reinforcement. Cumulative dose-response curves for the various drugs were determined before and during daily Delta 9-THC administration. All four drugs dose-dependently decreased responding both before and during daily dosing with Delta 9-THC (18 mg/kg/day). The dose-response curves for both Delta 9 THC and (R)-methanandamide were shifted to the right with daily dosing with Delta 9-THC, indicating tolerance to the effects of Delta 9-THC and cross-tolerance to the effects of (R)-methanandamide. The doses of d-amphetamine examined produced similar effects both before and during daily dosing with Delta 9-THC. The effects of SR 141716 were not consistently altered by daily Delta 9-THC administration. These results indicate that tolerance develops to the effects of Delta 9-THC, when Delta 9-THC is administered repeatedly. These results also indicate that cross-tolerance to (R)-methanandamide develops with repeated Delta 9-THC administration. PMID- 10854838 TI - Differential effects of ibogaine on behavioural and dopamine sensitization to cocaine. AB - To investigate a possible basis for the proposed anti-addictive property of ibogaine, the effects of ibogaine (40 mg/kg, i.p., 19 h earlier) on the expression of sensitization induced by cocaine were investigated. Ibogaine pretreatment potentiated the increase in the stereotypic effects of a cocaine challenge (20 mg/kg) in both sensitized (5 x 15 mg/kg, i.p.) and acutely treated rats. However, while ibogaine pretreatment did not significantly alter the dopamine response in the nucleus accumbens to acute cocaine, it abolished the expression of cocaine-induced dopamine sensitization. This result demonstrates that ibogaine pretreatment can reverse one of the neuroadaptations produced by chronic cocaine administration, an effect that may contribute to its putative anti-addictive property. PMID- 10854839 TI - Combined cardiac effects of cocaine and the anabolic steroid, nandrolone, in the rat. AB - Despite reports of an increase in the incidence of simultaneous cocaine and anabolic steroid abuse, potential adverse interactions between these two drugs on the cardiovascular system are largely unquantified. Cocaine has been reported to induce coronary vasoconstriction, cardiac arrhythmias and conduction delays. Anabolic steroids have been associated with cardiac hypertrophy and hypertension. Utilising both in vivo (radiotelemetry) and in vitro (isolated Langendorff perfused heart) techniques, our aim was to determine whether anabolic steroids cause cardiac hypertrophy and alter cardiac function, and consequently alter the response of the heart to cocaine. It was found that 15 days of treatment of rats with nandrolone decanoate (20 mg/kg, s.c.) was not sufficient to cause hypertrophy, alter cardiac function or the spread of electrical activity through the heart. However, nandrolone pretreatment was found to significantly potentiate the heart rate response to cocaine (45 mg/kg, i.p.) in vivo. This study indicates that nandrolone significantly elevates the heart rate response to high dose cocaine without changing heart morphology. The mechanism of this interaction remains uncertain. PMID- 10854840 TI - Effects of the I(K.ATP) blockers glibenclamide and HMR1883 on cardiac electrophysiology during ischemia and reperfusion. AB - Clinical evidence indicates an antiarrhythmic effect of sulfonylureas, which might be blunted by their vascular action. We wanted to investigate the effect of glibenclamide and the new sulfonylthiourea compound 1-[[5-[2-(5-chloro-o anisamido)ethyl]-2-methoxyphenyl]-sulfonyl]-3 -me thylthiourea (HMR1883) on cardiac electrophysiology in the course of regional ischemia and reperfusion. Isolated rabbit hearts (Langendorff-technique) were pretreated with either vehicle (n=14), 3 micromol/l glibenclamide (n=7) or 3 micromol/l HMR1883 (n=7) before regional ischemia was induced by left coronary artery branch occlusion (45 min) followed by 45 min reperfusion. Unipolar epicardial electrocardiograms were recorded from 256 epicardial AgCl electrodes. Coronary ligation resulted in a decrease in coronary flow (CF) by 35% and in left ventricular pressure (LVP) by 40% in all series. The occluded zone was 23+/-3% in all series. Ischemia led to shortening of the epicardial activation-recovery interval (ARI) in the ischemic area, which was inhibited by both drugs especially in the early phase. In the non ischemic area, ARIs remained stable and there was no effect of the drugs. Ischemia led to an increase in the regional difference in ARI between ischemic center and border zone. This increase was significantly inhibited by both substances during late ischemia and early reperfusion (until 15 min reperfusion). In addition, the dispersion of ARIs was reduced by both drugs during late ischemia and reperfusion. Ventricular fibrillation was observed in 7/14 (control), 0/7 (glibenclamide), and 0/7 (HMR1883). All ventricular fibrillation occurred during reperfusion. In glibenclamide but not in HMR1883-treated hearts recovery of CF upon reperfusion was significantly depressed (control: 25.5+/-4; HMR1883: 23+/-2.5; glibenclamide: 16+/-1 ml/min, values at 2 min reperfusion), while the elevation of ST-segments of the electrograms in early ischemia was fully prevented by both treatments. We conclude that both glibenclamide and HMR1883 exert an antiarrhythmic effect in this model, and reduce the shortening of the ARIs in the ischemic area, thus attenuating regional differences in ARIs between ischemic and non-ischemic area. Furthermore, unlike glibenclamide HMR1883 does not interfere with postischemic hyperemia. PMID- 10854841 TI - Role of endothelium/nitric oxide in atypical beta-adrenoceptor-mediated relaxation in rat isolated aorta. AB - The role of endothelium in the modulation of classical and atypical beta adrenoceptor-mediated vasorelaxation was investigated in ring preparations of rat isolated thoracic aorta. Rings were pre-constricted with a sub-maximal concentration of noradrenaline (1 microM) and relaxant responses to cumulative concentrations of beta-adrenoceptor agonists obtained. Endothelium removal or pretreatment with N(G)-nitro-L-arginine methyl ester (L-NAME, 100 microM) or 1H [1,2,4] oxadiazolol[4,3,-a] quinoxalin-1-one (ODQ, 10 microM) significantly reduced the relaxant effects of isoprenaline, but had less effect on relaxant responses to the atypical beta-adrenoceptor agonist, (+/-)-4-(3-t-butylamino-2 hydroxypropoxy)-benzimidazol-2-one hydrochloride (CGP 12177A). Sodium nitroprusside (3 nM) shifted the isoprenaline concentration-response curve to the left and restored the attenuated responses in the presence of L-NAME back to control levels. Sodium nitroprusside had little effect on the CGP 12177A concentration-response curve. The results show that the endothelium/nitric oxide (NO) pathway modulates beta-adrenoceptor-mediated vasorelaxation in rat aorta and that classical beta-adrenoceptors are modulated to a greater extent than atypical beta-adrenoceptors. PMID- 10854842 TI - Effects of carnitine palmitoyltransferase I inhibitors on hepatic hypertrophy. AB - We investigated the effect of two types of carnitine palmitoyltransferase I inhibitors, ethyl 2-(6-(4-chlorophenoxy)hexyl)oxirane-2-carboxylate (etomoxir) and (R)-3-carboxy-N,N, N-trimethyl-2-?[hydroxy(tetradecyloxy)phosphinyl]oxy?-1 propana minium hydroxide (SDZ CPI 975), on cardiac and hepatic hypertrophy in ddY mice. One-week administration of etomoxir caused cardiac and hepatic hypertrophy, 19% and 22% as a ratio to body weight, respectively. Although 4-week administration of etomoxir caused hepatic hypertrophy, there was no significant change in liver triglyceride content in the first or second week. In cultured HepG(2) cells, etomoxir treatment (1 week) did not cause triglyceride to accumulate. One-week administration of SDZ CPI 975 caused neither cardiac nor hepatic hypertrophy. In vitro, neither drug had selectivity for carnitine palmitoyltransferase I isozymes. These findings suggest that the hepatic hypertrophy following 1- or 2-week treatment with etomoxir is caused by mechanisms different from those responsible for triglyceride accumulation, and that inhibition of carnitine palmitoyltransferase I may not necessarily induce hepatic hypertrophy. PMID- 10854843 TI - Hyperosmolarity-induced relaxation and prostaglandin release in guinea pig trachea in vitro. AB - In this study, a tracheal perfusion apparatus was used to investigate the nature of the relaxing factor released by hyperosmolarity on the epithelial side of guinea pig trachea. NaCl induced concentration-dependent relaxation. This relaxation was not affected when the trachea was preincubated with a vasoactive intestinal peptide (VIP) receptor antagonist or with the nitric oxide synthesis inhibitor N(G)-monomethyl-L-arginine (L-NMMA). When the prostaglandin synthesis was prevented by preincubation with the phospholipase A(2)-inhibitor quinacrine, or the cyclooxygenase inhibitor indomethacin, the maximal relaxation induced by NaCl was suppressed by 50% (P<0.05). Moreover, the prostaglandin E(2) concentration was four times higher (P<0.05) in the organ bath during the relaxations, whereas the nitric oxide concentration remained unchanged. In conclusion, increased osmolarity on the airway surface leads to the release of prostaglandins, which are involved in part in the hyperosmolarity-induced relaxation of airway smooth muscle. This might be relevant for asthmatic patients since prostaglandin may modulate the bronchoconstrictive response to hyperosmolar stimuli and exercise. PMID- 10854844 TI - Substance P induces tumor necrosis factor-alpha release from human skin via mitogen-activated protein kinase. AB - Substance P plays an important role in neurogenic inflammation with granulocyte infiltration. To investigate cytokines involved in the substance P-induced inflammation and the mechanism of cell activation, we studied the release of TNF (tumor necrosis factor)-alpha and histamine from human skin slices in response to substance P and antigen. Substance P induced the release of histamine and TNF alpha in a dose-dependent manner at concentrations from 0.8 to 100 microM. PD 098059 (2'-amino-3'-methoxyflavone) selectively inhibited the release of TNF alpha, but not the release of histamine induced by either substance P or antigen. SB 203580 ([4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-++ +imida zole]) slightly inhibited TNF-alpha release induced by antigen, but not that induced by substance P, and slightly enhanced histamine release induced by either stimulation. The release of TNF-alpha in response to either stimulation was inhibited by 1 nM-1 microM dexamethasone, but histamine release was not affected. These results suggest that substance P, in addition to antigen, induced TNF-alpha release from human skin by a mitogen-activated protein (MAP) kinase, predominantly extracellular signaling-regulated protein kinase (ERK)-dependent, and dexamethasone-sensitive pathway, which is separate from that for histamine release from mast cells. PMID- 10854845 TI - Phenytoin inhibits both the first ovulation and uterine development in gonadotropin-primed immature rats. AB - This study was planned to determine the effects and possible mechanism of action of phenytoin on development of the reproductive tract and first ovulation in immature rats. Rats were injected s.c. with 5 IU of equine chorionic gonadotrophin (equine CG) on day 26 to induce ovarian and uterine development. Treatment with phenytoin (140 mg/kg) at 1200 h on day 28, which induces serum levels approximately twice those reached with the clinical dose as anticorvulsant drug for humans, was effective for inhibiting the first ovulation and normal secretion of serum follicle - stimulating hormone and luteinizing hormone (LH) on day 29 as well as the preovulatory gonadotrophin surge on day 28. The block of ovulation was overcome by administration of human chorionic gonadotrophin or LH releasing hormone on day 28. Simultaneous treatment with equine CG and phenytoin at 0800 h on day 26 did not affect either ovarian weight or ovarian hormones secretion, whereas phenytoin clearly inhibited the normal increase in uterine weight on day 27. Furthermore, phenytoin suppressed uterine growth after 17beta oestradiol injection. These results indicate that phenytoin inhibits the first ovulation by inhibiting the gonadotrophin surge and further, that the drug impairs the stimulatory effects of oestrogen on uterine proliferation in the gonadotropin-induced ovulation model. PMID- 10854846 TI - Application of linear free energy relationships to the serpin-proteinase inhibition mechanism. AB - Linear free energy relationships can be used to link the changes in rate constant for a reaction to changes in the equilibrium caused by alterations in structure. While they have most often been used in the analysis of chemical reactions, they have also been employed to resolve questions in enzymology and protein folding. Here we analyze the reaction of a serpin with a panel of six serine proteinases, and observe that a linear free energy relationship exists between the true second order rate constant for reaction, k(inh), and the inhibition constant, K(I), indicating that formation of the covalent serpin-enzyme complex may be reversible. PMID- 10854847 TI - Retrovirus-mediated gene delivery into male germ line stem cells. AB - The male germ line stem cell is the only cell type in the adult that can contribute genes to the next generation and is characterized by postnatal proliferation. It has not been determined whether this cell population can be used to deliberately introduce genetic modification into the germ line to generate transgenic animals or whether human somatic cell gene therapy has the potential to accidentally introduce permanent genetic changes into a patient's germ line. Here we report that several techniques can be used to achieve both in vitro and in vivo gene transfer into mouse male germ line stem cells using a retroviral vector. Expression of a retrovirally delivered reporter lacZ transgene in male germ line stem cells and differentiated germ cells persisted in the testis for more than 6 months. At least one in 300 stem cells could be infected. The experiments demonstrate a system to introduce genes directly into the male germ line and also provide a method to address the potential of human somatic cell gene therapy DNA constructs to enter a patient's germ line. PMID- 10854848 TI - Structural models for carcinoembryonic antigen and its complex with the single chain Fv antibody molecule MFE23. AB - MFE23 is a single chain Fv antibody that has a high affinity for carcinoembryonic antigen (CEA). A full homology model for CEA based on V-type, I-type and C2-type immunoglobulin folds, 28 oligosaccharides and the interdomain angle of CD2 was validated using solution scattering data. The superimposition of the intermolecular contacts observed in our recent crystal structure of MFE23 with the N-terminal domain of CEA permitted the MFE23-CEA complex to be modelled. Good surface and electrostatic complementarity and carbohydrate-unhindered access of MFE23 with the indentation between the first two CEA domains was observed. The model is supported by biochemical data and provides insight on the high affinity of MFE23 for CEA. PMID- 10854849 TI - Differential degradation of calpastatin by mu- and m-calpain in Ca(2+)-enriched human neuroblastoma LAN-5 cells. AB - In neuroblastoma LAN-5 cells during calpain activation, in addition to the two expressed 70 kDa and 30 kDa calpastatin forms, other inhibitory species are produced, having molecular masses of 50 kDa and 15 kDa. At longer times of incubation, both native and new calpastatin species disappear. The formation of these new calpastatins as well as the decrease in intracellular total calpastatin activity are mediated by calpain itself, as indicated by the effect of the synthetic calpain inhibitor I, which prevents both degradative processes. Analysis of the calcium concentrations required for the two processes indicates that the first conservative proteolytic event is mediated by micro-calpain, whereas the second one is preferentially carried out by m-calpain. The appearance of the 15 kDa form, containing only the calpastatin repetitive inhibitory domain and identified also in red cells of hypertensive rats as the major inhibitor form, can be considered a marker of intracellular calpain activation, and it can be used for the monitoring of the involvement of calpain in pathological situations. PMID- 10854850 TI - Creatine kinase isoenzymes specificities: histidine 65 in human CK-BB, a role in protein stability, not in catalysis. AB - Creatine kinases (CK) play a prominent role in cell energy distribution through an energy shuttle between mitochondria and other organelles. Human brain CK was cloned and overexpressed in COS-7 cells. We then deleted His-65 and/or Pro-66 situated near the center of a flexible loop as shown by X-ray crystallography on mitochondrial and cytosolic CK. The DeltaH65 mutant had nearly the same affinity for its substrates as wild isoenzyme, but its stability was very low. Unlike DeltaH65, DeltaH65P66 had a eightfold decreased affinity for creatine phosphate and was unable to dephosphorylate cyclocreatine phosphate. Our results demonstrate that, despite an overall similar shape of the proteins, this loop accounts for some subtle differences in isoenzyme functions. PMID- 10854851 TI - TSH regulates a gene expression encoding ERp29, an endoplasmic reticulum stress protein, in the thyrocytes of FRTL-5 cells. AB - This experiment was performed to evaluate the effect of thyroid-stimulating hormone (TSH) on the endoplasmic reticulum resident 29 kDa protein (ERp29) gene expression in the thyrocytes of FRTL-5 cells. Although ERp29 mRNA was constantly expressed, its expression began to increase remarkably from 10(-9) M TSH. On the other hand, the effect of TSH on the abundance of ERp29 mRNA started within 6 h and peaked at 8 h. Actinomycin D strongly blocked this effect while cycloheximide did not. The half-life of ERp29 mRNA was about 4-4.5 h in the presence or absence of TSH that was not affected by the stability of ERp29 mRNA. The effect of TSH on the ERp29 gene expression was specific, while other growth factors (transferrin, insulin and hydrocortisone) did not alter its expression. Our data indicate for the first time that the expression of ERp29 is regulated transcriptionally by TSH in thyrocytes. PMID- 10854852 TI - The APS adapter protein couples the insulin receptor to the phosphorylation of c Cbl and facilitates ligand-stimulated ubiquitination of the insulin receptor. AB - The APS adapter protein is rapidly tyrosine-phosphorylated following insulin stimulation. In insulin-stimulated 3T3-L1 adipocytes, APS co-precipitated with phosphorylated c-Cbl. In CHO.T-APS cells overexpressing the insulin receptor and APS, APS co-precipitated with c-Cbl but not in CHO.T cells which do not express APS. APS-mediated recruitment of c-Cbl to the insulin receptor led to rapid ubiquitination of the insulin receptor beta-subunit in CHO. T-APS but not in parental CHO.T cells. These results suggest that the function of APS is to facilitate coupling of the insulin receptor to c-Cbl in order to catalyse the ubiquitination of the receptor and initiation of internalisation or degradation. PMID- 10854853 TI - Catalase activity of oxygenase domain of rat neuronal nitric oxide synthase. Evidence for product formation from L-arginine. AB - Nitric oxide synthases (NOSs) catalyze the formation of nitric oxide from L arginine. We purified the heme containing, tetrahydrobiopterin-free, oxygenase domain of rat neuronal nitric oxide synthase (nNOSox) overexpressed in Escherichia coli. We found catalase activity in nNOSox. This is significant because H(2)O(2) may also be a product of nitric oxide synthases. We found H(2)O(2) assisted product formation from N-hydroxy-L-arginine and even from L arginine both in the presence and in absence of tetrahydrobiopterin. We propose how heme moiety of the oxygenase domain alone is sufficient to carry out both steps of the NOS catalysis. PMID- 10854854 TI - Rapid up-regulation of P2Y messengers during granulocytic differentiation of HL 60 cells. AB - HL-60 cells are human promyelocytic cells expressing two ATP receptors: the P2Y(2) and P2Y(11) subtypes. Our Northern blotting experiments have shown that P2Y(2) and P2Y(11) messengers were up-regulated in these cells, rapidly and independently of protein synthesis, following treatment with granulocytic differentiating agents such as retinoic acid, dimethylsulfoxide, granulocyte colony stimulating factor, dibutyryl cyclic AMP and ATP. AR-C67085 and adenosine 5'-O-(3-thiotriphosphate), two potent agonists of the recombinant P2Y(11) receptor, increased intracellular cAMP concentration in HL-60 cells more potently than ATP itself. These observations support the conclusion that the effect of ATP on HL-60 cell differentiation is mediated by the P2Y(11) receptor. PMID- 10854855 TI - Detection of the absorption of glucose molecules by living cells using atomic force microscopy. AB - A very small electrode (nanobiosensor) was constructed by immobilizing enzyme (glucose oxidase or hexokinase) on the surface of the cantilever of the atomic force microscope in order to detect the absorption of glucose molecules by living cells. If glucose is present, the nanobiosensor deflects, probably due to the reaction heat evolved in the process. Nanobiosensors built with inactivated enzyme or cantilevers without immobilized enzyme were not capable of producing this type of signal (deflection). This technique will be very useful in detecting the passage of specific molecules through a cell wall (or a cell membrane for other types of cells). PMID- 10854856 TI - Characterization of a C-terminal-type kinesin-related protein from Dictyostelium discoideum. AB - We have determined the full sequence of K2, a kinesin-related protein (KRP) in Dictyostelium discoideum. Sequence homology and domain organization placed K2 in the ncd/Kar3 subfamily of the C-terminal-type KRPs. Bacterially expressed, truncated K2 showed ATP-dependent binding to microtubules and microtubule stimulated ATPase activity. K2-null cells grew and developed normally, suggesting overlapping functions of K2 with other microtubule motor(s). Overexpression of K2 caused partial mitotic arrest. Green fluorescent protein-tagged full-length K2 localized in the nucleus at the interphase and on the mitotic spindle during mitosis. These results suggest that K2 is a microtubule-dependent motor which may play some roles in mitotic spindles. PMID- 10854857 TI - Shedding and cleavage of the urokinase receptor (uPAR): identification and characterisation of uPAR fragments in vitro and in vivo. AB - Applying a novel, highly specific and sensitive immunoabsorption/Western blotting technique we have identified in vitro in conditioned cell culture medium and in vivo in human urine different soluble forms of the urokinase-type plasminogen activator receptor (uPAR/CD87). These include the uPAR fragment D2D3 and the never before identified domain 1 (D1) fragment. These forms correspond to fragments previously characterised as biologically active as inducers of chemotaxis and cell adhesion. We find that stimulation of U937 cells is associated with increased uPAR expression, cleavage of surface uPAR, and release of soluble fragments to the culture medium suggesting that monocytes are a source of the circulating and urinary soluble uPAR fragments found in vivo. Our study demonstrates that potentially biologically active uPAR fragments are produced in the human body, indicating a possible function in the regulation of not only proteolysis but also signal transduction related processes. PMID- 10854858 TI - Thrombin stimulation of platelets causes an increase in phosphatidylinositol 5 phosphate revealed by mass assay. AB - Phosphatidylinositol 5-phosphate (PtdIns5P), a novel inositol lipid, has been shown to be the major substrate for the type II PtdInsP kinases (PIPkins) ?Rameh et al. (1997) Nature 390, 192-196. A PtdInsP fraction was prepared from cell extracts by neomycin chromatography, using a protocol devised to eliminate the interaction of acidic solvents with plasticware, since this was found to inhibit the enzyme. The PtdIns5P in this fraction was measured by incubating with ?gamma (32)PATP and recombinant PIPkin IIalpha, and quantifying the radiolabelled PtdInsP(2) formed. This assay was used on platelets to show that during 10 min stimulation with thrombin, the mass level of PtdIns5P increases, implying the existence of an agonist-stimulated synthetic mechanism. PMID- 10854859 TI - Crystal structure of SULT2A3, human hydroxysteroid sulfotransferase. AB - The crystal structure of SULT2A3 human hydroxysteroid sulfotransferase has been solved at 2.4 A resolution in the presence of 3'-phosphoadenosine 5'-phosphate (PAP). The overall structure is similar to those of SULT1 enzymes such as estrogen sulfotransferase and the PAP binding site is conserved, however, significant differences exist in the positions of loops Pro14-Ser20, Glu79-Ile82 and Tyr234-Gln244 in the substrate binding pocket. Moreover, protein interaction in the crystal structure has revealed a possible dimer-directed conformational alteration that may regulate the SULT activity. PMID- 10854860 TI - Cloning and characterization of a root specific high-affinity sulfate transporter from Arabidopsis thaliana. AB - Hst1At (accession number AB018695) was identified from the Arabidopsis thaliana sequencing project on BAC T3F12, and the corresponding cDNA was isolated by reverse transcription-PCR. Southern blot analysis reveals a single copy of this gene. The cDNA encodes a root specific sulfate transporter of 649 amino acids. Heterologous expression of hst1At in a sulfate transport deficient yeast mutant shows that this gene encodes a high-affinity transport system ( approximately 2 microM). The transcript relative abundance increases in roots in response to sulfate deprivation, which correlated with increased root SO(4)(2-) influx capacity. These patterns were reversed upon sulfate addition to the medium and were accompanied by an increased glutathione level in roots. Feeding plants with cysteine or glutathione led to similar responses. Using buthionine sulfoximine, an inhibitor of glutathione synthesis, we show that glutathione rather than cysteine controls hst1At expression. PMID- 10854861 TI - Low-molecular-weight heparin for thrombophilia in pregnant women. AB - OBJECTIVE: Low-molecular-weight heparin (LMWH) is the anticoagulant of choice during pregnancy because it is associated with a low incidence of osteoporosis and thrombocytopenia. Antithrombotic therapy has recently been used to prevent pregnancy loss in high-risk patients with evidence of acquired or congenital thrombophilia. The aim of the present study was to gain further information on the teratogenic potential of LMWH in this patient group. METHODS: The study population included 46 patients with a history of recurrent abortions, intrauterine fetal death or intrauterine growth restriction (IUGR) and severe early-onset preeclampsia. Patients with a history of thromboembolism or positive findings for thrombophilia were prescribed LMWH (enoxaparin sodium, 40 mg daily) in combination with low-dose aspirin (100 mg daily) in the first trimester (group 1, n=14) or the second trimester (group 2, n=17); the remaining 15 patients received low-dose aspirin alone (group 3). RESULTS: No significant differences were noted between the groups in the incidence of congenital malformations or abortions, IUGR or preterm deliveries. One infant in group 1 had familial bilateral postaxial polydactyly of the hands and one in group 3 had patent ductus arteriosus. CONCLUSION: Despite the small size of the study groups, our results support the assumption that the use of LMWH is safe, at least as a teratogenic agent, in patients with thrombophilia throughout pregnancy. PMID- 10854862 TI - Seasonal variation in risk of anemia among pregnant Nepali women. AB - OBJECTIVE: We aimed to investigate whether there is any seasonal variation in risk of anemia among pregnant Nepali women. METHODS: We studied the hematocrit values in pregnant women (n=5495) attending Patan Hospital, Kathmandu for the first antenatal visit in the 2-year period from January 1994 until December 1995. In a sub-sample of subjects (n=2706), additional information was obtained through interviews and clinical examinations. Logistic regression models were used to analyze data. RESULTS: Mean hematocrit values recorded in the monsoon period were significantly lower than hematocrit values recorded in the winter. The prevalence of moderate (hematocrit 25-33%) and severe (hematocrit<25%) anemia was highest in September. CONCLUSIONS: We found a clear seasonal variation in risk of anemia, which was associated with rainfall and temperature. The monsoon seems to be a period when interventions may give the most beneficial effects. PMID- 10854863 TI - Prevalence of 'high risk' human papillomavirus in the lower genital tract of Brazilian gravidas. AB - The presence of high-risk human Papillomavirus types 16, 18 and 33 was examined in 125 pregnant patients with abnormal Papanicolau smears. Specimens of cervicovaginal cells were analyzed by a simplified method of slot-blot hybridization. The overall prevalence of those viral sequences was 48%, being: 22.4% of HPV16, 17.6% of HPV18, 4.0% for double HPV16 and 18 infestation and 4.0% of HPV33. Their prevalence in HPV positive cervical sample was alone respectively 46.6%, 36.6%, 8.3% and 8.3%. Besides the high incidence of those carcinogenic types and intense viral proliferation, a rapid progression from CIN to carcinoma was clinically observed in four pregnant patients. Our data may reinforce the idea that progesterone has a positive role to the persistence and transformation of 'high risk' HPV, particularly of HPV16. The enhanced detection of potentially malignant types during pregnancy should warn on the importance of early diagnosis and treatment of papillomatosis. PMID- 10854864 TI - Reduction of the cesarean delivery rate in Ecuador. AB - OBJECTIVE: This quasi-experimental study tested a method to safely reduce the rate of cesarean delivery in Ecuador. METHOD: Hospital policy was modified to provide co-management for cesarean candidates at the major maternity hospital in Quito. Cesarean rates before (n=14743) and after (n=12351) the intervention were compared by chi-square and multiple logistic regression with other major maternity hospitals (before, n=12514; after, n=9590). Characteristics of cesarean candidates who had vaginal or cesarean deliveries in the intervention hospital were compared by chi-square (n=1584). RESULT: Cesarean rates declined by 4.5% (P<0.001) in the intervention hospital. A smaller (2.1%, P<0.01) reduction occurred in the other major public hospital in Quito where students of the co principal investigator attempted to reduce cesarean delivery. Cesarean rates were unchanged in the public maternity hospitals of other major cities. CONCLUSION: Case co-management, a simple, locally appropriate, and inexpensive intervention, safely reduced surgical delivery, hospital stay and cost of care. PMID- 10854865 TI - Ultrasound assessment of fetal biparietal diameter and femur length during normal pregnancy in Iranian women. AB - OBJECTIVES: To determine the pattern of intrauterine growth and predicted biparietal diameter (BPD) and femur length (FL) at point in gestation of Iranian fetuses. METHODS: In an extensive and long-standing prospective study in Tehran, Iran 15693 BPD and 15594 FL measurements were obtained from the fetuses of 1324 normal pregnant women. Weekly mean values and the standard deviations (S.D. ) were calculated for both BPD and FL from 12 to 40 weeks of pregnancy. Comparison was also made between our results and previous Western studies using t-test analysis. RESULTS: Iranian fetuses had smaller BPD and shorter FL measurements in comparison with Western studies (P<0.05). There is a lag in BPD growth of our fetuses. The lag in FL growth is even more than BPD. Growth of the BPD and FL showed an asymptotic curve like that of Western studies but both of our values were lower. CONCLUSION: Ethnicity may influence ultrasonic fetal biometric measurements. PMID- 10854866 TI - Prevalence of hysterectomy in Ireland. AB - OBJECTIVES: To assess factors influencing the prevalence of hysterectomy in Ireland. METHODS: Analysis of results of a questionnaire completed by a population-based cohort of 17735 women aged 50-65 years attending for breast screening. RESULTS: Prevalence of hysterectomy was 22.2%, was increased in proportion to parity and was higher in younger women, those who had previously used oral contraception and those who had private health insurance; peak age at operation was 45-49 years. CONCLUSION: The relatively high prevalence parity reflects contraceptive practices and utilization of health service resources. PMID- 10854867 TI - Speculoscopy vs. the acetic acid test for cervical neoplasia. AB - OBJECTIVE: To compare the specificity of speculoscopy and the acetic acid test (AAT). METHODS: A randomized controlled trial was performed on 1150 women in a primary healthcare clinic. The main end-points were the sensitivity and specificity of speculoscopy and the AAT. RESULTS: The mean age of the patients was 37.7 years in the speculoscopy group (n=545) and 37.5 years in the AAT group (n=605). Cervicography (the golden standard) was positive in 11.3% of the participants, speculoscopy in 20.4% and the AAT in 25.1%. The sensitivity, specificity and the kappa value for speculoscopy were 53.5%, 83.6% and 0.23, respectively. For the AAT, the corresponding figures were 51.1%, 77.3% and 0.15. No statistically significant differences were found between these two groups. Of clinical importance, however, was a finding that speculoscopy did not miss a single case of high grade squamous intra-epithelial lesion or cancer whilst the AAT missed five such lesions. CONCLUSION: Speculoscopy could not be shown to have an improved specificity when compared with the acetic acid test. The low specificity of both the tests will result in the referral of too many patients for colposcopy. PMID- 10854868 TI - Classification of female urinary incontinence by the scored incontinence questionnaire. AB - OBJECTIVES: Our purpose was to investigate whether urinary incontinence (UI) severity could be graded by UI scores. METHODS: The study included 168 UI patients diagnosed by conventional procedures as stress incontinence (SI, n=108) or urge incontinence (URI, n=60). A questionnaire containing 15 items scored for stress-score (s-s) and urge-score (u-s) was administered to patients to assess the correlation between pad test values (PTVs) and s-s in SI and u-s in URI. RESULTS: Significant correlations were found between log[PTVs] and both s-s and u s: log[PTV(SI)]=0.051s-s-0.207 (r=0.830, P<0.0001) and log[PTV(URI)]=0.064u-s 0.459 (r=0.827, P<0. 0001). Based on theoretical PTVs obtained using these equations, s-s of 26-24, 23-18, and 17-10 in SI were equivalent to severe, moderate, and mild, respectively, and u-s of 22-19 and 18-12 in URI were equivalent to moderate and mild, respectively, according to the classification of the International Continence Society. CONCLUSION: The UI score should be a simple and useful procedure for use by general gynecologists to grade UI severity clinically. PMID- 10854869 TI - Uterine septum misdiagnosed on hysterosalpingogram. AB - OBJECTIVE: To evaluate the correctness of diagnosis of bicornuate uterus made on hysterosalpingogram. METHOD: Thirty-six women diagnosed on hysterosalpingogram to have bicornuate uterus were subjected to laparoscopy and hysteroscopy. RESULT: Two out of 36 women had bicornuate uterus whereas the remaining 34 had intrauterine septum. CONCLUSION: Diagnosis of bicornuate uterus made on hysterosalpingogram by radiologists is often incorrect and needs to be revised. PMID- 10854870 TI - Exposure to human immunodeficiency virus among healthcare workers in South Africa. AB - There have been no reports in the literature on occupational hazards of HIV in developing countries. The aim of this study was to evaluate occupational exposure to HIV in healthcare workers in Durban, South Africa. Individuals with occupational exposure to HIV were interviewed. Thirteen percent of the staff reported injuries with HIV positive patients. Registrars in training were the highest risk group (60%). Of the injuries, 94% were percutaneous and 65% occurred during emergency surgery. The commonest place of injury was the operating theater (46%) and the commonest procedure associated with accidental exposure was cesarean section (57%). Fifty-one percent were not wearing eye protection during procedures and although 83% initiated post-exposure prophylaxis (PEP), 48% discontinued treatment due to side effects of the drugs. Occupational exposure to HIV is common in the developing world. Rectifiable factors identified in this study that contributes to the milieu of occupational acquisition of HIV include less than proper adherence to universal precaution; inadequate documentation procedures and failure of a large percentage of respondents to complete post exposure prophylaxis. PMID- 10854871 TI - Appendicitis during pregnancy. PMID- 10854872 TI - Fetal acoustic stimulation for early intrapartum assessment of fetal well-being. PMID- 10854873 TI - Preliminary results of the International HELLP-Multicenter-Study. PMID- 10854874 TI - The effect of continuous combined oral estradiol and norethisterone on the renal and internal carotid artery pulsatility indices in postmenopausal women. PMID- 10854875 TI - Citations from the literature. This is a selection of abstracts taken from the literature in the field of gynecology and obstetrics which the Journal's editors feel may be of interest to our readers(1) PMID- 10854876 TI - Volumetric changes of articular cartilage during stress relaxation in unconfined compression. AB - The time-dependent lateral expansion and load relaxation of cartilage cylinders subjected to unconfined compression were simultaneously recorded. These measurements were used to (1) test the assumption of incompressibility for articular cartilage, (2) measure the Poisson's ratio of articular cartilage in compression and (3) investigate the relationship between stress relaxation and volumetric change. Mechanical tests were performed on fetal, calf, and adult humeral head articular cartilage. The instantaneous Poisson's ratio of adult cartilage was 0.49+/-0.08 (mean+S.D.), thus confirming the assumption of incompressibility for this tissue. The instantaneous Poisson's ratio was significantly lower for calf (0. 38+/-0.04) and fetal cartilage (0.36+/-0.04). The equilibrium Poisson's ratio, i.e. true Poisson's ratio of the solid matrix, was significantly higher for the adult tissue (0.26+/-0.11) compared to both the fetal (0.09+/-0.02) and calf (0.11+/-0.03) cartilage. A linear relationship between time-matched load and lateral expansion after the first minute of stress relaxation was observed. PMID- 10854877 TI - Interphalangeal joint and tendon forces: normal model and biomechanical consequences of surgical reconstruction. AB - Soft tissue reconstructive surgery for rheumatoid-related proximal interphalangeal joint deformities frequently fails to produce the long-term predicted results. Detailed information on the biomechanics of this joint, under both normal and pathological conditions, is required to assess the efficacy of such surgical intervention. A biomechanical model of the proximal interphalangeal joint has been developed to investigate tendon and joint loading during real life three-dimensional activities. Based on a rigid body mechanics approach, the model uses high resolution MRI scans to obtain anatomical tendon and bone geometries in conjunction with three-dimensional kinematic and loading data. The model incorporates an optimisation routine which minimises overall maximum tendon stress in the eight individual elements considered. Radial and ulnar joint force components are included at the proximal interphalangeal joint level. Two simulated pathological versions of the mathematical model are developed to accommodate the altered anatomic relationships after tendon reconstructive surgery. Joint forces of up to 450N and common usage of the extensor mechanism during normal pinching and grasping activities are predicted. The ulnar lateral bands of the extensor tendon are generally loaded to a greater extent than the radial bands. Extensor tendon and joint forces in the simulated pathological models are significantly higher than those in the normal model. Combined with the poor tendon quality of rheumatoid arthritis patients generally, these amplified internal forces may lead to further joint deformation. PMID- 10854878 TI - Scapular kinematics: effects of altering the Euler angle sequence of rotations. AB - An analysis of Euler angle sequences is presented for the scapula. In vivo kinematics were collected with a magnetic tracking device on eight healthy volunteers. To ensure accurate representation of scapular motion, pins were rigidly drilled into the scapular spine. Three rotations of the scapula with respect to the thorax were recorded during humeral elevation in the scapular plane: posterior (or backward) tilting, upward (or lateral) rotation and external rotation (or retraction). Rotations using all six possible Euler angle sequences were calculated for which each angle was represented only once. The sequence proposed by an International Society of Biomechanics subcommittee on shoulder motion (external rotation, upward rotation, posterior tilting) is consistent with both research- and clinical-based two-dimensional representations of scapular motion. Results from the present study indicate that changing sequence results in significant alterations in the description of motion, with differences up to 50 degrees noted for some angles. Therefore, in order to compare results across different laboratories, it is recommended that the proposed standard sequence be adopted. PMID- 10854879 TI - Three-dimensional deformation and stress distribution in an analytical/computational model of the anterior cruciate ligament. AB - In this study, a constitutive equation for the ACL composite was formulated, and 3-D finite deformations and stress distributions of the ACL were calculated using a finite element method to simulate knee flexion. The mathematical model of the ACL was created by a structurally motivated phenomenological approach. It was assumed that the ACL can be ideally represented as a homogeneous hyperelastic matrix (Mooney-Rivin material) in which two families of densely distributed extensible fibers are embedded; the fibers in one family have a parallel orientation, the other fibers extend radially in eight equally spaced directions. The non-linear stress-strain characteristic exhibited by collagen fibers was represented by a tri-linear curve. Simulation was performed and the results provided some original data; the stress distribution within the ACL body as well as that over the surface, the 3-D deformations and stress distributions of the ACL viewed from other sides in addition to those from the medial side, and the variations of the stress distribution pattern in the ACL which occurred when the tibia was displaced anteriorly or posteriorly. PMID- 10854880 TI - A two-dimensional fluid-structure interaction model of the aortic valve [correction of value]. AB - Failure of synthetic heart valves is usually caused by tearing and calcification of the leaflets. Leaflet fiber-reinforcement increases the durability of these valves by unloading the delicate parts of the leaflets, maintaining their physiological functioning. The interaction of the valve with the surrounding fluid is essential when analyzing its functioning. However, the large differences in material properties of fluid and structure and the finite motion of the leaflets complicate blood-valve interaction modeling. This has, so far, obstructed numerical analyses of valves operating under physiological conditions. A two-dimensional fluid-structure interaction model is presented, which allows the Reynolds number to be within the physiological range, using a fictitious domain method based on Lagrange multipliers to couple the two phases. The extension to the three-dimensional case is straightforward. The model has been validated experimentally using laser Doppler anemometry for measuring the fluid flow and digitized high-speed video recordings to visualize the leaflet motion in corresponding geometries. Results show that both the fluid and leaflet behaviour are well predicted for different leaflet thicknesses. PMID- 10854881 TI - Biomechanical and morphological properties in rat large intestine. AB - Intestinal stress-strain distributions are important determinants of intestinal function and are determined by the mechanical properties of the intestinal wall, the physiological loading conditions and the zero-stress state of the intestine. In this study the distribution of morphometric measures, residual circumferential strains and stress-strain relationships along the rat large intestine were determined in vitro. Segments from four parts of the large intestine were excised, closed at both ends, and inflated with pressures up to 2kPa. The outer diameter and length were measured. The zero-stress state was obtained by cutting rings of large intestine radially. The geometric configuration at the zero-stress state is of fundamental importance because it is the basic state with respect to which the physical stresses and strains are defined. The outer and inner circumferences, wall thickness and opening angle were measured from digitised images. Subsequently, residual strain and stress-strain distributions were calculated. The wall thickness and wall thickness-to-circumference ratio increased in the distal direction. The opening angle varied between approximately 40 and approximately 125 degrees with the highest values in the beginning of proximal colon (F=1.739, P<0.05). The residual strain at the inner surface was negative indicating that the mucosa-submucosal layers of the large intestine in no-load state are in compression. The four segments showed stress-strain distributions that were exponential. All segments were stiffer in longitudinal direction than in the circumferential direction (P<0.05). The transverse colon seemed stiffest both in the circumferential and longitudinal directions. In conclusion, significant variations were found in morphometric and biomechanical properties along the large intestine. The circumferential residual strains and passive elastic properties must be taken into account in studies of physiological problems in which the stress and strain are important, e.g. large intestinal bolus transport function. PMID- 10854882 TI - Influence of load carrying on loads in internal spinal fixators. AB - After spinal stabilization with an implant, patients want to know whether carrying loads will endanger their spine and their implant. Therefore, the effect of carrying a weight on the loading of internal spinal fixation devices was determined in 10 patients using instrumented, telemeterized internal fixators. Patients carried a 5kg dumbbell in one hand as well as up to 20kg in both hands. Compared to normal standing carrying a weight caused only a slight increase of the fixator loads in all patients. The maximum flexion bending moment in the fixators was then lower than during walking. Carrying a weight even decreased the axial compression force in the fixators of the patients with the T11 or T12 vertebra bridged. Patients also raised their arms in anteversion while holding weights of up to 5kg with both hands. This increased the flexion bending moment in the fixators. Only a small part of a carried weight is taken over by internal fixators. Carrying a shopping bag only slightly increases the risk of pedicle screw breakage, however, since the spine has to support most of the additional weight, it represents a risk with regard to correction loss and bone sintering. PMID- 10854883 TI - Theoretical considerations on cocontraction of sets of agonistic and antagonistic muscles. AB - It is well known that static, non-linear minimization of the sum of the stress in muscles to a certain power cannot predict cocontraction of pairs of one-joint antagonistic muscles. In this report, we prove that for a single joint either all agonistic muscles cocontract or all are silent. For two-joint muscles, we show that lengthening and shortening of muscles corresponds closely to zero force and non-zero force states, respectively. This gives a new physiological interpretation of situations in which cocontraction of pairs of two-joint antagonistic muscles is predicted by these static non-linear optimization approaches. PMID- 10854884 TI - Sit-to-stand motor strategies investigated in able-bodied young and elderly subjects. AB - For the execution of a certain motor task, a motor strategy is chosen by each individual among those that are consistent with the structural and functional constraints of his/her locomotor system, and that tends to maximise the effectiveness of the motor act. The identification of this strategy allows for the assessment of the individual's functional status. This study aimed at identifying the motor strategies adopted for the execution of the sit-to-stand motor task, at different speeds and initial postures, in a sample of 35 community dwelling elders and in a sample of 16 young able-bodied individuals. This was done using a method, least perceivable to the test subject and "economical" for the experimenter, which entailed the recording of external forces only. A musculo skeletal system model, based on a telescopic inverted-pendulum (TIP) moved by a linear and two rotational muscle-equivalent actuators, was then used. Parameters describing the kinematics and dynamics of these actuators were extracted and submitted to statistical analysis. Different motor strategies were identified in the two age groups, as well as associated with both a different initial posture (ankle dorsiflexion angle) and speed of execution of the motor task. In particular, the elder group, as compared with the young group, prior to seat-off tended to flex the trunk more, thus bringing the CM closer to the base of support, and at a higher velocity, thus gaining a higher momentum. After seat off, elders rotated the body forward and, only after having brought their CM over the base of support, effectively started elevation. Both global muscular effort and coordination effort associated with the achievement of balance and raising were lower. However, maximal speed was also lower. The above results indicated that the elders could count on a lower functional reserve than the young individuals and, from the methodological viewpoint, that the TIP approach is a good candidate for subject-specific functional evaluation in a clinical context. PMID- 10854885 TI - A mathematical model of umbilical venous pulsation. AB - Pulsations in the fetal heart propagate through the precordial vein and the ductus venosus but are normally not transmitted into the umbilical vein. Pulsations in the umbilical vein do occur, however, in early pregnancy and in pathological conditions. Such transmission into the umbilical vein is poorly understood. In this paper we hypothesize that the mechanical properties and the dimensions of the vessels do influence the umbilical venous pulsations, in addition to the magnitude of the pressure and flow waves generated in the fetal atria. To support this hypothesis we established a mathematical model of the umbilical vein/ductus venosus bifurcation. The umbilical vein was modeled as a compliant reservoir and the umbilical vein pressure was assumed to be equal to the stagnation pressure at the ductus venosus inlet. We calculated the index of pulsation of the umbilical vein pressure ((max-min)/mean), the reflection and transmission factors at the ductus venosus inlet, numerically and with estimates. Typical dimensions in the physiological range for the human fetus were used, while stiffness parameters were taken from fetal sheep. We found that wave transmission and reflection in the umbilical vein ductus venosus bifurcation depend on the impedance ratio between the umbilical vein and the ductus venosus, as well as the ratio of the mean velocity and the pulse wave velocity in the ductus venosus. Accordingly, the pulsations initiated by the fetal heart are transmitted upstream and may arrive in the umbilical vein with amplitudes depending on the impedance ratio and the ratio between the mean velocity and the pulse wave velocity in the ductus venosus. PMID- 10854886 TI - Observations of microdamage around osteocyte lacunae in bone. AB - Tensile microdamage was examined using laser scanning confocal microscopy in beam specimens of bovine, equine and human long bones loaded in vitro and whole specimens of rat ulnae loaded in vivo. Microcracks were observed to initiate frequently at osteocyte lacunae. The implication is that osteocyte lacunae act as stress concentrating features in bone. This association provides a potential mechanism for the detection of strain and/ or damage by osteocytes in bone. PMID- 10854887 TI - A new ambulatory foot pressure device for patients with sensory impairment. A system for continuous measurement of plantar pressure and a feed-back alarm. AB - Abnormal and excessive plantar pressure is a major risk factor for the development of foot ulcers in patients with loss of protective pain sensation. Repeated pressure with each step can result in inflammation at specific points, followed by ulcer formation. Patients with peripheral nerve disease are unable to prevent the development of such lesions, which often lead to amputation. For this reason, it has been suggested that a fundamental therapeutic intervention should be the reduction of high plantar pressure. We have developed a portable, battery operated ambulatory foot pressure device (AFPD) which has two important functions: (1) to determine the areas of high plantar pressure, and (2) to provide an acoustic alarm, adjusted to a specific pressure load, which is triggered when weight-bearing exceeds the predetermined plantar pressure. A memory of plantar pressure parameters allows for downloading of the data and sequential analysis during the investigation period. Such an alarm device could replace the lack of pain sensation and may play an important role in the prevention of ulcer development and lower extremity amputation. PMID- 10854888 TI - Determining the optimal flexion-extension axis of the elbow in vivo - a study of interobserver and intraobserver reliability. AB - In the current study the interobserver and intraobserver reliability of a recently developed method to obtain the position and orientation vectors of the flexion-extension axis of the elbow in vivo is determined. The method uses the Flock of Birds six degrees-of-freedom electromagnetic tracking device. Ten subjects performed three trials comprising five flexion and extension cycles. The movements of the forearm with respect to the upper arm were recorded. Observer A measured two trials and observer B measured one trial. Optimal instantaneous helical axes were calculated in a humeral coordinate system for each trial. Intraclass correlation coefficients and 99% confidence intervals were computed to compare the three measurements. Zero was in the range of all the narrow confidence intervals, which is strong indication for resemblance. Interobserver intraclass correlation coefficients values for orientation vectors were good to excellent and intraobserver values were fair to good. The intraclass correlation coefficients values for position vectors were lower, probably due to the lack of variance between subjects. It is concluded that the method is reliable and can be used in certain clinical settings. PMID- 10854889 TI - The effect of the point of application of anterior tibial loads on human knee kinematics. AB - Coupled axial tibial rotation in response to an anterior tibial load has been used as a common diagnostic measurement and as a means to load the ligamentous structures during laboratory tests. However, the exact location of the point of application of these loads as well as the corresponding sensitivity of the coupled tibial rotation to this point can have an effect on the function of the soft tissues at the joint. Therefore, the purpose of this study was to determine the effects of four different points of application of the anterior tibial load on the anterior tibial translation and coupled axial tibial rotation. The four points include: (1) geometric point - midway between the collateral ligament insertion sites on the tibia, (2) clinical point - a position that attempts to simulate clinical diagnostic tests, (3) medial point - a position medial to the geometric point and (4) lateral point - a position lateral to the clinical point. A robotic/universal force-moment sensor testing system was used to apply the anterior tibial load at the four points of application and to record the resulting joint motion. Anterior tibial translation in response to an anterior tibial load of 100N was found not to vary between the four points of application of the anterior tibial load at all flexion angles examined. However, internal tibial rotation was found for the lateral point (13+/-10 degrees at 30 degrees of knee flexion) in all specimens and clinical point (8+/-10 degrees at 30 degrees of knee flexion) while external rotation resulted when the load was applied at the medial point (-8+/-7 degrees at 30 degrees of knee flexion). Both internal and external tibial rotations occurred throughout the range of flexion when the tibial load was applied at the geometric point. The results suggest that the clinical point should be used as the point of application of the anterior tibial load whenever clinical examinations are simulated and multi-degree-of-freedom joint and soft tissue function are examined. PMID- 10854890 TI - Tensile experiments and SEM fractography on bovine subchondral bone. AB - Subchondral bone undecalcified samples, extracted from bovine femoral heads, are subjected to a direct tensile load. The Young's modulus of each sample is determined from repeated tests within the elastic limit. In a last test, the tensile load is increased up to the specimen failure, determining the ultimate tensile strength. The investigation is performed on both dry and wet specimens. The measured Young's modulus for dry samples is 10.3+/-2.5GPa, while that of wet samples is 3.5+/-1.2GPa. The ultimate tensile strengths are 36+/-10 and 30+/ 7.5MPa for dry and wet specimens, respectively. SEM micrographs of failure surfaces show characteristic lamellar bone structures, with lamellae composed of calcified collagen fibers. Rudimentary osteon-like structures are also observed. Failure surfaces of wet samples show a marked fiber pull-out, while delamination predominates in dry samples. The obtained results are interpreted on the basis of the deformation mechanisms typical of fiber-reinforced laminated composite materials. PMID- 10854891 TI - A rigid body model of the forearm. AB - In this article the forearm, with its complex, continuous motion of masses during pronation/supination, was approximated by a rigid body model consisting of a radial segment rotating around an ulnar segment. The method used to obtain the model parameters is based on three-dimensional voxel data that include velocity information. We propose a criterion that allows the voxels to be attributed to either of the two segments. It is based on the notion that the rotational kinetic energy determined from the voxel data equals the kinetic energy of the rigid body model. To obtain a three-dimensional smoothing we further propose a parameterization of the shape of both segments. These shapes can then be used to determine the dynamic integrals of the segments, i.e. mass, center of mass, and inertia. Using this approach we determined all model parameters for a human forearm from three series of MRI scans in a supinated, a pronated, and an intermediate position. In the appendix, a procedure is described that allows the dynamic quantities to be scaled homogeneously without recalculation of the integrals. Thus, this article provides all essential parameters required for three-dimensional dynamic simulations of general movements of the forearm. PMID- 10854892 TI - Contribution, development and morphology of microcracking in cortical bone during crack propagation. AB - A fracture mechanics study of cortical bone is presented to investigate the contribution, development morphology of microcracking in cortical bone during crack propagation. Post-hoc analyses of microcrack orientation, crack propagation velocity and fracture surface roughness were conducted on previously tested human and bovine bone compact tension specimens. It was found that, consistent with its higher toughness, bovine bone formed significantly more longitudinal, transverse and inclined microcracks than human bone. However, in human bone more of the microcracks that formed were longitudinal than transverse or inclined, a feature that would optimise bone's toughness. Crack propagation velocity in human and bovine bone displayed the same characteristic pattern with crack extension, where an increase in velocity is followed by a consequent decrease and vice versa. On the basis of this pattern, a model or crack propagation has been proposed. It provides a detailed account of mocrocrack formation and contribution towards the propagation of a fracture crack. Analyses of fracture surfaces indicated that, consistent with its higher toughness, bovine bone displays a rougher surface than human bone but they both have the same basic fractured element, i.e. a mineralised collagen fibril. PMID- 10854893 TI - Stress and micromotion in the taper lock joint of a modular segmental bone replacement prosthesis. AB - The stress distribution within the components and the micromotion of the interface significantly influence the long-term function of the taper lock joint in a modular segmental bone replacement prosthesis. Bending-induced gap opening between the cone and the sleeve can lead to an inflow of biological fluids, and thus accelerate implant corrosion. Local areas of high stress can also accelerate the corrosive processes and initiate local yielding, which may lead to a fracture in one of the components. In this study, a 3-D finite element (FE) model of a modular segmental bone replacement prosthesis was developed to study the interface micromotion and component stress distribution under the maximum loads applied during gait for a taper lock joint with multiple material combinations. Bending was the main cause of the local high stresses and interface separation within the taper joint. For Ti6A14V components, cortical bone bridging and ingrowth across the taper lock gap reduced the peak stress by 45% and reduced the contact interface separation by 55%. Such tissue formation around the taper lock joint could also form a closed capsule to restrict the migration of potential wear particles and thus prevent the biologic process of bone resorption induced by metal debris. PMID- 10854894 TI - GABA potentiation: a logical pharmacological approach for the treatment of acute ischaemic stroke. AB - It has been shown that enhancing the function of the major inhibitory neurotransmitter GABA decreases glutamatergic activity in the brain. Since increased glutamatergic activity is the major primary event that results in cell death following an acute hypoxic-ischaemic stroke, GABAmimetic drugs might therefore be expected to be neuroprotective. This review examines the evidence that GABAergic function is acutely depressed following an ischaemic insult, and also reviews the data that suggest that increasing cerebral GABA concentration has a neuroprotective effect, as does the administration of some (but not all) GABAmimetic agents. The GABA uptake inhibitor CI-966, the GABA(A) agonist muscimol and the GABA(A)mimetic clomethiazole have all been shown to be neuroprotective in animal models of stroke when given after the ischaemic insult. In contrast, benzodiazepines and particularly barbiturates, although potent GABA(A) potentiators, have shown little promise as neuroprotectants. The diversity of GABA(A) receptor subtypes and the in vivo efficacy of certain GABA(A) receptor ligands in animal models of stroke suggests that GABAmimetic drugs are an undervalued approach to stroke therapy. PMID- 10854895 TI - Somato-synaptic variation of GABA(A) receptors in cultured murine cerebellar granule cells: investigation of the role of the alpha6 subunit. AB - Electrophysiological investigation of cultured cerebellar murine granule cells revealed differences between the GABA(A) receptors at inhibitory synapses and those on the cell body. Specifically, mIPSCs decayed more rapidly than cell body receptors deactivated, the mean single channel conductance at the synapse (32 pS) was greater than that at cell body (21 pS) and only cell body receptors were sensitive to Zn(2+) (150 microM), which depressed response amplitude by 82+/-5% and almost doubled the rate of channel deactivation. The GABA(A) receptor alpha6 subunit is selectively expressed in cerebellar granule cells. Although concentrated at synapses, it is also found on extrasynaptic membranes. Using a mouse line (Deltaalpha6lacZ) lacking this subunit, we investigated its role in the somato-synaptic differences in GABA(A) receptor function. All differences between cell body and synaptic GABA(A) receptors observed in wild-type (WT) granule cells persisted in Deltaalpha6lacZ cells, thus demonstrating that they are not specifically due to the cellular distribution of the alpha6 subunit. However, mIPSCs from WT and Deltaalpha6lacZ cells differed in both their kinetics (faster decay in WT cells) and underlying single channel conductance (32 pS WT, 25 pS Deltaalpha6lacZ). This provides good evidence for a functional contribution of the alpha6 subunit to postsynaptic GABA(A) receptors in these cells. Despite this, deactivation kinetics of mIPSCs in WT and Deltaalpha6lacZ granule cells exhibited similar benzodiazepene (BDZ) sensitivity. This suggests that the enhanced BDZ-induced ataxia seen in Deltaalpha6lacZ mice may reflect physiological activity at extrasynaptic receptors which, unlike those at synapses, display differential BDZ-sensitivity in WT and Deltaalpha6lacZ granule cells (Jones, A.M., Korpi, E.R., McKernan, R.M., Nusser, Z., Pelz, R., Makela, R., Mellor, J.R., Pollard, S., Bahn, S., Stephenson, F.A., Randall, A.D., Sieghart, W., Somogyi, P., Smith, A.J.H., Wisden, W., 1997. Ligand-gated ion channel partnerships: GABA(A) receptor alpha(6) subunit inactivation inhibits delta subunit expression. Journal of Neuroscience 17, 1350-1362). PMID- 10854896 TI - Characterisation of GABA(A) receptors in fetal, neonatal and adult ovine brain: region and age related changes and the effects of allopregnanolone. AB - Progesterone metabolites acting via GABA(A) receptors suppress central nervous system (CNS) activity. The aim of the present study was to examine binding characteristics of GABA(A) receptors in fetal, newborn and adult sheep brains using [(35)S]TBPS, and to determine the effects of allopregnanolone on this binding. Receptor affinity (K(D)) and density (B(MAX)) in the brainstem were not different in fetal, newborn (1-2 days old) and adult brains. In the hypothalamus K(D) and B(MAX) increased significantly in the fetus between 85 and 128 days gestation, and were then similar to postnatal and adult values. In the frontal cortex K(D) and B(MAX) increased progressively between 85 days and term ( approximately 147 days gestation), and were then not different from postnatal and adult values. The K(i) values for the GABA(A) receptor antagonist picrotoxin was similar at all ages. Allopregnanolone inhibited [(35)S]TBPS binding in the presence of 5 microM GABA, but enhanced binding in the absence of GABA. These results show that (i), functional GABA(A) receptors are present in the fetal brain from at least 85 days gestation; (ii), 3alpha-pregnane steroids modify receptor affinity in the late gestation fetal brain; and (iii) there are region specific changes in GABA(A) receptor binding parameters. Steroid modulation of the GABA(A) receptor in the fetal brain is likely to influence fetal CNS activity in late gestation. PMID- 10854898 TI - Clozapine inhibits synaptic transmission at GABAergic synapses established by ventral tegmental area neurones in culture. AB - Elucidation of the mechanism of action of the atypical antipsychotic clozapine is complicated by the finding that this molecule interacts with multiple targets including dopaminergic and serotonergic receptors. Binding studies have suggested that clozapine also antagonises GABA(A) receptors, but physiological evidence for such a block at functional synapses is lacking. In this study, we explored this antagonism by using electrophysiological techniques on GABAergic neurones of the ventral tegmental area in culture. Inhibitory post-synaptic currents (IPSCs) evoked in isolated GABAergic neurones were found to be dose-dependently inhibited by clozapine. Compatible with a post-synaptic mechanism, we found that membrane currents evoked by exogenous applications of GABA were similarly dose-dependently inhibited by clozapine. An analysis of miniature inhibitory post-synaptic currents (mIPSCs) showed that clozapine reduced the amplitude of quantal events in a way similar to SR-95531, a specific GABA(A) receptor antagonist. Both drugs caused a similar leftward shift of the cumulative probability distribution of mIPSC amplitudes. This suggests that clozapine acts on both synaptic and extrasynaptic GABA(A) receptors. In conclusion, our work demonstrates that clozapine produces a functional antagonism of GABA(A) receptors at synapses. Because this effect occurs at concentrations that could be found in the brain of patients treated with clozapine, a reduction in GABAergic synaptic transmission could be implicated in the therapeutic actions and/or side-effects of clozapine. PMID- 10854897 TI - Modulation of GABAergic synaptic transmission by the non-benzodiazepine anxiolytic etifoxine. AB - We have investigated the effects of 2-ethylamino-6-chloro-4-methyl-4-phenyl-4H 3,1-benzoxazine hydrochloride (etifoxine) on GABA(A) receptor function. Etifoxine displaced [(35)S]TBPS (t-butylbicyclophosphorothionate) from GABA(A) receptors of rat cortical membranes with an IC(50) of 6.7+/-0.8 microM and [(3)H]PK11195 from peripheral (mitochondrial)-type benzodiazepine receptors (PBRs) of rat heart homogenates with an IC(50) of 27.3+/-1.0 microM. Etifoxine displayed anxiolytic properties in an anticonflict test in rats, and potentiated GABA(A) receptor mediated membrane currents elicited by submaximal (5-10 microM) but not saturating (0.5 mM) concentrations of GABA in cultured rat hypothalamic and spinal cord dorsal horn neurones. In hypothalamic cultures, etifoxine induced a dose-dependent inward current for concentrations >1 microM which reflected the post-synaptic potentiation of a small ( approximately 20 pA) tonic and bicuculline-sensitive GABA(A) receptor-gated Cl(-) current. Etifoxine also increased the frequency of spontaneous and miniature GABAergic inhibitory post synaptic currents without changing their amplitude and kinetic characteristics. Both effects of etifoxine were insensitive to flumazenil (10 microM), an antagonist of central-type benzodiazepine sites present at GABA(A) receptors, but were partly inhibited by PK11195 (10 microM) an antagonist of PBRs which control the synthesis of neurosteroids. Our results indicate that etifoxine potentiates GABA(A) receptor-function by a direct allosteric effect and by an indirect mechanism involving the activation of PBRs. PMID- 10854899 TI - Protein phosphatase 2B inhibitors mimic the action of arachidonic acid and prolong the facilitation of glutamate release by group I mGlu receptors. AB - We have addressed the role of arachidonic acid in the facilitation of glutamate release by group I metabotropic glutamate (mGlu) receptors. The activation of these receptors with the specific agonist 3,5-dihydroxyphenylglycine (DHPG) failed to enhance the cumulative Ca(2+)-dependent release of glutamate evoked by a 5 min depolarization with 4-aminopyridine, in the absence but not in the presence of arachidonic acid. However, DHPG, in the absence of arachidonic acid, transiently enhanced diacylglycerol levels, transiently potentiated 4AP-evoked depolarization, and significantly enhanced the fast but not the slow component of glutamate release observed after prolonged stimulations of nerve terminals. Further evidence that DHPG was able to initiate release facilitation in the absence of arachidonic acid was obtained in experiments where the protein phosphatase 2B (cyclosporine A and cypermethrine) but not protein phosphatase 1 or 2A inhibitors (okadaic acid and calyculin A) facilited glutamate release to a maximal extent comparable to that induced by arachidonic acid. We conclude that an active protein phosphatase 2B (calcineurin) dephosphorylates the presynaptic target/s responsible for facilitation of glutamate release. PMID- 10854900 TI - Metabotropic glutamate autoreceptors of the mGlu(5) subtype positively modulate neuronal glutamate release in the rat forebrain in vitro. AB - In the present study we have examined the role of presynaptic group I metabotropic glutamate (mGlu) receptors in the control of neuronal glutamate release using rat forebrain slices pre-loaded with [(3)H]D-aspartate. We have also addressed the question of which group I mGlu receptor subtype, mGlu(1) or mGlu(5), mediates the facilitatory response observed by the use of a range of established and some more novel agonists and antagonists showing selectivity for these receptors. The electrically-stimulated release of pre-loaded [(3)H]D aspartate from rat forebrain slices was markedly potentiated by the potent group I mGlu receptor agonist, L-quisqualic acid (L-QUIS), in a concentration-dependent manner (EC(50) 17.31 microM). This response was inhibited by the mGlu receptor antagonists (S)-MCPG (100 microM) and (RS)-MTPG (100 microM) but not by the AMPA type ionotropic glutamate receptor antagonist, NBQX (100 microM). The selective group I mGlu receptor agonist (S)-3, 5-dihydroxyphenylglycine ((S)-DHPG) also enhanced electrically-stimulated efflux of label, although responses diminished with high (10-100 microM) concentrations of the agonist. Maximum responses were fully restored when (S)-DHPG (10 microM) was applied in the presence of the proposed mGlu(5) receptor desensitization inhibitor, cyclothiazide (10 microM). The positive modulatory response to (S)-DHPG (1 microM) was powerfully inhibited by (S)-MCPG (IC(50) 0.08 microM) but was resistant to the mGlu(1) receptor antagonists, (RS)-AIDA (1-500 microM), CPCCOEt (0.1-100 microM) and (+)-2-methyl 4-carboxyphenylglycine (LY367385) (0.1-10 microM). The recently developed, selective mGlu(5) receptor agonist (RS)-2-chloro-5-hydroxyphenylglycine ((RS) CHPG) enhanced electrically-stimulated [(3)H]D-aspartate efflux from rat forebrain slices with a similar concentration-response profile to that of (S) DHPG. Responses to this receptor subtype-selective agonist were also blocked by (S)-MCPG (IC(50) 1.13 microM) but were unaffected by (RS)-AIDA (500 microM), CPCCOEt (100 microM) or LY367385 (10 microM). These results indicate that the positive modulation of neuronal glutamate release seen in the rat forebrain in the presence of group I mGlu receptor agonists is mediated by presynaptically located mGlu(5) glutamate autoreceptors. The pharmacological profile of these receptors appears to be distinct from that of postsynaptic mGlu receptors. Novel antagonists acting at these presynaptic receptors may provide new drugs for the experimental therapy of a range of acute or chronic neurodegenerative disorders. PMID- 10854901 TI - Anticonvulsant activity of two metabotropic glutamate group I antagonists selective for the mGlu5 receptor: 2-methyl-6-(phenylethynyl)-pyridine (MPEP), and (E)-6-methyl-2-styryl-pyridine (SIB 1893). AB - The selective mGlu5 antagonists, MPEP, 2-methyl-6-phenylethynyl-pyridine, and SIB1893, (E)-6-methyl-2-styryl-pyridine, have been evaluated as antiepileptic drugs in DBA/2 mice and lethargic mice. Clonic seizures induced by the selective mGlu5 agonist, (R,S)-2-chloro-5-hydroxyphenylglycine (CHPG), 3 micromol intracerebroventricularly (i.c.v.), are potently suppressed by both compounds (MPEP, ED(50)=0.42 [0.28-0.62] mg/kg intraperitoneally (i.p.); SIB 1893 ED(50)=0.19 [0.11-0.33] mg/kg i.p. ). Clonic seizures induced by the mGlu1,5 agonist, 3, 5-dihydroxyphenylglycine (DHPG), 1.5 micromol i.c.v., are less potently suppressed by both compounds (MPEP, ED(50)=22 [13-38] mg/kg i.p., 110 [67-180] nmol i.c.v.; SIB1893, ED(50)=31 [18-54] mg/kg i.p. , 95 [82-110] nmol i.c.v.). Sound-induced seizures in DBA/2 mice are suppressed at 15 min by MPEP and SIB 1893 (MPEP ED(50) clonic seizures=18 [10-32] mg/kg i.p., 93 [69-125] nmol i.c.v.; tonic seizures=6.1 [4.5-8.3] mg/kg i.p., 46 [26-80] nmol i.c.v.; SIB 1893 ED(50) clonic seizures=27 [17-44] mg/kg i.p., 825 [615-1108] nmol i. c.v., tonic seizures=5.4 [3.4-8.6] mg/kg i.p., 194 [113-332] nmol i. c.v.). The ED(50) for MPEP for impaired rotarod performance is 128 [83-193] mg/kg i.p., at 15 min, i.e. a therapeutic index for sound-induced seizures of 5-20. In lethargic mice (lh/lh), a genetic absence model, MPEP, 50 mg/kg i.p., caused a marked reduction in the incidence of spontaneous spike-and-wave discharges. These selective antagonists of mGlu5 block seizures due to activation of mGlu5 at very low systemic doses. At rather higher doses they block convulsive and non-convulsive primary generalised seizures. PMID- 10854902 TI - LY377770, a novel iGlu5 kainate receptor antagonist with neuroprotective effects in global and focal cerebral ischaemia. AB - We have evaluated the neuroprotective effects of the decahydroisoquinoline LY377770, a novel iGlu5 kainate receptor antagonist, in two models of cerebral ischaemia. Global ischaemia, induced in gerbils by bilateral carotid artery occlusion (BCAO) for 5 min, produced a large increase in locomotor activity at 96 hr post-occlusion and a severe loss of CA1 cells in the hippocampus histologically at 120 hr post-occlusion. LY377770 (80 mg/kg i.p. 30 min before or 30 min after BCAO followed by 40 mg/kg i.p. administered at 3 and 6 hr after the initial dose) attenuated the ischaemia-induced hyperactivity and provided (92%) and (29%) protection in the CA1 cells respectively. This protection was greater than that seen with maximally tolerated doses of other glutamate receptor antagonists (CGS19755, CPP, MK-801, ifenprodil, eliprodil, HA-966, ACEA1021, L701,324, NBQX, LY293558, GYKI52466 and LY300164). Focal ischaemia was induced by infusing 200 pmol of endothelin-1 (Et-1) adjacent to the middle cerebral artery and LY377770 was administered at 80 mg/kg i.p. immediately, 1 or 2 hr post occlusion followed by 40 mg/kg i.p. 3 and 6 hr after the first dose. The infarct volume, measured 72 hr later, was reduced by LY377770 when given immediately (P<0.01), at 1 hr (P<0.05) but not significantly at 2 hr post-occlusion. Reference compounds, LY293558 (20 mg/kg i.p. and then 10 mg/kg as above) and MK 801 (2.5 mg/kg i.p. ), both administered immediately post-occlusion produced significant (P<0.05) but somewhat less neuroprotection. In parallel microdialysis studies, LY377770 (75 mg/kg i.p.) attenuated ischaemia-induced increases in extracellular levels of glutamate, but not of dopamine. In conclusion, these results indicated that iGlu5 kainate receptors play a central role in ischaemic brain damage following global and focal cerebral ischaemia. LY377770 is a novel, soluble, systemically active iGlu5 antagonist with efficacy in global and focal ischaemia, even when administered post-occlusion. LY377770 may therefore be useful as a neuroprotectant in man. PMID- 10854903 TI - The effect of the glycine/NMDA receptor antagonist, (+)-HA966, on morphine dependence in neuropathic rats. AB - We have previously shown that rats with a painful peripheral neuropathy develop dependence without tolerance after repetitive doses [3mg/kg subcutaneously (s.c.)] of morphine. After injections of a higher dose (10mg/kg s.c.) the animals develop tolerance that can be prevented by the glycine/N-methyl-D-aspartate (NMDA) receptor antagonist, (+)-HA966. This study examined whether (1) dependence develops also after repetitive doses of 10mg/kg of morphine and, if so, (2) whether (+)-HA966 prevents the development of dependence after both the low and the higher morphine pretreatment doses. A 4day pretreatment regimen (post operative days 12-16) with two daily s.c. injections of saline+saline, saline+morphine (3 or 10mg/kg), (+)-HA966 (2.5 or 5mg/kg)+morphine or (+)-HA966 (5mg/kg)+saline was used, and withdrawal was precipitated by an injection of naloxone [2mg/kg intravenously (i.v.)] at 17h after the last pretreatment injection. Three signs of withdrawal (exploring, writhing, ptosis) appeared after pretreatment with both doses of morphine alone, while other signs (teeth chattering, pilo-erection) developed only after injections at the 3mg/kg dose. One sign (penile grooming/erection) appeared only after the higher morphine dose. Pretreatment with the combination of (+)-HA966 and morphine at 3mg/kg prevented the development of all withdrawal signs. By contrast, except for exploring, (+) HA966 did not modify the incidence of the withdrawal signs observed after pretreatment with doses of 10mg/kg of morphine. The results suggest that prevention of the development of morphine dependence by glycine/NMDA receptor antagonism depends on the degree of morphine dependence. PMID- 10854904 TI - Verapamil prevents withdrawal excitation of oxytocin neurones in morphine dependent rats. AB - We investigated whether the full expression of morphine withdrawal excitation by supraoptic nucleus (SON) oxytocin neurones is a property of the neurones themselves or a partial function of their afferent inputs, by interrupting synaptic input activity via central administration of the L-type Ca(2+) channel blocker verapamil. In morphine-dependent rats, withdrawal-induced release of oxytocin from the posterior pituitary was suppressed by prior administration of intracerebroventricular (i.c.v.) verapamil (160 microg), as was release of oxytocin within the SON measured by microdialysis. During morphine withdrawal the increased electrical activity of SON neurones was also reduced both by i.c.v. verapamil and microdialysis application of verapamil or nifedipine into the SON. Oxytocin secretion evoked by electrical stimulation of the pituitary stalk was unaffected by i.c.v. verapamil suggesting a central site of action. To determine whether the inhibitory actions of verapamil were specific to morphine withdrawal, we also investigated the effects of verapamil on other oxytocin-secreting stimuli. I.C.V. verapamil given to morphine-naive rats abolished pituitary oxytocin release in response to activation of brainstem or rostral excitatory inputs by cholecystokinin (20 microg kg(-1), i.v.) and 1.5 M saline (4 ml kg(-1), i.p.) respectively, whilst in lactating rats, i.c.v. verapamil reduced suckling induced release of oxytocin within the SON. These results suggest that verapamil has a central site of action on stimulated oxytocin release (including an action within the SON) and that both pre and post-synaptic L-type Ca(2+) channels are required for the full expression of morphine withdrawal in SON oxytocin neurones. PMID- 10854905 TI - The ability of WAY100,635 to potentiate the neurochemical and functional actions of fluoxetine is enhanced by co-administration of SB224,289, but not BRL15572. AB - The present study employed a combined neurochemical and behavioural approach to address the question of whether blockade of (presynaptic) 5-HT(1B) or 5-HT(1D) receptors enhances the facilitatory influence of 5-HT(1A) autoreceptor antagonism upon the actions of selective serotonin re-uptake inhibitors (SSRI). In the presence of the selective 5-HT(1A) antagonist, WAY100,635, the fluoxetine-induced increase in dialysate levels of 5-HT in the frontal cortex (FCX) of freely-moving rats was significantly potentiated. The selective 5-HT(1B) antagonist, SB224,289, likewise potentiated the increase in 5-HT levels evoked by fluoxetine. Further, administered together, WAY100,635 and SB224,289, at least additively, potentiated the influence of fluoxetine upon 5-HT levels. This effect was selective inasmuch as, either alone or together, WAY100,635 and SB224,289 did not modify the influence of fluoxetine upon FCX levels of dopamine (DA) or noradrenaline (NA) quantified in the same dialysis samples. Co-administration of SB224,289 also enhanced the ability of WAY100,635 to potentiate the induction of head-twitches (HTW) by fluoxetine. This response reflects activation of 5-HT(2A) sites in FCX and was abolished by the selective 5-HT(2A) antagonist, MDL100,907. In contrast to SB224,289, the 5-HT(1D) antagonist, BRL15572, failed to enhance the facilitatory influence of WAY100,635 upon the neurochemical or behavioural actions of fluoxetine. In conclusion, co-joint blockade of 5-HT(1B) - but not 5 HT(1D) - with 5-HT(1A) autoreceptors markedly potentiates the neurochemical and functional actions of the SSRI, fluoxetine. PMID- 10854907 TI - Retinoic acid negatively regulates neuropeptide Y expression in human neuroblastoma cells. AB - Retinoids are involved in the regulation of development and differentiation in many tissues, including the nervous system, where they have been associated with some neurotransmitter systems. In the present study, we evaluated the effects of all-trans retinoic acid (RA) on the biosynthesis and secretion of neuropeptide Y (NPY), a widely expressed neuroregulatory peptide. The SH-SY5Y human neuroblastoma cell line has been used as the in vitro model system. Treatment with 10 microM RA induced a marked decrease in NPY gene expression after as little as 3-6 h of incubation and resulted in its almost complete suppression at 12-24 h and after a 6-day differentiating treatment. The NPY content in cell extracts and the NPY secreted and accumulated in the culture medium were also reduced by exposure to 10 microM RA at 12 and 24 h and at 6 days. Moreover, RA treatment for 6 days, but not for 24 h, resulted in a marked stimulation of proNPY processing to mature NPY. The presence of negative retinoic acid-response elements in the human NPY promoter (up to -1078 bp) was excluded by a computer search. When SH-SY5Y cells were treated simultaneously with 20 nM TPA and 10 microM RA for 24 h, the marked stimulatory effect of TPA alone was completely suppressed. These observations suggest that the expression of NPY in SH-SY5Y human neuroblastoma cells is negatively regulated by RA at the level of gene expression, probably by mechanisms involving the interaction of activated RARs with transcription factors (such as AP-1). PMID- 10854906 TI - Effects of sigma(1) receptor agonist SA4503 and neuroactive steroids on performance in a radial arm maze task in rats. AB - This study examined the effects of sigma(1) receptor agonist SA4503 and neuroactive steroids dehydroepiandrosterone sulfate (DHEAS), pregnenolone sulfate (PREGS) and progesterone (PROG) on spatial working and reference memory in a radial arm maze task in rats. The insertion of a 6-min delay between the 2nd and 3rd choices caused a specific decline in working memory, but had no effect on reference memory. This decline in working memory was improved by SA4503, but not by DHEAS, PREGS or PROG. A non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine significantly impaired both working and reference memory in the presence or absence of a delay. The dizocilpine-induced impairments in the presence of a 6-min delay were ameliorated by SA4503, DHEAS and PREGS, whereas PROG had no effect. The beneficial effects of SA4503, DHEAS and PREGS were antagonized by treatment with sigma(1) receptor antagonist N, N-dipropyl-2-(4 methoxy-3-(2-phenylethoxy)phenyl)-ethylamine hydrochloride (NE-100). Furthermore, PROG attenuated the ameliorating effects of SA4503, DHEAS and PREGS on dizocilpine-induced memory deficits. These results suggest that sigma(1) receptors play a significant role in short-term working memory. Furthermore, it is suggested that DHEAS and PREGS ameliorate dizocilpine-induced memory impairments by acting as sigma(1) receptor agonists, while PROG antagonizes their effects by acting as a sigma(1) receptor antagonist. PMID- 10854908 TI - Drugs of abuse modulate the phosphorylation of ARPP-21, a cyclic AMP-regulated phosphoprotein enriched in the basal ganglia. AB - ARPP-21 is a cyclic AMP-regulated phosphoprotein of M(r) 21 kDa that is enriched in the cell bodies and terminals of medium-sized spiny neurons in the basal ganglia. Using a new phosphorylation state-specific antibody selective for the detection of ARPP-21 phosphorylated on Ser(55), we have demonstrated that activation of dopamine D1 receptors increased the level of ARPP-21 phosphorylation in mouse striatal slices. Conversely, activation of D2 receptors caused a large decrease in ARPP-21 phosphorylation. Treatment of mice with either methamphetamine or cocaine resulted in increased ARPP-21 phosphorylation in vivo. Studies using specific inhibitors of protein phosphatases and experiments in mice bearing a targeted deletion of the gene for DARPP-32, a dopamine-activated inhibitor of protein phosphatase-1, indicated that protein phosphatase-2A is primarily responsible for dephosphorylation of ARPP-21 in mouse striatum. These results demonstrate that phosphorylation and dephosphorylation of ARPP-21 are tightly regulated in the striatum. We speculate that ARPP-21 might mediate some of the physiologic effects of dopamine and certain drugs of abuse in the basal ganglia. PMID- 10854909 TI - The dopamine antagonist sch 23390 reverses dizocilpine-induced blockade of cocaine sensitization. AB - The present work examined the effects of pre-treatment with Sch 23390, a selective D(1) receptor antagonist, on the dizocilpine-induced blockade of sensitization to the locomotor-stimulating effect of cocaine. Rats were given either cocaine [15mgkg(-1)day(-1), intraperitoneally (i.p.)] from day 1 to day 5 (cocaine-experienced rats) or vehicle (cocaine-naive rats). From day 6 to day 15, animals remained drug-free in their home cages. On day 16 rats received a challenge injection of cocaine (15mgkg(-1)) or vehicle, and were tested for sensitization to the locomotor-stimulating effect of cocaine. In cocaine-naive rats the acute effect of cocaine was a 1.5 times increase in locomotor activity. In cocaine-experienced rats, the acute effects of cocaine were considerably more pronounced than in cocaine-naive rats; the stimulating effect of cocaine in these animals was a doubling in locomotor activity. In cocaine-naive rats, pre treatment with dizocilpine (100microgkg(-1)), Sch 23390 (100microgkg(-1)) or a combination of the two drugs from day 1 to day 5 changed neither spontaneous locomotor activity nor cocaine stimulant activity. By contrast, cocaine experienced animals that had been given 100microgkg(-1) dizocilpine from day 1 to day 5 failed to show the increase in locomotor activity when challenged with cocaine on day 16. Pre-treatment with Sch 23390 (100microgkg(-1)day(-1), i.p.) from day 1 to day 5 was found to prevent completely the cocaine anti sensitization properties of 100microgkg(-1) dizocilpine, but failed to prevent cocaine sensitization. On the other hand, horizontal activity in cocaine experienced rats that had been given dizocilpine (100microgkg(-1)) 15min before cocaine challenge on day 16 was higher than in corresponding controls. It is concluded that prevention of cocaine sensitization by dizocilpine may be related to the events set into motion by the NMDA antagonist at the level of dopaminergic transmission involving D(1) receptors. PMID- 10854911 TI - Differential sensitivity to hydroxyl radicals of pre- and postjunctional neurovascular transmission in the isolated canine mesenteric vein. AB - In some pathophysiological conditions, the first target of reactive oxygen intermediates is the vascular system. Superoxide anions, when generated in the vascular circulation, may then escape into the extracellular space via an anion channel and, following dismutation to hydrogen peroxide (H(2)O(2)), form hydroxyl radicals (HO(*)). In an attempt to understand the role of HO(*) in the regulation of transmission at the sympathetic neurovascular junction, the effect of HO(*) at nerve terminals was examined by measuring the amount of noradrenaline (NA) released from isolated, spirally cut, superfused canine mesenteric vein during basal and electrical stimulation (ES; 5Hz, 2ms, 9V); tension development evoked by ES was also recorded simultaneously. HO(*) was generated from Fenton's reagent (1. 5x10(-4)M H(2)O(2) plus 10(-4)M FeSO(4)); generation of HO(*) from H(2)O(2)/FeSO(4) in the superfusate was monitored by electron spin resonance spectroscopy using the spin-trap 5, 5-dimethyl-1-pyrroline-N-oxide throughout the experimental time course. Exposure to HO(*) of the helical strips produced an irreversible decrease in tension development evoked by ES with no effect on NA release, suggesting that the observed effect is elicited postjunctionally. The susceptibility of the processes of NA-mediated contraction to HO(*) may differ greatly from that of the NA release mechanism at the prejunctional site. Exposure of the strip preparation to HO(*) leads to a substantial stimulation of basal release of NA without affecting ES-evoked NA release, possibly due to enhanced non-exocytotic Ca(2+)-independent release elicited by HO(*). A direct demonstration of this concept was obtained by showing a significant increase in the basal response of NA release in Ca(2+)-free solution. The major conclusion of the present study is that HO(*) can damage NA-mediated contraction of the vascular preparations at the postjunctional site, and may selectively induce a non-exocytotic release of NA from the prejunctional site of sympathetic neurotransmission. PMID- 10854910 TI - The role of cytokines and prostaglandin-E(2) in thymulin induced hyperalgesia. AB - We have recently reported that intraperitoneal (i.p.) injection of thymulin at low doses (50 ng) resulted in thermal and mechanical hyperalgesia and upregulation of the level of interleukin-1beta in the liver. In this study, we demonstrate that such injections of thymulin result in a significant elevation in the levels of TNF-alpha (P<0.01), NGF (P<0.01) and PGE(2) (P<0.01) in the liver of the treated rats, in addition to the increase in the levels of IL-1beta. Pretreatment with specific antagonists to each of these factors (polyclonal anti TNF-alpha, anti-NGF antiserum and IL-1 receptor antagonist) did not result in the abolition of the hyperalgesia as assessed by the paw pressure, hot plate, paw immersion and tail flick tests. However, pretreatment with a combination of the above antagonist and antisera almost completely prevented thymulin-induced hyperalgesia. The cyclooxygenase inhibitor, meloxicam, reversed in a dose dependent manner (0.2, 0.4 and 2 mg/kg) thymulin effects as assessed by the different pain tests. It also abolished the thymulin-induced increase in the level of cytokines and NGF in the liver. Our results indicate that PGE(2) could be the key mediator of the hyperalgesic action of thymulin and the observed upregulation of proinflammatory cytokines and NGF. PMID- 10854912 TI - Chloramphenicol induces apoptosis in the developing brain. AB - Programmed cell death was studied in the superior colliculus of the developing rat brain following injections of chloramphenicol. Neonatal rats were either subject to unilateral eye removal or left untouched. Following a 3-h post operative survival, the animals were perfused with fixatives and frozen sections of their brains were examined for apoptosis after either neutral-red staining, in situ nick-end labeling of fragmented DNA, or immunocytochemistry to activated caspase-3. Chloramphenicol induced apoptosis in control brains and potentiated cell death in deafferented superior colliculi. The results show that CMP has a general pro-apoptotic effect in the developing brain. PMID- 10854913 TI - The evolution of neural coding ideas in the chemical senses. AB - A brief review of the evolution of hypotheses about neural coding in the chemical senses provides some perspective on the current status of these fields, and implications for further development. PMID- 10854914 TI - Coding in taste receptor cells. The early years of intracellular recordings. AB - In the gustatory system, the mechanism of coding-the process of relaying the identity and intensity of the stimulus to the central nervous system-begins with the taste receptor cells. Ironically, although these are the first cells of the gustatory system to contact stimuli, they were the last from which neurophysiological recordings were obtained. How taste receptor cells decipher stimulus identity remains the subject of active research; its origins began with a series of intracellular studies. Prior to the first intracellular recording, it was unknown if taste receptor cells would be specialists, responding to only a single class of taste stimulus, or generalists, responding to multiple stimuli. The first reports established several major aspects of these cells' physiology. Taste receptor cells have varying response profiles to basic stimuli; they have obvious conductance changes during stimulation; they have low resting potentials. It became evident that multiple transduction schemes must underlie these responses, although the identity of these transduction schemes remained elusive. Additionally, these early recordings missed a major phenomenon-the presence of regenerative electrical events (i.e., the action potential) was not observed due to the low input resistance that accompanied this technique. Although intracellular recordings are essentially no longer used to study taste receptor cells, replaced by the superior method of patch-clamp recording, these early articles provided key insights into the then unknown electrical responses of taste receptor cells. PMID- 10854915 TI - Taste cell function. Structural and biochemical implications. AB - Maintenance of constant relations between receptor cell types and branching from a single gustatory nerve fiber during normal cell turnover and regeneration requires cell-cell recognition likely mediated by timed expression of molecules at surfaces of taste bud cells, nerve endings, and in extracellular matrix. These processes assure stability of gustatory quality representation during intragemmal remodeling. Coincidentally, features of gemmal cell lifespan, including elongation, differentiation, and migration prior to apoptosis, must also be orchestrated by molecular signals. This article reviews the potential roles played by a variety of molecular markers for some relevant classes of proteins, peptides, and enzymes, which were presumed to be important for carrying out these gustatory cellular functions. PMID- 10854917 TI - Neuron types, receptors, behavior, and taste quality. AB - Neurophysiological studies on chorda tympani (CT) single fibers and behavioral studies on generalization of learned aversions in hamsters (Mesocricetus auratus) are reviewed. The work on hamsters is compared to work on other species, including the laboratory rat and several primate species, including humans. This body of data demonstrates associations between response profiles of physiologically defined specialist CT neurons and behavioral stimulus generalizations on one hand, and characteristics of putative taste receptors, on the other. Response profiles of generalist CT neurons are similarly associated with receptor characteristics, but are not associated with specific behavioral discriminations. The associations of peripheral nerve data with both receptor and behavior strongly suggest specific codes for "sucrose-like" and "NaCl-like" taste qualities. Definitive conclusions regarding "patterns" or "labeled lines" requires an understanding of mechanisms of central neural processing of the several specialist and generalist taste-afferent inputs. PMID- 10854916 TI - Gustatory neuron types in the periphery: a functional perspective. AB - Robert P. Erickson's research and writings formed the intellectual backdrop and guiding force for much of the major research on sensory coding in taste. As articulated best by Erickson, consideration focused on the relative merits of labeled-line and across-fiber pattern theory. The present article focuses primarily on a review of the electrophysiological and behavioral studies on salt taste and salt taste-mediated behavior in rodents. The evidence clearly shows that the peripheral gustatory system consists of a few neuron types/groups with well-defined physiological response characteristics. Electrophysiological studies of the chorda tympani nerve define a physiological group of narrowly tuned neurons selectively responsive to NaCl stimuli. It appears that this is a sodium sensing module that functions primarily in the detection, recognition, and ingestion of NaCl. PMID- 10854918 TI - Issues of gustatory neural coding: where they stand today. AB - The basic issues of gustatory neural coding are revisited. Questions addressed and conclusions drawn are: (1) what is the physical dimension across which gustatory neurons are sensitive, and upon which taste perceptions are based? The dimension that unites the various taste qualities is not physical, but physiological: a dimension of well-being, bounded by toxins at one extreme and nutrients at the other. (2) How broadly tuned are taste cells across the dimension? There are instances of specificity, but most mammalian taste cells respond to a range of qualities. (3) Are there basic taste qualities? Sweet, salty, sour, and bitter are widely accepted as basic tastes. Umami and starch tastes are considered basic by some. (4) Is taste topographically organized? There is some degree of physical separation among neurons most responsive to different taste qualities, but this does not appear to be sufficient precision to act as a meaningful coding mechanism. (5) Are there gustatory neuron types? Neurons, separated into categories according to their response profiles, respond as members of their category to the challenges of conditioned aversions and preferences, sodium deprivation, hyperglycemia, and receptor blockade, while cells from other categories react differently. This indicates the existence of functionally distinct types of taste cells. (6) Is the quality signal coded within the activity of the single most appropriate category of neurons, or is it carried by the pattern of response across neuronal categories? Both the breadth of responsiveness and the logical ambiguity of the signal in any one category of neurons argue that the taste message is carried by a pattern of activity across gustatory neuron types. PMID- 10854919 TI - Neuronal cell types and taste quality coding. AB - Over the past 25 years, there have been two opposing views of how taste information is represented in the activity of gustatory neurons. One view, the across-fiber pattern (AFP) theory, postulates that taste quality is represented by the pattern of activity across the afferent population. Stimuli with similar tastes produce similar patterns of activity. The other view is that activity in a few distinct neuron types codes taste quality in a "labeled-line" fashion. Neurons responding best to sucrose, for example, would represent "sweetness," and those responding best to NaCl would code "saltiness." Some of these neuron types appear to have a biological significance, such as the NaCl-best cells, which receive input about sodium stimuli exclusively from an amiloride-sensitive epithelial ion channel. However, the relatively broad tuning of these neurons makes it unlikely that they are capable of unambiguously coding information about taste quality. Rather, these neuron types play a critical role in establishing unique AFPs that distinguish among taste stimuli. The relative activity across these cell types represent taste quality, much like the patterns of activity across broadly tuned photoreceptors code information about stimulus wavelength. PMID- 10854920 TI - The neural code for taste in the brain stem: response profiles. AB - In the study of the neural code for taste, two theories have dominated the literature: the across neuron pattern (ANP), and the labeled line theories. Both of these theories are based on the observations that taste cells are multisensitive across a variety of different taste stimuli. Given a fixed array of taste stimuli, a cell's particular set of sensitivities defines its response profile. The characteristics of response profiles are the basis of both major theories of coding. In reviewing the literature, it is apparent that response profiles are an expression of a complex interplay of excitatory and inhibitory inputs that derive from cells with a wide variety of sensitivity patterns. These observations suggest that, in the absence of inhibition, taste cells might be potentially responsive to all taste stimuli. Several studies also suggest that response profiles can be influenced by the taste context, defined as the taste stimulus presented just before or simultaneously with another, under which they are recorded. A theory, called dynamic coding, was proposed to account for context dependency of taste response profiles. In this theory, those cells that are unaffected by taste context would provide the signal, i.e., the information containing portion of the ANP, and those cells whose responses are context dependent would provide noise, i.e., less stimulus specific information. When singular taste stimuli are presented, noise cells would provide amplification of the signal, and when complex mixtures are presented, the responses of the noise cells would be suppressed (depending on the particular combination of tastants), and the ratio of signal to noise would be enhanced. PMID- 10854921 TI - The across-fiber pattern theory and fuzzy logic: a matter of taste. AB - This article discusses a Fuzzy Logic (FL)-based model of neural coding and integration, proposed to be a formal extension of the Across-Fiber Pattern (AFP) theory. FL integration is conceptually similar to Bayesian reasoning, thus providing close-to-optimal decisions, and is also robust in that it does not require complete information. As a formal extension of AFP theory, the FL model describes sensory integration given multiple sources of information. When applied to gustation, the FL model is suggested to describe integration of information at the level of real-time pattern of single neural responses, population coding, and taste perception, as well as to provide a suitable description of taste mixtures. PMID- 10854922 TI - Artificial neural networks estimate the contribution of taste neurons to coding. AB - Taste qualities are believed to be coded in the activity of populations of taste neurons. However, it is not clear whether all neurons are equally responsible for coding. To clarify the point the relative contribution of each taste neuron to coding was assessed by constructing simple three-layer neural networks with input neurons that represent cortical taste neurons of the rat. The networks were trained by the back-propagation learning algorithm to classify the neural response patterns to the basic taste stimuli (sucrose, HCl, quinine hydrochloride, and NaCl). The networks had four output neurons representing the basic taste qualities, the values of which provide a measure for similarity of test stimuli to the basic taste stimuli. We estimated relative contributions of input neurons to the taste discrimination of the network by examining their significance S(j), which is defined as the sum of the absolute values of the connection weights from the jth input neuron to the hidden layer. When the input neurons with a smaller S(j) (e.g., 15 out of 39 input units) were "pruned" from the trained network, the ability of the network to discriminate the basic taste qualities was not greatly affected. On the other hand, the taste discrimination of the network progressively deteriorated much more rapidly with pruning of input neurons with a larger S(j). These results suggest that cortical taste neurons differentially contribute to the coding of taste qualities. Input neurons with a larger S(j) tended to be with a larger variation of neural discharge rates to the basic taste stimuli. The variation of neural discharges may be important in the coding of taste qualities. PMID- 10854923 TI - Sensory coding, decoding, and representations. Unnecessary and troublesome constructs? AB - Reception of certain environmental energy patterns can allow organisms to successfully respond to present or future environmental conditions. Sensory systems are the means by which this reception occurs. The bioelectric components of this process are typically characterized as "sensory coding." "Coding" logically requires subsequent "decoding," and implies that the decoder has a special, commanding role. In contrast, the term "transformation," which is sometimes applied to the prebioelectric components of sensory systems, can connote a series of changes in the medium, form, or content of sensory processes. Neither coding nor decoding are implied, no notions of "representation" or "reconstruction" of the world are introduced, and distributed, parallel processing is a natural corollary of transformations. It is proposed that the problematic construct of sensory coding, with its concomitant decoding, be replaced with the more neutral and physical concept of transformations. Elimination of the notions of representation or reconstruction of the world in some special nervous system locus is also suggested. PMID- 10854924 TI - Taste quality and intensity: lessons from the Morrison technique. AB - The nontoxic and nonshock Morrison operant technique was used to evaluate taste quality in rat and marmoset: response to a tastant test solution in pursuit of a pellet reward was dependent on making a choice between two bars that had been linked in discrimination training to qualitatively different stimulus pairs (NaCl versus either HCl, QHCl, or NH(4)Cl). The percentage distribution of bar-press responses to test stimuli showed: (1) stability of quality across 0.069-0.3 M NaCl, 0.003-0.1 M HCl, and 0.0001-0.003 M QHCl; (2) for LiCl, a quality change consistent with human reports of a "sour" to "salty" shift; (3) a suggestion that the "salty-like" quality of NH(4)Cl and NaCl are not perceptually equivalent; (4) NaNO(3) shares NaCl-like, QHCl-like, and NH(4)Cl-like components; (5) CaCl(2), KCl, and MgCl(2) share QHCl-like and NH(4)Cl-like components; and (6) responses to HCl and QHCl were not hedonically driven in the rat. Comparison of rank order correlations of single-unit firing rates to the distribution of bar-press responses for the same test stimulus concentration revealed that (7) no single level of the gustatory pathway exclusively accounts for the operant response distribution pattern to either simple or complex tastants, and (8) discriminations between tastants, one of which may be qualitatively complex, are not necessarily mediated only at levels proximal to the solitary nucleus. Thus, the Morrison discrimination technique effectively yields statements about gustatory quality without use of negative reinforcers and largely uninfluenced by tastant hedonics. PMID- 10854925 TI - Orosensory factors in the ingestion of corn oil/sucrose mixtures by the rat. AB - The experiments reported here were designed to study the orosensory factors contributing to the ingestion of sucrose/corn oil mixtures. When a flavor aversion was conditioned to the sucrose/corn oil mixture, the subsequent aversion to the mixture strongly generalized to the corn oil but very little to the sucrose. Rats conditioned with corn oil show a more profound aversion to the sucrose/corn oil mixture than rats conditioned with sucrose, indicating that the salient feature of the sucrose/corn oil mixture is the oil. Aversion to the sucrose/corn oil mixture does not generalize to a sucrose/mineral oil mixture, giving evidence that the textural aspects of the oil do not play a major role in its perception. This flavor aversion to the mixture is further illustrated in very short-term tests where postingestive factors are minimized, indicating a role for the gustatory system in the detection of the sucrose/corn oil mixture. Preliminary experiments are reported where conditioning tests were run with mixtures of sucrose and linoleic acid, one of the fatty acids that is possibly derived from a breakdown of the corn oil in the oral cavity by lingual lipase from von Ebner's Gland. PMID- 10854926 TI - Taste quality and neural coding: implications from psychophysics and neurophysiology. AB - Historically, taste research has often been guided by the concept that there are only four (or possibly five) basic taste qualities (sweet, sour, salty, and bitter, and possibly "umami"). All other tastes have been presumed to be combinations of these basic tastes. This psychophysical concept has been extended to electrophysiological data. That is, the neural code for each basic taste is hypothesized to be coded by a dedicated channel of neurons (the "Labeled-Line" theory); i.e., one group of neurons signals "salty" and another separate group signals "sweet." Numerous psychophysical and electrophysiological findings, however, cannot be accomodated by this quadripartite theory, which limits taste to four basic qualities and four basic neuron types. Rather, the data described in this article suggest that the range of taste is more extensive than four or five basic tastes, and that this breadth of taste quality results initially from the activation of a broad array of ion channels, receptors, and second messengers associated with taste cell membranes. These findings have implications for neural organization and provide support for the "Across-Fiber Pattern" theory in which the neural code for taste is represented by the pattern of activity across all of the neurons, i.e., neurons are not exclusively labeled for a particular sensation but cooperate with the others in the ensemble to encode taste quality. PMID- 10854927 TI - Genetic variation and inferences about perceived taste intensity in mice and men. AB - The study of genetic variation in taste produces parallels between mice and men. In mice, genetic variation across strains has been documented with psychophysical and anatomical measures as well as with recordings from whole nerves. In humans, the variation has been documented with psychophysical and anatomical measures. Whole-nerve recordings from animals and psychophysical ratings of perceived intensities from human subjects have a similar logical limitation: absolute comparisons across individuals require a standard stimulus that can be assumed equally intense to all. Comparisons across whole-nerve recordings are aided by single-fiber recordings. Comparisons across psychophysical ratings of perceived intensity have been aided by recent advances in methodology; these advances now reveal that the magnitude of genetic variation in human subjects is larger than previously suspected. In females, hormones further contribute to variation in taste. There is evidence that the ability to taste (particularly bitter) cycles with hormones in women of child-bearing age, rises to a maximum early in pregnancy and declines after menopause. Taste affects food preferences, which in turn affect dietary behavior and thus disease risks. Valid assessment of taste variation now permits measurement of the impact of taste variation on health. Advances in psychophysical methodology were essential to understanding genetic variation in taste. In turn, the association of perceived taste intensities with tongue anatomy now provides a new tool for psychophysics. The ability of a psychophysical scale to provide across-subject comparisons can be assessed through its ability to show the fungiform papillae density-taste association. PMID- 10854929 TI - Patterns in the brain. Neuronal population coding in the somatosensory system. AB - The aim of this article is to review some basic principles of neural coding, with an emphasis on mechanisms of stimulus representation in ensembles of neurons. The theory of "across-neuron response patterns" (ANRPs), first suggested by Thomas Young (1802) and fully developed by Robert Erickson (1963-2000), is summarized and applied to the problem of coding in primary afferent fibers and cortical neurons of the somatosensory system. The basic premise of the theory is that precise information about stimulus features cannot be encoded by single neurons, but is encoded by patterns of activity across populations of neurons. Different stimuli produce uniquely different patterns of ensemble activity (ANRPs) discrimination between two stimuli is based on the absolute difference in total amount of activity (neural mass difference) of the ANRPs for those stimuli. Review of the literature shows that ANRPs and related population codes can accurately represent and differentiate among various stimulus parameters that cannot be distinguished by single neurons alone. Finally, the behavior of neuronal ensembles can be used to account for the sensory-perceptual changes associated with plasticity of thalamocortical circuits following selective sensorimotor deprivation or experience. PMID- 10854928 TI - Different responses to repeated applications of zingerone in behavioral studies, recordings from intact and cultured TG neurons, and from VR1 receptors. AB - When applied repetitively to the cornea, capsaicin, the pungent compound in hot pepper, causes an initial eye-wiping response that diminishes upon repeated exposure (tachyphylaxis). This diminution, however, is not observed upon repetitive application of its pungent analogue, zingerone, to the cornea or tongue. In addition, compared with capsaicin, the lingual application of zingerone produces a gustatory response with a shorter latency and duration. Because both the tongue and the cornea are innervated by the trigeminal nerve, and because zingerone and capsaicin are structurally related, it is not evident why the responses to these compounds should give such different behavioral and psychophysical endpoints. We have addressed this issue by measuring the neural responses from rat trigeminal ganglion neurons (TG) to repeated applications of zingerone applied to the cornea, from cultured rat TG neurons, and from cloned capsaicin receptors (VR1) expressed in Xenopus oocytes and then comparing these effects to those evoked by capsaicin. Extracellular recordings from the trigeminal ganglion revealed that the responses to repeated corneal applications of 30 mM zingerone show desensitization. Cultured TG neurons, and oocytes expressing VR1 receptors, were also desensitized by repeated applications of zingerone. Electrophysiological recordings revealed that these two vanilloids could activate the same receptor (VR1), currents in the same neuron, and crossdesensitize. The more rapid onset and shorter duration responses seen with zingerone (compared with capsaicin) provides a rationalization for its more rapid onset and shorter duration gustatory response. We attribute the different behavioral responses to periodic applications of these two agonists to two competing effects: one leading to sensitization, and the other to tachyphylaxis. Which of these dominates depends on the concentration, exposure time, and interstimulus interval. Consequently, whether or not zingerone will exhibit tachyphylaxis depends critically on the experimental conditions. PMID- 10854930 TI - Learning from spiny lobsters about chemosensory coding of mixtures. AB - Studies of the peripheral olfactory system of the Caribbean spiny lobster Panulirus argus and related decapod crustaceans have helped us understand mechanisms of coding of mixtures, some of which are discussed in this review. Although the number of cells in the lobster's olfactory system is much lower than in vertebrate olfactory systems, it is a highly complex system. The receptor neurons (RNs) of this olfactory system are complex processors that cannot be categorized into discrete cell types, but rather have a diversity of response profiles. Each RN can have different types of receptor proteins, second messengers, and/or ion channels, which undoubtedly contributes to the functional diversity of these neurons and makes them complex peripheral integrators. The RNs probably encode information about the quality of mixtures as a distributed or population code, providing a basis for behavioral discrimination of natural food stimuli. Analysis of distributed codes for a series of blend ratios of binary mixtures reveals that the qualities of individual compounds are probably not lost when mixed. Such peripheral processing allows spiny lobsters to perceive complex odors as a set of elemental cues if the salience of the components is sufficiently high. PMID- 10854931 TI - Neural population coding and auditory temporal pattern analysis. AB - Over the 2 decades that have elapsed since Robert Erickson first published his pioneering work on across-fiber patterns in the gustatory system, the idea that information is represented by a population code has become almost universally accepted among neuroscientists. Although the concept of a population code is an implicit theoretical assumption underlying most of the work done in neuroscience today, the details of how population codes operate in specific systems remain unclear in many respects. This article reviews electrophysiological studies of the auditory system of echolocating bats that show that information about sound is initially represented across both space and time by relative amounts of activity in populations of excitatory and inhibitory neurons with different discharge patterns, different sensitivity functions, and different latencies. At the next level, each neuron in the auditory midbrain receives convergent input from a specific population of these lower brainstem neurons and acts as a "readout" of activity within this population. As a result, midbrain neurons become selectively tuned to stimulus features, for example, signal duration, to which neurons at lower levels respond indiscriminately. Intracellular recordings from auditory midbrain neurons show some of the mechanisms by which population input is processed. The known projection patterns of the midbrain "readout" neurons indicate that they, in turn, must become part of a new spatio-temporal population code that is transmitted to neurons at the thalamus, where additional forms of selectivity and patterns of output arise. PMID- 10854932 TI - Age pattern of mortality in eight breeds of insured dogs in Sweden. AB - The objective of this study was to use several methods to describe the age patterns for risk of death in selected breeds of dogs insured for life in a Swedish animal-insurance company in 1996. Data on eight breeds were analyzed for age at death (including euthanasia). If dogs left the insurance for reasons other than death, they were regarded as censored. Dogs were only insured up to 10 years of age. Four analytical approaches were used. First, descriptive statistics of age distributions (e.g. breed-specific median ages at death, breed- and age specific mortality risks) were computed. Second, age-specific estimates of survival were calculated using the formula: survival=(1-risk(age<1 year))(1 risk(age 1<2 year))... (1-risk(age 9<1 0 year)). Third, Cox regression (proportional-hazards model) was used to estimate survival and hazard functions. Finally, hierarchically coded Poisson regression was used to determine age specific cut-points in the risk of death. The hazards from Cox and the incidence density rates from the hierarchically coded models were transformed to estimates of risk: risk=1-exp?-(hazard)? or 1-exp?-(incidence-density rate)?. The breeds studied were Beagle, Bernese mountain dog, Boxer, Cavalier King Charles spaniel, Drever, German shepherd dog, Mongrel and Poodle, together representing over 50000 dogs each year. The yearly breed-specific mortality risk varied between 1.7% (Poodle) and 6.5% (Bernese mountain dog). In all breeds, the risk of death increased with age but the pattern varied by breed. The probability of survival at 5 years of age varied between 94% (Cavalier King Charles spaniel and Poodle) and 83% (Bernese mountain dog, Drever, German shepherd dog) and the survival at 10 years between 83% (Poodle) and 30% (Bernese mountain dog). The survival estimates from Cox and those derived using the combined-risk formula were similar. The cut-point risk estimates provided a simplified picture of when the risk of death changed significantly compared to previous age categories. As anticipated, breeds differed widely in survival up to 10 years of age and there were marked differences in age patterns of mortality. The implications of these findings should be considered in multivariable analyses, where the confounding effect of age is often controlled for using a single age variable common to several breeds. PMID- 10854933 TI - A cross-sectional study of paratuberculosis in 1155 Danish dairy cows. AB - In a cross-sectional study on milk samples from 1155 cows from 22 Danish dairy herds, selected risk factors for paratuberculosis were identified. The diagnostic procedure used was an indirect enzyme-linked immuno-sorbent assay (ELISA) to detect antibodies against Mycobacterium avium subsp. paratuberculosis. A sample was considered test-positive if it had a corrected optical density >/=0. 025 (test sensitivity 71.4% and test specificity 89.7%). Of the 1155 samples, 8.8% (102/1155) were test-positive, and 19 out of the 22 dairy herds had >/=1 test positive cows. The significant risk factors in a multiple logistic regression analysis were: Jersey versus large breeds, high parity versus low parity, the first month after calving versus other months of lactation, and a large herd size compared to a small herd size. The highest probability (37-38%) of a positive test was observed among older cows (parity >4) and tested within the first month after calving (irrespective of breed). The lowest probability (2%) of a positive test-result was observed among first parity, large-breed cows tested before calving or later than one month after. PMID- 10854934 TI - Simulated effects on dairy cattle health of extending the voluntary waiting period with recombinant bovine somatotropin. AB - We simulated the effect of extending the voluntary wait period by 100 days on disorder-frequency measures that were based on cow-years (from lactations completed during the 4-year simulation horizon), metric tons of milk yield, and lactational incidence risks. A dynamic stochastic discrete-event simulation model that focuses on clinical and subclinical intramammary infections (IMI), plus clinical metabolic (left-displaced abomasum, ketosis, milk fever) and reproductive (cystic ovarian disease, dystocia, retained placenta, twinning, uterine infection) disorders in dairy herds was used. Although the voluntary wait period was increased by 100 days (50 vs. 150), the predicted difference in simulated days to conception was only 89 days for the extended voluntary wait period group (which we attributed to higher fertility later in lactation). Herds that had a voluntary wait period of 150 days (compared to the control herds' voluntary wait period of 50 days) were predicted to have significantly lower rates of metabolic and reproductive disorders and clinical mastitis on both cow year and milk-yield bases. Simulated control herds, on average, produced 8539 kg of milk in an average lactation of 325 days and simulated herds with a 150-day voluntary wait period 10893 kg of milk in an average lactation of 409 days. There was a significantly lower predicted rate and risk of culling for reproductive failure in the extended voluntary wait period group. The predicted lactational incidence risks for subclinical IMI were 18% higher for the extended voluntary wait period group - but extending the voluntary wait period by 100 days was predicted not to increase the risk of any of the other 10 disorders. PMID- 10854935 TI - Seroepidemiology of Brucella abortus infection in bovids in Sri Lanka. AB - From 1992 to 1995, 0.15% (n=3916) of the bovids (cattle and buffalo) in Sri Lanka were sampled, using a multi-stage sampling procedure. Serum antibodies for Brucella abortus were detected using the indirect enzyme-linked immunosorbent assay (ELISA). The age, the agroecological zone and the management system practiced in the farms of the sampled bovids were studied as risk factors for seropositivity. The overall seroprevalence of brucellosis in cattle was 4.7% (n=3076) and 4.2% in buffaloes (n=840). Bovids that were over 3 years of age, from the dry zone (annual rainfall 20-35 in.), and reared under an extensive management system had higher odds of being seropositive. Bovids from the dry zone were at approximately six times higher odds of being seropositive even after controlling for the possible effects of age and management system. Approximately 75% of the seropositive males were from the dry zone. Most bovids (84%) from the dry zone in this study were reared under an extensive management system (free grazing) which allows unrestricted contact between animals. These factors may have contributed to the spread of brucellosis in bovids in the dry zone. This infection might be an important cause of abortion in bovids in Sri Lanka. However, there is also evidence of other causes for abortion, repeat breeding and retained placenta. PMID- 10854936 TI - Foaling-management practices associated with the occurrence of enterocolitis attributed to Clostridium perfringens infection in the equine neonate. AB - Enterocolitis associated with Clostridium perfringens (C. perfringens) infection in neonatal foals is often severe and has been associated with a high case mortality risk. We designed a premises-based survey to evaluate the associations of regional foaling practices, premises environmental management, periparturient foal and brood-mare management, and periparturient brood-mare ration with the occurrence of neonatal enterocolitis attributed to C. perfringens infection. Potential risk factors individually associated with enterocolitis were breed type, housing type at foaling and in the first three days of life, ground/floor surface type at foaling and in the first three days of life, brood-mare ration before and after foaling, and the presence of livestock other than horses on the premises in the past. From the multivariable-logistic regression models, six variables were significantly associated with an increased risk of the outcome of interest (p<0.05): foals of the stock horse type, housing in a stall or drylot in the first three days of life, other livestock present on the premises in the past, foal born on dirt, sand or gravel surface, and low amounts of grass hay and grain fed post-partum. Low grain amounts fed pre-partum represented a decreased risk of the outcome of interest. PMID- 10854937 TI - Adjuvant radiotherapy after breast conserving surgery for breast cancer. Pro. PMID- 10854938 TI - Adjuvant radiotherapy after breast conserving surgery for breast cancer. Contra. PMID- 10854939 TI - Adjuvant radiotherapy after breast conserving surgery for breast cancer. Arbiter. PMID- 10854940 TI - Future prospects for the prevention and cure of breast cancer. PMID- 10854941 TI - False-positive findings in mammography screening induces short-term distress - breast cancer-specific concern prevails longer. AB - The aim of this study was to examine psychological distress in a mammography screening process as a consequence of screening after adjusting for background, personality and prescreening distress. Subjects, aged 50 years, were invitees at their first screening. There were three groups; normal findings (n=1407), false positive findings (n=492) and referents from outside the screening programme (n=1718, age 48-49 years). Distress was measured as illness worry, anxiety, depression, cancer beliefs and early detection behaviour. Measurements were one month before screening invitation with follow-ups at 2 and 12 months postscreening. At 2 months, there was a moderate multivariate effect of group on distress; and intrusive thinking and worry about breast cancer, in particular, were most frequent amongst the false positives. Intrusive thinking still prevailed at 12 months, in addition to a higher perceived breast cancer risk and susceptibility. Distress related to screening and false-positive findings seems to be moderate, but prevailing cancer-specific concerns call for improvements in screening programmes. PMID- 10854942 TI - Altered expression of E-cadherin in breast cancer. patterns, mechanisms and clinical significance. AB - Reduced cell adhesion brought about by altered surface expression of E-cadherin has been implicated in invasive and metastatic malignant growth. We investigated the patterns of immunohistochemical E-cadherin expression in 120 breast carcinomas. Furthermore, we analysed DNA from the same samples for loss of heterozygosity (LOH) using three separate microsatellite markers on chromosome 16q22.1. Finally, the clinical outcome was ascertained for 108 patients. 19% (18/97) of infiltrating ductal carcinomas showed complete loss of E-cadherin expression compared with 64% (9/14) of infiltrating lobular carcinomas. LOH was detected in 46% (24/52) of infiltrating ductal carcinomas and 89% (8/9) of infiltrating lobular carcinomas. In the infiltrating lobular carcinomas, LOH was associated with complete loss of cell membrane expression of E-cadherin, although a cytoplasmic expression pattern was evident. In contrast, this association was not seen in the infiltrating ductal carcinomas. In a multivariate analysis, loss of E-cadherin expression was shown to be a significant independent risk factor for a poorer disease-free survival (P=0.019), in particular in the node-negative subset of patients (P=0.029). Significance was also approached for breast cancer corrected survival (P=0.056). We conclude that different mechanisms are involved in the altered E-cadherin expression seen in different subtypes of breast carcinomas. Furthermore, we implicate loss of E-cadherin, regardless of the genetic causes, as an independent prognostic marker for disease recurrence, especially in node-negative breast cancer patients, irrespective of the histological type. PMID- 10854943 TI - A phase II study of docetaxel and vinorelbine combination chemotherapy in patients with advanced non-small cell lung cancer. AB - A phase II study was conducted to determine the efficacy and the safety of docetaxel combined with vinorelbine as first-line chemotherapy in patients with metastatic or unresectable non-small cell lung cancer (NSCLC). 39 patients, median age 54 years (range: 35-69), with stage IIIB (5 patients; 13%) or IV (34 patients; 87%) NSCLC were treated with 75 mg/m(2) docetaxel given intravenously (i. v.) over 1 h on day 1 and with 20 mg/m(2) vinorelbine given i.v. over 15 to 30 min on days 1 and 5. Cycles were repeated every 3 weeks. 9 of the 39 patients had a partial response (overall response rate 23.1%, 95% confidence interval (CI): 11.1-39.3%) with a median duration of response of 20 weeks (95% CI; 17-30). The median survival was 40 weeks (95% CI: 21-49 weeks) with a 1-year survival rate of 31% in the intent-to-treat population. Neutropenia grade IV occurred in 33 patients (92%). 16 patients (41%) experienced febrile neutropenia with a concomitant stomatitis in 9 patients (23%). One patient died due to febrile neutropenia associated with a grade 4 stomatitis and 1 patient due to a septicaemia concomitant with a grade 4 neutropenia. Although the combination of docetaxel and vinorelbine is feasible, the efficacy does not seem to be improved compared with single-agent docetaxel or vinorelbine and the rate of febrile neutropenia is unacceptable in this population with incurable disease. Therefore, different doses and/or schedules are to be explored. PMID- 10854944 TI - Genetic instability in intestinal metaplasia is a frequent event leading to well differentiated early adenocarcinoma of the stomach. AB - To understand the development of well-differentiated adenocarcinoma in the stomach, we examined genetic instability in 31 patients with stage Ia gastric cancer. Triplets of tissue specimens (normal/metaplasia/tumour) from 33 lesions were examined for microsatellite instability (MSI) and loss of heterozygosity (LOH), using nine microsatellite loci. Frameshift mutations in the transforming growth factor beta receptor type II (TGF-betaRII) (A)(10), Bcl-2-associated X protein (BAX) (G)(8), hMSH3 (A)(8) and hMSH6 (C)(8) genes were also studied. In this study, a high incidence of MSI (MSI-H) was defined as samples containing 30% or more MSI positive loci, and a low incidence of MSI (MSI-L) as samples which had less than 30% MSI. MSI-L was observed in 19 cancerous lesions (58%), and MSI H in three (9%). Eleven intestinal metaplasia lesions (33%) showed MSI-L, but no metaplasia lesions exhibited MSI-H. Frameshift mutation was observed in only one cancerous lesion (3%) at the (A)(10) tract of TGF-betaRII. In contrast, LOH was observed in 24 cancerous lesions (73%), and in 15 (45%) of intestinal metaplasia lesions. Intriguingly, these alterations tend to be coincident between metaplasia and cancerous lesions in the same sets of specimens, and there was no case that showed alterations in metaplasia, but not in cancerous lesions. These findings suggest that metaplasia and well-differentiated adenocarcinoma in the stomach may have the same molecular backgrounds, and that these two lesions may be chronologically connected. PMID- 10854945 TI - Immunohistochemical analyses of sporadic and familial (185delAG carriers) ovarian cancer in Israel. AB - A single germ line mutation in BRCA1, (185delAG) is detected in a substantial portion of Jewish Israeli patients with ovarian cancer. Whether disease phenotypes differ in BRCA1 mutation carriers and sporadic cases is presently a subject for debate. To gain insight into this issue, we analysed tumours from 65 Jewish women with ovarian cancer, 29 (45%) were 185delAG BRCA1 mutation carriers, and 36 (55%) were non-carriers of any of the predominant Jewish mutations in BRCA1 or BRCA2 (sporadic). In 19/29 mutation carriers (66%) diagnosis was made prior to age 60 years, compared with 14/36 (39%) of the non-carriers (P=0.03; Yates corrected P=0.06). Low malignant potential ('borderline') tumours were detected less frequently among carriers (2/29; 7%) than non-carriers (9/36; 25%) (P=0.03; one tail P=0.05). Immunohistochemical analysis in invasive carcinoma (n=54) showed that 17/27 carriers (63%) and 18/27 non-carriers (67%) had positive nuclear staining with a p53 antibody. In 4/27 carriers (15%) and 3/25 non carriers (12%), 25% or more of the tumour cells stained positive for Ki-67, an insignificant difference. Results were not altered by including borderline tumours (n=11) in these analyses. We conclude that the rate of TP53 inactivation and proliferative index in ovarian cancer, are similar for 185delAG BRCA1 mutation carriers and sporadic cases. PMID- 10854946 TI - Quality control in multicentric clinical trials. An experience of the EORTC Gynecological Cancer Cooperative Group. AB - Data Quality is a central requirement of scientific research and external monitoring is essential in multicentric clinical trials (MCT). A quality control (QC) study was conducted in the main Institutions participating in EORTC-GCCG Protocol number 55863 - randomised phase III trial of vindesine, cisplatin, bleomycin and mitomycin-C (BEMP) versus cisplatin (P) in disseminated squamous cell carcinoma of the uterine cervix - in order to assess the impact of variations in data quality on the conclusions of the trial. The reliability of the different centres in following the protocol was investigated by a questionnaire covering drug prescription, local facilities and the procedure for preparation and administration of chemotherapy. The 'treatment protocol adherence' was evaluated by recalculation of the ideal protocol dose and its comparison with the actual delivered dosage at each cycle of chemotherapy. 'Data quality control' was assessed by comparison of data on case report forms (CRFs) with the corresponding items in the medical records. Eleven centres participating in the trial were visited by the same team of reviewers. Striking differences were noted in the chemotherapy administration procedures and between the type and quality of hospital files. Overall, there was an acceptable level of data quality and protocol compliance. Data accuracy was 81.8% (range: 65. 6-97%) of the 4424 items checked. Incorrect data were found in 7.0% (2.3-14.5%), data were missing on the form in 3.6% of cases (0-12%) and data was on the form but not in the file in 7.6% of cases (0. 7-17.5%). Causes of inaccuracy were analysed. Both problems in data management but also in a lack of clarity of the protocol and/or CRFs were to blame. Training and supervision of data managers, precision in writing protocols, standardisation of some aspects of CRFs and the use of a checklist for chemotherapy data and treatment toxicities would have avoided many of these errors. The need for QC in all collaborative groups performing MCT is emphasised. A literature review on QC in MCT dealing with chemotherapy is included. PMID- 10854947 TI - Fatigue in patients with prostate cancer receiving hormone therapy. AB - The aim of this study was to determine the prevalence, severity and correlates of fatigue in a convenience sample of outpatients with prostate cancer prior to and following 3-months treatment with first-line hormone therapy (cyproterone acetate and goserelin). 'Severe fatigue' in the patients (n=62) was defined as a score on the Fatigue Severity Scale (FSS) greater than the 95th percentile of a group of elderly volunteers without cancer. Subjects also completed other questionnaires about fatigue and about quality of life, anxiety/depression and personality. Subjects underwent a nutritional assessment, tests of voluntary muscle function and attention. The prevalence of 'severe fatigue' at baseline was 8/58 (14%). Median FSS scores increased significantly after 3 months treatment. On multivariate analysis psychological distress explained 28% of the variance in fatigue scores. Treatment was associated with a reduction in voluntary muscle function, loss of muscle bulk, a decline in virility and potency, an improvement in pain and a reduction in nausea/vomiting. Fatigue is an important but under recognised side-effect of hormone therapy. PMID- 10854948 TI - Familial breast cancer in southern Finland: how prevalent are breast cancer families and can we trust the family history reported by patients? AB - We evaluate the validity of the family history of breast cancer reported by the patient and the number of families and individuals at risk and potentially benefiting from surveillance. Family history of cancer was systematically screened in three different series of breast cancer patients. Breast cancer families were defined by the selection criterion of at least three first- or second-degree relatives with breast or ovarian cancer (including the proband) and their genealogy and cancer diagnoses were confirmed. Family history of breast or ovarian cancer was reported by approximately 30% of the patients and 7-9% were classified as breast cancer families. On average, there were 3.1 healthy female (age: 20-70 years) first degree relatives per family potentially at high risk. Index patients reported 87% of all confirmed diagnoses and the primary site was correct in 93-95% of the reported cases. The incompleteness and errors in accuracy should be considered in epidemiological studies and verification of the diagnoses is important when deciding clinical management. PMID- 10854949 TI - Altered cell cycle response of drug-resistant lung carcinoma cells to doxorubicin. AB - The effect of doxorubicin treatment on cell cycle parameters in asynchronous populations of multidrug-resistant human lung carcinoma cell lines was investigated. A sensitive (DLKP-SQ) and three resistant (DLKP-SQ A250 10p#7, DLKP A2B and DLKP-A5F) variants of a human lung carcinoma cell line DLKP were exposed to equitoxic concentrations of doxorubicin. The latter three were 8-fold, 30-fold and 300-fold resistant to doxorubicin, respectively. Irreversible G2/M arrest in sensitive (DLKP-SQ) cells was observed 24 h after initiation of doxorubicin treatment. In resistant variants, G2/M arrest occurred at 12-16 h with a subsequent bypass of the G2/M arrest to re-emerge and accumulate in G1. This transient G2/M arrest and subsequent progression into G1 indicated an inefficient checkpoint for monitoring DNA damage induced by doxorubicin treatment. Caffeine treatment could bypass the G2/M block in DLKP-SQ cells. Doxorubicin treatment did not alter cyclin B or cdc2 protein levels, the ability of cdc2 to form complexes with cyclin B or the levels of cyclin B bound to cdc2. The G2/M arrest seen in sensitive cells was associated with an increase in inhibitory phosphorylation of Tyr15 on cdc2. In contrast, tyrosine 15 phosphorylation did not change in resistant variants after drug treatment and a general increase in cdc2 kinase activity was seen. Cdc25C levels were not altered following drug treatment. PMID- 10854950 TI - Inhibition of farnesyltransferase with A-176120, a novel and potent farnesyl pyrophosphate analogue. AB - Farnesylation of Ras is required for its transforming activity in human cancer and the reaction is catalysed by the enzyme farnesyltransferase. Recently, we discovered a novel chemical series of potent farnesyl pyrophosphate (FPP) analogues which selectively inhibited farnesyltransferase. Our most potent compound to date in this series, A-176120, selectively inhibited farnesyltransferase activity (IC(50) 1.2+/-0.3 nM) over the closely related enzymes geranylgeranyltransferase I (GGTaseI) (IC(50) 423+/-1.8 nM), geranylgeranyltransferase II (GGTaseII) (IC(50) 3000 nM) and squalene synthase (SSase) (IC(50)>10000 nM). A-176120 inhibited ras processing in H-ras-transformed NIH3T3 cells and HCT116 K-ras-mutated cells (ED(50) 1.6 and 0.5 microM, respectively). The anti-angiogenic potential of A-176120 was demonstrated by a decrease in Ras processing, cell proliferation and capillary structure formation of human umbilical vein endothelial cells (HUVEC), and a decrease in the secretion of vascular endothelial growth factor (VEGF) from HCT116 cells. In vivo, A-176120 reduced H-ras NIH3T3 tumour growth and extended the lifespan of nude mice inoculated with H- or K-ras-transformed NIH3T3 cells. A-176120 also had an additive effect in combination with cyclophosphamide in nude mice inoculated with K-ras NIH3T3 transformed cells. Overall, our results demonstrate that A 176120 is a potent FPP mimetic with both antitumour and anti-angiogenic properties. PMID- 10854951 TI - Analysis of growth factor-dependent signalling in human epithelioid sarcoma cell lines. clues To the role of autocrine, juxtacrine and paracrine interactions in epithelioid sarcoma. AB - Human epithelioid sarcoma (ES) is an extremely aggressive soft tissue tumour of unknown histogenesis. Although growth factor-dependent signalling cascades significantly affect the biological behaviour of malignant tumours, little is known so far about their role in human ES. The present investigation, therefore, analyses the coexpression and function of different growth factors and their receptors in the human ES cell line GRU-1 and its clonal subpopulations (GRU-1A, GRU-1B and GRU-1C). As shown by Northern blot, flow cytometry, immunocytochemistry and MTT assay, all ES cell lines expressed transforming growth factor (TGF)-alpha and the epidermal growth factor receptor (EGF-R). Although no response to exogenous TGF-alpha was observed, antagonistic anti-EGF-R antibodies (at 20 microg/ml) induced significant (P<0.05) growth inhibition in all cell lines. All cell lines showed coexpression of platelet-derived growth factor (PDGF)-A and the corresponding receptors. Neutralisation of ES-derived PDGF by anti-hPDGF antibodies resulted in significant (P<0.05) growth inhibition of all clonal subpopulations. Although all cell lines expressed TGF-beta(1) as well as TGF-beta type I and type II receptors (TGF-BI-R and TGF-BII-R), growth inhibition (P<0.05) by exogenous TGF-beta(1) was achieved in the clonal subpopulations only and not in the parental cell line. No ES cell line expressed acidic fibroblast growth factor (FGF) but stimulation of FGF type 3 and type 4 receptors (FGF-3R and FGF-4R) by exogenous acidic FGF (aFGF) resulted in a marked (P<0.05) acceleration of proliferation in all cell lines. In conclusion, our investigation suggests an intricate network of autocrine, juxtacrine and paracrine signalling between ES tumour cells and adjacent non-neoplastic stromal cells. PMID- 10854952 TI - Effect of carbogen breathing on the pharmacodynamics of 5-fluorouracil in a murine colon carcinoma. AB - To determine whether carbogen breathing has an effect on 5-fluorouracil (5-FU) uptake, retention and metabolism in C38 murine colon tumours grown in C57Bl/6 mice, we used in vivo 19F nuclear magnetic resonance (NMR) spectroscopy. Eleven tumour-bearing mice were treated with 150 mg/kg of 5-FU given intraperitoneally (i.p.). Five mice received carbogen gas (95% O(2) and 5% CO(2)) for 9.5 min, starting 1 min before 5-FU administration. We found increased levels of 5-FU and its anabolites and catabolites by sequential ?19F NMR spectroscopy in the group treated with 5-FU in combination with carbogen compared with the group treated with 5-FU alone. The maximum of normalised values of 5-FU and its metabolites, reached after carbogen breathing, was almost 2-fold higher than after treatment with 5-FU alone. Despite these increased concentrations no significant effect of carbogen on growth inhibition of the tumour by 5-FU was observed, which may be related to the size as well as the well vascularised and perfused conditions of the tumours studied. PMID- 10854953 TI - Using administrative data to identify associations between implanted medical devices and chronic diseases. PMID- 10854954 TI - An introduction to epidemiological research with medical databases. AB - In most regards, database research is like any other epidemiological endeavor: excellent research can be conducted, but there are many potential difficulties. Training in appropriate epidemiological and statistical methodology, together with knowledge of the databases and their coding systems, is likely to magnify the advantages of databases and also minimize the potential problems. As in all epidemiological investigations, the quality of the data and the methodology employed need to be carefully considered in the context of the research questions at hand. PMID- 10854955 TI - A retrospective cohort study of implanted medical devices and selected chronic diseases in Medicare claims data. AB - PURPOSE: Several case-control studies have observed associations of implanted medical devices and certain connective-tissue and neurologic diseases. We reexamined these and other associations using cohort comparisons. METHODS: We compared the incidence of 52 diseases in several retrospective cohorts constructed from Medicare claims data. Six cohorts were defined by implantation of medical devices (silicone, metal bone or joint implants, breast implants, penile implants, pacemakers, artificial heart valves), and four comparison cohorts were defined by surgeries not involving implants. RESULTS: We observed associations that were generally consistent with previous reports, including associations of bone and joint implants with connective-tissue diseases, and an association of penile implants with idiopathic progressive neuropathy. We also observed associations of breast implants and pacemakers with connective-tissue diseases. CONCLUSIONS: For the most part, our study confirms our previous case control results. Although confounding by presurgical conditions (such as diabetes) remains a plausible explanation of the findings, several associations are worthy of more detailed research. PMID- 10854956 TI - Race, socioeconomic status, and cause-specific mortality. AB - PURPOSE: Life expectancy for black Americans is five to eight years less than for Whites. The socioeconomic status (SES) of Blacks is also less than for Whites, and SES is associated with early mortality. This paper estimates the proportion of the racial difference in mortality attributable to SES by specific causes of death. METHODS: Data on 453,384 individuals in the National Longitudinal Mortality Study were used to estimate the hazard ratio associated with black race, with and without adjustment for income and education (measures of SES), in 38 strata defined by cause of death and age. RESULTS: For women, SES accounted for much (37-67%) of the black excess mortality for accidents, ischemic heart disease (ages 35-54), diabetes, and homicide; but not for hypertension, infections, and stomach cancers (11-17%). For men, SES accounted for much of the excess risk (30-55%) for accidents, lung cancer, stomach cancer, stroke, and homicide; but not for prostate cancer, pulmonary diseases, hypertension, and cardiomyopathy (0-17%). CONCLUSIONS: These results confirm those specific causes of death likely to underlie the overall excess mortality of Blacks, and identify those causes where SES may play a large role. PMID- 10854957 TI - Marital status and mortality: the national longitudinal mortality study. AB - PURPOSE: To examine the effect of marital status (married, widowed, divorced/separated, and never-married) on mortality in a cohort of 281,460 men and women, ages 45 years and older, of black and white races, who were part of the National Longitudinal Mortality Study (NLMS). METHODS: Major findings are based on assessments of estimated relative risk (RR) from Cox proportional hazards models. Duration of bereavement for the widowed is also estimated using the Cox model. RESULTS: For persons aged 45-64, each of the non-married groups generally showed statistically significant increased risk compared to their married counterparts (RR for white males, 1.24-1.39; white females, 1.46-1.49; black males, 1.27-1.57; and black females, 1. 10-1.36). Older age groups tended to have smaller RRs than their younger counterparts. Elevated risk for non married females was comparable to that of non-married males. For cardiovascular disease mortality, widowed and never-married white males ages 45-64 showed statistically significant increased RRs of 1.25 and 1.32, respectively, whereas each non-married group of white females showed statistically significant increased RRs from 1.50 to 1.60. RRs for causes other than cardiovascular diseases or cancers were high (for white males ages 45-64: widowed, 1.85; divorced/separated, 2.15; and never-married, 1.48). The importance of labor force status in determining the elevated risk of non-married males compared to non married females by race is shown. CONCLUSIONS: Each of the non-married categories show elevated RR of death compared to married persons, and these effects continue to be strong after adjustment for other socioeconomic factors. PMID- 10854958 TI - The effect of alcohol abuse on the risk of NSAID-related gastrointestinal events. AB - PURPOSE: Non-steroidal anti-inflammatory drugs (NSAIDs) are known to increase the risk of gastrointestinal (GI) complications. Excessive alcohol consumption may further increase this risk and the FDA is requiring warnings on over-the-counter (OTC) NSAIDs. Our objective is to evaluate the risk of NSAID-related GI events for persons with a history of alcohol abuse. METHODS: This case control study used data from Saskatchewan Health. Cases consisted of 1083 patients hospitalized for severe GI events, whereas the control group consisted of 14,754 persons without such hospitalizations. RESULTS: Five percent of cases and 1.9% of controls had a history of treatment for alcohol abuse. The presence of either NSAID use or a history of alcohol abuse led to an odds ratio (OR) of 2.9* for severe GI events, whereas the presence of both risk factors simultaneously led to an OR of 10.2* (additive would be 5.8). Similarly, the presence of ibuprofen and naproxen use, which are OTC in the USA, without alcohol abuse led to an OR of 1.9*, whereas alcohol abuse by itself led to an OR of 2.4*. The presence of both OTC NSAIDs and alcohol abuse simultaneously, led to an OR of 6.5 (additive would be 4.3). Thus with both risk factors present, the resulting risk ratio is greater than the additive risk of the separate risk factors. CONCLUSIONS: The Food and Drug Administration (FDA) warning concerns concurrent use of alcohol with NSAIDs, whereas the present study presents the effect of long term alcohol abuse. Further research is needed to separate these two issues to allow physicians to provide the best advice to their patients. *Statistically significant at p<0.05. PMID- 10854959 TI - Validity and coverage of estimates of relative accuracy. AB - PURPOSE: Studies comparing test accuracy often restrict the confirmation procedure to subjects classified as positive by either test. Relative sensitivity (RSN) and relative false-positive rate (RFP) are two estimable comparative measures of accuracy. This article evaluates the influence of sample size, disease prevalence, and test accuracy on the validity of point estimates of RSN and RFP, and on the coverage of their confidence intervals (CI). METHODS: For each combination of sample size, disease prevalence, test accuracy, and interdependence between tests 1,000 samples were generated using computer simulations. The percent bias in the RSN and RFP estimates was measured by comparing the means of the 1,000 values computed in each simulation (log transformed) with their theoretical values. Coverage of the estimated CI was measured by computing the proportion that actually included the theoretical values. Application of these methods was illustrated with data from a study comparing mammography and physical examination in screening for breast cancer. RESULTS: RSN estimates were valid if the true number of diseased cases exceeded 30, and RFP estimates were valid if the number of nondiseased subjects exceeded 200. When the numbers of diseased and nondiseased subjects exceeded 150 each, the 95% CI of RSN and RFP provided adequate coverage of the parameters (95 +/- 2%). CONCLUSION: Sample size is the most important variable for the validity and coverage of RSN and RFP estimates. For small samples, validity and coverage of RSN and RFP also depend on the accuracy of each test and on the degree of interdependence between the tests. PMID- 10854960 TI - Clinician's guide to the choice of instruments for quality of life assessment (http://www.qlmed.org) PMID- 10854961 TI - Adaptive local contrast enhancement method for medical images displayed on a video monitor. AB - It is known that light-box luminance is an important factor in the detection of objects on radiographs. Existing methods for image contrast enhancement do not consider the luminance effect so that the enhancement may be inappropriate for the processed image displayed on a video monitor. Based on the properties of human visual system (HVS), an adaptive local contrast enhancement (ALCE) method was developed to enhance the medical image displayed on the video monitor by investigating the influence on human eyes of display luminance difference between a conventional light-box and a video monitor. The HVS model indicated parts of a radiograph unclear to human eyes at a given display luminance. By calculating the contrast of the image pixel by pixel, we found the parts of an image that are needed to be amplified at low display luminance and provided these parts with adequate enhancement. The processed image displayed on a video monitor was visually equivalent to the raw radiograph displayed on a light-box. A quantitative evaluation method of image quality assessment was used to compare the ALCE-processed image with others. PMID- 10854962 TI - A fast and accurate segmentation technique for the extraction of gastrointestinal lumen from endoscopic images. AB - A novel region-growing algorithm for the segmentation of endoscopic images is proposed in this paper. The objective of the research work is fast and accurate segmentation of gastrointestinal lumen from the endoscopic images for real-time applications. The proposed technique consists of a dual-step methodology in which a quasi Region of Interest (qROI) is segmented first using a global thresholding technique and then the actual lumen is extracted using differential region growing. An adaptive progressive thresholding technique is used to obtain qROI for a given endoscopic image. The centre of mass of qROI acts as the seed for the region growing in the next step. A differential region growing technique is suggested which grows the region on the basis of a similarity criterion. A dynamic hill-clustering method is utilised to ensure the effectiveness of the terminating condition during the growth process. The proposed scheme is faster than the conventional gradient based region-growing technique. The accuracy and high speed response of the proposed technique is validated with several endoscopic images and the results are presented. PMID- 10854963 TI - Variation of recruitment nonlinearity and dynamic response of ankle plantarflexors. AB - The ankle plantarflexor muscles of paraplegics may be trained to provide balance without support from the hands (in the laboratory environment) with the controller based on a two-block Hammerstein muscle model. This paper presents data on the variations of the recruitment curve block and linear dynamics block with electrode position, among various individuals and with fatigue. The tests were conducted in six groups: 'a' tests of a neurologically-intact subject were repeated on one day several times to record the effect of muscle fatigue; 'b' the same individual kept electrodes attached for a week and the muscle was identified every day; 'c' the same subject attached electrodes at marked positions every day for a week prior to identification; 'd' another normal attached electrodes at notionally the same positions over a period of one week; 'e' three normals and 'f' two paraplegics. Measurements were made with the Wobbler apparatus, in which the subject is supported upright in a standing posture. When comparing tests of fresh muscle every day, little difference was found between the nonlinear recruitment curves and linear dynamics of groups 'b' and 'c'. In fatigued muscle the dynamics were slower. When the electrode position was not carefully reproduced, and over a longer period, significant differences in nonlinearity appear in the curve shapes (group 'd') and a similar amount of variation occurs between normals, between paraplegics, and from normals to paraplegics. The paraplegic curves show wider deadbands. The effect of prolonged stimulation on normals is slight but on paraplegics it is significant. PMID- 10854964 TI - An experimental method for the application of lateral muscle loading and its effect on femoral strain distributions. AB - Experimental models that have been used to evaluate hip loading and the effect of hip implants on bone often use only a head load and abductor load. Anatomic considerations and in vivo measurements have lead several investigators to suggest that these models are inaccurate because they do not incorporate the loads imposed by additional muscles. The aim of this study was to evaluate the strains in the proximal and mid diaphysis of the femur for five hip loading models, one with a head load and abductor load only and four which incorporated lateral muscle loads as well. Head load to body weight load ratios were used to evaluate the physiologic accuracy of these models and strains were compared to determine the extent of strain changes as a function of model complexity. All models which incorporated additional lateral muscle loads more accurately simulated head load to body-weight load ratios than the simple abductor-only model. The model which incorporated a coupled vastus lateralis and iliotibial band load in addition to the abductor load provided the simplest configuration with a reasonable body-weight to head-load ratio. PMID- 10854965 TI - A finite difference model of load-induced fluid displacements within bone under mechanical loading. AB - Load-induced fluid flow in the lacunocanalicular network, induced by the mechanical loading of bone, is believed to play an important role in bone modelling, remodelling and adaptation processes. There are strong indications that this fluid flow is responsible for the mechanotransduction from external mechanical loads to the cells responsible for bone apposition or removal. Since direct flow measurements (especially in compact bone, in vivo and in situ) are not yet possible, theoretical modelling offers an alternative approach to determine the fluid flow velocities, displacements and effects of interstitial fluid flow. In this model, the fluid displacements in a middiaphyseal slab of a rat tibia under a cyclic four-point-bending load were calculated by applying Biot's theory of poroelasticity. The resulting differential equations were solved numerically for the fluid displacement vectors using the finite difference method. Thereby, the cross section located in the middle between the two inner points of force application was chosen for examination, such that the problem, although formulated in three dimensions, reduced itself to an essentially planar form. The maximal fluid displacements for the vector components in the cross sectional plane were found in the proximity of the neutral axis of bending. The direction of the displacement vectors was from the lateral aspect, which was in compression in the examined loading situation, towards the medial aspect in tension. In a parameter study it was found that the fluid displacement pattern and the distribution of fluid displacements remained constant for all the examined parameters, while the magnitude was influenced by the model parameters Young's modulus, Poisson's ratio and porosity. This study represents a further step in the examination of load-induced fluid displacements in loaded bone using theoretical models, aiming to understand the relationship between mechanical loading and bone modelling, remodelling and functional adaptation. PMID- 10854966 TI - Canalicular fluid flow induced by bending of a long bone. AB - Interstitial fluid flow has been hypothesized to underly mechanotransduction within bone. Here, we present an analytical model of fluid flows induced at the level of osteocyte canaliculi when a long bone is subject to functionally relevant bending loads. Dynamic bending of cortical bone results in a non-uniform longitudinal normal strain environment in which strain magnitude varies both temporally (i.e., at a given location, strain varies as a function of time) and spatio-temporally (i.e., at each given point in time, strain varies between locations). To account for the complexity posed by these two aspects of the strain environment, canalicular fluid flows were decomposed into temporal and spatio-temporal components. In terms of distribution around the cortex, temporal and spatio-temporal flows in the radial direction were both maximal near sites of peak strain magnitude. Spatio-temporal flows in the circumferential direction, in contrast, were maximal near locations of minimal strain magnitude (i.e. near the neutral axis). All fluid flow components were maximal during the first load cycle and reached markedly reduced steady state levels during subsequent load cycles. The novelty of the described model is that it provides the first estimate of canalicular fluid flows induced within a complexly loaded long bone. As the model may be readily extended to provide a simplistic accounting of the fluid flow profiles induced during functional loading and other exogenous loading regimes, the approach will enhance the ability to examine fluid flow related mechanotransduction within bone. PMID- 10854967 TI - A comparative analysis of fundamental frequency estimation methods with application to pathological voices. AB - One of the basic parameters characterising voiced phonation is the fundamental frequency, named pitch, which is the rate of vibration of the folds. In pathological voices. pitch variations within an utterance are indicative of the patient status. As such voices are corrupted by 'noise', robust pitch estimation methods are required in order to track its variations. This paper aims to compare some pitch estimation methods, pointing out their main advantages and drawbacks for present application. For each method, modifications are proposed in order to enhance performance. The methods are tested on simulated signals and then applied to real signals, coming both from healthy and pathological voices. The latter we obtained from patients who have undergone surgery for vocal folds via laser or traditional lancet techniques. PMID- 10854969 TI - A method for determining the heat transfer and water vapour permeability of patient support systems. Graham P. Nicholson, john T. Scales, OBE, raymond P. Clark and mervyn L. de calcina-goff medical engineering & physics, 1999;21:701 712 PMID- 10854968 TI - A laser plantar pressure sensor for the diabetic foot. AB - This paper describes the design and development of a high-resolution pressure sensor for use in the field of foot pressure measurement. The device uses an interferometry technique that involves the use of a laser light directed onto a pressure sensitive plate. The pressure plate compresses when a load is applied, and it is this compression which is measured by the interferometry technique. The interference pattern (interferogram) produced represents the pressure distribution across the plate. The interferogram is inputted into the computer using a high pixel CCD camera and video capture card. A fringe analysis program then converts the interferogram into a three-dimensional image of the pressure distribution. A prototype system is described and the initial results show that the system can produce a high resolution image with dynamic capabilities. PMID- 10854974 TI - [ [In Process Citation] PMID- 10854973 TI - [ [In Process Citation] PMID- 10854975 TI - [Introduction and history of surgery of the third ventricle]. AB - In this annual report of the Societe de Neurochirurgie de Langue Francaise, the authors reviewed in a retrospective study 262 cases of tumors exclusively located in the third ventricle (thalamic, chiasmatic and pineal tumors were excluded). Modern neuroradiology easily discloses these lesions, and therapeutic management represents the main challenge : most of the patients present with few symptoms, and even in some cases the lesions are diagnosed fortuitously. Neuropsychological assessment appears necessary to evaluate the long-term outcome of the patients : unfortunately few cases in this series were informative for these data. Walter Dandy described the operative approach of these tumors in 1922, and he also reported the benefits of endoscopy. Almost 80 years later, technical advances such as CSF shunts, stereotaxis, neuronavigation have improved the results of the neurosurgical treatment of these deep-seated lesions. PMID- 10854976 TI - [History of the cerebral ventricles]. AB - Along the centuries, the cerebral ventricles were thought by Ancient anatomists to be involved in rational judgement, imagination, and memory. The role played by the cerebral cortex in these functions was only identified during the XIX(th) century. PMID- 10854977 TI - [Fundamental anatomy of the third ventricle]. AB - Comparative anatomy of the third ventricle shows that this diencephalic cavity is present in most vertebrates. In humans, the roof of the third ventricle has thinned and most of its diverticles have disappeared. Human embryology underlines the development of hemispheric vesicles and interhemispheric commissures, and both of these structures progressively dissimulate the third ventricle deep within brain tissue. The anatomy of the third ventricle, particularly the roof of the ventricle and the interventricular foramen are described, with special references to their relationship with surrounding structures and vessels. The knowledge of these anatomical landmarks is essential to operative approaches to the third ventricle. PMID- 10854978 TI - [Functional anatomy of the third ventricle. Neuropsychological data]. AB - The functional structures surrounding the third ventricle explain the occurrence of neuropsychological disorders when a tumor develops in this area. The functional environment of the third ventricle is involved in memory, motor executive control, endocrine and vegetative regulations. The types of memory deficit, the behavioural and emotional regulations, the interhemispheric transfer, and the regulation of executive functions are analyzed and correlated to the concerned anatomical structures. The review of the literature collected few specific considerations in neuropsychological dysfunctions occurring with tumors of the third ventricle. A retrospective study was conducted in 17 patients of the national series operated on for a third ventricle lesion : long-term memory and executive functions were frequently impaired in the patients, and the deficits were underestimated by usual follow-up. More systematic utilization of preoperative and postoperative test batteries is necessary for a better evaluation of neuropsychological disorders after third ventricle surgery. PMID- 10854979 TI - [Surgical anatomy and surgical approaches of the third ventricle]. AB - Careful analysis of MRI images is mandatory before any surgical procedure in the third ventricle. This analysis should take in account the relationship of the tumor itself, but also the grade of hydrocephalus and the main anatomical landmarks along the surgical approach. The first step is the access to the lateral ventricle, which may be achieved via transcortical or anterior transcallosal routes : these two operative procedures are detailed. The transforaminal entry to the third ventricle may be easy if hydrocephalus has widened the foramen of Monro. In other cases, a subchoroidal (or interthalamo trigonal) approach is necessary, and the division of the thalamostriate vein is sometimes required. In this series, the transcortical route has been favoured by neurosurgeons. The advantages and drawbacks of both transcortical and anterior transcallosal routes are discussed. The anterior interhemispheric and pterional approaches are briefly evoked, as they were used in very few cases of this series. The management of hydrocephalus is discussed. PMID- 10854980 TI - [Endoscopic anatomy of the third ventricle]. AB - According to the development of neurosurgical endoscopy (and especially for third ventriculostomy), the endoscopic anatomy in hydrocephalus should be well known and utilized for orientation. The endoscopic pictures are obtained with a 30; telescope, acquired by a digitalized camera and visualized on a video monitor. The pictures are then numerized on a DKR system. Endoscopic anatomy of the third ventricle is described with a particular focus on the anatomical landmarks and their variations around the foramen of Monro, the anterior and posterior walls of the third ventricle. The knowledge of this anatomy is essential for the safety and the reliability of intraventricular endoscopic procedures. PMID- 10854981 TI - [Tumors of the third ventricle: review of 262 cases]. AB - The data from 262 cases of third ventricle (V3) tumors treated in 21 Departments of Neurosurgery in France between 1980 and 1995 were collected in this series. These tumors were frequent in young adults, and 17.5% of the patients were children. Colloid cysts (55%) and gliomas (19%) were the most frequent lesions. Other tumors were rare, or exceptional. CLINICAL PRESENTATION: The duration of symptoms was short in time, despite these lesions were usually benign. Most of cases were revealed by intracranial hypertension (63%), sometimes with a paroxystic or positional evolution. Neuropsychological signs (48 %) were undoubtedly under-estimated, revealing the disease in only 10% of cases. Ophthalmologic signs and endocrine disorders were infrequent. This feature is related to the selection of patients in this series, as tumors arising from the floor of the third ventricle or from the optic chiasm were excluded. Endocrine disorders were frequent with gliomas (30 %). THERAPEUTIC MANAGEMENT: In half of the patients, hydrocephalus was absent or mild and was ruled out after the treatment of the ventricular lesion. However, 12% of patients required a shunt procedure after the treatment of the ventricular lesion. A stereotactic procedure was performed in 63 patients, 12 had ventriculoscopy, and a direct surgical approach to the V3 was performed in 200 patients, sometimes after the failure of stereotactic or endoscopic procedures. Thirty six patients received no treatment. The patients were operated on via a transcortical approach (159 cases), or via the anterior transcallosal route (35 patients). Postoperative course was uneventful in 67% of the patients, complications were recorded in 24% of patients. MORTALITY AND MORBIDITY: The overall mortality in the national series is 13.7 % (36/262 died). The death occurred before any treatment (4 patients), or was directly correlated to the surgical procedure (13 cases), to long-term complications of hydrocephalus (2 patients), to general complications (7 patients), or to recurrence of the tumor (10 cases). The final outcome analysis recorded neurological impairment in 29% of cases, neuropsychological deficit in 50% of patients, and residual endocrine disorders in 19%. Social independence was recovered by 86% of patients, 76% of them returned to work, 72% of students returned to normal school attendance. The long-term neurological outcome was better with the transcallosal approach. No conclusion was possible concerning neuropsychological outcome, as postoperative neuropsychological assessment was not available for most of the patients operated on with the transcortical approach. PROGNOSIS: The results of treatment were evaluated only for the most frequent lesions (colloid cysts and gliomas). The outcome was worse for gliomas when compared to colloid cysts, considering mortality (13% vs 8%), neurological impairment (36% vs 21%), residual endocrine disorders (34% vs 0%), and ability to return to work (83% vs 56%). PMID- 10854982 TI - [Imaging of tumors of the third ventricle]. AB - MR imaging now appears as the best tool for diagnosis and pre-therapeutic assessment of tumors arising in the third ventricle (V3), as MR images can be obtained in the axial, coronal, and sagittal planes. MRI makes certain the location of the tumor in the V3 and delineates the anatomical landmarks in order to plan the surgical approach. It also allows the evaluation of associated hydrocephalus. Colloid cysts, choroid plexus papillomas, cavernomas, subependymal giant cell astrocytomas and germ-cell tumors may exhibit specific radiological features. PMID- 10854983 TI - [Pathological anatomy of tumors of the third ventricle]. AB - The walls of the third ventricle contain a large variety of tissues, and the pathological processes in this area are extremely varied. The pathological lesions encountered in the third ventricle are analyzed with interest to their incidence in the national series. Colloid cysts have a single layer of epithelial cells, mimicking the bronchus epithelium, which may reveal their probable endodermic origin. Xanthogranulomas are not true neoplasms but are only reactional to local hemorrhages. Histological diagnosis may be difficult for glial tumors if the biopsy sampling is insufficient : pilocytic astrocytomas can be overgraded and confused with high grade astrocytomas. Subependymomas should be separated from ependymomas, and giant cell subependymal astrocytomas are specific lesions occurring in tuberous sclerosis. Craniopharyngiomas often show a papillary type when located in the third ventricle. Germinal tumors are associated with immunological markers. The variety of the encountered lesions in the third ventricle needs a close collaboration between neuropathologists and neurosurgeons, and a confrontation of histological data with clinical and radiological data. PMID- 10854984 TI - [Stereotactic applications in third ventricle tumors]. AB - Stereotactic procedures are especially indicated for deep lesions within the brain substance, surrounded by functional structures, such as third ventricle tumors. Despite recent technological advances, the basis for stereotaxis remains minimally traumatic procedures, with the safety of vascular and ventricular anatomical definitions. Stereotactic procedures were performed in 63 patients of the national series. The indications, procedures, results and complications were reviewed. Stereotaxis was used with a diagnosis purpose in 34 patients. In all cases, the histological diagnosis was identified, which allowed the definition of the therapeutic strategy, and avoidance of useless direct approach to the third ventricle. Stereotaxis was used with a therapeutic purpose in 29 other patients : the procedure allowed the evacuation of colloid or glial tumor cysts, ventriculostomy, intracavitary irradiation, or radiosurgery. The stereotactic procedure should take in account the grade of associated hydrocephalus. Stereotactic evacuation of colloid cysts in this series (25 cases) often resulted in a recurrence of the cyst. Endoscopic procedures, eventually associated with stereotactic guidance, should be preferred for the treatment of colloid cysts. PMID- 10854985 TI - [Neuronavigation in third ventricle tumors]. AB - The authors present the results of neuronavigation as a help to open neurosurgery for the tumors of the third ventricle. From January, 1995 to August, 1999, six image-guided surgical procedures were performed to remove third ventricle lesions : 4 colloid cysts, 1 ependymoma, and 1 craniopharyngioma. The operative approach was transcortical in 5 cases, and transcallosal in 1 case. The use of neuronavigation allows a decrease of the surgical trauma during the surgical approach. The procedure secures the neurosurgeon in the choice and execution of his pathway to the target. It becomes however less accurate after opening the ventricle, because of the brainshift induced by the loss of cerebrospinal fluid becomes important. Nevertheless, neuronavigation is useful in the surgery of the third ventricle, especially if it is used with neuroendoscopy. PMID- 10854986 TI - [Endoscopic surgery of third ventricle lesions]. AB - The endoscopic approach of the tumors of the third ventricle interests mainly the colloid cysts but also offers the possibilities of biopsies. Twenty two patients (16 men and 6 women, average age 41 years) presenting with hydrocephalus related to a tumor of the pineal area were treated by a ventriculostomy with attempt at biopsy : they are outside of the limits of this report. Twenty two other patients (15 men, 7 women, average age 39 years) were operated on from 1994 to 1999 for a colloid cyst, and 2 of them were admitted in emergency in sudden coma. The CT scan showed a colloid cyst (hyperdense in 16 patients) associated with an hydrocephalus, except for a patient previously shunted. The diameter of the cyst varied from 4 to 50 mm (average of 20 mm). All the patients were operated on using a rigid endoscope. Among the 20 patients presenting a tumor of the pineal area, a biopsy was possible only in 4 cases (20%). There were no hemorrhage nor neurological disorders. In all the cases, the size and the number of the specimens were sufficient to allow the histological diagnosis. For the patients presenting with colloid cyst, the average follow-up is 2 years. All the preoperative symptoms disappeared except for the memory disorders which were improved. The post-operative Evans index decreased significantly. No residual cyst was observed on the post-operative MRI in 14 patients (63%). Among these patients, an asymptomatic recurrence was observed and remained stable after 44 months of follow-up. A residual cyst was observed in 8 patients (36%), with a diameter from 5 to 25 mm (average 9 mm). No patient required a shunt procedure, and no patient presented hemorrhagic complication. Endoscopy is especially useful in the first line treatment of the colloid cysts of the third ventricle. PMID- 10854987 TI - [Tumors of the third ventricle: review of the literature]. AB - In this chapter, the authors compared the results of the national series (262 patients) with the relevant data from the literature, in order to establish the optimal therapeutic management. Colloid cysts represent the main tumor encountered within the third ventricle (145 cases). Sudden death from colloid cysts is documented in this series, but neurosurgeons should also be aware of the increasing number of fortuitous diagnosis of such lesions. The liquid consistency of the content of the cyst is the major argument to choose the surgical procedure. Liquid colloid cysts usually associate isodense appearance on CT Scan, hypo-intensity on T1 weighted MRI images, and hyperintensity on T2-weighted MRI images. Endoscopy now appears as an accurate surgical procedure. A surgical strategy is proposed. Among glial tumors, pilocytic astrocytomas are poorly documented in the literature. Low grade astrocytomas, ependymomas, subependymomas and giant cell subependymal astrocytomas, gangliogliomas, and oligodendrogliomas were reviewed with the relevant literature. Other tumors are rare in the third ventricle, except for choroid plexus papilloma and craniopharyngioma. The reality of purely intraventricular craniopharyngioma is confirmed by the experience of this series. Other lesions such as meningioma, cavernoma, germ-cell tumors, lymphomas, metastasis, epidermoid cysts, and exceptionnal lesions were also reviewed. PMID- 10854988 TI - [Tumors of the third ventricle in children: review of 46 cases]. AB - In the national series, 46 patients out of 262 (17.5%) were children under 15 years of age. Most of them presented with gliomas (50 %), choroid plexus tumors were recorded in 8 cases (17%). Intracranial hypertension revealed the tumor in 80% of children, endocrine disorders were recorded in 8 patients. A stereotactic procedure was performed in 7 patients, 1 had ventriculoscopy, and a direct surgical approach to the third ventricle was performed in 37 patients, in one case after the failure of the endoscopic procedure. Two patients with a radiological appearance of hamartoma received no treatment. Most of patients were operated on via a transcortical approach (27 cases). The overall mortality in this series is 9%. The death was directly correlated to the surgical procedure (1 case), or to recurrence of the tumor (3 cases). The final outcome analysis recorded neurological impairment in 50% of cases, neuropsychological deficit in 58 % of patients, and residual endocrine disorders in 41%. Social independence was recovered by 82% of the children, and 68% returned to normal school attendance. PMID- 10854990 TI - [ [In Process Citation] PMID- 10854989 TI - [Third ventricle tumors in children]. AB - Tumors of the third ventricle in children raise specific problems, owing to their clinical presentation, pathology, treatment, and outcome. The clinical presentation is often unspecific at that age, many cases presenting with isolated macrocrania or delayed milestones. Some histological types are specific of the pediatric group, such as the pilocytic astrocytoma or hypothalamic hamartoma, and others types which are exceptional in adults are relatively common in children, such as choroid plexus tumors. Surgery is especially dangerous in young children, because of the blood loss, and the hemispheric collapse associated with the large hydrocephalus which is the rule in these patients. External irradiation is associated with a high rate of complications, above all a progressive intellectual delay, and endocrine disorders. Chemotherapy has reduced the indication of irradiation, and, in some cases, can be performed preoperatively to reduce tumor volume and intraoperative blood loss. The therapeutic approach to these patients is thus pluridisciplinar, and should be tailored for each case, with a follow-up protracted well into the adult age. PMID- 10854991 TI - An assessment of the spread of HIV infection in Poland and Ukraine. Introduction. PMID- 10854992 TI - HIV infection in Ukraine (1987-96). AB - In Ukraine, the number of reported HIV infections increased extremely rapidly during the second half of the 1990s, from less than 50 per year until 1994 to more than 12,000 in 1996. The increase was initially observed and was particularly striking in the regions along the Black Sea. The majority of reported HIV infections were diagnosed in injecting drug users. The extend of HIV spread through sexual transmission is more difficult to assess because of the strong social stigma attached to homosexuality and the lack of information on sexual behaviour in general. The reported number of syphilis cases have also dramatically increased, from 3,000 cases in 1990 to nearly 80,000 cases in 1996. In this paper, we describe the surveillance systems for, and epidemiologic data on HIV infections, AIDS, and other STD in Ukraine from 1987 to 1996. We review the contributions of different vulnerable groups and we also discuss the factors influencing the past spread and the potential for future spread of HIV infection and make recommendations for surveillance, research and prevention. PMID- 10854993 TI - HIV infection in Poland (1985-96). AB - Reported numbers of HIV infections have provided an indication of the order of magnitude of the size of the HIV epidemic in Poland, and how it has evolved in time. To the end of 1996, most documented infections had been observed among intravenous drug users (IDUs, n=2, 933/3,470) and, to a lesser extent, men who have sex with men (MSM, 289/3,470). Reported infections among non-IDU prostitutes and other non-IDU heterosexuals remained low. In this paper we describe surveillance systems and epidemiologic data in Poland from 1985 to 1996 for AIDS cases, HIV infections and other STDs. We also discuss the contributions of different vulnerable groups. Finally we discuss factors influencing the past spread of HIV infection and the potential for future spread, and propose recommendations for surveillance and research. PMID- 10855006 TI - [Acinetobacter: multiresistance or survival?]. PMID- 10855007 TI - [Interactions between bacteria and antimicrobials during the postantibiotic effect phase]. PMID- 10855008 TI - [Peptides of eukaryotic origin with antimicrobial activity]. PMID- 10855009 TI - [HIV-1 resistance and the clinical laboratory]. PMID- 10855010 TI - [Evaluation of mutations that confer resistance to nucleoside analogs and protease inhibitors in HIV-1-infected patients. Study Group on Resistance to Antiretroviral Agents]. AB - Genotypes that confer drug resistance to reverse transcriptase inhibitors and protease inhibitors were evaluated in HIV-1 proviral DNA obtained from peripheral blood mononuclear cell samples. Fifty-three HIV-1-infected patients were studied, 19 of whom had not received antiretroviral treatment. In the other 34 patients, 9 had been treated with combinations of two reverse transcriptase inhibitors (AZT, ddI, d4T, 3TC) and 25 had been treated with triple antiretroviral therapy including a protease inhibitor (nelfinavir, indinavir, saquinavir, ritonavir). To determine the presence of mutations involved in the development of resistance to reverse transcriptase inhibitors a hybridization Microtiter assay was carried out. Mutations were detected in treated patients as well as in those without previous antiretroviral treatment, with the most frequent mutations being those that confer resistance to AZT, followed by those that develop cross-resistance to ddI/ddC and 3TC, which are the most commonly used drugs to date. No mutations were detected to any nucleoside analog in only 13 cases. To analyze the presence of mutations in the protease gene a dot-blot hybridization was carried out which included the mutations in codons 36, 82 and 90. Mutation 82 was detected in one case. Therefore, with the aim of determining the pattern of genotypic mutations in patients infected with HIV-1 and in order to make the best therapeutic choice, it would be recommended to consider carrying out genotypic resistance assays in clinical practice. PMID- 10855011 TI - [Should cefminox substitute cefoxitin in infections caused by bacteria susceptible to both drugs?]. AB - We studied the in vitro activity of cefminox (MIC by agar dilution) and cefoxitin against 477 facultative Gram-positive and Gram-negative isolates which were recovered from clinical specimens and identified by standard methods. Cefminox has proved to be 4-16 times more active than cefoxitin against enteric Gram negative bacilli commonly involved in cholecystitis, secondary peritonitis, intraabdominal abscesses and gynecological infections. On the contrary, cefoxitin has proved to be four times more active against Gram-positive cocci. Both have been ineffective against Enterobacter spp., Pseudomonas aeruginosa and Acinetobacter spp. The microbiological activity and pharmacokinetic parameters of cefminox make it one of the first choice treatments for community-acquired intraabdominal and pelvic infections with the presence of anaerobes. PMID- 10855012 TI - Antimicrobial susceptibility of anaerobic and aerobic bacteria isolated from patients with mixed infections in Nicaragua. AB - The agar dilution method was used to test the activity of ampicillin, benzylpenicillin, cefoxitin, imipenem, clindamycin, metronidazole, chloramphenicol, gentamicin, methicillin and vancomycin against 241 anaerobic and 227 aerobic bacteria isolated from 136 patients with intraabdominal infections and 49 with nonintraabdominal infections. Beta-lactamase production was tested in all strains. Overall, imipenem, metronidazole and chloramphenicol were the most active antimicrobial agents against anaerobic bacteria followed by clindamycin. Only the Bacteroides fragilis group was shown to be less susceptible to clindamycin (MIC90 8 mg/l). Ampicillin and cefoxitin were the least active beta lactam antibiotics against the most common isolated B. fragilis group strains (MIC(90) >1024 and 64 mg/l, respectively) and against Escherichia coli strains (MIC(90) >1024 and >1024 mg/l, respectively). Chloramphenicol showed low activity against the Gram-negative aerobic bacteria, while gentamicin had good activity against the aerobic bacteria tested, except for E. coli and Pseudomonas. Among the Gram-positive aerobic and anaerobic bacteria tested, Staphylococcus aureus was shown to be less susceptible to beta-lactam antibiotics (29% were methicillin resistant). No vancomycin-resistant S. aureus strains were found. A good correlation between beta-lactamase production and beta-lactam resistance was observed. PMID- 10855013 TI - [Relationship between antimicrobial susceptibility and virulence factors in Helicobacter pylori clinical isolates]. AB - The aim of this study was to determine the relationship between the antibiotic susceptibility and different virulence factors among Helicobacter pylori clinical isolates. One hundred and forty-five strains were isolated from biopsy cultures obtained from adult patients. Antimicrobial susceptibility to amoxicillin, clarithromycin, metronidazole and tetracycline were tested using an agar dilution method. cagA and iceA genes and s1 and s2 alleles of vacA were studied by polymerase chain reaction. A group of patients had been previously treated for H. pylori infection. We found a resistance rate of 28.7% and 16.5% to metronidazole and clarithromycin, respectively. We did not find any resistance to amoxicillin or tetracycline. The cagA gene and s1 allele were detected in 86.3% and 65.2% of the strains. One hundred and two (71.3%) strains were iceA+. cagA+ strains showed lower percentages of resistance to antibiotics, as did vacA s1 and iceA+ strains. The role of lower rates of resistance to clarithromycin and metronidazole in more virulent H. pylori strains may have favorable effects in their eradication in patients infected with these strains. PMID- 10855014 TI - [Phenotypes of macrolide, lincosamide, and streptogramin resistant Streptococcus viridans isolated from blood]. AB - Macrolide resistance has been widely studied in Streptococcus pyogenes and Streptococcus pneumoniae although not in viridans group streptococci (VGS). We studied 30 blood culture isolates of viridans group streptococci (25 resistant to erythromycin: 10 S. mitis, 8 S. milleri, 6 S. sanguis and 1 S. salivarius; and 5 susceptible: 2 S. mitis, 2 S. milleri and 1 S. sanguis). We carried out a double disk test and determined MICs. The susceptibility testing was carried out by agar dilution for 14-, 15- and 16-member lactone ring macrolides, as well as for clindamycin and quinupristin-dalfopristin. Fifty-six percent of the erithromycin resistant strains (6 S. mitis, 6 S. milleri and 2 S. sanguis) showed an MLS(B) phenotype, with a high level of intrinsic resistance to all the macrolides studied and clindomycin; 28% were of the M phenotype (4 S. sanguis, 2 S. mitis and 1 S. salivarius). We found a third resistance phenotype, which was present in 4 strains with MICs of 2-8 microg/ml, with resistance to macrolides and different degrees of resistance to clindamycin. All isolates were fully susceptible to quinupristin-dalfopristin. The MLS(B) and M phenotypes initially described in S. pyogenes and S. pneumoniae are also observed in VGS. PMID- 10855015 TI - [Multicenter study of the variability and adequacy of antimicrobial therapy for community-acquired pneumonia in adults]. AB - We performed a study to evaluate the variability and adequacy of prescribing antibiotics in community-acquired pneumonia (CAP) in 10 Spanish hospitals. We studied 452 patients with CAP. Initial empirical administration of antibiotics was prescribed in 90.7% of the cases, 82.5% as monotherapy. Macrolides and third and second generation cephalosporins were the most widely used groups of antibiotics. Penicillin and amoxicillin were only prescribed in 1. 7% of the patients. A significant variability between hospitals was observed. Reference patterns for the use of antibiotics in CAP were devised by a panel of experts. According to the recommendations of this panel, 29% of the total prescriptions were not adequate, with this percentage reaching 65% in outpatients older than 65 years or with comorbidity. This was mainly due to the fact that monotherapy with erythromycin, which was considered inadequate, was the most widely prescribed treatment. PMID- 10855016 TI - [Piperacillin: use at a general hospital]. AB - AIM: To analyze the use of piperacillin in a general hospital in 1996 and 1997. MATERIAL AND METHODS: The data from 338 patients who were treated with piperacillin between January 1996 and December 1997 were collected retrospectively. RESULTS: We analyzed 385 infectious episodes in 338 patients with a mean age of 63.1 years. The therapeutic indications included nosocomial infection, infection in neutropenic patients and other diagnoses such as respiratory, intraabdominal and osteoarticular infections. In most episodes, there were infections in neutropenic patients, representing 39.2% of all cases. In 86.2% of the episodes the combination of piperacillin and aminoglycoside was established. The mean duration of the treatment with piperacillin was 8.9 days. Positive cultures were presented in 39.7% of the episodes. The microorganisms most frequently isolated were the Gram-negative bacilli, such as Pseudomonas aeruginosa and Escherichia coli. The isolations were obtained from sputum (24.6%), blood (22.1%), urine (19.8) and exudate (8.8%). The activity of piperacillin and aminoglycoside was 93% and 90% respectively against P. aeruginosa. The global mortality of the series was 25.5% with 18.9% being related to the infectious episode. CONCLUSIONS: From the above analyses we can deduce that piperacillin maintains its indications in a general hospital, these being, basically, neutropenic infection and nosocomial pneumonia. Another indication is infection by Pseudomonas, against which it continues to show good activity. PMID- 10855017 TI - [Evolution of resistance to quinolones in Salmonella enterica in our setting]. AB - Salmonella enterica is mainly associated with acute gastroenteritis; however, it is also associated with other more severe disease processes, for which quinolones are the treatment of choice. We retrospectively studied the evolution of resistance to nalidixic acid and ciprofloxacin of all the clinical isolates of S. enterica from 1992 to 1998 in our hospital environment. A total of 848 strains from feces, blood and other locations were studied. We detected an increase in the resistance to nalidixic acid from 12% in 1992 to 21.3% in 1998, especially in the Enteritidis and Hadar serotypes. We did not detect resistance to ciprofloxacin, but there was in increase in the MIC in the nalidixic acid resistant strains. Although this is interpreted as sensitive according the the NCCLS criteria, if we apply the cutoff points established by MENSURA, 89.46% of the strains do not fit into this category (S <0.12 mg/l). This puts into question the utility of quinolones in the long-term treatment of severe disease processes produced by this type of strain. PMID- 10855018 TI - [Pneumonia caused by Haemophilus influenzae. Study in a series of 58 patients]. AB - Haemophilus influenzae tends to form part of the usual respiratory flora in adults, especially if they have a chronic underlying disease or are smokers. Pneumonia due to H. influenzae is frequently involved in respiratory infections and its level of resistance to ampicillin has remained stable over the last five years. Most of the literature on the subject was published more than 10 years ago. In this study, we describe the clinical features and evolution of 58 adult patients admitted to hospital for pneumonia due to H. influenzae over a 2-year period, with this group accounting for 6.5% of all the patients admitted with pneumonia during this time period. The etiological diagnosis was made using a good quality sputum sample. Forty patients (69%) were male. The mean age (+/- SD) of the group was 67 (+/-16.8) years and all the patients had at least one underlying disease. The mean duration of the symptoms was 6.7 days. All patients presented an increase in the quantity or purulence of the sputum. On admittance, respiratory failure was present in 52 patients (90%). Gram-negative coccus bacilli were observed in the direct sputum test and H. influenzae grew in the culture. In two cases, H. influenzae was recovered from the blood culture and in one from bronchial aspiration obtained through bronchoscopy. Another pathogen was identified in 28 patients (48%). In 21 it was another pyogenic bacteria (15 S. pneumoniae, 4 M. catharralis, 1 K. pneumoniae, 1 E. coli), an atypical microorganism in 5 (3 C. pneumoniae, 2 C. burnetii) and a respiratory virus in 2 (syncytial and influenza A). Atypical bacteria and respiratory virus were detected using serological techniques. The radiographic infiltrate was unilobar in 54 of the 58 patients and all showed an alveolar pattern. The empirical treatment included the administration of a third generation cephalosporin (or a fluoroquinolone in patients allergic to penicillin). The evolution was favorable in all the cases in which H. influenzae was the only pathogen or was accompanied by an atypical microorganism or a respiratory virus. Four patients with mixed bacterial pneumonia died (2 S. pneumoniae, 1 E. coli and 1 M. catharralis). The study indicates that pneumoniae due to H. influenzae affects a population with an underlying disease, preferably pulmonary, that it has a longer clinical period than that for pneumococcal pneumonia, that it is slightly bacteremic and, that, usually, it evolves benignly with a low mortality. PMID- 10855019 TI - [Antimicrobial therapy and evidence-based medicine]. PMID- 10855020 TI - [Eukaryotic antibiotic peptides: a evolutionary relic or therapeutic remedy?]. PMID- 10855021 TI - Methodology of case-control studies in the epidemiology of multidrug-resistant tuberculosis. PMID- 10855022 TI - [Activity of betalactamase inhibitors against Acinetobacter baumannii]. PMID- 10855023 TI - [National multicenter study of the in vitro activity of moxifloxacin against respiratory tract pathogens. Spanish Study Group on Moxifloxacin]. AB - The activity of moxifloxacin, a novel 8-methoxyquinolone, was evaluated against 1,218 respiratory pathogens isolated in nine Spanish hospitals and was compared with ciprofloxacin, sparfloxacin, amoxicillin, amoxicillin-clavulanic acid, cefuroxime, erythromycin and clarithromycin. Moxifloxacin exhibited an excellent in vitro activity against most tested isolates with MIC90 values of 0.25 mg/l for Streptococcus pneumoniae and viridans group streptococci; 0.12 mg/l for Streptococcus pyogenes; 0.25 mg/l for Streptococcus agalactiae; 0.06 and 4 mg/l for methicillin-susceptible and -resistant Staphylococcus aureus, respectively; 0.06 mg/l for Haemophilus influenzae and 0.12 mg/l for Moraxella catarrhalis. Moxifloxacin susceptibility rates were not affected by penicillin resistance in S. pneumoniae and S. viridans, by the betalactamase production in H. influenzae and M. catarrhalis or by macrolide resistance. Moxifloxacin was twice as active as sparfloxacin and four to sixteen times more active than ciprofloxacin against Gram-positive isolates. Sparfloxacin and ciprofloxacin were slightly more active than moxifloxacin when tested against H. influenzae and M. catarrhalis isolates. The microbiological data obtained confirm that moxifloxacin is a promising antimicrobial agent for treating respiratory tract infections. PMID- 10855024 TI - [Epidemiological aspects of ciprofloxacin-resistant Escherichia coli at a general hospital]. AB - The susceptibility to ciprofloxacin of 7288 Escherichia coli clinical isolates from 5667 patients was determined over a 4-year period (1995-1998). Information about the patients' age, sex, specimen type, date, origin and susceptibility to Escherichia coli isolates was studied, and the c2 test was used for statistical comparison. Overall, 1003 (17.70%) patients out of the 5667 included in the study presented ciprofloxacin resistance. The annual resistance observed over this 4-year period was not considered statistically significant. The resistant isolates were more frequent among men, in urine specimens and in outpatients, and increased with different age groups. The average age of patients with resistant isolates was 61.29 years (SD 21.56) and that of patients with susceptible isolates was 39.76 years (SD 27.41). A similar rate of resistance was observed among outpatients from health centers and those from hospital outpatient services. The higher resistance rates were found in outpatients from the urology department. The resistance to other fluoroquinolones remained the same and was not significant for norfloxacin; it increased starting from 1997 for pefloxacin at the expense of ciprofloxacin-sensitive isolates. PMID- 10855025 TI - [ROC curve analysis of factors predictive of non-response to interferon treatment in patients with chronic hepatitis C, genotype 1]. AB - The objectives of this study included: 1) to identify pretreatment variables predictive of absence of response in 107 patients with chronic hepatitis C, genotype 1, treated with interferon-a (IFN-a) at a dose of 3 MU three times weekly for 3-12 months and classified into two groups: group A, nonresponders vs. patients with a complete response, and group B, nonresponding and relapsing patients vs. patients with a sustained response; and 2) to establish a prognostic index using ROC curve analysis. The rate of sustained response was 6. 5% at the 24-month follow-up. The pretreatment characteristics with predictive value using ROC curves were as follows: in group A, age, GGT, serum ferritin, viral load, and grade and stage of the histological lesion; and in group B, known duration of infection, GPT, GGT, serum ferritin, viral load, and grade and stage of the histological lesion. In both group A and group B the positive predictive value (the probability of predicting an absence of response when the variable is present) was greater than the negative predictive value (mean: 84.3% vs. 41.1%, 99% vs. 16.5%, respectively). In group A, based on the prognostic index, the positive predictive value when three variables were present was 96% and the sensitivity was 63.5%, with the test being unequivocal in 6.5%, whereas when four or five variables were present, the positive predictive value was 97% and 100% and the sensitivity was 40.5% and 18%, respectively. In group B, the positive predictive value when two variables were present was 100% and the sensitivity was 87%, whereas when three, four, five and six variables were present the sensitivity was between 73% and 28%. In group A, age, GGT and ferritin were the predictive variables independently associated with an absence of response, with a relative risk of 6.5, 4.8 and 3.1, respectively, whereas in group B we did not find variables independently associated with an absence of response. It was concluded that in patients with genotype 1, it is possible to predict the absence of response to IFN therapy with a high degree of reliability. PMID- 10855026 TI - [Susceptibility to fluconazole and itraconazole in isolates of Candida spp. from HIV-positive and HIV-negative patients]. AB - We studied the possible differences in the pattern of susceptibility to fluconazole and itraconazole in 393 isolates of Candida spp. from the oral cavity of HIV-positive patients and 102 isolates from HIV-negative patients with candidemia or candiduria. We used the broth microdilution method according to the NCCLS guidelines. We observed a decrease in the susceptibility to fluconazole in the group of HIV-positive patients in comparison to those who were HIV negative, especially in Candida albicans (MIC(90) 32 mg/l vs. 1 mg/l and Candida glabrata (MIC(90) 64 mg/l vs. 16 mg/l). Furthermore, we did not find any resistant strains in the HIV-negative group. For itraconazole, the MIC(90) was two dilutions greater in the HIV-positive patients, except for C. albicans, which had a much higher MIC(90) (4 mg/l vs. 0.12 mg/.). Therefore, the decrease in the susceptibility of Candida spp. in the HIV-positive patients must be taken into account when establishing a specific antifungal therapy. PMID- 10855027 TI - [Guidelines for empirical antibiotic treatment of intraabdominal infections. Spanish Society of Chemotherapy]. PMID- 10855028 TI - Isolation of primary open-angle glaucomatous trabecular meshwork cells from whole eye tissue. AB - PURPOSE: Isolation and culture of human trabecular meshwork (TM) cells from primary open-angle glaucomatous (POAG) tissue has proven difficult. The objective of this study was to directly compare the utility of two different isolation methods to obtain viable human TM cells from POAG whole eye tissue. METHODS: Using a blunt dissection technique, human TM tissue was obtained from four pairs of donor eyes (67, 77, 81 and 82 years) with a documented history of POAG. TM tissue from one eye was explanted into tissue culture. TM from the contralateral eye was digested with a collagenase mixture and seeded onto culture plates. RESULTS: Primary cell isolates were obtained from all donors with both techniques. However, only cells obtained using the digestion method (3 of 4 TMs) could be passaged for expansion and freeze-downs (3 x 107 second passage cells/donor). None of the cells obtained from explanted TMs could be passaged. Cells from successful isolations were of uniform size, possessed typical TM morphology and had doubling times < 48 hours. CONCLUSION: These results demonstrate a clear advantage to digesting the extracellular matrix of glaucomatous TM tissue to obtain sufficient numbers of healthy cells for use in experiments. In contrast to cells obtained from explants, cells liberated from POAG TM tissue by digestion appear indistinguishable morphologically and behaviorally from "normal" TM cells. PMID- 10855029 TI - Assessment of ascorbate ocular disposition in the conscious rabbit: an approach using the microdialysis technique. AB - PURPOSE: This study was performed to evaluate the transport kinetics of ascorbate in aqueous humor of conscious rabbits. METHODS: Following the development of a spectrophotometric assay for ascorbate in serum, aqueous humor and microdialysate, and preliminary studies of ascorbate systemic disposition in the rabbit, microdialysis probes were placed into the anterior chamber of one eye, and the posterior chamber of the contralateral eye, of four New Zealand white rabbits. After a one-month recovery period, conscious rabbits were placed in restraining devices, and marginal ear veins were cannulated for repeat blood sampling and ascorbate administration. A tracer i.v. bolus of (14)C-ascorbate, followed by stepwise increasing i.v. infusions of unlabelled ascorbate, was administered. Estimates of basal ascorbate transport into aqueous were determined by analysis of tracer ( 14)C-ascorbate in microdialysis probe effluent and serum. Kinetic modeling was employed to assess ascorbate disposition during infusion. RESULTS: Systemic disposition of exogenously administered ascorbate was well characterized by a two-compartment model. Kinetic modeling returned physiologically realistic volumes for the posterior chamber, and reliable estimates governing ascorbate flux into, between, and from the posterior and anterior chambers. CONCLUSIONS: In vivo assessment of ascorbate kinetics in aqueous humor and blood of the rabbit was facilitated by the microdialysis technique. Contrary to reports in the literature, ascorbate saturable uptake from blood to aqueous was not observed at physiologic blood concentrations ( approximately 11 to 30 mg/L). PMID- 10855030 TI - Optimization of non-viral gene transfer to human primary retinal pigment epithelial cells. AB - PURPOSE: To optimise the high efficiency, non-viral transfer of DNA to retinal pigment epithelial (RPE) cells in vitro. METHODS: A mammalian expression vector (pcDNA3.1) containing a firefly luciferase (luc) cDNA was used to transfect RPE cells using different chemical methods; calcium phosphate, DEAE-dextran and, liposomes-based transfection techniques. Transfection was optimised for both dose and time of exposure. The efficiency of gene transfer and cytotoxicity was measured 48 hours post-transfection using luciferase and MTT assays, respectively. The percentage of transfected cells (using optimal conditions) was determined with a construct expressing a jellyfish green fluorescent protein (GFP) using flow cytometery. RESULTS: Calcium phosphate and DEAE-dextran techniques failed to transfect the vector and led to high cytotoxicity. Liposomes based methods successfully transferred the vector to RPE cells, but the efficiency varied for different liposomes; Tfx-50 > Lipofectin > Lipofectamine > Cellfectin > DMRIE-C. No significant cytotoxicity was observed with any of the liposome treatments. Optimal transfection was achieved with Tfx-50 at a 3:1 ratio of DNA:liposome; between 12-15% of cells being transfected. CONCLUSIONS: Efficient and non-toxic transfer of functional genes into primary RPE cells in vitro can be successfuly achieved by liposomes-based techniques. Tfx-50 appears to be a promising non-viral vector for RPE gene transfer. PMID- 10855031 TI - Apoptosis is associated with formation and persistence of the embryonic fissure. AB - PURPOSE: The role of apoptosis in the transitory ocular embryonic structures has not been clarified yet, therefore, in the present study we focused on one of the transitory ocular structures, the embryonic fissure. To elucidate the developmental mechanism of the embryonic fissure, we observed cell death by apoptosis in the optic cup during early development in mice. METHODS: Pairs of C57BL6N/Jcl mice, each comprising an estrous female and a potent male, were caged together overnight. Females that had vaginal plugs the next morning were considered at day 0 of pregnancy. The embryos or fetuses were removed by laparotomy on days 9, 10, 11, 12, 13, 14, 16, and 18 of gestation. Tissue blocks of the eyes were fixed and embedded in paraffin wax. Serial frontal sections of the eye were cut and stained with the TUNEL method and then counterstained by hematoxylin or methyl green solution. We examined TUNEL-positive cells in the optic cup by light microscopy. RESULTS: TUNEL-positive cells were seen at the lower nasal side of the optic cup, corresponding to the presumed embryonic fissure area, on day 9 of gestation before the formation of the embryonic fissure. Many TUNEL-positive cells were present at the lips of the embryonic fissure on days 10, 11, and 12. In contrast, TUNEL-positive cells were not detectable in the corresponding area on day 13 after the complete closure of the embryonic fissure. CONCLUSIONS: Apoptosis is anatomically closely associated, and appears to be essential for the formation and persistence of the embryonic fissure. PMID- 10855032 TI - Protein phosphatase inhibitors alter cellular microtubules and reduce carbachol dependent protein secretion in lacrimal acini. AB - PURPOSE: To further understand the regulation of microtubules and their function in the lacrimal gland, we investigated the effects of two serine/threonine phosphatase inhibitors, okadaic acid (300 nM-1 microM) and calyculin A (20-100 nM), on microtubules and stimulated secretion in lacrimal acini. METHODS: Primary rabbit lacrimal acini cultured for two days were utilized. Microtubule structure was probed using biochemical analysis and confocal fluorescence microscopy. Carbachol-stimulated and basal protein secretion were determined by measurement of released protein or, for pulse-chase studies, [(35)S]-protein. RESULTS: Biochemical analysis and confocal fluorescence microscopy showed that both inhibitors caused a major loss of cellular microtubules and also of acetylated (stable) microtubules. However, calyculin A was more potent than okadaic acid in causing microtubule loss. Because changes in microtubules can partially impair stimulated protein secretion in lacrimal acini, the effects of inhibitors on protein secretion were also evaluated. Both inhibitors caused a comparable dose dependent and significant (p or = 90 degrees and bilateral synostosis; in both cases, the disability must be very severe. The operative technique used appeared to be very well adapted to this disease: "reed" radial osteotomy and transverse ulnar osteotomy. No complications or relapses were observed in six operated patients with a mean follow-up of 10 years, with a mean loss of derotation of only 5 degrees. PMID- 10855299 TI - Incidence of overuse syndromes of the upper limb in young pianists and its correlation with hand size, hypermobility and playing habits. AB - The incidence of overuse syndromes of the elbow, wrist and hand was evaluated in a series of 66 pianists: 28 presented such a pathology. No significant correlation with starting age, or duration and intensity of practice was revealed. Smaller hands lead to more frequent overuse syndromes, in both males and females. Conditioning, sports activity, playing a second or third instrument, warming-up before playing and stretching afterwards did not influence the incidence of overuse syndromes. Pianists were not more hypermobile than a sex- and age-matched control group, nor was there any significant difference in hypermobility between pianists with and without overuse problems. There was no significant difference in incidence between the left and right side. PMID- 10855300 TI - Biomechanical evaluation of the elbow following radial head fracture. Comparison of open reduction and internal fixation vs. excision, silastic replacement, and non-operative management. AB - Thirty-six fractures of the radial head were evaluated following treatment with four different techniques: open reduction and internal fixation (ORIF), excision, silastic replacement, and non-operative management. Clinical assessment included calculation of the Clinical Performance Index based on parameters for motion, stability, pain, and daily function, as well as radiographic evaluation for fracture type (Mason) and the development of degenerative changes. Further assessment included isometric strength measurements for elbow flexion, extension, pronation, supination, and grip. Range of motion and strength measurements were similar, except for grip strength, which was significantly better for ORIF patients compared to excision or silastic replacement groups. Patients with ORIF had the best clinical scores with minimal radiographic changes. When possible, ORIF of displaced radial head fractures appears to provide the best functional result. PMID- 10855301 TI - [Distal traumatic avulsion of the triceps brachii. Apropos of a treated cases]. AB - Traumatic distal avulsion of triceps brachii is a rare injury, usually due to a fall on the outstre ched elbow, with sudden contraction of the triceps muscle. Examination reveals pain, swelling, and a palpable depression just proximal to the olecranon. Lateral X rays show avulsed osseous material. Surgical therapy was advisable for complete avulsion. In this case, the authors used absorbable sutures for this reinsertion. Immediate surgical repair always achieves good results. PMID- 10855302 TI - Extensor digiti minimi tendon "rerouting" transfer in permanent abduction of the little finger. AB - Permanent abduction of the little finger is a bothersome deformity which usually occurs in the context of sequelae of ulnar nerve palsy (Wartenberg's sign), but also in rheumatoid arthritis. The authors report an original technique for correction of this deformity. The extensor digiti minimi tendon is sectioned at its distal insertion and transferred in the wrist through the extensor retinaculum. The "rerouted" tendon is finally resutured distally on the radial aspect of the interosseous muscle. Side-to-side suture of the transferred tendon to the extensor digitorum tendon of the little finger further reinforces the solidity of the procedure. The distal insertion of the extensor digiti minimi tendon is consequently radialized. Its new direction eliminates the abduction component, and the tendon then behaves as an active adductor of the little finger. Five cases (2 cases of ulnar nerve palsy, 3 cases of rheumatoid arthritis) are reported with a mean follow-up of 19 months. All patients have complete active adduction of the little finger in extension, with a persistent capacity for abduction. The other correction techniques published in the literature are discussed. PMID- 10855303 TI - [Compression of the ulnar (cubital) nerve at the wrist by a supernumerary muscle]. AB - The authors report a case of anomaly of the flexor digiti minimi muscle, which extended into the forearm to be inserted 10 centimetres proximal to the carpal flexion crease on flexor carpi radialis. This anomaly was responsible for ulnar nerve compression when grasping objects with the hand. Cases of ulnar nerve compression at the wrist appear to be rare and the great majority of cases are secondary. Anatomical variants of muscles and nerves at the wrist are not exceptional, hence the importance of systematically looking for such anomalies in patients with ulnar nerve compression at the wrist. Excision of the muscle anomaly eliminated nerve compression and the associated symptoms. PMID- 10855304 TI - [Metacarpophalangeal dislocation of the index finger. Apropos of 7 operated cases]. AB - Complex dislocation of the metacarpophalangeal joint of the index finger is an uncommon injury. The authors report seven cases, including three neglected dislocations, two with associated osteochondral fractures and one with sesamoid entrapment. Open reduction was necessary in each of the seven cases. In recent dislocations (four cases), both approaches were successful in obtaining reduction and normal finger movements were obtained. The dorsal approach was simple and safe, while the palmar approach was difficult and had many disadvantages. In the case with sesamoid entrapment, the sesamoid had to be removed to achieve reduction. In neglected dislocations (three cases), the dorsal approach was successful in one case (three weeks), two incisions were necessary in one case (seven months), and one case was treated by Swanson prothesis (two years). None of these patients regained normal mobility postoperatively. PMID- 10855305 TI - [The radio-ulnar unit. Its functional importance in fractures of both bones of the forearm]. AB - The forearm has been composed of two bones since our very old ancestor Ichthyostega, which settled on dry land 300 million years ago. The fish fins, transformed into legs, already comprised two bones, which only became useful much later. This "frame", composed of 2 bones rotating around a longitudinal axis, is the most "practical" mechanical solution to allow longitudinal rotation of the distal extremity, but this rectangle must be pliable in a diagonal plane by means of the two conjoint radioulnar joints and a flexible hinge, the interosseous membrane. This system can only function when certain conditions are respected: strict alignment of the axes and simultaneous functioning of the two joints, maintenance of the "coordinated" shape of the two bones, an intact system of union of the two bones and coaptation of the joints. These anatomical and functional devices may be compromised by many pathological conditions, which is why the pronation-supination function of the forearm is so important and so vulnerable. PMID- 10855306 TI - Neurotization of the biceps muscle by end-to-side neurorraphy between ulnar and musculocutaneous nerves. A series of five cases. AB - Three patients with avulsed C5, C6, and C7 roots and two patients with avulsed C5 and C6 roots after trauma of the brachial plexus, were treated by neurotization of the biceps using nerve fibers derived from the ulnar nerve and obtained by end to-side neurorraphy between the ulnar and musculocutaneous nerves. The age of patients ranged from 19 to 45. The interval between the accident and surgery was 2 to 13 months. Return of biceps contraction was observed 4 to 6 months after surgery. Four patients recovered grade 4 elbow flexion. One 45-year-old patient did not obtain any biceps contraction after 9 months. PMID- 10855307 TI - Results of excision of heterotopic new bone around the elbow in patients with head injuries. A series of 25 cases. AB - Twenty patients were operated upon for heterotopic ossification around the elbow of neurogenic origin following intra-cranial trauma between 1993 and 1997. They did not receive any diphosphonates or radiotherapy. Pre-operative evaluation included a CT scan with enhancement using a dye injected intravenously and 3D reconstruction. Surgery was indicated in the presence of a clinical deficit in mobility or signs (clinical or electrical) of nerve compression. The average delay between the accident and the surgery was 34 months (5 months to 9 years). Associated procedure included lengthening of the brachialis (4 cases), lengthening of the triceps tendon (4 cases) and an anterior capsulotomy in 3 cases. 24 elbows were reviewed with an average follow-up period of 18 months (6 months to 4 years). In 58% of the cases, the result was very good (gain in mobility > 70%) while in the remaining 42% of cases, the improvement was between 40% and 70%. This study confirms the possibility of obtaining good results by excision of the masses of heterotopic ossification of neurogenic origin around the elbow before 1 year after the accident without the necessity of complementary treatment. PMID- 10855308 TI - Synovial osteochondromatosis at the elbow producing ulnar and median nerve palsy. Case report and review of the literature. AB - The authors present the case of a 53-year-old woman suffering from synovial osteochondromatosis of her right elbow responsible for ulnar and median nerve entrapment neuropathy. This condition is characterised by the formation of multiple cartilaginous nodules in the metaplastic synovium of otherwise normal joints, bursae or tendon sheaths. Treatment consisted of partial synovectomy, removal of loose bodies and microscopic nerve release. Synovial osteochondromatosis complicated by nerve compression syndromes has been rarely reported, usually with ulnar tunnel syndrome at the elbow. The literature on this subject is reviewed. PMID- 10855309 TI - Complications and outcome in open carpal tunnel release. A 6-year follow-up in 92 patients. AB - In this retrospective study, 92 patients (83% attendance rate) were examined 6 years after open carpal tunnel release. A questionnaire was answered by the patient and a physical examination was made by an independent observer. Five complications were found, of which 4 were early postoperative problems and one was a major complication with long-term disability. In one third of the patients numerous complaints were noted even after six years. Recurrences were found in 9 cases. No lacerations of nerves, tendons or vessels were seen and no patient developed reflex sympathetic dystrophy. The overall subjective outcome showed that 91% of the patients were free of symptoms or improved. The grip strength at follow-up was reduced, if the symptoms had been present for more than 12 months, as compared to less than 12 months, (p = 0.009) and when associated, unspecific brachialgia had been present (p = 0.02). No differences were found in conjunction with the operating surgeon being either an orthopaedic resident or a specialist in hand surgery. In conclusion, open carpal tunnel release had a subjectively favourable outcome, but due to complications and postoperative complaints, further investigations into alternative techniques seem necessary. PMID- 10855310 TI - Cross-chest radial nerve transfer in brachial plexus injuries. Experimental and anatomical basis. AB - Brachial plexus avulsion injuries are devastating injuries to the upper limb, and nerve transfer remains the only option in reconstruction. Despite the encouraging results concerning recovery of shoulder and elbow function, no option is available for treatment of the paralytic hand. In rats, we sectioned the radial nerve in the elbow region and transferred it across the chest to reinnervate the lesioned contralateral medial cord of the brachial plexus. Rats were then evaluated for motor and sensory recovery, electrophysiologically, behaviorally and morphologically. Forepaw functional recovery was estimated to be 90%. In cadavers, the radial nerve and profunda brachii artery were dissected. It was observed that the radial nerve vascularized by the profunda brachii artery was able to reach the contralateral brachial plexus distal to the shoulder region without nerve grafts. After sectioning the radial nerve, sensory loss is minimal and motor palsy can be easily restored by tendon transfers. The results of tendon transfer for radial nerve palsy are better than for any other nerve. Cross-chest radial nerve transfer might be of clinical interest in the reconstruction of hand function in entire injury to the brachial plexus. PMID- 10855311 TI - Are there still indications for the Krukenberg kineplasty? Report of two patients. AB - Krukenberg's operation was actually described by Vanghetti in 1989. Three cases, including one bilateral case, are reported. The first case was a 36-year-old patient from a developing country. At the sixth postoperative week, the patient had regained his independence and gained 1.5 kg. The second case was a child with multiple malformations, who presented, in addition to amputation of both legs, a high amputation of the left forearm and distal fold of the right forearm. Krukenberg's operation was performed at the age of 8 months. At the age of 2 years, the child was able to eat and drink independently. PMID- 10855312 TI - [Old Monteggia lesions in children. Apropos of 14 cases]. AB - Fourteen patients between the ages of 3.5 years and 11 years, with an old Monteggia lesion were managed in the Orthopaedics department between 1968 and 1997. Dislocation of the radial head was not diagnosed at the time of the initial trauma in 9 out of 14 cases and the diagnosis was made an average of 14 months after the trauma (range: 5 weeks to 84 months). Twelve of the 14 patients underwent surgical reduction of chronic dislocation of the radial head, mostly associated with proximal ulnar osteotomy and/or a procedure on the extensor retinaculum. The mean follow-up is 4 years 9 months. There were 66% excellent and good results, 25% of moderate results and 9% of poor results. Due to the difficulty of surgery of neglected lesions, the authors report their experience and emphasize the importance of the initial diagnosis. PMID- 10855313 TI - [An unusual complication of reimplantation of the hand: a false aneurysm of the radial artery anastomosis. Apropos of a treated case]. AB - The authors report a case of arterial pseudoaneurysm occurring 3 months after reimplantation of the hand in contact with the radial anastomosis. This complication, related to the initial mechanism of avulsion, although nonspecific, required rapid surgical revision as only one patent artery was present. PMID- 10855314 TI - [Finger replantation after 60 years of age. Apropos of 7 cases]. AB - The authors analysed retrospectively 7 cases of digital replantation in 7 men aged from 60 to 71 years, performed between 1985 and 1996. There were 2 amputations of the thumb, 1 of the index, 2 of the middle finger, 1 of the fourth and 1 of the fifth finger. 4 failures of replantation were noted. These 4 failures always concerned amputations of long digits by a circular saw with associated complex multidigital injuries of bad prognosis and in combination with a poor vascular status. We had 3 successful results: the 2 amputations of the thumb and the ring finger of the auricular. All these 3 patients recovered a good hand function. We found some common characteristics in this group of patients: excellent general condition, non smoker, good motivation and cooperation, injury of one digit, clear amputation (except the ring finger), correct conservation of the amputated part. The advanced patient's age does not represent a contraindication for digital replantation. The injury mechanism and the general condition of the patient represent major criteria of prognosis. In favourable circumstances, a good functional result can be expected. PMID- 10855315 TI - [Necrotizing fasciitis of the upper limb. Apropos of 4 cases]. AB - This article is based on the retrospective study of 4 cases of necrotic fasciitis of the upper extremity, in adult patients with a mean age of 57 years (range: 36 to 78 years) and with a male predominance (3 M/1 F). Presenting signs were variable: pain, febrile and inflammatory oedema, ecchymoses with inflammatory masses containing clear or haemorrhagic fluid. Treatment with antibiotics and anti-inflammatory drugs did not prevent progression to painless, necrotic ulcers. Rapid medical and surgical treatment constitutes an element essential of the prognosis and must include wide large debridement, antibiotics and intensive care. PMID- 10855316 TI - Evolution of our indications for neurotization. Our concept of functional restoration of the upper limb after brachial plexus injuries. PMID- 10855317 TI - Restoration of sensation over the contact surfaces of the thumb-index pinch grip using the terminal branches of the superficial branch of the radial nerve. AB - In huge median nerve losses and in some brachial plexus lesions, absence of sensation over the pulps of the index finger and the thumb preclude their use without visual control. Currently, end-to-side anastomosis is a new option available (when the ulnar nerve is intact) but we have reviewed the results of 7 cases of nerve anastomosis between the sensory branches of the radial nerve and the collateral nerves of the thumb (ulnar) and index finger (radial). Palmar translocation of the donor nerve, as classically performed, was used in two cases and the technique was subsequently modified to provide a better nerve suture by dorsal transfer of the collateral nerves of the thumb and index. Two sequellae of brachial plexus lesions and 5 cases of extensive defects of the median nerve were reviewed at a mean follow up of 5 years. With the classical technique the two point discrimination was 15 mm in one case and more in the other; with the modified technique, 4 patients achieved a thumb discriminaTion of 9 mm, 12 mm (2 cases) and 13 mm. PMID- 10855318 TI - Fractures of the base of the first metacarpal in children. Role of K-wire stabilisation. AB - In order to define the factors of instability of fractures of the base of the first metacarpal in children, the authors reviewed 30 children presenting this lesion with a follow-up greater than 10 months. Patients in whom the growth cartilage of the base of the first metacarpal was still open and presenting a fracture with angular displacement greater than 30 degrees or metaphyso epiphyseal sliding greater than 1 mm were included. Three groups were defined on the basis of radiographic findings: Group A: pure metaphyseal fractures (14 cases: 10 pinnings and 4 orthopaedic treatments); Group B: Salter II epiphyseal detachment fractures with a medial metaphyso-epiphyseal corner (10 cases: 1 pinning and 9 orthopaedic treatments); group C: Salter II epiphyseal detachment fractures with a lateral metaphyso-epiphyseal corner (6 cases: 2 pinnings and 4 orthopaedic treatments). The authors studied early secondary displacements as a function of the emergency treatment modality. No secondary displacement was observed for group B lesions regardless of treatment and for lesions stabilized immediately by intermetacarpal pinning. In contrast, one half of group A and C lesions treated orthopaedically subsequently became displaced, requiring surgical revision with stabilization by pinning. The authors recommend orthopaedic treatment for group B lesions and immediate surgical stabilization for group A and C lesions. PMID- 10855319 TI - [Reconstructive surgery of Blauth type III hypoplasia of the thumb]. AB - Thumb hypoplasia type III according to Blauth remains a rare congenital malformation. Recently Manske has promoted reconstruction versus pollicization in the sub-type IIIA where a first carpometacarpal joint is present. However we felt that pollicization is the solution for sub-type IIB where the basal joint is absent. We have reviewed 14 cases of thumb hypoplasia type III, four of them being type IIIB. After performing a first step with a free vascularized second metatarso-phalangeal joint transfer, the secondary steps were identical in both sub-groups. After a mean follow up of five years, no great difference was found in the two sub-groups and basal stability was even better in type IIIB. However the results were functionally and cosmetically inferior to the ones observed after pollicization. When the relatives refuse pollicization or the patient consults late for functional improvement, reconstruction remains worthwhile. PMID- 10855320 TI - Fracture of the distal part of the radius associated with severed ulnar nerve. AB - We report a case of a severed ulnar nerve after fracture of the distal part of the radius. The most likely hypothesis is stretching of the ulnar nerve fixed by Guyon's canal and severed on the sharp edge of the proximal radius. Although very rare, this lesion must be investigated particularly in cases with marked displacement, especially ulnar and/or volar. PMID- 10855321 TI - Avascular necrosis of multiple carpal bones. A case report. AB - A case of a 66-year-old female patient with hyperlipaemia, corticosteroid osteoporosis and chronic obstructive lung disease with avascular necrosis of the proximal row of the carpus and hamate is described. No other sites of avascular bone necrosis were found. A proximal row carpectomy was performed with an excellent outcome. PMID- 10855322 TI - Trapezo-metacarpal and metacarpo-phalangeal dislocation of the thumb associated with a carpo-metacarpal dislocation of the four fingers. AB - The authors report a case of combined dorsal fracture-dislocations of all 4 fingers, palmar trapezo-metacarpal dislocation and metacarpophalangeal dislocation of the thumb following a motorbike accident. These exceptional lesions were treated as an emergency by reduction and pinning. With a follow-up of 13 years, the patient still worked as an electrician. PMID- 10855323 TI - [Treatment of sections of the long flexor tendon of the thumb using the Rouhier technic. Apropos of 25 cases]. AB - We treated 28 cases of Flexor Pollicis Longus tendon injury by Rouhier's technique between 1989 and 1996 and reviewed 25 cases, with a minimum follow-up of 6 months and maximum of 8 years. Nine patients presented an associated collateral nerve section. We used the International Federation of Hand Surgery Societies (IFSSH) topographic classification. The surgical technique consisted of Flexor Pollicis Longus lengthening and transosseous pull-out in 16 cases and distal suture in 9. The immobilization time ranged from 4 to 7 weeks. The results were evaluated according to Tubiana's classification, with 2 very good, 14 good, 9 fair and no poor results. Flexor Pollicis Longus simple suture in T2 zone may produce suture blockade, tendon shortening and adherences. Suture through the digital canal must be avoided to decrease these complications. In the absence of bone injuries, better functional results can be obtained with Rouhier's technique than with simple suture. PMID- 10855324 TI - [Angiomyoma of the hand: a post-traumatic tumor?]. AB - A 69-year-old woman presented with a tumor of the lateral aspect of the proximal phalanx of the little finger of the right hand. The finger was injured by a ring during a handshake. Excision and histopathological examination revealed the diagnosis of angio-myoma, a benign vascular tumor originating from the smooth muscle cells of arterial or venous walls. Angiomyomas belong to the family of leiomyomas and rare, small tumors (less than one centimeter), preferentially occurring in women between the ages of 40 and 60 years. They may be painful. Only one case of sarcomatous degeneration has been described in the literature. Trauma has never been previously reported as a cause of angio-myoma, but in this case the traumatic origin of the tumor was not in doubt. PMID- 10855325 TI - [Anterior transbrachial approach of the coronoid apophysis]. AB - Fractures of the coronoid process of the ulna can cause elbow instability. Treatment of these fractures, sometimes surgical, raises a problem of the incision. The incisions described to date do not provide specific exposure, or require sometimes dangerous nerve and vascular dissections. The authors propose a strictly brachial midline anterior incision. The biceps tendon is retracted laterally and the brachialis muscle is then dissociated longitudinally providing direct exposure of the coronoid process, with no risk of nerve lesions, and allowing direct screwing of the fracture. PMID- 10855326 TI - Tenorrhaphy and tendinous venous envelope (TVE). A morphofunctional study in 24 beagles dogs. AB - A morphofunctional study was undertaken in 24 Beagle dogs subjected to a a tenorrhaphy, in order to evaluate the results obtained with 3 different surgical procedures (16 hands each)--"traditional" (TRAD); the "tendinous venous envelope" (TVE), and the "simulated procedure" (SHAM). The surgical tissue (n = 48) was the extensor digitorum communis muscle of the hand (forepaw) bilaterally. For TRAD procedure, the Bunnell suture technique was applied. For the TVE procedure, the tenorrhaphy site was wrapped in a segment of autologous femoral vein and for the SHAM procedure, the tendon was exposed over 3 cm without sectioning it and the wound was closed. In each case, the limb was immobilised for 21 days in a cast. Functional data (288 X-rays) was obtained on angle variations of the radio-carpal joint in 2 different positions--passive (physiological), and forced (with an additional weight of 150 g). For each position, the angle variation obtained for each one of the surgical procedures were compared at 3 different times: day zero (surgical intervention), day 21 (cast removal), and day 35 (surgical tissue removal). Morphological data (144 sections) was obtained by analyzing the surgical tissue (tendinous fibrosis; fusion of the tendon clusters with the epimysium and the neighboring tissues; synovitis and the virtual space between the tendon and the neosheath). Tests of variance analysis by posts (critical H = 5.99), showed that the TVE procedure produced better results as compared to TRAD (SHAM approximately equal to TVE > TRAD) on day 21 (H = 22.58) as well as on day 35 (H = 8.08). PMID- 10855327 TI - Morphometric study of the upper intercostal nerves: practical application for neurotizations in traumatic brachial plexus palsies. AB - The aim of the study was a morphometric evaluation of the intercostal nerves at different levels along their course in order to determine their adequacy in neurotizing the recipient nerves. The intercostal nerves were harvested from 5 cadavers. A biopsy of the nerve was obtained at 2 levels for each nerve in the parasternal region and at the level of the mid-axillary line. The musculocutaneous nerve was isolated at its origin from the lateral cord. Each harvested specimen was embedded in paraffin and sections were made using a microtome. These sections were then stained histochemically using HPS (Hematein, Phloxine, Safran). Real-time digitalisation of the video image under the microscope was performed. The sum of the different fascicular zones is the effective sensorimotor surface of the nerve at the level being studied. RESULTS: Direct suture of the upper three intercostal nerves to the musculocutaneous nerve is always possible upto the axillary fossa. The sixth intercostal nerve can be delivered upto this level in only 50% of cases without dissection of the musculocutaneous nerve upto its entry into the coracobrachialis. The musculocutaneous nerve presents a mean surface area of 2.64 mm2 while the nerve to the biceps has a mean surface area of 0.34 mm2 i.e. a ration of 1/8. The mean surface area of the intercostal nerves at the parasternal level is 0.23 mm2 while that at the axillary level is 0.34 mm2. Thus a loss of 33% in surface area occurs between the axillary and the parasternal levels. Our study confirms the insufficiency between the surface area of the intercostal nerves and the different nerve trunks to be neurotized. The relationship between the surface area of the musculocutaneous nerve and the three intercostal nerves is 26.72% with a minimum of 17.2%. If a fourth intercostal nerve is added, this ratio nerves appears to be a superior technique. We were able to deliver the sixth intercostal nerve for a direct suture to the musculocutaneous nerve in only half the cases. PMID- 10855328 TI - A biomechanical study of Tang's multiple locking techniques for flexor tendon repair. AB - This study was designed to biomechanically compare Tang's multiple looped locking techniques with various suture techniques for flexor tendon repair in the hand. Fifty flexor digitorum profondus tendons taken from pig toes were used as models; The tendons were transected in the middle part of zone 2 defined as the area beneath bifurcation of the flexor digitorum superficialis tendons, and were repaired by five different suture methods: (1) modified Kessler, (2) Tsuge's suture, (3) double Kessler, (4) modified Kessler plus Tsuge, and (5) Tang's suture. The repaired tendons were placed in an Instron tensile testing machine to determine the tensile properties of the repair. 2 mm gap formation force and ultimate tensile strength were measured during the test. Maximal work to failure were calculated according to area under the load-displacement curve of the test. 2 mm gap formation force was 21.5 N for the Kessler, 20.6 N for the Tsuge, 31.6 N for double Kessler, 30.9 N for the Kessler plus Tsuge and 41.4 N for the Tang. Ultimate tensile strength was 23.5 N for the Kessler, 22.9 N for the Tsuge, 34.5 N for the Kessler plus Tsuge and 45.6 N for the Tang. Statistically, Tang's suture had the greatest gap formation force, ultimate strength and energy for failure among the five techniques (p < 0.01 or p < 0.001). Gap formation force, ultimate strength and energy to failure for double Kessler or the Kessler plus Tsuge were significantly greater than those for the Kessler or the Tsuge (p < 0.05 or < 0.01). The tendons repaired by Tang's method tolerated a significantly higher tensile load (133 to 198% of the other techniques) than the other methods. Among the methods tested, Tang's multiple looped locking suture provides sufficient gap resistance and tensile strength that may be able to withstand early active mobilization after primary flexor tendon repair. PMID- 10855329 TI - Hyperostotic macrodactyly and lipofibromatous hamartoma of the median nerve associated with carpal tunnel syndrome. AB - A new case with 14-year follow-up of an extremely rare variety of congenital hand macrodactyly is presented. The disease characteristically presents a diffuse proliferation of fibrofatty tissue, but in this special type, osteocartilaginous deposits around the joints can also be found. The case presented included the troublesome feature of a lipofibromatous hamartoma in the median nerve at the wrist and its branches producing carpal tunnel syndrome. The patient obtained benefit from carpal tunnel release and epineurolysis. The hyperostotic development was managed with conservative resection of the periarticular osteochondromas. The literature reviewed suggests that the hyperostotic cases of macrodactyly do not differ from general cases of this congenital condition, except for the osteochondral deposits. These tumours develop during adulthood or after previous trauma, before epiphyseal closure. PMID- 10855330 TI - [Pure posterior luxation of the elbow in adults: immobilization or early mobilization. A randomized prospective study of 50 cases]. AB - INTRODUCTION: Pure posterior dislocation of the elbow is frequent in young subjects. The objective of treatment must be to reduce the dislocation and avoid complications, the most frequent being stiffness, but also elbow instability. The objective of this prospective study was to evaluate the functional and anatomical characteristics of two treatment modalities: plaster immobilization and early mobilization. MATERIAL AND METHODS: 50 cases of pure posterior dislocation of the elbow were included in a prospective study and randomized to two groups: Group I: twenty six cases were treated by reduction under general anaesthesia and plaster immobilization for three weeks, followed by rehabilitation. Group II: twenty four cases were treated by reduction under general anaesthesia, followed by early mobilization. RESULTS: We evaluated our results in terms of loss of amplitude of elbow movement (particularly extension), stiffness, instability, relapses, pain and ossification. This study demonstrated better recovery of elbow function in patients treated by early mobilization: 96% of good results with recovery of normal extension in group II versus 81% of cases in group I. Stiffness was observed in 19% of patients in group I versus 4% in group II; this difference was very significant. Comparison of pain revealed no significant difference and no relapses, instability or ossifications were observed in either of the two groups. DISCUSSION AND CONCLUSION: Early mobilization is superior to plaster immobilization, as it allows recovery of better quality elbow function without inducing instability or recurrence. PMID- 10855331 TI - [Perineural fibrosis of the median nerve at the wrist. Treatment by neurolysis and dermal-hypodermal graft]. AB - In the treatment of multiple recurrences of carpal tunnel syndrome due to fibrosis, following repeated nerve release, the authors propose interposition of a composite dermal fat graft raised from the inguinal region, and report four clinical cases using this technique. The objective of this study was to evaluate the efficacy of the graft in terms of clinical and electromyographic results and determine the course of the graft by a postoperative MRI study and a longer follow-up. CLINICAL RESULTS: Nocturnal paraesthesiae resolved in 2 patients, was improved in one patient and remained unchanged in another patient. Objectively, neurological examination showed improvement in 3 cases and no change in 1 case. Electromyographic results: At last follow-up, EMG was improved in only one case. Magnetic resonance imaging: MRI visualized the graft with a fat signal on T1 weighted sequences in every case, with an increase in size over time in 3 cases. The efficacy of the dermal fat graft may seem disappointing, as none of the patients were cured. However, with a mean follow-up of more than two years, we have not observed any deterioration of the clinical features, and no surgical revision for carpal tunnel syndrome. This technique appears to increase graft survival, as the blood supply to the subdermis is restored more rapidly via the dermis, which is anatomically a very vascular tissue. There is nevertheless a discrepancy between the clinical results, and EMG and MRI findings, which could be explained by the anatomical lesion of the median nerve, surgically released several times, and by alterations of the perineurium. PMID- 10855332 TI - [Rupture in the palm of a flexor tendon in a young man with osteogenesis imperfecta]. AB - Flexor tendon rupture in the palm of the hand is very infrequent. It usually occurs in association with an underlying pathological condition. We report a case of hand tendon rupture in a patient with osteogenesis imperfecta (OI). This disease is typically characterised by brittle bones but other tissues rich in collagen, such as tendons, may also be involved. The ruptured tendon was diagnosed one month post-injury and was treated by tendon graft. Six months later, tenolysis was performed and one year post-injury flexion was still limited at the distal interphalangeal joint. Few cases of tendon lesions and OI have been reported in the literature. Rupture occurs at the bony insertion and the outcome is good the poor outcome in our case could be due to alteration of the collagen tissue of the tendon. PMID- 10855333 TI - [Ulnar clasp failure or again "the nail file sign", a little known sign of disorders of the ulnar nerve]. AB - Besides the well known signs of the ulnar palsy, there is a new sign, unknown till now: the "failing ulnar hook". It could be named the "nail file sign", because it was discovered from a woman who could not file the nail of her little finger. Practically, the patient is asked, in a first time, to roll his little and ring fingers around the index of the same hand of the examinator. As the patient is resisting, the other index of the examinator tries to extend these two fingers. If they cannot resist, there is an ulnar palsy at an upper level. The explanation is that the flexor profundus of the ring and the little fingers are commanded by the ulnar nerf, whereas those of the index and the middle finger are by the median nerve. So, when this sign is present, the block is located from the elbow to the brachial plexus. When it is absent, whereas other signs of ulnar palsy are present, it evoques a lower palsy. This sign is of great interest because it confirms the diagnosis of ulnar palsy and allows the localization of the block. It is very useful to search it in every neurologic examination of the hand. In a carpal tunnel syndrome, it allows to eliminate or not an ulnar participation without using of electrical tests. PMID- 10855334 TI - [A correction technic using an osteotomy-graft in chronic impaction of the radial socket, called "die punch"]. AB - The central crush of the radial glena, so-called "Die-Punch", may be relatively easy to fix, when it is initially recognized. Unfortunately, it is often ignored, which compromises the future of the radio-carpal joint. When this "die-punch" is at the stage of malunion, the cartilage depression is very difficult to reach to. It is the goal of this described technique: through an antero-lateral way, making a sagittal osteotomy guided on a K-wire, aiming at the lateral limit of the crush; the lateral fragment comprising the radial styloide process is then turned like a door around a posterior hinge, opening the access to the medial bony cut, at the lower part of it, the depressed fragment is clearly visible. This fragment is lowered with a chisel of appropriate width, until it joins its proper level; then a fragment of cancellous bone taken in the upper part of the cut is crammed in the room above the "die-punch". The "door" is then closed and fixed with a screw, without any problem of consolidation, so as the rehabilitation may be initiated immediately. The practice of this procedure is till now limited to a few cases (two), but the results are very encouraging. This technique is worthy to be tried by other hand surgeons. PMID- 10855335 TI - [Anterior trans-scaphoid-lunate luxation of the wrist. Apropos of a case]. AB - About one uncommon case of anterior perilunate fracture-dislocation, the authors show the interest of posterior approach of the wrist to treat all the lesions. The scaphoid is screwing proximally to distally and a pin is used to stabilise the pyramidolunare articulation. The clinical results are good on motion, on strength; the lunate doesn't present necrosis on radiology at 4 years. PMID- 10855336 TI - [An uncommon occupational accident: tuberculous tenosynovitis of the extensor tendons of the hand]. AB - We present a case of tuberculous tenosynovitis of the extensor tendons of the hand. Our patient was a young doctor working in the respiratory medicine department. He was injured on the dorsal aspect of the hand with a needle used for pleural aspiration. The clinical features consisted of gradually swelling, mild pain and stiffness of the metacarpophalangeal joint. The diagnosis was made after synovectomy. Histological and bacteriological examinations revealed tuberculosis. Treatment consisted of synovectomy and appropriate antibiotics. The clinical course was excellent after one year follow-up. Tuberculous tenosynovitis of the hand is a rare manifestation of extrapulmonary tuberculosis occurring in fewer than 5% of all cases of skeletal tuberculosis. Thickening of the tendon or synovial sheath and local accumulation of fluid are the characteristic manifestations. The diagnosis must be confirmed by surgical biopsy. Antibiotics and synovectomy achieve a good functional result. PMID- 10855337 TI - [Is homeopathy superior to placebo? Controversy apropos of a meta-analysis of controlled studies]. AB - At a time when scientists support Evidence-Based Medicine, the Parliament of Belgium has recently decided to recognize four alternative medicines, among which homeopathy. Whereas this discipline does not rely on any scientific basis, it appears to be popular, especially in general practice. The homeopaths have recently taken arguments from a meta-analysis published in 1997 in the Lancet of 89 placebo-controlled trials. This study indeed concluded that the results are not compatible with the hypothesis that the clinical effects of homeopathy are completely due to placebo. However, this meta-analysis contains several methodological flaws. Furthermore, it is recognized that results of a meta analysis may not be confirmed in large well-performed clinical trials. Thus, homeopathy should still provide the evidence that conventional medicine has regularly brought during the last two decades in many fields of therapeutics, with the respect of the rigorous rules of "Good Clinical Practice", leading to "Evidence-Based Medicine". PMID- 10855338 TI - [Lupus erythematosus: from clinical aspects to immunopathology]. AB - The author highlights some of the clinical and serological signs associated with systemic lupus erythematosus and use them as keys to enter into the most recent advances in the immunopathology of the disease, in particular the mechanisms underlying the production of pathogenic autoantibodies. He stresses that a better knowledge of the pathophysiology of the disease, combined to the progresses in basic immunology and biotechnology, has raised hopes for a more targeted immunointervention. PMID- 10855339 TI - [Regulation of gonadal activity in the primate and in the human]. PMID- 10855340 TI - [20th and 21st century: evaluation of world health]. PMID- 10855341 TI - Cross-cultural differences in health-related quality of life of people with epilepsy: findings from a European study. AB - PURPOSE: To examine between-country differences in health-related quality of life (HRQOL) of adults with epilepsy across a large number of European countries. METHODS: Self-completion postal questionnaire sent to large sample of adults with epilepsy, recruited from epilepsy support groups or epilepsy outpatient clinics. The questionnaire was developed in English and translated. Back-translations from each language were checked for accuracy. The questionnaire sought information on clinical and socio-demographic details, and contained a number of previously validated scales of psychosocial well-being (the SF-36, the perceived impact of epilepsy scale, and a feelings of stigma scale). RESULTS: Controlling for socio demographic and clinical characteristics, significant between-country differences were found in scores on the perceived impact of epilepsy scale, on seven of the eight SF-36 domains, and on the feelings of stigma scale. Respondents in Spain and the Netherlands fared consistently better, whilst those in France fared poorest, compared to those in other countries in terms of the various HRQOL measures used. CONCLUSION: Several possible reasons for the cross-cultural differences in HRQOL are proposed. Clearly, there is no single explanation and there may also be reasons which we have overlooked. This study emphasises the need for further comprehensive research in order that the position of people with epilepsy in different countries be more thoroughly understood in the social context. PMID- 10855342 TI - Physical functioning and mental health in patients with chronic medical conditions. AB - There is evidence to suggest that a decline in physical functioning with advancing age is independent of mental health, which appears to remain relatively stable. There is additional evidence to suggest that those with a chronic disease also experience a decline in physical function while the mental health remains relatively stable. Using a cross-sectional design, data from the US population norms for the Medical Outcomes Study SF-36 are examined and compared to SF-36 data collected for four patient groups. Patient groups include kidney dialysis patients, multiple sclerosis patients, kidney transplant patients and patients with severe osteoarthritis of the hip prior to total hip replacement. Overall scores and scores within 10-year age groupings are examined in order to compare the physical functioning and mental health scores of the general population with those of the four patient groups. Results support the hypothesis that physical functioning declines with advancing age and with the development of chronic disease, but mental health remains remarkably stable regardless of chronic disease and/or advancing age. This observation suggests a process of psychological adjustment or adaptation to the physical difficulties encountered with advanced age or disability, and implies that this adjustment process may in fact be quite strong. PMID- 10855343 TI - The Illness Intrusiveness Rating Scale: a measure of severity in individuals with hyperhidrosis. AB - OBJECTIVE: We estimated the reliability and validity of the Illness Intrusiveness Ratings Scale (IIRS) in hyperhidrosis, using an electronic mail form of administration. METHODS: Recent contributors to an electronic mail discussion group on hyperhidrosis responded to the IIRS, questions about surgical history, items designed to assess severity, and demographic questions, on two occasions four weeks apart. A variety of hypotheses regarding the relationships between these variables were constructed a priori. RESULTS: Sixty-eight people replied on two occasions. Internal consistency was high (Cronbach's alpha 0.88), as was test retest reliability (kappa 0.89). The total IIRS score correlated with a global severity question (0.61; p < 0.001). Total IIRS score was lower in participants who had previously had surgery for hyperhidrosis, compared with those who had not (47 vs. 36; p = 0.02), and changed dramatically in the direction of diminished severity in four patients who underwent surgery during the course of the study (54 vs. 17; p = 0.01). Weak-to-moderate correlations were observed between total score and use of topical preparations, use of medications, number of clothing changes during a day, and limitations in choice of wardrobe. CONCLUSIONS: The IIRS is both reliable and valid in the assessment of patients with hyperhidrosis. A novel form of administration does not appear to affect its properties. PMID- 10855344 TI - The migraine work and productivity loss questionnaire: concepts and design. AB - Our objectives were to: (1) develop a self-report questionnaire for measuring the impact of migraine headache on work; and (2) qualitatively assess aspects of its performance. Two samples of migraine sufferers provided the data. Sample 1 (n = 18) participated in a structured discussion group designed to elicit examples of migraine's on-the-job impact. Sample 2 (n = 11) completed a mail survey and participated in in-depth phone interviews. Interviews addressed item comprehensibility, consistency of interpretation, the cognitive processes by which certain answers were generated and response burden. The participants were currently employed men and women, at least 18 years of age, who met the International Headache Society (IHS) criteria for migraine headache [1]. Discussion group participants indicated that migraine attacks substantially diminished their job performance. Pain, photophobia, phonophobia, mental impairment and fatigue were perceived as interfering with even routine or relatively simple job tasks. The Migraine Work and Productivity Loss Questionnaire, Version 1.0 (MWPLQ) was written. Next, it was assessed in the context of the in-depth interviews. Result indicated that the MWPLQ was comprehended without difficulty, interpreted consistently and easy to complete. Thus, qualitative results provide initial support for the new questionnaire. PMID- 10855345 TI - Health behaviors, social networks, and healthy aging: cross-sectional evidence from the Nurses' Health Study. AB - Physical function is a significant component of health-related quality of life among older adults. Potential correlates of healthy aging, including health behaviors and social network characteristics, were examined among 56,436 US women aged 55-72 in 1992. Healthy aging was assessed by maintenance of physical function measured by four subscales of the Medical Outcomes Study Short Form (SF) 36 Health Survey: physical functioning; role limitations; freedom from bodily pain; and vitality. Individual health behaviors, defined as current smoking, alcohol consumption, sedentary behavior, and being overweight each contributed to significant decrements in functioning across all age-groups. After controlling for these health behaviors and other confounders (age, race, education, and co morbid conditions), elements of a woman's social network were significantly correlated with functional status. Strong predictors of high functioning among older women were having close friends and relatives and presence of a confidant. For example, the absence of a confidant was associated with a 4.44 point reduction in physical functioning (95% CI: -7.0, -1.9), and a 5.68 point reduction in vitality (95% CI: -7.9, -3.4). These effects were comparable in magnitude to those observed among heavy smokers, or women in the highest category of body mass index. PMID- 10855346 TI - Quality of life and quality adjusted survival for breast cancer patients receiving adjuvant therapy. Eastern Cooperative Oncology Group (ECOG). AB - PURPOSE: The objective was to compare health related quality of life (QOL) in hormone receptor negative, node-positive breast cancer patients receiving adjuvant chemotherapy to determine whether a more intensive chemotherapy regimen has an adverse effect upon QOL that is not balanced by improvements in disease control or survival. Increased physical symptoms, including fatigue and the inconvenience of the dose intensive 16-week regimen, were expected to have a negative impact on QOL. DESIGN: QOL was measured in 163 patients, randomized to either a standard cyclophosphamide, doxorubicin and 5-flurouracil (CAF) or a 16 week multidrug regimen, using the Breast Chemotherapy Questionnaire (BCQ). The 30 item BCQ was self-administered prior to therapy, during therapy, and 4 months post treatment. RESULTS: BCQ scores decreased (worsened) more during therapy on the 16-week regimen, median change -1.4, than on CAF, median change -0.8 (p < 0.001). By 4 months post treatment, BCQ scores were higher than pre-treatment and equal in the two arms (CAF: 8.1 and 16 weeks: 8.2, p = 0.6). Over a period of 48 months, patients on the 16-week regimen averaged 1.4 fewer months of treatment with toxicity, 4.0 more months without symptoms and 0.7 fewer months post recurrence compared to patients on the CAF regimen. Given typical values for these health states, the gain in Q-TWiST observed for the CAF regimen during treatment shifted to the 16-week regimen after 1 year, with a gain of 2.0 to 2.4 months after 4 years. CONCLUSIONS: The hypothesized negative impact of the dose intensive 16-week regimen was confirmed by the BCQ assessments. However, Q-TWiST analysis suggests a small gain for the 16-week regimen. The later results should be interpreted with caution with the limited follow-up of 4 years. PMID- 10855347 TI - Using data from studies of health-related quality of life to describe clinical issues examples from a longitudinal study of patients with advanced stages of cervical cancer. AB - The focus of the paper is to describe how to present data from studies on health related quality of life (H-QoL) in a way that is simple and clinically relevant. Data from a longitudinal study of patients with advanced stages of cervix cancer are used. One hundred and eighteen patients filled out questionnaires (including EORTC QLQ-C30) 7 times over a period of 2 years. The following issues are considered: (1) The use of a panel for an initial overview of data. (2) The visual difference between using mean and median values. (3) Box-whisker plots to illustrate the variability of the data. (4) The effect of combining categorical data into fewer categories. (5) Individual patient profiles showing the wide variability among patients. (6) A table showing the change of scores over a one year period. (7) "Prognostic plots" dividing the initial scores and the following scores. (8) Plotting changes over time. (9) Illustration of the impact of non random dropout. (10) The effect of drop-out for the patients who fill out two sequential assessments. (11) The use of healthy controls to help answer the question "what is normal?". PMID- 10855348 TI - Feasibility of quality-of-life research on the Internet: a follow-up study. AB - OBJECTIVE: Assessment of the feasibility of conducting clinical research with patients using electronic mail and the World Wide Web. DESIGN: We re-contacted 463 patients with ulcerative colitis (UC) and 154 benign prostatic hyperplasia (BPH) patients who had provided us with e-mail addresses as part of a Web-based study. In an electronic mail message, we informed patients of a web site with a new study and invited them to participate. We then examined the factors associated with patients' participation in the new study. RESULTS: Completion rates were 28% for UC patients (single mailing) and 48% for BPH patients (up to four mailings in a two-month period). Some patients could not be contacted due to invalid e-mail addresses (23%). Those who completed the new study tended to be older, and less time had elapsed since their participation in the previous study. Furthermore, their health-related quality-of-life had significantly improved since the previous study. CONCLUSION: It is possible to use direct electronic mail contact to conduct follow-up research with patients. Response rates appear to be related to the number of messages sent, age of the recipients, and time since the initial contact. PMID- 10855349 TI - Use of relevancy ratings by target respondents to develop health-related quality of life measures: an example with African-American elderly. AB - BACKGROUND: Health-related quality of life (HRQOL) instruments assess functioning and well-being. Generic HRQOL measures are intended to be relevant to everyone whereas population-targeted measures are designed to be relevant to a particular population. METHODS: We asked 99 African-American elderly (mean age 72, 33% female, 47% less than high school education) to rate the relevancy of 33 HRQOL items drawn largely from existing instruments. We assessed the reliability of the relevancy ratings across respondents, rank-ordered the items by relevancy, and tested the significance of difference in relevancy ratings for each item compared to the average of all other items. We also examined the associations of the relevancy ratings with sociodemographic and clinical characteristics. RESULTS: The relevancy ratings were reliable (intraclass correlation = 0.71) and relevancy was generally distinct from HRQOL and demographic characteristics. Items assessing spirituality and weight-related health status were rated as significantly more relevant than other types of items. Generic HRQOL items were not rated as highly relevant. CONCLUSIONS: HRQOL measures assessing spirituality and weight-related concepts are important for future studies of HRQOL in African American elderly. The method of identifying these concepts used in this study should be valuable in developing new measures targeted to other sociodemographically or clinically defined subgroups. PMID- 10855350 TI - [Chronic active plasmacytic gastritis associated with cytomegalovirus]. AB - Chronic active plasmacytic gastritis (CAPG) is characterized by the presence of chronic inflammatory cell infiltrates, mainly formed by plasma cells, involving the neck of gastric glands. This lesion, as well as Menetrier disease, has been linked to cytomegalovirus (CMV). To test this association we evaluated the foveolar/glandular (F/G) index and the presence of CMV DNA (desoxirribonucleic acid) by means of polymerase chain reaction (PCR) in 12 cases of CAPG and 13 controls. Cases exhibiting CAPG included 2 with Menetrier disease, 6 with foveolar hyperplasia, and 3 with normal foveolar/glandular (F/G) index. None showed either lymphocytic gastritis or CMV inclusions. Three CAPG cases were associated with gastric carcinoma. The F/G index was less than 1 in all controls. Eleven out of the 12 cases with CAPG showed amplification for CMV DNA while all controls were negative. Findings suggest a very close association, probably in progressive stages, between CMV infection, CAPG, foveolar hyperplasia (with or without Menetrier disease) and gastric carcinoma. CAPG might be a histologic marker for CMV infection in the germinative zone of the neck of gastric glands. These findings resemble those of hepatitis B virus (HBV) infection, chronic hepatitis, cirrhosis, and hepatocellular carcinoma saga. PMID- 10855351 TI - [The time and pH at the moment of disappearance of pyrosis induced by of esophagus acid perfusion is an indicator of hyperalgesia]. AB - OBJECTIVES: 1) Describe and compare evolutionary features of the pyrosis symptom with a positive acid perfusion test in the entire study population, and divided by sex. 2) Describe the results of different tests according to sex (standard acid clearance, number of reflux episodes in a short testing period, period of time until the appearance of pyrosis during acid perfusion, period of time until its disappearance, and esophagus pH at the time). 3) Compare differences between control and patient cases regarding standard clearance and reflux episodes. 4) Establish a multivariance correlation between the results of the tests according to sex and historical data, looking for one or more regulating factors. 5) Establish the same descriptive and comparative assessments in both subgroups, with and without hiatal hernia (HH). STUDY POPULATION: 15 healthy subjects, 9 men and 6 women (control), and 50 patients with pyrosis, 23 men and 27 women, over 18 years old, with or without HH, matched for age and sex (p = NS), and consecutive. MATERIAL AND METHODS: Clinical data records, esophagogastroendoscopy with at least three biopsy samples, number of reflux episodes within 30 minutes, standard acid clearance, measurement of perfusion time, time it takes pyrosis to disappear, and esophagus pH at the time. This is a prospective, descriptive, comparative, experimental, longitudinal, single-blind study with control subjects. RESULTS: Eighty-two percent (82%) of the study population had pyrosis II. Average age: 39.9 +/- 13.3 years, with no differences between sexes (p = 0.31). Development period at study time: 5.7 +/- 5.5 years (p = 0.33). Fifty eight percent (58%) of patients showed endoscopic signs of esophagitis, and 52% had HH. The number of reflux episodes, and of deglutitions needed to reach a pH of up to 5 were statistically different between patients and control subjects (p = 0.0005). The time required by acid perfusion for pyrosis to recur was 3.67 +/- 3.26 minutes. The time until its disappearance was 2.2 +/- 1.78 minutes. pH at pyrosis spontaneous cessation was 2.10 +/- 0.83. There was a correlation between the age at the time the symptom appeared, and the time the induced pyrosis disappeared (p = 0.02), as well as between this point in time and its corresponding pH (p = 0.006, r = 0.45). The presence of HH was associated with the number of reflux episodes and the frequency of proven endoscopic injuries, but not with the other parameters. CONCLUSIONS: 1) Most patients complaining of pyrosis were middle-aged, but it might also appear in very old people. 2) An early history of pyrosis was slightly associated with a delay in suppression of induced pyrosis. 3) Longer duration of pyrosis at induced pyrosis test was associated with a higher pH pyrosis degree, a fact that points to an enhanced sensory perception. 4) An association was established between HH and abnormal findings in esophagoendoscopy/hystopathology, but not so with sensory parameters. Hyperalgesia and a low pH are not the only factors which determine pyrosis. The findings strongly support the hypothesis according to which pyrosis, gastroesophageal reflux, and endoscopically proven injuries are associated, but independent. PMID- 10855352 TI - [Gastrointestinal damage induced by celecoxib and rofecoxib in rats]. AB - Five experimental models were carried out in different groups of Wistar rats (n = 15) in order to study selective (cyclo-oxygenase) COX-2 non-steroid antiinflammatory inhibitors, such as celecoxib and rofecoxib, as follows: 1) Dose dependent oral celecoxib and rofecoxib for 5 days, and 24 hours after oral indomethacin. 2) Same as 1, but subcutaneously. 3) Gastric ulcer induced by means of glacial acetic acid. 4) Duodenal ulcer induced by means of cysteamine. 5) Stress due to being kept under restraint and immersion in water at 15 degrees C for 6 hours. Celecoxib and rofecoxib, either orally or subcutaneously, did not produce necrotic injuries in healthy gastrointestinal mucosa (0%), showing normal histology. On the other hand, the injuries previously induced by indomethacin worsened (90%, p < 0.001). Total necrosis of small intestine as well as increased ulcer and perforation of gastric and duodenal ulcers induced by acetic acid and cysteamine were observed. There was also worsening of gastric necrotic area with stress (60-90%, p < 0.05). Celecoxib and rofecoxib showed neutrophilia (5,000/mm3) similar to that presented by indomethacin, but there was no leukocyte infiltration in the gastric mucosa; thus we can consider it a COX-2 selective NSAID (non-steroidal anti-inflammatory drug). CONCLUSION: Dose-dependent administration of celecoxib and rofecoxib as COX-2 inhibitors and non-COX-1 inhibitors, respectively, did not produce toxic injuries on healthy gastrointestinal mucosa, thus providing a broad therapeutic spectre. On the other hand, when administered in presence of altered gastrointestinal mucosa, they worsened and complicated gastric ulcers, and also induced necrosis in the small intestine, thereby restricting their clinical use. PMID- 10855353 TI - Assessment of the Sydney System in Helicobacter pylori-associated gastritis in children. AB - OBJECTIVE: The aim of our study was to establish the usefulness of the Sydney System in grading H. pylori-associated chronic gastritis in biopsies from pediatric patients. STUDY DESIGN: Fifteen children (average age: 10.8 years) with histologically-proven gastritis were studied. Classification and grading of gastritis were performed according to the analogue visual scales described in the updated version of the Sydney System. A chart was specially designed to record the morphological grading in this study. Altogether, we studied 79 gastric biopsies. RESULTS: Neutrophilic infiltrates were absent in 27 biopsies, mild in 35, and moderate in 17. This feature was not marked in any of the biopsies. Mononuclear infiltrates were mild in 38 biopsies, moderate in 36, and marked in 5. Density of H. pylori was mild in 39 biopsies, moderate in 27, and marked in 2. In three post-treatment biopsies belonging to the same patient, no H. pylori was observed. Additional 8 biopsies (7 from the body and 1 from the antrum) showed no H. pylori, although organisms were simultaneously present in other stomach sites. Lymphoid follicles were present in 19/79 biopsies. Intestinal metaplasia was not seen on slides stained with hematoxylin-eosin (HE). However, the Alcian blue-PAS stain revealed isolated positive cells in 8 out of 15 patients. None of the gastric biopsies showed mucosal atrophy. CONCLUSION: The results show that the Sydney System is applicable for pediatric patients in case of H. pylori associated gastritis. However, the number of biopsies recommended in the Sydney System seems excessive for this age group. PMID- 10855354 TI - [Lymphangiectatic cysts of the intestine in autopsy]. AB - We report the postmortem findings of three patients with multiple pale polyps in the small bowell, which show the typical histological features of lymphangiectatic cysts. PMID- 10855355 TI - [Identification of Enterocytozoon bieneusi in a patient with sclerosing cholangitis and AIDS]. AB - Enterocytozoon bieneusi is the most common microsporidian parasite found in patients with AIDS. We report the clinical features of a patient with chronic diarrhea, pancreatitis, and AIDS-related sclerosing cholangitis. Ultrasonography and endoscopic retrograde cholangiopancreatography disclosed intrahepatic and extrahepatic bile duct changes identical to those seen in sclerosing cholangitis. Enterocytozoon bieneusi was found in duodenum and peripapillary duodenum by means of light microscopy, and confirmed by PCR amplification of paraffin-embedded tissues with species-specific primers. Microsporidian infection should be suspected in patients with advanced immunodeficiency and AIDS-related sclerosing cholangitis in our country. PMID- 10855356 TI - Primary mesothelioma. A practical tutorial review. AB - TARGET AUDIENCE: Gastroenterologists, internists, and surgeons LEARNING OBJECTIVES: After completion of this article, the reader will be able to understand the main diagnostic features of this disease and the differential diagnosis with other entities that have a more benign outcome. The lack of acceptable treatment is explained. PMID- 10855357 TI - [Natural history of gastroesophageal reflux at the dawn of the third millennium]. PMID- 10855358 TI - [Esophageal sensitivity: a silent challenge]. PMID- 10855359 TI - [Directions and destiny of internal medicine]. PMID- 10855360 TI - [Outcome of pregnancies in lupus: experience at one center]. AB - We determined the outcome of all pregnancies in SLE patients in our lupus cohort between 1991 and 1997. The women were advised that pregnancy was acceptable if the disease had been inactive for 6 months (SLEDAI < or = (4 at 2 serial examinations) and daily prednisone dose was below 10 mg. Patients were advised against pregnancy in case of active nephritis or neurolupus. In case of antiphospholipid antibodies, patients were treated with aspirin or heparin if previous fetal losses were documented. In case of anti-SSA ab, patients were monitored with ultrasound and given dexamethasone in case of atrioventricular block. Fifty-nine pregnancies were registered among 31 women: mean age at diagnosis of SLE was 25.3 +/- 3.7 years (range: 17-31); mean disease duration before pregnancy 4.4 +/- 3 years (0-14); mean ACR score 5.4 +/- 1.5 (4-9). Seven patients had ACL ab, 8 had anti-SSA ab. Pregnancies ended in: 13 early spontaneous abortions (9 not related to disease flare up, 4 related to SAPL); 7 elective abortions (patient decision in 5 cases, severe lupus flare up in 2); one in utero death; 19 full term births (> 38 weeks); and 19 preterm births. Cesarean section was performed in 11 cases (6 for fetal distress, dystocia and previous ceasarian; 5 for active lupus). Severe sepsis occurred in one premature infant who died at the age of 1 week. Intrauterine growth retardation was observed in 11 cases, mean APGAR score was 8.9 +/- 1.43. Child development was normal in all cases except one child with mild mental retardation. Severe lupus flare ups occurred in 6 cases, of which 4 were pregnancies in unadvised situations. Six mild flare ups were documented in the post partum. One fatal case of neonatal lupus with AVB was observed. In conclusion, in our experience, the live birth rate is similar to the general population and the risk of lupus flare up is low when the above mentioned criteria are applied. Systematic increase of steroid dose at pregnancy onset does not seem to be necessary. The high rate of prematurity remains a problem to be solved. PMID- 10855361 TI - [Gougerot-Sjogren syndrome disclosed by MALT lymphoma of the salivary glands. Report of 3 cases]. AB - Sjogren's syndrome is characterized by an increased risk of developing non Hodgkin's lymphoma. The lymphoma is most frequently extra-nodal, preferentially affecting the salivary gland: low-grade MALT lymphoma. Conversely, underlying Sjogren's syndrome has been recently identified by some authors in patients with non-Hodgkin's lymphoma. In the present report, we present three cases of Sjogren's syndrome disclosed by low-grade salivary gland MALT lymphoma. The patients were all women aged 33, 38 and 52 years. Extension work-up revealed nodal and bone marrow involvement in one case and no evidence of disseminated disease in the two others. Using the European criteria, all of our patients had certain Sjogren's syndrome. Labial salivary gland biopsy and immunopathological studies in newly diagnosed low-grade MALT lymphoma would be helpful in identifying the real frequency of this association. PMID- 10855362 TI - [Olfactory disorders in Alzheimer's disease and in Parkinson's disease. Review of the literature]. AB - Olfactory disorders in Alzheimer's disease and Parkinson's disease have been the topic of a large body of work over the last decades. Work devoted to olfactory disorders in Alzheimer's disease includes over 300 papers providing clinical and fundamental data. Anatomy studies in Alzheimer's disease have demonstrated a specific concentration of lesions in peripheral and central olfactory structures (senile plaques, neurofibrillary degeneration) as well as lesions in layers II and III of the entorhinal cortex. These neuropathological findings led to the development of the hypothesis that olfactory disorders in Alzheimer's disease would result from a toxic process. Observed olfactory deficits involve both identification and recognition of odors and detection thresholds. Nevertheless, patients with Alzheimer's disease rarely consult for sensorial deficits as the other signs of the disease predominate. Neuropathology data on the olfactory system are much more sparse in Parkinson's disease. Lewy bodies suggestive of Parkinson's disease have been observed in the olfactory bulb and pathways, but, unlike Alzheimer's disease, the olfactory disorders appear to be stable, changing little over time, as opposed to the evolution of neurological symptoms and cognition impairment. Clinicians should be aware that olfactory disorders are an integral part of Alzheimer's disease and Parkinson's disease. Screening for sensorial impairment however is a secondary objective in the context of these neurodegenerative diseases. PMID- 10855363 TI - [Graft safety and need satisfaction: searching for a crucial balance]. AB - In 1998, 3,033 organs were transplanted and 3,186 stem cell and 4,053 cornea grafts were performed in France. Organ shortage varies for each organ but is particularly critical for kidneys. Given the physiological and therapeutic efficacy of grafting, an increase in organ procurement is likely to raise safety problems. This paper concerns donor-transmitted infections, mainly viral infections. Regulations on health risks express a social and cultural refusal to accept any hazard. This collective concern is however in opposition with the interests of individual patients who are likely to accept a greater risk for an obvious benefit. Although the assessment of the risk/benefit ratio is clearly different between organ and tissue transplantation, further thought needs to be given to the problem of acceptance of infectious risk in organ transplantation. In France, special exceptions are now legally authorized for specific situations. Infectious agents transmitted by grafting have not been largely reported, mainly because screening tests were not available. The number of transmitted infections is increasing with the expansion of transport facilities and with the various geographical origins of the population. In some cases, search for histocompatibility between relatives living in endemic areas introduces new risks, for example tropical diseases. Diagnosis is often made by serological or molecular biological tests which may be difficult to analyze depending on their specificity. Organ transplantation is often an emergency requiring rapid decision making, for the donor or the recipient. However, with a quality assurance process based on accepted guidelines, the appropriate answers to operational problems can be given rapidly. Patient satisfaction can be achieved only if the public health challenge is met. Increasing organ procurement can clearly be expected to be beneficial, but at the cost of higher risks. Nevertheless, benefit and risk are quite different, benefit is for the time being while risk is delayed and may be uncertain depending on individual survival. Because of the variability of human disease, especially in situations where an ultimate therapy is involved, automatic application of a standard procedure must be avoided as it would inevitable lead to a cut-back in organ procurement in an attempt to achieve a theoretically safe situation which would be contrary to individual patient's self interest. Health care workers should have good knowledge of the epidemiology of these diseases in order to assess risk. The patient must be able to express informed consent. PMID- 10855364 TI - The use of G-CSF in normal neutrophil donors. AB - The efficacy of neutrophil transfusion therapy in the treatment of neutropenia associated infection is most likely dependent on the dose of neutrophils provided. Stimulation of neutrophil donors with G-CSF, with or without corticosteroids, causes marked donor neutrophilia and greatly increases the number of neutrophils collected by leukapheresis. Transfusion of these cells into severely neutropenic patients results in substantial increases in the patient's neutrophil count, levels which are sustained for 24 hours. These cells are capable of migrating to extravascular sites. These transfusion results are in marked contrast to those observed when fewer cells are administered and have led to great enthusiasm for this therapy on the part of many investigators. In spite of these preliminary encouraging results, the determination of clinical efficacy must await the results of large clinical trials. PMID- 10855365 TI - Indications of plasma exchanges in 2000. AB - In Hematology, Neurology, in renal diseases, and autoimmune diseases. PMID- 10855366 TI - [Intravenous immunoglobulins: anti-infection indications]. AB - Intravenous immunoglobulin (IVIg) are therapeutic preparations of intact IgG that are obtained from a pool of more than one thousand healthy blood donors and contain antibodies directed toward a large panel of microbial agents. IVIg contain high amounts of antibodies with opsonising activity and are indicated in substitutive treatment of patients with constitutive hypogammaglobulinemia. IVIg also contain antibodies able to neutralize certain bacterial toxins exerting superantigenic activity, and by this mechanism exert both anti-infectious and immunomodulating activity. During the last fifteen years, indications of IVIg in the prophylactic treatment of infections have been well defined: a) primary antibody deficiency; b) multiple myeloma and chronic lymphocytic leukemia with hypogammaglobulinemia and recurrent bacterial infections; c) bone marrow allograft; d) childhood AIDS; e) chicken pox seroprophylaxis in immunosuppressed patients and pregnant women. Therapeutic indications of IVIg in infectious diseases are limited to chronic parvovirus B19 infection associated or not with HIV infection. Other therapeutic indications are not well defined. PMID- 10855367 TI - [Unusual cause of chronic iron-deficiency anemia and insulin resistance: cholesterol embolism]. AB - BACKGROUND: Cholesterol embolism disease (CED) is an extremely polymorphous clinical entity. Rare reports mention digestive or endocrine manifestations. CASE REPORT: A 73 year-old female patient developed CED disclosed by iron deficiency anemia with chronic digestive bleeding associated with major insulin resistance. DISCUSSION: Solitary iron deficiency anemia as the inaugural sign of CED is exceptional. Rare reports in the literature suggest it is related to polymorphous nonspecific lesions (ulcers, pseudotumoral lesions...) leading to chronic bleeding. Careful histopathology study of the lesions of fragments of healthy mucosa provides the correct diagnosis showing biconcave and optically empty slits. Insulin resistance has not been reported although prior diabetes is often mentioned as a predisposing factor for atherosclerosis. In our case, the role of CED was not determined, but pancreatic or inflammatory involvement can be envisaged. PMID- 10855368 TI - [Spontaneous remission of cytomegalovirus retinitis in a patient infected with the human immunodeficiency virus]. AB - BACKGROUND: We report a case of CMV retinitis cured in an AIDS patient after two months of regular HAART therapy without CMV medications. CASE REPORT: A 37-year old patient (baseline CD4 count 47/ml) receiving HAART (2 NRTI and 1 IP) showed poor compliance and had increased his CD4 count for two months reaching 263/ml although HIV viral load remained high (71,840 copies/ml). His fundus was normal. He was evaluated 8 months after a period of loss to follow-up: his CD4 count was 65/ml, HIV viral load was 123,000 copies/ml, CMV serology for IgV and viruria were positive, viremia was negative, his fundus revealed a healed CMV retinitis. Six months later and without any CMV therapy, no relapse has been observed while regularly taking HAART medications. CONCLUSION: Few cases have been reported in which HAART therapy with good immune response and reduced HIV viral load lead to complete regression of CMV retinitis without specific CMV medication. This case suggests that, even with an uncontrolled HIV viral load, HAART via transient immune restoration may contribute to resolving CMV retinitis. PMID- 10855369 TI - [Infective endocarditis following incomplete ablation of a pacemaker]. AB - Pacemaker lead-related infective endocarditis is uncommon but mortality remains high. We report the case of a 63-year-old man who presented with a history of intermittent low-grade fever and no other sign for 15 months. Fever had developed after incomplete removal of a pacemaker with the ventricular lead left in situ followed by a new implantation of cardiac stimulation material. Positive blood cultures and transesophageal echocardiography showing a vegetation on a pacemaker lead gave the diagnosis. Initial antibiotic therapy was insufficient and complete surgical extraction of the pacemaker and leads was required. A huge vegetation was seen on the old ventricular lead. The other leads were not affected. Outcome was good. The paucity of symptoms in pacemaker lead-related infective endocarditis makes diagnosis difficult. It must however be suspected in pacemaker patients with low-grade intermittent fever. Transesophageal echocardiography is required. Treatment must combine antibiotic therapy with material extraction. PMID- 10855370 TI - [Study and treatment of drug addictions: an emergency that at last is considered]. PMID- 10855371 TI - [Abuse of flunitrazepam in opioid addicts]. AB - Fifty-three heroin addicts, nineteen of them methadone treated, were asked about their use of flunitrazepam. Drug abuse began after a medical prescription in 50% of cases; flunitrazepam was taken per os, but also i.v. in 19% of cases. The mean daily dose was about 20 mg/day, often associated with other psychotropic drugs, alcohol, opiates or cocaine. Attention is focused on the amnesia and increased feeling of power (11% of cases), often resulting in serious violent offenses. PMID- 10855372 TI - [Maintenance treatment for opioid dependence in care centers: the OPPIDUM program of the Evaluation and Information Centers for Drug Addiction]. AB - The aim of this study was to analyze information concerning multiple drug addiction, illicit behaviors and use of the venous route by maintenance treatment patients included in the October 1998 survey of the OPPIDUM program. Among 1,462 observations, 71% of the subjects were taking maintenance treatments (60% high dose buprenorphine and 40% methadone). High-dose buprenorphine was taken without medical supervision in 10% of cases. Indicators of abuse were high in this case: multiple drug addiction and intravenous use of buprenorphine (28%). Patients maintained by methadone were older and living in better socio-economic conditions than patients maintained by high-dose buprenorphine. However, in the two groups, the percentage of patients using the intravenous route was the same (15% and 21%). More cocaine was used by the methadone group (16% versus 7%). Thirty-seven percent of the subjects maintained on high-dose buprenorphine were followed by a general practitioner. They appeared to be more unbalanced and in more precarious condition than subjects treated in specialized care centers but they were not representative of the patients maintained by buprenorphine. It would be important to determine why these subjects consult a specialized care center. PMID- 10855373 TI - [Physical and sports activities in the history of patients treated for addictions. Report 1999 of the study sponsored by the Ministry of Youth and Sports (France)]. AB - Early February 1999, the French Ministere de la Jeunesse et des Sports (Youth and Sports Ministry) sponsored three different studies, aiming to prevent harmful behavior in the area of sport practices among youth. Two years earlier, our health care team working with drug users published reports on the meaningfulness of intensive sports activities in the history of our patients. The present work was performed to highlight the midterm results of one of these studies, to better understand and quantify the importance of physical training in the history of a group of outpatients seen for addictive disorders and comorbid pathologies. For 20 consecutive weeks, 3,040 self-administered questionnaires were available for persons consulting 20 health centers, 2 self-help groups and a general practitioner network working in the field of alcohol or heroine abuse. One thousand one hundred and eleven questionnaires were filled out (36.1%) and returned by mail for complete analysis: 86% of the answering persons had practiced at least one sports activity or participated in physical training, 10.5% had participated in a national or international level competition, and 10.6% reported stress fractures. In the intensive sports group, 36% had used illicit drugs intravenously and 16.4% said they had already used doping substances. Only 28.4% said they experienced dependence during their period of intensive sports activities compared with 15.2% before this time, and a majority (56.4%) thereafter. Intensive sports or physical training should not be seen as a protective factor nor as a way of improving addictive behaviors. More studies are needed to evaluate individual vulnerability factors and specific harm of overtraining and to determine the exact periods when men and women participating in sports activities are likely to abuse drugs, especially at the end of their career. PMID- 10855374 TI - [Follow-up of opioid addicts treated with high-dose buprenorphine in a health care network. National retrospective study. Experience of French general physicians]. AB - AIMS: This study was designed to examine the profile of drug-dependent outpatients treated by general practitioners working in a health care network and to evaluate the impact of treatment with high-dose buprenorphine on their medical and social status. METHODS: A retrospective study was undertaken by 71 general practitioners, selected at random from physicians in four health care networks. Data for the period between June and December 1997 concerning the initial prescription, the first stabilization prescription and the most recent prescription, was collected retrospectively. RESULTS: Among the outpatients included in this study, high-dose buprenorphine treatment resulted in a clear reduction in the use of heroin (69.9%) and benzodiazepine (57.1%). It also reduced associated risks of infection and social vulnerability. CONCLUSION: This retrospective study seems to show that care by general practitioners proceeds satisfactorily. The majority of opiate-dependent outpatients were compliant with treatment and successfully reintegrated into society. This method of treatment will be effective if specialised training is given to the general practitioners within the framework of a health care network. PMID- 10855375 TI - [Treatment of viral hepatitis in drug addicts]. AB - Management of viral hepatitis in drug uses must follow a multidisciplinary approach within the framework of overall psychosocial, alcohologic, and medical care. Substitution is an important but not indispensable aspect. In patients who have achieved social, psychological and medical stability (possible initiation of antiretroviral treatment), antiviral treatments can be started and followed in the same way as for other patients. Prescriptions should aim at improving the liver disease and avoid progression to cirrhosis and complications. Ribavirin/interferon-alfa appears to be the combination of choice for first line treatment of hepatitis C (while waiting for combinations with protease inhibitors). Anti-HBV vaccination is the treatment of choice for the prevention of hepatitis B and D. Coordination and information sharing between health and social care partners is crucial for the management of these patients. PMID- 10855376 TI - [Use of methadone as analgesic]. AB - Methadone is a synthetic opioid agonist which is mostly used for drug maintenance therapy in opioid addicts. It has also proven to be a powerful analgesic four times more powerful than oral morphine. The main indication is cancer pain as an alternative to oral morphine in case of tolerance or side effects due to accumulation of active metabolites. Methadone however shows wide inter-individual variability in half-life necessitating a carefully scheduled administration scheme. Side effects are similar for morphine and methadone. Methadone might be a safe and effective analgesic in patients with renal disease because its metabolites are almost exclusively excreted in the feces. PMID- 10855377 TI - Acute benzodiazepine withdrawal delirium after a short course of flunitrazepam in an intensive care patient. AB - Acute benzodiazepine and opioid withdrawal syndromes are increasingly described in intensive care unit patients. There is often some difficulty in distinguishing between the respective responsibilities of the sedative and analgesic agents. We report a case of acute benzodiazepine withdrawal delirium after a short course of flunitrazepam in an intensive care patient. Flunitrazepam was administered because of institution of mechanical ventilation. Recent data on the incidence and risk factors of the syndrome in intensive care unit patients are presented. Criteria for diagnosis and therapeutic guidelines are discussed. PMID- 10855378 TI - [Mental anorexia in aging. Report of a case]. AB - Elderly subjects who refuse to eat, and thus develop malnutrition, can be observed both in institutional and community settings. Causes include physiological changes related to the aging process, mental disorders and environmental factors. Most cases of anorexia nevrosa begin in adolescence but a sizeable number may develop at a later age. The term anorexia nevrosa in the elderly is proposed to describe this clinical entity observed in a woman who developed anorexia nevrosa at the age of 76 years. This entity is examined in comparison with prolonged earlier-onset anorexia nevrosa and late-onset anorexia. PMID- 10855379 TI - [Paroxetine withdrawal syndrome]. AB - Withdrawal syndrome after discontinuing serotonin re-uptake inhibitors, especially paroxetine, is largely unknown to most physicians. Variable incidence has been reported. Our aim was to stress the main clinical features of this syndrome. Serotonin re-uptake inhibitor withdrawal syndrome generally begins within 24 to 48 hours after discontinuing the drug. Signs reach their maximum on day 5 and usually resolve within 2 to 3 weeks. Withdrawal syndrome is more common with short half-life drugs (paroxetine, fluvoxamine). The intensity of the clinical signs depends on the daily dose and how long the drug has been given. The main signs are dizziness, vertigo, headache, nausea, and flu-like symptoms as well as anxiety, confusion, irritability, excessive dreaming and insomnia. Risk factors usually stressed are poor treatment compliance, previous withdrawal syndrome with another drug, concomitant medication and alcohol consumption. The syndrome can be prevented by tapering off the dose and patient education. When a withdrawal syndrome is present, it is advisable to reintroduce the drug then withdraw gradually. PMID- 10855380 TI - [Interview with Jean Clavreul by Mario Sanchez)]. PMID- 10855381 TI - [Strive for excellence and addiction to body movement: a risk model in high-level athletes]. AB - Champion athletes strive to attain a personal goal defined by a socially constructed image of psychomotor performance to be accomplished at the moment of the championship celebration. This intrapsychic process is initiated by a transformation of the body, programmed and controlled by repeated training. The athlete's body becomes accustomed to ritualized obsessive movements, favoring the feeling of self-fulfillment solely during muscular effort (contraction/relaxation, displacement). This social goal of excellence implies personal adaptation involving an addictive link to movement: a mechanism uniquely valid in high level sports. Twelve years experience in psychological support of high-level athletes participating in Olympic sports has led to an analysis of this adaptive mechanism and a proposed psychopathological model of its invasion of the athlete's psychic economy. PMID- 10855382 TI - [Economic approach to public health problems. Childhood accidents in France]. PMID- 10855383 TI - [Parental care-giving, the example of cystic fibrosis]. PMID- 10855384 TI - [Development of care activities for children with sickle cell disease at the Robert-Debre Hospital (Paris), between 1992 and 1996]. AB - AIM: In recent years, physicians at the Robert-Debre pediatric hospital in Paris perceived an increase in activity linked to sickle cell disease care. Our study had two objectives: first, to describe the evolution of care for children with sickle cell disease in the hospital, then, if a heavier intensity of care was shown, to try to investigate its causes. METHODS AND PATIENTS: We conducted a retrospective study using two strategies. On one hand, we compared the group of children followed up in 1992 with the group of children followed up in 1996 for their phenotype and hospitalizations (frequency, length of stay and type of complications), on the other hand, we described the course of complications and hospitalizations in the cohort of children followed from 1992 to 1996. Children were spotted through lists established by the 'Center for sickle cell disease', which coordinates the follow-up of all sickle cell patients in the hospital. Data came from the hospital's information system, and from all (medical, nursing, and social) individual records. RESULTS: The major result of this study shows an increasing activity linked to sickle cell disease care in this hospital: multiplication of hospitalizations and increasing work load for the healthcare teams. This situation is due to a larger recruitment, a higher emergency hospitalization rate, and an increasing rate of complications among the sickle cell children. The children's ageing is part of the explanation. The work load for the healthcare team linked to each hospitalization has also grown, as shown by an increasing rate of morphine prescription. PMID- 10855385 TI - [Patient-controlled analgesia for prolonged pain in the child. An open-label feasibility study of a standardized method]. AB - BACKGROUND: Patient-controlled analgesia (PCA) has been shown to be superior to a continuous morphine infusion for the treatment of ongoing pain in children over five years of age. Nevertheless, prescription parameters such as the bolus dosage and the possible association of a continuous background infusion have not yet been standardized. PATIENTS AND METHODS: Thirty-three children, aged four to 17, hospitalized in a pediatric hematology ward, benefited from PCA with a standardized prescription: a bolus dosage of at least 25 mg/kg, without a background infusion. Morphine consumption, side effects and efficacy on pain relief were followed. RESULTS: Median of mean morphine consumption was 0.32 mg.kg 1.d-1. Median of maximal consumption was 0.58 mg.kg-1.d-1. Mean duration was nine days. No important side effects were noted, except in two patients. They presented prolonged constipation and a poor quality of nocturnal sleep, but they also had a major depressive syndrome persisting after resolution of pain. Efficacy was comparable to a continuous intravenous infusion, and nocturnal sleep was of good quality for 31 children. CONCLUSION: This standardized technique of PCA can be used extensively in children over five years of age. It can be used as a reference for further studies. PMID- 10855386 TI - [Office-based pediatricians' knowledge and adherence to a consensus statement on acute rhinopharyngitis in the child]. AB - AIM: To assess the impact on community-based pediatricians of the conclusions of the 10th Conference de Consensus en Therapeutique Anti-Infectieuse (CCTAI) on the antibiotic treatment in acute nasopharyngitis (ANP). METHODS: Fifty-six pediatricians took part in the study. Over a period of 15 days in October 1998, all the children (n = 997) presenting an ANP were prospectively included. The prescription of an antibiotic treatment as well as the clinical criteria authorizing it, according to the conclusions of the 10th CCTAI, were recorded. The participants were not told the purpose of the study. RESULT: Sixty percent of the pediatricians questioned were familiar with the 10th CCTAI. Forty-five percent said they complied with it, but only 7% stated it had changed their day to-day clinical practice. Based on the conclusions of the 10th CCTAI, an antibiotic treatment would have been discussed for 38% of the children. Twenty four percent of them were given one. For 54% of the children that were given an antibiotic, such treatment was disapproved by the 10th CCTAI. No significant association has been established between familiarity with the 10th CCTAI and the following criteria: gender, age, hospital activity, years of practice, medical journals read, and belief in the existence of a license for one or more antibiotics for ANP in children. However, this last criterion was significantly (P = 0.03) associated with an increase in the percentage of antibiotics prescribed: 29% vs 16%. DISCUSSION AND CONCLUSION: The 10th CCTAI has had a moderate impact on the day-to-day practice of the pediatricians who took part in our study. Several explanations are discussed. The authors emphasize the virtual lack of indications of antibiotics in ANP. PMID- 10855387 TI - [Trends in infant mortality in France: frequency and causes from 1950 to 1997]. AB - OBJECTIVES: To present an analysis of the infant mortality trends and causes of death in France from the beginning of the 1950s, neonatal (0-27 days) and post neonatal mortality (27-364 days) being considered separately. MATERIAL AND METHODS: We used the data from the national registries of births computed by INSEE (National Institute of Statistics and Economic Surveys) and of causes of deaths computed by Inserm (National Institute of Health and Medical Research). We analysed the evolution of the infant death rates from 1950 to 1997, the overall mortality for males and the percentages of causes of death at three different periods. RESULTS: Mortality has changed according to neonatal or post-neonatal ages. A constant improvement was recorded for neonatal mortality up to 1995 (2.9 per 1,000), while there was a stagnation for post-neonatal mortality between 1979 and 1993, followed by a sharp decrease (2.0 per 1,000 in 1995). During the neonatal age the main causes of death are conditions generated in the neonatal period and congenital abnormalities, both decreasing regularly; during the post neonatal age the main cause is sudden infant death syndrome, which fell dramatically during the last four years. CONCLUSION: Several factors related to medical care, nursing and type of registration are contributing simultaneously to the important variations in mortality found in our results. PMID- 10855388 TI - [Sweet syndrome in an infant]. AB - BACKGROUND: Acute febrile neutrophilic dermatosis (Sweet's syndrome) is very uncommon in infancy. Systemic corticosteroid treatment is the standard therapy, usually leading to dramatic improvement within a few days. CASE REPORT: A seven month-old female infant was admitted for investigation of a rash developing over ten days with fever. Physical examination and skin biopsy led to a diagnosis of Sweet's syndrome. The relative inefficiency of systemic corticotherapy induced the parents to stop all treatment. CONCLUSION: This case report allows us: 1) to consider the clinical and biological features of Sweet's syndrome in infancy; 2) to describe a corticosteroid resistant disease; and 3) to observe the course of a spontaneous evolution of Sweet's syndrome. PMID- 10855389 TI - [Neonatal hyperthyroidism in a premature infant born to a mother with Grave's disease]. AB - BACKGROUND: Neonatal thyrotoxicosis is most commonly due to transplacental transfer of maternal thyroid-stimulating hormone receptor antibodies (TRAb). Bioassay of thyrotropin receptor antibodies may help to determine the risk for neonatal hyperthyroidism. CASE REPORT: Thyrotoxicosis developed in a premature infant born to a mother with Graves' disease, with a low level of TRAb by bioassay. The infant was treated with carbimazole for two months, until TRAb had disappeared. CONCLUSION: Bioassay TRAb is not always reliable for predicting the development of neonatal hyperthyroidism in infants born to mothers with Graves' disease. Thyroid function should be measured in all these neonates. PMID- 10855390 TI - [Respiratory distress in three newborns after mask disinfection with Endosporine. Probable role of glutaraldehyde]. AB - BACKGROUND: Disinfectants for medical devices are uncommonly a cause of iatrogenic adverse effects. Nevertheless, when misused, they can induce severe complications. Three cases of acute respiratory distress in newborns probably induced by glutaraldehyde are reported. CASE REPORTS: Three children born by Caesarean section between 8 and 19 May 1999 in the same hospital presented acute respiratory distress requiring hospitalization in the neonatal intensive care unit; one child was premature. The clinical appearance, which was initially normal, deteriorated with a respiratory distress in 30 to 60 minutes. Recovery was uneventful in all cases. The diagnosis considered was a hyaline membrane disease. The enquiry conducted after this cluster onset identified, as a main contributing factor, the disinfection procedure recently introduced in the surgical theater. CONCLUSION: Review of toxicologic data on glutaraldehyde shows this is a highly irritating chemical for the respiratory tract, even at low concentrations. Clinical and radiologic features in these three neonates are compatible with a pulmonary sub-edema on an immature alveolar setting. The hypothesis proposed is that glutaraldehyde, the active ingredient of the biocidal formula used to disinfect the respiratory masks, was massively desorbed from the rubber and foam of which masks are made. PMID- 10855391 TI - [Hereditary methemoglobinemias]. AB - The different forms of hereditary methemoglobinemia are described. Dominant hereditary methemoglobinemia (hemoglobin M diseases) is due to punctual mutations on the alpha or beta globin chain leading to its permanent oxidation. Recessive hereditary methemoglobinemia is due to a deficit of the NADH-cytochrome b5 reductase, with two different clinical presentations according to whether the deficit is limited to blood cells (type I) or is generalized (type II); the latter is extremely severe and justifies a prenatal diagnosis. Finally, the only known case of recessive hereditary methemoglobinemia due to a deficit of b5 cytochrome is quoted. PMID- 10855392 TI - [Antibiotic therapy in cystic fibrosis. I. Pharmacologic specifics of antibiotics]. AB - Antibiotherapy is one of the main treatment in cystic fibrosis. Antibiotic administration schedules are different from normal patients because of pharmacokinetic and pharmacodynamic particularities. In moderate disease, the digestive resorption of antibiotics is delayed and their half-life is reduced due to an increase in total clearance. In severe disease, the volume of distribution of antibiotics is increased due to the higher proportion of lean mass in these malnourished patients. Other particularities limit the action of antibiotics such as thick sputum, which limits drug penetration; the property of Pseudomonas aeruginosa to be surrounded by a biofilm; alteration of local antibacterial defense; and inhibition of antibiotics by local factors. Systematic prescription of a biotherapy beta-lactam-aminoglycoside and obtaining high antibiotic concentration in situ might limit this antagonism. In spite of particular therapeutic schedules such as single daily dose for aminoglycoside and continuous infusion for beta-lactams, the intervals between administrations must be narrowed for time-dependent antibiotics, and the total daily dose increased by 20 to 30% for concentration-dependent antibiotics. PMID- 10855393 TI - [Who are my parents? Filial adoption and the function of time and place]. AB - Despite a substantial development for several years, adoption frequently remains seen as an unnatural phenomenon which cannot replace a biological relationship. In all periods and everywhere in the world children have been and are being resettled, and history and anthropological studies show that there exist as many modes of adoption as societies. In some societies, such as in Polynesia, no differences exist between biological and adoptive parenthood. The most important thing is to allow every child to have a family, biological or not. The adopted child must be told about her/his adoption as early as possible in a natural manner. Concealing the truth may lead to a profound psychological wound when later learned by the child. PMID- 10855394 TI - [Radiologic case of the month. Constrictive bronchiolitis]. PMID- 10855395 TI - [Practical advice for women who want to breast feed]. AB - Breastfeeding is currently a rare practice in France. Pediatricians wanting to help those women who wish to breast feed should be able to provide them with proper information during the perinatal period, and to ensure that breast feeding begins under the best conditions, i.e., early initiation of breast feeding, then self-demand feeding, breast feeding without restrictions on duration and frequency, no supplementation of any kind, continuous rooming-in, and proper positioning of the baby. Most of the time this is sufficient to prevent complications such as engorgement, sore nipples, mastitis or poor weight gain of the baby. If difficulties appear, they have to be managed by means of simple advice and reassurance, without interrupting breast feeding. PMID- 10855396 TI - [Breastfeeding: pledging allegiance to ourselves]. AB - Since prehistoric times, how babies should be fed has been a constant for our species, which is one of the more than 4,600 varieties of milk-producing creatures collectively known as mammals. Each mammal produces its own unique milk, and thus feeding a baby something other than breast milk will always be a deviation from the biological norm for our species. Indeed, breast milk is the only truly universal food uniting all 6 billion of us, irrespective of geography and culture. While in extreme cases it may be necessary to feed a baby a breast milk substitute, this is not without considerable risk. In any case, it should remain the exception. Paediatricians and other professionals responsible for health services are particularly well placed to provide the leadership to sustain or, if necessary, to re-establish, a 'breastfeeding culture'. The WHO/UNICEF Baby friendly Hospital Initiative offers a useful framework for doing just this. In only eight short years, more than 16,000 hospitals in 171 countries have begun implementing the Initiative. It is hoped that hospitals in France will soon join this worldwide alliance. PMID- 10855397 TI - [Decisional procedure in school-age children with hyperkinetic syndrome]. AB - Hyperkinetic syndrome may be either secondary to an organic disease or a psycho effective disorder (mood and/or anxiety disorder), or primary as part of an attention deficit hyperactivity disorder. Precise diagnosis is essential before any therapeutic decision; this requires a complete anamnestic, behavioural, psychological, sensorial, and neurological evaluation. It is only when a reliable diagnosis has been made that a relevant therapeutic project can be proposed. An evaluation procedure and a decisional tree are presented. PMID- 10855398 TI - [Munchausen syndrome by proxy hits journalists!]. PMID- 10855399 TI - [Red cell parameters and iron status in 48 children with kwashiorkor in Dakar (Senegal)]. PMID- 10855400 TI - [Headache in malaria]. PMID- 10855401 TI - [Pseudomonas aeruginosa external otitis in four neonates]. PMID- 10855402 TI - [Duration of maternal breastfeeding in France]. PMID- 10855403 TI - [Neonatal presentation of Prader-Willi syndrome]. PMID- 10855404 TI - [Management of childhood heart disease in Madagascar. What perspective for tomorrow?]. PMID- 10855405 TI - [Main coronary lesion: indication for angioplasty?]. PMID- 10855406 TI - Spontaneous changes in ventricular tachycardia cycle length and their relation to earliest sites of epicardial activation in a canine model. AB - BACKGROUND: The purpose of this study was to examine the spontaneous changes in cycle length during episodes of sustained monomorphic (MVT) and polymorphic (PVT) ventricular tachycardias and to relate these changes with the earliest epicardial activation site of the beat. METHODS: Isochronal activation maps were obtained from 127 unipolar electrograms recorded from the surface of both ventricles with a sock electrode array in 24 open chest anesthetized dogs. After atrioventricular block, the left anterior descending coronary artery was occluded for 60 min under ventricular pacing (140/min), followed by reperfusion. In 7 dogs the left stellate ganglion was stimulated 5 min after reperfusion. RESULTS: In 7 MVTs (reperfusion) and 4 PVTs (sympathetic stimulation), cycle length changes showed an initial acceleration, reaching a minimum cycle length and then decelerating before termination. Isochronal maps showed radial spread from earliest activation, without conduction block. Cycle length (481 +/- 80 msec) in MVT had beat to beat variations of 15 +/- 17 msec corresponding to small shifts in sites of the earliest activation, clustered along the border of the ischemic myocardium. In PVTs the cycle length (352 +/- 90 msec, p < 0.01) had a variability of 62 +/- 23 msec, corresponding to wide changes in the sites of earliest activation in right and left ventricles. Linear regression analysis showed a strong and significant correlation between cycle length variability and the number of electrodes with the earliest activation (r = 0.77, p < 0.0001). CONCLUSION: In these models of monomorphic and polymorphic ventricular tachycardias, cycle length variability showed a significant correlation with the number of electrodes with the earliest activation. MVTs showed concentrated origins with regular cycle length, whereas PVTs showed dispersed origins with irregular cycle length. These results suggest that the earliest epicardial activation site of the beat could be a factor in determining the dynamics in the cycle length. PMID- 10855407 TI - [Is the concept of "jumping wave" still valid?]. AB - The concept of the "jumping wave" phenomenon, i.e. of the slow and difficult passage of activation fronts from one septal mass to the other through an "intraseptal barrier", is derived from experimental studies of the Mexican School of Electrovectorcardiography. OBJECTIVE: To confirm the existence of histologically bipartite interventricular septum and of the electric independence of both septal masses. METHODOLOGY: We examined the histological characteristics of both septal masses in rat, canine, and human hearts. We also analyzed the morphological and chronological data of intracavitary records in the presence of different degree proximal blocks, comparing these findings with those obtained when peripheral blocks existed. RESULTS: We found a medial, longitudinal band between the two septal masses in animal as well as in human hearts. The analysis of intracavitary electric records confirmed a slow and difficult transmission of the activation fronts from one septal mass to the other, in the presence of proximal blocks and ventricular arrhythmias. Morphological and chronological changes of intraventricular complexes could not be explained if the septal activation process were of syncytial type. CONCLUSIONS: Results of this study firmly support the validity of our approach to the septal activation process in the presence of ventricular conduction disorders and arrhythmias. This approach helps to detect the possible coexistence of dead septal tissue. PMID- 10855408 TI - [Simultaneous analysis of myocardial perfusion and ventricular function. Normal values in the Mexican population]. AB - Myocardial perfusion imaging is an useful procedure in the evaluation of patients with coronary artery disease, Gated SPECT technique evaluates simultaneously perfusion and ventricular function, left ventricular ejection fraction (LVEF), ventricular volumes and the transient ischemic dilatation of the left ventricle. OBJECTIVE: To evaluate the normal ventricle volumes and the ejection fraction of the LV obtained automatically with the Gated SPECT in Mexican population. METHODS: 100 patients were studied with low likelihood for CAD. All of them were studied with Tc-99m Sestamibi Gated SPECT. We obtained automatically the LVEF, and the end diastolic and systolic left ventricular volumes. RESULTS: Myocardial perfusion, regional motion and systolic thickening were normal in all patients. We obtained the mean values of EF and end diastolic and systolic volumes. These values were lower in female. CONCLUSION: Tc-99m Sestamibi myocardial perfusion SPECT is an useful procedure, with high accuracy for the simultaneous evaluation of myocardial perfusion and ventricular function. PMID- 10855409 TI - [Stent in protected and non-protected left main coronary artery: results and short-term follow-up]. AB - We examined the immediate and short-term outcomes after stenting protected and unprotected left main coronary artery (LMCA) stenoses, in patients with normal ventricular function. Left main coronary artery disease is regarded as an absolute contraindication for coronary angioplasty, because it has been associated with high procedural morbidity and poor mid-term results. Between february 1995 and february 1999, 596 procedures were performed in 468 patients. Ten patients who had disease involving the left main coronary artery were included. They were not candidates for coronary surgery. The post-stent antithrombotic regimens were aspirin and ticlopidine. The procedural success rate was 100% without episodes of subacute thrombosis. Three to six months follow-up angiography was performed in all, restenosis occurred only in two patients, there were two repeat PTCA (20%) and there were no deaths. CONCLUSIONS: Stenting of unprotected and protected left main coronary artery stenoses may be a safe and effective alternative to surgery in carefully selected patients with normal left ventricular function. The results of our study suggests that when patients have prohibitive surgical risks, elective LMCA angioplasty and/or stenting maybe undertaken with a high procedural success rate as an effective alternative to CABG in carefully selected patients. Further studies in larger patient populations are needed to assess late outcome. PMID- 10855410 TI - [Permanent rhythm and conduction disorders in patients surgically treated for atrial septal defect]. AB - Atrial septal defect (ASD) represents a congenital heart disease with good prognosis, however, atrial arrhythmias are well-documented complications. A retrospective study was performed to assess the prevalence, types and risk factors of arrhythmias in patients with ASD after surgical correction. 920 patients were analyzed retrospectively; cases with complex congenital heart disease or another systemic alteration conditioning atrial rhythm disturbances were excluded. 460 patients with ASD corrected by surgery were followed at least for six months after the procedure. 29.3% of patients were male and 70.7% female. Direct closure was performed in 63%, pericardial patch was installed in 27%. 29 patients (6.3%) had conduction and rhythm disturbances before surgery, the most common arrhythmias were atrial flutter (34.5%), first degree A-V block (31%) and low right atrial rhythm (27.6%). Six months after atrial defect closure, 65 patients (14.13%) had arrhythmias, 44.6% atrial flutter, 20% ectopic atrial rhythm, 10.8% sick sinus syndrome. The risk of atrial arrhythmias was related to age at surgical repair, pulmonary hypertension and atrial arrhythmias before surgery. PMID- 10855411 TI - [Radiofrequency transcatheter ablation in atrial tachycardia]. AB - Incessant atrial tachycardia is an infrequent arrhythmia. Specially difficult to treat medically. Radiofrequency catheter ablation has been used successfully to cure a variety of supraventricular tachycardias. The purpose of this work is to report our initial experience in the treatment of atrial tachycardia. Ten patients, mean age 28.7 +/- 15 year with conventional drug-resistant symptomatic atrial tachycardia were treated with selective ablation of the focus using radiofrequency energy. It was found an abnormal automaticity in 10 tachycardias and in only one patient intra-atrial reentrant was supported. Radiofrequency energy was successful in 10 of 11 tachycardias with a mean of 9.3 +/- 6.8 applications using the technique of local atrial electrogram activation time with a mean value of -54 +/- -31 milliseconds at the successful ablation sites. No complications were observed and one patient had an early clinical recurrence. All patients with successful ablation are symptom-free, in sinus rhythm and without antiarrhythmic medications after 1 to 28 months of follow-up. Our initial experience support that radiofrequency catheter ablation is a safe and effective therapeutic option for incessant atrial tachycardia. PMID- 10855412 TI - Endomyocardial fibrosis (Davies disease) coincidental with systemic lupus erythematosus. AB - This is the case of a 27 years-old woman with signs and symptoms of severe untreatable congestive heart failure, anemia, gingival mucosa ulcers, photosensitivity and alopecia. The electrocardiographic, echocardiographic, angiographic and hemodynamic data oriented the diagnosis of restrictive cardiomyopathy, mitral insufficiency secondary to mitral prolapse and bi-atrial dilation. The histologic study of the endomyocardial biopsy, performed during catheterization, showed signs of endomyocardial fibrosis, and immunological analysis was compatible with systemic lupus erythematosus. As far as we know, this is the first case of endomyocardial fibrosis (Davies disease) associated with systemic lupus erythematosus published in the medical literature. The etiology of Davies disease remains unrevealed and its association with systemic lupus erythematosus suggest a probable autoimmune origin. PMID- 10855413 TI - [Arthroplasty in congenital aortic coarctation in adults with balloon and endovascular prosthesis: immediate results and 6-month follow-up]. AB - Up to 1982, surgery was the treatment of aortic coarctation, with postsurgical recoarctation in 39% of cases. Since 1984 balloon aortoplasty has been performed successfully in adolescents and adult patients. We present the immediate results, and more than six months follow up of 6 patients with congenital aortic coarctation, who underwent this procedure. Five of the six patients were male, with an average age of 28.6 years (15-46), and in 4 of them a stent was placed. Systolic pressure of ascending aorta decreased from 187.1 mm of Hg (+/- 41.8) to 128 (+/- 25.4), and transaortic gradient from 66 mm of Hg (+/- 21.8) to 4.8 (+/- 7.6). Coarctation luminal diameter increased from 4.6 mm (+/- 1.41) to 14.3 (+/- 3) in patients with only balloon aortoplasty and to 17.8 mm with stent placement, p = NS. Angiography in three patients with stent at 6 months did not reveal restenosis, all six patients require less antihypertensive medications. Acute and chronic complications, percentage and time of restenosis, long term results, and possible benefit of stents are yet to be determined. PMID- 10855414 TI - [Unusual case: injury of the 4 heart valves in a patient with old heart injury caused by fire arm. Review of the literature]. AB - We review the literature on heart wounds. Along the history surgeons regarded heart wounds as forbidden place for therapeutic approach. We present a patient who sustained a bullet wound to the heart and two years later 3 knife' stabs in the same place injuring its 4 valves and inducing 2 fistulae: one from the aorta to the pulmonary artery, the other from left-ventricle and the pulmonary artery. Surgery was successful in repairing the damage without the need of valve replacement. PMID- 10855415 TI - [Management of chronic phase atherosclerosis]. PMID- 10855416 TI - [Guidelines for the diagnosis and treatment of unstable angina and non-Q myocardial infarction: proposed changes]. AB - In 1994, the United States Agency for Health Care Policy and Research issued clinical practice guidelines for the diagnosis and management of unstable angina and non-Q-wave myocardial infarction. In the past 5 years, rapid progress has been made in the management of patients with unstable coronary syndromes, and treatment guidelines should be revised to reflect these advances. An international forum of cardiology investigators convened to discuss areas in which the diagnosis and treatment of unstable angina/non-Q-wave myocardial infarction should be modified. Although there were areas of controversy, it was agreed that there is sufficient evidence to recommend the following changes: 1) the use of serum cardiac markers should be expanded to include troponin I and T levels as diagnostic and prognostic tools, 2) low-molecular-weight heparins should replace UH as antithrombin agents, 3) new classes of antiplatelet agents are recommended in addition to aspirin, 4) the use of cholesterol-lowering drugs is appropriate in the long-term management of these patients. The rationale for these suggested revisions, including evidence from pertinent clinical trials, is discussed in detail in the accompanying document. PMID- 10855417 TI - The source of feelings of familiarity: the discrepancy-attribution hypothesis. AB - Many investigators have observed that the feeling of familiarity is associated with fluency of processing. The authors demonstrated a case in which the feeling of familiarity did not result from fluency per se; they argued that it resulted instead from perceiving a discrepancy between the actual and expected fluency of processing (B. W. A. Whittlesea & L. D. Williams, 1998). In this article, the authors extend that argument. They observed that stimuli that are experienced as strongly familiar when presented in isolation are instead experienced as being novel when presented in a rhyme or semantic context. They interpreted that result to mean that in those other contexts, the subjects brought a different standard to bear in evaluating the fluency of their processing. This different standard caused the subjects to perceive their performance not as discrepant, but as coherent in one case and incongruous in the other. The authors suggest that the perception of discrepancy is a major factor in producing the feeling of familiarity. They further suggest that the occurrence of that perception depends on the task in which the person is engaged when encountering the stimulus, because that task affects the standard that the person will apply in evaluating their processing. PMID- 10855418 TI - Hindsight bias: a by-product of knowledge updating? AB - With the benefit of feedback about the outcome of an event, people's recalled judgments are typically closer to the outcome of the event than their original judgments were. It has been suggested that this hindsight bias may be due to a reconstruction process of the prior judgment. A model of such a process is proposed that assumes that knowledge is updated after feedback and that reconstruction is based on the updated knowledge. Consistent with the model's predictions, the results of 2 studies show that knowledge after feedback is systematically shifted toward feedback, and that assisting retrieval of the knowledge prior to feedback reduces hindsight bias. In addition, the model accounts for about 75% of cases in which either hindsight bias or reversed hindsight bias occurred. The authors conclude that hindsight bias can be understood as a by-product of an adaptive process, namely the updating of knowledge after feedback. PMID- 10855419 TI - ROC curves and confidence judgements in recognition memory. AB - Most models of recognition memory rely on a strength/familiarity-based signal detection account that assumes that the processes giving rise to a confidence judgment are the same as those giving rise to an old-new decision. Confidence is assumed to be scaled directly from the perceived familiarity of a probe. This assumption was tested in 2 experiments that examine the shape of confidence-based z receiver operating characteristic (zROC) curves under different levels of response bias induced by changing stimulus probabilities (Experiment 1) and payoffs (Experiment 2). Changes in the shape of the zROC curves with bias indicate that confidence is not scaled directly from perceived familiarity or likelihood. A model of information accumulation in recognition memory is proposed that can account for the observed effects. PMID- 10855420 TI - Negative transfer errors in sequential cognitive skills: strong-but-wrong sequence application. AB - Three experiments investigated the role of processing sequence knowledge in negative transfer within multistep cognitive skills. In Experiments 1 and 2, more training resulted in higher error rates when new processing sequences that resembled familiar ones were introduced in transfer. Transfer error responses were executed with the same speed as correct responses to familiar sequence trials, and the errors appeared to be undetected by the performers. Experiment 3 tested whether the effects of sequence learning were attributable to explicit or implicit knowledge of processing sequences. Evidence favored the implicit learning interpretation. Findings are discussed in relationship to earlier demonstrations of the einstellung effect and to current taxonomic theories of human error. PMID- 10855421 TI - Attention and perceptual priming in the perceptual identification task. AB - Prior research indicates that manipulations of attention during encoding sometimes affect perceptual implicit memory. Two hypotheses were investigated. One proposes that manipulations of attention affect perceptual priming only to the extent that they disrupt stimulus identification. The other attributes reduced priming to the disruptive effects of distractor selection. The role of attention was investigated with a variant of the Stroop task in which participants either read words, identified their color, or did both. Identifying the color reduced priming even when the word was also overtly identified. This result held regardless of whether color and word were presented as a single object (Experiments 1 and 2) or as separate objects (Experiment 4). When participants read and identified a color, the overt order of the responses did not matter; both conditions reduced priming relative to reading alone (Experiment 3). The results provide evidence against the stimulus-identification account but are consistent with the distractor-selection hypothesis. PMID- 10855422 TI - On the relationship between recognition speed and accuracy for words rehearsed via rote versus elaborative rehearsal. AB - Tacit within both lay and cognitive conceptualizations of learning is the notion that those conditions of learning that foster "good" retention do so by increasing both the probability and the speed of access to the relevant information. In 3 experiments, time pressure during recognition is shown to decrease accessibility more for words learned via elaborative rehearsal than for words learned via rote rehearsal, despite the fact that elaborative rehearsal is a more efficacious learning strategy as measured by the probability of access. In Experiment 1, participants learned each word using both types of rehearsal, and the results show that access to the products of elaborative rehearsal is more compromised by time pressure than is access to the products of rote rehearsal. The results of Experiment 2, in which each word was learned via either pure rote or pure elaborative rehearsal, exhibit the same pattern. Experiment 3, in which the authors used the response-signal procedure, provides evidence that this difference in accessibility owes not to differences in the rate of access to the 2 types of traces, but rather to the higher asymptotic level of stored information for words learned via elaborative rehearsal. PMID- 10855423 TI - Is perceptual salience needed in explanations of the isolation effect? AB - The isolation effect is a well-known phenomenon that has a well-accepted explanation: An item that is isolated on a list becomes perceptually salient, which leads to extra rehearsal that enhances memory for the isolate. To evaluate this hypothesis, the authors isolated an item near the beginning of a list. Immediately after each item was presented for study, participants judged the likelihood of recalling the item. Although the isolation effect occurred, participants did not judge the isolate as being more memorable than the preceding item, suggesting that the isolate was not salient. In a second experiment, participants rehearsed items aloud. Isolation at the beginning of the list did not produce extra rehearsal. By contrast, isolation in the middle of the list produced extra rehearsal; however, even when the isolate did not receive extra rehearsal, an isolation effect was evident. Thus, salience and extra rehearsal are not necessary for producing an isolation effect. PMID- 10855424 TI - Pop-out into memory: a retrieval mechanism that is enhanced with the recall of subject-performed tasks. AB - Subject-performed tasks (SPTs; i.e., carrying out the actions during study) improve free recall of action phrases without enhancing relational information. By this mechanism, items pop into a person's mind without active search, and this process especially extends the recency effect. The authors demonstrated the existence of the extended recency effect and its importance for the SPT recall advantage (Experiments 1 and 2). Carrying out the action and not semantic processing caused the effect (Experiment 3). The extended recency effect was also not a consequence of a deliberate last-in, first-out strategy (Experiment 4), and performing a difficult secondary task (an arithmetic task) during recall reduced memory performances but did not influence the extended recency effect (Experiment 5). These data support the theory that performing actions during study enhances the efficiency of an automatic pop-out mechanism in free recall. PMID- 10855425 TI - Item and order information in subject-performed tasks and experimenter-performed tasks. AB - This study investigated the enactment effect from the perspective of the item order hypothesis (e.g., M. Serra & J. S. Nairne, 1993). The authors assumed that in subject-performed tasks (SPTs), item encoding is improved but order encoding is disrupted compared with experimenter-performed tasks (EPTs), that order encoding of EPTs is only better in pure lists, and that the item--order hypothesis is confined to short lists. Item information was tested in recognition memory tests, order information in order reconstruction tasks, and both item and order information in free-recall tests. The results of 5 experiments using short (8 items) and long lists (24 items) in a design with list type (pure, mixed) and encoding condition (EPT, SPT) as factors supported the hypotheses. PMID- 10855426 TI - Organization of visual short-term memory. AB - The authors examined the organization of visual short-term memory (VSTM). Using a change-detection task, they reported that VSTM stores relational information between individual items. This relational processing is mediated by the organization of items into spatial configurations. The spatial configuration of visual objects is important for VSTM of spatial locations, colors, and shapes. When color VSTM is compared with location VSTM, spatial configuration plays an integral role because configuration is important for color VSTM, whereas color is not important for location VSTM. The authors also examined the role of attention and found that the formation of configuration is modulated by both top-down and bottom-up attentional factors. In summary, the authors proposed that VSTM stores the relational information of individual visual items on the basis of global spatial configuration. PMID- 10855427 TI - Evidence for 40-Hz oscillatory short-term visual memory revealed by human reaction-time measurements. AB - Four experiments show that presentation of a synchronous premask frame within a 40-Hz, flickering premask matrix primes subsequent detection of a Kanizsa-type square by generation of a 40-Hz prime. Reaction time (RT) priming effects indicated a 150-200-ms prime duration following premask display. RTs were also found to be sensitive to the phase relationship between offset of the premask display relative to the onset time of the target: Priming effects were maximal when the target was presented out of phase with premask presentation (i.e., at interstimulus intervals displaced by 180 degrees relative to the 40-Hz rhythm of premask-frame presentation). Taken together, these results demonstrate the existence of a very short-term visual memory that oscillates at 40 Hz. The findings are discussed in the context of complementary psychological and neurophysiological findings related to visual-object coding and the role of gamma band activity in the brain. PMID- 10855428 TI - Strategic control in word reading: evidence from speeded responding in the tempo naming task. AB - To investigate strategic control over response initiation in word reading, the authors introduce the tempo-naming task. Relative to baseline performance in the standard-naming task, participants were induced to respond with faster latencies, shorter durations, and lower levels of accuracy by instructing them to time response initiation with an experimentally controlled tempo. The tempo response cue attenuated stimulus effects, and as faster tempos reduced latencies, the number of spelling-sound errors remained constant, whereas the number of word, nonword, and articulatory errors increased. To explain these results, the authors propose input gain as a mechanism of control over processing speed. The experimenters sketch how input gain could account for the current results as well as for the results from stimulus-blocking experiments testing the route emphasis and time criterion hypotheses of strategic control. PMID- 10855429 TI - Decomposing morphologically complex words in a nonlinear morphology. AB - Most Hebrew words are composed of 2 intertwined morphemes: a triconsonantal root and a phonological word pattern. Previous research with conjugated verb forms has shown consistent priming from the verbal patterns, suggesting that verbal forms are automatically parsed by native speakers into their morphemic constituents. The authors investigated the decomposition process, focusing on the structural properties of verbal forms that are perceived and extracted during word recognition. The manipulations consisted of using verbal forms derived from "weak" roots that have one consonant missing in some of the forms. The results demonstrated that if 1 consonant is missing, the parsing system collapses, and there is no evidence for morphological priming. In contrast, when a random consonant is inserted into the weak form, the verbal-pattern priming re-emerges. This outcome suggests that the constraint imposed on the decomposition process is primarily structural and abstract. Moreover, the all-or-none pattern of results is characteristic of rule-based behavior and not of simple correlational systems. PMID- 10855430 TI - Language comprehension and probe-list memory. AB - Experiments were performed using probe-word recognition methodology in which participants read sentences that were presented 1 word at a time and were then shown a probe word and had to make a speeded response indicating whether the word had occurred in the sentence. One experiment showed that response times to probe words increased with the size of the set of candidate probes. The other experiments showed that the effects caused by name repetition in circumstances in which the repeated name was co-referential also occurred when the repeated name was not co-referential and when the order of words in a sentence was scrambled. The results suggest that responses in the task can be based on probe-list memory, a mental representation created to keep track of those words that the participant believes are likely to be probed, and that the use of the task to make inferences about language comprehension should be accompanied by controls ruling out such strategies. PMID- 10855431 TI - Inference using categories. AB - How do people use category membership and similarity for making inductive inferences? The authors addressed this question by examining the impact of category labels and category features on inference and classification tasks that were designed to be comparable. In the inference task, participants predicted the value of a missing feature of an item given its category label and other feature values. In the classification task, participants predicted the category label of an item given its feature values. The results from 4 experiments suggest that category membership influences inference even when similarity information contradicts the category label. This tendency was stronger when the category label conveyed class inclusion information than when the label reflected a feature of the category. These findings suggest that category membership affects inference beyond similarity and that category labels and category features are 2 different things. PMID- 10855432 TI - The effects of word co-occurrence on short-term memory: associative links in long term memory affect short-term memory performance. AB - In immediate serial recall tasks, high-frequency words are recalled better than low-frequency words. This has been attributed to high-frequency words' being better represented and providing more effective support to a redintegration process at retrieval (C. Hulme et al., 1997). In studies of free recall, there is evidence that frequency of word co-occurrence, rather than word frequency per se, may explain the recall advantage enjoyed by high-frequency words (J. Deese, 1960). The authors present evidence that preexposing pairs of low-frequency words, so as to create associative links between them, has substantial beneficial effects on immediate serial recall performance. These benefits, which are not attributable to simple familiarization with the words per se, do not occur for high-frequency words. These findings indicate that associative links between items in long-term memory have important effects on short-term memory performance and suggest that the effects of word frequency in short-term memory tasks are related to differences in interitem associations in long-term memory. PMID- 10855433 TI - Limb amputation on renal replacement therapy. AB - Ten (10) diabetic and 7 non-diabetic patients on renal replacement therapy have undergone limb amputation in the authors' unit in the 1988 to 1996 period. The article examines the course of illness and survival patterns in this distinct and increasing sub-set in the amputee population. Rehabilitation and survival were significantly better in the diabetic group and it is recommended that it would be helpful for both prognosis and analysis if the sub-set of amputees on treatment for chronic renal failure is further divided into diabetic and non-diabetic sub sets. PMID- 10855434 TI - Lower limb deficient children in The Netherlands: epidemiological aspects. AB - Information on the characteristics of children with limb deficiencies and amputations in The Netherlands is largely lacking. The present study aimed to collect data about the prevalence of congenital deficiencies, the ratio of congenital to acquired limb deficiencies, types of lower leg deficiency or amputation and male/female ratios. Data were obtained from a regional birth defects registry for the northern part of The Netherlands (EUROCAT-NNL) and from a national survey. Inclusion criteria for the selection of the EUROCAT data were: children/foetuses with lower leg deficiencies born in 1981-1986. Inclusion criteria for the survey data were: children aged 1-18 years with congenital deficiencies or acquired amputations of the leg, excluding toe deficiencies/amputations. Both the regional birth defects registry and the national survey only yielded small numbers of children, which limits the validity of the authors' findings. The Eurocat data show a prevalence of lower leg deficiencies at birth of 2.07/10,000. Fifty-five (55) children/foetuses were included in the present study. The male/female ratio was 1:1. Of the live-born children, 30% also had defects of the upper limbs, while 38% had bilateral lower limb deficiencies. The national survey included 89 children, of whom 73% had congenital deficiencies, while the others had undergone amputations: of which 37% were due to malignancies, 29% to traumata, 13% to infections and 21% to other pathology. The male/female ratio was 7:3 for the children with congenital deficiencies versus 6:4 for the children with acquired amputations. In the group of congenital deficiencies, fibula deficiency was most frequently seen (36%), while in the group with acquired amputations trans-femoral amputation, knee disarticulation and trans-tibial amputation were seen with equal frequency (21%). In 40% of the children with congenital deficiency and in 8% of the children with acquired amputations the arm was also affected. Both legs were affected in 37% of the children with congenital deficiencies and in 8% of the children with acquired amputations. PMID- 10855435 TI - Children with congenital deficiencies or acquired amputations of the lower limbs: functional aspects. AB - The aim of the study was to evaluate the use of prostheses, some secondary complications and functional aspects among children who had a congenital leg deficiency or an acquired leg amputation. Rehabilitation physicians were asked to refer children, aged 1-18 years, with a leg deficiency or amputation. Mentally retarded children, children who had only had a toe amputated and children within one year after amputation were excluded. A total of 88 children were included; 64 with a congenital deficiency and 24 with an acquired amputation. In 25 of these 88, both legs were affected; 28 children also had an arm deficiency or amputation. A structured interview was held and the Child-HAQ assessed. Five (5) questions from the Child-HAQ, all relating to leg functions, were analysed. All but 7 children had had a prosthesis fitted, most (89%) using it for almost the entire day. In the children with congenital deficiencies, the first prosthesis had been fitted at an average age of approximately 18 months. Four (4) of the 7 children without prostheses used orthopaedic footwear. The 10 children with congenital deficiencies necessitating prostheses with articulated knees had the first knee of this type fitted at an average age of approximately 37 months. Forty-seven (47) of the 88 children had needed one or more (secondary) operations. In the children with congenital deficiencies, this was usually a conversion procedure, while the children with an acquired amputation had usually been operated on for osseous overgrowth. Twenty (20) of the 88 children experienced or had previously experienced phantom sensations, 5 children phantom pain. Skin problems were common. Most children (95%) were able to walk, most of them (93%) more than 100 m and 93% of the children aged 4 years or over were able to cycle. Most children (94%) aged 6 years or over were able to don and doff their prostheses independently. Some 90% of the children aged 4 years or over attended a normal primary or secondary school. Most (93%) of the children were able to take part in the physical education programme at school, although frequently (47%) with some degree of difficulty. The functional abilities of 88 Dutch children with congenital leg deficiencies or leg amputations were found to be generally satisfactory. Most of the children used prostheses in their daily activities. Secondary complications were, however, frequent. PMID- 10855436 TI - A study of technical changes to lower limb prostheses after initial fitting. AB - There is little published material in recent years about the use of lower limb prostheses in an elderly amputee population. In this study the authors were interested in the technical changes to lower limb prostheses after a first limb fitting procedure in a post-rehabilitation population in The Netherlands. The process of fitting a prosthesis and the technical changes to the artificial limb in the first year afterwards are studied. PMID- 10855437 TI - An evaluation of the Amputee Mobility Aid (AMA) early walking aid. AB - The most popular early walking aid (EWA) in the United Kingdom (UK) is the Pneumatic Post-Amputation Mobility aid or PPAM aid. A disadvantage of this device is that it does not allow a trans-tibial amputee to flex or extend the knee during walking. The Amputee Mobility Aid (AMA) was developed to allow knee movement, enabling trans-tibial amputees to practise a more natural gait. The benefits of using EWAs include early walking, reduction in post-operative oedema and improvement in patient morale. This pilot study investigated the pneumatic bag/stump interface pressures of the PPAM aid and the AMA. In addition, the range of motion of the knee on the amputated side and the mechanical knee of the AMA were compared. The AMA was found to have higher interface pressures than the PPAM aid during standing and similar pressures during supported walking. Subjects using the AMA did flex and extend their knee during walking but through a reduced range of motion. There were no significant differences between the angular movements of the AMA's mechanical knee and the patient's knee within it. PMID- 10855438 TI - Evaluation of polypropylene prostheses designed by the International Committee of the Red Cross for trans-tibial amputees. AB - Thirty-two (32) trained prosthesis users with 34 trans-tibial amputations, mostly due to war, were fitted with prostheses fabricated from polypropylene (PP) prosthetic components designed and manufactured by the International Committee of the Red Cross (ICRC). The patients were followed prospectively for 10 and 19 months. All but one patient had at least one other type of prosthesis to compare with. Twenty-eight (28) patients were satisfied with the PP prosthesis. Among these 23 found the PP prosthesis the preferred artificial limb, and one patient found the PP limb equal to the aluminium prosthesis previously in use. In 6/28 patients having an aluminium (ALU) prostheses this was found the best, and the 1 already mentioned found it equivalent to the new technology. In only 1/20 cases having an Automated Fabrication of Mobility Aids (AFMA) prosthesis available this was found the best. One (1) double-amputee found all three designs equal. Minor failures of the PP prostheses were encountered; in 4 cases small cracks in the hard socket; in 3 cases cracks of the cosmetic socket seam. From an overall prospect the PP technology can be recommended for trans-tibial prostheses. PMID- 10855439 TI - The calibration of ultrasound transducers used to monitor motion of the residual femur within a trans-femoral socket during gait. AB - During gait the motion of the residual femur within a trans-femoral socket may be estimated using video recorded data from two ultrasound transducers attached to the socket wall. This paper reviews possible measurement errors and identifies the magnitude of the inaccuracies. Inaccuracies due to equipment limitations and those due to human error are identified and quantified. Ranges of flexion/extension and abduction/adduction of the residual femur within the socket during gait have been estimated with a cumulative level of inaccuracy of <1 degree. PMID- 10855440 TI - Scientific validation of two commercial pressure sensor systems for prosthetic socket fit. AB - The concept of measuring pressure at the interface between the stump and the prosthetic socket could provide valuable information in the process of prosthetic socket fabrication, modification, and fit. Two systems, the Rincoe Socket Fitting System (SFS) and Tekscan's F-Socket Pressure Measurement System, have been commercially designed for in situ interface pressure measurement over the past decade. Their use is not common in prosthetic practice, perhaps due to questions of cost effectiveness and the difficulties of interpreting the data. Another concern is the use of sensors for pressure measurements in areas of high contour and complex geometries such as the stump. Before these systems can be used in a clinical setting, it is necessary to determine the reliability and accuracy of each system. In order to assess the clinical validity of the Rincoe SFS and F Socket systems, a series of trials was conducted to evaluate different aspects of sensor performance, namely; accuracy, hysteresis, drift and the effect of curvature. The sensors were subjected to tests in flatbed and custom-designed pressure vessels. Overall results indicated an accuracy error for the Rincoe SFS system of 25% (flatbed) and 33% (mould), with a corresponding 15% (flatbed) and 23% (mould) error in hysteresis, and 7% (flatbed) and 11% (mould) drift errors. The F-Socket system demonstrated an 8% (flatbed) and 11% (mould) accuracy errors, 42% (flatbed) and 24% (mould) hysteresis errors, and 12% (flatbed) and 33% (mould) drift errors. These findings indicate favourable results for the F-Socket Pressure Measurement System compared to the Rincoe Socket Fitting System with respect to its accuracy errors only. Nevertheless, it is the authors' belief that these systems are adequate in indicating areas of high pressure at the stump socket interface for clinical purposes, but both systems should be used with caution. PMID- 10855441 TI - A device to facilitate donning and cleaning of a roll-on socket in a trans-tibial amputee with a non-functional arm. AB - Rehabilitation of a person with a non-functional arm and a trans-tibial amputation with a short stump with fragile skin, multiple scars and a limited knee function presents a considerable problem. The best prosthesis was considered to be one with a roll-on socket. Donning and cleaning of the socket however could not be done by the patient himself and it was therefore necessary to develop a device to permit the patient to perform these tasks independently. A case history is presented and the new device described. PMID- 10855442 TI - Trans-tibial amputation for reflex sympathetic dystrophy: postoperative management. AB - This paper describes the experience with a trans-tibial amputation due to reflex sympathetic dystrophy. Because of lack of information about postoperative management in these cases, the medical history is provided together with a description of early mobilisation and technical information about prosthetic equipment. PMID- 10855443 TI - Internet as a meeting place concerning partial foot amputation. PMID- 10855444 TI - Management of peripheral arterial disease (PAD). TransAtlantic Inter-Society Consensus (TASC). PMID- 10855445 TI - Motion times, hand forces, and trunk kinematics when using material handling manipulators in short-distance transfers of moderate mass objects. AB - The risk of musculoskeletal injury associated with manual materials handling tasks has led in part to the use of material handling manipulators, yet there is limited empirical data to facilitate selection, design, and evaluation of these devices. A laboratory study of two types of mechanical manipulators (articulated arm and overhead hoist) was conducted of short-distance transfers of moderate loads, and the influence of various task parameters and transfer method on motion times, peak hand forces, and torso kinematics was obtained. Use of manipulators increased elemental motion times for symmetric sagittal plane transfers by 36 63%, and asymmetric transfers (in the frontal plane) by 62-115%, compared to similar transfers performed manually. Peak hand forces were significantly lower with both manipulators (40-50%), and approximately 10% higher for asymmetric versus symmetric transfers. Overall torso kinematics were grossly similar with and without a manipulator. These results suggest that for self-paced job tasks, moderate mass objects will be transferred slower over short distances and with lower levels of external (hand) forces when using mechanical aids. These simple effects, however, were influenced by object mass and transfer height. PMID- 10855446 TI - Computer use in cold environments. AB - This study addresses computer work in cold environments with the two-fold aim to explore conditions for such work, and to add knowledge about the use of fingers at data entry in the cold. Five workplaces were visited and work contents and use of computers are briefly described. Effects of work in the cold were in line with those mentioned in the literature, and manual lifting of heavy goods the most impairing activity. Subjects contended with strenuous working postures--holding the computers in their hands or arms--and with cold fingers. Individual fingering for data input was noted. Forefinger or a pen were used, and a pen is recommendable for input, either as a touch pen or, simply to press the keys. A supportive rack could be recommended for portable workstations. PMID- 10855447 TI - System to measure the use of the backrest in sitting-posture office tasks. AB - This paper presents an inexpensive and simple system to measure the way of use of the backrest. The system can be also used in field studies. It is based on a set of electrodes which, attached to the subject's back and the backrest, allows the contact area to be measured. A laboratory test was performed to validate the system. In the test the spontaneous use of the backrest in standard office chairs and tasks was studied. Four different types of backrest use have been detected, and it has been shown that they determine the lumbar curvature and pelvic inclination angles, as well as postural mobility. The comfort levels observed in the four types of backrest use were also different. Consequently, the system can be used as an indicator of back posture and comfort. PMID- 10855448 TI - Difference thresholds for automobile seat vibration. AB - Reductions in vehicle vibration that may contribute to improvements in overall vehicle ride could individually be too small to be detected by drivers or passengers. This study investigated the 'difference threshold' (the difference in magnitude between two stimuli which is just sufficient for their difference to be detected) required for a change in vehicle ride to be perceived and whether this was consistent with Weber's Law. Ten male and 10 female subjects sat in a car seat and were exposed to four different reproductions of the vertical vibration recorded on the seat of a car. Three of the stimuli had the same waveform recorded while the car traversed a tarmac surface. This waveform was reproduced using three different magnitudes of vibration at the seat: 0.2, 0.4 and 0.8 m s( 2) r.m.s. (Wb weighted). The other stimulus was recorded with the car traversing a 'pave' surface that gave a different waveform that was reproduced at a magnitude of 0.4 m s(-2) r.m.s. (Wb weighted). There were significant differences in the absolute difference thresholds measured using the same waveform at the three different magnitudes. When the difference thresholds were expressed in relative terms (the proportion by which two stimuli must differ in magnitude to be discriminated), the relative difference thresholds were approximately 13%, and independent of both the vibration magnitude and the vibration waveform. The results are therefore consistent with Weber's Law. No consistent differences were observed between the responses of male and female subjects. PMID- 10855449 TI - The application of ergonomics in rural development: a review. AB - The importance of ergonomics issues in rural development is highlighted in this paper. Some examples are given of the contribution that ergonomics has already made to industrially developing countries, cases which are mainly concentrated in the industrial sector. Key areas for future ergonomics research are identified, focusing on the needs of communities living and working in the agricultural sector where most of the population in the industrially developing world is located. PMID- 10855450 TI - A new approach to applying feedforward neural networks to the prediction of musculoskeletal disorder risk. AB - A new and improved method to feedforward neural network (FNN) development for application to data classification problems, such as the prediction of levels of low-back disorder (LBD) risk associated with industrial jobs, is presented. Background on FNN development for data classification is provided along with discussions of previous research and neighborhood (local) solution search methods for hard combinatorial problems. An analytical study is presented which compared prediction accuracy of a FNN based on an error-back propagation (EBP) algorithm with the accuracy of a FNN developed by considering results of local solution search (simulated annealing) for classifying industrial jobs as posing low or high risk for LBDs. The comparison demonstrated superior performance of the FNN generated using the new method. The architecture of this FNN included fewer input (predictor) variables and hidden neurons than the FNN developed based on the EBP algorithm. Independent variable selection methods and the phenomenon of 'overfitting' in FNN (and statistical model) generation for data classification are discussed. The results are supportive of the use of the new approach to FNN development for applications to musculoskeletal disorders and risk forecasting in other domains. PMID- 10855451 TI - Evaluating factors that influence hand-arm stress while operating an electric screwdriver. AB - The objective of this study is to evaluate the factors contributing toward hand arm stress while operating an electric screwdriver. Hand-arm stress was investigated in terms of individual finger force exertion, flexor digitorum EMG, and hand-transmitted vibration. Two activation modes (push and push plus trigger (P + T)), two preset shut-off torque levels (low and high) and three horizontal operating distances (far, middle, and near) were evaluated. Thirteen healthy male subjects drove screws into a horizontally mounted iron plate with pre-tapped screw holes using an in-line electric screwdriver in randomly ordered experimental combinations. The results indicate that using push-to-start mode at low torque level was better than the other combinations of activation mode x torque because it resulted in less hand-arm stress. In addition, the far distance level (33-45 cm away from the work table edge) caused greater stress than the middle and near distances, and hence is best avoided. While operating an in-line electrical screwdriver, the force contribution of the small finger was greatest, followed by the ring finger. The average force contributions of the index, middle, ring, and small fingers were 19, 25, 27, and 30%, respectively, while operating with push-to-start mode. PMID- 10855452 TI - Computer-aided ergonomics: a case study of incorporating ergonomics analyses into workplace design. AB - One of the primary goals of computer-aided ergonomics is to develop software tools that allow ergonomics information to be accessed at the earliest stages of design. This case study discusses a PC-based software program that allows a designer to quantify a worker's biomechanical risk for injury based on a proposed workplace design. The program couples an established software tool for biomechanical analysis, the Three-Dimensional Static Strength Prediction Program (3DSSPP), with a widely used computer-aided design software package, AutoCAD. The use of this "3DSSPP/AutoCAD interface" in the proactive analysis of an automotive assembly task is described and the results compared with an independent assessment using observations of workers performing the same task. Both studies yield similar conclusions, suggesting that proactive use of software such as the 3DSSPP/AutoCAD interface may be a valid tool in evaluating proposed workplace designs. In this context, issues in the analysis of workplace designs regarding the use of supporting ergonomic tools, assumptions, and posture selection are discussed. PMID- 10855453 TI - Impact of physical exposure on neck and upper limb disorders in female workers. AB - Physical workload [muscular load of the trapezius and infraspinatus muscles using electromyography (EMG), wrist positions and movements by electrogoniometers] and neck and upper limb disorders (from, for example, a physical examination) were studied in women with repetitive industrial work (n = 95) and referents (n = 74). The repetitive work displayed higher ratings for wrist movements, but not for EMG. The prevalences of neck, shoulder and wrist/hand disorders were elevated for women with repetitive work [age-adjusted prevalence odds ratios (PORs) 2.0-7.5]. For the left hand, high frequency of wrist movements (mean power frequency 0.53 Hz) was associated with a high prevalence of disorders (56%), as compared to low (0.28 Hz and 26%; POR 3.5). We found no consistent and significant effect of muscular load, on either neck or shoulder disorders. However, selection and other bias may have diminished our possibility to observe such effects. Psychosocial work environment factors were not confounding the results. Measurements of wrist movements may be used for identification of high-risk work tasks. PMID- 10855454 TI - Development of an ergonomic evaluation facility for Indian tractors. AB - The design of tractors manufactured in low-income countries like India has not changed much in the past two decades, especially from an ergonomic point of view. Moreover, in these countries tractors are used for transportation purposes in addition to farming operations. Therefore, the design criteria for these tractors need to be different from those in high-income countries. This paper describes the development of an ergonomic facility for improvement of tractor design. An ergonomic evaluation facility has been developed consisting of a work place envelope for the Indian population, a layout measuring device and an ergonomic rig. This facility can be used for comparative evaluation of the display and control layouts of different tractors in order to develop an optimum layout. The ergonomic rig has the facility to simulate the improved layout for subjective evaluation. PMID- 10855455 TI - Ergonomics, gerontechnology, and design for the home-environment. AB - An ergonomic approach could improve the quality of life and activities in daily living. Gerontechnology reduces the effects of age-related impairments with technological devices and particular design for the home-environment. Physiological decline with increasing age renders the daily activities at home more difficult. This paper highlights some "common sense" and specific design suggestions in the entrance and kitchen, aimed to increase the self-sufficiency of elderly people. We suggest that gerontechnology may have a particular role in the improvement of comfort and safety for aged people. PMID- 10855456 TI - Ride vibration on tractor-implement system. AB - India is the largest manufacturer of tractors in the world. They are used for primary and secondary tillage operations and as a means of transportation. Vibration in tractor driving can cause deafness and disorders of the spinal column and stomach. The effect of implements on tractor ride is not well understood in India. The present study was undertaken to quantify ride vibration of a low horsepower tractor-implement system. Tractor ride vibration levels have been measured at the person-seat interface along three mutually perpendicular axes, longitudinal, lateral and vertical, under different operating conditions. It was observed that the acceleration levels increased as forward speed of travel increased under most of the operating conditions. There was no conclusive difference in measured acceleration levels on a tar-macadam road and a farm road during transport mode. The measured ride vibration levels under different operating conditions were evaluated as per ISO 2631/1 (1985), Geneva, and BS 6841 (1987), London, standards. On the basis of this study, it is concluded that the exposure time for the tractor operator should not exceed 2.5 h during ploughing and harrowing operations. Increasing exposure time may cause severe discomfort, pain and injury. PMID- 10855457 TI - Anthropometric data of elderly people in Australia. AB - An anthropometric data set on 171 elderly people in Australia has been presented. The data set consists of 22 body dimensions relevant to design of living facilities, equipment and workplaces for the elderly people. The study was carried out in metropolitan Sydney, New South Wales, Australia. A comparative analysis with other populations indicates significant differences in the body dimensions. PMID- 10855458 TI - Body weight gain after administration of antipsychotic drugs: correlation with leptin, insulin and reproductive hormones. AB - Excessive body weight gain, hyperprolactinemia and low gonadal steroid serum levels are often observed during chronic administration of antipsychotic drugs (AP). Clinical and experimental findings suggest that leptin, the peptidic hormone involved in long-term body weight regulation, and reproductive hormones are interrelated. Therefore, we assessed circulating leptin levels in healthy, lean women (n = 12) and men (n = 7) before and after short-term administration of the AP sulpiride (SUL, 200 mg/day). In addition, we studied psychotic obese (n = 9) and lean women (n = 13) under chronic treatment with diverse AP. No significant weight changes were observed after SUL administration in healthy women--initial weight: 54.9+/-2.6 Kg; final weight: 55.04+/-2.6, NS. Leptin levels did not change either: 11.9+/-1.5 ng/ml. vs. 10.6+/-1.3, NS. By contrast, a small, but significant weight gain was found in SUL-treated men--60.6+/-1.9 Kg. vs. 61.3+/-2.1, p = 0.004. Leptin and insulin levels were significantly higher after SUL administration--leptin: 2.77+/-0.22 ng/ml. vs. 13.9+/-2.5, p=0.035; insulin: 3.59+/-0.17 mIU/ml vs. 8.81+/-0.81, p = 0.0001. In these subjects, leptin levels positively correlated with body weight change (p = 0.006), and serum prolactin change (p = 0.001). Obese psychotic women (Body Mass Index, BMI, Kg/m2 = 31.5+/-1.03) displayed higher leptin levels than non-obese psychotic women (BMI = 25.5+/-0.52): 26.8+/-4.8, vs. 12.8+/-3.4 ng/ml, p = 0.006. In these women, a significant positive correlation was found between leptin levels and BMI (p = 0.0001), and between leptin and basal insulin levels (p = 0.001). These results show that the expected circulating leptin elevation which is observed when body weight raises, is preserved in people treated with AP drugs. PMID- 10855459 TI - Cyclandelate in the treatment of patients with mild to moderate primary degenerative dementia of the Alzheimer type or vascular dementia: experience from a placebo controlled multi-center study. AB - A 24-week, double-blind, multi-center, randomised parallel group study compared the efficacy and safety of 800 mg bid cyclandelate with placebo in patients with mild to moderate dementia of primary degenerative or vascular origin. A total of 196 patients entered the study, 147 patients completed treatment in adherence with the protocol. Primary outcome measures were the cognitive score of the Alzheimer's Disease Assessment Scale (ADAS-Cog), the subscale Instrumental Activities of Daily Living of the Nurses' Observation Scale for Geriatric Patients (NOSGER-IADL) and the Clinical Global Impressions of Change (CGI-C). Safety assessments included adverse events, vital signs, ECG and clinical laboratory parameters. The primary efficacy results based on a multi-level responder analysis including ADAS-Cog, NOSGER-IADL and CGI-C failed to demonstrate statistical superiority of cyclandelate in comparison to placebo. The direction of changes favored cyclandelate in each of the variables, but the differences to placebo were small and varied considerably between patients and centers. Retrospective exploratory analyses suggested that efficacy of cyclandelate might be dependent on the severity of the disease. The treatment effects in favor of cyclandelate were statistically significant in the subgroup of moderately impaired patients (MMSE at baseline <18) for ADAS-Cog (delta = -4.0 points, p = 0.015) and CGI-C (delta = -0.4 points, p = 0.043) but not for NOSGER IADL (delta = -1.6 points, p = 0.059). When patients were stepwise selected for the severity of the disease according to ADAS-Cog at baseline (>15, >20, >25 points), statistical significance was reached for ADAS-Cog and NOSGER-IADL beginning with the step ADAS-Cog >20 points: delta ADAS-Cog = -3.9 points, p = 0.044; delta NOSGER-IADL = -1.0, p = 0.023. The treatment differences increased further with the step ADAS-Cog >25 points: delta ADAS-Cog = -7.0 points, p = 0.008; delta NOSGER-IADL = -1.7, p = 0.003. Treatment differences in CGI-C increased marginally with the stepwise selection but did not reach statistical significance. The drug was safe and well tolerated. PMID- 10855460 TI - How common is violence in schizophrenia despite neuroleptic treatment? AB - There is a large body of literature about increased figures for violence in schizophrenic in-patients and out-patients. The therapeutic efficacy of neuroleptics in coping with violent behaviour in schizophrenics is well documented. However, little data is available about the treatment given to schizophrenics who behave violently. We performed an extensive chart review in an unselected sample of n = 138 patients with schizophrenic or schizoaffective disorder (ICD10) and first admission between 1990 and 1993, including in-patient episodes in the first two years after first admission. Staff records were reviewed for all notes on aggressive behaviour (threats, physical aggression against persons and objects, selfdirected aggression) and coercive measures. For each incident, the number of days after the beginning of neuroleptic treatment was coded. 258 inpatient treatment periods with an average length of 78.5 days were evaluated; 226 lasted more than one week. 142 aggressive incidents were observed, of these 66% within the first week of neuroleptic treatment, 9% within the second. The day-by-day decline of aggressive incidents after the start of neuroleptic treatment was highly significant (trend-test: Spearman rank correlation r=0.964; df=5; t<-8.1; p<0.001). The results support the assumption that the increased figures for violence by schizophrenics are, at least in part, due to the lack of adequate treatment. PMID- 10855461 TI - A prospective PMS study to validate the sensitivity for change of the D-scale in advanced stages of dementia using the NMDA-antagonist memantine. AB - The present postmarketing surveillance (PMS) study is the first large scale systematic and prospective clinical trial of pharmacotherapeutic intervention in advanced stages of dementia. Within a validation program this study aimed at demonstrating the sensitivity of the D-Scale of change (DS-C) for measuring ADL function. Efficacy of treatment with the NMDA-antagonist memantine was investigated in 531 patients with advanced dementia employing a parallel group design that stratified patient cohorts by severity according to GDS stages (Reisberg, 1992). Efficacy was determined on two independent levels: by the assessment of the physicians' Clinical Global Impression of Change (CGI-C) at the end of a 6-week observation period, and by the assessment of change in elementary ADL-functions by the caregivers using the D-Scale-of-Change. With the D-Scale-of Change the caregivers can assess a change in broad functional items, i.e. cognitive and motor functions and also elementary functions of daily life. The effect size of this improvement increased constantly during the observation period. Even in patients of GDS stage 7 an improvement could be measured. These results were also seen by the physicians, who recorded an overall clinical improvement in 75.5% of the patients after 6 weeks. Tolerability evaluations resulted in the ratings "very well" by 59.5% or "well" by 35.0% of the patients. No serious adverse drug reactions occurred. A correlation analysis demonstrated a high congruency of both assessments. Furthermore, the observed time course of these improvements paralleled with the time course of symptomatic benefit by effects of memantine that had been repeatedly demonstrated in randomised, double blind, placebo-controlled studies in mild to moderate dementia. Together with the evaluation of the scale-properties of the D-Scale for assessment of severity the D-Scale and the D-Scale-of-Change can be regarded as validated. PMID- 10855462 TI - A comparison of the effects of clozapine and olanzapine on the EEG in patients with schizophrenia. AB - Clozapine is known to induce epileptic seizures and changes in EEG-patterns, including slowing and the appearance of epileptiform activity. Olanzapine, a new antipsychotic drug, shares many pharmacological and clinical properties with clozapine. However, in patients treated with olanzapine, no case of seizure induction has been reported so far, and the EEG has not been studied systematically. We examined the EEGs of patients with schizophrenia treated with either olanzapine (N = 9) or clozapine (N = 9) prior to medication and 3 to 7 weeks afterwards. Clozapine induced significant EEG slowing present in 78% of the patients, and definite epileptiform activity appeared in 33%. Olanzapine also induced significant EEG slowing, but less frequently (in 44% of the patients) and less pronounced than clozapine. Olanzapine had no significant effect an epileptiform activity, but in one patient, an isolated sharp/slow-wave complex was observed. These preliminary data suggest that olanzapine induces EEG slowing to a lower extent than clozapine. Olanzapine's possible effect an the seizure threshold deserves further attention. PMID- 10855463 TI - Marked increase of venlafaxine enantiomer concentrations as a consequence of metabolic interactions: a case report. AB - On three occasions, unusually high trough plasma concentrations of venlafaxine were measured in a patient phenotyped and genotyped as being an extensive CYP2D6 metabolizer and receiving 450 mg/day of venlafaxine and multiple comedications. Values of 1.54 and of 0.60 mg/l of venlafaxine and O-desmethylvenlafaxine, respectively, were determined in the first blood sample, giving an unusually high venlafaxine to O-desmethylvenlafaxine ratio. This suggests an impaired metabolism of venlafaxine to O-desmethylvenlafaxine, and is most likely due to metabolic interactions with mianserin (240 mg/day) and propranolol (40 mg/day). Concentration of (S)-venlafaxine measured in this blood sample was almost twice as high as (R)-venlafaxine ((S)/(R) ratio: 1.94). At the second blood sampling, after addition of thioridazine (260 mg/day), which is a strong CYP2D6 inhibitor, concentrations of venlafaxine were further increased (2.76 mg/l), and concentrations of O-desmethylvenlafaxine decreased (0.22 mg/l). A decrease of the (S)/(R)-venlafaxine ratio (-20%) suggests a possible stereoselectivity towards the (R)-enantiomer of the enzyme(s) involved in venlafaxine O-demethylation at these high venlafaxine concentrations. At the third blood sampling, after interruption of thioridazine, concentrations of venlafaxine and O desmethylvenlafaxine were similar to those measured in the first blood sample. This case report shows the importance of performing studies on the effects of either genetically determined or acquired deficiency of metabolism on the kinetics of venlafaxine. PMID- 10855464 TI - Association of paroxetine with suicide attempt in obsessive-compulsive disorder. PMID- 10855465 TI - Galactorrhea after paroxetine treatment. PMID- 10855466 TI - MR imaging of clear cell sarcoma (malignant melanoma of the soft parts): a multicenter correlative MRI-pathology study of 21 cases and literature review. AB - OBJECTIVE: To evaluate MR imaging and pathology findings in order to define the characteristic features of clear cell sarcoma of the soft tissues (malignant melanoma of the soft parts). DESIGN AND PATIENTS: MR examinations of 21 patients with histologically proven clear cell sarcoma of the musculoskeletal system were retrospectively reviewed and assessed for shape, homogeneity, delineation, signal intensities on T1- and T2-weighted images, contrast enhancement, relationship with adjacent fascia or tendon, secondary bone involvement, and intratumoral necrosis. In 19 cases the pathology findings were available for review and for a comparative MR-pathology study. RESULTS: On T1-weighted images, lesions were isointense (n=3), hypointense (n=7) or slightly hyperintense to muscle (n=11). Immunohistochemical examination was performed in 17 patients. All 17 specimens showed positivity for HMB-45 antibody. In nine of 11 lesions with slightly increased signal intensity on T1-weighted images, a correlative MR imaging pathology study was possible. All nine were positive to HMB-45 antibody. CONCLUSIONS: Clear cell sarcoma of the musculoskeletal system often has a benign looking appearance on MR images. In up to 52% of patients, this lesion with melanocytic differentiation has slightly increased signal intensity on T1 weighted images compared with muscle. As the presence of this relative higher signal intensity on T1-weighted images is rather specific for tumors displaying melanocytic differentiation, radiologists should familiarize themselves with this rare entity and include it in their differential diagnosis when confronted with a well-defined, homogeneous, strongly enhancing mass with slightly higher signal intensity compared with muscle on native T1-weighted images. PMID- 10855467 TI - A pilot study using magnetic resonance imaging to determine the pattern of muscle group recruitment by rowers with different levels of experience. AB - OBJECTIVE: To determine whether it was possible using magnetic resonance imaging (MRI) to define the pattern of muscle recruitment in a specific sport (rowing) and to see whether there were differences in this pattern between athletes of different experience. DESIGN AND METHOD: It has been shown that during vigorous exercise the water content of muscle increases transiently. This can be observed using MRI, where the prolonged T2 relaxation time of muscle can be demonstrated. In this study we have exploited the increase in signal seen in exercised muscle on short TI inversion recovery (STIR) sequences, to show how rowers of different experience use different muscle groups. RESULTS: We have shown that trained athletes recruit selected muscle groups to carry out a given task, which they carry out more efficiently than untrained or less experienced athletes. CONCLUSION: We have provided the basis of potential research to refine training methods, in order to develop specific muscle groups in athletes, in the hope of achieving a higher level of performance at an earlier stage in their training. We have also defined a technique that may be of clinical value in cases of muscle dysfunction. PMID- 10855468 TI - Posterior glenoid rim deficiency in recurrent (atraumatic) posterior shoulder instability. AB - OBJECTIVE: To assess the shape of the posterior glenoid rim in patients with recurrent (atraumatic) posterior instability. DESIGN AND PATIENTS: CT examinations of 15 shoulders with recurrent (atraumatic) posterior instability were reviewed in masked fashion with regard to abnormalities of the glenoid shape, specifically of its posterior rim. The glenoid version was also assessed. The findings were compared with the findings in 15 shoulders with recurrent anterior shoulder instability and 15 shoulders without instability. For all patients, surgical correlation was available. RESULTS: Fourteen of the 15 (93%) shoulders with recurrent (atraumatic) posterior shoulder instability had a deficiency of the posteroinferior glenoid rim. In patients with recurrent anterior instability or stable shoulders such deficiencies were less common (60% and 73%, respectively). The craniocaudal length of the deficiencies was largest in patients with posterior instability. When a posteroinferior deficiency with a craniocaudal length of 12 mm or more was defined as abnormal, sensitivity and specificity for diagnosing recurrent (atraumatic) posterior instability were 86.7% and 83.3%, respectively. There was a statistically significant difference in glenoid version between shoulders with posterior instability and stable shoulders (P=0.01). CONCLUSION: Recurrent (atraumatic) posterior shoulder instability should be considered in patients with a bony deficiency of the posteroinferior glenoid rim with a craniocaudal length of more than 12 mm. PMID- 10855469 TI - Percutaneous Ethibloc injection in aneurysmal bone cysts. AB - OBJECTIVE: To investigate whether the injection of Ethibloc into aneurysmal bone cysts can be an effective treatment modality. DESIGN AND PATIENTS: Ethibloc is an alcoholic solution of zein (corn protein) which has thrombogenic and fibrogenic properties. Ten patients with aneurysmal bone cysts were treated with CT-guided percutaneous injection of Ethibloc into the cyst cavity. Ethibloc injection was the primary treatment in five patients. Four patients had recurrence following previous curettage and bone grafting and one patient had not responded to injection into the lesion of autologous iliac crest bone marrow aspirate. Three patients needed a second injection. The median follow-up was 27 (6-60) months. RESULTS AND CONCLUSION: Symptoms were relieved in all patients. At imaging, seven patients had resolution of the lesion and three had partial response at the most recent follow-up. Complications consisted of a local transitory inflammatory reaction in two patients and an aseptic abscess in one patient. This relatively simple, minimally invasive procedure makes an operation unnecessary by stopping the expansion of the cyst and inducing endosteal new bone formation. This technique may be used as the primary management of aneurysmal bone cysts excluding spinal lesions. PMID- 10855470 TI - Effect of low back posture on the morphology of the spinal canal. AB - OBJECTIVE: To define the possible mechanism of posture-dependent symptoms of spinal stenosis by measuring the effect of low back posture on morphologic changes of the intervertebral discs and spinal canal in healthy young people. DESIGN: Twenty healthy young volunteers underwent magnetic resonance imaging while supine with their spine in neutral, flexed, extended, and right and left rotational positions. The axial MR images at the middle of the intervertebral discs of L3-4 and L4-5 were analyzed to measure the difference in the size and shape of the intervertebral discs and spinal canal in each posture. RESULTS: Extension or rotation decreased the sagittal diameters and cross-sectional areas of the dural sac and spinal canal and increased the thickness of the ligamentum flavum, whereas flexion had the opposite effects. The gap between the convex posterior disc margin and the anterior margin of the facet joint on each side, represented as the subarticular sagittal diameter, increased with flexion and decreased with extension or rotation. The direction of rotation did not result in asymmetry of the subarticular sagittal diameter, but right rotation caused thickening of the right ligamentum flavum, and vice versa. The shape and dimensions of the disc did not change significantly according to the positions of the low back. CONCLUSIONS: With extension or rotation, the thickness of the ligamentum flavum increased and the posterior margin of the intervertebral disc was approximated to the facet joint without any change in shape and size of the disc. These phenomena result in a decrease in the size of the spinal canal and dural sac in extension or rotation postures in young healthy people without disc degeneration, and may explain the posture-dependent symptom of spinal stenosis. PMID- 10855471 TI - Intraosseous schwannoma of the metacarpal. AB - Intraosseous schwannoma (neurilemmoma) is an extremely rare, benign neoplasm, constituting less than 0.2% of primary bone tumors. It infrequently involves the bones of the hand. We present a case of intraosseous neurilemmoma of the metacarpal. PMID- 10855472 TI - Erdheim-Chester disease with intramuscular lipogranuloma. AB - We report on a rare manifestation of Erdheim-Chester disease with intramuscular lipogranuloma. The patient was a 66-year-old man who noted a soft tissue mass in the right quadriceps femoris muscle. Radiographs revealed symmetrical osteosclerosis in the diametaphysis of both femora and tibiae. An open biopsy revealed a proliferation of lipid-laden histiocytes in the femoral bone marrow and the quadriceps femoris muscle. To our knowledge, this is the second case of Erdheim-Chester disease involving muscle. PMID- 10855473 TI - Hemangioma of bone arising in the ulna: imaging findings with emphasis on MR. AB - An 18-year-old woman presented with left elbow joint pain. Radiographs and computed tomographic scan showed a well-defined osteolytic lesion of the left ulna associated with a honeycomb appearance on the radiographs. Magnetic resonance images showed intermediate signal intensity on T1-weighted images and mixed intermediate and high signal intensities on T2-weighted images. Only the periphery of the lesion enhanced with intravenously injected gadolinium diethylenetriamine pentaacetic acid. The lesion was curetted to avoid pathologic fracture, and a histologic diagnosis of cavernous hemangioma of bone was made. Hemangioma involving the ulna is rare, but should be included in the differential diagnosis of a radiographic osteolytic lesion with a honeycomb appearance. PMID- 10855474 TI - MR appearances in a case of femoral ecchinococcosis. AB - Magnetic resonance (MR) findings of a rare case of osseous echinococcosis with involvement of the femur are described. Attention is drawn to the typical MR appearances and therapeutic management. PMID- 10855475 TI - Jansen type of spondylometaphyseal dysplasia. AB - Metaphyseal dysplasia, type Jansen (JMD), is a rare skeletal dysplasia with characteristic radiographic abnormalities. Of the various types of metaphyseal dysplasia, JMD shows the most severe alteration in metaphyseal architecture. All of the long tubular bones, including those of the hands and feet, show metaphyseal irregularity with a fragmented appearance and slight widening. The adjacent physes are abnormally widened, while the epiphyses tend to be slightly enlarged, rounded but otherwise normal. The spine in infancy and childhood usually appears normal. This report describes a young girl with metaphyseal changes typical of JMD except for the hands and feet, which appeared normal. She also showed very unusual abnormalities of the spine. This appears, therefore, to represent a unique osteochondrodysplasia for which we propose the term spondylometaphyseal dysplasia, type Jansen. PMID- 10855476 TI - Congenital transverse deficiency of the tibia and fibula: a report of two cases. AB - We report two similar, but unrelated, patients with congenital bilateral partial deficiencies of the tibia and fibula associated with intact feet. In both patients, the tibia and fibula were absent on initial radiographs, while the femur and the tarsal bones were well developed and there was bilateral teratologic dislocation of the hips. Ultrasound and magnetic resonance imaging (MRI) studies suggested the presence of cartilaginous remnants of the tibia and fibula. There were multidirectional instabilities in the knees and ankles. The clinical and radiological features of these cases are distinct from those of congenital longitudinal deficiency of the tibia, in which the fibula is always preserved, and from longitudinal deficiency of the fibula, in which the tibia is present and the foot is usually involved. We suggest that the bilateral partial deficiencies of the tibia and fibula associated with the intact foot and teratologic dislocation of the hips is a single-entity disorder, possibly categorized as an intercalary transverse deficiency of the lower limb. PMID- 10855477 TI - Introduction: How can research discover how to do psychotherapy? AB - It is almost exceptional that psychotherapy researchers set out to answer questions like these: What are some new and better ways of doing actual in session psychotherapeutic work? What are some new and better changes that psychotherapy can help bring about, and what are some new and better ways of helping to bring them about? The purpose of this special series of papers is to provide a forum for researchers to tell how to do research to answer those questions, i.e., to present practical and useful methodologies, designs, and strategies that are helpful in answering these questions. PMID- 10855478 TI - Traditional research and psychotherapy practice. AB - Practitioners of psychotherapy hold different assumptions about the predictability and constancy of human actions than those required by the inferential logic employed in traditional research. Psychotherapy requires the use of judgment by therapists that goes beyond the direct application of generalized knowledge. Judgment is sensitive to the particular, contextual, and changing situation characteristic of therapy practice. Therapists decisions about what to do and say are informed by, but go beyond, that which is available in their knowledge bases. Therapists' knowledge bases include their own, as well as vicarious, experiences. Therapists can parse traditional research into descriptive and inferential components, and by treating the descriptive component as vicarious experiences, they can incorporate it into their knowledge base. PMID- 10855479 TI - Discovering markers of assimilation stages: the fear-of-losing-control marker. AB - One way research can discover how to do successful psychotherapy is by intensive examination of the process of psychological change as it occurs within single clients. The assimilation model describes a series of stages through which clients' problematic experiences are hypothesized to progress during successful psychotherapy. Markers of assimilation stages are recognizable types of events in psychotherapy discourse that are linked empirically and theoretically to those stages. Validated markers may be clinically useful signs of clients' progress and current therapeutic requirements, as well as useful tools for research on the assimilation model. We describe how we discovered the "fear-of-losing-control" marker, which signals the emergence of unwanted thoughts, an early stage in the hypothesized sequence. Analysis of examples of the fear-of-losing-of-control marker suggests mechanisms by which client progression may be suspended between warding off and exploration of problematic experiences. PMID- 10855480 TI - Methodological pluralism: a framework for psychotherapy research. AB - A new level of sophistication is advocated for psychotherapy research- methodological pluralism. This pluralism recognizes the number of unexamined and untested assumptions that are embedded within any method, including traditional scientific methods. These assumptions restrict the questions that can be asked and the variables that can be tested before any investigation of psychotherapy has occurred. Such premature closure is ultimately unscientific and hinders truly creative investigations of psychotherapy. Consequently, a framework is described that allows multiple methods--with multiple underlying philosophies--to complement one another in psychotherapy research. Five essential benefits of this pluralistic framework are shown: advancement of science, increased objectivity, a level playing field, greater scientific freedom, and an end to the burgeoning "method wars" of psychotherapy research. PMID- 10855481 TI - Ideal psychotherapy research: a study of significant change processes. AB - The research approach I recommend involves the intensive analysis of concrete change performances using both intensive observation and measurement of in session behavior, as well as the investigation of participants' subjective recall of their experience. The goal is to build models of client-change processes and the therapist interventions that set these in motion. Examples of research efforts to study the allowing of emotional pain, the process of interruption of emotion, and the process of resolution of hopelessness are given. PMID- 10855482 TI - How to do discovery-oriented psychotherapy research. AB - A discovery-oriented approach to psychotherapy research relies on discovery oriented research questions. Four such questions are illustrated, together with the logistics of this approach to research, and the practical working steps in carrying out a program of discovery-oriented research. A case is made for the superior elegance of the discovery-oriented approach to psychotherapy research. PMID- 10855483 TI - Getting our act together: lessons on meaningful psychotherapy research from the philosophy of science. AB - Psychotherapy research should concentrate on building up a scientifically validated, theoretical knowledge base by means of disciplined empirical research. The normal nested relationship between technology, theory, and research has not been the norm in the world of psychotherapy. Psychotherapy researchers should learn from the history of science and concentrate on building basic theory. Investigations into the causal relations underpinning psychotherapy is the best way that research can help us do psychotherapy better. These investigations must conform to the canons of inductive reasoning. Conventional use of clinical data to underwrite psychotherapeutic theory is vulnerable to Grunbaum's critique. A research proposal based on Langs' communicative approach to psychotherapy is presented predicting measurable unconscious responses to brief, time-limited psychotherapy. PMID- 10855484 TI - Improving psychotherapeutic services via randomized clinical trials, treatment manuals, and component analysis designs. AB - In order to advance the practice of psychotherapy and to discover how to improve on the benefits of psychotherapy, we propose that practitioners and researchers should rely more on the results of randomized clinical trials research. On the practice side, the routine use by practitioners of manuals for empirically supported treatments holds the promise of providing scientifically based clinical services to clients. On the research side, there is much to be gained in using component analysis designs to determine the effective elements of a treatment that has been established as empirically supported in randomized clinical trials. Once these elements are identified, the next version of the treatment manual then could highlight the crucial aspects of treatment to be included in clinical practice. thus further refining scientifically based psychotherapeutic services. PMID- 10855485 TI - How valuable are psychotherapy experiments?: The idiographic problem. AB - The idiographic problem is said to arise because psychotherapy experiments study groups of subjects, whereas the clinician wants useful etiological or treatment information concerning a single, unique client. One response is to place little value on psychotherapy experiments and to argue for nonexperimental methods for obtaining clinical knowledge. It is argued here that these alternative methods are defective and that the idiographic problem can be solved without renouncing experimental methods. PMID- 10855486 TI - Multiple-case depth research: bringing experience-near closer. AB - The term "experience-near" has become associated with a variety of alternatives to mainstream clinical research. These alternatives converge on one basic methodological goal-faithfulness to clinical phenomena as lived. This article presents one approach to lived clinical phenomena that I term multiple-case depth research or MCDR. MCDR is a novel and highly sensitive methodology that combines both in-depth case investigation with experiential therapeutic principles. To illustrate the power of MCDR, I present a hypothetical process and outcome study involving three client cohorts (those who undergo respectively cognitive behavioral therapy, intersubjective psychoanalytic therapy, and existential humanistic therapy). I detail the structure of this hypothetical study, the steps by which it proceeds, and the yield that it portends. I conclude that, if conducted properly, MCDR can provide rich, valid, and unprecedented insight into effective psychotherapy. PMID- 10855487 TI - MMPI-2 with Mexican psychiatric inpatients: basic and content scales. AB - This paper presents the results of a study of the Mexican Spanish version of the MMPI-2 with a clinical sample of 233 patients who were diagnosed as having psychological disturbances or personality disorders, according to DSM-III R criteria. Inpatient scores were obtained from four psychiatric hospitals, located in Mexico City. The scores of the patients were compared with those of Mexican college students, which is the largest Mexican normative sample collected to date, consisting of 813 men and 1,137 women. Results of this study show that the MMPI-2 can accurately differentiate between normal and non-normal groups in Mexican populations and demonstrate that the inventory maintains its construct validity in this clinical sample. PMID- 10855488 TI - Differential patterns of responding among three groups of chronic, psychotic, forensic outpatients. AB - Elements of response style were examined among three groups of chronic, psychotic, forensic patients: paranoid schizophrenics (N = 89); undifferentiated disorganized schizophrenics (N = 38); and schizoaffective patients (N = 53). Forensic patients with elevated MMPI-2 L Scales produced increased percentages of Pure Form (F%) on the Rorschach. A similar relationship occurred when the Rorschach was used as the independent measure. Schizoaffective patients reported more psychotic symptoms on the MMPI-2 and lower F% (Rorschach) than both schizophrenic groups. Although undifferentiated schizophrenics evidenced the most psychopathology on the Rorschach (impaired reality testing and perceptual accuracy disturbance), all three groups produced lower than expected frequencies for Rorschach variables commonly associated with thought disorder and poor reality testing (Exner, 1995b). The clinical importance of using the MMPI-2 and Rorschach in tandem with forensic psychiatric patients is discussed. Our empirical findings suggest the need for forensic evaluators to consider the important relationship between psychiatric diagnosis and response style (defensiveness, denial, illness chronicity, medications, and concurrent Axis II psychopathology) when interpreting often-constricted psychological testing protocols in chronic forensic patient populations. PMID- 10855489 TI - Patient Impairment Lexicon: a validation study. AB - Growth of managed behavioral health organizations (MBHOs) has increased the need for a standardized diagnostic language. The Patient Impairment Lexicon (PIL; Goodman, Brown, & Deitz, 1992, 1996) is intended to address this need. Augmenting previous psychometric assessment (Klewicki, Bjorck, Leucht, & Goodman, 1998), the current study evaluated the PIL's construct validity. Sixteen raters completed impairment inventories; diagnoses based on the Diagnostic and Statistical Manual of Mental Disorders, Third Edition, Revised (DSM-III-R; American Psychiatric Association, 1987); and independent measures of psychiatric functioning for patients in 20 vignettes. Overall rater accuracy was significantly higher for PIL impairments than for diagnoses. As predicted, there were positive correlations between PIL impairments and psychiatric functioning for all 20 vignettes, 11 of which were significant. Results remained significant after controlling variance due to: (a) raters' past experience with similar patients and (b) vignette imaginability. Findings are discussed in terms of MBHO applications and future PIL research. PMID- 10855490 TI - Hanging on to your homolog: the roles of pairing, synapsis and recombination in the maintenance of homolog adhesion. AB - Homologous chromosomes initially undergo weak alignments that bring homologous sequences into register during meiosis. These alignments can be facilitated by two types of mechanisms: interstitial homology searches and telomere-telomere alignments. As prophase (and chromatin compaction) proceeds, these initial pairings or alignments need to be stabilized. In at least some organisms, such as Saccharomyces cerevisiae and S. pombe, these pairings can apparently be maintained by the creation of recombination intermediates. In contrast, synapsis during zygotene may be able to facilitate and/or maintain chromosome pairing even in the absence of exchange in several higher organisms. It thus seems possible that the synaptonemal complex plays a role both in maintaining homolog adhesion during meiotic prophase and, more speculatively, in facilitating meiotic exchange. PMID- 10855491 TI - Sister chromatid cohesion and recombination in meiosis. AB - Sister chromatids are associated from their formation until their disjunction. Cohesion between sister chromatids is provided by protein complexes, of which some components are conserved across the kingdoms and between the mitotic and meiotic cell cycles. Sister chromatid cohesion is intimately linked to other aspects of chromosome behaviour and metabolism, in particular chromosome condensation, recombination and segregation. Recombination, sister chromatid cohesion and the relation between the two processes must be regulated differently in mitosis and meiosis. In meiosis, cohesion and recombination are modified in such a way that reciprocal exchange and reductional segregation of homologous chromosomes are ensured. PMID- 10855492 TI - Destruction of the securin Pds1p occurs at the onset of anaphase during both meiotic divisions in yeast. AB - Sister chromatid cohesion is established during DNA replication and depends on a multiprotein complex called cohesin. At the onset of anaphase the cohesive structures that hold sisters together must be destroyed to allow segregation of sisters. In the budding yeast Saccharomyces cerevisiae loss of sister chromatid cohesion depends on a separating protein (separin) called Esp1. At the metaphase to anaphase transition, separin is activated by proteolysis of its inhibitory subunit (securin) called Pds1. This process is mediated by the anaphase promoting complex and an accessory protein Cdc20. In meiosis a single round of DNA replication is followed by two successive rounds of segregation. Thus loss of cohesion is spun out over two divisions. By studying the mechanisms that initiate anaphase in meiotic division we show that the yeast securin Pds1p is present in meiotic nuclei and is destroyed at the onset of each meiotic division. We also show that securin destruction depends on Cdc20p which accumulates within nuclei around the time of Pds1p's disappearance. PMID- 10855493 TI - Meiotic sister chromatid cohesion in holocentric sex chromosomes of three heteropteran species is maintained in absence of axial elements. AB - It has been suggested that in species with monocentric chromosomes axial element (AE) components may be responsible for sister chromatid cohesion during meiosis. To test this hypothesis in species with holocentric chromosomes we selected three heteropteran species with different sex-determining mechanisms. We observed in surface-spreads and sections using transmission electron microscopy that the univalent sex chromosomes form neither AEs nor synaptonemal complexes (SCs) during pachytene. We also found that a polyclonal antibody recognizing SCP3/Cor1, a protein present at AEs and SC lateral elements of rodents, labels the autosomal SCs but not AEs or SC stretches corresponding to the sex chromosomes. Cytological analysis of the segregational behaviour of the sex univalents demonstrates that although these chromosomes segregate equationally during anaphase I they never show precocious separation of sister chromatids during late prophase I or metaphase I. These results suggest that AEs are not responsible for sister cohesion in sex chromosomes. The segregational behaviour of these chromosomes during both meiotic divisions also indicates that different achiasmate modes of chromosome association exist in heteropteran species. PMID- 10855494 TI - mei-41 is required for precocious anaphase in Drosophila females. AB - This paper reports on a new role for mei-41 in cell cycle control during meiosis. This function is revealed by the requirement of mei-41 for the precocious anaphase observed in crossover-defective mutants. Normally in Drosophila oocytes, tension on the meiotic spindle causes a metaphase I arrest. This tension results because crossovers, and the resulting chiasmata, hold homologs together that are being pulled by kinetochore microtobules toward opposite spindle poles. In the absence of tension, such as in a recombination-defective mutant, metaphase arrest is not observed and meiosis proceeds through the two divisions. Here we show that in some recombination-defective mutants, the precocious anaphase requires the mei 41 gene product. For example, metaphase arrest is not observed in mei-218 mutants because of the severe reduction in crossing over. In mei-41 mei-218 double mutants, however, metaphase arrest was restored. The effect of mei-41 is dependent on double-strand break formation. Thus, in mutants that fail to initiate meiotic recombination the absence of mei-41 has no effect. PMID- 10855495 TI - Cloning and characterization of the Kluyveromyces lactis homologs of the Saccharomyces cerevisiae RED1 and HOP1 genes. AB - The synaptonemal complex (SC) is a meiosis-specific proteinaceous structure that holds homologous chromosomes close together along their length during the pachytene stage of meiotic prophase. The SC is observed in sexually reproducing fungi, plants and animals and is highly conserved at the cytological level. Despite this striking conservation of structure, however, the known protein components of the SC do not appear to be highly conserved across species. In Saccharomyces cerevisiae, the products of the RED1 and HOP1 genes are associated with the lateral elements of the SC. Using a functional complementation strategy, we have isolated homologs of these genes from the related yeast, Kluyveromyces lactis. The predicted K. lactis Red1 protein is 26% identical to the S. cerevisiae Red1 protein, and the K. lactis Hop1 protein is 40% identical to the S. cerevisiae Hop1 protein. The K. lactis RED1 gene fully complements the S. cerevisiae red1 mutant, both when overexpressed and when present in two copies in a diploid. However, the K. lactis HOP1 gene complements a hop1 mutant poorly when overproduced and not at all when present in two copies in a diploid. Unlike the S. cerevisiae RED1 gene, the K. lactis RED1 contains an intron; the transcript of the K. lactis gene is efficiently spliced during meiosis in S. cerevisiae. PMID- 10855496 TI - A homologue of the yeast HOP1 gene is inactivated in the Arabidopsis meiotic mutant asy1. AB - Synapsis of homologous chromosomes is a key event in meiosis as it is essential for normal chromosome segregation and is implicated in the regulation of crossover frequency. We have previously reported the identification and cytological characterisation of a T-DNA-tagged asynaptic mutant of Arabidopsis thaliana. We have demonstrated that this mutant, asy1, is defective in meiosis in both males and females. Cloning and nucleotide sequencing of the ASY1 gene has revealed that it encodes a polypeptide of 596 amino acids that exhibits similarity to the HOP1 gene of Saccharomyces cerevisiae, which is known to encode a protein essential for synaptonemal complex assembly and normal synapsis. Expression studies indicate that, in common with a number of other Arabidopsis meiotic genes, ASY1 exhibits low-level expression in a range of plant tissues. Southern analysis coupled with database searching has resulted in the identification of an ASY1 homologue, ASY2. Although asy1 exhibits a strong asynaptic phenotype, a residual low level of synapsis indicates that ASY1 and ASY2 may exhibit a low degree of functional redundancy. PMID- 10855497 TI - Spo1, a phospholipase B homolog, is required for spindle pole body duplication during meiosis in Saccharomyces cerevisiae. AB - The SPO1 gene was cloned and shown to encode an early meiotic transcript specifying a nuclear protein with extensive similarity to fungal and vertebrate phospholipase enzymes. Alteration of a conserved serine residue in the putative phospholipase active site, and presence of the spo1-1 temperature-sensitive mutation, which resides near this site, each result in loss of SPO1 function. The phenotype of a complete deletion indicates that SPO1 is dispensable for vegetative growth, premeiotic DNA synthesis and meiotic recombination. In contrast, it is required for Meiosis I (MI) and Meiosis II (MII) chromosome segregation and spore formation. In a null mutant approximately 75% of cells arrest early at MI spindle pole body (SPB) duplication, approximately 20% arrest at MII, and approximately 5% arrest at spore formation. Progression beyond the first arrest point suggests the existence of functions partially redundant to Spo1 and that Spo1 is required at multiple stages. At present SPO1 is the only known gene required for SPB duplication in meiosis but not in mitosis. Its product may thus play a regulatory (rather than a structural) role in SPB function. The transcriptional program in the spo1 null is similar to the wild type early in meiosis but is significantly delayed at later stages of sporulation. A single gene, CWP1, was recovered as a multicopy suppressor of the spo1 null. CWP1 encodes a cell wall protein with a glycolipid moiety. We propose that, when modified by other lipases, this moiety may substitute for the product(s) of Spo1p lipase activity in meiosis. Based on the similarity of Spo1p to phospholipase B enzymes, its unique role in SPB duplication, and pleiotropic effects on MII, late gene expression and spore formation, we propose that the Spo1 protein participates in a novel meiotic pathway that functions through the SPB to coordinate nuclear division with spore development. PMID- 10855498 TI - Meiotic recombination in RAD54 mutants of Saccharomyces cerevisiae. AB - The Rad54 protein is an important component of the recombinational DNA repair pathway in vegetative Saccharomyces cerevisiae cells. Unlike those in other members of the RAD52 group, the meiotic defect in rad54 is rather mild, reducing spore viability only to 26%-65%. A consistently greater requirement for Rad54p during meiosis was observed in hybrid strains, suggesting that Rad54p has a certain role in interhomolog interactions. Such a role is probably minor as no recombination defects were found in the surviving gametes in three genetic intervals on chromosome V. Also, the spore viability pattern in tetrads did not reflect an increase in nondisjunction at meiosis I indicative of a meiotic recombination defect. We suggest that the meiotic defect of rad54 cells lies in the failure to repair meiosis-specific double-strand breaks outside the context of the highly differentiated pathway leading to interhomolog joint molecules and meiotic crossovers that ensure accurate segregation at meiosis I. PMID- 10855499 TI - EXO1 and MSH4 differentially affect crossing-over and segregation. AB - The 5'-3' exonuclease Exo1p from Saccharomyces cerevisiae is required for wild type levels of meiotic crossing-over and normal meiotic chromosome segregation as is the meiosis-specific MutS homologue, Msh4p. Mutations in both genes reduce crossing-over by approximately two-fold, but deltamsh4 strains have significantly lower viability and a higher frequency of meiosis I non-disjunction. Epistasis analysis indicates a complex interaction between the two genes. Although crossing over was not detectably lower in the double mutant, viability was significantly worse than either single mutant. Such a result suggests that the two genes are affecting meiotic viability by distinct mechanisms. We propose that deltaexo1 affects chromosome segregation by reducing crossing-over, while deltamsh4 affects both the frequency and distribution of crossovers. Mutation in EXO1 reduces gene conversion frequencies significantly at some but not all loci, suggesting that other enzymes are also involved in DNA resection. We propose that Exo1p plays an early role in establishing some recombination intermediates by generating single stranded tails. The role of Msh4p is suggested to be in determining whether some recombination intermediates are resolved as crossover events and in generating crossover interference. The synergistic effect of deltaexo1deltamsh4 on spore viability suggests that the two genes have partially compensatory roles in a process affecting meiotic success. PMID- 10855500 TI - Characterization of fission yeast meiotic mutants based on live observation of meiotic prophase nuclear movement. AB - We characterized four meiotic mutants of the fission yeast Schizosaccharomyces pombe by live observation of nuclear movement. Nuclei were stained with either the DNA-specific fluorescent dye Hoechst 33342 or jellyfish green fluorescent protein (GFP) fused with the N-terminal portion of DNA polymerase alpha. We first followed nuclear dynamics in wild-type cells to determine the temporal sequence of meiotic events: nuclear fusion in the conjugated zygote is immediately followed by oscillatory nuclear movements that continue for 146 min; then, after coming to rest, the nucleus remains in the center of the cell for 26 min before the first meiotic division. Next we examined nuclear dynamics in four meiotic mutants: mei1 (also called mat2), mei4, dhc1, and taz1. Mei1 and mei4 both arrest during meiotic prophase; our observations, however, show that the timing of mei1 arrest is quite different from that of mei4: the mei1 mutant arrests after nuclear fusion but before starting the oscillatory nuclear movements, while the mei4 mutant arrests after the nucleus has completed the oscillatory movements but before the first meiotic division. We also show examples of the dynamic phenotypes of dhc1 and taz1, both of which complete meiosis but exhibit impaired nuclear movement and reduced frequencies of homologous recombination: the dhc1 mutant exhibits no nuclear movement after nuclear fusion, while the taz1 mutant exhibits severely impaired nuclear movement after nuclear fusion. PMID- 10855501 TI - Methods for immunoelectron microscopic and fine structural analysis of synaptonemal complexes and nodules in yeast. AB - Several gene products involved in meiotic chromosome pairing and recombination in yeast have been identified in recent years. Two nuclear structures play key roles in the meiotic processes: the synaptonemal complex (SC), which is essential for the pairing of the chromosomes, and the recombination nodules (RNs), which mark the sites of recombination. Good morphological representation of the yeast SC and RNs is needed in order to show structural changes caused by specific mutations in protein-coding genes and for fine localization of proteins using immunoelectron microscopy (immuno-EM). This paper presents a newly developed preparation method for EM and immuno-EM that allows analysis of fine structural details and localization of proteins in the SC and RNs in yeast. Structural components of the SC are clearly seen and appear strikingly similar to those in the SC in other organisms. Antibodies against the SC protein Zip1, a transverse filament protein, label the central region of the SC strongly and specifically as expected. The improved method will be an important tool in high-resolution determination of the location of proteins in the meiotic yeast nucleus. PMID- 10855502 TI - Fate of meiotic lamin C2 in rat spermatocytes cultured in the presence of okadaic acid. AB - The fate of nuclear envelope proteins during the pachytene/metaphase I transition was investigated in rat spermatocytes cultured in vitro in the presence of the phosphatase inhibitor okadaic acid (OA). Under these experimental conditions lamin B1 and the lamina-associated proteins 2 (LAPs2) behave as already described in other cell types. In contrast to these results, meiotic lamin C2 appears to be degraded after addition of OA to the spermatocyte culture medium as this lamin was no longer detectable by immunofluorescence microscopy or by immunoblotting. Taking into account the peculiarities of the lamin C2 primary structure, it is tempting to speculate that degradation of this protein represents a critical step in the process of disassembly of the spermatocyte nuclear envelope. PMID- 10855503 TI - Localisation of RAD50 and MRE11 in spermatocyte nuclei of mouse and rat. AB - Synaptonemal complexes (SCs) are zipperlike structures that are assembled between homologous chromosomes during meiotic prophase. They consist of two axial elements (AEs) (one along each of the two homologous chromosomes), which, in mature SCs, are connected by numerous transverse filaments along their length. Several proteins involved in the later steps of meiotic recombination most probably function in close association with the AEs of SCs, because the proteins involved in these steps have all been localised along AEs or SCs by immunocytochemical methods. It is not known at which step in meiotic recombination this association with the AEs is established. In order to shed some light on this issue, we analysed the localisation of two proteins that are involved in early steps of meiotic recombination, RAD50 and MRE11, relative to AEs and SCs by immunofluorescence labelling of paraffin sections of the mouse testis, using affinity-purified polyclonal antibodies against RAD50 and MRE11, and monoclonal and polyclonal antibodies against SC components. The localisation patterns of MRE11 and RAD50 within spermatocytes were very similar. MRE11 and RAD50 appeared in high abundance in preleptotene spermatocytes, just before SC components could be detected. From preleptotene until early zygotene they were present throughout the nucleus. In mid and late zygotene, MRE11 and RAD50 concentrated in distinct areas; in early pachytene the two proteins had almost disappeared from the nucleus, except from the sex vesicle (the chromatin of the XY bivalent), where they persisted in high abundance until diplotene. We propose that MRE11 and RAD50, together with other proteins, prepare chromatin throughout the early meiotic prophase nucleus for the initiation of meiotic recombination. Possibly, only a small fraction of the RAD50- and MRE11-containing (pre)recombination complexes associates transiently with AEs, where further steps in meiotic recombination can take place. PMID- 10855504 TI - Identification and characterization of an SPO11 homolog in the mouse. AB - The SPO11/TOPVIA family includes proteins from archaebacteria and eukaryotes. The protein member from the archaebacterium Sulfulobus shibatae is the catalytic subunit of TopoVI DNA topoisomerase. In Saccharomyces cerevisiae, Schizosaccharomyces pombe, Caenorhabditis elegans and Drosophila melanogaster, SPO11 is required for meiotic recombination, suggesting a conserved mechanism for the initiation step of this process. Indeed, S. cerevisiae SPO11 has been shown to be directly involved in the formation of meiotic DNA double-strand breaks that initiate meiotic recombination. Here, we report the identification of a Mus musculus Spo11 cDNA, which encodes a protein closely related to all members of the SPO11/TOPVIA family. cDNAs resulting from alternative splicing were detected, suggesting that there are potential variants of the mouse SPO11 protein. By RNA blotting analysis, expression of the mouse Spo11 gene was detected only in the testis, in agreement with its predicted function in the initiation of meiotic recombination. We mapped the mouse Spo11 gene to chromosome 2, band H2-H4. PMID- 10855505 TI - Distribution of Atr protein in primary spermatocytes of a mouse chromosomal mutant: a comparison of preparation techniques. AB - In this study, we examined the suitability of a three dimensional preparation technique for studying chromosome behaviour in the first meiotic prophase in the mouse chromosomal mutant T(1;13)H/T(1;13)Wa. To preserve cellular shape, primary spermatocytes were encapsulated in a fibrin clot. Conventionally sedimented prophase nuclei served as controls. Axial elements and lateral synaptonemal complex components were subsequently stained by immunofluorescence and the presence of axial elements at the pachytene stage was highlighted with indirect immunofluorescence against the Atr protein. We compared the distribution of Atr signal in the fibrin-embedded spermatocytes with surface-spread preparations and immunohistochemically stained histological sections of seminiferous tubules. Furthermore, fluorescence in situ hybridisation of the mouse minor satellite DNA was done on fibrin-embedded spermatocytes. The Atr signal is most conspicuous in fibrin-embedded nuclei on unpaired axial elements during pachytene, both for sex chromosomal and for autosomal segments, and expanding from these elements into the surrounding chromatin. Both spread and encapsulated zygotene nuclei with extended axial element formation proved to be positive for Atr. Mid- to late zygotene nuclei were devoid of 3,3'-diaminodibenzene deposition in the histological sections. Highlighting the unpaired axial elements in the small heteromorphic 1(13)H;1(13)Wa bivalent with an Atr signal enabled meiotic analysis of this bivalent to be carried out in a three-dimensional context. Thus, proximity of this bivalent with the sex chromosomes is found more often in three dimensional preparations than in spread preparations. Furthermore, the development of the Atr signal over the sex chromosomes as pachytene proceeds helps in substaging of this long and heterogeneous meiotic phase, in sedimented but especially in fibrin-encapsulated nuclei. PMID- 10855506 TI - The Su(Hw) chromatin insulator protein alters double-strand break repair frequencies in the Drosophila germ line. AB - P element-induced double-strand breaks (DSBs) on the X chromosome of Drosophila melanogaster were repaired up to four times more frequently when functional Su(Hw) chromatin insulator protein was removed from all genomic binding sites. Simultaneous comparisons of interallelic gap repair frequencies at two target loci on the X chromosome confirmed that a Su(Hw) binding site nested within a template had no effect on DSB repair efficiency. The results suggest that the genome-wide homology search of broken ends for homologous template sequences is affected because it is the only step in the recombinational repair process with an apparent genome-wide interaction. We propose that the searching 3'-hydroxy ends gain a higher degree of freedom for the search in a su(Hw) mutant background. PMID- 10855507 TI - Is grand multiparity associated with offsprings' hospital-treated mental disorders? A 28-year follow-up of the North Finland 1966 birth cohort. AB - BACKGROUND: A child born to a grand multiparous (GMP) mother (i.e. a mother who has undergone six or more deliveries) is at increased risk of perinatal complications, but it is not known whether or not GMP status is associated with child's adulthood mental disorders. METHODS: The data were obtained from the unselected, general population Northern Finland 1966 Birth Cohort (n = 11,017). The cohort members (children) were followed up prospectively to the age of 28 years. Using the National Hospital Discharge Register, a total of 89 DSM-III-R schizophrenia cases were identified, as well as 55 other psychoses, 87 personality disorders, 36 cases of alcoholism, 53 depressive disorders, and 67 anxiety and other non-psychotic disorders. The association between the mother's grand multiparity and the offspring's adult hospital-treated psychiatric morbidity was analysed using a continuation ratio model, which is a modification of logistic regression. Odds ratios were adjusted for social class, maternal antenatal depression, and wantedness of pregnancy. RESULTS: A total of 1320 mothers (12%) were GMPs. Maternal GMP status was not associated with offspring's schizophrenia, anxiety or other non-psychotic disorders. The risk of other psychoses (OR 2.3; 95% CI 1.2-4.7), alcoholism (OR 2.0; 95% CI 0.8-4.7) and depressive disorder (OR 2.2; 95% CI 1.0-4.5) was elevated among offspring of GMP mothers. CONCLUSIONS: It is possible that the mother's GMP status and the large family size associated with this are causal factors in the development of other psychoses than schizophrenia, alcoholism and depression among adult offspring. PMID- 10855508 TI - Family burden and coping strategies in schizophrenia: 1-year follow-up data from the BIOMED I study. AB - BACKGROUND: To date, only few data are available on how family burden in schizophrenia changes over time. In addition, no study has explored how such factors as coping styles and social support influence burden over time. This paper presents the 1-year follow-up data from the BIOMED I study on family burden and coping strategies in schizophrenia. METHODS: A sample of 159 relatives of patients with schizophrenia living in five European countries was followed up prospectively for 1 year with regard to burden and coping strategies, using validated questionnaires. RESULTS: In the sample as a whole, the burden was stable. A reduction of family burden over time was found among relatives who adopted less emotion-focused coping strategies and received more practical support from their social network. In addition, family burden decreased in relation to the improvement of patient's social functioning. CONCLUSIONS: When relatives of patients with schizophrenia are able to improve their coping strategies, it is possible for burden to be reduced even after several years. This points to the necessity to provide families of chronic psychotic patients with psychoeducational interventions emphasising the adoption of an effective coping style. PMID- 10855509 TI - Household crowding and compliance with outpatient treatment in patients with non affective functional psychoses in Sao Paulo, Brazil. AB - BACKGROUND: Compliance with outpatient treatment can reduce the use of psychiatric inpatient services by people with severe mental disorders. In developing countries, socio-economic factors may be associated with compliance with outpatient treatment. METHODS: A 2-year prospective cohort study was conducted of 99 patients with non-affective functional psychoses who were discharged from hospital. Standardized assessments were used for psychopathology, social functioning and use of psychiatric services. RESULTS: Forty-two patients (42.4%) missed all outpatient appointments for at least 2 consecutive months. Household crowding was the only variable associated with poor compliance, patients living in very crowded homes being more than twice as likely to show poor compliance as those living in less crowded homes. CONCLUSIONS: In large urban centres in developing countries, strategies to improve compliance with outpatient treatment targeted towards those living in overcrowded households may reduce use of psychiatric beds. PMID- 10855510 TI - The reliability of case register diagnoses: a birth cohort analysis. AB - BACKGROUND: Few studies assess the reliability of case register diagnoses, despite their widespread use in psychiatric research. This study investigates case register diagnostic reliability in comparison to casenote derived diagnoses in a birth cohort. METHODS: Diagnostic information from the case register and casenotes of 449 individuals was extracted. The incident and lifetime register diagnoses were compared with those derived from the casenotes. RESULTS: Inter rater reliability was good (kappa = 0.71). Agreement between casenote and incident register diagnosis was moderate (kappa = 0.52), as was agreement between casenote and register lifetime diagnosis (kappa = 0.58). Case register diagnoses were insufficiently accurate to stand alone. Case register diagnoses for organic disorder, schizophrenia, alcoholism, learning disability, personality disorder and transient or no psychiatric disorder were reliable enough for the case register to act as a useful screening instrument. The case register was not acceptable, even as a screening instrument, for the diagnoses of neurotic or affective disorders. CONCLUSIONS: Studies relying only on case register diagnoses may be flawed if diagnoses are not independently verified. National statistics derived from case register data, especially for neurosis and affective disorder, may be unreliable. PMID- 10855511 TI - Suicide amongst Cambridge University students 1970-1996. AB - BACKGROUND: Anecdote, media coverage and earlier research suggest that the rate of suicide amongst students at Cambridge and Oxford Universities is unduly high. There is also a popular belief that student suicide is common at examination times. METHOD: Student deaths at the University of Cambridge were identified using the University database. The cause of death was determined by reference to death certificates and coroners' inquest records. RESULTS: We identified 157 student deaths during academic years 1970-1996, of which 36 appeared to be suicides. The overall suicide rate was 11.3/100,000 person years at risk. Suicide rates were similar to those seen amongst 15- to 24-year-olds in the general population. There were non-significant trends for male postgraduates to be over represented and first-year undergraduates under-represented. Examination times were not associated with excess suicide. CONCLUSIONS: Suicide rates in University of Cambridge students do not appear to be unduly high. PMID- 10855512 TI - Psychosocial predictors of first-onset depression in Chinese Americans. AB - BACKGROUND: This study examines the longitudinal and concurrent risk factors associated with first-onset major depression in a community sample of 1747 Chinese Americans in Los Angeles. METHODS: The relative contributions of demographic, health, psychiatric, psychosocial, and cultural variables were assessed in a series of longitudinal and concurrent hierarchical multivariable analyses. RESULTS: Results of the longitudinal analyses indicated that the risk for experiencing a first major depressive episode at 18-months follow-up was higher for those who initially rated their health as poor, reported higher depressive symptoms, and perceived higher levels of social support. After controlling for prior health and psychiatric and psychosocial status at time 1, the results of the concurrent analyses indicated that the risk for experiencing a first major depressive episode at time 2 was higher for those who rated their health as poor, had at least one other psychiatric disorder, were bilingual, experienced high levels of life stress, and perceived themselves as having low and/or decreased social supports. CONCLUSIONS: The results of this study confirm previous evidence that psychosocial vulnerabilities, including higher acculturation, greater stress exposure and reduced social supports, are important predictors of risk for first-onset depressive episodes. Prevention and treatment implications are addressed, and future directions for research are offered. PMID- 10855513 TI - Prolonged survival and childhood-onset epilepsy in alobar holoprosencephaly. PMID- 10855514 TI - Posterior fossa tension pneumocephalus. AB - Posterior fossa tension pneumocephalus (PFTP) is a very rare clinical entity--OFF few case reports available prove how rare. Five patients with PFTP are presented. All were operated on for posterior fossa lesions. One was operated on in the lateral position and the rest in a sitting position. All the patients had postoperative (early) neurological deterioration, and computed tomography (CT) scans revealed PFTP in the left cerebellopontine angle in one case and in the IV ventricle in the rest. Air was aspirated from the posterior fossa in one case. All the cases were electively ventilated for 48-72 h. All the patients had an immediate onset of progressive neurological recovery after early intervention and elective ventilation. One patient died 10 days later because of a recurrence of empyema and the development of meningitis. PFTP is a rare but important complication of posterior fossa surgery. Early intervention and elective ventilation can produce a good recovery. PMID- 10855515 TI - Syringomyelia in children with primary scoliosis. AB - The clinical notes of 35 children presenting with scoliosis were reviewed; all 35 had been investigated with MRI. Seven were found to have syringomyelia, and six of these had Chiari malformation. Correction of the syrinx resulted in improvement or stabilisation of the spinal curvature. We recommend that all cases presenting with primary scoliosis should have MRI and should be treated if a syrinx is found. PMID- 10855516 TI - Serial lumbar CSF presure measurements and cranial computed tomographic findings in childhood tuberculous meningitits. AB - Intracranial pressure (ICP) was monitored in 218 consecutive children with hydrocephalus secondary to tuberculous meningitis (TBM). All children underwent cranial computerized tomographic (CT) scanning and continuous lumbar cerebrospinal (CSF) pressure monitoring on admission. Noncommunicating hydrocephalus (37 children), as determined by air encephalography, was treated by ventriculoperitoneal (VP) shunting and communicating hydrocephalus (181 children), by means of daily acetazolamide and frusemide. Response of ICP to treatment in the group with communicating hydrocephalus was assessed by means of repeated CSF pressure monitoring and CT scanning. One hundred and eighty-five of the 218 patients survived the 1st month of treatment. The aim of this study was the retrospective determination of (1) the relationship between ICP measurements and CT findings on admission and (2) the characteristics of the ICP recording which correlated best with the CT criteria of compensated hydrocephalus after the 1st month of treatment. No relationship was found between the level of baseline CSF pressure and the degree of hydrocephalus, as demonstrated by CT scanning, on admission. Seventy-five per cent of the patients with communicating hydrocephalus that survived the 1st month of treatment complied with the CT criteria for compensated hydrocephalus. All these patients had a baseline CSF pressure below 15 mmHg and absence of high-amplitude B waves on the pressure recording done at the end of the 1st month. In this study repeated lumbar CSF pressure monitoring proved to be an effective instrument to assess the response of communicating tuberculous hydrocephalus to medical treatment and also accurately predicted the timing of compensation of the hydrocephalus. PMID- 10855517 TI - Zero rate of shunt infection in the first postoperative year in children--dream or reality? AB - The rate of infectious complications following shunt implantations at the Pierre Wertheimer Hospital was 6.4% in 1992-1994. In order to improve this infection rate, new recommendations for surgery were applied and a new type of valve was used. The effects of these measures after a 1-year follow-up were analyzed in 70 patients. The rate of infection was zero, 2.8, and 4.3% at 2, 6, and 12 months, respectively. A case-control study did not reveal any significant risk factor among the patient and surgical factors analyzed. PMID- 10855518 TI - Membranous occlusion of the foramen of Monro following ventriculoperitoneal shunt insertion: a role for endoscopic foraminoplasty. AB - We report two cases of lateral ventricle dilatation due to membranous occlusion of the foramen of Monro following ventriculoperitoneal shunt insertion. Both cases were treated successfully by endoscopic foraminoplasty of the obstructed foramen of Monro and III ventriculostomy. One child had meningomyelocele and hydrocephalus. She had CSF infection after repair of the back lesion. Isolated left lateral ventricle occurred after insertion of a right ventriculoperitoneal shunt for hydrocephalus when the girl was 2 months old. A right ventriculoperitoneal shunt was then inserted. Chronic shunt infection with abdominal pseudocyst was found 8 years later. The shunts were exteriorized. Membranous obstruction of the foramen of Monro was found endoscopically. Fenestration of the membranous obstruction along with a III ventriculostomy was performed. After the endoscopic procedure, the exteriorized ventriculoperitoneal shunt was removed 2 weeks later. The patient was still symptom free without shunting 14 months after the operation. The other child had hydrocephalus after a premature birth and hemorrhage. Repeated ventriculoperitoneal shunt infections contributed to membranous obstruction of bilateral foramen of Monro. After the shunt infection was treated this patient's shunting procedure was simplified by endoscopic foraminoplasty of the left and right foramen of Monro along with a III ventriculostomy. He was symptom free with a new ventriculoperitoneal shunt 9 months after the operation. PMID- 10855519 TI - Causes and treatment of intracranial haemorrhage complicating shunting for paediatric hydrocephalus. AB - Intracranial haematomas are a well-known complication of shunting procedures for hydrocephalic patients. Most are subdural haematomas, and epidural haematomas are much less common in this setting. Their aetiology is thought to be due to an overdrainage of cerebrospinal fluid and a rapid lowering of intracranial pressure, leading to the development of these haematomas. Since the advent of modern neuroimaging techniques, prompt diagnosis of postshunting intracranial haematoma has been possible even in asymptomatic patients. The choice between surgical and nonsurgical management of postshunting intracranial haematoma is a difficult and controversial issue, especially in asymptomatic patients. Several therapeutic options have been proposed for the treatment of postshunting intracranial haematoma. Evacuation of the haematoma by conventional neurosurgical methods with the implantation of a higher pressure valve system is the most common option adopted. Intraventricular haemorrhage is occasionally reported, chiefly in children with hydrocephalus associated with vein of Galen malformation. PMID- 10855520 TI - Issues of medical management in adults with spina bifida. AB - The objective of this study was to establish the range of medical issues among those attending a clinic for adults with spina bifida (SB) and hydrocephalus (CASBAH). Owing to improvements in medical care in the past 3-4 decades, an increasing number of SB patients go on to prosper in adult life. Since 1990 there has been a CASBAH service in Belfast serving Northern Ireland on a regional basis. There are now 237 patients with SB remaining on the live register at CASBAH. All records were reviewed with regard to site of lesion, ambulatory ability, shunt placement and regularity of attendance. The records of 193 were also reviewed with reference to musculoskeletal problems, bladder function, bowel function, renal impairment and hypertension, skin breakdown, epilepsy and the incidence of clinically significant Chiari/hydrosyringomyelia. Of the patients on the register, 36% are wheelchair dependent, 8% have some ambulatory capacity but are largely wheelchair dependent, 22% are ambulatory with aid and 34% are independently ambulatory. Bladder function is normal in 8%, whilst 32% describe normal bowel function. Renal impairment is present in 48% of patients, and 15% are on anti-hypertensive therapy. Epilepsy is an active issue in 9%, and intraventricular shunts are in situ in 37% of patients. Scoliosis is present in 50% and 70% have joint deformities or contractures. Five patients have become symptomatic from the Chiari/hydrosyringomyelia complex. These data reflect the considerable range of disability in adult SB patients, the challenges presented in long-term management and the need for organised follow-up. PMID- 10855521 TI - Rhabdoid tumors of the central nervous system. AB - Rhabdoid tumors of the central nervous system are rare malignancies with a still almost uniformly fatal outcome. There is still no proven curative therapy available. We report our experience with nine patients with central nervous system rhabdoid tumors. Gross complete surgical removal of the tumor was achieved in six patients. Seven patients received intensive chemotherapy. Four of these were treated in addition with both neuroaxis radiotherapy and a local boost directed to the tumor region, while two patients received local radiotherapy only. The therapy was reasonably well tolerated in most cases. Despite the aggressive therapy, eight of the nine patients died from progressive tumor disease, and one patient died from hemorrhagic brain stem lesions of unknown etiology. The mean survival time was 10 months after diagnosis. Conventional treatment, although aggressive, cannot change the fatal prognosis of central nervous system rhabdoid tumors. As these neoplasms are so rare, a coordinated register would probably be a good idea, offering a means of learning more about the tumor's biology and possible strategies of treatment. PMID- 10855522 TI - Magnetic resonance spectroscopy of brain in epilepsy. AB - The current applications of magnetic resonance spectroscopy (MRS) in the clinical management of epileptic patients are reviewed. A major contribution of MRS to epilepsy is its ability to determine lateralisation before surgical resection of the diseased brain region. Phosphorus-31 and proton single-voxel MRS identify abnormalities in high-energy metabolism, neuronal function and neurotransmitter levels, but information can only be obtained from restricted regions of the brain. Spectroscopic imaging techniques (also known as chemical shift imaging) provide a metabolic mapping of the whole brain. They expand the range of applications of MRS to other types of epilepsy (neocortical, frontal) than temporal lobe epilepsy, which is the most often studied. Also, spectral editing techniques in proton MRS make it possible to detect and monitor drug-induced variations of GABA in the human brain, opening new insights into patient response to drug therapy of epilepsy. MRS is playing an increasing role in the noninvasive characterisation and management of epileptic patients. PMID- 10855523 TI - The treatment of infantile hydrocephalus: "differential-pressure" or "flow control" valves. A pilot study. AB - The choice of shunt valve in the treatment of hydrocephalus in children remains controversial. We embarked on a pilot study to determine the differences in outcome between differential-pressure and flow-regulating valves. Prospective data collected on 50 consecutive first-time shunt insertions, performed between June 1993 to June 1996, was analysed. Children with tumour-related hydrocephalus and Dandy-Walker malformations as well as children who had external ventricular drainage prior to definitive shunt insertion were excluded from the study. The defining event was the first complication necessitating surgery, including obstruction, over-drainage and infection. Of the 50 children (31 males), 23 had differential pressure (medium-pressure) and 27 had Delta (performance level 2) valves inserted. The mean age at shunt insertion was 26.4 months. The mean follow up was 53.8 months. The overall cumulative shunt survival at 5 years was 58.6% for the differential pressure and 58.7% for the Delta valves. The mean shunt life was 37.1 months for the differential pressure group and 34.6 months for the Delta group. This difference was not statistically significant (P=0.72, t-test). Both valves had a similar outcome with respect to obstruction (including proximal, valve, distal). The main differences between the two valves were with respect to the incidence of over-drainage and infection. Amongst the differential pressure valves, there were 4 instances of overdrainage (3 slit-ventricle syndrome, 1 bilateral subdural collection)--all occurring within the first 36 months. The Delta valve group had only one instance of over-drainage (bilateral subdural collection). There were no infections in the differential pressure valve group, whereas 3 of the Delta valve shunts got infected, all within the first month. Whereas both shunt types seemed to have a similar overall survival, there was a relatively higher incidence of over-drainage amongst the differential pressure valves. The Delta valves, on the other hand, had higher rates of infection. Similar studies with larger numbers could suggest whether the choice of shunt type will ultimately have to be a compromise accepting one or the other complication. PMID- 10855524 TI - Repair of a congenital cranial defect in a newborn with autologous calvarial bone. AB - A case is reported of early repair of a cranial defect associated with aplasia cutis congenita using a full-thickness autologous calvarial graft. New bone regenerated to fill the defect at the donor site, such that by 6 weeks of life no calvarial defects remained. This simple technique is recommended as an alternative to a delayed acrylic or split cranial bone cranioplasty as a second operation. A similar technique could be considered for other congenital cranial defects, such as those associated with encephaloceles or bilateral parietal foramina. PMID- 10855525 TI - Moyamoya disease associated with a brain stem glioma. AB - An 8-year-old boy was found to have primary moyamoya disease associated with a brain stem glioma. For over 3 years the child had experienced transient ischemic attacks induced by hyperventilation. One month before referral to our hospital he had presented with progressive left facial nerve palsy. Magnetic resonance imaging showed a cystic mass in the lower pons. Angiography revealed severe bilateral stenosis of the internal carotid arteries and prominent moyamoya vessels in the basal ganglia. Partial resection of the tumor yielded a histological diagnosis of pilocytic astrocytoma. Local radiation therapy reduced the size of the tumor. Anastomosis of the superficial temporal arteries and middle cerebral arteries on both sides was then performed. After direct bypass surgery, the patient remained in a good condition for a 5-year follow-up period. Clinical investigation of the coincidence of primary moyamoya disease and brain stem glioma led the authors to conclude that these two diseases coexisted independently. PMID- 10855526 TI - Malignant triton tumor in the thoracic spine. AB - We present a 15-year-old patient diagnosed with peripheral neurofibromatosis (NF 1), who was admitted with paraparesis caused by a large intrathoracic tumor with an intracanalicular component that affected the spinal cord. After surgery his condition improved, but a year later he suffered a relapse and died. Histologically the tumor was diagnosed as malignant with neurogenic and myogenic differentiation ("malignant triton tumor"). Malignant triton tumors (MTT) are infrequent; those found in the head and neck and the upper or lower extremities have a better prognosis than those in the retroperitoneum, buttock, or trunk. It is not clear whether this variation is due to a difference in tumor grade, stage, or resectability, or whether it is a consequence of therapy. PMID- 10855527 TI - New insights into sympathetic regulation of glucose and fat metabolism. AB - The autonomic nervous system modulates glucose and fat metabolism through both direct neural effects and hormonal effects. This review presents recent concepts on the sympathetic regulation of glucose and fat metabolism. Focally released norepinephrine from sympathetic nerves is likely to increase glucose uptake in skeletal muscle and adipose tissues independent of insulin but norepinephrine does not contribute so much as epinephrine to hepatic glucose production. Epinephrine increases hepatic glucose production and inhibits insulin secretion and the glucose uptake by tissues that is induced by insulin. Additionally, catecholamines can increase thermogenesis and lipolysis, leading to increased energy expenditure and decreased fat stores. It is likely that beta-(beta3) adrenergic receptors mediate these responses. Alterations of central neurotransmission and environmental factors can change the relative contribution of sympathetic outflow to the pancreas, liver, adrenal medulla and adipose tissues, leading to the modulation of glucose and fat metabolism. Recent studies have proposed that leptin, an adipocyte hormone, affects the central nervous system to increase sympathetic outflow independent of feeding. The effects of leptin on glucose and fat metabolism could be in part mediated by the sympathetic nervous system. Studies using mice with a genetic disruption of serotonin 5-HT2c receptor indicate that central neural mechanisms in the regulation of sympathetic outflow and satiety could be dissociated. Abnormalities of sympathetic effects, including disturbances of leptin and beta3-adrenergic receptor signalling, are likely to cause obesity and impaired glucose tolerance in rodents and humans. These findings indicate that dysfunction of the sympathetic nervous system could predispose to obesity and Type II (non-insulin-dependent) diabetes mellitus. PMID- 10855528 TI - The importance of lipid-derived malondialdehyde in diabetes mellitus. AB - Malondialdehyde (MDA) is a highly toxic by-product formed in part by lipid oxidation derived free radicals. Many studies have shown that its concentration is increased considerably in diabetes mellitus. Malondialdehyde reacts both irreversibly and reversibly with proteins and phospholipids with profound effects. In particular, the collagen of the cardiovascular system is not only stiffened by cross-links mediated by malondialdehyde but then becomes increasingly resistant to remodelling. It is important in diabetes mellitus because the initial modification of collagen by sugar adducts forms a series of glycation products which then stimulate breakdown of the lipids to malondialdehyde and hence further cross-linking by malondialdehyde of the already modified collagen. Some progress is being made into the mechanisms of formation and the nature of the intermolecular cross-links induced by malondialdehyde which result in the stiffening of the collagenous tissues. Our recent studies indicate the formation of pyridyl cross-links. Malondialdehyde has been shown to react several orders of magnitude faster with the pre-existing collagen enzymic cross links than the amino acid side-chains. Malondialdehyde modification of basic amino-acid side-chains also results in a change in properties, for example, in the charge profile of the molecule resulting in modified cell-matrix interactions. Although aspects of the biochemistry of malondialdehyde are still not fully understood its complex chemistry is being unravelled and this should lead to ways of preventing its damaging reactions, for example, through antioxidant therapy. PMID- 10855529 TI - Increased mortality in Type II diabetic patients using sulphonylurea and metformin in combination: a population-based observational study. AB - AIMS/HYPOTHESIS: This study analysed cause-specific mortality in Type II (non insulin-dependent) diabetic patients using either sulphonylurea alone or in combination with metformin. METHODS: Patients were followed from the first day they were taking either the combination or sulphonylurea alone. Odds ratios by Cox regression analyses were adjusted for age, sex, duration of diabetes, study area, year of inclusion and fasting blood glucose at inclusion. RESULTS: We included 169 patients taking sulphonylurea and metformin in combination and 741 patients taking only sulphonylurea. Mean (range) follow-up time was 6.1 (0.1 13.0) years. The adjusted odds ratio for overall mortality was 1.63 (95% confidence interval 1.27-2.09) in patients taking sulphonylurea and metformin combination vs those using sulphonylurea alone. For mortality from ischaemic heart disease and stroke the adjusted odds ratios were 1.73 (95% confidence interval 1.17-2.55) and 2.33 (95% confidence interval 1.17-4.63), respectively. CONCLUSION/INTERPRETATION: There was a higher cardiovascular mortality in Type II diabetic patients taking sulphonylurea and metformin in combination than in those taking only sulphonylurea. Hence, it cannot be excluded that this kind of combination therapy possibly increases cardiovascular mortality. It is feasible that the increased mortality was secondary to a more aggressive type of diabetes in the patients using sulphonylurea and metformin in combination. Combination therapy is known to promote additional blood glucose reduction but there is as yet no evidence that a sulphonylurea and metformin combination is more beneficial on micro- or macrovascular disease than sulphonylurea or metformin alone. PMID- 10855530 TI - Glucose tolerance and other determinants of cardiovascular autonomic function: the Hoorn Study. AB - AIMS/HYPOTHESIS: Currently, three categories of measures are used to assess cardiovascular autonomic dysfunction: measures of the Ewing-test, measures of heart-rate variability, and measures of baroreflex sensitivity. We studied the determinants of these measures obtained from cardiovascular autonomic function tests in the Hoorn Study. METHODS: The study group (n = 631) consisted of a glucose-tolerance-stratified sample from a 50- to 75-year-old group of people. Cardiac cycle duration (RR interval) and continuous finger arterial pressure were measured under three conditions: during (a) spontaneous breathing, (b) six deep breaths over one minute, and (c) an active change in position from lying to standing. From these readings, ten measures of autonomic function were assessed (three Ewing, six heart-rate variability and one baroreflex sensitivity). As possible determinants we considered age, sex, glucose tolerance, cardiovascular disease, use of anti-hypertensive drugs, anthropometric factors, metabolic factors and lifestyle factors. RESULTS: Multivariate analysis showed that eight of ten cardiovascular autonomic function measures were most strongly associated with glucose tolerance. Furthermore, measures were moderately associated with age, sex, waist-to-hip ratio, use of anti-hypertensive drugs, and insulin. The measures were weakly associated with coronary artery disease but not with lipids. The strongest determinants seemed to differ between subjects with and without diabetes: in the non-diabetic subjects the most strongly associated were age and use of anti-hypertensive drugs and in subjects with diabetes, insulin. No consistent differences in association between the three categories of measures were observed. CONCLUSION/INTERPRETATION: The strongest determinants of autonomic function were age, presence of diabetes and use of anti-hypertensive drugs. PMID- 10855531 TI - The effect of acute hyperglycaemia on QTc duration in healthy man. AB - AIMS/HYPOTHESIS: Prolongation of heart rate-adjusted QT (QTc) is associated with an increased risk of coronary heart disease and sudden death. The objective of this study was to investigate whether acute increases of plasma glucose concentrations in healthy subjects could influence QTc and QTc dispersion. METHODS: Plasma glucose concentrations were quickly raised to 15 mmol/l in 20 healthy subjects (10 men/10 women) and maintained for 2 h. On another occasion, and in random order, all subjects underwent the same hyperglycaemic clamp as above and an infusion of the somatostatin analogue octreotide (25 microg as iv bolus followed by a 0.5 g/min infusion) to block the release of endogenous insulin. RESULTS: Systolic and diastolic blood pressures, heart rate and plasma catecholamine concentrations showed significant increases (p < 0.05) starting after 60 min of hyperglycaemia. QTc, QTc dispersion and PR interval also showed significant increments at 120 min of the hyperglycaemic clamp. The infusion of octreotide did not influence QTc duration, QTc dispersion, PR interval and the haemodynamic effects of acute hyperglycaemia. CONCLUSION/INTERPRETATION: The results show that acute hyperglycaemia produces significant increments of QTc and QTc dispersion in normal subjects. In this context, endogenously released insulin during acute hyperglycaemia seems to play a minor part. PMID- 10855532 TI - Effects of gestational diabetes on human placental glucose uptake, transfer, and utilisation. AB - AIMS/HYPOTHESIS: Gestational diabetes is associated with complications for the offspring before, during and after delivery. Poor maternal glucose control, however, is a weak predictor of these complications. Given its position at the interface of the maternal and fetal circulations, the placenta possibly plays a crucial part in protecting the fetus from adverse effects from the maternal diabetic milieu. We hypothesised that gestational diabetes may result in changes in placental function, particularly with respect to the uptake, transfer, and/or utilisation of glucose. We aimed to examine glucose transport and utilisation in intact human placental lobules from women with gestational diabetes and those from normal pregnancies. METHOD: Dual perfusion of an isolated placental lobule was done on placentae from diet treated gestational diabetic (n = 7) and normal pregnant patients (n = 9) using maternal glucose concentrations of 4, 8, 16 and 24 mmol/l in random order over a 4-h experiment. Results were expressed in micromol x min(-1) x g(-1). RESULTS: D-glucose uptake from the maternal circulation (control 0.492 vs gestational diabetes mellitus 0.248, at 8 mmol/l maternal glucose), D-glucose utilisation by the placenta (0.255 vs 0.129), D glucose transfer to the fetal circulation (direct 0.979 vs 0.402; net transfer 0.269 vs 0.118) and L-lactate maternal release into both the fetal (0.052 vs 0.042) and maternal (0.255 vs 0.129) circulation were significantly reduced during in vitro perfusion of placentae from patients with gestational diabetic pregnancies. Transfer of 3H-L-glucose also significantly reduced in the diabetic group (8.1% vs 2.6%). CONCLUSION/INTERPRETATION: These results suggest that placental transport and metabolism of D-glucose is altered during gestational diabetes. PMID- 10855533 TI - Short-term treatment with GLP-1 increases pulsatile insulin secretion in Type II diabetes with no effect on orderliness. AB - AIMS/HYPOTHESIS: The enteric incretin hormone, glucagon-like peptide-1 (GLP-1), is a potent insulin secretagogue in healthy humans and patients with Type II (non insulin-dependent) diabetes mellitus. In this study we assessed the impact of short-term GLP-1 infusion on pulsatile insulin secretion in Type II diabetic patients. METHODS: Type II diabetic patients (n = 8) were studied in a randomised cross-over design. Plasma insulin concentration time series were obtained during basal conditions and during infusion with saline or GLP-1 (1.2 pmol/l x kg(-1) x min(-1)) on 2 separate days. Plasma glucose was clamped at the initial concentration by a variable glucose infusion. Serum insulin concentration time series were evaluated by deconvolution analysis, autocorrelation analysis, spectral analysis and approximate entropy. RESULTS: Serum insulin concentrations increased by approximately 100% during GLP-1 infusion. Pulsatile insulin secretion was increased as measured by secretory burst mass (19.3 +/- 3.8 vs 53.0 +/- 10.7 pmol/l/ pulse, p = 0.02) and secretory burst amplitude (7.7 +/- 1.5 vs 21.1 +/- 4.3 pmol/l/min, p = 0.02). A similar increase in basal insulin secretion was observed (3.6 +/- 0.9 vs 10.2 +/- 2.2 pmol/l/min, p = 0.004) with no changes in the fraction of insulin delivered in pulses (0.50 +/- 0.06 vs 0.49 +/- 0.02, p = 0.84). Regularity of secretion was unchanged as measured by spectral analysis (normalised spectral power: 5.9 +/- 0.6 vs 6.3 +/- 0.8, p = 0.86), autocorrelation analysis (autocorrelation coefficient: 0.19 +/- 0.04 vs 0.18 +/- 0.05, p = 0.66) and the approximate entropy statistic (1.48 +/- 0.02 vs 1.51 +/- 0.02, p = 0.86). CONCLUSION/INTERPRETATION: Short-term stimulation with GLP-1 jointly increases pulsatile and basal insulin secretion, maintaining but not improving system regularity in Type II diabetic patients. PMID- 10855534 TI - Impaired glycogen synthesis in hepatocytes from Zucker fatty fa/fa rats: the role of increased phosphorylase activity. AB - AIMS/HYPOTHESIS: The Zucker fatty fa/fa rat develops hyperinsulinaemia, insulin resistance and severe obesity as a result of a homozygous mutation in the leptin receptor gene. The aim was to characterise the metabolic defect(s) in hepatocytes from fa/fa rats. METHODS: Glucose metabolism and key regulatory enzymes were investigated in hepatocytes from fa/fa and Fa/? rats after short-term culture in the absence of insulin. RESULTS: Hepatocytes from fa/fa rats have higher glucokinase activity and expression of the glucokinase regulatory protein and higher rates of glycolysis and lipogenesis, but lower rates of glycogen synthesis than hepatocytes from Fa/? controls. Insulin caused a similar stimulation of glycogen synthesis in hepatocytes from fa/fa rats as in controls ( > twofold) but did not restore the impaired glycogen synthesis in cells from fa/fa rats. Adenovirus-mediated glucokinase overexpression stimulated glycogen synthesis and glycolysis but aggravated rather than abolished the relative impairment of glycogen synthesis in cells from fa/fa rats. Inhibition of glycolysis with 2,5 anhydromannitol, an inhibitor of glycolysis and gluconeogenesis, increased glucose 6-phosphate concentrations and glycogen synthesis in hepatocytes from Fa/? and fa/fa rats but did not restore the impaired glycogen synthesis in cells from fa/fa rats. Hepatocytes from fa/fa rats had a higher activity of phosphorylase a in the basal state and after incubation with insulin or glucagon and higher total phosphorylase. CONCLUSION/INTERPRETATION: The increased activity of phosphorylase is a major contributing factor to the impaired glycogen synthesis in hepatocytes from fa/fa rats and could contribute to the lipogenic state by a glycogenolytic-glycolytic-lipogenic pathway. PMID- 10855535 TI - Peptide mapping and characterisation of glycation patterns of the glima 38 antigen recognised by autoantibodies in Type I diabetic patients. AB - AIMS/HYPOTHESIS: Glima 38 is an N-glycated neuroendocrine membrane protein of M(r) 38,000, which is recognised by autoantibodies in approximately 20% of patients with Type I (insulin-dependent) diabetes mellitus. The aim of this study was to characterise the carbohydrate moiety and generate peptide maps of glima 38. METHODS: Sera of high immunoreactivity to glima 38 were used to isolate 35-S methionine-labelled protein from betaTC-3 cells and a neuronal cell line GT1.7. Tunicamycin was used to inhibit N-glycation of glima 38 and define the core protein. The carbohydrate moiety was characterised for tunicamycin sensitivity, lectin binding and susceptibility to different endoglycosidases. The protein moiety was subjected to digestion by proteases to define peptide maps. RESULTS: The autoreactive epitopes in glima 38 recognised by Type I diabetic sera are conformational and independent of the carbohydrate moiety. Inhibition of N glycation of glima 38 in vivo, shows a protein core of M(r) 22,000 in both pancreatic beta-(betaTC3) and neuronal (GT1.7) cell lines. The carbohydrate moieties in the two cell types are distinct but contain a similar amount of terminal sialic acid residues and at least five oligosaccharide chains Glima 38 binds Triticum vulgare and Ricinus communis I lectins. Endoproteinase treatment of the M(r) 22,000 core protein results in peptides of M(r) 4500 and M(r) 20,000 with Lys-C, and peptides of M(r) 4000 and M(r) 11,000-12,000 with Glu-C/V8 and Asp-N proteases. CONCLUSION/INTERPRETATION: The biochemical properties of glima 38 define it as a new autoantigen in Type I diabetes and provide a basis for its purification. PMID- 10855536 TI - Modelling of the MHC II allele I-A(g7) of NOD mouse: pH-dependent changes in specificity at pockets 9 and 6 explain several of the unique properties of this molecule. AB - AIMS/HYPOTHESIS: We modelled the three-dimensional structure of I-A(g7), the chief genetic component of diabetes in non-obese diabetic mice, to understand the unusual properties of this molecule. METHODS: Modelling was done, in complex with established antigenic peptides, based on the structure of I-A(k). RESULTS: The selectivity of the I-A(g7) molecule changes greatly at pockets 9 and 6 but hardly at all at pockets 1, 4 and 7, between endosomal pH (5.0) and extracellular pH (7.0), in agreement with previous results. This selectivity is attributed to the unique combination of beta9His, beta56His and beta57Ser. The positive charges in and around pocket 9 at pH 5, favour binding by negatively charged residues. At pH 7 however, the uncharged alpha68, beta9 and beta56 histidines favour the accommodation of the bulky residues lysine, arginine, phenylalanine and tyrosine at pocket 9. The combination of beta9His and alpha66Glu is responsible for the pH dependent selectivity at pocket 6. Furthermore, the lack of repulsion between beta56His and alpha76Arg at pH 7 leads to a more stable ternary complex. CONCLUSION/INTERPRETATION: These results reconcile previous conflicts over the peptide binding ability of I-A(g7) and its motif. They furthermore provide possible explanations for the short lifetime of cell-surface I-A(g7) complexes in vivo, the higher threshold of thymic negative selection and inherent self reactivity shown by immunocytes in these mice and the protection from diabetes afforded to them by several transgenically expressed mouse class II alleles. This contributes to an understanding of the pathogenesis of Type I (insulin-dependent) diabetes mellitus in this animal. PMID- 10855537 TI - Contribution of adenoviral-mediated superoxide dismutase gene transfer to the reduction in nitric oxide-induced cytotoxicity on human islets and INS-1 insulin secreting cells. AB - AIMS/HYPOTHESIS: Vulnerability of pancreatic islets to oxygen free radicals and nitric oxide contributes to islet transplantation obstacles. This susceptibility can be linked to the low expression levels of antioxidant enzymes in islets. Our aim was to investigate the effect of overexpressing Cu/Zn superoxide dismutase in human islets through a simple procedure on the cytotoxic effects of two nitric oxide donors: 3-morpholinosydnonimine (SIN-1) and S-Nitroso-N-acetyl-D,L penicillamine (SNAP). METHODS: Cultured human islets and INS-1 rat-derived insulin-secreting cells were transfected by an E1-deleted adenovirus carrying Cu/Zn SOD cDNA under the control of a cytomegalovirus (CMV) promoter (AdSOD). The viability of the cells was tested by the WST-1 assay (Roche, Indianapolis, Ind., USA). RESULTS: The AdSOD procedure allowed SOD activity to increase by twofold to threefold for 2 to 8 days following transfection. Adenovirus-driven SOD overexpression was associated with a significant reduction of SIN-1-induced cytotoxicity on human islets (69.9 +/- 10.5% protection at 200 micromol/l and 40.5 +/- 8.9% protection at 400 micromol/l) and INS-1 cells (82.2 8.8% protection at 200 micromol/l and 31.1 +/- 5.8% protection at 400 micromol/l). Protection against increasing doses of SNAP was AdSOD dose-dependent. Transfected islets released significantly more insulin than control islets in glucose-theophylline stimulated conditions, without or following exposure to SNAP. CONCLUSIONS/INTERPRETATION: We thus established that adenoviral-induced overexpression of Cu/Zn SOD can be beneficial to human islet endocrine function and resistance to nitric oxide cytotoxicity. These data could be relevant for the development of new strategies aimed at preventing NO-induced beta-cell damage in an islet transplantation setting. PMID- 10855538 TI - Coronary atherosclerosis in Type II diabetes: angiographic findings and clinical outcome. AB - AIMS/HYPOTHESIS: Prevalence and incidence of coronary heart disease (CHD) are increased in patients with Type II (non-insulin-dependent) diabetes mellitus; whether this is entirely due to more extensive coronary atherosclerosis is, however, controversial. METHODS: We analysed the clinical, angiographic and follow-up data of 2253 consecutive patients undergoing coronary angiography over the decade 1983-1992. RESULTS: Abnormal coronary arteries (> or =50% stenosis) were found more frequently in diabetic than in non-diabetic subjects (85 vs 67%, p < 0.0001), the excess being explained by a higher prevalence of three-vessel disease (36 vs 17%, p < 0.0001). The sum of all angiographically detectable lumen stenoses (atherosclerosis score, ATS) was higher in diabetic than in non-diabetic subjects (352 +/- 232 vs 211 +/- 201 units, p < 0.0001). After adjusting for measured cardiovascular risk factors, diabetes was still associated with an excess ATS (114 units in men and 187 units in women, p < 0.0001 for both, p < 0.03 for the interaction ATS x sex). Within the diabetic group, the only variable that was independently (of sex and age) associated with ATS was serum cholesterol, whereas plasma glucose concentration, disease duration and type of treatment were not correlated with the severity of coronary atherosclerosis. In contrast, clinical grade proteinuria was not associated with a more diffuse coronary atherosclerosis either in diabetic (366 +/- 243 vs 354 +/- 233 units) or non-diabetic subjects (231 +/- 201 vs 207 +/- 197 units). Over a mean follow-up period of 88 months, 19% of diabetic patients compared with 10% of non-diabetic patients died of a cardiac cause (age and sex-adjusted odds ratio OR = 1.34 [1.14 1.57]). In a Cox model adjusting for age, sex and all major risk factors, diabetes was still associated with a significant excess risk of dying of a cardiac cause (OR = 1.37 [1.14-1.60]); this excess was similar to, and independent of, that carried by the presence of prior myocardial infarction in the whole population (OR = 1.42 [1.25-1.62]). Proteinuria was associated with a higher risk of cardiac death, particularly in diabetic patients, independently of coronary atherosclerosis (adjusted OR = 1.46 [1.03-1.99]). CONCLUSION/INTERPRETATION: In patients undergoing angiography, diabetes, especially in women, is associated with more severe and diffuse coronary atherosclerosis which is not explained by either the traditional risk factors or the presence of proteinuria. On follow-up, these patients experience an excess of cardiac deaths, to which coronary atherosclerosis and proteinuria make independent, quantitative contributions. PMID- 10855539 TI - High glucose concentration inhibits the expression of membrane type metalloproteinase by mesangial cells: possible role in mesangium accumulation. AB - AIMS/HYPOTHESIS: High glucose concentration decreases the degradation of mesangium matrix, an action substantially mediated by a reduction in the activities of the matrix metalloproteinases (MMPs). Metalloproteinase-2 is unique in that it is activated on the cell surface by one of the membrane type metalloproteinases (MT1-MMP), a process involving complex interactions with tissue inhibitor of metalloproteinase-2. The aim of this study was investigate the effects of glucose concentration on mesangial cell gene expression of MT1-MMP and its ability to modulate the activation of metalloproteinase-2. METHODS: Gene expression was determined using competitive RT-PCR, protein expression of MMP-2 was measured by western blot and its activation by zymography. Concanavalin A, known to increase MT1-MMP expression was added in some experiments. RESULTS: High glucose concentration decreased MT1-MMP gene expression (11.52 +/- 1.63 and 4.84 +/- 0.72 amol/microg RNA, 5 vs 25 mmol/l glucose, respectively) and decreased activation of MMP-2 by 30% despite a twofold increase in gene expression of MMP 2. Concanavalin A increased expression of MT1-MMP and activation of MMP-2. Irrespective of whether MMP-2 was from endogenous or exogenous source there was an excellent correlation between the MT1-MMP expression and degree of MMP-2 activation, whereas the gene expression of TIMP-2 was not significantly altered by high glucose concentration or concanavalin A. CONCLUSION/INTERPRETATION: Our results indicate that in a high glucose milieu, suppression of MT1-MMP expression could explain the low MMP-2 activity in the presence of high MMP-2 expression. This process could contribute to the mesangium matrix accumulation in diabetic nephropathy. PMID- 10855540 TI - Neuronal and endothelial nitric oxide synthase expression in outer medulla of streptozotocin-induced diabetic rat kidney. AB - AIMS/HYPOTHESIS: Several investigations have shown that the renal medulla has a greater capacity to generate nitric oxide than the renal cortex. To further evaluate the changes of nitric oxide synthesis in the kidney, particularly in the outer medulla, in disorders involving fluid and electrolyte imbalances, we sought to determine renal nitric oxide synthase expression in the diabetic rats. METHODS: We determined renal nitric oxide synthase mRNA and urinary nitrite/nitrate excretion in 12 normal and 12 streptozotocin-induced diabetic rats by reverse transcription-polymerase chain reaction with Southern blot hybridization and with Griess reaction, respectively. Nitric oxide synthase immunoreactivity was detected by immunohistochemistry in four normal and four diabetic rats. RESULTS: Neuronal and endothelial nitric oxide synthase mRNA were 3.5-fold and 1.8-fold increased in the outer medulla of 12 diabetic rats with no difference found in the cortex and inner medulla when compared with 12 normal rats. Urinary nitrite/nitrate excretion was significantly increased from the first week after diabetic induction. In normal rats, immunohistochemical studies showed positive neuronal and endothelial nitric oxide synthase immunostaining in almost all segments of renal tubules. Diabetic rats had the greatest enhancement of immunostaining for neuronal and endothelial nitric oxide synthase in the proximal straight tubule and medullary thick ascending limb. CONCLUSION/INTERPRETATION: Our results indicate that increases in neuronal and endothelial nitric oxide synthase synthesis in the kidney, particularly in the outer medulla, possibly play an important part in the adaptation of renal function to hyperglycaemia and hyperosmolality in diabetes. PMID- 10855541 TI - The cross-link breaker, N-phenacylthiazolium bromide prevents vascular advanced glycation end-product accumulation. AB - AIMS/HYPOTHESIS: Advanced glycation is postulated to have a pivotal role in mediating diabetic vascular complications. The emergence of thiazolium compounds such as N-phenacylthiazolium bromide which cleave preformed advanced glycation end products (AGEs) has allowed us to explore the effects of these agents on the vascular AGE accumulation and hypertrophy associated with diabetes. METHODS: Control and streptozotocin diabetic rats were selected at random for no treatment or treatment with N-phenacylthiazolium bromide (10 mg/kg intraperitoneally) and followed for 3 weeks. In a separate study, intervention with N-phenacylthiazolium bromide was delayed until after 3 weeks of diabetes and then given for 3 weeks (total of 6 weeks). RESULTS: Diabetes was associated with increased mesenteric vascular advanced glycation end products, as assessed by radioimmunoassay and immunohistochemistry. This increase in vascular AGE accumulation was prevented by N-phenacylthiazolium bromide treatment. Diabetes-associated mesenteric vascular hypertrophy was attenuated by treatment with N-phenacylthiazolium bromide only if given from the time of induction of diabetes. CONCLUSION/INTERPRETATION: Cross link breakers seem to be effective in preventing or reversing accumulation of advanced glycation end-products in blood vessels and have the potential to play a part in the treatment of diabetic vascular complications. PMID- 10855542 TI - Variables of the insulin resistance syndrome are associated with reduced arterial distensibility in healthy non-diabetic middle-aged women. AB - AIMS/HYPOTHESIS: The insulin resistance syndrome is related to arterial stiffness in diabetic subjects. Whether the insulin resistance syndrome is also related to arterial stiffness in non-diabetic subjects is less clear. We studied the association between variables of the insulin resistance syndrome in relation to arterial distensibility in healthy middle-aged non-diabetic women. METHODS: This study was done in 180 non-diabetic women, aged 43-55, selected from the general population. Arterial distensibility was assessed in the carotid artery. The associations were evaluated using linear regression analyses. RESULTS: Strong associations were found between arterial distensibility and the variables of the insulin resistance syndrome: body mass index, waist-to-hip ratio, high-density lipoprotein-cholesterol, triglycerides, glucose, insulin, apolipoprotein A1, plasminogen activator inhibitor-1-antigen and tissue-type plasminogen activator antigen. After additional adjustment for mean arterial pressure, common carotid arterial distensibility remained associated with body mass index: beta coefficient (95% confidence interval) per kg/m2: -0.24 (-0.42; -0.06); waist-to hip ratio: -26.62 (-40.59; -12.65) per m/m; triglycerides: -1.42(-2.77; -0.08) per mmol/l; plasminogen activator inhibitor-1-antigen: -0.01 (-0.02; -0.00) per ng/ml and borderline significant associated with high-density-lipoprotein cholesterol: 1.93 (-0.01; 3.87; p = 0.07) per mmol/l. Clustering of variables of the insulin resistance syndrome was strongly related to decreased arterial distensibility which remained after adjustment for mean arterial pressure. No association was found between arterial distensibility and variables that are not part of the insulin resistance syndrome: total cholesterol, LDL-cholesterol and apolipoprotein B. CONCLUSION/INTERPRETATION: The results of this study show that variables of the insulin resistance syndrome are associated with decreased arterial distensibility of the common carotid artery in healthy non-diabetic subjects. PMID- 10855543 TI - Variation in the PPARalpha gene is associated with altered function in vitro and plasma lipid concentrations in Type II diabetic subjects. AB - AIMS/HYPOTHESIS: Peroxisome proliferator activated receptor alpha (PPARalpha) regulates genes involved in lipid metabolism, haemostasis and inflammation, in response to fatty acids and fibrates, making it a candidate gene for risk of dyslipidaemia, atherosclerosis and coronary artery disease. Plasma non-esterified fatty acids are increased in subjects with Type II (non-insulin-dependent) diabetes mellitus, suggesting that PPARalpha could link Type II diabetes and dyslipidaemia, and affect response to fibrates. This has been investigated in association studies in healthy and diabetic subjects and in vitro studies. METHODS: The human PPARalpha gene was isolated and screened for variation by single strand conformation polymorphism analysis. Genotypes were determined for 129 Type II diabetic subjects and 2508 healthy men. The association with plasma lipid concentrations was examined. The function of the V162 variant was examined in co-transfection assays. RESULTS: We identified two polymorphisms, one in intron 3 and a missense mutation, leucine 162 to valine, in the DNA binding domain. In Type II diabetic patients, V162 allele carriers had higher total cholesterol, HDL cholesterol and apoAI whereas intron 3 rare allele carriers had higher apoAI concentrations. By contrast, no effect was observed in healthy rare allele carriers. In vitro, the V162 variant showed greater transactivation of a reporter gene construct. CONCLUSION/INTERPRETATION: Naturally occurring variation alters PPARalpha function, influencing plasma lipid concentrations in Type II diabetic patients but not healthy people. This demonstrates that PPARalpha is a link between diabetes and dyslipidaemia, and so could influence the risk of coronary artery disease, the greatest cause of morbidity and mortality in Type II diabetes. PMID- 10855544 TI - The exon 16-3t variant of the sulphonylurea receptor gene is not a risk factor for Type II diabetes mellitus in the Dutch Breda cohort. PMID- 10855545 TI - Rising incidence of childhood diabetes is seen at all ages and in urban and rural settings in Yorkshire, United Kingdom. PMID- 10855546 TI - Vaccinations as risk factors for Type I diabetes mellitus. PMID- 10855547 TI - Gut-liver axis. AB - The gut and the liver are the key organs in nutrient absorption and metabolism. Bile acids, drugs, and toxins undergo extensive enterohepatic circulation. Bile acids play a major role in several hepatic and intestinal diseases. Endotoxins deriving from intestinal Gram-negative bacteria are important in the pathogenesis of liver and systemic diseases. Chronic liver diseases can influence gastrointestinal motility, which together with other factors may contribute to bacterial overgrowth and in patients with ascites to an increased risk of spontaneous bacterial peritonitis. Patients with end-stage liver disease frequently develop portal hypertension leading to varices, gastric vascular ectasia, and portal hypertensive gastroenteropathy. Several liver and biliary abnormalities are observed in patients with inflammatory bowel disease (primary sclerosing cholangitis, autoimmune hepatitis, cholelithiasis). The primary defect in hemochromatosis is located in the intestine, causing an inappropriate increase in iron absorption, and the liver is the site of earliest and heaviest iron deposition. Elevated transaminases are observed in many patients with celiac disease, and steatohepatitis frequently develops in patients with jejunoileal bypass and short bowel syndrome. Furthermore, the liver is the primary organ for metastasis of intestinal cancer. Many viral, bacterial, fungal, and parasitic diseases affect the intestine as well as the liver and the biliary tract. PMID- 10855548 TI - Genetic analysis of multiple sporadic colon carcinomas from a single patient. AB - At least two separate genetic pathways of carcinogenesis in sporadic colon cancer involving the accumulation of mutations at various genetic loci have been described. About 15% of sporadic colorectal carcinomas arise via a mechanism associated with microsatellite instability (MSI) and mutations in transforming growth factor beta receptor II (TGFbetaRII), insulin-like growth factor II receptor (IGFIIR) and BAX, whilst the remaining 85% are associated with aneuploidy and gross chromosomal rearrangements. An 81-year-old woman had a sigmoid colon carcinoma resected and 18 months later developed two additional carcinomas of the caecum and transverse colon. To investigate whether there was a common genetic mechanism of carcinogenesis for the three lesions, MSI status was assessed, TGFbetaRII, IGFIIR and BAX were analysed for mutations and protein expression of transforming growth factor beta1 (TGFbeta1) and p53 were studied using immunohistochemistry. The caecal and transverse colonic carcinomas were both MSI positive but different mutations were identified in each lesion. No genetic abnormalities were identified in the sigmoid colonic carcinoma. This suggests that each carcinoma arose via a separate genetic mechanism of carcinogenesis. PMID- 10855549 TI - Relationship between sphincter morphology on endoanal MRI and histopathological aspects of the external anal sphincter. AB - Atrophy of the external anal sphincter can be shown only on endoanal magnetic resonance imaging (MRI). Until now no study has compared the morphological endoanal MRI findings with histopathological aspects of the external anal sphincter. The aim of this study was to validate the MRI interpretation of the external anal sphincter using histology as a "gold standard." In this prospective study 25 consecutive unselected women (median age 48 years, range 27-72) with fecal incontinence due to obstetric trauma were assessed preoperatively with endoanal MRI. All patients underwent anterior sphincteroplasty within 6 months of the preoperative assessment. During sphincter repair, a biopsy specimen was taken both from the left and right lateral parts of the external anal sphincter. Interpretation of MRI was performed by one of the radiologists (J.S.), and biopsy specimens were evaluated by the pathologist (W.J.M.). Both were blinded to the interpretation of the other. MRI revealed external anal sphincter atrophy in 9 of the 25 patients (36%). Histopathological investigation confirmed these findings in all but one. In one additional patient atrophy was detected on histological investigation while the morphology of the external anal sphincter was classified as normal on MRI. In detecting sphincter atrophy endoanal MRI showed 89% sensitivity, 94% specificity, 89% positive predictive value, and 94% negative predictive value. MRI correctly identified sphincter morphology in 23 of 25 cases (92%). This study demonstrates that endoanal MRI accurately identifies normal and abnormal external anal sphincter morphology. Endoanal MRI is therefore a valuable preoperative diagnostic tool. PMID- 10855550 TI - A pilot assessment of whether external coil MRI is useful to assess evacuatory disorders. AB - This study assessed the value of common surface coil magnetic resonance imaging (MRI) in patients with evacuatory disorders including fecal incontinence and constipation. These findings were then compared with those from other standard physiological examinations and/or surgical findings. From July 1996 to June 1997, 14 consecutive patients underwent surface coil MRI for evaluation of either fecal incontinence (n=5) or constipation (n=9). In patients with incontinence we compared the findings from endoanal ultrasound (EAUS), anal MRI, and surgery regarding morphopathological findings of the internal and external anal sphincter components. In constipated patients the findings of videoprography and dynamic pelvic MRI were compared regarding the presence of rectocele, rectoanal intussusception, and sigmoidocele as well as the measurements of anorectal angle and perineal descent. The five incontinent patients were all women, with a median age of 67 years (range 43-77). EAUS revealed an anterior sphincter defect in two patients, a posterior defect in one, and normal anal sphincter images in two. Surgical findings confirmed an anterior external anal sphincter scar in two patients, an internal anal sphincter defect in one, and an anatomically normal anal sphincter in two. In one patient, although anal MRI showed posterior external anal sphincter defect, EAUS and surgery revealed normal external anal sphincter appearance. The accuracy rate between EAUS and anal MRI was only 20%, that between surgery and anal MRI 40%, and that between surgery and EAUS 80%. Thus EAUS was more accurate than anal MRI in incontinent patients. The nine constipated patients were all women, with a mean age of 59 years (range 40-78). Videoproctography revealed an anterior rectocele in six patients, rectoanal intussusception in three, and sigmoidocele in five; no abnormalities were identified in two patients. On dynamic pelvic MRI anterior rectocele was seen in three patients and sigmoidocele in two, and five studies were interpreted as normal. One of the patients underwent sigmoidectomy for sigmoidocele, and five patients were treated by biofeedback. Thus the accuracy rate of dynamic pelvic MRI against videoproctography was 60% for anterior rectocele, 40% for sigmoidocele, and zero for rectoanal intussusception. In conclusion, neither MRI for the evaluation of patients with fecal incontinence nor for the evaluation of patients with constipation added any significant information that would warrant its continued use in these patient groups. Perhaps the more widespread availability of an endoanal coil will alter this conclusion; however, at the present time we cannot routinely endorse the expense, time, or inconvenience of these MRI investigations in patients with these diagnoses. Larger prospective comparative studies are required prior to endorsing the technique. PMID- 10855551 TI - Complications following formalin installation in the treatment of radiation induced proctitis. AB - Formalin installation has been safely and effectively used to treat refractory bleeding caused by radiation proctitis. This study evaluated the results of such treatment in terms of outcome and complications. All four patients who underwent formalin irrigation for transfusion-dependent radiation proctitis over a 15-month period were evaluated retrospectively. The procedure was performed under sedation in the operating room, with patients in the prone jack-knife position. A solution of 4% formalin was introduced in aliquots of 50 ml kept in contact with the mucosa for 30 s and then cleared away using saline irrigation; five to six aliquots were used in each session. In a fifth patient formalin-soaked gauze pads were applied directly to the injured mucosa. At a mean follow-up of 18 months (range 6-26) two patients had repeat episodes of bleeding, one underwent successful repeat irrigation, and the other refused further treatment. One patient suffered from severe anococcygeal pain and worsening of incontinence after the procedure. The pain was treated with lidocaine ointment and sitz baths with partial success. Another patient developed severe formalin-induced colitis 5 days after the procedure, which required intravenous antibiotics and hydration. Formalin installation may be effective in controlling refractory bleeding due to radiation induced proctitis. The procedure, however, is not risk free and may induce major complications such as acute colitis. PMID- 10855552 TI - Rectal pacing in patients with constipation due to rectal inertia: technique and results. AB - In a previous study we determined the rectal pacing parameters needed for rectal evacuation in patients with rectal inertia. Here we investigated the effect of rectal pacing on rectal myoelectric activity, motility, and evacuation in ten patients with constipation due to rectal inertia. A pacemaker was implanted in a subcutaneous pocket above the inguinal area, with a lead threaded in the anal submucosa to be hooked at the rectosigmoid junction. The effect of rectal pacing on rectal electric activity was investigated by inserting two recording electrodes to the rectal mucosa. The patients were then trained for home pacing. No waves were recorded from the rectum at rest. On rectal pacing, slow waves or pacesetter potentials (mean frequency 2.3+/-1.1 cpm, amplitude 0.86+/-0.1 mV, velocity 3.4+/-1.6 ms) were registered after a latency period of 5.2+/-1.6 min. Rectal evacuation, on pacing, occurred in seven of the ten patients. The three who showed no significant response exhibited low wave parameters. Three of seven patients were able to evacuate spontaneously without pacing after having performed daily pacing for 5-6 months. The pacemaker was removed in six patients (three failures and three after spontaneous defecation). Thus rectal pacing succeeded in inducing rectal evacuation in 70% of the patients. The procedure failed in three patients. Three had spontaneous defecation after a few months of rectal pacing. No complications were encountered, and the method was tolerated and acceptable. Further studies on a large group of patients are required. PMID- 10855553 TI - Role of caspase-3 in apoptosis of colon cancer cells induced by nonsteroidal anti inflammatory drugs. AB - Epidemiological studies have demonstrated that nonsteroidal anti-inflammatory drugs (NSAIDs) decrease the incidence of and mortality from colon cancer. In addition, NSAIDs reduce the number and the size of polyps in patients with familial adenomatous polyposis. The mechanisms responsible for the antineoplastic effect of NSAIDs are not yet completely understood, but one of the possible mechanisms is an induction of apoptosis. We explored the role of caspase-3, a major apoptosis-executing enzyme, in NSAID-induced apoptosis of colon cancer cell line HT-29. Treatment of HT-29 cells with indomethacin induced a dramatic increase in caspase-3-like protease activity measured by a cleavage of the fluorogenic substrate Ac-DEVD-AMC. Western blot analysis showed that indomethacin treatment led both to decrease in procaspase-3 and to cleavage of its substrate poly(ADP-ribose) polymerase (PARP). Furthermore, the caspase-3-like protease inhibitor Ac-DEVD-CHO attenuated indomethacin-induced DNA fragmentation dose dependently. However, mRNA expression of CASP genes was not affected by the addition of indomethacin, highlighting the importance of posttranslational modification of this enzyme for the activation. These results suggest that NSAIDs, including indomethacin, induce apoptosis in colon cancer cells through a caspase-3 dependent mechanism which may contribute to the chemopreventive functions of these agents. PMID- 10855554 TI - The dilemma of gastroenterological oncology: we know a lot but we still achieve too little. PMID- 10855555 TI - Comparison of anastomotic microcirculation in coloanal J-pouches versus straight and side-to-end coloanal reconstruction: an experimental study in the pig. AB - Several studies have shown a lower rate of anastomotic leakages in patients with coloanal J-pouch reconstruction than in those with straight coloanal anastomosis following anterior resection of the rectum. This study investigated whether this difference is due to a better anastomotic microcirculation. Thirty-two healthy, adult Gottinger mini-pigs underwent anterior rectal resection. They were subsequently randomized to following four groups (eight pigs per group): straight end-to-end, side-to-end, small pouch (4 cm), and large pouch (8 cm) coloanal anastomosis. Bowel perfusion was measured before and after vessel ligature at predefined locations using laser Doppler flowmetry. After completion of the anastomosis microcirculation was investigated 1 cm above, below, and directly at the anastomotic site. Following vessel ligature there was a 25% drop in blood flow. After completion of the anastomosis there was a further decrease of 25% in the distal segment, while no changes were observed above the anastomosis. There were no statistical differences either before or after completion of the anastomosis between the various groups. It is concluded that anastomotic blood flow does not depend on the type of coloanal reconstruction in healthy pigs. PMID- 10855556 TI - Extraintestinal polyps in Peutz-Jeghers syndrome: presentation of four cases and review of the literature. Deutsche Peutz-Jeghers-Studiengruppe. AB - Peutz-Jeghers syndrome (PJS) is a rare hereditary disorder characterized by hamartomatous polyps in the gastrointestinal tract and typical pigment lesions. Extraintestinal polyps have rarely been reported. Possible sites include the respiratory tract, urogenital tract, and gallbladder. We here describe four cases of extraintestinal polyps in PJS patients and review the literature on the need for operative therapy of extraintestinal polyps in PJS. Three nonrelated patients were examined who had PJS and polyps in the gallbladder; the fourth patient had PJS and recurrent choanal polyps. Surgery has so far been performed only for symptomatic polyps: one laparoscopic cholecystectomy and removal of the choanal polyps for recurrent infections of the respiratory tract. The remaining two patients reported no symptoms from the extraintestinal polyps. No malignant transformation was found in these patients, nor has such been reported in the literature on PJS. The frequent observation of this manifestation in our patients raises the question of clinical management: Is prophylactic surgery indicated? Since malignant transformation of PJS polyps in the intestine is extremely rare we see no reason for operative therapy as long as the polyps are small and asymptomatic. Regular sonographic controls are recommended since the risk of malignant transformation cannot be ruled out at present. PMID- 10855557 TI - High level of lipoprotein(a) is a strong predictor for progression of coronary artery disease. AB - Elevated levels of serum lipoprotein(a) [Lp(a)] are reported to be associated with risk of atherosclerosis and thrombosis. Little is known about the influence of Lp(a) on the progression of coronary artery disease. We evaluated the association of serum Lp(a) and the long-term changes of angiographic severity in patients who underwent repeated coronary angiography at intervals of more than 2 years. We evaluated 70 patients, and divided them into 3 groups by angiographic findings. Median Lp(a) concentration was significantly higher in the progression group (N=36) than in the no-change group (N=23) or the regression group (N=11) (32.4 vs 22, 19.3 mg/dl, p<0.05). Furthermore, the progression group had more patients whose Lp(a) levels were greater than 30 mg/dl (p=0.006), while in the regression group all patients were under 30 mg/dl. Stepwise logistic regression analysis for progression of lesions showed that Lp(a) > or =30 mg/dl remained significant, giving an estimated odds ratio (OR) of 2.46 (p= 0.005). In the subgroup analysis, OR in patients with mild lesions was reduced to 2.05 (p<0.05) while in patients with severe lesions OR was increased to 3.39 (p=0.003). The serum Lp(a) level has a close correlation with angiographic progression, and may be an important predictor for progression. PMID- 10855558 TI - Effect of nephropathy on the composition of apolipoprotein-containing particles in NIDDM. AB - We investigated the effect of nephropathy on the composition of apolipoprotein containing particles in non-obese NIDDM patients with normocholesterolemia. Sixty seven normal control subjects (group A), 48 NIDDM patients without nephropathy (group B) and 36 NIDDM patients with nephropathy (group C) were studied. Apolipoprotein AI or B100 containing particles (Apo AI or Apo B100) were isolated by immunoaffinity columns prepared with monoclonal antibodies. The total cholesterol (CH), esterified cholesterol (EC) and free cholesterol (FC) content in these particles was analyzed. Both the EC/FC ratio levels in Apo AI and in Apo B100 in group C were significantly higher than those in group A or B. Both the CH in Apo AI/apolipoprotein AI ratio and in Apo B100/apolipoprotein B100 ratio levels in group C were significantly lower than those in group A or B. The insulin resistance index showed significant positive correlation with the EC/FC ratio levels in Apo AI and in Apo B100, and showed significant negative correlation with the CH levels in Apo AI/apolipoprotein AI ratio and the CH levels in Apo B100/apolipoprotein B100 ratio levels in group C. Those compositional changes of lipoproteins in NIDDM patients with nephropathy may reflect partial insulin resistance and deteriorating atherosclerosis. PMID- 10855559 TI - Age-related change in relationship between body-mass index, serum sialic acid, and atherogenic risk factors. AB - In order to clarify the significance of obesity in atherosclerotic risk at different ages, the relationship between the body-mass index, serum sialic acid concentration, and various atherosclerotic risk factors were investigated for healthy subjects in three different age groups, 35-39, 40-49 and 50-59 years old. The body-mass index correlated significantly with mean arterial blood pressure, fasting blood sugar, serum triglycerides, atherogenic index and serum uric acid in all age groups. The magnitudes of the association of body-mass index with mean arterial blood pressure, fasting blood sugar and serum triglycerides decreased with age, while those of the association with atherogenic index and serum uric acid were not different among the three age groups. Body-mass index did not show significant correlation with white blood cell count, platelet count or smoking in any of the age groups. Simple and multiple regression analyses showed that body mass index was significantly correlated with serum sialic acid in the 35-39-year old group, but not in the other two groups. Neither the percentage of obese subjects (body-mass index > 26.4) nor the mean values of body-mass index were different among the three groups. These results suggest that for younger people, the body-mass index is related more closely with some atherosclerotic risk factors (e.g. blood pressure, blood sugar and serum triglycerides), and obesity may be possibly more involved in the progression of atherosclerosis, compared to more elderly people. PMID- 10855560 TI - Cytotoxic effect of oxidized low density lipoprotein on macrophages. AB - Macrophage or macrophage-derived foam cell death is one of the characteristic events in the development of cell-poor lipid-rich cores of the advanced atherosclerotic plaques. Although the in vivo mechanism for the death of macrophages is unclear, one possible candidate for the agent which induces macrophage cell death is oxidized low density lipoprotein (Ox-LDL). To investigate the mechanism of Ox-LDL-induced macrophage cell death, we have recently employed macrophage cell genetics and isolated mutant cells resistant to the cytotoxic effect of Ox-LDL from mutagenized populations of murine macrophage derived J774 cells (Hakamata, H., Miyazaki, A., Sakai, M., Matsuda, H., Suzuki, H., Kodama, T., and Horiuchi, S. (1998) J. Lipid Res. 39, 482-494). The results obtained showed that one mutant form, JO21b cells, was characterized by reduced expression of type I and type II class A macrophage scavenger receptors (MSR AI/AII) with a concomitant decrease in the uptake of Ox-LDL. Moreover, peritoneal macrophages obtained from MSR-AI/AII-knockout mice showed a higher resistance to the cytotoxic effect of Ox-LDL compared to those of their wild-type littermates. From these results, we have concluded that Ox-LDL cytotoxicity to macrophages is enhanced by effective endocytic uptake of Ox-LDL through MSR-AI/AII. These findings imply a possibility that formation of the cell-poor lipid-rich core is also enhanced by MSR-AI/AII-mediated uptake of Ox-LDL and subsequent macrophage cell death in atherosclerotic lesions. PMID- 10855561 TI - Diminished fibrinolysis and thrombosis: clinical implications for accelerated atherosclerosis. AB - Obesity is associated with an increased risk of atherosclerotic coronary artery disease. Cytokines and oxygen-centered free radicals implicated in insulin resistance stimulate adipocyte and endothelial production of plasminogen activator inhibitor type-1 (PAI-1), the primary physiologic inhibitor of fibrinolysis, in vitro. In obese hyperinsulinemic animal models simulating insulin resistance, plasma PAI-1 activity is increased. As the cardiovascular risk profile in specific populations may differ, endogenous fibrinolysis in lean and obese subjects was characterized and the mechanisms underlying differences were identified. Obese subjects (body mass index > 26) exhibited increased blood levels of PAI-1 antigen compared with corresponding values in lean controls. Blood t-PA antigen differed as well, yet basal endogenous fibrinolytic activity was decreased because of the high PAI-1 activity. The increased PAI-1 level was associated with increased levels of immunoreactive insulin (IRI). In diabetic subjects, coronary atherectomy specimens exhibited strong positive PAI-1 immunostaining, suggesting that in the diabetic vascular wall, intramural fibrinolytic activity is diminished. Using the oral glucose tolerance test, patients with significant stenosis confirmed by coronary angiography exhibited increased sigmaIRI, sigmaBS, sigmaIRI/sigmaBS, and IRI at 120 min compared to subjects without significant stenosis. IRI at 120 min was closely correlated with the severity of coronary artery disease. These results indicate that adipocyte overproduction of PAI-1 by insulin induces decreased endogenous fibrinolytic activity and contributes to the accelerated coronary macroangiopathy in hyperinsulinemic obese subjects with insulin resistance. PMID- 10855562 TI - Idiopathic sudden sensorineural hearing loss: temporal bone histopathologic study. AB - We microscopically examined the temporal bones of 12 ears with idiopathic sudden sensorineural hearing loss (iSSNHL), 10 ears with presbycusis, 11 ears with normal hearing, and 8 unaffected contralateral ears of patients with iSSNHL. The degeneration of the spiral ligament, vascular stria, hair cells, dendrites, and apical spiral ganglion cells was greater in ears with iSSNHL than in the other groups. The apical ganglion cells were significantly more affected than the basal ganglion cells, and the spiral ganglion cell loss increased as a function of duration of iSSNHL. Cochlear ossification was found in 1 ear with iSSNHL, and hydrops in 2. These findings suggest a viral rather than a vascular or ruptured inner ear membrane origin for iSSNHL. PMID- 10855563 TI - Mathematical model explaining the sources of error in certain estimates of the gas exchange constants for the middle ear. AB - Mover-Lev and colleagues reported a carbon dioxide-oxygen time-constant ratio of 3.9 for transmucosal gas exchange in guinea pigs under conditions of a large positive oxygen pressure gradient and a negative carbon dioxide gradient. That ratio is much less than the value of 19 reported previously for monkeys and used in predictive models of middle ear pressure regulation. In this report, the mathematics that underlie models of transmucosal gas exchange are developed and the conditions that allow accurate estimation of time constants are defined. The results demonstrate that the experimental and analytic methods used by Mover-Lev et al do not control for certain confounding effects or concurrently measure all required system parameters. Under the most realistic conditions, their ratio of 3.9 represents a significant underestimation of a true value on the order of 10. Also, their expectation of nonvarying, transmucosal time constant ratios for pairings that include reactive gases is simplistic and true only for identical experimental contexts. PMID- 10855564 TI - Length of the eustachian tube and its postnatal development: computer-aided three dimensional reconstruction and measurement study. AB - Eleven normal human temporal bone-eustachian tube (ET) specimens obtained from 11 individuals whose ages were 3 months to 88 years were studied to investigate the path length along the ET lumen and its postnatal development by means of a computer-aided 3-dimensional reconstruction and measurement method. The path length of the ET lumen of the 3-month-old infant was 21.2 mm, and its growth was in correlation with age to attain its adult length (average, 37.00 +/- 4.16 mm). The ratio of the length of the cartilaginous portion together with the junctional portion to the length of the bony portion was 8:1 in an infant at the age of 3 months and 4:1 in adults. That the bony portion of the ET develops relatively more than the cartilaginous and junctional portions may cause this finding. In addition, there is a developmental shift in the orientation of the cartilaginous portion with respect to the bony portion of the ET. In children, the cartilaginous and bony portions are both aligned with the line that connects the pharyngeal orifice and the tympanic orifice. In adults, however, the cartilaginous portion is angled inferiorly and laterally from the bony portion. This change may reflect the relative growth of the face. PMID- 10855565 TI - Effect of WEB 2170 BS, platelet activating factor receptor inhibitor, in the guinea pig model of middle ear inflammation. AB - Platelet activating factor (PAF), a potent inflammatory mediator, is a biologically active phospholipid. Recent studies have shown that PAF may play an important role in the pathogenesis of otitis media (OM). WEB 2170 BS has been shown to be a PAF antagonist both in vitro and in vivo. In this study, the anti inflammatory effects of WEB 2170 BS were investigated in a guinea pig model of OM induced by middle ear (ME) inoculation of killed Staphylococcus aureus. The outcome of treatment was determined by measurement of myeloperoxidase activity in the samples of ME mucosa, evaluation of temporal bone histopathology, and the presence of ME fluid. The myeloperoxidase activity in the WEB 2170 BS-treated group was found to be significantly lower than that in the control group. Histopathology of the temporal bones indicated decreased inflammation in the treated group as compared to the controls. In addition, ME fluid was absent in 16 of the 20 ears of the 10 treated animals. These results demonstrate that WEB 2170 BS can produce significant anti-inflammatory effects in this model of OM. PMID- 10855566 TI - Laryngeal radionecrosis and hyperbaric oxygen therapy: report of 18 cases and review of the literature. AB - Laryngeal radionecrosis is a difficult late complication of radiotherapy. It is associated with hoarseness, edema, pain, weight loss, and upper airway obstruction. The medical treatment options are limited, and in severe cases, the patient may require tracheostomy or laryngectomy. We report clinical results in 18 patients treated with adjunctive hyperbaric oxygen (HBO) therapy for severe radionecrosis of the larynx. Of these 18 patients, 2 had grade 3 and 16 had grade 4 radionecrosis. The patients received a mean number of 41 HBO treatments (range, 6 to 80) at 2 atmospheres absolute for 2 hours, twice a day, 6 days a week. Thirteen patients (72.2%) had a major improvement after HBO therapy, and none of them required total laryngectomy. All patients preserved their voice and deglutition in good or normal condition. Five patients (27.8%) failed to have a good response to HBO and underwent total laryngectomy. One of these patients had local recurrence of his cancer 4 months later, and the other 3 had significant concurrent medical problems. The remaining patient received only 6 HBO treatments because of emergency heart surgery. These encouraging results are comparable to those of smaller previous studies suggesting that HBO has a beneficial effect in the management of advanced laryngeal radionecrosis. PMID- 10855567 TI - Airway flow dynamics and voice acoustics after autologous fascia augmentation of paralyzed vocal fold. AB - The aim of this study was to assess the effect of vocal fold medialization, accomplished by injection of autologous fascia, on airflow dynamics and voice acoustics. Ten patients with unilateral vocal fold paralysis were included. Flow volume spirometry, body plethysmography, and acoustic analysis of voice were performed within 1 week before injection of autologous fascia and 4 to 14 months after operation. Medialization of the paralyzed vocal fold decreased the mean peak inspiratory flow (PIF) from 4.63 L to 4.10 L (p = .012). The acoustic characteristics of the voice improved: the values of jitter, shimmer, and mean noise-to-harmonics ratio decreased significantly (p = .006, p = .017, and p = .047, respectively), and the mean maximal phonation time almost doubled (p = .002). Changes in PIF and shimmer showed a negative correlation (r = -.857, p = .007). In conclusion, injection of autologous fascia improves voice acoustics, but induces a slight abnormal limitation on PIF. The results also suggest that improvement in voice acoustics is most prominent in subjects with the least deterioration in inspiratory airflow. PMID- 10855568 TI - Effects of dehydration on phonation in excised canine larynges. AB - The effects of exposure to dry air on phonation were measured in an ex vivo model of vocal fold vibration. Excised canine larynges were mounted on an apparatus and made to phonate at a constant subglottal pressure by means of unhumidified airflow. The phonation threshold pressure (PTP), glottal airflow, sound intensity of the acoustic output, and effects on vocal efficiency were also assessed. Student's t-test was performed on the results. In 17 larynges, the average PTP increased from 10.0 cm H2O to 15.0 cm H2O after exposure to dry airflow (p < .001). In addition, the average flow increased from 585 mL/s to 801 mL/s at a constant suprathreshold subglottal pressure (p < .001), and from 323 mL/s to 610 mL/s at the PTP (p < .001). The average acoustic output levels, measured during stable phonation segments, markedly decreased with exposure to the dry airflow, from 91.5 dB to 88.5 dB (p < .001). The average vocal efficiency decreased from 3.63 x 10(-4) to 7.00 x 10(-5) (p < .001). No such changes were seen in control larynges phonated with 100% humidified air used for driving the airflow. The results support previously reported modeling and experimental findings that dehydration of the vocal fold generally degrades laryngeal performance. PMID- 10855569 TI - Laryngeal reflexomyographic responses in rabbits: a neurolaryngological study of glottal movement. AB - The role of the laryngeal reflex in glottal movement has been reported, but its mechanism remains unclear. To further investigate the neurophysiological characteristics of glottal movement, we recorded the laryngeal reflexomyographic responses (LRMRs) to electrical stimulation of the superior laryngeal nerve (SLN) in rabbits. The procedure involved simultaneous recording of the LRMRs from the thyroarytenoid muscles by means of bipolar hooked wire electrodes after electrical stimulation to the SLN. The results demonstrated characteristic patterns of the responses, consisting of R1 and R2, similar to those found in humans. The R1 response was obtained with a latency of 10.7 +/- 0.78 ms. The ipsilateral R2 response was obtained with a latency of 43.76 +/- 4.67 ms in all rabbits, and the contralateral R2 response with a latency from 42.6 to 50.2 ms in 4 rabbits. It was concluded that LRMRs may serve as a potential central laryngeal function test in the investigation of glottal movement control. PMID- 10855570 TI - Site of airway collapse in obstructive sleep apnea after uvulopalatopharyngoplasty. AB - The objective of this prospective study was to determine the site and pattern of upper airway collapse by a multiple-catheter technique in subjects demonstrated to have obstructive sleep apnea (OSA) after uvulopalatopharyngoplasty (UPPP). Standard diagnostic nocturnal polysomnography (PSG) was done on all subjects. The PSG recordings included electroencephalogram, electrooculogram, electrocardiogram, chin and leg electromyograms, nasal and oral airflow, and abdominal effort. Polysomnography with a multiport flexible airway Gaeltec catheter was performed in 22 subjects. The Gaeltec flexible airway catheter has 4 high-fidelity pressure sensors to aid in determining the primary site of airway collapse. The primary site of airway collapse was determined by differential pressure gradients between pressure ports and by visual inspection of the pressure tracings. Forty-two subjects with prior UPPP from a total of 60 (39 men and 3 women, ages 33 to 61) agreed to be to studied by the standard PSG technique. Thirty-five subjects complained of excessive daytime sleepiness. Ten had mild OSA, 10 had moderate OSA, 12 had severe OSA, and 10 were "normal." Of the 22 subjects who had airway catheter monitoring, 3 of the normals were reclassified as having upper airway resistance (mean peak negative esophageal pressure of -28 cm H2O); 2 patients demonstrated airway obstruction in the nasopharynx, 2 at the oropharynx, and 11 at the level of the hypopharynx. Postoperative nocturnal PSG data were compared to data gathered prior to UPPP. The mean respiratory disturbance index (RDI) for the catheter group was 54 events per hour prior to UPPP, and the mean RDI after surgery was 44. There was no correlation between the severity of OSA and the stage of sleep. We conclude that the majority of patients who complain of excessive daytime sleepiness following UPPP have OSA with the primary site of obstruction at the level of the hypopharynx. The severity of airway collapse is variable during each stage of sleep. Esophageal pressure monitoring during sleep should be considered when evaluating symptoms of persistent OSA in patients who have had UPPP. PMID- 10855571 TI - Pharyngeal pressure analysis by the finite element method during liquid bolus swallow. AB - The human pharynx is unique, acting as a complex interchange between the oral cavity and the esophagus, and between the nasal cavity and the lungs. It is actively involved in the transport of food and liquid, producing the forces that guide the bolus into the upper esophagus and away from the adjacent larynx and lungs. This study developed a biomechanical computer model of the human pharynx, utilizing a finite element method (FEM). Control 2-dimensional cine computed tomography images were obtained during 10-mL barium paste swallows at 8 levels extending from the tongue base to the cricopharyngeal level in order to encompass the entire pharynx. Three-dimensional finite element models of the pharynx were reconstructed from the geometric information obtained from the images at each level. Using an inverse dynamic approach with the addition of known tissue properties, we analyzed the 8 models under estimated pressure histories during swallow. Within each model, changes in the cross-sectional intraluminal area were calculated and compared with the area from the computer-generated FEM model. Area matching allowed estimation of intraluminal pressure gradients during swallow. The estimated pressure gradients were distributed through a range from 10 to 55 mm Hg, varying from one region to another and showing different patterns for the upper 4 levels and the lower 4 levels. The contraction velocity for the upper 4 levels was much higher than that for the lower 4 levels. The higher contraction velocities and pressure gradients in the upper levels are consistent with the bolus velocities required for efficient swallow. PMID- 10855572 TI - Carcinoma of the hypopharynx and cervical esophagus in young adults. AB - Squamous cell carcinoma of the hypopharynx and cervical esophagus usually presents in the late-middle-aged and elderly. When diagnosed in young adults, the disease process is often thought to be more aggressive and have a worse long-term outcome. Four hundred ninety patients presented to the Christie Hospital and Manchester Royal Infirmary between 1981 and 1990 with squamous cell carcinoma of the hypopharynx and cervical esophagus. Of this group, 24 patients (5%) received their diagnosis before the age of 45. A comparison is made with a control group of 156 (32%) patients who presented between the ages of 60 and 69 years. Analysis of tumor and nodal staging at presentation demonstrates no statistically significant difference between the 2 age groups. There is a higher incidence of a combination of smoking and alcohol abuse in the older age group, but it is of no statistical significance. There is no difference in 5-year survival results between the 2 groups. We conclude that patients with squamous cell carcinoma of the hypopharynx and cervical esophagus who receive their diagnosis under the age of 45 show no difference in tumor stage or long-term outcome when compared with a control group encompassing the mean age of presentation. PMID- 10855573 TI - Secretory differentiation of serially passaged normal human nasal epithelial cells by retinoic acid: expression of mucin and lysozyme. AB - The purpose of this study was to subculture normal human nasal epithelial (NHNE) cells without compromising their ability to differentiate into secretory and ciliated cells and to study the effect of retinoic acid on mucous and serous secretions in passaged cells and to compare the expression of mucin and lysozyme in cultured cells with those in in vivo nasal epithelium. The subcultured cells were tested after every passage for secretory differentiation in air-liquid interface cultures. The cultured NHNE cells secreted mucin and lysozyme. The cells became squamous and mucin secretion decreased when retinoic acid was deleted from the culture media. Cells from passage 1 through passage 2 remained able to differentiate into mucous or squamous cells. Mucin gene 4 (MUC4), MUC5AC, MUC7, MUC8, and lysozyme messenger RNAs were expressed in passage 2 NHNE cells. In conclusion, passage 2 NHNE cell cultures retain features of normal epithelium and are suitable for many studies of upper airway cell biology. PMID- 10855574 TI - Laryngeal diversion and tracheotracheal speech fistula for chronic aspiration. AB - Intractable aspiration is a life-threatening problem and often requires a procedure for blocking or separating the larynx from the bronchial tree. The disadvantage of these techniques is a compromise of phonation. We report the use of a speech fistula after laryngotracheal diversion to restore voice. It allows for the definitive treatment of aspiration, while maintaining the use of the vocal folds for phonation. PMID- 10855575 TI - Actinomycosis imitating nasopharyngeal carcinoma. AB - The incidence of nasopharyngeal actinomycosis is exceedingly rare. To our knowledge, only 1 case has been reported previously. This article presents 4 cases of actinomycosis involving the nasopharynx that imitated nasopharyngeal carcinoma. Acidic gas exposure and farming of aquatic products are possibly involved in the pathogenesis of nasopharyngeal actinomycosis. Patients in the case studies completely recovered after 4 weeks of oral antibiotic treatment. We recommend including actinomycosis in the differential diagnosis of nasopharyngeal neoplasms. PMID- 10855576 TI - Sphenoethmoidal mucocele presenting with bilateral visual compromise. PMID- 10855577 TI - Herpes simplex viral laryngitis. AB - The true incidence of herpetic infections of the larynx is unknown. This entity may be underreported because of the difficulty in establishing the diagnosis. This report describes an immune-competent patient in whom extubation failed because of mass lesions of the posterior glottis. A biopsy specimen of the lesions revealed herpes simplex virus. We review the clinical presentation and histopathologic findings in this patient. PMID- 10855578 TI - Poverty--still a health hazard. PMID- 10855579 TI - Tuberculosis in New Zealand: why do we have twice as much as Australia? PMID- 10855580 TI - Prioritisation and cardiac events while waiting for coronary bypass surgery in New Zealand. PMID- 10855581 TI - A school and community outbreak of tuberculosis in Auckland. AB - AIM: To describe a school and community outbreak of tuberculosis in South Auckland in 1997/8. METHODS: Cases were diagnosed according to national guidelines at Middlemore, Green Lane and Starship Hospitals. Public health follow up was conducted by Auckland Healthcare. RESULTS: Twelve cases were diagnosed during the outbreak. Nine cases were from the same South Auckland secondary school; six reported no association outside school. Three cases were in younger children who had close household contact with two of the school cases. Nine cases (including eight from the school) had identical Mycobacterium tuberculosis isolates on restriction fragment length polymorphism testing. No microbiological culture was obtained from the three remaining cases. Contact investigation detected five of the cases. Chemoprophylaxis was prescribed for twenty-six school students, two adult staff, and nine household contacts. CONCLUSION: This is the first published account of a tuberculosis outbreak in a New Zealand school setting for decades. Recognition of the outbreak was delayed. DNA fingerprinting played a valuable role in the investigation. The source case may have been a school student. The social impact of the outbreak and preventability with routine adolescent BCG vaccination are discussed. PMID- 10855582 TI - Problem drinking profiles of patients presenting to general practitioners: analysis of Alcohol Use Disorders Identification Test (AUDIT) scores for the Auckland area. AB - AIM: To quantify the prevalence and demography of at-risk and problematic drinkers in the population attending a random selection of general practices and to compare this with similar studies. METHOD: A study examining the uptake and utilisation of the "DRINKLESS" package to 369 New Zealand general practitioners was conducted during 1995/6. The "DRINKLESS" package was developed with the World Health Organisation collaborative study for brief intervention for at-risk alcohol consumption. The package uses the Alcohol Use Disorders Identification Test (AUDIT). There were 15,670 completed AUDIT questionnaires collected during the study. These were analysed to ascertain the prevalence and demography of at risk and problematic drinkers attending general practitioners. RESULTS: There were 16% of patients identified as having either "risky drinking" or "problematic or dependent drinking". This pattern varied according to the occupation, age and gender of patients. CONCLUSIONS: The data confirm that large numbers of patients presenting to general practitioners experience alcohol problems of varying degrees. This study also suggests that the AUDIT will have satisfactory detection rates in a primary care setting. PMID- 10855583 TI - Blood donation by healthy individuals with haemochromatosis. AB - AIM: To determine how many individuals with haemochromatosis undergoing therapeutic venesections in our department might be eligible for blood donation. METHODS: Patients with genetic haemochromatosis were assessed with respect to complications of their disorder, the presence of other medical conditions and their suitability to be blood donors. RESULTS: Of 74 patients, 53 have been tested and shown to be homozygous for the Cys282Tyr mutation, and 30 of these (40%) fulfilled criteria for blood donation. This group is having 409 units of blood removed annually or 13-units per individual that is currently discarded. Of these 30 patients, all (100%) were keen to be blood donors. CONCLUSION: It is timely to review the policy regarding use of this wasted blood for transfusion. PMID- 10855584 TI - Laboratory expenditure in Pegasus Medical Group: a comparison of high and low users of laboratory tests with academics. AB - AIMS: To determine, through the use of clinical vignettes, whether low and high cost users of laboratory tests in Pegasus Medical Group (Pegasus) differed in their choice of laboratory tests from academics as a means of further investigating issues relating to quality and cost in laboratory testing. METHODS: Seven clinical vignettes were drawn up and sent to 30 selected members in Pegasus whose actual laboratory expenditure per consultation ranged from a mean of $2.3 in a low cost group (15 members) to $12.2 in a high cost group (15 members). The vignettes were also sent to 15 general practitioner academics. Respondents were requested to complete a laboratory form as to which tests they would use for each individual scenario. The answers were analysed for overall cost as well as numbers of laboratory tests requested. RESULTS: There were 14 academic responses and 13 each from the bottom and top laboratory users. Overall results for the seven vignette cases showed that low cost laboratory users would spend a total of $176.3, the academics $188.8, and the high cost users $219.5 on the cases. The mean per case costs were $25.2, $27.0 and $31.4 respectively. There was a clear tendency for high volume users of tests in each vignette to be high in others suggesting that doctor rather than patient factors were the main explanation of the variation. CONCLUSIONS: Clinical vignettes do not appear to be a useful strategy in clarifying issues related to quality and cost in laboratory utilisation. Test ordering behaviour appears, from the international literature and this study, to be determined more by personal doctor factors than by objective evidence and clinical need. Further work is needed to clarify the relationship between quality and the wide variation observed in utilisation and expenditure. PMID- 10855585 TI - Cricketing injuries in children: from the trivial to the severe. AB - AIM: To describe the nature of acute cricketing injuries in children presenting to the emergency department of a tertiary level children's hospital. Two cases of severe injuries during a cricket game are reported. METHOD: A retrospective review of presentations to the emergency department from 1993 to April 1998. RESULTS: Sixty cases of cricketing injuries were reviewed. Injuries to the head, hands and forearms were most common. Most injuries were caused by being hit by a ball. A high proportion of cases required operative intervention. Length of stay in hospital was only overnight in most cases. The two case reports highlight unusual but severe injuries that caused significant morbidity to the patients involved. CONCLUSION: Although cricket is, by and large, a safe sport, this report will raise awareness of the variety of injuries that can be suffered by children playing the game. PMID- 10855586 TI - Live donors for liver transplantation? PMID- 10855587 TI - Thrombocytopenia with the platelet aggregation inhibitor, Reopro. PMID- 10855588 TI - Altruism can have its rewards. PMID- 10855589 TI - The Locum's lament. PMID- 10855590 TI - Hepatitis B vaccine: why offer boosters? PMID- 10855591 TI - Hepatitis B vaccine: why offer boosters? PMID- 10855592 TI - Venlafaxine and fibromyalgia. PMID- 10855593 TI - Oral contraceptives and venous thromboembolism continued. PMID- 10855594 TI - Physician perceptions of HMO care for older persons. Health Maintenance Organization. AB - OBJECTIVE: Physician attitudes may be a key factor in effective managed care for older patients. We sought to explore physicians' views of the influence of health maintenance organization (HMO) policies on the care of their older patients. DESIGN: A self-administered one-page questionnaire consisting of questions about physician demographics, the impact of HMOs on physician practice, patient care, HMO policies, and respondents' personal use of managed health care plans. PARTICIPANTS: The survey was mailed to 838 randomly selected primary care physicians affiliated with two large, nonprofit, academically-oriented, Medicare HMOs in Massachusetts. RESULTS: Completed surveys were received from 516 of 797 eligible primary care physicians, affiliated with either Secure Horizons (Tufts Associated Health Plan) or First Seniority (Harvard Pilgrim Health Care). About half (55%) of the physician respondents reported they had sufficient time to spend with their older patients. Most (81%) respondents indicated that overall, patients aged 65 and older received either better care or no change in care after joining an HMO. The majority of physicians reported that HMO affiliation had increased the frequency with which they addressed geriatric issues with their older patients. There were several patterns of response that emerged with respect to beliefs about key HMO policies. CONCLUSIONS: The majority of physicians working in two nonprofit, academically oriented Medicare HMOs in Massachusetts believed that the overall quality of care that older patients received after joining an HMO either did not change or improved. PMID- 10855595 TI - Depressed mood and body mass index as predictors of muscle strength decline in old men. AB - OBJECTIVE: To study depressed mood as a predictor of strength decline within body weight categories over a 3-year follow-up period. DESIGN: A prospective cohort study over 3 years. SETTING: Honolulu, Hawaii. PARTICIPANTS: The subjects were 2275 men participating in the Honolulu Heart Program with an average age of 77.1 years (range 71-92 years), who were not cognitively impaired at baseline (Exam 4), and who participated in maximal hand grip strength measurements at baseline and 3 years later (Exam 5). MEASUREMENTS: Hand grip strength was measured using a dynamometer. Depressive symptoms were studied using an 11-item version of Center for Epidemiologic Studies Depression Scale with 9 as a cutoff. Body weight categories were formed on the basis of body mass index (BMI) (BMI = weight/height2; underweight: BMI < 20; normal weight: BMI 20-24.99, overweight: BMI > or = 25). MAIN RESULTS: At baseline, 9.4% of the participants were rated as having depressed mood. The mean individual strength change over 3 years was - 6.9% (standard deviation 14.0). Steep strength decline was determined as losing > or = 14% (lowest quartile). The proportions of those with steep strength decline in the groups based on combined distributions of BMI and depressed mood were: underweight/ depressed (n = 22) 41%, underweight/not depressed (n = 200) 28%, normal weight/depressed (n = 127) 30%, normal weight/not depressed (n = 1181) 25%, overweight/depressed (n = 55) 31%, overweight/not depressed (n = 675, referent) 21%. After adjusting for baseline strength, age, height, sociodemographic variables and diseases, the odds ratio for steep strength decline was more than four times greater among those who were depressed and underweight, and twice as great among people who were depressed and normal weight compared with those who were nondepressed and overweight. The risks of nondepressed under- and normal weight people and depressed overweight people did not differ from the reference group. CONCLUSIONS: Depressed mood was associated with increased risk of steep strength decline, in particular in older men with low body weight. Low body weight in combination with depressed mood may be an indicator of frailty or severe disease status that leads to accelerated strength loss and disability. PMID- 10855596 TI - Delirium is independently associated with poor functional recovery after hip fracture. AB - OBJECTIVE: To evaluate the role of delirium in the natural history of functional recovery after hip fracture surgery, independent of prefracture status. DESIGN: Prospective cohort study. SETTING: Orthopedic surgery service at a large academic tertiary hospital, with follow-up extending into rehabilitation hospitals, nursing homes, and the community. PARTICIPANTS: One hundred twenty-six consenting subjects older than 65 years (mean age 79 +/- 8 years, 79% women) admitted emergently for surgical repair of hip fracture. MEASUREMENTS: Detailed assessment at enrollment to ascertain prefracture status through interviews with the patient and designated proxy and review of the medical record. Interviews included administration of standardized instruments (Activities of Daily Living (ADL) Scale, Blessed Dementia Rating Scale, Delirium Symptom Interview) and assessment of ambulation, and prefracture living situation. Medical comorbidity, the nature of the hip fracture, and the surgical repair were obtained from the medical record. All subjects underwent daily interviews for the duration of the hospitalization, including the Mini-Mental State Examination and Delirium Symptom Interview, and delirium was diagnosed using the Confusion Assessment Methods algorithm. Patients and proxies were recontacted 1 and 6 months after fracture, and underwent interviews similar to those at enrollment to determine death, persistent delirium, decline in ADL function, decline in ambulation, or new nursing home placement. RESULTS: Delirium occurred in 52/126 (41%) of patients, and persisted in 20/52 (39%) at hospital discharge, 15/52 (32%) at 1 month, and 3/52 (6%) at 6 months. Patients aged 80 years or older, and those with prefracture cognitive impairment, ADL functional impairment, and high medical comorbidity were more likely to develop delirium. However, after adjusting for these factors, delirium was still significantly associated with outcomes indicative of poor functional recovery 1 month after hip fracture: ADL decline (odds ratio (OR) = 2.6; 95% confidence interval (95% CI), 1.1- 6.1), decline in ambulation (OR = 2.6; 95% CI, 1.03-6.5), and death or new nursing home placement (OR = 3.0; 95% CI, 1.1-8.4). Patients whose delirium persisted at 1 month had worse outcomes than those whose delirium had resolved. CONCLUSIONS: Delirium is common, persistent, and independently associated with poor functional recovery 1 month after hip fracture even after adjusting for prefracture frailty. Further research is necessary to identify the mechanisms by which delirium contributes to poor functional recovery, and to determine whether interventions designed to prevent or reduce delirium can improve recovery after hip fracture. PMID- 10855597 TI - Epidemiology of sarcopenia. AB - OBJECTIVES: To examine patterns of muscle mass change with aging and to estimate the prevalence of sarcopenia. DESIGN: Cross-sectional survey. SETTING: Population based study in Rochester, Minnesota. PARTICIPANTS: Age-stratified sample of men and women from the community. MEASUREMENTS: Muscle mass estimated from total body scans by dual-energy X-ray absorptiometry. Sarcopenia was defined as muscle mass more than 2 standard deviations below the sex-specific young-normal mean. RESULTS: Total lean body mass (exclusive of bone) and total skeletal muscle mass both were greater in men than women and declined linearly with age as judged from these cross-sectional data. Adjustment for height reduced the gender difference. The age- and sex-adjusted prevalence of sarcopenia varied from 6 to 15% among subjects 65 years of age or over, depending on the muscle mass parameter that was evaluated, but prevalence rates were quite sensitive to the normal values used to define cutoff levels. Subjects with sarcopenia appeared to have more physical limitations than the others. CONCLUSIONS: Late in life, a substantial portion of the population reaches low levels of muscle mass that are associated with increased physical disability. However, additional efforts are needed to validate an operational definition of sarcopenia. PMID- 10855598 TI - Predicting 10-year care requirements for older people with suspected Alzheimer's disease. AB - OBJECTIVE: To describe the types and costs of care received for 10 years after the identification of an older person with suspected Alzheimer's disease (AD) by using data from 3254 patients with suspected AD who participated in the National Long Term Care Survey (NLTCS). METHODS: By using a Markov model derived using grade of membership techniques, the following were determined: survival probabilities at 10 years; years of survival during the 10 years; years in institutions; years with two or more impairments in basic activities of daily living; hours of paid and informal care while the older person lived in the community; and costs of paid community, institutional, and medical care. RESULTS: Greater degrees of cognitive impairment present when AD was identified were associated with reduced predicted probability of surviving 10 years, increased predicted number of years spent in institutions, increased hours of care required while affected individuals remained in the community, and increased costs of paid community, institutional, and medical care. Substantial differences between men and women were seen: severity-adjusted 10-year costs were almost two times higher for women with AD than for men ($75,000 compared with $44,000); according to sensitivity analysis, average 10-year costs might be as high as $109,000 for women and $67,000 for men. CONCLUSIONS: AD imposes a substantial burden on older persons. Interventions that slow the progression of the disease may therefore affect community survival as well as healthcare costs. PMID- 10855599 TI - How much do persons with Alzheimer's disease cost Medicare? AB - BACKGROUND: Medicare claims are increasingly being used to identify persons with chronic diseases such as Alzheimer's disease (AD) for the purpose of determining the cost to Medicare of caring for such persons. Past work has been limited by the use of only 1 or 2 years of claims data to identify cases, leading to worries that this might lead to an undercount of prevalent cases and bias cost findings. OBJECTIVES: To analyze the average total cost to the Medicare program in 1994 of persons with a claims-based diagnosis of AD, using a 12-year period of claims history to identify prevalent cases, and to investigate the effect on cost of time since diagnosis. DESIGN: A cross-sectional design with a 12-year retrospective period to identify persons with AD. SETTING: Medical care practices, hospitals, and other providers of services to Medicare beneficiaries in the US in 1994. SUBJECTS: Respondents to the screener (n = 10,858) and community (5429) and institutional (n = 1341) questionnaire of the 1994 National Long Term Care Survey, with and without a claims-based diagnosis of AD. MEASUREMENTS: Average total cost to Medicare in 1994, measured as the actual amount Medicare paid for inpatient, outpatient, home health, skilled nursing facility, hospice, and Part B services, including payments to physicians, and other items such as durable medical equipment. We also measured disability in a variety of ways, including cognition, activity limitations, and residence in a nursing home. RESULTS: The average total cost to Medicare of persons with a claims-based diagnosis of AD was $6021 versus $2310 (P < .001) for persons without a diagnosis. When adjusting for patient characteristics, the ratio of cost between persons with AD and those without was reduced to about 1.6 to 1. Time since diagnosis was an important predictor of average total cost in 1994, with each additional year since diagnosis resulting in a $248 (P = .04) decrease in total cost (about 10% of the total sample mean cost of $2426). There was mixed evidence that persons with a diagnosis of AD incurred less cost than otherwise similarly disabled Medicare beneficiaries. CONCLUSIONS: Time since diagnosis with AD is an important predictor of cost and one that should be explicitly included in any rate-setting formula. Expanding the period used to identify cases resulted in an increase in the unadjusted ratio of cost of a Medicare beneficiary with AD relative to one without primarily because our control group costs are lower compared with those of past work. PMID- 10855600 TI - Silent ischemia during voluntary detraining and future cardiac events in master athletes. AB - OBJECTIVES: To determine whether exercise-induced silent ischemia in older master athletes following a 3-month period of deconditioning is a predictor of future cardiovascular events. DESIGN: A longitudinal study of a cohort of master athletes. SETTING: The Geriatric Research Education and Clinical Center (GRECC), Baltimore VA Medical Center, Baltimore, Maryland. PARTICIPANTS: Ten older (59 +/- 8 years, mean +/- SD), highly conditioned (maximal aerobic capacity VO2max 50 +/- 5 mL/kg/min), aerobically trained athletes. INTERVENTION: Five to eight years of longitudinal follow-up of athletes who had previously participated in a 3-month long detraining intervention. MEASUREMENTS: At baseline, all 10 athletes had their history taken and underwent physical examinations, metabolic testing, electrocardiogram at rest, exercise treadmill tests, exercise thallium scintigrams, and exercise multigated acquisition scans. After 3 months of deconditioning, they had repeat maximal exercise stress tests. After 5 to 8 years of follow-up, they were re-evaluated, including history and physical examination and measurement of their VO2max. RESULT: All 10 master athletes had normal studies at baseline. At the end of 3 months of detraining, three of these athletes had exercise-induced silent ischemia, which disappeared after retraining in two subjects and persisted at a higher heart rate in one subject. Over a 5- to 8-year period of observation, two of these three athletes with silent ischemia experienced major cardiac events (sudden death, cardiac bypass surgery). The other seven athletes did not have any cardiovascular events. CONCLUSIONS: Exercise-induced silent ischemia after a short period of detraining in highly trained older athletes may be a predictor of future cardiac events. A study with a larger cohort is warranted. PMID- 10855601 TI - Underutilization of controller and rescue medications among older adults with asthma requiring hospital care. AB - BACKGROUND: Asthma causes serious morbidity in older people, but pharmacologic therapy in older people with asthma has never been studied, at least in part because of the difficulty of defining asthma in this population. OBJECTIVE: To determine if older persons enrolled in Medicaid and hospitalized with an exacerbation of asthma receive appropriate outpatient asthma care. DESIGN: Descriptive pharmacoepidemiology of a group of older adults with asthma. SETTING: The Tennessee Medicaid Program. PARTICIPANTS: Persons aged 65 and older, enrolled in the Tennessee Medicaid program, identified through Medicaid's computerized database as having a hospital care visit for asthma in 1992 and who had their diagnosis confirmed by chart review. MEASUREMENT: Medication utilization. RESULTS: The source population included 93,686 persons aged 65 or older enrolled in the Tennessee Medicaid program. The group meeting study criteria included 512 patients with chronic asthma who had a hospital care visit for an asthma exacerbation. Eighty-one percent of these 512 persons with an asthma hospitalization confirmed by chart review were classified as having moderate to severe or potentially fatal asthma. These patients had had a median of 15 outpatient visits in the previous year, and more than half of them had an outpatient visit in the 14 days before their hospitalization. However, among those with moderate to severe or near fatal asthma only 25% filled prescriptions for inhaled corticosteroids, whereas 52% were taking theophylline, the most commonly prescribed asthma medication in this group. There was also high use of antibiotics (29%) and low use of rescue corticosteroids (5%) before the hospital care visit, despite frequent medical encounters. CONCLUSIONS: Despite widespread promulgation of the National Asthma Education Prevention Program guidelines, our study suggests that providers caring for indigent older subjects with moderate to severe or potentially fatal asthma were not following these guidelines. There was significant underutilization of inhaled anti-inflammatory agents, beta-agonists, and rescue corticosteroids in this population despite frequent outpatient medical care visits. PMID- 10855602 TI - Racial variations in end-of-life care. AB - OBJECTIVES: To identify differences in advanced care planning and end-of-life decision-making between whites and blacks aged 70 and older. DESIGN: The Asset and Health Dynamics Among the Oldest Old (AHEAD) study is a nationally representative survey of adults who were aged 70 and older in 1993. Relatives (proxy respondents) for 540 persons who died between the first (1993) and second (1995) waves of the study were surveyed about advanced care planning and end-of life decisions that were made for their family member who died. SETTING: Respondents were interviewed at home by telephone (n = 444) or in person (n = 95). PARTICIPANTS: The 540 proxy respondents included 454 whites and 86 blacks. MEASUREMENTS: Questions were asked about advance care planning and end-of-life decisions. RESULTS: Whites were significantly more likely than blacks to discuss treatment preferences before death (P = .002), to complete a living will (P = .001), and to designate a Durable Power of Attorney for Health Care (DPAHC) (P = .032). The treatment decisions for whites were more likely to involve limiting care in certain situations (P = .007) and withholding treatment before death (P = .034). In contrast, the treatment decisions for blacks were more likely to be based on the desire to provide all care possible in order to prolong life (P = .013). Logistic regression models revealed that race continued to be a significant predictor of advance care planning and treatment decisions even after controlling for sociodemographic factors. CONCLUSIONS: These findings suggest that there are important differences between blacks and whites regarding advanced care planning and end-of-life decision-making. Health professionals need to understand the diverse array of end-of-life preferences among various racial and ethnic groups and to develop greater awareness and sensitivity to these preferences when helping patients with end-of-life decision-making. PMID- 10855603 TI - The C677T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene and vascular dementia. AB - OBJECTIVE: To determine the association between the C677T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene and vascular dementia in Ashkenazi and non-Ashkenazi Jews. DESIGN: A case-control study. SETTING: Nursing homes in Jerusalem, Israel. PARTICIPANTS: Two hundred fifty nine Jewish people of Ashkenazi and non-Ashkenazi origin, older than age 70, who have vascular dementia (VD) (n = 85), Alzheimer's disease (AD) (n = 92), and who are cognitively intact (n = 82) with no clinical evidence of atherosclerotic vascular disease. MEASUREMENTS: The frequencies of the mutant allele (T allele) and homozygotes for the C677T MTHFR mutation (T/T genotype). The total plasma homocysteine (tHCT) level in 75 subjects. RESULTS: There were no significant differences in the frequencies of the T/T genotype or T allele among VD, AD, and cognitively intact older people of the same ethnic origin (0.15, 0.19, 0.25 T/T genotype and 0.42, 0.46, 0.47 T allele in Ashkenazi; 0.08, 0.06, 0.10 T/T genotype and 0.28, 0.32, 0.33 T allele in non-Ashkenazi with VD and AD, and in cognitively intact older people, respectively). The relative risk of VD associated with the T/T genotype versus the C/C genotype was 0.62 (95% CI, 0.19-1.19) in Ashkenazi and 0.65 (95% CI, 0.11-3.7) in non-Ashkenazi, respectively. The relative risk of AD associated with the T/T genotype was 0.85 (95% CI, 0.29-2.45) in Ashkenazi and 0.62 (95% CI, 0.1-4.3) in non-Ashkenazi, respectively. The frequencies of mutant homozygotes and allele were significantly higher in Ashkenazi than in non-Ashkenazi Jews (19.9% vs 7.5% T/T genotype, chi2 = 6.2, P = .01, 0.45 vs 0.31 T allele, chi2 = 9.77, P = .002 in Ashkenazi vs non-Ashkenazi, respectively). There were no differences in mean tHCT concentration among VD, AD, and cognitively intact older people. CONCLUSIONS: The MTHFR C677T is not associated with an increased risk of vascular dementia or Alzheimer's disease. The frequency of the mutation is significantly higher in Ashkenazi compared with non-Ashkenazi Jews. PMID- 10855604 TI - Hormone replacement therapy use in urban older women attending meal sites: associations with sociodemographic and health characteristics and use of preventive services. AB - OBJECTIVE: To examine sociodemographic, health and preventive health practices associated with hormone replacement therapy (HRT) use in urban community-dwelling older women. DESIGN: Survey. SETTING: Community-based meal sites throughout the city of Los Angeles. PARTICIPANTS: A convenience sample of 705 community-dwelling women older than age 60 who completed questionnaires for the Prevention for Elderly Persons Program. MEASUREMENTS: Demographic and life style characteristics, functional status, preventive practices, and current and past use of HRT. RESULTS: Among the 705 women surveyed, 13% reported current use and 17% reported past use of HRT. Current users were more likely to be younger and more likely to report a history of osteoporosis, hysterectomy, and calcium use than never users. White women were more likely to be current users than black women. CONCLUSIONS: Only a small proportion of the older urban women studied are currently using HRT. In particular, efforts to increase the use of these preventive services need to focus on black women and women who do not have a prior history of osteoporosis. PMID- 10855605 TI - Dually incontinent nursing home residents: clinical characteristics and treatment differences. AB - OBJECTIVE: Previous studies have described urinary and fecal incontinence in nursing homes and their separate effects on healthcare utilization. However, little is known about those who are incontinent of both. DESIGN: Retrospective chart review. SETTINGS: Twenty sites in three states PARTICIPANTS: A total of 413 nursing home residents were categorized as having neither fecal nor urinary incontinence (C, n = 114), urinary incontinence only (UI, n = 53), fecal incontinence only (FI, n = 9), or were dually incontinent (DI, n = 237). MEASUREMENTS: Charts were abstracted for sociodemographic information and health status information as well as utilization for the year before the date of abstraction. We then compared these characteristics across groups using ANOVA with pairwise comparisons and multiply adjusted regression. RESULTS: Almost all patients with DI were cognitively and mobility impaired. However, there were no significant differences between the groups with respect to age and number of diagnoses. A diagnosis of stroke was also more common among those with DI compared with C. When examining healthcare utilization in multiply adjusted regression, dually incontinent residents received significantly fewer days of hospital care than those with UI. CONCLUSIONS: Dual incontinence in NH residents is likely to have an important functional component. These residents seem to be treated less aggressively with respect to hospitalization compared with those with UI alone. The reasons for these differences need to be explored further. PMID- 10855606 TI - The involvement of physicians in VA home care: results from a national survey. AB - OBJECTIVES: To examine the role of physicians in the Veteran Affairs (VA) home based primary care (HBPC) program and to identify variables that predict whether physicians make home visits and volume of home visits made. DESIGN: Descriptive and regression analyses of responses from a mail survey. PARTICIPANTS: Forty-five physicians affiliated with VA HBPC programs. MAIN SURVEY TOPICS: Self-reported work load, attitudes toward home care, reasons for home visits, administrative policies regarding physicians' role in patient care management, and time commitment to home care. RESULTS: A majority of physicians believed strongly in the importance of home care and made home visits for reasons consistent with their training. Physician attitude toward home care and preoccupation with office or hospital practice were related to whether or not physicians made home visits. Degree of preoccupation with office practice and amount of salary support from VA HBPC were significant predictors of the number of visits made (R2 = 0.44). CONCLUSIONS: These findings indicate that most physicians will make home visits if they believe that home care is valuable and if their time commitment is supported financially. Managed care plans that own and operate home care programs and have the capacity to transfer primary care management to physicians who derive financial support from the programs should find this information particularly relevant. PMID- 10855607 TI - Benzodiazepines and the risk of falls in nursing home residents. AB - CONTEXT: For nursing home residents who require a benzodiazepine, short-acting agents are recommended, primarily to avoid increased risk of falls and other injuries associated with the long-acting agents. However, much of the data for the clinical outcomes of falls and injuries comes from community-dwelling older people. OBJECTIVE: To quantify the rate of falls among nursing home residents taking benzodiazepines and how this varies with drug elimination half-life. DESIGN: Historical cohort study. POPULATION: A total of 2510 residents of 53 Tennessee nursing homes, classified according to benzodiazepine use on each day of follow-up. OUTCOME MEASURES: Falls occurring during study follow-up. RESULTS: After adjustment for differences in resident characteristics, benzodiazepine users had a 44% increased rate of falls (adjusted rate ratio 1.44 [95% confidence interval, 1.33-1.56]). The adjusted rate ratio increased from 1.30 (1.12-1.52) for a dose equivalent to < or = 2 mg of diazepam, to 2.21 (1.89-2.60, P < .001) for a dose of > 8 mg. The rate of falls was greatest in the 7 days after the benzodiazepine was started (rate ratio of 2.96 [2.33-3.75]) but remained elevated (1.30 [1.17-1.44]) after the first 30 days of therapy. Drugs with elimination half-lives of <12, 12-23, and > or = 24 hours had adjusted rate ratios of 1.15 (0.94-1.40), 1.45 (1.33-1.59), and 1.73 (1.40-2.14), respectively. Users of hypnotics with elimination half-lives <12 hours had an increased rate of falls occurring during the night (adjusted rate ratio 2.82 [2.02-3.94]). CONCLUSIONS: Although the risk of falls among nursing home residents receiving short-acting benzodiazepines is less than that for the long-acting agents, these drugs are associated with a materially increased risk of nocturnal falls. PMID- 10855608 TI - Revisiting the one-year geriatric fellowship option: a preliminary assessment. AB - OBJECTIVE: To ascertain early opinions of geriatric fellowship program directors regarding the 1-year clinical training option. DESIGN: A brief mail survey of the 100 program directors listed in Graduate Medical Education, 1998-1999. MEASUREMENTS: Answers (favorable (+), no effect (0), or negative (-)) to questions regarding the effect on the training program, position of the director, and geriatrics as a discipline. RESULTS: This preliminary assessment produced decidedly mixed results. Seventy-six program directors responded (76% response rate). The clearest positive effect on the program itself was on the quantity of applicants (63% +, 4% -) but less so on their quality (33% +, 15% -). The clearest negative impact was on research aspects of the fellowship (0% +, 67%-). Other effects on the program were either null (clinical aspects and faculty morale) or moderately negative (educational, administrative, and financial aspects and the position of the program director), with few effects on the geriatrics division as a whole. Overall, the 1-year option was felt to have adversely affected geriatrics as a respected professional field (11% +, 41% -). Nevertheless, the majority (61%) answered that introduction of the 1-year option was wise. Although not attributed to this training pathway, program directors overwhelmingly perceived (75 % +) that geriatrics has enjoyed enhanced strength and attractiveness as a discipline since introduction of the 1-year option in 1992. CONCLUSIONS: The 1-year geriatric fellowship option has, at best, proved a mixed blessing to program directors. However, optimism regarding the future would seem to justify continuing to offer this option because of several observations and trends: (1) the increase in quantity and also perhaps quality of applicants; (2) growth in numbers of fellows and higher percentage program fill rates; (3) success of both 1- and two-year fellows in passing the Certificate of Added Qualifications examination and consequent increased generation of certified fellowship-trained geriatricians; (4) strong sentiment that geriatrics is gaining strength and attractiveness; and (5) that introduction of the 1-year option was wise. PMID- 10855609 TI - A ray of hope for older Nigerians. PMID- 10855610 TI - Catechol-O-methyltransferase (COMT) inhibitors in Parkinson's disease. AB - Catechol-O-methyltransferase (COMT) inhibitors are a new therapeutic option in the treatment of patients with Parkinson's disease. COMT inhibitors act by extending the duration of action of levodopa, thus improving the amount of time a patient can experience benefit from levodopa. COMT inhibitors are only used in conjunction with levodopa. They do have a propensity to augment dopaminergic effects, such that levodopa doses might need to be adjusted downward. Other side effects of COMT inhibitors include diarrhea and liver function abnormalities. Due to the latter, recent guidelines have been developed to monitor patients on tolcapone for this rare side effect, and these guidelines will be discussed. This article also provides representative case histories for the appropriate use of COMT inhibitors that illustrate how these drugs can be used to manage patients with a fluctuating response to levodopa. PMID- 10855611 TI - A statement of principles: toward improved care of older patients in surgical and medical specialties. PMID- 10855612 TI - The new frontier: increasing geriatrics expertise in surgical and medical specialties. PMID- 10855613 TI - The patient as teacher: a tribute to Clark M. Clifford. PMID- 10855614 TI - Quetiapine for sexually inappropriate behavior in dementia. PMID- 10855615 TI - Gait, balance, and self-efficacy in older black and white American women. PMID- 10855616 TI - A novel accelerometry-based method for the quantification of balance and postural sway. PMID- 10855617 TI - In re MediCaring. PMID- 10855618 TI - MIRDOSE 3 distribution suspended. PMID- 10855619 TI - The role of exercise radionuclide angiocardiography in predicting future cardiac events in patients with acute myocardial infarction. AB - Left ventricular ejection fraction (LVEF) during exercise radionuclide angiocardiography is a useful prognostic index for patients with acute myocardial infarction (AMI). However, most previous studies were performed before reperfusion therapies (i.e., thrombolysis and coronary angioplasty) were widely used. Therefore, because reperfusion therapy has become a standard therapeutic option, we reexamined the prognostic value of rest LVEF and exercise LVEF determined by radionuclide angiocardiography in patients with AMI at the time of hospital discharge. METHODS: The retrospective analysis included 419 consecutive patients with AMI who underwent ergometric stress radionuclide angiocardiography before hospital discharge, 44 +/- 14 d after the onset of AMI. RESULTS: During a mean follow-up of 4.6 y, cardiac events occurred in 101 (24.1%) patients. Cardiac events included recurrent MI (33 patients, 7.9%), unstable angina (49 patients, 11.7%), congestive heart failure (16 patients, 3.8%), and ventricular tachycardia (3 patients, 0.7%). The LVEF at peak exercise was significantly lower in the group with cardiac events (P = 0.0140). However, no significant difference was observed in the rest LVEF between patients with and without cardiac events. On the basis of multivariate analysis using a Cox proportional hazards model, only peak LVEF (P = 0.0246) was found to be an independent predictor of cardiac events. In the patient subsets with a peak LVEF >50% or <50%, the event-free rate was 81.0% versus 62.4% (P = 0.0007), respectively. Regardless of the presence or absence of reperfusion therapy, the lower peak LVEF was associated with a decrease in the event-free survival rate. CONCLUSION: In the current reperfusion era, the lower peak LVEF as measured by radionuclide angiocardiography at the time of discharge is a useful predictor of subsequent cardiac events in patients with AMI. PMID- 10855620 TI - Effects of left bundle branch block on myocardial FDG PET in patients without significant coronary artery stenoses. AB - Cardiac PET studies in patients with left bundle branch block (LBBB) are few, and the results are conflicting. In particular, even if a reduced uptake of FDG is reported, confirmation in a large group of patients and exact understanding of the underlying cause are lacking. METHODS: We selected 29 consecutive patients who had complete LBBB and no significant stenosis on coronary angiography scheduled for FDG and 13N-NH3 PET for myocardial viability evaluation at our center. Wall motion was evaluated using 2-dimensional echocardiography. Ten volunteers without coronary stenosis or LBBB served as a control group. RESULTS: All LBBB patients had a reverse mismatch in the septum, defined as reduced uptake of FDG in comparison with 13N-NH3. The mismatch extended to the anterior and inferior walls in 17 patients. The mean (+/-SD) septal-to-lateral ratio was 0.57 +/- 0.11 for FDG (range, 0.28-0.76) and 0.99 +/- 0.12 for 13N-NH3 (range, 0.75 1.18), with P < 0.0001. In contrast, no significant differences in uptake were seen in the control group, which had a septal-to-lateral ratio of 0.95 +/- 0.13 for FDG (range, 0.78-1.15; P < 0.01 with respect to LBBB patients) and 0.94 +/- 0.11 (range, 0.85-1.20) for 13N-NH3. CONCLUSION: Our study suggests that in LBBB patients without significant coronary stenosis, FDG uptake in the septum changes without a correlating change in perfusion. To avoid possible overestimation of necrosis, especially in the LAD territory, this phenomenon must be considered in evaluations of myocardial viability using FDG images. PMID- 10855621 TI - Coronary microangiopathy in type 2 diabetic patients: relation to glycemic control, sex, and microvascular angina rather than to coronary artery disease. AB - Coronary microangiopathy is a major complication in diabetics. However, the presence of independent factors in association with coronary microangiopathy in patients with non-insulin-dependent diabetes mellitus (NIDDM) or the difference in coronary microangiopathy between diabetics with coronary artery disease (CAD) and those with microvascular angina is unclear. METHODS: Nineteen patients with NIDDM and microvascular angina, 18 patients with NIDDM and CAD, and 17 age matched control subjects were studied. Myocardial segments that were perfused by angiographically normal coronary arteries were studied. The baseline myocardial blood flow (MBF) and the MBF during dipyridamole administration were measured using PET and 13N-ammonia, after which the myocardial flow reserve (MFR) was calculated to assess coronary microangiopathy. RESULTS: The baseline MBF was comparable among NIDDM patients with microvascular angina, NIDDM patients with CAD, and control subjects. However, the MBF during dipyridamole administration was significantly lower in NIDDM patients with microvascular angina (126 +/- 42.7 mL/min/100 g) than that in either NIDDM patients with CAD (210 +/- 70.1 mL/min/100 g; P < 0.01) or control subjects (293 +/- 159 mL/min/100 g; P < 0.01), as was the MFR (NIDDM with microvascular angina, 1.90 +/- 0.73; NIDDM with CAD, 2.59 +/- 0.81 [P < 0.01]; control subjects, 3.69 +/- 1.09 [P < 0.01]). Multivariate stepwise regression analysis showed that, among the factors considered, glycemic control was independently related to the MFR (r = 0.838; P < 0.05). CONCLUSION: Glycemic control appears to be essential for coronary microangiopathy in NIDDM. PMID- 10855622 TI - How accurate is dimercaptosuccinic acid scintigraphy for the diagnosis of acute pyelonephritis? A meta-analysis of experimental studies. AB - The purpose of this study was to evaluate the performance of dimercaptosuccinic acid (DMSA) scintigraphy in the diagnosis of acute pyelonephritis and to compare the test performance of the standard technique, planar DMSA, with the newly introduced technique, SPECT DMSA. METHODS: All published animal studies in which DMSA scintigraphy was compared with histopathology, the reference standard for acute pyelonephritis, were identified using a comprehensive search strategy with the MEDLINE and EMBASE databases. Test performances of all DMSA methods and SPECT versus planar DMSA were analyzed using summary receiver operating characteristic (sROC) curves. RESULTS: Seven studies were identified, including 2 of SPECT DMSA. Problems in study design or reporting were common, with numerical errors in 4 studies. Overall, at a sensitivity of 86%, specificity was estimated to be 91%. Detection of acute pyelonephritis was at a lower threshold for SPECT than for planar DMSA (sensitivity/specificity values of 97%0/66% compared with 82%/ 97%), and the overall test performance of SPECT was not demonstrably better than that of planar DMSA. When applied to a group of children with a prevalence of renal damage of 40%, this means that 98% of children with abnormal planar DMSA scans will have renal damage, whereas only 65% of those with abnormal SPECT scans will have renal damage. Planar and SPECT DMSA will miss 11% and 3% of children with renal damage, respectively. Out of 100 children in the hypothetical group with 40% experiencing renal damage, SPECT will identify 6 extra true cases of renal damage at the expense of 19 extra false positives, when compared with planar DMSA. CONCLUSION: Published studies of DMSA test performance are few in number and have significant methodologic problems that should be avoided in future studies. DMSA, particularly the planar technique, performs well for the diagnosis of acute pyelonephritis. Using test performance criteria, SPECT DMSA alone has not been shown to be preferable to the established planar method and will result in a small number of true-positives at the expense of a larger number of false positives. PMID- 10855623 TI - Decreased dopamine transporter binding in Machado-Joseph disease. AB - The aim of this study was to use 99mTc-TRODAT-1 brain SPECT for investigation of the binding of dopamine transporter (DAT) in the nigrostriatal dopaminergic pathway of symptomatic Machado-Joseph disease (MJD) and to compare the results with the abnormal cytidylate, adenylate, and guanylate (CAG) expansion in the MJD1 gene and other clinical factors. METHODS: Ten symptomatic MJD patients (8 women, 2 men; age range, 20-71 y; mean age +/- SD, 36.4 +/- 10.6 y; mean duration of illness, 9.8 +/- 5.4 y) and 21 healthy volunteers (age range, 24-71 y; mean age, 47.6 +/- 20.1 y) were examined. Brain SPECT images were acquired 4 h after injection. The ratio of specific to nonspecific nigrostriatal 99mTc-TRODAT-1 binding was measured and compared with the clinical symptoms, duration of illness, and size of abnormal expanded CAG repeats. RESULTS: All nigrostriatal 99mTc-TRODAT-1 ratios were significantly lower in MJD patients than in healthy volunteers (P < 0.05). Discriminant function analysis of all MJD patients showed that the decreased binding of 99mTc-TRODAT-1 in the putamen was not significantly different from that in the caudate nucleus. Eight of 10 MJD patients had significantly decreased 99mTc-TRODAT-1 uptake. Of these 8, 2 had extrapyramidal signs and 6 had no obvious extrapyramidal signs. The other 2 patients, who had normal 99mTc-TRODAT-1 uptake, had no obvious extrapyramidal signs. CONCLUSION: Our findings indicate that 99mTc-TRODAT-1 brain SPECT is an appropriate method for evaluating damage to the nigrostriatal DAT in symptomatic MJD patients with and without extrapyramidal signs. The decreased binding of 99mTc-TRODAT-1 in the nigrostriatal dopaminergic pathway in symptomatic MJD patients correlates with the phenotype of extrapyramidal signs but not with the abnormal CAG repeat length, age at disease onset, or disease duration. PMID- 10855624 TI - Metabolic response of non-Hodgkin's lymphoma to 131I-anti-B1 radioimmunotherapy: evaluation with FDG PET. AB - 131I-anti-B1 (CD20) radioimmunotherapy (RIT) is a promising approach for treatment of non-Hodgkin's lymphoma (NHL). We assessed the tumor metabolic response to RIT using FDG PET. METHODS: We examined 14 patients with NHL, who were given first a tracer dose of 131I-anti-B1 and then RIT, each preceded by infusion of unlabeled anti-B1. In 8 of 14 patients, PET was performed at baseline and 33-70 d after RIT. The other 6 patients underwent PET at baseline, 6-7 d after the tracer dose, and 5-7 d after RIT to estimate the early response to tracer dose and RIT. To assess tumor FDG uptake, standardized uptake value normalized for lean body mass (SUV-lean) was measured 1 h after FDG injection. RESULTS: After RIT, complete response was observed in 6 patients, partial response in 6, and no response in 2. At 33-70 d after RIT, mean SUV-lean of 6 responders markedly declined to 41% of the baseline value (P < 0.002). Soon after tracer dose and after RIT, mean SUV-lean of the other 6 responders decreased to 79% and 62% of the baseline values, respectively (P < 0.05). In 2 nonresponders, SUV-lean did not significantly decline from the baseline value at 37 d after RIT. CONCLUSION: FDG PET metabolic data obtained 1-2 mo after RIT correlate well with the ultimate best response of NHL to RIT, more significantly than the early data after tracer dose or RIT. FDG uptake in NHL may decline gradually after RIT in responding patients. PMID- 10855625 TI - Thyroid cancer prevalence after radioiodine treatment of hyperthyroidism. AB - The definitive treatment of hyperthyroidism in Europe is quite different from that in the United States. In Europe, the surgical approach is often preferred and considered safer than radioiodine treatment. European doctors usually prefer to surgically remove the thyroid and perform a pathologic examination of it. They consider it to be an essential diagnostic tool to identify possible diseases that might be associated with hyperthyroidism and even to detect the rare thyroid tumors that might be associated with thyroid hyperfunction. The aim of this study was to evaluate whether radioiodine therapy could be a risk factor for the misdiagnosis of thyroid cancer. METHODS: We performed a retrospective revision of data we collected from 6647 patients (1171 [17.5%] men, 5476 [82.5%] women), all of whom underwent 1311 therapy for hyperthyroidism from 1970 to 1997. Of the whole group, 6.5% were younger than 40 y, 33.5% were 40-60 y old, and 60% were older than 60 y. Moreover, 5061 (76%) patients had either an autonomously functioning node or a toxic multinodular goiter. The other 1586 (24%) patients had Graves' disease. RESULTS: After treatment, thyroid cancer was discovered in 10 (0.15%) patients, none of whom belonged to the group of patients with Graves' disease. Five of these patients were treated during a period from 1970 to 1980, when sonography was not routinely available. The incidence of thyroid cancer in the series of radioiodine-treated patients (150/100,000 over a 27-y period) was not significantly different from its incidence in the general population. The expected rate is 124.88 per 100,000 over a 27-y period. CONCLUSION: An accurate preliminary evaluation (clinical examination, sonography, and cytologic evaluation of fine-needle aspiration) is fundamental for a proper choice between radioiodine and surgical therapy. PMID- 10855626 TI - FDG PET of recurrent or metastatic 131I-negative papillary thyroid carcinoma. AB - This study reports on the use of FDG PET in the follow-up of papillary thyroid cancer patients with negative findings on 131I total body scans and elevated levels of thyroglobulin after total thyroidectomy. METHODS: Eleven asymptomatic patients with previous papillary thyroid cancer, total thyroidectomy, 131I ablation, and treatment of all known metastases had negative findings on 131I total body scans after therapy but persisting elevations of thyroglobulin when not receiving thyroid hormone. All imaging before PET failed to show persisting tumor. FDG PET was performed on all patients while receiving full thyroid hormone replacement, except for the repeated scan of 1 patient (patient 6). After the PET scan, all patients were referred for supplementary CT, sonography, or biopsy of lesions in the neck. RESULTS: All 11 patients showed FDG uptake in the neck or upper mediastinum-in the initial scan in 10 and in a repeated scan in 1. Sonographically guided biopsy confirmed malignancy in 6, was nondiagnostic in 2, and showed normal findings in 1. In 2 patients, the sonographic results were normal and no biopsy was attempted. FDG imaging redirected the treatment of 7 patients, resulting in surgery and external beam radiotherapy in 3, surgery in 1, and external beam radiotherapy in 2. One patient declined further recommended surgery. The other 4 patients remain under observation. Surgical histopathology confirmed thyroid tumor in all 4 surgically treated patients. Retrospective review of the original histopathology slides showed no preponderance of aggressive histology. CONCLUSION: FDG PET is able to guide further evaluation of thyroid cancer patients who have elevated thyroglobulin levels and normal findings on 131I whole-body scanning. PMID- 10855627 TI - Safety and efficacy of arcitumomab imaging in colorectal cancer after repeated administration. AB - In pivotal phase III clinical trials for detecting recurrent or metastatic colorectal cancer, most patients received a single arcitumomab injection. However, the early detection of postsurgical recurrence or metastases with arcitumomab will necessitate serial studies for surveillance. We present immunogenicity, safety, and imaging data supporting the use of multiple administrations of arcitumomab. METHODS: Human antimouse antibody (HAMA) response, adverse events, clinical laboratory values, and diagnostic imaging results were evaluated in 44 patients (24 men, 20 women; age range, 2878 y) after repeated arcitumomab administration (44 second and 3 third injections). Most patients initially had Dukes' class B or C colorectal cancer and had known or occult disease recurrence and elevated serum carcinoembryonic antigen levels at the time of the repeated injection. RESULTS: At the repeated injection, in no patient did elevated HAMA titers develop, hematology and serum chemistry changes were clinically insignificant, and only 1 adverse event (eosinophilia) was judged at least possibly related to arcitumomab. Arcitumomab imaging results at the second injection were comparable with those obtained in phase III trials after a single injection of arcitumomab, having a 78% per-lesion concordance with CT in the abdomen and pelvis and a 73% sensitivity and 94% specificity based on 9 patients with cancer confirmed surgically at 11 anatomic sites and excluded at 16 sites. CONCLUSION: These data indicate that at least 2 injections of arcitumomab can be given safely to patients with colorectal cancer, without increased immunogenicity and with imaging efficacy equivalent to the first administration. PMID- 10855628 TI - Decreased serum E-selectin concentration after 89Sr-chloride therapy for metastatic prostate cancer bone pain. AB - Palliative systemic radionuclide therapy with 89Sr-chloride is a useful intervention for patients with bone pain from metastatic prostatic cancer. Although this radionuclide is highly effective, its mechanism of action remains unresolved. This investigation sought to determine whether systemic radionuclide therapy decreases the production of cell adhesion molecules (E-selectins) that participate in the metastatic process. METHODS: Sera were collected from 25 men with metastatic (stage IV) prostate carcinoma who received 89Sr-chloride palliative therapy and from 10 age-matched healthy volunteers. The serum concentration of E-selectin was quantified by an enzyme-linked immunosorbent assay. Sera from 5 patients who received 2 courses of radionuclide therapy were also included in the analysis. RESULTS: A 2.8-fold decrease in serum E-selectin concentration occurred within 2 mo of radionuclide therapy (P < 0.0001). At 10 mo, however, the concentration increased to a mean (+/- SD) of 151.2 +/- 51.3 ng/mL, surpassing the baseline concentration. This pattern coincided with symptomatic improvement and subsequent health status deterioration. For patients who received 2 courses of radionuclide therapy, a second fall in serum E-selectin concentration followed the second radionuclide treatment. CONCLUSION: A significant decrease in serum E-selectin concentration was observed after systemic radionuclide therapy. This finding suggests that expression of cell adhesion molecules, an important determinant of metastatic progression, may be inhibited by 89Sr-chloride. PMID- 10855629 TI - Kidney function after radical nephrectomy: assessment by quantitative SPECT of 99mTc-DMSA uptake by the kidneys. AB - To determine the function of the remaining contralateral kidney after the removal of a functioning kidney, 30 consecutive patients (18 men, 12 women; average age, 67 y; age range, 34-87 y) who were undergoing unilateral radical nephrectomy were evaluated by sequential quantitative 99mTc-dimercaptosuccinic acid (DMSA) SPECT (QDMSA) studies. METHODS: The 30 patients were undergoing radical nephrectomy for renal tumors. The first study was done before surgery. Follow-up studies were performed 2-23 mo after surgery. Clinical evaluations and determinations of serum creatinine level were performed at the same time as the QDMSA studies. RESULTS: The relative contribution of the resected kidneys to the global renal function before surgery was 43.2% +/- 7.3%. After surgery the uptake of the remaining kidney increased from 13.4% +/- 4.0% to 18.3% +/- 5.8% (t = 5.7; P = 0.0000). The relative function of the remaining kidney increased from 56.8% +/- 7.1% to 79.1% +/- 23.6% (t = 4.9; P < 0.0001) of the global renal function before nephrectomy. Increases in the renal volume (from 211 +/- 62 cm3 to 229 +/- 68 cm3; t = 4.5; P = 0.0001) and in the percentage injected dose per cubic centimeter (%ID/cm3) of the remaining kidney (from 0.066 +/- 0.02 % ID/cm3 to 0.085 +/- 0.03 %ID/cm3; t = 4.6; P = 0.0001) were associated with this change. Nine patients had 2 follow-up studies performed 3-4 mo after surgery and 12-14 mo after surgery. The volume of the remaining kidney (209.22 +/- 46.20 cm3 versus 217.88 +/- 58.85 cm3; t = 0.962; P = 0.364), the %ID/cm3 (0.09 +/- 0.016 %ID/cm3 versus 0.093 +/- 0.025 %ID/cm3; t = 0.362; P = 0.726), and the percentage uptake (19.26% +/- 4.45% versus 20.11% +/- 7.01%) did not change significantly between these 2 QDMSA studies. CONCLUSION: The results of this study suggest that adaptive changes causing hyperfunction of the remaining kidney may occur after nephrectomy of a functioning kidney in adults. These changes occur soon after surgery, persist for at least 1 y, and are evident on QDMSA studies. PMID- 10855630 TI - Diuretic renography with the addition of quantitative gravity-assisted drainage in infants and children. AB - The aim of this study was to evaluate the use of quantitative gravity-assisted drainage (GAD) using >50% residual activity as an indicator to confirm obstruction in diuretic renography in the investigation of hydronephrosis and hydroureteronephrosis in infants and children. This was evaluated in 2 groups: furosemide clearance half-time (t 1/2) > 20 min (obstructed range) and t 1/2 = 10 20 min (indeterminate range). METHODS: Two hundred children (155 boys, 45 girls; age range, 2 d to 16 y; median age, 26 wk) were studied over a 2-y period. One hundred thirty-five F+20 (diuretic given 20 min after radiopharmaceutical) and 65 F+0 (simultaneous administration of diuretic and radiopharmaceutical) studies were performed with intravenous administration of 99mTc-mercaptoacetyltriglycine (MAG3) and furosemide. At the end of the 20-min diuretic phase, a 5-min post-GAD image was obtained, and the percentage of residual activity was calculated by comparison with the last 5 min of the diuretic phase. All patients were monitored for 6-12 mo, and the final diagnoses were based on either surgical findings or conservative management with follow-up sonography or 99mTc-MAG3 studies. Results of the diuretic renography using quantitative GAD were then compared with the final diagnoses. RESULTS: A renal unit was defined as a kidney and its ureter. In the 200 patients studied, 256 hydronephrotic renal units were analyzed: 10 units showed no function, 1 unit showed poor function, 131 units had t 1/2 < 10 min, 62 units had t 1/2 > 20 min, and 52 units had t 1/2 = 10-20 min. Of the 131 renal units with t 1/2 < 10 min, there was only 1 case of obstruction. Using GAD > 50% residual activity for the diagnosis of obstruction in 62 renal units with t 1/2 > 20 min, the sensitivity was 88.4%, the specificity was 73.7%, and the accuracy was 83.9%. Similarly, using GAD > 50% residual activity for the diagnosis of obstruction in 52 units with t 1/2 = 10-20 min, the sensitivity was 100%, the specificity was 79.5%, and the accuracy was 82.7%. CONCLUSION: The quantitation of GAD > 50% residual activity in diuretic renography can help to differentiate between obstruction and nonobstruction in renal units with t 1/2 > 20 min and t 1/2 = 10-20 min. The quantitation of GAD when t 1/2 < 10 min is not useful because obstruction has already been excluded. PMID- 10855631 TI - Diuretic MAG3 scintirenography in children with HIV nephropathy: diffuse parenchymal dysfunction. AB - HIV nephropathy (HIVN) is prevalent in 15%-56% of HIV-infected children and induces mild to severe progressive nephropathy. METHODS: A total of 33 renal diuretic scintirenographic studies with 99mTc-mercaptoacetyltriglycine (MAG3) were reviewed and analyzed from 23 HIV pediatric patients, 21 of whom had HIVN with varying degrees of renal impairment. Results were compared with 10 studies of control patients of matching ages. Visual interpretation of images and renograms as well as semiquantitative analyses were performed. Variables compared were size of kidneys, time of peak and one-half peak activities, residual (or retained) cortical activity at 20 min, ratio of cortical activity at 2.5-20 min, and ratio of kidney activity to kidney plus background activity at 2 min. The results of MAG3 renal studies were also compared with laboratory data pertaining to creatinine clearance in all patients and with sonography in 17 patients. RESULTS: In most patients with HIVN (18/21), the kidneys were larger than normal, with a diffuse parenchymal dysfunction (decreased uptake, slow processing, and increased retention of activity) and flat renograms, findings similar to those observed in other diffuse parenchymal diseases. In all patients with HIVN, semiquantitative analysis (paired t test) showed statistically significant differences from control patients for all variables. On ANOVA, a statistically significant correlation was found between most scintigraphic parameters and the severity of renal impairment. Of the 17 concurrent sonographic studies in HIVN patients, 7 showed no abnormalities, whereas the results of scintigraphy were abnormal. CONCLUSION: Diuretic MAG3 scintirenography shows nonspecific diffuse parenchymal dysfunction in pediatric patients with HIVN. Such dysfunction may provide corroborative evidence of HIVN and should be recognized when the test is performed for standard indications. Further work is necessary to prove that the test has indeed the high sensitivity and good correlation with the seventy of HIVN suggested in this population; the test may be useful to follow up the progression of disease and the effect of treatment. PMID- 10855632 TI - Incomplete recovery of lung perfusion after 3 months in patients with acute pulmonary embolism treated with antithrombotic agents. THESEE Study Group. Tinzaparin ou Heparin Standard: Evaluation dans l'Embolie Pulmonaire Study. AB - We assessed the time course of lung perfusion after 3 mo of anticoagulant therapy for acute pulmonary embolism (APE) on the basis of perfusion lung scan (PLS) findings for 157 patients included in the Tinzaparin ou Heparin Standard: Evaluation dans l'Embolie Pulmonaire Study (THESEE), a multicenter, randomized, nonmasked trial comparing standard, continuous, adjusted-dose intravenous heparin with once-daily, subcutaneous, low-molecular-weight heparin in patients with APE. METHODS: We calculated the percentage-of-vascular-obstruction score (PVOs) on PLSs on the day of diagnosis of APE (PVOsD1), on day 8 (PVOsD8), and after 3 mo (PVOsM3) and the mean relative changes in PVOs on day 8 versus the day of diagnosis and after 3 mo versus the day of diagnosis. RESULTS: Mean PVOsD1 +/- SD was 49% +/- 20%, PVOsD8 was 29% +/- 18%, and PVOsM3 was 19% +/- 18%. PVOsD1 was at least 50% in 49% of patients. Reperfusion did not correlate with age, importance of initial obstruction, or clinical severity of disease at inclusion in THESEE. Relative change after 3 mo versus at diagnosis was lower in the 87 patients with associated prior cardiopulmonary disease than in those without. In the 43 patients with a history of thromboembolic disease, neither mean PVOsD1 nor the time course of PVOs was different from those in patients without a history of thromboembolic disease. Residual defects after 3 mo were observed in 104 patients (66%), including 13 with a PVOs of at least 50%. CONCLUSION: These results emphasize the need for a control PLS at completion of anticoagulant therapy for APE, even in patients with full resolution of symptoms. PMID- 10855633 TI - The chronic perfusion defect: our knowledge is still hazy, but the message is clear. PMID- 10855634 TI - Scintillation crystals for PET. AB - In PET, inorganic scintillator crystals are used to record gamma-rays produced by the annihilation of positrons emitted by injected tracers. The ultimate performance of the camera is strongly tied to both the physical and scintillation properties of the crystals. For this reason, researchers have investigated virtually all known scintillator crystals for possible use in PET. Despite this massive research effort, only a few different scintillators have been found that have a suitable combination of characteristics, and only 2 (thallium-doped sodium iodide and bismuth germanate) have found widespread use. A recently developed scintillator crystal, cerium-doped lutetium oxyorthosilicate, appears to surpass all previously used materials in most respects and promises to be the basis for the next generation of PET cameras. PMID- 10855635 TI - Imaging pulmonary emboli and deep venous thrombi with 99mTc-bitistatin, a platelet-binding polypeptide from viper venom. AB - An imaging test that could locate both pulmonary emboli (PE) and their source, active deep venous thrombi (DVT), would be valuable in patient management. Bitistatin, an 83-amino-acid polypeptide isolated from Bitis arietans venom, binds avidly to the glycoprotein IIb/IIIa receptor on platelets. The goal of this study was to label bitistatin with 99mTc and assess its potential for imaging thrombi and emboli in vivo. METHODS: Molecular modeling of bitistatin indicated that its primary amines are located on the opposite side of the molecule from the receptor-binding domain. The primary amines were reacted with succinimidyl-4 hydrazino nicotinate hydrochloride to place 2.4 hydrazino nicotinate (Hn) chelating groups per peptide molecule. Hn-bitistatin was labeled by incubation with 99mTc-glucoheptonate to 96 TBq/mmol and then tested for binding to platelets in vitro and for imaging of 24-h-old DVT and PE in a canine model used previously for other thrombus tracers. RESULTS: 99mTc-Hn-bitistatin bound to stimulated platelets with a dissociation constant (Kd) = 32 nmol/L, similar to that of 125I bitistatin (Kd = 41 nmol/L). In vivo, focal uptake was observed in planar images as early as 30 min (DVT) and 60 min (PE) after injection. Lesion uptake of 99mTc Hn-bitistatin at 4 h after injection was calculated in terms of percentage injected dose per gram (%ID/g) of tissue and averaged 0.89 %ID/g PE and 0.79 %ID/g DVT. Lesion-to-background ratios averaged 34:1 (PE-to-lung), 18:1 (DVT-to blood), and 284:1 (DVT-to-muscle). These values were not significantly different from iodinated bitistatin, but uptakes were higher than other tracers tested in the same model. CONCLUSION: 99mTc-Hn-bitistatin retains the functional activity of the iodinated peptide, has higher DVT and PE uptakes than other thrombus tracers in this standardized model, and has target-to-background characteristics suitable for imaging both PE and DVT in a single test. PMID- 10855636 TI - Use of radiolabeled peptides to image deep venous thrombosis and pulmonary embolism. PMID- 10855637 TI - PET with a dual-head coincidence camera: spatial resolution, scatter fraction, and sensitivity. AB - Scintillation cameras with options for detecting positron annihilation quanta in the coincidence acquisition mode may be the most innovative diagnostic devices introduced in nuclear medicine during the last few years. Besides conventional low-energy imaging in the collimated single-photon mode, these options offer a relatively inexpensive opportunity to perform uncollimated PET by switching into the coincidence acquisition mode. Instead of collimators, scatter frames (with 2 optional configurations: axial or open scatter frame) can be mounted to reduce the amount of quanta reaching the detectors from parts of the patient's body outside the field of view. This study investigates the coincidence imaging properties of the scintillation camera by measuring spatial resolution, scatter fraction, sensitivity, and count-rate response for 18F. METHODS: A needle in air and a plastic tube in water, each filled with 18F, were oriented axially and transversally to measure the transverse and axial spatial resolutions, respectively. Using either the axial or the open scatter frame, a standard cylinder filled homogeneously with activity was studied over several half-life periods to deduce the respective scatter and random fractions of the system by means of a sinogram technique. The activity of the cylinder was kept low to determine the sensitivity to coincidence events for both scatter frames. RESULTS: Depending on the distance between the line source and the axis of rotation and on the choice of the axial acceptance angle used to reconstruct the coincidence events (single-slice rebinning algorithm), the axial resolution was found to be between 6 and 10 mm (full width at half maximum) with the axial scatter frame mounted. The transversal resolution was 6-6.5 mm on the axis of rotation, independent of the scatter frame used. The scatter fraction amounted to roughly 25% for the axial and 38% for the open scatter frames. The sensitivity when measuring true coincidence pairs ran to nearly 650 Hz/kBq/mL, when acquisition was performed with the axial scatter frame using a 30%-wide photopeak energy window. When acquiring with the open scatter frame, the sensitivity increased to nearly 3000 Hz/kBq/mL. Using the axial scatter frame, the homogeneously filled cylinder could be scanned with a maximum true coincidence rate of 2000 Hz for an activity of 55-60 MBq. Although this maximum true coincidence counting rate did not change significantly when the acquisition was performed with the open scatter frame, the respective activity in the standard cylinder was decreased to 10-15 MBq. CONCLUSION: The spatial resolution of the scintillation camera is sufficient for high-resolution coincidence imaging. Compared with a dedicated PET scanner, the scatter fraction is relatively high and should therefore be corrected adequately. The relatively low sensitivity and the rather low maximum true coincidence counting rate are considerably inferior compared with a conventional PET scanner. However, these drawbacks can be partially compensated for, facilitating its clinical use. PMID- 10855638 TI - Spatial and temporal registration of CT and SPECT images: development and validation of a technique for in vivo three-dimensional semiquantitative analysis of bone. AB - The combined use of postoperative 3-dimensional CT and SPECT imaging provides a means of relating anatomy and physiology for the semiquantitative in vivo analysis of bone. This study focuses on the development and validation of a technique that accomplishes this through the registration of SPECT data to a 3 dimensional volume of interest (VOI) interactively defined on CT images. METHODS: Five human cadaver heads served as anthropomorphic models for all experiments. Four cranial defects were created in each specimen with inlay and onlay split skull bone grafts reconstructed to skull and malar recipient sites. To acquire all images, each specimen was landmarked with 1.6-mm ball bearings and CT scanned. Bone surfaces were coated with 99mTc-doped paint. The locations of the ball bearings were marked with paint doped with 111In. Separate SPECT scans were acquired using the energy windows of 99mTc and 111In. RESULTS: Serial SPECT images aligned with an average root-mean-square (RMS) error of 3.8 mm (i.e., <1 pixel). CT-to-SPECT volume matching aligned with an RMS error of 7.8 mm. Total counts in CT-defined VOIs applied to SPECT data showed a strong linear correlation (r2 = 0.86) with true counts obtained from a dose calibrator. CONCLUSION: The capability of this multimodality registration technique to anatomically localize and quantify radiotracer uptake is sufficiently accurate to warrant further assessment in an in vivo trial. PMID- 10855639 TI - Radioiodine uptake in thyroid remnants during therapy after tracer dosimetry. AB - Our objective was to evaluate the effect of a diagnostic tracer dose of 131I on the uptake of the therapeutic dose of 1311 in the ablation of a thyroid remnant or residual tumor in patients with differentiated thyroid cancer. METHODS: Twelve consecutive patients referred for a dosimetric study and subsequent radioiodine treatment of focal neck uptake of 131I were studied. The 24-h (in 1 case, 48-h) neck activity was calculated by the region-of-interest method, after both dosimetric and therapeutic administrations. The focal activity in the neck was corrected for decay and compared with the total activity administered to obtain the percentage uptake at 24 h. This procedure was performed for both the scanning dose (range, 19.8-196.1 MBq; mean, 85.1 MBq; median, 40 MBq) and the therapeutic dose (range, 1.073-5.713 GBq; mean, 2.991 GBq). The uptake of the therapeutic dose was then expressed as a percentage of the uptake of the diagnostic dose (%T/D). Counting rate linearity was established up to 350 MBq in the field of view of the gamma camera used in the study. RESULTS: Thirteen of a total of 16 lesions exhibited reduced uptake from the therapeutic dose, 2 remained the same, and in 1 the uptake actually increased from 0.26% to 1.01%. The %T/D ranged from 7.0% to 388.5%, with a mean of 71%. If the lesion with increased uptake is excluded, the range becomes 7.0%-102.1%, with a mean of 50%. Linear regression between the percentage uptake of the diagnostic dose to that of the therapeutic dose results in a slope of 0.42, with a correlation coefficient of only 0.75. We were unable to accurately calculate the radiation dose to the lesion from the diagnostic activity of 131I, because of uncertainty about the tumor mass. CONCLUSION: The percentage uptake of the therapeutic dose is on average only one half of that predicted from the dosimetric uptake in thyroid remnants after surgery, even though the median dosimetric dose was only 40 MBq. This reduced uptake should be accounted for in the therapeutic prescription for thyroid ablation or treatment of residual thyroid cancer. We postulate that this effect is caused by radiation damage from the tracer dose during dosimetry. PMID- 10855640 TI - Comparison of 4 methods for quantification of dopamine transporters by SPECT with [123I]IACFT. AB - 2Beta-carbomethoxy-3beta-(4-fluorophenyl)-n-(1-iodoprop-1-en -3-yl) nortropane (IACFT) is a highly selective ligand for dopamine transporter (DAT) sites in the striatum. Recent reports have described the basic kinetics, neurobiology, and imaging properties of [123I]IACFT. This report focuses on the structural (i.e., the ability to produce consistent binding estimates) validity of 4 methods to quantify striatal binding potential (BP) for IACFT. METHODS: Seven healthy volunteers and 8 patients with Parkinson's disease were subjects for this study. Dynamic SPECT images and arterial blood samples were acquired during the 1.5-2 h after injection of 185-370 MBq [123I]IACFT. Plasma radioactivity was analyzed chromatographically to obtain metabolite-corrected arterial input functions. The k3/k4 ratio (BP) for striatal DAT sites was calculated by 4 methods. In the first method, tissue time-activity curves and metabolite-corrected arterial input functions were analyzed by a linear graphic method developed for reversible receptor ligands. The second method was also graphic; however, the occipital cortex time-activity curve was used as the input function. In the third method, the difference between the striatal and occipital cortex time-activity curves at secular equilibrium was taken to represent bound tracer, the occipital cortex time-activity curve was used to represent tracer in the free and nonspecifically bound state, and equilibrium receptor equations were used to determine BP. The fourth method used the occipital cortex time-activity curve to mathematically derive an input function for fitting the striatal time-activity curve and to determine BP. RESULTS: Analysis of the dynamic SPECT data by methods 1 and 2 resulted in highly linear plots (after approximately 15 min), supporting the reversibility of the tracer. A high linear correlation was found for BP determined by all 4 methods. ANOVA showed that methods 1-3 were indistinguishable; method 4 yielded lower BPs than did methods 1-3. CONCLUSION: These results show that BP can be estimated consistently using 4 different methods. This finding lends support to the modeling assumptions and provides methods suitable for clinical investigation. PMID- 10855641 TI - DNA cleavage by 111In-labeled oligodeoxyribonucleotides. AB - We studied the fine structure of DNA damage produced by the decay of 111In incorporated into duplex and triplex DNA strands to evaluate the usefulness of this radionuclide for sequence-specific DNA cleavage. METHODS: Oligodeoxyribonucleotides (ODNs) were prepared with 111In attached by diethylenetriaminepentaacetic acid (DTPA) at the 5' end or 3' end through a long chemical linker or to an internal nucleotide position through a short linker. Subsequent formation of DNA duplexes and triplexes was confirmed by gel electrophoresis. The 111In-induced breaks were assayed in denaturing polyacrylamide gel electrophoresis with a single-nucleotide resolution. RESULTS: 111In-labeled oligonucleotides of high specific activity (740-1554 TBq/mmol) were synthesized. The presence of the bulky 111In-DTPA group did not impede duplex or triplex formation. Localized DNA breaks were observed in all duplexes and triplexes formed. The majority of DNA breaks in duplex formations were located within +/- 10 nucleotides from the site of attachment of the 111In-bearing linker. The yield of DNA breaks per decay was 0.38 in a duplex with internally modified ODNs. This is nearly 2 times less than the yield of DNA breaks in the same duplex with 1251 attached through the same linker. The yield of DNA breaks in the pyrimidine and purine strands of DNA triplexes with 111In attached to the triplex-forming ODNs through the linkers of different length varied from 0.05 to 0.10. The distribution of DNA breaks was wider in comparison with the duplex experiment. The lower yields of breaks per 111In decay compared with 125I may be not only the result of lower deposited energy but also of the ionic repulsion of the negatively charged 111In-DTPA group from the DNA strands. CONCLUSION: We have shown that decay of 111In produces highly localized DNA breaks. 111In introduced into triplex- and duplex-forming ODNs through hydrocarbon linkers produces sequence-specific DNA strand breaks with an efficiency nearly comparable with that of 1251. These findings are supportive of our proposed use of 111In-ODNs for gene-specific radiotherapy. PMID- 10855642 TI - Kinetics of a putative hypoxic tracer, 99mTc-HL91, in normoxic, hypoxic, ischemic, and stunned myocardium. AB - 99mTc-4,9-diaza-3,3,10,10-tetramethyldodecan-2,11-dione dioxime (HL91) was developed as a putative hypoxic reagent. This study focused on the myocardial kinetics of 99mTc-HL91 in various oxygen levels and perfusion states. METHODS: The time-activity curve of 99mTc-HL91 was measured in isolated perfused rat heart after the bolus infusion. RESULTS: 99mTc-HL91 was cleared quickly from normoxic hearts, and retention at 30 min after injection was 0.18 +/- 0.02 percentage injected dose per gram of wet weight (mean +/- SE; n = 6). When the concentration of oxygen bubbling through the perfusate was reduced from 100% to 50%, 20%, 5%, and 0%, retention of 99mTc-HL91 increased to 0.47 +/- 0.03 (n = 5), 0.48 +/- 0.03 (n = 5), 0.71 +/- 0.01 (n = 5), and 0.70 +/- 0.02 (n = 5), respectively (P < 0.05). Compartment analysis revealed that the trapping mechanism, which was dependent on tissue oxygen concentration, determined the retention rate. Although not retained in stunned myocardium (0.17 +/- 0.02, n = 5; P = not significant), 99mTc-HL91 was significantly retained when injected before ischemia (1.06 +/- 0.06, n = 5; P < 0.05). CONCLUSION: These results indicate that retention of 99mTc-HL91 correlates well with oxygen level in the perfusate, suggesting that the agent may be a useful marker of the severity of myocardial hypoxia. PMID- 10855643 TI - Interferon-alpha-2b immunoconjugate for improving immunoscintigraphy and immunotherapy. AB - A pretreatment with a single dose of an immunoconjugate (IC) that promises to enhance tumor uptake and decrease liver uptake of radiolabeled monoclonal antibodies (MAbs) might be of use in radioimmunodetection and radioimmunotherapy (RIT). We have shown previously that an interferon (IFN)-MAb (1:1) immunoconjugate (IC) enhances tumor uptake by a factor of 2 or more and reduces liver uptake by 50% in nude mice bearing human tumors. The aim of this study was to determine whether IFN modulates antigenic expression and to ascertain the most effective route of its administration, the optimal quantity to be administered, and the optimal duration of time to lapse between the administration of IC and the radiolabeled MAb. METHODS: IFN-alpha-2b and anticarcinoembryonic antigen-F6 (IgG2a) MAb were conjugated (1:1), and F(ab')2 of the MAb was labeled with 99mTc. Human colorectal tumors were grown in nude mice by implanting 5 x 10(6) LS174T confluent cells grown in culture. Mice, 5 in each group, received 20 x 10(3) IU intravenously, intramuscularly, or intraperitoneally and 40 x 10(3), 60 x 10(3), and 80 x 10(3) IU intravenously 30 min before the intravenous administration of 25.9 MBq 99mTc/20 microg F(ab')2. Mice in the control groups received 99mTc F(ab')2 but not the conjugate. Twenty-four hours later mice were killed and imaged, and tissues were removed for quantitative (percentage injected dose/g [% ID/g]) distribution of 99mTc. RESULTS: In all conjugate-receiving mice, the tumor uptake was higher and the liver uptake was lower (P < 0.01) than that in the control mice with the exception of liver uptake, which was not significantly different in mice receiving 80 x 10(3) IU conjugate. The optimal results were apparent in mice pretreated with 40 x 10(3) IU conjugate in which tumor uptake was enhanced by a factor of 2.3 (4.8 +/- 0.5 %ID/g versus 11 +/- 0.7 %ID/g; P < 0.01). The renal uptake remained unchanged, and the tumor-to-muscle ratios increased from 11.5 +/- 6.8 to 14.6 +/- 3.9, and the tumor-to-blood ratios increased from 4.4 +/- 1.8 to 8.3 +/- 2.4. The liver uptake decreased from 9.5% +/- 1% to 5% +/- 1.6%. Results were attributed to enhanced tumor blood flow, increased antigenic expression, and blocking of hepatic nonspecific Fc receptors. CONCLUSION: A pretreatment with IFN-MAb conjugate is a worthwhile approach to consider in radioimmunoscintigraphy and RIT. PMID- 10855644 TI - 99mTc-HYNIC-[Tyr3]-octreotide for imaging somatostatin-receptor-positive tumors: preclinical evaluation and comparison with 111In-octreotide. AB - In this paper we describe the preclinical evaluation of 99mTc-hydrazinonicotinyl Tyr3-octreotide (HYNIC-TOC) using different coligands for radiolabeling and a comparison of their in vitro and in vivo properties with 111In diethylenetriaminepentaacetic acid (DTPA)-octreotide. METHODS: HYNIC-TOC was radiolabeled at high specific activities using tricine, ethylenediaminediacetic acid (EDDA), and tricine-nicotinic acid as coligand systems. Receptor binding was tested using AR42J rat pancreatic tumor cell membranes. Internalization and protein binding studies were performed, and biodistribution and tumor uptake were determined in AR42J tumor-bearing nude mice. RESULTS: All 99mTc-labeled HYNIC peptides showed retained somatostatin-receptor binding affinities (Kd < 2.65 nM). Protein binding and internalization rates were dependent on the coligand used. Specific tumor uptake between 5.8 and 9.6 percentage injected dose per gram (%ID/g) was found for the 99mTc-labeled peptides, compared with 4.3 %ID/g for 111In-DTPA-octreotide. Tricine as coligand showed higher activity levels in muscle, blood, and liver, whereas tricine-nicotinic acid produced significant levels of activity in the gastrointestinal tract. EDDA showed the most promising overall biodistribution profile, with tumor-to-liver and tumor-to gastrointestinal tract ratios similar to those obtained with 111In-DTPA octreotide, lower ratios in blood and muscle, but considerably higher tumor-to kidney ratios. CONCLUSION: TOC can be radiolabeled to high specific activities using HYNIC as a bifunctional chelator. The high specific tumor uptake, rapid blood clearance, and predominantly renal excretion make 99mTc-EDDA-HYNIC-TOC a promising candidate for an alternative to 111In-DTPA-octreotide for tumor imaging. PMID- 10855645 TI - 99mTc-tetrofosmin scintigraphy in management of pulmonary tuberculosis. PMID- 10855646 TI - Left ventricular ejection fraction and gated SPECT. PMID- 10855647 TI - Outcome and input. PMID- 10855648 TI - The gross motor function classification system for cerebral palsy: a study of reliability and stability over time. AB - Children with cerebral palsy (CP) experience a change in motor function with age and development. It is important to consider this expected change in offering a prognosis, or in assessing differences in motor function after an intervention. The Gross Motor Function Classification System for CP (GMFCS) has been developed for these purposes. This study was based on a retrospective chart review of 85 children with CP followed from < or =2 to > or =12 years of age. The GMFCS was applied to clinical notes by two blinded raters four times throughout the study. Interrater reliability was high (G=0.93). Test-retest reliability was high (G=0.79). The positive predictive value of the GMFCS at 1 to 2 years of age to predict walking by age 12 years was 0.74. The negative predictive value was 0.90. The GMFCS can validly predict motor function for children with CP. The results are discussed in terms of their implications for clinical practice and future research. PMID- 10855649 TI - Changes in synergistic movement patterns after selective dorsal rhizotomy. AB - The purpose of this investigation was to quantitatively compare synergistic movement patterns between seven children (four male, three female; aged 3 to 17 years; mean 6.7, SD 5.3) without cerebral palsy (CP) (controls) and 27 children (15 male, 12 female; aged 2 to 16 years; mean 5.7, SD 3.7) with spastic diplegic CP before and after selective dorsal rhizotomy (SDR). The study design was also descriptive, comparing results of before and after SDR to control children. A two dimensional video system and retroreflective markers were used to obtain sagittal plane angles for the hip, knee, and ankle during maximum active knee flexion and extension. Correlations were calculated between the knee and hip and between the knee and ankle joint pairs. Control children demonstrated non-synergistic movement patterns (-0.75 and -0.61). These results were significantly different from children with CP (0.40 and 0.43, p<0.05). Eight months after SDR, synergistic patterns did not significantly change from preoperative results (0.23 and 0.36, p>0.05) and remained significantly different from control children (p<0.05). We conclude that it may not be possible to significantly alter synergistic patterns after SDR. PMID- 10855650 TI - Explicit memory in low-risk infants aged 19 months born between 27 and 42 weeks of gestation. AB - The aim of this study was to determine whether there are primary effects of prematurity on the development of explicit memory. Elicited imitation of action sequences was used to compare immediate and 15-minute delayed memory in term and preterm infants (19 months corrected age; n=48) who were at low risk: none had experienced the medical or social risk factors often associated with preterm birth. Relative to infants born at term (38 to 40 weeks' gestation), children who had been born at 27 to 34 weeks' gestation showed lower levels of ordered recall; performance of healthy infants born at 35 to 37 weeks' gestation was intermediate and did not differ significantly from that of the other groups. These results suggest that specific cognitive deficits can occur as a function of preterm birth even in low-risk infants. PMID- 10855651 TI - Prognosis of ischemic stroke in childhood: a long-term follow-up study. AB - Little is known about long-term physical sequelae, cognitive functioning, and quality of life in children who have experienced ischemic stroke. Thirty-seven patients under 16 years of age were studied; the median interval after stroke was 7 years. CT-scans were reassessed to determine the type of infarction at baseline. Occurrences of death, of new cardiovascular events, and of seizures during follow-up were recorded. Surviving patients were invited for a follow-up examination, including physical check-up, global screening of cognition, and an inventory of subjective health perception. Only two patients were lost to follow up. During follow-up four died, nine developed seizures, eight had transient ischemic attacks, and two experienced a recurrent ischemic stroke. None of the patients had cardiac complications during follow-up. In 11 of 27, no functional impairment was found, in 15 there was a hemiparesis of varying severity, and in one a paraplegia. There was a significant shift in cognitive functioning towards lower levels, especially in children with epilepsy. Remedial teaching was frequently needed. Many of the parents' perceived their child's behavior to be very changeable. Three-quarters of the children considered themselves as healthy as other children, and almost all of them as happy. The physical and functional prognosis after ischemic stroke in childhood is relatively good, particularly in children with no serious causative illness, but special education is often needed and social changes occur. PMID- 10855652 TI - Epileptic syndromes, cognitive assessment and school placement: a study of 251 children. AB - Two-hundred and fifty-one children (98 girls and 153 boys, aged from 3 to 17 years) with documented diagnosis of epileptic syndrome, IQ measurement, and information on school placement were included in this retrospective study. The relations between these three parameters as well as effects of age at onset and duration of epilepsy, seizure frequency, and number of antiepileptic drugs (AEDs) were analysed. Both IQ and schooling were univariately related to epileptic syndrome, age at onset and duration of epilepsy, and number of AEDs; seizure frequency was related to IQ but not to school placement. Multiple regression showed that IQ was independently related to epileptic syndrome and AED; multiple logistic regression showed that type of school (mainstream versus adapted or special) was independently related to IQ and AED. Children with idiopathic generalised or with localisation-related epilepsy had higher IQ scores and higher probability of mainstream schooling than those with symptomatic or cryptogenic generalised epilepsies or epileptic syndromes which were undetermined. Subtests profile of intelligence scale in localisation-related epilepsies showed different specific cognitive deficits, according to the location of the epileptic focus. PMID- 10855653 TI - Ten-year follow-up of adolescent-onset anorexia nervosa: physical health and neurodevelopment. AB - To study the development of physical health and some neuromotor functions in anorexia nervosa (AN) 51 individuals (48 females, three males) with a mean AN onset of 14 years, recruited after community screening, were followed prospectively together with 51 age-, sex-, and school-matched individuals without AN (controls). About 10 years after AN onset, all individuals were examined in respect of physical health and neurodevelopment. There were no deaths. Weight and height had normalised, except in three participants with persistent AN. Significantly more participants with AN had a physical complaint/disorder, including hirsutism. This might be a long-term complication in weight restored AN. Dysdiadochokinesis occurred almost exclusively among individuals with former AN in accordance with our previous studies. PMID- 10855654 TI - Body composition in nutritionally adequate ambulatory and non-ambulatory children with cerebral palsy and a healthy reference group. AB - Bone-mineral content (BMC; g) and density (BMD; g/cm2) were measured by dual energy X-ray absorptiometry in the proximal femur, femoral neck, and total body of nutritionally adequate children (n=17; 11 girls, six boys; aged 7.6 to 13.8 years) with spastic cerebral palsy (CP). Bone-mineral-free lean tissue (BMFL; g) and fat mass (FM; g) were obtained from total body scans. Chronological and developmental age-based z scores for the children with CP were derived from a pediatric database (n=894). Children with CP had BMC z scores from -1.8 (total body) to -3.2 (femoral neck) SDs below the normative sample. Non-independent ambulators had lower z scores for total body BMD, femoral neck BMD, and BMC than independent ambulators. The BMFL z score of individuals with CP was 2 SDs below that of the reference group and higher in the independent ambulators than in the non-independent ambulators, whereas FM deviated little. These findings suggest that non-nutritional factors, such as ambulation, account for the low BMC, BMD, and BMFL tissue observed in this population. PMID- 10855655 TI - Multicore myopathy: respiratory failure and paraspinal muscle contractures are important complications. AB - Three ambulant males with multicore myopathy, a rare congenital myopathy, are reported with nocturnal hypoventilation progressing to respiratory failure at the age of 9, 13, and 21 years. Deterioration in these individuals occurred over several months without any precipitating event. Patients had clinical evidence of nocturnal hypoventilation with hypoxaemia and hypercapnia. Forced vital capacity was significantly reduced (20 to 43% of predicted level). These parameters improved on institution of overnight ventilation using a BiPAP pressure support ventilator with face mask or nasal pillows with O2 saturation maintained above 90% overnight and an increase in forced vital capacity by as much as 100% (0.3 to 0.6 litres). This was matched by a symptomatic and functional improvement. Also present in these patients and not previously reported is the association of multicore myopathy with paraspinal contractures which produce a characteristic scoliosis described as a 'side-sliding' spine. This may be improved by spinal bracing or surgery. PMID- 10855656 TI - Recurrent artery of Heubner infarction in infancy. AB - Classically, acquired occlusion of the recurrent artery of Heubner (RAH) results in hemiparesis with faciobrachial predominance. Infarction in the territory of the RAH represents a specific stroke syndrome not yet described in infancy with a range of motor and functional manifestations. An infant is described with apparent congenital infarction of the recurrent artery of Heubner. The child had prominent involvement of the contralateral upper extremity with athetosis. Neuroimaging changes were evident in the vascular territory classically attributed to the RAH. The clinician should suspect congenital RAH infarction in those infants with congenital upper-extremity athetosis. PMID- 10855657 TI - Congenital skull fracture as a presentation of Menkes disease. AB - We report the rare presentation of Menkes disease with a congenital skull fracture, intracerebral bleeding, and seizures. The diagnosis was made at 3 months of age based on the characteristic features of the syndrome, by which time the child experienced uncontrollable seizures. Following progressive neurodegeneration, death occurred at 3 years of age. The prognosis in Menkes disease is dependent on early copper-histidine therapy. Effective treatment has led to children surviving into adulthood. Diagnosing the syndrome during the neonatal period is difficult. There are no published reports of congenital skull fracture as a presenting sign of Menkes disease. It is concluded that Menkes disease should be considered in any child who presents with congenital skull fracture as early diagnosis and treatment significantly improve the outcome. PMID- 10855658 TI - Autistic regression and Landau-Kleffner syndrome: progress or confusion? PMID- 10855659 TI - Quality of life in families of children with disabilities. PMID- 10855660 TI - Experimental inoculation of calves with laboratory strains of bovine viral diarrhea virus. AB - Diarrhea, erosions and ulcers of the oral mucosa, with conjunctival and nasal discharges, were observed in six calves inoculated with a mixture of two laboratory cytopathic reference strains of bovine viral diarrhea virus (BVDV) Oregon C24 V and NADL. The clinical picture was accompanied by biphasic body temperature elevation, transient leukopenia and a decrease in the number of lymphocytes. High dose of viruses and multiple routes of inoculation promoted the development of clinical and hematological changes typical for BVDV infection although laboratory strains were used. PMID- 10855661 TI - Binding of bovine growth hormone to. Mycobacterium avium ss paratuberculosis. AB - Mycobacterium avium ss paratuberculosis causes a chronic progressive enteritis in cattle and other ruminants referred to as Johne's disease. It also has been suggested by some as possibly being associated with Crohn's disease in humans. In a previous study we observed that incubation of bovine monocytes with recombinant bovine growth hormone (bGH) altered the ingestion and intracellular growth of M. avium ss paratuberculosis in vitro. This led us to investigate whether bGH also has a direct effect on M. avium ss paratuberculosis. We observed that addition of bGH (5 microg/ml) had a direct inhibitory effect on the growth of M. avium ss paratuberculosis in Middlebrook 7H9 broth. In contrast, the growth of Mycobacterium smegmatis was unaffected, even at a bGH concentration of 50 microg/ml. Using 125I-bGH we observed high affinity binding (Kd = 1.32 nM) of bGH to M. avium ss paratuberculosis, with an estimated 204 binding sites per bacillus. To the best of our knowledge, this is the first report of a mammalian hormone binding to this important enteric pathogen. PMID- 10855662 TI - The use of immunomodulators in the control of infectious bovine rhinotracheitis. AB - Three experiments have been carried out to verify the effectiveness of an immunomodulator, Baypamun (Bayer AG) in limiting the spread of Bovine herpesvirus 1 (BHV-1), the causal agent of infectious bovine rhinotracheitis (IBR). In the first experiment, four calves infected with BHV-1 developed severe disease whereas four calves given Baypamun simultaneously with the virus had less severe disease. Four other calves in contact with the infected calves became severely ill but another four given Baypamun were only mildly affected. In the second experiment three calves infected with BHV-1, which reacted with typical disease, were allowed to remain in contact with six calves. All six calves were given Baypamun at various times following the exposure to BHV-1 infection and all showed a much reduced reaction with two treated for 4 days developing no clinical disease. Finally, in the third experiment one calf vaccinated one month before the start of the experiment did not develop any signs of disease when housed together with a calf experimentally infected with BHV-1. Of four other calves, vaccinated when the infected calf showed the first signs of disease, only the two given Baypamun in addition to the vaccine, were protected from clinical disease whereas the two given vaccine only developed classical signs of IBR. In the three experiments the virus shedding by the Baypamun-treated calves resulted to be significantly reduced. PMID- 10855663 TI - Rotavirus and concurrent infections with other enteropathogens in neonatal diarrheic dairy calves in Spain. AB - Faeces samples from 218, one to 30 days old, diarrheic dairy calves in 65 dairy herds were screened for the presence of rotavirus and concurrent infections with coronavirus, Cryptosporidium, F5+ Escherichia coli and Salmonella spp. Calves were grouped according to their age as follows: 1-7, 8-14, 15-21 and 22-30 days. Rotavirus infection was detected in 46.9%, 45.6%, 33.8% and 48.3% of the calves in the respective age-groups. No significant differences in the detection rate of rotavirus were found among calves on the different age-groups. Rotavirus was the only enteropathogen detected in 39 of the 93 (41.9%) diarrheic calves positive to this agent. Concurrent infections with other enteropathogen(s) were detected in 31.3%, 33.3%, 20.6% and 3.4% of the rotavirus infected calves in the age-groups 1 7, 8-14, 15-21 and 22-30 d, respectively. A significant age-associated decrease in the detection rate of mixed infections (p < 0.01) was found. The detection rates of the other enteropathogens considered in calves with rotavirus infection were 20.4% for coronavirus, 85.2% for Cryptosporidium, 16.7% for F5+ E. coli and 1.8% for Salmonella. PMID- 10855664 TI - Maternal antibody passively transferred interferes with rabies vaccination in hamsters. AB - Transference and interference of maternal immunity to offspring after rabies vaccination were studied in hamsters. Females were vaccinated or not before mating and offspring were vaccinated at the age of 7, 14, 21 and 30 days. Other pups were maintained as controls. Thirty days after vaccination pups were challenged intracerebrally with CVS virus. Mouse neutralization tests were used to verify antibody titers. Mortality of 97.0, 76.9, 60.9 and 24.0% was observed in pups vaccinated at 7, 14, 21 and 30 days respectively, born from vaccinated dams, while in pups from non-vaccinated dams, mortality was 51.4, 28.6, 8.7 and 0.0%. Statistically significant associations were found between mortality and age at vaccination, by simple linear regression with y=-3.1169x + 120.8 (p = 0.008; r2=0.98) for litters vaccinated and born from vaccinated dams and y=-2.2541x + 62.7495 (p = 0.03; r2=0.93) for pups vaccinated and born from non-vaccinated dams. Immunological response to vaccination in pups born from vaccinated mothers was delayed 11 days, when compared to that observed in pups of non-vaccinated mothers. PMID- 10855665 TI - Cellular immune response cytokine expression during the initial stage of bovine leukemia virus (BLV) infection determines the disease progression to persistent lymphocytosis. AB - We have established experimental models of bovine leukemia virus (BLV) infection followed by progression to persistent lymphocytosis (PL) positive (BLV+PL+) or PL negative (BLV+PL-) stages of infection. Two out of six BLV infected animals developed PL+ 4 weeks after BLV infection. One other animal became PL+ late in the course of infection and three infected animals stayed PL-. These animals (PL ) exhibited transient lymphocytosis 3-4 weeks after infection and sustained PL- lymphocyte counts up to 24 weeks after infection. Competitive RT-PCR analysis of IFN-gamma mRNA expression revealed that peripheral blood mononuclear cells (PBMC) of animals with PL+ status developed by 4 weeks after infection had augmented IFN gamma mRNA expression 3-4 weeks after BLV infection. However PBMC of animals that sustained a long-termed PL- lymphocyte count had elevated IFN-gamma mRNA expression 1-24 weeks after infection. Competitive RT-PCR analysis of IL-2 mRNA expression showed an increase in the levels of IL-2 mRNA in PL animals. Interleukin-10 (IL-10) mRNAs expression were elevated both in PL+ and PL- animals from 3 and 12 weeks after infection respectively. We suggest that early and extended expression of cellular response cytokines may delay the progression to PL+ in enzootic bovine leukemia. PMID- 10855666 TI - Antigenic characterization of the nucleocapsid protein of Newcastle disease virus by means of a new panel of monoclonal antibodies. AB - Nine monoclonal antibodies (MAB) against nucleocapsid protein (NP) of Newcastle disease virus (NDV) have been prepared and characterized. All the MABs were classified into three groups by means of the competitive binding assay. At least three antigenic sites were delineated on the NP. The 1st site includes two closely located epitopes; the 2nd site includes two related and two distinct epitopes; the 3rd site includes two closely related and one distinct epitopes. PMID- 10855667 TI - Production and separation of carrier-free 145,146Eu from a CsNO3 target using a 16O beam. AB - Carrier free europium isotopes 145,146Eu have been produced by irradiating a CsNO3 target with an 80 MeV 16O beam. Radiochemical separation of the produced europium isotopes from the bulk target matrix of CsNO3 have effectively been done using HDEHP, the liquid cation exchanger. PMID- 10855668 TI - Separation of carrier-free hafnium and lutetium radionuclides produced in 16O activated terbium metal target. AB - Charged particle activation with approximately 88 MeV 16O7+ beam on natural terbium metal foil leads to the production of the short lived carrier-free radioisotopes 170,171Ta and their corresponding daughter products 170,171Hf and 170,171Lu in the target matrix. Liquid-liquid extraction with HDEHP diluted in cyclohexane was carried out for the separation of 170,171Hf and 170,171Lu from the bulk terbium in an aqueous HCl medium. PMID- 10855669 TI - Removal of 125I from radioactive experimental waste with an anion exchange paper membrane. AB - The behavior of radioactive iodide and chloride ions through an anion exchange paper membrane to remove 125I from radioactive experimental waste has been studied with nonequilibrium thermodynamic analyses. Anion exchange paper membrane was found to be electroconductively more permeable to iodide ion than to chloride ion. The iodide ion bound more strongly to the anion exchange site within a membrane phase than the chloride ion by more than twice. The results suggested that an anion exchange paper membrane was appropriate for the filtration removal system. PMID- 10855670 TI - Practical reactor production of 41Ar from argon clathrate. AB - The radionuclide 41Ar has many ideal properties as a gas flow tracer. However, the modest cross-section of 40Ar for thermal neutron activation makes preparation of suitable activities of 41Ar technically difficult particularly for low flux reactors. Argon can however be trapped in a molecular complex called a clathrate that can then be irradiated. We prepared argon clathrate and explored its irradiation and stability characteristics. Argon clathrate can be used to provide gigabecquerel quantities of 41Ar even with low power reactors. PMID- 10855671 TI - Numerical simulation of a TLD pulsed laser-heating scheme for determination of shallow dose and deep dose in low-LET radiation fields. AB - A new method is described to determine the depth-dose distribution in low-LET radiation fields using a thick thermoluminescent dosimeter (TLD) with a pulsed laser-heating scheme to obtain TL glow output. The computational simulation entails heat conduction and glow curve production processes. An iterative algorithm is used to obtain the dose distribution in the detector. The simulation results indicate that the method can predict the shallow and deep dose in various radiation fields with relative errors less than 20%. PMID- 10855672 TI - Improved target system for production of high purity [18F]fluorine via the 18O(p,n)18F reaction. AB - An improved aluminium target system for production of elemental fluorine via the 18O(p,n)18F reaction using a two-step irradiation protocol is described. In the first step highly enriched gaseous oxygen-18 is irradiated with protons to form fluorine-18 which gets deposited on the inner target surface. In the second step, after cryogenic recovery of oxygen-18 target gas, a mixture of elemental 'cold' fluorine and krypton is introduced and a short proton irradiation is done, whereby an isotopic exchange between the gaseous fluorine and the deposited radiofluorine occurs. The second step leads to the recovery of the radiofluorine as [18F]fluorine. Optimisation studies were performed regarding the yield and specific radioactivity of [18F]fluorine. Furthermore, some irradiation parameters relevant to the recovery step were investigated. It was found that a 15 to 20 min irradiation with a beam current of 20 microA is sufficient for the isotopic exchange between the fluorine-carrier and the 18F-radioactivity deposited on the inner wall of the target. The distribution of the 18F-radioactivity deposited on the inner target surface is inhomogeneous, probably due to convection effects. Extensive radioanalytical techniques were applied to characterise the reactivity of [18F]fluorine and to identify undesired nonreactive 18F-compounds, mainly [18F]tetrafluoromethane and [18F]nitrogentrifluoride. The [18F]fluorine produced in the system used has the distinction of having a negligible contamination from those inert 18F-compounds. This is a combined effect of the use of highest purity gases and a welded target construction, which avoids any contact of the gases with organic material during irradiations. The target has proved to be very reliable for production of [18F]fluorine in high yields of up to 34 GBq and specific activities of 350-600 GBq/mmol, both at 30 min after end of activation bombardment. PMID- 10855673 TI - Determination of the retention of 47Ca by whole-body counting. AB - Retention of intravenously or orally administered 47Ca in the human body are described by a two-parameter function. It is then sufficient to make only a few whole-body measurements to determine the retention function, avoiding faeces sampling and stool markers. Seven days after intake the non-absorbed calcium was excreted and the model agreed with the measured relative retention. Absorption of calcium could then, in some cases (e.g. comparative studies), be described by relative retention at the 7th day after intake. PMID- 10855674 TI - A concept of metrological assurance for geophysical methods used to solve geological problems. PMID- 10855675 TI - Estimation of thermal neutron absorption cross-section from K, U and Th concentrations for Miocene rocks from the Carpathian Piedmont in Poland using artificial neural networks. AB - The radiometric K, U and Th concentrations and neutron absorption cross-section sigma a of rock samples obtained from coring are analysed. The cores are from wellbores located in the Sucha-Jordanow region (Carpathian Mountains) and from gas producing Miocene formations in the Carpathian foothills. Correlation coefficients between the neutron absorption cross-section (sigma a) and K, U and Th concentrations are presented. Neural network representation of the function sigma a = f(K, U, Th) obtained for a region can later be used for sigma a estimation from spectrometric probe results in uncored wells. PMID- 10855676 TI - An examination of psychosocial variables moderating the relationship between life stress and injury time-loss among athletes of a high standard. AB - Based on Williams and Andersen's model of stress and athletic injury, six psychosocial variables were assessed as possible moderators of the relationship between life stress and injury among 121 athletes (65 males, 56 females) competing in a variety of sports at state, national or international level. No significant effects of the sex of the participants were evident. Correlational analyses revealed moderator effects of several variables. Specifically, dispositional optimism and hardiness were related to decreased injury time-loss in athletes when positive life change increased, and global self-esteem was associated with decreased injury time-loss when both negative life change and total life change increased. The results indicate that athletes with more optimism, hardiness or global self-esteem may cope more effectively with life change stress, resulting in reduced injury vulnerability and recovery rates. PMID- 10855677 TI - The effect of knee angle on the external validity of isometric measures of lower body neuromuscular function. AB - The aim of this study was to evaluate the effect of varying knee angle (120 degrees and 90 degrees) on the external validity of an isometric leg press test with reference to vertical jump performance. Isometric peak force (PF120 and PF90), rate of force development (RFD120 and RFD90), and maximum height reached with a squat jump and counter-movement jump were measured in 14 males. Although RFD120 was significantly correlated with squat jump and counter-movement jump performance (r = 0.71 and 0.69), and the correlations with PF120 approached statistical significance (r = 0.53 and 0.50), neither PF90 nor RFD90 was significantly related to vertical jump performance. Furthermore, although both RFD120 and PF120 were significantly different between the best five and the worst five jumpers, RFD90 and PF90 did not differentiate between individuals' vertical jump performance. We conclude that the choice of joint angle affects the external validity of isometric strength testing. Based on our results, we recommend accurate control of biomechanical specificity and assessment at different angles to find the position at which isometric strength testing is most comfortable. PMID- 10855678 TI - Kinematic analysis of shot-putting performed by wheelchair athletes of different medical classes. AB - The aim of this study was to identify those kinematic characteristics that are most closely related to an athlete's medical classification and measured distance of a put. Two S-VHS camcorders (60 fields per second) were used to record the performance of 17 males of different classes. Each participant performed six trials and the best trial for each was selected for analysis. Three-dimensional kinematics of the shot and upper body segments at the instant of release and during the forward thrust (delivery) were determined. The average speeds and angles of the shot at release for different classes (5.3-7.8 m x s(-1) and 21.2 to 34.4 degrees, respectively) were smaller than those exhibited by elite male able-bodied throwers. The height of the shot at release, the angular speed of the upper arm at release, the range of motion of the shoulder girdle during the delivery, and the average angular speeds of the trunk, shoulder girdle and upper arm during the delivery, were all significantly correlated with both the classification and measured distance (P < 0.05). The results indicate the importance of achieving a high average angular speed for each upper body segment during the delivery. PMID- 10855679 TI - Reproducibility of the maximum accumulated oxygen deficit and run time to exhaustion during short-distance running. AB - The aim of this study was to determine the reproducibility of the maximal accumulated oxygen deficit and the associated exercise time to exhaustion during short-distance running. Fifteen well-trained males (mean +/- s: VO2max = 58.0+/ 4.6 ml x kg(-1) x min(-1)) performed the maximum accumulated oxygen deficit test at an exercise intensity equivalent to 125% VO2max. The test was repeated at the same time of day on three occasions within 3 weeks. There was no significant systematic bias between trials for either maximum accumulated oxygen deficit (man +/- s: trial 1 = 69.0+/-13.1; trial 2 = 71.4+/-12.5; trial 3 = 70.4+/-15.0 ml O2 Eq x kg(-1); ANOVA, F = 0.70, PP= 0.51) or exercise time to exhaustion (trial 1 = 194 + 31.1; trial 2 = 198 + 33.2; trial 3 = 201 + 36.8 s; F= 1.49, P = 0.24). In addition, other traditional measures of reliability were also favourable. These included intraclass correlation coefficients of 0.91 and 0.87, and sample coefficients of variation of 6.8% and 5.0%, for maximum accumulated oxygen deficit and exercise time to exhaustion respectively. However, the '95% limits of agreement' were 0+/-15.1 ml O2 Eq (1.01 multiply/divide 1.26 as a ratio) and 0+/ 33.5 s (1.0 multiply/divide 1.18 as a ratio) for maximum accumulated oxygen deficit and exercise time to exhaustion respectively. We estimate that the sample sizes required to detect a 10% change in exercise time to exhaustion and maximum accumulated oxygen deficit after a repeated measures experiment are 10 and 20 respectively. Unlike the results of previous maximum accumulated oxygen deficit studies, we conclude that it is not a reliable measure. PMID- 10855680 TI - The effects of substrate and fluid provision on thermoregulatory and metabolic responses to prolonged exercise in a hot environment. AB - A high ambient temperature reduces the capacity to perform prolonged exercise. Total carbohydrate oxidation is less, and thus glycogen depletion is not limiting. Fluid ingestion in the heat should, therefore, focus on maintenance of hydration status rather than on substrate provision. Six healthy males cycled to exhaustion at 60% of maximum oxygen consumption (VO2max) with no drink, ingestion of a 15% carbohydrate-electrolyte drink (1.45+/-0.29 litres) or ingestion of a 2% carbohydrate-electrolyte drink (3.12+/-0.47 litres). The ambient temperature was 30.2+/-0.6 degrees C (mean +/- s), with a relative humidity of 71+/-1% and an air speed of approximately 0.7 m x s(-1) on all trials. Weighted mean skin temperature, rectal temperature and heart rate were recorded and venous samples drawn for determination of plasma volume changes, blood metabolites, serum electrolytes and osmolality. Expired gas was collected to estimate rates of fuel oxidation. Exercise capacity was significantly (P < 0.05) different in all trials. The median (range) time to exhaustion was 70.9 min (39.4-97.4 min) in the no-drink trial, 84.0 min (62.7-145 min) in the 15% carbohydrate trial and 118 min (82.6-168 min) in the 2% carbohydrate trial. The 15% carbohydrate drink resulted in significantly (P < 0.05) elevated blood glucose and total carbohydrate oxidation compared with the no-drink trial. The 2% carbohydrate drink restored plasma volume to pre-exercise values by the end of exercise. No differences were observed in other thermoregulatory or cardiorespiratory responses between trials. These results suggest that fluid replacement with a large volume of a dilute carbohydrate drink is beneficial during exercise in the heat, but the precise mechanisms for the improved exercise capacity are unclear. PMID- 10855681 TI - The relationship between evaluative concerns and sport competition state anxiety among youth skiers. AB - Thirty-four youth competitive skiers (mean age = 13.74 years) completed measures of social evaluative concern and competitive anxiety. Consistent with past research, regression analyses showed that cognitive anxiety was related to performance-specific evaluative concerns. However, contrary to current conceptualizations of sport competition anxiety, somatic anxiety was correlated with concerns about evaluation of other non-performance aspects of ski racing. Competitive skiers were most concerned about parents' and friends' evaluations of their performance, and other competitors' and friends' evaluations of their skiing in general. These findings are discussed in relation to the theory and management of sport competition state anxiety. PMID- 10855682 TI - Biomechanical loading in the triple jump. AB - The triple jump is a demanding field event in which a jumper must tolerate extremely high impact forces while maintaining high horizontal speed. The present study was designed to clarify the mechanical loading characteristics and the role of neuromuscular function in the triple jump. Seven national triple jumpers (4 males, 3 females) volunteered to perform 3-6 jumps. The mean best performances were 14.32+/-0.45 m and 11.90+/-0.28 m for males and females, respectively. The three longest triple jumps for each jumper were selected for final analysis. The mean contact times were 0.139 s (hop), 0.157 s (step) and 0.177 s (jump). The largest ground reaction forces were observed in the step (15.2 times body weight), while the highest peak pressures were recorded under the heel and forefoot. The plantar pressure of the lateral side of the forefoot was highly related to the length of the triple jump (P < 0.05-0.01). In addition, electromyograms of both legs Suggested that mechanical loading places high demands on the neuromuscular system, as characterized by the high rate of activation in the pre-activity phase followed by high eccentric activity. Thus, the high activities of the gastrocnemius, vastus lateralis and hip extensor muscles seem to play an important role in preventing unnecessary yielding of the jumper during the braking phase. PMID- 10855683 TI - Angiogenesis in the corpus luteum. AB - The ovarian corpus luteum plays a critical role in reproduction because it is the primary source of circulating progesterone. After ovulation, as the corpus luteum forms from the wall of the ruptured follicle, it grows and vascularizes extremely rapidly. In fact, the rates of tissue growth and angiogenesis in the corpus luteum rival those of even the fastest growing tumors. Thus, the corpus luteum provides an outstanding model for studying the factors that regulate the angiogenic process, which is critical for normal tissue growth, development, and function. In agreement with data from other tissues, vascular endothelial growth factors (VEGF) seem to be a major angiogenic factor responsible for vascularization of the developing corpus luteum. Recent data suggest that luteal expression of VEGF occurs primarily in specific perivascular cells, including arteriolar smooth muscle and capillary pericytes, and is regulated primarily by oxygen levels. In addition, soon after ovulation, pericytes derived from the thecal compartment appear to be the first vascular cells to invade the developing luteal parenchyma. The granulosa-derived cells produce a factor that stimulates pericyte migration. Moreover, nitric oxide (NO), which is a potent vasodilator and can stimulate VEGF production and angiogenesis, is expressed in endothelial cells of luteal arterioles and capillaries, often in association with expression of VEGF by luteal perivascular cells. Thus, we have proposed a model for the initial process of luteal vascularization in which hypoxia plays a major role. In this model, which we believe will apply to other tissues as well, a paracrine loop exists between the vascular endothelial cells, which produce NO, and the peri-endothelial cells (vascular smooth muscle and pericytes), which produce VEGF, to ensure coordinate regulation of luteal vasodilation and angiogenesis. PMID- 10855684 TI - Large goiter and multiple rib tumors. AB - We report an interesting case of a 47-yr-old who had a large goiter and multiple rib tumors. The patient was initially suspected of having thyroid cancer, which had metastasized on the ribs, based on imaging studies. However, laboratory tests revealed a high level of ionized calcium and parathyroid hormone (PTH). The large goiter was diagnosed as having parathyroid tumors owing to the high level of PTH in the tissue fluid. The biopsy specimen from a rib tumor was diagnosed as containing brown tumors associated with primary hyperparathyroidism (PHP). The patient also had prolactinoma and pancreatic gastrinoma. Her daughter had both prolactinoma and PHP, and her brother and her father had PHP. Thus, the patient was diagnosed as having multiple endocrine neoplasia type 1. PMID- 10855685 TI - Adrenomedullin expression in the human endometrium. AB - Immunohistochemical studies were performed using a specific antibody to human adrenomedullin (AM) to determine its presence and cellular localization in the human endometrium, in the different phases of the menstrual cycle, and in the postmenopausal period. Specimens were obtained from 21 patients who underwent abdominal hysterectomy for various reasons. The endometrium had no pathological lesion in all cases. In the early and mid proliferative phases of the menstrual cycle, no immunostaining for AM was noted in the endometrium. AM immunostaining in the endometrium became apparent in the late proliferative phase. The staining intensity of AM in the endometrium became more abundant in the secretory phase. No appreciable difference in the staining intensity of AM in the endometrium was noted among early, mid, and late secretory phases. Immunostaining for AM was evident in both the epithelial and stromal compartments of the endometrium. In the postmenopausal endometrium, there was intense immunostaining for AM only in the stromal compartment. This is the first study to demonstrate the expression of AM in the endometrium in relation to the menstrual cycle. The results obtained suggest the participation of AM in the growth and differentiation of the endometrium. PMID- 10855686 TI - Circulating endothelium-related adhesion molecules in black South African patients with Graves' disease. AB - Elevated serum concentrations of endothelium-associated adhesion molecules occur in Graves' disease. However, no data exist in African subjects, among whom the incidence is rising. Therefore, 20 black South Africans with Graves' hyperthyroidism were evaluated and 10 healthy controls were also studied. Quantitative determinations of soluble intercellular adhesion molecule-1 (sICAM 1), soluble vascular cell adhesion molecule-1 (sVCAM-1), and soluble E-Selectin (sE-Selectin) were performed in serum samples by an enzyme-linked immunosorbent assay. Mean levels of sVCAM-1 were significantly increased in the thyrotoxic patients compared to controls, but this did not apply to the other adhesion molecules. The presence of ophthalmopathy in 12 patients did not further increase the mean sVCAM-1 concentration, and the administration of antithyroid medication in 5 patients had no measurable effect. In conclusion, sVCAM-1 appears to be a useful marker of active Graves' disease in black South Africans although it does not seem to reflect the occurrence of eye involvement in such patients. PMID- 10855687 TI - Regulation of PTH/PTH-related protein receptor expression by endogenous PTH related protein in the rat osteosarcoma cell line ROS 17/2.8. AB - We have utilized clonal lines of the rat osteoblastic cell line ROS 17/2.8 stably transfected with full-length parathyroid hormone-related protein (PTHrP) cDNA in a sense or an antisense orientation to examine the effects of alteration in the production of endogenous PTHrP on expression of the PTH/PTHrP receptor. In the stably transfected clonal cell lines, changes in PTH/PTHrP receptor expression were evaluated by Northern blot analysis, whole-cell ligand binding of 125I [Tyr36] PTHrP (1-36), and exogenous PTHrP (1-34)-stimulated cyclic adenosine monophosphate (cAMP) accumulation. Compared to control (vector-transfected) cells, PTHP-overproducing (sense-transfected) cells exhibited a marked decrease in the expression of PTH/PTHrP receptor mRNA and PTHrP ligand binding, as well as a corresponding decrease in the PTHrP (1-34)-stimulated cAMP response. By contrast, the antisense-transfected cells showed a marked increase in expression of PTH/PTHrP receptor mRNA and PTHrP (1-34) ligand binding, but a significant increase in the PTHrP (1-34)-stimulated cAMP response was not detected. Using antisense-transfected ROS cells, PTH/PTHrP receptor mRNA expression and 125I [Tyr36] PTHrP (1-36) binding were downregulated by treatment for 24 h with exogenous PTHrP (1-36), forskolin, or dibutyryl cAMP. The findings extend those of earlier studies showing receptor downregulation by exogenous PTH by indicating that endogenous PTHrP, as well as circulating PTH, may help regulate receptor production; and suggesting that even very low concentrations of the peptide may influence receptor production. PMID- 10855688 TI - Sex difference in serum luteinizing hormone postgonadectomy in the rat: role of gamma-aminobutyric acid-ergic inhibition. AB - In adult male rats, serum luteinizing hormone (LH) rises within a few hours of castration. By contrast, in adult female rats, serum LH does not increase reliably until 4-6 d after ovariectomy. The release of gonadotropin-releasing hormone (GnRH) declines in female rats postovariectomy, suggesting an increase in inhibition of the release of GnRH. We investigated whether differences in gamma aminobutyric acid (GABA)-ergic transmission, which inhibits GnRH release, accounts for the sex difference in the response of serum LH to gonadectomy. We examined the effects of GABA-A receptor antagonist bicuculline methiodide (BMI), GABA-B receptor antagonist phaclofen, and transaminase inhibitor aminooxyacetic acid (AOAA), injected subcutaneously, on the postgonadectomy rise in LH. AOAA prevented the postcastration rise in male rats (p < 0.05). Female rats treated with BMI, phaclofen, or both BMI and phaclofen (p < 0.05) showed a significant increase in LH postovariectomy. BMI had no effect in male rats. GnRH antagonist blocked the BMI-induced increase in serum LH. We conclude that the delay in the rise of serum LH in female rats postovariectomy is at least partly owing to GABAergic inhibition of the release of GnRH. PMID- 10855689 TI - Novel studies on influence of gonadotropins and insulin-like growth factor-I on growth of cumulus oophorus in the rat. AB - In the final developmental stage of a Graafian follicle, there are two functionally distinct types of granulosa cells: the cumulus cells (CCs) and the mural granulosa cells (MGCs). Previous studies focused on follicle-stimulating hormone (FSH) and insulin-like growth factor I (IGF-I) interactions in MGCs. Our goal was to study these interactions in CC proliferation. Immature rats received in vivo treatments of either saline, equine chorionic gonadotropin (eCG) with high FSH activity, an IGF-I analog (LR3-IGF-I) with poor binding to IGF-I binding proteins, or a combination of both hormones. CCs from each in vivo treatment were then cultured and treated in vitro with either saline, FSH, or IGF-I. CCs proliferation were assessed by measurement of 3H-thymidine incorporation. Prior in vivo treatment with eCG resulted in the highest proliferative activity of CCs when combined with FSH in vitro treatment. In vivo treatment with LR3-IGF-I had no effect on CC replication. CC replication was higher in FSH in vitro treatment than of IGF-I. The combination of eCG and LR3-IGF-I was the only in vivo treatment to stimulate higher CC proliferation with IGF-I in vitro treatment. This study suggests that FSH does not act through IGF-I, a mechanism previously proposed. PMID- 10855690 TI - Mitogen-activated protein kinase activates human placental lactogen-B enhancer by an NF-IL6-dependent pathway. AB - Computer analysis of the human placental lactogen-B (hPL-B) enhancer reveals two putative binding sites for the transcription factor NF-IL6, but the role of NF IL6 in the regulation of the enhancer is unknown. Using gel mobility shift and supershift assays, we demonstrated that NF-IL6 binds to both enhancer sites. Transient transfection studies indicated that the transcription factor NF-IL6 stimulates hPL-B enhancer activity by 4.4-fold in primary cultures of human trophoblast cells and by 32.0- and 8.4-fold in JAR and BeWo choriocarcinoma cells, respectively. Overexpression of MEK (mitogen-activated protein [MAP] kinase kinase), which is known to stimulate phosphorylation of NF-IL6, induced a 3.6-fold increase in hPL-B enhancer activity. The induction by MEK was completely inhibited by an expression plasmid for a dominant/negative mutant of NF-IL6 or by mutation of the NF-IL6 binding sites on the enhancer. PD98059, an inhibitor of MEK, inhibited hPL release from cultured trophoblast cells by about 50%. Taken together, these results indicate that MAP kinase stimulates the hPL-B enhancer by an NF-IL-6-dependent pathway. PMID- 10855691 TI - Ovarian action of leptin: effects on insulin-like growth factor-I-stimulated function of granulosa and thecal cells. AB - To test the hypothesis that leptin signals metabolic information to the reproductive system in cattle by directly affecting IGF-I-induced ovarian cell function, granulosa and thecal cells from bovine ovarian follicles were cultured for 2 d in serum-free medium with added hormones. Recombinant human leptin at 30 and 300 ng/mL had no effect on basal thecal cell steroidogenesis or thecal cell numbers. However, 300 but not 30 ng/mL of leptin attenuated (p < 0.05) luteinizing hormone-induced androstenedione production by 24% in the absence of IGF-I and by 16% in the presence of IGF-I. Leptin had no effect on IGF-I-induced estradiol production in the presence of follicle-stimulating hormone (FSH), but at 100 ng/mL, leptin inhibited (p < 0.05) FSH plus IGF-I-induced progesterone production and granulosa cell proliferation by 29 and 31%, respectively. Leptin did not compete for 125I-IGF-I binding to granulosa or thecal cells, whereas unlabeled IGF-I did. In conclusion, leptin has weak inhibitory effects on gonadotropin- and/or IGF-I-induced steroidogenesis of thecal and granulosa cells. PMID- 10855692 TI - Comparative analysis of the effects of gonadotropin-releasing hormone agonist on the proliferative activity, apoptosis, and steroidogenesis in cultured porcine granulosa cells at varying stages of follicular growth. AB - This study was conducted to analyze comparative effects of gonadropin-releasing hormone (GnRH) agonist on the proliferation, apoptosis, and differentiated function of cultured porcine granulosa cells from varying follicle stages. Comparative analyses of porcine granulosa cells from varying follicle stages to respond to GnRH agonist were performed in terms of proliferating cell nuclear antigen (PCNA) expression, occurrence of apoptosis, and 17beta-estradiol (E2) and progesterone (P) secretion. PCNA expression was examined by the avidin/biotin immunoperoxidase method with a monoclonal antibody to PCNA, and apoptosis was assessed by in situ DNA 3'-end labeling method and DNA fragmentation analysis. E2 and P were measured by radioimmunoassays. The PCNA positive rate of granulosa cells cultured in the presence of GnRH agonist (10(-9) M) was lower compared with that of cells cultured in the absence of GnRH agonist. However, the apoptosis positive rate was higher, and E2 and P secretion by cultured granulosa cells was lower in the presence of GnRH agonist (10(-9) M) compared with that in the absence of GnRH agonist. The inhibitory effect of GnRH agonist on PCNA positive rate of cultured cells was prominent in granulosa cells from small and medium but not from large follicles. By contrast, the inhibitory effect of GnRH agonist on E2 and P secretion by cultured cells was prominent in granulosa cells from large but not small and medium follicles. The stimulatory effect of GnRH agonist on apoptosis positive rate of cultured cells was, however, uniform regardless of the stages of follicular growth. These results demonstrate that GnRH agonist exerts diverse actions on granulosa cells over the course of follicular growth. One downregulates granulosa proliferation in immature follicles as well as steroidogenesis in mature follicles, and the other upregulates apoptosis of granulosa cells regardless of the stages of follicular growth. PMID- 10855693 TI - Isolation and characterization of the androgen receptor mutants with divergent transcriptional activity in response to hydroxyflutamide. AB - A yeast genetic screening was developed to isolate androgen receptor (AR) mutants with divergent transactivation characteristics in response to hydroxyflutamide (HF), an active metabolite of flutamide used for prostate cancer treatment. Two mutants carrying the substitution C685Y or E708K were isolated and characterized. Substitution of C685Y for wild-type AR (wtAR) rendered the receptor supersensitive to androgenic activity from HF and female hormones such as 17beta estradiol (E2) and progesterone (P). Similar effects were observed in the AR mutant, named T876AAR, isolated from LNCaP cells. Surprisingly, we found that C685YAR7, but not T876AAR7, could be activated by casodex (bicalutamide), a nonsteroidal pure antiandrogen, with an induction fold 3- to 5-fold times higher than that for wild type or T876AAR. By contrast, although replacement of E708K for wtAR showed little effect on dihydrotestosterone-mediated transactivation, E708KAR lost its transcriptional response from many other ligands. The effects of ligands on E708KAR could be controlled at the DNA-binding level owing to the finding of a significant decrease in the DNA-binding ability once E708KAR was bound to HF, E2, or P. Together, these results suggest that C685YAR can be a novel tool for assaying the androgenic activity from antiandrogens, and the mechanism revealed from E708KAR could provide a possible explanation for the partial androgen insensitivity syndrome in men with a natural E708KAR mutation. PMID- 10855694 TI - Detection and characterization of endothelin in transformed human osteoblast cell culture medium. AB - Endothelin-1 (ET-1), a 21 amino acid peptide originally purified from conditioned medium of cultures of porcine aortic endothelial cells, is recognized also as a product of many other cells such as epithelial cells, glial cells, and neurons. It is now recognized that at least ET-1 plays an important role in bone metabolism. It has been shown that ET-1 inhibits osteoclast bone resorption by a direct effect on cell motility and it can also activate phospholipase C in the osteoblast. Furthermore, several studies have shown that ET-1 stimulates the formation of inositol phosphates, the synthesis of DNA, the mobilization of calcium from extra- and intracellular pools, the activation of phospholipase D, and the stimulation of tyrosine phosphorylation in osteoblast-like (MC3T3-E1 and UMR-106) cells. The aim of the present study was to detect and characterize the presence of endothelin in transformed human osteoblast cell culture medium (HTb96) by radioimmunoassay and chromatography methods. Immunoreactive endothelin (IR-ET) was undetectable in the medium incubated at 0.5 and 1 h and was 3.2 +/- 0.2 fmol/10(5) cells (mean +/- SEM, n = 6) at 2 h, 9.5 +/- 0.5 fmol/10(5) cells at 6 h, 19.8 +/- 2.1 fmol/10(5) cells at 24 h, and 23.7 +/- 2.0 fmol/10(5) cells at 48 h, respectively. Sephadex G-25 superfine chromatography and fast protein liquid chromatography studies showed that >90% of IR-ET in the culture medium coeluted with synthetic ET-1. These results show that ET-1 could be formed by transformed human osteoblasts. Further studies should be conducted to elucidate the physiological role of endothelins as possible autocrine, paracrine, or endocrine factors in calcium and bone metabolism. PMID- 10855695 TI - Influence of estrogen deficiency and replacement on T-cell populations in rat lymphoid tissues and organs. AB - Estrogen deficiency following ovariectomy or menopause results in bone loss. Although evidence strongly suggests that the immune system is involved in the pathogenesis of estrogen-deficient osteoporosis, it is not clear what role, if any, the T-lymphocyte plays in this process. Therefore, we examined the distribution of T-cell subsets in lymphoid organs and tissues, under varying estrogenic states in the rat. Six-month-old female Sprague-Dawley rats, ovariectomized (Ovx) and sham-operated, were randomized 5 d post-surgery into six groups to receive the following treatments: (A) sham/placebo; (B) sham/low-dose E2; (C) sham/high-dose E2; (D) Ovx/placebo; (E) Ovx/low-dose E2; (F) Ovx/high dose E2. Half of the treated rats (groups A-F) were sacrificed on d 14; the remainder on d 28. Following euthanasia, mononuclear cells were isolated from the thymus, peripheral blood, spleen, lymph node and bone marrow, and were labeled for flow cytometric analysis using mouse anti-rat monoclonal antibodies directed against CD5, CD4, and CD8 antigenic markers. In the thymus, ovariectomy caused a dramatic increase and E2 treatment caused a dose-dependent decrease in weight that was proportional to the number of thymocytes. In the bone marrow, ovariectomy caused a significant reduction in the percentage of all T-cell subsets examined and this effect persisted throughout the duration of the study. Estrogen replacement therapy at the low-dose reversed the effects of ovariectomy and high-dose E2 treatment caused an increase in T-cell subsets in both the sham and Ovx groups, an effect that was more pronounced at d 14 compared with d 28. Although the percentages of some T-cell subsets in the other lymphoid organs/tissues were altered by ovariectomy or E2 treatment at d 0 and 14, all these changes had normalized by d 28 except for CD5 and CD4 cells in peripheral blood. In summary, with the exception of T-lymphocytes in the bone marrow, the effects of varying estrogenic states on T-cells were variable and transient. The influence of estrogen status on bone marrow T-lymphocytes suggests that these cells may play a role in mediating the effects of estrogen on bone turnover and warrant additional studies focusing on the functional role of T-cells in the bone marrow compartment. PMID- 10855696 TI - Identification of an essential cis-acting element (TR2-PACE) in the 5' promoter of human TR2 orphan receptor gene. AB - The human TR2 orphan receptor (TR2) is a member of the steroid/thyroid hormone receptor superfamily. It has been shown to be expressed in a wide variety of tissues during development. Using deletion mutation analyses and transient transfection CAT assays, we demonstrated here that a DNA fragment of 103 bp, with a sequence from +65 to -38, containing an initiator is capable of serving as a core promoter to initiate basal level transcription; further extending of this core promoter sequence up to -441 maximizes the reporter gene expression. Within this positive regulatory region (-441/+65), we were able to narrow the regulation responsible sequence down to a small 64-bp (-263/-201) DNA fragment named the TR2 promoter activating cis-element (TR2-PACE). Further deletion mutagenesis and shifting of the insert position followed by reporter assays demonstrated that this TR2-PACE is essential for high-level induction of a heterologous core promoter's activity in a position-dependent nature. In addition, orientation tests indicated that the sense, but not antisense orientation increased the TR2 core promoter activity. Moreover, electrophoresis mobility shift assays and Southwestern analyses suggested that TR2-PACE may interact with unknown specific nuclear proteins for its enhancer activity. Together, our data suggest that TR2 PACE is a position-dependent and, in the case of TR2 core promoter (TATA-less), an orientation-dependent cis-activating element required for maximal expression of the TR2 gene. PMID- 10855697 TI - Functional morphology of the gas-gland cells of the air-bladder of Oreochromis alcalicus grahami (teleostei: cichlidae): an ultrastructural study on a fish adapted to a severe, highly alkaline environment. AB - Oreochromis alcalicus grahami is a small cichlid fish that lives in shallow peripheral lagoons of Lake Magadi, Kenya. The internal surface of the air-bladder is highly vascularized, illustrating possible utilization as an accessory respiratory organ. The wall of the bladder consists of five distinct tissue layers. From the outer to the inner surfaces are: a squamous, undifferentiated epithelial cell; a collagen-elastic tissue space; a smooth muscle tissue band; an inner connective tissue space; and columnar gas-gland cells projecting into the lumen. The cell membrane over the perikarya of the gas-gland cells was sporadically broken. The disruptions were ascribed to possible physical damage by discharging gas-bubbles. Juxtaluminally, the gas-gland cells attached across tight junctions. The cells have highly euchromatic nuclei and conspicuously large intracytoplasmic secretory bodies. On the blood capillary facing (basal) aspect, the cell membrane of the gas-gland cells is highly amplified. The cells insert onto a smooth muscle tissue band. The morphological features and the topographical arrangement of the gas-gland cells in O. a. grahami are indicative of an operative exchange of materials between them and the underlying tissue components especially the blood capillaries. For a fish that subsists in hot, highly saline and alkaline water heavily invested by avian predators and where the partial pressure of oxygen diurnally shifts from virtual anoxia to hyperoxia, development of a versatile air-bladder for efficient buoyancy control conforms to the functional demands placed on it by a unique environment. Illustratively, instead of the gas-gland morphology in O. a. grahami resembling that in the freshwater fishes, the group from which the fish has evolved, it compares more closely to that of the marine fish. This similarity suggests amazing parallel evolutionary adaptation to biophysically corresponding aquatic milieus. PMID- 10855698 TI - Connexin43 in porcine myocardium and non-pregnant myometrium. AB - It is generally accepted that connexin43 (Cx43) is a major constituent of heart and myometrial gap junctions. However, the presence of Cx43 gap junctions in non pregnant myometrium is still poorly documented. Tissue sections of porcine heart and non-pregnant uterus and myometrial smooth muscle cell cultures were immunostained with monoclonal antibody against Cx43. In the heart, intensive immunostaining was confined to the intercalated discs as previously reported. In the non-pregnant uterus, punctuate immunostaining of Cx43 was seen throughout the myometrium along cell interfaces between myocytes. The expression of Cx43 was sustained in cultured smooth muscle cells isolated from non-pregnant myometrium. Western blotting has detected single isoform of Cx43 in both, cardiac and myometrial tissues. The electrophoretic mobility of porcine heart Cx43 was similar to that of myometrial isoform but different from the pattern of mobility of Cx43 of the rat heart. Hence, porcine myometrium may provide attractive model for studying cellular mechanisms triggering expression of gap junction protein in normal (non-pregnant) uterus. PMID- 10855699 TI - Immunolocalization of stress proteins and extracellular matrix proteins in the rat tibia. AB - Stress proteins (heat shock proteins [hsps]) serve a number of protective functions, including protection from apoptosis and acting as chaperones during protein biosynthesis. For example, hsp 27 has been defined as a chaperone for the G3 domain of aggrecan, while hsp 47 is the chaperone for type I collagen. Separate cytoprotective roles for hsp 27 and hsp 70 have been demonstrated. The aim of this study was to define the expression of hsps in osteoblastic and chondrocytic cells of the growing rat long bone in relationship to the immunohistochemical localization of aggrecan, type I collagen and the presence of fragmented DNA that defines apoptotic events. Tibiae were harvested from Fisher 344 rats (n=6) and fixed in 10% buffered formalin. Samples were decalcified in 10% EDTA, bisected, and processed for histologic examination. Sections (5 mm) were immunohistochemically stained using a streptavidin-biotin detection method. Co-localization of hsps with apoptosis was achieved using the TUNEL procedure. In the rat tibia growth plate, aggrecan was generally distributed throughout cartilage and chondrocytes. However, hsp 27 expression was observed only in the lower hypertrophic chondrocytes. hsp27 was present in osteoblasts lining newly formed bone. hsp 47 staining was also prominent within these osteoblasts where collagen type I immunolocalization occurred. The inducible form of hsp 70 was localized to the osteoblastic cells lining new bone in the primary spongiosa. In cartilage, DNA fragmentation was restricted to the hypertrophic, hsp27-positive, chondrocytes. In contrast, DNA fragmentation was not co-localized with hsp27 positive osteoblastic cells of the primary spongiosa, although occasional apoptotic cells were identified. These results indicate that apoptosis is a mechanism by which hypertrophic chondrocytes are eliminated from cartilage prior to calcification, but that other mechanisms are also likely to be involved. They also suggest that hsps have cytoprotective and biosynthetic functions within osteoblasts and chondrocytes, but apoptotic signals may override these effects in some instances, resulting in apoptosis. PMID- 10855700 TI - Immunoidentification of CRF-like material in the intermaxillary glands during Bufo arenarum development. AB - The presence of corticotropin-releasing factor-like material in the intermaxillary glands was studied by immunocytochemical techniques during the metamorphosis of Bufo arenarum. The intermaxillary glands appeared at stage XV (midprometamorphosis) with CRF-like material slightly immunoreactive. These glands are located posterior to the premaxillae and between the nasal capsules in the roof of the mouth and are formed of alveoli or tubules. During metamorphic climax, corticotropin-releasing factor-like material was identified strongly immunostained at the apices of the secretory cells. It was observed that collecting ducts of the gland open to the anterior palatal surface suggesting that the secretion could be ingested by tadpoles. Our results clearly showed that ir-CRF-like material present in the intermaxillary glands is ingested by tadpoles during metamorphosis and could play an important role during amphibian development. PMID- 10855701 TI - Ultrastructural localization of alpha-keratins in the regenerating epidermis of the lizard Podarcis muralis during formation of the shedding layer. AB - In the epidermis of lizards, alpha- and beta-keratins are sequentially produced during a shedding cycle. Using pre- and post-embedding immunocytochemistry this study shows the ultrastructural distribution of 3 alpha-keratin antibodies (AE1, AE2, AE3) in the renewing epidermis and in the shedding complex of the regenerating tail of the lizard Podarcis muralis. The AE1 antibody that recognizes acidic low MW keratins is confined to tonofilament bundles in basal and suprabasal cells but is not present in keratinizing beta- and alpha-cells. The AE2 antibody that recognises higher MW keratins weakly stains pre-keratinized cells and intensely keratinized alpha-layers. A weak labeling is present in small electrondense areas within the beta-layer. The AE3 antibody, that recognizes low and high MW basic keratins, immunolabels tonofilament bundles in all epidermal layers but intensely the alpha-keratinizing and keratinized layers (mesos, alpha , lacunar and clear). Keratohyalin-like granules, present in the clear cells of the shedding layer, are negative to these antibodies so that the cornified clear layer contains keratins mixed with non-keratin material. The AE3 antibody shows that the mature beta-layer and the spinulated folds of the oberhautchen are labeled only in small dense areas among the prevalent electron-pale beta-keratin material. Therefore, some alpha-keratin is still present in the beta-layer, and supports the idea that alpha-keratins (basic) function as scaffold for beta keratin deposition. PMID- 10855702 TI - The in vitro life-span of human periodontal ligament fibroblasts. AB - The in vitro life-span of human periodontal ligament fibroblasts (PDLF) was studied on clones from periodontium of teeth extracted due to periodontitis and dental caries (69 clones/192 individuals, aged 20-80 years) and from periodontium of teeth extracted for orthodontic reasons (23 clones/26 individuals, aged 15-19 years). In the primary cultures the ratio of the number of cells expressing senescence-associated beta-galactosidase (SA-beta-Gal) to the total number of cells is significantly larger in PDLF (92 clones; 11.1+/-4.9%) than in human gingival fibroblasts (GF) (10 clones; 0.5+/-0.1 %). The finite population doubling numbers (PD) of PDLF are not age-matched and the mean PD of PDLF (7.1+/ 2.9) is significantly smaller than GF (28.5+/-3.2), IMR-90 (human lung fibroblasts, 5 clones; 44.3 +/- 2.2), and human osteoblasts (5 clones; 19.7+/ 1.4). Comparing the ratio of the number of SA-beta-Gal positive cells to the total number of cells in primary culture, and the finite PD in PDLF cultures: 1) the ratio of 15-19 years old donor group is significantly smaller than in the other donor groups (20-29, 30-39, 40-49, 50-59 and 60-80 years old), and 2) there were no statistically significant differences among the 20-29, 30-39, 40-49 and 50-59 year old donor groups, and the 30-39, 40-49, 50-59 and 60-80 year old donor groups. These findings suggest that the in vitro life-span of PDLF is shorter than other fibroblasts in the connective tissues and that PDLF may undergo senescence in adult clones without relation to donor's age. There may be more aged fibroblasts in periodontium than in other tissues, such as gingiva and lung. PMID- 10855703 TI - Ontogeny of adenosine deaminase in developing trophoblast and decidual cells of rat and hamster. AB - The enzyme adenosine deaminase (ADA) is expressed at high level in the tissue of foeto-maternal interface during early pregnancy. As the main constituents of this interface are trophoblast (TR) and decidual cells (DC), the enzyme was estimated in isolated TR and DC to determine the extent of contribution by the respective cells. The enzyme level was estimated in cytosolic fraction, cell lysate and in conditioned media of these cells in rat and hamster. In both species the concentration of ADA was found to be markedly high in cytosolic fraction over to the cell lysate and the conditioned media in both TR and DC. Species-wise, it was higher in hamster. Cell-wise, the enzyme activity was significantly higher in TR than DC in rat but equal in hamster. In the conditioned medium, also, the enzyme activity was higher in TR in both species. The inference drawn from the results are: 1) the maximum enzyme activity in cytosolic fraction of TR and DC of both species clearly indicates equal involvement of the cells that constitute foeto maternal unit, 2) the enhanced level of enzyme in TR and DC of hamster over to those of rat is possibly due to the higher proliferative activity in the cells of this species because of shorter gestation (16-17 days in hamster and 22-23 days in rats). PMID- 10855704 TI - The fine structure of the axostyle and its associations with organelles in Trichomonads. AB - The fine structure of the axostyle in the protists Tritrichomonas foetus and Monocercomonas sp is described using transmission electron microscopy after quick freezing techniques and immunocytochemistry. The axostyle microtubules presents a lateral projection formed by two protofilaments in addition to the 13 protofilaments normally found in microtubules. The axostyle is associated with other cell structures such as hydrogenosomes, endoplasmic reticulum, sigmoid filaments and glycogen particles. The microtubules of the pelta-axostylar system are connected to each other by bridges regularly spaced with an interval of 9 nm. Labeling of the axostyle was observed after cell incubation with monoclonal antibodies recognizing alpha-tubulin and acetylated-tubulin. PMID- 10855705 TI - Morphogenesis of the giant sperm axoneme in Asphondylia ruebsaameni Kertesz (Diptera, Cecidomyiidae). AB - The formation of the sperm giant axoneme of the gall-midge fly Asphondylia ruebsaameni is described here. The axoneme consists of a great number of microtubular doublets (up to 2,500) arranged in a double spiral wrapping around an axial cluster of mitochondria. Each microtubular doublet is provided with an outer arm only. In the early spermatid the occurrence of a large system of curved multi-layered filamentous material associated with membranous cisternae has been observed in the perinuclear region. Such a system extends throughout the cytoplasm to contact the plasma membrane. The filamentous material appears to act as a nucleating centre for the assembly of the microtubular doublets, which initially have a submembranous location and later are distributed in the interior of the cell. After their assembly, microtubular doublets are associated pairwise and are arranged in a single microtubular row with a zig-zag configuration. This configuration changes during spermiogenesis as a consequence both of a rotation of the microtubular doublet pairs and a compaction of the axonemal complex due to the elimination of the excess cytoplasm. As a result of this process, a double parallel spiral of microtubular doublets is formed. PMID- 10855706 TI - Impairment of osteocalcin production in senescent periodontal ligament fibroblasts. AB - Osteocalcin production of senescent periodontal ligament fibroblasts (PDLF) with the expression of senescence-associated beta-galactosidase (SA-beta-Gal) was investigated on clones from 50-80 years old donors (n=20) with teeth extracted due to periodontitis and dental caries, and from 15-19 year old donors (n=20) with normal teeth extracted for orthodontic reasons. Immunohistochemically, the nonsenescent PDLF in all cultures in passage 2 showed strong reactivity with anti osteocalcin. The reactive intensity of PDLF (passage 2, PD 3.0) was significantly stronger in 50-80 year old donor group than in 15-19 year old donor group, suggesting that osteocalcin production of PDLF cultured in early passage is larger in cells from adult population than in cells from adolescent population. In PDLF cultures in passage 2 from 50-80 year old donor, two types of senescent cells were found: one with strong reactivity to anti-osteocalcin and the other with little detectable reactivity. The culture consisted of senescent PDLF (passage 8, PD 14.8) did not include cells which have a detectable reactivity with anti-osteocalcin immunohistochemically and the reactive intensity was significantly weaker in the senescent culture than in the culture in passage 2 by ELISA. This suggests that the production potential of osteocalcin is impaired in PDLF with aging in culture. Further, the reactive intensity with anti-osteocalcin of PDLF in passage 2 deprived of serum for 48 h was 6% of that of cells cultured with serum and the reaction increased after serum stimulation, suggesting that the osteocalcin production in PDLF in early passage is implicated in mitogenic stimulation. PMID- 10855707 TI - Potential commercial applications of microbial surfactants. AB - Surfactants are surface-active compounds capable of reducing surface and interfacial tension at the interfaces between liquids, solids and gases, thereby allowing them to mix or disperse readily as emulsions in water or other liquids. The enormous market demand for surfactants is currently met by numerous synthetic, mainly petroleum-based, chemical surfactants. These compounds are usually toxic to the environment and non-biodegradable. They may bio-accumulate and their production, processes and by-products can be environmentally hazardous. Tightening environmental regulations and increasing awareness for the need to protect the ecosystem have effectively resulted in an increasing interest in biosurfactants as possible alternatives to chemical surfactants. Biosurfactants are amphiphilic compounds of microbial origin with considerable potential in commercial applications within various industries. They have advantages over their chemical counterparts in biodegradability and effectiveness at extreme temperature or pH and in having lower toxicity. Biosurfactants are beginning to acquire a status as potential performance-effective molecules in various fields. At present biosurfactants are mainly used in studies on enhanced oil recovery and hydrocarbon bioremediation. The solubilization and emulsification of toxic chemicals by biosurfactants have also been reported. Biosurfactants also have potential applications in agriculture, cosmetics, pharmaceuticals, detergents, personal care products, food processing, textile manufacturing, laundry supplies, metal treatment and processing, pulp and paper processing and paint industries. Their uses and potential commercial applications in these fields are reviewed. PMID- 10855708 TI - Three biotechnical processes using Ashbya gossypii, Candida famata, or Bacillus subtilis compete with chemical riboflavin production. AB - Chemical riboflavin production, successfully used for decades, is in the course of being replaced by microbial processes. These promise to save half the costs, reduce waste and energy requirements, and use renewable resources like sugar or plant oil. Three microorganisms are currently in use for industrial riboflavin production. The hemiascomycetes Ashbya gossypii, a filamentous fungus, and Candida famata, a yeast, are naturally occurring overproducers of this vitamin. To obtain riboflavin production with the gram-positive bacterium Bacillus subtilis requires at least the deregulation of purine synthesis and a mutation in a flavokinase/FAD-synthetase. It is common to all three organisms that riboflavin production is recognizable by the yellow color of the colonies. This is an important tool for the screening of improved mutants. Antimetabolites like itaconate, which inhibits the isocitrate lyase in A. gossypii, tubercidin, which inhibits purine biosynthesis in C. famata, or roseoflavin, a structural analog of riboflavin used for B. subtilis, have been applied successfully for mutant selections. The production of riboflavin by the two fungi seems to be limited by precursor supply, as was concluded from feeding and gene-overexpression experiments. Although flux studies in B. subtilis revealed an increase both in maintenance metabolism and in the oxidative part of the pentose phosphate pathway, the major limitation there seems to be the riboflavin pathway. Multiple copies of the rib genes and promoter replacements are necessary to achieve competitive productivity. PMID- 10855709 TI - Response of the thermophile Thermus sp. RQ-1 to hyperbaric air in batch and fed batch cultivation. AB - The effects of increased air pressure in a culture of the thermophilic microorganism Thermus sp. RQ-1 were investigated. Cell growth dependence on oxygen supply was investigated in a fermenter at atmospheric pressure. Total oxygen depletion from the medium for low values of kLa was observed during the exponential growth phase. It was possible with this strain to enhance the oxygen transfer rate by increasing the air pressure. Cell productivity was improved by pressurisation up to 0.56 MPa for batch cultivation; and an induction of the antioxidant enzymes, superoxide dismutase and catalase, was observed with the rise in pressure. Cell pre-cultivation under pressurised conditions conferred to the cells more resistance to an exposure to hydrogen peroxide and more sensitivity to paraquat (methyl viologen). The usefulness of bioreactor pressurisation on the cultivation of Thermus sp. RQ-1 was demonstrated for fed batch operation, with the attainment of higher cell densities. A two-fold increase in cell mass productivity was obtained by the use of hyperbaric air (0.5 MPa). With the pressurisation of the head-space in the reactor, it was also possible to eliminate the loss of liquid by evaporation, which amounted to more than 10% at 70 degrees C and atmospheric pressure. PMID- 10855710 TI - Biosynthesis of citric acid by Yarrowia lipolytica repeat-batch culture on ethanol. AB - After analysis of batch culture and identification of the ways for prolongation of citric acid active synthesis by yeast, repeat-batch (RB) cultivation was suggested. Yarrowia lipolytica strain RB cultivation was studied and optimal conditions for cultivation selected. It was shown that when applying RB cultivation, better results were obtained than for batch cultivation. The activity of the culture remained stable after cultivation for more than 700 h. Comparative analysis of enzyme activities confirmed the regularity of the effect described, as the activity of practically of all the enzymes participating in ethanol oxidation and citric acid biosynthesis remained stable over time during RB cultivation. Advantages of RB cultivation for the production of citric acid by yeast are discussed. PMID- 10855711 TI - Optimization of culture medium for the continuous cultivation of the microalga Haematococcus pluvialis. AB - The freshwater microalga Haematococcus pluvialis is one of the best microbial sources of the carotenoid astaxanthin, but this microalga shows low growth rates and low final cell densities when cultured with traditional media. A single variable optimization strategy was applied to 18 components of the culture media in order to maximize the productivity of vegetative cells of H. pluvialis in semicontinuous culture. The steady-state cell density obtained with the optimized culture medium at a daily volume exchange of 20% was 3.77 x 10(5) cells ml(-1), three times higher than the cell density obtained with Bold basal medium and with the initial formulation. The formulation of the optimal Haematococcus medium (OHM) is (in g l(-1)) KNO3 0.41, Na2HPO4 0.03, MgSO4 x 7H2O 0.246, CaCl2 x 2H2O 0.11, (in mg l(-1)) Fe(III)citrate x H2O 2.62, CoCl2 x 6H2O 0.011, CuSO4 x 5H2O 0.012, Cr2O3 0.075, MnCl2 x 4H2O 0.98, Na2MoO4 x 2H2O 0.12, SeO2 0.005 and (in microg l(-1)]) biotin 25, thiamine 17.5 and B12 15. Vanadium, iodine, boron and zinc were demonstrated to be non-essential for the growth of H. pluvialis. Higher steady-state cell densities were obtained by a three-fold increase of all nutrient concentrations but a high nitrate concentration remained in the culture medium under such conditions. The high cell productivities obtained with the new optimized medium can serve as a basis for the development of a two-stage technology for the production of astaxanthin from H. pluvialis. PMID- 10855712 TI - On-line monitoring of growth of Escherichia coli in batch cultures by bioluminescence. AB - Bioluminescence was used to monitor growth of Escherichia coli in batch cultures on-line. Light emission of a strain engineered for constitutive bioluminescence was monitored with a simple set-up consisting of a photodiode, a photodetector amplifier and a recorder. Bioluminescence and colony forming units (CFU) of the cultures increased and decreased proportionally and were correlated during every growth phase at temperatures between 28 degrees C and 40 degrees C. Up to the late log (deceleration) phase, both light emission and CFU increased rapidly. Beyond the stationary phase these characteristics decreased very slowly at lower temperatures, while at higher ones they declined more rapidly. Towards the end of the cultivation, light emission of the cultures dropped to undetectable levels, even though CFU were recovered. This was particularly marked at lower temperatures where non-luminescent cultures retained very high CFU. This indicates that the actual metabolism of cells in a culture can be at a very low level or completely shut down, yet cells retain their capability to be culturable. The on-line technology described here has a number of potential uses in the laboratory and industry. PMID- 10855713 TI - Suspended rice particles for cultivation of Monascus purpureus in a tower-type bioreactor. AB - Cultivation of Monascus purpureus (CCRC 31615) for the production of natural pigments was investigated. Traditionally, Monascus species were grown on rice by solid-state culture. For large-scale cultivation, solid-state cultures were associated with some problems such as contamination and scale-up. By using submerged cultures with rice particles, a stirred-tank fermentor was not suitable for submerged cultures as the impeller tended to break the particles into small pieces. A conventional bubble column was also unsuitable as its mixing capability was poor. In the present study, a modified bubble column with wire-mesh draft tubes was employed for the cultivation of M. purpureus. The proposed column had a shorter mixing time and a higher oxygen transfer rate relative to the conventional bubble column. The production of pigments using the proposed column was up to 80% higher than that achieved using the conventional bubble column. PMID- 10855714 TI - Exploitation of butyrate kinase and phosphotransbutyrylase from Clostridium acetobutylicum for the in vitro biosynthesis of poly(hydroxyalkanoic acid). AB - Active butyrate kinase (Buk) and phosphotransbutyrylase (Ptb) were purified in three steps: ammonium sulfate precipitation, hydrophobic chromatography on phenyl Sepharose and affinity chromatography on Matrex Red A from recombinant Escherichia coli K2006 (pJC7). They were then successfully exploited for in vitro synthesis of 3-hydroxybutyryl-CoA (3HBCoA), 4-hydroxybutyryl-CoA (4HBCoA), 4 hydroxyvaleryl-CoA (4HVCoA) and poly(hydroxyalkanoic acid) (PHA). In addition, the ability of the PHA synthase of Chromatium vinosum, PhaEC(Cv), to use these CoA thioesters was evaluated. Combination of Buk and Ptb with PhaEC(Cv) established a new system for in vitro synthesis of poly(3-hydroxybutyric acid) [poly(3HB)]. In this system, 3-hydroxybutyric acid was converted to 3HBCoA by Buk and Ptb at the expense of ATP. Formation of 3HBCoA was further driven by the polymerization of 3HBCoA molecules to poly(3HB) by PHA synthase, and the released CoA was recycled by Ptb. This system therefore also ensured the regeneration of CoA. With ATP as the energy supply, which was hydrolyzed to ADP and phosphate, 2.6 mg poly(3HB) was obtained from a 1-ml reaction mixture containing 7.6 mg 3 hydroxybutyrate at the beginning. Studies showed that Ptb and PHA synthase were the rate-limiting steps in this system, and initial CoA concentrations ranging from 1 to 7 mM did not inhibit poly(3HB) synthesis. Synthesis of various polyesters of 3HB and 4HB with this system was also tested, and copolyesters containing 4HB of 1-46 mol % were obtained. PMID- 10855715 TI - A simple and rapid method to gain high amounts of manganese peroxidase with immobilized mycelium of the agaric white-rot fungus Clitocybula dusenii. AB - The agaric basidiomycete Clitocybula dusenii was used for the production of the extracellular ligninolytic enzyme, manganese (Mn) peroxidase. An immobilization technique is described using cellulose and polypropylene as carrier for the fungal mycelium. High amounts of Mn peroxidase were obtained with agitated cultures of immobilized fungus (up to 3,000 U l(-1)) while the biomass was recovered and used for further production cycles. Purification of Mn peroxidase revealed the existence of two forms: MnP1 (molecular mass 43 kDa, pI 4.5) and MnP2 (42 kDa, pI 3.8). PMID- 10855716 TI - Development of DNA delivery system using Pseudomonas exotoxin A and a DNA binding region of human DNA topoisomerase I. AB - Gene therapy is defined as the delivery of a functional gene for expression in somatic tissues with the intent to cure a disease. Thus, highly efficient gene transfer is essential for gene therapy. Receptor-mediated gene delivery can offer high efficiency in gene transfer, but several technical difficulties need to be solved. In this study, we first examined the DNA binding regions of the human DNA topoisomerase I (Topo I), using agarose gel mobility shift assay, in order to identify sites of noncovalent binding of human DNA Topo I to plasmid DNA. We identified four DNA binding regions in human DNA Topo I. They resided in aa 51 200, 271-375, 422-596, and 651-696 of the human DNA Topo I. We then used one of the four regions as a DNA binding protein fragment in the construction of a DNA delivery vehicle. Based on the known functional property of each Pseudomonas exotoxin A (PE) domain and human DNA Topo I, we fused the receptor binding and membrane translocation domains of PE with a highly positively charged DNA binding region of the N-terminal 198 amino acid residues of human DNA Topo I. The resulting recombinant protein was examined for DNA binding in vitro and transfer efficiency in cultured cells. The results show that this DNA delivery protein is a general DNA delivery vehicle without DNA sequence, topology, and cell-type specificity. The DNA delivery protein could be used to target genes of interest into cells for genetic and biochemical studies. Therefore, this technique can potentially be applied to cancer gene therapy. PMID- 10855717 TI - Overexpression of the OLE1 gene enhances ethanol fermentation by Saccharomyces cerevisiae. AB - The fermentation characteristics of Saccharomyces cerevisiae strains which overexpress a constitutive OLE1 gene were studied to clarify the relationship between the fatty acid composition of this yeast and its ethanol productivity. The growth yield and ethanol productivity of these strains in the medium containing 15% dextrose at 10 degrees C were greater than those of the control strains under both aerobic and anaerobic conditions but this difference was not observed under other culture conditions. During repeated-batch fermentation, moreover, the growth yield and ethanol productivity of the wild-type S. cerevisiae increased gradually and then were similar to those of the OLE1 overexpressing transformant in the last batch fermentation. However, the unsaturated fatty acid content (77.6%) of the wild-type cells was lower than that (86.2%) of the OLE1-recombinant cells. These results suggested that other phenomena caused by the overexpression of the OLE1 gene, rather than high unsaturated fatty acid content, are essential to ethanol fermentation by this yeast. PMID- 10855718 TI - Proteolytic stability of recombinant human serum albumin secreted in the yeast Saccharomyces cerevisiae. AB - In order to direct the persistent expression of recombinant human serum albumin (HSA) from the GAL10 promoter in the yeast Saccharomyces cerevisiae, we carried out periodic feeding of galactose during shake-flask cultures. Unexpectedly, the recombinant protein secreted was observed to undergo rapid degradation, which was apparently accelerated by carbon-source feeding. The extracellular degradation of HSA occurred even in the strain deficient in the major vacuolar proteases PrA and PrB, and in the strain lacking the acidic protease Yap3p (involved in the generation of HSA-truncated fragments). Interestingly, the degradation correlated closely with the acidification of extracellular pH and thus was significantly overcome either by buffering the culture medium above pH 5.0 or by adding amino acid-rich supplements to the culture medium, which could prevent the acidification of medium pH during cultivation. Addition of arginine or ammonium salt also substantially minimized the degradation of HSA, even without buffering. The extracellular degradation activity was not detected in the cell-free culture supernatant but was found to be associated with intact cells. The results of the present study strongly suggest that the HSA secreted in S. cerevisiae is highly susceptible to the pH-dependent proteolysis mediated by cell-bound protease(s) whose activity and expression are greatly affected by the composition of the medium. PMID- 10855719 TI - Genes encoding cytochrome P450 and monooxygenase enzymes define one end of the aflatoxin pathway gene cluster in Aspergillus parasiticus. AB - The identification of overlapping cosmids resulted in the discovery of the aflatoxin biosynthetic pathway gene cluster in Aspergillus flavus and A. parasiticus. This finding led to the cloning and characterization of one regulatory and 16 structural genes involved in aflatoxin biosynthesis, including the most recent report on the gene, ordA, which has been identified to be involved in the formation of four aflatoxins (B1, B2, G1 and G2). However, these genes do not account for all the identified chemical/biochemical steps in aflatoxin synthesis and efforts are underway to identify the genes controlling the other steps. We are also attempting to define the outer boundaries of the aflatoxin pathway gene cluster in the Aspergillus genome. For this goal, we extended sequencing in both directions from the existing (60 kb) aflatoxin pathway gene cluster, beyond the pksA gene at one end and the omtA gene at the other. Within the 25-kb genomic DNA sequence determined at the omtA end of the cluster, several new gene sequences were identified. The recently reported genes, vbs and ordA, were found within this 25-kb region. Two additional genes were also found in this region, a cytochrome P450 monooxygenase encoding gene, tentatively named cypX, and a monooxygenase encoding gene, tentatively named moxY, and these are also reported in this study. The sequence beyond these genes showed a 5-kb non-coding region of DNA followed by the presence of a cluster of genes probably involved in sugar metabolism. Northern blot analysis and reverse transcriptase polymerase chain reaction (RT-PCR) studies demonstrated that the genes, cypX and moxY, are expressed concurrently with genes involved in aflatoxin biosynthesis. Therefore, the two putative aflatoxin pathway genes cypX and moxY followed by a 5 kb non-coding region of DNA define one end of the boundary of the aflatoxin pathway gene cluster in A. parasiticus. PMID- 10855720 TI - Isolation of a delta7-cholesterol desaturase from Tetrahymena thermophila. AB - Cell-free preparations of Tetrahymena thermophila catalyze the direct desaturation of cholesterol to delta7-dehydrocholesterol (provitamin D3). The activity was isolated in the microsomal fraction from Tetrahymena homogenates. Delta7-desaturase activity was stimulated fivefold by the addition of 6 mM ATP. Other cofactors assayed, including NAD, NADP, NADH or NADPH, had no significant effect. The activity was found in microsomes prepared from stationary-phase cultures of the ciliate, grown either with or without added cholesterol, thus indicating that it is constitutively expressed in T. thermophila cells. PMID- 10855721 TI - Purification and characterization of isoamyl acetate-hydrolyzing esterase encoded by the IAH1 gene of Saccharomyces cerevisiae from a recombinant Escherichia coli. AB - The IAH1 gene of Saccharomyces cerevisiae encodes an esterase that preferentially acts on isoamyl acetate; however, the enzyme has not yet been completely purified from the yeast S. cerevisiae. We constructed the IAH1 gene expression system in Escherichia coli, and purified the IAH1 gene product (Iah1p). The amount of Iah1p produced by recombinant E. coli was more than 40% of total cellular proteins. The molecular size of Iah1p was 28 kDa by SDS-polyacrylamide gel electrophoresis. Judging from the molecular weight estimation by gel filtration of purified Iah1p, the enzyme was thought to be a homodimer. The Km values for isoamyl acetate and isobutyl acetate were 40.3 mM and 15.3 mM, respectively. The enzyme activity was inhibited by Hg2+, p-chloromercuribenzoate, and diisopropylfluorophosphate. PMID- 10855722 TI - Biotransformation of methyl ent-17-hydroxy-16beta-kauran-19-oate by Rhizopus stolonifer. AB - Methyl ent-17-hydroxy-16beta-kauran-19-oate was fed to a 2-day-old culture of the fungus Rhizopus stolonifer, fermenting at room temperature (25 degrees C) in an orbital shaker (21). After 11 days, both broth and mycelia were extracted with ethyl acetate. Two novel compounds were isolated from this experiment: methyl ent 9alpha,17-dihydroxy-16beta-kauran-19-oate and methyl ent-7alpha,17-dihydroxy 16beta-kauran-19-oate. Their structures were fully confirmed by spectroscopic methods. PMID- 10855723 TI - Molecular and physiological aspects of aflatoxin and sterigmatocystin biosynthesis by Aspergillus tamarii and A. ochraceoroseus. AB - Until recently, only three species (Aspergillus flavus, A. parasiticus and A. nomius) have been widely recognized as producers of aflatoxin. In this study we examine aflatoxin production by two other species, A. tamarii and A. ochraceoroseus, the latter of which also produces sterigmatocystin. Toxin producing strains of A. tamarii and A. ochraceoroseus were examined morphologically, and toxin production was assayed on different media at different pH levels using thin layer chromatography and a densitometer. Genomic DNA of these two species was probed with known aflatoxin and sterigmatocystin biosynthesis genes from A. flavus, A. parasiticus and A. nidulans. Under the high stringency conditions, A. tamarii DNA hybridized to all four of the A. flavus and A. parasiticus gene probes, indicating strong similarities in the biosynthetic pathway genes of these three species. The A. ochraceoroseus DNA hybridized weakly to the A. flavus and A. parasiticus verB gene probe, and to two of the three A. nidulans probes. These data indicate that, at the DNA level, the aflatoxin and sterigmatocystin biosynthetic pathway genes for A. ochraceoroseus are somewhat different from known pathway genes. PMID- 10855724 TI - Study on the production of 6-pentyl-alpha-pyrone using two methods of fermentation. AB - The lactone 6-pentyl-alpha-pyrone has a characteristic coconut aroma and is produced by Trichoderma species. A study on the fermentative production of 6 pentyl-alpha-pyrone in both surface and submerged conditions by Trichoderma harzianum was carried out. Maximum concentrations of 455 mg/l and 167 mg/l after 96 h and 48 h of fermentation in surface and submerged conditions, respectively, were obtained without using any additional recovery operations. The resultant yields are higher than those previously reported in the literature, which may be attributable to strain characteristics in combination with the choice of fermentation conditions employed in the present study. Enough scope exists for further improvement in the yields by optimizing the cultural and nutritional parameters. PMID- 10855725 TI - Biochemical mediator demand--a novel rapid alternative for measuring biochemical oxygen demand. AB - The biochemical oxygen demand (BOD) test (BOD5) is a crucial environmental index for monitoring organic pollutants in waste water but is limited by the 5-day requirement for completing the test. We have optimised a rapid microbial technique for measuring the BOD of a standard BOD5 substrate (150 mg glucose/l, 150 mg glutamic acid/l) by quantifying an equivalent biochemical mediator demand in the absence of oxygen. Elevated concentrations of Escherichia coli were incubated with an excess of redox mediator, potassium hexacyanoferrate(III), and a known substrate for 1 h at 37 degrees C without oxygen. The addition of substrate increased the respiratory activity of the microorganisms and the accumulation of reduced mediator; the mediator was subsequently re-oxidised at a working electrode generating a current quantifiable by a coulometric transducer. Catabolic conversion efficiencies exceeding 75% were observed for the oxidation of the standard substrate. The inclusion of a mediator allowed a higher co substrate concentration compared to oxygen and substantially reduced the incubation time from 5 days to 1 h. The technique replicates the traditional BOD5 method, except that a mediator is substituted for oxygen, and we aim to apply the principle to measure the BOD of real waste streams in future work. PMID- 10855727 TI - Blood pressure @ Internet-www.bloodpressure.nu. PMID- 10855726 TI - Biological degradation of selected hydrocarbons in an old PAH/creosote contaminated soil from a gas work site. AB - An old PAH/creosote contaminated soil (total approximately 300 microg PAH/g soil) from a former gas work site in Stockholm, Sweden, has been treated at 20 degrees C with the addition of various nutrients and inoculated with bacteria (isolated from the soil) to enhance the degradation of selected hydrocarbons. Microcosm studies showed that the soil consisted of two contaminant fractions: one available, easily degraded fraction and a strongly sorbed, recalcitrant one. The bioavailable fraction, monitored by headspace solid phase microextraction, contained aromatics with up to three rings, and these were degraded within 20 days down to non-detectable levels (ng PAH/g soil) by both the indigenous bacteria and the externally inoculated samples. The nutrient additives were: a minimal medium (Bushnell-Haas), nitrate, nitrite, potting soil (Anglamark, Sweden), sterile water and aeration with Bushnell-Haas medium. After 30 days treatment most of the sorbed fractions were still present in the soil. Stirring or mechanical mixing of the soil slurries had the greatest effect on degradation, indicating that the substances were too strongly sorbed for the microorganisms. When stirring the choice of nutrient seemed less important. For the non-stirred samples the addition of nitrate with the bacterial inoculum showed the best degradation, compared to the other non-stirred samples. At the end of the experiments, accumulations of metabolites/degradation products, such as 9H fluorenone, 4-hydroxy-9H-fluorenone, 9,10-phenanthrenedione and 4H cyclopenta[def]phenanthrenone were detected. The metabolite 4-hydroxy-9H fluorenone increased by several orders of magnitude during the biological treatments. Microbial activity in the soil was measured by oxygen consumption and carbon dioxide production. PMID- 10855728 TI - Hypertension and the metabolic syndrome: closely related central origin? AB - In primary hypertension a mild hyperresponsiveness of hypothalamic, sympatho hormonal centres to psychosocial stimuli forms a major pathogenetic element, although high salt intake in some subjects may contribute via volume expansion. Hypertension is often associated with another "civilisation" disorder, the metabolic syndrome, defined as abdominal obesity, insulin resistance and dyslipidaemia. According to recent research, the metabolic syndrome has in all likelihood a central neuroendocrine origin in the form of enhanced engagement of the hypothalamic-pituitary-adrenal (HPA) axis. Here the peripheral endocrine perturbations act as triggers for both central obesity and the metabolic abnormalities. The reaction pattern characterising early primary hypertension is identical with, or closely related to, the "defence reaction", while that leading to the metabolic syndrome is similar to that of the "defeat reaction". Both belong to the primitive survival reactions, common to all mammals, though man can control, or at least mask, his outward-behavioural part but not the neuro hormonal expressions. Animal experiments show how frequent or chronic mental challenges are capable of engaging these limbic-hypothalamic centres, affecting blood pressure regulation as well as endocrine-metabolic regulation. Furthermore, these centres are tightly coupled functionally, and their signals to the periphery often combined. On a long-term basis their engagements appear to be decisive for the development of both primary hypertension and the metabolic syndrome, as suggested by intervention studies. In both these "disorders of civilisation", observations strongly indicate that psychosocial stress, socioeconomic handicaps, lack of exercise, abuse and also psychiatric traits are involved. Such factors, characteristic of current competitive society, probably cause mixed engagements of the two above-mentioned neuro-hormonal patterns, and thereby, with time, primary hypertension and the metabolic syndrome, with end points such as coronary artery disease, diabetes mellitus type2 and stroke. Susceptibility to such developments is probably enhanced by genetic factors. This overview of recent developments therefore serves to emphasise how both primary hypertension and the metabolic syndrome seem to have a common central origin. Central regulatory factors are often overlooked, partly because it is not realised that limbic-hypothalamic centres are the major regulators of both circulatory and metabolic events, and partly because of the long period of time required before these disease end-points are reached. PMID- 10855729 TI - Blood viscosity, plasma adrenaline and fasting insulin in hypertensive patients with left ventricular hypertrophy. ICARUS, a LIFE Substudy. Insulin CARotids US Scandinavica. AB - We have seen relationships between whole blood viscosity (WBV) and components of the metabolic cardiovascular syndrome in borderline hypertensive young men and suggested that sympathetic nervous system (SNS) activity may be a mediator. In the present study we aimed to test this hypothesis in established hypertension and to investigate the relationship between WBV and cardiac dimensions. Unmedicated patients (n = 42) with stage II-III hypertension and electrocardiographic left ventricular hypertrophy (LVH) underwent hyperinsulinemic isoglycemic glucose clamp to assess glucose disposal rate (GDR) and echocardiographic studies. WBV, plasma catecholamines and insulin were measured in arterialized venous blood. WBV at high shear rate correlated with baseline plasma adrenaline (r = 0.33, p = 0.04) and fasting insulin (r = 0.34, p = 0.04) while there was a negative trend for GDR (r = -0.21, p = 0.2). WBV at low shear rate correlated with plasma adrenaline (r = 0.49, p = 0.002) and resting heart rate (r = 0.36, p = 0.02). WBV was higher in smokers than in non-smokers (p = 0.02) and in males than in females (p = 0.02). Fasting insulin independently explained 12% of the variation in WBV at high shear, while baseline adrenaline independently explained 17% of the variation in WBV at low shear. Systolic blood pressure explained 31% of the variation in LV mass index. Thus, we demonstrate positive relationships between blood viscosity versus plasma adrenaline and fasting insulin in hypertensive patients with LVH. We suggest that adrenergic activity may increase hematocrit and viscosity and hence reduce insulin sensitivity. PMID- 10855730 TI - Blood pressure level and relation to other cardiovascular risk factors in male hypertensive patients without clinical evidence of ischemic heart disease. AB - Arterial hypertension is accompanied by increased morbidity and mortality and constitutes a substantial part of medical care. Antihypertensive intervention reduces the cardiovascular morbidity and mortality. The aims of the study were to evaluate the relationship between cardiovascular risk factors and the blood pressure (BP), and to evaluate the percentage of patients who had achieved a BP level as recommended by the sixth report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC VI). BP was evaluated in relation to age, body mass index, duration of hypertension, cholesterol and triglyceride level, smoking status, information of regular exercise, a family history of ischemic heart disease (IHD) and drug treatment, in 220 men treated for arterial hypertension. In the univariate analyses we found a higher systolic blood pressure (SBP) with older age, higher SBP in smoking patients and lower SBP in patients with regular exercise. In a multivariate model age (p = 0.0004), smoking status (p = 0.01) and regular exercise (p= 0.06) were independently associated with SBP. There was a lower diastolic blood pressure (DBP) with older age, and age was independently associated with DBP. Office SBP was above 140 mmHg in 83% and above 160 mmHg in 44% of patients. During ambulatory blood pressure monitoring (AMBP), SBP was above 135 mmHg in 40% and above 155 mmHg in 15% of patients. In addition to male sex and hypertension there was a high percentage of other cardiovascular risk factors--43% was smoking, 21% had a family history of IHD, 77% had a se-cholesterol above 5.5 mmol/l and 48% had a se-triglyceride above 1.6 mmol/l. In a consecutive group of asymptomatic male treated hypertensive patients SBP is independently associated with age and smoking status, and DBP with age. A high percentage of the patients do not have a well controlled BP, and a high percentage have additional risk factors such as smoking, hypercholesterolaemia, hypertriglyceridaemia and a family history of IHD. This means that there is room for much improvement in the control of hypertension. PMID- 10855731 TI - More non-Q-wave myocardial infarctions but similar infarct sizes in patients with hypertension. AB - Of 350 consecutive patients without previous symptoms of coronary artery disease, admitted to hospital with an acute myocardial infarction, 109 of them (31%) reported a history of previous hypertension. Hypertensive patients were older than their normotensive counterparts, more of them were females, and thrombolytic treatment was administered to significantly fewer. Blood pressure values at admission to hospital were higher in hypertensive patients; this difference was significant in hypertensive males. Altogether 44 out of 49 female (90%) and 42 out of 60 male hypertensive patients (70%) reported using antihypertensive medication. A previous history of hypertension did not change infarct size as assessed by peak enzyme levels, neither in the bivariate nor in the multivariate analysis. In contrast to this, the adjusted odds ratio for developing a non-Q wave infarct was 2.51 (p=0.003), i.e. the chance of developing a non-Q-wave infarct in hypertensives was increased by 151%. Thus, in spite of similar infarct size in normotensive and hypertensive patients, a relative smaller proportion of the probably hypertrophied left ventricular wall developed necrosis in the hypertensive population. The propensity towards non-Q-wave infarctions may contribute to the observed less use of fibrinolytic drug treatment in the presently observed patients with hypertension. PMID- 10855732 TI - Angiotensin-converting enzyme gene I/D polymorphism in malignant hypertension. AB - BACKGROUND: The mechanism of the rapid transition of a stable benign hypertensive disease to a severe and devastating malignant hypertension is not fully understood. However, the renin angiotensin system, which is highly activated in malignant hypertension, is established as an important pathogenetic factor in different cardiovascular and renal diseases. Over the last decade, a polymorphism in genes regulating this system has been found. This includes the 287 bp sequence deletion (D)/insertion (I) polymorphism in the angiotensin-converting enzyme (ACE) gene and the methionine (M) to threonine (T) point mutation polymorphism in the angiotensinogen (AGT) gene. These gene polymorphisms have been associated with various cardiovascular and renal diseases and the aim of this study was to investigate whether they were linked to malignant hypertension. METHODS: Forty two patients with malignant hypertension (mean age 55 years), 42 patients with non-malignant hypertension (mean age 57 years) and 85 normotensive control subjects (mean age 42 years) were investigated with respect to ACE I/D and AGT M/T genotypes. DNA was prepared by standard methods from isolated white blood cells and analysed by the PCR technique. The PCR reaction used in the detection of the ACE I/D polymorphism was optimized for an equal amplification of the I and D alleles. RESULTS: The frequency of the DD genotype was significantly increased in patients with malignant hypertension (43%) compared with patients with non malignant hypertension (14%) and normotensive control subjects (18%) (p <0.01) for both. The frequency distribution of AGT M/T genotype did not differ between patients with malignant and non-malignant hypertension. CONCLUSION: The DD genotype of the ACE gene occurred more than twice as often in malignant hypertension than in non-malignant hypertension and indicates that ACE gene polymorphism is a significant risk factor for initiation of malignant hypertension. PMID- 10855733 TI - Ambulatory blood pressure and endothelium-dependent vasodilation in hypertensive patients. AB - Endothelial function is important for local vascular regulation and an abnormal endothelium-dependent vasodilatation (EDV) has been observed in subjects with essential hypertension. As ambulatory blood pressure (ABP) is more closely related to target organ damage than office blood pressure, this study investigated also if 24-h ABP is more closely related to an impaired EDV than office blood pressure recordings. In a group of 25 untreated patients with essential hypertension and an age- and sex-matched control group (n = 21) endothelial function was evaluated by measurements of forearm blood flow (FBF) during local intra-arterial infusions of metacholine (evaluating EDV) and sodium nitroprusside (evaluating endothelium independent vasodilation, EIDV). FBF was measured with venous occlusion plethysmography. Both office mean artery pressure (MAP; r= -0.57, p < 0.001) and 24-h ABP (r = 0.40, p < 0.01) were related to the endothelial vasodilator function (EDV to EIDV ratio) in an inverse way, but ABP was not superior to office blood pressure recordings. Within the hypertensive group, pronounced white-coat effect (office minus daytime ABP) was associated with a reduced,EDV (r= 0.41, p < 0.05). The degree of night-time decline in blood pressure ("dipping") showed no correlation to EDV. In conclusion, the finding that ABP was no more closely related to the endothelial vasodilator function than office blood pressure recordings might be due to an increased mental alertness affecting EDV in some hypertensive subjects, as suggested by the finding of a reduced EDV in those with a pronounced white-coat effect. PMID- 10855734 TI - Blood pressure changes in relation to sodium and calcium status in induced hyperinsulinemia. AB - Insulin increases renal sodium reabsorption which may contribute to hypertension. However, acute insulin administration may result in vasodilation. The aim of the present study was to investigate effects on blood pressure and alterations in ion status during hyperinsulinemia. Blood pressure and serum sodium and ionized calcium concentrations were measured before and at the end of euglycemic hyperinsulinemic clamp tests performed in 45 patients with essential hypertension. Both the systolic and the diastolic blood pressure decreased, by 4% (p < 0.05) and 3% (p < 0.05), respectively. Circulating ionized calcium concentration increased by 2% (p < 0.001), and the ratio between circulating sodium and ionized calcium concentrations decreased. The changes in circulating sodium concentration correlated to changes in systolic blood pressure (SBP; r = 0.36, p = 0.05). Both ionized calcium concentrations and the ratio between circulating sodium and ionized calcium concentrations correlated to changes in SBP during hyperinsulinemia (r = -0.41, p = 0.03, r = 0.56, p < 0.01, respectively). The changes in ion status were not significantly correlated to age, body mass index or insulin sensitivity. In conclusion, a more pronounced increase in circulating ionized calcium concentration and reduction in the ratio between sodium and ionized calcium concentrations was associated with a greater blood pressure decline during the hyperinsulinemic clamp test when performed in hypertensive patients. PMID- 10855735 TI - Assessment of insulin sensitivity by 90 min isoglycaemic hyperinsulinaemic glucose clamp in healthy young men. AB - We aimed to perform a detailed analysis of the isoglycaemic hyperinsulinaemic glucose clamp in relation to the time spent in performing the procedure, and analysed two series performed by independent investigators on different groups (n = 19 and n = 28) of healthy, young men. We calculated glucose disposal rates (GDR) during 20-min periods at different time points during the clamp. There was no difference in 90- and 120-min GDR when comparing the two series. The differences between 90- and 120-min GDR were (mean +/- SD) 0.48 +/- 1.10 mg/kg/min (p = 0.73) and 0.37 +/- 1.05 mg/kg/min (p = 0.71), respectively. The correlations between 90- and 120-min GDR were 0.94 (p < 0.001) and 0.89 (p < 0.001). Correlations between GDR during the second hour of the clamp and fasting plasma insulin ranged from -0.53 (p = 0.020) to -0.55 (p = 0.016) and from -0.44 (p = 0.020) to -0.54 (p = 0.003), respectively, and did not improve after 60 min of clamping. These data suggest that reliable indices of insulin sensitivity in healthy young men may appear even when the isoglycaemic hyperinsulinaemic clamp procedure is shortened from 120 to 90 min. A shorter procedure is time-effective and less expensive, but may be limited to healthy, young Caucasian men. PMID- 10855736 TI - Forearm blood flow responses to neuropeptide Y, noradrenaline and adenosine 5' triphosphate in hypertensive and normotensive subjects. AB - Neuropeptide Y (NPY), noradrenaline (NA) and adenosine 5'-triphosphate (ATP) are important co-transmitters in the sympathetic nervous system, which has a central role in cardiovascular control. In order to evaluate if hypertension is associated with alterations in vascular responses to sympathetic co-transmitters we studied the effects of intra-arterial infusion of NPY, NA and ATP on forearm blood flow. Blood flow was measured by venous occlusion plethysmography in six hypertensive (mean arterial blood pressure (MAP) 113 +/- 4 mmHg) and six matched normotensive subjects (MAP 97 +/- 3 mmHg). NPY and NA significantly reduced forearm blood flow, while a powerful increase was seen with ATP. Forearm vascular resistance, calculated as MAP divided by forearm blood flow, was significantly increased by NPY and NA and strongly reduced by ATP. There was no difference between hypertensive and normotensive subjects in response to either transmitter. In conclusion, vascular reactivity to intra-arterial administration of NPY, NA and ATP seems to be intact in hypertensive patients without metabolic aberrations. PMID- 10855737 TI - A possible link between endothelial dysfunction and insulin resistance in hypertension. A LIFE substudy. Losartan Intervention For Endpoint-Reduction in Hypertension. AB - BACKGROUND: We wanted to investigate whether insulin resistance and time to steady state during isoglycemic clamp were associated with endothelial dysfunction, peripheral vascular remodeling and forearm blood flow (FBF) in patients with longstanding hypertension. METHODS: In 43 unmedicated, hypertensive patients with electrocardiographic-defined left ventricular hypertrophy we performed a 2-h oral glucose tolerance test and a 3-h isoglycemic hyperinsulinemic clamp with measurements of circulating plasma epinephrine and FBF by plethysmography. Delayed steady state was assessed by measuring the increase in insulin sensitivity from the second to the third hour of clamping. We measured 24-h ambulatory blood pressure, minimal forearm vascular resistance (MFVR) by plethysmography, media:lumen ratio (MLR) and acetylcholine-induced relaxation (AIR) in isolated, subcutaneous resistance arteries by myography. RESULTS: Insulin sensitivity after 2 and 3 h of clamping was not related to maximal AIR, MLR, MFVR or FBF. The increase in insulin sensitivity in men was negatively correlated to maximal AIR (r = -0.36, p < 0.05), and was independently correlated to relative changes in FBF (beta = 0.46) and in circulating epinephrine (beta = 0.33; adj. R = 0.33, p < 0.001). CONCLUSIONS: Insulin sensitivity was not correlated to parameters of peripheral vascular remodeling, endothelial function or microvascular rarefaction in patients with longstanding hypertension and left ventricular hypertrophy. However, the action of insulin on peripheral glucose uptake was influenced by endothelial dysfunction (delayed transcapillary insulin transport) and by changes in and/or redistribution of blood flow suggesting a link between vascular function and insulin sensitivity. PMID- 10855738 TI - Efficacy and safety of a new long-acting drug combination, trandolapril/verapamil as compared to monotherapy in primary hypertension. Swedish TARKA trialists. AB - OBJECTIVE: To evaluate the clinical efficacy and safety of a new antihypertensive drug combination of trandolapril/verapamil compared to monotherapy with verapamil or trandolapril, in patients with mild to moderate primary hypertension. DESIGN: A multicentre, prospective, randomized, double-blind, controlled cross-over study with specific statistical considerations. SETTINGS: Eighteen primary health care centres and out-patient hospital clinics in Sweden. PATIENTS: Two hundred and twenty-six outpatients with uncomplicated primary hypertension with a baseline sitting diastolic blood pressure (BP) between 95 and 115 mmHg. INTERVENTIONS: After a 4-week placebo period, patients were randomized to treatment for 8 weeks with trandolapril/verapamil (2 mg/180 mg) or each drug alone (verapamil 240 mg, trandolapril 2 mg) for 8 weeks. MAIN OUTCOME MEASURES: Treatment responses (blood pressure (BP) fall and rate pressure product) to the three regimens with statistical comparison and also in relation to plasma concentrations of active renin (AR). Adverse events and safety were also evaluated. RESULTS: The mean BP fall was significantly greater with the combination (20/15 mmHg), p < 0.00054, as compared to both trandolapril (14/11 mmHg) or verapamil (13/11) mmHg. The difference between verapamil and trandolapril was not significant. Rate pressure product decreased significantly more on the combination, p < 0.001, than on trandolapril or verapamil alone. Treatment response to trandolapril was positively correlated to initial AR (r = 0.30-0.43). All treatments were well tolerated and safe. CONCLUSIONS: The new fixed drug combination trandolapril/verapamil was superior to monotherapy with either of these drugs alone regarding reduction of both BP and rate pressure product. This combination can be safely and effectively used for the treatment of mild to moderate primary hypertension. PMID- 10855739 TI - Study on COgnition and Prognosis in the Elderly (SCOPE): baseline characteristics. AB - The Study on COgnition and Prognosis in the Elderly (SCOPE) is a multi-centre, prospective, randomized, double-blind, parallel-group study. The primary objective of SCOPE is to assess the effect of the angiotensin II type 1 (AT1) receptor blocker, candesartan cilexetil 8-16 mg once daily, on major cardiovascular events in elderly patients (70-89 years of age) with mild hypertension (DBP 90-99 and/or SBP 160-179 mmHg). The secondary objectives of the study are to test the hypothesis that antihypertensive therapy can prevent cognitive decline (as measured by the Mini Mental State Examination, MMSE) and dementia, and to assess the effect of therapy on total mortality, myocardial infarction (MI), stroke, renal function, and hospitalization. A total of 4964 patients from 15 participating countries were recruited during the randomization phase of SCOPE, exceeding the target population of 4000. The mean age of the patients at enrolment was 76 years, the ratio of male to female patients was approximately 1:2, and 52% of patients were already being treated with an antihypertensive agent at enrolment. The majority of patients (88%) were educated to at least primary school level. At randomization, mean sitting blood pressure values were SBP 166 mmHg and DBP 90 mmHg, and the mean MMSE score was 28. Previous cardiovascular disease in the study population included myocardial infarction (4%), stroke (4%) and atrial fibrillation (4%). Men, more often than women, had a history of previous MI, stroke and atrial fibrillation. A greater percentage of men were smokers (13% vs 6% in women) and had attended university (11% vs 3% of women). Of the randomized patients, 21% were 80 years of age. In this age group smoking was less common (4% vs 10% for 70-79-year-olds) and fewer had attended university (4% vs 7% for 70-79-year-olds). The incidence of MI was similar in both age groups. However, stroke and atrial fibrillation had occurred approximately twice as frequently in the older patients. The patients' mean age at baseline was similar in the participating countries, and most countries showed the approximate 1:2 ratio for male to female patients. There was also little inter-country variation in terms of mean SBP, DBP or MMSE score. However, there was considerable regional variation in the percentage of patients on therapy prior to enrolment. PMID- 10855740 TI - Downregulation of adenosine and P2X receptor-mediated cardiovascular responses in heart failure rats. AB - Neurohormonal changes in congestive heart failure (CHF) include an enhanced peripheral sympathetic nerve activity which results in increased release of noradrenaline, neuropeptide Y and ATP. To examine if such changes in CHF would modulate peripheral pre- and postsynaptic receptors of ATP and its degradation product adenosine, experiments were performed in a rat model of ischaemic CHF. In this model, ischaemia was induced in rats by ligation of the left coronary artery. Our results demonstrate that there is a selective downregulation of P2X receptor-mediated pressor effects, while the hypotensive effects mediated by the endothelial P2Y receptors are unaffected in CHF. Moreover, the adenosine-mediated inhibitory effects on heart rate and blood pressure were also attenuated in the CHF rats. The most important changes in adenosine and P2-receptor function induced by ischaemic CHF were the reduced pressor effect mediated by the P2X receptor and the increased heart rate due to an attenuated inhibitory effect of adenosine. PMID- 10855741 TI - Development of chronic allograft rejection and arterial hypertension in Brown Norway rats after renal transplantation. AB - The cardiovascular and renal pathophysiology associated with chronic renal allograft rejection under triple drug immunosuppressive treatment was studied using a recently developed model (Brown Norway (BN) rats) in a 6-week experiment. Renal transplantation was performed to 10-week-old rats in a rat strain combination of Dark Agouti (DA) --> BN. The right kidney was removed from another group of BN rats (uninephrectomized). A triple drug treatment comprising cyclosporine (10 mg/kg subcutaneously, s.c.), azathioprine (2 mg/kg s.c.) and methylprednisolone (1.6 mg/kg s.c.) was given to each rat daily for 6 weeks. A control group underwent no operations nor drug treatment. After the transplantation, the systolic blood pressure in this group was increased from 116 +/- 2 to 166 +/- 2 mmHg, while in the uninephrectomized group the rise was from 115 +/- 4 to 146 +/- 4 mmHg, and no change was observed in the blood pressures of the control group. The vascular relaxation responses of mesenteric arterial rings in vitro to acetylcholine were inhibited in both the transplantation group and the uninephrectomized group as compared with the control group, but few significant differences were found in the contraction responses to noradrenaline and potassium chloride. Graft histology was examined after 6 weeks, quantified by using the chronic allograft damage index (CADI). Changes specific to a chronic rejection reaction were observed in the allografts (CADI mean 6.0) but no injuries were seen in the rats' own kidneys (CADI mean 1.2). Our findings show that allograft rejection in BN rats after renal transplantation is associated with the development of arterial hypertension. The combination of cyclosporine, methylprednisolone and azathioprine also rises blood pressure in uninephrectomized BN rats. The hypertensive effects of the drug treatment and graft rejection are associated with endothelial dysfunction. PMID- 10855742 TI - Regional renal haemodynamics of angiotensin II infusion under prostaglandin, kinin or converting enzyme inhibition in the Wistar rat. AB - AIMS: Renal medullary blood flow is important in blood pressure regulation and is surprisingly unaffected by the vasoconstrictor action of angiotensin II (Ang II). This study tested if the effect of Ang II on the renal papillary circulation is modulated by bradykinins, prostaglandins or NO (NO). In anaesthetised Wistar rats, total renal blood flow (RBF) was measured, as was cortical (CBF) and papillary (PBF) blood flow, using the laser-Doppler technique, in responses to Ang II (30 ng kg(-1) min(-1)) alone and after ACE inhibition (enalapril) or bradykinin/prostaglandin synthesis inhibition (ketoprofen, aprotinin). PBF was also measured after blockade of NO formation with or without pretreatment with an Ang II receptor antagonist (losartan). MAJOR FINDINGS: (i) PBF did not change in response to Ang II infusion but MAP increased (+ 10%) and RBF and CBF decreased. (ii) Treatment with aprotinin and ketoprofen left MAP, RBF and CBF unchanged but decreased PBF. Ang II did not decrease PBF further but a significant increase in MAP was seen. (iii) Enalapril treatment left PBF unchanged but decreased MAP and increased RBF and CBF. When Ang II was infused PBF and MAP increased markedly. (iv) L-NAME reduced PBF independently of losartan treatment. PRINCIPAL CONCLUSION: Bradykinin and prostaglandins do not appear to cause the lack of renal papillary vasoconstriction to Ang II. However, the increase in PBF to Ang II seen after enalapril treatment suggests that enalapril treatment, possibly via its effects on kinin breakdown and subsequent NO formation, might affect the sensitivity of renal papillary autoregulation. This may be an important aspect of the blood pressure lowering effect of ACE inhibitors. PMID- 10855743 TI - Osteoarthritis and traumatic arthritis of the upper quadrant. PMID- 10855744 TI - Osteoarthritis of the hand: a modifiable disease. AB - Osteoarthritis remains an exciting challenge in terms of an understanding of its pathophysiology, the development of disease-modifying drugs, and the search for more effective treatment for pain. With the aging of the general population, the impetus for research will increase, making the treatment of osteoarthritis, once considered an unalterable disease, an area of dynamic interest. PMID- 10855745 TI - Arthritis of the thumb basal joint complex. AB - Osteoarthritis at the base of the thumb is a common and extremely disabling condition that severely compromises function of the entire hand. Successful treatment is based on an understanding of the specific anatomy and the unique functional attributes of the human hand and thumb. Preservation of the web space is a priority in nonsurgical care and splinting as well as a principal goal of surgical reconstruction. Exercise regimens are designed to emphasize thenar strengthening to encourage preservation of the web space. Activity analysis and modification are focused on joint protection and the avoidance of positions that will accentuate the pathologic condition of trapeziometacarpal subluxation accompanying the retropulsion of the thumb that occurs with contracture of the web space. Surgical treatment is directed toward restoring the thumb-index web space and stabilizing the newly fabricated basal joint by reconstructing the beak ligament and providing a suitable interposition material. After-care likewise emphasizes restoration of the thumb web space, joint mobilization, and strengthening of the supporting thenar musculature. A well-integrated surgical and therapy team will produce uniformly good functional results in the treatment of this disabling condition at the base of the thumb that differentiates us from our simian ancestors. PMID- 10855746 TI - Osteoarthritis of the fingers. AB - Osteoarthritis of the fingers is an especially common condition in postmenopausal women. Many consider it a normal part of aging or a relatively minor disease. Osteoarthritis of the fingers is a disease process that destroys interphalangeal cartilage and results in pain, swelling, decreased finger motion, joint deformities, and difficulty performing activities that require grip or pinch. Medication, rest, and supportive splinting alternated with mobility and strengthening exercises, patient education, and incorporation of joint protection techniques may alleviate symptoms and improve functional abilities. Surgery is indicated when these methods have failed to control pain, instability, and deformities or improve ease of function. Joints may be fused (arthrodesis) or replaced (arthroplasty) to manage pain and produce finger joints capable of withstanding the force of normal use without pain or further joint damage. Postoperative therapy includes splinting to protect healing structures along with range-of-motion programs to maximize functional outcomes. PMID- 10855747 TI - Traumatic arthritis and osteoarthritis of the wrist. PMID- 10855748 TI - Osteoarthritis and traumatic arthritis of the elbow. PMID- 10855749 TI - Osteoarthritis and traumatic arthritis of the shoulder. AB - Glenohumeral osteoarthritis and traumatic arthritis result in a painful shoulder with impairments, functional deficits, and disability. Conservative treatment includes oral inflammatory medication, cortisone injection, or rehabilitation. Rehabilitation of the shoulder can be beneficial, but if joint destruction is advanced, surgery may be required. Postoperative rehabilitation requires the therapist to know the basics of the surgical technique so that safe and effective therapeutic intervention can be made. A successful outcome depends on effective communication and interaction among the physician, therapist, and patient. Each "team" member has a defined role in rehabilitation, and all three must fulfill their responsibilities for the desired outcome to be achieved. PMID- 10855750 TI - The degenerative cervical spine: pathogenesis and rehabilitation concepts. AB - The degenerative process associated with spondylosis in the cervical spine has been reviewed. The two compressive syndromes commonly associated with spondylosis, radiculopathy and myelopathy, are briefly reviewed. Except for more severe, multilevel degenerative changes producing neurologic compromise, correlation between degenerative changes and patient symptoms or functional limitations is generally poor. A conceptual scheme for guiding rehabilitation of mechanical neck pain, based on irritability level and the effects of mechanical stress on symptoms, is proposed. Further research is required to test the reliability and validity of categorization schemes like the one proposed. Such schemes based on history and effects of mechanical stresses, rather than solely on degenerative radiographic findings, are necessary to classify patients in meaningful ways that help guide specific rehabilitation strategies and tactics. When meaningful classification schemes exist, treatments matched with specific categories of dysfunction can be tested for effectiveness. PMID- 10855751 TI - Exercise for the patient with upper quadrant osteoarthritis. PMID- 10855752 TI - Community resources and assistive devices for people with arthritis. AB - Helping patients obtain support from outside sources--both community resources and assistive technologies--can be a very rewarding part of the practice of therapy. During rehabilitation, patients with chronic diseases, in particular, need to develop self-management skills that will help them improve their quality of life. PMID- 10855753 TI - Low use of analgesics in Alzheimer's disease: possible mechanisms. AB - This article discusses three possible mechanisms that might underlie the often observed low use of nonsteroidal anti-inflammatory drugs (NSAIDs) and other analgesics in patients with Alzheimer's disease (AD), compared with nondemented elderly: (a) AD patients are progressively less able to communicate about pain; (b) AD patients suffer from fewer painful conditions than nondemented elderly subjects; and (c) considering the neuropathology, AD patients might actually experience pain to a lesser extent. Suggestions for future pain assessment in AD are made. PMID- 10855754 TI - Alzheimer's: no pain or no complain? PMID- 10855755 TI - Is the (relatively) low use of analgesics in elderly AD patients a problem? PMID- 10855756 TI - Memory, pain, agitation, and dissociation. PMID- 10855757 TI - Residential caregiver attitudes toward seriously mentally ill persons. AB - Previous research has found that caregiver attitudes are associated with the course of illness of seriously mentally ill (SMI) persons. This study examined whether variation in caregiver attitudes could be accounted for by (a) staff caregivers and/or (b) SMI persons. Group home staff were asked to describe each SMI group home resident and to describe the relationship they had with each resident. We recorded the number of positive and negative statements made by each staff member about each resident's character, behaviors, and the interactions staff had with them. Overall, the variation in positive staff statements about group home residents was significantly accounted for by the residents. In general, the variation in negative staff statements was significantly accounted for by staff. However, the variation in negative staff statements about the character of residents was accounted for by both staff and residents. PMID- 10855758 TI - Affect and cognition in dreams: a critique of the cognitive role in adaptive dream functioning and support for associative models. AB - Dream affect and cognition are examined in a 4-month longitudinal study of depressed, recently divorced subjects. Several measures of cognition in dream reports were used to determine whether remitting subjects were more likely to utilize certain cognitive strategies than nonremitters. No differences were found in the cognition between those who were remitting and nonremitting subjects. Levels of cognition in dreaming were directly related to the emotional intensity of the dreams across all subjects. In subjects with higher depression scores (Beck Depression Inventory > 20), depression levels were inversely related to both dream affect and cognition. It is argued that cognition plays a secondary role in dream production. A modification of David Foulkes's model of dreaming is proposed, in which emotional intensity drives associative processes, which in turn require more elaborate cognitive devices to relate diverse activated mnemonic units. PMID- 10855759 TI - A conceptualization of the repetition compulsion. AB - The concept of the repetition compulsion remains an enigma. Its etiology is not fully understood and the purpose it serves continues to be a mystery. Although it is often theorized that the compulsion to repeat may function to facilitate mastery of a past trauma, mastery is rarely achieved. In this article the concept of the repetition compulsion is reviewed and the unanswered questions that continue to exist about this phenomenon are summarized. A way to conceptualize the compulsion to repeat is then offered. The compulsion to repeat as it specifically relates to the attempt to master a previous trauma is reviewed, followed by a examination of the relationship between the compulsion to repeat and reenactments. Finally, how the compulsion to repeat can be viewed as a posttraumatic stress response and the implications of understanding it in this fashion is then examined. PMID- 10855760 TI - Generalized anxiety disorder in adolescents and young adults with mild mental retardation. AB - This report examines clinical features of generalized anxiety disorder in adolescents and young adults with mild mental retardation (MR), compared with children and adolescents with normal IQ. Frequency of symptoms, comorbidity, agreement between reports of subjects and parents, correlation between IQ and severity of disorder, and comparison between frequency of symptoms in the experimental and control groups are described. Twenty-two subjects with MR (12 males and 10 females aged 11-25 years; mean age = 16.3), 30 children (19 males and 11 females aged 7-11.11; mean age = 10), and 30 adolescents (18 males and 12 females aged 12.1-18; mean age = 15.2) participated in the study. All the subjects were comprehensively diagnosed with diagnostic interviews (K-SADS or DICA-R). According to our data, generalized anxiety disorder can be diagnosed in adolescents with mild MR, with high agreement between self-reports and parent reports. Phenomenology of GAD in mildly developmentally delayed persons grossly paralleled that of normal IQ people, except for brooding, somatic complaints, and sleep disorders. Number and severity of symptoms did not correlate with Full Scale and Verbal IQs. High rates of comorbidity with depression were evident both in normal IQ and in developmentally delayed subjects. PMID- 10855761 TI - Higher brain blood flow at amygdala and lower frontal cortex blood flow in PTSD patients with comorbid cocaine and alcohol abuse compared with normals. AB - Posttraumatic stress disorder (PTSD) patients with histories of cocaine and alcohol abuse (CA-PTSD) were compared with normal volunteers. Positron emission tomography (PET) scans with 15O-butanol were used to compare regional cerebral blood flow (rCBF) between the groups during rest and during an auditory continuous performance task (ACPT). CA-PTSD patients had significantly higher rCBF in right amygdala and left parahippocampal gyrus than normals during the ACPT. Normals had higher rCBF at frontal cortex during the resting scan and during the ACPT. The role of the amygdala in attention and fear conditioning suggests that increased amygdala rCBF may be related to clinical features of PTSD. Cocaine use may be associated with increased amygdala rCBF in PTSD patients. Amygdala and frontal cortex attention system components may be reciprocally related and their relative contributions to processing of neutral stimuli perturbed in CA-PTSD. PMID- 10855762 TI - Attachment and developmental psychopathology. AB - This article describes the case of an 8-year-old boy who was first brought to clinical attention at age 2 1/2. He was followed by several clinicians over a 6 year period and was assigned a wide variety of diagnoses. From the first assessment to hospitalization at age 8, a pattern of mother-child relationship disturbance is evident. The use of principles and recent research derived from attachment theory provides a useful basis for formulation in this complex case. This report focuses on the relationship between the patient's mother's internal working models of attachment and his behavior in infancy and early childhood. PMID- 10855763 TI - Reverse Othello syndrome subsequent to traumatic brain injury. AB - Delusional syndromes that occur following head injury are frequently ascribed directly to the consequences of organic insult and seen as empty of psychological significance. The presence of an organic factor, however, does not necessarily indicate that delusional ideation is a direct product of that factor. In this article a detailed report is given of Reverse Othello syndrome (a delusional belief in the fidelity of a romantic partner) appearing in a 49-year-old male following extremely severe traumatic brain injury. This case report highlights the interaction and interpenetration of a complex array of biological, psychological, and social factors in the crystallization of a delusion system. It is argued, following Jaspers, that the emergence of erotically themed delusions following trauma may represent an active attempt to regain intrapsychic coherence and to confer meaning on otherwise catastrophic loss or emptiness. PMID- 10855764 TI - The Kleine-Levin syndrome as a neuropsychiatric disorder: a case report. AB - The Kleine-Levin syndrome (KLS) is characterized by periodic, sudden-onset episodes of hypersomnia, compulsive hyperphagia, and behavioral-emotional disorders (typically indiscriminate hypersexuality, irritability, impulsive behaviors), lasting from a few days to a few weeks, with almost complete remission in the intercritical periods. Depression, confusion, and thought disorders are frequently associated with the critical symptomatology, and they may suggest other psychiatric diagnoses (schizophrenia, mood disorder, conversion disorder) or a substance abuse. A diencephalic-hypothalamic dysfunction is suspected, even if this composite symptomatology cannot easily be linked to a simple mechanism. The aim of this article is to illustrate problems in differential diagnosis, using a case approach. History, course, and therapeutic intervention in a 21-year-old patient with KLS, associated with a clear psychiatric symptomatology and a critical affective pattern, is reported. Psychiatric correlates of KLS are discussed, including the relationship with affective disorders and the possible emotional impact of the attacks. Implications regarding a combined psychological and pharmacological treatment are also discussed. PMID- 10855765 TI - The mind-body harmony. PMID- 10855766 TI - Herpes simplex virus: the importance of asymptomatic shedding. AB - Herpes simplex virus (HSV) is frequently shed after infection of the genital or perianal area. HSV shedding, as determined by culture, occurs on about 3% of days for immunocompetent women and men, and more for persons with HIV infection or if measured by polymerase chain reaction (PCR). Most horizontal and vertical transmission of HSV occurs during unrecognized or asymptomatic shedding, and the majority of HSV-2-infected persons are unaware of their infection. Many persons with 'asymptomatic' HSV-2 infection can learn to recognize genital signs and symptoms as recurrences of HSV-2 infection. However, some shedding episodes remain truly asymptomatic even after patient education. Antiviral therapy dramatically reduces asymptomatic shedding, and trials to evaluate its effect on HSV transmission are underway. PMID- 10855767 TI - Testing for type-specific antibody to herpes simplex virus--implications for clinical practice. AB - Recently, assays that can distinguish between antibody to herpes simplex virus (HSV) types 1 and 2 have become available. These tests not only make it possible to better define infection in symptomatic patients and their sexual partners but also to identify asymptomatic infected individuals who may, nevertheless, be infectious. Type-specific antibody tests for HSV have several potential applications. They have a clear role in helping to define the worldwide distribution and pattern of HSV infection, and a potential role in the management of individual patients, although this has yet to be formally established and evaluated. Because of the high costs and potential disadvantages of targeted screening, particularly in the absence of effective interventions to prevent acquisition or transmission of infection, the public health benefits of screening need to be formally evaluated before its widespread introduction. PMID- 10855768 TI - Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8): epidemiology and pathogenesis. AB - Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus type 8 (HHV 8), is a gamma2 herpesvirus (rhadinovirus) and the most recently discovered human tumour virus. It is involved in the pathogenesis of Kaposi's sarcoma (KS), primary effusion lymphoma and the plasma cell variant of multicentric Castleman's disease. KSHV is not pathogenic in most otherwise healthy individuals but is highly oncogenic in HIV-1-infected and iatrogenically immunosuppressed individuals. It establishes a latent infection in most KS spindle (endothelial tumour) cells and in the neoplastic B cells of primary effusion lymphomas. The KSHV genome contains several homologues of cellular genes known to regulate cell growth and differentiation. Although some of these have transforming properties in vitro, their precise role in oncogenesis is still under investigation. Other co-factors may be involved in the pathogenesis of KS in HIV-uninfected, immunocompetent individuals, e.g. in African endemic KS, but none have been identified yet. Transmission of KSHV among homosexual men appears to occur through sexual contact, but in endemic countries transmission is frequent in childhood and may occur through close contact within families. Four major variants of KSHV have been defined, on the basis of variability in the K1 gene; they may have co-evolved with certain human populations. In addition, some KSHV strains may have resulted from a recombination event with a related, but not yet identified, gamma2 herpesvirus. PMID- 10855769 TI - Betaherpesviruses in transplant recipients. AB - The three betaherpesviruses known to infect humans are cytomegalovirus (CMV) and human herpesviruses 6 and 7 (HHV-6 and -7). All three viruses can infect opportunistically after organ transplantation. CMV causes a variety of end-organ diseases, including pneumonitis, hepatitis and gastrointestinal ulceration. Patients who develop overt CMV disease have significantly higher CMV viral loads than infected patients without evidence of clinical disease. A high CMV viral load largely explains the previously described risk factors for the development of CMV disease, which include donor/recipient serostatus before transplant and viraemia after transplant. CMV also causes some cases of allograft rejection, which can be prevented by antiviral prophylaxis. Application of similar quantitative methods for the study of HHV-6 and -7 have shown that HHV-6 and CMV are significantly and independently associated with biopsy-proven graft rejection after liver transplantation. The full clinicopathological significance of the betaherpesviruses may, thus, be greater than is currently appreciated. PMID- 10855770 TI - T-cell therapy of cytomegalovirus and human immunodeficiency virus infection. AB - Acute and persistent virus infections are limited by the development and maintenance of host T-cell responses to viral antigens. Individuals with congenital or acquired immunodeficiencies are at risk of progressive and often life-threatening infection. Recent studies have provided insight into the nature of protective T-cell responses to viruses and advances in T-cell culture technology have made it possible to evaluate the adoptive transfer of T-cell clones of defined antigen specificity and function to restore deficient responses in immunocompromised hosts. The progress of these studies in cytomegalovirus and human immunodeficiency virus infection is the subject of this review. PMID- 10855771 TI - The U.S.-Russian radiation health effects research program in the Southern Urals. AB - The Joint Coordinating Committee for Radiation Effects Research (JCCRER) was established through a bilateral U.S.-Russian agreement to support research and exchange information on radiation health effects. The U.S. member agencies include the Department of Energy (DOE), Nuclear Regulatory Commission (NRC), Department of Health and Human Services (DHHS), Department of Defense (DoD), National Aeronautics and Space Administration (NASA), and Environmental Protection Agency (EPA). The Russians are represented by the Ministries of Emergencies (EMERCOM), Atomic Energy (MINATOM) and Health (MINZDRAV), and the Russian Academy of Sciences (IBRAE). The focus of this research is on the workers from the Mayak Production Association (MAYAK) in the Southern Urals and on the neighboring populations along the Techa River exposed to contamination from the plant. The goal of the program is to better define the relationship between the health effects and the chronic low dose and dose-rate exposure, these data being essential to validate current radiation protection standards and practices. The current primary areas of JCCRER research include dose reconstruction, epidemiologic health studies, molecular epidemiology/biodosimetry, and the creation of tissue banks. The organization of the ongoing research conducted under the aegis of the JCCRER and the rationale for this work are described. PMID- 10855772 TI - The basic directions and results of activities of branch no. 1 of the state research center of the Russian Federation "Biophysics Institute" (FIB-1). AB - During the 1950's, in the early years at the Mayak Production Association (MAYAK) in the Southern Urals, there was a rapid expansion of plutonium output. This was carried out when nuclear technology was still being developed and knowledge of the effects of radiation exposure on humans was not well understood. As result of the discharge of liquid waste into the Techa River, there was large scale overexposure to both the workers at the facility and the population of riverside localities on the Techa River. There were also other environmental discharges, due to an accident and problems of waste storage, which contributed to exposure of other populations around MAYAK. Although all information on the MAYAK operation and its consequences for both workers and population was kept secret, studies on nuclear technology improvement, the handling of radioactive waste, of medical sequence of radiation exposure, radiation safety improvement, and prevention and treatment of radiation injuries were initiated virtually from the onset the of nuclear weapons production program. PMID- 10855773 TI - Radiological medical data preservation in the Southern Urals. AB - Information is being microfilmed and scanned in the Southern Urals to ensure the archival preservation of the unique, important and irreplaceable records documenting chronic low-level radiation exposure to workers and neighboring populations of the Mayak Production Association (MAYAK), as well as health effects. The records include dosimetric and epidemiologic information maintained on workers and neighboring populations located at facilities in Ozyorsk and Chelyabinsk. Microfilming is being done in Ozyorsk, and on a more limited basis in Chelyabinsk, where a scanning project has also recently been initiated. Over 1,800 rolls of film have been produced as of the fall of 1999. This article describes the background and results of this data preservation effort and includes brief summary tables describing the types of records being preserved. Researchers interested in access to these records should contact Paul Seligman, Deputy Assistant Secretary for the Office of Health Studies at the Department of Energy (DOE) for more information. PMID- 10855774 TI - Reconstruction of radionuclide contamination of the Techa River caused by liquid waste discharge from radiochemical production at the Mayak Production Association. AB - Because of its importance to reconstructing radiation doses for ongoing epidemiological studies, a feasibility study was undertaken to determine if the source term of radioactive materials released to the Techa River from the Mayak Production Association, the first facility in the former Soviet Union for the production of plutonium, could be reconstructed from historical measurements made at a limited number of downriver locations. The feasibility study used historically measured water flow rates and total-beta radioactivity measurements, and considered the processes of radioactive decay and of sorption/desorption. A simple radionuclide mass balance approach was used. To determine the rate of input of radionuclides to the Techa River system, the Techa River was depicted as a series of segments for which measurements are available. For each segment of the river, a system of recurrent (with time) equations was compiled for radioactivity balance accounting for the radioactivity inflow at the inflowing end, activity discharge with water at the outflowing end, and the reduction of activity because of radioactive decay. The equations change with time to account for the changing nature of the river regime. Effective sorption constants for 90Sr and 137Cs, which characterize the transport of radionuclides among the river system components (water and bottom sediments), were defined based on the inventory of these radionuclides deposited at each of the studied river segments and data on water concentration and radioactive removal. All the information on radioactive contamination of the river system components during the period 1949 1996 was used. Solution of the series of equations provided information on the rate of input of these radionuclides into the upper end of the river. The pilot study indicated that it is possible to determine the historical releases of a wider suite of radionuclides using the historical monitoring data from numerous locations along the river, rather than relying on a more uncertain reconstruction of quantities released at the point of discharge. Radionuclides considered include 90Sr, 106Ru, 137Cs, and 144Ce. Estimated concentrations of selected radionuclides at various times are presented. PMID- 10855775 TI - The Techa River dosimetry system: methods for the reconstruction of internal dose. AB - The Mayak Production Association (MPA) was the first facility in the former Soviet Union for the production of plutonium. Significant worker and population exposures occurred as a result of failures in the technological processes in the late 1940's and early 1950's. Residents of many villages downstream on the Techa River were exposed via a variety of pathways; the more significant included drinking of water from the river and external gamma exposure due to proximity to contaminated bottom sediment and shoreline. After the extent of the major contamination of the Techa River became known, several villages on the upper part of the Techa River were evacuated. Organ doses are being reconstructed on the basis of derivation of an historical source term and a simple river model used to simulate the transport of radionuclides downstream and their retention on sediments; measurements of 90Sr content in teeth and the whole body of half of the members of the cohort; and development of the "Techa River Dosimetry System" for computation of the doses. PMID- 10855776 TI - Radioecological impacts of the Techa River contamination. AB - During the years 1949-1952, the Mayak Production Association (MAYAK), which was processing weapons-grade plutonium, was discharging radioactive wastes into the Techa River. As a result, all components of the river system (water, bottom soils, and flood plains) were exposed to massive radioactive contamination. The protective measures taken in the 1950's resulted in the improvement of the radioecological conditions in the Techa River. After 1952, the radioecological conditions in the area were mostly determined by the long-lived radionuclides 9OSr and 137Cs. This article focuses on the dependencies governing the migration of radionuclides along the vertical and horizontal planes in different components of the river system over a 40-y period. Until the 1990's, a decrease in 90Sr and 137Cs contents was noted in environmental samples and foodstuffs produced in riverside villages. In the subsequent years, the radioecological situation on the Techa stabilized. The sources of the current contamination of the river are represented by the runoffs from by-pass canals and swampy upper reaches. PMID- 10855777 TI - Deterministic effects from occupational radiation exposures in a cohort of Mayak PA workers: data base description. AB - Project 2.3 of the Joint Coordinating Committee on Radiation Effects Research (JCCRER) is a study of deterministic health effects among a cohort of Russia nuclear workers. The preliminary study population includes a stratified random sample of 221 radiation workers who were employed in a cohort of 8,055 workers at the Mayak PA facilities for at least one year during the period from 1948 to 1958. High annual doses, approaching 1 Gy per year from external and internal radiation sources, were reported for a significant proportion of the workers in this cohort. The present data set includes 96 cases of chronic radiation sickness (CRS), 14 cases of acute radiation syndrome (ARS) and 13 cases of plutonium pneumosclerosis (PPn). The remainder of the sample consists of "uninjured workers" who had no known history of radiation illness or injury; however, the uninjured workers are not "controls" for radiation exposure. The data base is currently being expanded to 600 individuals sampled from the cohort of workers from 1948 to 1958 to allow a more complete analysis of the deterministic health effects and comparisons with existing health effect models. The final data base will be used with state-of-the-art modeling techniques to determine threshold doses and dose-response relationships for key clinical diagnostic variables. PMID- 10855778 TI - Stochastic effects of environmental radiation exposure in populations living near the Mayak Industrial Association: preliminary report on study of cancer morbidity. AB - The Mayak Industrial Association, located in the South Ural Mountains, began operation in 1948 and was the first Russian site for the production and separation of plutonium. During the early days of operation, technological failures resulted in the release of large amounts of radioactive waste into the Techa River. Residents who lived in villages on the banks of the Techa and Iset Rivers were exposed to varying levels of radioactivity. The objective of this study is to assess stochastic (carcinogenic) effects in populations exposed to offsite releases of radioactive materials from the Mayak nuclear facility in Russia. Subjects of the present study are those individuals who lived during the period January 1950 through December 1960 in any of the exposed villages along the Techa River in Chelyabinsk Oblast. Death certificates and cancer incidence data have been routinely collected in the past from a five-rayon catchment area of Chelyabinsk Oblast. The registry of exposed residents along the Techa River assembled and maintained by the Urals Research Center for Radiation Medicine for the past 40 y is the basis for identifying study subjects for this project. Specific study objectives are to evaluate the incidence of cancer among current and former residents of Chelyabinsk Oblast who are in the exposed Techa River cohort; integrate results from the dose-reconstruction study to estimate doses for risk assessment; and develop a structure for maintaining continued follow-up of the cohort for cancer incidence. In the earlier part of our collaborative effort, the focus has been to enhance the cancer morbidity registry by updating it with cancer cases diagnosed through 1997, to conduct a series of validation procedures to ensure completeness and accuracy of the registry, and to reduce the numbers of subjects lost to follow-up. A feasibility study to determine cancer morbidity in migrants from the catchment area has been proposed. Our preliminary analyses of cancer morbidity underscore the importance of examining both cancer mortality and cancer morbidity in conducting a comprehensive analysis of the occurrence of cancer in this important cohort. PMID- 10855779 TI - Metabolism and dosimetry of actinide elements in occupationally-exposed personnel of Russia and the United States: a summary progress report. AB - The United States Transuranium and Uranium Registries (USTUR) and the Dosimetry Registry of the Mayak Industrial Association (DRMIA) have been independently collecting tissues at autopsy of plutonium workers in their respective countries for nearly 30 y. The tissues are analyzed radiochemically and the analytical data are used to develop, modify, or refine biokinetic models that describe the depositions and translocations of plutonium and transplutonium elements in the human body. The purpose of this collaborative research project is to combine the unique information on humans, gathered by the two Registries, into a joint database and perform analyses of the data. A series of project tasks are directly concerned with dosimetry in Mayak workers and involve biokinetic modeling for actinide elements. Transportability coefficients derived from in-vitro solubility measurements of actinide-containing aerosols (as measured by the DRMIA) were related to specific workplaces within Mayak facilities. The transportability coefficients of inhaled aerosols significantly affected the translocation rates of plutonium from the respiratory tract to the systemic circulation. Parameters for a simplified lung model, used by Branch No. 1, Federal Research Center Institute of Biophysics (FIB-1) and the Mayak Production Association for dose assessment at long times after inhalation of plutonium-containing aerosols, were developed on the basis of joint USTUR and DRMIA data. This model has separate sets of deposition and transfer parameters for three aerosol transportability groups, allowing work histories of the workers to be considered in the dose assessment process. FIB-1 biokinetic models were extended to include the distributions of actinide elements in systemic organs of workers, and a relationship between the health of individual workers and plutonium distribution in tissues was determined. Workers who suffered from liver diseases generally had a smaller fraction of systemic plutonium in the liver at death and a larger fraction in the skeleton than did relatively healthy workers. Also, the fraction of total systemic plutonium excreted per day was significantly greater for workers with liver diseases than for relatively healthy workers. These observations could have a considerable effect on organ dosimetry in health impaired workers whose dose assessments were based solely on urinary excretion rates. A comparison of this model to other biokinetic models, such as those published by the International Commission for Radiological Protection, is currently underway as is the documentation of uncertainty estimates associated with the model. PMID- 10855780 TI - Development of an improved dosimetry system for the workers at the Mayak Production Association. AB - Databases are being created that contain verified and updated dosimetry and worker history information for workers at the Mayak Production Association. Many workers had significant external and internal exposures, particularly during the early years (1948-1952) of operation. These dosimetric and worker history data are to be used in companion epidemiology studies of stochastic and deterministic effects. The database contains both external and internal dose information and is being constructed from other databases that include radiochemical analyses of tissues, bioassay data, air sampling data, whole body counting data, and occupational and worker histories. The procedures, models, methods, and operational uncertainties will be documented and included in the database, technical reports, and publications. The cohort of the stochastic epidemiological study is expected to include about 19,000 persons while the cohort for the deterministic epidemiological study is expected to include about 600 persons. For external dosimetry, workplace gamma, beta, and neutron doses are being reconstructed. The models used for this incorporate issues such as known isotopes, composition, shielding, further analysis of film badge sensitivities, and records of direct measurements. Organ doses from external exposures are also being calculated. Methods for calculating dose uncertainties are being developed. For internal dosimetry, the organ doses have been calculated using the established FIB-1 biokinetic model. A new biokinetic model is being developed that includes more information of the solubility and biokinetics of the different chemical forms and particulate sizes of plutonium that were in the workplace. In addition, updated worker histories will be used to estimate doses to some workers where direct measurements were not made. A rigorous quality control procedure is being implemented to ensure that the correct dosimetry data is entering the various databases being used by the epidemiologists. PMID- 10855781 TI - The promise of molecular epidemiology in defining the association between radiation and cancer. AB - Molecular epidemiology involves the inclusion in epidemiologic studies of biologic measurements made at a genetic and molecular level and aims to improve the current knowledge of disease etiology and risk. One of the goals of molecular epidemiology studies of cancer is to determine the role of environmental and genetic factors in initiation and progression of malignancies and to use this knowledge to develop preventive strategies. This approach promises extraordinary opportunities for revolutionizing the practice of medicine and reducing risk. However, this will be accompanied by the need to address and resolve many challenges, such as ensuring the appropriate interpretation of molecular testing and resolving associated ethical, legal, and social issues. Traditional epidemiologic approaches determined that exposure to ionizing radiation poses significantly increased risk of leukemia and several other types of cancer. Such studies provided the basis for setting exposure standards to protect the public and the workforce from potentially adverse effects of ionizing radiation. These standards were set by using modeling approaches to extrapolate from the biological effects observed in high-dose radiation studies to predicted, but mostly unmeasurable, effects at low radiation doses. It is anticipated that the addition of the molecular parameters to the population-based studies will help identify the genes and pathways characteristic of cancers due to radiation exposure of individuals, as well as identify susceptible or resistant subpopulations. In turn, the information about the molecular mechanisms should aid to improve risk assessment. While studies on radiogenic cancers are currently limited to only a few candidate genes, the exponential growth of scientific knowledge and technology promises expansion of knowledge about identity of participating genes and pathways in the future. This article is meant to provide an introductory overview of recent advances in understanding of carcinogenesis at the molecular level, with an emphasis of the aspects that may be of use in establishing the association between radiation and cancer. PMID- 10855783 TI - Report on routine discharges from UK civil nuclear sites. PMID- 10855782 TI - NOAA space weather alerts. National Oceanic and Atmospheric Administration. PMID- 10855784 TI - Engineering antibodies for the clinic. AB - In the last ten years recombinant 'protein drugs' such as erythropoietin or tissue plasminogen activator have become widely used in the clinic. After some early setbacks antibodies look well placed to join them. A decade of antibody engineering is finally beginning to pay off with a string of chimeric and humanized antibodies gaining the Food and Drug Administration approval in the last two years. Here we will report on recent developments in the clinical application of antibodies, in particular, in the treatment of malignant lymphoma. We will also discuss some of the current strategies for the engineering of both whole antibodies (IgG) and recombinant antibody fragments for the next generation of antibody therapeutics. PMID- 10855785 TI - Transgenic mice as a source of fully human antibodies for the treatment of cancer. AB - The last two years have seen a renaissance of monoclonal antibodies for the treatment of disease. Of the eight antibodies currently approved for human therapy, two are for the treatment of cancer. In large part, the revival of antibodies has been driven by technology developments geared toward making antibodies less likely to elicit an anti-antibody response in humans. The development of transgenic mice, XenoMouse animals, capable of making fully human antibodies offers new opportunities for generating antibodies of therapeutic quality. Recently, this technology has been applied to the generation of a fully human antibody to the epidermal growth factor receptor. A description of the development of this antibody serves to illustrate the power and ease of use of XenoMouse technology. PMID- 10855786 TI - Role of an anti-epidermal growth factor receptor in treating cancer. AB - Recent technological advances, together with the discovery of the important role many growth factors play in modulating cell proliferation and differentiation, have led to the development of novel therapeutic agents for the treatment of cancer. In particular, advances in hybridoma technology and molecular engineering have permitted the development of humanized or chimeric monoclonal antibodies capable of interfering with growth factor signaling pathways. One promising target of interest is the epidermal growth factor receptor (EGFr), which is activated by the ligands EGF and TGF-alpha. This ligand receptor interaction plays a crucial role in the growth and survival of many human cancers. A chimeric (human/mouse) monoclonal antibody p6tuximab (IMC-C225) targets the EGFr and has potential clinical value as an anticancer agent. PMID- 10855787 TI - Target antigens for prostate cancer immunotherapy. AB - The detection and treatment of prostate cancer has been markedly improved by the use of Prostate-Specific Antigen (PSA) as a serological biomarker for disease. However, even after surgical intervention and hormone ablation therapy, a significant proportion of patients progress to advanced metastatic disease, for which there is no cure. An important goal has become the identification of antigens in advanced stage prostate cancer that represent targets for therapy. Recently, great progress has been made to utilize immunological therapies to treat cancer. Monoclonal antibody therapy has been successfully approved for the treatment of breast cancer and B-cell lymphoma, and multiple clinical trails are currently in progress in a variety of cancers, including prostate cancer. Pre clinical and clinical studies are also underway to evaluate cancer vaccine approaches directed against antigens that are highly expressed in prostate and other cancers. This article describes several target antigens expressed in prostate cancer and immunological approaches directed against them that may be effective for treating prostate cancer patients. PMID- 10855788 TI - Monoclonal antibody therapy of human gliomas: current status and future approaches. AB - The development of immunotherapeutic protocols for the treatment of human CNS neoplasia over the past two decades has been impressive. Several crucial aspects have been defined, characterized, and in many cases, optimized (Wikstrand CJ, Zalutsky MR, Bigner DD: In: Liau LM, Bigner DD (eds) Brain Tumor Immunotherapy. Humana Press (in press), 2000). Specific Mabs or constructs reacting with targetable antigens are currently available and in clinical trial. In addition, additional antigens currently under study (angiogenesis-related markers, developmentally associated antigens for medulloblastoma such as L1, and the identification of new targets by SAGE, just in its infancy, will provide a veritable library of available targets for therapy. The molecular engineering and affinity maturation techniques being applied to Mab fragment optimization have already been rapidly effective in generating a variety of Mab constructs of appropriate affinity for clinical trial; as new targets are defined, and experience is accrued with the various constructs currently and prospectively available, the optimal targeting of a multitude of antigens will be possible. PMID- 10855789 TI - Infusion reactions associated with the therapeutic use of monoclonal antibodies in the treatment of malignancy. AB - With the FDA approval of Rituximab in 1998 for the treatment of lymphoma, and Trastuzumab in 1999 for the treatment of breast cancer, monoclonal antibodies were officially added to the therapeutic armamentarium against malignancy. Most of the side effects associated with these agents are due to antigen-antibody interactions on specific cells and tissues. One of the most predictable side effects of these products is a constellation of various systemic effects including flu-like symptoms such as headache, fever, sweats, skin rash, shortness of breath, hypotension, nausea, and asthenia that occurs with the first infusion of such products. Rarely severe hypotension, bronchospasm, and hypoxia and even death have occurred. The pathophysiology of these reactions appears to be secondary to the release of cytokines as the antibodies bind do circulating antigen-expressing cells that are then removed in the reticuloendothelial system of the lungs, spleen and liver. In patients with large numbers of antigen-dense cells that have a high mitotic index, such as prolymphocytic leukemia, mantle cell lymphoma, or lymphosarcoma cell leukemia, there is a risk of true tumor lysis syndrome. One should be particularly cautious when treating patients with high numbers of circulating antigen-expressing cells in the setting of underlying cardiovascular or respiratory disease. PMID- 10855790 TI - Targeting vascular endothelial growth factor (VEGF) for anti-tumor therapy, by anti-VEGF neutralizing monoclonal antibodies or by VEGF receptor tyrosine-kinase inhibitors. AB - Vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) is an important mediator of tumor-induced angiogenesis and represents a potential target for innovative anticancer therapy. In several animal models, neutralizing anti-VEGF/VPF antibodies have shown encouraging inhibitory effects on solid tumor growth, ascites formation and metastatic dissemination. Targeting the VEGF signaling pathway by means of VEGF receptor tyrosine-kinase inhibitors has shown similar efficacy in animal tumor models. Several of these anti-VEGF therapies are currently being tested in clinical trials in cancer patients. The profiles and effects of the neutralizing anti-VEGF/VPF antibodies and the VEGF receptor tyrosine-kinase inhibitors in animal models are reviewed and of the risks and benefits of VEGF blockade by one or the other treatments are discussed. PMID- 10855791 TI - Prostate-specific membrane antigen (PSMA)-specific monoclonal antibodies in the treatment of prostate and other cancers. AB - Prostate-specific membrane antigen (PSMA) is a cell surface glycoprotein that is expressed by prostate epithelial cells. PSMA-specific monoclonal antibodies have been utilized to characterize the biologic function and in vivo biodistribution of PSMA. PSMA is an attractive target protein for monoclonal antibody directed imaging or therapeutics for prostate cancer since its expression is relatively restricted to prostate epithelial cells and is over-expressed in prostate cancer, including in advanced stages. Currently, clinical usage of PSMA specific monoclonal antibodies has been limited to diagnostic immunohistochemistry and imaging of patients with prostate cancer. Novel applications for these antibodies will be discussed. PMID- 10855792 TI - VCP, a weak ATPase involved in multiple cellular events, interacts physically with BRCA1 in the nucleus of living cells. AB - BRCA1, a breast/ovarian cancer susceptibility gene, undergoes mutations in as many as 50% of familial breast tumors. Recent studies indicate that BRCA1 may be involved in DNA damage repair. Here, we demonstrate that the BRCA1 protein physically associates with valosin-containing protein (VCP), a member of the ATPases associated with a variety of cellular activities (AAA) superfamily. In vitro studies revealed that VCP, via its N- terminal region, binds to amino acid residues 303-625 in the BRCA1 protein. Although found predominantly in the cytoplasm and, less abundantly, in the nucleus, VCP can be translocated from the nucleus after stimulation with epidermal growth factor. Collectively, our results suggest that VCP, by binding to BRCA1, participates in the DNA damage-repair function as an ATP transporter, possibly facilitating the transcription-coupled repair. PMID- 10855793 TI - Characterization of trinucleotide- and tandem repeat-containing transcripts obtained from human spinal cord cDNA library by high-density filter hybridization. AB - In order to identify trinucleotide- and tandem repeat-containing transcripts in human spinal cord, hybridization of a high-density spinal cord cDNA library filter was carried out using a radioactively labeled degenerate oligonucleotide designed to detect different trinucleotide repeats including those known to occur in disease-associated expansions, in a single step. The sequence analysis of the trinucleotide repeat-containing transcripts (TNRTs) revealed 23 known mammalian genes with trinucleotide repeat-containing regions (TNRs), some of which were not previously reported to contain TNRs, and 18 cDNA clones with no or insignificant sequence homology to known genes. Amongst the known genes detected was the fragile X gene (FMR-1) containing (CGG)30. Other genes containing extended TNRs of 9 to 21 repeats were calcium-dependent protease, ATBF1-A, ferritin H chain, and the G protein Gsalpha2. Ten sequences containing perfect TNRs and two sequences containing perfect tandem repeats (derived from 11 TNRTs) were further analyzed for allelic variation using primers flanking the TNR, and five were shown to exhibit two to five alleles per TNR. These transcripts were further investigated for their chromosomal localization where unknown or only partially characterized. The transcripts that were polymorphic in the TNR region were ATBF1 A (a homeodomain protein), clone 390013 on chromosome Xp11, a member of the family of the 14.3.3 protein kinase C regulators, a human translation initiation factor (an isolog of the yeast Suilisol gene 1), and a novel sequence (TR21). Only the first two transcripts showed the presence of rare expanded alleles. Characterization of polymorphic TNRs in novel and even known genes expressed in human spinal cord is likely to help in the identification of new candidates for genes involved in neurodegenerative disorders. PMID- 10855794 TI - BMP-2 and sonic hedgehog have contrary effects on adipocyte-like differentiation of C3H10T1/2 cells. AB - The signaling pathways of bone morphogenic protein 2 (BMP-2) and Sonic hedgehog (Shh) are related during embryogenesis. Both proteins have been implicated as important components during osteogenic differentiation; e.g., considering their in vitro effects in the pluripotent C3H10T/1/2 cell system. Also, BMP-2 has been frequently reported to stimulate adipogenesis as well as osteogenesis in these cells. We investigated the relative potencies of Shh and BMP-2 with regard to adipogenesis. We performed differentiation experiments by stimulating C3H10T1/2 cells with BMP-2, Shh, or a combination. We monitored adipocyte-like differentiation via gene expression analysis and cytologic staining. An adipocytic phenotype was observed in BMP-2-treated cells, as shown by upregulation of two adipocytic marker mRNAs, PPAR-gamma and aP2, and by staining of lipid-filled cell vesicles with Oil Red O. In contrast, no adipocyte-like differentiation could be detected either after treatment with Shh or after exposure to a combination of Shh and BMP-2. Our results demonstrate for the first time that Shh and BMP-2 have contrary effects on adipocyte-like differentiation. Whereas BMP-2 promotes the adipocytic lineage, Shh suppresses the expression of the BMP-2-induced fat-cell phenotype. PMID- 10855795 TI - Regulation of DNA synthesis by the higher-order chromatin structure. AB - Mouse erythroleukemia cells were treated with the topoisomerase II poison VP-16, the intrastrand crosslinking agent cis-DDP, and the ribonucleotide reductase inhibitor hydroxyurea. In all cases, the rate of DNA synthesis decreased as a result of the treatment. To study the mechanism of inhibition of DNA chain elongation, we determined DNA synthesis in a cell-free replication system containing isolated nuclei and cytoplasmic extracts. The rate of DNA synthesis in the reactions containing nuclei isolated from untreated cells and extracts from cells treated with the three drugs were slightly reduced and did not show significant differences between the drugs. In the systems containing nuclei from cells treated with cis-DDP, DNA synthesis was again slightly inhibited; synthesis in nuclei treated with hydroxyurea was enhanced, and synthesis in the systems containing nuclei from cells treated with VP-16 was significantly reduced. DNA synthesis was reduced to the same extent in a system containing nuclei isolated from untreated cells that had been briefly sonicated to introduce a limited number of double-strand breaks in the DNA. As VP-16 and sonication mediate changes in chromatin topology, these results suggest that, along with the trans acting signal transduction pathways, there is a topologic mechanism for regulation of DNA synthesis in the S phase of the cell cycle. PMID- 10855796 TI - Genomic structure and promoter analysis of the mouse EphA8 receptor tyrosine kinase gene. AB - The gene encoding the mouse EphA8 receptor tyrosine kinase has been isolated from a mouse genomic library, and its complete genomic structure has been determined. This gene spans approximately 28 kb and consists of 17 exons. This gene structure is similar to the structure of the chick EphB2 (Cek5) gene, except for one intron present between the first two exons encoding the EphA8 kinase domain. This difference may reflect an evolutionary divergence of the catalytic domain between EphA and EphB subgroup receptors. The site for transcription initiation has been mapped to the 19th nucleotide upstream from the translation start codon ATG. A feature of this gene is an unmethylated CpG island spanning exon 1 and the flanking sequence. The putative promoter of the EphA8 gene lacks a TATA box and contains multiple copies of the sequence GGGCGG, the core sequence of the putative Sp1-binding site. The 3.5-kb upstream genomic region containing part of the first exon showed strong promoter activity in NG108-15 neuroblastoma cells but much less in 293T cells, suggesting that this fragment is sufficient for neural cell-directed promoter activity. By deleting the genomic region containing the five GC boxes, it was shown that the minimal promoter region is primarily comprised of five copies of the Sp1-binding site located upstream from the transcription initiation site. Finally, in situ RNA hybridization studies revealed a very specific pattern of EphA8 gene expression restricted to the rostral region of midbrain tectum during embryonic development. Isolation of a functional promoter for the EphA8 gene is a first step in understanding how expression of this gene is controlled at the molecular level. PMID- 10855797 TI - Absence of PTEN/MMAC1 pseudogene in mice. AB - The PTEN gene encodes a phosphatase that acts as a tumor-suppressor gene and is mutated in a variety of human cancers. Alterations of the PTEN gene in these tumor samples were identified using exon-by-exon analysis of the gene using single-stranded conformational polymorphism or direct sequencing of PTEN cDNA. However, in humans, mutational analysis of a PTEN cDNA template can produce false results because of a highly conserved PTEN processed pseudogene that shares more than 98% homology with the coding region of functional PTEN. PTEN-knockout mice develop tumors, suggesting that mouse tumor models are useful in vivo model systems to study PTEN function. Any mutational analysis of mouse PTEN cDNA may also produce false results if mice contain a highly conserved PTEN pseudogene. In this paper, we demonstrate the absence of any PTEN pseudogene in the mouse and discuss the significance of this observation for the mutational studies of the PTEN gene in mouse tumor models. PMID- 10855798 TI - Intron-exon organization and phylogeny in a large superfamily, the paralogous cytochrome P450 genes of Arabidopsis thaliana. AB - The cytochrome P450 gene superfamily is represented by 80 genes in animal genomes and perhaps more than 300 genes in plant genomes. We analyzed about half of all Arabidopsis P450 genes, a very large dataset of truly paralogous genes. Sequence alignments were used to draw phylogenetic trees, and this information was compared with the intron-exon organization of each P450 gene. We found 60 unique intron positions, of which 37 were phase 0 introns. Our results confirm the polyphyletic origin of plant P450 genes. One group of these genes, the A-type P450s, are plant specific and characterized by a simple organization, with one highly conserved intron. Closely related A-type P450 genes are often clustered in the genome with as many as a dozen genes (e.g., of the CYP71 subfamily) on a short stretch of chromosome. The other P450 genes (non-A-type) form several distinct clades and are characterized by numerous introns. One such clade contains the two CYP51 genes, which are thought to encode obtusifoliol 14a demethylase. The two CYP51 genes have a single intron that is not shared with CYP51 genes from vertebrates or fungi, or with any other Arabidopsis P450 gene. Only a few of the Arabidopsis P450 genes are intronless (e.g., the CYP710A and CYP96A subfamilies). There was a relatively good correlation between intron conservation and phylogenetic relationships between members of the P450 subfamilies. Gene organization appears to be a useful tool in establishing the evolutionary relatedness of P450 genes, which may help in predictions of P450 function. PMID- 10855799 TI - Social work practice in a rural community collaborative to improve perinatal care. AB - U.S. rural women encounter many logistical and psychosocial barriers to prenatal care, including high poverty rates, high rates of inadequate health insurance, health care provider shortages, transportation problems and health care systems that may be inadequate or unresponsive to the needs of poor women. The purpose of this article is to describe the role of the social worker in rural communities in the development, implementation and evaluation of a community effort that sought to improve prenatal care through a collaborative of health and human services organizations. The nature of social work participation and the implications for social work are discussed. PMID- 10855800 TI - Evaluation research on the effectiveness of social work intervention on dialysis patients: the first three months. AB - The research reported in this article examined the effectiveness of a dedicated, timed, recurring Master's level social work intervention with patients new to dialysis. Study participants were evaluated for depression and levels of adjustment when first beginning dialysis and three months later. A control group received standard, mandated social work services while the experimental group received an additional counseling component. The experimental group showed statistically significant changes over time in lowered levels of depression and maladjustment. PMID- 10855801 TI - Anti-anxiety medications: a review for social workers. AB - Social workers in mental health settings frequently participate in the treatment of clients with incapacitating anxiety. Medication is often included among the interventions for these clients. Social workers, who specialize in psychosocial interventions, also have key roles in the assessment and monitoring of medication effects. The purpose of this article is to provide social workers with an overview of the pharmacological treatment of anxiety so that they can carry out their range of interventions more effectively. Included is information about types of medication and their dosages, positive and adverse effects, interactions of medication with other interventions, and special concerns with children and adolescents. PMID- 10855802 TI - Hospital social workers' attitudes toward euthanasia and assisted suicide. AB - Euthanasia and assisted suicide are the subjects of increasing controversy in the health care setting. In this study of 122 hospital social workers' attitudes toward euthanasia and assisted suicide, many respondents reported agreement that both practices may be ethical, should be legal in some situations, and that they would be willing to participate in the practices. Almost one-fourth of the respondents have been asked by patients and families during their careers to discuss euthanasia and assisted suicide. The social workers also identified situations in which euthanasia and assisted suicide may be appropriate and safeguards that should apply if practices are legalized. Preparation for requests to discuss these practices, through awareness of their own beliefs and attitudes and becoming knowledgeable about current controversies, policies, and practices, is essential. By doing so, social workers will be ready to seize the opportunity to emerge as leaders of multidisciplinary discussion of complex ethical issues in health care. PMID- 10855803 TI - The role of values and experience in determining social workers' attitudes toward euthanasia and assisted suicide. AB - This article reports results of a study examining the impact of personal and professional values and experience on 122 hospital social workers' attitudes toward euthanasia and assisted suicide. Respect for self-determination was rated as the most important consideration in end-of-life issues. Predictors of social workers' agreement that euthanasia should be legal were: self-determination, religious beliefs, educational level (BSW/MSW), and for assisted suicide were: religious beliefs, belief in the potential for abuse, educational level and participation in ethics training. The findings underscore the need for social workers' awareness of how an interplay of personal and professional factors in potentially explosive ethical issues may influence practice in health care settings. PMID- 10855804 TI - An automated and modified technique for testing the retinal function (Arden test) by use of the electro-oculogram (EOG) for clinical and research use. AB - The electro-oculogram (EOG) can be recorded by use of skin electrodes positioned near the eye in the horizontal or vertical plane and is dependent on the position of the eye relative to the electrodes and the size of the corneo-retinal potential (CRP). By use of the functional relation between the recorded potential and the eye position - CRP is considered to be constant - the EOG is the eye movement recording method most often used. Changes in CRP dependent on luminance or metabolic influences can be measured, on the other hand, by means of the EOG when eye movements of constant amplitudes are performed. The EOG in this paper is used in the latter sense. The widely used clinical test of rentinal function based on the electro-oculogram firstly described by Arden (Arden index) has been modified to achieve more precise and reliable results by including a gradual dark adaptation procedure and a special data analysis to detect fixation periods. Using modern techniques, the complete procedure has been automated. The efficiency of the procedure and the device is demonstrated in various ways. PMID- 10855805 TI - Visual half-field contrast sensitivity in children with dyslexia. AB - We address the question whether left-hemispheric and/or right-hemispheric contrast thresholds differ between children with dyslexia and controls, and whether there are interhemispheric differences. In order to answer these questions we examined [1] thresholds for the detection of Contrast-Defined (CD) forms to left and right half-field stimulation, and [2] half-field pattern onset Evoked Potentials (EPs) as a function of stimulus contrast in 21 children with dyslexia of 8-15 years of age and in 17 age-matched healthy controls. It was found that (A) children with dyslexia were less sensitive to CD forms than controls irrespective of half field of stimulation. In children with dyslexia as well as controls the forms were more easily detected in the right visual half field than in the left one: (B) Peak latencies of the pattern onset EPs to contrast levels above 20% were up to 10 ms longer in children with dyslexia than in controls irrespective of half field of stimulation, and (C) children with dyslexia had smaller mean amplitudes for all contrast levels used. (D) Neither children with dyslexia nor controls showed hemispheric differences in the pattern onset EPs. PMID- 10855806 TI - Normal EOG values in intraretinal metastasis from cutaneous melanoma: a case report. AB - PURPOSE: The electro-oculogram (EOG) is a powerful test to diagnose primary and metastatic choroidal tumors. While in benign tumors light-peak to dark-trough ratio values are in the range of normal subjects, these values appear highly altered in eyes affected by malignant choroidal tumors. Here we report a clinical case of a patient with intraretinal metastasis from cutaneous melanoma; notwithstanding the malignancy of the tumor, the EOG doesn't present alterations. METHODS AND RESULTS: Standard electro-oculographic recordings were performed before and after local excision of the tumor: recordings from the normal eye were taken as control. The EOG values were always normal in both eyes. Histological sections showed no evident change in the cell population of the retinal pigment epithelium (RPE) and Bruch's membrane. CONCLUSION: Our results suggest that the presence of an intact RPE is a crucial requirement to obtain a normal EOG. PMID- 10855807 TI - The mouse ERG before and after light damage is independent of p53. AB - Death of retinal photoreceptors by apoptosis is observed under many physiological and pathological conditions such as histogenesis, retinal dystrophies and light induced photoreceptor degeneration. To date, little is known about regulatory mechanisms for apoptosis in the retina. The tumor suppressor gene p53 is a regulator of apoptosis in a number of systems, however, p53-independent apoptosis has also been described. We have therefore investigated whether the lack of p53 influences the dark-adapted ERG in C57BL/6 p53-/- mice compared to p53+/+ control littermates under physiological (regular light-dark cycle) conditions. We also recorded ERGs at 12 to 14 h in darkness following diffuse bright light exposure to 8,000 or 15,000 lux for 2 h. ERG analysis over a range of 6 logarithmic units of light intensity revealed normal and virtually identical a-, b-, c-waves and oscillatory potentials in dark-adapted p53+/+ and p53-/- mice. After exposure to diffuse white fluorescent light strong decreases of all ERG components were found to be very similar in both genotypes. These data support the notion that the p53 protein is neither essential for normal retinal function nor for processes involved in light-induced depression of the ERG in mice. PMID- 10855808 TI - Pattern reversal visual evoked response in retinitis pigmentosa. AB - PURPOSE: To determine if the pattern reversal visual evoked response (PVER) can objectively evaluate visual function in retinitis pigmentosa (RP). METHODS: We retrospectively reviewed the medical records of 29 patients (29 eyes) with RP, a visual acuity (VA) better than 20/100, and for whom an electroretinogram (ERG) and a PVER had been recorded. A steady-state PVER was measured using five check sizes: 160, 80, 40, 20 and 10 min of arc. The best-corrected VA was measured using the ETDRS chart. Visual fields (VFs) were measured using standard Goldmann perimetry. To quantify the VFs, we measured the field size (isopter IV-2) in four meridians and averaged them. Twenty-five eyes of 25 normal subjects served as controls. RESULTS: The mean and median VAs were 20/40 (logMAR = 0.30 +/- 0.20). The average VF (isopter IV-2 ) was 20 degrees (median, 9 degrees: range. 2 degrees to 62 degrees). All patients had a recordable PVER. Only 12 (41 %) patients had a recordable ERGs, all subnormal. The PVER amplitudes showed a relatively good correlation with VA in the intermediate check sizes (40 min of arc, r = 0.611. P = 0.0004: 20 min of arc, r = 0.596. P = 0.0007). The PVER amplitude-check size function had a flattened inverted V shape in the RP patients and an inverted U shape in the normal subjects, with the mean amplitudes for the RP patients significantly smaller than the normals at all check sizes. CONCLUSION: The PVER is a useful method for objectively evaluating the visual function in RP patients whose central vision is still preserved. PMID- 10855809 TI - Rod and rod mediated function in patients with beta-thalassemia major. AB - An electroretinographic (ERG) study was undertaken to test the hypothesis that scotopic retinal function is altered in transfused thalassemics on chronic Deferoxamine (DFO). ERG a- and b-wave responses and dark adapted visual thresholds were obtained from 11 patients with beta-thalassemia major, ages 7 to 38 (median 17) years. A quantitative model of the activation of phototransduction was fitted to the a-waves to estimate the gain of the transduction processes and the saturated amplitude of the rod photoresponse. From b-wave stimulus/response functions. the saturated b-wave amplitude and an index of b-wave sensitivity (log sigma ) were calculated. The patients' data were compared to those of normal subjects. The relations of the ERG parameters to age. average ferritin level, and duration of transfusion without DFO as well as other clinical parameters were examined. Longitudinal measures of b-wave responses and dark adapted visual thresholds. available for nine of the patients, were examined for significant change over time. For all patients both the gain and saturated amplitude of the rod response are normal. In two patients log sigma is below the 99% prediction interval for normal. One has low scotopic visual sensitivity. The duration of transfusion therapy unprotected by DFO chelation therapy was correlated with log a. These results suggest iron accumulation rather than DFO toxicity underlies scotopic dysfunction in older thalassemics. some of whom may have had extended periods of transfusion without the protection of chelation. Thus, monitoring of retinal function is recommended in such patients. PMID- 10855810 TI - Electroretinogram and visual-evoked potentials in children with optic nerve coloboma. AB - The aim was to evaluate alterations in Visual-Evoked Potentials (VEP) and Electroretinogram (ERG) and discover whether these tests are useful for determining residual visual acuity in cases where a patient is unable to collaborate. Flash and, when possible, Transient Pattern Reversal Visual-Evoked Potentials and Maximal Response ERG were recorded in 8 children (under three years old) affected by different aspects of optic nerve coloboma. None of them had visual acuity evaluated because of poor collaboration. All examinations were carried out using skin electrodes. Amplitude of the a and b component of ERG, amplitude, morphology and latency of the major components of Flash VEP and amplitude and latency of P100 Pattern Reversal VEP were evaluated. Four of the patients were examined three years later and visual acuity was compared with the previous electrofunctional results. Alterations in ERG were found only in the case of significant retinal anomalies (great coloboma, retinal detachment), huge alterations were found in both Flash VEP and in Pattern Reversal VEP. The retrospective study of VEP traces and visual acuity showed a good correlation between electrofunctional data and visual capability. Electrofucntional examinations can identify important conductive retinocortical anomalies. The possibility of correlating electrophysiological results with residual visual acuity is important for rehabilitative management in such optic disc malformations. PMID- 10855811 TI - Time courses for maturation of electroretinogram responses from infancy to adulthood. AB - The purpose of this study was to determine how responses in the normal human electroretinogram (ERG) change with subject age. We studied 62 children, 10 days to 15 years old, and 30 subjects 15-37 years old, using the standard protocol established by the International Society for Clinical Electrophysiology of Vision, with Burian-Allen bipolar contact-lens electrodes. We measured rod response, maximal response, oscillatory potentials (OPs), cone response, flicker response, and b-wave amplitude/log intensity (V/log I) curve. A logistic growth curve was used to describe the developmental changes. Dark- and light-adapted ERG a- and b-wave amplitudes reached adult levels by three to five years of age. although b-wave amplitudes of scotopic rod-mediated responses were slower to reach maturity than mixed rod-cone mediated responses. In early infancy OPs were the most immature of the ERG responses, although the rate of development thereafter exceeded that of the other responses such that OP amplitudes were within adult levels by two years of age. Amplitudes of the ERG responses in 21 children sedated with chloral hydrate did not differ significantly from 21 who had not been sedated. ERG responses developed at varying rates, reflecting different developmental stages in photoreceptors, middle retinal layers and more proximal retina. PMID- 10855812 TI - Streptococcal meningitis in a five-month-old male lamb. PMID- 10855813 TI - Comments on 'Ketamine hydrochloride--an adjunct for analgesia in dogs with burn wounds'. PMID- 10855814 TI - Vacuum phenomenon in equine carpal, metacarpophalangeal and metatarsophalangeal joints. AB - In order to establish the incidence of vacuum phenomenon in horses' carpal, metacarpophalangeal and metatarsophalangeal joints, stress-flexed radiographs were made of normal joints, joints with known pathology and in anaesthetised horses with joints under traction. Focal intra-articular radiolucencies were identified in normal stress-flexed carpal, metacarpophalangeal and metatarsophalangeal joints. These radiolucencies can be confused with those associated with pathological conditions. PMID- 10855815 TI - The influence of dietary crude protein intake on bone and mineral metabolism in sheep. AB - Increased dietary protein consumption is thought to cause calciuresis, a negative calcium balance and increased bone loss that may result in skeletal deformities and fracture. To explore this hypothesis, 40 approximately 100-day-old meat-type Merino ram lambs were fed, for 6 months, diets with an increasing crude protein (CP) content (114, 142, 171 and 190 g/kg DM) but approximately on an iso-nutrient basis with regard to metabolisable energy, calcium and phosphorus. Increased protein consumption modestly (NS) enhanced calciuresis and resulted in significant (P < or = 0.01) limb skewness. This could not, however, be ascribed to osteopaenic bones, and compared with animals consuming lower protein rations, the bone mineral density (BMD) and vertebral trabecular bone volume of animals fed high protein diets were significantly increased: the BMD of thoracic vertebrae was positively related to the CP intake (r = 0.62; P < or = 0.001). In animals consuming higher protein diets, skeletal radiology and quantitative bone histology revealed no evidence of increased bone turnover as would be expected in animals that are in negative calcium balance. No relationship existed between limb skewness and the growth rate of lambs. However, the ratio of Ca:P in the forelimb (r = -0.98), vertebrae (r = -0.72) and rib (r = -0.42) was found to be inversely correlated with increased protein intake and resulted from an increase in the phosphorus content of bone, while the amount of bone calcium was unaffected. We conclude that qualitative micro-architectural abnormalities, and not mere bone loss, may underlie the skeletal deformities induced by increased protein consumption in sheep. PMID- 10855816 TI - An experimental rat model of salt-sensitive hypertension; biochemical and morphological parameters and sympathetic nervous system. AB - The objective of the study was to outline the characteristics of the development of hypertension and some neurohumoral, haematological and morphological factors contributing to development of high blood pressure in a genetic model of salt sensitive rat. Characteristics of Dahl salt-sensitive (DS) rats, as compared to their Dahl salt-resistant (DR) controls were as follows: 1) DS rats display higher blood pressure and lower heart rate compared to DR rats as early as 1 month of age at weaning). They gradually develop hypertension at 2 months of age, irrespective of diet. Low-Na diet (0.5% NaCl) does not prevent hypertension but delays its development and ameliorates it. High Na-diet (8% NaCl) exacerbates hypertension. 2) DS rats have retardation in body weight gain. They develop mild hypochromic anaemia. 3) After 2 months of Na loading (3 months of age), DS rats express significantly increased Na and water retention and increased plasma volume by 15% compared to 2.8% increase in DR rats on high-Na diet. 4) DS rats showed renal parenchymal lesions, more pronounced after Na-loading, focal atrophy of cortical tubules, mesangial matrix expansion and glomerulosclerosis. Consistent with high blood pressure were changes in renal arterioles, fibromuscular proliferation, deposition of fibrinoid material in intima. 5) Sodium loading produced increased activity of the sympathetic nervous system (SNS), and sodium restriction reduced SNS responsiveness. PMID- 10855817 TI - Semen characteristics of goats with subacute, acute and chronic besnoitiosis. AB - A study on the semen obtained from breeding goats suffering from mild to severe chronic besnoitiosis revealed marked changes in semen volume, colour, density, concentration, mass and individual motility and percentage live. There were also many neutrophils and spermatozoa with primary and secondary defects, including missing tails and deformed heads and tails. The observed changes were considered to be severe enough to account for the infertility observed in the flock. Sections of testes obtained for histopathology were characterised by massive blockage of the pampiniform plexus, degeneration of the germinal epithelium, tubular necrosis with an inflammatory infiltrate and, in some cases, accumulation of haemosiderin-like material in the tunica vaginalis. PMID- 10855818 TI - A primary animal health care approach to treatment and control of flea (Ctenocephalides felis) infestation in indigenous goats kept on communal grazing. AB - This paper describes a primary health care approach to an infestation of indigenous goats by the common cat flea, Ctenocephalides felis. The flea species was identified using scanning electron microscopy. The infested goats were kept on communal grazing at Winterveld in the North-West Province. They were penned at night in housing made of wire and corrugated iron. The owner complained that the goats were lethargic. Fleas were found on the goats and flea larvae were found in the kraal. Haematology and blood biochemistry performed on the infested goats revealed no abnormalities; however, infestation caused irritation that made the animals lethargic. Available flea control methods for domestic animals were appraised in terms of cost, availability and ease of administration at a primary animal health care level using participatory extension methods. It was found that a carbamate powder was available, affordable and effective for flea control in this small flock of goats kept under communal grazing conditions. Although the authors had observed fleas on goats kept under similar conditions elsewhere in Mpumalanga and the North-West Province, this was the 1st time that the species had been identified. PMID- 10855819 TI - Spinal nephroblastoma in an Irish wolfhound. AB - A 1-year-old Irish wolfhound was presented with a history of slowly progressive left pelvic limb paresis. A neurological examination demonstrated bilateral deficits referable to the thoracolumbar spinal cord. Lumbar cerebrospinal fluid contained neoplastic cells. An intradural, extramedullary mass was demonstrated by myelography at the caudal aspect of T13. Surgical excision was abandoned owing to severe macroscopic damage to, and apparent infiltration of, the cord, and the dog was euthanased. The tumour was diagnosed histologically as an extrarenal nephroblastoma. Nephroblastoma should be suspected in young, large-breed dogs with intradural extramedullary masses over spinal segments T10-L2. The prognosis for complete recovery after surgical excision is guarded to poor. PMID- 10855820 TI - Three case studies involving Leptospira interrogans serovar pomona infection in mixed farming units. AB - Three case studies involving Leptospira interrogans serovar pomona outbreaks within mixed farming systems in South Africa are described. On 2 farms, pigs constituted the main enterprise with cattle and sheep of secondary importance. On each of these 2 farms, abortion due to L. pomona in sows was confirmed by culture, and antibody titres to pomona were detected in cattle, sheep, horses and dogs. On the 3rd farm, a piggery was of secondary importance to cattle farming. Abortion and death in cows occurred on this farm and serology showed titres to various serovars, including pomona. L. pomona was also isolated from bovine urine, an aborted bovine foetus and kidneys from slaughtered pigs. This particular case study was regarded as clinically atypical in that adult Jersey cattle died of acute leptospirosis in a semiarid region of South Africa. In all 3 case studies, the poor management of pig effluent and of the drinking water and its sources played a pivotal role in the transmission of the disease. Inadequate vaccination of animals against Leptospira and poor record-keeping within the secondary farming enterprises were also contributing factors to the spread of leptospirosis. PMID- 10855821 TI - Cryptococcosis in captive cheetah (Acinonyx jubatus): two cases. AB - Cryptococcus neoformans is a yeast-like organism associated with pulmonary, meningoencephalitic, or systemic disease. This case report documents 2 cases of cryptococcosis with central nervous system involvement in captive cheetah (Acinonyx jubatus). In both cases the predominant post mortal lesions were pulmonary cryptococcomas and extensive meningoencephalomyelitis. Both cheetahs tested negative for feline immunodeficiency virus and feline leukaemia virus. The organism isolated in Case 2 was classified as Cryptococcus neoformans var. gattii, which is mainly associated with disease in immunocompetent hosts. PMID- 10855822 TI - Chlamydia-induced septic polyarthritis in a dog. AB - A systemic disease associated with pyrexia, lymphadenopathy, and arthropathy of several joints of the appendicular skeleton in a dog is described. Chlamydia-like organisms were detected on light-microscopic examination of a smear made from joint fluid aspirated from one of the affected joints. A group-specific lipopolysaccharide antigen shared by all Chlamydia spp. was demonstrated by direct fluorescent antibody staining of joint fluid, which also proved positive for chlamydia by means of the relevant polymerase chain reaction test. An indirect fluorescent antibody test on serum was also positive, although the complement fixation test was negative. Attempts to grow the organism from joint aspirates in the yolk sac of embryonating hens' eggs and on appropriate tissue cultures, however, failed. Chlamydia spp. are considered to have played an aetiological role in this case, making it the first substantiated case of naturally-occurring arthropathy in a dog due to chlamydiosis. The origin of the infection could not be traced. PMID- 10855823 TI - A focus of feline babesiosis at Kaapschehoop on the Mpumalanga escarpment. PMID- 10855824 TI - The pharmacology of halogenated salicylanilides and their anthelmintic use in animals. AB - The halogenated salicylanilides are a large group of compounds developed mainly for their antiparasitic activity in animals. Several halogenated salicylanilides with potent antiparasitic activity have been synthesised of which only closantel, niclosamide, oxyclozanide, rafoxanide and resorantel are commercially available. Closantel and rafoxanide, which represent the most important drugs in the group, are used extensively for the control of Haemonchus spp. and Fasciola spp. infestations in sheep and cattle and Oestrus ovis in sheep in many parts of the world. Niclosamide is used extensively for its anticestodal activity in a wide range of animals. Antiparasitic activity of the halogenated salicylanilides has also been demonstrated against a large number of other internal parasites, in particular haematophagous helminths, and external parasites including ticks and mites, in a variety of animal species. Several cases of toxicity and mortality have been reported for closantel and rafoxanide in sheep and goats. Their unique pharmacokinetic behaviour appears to play an important role in the efficacy and safety of these compounds. The chemical and physical characteristics, mode of action, pharmacokinetics, antiparasitic activity and toxicity of the halogenated salicylanilides in animals are reviewed. PMID- 10855825 TI - Relative bioavailability of rafoxanide following intraruminal and intra-abomasal administration in sheep. AB - The bioavailability of rafoxanide was compared after intraruminal and intra abomasal administration in healthy adult sheep (n = 6) in a single dose, 2 parallel group study at 7.5 mg/kg. Rafoxanide concentrations in plasma were measured by means of HPLC analysis. Primary pharmacokinetic parameters for bioavailability and disposition of rafoxanide in plasma for both routes of administration were determined by non-compartmental and non-linear, 1 compartmental pharmacokinetic analysis, respectively. Significantly (P < or = 0.05) higher peak plasma concentrations (c(max)) of rafoxanide and a more rapid rate of absorption (c. 3.5 times) was observed in sheep after intra-abomasal (i a) administration compared to intraruminal (i.r.) administration. A significantly (P < or = 0.05) longer lag period (t(lag)) before absorption (6.8 +/- 2.9 h) occurred after i.r. than after i-a treatment (1.9 +/- 0.6 h). There was no significant difference (P > 0.05) in AUC, MRT and in the rates of elimination (k10-HL and t(1/2beta)) between the i.r. and i-a routes of administration. The results of the study demonstrated the important influence of the rumino-reticulum on absorption of rafoxanide in sheep. PMID- 10855826 TI - Intravascular plasma disposition and salivary secretion of closantel and rafoxanide in sheep. AB - The plasma and salivary disposition of closantel and rafoxanide were examined following intravenous administration in adult sheep. Two studies were conducted with rafoxanide at 7.5 mg/kg and 1 with closantel using 2 doses (5 and 15 mg/kg). The pharmacokinetic profile of both drugs in plasma were best described by a 2 compartmental model with 1st-order rate constants. Plasma disposition of closantel and rafoxanide were characterised by a rapid distribution (t1/2(alpha)) of <30 min), long elimination half-life (t1/2(beta)) of 17.0 +/- 4.0 days for closantel and 7.2 +/- 0.6 days for rafoxanide), small apparent volume of distribution (V(SS) of <0.15 l/kg) and a slow rate of total body clearance (Cl of <0.01 ml/min/kg). The area under the drug plasma concentration curve (AUC) of closantel at 5 mg/kg was nearly twice as large as that of rafoxanide at 7.5 mg/kg resulting from the slower t1/2(beta) observed with closantel compared to rafoxanide. Large individual differences were observed in the rate measurements of distribution (k12, k21 and t1/2(alpha)), whereas the parameters of elimination (k10, t1/2(beta) and Cl), were more consistent between animals. A dose proportional increase in AUC was observed for closantel administered at 5 and 15 mg/kg. A low, constant salivary concentration of closantel (mean of 0.04 +/- 0.05 microg/mL) and rafoxanide (mean of 0.07 +/- 0.04 microg/mL) was observed during the 24-h examination period after dosing. PMID- 10855827 TI - A preliminary attempt to use climate data and satellite imagery to model the abundance and distribution of Culicoides imicola (Diptera: Ceratopogonidae) in southern Africa. AB - Abundances of Culicoides imicola, the insect vector of several livestock viruses, including bluetongue and African horse sickness, were recently published for 34 sites in southern Africa, together with associated climate data. Here, these data are analysed statistically in combination with certain satellite-derived variables, with the aim of developing predictive models of C. imicola abundance. Satellite-derived variables were the land surface temperature (LST, a measure of temperature at the earth's surface) and the normalised difference vegetation index (NDVI, a measure of photosynthetic activity). Two models were developed: (1) climatic variables only and (2) satellite-derived and climatic variables. For model I, the best model used a single predictor variable (the mean daily minimum temperature) only, and accounted for nearly 34% of the variance in C. imicola abundance. Two variable climatic models did not perform significantly better. For model II, the best 1-variable model used the annual minimum LST as a predictor of C. imicola abundance, and accounted for nearly 40% of the variance in C. imicola abundance. The best 2-variable model, which gave a significantly better fit than the 1-variable model, combined the minimum LST and minimum NDVI as predictors of C. imicola abundance, and accounted for nearly 67% of variance. A map of predicted C. imicola abundances is produced on the basis of this 2nd model which, despite some anomalies, agrees largely with what is currently known of the prevalence of C. imicola in the region. PMID- 10855828 TI - Could treatment of pregnant mares prevent abortions due to equine piroplasmosis? AB - Treatment of pregnant mares to prevent abortions due to equine piroplasmosis is a novel idea practised empirically at some Thoroughbred studs in South Africa. This paper presents the results of an investigation to ascertain whether imidocarb dipropionate crosses the equine placenta. Three pregnant mares were injected intramuscularly with imidocarb and their foetuses were mechanically aborted at varying time intervals thereafter. Imidocarb was found in foetal blood at a level similar to that in the dam's blood, suggesting that imidocarb administered to the dam would be available for anti-parasitic activity in the foetal circulation. Uncertainty concerning the time of treatment to achieve the desired effect currently makes this a questionable exercise. PMID- 10855829 TI - Helminth parasites of dogs from two resource-limited communities in South Africa. AB - Biological samples were collected from dogs in resource-limited communities in the North-West and Gauteng Provinces of South Africa to assess the prevalence of helminth parasitism. These samples included adhesive tape peri-anal skin swabs and fresh faecal samples for helminth examination, and thick and thin blood films (smears) and whole-blood samples in anticoagulant for examination of filarial nematode microfilariae and haemoprotozoa. The eggs of Ancylostoma caninum, Toxocara canis, Toxascaris leonina, Dipylidium caninum and taeniids were identified. None of the blood samples and smears tested positive for microfilariae of Dirofilaria immitis or Dipetalonema spp. or for haemoprotozoa. The adhesive tape swabs were negative for cestode eggs and segments. Most of the helminth parasites identified in this study are zoonotic and consequently are regarded as a public health hazard. PMID- 10855830 TI - The incidence of gastroenteritis diagnosis among sick dogs presented to the Onderstepoort Veterinary Academic Hospital correlated with meteorological data. AB - The number of sick dogs diagnosed with and without gastroenteritis presented to the Onderstepoort Veterinary Academic Hospital situated north of Pretoria is reported from counts extracted from the records of the Outpatients clinic for 6 years, 1988 to 1993. The average percentage of sick dogs diagnosed with gastroenteritis was 11.51% and the average percentage of sick dogs that were admitted to the parvovirus isolation hospital ward was 2.8%. A strong correlation exists between the number of dogs admitted to the parvovirus ward and average monthly wind speed and inverse humidity values. PMID- 10855831 TI - Streptococcus phocae infections associated with starvation in Cape fur seals. AB - Mortalities and abortions associated with starvation occurred at Cape Cross, Namibia, in Cape fur seals (Arctocephalus pusillus pusillus). Affected seals showed lethargy and emaciation, and the most common pathological signs were those of a respiratory infection, both in adults and offspring. Streptococcus phocae was isolated from adult seals, a cub and aborted foetuses. PMID- 10855832 TI - An unusual presentation of suspected oedema disease of swine in Kenya. AB - From a group of 11 recently weaned pigs, 4 were reported to be sick. Clinical examination of the sick pigs revealed marked dyspnoea, bluish-red discolouration of the skin, incoordination and difficulty in walking. Bacteriological examination of the gut contents of 2 pigs that had died earlier yielded pure cultures of haemolytic Escherichia coli. Post mortem examination of the remaining 2 pigs that died subsequently revealed progressive pulmonary collapse. One of these also showed subcutaneous oedema of the head and marked oedema of the mesentery of the spiral colon and oedema of the brain. Microscopically there was pulmonary alveolar collapse and degenerative changes in the liver. On the basis of the clinical signs, isolation of haemolytic E. coli and the post mortem findings, a diagnosis of oedema disease was made. PMID- 10855833 TI - Monensin poisoning in ostriches. PMID- 10855834 TI - The hidden dangers of anaesthetic machines. PMID- 10855835 TI - The effect of gestation and lactation on bone calcium, phosphorus and magnesium in dairy cows. AB - A study was conducted to monitor changes in cortical bone mineral in the dairy cow in response to demands of lactation and pregnancy using rib bone biopsies in serial sampling. Sixteen Friesian cows from the University dairy herd were used to collect 9 samples during the lactation period and 5 samples during the dry period. The data were analysed using a split-plot design analysis of variance. There were no significant (P > 0.05) differences in cortical bone phosphorus concentrations in rib bone during the lactation period, but calcium concentrations in cortical bone were significantly (P < 0.05) higher at parturition and during the first 30 days of lactation compared to the next 30 days and between 90 and 120 days. Results reported here indicate that the cow resorbs cortical bone during the middle of the lactation period and not during the periparturient period as previously thought. Magnesium concentrations were also significantly (P < 0.05) higher at the beginning of lactation compared to some of the other sampling times, but cortical bone was significantly (P < 0.05) thinner at the beginning of lactation compared to several of the other sampling times. There were no significant (P > 0.05) differences in cortical bone Ca or Mg concentrations during the gestation period. Cortical bone P concentrations significantly (P < 0.05) decreased during the first 180 days, but significantly (P < 0.05) increased at 181-230 days and significantly (P < 0.05) decreased again at 231 days to term. Cortical bone thickness decreased significantly (P < 0.05) from the beginning of gestation to term. There were no significant (P > 0.05) differences in cortical bone thickness or Ca or Mg concentrations in cortical bone during the dry period, but cortical bone P concentrations were significantly (P < 0.05) greater at the end of the dry period compared to the first 30 days of the period. In general, cortical bone Ca and Mg values decreased as milk production increased up to 20 kg/day and cortical bone P values and bone thickness increased. In animals producing over 20 kg/day, however, cortical bone mineral values were greater and cortical bone thickness was lower compared to those animals producing less than 20 kg. PMID- 10855836 TI - The effect of an angiotensin-converting enzyme inhibitor on water and electrolyte balance in water-restricted sheep. AB - The importance of angiotensin II in the regulation of water and electrolyte balance in sheep is questionable. In this trial the effects of an angiotensin converting enzyme (ACE) inhibitor were quantified in sheep on restricted water intake. Comparing the phase of water restriction only with that of water restriction plus ACE inhibition, significant increases were observed during the latter phase in urine volume, sodium and potassium excretion via the urine, sodium concentration in the plasma and osmolar clearance. Urine osmolarity decreased with inhibition of angiotensin II formation while variables such as water, sodium and potassium loss via the faeces were unaffected. Most of the renal effects of ACE inhibition, except the increase in urinary potassium excretion, were explicable in terms of the established functions of angiotensin II. Furthermore, results of this trial indicate that angiotensin II has no significant effect on the intestine in regulating water and electrolyte excretion via the faeces. PMID- 10855837 TI - Prevalence of besnoitiosis in domestic ruminants in Kenya: a preliminary survey. AB - A preliminary survey on the prevalence of besnoitiosis in domestic ruminants in Kenya based on field and farm visits, clinical and post mortem examinations and histopathological examination of tissues and biopsies, showed that goats are the most affected, followed by cattle, while sheep were unaffected. Caprine besnoitiosis occurred in a continuous belt in 5 of the 8 provinces in Kenya stretching from the Coast, Eastern, North Eastern, Nairobi and the Rift Valley Provinces. Mandera, in the North Eastern Province, had the highest prevalence rate of 36%, followed by Kwale (35%), Isiolo (35%), Marsabit (33%), Wajir (28%), Nairobi (26%), Meru (24%), Garissa (21%), Taita Taveta (18%), Embu (17%), Kitui (9%), Machakos (7%), Laikipia (3%), Kajiado (2%) and Turkana and Elgeyo-Marakwet (1% each). In all flocks where the prevalence rates were over 6%, kids were observed to be affected. There were no significant differences (P < 0.05) between the prevalence rates in bucks and does (18% and 18.4, respectively), but kids were less (4%) affected. Bovine besnoitiosis was found only in the Tana River District, with an infection rate of 11%. PMID- 10855838 TI - Isolation of Bartonella henselae from a serologically negative cat in Bloemfontein, South Africa. AB - Sera collected from apparently healthy 6-12-month-old cats (n = 31) presented to the Society for the Prevention of Cruelty to Animals Veterinary Clinic in Bloemfontein for neutering were tested for antibodies reactive to Bartonella henselae (Houston-1 strain) by indirect fluorescent antibody testing. Whole blood collected from the cats was used in isolation experiments and subsequent identification of Bartonella species was based on comparison of the nucleotide base sequence of polymerase chain reaction-amplified citrate synthase gene fragments. While none of the cats had antibodies reactive with B. henselae at titres > or =1/64, an organism with a partial citrate synthase gene sequence identical to that of B. henselae (Houston-1) was isolated from 1 cat. PMID- 10855839 TI - Teat lesions and their relationship to intramammary infections on small-scale dairy farms in Kiambu district in Kenya. AB - Mammary gland quarters of 139 lactating dairy cows from small-scale dairy herds were examined visually and by palpation for teat lesions and by California mastitis test (CMT) and bacterial culture for subclinical mastitis. Teat lesions were observed in 97 teats. These included teat chaps (39.2%), teat papillomas (23.7%), teat erosions (22.7%), teat fistulae (5.1%), inverted teats (5.1%) and blocked teats (4.2%). According to the CMT, the prevalence of subclinical mastitis was 33.4% in all the mammary gland quarters, 71.0% in quarters with teat lesions and 24.5% in quarters without teat lesions. There was a significant (P < 0.01) association between teat lesions and the prevalence of subclinical mastitis. The mammary gland quarters with teat lesions were 7.2 times more likely to have a positive CMT (P < 0.01) and 5.6 times more likely to have bacterial organisms (P < 0.01) isolated from them than those without any teat lesions. The bacterial organisms most frequently isolated from the CMT-positive milk samples from both the mammary gland quarters with teat lesions and those without teat lesions were Staphylococcus aureus (50.0%), Streptococcus spp. (34.8%) and Arcanobacterium pyogenes (6.2%). PMID- 10855840 TI - Renal pathology in working dogs in the South African National Defence Force. AB - Urine analysis, serum biochemical profile and a cortical wedge biopsy for histopathological examination was performed on 42 South African National Defence Force (SANDF) dogs from around the country. The only significant finding on urine analysis and serum biochemistry was a relatively large number (16/42) of dogs with elevated serum inorganic phosphate levels. Histopathology revealed that only 9 of the animals had normal kidneys reflected in the wedge biopsy material, with over 50% of them showing signs of glomerular pathology (primarily mesangioproliferative glomerulonephritis). Other conditions detected histopathologically were haemosiderosis (47% of animals), focal nephrosis (2.4%), membranoproliferative glomerulonephritis (2.4%), focal interstitial nephritis (4.7%) and acute tubular nephrosis (4.7%). The lesions observed were of limited distribution and extent; this histopathological finding may account for the absence of significant abnormalities on urine analysis or serum biochemistry profiles. It appears from these results that a large percentage of the SANDF population would be expected to have mild renal lesions, but that these lesions are not severe enough to lead to clinical signs. The findings of this study are similar to those of randomly selected populations of non-military dogs performed in other areas of the world, which also demonstrated an unexpectedly high incidence of histopathological renal pathology in dogs considered healthy. These lesions may well, however, play a role in later life, and it is recommended that military veterinarians maintain an index of suspicion for renal disease, particularly glomerular disease. The aetiology of the histopathological lesions is unknown. PMID- 10855841 TI - Experimental transmission of Besnoitia caprae in goats. AB - Experimental transmission of Besnoitia caprae from naturally chronically-infected goats to susceptible ones was achieved by intra-nasal instillation and intra conjunctival inoculation of cystozoite-containing suspensions, subcutaneous implantation of fascia containing cysts and alternate needle pricking between the infected and non-infected goats. Typical chronic symptoms developed in the fascia infected does. Cystozoite inoculation into the eyes and mouth did not result in infection. Kids born of dams with acute and chronic besnoitiosis did not contract the infection in utero, suggesting that intra-uterine transmission may not occur. In contrast to does with acute besnoitiosis, which occasionally aborted, the does with chronic besnoitiosis gave birth to healthy kids. Kids below the age of 4 months (pre-weaned period) born of both infected and non-infected does were susceptible to besnoitiosis but appeared to be more resistant than adult goats. PMID- 10855842 TI - Etorphine-halothane anaesthesia in two five-year-old African elephants (Loxodonta africana). AB - Anaesthesia of 2 five-year-old female African elephants (Loxodonta africana) was required for dental surgery. The animals were each premedicated with 120 mg of azaperone 60 min before transportation to the hospital. Before offloading, 1 mg etorphine was administered intramuscularly (i.m.) to each elephant to facilitate walking them to the equine induction/recovery room. For induction, 2 mg etorphine was administered i.m. to each animal. Induction was complete within 6 min. Surgical anaesthesia was induced with halothane-in-oxygen after intubation of the trunk. During surgery the mean heart rate was 61 and 45 beats/min respectively. Systolic blood pressures increased to 27.5 and 25.6 kPa respectively, and were treated with intravenous azaperone. Blood pressure decreased thereafter to a mean systolic pressure of 18.1 and 19.8 kPa, respectively. Rectal temperature was 35.6 and 33.9 degrees C at the onset of surgery, and decreased to 35.3 and 33.5 degrees C, respectively, at the end of anaesthesia. Etorphine anaesthesia was reversed with 5 mg diprenorphine at the completion of 90 min of surgery. PMID- 10855843 TI - Laminitis and dermatitis in heifers associated with excessive carbohydrate intake: skin lesions and biochemical findings. AB - The effects of a sudden addition of a large quantity of readily fermentable carbohydrate to the feed ration of pregnant heifers are described. Clinical and pathological changes caused by the resulting disease were confined to the digits and skin. The 4 acutely affected heifers were reluctant to get up or move (group II). They tended to lie down or stand with feet bunched together and the back arched, often shifting weight from limb to limb. They walked stiffly with great tenderness and pain in the digits. Extreme pain was noticed when the digits were examined. In 4 of 8 heifers, separation of the sole at the heel, with leakage of exudate, and under-running of the sole were observed. Necrotic dermatitis of the legs, alopecia and hyperkeratosis of the tail were noticed in all 8 heifers. Skin lesions appeared simultaneously. Four of the heifers (group I) recovered, and the other 4 (group II) were sent to slaughter. No post mortem examination was performed. The biochemical findings revealed a significantly higher concentration of total serum globulins and sodium, and increased activity, in CK, LDH and AST. A significantly decreasing pattern was noted in blood urea concentration, cholesterol, triglycerides, albumin and calcium. No significant differences among the various groups were found in the activities of amylase, GGT, and concentration of creatinine, total bilirubin, inorganic phosphorus, magnesium and potassium. PMID- 10855844 TI - Drug choice and therapeutic drug monitoring in the management of canine primary epilepsy. AB - Therapeutic drug monitoring is an underutilised resource in the management of canine primary epilepsy. Many of the anti-epileptic drugs, including phenobarbitone, have variable pharmacokinetic profiles in different dogs, with each individual animal showing variable rates of absorption, distribution, metabolism and excretion. This results in variable serum drug concentrations with the same oral dose. Many clinicians interpret this situation as therapeutic failure and classify these patients as refractory to treatment. By measuring blood concentrations of drugs at appropriate times, it is possible to explain the efficacy or failure of treatment, and also to prevent serum concentrations from reaching toxic levels. By analysing paired samples, key pharmacokinetic parameters may be calculated for each patient and a profile for the disposition of the drug obtained. Individual optimal drug dosage can be calculated for each patient at little cost to the pet owner. PMID- 10855845 TI - Water intoxication in cattle. AB - Water intoxication is a condition that is common in cattle, and has also been reported in other domestic animals and man. A comprehensive description of the condition is lacking. For a better understanding of the condition, this paper reviews work that has been reported previously by various authors. PMID- 10855847 TI - Age at diagnosis, extent of disease and breast cancer survival: a population based study in Florence, Italy. AB - BACKGROUND: The effect of age at diagnosis on the prognosis of breast cancer is still controversial. The study described the variation by age at diagnosis of some clinical-pathologic features and evaluated the relationship between age and survival, taking into account the effect of extent of disease. MATERIALS: The study comprised a large population-based series of 1,182 invasive breast cancers, incident in the period 1985-1986 in the province of Florence. RESULTS: The proportion of cases without nodal involvement progressively lowered from 59% in the age group < or =39 years to 22% in the age group > or =80 years. The extent of disease was unknown in 14% of cases aged 70-79 years and in 43% of those aged > or =80 years (other age groups: 3%-5%). A lower rate of surgical treatment and axillary surgery were the main reasons for inadequate staging in the elderly. Ten year observed survival progressively decreased from 71% for age < or =39 years to 12% for age > or =80 years. Ten-year relative survival showed less evident differences, dropping from 72% for age < or =39 years to 57% for age > or =80 years. In the relative survival analysis, the differences in relative risks of death among age groups were not significant, either in the univariate or multivariate analysis. Nevertheless, the model with adjustment for extension of disease showed a flattening of the estimated relative risks in age groups over 59 years. CONCLUSIONS: Age at diagnosis was not significantly related to 10-year breast cancer relative survival, suggesting that the worse prognosis in the elderly was largely related to the risk of death from other causes, rather than to a different malignant potential of the tumor. The worse distribution by extent of disease in older women indirectly suggested that diagnostic delays also influenced the different prognosis observed among age groups. PMID- 10855846 TI - The epidemiology of selenium and human cancer. AB - The relation between the trace element selenium and the etiology of cancer in humans remains elusive and intriguing, despite the number of epidemiologic studies published on the topic. We address some methodologic issues, such as misclassification of exposure, particularly to single selenium compounds, effect modification, confounding, and other sources of bias, which may explain the inconsistencies in the literature. We also review the results of cohort studies, which have yielded either inverse or null or direct associations between selenium exposure and subsequent cancer risk. To date, no beneficial effect on cancer incidence at major sites, including prostate cancer, has emerged from the Finnish program begun in 1984 to increase the average selenium intake in its population. Populations exposed to unusually high or low levels of environmental selenium might offer unique opportunities to investigate if selenium exposure is related to the etiology of human cancer. PMID- 10855848 TI - General practitioners and mammographic screening uptake: influence of different modalities of general practitioner participation. Working Group. AB - AIMS AND BACKGROUND: To compare the impact of different modalities of general practitioner (GP) involvement, including the introduction of target payments, on the attendance rate of organized population-based screening programs for breast cancer in Italy. STUDY DESIGN: The study was conducted between 1994 and 1996 in four Italian cities where mammographic screening programs are active: Caltanissetta (CL), Firenze (FI), Modena (MO) and Torino (TO). The impact on attendance rate of different invitation strategies based on active GP involvement was tested in each center. The additional effect of economic incentives was also assessed. The incentives were proportional to the level of compliance attained by each GP and weighted by the size of his eligible patients' list. RESULTS: In the Firenze project, an invitation signed by the GP and the project co-ordinator attained a statistically significant higher participation (difference: 4.2%, chi2 = 7.42, P = 0.006). In Caltanissetta and Torino there was a significant increase of about 7% in the response rate to the postal reminder in the groups contacted by the GPs. No difference was observed in the Modena project between the two groups. CONCLUSIONS: The main contributions of GP involvement can be: "cleaning up" the invitation lists, especially when computerized archives with the mammographic history of the target population are not available; increasing the women's participation by signing the invitation letter, by counseling and active participation in the invitation phase; co-operating in the reminder phase by recalling women non responders at first invitation. The offer of target payment had a certain impact on the screening uptake, but not easily distinguishable from GP signature of the invitation letter; further studies of appropriate design should be planned. Organizational factors, such as availability of a list of non responders, might be crucial in order to enhance the effect of the GPs' action. PMID- 10855849 TI - The availability of histologic grading among 5,923 Italian cancer patients and its relationship with survival: a population-based study. AB - AIM: The specific goal of the study was to evaluate the availability of the histologic grading of cancer and its effect on survival in an Italian population based cancer series. METHODS: Data were drawn from the Tuscany Cancer Registry, active in central Italy since 1985. Among the cases incident during the period 1985 to 1989, bladder, prostate, colon, corpus uteri, rectum and stomach cancers, for which the proportion of graded cases exceeded 50%, were analyzed. Overall, 5,923 cancer cases were included. Ten-year relative survival rates by grade were computed. RESULTS: Overall, data on histologic grading was available only for 38% of cases. The sites most frequently graded were urinary bladder (80%), prostate (73%), colon (71%), corpus uteri (69%), rectum (65%) and stomach (56%). For all the cancer sites analyzed, the 10-year relative survival rates increased as the histologic grading improved. The grade distribution resulted related also to the disease extension, more limited the extension higher the proportion of well differentiated cases. CONCLUSIONS: Due to the evidenced importance of histologic grading as a valuable prognostic factor, it should be requested by clinicians and reported by pathologists more frequently than has been done in the area. PMID- 10855850 TI - Evaluation of quality of life in patients with malignant dysphagia. AB - BACKGROUND: In the last 10 years of clinical research there has been increasing interest in the evaluation of quality of life. Several generic and specific instruments have been developed for this purpose. EORTC QLQ C-30 is a cancer specific questionnaire translated into various languages and validated in several European countries including Italy, where the impact of malignant disease on different areas of quality of life is poorly documented. METHODS: The EORTC QLQ C 30 was administered to 109 patients referred to the endoscopy division of the Istituto Nazionale Tumori, Milan, for endoscopic palliative treatment of malignant dysphagia to test its characteristics in terms of acceptability and clinical validity. RESULTS: In this group of patients the impact of advanced esophageal cancer was highly evident for Emotional and Physical Functioning, Fatigue and Global QoL scales. Dysphagia is a serious problem for many patients; there is a correlation between grade of dysphagia and four QoL dimensions. CONCLUSIONS: QoL assessment is an important tool to evaluate the adequate management of patients with esophageal cancer. The EORTC QLQ-C30 questionnaire proved to be valid and reliable also in this population. PMID- 10855851 TI - Preoperative endoscopic ultrasonography in patients with gastric cancer. AB - AIMS AND BACKGROUND: There is a need to assess the accuracy of endoscopic ultrasonography (EUS) in the diagnosis and staging of gastric cancer, especially in the early and very advanced stages of the disease when the therapeutic approach is still controversial. METHODS: A retrospective study was performed on 79 patients with gastric cancer in order to compare the stage defined by preoperative EUS with that assessed histopathologically. All patients underwent laparotomy for final diagnosis, staging, and eventually treatment. The results of EUS were correlated with the histologic findings of the resected specimens. RESULTS: In the uT1 group, which corresponds to early gastric cancer, the diagnosis was histologically confirmed in 85.7% of the cases. In patients with advanced tumors defined as uT3-uT4, i.e., tumors infiltrating the serosa or neighboring structures, the diagnostic concordance was 91.1%. In contrast, concordance for less advanced lesions confined to the muscular layer was only 31.2%. As regards the lymph nodes, they were defined metastatic in 31 patients and confirmed to be histologically involved in 77.4%. In contrast, when the lymph nodes were assessed as negative at EUS, they proved to be metastatic in more than half the cases. CONCLUSIONS: From the data it appears that EUS has proven to be valuable in correctly staging most of the patients. EUS shows not only tumor depth and local spread but also the passage from a pathologic to a normal wall and lymph node metastasis. EUS appears to represent an important advance in the staging and follow-up of patients with gastric cancer. Instruments and techniques will continue to evolve, but the next level of research should be designed to show that the improved staging provided by EUS has clinical utility and can affect patient outcome. It is noteworthy that the highest accuracy of EUS has been shown in those conditions (uT1 and uT3-4) which currently are under consideration for a therapeutic approach that differs from the standard one. PMID- 10855852 TI - Feasibility of a cell kinetic-based adjuvant chemotherapy trial in axillary node negative breast cancer. AB - AIMS AND BACKGROUND: Accumulated information on biologic prognostic indicators and predictors of response to different types of treatment in patients with different tumor characteristics has made it possible to design clinical protocols on biologic bases. Among cell proliferation indices, the thymidine labelling index (TLI) has proved to be an independent and consistent prognostic indicator over time. Moreover, experimental and retrospective analyses of clinical studies have revealed a direct relation between TLI and response to chemotherapy. On the basis of the results, a prospective clinical protocol on axillary node-negative breast cancer was activated in Italy in 1989. METHODS: Patients with low TLI tumors were treated with local-regional therapy alone, whereas patients with high TLI tumors were randomized to receive local-regional therapy followed or not by adjuvant chemotherapy consisting of 6 cycles of CMF. RESULTS AND CONCLUSIONS: The present paper reports on the feasibility of a prospective clinical protocol based on a subgroup of patients with specific pathologic (node negative) and biologic (rapidly proliferating) breast cancers. However, patient eligibility was only 11%. PMID- 10855853 TI - In vitro interleukin-8 production by monocytes treated with lithium chloride from breast cancer patients. AB - AIMS AND BACKGROUND: Since interleukin-8 (IL-8) has a suppressive effect on hematopoiesis, lithium induces leukocytosis and granulocytosis and mononuclear cells are defective in patients affected by neoplastic disease, we analyzed IL-8 production by monocytes obtained from patients with nonmetastatic breast cancer (BCaM0) and metastatic breast cancer (BCaM1) and the effect of lithium chloride (LiCl) on these cells. Lithium salt compounds are used to limit the degree and duration of neutropenia in patients receiving chemotherapy for cancer and acute leukemia. Lithium influences the hematopoietic system, which is known to be regulated by numerous cytokines including IL-8. METHODS: We selected three groups of subjects (15 per group): patients affected by BCaM0, BCaM1 and healthy donors (HD) matched for sex and age. IL-8 release was assessed in supernatants of lipopolysaccharide (LPS) and/or LiCl-treated monocyte cultures. RESULTS: Monocytes from BCaM1 released higher IL-8 levels than monocytes from BCaM0 (P <0.0001); the IL-8 levels of both groups were significantly higher (P <0.0001) than those of HD. In vitro LiCl treatment reduced IL-8 production by monocytes obtained from all subjects compared to the same cells when untreated or LPS treated. The suppressive effect of LiCl on IL-8 production by monocytes from breast cancer patients was particularly marked in monocytes from BCaM0 with respect to those from BCaM1. LPS treatment increased the IL-8 production more in BCaM1 monocytes than in BCaM0 monocytes. Moreover, combined LPS/LiCl treatment of monocytes significantly (p <0.0001) downregulated the release of IL-8 compared to treatment with LPS alone. CONCLUSIONS: Our data demonstrate that monocytes from BCaM0 release larger amounts of IL-8 than monocytes from BCaM0 and from HD. Lithium was able to downregulate IL-8 production by monocytes from different subgroups. Further studies are needed to clarify if the improvement of the hematopoietic system in vivo observed following lithium therapy could reside, at least in part, in the ability of lithium to downregulate this chemokine. PMID- 10855854 TI - Evaluation of cisplatin combined with ondansetron in Ehrlich ascites carcinoma in vitro and in vivo. AB - AIMS AND BACKGROUND: Nausea and vomiting occur in the majority of patients receiving cisplatin (CDDP) chemotherapy. Ondansetron, a new 5-HT3 receptor antagonist, has been used effectively to control CDDP-induced nausea and vomiting. This study examined the potential of ondansetron to interfere with CDDP antitumor activity and toxicity in Ehrlich ascites carcinoma (EAC). METHODS: The influence of ondansetron on CDDP cytotoxicity was evaluated using EAC cells in culture. In addition, the influence of ondansetron pretreatment on CDDP-induced antitumor activity and host tissue toxicity was studied in EAC-bearing mice. RESULTS: Ondansetron (0.25 microM) enhanced CDDP (0-32 microM) cytotoxicity against EAC cells in vitro. In EAC-bearing mice ondansetron (0.2 mg/kg,ip) administered 1 h before CDDP (7 mg/kg, ip) did not modify the antitumor activity of CDDP. CDDP (7 mg/kg, ip) single treatment induced significant increases in blood urea nitrogen (2-fold) and serum creatinine (2.5-fold) and significant decreases in hematocrit (25%) and white blood cell count (39%) compared to saline treatment. Mice receiving ondansetron 1 h before CDDP showed no significant enhancement of CDDP-induced nephrotoxicity or myelosuppression compared to those pretreated with saline receiving the same dose of CDDP. CONCLUSIONS: This study suggests that the use of ondansetron to control CDDP-induced nausea and vomiting does not affect CDDP antitumor efficacy. PMID- 10855855 TI - Modulatory effects of melatonin and vitamin E on doxorubicin-induced cardiotoxicity in Ehrlich ascites carcinoma-bearing mice. AB - Doxorubicin (Dox), an anthracycline antibiotic, has a wide spectrum of antitumor activity with dose-limiting cardiotoxicity. The drug's toxicity is known to be closely related to the generation of active oxygen free radicals. In our study the normal cardiac tissue contents of total protein, glutathione (GSH) and malondialdehyde (MDA) were significantly decreased, by 25%, 33% and 92%, respectively, in the group of mice bearing Ehrlich ascites carcinoma (EAC) and treated with Dox (4 mg/kg/week x 2, ip). Administration of melatonin (5 mg/kg/day x 15, po) starting 24 hours prior to Dox treatment significantly increased the cardiac contents of total protein and GSH as well as the superoxide dismutase (SOD) activity, by 31%, 36% and 39%, respectively, compared to treatment with Dox only, while the content of MDA was decreased by 26%. Similarly, administration of vitamin E (250 mg/kg/day x 15, po) starting 24 hours prior to Dox treatment significantly increased the cardiac contents of total protein, GSH and SOD, by 23%, 26% and 42%, respectively, while the cardiac content of MDA was decreased by 35% compared with the Dox-only-treated group. As to the oncolytic activity of Dox, pretreatment of EAC-bearing mice with melatonin (5 mg/kg/day x 30, po) or vitamin E (250 mg/kg/day x 30, po) 24 hours prior to Dox administration (4 mg/kg/week x 4, ip) improved the antitumor activity of Dox as indicated by the increase in the average life span of the animals and the number of long-term survivors as well as the decrease in body weight loss induced by Dox treatment. It is clear from these results that administration of melatonin not only protects against the cardiotoxicity induced by Dox treatment but also enhances its antitumor activity to a more significant extent than does vitamin E. PMID- 10855856 TI - Electrochemotherapy with bleomycin in the treatment of hypernephroma metastasis: case report and literature review. AB - A metastasis of hypernephroma was treated by electrochemotherapy with bleomycin. Electrochemotherapy consists of chemotherapy followed by local application of electric pulses to the tumor in order to increase drug delivery to the cells. In the course of a four-week treatment period, the subcutaneous metastasis of the hypernephroma was treated with three sessions of electrochemotherapy, each consisting of 9-10 runs of 8 electric pulses, 10 min after intravenous injection of bleomycin. This treatment resulted in stabilization of the tumor volume for 12 months, whereas the subcutaneous metastasis next to the electrochemotherapy treated one that did not receive electric pulses progressed immediately. PMID- 10855857 TI - Hemangiosarcoma of the breast followed by term pregnancy. AB - AIMS AND BACKGROUND: Primary angiosarcoma of the breast is an unusual neoplasm which is generally associated with a dismal prognosis. Given the poor prognosis and the predominance in the third and fourth decades of life when fertility tends to decrease, reports of term pregnancies after treatment of this disease are rare. METHODS AND STUDY DESIGN: We report a case of angiosarcoma of the breast in a 17-year-old patient with a recurrent breast nodule treated by local surgery only. After three years of follow-up without evidence of disease recurrence she became pregnant. RESULTS: A girl weighing 2,430 g was delivered at 38 weeks and two days. The patient has been free of disease for five years now. CONCLUSIONS: Our experience of the present case shows the lack of full information about this tumor. Pregnancy does not seem to interfere negatively with the course of the disease. PMID- 10855858 TI - Breast metastases from cutaneous melanoma: a report of three cases. AB - In this report we describe three female patients with breast metastases from cutaneous melanoma (CM) who were treated in Slovenia in the period from 1988 to 1991. We found that in 476 Slovenian patents with this disease diagnosed in the given period CM disseminated to the breast less frequently than in other series. In one pregnant and one perimenopausal patient breast involvement by CM was confirmed at the time of widespread dissemination of the disease. Treatment was not effective and the survival of these patients was four months and two weeks and six months, respectively. A slightly better outcome was observed in a normally menstruating patient referred from another country with an isolated solitary breast metastasis from CM. In this patient quadrantectomy seemed to have been sufficient to achieve a disease-free interval of more than eight months. Special attention should therefore be given to a small subset of patients with isolated solitary breast metastases from CM, since their prognosis may be less dismal than in patients with massively infiltrated breasts and disseminated disease. PMID- 10855859 TI - Small cell carcinoma of the esophagus: a case report and review of the literature. AB - We present a case of small cell esophageal carcinoma (SCEC) treated with alternated chemotherapy (including cisplatin, etoposide, vincristine, cyclophosphamide and doxorubicin) and irradiation (36 Gy) followed by surgery. Despite a pathological complete response, the patient died of regional disease recurrence 29 months after the diagnosis. We reviewed the available literature on SCEC with regard to the incidence, clinical symptoms, radiological signs, diagnostic workup, therapeutic modalities and prognosis of this malignancy. PMID- 10855860 TI - A rare case of ceruminous tumor of the cerebellopontine angle. AB - In this case report we describe the development in the cerebellopontine angle of a very rare tumor, ceruminous adenoma. In the few cases described in the literature this tumor occurred in the external acoustic meatus. In four cases it developed in the cerebellopontine angle by infiltration of the petrous bone or by subcutaneous spread. In the present case no connection was found between the cerebellopontine angle and the external acoustic meatus. The most likely pathogenetic hypothesis in this case is that of a tumor of dysembryogenetic origin. PMID- 10855861 TI - [Oxaliplatinum in combination with 5-FU/FA in first-line chemotherapy of metastatic colorectal cancer: a new era in therapeutic strategy]. PMID- 10855862 TI - Whither thromboprophylaxis after total hip replacement? PMID- 10855863 TI - Are recommendations for the routine use of pharmacological thromboprophylaxis in total hip arthroplasty justified? PMID- 10855864 TI - Risks and benefits of prophylaxis against venous thromboembolism in orthopaedic surgery. PMID- 10855865 TI - The contrast between venographic and clinical endpoints in trials of thromboprophylaxis in hip replacement. PMID- 10855866 TI - Thromboprophylaxis--which treatment for which patient? PMID- 10855867 TI - The orthopaedic surgeon and post-traumatic psychopathology. PMID- 10855868 TI - Resection arthrodesis of the knee with a vascularised fibular graft. Medium- to long-term results. AB - We present the results in 12 patients of arthrodesis of the knee using a vascularised fibular graft after resection of a malignant bone tumour. At a mean follow-up of 95 months (60 to 178) all patients were free from disease although 11 had had at least one complication, with stress fracture of the graft in five patients, nonunion in two and deep infection requiring above-knee amputation in one. Despite the high rate of complications, satisfactory results can be obtained using this technique. Careful preoperative counselling is required. PMID- 10855869 TI - Results of open meniscus repair. Long-term follow-up study with a matched uninjured control group. AB - We have followed for 13 years a consecutive series of 31 patients who had open repair of a torn meniscus. They were between 13 and 43 years of age at the time of operation and all had intact stabilising ligaments. Comparison was made with a matched group of normal subjects of similar age and level of activity. The total rate of failure after meniscal repair was 29%; three of the repaired menisci did not heal and six reruptured during the follow-up period. At follow-up 80% of the patients had normal knee function for daily activities. Radiological changes were found in seven. Two had reduction of the joint space (Ahlback grade 1), one with successful and one with failed repair. In the control group of uninjured subjects one knee showed Fairbank changes but none had changes according to Ahlback. The incidence of radiological changes did not differ between the group with meniscal repair and the control group but knee function was reduced after meniscal repair (p < 0.001). We conclude that the long-term results of meniscal repair in stable knees are good with nearly normal function and a low incidence of low-grade radiological changes. PMID- 10855870 TI - In vivo kinematics of total knee arthroplasty. Concave versus posterior stabilised tibial joint surface. AB - We studied the kinetics of the knee in 20 patients (22 knees) 12 months after total knee arthroplasty (TKA), by using three-dimensional radiostereometry and film-exchanger techniques. Eleven knees had a concave (constrained) tibial implant and 11 a posterior-stabilised prosthesis. Eleven normal knees served as a control group. In the posterior-stabilised knees there was less proximal and posterior displacement of the centre of the tibial plateau during extension from 45 degrees to 15 degrees, with a decrease in the anterior translation of the femoral condyles of 4 mm at 45 degrees. There was less internal tibial rotation and increased distal positioning of the centre of the tibial plateau with both designs when compared with the normal knees, and in both the centre of the plateau was displaced posteriorly by more than 1 cm. Increased AP translation has been recorded in all prosthetic designs so far studied by radiostereometry. The use of a posterior-stabilised design of tibial insert could reduce this translation but not to that of the normal knee. PMID- 10855871 TI - Ten-year survival of the cemented Freeman-Samuelson primary knee arthroplasty. Data from the Swedish Knee Arthroplasty Register and the Royal London Hospital. AB - We report a ten-year rate of survival of 96% for the cemented Freeman-Samuelson knee arthroplasty in patients from the Swedish Knee Registry and the Royal London Hospital with revision for aseptic loosening as the criterion for failure. PMID- 10855872 TI - Charnley low-friction arthroplasty with an autograft of the femoral head for developmental dysplasia of the hip. The 10- to 15-year results. AB - Between 1983 and 1988 we carried out 45 Charnley low-friction arthroplasties with autografts from the femoral head in 41 patients for developmental dysplasia of the hip. The preoperative radiographs were assessed for the severity of DDH according to the classifications of Crowe et al, Hartofilakidis et al and Sharp. The postoperative and follow-up radiographs were analysed for coverage of the socket by the graft, for loosening and for the outcome of the fixation of the bone graft. Two patients died (two hips) at four and seven years after THR from causes unrelated to the surgery and were excluded from the final radiological analysis. The mean age of the patients at the time of operation was 46 years 3 months. The autograft of the femoral head covered a mean 26% (16 to 35) of the acetabular component. All the grafts united. Some degree of resorption of the bone graft occurred in 27 patients, and always involved the lateral part of the graft, which was beyond the margin of the socket. After a mean follow-up of 11 years there had been no revisions and 38 patients had no pain or only slight discomfort. One socket migrated and four others were fully demarcated. Our findings indicate that the Charnley LFA with an autograft of the femoral head for DDH remains successful at a follow-up of 15 years. PMID- 10855873 TI - Prediction of the outcome of transtrochanteric rotational osteotomy for osteonecrosis of the femoral head. AB - We have studied the correlation between the prevention of progressive collapse and the ratio of the intact articular surface of the femoral head, after transtrochanteric rotational osteotomy for osteonecrosis. We used probit analysis on 125 hips in order to assess the ratio necessary to prevent progressive radiological collapse over a ten-year period. The results show that a minimum postoperative intact ratio of 34% was required. This critical ratio may be useful for surgical planning and in assessing the natural history of the condition. PMID- 10855874 TI - Factors influencing the outcome of Chiari pelvic osteotomy: a long-term follow up. AB - We have reviewed 103 of 126 Chiari osteotomies carried out in our department between 1956 and 1987. The cases were graded radiologically, using the Japanese Orthopaedic Association (JOA) system, into a pre/early osteoarthritis (OA) group and an advanced OA group. In the pre/early group there were 86 hips. The mean follow-up was for 17.1 years (4 to 37). Preoperatively, 51 hips had an average JOA clinical score of 78.6+/-8.4 points and the final mean JOA clinical score was 89.4+/-12.5 points. Advanced degenerative change developed in 33.7% and one hip required a total replacement arthroplasty (TRA). Chiari osteotomy alone, without accompanying intertrochanteric osteotomy, was performed on 62 hips. For these the median survival time was 26.0+/-2.5 years, using as the endpoint progression to advanced OA. Differences in survivorship curves related significantly to the severity of the preoperative OA, the shape of the femoral head and the level of osteotomy. In the advanced OA group, we followed up 17 hips for a mean of 16.2 years (1 to 27). Before operation, the mean JOA clinical score in 13 hips was 63.2+/-7.9 points and the final score 84.0+/-12.0 points. TRA was eventually carried out on four hips. Our findings suggest that the Chiari osteotomy remains radiologically effective for about 25 years. The procedure is best suited to subluxated hips with round or flat femoral heads and early or no degenerative change. Intra-articular osteotomy can lead to osteonecrosis, and should be avoided. In hips with advanced OA, the Chiari procedure creates an acetabulum which facilitates later TRA, and may delay the need for this procedure in younger patients. PMID- 10855875 TI - CT evaluation of periacetabular osteotomies. AB - We have previously described a simple and reproducible three-dimensional technique of CT for the measurement of the cover of the femoral head in acetabular dysplasia in adults. We now describe the application of this technique in ten patients with symptomatic dysplasia to assess the degree and direction of dysplasia and to measure the cover obtained at acetabular osteotomy. The indices obtained gave a useful indication of the degree and direction of the dysplasia and confirmed which components had been used most efficiently to achieve cover. The information is easily presented in graphical form and gives a clearer indication of the cover obtained than the indices derived from plain radiographs. PMID- 10855876 TI - The anterior centre-edge angle. A cadaver study. AB - The anterior centre-edge (VCA) angle quantifies the anterior cover of the femoral head, and angles of less than 20 degrees are considered abnormal. We have measured the VCA angles in hips without osteoarthritic changes. We took bilateral false-profile radiographs of nine female and 30 male cadavers without signs of osteoarthritis. The mean age at the time of death was 72 years (46 to 92). The mean VCA angle was 32.8 degrees (17.7 to 53.6). The SD was 7.9 degrees. Our findings suggest that the threshold of abnormality of the VCA angle may be slightly lower than previously thought. This information may be useful in counselling patients with asymptomatic acetabular dysplasia. PMID- 10855877 TI - The Baumann procedure for fixed contracture of the gastrosoleus in cerebral palsy. Evaluation of function of the ankle after multilevel surgery. AB - We treated 22 children (28 limbs) with diplegic cerebral palsy who were able to walk by the Baumann procedure for correction of fixed contracture of the gastrosoleus as part of multilevel single-stage surgery to improve gait. The function of the ankle was assessed by clinical examination and gait analysis before and at two years (2.1 to 4.0) after operation. At follow-up the ankle showed an increase in dorsiflexion at initial contact, in single stance and in the swing phase. There was an increase in dorsiflexion at initial push-off without a decrease in the range of movement of the ankle, and a significant improvement in the maximum flexor moment in the ankle in the second half of single stance. There was also a change from abnormal generation of energy in mid stance to the normal pattern of energy absorption. Positive work during push-off was significantly increased. Lengthening of the gastrocnemius fascia by the Baumann procedure improved the function of the ankle significantly, and did not result in weakening of the triceps surae. We discuss the anatomical and mechanical merits of the procedure. PMID- 10855878 TI - Toe-walking in children younger than six years with cerebral palsy. The contribution of serial corrective casts. AB - Our aim in this retrospective study was to analyse the value of serial corrective casts in the management of toe-walking in children aged less than six years with cerebral palsy. A total of 20 children (10 hemiplegic and 10 diplegic) had elongation of the triceps surae by serial casting at a mean age of four years and one month. The mean passive dorsiflexion of the foot with the knee in extension was 3 degrees (-10 to +5) and 12 degrees (0 to +15) with the knee in flexion. After removal of the cast passive dorsiflexion was 20 degrees (+10 to +30) with the knee in extension, and 28 (+10 to +35) with the knee in flexion. At a mean follow-up of 3.08 years (2.08 to 4.92), passive dorsiflexion was 9 degrees (-10 to +20) with the knee in extension and 18 degrees (0 to +30) with the knee in flexion. Serial corrective casts are useful for the treatment of equinus in young children as the procedure is simple and the results are at least equal to those of other non-operative techniques. It is a safe alternative to surgical procedures especially in young children. If the equinus recurs operation can be undertaken on a tendon which is not scarred. PMID- 10855879 TI - Instability of the hip in neonates. An ethnic and geographical study in 24,101 newborn infants in Malmo. AB - In a prospective study conducted between 1990 and 1997, 24 101 newborn infants were examined for neonatal instability of the hip and classified by the ethnic origin of their parents. In 63% their mother and father were of Swedish extraction and in 24% they were born in a foreign country. Those of foreign extraction were split into ethnic and geographical subgroups. Although the incidence of treated (dislocatable-unstable) hips was greater in Swedes (7.6/thousand), than in other geographical groups (5.8/thousand) it was not significantly different (p = 0.065). A total of 12.7/thousand were referred from the neonatal ward to the orthopaedic clinic with suspected dislocatable or unstable hips; 6.8/thousand were treated (5.4/thousand dislocatable, 1.4/thousand unstable), but 5.9/thousand were not treated since their ultrasound examination was normal. Two hips were diagnosed late and one case of mild avascular necrosis was found. Examination by dynamic ultrasound decreased the number of treated cases by 5.9/thousand but was not an absolute guarantee of diagnosis. PMID- 10855880 TI - Valgus extension osteotomy for 'hinge abduction' in Perthes' disease. Results at maturity and factors influencing the radiological outcome. AB - Valgus extension osteotomy (VGEO) is a salvage procedure for 'hinge abduction' in Perthes' disease. The indications for its use are pain and fixed deformity. Our study shows the clinical results at maturity of VGEO carried out in 48 children (51 hips) and the factors which influence subsequent remodelling of the hip. After a mean follow-up of ten years, total hip replacement has been carried out in four patients and arthrodesis in one. The average Iowa Hip Score in the remainder was 86 (54 to 100). Favourable remodelling of the femoral head was seen in 12 hips. This was associated with three factors at surgery; younger age (p = 0.009), the phase of reossification (p = 0.05) and an open triradiate cartilage (p = 0.0007). Our study has shown that, in the short term, VGEO relieves pain and corrects deformity; as growth proceeds it may produce useful remodelling in this worst affected subgroup of children with Perthes' disease. PMID- 10855881 TI - Acetabular dysplasia associated with hereditary multiple exostoses. A case report. AB - Hereditary multiple exostoses is an autosomal dominant disorder characterised by multiple osteochondromata, most commonly affecting the forearm, knee and ankle. Osteochondromata of the proximal femur have been reported to occur in 30% to 90% of affected patients with coxa valga in 25%. Acetabular dysplasia is rare but has been described. This is the first report of a patient requiring surgical intervention. A girl was seen at the age of nine with hereditary multiple exostoses and when 12 developed bilateral pain in the groin. Radiographs showed severely dysplastic acetabula with less than 50% coverage of the femoral heads and widening of the medial joint space. Large sessile osteochondromata were present along the medial side of the femoral neck proximal to the lesser trochanter, with associated coxa valga. The case illustrates the importance of obtaining initial skeletal surveys in children with hereditary multiple exostoses to identify potential problems such as acetabular dysplasia and subluxation of the hip. PMID- 10855882 TI - Reconstruction of part of the distal tibial growth plate with an autologous graft from the iliac crest. AB - We describe the reconstruction of a defect of the medial malleolus which was the result of an accident in a ten-year-old child. A graft from the iliac crest, with the apophyseal cartilage and perichondrium, was used for reconstruction of the medial malleolus, the growth plate and the adjacent metaphyseal defect, respectively. The soft-tissue defect was covered with a free scapular flap with microvascular anastomosis. Three years after the injury stability of the ankle is excellent with adequate growth of the reconstructed epiphyseal plate. PMID- 10855883 TI - Wound infection in hip and knee arthroplasty. AB - We have studied prospectively the outcome of wound discharge in patients after arthroplasty of the hip and knee. Over a period of 3.5 years 530 primary arthroplasties were carried out in one hospital. Postoperative wound infections developed in 82. At a mean follow-up of two years a comparison was made between these patients and 82 with healthy wounds, in terms of symptoms and signs of deep infection. There was an incidence of 1.1% of early deep infection, within six weeks in all cases. The rate of 'superficial' infection was 17.3% in the hips, 10.5% in the knees and 14.3% in total. At a mean follow-up of 26 months, there were no significant differences between the patients with infected wounds and a matched group of patients with healthy wounds in terms of the ESR, level of C reactive protein, white cell count and radiological scores, but clinical scores were significantly worse in the patients with infected knees (p < 0.05). The length of stay was also significantly longer in this group (mean 14.6 days in the healthy wound group, 19 days in the problem group; p < 0.005). There was, however, no convincing evidence that these wound infections led to deep infection and early revision in the early to medium follow-up period. A larger and longer prospective trial would be necessary to shed more light on this potential problem. PMID- 10855884 TI - Arthrodesis of flail or partially flail wrists using a dynamic compression plate without bone grafting. AB - Between June 1991 and May 1996 we carried out arthrodesis on 15 patients with flail or partially flail wrists using an AO/ASIF dynamic compression plate (DCP) without a bone graft. The wrist was approached through the second extensor compartment. The minimum follow-up was for 24 months with a mean of 34.2 months. All 15 wrists fused without major complications at a mean of 11.9 weeks. Stabilisation improved the function of the hand affected with paralysis and the appearance of the extensively paralysed upper limb with a flail hand. In the absence of bony abnormality fusion can be obtained with a DCP alone without the need for bone grafting. PMID- 10855885 TI - Giant-cell tumour of the tendon sheath. Is radiotherapy indicated to prevent recurrence after surgery? AB - Giant-cell tumour of the tendon sheath, also called pigmented villonodular synovitis, is a benign tumour with a high incidence of recurrence. We have tried to identify risk factors for recurrence. Of the 48 patients included in the study, 14 received radiotherapy after surgery. Only two (4%) had a recurrence. This compares favourably with previously reported incidences of between 25% and 45%. PMID- 10855886 TI - Talocalcaneal coalition. Diagnosis with the C-sign on lateral radiographs of the ankle. AB - We analysed 42 weight-bearing lateral radiographs of the ankle, 20 of which were from patients with a clinical and plain radiological diagnosis of talocalcaneal coalition (TCC) who subsequently had CT. The remainder were from 22 healthy volunteers with no clinical findings suggestive of hindfoot pathology. Four observers, blinded to the CT findings, independently evaluated the radiographs on two separate occasions. With the 95% confidence interval and using the CT findings as the comparison we calculated the sensitivity, specificity, accuracy, and positive and negative predictive values for the C-sign, and for other signs known to be associated with TCC. Similarly, we also calculated the interobserver and intraobserver reliability for these signs using the kappa statistic. Our results suggest that the C-sign is highly sensitive and specific for TCC. It is an accurate indicator and significantly more reliable than other previously recognised radiological signs of TCC. Features of the C-sign, however, cannot be relied upon to indicate whether the TCC is fibrous or bony. PMID- 10855887 TI - Reconstruction of the lateral ligaments of the ankle using solvent-dried and gamma-irradiated allogeneic fascia lata. AB - We have described a method of anatomical reconstruction of the lateral ligaments of the ankles with instability using allogeneic fascia lata dried with solvents and sterilised with gamma irradiation. Twenty ankles of 20 patients were assessed objectively and subjectively after a mean follow-up of 4.2 years (3.1 to 10). The result was excellent in 12 (60%), good in seven (35%) and fair in one (5%); none had a poor result. Stress radiography showed that the angle of talar tilt improved from 12.3+/-4.2 degrees (mean +/- SD) to 5.9+/-3.0 degrees and that the anterior drawer distance decreased from 9.2+/-3.9 mm to 4.4+/-2.5 mm. Neither infection nor limitation of movement occurred after operation. Fascia lata allografts provide a good alternative to autogenous grafts such as the peroneus brevis tendon. PMID- 10855888 TI - Haemorrhagic lumbar synovial cyst. A cause of acute radiculopathy. AB - A total of 254 cases of synovial cysts of the spine have been reported in the English literature, but only eight have been associated with haemorrhage. We describe a 55-year-old man with acute radiculopathy resulting from haemorrhage involving a synovial cyst at a lumbar facet joint. Traumatic factors could have caused bleeding around or into the synovial cyst. Treatment by resection of the cyst and evacuation of the haematoma led to complete neurological recovery. PMID- 10855889 TI - Neuromuscular effects of two types of fracture treatment. AB - Immobilisation causes denervation-like changes in the motor endplates, decreases the content of IGF-I, and increases the number of IGF-I receptors in the spinal cord. In the rat we investigated whether similar changes occur after a fracture of the midshaft of the femur which had been treated by intramedullary fixation with adequate or undersized pins. A more pronounced reduction in muscle wet weight was seen after fixation by undersized pins as well as decreased ash density of the ipsilateral tibia which did not completely return to normal within the 12-week experimental period. The nicotinic cholinergic receptors in the motor endplates of tibialis anterior were increased (p < 0.01) and there was a significant increase (p < 0.02) in IGF-I receptors in the lumbar spinal cord ipsilateral to the fracture after treatment by undersized nails. These changes may be associated with the impaired proprioception, co-ordination and motor activity which are sometimes seen after fractures. PMID- 10855890 TI - In-vivo strain measurements to evaluate the strengthening potential of exercises on the tibial bone. AB - Mechanical loading during physical activity produces strains within bones. It is thought that these forces provide the stimulus for the adaptation of bone. Tibial strains and rates of strain were measured in vivo in six subjects during running, stationary bicycling, leg presses and stepping and were compared with those of walking, an activity which has been found to have only a minimal effect on bone mass. Running had a statistically significant higher principal tension, compression and shear strain and strain rates than walking. Stationary bicycling had significantly lower tension and shear strains than walking. If bone strains and/or strain rates higher than walking are needed for tibial bone strengthening, then running is an effective strengthening exercise for tibial bone. PMID- 10855891 TI - The effects of the concentration of high-density polyethylene particles on the bone-implant interface. AB - We used a rat model in vivo to study the effects of the concentration of polyethylene particles on the bone-implant interface around stable implants in the proximal tibia. Intra-articular injections of 10(4), 10(6) or 10(8) high density polyethylene (HDPE) particles per joint were given 8, 10 and 12 weeks after surgery. The animals were killed after 14 and 26 weeks and the response at the interface determined. Fibrous tissue was seen at the bone-implant interface when the head of the implant was flush with the top of the tibia but not when it was sunk below the tibial plateau. In the latter case the implant was completely surrounded by a shell of bone. The area of fibrous tissue and that of the gap between the implant and bone was related to the concentration of particles in the 14-week group (p < 0.05). Foreign-body granulomas containing HDPE particles were seen at the bone-implant interface in animals given 10(8) particles. The pathology resembles that seen around prostheses with aseptic loosening and we suggest that this is a useful model by which to study this process. PMID- 10855892 TI - Physiological cell death of chondrocytes in vivo is not confined to apoptosis. New observations on the mammalian growth plate. AB - Chondrocytes at the lower zone of the growth plate must be eliminated to facilitate longitudinal growth; this is generally assumed to involve apoptosis. We attempted to provide definitive electron-microscopic evidence of apoptosis in chondrocytes of physes and chondroepiphyses in the rabbit. We were, however, unable to find a single chondrocyte with the ultrastructure of 'classical' apoptosis in vivo, although such a cell was found in vitro. Instead, condensed chondrocytes had a convoluted nucleus with patchy chromatin condensations while the cytoplasm was dark with excessive amounts of endoplasmic reticulum. These cells were termed 'dark chondrocytes'. A detailed study of their ultrastructure combined with localisation methods in situ suggested a different mechanism of programmed cell death. In addition, another type of death was identified among the immature chondrocytes of the chondroepiphysis. These cells had the same nucleus as dark chondrocytes, but the lumen of the endoplasmic reticulum had expanded to fill the entire non-nuclear space, and all cytoplasm and organelles had been reduced to dark, worm-like inclusions. Since these cells appeared to be 'in limbo', they were termed 'paralysed' cells. It is proposed that 'dark chondrocytes' and 'paralysed cells' are examples of physiological cell death which does not involve apoptosis. It is possible that the confinement of chondrocytes within their lacunae, which would prevent phagocytosis of apoptotic bodies, necessitates different mechanisms of elimination. PMID- 10855893 TI - Porous apatite-wollastonite glass-ceramic as an intramedullary plug. AB - We evaluated the efficacy and biocompatibility of porous apatite-wollastonite glass ceramic (AW-GC) as an intramedullary plug in total hip replacement (THR) for up to two years in 22 adult beagle dogs. Cylindrical porous AW-GC rods (70% porosity, mean pore size 200 microm) were prepared. Four dogs were killed at 1, 3, 6 and 12 months each and six at 24 months after implantation. Radiological evaluation confirmed the efficacy of porous AW-CG as an intramedullary plug. Histological evaluation showed osteoconduction at one month and resorption of the porous AW-GC, which was replaced by newly-formed bone, at 24 months. Our findings indicate that porous AW-GC can be used clinically as an intramedullary plug in THR. PMID- 10855894 TI - Percutaneous repair of ruptured tendo Achillis. PMID- 10855895 TI - The use of ultrasound in determining the initiation of treatment in instability of the hip in neonates. PMID- 10855896 TI - Chlorhexidine and chondrolysis in the knee. PMID- 10855897 TI - Smoking cessation in old age: closing the stable door? PMID- 10855898 TI - Cerebral and systemic pathophysiological responses to acute stroke. PMID- 10855899 TI - The electrocardiogram in heart failure. PMID- 10855900 TI - Carnosine: physiological properties and therapeutic potential. PMID- 10855901 TI - Influenza immunization coverage in older hospitalized patients during winter 1998 99 in Carmarthenshire, UK. AB - OBJECTIVE: to determine the coverage of influenza vaccination in hospitalized elderly patients in view of the 1998 recommendations of the UK Department of Health, and the reasons for refusal in those unvaccinated. DESIGN: questionnaire based interview. SETTING: acute elderly-care wards in a district general hospital in South Wales, UK. SUBJECTS: 443 consecutive patients aged over 65 hospitalized during December 1998 and January 1999. RESULTS: of 383 patients in whom influenza vaccination was recommended according to Department of Health guidelines, only 48% received it during the winter of 1998-99. The commonest reason given by those unvaccinated was lack of information from the general practitioner (in 26% of cases). Other reasons were concern about vaccine side effects (21%), perceived good health (16%) and concern about vaccine efficacy (11%). CONCLUSIONS: influenza vaccine uptake in high-risk older hospitalized patients is still unsatisfactory. Improved education of the health-care staff and the general public about the benefits of vaccine is necessary to improve uptake. PMID- 10855902 TI - IgE-mediated and age-related bronchial hyperresponsiveness in patients with asthma. relationship to family history of the disease. AB - OBJECTIVE: to uncover any differences in the age-related and IgE-mediated pathophysiology of the airways in asthmatics. METHODS: we examined the relationship of both IgE-mediated bronchial hyperresponsiveness and the cell content of bronchoalveolar lavage fluid with a family history of asthma in 263 patients with asthma classified according to age at onset. RESULTS: bronchial hyperresponsiveness decreased significantly as age at onset increased in those without a family history. Responsiveness was significantly higher in patients who were > or = 60 years of age at onset who had a family history than in those who did not (P < 0.05). The proportion of lymphocytes in bronchoalveolar lavage fluid was significantly higher in patients between 50 and 59 years old at onset who had a family history than those who did not (P < 0.05). These results suggest that bronchial hyperresponsiveness and the proportion of bronchoalveolar lavage lymphocytes differ according to the presence or absence of a family history, a finding which is closely related to IgE-mediated allergy in elderly patients at onset. CONCLUSIONS: our findings suggest (i) the possibility of asthma induced by non-IgE-mediated allergy in elderly patients and (ii) that bronchial responsiveness is also influenced by IgE-mediated allergy and age at onset. PMID- 10855903 TI - Correctable visual impairment in stroke rehabilitation patients. AB - BACKGROUND: after stroke, visual impairment may exacerbate the impact of other impairments on overall disability and negatively influence rehabilitation. OBJECTIVE: to examine the visual status of patients after stroke and determine whether this can be improved by simple interventions. DESIGN: prospective study. SETTINGS: stroke rehabilitation unit in a Belfast teaching hospital. SUBJECTS: 77 consecutive patients admitted for rehabilitation after acute stroke. METHODS: full optometric and ophthalmic assessment within 2 weeks of admission. RESULTS: of 70 patients with glasses, 19 did not have their glasses in hospital before prompting and 18 had glasses in unacceptable condition. Twenty patients had impaired visual acuity (6/12 or worse) with existing glasses (if helpful); 11 of these improved to better than 6/12 with refractive correction. CONCLUSIONS: stroke professionals need to enquire about patients' spectacles and assess their condition. Patients with reduced visual acuity in the absence of significant non refractive disease should be referred to an optometrist: in this series 14% of patients had visual impairment which benefited from refractive correction. PMID- 10855904 TI - Reliability of measurements of muscle tone and muscle power in stroke patients. AB - OBJECTIVES: to establish the reliability of the modified Ashworth scale for measuring muscle tone in a range of muscle groups (elbow, wrist, knee and ankle; flexors and extensors) and of the Medical Research Council scale for measuring muscle power in the same muscle groups and their direct antagonists. DESIGN: a cross-sectional study involving repeated measures by two raters. We estimated reliability using the kappa statistic with quadratic weights (Kw). SETTING: an acute stroke ward, a stroke rehabilitation unit and a continuing care facility. SUBJECTS: people admitted to hospital with an acute stroke-35 patients, median age 73 (interquartile range 65-80), 20 men and 15 women. RESULTS: inter- and intra-rater agreement for the measurement of power was good to very good for all tested muscle groups (Kw = 0.84-0.96, Kw = 0.70-0.96). Inter- and intra-rater agreement for the measurement of tone in the elbow, wrist and knee flexors was good to very good (Kw = 0.73-0.96, Kw = 0.77-0.94). Inter- and intra-rater agreement for the measurement of tone in the ankle plantarflexors was moderate to good (Kw = 0.45-0.51, Kw = 0.59-0.64). CONCLUSIONS: the Medical Research Council scale was reliable in the tested muscle groups. The modified Ashworth scale demonstrated reliability in all tested muscle groups except the ankle plantarflexors. If reliable measurement of tone at the ankle is required for a specific purpose (e.g. to measure the effect of therapeutic intervention), further work will be necessary. PMID- 10855905 TI - The unreliability of clinical measures of muscle tone: implications for stroke therapy. AB - BACKGROUND: the central tenet of the neurofacilitatory approach to stroke therapy is that muscle tone needs to be normal before normal movement can occur. A reliable clinical measure of the full spectrum of muscle tone is needed to test: (i) the purported relationship between muscle tone, other motor impairments and disability, and (ii) the effectiveness of stroke therapy to restore movement. AIM: the purpose of the study was to test the inter-rater reliability of clinical categorization of muscle tone (spastic/normal/flaccid) and also a visual analogue scale with anchor points of 'lowest tone possible' (score 0) and 'highest tone possible' (score 100). METHODS: four independent raters assessed tone of elbow flexors and knee extensors of 14 stroke rehabilitation inpatients using the categorical scale. Six independent raters assessed tone of elbow flexors and knee extensors of 25 chronic stroke patients and two healthy volunteers using the visual analogue scale. All assessment orders were randomized. RESULTS: both scales were unreliable, with K coefficients for the categorical scale ranging from -0.046 to 0.56 for the categorical scale, and intra-class correlation coefficients for the visual analogue scale of 0.595 for elbow flexors and 0.451 for knee extensors. Assessment order effects for the visual analogue scale were non-significant for elbow flexors (P= 0.545) and knee extensors (P= 0.911). CONCLUSIONS: these results, and those of earlier studies, suggest that clinical measures of muscle tone are consistently unreliable. Systematic investigation of the therapy rationale for planning and evaluating treatment is required before relevant clinical measures can be developed. PMID- 10855906 TI - Relationships between physical performance measures, age, height and body weight in healthy adults. AB - OBJECTIVE: we measured muscle strength and functional mobility in healthy men and women over the adult age range to investigate the changes with age and sex, and to establish the effects of the anthropometric indices height and weight. DESIGN: cross-sectional study. SUBJECTS AND METHODS: we recruited 74 healthy women (mean age 49.0, range 20-90) and 81 healthy men (mean age 51.6, range 20-90). We measured maximum isometric knee extension strength, handgrip strength and explosive leg extensor power. We assessed functional mobility quantitatively with the timed 'get up and go' test and the modified Cooper test. RESULTS: older subjects had lower values for muscle strength and muscle power than young subjects. Times for the timed 'get up and go' test were longer and distances in the modified Cooper test shorter. At about the age of 55, women showed an acceleration in the decline of isometric knee extension strength and handgrip strength (between 20 and 55 years, knee strength decreased by 10.3% and handgrip strength decreased by 8.2%, between 55 and 80 years the decreases were 40.2% and 28% respectively). Men showed a more gradual declines over the adult age range, with decreases in knee and handgrip strength of 24% and 19.6% between 20 and 55 years, and 23% and 17.4% between 55 and 80 years. The age-related decline is partly associated with differences in height and body weight. Women had higher correlations between muscle strength and functional mobility tests than men. CONCLUSIONS: muscle strength and functional mobility decline with age in healthy people; in women we observed an accelerated decrement in muscle strength above the age of 55. Lower values in healthy old subjects are partly associated with differences in height and body weight. PMID- 10855907 TI - The association between low diastolic blood pressure in middle age and cognitive function in old age. A population-based study. AB - BACKGROUND: previous longitudinal studies have shown an inverse relation between blood pressure and cognitive function. OBJECTIVE: to determine the association between mid-life blood pressure and performance in different areas of cognitive function in late life. SUBJECTS AND METHODS: we recruited 502 men, aged 69-74 years, from a population-based cohort in Uppsala, Sweden. Blood pressure had been measured at age 50 and we examined performance in 13 psychometric tests about 20 years later. RESULTS: after the 39 men with a previous stroke had been excluded, there was an inverse relation between diastolic blood pressure at age 50 and performance 20 years later in the digit span test, the trail-making tests and in verbal fluency. The relationships were significant, independently of age, education and previous occupational level. Men within the lowest category of diastolic blood pressure (< or = 70 mmHg, n = 59) showed the best results. Baseline blood pressure levels were not linked to performance in tasks on vocabulary, verbal learning and memory or figure copying. CONCLUSIONS: low blood pressure in mid-life indicates a low long-term cerebrovascular risk and is associated with higher late-life performance in cognitive tests that mainly assess subcortico-frontal cognitive functions. PMID- 10855908 TI - Age, costs of acute and long-term care and proximity to death: evidence for 1987 88 and 1994-95 in British Columbia. AB - BACKGROUND: the consequences of ageing populations for health care costs have become a concern for governments and health care funders in most countries. However, there is increasing evidence that costs are more closely related to proximity to death than to age. This means that projections using age-specific costs will exaggerate the impact of ageing. Previous studies of the relationship of age, proximity to death and costs have been restricted to acute medical care. OBJECTIVE: to assess the effects of age and proximity to death on costs of both acute medical care and nursing and social care, and to assess if this relationship was stable in a time of rapid change in health care expenditure. DESIGN AND METHODS: we compared all decedents in the chosen age categories for the years 1987-88 and 1994-95 with all survivors in the same age groups. We measured use of health and social care for each individual using the British Columbia linked data, and costs of care assessed by multiplying the number of services by the unit cost of each service. SETTING: the Province of British Columbia. SUBJECTS: all decedents in 1987-88 and 1994-95 in British Columbia in the chosen age groups, and all survivors in the same age groups. RESULTS: costs of acute care rise with age, but the proximity to death is a more important factor in determining costs. The additional costs of dying fall with age. In contrast, costs of nursing and social care rise with age, but additional costs for those who are dying increase with age. Similar patterns were found for the two cohorts. CONCLUSIONS: age is less important than proximity to death as a predictor of costs. However, the pattern of social and nursing care costs is different from that for acute medical care. In planning services it is important to take into account the relatively larger impact of ageing on social and nursing care than on acute care. PMID- 10855909 TI - Dependency in older people recently admitted to care homes. AB - OBJECTIVE: to investigate dependency and general health status of a cohort of older people admitted to residential or nursing homes for long-term care. METHOD: we assessed 308 people aged over 65 years within 2 weeks of admission for long term care to one of 30 nursing or residential homes in north-west England. Dependency was assessed using the Barthel activities of daily living index and the Crichton Royal Behaviour Rating Scale. We collected information from the homes' records on diagnosed conditions and current medication. RESULTS: 50% of the cohort were in a 'low dependency' band (Barthel score 13 - 20): 31% of those in nursing homes and 71% of those in residential homes. In nursing homes, low dependency residents were more likely to be self-funding than those with higher dependency. Of a number of broad diagnostic groupings, only a diagnosis of dementia was associated with nursing- rather than residential-home admission. Of 47 residents who scored 9 or less on the Mini-Mental State Examination (indicating severe cognitive impairment), 85% had no diagnosis of dementia, neurological disorder or other psychiatric disorder. DISCUSSION: the high proportion of new admissions of subjects with low dependency needs raises questions about the effective targeting of resources and about management of the boundary between home-based and institutional care. The existence of an important group of self-funded, low-dependency new admissions to nursing homes suggests a need to provide better assessment and placement services for those who are financially independent of local authorities. Many new admissions had conditions which might benefit from rehabilitation but there were almost no therapy staff in the studied homes. In some cases where severe cognitive impairment was evident, there was no evidence that the result of any formal pre-admission psychiatric evaluation had been communicated to nursing or care staff. PMID- 10855910 TI - An unusual cause of bruising in an 80-year-old woman. AB - PRESENTATION: a previously fit 80-year-old woman presented with a 2-week history of spontaneous and extensive bruising affecting all four limbs. The severity was such that she required a transfusion of 8 units of blood. RESULTS OF INVESTIGATIONS: a markedly prolonged activated partial thromboplastin time which was only partially corrected with normal plasma; tests for lupus anticoagulant were negative. Factor VIII levels were reduced and the Bethesda assay indicated an acquired inhibitor to factor VIII. She was treated with a combination of intravenous immunoglobulin and immunosuppression. OUTCOME: the response to treatment was excellent, with a marked reduction in anti-factor VIII antibody levels and resolution of the bruising over the next few weeks. PMID- 10855911 TI - Do patient age and medical condition influence medical advice to stop smoking? AB - OBJECTIVE: to determine whether the age and medical condition of a patient influences hospital-based doctors' decision making when advising patients to stop smoking cigarettes. METHODS: we presented 142 doctors from four grades (consultant, registrar, senior house officer and house officer) and four specialities (medicine, surgery, psychiatry and anaesthetics), based in a Dublin teaching hospital, with 20 clinical vignettes. Each vignette described a patient from one of five age groups with one of four levels of health. The vignettes were randomly mixed. We asked doctors to say whether they would advise the patient in each case to quit smoking. RESULTS: hospital-based doctors are significantly less likely to advise patients aged over 65 years than younger patients of the hazards of cigarette smoking, irrespective of the person's physical or mental health (P < 0.001). CONCLUSION: the advice given to patients about their cigarette smoking habits by hospital doctors is strongly influenced not only by the patient's health, but also by the patient's chronological age. PMID- 10855912 TI - Association of individual activities of daily living with self-rated health in older people. AB - OBJECTIVE: to evaluate the associations of 18 activities of daily living with self-rated health in older people. DESIGN AND SETTING: cross-sectional study of a representative sample of 781 people aged 65 or over (response rate: 89.9%). METHODS: self-rated health was assessed by the question: "Overall, how would you rate your current health status-very good, good, fair, poor or very poor?" We used the Barthel index and Lawton and Brody's index for basic and instrumental activities of daily living, respectively. We classified subjects into three groups according to their Barthel index score: level 1 (score 100), level 2 (score 91-99) and level 3 (score 0-90). Logistic regression was used to identify associations between each activity and self-rated health. RESULTS: use of stairs [odds ratio (OR) = 4.28, 95% confidence interval (95% CI): 2.82-6.52], ambulation (OR = 3.67, 95% CI: 2.39-5.64) and chair/bed transfer (OR = 3.00, 95% CI: 1.68 5.36) were the basic activities of daily living best associated with self-rated health. Among instrumental activities of daily living, ability to handle finances (OR = 2.20), laundry (OR = 2.15) and transport (OR = 2.12) were associated with self-rated health. On the Barthel index, only transport was associated with self rated health in subjects at levels 1 (OR = 2.55) and 2 (OR = 2.72). For subjects with poor functional status (level 3), no instrumental activities of daily living were related to self-rated health. CONCLUSION: in terms of self-rated health, the most important activities of daily living were those involving mobility. The effect of each instrumental activity of daily living on self-rated health depends on the level of functional capacity in basic activities of daily living. PMID- 10855913 TI - The effects of supporting discharge from hospital to home in older people. AB - OBJECTIVE: to investigate the effects of supported discharge after an acute admission in older people with undifferentiated clinical problems. DESIGN: a systematic review of randomized controlled trials. METHODS: we searched MEDLINE, CINAHL, the Cochrane Library, PsycLit and the Social Science Citation Index up to the end of 1997. This was augmented by hand-searching, follow-up of bibliographies and.direct enquiry of authors of included studies. Application of inclusion decisions, quality assessment and data abstraction were carried out independently by at least two of the reviewers. We tabulated the results of the included studies and used meta-analysis where appropriate to refine conclusions. RESULTS: we finally included nine studies in the review, assessment of which revealed that bias was present, dictating the need for caution in interpreting results. Despite this, there was relative certainty that the proportion of those at home 6-12 months after admission is greater with supported discharge (odds ratio 1.4, 95% confidence interval 1.1- 2.0). This was associated with a consistent pattern of reduction in admission to long-stay care over the same period, without apparent increases in mortality. There was uncertainty about the effect of supported discharge on hospitalization. There were no rigorous research data on functional status, patient and carer satisfaction, and, in consequence, uncertainty about the overall effectiveness of supported discharge. CONCLUSIONS: we believe that the results of this review provide reassurance that supporting discharge from hospital to home is of value. However, important sources of uncertainty remain, suggesting the need for further research. PMID- 10855914 TI - Respiratory disease in old age: research into Ageing Workshop, London, 1998. PMID- 10855915 TI - Osteoporosis management in elderly subjects--a UK survey of geriatricians. PMID- 10855916 TI - Chronic obstructive pulmonary disease and depression: analysis of depressive symptoms. PMID- 10855917 TI - Doctors' training in manual handling. PMID- 10855918 TI - Does everyone in heart failure need echocardiography? PMID- 10855919 TI - Does everyone in heart failure need echocardiography? PMID- 10855920 TI - Comprehensive integrated primary mental health care for South Africa. Pipedream or possibility? AB - While the vision for restructuring health care in South Africa is based on a comprehensive primary health care system, care at the primary level remains largely biomedical in orientation. Given this, I argue that whilst adding mental health care to primary level care may increase accessibility of psychiatric care. it will not, however, provide for comprehensive integrated primary mental health care as planned. This would require a paradigm shift towards a comprehensive discourse of care which includes mental health care. While efforts towards reorienting health care personnel in South Africa towards the primary health care approach have been initiated, an examination of the primary health care system in one sub-district in South Africa, reveals that the delivery of biomedical care is sustained by a number of factors within the primary health care system as well as within the macro-context. A shift in the paradigm of care provided would therefore require the transformation of the system on many fronts. Of central importance would be the restructuring of the primary health care system to be supportive of emotional labour, health promotion, empowerment of service users and of care which takes the subjectivity of the illness experience for the patient into account. PMID- 10855921 TI - When providers and community leaders define health priorities: the results of a Delphi survey in the canton of Geneva. AB - The Delphi method was used to determine the health priorities in one Swiss canton. The opinion of various groups concerned, either as health professionals or as representatives of the general population, was gathered to identify the health determinants and health problems perceived as most important, to clarify the reasons for these choices, and to recommend interventions to be undertaken in order to improve the situation in the identified priority areas. Five panels, including health professionals as well as selected leaders of community groups with no direct involvement in health, were given the opportunity to reply to two rounds of questionnaires. There was a high convergence of opinion on health determinants and problems to be given priority between panels and between the first and second round. Priorities identified are mainly physical problems (cardiovascular disease, respiratory and breast cancer, AIDS, injuries due to road accidents, chronic back pain), psychosocial disorders (depression, suicide, violence in the family, stress), and problems of substance abuse (alcohol and tobacco). Unemployment and social isolation were chosen because of their perceived impact on health. Very few interventions were proposed in the medical technical or research areas. This may be due partly to the fact that good quality care is widely available and accessible in Geneva, whereas preventive programmes have not received enough attention in the past. Through the identified priorities and the proposed activities, a new vision of health emerges which gives more importance to psychosocial problems and the social environment. In this context, health promotion is seen as essential, acknowledging that sustained change in individual behaviours can only occur if the social and cultural context is taken into consideration. In conclusion, the results of this survey show that the Delphi method is a useful tool to reach consensus on health priorities and corresponding activities among a variety of actors. PMID- 10855922 TI - Appropriateness measurement: application to advice-giving in community pharmacies. AB - Awareness of variations in the delivery of medical care has resulted in considerable research activity focused on developing measures to assess the appropriateness of health service provision both internationally and within Great Britain. As in other areas of health service provision there is evidence of variation in advice provided alongside sales of non-prescription medicines and variation in response to requests for advice about the treatment of minor ailments within community pharmacies in Great Britain. However, there is little research which has explored the extensive methodological problems associated with developing criteria to assess the appropriateness of these-two activities. Following a critical review of relevant existing research, this paper describes a methodology and empirical findings from a study which aimed to develop criteria to measure the appropriateness of advice provided in community pharmacies. Firstly, details of advice-giving episodes occurring between consumers and pharmacists or medicines counter assistants were captured and documented using a combination of audio tape-recording and non-participant observation. Secondly, the nominal group technique was used to develop a set of explicit criteria for assessing the appropriateness of advice. Thirdly, an assessment instrument was developed in order to operationalise the criteria. The devised criteria include both process and output components. We discuss the utility of these criteria in relation to developments in self-medication practice affecting community pharmacy and the deregulation of medicines within the UK. The criteria have been subject to rigorous statistical testing to establish standards of validity and reliability (Ward, Bissell & Noyce, 2000a [Ward, P. R., Bissell, P. & Noyce, P. R. (2000a). Criteria for assessing non-prescription drug therapy in community pharmacy, Annals of Pharmacotherapy (in press).]). The developed criteria will allow us to identify dimensions of both appropriate and inappropriate advice provided in community pharmacies and provide the basis for education and training initiatives identified as a result of the research. In addition, we suggest that this research is highly relevant to informing the content, structure and operationalisation of protocols and/or guidelines associated with the management of minor ailments and the sale of medicines through community pharmacies. PMID- 10855923 TI - Gender, socioeconomic development and health-seeking behaviour in Bangladesh. AB - In efforts to reduce gender and socioeconomic disparities in the health of populations, the provision of medical services alone is clearly inadequate. While socioeconomic development is assumed important in rectifying gender and socioeconomic inequities in health care access, service use and ultimately, outcomes, empirical evidence of its impact is limited. Using cross-sectional data from the BRAC-ICDDR,B Joint Research Project in Matlab, Bangladesh, this paper examines the impact of membership in BRAC's integrated Rural Development Programme (RDP) on gender equity and health-seeking behaviour. Differences in health care seeking are explored by comparing a sample of households who are BRAC members with a sample of BRAC-eligible non-members. Individuals from the BRAC member group report significantly less morbidity (15-day recall) than those from the non-member group, although no gender differences in the prevalence of self reported morbidity are apparent in either group. Sick individuals from BRAC member households tend to seek care less frequently than non-members. When treatment is sought, BRAC members rely to a greater extent on home remedies, traditional care, and unqualified allopaths than non-member households. While reported treatment seeking from qualified allopaths is more prevalent in the BRAC group, non-members use the para-professional services of community health care workers almost twice as frequently. In both BRAC member and non-member groups, women suffering illness report seeking care significantly less often than men. The policy and programmatic implications of between group and gender differences in care seeking are discussed with reference to the literature. PMID- 10855924 TI - Violence and fear of violence in East and West Germany. AB - The purpose of this research was to assess the effect of major social changes in Germany since 1989 on mortality due to intentional injury. Mechanisms and types of fatal intentional injury in East and West Germany between 1970 and 1995 were determined from death certificates and compared with judicial data on violent crime convictions and recent public survey data on citizen fear of crime. The number of homicides among East German males increased between 1989 and 1991, and the homicide rate remains high when compared with West German males (although lower than that of American males). Homicide among German females is less common, presently about equally likely in East and West. Violent crime in general has become more frequent in Germany, and citizen fear of crime has increased markedly, especially in the East. Non-citizens are convicted for an increasing number of homicides and assaults. Rates of suicide were declining in East and West before reunification, and these rates have continued to decline. Social changes in Europe since 1989 have led to noticeable increases in violence and homicide in Germany, which in turn have reduced feelings of security among German citizens, especially in the East. Suicide rates have not been affected. PMID- 10855925 TI - Self-identity in older persons suffering from dementia: preliminary results. AB - In this study, we explored the role-identity of nursing home residents suffering from dementia, as well as the potential for utilizing their enduring sense of self-identity for enhancing their quality of life. Four types of role-identity were explored: professional, family-role, leisure activities, and personal attributes. The methodology included structured interviews and a case study. Participants for the interviews were 38 residents of two nursing homes in Israel. Residents, relatives, and staff members were interviewed to provide information about past roles and the degree to which those roles are maintained in the present, and about strategies for bolstering the sense of self-identity. A large range of roles were identified. All role identities deteriorated significantly, with family roles retaining the greatest prominence in the present. However, much heterogeneity was manifested in all roles. Both staff members and relatives felt that a sense of identity in residents could be enhanced in most of the residents, which would exert a beneficial effect on their well-being. Caregiving respondents anticipated that this improvement would be substantial for about half of the residents. The case study illustrates how self-identity can change throughout dementia, and how it can be utilized to improve quality of life. PMID- 10855926 TI - Money illusion among health care providers: should we adjust for inflation in analyses of provider behavior? AB - This analysis questions the appropriateness of inflation adjustment in analyses of provider behavior by comparing results from estimations using adjusted financial variables with those from estimations using unadjusted financial variables. Using Medicaid claims from 1984-1991, we explored the effects of Medicaid reimbursement increases on dentists' participation. Using results from inflation adjusted analyses, we would conclude that a 23% nominal increase in Medicaid reimbursement rates yields no increase in the number of Medicaid children seen by dentists. In contrast, estimations based on unadjusted reimbursement rates suggest that this same 23% nominal increase in reimbursement leads to an expected 16-person (15.4%) increase in the number of Medicaid patients seen per provider per year. These analyses demonstrate that results are sensitive to adjustment for inflation. While adjusting for inflation is a generally accepted practice in health services research, doing so without evidence that providers respond to adjusted reimbursement may be unjustified. More research is needed to determine the appropriateness of inflation adjustment in analyses of provider behavior, and the circumstances under which it should or should not be done. PMID- 10855927 TI - Discounted lives? Weighing disability when measuring health and ruling on "compassionate" murder. AB - This paper examines the politics of "suffering" by considering the Disability Adjusted Life Year (DALY) alongside a controversial Canadian murder case, involving the killing of a child with disabilities by her father. The DALY aims to measure health and death correctly so as to allocate resources fairly, and eventually achieve better living conditions among the world's people. The Latimer controversy centres on the contention that some lives are not worth living, which the DALY's formula also implies. By ranking types of people according to their degree of disability, the DALY rates the lives of some people as worse than "a state equivalent to death". By examining the politics of "suffering" in the DALY and the Latimer affair, this paper underlines a valorisation of the "normal" body in much of the social science literature on health, medicine and suffering. PMID- 10855928 TI - Life quality vs the 'quality of life': assumptions underlying prospective quality of life instruments in health care planning. AB - Quality of Life is a broad construct used in health planning, health economics, and medical decision-making. It is also a term that has a long currency in social and sociological literatures. This paper considers the assumptions underlying prospective QL instruments in an historical and contemporary context. It argues that as a tool in health planning and in clinical decision making life quality as a measurement has its origins in the early eugenics literature and the social policies that derived from it in first North America, the primary focus of this paper, and later in Europe. Reference to narrative and social literatures, as well as those involving coping and adaptation, are then used to critique the assumptions underlying this class of QL instruments. It concludes that to the degree now current prospective instruments reflect a purely physical perspective of "disease burden" irrespective of social conditions they create a context that works against life quality, and in some cases, the continuance of persons with physical differences. PMID- 10855929 TI - Consultation with another physician on euthanasia and assisted suicide in the Netherlands. AB - Consultation with another physician is considered to be an important safeguard of the practice of euthanasia and physician-assisted suicide. The objective is to describe the frequency and characteristics of consultation in cases of euthanasia or physician-assisted suicide (EAS) in the Netherlands. Data from two cross sectional descriptive nationwide surveys, carried out in 1995, were used. Questionnaires were mailed to physicians attending 6060 deaths, identified from death certificates, and a stratified sample of 405 physicians were interviewed. In 1990, a cross-sectional descriptive postal survey of a random sample of 1042 general practitioners took place. Consultation took place in 63% of cases of EAS in the Netherlands, in 99% of the cases reported to the public prosecutor and in approximately 37% of unreported cases. In almost half of the unreported cases the decision had been discussed less formally with at least one colleague. In 1990, 7% of general practitioners met all 8 criteria for good consultation; this increased to 64% in 1995. Of the respondents, 26% had at some time advised against performing euthanasia or assisted suicide when acting as a consultant. This study shows that approximately two thirds of all cases of EAS are safeguarded by consultation. Although in the majority of these cases the consultation is of good quality, there is certainly still room for improvement. The quality of consultation could be improved, for instance, by appointing independent and specifically trained consultants. PMID- 10855930 TI - The understanding of their illness amongst people with irritable bowel syndrome: a Q methodological study. AB - Irritable Bowel Syndrome (IBS) refers to a collection of gastrointestinal symptoms which affect up to 22% of the Western population. Although the disorder costs the British National Health Service and employers vast sums of money in terms of repeated physician visits, medications, and loss of productivity, the cause or causes of IBS are still unknown, and there is no cure which is lastingly effective. Since IBS is not life-threatening, and the symptoms can be hidden from others, many consider it a trivial disorder. For an individual with IBS, however, the uncertainty regarding cause, diagnosis and treatment may lead to anxiety and constant searching for causes, or to hopelessness and resignation. The present study aims to help clarify these problems by discovering how those who suffer from IBS understand the nature and causality of their own illness. Through use of Q methodology with a sample of 60 people with IBS, a taxonomy of 7 clear and distinct accounts is identified and described. These data (based on Q factor analysis) are described in qualitative detail and discussed in relation to the problem of improving communication with doctors, and untangling issues of responsibility for illness. PMID- 10855931 TI - Information presentation and decisions to enter clinical trials: a hypothetical trial of hormone replacement therapy. AB - We examined recruitment to an imaginary trial of hormone replacement therapy (HRT) following two different styles of information about HRT. We predicted that for treatments which, like HRT, are available outside a trial, people offered the facts as currently known would be less likely to remain unsure about the relative costs and benefits, and so less likely to agree to enter a randomised trial. In contrast, when the information provided reflected the current state of uncertainty which justified the trial, we predicted that people would be less likely to form a preference for one treatment arm over the other, and so more likely to agree to enter a trial. One hundred women aged 25-40 years were informed about HRT via a video and an information leaflet. For half the participants the information was framed in a way which emphasised the current state of uncertainty about the relative costs and benefits of HRT, and in that respect it reflected the justification for a trial. This version was considered to be similar in style to information commonly provided to potential trial participants. For half the participants the same information was framed in a way which offered explicit numerical detail about currently known facts, and in that respect it was considered to be similar in style to information commonly available to doctors prior to a trial. Women learned as much about HRT in the two conditions, but women given the explicit versions were more likely (i) to hold a stronger view about whether or not they would take HRT (ratings were not elicited from the first 30 participants in this condition. N = 20, p < 0.05 1 tailed) and (ii) to refuse entry to the trial (N = 50, p < 0.05 2 tailed). Those who, given the explicit version, agreed rather than refused to enter the trial, scored higher on believing that others control their health (p < 0.01 2 tailed). PMID- 10855932 TI - Patient education literature and help seeking behaviour: perspectives from an evaluation in the United Kingdom. AB - Decisions by patients upon when to use health care services are a major influence on the consumption of health care resources. Patient education--often based upon written information on how to identify symptoms of common illnesses, when to seek help and how to self-treat--is an increasingly popular strategy to rationalise demand. A large body of literature, in evaluating the impact of such written information, has though overlooked the possession or acquisition of comparable publications by respondents in the course of the studies. This study attempted to overcome this limitation in considering the impact of a prominent patient education booklet that makes reference to over 40 common ailments. Questionnaire data from a cohort sample of residents (n = 495) in an area within which the booklet was circulated and that from a comparison area (n = 509) suggests that such literature exerts a modest influence in orienting patients towards "appropriate" self-referral and self-care behaviour. Reasons for this limited impact emerged however in semi-structured interviews with a sub-sample of respondents (n = 85). These data show that understanding of the way in which written advice for patients is perceived has to focus upon the ways in which diverse sub-populations process and attribute meaning to "official" and "unofficial" sources of advice. More fundamentally, the increasingly sophisticated and specialised nature of medical and scientific knowledge may be distancing expert knowledge from individuals and society such that "lay" responses to "expert" advice now reflect a continuing process of risk assessment, trust or the withholding of trust. PMID- 10855933 TI - ANCA glomerulonephritis and vasculitis: a Chapel Hill perspective. AB - Microscopic polyangiitis, Wegener's granulomatosis, Churg-Strauss syndrome, and pauci-immune necrotizing glomerulonephritis share pathogenic, pathological, and clinical features. They all involve capillaries, venules, arterioles, and small arteries. Approximately 90% of patients have autoantibodies either to myeloperoxidase (MPO-ANCA) or to proteinase 3 (PR3-ANCA). The clinical manifestations of ANCA-small vessel vasculitis are protean. These can be limited to the kidney alone, or may involve the upper respiratory tract, the lungs, the skin, or a number of other organs in various combinations. The characteristic feature of the glomerular lesion is a focal necrotizing glomerulonephritis associated with crescent formation and little or no glomerular staining for immunoglobulin by immunofluorescence microscopy. The renal manifestations can present as a rapidly progressive glomerulonephritis or that of a more indolent, remitting, and relapsing course that leads to substantial glomerulosclerosis. The two main prognostic markers of the long-term outcome are the presence of pulmonary hemorrhage (which accounts for at least half of all deaths) and the entry serum creatinine. The higher the entry serum creatinine, the higher the risk of developing end-stage renal disease. The treatment of ANCA-small vessel vasculitis and glomerulonephritis rests primarily on the use of induction high dose corticosteroids and cyclophosphamide. Patients with pulmonary hemorrhage also benefit from plasmapheresis. With the use of an alkylating agent, the rate of remission is of the order of 75%, but relapses occur in about 30% of patients who achieve a remission, and in about 17% of patients after renal transplantation. Despite the improved outcome of patients with ANCA vasculitis in the recent decade, their long-term prognosis continues to be primarily determined by a rapid diagnosis, and the prompt institution of therapy. PMID- 10855934 TI - Treatment of glomerular diseases: ANCA-negative RPGN. AB - Classification of rapidly progressive glomerulonephritis (RPGN) has evolved over the last decade into a variety of categories, some are which are more amenable to treatment than others. The advent of testing for antineutrophilic cytoplasmic antibody (ANCA) has further defined RPGN and aided in the diagnosis and treatment of these diseases. Although RPGN has traditionally carried a poor prognosis, this has been markedly improved by early diagnosis and intervention with aggressive therapy. The current chapter provides an oversight of ANCA-negative RPGN, and delineates an approach to diagnosis and therapy of various subtypes of this entity. Although current therapeutic options still encompass broad-spectrum immunosuppressive modalities, the future holds great hope for specific directed interventions at discrete points in the pathogenic progress. This offers the possibility of abrogation of the immune response leading to RPGN with less toxic general side effects then with treatments currently used. The next millennium will be marked by improved prognosis for ANCA-negative glomerulonephritis compared with that observed with this devastating disease in past decades. PMID- 10855935 TI - Treatment of glomerulonephritis in the elderly. AB - With the increasing use of renal biopsy in the elderly, glomerulonephritis is now known to be a common finding. Whereas membranous glomerulonephritis and minimal change disease are common in younger and older adults, primary amyloidosis and crescentic glomerulonephritis are more common in the elderly. Other glomerulonephritides such as focal segmental glomerulosclerosis or IgA nephropathy are very uncommon in the elderly. Because of the serious consequences of the nephrotic syndrome and acute and chronic renal failure in the elderly, aggressive treatment with immunosuppression should not be withheld. Caution should always be taken because of the presumed greater morbidity and mortality from such treatment in the elderly. PMID- 10855936 TI - Treatment of lupus nephritis. AB - Patients with lupus nephritis pose a therapeutic challenge and stimulate investigation of innovative treatment strategies. Although patient survival and renal function outcomes have improved over the last 4 decades, contemporary immunosuppressive regimens are not consistently effective and often require extended courses associated with insidious toxicities. Several strategies are under investigation to induce remissions more rapidly and to reduce the risk of long courses of cytotoxic drug therapy. The combination of pulse methylprednisolone and pulse cyclophosphamide may be more effective than pulse cyclophosphamide alone for patients with relatively severe proliferative lupus nephritis. Ongoing clinical studies evaluate the risk/benefit of other intensive induction regimens (eg, combination fludarabine with relatively low-dose pulse cyclophosphamide). A particularly vigorous strategy employs immunoablative cyclophosphamide with or without stem cell rescue. Several studies of sequential immunosuppressive therapy are in progress. It is anticipated that long-term toxicities can be lessened by substituting various maintenance agents (eg, azathioprine or mycophenolate mofetil) after initial cyclophosphamide therapy has induced a renal response. Additional information is needed to determine the role of this strategy. Furthermore, a number of standard and experimental immunosuppressive regimens (that do not include cyclophosphamide) are under investigation as well. Innovative approaches (eg, costimulatory blockade) offer the hope of more effective treatments without the risks of contemporary regimens. PMID- 10855937 TI - Treatment of IgA nephropathy. AB - IgA nephropathy (Berger's disease) is the most common primary glomerulonephritis worldwide and was once equated with benign recurrent hematuria. Longer observation showed that 15% to 30% of patients progress to end-stage renal failure after 20 years of clinical manifestations. Because the pathogenesis remains enigmatic, therapy to ameliorate disease progression can not be disease specific. Several approaches to treatment have generated increasing interest in the last few years, including angiotensin inhibition, glucocorticoids, fish oil, cyclophosphamide, tonsillectomy and mycophenolate mofetil. For patients reaching end-stage renal failure, recurrent disease after transplantation remains a clinically important problem, despite immunosuppression since engraftment. PMID- 10855938 TI - Treatment of hepatitis C-associated glomerular disease. AB - Hepatitis C virus (HCV) infection can lead to chronic active hepatitis, cirrhosis, and liver failure; however it is also associated with a wide range of extrahepatic features. Renal manifestations include cryoglobulinemic membranoproliferative glomerulonephritis and membranous nephropathy. Treatment of HCV with alpha-interferon is only moderately effective and suffers from a high relapse rate. More recently, combination therapy with ribavirin has led to improved suppression of HCV RNA levels. In this review we briefly describe the features of renal disease associated with HCV infection and discuss the therapeutic options. PMID- 10855939 TI - Treatment of HIV-associated nephropathy. AB - HIV-Associated Nephropathy (HIVAN) is the most common cause of chronic renal disease in HIV-1 infected patients. The disease occurs predominantly in blacks between the ages of 20 and 64. In this population it is currently the third leading cause of end-stage renal disease. The majority of patients with HIVAN have an AIDS-defining condition when the kidney disease is diagnosed. Without treatment they progress to end-stage renal disease within weeks to months. Patients with HIVAN should be treated with highly active antiretroviral therapy (HAART). Treatment should prolong survival and may improve or stabilize kidney function. Steroids have short-term benefits but long-term benefits have not been shown. Converting enzyme inhibitors (CEI) seem to stabilize kidney function and appear to be most effective when administered early in the course of HIVAN. A randomized controlled trial comparing HAART therapy to HAART and CEI should be performed. PMID- 10855940 TI - The treatment of idiopathic membranous nephropathy. AB - Membranous nephropathy remains the most common cause of the nephrotic syndrome in adults. Most patients do well with long-term natural history studies reporting a 10-year renal survival of 70% to 90% but the remainder progress to end stage renal failure. This plus the associated morbidity of those with persistent high grade proteinuria makes the decision about the timing and type of treatment difficult. Models have been developed to help predict at an early stage of the disease those at the highest risk of progression and their use is encouraged. The use of nonspecific, nontoxic therapy, ie, angiotensin-converting enzyme inhibitor (ACEI), for both hypertension control and their renoprotective effect is supported by evidence from high-quality studies. Modest dietary protein restriction may be of use but its effect is more controversial. If subnephrotic proteinuria plus normal renal function is present or inducible, conservative therapy and ongoing observation is probably all that is warranted. If high-grade proteinuria (>3.5 g/d) persists then the Italian regime consisting of cytotoxic therapy alternating monthly with prednisone treatment for three cycles has shown the best evidence of long-term induction of remission of proteinuria and preservation of renal function. If this fails or is judged too toxic then a 6- to 12-month course of cyclosporine seems warranted, especially if renal function is deteriorating. Introduction of treatment for risk reduction of both secondary effects of the disease and for modification of the adverse effects of immunosuppressive drugs should be considered in cases with high-grade persistent proteinuria. PMID- 10855941 TI - Treatment of focal segmental glomerulosclerosis. AB - Focal segmental glomerulosclerosis (FSGS) has been increasing in incidence over the past 2 decades and may currently be the most common form of primary nephrotic syndrome in the United States. Nephrotic patients with FSGS who do not achieve a remission in proteinuria usually advance to end-stage renal disease within 5 to 10 years. Although initially felt to be a steroid-resistant disease, especially in adults, studies show significant responsiveness to more prolonged courses of steroids. For patients with steroid-resistant or steroid-dependent FSGS, cyclosporine A and cytotoxic agents have shown efficacy in clinical trials. Other agents used include pulse methylprednisolone, azathioprine, tacrolimus, mycophenolate mofetil, and combination therapy. For recurrent FSGS after renal transplantation, plasmapheresis is often used but appears not to be as efficacious in adults as in the pediatric population. PMID- 10855942 TI - Lung opioid receptors: pharmacology and possible target for nebulized morphine in dyspnea. AB - Opioid receptors are located throughout the respiratory tract. Yet, these have received relatively scant attention compared to other opioid receptors. The most abundant sites within the respiratory tract appear localized within the alveolar walls, other sites appear to line the smooth muscle within the trachea and main bronchi near the lumen. There is about 100-times greater [3H]morphine binding density within the bronchioles and lobes than in the main bronchi or trachea. In addition to the usual mu, delta and kappa types of opioid receptors, 'non conventional' opioid binding sites have been suggested, although the function of these or of the other opioid receptors in the pulmonary tract is not known. However, they might explain the otherwise counterintuitive apparent utility of morphine treatment of dyspnea. Dyspnea is a common and distressing symptom in terminally-ill cancer patients and patients with chronic lung disease. It results from multiple causes, is difficult to treat and is a significant precipitating factor for late-stage hospital or hospice admissions. Nebulized morphine or other opioids have been reported to have beneficial effect, but the mechanism by which opioids might produce this seemingly contradictory effect is not clear. We review here lung opioid receptor distribution, pharmacology and possible clinical relevance in the treatment of dyspnea. PMID- 10855943 TI - Possible involvement of 5-HT and 5-HT2 receptors in acceleration of gastrointestinal transit by escin Ib in mice. AB - We have reported previously that escin Ib accelerated gastrointestinal transit (GIT) in mice, and that its effect may be mediated by the release of endogenous prostaglandins (PGs) and nitric oxide (NO). In this study, the possible involvement of 5-HT and 5-HT receptors in the GIT acceleration of escin Ib was investigated in mice. The acceleration of GIT by escin Ib (25 or 50 mg/kg, p.o.) was attenuated by pretreatment with ritanserin (0.5-5 mg/kg, s.c., a 5 HT(2A/2C/2B) receptor antagonist), but not with MDL 72222 (1 and 5 mg/kg, s.c.) and metoclopramide (10 mg/kg, s.c.) (5-HT3 receptor antagonists) or tropisetron (1 and 10 mg/kg, s.c., a 5-HT(3/4) receptor antagonist). Furthermore, pretreatment with ketanserin (0.05-5 mg/kg, s.c.), haloperidol (1-5 mg/kg, s.c.) and spiperone (0.5-5 mg/kg, s.c.) (5-HT2A receptor antagonists), as well as a bolus of dl-p-chlorophenylalanine methyl ester (PCPA, 1000 mg/kg, p.o., 1, 6 or 24 h before administration of the sample) (an inhibitor of 5-HT synthesizing enzyme tryptophan hydroxylase) and reserpine (5 mg/kg, p.o.) (a 5-HT depletor), but not 6-hydroxydopamine (80 mg/kg, i.p., a dopamine depletor) or repeated PCPA (300 mg/kg x2, p.o., 72 and 48 h before administration of the sample), also attenuated the effects of escin Ib. It is postulated that escin Ib accelerates GIT, at least in part, by stimulating the synthesis of 5-HT to act through 5-HT2, possibly 5-HT2A receptors, which in turn causes the release of NO and PGs. PMID- 10855944 TI - The spinal antinociceptive effect of FR140423 is mediated through kyotorphin receptors. AB - We investigated the antinociceptive effect of a novel anti-inflammatory and analgesic drug, 3-(difluoromethyl)-1-(4-methoxyphenyl)-5-[4 (methylsulfinyl)phenyl]pyraz ole (FR140423), in the tail-pinch test in mice, and evaluated the mechanism of action of FR140423 using L-leucyl-L-arginine (Leu Arg), a kyotorphin (endogenous Met-enkephalin releaser) receptor antagonist, L-NG nitroarginine methylester (L-NAME), an inhibitor of nitric oxide (NO) synthase, and methylene blue (MB), an inhibitor of activation of guanylate cyclase. Oral administration of FR140423, at doses of 5-80 mg/kg, produced a dose-dependent antinociceptive effect with an ED50 value of 18 mg/kg. This antinociception was reversed by intrathecal (i.t.) (10 microg/mouse), but not by intracerebroventricular (i.c.v.) (100 microg/mouse), injection of Leu-Arg. Moreover, the antinociceptive effect of i.t. injection of FR140423 with an ED50 value of 3.7 microg/mouse was completely antagonized by co-administered Leu-Arg 10 microg/mouse. However, L-NAME (2000 mg/kg s.c.) and MB (200 mg/kg s.c.) did not antagonize the antinociception of FR140423. These findings suggest that FR140423 plays a role in nociceptive modulation in the spinal cord, being antinociceptive via the kyotorphin-Met-enkephalin pathway but not via the peripheral NO-cyclic GMP pathway. PMID- 10855945 TI - Adrenocorticotropin inhibits nitric oxide synthase II mRNA expression in rat macrophages. AB - During hemorrhagic shock there is a massive overproduction of nitric oxide (NO). In such conditions, the intravenous (i.v.) injection of melanocortin peptides in nanomolar amounts produces a long-lasting restoration of cardiovascular and respiratory functions associated with the normalization of NO blood levels. To clarify the mechanism of such melanocortin-induced inhibition of NO overproduction, the influence of the adrenocorticotropin fragment 1-24 [ACTH-(1 24)] on the NO synthesizing activity of rat macrophages was studied in vitro. Nitrite production, an indicator of NO synthesis, was measured in the supernatant of rat macrophages whose inducible NO synthase (NOS II, iNOS) had been stimulated by the addition of S. enteritidis lipopolysaccharide (LPS, 50 microg/ml). ACTH-(1 24) (25, 50 and 100 nM) inhibited nitrite production when incubated together with LPS, but had no effect when applied 6 h after LPS. Further, the effect of ACTH-(1 24) on the expression of iNOS mRNA in rat macrophages activated with LPS was studied by means of a reverse transcriptase-polymerase chain reaction assay. ACTH (1-24) (25, 50 and 100 nM), applied together with LPS, dose-dependently suppressed iNOS gene activation. The present data suggest that the melanocortin induced normalization of NO blood levels during hemorrhagic shock is due, at least in part, to a direct inhibition of iNOS induction, at the level of mRNA transcription. PMID- 10855946 TI - Induction of apoptosis in mouse brain capillary endothelial cells by cyclosporin A and tacrolimus. AB - Although cyclosporin A and tacrolimus are used clinically as potent immunosuppressants, there have been reports of neurotoxicity and encephalopathy. A possible mechanism is that these drugs damage the blood-brain barrier (BBB), inducing dysfunction and increased permeability, and are then able to enter the brain. We studied the cytotoxicity of cyclosporin A and tacrolimus, focused on apoptosis induction, using an immortalized cell line established from BALB/c mouse cerebral microvessel endothelial cells (MBEC4). We found that these two drugs induced cell shrinkage, chromatin condensation and DNA fragmentation, which are characteristics of apoptosis. Our data suggest that the induction of apoptosis on the brain capillary endothelial cells may be at least partly involved in the occurrence of immunosuppressant-induced encephalopathy. PMID- 10855947 TI - Aluminum inhibits the lysosomal proton pump from rat liver. AB - Lysosomes are cytoplasmatic organelles, delimitated by a single lipoprotein membrane, that contain several enzymes mostly belonging to the hydrolases in that they function mainly for intracellular digestion. Lysosomal internal pH is characteristically acidic and it is maintained around pH 4.5 by a proton pump, an ATPase, that uses energy from ATP hydrolysis to translocate H+ ions into lysosomes. In the presence of Al3+ the proton pump activity is markedly reduced compromising acidic vesicles functionality. Among different species utilized, Al2(SO4)3 and AlF3 were the most effective. Aluminum effect was not observed when the delta pH was produced artificially by nigericin. PMID- 10855948 TI - Plasma homocysteine concentrations are positively associated with hostility and anger. AB - Homocysteine is a sulphur amino acid that is positively associated with risk of vascular disease. Very few behavioral or psychological factors have been studied in relationship to homocysteine levels, despite the fact that several psychological factors have also been linked with risk for cardiovascular disease. One psychological attribute showing a strong association with risk is hostility, which is prospectively predictive of future cardiovascular disease endpoints. Another related psychological factor is anger expression; coronary heart disease risk is associated with both heightened expression and inhibition of anger. The purpose of this study was to test the relationship of hostility and anger expression with homocysteine concentrations in a sample of healthy, middle-aged men and women. Participants completed the Cook-Medley hostility questionnaire, the Speilberger Anger Expression questionnaire, and had blood taken for the assessment of plasma homocysteine concentrations. Results indicated positive and significant associations between hostility and homocysteine levels for all participants, and positive and significant correlations between anger-in and homocysteine levels for men only. These data are among the first to test the relationship between homocysteine and psychological risk factors for cardiovascular diseases, and suggest one potential mechanism for the increased cardiovascular risk associated with hostility and anger expression. PMID- 10855949 TI - Effects of L-dopa treatment on methylation in mouse brain: implications for the side effects of L-dopa. AB - The effects of L-dopa on methylation process in the mouse brain were investigated. The study is based on recent findings that methylation may play an important role in Parkinson's disease (PD) and in the actions of L-dopa. The methyl donor, S-adenosylmethionine (SAM) and a product of SAM, methyl beta carboline, were shown to cause PD-like symptoms, when injected into the brain of animals. Furthermore, large amounts of 3-O-methyl dopa, the methyl product of L dopa, are produced in PD patients receiving L-dopa treatment, and L-dopa induces methionine adenosyl transferase, the enzyme that produces SAM. The results show that, at 0.5 hr, L-dopa (100 mg/kg) decreased the methyl donor, S adenosylmethionine (SAM) by 36%, increased its metabolite S-adenosylhomocysteine (SAH) by 89% and increased methylation (SAH/SAM) by about 200%. All parameters returned to control values within 4 hr. But 2, 3 and 4 consecutive injections of L-dopa, given at 45 min intervals, depleted SAM by 60, 64 and 76% and increased SAM/SAH to 818, 896, and 1524%. L-dopa (50, 100 and 200 mg/kg) dose-dependently depleted SAM from 24.9 +/- 1.7 nmol/g to 13.0 +/- 0.8, 14.7 +/- 0.8 and 7.7 +/- 0.7 nmol/g, and increased SAH from 1.88 +/- 0.14 to 3.43 +/- 0.26, 4.22 +/- 0.32 and 6.21 +/- 0.40 nmol/g. Brain L-dopa was increased to 326, 335 and 779%, dopamine to 138, 116 and 217% and SAH/SAM to 354, 392 and 1101%. The data show that L-dopa depletes SAM, and increases methylation 4-5 times more than dopamine, therefore, methylation may play a role in the actions of L-dopa. This and other studies suggest that the high level of utilization of methyl group by L-dopa leads to the induction of enzymes to replenish SAM and to increase the methylation of L-dopa as well as DA. These changes may be involved in the side effects of L-dopa. PMID- 10855950 TI - Novel effects of clotrimazole on Ca2+ signaling in Madin Darby canine kidney cells. AB - The effect of clotrimazole on Ca2+ signaling in Madin Darby canine kidney (MDCK) cells was investigated by using fura-2 as a Ca2+ indicator. Clotrimazole (1-30 microM) induced a concentration-dependent [Ca2+]i increase. The [Ca2+]i increase comprised an initial rise and a slow decay. External Ca2+ removal partly inhibited the Ca2+ signals by reducing both the initial rise and the decay phase, indicating that clotrimazole triggered both Ca2+ influx and Ca2+ release. Pretreatment with 30 microM clotrimazole in Ca2+-free medium abolished the Ca2+ release induced by thapsigargin (1 microM), an endoplasmic reticulum Ca2+ pump inhibitor, and conversely, pretreatment with thapsigargin prevented clotrimazole from releasing more Ca2+. This suggests that the thapsigargin-sensitive Ca2+ store is the source of clotrimazole-induced Ca2+ release. Clotrimazole (10 microM) triggered Mn2+ quench of fura-2 fluorescence which was partly inhibited by 1 mM La3+. Addition of 3 mM Ca2+ induced a [Ca2+]i increase after preincubation with 10 microM clotrimazole in Ca2+-free medium, indicating that clotrimazole activated capacitative Ca2+ entry. However, 10 and 30 microM clotrimazole inhibited 1 microM thapsigargin-induced capacitative Ca2+ entry by 21% and 74%, respectively. Pretreatment with 40 microM aristolochic acid to inhibit phospholipase A2 reduced 30 microM clotrimazole-induced Ca2+ release by 51%, but inhibiting phospholipase C with 2 microM U73122 had little effect. This implies that clotrimazole induces Ca2+ release in an IP3-independent manner, which could be modulated by phospholipase A2-coupled events. PMID- 10855951 TI - Effects of methotrexate on nucleotide pools in normal human T cells and the CEM T cell line. AB - It has been proposed that the clinical utility of methotrexate (MTX) in the treatment of rheumatoid arthritis may be due, in part, to inhibition of 5-amino imidazole-4-carboxamide ribonucleotide formyltransferase (AICARFT) by polyglutamated forms of MTX. AICARFT is the second folate dependent enzyme in de novo purine biosynthesis. In this study, the effects of MTX on de novo purine biosynthesis as well as total nucleotide pools were evaluated in both the human T cell line, CEM, and phytohemagglutinin-activated normal human T lymphocytes. De novo synthesized purines were metabolically labeled with 14C-glycine after MTX treatment and analyzed by HPLC. In normal T cells, MTX produced a dose-dependent reduction in de novo adenosine and guanosine pools with maximal effects (>50%) at 1 microM MTX. In CEM cells, de novo purine synthesis was almost completely blocked by 1 microM MTX. Total purine pools were also reduced in both cell types after MTX treatment. Since 1 microM MTX caused almost complete growth inhibition in CEM cells, we evaluated whether growth could be reconstituted with exogenous purine bases and pyrimidine nucleosides which can be utilized via salvage pathways. The combination of hypoxanthine and thymidine substantially reversed growth inhibition with 1 microM MTX in CEM cells. Taken together, these results demonstrate that MTX inhibits de novo nucleotide synthesis in T cells and suggest that AICARFT inhibition may be one aspect of the multi-site mechanism of MTX action in the treatment of rheumatoid arthritis. PMID- 10855952 TI - Amphotericin B-induced apoptosis and cytotoxicity is prevented by the Na+, K+, 2Cl(-)-cotransport blocker bumetanide. AB - Amphotericin B is the most commonly used antifungal drug although it exhibits poor effectiveness and considerable toxicity during treatment. It acts as a ionophore inducing cellular potassium efflux. The efflux of potassium, which is necessary for cell shrinkage during apoptosis, is counteracted by increased inward pumping of potassium ions. Modulation of potassium pump activity could therefore interact with programmed cell death depending on the nature of the disruption of cellular potassium homeostasis and subsequently affect the cytotoxicity of various drugs. We explored the role of apoptosis in amphotericin B-induced cytotoxicity in a mesothelioma cell line (P31) and investigated the role of K+ influx inhibitors of Na+, K+, ATPase and Na+, K+, 2Cl(-)-cotransport in these processes. Clone formation was used to determine the cytotoxicity of amphotericin B, ouabain (Na+, K+, ATPase blocker), and bumetanide (Na+, K+, 2Cl( )-cotransport blocker), alone or in combination. Apoptosis was estimated by quantifying free nucleosomes. Amphotericin B (3.2 micromol/L, 3 mg/L) per se reduced the percentage of surviving clones to 64% and increased the number of nucleosomes by 31% compared to untreated control. When ouabain (100 micromol/L) was added to amphotericin B a less than additive effect on clone formation was seen but no reduction of nucleosomes was noted. Bumetanide (100 micromol/L) per se was not cytotoxic but increased cellular nucleosome expression. Bumetanide eradicated amphotericin B-induced reduction of formed clones and generated nucleosomes. In conclusion, the induction of apoptosis seems to be of significant importance in amphotericin B-induced cytotoxicity. Amphotericin B-induced cytotoxicity and apoptosis was eradicated by the Na+, K+, 2Cl(-)-cotransport inhibitor bumetanide. The changes of cellular K+ fluxes induced by bumetanide combined with amphotericin B needs further elucidation. Bumetanide could possibly be used in antifungal therapy to increase amphotericin B effectiveness doses without increasing its adverse effects. PMID- 10855953 TI - Effect of dothiepin on gastric ulceration mediated by lipid derived eicosanoids. AB - Dothiepin, a tricyclic antidepressant, significantly inhibited the development of gastric ulcers induced by alcohol, aspirin, indomethacin and Shay's pyloric ligation. Antisecretory studies in pyloric ligated rats revealed that the drug at a dose of 100 mg/kg significantly reduced total acidity, gastric output and protein content. In another set of experiments, dothiepin significantly reduced gastric output, total acidity and protein content in pyloric ligated rats which received 50% alcohol (v/v) 30 minutes after the administration of dothiepin. PMID- 10855954 TI - Skeletal manifestations of bear scavenging. AB - In many partially or fully skeletonized forensic cases, postmortem animal damage is simply attributed to rodents or carnivores; little effort is made to determine the general size or assign a genus to the scavenger. As one of the largest wild carnivores to inhabit mountainous and forested areas throughout the continental United States, Alaska, and Canada, black bears (Ursus americanus) must be considered possible suspects when skeletonized remains are located showing marks of carnivore damage. Since 1995, three cases of known bear scavenging have been referred to the Maxwell Museum's Laboratory of Human Osteology by the New Mexico Office of the Medical Investigator for skeletal analysis. These cases comprise a total of seven individuals, and all of the remains were deposited in high altitude forests of New Mexico along the western border with Arizona with a minimum of 4 months exposure before recovery. When analyzed, all cases shared a similar pattern of element survivorship and damage. We suggest that bears can be distinguished from members of the canid family, the other common scavenger of human remains, based on the representation of skeletal elements at the scene. Rates and patterns of damage are not as accurate as element recovery in the discrimination of scavenger genus. Use of this information should allow forensic anthropologists to better understand the postmortem taphonomic processes that shaped the skeletal remains, and hopefully prevent misdiagnoses of perimortem trauma on elements not typically scavenged by canids. PMID- 10855955 TI - Detecting psychoactive drugs in the developmental stages of mushrooms. AB - The following questions regarding the detection of psychoactive drugs in mushrooms are addressed: At what stage of the mushroom development can the psychoactive drugs psilocyn and psilocybin be identified, and what effect does light have on the growth of these mushrooms. To answer these questions, Psilocybe cyanescens Wakefield mushrooms were grown from their spores in a controlled setting. At various times of their development, samples were taken and analyzed for psilocyn and psilocybin. Knowing what stage of development the psychoactive drugs can be identified may be useful to law enforcement personnel and forensic chemists. Methanolic extracts of various samples were analyzed by TLC and by GC/MS. It was determined that the mycelium knot stage of the mushroom was the earliest stage at which the psychoactive drugs could be detected. It was observed that light affected the time of development and the appearance of these mushrooms. PMID- 10855956 TI - Reagents for the chemical development of latent fingerprints: scope and limitations of benzo[f]ninhydrin in comparison to ninhydrin. AB - Benzo[f]ninhydrin was compared to ninhydrin for fingerprint development on paper. Overall, the performance of ninhydrin on exhibits was slightly better than that of benzo[f]ninhydrin. The significant advantages of the benzo[f]ninhydrin over ninhydrin were the much stronger fluorescence it gave after treatment with zinc salts and a slightly quicker reaction under ambient conditions. This fluorescence is, however, similar to that obtained with other reagents, such as DFO or ninhydrin analogs. These advantages apparently are not sufficient to justify regular usage of benzo[f]ninhydrin, especially when one considers its low solubility and high cost. PMID- 10855957 TI - Photoluminescent semiconductor nanocrystals for fingerprint detection. AB - The concept of utilizing photoluminescent semiconductor nanocrystals for latent fingerprint detection, especially in concert with phase-resolved imaging for background fluorescence suppression, is reduced to practice with CdS nanocrystals that are capped with dioctyl sulfosuccinate. The nanocrystals are dissolved in heptane or hexane and are applied in much the same way as staining with fluorescent dye, on articles that have been pre-fumed with cyanoacrylate ester and also on the sticky side of electrical tape without pre-fuming. Since CdS can form a photoluminescent nanocomposite with dendrimers, a feasibility examination of dendrimer tagging of fingerprints has also been conducted. PMID- 10855958 TI - Geographic origin determination of heroin and cocaine using site-specific isotopic ratio deuterium NMR AB - SNIF-NMR (Site-specific natural isotopic fractionation measured by deuterium NMR) was employed on 36 heroin samples from seven different known origins, and two cocaine samples from two different known origins. Heroin has two "synthetic" deuterium labeled sites (the two acetyls from acetic anhydride, each representing three equivalent nuclei) and 15 "natural" deuterium labeled sites (originating from the morphine produced in the opium plant). The "natural" sites have the potential for determining geographic location of the original opium plant, while the "synthetic" sites could assist in giving information about the commercial source of acetic anhydride used to convert morphine to heroin. Cocaine has 15 "natural" deuterium labeled sites. This study shows that SNIF-NMR has some use in determining the geographic origin of heroin and also has good potential for determining the geographical origin of cocaine. PMID- 10855959 TI - A scanning beam time-resolved imaging system for fingerprint detection. AB - A highly sensitive confocal scanning-beam system for time-resolved imaging of fingerprints is described. Time-resolved imaging is a relatively new forensic procedure for the detection and imaging of latent fingerprints on fluorescent substrates such as paper, cardboard, and fluorescent paint. Ordinary fluorescent imaging of latent fingerprints on these surfaces results in poor contrast. Instead, the specimens are treated with a phosphorescent dye that preferentially adheres to the fingerprint which allows time-resolved discrimination between the fingerprint phosphorescence and the background fluorescence. Time resolved images are obtained by synchronizing the digital sampling of the specimen luminescence with the on-off cycle of the chopped illumination beam. The merit of this technique is illustrated with high contrast images of fingerprints obtained from the fluorescent painted surface of a Coke can. PMID- 10855960 TI - Clothing damage analysis and the phenomenon of the false sexual assault. AB - This paper describes three recent false sexual assaults examined at the Victoria Forensic Science Centre laboratory where clothing damage analysis assisted in the resolution of the case. Suspected false reports of sexual assaults are often sensitive cases with little other forensic evidence. Any evidential value that can be obtained is thus valuable in order to minimize any ordeal to the complainant and any suspect and to conserve valuable resources. The findings illustrate the application of clothing damage analysis in a cross section of confirmed false sexual assault reports and the fact that the forensic examiner should be aware of the potential evidential value of this kind of analysis. Furthermore, the corroboration of a victim's scenario when the investigator has doubts may be no less valuable as it may minimize the adversarial ordeal that is often faced by a rape victim. PMID- 10855961 TI - On the assessment of children in suspected child sexual abuse in light of Daubert and Frye: limitations of profiles and interviews as scientifically grounded evidence. AB - Practice with children and families entails the higher probability of encountering forensic issues of child sexual abuse (CSA) assessments for which relatively few psychologists, allied mental health and legal practitioners are sufficiently well equipped. The current paper reviews some of the key psycholegal issues bearing on the assessment of suspected CSA in the contexts of: (a) recent psycholegal precedence and common law rules of reliability and admissibility of CSA profile evidence; (b) the empirical problems with CSA syndromes; and (c) the problems with children's interviews as evidence, and suggestions for valid interviewing guidelines supporting free recall. These psycholegal issues are presented in terms of the Frye standard for expert testimony and the Federal Rules of Evidence, with recent American and Canadian case illustrations, such as Daubert v. Merrell Dow Pharmaceuticals, Hadden v. State of Florida (1997), Bighead v. The United States of America (1997), Diocese of Winona v. Interstate Fire & Cas. Co. (1994), and R. v. Simpson (1996). PMID- 10855962 TI - A cross-cultural review of sudden mass assault by a single individual in the oriental and occidental cultures. AB - A nonrandom sample of North American cases of sudden mass assault by a single individual (SMASI, n = 30) is compared with a nonrandom sample of Laotian amok cases (n = 18) and other amok studies. Perpetrators in both studies show evidence of social isolation, loss, depression, anger, pathological narcissism, and paranoia, often to a psychotic degree. The term "innovative perpetrator" is reintroduced and expanded upon. Similarities among samples far outweigh differences, leading the authors to conclude that SMASI and its appearance in different cultures is not a culture-bound syndrome. PMID- 10855963 TI - Burned beyond recognition: systematic approach to the dental identification of charred human remains. AB - Forensic dental evaluation methods for use in a systematic approach to the dental identification of charred human remains are described. A systematic, conservative approach prevents the loss of valuable dental information before a thorough picture of the individual's dental remains has been adequately documented. The presenting conditions of fire victims are explained and illustrated with photographs, and a series of illustrations and text describe the damage seen in the dentition of the fire victim. A systematic four-stage process for gaining access to the intraoral structures of charred human remains is outlined and illustrated. Utilizing methods of access to the oral structures that maintain the integrity of the dentition through each stage of the evaluation of charred remains will prevent the loss of potential dental information before a thorough dental charting, intra-oral photographs, and radiographs can be obtained. PMID- 10855964 TI - Radiography of perforating centerfire rifle wounds of the trunk. AB - All deaths resulting from perforating centerfire rifle wounds of the chest and abdomen, investigated by the Office of the Chief Medical Examiner for the Province of Alberta from 1988 to 1995. were reviewed retrospectively to determine whether the radiographic distribution of bullet fragments in such cases is a useful predictor of bullet trajectory. Study cases were limited to single gunshot wounds without surgical intervention or intermediate targets, and for which adequate radiography was available. Three pathologists individually viewed the radiographs on two separate occasions; wound locations were provided for the second viewing (Group 2). Differences in opinion regarding direction of fire were resolved by consensus review. A trauma radiologist independently made two sets of interpretations in the same way. Comparisons of these groups of interpretations were made with the actual bullet direction determined at autopsy. Of 21 cases included in the study, only three (14.3%) did not require consensus resolution in either group. Accuracy of pathologists' interpretation improved from 38.1% (8/21) to 76.2% (16/21) with provision of wound locations (p = 0.012). The radiologist achieved similar improvement, from 28.6% (6/21) to 47.6% (10/21). The rate of agreement between radiologist and pathologists increased from 42.9% (9/21) to 61.9% (13/21) between Groups 1 and 2. Both the pathologists and radiologist interpreted several cases the same way in both groups; of those cases interpreted differently, the second interpretation was occasionally incorrect after correct interpretation in Group 1. We conclude that bullet direction for perforating centerfire rifle wounds cannot be accurately determined from postmortem radiographs. When wound location is known, the ability to predict bullet direction improves but is still subject to error, including a lack of consistency between observers. PMID- 10855965 TI - Petechiae of the baby's skin as differentiation symptom of infanticide versus SIDS. AB - The successive killing of three siblings by their biological mother at two-year intervals is described. The children were 367 days, 75 days and 3 years old. Although sudden infant death syndrome (SIDS) or interstitial pneumonia could not be ruled out as the cause of death in the two younger children, who were killed first, the third child exhibited discrete signs of violence in the mouth and throat area which were interpreted as proof of infanticide. All three children had petechiae of the skin of the face and throat, the upper thorax, the shoulders and the mucous membranes of the mouth. None of the children exhibited signs of a disease-related hemorrhagic tendency. After the mother was convicted of murdering the three-year-old boy by smothering in combination with compression of the thorax, she confessed to having killed the other two children in a similar manner. In the absence of hemostatic disease, the presence of petechiae of the skin extending over the entire drainage area of the Vena cava superior can be regarded as evidence of an increase in pressure in the thoracic cavity secondary to obstruction of the airways with simultaneous chest compression. PMID- 10855966 TI - Suffocation using plastic bags: a retrospective study of suicides in Ontario, Canada. AB - One hundred and ten cases of suicidal suffocation using a plastic bag were identified in the files of the Office of the Chief Coroner of Ontario, Canada, between 1993 and 1997. The records were reviewed to determine the demographic characteristics of this group compared with all cases of suicide in Ontario, the scene information, autopsy findings and toxicology results. Most suicides occurred in people over 60 years of age, with older women making up a considerable proportion of cases as compared with other methods of suicide. In 40% of cases the deceased was suffering from a serious illness. Autopsy findings were usually minimal, with facial, conjunctival and visceral petechiae present in a minority of cases. One or more drugs were detected in the blood in 92.6% of cases where toxicologic testing was performed. Benzodiazepines, diphenhydramine and antidepressants were the most common drugs found, with diphenhydramine the most common drug present at an elevated concentration. Information at the scene from "right to die" societies was uncommon. One quarter of decedents took additional measures, besides the use of drugs or alcohol, to ensure the rapidity, certainty or comfort of their death. This study further elucidates the characteristics of this uncommon method of suicide. It emphasizes additional scene findings, such as the presence of dust masks, physical restraints and modification of the plastic bag that may be of use to death investigators in determining the correct manner of death. PMID- 10855967 TI - A triad of laryngeal hemorrhages in strangulation: a report of eight cases. AB - The results of histological studies on larynges from eight cases of manual strangulation, all that had intracartilaginous laryngeal hemorrhages, a recently described and under-recognized lesion associated with strangulation, are reported. Formalin-fixed larynges were examined in serial section using a standardized protocol. In all cases, intracartilaginous laryngeal hemorrhages were associated with subepithelial laryngeal hemorrhages, and intralaryngeal muscular hemorrhages forming a "triad of hemorrhages." In five cases, the triad was found in the presence of laryngeal cartilage microfractures. Since cartilage microfractures can be causally related to mechanical injury to the neck, it is likely that the triad of hemorrhages has diagnostic value as an independent morphological criterion for the postmortem diagnosis of strangulation. Since a proportion of cases of strangulation lack characteristics that are self-evidently due to violent application of pressure on the neck, recognition of the triad may have important implications for the postmortem diagnosis of strangulation. PMID- 10855968 TI - Carisoprodol, meprobamate, and driving impairment. AB - This paper considers the pharmacology of the centrally acting muscle relaxant carisoprodol, and its metabolite meprobamate, which is also administered as an anxiolytic in its own right. Literature implicating these drugs in impaired driving is also reviewed. A series of 104 incidents in which these drugs were detected in the blood of drivers involved in accidents or arrested for impaired driving was considered, with respect to the analytical toxicology results, patterns of drug use in these subjects, the driving behaviors exhibited, and the symptoms observed in the drivers. Symptomatology and driving impairment were consistent with other CNS depressants, most notably alcohol. Reported driving behaviors included erratic lane travel, weaving, driving slowly, swerving, stopping in traffic, and hitting parked cars and other stationary objects. Drivers on contact by the police displayed poor balance and coordination, horizontal gaze nystagmus, bloodshot eyes, unsteadiness, slurred speech, slow responses, tendency to doze off or fall asleep, difficulty standing, walking or exiting their vehicles, and disorientation. Many of these cases had alcohol or other centrally acting drugs present also, making difficult the attribution of the documented impairment specifically to carisoprodol and meprobamate. In 21 cases, however, no other drugs were detected, and similar symptoms were present. Impairment appeared to be possible at any concentration of these two drugs; however, the most severe driving impairment and most overt symptoms of intoxication were noted when the combined concentration exceeded 10 mg/L, a level still within the normal therapeutic range. PMID- 10855969 TI - Beta-hydroxybutyric acid--an indicator for an alcoholic ketoacidosis as cause of death in deceased alcohol abusers. AB - We analyzed the postmortem blood of a total of 100 fatal cases for beta hydroxybutyric acid (BHBA). In 25 cases of sudden and unexpected death of alcoholics we found pathologically increased levels of BHBA of 1260 to 47200 (median 8000) micromol/L. This led us to the diagnosis of an alcoholic ketoacidosis (AKA) as cause of death in these cases. The control group of 69 postmortem cases revealed that BHBA concentrations below 500 can be regarded as normal, and values up to 2500 micromol/L as elevated. Our study shows that BHBA values over 2500 micromol/L could lead to death, if no medical attention is sought. During storage we did not find any indication of postmortem formation or decomposition of BHBA in blood in vitro or in the corpses. In our opinion, BHBA should be considered the diagnostic marker of choice for the postmortem determination of alcoholic ketoacidosis (AKA) as the cause of death. The classical indications of such deaths are: unexpected death of a chronic alcoholic; none or only traces of ethanol in the blood; increased acetone blood concentration; and neither autopsy, histology, microbiology, nor toxicology reveal the cause of death. In six further cases a diabetic ketoacidosis (DKA) was diagnosed as the cause of death. PMID- 10855970 TI - Postmortem forensic toxicology of selective serotonin reuptake inhibitors: a review of pharmacology and report of 168 cases. AB - This paper reviews the complex pharmacology of the new class of antidepressant medications exhibiting selective inhibition of serotonin reuptake. The four selective serotonin reuptake inhibitors (SSRIs) considered--fluoxetine, fluvoxamine, sertraline and paroxetine--can result in toxicity and death through contributing to serotonergic excess resulting in serotonin syndrome, inhibiting the metabolism of other centrally acting drugs, leading to accumulation of toxic concentrations, and exerting complex vasoactive effects on the vascular smooth muscle. This latter feature is of particular concern in patients with preexisting heart disease. An analytical method involving isolation of the drugs by liquid/liquid extraction at alkaline pH into n-butyl chloride, and analysis by gas chromatography/mass spectrometry (GC/MS) is described, together with some of its limitations. Toxicologic and cause and manner of death data were examined in 60 deaths involving fluoxetine, 5 involving fluvoxamine, 75 involving sertraline, and 28 involving paroxetine. Deaths involving drug toxicity were generally a result of ingestion of multiple drugs, and in only a small number of the cases was death attributed principally to the SSRI involved. The potential for drug interactions between members of this class of drugs is discussed as well as their metabolites and a variety of other therapeutic and abused drugs which can contribute to their toxicity. In the absence of other risk factors, the lowest concentrations determined to have resulted in death were 0.63 mg/L for fluoxetine, 0.4 mg/L for paroxetine, and 1.5 mg/L for sertraline. We had insufficient data to make even this crude assessment for fluvoxamine. Drug induced elevation of serotonin concentrations may be a significant risk factor for patients with atherosclerotic cardiovascular disease (ASCVD). Other factors including preexisting disease and the presence of other drugs and their pharmacology need to be carefully considered before determining the appropriate cause and manner of death in these cases. PMID- 10855971 TI - Drug-related deaths among recently released prisoners in the Strathclyde Region of Scotland. AB - Drug abuse and its consequences are everyday problems encountered globally, and Scotland is no exception. During a study of drug-related deaths in the Strathclyde region of Scotland it was noted that known drug users who had recently been released from prison were at high risk of dying from a drug overdose. The majority of deaths occurred within one week of the release date and polydrug use was prevalent. Morphine was the most frequently encountered drug and this was found in combination with benzodiazepines in a significant number of cases. This paper highlights the dangers of resuming drug consumption following a period of abstinence. PMID- 10855972 TI - Ketamine in non-hospital and hospital deaths in New York City. AB - We reviewed all ketamine-positive deaths (87) examined at the New York City Office of Chief Medical Examiner over a two-year period (1997 to 1999). There were 15 non-hospital deaths with 12 due to acute multidrug intoxications, one due to sarcoidosis, and two due to physical injury (blunt and thermal). In no instance was a fatal intoxication caused exclusively by ketamine. Opiates (10/15), followed by amphetamines (7/15) and cocaine (6/15), were the most frequent co-intoxicants. Ethanol was found in only one death. The race of all decedents was white and the majority were men (11/15) between the ages of 18 and 30 years. The remaining 72 instances of positive ketamine findings were hospital deaths following surgical procedures or burns. PMID- 10855973 TI - 13C4-secobarbital as the internal standard for the quantitative determination of secobarbital--a critical evaluation AB - In this study, 13C4-secobarbital was used as an exemplar compound to illustrate the mechanism based on which the effectiveness of a proposed internal standard (IS) could be evaluated. A deuterated analog, 2H5-secobarbital, was also studied in parallel for comparison purposes. Well-established solid-phase extraction and methylation procedures were used prior to the GC/MS measurement step. The contribution of the intensity of an ion designated for the analyte (secobarbital) by the proposed IS, and similarly, the contribution of the intensity of an ion designated for the IS by the analyte--a phenomenon termed "cross-contribution" were evaluated based on a "direct measurement" procedure in which equimolar amounts of the analyte and the IS were used to generate intensity data. These intensity data were then used as the basis for the calculation of "cross contribution" (in percentages) of ions designated for the analyte and the IS. Cross-contribution data were compared with the linearity data resulting from two series of standards containing 25 to 9600 ng/mL secobarbital using two sets of quantitation ion pairs--m/z 196/200 and 195/199 with 13C4-secobarbital as the IS and m/z 196/201 and 195/200 with 2HS-secobarbital as the IS. 13C4-secobarbital was found to be much less problematic and thus can serve as a very effective IS. Cross-contribution data alone cannot fully explain the observed differences resulting from the use of these two ISs; further systematic study is needed to provide better understanding of the underlying interference mechanism. PMID- 10855974 TI - Microchemical identification of gamma-hydroxybutyrate (GHB) AB - A new microcrystal test for the detection of gamma-hydroxybutyrate (GHB) is described. The silver/copper reagent consists of an aqueous solution of 0.1 g of cupric nitrate and 0.1 g of silver nitrate in 10.0 mL water. While some crystals form upon evaporation of the reagent, the test forms distinctive crystals for GHB and does not form crystals with some commonly encountered controlled substances. The reagent was also tested against some controlled substances that have similar biological activity to GHB, including flunitrazepam, and some barbiturates. No crystals were observed with these compounds. A blind test was performed to determine if GHB could be discriminated from the other compounds. Two of ten unknowns were correctly identified as GHB--one solid, one liquid. One GHB sample was not identified as GHB and the remaining seven non-GHB samples were not identified as GHB. The reagent is therefore selective for GHB, but not extremely sensitive. PMID- 10855975 TI - Evaluation of an alkaline lysis method for the extraction of DNA from whole blood and forensic stains for STR analysis. AB - A modified alkaline lysis protocol for extracting DNA from forensically relevant specimens is evaluated and compared with the chelex 100 method. For whole blood, bloodstains and sperm stains, both methods yielded comparable results after amplification for a pentameric STR locus (HumCD4). The main advantages of the new method are: only approximately ten minutes and two pipetting steps are necessary and the expenses for the extraction are extremely low as only NaOH, TrisHCl buffer and a single microcentrifuge tube are required. Alkaline lysis also proved to yield DNA suitable for typing longer STRs by using dye-labeled primers and capillary electrophoresis. These advantages should render this protocol especially interesting for high-throughput laboratories in combination with multiplex PCR and fluorescent dye technology, although the storability of the extracts proved to be problematic. PMID- 10855976 TI - Usefulness of a toothbrush as a source of evidential DNA for typing. AB - We investigated the usefulness of a toothbrush as a source of DNA for an unidentified cadaver. Ten toothbrushes were obtained from ten individuals along with their peripheral blood. We recovered from 10 to 430 ng of DNA from all but one of the toothbrushes. All ten toothbrushes, including the one containing no detectable DNA by fluorometry, were typed correctly at all of the loci tested, including nine STRs. Three toothbrushes obtained in two actual deaths also identified two victims and one suspect. Therefore, toothbrushes seem to be useful as a source of evidential DNA for personal identification. PMID- 10855977 TI - Validation of the PowerPlex 1.1 loci for use in human identification. AB - STR typing is now the favored method of DNA analysis for the purposes of human identification in the forensic community. The Forensic Services Division of the Detroit Police Department has completed its validation of the PowerPlex 1.1 loci (CSF1PO, TPOX, THO1, vWA, D16S539, D7S820, D13S317, and D5S818) for use in forensic casework. Detroit Metro Area Red Cross samples were typed from each of five racial/ethnic groups--the Hispanic, Caucasian, African American, Asian, and American Indian populations--and allele and genotype frequencies were calculated. A rare off-ladder variant (9.1 allele at D7S820) was identified among the database samples. A number of validation studies were performed. DNA was extracted from different substrates and typed as expected, except for the DNA extracted from leather (signal absent from the D16S539, D7S820, D13S317, CSF1PO, and TPOX loci) and from dirt (no PCR product generated). The minor contributor in the mixture study (250 pg input DNA) was facile to discern. The Concordance study, the variety of fluids from the same individual, and NIST standards studies all produced the expected results. Finally, STR data confirmed previous DNA typing results from adjudicated casework samples. PMID- 10855978 TI - Population genetics of HPRTB, F13B, and LPL in Pernambuco, Northeast Brazil. AB - One hundred thirty-four unrelated Northeast Brazilian individuals were typed for the HPRTB, F13B, and LPL short tandem repeats (STRs). DNA was amplified by specific primers and identified by silver staining of polyacrylamide gels. The allelic frequencies of these loci were in agreement with Hardy-Weinberg proportions. The most frequent alleles were HPRTB*13, F13B*10, LPL*10. The combined probability of paternity and the discrimination power of these 3 STRs were high, permitting their utilization for forensic science purposes. PMID- 10855979 TI - Fingerprints and DNA: STR typing of DNA extracted from adhesive tape after processing for fingerprints. AB - An exhibit that is often received for examination in cases of robbery or terrorist activity is adhesive tape. This type of exhibit can often, but not always, be successfully processed for fingerprints. The question arises whether or not it is possible to extract and type DNA after the tape has been sequentially processed for fingerprints. In this work, various donors left fingerprints on the adhesive side of tapes. The tapes were then sequentially processed for fingerprints using an alternate light source, cyanoacrylate fuming, and staining with BY-40 and then crystal violet. DNA was subsequently successfully extracted, amplified and typed for six STR loci. PMID- 10855980 TI - The Slovenian population data on the PCR based loci HLA-DQA1, LDLR, GYPA, HBGG, D7S8, GC, and D1S80. AB - Allele frequencies for the loci HLA-DQA1, LDLR, GYPA, HBGG, D7S8, GC, and D1S80 were determined for a sample population of unrelated individuals from Slovenia. All loci meet Hardy-Weinberg expectations, except the loci GYPA (p = 0.041) and D1S80 (p = 0.009). There is little evidence for association of alleles among the seven loci. Only one out of 21 pairwise comparisons demonstrated departures from independence (HLA-DQA1/HBGG, p = 0.008). The allelic frequency data generally are similar to that of U.S. Caucasians. PMID- 10855981 TI - Population studies and validation of paternity determinations by six microsatellite loci. AB - A single locus system of 6 microsatellite markers was evaluated for paternity testing. A nonradioactive method based on peroxidase labeling of a DNA probe was used to estimate the allele frequency of markers D1S216, D3S1217, D7S480, D9S157, D13S153, and D16S422 by genotyping 1134-1698 chromosomes. The number of detected alleles were 22, 15, 23, 10, 16, and 19, respectively, and the allele frequency varied from 0.001 to 0.317. The genotype of 87 families, consisting of mother, father, and child was determined. The probability that a random individual will give a positive paternity was evaluated. We conclude that the markers can be reliably typed and give sufficient and reliable information for paternity testing. PMID- 10855982 TI - Allele and genotype frequencies for D1S80 locus in a Brazilian population sample. AB - Gene and genotype frequencies in relation to the D1S80 locus were determined in a sample of 197 unrelated individuals (144 Caucasians and 53 Mulattoes), living in the city of Sao Paulo, Brazil. The Mulatto group was composed by mixed individuals who presented at least one negroid physical characteristic or declared themselves to be of mixed (Black-White) ancestry. Nineteen different alleles were detected in the Caucasian sample and 15 among Mulattoes. Alleles 18 and 24 were found to be the most common ones in the Caucasian population with frequencies of 0.173 and 0.357 respectively; the sample heterozygote frequency was estimated in 0.824. Alleles 18, 24, and 28 were found to be the most common alleles among Mulattoes with respective frequencies of 0.150, 0.349, and 0.113; the sample heterozygote frequency was 0.759. Fifty-five different genotypes were detected among Brazilian Caucasians whereas the respective figure among Mulattoes was 31. No significant deviations from Hardy-Weinberg equilibrium were found in both population samples. PMID- 10855983 TI - Anger experience, styles of anger expression, sadistic personality disorder, and psychopathy in juvenile sexual homicide offenders. AB - Sexual homicide by juveniles is a rare phenomenon, and information regarding the psychological and behavioral characteristics of this group is limited. No studies exist which have investigated anger experience and styles of anger expression, and the relationship between anger, sadistic personality disorder, and psychopathy, in this type of youthful offender. These areas were explored by evaluating 14 juvenile sexual homicide offenders through clinical assessment, the State-Trait Anger Expression Inventory (STAXI), the Schedule for Nonadaptive and Adaptive Personality (SNAP), the Revised Psychopathy Checklist (PCL-R), and review of correctional records. Descriptive information for the STAXI scales and internal consistency data are presented. Trait Anger was significantly higher than State Anger for the youth, but still comparable to adolescent norms. The difference between Anger-In and Anger-Out scale scores was not significant. Unexpectedly, Anger Control scale scores were significantly higher than Anger Out scale scores, clinically consistent with efforts by some of these boys to resist sadistic impulses. Those four (31%) participants who met criteria for sadistic personality had significantly higher Anger-Out scale scores than those without the disorder, and were also higher on Trait Anger to a marginally significant degree. Psychopathy was significantly negatively associated with Anger Control. This study is intended to contribute to the scant literature on juvenile sexual homicide, and lends some support to the validity and utility of sadistic personality disorder as a diagnosis in younger forensic populations. The findings did not support the contention that this form of violence is necessarily an outgrowth of excessive anger. PMID- 10855984 TI - The comparison of toluene determination between headspace-solid phase microextraction and headspace methods in glue-sniffer's blood and urine samples. AB - An accurate and simple method was developed to determine the level of toluene in urine and blood quantitatively by using the gas chromatography/mass spectrometry (GC/MS) with headspace--solid phase microextraction (HS-SPME) technique. An assembly of SPME with a replaceable extraction fiber, coated with 100 microm polydimethylsiloxane, was used. The detection limit of toluene in blood and urine with HS-SPME technique was 10 times higher than that with headspace (HS) technique. To compare the HS-SPME with HS technique for the determination of toluene in biological fluids, blood and urine samples from glue sniffers were analyzed by both methods. The level of toluene by the two techniques was highly correlated: the correlation coefficient (r2) between the two sets of values were 0.98 and 0.96 in urine and blood, respectively. PMID- 10855985 TI - Achiral and chiral quantification of methamphetamine and amphetamine in human urine by semi-micro column high-performance liquid chromatography and fluorescence detection. AB - In this paper, miniaturized achiral and chiral high-performance liquid chromatographic procedures for the determination of methamphetamine and amphetamine in human urine are described. After a simple pretreatment of human urine (i.e., 10 microL of urine or diluted urine were acidified and dried-up under N2 at room temperature) and fluorescence derivatization with 4-(4,5 diphenyl-1H-imidazol-2-yl)-benzoyl chloride under mild conditions (pH 9.0, 10 min at room temperature), the derivatives were isocratically separated on a semi micro ODS column with Tris-HCl buffer (0.1 M, pH 7.0): acetonitrile (45 + 55 v/v) at a flow rate of 0.2 mL/min or their enantiomers were separated on a semi-micro OD-RH column with sodium hexafluorophosphate (0.3 M aq.): acetonitrile (44 + 56 v/v) at a flow rate of 0.1 mL/min as the mobile phase. Wide-ranged calibration curves were obtained with detection limits for the achiral and chiral analyses in the atto and femtomol levels, respectively, per injected volume. Satisfactory within- and between-day reproducibility data were obtained with both the methods with the highest relative standard deviation being 9.6%. The methods were applied to the determination of methamphetamine and amphetamine in human urine samples and the concentrations determined by the two methods were well correlated (r = 0.994). PMID- 10855986 TI - The search for "Yvonne": a case example of the delineation of a grave using near surface geophysical methods. AB - Shallow electromagnetic (EM) and ground-penetrating radar (GPR) surveys were conducted in an area north of Auckland, New Zealand to assist the search for human remains. The body had been buried for almost 12 years in a plantation forest that was irregularly disrupted and modified by tree harvesting and the partial removal of stumps. EM identified anomalous areas of potential interest, because a target need only be nearby to generate an EM response. GPR was then used to map subsurface layering, layering disruption, and buried objects, immediately adjacent to an EM anomaly. Because of the nature of the site, numerous geophysical anomalies were present. GPR was particularly sensitive to site disturbance resulting from the forestry operations. An isolated EM anomaly on the fringes of an expanded survey area was coincident with the location of the body. Whether for criminal investigations or for archaeological work, a combination of geophysical techniques is recommended. PMID- 10855987 TI - Physical match of fragmented bullets AB - The technique of physical match, which is widely used with materials such as plastic and glass, is applied to fragmented bullets for determination of common origin. The more conventional method, of tool marks comparison, is later used to connect the bullet with a firearm. PMID- 10855988 TI - Hyperparathyroidism and psychosis: possible prelude to murder. AB - The authors present a case of a middle aged attorney who suffered from hyperparathyroidism and a psychotic disorder. It is possible that the hyperparathyroidism may have precipitated an acute psychotic delusional rage leading to an attempted mass murder. They discuss the relationship between hyperparathyroidism and neuropsychiatric symptoms in consideration of available research. PMID- 10855989 TI - Sudden unexpected infant death due to fibroma of the heart. AB - A 7-month-old previously healthy female infant was found dead in her crib by her mother shortly after having been laid down to sleep following the noontime feeding. Because the child did not suffer from an acute illness and no other evidence pointed to a cause of death, it was initially assumed by the police that she had died of sudden infant death syndrome. At autopsy, however, the cause of death was determined to be cardiac arrhythmia secondary to fibroma of the heart. PMID- 10855990 TI - Homicidal commotio cordis in two children. AB - This paper's objective is to describe two cases of fatal commotio cordis resulting from the deliberate striking of children's chests by adults with their fists. These deaths involve two male children, ages 3 years and 14 months. The clinical histories, events in the households prior to the deaths, behaviors of the children, autopsy findings, and investigation results are all similar. In both cases, fatal blows were delivered to the anterior chest with a closed fist. Both children collapsed immediately, unable to be resuscitated. Confessions were obtained in both cases by investigators soon after the children's deaths. Autopsies showed chest contusions in only one child, presumably due to knuckle impact. The cardiac rhythms noted by paramedics were ventricular fibrillation and asystole. Due to the lack of physical findings, an immediate and thorough investigation is critical. An accurate history of events preceding death must be obtained. PMID- 10855991 TI - Fatal poisoning from Nicotiana glauca leaves: identification of anabasine by gas chromatography/mass spectrometry. AB - Death of a worker occurred after ingestion of unknown amounts of Nicotiana glauca G leaves. The leaves were cooked after having been mistakenly considered to be spices of a type which grow in Thailand. After ingestion, two Thai workers collapsed, one with asystolia. Resuscitation efforts were successful only for one of the victims. A GC/MS method was used for the identification of anabasine as the main constituent in the leaves, food extract, blood, and the urine of the deceased. Lacking a standard, it was necessary to interpret the GC/MS spectrum to identify anabasine and establish its presence. PMID- 10855992 TI - Gene frequencies for six STR loci in a Chilean population of mixed ancestry. PMID- 10855993 TI - Allele frequencies for nine STR loci in African American and Caucasian populations from Marion County, Indiana, USA. PMID- 10855994 TI - Commentary on Amar A, Brautbar C, Motro U, et al. Genetic variation of three tetrameric tandem repeats in four distinct Israeli ethnic groups. J Forensic Sci 1999;44(5):983-6. PMID- 10855995 TI - Commentary on George JR, Davis GG. Comparison of antiepileptic drug levels in different cases of sudden death. J Forensic Sci 1998;43:595-603. PMID- 10855996 TI - Commentary on the American Board of Criminalistics (ABC) Certification Process. PMID- 10855997 TI - Palliative care for HIV disease in the era of highly active antiretroviral therapy. PMID- 10855998 TI - Impact of HAART on causes of death of persons with late-stage AIDS. AB - CONTEXT: The increasing use of highly active antiretroviral therapies (HAARTs) has changed the course of AIDS-related illnesses and enhanced the quality of life of patients infected with human immunodeficiency virus (HIV) and may have changed the causes of deaths in patients with acquired immunodeficiency syndrome (AIDS). OBJECTIVE: The aim of the present study was to investigate causes of deaths in long-term care hospital patients with late-stage AIDS who expired at the Coler Goldwater Memorial Hospital in New York City in 1995, and in 1998 and 1999, that is, immediately before and the two most recent years after the advent of HAART. METHODS: Analysis of causes of deaths as recorded on the death certificates of 232 AIDS patients. RESULTS: The overall mortality rate declined from 75.6 deaths per 100 person-years in 1995 to 33.2 deaths per 100 person-years in 1998-1999 (P < .001). The number of AIDS patients who expired because of sepsis and opportunistic infections, which included Pneumocystis carinii pneumonia (PCP), decreased significantly from 30 (26.1%) and 24 (20.9%) in 1995 to 15 (12.8%) and 10 (8.5%) in 1998-1999, respectively (P < .05). In contrast, deaths from hepatic failure increased from 0 (0%) in 1995 to 7 (6%) in 1998-1999 (P < .05). Increases, although not significant statistically, were associated with pneumonias excluding PCP, end-stage AIDS, renal failure, and malignancies. Analysis of cause-specific mortality by gender between 1995 and 1998-1999 revealed very little difference between men and women. This analysis showed, however, that the infectious processes taken together (pneumonias excluding PCP, sepsis, and opportunistic infections including PCP) were significantly less frequent causes of death in 1998-1999 than in 1995 (P < .01). CONCLUSION: These findings indicate that HAART affected the causes of deaths in patients with AIDS, with "traditional" opportunistic infections diminishing in importance relative to chronic medical conditions and malignancies. PMID- 10855999 TI - Long-term continuum of care for people living with HIV/AIDS. AB - The introduction of highly active antiretroviral therapy (HAART), has created new options for those infected and affected by human immunodeficiency virus/ acquired immunodeficiency syndrome (HIV/AIDS). Most HIV-infected persons no longer die within months of diagnosis. There is now a long-term continuum of care that can end in misery or relative comfort. The introduction of palliative care in concert with curative therapies throughout the disease trajectory should be the standard of care for all persons. At the very least, the introduction of palliative care and hospice at the end of life is important to the holistic care of persons living with HIV/AIDS. PMID- 10856000 TI - HIV as a chronic disease: implications for long-term care at an AIDS-dedicated skilled nursing facility. AB - OBJECTIVE: To describe the characteristics and outcomes of the first 3 years of admissions to a dedicated skilled nursing facility for people with acquired immunodeficiency syndrome (AIDS). METHODS: Systematic chart review of consecutive admissions to a 30-bed, AIDS-designated long-term care facility in New Haven, Connecticut, from October 1995 through December 1998. RESULTS: The facility has remained filled to 90% or more of its bed capacity since opening. Of 180 patients (representing 222 admissions), 69% were male; mean age was 41 years; 57% were injection drug users; 71% were admitted directly from a hospital. Leading reasons for admission were (1) the need for 24-hour nursing/medical supervision, (2) completion of acute medical treatment, and (3) terminal care. On admission, the median Karnofsky score was 40, and median CD4+ cell count was 24/mm3; 48% were diagnosed with serious neurologic disease, 44% with psychiatric illness; patients were receiving a median of 11 medications on admission. Of 202 completed admissions, 44% of patients died, 48% were discharged to the community, 8% were discharged to a hospital. Median length of stay was 59 days (range 1 to 1,353). Early (< or = 6 months) mortality was predicted by lower admission CD4+ count, impairments in activities of daily living, and the absence of a psychiatric history; long-term stay (> 6 months) was predicted by total number of admission medications, neurologic disease, and dementia. Comparison of admissions from 1995 to 1996 to those in 1997 to 1998 indicated significantly decreased mortality rates and increased prevalence of psychiatric illness between the two periods (P < .01). CONCLUSIONS: A dedicated skilled nursing facility for people with AIDS can fill an important service need for patients with advanced disease, acute convalescence, long-term care, and terminal care. The need for long-term care may continue to grow for patients who do not respond fully to current antiretroviral therapies and/or have significant neuropsychiatric comorbidities. This level of care may be increasingly important not only in reducing lengths of stay in the hospital, but also as a bridge to community-based residential options in the emerging chronic disease phase of the AIDS epidemic. PMID- 10856001 TI - Chronic-treated HIV: a neurologic disease. AB - The concept of slow virus diseases was developed by Sigurdsson in the 1950s in studies of infections of Icelandic sheep, including Visna, a slow (lenti) viral infection of the central nervous system. Human immunodeficiency virus (HIV) belongs to the same lentivirus subfamily of retroviruses and causes significant dysfunction of all levels of the nervous system. Highly active antiretroviral therapy should allow host control of opportunistic infections, producing a clinical state of chronic-treated HIV. However, viral persistence may occur in the sanctuary of the central nervous system. As a consequence, major disabilities in the chronic-treated phase of the HIV epidemic may include cognitive impairment, gait disorders, and various pain syndromes. Policy planning will need to take into account the long-term residential, social, and health care needs of this population. PMID- 10856002 TI - Psychiatric comorbidity and the long-term care of people with AIDS. AB - OBJECTIVES: To examine the association of comorbid psychiatric disorders with admission and discharge characteristics for patients residing at a long-term care facility designated for acquired immunodeficiency syndrome (AIDS). METHODS: Demographic and clinical characteristics were obtained by systematic chart review for all patients (N = 180) admitted to the facility from its opening in October 1995 through December 1999. Lifetime history of severe and persistent psychiatric disorders (major depression, bipolar and psychotic disorders) was determined by current diagnosis on baseline clinical evaluation or a documented past history. RESULTS: Forty-five patients (25%) had comorbid psychiatric disorders. At admission, patients with comorbidity were more likely to be ambulatory (80% vs. 62%, P = .03) and had fewer deficits in activities of daily living (27% vs. 43%, P = .05). After controlling for human immunodeficiency virus (HIV) disease severity, patients with comorbidity had significantly lower discharge rates (relative risk = 0.43, 95% confidence interval 0.23-0.78, P = .0001) and death rates (relative risk = 0.53, 95% confidence interval 0.42-0.68, P = .009). CONCLUSIONS: Patients with AIDS and comorbid psychiatric disorders at this facility had more favorable admission characteristics and were less likely to be discharged or to die. They may have been admitted earlier in their disease course for reasons not exclusively due to HIV infection. Once admitted, community-based residential alternatives may be unavailable as a discharge option. These findings are unlikely to be an anomaly and may become more pronounced with prolonged survival due to further therapeutic improvements in HIV care. Health services planners must anticipate rising demands on the costs of care for an increasing number of patients who may require long-term care and expanded discharge options for the comanagement of HIV disease and chronic psychiatric disorders. PMID- 10856003 TI - The difficult, demanding, and demented AIDS patient in long-term care. AB - Demented AIDS patients in long-term care present interconnected medical, ethical, and management problems. The patient's right to care must be considered in the context of the obligations owed to other residents and to staff members. A principled analysis should focus on substantive and procedural issues: the concept of autonomy must be modified by notions of accommodation to the needs of others; procedural fairness should guide discussions. A dynamic analysis should identify the various parties, their conflicting interests, and possible routes for resolving differences. PMID- 10856004 TI - HIV/AIDS and long-term care: a state perspective. AB - Long-term care services for people with human immunodeficiency virus/ acquired immunodeficiency syndrome (HIV/AIDS) were fostered in New York State by passage of HIV-specific regulations that set program standards and authorized reimbursement rates sufficient to support these standards. A rapid expansion of HIV-specific capacity has occurred. Demographic and selected clinical characteristics of the populations in AIDS residential health care facilities and AIDS adult day health care programs in New York State are presented. Aspects of the service models for these two program types that have changed to meet new needs are discussed. PMID- 10856005 TI - Improving HIV/AIDS services through palliative care: an HRSA perspective. Health Resources and Services Administration. AB - There has been a dramatic shift of the human immunodeficiency virus/ acquired immunodeficiency syndrome (HIV/AIDS) epidemic into poor, marginalized, and minority communities in the US. At the same time, the availability of new highly active antiretroviral treatments has made it possible for a large number of individuals to live for a much longer time with their disease. A net result is that the US is faced with an increasing number of people who are living with HIV/AIDS and are dependent on publicly supported health care services. In this paper, we review the palliative care efforts of the federal agency, the Health Resources and Services Administration (HRSA), responsible for providing Ryan White CARE Act HIV/AIDS care to medically underserved populations. In addition to supporting traditional hospice care, HRSA's HIV/AIDS Bureau has begun a series of initiatives that apply a broader concept of palliative care to its HIV programs in hospital- and community-based settings. Our interest is not to substitute palliation for access to new HIV therapies, such as highly active antiretroviral treatments, but to ensure that our health delivery systems attend to the alleviation of symptoms and suffering along with the provision of antiretroviral and other necessary treatments. HRSA's HIV/AIDS Bureau is organizing a broader provision of palliative care for its clients and actively contributing to improving care for the disenfranchised internationally. PMID- 10856006 TI - New York City AIDS case mortality rates in the era of potent antiretroviral therapy. PMID- 10856007 TI - Convalescence and chronic illness. PMID- 10856008 TI - Effect of asthma on the quality of life among children and their caregivers in the Atlanta Empowerment Zone. AB - BACKGROUND AND OBJECTIVE: Asthma is the most common chronic pediatric disease and exacts a toll on the health-related quality of life of affected children and their primary caregivers. This investigation describes the relationship between the clinical severity of asthma among inner-city children and their quality of life and that of their primary adult caregivers. METHODS: Telephone interview data were collected from individual adult caregivers of 5-12-year-old children with asthma. Questions addressed the history, diagnosis, and management of the child's asthma, the child's family and social background, the family's socioeconomic status, the caregiver's knowledge and attitude about asthma, and the health-related quality of life of both the child and the caregiver. An asthma severity score was calculated from the caregiver's responses to questions about their child's wheezing frequency, nocturnal and early morning symptoms, and speaking during an asthma attack, as well as the impact of the disease on their child's physical activity and breathing during the prior 4-month period. A clinical asthma triage score was determined from information collected at the emergency department about the child's oxygen saturation, alertness, use of accessory respiratory muscles, extent of breathlessness, and peak expiratory flow. Spearman correlation coefficients were used to identify association between quality of life and disease severity, caretaker's asthma knowledge, and functional impact of asthma symptoms. RESULTS: Data from 240 of 755 eligible children were analyzed. Most children were younger than 11 years, male, black, and non-Hispanic. The children's median duration of asthma diagnosis was 86% of their life (range less than 1 to 11.3 years, median 5.0 years). Of the primary caregivers, 69% had at least completed high school, and 90% reported a total monthly household income of $1,600 or less. The maximum possible quality-of-life score and the median for caregivers were 91 and 70, respectively; for children, the same scores were 69 and 58, respectively. In addition, there was significant negative correlation of the quality-of-life scores of both the caregivers and children with the number of schooldays the children missed (r = -0.24 and r = 0.26, respectively, P < .001 for both) and the caregivers' and children's asthma severity scores (r = -0.39 and r = -0.47, respectively, P < .001 for both). The quality-of-life scores of the children and caregivers did not correlate significantly with the asthma triage scores. CONCLUSIONS: The questionnaires captured baseline quality-of-life information about this urban population and will facilitate longitudinal monitoring. The fact that the quality-of-life scores of children with asthma correlated with those of their adult caregivers, but not with their clinical triage scores, highlights the impact of asthma on families and the importance of having a long-term comprehensive management plan that is not based on exacerbations, but that includes both the children and their primary caregivers. PMID- 10856010 TI - U-Pb Geochronologic, Nd Isotopic, and Geochemical Evidence for the Correlation of the Chopawamsic and Milton Terranes, Piedmont Zone, Southern Appalachian Orogen. AB - We report U-Pb crystallization ages from four metavolcanic rocks and two granitic gneiss samples as well as whole-rock chemical analyses and Sm-Nd isotopic ratios from 25 metaigneous and metasedimentary rocks from the Chopawamsic and Milton terranes, southern Appalachian Orogen. A metarhyolite sample from the Chopawamsic Formation and a metabasalt sample from the Ta River Formation in the Chopawamsic terrane have indistinguishable U-Pb crystallization ages of 471.4+/-1.3 Ma and 470.0+1.3/-1.5 Ma, respectively. A sample from the Prospect granite that intruded metavolcanic rocks of the Ta River Formation yields a younger U-Pb date of 458.0+/-1 Ma. Metarhyolite and granitic gneiss samples from the northern part of the Milton terrane yield U-Pb dates of 458.5+3.8/-1.0 Ma and 450+/-1.8 Ma, respectively. Metavolcanic and metaplutonic rocks from both terranes span a range in major element composition from basalt to rhyolite. Trace element concentrations in these samples show enrichment in large-ion lithophile elements K, Ba, and Rb and depletion in high field strength elements Ti and Nb, similar to those from island arc volcanic rocks. Initial epsilon(Nd) values and T(DM) ages of the metaigneous and metasedimentary samples range from 0.2 to -7.2 and from 1200 to 1700 Ma for the Chopawamsic terrane and from 3.7 to -7.2 and from 850 to 1650 Ma for the Milton terrane. The crystallization ages for the metavolcanic and metaplutonic samples from both terranes indicate that Ordovician magmatism occurred in both. Similar epsilon(Nd) values from representative samples from both terranes suggest that both were generated from an isotopically similar source. Xenocrystic zircons from metavolcanic rocks in the Chopawamsic terrane have predominately Mesoproterozoic (207)Pb/(206)Pb ages (600-1300 Ma), but a single Archean (2.56 Ga) core was also present. The xenocrystic zircons and the generally negative epsilon(Nd) values indicate that both terranes are composed of isotopically evolved continental crust. PMID- 10856011 TI - Integrated Paleomagnetism and U-Pb Geochronology of Mafic Dikes of the Eastern Anabar Shield Region, Siberia: Implications for Mesoproterozoic Paleolatitude of Siberia and Comparison with Laurentia. AB - This article reports the first joint paleomagnetic and U-Pb geochronologic study of Precambrian diabase dikes in the Anabar Shield and adjacent Riphean cover of Siberia. It was undertaken to allow comparison with similar published studies in Laurentia and to test Proterozoic reconstructions of Siberia and Laurentia. An east-trending Kuonamka dike yielded a provisional U-Pb baddeleyite emplacement age of 1503+/-5 Ma and a virtual geomagnetic pole at 16 degrees S, 221 degrees E (dm=17&j0;, dp=10&j0;). A paleomagnetic pole at 6 degrees N, 234 degrees E (dm=28&j0;, dp=14&j0;) was obtained from five Kuonamka dikes. An east-southeast trending Chieress dike yielded a U-Pb baddeleyite emplacement age of 1384+/-2 Ma and a virtual geomagnetic pole at 4 degrees N, 258 degrees E (dm=9&j0;, dp=5&j0;). Kuonamka and Chieress poles are interpreted to be primary but do not average out secular variation. Assuming that the Siberian Plate has remained intact since the Mesoproterozoic, except for mid-Paleozoic opening of the Viljuy Rift, then the above results indicate that the Siberian Plate was in low latitudes at ca. 1503 and 1384 Ma, broadly similar to low latitudes determined for Laurentia from well-dated paleopoles at 1460-1420, 1320-1290, and 1267 Ma. This would allow Laurentia and Siberia to have been attached in the Mesoproterozoic, as suggested in several recent studies based on geological criteria. However, because paleomagnetic results from the Anabar Shield region do not average out secular variation and the ages of poles from Siberia and Laurentia are not well matched, it is not yet possible to distinguish between these reconstructions or to rule out other configurations that also maintain the two cratons at low paleolatitudes. PMID- 10856009 TI - A randomized field trial of ACINDES: a child-centered training model for children with chronic illnesses (asthma and epilepsy). AB - A randomized field trial of a child-centered model of training for self management of chronic illnesses was conducted of 355 Spanish-speaking school-aged children, between 6 and 15 years old, with moderate to severe asthma and epilepsy, in Buenos Aires, Argentina. The model, based on play techniques, consists of five weekly meetings of 8-10 families, with children's and parents' groups held simultaneously, coordinated by specially trained teachers and outside the hospital environment. Children are trained to assume a leading role in the management of their health; parents learn to be facilitators; and physicians provide guidance, acting as counselors. Group activities include games, drawings, stories, videos, and role-playing. Children and parents were interviewed at home before the program and 6 and 12 months after the program, and medical and school records were monitored for emergency and routine visits, hospitalizations, and school absenteeism. In asthma and epilepsy, children in the experiment showed significant improvements in knowledge, beliefs, attitudes, and behaviors compared to controls (probability of experimental gain over controls = .69 for epilepsy and .56 for asthma, with sigma2 = .007 and .016, respectively). Parent participants in the experiment had improved knowledge of asthma (39% before vs. 58% after) and epilepsy (22% before vs. 56% after), with a probability of gain = .62 (sigma2 = .0026) with respect to the control group. Similar positive outcomes were found in fears of child death (experimental 39% before vs. 4% after for asthma, 69% before vs. 30% after for epilepsy), as well as in disruption of family life and patient-physician relationship, while controls showed no change. Regarding clinical variables, for both asthma and epilepsy, children in the experimental group had significantly fewer crises than the controls after the groups (P = .036 and P = .026). Visits to physicians showed a significant decrease for those with asthma (P = .048), and emergency visits decreased for those with epilepsy (P = .046). An 18-item Children Health Locus of Control Scale (CHLCS) showed a significant increase in internality in experimental group children with asthma and epilepsy (P < .01), while controls did not change or performed worse 12 months after the program. School absenteeism was reduced significantly for those with asthma and epilepsy (for the group with asthma, fall/winter P = .006, and spring P = .029; for the group with epilepsy, P = .011). CONCLUSION: The program was successful in improving the health, activity, and quality of life of children with asthma and epilepsy. The data suggested that an autonomous (Piagetian) model of training is a key to this success, reinforcing children's autonomous decision making. PMID- 10856012 TI - A New Late Middle Cambrian Paleomagnetic Pole for the Ellsworth Mountains, Antarctica. AB - A paleomagnetic study of the late Middle to possibly early Late Cambrian Liberty Hills Formation in the Ellsworth Mountains, Antarctica, reveals a stable magnetization with positive fold and reversal tests. The paleopole is based on 16 sites from volcanic and sedimentary rocks and lies at lat 7.3 degrees N and long 326.3 degrees E (A95=6.0&j0;). The new paleomagnetic data support the view that the Ellsworth Mountains are part of a microplate-the Ellsworth-Whitmore Mountains crustal block-that rotated independently of the main Gondwana continental blocks during breakup. The Liberty Hills pole differs from both previous poles recovered from Cambrian rocks in the Ellsworth Mountains and from the available Gondwana reference pole data. Our pole indicates a more northerly prebreakup position for the Ellsworth Mountains than previously suggested, contradicting the overwhelming geologic evidence for a prebreakup position close to southern Africa. The reasons for this are uncertain, but we suggest that problems with the Gondwana apparent polar wander path may be important. More well constrained, early Paleozoic paleomagnetic data are required from the Ellsworth Mountains and the Gondwana continents if the data are to constrain further the Middle-Late Cambrian location of the Ellsworth-Whitmore Mountains block. PMID- 10856013 TI - Constraining the Late Mesozoic and Early Tertiary Tectonic Evolution of Southern Mexico: Structure and Deformation History of the Tierra Caliente Region. AB - We analyze the structure and assess the deformation history of the Tierra Caliente Metamorphic Complex (TCMC) of southern Mexico, where Laramide accretion of exotic terranes is in debate. The TCMC consists of a south-plunging antiform fault that is bounded on both its eastern and western flanks. Tierra Caliente Metamorphic Complex rocks show at least two phases of compressional deformation. The first and most prominent records a mean tectonic transport direction of 068 degrees. This phase is responsible for east-verging asymmetrical folding and thrusting of both metamorphic and superjacent sedimentary rocks. The second phase has an average transport direction of 232 degrees and is restricted to the western portion of the TCMC. A third phase is responsible for normal faulting. Lack of discernible deformation before Late Cretaceous time indicates that the main deformation phase is coincident with Laramide orogenesis elsewhere in the North American Cordillera. The stratigraphy, structure, and deformational history of the TCMC do not require accretion of exotic terranes. We explain the Mesozoic tectonostratigraphic evolution of the TCMC in terms of deposition and deformation of Mesozoic volcanic and sedimentary strata over the attenuated continental crust of the North American plate. PMID- 10856014 TI - A Lost Realm in the Internal Domains of the Betic-Rif Orogen (Spain and Morocco): Evidence from Conglomerates and Consequences for Alpine Geodynamic Evolution. AB - The Malaguide-Ghomaride Complex is capped by Upper Oligocene-Aquitanian clastic deposits postdating early Alpine orogenesis but predating the main tectonic metamorphic evolution, end of nappe emplacement, unroofing, and exhumation of the metamorphic units of the Betic-Rif Orogen. Two conglomerate intervals within these deposits are characterized by clasts of sedimentary, epimetamorphic, and mafic volcanic rocks derived from Malaguide-Ghomaride units and by clasts of acidic magmatic and orthogneissic rocks of unknown provenance, here studied. Magmatic rocks originated from late-Variscan two-mica cordierite-bearing granitoids and, subordinately, from aplitic dikes. Orthogneisses derive from similar plutonic rocks but are affected by an Alpine metamorphic overprint evolving from greenschist (T=510&j0;-530 degrees C and P=5-6 kbar) to low temperature amphibolite facies (T>550&j0;C and P<3 kbar). Such a plutonic rock suite is unknown in any Betic-Rif unit or in the basement of the Alboran Sea, and the metamorphic evolution in the orthogneisses is different from (and older than) that of Alpujarride-Sebtide rocks to which they were formerly ascribed. Magmatic and metamorphic rocks very similar to those studied characterize the basements of some Kabylia and Calabria-Peloritani units. Therefore, the source area is a currently lost continental-crust realm of Calabria-Peloritani-Kabylia type, located to the ESE of the Malaguide-Ghomaride Domain and affected by a pre-latest Oligocene Alpine metamorphism. Increasingly active tectonics transformed this realm into rising areas from which erosion fed small subsiding synorogenic basins formed on the Malaguide-Ghomaride Complex. This provenance analysis demonstrates that all these domains constituted a single continental-crust block until Aquitanian-Burdigalian times, before its dispersal around nascent western Mediterranean basins. PMID- 10856015 TI - The Kyonggi Shear Zone of the Central Korean Peninsula: Late Orogenic Imprint of the North and South China Collision. AB - The crustal-scale Kyonggi shear zone of central Korea has been identified as a major boundary between the Precambrian Kyonggi massif in the south and the Imjingang belt in the north. The latter is an eastward extension of the Qinling Dabie-Sulu collisional belt of China. Field observations and microstructural analysis indicate that the extensional shear zone evolved from a deep crustal ductile regime to a shallow crustal brittle regime, associated with a rapid uplift of the Kyonggi massif following the Late Permian-Early Triassic collision between the Sino-Korean and Yangtze cratons. A Rb-Sr muscovite age (226+/-1.2 Ma) of the mylonite suggests that the extensional ductile shearing occurred during the Late Triassic. PMID- 10856016 TI - Ultrahigh-Temperature Metamorphism in Madurai Granulites, Southern India: Evidence from Carbon Isotope Thermometry. AB - Ultrahigh-temperature (UHT) metamorphism in the Madurai Block of the southern Indian granulite terrain has been verified using the calcite-graphite isotope exchange thermometer. Carbon isotope thermometry has been applied to marbles from a locality near the reported occurrence of sapphirine granulites that have yielded temperature estimates of around 1000 degrees C. The delta(13)C and delta(18)O values of calcite are homogenous, implying equilibration of the isotopes during metamorphism. However, the delta(13)C values of single graphite crystals show variations in the order of 1 per thousand within a hand specimen. Detailed isotopic zonation studies indicate that graphite preserves either the time-integrated crystal growth history or reequilibrium fractionation during its cooling history. The graphite cores preserve higher delta(13)C values than the rims. The fractionation between calcite and graphite cores gives the highest metamorphic temperature of about 1060 degrees C, which matches the petrologically inferred temperature estimates in the high-magnesian pelites. The fractionation between graphite rims and calcite suggests a temperature of around 750 degrees C, which is interpreted to reflect retrograde cooling. This event is also observed in the sapphirine granulites. Calcite-graphite thermometry thus provides a useful tool to define UHT metamorphism in granulite terrains. PMID- 10856017 TI - Major Element, REE, and Other Trace Element Behavior in Amphibolite Weathering under Semiarid Conditions in Southern India. AB - A body of komatiitic amphibolite, an enclave within the Archean high-grade orthogneisses in southern India, shows mild chemical weathering under semiarid conditions. Along fractures, chemical weathering has advanced (Chemical Index of Alteration &sqbl0;CIA&sqbr0;=53; CIA of fresh rock approximately 26) to the extent that secondary Mg-Fe-Al clay minerals have formed and the rock has turned brownish red, soft, and fine grained. The weathering process has resulted in the mobilization and redistribution of the so-called immobile elements Fe, Al, Ti, and REE effected by the nature of secondary mineral formation (talc vs. aluminous clay minerals) and also possibly by soil microbes. In the initial stages of secondary mineral formation, there is a small loss of Fe, Al, and REE (noticeably Eu). However, in the fracture zone as well as in the incipiently altered zone, there is significant REE enrichment, probably affected by a different precipitation mechanism. Mobilized REE may have come from a minor alteration of clinopyroxene. PMID- 10856018 TI - A tiered approach for assessing children's exposure. AB - Recently, intense attention has been given to children's health issues, particularly in the use of consumer products. Because of this attention, researchers have been planning and initiating studies specifically aimed at developing both toxicology data and exposure data directed to improve our understanding of industrial and consumer product chemical impacts on children's health. To ensure that this research is focused on the highest priority chemicals, we present a methodology for determining and prioritizing the higher hazard chemicals and scenarios for which children could be disproportionately or highly exposed. This tiered approach includes a screening step for initial chemical selection, a hazard assessment based on no- or lowest-observed-adverse effect levels, and a margin of exposure (MOE) calculation. The initial chemical screen focuses on the chemical presence in specific media that are special to children, such as foods children regularly eat and drink, residential or school air, products children use, and soil and dust in and around residences. Data from the literature or from models serve as the initial exposure estimate. This methodology would allow us to focus on those chemicals to which children are most exposed that are also associated with, potentially, the highest risk. Use of the MOE calculation allows for comparison among chemicals, prioritization of chemicals for evaluation and testing, and identification of significant data gaps. PMID- 10856019 TI - Children's exposure assessment: a review of factors influencing Children's exposure, and the data available to characterize and assess that exposure. AB - We review the factors influencing children's exposure to environmental contaminants and the data available to characterize and assess that exposure. Children's activity pattern data requirements are demonstrated in the context of the algorithms used to estimate exposure by inhalation, dermal contact, and ingestion. Currently, data on children's exposures and activities are insufficient to adequately assess multimedia exposures to environmental contaminants. As a result, regulators use a series of default assumptions and exposure factors when conducting exposure assessments. Data to reduce uncertainty in the assumptions and exposure estimates are needed to ensure chemicals are regulated appropriately to protect children's health. To improve the database, advancement in the following general areas of research is required: identification of appropriate age/developmental benchmarks for categorizing children in exposure assessment; development and improvement of methods for monitoring children's exposures and activities; collection of activity pattern data for children (especially young children) required to assess exposure by all routes; collection of data on concentrations of environmental contaminants, biomarkers, and transfer coefficients that can be used as inputs to aggregate exposure models. PMID- 10856021 TI - Risk of childhood leukemia associated with diagnostic irradiation and polymorphisms in DNA repair genes. AB - The purpose of the study was to measure risk of childhood acute lymphoblastic leukemia associated with reported postnatal diagnostic X rays and to determine if it was modified in the presence of variants in genes involved in DNA repair. We conducted a population-based case-control study with 491 cases and 491 healthy controls among children 0-9 years of age at diagnosis. To evaluate gene environment interaction, we used a subgroup of 129 cases. The adjusted odds ratio (OR) for one reported postnatal child X ray versus none was 1.04 [95% confidence interval (CI), 0.72-1.49], whereas the OR for two or more X rays was 1.61 (CI, 1.13-2.28). Among girls, the former ORs were 1.14 (CI, 0.66-1.96) and 2.26 (1. 20 4.23), respectively. Among girls who carried the hMSH3 [exon (ex) 23] variant, the ORs were 3.33 (CI, 0.75-14.82) for one X ray and 0. 27 (CI, 0.05-1.57) for two or more X rays, whereas among those who carried the XRCCI (ex 6) variant, the ORs were 1.45 (0.11-19.08) and 6.66 (0.78-56.63), respectively. On the other hand, at low levels of exposure, boys seemed protected by the variant hMLH1 (ex 8). The latter results must be interpreted with caution but suggest that the effect of diagnostic X rays could be modified by variants in repair genes according to sex. Few studies have evaluated the risk of postnatal diagnostic irradiation, which was moderately strong here; we are not aware of any studies that also considered the effect of polymorphisms in DNA repair genes. Based on the present results, both aspects deserve further study. PMID- 10856020 TI - Endocrine disruptors and human health--is there a problem? An update. AB - It has been hypothesized that environmental exposure to synthetic estrogenic chemicals and related endocrine-active compounds may be responsible for a global decrease in sperm counts, decreased male reproductive capacity, and breast cancer in women. Results of recent studies show that there are large demographic variations in sperm counts within countries or regions, and analyses of North American data show that sperm counts have not decreased over the last 60 years. Analyses of records for hypospadias and cryptorchidism also show demographic differences in these disorders before 1985; however, since 1985 rates of hypospadias have not changed and cryptorchidism has actually declined. Temporal changes in sex ratios and fertility are minimal, whereas testicular cancer is increasing in most countries; however, in Scandinavia, the difference between high (Denmark) and low (Finland) incidence areas are not well understood and are unlikely to be correlated with differences in exposure to synthetic industrial chemicals. Results from studies on organochlorine contaminants (DDE/PCB) show that levels were not significantly different in breast cancer patients versus controls. Thus, many of the male and female reproductive tract problems linked to the endocrine-disruptor hypothesis have not increased and are not correlated with synthetic industrial contaminants. This does not exclude an endocrine-etiology for some adverse environmental effects or human problems associated with high exposures to some chemicals. PMID- 10856022 TI - Childhood cancer in the offspring of male sawmill workers occupationally exposed to chlorophenate fungicides. AB - The objective of this study was to determine whether paternal occupational exposure to chlorophenol fungicides and their dioxin contaminants is associated with childhood cancer in the offspring of sawmill workers. We used data from 23,829 British Columbian sawmill workers employed for at least 1 continuous year between 1950 and 1985 in 11 sawmills that used chlorophenates. Probabilistic linkage of the sawmill worker cohort to the provincial marriage and birth files produced an offspring cohort of 19,674 children born at least 1 year after the initiation of employment in the period 1952-1988. We then linked the offspring cohort to the British Columbia Cancer Registry. We included all malignancies in cases younger than 20 years of age that appeared on the cancer registry between 1969 and 1993. We calculated standardized incidence ratios (SIRs) using the British Columbia population as a reference. A nested case-control analysis assessed the effects of paternal cumulative exposure and windows of exposure on the risk of developing cancer in the offspring. We identified 40 cases of cancer during 259,919 person-years of follow-up. The all-cancer SIR was 1.0 [95% confidence interval (CI), 0.7-1.4]; the SIR for leukemia was 1.0 (CI, 0.5-1.8); and the SIR for brain cancer was 1.3 (CI, 0.6-2.5). The nested case-control analysis showed slightly increased risks in the highest categories of chlorophenol exposure, although none was statistically significant. Our analyses provide little evidence to support a relationship between the risk of childhood cancer and paternal occupational exposure to chlorophenate fungicides in British Columbian sawmills. PMID- 10856023 TI - A modeling framework for estimating children's residential exposure and dose to chlorpyrifos via dermal residue contact and nondietary ingestion. AB - To help address the Food Quality Protection Act of 1996, a physically based probabilistic model has been developed to quantify and analyze dermal and nondietary ingestion exposure and dose to pesticides. The Residential Stochastic Human Exposure and Dose Simulation Model for Pesticides (Residential-SHEDS) simulates the exposures and doses of children contacting residues on surfaces in treated residences and on turf in treated residential yards. The simulations combine sequential time-location-activity information from children's diaries with microlevel videotaped activity data, probability distributions of measured surface residues and exposure factors, and pharmacokinetic rate constants. Model outputs include individual profiles and population statistics for daily dermal loading, mass in the blood compartment, ingested residue via nondietary objects, and mass of eliminated metabolite, as well as contributions from various routes, pathways, and media. To illustrate the capabilities of the model framework, we applied Residential-SHEDS to estimate children's residential exposure and dose to chlorpyrifos for 12 exposure scenarios: 2 age groups (0-4 and 5-9 years); 2 indoor pesticide application methods (broadcast and crack and crevice); and 3 postindoor application time periods (< 1, 1-7, and 8-30 days). Independent residential turf applications (liquid or granular) were included in each of these scenarios. Despite the current data limitations and model assumptions, the case study predicts exposure and dose estimates that compare well to measurements in the published literature, and provides insights to the relative importance of exposure scenarios and pathways. PMID- 10856024 TI - Biologically based pesticide dose estimates for children in an agricultural community. AB - Current pesticide health risk assessments in the United States require the characterization of aggregate exposure and cumulative risk in the setting of food tolerances. Biologic monitoring can aggregate exposures from all sources and routes, and can integrate exposures for chemicals with a common mechanism of action. Its value was demonstrated in a recent study of organophosphorus (OP) pesticide exposure among 109 children in an agricultural community in Washington State; 91 of the children had parents working in agriculture. We estimated individual OP pesticide doses from urinary metabolite concentrations with a deterministic steady state model, and compared them to toxicologic reference values. We evaluated doses by assuming that metabolites were attributable entirely to either azinphos-methyl or phosmet, the two OP pesticides used most frequently in the region. Creatinine-adjusted average dose estimates during the 6 to 8-week spraying season ranged from 0 to 36 microg/kg/day. For children whose parents worked in agriculture as either orchard applicators or as fieldworkers, 56% of the doses estimated for the spray season exceeded the U.S. Environmental Protection Agency (EPA) chronic dietary reference dose, and 19% exceeded the World Health Organization acceptable daily intake values for azinphos-methyl. The corresponding values for children whose parents did not work in agriculture were 44 and 22%, respectively. The percentage of children exceeding the relevant reference values for phosmet was substantially lower (< 10%). Single-day dose estimates ranged from 0 to 72 microg/kg/day, and 26% of these exceeded the EPA acute reference dose for azinphos-methyl. We also generated dose estimates by adjustment for total daily urine volume, and these estimates were consistently higher than the creatinine-adjusted estimates. None of the dose estimates exceeded the empirically derived no-observable-adverse-effect levels for these compounds. The study took place in an agricultural region during a period of active spraying, so the dose estimates for this population should not be considered representative of exposures in the general population. The findings indicate that children living in agricultural regions represent an important subpopulation for public health evaluation, and that their exposures fall within a range of regulatory concern. They also demonstrate that biologically based exposure measures can provide data for health risk evaluations in such populations. PMID- 10856025 TI - Biologic monitoring of exposure to organophosphorus pesticides in 195 Italian children. AB - One hundred ninety-five 6- to 7-year-old children who lived in the municipality of Siena (Tuscany, Italy), underwent biologic monitoring to evaluate urinary excretion of several alkylphosphates that are metabolites of organophosphorus pesticides. We evaluated dimethylphosphate (DMP), dimethylthiophosphate (DMTP), dimethyldithiophosphate (DMDTP), diethylphosphate (DEP), diethylthiophosphate (DETP), and diethyldithiophosphate (DEDTP). We obtained urine samples taken in the children's schools, and each sample was accompanied by a questionnaire about lifestyle and dietary habits. We found DMP and DMTP in detectable concentrations in the greatest number of samples (96 and 94%, respectively). The DMP values were geometric mean (GM) 116.7, [geometric standard deviation (GSD) 2.5], and a range of 7.4-1,471.5 nmol/g creatinine. The corresponding DMTP values were GM 104.3 (GSD 2.8) and a range of 4.0-1,526.0 nmol/g creatinine. DMDTP, DEP, DETP, and DEDTP concentrations were GM 14.1, (GSD 3.0), and a range of 3.3-754.6 nmol/g creatinine in 34% of the children; GM 33.2, (GSD 2.4), and a range of 5.1-360.1 nmol/g creatinine in 75% of the children; GM 16.0, (GSD 2.9), and a range of 3.1 284.7 in 48% of the children; and GM 7.7, (GSD 2.1), and a range of 2.3-140.1 in 12% of the children, respectively. The significant variable for urinary excretion of these metabolites in children was pest control operations performed inside or outside the house in the preceding month; however, the presence of a vegetable garden near the house rarely emerged. The urinary excretion of alkylphosphates in children was significantly higher than in a group of the adult population resident in the same province. PMID- 10856026 TI - Effects of lead exposure before pregnancy and dietary calcium during pregnancy on fetal development and lead accumulation. AB - Millions of women of child-bearing age have substantial bone lead stores due to lead exposure as children. Dietary calcium ingested simultaneously with lead exposure can reduce lead absorption and accumulation. However, the effects of dietary calcium on previously accumulated maternal lead stores and transfer to the fetus have not been investigated. We studied the effects of lead exposure of female rats at an early age on fetal development during a subsequent pregnancy. We gave 5-week-old female Sprague-Dawley rats lead as the acetate in their drinking water for 5 weeks; controls received equimolar sodium acetate. This was followed by a 1-month period without lead exposure before mating. We randomly assigned pregnant rats (n = 39) to diets with a deficient (0.1%) or normal (0.5%) calcium content during pregnancy. A total of 345 pups were delivered alive. Lead exposed dams and their pups had significantly higher blood lead concentrations than controls, but the concentrations were in the range of those found in many pregnant women. Pups born to dams fed the calcium-deficient diet during pregnancy had higher blood and organ lead concentrations than pups born to dams fed the 0. 5% calcium diet. Pups born to lead-exposed dams had significantly (p<0.0001) lower mean birth weights and birth lengths than controls. There were significant inverse univariate associations between dam or pup organ lead concentrations and birth weight or length. The 0.5% calcium diet did not increase in utero growth. Stepwise regression analysis demonstrated that greater litter size and female sex were significantly associated with reduced pup birth weight and length. However, lead exposure that ended well before pregnancy was significantly (p<0.0001) associated with reduced birth weight and length, even after litter size, pup sex, and dam weight gain during pregnancy were included in the regression analysis. The data demonstrate that an increase in dietary calcium during pregnancy can reduce fetal lead accumulation but cannot prevent lead-induced decreases in birth weight and length. The results provide evidence that dietary nutrients can influence the transfer of toxins to the fetus during pregnancy. If these results are applicable to women, an increase in diet calcium during pregnancy could reduce the transfer of lead from prepregnancy maternal exposures to the fetus. PMID- 10856027 TI - Serum clara cell protein: a sensitive biomarker of increased lung epithelium permeability caused by ambient ozone. AB - Ozone in ambient air may cause various effects on human health, including decreased lung function, asthma exacerbation, and even premature mortality. These effects have been evidenced using various clinical indicators that, although sensitive, do not specifically evaluate the O(3)-increased lung epithelium permeability. In the present study, we assessed the acute effects of ambient O(3) on the pulmonary epithelium by a new approach relying on the assay in serum of the lung-specific Clara cell protein (CC16 or CC10). We applied this test to cyclists who exercised for 2 hr during episodes of photochemical smog and found that O(3) induces an early leakage of lung Clara cell protein. The protein levels increased significantly into the serum from exposure levels as low as 0.060-0.084 ppm. Our findings, confirmed in mice exposed to the current U.S. National Ambient Air Quality Standards for O(3) (0.08 ppm for 8 hr) indicate that above the present natural background levels, there is almost no safety margin for the effects of ambient O(3) on airway permeability. The assay of CC16 in the serum represents a new sensitive noninvasive test allowing the detection of early effects of ambient O(3) on the lung epithelial barrier. PMID- 10856028 TI - Onset of spermatogenesis is accelerated by gestational administration of 1,2,3,4,6,7-hexachlorinated naphthalene in male rat offspring. AB - We treated pregnant rats with 1 microg/kg body weight/day 1,2,3,4,6,7 hexachlorinated naphthalene (1,2,3,4,6,7-HxCN) on days 14-16 of gestation and examined the effects on the reproductive systems of their male offspring at various phases of sexual maturation. Sperm count in the cauda epididymidis did not change in 1,2,3,4,6, 7-HxCN-treated rats on postnatal day 89, the age of sexual maturity, but the sperm count in the cauda epididymidis did increase to approximately 180% of the control value on postnatal day 62. In addition, homogenization-resistant testicular spermatids increased to approximately 160% of the control value on postnatal day 48, and the percent of postmeiotic tubules increased to approximately 190% of the control value on postnatal day 31 in this group. These results indicate that the onset of spermatogenesis was accelerated in the 1,2,3,4,6,7-HxCN rats. Serum concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) had already reached the maximum level on postnatal day 31 in the 1,2,3,4,6, 7-HxCN group, suggesting that the onset of LH and FSH secretions from the pituitary gland was also accelerated and that this endocrine disruption was the cause of early onset of spermatogenesis in this group. In the fat of 1,2,3,4,6,7-HxCN-treated dams, 5.75+/-2.81 ppb 1,2,3,4,6,7 HxCN was detected when offspring were weaned. This concentration was 5-10 times higher than that found in human adipose tissue. PMID- 10856029 TI - Mortality among the residents of the Three Mile Island accident area: 1979-1992. AB - The largest U.S. population exposed to low-level radioactivity released by an accident at a nuclear power plant is composed of residents near the Three Mile Island (TMI) Plant on 28 March 1979. This paper (a collaboration of The University of Pittsburgh and the Pennsylvania Department of Health) reports on the mortality experience of the 32,135 members in this cohort for 1979-1992. We analyzed standardized mortality ratios (SMRs) using a local comparison population and performed relative risk regression modeling to assess overall mortality and specific cancer risks by confounding factors and radiation-related exposure variables. Total mortality was significantly elevated for both men and women (SMRs = 109 and 118, respectively). All heart disease accounted for 43.3% of total deaths and demonstrated elevated SMRs for heart disease of 113 and 130 for men and women, respectively; however, when controlling for confounders and natural background radiation, these elevations in heart disease were no longer evident. Overall cancer mortality was similar in this cohort as compared to the local population (male SMR = 100; female SMR = 101). In the relative risk modeling, there was a significant effect for all lymphatic and hematopoietic tissue in males in relation to natural background exposure (p = 0.04). However, no trend was noted. We found a significant linear trend for female breast cancer risk in relation to increasing levels of TMI-related likely [gamma]-exposure (p = 0.02). Although such a relationship has been noted in other investigations, emissions from the TMI incident were significantly lower than in other documented studies. Therefore, it is unlikely that this observed increase is related to radiation exposure on the day of the accident. The mortality surveillance of this cohort does not provide consistent evidence that radioactivity released during the TMI accident has a significant impact on the mortality experience of this cohort to date. However, continued follow-up of these individuals will provide a more comprehensive description of the morbidity and mortality experience of the cohort. PMID- 10856030 TI - Comparison of chemical-activated luciferase gene expression bioassay and gas chromatography for PCB determination in human serum and follicular fluid. AB - We assessed exposure to dioxin-like compounds using chemical and bioassay analysis in different matrices in a female population. A total of 106 serum and 9 follicular fluid samples were collected from infertile women attending Centers for Reproductive Medicine in Belgium from 1996 to 1998. Major polychlorinated biphenyl (PCB) congeners were quantified by chemical analysis using gas chromatography with electron-capture detection, and the chemical-activated luciferase gene expression (CALUX) bioassay was used to determine the total dioxin-like toxic equivalence (TEQ) of mixtures of polyhalogenated aromatic hydrocarbons present in body fluids, such as serum and follicular fluid. To the best of our knowledge, this is the first investigation to determine TEQ values by the CALUX bioassay in follicular fluid. The TEQ levels in both matrices are well correlated (r = 0.83, p = 0.02). As the chemical and bioassay analysis executed in this study do not cover the same span of polyhalogenated aromatic hydrocarbons, we did not expect totally correlated results. Moreover, the sample workup and quantification of the analytes differed completely. Nonetheless, the TEQ values in human extracts correlated well with the sum of four major PCB congeners chemically determined in both serum and follicular fluid. These results indicate that the CALUX bioassay may serve as a simple, relatively inexpensive prescreening tool for exposure assessment in epidemiologic surveys. PMID- 10856031 TI - Genotoxic effects of alpha-endosulfan and beta-endosulfan on human HepG2 cells. AB - alpha-Endosulfan and ss-endosulfan are isomers of endosulfan, a pesticide used worldwide. In this study, we examined the genotoxicity of [alpha]- and ss endosulfan in vitro with a HepG2 cell line. We used sister chromatid exchanges (SCE), micronuclei (MN), and DNA strand breaks as detected by single-cell gel electrophoresis (SCG) assays as biomarkers to judge the genotoxicity of [alpha]- and ss-endosulfan at concentrations from 1 times 10(-12) M to 1 times 10(-3) M. After treating HepG2 cells for 48 hr with ss-endosulfan, SCE showed a significant increase at concentrations from 1 times 10(-7) M to 1 times 10(-5) M, and MN showed a significant increase at concentrations from 5 times 10(-5) M to 1 times 10(-3) M. [alpha]-Endosulfan failed to show significant effects in both the SCE and MN assays. After treating HepG2 cells with [alpha]- or ss-endosulfan for 1 hr, DNA strand breaks were significantly induced by [alpha]-endosulfan at concentrations from 2 times 10(-4) M to 1 times 10(-3) M, and by ss-endosulfan at 1 times 10(-3) M. The results of this study suggest that both [alpha]- and ss endosulfan are genotoxic to HepG2 cells and that the genotoxicity of ss endosulfan seems stronger than that of [alpha]-endosulfan. PMID- 10856032 TI - Assessing confounding, effect modification, and thresholds in the association between ambient particles and daily deaths. AB - I examined the relationship between daily deaths and airborne particles in 10 U.S. cities with varying climatic conditions and seasons in which particle concentrations were high. Airborne particles were associated with significant increases in daily deaths [0.67% increase for a 10 microg/m(3) increase in particles; 95% confidence interval (CI), 0.52-0.81%]. This association was the same in summer and winter. To examine potential confounding by other pollutants, I regressed city- and season-specific effect sizes against the relationship between airborne particles and other pollutants. Controlling for other pollutants did not substantially (or significantly) change the estimated effect of airborne particles. Socioeconomic differences between cities likewise did not modify the effect. The increase in daily deaths that occurred out of hospitals (0.89% per 10 microg/m(3); CI, 0.67-1.10%) was substantially greater than the increase in deaths in hospitals (0. 49%; CI, 0.31-0.68%). This is consistent with results previously reported in Philadelphia, Pennsylvania, and suggests that the particle associated deaths are not just being brought forward by a few days. It is also consistent with recent animal and human studies of the mechanisms of particle toxicity. PMID- 10856033 TI - Variability in airborne and biological measures of exposure to mercury in the chloralkali industry: implications for epidemiologic studies. AB - Exposure assessment is a critical component of epidemiologic studies, and more sophisticated approaches require that variation in exposure be considered. We examined the intra- and interindividual sources of variation in exposure to mercury vapor as measured in air, blood, and urine among four groups of workers during 1990-1997 at a Swedish chloralkali plant. Consistent with the underlying kinetics of mercury in the body, the variability of biological measures was dampened considerably relative to the variation in airborne levels. Owing to the effects of intraindividual variation, estimating workers' exposures from a few measurements can attenuate measures of effect. To examine such effects on studies relating long-term exposure to a continuous health outcome, we evaluated the utility of each exposure measure by comparing the necessary sample sizes required for accurate estimation of a slope coefficient obtained from a regression analysis. No single measure outperformed the others for all groups of workers. However, when workers were evaluated together, creatinine-corrected urinary mercury better discriminated workers' exposures than airborne or blood mercury levels. Thus, pilot studies should be conducted to examine variability in both air and biomonitoring data because quantitative information about the relative magnitude of the intra- and interindividual sources of variation feeds directly into our efforts to design an optimal sampling strategy when evaluating health risks associated with occupational or environmental contaminants. PMID- 10856034 TI - A cluster of pediatric metallic mercury exposure cases treated with meso-2,3 dimercaptosuccinic acid (DMSA) AB - Nine children and their mother were exposed to vapors of metallic mercury. The source of the exposure appears to have been a 6-oz vial of mercury taken from a neighbor's home. The neighbor reportedly operated a business preparing mercury filled amulets for practitioners of the Afro-Caribbean religion Santeria. At diagnosis, urinary mercury levels in the children ranged from 61 to 1,213 microg/g creatinine, with a geometric mean of 214.3 microg/m creatinine. All of the children were asymptomatic. To prevent development of neurotoxicity, we treated the children with oral meso-2,3-dimercaptosuccinic acid (DMSA). During chelation, the geometric mean urine level rose initially by 268% to 573.2 microg mercury/g creatinine (p<0.0005). At the 6-week follow-up examination after treatment, the geometric mean urine mercury level had fallen to 102.1 microg/g creatinine, which was 17.8% of the geometric mean level observed during treatment (p<0.0005) and 47.6% of the original baseline level (p<0.001). Thus, oral chelation with DMSA produced a significant mercury diuresis in these children. We observed no adverse side effects of treatment. DMSA appears to be an effective and safe chelating agent for treatment of pediatric overexposure to metallic mercury. PMID- 10856035 TI - Children's health: a mixed review. PMID- 10856036 TI - Do urban environmental pollutants exacerbate childhood lung diseases? PMID- 10856037 TI - NIEHS teaches by example. PMID- 10856038 TI - Getting in touch with your inner-city child. PMID- 10856039 TI - NIEHS investigates Arctic health issues. PMID- 10856040 TI - Disturbing behavior: neurotoxic effects in children. PMID- 10856041 TI - Chemical regulation and kids: In search of a better fit. PMID- 10856042 TI - Midline versus mediolateral episiotomy. PMID- 10856043 TI - Fix what's wrong, not what's right, with general practice in Britain. PMID- 10856044 TI - Suicidal behaviour in gay, lesbian, and bisexual youth. PMID- 10856045 TI - How living wills can help doctors and patients talk about dying. PMID- 10856046 TI - Towards better treatment of glaucoma. PMID- 10856047 TI - One in six children live in relative poverty. PMID- 10856048 TI - Senior doctors split over general medical council. PMID- 10856049 TI - Pallidotomy relieves some symptoms of Parkinson's disease PMID- 10856050 TI - Antibodies can repair damaged myelin in model of MS PMID- 10856051 TI - In brief PMID- 10856053 TI - Chinese herb may cause cancer PMID- 10856052 TI - Gynaecologist banned in Canada appears before GMC. PMID- 10856054 TI - WHO warns of threat of "superbugs". PMID- 10856055 TI - Clinical evidence to go to 400000 US doctors. PMID- 10856056 TI - Supply of generic drugs still unreliable. PMID- 10856057 TI - Budget cuts may have led to E coli outbreak. PMID- 10856058 TI - Private finance scheme for Worcester "will cut beds". PMID- 10856059 TI - Clinton orders medicare to cover patients in clinical trials. PMID- 10856060 TI - More doctors criticise the GMC. PMID- 10856061 TI - Claim that events before birth cause cerebral palsy is disputed. PMID- 10856062 TI - Cost effectiveness analysis of screening for sight threatening diabetic eye disease. AB - OBJECTIVE: To measure the cost effectiveness of systematic photographic screening for sight threatening diabetic eye disease compared with existing practice. DESIGN: Cost effectiveness analysis. SETTING: Liverpool. SUBJECTS: A target population of 5000 diabetic patients invited for screening. MAIN OUTCOME MEASURES: Cost effectiveness (cost per true positive) of systematic and opportunistic programmes; incremental cost effectiveness of replacing opportunistic with systematic screening. RESULTS: Baseline prevalence of sight threatening eye disease was 14.1%. The cost effectiveness of the systematic programme was 209 pound sterling (sensitivity 89%, specificity 86%, compliance 80%, annual cost 104996 pound sterling) and of the opportunistic programme was 289 pound sterling (combined sensitivity 63%, specificity 92%, compliance 78%, annual cost 99 981 pound sterling). The incremental cost effectiveness of completely replacing the opportunistic programme was 32 pound sterling. Absolute values of cost effectiveness were highly sensitive to varying prevalence, sensitivity and specificity, compliance, and programme size. CONCLUSION: Replacing existing programmes with systematic screening for diabetic eye disease is justified. PMID- 10856063 TI - Disability in young people and adults one year after head injury: prospective cohort study. AB - OBJECTIVE: To determine the frequency of disability in young people and adults admitted to hospital with a head injury and to estimate the annual incidence in the community. DESIGN: Prospective, hospital based cohort study, with one year follow up of sample stratified by coma score. SETTING: Five acute hospitals in Glasgow. SUBJECTS: 2962 patients (aged 14 years or more) with head injury; 549 (71%) of the 769 patients selected for follow up participated. MAIN OUTCOME MEASURES: Glasgow outcome scale and problem orientated questionnaire. RESULTS: Survival with moderate or severe disability was common after mild head injury (47%, 95% confidence interval 42% to 52%) and similar to that after moderate (45%, 35% to 56%) or severe injury (48%, 36% to 60%). By extrapolation from the population identified (90% of whom had mild injuries), it was estimated that annually in Glasgow (population 909 498) 1400 young people and adults are still disabled one year after head injury. CONCLUSION: The incidence of disability in young people and adults admitted with a head injury is higher than expected. This reflects the high rate of sequelae previously unrecognised in the large number of patients admitted to hospital with an apparently mild head injury. PMID- 10856064 TI - US women's attitudes to false positive mammography results and detection of ductal carcinoma in situ: cross sectional survey. AB - OBJECTIVE: To determine women's attitudes to and knowledge of both false positive mammography results and the detection of ductal carcinoma in situ after screening mammography. DESIGN: Cross sectional survey. SETTING: United States. PARTICIPANTS: 479 women aged 18-97 years who did not report a history of breast cancer. MAIN OUTCOME MEASURES: Attitudes to and knowledge of false positive results and the detection of ductal carcinoma in situ after screening mammography. RESULTS: Women were aware that false positive results do occur. Their median estimate of the false positive rate for 10 years of annual screening was 20% (25th percentile estimate, 10%; 75th percentile estimate, 45%). The women were highly tolerant of false positives: 63% thought that 500 or more false positives per life saved was reasonable and 37% would tolerate 10 000 or more. Women who had had a false positive result (n=76) expressed the same high tolerance: 39% would tolerate 10 000 or more false positives. 62% of women did not want to take false positive results into account when deciding about screening. Only 8% of women thought that mammography could harm a woman without breast cancer, and 94% doubted the possibility of non-progressive breast cancers. Few had heard about ductal carcinoma in situ, a cancer that may not progress, but when informed, 60% of women wanted to take into account the possibility of it being detected when deciding about screening. CONCLUSIONS: Women are aware of false positives and seem to view them as an acceptable consequence of screening mammography. In contrast, most women are unaware that screening can detect cancers that may never progress but feel that such information would be relevant. Education should perhaps focus less on false positives and more on the less familiar outcome of detection of ductal carcinoma in situ. PMID- 10856065 TI - Views of elderly people on living wills: interview study. PMID- 10856066 TI - Two case histories PMID- 10856067 TI - Effect of anti-inflammatory drugs on overall risk of common cancer: case-control study in general practice research database. AB - OBJECTIVE: To examine whether anti-inflammatory drug treatment protects against the commoner cancers in the United Kingdom. DESIGN: Case-control study using the general practice research database. SETTING: Practices throughout United Kingdom providing data to the database. SUBJECTS: Patients who had a first diagnosis of five gastrointestinal (oesophagus, stomach, colon, rectum, and pancreas) cancers and four non-gastrointestinal (bladder, breast, lung, and prostate) cancers in 1993-5 for whom prescription data were available for the at least the previous 36 months. Each case was matched for age, sex, and general practice with three controls. MAIN OUTCOME MEASURE: Risk of cancer. RESULTS: In 12 174 cancer cases and 34 934 controls overall risk of the nine cancers was not significantly reduced among those who had received at least seven prescriptions in the 13-36 months before cancer diagnosis (odds ratio 0.98, 95% confidence interval 0.89 to 1.07). Findings were nevertheless compatible with protective effects from anti inflammatory drugs against cancers of the oesophagus (0.64, 0. 41 to 0.98), stomach (0.51, 0.33 to 0.79), colon (0.76, 0.58 to 1. 00), and rectum (0.75, 0.49 to 1.14) with dose related trends. The risk of pancreatic cancer (1.49, 1.02 to 2.18) and prostatic cancer (1.33, 1.07 to1.64) was increased among patients who had received at least seven prescriptions, but the trend was dose related for only pancreatic cancer. CONCLUSIONS: Anti-inflammatory drugs may protect against oesophageal and gastric cancer as well as colon and rectal cancer. The increased risks of pancreatic and prostatic cancer could be due to chance or to undetected biases and warrant further investigation. PMID- 10856068 TI - There's more inside the tablet PMID- 10856070 TI - A wise headmaster PMID- 10856069 TI - Diagnosis and management of porphyria. PMID- 10856071 TI - ABC of oral health. Oral health and disease. PMID- 10856072 TI - A message from holland PMID- 10856075 TI - The BMA from a general practitioner's point of view PMID- 10856073 TI - Appropriate feeding methods for infants of HIV infected mothers in sub-Saharan Africa. PMID- 10856074 TI - Patient care (empowerment): the view from a national society. PMID- 10856078 TI - Public health conference PMID- 10856077 TI - Obituaries PMID- 10856076 TI - Patient care (empowerment): a local view. PMID- 10856079 TI - Teaching made easy: A manual for health professionals PMID- 10856081 TI - Tarring consultants with the ledward brush PMID- 10856080 TI - The origin of captured images PMID- 10856082 TI - Transplant organs PMID- 10856083 TI - Priority setting in health care: should we ask the tax payer? PMID- 10856084 TI - Why seeing a patient after the counsellor is so difficult PMID- 10856085 TI - A week of overexcitement PMID- 10856086 TI - Systematic screening for diabetic eye disease is cost effective PMID- 10856087 TI - Almost half of adults with a head injury suffer long term disability PMID- 10856089 TI - Most elderly people inpatients haven't heard of living wills PMID- 10856088 TI - Women are unaware that mammography may detect non-progressive cancer PMID- 10856090 TI - NSAIDs may reduce risk of gastrointestinal cancers PMID- 10856091 TI - Mixed feeding may be the most dangerous option for babies of HIV positive mothers in africa PMID- 10856092 TI - What does an R&D lead do? PMID- 10856093 TI - The policing of clinical trials PMID- 10856095 TI - Phase I/II trial of cyclophosphamide, mitoxantrone, and escalated doses of carboplatin supported by peripheral-blood stem cells in women with metastatic breast cancer. AB - PURPOSE: To intensify a regimen of high-dose cyclophosphamide, mitoxantrone, and carboplatin that had previously produced high complete and overall response rates in metastatic breast cancer (MBC). PATIENTS AND METHODS: Forty-four patients with a median age of 43 years (range, 25 to 57 years) and previously untreated MBC who were responding to anthracycline-based or single-agent taxane chemotherapy received cyclophosphamide 1.5 g/m(2)/d and mitoxantrone 16 mg/m(2)/d combined with escalating doses of carboplatin 200 to 500 mg/m(2)/d, each given daily from days -6 to -3. Hematopoiesis was supported by mobilized peripheral-blood stem cells infused on day 0 and by use of granulocyte-macrophage colony-stimulating factor 300 microg/d subcutaneously starting on day 1. RESULTS: A total of six dose levels of carboplatin were examined. Grades 3 and 4 toxicity occurred in 10 patients and one patient, respectively, with grade 3 toxicity occurring in only five of 31 patients treated with < or = 400 mg/m(2) of carboplatin. Major dose limiting toxicities were cardiac, pulmonary, and renal. Four patients developed congestive heart failure: two had persistently low ejection fraction 11 and 36 months after peripheral-blood stem-cell transplantation (PBSCT), and two recovered. Hematologic recovery to an absolute neutrophil count of greater than 0.5 x 10(9)/L occurred at a median of 11 days (range, 8 to 25 days) and to a platelet count of greater than 20 x 10(9)/L at a median of 10.5 days (range, 6 to 60 days). There were two toxic deaths from sepsis: one on day 27 (level 5) and one from cardiac arrest on day 42 (level 6). CONCLUSION: The maximum-tolerated dose of carboplatin was 400 mg/m(2)/d in combination with mitoxantrone 16 mg/m(2)/d and cyclophosphamide 1,500 mg/m(2), all drugs given over 4 days. This regimen is being tested in a phase III trial of high-dose chemotherapy and PBSCT versus standard treatment. PMID- 10856094 TI - Randomized phase III trial of docetaxel versus vinorelbine or ifosfamide in patients with advanced non-small-cell lung cancer previously treated with platinum-containing chemotherapy regimens. The TAX 320 Non-Small Cell Lung Cancer Study Group. AB - PURPOSE: To confirm the promising phase II results of docetaxel monotherapy, this phase III trial was conducted of chemotherapy for patients with advanced non small-cell lung cancer (NSCLC) who had previously failed platinum-containing chemotherapy. PATIENTS AND METHODS: A total of 373 patients were randomized to receive either docetaxel 100 mg/m(2) (D100) or 75 mg/m(2) (D75) versus a control regimen of vinorelbine or ifosfamide (V/I). The three treatment groups were well balanced for key patient characteristics. RESULTS: Overall response rates were 10.8% with D100 and 6.7% with D75, each significantly higher than the 0.8% response with V/I (P =.001 and P =.036, respectively). Patients who received docetaxel had a longer time to progression (P =.046, by log-rank test) and a greater progression-free survival at 26 weeks (P =.005, by chi(2) test). Although overall survival was not significantly different between the three groups, the 1 year survival was significantly greater with D75 than with the control treatment (32% v 19%; P =.025, by chi(2) test). Prior exposure to paclitaxel did not decrease the likelihood of response to docetaxel, nor did it impact survival. There was a trend toward greater efficacy in patients whose disease was platinum resistant rather than platinum-refractory and in patients with performance status of 0 or 1 versus 2. Toxicity was greatest with D100, but the D75 arm was well tolerated. CONCLUSION: This first randomized trial in this setting demonstrates that D75 every 3 weeks can offer clinically meaningful benefit to patients with advanced NSCLC whose disease has relapsed or progressed after platinum-based chemotherapy. PMID- 10856096 TI - Phase II trial of doxorubicin and docetaxel plus granulocyte colony-stimulating factor in metastatic breast cancer: Eastern Cooperative Oncology Group Study E1196. AB - PURPOSE: The purpose of this multi-institutional phase II trial was to evaluate the efficacy and toxicity of doxorubicin and docetaxel plus granulocyte colony stimulating factor (G-CSF) in patients with metastatic breast cancer. The primary objective was to determine whether the combination produced a response rate of at least 50%. PATIENTS AND METHODS: Fifty-four patients with metastatic breast cancer received doxorubicin (60 mg/m(2) by intravenous [IV] injection) followed 1 hour later by docetaxel (60 mg/m(2) by IV infusion over 1 hour) every 3 weeks for up to eight cycles. All patients also received G-CSF. RESULTS: Objective response occurred in 29 (57%) of 51 eligible patients (95% confidence interval [CI], 42% to 70%), including three patients who had a complete response (6%; 95% CI, 1% to 16%). The median response duration was 7 months (95% CI, 6.0 to 15.0 months), median time to treatment failure was 7. 6 months (95% CI, 6.2 to 9.9 months), and the median survival was 27. 5 months (95% CI, 21.5 months to upper limit not reached). The median cumulative doxorubicin dose was 395 mg/m(2) (range, 60 to 480 mg/m(2)). Fifteen patients (28%) were documented to have a decrease in the left ventricular ejection fraction below normal, and three patients (6%; 95% CI, 1% to 15%) developed congestive heart failure. CONCLUSION: Using criteria that we had defined a priori, the doxorubicin-docetaxel regimen as used in this study was sufficiently active and tolerable to justify a phase III comparison with doxorubicin-cyclophosphamide in early-stage breast cancer. PMID- 10856097 TI - Low-dose oral fluorouracil with eniluracil as first-line chemotherapy against advanced breast cancer: a phase II study. AB - PURPOSE: Eniluracil (776C85) is an effective inactivator of dihydropyrimidine dehydrogenase that allows continuous low-dose oral fluorouracil (5-FU) to be given with predicable oral bioavailability. We have assessed this as first-line oral chemotherapy for patients with advanced/metastatic breast cancer. PATIENTS AND METHODS: Patients with histologically proven, locally advanced or metastatic breast cancer without previous chemotherapy for advanced disease were entered onto this open-label phase II study. Patients received oral 5-FU 1.0 mg/m(2) with eniluracil 10 mg/m(2), both given twice daily for the first 28 days of each 35 day cycle, continuing until disease progression or unmanageable toxicity. RESULTS: Thirty-three patients were entered, with a median age of 53 years. Sixteen partial responses were seen in twenty-nine assessable patients (55%; 95% confidence interval, 37% to 73%), including responses in four (40%) out of 10 patients who had received prior adjuvant 5-FU. Seven patients had stable disease for at least 3 months with symptom improvement. Median response duration was 14 months (range, 10 to 18+ months). Toxicity was low. There were only two episodes of drug-related grade 3 nonhematologic toxicity (diarrhea and infection), and only 6%, 3%, and 3% of patients developed granulocytopenia, thrombocytopenia, and neutropenic sepsis, respectively. Mild (grade 1/2) diarrhea occurred in 39% of patients, hand-foot syndrome in 15%, nausea in 27%, and mucositis in 18%. Toxicity-associated delay and dose reduction occurred in only 2% and 5% of courses, respectively. CONCLUSION: First-line treatment with the combination of oral 5-FU and eniluracil has high activity in patients with advanced breast cancer comparable with the most active conventional cytotoxic agents but with strikingly less toxicity. PMID- 10856098 TI - Phase III comparative study of vinorelbine combined with doxorubicin versus doxorubicin alone in disseminated metastatic/recurrent breast cancer: National Cancer Institute of Canada Clinical Trials Group Study MA8. AB - PURPOSE: This phase III study was performed to determine the superiority of doxorubicin (DOX) and vinorelbine (VNB) (arm 1) versus DOX alone (arm 2) in metastatic breast cancer (MBC) for overall survival (OS), time to treatment failure (TTF), toxicity, and quality of life (QOL). PATIENTS AND METHODS: Three hundred three patients were randomized to DOX 50 mg/m(2) intravenously (IV) on day 1 and VNB 25 mg/m(2) IV on days 1 and 8 (arm 1) or DOX 70 mg/m(2) IV on day 1 (arm 2). Both regimens were given every 3 weeks until a cumulative DOX dose of 450 mg/m(2). After 16 of the first 65 randomized patients experienced febrile neutropenia (FN), the doses were reduced to DOX 40 mg/m(2) on day 1 and VNB 20 mg/m(2) on days 1 and 8 versus DOX 60 mg/m(2) on day 1. Eligible patients were vinca alkaloid and anthracycline naive. Chemotherapy was first-line or second line for MBC. RESULTS: Three patients were ineligible. Thus, 300 patients were assessable for toxicity and to determine time to disease progression (TTP), TTF, and OS. Two hundred eighty-nine patients were assessable for response, and 99 responders were assessable for response duration (RD). The response rates, QOL, and median RD, TTP, and TTF were not significantly different between the arms. Median OS was 13.8 months for arm 1 versus 14.4 months for arm 2 (P =.4). Grade 3 or 4 granulocytopenia was equivalent in both arms but more grade 3/4 neurotoxicity, mild venous toxicity, and FN were seen on arm 1. CONCLUSION: The survival with DOX and VNB is not superior to DOX alone in MBC. PMID- 10856099 TI - Palliative effect of chemotherapy: objective tumor response is associated with symptom improvement in patients with metastatic breast cancer. AB - PURPOSE: Because one of the goals of chemotherapy for metastatic breast cancer is to provide symptom palliation, we were interested in identifying the relationship between tumor shrinkage and improvement in disease-related symptoms. PATIENTS AND METHODS: Three hundred patients enrolled onto a randomized trial of metastatic breast cancer formed the basis of our study. The nine most common baseline symptoms were identified and followed. Changes from baseline (improvement, stable, worsening) were defined using patient responses to a quality-of-life (QoL) questionnaire (the European Organization for Research and Treatment of Cancer EORTC QLQ-C30) as well as using graded toxicity data collected on case report forms (CRFs). The association between symptom improvement and tumor response was assessed using a linear trend test via a logistic regression model. RESULTS: The most commonly reported baseline symptoms were cancer pain in 38% (CRF data) and 81% of patients (QoL data) and tiredness in 26% (CRF data) and 89% (QoL data) of patients. Three symptoms-cancer pain, shortness of breath, and abnormal mood-showed a significant relationship between improvement and objective response, using both CRF and QoL assessments. Constipation, anorexia, and nausea showed a similar trend when QoL data were used but not when CRF information was used. The converse was seen for lethargy. There was no correlation between symptom change and response for cough and insomnia. CONCLUSION: For some symptoms, we found a significant association between symptom improvement and objective tumor regression. In these cases, symptom improvement was greatest in those patients who had complete or partial responses, followed by those with stable disease and then those with progressive disease. Further work in this area will be useful in determining the surrogate value of objective tumor response in identifying the efficacy of palliative chemotherapy. PMID- 10856100 TI - Second malignancies after treatment of early-stage breast cancer: lumpectomy and radiation therapy versus mastectomy. AB - PURPOSE: To determine the risk of second malignancies after lumpectomy and radiation therapy (LRT), and to compare it with that in a similar cohort of early stage breast cancer patients undergoing mastectomy without radiation (MAST). PATIENTS AND METHODS: Between January 1970 and December 1990, 1,029 breast cancer patients at our institution underwent LRT. A cohort of 1,387 breast cancer patients who underwent surgical treatment by mastectomy (MAST), and who did not receive postoperative radiation during the same time period, served as a comparison group. Second malignancies were categorized as contralateral breast versus nonbreast. In the cohort of patients undergoing LRT, a detailed analysis was carried out with respect to age, disease stage, smoking history, radiation therapy technique, dose, the use of chemotherapy or hormone therapy, and other clinical and/or pathologic characteristics. RESULTS: As of March 1999, the median follow-up was 14.6 years for the LRT group and 16 years for the MAST group. The 15-year risk of any second malignancy was nearly identical for both cohorts (17.5% v 19%, respectively). The second breast malignancy rate at 15 years was 10% for both the MAST and LRT groups. The 15-year risk of a second nonbreast malignancy was 11% for the LRT and 10% for the MAST group. In the subset of patients 45 years of age or younger at the time of treatment, the second breast and nonbreast malignancy rates at 15 years were 10% and 5% for patients undergoing LRT versus 7% and 4% for patients undergoing mastectomy (P, not statistically significant). In the detailed analysis of LRT patients, second lung malignancies were associated with a history of tobacco use. There were fewer contralateral breast tumors in patients undergoing adjuvant hormone therapy, although this did not reach statistical significance. The adjuvant use of chemotherapy did not significantly affect the risk of second malignancies. CONCLUSION: There seems to be no increased risk of second malignancies in patients undergoing LRT using modern techniques, compared with MAST. Continued monitoring of these patient cohorts will be required in order to document that these findings are maintained with even longer follow-up periods. With nearly 15 years median follow-up periods, however, these data should be reassuring to women who are considering LRT as a treatment option. PMID- 10856101 TI - Paclitaxel, ifosfamide, and cisplatin second-line therapy for patients with relapsed testicular germ cell cancer. AB - PURPOSE: To evaluate the dose, toxicity, and efficacy of paclitaxel in combination with ifosfamide and cisplatin as salvage therapy for patients with relapsed testicular germ cell tumors (GCTs). PATIENTS AND METHODS: Thirty patients with previously treated GCTs were treated with paclitaxel and ifosfamide plus cisplatin (TIP) as second-line therapy. All had favorable prognostic features for response (testis primary tumor site and prior complete response to first-line chemotherapy program). Four cycles of paclitaxel, ifosfamide 5 g/m(2), and cisplatin 100 mg/m(2) were given 21 days apart with granulocyte colony stimulating factor support, followed by resection of radiographic residua. The dose of paclitaxel was increased among cohorts with dose levels of 175, 215, and 250 mg/m(2); the largest dose was selected for the phase II part of the trial. RESULTS: Twenty-three (77%) of 30 patients achieved a complete response to chemotherapy alone, and one patient achieved a durable partial response with normal tumor markers. Therefore, 24 (80%) achieved a favorable response. Eleven patients with normalized markers after chemotherapy underwent resection of residual tissue, with only necrosis found in 10 and mature teratoma in one. Two patients relapsed, and 22 (73%) of the favorable responses remain durable at a median follow-up duration of 33 months. Myelosuppression was the major toxicity, and two patients had grade 3 neurotoxicity. CONCLUSION: Four cycles of TIP was associated with a high proportion of patients who achieved a complete response, a lack of relapse, and relative tolerability as an ifosfamide-containing salvage regimen for testicular GCTs. The high durable complete response proportion emphasizes the importance of patient selection according to prognostic factors for a favorable outcome to conventional-dose salvage therapy. PMID- 10856102 TI - Phase II trial of weekly intravenous gemcitabine with continuous infusion fluorouracil in patients with metastatic renal cell cancer. AB - PURPOSE: To determine the clinical response rate of the combination of weekly intravenous (IV) gemcitabine with continuous infusion fluorouracil (5-FU) in patients with metastatic renal cell carcinoma (RCC). PATIENTS AND METHODS: Between June 1998 and February 1999, 41 patients with metastatic RCC were enrolled onto this multi-institutional phase II study of gemcitabine 600 mg/m(2) over 30 minutes on days 1, 8, and 15 and 5-FU 150 mg/m(2)/d via continuous IV infusion through a permanent catheter on days 1 to 21 of a 28-day cycle. Patients had a Cancer and Leukemia Group B performance status of 0 or 1, with a median time since diagnosis of metastatic disease of 10 months (range, 0 to 129 months). Thirty-three patients (80%) had multiple metastatic sites, and 34 patients (83%) had prior chemotherapy or immunotherapy. RESULTS: Of the 39 assessable patients, there were no complete responses but seven partial responses (objective response rate = 17%; 95% confidence interval, 8% to 34%). Five minor responses (25% to 50% decreased tumor size) were also observed. The duration of response for the seven partial responders was 2, 3, 7, 8, 10, 11, and 14 months. Median progression-free survival for the gemcitabine/5-FU group was 28.7 weeks versus 8 weeks for a similar cohort of patients treated on previous phase II studies at the University of Chicago (P =.008). The regimen was well tolerated, with fatigue, mucositis, nausea/vomiting, and grade 2 hematologic toxicities being most common. CONCLUSION: Weekly gemcitabine with continuous infusion 5-FU is an active combination in patients with metastatic RCC. Therapy was well tolerated and produced an improvement in progression-free survival over historical controls. PMID- 10856103 TI - Benefit of intensified therapy for patients with local or regional embryonal rhabdomyosarcoma: results from the Intergroup Rhabdomyosarcoma Study IV. AB - PURPOSE: To compare failure-free survival (FFS) and survival for patients with local or regional embryonal rhabdomyosarcoma treated on the Intergroup Rhabdomyosarcoma Study (IRS)-IV with that of comparable patients treated on IRS III. PATIENTS AND METHODS: Patients were retrospectively classified as low- or intermediate-risk. Low-risk patients were defined as those with primary tumors at favorable sites, completely resected or microscopic residual, or orbit/eyelid primaries with gross residual disease and tumors less than 5 cm at unfavorable sites but completely resected. Intermediate-risk patients were all other patients with local or regional tumors. RESULTS: Three-year FFS improved from 72% on IRS III to 78% on IRS-IV for patients with intermediate-risk embryonal rhabdomyosarcoma (P =.02). Subset analysis revealed two groups that benefited most from IRS-IV therapy. FFS at 3 years for patients with resectable node positive or unresectable (group III) embryonal rhabdomyosarcoma arising at certain favorable sites (head and neck [not orbit/eyelid or parameningeal] and genitourinary [not bladder or prostate]) improved from 72% on IRS-III to 92% on IRS-IV (P =.01). Similarly, 3-year FFS for patients with completely resected tumor or with only microscopic disease remaining (group I or II) at unfavorable sites improved from 71% on IRS-III to 86% on IRS-IV (P =.04). Only patients with unresectable embryonal rhabdomyosarcoma (group III) at unfavorable sites had no improvement in outcome on IRS-IV (3-year FFS for IRS-III and IRS-IV, 72% and 75%, respectively; P =.31). CONCLUSION: IRS-IV therapy benefited certain subgroups of patients with intermediate-risk embryonal rhabdomyosarcoma. A doubling of the intensity of cyclophosphamide (or ifosfamide equivalent) dosing per cycle between IRS-III and IRS-IV is thought to be a key contributing factor for this improvement. PMID- 10856104 TI - Second cancers among long-term survivors of Hodgkin's disease diagnosed in childhood and adolescence. AB - PURPOSE: To quantify the risk of second cancers among long-term survivors of Hodgkin's disease (HD) diagnosed before 21 years of age and to explore sex-, age , and site-related differences. PATIENTS AND METHODS: We analyzed data from 5,925 pediatric HD patients, including 2,646 10-year and 755 20-year survivors, who were reported to 16 population-based cancer registries in North America and Europe between 1935 and 1994. RESULTS: A total of 157 solid tumors (observed/expected ratio [O/E] = 7.0; 95% confidence interval [CI], 5.9 to 8.2.) and 26 acute leukemias (O/E = 27.4; 95% CI, 17.9 to 40. 2) were reported. Risk of solid tumors remained significantly increased among 20-year survivors (O/E = 6.6, observed [O] = 40, cumulative risk = 6.5%) and persisted for 25 years (O/E = 4.6, O = 15, cumulative risk = 11.7%). Temporal trends for cancers of thyroid, female breast, bone/connective tissue, stomach, and esophagus were consistent with the late effects of radiotherapy. Greater than 50-fold increased risks were observed for tumors of the thyroid and respiratory tract (one lung and one pleura) among children treated before age 10. At older ages (10 to 16 years), the largest number of second cancers occurred in the digestive tract (O/E = 19.3) and breast (O/E = 22.9). Risk of solid tumors increased with decreasing age at HD on a relative but not absolute scale. CONCLUSION: Children and adolescents treated for HD experience significantly increased risks of second cancers at various sites for 2 to 3 decades. Although our results reflect the late effects of past therapeutic modalities, they underscore the importance of lifelong follow-up of pediatric HD patients given early, more aggressive treatments. PMID- 10856105 TI - High- and low-dose interferon alfa-2b in high-risk melanoma: first analysis of intergroup trial E1690/S9111/C9190. AB - PURPOSE: Pivotal trial E1684 of adjuvant high-dose interferon alfa-2b (IFNalpha2b) therapy in high-risk melanoma patients demonstrated a significant relapse-free and overall survival (RFS and OS) benefit compared with observation (Obs). PATIENTS AND METHODS: A prospective, randomized, three-arm, intergroup trial evaluated the efficacy of high-dose IFNalpha2b (HDI) for 1 year and low dose IFNalpha2b (LDI) for 2 years versus Obs in high-risk (stage IIB and III) melanoma with RFS and OS end points. RESULTS: A total of 642 patients were enrolled (608 patients eligible), of whom a majority (75%) had nodal metastasis (50% had nodal recurrence). Unlike E1684, E1690 allowed entry of patients with T4 (> 4 mm) deep primary tumors, regardless of nodal dissection, and 25% of the patients entered onto this trial had deep primary tumors (compared with 11% in E1684). At 52 months' median follow-up, HDI demonstrated an RFS benefit exceeding that of LDI compared with Obs. The 5-year estimated RFS rates for the HDI, LDI, and Obs arms were 44%, 40%, and 35%, respectively. The hazards ratio for the intent-to-treat analysis of HDI versus Obs was 1.28 (P(2) =.05); for LDI versus Obs, it was 1.19 (P(2) =.17). By Cox analysis, the impact of HDI on RFS achieved significance (P(2) =.03). The RFS benefit was equivalent for node-negative and node-positive patients. Neither HDI nor LDI has demonstrated an OS benefit compared with Obs at this time. A major improvement in the median OS of patients in the E1690 Obs arm was noted in comparison with E1684 (6 years v 2.8 years). An analysis of salvage therapy for patients who relapsed on E1690 demonstrated that a significantly larger proportion of patients in the Obs arm received IFNalpha containing salvage therapy compared with the HDI arm; this therapy was unavailable to patients during E1684, and patients with undissected regional nodes were not included in E1684. This study did not specify therapy at recurrence. Analysis of treatments received at recurrence demonstrated significantly more frequent use of IFNalpha2b at relapse from Obs than from HDI, which may have confounded interpretation of the survival benefit of assigned treatments in E1690. CONCLUSION: The results of the intergroup E1690 trial demonstrate an RFS benefit of IFNalpha2b that is dose-dependent and significant for HDI by Cox multivariable analysis. PMID- 10856106 TI - Population pharmacokinetic model for topotecan derived from phase I clinical trials. AB - PURPOSE: To characterize the pharmacokinetics of topotecan in a population model that would identify patient variables or covariates that appreciably impacted on its disposition. PATIENTS AND METHODS: All data were collected from 82 patients entered in four different phase I trials that were previously reported as separate studies from 1992 to 1996. All patients received topotecan as a 30 minute constant-rate infusion on a daily-times-five schedule and were selected for this study because their daily dose did not exceed 2.0 mg/m(2). Among the 82 patients were 30 patients classified as having renal insufficiency and 13 patients with hepatic dysfunction. The population pharmacokinetic model was built in sequential manner, starting with a covariate-free model and progressing to a covariate model with the aid of generalized additive modeling. RESULTS: A linear two-compartment model characterized total topotecan plasma concentrations (n = 899). Four primary pharmacokinetic parameters (total clearance, volume of the central compartment, distributional clearance, and volume of the peripheral compartment) were related to various combinations of covariates. The relationship for total clearance (TVCL [L/h] = 32.0 + [0.356(WT - 71) + 0.308(HT - 168.5) - 8.42(SCR - 1.1)] x [1 + 0.671 sex]) was dependent on the patients' weight (WT), height (HT), serum creatinine (SCR), and sex and had a moderate ability to predict (r(2) = 0.64) each patient's individual clearance value. The addition of covariates to the population model improved the prediction errors, particularly for clearance. Removal of 10 outlying patients from the analysis improved the ability of the model to predict individual clearance values (r(2) = 0.77). CONCLUSION: A population pharmacokinetic model for total topotecan has been developed that incorporates measures of body size and renal function to predict total clearance. The model can be used prospectively to obtain a revised and validated model that can then be used to design individualized dosing regimens. PMID- 10856107 TI - Phase I and pharmacokinetic study of oral paclitaxel. AB - PURPOSE: To investigate dose escalation of oral paclitaxel in combination with dose increment and scheduling of cyclosporine (CsA) to improve the systemic exposure to paclitaxel and to explore the maximum-tolerated dose (MTD) and dose limiting toxicity (DLT). PATIENTS AND METHODS: A total of 53 patients received, on one occasion, oral paclitaxel in combination with CsA, coadministered to enhance the absorption of paclitaxel, and, on another occasion, intravenous paclitaxel at a dose of 175 mg/m(2) as a 3-hour infusion. RESULTS: The main toxicities observed after oral intake of paclitaxel were acute nausea and vomiting, which reached DLT at the dose level of 360 mg/m(2). Dose escalation of oral paclitaxel from 60 to 300 mg/m(2) resulted in significant but less than proportional increases in the plasma area under the concentration-time curve (AUC) of paclitaxel. The mean AUC values +/- SD after 60, 180, and 300 mg/m(2) of oral paclitaxel were 1.65 +/- 0.93, 3.33 +/- 2.39, and 3.46 +/- 1.37 micromol/L.h, respectively. Dose increment and scheduling of CsA did not result in a further increase in the AUC of paclitaxel. The AUC of intravenous paclitaxel was 15.39 +/- 3.26 micromol/L.h. CONCLUSION: The MTD of oral paclitaxel was 300 mg/m(2). However, because the pharmacokinetic data of oral paclitaxel, in particular at the highest doses applied, revealed nonlinear pharmacokinetics with only a moderate further increase of the AUC with doses up to 300 mg/m(2), the oral paclitaxel dose of 180 mg/m(2) in combination with 15 mg/kg oral CsA is considered most appropriate for further investigation. The safety of the oral combination at this dose level was good. PMID- 10856108 TI - Pharmacoeconomic analysis of liposomal amphotericin B versus conventional amphotericin B in the empirical treatment of persistently febrile neutropenic patients. AB - PURPOSE: In a randomized, double-blind, comparative, multicenter trial, liposomal amphotericin B was equivalent to conventional amphotericin B for empirical antifungal therapy in febrile neutropenic patients, using a composite end point, but was more effective in reducing proven emergent fungal infections, infusion related toxicities, and nephrotoxicity. The purpose of this study was to compare the pharmacoeconomics of liposomal versus conventional therapy. PATIENTS AND METHODS: Itemized hospital billing data were collected on 414 patients from 19 of the 32 centers that participated in the trial. Hospital length of stay and costs from the first dose of study medication to the time of hospital discharge were assessed. RESULTS: Hospital costs from the time of first dose to discharge were significantly higher for all patients who received liposomal amphotericin B ($48,962 v $43,183; P =.022). However, hospital costs were highly sensitive to the cost of study medication ($39,648 v $43,048 when drug costs were not included; P =.416). Using decision analysis models and sensitivity analyses to vary the cost of study medications and the risk of nephrotoxicity, the break-even points for the cost of liposomal therapy were calculated to range from $72 to $87 per 50 mg for all patients and $83 to $112 per 50 mg in allogeneic bone marrow transplant patients. CONCLUSION: The cost of liposomal amphotericin B and patient risk for developing nephrotoxicity play large roles in determining whether liposomal amphotericin B is cost-effective as first-line empirical therapy in persistently febrile neutropenic patients. PMID- 10856109 TI - What would you do? Specialists' perspectives on cancer genetic testing, prophylactic surgery, and insurance discrimination. AB - PURPOSE: To examine what cancer genetics specialists predict they would do personally if they were at 50% risk of carrying a mutation that predisposes to hereditary breast/ovarian cancer (BRCA1/BRCA2) and hereditary nonpolyposis colon cancer (HNPCC). METHODS: Questionnaire survey of the membership of the National Society of Genetic Counselors (NSGC) Special Interest Group (SIG) in Cancer. RESULTS: Of the 296 active members of the NSGC Cancer-SIG surveyed, 163 (55%) responded. Eighty-five percent predicted that if they had a 50% risk of carrying a BRCA1/BRCA2 mutation, they would pursue genetic testing. If they tested positive for a mutation at age 35, 25% predicted they would pursue prophylactic bilateral mastectomies and 68%, prophylactic oophorectomy. Ninety-one percent of respondents believe they would pursue genetic testing for HNPCC, and 17% would elect prophylactic colectomy; 54%, prophylactic hysterectomy; and 52%, prophylactic oophorectomy if they tested positive for a mutation. The majority (68%) would not bill their insurance companies for genetic testing because of fear of discrimination, and 26% would use an alias when undergoing testing. Fifty seven percent of counselors would seek professional psychologic support to help them cope with the results of testing. CONCLUSION: A large percentage of cancer genetic counseling providers predicted they would opt for prophylactic surgery at a young age if they carried a BRCA or HNPCC mutation, and most would seek professional psychologic assistance when undergoing testing. More than half of respondents would not bill their insurance companies for genetic testing, largely because of fear of genetic discrimination. The vast majority of those providers most familiar with cancer genetic testing and its associated medical, psychologic, and legal implications would still pursue genetic testing. PMID- 10856110 TI - Chemotherapy in neuroendocrine/Merkel cell carcinoma of the skin: case series and review of 204 cases. AB - PURPOSE: To study the use of chemotherapy for Merkel cell carcinoma (MCC) of the skin. PATIENTS AND METHODS: Twenty-five cases of MCC were treated at the London Regional Cancer Center between 1987 and 1997. Thirteen cases treated with chemotherapy were reviewed with 191 cases from the literature. RESULTS: At presentation, 24 patients had localized skin lesions (stage I) and one had locoregional involvement (stage II). Among the nine cases with recurrent nodal disease, six had chemotherapy as a component of salvage treatment. They were all free of disease at a median of 19 months (range, 12 to 37 months). In contrast, two patients who had salvage radiotherapy alone died of disease. Overall survival (OS) and disease-free survival (DFS) were 59% and 43%, respectively, at two years. Median OS and DFS were 29 months (range, 1 to 133 months) and 9 months (range, 1 to 133 months), respectively. Nodal disease developed in 12 (50%) of 24 patients with stage I disease, and distant metastases developed in six (25%) of 24. Including those from the literature, there were 204 cases treated with chemotherapy. Cyclophosphamide/doxorubicin (or epirubicin)/vincristine combination +/- prednisone was the most commonly used chemotherapy regimen (47 cases), with an overall response rate of 75.7% (35.1% complete, 35. 1% partial, and 5.4% minor responses). Etoposide/cisplatin (or carboplatin) was the next most commonly used regimen (27 cases), with an overall response rate of 60% (36% complete and 24% partial responses). The difference in response rate was not statistically significant (P =.19). Among the 204 cases, there were seven (3.4%) toxic deaths. CONCLUSION: Chemoradiation for locally recurrent or advanced disease may be an option for patients with a good performance status. PMID- 10856111 TI - K(+) channels and their distribution in large cortical pyramidal neurones. PMID- 10856112 TI - Mammalian brainstem chemosensitive neurones: linking them to respiration in vitro. AB - Neurones which are excited by CO2 or H+ are present in a number of brainstem structures in addition to the ventrolateral region of the medulla, the site at which the respiratory response to hypercapnia is traditionally believed to originate. In this review we examine recent work concerned with establishing the relationship between these chemosensitive neurones and respiration, the emphasis being placed on the use for this purpose of in vitro preparations of the mammalian brainstem. PMID- 10856113 TI - Disruption of actin impedes transmitter release in snake motor terminals. AB - To investigate the role of actin in vertebrate nerve terminals, nerve-muscle preparations from garter snake (Thamnophis sirtalis) were treated with the actin depolymerizing agent latrunculin A. Immunostaining revealed that actin filaments within presynaptic motor terminal boutons were disrupted by the drug. In preparations loaded with the optical probe FM1-43, destaining was reduced by latrunculin treatment, suggesting that transmitter release was partially blocked. Latrunculin treatment did not influence the amplitude or time course of spontaneous miniature endplate potentials (MEPPs). Similarly, endplate potentials (EPPs) evoked at low frequency were comparable in control and latrunculin-treated curarized preparations. Brief tetanic stimulation of the muscle nerve (25 Hz, 90 s) depressed EPP amplitudes in both control and latrunculin-treated preparations. After tetanus, EPPs elicited at 0. 2 Hz in control preparations recovered rapidly (0-5 min) and completely (usually potentiating to above pre-tetanus levels; 130 +/- 11 %, mean +/- s.e.m.). In contrast, EPPs evoked in latrunculin-treated preparations recovered slowly (8-10 min) and incompletely (84 +/- 8 %). The influence of latrunculin on post-tetanic EPPs depended on its concentration in the bath (KD = 3. 1 microM) and on time of incubation. These observations argue that actin filaments facilitate transmitter release rather than impede it. Specifically, actin may facilitate mobilization of vesicles towards the releasable pools. PMID- 10856114 TI - In vivo formation of a proton-sensitive K+ channel by heteromeric subunit assembly of Kir5.1 with Kir4.1. AB - Kir5.1 is an inwardly rectifying K+ channel (Kir) subunit, whose physiological function is unknown. Human embryonic kidney HEK293T cells co-transfected with rat Kir5.1 and Kir4.1 cDNA expressed a functional K+ channel, whose properties were significantly different from those of the homomeric Kir4.1 channel. Formation of a Kir4. 1/Kir5.1 assembly in HEK293T was confirmed biochemically. We found that heteromeric Kir4.1/Kir5.1 channel activity was affected by internal pH levels between 6.0 and 8.0, when the homomeric Kir4.1 channel activity was relatively stable. Changing external pH in this range had no effect on either Kir channel. Western blot analysis using specific antibodies revealed that Kir4.1 and Kir5.1 proteins were expressed in kidney and brain, but co-immunoprecipitated only from kidney. These results indicate that the co-assembly of Kir5.1 with Kir4.1 occurs in vivo, at least in kidney. The heteromeric Kir4. 1/Kir5.1 channel may therefore sense intracellular pH in renal epithelium and be involved in the regulation of acid-base homeostasis. PMID- 10856115 TI - Properties of voltage-gated potassium currents in nucleated patches from large layer 5 cortical pyramidal neurons of the rat. AB - Voltage-gated potassium currents were studied in nucleated outside-out patches obtained from large layer 5 pyramidal neurons in acute slices of sensorimotor cortex from 13- to 15-day-old Wistar rats (22-25 C). Two main types of current were found, an A-current (IA) and a delayed rectifier current (IK), which were blocked by 4-aminopyridine (5 mM) and tetraethylammonium (30 mM), respectively. Recovery from inactivation was mono-exponential (for IA) or bi-exponential (for IK) and strongly voltage dependent. Both IA and IK could be almost fully inactivated by depolarising prepulses of sufficient duration. Steady-state inactivation curves were well fitted by the Boltzmann equation with half-maximal voltage (V ) and slope factor (k) values of -81.6 mV and -6.7 mV for IA, and 66.6 mV and -9.2 mV for IK. Peak activation curves were described by the Boltzmann equation with V and k values of -18.8 mV and 16.6 mV for IA, and -9.6 mV and 13.2 mV for IK. IA inactivated mono-exponentially during a depolarising test pulse, with a time constant ( approximately 7 ms) that was weakly dependent on membrane potential. IK inactivated bi-exponentially with time constants ( approximately 460 ms, approximately 4.2 s) that were also weakly voltage dependent. The time to peak of both IA and IK depended strongly on membrane potential. The kinetics of IA and IK were described by a Hodgkin-Huxley-style equation of the form mNh, where N was 3 for IA and 1 for IK. These results provide a basis for understanding the role of voltage-gated potassium currents in the firing properties of large layer 5 pyramidal neurons of the rat neocortex. PMID- 10856116 TI - Distribution and activation of voltage-gated potassium channels in cell-attached and outside-out patches from large layer 5 cortical pyramidal neurons of the rat. AB - Voltage-gated potassium channels were studied in cell-attached and outside-out patches from the soma and primary apical dendrite of large layer 5 pyramidal neurons in acute slices of rat sensorimotor cortex (22-25 degrees C). Ensemble averages revealed that some patches contained only fast, I(A)-like channels, other contained only I(K)-like channels that did not inactivate or inactivated slowly, and the remainder contained mixtures of both types. I(A) and I(K) channels had mean unitary conductances of 8.5 and 20.3 pS, respectively, and had distinctive patterns of gating. Peak activation curves for ensemble-averaged currents were described by the Boltzmann equation with half-maximal voltage [V(1/2)] and slope factor (k) values of -24.5 mV and 16.9 mV for I(A) and -7.6 mV and 10.1 mV for I(K) (patches < 250 microm from the soma) or -22.9 mV and 16.2 mV for I(A) (patches > 250 microm from the soma). The steady-state inactivation curve for I(A) gave V(1/2) and k values of -72.3 mV and -5.9 mV (< 250 microm from the soma) or -83.1 mV and -6.5 mV (> 250 microm from the soma). These values were similar to the corresponding data for I(A) and I(K) in nucleated patches from the same cell. The amount of I(A) and I(K) present in patches depended weakly on distance along the primary apical dendrite from the soma. The amplitude of I(A) increased, on the average, by 2.3 pA per 100 microm, while the amplitude of I(K) decreased by 0.4 pA per 100 microm. I(A) and I(K) channels in dendritic cell-attached patches were activated by the passage of a back-propagating action potential past the tip of the patch electrode. These results show directly that these potassium channels participate in action potential repolarisation, and thus contribute to the process of synaptic integration in these neurons. PMID- 10856117 TI - Voltage-gated K+ channels in layer 5 neocortical pyramidal neurones from young rats: subtypes and gradients. AB - We investigated the types and distribution of voltage-gated K+ channels in the soma and apical dendrite of layer 5 (L5) neocortical pyramidal neurones, of young rats (postnatal days 13-15), in acute brain slices. A slow inactivating outward K+ current and a fast inactivating outward K+ current were detected in nucleated patches. The slow K+ current was completely blocked by tetraethylammonium (TEA) with an IC50 of 5 +/- 1 mM (mean +/- s.e.m.) and was partially blocked by 4 aminopyridine (4-AP). The fast K+ current was blocked by 4-AP with an IC50 of 4.2 +/- 0.5 mM, but was not blocked by TEA. The activation kinetics of the slow K+ current were described by a second order Hodgkin-Huxley model. The slow K+ current displayed bi-exponential inactivation. A fourth order Hodgkin-Huxley model for activation and first order for inactivation described the kinetics of the fast K+ current. In somatic cell-attached recordings, three classes of single K+ channels could be differentiated based on their unitary conductance and inactivation kinetics, a fast inactivating channel having a conductance of 13 +/- 1 pS, a slow inactivating channel having a conductance of 9.5 +/- 0.5 pS, and a very slowly inactivating channel having a conductance of 16 +/- 1 pS. The inactivation time constants of the slow and of the very slow K+ channel corresponded to the two inactivation time constants of the slow K+ current observed in nucleated patches. This suggested that two distinct K+ channels mediated the slow K+ current in nucleated patches. The three subtypes of K+ channels that were observed in somatic recordings were present along the apical dendrite. The amplitude of ensemble K+ currents in cell-attached patches decreased along the apical dendrite as the distance from the soma increased, with a slope of -0.9 +/- 0.3 pA per 100 microm. The results suggest that the decrease of the voltage-gated K+ channel density from the soma along the apical dendrite of L5 pyramidal neurones helps to define a distal, low threshold region for the initiation of dendritic regenerative potentials. PMID- 10856118 TI - Gentamicin blocks ACh-evoked K+ current in guinea-pig outer hair cells by impairing Ca2+ entry at the cholinergic receptor. AB - Aminoglycoside antibiotics such as gentamicin are known to block the medial olivocochlear efferent system. In order to determine whether this inhibition takes place at the postsynaptic cholinergic receptors in outer hair cells (OHCs), we studied the effects of these polycationic molecules on cholinergic currents evoked in isolated guinea-pig OHCs. The cholinergic response of OHCs involves nicotinic-like receptors (nAChRs) permeable to Ca2+ ions that activate nearby Ca2+-sensitive K+ channels (KCa(ACh) channels). The extracellular application of gentamicin and neomycin reversibly blocked ACh-evoked K+ current (IK(ACh)) with IC50 values of 5.5 and 3.2 microM, respectively. The results showed that the blocking mechanism of IK(ACh) was due to inhibition of Ca2+ influx via nAChRs. Our study also provides interesting insights into the functional coupling between nAChRs and KCa(ACh) channels in OHCs. By directly recording the cation current flowing through nAChRs (In(ACh)) using an intracellular solution containing 10 mM BAPTA, we measured an EC50 near 110 microM for ACh-evoked In(ACh). This EC50 for ACh is one order of magnitude higher than that measured indirectly on IK(ACh). This reveals a rather low affinity of ACh for its receptor but a very efficient coupling between nAChRs and KCa(ACh) channels. We also show that a high external Ca2+ concentration reverts the gentamicin inhibition of IK(ACh) and that gentamicin directly alters the cation current flowing through the nAChRs of OHCs. We propose that gentamicin acts as a non-competitive cholinergic blocker by displacing Ca2+ from specific binding sites at the nAChRs. This block of the nAChRs at the level of the postsynaptic membrane in OHCs could explain the inhibitory effect of gentamicin reported on the crossed medial olivocochlear efferent system in vivo. PMID- 10856119 TI - Adenosine A1 receptors modulate high voltage-activated Ca2+ currents and motor pattern generation in the xenopus embryo. AB - Adenosine causes voltage- and non-voltage-dependent inhibition of high voltage activated (HVA) Ca2+ currents in Xenopus laevis embryo spinal neurons. As this inhibition can be blocked by 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX) and mimicked by N6-cyclopentyladenosine (CPA) it appears to be mediated by A1 receptors. Agents active at A2 receptors either were without effect or could be blocked by DPCPX. AMP had no agonist action on these receptors. By using omega conotoxin GVIA we found that adenosine inhibited an N-type Ca2+ current as well as a further unidentified HVA current that was insensitive to dihydropyridines, omega-agatoxin TK and omega-conotoxin MVIIC. Both types of current were subject to voltage- and non-voltage-dependent inhibition. We used CPA and DPCPX to test whether A1 receptors regulated spinal motor pattern generation in spinalized Xenopus embryos. DPCPX caused a near doubling of, while CPA greatly shortened, the length of swimming episodes. In addition, DPCPX slowed, while CPA greatly speeded up, the rate of run-down of motor activity. Our results demonstrate a novel action of A1 receptors in modulating spinal motor activity. Furthermore they confirm that adenosine is produced continually throughout swimming episodes and acts to cause the eventual termination of activity. PMID- 10856120 TI - Voltage-independent calcium entry in hypoxic pulmonary vasoconstriction of intrapulmonary arteries of the rat. AB - It has been proposed that hypoxic pulmonary vasoconstriction (HPV) is mediated via K+ channel inhibition and Ca2+ influx through voltage-gated channels. HPV depends strongly on the degree of preconstriction, and we therefore examined the effect of Ca2+ channel blockade on tension and intracellular [Ca2+] ([Ca2+]i) during HPV in rat intrapulmonary arteries (IPAs), whilst maintaining preconstriction constant. We also investigated the role of intracellular Ca2+ stores. HPV demonstrated a transient constriction (phase I) superimposed on a sustained constriction (phase II). Nifedipine (1 microM) partially inhibited phase I, but did not affect phase II. In arteries exposed to 80 mM K+ and nifedipine or diltiazem the rises in tension and [Ca2+]i were blunted during phase I, but were unaffected during phase II. At low concentrations (< 3 microM), La3+ almost abolished the phase I constriction and rise in [Ca2+]i, but had no effect on phase II, or constriction in response to 80 mM K+. Phase II was inhibited by higher concentrations of La3+ (IC50 approximately 50 microM). IPA treated with thapsigargin (1 microM) in Ca2+-free solution to deplete Ca2+ stores showed sustained constriction upon re-exposure to Ca2+ and an increase in the rate of Mn2+ influx, suggesting capacitative Ca2+ entry. The concentration dependency of the block of constriction by La3+ was similar to that for phase I of HPV. Pretreatment of IPA with 30 microM CPA reduced phase I by > 80 %, but had no significant effect on phase II. We conclude that depolarization-mediated Ca2+ influx plays at best a minor role in the transient phase I constriction of HPV, and is not involved in the sustained phase II constriction. Instead, phase I appears to be mainly dependent on capacitative Ca2+ entry related to release of thapsigargin-sensitive Ca2+ stores, whereas phase II is supported by Ca2+ entry via a separate voltage-independent pathway. PMID- 10856121 TI - Effects of the immunosuppressant FK506 on intracellular Ca2+ release and Ca2+ accumulation mechanisms. AB - The immunophilin FKBP12 associates with intracellular Ca2+ channels and this interaction can be disrupted by the immunosuppressant FK506. We have investigated the effect of FK506 on Ca2+ release and Ca2+ uptake in permeabilized cell types. Changes in medium free [Ca2+] were detected by the fluorescent Ca2+ indicator fluo-3 in digitonin-permeabilized SH-SY5Y human neuroblastoma cells, DT40 and R23 11 (i.e. triple inositol 1,4,5-trisphosphate (IP3) receptor knockout cells) chicken B lymphocytes and differentiated and undifferentiated BC3H1 skeletal muscle cells. 45Ca2+ fluxes were studied in saponin-permeabilized A7r5 rat smooth muscle cells. Addition of FK506 to permeabilized SH-SY5Y cells led to a sustained elevation of the medium [Ca2+] corresponding to approximately 30 % of the Ca2+ ionophore A23187-induced [Ca2+] rise. This rise in [Ca2+] was not dependent on mitochondrial activity. This FK506-induced [Ca2+] rise was related to the inhibition of the sarcoplasmic/endoplasmic reticulum Ca2+-Mg2+-ATPase (SERCA) Ca2+ pump. Oxalate-facilitated 45Ca2+ uptake in SH-SY5Y microsomes was inhibited by FK506 with an IC50 of 19 microM. The inhibition of the SERCA Ca2+ pump was not specific since several macrocyclic lactone compounds (ivermectin > FK506, ascomycin and rapamycin) were able to inhibit Ca2+ uptake activity. FK506 (10 microM) did not affect IP3-induced Ca2+ release in permeabilized SH-SY5Y and A7r5 cells, but enhanced caffeine-induced Ca2+ release via the ryanodine receptor (RyR) in differentiated BC3H1 cells. In conclusion, FK506 inhibited active Ca2+ uptake by the SERCA Ca2+ pump; in addition, FK506 enhanced intracellular Ca2+ release through the RyR, but it had no direct effect on IP3-induced Ca2+ release. PMID- 10856122 TI - The mechanisms of sarcoplasmic reticulum Ca2+ release in toad pacemaker cells. AB - The mechanisms of sarcoplasmic reticulum (SR) Ca2+ release in pacemaker cells from the sinus venosus of the cane toad (Bufo marinus) were studied. Single, isolated cells were voltage clamped using a nystatin-perforated patch. Ionic currents and intracellular Ca2+ concentration ([Ca2+]i) were recorded simultaneously. Depolarizations of 300 ms duration from a holding potential of 55 mV produced an inward current which had a bell-shaped relationship with voltage. Inward current first appeared at about -45 mV, reached a maximum of -343 +/- 46 pA at -15 mV and reversed at +45 mV. In contrast the amplitude of the increase in [Ca2+]i caused by depolarization (Ca2+ transient) increased monotonically with the increasing depolarization. At -15 mV the amplitude of the Ca2+ transient was 243 +/- 33 nM and at +45 mV it was 411 +/- 43 nM. The inward current produced by depolarizations to -5 mV was largely eliminated by the L-type Ca2+ channel blocker nifedipine (10 microM) while 37 +/- 7 % of the Ca2+ transient persisted. A significantly larger proportion of the Ca2+ transient (56 +/- 5 %) remained at +85 mV in the presence of nifedipine. The SR Ca2+ pump inhibitor 2, 5-di(tert-butyl)-1,4-hydroquinone (10 microM), which causes depletion of the SR Ca2+, reduced the amplitude of the Ca2+ transient to 34 +/- 1 % of control, irrespective of the voltage. Brief exposure to extracellular Ca2+ free solution abolished the Ca2+ transients caused by depolarization while the caffeine-induced Ca2+ release persisted. Tetrodotoxin (1 microM) had no effect on the amplitude of the depolarization-induced Ca2+ transient, although it reduced the fast component of the inward current. In contrast, Ni2+ (5 mM) abolished the Ca2+ transients at any given voltage. Ni2+ also abolished spontaneous Ca2+ transients. In conclusion, in toad pacemaker cells Ca2+ release from SR contributes approximately 66 % of the Ca2+ involved in the Ca2+ transient and requires extracellular Ca2+ influx to trigger its release. The L-type Ca2+ channels and Na+-Ca2+ exchange are major sources of Ca2+ influx under physiological conditions. PMID- 10856123 TI - Control of InsP3-induced Ca2+ oscillations in permeabilized blowfly salivary gland cells: contribution of mitochondria. AB - Many agonists linked to the generation of inositol 1,4, 5-trisphosphate (InsP3) and release of Ca2+ from intracellular stores induce repetitive transients in cytosolic Ca2+ whose frequency increases over a certain range of agonist concentrations. In order to investigate the mechanisms underlying this frequency modulation, the fluorescent Ca2+ sensor mag-fura-2 was loaded into intracellular calcium stores and used to monitor InsP3-induced dynamics of the intraluminal calcium concentration ([Ca2+]L) in secretory cells of permeabilized blowfly Calliphora vicina salivary glands. In this preparation, increasing concentrations of InsP3 induced graded decreases in [Ca2+]L that were often superimposed with repetitive [Ca2+]L transients produced by sequential Ca2+ release and re-uptake. These [Ca2+]L oscillations developed at frequencies of 3-11 min-1 unrelated to the concentration of InsP3 present. In contrast, incremental concentrations of InsP3 applied in the presence of the oxidizable mitochondrial substrates citrate, succinate, or pyruvate-malate induced repetitive [Ca2+]L transients whose frequency increased with the concentration of InsP3. This InsP3 concentration dependent modulation of oscillation frequency was abolished after dissipating the mitochondrial membrane potential (Delta psi m) by combined treatment with carbonyl cyanide p-trifluoromethoxyphenyl hydrazone + oligomycin or after application of Ruthenium Red, an inhibitor of mitochondrial Ca2+ uptake. Taken together, the data indicate that energized mitochondria exert negative control over the frequency of InsP3-induced Ca2+ oscillations. It is concluded that mitochondria play a crucial role in determining the duration of the interspike period and, therefore, for the encoding of amplitude-modulated, InsP3-liberating stimuli into the frequency of cytosolic Ca2+ oscillations. PMID- 10856124 TI - Activation of volume-regulated Cl(-) channels by ACh and ATP in Xenopus follicles. AB - Osmolarity-dependent ionic currents from follicle-enclosed Xenopus oocytes (follicles) were studied using electrophysiological techniques. Whole follicle currents were monitored using a two-electrode voltage clamp and single-channel activity was measured using the patch-clamp technique. In follicles held at -60 mV two chloride currents were activated in external hyposmotic solutions. One was the habitual volume-regulated current elicited by external hyposmolarity (ICl,swell), and the second was a slow and smooth current (Sin) generated by ACh or ATP application. In follicles, the permeability ratios for different anions with respect to Cl- were similar for both ICl,swell and Sin, with a sequence of: SCN- > I- > Br- >= NO3- >= Cl- > gluconate >= cyclamate > acetate > SO42-. Extracellular ATP blocked the outward component of Sin. Also, extracellular pH modulated the inactivation kinetics of Sin elicited by ACh; e.g. inactivation at +80 mV was approximately 100 % slower at pH 8.0 compared with that at pH 6.0. Lanthanides inhibited ICl, swell and Sin. La3+ completely inhibited ICl,swell with a half-maximal inhibitory concentration (IC50) of 17 +/- 1.9 microM, while Sin was blocked up to 55 % with an apparent IC50 of 36 +/- 2.6 microM. Patch clamp recordings in follicular cells showed that hyposmotic challenge opened inward single-channel currents. The single channel conductance (4.7 +/- 0.4 pS) had a linear current-voltage relationship with a reversal membrane potential close to -20 mV. This single-channel activity was increased by application of ACh or ATP. The ICl,swell generation was not affected by pirenzepine or metoctramine, and did not affect the purinergic activation of the chloride current named Fin. Thus, ICl,swell was not generated via neurotransmitters released during cellular swelling. All together, equal discrimination for different anions, similar modulatory effects by extracellular pH, the blocking effects by ATP and La3+, and the same single-channel activity, strongly suggest that ICl,swell and Sin currents depend on the opening of the same type or a closely related class of volume-regulated chloride channels. PMID- 10856125 TI - Two distinct classes of functional 7-containing nicotinic receptor on rat superior cervical ganglion neurons. AB - Nicotinic acetylcholine receptors (nAChRs) that bind alpha-bungarotoxin (alpha Bgt) were studied on isolated rat superior cervical ganglion (SCG) neurons using whole-cell patch clamp recording techniques. Rapid application of ACh onto the soma of voltage clamped neurons evoked a slowly desensitizing current that was reversibly blocked by alpha Bgt (50 nM). The toxin-sensitive current constituted on average about half of the peak whole-cell response evoked by ACh. Nanomolar concentrations of methyllycaconitine blocked the alpha Bgt-sensitive component of the ACh-evoked current as did intracellular dialysis with an anti-alpha 7 monoclonal antibody. The results indicate that the slowly reversible toxin sensitive response elicited by ACh arises from activation of an unusual class of alpha 7-containing receptor (alpha 7-nAChR) similar to that reported previously for rat intracardiac ganglion neurons. A second class of functional alpha 7-nAChR was identified on some SCG neurons by using rapid application of choline to elicit responses. In these cases a biphasic response was obtained, which included a rapidly desensitizing component that was blocked by alpha Bgt in a pseudo irreversible manner. The pharmacology and kinetics of the responses resembled those previously attributed to alpha 7-nAChRs in a number of other neuronal cell types. Experiments measuring the dissociation rate of 125I-labelled alpha Bgt from SCG neurons revealed two classes of toxin-binding site. The times for toxin dissociation were consistent with those required to reverse blockade of the two kinds of alpha Bgt-sensitive response. These results indicate that rat SCG neurons express two types of functional alpha 7-nAChR, differing in pharmacology, desensitization and reversibility of alpha Bgt blockade. PMID- 10856126 TI - Voltage- and time-dependent properties of the recombinant rat vanilloid receptor (rVR1). AB - Whole-cell voltage-clamp techniques were used to investigate the capsaicin-, voltage- and time-dependent properties of the rat vanilloid receptor (rVR1) stably expressed in human embryonic kidney (HEK) 293 cells. At a holding potential of -70 mV, application of capsaicin (0.03-30 microM) to HEK 293 cells expressing the rVR1 receptor led to the appearance of inward currents (EC50, 497 nM; Hill coefficient, nH, 2.85) which were reversibly antagonized by 10 microM capsazepine. Current-voltage relationships, determined using depolarizing or hyperpolarizing voltage ramps, had reversal potentials close to 0 mV, exhibited substantial outward rectification and possessed a region of negative slope conductance at holding potentials negative to around -70 mV. Further experiments indicated that the outward rectification and the region of negative slope conductance did not result from external block of the channel by either Ba2+, Ca2+ or Mg2+. During our characterization of rVR1, it became apparent that the rectification behaviour of this receptor was not entirely instantaneous as might be expected for a ligand-gated ion channel, but rather displayed clear time dependent components. We characterized the kinetics of these novel gating properties in a series of additional voltage-step experiments. The time-dependent changes in rVR1-mediated conductance due to membrane depolarization or repolarization occurred with bi-exponential kinetics. On depolarization to +70 mV the time-dependent increase in outward current developed with mean time constants of 6.7 +/- 0.7 and 51.8 +/- 18.4 ms, with the faster time constant playing a dominant role (64.4 +/- 3.8 %). Similar kinetics also described the decay of 'tail currents' observed on repolarization. Furthermore, these time-dependent changes appeared to be unaffected by the removal of extracellular divalent cations and were not significantly voltage dependent. Our data reveal that rVR1 exhibits substantial time- and voltage-dependent gating properties that may have significance for the physiology of sensory transduction of nociceptive signals. PMID- 10856127 TI - Regulation of the endothelin system by shear stress in human endothelial cells. AB - In this study, the effect of shear stress on the expression of genes of the human endothelin-1 system was examined. Primary cultures of human umbilical vein endothelial cells (HUVEC) were exposed to laminar shear stress of 1, 15 or 30 dyn cm-2 (i.e. 0.1, 1.5 or 3 N m-2) (venous and two different arterial levels of shear stress) in a cone-and-plate viscometer. Laminar shear stress transiently upregulates preproendothelin-1 (ppET-1) mRNA, reaching its maximum after 30 min (approx 1.7-fold increase). In contrast, long-term application of shear stress (24 h) causes downregulation of ppET-1 mRNA in a dose-dependent manner. Arterial levels of shear stress result in downregulation of endothelin-converting enzyme-1 isoform ECE-1a (predominating in HUVEC) to 36.2 +/- 8.5 %, and isoform ECE-1b mRNA to 72.3 +/- 1.9 % of static control level. The endothelin-1 (ET-1) release is downregulated by laminar shear stress in a dose-dependent manner. This downregulation of ppET-1 mRNA and ET-1 release is not affected by inhibition of protein kinase C (PKC), or tyrosine kinase. Inhibition of endothelial NO synthase (L-NAME, 500 microm) prevents downregulation of ppET-1 mRNA by shear stress. In contrast, increasing degrees of long-term shear stress upregulate endothelin receptor type B (ETB) mRNA by a NO- and PKC-, but not tyrosine kinase-dependent mechanism. In conclusion, our data suggest the downregulation of human endothelin synthesis, and an upregulation of the ETB receptor by long-term arterial laminar shear stress. These effects might contribute to the vasoprotective and anti arteriosclerotic potential of arterial laminar shear stress. PMID- 10856128 TI - Role of aquaporins in alveolar fluid clearance in neonatal and adult lung, and in oedema formation following acute lung injury: studies in transgenic aquaporin null mice. AB - Aquaporin (AQP) water channels provide a major pathway for osmotically driven water movement across epithelial and microvascular barriers in the lung. We used mice deficient in each of the three principal lung aquaporins, AQP1, AQP4 and AQP5, to test the hypothesis that aquaporins are important in neonatal lung fluid balance, adult lung fluid clearance and formation of lung oedema after acute lung injury. Wet-to-dry weight ratios (W/D) in lungs from wild-type mice decreased from 7.9 to 5.7 over the first hour after spontaneous delivery. AQP deletion did not significantly affect W/D at 45 min after birth. Alveolar fluid clearance was measured in living ventilated mice in which 0.5 ml saline containing radiolabelled albumin was instilled into the airspaces. Fluid clearance was 17.4 % in 15 min and inhibited >90 % by amiloride, but clearance was not affected by AQP deletion. W/D was measured in established models of acute lung injury - acid aspiration and thiourea administration. Two hours after intratracheal administration of HCl, W/D increased from 3.7 to 7.5 but was not affected by AQP deletion. Three hours after intraperitoneal infusion of thiourea, W/D increased to 5.5 and marked pleural effusions appeared, but there were no differences in wild-type and AQP knockout mice. Hyperoxic subacute lung injury was induced by 95 % oxygen. Neither mean survival (143 h) nor W/D at 65 h (5.1) were significantly affected by AQP deletion. Despite their role in osmotically driven lung water transport, aquaporins are not required for the physiological clearance of lung water in the neonatal or adult lung, or for the accumulation of extravascular lung water in the injured lung. PMID- 10856129 TI - Functional role of muscle reflexes for force generation in the decerebrate walking cat. AB - To quantify the importance of reflexes due to muscle length changes in generating force during walking, we studied high decerebrate cats that walked on a treadmill. One leg was denervated except for the triceps surae and a few other selected muscles. The triceps surae muscles are ankle extensor muscles that attach to the Achilles' tendon which was cut and connected to a muscle puller. In some steps the EMG activity triggered the puller to move the muscle through the pattern of length changes that are normally produced by ankle movements in intact cats walking over ground (simulated walking). In other steps the muscles were held isometrically. The EMG and force produced during the two types of steps were compared. On average about 50 % more EMG was generated during the E2 part of the simulated stance phase in the triceps surae muscles, but not in other muscles studied. Force was increased significantly over the entire stance phase by about 20 %, when muscle stretches simulating walking were applied. However, during much of the stance phase the triceps surae muscles are shortening and so would produce less force. The effect of shortening was assessed in control experiments in which these muscles were stimulated at a constant frequency, either isometrically or during simulated walking movements. By combining data from the walking and control experiments, we estimate that about 35 % of the force produced in the cat ankle extensors during stance is produced by reflexes due to muscle length changes. Other sensory inputs may also contribute to force production, but the total reflex contribution will vary under different conditions of speed, length, loading, task difficulty, etc. Since a substantial percentage of the force in the stance phase of walking is normally produced by muscle reflexes, this force can be continuously adjusted up or down, if the muscles receive extra stretch or unloading during a particular step cycle. PMID- 10856130 TI - Ischaemia after exercise does not reduce responses of human motoneurones to cortical or corticospinal tract stimulation. AB - Motor unit firing rates and voluntary activation of muscle decline during sustained isometric contractions. After exercise, the responses to motor cortical and corticospinal stimulation are reduced. These changes may reflect motoneuronal inhibition mediated by group III and IV muscle afferents. To determine whether the post-contraction depression of the responses to corticospinal or motor cortical stimulation could be maintained by continued firing of ischaemically sensitive group III and IV muscle afferents, we examined responses in muscles that were held ischaemic after exercise. Following a sustained maximal voluntary contraction (MVC) of the elbow flexors lasting 2 min, the response to stimulation of the corticospinal tract was reduced but the usual recovery (over approximately 2 min) was not delayed when the muscles were maintained ischaemic for 2 min after the contraction. Following a sustained MVC, the time course of the reduction in the response to motor cortical stimulation (a gradual decrease over approximately 2 min, maintained for > 10 min) was also not altered if the muscle was held ischaemic. Mean arterial blood pressure rose to 155 +/- 12 mmHg during the 2 min MVC, declined to 125 +/- 9 mmHg immediately after it, but remained at this level without returning to pre-exercise levels (102 +/- 10 mmHg) until circulation to the arm was restored. This confirms that the sustained MVC activated a reflex dependent on group III and IV muscle afferents. This study shows that ischaemically sensitive group III and IV muscle afferents do not mediate depression of responses to motor cortical or corticospinal stimulation after fatiguing exercise. It also suggests that firing of such afferents does not directly inhibit motoneurones or motor cortical output cells. PMID- 10856131 TI - Neck muscle vibration makes walking humans accelerate in the direction of gaze. AB - We studied the effect of the continuous vibration of symmetrical dorsal neck muscles in seven normal subjects during (a) quiet standing, (b) stepping in place movements and (c) walking on the treadmill. The experiments were performed in a darkened room and the subjects were given the instruction not to resist the applied perturbation. In one condition the velocity of the treadmill was controlled by feedback from the subject's current position. Head, trunk and leg motion were recorded at 100 Hz. In normal standing, neck vibration elicited a prominent forward body sway. During stepping in place, neck vibration produced an involuntary forward stepping at about 0.3 m s-1 without modifying the stepping frequency. If the head was turned horizontally 45 and 90 deg to the right or to the left, neck muscle vibration caused stepping approximately in the direction of the head naso-occipital axis. For lateral eye deviations, the direction of stepping was roughly aligned with gaze direction. In treadmill locomotion, neck vibration produced an involuntary step-like increase of walking speed (by 0.1-0.6 m s-1), independent of the initial walking speed. During backward locomotion, the walking speed tended to decrease during neck vibration. Thus, continuous neck vibration evokes changes in the postural reference during quiet standing and in the walking speed during locomotion. The results suggest that the proprioceptive input from the neck is integrated in the control of human posture and locomotion and is processed in the context of a viewer-centred reference frame. PMID- 10856132 TI - [Recommendations from MENSURA for selection of antimicrobial agents for susceptibility testing and criteria for the interpretation of antibiograms]. AB - This document includes the recommendations from the Spanish antibiogram committee (The MENSURA group, Mesa Espanola de Normalizacion de la Sensibilidad y Resistencia a los Antimicrobianos, under the auspices of the Sociedad Espanola de Quimioterapia and Sociedad Espanola de Enfermedades Infecciosas y Microbiologia Clinica) for the selection of antimicrobials for susceptibility testing. Separate tables for each group of organism with proposed susceptibility and resistance breakpoints are updated and comparatively presented with those of other groups, such us NCCLS, CA-SFM and BSAC. The susceptibility breakpoint tends to identify the fully susceptible population, which probably lacks any specific resistance mechanism. The analysis of MIC distributions for different homogeneous populations (same species) is used to define breakpoints for susceptibility. The resistance breakpoint is based on pharmacological and clinical data obtained when the corresponding antibiotic is administered with a conventional schedule. The primary objective of the Spanish MENSURA group is to contribute to the international consensus on the establishment of breakpoints. PMID- 10856134 TI - Tissue remodeling as a feature of persistent asthma. AB - A chronic inflammatory process is almost invariably associated with tissue damage and healing. Healing results in repair and replacement of dead or damaged cells by viable cells. Repair usually involves 2 distinct processes: regeneration, which is the replacement of injured tissue by parenchymal cells of the same type, and replacement by connective tissue and its eventual maturation into scar tissue. In many instances both processes contribute to the healing response. Chronic inflammatory disease can therefore lead to a wide variety of consequences, from complete or partial restitution of organ structure and function to fibrosis. Asthma is characterized by a chronic inflammatory process of the airways. The ensuing healing process results in structural alterations referred to as a remodeling of the airways. The mechanisms underlying these structural alterations are still largely unknown. They are likely to be heterogeneous, leading-through the highly dynamic process of cell de differentiation, migration, differentiation, and maturation-to changes in connective tissue deposition and to the altered restitution of airways structure, resulting in mucus gland hyperplasia, neovascularization, fibrosis, and an increase in smooth muscle mass. PMID- 10856135 TI - Latex allergy. AB - Latex allergy continues to be an important medical problem. In this review we re examine the definition of latex allergy, the offending allergens, the factors that enhance sensitization, the threshold levels that sensitize and elicit reactions in sensitized individuals, current diagnostic techniques, avoidance measures, the barrier properties of nonlatex alternatives, and the roles of premedication and immunotherapy. Twenty years after its resurgence, latex allergy is a well-defined condition with established diagnostic criteria and rational treatment and prevention strategies. However, in spite of advances associated with molecular studies of latex allergens and improved understanding of immunotherapy, avoidance remains the only effective treatment. PMID- 10856136 TI - IL-4/IL-13 signaling beyond JAK/STAT. AB - In the past several years, extensive studies on the mechanisms underlying IL-4 and IL-13 signaling have enabled us to gain insight into how these cytokines regulate immune responses. Because both IL-4 and IL-13 use the IL-4Ralpha as a receptor component, these cytokines activate many common signaling pathways. Both of these cytokines use Janus kinases (JAKs) to initiate signaling and activate signal transducer and activator of transcription-6 (STAT6), which is a transcription factor required for many of their biologic functions. In addition to JAK/STAT, these cytokines also activate a variety of other signaling molecules that are important in regulating IL-4-induced proliferation and protection from apoptosis. Suppressor of cytokine signaling-1 (SOCS-1) is a molecule that can inhibit the activation of IL-4 signaling through the inhibition of JAKs. The Fes tyrosine kinase is activated by IL-4 and appears to be important in regulating IL 4-induced proliferation through the phosphorylation of insulin receptor substrate (IRS) molecules. IRS molecules are essential for IL-4-induced proliferation through their ability to recruit phosphoinositol-3 kinase to the activated IL-4 receptor kinase. In addition, IL-4 can activate a number of phosphatases including SH2-containing inositol phosphatase (SHIP), SHP-1, and SHP-2. Finally, B-cell lymphoma gene-6 (BCL-6) appears to regulate a subset of IL-4-induced genes. Thus the biologic responses induced by IL-4/IL-13 require a complex interaction of signaling pathways and regulators. PMID- 10856137 TI - Induced sputum--a tool with great potential but not without problems. PMID- 10856138 TI - It is not yet time to stop skin testing, but... PMID- 10856139 TI - Quantitative IgE antibody assays in allergic diseases. AB - During the past several years, immunoassays for specific IgE antibodies have been refined to permit reporting results in mass units. Thus quantitative immunoassays for IgE antibodies may be an adjunct to skin tests. In cases of food allergy among children with atopic dermatitis, cutoff values for IgE antibody concentrations to egg, milk, peanut, and fish have been derived to provide 95% positive and 90% negative predictive values. Food-specific IgE antibody determinations can also be used to predict which food allergies are resolving spontaneously. Elevated egg-specific IgE antibody levels in infancy are associated with significantly increased risk for development of inhalant allergies later in childhood. In cases of inhalant allergy, specific IgE antibody levels correlate closely with results of inhalation challenge studies in cat sensitive persons. Also, mite-specific IgE antibody levels correlate significantly with the mite allergen contents of reservoir dust in the homes of mite-sensitive persons. Immunoassays for quantitation of specific IgE antibodies may be used to document allergen sensitization over time and to evaluate the risk of reaction on allergen exposure. However, immunoassays and skin tests are not entirely interchangeable, and neither will replace the other in appropriate circumstances. PMID- 10856140 TI - Use of phage display technology to investigate allergen-antibody interactions. AB - Phage display is an advanced technology that can be used to characterize the interactions of antibody with antigen at the molecular level. It provides valuable data when applied to the investigation of IgE interaction with allergens. The aim of this rostrum article is to provide an explanation of the potential of phage display for increasing the understanding of allergen-IgE interaction, the discovery of diagnostic reagents, and the development of novel therapeutics for the treatment of allergic disease. The significance of initial studies that have applied phage display technology in allergy research will be highlighted. Phage display has been used to clone human IgE to timothy grass pollen allergen Phl p 5, to characterize the epitopes for murine and human antibodies to a birch pollen allergen Bet v 1, and to elucidate the epitopes of a murine mAb to the house dust mite allergen Der p 1. The technology has identified peptides that functionally mimic sites of human IgE constant domains and that were used to raise antiserum for blocking binding of IgE to the FcepsilonRI on basophils and subsequent release of histamine. Phage display has also been used to characterize novel peanut and fungal allergens. The method has been used to increase our understanding of the molecular basis of allergen-IgE interactions and to develop clinically relevant reagents with the pharmacologic potential to block the effector phase of allergic reactions. Many advances from these early studies are likely as phage display technology evolves and allergists gain expertise in its research applications. PMID- 10856141 TI - Identification of SAF-2, a novel siglec expressed on eosinophils, mast cells, and basophils. AB - BACKGROUND: Eosinophils, basophils, and mast cells are believed to be the central tenet cells in allergic conditions including allergic rhinitis, asthma, and eczema. The molecular mechanisms underlying the recruitment of these cells to sites of allergic inflammation are poorly understood. OBJECTIVES: Our aim was to identify a common adhesion molecule that could potentially be responsible for mediating the recruitment of the allergic cell types to the lungs and other sites of allergy. METHODS: We have cloned a sialoadhesin molecule from a human eosinophil library with the use of expressed sequence tag technology and characterized its expression on allergic cells by the use of flow cytometry and specific mAbs. RESULTS: With the use of expressed sequence tag sequencing, we have identified a novel siglec molecule, SAF-2. SAF-2 has homology with other sialoadhesin family members (CD33 and siglec-5) and belongs to a subgroup of the Ig superfamily. SAF-2 is a 431-amino acid protein composed of 3 Ig domains with a 358-amino acid extracellular domain and a 47-amino acid tail. SAF-2 is highly restricted to eosinophils, basophils, and mast cells. Antibodies to SAF-2 do not modulate Ca(++) mobilization or chemotaxis of human eosinophils induced by eotaxin. CONCLUSION: SAF-2 is a highly restricted sialoadhesin molecule, which may be useful in the detection and/or modulation of allergic cells. PMID- 10856142 TI - Comparison of once-daily ebastine 20 mg, ebastine 10 mg, loratadine 10 mg, and placebo in the treatment of seasonal allergic rhinitis. The Ebastine Study Group. AB - BACKGROUND: Ebastine and loratadine are 2 nonsedating second-generation H(1) antihistamines with once-daily dosing. OBJECTIVE: We compared the efficacy and safety of ebastine 20 mg and 10 mg, loratadine 10 mg, and placebo administered once daily for 4 weeks in controlling the symptoms of seasonal allergic rhinitis (SAR). METHODS: In a double-blind, placebo-controlled, randomized, parallel-group study, 565 patients with ragweed SAR, ages 12 to 70 years, received either ebastine 20 mg, ebastine 10 mg, loratadine 10 mg, or placebo once daily for 4 weeks. Patients recorded morning and evening reflective scores (past 12 hours) as well as snapshot scores (at time of recording) for nasal discharge, congestion, sneezing, itching, and total eye symptoms. Total symptom score (TSS) is the sum of these 5 scores. RESULTS: Ebastine 20 mg produced significantly greater (P <.05) reductions from baseline compared with loratadine 10 mg over the entire treatment period in the mean daily reflective (42.5% vs 36.3%) and mean morning snapshot (40.3% vs 31.3%) TSS. The overall improvement in daily reflective and morning snapshot TSS was comparable between ebastine 10 mg and loratadine 10 mg and significantly better than placebo (P <.05). The total percent of patients with adverse events was similar among all 4 treatment groups (P =.78). CONCLUSION: Ebastine 20 mg given once daily was significantly superior to loratadine 10 mg given once daily at improving the rhinitis total symptom score throughout the day and at awakening over a 4-week period. Ebastine 20 mg and 10 mg doses were both efficacious and well tolerated in the treatment of SAR. PMID- 10856143 TI - Salmeterol and fluticasone propionate combined in a new powder inhalation device for the treatment of asthma: a randomized, double-blind, placebo-controlled trial. AB - BACKGROUND: Many patients with persistent asthma need both long-acting bronchodilators and inhaled corticosteroids for optimal asthma control. OBJECTIVE: Our purpose was to compare the efficacy and safety of salmeterol 50 microg combined with fluticasone 100 microg (in a combination dry powder product) with that of placebo, fluticasone, or salmeterol alone. METHODS: A 12-week randomized, double-blind, multicenter study was conducted in 356 patients aged 12 years or older with asthma. After a 14-day screening period, patients were randomized to treatment with salmeterol 50 microg combined with fluticasone 100 microg (combination product), salmeterol 50 microg, fluticasone 100 microg, or placebo administered in the Diskus dry powder inhaler (GlaxoWellcome, UK) twice daily. RESULTS: Mean change in FEV(1) at end point was significantly (P < or =.003) greater with the combination product (0.51 L) compared with placebo (0.01 L), salmeterol (0.11 L), and fluticasone (0.28 L). The combination product significantly increased (P < or =.013) area under the curve compared with placebo and fluticasone on day 1 and compared with placebo, salmeterol, and fluticasone at week 1 and week 12. Patients in the combination product group were less likely to withdraw from the study because of worsening asthma compared with those in the other groups (P < or =.020). The combination product significantly increased (P < or =.012) morning PEF (combination, 52.5 L/min; placebo, -23.7 L/min; salmeterol, -1.7 L/min; fluticasone, 17.3 L/min) and evening PEF at end point compared with the other groups. The combination product significantly (P < or =.025) reduced symptom scores and albuterol use compared with the other treatments and increased the percentage of nights with no awakenings and the percentage of days with no symptoms compared with placebo and salmeterol. All treatments were equally well tolerated. CONCLUSION: Salmeterol 50 microg and fluticasone 100 microg combined in the Diskus powder delivery device offers significant clinical advantages over salmeterol or fluticasone alone at the same doses. PMID- 10856144 TI - Development of immunologic memory against tetanus toxoid and pertactin antigens from the diphtheria-tetanus-pertussis vaccine in atopic versus nonatopic children. AB - BACKGROUND: Recent findings suggest that a hallmark of the atopic phenotype is reduced capacity to respond to vaccine antigens, as well as to environmental allergens, during infancy. This deficiency, which is most marked for the cytokine IFN-gamma, appears transient but can result in a long-lasting imbalance within T helper cell (T(H)) memory responses to allergens. Indirect evidence suggests that parallel effects may occur within immunologic memory responses against vaccine antigens in atopic children. OBJECTIVE: Our purpose was to compare vaccine antigen-specific T(H) memory responses in atopic and nonatopic children. METHODS: We analyzed specific serum IgG and cytokine responses to pertactin and tetanus antigens as well as to mitogen (PHA) and house dust mite (HDM) allergen in 25 HDM sensitized atopic and 25 nonatopic 6-year-old children who were vaccinated and boosted with diphtheria-tetanus-pertussis (DTP) vaccine. RESULTS: PBMCs from the atopic subjects produced higher levels of T(H)1 and T(H)2 cytokines to HDM allergen and PHA. Vaccine antibody titers were normal in the atopic subjects; vaccine-specific T(H)2 responses were rarely detectable, yet T(H)1 (IFN-gamma) responses, in particular against tetanus, were frequent and higher in the atopic subjects (121.5 [SE 64.3] vs 8.0 [3.5] pg/mL culture fluid, P =.04). Corresponding pertactin responses were comparable in both groups. CONCLUSIONS: At the completion of the full primer-booster DTP vaccination regimen, levels of vaccine-specific immunity in atopic 6-year-old children are at least equivalent to their nonatopic counterparts, indicating that the transient atopy-associated deficiency in T(H)1 function in childhood can be successfully overcome by appropriate vaccination and boosting regimens. PMID- 10856145 TI - Low-dose inhaled fluticasone propionate versus oral zafirlukast in the treatment of persistent asthma. AB - BACKGROUND: Few studies have compared the efficacy of inhaled corticosteroids and leukotriene modifiers for the treatment of persistent asthma. OBJECTIVE: Our purpose was to compare the efficacy of a low dose of inhaled fluticasone propionate (FP) with that of oral zafirlukast in the treatment of persistent asthma previously treated with short-acting beta(2)-agonists alone. METHODS: A 12 week, randomized, double-blind, double-dummy, multicenter study was conducted in 451 patients aged 12 years and older with asthma who were symptomatic on short acting beta(2)-agonists alone. After an 8- to 14-day run-in period, patients were randomized to treatment with FP 88 microg twice daily or zafirlukast 20 mg twice daily. RESULTS: Treatment with FP was more effective than treatment with zafirlukast in increasing morning FEV(1) (by 0.42 L vs 0.20 L over baseline, P <.001), morning peak expiratory flow (by 49.94 L/min vs 11.68 L/min over baseline, P <. 001), and evening PEF (by 38.91 L/min vs 10.50 L/min over baseline, P <.001). Statistically significant differences between the two treatments in FEV(1) were noted after the first observation (week 4) and in morning and evening peak expiratory flow by week 2. Mean change in percentage of symptom-free days was greater with FP than with zafirlukast (28.5% of days vs 15.6% of days, P <.001) and FP significantly increased the percentage of rescue free days by 40.4% of days compared with 24.2% of days with zafirlukast (P <.001). Treatment with FP significantly reduced albuterol use by 2.39 puffs per day compared with 1.45 puffs per day (P <.001) and increased the percentage of nights with no awakenings by 21.2% of nights compared with 8.0% of nights with zafirlukast (P <.001). CONCLUSION: The clinical effectiveness of a low dose of FP as first-line therapy in patients with persistent asthma who are symptomatic on beta(2)-agonists alone is superior to that of zafirlukast. PMID- 10856146 TI - Domestic control of house dust mite allergen in children's beds. AB - BACKGROUND: House dust mite allergen levels in humid coastal regions of Australia are high, particularly in beds. Because high allergen levels in beds are associated with more severe asthma, reduction of levels may be important for asthma control. OBJECTIVE: We tested the effectiveness of an acaricidal treatment of bedding in combination with occlusive mattress and pillow encasings in reducing allergen levels in children's beds in a community setting. METHODS: A total of 14 beds of children were selected for the active intervention. In each home the bed of a sibling of nearest age was selected as the control. Dust was vacuumed from beds by using a standard protocol, and Der p 1 levels were measured by using ELISA. Adjacent settling dust was collected by using opened Petri dishes. The intervention consisted of encasing mattresses and pillows in occlusive covers and washing all bedding with Acaril, an acaricidal additive. The acaricidal wash was repeated twice in 7 households at 2-month intervals. Control beds were not treated. RESULTS: The mean Der p 1 concentration at baseline was 27.9 microg/g in the active beds and 18.1 microg/g in the control beds. At 4 days after intervention, Der p 1 decreased to 3.2 microg/g and 15.7 microg/g in active and control beds, respectively. The average difference (active minus control) over the first 8-week cycle was 78.5% (P <.0001), and the difference over 3 washing cycles was 125.1% (P <.05). The mean rate of settling Der p 1 adjacent to the actively treated beds decreased from 24.4 ng.m(-2).d(-1) at baseline to 10.0 ng.m(-2).d(-1) after intervention (P <.01). CONCLUSION: A substantial reduction in Der p 1 levels in beds and in airborne dust in a humid region with naturally high house dust mite allergen levels can be achieved and sustained in a community setting with use of occlusive covers and a rigorous washing routine. PMID- 10856147 TI - Effect of inhaled indomethacin in asthmatic patients taking high doses of inhaled corticosteroids. AB - BACKGROUND: Cyclooxygenase products of arachidonic acid may play a part in bronchoconstriction and airway inflammation in asthma. OBJECTIVE: We sought to determine the effect of inhaled indomethacin on asthma control and asthma exacerbations during reduction of inhaled corticosteroids in patients with moderate-to-severe steroid-dependent asthma. METHODS: We conducted a double blind, randomized, parallel-group, multicenter study in 38 patients with asthma taking high doses (> or =1500 microg/d) of beclomethasone dipropionate (BDP). After a run-in period, patients were assigned inhaled indomethacin (50 mg/d) or placebo for 6 weeks, during which the daily doses of BDP were reduced to half at week 2 and then to one third of the baseline dose at week 4. RESULTS: Data were available from 34 patients. After the reduction of BDP doses, FEV(1), peak expiratory flow, asthma symptoms, and exhaled nitric oxide concentrations deteriorated in both treatment groups, but these effects were less pronounced in the indomethacin group compared with the placebo group. During the 6-week treatment period, 89% of the patients receiving placebo had relapse of asthma, whereas only 38% of those receiving inhaled indomethacin did so (P =.003). CONCLUSION: Inhalation of indomethacin can reduce asthma exacerbations induced by reduction of high-dose inhaled corticosteroid in steroid-dependent asthma. PMID- 10856148 TI - Expulsion of allergen-containing materials from hydrated rye grass (Lolium perenne) pollen revealed by using immunogold field emission scanning and transmission electron microscopy. AB - BACKGROUND: Several studies demonstrated episodes of grass pollen-induced allergic asthma after heavy rainfalls. It has been hypothesized that these asthma attacks might be due to the release of respirable allergen-bearing particles from pollen cytoplasm. OBJECTIVE: In this study we investigated the release mechanism of the most potent and frequently recognized grass pollen allergens, group 1 and group 5, from freshly harvested and subsequently hydrated rye grass pollen at the ultrastructural level. METHODS: Rabbit antisera against purified recombinant group 1 and group 5 allergens were used to investigate, by using field emission scanning and transmission immunogold electron microscopy, the allergen release from rye grass pollen grains into isotonic aqueous solutions or water. RESULTS: Pollen grains exposed to isotonic aqueous solutions remained intact and released allergens by means of diffusion. However, pollen grains hydrated in distilled water or rainwater expelled starch grains and cytoplasmic debris of respirable size. Group 1 and group 5 allergens were observed on and within these materials. CONCLUSIONS: Exposure of rye grass pollen to water leads to an expulsion of subcellular allergen-containing pollen components of respirable size. Our ultrastructural data thus support the idea that this release of allergen containing respirable pollen materials may be a cause of asthma attacks after heavy rainfalls. PMID- 10856149 TI - Upregulation of the transcription factor GATA-3 in upper airway mucosa after in vivo and in vitro allergen challenge. AB - BACKGROUND: Allergic rhinitis is a complex upper airways disorder characterized by the infiltration of eosinophils and T(H2)-type T lymphocytes. GATA-3 is a novel transcription factor recently shown to regulate IL-5 and, possibly, IL-4 gene expression. We previously reported that GATA-3 is increased within the bronchial mucosa of allergic asthmatic subjects compared with control subjects. OBJECTIVE: In the present study we set out to determine whether there is also an increased number of cells expressing GATA-3 messenger (m)RNA within the nasal mucosa of patients with allergic rhinitis. METHODS: Inferior turbinate biopsy specimens were obtained from patients with allergic rhinitis and nonatopic control subjects before and after local allergen provocation in vivo. To assess the contribution of resident cells expressing GATA-3 mRNA, we also performed isolated explant studies in which nasal mucosal tissue from subjects with allergic rhinitis and nonatopic control subjects was cultured in allergen-treated medium. The presence of mRNA coding for GATA-3, IL-5, IL-4, IL-13, and GM-CSF was assessed by using in situ hybridization. RESULTS: The number of GATA-3 mRNA(+) cells was increased after local allergen provocation in vivo (increase in GATA-3 mRNA(+) cells [mean +/- SEM]: subjects with allergic rhinitis, 11.3 +/- 8.7; control subjects, 1.2 +/- 4.1; P <.05) and in explanted nasal mucosa in vitro (subjects with allergic rhinitis, 10. 2 +/- 3.8; control subjects, 2.7 +/- 4.4; P <.05). The gene expression of GATA-3 was significantly correlated to the numbers of IL-5 (r = 0.87) and GM-CSF (r = 0.79) mRNA(+) cells but not with IL-4 or IL-13 mRNA(+) cells. CONCLUSION: In summary, the expression of the transcription factor GATA-3 was increased after allergen challenge, and this was evident in the absence of de novo inflammatory cell recruitment. GATA-3 may be a potential target in the treatment of allergic diseases, such as rhinitis. PMID- 10856150 TI - Comparison of once- and twice-daily dosing of fluticasone propionate 200 micrograms per day administered by diskus device in patients with asthma treated with or without inhaled corticosteroids. AB - BACKGROUND: There are limited published data regarding the efficacy of once- versus twice-daily administration of flutica-sone propionate. OBJECTIVE: Our purpose was to evaluate the effectiveness of fluticasone propionate powder 200 microg/d administered as a once- or twice-daily dosage regimen in patients who were currently being treated with bronchodilators only (BD patients) and in patients who required inhaled corticosteroids for maintenance treatment of asthma (ICS patients). METHODS: Five hundred seventy patients were randomly assigned to receive one of the following inhaled treatments through the Diskus device (Glaxo Wellcome, Research Triangle Park, NC) for 12 weeks: fluticasone propionate 100 microg twice daily (FP100BID) or 200 microg once daily (FP200QD) or placebo. RESULTS: BD patients treated with FP100BID, FP200QD, and placebo had mean increases in FEV(1) from baseline to end point of 0. 49 L, 0.37 L, and 0.21 L, respectively (P <.001, FP100BID vs placebo; P =.05, FP200QD vs placebo). ICS patients treated with FP100BID and FP200QD had mean increases in FEV(1) of 0.27 L and 0.11 L, respectively, compared with a decrease in FEV(1) of -0.08 L with placebo (P <.001, FP100BID vs placebo; P =.023, FP200QD vs placebo). BD patients treated with FP100BID and FP200QD had mean increases in morning peak expiratory flow from baseline to end point of 31 L/min and 27 L/min, respectively, compared with a 1 L/min increase in patients treated with placebo. ICS patients treated with FP100BID had a mean increase in morning peak expiratory flow (from baseline to end point) of 18 L/min compared with mean decreases of -3 L/min and -12 L/min in the FP200QD and placebo groups, respectively. More patients were withdrawn from placebo (26% and 48%, in BD and ICS patients, respectively) than from fluticasone propionate (7%-9% [BID-QD] and 18%-32% [BID-QD], in BD and ICS patients, respectively) because of failure to meet predetermined asthma stability criteria. CONCLUSION: The efficacies of FP100BID and FP200QD were comparable with regard to improvement in pulmonary function and asthma stability in BD patients. In ICS patients, asthma control was maintained with FP200QD, whereas FP100BID provided greater improvements in pulmonary function and asthma stability. PMID- 10856151 TI - Induced sputum: validity of fluid-phase IL-5 measurement. AB - BACKGROUND: IL-5 measurement in the fluid phase of induced sputum is considered to be important in the assessment of asthma, but the validity of these measurements is uncertain. OBJECTIVE: We investigated the validity of sputum IL-5 measurements through a series of spiking experiments and examined the effect of dithiothreitol (DTT) on these measurements. METHODS: Induced sputum from 26 asthmatic subjects was spiked with IL-5 and processed, and the percentage of recovery was measured by means of immunoassay. In 6 of the 26 samples the effect of adding albumin to the processing fluids was studied. In 3 separate samples radiolabeled IL-5 was added, and the recovery measured by means of gamma counting and immunoassay were compared. In addition, the effect of DTT on the immunoassay was examined. RESULTS: The mean +/- SD recovery of spiked IL-5 was 26.1% +/- 14.6% measured by means of immunoassay; adding albumin increased the recovery to 47.7% +/- 8.0% (P <.001). The mean recovery measured by means of gamma counting was 84.8% +/- 5.7% (P <.001); adding albumin had no effect on recovery. DTT had no significant effect on IL-5 measurement. CONCLUSION: The validity of IL-5 measurement by means of current methods is poor. The discrepancy in recovery as measured by gamma counting compared with immunoassay suggests that there is a problem with the recognition of IL-5 epitopes by immunoassay in induced sputum. This cannot be attributed to DTT but may be due to other interfering substances present in sputum, such as sputum proteases, soluble receptors, or autoantibodies. PMID- 10856152 TI - The effect of an experimental rhinovirus 16 infection on bronchial lavage neutrophils. AB - BACKGROUND: Viral respiratory tract infections are the most frequent cause of asthma exacerbations. Of the respiratory viruses associated with these exacerbations, rhinovirus (RV) is the most common. It is proposed that these RV infections may enhance airway inflammation and thus provoke asthma. OBJECTIVE: It is our hypothesis that RV infections generate nasal proinflammatory mediators that are associated with an initial increase in circulating leukocytes and may contribute to later development of neutrophilic airway inflammation. METHODS: To evaluate this hypothesis, subjects with a history of allergic asthma were experimentally inoculated with strain 16 RV (RV16). The effect of this experimental infection was evaluated on circulating leukocytes, nasal-derived mediators, and markers of bronchial inflammation that were obtained by bronchoscopy and lavage. RESULTS: RV16 inoculation was associated with an initial increase in circulating neutrophils. Paralleling these acute changes in circulating neutrophils was an increase in nasal concentrations of IL-8 and granulocyte-colony-stimulating factor (G-CSF). The RV16-associated changes in circulating and nasal G-CSF correlated with increases in peripheral blood neutrophils (r(s) = 0.874, P <. 001 and r(s) = 0.898, P <.001, respectively). Bronchial lavage samples showed no increase in neutrophils 48 hours after RV16 inoculation; however, 96 hours after RV inoculation there was a significant increase in bronchial neutrophils compared with preinoculation values. CONCLUSIONS: These results suggest that the production of nasal mediators associated with the RV infection, particularly G-CSF, may be important to the eventual development of neutrophilic bronchial inflammation and thus contribute to asthma exacerbations. PMID- 10856153 TI - Immunofluorescence analysis of cytokine and granule protein expression during eosinophil maturation from cord blood-derived CD34 progenitors. AB - BACKGROUND: In allergic inflammation and asthma, eosinophils are major effector cells. They have been shown to synthesize at least 23 cytokines, some of which are stored intracellularly in their unique crystalloid granules together with cationic granule protein. Little is known about the synthesis and storage of cytokines relative to cationic granule proteins in maturing eosinophils during eosinophilopoiesis. OBJECTIVE: Our purpose was to analyze the expression of eosinophil-derived mediators, major basic protein (MBP), eosinophil cationic protein (ECP), IL-6, and RANTES, during early stages of eosinophil maturation in CD34(+) cell-derived colonies. METHODS: Purified human cord blood CD34(+) cells were grown in methylcellulose cultures in the presence of recombinant human IL-3 and IL-5. By confocal laser scanning microscopy, the coexpression of eosinophil granular proteins MBP and ECP was determined concurrently with IL-6 and RANTES during eosinophil maturation on days 16, 19, 23, and 28 of culture. RESULTS: Immunoreactivity against MBP, ECP, IL-6, and RANTES was not detectable in freshly purified CD34(+) cells. Maturing eosinophils (>95%) exhibited positive immunostaining for all these proteins between days 16 and 28 of culture. At early stages of culture, discrete immunostaining was observed around the periphery but not in the center of granular structures. By day 28 cultured eosinophil-like cells showed evidence of the acquisition of crystalloid granule-like structures, analogous to those observed in mature peripheral blood eosinophils. CONCLUSIONS: Eosinophils express and store cytokines simultaneously with cationic granule proteins during the process of maturation. We propose that the storage of cytokines during the development of eosinophils is an early event and it may be integral to inflammatory responses involving these cells. The results of this study suggest a potential immunoregulatory function for maturing eosinophils. PMID- 10856154 TI - Protease-dependent activation of epithelial cells by fungal allergens leads to morphologic changes and cytokine production. AB - BACKGROUND: Proteases in extracts of Aspergillus fumigatus cause epithelial cell desquamation and release of proinflammatory cytokines. OBJECTIVE: We sought to assess protease activity in Alternaria alternata, Cladosporium herbarum, and Aspergillus fumigatus extracts and study the ability of these extracts to cause desquamation and release of proinflammatory cytokines from epithelial cells. METHODS: Protease activities of the fungal extracts were quantified. Changes with respect to cell morphology, cell desquamation, and cytokine production (IL-6 and IL-8) were measured in the absence and presence of the fungal extracts in an airway-derived epithelial cell line (A549) and primary epithelial nasal cells. RESULTS: Fungal proteases differentially induced morphologic changes, cell desquamation, and production of IL-6 and IL-8 in a dose- and time-dependent fashion. Alternaria alternata extracts induced cell shrinking and cell desquamation and strongly enhanced the production of IL-6 and IL-8 at higher concentrations. Aspergillus fumigatus extracts caused cell shrinking, cell desquamation, and production of IL-6 and IL-8, even at low concentrations. The Aspergillus fumigatus-derived extract grown on collagen medium induced a strong dose-dependent decline in cytokine production at higher concentrations. Cladosporium herbarum extracts did not induce morphologic changes or cell desquamation but enhanced IL-6 and IL-8 productions at higher concentrations. The dependence of these effects on intact protease activity was shown by their abrogation by protease inhibitors. CONCLUSION: Proteases present in fungal extracts interact with epithelial cells, leading to morphologic changes, cell desquamation, and induction of proinflammatory cytokines. It is proposed that these fungal proteases may activate epithelial cells through a protease-activated receptor type 2-driven mechanism. PMID- 10856155 TI - Langerhans-like dendritic cells generated from cord blood progenitors internalize pollen allergens by macropinocytosis, and part of the molecules are processed and can activate autologous naive T lymphocytes. AB - BACKGROUND: The safety and efficacy of sublingual immunotherapy have been demonstrated in moderate allergic asthma and seasonal rhinitis. However, not much is known about the precise mechanism of action of the allergen when it crosses the oral mucosa. OBJECTIVE: To define this mechanism, we investigated the role of Langerhans' cells in the capture and internalization of allergens. METHODS: We generated dendritic cells in vitro with the phenotypic characteristics of Langerhans-like dendritic cells (LLDCs) from cord blood CD34(+) progenitors. We used two recombinant major allergens: Bet v 1 and Phl p 1 labeled with FITC. RESULTS: Internalization of allergens and control proteins was dose- and time dependent and related to the immature state of the cells. LLDCs internalized allergens with a high efficiency in comparison with control molecules. Allergens were only internalized by macropinocytosis, as demonstrated by the use of various inhibitors. Addition of intracellular pH-modifying molecules indicated that only a part of the allergens was accumulated in acidic vesicles, whereas the majority remained in other cytoplasmic structures. Pulse-chase experiments calculated a half-life of 4 hours, suggesting that part of the molecules were not metabolized in the lysosome. Allergen internalization by LLDCs might be followed by processing in some experiments, as demonstrated by activation of autologous T lymphocytes in 4 of 9 experiments. CONCLUSION: These elements showed that Langerhans' cells present in mucosa might play an active role in immune responses to allergens. PMID- 10856156 TI - Ligation of IgE receptors causes an anaphylactic response and neutrophil infiltration but does not induce eosinophilic inflammation in mice. AB - BACKGROUND: Eosinophils are selectively recruited into the tissues during chronic allergic inflammation. IgE is considered an initiator of the allergic reaction; however, the roles of IgE in allergic inflammation are not fully understood. OBJECTIVE: We tested the hypothesis that antigen interaction with specific IgE antibody provokes eosinophilic inflammation. METHODS: BALB/c mice were actively sensitized with ragweed extract and passively sensitized with anti-dinitrophenyl (anti-DNP) mouse IgE and challenged intraperitoneally by injecting either ragweed extract or DNP-ovalbumin (OVA). Immediate anaphylactic responses were examined by monitoring vascular permeability and by measuring histamine content in peritoneal lavage fluids. Late-phase allergic responses were examined by total cell counts and cell differentials. RESULTS: Mice sensitized and challenged with ragweed showed immediate anaphylactic responses followed by temporal increases in neutrophils at 3 to 12 hours and sustained increases in eosinophils in their peritoneal cavities after 24 hours. Double-sensitized mice (ie, sensitized actively for ragweed and passively for DNP-OVA) challenged with ragweed showed immediate anaphylactic responses and peritoneal eosinophilia at 48 hours. Double sensitized mice challenged with DNP-OVA showed comparable immediate anaphylactic responses but no peritoneal eosinophilia. Furthermore, at 8 hours, ragweed challenged animals recruited both eosinophils and neutrophils, but DNP-OVA challenged animals recruited only neutrophils. Finally, after active sensitization and challenge with ragweed, mast cell-deficient mice (WBB6F1 W/W(v)) lacked the immediate response but showed comparable eosinophil accumulation as their litter mate controls (WBB6F1-+/+). CONCLUSION: Interaction of antigen with IgE antibody is insufficient to provoke eosinophilic inflammation in mice. PMID- 10856157 TI - Autoantibodies to DFS 70 kd/transcription coactivator p75 in atopic dermatitis and other conditions. AB - BACKGROUND: Sera of patients with atopic dermatitis (AD) were found to have autoantibodies that reacted with tissue culture cell substrates in immunohistochemistry to display a characteristic pattern of nuclear distribution of dense fine speckles. The sera also recognized a 70-kd protein on Western immunoblots, and the antigen was termed dense fine speckles 70 kd (DSF70). OBJECTIVE: Because spontaneously occurring autoantibodies could be immune responses to proteins that might be participating in the disease process, it was of interest to identify the antigens driving the autoimmune antibody response. METHODS: A serum containing high-titer antibodies to DFS70 was used to immunoscreen a complementary (c)DNA expression library to isolate cDNA encoding the antigen. After the cDNA was isolated, this was used to express recombinant protein to determine the prevalence of antibody in AD and other conditions. RESULTS: Thirty percent of patients with AD were found to have antibody to recombinant DFS70 in Western immunoblots. Sixteen percent of patients with asthma and 9% of patients with interstitial cystitis had antibodies of the same specificities. The cDNA encoding DFS70 was identical to a transcription coactivator called p75, which had been shown to be required for RNA polymerase II dependent transcription. Another important finding was that IgE antibodies to DFS70 were also present in AD sera. CONCLUSION: It is suggested that a common basis for the presence of autoantibodies to DFS70 might be related to AD in asthma, interstitial cystitis, and other conditions. A possible role of this antigen-antibody system in pathogenesis remains to be demonstrated, but it appears to be a marker for a subset of patients with AD. PMID- 10856158 TI - Analytic precision and accuracy of commercial immunoassays for specific IgE: establishing a standard. AB - BACKGROUND: Different laboratory assays are used to detect and measure specific IgE antibodies. No standard exists to assess their analytic performance. OBJECTIVE: We sought to analyze reported specific IgE results from different laboratories on the same serum samples for their accuracy and precision. METHODS: Blinded serum samples (26) containing variable levels of specific IgE to 17 common aeroallergens were sent on 3 different occasions through normal channels to 6 laboratories that used 5 different test procedures. Six samples were presented as a dilution series. Laboratory-assay performance was assessed by analyzing the reported results (n = 12, 708) by using ordinary least squares regression with slope coefficients, the t statistic, SEs, confidence intervals, and R (2) values. These were compared with a theoretic ideal assay as the reference. RESULTS: Analysis revealed that one system used in two different laboratories performed nearly as well as the ideal standard, with an overall average slope (0.97; range, 0.91-1.01), SE (0.05; range, 0.02-0.16), R (2) value (93%; range, 0.64-0.99), and coefficient of variation (10.3%; range, 6%-14%). Extensive variability was observed in the other 4 laboratory-assay systems with respect to overall average slope (0.76; range, 0.11-1.24), SE (0.19; range, 0.03 0.95), R (2) value (53%; range, 0.00-0.98), and coefficient of variation (19%; range, 5%-49%). For some specific allergens, some laboratories-assays were not able to detect serial dilutions of the same sample. CONCLUSIONS: One commercial system used in two different laboratories performed nearly as well as the ideal standard. Four of the laboratories-assays for specific IgE antibodies demonstrated substandard overall performance with multiple instances of poor precision and accuracy, particularly for certain allergens, such as weeds and molds. PMID- 10856159 TI - Safety of a two-day ultrarush insect venom immunotherapy protocol in comparison with protocols of longer duration and involving a larger number of injections. AB - BACKGROUND: Insect venom immunotherapy (VIT) is initiated by a dose increase protocol administered usually over 7 to 9 days. Shorter protocols have the advantage of reducing the patient's stay in the hospital. Very few data are currently available on the safety of shorter VIT dose increase protocols. OBJECTIVE: The aim of this study was to investigate whether a reduction in the duration of the VIT dose increase protocol from 7 to 9 days to 2 days causes an increase in the incidence and severity of adverse reactions. METHODS: Between 1992 and 1997 we administered VIT to 1055 patients allergic to bee or wasp venom. We shortened the 7- to 9-day rush protocol stepwise to 2 days by reducing the number of injections and increasing the initial dose and compared the incidence and severity of adverse reactions. The patients were retrospectively divided into 3 cohorts: 20 injections over 7 to 9 days (cohort 1, 317 patients), 10 to 14 injections over 3 to 6 days (cohort 2, 335 patients), and 9 injections over 2 days (cohort 3, 403 patients). RESULTS: We observed no severe adverse reactions in any of the cohorts during VIT. Adverse reactions were treated in 7.1% of the patients by oral and in 2.9% by intravenous antihistamines and in 0.8% by systemic corticosteroids. The incidence of adverse reactions declined significantly from 22.4% in cohort 1 to 13.7% in cohort 2 and 10.7% in cohort 3 with reduced number of injections (P <.001). CONCLUSION: The incidence and severity of adverse reactions decline if the VIT dose increase protocol is shortened to 2 days. PMID- 10856160 TI - Detection of IgA antibodies to cat, beta-lactoglobulin, and ovalbumin allergens in human milk. AB - BACKGROUND: The relationship between the development of allergy during infancy and breast-feeding remains controversial. This controversy may be due to individual variations in the composition of human milk. Antibodies to food antigens to which the mother is commonly exposed are present in the milk, but their relationship to allergy is still unknown. IgA antibodies to inhalant allergens have not been previously detected. OBJECTIVE: Our purpose was to analyze secretory IgA antibody levels to cat, beta-lactoglobulin, and ovalbumin allergens in colostrum and mature milk in relation to maternal allergy. METHODS: Colostrum and samples of mature milk were obtained after 1 and 3 months of lactation from 53 nursing mothers (17 allergic and 36 nonallergic mothers) and were analyzed for total secretory IgA levels by ELISA and secretory IgA antibodies to cat, beta-lactoglobulin, and ovalbumin by an enzyme-amplified ELISA. The specificity of the assays was confirmed by inhibition experiments. RESULTS: Secretory IgA to cat, beta-lactoglobulin, and ovalbumin allergens were detected in colostrum as well as mature milk. The levels of secretory IgA to ovalbumin were lower in colostrum from allergic mothers with P =.016, whereas the levels to beta-lactoglobulin and cat were similar in the 2 groups. IgA antibodies to ovalbumin were detected in 94% of the colostrum samples from allergic and in all samples from nonallergic mothers, in 82% and 96%, respectively at 1 month, and 53% and 65% at 3 months. Fewer samples had detectable secretory IgA antibodies to beta-lactoglobulin than to ovalbumin and cat, and only 33% and 10% of the samples from the allergic and nonallergic mothers, respectively, remained positive at 3 months. All the allergic mothers had detectable IgA to cat in colostrum, whereas 83% and 73% of the samples were positive at 1 and 3 months. The corresponding numbers were 93%, 81%, and 81% in the nonallergic mothers (not significant). CONCLUSION: Even a low level of exposure of the mucosa (eg, by inhalant allergens) can induce antibody secretion into the milk, both in allergic and nonallergic mothers. PMID- 10856161 TI - Occupational asthma caused by a natural food colorant derived from Monascus ruber. PMID- 10856163 TI - Mechanisms and advances in allergic diseases. AB - There have been many attempts to explain the increases in the incidence of allergic diseases, including hay fever and allergic asthma, that have been documented worldwide in recent decades. Epidemiologic studies offer rich opportunities to uncover sometimes unexpected correlations between lifestyle, environmental exposures, temporal development of the immune system, and genetics. Examples include the differing prevalence of atopy, bronchial hyperresponsiveness, and asthma in East and West Germany around the time of reunification, which suggests that a "western lifestyle presents a greater risk for the development of allergic responses than the more traditionally suspected factor of outdoor air pollutant levels. Other epidemiologic studies suggest how infections may interface with an atopic patterning: Evidence from natural measles exposure and nonwheeze-inducing lower respiratory tract infections in young children implicate early childhood viral infections as protective against the development of atopy and airway allergic sensitivity, although in later life viral airway infections exacerbate asthma symptoms. These studies and others involving the scrutiny of lymphocyte subtypes in atopic individuals, notably T(H1) and T(H2) cells, are helping to formulate a theory of interdependence between the early development of the immune system, allergen exposure, and the diverse community of airway cells whose secretory products generate the final physiologic response pattern. PMID- 10856164 TI - Unique mechanistic features of allergic rhinitis. AB - Symptoms of allergic rhinitis are produced by inflammatory mediators that are released upon activation of mast cells by antigen-IgE interaction. These mediators target the end organs directly or indirectly. Stimulation of sensory nerves by histamine, for example, leads to sneezing, pruritus, rhinorrhea, and nasal congestion. The clinical presentation of allergic rhinitis is also characterized by the phenomenon of hyperresponsiveness to nonallergic stimuli, such as cold air and various irritants. This phenomenon is believed to result from the effect of allergic inflammation on the sensory nerves that supply the upper airway mucosa. Various nonallergic triggers have been shown to act on the nasal mucosa through sensorineural stimulation. In allergic rhinitis, responsiveness to these stimuli is increased compared with the healthy state. A similar phenomenon is observed against such products of the allergic reaction as his-tamine and bradykinin. Also, in allergic rhinitis, stimulation of sensory nerves per se can produce inflammatory changes, a phenomenon known as neurogenic inflammation. The mechanism behind the development of sensorineural hyperresponsiveness and of increased propensity for neurogenic inflammation is unknown. However, evidence exists that the neurotrophin nerve growth factor, which can induce all these changes on sensory nerves, is produced in the human nasal mucosa and found in higher quantities in nasal secretions of patients with perennial allergic rhinitis as compared with healthy control subjects. Also, nerve growth factor is acutely released into nasal fluids after allergen provocation of patients with allergic disease. In patients with asthma of atopic origin, allergic rhinitis is almost ubiquitous. Because the nose is the air conditioner of the respiratory system, its dysfunction may negatively affect the lower airways. In addition to the conditioning of inhaled air, the association between allergic rhinitis and asthma may involve various mechanisms. For example, allergen provocation in the nose of a patient with asthma can lead to reductions in pulmonary function and to increased lower airway responsiveness after several hours. Also, nasal inflammation may propagate through a systemic route to affect the lower airways. PMID- 10856165 TI - Complications of allergic rhinitis. AB - Upper and lower respiratory diseases, including asthma, sinusitis, and otitis media with effusion, frequently complicate allergic rhinitis. The close association of nasal allergies with these conditions has been supported by extensive epidemiologic evidence. Similar models have been proposed to explain the pathophysiologic links between allergic rhinitis and both sinusitis and otitis media with effusion. In these models, inflammation caused by nasal allergy and/or viral infection leads to obstruction, fluid accumulation, bacterial infection, and acute disease. If these diseases are unsuccessfully treated, a chronic state of inflammation, obstruction, and infection develops that can cause mucosal damage and, ultimately, chronic disease. A number of studies have investigated the roles and interactions of viruses and allergens in the development of otitis media with effusion. Diagnosing and prophylactically treating nasal allergies in patients with this condition may help prevent recurrent episodes and improve the response to therapy. PMID- 10856166 TI - Allergic rhinitis: treating the adult. AB - Allergic rhinitis is now recognized as a chronic medical condition that markedly affects patient quality of life and is a cause of substantial medical care expenditures. Effective treatment of adults with allergic rhinitis usually requires an integrated regimen that combines allergen avoidance measures, pharmacotherapy, and possible specific-allergen immunotherapy. This approach can control bothersome symptoms with minimal adverse effects in most patients. New medications, such as anti-immunoglobulin E therapy and cytokine antagonists, may provide relief to patients who are refractory to or do not tolerate currently available treatments. PMID- 10856167 TI - Therapeutic approaches to allergic rhinitis: treating the child. AB - Allergic rhinitis is currently the most common of all chronic diseases in children. However, children frequently lack the ability to verbalize their symptoms, with the result that the condition may go undiagnosed and untreated. Unfortunately, untreated allergic rhinitis not only detrimentally affects children's physical and psychosocial well-being, quality of life, and capacity to function and learn, but it is also associated with and may contribute to potentially serious sequelae, including asthma, sinusitis, and otitis media. Because children may not accurately describe their symptoms, the classic signs of allergic rhinitis in the pediatric population, including the allergic shiner, the allergic crease, and the allergic salute, are particularly important in enabling the clinician to recognize those children who may have this condition; other significant signs include mouth breathing, snoring, chronic cough, and continual throat clearing. The options for treating allergic rhinitis in the child are the same as those for the adult, and the clinician can expect the same level of efficacy. Environmental control for allergen avoidance is an important goal, but the clinician must prescribe it within the context of the family's lifestyle to obtain compliance. Complete avoidance of inhalant allergens is not always feasible, and medications are necessary. Oral antihistamines remain the mainstay of initial treatment for allergies. Given evidence of the significant deleterious effects of the sedating antihistamines on learning, the clinician should prescribe nonsedating second-generation agents whenever possible. Decongestants may be needed. Intranasal corticosteroids are a most effective option, and these agents lack the systemic side effects associated with orally administered steroids. In persistent disease, allergen immunotherapy injections may be considered. In all cases, the clinician should consider issues that are likely to influence compliance in the pediatric population. PMID- 10856168 TI - The effects of antihistamines on cognition and performance. AB - Allergic diseases are responsible for substantially more disability than is generally realized. Allergic rhinitis alone results in 3.5 million lost workdays and 2 million missed school days in the United States each year. Comorbid conditions such as asthma and sinusitis can be disabling as well, resulting each year in more than 10 million missed school days and more than 73 million days of restricted activity, respectively. Antihistamines continue to be the mainstay of treatment for allergic disorders. In the case of the first-generation antihistamines, however, the treatment may well be worse than the disease. Although these agents are effective H(1)-receptor antagonists, they are also highly lipophilic and readily cross the blood-brain barrier, causing considerable sedation. The second-generation agents are more lipophobic and possess different ionic charges than the first-generation antihistamines. As a result, they are far less likely to cross the blood-brain barrier and, for that reason, cause little if any sedation. In a recent comparative trial, subjects who were treated with the first-generation agent diphenhydramine were found to have significant performance deficits on tests of divided attention, working memory, vigilance, and speed. By contrast, subjects who were treated with the second-generation antihistamine loratadine performed as well as subjects who were treated with placebo. The sedative effects of the first-generation agents persist well into the next day and thus can potentially interfere with daytime performance and safety even when taken the night before. It is therefore recommended that patients whose occupations require vigilance, divided attention, or concentration receive only second-generation antihistamines. PMID- 10856169 TI - The importance of allergens in the development of asthma and the persistence of symptoms. AB - The importance of allergies and allergens in the development and persistence of asthma is suggested by 3 lines of evidence. First, a number of epidemiologic studies demonstrate that sensitization to indoor allergens and the spores of the outdoor seasonal fungus Alternaria is a risk factor for the development of asthma in both children and adults. Sensitivity to pollens, on the other hand, rarely constitutes a risk for asthma but does constitute a risk for seasonal allergic rhinitis. Second, several studies, again in both children and adults, have demonstrated that, in persons sensitive to indoor allergens, the severity of asthma symptoms will vary with the level of exposure. Third, the elimination of exposure to house-dust mites has produced a remarkable reversal of asthma in sensitive children and adults. Not only have symptoms and pulmonary function improved, but there has also been evidence of a reduction in airway inflammation and hyperresponsiveness. Taken together, these studies make a strong argument for the importance of allergy and allergen exposure as aggravating factors in asthma in both children and adults and reinforce the importance of the identification and treatment of these allergen sensitivities. PMID- 10856170 TI - Inflammatory events in asthma: an expanding equation. AB - Asthma is a chronic inflammatory disorder that can lead to progressive, potentially irreversible declines in lung function in some patients. Asthmatic inflammation develops when the sequential interaction of inflammatory cells with resident cells generates a cascade of events that contribute to the chronic inflammation and clinical manifestations associated with the disease, including further inflammation, airway smooth muscle spasm (bronchospasm), airway mucus secretion, airway edema and narrowing, and bronchial epithelial damage. Because of the chronic, progressive nature of asthmatic inflammation and the early age of onset, the ability to evaluate inflammation in children would be useful. Several procedures that quantify inflammatory mediators (in peripheral blood, induced sputum, bronchoalveolar lavage fluid, and bronchial biopsies) have shown potential usefulness in the evaluation of and the monitoring of disease severity in children (and, by extension, adults) with asthma. Further research needs to be devoted to the elucidation of when the inflammatory process starts and how it changes over time, to the determination of whether the inflammatory process is the same in all patients with wheezing, regardless of the stimulus, to the definition of the relationship between atopy and asthma, and to the establishment of the usefulness of testing for inflammatory markers to help identify individual asthmatic phenotypes, to evaluate disease severity, to measure therapeutic response, and/or to predict potential outcomes. PMID- 10856172 TI - Evolutionary bases for cerebral localization of higher cognitive functions PMID- 10856171 TI - Understanding drug allergies. AB - At this time, the incidence of adverse drug reactions can only be estimated because the intensive monitoring and documenting that is required to make this determination does not exist at most hospitals and clinics. Despite these limitations, a meta-analysis of prospective studies has estimated the incidence of serious adverse drug reactions in hospitalized patients to be 6.7% and the incidence of fatal adverse drug reactions to be 0.32%. When evaluating and managing the condition of a patient who has experienced an adverse drug reaction, the physician first obtains an accurate history and performs a careful physical examination to determine whether the reaction was immunologic in nature. Drug reactions that are immunologically mediated (1) require a period of sensitization, (2) occur in a small proportion of the population, (3) are elicited at drug doses far below the therapeutic range, and (4) subside after drug discontinuation in most instances. All possible culprit drugs should be identified, with dosages and dates of administration and discontinuation, and the patient should be asked about any previous drug exposure history. Although immunodiagnostic tests for allergic drug reactions are limited, several tests do exist and may be useful in the identification of drug-specific antibodies, drug specific T lymphocytes, or mediators from activated cells. If the reaction was not consistent with an IgE-mediated event and if it did not involve serious organ damage, cautious rechallenge may be considered. For those reactions that appear to be IgE-mediated and for which there is no reliable skin test reagent, drug desensitization may be performed by allergists who are trained in this procedure. PMID- 10856173 TI - On the evolution of handedness: a speculative analysis of Darwin's views and a review of early studies of handedness in "the nearest allies of man". AB - Scientists today who seek clues into the evolutionary origins of human handedness make extensive use of evidence from comparative studies, that is, studies that ask whether handedness occurs in other species, especially apes and monkeys, as the Darwinian principle of continuity would seem to imply, or whether it is uniquely human. Early investigations had the same goal and drew on much the same kind of evidence. In this article, I review studies of animal handedness in the period before 1859, when Darwin published On the Origin of Species, and afterward, through the 1st decade of the 20th century. Inasmuch as Darwin's published writings contain hardly any statements about handedness and none at all about its evolution and continuity across species, I also speculate about what Darwin himself might have said on the subject. To do this, I draw on his statements on related matters, such as the form and structure of the hand and the transition from a quadrupedal to bipedal stance, on other writers' reports and opinions about handedness with which he was familiar or likely to have been familiar, and finally, on clues from his own and only statement about animal handedness in an unpublished letter. I conclude by asking whether and how early investigators, lacking any statement by Darwin on the evolution of handedness, invoked his theory of evolution and his views on related matters in the interpretation of their findings. PMID- 10856174 TI - Comparative neuropsychology of the dual brain: a stroll through animals' left and right perceptual worlds. AB - Perceptual asymmetries in humans typically manifest themselves under quite unnatural settings (e.g., tachistoscopic viewing and dichotic listening) and this has put into question their real biological significance. In animals with laterally placed eyes, however, perceptual asymmetries are ubiquitous in the normal, everyday behavior, as revealed by the differential use of the lateral visual field of the left and right eye in a variety of tasks. Data are presented showing how preferential use of the left and right eyes influences visual discrimination learning and detour behavior in chicks; similarities with detour tests performed in fish and evidence for asymmetries in eye use in animals with larger binocular overlap (e.g., anuran amphibians) are discussed. Implications of these perceptual asymmetries on the formation and fate of memory traces are put forward, with examples from unihemispheric sleep and lateralization of spatial memory in chicks. Finally, speculations about the evolutionary origins and possible adaptive advantages of perceptual asymmetries in vertebrates are presented. PMID- 10856175 TI - Motor control by vision and the evolution of cerebral lateralization. AB - Chicks (4 or 5 days old), which are able to use either eye freely, use the right eye (RE) preferentially in approach to a food dish when a lid, which has to be removed, is visible during approach. They use the left eye (LE) instead when no manipulation is required, but the same dish is similarly visible. The RE is also used preferentially in selecting food grains scattered over the floor; RE use in these two contexts is thus associated with visual control which brings the bill in planned contact with a visible target rather than with approach to a site where it is anticipated that feeding will occur. Zebrafish also use the RE preferentially when preparing to bite a target; during purely visual examination of the same target, this preference disappears. This evidence is used together with evolutionary evidence to support a new hypothesis for the origin of cerebral lateralization: paired anterior eyes evolved in filter-feeding ancestors of the vertebrates as part of the acquisition of prey catching. A key use for early vision was to predict likely contact with prey so as to inhibit reflexes of rejection and avoidance normally elicitated by tactile input to the mouth and so to allow ingestion. Innervation of mouth structures by the left side of the CNS caused control of mouth reflexes to become predominantly a left CNS affair. As visual abilities developed this starting condition meant that control of manipulation (which is by the mouth for most vertebrates) remained predominantly with the left side of the CNS. PMID- 10856176 TI - Evolution of hemispheric specialization: advantages and disadvantages. AB - Lateralization of the brain appeared early in evolution and many of its features appear to have been retained, possibly even in humans. We now have a considerable amount of information on the different forms of lateralization in a number of species, and the commonalities of these are discussed, but there has been relatively little investigation of the advantages of being lateralized. This article reports new findings on the differences between lateralized and nonlateralized chicks. The lateralized chicks were exposed to light for 24 h on day 19 of incubation, a treatment known to lead to lateralization of a number of visually guided responses, and the nonlateralized chicks were incubated in the dark. When they were feeding, the lateralized chicks were found to detect a stimulus resembling a raptor with shorter latency than nonlateralized chicks. This difference was not a nonspecific effect caused by the light-exposed chicks being more distressed by the stimulus. Instead, it appears to be a genuine advantage conferred by having a lateralized brain. It is suggested that having a lateralized brain allows dual attention to the tasks of feeding (right eye and left hemisphere) and vigilance for predators (left eye and right hemisphere). Nonlateralized chicks appear to perform these dual tasks less efficiently than lateralized ones. Reference is made to other species in discussing these results. PMID- 10856177 TI - Lateralized interhemispheric transfer of color cues: evidence for dynamic coding principles of visual lateralization in pigeons. AB - Visual feature discrimination tasks in pigeons reveal a right eye/left hemisphere dominance at the population level. Anatomical studies and lesion data show this visual lateralization to be related to asymmetries of the tectofugal system, which ascends from the tectum over the n. rotundus to the forebrain. Anatomically, this system is characterized by numerous morphological and connectional asymmetries which result in a bilateral visual representation in the dominant left hemisphere and a mostly contralateral representation in the subdominant right hemisphere. Ontogenetically, visual lateralization starts with an asymmetrical embryonic position within the egg, which leads to asymmetries of light stimulation. Differences in exposure to light stimulation between the eyes result in activity differences between the ascending tectofugal pathways of the left and the right hemisphere, which are transcribed during a critical time span into morphological asymmetries. The asymmetries established after this transient period finally start to determine the lateralized processes of the visual system for the entire life span of the individual. We now can show that these anatomical lateralizations are accompanied by asymmetries of interocular transfer, which enable a faster shift of learned color cues from the dominant right to the left eye than vice versa. In summary, our data provide evidence that cerebral asymmetries are based both on "static" anatomical and on "dynamic" process dependent principles. PMID- 10856178 TI - Evolution proposes and ontogeny disposes. AB - Genes, the basic building blocks of evolution, are highly conserved. For example, the mouse and human have approximately the same number of genes, and around 94% are identical in the two species. Since species differ on multiple dimensions (e.g., anatomy, physiology, and behavior), it follows that identical genes may subserve different functions in different species. Two reasons for this are gene gene interaction and gene-environment interaction (and it is the presence of these interactions which prevents one from making deterministic statements about genetics, thus rendering obsolete the nature-nurture controversy). Behavioral examples of both types of interactions are presented, including studies showing that (1) the uterine environment enhances later cognitive competence, (2) early postnatal experiences affect learning and emotionality and can extend into future generations, (3) maternal behavior changes the offspring's later behavior and physiology, and (4) knocking out one gene results in an animal less competent in one learning process but more competent in a complementary learning process. PMID- 10856179 TI - The neurodevelopmental frontostriatal disorders: evolutionary adaptiveness and anomalous lateralization. AB - The frontostriatal system (dorsolateral prefrontal cortex, lateral orbitofrontal cortex, anterior cingulate, supplementary motor area, and associated basal ganglia structures) is subject to a range of neurodevelopmental disorders: Tourette's syndrome (TS), obsessive compulsive disorder (OCD), attention deficit hyperactivity disorder (ADHD), schizophrenia (SCZ), autism, and probably depression. The system is responsible for our adaptive responses (initiation, execution, or withholding) to environmental situations, and the above disorders, involving effectively excessive release or withholding of various types of response, are all a consequence of changes in specific frontostriatal regions. The disorders all have a genetic component, and their persistence in the genome indicates that their clinical manifestations may also be associated, perhaps in low levels in close relatives, with certain adaptive advantages in given situations. Thus autism is associated with computational careers, depression with literary creativity, SCZ with lateral thinking and the Odyssean personality, ADHD with an Ice-Age readiness to respond, OCD with a focused range of interests, and TS with competitive sports and jazz improvisation. The disorders are all highly comorbid, and which one predominantly manifests may depend on how the frontostriatal system happens to be compromised as a result of inherited genetic predispositions and environmental contingency. We review the adaptive nature of the various subclinical manifestations and the evidence for concomitant phenomena (possibly epiphenomena): alterations in structural, functional, and behavioral lateralization in each syndrome. Indeed it is not clear that altered lateralization in frontostriatal disorders of a neurodevelopmental origin generally has any adaptive significance; it may often simply serve as a marker for altered regulatory function of the frontostriatal system, alterations which in low genetic dosage or penetrance continue to play an adaptive role in clinically unaffected close relatives of probands, but which, in high dosage or penetrance in the probands themselves, are generally deleterious. PMID- 10856180 TI - Health surveillance in the population living in a methyl mercury-polluted area over a long period. AB - It is important to follow up on the health status of inhabitants living in the methyl mercury-polluted area surrounding Minamata City, paying particular attention to diseases not only of the central nervous system but also of other organs. We have been carrying out such concentric studies for more than 10 years. We have previously studied the cause-specific standard mortality ratios in Minamata disease patients and reported that the SMRs for liver disease and renal disease were significantly raised in male and female patients, respectively. It was also found that complications arising from diabetes could be due to the large number of old people among the autopsy cases. The next step was to clarify the actual prevalence and incidence of liver disease, renal disease, and diabetes mellitus epidemiologically among the population in this area. The aim of this study was to determine the actual prevalence of these diseases and complaints, and to investigate the contribution of various risk factors to these diseases in this area. The study was a population-based cross-sectional mass screening survey. A case-control study was designed to estimate the role of various risk factors including methyl mercury exposure for these diseases. A mass multiple health examination survey was performed in 1500 subjects aged 40 years and older in Tsunagi Town, neighboring Minamata City, every summer since 1984. Tsunagi Town is located in a methyl mercury-polluted area and there are 36.9 certified Minamata disease patients per 1000 population. Data concerning liver disease, renal disease, and diabetes mellitus were collected on the basis of urine, hematological, physical, and ultrasonographic examinations. Data on risk factors and subjective complaints were collected by interview and other measures. The prevalence of these diseases was not higher in this methyl mercury-polluted area compared with other areas in Japan, contrary to what was expected based on standard mortality ratios and pathological findings. There were no positive correlations between those diseases and methyl mercury exposure. On the other hand, the population in the polluted area had more and a greater variety of complaints than those in the nonpolluted area. It is possible that not only neurological subjective complaints but also nonspecific complaints of the population in the polluted area might be influenced by past methyl mercury exposure. This health surveillance in the population living in a methyl mercury polluted area must be maintained in the future. PMID- 10856181 TI - Mercury distribution in the neonatal and adult cerebellum after mercury vapor exposure of pregnant squirrel monkeys. AB - The objectives of the study were (1) to map the detailed localization of mercury in the monkey cerebellum after mercury vapour exposure; (2) to investigate whether there is any difference in mercury distribution between neonatal and adult cerebellum after mercury vapor exposure; (3) to investigate the ability of mercury to accumulate in the cerebellum years after the end of exposure. Pregnant squirrel monkeys were exposed 5 days/week to mercury vapor at a concentration of 0.5 mg Hg/m(3) air 4 or 7 h/day or 1 mg Hg/m(3) air for 4 or 7 h/day. Mercury concentration in the offspring and maternal brains was examined by cold vapor, flameless atomic absorption spectrophotometry. Mercury distribution was examined by processing cerebellar sections for autometallographic (AMG) silver enhancement. Mercury concentration in the offspring cerebral occipital pole ranged between 0.20 and 0.70 microg Hg/g tissue, and in the maternal between 0.80 and 2.58 microg/Hg tissue in animals killed immediately after the end of exposure. AMG revealed that the external granule cell layer of offspring cerebellar tissue contained small amounts of mercury. The molecular layer contained mercury in some of the mercury-exposed monkeys. In the Purkinje cell layer, the Bergmann glial cells together with the Purkinje cells contained mercury. The granule cells and the Golgi cells contained small amounts of mercury. The astrocytes of the medullary layer, identified by immunohistochemistry, contained considerable amounts of mercury, but the cerebellar nuclei accumulated the highest amounts of mercury. No correlation was found between cellular accumulation and maturity of the brain; that is, the cellular localization of mercury did not differ between adult and neonatal brain, except for the amount of visualized mercury. This pattern corresponded well to the mercury concentrations found in the cerebral occipital pole. The differences found in mercury accumulation were instead considered to be dose-related. The results demonstrate that the distribution of mercury in the cerebellum after mercury vapor exposure is similar to the distribution pattern obtained after methyl mercury exposure and that mercury is trapped in the cerebellum over a long period of time. PMID- 10856182 TI - Mercury distribution in the squirrel monkey retina after in Utero exposure to mercury vapor. AB - Pregnant squirrel monkeys were exposed to mercury vapor during approximately 2/3 of a pregnancy, at a concentration of 0.5 or 1 mg Hg/m(3) air for 4 or 7 h a day, 5 days a week. The offspring were sacrificed at different ages (gestational week 16 to 5 years). The eyes were enucleated and horizontal sections of the retina, comprising the optic disc and the fovea, were processed for autometallographic (AMG) silver enhancement. The AMG mercury distribution was mapped using light and epipolarization microscopy. In young offspring (16-week-old fetus to 3 days old), mercury was detected mainly in the optic nerve, retinal pigment epithelium, inner plexiform layer, vessel walls, and ganglion cells. Three and a half months later, the amount of visualized mercury had decreased in all areas except for the retinal pigment epithelium. In adult monkeys that had survived for 2 to 5 years, only a faint AMG staining was seen in the retinal pigment epithelium, the optic nerve, and in some vessel walls. In conclusion, in offspring sacrificed in utero or shortly after birth, the structures accumulating mercury were the same as those which accumulate mercury following direct exposure through the lungs, as reported previously (K. Warfvinge and A. Bruun, 1996, Toxicology 107, 189-200), although the amount of AMG staining was less in transplacental animals. This demonstrates that inorganic mercury penetrates the blood-retina barrier. In monkeys that had survived 3 to 5 years, only tiny amounts of mercury were detected, which is in contrast to findings from direct exposure, in which large amounts were still found 3 years after exposure. This may suggest that the elimination process in the retina is more efficient in young animals, but a possible adverse effect of mercury on retinal development cannot be ruled out. PMID- 10856183 TI - A program for the control of indoor pollution by metallic mercury. AB - The objective of this study was to develop an educational program for preventing metallic mercury emissions due to the burning of mercury-gold amalgams inside houses. The main participants were adolescents from a school in the city of Pocone, State of Mato Grosso. The program was developed in five stages, including discussions of the methods and exhibition of slides showing people working in activities including the garimpos, planning activities as dramatizations, making posters and preparing a screenplay for the production of a video, discussing how to prevent indoor burning of gold-mercury amalgams, and a final evaluation of the adolescents about what they had learned during the program. The evaluation of the impact was done through a comparison of correct answers from a questionnaire before and after the development of the educational activities and by means of a comparison of urinary mercury in school students and a group of residents (women) at three different times: before the program (2.30 microgram/L), 6 months after (2.90 microgram/L), and 11 months after (1.49 microgram/L). PMID- 10856184 TI - Mercury contamination in fish from Santarem, Para, brazil. AB - This paper reports on total mercury concentrations in edible tissue from 11 fish species caught in the Municipality of Santarem, Tapajos River Basin, Para State, which are most consumed by the local population and investigates the influence of the distance between the goldmining areas and Santarem city on fish contamination by mercury. It was found that the carnivorous species reached an average of 222.1 ng.g(-1) (n=69), higher than the herbivorous species with 31.9 ng.g(-1) (n=30) and the omnivorous species with 68.7 ng.g(-1) (n=10). Significant relationships are found between fish weight and total mercury concentrations by using descriptive statistical and regression analysis for the two species, the carnivorous Pellona sp. (Sarda, r=0.73) and Pseudoplatystoma sp. (Surubim, r=0.63). PMID- 10856185 TI - Mercury intolerance in relation to superoxide dismutase, glutathione peroxidase, catalase, and the nitroblue tetrazolium responses. AB - Through percutaneous provocation with metallic mercury and phenyl mercuric acetate in patients stating the presence of subjective psychosomatic symptoms following dental amalgam treatment, it has been possible to categorize and score two extreme groups of patients, mercury-intolerant and mercury-tolerant patients reacting and not reacting, respectively, to low doses of mercury. The intolerant patients had a high psychosomatic score and the tolerant patients had a low or null score when exposed to low doses of the two mercury compounds. Determination of the scavenger enzymes superoxide dismutase, glutathione peroxidase, and catalase showed no significant differences between the mercury-intolerant and the mercury-tolerant patients and the controls. The activity of superoxide dismutase and the quantitative psychosomatic score elicited by either metallic mercury or phenyl mercuric acetate showed a positive correlation. On the other hand, analyses of the psychosomatic score and the areas under the curves of the nitroblue tetrazolium test response showed negative correlations. The results indicate that the oxidative metabolism and, in particular, superoxide dismutase may be perturbed in mercury-intolerant patients. PMID- 10856186 TI - Mercury biogeochemistry in the Idrija river, Slovenia, from above the mine into the Gulf of Trieste. AB - The Idrija Mine is the second largest Hg mine in the world which operated for 500 years. Mercury (Hg)-laden tailings still line the banks, and the system is a threat to the Idrija River and water bodies downstream including the Soca/Isonzo River and the Gulf of Trieste in the northern Adriatic Sea. A multidisciplinary study was conducted in June 1998 on water samples collected throughout the Idrija and Soca River systems and waters and sediments in the Gulf. Total Hg in the Idrija River increased >20-fold downstream of the mine from <3 to >60 ng liter( 1) with methyl mercury (MeHg) accounting for approximately 0.5%. Concentrations increased again downstream and into the estuary with MeHg accounting for nearly 1.5% of the total. While bacteria upstream of the mine did not contain mercury detoxification genes (mer), such genes were detected in bacteria collected downstream. Benthic macroinvertebrate diversity decreased downstream of the mine. Gulf waters near the river mouth contained up to 65 ng liter(-1) total Hg with approximately 0.05 ng liter(-1) MeHg. Gulf sediments near the river mouth contained 40 microgram g(-1) total Hg with MeHg concentrations of about 3 ng g( 1). Hg in sediment pore waters varied between 1 and 8 ng liter(-1), with MeHg accounting for up to 85%. Hg methylation and MeHg demethylation were active in Gulf sediments with highest activities near the surface. MeHg was degraded by an oxidative pathway with >97% C released from MeHg as CO(2). Hg methylation depth profiles resembled profiles of dissolved MeHg. Hg-laden waters still strongly impact the riverine, estuarine, and marine systems. Macroinvertebrates and bacteria in the Idrija River responded to Hg stress, and high Hg levels persist into the Gulf. Increases in total Hg and MeHg in the estuary demonstrate the remobilization of Hg, presumably as HgS dissolution and recycling. Gulf sediments actively produce MeHg, which enters bottom waters and presumably the marine food chain. PMID- 10856187 TI - Gender differences in meal patterns: role of self-caught fish and wild game in meat and fish diets. AB - The hypothesis that there are gender differences in consumption patterns of self caught fish and wild game in the meat and fish diet was examined for 415 people attending the Palmetto Sportsmen's Classic in Columbia, South Carolina. Women were less likely to eat most types of wild fish and game than were men, although there were no gender differences in the percentage eating beef, chicken, pork, and restaurant and store-bought fish. Similarly, women consumed significantly fewer meals of wild-caught fish and game than did men, although the number of meals of most store-bought foods did not differ. Both men and women who ate more meals of fish ate a higher percentage of wild-caught fish than either store bought or restaurant fish. People with low number of fish and meat meals ate mainly fish; people eating over 30 meals of meat and fish a month ate mainly meat. Only about 9% of those interviewed said that they changed their fish consumption patterns when they, or their spouse, were pregnant. These gender specific data on protein consumption can be used for exposure assessment and risk management decisions regarding consumption advisories for wild-caught fish and game. PMID- 10856188 TI - Linear regression models of methyl mercury exposure during prenatal and early postnatal life among riverside people along the upper Madeira river, Amazon. AB - This research is focused on prenatal and early postnatal mercury (Hg) exposure among the riverside people along the Upper Madeira river in the Amazon. Linear regression models were developed to predict the hair Hg concentration in infants. The independent variables included in the model of Group 1 (87 pairs of mothers and their infants) were the average maternal hair Hg concentration and maternal age. Group 2 (31 pairs) included maternal segmental hair Hg concentrations. For the segmental hair Hg analysis over time, it was assumed that hair grows at a rate of 11 cm per month. Thus, information on the timing of the dates of pregnancy and breast feeding from the birth history was used to cut the hair strands into segments, making them correspond to the mother's reproductive stage of life (31 pairs of mothers and their infants). Breast milk Hg concentration results were included with segmental and average maternal hair Hg concentration values (22 and 44 pairs of mothers and their infants, respectively). The models including the breast milk Hg concentration indicated that 61 and 55% of the variability of the infant hair Hg concentrations were due to the independent variables: segmental maternal hair Hg with breast milk Hg and average maternal hair Hg with breast milk Hg, respectively. The regression coefficients were in the range of 0.19 to 0.90, and P values were in the range of 0.0001 to 0.1490. Further recommendations include fish advisories to prevent critical Hg exposures during reproductive life and investigation of neurobehavioral performance of this study population. PMID- 10856189 TI - An evaluation of employee exposure to volatile organic compounds in three photocopy centers. AB - Personal and area samples from three copy centres were collected in thermal desorption tubes and analyzed using gas chromatography-mass spectrometry. Real time personal total volatile organic compounds (TVOC) were measured using a data logging photoionization detector. Fifty-four different VOCs were detected in the area samples. The maximum concentration measured was 1132.0 ppb (toluene, copy center 3, day 1). Thirty-eight VOCs were detected in the personal samples and concentrations ranged from 0.1 ppb (1,1-biphenyl, p-dichlorobenzene, propylbenzene, styrene, and tetrachloroethylene) to 689.6 ppb (toluene). Real time TVOC measurements indicated daily fluctuations in exposure, ranging from <71 to 21,300 ppb. The time-weighted average exposures for the photocopier operators on days 1 and 2 were 235 and 266 ppb and 6155 and 3683 ppb, in copy centers 2 and 3, respectively. Personal exposure measurements of individual VOCs were below accepted occupational standards and guidelines. For example, the maximum concentration was 0.3% of the permissible exposure limits (toluene, copy center 3). Exposures were highest in copy center 3; this is likely due to the presence of offset printing presses. It is concluded that photocopiers contribute a wide variety of VOCs to the indoor air of photocopy centers; however, exposures are at least 100 times below established standards. PMID- 10856190 TI - A portable x-ray fluorescence instrument for analyzing dust wipe samples for lead: evaluation with field samples. AB - Dust wipe samples collected in the field were tested by nondestructive X-ray fluorescence (XRF) followed by laboratory analysis with flame atomic absorption spectrophotometry (FAAS). Data were analyzed for precision and accuracy of measurement. Replicate samples with the XRF show high precision with an intraclass correlation coefficient (ICC) of 0.97 (P<0.0001) and an overall coefficient of variation of 11.6%. Paired comparison indicates no statistical difference (P=0.272) between XRF and FAAS analysis. Paired samples are highly correlated with an R(2) ranging between 0.89 for samples that contain paint chips and 0.93 for samples that do not contain paint chips. The ICC for absolute agreement between XRF and laboratory results was 0.95 (P<0.0001). The relative error over the concentration range of 25 to 14,200 microgram Pb is -12% (95% CI, 18 to -5). The XRF appears to be an excellent method for rapid on-site evaluation of dust wipes for clearance and risk assessment purposes, although there are indications of some confounding when paint chips are present. PMID- 10856192 TI - Environmental research, section B PMID- 10856191 TI - Utilization of breath analysis for exposure estimates of benzene associated with active smoking. AB - This study included three different experiments for benzene exposures associated with active smoking. In the first experiment, the mean exhaled breath benzene concentrations measured 1 min after an active smoke ranged from 58.1 to 81.3 microgram/m(3), depending on the commercial cigarette brand, while those measured prior to an active smoke ranged from 15.9 to 19.2 microgram/m(3). The postexposure breath concentrations were much higher than the mean breath concentrations reported by some previous studies whose exposure conditions and postsampling times were not controlled. Similar to some previous decay studies conducted for different volatile organic compounds in different microenvironments, our second experiment showed that there was a rapid fall in the breath concentration and thereafter the decrease was much slower. One compartment half-lives ranged from 30.1 to 57.8 min. Two-compartment half-lives ranged from 3.2 to 25.7 min for the first half-life and from 67 to 462 min for the second half-life. In the final repeated smoke experiment conducted with two specified time intervals, the breath concentrations showed increasing trends for both the pre- and the post exposure concentrations, with few exceptions. However, none of the changes were statistically significant at P<0.05. PMID- 10856193 TI - Imprinting of human GRB10 and its mutations in two patients with Russell-Silver syndrome. AB - Documentation of maternal uniparental disomy of chromosome 7 in 10% of patients with Russell-Silver syndrome (RSS), characterized by prenatal and postnatal growth retardation and dysmorphic features, has suggested the presence of an imprinted gene on chromosome 7 whose mutation is responsible for the RSS phenotype. Human GRB10 on chromosome 7, a homologue of the mouse imprinted gene Grb10, is a candidate, because GRB10 has a suppressive effect on growth, through its interaction with either the IGF-I receptor or the GH receptor, and two patients with RSS were shown to have a maternally derived duplication of 7p11 p13, encompassing GRB10. In the present study, we first demonstrated that the GRB10 gene is also monoallelically expressed in human fetal brain tissues and is transcribed from the maternally derived allele in somatic-cell hybrids. Hence, human GRB10 is imprinted. A mutation analysis of GRB10 in 58 unrelated patients with RSS identified, within the N-terminal domain of the protein, a P95S substitution in two patients with RSS. In these two cases, the mutant allele was inherited from the mother. The fact that monoallelic GRB10 expression was observed from the maternal allele in this study suggests but does not prove that these maternally transmitted mutant alleles contribute to the RSS phenotype. PMID- 10856194 TI - Congenital hereditary lymphedema caused by a mutation that inactivates VEGFR3 tyrosine kinase. AB - Hereditary lymphedema is a chronic swelling of limbs due to dysfunction of lymphatic vessels. An autosomal dominant, congenital form of the disease, also known as "Milroy disease," has been mapped to the telomeric part of chromosome 5q, in the region 5q34-q35. This region contains a good candidate gene for the disease, VEGFR3 (FLT4), that encodes a receptor tyrosine kinase specific for lymphatic vessels. To clarify the role of VEGFR3 in the etiology of the disease, we have analyzed a family with hereditary lymphedema. We show linkage of the disease with markers in 5q34-q35, including a VEGFR3 intragenic polymorphism, and we describe an A-->G transition that cosegregates with the disease, corresponding to a histidine-to-arginine substitution in the catalytic loop of the protein. In addition, we show, by in vitro expression, that this mutation inhibits the autophosphorylation of the receptor. Thus, defective VEGFR3 signaling seems to be the cause of congenital hereditary lymphedema linked to 5q34-q35. PMID- 10856195 TI - Pathogen-Driven Outbreaks in Forest Defoliators Revisited: Building Models from Experimental Data. AB - Models of outbreaks in forest-defoliating insects are typically built from a priori considerations and tested only with long time series of abundances. We instead present a model built from experimental data on the gypsy moth and its nuclear polyhedrosis virus, which has been extensively tested with epidemic data. These data have identified key details of the gypsy moth-virus interaction that are missing from earlier models, including seasonality in host reproduction, delays between host infection and death, and heterogeneity among hosts in their susceptibility to the virus. Allowing for these details produces models in which annual epidemics are followed by bouts of reproduction among surviving hosts and leads to quite different conclusions than earlier models. First, these models suggest that pathogen-driven outbreaks in forest defoliators occur partly because newly hatched insect larvae have higher average susceptibility than do older larvae. Second, the models show that a combination of seasonality and delays between infection and death can lead to unstable cycles in the absence of a stabilizing mechanism; these cycles, however, are stabilized by the levels of heterogeneity in susceptibility that we have observed in our experimental data. Moreover, our experimental estimates of virus transmission rates and levels of heterogeneity in susceptibility in gypsy moth populations give model dynamics that closely approximate the dynamics of real gypsy moth populations. Although we built our models from data for gypsy moth, our models are, nevertheless, quite general. Our conclusions are therefore likely to be true, not just for other defoliator-pathogen interactions, but for many host-pathogen interactions in which seasonality plays an important role. Our models thus give qualitative insight into the dynamics of host-pathogen interactions, while providing a quantitative interpretation of our gypsy moth-virus data. PMID- 10856196 TI - The Evolution of Plasticity and Nonplastic Spatial and Temporal Adaptations in the Presence of Imperfect Environmental Cues. AB - A model for the evolution of plasticity is considered in which the phenotype, undergoing stabilizing selection, is modeled as a linear function of an environmental cue correlated with the phenotypic optimum, with the coefficients z0 and z1 evolving according to standard quantitative genetic theory. In contrast to previous theoretical models, as the rate of migration between demes or the rate of cyclic fluctuations in the optimum increases, the amount of plasticity [Formula: see text] at equilibrium is shown to increase gradually, in part accounting for the effect of reduced nonplastic adaptation and reaching a maximum equal to the squared correlation between the environmental cue and the phenotypic optimum. Given that information available to the organism is limited, this bias of the expressed phenotype toward the global optimum is still optimal, however, in a certain decision-theoretic sense. When genetic variation in the plastic component of the trait is small so that spatial or temporal differentiation in plasticity is small, the effect of plasticity on nonplastic adaptation is to reduce the effects of variation in the phenotypic optimum by a factor [Formula: see text] only. Information acquisition costs and joint evolution of sensory systems are discussed. PMID- 10856197 TI - The Dispersal System of a Butterfly: A Test of Source-Sink Theory Suggests the Intermediate-Scale Hypothesis. AB - Theory predicts source-sink dynamics can occur in species with the ideal preemptive distribution but not with the ideal free distribution. Source-sink dynamics can also occur in species with passive dispersal, in which a fixed fraction of the population disperses each generation. However, in nature, dispersal often approximates random diffusion rather than ideal choices or fixed probabilities. Here, I ask which dispersal system occurred in a butterfly (Euphydryas editha) known to have source-sink dynamics. The study used 13 experimental sites, where vacant and occupied habitat patches were juxtaposed. I estimated movement during the flight season and tested hypotheses about the type of dispersal system. Ideal free and ideal preemptive models were rejected because per capita movement rates were density independent. Passive dispersal was rejected because per capita rates were related to patch area and habitat preference. The diffusion model best explained the data because it predicted both the area relationship and an odd feature of the habitat preference: immigration was not higher in preferred habitat; rather, emigration was lower. The diffusion model implied that source-sink dynamics were driven by diffusion from areas of high to low population density. Existing source-sink theory assumes fine-scale patchiness, in which animals have perfect knowledge and ease of mobility. The results from the butterfly suggest that source-sink dynamics arise at coarser spatial scales, where diffusion models apply. PMID- 10856198 TI - Exploring the Phylogenetic Structure of Ecological Communities: An Example for Rain Forest Trees. AB - Because of the correlation expected between the phylogenetic relatedness of two taxa and their net ecological similarity, a measure of the overall phylogenetic relatedness of a community of interacting organisms can be used to investigate the contemporary ecological processes that structure community composition. I describe two indices that use the number of nodes that separate taxa on a phylogeny as a measure of their phylogenetic relatedness. As an example of the use of these indices in community analysis, I compared the mean observed net relatedness of trees (>=10 cm diameter at breast height) in each of 28 plots (each 0.16 ha) in a Bornean rain forest with the net relatedness expected if species were drawn randomly from the species pool (of the 324 species in the 28 plots), using a supertree that I assembled from published sources. I found that the species in plots were more phylogenetically related than expected by chance, a result that was insensitive to various modifications to the basic methodology. I tentatively infer that variation in habitat among plots causes ecologically more similar species to co-occur within plots. Finally, I suggest a range of applications for phylogenetic relatedness measures in community analysis. PMID- 10856199 TI - Hot Spots, Cold Spots, and the Geographic Mosaic Theory of Coevolution. AB - Species interactions commonly coevolve as complex geographic mosaics of populations shaped by differences in local selection and gene flow. We use a haploid matching-alleles model for coevolution to evaluate how a pair of species coevolves when fitness interactions are reciprocal in some locations ("hot spots") but not in others ("cold spots"). Our analyses consider mutualistic and antagonistic interspecific interactions and a variety of gene flow patterns between hot and cold spots. We found that hot and cold spots together with gene flow influence coevolutionary dynamics in four important ways. First, hot spots need not be ubiquitous to have a global influence on evolution, although rare hot spots will not have a disproportionate impact unless selection is relatively strong there. Second, asymmetries in gene flow can influence local adaptation, sometimes creating stable equilibria at which species experience minimal fitness in hot spots and maximal fitness in cold spots, or vice versa. Third, asymmetries in gene flow are no more important than asymmetries in population regulation for determining the maintenance of local polymorphisms through coevolution. Fourth, intraspecific allele frequency differences among hot and cold spot populations evolve under some, but not all, conditions. That is, selection mosaics are indeed capable of producing spatially variable coevolutionary outcomes across the landscapes over which species interact. Altogether, our analyses indicate that coevolutionary trajectories can be strongly shaped by the geographic distribution of coevolutionary hot and cold spots, and by the pattern of gene flow among populations. PMID- 10856200 TI - Optimal Egg Size in Marine Invertebrates: Theory and Phylogenetic Analysis of the Critical Relationship between Egg Size and Development Time in Echinoids. AB - Life-history models for marine invertebrate larvae generally predict a dichotomy in egg size in different species: eggs should be either minimal in size or large enough to support development fully without larval feeding. This prediction is contradicted, however, by the empirical observation of wide, continuous variation in egg size between these extremes. The prediction of dichotomy rests on the assumption of a negative linear relationship between egg size and development time. Here, I present a simple model in which development time is inversely proportional to egg size. Incorporating this relationship into an optimality model produces predictions of intermediate rather than extreme egg size. Modeled variations in mortality, food availability, fertilization rates, and temperature all produce continuous shifts in the value of the intermediate optimal size, in direct contrast to those produced by previous models, which predict shifts between two extreme optima. Empirical data on echinoid egg size and development time strongly support the model's assumption of an inverse proportional relationship between egg size and development time. A composite phylogeny is constructed of the 37 species for which egg size, development time, water temperature, and phylogenetic relatedness are known. Independent contrasts are made of the evolutionary changes in egg size and development time. This analysis indicates that evolutionary shifts in development time are correlated with the inversely proportional shifts in egg size assumed in the model. The assumption of a negative linear relationship used in previous models is rejected. This model provides a potential explanation for intraspecific variation in egg size along environmental gradients, sympatric differences in egg size among species, and biogeographic trends in egg size and development mode across taxa. PMID- 10856201 TI - Costs and Benefits of Subadult Plumage in Mute Swans: Testing Hypotheses for the Evolution of Delayed Plumage Maturation. AB - In some avian species, young birds capable of reproducing diminish their prospects of doing so by molting into a subadult plumage that accurately signals their subadult status. Several hypotheses have been proposed to explain the evolution of delayed plumage maturation, but testing them usually has involved interspecific comparisons that are hard to interpret. Mute swans (Cygnus olor) exhibit two phenotypes that differ in whether the birds have a gray subadult plumage (SAP phenotype) or molt immediately into an all white adult plumage (AP phenotype). The AP phenotype results from a recessive allele on the X chromosome; both phenotypes occur in the same population and even in the same brood. We compared costs and benefits of both phenotypes in mute swans on the Chesapeake Bay in 1972-1980 and on Long Island Sound in 1982-1989. Swans with the SAP phenotype had higher survival rates from hatching to fledging than AP swans. In the fall, when AP cygnets began to molt into their white plumage, their parents often attacked and drove them off while allowing SAP cygnets from the same brood to remain on their territories for several more months. SAP males had higher survival rates during their first 2 yr of life than AP males, but AP swans bred at a younger age than SAP swans. The only proposed hypothesis for the evolution of delayed plumage maturation that can explain its occurrence in mute swans is the status-signaling hypothesis. This hypothesis argues that males with subadult plumage honestly advertise their age and subordinate status while AP swans are cheaters and engaging in dishonest communication. SAP males acquire a longer period of parental care, suffer less aggression from older birds, and increase their survival but forgo the opportunity to breed at an early age. This is a unique example of how a single gene resulted in either honest or dishonest communication, changed a bird's relationship with its parents and potential mates, and altered the bird's chances to survive and to reproduce. PMID- 10856202 TI - The Zoogeography of Mammalian Basal Metabolic Rate. AB - Zoogeographical effects on the basal metabolic rate (BMR) of 487 mammal species were analyzed using conventional and phylogenetically independent ANCOVA. Minimal BMR variance occurred at a "constrained body mass" of 358 g, whereas maximum variance occurred at the smallest and largest body masses. Significant differences in BMR were identified for similar-sized mammals from the six terrestrial zoogeographical zones (Afrotropical, Australasian, Indomalayan, Nearctic, Neotropical, and Palearctic). Nearctic and Palearctic mammals had higher basal rates than their Afrotropical, Australasian, Indomalayan, and Neotropical counterparts. Desert mammals had lower basal rates than mesic mammals. The patterns were interpreted with a conceptual model describing geographical BMR variance in terms of the influence of latitudinal and zonal climate variability. Low and high basal rates were explained in unpredictable and predictable environments, respectively, especially in small mammals. The BMR of large mammals may be influenced in addition by mobility and predation constraints. Highly mobile mammals tend to have high BMRs that may somehow facilitate fast running speeds, whereas less mobile mammals are generally dietary specialists and are often armored. The model thus integrates physiological and ecological criteria and makes predictions concerning body size and life-history evolution, island effects, and locomotor energetics. PMID- 10856203 TI - The solar wind-magnetosphere-ionosphere system AB - The solar wind, magnetosphere, and ionosphere form a single system driven by the transfer of energy and momentum from the solar wind to the magnetosphere and ionosphere. Variations in the solar wind can lead to disruptions of space- and ground-based systems caused by enhanced currents flowing into the ionosphere and increased radiation in the near-Earth environment. The coupling between the solar wind and the magnetosphere is mediated and controlled by the magnetic field in the solar wind through the process of magnetic reconnection. Understanding of the global behavior of this system has improved markedly in the recent past from coordinated observations with a constellation of satellite and ground instruments. PMID- 10856204 TI - Status and improvements of coupled general circulation models AB - Coupled general circulation models (CGCMs) integrate our knowledge about atmospheric and oceanic circulation. Different versions of CGCMs are used to provide a better understanding of natural climate variability on interannual and decadal time scales, for extended weather forecasting, and for making seasonal climate scenario projections. They also help to reconstruct past climates, especially abrupt climate change processes. Model intercomparisons, new test data (mainly from satellites), more powerful computers, and parameterizations of atmospheric and oceanic processes have improved CGCM performance to such a degree that the model results are now used by many decision-makers, including governments. They are also fundamental for the detection and attribution of climate change. PMID- 10856205 TI - Is El Nino changing? AB - Recent advances in observational and theoretical studies of El Nino have shed light on controversies concerning the possible effect of global warming on this phenomenon over the past few decades and in the future. El Nino is now understood to be one phase of a natural mode of oscillation-La Nina is the complementary phase-that results from unstable interactions between the tropical Pacific Ocean and the atmosphere. Random disturbances maintain this neutrally stable mode, whose properties depend on the background (time-averaged) climate state. Apparent changes in the properties of El Nino could reflect the importance of random disturbances, but they could also be a consequence of decadal variations of the background state. The possibility that global warming is affecting those variations cannot be excluded. PMID- 10856206 TI - Mantle convection and plate tectonics: toward an integrated physical and chemical theory AB - Plate tectonics and convection of the solid, rocky mantle are responsible for transporting heat out of Earth. However, the physics of plate tectonics is poorly understood; other planets do not exhibit it. Recent seismic evidence for convection and mixing throughout the mantle seems at odds with the chemical composition of erupted magmas requiring the presence of several chemically distinct reservoirs within the mantle. There has been rapid progress on these two problems, with the emergence of the first self-consistent models of plate tectonics and mantle convection, along with new geochemical models that may be consistent with seismic and dynamical constraints on mantle structure. PMID- 10856207 TI - Earth's core and the geodynamo AB - Earth's magnetic field is generated by fluid motion in the liquid iron core. Details of how this occurs are now emerging from numerical simulations that achieve a self-sustaining magnetic field. Early results predict a dominant dipole field outside the core, and some models even reproduce magnetic reversals. The simulations also show how different patterns of flow can produce similar external fields. Efforts to distinguish between the various possibilities appeal to observations of the time-dependent behavior of the field. Important constraints will come from geological records of the magnetic field in the past. PMID- 10856209 TI - Negative Poisson's ratios for extreme states of matter AB - Negative Poisson's ratios are predicted for body-centered-cubic phases that likely exist in white dwarf cores and neutron star outer crusts, as well as those found for vacuumlike ion crystals, plasma dust crystals, and colloidal crystals (including certain virus crystals). The existence of this counterintuitive property, which means that a material laterally expands when stretched, is experimentally demonstrated for very low density crystals of trapped ions. At very high densities, the large predicted negative and positive Poisson's ratios might be important for understanding the asteroseismology of neutron stars and white dwarfs and the effect of stellar stresses on nuclear reaction rates. Giant Poisson's ratios are both predicted and observed for highly strained coulombic photonic crystals, suggesting possible applications of large, tunable Poisson's ratios for photonic crystal devices. PMID- 10856208 TI - Gene targeting by homologous recombination in Drosophila. AB - Drosophila offers many advantages as an experimental organism. However, in comparison with yeast and mouse, two other widely used eukaryotic model systems, Drosophila suffers from an inability to perform homologous recombination between introduced DNA and the corresponding chromosomal loci. The ability to specifically modify the genomes of yeast and mouse provides a quick and easy way to generate or rescue mutations in genes for which a DNA clone or sequence is available. A method is described that enables analogous manipulations of the Drosophila genome. This technique may also be applicable to other organisms for which gene-targeting procedures do not yet exist. PMID- 10856210 TI - Viscosity mechanisms in accretion disks AB - The self-sustained turbulence that develops in magnetized accretion disks is suppressed in the weakly ionized, quiescent disks of close binary stars. Because accretion still proceeds during quiescence, another viscosity mechanism operates in these systems. An anticorrelation of the recurrence times of SU UMa dwarf novae with their mass ratio supports spiral waves or shockwaves tidally induced by the companion star as the main process responsible for accretion in the quiescent disks. Other weakly ionized gaseous disks in systems lacking a massive companion must rely on yet another transport mechanism, or they could be essentially passive. PMID- 10856211 TI - Step-by-step engineered multiparticle entanglement AB - After quantum particles have interacted, they generally remain in an entangled state and are correlated at a distance by quantum-mechanical links that can be used to transmit and process information in nonclassical ways. This implies programmable sequences of operations to generate and analyze the entanglement of complex systems. We have demonstrated such a procedure for two atoms and a single photon cavity mode, engineering and analyzing a three-particle entangled state by a succession of controlled steps that address the particles individually. This entangling procedure can, in principle, operate on larger numbers of particles, opening new perspectives for fundamental tests of quantum theory. PMID- 10856212 TI - Assessment of oceanic productivity with the triple-isotope composition of dissolved oxygen. AB - Plant production in the sea is a primary mechanism of global oxygen formation and carbon fixation. For this reason, and also because the ocean is a major sink for fossil fuel carbon dioxide, much attention has been given to estimating marine primary production. Here, we describe an approach for estimating production of photosynthetic oxygen, based on the isotopic composition of dissolved oxygen of seawater. This method allows the estimation of integrated oceanic productivity on a time scale of weeks. PMID- 10856213 TI - A low-operating-temperature solid oxide fuel cell in hydrocarbon-Air mixtures AB - The performance of a single-chamber solid oxide fuel cell was studied using a ceria-based solid electrolyte at temperatures below 773 kelvin. Electromotive forces of approximately 900 millivolts were generated from the cell in a flowing mixture of ethane or propane and air, where the solid electrolyte functioned as a purely ionic conductor. The electrode-reaction resistance was negligibly small in the total internal resistances of the cell. The resulting peak power density reached 403 and 101 milliwatts per square centimeter at 773 and 623 kelvin, respectively. PMID- 10856214 TI - Discovery of a basaltic asteroid in the outer main belt AB - Visible and near-infrared spectroscopic observations of the asteroid 1459 Magnya indicate that it has a basaltic surface. Magnya is at 3. 15 astronomical units (AU) from the sun and has no known dynamical link to any family, to any nearby large asteroid, or to asteroid 4 Vesta at 2.36 AU, which is the only other known large basaltic asteroid. We show that the region of the belt around Magnya is densely filled by mean-motion resonances, generating slow orbital diffusion processes and providing a potential mechanism for removing other basaltic fragments that may have been created on the same parent body as Magnya. Magnya may represent a rare surviving fragment from a larger, differentiated planetesimal that was disrupted long ago. PMID- 10856215 TI - Hierarchical self-assembly of F-actin and cationic lipid complexes: stacked three layer tubule networks. AB - We describe a distinct type of spontaneous hierarchical self-assembly of cytoskeletal filamentous actin (F-actin), a highly charged polyelectrolyte, and cationic lipid membranes. On the mesoscopic length scale, confocal microscopy reveals ribbonlike tubule structures that connect to form a network of tubules on the macroscopic scale (more than 100 micrometers). Within the tubules, on the 0.5 to 50-nanometer length scale, x-ray diffraction reveals an unusual structure consisting of osmotically swollen stacks of composite membranes with no direct analog in simple amphiphilic systems. The composite membrane is composed of three layers, a lipid bilayer sandwiched between two layers of actin, and is reminiscent of multilayered bacterial cell walls that exist far from equilibrium. Electron microscopy reveals that the actin layer consists of laterally locked F actin filaments forming an anisotropic two-dimensional tethered crystal that appears to be the origin of the tubule formation. PMID- 10856216 TI - Impacts of a global climate cycle on population dynamics of a migratory songbird. AB - Progress toward understanding factors that limit abundances of migratory birds, including climate change, has been difficult because these species move between diverse locations, often on different continents. For black-throated blue warblers (Dendroica caerulescens), demographic rates in both tropical winter quarters and north temperate breeding grounds varied with fluctuations in the El Nino Southern Oscillation. Adult survival and fecundity were lower in El Nino years and higher in La Nina years. Fecundity, in turn, was positively correlated with subsequent recruitment of new individuals into winter and breeding populations. These findings demonstrate that migratory birds can be affected by shifts in global climate patterns and emphasize the need to know how events throughout the annual cycle interact to determine population size. PMID- 10856217 TI - Designing small-molecule switches for protein-protein interactions. AB - Mutations introduced into human growth hormone (hGH) (Thr175 --> Gly-hGH) and the extracellular domain of the hGH receptor (Trp104 --> Gly-hGHbp) created a cavity at the protein-protein interface that resulted in binding affinity being reduced by a factor of 10(6). A small library of indole analogs was screened for small molecules that bind the cavity created by the mutations and restore binding affinity. The ligand 5-chloro-2-trichloromethylimidazole was found to increase the affinity of the mutant hormone for its receptor more than 1000-fold. Cell proliferation and JAK2 phosphorylation assays showed that the mutant hGH activates growth hormone signaling in the presence of added ligand. This approach may allow other protein-protein and protein-nucleic acid interactions to be switched on or off by the addition or depletion of exogenous small molecules. PMID- 10856218 TI - Proliferation, but not growth, blocked by conditional deletion of 40S ribosomal protein S6. AB - Because ribosome biogenesis plays an essential role in cell proliferation, control mechanisms may have evolved to recognize lesions in this critical anabolic process. To test this possibility, we conditionally deleted the gene encoding 40S ribosomal protein S6 in the liver of adult mice. Unexpectedly, livers from fasted animals deficient in S6 grew in response to nutrients even though biogenesis of 40S ribosomes was abolished. However, liver cells failed to proliferate or induce cyclin E expression after partial hepatectomy, despite formation of active cyclin D-CDK4 complexes. These results imply that abrogation of 40S ribosome biogenesis may induce a checkpoint control that prevents cell cycle progression. PMID- 10856219 TI - A single-molecule study of RNA catalysis and folding. AB - Using fluorescence microscopy, we studied the catalysis by and folding of individual Tetrahymena thermophila ribozyme molecules. The dye-labeled and surface-immobilized ribozymes used were shown to be functionally indistinguishable from the unmodified free ribozyme in solution. A reversible local folding step in which a duplex docks and undocks from the ribozyme core was observed directly in single-molecule time trajectories, allowing the determination of the rate constants and characterization of the transition state. A rarely populated docked state, not measurable by ensemble methods, was observed. In the overall folding process, intermediate folding states and multiple folding pathways were observed. In addition to observing previously established folding pathways, a pathway with an observed folding rate constant of 1 per second was discovered. These results establish single-molecule fluorescence as a powerful tool for examining RNA folding. PMID- 10856220 TI - Role of CD47 as a marker of self on red blood cells. AB - The immune system recognizes invaders as foreign because they express determinants that are absent on host cells or because they lack "markers of self" that are normally present. Here we show that CD47 (integrin-associated protein) functions as a marker of self on murine red blood cells. Red blood cells that lacked CD47 were rapidly cleared from the bloodstream by splenic red pulp macrophages. CD47 on normal red blood cells prevented this elimination by binding to the inhibitory receptor signal regulatory protein alpha (SIRPalpha). Thus, macrophages may use a number of nonspecific activating receptors and rely on the presence or absence of CD47 to distinguish self from foreign. CD47-SIRPalpha may represent a potential pathway for the control of hemolytic anemia. PMID- 10856221 TI - Selectivity for 3D shape that reveals distinct areas within macaque inferior temporal cortex. AB - The anterior part of the macaque inferior temporal cortex, area TE, occupies a large portion of the temporal lobe and is critical for object recognition. Thus far, no relation between anatomical subdivisions of TE and neuronal selectivity has been described. Here, we present evidence that neurons selective for three dimensional (3D) shape are concentrated in the lower bank of the superior temporal sulcus, whereas neurons in lateral TE are generally unselective for 3D shape, though equally selective for 2D shape. These findings reveal that TE consists of at least two distinct areas, one of which processes a specific object property. PMID- 10856222 TI - Disruption of AMPA receptor GluR2 clusters following long-term depression induction in cerebellar Purkinje neurons. AB - Cerebellar long-term depression (LTD) is thought to play an important role in certain types of motor learning. However, the molecular mechanisms underlying this event have not been clarified. Here, using cultured Purkinje cells, we show that stimulations inducing cerebellar LTD cause phosphorylation of Ser880 in the intracellular C-terminal domain of the AMPA receptor subunit GluR2. This phosphorylation is accompanied by both a reduction in the affinity of GluR2 to glutamate receptor interacting protein (GRIP), a molecule known to be critical for AMPA receptor clustering, and a significant disruption of postsynaptic GluR2 clusters. Moreover, GluR2 protein released from GRIP is shown to be internalized. These results suggest that the dissociation of postsynaptic GluR2 clusters and subsequent internalization of the receptor protein, initiated by the phosphorylation of Ser880, are the mechanisms underlying the induction of cerebellar LTD. PMID- 10856223 TI - Impaired learning with enhanced hippocampal long-term potentiation in PTPdelta deficient mice. AB - Protein tyrosine phosphatase delta (PTPdelta) is a receptor-type PTP expressed in the specialized regions of the brain including the hippocampal CA2 and CA3, B lymphocytes and thymic medulla. To elucidate the physiological roles of PTPdelta, PTPdelta-deficient mice were produced by gene targeting. It was found that PTPdelta-deficient mice were semi-lethal due to insufficient food intake. They also exhibited learning impairment in the Morris water maze, reinforced T-maze and radial arm maze tasks. Interestingly, although the histology of the hippocampus appeared normal, the magnitudes of long-term potentiation (LTP) induced at hippocampal CA1 and CA3 synapses were significantly enhanced in PTPdelta-deficient mice, with augmented paired-pulse facilitation in the CA1 region. Thus, it was shown that PTPdelta plays important roles in regulating hippocampal LTP and learning processes, and that hippocampal LTP does not necessarily positively correlate with spatial learning ability. To our knowledge, this is the first report of a specific PTP involved in the regulation of synaptic plasticity or in the processes regulating learning and memory. PMID- 10856224 TI - A mutation in the ovine cathepsin D gene causes a congenital lysosomal storage disease with profound neurodegeneration. AB - The neuronal ceroid lipofuscinoses (NCLs) constitute a group of neurodegenerative storage diseases characterized by progressive psychomotor retardation, blindness and premature death. Pathologically, there is accumulation of autofluorescent material in lysosome-derived organelles in a variety of cell types, but neurons in the central nervous system appear to be selectively affected and undergo progressive death. In this report we show that a novel form of NCL, congenital ovine NCL, is caused by a deficiency in the lysosomal aspartyl proteinase cathepsin D. A single nucleotide mutation in the cathepsin D gene results in conversion of an active site aspartate to asparagine, leading to production of an enzymatically inactive but stable protein. This results in severe cerebrocortical atrophy and early death, providing strong evidence for an important role of cathepsin D in neuronal development and/or homeostasis. PMID- 10856225 TI - Phosphatidylinositol 3,4,5-trisphosphate regulates Ca(2+) entry via btk in platelets and megakaryocytes without increasing phospholipase C activity. AB - The role of phosphatidylinositol 3,4,5-trisphosphate (PI3,4,5P(3)) and Btk in signalling by the collagen receptor glycoprotein VI was investigated. PI3,4,5P(3) was increased in platelets from mice deficient in the SH2 domain-containing inositol 5-phosphatase (SHIP), in response to collagen related peptide (CRP). Tyrosine phosphorylation and activation of phospholipase Cgamma2 (PLCgamma2) were unaltered in SHIP(-/-) platelets, whereas Btk was heavily tyrosine phosphorylated under basal conditions and maximally phosphorylated by low concentrations of CRP. There was an increase in basal Ca(2+), maximal expression of P-selectin, and potentiation of Ca(2+) and aminophospholipid exposure to CRP in SHIP(-/-) platelets in the presence of Ca(2+) (1 mM). Microinjection of PI3,4, 5P(3) into megakaryocytes caused a 3-fold increase in Ca(2+) in response to CRP, which was absent in X-linked immunodeficiency (Xid) mice, which have a mutation in the PH domain of Btk. There was a corresponding partial reduction in the sustained level of intracellular Ca(2+) in response to CRP in Xid mice but no change in PLC activity. These results demonstrate a novel pathway of Ca(2+) entry that involves PI3,4,5P(3) and Btk, and which is independent of increased PLC activity. PMID- 10856226 TI - Type 1 pilus-mediated bacterial invasion of bladder epithelial cells. AB - Most strains of uropathogenic Escherichia coli (UPEC) encode filamentous adhesive organelles called type 1 pili. We have determined that the type 1 pilus adhesin, FimH, mediates not only bacterial adherence, but also invasion of human bladder epithelial cells. In contrast, adherence mediated by another pilus adhesin, PapG, did not initiate bacterial internalization. FimH-mediated invasion required localized host actin reorganization, phosphoinositide 3-kinase (PI 3-kinase) activation and host protein tyrosine phosphorylation, but not activation of Src family tyrosine kinases. Phosphorylation of focal adhesin kinase (FAK) at Tyr397 and the formation of complexes between FAK and PI 3-kinase and between alpha actinin and vinculin were found to correlate with type 1 pilus-mediated bacterial invasion. Inhibitors that prevented bacterial invasion also blocked the formation of these complexes. Our results demonstrate that UPEC strains are not strictly extracellular pathogens and that the type 1 pilus adhesin FimH can directly trigger host cell signaling cascades that lead to bacterial internalization. PMID- 10856227 TI - Targeted mutagenesis of Plasmodium falciparum erythrocyte membrane protein 3 (PfEMP3) disrupts cytoadherence of malaria-infected red blood cells. AB - Adhesion of parasite-infected red blood cells to the vascular endothelium is a critical event in the pathogenesis of malaria caused by Plasmodium falciparum. Adherence is mediated by the variant erythrocyte membrane protein 1 (PfEMP1). Another protein, erythrocyte membrane protein-3 (PfEMP3), is deposited under the membrane of the parasite-infected erythrocyte but its function is unknown. Here we show that mutation of PfEMP3 disrupts transfer of PfEMP1 to the outside of the P.FALCIPARUM:-infected cell. Truncation of the C-terminal end of PfEMP3 by transfection prevents distribution of this large (>300 kDa) protein around the membrane but does not disrupt trafficking of the protein from the parasite to the cytoplasmic face of the erythrocyte membrane. The truncated PfEMP3 accumulates in structures that appear to be associated with the erythrocyte membrane. We show that accumulation of mutated PfEMP3 blocks the transfer of PfEMP1 onto the outside of the parasitized cell surface and suggest that these proteins traffic through an erythrocyte membrane-associated compartment that is involved in the transfer of PfEMP1 to the surface of the parasite-infected red blood cell. PMID- 10856228 TI - Sphingoid base synthesis requirement for endocytosis in Saccharomyces cerevisiae. AB - The internalization step of endocytosis in yeast requires actin and sterols for maximum efficiency. In addition, many receptors and plasma membrane proteins must be phosphorylated and ubiquitylated prior to internalization. The Saccharomyces cerevisiae end8-1 mutant is allelic to lcb1, a mutant defective in the first step of sphingoid base synthesis. Upon arrest of sphingoid base synthesis a rapid block in endocytosis is seen. This block can be overcome by exogenous sphingoid base. Under conditions where endogenous sphingosine base synthesis was blocked and exogenous sphingoid bases could not be converted to phosphorylated sphingoid bases or to ceramide, sphingoid bases could still suppress the endocytic defect. Therefore, the required lipid is most likely a sphingoid base. Interestingly, sphingoid base synthesis is required for proper actin organization, but is not required for receptor phosphorylation. This is the first case of a physiological role for sphingoid base synthesis, other than as a precursor for ceramide or phosphorylated sphingoid base synthesis. PMID- 10856229 TI - Increased protein kinase or decreased PP2A activity bypasses sphingoid base requirement in endocytosis. AB - Lipids have been implicated in signal transduction and in several stages of membrane trafficking, but these two functions have not been functionally linked. In yeast, sphingoid base synthesis is required for the internalization step of endocytosis and organization of the actin cytoskeleton. We show that inactivation of a protein phosphatase 2A (PP2A) or overexpression of one of two kinases, Yck2p or Pkc1p, can specifically suppress the sphingoid base synthesis requirement for endocytosis. The two kinases have an overlapping function because only a mutant with impaired function of both kinases is defective in endocytosis. An ultimate target of sphingoid base synthesis may be the actin cytoskeleton, because overexpression of the kinases and inactivation of PP2A substantially corrected the actin defect due to the absence of sphingoid base. These results suggest that sphingoid base controls protein phosphorylation, perhaps by activating a signal transduction pathway that is required for endocytosis and proper actin cytoskeleton organization in yeast. PMID- 10856230 TI - Zinc transporters that regulate vacuolar zinc storage in Saccharomyces cerevisiae. AB - All cells regulate their intracellular zinc levels. In yeast, zinc uptake is mediated by Zrt1p and Zrt2p, which belong to the ZIP family of metal transporters. Under zinc limitation, ZRT1 and ZRT2 transcription is induced by the Zap1p transcriptional activator. We describe here a new component of zinc homeostasis, vacuolar zinc storage, that is also regulated by Zap1p. Zinc-replete cells accumulate zinc in the vacuole via the Zrc1p and Cot1p transporters. Our results indicate that another zinc transporter, Zrt3p, mobilizes this stored zinc in zinc-limited cells. ZRT3 is a Zap1p-regulated gene whose transcription increases in low zinc. Zrt3p is also a member of the ZIP family and it localizes to the vacuolar membrane. The effects of ZRT3 mutation and overexpression on cell growth, cellular zinc accumulation and intracellular labile zinc pools are all consistent with its proposed role. Furthermore, we demonstrate that zrt3 mutants inefficiently mobilize stored zinc to offset deficiency. Thus, our studies define a system of zinc influx and efflux transporters in the vacuole that play important roles in zinc homeostasis. PMID- 10856231 TI - Peptide antagonists of the plasmodesmal macromolecular trafficking pathway. AB - In plants, cell-to-cell transport of endogenous and viral proteins and ribonucleoprotein complexes (RNPCs) occurs via plasmodesmata. Specificity of this transport pathway appears to involve interaction between such proteins/RNPCs and plasmodesmal chaperones/receptors. Here, KN1 and the cucumber mosaic virus movement protein (CMV-MP) were used, in a modified phage-display screening system, to identify peptides capable of interacting with proteins present in a plasmodesmal-enriched cell wall fraction. Binding/competition assays and microinjection experiments revealed that these phage-displayed peptides and homologous synthetic oligopeptides function as ligand-specific antagonists of macromolecular trafficking through plasmodesmata. A KN1 peptide antagonist had the capacity to interact with a motif involved in the dilation of plasmodesmal microchannels. Although KN1 could still achieve limited movement through plasmodesmata when this SEL motif was blocked, KN1-mediated transport of KN1 sense RNA was fully inhibited. These findings provide direct support for the hypothesis that KN1 requires, minimally, two physically separated signal motifs involved in the dilation of, and protein translocation through, plasmodesmal microchannels, and provide direct proof that plasmodesmal dilation is a prerequisite for the cell-to-cell transport of an RNPC. PMID- 10856232 TI - 14-3-3s regulate global cleavage of their diverse binding partners in sugar starved Arabidopsis cells. AB - Despite 14-3-3 proteins being implicated in the control of the eukaryotic cell cycle, metabolism, cell signalling and survival, little is known about the global regulation or functions of the phosphorylation-dependent binding of 14-3-3s to diverse target proteins. We identified Arabidopsis cytosolic proteins that bound 14-3-3s in competition with a 14-3-3-binding phosphopeptide, including nitrate reductase, glyceraldehyde- 3-phosphate dehydrogenase, a calcium-dependent protein kinase, sucrose-phosphate synthase (SPS) and glutamyl-tRNA synthetase. Remarkably, in cells starved of sugars or fed with non-metabolizable glucose analogues, all 14-3-3 binding was lost and the target proteins were selectively cleaved into proteolytic fragments. 14-3-3 binding reappeared after several hours of re-feeding with sugars. Starvation-induced degradation was blocked by 5-amino imidazole-4-carboxamide riboside (which is converted to an AMP-mimetic) or the protease inhibitor MG132 (Cbz-leu-leu-leucinal). Extracts of sugar-starved (but not sugar-fed) Arabidopsis cells contained an ATP-independent, MG132-sensitive, neutral protease that cleaved Arabidopsis SPS, and the mammalian 14-3-3-regulated transcription factor, FKHR. Cleavage of SPS and phosphorylated FKHR in vitro was blocked by binding to 14-3-3s. The finding that 14-3-3s participate in a nutrient sensing pathway controlling cleavage of many targets may underlie the effects of these proteins on plant development. PMID- 10856233 TI - Crystal structure of a gamma-herpesvirus cyclin-cdk complex. AB - Several gamma-herpesviruses encode proteins related to the mammalian cyclins, regulatory subunits of cyclin-dependent kinases (cdks) essential for cell cycle progression. We report a 2.5 A crystal structure of a full-length oncogenic viral cyclin from gamma-herpesvirus 68 complexed with cdk2. The viral cyclin binds cdk2 with an orientation different from cyclin A and makes several novel interactions at the interface, yet it activates cdk2 by triggering conformational changes similar to cyclin A. Sequences within the viral cyclin N-terminus lock part of the cdk2 T-loop within the core of the complex. These sequences and others are conserved amongst the viral and cellular D-type cyclins, suggesting that this structure has wider implications for other cyclin-cdk complexes. The observed resistance of this viral cyclin-cdk complex to inhibition by the p27(KIP:) cdk inhibitor is explained by sequence and conformational variation in the cyclin rendering the p27(KIP:)-binding site on the cyclin subunit non-functional. PMID- 10856234 TI - SH3 domain recognition of a proline-independent tyrosine-based RKxxYxxY motif in immune cell adaptor SKAP55. AB - Src-homology 3 (SH3) domains recognize PXXP core motif preceded or followed by positively charged residue(s). Whether SH3 domains recognize motifs other than proline-based sequences is unclear. In this study, we report SH3 domain binding to a novel proline-independent motif in immune cell adaptor SKAP55, which is comprised of two N-terminal lysine and arginine residues followed by two tyrosines (i.e. RKxxYxxY). Domains capable of binding to class I proline motifs bound to the motif, while the class II domains failed to bind. Peptide precipitation, alanine scanning and in vivo co-expression studies demonstrated a requirement for the arginine, lysine and tandem tyrosines of the motif. Two dimensional NMR analysis of the peptide bound FYN-SH3 domain showed overlap with the binding site of a proline-rich peptide on the charged surface of the SH3 domain, while resonance signals for other residues (W119, W120, Y137) were not perturbed by the RKGDYASY based peptide. Expression of the RKGDYASY peptide potently inhibited TcRzeta/CD3-mediated NF-AT transcription in T cells. Our findings extend the repertoire of SH3 domain binding motifs to include a tyrosine based motif and demonstrate a regulatory role for this motif in receptor signaling. PMID- 10856235 TI - Ras mediates the cAMP-dependent activation of extracellular signal-regulated kinases (ERKs) in melanocytes. AB - In melanocytes and melanoma cells, cAMP activates extracellular signal-regulated kinases (ERKs) and MEK-1 by an unknown mechanism. We demonstrate that B-Raf is activated by cAMP in melanocytes. A dominant-negative mutant of B-Raf, but not of Raf-1, blocked the cAMP-induced activation of ERK, indicating that B-Raf is the MEK-1 upstream regulator mediating this cAMP effect. Studies using Clostridium sordelii lethal toxin and Clostridium difficile toxin B have suggested that Rap-1 or Ras might transduce cAMP action. We show that Ras, but not Rap-1, is activated cell-specifically and mediates the cAMP-dependent activation of ERKs, while Rap-1 is not involved in this process in melanocytes. Our results suggest a novel, cell specific mechanism involving Ras small GTPase and B-Raf kinase as mediators of ERK activation by cAMP. Also, in melanocytes, Ras or ERK activation by cAMP is not mediated through protein kinase A activation. Neither the Ras exchange factor, Son of sevenless (SOS), nor the cAMP-responsive Rap-1 exchange factor, Epac, participate in the cAMP-dependent activation of Ras. These findings suggest the existence of a melanocyte-specific Ras exchange factor directly regulated by cAMP. PMID- 10856236 TI - Active ERK/MAP kinase is targeted to newly forming cell-matrix adhesions by integrin engagement and v-Src. AB - Integrin engagement generates cellular signals leading to the recruitment of structural and signalling molecules which, in concert with rearrangements of the actin cytoskeleton, leads to the formation of focal adhesion complexes. Using antisera reactive either with total ERK or with phosphorylated/activated forms of ERK, in rat embryo fibroblasts and embryonic avian cells that express v-Src, we found that active ERK is targeted to newly forming focal adhesions after integrin engagement or activation of v-Src. UO126, an inhibitor of MAP kinase kinase 1 (MEK1), suppressed focal adhesion targeting of active ERK and cell spreading. Also, integrin engagement and v-Src induced myosin light chain kinase (MLCK) dependent phosphorylation of myosin light chain downstream of the MEK/ERK pathway, and MLCK and myosin activities are required for the focal adhesion targeting of ERK. The translocation of active ERK to newly forming focal adhesions may direct specificity towards appropriate downstream targets that influence adhesion assembly. These findings support a role for ERK in the regulation of the adhesion/cytoskeletal network and provide an explanation for the role of ERK in cell motility. PMID- 10856237 TI - A phosphoserine-regulated docking site in the protein kinase RSK2 that recruits and activates PDK1. AB - The 90 kDa ribosomal S6 kinase-2 (RSK2) is a growth factor-stimulated protein kinase with two kinase domains. The C-terminal kinase of RSK2 is activated by ERK type MAP kinases, leading to autophosphorylation of RSK2 at Ser386 in a hydrophobic motif. The N-terminal kinase is activated by 3-phosphoinositide dependent protein kinase-1 (PDK1) through phosphorylation of Ser227, and phosphorylates the substrates of RSK. Here, we identify Ser386 in the hydrophobic motif of RSK2 as a phosphorylation-dependent docking site and activator of PDK1. Treatment of cells with growth factor induced recruitment of PDK1 to the Ser386 phosphorylated hydrophobic motif and phosphorylation of RSK2 at Ser227. A RSK2 S386K mutant showed no interaction with PDK1 or phosphorylation at Ser227. Interaction with Ser386-phosphorylated RSK2 induced autophosphorylation of PDK1. Addition of a synthetic phosphoSer386 peptide (RSK2(373-396)) increased PDK1 activity 6-fold in vitro. Finally, mutants of RSK2 and MSK1, a RSK-related kinase, with increased affinity for PDK1, were constitutively active in vivo and phosphorylated histone H3. Our results suggest a novel regulatory mechanism based on phosphoserine-mediated recruitment of PDK1 to RSK2, leading to coordinated phosphorylation and activation of PDK1 and RSK2. PMID- 10856238 TI - Spatial and temporal regulation of protein kinase D (PKD). AB - Protein kinase D (PKD; also known as PKCmicro) is a serine/threonine kinase activated by diacylglycerol signalling pathways in a variety of cells. PKD has been described previously as Golgi-localized, but herein we show that it is present within the cytosol of quiescent B cells and mast cells and moves rapidly to the plasma membrane after antigen receptor triggering. The membrane redistribution of PKD requires the diacylglycerol-binding domain of the enzyme, but is independent of its catalytic activity and does not require the integrity of the pleckstrin homology domain. Antigen receptor signalling initiates in glycosphingolipid-enriched microdomains, but membrane-associated PKD does not co localize with these specialized structures. Membrane targeting of PKD is transient, the enzyme returns to the cytosol within 10 min of antigen receptor engagement. Strikingly, the membrane-recycled PKD remains active in the cytosol for several hours. The present work thus characterizes a sustained antigen receptor-induced signal transduction pathway and establishes PKD as a serine kinase that temporally and spatially disseminates antigen receptor signals away from the plasma membrane into the cytosol. PMID- 10856239 TI - Phosphorylation status of the SCR homeodomain determines its functional activity: essential role for protein phosphatase 2A,B'. AB - Sex combs reduced (SCR) is a Drosophila Hox protein that determines the identity of the labial and prothoracic segments. In search of factors that might associate with SCR to control its activity and/or specificity, we performed a yeast two hybrid screen. A Drosophila homologue of the regulatory subunit (B'/PR61) of serine-threonine protein phosphatase 2A (dPP2A,B') specifically interacted with the SCR homeodomain. The N-terminal arm within the SCR homeodomain was shown to be a target of phosphorylation/dephosphorylation by cAMP-dependent protein kinase A and protein phosphatase 2A, respectively. In vivo analyses revealed that mutant forms of SCR mimicking constitutively dephosphorylated or phosphorylated states of the homeodomain were active or inactive, respectively. Inactivity of the phosphorylated mimic form was attributed to impaired DNA binding. Specific ablation of dPP2A,B' gene activity by double-stranded RNA-mediated genetic interference resulted in embryos without salivary glands, an SCR null phenotype. Our data demonstrate an essential role for Drosophila PP2A,B' in positively modulating SCR function. PMID- 10856240 TI - The evi-1 oncoprotein inhibits c-Jun N-terminal kinase and prevents stress induced cell death. AB - Evi-1 encodes a nuclear protein involved in leukemic transformation of hematopoietic cells. Evi-1 possesses two sets of zinc finger motifs separated into two domains, and its characteristics as a transcriptional regulator have been described. Here we show that Evi-1 acts as an inhibitor of c-Jun N-terminal kinase (JNK), a class of mitogen-activated protein kinases implicated in stress responses of cells. Evi-1 physically interacts with JNK, although it does not affect its phosphorylation. This interaction is required for inhibition of JNK. Evi-1 protects cells from stress-induced cell death with dependence on the ability to inhibit JNK. These results reveal a novel function of Evi-1, which provides evidence for inhibition of JNK by a nuclear oncogene product. Evi-1 blocks cell death by selectively inhibiting JNK, thereby contributing to oncogenic transformation of cells. PMID- 10856241 TI - JunB suppresses cell proliferation by transcriptional activation of p16(INK4a) expression. AB - A role for the transcription factor JunB in proliferation control was investigated in genetically modified mouse fibroblasts. Increased JunB expression induced high levels of the cyclin-dependent kinase inhibitor p16(INK4a), leading to premature senescence in primary cells and reduced proliferation in 3T3 cells, whereas lack of JunB expression results in decreased p16 levels. Furthermore, JunB-mediated p16 induction in 3T3 cells completely abolished cyclin D-associated kinase activity, resulting in reduced pRb hyperphosphorylation and G(1)-phase extension. Moreover, three AP1-like binding sites were identified in the p16 promoter through which JunB directly activates p16 transcription. Elevated JunB expression in 3T3 cells also inhibited Ras- and Src-mediated transformation and tumour growth in vivo. The suppressive effect of JunB on cell proliferation was shown to be dependent on p16 since it did not occur in INK4a(-/-) fibroblasts that lack both p16 and p19(ARF). These results demonstrate that p16 is a direct transcriptional target gene of JunB and identify JunB as a negative regulator of cell proliferation. PMID- 10856242 TI - One enhancer mediates mafK transcriptional activation in both hematopoietic and cardiac muscle cells. AB - Members of the small Maf family of transcription factors play important roles in hematopoiesis. Using transgenic assays, we discovered a tissue-specific enhancer 3' to the mafK gene. This enhancer directs mafK transcription in hematopoietic as well as in developing cardiac muscle cells, and was thus designated the hematopoietic and cardiac enhancer of mafK (HCEK). Only two of four GATA consensus motifs identified within HCEK contributed to enhancer activity, and both of these sites were required for both cardiac and hematopoietic transcriptional activation. The expression profile of MafK significantly overlapped that of GATA-1 in hematopoietic cells and of GATA-4/-6 in cardiac tissues. Each of these GATA factors bound with high specificity to both of the critical GATA sites in HCEK. Hence, the mafK gene is regulated by different GATA proteins in the hematopoietic and cardiac compartments through the same two GATA binding sites in HCEK. These data provide the first in vivo demonstration that distinct members of a related transcription factor family activate the tissue specific expression of a single target gene using the same cis-regulatory element. PMID- 10856243 TI - A distal Schwann cell-specific enhancer mediates axonal regulation of the Oct-6 transcription factor during peripheral nerve development and regeneration. AB - The POU domain transcription factor Oct-6 is a major regulator of Schwann cell differentiation and myelination. During nerve development and regeneration, expression of Oct-6 is under the control of axonal signals. Identification of the cis-acting elements necessary for Oct-6 gene regulation is an important step in deciphering the complex signalling between Schwann cells and axons governing myelination. Here we show that a fragment distal to the Oct-6 gene, containing two DNase I-hypersensitive sites, acts as the Oct-6 Schwann cell-specific enhancer (SCE). The SCE is sufficient to drive spatially and temporally correct expression, during both normal peripheral nerve development and regeneration. We further demonstrate that a tagged version of Oct-6, driven by the SCE, rescues the peripheral nerve phenotype of Oct-6-deficient mice. Thus, our isolation and characterization of the Oct-6 SCE provides the first description of a cis-acting genetic element that responds to converging signalling pathways to drive myelination in the peripheral nervous system. PMID- 10856244 TI - Structural basis for the heterodimeric interaction between the acute leukaemia associated transcription factors AML1 and CBFbeta. AB - Mutations in the genes encoding the interacting proteins AML1 and CBFbeta are the most common genetic abnormalities in acute leukaemia, and congenital mutations in the related AML3 gene are associated with disorders of osteogenesis. Furthermore, the interaction of AML1 with CBFbeta is essential for haematopoiesis. We report the 2.6 A resolution crystal structure of the complex between the AML1 Runt domain and CBFbeta, which represents a paradigm for the mode of interaction of this highly conserved family of transcription factors. The structure demonstrates that point mutations associated with cleidocranial dysplasia map to the conserved heterodimer interface, suggesting a role for CBFbeta in osteogenesis, and reveals a potential protein interaction platform composed of conserved negatively charged residues on the surface of CBFbeta. PMID- 10856245 TI - Structure of the C-terminal domain of Tup1, a corepressor of transcription in yeast. AB - The Tup1-Ssn6 corepressor complex regulates the expression of several sets of genes, including genes that specify mating type in the yeast Saccharomyces cerevisiae. Repression of mating-type genes occurs when Tup1-Ssn6 is brought to the DNA by the Matalpha2 DNA-binding protein and assembled upstream of a- and haploid-specific genes. We have determined the 2.3 A X-ray crystal structure of the C-terminal domain of Tup1 (accesion No. 1ERJ), a 43 kDa fragment that contains seven copies of the WD40 sequence motif and binds to the Matalpha2 protein. Moreover, this portion of the protein can partially substitute for full length Tup1 in bringing about transcriptional repression. The structure reveals a seven-bladed beta propeller with an N-terminal subdomain that is anchored to the side of the propeller and extends the beta sheet of one of the blades. Point mutations in Tup1 that specifically affect the Tup1-Matalpha2 interaction cluster on one surface of the propeller. We identified regions of Tup1 that are conserved among the fungal Tup1 homologs and may be important in protein-protein interactions with additional components of the Tup1-mediated repression pathways. PMID- 10856246 TI - sigma factor selectivity of Escherichia coli RNA polymerase: role for CRP, IHF and lrp transcription factors. AB - osmY is a stationary phase-induced and osmotically regulated gene in Escherichia coli that requires the stationary phase RNA polymerase (Esigma(S)) for in vivo expression. We show here that the major RNA polymerase, Esigma(70), also transcribes osmY in vitro and, depending on genetic background, even in vivo. The cAMP receptor protein (CRP) bound to cAMP, the leucine-responsive regulatory protein (Lrp) and the integration host factor (IHF) inhibit transcription initiation at the osmY promoter. The binding site for CRP is centred at -12.5 from the transcription start site, whereas Lrp covers the whole promoter region. The site for IHF maps in the -90 region. By mobility shift assay, permanganate reactivity and in vitro transcription experiments, we show that repression is much stronger with Esigma(70) than with Esigma(S) holoenzyme. We conclude that CRP, Lrp and IHF inhibit open complex formation more efficiently with Esigma(70) than with Esigma(S). This different ability of the two holoenzymes to interact productively with promoters once assembled in complex nucleoprotein structures may be a crucial factor in generating sigma(S) selectivity in vivo. PMID- 10856247 TI - Conservation of sigma-core RNA polymerase proximity relationships between the enhancer-independent and enhancer-dependent sigma classes. AB - Two distinct classes of RNA polymerase sigma factors (sigma) exist in bacteria and are largely unrelated in primary amino acid sequence and their modes of transcription activation. Using tethered iron chelate (Fe-BABE) derivatives of the enhancer-dependent sigma(54), we mapped several sites of proximity to the beta and beta' subunits of the core RNA polymerase. Remarkably, most sites localized to those previously identified as close to the enhancer-independent sigma(70) and sigma(38). This indicates a common use of sets of sequences in core for interacting with the two sigma classes. Some sites chosen in sigma(54) for modification with Fe-BABE were positions, which when mutated, deregulate the sigma(54)-holoenzyme and allow activator-independent initiation and holoenzyme isomerization. We infer that these sites in sigma(54) may be involved in interactions with the core that contribute to maintenance of alternative states of the holoenzyme needed for either the stable closed promoter complex conformation or the isomerized holoenzyme conformation associated with the open promoter complex. One site of sigma(54) proximity to the core is apparently not evident with sigma(70), and may represent a specialized interaction. PMID- 10856248 TI - Two histone fold proteins, CHRAC-14 and CHRAC-16, are developmentally regulated subunits of chromatin accessibility complex (CHRAC). AB - The ISWI ATPase of Drosophila is a molecular engine that can drive a range of nucleosome remodelling reactions in vitro. ISWI is important for cell viability, developmental gene expression and chromosome structure. It interacts with other proteins to form several distinct nucleosome remodelling machines. The chromatin accessibility complex (CHRAC) is a biochemical entity containing ISWI in association with several other proteins. Here we report on the identification of the two smallest CHRAC subunits, CHRAC-14 and CHRAC-16. They contain histone fold domains most closely related to those found in sequence-specific transcription factors NF-YB and NF-YC, respectively. CHRAC-14 and CHRAC-16 interact directly with each other as well as with ISWI, and are associated with functionally active CHRAC. The developmental expression profiles of both subunits suggest specialized roles in chromatin remodelling reactions in the early embryo for both histone fold subunits. PMID- 10856249 TI - Overlapping roles for the histone acetyltransferase activities of SAGA and elongator in vivo. AB - Elp3 and Gcn5 are histone acetyltransferases (HATs) that function in transcription as subunits of Elongator and SAGA/ADA, respectively. Here we show that mutations that impair the in vitro HAT activity of Elp3 confer typical elp phenotypes such as temperature sensitivity. Combining an elp3Delta mutation with histone H3 or H4 tail mutations confers lethality or sickness, supporting a role for Elongator in chromatin remodelling in vivo. gcn5Deltaelp3Delta double mutants display a number of severe phenotypes, and similar phenotypes result from combining the elp mutation with mutation in a gene encoding a SAGA-specific, but not an ADA-specific subunit, indicating that Elongator functionally overlaps with SAGA. Because concomitant active site alterations in Elp3 and Gcn5 are sufficient to confer severe phenotypes, the redundancy must be specifically related to the HAT activity of these complexes. In support of this conclusion, gcn5Deltaelp3Delta phenotypes are suppressed by concomitant mutation of the HDA1 and HOS2 histone deacetylases. Our results demonstrate functional redundancy among transcription-associated HAT and deacetylase activities, and indicate the importance of a fine-tuned acetylation-deacetylation balance during transcription in vivo. PMID- 10856250 TI - Separate domains in E1 and E2 proteins serve architectural and productive roles for cooperative DNA binding. AB - The E1 and E2 proteins from bovine papillomavirus bind cooperatively to binding sites in the viral origin of DNA replication. The DNA-binding domains (DBDs) of the two proteins interact with each other, and the E2 transactivation domain interacts with the helicase domain of E1. Mutations that disrupt the interaction between the two DBDs also disrupt the interaction between the E2 activation domain and the E1 helicase domain, demonstrating interdependence of the two interactions. Cooperative binding of the two DBDs generates a sharp bend in the DNA that is required for interaction between the E2 activation domain and E1. This indicates that interaction between the two DBDs plays an architectural role, 'triggering' a productive interaction between the E2 transactivation domain and E1 through introduction of a sharp bend in the DNA. This two-step mechanism may be a required feature for cooperative DNA binding to proximal binding sites. PMID- 10856251 TI - The Epstein-Barr virus lytic program is controlled by the co-operative functions of two transactivators. AB - The propagation of herpesviruses has long been viewed as a temporally regulated sequential process that results from the consecutive expression of specific viral transactivators. As a key step in this process, lytic viral DNA replication is considered as a checkpoint that controls the expression of the late structural viral genes. In a novel genetic approach, we show that both hypotheses do not hold true for the Epstein-Barr virus (EBV). The study of viral mutants of EBV in which the early genes BZLF1 and BRLF1 are deleted allowed a precise assignment of the function of these proteins. Both transactivators were absolutely essential for viral DNA replication. Both BZLF1 and BRLF1 were required for full expression of the EBV proteins expressed during the lytic program, although the respective influence of these molecules on the expression of various viral target genes varied greatly. In replication-defective viral mutants, neither early gene expression nor DNA replication was a prerequisite for late gene expression. This work shows that BRLF1 and BZLF1 harbor distinct but complementary functions that influence all stages of viral production. PMID- 10856252 TI - Characterization of mammalian RAD51 double strand break repair using non-lethal dominant-negative forms. AB - In contrast to yeast RAD51, mammalian mRAD51 is an essential gene. Its role in double strand break (DSB) repair and its consequences on cell viability remain to be characterized precisely. Here, we used a hamster cell line carrying tandem repeat sequences with an I-SCE:I cleavage site. We characterized conservative recombination after I-SCE:I cleavage as gene conversion or intrachromatid crossing over associated with random reintegration of the excised reciprocal product. We identified two dominant-negative RAD51 forms that specifically inhibit conservative recombination: the yeast ScRAD51 or the yeast-mouse chimera SMRAD51. In contrast, the mouse MmRAD51 stimulates conservative recombination. None of these RAD51 forms affects non-conservative recombination or global DSB healing. Consistently, although resistance to gamma-rays remains unaffected, MmRAD51 stimulates whereas ScRAD51 or SMRAD51 prevents radiation-induced recombination. This suggests that mRAD51 does not significantly affect the global DSB repair efficiency but controls the classes of recombination events. Finally, both ScRAD51 and SMRAD51 drastically inhibit spontaneous recombination but not cell proliferation, showing that RAD51-dependent spontaneous and DSB-induced conservative recombination can be impaired significantly without affecting cell viability. PMID- 10856253 TI - Mechanisms of accurate translesion synthesis by human DNA polymerase eta. AB - The XPV (xeroderma pigmentosum variant) gene encodes human DNA polymerase eta (pol eta), which is involved in the replication of damaged DNA. Pol eta catalyzes efficient and accurate translesion synthesis past cis-syn cyclobutane di-thymine lesions. Here we show that human pol eta can catalyze translesion synthesis past an abasic (AP) site analog, N-2-acetylaminofluorene (AAF)-modified guanine, and a cisplatin-induced intrastrand cross-link between two guanines. Pol eta preferentially incorporated dAMP and dGMP opposite AP, and dCMP opposite AAF-G and cisplatin-GG, but other nucleotides were also incorporated opposite these lesions. However, after incorporating an incorrect nucleotide opposite a lesion, pol eta could not continue chain elongation. In contrast, after incorporating the correct nucleotide opposite a lesion, pol eta could continue chain elongation, whereas pol alpha could not. Thus, the fidelity of translesion synthesis by human pol eta relies not only on the ability of this enzyme to incorporate the correct nucleotide opposite a lesion, but also on its ability to elongate only DNA chains that have a correctly incorporated nucleotide opposite a lesion. PMID- 10856254 TI - Crystal structure of NaeI-an evolutionary bridge between DNA endonuclease and topoisomerase. AB - NAE:I is transformed from DNA endonuclease to DNA topoisomerase and recombinase by a single amino acid substitution. The crystal structure of NAE:I was solved at 2.3 A resolution and shows that NAE:I is a dimeric molecule with two domains per monomer. Each domain contains one potential DNA recognition motif corresponding to either endonuclease or topoisomerase activity. The N-terminal domain core folds like the other type II restriction endonucleases as well as lambda exonuclease and the DNA repair enzymes MutH and Vsr, implying a common evolutionary origin and catalytic mechanism. The C-terminal domain contains a catabolite activator protein (CAP) motif present in many DNA-binding proteins, including the type IA and type II topoisomerases. Thus, the NAE:I structure implies that DNA processing enzymes evolved from a few common ancestors. NAE:I may be an evolutionary bridge between endonuclease and DNA processing enzymes. PMID- 10856255 TI - Three telomerases with completely non-telomeric template replacements are catalytically active. AB - Telomerase is a reverse transcriptase minimally composed of a reverse transcriptase protein subunit and an internal RNA component that contains the templating region. Point mutations of template RNA bases can cause loss of enzymatic activity, reduced processivity and misincorporation in vitro. Here we report the first complete replacement of the nine base TETRAHYMENA: thermophila telomerase templating region in vivo with non-telomeric sequences. Rather than ablating telomerase activity, three such replaced telomerases (U9, AUN and AU4) were effective in polymerization in vitro. In vivo, the AU4 and AUN genes caused telomere shortening. We demonstrated the fidelity of the AUN and U9 telomerases in vitro and utilized AUN telomerase to demonstrate that 5' end primer recognition by telomerase is independent of template base pairing. However, the mutant AUN template telomerase catalyzed an abnormal DNA cleavage reaction. For these U-only and AU- substituted templates, we conclude that base-specific interactions between the telomerase template and protein (or distant parts of the RNA) are not absolutely required for the minimal core telomerase functions of nucleotide addition and base discrimination. PMID- 10856256 TI - Structure of tandem RNA recognition motifs from polypyrimidine tract binding protein reveals novel features of the RRM fold. AB - Polypyrimidine tract binding protein (PTB), an RNA binding protein containing four RNA recognition motifs (RRMs), is involved in both pre-mRNA splicing and translation initiation directed by picornaviral internal ribosome entry sites. Sequence comparisons previously indicated that PTB is a non-canonical RRM protein. The solution structure of a PTB fragment containing RRMs 3 and 4 shows that the protein consists of two domains connected by a long, flexible linker. The two domains tumble independently in solution, having no fixed relative orientation. In addition to the betaalphabetabetaalphabeta topology, which is characteristic of RRM domains, the C-terminal extension of PTB RRM-3 incorporates an unanticipated fifth beta-strand, which extends the RNA binding surface. The long, disordered polypeptide connecting beta4 and beta5 in RRM-3 is poised above the RNA binding surface and is likely to contribute to RNA recognition. Mutational analyses show that both RRM-3 and RRM-4 contribute to RNA binding specificity and that, despite its unusual sequence, PTB binds RNA in a manner akin to that of other RRM proteins. PMID- 10856258 TI - State of hypertension as We enter the 21st century : A societal conundrum PMID- 10856257 TI - A novel shuttling protein, 4E-T, mediates the nuclear import of the mRNA 5' cap binding protein, eIF4E. AB - The eukaryotic translation initiation factor 4E (eIF4E) plays an important role in the control of cell growth. eIF4E binds to the mRNA 5' cap structure m(7)GpppN (where N is any nucleotide), and promotes ribosome binding to the mRNA in the cytoplasm. However, a fraction of eIF4E localizes to the nucleus. Here we describe the cloning and functional characterization of a new eIF4E-binding protein, referred to as 4E-T (eIF4E-Transporter). We demonstrate that 4E-T is a nucleocytoplasmic shuttling protein that contains an eIF4E-binding site, one bipartite nuclear localization signal and two leucine-rich nuclear export signals. eIF4E forms a complex with the importin alphabeta heterodimer only in the presence of 4E-T. Overexpression of wild-type 4E-T, but not of a mutant defective for eIF4E binding, causes the nuclear accumulation of HA-eIF4E in cells treated with leptomycin B. Taken together, these results demonstrate that the novel nucleocytoplasmic shuttling protein 4E-T mediates the nuclear import of eIF4E via the importin alphabeta pathway by a piggy-back mechanism. PMID- 10856259 TI - Hypertension in the political arena. PMID- 10856260 TI - Angiotensin II and the heart : on the intracrine renin-angiotensin system. AB - -The active end product of the renin-angiotensin system, angiotensin II (Ang II), through the activation of specific Ang II receptors, regulates cardiac contractility, cell coupling, and impulse propagation and is involved in cardiac remodeling, growth, and apoptosis. We review these subjects, as well as the second messengers that are involved, and the synthesis of Ang II in the heart under normal and pathological conditions. Finally, we discuss the possibility that there is an intracrine renin-angiotensin system in the heart that plays a role in the control of cell communication and inward Ca(2+) current. PMID- 10856261 TI - On the biological actions of intracellular angiotensin. PMID- 10856262 TI - Cardiotrophin-1 increases angiotensinogen mRNA in rat cardiac myocytes through STAT3 : an autocrine loop for hypertrophy. AB - -Cardiotrophin-1, an interleukin-6-related cytokine, stimulates the Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway and induces cardiac myocyte hypertrophy. In this study, we demonstrate that cardiotrophin-1 induces cardiac myocyte hypertrophy in part by upregulation of a local renin-angiotensin system through the JAK/STAT pathway. We found that cardiotrophin-1 increased angiotensinogen mRNA expression in cardiac myocytes via STAT3 activation. Tyrosine phosphorylation of STAT3 by cardiotrophin-1 treatment resulted in STAT3 homodimer binding to the St-domain in the angiotensinogen gene promoter, which lead to promoter activation in a transient transfection assay. Cardiotrophin-1-induced STAT3 tyrosine phosphorylation and binding to the St domain were suppressed by AG490, a specific JAK2 inhibitor, which also attenuated cardiotrophin-1-stimulated angiotensinogen promoter activity. Cardiotrophin-1 did not activate the angiotensinogen gene promoter that contained a substitution mutation within the St-domain. Finally, losartan, an angiotensin II type 1 receptor antagonist, significantly attenuated cardiotrophin-1-induced hypertrophy of neonatal rat cardiac myocytes. Angiotensin II is known to induce cardiac myocyte hypertrophy by activating the G-protein-coupled angiotensin II type 1 receptor. Our results suggest that upregulation of angiotensinogen and angiotensin II production contribute to cardiotrophin-1-induced cardiac myocyte hypertrophy and emphasize an important interaction between G-protein-coupled and cytokine receptors. PMID- 10856263 TI - Chronic AT(1) blockade stimulates extracellular collagen type I degradation and reverses myocardial fibrosis in spontaneously hypertensive rats. AB - It has been suggested that left ventricular fibrosis in spontaneously hypertensive rats (SHR) is the result of both exaggerated collagen synthesis and insufficient collagen degradation. We have shown previously that chronic treatment with the angiotensin II type 1 receptor antagonist losartan results in diminished synthesis of collagen type I molecules and reversal of myocardial fibrosis in SHR. This study was designed to investigate whether losartan also affects the extracellular degradation of collagen type I fibers in the left ventricle of SHR. The study was performed in 30-week-old normotensive Wistar Kyoto rats (WKY), untreated SHR, and SHR treated with orally administered losartan (20 mg/kg per day) for 14 weeks before they were killed. Ventricular collagenase activity was determined by degradation of [(14)C]collagen with tissue extracts. Ventricular expression of tissue inhibitor of metalloproteinases 1 (TIMP-1) mRNA was analyzed by Northern blot. A histomorphometric study of the left ventricle was performed in all rats. Compared with WKY, SHR exhibited left ventricular hypertrophy, increased (P<0.05) blood pressure, left ventricular collagen volume fraction and TIMP-1 mRNA, and diminished (P<0.05) collagenase activity. After the treatment period, blood pressure was higher (P<0.05) in losartan-treated SHR than in WKY, and no significant differences were noted in the remaining parameters between the 2 strains of rats. Compared with untreated SHR, treated SHR showed no left ventricular hypertrophy, diminished (P<0.05) blood pressure, left ventricular collagen volume fraction and TIMP-1 mRNA, and increased (P<0.05) collagenase activity. These results suggest that the transcription of the TIMP-1 gene is upregulated in the hypertrophied and fibrotic left ventricle of adult SHR. Upregulation of TIMP-1 may account for diminished collagenase activity in the myocardium of those rats. Chronic angiotensin II type 1 receptor blockade with losartan resulted in inhibition of TIMP-1 expression and stimulation of collagenase activity in the left ventricle of SHR. It is proposed that angiotensin II may facilitate myocardial fibrosis in SHR by depressing the collagenase-mediated extracellular degradation of collagen fibers. PMID- 10856264 TI - Altered inotropic responsiveness and gene expression of hypertrophied myocardium with captopril. AB - Inotropic responsiveness to beta-adrenergic stimulation is generally found to be impaired in left ventricular (LV) hypertrophy and failure. To investigate the mechanisms by which angiotensin-converting enzyme inhibitor therapy may modulate inotropic responsiveness with long-term pressure overload, we studied the effects of captopril treatment on cardiac gene expression, LV muscle mechanical contraction, and intracellular calcium (Ca(2+)) transients from spontaneously hypertensive rats (SHR). LV papillary muscles from untreated SHR, age-matched normotensive Wistar-Kyoto rats (WKY), and SHR treated with captopril (CAP(Rx) started at 12, 18, and 21 months of age) were studied. All animals were studied at 24 months of age or when heart failure developed. In untreated SHR, alpha myosin heavy chain (MHC) gene expression and protein were decreased, the Ca(2+) transient (with the bioluminescent indicator aequorin) was prolonged, and abundance of Na(+)/Ca(2+) exchanger mRNA levels increased in comparison to WKY. Active stress development at L(max) and the maximum rate of stress development were depressed and contractile duration prolonged in SHR relative to WKY. Isoproterenol administration further decreased active stress in untreated SHR despite an increase in intracellular Ca(2+) levels. In CAP(Rx) SHR, alpha-MHC gene expression and protein levels were increased, the Ca(2+) transient was not prolonged, Na(+)/Ca(2+) exchanger expression was downregulated, and papillary muscle function demonstrated increased active stress and maximum rate of stress development in response to isoproterenol. The increased abundance of alpha-MHC mRNA in conjunction with an increase in V(1) myosin isozyme suggests that captopril affects transcriptional regulation of cardiac gene expression. Restored LV inotropic responsiveness to beta-adrenergic stimulation in CAP(Rx) SHR appears to be coupled to normalization of Na(+)/Ca(2+) exchanger mRNA expression, upregulation of V(1) myosin isozyme levels, and increased speed of contraction. PMID- 10856265 TI - Enhanced adrenomedullin production by mechanical stretching in cultured rat cardiomyocytes. AB - Adrenomedullin (AM) is secreted from cultured cardiac myocytes. In this study, we examined whether mechanical stretching stimulates AM production in cardiac myocytes, and if so, whether angiotensin II (Ang II) is involved in that mechanism. Neonatal rat cardiac myocytes cultured in serum-free medium were stretched 10% or 20% on flexible silicone rubber culture dishes, and AM mRNA expression was examined by quantitative polymerase chain reaction. The AM mRNA levels in the myocytes stretched 10% and 20% for 24 hours significantly increased by 56% (P<0.05) and 88% (P<0.01), respectively, when compared with the levels in nonstretched cells. AM secretion into the medium after the myocytes were stretched 10% and 20% increased by 22% (P<0.05) and 45% (P<0.01), respectively. In nonstretched myocytes incubated with 10(-6) mol/L Ang II for 24 hours, AM mRNA and secretion increased by 86% (P<0.05) and 36% (P<0. 01), respectively. These effects of Ang II were abolished by 10(-6) mol/L CV-11974, an Ang II type I (AT(1)) receptor antagonist, but not by 10(-6) mol/L PD-123319, an Ang II type II antagonist. Stretch-induced increases of AM gene expression and secretion were significantly inhibited (P<0.05) in the presence of 10(-6) mol/L CV-11974 by 46% and 52%, respectively; however, they were not affected by 10(-6) mol/L PD-123319. These findings indicate that AM production from cardiac myocytes is augmented by mechanical stretching, partially through the AT(1) receptors, which suggests a local interaction between AM and the renin-angiotensin system in stretched cardiac myocytes. PMID- 10856266 TI - Atrial natriuretic peptide is involved in renal actions of moxonidine. AB - Moxonidine, an antihypertensive imidazoline compound, reduces blood pressure by selective activation of central imidazoline I(1)-receptors and inhibition of sympathetic nerve activity and by direct actions on the kidney, with both mechanisms resulting in diuresis and natriuresis. We hypothesized that the hypotensive and renal actions of moxonidine may be mediated by atrial natriuretic peptide (ANP), a cardiac peptide involved in pressure and volume homeostasis through its vasodilatory, diuretic, and natriuretic actions. Renal parameters were measured on an hourly basis over a period of 4 hours in conscious rats that received bolus intravenous injections of moxonidine (1 to 150 microg/300 microL saline). During the first hour, moxonidine dose-dependently stimulated diuresis, natriuresis, kaliuresis, and urinary cGMP, the index of ANP activity. Moxonidine (50 microg) significantly (P<0.001) stimulated urinary volume (0.35+/-0.04 versus 1.05+/-0.09 mL/h per 100 g), sodium (14. 3+/-2.5 versus 51.8+/-6.5 micromol/h per 100 g), potassium (10.5+/-2. 3 versus 32.3+/-3.2 micromol/h per 100 g), and cGMP (325+/-52 versus 744+/-120 pmol/h per 100 g). Pretreatment with a selective imidazoline receptor antagonist, efaroxan, dose-dependently inhibited moxonidine stimulated renal parameters. Efaroxan (25 microg per rat) significantly inhibited moxonidine-stimulated diuretic and natriuretic effects and urinary cGMP excretion (744+/-120 versus 381+/-137 pmol/h per 100 g, P<0.02). The alpha(2)-adrenoceptor antagonist yohimbine (50 microg per rat) partially yet significantly inhibited moxonidine-stimulated diuresis and natriuresis but not cGMP excretion. Plasma ANP was dose-dependently increased by moxonidine and was inhibited by pretreatment with efaroxan (220.8+/-36.9 versus 100.3+/-31.7 pg/mL, P<0.03) but not by yohimbine. In conclusion, selective in vivo activation of imidazoline receptors by moxonidine is associated with dose-dependent diuresis, natriuresis, and kaliuresis as well as stimulated plasma ANP and urinary cGMP excretion, thus implicating ANP in the renal actions of moxonidine. PMID- 10856267 TI - Vasopeptidase inhibition has potent effects on blood pressure and resistance arteries in stroke-prone spontaneously hypertensive rats. AB - The antihypertensive agent omapatrilat represents a novel approach to antihypertensive therapy, namely vasopeptidase inhibition. Omapatrilat (BMS 186716) concomitantly inhibits neutral endopeptidase and angiotensin-converting enzyme, leading to protection from degradation of natriuretic and other hypotensive peptides in addition to interruption of the renin-angiotensin system. Although the potency of omapatrilat on reduction of blood pressure has been reported, its effects on resistance artery structure and function were unknown. We tested omapatrilat in stroke-prone spontaneously hypertensive rats (SHRSP), a malignant model of hypertension, with the hypothesis that it would improve the structure and endothelial function of mesenteric resistance arteries. Ten-week old SHRSP were treated orally for 10 weeks with omapatrilat (40 mg/kg per day). Mesenteric arteries (lumen <300 microm) were studied on a pressurized myograph. After 10 weeks, untreated SHRSP had a systolic blood pressure of 230+/-2 mm Hg that was significantly reduced (P<0.05) by omapatrilat (145+/-3 mm Hg). Omapatrilat treatment improved endothelium-dependent relaxation of resistance arteries as elicited by acetylcholine (10(-5) mol/L) but had no significant effect on endothelium-independent relaxation produced by a nitric oxide donor (sodium nitroprusside). This suggested that there existed endothelial dysfunction in SHRSP that was corrected by vasopeptidase inhibition, probably in part caused by the potent blood pressure-lowering effect of omapatrilat. Media width and media/lumen ratio were significantly decreased (P<0.05) by omapatrilat, and a trend (P=0.07) to increase lumen diameter was observed. Vascular stiffness (slope of the elastic modulus versus stress curve) was unaltered by omapatrilat. In conclusion, omapatrilat, acting as a potent antihypertensive agent, may improve structure and endothelial function of resistance arteries in SHRSP, a severe form of genetic hypertension. PMID- 10856268 TI - In vitro and in vivo inhibition of the 2 active sites of ACE by omapatrilat, a vasopeptidase inhibitor. AB - The vasopeptidase inhibitor omapatrilat inhibits both neutral endopeptidase and angiotensin-converting enzyme (ACE). The in vitro and in vivo inhibitory potency of omapatrilat and the specific ACE inhibitor fosinopril toward the 2 active sites of ACE (called N- and C-domains) was investigated with the use of 3 substrates: angiotensin I, which is equally cleaved by the 2 ACE domains; hippuryl-histidyl-leucine, specific synthetic substrate of the C-domain in high- salt conditions; and a newly synthesized specific substrate of the N-domain designed by acetylating the lysine residue of AcSDKP. In vitro, omapatrilat was 5 times more potent than fosinoprilat in inhibiting angiotensin I hydrolysis. Omapatrilat inhibited similarly both N- and C-domain hydrolysis, whereas fosinoprilat was slightly more specific for the N-domain. The in vivo selective inhibitory potency of single oral doses of 10 mg omapatrilat and 20 mg fosinopril were investigated in a double-blind, placebo-controlled, cross-over study in 9 mildly sodium-depleted normotensive subjects. In accordance with the in vitro results, fosinopril appeared to be more specific for the N-domain than the C domain in vivo, since plasma and urine AcSDKP concentrations were significantly higher than those observed with omapatrilat. This study shows that it is possible to assess separately in vitro and in vivo the selectivity of ACE or ACE/neutral endopeptidase inhibitors. A differential selectivity may explain some peculiar properties observed with some ACE inhibitors. PMID- 10856269 TI - Peroxisome proliferator-activated receptor-gamma ligands inhibit nitric oxide synthesis in vascular smooth muscle cells. AB - Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a key player in glucose metabolism. If PPARgamma ligands modulate nitric oxide (NO) synthesis in the vascular tissue, they may affect the process of plaque formation and postangioplasty restenosis. We investigated the effects of PPARgamma ligands on NO synthesis in vascular smooth muscle cells. Incubation of cultures with interleukin-1beta (10 ng/mL) for 24 hours caused a significant increase in the production of nitrite, a stable metabolite of NO, in cultured rat vascular smooth muscle cells. The PPARgamma agonists troglitazone and 15-deoxy-triangle up(12,14) prostaglandin J(2) (15d-PG J(2)) dose-dependently inhibited nitrite production by interleukin-1beta-stimulated vascular smooth muscle cells. Decreased interleukin 1beta-induced nitrite production by the PPARgamma agonists was accompanied by decreased inducible NO synthase mRNA and protein accumulation. Interleukin-1beta induced nuclear factor-kappaB activation in vascular smooth muscle cells, and both troglitazone and 15d-PG J(2) markedly suppressed this nuclear factor-kappaB activation. PPARgamma ligands inhibit NO synthesis in cytokine-stimulated vascular smooth muscle cells, suggesting that these agonists may act directly on the vascular smooth muscle and influence the process of atherosclerosis and restenosis. PMID- 10856270 TI - Interactions between nitric oxide and endothelin in the regulation of vascular tone of human resistance vessels in vivo. AB - Endothelial release of nitric oxide (NO) contributes to the regulation of vascular tone by inducing vascular relaxation. In addition, NO may inhibit the synthesis and hemodynamic effects of endothelin-1 (ET-1), a powerful endothelium derived vasoconstrictor peptide that may stimulate NO production. However, whether NO and ET-1 physiologically interact to regulate vascular tone in humans has not been defined. In this study, the interactions between the L-arginine NO pathway and the ET-1 system in the regulation of vascular tone in human forearm resistance vessels were examined in vivo. Vasomotor response to the NO synthase inhibitor N(G)-monomethyl-L-arginine (L-NMMA, 4 micromol/min for 30 minutes) was measured during either saline infusion or blockade of ET-1 receptors. Endothelin A (ET(A)) and endothelin-B (ET(B)) receptor blockade was achieved by infusion of BQ-123 (100 nmol/min) and BQ-788 (50 nmol/min), respectively, separately and in combination. Drugs were infused into the brachial artery, and the forearm blood flow (FBF) response was measured by strain-gauge plethysmography. During saline infusion, L-NMMA administration significantly decreased FBF (25%, P<0.01 versus baseline). This effect was significantly blunted during nonselective blockade of ET-1 receptors (7% decrease in FBF, P=0.02 versus the effect of L-NMMA during saline infusion). Selective ET(A) blockade did not modify the vasoconstrictor response to L-NMMA (26% decrease in FBF, P=0.66 versus the effect of L-NMMA during saline infusion), but selective ET(B) receptor antagonism caused significant diminution of the hemodynamic response to NO inhibition (8% decrease in FBF, P=0.04 versus the effect of L-NMMA during saline infusion). Thus ET-1 contributes to the regulation of vascular tone by stimulating NO activity. This effect is mediated through endothelial ET(B) receptors and may be relevant in conditions associated with endothelial dysfunction. PMID- 10856271 TI - Aging increases PGHS-2-dependent vasoconstriction in rat mesenteric arteries. AB - During aging, the vascular endothelium changes functionally and morphologically. Although previous studies have shown that endothelium-derived eicosanoids increase vessel tone in aging, the precise mechanism(s) has not been fully determined. We hypothesized that aging would increase prostaglandin H synthase (PGHS)-dependent vasoconstriction as well as decrease nitric oxide-dependent relaxation. Mesenteric arteries from 3-month-old (n=9) and 12-month-old (n=14) female Sprague-Dawley rats were studied in a myograph system. Aging significantly blunted the endothelium-dependent relaxation response to methacholine compared with young rats (EC(50)=7.77x10(-8) versus 2.68x10(-8) mol/L, P<0. 05). Nitric oxide synthase inhibition reduced methacholine-induced relaxation in the young (P<0.05) but had no effect in the aging group. Specific inhibition of the PGHS-1 isoform did not significantly affect methacholine-mediated relaxation in the young or aged groups. However, PGHS-2 inhibition greatly enhanced relaxation to methacholine (1.59x10(-8) versus 7.77x10(-8) mol/L, P<0.01) in the aged group only, restoring vessel function to that of the young. In the aged group, inhibition of the prostaglandin H(2)/thromboxane A(2) receptor enhanced methacholine-dependent relaxation similar to that of PGHS-2 inhibition. Moreover, arterial expression of PGHS-2 protein increased with age. In summary, nitric oxide-dependent modulation of vessel function decreased with age, PGHS-1 did not significantly affect vessel tone in either the young or aging group, and PGHS-2 greatly increased vasoconstriction in aging. Thus, we have identified enhanced PGHS-2-mediated vasoconstriction in aging and therefore suggest that inhibition of this isoform is potentially a new target for therapeutic intervention to improve vascular function. PMID- 10856272 TI - Angiotensin-(1-7) reduces norepinephrine release through a nitric oxide mechanism in rat hypothalamus. AB - Angiotensin (Ang)-(1-7) elicits a facilitatory presynaptic effect on peripheral noradrenergic neurotransmission, and because biological responses to the heptapeptide on occasion are tissue specific, the present investigation was undertaken to study its action on noradrenergic neurotransmission at the central level. In rat hypothalamus labeled with [(3)H]-norepinephrine, 100 to 600 nmol/L Ang-(1-7) diminished norepinephrine released by 25 mmol/L KCl. This effect was blocked by the selective angiotensin type 2 receptor antagonist PD 123319 (1 micromol/L) and by the specific Ang-(1-7) receptor antagonist ([D-Ala(7)]Ang-(1 7) (1 micromol/L) but not by losartan (10 nmol/L to 1 micromol/L), a selective angiotensin type 1 receptor antagonist. The inhibitory effect on noradrenergic neurotransmission caused by Ang-(1-7) was prevented by 10 micromol/L N(omega) nitro-L-arginine methylester, an inhibitor of nitric oxide synthase activity, and was restored by 100 micromol/L L-arginine, precursor of nitric oxide synthesis. Methylene blue (10 micromol/L), an inhibitor of guanylate cyclase considered as the target of nitric oxide action, as well as Hoe 140 (10 micromol/L), a bradykinin B(2)-receptor antagonist, prevented the inhibitory effect of the heptapeptide on neuronal norepinephrine release, whereas no modification was observed in the presence of 0.1 to 10 micromol/L indomethacin, a cyclooxygenase inhibitor. Our results indicate that Ang-(1-7) has a tissue-specific neuromodulatory effect on noradrenergic neurotransmission, being inhibitory at the central nervous system by a nitric oxide-dependent mechanism that involves angiotensin type 2 receptors and local bradykinin production. PMID- 10856273 TI - Modulatory effects of carbon monoxide on baroreflex activation in nucleus tractus solitarii of rats. AB - Recent studies suggest that carbon monoxide (CO), which is produced in significant quantities in many brain regions, may function as a neurotransmitter. Heme oxygenase catalyzes the metabolism of heme to CO and biliverdin; however, the physiological role of CO in central cardiovascular regulation was not well understood. In the present study, we evaluated the baroreflex response of CO in the nucleus tractus solitarii (NTS) of rats. Male Sprague-Dawley rats were anesthetized with urethane, and blood pressure and heart rate were monitored intra-arterially. Unilateral microinjection (60 nL) of hematin, a heme molecule cleaved by heme oxygenase to yield CO, into the NTS produced prominent dose related depressor and bradycardic effects. Baroreflex responses were elicited by increasing doses of phenylephrine (10 to 30 microg/kg IV) before and after intra NTS administration of zinc deuteroporphyrin 2,4-bis-glycol (ZnDPBG) (1 nmol), an inhibitor of heme oxygenase activity, or vehicle alone. The reflex bradycardia elicited by phenylephrine was significantly inhibited by pretreatment with ZnDPBG. Furthermore, the inhibitory effect of ZnDPBG on baroreflex activation was dose dependent. These results suggest CO formed by brain heme oxygenase plays a significant role in central cardiovascular regulation and that inhibition of heme oxygenase attenuated baroreflex activation. PMID- 10856274 TI - Study of arterial and autonomic effects of cyclosporine in humans. AB - Altered sympathetic activity and peripheral vascular function are suspected as a mechanism of the development of arterial hypertension in organ transplantation recipients treated with cyclosporine. We assessed whether cyclosporine might alter peripheral vascular properties or autonomic modulation of the sinus node and the vasculature during rest and standing. We examined 17 orthotopic heart transplantation recipients, 8 solid organ transplantation recipients, 17 patients with essential hypertension, and 42 normotensive control subjects. All except the normotensive control subjects were treated with a long-acting dihydropyridine calcium entry blocker; transplantation recipients also received cyclosporine based immunosuppression. Radial artery compliance was reduced in patients with essential hypertension and in patients with heart and solid organ transplantation as compared with normotensive control subjects, with this reduction being more marked in heart transplantation recipients. At rest, R-R variance was lowest in heart transplantation recipients, denoting denervation. The spectral profile of both R-R and systolic blood pressure variability as well as the index of baroreflex gain was normal at rest in patients with solid organ transplantation. On standing, both transplantation groups demonstrated reduced responsiveness in markers of autonomic modulation. The decrease in arterial compliance in cyclosporine-induced hypertension seems to imply a degree of ventricular vascular uncoupling more apparent in heart transplantation recipients. These changes are associated with alterations in autonomic modulation that are evidenced by an orthostatic stimulus. PMID- 10856275 TI - Chronic I(1)-imidazoline agonism : sympathetic mechanisms in hypertension. AB - Evidence exists for a state of sympathetic hyperactivity in essential hypertension, and moxonidine, a new central sympathetic inhibitor, has been introduced for its treatment. Acute administration of moxonidine lowers peripheral sympathetic neural output. This study examined the effect of chronic moxonidine therapy, at increasing therapeutic doses, on resting peripheral sympathetic activity and vascular resistance and their responses to physiological reflex maneuvers. Twelve newly diagnosed patients with essential hypertension were studied sequentially at least 1 month apart, initially on no therapy, then on 200 microg, and finally on 400 microg of oral moxonidine daily. Changes in heart rate, arterial blood pressure, calf vascular resistance, and peripheral sympathetic drive were assessed at rest and during reflex maneuvers. Peroneal microneurography was used to quantify peripheral sympathetic vasoconstrictor activity by single-unit and multiunit techniques. Moxonidine therapy progressively reduced resting mean arterial pressure (P<0.0001) without affecting heart rate. At 200 microg daily, there was a significant reduction in sympathetic nerve activity (P<0.001) and calf vascular resistance (P<0.01). At 400 microg daily, further reductions were smaller and insignificant. Responses to cold stimulus and isometric handgrip exercise showed a similar pattern, with the greatest magnitude of change at 200 microg daily. In patients with essential hypertension, chronic moxonidine therapy inhibited resting sympathetic vasoconstrictor drive and also its reflex responses. The magnitude of inhibition became less as the therapeutic dose was increased, suggesting that moxonidine may be more effective under conditions of high sympathetic activity. PMID- 10856276 TI - Physiology and pathophysiology of the adipose tissue renin-angiotensin system. AB - The renin-angiotensin system has long been recognized as an important regulator of systemic blood pressure and renal electrolyte homeostasis, and local renin angiotensin systems have also been implicated in pathological changes of organ structure and function by modulation of gene expression, growth, fibrosis, and inflammatory response. Recently, substantial data have been accumulated in support of the notion that adipose tissue, besides other endocrine functions, also hosts a local renin-angiotensin system. In the first part of this review, we describe the components of the adipose tissue renin-angiotensin system in human and rodent animal models with respect to regulation of angiotensinogen expression and secretion, formation of angiotensin peptides, and the existence of angiotensin II receptors. In the second part, we describe the role of the adipose tissue renin-angiotensin system in the process of adipogenic differentiation and in the regulation of body weight. We also detail the differential regulation of the adipose tissue renin-angiotensin system in obesity and hypertension and thereby also speculate on its possible role in the development of obesity associated hypertension. Although some findings on the adipose tissue renin angiotensin system appear to be confusing, its involvement in the physiology and pathophysiology of adipose tissue has been confirmed by several functional studies. Nevertheless, future studies with more carefully described phenotypes are necessary to conclude whether obesity (by stimulation of adipogenic differentiation) and hypertension are associated with changes of renin angiotensin system activity in adipose tissue. If so, the physiological relevance of this system in animal models and humans may warrant further interest. PMID- 10856277 TI - Hydrolysis by cathepsin B of fluorescent peptides derived from human prorenin. AB - Cathepsin B is a lysosomal thiolprotease that, because of its colocalization with renin and its ability to activate prorenin, has been proposed as a prorenin processing enzyme. To characterize the biochemical aspect of this potential cathepsin B activity in more detail, we synthesized and assayed with human cathepsin B the internally quenched fluorescent peptide Abz-FSQPMKRLTLGNTTQ-EDDnp (Abz, ortho-aminobenzoic acid fluorescent group and EDDnp, N-?2, 4-dinitrophenyl ethylenediamine quencher group) that contains 7 amino acids for each side of the R-L bond that is the processing site of human prorenin. Human cathepsin B hydrolyzed this peptide at the correct site (R-L bond), with k(cat)/K(m)=75 mmol/L(-1) s(-1). Analogues of this peptide obtained by Ala scanning at positions P(5) to P(5)' were also synthesized and assayed as substrates for human cathepsin B. The obtained specificity constant (k(cat)/K(m)) values have a significant parallel with the previous data of prorenin activation by AtT-20 cells and in vitro by cathepsin B. In addition, we demonstrated the presence of cathepsin B like activity in rat mesangial cells and the ability of its whole soluble fraction lysates, as well as that of purified cloned rat cathepsin B, to hydrolyze Abz-IKKSSF-EDDnp at the K-S bond, which contains 6 amino acids of rat prorenin processing site. The specificity data of cathepsin B toward peptides derived from prorenin processing site support the view that human or rodent cathepsin B could be involved in the intracellular processing of prorenin that is locally synthesized or taken up from the extracellular compartment. PMID- 10856278 TI - Angiotensin I-converting enzyme isoforms (high and low molecular weight) in urine of premature and full-term infants. AB - Angiotensin I-converting enzyme (ACE) isoforms in urine from healthy and mildly hypertensive untreated patients have been described in the literature. Healthy subjects have high- and low-molecular-weight ACEs (170 and 65 kDa), whereas mildly hypertensive untreated patients have only low-molecular-weight ACEs (90 and 65 kDa), both of which resemble ACE from the N-terminal domain. Previous studies have shown that ACE is regulated during development, and renal tubules of premature human infants are not completely mature, given that nephrogenesis is not complete until the 36th week of gestation. The aim of the present study was to purify and characterize ACE isoforms from urine of premature and full-term infants and to detect the presence of the N-domain form of ACE during prenatal development. Urine from premature and full-term infants was concentrated in an Amicon concentrator, dialyzed in the same equipment against 50 mmol/L Tris-HCl buffer (pH 8.0) that contained 150 mmol/L NaCl, and submitted to gel filtration on an AcA-34 column equilibrated with the buffer described above. Two peaks (P1 and P2 for premature infants; TP1 and TP2 for full-term infants) with ACE activity on hippuryl-His-Leu (K(m), 3 mmol/L) were detected. All enzymes were Cl( ) dependent and inhibited by captopril and EDTA. The peptides angiotensin-(1-7) and N-acetyl-Ser-Asp-Lys-Pro, described as specific for N-domain ACE, were hydrolyzed by P2 and TP2, which suggests that both enzymes are N-domain ACE. In premature infants, P1 activity with hippuryl-His-Leu was 12-fold lower than P2 activity, but in full-term infants, the difference between TP1 and TP2 was 1.6 fold. Chromatography profiles of urine from premature infants were analyzed on days 1, 3, 7, 14, 21, and 30 after birth. The P1 of ACE was detected around the 21st and 30th days, whereas P2 was detected from day 1. These results suggest that ACE activity is related to renal development and that N-domain ACE as well as full-length ACE is present in urine from premature infants. This may indicate that healthy subjects produce and secrete the N-domain form of ACE even before term development. PMID- 10856279 TI - A genome-wide search for susceptibility loci to human essential hypertension. AB - We undertook a systematic search of the entire human genome with the affected sibling-pair model to identify major susceptibility loci to essential hypertension. Affected nuclear families (n=263) were recruited and divided according to definite or probable genetic contribution to hypertension depending on number of hypertensive siblings. The largest nuclear families were first screened with a set of microsatellite markers. Regions on the genome with P<0.05 were tested against the second set of smaller families. An exclusion map was generated to identify regions in which hypertension-causing genes are unlikely to reside. Sibling-pair linkage analysis identified a single locus on chromosome 11q (P<0.004) in the first pass. A second pass with nuclear families that had only affected sibling pairs was, as expected, insufficient to support linkage to 11q. Multipoint exclusion-linkage analysis showed that 3 genetic loci are necessary to explain familial aggregation of essential hypertension. Our preliminary findings suggest that no single region within the human genome contains genes with a major contribution to essential hypertension. We show that the disease is indeed polygenic, with each gene providing a relatively small risk. Our exclusion map will help future investigators to concentrate on areas likely to contain these genes. The region on chromosome 11 is the first to point to a new candidate gene for hypertension that has arisen out of a genome search, but replication of these results at a higher significance is necessary before positional cloning can be justified. PMID- 10856280 TI - Lack of association of 3 functional gene variants with hypertension in African Americans. AB - African Americans are a critical population in which to study the impact of physiologically important candidate gene mutations on the occurrence of hypertension. African Americans not only have a higher prevalence of hypertension, but the disease strikes earlier, with greater severity, and often results in death at an earlier age compared with whites in the United States. In this study, 3 physiologically important candidate gene mutations (angiotensinogen A[-6], alpha-Adducin Gly460Trp, and G-Protein beta(3)-subunit C825T) were examined for their association with hypertension status in a sample of 904 African Americans from Jackson, Mississippi. Tests of simple association and multivariate logistic regression analyses revealed no association between hypertension status and any of the studied polymorphisms. This lack of association persisted after stratification of the sample by gender and body size. These data indicate that these polymorphisms do not contribute in a significant way to interindividual variation in the risk of hypertension in this sample of African Americans, and further genome-wide studies should be performed to identify genes that may influence blood pressure levels in this population. PMID- 10856281 TI - Low birth weight predicts elevated plasma cortisol concentrations in adults from 3 populations. AB - Low birth weight is linked with raised blood pressure in adult life. Recent evidence has suggested that a neuroendocrine disturbance involving the hypothalamic-pituitary-adrenal axis could mediate this link. We therefore investigated the relation between birth weight and fasting plasma cortisol concentrations and the association of cortisol with current blood pressure in population samples of 165 men and women born in Adelaide, South Australia, from 1975 to 1976, 199 men and women born in Preston, UK, from 1935 to 1943, and 306 women born in East Hertfordshire, UK, from 1923 to 1930. Fasting plasma cortisol was measured in plasma samples obtained between 8 and 10 AM. Blood pressure was measured with an automated sphygmomanometer. Low birth weight was associated with raised fasting plasma cortisol concentrations in all 3 populations. A combined analysis that allowed for differences in the gender composition, age, and body mass index between the studies showed that cortisol concentrations fell by 23.9 nmol/L per kilogram increase in birth weight (95% CI 9.6 to 38.2, P<0.001). Fasting plasma cortisol concentrations also correlated positively with the subjects' current blood pressure. However, the association between cortisol and blood pressure was most marked in subjects who were obese (P=0.038 for interaction between body mass index and cortisol, P=0.01 for interaction between waist-to-hip ratio and cortisol). These results show that low birth weight is associated with raised fasting plasma cortisol concentrations. Increased activity of the hypothalamic-pituitary-adrenal axis may link low birth weight with raised blood pressure in adult life. PMID- 10856282 TI - 8-Iso-prostaglandin f(2alpha) reduces trophoblast invasion and matrix metalloproteinase activity. AB - Preeclampsia is a common pregnancy complication in the latter half of gestation diagnosed by hypertension and proteinuria. A key feature of preeclampsia is an altered placentation with reduced trophoblast invasion. Normal placentation requires controlled invasion of trophoblasts into the maternal uterine wall, with secretion of specific proteolytic enzymes able to degrade basement membranes and extracellular matrix, such as the matrix metalloproteinases (MMPs). 8-Iso prostaglandin F(2alpha) (8-iso-PGF(2alpha)) is a marker of oxidative stress in vivo and is biologically active. We have recently reported an elevated content of free 8-iso-PGF(2alpha) in preeclamptic gestational tissue at delivery. Assuming an elevated level of 8-iso-PGF(2alpha) during the invasion period of the pregnancy, we hypothesized that 8-iso-PGF(2alpha) could reduce invasion of JAR cells, a choriocarcinoma cell line. We investigated JAR cell invasion with 2 types of Transwell assays and demonstrated that 8-iso-PGF(2alpha) (10 micromol/L) resulted in reduced cell invasion in both the colorimetric and radioactivity Transwell assays (P<0.01). Zymograms revealed reduced MMP-2 and MMP-9 activity in conditioned media from JAR cells incubated with 8-iso-PGF(2alpha) (10 micromol/L) (P<0.02). 8-Iso-PGF(2alpha) (10 micromol/L) also reduced the collagenase type IV activity in the conditioned media of JAR cells (P=0.04). No effects on MMP-2 and MMP-9 mRNA levels were observed after incubation with 8-iso-PGF(2alpha) (10 micromol/L), whereas protein levels were significantly decreased (P<0.02), suggesting a posttranscriptional regulation. We hypothesize a potential role for 8-iso-PGF(2alpha) in the reduced trophoblast invasion in preeclampsia. PMID- 10856283 TI - Bradykinin-induced vasodilation of human forearm resistance vessels is primarily mediated by endothelium-dependent hyperpolarization. AB - Bradykinin (BK) stimulates endothelial cells to release a number of relaxing factors, such as NO, prostanoids (PGs), and an endothelium-derived hyperpolarizing factor (EDHF). However, the contributions of NO, PG, and EDHF in the vascular relaxation to BK vary with species and anatomic origin of blood vessels used. Therefore, the present study was designed to investigate the contributions of NO, PG, and EDHF in vasodilation caused by BK in human forearm resistance vessels. Forearm blood flow (FBF) was recorded with venous occlusion plethysmography in healthy nonsmoking subjects. At first, studies were performed to validate the NO clamp technique for its ability to inhibit endogenous NO generation. Brachial artery infusion of serotonin (0.6, 1.8, and 6 ng. 100 mL forearm volume [FAV](-1). min(-1)) caused significant forearm vasodilation (2.6 to 4.6 mL. 100 mL FAV(-1). min(-1)), which is known to be NO mediated. Indeed, during the NO clamp, cumulative doses of serotonin caused no vasodilation (2.4 to 2.6 mL. 100 mL FAV(-1). min(-1)), indicating that the generation of endogenous NO was completely blocked. Thereafter, the vasodilative actions of BK were investigated. Brachial artery infusion of BK (50, 100, and 200 ng. 100 mL FAV( 1). min(-1)) caused significant forearm vasodilation in all studies (from 3.1 to 20.4 mL. 100 mL FAV(-1). min(-1)). After the inhibition of cyclooxygenase and NO synthase activity through the use of aspirin and the NO-clamp technique, BK increased FBF in a similar manner (3.9 to 18.9 mL. 100 mL FAV(-1). min(-1)), indicating that the vasodilative actions of BK are independent of NO and PG generation. However, vasodilation caused by the 2 lower doses of BK were significantly attenuated after K(Ca) channel activity was blocked with tetraethylammonium chloride (0.1 mg. 100 mL FAV(-1). min(-1)), suggesting that in the lower dose range, BK mediates vasodilation through the opening of vascular potassium channels. In conclusion, BK is a potent vasodilator peptide in human forearm resistance vessels, causing vasodilation through hyperpolarization of the vascular wall independent of NO and PG production. In addition, the NO-clamp technique is a valid instrument to investigate the contribution of NO in the vasodilative response to different agents. PMID- 10856284 TI - Antagonist-induced intracellular sequestration of rabbit bradykinin B(2) receptor. AB - In a contractility assay based on the rabbit jugular vein, the structurally related drugs NPC 17731 or icatibant (1 to 3 nmol/L) were insurmountable antagonists of bradykinin (BK) B(2) receptors (B(2)Rs). After ample washing (3 hours), the antagonism exerted by these peptides was not reversible. By contrast, the antagonist LF 16. 0687 (30 to 100 nmol/L) was competitive and reversible. A rabbit B(2)R-green fluorescent protein (B(2)R-GFP) conjugate was expressed in mammalian cells. In COS-1 cells, it exhibited an affinity for [3H]BK (K(D)=1.61 nmol/L) similar to that of the wild-type rabbit B(2)R. The stably expressed construction in HEK-293 cells was functionally active (phospholipase A(2) assay), and the antagonists mentioned above retained their respective surmountable or insurmountable behavior. Competition of [(3)H]BK binding to B(2)R-GFP by the antagonists or BK was largely reversible after a 3-hour washout period at 0 degrees C; at 37 degrees C, icatibant or NPC 17731 effects were not reversible. B(2)R-GFP was visualized in the plasma membranes of HEK-293 cells and rapidly internalized in response to BK. NPC 17731 or icatibant slowly translocated B(2)R GFP into cells over 24 hours, whereas LF 16.0687 had no effect on the subcellular distribution of B(2)R-GFP. Cell extract immunoblotting with anti-GFP antibodies revealed a 101- to 105-kDa protein that was not significantly degraded on 24 hours of cell treatment with any of the ligands but was translocated in part to the 15 000-g pellet of the extract on treatment with BK or the noncompetitive antagonists. NPC 17731 and icatibant are noncompetitive, nonequilibrium antagonists that promote the cellular sequestration of rabbit B(2)R expressed in an heterologous system. PMID- 10856285 TI - Hypertension online only : june 2000 PMID- 10856287 TI - Structure of GATE-16, membrane transport modulator and mammalian ortholog of autophagocytosis factor Aut7p. AB - The GATE-16 protein participates in intra-Golgi transport and can associate with the N-ethylmaleimide-sensitive fusion protein and with Golgi SNAREs. The yeast ortholog of GATE-16 is the autophagocytosis factor Aut7p. GATE-16 is also closely related to the GABA receptor-associated protein (GABARAP), which has been proposed to cluster neurotransmitter receptors by mediating interaction with the cytoskeleton, and to the light chain-3 subunit of the neuronal microtubule associated protein complex. Here, we present the crystal structure of GATE-16 refined to 1.8 A resolution. GATE-16 contains a ubiquitin fold decorated by two additional N-terminal helices. Proteins with strong structural similarity but no detectable sequence homology to GATE-16 include Ras effectors that mediate diverse downstream functions, but each interacts with Ras by forming pseudo continuous beta-sheets. The GATE-16 surface suggests that it binds its targets in a similar manner. Moreover, a second potential protein-protein interaction site on GATE-16 may explain the adapter activity observed for members of the GATE-16 family. PMID- 10856288 TI - Selective activation of p38 MAPK cascade and mitotic arrest caused by low level oxidative stress. AB - Apoptosis induced by high level oxidative stress accompanies diverse cellular biochemical events including activation of the stress signal cascades of JNK and NF-kappaB. We report here selective activation of p38 MAPK cascade and mitotic arrest under a low level oxidative stress that lacks apoptosis induction. U937 human lymphoid cells treated with low dose (0.02 mm) H(2)O(2) rapidly caused p38 MAPK cascade activation detectable by phosphorylation of MKK3/6, p38 MAPK, activating transcription factor-2, and cAMP-responsive element-binding protein, leaving the JNK and NF-kappaB cascades unaffected. The p38 kinase activation was sustained for 24 h under the low level stress conditions and led to formation of polyploid nuclei. N-Acetyl-l-cysteine, a precursor of anti-oxidant glutathione, canceled both p38 MAPK activation and abnormal cell cycle progression, whereas blockage of the kinase by specific inhibitor SB203580 allowed the appearance of apoptotic cells. Thus, mimicking the effects of nocodazole, the low level oxidative stimulus caused inhibition of cell division in the M phase through p38 MAPK activation. The kinase cascade may serve as a primary transducer of cytoplasmic oxidative signals to nucleus for stress-relieving gene expression and cell cycle control before apoptosis-inducing signals are transduced. This is the first report demonstrating that oxidative stress can participate in cell cycle control by induction of a signal cascade. PMID- 10856289 TI - Activation of the human asparagine synthetase gene by the amino acid response and the endoplasmic reticulum stress response pathways occurs by common genomic elements. AB - The human asparagine synthetase (AS) gene is transcriptionally regulated by amino acid deprivation (amino acid response, AAR) and the endoplasmic reticulum stress response (ERSR), also known as the unfolded protein response pathway. The results reported here document the novel observation that induction of the AS gene by the AAR and ERSR pathways occurs via the same set of genomic elements. Data supporting this conclusion include transient transfection of AS promoter/reporter gene constructs that illustrate that the transcriptional control elements used by both pathways are contained with nucleotides -111 to -34 of the AS promoter. In vivo footprinting analysis of this region identified six specific protein-binding sites. Within two of these sites, altered footprinting was observed following amino acid or glucose deprivation, but the patterns were identical for both the AAR and the ERSR pathway. Site-directed mutation of individual nucleotides within these two binding sites confirmed their importance for regulated transcription, and none of the mutations resulted in loss of response of only one pathway. Neither of these two sites corresponds to a recently identified ERSR cis-element, nor do they contain consensus sequences for known transcription factors. Collectively, the data document that there are at least two independent transcriptional mechanisms for gene activation by the ERSR pathway, one of which terminates at the same genomic elements used by the AAR pathway. PMID- 10856290 TI - The role for zinc in replication protein A. AB - Heterotrimeric human single-stranded DNA (ssDNA)-binding protein, replication protein A (RPA), is a central player in DNA replication, recombination, and repair. The C terminus of the largest subunit, RPA70, contains a putative zinc binding motif and is implicated in complex formation with two smaller subunits, RPA14 and RPA32. The C-terminal domain of RPA70 (RPA70-CTD) was characterized using proteolysis and x-ray fluorescence emission spectroscopy. The proteolytic core of this domain comprised amino acids 432-616. X-ray fluorescence spectra revealed that RPA70-CTD possesses a coordinated Zn(II). The trimeric complex of RPA70-CTD, the ssDNA-binding domain of RPA32 (amino acids 43-171), and RPA14 had strong DNA binding activity. When properly coordinated with zinc, the trimer's affinity to ssDNA was only 3-10-fold less than that of the ssDNA-binding domain in the middle of RPA70. However, the DNA-binding activity of the trimer was dramatically reduced in the presence of chelating agents. Our data indicate that (i) Zn(II) is essential to stabilize the tertiary structure of RPA70-CTD; (ii) RPA70-CTD possesses DNA-binding activity, which is modulated by Zn(II); and (iii) ssDNA binding by the trimer is a synergistic effect generated by the RPA70-CTD and RPA32. PMID- 10856291 TI - Human La antigen is required for the hepatitis C virus internal ribosome entry site-mediated translation. AB - The 5'-noncoding region (5'-NCR) of the hepatitis C virus (HCV) RNA genome serves as an internal ribosome entry site (IRES) and mediates translation initiation in a cap-independent manner. Previously, we reported the interaction between La antigen and the HCV IRES, which appeared to occur in the context of initiator AUG. It was further shown that HCV IRES-mediated translation was stimulated in the presence of human La antigen. In this study, we have defined the cis- and trans-acting elements responsible for La-5'-NCR interactions and established the dependence of the HCV IRES efficiency on cellular La antigen. During the La-IRES interaction, initiator AUG but not the neighboring codons was found to be the direct target of La binding. The C terminus effector domain-dependent modulation of La binding to the HCV IRES is demonstrated by deletion and substitution mutagenesis of the protein. An RNA systematic evolution of ligands by exponential enrichment (SELEX), generated against La protein that selectively binds La in HeLa lysates and competes for the protein binding to the 5'-NCR, was used to demonstrate the requirement of La for the HCV IRES function in the context of mono- and dicistronic mRNAs. Sequestration of La antigen by the RNA SELEX in HeLa translation lysates blocked the HCV and poliovirus IRES-mediated translation in vitro. The functional requirement of La protein for the HCV IRES activity was further established in a liver-derived cell line and in an add-back experiment in which the inhibited IRES was rescued by recombinant human La. These results strongly argue for the novel role of La protein during selection of the initiator AUG and its participation during internal initiation of translation of the HCV RNA genome. PMID- 10856292 TI - Regulation of human COL2A1 gene expression in chondrocytes. Identification of C Krox-responsive elements and modulation by phenotype alteration. AB - To identify control motifs involved in human type II collagen gene transcription in both differentiated and dedifferentiated rabbit articular chondrocytes, transient transfection experiments were performed. A 715-base pair (bp) region of the first intron (+2127/+2842), including a 153-bp sequence so far uncharacterized (+2689/+2842), was found to mediate enhancer activity. In dedifferentiated chondrocytes, this enhancer activity was shown to be less effective than in primary cultures but still present. We then demonstrated that a zinc finger protein, C-Krox, activates COL2A1 gene transcription in differentiated chondrocytes through the enhancer region, whereas in subcultured cells, it inhibited the gene activity via a 266-bp promoter. Multicopies of the C Krox binding site were found to mediate transactivation in both primary cultures and passaged cells, whereas C-Krox overexpression inhibited transcription in dedifferentiated chondrocytes. Additionally, we showed that C-Krox binds to several cis sequences that mediate its transcriptional effects. During chondrocyte dedifferentiation, the protein levels and binding activity of C-Krox were reduced, whereas those of NF-kappaB were increased. This was not associated with variations of mRNA levels, suggesting that post-transcriptional regulatory mechanisms could be involved in C-Krox expression. These results suggest that C Krox plays a major role in type II collagen expression and the chondrocyte phenotype modulation. PMID- 10856293 TI - Glycogen synthase kinase 3beta negatively regulates both DNA-binding and transcriptional activities of heat shock factor 1. AB - Stress activation of heat shock factor (HSF1) involves the conversion of repressed monomers to DNA-binding homotrimers with increased transcriptional capacity and results in transcriptional up-regulation of the heat shock protein (hsp) gene family. Cells tightly control the activity of HSF1 through interactions with hsp90 chaperone complexes and through integration into a number of different signaling cascades. A number of studies have shown that HSF1 transcriptional activity is negatively regulated by constitutive phosphorylation in the regulatory domain by glycogen synthase kinase (GSK3) isoforms alpha/beta. However, previous studies have not examined the ability of GSK3 to regulate the DNA-binding activity of native HSF1 in vivo under heat shock conditions. Here we show that GSK3beta inhibits both DNA-binding and transcriptional activities of HSF1 in heat-shocked cells. Specific inhibition of GSK3 increased the levels of DNA binding and transcription after heat shock and delayed the attenuation of HSF1 during recovery. In contrast, the overexpression of GSK3beta resulted in significant reduction in heat-induced HSF1 activities. These results confirm the role of GSK3beta as a negative regulator of HSF1 transcription in cells during heat shock and demonstrate for the first time that GSK3beta functions to repress DNA binding. PMID- 10856294 TI - Lipopolysaccharide inhibits long term potentiation in the rat dentate gyrus by activating caspase-1. AB - Lipopolysaccharide, a component of the cell wall of Gram-negative bacteria, may be responsible for at least some of the pathophysiological sequelae of bacterial infections, probably by inducing an increase in interleukin-1beta (IL-1beta) concentration. We report that intraperitoneal injection of lipopolysaccharide increased hippocampal caspase-1 activity and IL-1beta concentration; these changes were associated with increased activity of the stress-activated kinase c Jun NH(2)-terminal kinase, decreased glutamate release, and impaired long term potentiation. The degenerative changes in hippocampus and entorhinal cortical neurones were consistent with apoptosis because translocation of cytochrome c and poly(ADP-ribose) polymerase cleavage were increased. Inhibition of caspase-1 blocked these changes, suggesting that IL-1beta mediated the lipopolysaccharide induced changes. PMID- 10856295 TI - Junctional adhesion molecule interacts with the PDZ domain-containing proteins AF 6 and ZO-1. AB - We have identified the PDZ domain protein AF-6 as an intracellular binding partner of the junctional adhesion molecule (JAM), an integral membrane protein located at cell contacts. Binding of AF-6 to JAM required the presence of the intact C terminus of JAM, which represents a classical type II PDZ domain-binding motif. Although JAM did not interact with the single PDZ domains of ZO-1 or of CASK, we found that a ZO-1 fragment containing PDZ domains 2 and 3 bound to JAM in vitro in a PDZ domain-dependent manner. AF-6 as well as ZO-1 could be coprecipitated with JAM from endothelial cell extracts, demonstrating the association of the endogenously expressed molecules in vivo. Targeting of JAM to sites of cell contacts could be affected by the loss of the PDZ domain-binding C terminus. Full-length mouse JAM co-distributed with endogenous AF-6 in human Caco 2 cells at sites of cell contact independent of whether adjacent cells expressed mouse JAM as an extracellular binding partner. In contrast, truncated JAM lacking the PDZ domain-binding C terminus did not co-distribute with endogenous AF-6, but was restricted to cell contacts between cells expressing mouse JAM. Our results suggest that JAM can be recruited to intercellular junctions by its interaction with the PDZ domain-containing proteins AF-6 and possibly ZO-1. PMID- 10856296 TI - DNA-damaging aryl hydrocarbons induce Mdm2 expression via p53-independent post transcriptional mechanisms. AB - During previous studies, we found that mdm2 mRNA levels were elevated in benzo[a]pyrene (BaP, a polycyclic aryl hydrocarbon)-treated cells under conditions of DNA damage-induced cell cycle arrest (Vaziri, C., and Faller, D. V. (1997) J. Biol. Chem. 272, 2762-2769). We have identified potential aryl hydrocarbon receptor-binding sites in the mdm2 promoter. However, we show that induction of mdm2 mRNA by BaP is entirely dependent upon aryl-hydrocarbon-induced genotoxicity and does not involve direct aryl-hydrocarbon receptor-mediated transcriptional activation of the mdm2 gene. Heterologous mdm2 promoter-reporter constructs containing p53-response elements were not responsive to BaP treatment. Therefore the p53-response elements in the mdm2 promoter are insufficient to confer DNA damage-dependent expression of mdm2. Furthermore, mdm2 transcripts were induced by BaP in p53 null cells from transgenic mice (although both basal and BaP-induced mdm2 expression levels were reduced in these cells relative to p53(+/+) cultures). These data show that p53-mediated mechanisms cannot account for BaP/DNA damage-induced mdm2 expression. Mdm2 promoter-reporter gene assays and nuclear run-off analyses of nascent mdm2 transcripts showed that transcriptional induction was unable to account for the large changes in mdm2 transcript levels following BaP treatment. However, mdm2 mRNA half-life measurements showed stabilization of the mdm2 transcript (from approximately 1 h to >4 h) in response to BaP. To our knowledge, this is the first report of control of mdm2 at the post-transcriptional level and in a p53-independent manner. Transient ectopic expression of mdm2 strongly augmented aryl-hydrocarbon induced apoptosis, demonstrating that mdm2 levels can have a profound effect on the cellular response to DNA damage. Overall, our results suggest a potentially important link between DNA damage signaling and RNA stability that may be relevant to cell cycle regulation, tumor suppression, and environmental carcinogenesis. PMID- 10856297 TI - Peptide specificity of RT1-A1(c), an inhibitory rat major histocompatibility complex class I natural killer cell ligand. AB - The rat major histocompatibility complex class Ia allelomorph RT1-A1(c) is a potent ligand for the recently identified inhibitory rLy-49 receptor, STOK-2. With the ultimate objective of studying the interactions of these molecules using structural and functional methods, we undertook a detailed study of its peptide specificity. The study revealed that designing an "ideal peptide" by choosing the most abundant residues in the "binding motif" obtained by pool sequencing does not necessarily yield an optimal binding peptide. For RT1-A1(c), as many as four positions, P2, P4, P5, and P9, were detected as putative anchors. Since this molecule displays a preference for highly hydrophobic peptides, we tested binding of peptides derived from the known leader peptide sequences of other rat histocompatibility complex class I molecules. One such peptide, found to bind well, requiring 1.6 microm peptide to achieve 50% stabilization, was searched for in vivo. Natural RT1-A1(c) binding peptides were purified from rat splenocytes and characterized by mass spectrometry using a combined matrix-assisted laser desorption ionization/time-of-flight and quadrupole time-of-flight approach. Results showed that the signal sequence-derived peptide was not detectable in the purified peptide pool, which was composed of a complex spectrum of peptides. Seven of these self-peptides were successfully sequenced. PMID- 10856298 TI - Molecular analysis of murine leukemia cell lines resistant to 5, 10 dideazatetrahydrofolate identifies several amino acids critical to the function of folylpolyglutamate synthetase. AB - Four L1210 murine leukemia cell lines resistant to 5, 10-dideazatetrahydrofolate (DDATHF) and other folate analogs, but sensitive to continuous exposure to methotrexate, were developed by chemical mutagenesis followed by DDATHF selective pressure. Endogenous folate pools were modestly reduced but polyglutamate derivatives of DDATHF and ALIMTA (LY231514, MTA) were markedly decreased in these mutant cell lines. Membrane transport was not a factor in drug resistance; rather, folypolyglutamate synthetase (FPGS) activity was decreased by >98%. In each cell line, FPGS mRNA expression was unchanged but both alleles of the FPGS gene bore a point mutation in highly conserved domains of the coding region. Four mutations were in the predicted ATP-, folate-, and/or glutamate-binding sites of FPGS, and two others were clustered in a peptide predicted to be beta sheet 5, based on the crystal structure of the Lactobacillus casei enzyme. Transfection of cDNAs for three mutant enzymes into FPGS-null Chinese hamster ovary cells restored a reduced level of clonal growth, whereas a T339I mutant supported growth at a level comparable to that of the wild-type enzyme. The two mutations predicted to be in beta sheet 5, and one in the loop between NH(2)- and COOH terminal domains did not support cell growth. When sets of mutated cDNAs were co transfected into FPGS-null cells to mimic the genotype of drug-selected resistant cells, clonal growth was restored. These results demonstrate for the first time that single amino acid substitutions in several critical regions of FPGS can cause marked resistance to tetrahydrofolate antimetabolites, while still allowing cell survival. PMID- 10856299 TI - Mutation of conserved aspartates affects maturation of both aspartate mutant and endogenous presenilin 1 and presenilin 2 complexes. AB - Presenilin (PS1 and PS2) holoproteins are transiently incorporated into low molecular weight (MW) complexes. During subsequent incorporation into a higher MW complex, they undergo endoproteolysis to generate stable N- and C-terminal fragments. Mutation of either of two conserved aspartate residues in transmembrane domains inhibits both presenilin-endoproteolysis and the proteolytic processing of beta-amyloid precursor protein and Notch. We show that although PS1/PS2 endoproteolysis is not required for inclusion into the higher MW N- and C-terminal fragment-containing complex, aspartate mutant holoprotein presenilins are not incorporated into the high MW complexes. Aspartate mutant presenilin holoproteins also preclude entry of endogenous wild type PS1/PS2 into the high MW complexes but do not affect the incorporation of wild type holoproteins into lower MW holoprotein complexes. These data suggest that the loss of function effects of the aspartate mutants result in altered PS complex maturation and argue that the functional presenilin moieties are contained in the high molecular weight complexes. PMID- 10856300 TI - Activation of ATF6 and an ATF6 DNA binding site by the endoplasmic reticulum stress response. AB - ATF6 is a member of the basic-leucine zipper family of transcription factors. It contains a transmembrane domain and is located in membranes of the endoplasmic reticulum. ATF6 has been implicated in the endoplasmic reticulum (ER) stress response pathway since it can activate expression of GRP78 and other genes induced by the ER stress response. ER stress appears to activate ATF6 by cleavage from the ER membrane and translocation to the nucleus. However, direct DNA binding by ATF6 had not been demonstrated. In this report, we have identified a consensus DNA binding sequence for ATF6. This site is related to but distinct from ATF1/CREB binding sites. The site was placed in a reporter gene and was specifically activated by ATF6 overexpression and was strongly induced by the ER stress response. A dominant negative form of ATF6 blocked ER stress induction of both ATF6 site and GRP78 reporter genes. We further found that GAL4-ATF6 could be activated by ER stress. These results demonstrate that ATF6 is a direct target of the ER stress response. A proximal sensor of the ER stress response, human IRE1 (hIRE1), was sufficient to activate the ATF6 reporter gene, while a dominant negative form of hIRE1 blocked ER stress activation, suggesting that hIRE1 is upstream of ATF6 in the ER stress signaling pathway. PMID- 10856301 TI - Glycogen synthase association with the striated muscle glycogen-targeting subunit of protein phosphatase-1. Synthase activation involves scaffolding regulated by beta-adrenergic signaling. AB - Glycogen-binding subunits for protein phosphatase-1 (PP1) target the PP1 catalytic subunit (PP1C) to glycogen particles, where the enzymes glycogen synthase and glycogen phosphorylase are concentrated. Here we identify sites within the striated muscle glycogen-binding subunit (G(M)) that mediate direct binding to glycogen synthase. Both PP1C and glycogen synthase were coimmunoprecipitated with a full-length FLAG-tagged G(M) transiently expressed in COS7 cells or C2C12 myotubes. Deletion and mutational analysis of a glutathione S transferase (GST) fusion of the N-terminal domain of G(M) (residues 1-240) identified two putative sites for binding to glycogen synthase, one of which is the WXNXGXNYX(I/L) motif that is conserved among the family of PP1 glycogen binding subunits. Either deletion of this motif or Ala substitution of Asn-228 in this motif disrupted the binding of glycogen synthase. Expression of full-length FLAG-G(M) in cells increased the activity of endogenous glycogen synthase, but protein disabled in either PP1 binding or glycogen synthase binding did not produce synthase activation. The results show that efficient activation of glycogen synthase requires a scaffold function of G(M) that involves simultaneous binding of both PP1C and glycogen synthase. Isoproterenol and forskolin treatment of cells decreased glycogen synthase binding to FLAG-G(M), thereby limiting synthase activation by PP1. This response was insensitive to inhibition by H-89, therefore probably not involving cAMP-dependent protein kinase, but did require inclusion of microcystin-LR during cell lysis, implying that phosphorylation was modulating binding of glycogen synthase. Phosphorylation control of binding to a scaffold site on the G(M) subunit of PP1 offers a new mechanism for regulation of muscle glycogen synthase in response to beta-adrenergic signals. PMID- 10856302 TI - Structure of tuberoinfundibular peptide of 39 residues. AB - The recently identified natural peptide ligand, tuberoinfundibular peptide of 39 residues (TIP39) for the parathyroid hormone-2 (PTH2) receptor has been structurally characterized by high resolution NMR, circular dichroism, and computer simulations. The structural features of TIP39, determined in the presence of a zwitterionic lipid to mimic the membrane environment of the G protein-coupled PTH2 receptor, consist of two alpha-helices, Ala(5)-Arg(21) and Leu(26)-Val(35). Although TIP39 shares limited sequence homology with parathyroid hormone (PTH), a comparison of the structural features of TIP39 and PTH illustrates a similar topological display of residues of the N-terminal helix important for PTH2 receptor activation. The C-terminal helix of TIP39 differs from that of PTH with respect to size and amphipathicity, suggesting an altered mode of binding for TIP39, consistent with the receptor chimera and ligand truncation studies presented in the accompanying paper (Hoare, S. R. J., Clark, J. A., and Usdin, T. B. (2000) J. Biol. Chem. 275, 27274-27283). The structural characterization of TIP39 also provides some insight into the lack of affinity of this novel ligand for the PTH1 receptor. PMID- 10856303 TI - Purification and mass spectrometry of six lipid A species from the bacterial endosymbiont Rhizobium etli. Demonstration of a conserved distal unit and a variable proximal portion. AB - Lipid A of Rhizobium etli CE3 differs dramatically from that of other Gram negative bacteria. Key features include the presence of an unusual C28 acyl chain, a galacturonic acid moiety at position 4', and an acylated aminogluconate unit in place of the proximal glucosamine. In addition, R. etli lipid A is reported to lack phosphate and acyloxyacyl residues. Most of these remarkable structural claims are consistent with our recent enzymatic studies. However, the proposed R. etli lipid A structure is inconsistent with the ability of the precursor (3-deoxy-D-manno-octulosonic acid)(2)-4'-(32)P-lipid IV(A) to accept a C28 chain in vitro (Brozek, K. A., Carlson, R. W., and Raetz, C. R. H. (1996) J. Biol. Chem. 271, 32126-32136). To re-evaluate the structure, CE3 lipid A was isolated by new chromatographic procedures. CE3 lipid A is now resolved into six related components. Aminogluconate is present in D-1, D-2, and E, whereas B and C contain the typical glucosamine disaccharide seen in lipid A of most other bacteria. All the components possess a peculiar acyloxyacyl moiety at position 2', which includes the ester-linked C28 chain. As judged by mass spectrometry, the distal glucosamine units of A through E are the same, but the proximal units are variable. As described in the accompanying article (Que, N. L. S., Ribeiro, A. A., and Raetz, C. R. H. (2000) J. Biol. Chem. 275, 28017-28027), the discovery of component B suggests a plausible enzymatic pathway for the biosynthesis of the aminogluconate residue found in species D-1, D-2, and E of R. etli lipid A. We suggest that the unusual lipid A species of R. etli might be essential during symbiosis with leguminous host plants. PMID- 10856304 TI - Two-dimensional NMR spectroscopy and structures of six lipid A species from Rhizobium etli CE3. Detection of an acyloxyacyl residue in each component and origin of the aminogluconate moiety. AB - The chemical structures of six lipid A species (A, B, C, D-1, D-2, and E) purified from Rhizobium etli CE3 were investigated by one- and two-dimensional NMR spectroscopy. The R. etli lipid A subtypes each contain an unusual acyloxyacyl residue at position 2' as part of a conserved distal glucosamine moiety but differ in their proximal units. All R. etli lipid A species lack phosphate groups. However, they are derivatized with an alpha-linked galacturonic acid group at position 4', as shown by nuclear Overhauser effect spectroscopy. Component B, which had been not been reported in previous studies, features a beta, 1'-6 linked disaccharide of glucosamine acylated at positions 2, 3, 2', and 3' in a pattern that is typical of lipid A found in other Gram-negative bacteria. D-1 contains an acylated aminogluconate unit in place of the proximal glucosamine residue of B. C and E lack ester-linked beta-hydroxyacyl chains at position 3, as judged by their H-3 chemical shifts, and may be synthesized from B and D-1, respectively, by the R. etli 3-O-deacylase. D-2 is an isomer of D-1 that forms nonenzymatically by acyl chain migration. A may be an elimination product derived from D-1 during hydrolysis at 100 degrees C (pH 4.5), a step needed to release lipid A from lipopolysaccharide. Based on these findings, we propose a biosynthetic scheme for R. etli lipid A in which B is generated first by a variation of the E. coli pathway. The aminogluconate unit of D-1 could then be made from B by enzymatic oxidation of the proximal glucosamine. As predicted by our hypothesis, enzyme(s) can be demonstrated in extracts of R. etli that convert (14)C-labeled B to D-1. PMID- 10856305 TI - Protein kinase C phosphorylation of syntaxin 4 in thrombin-activated human platelets. AB - We postulated that the syntaxins, because of their key role in SNARE complex formation and exocytosis, could be important targets for signaling by intracellular kinases involved in secretion. We found that syntaxin 4 was phosphorylated in human platelets treated with a physiologic agent that induces secretion (thrombin) but not when they were treated with an agent that prevents secretion (prostacyclin). Syntaxin 4 phosphorylation was blocked by inhibitors of activated protein kinase C (PKC), and, in parallel assays, PKC inhibitors also blocked secretion from thrombin-activated platelets. In platelets, cellular activation by thrombin or phorbol 12-myristate 13-acetate decreased the binding of syntaxin 4 with SNAP-23, another platelet t-SNARE. Phosphatase inhibitors increased syntaxin 4 phosphorylation and further decreased syntaxin 4-SNAP-23 binding induced by cell activation. Conversely, a PKC inhibitor blocked syntaxin 4 phosphorylation and returned binding of syntaxin 4-SNAP-23 to that seen in nonstimulated platelets. In vitro, PKC directly phosphorylated platelet syntaxin 4 and recombinant syntaxin 4. PKC phosphorylation in vitro inhibited (71 +/- 8%) the binding of syntaxin 4 to SNAP-23. These results provide evidence that extracellular activation can be coupled through intracellular PKC signaling so as to modulate SNARE protein interactions involved in platelet exocytosis. PMID- 10856306 TI - Generation of truncated C/EBPbeta isoforms by in vitro proteolysis. AB - Multiple forms of the transcriptional regulator CCAAT/enhancer-binding protein beta (C/EBPbeta) with molecular masses of approximately 38, 34, 20, and 14 kDa have been observed in cell extracts. It has been proposed that these proteins arise by alternative initiation at in-frame AUG codons. The truncated C/EBPbeta isoforms (p14 and p20/LIP) lack transactivation domains but retain DNA-binding and dimerization sequences and are therefore assumed to function as competitive inhibitors of C/EBP-mediated transcription in vivo. By comparing various extraction procedures to analyze endogenous and overexpressed C/EBPbeta proteins, we determined that p20-C/EBPbeta is generated predominantly by in vitro proteolytic cleavage during isolation from cells and that p14-C/EBPbeta is produced exclusively by this mechanism. In transfected cells, the full-length (p34 and p38) isoforms but not the truncated proteins were detectable in the cytoplasm, indicating that the latter are not primary translation products. In addition, the C/EBPbeta leucine zipper dimerization domain was essential for the appearance of the truncated species, demonstrating that protein folding or dimerization are critical determinants of proteolytic sensitivity. Our findings suggest that the presence of truncated C/EBPbeta proteins in cell extracts must be interpreted with caution and that assumptions about the in vivo relevance of these isoforms should be re-evaluated. PMID- 10856307 TI - Non-replicating Epstein-Barr virus-based plasmids extend gene expression and can improve gene therapy in vivo. AB - To date, no gene transfer vector has produced prolonged gene expression following a single intravenous injection and then efficiently re-expressed the delivered gene following repeated systemic injection into immunocompetent hosts. To overcome these limitations, a gene therapy regimen using non-replicating Epstein Barr virus (EBV)-based expression plasmids was developed. One plasmid contains the FR (EBV family of repeats) sequence and the expressed gene. The other encodes Epstein-Barr nuclear antigen 1 (EBNA-1), but lacks FR. Although unable to replicate in mice, intravenous co-injection of EBV-based plasmids in cationic liposome-DNA complexes (CLDCs) substantially prolonged luciferase gene expression. The use of a two-vector system limited host exposure to the EBNA-1 gene product. Furthermore, this EBV-based vector system could be intravenously re injected multiple times into immunocompetent mice without loss of transfection efficiency. Use of this vector system significantly improved the therapeutic efficacy of the biologically important human granulocyte colony-stimulating factor gene. Delivery of the human granulocyte colony-stimulating factor gene in EBV-based plasmids increased circulating white blood counts for at least 2 months following a single CLDC-based intravenous co-injection. Conversely, white blood counts were never elevated following injection of CLDCs lacking EBV-derived elements. Thus, this EBV-based plasmid vector system both markedly prolongs gene expression at therapeutic levels and efficiently and repeatedly re-transfects immunocompetent hosts. These properties of EBV-based plasmid vectors appear to be due, at least in part, to the documented abilities of the EBNA-1 protein both to retain FR-containing DNA intracellularly and within the nucleus and to block anti EBNA-1 cytotoxic T cell responses. PMID- 10856308 TI - Angiotensin II initiates tyrosine kinase Pyk2-dependent signalings leading to activation of Rac1-mediated c-Jun NH2-terminal kinase. AB - Ca(2+)-sensitive tyrosine kinase Pyk2 was shown to be involved in angiotensin (Ang) II-mediated activation of extracellular signal-regulated kinase (ERK) via transactivation of epidermal growth factor receptor (EGF-R). In this study, we tested the involvement of Pyk2 and EGF-R in Ang II-induced activation of JNK and c-Jun in cardiac fibroblasts. Ang II markedly stimulated JNK activities, which were abolished by genistein and intracellular Ca(2+) chelators but partially by protein kinase C depletion. Inhibition of EGF-R did not affect Pyk2 and JNK activation by Ang II. Stable transfection with a dominant negative (DN) mutant for Pyk2 (PKM) completely blocked JNK activation by Ang II. DN mutants of Rac1 (DN-Rac1) and MEK kinase (DN-MEKK1) also abolished it, whereas those of Cdc42, RhoA, and Ha-Ras had no effect. Induction of c-Jun gene transcription by Ang II was abolished in PKM, DN-Rac1, and DN-MEKK1, in which Ang II-induced binding of ATF2/c-Jun heterodimer to the activator protein-1 sequence at -190 played a key role. These results suggest that 1) in cardiac fibroblasts activation of JNK and c-Jun by Ang II is initiated by Pyk2-dependent signalings but not by downstream signals of EGF-R or Ras, 2) Rac1 but not Cdc42 is required for JNK activation by Ang II upstream of MEKK1, and 3) ATF-2/c-Jun binding to the activator protein-1 sequence at -190 plays a key role for induction of c-Jun gene by Ang II. PMID- 10856309 TI - Supplementary oxygen therapy in COPD: is it really useful? PMID- 10856310 TI - Supplemental oxygen during pulmonary rehabilitation in patients with COPD with exercise hypoxaemia. AB - BACKGROUND: Supplemental oxygen in patients with chronic obstructive pulmonary disease (COPD) and exercise hypoxaemia improves exercise capacity and dyspnoea. However, the benefit of oxygen during pulmonary rehabilitation in these patients is still unknown. METHODS: Twenty five patients with stable COPD (mean (SD) forced expiratory volume in one second (FEV(1)) 0.76 (0.29) l and 30.0 (9.89)% predicted, arterial oxygen tension (PaO(2)) 8.46 (1.22) kPa, arterial carbon dioxide tension (PaCO(2)) 6.32 (1.01) kPa) and significant arterial desaturation on exercise (82.0 (10.4)%) were entered onto a pulmonary rehabilitation programme. Patients were randomised to train whilst breathing oxygen (OT) (n = 13) or air (AT) (n = 12), both at 4 l/min. Assessments included exercise tolerance and associated dyspnoea using the shuttle walk test (SWT) and Borg dyspnoea score, health status, mood state, and performance during daily activities. RESULTS: The OT group showed a significant reduction in dyspnoea after rehabilitation compared with the AT group (Borg mean difference -1.46 (95% CI -2.72 to -0.19)) but there were no differences in other outcome measures: SWT difference -23.6 m (95% CI -70.7 to 23.5), Chronic Respiratory Disease Questionnaire 3.67 (95% CI -7.70 to 15.1), Hospital Anxiety and Depression Scale 1. 73 (95% CI -2.32 to 5.78), and London Chest Activity of Daily Living Scale 2.18 (95% CI -7.15 to 2.79). At baseline oxygen significantly improved SWT (mean difference 27.3 m (95% CI 14.7 to 39.8) and dyspnoea (-0.68 (95% CI -1.05 to 0.31)) compared with placebo air. CONCLUSIONS: This study suggests that supplemental oxygen during training does little to enhance exercise tolerance although there is a small benefit in terms of dyspnoea. Patients with severe disabling dyspnoea may find symptomatic relief with supplemental oxygen. PMID- 10856311 TI - A pragmatic assessment of the placement of oxygen when given for exercise induced dyspnoea. AB - BACKGROUND: It is uncertain whether patients with chronic obstructive pulmonary disease (COPD) given oxygen for symptom relief should be advised to use it before or after exertion. METHODS: Eighteen patients with smoking related COPD who desaturated on exercise were given oxygen or air from a cylinder in a single blind manner and in an order determined by Latin square randomisation, before and after ascending stairs. The time of ascent, desaturation, and dyspnoea associated with the ascent was compared across the treatment groups. RESULTS: Oxygen given before or after the ascent reduced maximal dyspnoea from 49.1 mm (95% CI 35.5 to 62.7) to 41.7 mm (95% CI 34.3 to 49.1) of a 100 mm visual analogue scale, reduced desaturation (oxygen before 4.9% (95% CI 3.6 to 6.2), oxygen after 6. 4% (5.3 to 7.5), air before and after 8.2% (6.7 to 9.7%)), but did not affect time of ascent (air before: 5.1 s (95% CI 3.4 to 6.9) reduction from training ascent; oxygen before: 6.1 s (95% CI 2.9 to 9.2) reduction). CONCLUSIONS: Oxygen prescribed for symptomatic relief of dyspnoea benefits selected patients with COPD, but it seems not to matter whether it is taken before or after exertion. PMID- 10856312 TI - Inspiratory pressure support prolongs exercise induced lactataemia in severe COPD. AB - BACKGROUND: A physiological benefit from pulmonary rehabilitation in chronic obstructive pulmonary disease (COPD) is more probable if exercise is performed above the lactate threshold. This study was undertaken to investigate whether it was possible to extend the lactataemia of exercise using non-invasive inspiratory pressure support (IPS). METHODS: Plasma lactate levels were measured in eight men with severe COPD who performed two treadmill walks at an identical constant work rate to a condition of severe dyspnoea; the second walk was supported by IPS. RESULTS: Mean plasma lactate levels before the free and IPS assisted walks were 1.65 mmol/l and 1. 53 mmol/l, respectively (p = NS). Lactate levels increased during both walks to 2.96 mmol/l and 2.42 mmol/l, respectively (p = 0.01 for each) but the duration of the IPS assisted walk was significantly greater than the free walk (13.6 minutes versus 5.5 minutes, p = 0.01). CONCLUSIONS: Patients with severe COPD can sustain exercise induced lactataemia for longer if assisted with IPS. This technique may prove to be a useful adjunct in pulmonary rehabilitation. PMID- 10856313 TI - One year period prevalence study of respiratory acidosis in acute exacerbations of COPD: implications for the provision of non-invasive ventilation and oxygen administration. AB - BACKGROUND: Non-invasive ventilation (NIV) reduces mortality and intubation rates in patients with chronic obstructive pulmonary disease (COPD) admitted to hospital with respiratory acidosis. This study aimed to determine the prevalence of respiratory acidosis in patients admitted with COPD, to draw inferences about oxygen therapy, and to determine the need for NIV services for acute COPD in typical UK hospitals. METHODS: This one year prospective prevalence study identified patients with COPD aged 45-79 years inclusive who were admitted to Leeds General Infirmary, St James's University, and Killingbeck Hospitals, Leeds between 1 March 1997 and 28 February 1998. The prevalence of respiratory acidosis and the relationship with oxygenation are described. Other outcomes included intensive care use and in hospital mortality. From this data population prevalence estimates were determined for respiratory acidosis, from which the need for NIV in a typical district general hospital was modelled. RESULTS: 983 patients were admitted, 11 of whom required immediate intubation. 20% of the remaining 972 had a respiratory acidosis. Acidosis was associated with subsequent admission to the intensive care unit (ICU): pH<7.25, OR 6.10 (95% confidence interval (CI) 1.19 to 31.11); pH 7.25-7.30, OR 8.73 (95% CI 2.11 to 36.06). pH was inversely correlated with arterial oxygen tension (PaO(2)) in the 47% of patients who were hypercapnic, with a PaO(2) of >10 kPa being associated with acidosis in most hypercapnic patients. 80% remained acidotic after initial treatment, giving an age/sex specific prevalence for England and Wales of 75 (95% CI 61 to 90)/100 000/year for men aged 45-79 years and 57 (95% CI 46 to 69)/100 000/year for women. Modelling the need for NIV for all COPD patients indicates that a typical UK hospital will admit 90 patients per year with acidosis of which 72 will require NIV. CONCLUSIONS: In patients with acute COPD the PaO(2) should be maintained at 7.3-10 kPa (SaO(2) 85-92%) to avoid the dangers of hypoxia and acidosis. If all COPD patients with a respiratory acidosis (pH<7.35) after initial treatment are offered NIV, a typical UK hospital will treat 72 patients per year. PMID- 10856314 TI - Atopy, lung function, and obstructive airways disease after prenatal exposure to famine. AB - BACKGROUND: Associations have been found between a large head size at birth and atopy, and between low birth weight and obstructive airways disease. A study was undertaken of people born around the time of the Dutch famine in 1944-5 to determine the effects of maternal malnutrition during specific periods of gestation on the prevalence of obstructive airways disease and atopy. METHODS: Nine hundred and twelve people aged about 50, born at term between November 1943 and February 1947 in Amsterdam, were asked about their medical history. Lung function was measured in 733 and serum concentrations of total IgE and specific IgE against mite, pollen and cat were measured in 726. Those exposed in late, mid, and early gestation (exposed participants) were compared with those born before or conceived after the famine (non-exposed participants). RESULTS: Exposure to famine during gestation affected neither the concentrations of total or specific IgE nor lung function values. The prevalence of obstructive airways disease was increased in people exposed to famine in mid gestation (odds ratio adjusted for sex 1.7, 95% confidence interval (CI) 1.1 to 2.6) and tended to be higher in those exposed in early gestation (odds ratio 1.5, 95% CI 0. 9 to 2.6). CONCLUSIONS: The observed increase in the prevalence of obstructive airways disease in people exposed to famine in mid and early gestation was not parallelled by effects on IgE concentrations or lung function. The link between exposure to famine in mid and early gestation and obstructive airways disease in adulthood suggests that fetal lungs can be permanently affected by nutritional challenges during periods of rapid growth. PMID- 10856315 TI - Underdiagnosis of asthma: is the doctor or the patient to blame? The DIMCA project. AB - BACKGROUND: It is important to diagnose asthma at an early stage as early treatment may improve the prognosis in the long term. However, many patients do not present at an early stage of the condition so the physician may have difficulty with the diagnosis. A study was therefore undertaken to compare the proportion of patients who underpresented their respiratory symptoms with the proportion of underdiagnosed cases of asthma by the general practitioner (GP). A secondary aim was to investigate whether bad perception of dyspnoea by the patient was a determining factor in the underpresentation of asthma symptoms to the GP. METHODS: A random sample of 1155 adult subjects from the general population in the eastern part of the Netherlands was screened for respiratory symptoms and lung function and the results were compared with the numbers of asthma related consultations registered in the medical files of the GP. In subjects with reduced lung function the ability to perceive dyspnoea was investigated during a histamine provocation test in subjects who did and did not report their symptoms to their GP. RESULTS: Of the random sample of 1155 subjects 86 (7%) had objective airflow obstruction (forced expiratory volume in one second (FEV(1)) below the reference value corrected for age, length, and sex minus 1.64SD on two occasions) and had symptoms suggestive of asthma. Of these 86 subjects only 29 (34%) consulted the GP, which indicates underpresentation by 66% of patients. Of all subjects with objective airflow obstruction who presented to their GP with respiratory symptoms, 23 (79%) were recorded in the medical files as having asthma, indicating underdiagnosis by the GP in 21% of cases. Of the subjects with objective airflow obstruction who visited the GP with respiratory symptoms 6% had bad perception of dyspnoea compared with 26% of those who did not present to the GP in spite of airflow obstruction (chi(2) = 3.02, p = 0.08). CONCLUSIONS: Underpresentation to GPs of respiratory symptoms by asthmatic patients contributes significantly to the problem of underdiagnosis of asthma. Underdiagnosis by the GP seems to play a smaller role. Furthermore, there are indications that underpresentation of symptoms by the patient is at least partly explained by a worse perception of dyspnoea. PMID- 10856316 TI - Factors associated with hospital admissions and repeat emergency department visits for adults with asthma. AB - BACKGROUND: A small proportion of patients with asthma account for a disproportionate number of acute health service events. To identify whether factors other than severity and low socioeconomic status were associated with this disproportionate use, a prospective study was undertaken to examine management and psychosocial factors associated with increased risk for admission to hospital with asthma and repeat visits to the emergency department over a 12 month period. METHODS: A total of 293 patients with moderate or severe asthma managed at least in part at two teaching hospitals completed surveys of clinical status, acute events, sociodemographic, and psychological variables. RESULTS: Twenty three percent had a single admission to hospital and 16% had two or more hospital admissions. Twenty six percent had one emergency department visit and 32% had two or more visits to the emergency department. In a multiple logistic regression model, adjusted for age, sex, education and income, odds ratios (95% CI) for baseline factors associated with hospital admissions over the next 12 months were: moderate severity compared with severe asthma 0.6 (0.2 to 0.9); no hospital admissions in the past 12 months 0.1 (0.01 to 0.2); not possessing a written asthma action plan 4.0 (1.5 to 10.7); less use of an avoidance coping style 0.4 (0.3 to 0.7); lower preferences for autonomy in asthma management decisions 1.4 (0.96 to 2.0). Adjusted odds ratios (95% CI) for repeat emergency department visits were: moderate asthma severity 0.3 (0.1 to 0.8); current regular use of oral corticosteroids 10.0 (3.1 to 32.4); a hospital admission in the past 12 months 2.9 (1.8 to 4.8); not possessing a written asthma action plan 2.2 (1.1 to 5.6); less dislike of asthma medications 0.7 (0.5 to 0.9). CONCLUSIONS: In addition to factors relating to severity, not possessing a written asthma action plan, avoidance coping, and attitudes to self-management were related to acute use of health services in this at risk group. Interventions need to address or take these factors into account to reduce asthma morbidity. PMID- 10856317 TI - Oestrogen metabolism in lymphangioleiomyomatosis: catechol-O-methyltransferase pathway is not involved. AB - BACKGROUND: Lymphangioleiomyomatosis (LAM) is an uncommon lung disease for which no effective method of treatment has been found. The predilection of LAM for premenopausal women has led to the assumption that hormonal factors play an important role in the pathogenesis of this disease. The aim of this study was to determine if women with LAM manifest alterations in the catechol-O methyltransferase (COMT) pathway which is essential for preventing the generation of oestrogen derived reactive oxygen species (ROS). METHODS: Blood samples were collected from 15 women with LAM and compared with appropriate controls. The distribution of high and low activity alleles of COMT was determined with a PCR based RFLP assay. The enzymatic activity of COMT was measured in each sample and the potential presence of a circulating inhibitor of COMT was determined. Since an alteration in the COMT pathway could increase the oxidative stress, the plasma concentration of malondialdehyde (MDA), a secondary product generated from lipid peroxidation, has been used as an internal marker. RESULTS: The distribution of high and low activity alleles of COMT (named COMT(HH), COMT(LL), and COMT(HL)) was similar in the two groups with proportions of 40%, 7%, and 53%, respectively, in the women with LAM and 38%, 6%, and 56% in the control subjects. The mean (SD) COMT activity was 24.2 (12.3) pmol/min/mg protein in women with LAM and 24.1 (6.3) pmol/min/mg protein in the control group. Incubation of plasma from women in the two groups with a preparation of commercial COMT showed that no detectable COMT inhibitor was present. The plasma concentration of MDA in the women with LAM was also not significantly different from control subjects. CONCLUSIONS: This study shows that there are no significant alterations in the COMT pathway of women with LAM. It is therefore unlikely that alterations in oestrogen mediated cell signalling pathways are mediated by oxidants derived from an excess of catecholoestrogens in LAM. PMID- 10856318 TI - Enhanced binding of Aspergillus fumigatus spores to A549 epithelial cells and extracellular matrix proteins by a component from the spore surface and inhibition by rat lung lavage fluid. AB - BACKGROUND: Aspergillus fumigatus is a pathogenic fungus which causes a range of diseases, particularly in the human lung. The pathological mechanism is unknown but may involve a complex mixture of biomolecules which can diffuse from the spore surface. This material is known as A fumigatus diffusate (AfD) and has previously been shown to have a range of immunosuppressive functions. It is hypothesised that AfD may influence the binding of spores to extracellular matrix (ECM) proteins and lung epithelial cells, thereby affecting the ability of the fungus to cause infection. METHODS: The binding of spores to ECM proteins and to epithelial cells was carried out using a direct binding assay in microtitre plates and spores were counted by phase contrast microscopy. Rat bronchoalveolar lavage (BAL) fluid was enriched for surfactant protein D (SP-D) using maltose agarose affinity chromatography. The effects of AfD and the SP-D enriched BAL fluid were assessed by pre-incubation with ECM proteins or epithelial cells in the direct binding assay. RESULTS: AfD enhanced the binding of spores to laminin by 137% and to A549 epithelial cells by 250%. SP-D enriched BAL fluid inhibited spore binding to ECM proteins and epithelial cells. Pre-incubation of ECM proteins and epithelial cells with SP-D enriched BAL fluid prevented the enhancement of spore binding by AfD, and pre-incubation of ECM proteins and epithelial cells with AfD prevented the inhibition of spore binding by SP-D enriched BAL fluid. This pretreatment did not prevent the enhancement of spore binding, giving an increase of 95% for collagen I, 80% for fibronectin, 75% for laminin, and 150% for A549 cells. CONCLUSIONS: The hypothesis that AfD would affect spore binding to ECM proteins and epithelial cells was confirmed. Rat BAL fluid, with SP-D as the possible bioactive agent, prevented this enhancement. The in vivo significance is unclear but the enhanced binding of spores may increase the chance of fungal infection in the lung which could be prevented by the protective effects of lung surfactant components (possibly SP-D). The results suggest that there may be competition between AfD and a BAL fluid component (possibly SP-D) for the same or similar binding sites on ECM proteins and epithelial cells. Whether this competition occurs in vivo requires further investigation. PMID- 10856319 TI - Introduction PMID- 10856320 TI - Pediatric origins of adult lung disease. 1. The contribution of airway development to paediatric and adult lung disease. AB - In summary, factors that affect airway growth early in development appear to cause physiological effects that can be persistent. Reduced airway function early in life does not necessarily result in persistent symptoms, but the long term effects and impact on the development of chronic airflow limitation in adults are yet to be determined. Generally, long term sequelae seem to be related to the severity of the initial insult, but the development of persistent increased bronchial responsiveness is an independent risk factor for symptoms and abnormal lung function in later life. In addition, there appear to be separate genetic factors that influence atopy, airway development, and bronchial responsiveness. PMID- 10856321 TI - Interactions between corticosteroids and beta agonists. PMID- 10856323 TI - Alpha-1-antitrypsin deficiency: what next? PMID- 10856324 TI - Anaemia in lung transplant patient caused by parvovirus B19. AB - The case history is presented of a lung transplant patient who developed prolonged parvovirus B19 infection with severe transfusion dependent anaemia. The patient was treated with intravenous immunoglobulin after which the haemoglobin rose, together with a reticulocytosis. The patient then remained transfusion free and the virus cleared more than three months after the initial immunoglobulin treatment. The clinical and social implications for this group of patients are discussed. PMID- 10856322 TI - Molecular mechanisms of glucocorticoid action: what is important? PMID- 10856325 TI - Pneumothorax in adults with cystic fibrosis dependent on nasal intermittent positive pressure ventilation (NIPPV): a management dilemma. AB - The management of pneumothorax in three adult patients with cystic fibrosis dependent on nasal intermittent positive pressure ventilation is described. PMID- 10856326 TI - A case of Haemophilus parainfluenzae pneumonia. AB - A 41 year old woman presented with community acquired pneumonia (CAP) which failed to resolve following treatment with amoxycillin and cefaclor prior to referral. Quantitative culture of sputum revealed a pure growth of Haemophilus parainfluenzae and, following antibiotic susceptibility testing of the isolate, ciprofloxacin was prescribed resulting in resolution of the infection. Immunological investigations showed that the patient had a high titre of H parainfluenzae specific IgM. The combination of a pure growth of H parainfluenzae, a response to appropriate antimicrobial therapy, and the presence of a specific antibody response indicated that this organism had a pathogenic role in the patient's pneumonia and should be considered in the differential diagnosis of CAP. PMID- 10856327 TI - Subacute hypersensitivity pneumonitis in an HIV infected patient receiving antiretroviral therapy. AB - Abnormal pulmonary immune response to various antigens can lead to hypersensitivity pneumonitis. This disease has not previously been reported in HIV infected patients. This case report describes an HIV infected woman who developed subacute hypersensitivity pneumonitis in response to bird exposure. The disease manifested itself only after the patient experienced an improvement in her CD4 positive T lymphocyte count secondary to antiretroviral therapy. This case emphasises the need to consider non-HIV associated diseases in patients with HIV and suggests that diseases in which host immune response plays an essential role in pathogenesis may become more prevalent in HIV infected patients receiving effective antiretroviral therapy. PMID- 10856328 TI - Pharmacology of asthma PMID- 10856329 TI - Obituary PMID- 10856330 TI - Effect of additional carbon sources on biodegradation of phenol. PMID- 10856331 TI - Role of Acinetobacter for biodegradability of quaternary ammonium compounds. PMID- 10856332 TI - Biokinectic parameter investigation and biological treatment of coffee berry effluents. PMID- 10856333 TI - Photodegradation of the herbicides butachlor and ronstar using natural sunlight and diethylamine. PMID- 10856334 TI - Soil ergosterol, dimethyl sulphide reduction, and microbial biomass along a Zn concentrations gradient in soils from a mine spoil tip. PMID- 10856335 TI - Comparison of fluoride ion-selective electrode based potentiometric methods of fluoride determination in human urine. PMID- 10856336 TI - Changes in hepatic cytochrome P450 enzymes by biodegradation products of 4-tert octylphenol polyethoxylate in rats. PMID- 10856337 TI - Organochlorine pesticides and polychlorinated biphenyls in ambient air collected in Zagreb, Croatia. PMID- 10856338 TI - Aliphatic aldehydes produced by heating Chinese cooking oils. PMID- 10856339 TI - Method for the simultaneous extraction and analysis of two current use pesticides, atrazine and lambda-cyhalothrin, in sediment and aquatic plants. PMID- 10856340 TI - Exposure factor and toxicity data for California wildlife: data availability and sources of uncertainty for ecological risk assessments. PMID- 10856341 TI - Time efficient method to renew water in static sediment toxicity tests. PMID- 10856342 TI - Macroalgae as biomonitors of heavy metal availability in coastal lagoons from the subtropical Pacific of Mexico. PMID- 10856343 TI - Stress-protein response in tributyltin-exposed clams. PMID- 10856344 TI - Quantitative structure-activity relationships for the toxicity to the tadpole Rana japonica of selected phenols. PMID- 10856345 TI - Oil pollution of marine algae. PMID- 10856346 TI - Impact of mechanical pulp mill effluent on egg hatchability of brown trout. PMID- 10856370 TI - Low-substrate regulated microaerophilic behavior as a stress response of aquatic and soil bacteria. AB - Low-substrate regulated microaerophilic behavior (LSRMB) was observed in 10-54% of the bacteria isolated from several fresh-water lakes or ponds, subsurface soils, activated sludge, and Antarctic dry valley soils. Five Pseudomonas and two Bacillus type species showed LSRMB. A subsurface Pseudomonas jessenii strain was used as a model to show the metabolic interaction between substrate and oxygen concentrations, cell band movement, and the appearance of unique stress lipids and proteins. When the oxygen in the P. jessenii culture medium was increased from 11% to 100% saturation under atmospheric condition, the concentration of 17:0 cyclopropane fatty acid, a stress indicator, increased five-fold, and four unique proteins were also detected. This stress response occurred only in low substrate media. It is our hypothesis that LSRMB is a common but under appreciated trait of many aquatic and soil bacteria. PMID- 10856371 TI - Phylogeny of marine Bacillus isolates from the Gulf of Mexico. AB - The phylogeny of 11 pigmented, aerobic, spore-forming isolates from marine sources was studied. Forty-two biochemical characteristics were examined, and a 16S rDNA sequence was obtained for each isolate. In a phylogenetic tree based on 16S sequencing, four isolates (NRRL B-14850, NRRL B-14904, NRRL B-14907, and NRRL B-14908) clustered with B. subtilis and related organisms; NRRL B-14907 was closely related to B. amyloliquefaciens. NRRL B-14907 and NRRL B-14908 were phenotypically similar to B. amyloliquefaciens and B. pumilus, respectively. Three strains (NRRL B-14906, NRRL B-14910, and NRRL B-14911) clustered in a clade that included B. firmus, B. lentus, and B. megaterium. NRRL B-14910 was closely related phenotypically and phylogenetically to B. megaterium. NRRL B-14905 clustered with the mesophilic round spore-producing species, B. fusiformis and B. sphaericus; the isolate was more closely related to B. fusiformis. NRRL B-14905 displayed characteristics typical of the B. sphaericus-like organisms. NRRL B 14909 and NRRL B-14912 clustered with the Paenibacillus species and displayed characteristics typical of the genus. Only NRRL B-14851, an unusually thin rod that forms very small spores, may represent a new Bacillus species. PMID- 10856372 TI - Influence of yeast flocculation on the rate of Jerusalem artichoke extract fermentation. AB - Variations in residual sugar composition have been observed during Jerusalem artichoke extract fermentations by using Saccharomyces diastaticus NCYC 625, a flocculating yeast strain. In batch cultures, these differences were due to the inulin polymer size distribution of the extracts: measurements of enzymatic activities on different polymerized substrates have shown that the hydrolysis and fermentation yield decreased when the fructose/glucose ratio of the extract increased. Inulin hydrolysis appeared to be the limiting factor of the fermentation rate. A comparison of continuous and batch cultures with the same extract showed that fermentability differences were related to the structure and size of the yeast flocs. This led to an hydrolysis selectivity of the inulin polymers according to their size: the chemostat culture in which the floc average size was larger gave longer chained residual sugars. PMID- 10856373 TI - wsp gene sequences from the Wolbachia of filarial nematodes. AB - Wolbachia endosymbiotic bacteria are widespread in arthropods and are also present in filarial nematodes. Almost all filarial species so far examined have been found to harbor these endosymbionts. The sequences of only three genes have been published for nematode Wolbachia (i.e., the genes coding for the proteins FtsZ and catalase and for 16S rRNA). Here we present the sequences of the genes coding for the Wolbachia surface protein (WSP) from the endosymbionts of eight species of filaria. Complete gene sequences were obtained from the endosymbionts of two different species, Dirofilaria immitis and Brugia malayi. These sequences allowed us to design general primers for amplification of the wsp gene from the Wolbachia of all filarial species examined. For these species, partial WSP sequences (about 600 base pairs) were obtained with these primers. Phylogenetic analysis groups these nematode wsp sequences into a coherent cluster. Within the nematode cluster, wsp-based Wolbachia phylogeny matches a previous phylogeny obtained with ftsZ gene sequences, with a good consistency of the phylogeny of hosts (nematodes) and symbionts (Wolbachia). In addition, different individuals of the same host species (Dirofilaria immitis and Wuchereria bancrofti) show identical wsp gene sequences. PMID- 10856374 TI - An ACC deaminase minus mutant of Enterobacter cloacae UW4 no longer promotes root elongation. AB - The ACC deaminase gene (acdS) from Enterobacter cloacae UW4 was replaced by homologous recombination with the acdS gene with a tetracycline resistance gene inserted within the coding region. Upon characterization of this AcdS minus mutant, it was determined that both ACC deaminase activity and the ability to promote the elongation of canola roots under gnotobiotic conditions were greatly diminished. This result is consistent with a previously postulated model that suggests that a major mechanism utilized by plant growth-promoting bacteria involves the lowering of plant ethylene levels, and hence ethylene inhibition of root elongation, by bacterial ACC deaminase. PMID- 10856375 TI - Variation in sic gene encoding complement-inhibiting protein of Streptococcus pyogenes serotype M1 isolates in Japan. AB - DNA sequencing of the gene encoding a complement-inhibiting protein of Streptococcus pyogenes (streptococcal inhibitor of complement, Sic) was carried out on 49 strains of S. pyogenes serotype M1. Those strains were obtained from patients and asymptomatic carriers in Japan from 1969 to 1997 and had various pulsed-field gel electrophoresis (PFGE) patterns. Four identical polymorphic sites were found in the strains with the same PFGE pattern (Ia), but not in those giving the pattern IIa. The other identical sites were found in the strains with the PFGE pattern IIa, but not in those with the pattern Ia. These observations suggest that each of PFGE patterns was restricted to a set of variation in the sic gene. PMID- 10856376 TI - High-resolution physical map of the pSymb megaplasmid and comparison of the three replicons of Sinorhizobium meliloti strain 1021. AB - A high-resolution physical map of the larger megaplasmid (pSymb) of Sinorhizobium meliloti strain 1021 has been constructed by using BAC libraries and an original two-step PCR screening method. This method, previously used to map both the chromosome and the smaller megaplasmid (pSyma), allowed us to position over the genome a total of 842 markers with an average density of one marker every 8.3 kb. In addition, we used BLASTX and PRODOM analysis to predict a function for a number of STSs. This work led to the discovery of several interesting loci and to a comparison of the genetic information carried by each replicon. The two main results emerging from this study are (i) a biased distribution of housekeeping genes, mainly detected on chromosome, and (ii) the presence of an unexpected number of transporters, mainly belonging to the ABC superfamily. These are broadly distributed across the whole genome, but particularly found on pSymb. PMID- 10856377 TI - Nitrate reductase and nitrous oxide production by Fusarium oxysporum 11dn1 under aerobic and anaerobic conditions. AB - The fungus Fusarium oxysporum 11dn1 was found to be able to grow and produce nitrous oxide on nitrate-containing medium in anaerobic conditions. The rate of nitrous oxide formation was three to six orders of magnitude lower than the rates of molecular nitrogen production by common denitrifying bacteria. Acetylene and ammonia did not affect the release of nitrous oxide release. It was shown that under anaerobic conditions fast increase of nitrate reductase activity occurred, caused by the synthesis of enzyme de novo and protein dephosphorylation. Reverse transfer of the mycelium to aerobic conditions led to a decline in nitrate reductase activity and stopped nitrous oxide production. The presence of two nitrate reductases was shown, which differed in molecular mass, location, temperature optima, and activity in nitrate- and ammonium-containing media. Two enzymes represent assimilatory and dissimilatory nitrate reductases, which are active in aerobic and anaerobic conditions, respectively. PMID- 10856378 TI - Effects of hydrogel/silver coatings on in vitro adhesion to catheters of bacteria associated with urinary tract infections. AB - Sections of sterile all-silicone-, hydrogel/silver-all-silicone-, and hydrogel/silver-latex-Foley urinary catheters were exposed to suspensions of bacteria and Candida albicans associated with urinary tract infections. The adhesion of these microorganisms to the catheters was determined with a radiolabel-cell procedure and scanning electron microscopy. Anomalous data with the radiolabel procedure were produced with the hydrogel/silver-latex catheters for certain species. These aberrant data were related to adhesion on the untreated cut ends of the latex catheter. Radiolabel-cell-adhesion procedures that involve sections of coated materials may need to be supplemented with additional procedures such as scanning electron microscopy for valid interpretations of the data. Adhesion to the hydrogel/silver catheters by both Gram-positive- and Gram-negative bacteria most commonly associated with nosocomial urinary tract infections, including a strain of Pseudomonas aeruginosa noted for its superior adhesion capacity, was significantly lower than the adhesion to the control all-silicone catheter. PMID- 10856379 TI - Specific inhibition of 50S ribosomal subunit formation in Staphylococcus aureus cells by 16-membered macrolide, lincosamide, and streptogramin B antibiotics. AB - The translational functions of the bacterial ribosome are the target for a large number of antimicrobial agents. The 14- and 16-membered macrolides, the lincosamides, and the streptogramin B type antibiotics are thought to share certain inhibitory properties, based on both biochemical and genetic studies. We have shown previously that the 14-membered macrolides, like erythromycin, have an equivalent inhibitory effect on translation and the formation of the 50S ribosomal subunit in growing bacterial cells. To extend this work, we have now tested the 16-membered macrolides spiramycin and tylosin, the lincosamides lincomycin and clindamycin, and 3 streptogramin B compounds pristinamycin I(A), virginiamycin S, and CP37277. Each of these was a specific inhibitor of 50S subunit formation, in addition to having an inhibitory effect on translation. By contrast, two streptogramin A compounds, virginiamycin M1 and CP36926, as well as chloramphenicol, were effective inhibitors of translation without showing a specific effect on the assembly of the large ribosomal subunit. A combination of an A and B type streptogramin (virginiamycin M1 and pristinamycin I(A)) demonstrated a synergistic inhibition of protein synthesis without exhibiting a specific inhibition of 50S subunit formation. These results extend our observations on 50S assembly inhibition to the entire class of MLS(B) antibiotics and reinforce other suggestions concerning their common ribosome-binding site and inhibitory functions. PMID- 10856380 TI - Adherence of Lactobacillus to intestinal 407 cells in culture correlates with fibronectin binding. AB - Lactobacilli are members of the normal mucosal microflora of most animals. Many isolates of Lactobacillus spp. are adherent to epithelial cells. In this study, using Lactobacillus acidophilus and L. agilis, we detected adherence in a pattern that suggested that the bacteria were binding to extracellular matrix proteins. Fluorescent microscopy, by using anti-fibronectin antibody, demonstrated that the isolates localize in those areas where fibronectin was detected. In addition, fibronectin pretreatment of the bacterial cells decreased adherence to Intestinal 407 epithelial cell monolayers. Cellular binding to fibronectin was detected by enzyme-linked immunosorbent assay and affinity binding to radio-labeled fibronectin. Fibronectin may be one of the eukaryotic receptors mediating attachment of Lactobacillus to mucosal surfaces. PMID- 10856381 TI - Numerical analysis of Astragalus cicer microsymbionts. AB - Thirty-seven rhizobium strains, isolated from root nodules of Astragalus cicer (L.) (cicer milkvetch) deriving from different geographic regions, were compared with the representative strains of the known rhizobial species and genera by numerical analysis of phenotypic characteristics. Our results indicated that Astragalus cicer rhizobia were related to the bacteria of Mesorhizobium species and formed two major phena. One phenon, localized on Mesorhizobium loti branch, contained strains from Poland. Another cluster, placed in the vicinity of M. tianshanense, M. mediterraneum, M. ciceri, and M. huakuii, comprised cicer milkvetch nodule isolates from Canada, Ukraine, and one strain from Poland. The relationship of Astragalus cicer microsymbionts to bacteria of the Mesorhizobium species was also supported by phage typing. PMID- 10856382 TI - Carotid artery atherosclerosis in type-2 diabetic and nondiabetic subjects with and without symptomatic coronary artery disease (The Insulin Resistance Atherosclerosis Study). AB - Type-2 diabetes mellitus is associated with a 2- to 4-fold increase in the risk of clinical coronary artery disease (CAD). It has been suggested that diabetic subjects without clinical CAD should be treated as aggressively for cardiovascular risk factors as subjects with CAD. This would be warranted if diabetic subjects without clinical CAD would have accelerated CAD similar to that of nondiabetic subjects with symptomatic CAD. To assess this suggestion, we compared the intima-media wall thickness in the common carotid artery (CCA) and internal carotid artery (ICA) in 43 diabetic subjects with clinical CAD, 446 diabetic subjects without clinical CAD, 47 nondiabetic subjects with clinical CAD, and 975 nondiabetic subjects without clinical CAD (all aged 40 to 70 years) in the Insulin Resistance Atherosclerosis Study. All data were adjusted for age, gender, ethnicity, and clinical results. Both diabetes and CAD were associated with increased atherosclerosis in the CCA. Likewise, diabetes was significantly associated with increased atherosclerosis in the ICA; however, CAD was not associated with ICA intima-media wall thickness. As expected, diabetic subjects with CAD had the greatest intima-media wall thickness, whereas nondiabetic subjects without CAD had the least atherosclerosis. Subjects with diabetes but without CAD had slightly greater intima-media wall thickness than nondiabetic subjects with CAD, although these differences were not statistically significant. Thus, diabetic subjects even without CAD had extensive atherosclerosis in the carotid artery. These results support the suggestion that diabetic subjects should be treated as aggressively for cardiovascular risk factor management as subjects with pre-existing CAD. PMID- 10856383 TI - Gender-related differences in thrombogenic factors predicting recurrent cardiac events in patients after acute myocardial infarction. The THROMBO Investigators. AB - Thrombosis contributes to recurrent coronary events in patients after acute myocardial infarction (AMI), but prognostic significance of thrombogenic factors by gender is unknown. This study aimed to determine gender-related differences in the prognostic significance of thrombogenic factors for predicting cardiac events (nonfatal reinfarction or cardiac death) in postinfarction patients. Blood levels of the following factors were measured 2 months after AMI in 791 men and 254 women: fibrinogen, von Willebrand factor, factor VII and VIIa, plasminogen activator inhibitor, D-dimer, cholesterol, apolipoprotein A-1, apolipoprotein B, lipoprotein(a), triglycerides, and high-density lipoprotein cholesterol. After adjustment for clinical covariates, levels of apolipoprotein A, high-density lipoprotein cholesterol, fibrinogen, and factor VIIa were significantly higher in postinfarction women than men. During a mean 26-month follow-up, there were 67 cardiac events (8.5%) in men and 14 (5.5%) in women (p = 0.11). In the multivariate Cox model, elevated levels of factor VIIa were a significant predictor of cardiac events in women (p = 0.022) but not in men (p = 0.80), with significant gender-related effect (hazard ratio 2.80 vs 0.92, respectively; p <0.05). D-dimer had prognostic value in men (p = 0. 006) but not in women (p = 0.36), although the difference between hazard ratios for men and women was not significant (2.35 vs 1.58, respectively; p = 0.49). In conclusion, elevated levels of factor VIIa are associated with an increased risk of recurrent cardiac events in postinfarction women, but not in men. D-dimer is more predictive for cardiac events in postinfarction men than women. These observations indicate possible gender-related differences in the pathophysiologic mechanisms of recurrent cardiac events. PMID- 10856384 TI - Degree of residual stenosis in the culprit coronary artery after thrombolytic administration (Thrombolysis In Myocardial Infarction [TIMI] trials). AB - This study was undertaken to characterize residual stenosis after thrombolytic administration and to evaluate clinical and angiographic features and early outcomes of patients with mild residual obstruction after thrombolytic administration. Patients who underwent angiography at 90 minutes after thrombolytic administration in the Thrombolysis In Myocardial Infarction 4, 10A, 10B, and 14 trials were divided into 3 groups according to the degree of residual stenosis measured by quantitative coronary angiography: patients with a patent culprit artery with <50% stenosis, patients with patent arteries and residual stenosis > or =50%, and patients with occluded arteries. Only 8.9% of the patients (188 of 2,119) had an infarct-related artery luminal diameter stenosis of <50% 90 minutes after thrombolysis. Compared with patients with patent arteries and > or =50% stenosis, patients with mild residual obstruction were younger (56.8 vs 58.6 years; p = 0.03), had fewer prior myocardial infarctions (6.9% vs 13.3%; p = 0.01), fewer eccentric (19.8% vs 42.1%; p <0.0001), ulcerated (7.5% vs 13.2%; p = 0.03), and collateralized (6.6% vs 13.2%, p = 0.01) lesions, but a greater thrombus burden (29.7% vs 18.3%, p = 0.0002). Among patients with patent arteries, a residual stenosis of <50% was associated with a significantly lower composite of in-hospital death, myocardial infarction, and congestive heart failure (2.8% vs 7.1%, p = 0.03). Thus, a minority of patients have a mild residual obstruction at 90 minutes after thrombolytic administration. These patients have less complex lesions with greater thrombus burdens and better clinical outcomes. PMID- 10856385 TI - Effects of a single intracoronary injection of basic fibroblast growth factor in stable angina pectoris. AB - We sought to evaluate safety, tolerability, pharmacokinetics, and pharmacodynamics of basic fibroblast growth factor (bFGF), administered as a single intracoronary injection, to subjects with stable angina pectoris secondary to coronary artery disease. bFGF, an angiogenic growth factor, has been shown to enhance collateral development in animal models of progressive coronary occlusion. To our knowledge, this study represents the initial introduction of parenteral bFGF into humans. This was a phase 1, randomized, dose-escalation trial of bFGF in 25 subjects with coronary artery disease and stable angina. Subjects were randomized 2:1 to a single dose of bFGF or placebo, injected into the left main coronary artery. bFGF doses ranged from 3 to 100 microg/kg, increasing in half-log increments. bFGF was generally well tolerated at doses of 3 to 30 microg/kg. Plasma clearance was 20 +/- 2 ml/kg/min, with an elimination half-life of 85 +/- 11 minutes. bFGF caused acute hypotension ( approximately 10%) that did not appear to be dose-related through the dose range studied. Of the 9 subjects who received 30 to 100 microg/kg bFGF, 2 had sustained hypotension, mild to moderate in severity, lasting 1 to 3 days, and 3 subjects developed bradycardia hours to days after bFGF administration. bFGF dilated epicardial coronary arteries (7.4 +/- 2.5% mean diameter increase, p <0.02). Transient mild thrombocytopenia and proteinuria were observed in some subjects in the 30-microg/kg cohort. No subject had signs suggesting systemic angiogenesis. Thus, intracoronary bFGF, at doses of 3 to 30 microg/kg, was generally well tolerated in subjects with stable angina. PMID- 10856386 TI - Late-onset heart failure as a mechanism for adverse long-term outcome in diabetic patients undergoing revascularization (a 13-year report from the Lady Davis Carmel Medical Center registry). AB - The adverse long-term prognosis following myocardial revascularization in diabetic patients has been ascribed to accelerated coronary disease, a higher incidence of late coronary restenosis after revascularization, and myocardial dysfunction. To examine the development of heart failure and its prognostic implications in diabetic patients, we analyzed the long-term (13-year) follow-up data of 363 patients-193 percutaneous transluminal coronary angioplasties and 170 coronary artery bypass operations-revascularized in a single cardiovascular center from 1984 to 1986. Baseline characteristics (age, previous infarction, baseline ventricular function) were similar in the 80 diabetic and 283 nondiabetic patients; multivessel disease and hypertension were marginally more common in diabetics (p = NS). Cumulative incidence of hospitalization for heart failure was high in the diabetic cohort (25% vs 11%, p = 0.001), with a rapidly increasing incidence after 5 years. Survival after first hospitalization for heart failure was markedly reduced in diabetics (11 of 20 [55%] vs 25 of 31 [81%] at 3 years; p = 0.04), as was survival free of further hospitalization for heart failure (5 of 20 [25%] vs 20 of 30 [63%]; p <0.005). Long-term 13-year survival (43% vs 78%, p <0.0001) and survival free of heart failure (33% vs 71%, p <0.0001) were decreased in diabetics, especially those with reduced ventricular function at baseline (17% vs 42%, p = 0.07). Multivariate analysis showed diabetes to be the strongest independent predictor of decreased survival (odds ratio 3. 6, 95% confidence interval 2.0 to 6.2; p <0.0001) and survival free of heart failure (odds ratio 4.0, 95% confidence interval 2.2 to 7. 1; p <0.0001) in patients undergoing revascularization. In summary, late-onset heart failure was frequent in diabetic patients after percutaneous transluminal coronary angioplasty or coronary artery bypass grafting, and once present heralded an unrelenting progressive downhill clinical course. PMID- 10856387 TI - Predictors of clinical outcome following percutaneous intervention for in-stent restenosis. AB - Percutaneous intervention for the first episode of in-stent restenosis was performed in 177 patients 5.4 +/- 0.3 months after native coronary stent implantation. Medical records were reviewed and patients contacted 13.3 +/- 1.2 months after in-stent intervention to ascertain the subsequent clinical course. The effects of demographic, procedural, and angiographic variables on clinical outcomes were determined. At 2 years, Kaplan-Meier estimated survival was 93 +/- 3% and freedom from death, myocardial infarction, and a third target artery revascularization (TAR) was 67 +/- 4%. The actuarial frequency of a third TAR was 26 +/- 4% at 1 year. Stratification of outcomes according to timing of in-stent intervention revealed an approximate twofold higher frequency of adverse events among patients with early (5%. Improvement in EF during dobutamine infusion predicted RNV-LVEF recovery after CABG significantly better than segmental wall motion changes (72% vs 53%, p = 0.03). A biphasic EF response (i.e., improvement in > or =10% at low dose and subsequent worsening at peak stress) had the highest predictive value (80%) for late functional recovery. In conclusion, EF response to dobutamine infusion was superior to segmental wall motion changes in predicting RNV-LVEF recovery after CABG. PMID- 10856390 TI - Temporal patterns of paroxysmal atrial fibrillation following DDDR pacemaker implantation. AB - Paroxysmal atrial fibrillation (AF) episodes have been reported to be randomly distributed. However, because patients are not always symptomatic, it has been difficult to study temporal patterns of AF. Newer implantable pulse generators have data-logging capabilities that permit the detection and analysis of temporal patterns of AF. This study tested the hypothesis that AF episodes occur in clusters over time and that these episodes are not randomly distributed in individual patients. The date and time of 582 episodes of AF were recorded from the data logs of 16 patients with a Medtronic Thera DR followed 6 weeks and 6 and 12 months after pulse generator implant. The probability of AF recurrence and the interevent intervals between successive episodes of AF were fitted to monoexponential and Weibull distributions. A Weibull distribution best described the nonrandom distribution of AF for 67% of follow-up visits. Temporal clustering of AF (interevent intervals <24 hours) declined during follow-up (95 +/- 10%, 90 +/- 11%, and 74 +/- 28% at the 6-week and 6- and 12-month visits, respectively; p <0.05). The average duration of an episode of AF tended to increase over time (0.31 hour, 95% confidence intervals [CI] 0.17 to 0.58 hours; 0.36 hours, 95% CI 0. 17 to 0.78 hours; 0.65 hours, 95% CI 0.29 to 1.45 hours [p = 0.07] at the 6 week and 6- and 12-month visits, respectively). Paroxysmal AF recurrence is nonrandomly distributed over the long term in many patients. The temporal patterns of AF change over time after pacemaker implantation. This has implications for the selection of study end points in AF clinical trials. PMID- 10856391 TI - Time to recover from atrial hormonal, mechanical, and electrical dysfunction after successful electrical cardioversion of persistent atrial fibrillation. AB - Although transient atrial dysfunction has been reported after electrical cardioversion of atrial fibrillation (AF), the difference in the time to recover from the atrial hormonal, mechanical, and electrical dysfunction has not been described. Thus, we evaluated the time course of recovery from atrial hormonal, mechanical, and electrical dysfunction after cardioversion in patients with nonvalvular AF. We attempted electrical cardioversion in 87 consecutive patients with nonvalvular AF that had persisted for > or =6 months, and in 24 patients (28%) with maintained sinus rhythm for > or =6 months. To evaluate atrial hormonal, mechanical, and electrical dysfunction in these 24 patients, we measured plasma concentration of atrial natriuretic peptide, the atrial peak velocity in transmitral flow, and the ratio of peak systolic-to-diastolic pulmonary venous flow (S/D ratio) using echocardiography, and the duration and the root mean voltage for the terminal 20 ms (LP20) of the filtered P wave using P-wave signal-averaged electrocardiography. Atrial natriuretic peptide rapidly returned to baseline within 1 day after cardioversion, and maintained these levels for 6 months. Atrial peak velocity in transmitral flow and S/D ratio were significantly increased at 2 weeks, and continued to increase until 1 month, and then reached a plateau. The duration and LP20 began to recover only 6 months after cardioversion. One to 3 years after conversion, the duration and LP20 had nearly reached a plateau, but the latter value remained below normal. In patients with nonvalvular AF of prolonged duration, recovery from atrial electrical dysfunction after sinus conversion took much longer than that from either atrial hormonal or mechanical dysfunction. PMID- 10856392 TI - Impact of blunted pulmonary venous flow on the outcome of patients with left ventricular systolic dysfunction secondary to either ischemic or idiopathic dilated cardiomyopathy. AB - The intravenous administration of echo contrast agents enhances the Doppler signal and makes the study of pulmonary venous flow (PVF) easily achievable by transthoracic echocardiography. The aim of this study was to evaluate whether PVF patterns play a role in predicting the outcome of patients with left ventricular (LV) systolic dysfunction. Thus, 115 patients (79 men, mean age 69 years) with LV dysfunction (ejection fraction [EF] <45%) due to either ischemic or idiopathic dilated cardiomyopathy were studied and followed-up for 1 year. A quantitative interrogation of all components of PVF was feasible in 69% of patients at standard transthoracic examination; after contrast enhancement, anterograde and retrograde flow velocities were measurable in 100% and 92% of patients, respectively. A blunted PVF (defined by a systolic-to-diastolic peak velocity ratio <1) was identified in 48 patients (42%), who had a worse clinical status, a lower LVEF, and a more severe pulmonary hypertension. Thirty-six patients had cardiac events at follow-up: sudden death in 4, progressive heart failure in 12, and hospitalization for worsening heart failure in 20 patients. Multivariate Cox proportional-hazards analysis revealed that advanced New York Heart Association class, male gender, and older age were independent predictors of mortality. However, blunted PVF, reduced LVEF, older age, and increased heart rate in descending order of power were independent predictors of heart failure hospitalizations and deaths from end-stage heart failure. In conclusion, the assessments of PVF may effectively contribute to the characterization of patients with LV dysfunction and to the prediction of their outcome. PMID- 10856393 TI - Furosemide withdrawal in elderly heart failure patients with preserved left ventricular systolic function. AB - To explore the possibilities of furosemide withdrawal in elderly heart failure (HF) patients with intact left ventricular (LV) systolic function and assess its effects on functional status and orthostatic blood pressure homeostasis, we performed a placebo-controlled pilot trial of furosemide withdrawal with 3 months of follow-up in 32 HF patients (aged 75.1 +/- 0.7 years [mean +/- SEM]) with a LV ejection fraction of 60 +/- 2% and without overt congestion. Investigations included repeated clinical assessment, spirometry, standardized 6-minute walking test, and chest x-rays. Measurements of blood pressure response on active standing and Doppler echocardiography were performed before and 3 months after furosemide withdrawal. Recurrent congestive HF occurred in 2 of 21 patients (10%) who discontinued furosemide use, and in 1 of 11 patients (9%) who continued furosemide (p = NS). Three patients restarted furosemide for ankle edema and 1 for blood pressure levels >180/100 mm Hg. After 3 months, there were no differences regarding HF symptom scores, blood pressure, heart rate, spirometric results, 6-minute walking distance, or quality of life scores between patients who discontinued use and patients who continued the therapy. In patients successfully withdrawn, Doppler E/A ratio increased from 0.68 +/- 0.05 to 0.79 +/ 0.06 after withdrawal (p <0.01), and maximum blood pressure decrease on active standing changed from -8 +/- 5 mm Hg to +5 +/- 3 mm Hg systolic (p <0.05). Thus, in this pilot investigation of furosemide withdrawal in elderly HF patients without overt congestion and with a normal LV systolic function, withdrawal was successful in almost all patients and was associated with improvement of LV diastolic filling and blood pressure homeostasis on active standing. PMID- 10856394 TI - Treatment of heart failure with celiprolol, a cardioselective beta blocker with beta-2 agonist vasodilatory properties. The CELICARD Group. AB - Treatment with beta blockers results in improvement in functional status, and reduces mortality in patients with heart failure. A number of differences in the results noted could be due to additional properties of the specific beta blockers studied: absence of cardioselectivity, and existence of a vasodilator effect and of an associated antioxidant effect. We studied the effects of celiprolol, a cardioselective beta blocker with a stimulant effect on beta2 receptors. One hundred thirty-two patients presenting with chronic heart failure of various etiologies, with an ejection fraction of <40% and New York Heart Association cardiac functional status grades II and III were included in a randomized, double blind, placebo-controlled study. The maximum dose of celiprolol (100 mg) was attained after 1 month. The study lasted 1 year. The primary evaluation criterion was functional class as evaluated using the Goldman questionnaire. There was no difference in efficacy between the 2 treatment groups in terms of functional class (p = 0.56). With regard to the secondary evaluation criteria, an improvement in DiBianco functional score was seen with celiprolol (p = 0.03), as well as a significant reduction in heart rate (p = 0.01). Ejection fraction increased in both groups (p = 0.15). There was no difference regarding improvement in left ventricular volume as determined at echocardiography or in exercise capacity. The safety profile of celiprolol was excellent. There was no difference in terms of cardiovascular mortality (2 receiving celiprolol vs 4 placebo), onset of arrhythmias (2 receiving celiprolol vs 3 placebo), worsening of heart failure (26 receiving celiprolol vs 23 placebo), or noncardiovascular adverse events (9 receiving celiprolol vs 14 placebo). The absence of a significant efficacy of celiprolol, a beta blocker with vasodilator properties, but exerting stimulation of beta2 receptors, suggests an unfavorable role of this latter property in heart failure. However, the safety profile of celiprolol was excellent. This beta blocker may consequently be used for its other indications, hypertension and angina, in patients presenting with altered cardiac function. PMID- 10856395 TI - Impact of anatomic closure on somatic growth among small, asymptomatic children with secundum atrial septal defect. AB - We retrospectively identified 92 children aged dipyridamole>/=hexobendine inhibited [3H]adenosine incorporation at low micromolar concentrations. The nucleosides inosine and uridine were weak inhibitors of this system. The adenosine receptor ligands N(6)-phenylisopropyladenosine (PIA) and 2 chloroadenosine inhibited the uptake only at micromolar concentrations. Neither 5'-(N-ethylcarboxamido)-adenosine (NECA) nor theophylline inhibited adenosine uptake by more than 60% but the mitochodrial benzodiazepine receptor ligands 4' chloro-diazepam (Ro 5-4864) and 1-(2-chlorophenyl)-N-methyl-N-(1-methyl-propyl) isoquinoline carboxamide (PK 11195) were able to inhibit it. The lack of inhibition by the blockers of the mitochondrial adenine-nucleotide carrier, atractyloside and alpha, beta-methylene-ATP, indicates that [3H]adenosine uptake occurs via a transporter other than this carrier. All these results support the existence of an equilibrative adenosine transport system, which might mediate the passage of adenosine formed in the mitochondria to the cytoplasm. PMID- 10856446 TI - Effects of buspirone on brain indoleamines and catecholamines in wild-type mice and Lurcher mutants. AB - The effects of a chronic serotoninergic stimulation on brain monoamine levels and metabolism were studied in wild-type (+/+) mice and Lurcher (Lc/+) mutants. Endogenous serotonin, dopamine, noradrenaline and some of their major metabolites were measured in the frontal cortex, neostriatum, thalamus, brainstem, cerebellum and spinal cord. In +/+ mice, buspirone (1 mg/kg; i.p.) treatment during 40 days increased indoleamines, albeit with moderate changes in the ratios between tissue serotonin metabolites and endogenous serotonin, augmented noradrenaline contents in the spinal cord, and caused elevations of dopamine metabolites in most regions. In Lc/+ mutants, the effects of buspirone were attenuated, but higher L tryptophan and indoleamine levels, suggest a storage of serotonin in a non releasable compartment. In the hypoplastic Lc/+ cerebellum, indoleamine content was accrued, but with a decreased [serotonin metabolites]/[serotonin] ratio, indicating that the reorganized nerve terminals in Lc/+ mutants although they can synthesize and accumulate serotonin, may not utilize it efficiently in synaptic transmission. PMID- 10856447 TI - A microdialysis study on the mechanism of action of gabapentin. AB - To gain insight into the mechanism of action of the anti-epileptic, gabapentin, the effects of gabapentin on the in vivo extracellular gamma-aminobutyric acid (GABA) levels in the rat substantia nigra reticulata were studied using microdialysis. In order to investigate possible interference with different GABA ergic compartments in the substantia nigra reticulata, we studied the effects of gabapentin under basal, K(+)-, nipecotic acid- and glutamate-stimulated conditions. Intraperitoneally (i.p.) administered gabapentin, at a dose of 100 mg/kg, did not significantly affect extracellular GABA levels under any condition. Thus, our data do not support the involvement of nigral GABA release in the mechanism of action of the anti-epileptic gabapentin. PMID- 10856448 TI - 5-HT(2C) receptor antagonists enhance the behavioural response to dopamine D(1) receptor agonists in the 6-hydroxydopamine-lesioned rat. AB - Non-dopaminergic therapies are of potential interest in the treatment of Parkinson's disease given the complications associated with current dopamine replacement therapies. In this study we demonstrate that SB 206553 (5-methyl-1-(3 pyridylcarbamoyl)-1,2,3, 5-tetrahydropyrrol[2,3-f]indole) (20 mg/kg) enhanced the actions of the dopamine D(1) receptor agonist, SKF 82958 ((+)-6-chloro-7, 8 dihydroxy-3-allyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide) (1 mg/kg), in eliciting locomotion in the 6-hydroxydopamine-lesioned rat model of Parkinson's disease. This action was only seen following prior priming with L DOPA (L-3, 4-dihydroxyphenylalanine). SB 206553 had no effect on rotational behaviour when given alone. 5-HT(2C) receptor antagonists may have potential as a means of reducing reliance on dopamine replacement in the treatment of Parkinson's disease. PMID- 10856449 TI - Huperzine A improves cognitive deficits caused by chronic cerebral hypoperfusion in rats. AB - The effects of (-)-huperzine A, a promising therapeutic agent for Alzheimer's disease, on learning behavior and on alterations of the cholinergic system, the oxygen free radicals and energy metabolites induced by permanent bilateral ligation of the common carotid arteries were investigated in rats. Daily oral administration of huperzine A produced a significant improvement of the deficit in the learning of the water maze task, beginning 28 days after ischemia, correlating to about 33-40% inhibition of acetylcholinesterase activity in cortex and hippocampus. Huperzine A significantly restored the decrease in choline acetyltransferase activity in hippocampus and significantly reduced the increases in superoxide dismutase, lipid peroxide, lactate and glucose to their normal levels. The present findings demonstrate that the improvement by huperzine A of the cognitive dysfunction in the late phase in chronically hypoperfused rats is due to its effects, not only on the cholinergic system, but also on the oxygen free radical system and energy metabolism. Our results strongly suggest that huperzine A has therapeutic potential for the treatment of dementia caused by cholinergic dysfunction and/or decrease of cerebral blood flow. PMID- 10856450 TI - The in vivo pharmacological profile of eletriptan (UK-116,044): a potent and novel 5-HT(1B/1D) receptor agonist. AB - The anti-migraine drug, eletriptan [(R)-3-(1-methyl-2-pyrrolidinylmethyl)-5-[2 (phenylsulphonyl )ethyl]-1 H-indole; UK-116,044], is a novel 5-HT(1B/1D) receptor agonist. In this paper, the regional vasoconstrictor profile of eletriptan, in comparison with sumatriptan, was examined in the anaesthetised dog. The inhibitory actions of eletriptan on neurogenic inflammation in rat dura mater were also assessed. In the anaesthetised dog, eletriptan (1-1000 microg kg(-1) i.v.) produced a dose-dependent reduction of carotid arterial blood flow with a similar potency and maximum effect to sumatriptan (ED(50) values: eletriptan and sumatriptan, 12 and 9 microg kg(-1), i.v., respectively). However, eletriptan exhibited a significantly lower potency than sumatriptan in reducing coronary artery diameter (ED(50) values: 63 and 19 microg kg(-1), i.v., respectively, P<0.05). In the femoral circulation, sumatriptan caused a significant reduction in arterial blood flow (ED(50) 35 microg kg(-1) i.v.) whereas eletriptan (1-1000 microg kg(-1) i.v.) had no significant effect upon femoral arterial blood flow when compared to vehicle-treated animals. In rats, eletriptan (30-300 microg kg( 1) i.v.) administered prior to electrical stimulation of the trigeminal ganglion produced a dose-related and complete inhibition of plasma protein extravasation in the dura mater (mean extravasation ratio: control 1.9; eletriptan 1.0, minimum effective dose 100 microg kg(-1), P<0.05). The potency and maximum effect of eletriptan was identical to that of sumatriptan in this model. When administered during a period of continual stimulation of the trigeminal nerve, eletriptan (100 microg kg(-1) i.v.) produced a complete inhibition of plasma protein extravasation. The ability to reduce canine carotid arterial blood flow and inhibit neurogenic inflammation in rat dura mater suggests that vascular and neurogenic mechanisms may contribute to eletriptan's clinical efficacy in migraine patients. In addition, eletriptan exhibits some selectivity for reducing carotid arterial blood flow when compared with femoral arterial blood flow and coronary artery diameter, in the anaesthetised dog. PMID- 10856451 TI - Protective effect of diltiazem against ischemia-induced decreases in regional myocardial flow in rat heart. AB - Diltiazem has cardioprotective properties following myocardial ischemic injury. However, there are controversial results regarding the beneficial effects of diltiazem on regional myocardial flow after ischemia. Therefore, we investigated the effect of diltiazem on changes in regional myocardial flow due to ischemia for different periods. Non-radioactive colored microspheres were used for this measurement in isolated rat heart. After 20 or 40 min of global ischemia and 40 min of reperfusion, regional myocardial flow was decreased, especially in the endocardial layer. The endocardial/epicardial ratio was also decreased. The decreases in endocardial flow and the endocardial/epicardial ratio were more remarkable after 40 min of ischemia than after 20 min of ischemia. Diltiazem (10( 6) M), which was administered 15 min before ischemia, prevented only the decrease in endocardial flow and endocardial/epicardial ratio after 20 min of ischemia, whereas it did not prevent that after 40 min of ischemia. Nifedipine (2x10(-6) M) did not exert a cardioprotective effect. These findings suggested that the effect of ischemia is marked in the endocardium and, also, that the protective effect of diltiazem is seen only during a decrease in endocardial flow following short-term and reversible ischemia. PMID- 10856452 TI - Effects of vitamin E and sodium selenate on neurogenic and endothelial relaxation of corpus cavernosum in the diabetic mouse. AB - We studied the effect of vitamin E and sodium selenate treatment on the neurogenic and endothelium-dependent relaxation of isolated corpus cavernosum obtained from streptozotocin-induced diabetic mice. Relaxant responses of corpus cavernosum precontracted by phenylephrine to electrical field stimulation and to acetylcholine were significantly decreased in diabetic mice. There was no significant difference between diabetic and non-diabetic groups for the relaxant response of corpus cavernosum to sodium nitroprusside and papaverine. Treatment with sodium selenate, but not vitamin E, partially prevented the impairment of the neurogenic relaxation, whereas both had a significant, partial restorative action on endothelial dysfunction in corpus cavernosum obtained from diabetic groups. Neither agent exhibited a significant action on the relaxant responses of corpus cavernosum obtained from non-diabetic mice. A decrease in the sensitivity of the neurogenic impairment to antioxidant action may develop more rapidly than that of endothelial dysfunction in streptozotocin-induced diabetic mice. PMID- 10856453 TI - Enalapril and valsartan improve cyclosporine A-induced vascular dysfunction in spontaneously hypertensive rats. AB - Cyclosporine A causes hypertension and nephrotoxicity in spontaneously hypertensive rats (SHR). In the present study, arterial function was investigated using in vitro vascular preparations after long-term treatment with cyclosporine A. SHR received cyclosporine A (5 mg kg(-1) day(-1) s.c.) and high-Na(+) diet for 6 weeks during the developmental phase of hypertension. Part of the rats were treated concomitantly either with the angiotensin converting enzyme inhibitor enalapril (30 mg kg(-1) day(-1) p.o.) or with an angiotensin AT(1) receptor antagonist valsartan (3 or 30 mg kg(-1) day(-1) p.o.). In renal arteries, contractile responses to noradrenaline and angiotensin II, as well as relaxation responses to acetylcholine (endothelium-dependent) and to sodium nitroprusside (endothelium-independent), were severely impaired by cyclosporine A-treatment. There was also a trend for the dysfunction of the mesenteric arteries, but the impairment did not reach statistical difference. Enalapril and valsartan improved the impaired renal arterial functions. Cyclosporine A-induced hypertension and nephrotoxicity seem to be associated with renal arterial dysfunction in SHR on high-Na(+) diet. Antagonism of the renin-angiotensin system protects from vascular toxicity of cyclosporine A. PMID- 10856454 TI - Antiarrhythmic and cardiohemodynamic effects of a novel Ca(2+) channel blocker, AH-1058, assessed in canine arrhythmia models. AB - The antiarrhythmic profile and cardiohemodynamic effect of a novel Ca(2+) channel blocker, 4-(5H-Dibenzo[a, d]cyclohepten-5-ylidene)-1-[(E)-3-(3-methoxy-2 nitro)phenyl-2-p ropeny l]piperidine hydrochloride (AH-1058), were analyzed using the epinephrine-, digitalis- and two-stage coronary ligation-induced canine ventricular arrhythmia models. Intravenous administration of AH-1058 (100 microg/kg) effectively suppressed each of the ventricular arrhythmias accompanied by weak hypotensive effects. The results contrast well with those of a typical Ca(2+) channel blocker, verapamil, which suppresses only the epinephrine-induced ventricular arrhythmia with severe hypotension. These results indicate that AH 1058 may possess a more selective inhibitory action on Ca(2+) channels in the heart than on those in the vessels. Furthermore, the antiarrhythmic actions of AH 1058 were slower in onset and longer-lasting, than those in our previous studies using other antiarrhythmic drugs, including Na(+) and Ca(2+) channel blockers. The antiarrhythmic effects of AH-1058 did not correlate with its plasma concentrations when administered either intravenously or orally. These results suggest that AH-1058 can become a long-acting Ca(2+) channel blocker with unique antiarrhythmic properties, and that AH-1058 may be used in certain pathological processes, for which selective inhibition of the cardiac Ca(2+) channels is essential. PMID- 10856455 TI - Protein kinase cdelta and alpha are involved in the development of vasospasm after subarachnoid hemorrhage. AB - We have previously shown the enhanced activity of protein kinase C in the membrane fraction of the canine vasospastic artery after subarachnoid hemorrhage, which increased with progression of angiographic vasospasm. This study examined identification of protein kinase C isoforms in the canine basilar artery, and the changes in expression and/or translocation of each isoform during the development of vasospasm. Vasospasm was produced by using the "two-hemorrhage" canine model in the basilar artery, and angiographic progression of vasospasm was assessed consecutively. Two isoforms, protein kinase Calpha and delta were identified in basilar arteries by Western blotting. Densitometric analysis showed that the expression of protein kinase Cdelta in the membrane fraction was significantly increased in the earlier stage, and protein kinase Calpha was increased later as vasospasm progressed. These results indicate that protein kinase Cdelta and alpha isoforms may play a significant role in the development and maintenance of vasospasm. PMID- 10856456 TI - Acute dependence on, but not tolerance to, heroin and morphine as measured by respiratory effects in rhesus monkeys. AB - Acute dependence on and tolerance to heroin and morphine were assessed in rhesus monkeys using measures of respiration. Respiratory frequency (f) and minute volume (V(e)) were measured in monkeys breathing air or 5% CO(2) in air using a pressure-displacement plethysmograph. Cumulative doses of naltrexone (0.0001-1.0 mg/kg, i.m) did not alter these parameters in untreated monkeys. Twenty-four hours after a cumulative dose of heroin (1 mg/kg, i.m.), naltrexone produced an increase in both f and V(e) when monkeys were breathing air or 5% CO(2). Following 1 to 3 days of treatment with heroin (0.5 mg/kg/day, i.m.) or morphine (16 mg/kg/day, i.m.), naltrexone produced an increase in f and V(e) that was related to the dose of naltrexone and the number of days of agonist administration. Two days following termination of heroin administration, naltrexone-induced respiratory stimulation declined and had disappeared completely by the fifth day. In tolerance studies, heroin (0.032-0.5 mg/kg, i.m.) and morphine (1-16 mg/kg, i. m.) were injected cumulatively each day for three consecutive days. These drugs suppressed f and V(e) to nearly the same extent on day 3 as they had on day 1 of administration. These results suggest that dependence to morphine and heroin can be measured under conditions of acute 1 to 3 day administration conditions in primates using f and V(e) as reliable and quantitative indicators of opioid withdrawal. Under these conditions, tolerance does not occur. PMID- 10856457 TI - Hoe 140 and pseudo-irreversible antagonism in the rat vas deferens in vitro. AB - The effects of bradykinin and the bradykinin B(2) receptor antagonists D-Arg [Hyp(3),Thi(5,8),D-Phe(7)]-bradykinin (NPC 349) and D-Arg-[Hyp(3),Thi(5),D Tic(7),Oic(8)]-bradykinin (Hoe 140) were examined in the electrically-stimulated rat vas deferens. Cumulative additions of bradykinin (1-3000 nM) produced two distinct responses: an enhancement in the magnitude of the basal electrically induced twitch response (neurogenic response) and an increase in the baseline tension (musculotropic response). NPC 349 (10-100 microM) produced concentration dependent surmountable rightward shifts of both the bradykinin neurogenic and musculotropic response curves. In contrast, while Hoe 140 (10-100 nM) caused an apparently surmountable antagonism of the bradykinin neurogenic response, it caused an apparent insurmountable antagonism of the bradykinin musculotropic response. Interestingly, co-incubation of Hoe 140 (30 nM) with NPC 349 (30 and 100 microM) resulted in a concentration-related upwards displacement of the Hoe 140-suppressed bradykinin musculotropic response curve. Thus, Hoe 140 can be described as a pseudo-irreversible antagonist against the bradykinin musculotropic response. No time-dependent changes were observed in the maximum bradykinin musculotropic response attainable when NPC 349 (100 microM) additions were made for the final 2 or 18 min of the Hoe 140 incubation (20 min). These findings indicate that slow reversibility of Hoe 140 from the bradykinin B(2) receptor is unlikely to be the mechanism responsible for the pseudo-irreversible antagonism of the bradykinin-induced musculotropic response. Instead, we propose an alternative explanation involving a third, unstable and inactive form of the bradykinin B(2) receptor. PMID- 10856458 TI - Early changes in lipid peroxidation and antioxidative defense in diabetic rat retina: effect of DL-alpha-lipoic acid. AB - This study was designed to (1) evaluate retinal lipid peroxidation in early diabetes by the method specific for free malondialdehyde and 4-hydroxyalkenals, (2) identify impaired antioxidative defense mechanisms and (3) assess if enhanced retinal oxidative stress in diabetes is prevented by the potent antioxidant, DL alpha-lipoic acid. The groups included control and streptozotocin-diabetic rats treated with or without DL-alpha-lipoic acid (100 mg kg(-1) day(-1), i.p., for 6 weeks). All parameters were measured in individual retinae. 4-Hydroxyalkenal concentration was increased in diabetic rats (2.63+/-0.60 vs. 1.44+/-0.30 nmol/mg soluble protein in controls, P<0.01), and this increase was prevented by DL-alpha lipoic acid (1.20+/-0.88, P<0.01 vs. untreated diabetic group). Malondialdehyde, reduced glutathione (GSH) and oxidized glutathione (GSSG) concentrations were similar among the groups. Superoxide dismutase, glutathione peroxidase (GSHPx), glutathione reductase (GSSGRed) and glutathione transferase (GSHTrans) activities were decreased in diabetic rats vs. controls. Quinone reductase was upregulated in diabetic rats, whereas catalase and cytoplasmic NADH oxidase activities were unchanged. DL-alpha-Lipoic acid prevented changes in superoxide dismutase and quinone reductase activities induced by diabetes without affecting the enzymes of glutathione metabolism. In conclusion, accumulation of 4-hydroxyalkenals is an early marker of oxidative stress in the diabetic retina. Increased lipid peroxidation occurs in the absence of GSH depletion, and is prevented by DL-alpha lipoic acid. PMID- 10856459 TI - Role of gastric acid secretion in progression of acute gastric erosions induced by ischemia-reperfusion into gastric ulcers. AB - Ischemia followed by reperfusion is known to produce gastric lesions due to oxidative stress, but the role of gastric H(+) secretion in the formation of this mucosal injury remains unknown. We studied alterations in gastric acid secretion and gastric histamine content, as well as the expression of histidine decarboxylase and interleukin-1beta during the mucosal recovery from ischemia reperfusion erosions. Gastric secretion was studied in rats (series A) with gastric fistula before, during and after the ischemia induced by clamping of celiac artery for 0.5 h followed by reperfusion in animals pretreated with vehicle (saline), omeprazole, a proton pump inhibitor, or ranitidine, a histamine (H(2)) receptor antagonist. In series B, the animals were submitted to 0.5 h of ischemia followed by 1 h of reperfusion and then anesthetized at 0, 3, 12 and 24 h or 3, 5, 10 or 15 days after the end of ischemia-reperfusion to determine gastric blood flow by H(2)-gas clearance technique, area of gastric lesions, plasma gastrin and interleukin-1beta levels, histamine content by radioimmunoassay (RIA) and expression of histidine-decarboxylase and interleukin 1beta mRNA by reverse transcription polymerase chain reaction. Clamping of celiac artery caused cessation of gastric blood flow and almost complete suppression of basal gastric acid secretion (series A) that returned gradually to the control value at day 3 after ischemia-reperfusion, accompanied by the rise in plasma gastrin levels, pronounced expression of histidine-decarboxylase mRNA and increased mucosal histamine content. Ischemia, followed by 1 h of reperfusion, produced gastric erosions (series B) that reached maximum at 12 h, but then declined at 24 h. These erosions progressed at day 3 into deeper ulcers whose area declined progressively within the next 5-15 days. The gastric blood ceased to flow (series B) during 30 min of clamping and was reduced throughout the period of healing of acute erosions, being accompanied by a gradual rise in mucosal interleukin-1beta mRNA content and in plasma interleukin-1beta levels. Treatment with omeprazole or ranitidine, which completely suppressed gastric acid secretion and significantly raised plasma gastrin level, greatly reduced the formation of erosive lesions preventing the progression of these lesions to chronic gastric ulcers, and this was accompanied by the rise in gastric blood flow and plasma gastrin levels and the significant attenuation of plasma interleukin-1beta levels. The ranitidine and omeprazole-induced suppression of ischemia-reperfusion erosions were abolished by the instillation of exogenous 0.2 N HCl into the stomach of these rats. The histidine-decarboxylase was faintly expressed in the intact gastric mucosa, but strongly upregulated during mucosal recovery from the damage induced by ischemia-reperfusion. We conclude that following ischemia-reperfusion: (1) gastric acid secretion, gastric microcirculation and histamine production markedly decline, while interleukin 1beta release significantly increases, probably playing an important role in the progression of acute lesions into chronic gastric ulcerations; (2) the suppression of gastric acid secretion by omeprazole and ranitidine, that induces hypergastrinemia, prevents the progression of gastric erosions into ulcers; and (3) the addition of exogenous acid restores the progression of the acute lesions into gastric ulcers, indicating that gastric acid plays a key role in ulcerogenesis induced by ischemia-reperfusion. PMID- 10856460 TI - Epidermal growth factor accelerates pancreatic recovery after caerulein-induced pancreatitis. AB - We examined the influence of endogenous and exogenous epidermal growth factor (EGF) on pancreatic repair after acute pancreatitis. Caerulein-induced pancreatitis was evoked in rats with intact or removed salivary glands and EGF (10 microg/kg) was administered starting 24 h after cessation of caerulein infusion. The dose of EGF 10 microg/kg was chosen because it was the most effective in preliminary experiments when 1, 10 or 50 microg/kg of EGF was used. Caerulein administration caused acute edematous pancreatitis with biochemical and histological manifestation of pancreatic damage, followed by spontaneous regeneration. The effect of salivectomy on the course of acute pancreatitis was slight, resulting in additional reduction in pancreatic blood flow, DNA synthesis and in an increase in plasma interleukin 1beta level. Treatment with EGF accelerated the healing of pancreatic damage, causing an increase in pancreatic blood flow and DNA synthesis. EGF caused faster normalization of plasma amylase and lipase activity and plasma interleukin 1beta concentration, as well as, this peptide accelerated the restoration of pancreatic amylase activity. On histological examination, EGF caused reduction of pancreatic damage and acceleration of tissue repair. We conclude that EGF reduces the severity of pancreatic damage evoked by caerulein-induced pancreatitis-related pancreatic damage and accelerates tissue repair. The beneficial effects of EGF appear to depend, at least in part, on the improvement of pancreatic blood flow, as well as on an increase of pancreatic cell growth and limitation of the activation cytokine release. PMID- 10856462 TI - Medical management of endometriosis and infertility. AB - OBJECTIVE: To review the literature on the use of medical management of endometriosis and infertility. DESIGN: Literature review. RESULT(S): Endometriosis is a common finding in women with infertility, but the mechanism by which it renders a woman infertile remains unclear. Despite many years of controversy and debate, there remains a strong bias against medical treatment for endometriosis-associated infertility. A review of the current literature suggests that medical management of endometriosis may be effective in selected patients and in certain settings, including patients undergoing IVF. CONCLUSION(S): A closer look at the question of medical management of endometriosis reveals that much remains to be learned before a final decision can be made about the use of medical therapies, such as GnRH agonists, for endometriosis and associated infertility. PMID- 10856461 TI - Effects of low-dose VOSO(4) on age-related changes in glucose homeostasis in rats. AB - The effects of low doses of vanadyl sulfate (0.2 mg/ml in the drinking water) on the age-related impairment of glucose homeostasis in Sprague-Dawley rats were investigated. VOSO(4) administration was initiated in 5-month-old animals and lasted 3 months. Thus, in 8-month-old rats, we investigated glucose metabolism in vivo and insulin secretory function in vitro. Results showed that VOSO(4) allowed the disposal of an oral glucose load at lower insulin levels than in age-matched controls. No significant changes were found in muscle glucose transporter (GLUT 4) levels or in glycogen content upon VOSO(4) treatment. Islets isolated from VOSO(4)-treated rats released less insulin than control islets, but showed a better preserved sensitivity to secretagogues, in terms of incremental release over basal release, secretory efficiency, and maintenance of the priming effect of glucose. In conclusion, chronic low-dose VOSO(4) treatment facilitates insulin action by a mechanism independent of muscle GLUT-4 levels and helps preserve the appropriate sensitivity of beta cells to stimuli, thereby preventing age dependent functional alterations. PMID- 10856463 TI - Insulin sensitizers and polycystic ovary syndrome: can a diabetes medication treat infertility? PMID- 10856464 TI - A comparison of the morphology of testicular, epididymal, and ejaculated sperm from fertile men and men with obstructive azoospermia. AB - OBJECTIVE: To assess the morphology of testicular, epididymal, and ejaculated sperm. DESIGN: Morphology of the three types of sperm was assessed by using Tygerberg strict criteria. SETTING: The Regional Fertility Center, Royal Maternity Hospital, Belfast, Northern Ireland, United Kingdom. PATIENT(S): Thirty two men with obstructive azoospermia and 10 fertile men. INTERVENTION(S): Trucut needle testicular biopsy and percutaneous epididymal sperm aspiration under local anesthetic. MAIN OUTCOME MEASURE(S): Percentages of normal sperm and sperm with head, midpiece, and tail defects for testicular, epididymal, and ejaculated sperm. Testicular sperm morphology in men with obstructive azoospermia was compared with that of fertile men. RESULT(S): The percentage of normal testicular sperm (4.3%) differed significantly from the percentages of normal epididymal (10. 8%) and ejaculated sperm (9.6%). Testicular sperm morphology in men with obstructive azoospermia did not differ from that in fertile men. CONCLUSION(S): Tygerberg strict criteria are not suitable for the assessment of testicular sperm morphology. PMID- 10856465 TI - Expression pattern of germ cell-specific genes in the testis of patients with nonobstructive azoospermia: usefulness as a molecular marker to predict the presence of testicular sperm. AB - OBJECTIVE: To analyze the expression pattern of testis-specific genes of patients with various spermatogenic defects and their usefulness as a molecular marker to predict the presence of testicular spermatozoa in patients with nonobstructive azoospermia undergoing IVF. DESIGN: Prospective, controlled study. SETTING: Hospital-based infertility research laboratory. PATIENT(S): Fifty-eight men with azoospermia or severe oligozoospermia. INTERVENTION(S): Testicular biopsy was done in the patients with obstructive or nonobstructive azoospermia, including Sertoli cell-only syndrome, maturation arrest, severe hypospermatogenesis, and normal spermatogenesis. MAIN OUTCOME MEASURE(S): Reverse transcriptase polymerase chain reaction was performed using 1 microgram of total RNA extracted from testicular tissues. Three pairs of primers were used for amplification of male germ cell-specific genes (DAZ, transcribed in male germ cells; PGK2, in late spermatocytes and spermatids; protamine-2, in spermatids) as molecular markers. Testicular sperm was obtained by multiple testicular sperm extraction. RESULT(S): The DAZ, PGK2, and protamine-2 genes were expressed in 38, 30, and 21 of the 43 patients with nonobstructive azoospermia, respectively. Testicular spermatozoa were successfully extracted in 4 of 43 patients with nonobstructive azoospermia with the use of multiple testicular sperm extraction. Detection of protamine-2 transcripts predicted the presence or absence of spermatozoa in the testicular tissue in 39 of 43 patients (91%). CONCLUSION(S): Expression of the protamine-2 gene may be a useful molecular marker to predict the presence of testicular sperm in patients with nonobstructive azoospermia. PMID- 10856466 TI - Factors associated with the formation of triploid zygotes after intracytoplasmic sperm injection. AB - OBJECTIVE: To determine whether clinical or laboratory factors influence development of triploid (3PN) zygotes after ICSI. DESIGN: Retrospective review. SETTING: The assisted reproductive technology program of Brigham and Women's Hospital. PATIENT(S): Patients undergoing ICSI. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Cycles were divided into two groups: group A, cycles with one or more 3PN zygotes after ICSI, and group B, cycles with no 3PN zygotes. Age, amount of gonadotropin administered, peak estradiol levels, number of follicles, number of oocytes retrieved and injected, time between retrieval and injection, oocyte abnormalities, sperm type and motile count, percentage of diploid zygotes, and ongoing pregnancy rates were compared between groups. RESULT(S): Compared with patients in group B, those in group A received fewer ampoules of gonadotropins, had higher estradiol levels, and had more follicles on the day of hCG administration, oocytes, immature oocytes and oocytes injected and lower percentages of diploid zygotes. However, ongoing pregnancy rates did not differ between groups. CONCLUSION(S): Patients who produce 3PN zygotes after ICSI are high responders to ovarian stimulation. The appearance of such embryos is not associated with lower ongoing pregnancy rates and should not necessarily dictate alterations in ovarian stimulation protocols. PMID- 10856467 TI - Rescue intracytoplasmic sperm injection (ICSI)-salvaging in vitro fertilization (IVF) cycles after total or near-total fertilization failure. AB - OBJECTIVE: To evaluate the effectiveness of delayed oocyte reinsemination by ICSI (rescue ICSI) after total or near-total fertilization failure (/=10 mm were found. The mean (+/-SD) number of follicles >/=8 mm on the day of maximum stimulation in the 150 IU and 225 IU s. c. and 150 IU i.m. groups were 14.0 +/- 7.1, 14.3 +/- 8.2, and 6.5 +/- 4.7. CONCLUSION(S): Pharmacokinetics of recombinant FSH were dose proportional within the dose range studied (75-225 IU). Subcutaneous and i.m. administration of 150 IU was bioequivalent with respect to pharmacokinetics, but after s.c. administration the number of growing follicles and estradiol response were higher. PMID- 10856482 TI - Interleukin-3 and human trophoblast: in vitro explanations for the effect of interleukin in patients with antiphospholipid antibody syndrome. AB - OBJECTIVE: To examine the effect of interleukin (IL)-3 on in vitro trophoblast differentiation, hormone production, and invasiveness affected by antiphospholipid antibodies. DESIGN: Primary cytotrophoblast cell cultures. SETTING: Obstetrics and Gynecology Department of the Catholic University, Rome, Italy. PATIENT(S): Five normal pregnant women underwent uncomplicated vaginal delivery at 36 weeks of gestation. INTERVENTION(S): Immunoglobulin (Ig) G antibodies were isolated from the plasma of two patients with antiphospholipid syndrome and two normal control subjects with the use of protein-G Sepharose columns. Cytotrophoblast cells were dispersed in Ringer's bicarbonate buffer containing trypsin and DNAseI, filtered, and layered over a Percoll gradient in Hank's balanced salt solution. MAIN OUTCOME MEASURE(S): We investigated the effects of IL-3 and antiphospholipid antibodies on trophoblast cell invasiveness, differentiation, and hormone secretion. RESULT(S): IgG obtained from patients with antiphospholipid syndrome bound to trophoblast cells, with inhibitory effects on the cells' invasiveness, differentiation, and hCG secretion. IL-3 was able to restore in vitro placental functions. CONCLUSION(S): These results imply that IL-3 favorably affects human trophoblast implantation and development. PMID- 10856483 TI - Association of histologic features and cytogenetic abnormalities in ectopic pregnancies. AB - OBJECTIVE: To evaluate the association between specific histologic features and cytogenetic abnormalities in ectopic pregnancies. DESIGN: Blinded histologic analysis. SETTING: University hospital. PATIENT(S): Fifty-four patients with ectopic pregnancy for whom successful karyotypes and sufficient histologic material were available. INTERVENTION(S): Histologic evaluation of chorionic villi from ectopic pregnancies was done by two pathologists who were unaware of the cytogenetic outcome. Seventeen histologic features were evaluated: villus size, villus contour, ghost villi, hydropic villi, trophoblastic hyperplasia, trophoblastic hypoplasia, syncytial knots, Hofbauer cells, blood vessels, trophoblastic lacunae, trophoblastic inclusions or cisterns, degeneration, fibrohyalinization, microcalcifications, and perivillous and intervillous fibrin deposits. MAIN OUTCOME MEASURE(S): The association between histopathologic features and cytogenetic outcome. RESULT(S): The presence of ghost villi and intervillous or perivillous fibrin was found to be associated with cytogenetic abnormalities. These features are associated with previous fetal cell death. CONCLUSION(S): This study does not support an association between specific histologic features of chorionic villi and cytogenetic abnormalities in ectopic pregnancies. The only histologic features that were associated with cytogenetic abnormalities (i.e., ghost villi and intervillous and perivillous fibrin) are merely a result of previous fetal cell death. PMID- 10856484 TI - Evidence of a T(H) 1 type response associated with recurrent miscarriage. AB - OBJECTIVE: To determine whether the T(H) 1 cytokine interferon (IFN)-gamma is associated with miscarriage whereas the T(H) 2 cytokine interleukin (IL)-10 is associated with successful pregnancy. DESIGN: Controlled clinical study. SETTING: Healthy volunteers in an academic setting. PATIENT(S): Group 1 comprised 10 nonpregnant women; group 2, 10 first-trimester primigravid women; group 3, 10 first-trimester primigravid women suffering spontaneous abortion; and group 4, 10 first-trimester pregnant women with a history of miscarriage. All women were pregnant at the time of sampling, but 5 miscarried later in the first trimester. INTERVENTION(S): None of the patients received any medication. MAIN OUTCOME MEASURE(S): Serum levels of IL-10 and IFN-gamma. RESULT(S): Levels of IL-10 were significantly raised in normal pregnancy. Levels of IFN-gamma were raised in the recurrent-miscarriage group as compared with normal pregnancy. When patients in group 4 were divided into those whose pregnancies went to term and those who miscarried, we found that successful pregnancy was associated with a statistically significant increase in IL-10, whereas miscarriage was associated with significantly increased levels of IFN-gamma. CONCLUSION(S): These results support the view that miscarriage is associated with a T(H) 1 type response. PMID- 10856485 TI - Outcome of preimplantation genetic diagnosis of translocations. AB - OBJECTIVE: To review 35 cases of preimplantation genetic diagnosis (PGD) of translocations with several methods, including telomeric probes. DESIGN: Retrospective study. SETTING: Clinical IVF laboratory. PATIENT(S): Thirty-five couples with one partner carrying a chromosomal translocation. INTERVENTION(S): PGD of translocation after polar-body or embryo biopsy. MAIN OUTCOME MEASURE(S): Pregnancy outcome. RESULT(S): Several trends were observed. First, PGD can achieve a statistically significant reduction in spontaneous abortion, from 95% to 13%. Second, the chances of achieving pregnancy are correlated with 50% or more of the embryos being chromosomally normal. Third, patients with robertsonian translocations produced fewer abnormal gametes and more pregnancies than did patients with reciprocal translocations. Fourth, a new fluorescence in situ hybridization protocol for PGD of translocations, which involves applying telomeric probes, has proved adequately reliable with a 6% average error rate. CONCLUSION(S): PGD of translocations achieves a statistically significant reduction in spontaneous abortion, both for polar-body and blastomere biopsy cases. Pregnancy outcome depended on the number of normal embryos available for transfer, with patients having <50% abnormal embryos achieving the most pregnancies. Because robertsonian translocations caused fewer abnormal embryos than reciprocal translocations, they also resulted in higher rates of implantation. PMID- 10856486 TI - Characterization of messenger RNA expression of estrogen receptor-alpha and -beta in patients with ovarian endometriosis. AB - OBJECTIVE: To evaluate the expression of estrogen receptor (ER)-alpha and ER-beta messenger RNA (mRNA) in ovarian endometriosis. DESIGN: Prospective study. SETTING: University hospital. PATIENT(S): Patients with endometriosis and patients with uterine leiomyoma or carcinoma in situ. INTERVENTION(S): A sample of ovarian endometriotic cyst tissue was obtained from each of the 32 patients during laparoscopic cystectomy. Samples of ovarian tissue and endometrium were obtained from 15 patients during or just after surgery. MAIN OUTCOME MEASURE(S): Expression of mRNA for ER-alpha and ER-beta with use of the reverse transcription polymerase chain reaction (RT-PCR) and nonradioactive in situ hybridization (ISH) techniques. RESULT(S): Expression of mRNA for ER-alpha and ER-beta was observed in all of the control tissues from the normal endometrium and normal ovaries in the RT-PCR and ISH analyses, although the distribution of positive signals changed in the ISH analysis during different phases of the menstrual cycle. Messenger RNA for ER-alpha was detected in all of the ovarian endometriotic cysts analyzed (19 of 19), but mRNA for the ER-beta was limited (12 of 19) in the RT PCR analysis. The ISH analysis confirmed the RT-PCR results and revealed that the two estrogen receptors were localized in both epithelial and stromal cells of endometriotic tissues. CONCLUSION(S): The results suggest that predominant expression of ER-alpha in both glandular epithelial and stromal cells may be essential to the development and growth of ovarian endometriosis. PMID- 10856487 TI - Seminal plasma characteristics as indicators of cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations in men with obstructive azoospermia. AB - OBJECTIVE: To determine the prognostic value of seminal plasma volume, pH, fructose, and alpha-glucosidase for the detection of cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations. DESIGN: Retrospective data analysis. SETTING: University infertility clinic (referral center). PATIENT(S): Fifty-nine men with obstructive azoospermia. INTERVENTION(S): Semen analysis including seminal plasma volume, pH, fructose, alpha-glucosidase, molecular genetic diagnosis of CFTR mutations and FSH measurement. MAIN OUTCOME MEASURE(S): Sensitivity and specificity of seminal plasma markers for the detection of CFTR mutations. RESULT(S): A CFTR mutation was detected in 26 of 59 patients with obstructive azoospermia. Patients carrying a mutation had significantly lower seminal plasma volume (mean +/- SEM: 1.5 +/- 1.4 mL vs. 2.8 +/- 2.2 mL), lower pH levels (25th percentile, median, 75th percentile: 6.5, 6.8, 7.5 vs. 7.7, 7. 9, 7.9) and lower fructose content (1.0, 1.1, 3.7 vs. 5.8, 20.0, 83. 0 micromol/ejaculate) than those without mutations. Diagnostic efficacy for detection of mutations was best (pH 81.4%, fructose 81. 8%) at a cutoff level for pH of 7.4 and fructose of 2 micromol/ejaculate. CONCLUSION(S): Seminal plasma markers provide an effective, noninvasive method to predict CFTR mutations in men with obstructive azoospermia. PMID- 10856488 TI - Interstitial pregnancy. PMID- 10856489 TI - Stepwise pain score analysis of the effect of local lignocaine on outpatient hysteroscopy: a randomized, double-blind, placebo-controlled trial. AB - OBJECTIVE: To assess the efficacy of lignocaine gel in reducing the overall pain and pain of individual steps during outpatient hysteroscopy in comparison with placebo (no anesthesia). DESIGN: A prospective, randomized, double-blind, placebo controlled trial. SETTING: Outpatient hysteroscopy clinic in a regional hospital in Hong Kong. PATIENT(S): A total of 500 Chinese patients undergoing outpatient hysteroscopy. INTERVENTION(S): Application of lignocaine gel to the cervix during outpatient hysteroscopy. MAIN OUTCOME MEASURE(S): Mean pain score using present pain intensity, overall pain score measured by total area under the curve, and the pain score of individual steps in the procedure in patients receiving lignocaine gel were compared with those of patients having no anesthesia. The failure rate and poor-view rate in both groups were also compared. RESULT(S): There were no statistically significant differences in mean pain score, overall pain score, and pain score of individual steps between the lignocaine group and controls. The failure rate and poor-view rate also showed no statistically significant differences. CONCLUSION(S): Outpatient hysteroscopy without anesthesia is acceptable to most Chinese women, and the local application of lignocaine gel is not effective in reducing pain. PMID- 10856490 TI - Laparoscopic repair of bilateral inguinal hernias in a patient with mullerian agenesis. AB - OBJECTIVE: To report a case of laparoscopic repair of bilateral inguinal hernias in a patient with mullerian agenesis. DESIGN: Case report. SETTING: Hospital. PATIENT(S): An 18-year-old woman with mullerian agenesis who presented with symptomatic inguinal hernias containing ovaries. INTERVENTION(S): Laparoscopic repair of bilateral inguinal hernias. MAIN OUTCOME MEASURE(S): Hernia repair was accomplished laparoscopically. RESULT(S): Outpatient laparoscopic repair of bilateral inguinal hernias with reduction of ovaries was performed using an extraperitoneal approach without complication. CONCLUSION(S): Laparoscopic approach of inguinal hernias can be an option for surgical repair in patients with mullerian agenesis even when the hernias contain ovaries. PMID- 10856491 TI - Pregnancy after embolization of uterine myoma: report of 12 cases. AB - OBJECTIVE: To treat uterine myomas with embolization, to look for pregnancy induced myoma recurrences, and to assess pregnancy course and outcome after embolization. DESIGN: Observational clinical study. SETTING: University of Paris VII hospital. PATIENT(S): Nine women had embolization for symptomatic myoma, with 12 pregnancies observed. INTERVENTION(S): Embolizations were highly selective and performed bilaterally through the uterine arteries with polyvinyl alcohol. MAIN OUTCOME MEASURE(S): Pregnant women were evaluated by physical and sonographic examinations. RESULT(S): Before embolization, the mean uterine volume was 450 cm(3), and in six patients polymyomas were present. The median age at embolization was 40 years; the median delay before pregnancy was 9 months; and the median age at first pregnancy outcome was 41 years. Five early miscarriages occurred. The seven other pregnancies were uneventful, although three premature births and one case of late toxemia occurred unrelated to previous embolization. Three women delivered vaginally and four by cesarean section. Neither myoma recurrence nor abnormality in uterine function was observed. CONCLUSION(S): The results of this first series of 12 pregnancies after myoma embolization are promising. If these preliminary results are confirmed, embolization could be a major breakthrough in the management of myoma and could replace conventional medical and surgical treatments. PMID- 10856492 TI - Two consecutive pregnancies during inadvertent gonadotropin-releasing hormone agonist desensitization. AB - OBJECTIVE: To describe two consecutive spontaneous pregnancies during luteal phase down-regulation with a GnRH-a. DESIGN: Case report. SETTING: University affiliated reproductive medicine unit. PATIENT(S): A 33-year-old woman with a history of failed uterine tubal reanastomosis. INTERVENTION(S): Four IVF cycles. MAIN OUTCOME MEASURE(S): Pregnancy. RESULT(S): The patient had two spontaneous pregnancies during luteal-phase down-regulation. CONCLUSION(S): Although midluteal administration of GnRH-a is not established as a treatment, there might be a beneficial effect on the endometrium via the medication's evoked gonadotropin flare. PMID- 10856493 TI - Decreased in vitro fertilization and cleavage rates after an equipment error during CO(2) calibration. AB - OBJECTIVE: To report the decreased IVF rate of oocytes and the reduced cleavage rate of pronuclear stage embryos after gas has backflowed from a gas analyzer into an incubator during the calibration of the CO(2) concentration. DESIGN: Case report. SETTING: Clinical research unit for reproductive medicine in a hospital. INTERVENTION(S): Backflow of gas from a gas analyzer into an incubator during calibration of the CO(2) concentration. MAIN OUTCOME MEASURE: The IVF rate of oocytes and the cleavage rate of pronuclear stage embryos. RESULT(S): Thirty-two oocytes were retrieved from four infertile female patients. Nine oocytes were fertilized and four fertilized oocytes were cleaved. In a related animal experiment, the cleavage rate was 57.1% (36/63) in the control group and 25.4% (16/63) in the study group (P =.00026). CONCLUSION(S): The backflow of gas from the gas analyzer adversely affected the fertilization of oocytes and the cleavage of pronuclear stage embryos. PMID- 10856494 TI - Vertebral osteomyelitis: a rare complication of transvaginal ultrasound-guided oocyte retrieval. AB - OBJECTIVE: To present a case of vertebral osteomyelitis as a complication of transvaginal oocyte retrieval. DESIGN: Case report. SETTING: The IVF unit of a university-affiliated hospital. PATIENT(S): A 41-year-old woman who underwent IVF ET treatment. INTERVENTION(S): Standard IVF-ET treatment cycles with the use of transvaginal ultrasound for oocyte retrieval and computed tomography-guided needle aspiration. MAIN OUTCOME MEASURE(S): Recovery of the patient, sequelae, and recurrence. RESULT(S): Vertebral osteomyelitis was diagnosed and treated with antibiotics. CONCLUSION(S): When severe low back pain occurs after ovum retrieval, vertebral osteomyelitis should be considered. Early diagnosis requires a high index of suspicion. PMID- 10856495 TI - Ovarian hyperstimulation after administration of triptorelin therapy to a patient with polycystic ovary syndrome. PMID- 10856496 TI - Presence of stem cell factor in follicular fluid and its expression in the human ovary. PMID- 10856497 TI - Polycystic ovary syndrome nomenclature: chaos? PMID- 10856498 TI - All that chocolate--but where did it come from? PMID- 10856499 TI - Bridges yet to cross. PMID- 10856500 TI - Insurance for infertility treatments--different perspectives. PMID- 10856501 TI - Chromatin remodelling during the life cycle of trypanosomatids. AB - The mechanisms which control the expression of developmentally regulated genes in trypanosomatids remain unclear. The genes are grouped together into transcription units that are co-transcribed to yield polycistronic RNAs. Trans-splicing and polyadenylation give rise to mature, monocistronic mRNAs. It is difficult to imagine that expression of these genes is controlled at the level of transcription initiation because this would suggest that the genes are transcribed at the same rate. This is not the case, because at any given developmental stage in trypanosomes or Leishmania, genes transcribed from the same transcription unit are expressed at different levels within the cell. Consequently, these parasites must rely on post-transcriptional or post translational mechanisms to generate the appropriate levels of gene product within the cell. There are no well-established examples of RNA polymerase II promoters in trypanosomes or Leishmania. However, the promoters for genes encoding the variant surface glycoprotein (VSG) and the procyclic acidic repetitive protein (PARP) have been identified and resemble ribosomal RNA polymerase I promoters. In higher eukaryotes where the mechanisms regulating transcription are clearer, there is increasing evidence that epigenetic factors, such as histones and modified bases, influence gene expression. Chemical modification of these factors can restructure chromatin and lead to gene activation or silencing. In trypanosomatids, an epigenetic mechanism for the control of developmentally expressed genes is a possibility. In this review, chromatin remodelling during the life and cell cycle of trypanosomes and Leishmania is explored, and the influence of epigenetic factors such as histones and modified bases on this process is discussed. PMID- 10856502 TI - Characterisation of the carbohydrate components of Taenia solium metacestode glycoprotein antigens. AB - Human neurocysticercosis is caused by Taenia solium metacestodes. It usually affects the central nervous system of humans and can be confused with other brain pathologies. The Lens culinaris-binding glycoproteins from this parasite have been shown to be ideal targets for the development of a highly specific immunoassay for the diagnosis of neurocysticercosis. In the present study we characterised the carbohydrates associated with five antigenic glycoproteins of T. solium metacestodes in the range of 12-28 kilodaltons. Lectin-affinities and enzymatic deglycosylations suggested that each of the five antigens contain various glycoforms of asparagine-linked carbohydrates of the hybrid, complex and probably high mannose type. These carbohydrates accounted for at least 30-66% of the apparent molecular mass of the glycoconjugates. In contrast, there was no evidence for the presence of O-linked carbohydrates. Lectin affinity patterns suggested that the sugars are short and truncated in their biosynthetic route, and that some contain terminal galactose moieties. Elucidating the precise structure of the carbohydrates and establishing their role in antigenicity will be essential to design strategies to produce them in large and reproducible amounts for the development of improved immunoassays. PMID- 10856503 TI - Metazoan parasite species richness and genetic variation among freshwater fish species: cause or consequence? AB - The factors responsible for the maintenance of genetic variation among natural populations remain a mystery. Recent models of host-parasite co-evolution assume that parasites exert frequency-dependent selection on their hosts by favouring rare alleles that may confer resistance against infection. We tested this prediction in a comparative analysis that sought relationships between levels of genetic variation and the number of metazoan parasite species exploiting each host species. We used data on 40 species of North American freshwater fishes. After controlling for sampling effort and phylogenetic influences, we found no relationship between genetic polymorphism and parasite species richness among fish species. However, we found a marginal negative correlation between parasite species richness and heterozygosity. This result goes against the prediction that increased selective pressure by parasites should be associated with higher levels of genetic variation. Instead, it suggests that parasites may be colonising host species showing low levels of genetic variation with greater success than genetically more variable host species. PMID- 10856504 TI - Identification and characterisation of the excreted/secreted serine proteases of larvae of the old world screwworm fly, Chrysomya bezziana. AB - Serine proteases are the major proteolytic activity excreted or secreted from Chrysomya bezziana larvae as demonstrated by gelatin gel analyses and the use of specific substrates, benzoyl-Arg-p-nitroanilide and succinyl-Ala-Ala-Pro-Phe-p nitroanilide. Serine proteases were identified through their inhibition by 4-(2 aminoethyl)-benzene sulphonyl fluoride and classified as trypsin- and chymotrypsin-like on the basis of inhibition by tosyl-L-lysine chloromethyl ketone and tosyl-L-phenylalanine chloromethyl ketone, respectively. Like most insect serine proteases, the C. bezziana enzymes were active over broad pH range from mildly acidic to alkaline. The excreted or secreted serine proteases were purified by affinity chromatography using soybean trypsin inhibitor. A different subset of the serine proteases was isolated by salt elution from washed larval peritrophic matrices. Amino-terminal sequencing identified both trypsin and chymotrypsin-like sequences in the excreted or secreted pool with the latter being the dominant protease, whereas trypsin was the dominant species in the peritrophic matrix eluant. These results suggest that trypsin was possibly preferably adsorbed by the peritrophic matrix and may act as a final proteolytic processing stage as partially digested and ingested polypeptides pass through the peritrophic matrix. Immunoblot analysis on dissected gut tissues indicated that the anterior and posterior midguts were the main source of the serine proteases, although a novel species of 32 kDa was predominantly associated with the peritrophic matrix. Proteases are a target for a partially protective immune response and understanding the complexity of the secreted and digestive proteases is a necessary part of understanding the mechanism of the host's immunological defence against the parasite. PMID- 10856505 TI - Studies on codon usage in Entamoeba histolytica. AB - Codon usage bias of Entamoeba histolytica, a protozoan parasite, was investigated using the available DNA sequence data. Entamoeba histolytica having AT rich genome, is expected to have A and/or T at the third position of codons. Overall codon usage data analysis indicates that A and/or T ending codons are strongly biased in the coding region of this organism. However, multivariate statistical analysis suggests that there is a single major trend in codon usage variation among the genes. The genes which are supposed to be highly expressed are clustered at one end, while the majority of the putatively lowly expressed genes are clustered at the other end. The codon usage pattern is distinctly different in these two sets of genes. C ending codons are significantly higher in the putatively highly expressed genes suggesting that C ending codons are translationally optimal in this organism. In the putatively lowly expressed genes A and/or T ending codons are predominant, which suggests that compositional constraints are playing the major role in shaping codon usage variation among the lowly expressed genes. These results suggest that both mutational bias and translational selection are operational in the codon usage variation in this organism. PMID- 10856506 TI - Inter- and intra-strain variation in the 5.8S ribosomal RNA and internal transcribed spacer sequences of Entamoeba histolytica and comparison with Entamoeba dispar, Entamoeba moshkovskii and Entamoeba invadens. AB - The ribosomal RNA genes in Entamoeba histolytica are located on circular DNA molecules in about 200 copies per genome equivalent. Nucleotide sequence analysis of the 5.8S rRNA gene and the flanking internal transcribed spacers was carried out to determine the degree of sequence divergence in the multiple rRNA gene copies of a given strain; amongst three different E. histolytica strains (HM 1:IMSS, Rahman and HK-9); and amongst four species of Entamoeba (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii and Entamoeba invadens). The results show that all rRNA gene copies of a given strain are identical. Few nucleotide positions varied between strains of a species but the differences were very pronounced amongst species. In general, the internal transcribed spacer 2 sequence was more variable and may be useful for strain- and species identification. The 5.8S rRNA gene and the internal transcribed spacer 2 of E. invadens were unusually small in size. PMID- 10856507 TI - Molecular cloning of a gene encoding a 20S proteasome beta subunit from Plasmodium falciparum. AB - A novel gene was cloned from Plasmodium falciparum. Database searches indicated this gene to be a member of the 20S proteasome beta-subunit family. Comparison of the gene's genomic DNA sequence with cDNA sequence revealed a 156-bp intron 85 bp downstream from the start codon. The nucleotide sequence of the gene contains one open reading frame encoding 265 amino acids with a predicted molecular mass of 30.9 kDa and a pI of 6.2. Northern blot analysis showed the transcript size to be approximately 1.6 kb indicating that some 800 bp of the transcript is non-coding. PMID- 10856508 TI - Localisation of parasite antigens and inflammatory responses in experimental opisthorchiasis. AB - The time course localisation of parasite antigens and related host pathology were studied in hamsters infected with 100 metacercariae of Opisthorchis viverrini for up to 6 months. Parasite antigens, as detected by immunofluorescence and/or immunoperoxidase-staining, were first observed in the flukes and the biliary epithelium of the intrahepatic and extrahepatic bile ducts as early as day 3 p.i. Antigens increased as the parasite matured, both in tissues in direct contact with the flukes and those surrounding the infection. Opisthorchis antigens were also observed in the first order bile ducts (small bile ducts) of the liver, which are not normally inhabited by flukes. In addition, they were found in damaged liver cells, Kupffer cells, macrophages, and within epithelioid and giant cells in the egg granuloma. The presence of the antigens was associated with heavy inflammatory cell infiltration, particularly with mononuclear cells. The results strongly support the role of fluke-associated antigens and local parasite specific immune responses in the pathogenesis of opisthorchiasis. PMID- 10856509 TI - Comparison of ribosomal DNA sequences of Lamellodiscus spp. (monogenea, diplectanidae) parasitising Pagellus (sparidae, teleostei) in the North Mediterranean Sea: species divergence and coevolutionary interactions. AB - We sequenced DNA fragments from four monogenean species of the genus Lamellodiscus and their three fish host species from the genus Pagellus in the North Mediterranean Sea, in order to estimate the molecular divergence and the coevolutionary interactions in this association. By comparing the ITS1 sequences of the parasites, we assessed their level of interspecific differences and tested the phylogenetic status of Lamellodiscus virgula and Lamellodiscus obeliae, formerly described as two different species. Moreover, we wanted to know if closely related parasites used closely related hosts, to investigate the coevolutionary interactions in this complex. Phylogenetic relationships among Lamellodiscus species were estimated with partial 18S ribosomal DNA sequences while mitochondrial cytochrome-b DNA sequences were used for their fish hosts. The ITS1 sequences appear to be highly variable among Lamellodiscus species, except L.virgula and L.obeliae, suggesting an old divergence time or a rapid molecular evolution within this genus. This fish-parasite association seems to exhibit coevolutionary interactions. L.virgula and L.obeliae are proposed to be a single species on the basis of their almost identical ITS1 sequences. PMID- 10856510 TI - The life cycle of Opecoeloides columbellae (Pagenstecher, 1863) n. comb. (Digenea, opecoelidae): evidence from molecules and morphology. AB - The cosmopolitan digenean family Opecoelidae comprises several hundred species, whose adults live in the digestive tract of marine and freshwater fishes. The genus Opecoeloides Odhner, 1928 is represented in the Mediterranean by a single species, Opecoeloides furcatus (Bremser in Rudolphi, 1819), that has been recorded from six definitive hosts species. To see if this broad host range could be the result of an underestimation of species diversity, we obtained ITS1 ribosomal DNA sequences as well as morphological data from adult specimens of O. furcatus isolated from two definitive hosts species: Mullus surmuletus and Gaidropsarus mediterraneus. Sequence and morphological data were also obtained from several opecoelid cercariae and metacercariae occurring in different invertebrate hosts. The data presented here provide striking evidence that O. furcatus specimens isolated from the two host fishes represent distinct species. This argument is reinforced by the fact that cercariae corresponding to each of these adult species were found in two molluscan host-species, Columbella rustica and Mitrella scripta. These parasite species differ by several nucleotide substitutions and a 60 bp-long insertion in the ITS1. They also show clear morphological differences in testis and ovary shape, as well as in their mean dimensions. Here, we attribute the adult specimens found in G. mediterraneus to Opecoeloides columbellae (Pagenstecher, 1863) n. comb. This species was described and compared with O. furcatus from M. surmuletus. ITS1 sequence comparison allowed identification of the cercaria (occurring in C. rustica) and metacercaria (occurring in Hippolyte inermis) of O. columbellae n. comb. PMID- 10856511 TI - Anti-malarial effect of histone deacetylation inhibitors and mammalian tumour cytodifferentiating agents. AB - The histones of Plasmodium falciparum represent a potential new target for anti malarial compounds. A naturally occurring compound, apicidin, has recently been shown to inhibit the in vitro growth of P. falciparum. Apicidin was shown to hyperacetylate histones, suggesting that its mode of action is through histone deacetylase inhibition. We have tested the ability of known histone deacetylase inhibitors, mammalian tumour suppressor compounds, and cytodifferentiating agents to inhibit the in vitro growth of a drug sensitive and resistant strain of P. falciparum. Seven of the tested compounds had microM IC50 values, and trichostatin A, a histone deacetylation inhibitor and cytodifferentiating agent, was active at low nM concentrations. One compound, suberic acid bisdimethylamide, which selectively arrests tumour cells as opposed to normal mammalian cells, had an in vivo cytostatic effect against the acute murine malaria Plasmodium berghei, and one round of treatment with the compound failed to select for resistant mutations. These results suggest a promising role for histone deacetylase inhibitors and cytodifferentiating agents as antimalarial drug candidates. PMID- 10856512 TI - The binding of closantel to ovine serum albumin, and homogenate fractions of Haemonchus contortus. AB - Closantel binds to the serum proteins of the host and affects blood sucking parasites when they ingest the blood of treated hosts. Closantel binds specifically to ovine serum albumin (K(a) of 9. 3x10(6)M(-1)) at site I, the warfarin/phenylbutazone binding site of albumin Closantel also binds to invertebrate haemocyanin and haemolymph. The strongest binding of closantel in homogenates of H. contortus is found in fractions containing soluble proteins. This binding is of low affinity and, because the site itself is not fully denaturable, it may not be proteinaceous. There is no detectable difference in binding affinity between homogenate fractions from closantel susceptible and resistant isolates of adult or larval worms suggesting that closantel resistance is not due to changes in the closantel receptor or carrier. PMID- 10856513 TI - Therapeutic evaluation of free and liposome-encapsulated atovaquone in the treatment of murine leishmaniasis. AB - The use of drug delivery systems may reduce the toxicity and improve the activity of anti-leishmanial compounds. The activity of atovaquone (ATV)-loaded liposomes was compared by determination of median effective doses (ED(25) and ED(50)), with that of free ATV in a murine model of visceral leishmaniasis induced by Leishmania infantum. On day 0, mice were infected intravenously with 4.10(7) promastigotes and treated via the tail vein on days 15, 17 and 19 by free drug in a DMSO/cremophor/water solution (0.2 to 1.6 mg/kg body weight) or by liposomal drug (0.04 to 0.32 mg/kg body weight). Mice were killed and livers and spleens were removed and weighed on day 21 p.i. and liver parasite burdens evaluated using the Stauber method. Effective doses were determined using the Hill representation relating the percentage of parasite suppression to the dose. Liposomal ATV was significantly more effective than the free drug in reducing liver parasites (61.6% of parasite suppression at a dose of 0.32 mg/kg vs 34.9% at a dose of 1.6mg/kg). Liposomal ATV was 23 times more active than the free drug (ED(25) value=0. 02+/-0.01 mg/kg vs 0.46+/-0.15 mg/kg for free drug). It was not possible to obtain the ED(50) for free ATV because the dose-response curve reached a plateau around 33% of parasite suppression. Conversely, the ED(50) for liposomal ATV was 0.17+/-0.05 mg/kg. 100% efficacy of bound ATV could be obtained with a concentration of 1. 77+/-0.35 mg/kg. A significant decrease in spleen weights was also observed reflecting a leishmanicidal activity of ATV. These results suggest that liposome loaded ATV is more efficacious than the free drug against Leishmania infantum in this murine model. PMID- 10856514 TI - Host specificity in blood feeding parasites: a defining contribution by haemoglobin-degrading enzymes? AB - A hypothesis is presented that proposes that the compatibility between species specific variants of haemoglobin-degrading proteases of blood-feeding parasites (e.g. hookworms, schistosomes, malarial parasites, etc.), and their natural substrates, i.e. haemoglobins from diverse species of mammals, has influenced to evolution of the host range of these parasites. Support for the hypothesis was drawn from molecular modelling studies of the three dimensional structure of an aspartic protease, Acasp, from the canine hookworm Ancylostoma caninum, and models of canine and human haemoglobins docked with the active site of Acasp. The molecular modelling suggested that Acasp, from a canine-specific hookworm, would have a higher substrate affinity for canine haemoglobin than for human haemoglobin. PMID- 10856515 TI - Relationship of dietary fat to glucose metabolism. AB - The relationship between dietary fat and glucose metabolism has been recognized for at least 60 years. In experimental animals, high fat diets result in impaired glucose tolerance. This impairment is associated with decreased basal and insulin stimulated glucose metabolism. Impaired insulin binding and/or glucose transporters has been related to changes in the fatty acid composition of the membrane induced by dietary fat modification. In humans, high-fat diets, independent of fatty acid profile, have been reported to result in decreased insulin sensitivity. Saturated fat, relative to monounsaturated and polyunsaturated fat, appears to be more deleterious with respect to fat-induced insulin insensitivity. Some of the adverse effects induced by fat feeding can be ameliorated with omega-3 fatty acid. Epidemiological data in humans suggest that subjects with higher intakes of fat are more prone to develop disturbances in glucose metabolism, type 2 diabetes or impaired glucose tolerance, than subjects with lower intakes of fat. Inconsistencies in the data may be attributable to clustering of high intakes of dietary fat (especially animal fat) with obesity and inactivity. Metabolic studies suggest that higher-fat diets containing a higher proportion of unsaturated fat result in better measures of glucose metabolism than high-carbohydrate diet. Clearly, the area of dietary fat and glucose metabolism has yet to be fully elucidated. PMID- 10856516 TI - Atherosclerotic arterial remodeling and the localization of macrophages and matrix metalloproteases 1, 2 and 9 in the human coronary artery. AB - Atherosclerotic luminal narrowing is determined by plaque mass and the mode of geometrical remodeling. Recently, we reported that the type of atherosclerotic remodeling is associated with the presence of histological markers for plaque vulnerability. Inflammation and matrix degrading proteases (MMPs) may play a role in both plaque vulnerability and in expansive arterial remodeling. The aim of the present study was to investigate the association between the remodeling mode and the localization of macrophages and MMPs in coronary atherosclerotic segments. From 36 atherosclerotic coronary arteries, 45 and 51 segments were selected with a vessel area that was >10% smaller and larger compared with the adjacent segments, respectively. No significant difference in staining for macrophages was observed between segments with expansive and constrictive remodeling. More MMP-2 and MMP-9 staining was observed in plaques of expansively remodeled segments compared with constrictively remodeled segments. In general, MMP-staining was less evident in the adventitial layer compared with the plaque. Zymography revealed more active MMP-2 in expansively remodeled segments compared with constrictively remodeled segments (340+/-319 vs. 199+/-181 (adjusted counts/mm(2)), respectively, P=0.019). Zymography did not show differences in inactive MMP-2 or MMP-9 among groups. It might be postulated that MMPs within the plaque play a causal role not only in plaque vulnerability but also in de novo atherosclerotic remodeling. PMID- 10856517 TI - The effects of fatty acids on apolipoprotein B secretion by human hepatoma cells (HEP G2). AB - We have investigated the effect of fatty acids on the rate of apolipoprotein B (apo B) secretion by human hepatoma cells (Hep G2). When Hep G2 cells were maintained in tissue culture flasks oleic acid up to 0.4 mM increased apo B secretion in a dose-dependent manner, whereas increases in triacylglycerol (TG) were smaller and dose dependency was less evident. In the absence of oleic acid, apo B accumulating in the tissue culture medium was predominantly in lipoproteins of higher density than very low density lipoproteins (VLDL). However, when the rate of secretion was stimulated with oleic acid the apo B-containing lipoproteins became lower in density. We postulated that there was a high rate of lipolysis of newly secreted VLDL by Hep G2 cells, which would account both for the relatively smaller effect of oleic acid on TG as opposed to apo B accumulating in the culture medium and the predominance of apo B in lipoproteins of a higher density than VLDL, which became less evident when VLDL secretory rates were stimulated by oleic acid. To test this hypothesis, cultured Hep G2 cells were transferred to columns containing Cytodex beads, permitting their continuous perfusion with culture medium so that newly secreted VLDL did not remain in contact with the cells. Apo B recovered from the perfusate was largely in VLDL range lipoproteins and the TG measured in the perfusate indicated that the true secretory rate of TG-rich lipoproteins was substantially higher than had been reflected by TG accumulating in culture medium left in contact with cells. Apo B measured in the culture medium of Hep G2 cells may thus be a better reflection of VLDL secretion, even though it is contained in higher density lipoproteins due to removal of TG by lipolysis. The effects of saturated fatty acids (SFA), monounsaturated fatty acids (MUFA) and polyunsaturated fatty acids (PUFA) on apo B (apo B) secretion by Hep G2 cells maintained in tissue culture flasks were next investigated. SFA (0.4 mM), with the exception of stearic acid (C18:0), increased apo B secretion. Lauric acid (C12:0) increased apo B secretion by 32%, myristic acid (C14:0) by 41% (P<0.005), palmitic acid (C16:0) by 154% (P<0.025), and arachidic acid (C20:0) by 186% (P<0.005). The effect of MUFA (0.4 mM) was to increase apo B secretion, oleic acid (C18:1) by 239% ((P<0.0005) and palmitoleic acid (C16: 1) by 125% (P<0.005). Of the PUFA investigated, linolenic acid (C18:3) (0.4 mM) did not have any significant effect on apo B secretion, whereas linoleic acid (C18:2) (0.4mM) arachidonic acid (C20:4) (0.1 mM) and eicosapentaenoic acid (C20:5) (0.1 mM) caused significant increases of 164, 171 and 171%, respectively (P<0.005). The fatty acids studied increased intracellular TG and cholesteryl ester concentrations to varying extents. The increase in intracellular TG produced by the different fatty acids correlated with the rate of apo B secretion (r=0.6; P<0.05). In this human hepatoma cell line, with the exception of the saturated fatty acids, the rate of secretion of apo B-containing lipoproteins does not follow the same pattern as changes in circulating low density lipoprotein (LDL) concentrations reported with dietary manipulation in man. If our findings reflect the in vivo situation, we suggest that whilst the dietary effects of SFA on serum LDL may in part be determined by the hepatic apo B secretory rate, the effects of MUFA and PUFA must be largely mediated through a catabolic effect rather than an effect on hepatic secretion. The marked increase in apo B secretion with the more highly polyunsaturated fatty acids, such as eicosapentaenoic acid, may also explain why they do not lower circulating LDL, despite reports of their apparently favourable effect on LDL-receptor mediated clearance. PMID- 10856518 TI - The effect of carotenoids on the expression of cell surface adhesion molecules and binding of monocytes to human aortic endothelial cells. AB - Several large epidemiological studies have shown a correlation between elevated plasma carotenoid levels and decreased risk of cardiovascular disease (CVD). One proposed mechanism for the beneficial effect of carotenoids is through functional modulation of potentially atherogenic processes associated with the vascular endothelium. To test this, we incubated confluent human aortic endothelial cell (HAEC) cultures (passages 4-8) for 24 h with each of the five most prevalent carotenoids in human plasma, which are alpha-carotene, beta-carotene, beta cryptoxanthin, lutein, and lycopene, at an approximate concentration of 1 micromol/l. Carotenoids were solubilized in 0.7% (v/v) tetrahydrofuran and incorporated into FBS before adding to cell culture medium. Due to disparate solubilities in aqueous medium, final concentrations of alpha-carotene, beta carotene, beta-cryptoxanthin, lutein, and lycopene were 1.7, 1.1, 0.7, 0.9, and 0.3 micromol/l and monolayers accumulated 647, 158, 7, 113, and 9 pmol/mg protein, respectively. Monolayers were then stimulated with IL-1beta (5 ng/ml) for 6 h with subsequent determination of cell surface expression of adhesion molecules as measured by an enzyme-linked immunosorbent assay (ELISA). To assess endothelial cell adhesion to monocytes, IL-1beta-stimulated monolayers were incubated for 10 min with 51Cr-labeled U937 monocytic cells and adhesion determined by isotope counting. Pre-incubation of HAEC with beta-carotene, lutein and lycopene significantly reduced VCAM-1 expression by 29, 28, and 13%, respectively. Pre-incubation with beta-carotene and lutein significantly reduced E-selectin expression by 38 and 34%, respectively. Pre-treatment with beta carotene, lutein and lycopene significantly reduced the expression of ICAM-1 by 11, 14, and 18%, respectively. While other carotenoids were ineffective, lycopene attenuated both IL-1beta-stimulated and spontaneous HAEC adhesion to U937 monocytic cells by 20 and 25%, respectively. Thus, among the carotenoids, lycopene appears to be most effective in reducing both HAEC adhesion to monocytes and expression of adhesion molecules on the cell surface. PMID- 10856519 TI - Ascorbic acid enhances 17 beta-estradiol-mediated inhibition of oxidized low density lipoprotein formation. AB - Postmenopausal women who use estrogen appear to be protected from coronary heart disease (CHD). Studies have demonstrated that estrogen can lower low-density lipoprotein (LDL) levels and the antioxidant activity of 17 beta-estradiol can prevent the oxidation of this LDL. Ascorbic acid is regarded as a major hydrophylic antioxidant, however, its impact on the prevention of CHD has yet to be clearly demonstrated. Modified low density lipoprotein (LDL(-)) is an important marker of LDL oxidation in vivo, since it contributes to the oxidative susceptibility of low density lipoprotein, and at physiological levels displays pro-inflammatory and cytotoxic properties. Previously we showed that women taking estrogen replacement therapy have lower LDL(-) levels along with lower predisposition of the LDL to oxidize. In this study, we evaluated the potential action of 17 beta-estradiol (E(2)) in combination with ascorbic acid (AA) measured on the basis of LDL oxidative susceptibility in vitro and in the presence of cultured cells. High concentrations of E(2) were able to inhibit LDL oxidation, whereas in the presence of ascorbic acid nano- to picomolar levels of E(2) were sufficient to suppress LDL oxidation (P<0.05). Preconditioning male aortic endothelial cells (RAEC) with 5 ng/ml of E(2) (E(2)RAEC) reduced the formation of LDL(-) (P<0.005), and a more extensive inhibition was found in the presence of AA (P<0.0001). Interestingly, E(2) enhanced the uptake of LDL in the absence or presence of AA, however, this was not seen for the uptake of LDL(-). These results provide the first evidence that ascorbic acid can enhance the antioxidant effect of E(2) by preventing LDL oxidation by copper ions or cells. The cytoprotective and antiatherogenic effect of E(2) appears to involve a reduction in the extent of oxidized LDL formation and uptake. The enhanced activity of E(2) in the presence of ascorbate indicates that the antioxidant and antiatherosclerosis activity of E(2) may occur at concentrations within the physiological range. PMID- 10856520 TI - The effects of N-6 polyunsaturated fatty acid supplementation on the lipid composition and atherogenesis in mouse models of atherosclerosis. AB - Despite numerous studies, the precise role of dietary n-6 polyunsaturated fatty acids in the pathogenesis of atherosclerosis remains controversial. It has been shown that feeding an n-6-enriched diet resulted in decreased atherosclerosis in African green monkeys and was associated with a reduction in LDL levels. However, other authors reported that n-6 supplementation increased the oxidative stress and the susceptibility of LDL to undergo in vitro oxidation, thus potentially enhancing atherosclerosis. The present study was designed to investigate the effect of dietary supplementation of n-6 polyunsaturated fats (safflower oil), as compared with a saturated fat-rich diet (Paigen), on the blood lipid profile and atherosclerosis in two mouse models. In the first experiment, female C57BL/6 mice (n=23-30 per group) were fed a cholate containing Paigen diet, a safflower oil rich diet (with cholate), or normal chow for 15 weeks. No significant differences between the high fat diet groups were evident with respect to total cholesterol, LDL, HDL or triglyceride levels. The extent of aortic sinus fatty streaks did not differ significantly between the two groups. In the second experiment, LDL receptor-deficient (LDL-RD) mice (n=20-30 per group) were randomized into similar dietary regimens. Mice consuming a safflower oil-enriched diet developed significantly less atherosclerosis, in comparison with Paigen diet-fed mice. A reduction in LDL levels, although not of a similar magnitude as the reduction in atherosclerosis, was evident in the safflower oil-fed mice when compared to the Paigen diet-fed littermates. In both mouse models of atherosclerosis, LDL isolated from the plasma of mice on the n-6 polyunsaturated diet was rendered slightly more susceptible to oxidation in vitro, as indicated by a shorter lag period for diene formation. Thus, the effects of n-6 fatty acids on the lipoprotein composition and other potential influences may have contributed to the anti-atherogenic effect in the LDL-RD mouse model. PMID- 10856521 TI - A polymorphism upstream from the human paraoxonase (PON1) gene and its association with PON1 expression. AB - Human serum paraoxonase (PON1) is an esterase that has been shown to decrease the susceptibility of lipoproteins to lipid peroxidation. We found a polymorphism of cytosine/thymidine (-108C/T, the number is from the ATG codon) in the upstream region of the PON1 gene. The luciferase activity was lower in the -108T allele than in the -108C allele. The serum PON1 concentrations in 132 normal subjects were as follows: -108CC>-108CT and>-108TT genotypes. The polymorphism upstream from the PON1 gene is associated with transcription of the PON1 gene and the serum PON1 concentration. PMID- 10856522 TI - Glycation amplifies lipoprotein(a)-induced alterations in the generation of fibrinolytic regulators from human vascular endothelial cells. AB - Increased lipoprotein(a) [Lp(a)] in plasma is an independent risk factor for premature cardiovascular diseases. The levels of glycated Lp(a) are elevated in diabetic patients. The present study demonstrated that glycation enhanced Lp(a) induced production of plasminogen activator inhibitor-1 (PAI-1), and further decreased the generation of tissue-type plasminogen activator (t-PA) from human umbilical vein endothelial cells (HUVEC) and human coronary artery EC. The levels of PAI-1 mRNA and its antigen in the media of HUVEC were significantly increased following treatments with 5 microgram/ml of glycated Lp(a) compared to equal amounts of native Lp(a). The secretion and de novo synthesis of t-PA, but not its mRNA level, in EC were reduced by glycated Lp(a) compared to native Lp(a). Treatment with aminoguanidine, an inhibitor for the formation of advanced glycation end products (AGEs), during glycation normalized the generation of PAI 1 and t-PA induced by glycated Lp(a). Butylated hydroxytoluene, a potent antioxidant, inhibited native and glycated Lp(a)-induced changes in PAI-1 and t PA generation in EC. The results indicate that glycation amplifies Lp(a)-induced changes in the generation of PAI-1 and t-PA from venous and arterial EC. This may attenuate fibrinolytic activity in blood circulation and potentially contributes to the increased incidence of cardiovascular complications in diabetic patients with hyperlipoprotein(a). EC-mediated oxidative modification and the formation of AGEs may be implicated in glycated Lp(a)-induced alterations in the generation of fibrinolytic regulators from vascular EC. PMID- 10856523 TI - Effect of hypercholesterolemia on the sequential changes of apoptosis and proliferation after balloon injury to rabbit iliac artery. AB - To evaluate the effect of hypercholesterolemia on apoptosis and proliferation after vascular injury, iliac arteries of hypercholesterolemic (HC) and normocholesterolemic (NC) rabbits were examined after balloon injury using TUNEL, immunohistochemical staining of PCNA, macrophages, smooth muscle actin and p53. In media, apoptosis occurred massively early after injury and then decreased. HC did not affect this early post-injury apoptosis but significantly increased apoptosis 14 days later (D14). Immediate apoptosis in media was followed by active proliferation. HC sustained a high activity of proliferation until D14. The changes of immunoreactivity to p53 over the same 14 day period parallel that of apoptosis. In intima, where cells were scarce initially, proliferative activity reached a peak at D7 and then decreased. HC significantly enhanced proliferation at D14. In intima proliferation was accompanied by a later low level apoptosis. HC significantly enhanced this low-level apoptosis at D14. These effects of HC resulted in significantly increased areas of intima and media. The fundamental difference between HC and NC was the infiltration of macrophages in HC. In conclusion, balloon injury induces early massive p53-associated apoptosis followed by proliferation in media, whereas in intima, it induces active proliferation followed by a low-level apoptosis. Hypercholesterolemia does not affect the early post-injury apoptosis but enhances proliferation and low-level apoptosis at a later stage, which in turn results in intimal and medial hyperplasia. PMID- 10856524 TI - High resolution ex vivo magnetic resonance imaging of in situ coronary and aortic atherosclerotic plaque in a porcine model. AB - Atherosclerotic plaque composition is central to the pathogenesis of plaque disruption and acute thrombosis. Thus, there is a need for accurate imaging and characterization of atherosclerotic lesions. Even though there is no ideal animal model of atherosclerosis, the porcine model is considered to most closely resemble human atherosclerosis. We report the feasibility of MR imaging and characterizing of atherosclerotic lesions from in situ coronary arteries and aortas in an ex vivo setting and validate this with histopathology. Coronary and aortic atherosclerosis was induced in Yucatan mini-swine (n=4) by a combination of atherogenic diet (6 months) and balloon injury. All coronary arteries were imaged ex vivo on the intact heart, preserving the curvature of their course. The aorta also underwent MR imaging. The MR images were correlated with the matched histopathology sections for both the coronary arteries (n=54) and the aortas (n=43). MR imaging accurately characterized complex atherosclerotic lesions, including calcified, lipid rich, fibrocellular and hemorrhagic regions. Mean wall thickness for the coronary arteries (r=0.94, slope: 0.81) and aortas (r=0.94, slope: 0.81) as well as aortic plaque area (r=0.97, slope: 0.90) was accurately determined by MR imaging (P<0.0001). Coronary artery MR imaging is not limited by the curvature of the coronary arteries in the heart. MR imaging accurately quantifies and characterizes coronary and aortic atherosclerotic lesions, including the vessel wall, in this experimental porcine model of complex atherosclerosis. This model may be useful for future study of MR imaging of atherosclerosis in vivo. PMID- 10856525 TI - Differences in the effects of HMG-CoA reductase inhibitors on proliferation and viability of smooth muscle cells in culture. AB - We investigated the influence of lovastatin, simvastatin and pravastatin on proliferation and viability of vascular smooth muscle cells (SMC) in vitro and studied the effects of lovastatin on a mouse SMC line transgenic for a temperature-sensitive mutant of SV40 large T antigen (TAg), known to inhibit the function of p53 and pRb family members. We found that lovastatin and simvastatin inhibited cell proliferation by provoking G0/G1 phase arrest with concomitant depression of the proliferation antigen Ki-67/MIB-1. Lovastatin at high concentrations of 20 micromol/l caused cell death in the presence of serum but not under serum starved conditions, which was verified on the basis of increased DNA strand breaks, decreased DNA content and morphological alterations seen by electron microscopy. Cell death was also found for simvastatin, whereas pravastatin did not exhibit antiproliferative or cytotoxic effects. Mouse SMC transgenic for TAg did not show any impaired sensitivity to the antiproliferative and cell death inducing effect of lovastatin, but both effects could be antagonized by the supplementation of mevalonate. The data indicate that antiproliferative and cytotoxic effects of lovastatin are caused by the using up of products of mevalonate metabolism and do not require the presence of p53 or pRb. PMID- 10856526 TI - In vivo and in vitro evidence for the glycoxidation of low density lipoprotein in human atherosclerotic plaques. AB - Although there have been suggestions that the glycation and oxidation of low density lipoprotein (LDL) might increase its atherogenic potential, little is known about the presence of glycoxidative LDL in human atherosclerotic lesions. We developed specific antibodies against different immunological epitopes of AGE structures, including N(epsilon)-(carboxymethyl)lysine-protein adduct (CML), a glycoxidation product, and structure(s) other than CML (nonCML), and a monoclonal antibody against oxidized phosphatidylcholine (oxPC), as an epitope of oxidized LDL. Immunohistochemical analysis demonstrated that the CML- and oxPC-epitopes were accumulated mainly in macrophage-derived foam cells in atherosclerotic lesions, including fatty streaks and atherosclerotic plaques. On the other hand, the nonCML-epitope and apolipoprotein B were localized mainly in extracellular matrices of atherosclerotic lesions. The CML- and oxPC-epitopes were characterized by a model antigen-generating system using the copper ion-induced peroxidation and/or glucose-induced glycation of LDL. The glycoxidation of LDL caused the formation of CML-epitope with increasing concentrations of copper ion and glucose. It was also formed to some extent in LDL incubated with high concentrations (500 mM) of glucose. However, no CML-epitope was observed in oxidized LDL induced by copper ion alone. On the other hand, the formation of oxPC-epitope in LDL was dependent on copper ion-induced peroxidation, but independent of glucose-induced glycation. The addition of chelators, ethylenediaminetetraacetic acid and diethylenetriaminepentaacetic acid, reduced the increase in electrophoretic mobility and TBARS caused by the peroxidation and glycoxidation of LDL, but had no effects on the formation of fructosamine caused by the glycation and glycoxidation of LDL. Chelators as well as aminoguanidine protected the formation of CML-epitope in glycated or glycoxidative LDL. Although the formation of oxPC-epitope was completely inhibited by the addition of chelators, it was partially protected by aminoguanidine. These in vitro results suggest that the glycoxidative modification of LDL may occur in the arterial intima, and may contribute to the development of human atherosclerotic lesions. PMID- 10856527 TI - Calcium and lipoprotein lipase synergistically enhance the binding and uptake of native and oxidized LDL in mouse peritoneal macrophages. AB - The influence of Ca(2+) and Mg(2+), together with lipoprotein lipase (LPL), on the binding and uptake of Eu(3+)-labeled native and oxidized low density lipoprotein (LDL) to mouse peritoneal macrophages (MPM), and on the deposition of esterified cholesterol in these macrophages, were studied. We found that both LPL and Ca(2+) (but not Mg(2+)) increased the binding and uptake of native and mildly or moderately oxidized LDL, and the subsequent deposition of cholesterol esters in MPM. When added together, LPL and Ca(2+) synergistically increased the binding and uptake of native and oxidized LDL, and the deposition of esterified cholesterol derived from native and mildly or moderately oxidized LDL, in MPM. Since both calcium and LPL are found in the atherosclerotic lesions, our results suggest that Ca(2+) and LPL may synergistically promote foam cell formation and atherogenesis. Furthermore, future research in the metabolism of lipoproteins should take into account the calcium levels in the experimental conditions. PMID- 10856528 TI - Relationships between fibrinogen and insulin resistance. AB - A relationship between plasma fibrinogen levels and insulinemia, as well as the different parameters of the insulin resistance syndrome has been described. The aim of the present paper was to investigate whether plasma fibrinogen concentrations were linked to plasma insulin levels or to the degree of insulin resistance. For this purpose, 62 nondiabetic, nonhypertensive patients, 30 men and 32 women, with body mass indexes (BMIs) and ages ranging from 18.6 to 50.2 kg/m(2) and from 19 to 60 years, respectively, were studied. Insulin sensitivity was quantified by the minimal model procedure over a 180-min intravenous glucose tolerance test with iterative sampling. Plasma insulin was determined by radioimmunoassay without cross-reactivity to human proinsulin, and fibrinogen by the method of Clauss. Insulin sensitivity ranged from 0.009 to 23.2 min( 1)/(microU/ml)x10(-4), covering the whole range of insulin sensitivities. Fibrinogen ranged from 1.70 to 5.07 g/l. There was a significant negative correlation between fibrinogen and insulin sensitivity (r=-0.76,P<0.0001) and a positive correlation between fibrinogen and basal insulin (r=0.56,P<0.0001). After adjustment for BMI, body fat mass and waist-to-hip ratio, these two relationships remained significant. In addition, a multiple regression analysis was performed to assess the independent effect of the following related variables: fibrinogen, insulin sensitivity, insulinemia and BMI. Only insulin sensitivity appeared to account for the ability to predict fibrinogen values. Thus, we hypothesized it was likely that the state of insulin resistance rather than hyperinsulinemia per se was related to hyperfibrinogenemia. We proposed an interpretation of these data in connection with some factors like free fatty acids or tumor necrosis factor-alpha, which have been implicated in the pathogenesis of insulin resistance. Nevertheless, prospective and intervention studies are needed to assess whether there is a simple association or a causal relationship between insulin resistance and hyperfibrinogenemia. PMID- 10856529 TI - Correlations between measures of atherosclerosis change using carotid ultrasonography and coronary angiography. AB - Few studies have examined the correlation between change in carotid artery intima media thickness (IMT) and change in coronary artery disease. In the Cholesterol Lowering Atherosclerosis Study, current nonsmoking men with coronary artery disease were randomized to colestipol-niacin or placebo. Among 133 subjects with baseline and on-trial coronary angiography and carotid ultrasonography, colestipol-niacin treatment significantly reduced progression of atherosclerosis by both end point measures (2-year average change in percent diameter stenosis by coronary angiography and rate of change in carotid IMT). Significant correlations between change in common carotid artery IMT and quantitative coronary angiographic measures of change were evident over all coronary artery lesions, and in mild/moderate (<50% diameter stenosis), but not severe (>/=50% diameter stenosis) coronary artery lesions. In mild/moderate lesions, correlations with change in common carotid IMT were: percent diameter stenosis (r=0.28, P=0.002), minimum lumen diameter (r=-0.28, P=0.002), and vessel edge roughness (r=0.25, P=0.003). While measures obtained by carotid ultrasonography and coronary angiography are correlated, they each assess different aspects of atherosclerosis change. PMID- 10856530 TI - Comparison of various lipid, lipoprotein, and bilirubin combinations as risk factors for predicting coronary artery disease. AB - Studies were performed to determine if serum bilirubin, when combined with various lipid and lipoprotein risk factors, enhances our ability to predict coronary artery disease (CAD). This hypothesis was tested in a retrospective study of 644 middle-aged males who had undergone coronary angiography. The traditional risk factors of cholesterol, high density lipoprotein cholesterol (HDL-C), cholesterol/HDL-C ratios, triglycerides, age, cigarette smoking, and systolic blood pressure were tested by discriminant analysis, as were various cholesterol/bilirubin, cholesterol/(HDL-C+bilirubin), and low-density lipoprotein cholesterol (LDL-C)/(HDL-C+bilirubin) ratios. Each of these bilirubin-containing ratios was found to be an independent risk predictor when tested with the traditional risk factors. When the LDL-C/(HDL-C+bilirubin) ratio was included with the traditional risk predictors, it improved the prediction of severe CAD from 28.4 to 35.3% and the overall correct classification of CAD from 68.3 to 71.1%. When the 75th percentile was used as a cut-point, the diagnostic sensitivities obtained with cholesterol/(HDL-C+bilirubin) ratios (52.1%) and LDL C/(HDL-C+bilirubin) ratios (51.7%) were better than those obtained with cholesterol/HDL-C ratios (40.4%) (P=0.033 and 0.048, respectively). LDL-C/(HDL C+bilirubin) ratios also improved the prediction of severe CAD over those obtained with LDL-C/HDL-C ratios (43.4%); however, the changes were not statistically significant (P=0.096). If confirmed in other populations, serum bilirubin may be combined with LDL-C/HDL-C ratios, cholesterol/HDL-C ratios, cholesterol, or with various apolipoproteins to improve the prediction of CAD. PMID- 10856531 TI - Lipoprotein (a) and the risk of ischemic stroke in young women. AB - BACKGROUND AND PURPOSE: lipoprotein (a) (lp (a)) is a lipid-containing particle similar to LDL which has been found in atherosclerotic plaque. The role of lp (a) in ischemic stroke remains controversial, but some studies suggest lp (a) is particularly important as a risk factor for stroke in young adults. We investigated the role of lp (a) as a risk factor for stroke in young women enrolled in the Stroke Prevention in Young Women Study. METHODS: subjects were participants in a population-based, case-control study of risk factors for ischemic stroke in young women. Cases were derived from surveillance of 59 regional hospitals in the central Maryland, Washington DC, Pennsylvania and Delaware area. Lp (a) was measured in 110 cases and 216 age-matched controls. Demographics, risk factors, and stroke subtype were determined by interview and review of medical records. RESULTS: lp (a) values were higher in blacks than whites, but within racial groups, the distribution of lp (a) values was similar between cases and controls. After adjustment for age, race, hypertension, diabetes, cigarette smoking, coronary artery disease, total cholesterol and HDL cholesterol, the odds ratio for an association of lp (a) and stroke was 1.36 (95% CI 0.80-2.29). There was no dose-response relationship between lp (a) quintile and stroke risk. Among stroke subtypes, only lacunar stroke patients had significantly elevated lp (a) values compared to controls. CONCLUSIONS: we found no association of lp (a) with stroke in a population of young women with ischemic stroke. Small numbers of patients limit conclusions regarding risk in ischemic stroke subtypes, but we could not confirm previous suggestions of an association of lp (a) with atherosclerotic stroke in young adults. PMID- 10856532 TI - Intima-media thickness of peripheral arteries in asymptomatic cigarette smokers. AB - BACKGROUND AND PURPOSE: Although it is known that smoking is associated with an increase in arterial wall thickness, most studies have been performed in heterogeneous groups of older age, already suffering from atherosclerotic diseases or having additional cardiovascular risk factors. The purpose of this study is to assess the effect on arterial wall thickness of the carotid and femoral artery in cigarette smokers. METHODS: In a cross-sectional study, intima media thickness of the common and internal carotid artery, carotid bulb and common femoral artery was determined with the use of a B-mode ultrasound device, in 184 (44.3+/-9.0 years) cigarette smokers for whom smoking is the single cardiovascular risk factor. Comparisons were made with 56 non-smokers, matching in age and gender. RESULTS: The posterior walls of both carotid bulbs (right: P=0.0005; left: P=0.02) and of the internal carotid arteries (right: P=0.004; left: P=0.003) as well as the posterior wall of the right common carotid artery (P=0.02) and of the right common femoral artery (P<0.0001) were thicker in smokers. CONCLUSIONS: Cigarette smoking as the single cardiovascular risk factor causes wall thickening of the carotid and femoral arteries, which indicates that early atherosclerosis is already present in smokers entering middle age. PMID- 10856533 TI - Relationship between classic risk factors, plasma antioxidants and indicators of oxidant stress in angina pectoris (AP) in Tehran. AB - Cardiovascular disease (CVD) in general seems to be the leading cause of death in the Eastern Mediterranean Region (EMR) including Iran. This may be due to classic risk factors such as high triglyceride (TG), high total cholesterol (TC), and low levels of high density lipoprotein cholesterol (HDL-C). The impact of antioxidants as potentially protective risk factors against early coronary heart disease (CHD) is unknown in Iran. Therefore, relationships between angina and plasma antioxidants and indicators of lipid peroxidation were investigated in a case-control study. In this study, 82 cases of previously undiagnosed angina pectoris (AP), identified by a modified WHO Rose chest pain questionnaire and verified by electrocardiography during treadmill exercise testing, were compared with 146 controls selected from the same population of over 4000 male civil servants aged 40-60 years. Subjects with AP declared significantly less physical activity and had higher serum TG [means (S.E.M.) 2.32 (0.18) versus 1.61 (0.07) mmol/l] but lower HDL-C [1.01 (0.04) versus 1.18 (0.03) mmol/l] than age-matched controls. Levels of total serum cholesterol, low-density lipoprotein cholesterol (LDL-C) and lipoprotein(a) [Lp(a)] were not significantly different between the two groups, while the ratio of LDL-C/HDL-C was significantly higher [4.51 (0.23) versus 3.54 (0. 11)] for subjects with AP than for the controls. There was no significant difference in plasma levels of alpha-tocopherol, vitamin C, alpha- and beta-carotene. However, retinol [1.90 (0.06) versus 2. 09 (0.05)] and beta cryptoxanthin [0.398 (0.04) versus 0.467 (0.03)] were significantly lower in AP. Furthermore, angina cases exhibited a higher index of lipid peroxidation than controls (e.g. malondialdehyde, MDA; 0.376 (0.010) versus 0.337 (0.009) micromol/l). On multiple logistic regression analysis, retinol with odds ratio (OR) of 0.644 [95% confidence interval (CI; 0.425-0.978)], beta-cryptoxanthin, with an OR of 0.675 (CI; 0.487-0.940), oxidation indices, MDA with OR of 1.612 (95% CI; 1.119-2.322) and LDL-C/HDL-C ratio with OR of 2.006 (95% CI; 1.416 2.849) showed the most significant independent associations with AP in this group of Iranians. In conclusion, the state of lipid peroxidation as well as the status of special antioxidants may be co-determinants of AP in Iran, in parallel with the influence of classical risk factors for cardiovascular disease. PMID- 10856534 TI - Lipoprotein-associated phospholipase A(2), platelet-activating factor acetylhydrolase: a potential new risk factor for coronary artery disease. AB - A specific and robust immunoassay for the lipoprotein-associated phospholipase A(2) (Lp-PLA(2)), platelet-activating factor acetylhydrolase, is described for the first time. The immunoassay was used to evaluate possible links between plasma Lp-PLA(2) levels and atherosclerosis risk amongst susceptible individuals. Such an investigation was important because Lp-PLA(2) participates in the oxidative modification of low density lipoprotein by cleaving oxidised phosphatidylcholines, generating lysophosphatidylcholine and oxidised free fatty acids. The majority of Lp-PLA(2) was found associated with LDL (approximately 80%) and, as expected, enzyme levels were significantly positively correlated to LDL cholesterol. Plasma Lp-PLA(2) levels were significantly elevated in patients with angiographically proven coronary artery disease (CAD) when compared with age matched controls, even though LDL cholesterol levels did not differ significantly. Indeed, when included in a general linear model with LDL cholesterol and other risk factors, Lp-PLA(2) appeared to be an independent predictor of disease status. We propose, therefore, that plasma Lp-PLA(2) mass should be viewed as a potential novel risk factor for CAD that provides information related to but additional to traditional lipoprotein measurements. PMID- 10856535 TI - Inhibition of cholesterol synthesis by atorvastatin in homozygous familial hypercholesterolaemia. AB - Patients with homozygous familial hypercholesterolaemia (HoFH) have markedly elevated low density lipoprotein (LDL) cholesterol levels that are refractory to standard doses of lipid-lowering drug therapy. In the present study we evaluated the effect of atorvastatin on steady state concentrations of plasma lipids and mevalonic acid (MVA), as well as on 24-h urinary excretion of MVA in patients with well characterized HoFH. Thirty-five HoFH patients (18 males; 17 females) received 40 mg and then 80 mg atorvastatin/day. The dose of atorvastatin was increased further to 120 mg/day in 20 subjects and to 160 mg/day in 13 subjects who had not achieved LDL cholesterol goal, or in whom the dose of atorvastatin had not exceeded 2.5 mg/kg body wt per day. LDL cholesterol levels were reduced by 17% at the 40 mg/day and by 28% at the 80 mg/day dosage (P<0.01). Reduction in LDL cholesterol in the five receptor negative patients was similar to that achieved in the 30 patients with residual LDL receptor activity. Plasma MVA and 24-h urinary excretion of MVA, as markers of in vivo cholesterol synthesis, were elevated at baseline and decreased markedly with treatment. Urinary MVA excretion decreased by 57% at the 40 mg/day dose and by 63% at the 80 mg/day dosage (P<0. 01). There was a correlation between reduction in LDL cholesterol and reduction in urinary MVA excretion; those patients with the highest basal levels of MVA excretion and thus the highest rates of cholesterol synthesis having the greatest reduction in LDL cholesterol (r=0.38; P=0.02). Increasing the dose of atorvastatin to 120 and 160 mg/day did not result in any further reduction in LDL cholesterol or urinary MVA excretion suggesting a plateau effect with no further inhibition of cholesterol synthesis at doses of atorvastatin greater than 80 mg/day. PMID- 10856536 TI - Efficacy and safety of a combination of fluvastatin and bezafibrate in patients with mixed hyperlipidaemia (FACT study). AB - Preliminary data suggest that fluvastatin may be safely combined with fibrates. The Fluvastatin Alone and in Combination Treatment Study examined the effects on plasma lipids and safety of a combination of fluvastatin and bezafibrate in patients with coronary artery disease and mixed hyperlipidaemia. A total of 333 patients were randomly allocated in this multicentre double-blind trial to receive 40 mg fluvastatin alone (n=80), 400 mg bezafibrate (n=86), 20 mg fluvastatin+400 mg bezafibrate (n=85) or 40 mg fluvastatin+400 mg bezafibrate (n=82) for 24 weeks. Low-density lipoprotein (LDL)-cholesterol decreased >20% in all fluvastatin-containing regimens, with significantly greater decreases compared with bezafibrate alone (P<0.001). Bezafibrate alone and fluvastatin+bezafibrate combinations resulted in greater increases in high density lipoprotein (HDL)-cholesterol and decreases in triglycerides compared with fluvastatin alone (P<0.001). Fluvastatin (40 mg)+bezafibrate was the most effective for all lipid parameters with a decrease from baseline at endpoint in LDL-cholesterol of 24%, a decrease in triglycerides of 38% and an increase in HDL cholesterol of 22%. All treatments were well tolerated with no increase in adverse events for combination therapy versus monotherapy, or between combination regimens. No clinically relevant liver (aspartate aminotransferase [ASAT] or alanine aminotransferase [ALAT]) greater than three times the upper limit of normal) or muscular (creatine phosphokinase (CPK) greater than four times the upper limit of normal) laboratory abnormalities were reported. This large study shows 40 mg fluvastatin in combination with 400 mg bezafibrate to be highly effective and superior to either drug given as monotherapy in mixed hyperlipidaemia, and to be safe and well tolerated. PMID- 10856538 TI - Introduction PMID- 10856539 TI - Laparoscopic donor nephrectomy. PMID- 10856540 TI - Kidney graft loss after laparoscopic live donor nephrectomy. PMID- 10856541 TI - Real time monitoring of intraoperative allograft vitality. PMID- 10856542 TI - Thrombotic microangiopathy early after kidney transplantation: hemolytic uremic syndrome or vascular rejection? PMID- 10856543 TI - Successful interspousal swap of ABO incompatible living donor kidneys. PMID- 10856544 TI - Peripheral microchimerism in living donor kidney transplantation-correlation with posttransplantation course and long-term graft survival. PMID- 10856545 TI - Lack of correlation between HLA match and long-term survival in living donor kidney transplantation. PMID- 10856546 TI - Effect of immunosuppressive therapy on DNA repair and cancer incidence in renal transplant recipients. PMID- 10856547 TI - Speech disturbances in a child after living related liver transplant. PMID- 10856548 TI - Plasma zinc and copper concentrations after orthotopic liver transplantation. PMID- 10856549 TI - Coronary vein "steal and portal vein thrombosis after orthotopic liver transplantation. PMID- 10856550 TI - Liver transplantation in children: experience at Schneider's Children's Medical Center, Israel. PMID- 10856551 TI - Monitoring of Epstein-Barr virus serology in children after liver transplant: lack of clinical correlation. PMID- 10856552 TI - Portosystemic shunt for postorthotopic liver transplantation portal hypertension. PMID- 10856553 TI - Conversion of liver allograft recipients from cyclosporine A to FK 506 immunosuppressive therapy. PMID- 10856554 TI - The course of hepatitis B virus after liver transplantation. PMID- 10856555 TI - Safe vaccination against hepatitis B virus and discontinuation of hepatitis B immune globulin treatment in a liver transplanted patient. PMID- 10856556 TI - Combination of interferon-alpha and ribavirin therapy for recurrent hepatitis C virus infection after liver transplantation. PMID- 10856557 TI - Skin granulomas after liver transplantation: unusual presentation of recurrent primary biliary cirrhosis. PMID- 10856558 TI - Alveolar bone height in patients after liver transplantation. PMID- 10856559 TI - Detection of transplanted liver cells to the spleen by semiquantitative analysis using PCR for the Sry region on the Y chromosome. PMID- 10856560 TI - Cardiothoracic ratio: important prognostic tool in heart failure patients who are candidates for heart transplantation. PMID- 10856561 TI - Role of cyclosporine in inducing hyperuricemia in heart transplant patients. PMID- 10856562 TI - Functional status and quality of life of heart transplant recipients surviving beyond 5 years. PMID- 10856563 TI - The sky is the limit: exercise capacity 10 years post-heart-lung transplantation. PMID- 10856564 TI - Natural history of left-sided valves after heart transplantation. PMID- 10856565 TI - Immunosuppression induction with rabbit antithymocyte globulin in heart transplantation patients: 6-year experience at the Sheba Medical Center. PMID- 10856566 TI - Long-term physical training in cardiac transplant candidates: is it feasible? PMID- 10856567 TI - Correlation between maximal exercise capacity and right-ventricular function in candidates for heart transplantation. PMID- 10856568 TI - Extracorporeal photopheresis induces lymphocyte but not monocyte apoptosis. PMID- 10856570 TI - Four year experience with brain-death determinations. PMID- 10856569 TI - Effect of peripheral immune tolerance on liver-associated lymphocytes expressing NK1.1. PMID- 10856571 TI - Rapid en bloc liver-pancreas-kidney procurement: more pancreas with better vascular supply. PMID- 10856572 TI - Attitudes of negev beduins toward organ donation: a field survey. PMID- 10856573 TI - The organ donation process-workshop. PMID- 10856574 TI - Psychosocial model for evaluation and intervention with candidates for organ transplantation. PMID- 10856575 TI - The role of urology in transplantation and of transplantation in urology. PMID- 10856576 TI - Nonimmunologic renal allograft injury and delayed graft function: clinical strategies for prevention and treatment. PMID- 10856577 TI - Different graft outcome of paired kidneys from same donors-why? PMID- 10856578 TI - Successful long-term outcome utilizing existing native cutaneous ureterostomy for renal transplant drainage without ipsilateral native nephrectomy. PMID- 10856579 TI - Effect of different anti-rejection regimens on the expression of differentiation and activation markers on the surface of host lymphocytes. PMID- 10856580 TI - Extraperitoneal access for kidney transplantation in children weighing less than 20 KG. PMID- 10856581 TI - Complications of laparoscopic live donor nephrectomy: the first 175 cases. PMID- 10856582 TI - Morbidity of flank incision for renal donors. PMID- 10856583 TI - Significance of serum creatinine pattern and area under the creatinine versus time curve during the first acute renal transplant rejection. PMID- 10856584 TI - Histologic outcome of acute cellular rejection in kidney transplantation after treatment with methylprednisolone. PMID- 10856585 TI - A 3-hour postdose cyclosporine level during the first week after kidney transplantation predicts acute rejection and cyclosporine nephrotoxicity more accurately than trough levels. PMID- 10856586 TI - Gasless laparoscopy-assisted live donor nephrectomy: the initial 23 cases. PMID- 10856587 TI - Daclizumab in live donor renal transplantation. PMID- 10856588 TI - Intragraft expression of chemokine gene occurs early during acute rejection of allogeneic cardiac grafts. PMID- 10856589 TI - RANTES is produced by CD8+ T cells during acute rejection of skin grafts. PMID- 10856590 TI - Synergy of mycophenolate mofetil and bioflavonoids in prevention of immune and ischemic injury. PMID- 10856591 TI - Kidney splitting in miniature swine: a new animal model in renal transplantation. PMID- 10856592 TI - Urinary excretion of endothelin is elevated during hypothermic perfusion preservation in kidneys subjected to preretrieval warm ischemic injury. PMID- 10856593 TI - Oxidant stress in cadaveric and living kidney donors as markers of renal injury: utility of total antioxidant capacity and isoprostane levels in urine. PMID- 10856594 TI - Comparison of preservation solution RPS-96 with university of wisconsin solution in rat renal ischemia-reperfusion injury. PMID- 10856595 TI - Effect of erythromycin on ischemia-reperfusion injury of liver in rats. PMID- 10856596 TI - Effect of aminophylline on urine flow in children with tacrolimus-induced renal insufficiency. PMID- 10856597 TI - Small-volume resuscitation improves indocyanine green kinetics in patients with liver graft dysfunction. PMID- 10856598 TI - Improved post-cryopreservation recovery following encapsulation of islets in chitosan-alginate microcapsules. PMID- 10856600 TI - Distribution and function of prostanoid receptors: studies from knockout mice. AB - Recent developments in the molecular biology of the prostanoid receptors has allowed the investigation of the physiological roles of each individual receptor type and subtype. The following article reports the prostanoid receptor distributions deduced from Northern blot and in situ hybridization analyses, summarizes the phenotypes of each receptor knockout mice, and discusses recent studies investigating the effects of each receptor deficiency on the inflammatory response and female reproductive processes. The combination of expression pattern and knockout analyses enabled us to determine which receptor expressed in a particular cell is important for the maintenance of normal and/or pathological physiology. The results from these analyses may be useful in the development of novel therapeutics that can selectively manipulate prostanoid-mediated actions. PMID- 10856601 TI - Lipid metabolism in vertebrate retinal rod outer segments. PMID- 10856602 TI - Effects of xenoestrogen treatment on zona radiata protein and vitellogenin expression in Atlantic salmon (Salmo salar). AB - Zonagenesis (zona radiata protein synthesis) and vitellogenesis (yolk protein synthesis) are two reproductive responses that are integral aspects of fish oogenesis. This study examines the responses of eggshell zona radiata proteins (Zrp) and vitellogenin (Vtg) to five environmental pollutants; 4-nonylphenol (NP) and o,p'-DDT [both at 25 mg/kg], lindane (gamma-HCH) [10 mg/kg], a technical PCB mixture (Aroclor 1254; A1254) and bisphenol A (BPA) [both at 5 mg/kg] in juvenile Atlantic salmon (Salmo salar). Fish were given intraperitoneal injections of o,p' DDT, gamma-HCH, A1254 or BPA; singly, in combination with NP, and as a cocktail of all five chemicals, and later compared to NP-treated and untreated fish. In a separate experiment, fish were exposed to BPA in a dose-response manner (1, 5, 25 and 125 mg/kg fish). Based on previous studies, blood and liver samples were collected 2 weeks after injection. Zrp and Vtg levels were analyzed in plasma using immunoblotting and enzyme-linked immunosorbent assay. Liver cytochrome P4501A was analyzed by 7-ethoxyresorufin O-deethylase (EROD) activity. NP caused pronounced elevations in plasma Zrp and Vtg levels. In comparison, when NP was given in combination with gamma-HCH, Vtg levels was significantly reduced, compared to NP treatment alone. Using o,p'-DDT, A1254 and BPA, significant elevations of plasma Zrp and Vtg were seen when chemicals were given in combination with NP, but not when administered by themselves. An apparent dose response induction of Vtg and Zrp levels were observed in BPA treated juvenile salmon. In immunoblots, one component of molecular weight approximating the Zrp beta was detected when either o,p'-DDT, gamma-HCH, A1254 or BPA were given singly. A1254 significantly induced hepatic EROD activity when administered alone. However, when given as a mixture with all the other xenobiotics, reduction of EROD activity was observed. The data suggest a pattern of xenobiotics action which may complicate assessment of their reproductive effects. Zrp (the beta monomer) was more responsive to the xenoestrogens than Vtg, and provides a sensitive means of detecting exposure to environmental estrogens. PMID- 10856603 TI - Paraquat induced embryotoxicity on Xenopus laevis development. AB - Paraquat (PQ, 1-1'-dimethyl-4,4'-bipyridylium dichloride) is an effective herbicide widely used in agriculture with a rate of application for aquatic weed control ranging from 0.1 to 2 parts per million. Considering its wide-spread presence in Italian wetlands, we studied its embryotoxic effects with the Frog Embryo Teratogenesis Assay-Xenopus (FETAX). The percentage of mortality as well as the percentage of malformed larvae was investigated by probit analysis. The results showed that PQ was highly embryolethal. From a LC(50) of 0.138 mg/l and TC(50) of 0.267 mg/l, a TI(50) of 0.52 was derived; indicating that PQ is to be considered a non-teratogenic compound. Remarkable was the presence of a specific malformation, classified as ventral tail flexure, in the 29% of living larvae exposed to 0.125 mg/l PQ concentration. Their histological examination showed several zones of abnormal somites containing severely affected myocytes. This confirmed the molecular mechanism of PQ toxicity in cell microfilaments. Even at the lowest concentration of 0.0625 mg/l, the difference between the mean head tail length of control and exposed larvae was statistically significant, a sign of growth retardation. All our data emphasize that PQ must be consider highly embryotoxic on amphibian development, and suggest that this herbicide should be strictly regulated in weed control programs. PMID- 10856604 TI - In vitro effect of gossypol acetate on yellow perch (Perca flavescens) spermatozoa. AB - Gossypol, a known antispermatogenic agent from the cotton plant genus Gossypium, was found to inhibit yellow perch sperm motility in vitro and lactate dehydrogenase activity in spermatozoa when used in a dose-dependent manner. This toxin also significantly decreased sperm fertilizing ability. Exposure of sperm suspension to a 200-uM concentration of gossypol caused sperm immobilization and a consequent lack of fertilization. Effects of gossypol on sperm motility and fertilizing ability were partially reversible when sperm suspensions were transferred to solutions without gossypol. This reversibility was proportional to the gossypol concentration. We observed a significant increase of abnormal embryos produced by the fertilization of spermatozoa pretreated with gossypol. This implies that gossypol could affect the sperm genome. The results indicate a usefulness for yellow perch sperm in studies of gossypol action mechanisms. Results of in vitro experiments suggest a potential for reproductive impairment in fish when using cottonseed-containing diets or organic fertilizers in yellow perch aquaculture. However, these observations should be confirmed in in vivo experiments with yellow perch and other species. PMID- 10856605 TI - Toxicity of the cyanobacterial cyclic heptapeptide toxins microcystin-LR and -RR in early life-stages of the African clawed frog (Xenopus laevis). AB - Numerous cyanobacterial species are capable of producing potent toxins, which have been known to cause intoxications and fatalities in wildlife, livestock and humans. Microcystis is amongst the most ubiquitously distributed blue-green algal genus and almost invariably produces cyclic heptapeptide toxins called microcystins (MC). These toxins are highly persistent in water (several weeks). Highest concentrations are found in shallow littoral areas, the primary environment for aquatic early life-stage development. Therefore, the present study focussed on the potential embryotoxic effects of MC (MC-LR and -RR) in early life-stages of the amphibian Xenopus laevis. The endpoints chosen were mortality, malformation and growth inhibition. To achieve an improved dose response relationship the uptake of MC was quantified simultaneously, using a radiolabeled derivative of MC-LR. As one of the best described molecular mechanisms of MC toxicity involves the specific inhibition of serine/threonine protein phosphatases-1 and -2A (PP), essential enzymes involved in the mechanisms of cell cycle regulation and maintenance of cellular morphology, the inhibition of PP in X. laevis exposed to MC was monitored. For this the presence of both PP 1 and PP-2A was confirmed by means of SDS-PAGE and immunoblotting. Second, the capacity of MC to inhibit X. laevis embryo-larval PP was corroborated by in vitro incubation of embryo-larval homogenates with MC-LR and -RR and subsequent determination of PP-inhibition. No increased mortality, malformation, or growth inhibition was observed even at the highest MC concentrations employed. MC had neither a demonstrable inhibitory effect on X. laevis PP-activity in vivo in the first 96 h of exposure. However, as of 96 and 120 h exposure a significant inhibition of PP activity was observed at the highest dose (2000 ug/l) in MC-LR and MC-RR exposed embryo-larvae, respectively. By the same token, no notable amounts of radiolabeled [3H]-MC-LR were taken up during the first 96 h, whereas a drastic increase in [3H]-MC-LR was observed after feeding of the larvae had commenced. The [3H]-MC-LR concentration was consistently found to be highest in the viscerothoracal sections of the larvae (2112+/-429 ug MC/kg dry weight after 120 h). The present findings indicate that transchorional/transdermal absorption of MC in X. laevis is minimal or absent and that oral uptake of MC with ambient water is necessary for the development of MC related toxicity. Furthermore, the comparison of the MC doses used in this study with the concentrations reported in surface waters indicate that early life-stages of amphibians (up to 5 days of development) are unlikely to be affected by cyanobacterial blooms producing MC-LR and -RR. PMID- 10856606 TI - Fate of sediment-associated pyrene and benzo AB - Bioaccumulation, depuration and biotransformation of radiolabelled pyrene and benzo[a]pyrene were studied in the freshwater oligochaete Lumbriculus variegatus in spiked Lake Mekrijarvi (Eastern Finland) sediment in two sets of experiments (I and II). In experiment I bioaccumulation, depuration and biotransformation of PAHs were studied. In experiment II biotransformation was investigated further using three different solvent extractions and the ability of L. variegatus to biotransform was based on the change of proportion of the parent PAH in tissue. Bioaccumulation of both chemicals was fast and an apparent steady level was reached within a week. Biotransformation results obtained by solvent extractions were in agreement with each other, although hexane appeared to be less efficient solvent for B[a]P than chloroform:methanol or ethyl acetate:acetone/cyclohexane. The relative proportion of parent PAHs in tissues decreased continously during the 504 and the 336 h exposures in experiments I and II, respectively. After 336 h exposure in experiment II, approximately 60% of pyrene and 90% of B[a]P associated radioactivity was still in the parent compound. Depuration of the parent compounds in clean sediment was fast. Most of the parent chemicals were depurated within 72 h but the possible biotransformation products remained mainly in tissues. Feeding behavior of the animals (sediment ingesting vs. noningesting) did not affect pyrene biotransformation but the proportion of parent B[a]P in tissues was larger in feeding animals. This was probably due to faster uptake rate of B[a]P to feeders than nonfeeders and slow biotransformation rate of the chemical. Our results suggest that biotransformation of pyrene and B[a]P in L. variegatus is likely and it should be taken into account when studying bioaccumulation of PAHs in assessing hazard of sediment contamination. PMID- 10856607 TI - Combined effects of pentachlorophenol and salinity stress on phagocytic and chemotactic function in two species of abalone. AB - The effect of pentachlorophenol (PCP) combined with salinity stress on specific aspects of cellular immunity was examined in two species of abalone. Chemotactic and phagocytic ability (percent phagocytosis, %P; and phagocytic index, PI), as well as gross morphological characteristics, of hemocytes withdrawn from red (Haliotis rufescens) and black abalone (H. cracherodii) after in vivo exposure to 25, 35, or 45 per thousand seawater salinity plus 1.2 mg/l PCP were determined. Abalone exposures of 3.5 and 6.5 h, respectively, were based on species-specific metabolic endpoints (MEPs) derived from previous NMR data. Hemocyte chemotaxis was found to be augmented by low salinity in red but not black abalone, while high salinity seemed to reduce chemotactic ability in both species. PCP did not potentiate the effects demonstrated by salinity variations alone in red abalone, although chemotaxis was further compromised in black abalone exposed to high salinity stress combined with PCP. Although chemotactic ability was similar among both species, both %P and PI was greater among black abalone. Low salinity may offset the effects of PCP in red but not black abalone. Whereas red abalone demonstrated a reduction in phagocytic ability upon exposure to high salinity alone, black abalone appeared more resilient to high salinity stress alone. Furthermore, while the effect of high salinity plus PCP is subadditive among red abalone, high salinity seems to potentiate the effect of PCP on black abalone. Finally, hemocytes exposed to acute salinity variations plus PCP showed varying morphological characteristics when compared to controls. Over 30% of hemocytes of high salinity plus PCP exposed abalone remained rounded and generally unattached to glass slides with only a few pseudopod extensions. Hemocyte preparations incubated in rhodamine-conjugated phalloidin were observed with fluorescence microscopy and reveal an altered actin filament pattern. Overall, black abalone demonstrate greater phagocytic ability than red abalone both in the presence and absence of PCP. Furthermore, increases in salinity are accompanied with reduced phagocytic ability with red abalone demonstrating particular sensitivity to salinity variations. While PCP inhibits ATP synthesis, salinity variations may impose an additional stress on intertidal animals exposed to transient or seasonal environmental changes. PMID- 10856608 TI - Current concepts in neuroanatomical tracing. AB - The development of new axonal tract tracing and cell labelling methods has revolutionised neurobiology in the last 30 years. The aim of this review is to consider some of the key methods of neuroanatomical tracing that are currently in use and have proved invaluable in charting the complex interconnections of the central nervous system. The review begins with a short overview of the most frequently used tracers, including enzymes, peptides, biocytin, latex beads, plant lectins and the ever-increasing number of fluorescent dyes. This is followed by a more detailed consideration of both well established and more recently introduced neuroanatomical tracing methods. Technical aspects of the application, uptake mechanisms, intracellular transport of tracers, and the problems of subsequent signal detection, are also discussed. The methods that are presented and discussed in detail include: (1) anterograde and retrograde neuroanatomical labelling with fluorescent dyes in vivo, (2) labelling of post mortem tissue, (3) developmental studies, (4) transcellular tracing (phagocytosis dependent staining of glial cells), (5) electrophysiological mapping combined with neuronal tract tracing, and (6) simultaneous detection of more than one axonal tracer. (7) Versatile protocols for three-colour labelling have been developed to study complex patterns of connections. It is envisaged that this review will be used to guide the readers in their selection of the most appropriate techniques to apply to their own particular area of interest. PMID- 10856609 TI - Elucidation of the neurobiology of depression: insights from a novel genetic animal model. AB - Development of drugs for the effective treatment of depressive disorders requires elucidation of factors that are critical for clinically antidepressant effects. During the past 4 years, we have studied in situ neurochemical alterations in the brain that may underlie depressive behavior. This was achieved using the genetically-selected Flinders Sensitive Line (FSL) of rats (a unique animal model of depression), before and after chronic antidepressant treatment. This line of rats exhibits behavioral features characteristic of depression, and responds to chronic, but not acute, antidepressant treatments. This review summarizes our findings concerning the local neuro-dynamics in the brain during manifestation of depressive behavior and effective antidepressant treatment in this animal model of depression. Understanding the abnormalities manifested in neurochemical pathways during depressive disorders and the dynamic effects of these abnormalities on the onset of action and efficacy of pharmacological treatments are crucial for the development of effective antidepresssant drugs and therapeutic strategies. PMID- 10856610 TI - Why we sleep: the evolutionary pathway to the mammalian sleep. AB - The cause of sleep is a complex question, which needs first, a clear distinction amongst the different meanings of a causal relationship in the study of a given behavior, second, the requisites to be met by a suggested cause, and third, a precise definition of sleep to distinguish behavioral from polygraphic sleep. This review aims at clarifying the meaning of the question and at showing the phylogenetic origin of the mammalian and avian sleep. The phylogenetic appearance of sleep can be approached through a study of the evolution of the vertebrate brain. This began as an undifferentiated dorsal nerve, which was followed by the development of an anterior simplified brain and ended with the formation of the multilayered mammalian neocortex or the avian neostriate. The successive stages in the differentiation of the vertebrate brain produced, at least, two different waking types. The oldest one is the diurnal activity, bound to the light phase of the circadian cycle. Poikilotherms control the waking from the whole brainstem, where their main sensorymotor areas lie. Mammals developed the thalamocortical lines, which displaced the waking up to the cortex after acquiring homeothermy and nocturnal lifestyle. In order to avoid competence between duplicate systems, the early waking type, controlled from the brainstem, was suppressed, and by necessity was turned into inactivity, probably slow wave sleep. On the other hand, the nocturnal rest of poikilotherms most probably resulted in rapid eye movement sleep. The complex structure of the mammalian sleep should thus be considered an evolutionary remnant; the true acquisition of mammals is the cortical waking and not the sleep. PMID- 10856611 TI - Modulation of taurine release by glutamate receptors and nitric oxide. AB - Taurine is held to function as a modulator and osmoregulator in the central nervous system, being of particular importance in the immature brain. In view of the possible involvement of excitatory pathways in the regulation of taurine function in the brain, the interference of glutamate receptors with taurine release from different tissue preparations in vitro and from the brain in vivo is of special interest. The release of taurine from the brain is enhanced by glutamate receptor agonists. This enhancement is inhibited by the respective receptor antagonists both in vitro and in vivo. The ionotropic N-methyl-D aspartate (NMDA) and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor agonists appear to be the most effective in enhancing taurine release, their effects being receptor-mediated. Kainate is less effective, particularly in adults. Of the glutamate receptors, the NMDA class seems to be the most susceptible to modulation by nitric oxide. Nitric oxide also modulates taurine release, enhancing the basal release in both immature and mature hippocampus, whereas the K(+)-stimulated release is generally inhibited. Metabotropic glutamate receptors also participate in the regulation of taurine release, group I metabotropic glutamate receptors potentiating the release in the developing hippocampus, while group III receptors may be involved in the adult. Under various cell-damaging conditions, including ischemia, hypoxia and hypoglycemia, taurine release is enhanced, together with an enhanced release of excitatory amino acids. The increase in extracellular taurine upon excessive stimulation of glutamate receptors and under cell-damaging conditions may serve as an important protective mechanism against excitotoxicity, being particularly effective in the immature brain. PMID- 10856612 TI - Calcium channels and the release of large dense core vesicles from neuroendocrine cells: spatial organization and functional coupling. AB - The release of large dense core vesicles (LDCV) by neuroendocrine cells displays a very similar calcium dependence as found in synapses, yet, the organization of channels and vesicles is quite different. Various biophysical properties of the release process, notably a large delay (>10 ms) between excitation and release and a high impact of mobile calcium buffers, suggest that, generally, vesicles and channels do not co-localize as in synapses, but are separated by a distance of 100-300 nm. This review focuses on the consequences of this organization for the functional coupling of calcium channels to LDCV-release in neuroendocrine cells. The large distance between LDCV and calcium channels in neuroendocrine cells obviates molecular interactions between channels and fusion peptides and implies that each type of calcium channel may be involved in release. Thus, preferential functional coupling of specific calcium channel types to the exocytotic process may be completely lacking, as in melanotropes. Alternatively, it may be present to some extent to induce differences in coupling efficacy between channel types, as in calf chromaffin cells and mouse pancreatic beta cells. Physiological mechanisms, like recruitment of channels through facilitation processes or suppression of channels through inactivation, may change coupling characteristics during activity. Due to the large distance between channels and vesicles, single action potentials (APs) are usually insufficient to elicit release, and the coupling between individual APs and release is loose. Most neuroendocrine cells are therefore seen to fire in bursts, like pancreatic beta-cells. Furthermore, a large variation in shape and duration of the APs, with APs of up to 300 ms as in melanotropes, acts as another mechanism to enhance stimulus secretion coupling. PMID- 10856613 TI - Intracorporeal imaging and differentiation of living tissue with an ultra-high frequency ultrasound probe. AB - Intraoperative diagnostic tissue differentiation is expected to be useful clinically. We have fabricated a 3-mm diameter rod-shaped ultrasound (US) probe mounted with a 120-MHz transducer whose lateral resolution is the same as the cellular size of 13 microm. The probe can image a microscopic structure (i.e., the cellular arrangement inside intracorporeal living tissue). We imaged normal kidney tissue of a living mouse and tumor tissue implanted in another mouse kidney. We anesthetized the mice, exteriorized the kidneys, and punctured the kidneys with the probe. Renal corpuscle-like structures were seen in the healthy kidney, but a wavy spindle-like structure was seen in the tumor. The similarity between the ultrasonic images and histological sections taken from the imaged organs demonstrates the possibility of real-time tissue differentiation by ultra high-frequency US. PMID- 10856614 TI - Three-dimensional ultrasound imaging of the rotator cuff: spatial compounding and tendon thickness measurement. AB - Three-dimensional (3-D) volume reconstructions of the shoulder rotator cuff were generated from freehand ultrasound (US) scans acquired with a magnetic tracking system. Image stacks acquired with lateral overlap from multiple acoustic windows were spatially compounded to provide an extended representation of the rotator cuff tendons. A semiautomated technique was developed for measuring rotator cuff thickness from the 3-D compound volumes. Scans of phantoms and volunteer subjects were used to evaluate the accuracy and repeatability of the thickness measurements. For an in vitro phantom with known thickness, the mean difference between the true value and the automatic measurements was 0.05 +/- 0.28 mm. Thickness measurements made manually from 2-D images and automatically from 3-D volumes were different by 0.03 +/- 0.44 mm in vitro and -0.06 +/- 0.36 in vivo. Repeated thickness measurements in vivo differed by 0.06 +/- 0.36 mm. The 3-D measurement technique offers a promising method for evaluating rotator cuff tendons. PMID- 10856615 TI - Three-dimensional automatic quantitative analysis of intravascular ultrasound images. AB - Intravascular ultrasound (IVUS) has established itself as a useful tool for coronary assessment. The vast amount of data obtained by a single IVUS study renders manual analysis impractical for clinical use. A computerized method is needed to accelerate the process and eliminate user-dependency. In this study, a new algorithm is used to identify the lumen border and the media-adventitia border (the external elastic membrane). Setting an initial surface on the IVUS catheter perimeter and using active contour principles, the surface inflates until virtual force equilibrium defined by the surface geometry and image features is reached. The method extracts these features in three dimensions (3 D). Eight IVUS procedures were performed using an automatic pullback device. Using the ECG signal for synchronization, sets of images covering the entire studied region and corresponding to the same cardiac phase were sampled. Lumen and media-adventitia border contours were traced manually and compared to the automatic results obtained by the suggested method. Linear regression results for vessel area enclosed by the lumen and media-adventitia border indicate high correlation between manual vs. automatic tracings (y = 1.07 x -0.38; r = 0.98; SD = 0.112 mm(2); n = 88). These results indicate that the suggested algorithm may potentially provide a clinical tool for accurate lumen and plaque assessment. PMID- 10856616 TI - Body-centered visualisation for freehand 3-D ultrasound. AB - Three-dimensional (3-D) ultrasound (US) data is typically visualised by any-plane slicing, volume rendering or surface rendering. Typical implementations of these techniques do not readily convey the spatial relationship between the visualised data and the patient's body, something that is particularly important when the data are reviewed after the scan has taken place, perhaps by a remote expert who did not even perform the scan. This paper describes a facility to register the 3 D US data to the patient's body and then display the data correctly superimposed on a rendered mannequin (rigid computer model). This way, the user can appreciate the position and orientation of any visualisation with respect to the patient's body. The facility relies on efficient implementation of progressive meshes to manage the level of detail of the mannequin model. PMID- 10856617 TI - Optimum scan spacing for three-dimensional ultrasound by speckle statistics. AB - A technique is introduced to determine the spacing between acquired two dimensional (2-D) planes necessary to maintain resolution in the scan direction while maximizing scan speed for 3-D ultrasound (US) imaging. We performed statistical analysis on a series of 3-D scans of agar blocks with different image spacings, quantifying speckle size (S(c)) as the full-width half-maximum (FWHM) of the autocovariance function. S(c) was approximately constant at small 2-D image-plane spacings, but increased after the scan spacing surpassed some optimum point. Optimum spacing ranged between 0.075 and 0.4 mm, increasing with the axial depth and the number of focal zones. This latter dependence is a result of successive sampling by the US machine and the digitizer at unequal rates, as demonstrated through an analysis of the noise power spectra. This analysis predicted that S(c) may be significantly reduced by using a transducer sampling rate that is twice the digitization rate. This technique permits the calculation of the most efficient scan spacings for 3-D US imaging. PMID- 10856618 TI - Evaluation of progression in nonrheumatic aortic valvular stenosis by scanning acoustic microscopy. AB - To investigate distributions of hardness and thickness in nonrheumatic aortic stenosis (AS), scanning acoustic microscopy was used. The acoustic propagation speed (APS: m/s) and thickness at three sites (tip, middle and base) of aortic valve were measured in 18 cusps from 7 surgical patients with AS (late lesion), 27 showing mild lesions from 9 autopsy cases (early lesion) and 18 healthy from 6 autopsy cases (healthy). These were measured in each layer of cusps: fibrosa (F), spongiosa (S) or ventricularis (V). In early lesions, an increase in APS preceded the thickening and distributed in the tip (1666 +/- 107), the three layers of the middle (F: 1782 +/- 121; S: 1590 +/- 38; V: 1636 +/- 59) and the fibrosa of the base (1736 +/- 203). In late lesions, APS of the tip and three layers of the base were markedly increased. Progressive nonrheumatic AS is characterized by increased hardness that precedes the thickening, and its distribution may be related to mechanical stress. PMID- 10856619 TI - A new approach for glomerular lesions: evaluation of scanning acoustic microscopy (SAM) for experimental glomerular disease in rats. AB - Most pathological evaluations using ordinary optical microscopy are usually qualitative and subjective. The beneficial properties of scanning acoustic microscopy (SAM) include not only observation of microstructure but also quantitative measurement of acoustic propagation speed, indicating the tissue elasticity. In this study, we examined the capability of SAM to evaluate pathological findings in glomeruli using anti-Thy.1 glomerulonephritis. Light microscopic observations of the glomeruli showed severe cell proliferation and mesangial matrix expansion at 10 days after induction of glomerulonephritis and, yet, to a lower extent at day 21 with onset of healing. C-mode scanning of SAM enabled imaging of glomerular structure compatible to findings of ordinary light microscopy. In addition, glomerular propagation speed in nephritic rats was significantly increased at day 10, and then decreased at day 21. These results indicate that SAM evaluation may be a new, useful technique for quantitative evaluation of proliferative glomerular lesions. PMID- 10856620 TI - Detection of stenoses in the anterior circulation using frequency-based transcranial color-coded sonography. AB - Atherosclerotic stenoses of the intracranial vessels are less frequent than those of the extracranial vessels, but they are associated with a considerable annual stroke rate. The aim of the present study was to investigate the usefulness of frequency-based transcranial color-coded sonography (TCCS), transcranial Doppler sonography (TCD) and digital subtraction angiography (DSA) in patients with middle cerebral artery (MCA) and intracranial internal carotid artery (ICA) stenosis. Forty patients presenting with 48 intracranial stenoses of the anterior circulation were involved in the study. The stenoses were detected in the neurovascular laboratory during routine TCD examinations. All patients underwent an additional frequency-based TCCS examination. Both the axial and coronal planes were obtained to allow the exact localization of MCA stenosis and differentiation from intracranial ICA stenosis. Angle-corrected flow velocity measurements were performed if straight vessel compartments were 20 mm or more in length. A total of 18 stenoses (44%) were investigated additionally with DSA. According to the investigation with TCD, 20 (42%) stenoses were low-grade, 12 (25%) were moderate, and the remaining 16 (33%) were severe. Angle-corrected flow velocity measurements obtained with the integrated pulse-wave Doppler device of the TCCS machine were highly correlated (0.912, p < 0.001) with those obtained with TCD. TCCS achieved a reliable differentiation of MCA main stem stenosis vs. intracranial ICA stenosis in 7 patients and vs. MCA branch stenosis in 4 patients, but TCD failed in these two subgroups. The agreement between DSA and TCCS to evaluate semiquantitatively 18 intracranial stenoses resulted in a weighted-kappa value of 0.764. The major clinically relevant advantages of TCCS over TCD in MCA stenosis are its ability to differentiate MCA trunk stenosis from terminal ICA or MCA branch stenosis reliably and to perform angle-corrected flow velocity measurements. PMID- 10856621 TI - Application of autoregressive methods to multigate spectral analysis. AB - Multigate analysis is known to be capable of detecting accurate blood velocity profiles from human vessels. Experimental systems so far presented in the literature use time-domain frequency estimations and, more recently, the fast Fourier transform (FFT) for real-time analysis of Doppler signals from multiple range cells. This experimental study is aimed at evaluating the application of an autoregressive (AR) method (Burg algorithm) to multigate Doppler analysis. Both in vitro and in vivo results were collected with a commercial Duplex scanner coupled with a prototype multigate unit developed in our laboratory. The same multigate signals are, thus, processed according to both the FFT and the Burg algorithms. The related spectral and maximum frequency profiles are reported and statistically compared. AR, implemented with the Burg algorithm, is demonstrated to be a way to perform multigate spectral analysis with reduced spectral variance, suitable for maximum velocity profile extraction through a simple threshold. PMID- 10856622 TI - Using doppler signal power to detect changes in vessel size: a feasibility study using a wall-less flow phantom. AB - The power of a Doppler signal is theoretically proportional to the volume of blood within the sample volume of an ultrasound (US) beam and, hence, may provide a means of detecting in vivo changes in the cross-sectional area of cerebral vessels, such as the middle cerebral artery. The purpose of this study was to examine the relationship between power and vessel size for signals recorded from a wall-less flow phantom. The results demonstrate the importance for the in vitro case of maximising the received signal power for each channel to obtain the true relationship between power and size, and show that a nonproportional relationship observed between the two parameters is primarily caused by high-pass filtering and nonuniform insonation. In addition, an investigation of the reproducibility of power values after transducer repositioning has shown that variation occurs even when extreme care is taken to maximise the received signal intensity. The implications of these results for the in vivo use of the Doppler signal power method are discussed. PMID- 10856623 TI - From digital image processing of colour Doppler M-mode maps to noninvasive evaluation of the left ventricular diastolic function: a dedicated software package. AB - Noninvasive estimation of diastolic pressure gradients has recently been validated using the space-temporal velocity distribution available from colour Doppler M-mode (CDM). However, the methods currently applied for analysing CDM patterns of left ventricular (LV) filling have limitations, such as lack of automation, subjective variability and limited use of digital velocity map. For this reason, we have developed software able to acquire and process the CDM maps; thus, providing an easily interpretable graphical and numerical display. The pressure field is obtained by approximating the derivatives with centred finite differences via the incompressible Navier-Stokes equations. After digital filtering of the noise and the removal of the colour black spots, the velocity field is utilised to compute the pressure gradient field and the pressure values by spatial integration. It is concluded that automatic quantification of colour CDM patterns is feasible and will be a strategic tool in the investigation of one of the most intriguing topics in cardiology. PMID- 10856624 TI - Delineation of the extracellular determinants of ultrasonic scattering from elastic arteries. AB - Elastic arteries consist of three primary components: elastin fibers, extracellular collagen matrix and smooth muscle cells. However, the relative contribution of elastin and collagen fibers to overall ultrasonic scattering from an intact arterial wall is poorly understood. To define the principal source of extracellular scattering from the medial layer of elastic arteries, canine ascending aortas (n = 10) were excised, fixed and sectioned for insonification. Subsequently, aortic specimens were restudied after treatment to dissolve all tissue components except extracellular collagen matrix (n = 5) and elastin fibers (n = 5). Histological staining revealed very few elastin fibers and sparse intact collagen in collagen-isolated and elastin-isolated tissues, respectively. Integrated backscatter, attenuation and backscatter coefficients differentiated these two treated tissues. The backscatter coefficient for elastin-isolated tissue demonstrated a fivefold increase over collagen-isolated tissue, suggesting that elastin fibers represent a primary scattering component within elastic arteries, and the collagen fibers may provide a secondary component of scattering. PMID- 10856625 TI - Influence of environmental conditions on a new surfactant-based contrast agent: ST68. AB - Environmental influences on the new surfactant-stabilized bubbles, ST68, were investigated. We have developed a new surfactant-based contrast agent ST68, which is prepared by insonating buffered mixtures of Span 60 and Tween 80 in the presence of either air, PFC, or SF(6) gas. The effect of dilution, shear, and sonication on size distribution of ST68 showed that PFC-containing bubbles (ST68 PFC) were most stable. ST68-PFC bubbles lasted more than 15 min with approximately 30 dB backscatter enhancement in degassed phosphate-buffered saline, (pH 7.4), and air bubbles lasted approximately 3 s, suggesting the effects of diffusion. Additionally, it was found that the ionic strength of the suspending medium (for example, PBS), did not have any effect on ST68 bubbles containing SF(6) or PFC, but had a dramatic impact on bubbles containing air. PMID- 10856626 TI - Quantification of microbubble destruction of three fluorocarbon-filled ultrasonic contrast agents. AB - The assessment of myocardial blood velocity using ultrasonic contrast agents is based on the premise that the vast majority of contrast microbubbles within a myocardial region can be destroyed by an acoustic pulse of sufficient magnitude. Determination of the period of time after destruction that a region of myocardium needs to reperfuse may be used to assess myocardial blood velocity. In this study, we investigated the acoustic pressure sensitivity of three solutions of intravenous fluorocarbon-filled contrast agents and the magnitude of acoustic pulse required to destroy the contrast agent microbubbles. A novel tissue mimicking phantom was designed and manufactured to investigate the relationships between mean integrated backscatter, incident acoustic pressure and number of frames of insonation for three fluorocarbon-filled contrast agents (Definity(R), Optison(R), and Sonazoid(R), formerly NC100100). Using a routine clinical ultrasound (US) scanner (Acuson XP-10), modified to allow access to the unprocessed US data, the contrast agents were scanned at the four acoustic output powers. All three agents initially demonstrated a linear relationship between mean integrated backscatter and number of frames of insonation. For all three agents, mean integrated backscatter decreased more rapidly at higher acoustic pressures, suggesting a more rapid destruction of the microbubbles. In spite of the fact that there was no movement of microbubbles into or out of the beam, only the results from Definity(R) suggested that a complete destruction of the contrast agent microbubbles had occurred within the total duration of insonation in this study. PMID- 10856627 TI - Measurements of phase velocity and group velocity in human calcaneus. AB - Ultrasonic velocity in calcaneus correlates highly with bone mineral density, which is a good predictor of osteoporotic fracture risk. Several commercial bone sonometers perform a velocity measurement based on the transit time of a broadband pulse to assess skeletal status. This approach is somewhat problematic, however, because several authors have reported ambiguities in measurements in calcaneus. Phase velocity is an alternative that may be less dependent on device spectral characteristics. In addition, dispersion (the frequency-dependence of phase velocity) is a fundamental property worth investigating to increase understanding of interaction between ultrasound and bone. To compare two group velocity measurement methods and one phase-velocity measurement method, a polycarbonate sample (for method validation) and 24 human calcanei were investigated in vitro. Phase velocity in calcaneus at 500 kHz was 1511 m/s +/- 30 m/s (mean +/- standard deviation). Average phase velocity decreased approximately linearly with frequency (-18 m/s MHz). The two group velocity measurements were comparable and tended to be slightly lower than phase velocity. The magnitude of dispersion showed little correlation with bone mineral density. PMID- 10856628 TI - The influence of variations in blood flow on pulsed doppler ultrasonic heating of the cerebral cortex of the neonatal pig. AB - Pulsed Doppler ultrasound examination of the fetal cerebral circulation may cause potentially harmful temperature elevations in brain tissue immediately beneath the insonated segment of the skull. This study measured the effect of variations in cerebral blood flow on ultrasonic heating of the cerebral cortex of anaesthetised, neonatal pigs. Wide and narrow ultrasound beams were used. Pulsed ultrasound exposures were delivered in 90 s bursts at 5.8 micros pulse length, pulse repetition frequency 8 kHz and centre frequency 3.5 MHz. Studies were performed with the target at the focus of a fixed, stationary beam of 0.3 cm -6 dB beam width (narrow beam) and I(spta) 1.4 W/cm(2) (n = 11), or with the target in the near field of a fixed, stationary beam of 1.6 cm -6 dB beam width (wide beam) and I(spta) 3.6 W/cm(2)(n = 5). The 90 s ultrasound exposures were performed under three different conditions of ambient cerebral blood flow: baseline (during normocarbic, normoxic conditions), increased (during hypercarbic, hypoxic conditions) and absent (postmortem). Cerebral blood flow was measured using the radiolabelled microsphere technique. In the narrow beam studies, cerebral blood flow during baseline was 34 +/- 4 ml/min/100 g, rising to 109 +/- 32 ml/min/100 g during the increased phase (p < 0.001); in the wide beam studies baseline flows were 29 +/- 9 ml/min/100 g, whereas flows in the increased phase were 128 +/- 32 ml/min/100 g (p < 0.001). There was no difference in the heating curves for normal, increased and absent cerebral blood flow for exposure to the narrow beam, when mean temperature increases of 1.5 degrees C at 90 s were recorded in each case (p > 0.21, power > 0.8). However, the heating curves for the wide beam were significantly different for the three rates of blood flow with mean temperature increases of 1.9 degrees C (normal flow), 1.7 degrees C (increased flow) and 2.4 degrees C (no flow) recorded at 90 s (p < 0.05). PMID- 10856629 TI - Ultrasonic wave action upon the red blood cell agglutination in vitro. AB - The ultrasonic wave action upon immune red blood cell (RBC) aggregation in vitro was investigated by means of the elastic light-scattering method. The RBC agglutination process in the ultrasonic wave presence was analysed as a function of irradiation time, the antiserum series, the whole blood and antiserum dilution degree and blood group types. It was experimentally shown that the ultrasonic irradiation accelerated the agglutination reaction in the blood and antiserum mixture. The sedimentation process showed a gain in the resolution of the optical method's ability to detect the induced RBC complex formation. It revealed that the ultrasonic application led to significant diagnostic test time shortening. The results may be used to develop new diagnostic techniques and devices. PMID- 10856630 TI - Sonoporation of monolayer cells by diagnostic ultrasound activation of contrast agent gas bodies. AB - Human (A431 epidermoid carcinoma) cells were grown as monolayers on 5 microm thick Mylar sheets, which formed the upper window for a 1-mm thick, 23-mm diameter disc-shaped exposure chamber. A 3.5-MHz curved linear-array transducer was aimed upward at the chamber, 7 cm away, in a 37 degrees C water bath. The chamber contained phosphate-buffered saline (PBS) with 10 mg/mL fluorescent dextran and 1% Optison ultrasound (US) contrast agent. Significant fluorescent cell counts, indicative of membrane damage (i.e., sonoporation), up to about 10% of cells within a 1-mm diameter field of view, were noted for spectral Doppler and two-dimensional (2-D) scan mode with or without a tissue-mimicking phantom. The effect was only weakly dependent on pulse-repetition frequency or exposure duration, but was strongly dependent on contrast agent concentration below 2%. Thus, diagnostic US activation of contrast-agent gas bodies can produce cell membrane damage. PMID- 10856631 TI - Development of an interstitial ultrasound applicator for endoscopic procedures: animal experimentation. AB - Biliary cancer is very difficult to treat, mainly because of the advanced stage at which such tumours are detected and the low efficacy of systemic therapeutic modalities like radiotherapy. Palliative measures designed to clear the duct (either by means of surgery or an endoscopic procedure) are presently performed. A relatively noninvasive alternative could be developed to fill this gap in the therapeutic arsenal. To this end, we have designed an interstitial ultrasound (US) applicator suitable for use with a digestive endoscope. This applicator is based on a water-cooled plane transducer that operates at 10 MHz. Although, because the target zone is cylindrical in shape, it might have seemed more logical to use a cylindrical transducer. Nevertheless, a plane transducer was chosen because the pressure field from this kind of emitter decreases less quickly, which means faster and deeper heating. However, to generate coagulation necrosis all around the duct, the applicator has to be rotated around its axis; this is achieved by means of a flexible metallic shaft (2 m in length and 3. 8 mm in diameter) that joins the device's active head (which contains the transducer) to the casing with all the connectors. A holder is fixed at the endoscope channel inlet; it controls the rotation of the applicator. Trials were conducted on pigs. The duodenoscope was introduced via the oesophagus down through the duodenum as far as the hepatopancreatic ampulla. Using a guide wire, the applicator was navigated into the duct via the endoscope instrument channel. Well defined, reproducible volumes of coagulation necrosis with a diameter of 20 mm were generated in the biliary tissue and the liver. These promising results indicate that this kind of endoscopic US delivery system may represent an effective tool for the treatment of biliary tumours in humans. An Independent Ethics Committee recently approved preliminary clinical trials of this applicator. PMID- 10856632 TI - Evolutionary relationships among 12 species belonging to three genera of the family Microhylidae in Papua New Guinea revealed by allozyme analysis. AB - To elucidate the potential of electrophoretic analysis for understanding relationships among microhylid frogs in Papua New Guinea, an allozyme analysis was conducted. A total of 119 individuals from nine species of Cophixalus, two species of Sphenophryne and one species of Barygenys, all of which belong to the family Microhylidae, were studied. Fourteen enzymes extracted from skeletal muscles and livers were analyzed by starch-gel electrophoresis. These enzymes were encoded by genes at 20 loci. There were 2-15 phenotypes produced by 2-12 alleles at these loci. The mean proportion of heterozygous loci per individual, mean proportion of polymorphic loci per population, and mean number of alleles per locus in 12 species were 6.1%, 17.1% and 1.17a on average, respectively. The NJ and ML trees constructed from Nei's genetic distances showed that the genus Sphenophryne can be distinguished biochemically from Cophixalus and Barygenys, and that the species groups of Cophixalus, which are similar in external morphology, can be divided biochemically into several species. PMID- 10856633 TI - High level of genetic divergence between sympatric color morphs of the littoral sea anemone Anthopleura orientalis (Anthozoa: Actiniaria). AB - Using enzyme electrophoresis and nematocyst analysis, the sympatrically occurring "light" and "dark" color morphs of the sea anemone Anthopleura orientalis from the Sea of Japan were shown to be two valid species. The "light" morph was identified as A. orientalis (Averintsev, 1967 Issledovaniya fauny morei: Vyp. 5 (13). Nanka, Leningrad, pp. 62-77), while the "dark" morph was designated as Anthopleura sp. The analysis of 21 isozyme loci revealed high value of Nei's genetic distance (D=1.284) between the two species, which are indistinguishable in their external morphology. The mean values of observed and expected heterozygosities for A. orientalis and Anthopleura sp. are high (H(o)=0.252+/ 0.061, H(e)=0.250+/-0.061 and H(o)=0.327+/-0.052, H(e)=0.351+/-0.054, respectively). The species differ significantly in the size of spirocysts and nematocysts, among which the atrichs from acrorhagi and the basitrichs from actinopharynx contribute most to the observed difference. Strong qualitative difference is revealed between distributions of nematocysts in mesenteric filaments of the two sea anemone species studied. The possible conspecificity of Anthopleura sp. with Anthopleura artemisia (Dana, 1848) is discussed and the conclusion made that these are two separate species. PMID- 10856634 TI - Lipophilic exudates of Pteridaceae - chemistry and chemotaxonomy. AB - A number of fern species, belonging to several genera of Pteridaceae, exhibit a more or less conspicuous farinose wax, which is mostly located on the lower leaf surface. Production of these waxes is often correlated with the presence of glandular trichomes. Particularly during the past two decades, a series of publications appeared on the chemical composition of these exudates. The major components were found to be flavonoids (chalcones, dihydrochalcones, flavanones, dihydroflavonols, flavones, flavonols), some of them with a complex substitution pattern, including esters and C-methyl derivatives, and even bisflavonoids. Diterpenoids and triterpenoids can also occur in such exudates. It is the purpose of this paper to survey the chemical composition of Pteridaceae exudates and their occurrence within the genera of the family. The chemotaxonomic significance of the flavonoid aglycones at the generic, specific and populational level is briefly discussed. PMID- 10856635 TI - Discrimination and identification of coastal Douglas-fir clones using needle flavonoid fingerprints. AB - This paper describes a method for discriminating and identifying 10 successful Douglas-fir (Pseudotsuga menziesii var. menziesii) clones using foliar flavonoids. All the 101 individuals analyzed by high performance liquid chromatography contained two proanthocyanidins: prodelphinidin and procyanidin and six flavonols: myricetin, quercetin, larycitrin, kaempferol, isorhamnetin and syringetin, but in different proportions. The experimental protocol used was very reproducible since the variation coefficients for each flavonoid did not exceed 9%. Submission of the flavonoid data to multivariate discriminant analysis allowed excellent discrimination of the 10 clones with 89% of the individuals being well-grouped. Then a clonal bank was established in which the fingerprint of each clone is defined by its position in the multidimensional space of the discriminant analysis. The clonal identity of several unknown individuals was determined with success by projecting their flavonoid data in a subsequent discriminant analysis. PMID- 10856636 TI - Tryptophol, a plant auxin isolated from the marine sponge Ircinia spinulosa. PMID- 10856637 TI - (-)-Amuronine from the leaves of Croton flavens L. (Euphorbiaceae). PMID- 10856638 TI - Calendin, tyrosol and two benzoic acid derivatives from Veronica persica (Scrophulariaceae). PMID- 10856639 TI - Flavones from Leaves of Tecomella undulata (Bignoniaceae). PMID- 10856640 TI - Quercetin 3-O-(6"-galloyl)-beta-D-galactoside from Polygonum viscosum (Polygonaceae). PMID- 10856641 TI - Flavonoid compounds of Lamyropsis cynaroides. PMID- 10856642 TI - Coelichelin, a new peptide siderophore encoded by the Streptomyces coelicolor genome: structure prediction from the sequence of its non-ribosomal peptide synthetase. AB - A gene cluster for the non-ribosomal synthesis of a peptide of unknown structure has been identified in the partial genome sequence of Streptomyces coelicolor. Using molecular and computational analyses, the total structure of a tripeptide siderophore synthesized by the non-ribosomal peptide synthetase within the cluster has been deduced from the translated sequence of its encoding gene. This represents a novel method for the structural assignment of natural products from genome sequence data. PMID- 10856643 TI - Regulation of the divergent guaBA and xseA promoters of Escherichia coli by the cyclic AMP receptor protein. AB - The gua promoter (guaP) of Escherichia coli resembles those for ribosomal RNA (rrn) operons and lies in a close back-to-back arrangement with the promoter for xseA (xseP). Transcription from guaP is subject to stringent control and growth rate-dependent regulation, and to repression by DnaA and PurR. In addition, transcription from guaP is regulated by the cyclic AMP receptor protein (CRP). Plasmid-borne promoter fusions to the receptor gene for chloramphenicol acetyl transferase were used to assess the role of CRP in controlling transcription from guaP and xseP following a downshift of cultures from rich into minimal medium. CRP is required to activate guaBA transcription and repress xseA transcription following downshift. Bandshift assays with a DNA fragment carrying the divergent promoters revealed specific binding of CRP. We propose that CRP, binding to a near-consensus site centred at -117.5, activates transcription from guaP and obstructs transcription from the xseA promoter. PMID- 10856644 TI - Desulfurization of alkylated forms of both dibenzothiophene and benzothiophene by a single bacterial strain. AB - Thirty-five bacterial strains capable of converting dibenzothiophene into 2 hydroxybiphenyl were isolated. Among them Rhodococcus erythropolis KA2-5-1 was chosen for further characterization because of its ability to retain high desulfurization activity stably. PCR cloning and DNA sequencing of a KA2-5-1 genomic DNA fragment showed that it was practically identical with dszABC genes from Rhodococcus sp. IGTS8, a representative carbon-sulfur-bond-targeted dibenzothiophene-degrading bacterium. KA2-5-1 desulfurized a variety of alkyl dibenzothiophenes through the specific cleavage of their C-S bonds. In addition, unexpectedly, KA2-5-1 also attacked alkyl benzothiophenes in a C-S-bond-targeted fashion. The purified monooxygenase, encoded by dszC of KA2-5-1, converted benzothiophene and dibenzothiophene into benzothiophene sulfone and dibenzothiophene sulfone, respectively, with the aid of an NADH-dependent oxidoreductase. This result raises the possibility that the same enzymatic step may be involved in desulfurization of alkylated forms of both dibenzothiophene and benzothiophene in KA2-5-1 cells. PMID- 10856645 TI - The FAPY-DNA glycosylase (Fpg) is required for survival of the cyanobacterium Synechococcus elongatus under high light irradiance. AB - The gene for the DNA repair enzyme Fpg from Synechococcus elongatus was detected immediately downstream of the photosystem I gene psaE. fpg is likely expressed together with psaE by transcriptional readover while psaE is mostly expressed independently. Segregated psaE and fpg deletion strains were obtained upon insertional inactivation of both genes in S. elongatus. These mutants are viable under photoautotrophic conditions, but fail to grow under high light regimes that likely cause oxidative stress. These high light sensitive phenotypes suggest that the Fpg protein, which has been shown to repair DNA lesions caused by reactive oxygen species in Escherichia coli, may be involved in the photoprotection of cyanobacteria against oxidative damage caused under high irradiance. PMID- 10856646 TI - Isolation and analysis of a circular form of the IncJ conjugative transposon-like elements, R391 and R997: implications for IncJ incompatibility. AB - The incompatibility between the chromosomally integrating, conjugative transposon like, IncJ elements R997 (ampicillin resistant) and R391 (kanamycin resistant) was examined by constructing strains harbouring both elements. Unusually, recA(+) strains harbouring the resistance determinants of both elements could be isolated but all strains lacked detectable extrachromosomal DNA. The phenotypic characteristics and transfer patterns observed suggested the formation of recombinant hybrids rather than strains harbouring both elements independently. Formation of strains harbouring two IncJ elements in a recA background was thus examined and resulted in the visualisation of extrachromosomal DNA. When R391 was transferred to a recA strain containing integrated R997, both elements co-existed stably and resulted in the isolation of a plasmid of 93.9 kb. When R997 was transferred to a recA strain harbouring an integrated R391, a plasmid of 85 kb was isolated. Comparison of restriction patterns for both elements revealed many common and several distinct fragments indicating a close physical relationship. These data suggest that although IncJ elements normally integrate at a unique site in the Escherichia coli chromosome, they possess the ability for autonomous replication which becomes manifest in a recA background when this site is occupied. This observation has implications for the nature of the incompatibility associated with IncJ elements and also provides a reliable method for isolating IncJ elements for molecular characterisation. PMID- 10856647 TI - Invasion of endothelial and epithelial cells by strains of Porphyromonas gingivalis. AB - Porphyromonas gingivalis is a periodontal pathogen that may also be involved in the pathogenesis of coronary heart disease. This microorganism has the ability to invade several cell lines. In this study, 26 different strains of P. gingivalis were tested for invasion of human umbilical vein endothelial cells and KB cells, a human oral epidermoid cell line. Abilities to invade both cell lines by an individual strain were similar, and their invasion efficiencies could be assembled into four groups: high, moderate, low and non-invasive. Of the 26 strains, only P. gingivalis AJW4 was non-invasive. Since the fimbriae are implicated as having a key role in invasion by this species, the presence of fimbriae on strain AJW4 was investigated. Using polymerase chain reaction (PCR), strain AJW4 was found to contain the fimA gene. Sequence analysis revealed it to be type IV according to the typing scheme developed by Amano et al. Further, fimA is transcribed in this strain as demonstrated by reverse transcription PCR and is expressed on the cell surface as visualized by negative staining and electron microscopy. The adherence+invasion of strain AJW4 was 38.7% of the most invasive strain (strain 381). However, the CFU ml(-1) of strain AJW4 recovered from within cells was 2.9% of strain 381. Even though strains AJW4 and W50 have the same type IV fimbriae, strain AJW4 is 8.9-fold more adhesive yet is internalized 170-fold less. These data indicate that the invasion efficiency of P. gingivalis is variable among the different strains, and that the expression of FimA is not sufficient for invasion. PMID- 10856648 TI - Virulence and mutation rates of Salmonella typhimurium strains with increased mutagenic strength in a mouse model. AB - Two strains of Salmonella typhimurium presenting increased mutation rates, either spontaneous or mediated by DNA damage, have been constructed. One of the strains carries a null mutS mutation, while the other harbors plasmid pRW30, which contains the Escherichia coli umuDC operon. The virulence of these strains has been determined by inoculating BALB/c or Swiss mice. The 50% lethal dose of both strains is identical to that obtained for the wild-type. Likewise, the two strains and the wild-type contribute equally to animal death in mixed infections. The frequency of Nal(R) mutants recovered from animals inoculated with either wild-type or MutS(-) cells was not affected by the presence of pRW30. These results indicate that the DNA damage which S. typhimurium cells can suffer during the infectious process by host cell metabolites does not cause induction of the SOS response at levels able to trigger the error-prone DNA repair pathway. PMID- 10856649 TI - Demonstration of the carbon-sulfur bond targeted desulfurization of benzothiophene by thermophilic Paenibacillus sp. strain A11-2 capable of desulfurizing dibenzothiophene. AB - Paenibacillus sp. strain A11-2, which had been primarily isolated as a bacterial strain capable of desulfurizing dibenzothiophene to produce 2-hydroxybiphenyl at high temperatures, was found to desulfurize benzothiophene more efficiently than dibenzothiophene. The desulfurized product was identified as o-hydroxystyrene by GC-MS and 1H-NMR analysis. Benzothiophene was assumed to be degraded in a way analogous to the 4S pathway, which has been well-known as a mode of dibenzothiophene degradation. These results suggest that benzothiophene desulfurization may share at least partially the reaction mechanism with dibenzothiophene desulfurization. PMID- 10856650 TI - Prevalence of the multiple antibiotic resistance operon (marRAB) in the genus Salmonella. AB - The multiple antibiotic resistance operon (marRAB) is a member of the multidrug resistance (mdr) systems. Similar to other mdr systems, this operon when induced encodes resistance to structurally and functionally unrelated antibiotics. marRAB has been shown to be conserved in the family Enterobacteriaceae, but within the genus Salmonella certain species appeared to be lacking this operon. To investigate how conserved the marRAB operon was in Salmonella, 30 veterinary isolates were examined by PCR, Southern blot, and dot blot analysis. Using DNA primers based on the marRAB operon of S. typhimurium, a predicted 2.3-kb amplicon resulted after PCR in 16 of the 30 organisms. The 2.3-kb DNA band from S. enteritidis was cloned and sequenced and shown to possess 99% sequence homology to marRAB from S. typhimurium. Using a labeled marRAB gene probe from S. enteritidis, Southern blot and dot blot analysis confirmed the presence of the operon in all 30 Salmonella species examined. Furthermore, when these isolates were induced with low levels of either tetracycline or chloramphenicol, increased antimicrobial resistance was observed to structurally and functionally unrelated antibiotics. Thus, the widespread occurrence of the marRAB locus in this genus prescribes judicious use of antimicrobials to avoid induction of a mdr phenotype. PMID- 10856651 TI - Protective effects of alpha-tocopherol and beta-carotene on para-nonylphenol induced inhibition of cell growth, cellular respiration and glucose-induced proton extrusion of bacteria. AB - para-Nonylphenol (NP) showed a dose-dependent inhibition against the cell growth of Bacillus subtilis, Micrococcus luteus, Pseudomonas aeruginosa and Staphylococcus aureus at 5-100 microM. However, other typical plastic-derived endocrine disruptors such as bisphenol A and di-2-ethylhexyl phthalate (DEHP) did not significantly affect the cell growth of these bacteria at 5-100 microM. The NP-induced cell growth inhibition was restored when concomitantly supplemented with lipophilic antioxidants such as alpha-tocopherol and beta-carotene, but not with hydrophilic antioxidants, ascorbic acid and (-)-epigallocatechin gallate (EGCG). NP also suppressed in a dose-dependent manner cellular oxygen consumption and glucose-induced proton extrusion of these bacteria at 10-100 microM. Both effects were prevented when added with alpha-tocopherol and beta-carotene, but not with ascorbic acid and EGCG. The significance of these results is discussed from the viewpoint of environmental microbiology and a possible biochemical mechanism of the inhibitory effect of NP is suggested. PMID- 10856652 TI - Rapid identification of 11 human intestinal Lactobacillus species by multiplex PCR assays using group- and species-specific primers derived from the 16S-23S rRNA intergenic spacer region and its flanking 23S rRNA. AB - Rapid and reliable two-step multiplex polymerase chain reaction (PCR) assays were established to identify human intestinal lactobacilli; a multiplex PCR was used for grouping of lactobacilli with a mixture of group-specific primers followed by four multiplex PCR assays with four sorts of species-specific primer mixtures for identification at the species level. Primers used were designed from nucleotide sequences of the 16S-23S rRNA intergenic spacer region and its flanking 23S rRNA gene of members of the genus Lactobacillus which are commonly isolated from human stool specimens: Lactobacillus acidophilus, Lactobacillus crispatus, Lactobacillus delbrueckii (ssp. bulgaricus and ssp. lactis), Lactobacillus fermentum, Lactobacillus gasseri, Lactobacillus jensenii, Lactobacillus paracasei (ssp. paracasei and ssp. tolerans), Lactobacillus plantarum, Lactobacillus reuteri, Lactobacillus rhamnosus and Lactobacillus salivarius (ssp. salicinius and ssp. salivarius). The established two-step multiplex PCR assays were applied to the identification of 84 Lactobacillus strains isolated from human stool specimens and the PCR results were consistent with the results from the DNA-DNA hybridization assay. These results suggest that the multiplex PCR system established in this study is a simple, rapid and reliable method for the identification of common Lactobacillus isolates from human stool samples. PMID- 10856653 TI - A single amino acid substitution beyond the C2H2-zinc finger in Ros derepresses virulence and T-DNA genes in Agrobacterium tumefaciens. AB - Ros is a chromosomally-encoded repressor containing a novel C2H2 zinc finger in Agrobacterium tumefaciens. Ros regulates the expression of six virulence genes and an oncogene on the Ti plasmid. Constitutive expression of these genes occurs in the spontaneous mutant 4011R derived from the octopine strain Ach-5, resulting in T-DNA processing in the absence of induction, and in the biosynthesis of cytokinin. Interestingly, the mutation in 4011R is an Arg to Cys conversion at amino acid residue 125 near the C-terminus well outside the zinc finger of Ros. Yet, Ros bearing this mutation is unable to bind to the Ros-box and is unable to complement other ros mutants. PMID- 10856654 TI - Phosphorylation of the GTS1 gene product of the yeast Saccharomyces cerevisiae and its effect on heat tolerance and flocculation. AB - The GTS1 gene from the yeast Saccharomyces cerevisiae showed pleiotropic effects on yeast phenotypes, including an increase of heat tolerance in stationary-phase cells and an induction of flocculation. Here, we found that the GTS1 product, Gts1p, was partially phosphorylated at some serine residue(s) in cells grown on glucose. Studies using mutants of protein kinase A (PKA) and CDC25, the Ras-GTP exchange activator, showed that PKA positively regulated the phosphorylation level of Gts1p. Overexpression of Gts1p in a mutant with attenuated PKA activity did not show any increase of heat tolerance and partially decreased flocculation inducibility, suggesting that phosphorylation of Gts1p is required for induction of these phenomena. PMID- 10856655 TI - A multidrug efflux transporter in Listeria monocytogenes. AB - A chromosomal gene (mdrL) was found in Listeria monocytogenes L028, showing a high degree of similarity with multidrug efflux transporters of the major facilitator superfamily (family 2). An allele-substituted mutant of this gene failed to pump out ethidium bromide and presented lower minimal inhibitory concentrations of macrolides, cefotaxime and heavy metals. This is the first multidrug efflux pump described in Listeria. PMID- 10856656 TI - Neutrophilic dermatoses: an overview. PMID- 10856657 TI - Polymorphonuclears: structure, function, and mechanisms of involvement in skin diseases. PMID- 10856658 TI - Chemoattractants as mediators of neutrophilic tissue recruitment. PMID- 10856659 TI - Sweet's syndrome: a neutrophilic dermatosis classically associated with acute onset and fever. PMID- 10856660 TI - Pyoderma gangrenosum. PMID- 10856661 TI - Erythema elevatum diutinum. PMID- 10856662 TI - Subcorneal pustular dermatosis. PMID- 10856663 TI - Dermatoses with intraepidermal IgA deposits. PMID- 10856664 TI - Neutrophilic eccrine hidradenitis. PMID- 10856665 TI - Neutrophilic drug eruptions. PMID- 10856666 TI - The extracutaneous involvement in the neutrophilic dermatoses. PMID- 10856667 TI - Pustular eruptions in systemic disease. PMID- 10856668 TI - Neutrophilic dermatoses associated with hematologic disorders. PMID- 10856669 TI - Pharmacologic modulation of neutrophil functions. PMID- 10856670 TI - Bacteroides, Prevotella, and Porphyromonas species: a review of antibiotic resistance and therapeutic options. AB - Recent basic and clinical research efforts have shed more light on the taxonomy, microbiology, epidemiology, antimicrobial susceptibility and treatment of Bacteroides, Prevotella, and Porphyromonas species. Of all anaerobic bacteria, Bacteroides is the most frequently isolated pathogen from clinical specimens, including blood. Bacteroides, Prevotella and/or Porphyromonas species have been isolated from clinical specimens in cases of infection from almost all anatomic sites. Several multicentre surveys have documented an alarming gradual increase of resistance rates of Bacteroides, Prevotella and Porphyromonas species worldwide. Antimicrobial agents active against >99% of clinical isolates of Bacteroides are metronidazole, chloramphenicol and carbapenems. Agents active against 95-99% of Bacteroides fragilis isolates are the beta-lactam/beta lactamase inhibitor combinations. B. fragilis group species other than B. fragilis are more likely to be resistant to beta-lactam/beta-lactamase inhibitor combinations than B. fragilis. PMID- 10856671 TI - Colorimetric in vitro susceptibility testing of penicillins, cephalosporins, macrolides, streptogramins, tetracyclines, and aminoglycosides against Borrelia burgdorferi isolates. AB - The spectrum of antibiotic susceptibility of Borrelia burgdorferi has been only partially defined. In the present study the effectiveness of 12 antimicrobials, belonging to six different antibiotic classes have been tested against Borrelia burgdorferi s.s. (N=3), Borrelia garinii (N=3), Borrelia afzelii (N=3), Borrelia valaisiana (N=1), and Borrelia bissettii (N=1) isolates. These isolates were analysed by a new standardised colorimetric minimal inhibitory concentration (MIC) method based upon colour changes that result from actively metabolizing spirochaetes after 72 h of incubation. Piperacillin (MIC90: 0.08 mg/l), ceftriaxone (MIC90: 0. 04 mg/l), cefotaxime (MIC90: 0.15 mg/l), azithromycin (MIC90: 0.015 mg/l), roxithromycin (MIC90: 0.05 mg/l) and quinupristin/dalfopristin (MIC90: 0.12 mg/l) gave the lowest MIC values. Minimal inhibibitory activity of amoxycillin (MIC90: 1.04 mg/l), cefixime (MIC90: 1.33 mg/l), cefoperazone (MIC90: 0.83 mg/l) tetracycline (MIC90: 0.29 mg/l) and minocycline (MIC90: 0.30 mg/l) was slightly lower, whereas borrelia were resistant to amikacin (MIC90: >128 mg/l). Mean minimal borreliacidal concentrations (MBCs) were representatively determined for piperacillin (MBC: 1.8 mg/l), ceftriaxone (MBC: 2.0 mg/l), azithromycin (MBC: 0.82 mg/ml), roxithromycin (MBC: 1.8 mg/l), quinupristin/dalfopristin (MBC: 5.0 mg/l), minocycline (MBC: 5.8 mg/l), and amikacin (MBC: >128 mg/l) by using conventional subculture for three weeks in combination with dark-field microscopy. B. garinii proved to be the most susceptible of the genospecies tested. Our study showed excellent in vitro antimicrobial activity of all classes of antibiotics tested, except the aminoglycosides and hence their suitability for therapy of Lyme disease. PMID- 10856672 TI - Rapid detection of methicillin resistance in staphylococci using a slide latex agglutination kit. AB - The slide latex agglutination test, MRSA-Screen, was compared with the mecA polymerase chain reaction (PCR) and traditional susceptibility test methods for the detection of methicillin resistance in Staphylococcus aureus and coagulase negative staphylococci. The MRSA-Screen test detected the same number of methicillin-resistant S. aureus as the mecA PCR and the traditional susceptibility tests. It correctly identified all 21 methicillin-susceptible S. aureus as being sensitive. It also produced the same result as the mecA PCR in identifying a methicillin-resistant S. aureus among six isolates classified as borderline resistant by traditional susceptibility tests. The MRSA-Screen test and mecA PCR detected methicillin resistance in 10 and 15 of 17 methicillin resistant coagulase-negative staphylococci, respectively. From these results, it is concluded that the MRSA-Screen is a very accurate, reliable and rapid method of detecting methicillin resistance in S. aureus and is suitable for use in clinical microbiology laboratories. Further study of its use in detecting methicillin resistance in coagulase-negative staphylococci is required. PMID- 10856673 TI - Comparison of vancomycin pharmacodynamics (1 g every 12 or 24 h) against methicillin-resistant staphylococci. AB - This study compared the duration of serum bactericidal activity for vancomycin, 1 g every 12 or 24 h at steady state, against methicillin-resistant Staphylococcus aureus (MRSA) and coagulase-negative staphylococci (MR-CNS). All four test isolates were susceptible to vancomycin with minimal inhibitory concentration (MIC) values of either 2 or 4 mg/l. Serum bactericidal titres (SBTs) were run in duplicate and serum bactericidal activity (SBA) was defined as the time points at which all subject SBTs were greater than or equal to 1:2. For the every 12-h regimen, SBA was 10-12 h. With the every 24-h regimen, the duration of SBA was 10 16 h for MRSA and 8-10 h for MR-CNS. The pharmacodynamic data suggest that for those with good renal function a Q12h dosing interval is most appropriate for MR CNS or staphylococcal isolates with MICs of 4. PMID- 10856674 TI - Quantitative RT-PCR analysis of multiple genes encoding putative metronidazole nitroreductases from Helicobacter pylori. AB - Metronidazole (Mtz), a pro-drug, requires reductive activation by ferredoxin-like electron carrier proteins to kill bacteria and Mtz resistance is associated with a decrease or deficiency of Mtz nitroreductase activities in a target cell. Several genes encoding ferredoxin-like or -linked proteins such as pyruvate oxidoreductase (POR), ferredoxin oxidoreductase (FOR), ferredoxin (FdxA), ferredoxin-like protein (FdxB), flavodoxin (FldA) and oxygen insensitive nitroreductase (RdxA) have been identified from the complete genomic sequence of Helicobacter pylori. To understand the roles of these genes in H. pylori Mtz resistance, the gene expression for the proteins was examined using a method optimized for quantitative reverse transcription polymerase chain reaction (RT PCR). The RT-PCR products of FOR and RdxA were significantly decreased in the total RNA prepared from H. pylori cultured in the presence of Mtz as compared to the total RNA prepared from H. pylori cultured without Mtz in the media. A slight decrease, however, in band intensity of the RT-PCR products of the POR and, to a lesser extent, FdxB was obtained in the presence of Mtz. In contrast, the RT-PCR products of the FdxA, FldA, and GalE (UDP-galactose 4-epimerase; a control gene) were unchanged in total RNA prepared from H. pylori cultured with or without Mtz in the culture media. These results suggest that Mtz resistance may also be acquired by decreasing the transcription of some genes involved in Mtz reductive activation, in addition to the mutation in some individual genes such as rdxA. PMID- 10856675 TI - Molecular basis of extended-spectrum beta-lactamase production in nosocomial isolates of Klebsiella pneumoniae from Merida, Venezuela. AB - Twelve Klebsiella pneumoniae isolates resistant to expanded-spectrum cephalosporins and aztreonam, from patients with nosocomial septicaemia at the intensive care unit of the Andes University Hospital, Merida, Venezuela, were studied for production of extended-spectrum beta-lactamase (ESbetaL) activity. All were also resistant to gentamicin, kanamycin, tetracycline and chloramphenicol but sensitive to cefoxitin, imipenem, amikacin and tobramycin. Production of ESbetaL activity was confirmed by restoring susceptibility to ceftazidime in the presence of clavulanic acid. All isolates carried an identical plasmid of approximately 87 kb. Resistance to beta-lactams, aminoglycosides, tetracycline and chloramphenicol was lost en bloc after plasmid curing by treatment with acridine orange and was transferable en bloc to Escherichia coli by conjugation. Transconjugants always showed the same plasmid profile as that of Klebsiella donors. Isoelectric focusing analysis of the crude extracts of transconjugants showed in all cases, the presence of two beta-lactamases of pI 5.4 and 8.2. Analysis of the plasmid carried by one of the transconjugants by means of hybridization assays, revealed the presence of both bla(TEM) and bla(SHV) determinants. Cloning and sequencing of each determinant identified them as bla(TEM-1) and bla(SHV-5), respectively, the latter being responsible for the ESbetaL activity. Results of this study indicate that ESbetaL determinants of the SHV-type carried by transferable elements, are spreading among nosocomial isolates of K. pneumoniae in Merida, Venezuela. PMID- 10856676 TI - Prevention by L-alpha-phosphatidylcholine of antifungal activity in vitro of liposome-encapsulated imidazoles determined by using time-killing curves. AB - The antifungal activity of the imidazole derivatives miconazole and ketoconazole was reduced when they were entrapped in liposomal structures and significant differences were detected between small unilamellar vesicles (SUV) and multilamellar vesicles (MLV). To understand which component of liposomes interfered with the antifungal activity of miconazole and ketoconazole, we examined the influence of pure egg and soy L-alpha-phosphatidylcholine and cholesterol on activity against Candida albicans ATCC E10231 by time killing curves. Association of phospholipids-cholesterol-imidazole leads to an inhibitory effect on the antifungal activity comparable to that shown when miconazole or ketoconazole were entrapped in SUV liposomes or when miconazole and ketoconazole were incubated in the presence of L-alpha-phosphatidylcholine. The antifungal activity determined in the presence of cholesterol was comparable to that observed with the free drugs. Inhibition of the antifungal activity of miconazole and ketoconazole by phospholipids is dependent on the phospholipid concentration but is independent of the source of phospholipids (egg or soy). Cholesterol had no influence on the antifungal activity of the imidazoles, unlike the effect on other antifungal drugs, such as amphotericin B. PMID- 10856677 TI - A comparison of the bactericidal effects and cytotoxic activity of three types of oxidizing water, prepared by electrolysis, as chemical dental plaque control agents. AB - Acid oxidizing water (AOW), neutral oxidizing water (NtOW) and acid oxidizing water with a low available chlorine concentration (AOW-LC) may be obtained by electrolyzing a solution of tap water containing various quantities of NaCl and HCl. This study compared the bactericidal effects of these waters on cariogenic and periodontopathogenic bacteria and their cytotoxicities against epithelial cells. AOW, NtOW and AOW-LC showed considerable bactericidal effects. The cytotoxicity of AOW-LC was significantly lower than the other solutions tested (P<0.0001). The results indicated that the three types of oxidizing water had similar activity in inhibiting bacterial plaque formation as conventional chemical plaque-control agents. PMID- 10856678 TI - Activity of poloxamer CRL-1072 against drug-sensitive and resistant strains of Mycobacterium tuberculosis in macrophages and in mice. AB - The present experiments evaluated a new, highly refined poloxamer, CRL-1072, alone and in combination with antibiotics against drug-sensitive and -resistant organisms. In macrophage culture, CRL-1072 reduced the drug concentration inhibiting 99% of control growth of isoniazid (INH) from 10 to 0.15 mg/l (fractional inhibitory concentration=0.07) for a drug-resistant strain. CRL-1072 also increased the susceptibility of drug-resistant strains of Mycobacterium tuberculosis to INH, streptomycin, rifampicin, pyrazinamide, ethambutol, PAS, thiacetazone and ethionamide. Fractional inhibitory concentration values of <0.5 indicated significant synergistic activity. In studies of acute infection in mice, CRL-1072 was only weakly bacteriostatic when used as a single agent but increased the bactericidal activity of INH, streptomycin, rifampicin, pyrazinamide and clindamycin, but not that of ethambutol. CRL-1072 enhanced the bactericidal activity of streptomycin against a streptomycin resistant strain of M. tuberculosis in a murine infection. PMID- 10856679 TI - Comparative pharmacoeconomic study of vancomycin and teicoplanin in intensive care patients. AB - Randomized clinical trials and meta-analyses have not demonstrated any statistically significant differences between teicoplanin and vancomycin with regard to efficacy. A cost-minimization analysis was conducted to compare the economical impact of the treatment with vancomycin and teicoplanin in intensive care patients. Information on resource utilization was retrospectively collected from 100 consecutive clinical histories of patients hospitalized in a Spanish Intensive Care Unit, who had been given a glycopeptide antibiotic (50 teicoplanin and 50 vancomycin) for the treatment of a suspected or proven infection. Although personnel, material, and monitoring costs were higher in the vancomycin group, the acquisition costs and the total costs were much lower in this group, so the resulting total costs per day were 5508 ptas (33 euros) for vancomycin-treated patients and 9893 ptas (59.5 euros) for teicoplanin-treated patients. The savings with vancomycin for a 10-day course of treatment would be approximately 40697 ptas (244.5 euros) per patient. Results were consistent for a variety of conditions that were included in the sensitivity analysis. PMID- 10856680 TI - In vitro activity of four fluoroquinolones against clinical isolates of Pseudomonas aeruginosa determined by the E test. AB - Ciprofloxacin, levofloxacin, ofloxacin, and trovafloxacin were tested by the E test against 100 clinical isolates of Pseudomonas aeruginosa. Ciprofloxacin was the most active of the tested agents with 82% of isolates having a MIC 8). Levofloxacin and trovafloxacin had nearly identical potency: 75% and 76% of the isolates were inhibited by 8 for levofloxacin; 0.19->8 for trovafloxacin). Ofloxacin was the least active of the four quinolones, with 43% of the isolates having a MIC >2 mg/l. All isolates resistant to ciprofloxacin were also resistant to the other agents, i.e. resistance to ciprofloxacin predicted resistance to all the quinolones tested in every case. This data demonstrates that fluoroquinolones are active agents against P. aeruginosa. In vitro susceptibility testing, however, is crucial to assess the resistance pattern in any specific location and for each individual agent. PMID- 10856681 TI - Intravenous pharmacokinetics of ciprofloxacin in goats. AB - The pharmacokinetics of ciprofloxacin in goats was studied following intravenous administration. A single dose of 10 mg/kg was administered as an intravenous bolus, and serum samples were collected at predetermined intervals over a 24 h period. Ciprofloxacin levels were measured using high pressure liquid chromatography and the resulting concentration versus time curve was analyzed using a non linear regression analysis. The data were best represented by a two compartment pharmacokinetic model with a mean elimination half-life of 2.72+/ 1.04 h. Mean pharmacokinetic parameters obtained were area under the curve (AUC: 10.320+/-5.137 microg/ml per h), mean residence time (MRT: 3.334+/-1.428 h), apparent volume of distribution (Vdss: 3.373+/-0.893 l/kg) and total body drug clearance (tCl 19.596+/-9.059 ml/min per kg). Ciprofloxacin in goats showed the general pharmacokinetic characteristics of a typical fluoroquinolone antibacterial agent and we recommend a dose of 10 mg/kg to be administered intravenously at 12 h intervals to goats. PMID- 10856682 TI - Review of dosimetric functions for meterset calculations. AB - Mathematical expressions used to calculate doses in a patient, based on data measured in a phantom, have to be simple, understandable, and reliable to minimize possible calculational error. In light of this concern, this paper reviews the dosimetric functions used in meterset calculations to determine the treatment times or monitor units for a prescribed dose. The dosimetric functions are the percent depth dose, the tissue-air ratio, the tissue-phantom ratio, and the inverse square law. This review examined the definition of the dosimetric functions, the inter-relationships among the dosimetric functions, and the mathematical expressions used in meterset calculations for nonstandard source-to surface distances in a phantom. PMID- 10856683 TI - Application of enhanced dynamic wedge to stereotactic radiotherapy. AB - Stereotactic radiotherapy has developed into a useful treatment technique in which conformal dose distributions can be delivered with precision and accuracy. In some cases, the position of the target volume relative to surrounding critical structures demands careful evaluation of fixed beam paths so that dose to these critical structures can be minimized. Micromultileaf collimators aid in conforming dose to the target volume but may not allow adjustment of an individual beam's intensity (intensity modulation) in an effort to achieve dose uniformity throughout the treatment volume. Enhanced dynamic wedge (EDW) is demonstrated to be a valuable tool in improving the dose distribution in stereotactic radiotherapy treatments in which these fixed, conformal fields must be used due to constraints in beam trajectories. Four cases are presented which show the potential for gain in dose uniformity with the addition of EDW. These cases represent typical applications of EDW to conformal stereotactic radiotherapy. PMID- 10856684 TI - BIOPLAN: software for the biological evaluation of. Radiotherapy treatment plans. AB - Distributions of absorbed dose do not provide information on the biological response of tissues (either tumor or organs at risk [OAR]) to irradiation. BIOPLAN (BiOlogical evaluation of PLANs) has been conceived and developed as a PC based user-friendly software that allows the user to evaluate a treatment plan from the (more objective) point of view of the biological response of the irradiated tissues, and at the same time, provides flexibility in the use of models and parameters. It requires information on dose-volume histograms (DVHs) and can accept a number of different formats (including DVH files from commercial treatment planning systems). BIOPLAN provides a variety of tools, such as tumor control probability (TCP) calculations (using the Poisson model), normal tissue complication probability (NTCP) calculations (using either the Lyman-Kutcher Burman or the relative seriality models), the ATCP method, DVH subtraction, plots of NTCP/TCP as a function of prescription dose, tumor and OAR dose statistics, equivalent uniform dose (EUD), individualized dose prescription, and parametric sensitivity analysis of the TCP/NTCP models employed. PMID- 10856685 TI - Point dose variations with time during traditional brachytherapy for cervical carcinoma. AB - In traditional brachytherapy for carcinoma of the cervix, doses are often prescribed to specifically chosen points (A and B) and the normal tissue tolerance calculated at specific reference points in the bladder and rectum. These tolerance doses are often used to modify the brachytherapy treatment plan. It is inherently assumed that the position of the brachytherapy applicator does not change in relation to the relevant anatomical structures over the time-course of an implant. To assess the accuracy of this assumption, 2 sets of localization films were obtained for each implant in 28 patients, 1 prior to loading and another after the removal of the radioactive sources. Significant applicator movement and, consequently, significant dose variations were ob: served. Therefore, isolated one-time dose measurements to normal critical structures should not be used as the sole basis for making therapeutic decisions. The magnitude of dose variations and their clinical significant are discussed. PMID- 10856686 TI - Implementation of enhanced dynamic wedge in the focus rtp system. AB - The FOCUS RTP system implementation of Varian's enhanced dynamic wedge (EDW) is presented. Calculations of both dose distributions and wedge factors (WFs) are based on segmented treatment tables (STTs). Calculating dose requires a "transmission matrix" derived from an STT to model the modified fluence from the source. The dose calculation is then performed using either the Clarkson or convolution/superposition algorithms. An initial "primary dose/monitor unit (MU) fraction" WF estimate at the weight point of symmetric and asymmetric fields is calculated from the STT as the ratio of MU delivered on the axis of the weight point divided by total MU delivered for the treatment field. In our approach, we go beyond this initial estimate with a "scatter dose" correction. This requires measured 60 degrees WFs for 5 fields. Scatter corrections derived from measured WFs are interpolated for other wedge angles and field sizes in much the same way as arbitrary wedge angle STTs are derived from a "golden STT" using the "ratio of tangents" formalism. Dose comparisons with measured distributions show good agreement to within 3% or 3 mm for 6-MV beams and all EDW angles. Agreement with measurements to within 1% is obtained for WFs in all symmetric and asymmetric fields for 6- and 10-MV beams. For large wedge angles and field sizes, this represents a significant improvement over the 3% to 4% errors often observed using the MU fraction model alone. PMID- 10856687 TI - Adapting the waterproof BMS-96 diode array for isodose determination of dynamic beams. AB - We report the application of the Schuster BMS-96 waterproof linear diode array for isodose determination of dynamic beams. The array recorded beam profiles correctly, while depth dose distributions of dynamic beams with large variations in dose rate were registered erroneously. The deviations could be eliminated by appropriate software modifications. Until the software is revised, true isodoses can be obtained by rescaling each individual profile to the depth dose curve as measured with a single ionization chamber. After the corrections presented in the paper, isodoses interpolated from these corrected data sets agreed with ionization chamber measurements within 1-2%. PMID- 10856688 TI - Application of a genetic algorithm to optimizing radiation therapy treatment plans for pancreatic carcinoma. AB - The performance of an automated treatment planning algorithm was tested using cases of patients with pancreatic carcinoma; the system implements optimization tools that suggest high-quality plans for consideration by the planner and physician, making best use of the capabilities of a conventional linear accelerator: isocentric setup, shaped fields, and wedges. Ten consecutive patients presenting with pancreatic cancer were first planned using a conventional 3-field protocol to provide a basis for comparison. Each was then planned using an automated optimization technique using a genetic algorithm and a dose-based score function subject to volume-dose constraints. Two sets of optimized plans were created, 1 using only axial beams and the other permitting non-axial beams. The improvement afforded by the optimization was assessed by comparing the score function results and by computing the combined normal tissue complication probability (NTCP) for a constant isocenter dose. In all 10 cases, optimization improved the dose-based score function. In 9 cases, the non-axial plan scored higher than the axial plan. Optimization driven by the dose-based score function improved or equaled the predicted NTCP in 8 axial and 9 nonaxial plans. This study demonstrates progress toward the goal of developing an automated planning tool that can robustly suggest high-quality plans. PMID- 10856689 TI - Where is the light field edge: perception of different operators on different surfaces. AB - On most radiotherapy treatment units, a light field indicates on the patient's skin where the treatment field will irradiate the patient. It was the aim of the present study to investigate the perception of the light field edge by different operators on different surfaces under different lighting conditions. Ten radiation therapists and physicists were asked to mark the light field edge of an 8 x 10-cm2 radiation field from a linear accelerator on prepacked radiographic film. Each operator marked the field 4 times each with the room light turned on and dimmed as usual for patient setup. Two operators marked the field on 5 different surfaces (film envelope, brown solid water, clear plastic used for the manufacturing of immobilization shells, black rubber, and Orfit patient immobilization material). The interoperator reproducibility (+/- 0.39 mm, ISD) was larger than the intraoperator reproducibility (+/- 0.27 mm). The light field was judged consistently to be 0.6 mm smaller in the light room than under dimmed light conditions and the physicists judged the field to be approximately 0.4 mm smaller compared to the radiation therapists' judgement. Compared to the yellow film wrapping, the light field on solid water, black rubber, and the clear plastic were judged to be 0.6 mm smaller by both operators. The same observation was made using a slotted block tray, which also gave the worst reproducibility of perceived field edges. While these systematic errors are relatively small and difficult to correct for, it appears to be important to be at least aware of them, in particular if the light field is used to junction radiation fields with steep penumbras, as commonly done in megavoltage treatments of head & neck and breast cancer. PMID- 10856690 TI - Preoperative assessment and management of pediatric heart transplantation. AB - The period of preoperative management of the pediatric cardiac transplant patient can be divided into three phases: determination of transplant feasibility, listing, and medical management. Chronic infection, irreversible elevation of pulmonary vascular resistance, and intractable disease in other organ systems may all be contraindications for transplantation. The United Network for Organ Sharing has recently changed its listing guidelines. Adolescent donors are now preferentially, to some extent, allocated to adolescent recipients. Management of pediatric patients awaiting cardiac transplantation encompasses optimization of cardiac output through the use of vasodilators and oral and intravenous inotropic agents. For those patients listed for transplantation who have single ventricle lesions, such as hypoplastic left heart syndrome, management of heart failure also includes balancing systemic and pulmonary blood flows. Mechanical support of the circulation with extracorporeal membrane oxygenation or ventricular assist devices can be used as a bridge to transplant in pediatric patients. PMID- 10856691 TI - Survival after listing for cardiac transplantation in children. AB - Despite improvement in surgical and medical management of children with congenital and acquired heart disease, cardiac transplantation remains an important therapeutic option for infants and children with end-stage heart disease. Ultimate survival in patients who are listed for transplantation is a function of both mortality while awaiting transplantation and survival after transplantation. Survival of heart transplantation is affected by the severity of illness before transplantation, the unique pathophysiology of certain defects, and the availability of donor hearts. Outcome following listing for transplantation is best studied with the use of recent modifications in statistical methods of competing outcomes analysis. By this analysis a predicted mortality while waiting among all pediatric patients is 20% at 1 year, with 67% undergoing transplantation, 10% still on the list awaiting transplant, and 3% removed from the list. Among infants, most of them with hypoplastic left heart syndrome, 60% will have transplantation by 6 months after listing, with 27% of patients dying while waiting. In infants the major risk factors for death while waiting are the need for inotropic support at listing, smaller size, and recipient blood type. In older children risk factors for death while waiting are Status 1 at listing and a need for mechanical ventilation. Intermediate-term survival after transplant is excellent in all age groups with 86% alive at 6 months, 84% at 1 year, and 73% at 5 years. Survival after transplant in infants is comparable to survival in older children, although the early mortality after transplantation is greater. Infants who have recently undergone sternotomy or received organs from donors who did not die of closed head trauma are more likely to die early after transplant. Among older children risk factors for death after transplantation include the need for a mechanical support device or a younger age in patients greater than 1 year of age. Death following transplantation is primarily related to early graft failure in infants, whereas rejection, infection, and sudden death account for the majority of deaths in older children. Although improved immunosuppressive agents promise to lead to even better survival rates after transplantation, greater access to donors is essential if overall survival is to be improved. PMID- 10856692 TI - Transplantation in complex congenital heart disease. AB - Three hundred and forty-five pediatric patients underwent orthotopic heart transplantation at Loma Linda University Medical Center throughout August 1999. Seventy-five percent of these patients had the diagnosis of congenital heart disease as the primary indication for transplantation. Two hundred and fifty seven were infants and of these, 91 were neonates. Forty-nine percent had the primary diagnosis of hypoplastic left heart syndrome or its equivalent. Thirty patients had variant situs and 15 of these had situs inversus. Seventy-seven children had undergone one or more previous cardiac and/or thoracic procedures. There appeared to be a survival advantage with neonatal as compared to infant heart transplants (77% vs. 63%). Technical considerations for reconstruction have made congenital heart transplantation a low perioperative risk factor as compared to the child with cardiomyopathy. Attention to (1) recipient selection; (2) adequate en bloc removal of the heart and accompanying vessels; (3) use of low flow bypass techniques with limited circulatory rest; and (4) multidisciplinary management have been the fundamental approach at this institution. Although challenging, high perioperative survival and long-term success can be achieved with heart transplantation in patients with high-risk or inoperable congenital heart disease. PMID- 10856693 TI - Immunosuppression for pediatric cardiac transplantation in the modern era. AB - With the advent of the T cell activation inhibitors such as cyclosporine, heart transplant success rates for pediatric patients have improved to the point that the initially restricted ages and indications have expanded considerably. Currently the half-life (50% still alive) for children transplanted in the early 1980s is approximately 12-14 years. Decades-long survival seems likely. Components and functions of the immune system are naive and change during postnatal development. Maturation occurs not only in the first years of life, but well through adolescence and even into adult life. These age-dependent changes within the immune system greatly complicate any attempt to assess immune implications for the use of immunosuppression in children. Since the introduction of cyclosporine, immunosuppression regimens have been virtually unchanged through the 1990s. Recently, there have been significant new immune pharmacological agents which are now commercially available, or still in investigational stages of development. The new maintenance immunosuppressive drugs are either inhibitors of de novo synthesis of nucleotides (purines or pyrimidines), or are immunophilin binding drugs that inhibit signal transduction in lymphocytes. The newer inhibitors of de novo nucleotide synthesis include mycophenolate mofetil, mizoribine, brequinar and leflunomide. The immunophilin-binding drugs are cyclosporine, tacrolimus and rapamycin. Antibody preparations such as ATG, ATGAM and OKT3, as well as the newer biological agents, which specifically bind to the IL-2 receptor, basiliximab and daclizumab, are discussed. The potential for biologicals which inhibit co-stimulation are also discussed. There may be dramatic changes in protocols used clinically as a result of these new agents over the next decade. The increasing understanding of the alloimmune response as well as the clinical use of these newer drugs promise even better long-term results. PMID- 10856694 TI - Pulmonary hypertension in pediatric heart transplantation. AB - Pulmonary hypertension can pose a significant problem in the management of children with congestive heart failure. Assessment of pulmonary artery anatomy, pressures and (when possible) pulmonary vascular resistance is critically important in the evaluation of these children when they are under consideration for heart transplantation. Severe, fixed elevation of the pulmonary vascular resistance is a contraindication to heart transplantation because of concerns of acute post-transplant donor right ventricular failure. However, even modest degrees of pulmonary hypertension can complicate the post-operative management of pediatric heart transplant recipients. This review will provide information regarding the recognition, diagnosis, and pre-operative and post-operative management of pulmonary hypertension in patients under consideration for heart transplantation. PMID- 10856695 TI - Transplant coronary artery disease in children. AB - Transplant coronary artery disease is an accelerated vasculopathy that occurs in adult and pediatric heart transplant recipients, and it is a leading cause of death among late survivors. This form of coronary disease, also known as graft coronary disease, differs from classical atherosclerosis in both histologic and angiographic features and it progresses much more rapidly. Although its pathogenesis has not been determined precisely, both immune and non-immune mechanisms appear to contribute, with a final common pathway of endothelial injury due to both antigen-dependent and antigen-independent factors. Many investigators believe both cellular and/or humoral rejection play a direct role in its etiology. In children the true incidence of the condition is unknown, although a multicenter survey identified 58 (7.2%) patients among 815 transplant recipients at 17 centers. Detection remains difficult. In the past, non-invasive methods have been unsatisfactory, although recent experience has suggested that Dobutamine stress echocardiography may be promising. Once a diagnosis is made, treatment has been limited to palliation by either intracoronary interventional procedures or surgical coronary bypass grafting, and to cardiac retransplantation with its own set of problems. Current efforts are directed at prevention. Blood levels of cholesterol have been reduced in adults treated with Pravastatin, but there have been no reports of its use in children. In adults additional agents with potential benefit have included calcium channel blockers and ACE inhibitors. A multicenter trial in children is needed to answer the many remaining questions regarding transplant coronary disease in this age group. PMID- 10856696 TI - Post-transplantation lymphoproliferative disorders: advances in diagnosis, prevention and management in children. AB - With improving operative and postoperative survival after pediatric thoracic transplantation, attention has appropriately begun to focus on complications of long-term immunosuppression. One important complication is post-transplantation lymphoproliferative disorders. Much has been learnt about this spectrum of disorders over the last 15 years, including the pivotal role of primary Epstein Barr virus infection in the etiology of most cases. Despite these advances, nomenclature remains confusing for the clinician, prediction of outcome is imprecise and treatment strategies are poorly defined. Indeed, no treatments have been subjected to comparative prospective clinical trials. Only recently has attention focussed on strategies for prevention. This article will review the current state of knowledge of post-transplantation lymphoproliferative disorders, with emphasis on recent advances in diagnosis, prevention and management. PMID- 10856697 TI - Developmental outcomes and cognitive functioning in infant and child heart transplant recipients. AB - Pediatric heart transplantation has become a mainstay in the treatment of end stage heart disease in infants and children. There is, however, sparse information on post-transplant developmental and cognitive functioning. At Loma Linda University Children's Hospital 223 infants have undergone heart transplantation surgery with a survival rate of 73% (n=165). This article reports on four areas of investigation of this cohort of infants. Infant development: Two studies were done utilizing the Bayley Scales of Infant Development (mean 100, S.D.+/-15). The first study (n=48) showed mean values within the normal range [Mental Development Index (MDI) 87; Psychomotor Developmental Index (PDI) 90]. The second study (n=23) showed developmental scores within normal limits in 4-8 month olds with a tendency for decline in development at 12-24 months (MDI 83, PDI 77). Child development: Ninty-one infant recipients were greater than 5 years old. Forty-five children were excluded because of long distances from the hospital, second transplantation, abnormal karyotype, primary language non English, or invalid testing. The Wechsler Preschool and Primary Scale of Intelligence-Revised testing of 5-6-year-olds (n=23) showed a Full Scale IQ of 74, Performance IQ of 76, and Verbal IQ of 77. The Wechsler Intelligence Scale for Children-III testing of 7-10-year-olds (n=23) showed a Full Scale IQ of 86, Performance IQ of 89, and Verbal IQ of 86. The Wechsler Individual Achievement Test (mean 100, S.D.+/-15) yielded a Total Composite of 91, Mathematics Composite of 86, Language Composite of 98, and a Reading Composite of 94. Visual spatial skills: Visual motor integration (mean 100, S.D.+/-15) was evaluated in 5-10-year olds with a mean of 87; however, 52% of the children had scores below 1 S.D. Subtests from the Wechsler scales that assess visual motor and visual spatial skills indicated significant deficits. Behavior: Younger children (n=33) demonstrated behaviors indicative of social isolation. Older children (n=36) showed behavior that was within the normal limits, but depression was noted in a significant number of them. Conclusions: Infant heart transplant recipients demonstrate IQ and achievement levels within the normal range, but there is a significant amount of variability with more children than would be expected scoring in the lower ranges. Children with heart transplantation are at risk for visual spatial skill deficits. Young children are at risk for social isolation while symptoms of depression are noted in older children. PMID- 10856698 TI - Clinical outcome 10 years after infant heart transplantation. AB - The feasibility of heart transplantation for infants has now been established. Clinical outcome data is necessary to assist in targeting areas for improvement and for counseling families considering this option. This report describes clinical outcome in 29 infant heart transplant recipients who have survived at least 10 years. A query of the transplant database, referring physicians and parental questionnaire was performed. Patient survival for the overall infant population is 64% at 13 years. Parents of 19/29 (55%) children described them as developmentally normal. Three children have had a severe developmental outcome. Sixteen of 29 children are in mainstream school environments. Four have repeated one grade in school. Speech delay was present in 10/26 (38%). Somatic growth is normal in 88%. All children are NYHA class I. Renal function shows only modest insufficiency with most recent BUN (mean+/-S.D.)=25+/-7 mg/dl and serum creatinine=0.8+/-0.2 mg/dl. Only four children have creatinine levels >1 mg/dl. No child requires dialysis. No children have developed post-transplant lymphoproliferative disease beyond 10 years. Four children have experienced rejection beyond 10 years with one mortality due to rejection and transplant coronary artery disease. Conclusion: Heart transplantation during infancy is technically feasible and results in good survival. Many children have some degree of learning disability but most are mild and the children function well in society. Improvements in surgical techniques may improve developmental outcome. Other side-effects of immunosuppression are manageable and most survivors have a good functional outcome. PMID- 10856699 TI - CD154 (CD40 ligand). AB - CD40 ligand, a type II transmembrane protein recently renamed CD154, was originally considered restricted to activated T lymphocytes, functioning as a mediator of T cell-dependent B cell activation, proliferation, and differentiation. However, the spectrum of CD154 expression and function has broadened considerably during recent years, establishing new roles as a central mediator of immunity and inflammation for this member of the tumor necrosis factor (TNF) gene superfamily. The emerging picture indicates that ligation of the receptor CD40 via CD154, most potently in its trimeric form, functions in two ways. CD154 modulates physiologic processes, such as T cell-mediated effector functions and general immune responses required for appropriate host defense, but also triggers the expression of pro-inflammatory mediators, such as cytokines, adhesion molecules, and matrix degrading activities, all of which are associated with the pathogenesis of chronic inflammatory diseases, e.g., autoimmune disorders, arthritis, atherosclerosis, and cancer. Accordingly, CD40/CD154 interactions have advanced as a potential therapeutic target for these diseases, whereby two opposing strategies, interruption as well as enhancement of CD40 signaling, are explored for beneficial outcomes. PMID- 10856700 TI - A large "footprint" at the boundary of the human beta-globin locus control region hypersensitive site-2. AB - The 5'-boundary region of the human beta-globin locus control region hypersensitive site-2 (HS-2) was examined for protein-DNA interactions. The HS-2 is an erythroid specific DNase I hypersensitive site that extends for approximately 600 bp. Erythroid K562 cells and non-erythroid HeLa cells were damaged by bleomycin and hedamycin--these agents are able to "footprint" nucleosome cores and proteins bound to DNA. The fragments generated by DNA damage were amplified by the ligation-mediated polymerase chain reaction with primers specific for the 5'-boundary region of HS-2 and examined at base pair resolution on DNA sequencing gels. The intensity of damage in intact cells was compared with that in purified DNA. The comparison between intact cells and purified DNA revealed a protected region of 226 bp with bleomycin and 182 bp with hedamycin in K562 cells. The length of the protected region was consistent with the presence of a nucleosome core. We postulate that an erythroid-specific protein binds next to the positioned nucleosome at the boundary of HS-2 to prevent sliding of the nucleosome into the hypersensitive site--this would also account for the large size of the protected region. HeLa cells (lacking a hypersensitive site in the beta-globin cluster) did not have an area of protection in this region. PMID- 10856701 TI - Inhibition of polyamine oxidase enhances the cytotoxicity of polyamine oxidase substrates. A model study with N1-(n-octanesulfonyl)spermine and human colon cancer cells. AB - N(1)-(n-octanesulfonyl)spermine (N(1) OSSpm) is a substrate of polyamine oxidase. It shares several properties with spermine, such as antagonism of NMDA-type glutamate receptors, calmodulin antagonism, and cytotoxicity, but it is more potent by orders of magnitude in these regards than spermine. The human colon carcinoma-derived cell line CaCo-2 was used as a model to study the toxicity of N(1) OSSpm as a function of polyamine oxidase (PAO) activity and differentiation. If the formation of hydrogen peroxide and aminoaldehyde by the PAO-catalysed reactions was prevented by selective inactivation of the enzyme with MDL 72527, cytotoxicity of N(1)OSSpm was not diminished, but on the contrary, enhanced. Exponentially growing CaCo-2 cells were considerably more sensitive to N(1)OSSpm than differentiating cells. The results suggest that cytotoxic substrates of PAO exhibit enhanced cytotoxicity in cells, if PAO activity is inhibited. Since tumour cells are known to have lower polyamine oxidase activities than their normal counterparts, it will be interesting to explore whether cytotoxic substrates of polyamine oxidase, for which N(1)OSSpm is an example, are suited to preferentially kill tumour cells. PMID- 10856702 TI - Kinetics of inhibition of beta-glucosidase from Ampullarium crossean by bromoacetic acid. AB - The kinetics of inhibition of beta-glucosidase from Ampullarium crossean by bromoacetic acid (BrAc) has been studied. The results show that the enzyme can be irreversibly and completely inactivated at high BrAc concentration, while at low BrAc concentration, inhibition of the enzyme is a slow, reversible reaction. The microscopic rate constants for the reactions of BrAc with the enzyme were determined. The presence of the substrate offers obvious protection of the enzyme against inhibition by BrAc. The above results suggest that the histidine residue is essential for activity and is situated at or near the active site of the enzyme. PMID- 10856703 TI - Structural characterization of acid-induced intermediates of human glutathione transferase P1-1. AB - The acid denaturation of human glutathione transferase P1-1 (hGSTP1-1) has been performed to investigate the unfolding intermediates of the protein and their possible involvement in the refolding mechanism. The acid-induced structures of GSTP1-1 have been characterized by activity, gel filtration, intrinsic fluorescence and far-u.v. circular dichroism (CD) techniques. Because of the non identity of the different transitions monitored, the acid denaturation of hGSTP1 1 appears to be a multistep process during which several intermediates coexist in equilibrium. The dependence of inactivation on the protein concentration, as well as gel-filtration experiments, indicate that the inactivation transition, centred at about pH 4.0, corresponds to the monomerization of the protein. At pH 2.0, when the enzyme is completely inactive, the protein retains a small, but significant, amount of secondary structure. This means that the dimeric arrangement of the molecule is important for maintaining the native-like secondary structure of the monomer. The results show that, at low pH, the compact state of the GST monomer, even upon the addition of salts, does not possess native-like secondary structure as described for many monomeric proteins (molten globule). In the presence of physiological concentrations of salts, the protein solution at pH 2.0 leads to a dead-end aggregation process, suggesting that this compact state cannot represent a productive intermediate of the refolding pathway. PMID- 10856704 TI - Analysis of the protein-protein interactions between the human acidic ribosomal P proteins: evaluation by the two hybrid system. AB - The surface acidic ribosomal proteins (P-proteins), together with ribosomal core protein P0 form a multimeric lateral protuberance on the 60 S ribosomal subunit. This structure, also called stalk, is important for efficient translational activity of the ribosome. In order to shed more light on the function of these proteins, we are the first to have precisely analyzed mutual interactions among human P-proteins, employing the two hybrid system. The human acidic ribosomal P proteins, (P1 or P2,) were fused to the GAL4 binding domain (BD) as well as the activation domain (AD), and analyzed in yeast cells. It is concluded that the heterodimeric complex of the P1/P2 proteins is formed preferentially. Formation of homodimers (P1/P1 and P2/P2) can also be observed, though with much less efficiency. Regarding that, we propose to describe the double heterodimeric complex as a protein configuration which forms the 60 S ribosomal stalk: P0 (P1/P2)(2). Additionally, mutual interactions among human and yeast P-proteins were analyzed. Heterodimer formation could be observed between human P2 and yeast P1 proteins. PMID- 10856705 TI - Separation of cathepsin A-like enzyme and the proteasome: evidence that lactacystin/beta-lactone is not a specific inhibitor of the proteasome. AB - Previous studies have described a human platelet cathepsin A-like enzyme with a number of similarities to the "acidic" and "neutral" chymotrypsin-like activities of the proteasome. This includes its strong inhibition by the highly specific proteasome inhibitor Lactacystin/beta-lactone, suggesting that either the Cbz-Phe Ala-hydrolyzing activity attributed to cathepsin A was due to the chymotrypsin like activity of the proteasome or that lactacystin was not a specific inhibitor of the proteasome. In the present study we discard the first possibility on the basis of the following findings: (a) human platelet cathepsin A, unlike proteasome, binds to concanavalin A, and does not bind to Heparin-Sepharose at pH 7.4; (b) neither the chymotrypsin-like activity of the proteasome, nor proteasome antigens are detected in the cathepsin A preparation; (c) purified proteasome does not exhibit Cbz-Phe-Ala-hydrolyzing activity; (d) Z-lle-Glu-(Ot-Bu)Ala leucinal (PSI), a compound that selectively inhibits the chymotrypsin-like activity of the proteasome at a concentration of 10 microM has no inhibitory effect on the carboxypeptidase activity of cathepsin A; (e) cathepsin A, free of the proteasome, is completely inhibited by micromolar concentrations of lactacystin/beta-lactone. It is therefore concluded that lactacystin/beta-lactone is not a specific inhibitor of the proteasome. PMID- 10856706 TI - Increased binding of chlorin e6 to lipoproteins at low pH values. AB - It is well known that the extracellular pH in tumors is lower than that of normal tissue. This has been proposed to be one of the reasons for the tumor selective uptake of several photosensitizers. Photosensitizers like chlorin e(6) are bound to blood components and delivered to different sites in the organism. Thus, the effect of pH on their interaction with human plasma needs to be studied in order to understand a possible role of the acidic microenvironment in tumors for the drug distribution. Increasing amounts of human plasma in the sample resulted in a gradual red shift of the fluorescence emission maxima of chlorin e(6), indicating binding of the drug to some of the plasma components. Titration showed that the drug-plasma interaction was pH-dependent. The titration curve had an inflection point at 7.4+/-0.1. The relative distribution of the drug among plasma components, as found after ultracentrifugation of chlorin e(6)-doped plasma in a salt gradient, showed more binding of the drug to nonlipoproteins than to lipoprotein classes at both pH values studied (6.5 and 7.4). A decrease in the pH was connected with a significant increase in drug-lipoprotein binding. The pH of the environment affects chlorin e(6)-plasma interaction and the distribution of the drug among different plasma components. The results of this study indicate a possible role of the acidic microenvironment in tumors for the preferential uptake and retention of several photosensitiziers. PMID- 10856707 TI - A fatty acid-binding protein and a protein disulphide isomerase-related protein expressed in urochordate gonad cytosol. AB - Despite the evolutionary-tree data suggesting that gene duplication leading to the divergence of the three branches which heart, liver and intestinal fatty acid binding proteins belong to must have occurred before the vertebrate/invertebrate split, only the heart fatty acid-binding protein has been reported for invertebrates. In an attempt to shed light on this apparent inconsistency the presence of the other two branch members was investigated in the Urochordata Molgula pedunculata, an ascidian species close to vertebrates. The mantle-, gonad and digestive tube-cytosolic fractions, obtained by centrifugation at 106,000 g, were incubated separately with [1-(14)C]palmitic acid and then fractionated on a Sephadex G-75 column. In the case of gonads and digestive tube, radioactive peaks corresponding to a molecular mass of 14-16 kDa, characteristic of fatty acid binding proteins, were detected. When the experiment was performed on the mantle, this peak showing fatty acid binding capacity was absent. Western Blot of the radioactive 14-16 kDa Sephadex fraction from the urochordate gonad cross-reacted with rat liver fatty acid-binding protein anti-serum but did not do so with anti rat intestinal, adipocyte or heart fatty acid-binding protein antisera. The material from the digestive tube was not recognized by any of the antisera. The most abundant protein in said 14-16 kDa fraction was a protein disulphide isomerase-related protein. Its partial amino acid sequence was determined. PMID- 10856708 TI - Phospholipase A2-mediated superoxide production of murine peritoneal macrophages induced by chrysotile stimulation. AB - In the present study, we investigated how chrysotile-stimulated macrophages generate superoxide using murine peritoneal macrophages, with special attention to the modulatory role of phospholipase A(2) (PLA(2)). We examined the effects of the following inhibitors and antagonists for signaling molecules on the superoxide anion (O(2)(-)) production of chrysotile-stimulated macrophages: p bromophenacyl bromide (pBPB) and mepacrine for PLA(2); islet-activating protein (IAP) for G-protein; H-7 for protein kinase C (PKC); AA861 for 5-lipoxygenase (5 LO); indomethacin for cyclo-oxygenase (COX); ETYA for both 5-LO and COX; hexanolamine PAF for platelet-activating factor (PAF). The PLA(2) and PKC inhibitors effectively inhibited the chrysotile-induced superoxide anion production of macrophages, but not the G-protein inhibitor, the 5-LO and COX inhibitors, and the PAF antagonist. We also examined the effects of the PLA(2) inhibitors on macrophages stimulated by phorbol 12-myristate 13-acetate (PMA) which directly activates PKC. The two structurally different PLA(2) inhibitors showed differential effects on the PMA-induced superoxide generation: pBPB inhibited it but mepacrine did not. These results suggested that (1) PLA(2) and PKC modulate the chrysotile-induced O(2) production, and (2) two different kinds of PLA(2) work upstream and downstream of PKC, but (3) G-protein, 5-LO and COX metabolites, and PAF have no modulatory role in the reaction. PMID- 10856709 TI - Fatty acid import into mitochondria. AB - The mitochondrial carnitine system plays an obligatory role in beta-oxidation of long-chain fatty acids by catalyzing their transport into the mitochondrial matrix. This transport system consists of the malonyl-CoA sensitive carnitine palmitoyltransferase I (CPT-I) localized in the mitochondrial outer membrane, the carnitine:acylcarnitine translocase, an integral inner membrane protein, and carnitine palmitoyltransferase II localized on the matrix side of the inner membrane. Carnitine palmitoyltransferase I is subject to regulation at the transcriptional level and to acute control by malonyl-CoA. The N-terminal domain of CPT-I is essential for malonyl-CoA inhibition. In liver CPT-I activity is also regulated by changes in the enzyme's sensitivity to malonyl-CoA. As fluctuations in tissue malonyl-CoA content are parallel with changes in acetyl-CoA carboxylase activity, which in turn is under the control of 5'-AMP-activated protein kinase, the CPT-I/malonyl-CoA system is part of a fuel sensing gauge, turning off and on fatty acid oxidation depending on the tissue's energy demand. Additional mechanism(s) of short-term control of CPT-I activity are emerging. One proposed mechanism involves phosphorylation/dephosphorylation dependent direct interaction of cytoskeletal components with the mitochondrial outer membrane or CPT-I. We have proposed that contact sites between the outer and inner mitochondrial membranes form a microenvironment which facilitates the carnitine transport system. In addition, this system includes the long-chain acyl-CoA synthetase and porin as components. PMID- 10856710 TI - Transport of fatty acids and metabolites across the peroxisomal membrane. AB - The peroxisomal membrane forms a permeability barrier for a wide variety of metabolites required for and formed during fatty acid beta-oxidation. To communicate with the cytoplasm and mitochondria, peroxisomes need dedicated proteins to transport such hydrophilic molecules across their membranes. Genetic and biochemical studies in the yeast Saccharomyces cerevisiae have identified enzymes for redox shuttles as well as the first peroxisomal membrane transporter. This peroxisomal ATP-binding cassette transporter (Pat) is highly homologous to the gene mutated in X-linked adrenoleukodystrophy (X-ALD). The yeast Pat is required for import of activated fatty acids into peroxisomes suggesting that this is the primary defect in X-ALD. PMID- 10856711 TI - The fatty acid transport function of fatty acid-binding proteins. AB - The intracellular fatty acid-binding proteins (FABPs) comprise a family of 14-15 kDa proteins which bind long-chain fatty acids. A role for FABPs in fatty acid transport has been hypothesized for several decades, and the accumulated indirect and correlative evidence is largely supportive of this proposed function. In recent years, a number of experimental approaches which more directly examine the transport function of FABPs have been taken. These include molecular level in vitro modeling of fatty acid transfer mechanisms, whole cell studies of fatty acid uptake and intracellular transfer following genetic manipulation of FABP type and amount, and an examination of cells and tissues from animals engineered to lack expression of specific FABPs. Collectively, data from these studies have provided strong support for defining the FABPs as fatty acid transport proteins. Further studies are necessary to elucidate the fundamental mechanisms by which cellular fatty acid trafficking is modulated by the FABPs. PMID- 10856712 TI - Sterol carrier protein-2. AB - The compartmentalization of cholesterol metabolism implies target-specific cholesterol trafficking between the endoplasmic reticulum, plasma membrane, lysosomes, mitochondria and peroxisomes. One hypothesis has been that sterol carrier protein-2 (SCP2, also known as the non-specific lipid transfer protein) acts in cholesterol transport through the cytoplasm. Recent studies employing gene targeting in mice showed, however, that mice lacking SCP2 and the related putative sterol carrier known as SCPx, develop a defect in peroxisomal beta oxidation. In addition, diminished peroxisomal alpha-oxidation of phytanic acid (3,7,11, 15-tetramethylhexadecanoic acid) in these null mice was attributed to the absence of SCP2 which has a number of properties supporting a function as carrier for fatty acyl-CoAs rather than for sterols. PMID- 10856713 TI - Phosphatidylinositol/phosphatidylcholine transfer proteins in yeast. AB - Phosphatidylinositol transfer proteins (PITPs) are now becoming widely recognized as intriguing proteins that participate in the coordination and coupling of phospholipid metabolism with vesicle trafficking, and in the regulation of important signaling cascades. Yet, only in one case is there a large body of evidence that speaks to the precise identities of PITP-dependent cellular reactions, and to the mechanisms by which PITPs execute function in eukaryotic cells. At present, yeast provide the most powerful system for analysis of the physiology of PITP function in vivo, and the mechanism by which this function is carried out. Here, we review the recent progress and remaining questions in the area of PITP function in yeast. PMID- 10856714 TI - Progress towards understanding the role of microsomal triglyceride transfer protein in apolipoprotein-B lipoprotein assembly. AB - The microsomal triglyceride transfer protein (MTP) is necessary for the proper assembly of the apolipoprotein B containing lipoproteins, very low density lipoprotein and chylomicrons. Recent research has significantly advanced our understanding of the role of MTP in these pathways at the molecular and cellular level. Biochemical studies suggest that initiation of lipidation of the nascent apolipoprotein B polypeptide may occur through a direct association with MTP. This early lipidation may be required to allow the nascent polypeptide to fold properly and therefore avoid ubiquitination and degradation. Concerning the addition of core neutral lipids in the later stages of lipoprotein assembly, cell culture studies show that MTP lipid transfer activity is not required for this to occur for apolipoprotein B-100 containing lipoproteins. Likewise, MTP does not appear to directly mediate addition of core neutral lipid to nascent apoB-48 particles. However, new data indicate that MTP is required to produce triglyceride rich droplets in the smooth endoplasmic reticulum which may supply the core lipids for conversion of nascent, dense apoB-48 particles to mature VLDL. In addition, assembly of dense apolipoprotein B-48 containing lipoproteins has been observed in mouse liver in the absence of MTP. As a result of these new data, an updated model for the role of MTP in lipoprotein assembly is proposed. PMID- 10856715 TI - The synthesis and transport of lipids for axonal growth and nerve regeneration. AB - Neurons are unique polarized cells in which the growing axon is often located up to a meter or more from the cell body. Consequently, the intracellular movement of membrane lipids and proteins between cell bodies and axons poses a special challenge. The mechanisms of lipid transport within neurons are, for the most part, unknown although lipid transport via vesicles and via cholesterol- and sphingolipid-rich 'rafts' are considered likely mechanisms. Very active anterograde and retrograde transport of lipid-containing vesicles occurs between the cell body and distal axons. However, it is becoming clear that the axon need not obtain all of its membrane constituents from the cell body. For example, the synthesis of phosphatidylcholine, the major membrane phospholipid, occurs in axons, and its synthesis at this location is required for axonal elongation. In contrast, cholesterol synthesis appears to occur only in cell bodies, and cholesterol is efficiently delivered from cell bodies to axons by anterograde transport. Cholesterol that is required for axonal growth can also be exogenously supplied from lipoproteins to axons of cultured neurons. Several studies have suggested a role for apolipoprotein E in lipid delivery for growth and regeneration of axons after a nerve injury. Alternatively, or in addition, apolipoprotein E has been proposed to be a ligand for receptors that mediate signal transduction cascades. Lipids are also transported from axons to myelin, although the importance of this process for myelination is not clear. PMID- 10856716 TI - Interorganelle transport of aminoglycerophospholipids. AB - The aminoglycerophospholipids of eukaryotic cells, phosphatidylserine (PtdSer), phosphatidylethanolamine (PtdEtn), and phosphatidylcholine (PtdCho), can be synthesized by multiple pathways. The PtdSer pathway encompasses the synthesis of PtdSer, its decarboxylation to PtdEtn and subsequent methylation reactions to form PtdCho. The Kennedy pathways consist of the synthesis of PtdEtn and PtdCho from Etn and Cho precursors via CDP-Etn and CDP-Cho intermediates. The reactions along the PtdSer pathway are spatially segregated with PtdSer synthesis occurring in the endoplasmic reticulum or mitochondria-associated membrane (MAM), PtdEtn formation occurring in the mitochondria and Golgi/vacuole compartments and PtdCho formation occurring in the endoplasmic reticulum or MAM. The organelle-specific metabolism of the different lipids in the PtdSer pathway has provided a convenient biochemical means for defining events in the interorganelle transport of the aminoglycerophospholipids in intact cells, isolated organelles and permeabilized cells. Studies with both mammalian cells and yeast demonstrate many significant similarities in lipid transport processes between the two systems. Genetic experiments in yeast now provide the tools to create new strains with mutations along the PtdSer pathway that can be conditionally rescued by the Kennedy pathway reactions. The genetic studies in yeast indicate that it is now possible to begin to define genes that participate in the interorganelle transport of the aminoglycerophospholipids. PMID- 10856717 TI - Identification and purification of aminophospholipid flippases. AB - Transbilayer phospholipid asymmetry is a common structural feature of most biological membranes. This organization of lipids is generated and maintained by a number of phospholipid transporters that vary in lipid specificity, energy requirements and direction of transport. These transporters can be divided into three classes: (1) bidirectional, non-energy dependent 'scramblases', and energy dependent transporters that move lipids (2) toward ('flippases') or (3) away from ('floppases') the cytofacial surface of the membrane. One of the more elusive members of this family is the plasma membrane aminophospholipid flippase, which selectively transports phosphatidylserine from the external to the cytofacial monolayer of the plasma membrane. This review summarizes the characteristics of aminophospholipid flippase activity in intact cells and describes current strategies to identify and isolate this protein. The biochemical characteristics of candidate flippases are critically compared and their potential role in flippase activity is evaluated. PMID- 10856718 TI - ABC transporters in lipid transport. AB - Since it was found that the P-glycoproteins encoded by the MDR3 (MDR2) gene in humans and the Mdr2 gene in mice are primarily phosphatidylcholine translocators, there has been increasing interest in the possibility that other ATP binding cassette (ABC) transporters are involved in lipid transport. The evidence reviewed here shows that the MDR1 P-glycoprotein and the multidrug resistance ( associated) transporter 1 (MRP1) are able to transport lipid analogues, but probably not major natural membrane lipids. Both transporters can transport a wide range of hydrophobic drugs and may see lipid analogues as just another drug. The MDR3 gene probably arose in evolution from a drug-transporting P-glycoprotein gene. Recent work has shown that the phosphatidylcholine translocator has retained significant drug transport activity and that this transport is inhibited by inhibitors of drug-transporting P-glycoproteins. Whether the phosphatidylcholine translocator also functions as a transporter of some drugs in vivo remains to be seen. Three other ABC transporters were recently shown to be involved in lipid transport: ABCR, also called Rim protein, was shown to be defective in Stargardt's macular dystrophy; this protein probably transports a complex of retinaldehyde and phosphatidylethanolamine in the retina of the eye. ABC1 was shown to be essential for the exit of cholesterol from cells and is probably a cholesterol transporter. A third example, the ABC transporter involved in the import of long-chain fatty acids into peroxisomes, is discussed in the chapter by Hettema and Tabak in this volume. PMID- 10856719 TI - Sphingolipid transport in eukaryotic cells. AB - Sphingolipids constitute a sizeable fraction of the membrane lipids in all eukaryotes and are indispensable for eukaryotic life. First of all, the involvement of sphingolipids in organizing the lateral domain structure of membranes appears essential for processes like protein sorting and membrane signaling. In addition, recognition events between complex glycosphingolipids and glycoproteins are thought to be required for tissue differentiation in higher eukaryotes and for other specific cell interactions. Finally, upon certain stimuli like stress or receptor activation, sphingolipids give rise to a variety of second messengers with effects on cellular homeostasis. All sphingolipid actions are governed by their local concentration. The intricate control of their intracellular topology by the proteins responsible for their synthesis, hydrolysis and intracellular transport is the topic of this review. PMID- 10856720 TI - Intracellular cholesterol trafficking: role of the NPC1 protein. PMID- 10856721 TI - Intramitochondrial cholesterol transfer. AB - Cholesterol serves as the initial substrate for all steroid hormones synthesized in the body regardless of the steroidogenic tissue or final steroid produced. The first steroid formed in the steroidogenic pathway is pregnenolone which is formed by the excision of a six carbon unit from cholesterol by the cytochrome P450 side chain cleavage enzyme system which is located in the inner mitochondrial membrane. It has long been known that the regulated biosynthesis of steroids is controlled by a cycloheximide sensitive factor whose function is to transfer cholesterol from the outer to the inner mitochondrial membrane, thus, the identity of this factor is of great importance. A candidate for the regulatory factor is the mitochondrial protein, the steroidogenic acute regulatory (StAR) protein. Cloning and sequencing of the StAR cDNA indicated that it was a novel protein, and transient transfections with the cDNA for the StAR protein resulted in increased steroid production in the absence of stimulation. Mutations in the StAR gene cause the potentially lethal disease congenital lipoid adrenal hyperplasia, a condition in which cholesterol transfer to the cytochrome P450 side chain cleavage enzyme, P450scc, is blocked, filling the cell with cholesterol and cholesterol esters. StAR knockout mice have a phenotype which is essentially identical to the human condition. The cholesterol transferring activity of StAR has been shown to reside in the C-terminal part of the molecule and a protein sharing homology with a region in the C-terminus of StAR has been shown to display cholesterol transferring capacity. Recent evidence has indicated that StAR can act as a sterol transfer protein and it is perhaps this characteristic which allows it to mobilize cholesterol to the inner mitochondrial membrane. However, while it appears that StAR is the acute regulator of steroid biosynthesis via its cholesterol transferring activity, its mechanism of action remains unknown. PMID- 10856722 TI - Intracellular transport of bile acids. AB - Bile acids originate from the liver and are transported via bile to the intestines where they perform an important role in the absorption of lipids and lipid-soluble nutrients. Most of the bile acids are reclaimed from the terminal ileum and returned to the liver via portal blood for reuse. The transport of bile acids is vectorial in both liver and intestinal cells, originating and terminating at opposite poles. Bile acids enter through the basolateral pole in liver cells, and through the apical pole in intestinal cells. During the past decade, much has been learned about the mechanisms by which bile acids enter and exit liver and intestinal cells. By contrast, the mechanisms by which bile acids are transported across cells remain poorly understood. The current body of evidence suggests that bile acids do not traverse the cell by vesicular transport. Although a carrier-mediated mechanism is a likely alternative, only a handful of intracellular proteins capable of binding bile acids have been described. The significance of these proteins in the intracellular transport of bile acids remains to be tested. PMID- 10856724 TI - The Euro Heart Failure Survey of the EUROHEART survey programme. A survey on the quality of care among patients with heart failure in Europe. The Study Group on Diagnosis of the Working Group on Heart Failure of the European Society of Cardiology. The Medicines Evaluation Group Centre for Health Economics University of York. AB - BACKGROUND: The EUROHEART programme is a rolling programme of cardiovascular surveys among the member nations of the European Society of Cardiology (ESC). These surveys will provide information on the nature of cardiovascular disease and its management. This manuscript describes a survey into the nature and management of heart failure. AIMS: The EuroHeart Failure survey aims to describe the quality of hospital care, diagnostic and therapeutic, for patients with suspected or confirmed heart failure in ESC member countries. Patients will be interviewed subsequent to hospital discharge to assess their understanding of their condition, side effects from and their compliance with therapy and their satisfaction with the management for heart failure. The quality of management will be judged against the recommendations contained in the ESC guidelines on diagnosis and treatment of heart failure. Outcome will be further assessed by repeat interviews in 6-12 months time. A further survey of heart failure in 2001/2002 is also planned. METHODS: A prospective survey of all deaths and discharges from medical (cardiology, internal medicine and geriatric medicine) and cardiac surgical wards to identify patients with heart failure, suspected or confirmed. Approximately 70 hospital clusters, comprising two to six hospitals in each cluster, in 24 member countries of the ESC are conducting the study. At the time of writing, approximately 30000 deaths and discharges have been screened and approximately 4000 patients have been enrolled. CONCLUSIONS: The EuroHeart Survey will allow actual practice to be compared to ESC guidelines on the diagnosis and treatment of heart failure. The surveys and guidelines should prove mutually informative. The main EuroHeart Failure project will be completed by late 2000. However, new centres volunteering to participate in the study (contact corresponding author) may be accepted providing they have the necessary research personnel and provided funding can be agreed for statistical support and administration. PMID- 10856725 TI - Heart failure in the elderly: a diastolic problem? PMID- 10856726 TI - Long-term prognosis of acute pulmonary oedema--an ominous outcome. AB - BACKGROUND: Acute pulmonary oedema (APOE) is a major health problem, leading to poor hospital and long-term outcomes. There is a relative paucity of studies describing prognosis of consecutive unsolicited patients diagnosed with APOE and hospitalized in internal medicine departments. AIMS: To describe the clinical profile and outcome (in hospital and 1-year prognosis) of successive unselected patients with APOE, in a prospective observational study. METHODS AND RESULTS: The study population included 150 consecutive unsolicited patients (90 men, 60 women; median age 75 years) with APOE all hospitalized in an internal medicine department, in a 900-bed care centre. Ischaemic heart disease (IHD), hypertension and diabetes were present in 85%, 70% and 52% of patients, respectively. The most common precipitating factors for APOE included high blood pressure (29%), rapid atrial fibrillation (29%), unstable angina pectoris (25%), infection (18%) and acute myocardial infarction (MI; 15%). Eighteen patients (12%) died in hospital, with 82% of these deaths attributed to cardiac pump failure. Predictors for an increased in-hospital mortality included: diabetes (P<0.05), orthopnoea (P<0. 05), echocardiographic finding of depressed global left ventricular systolic function (P<0.001), acute MI during hospital stay (P<0.001), hypotension/shock (P<0.05), and the need for mechanical ventilation (P<0.001). After a median hospital stay of 10 days, 132 patients were discharged home. The 1-year mortality was 40%. Only the presence of pleural effusion was found as a predictor for 1 year mortality. CONCLUSION: Most patients with APOE in this study are elderly, and have IHD, hypertension, diabetes and a previous history of APOE. The overall mortality is high (in-hospital, 12%: 1-year, 40%). Left ventricular dysfunction was associated with high in-hospital mortality, but not with long-term prognosis. PMID- 10856727 TI - Cardiovascular disease and outcome of acute stroke: influence of pre-existing cardiac failure. AB - BACKGROUND AND AIMS: Whilst a number of variables, mostly a consequence of a stroke, are known to predict mortality of acute stroke there is limited information on the significance of pre-existing cardiovascular variables on stroke mortality. We have investigated the influence of pre-existing cardiovascular factors in one cohort of stroke patients. METHODS: We studied 295 patients, mean age 74+/-10 (range 34-96) years; 133 males, presenting with acute stroke for pre-existing cardiovascular disease (CVD) defined as hypertension, atrial fibrillation (AF), ischaemic heart disease (IHD) and cardiac failure (CF). In addition, data were collected on epidemiological and neurological variables known to influence stroke mortality. The most significant of the cardiovascular factors was further investigated against all the other cardiovascular groups together and against those without any CVD. Outcome was measured as their influence on acute phase and 3-month mortality. RESULTS: There was no significant difference in 3-month mortality with hypertension (P=0.62) and IHD (P=0.33) but there was a significant higher mortality in patients with AF (P=0.05) and CF (P <0.001). CF was more significant than all other CVD (hypertension+AF+IHD) together without the failure (P<0.001); odds ratio of 4.5 (95% CI 2.28-9.07). Partial correlation coefficient revealed CF to be an independent significant variable to influence stroke mortality when controlled with AF, stroke syndromes, age, incontinence, pyrexia, dysphagia and Glasgow coma score. CONCLUSIONS: Pre existing CF has an adverse influence on stroke mortality independent of other known factors. Cardiovascular factors without failure do not have such an effect except the marginal effect of AF. PMID- 10856728 TI - Doppler tissue imaging in congestive heart failure patients due to diastolic or systolic dysfunction: a comparison with Doppler echocardiography and the atrio ventricular plane displacement technique. AB - BACKGROUND: Doppler tissue imaging (DTI) is an echocardiographic technique by which regional contractility, relaxation properties and time intervals are obtained easily. DTI has been reported to be relatively pre-load independent and could, in comparison with the commonly used mitral pulse wave Doppler (MPWD) method, be of clinical interest for identification of patients with diastolic dysfunction. The atrio-ventricular plane displacement (AVPD) method is an established technique to assess left ventricular systolic function. AIMS: To determine the pulsed Doppler DTI-pattern in patients with heart failure and to examine whether it has a similar capacity as MPWD and AVPD to diagnose diastolic dysfunction. METHODS: We studied 15 controls without congestive heart failure (CHF), 15 patients with diastolic (EF>45%+CHF) and 15 patients with systolic (EF<35%+CHF) left ventricular dysfunction and CHF. RESULTS: The DTI maximal velocities during systole (s), early filling wave (e) and atrial filling wave (a), decrease with reduced left ventricular ejection fraction, r=0.75, r=0.56 and r=0.66 (P<0.001) and regional isovolumetric contraction and intraventricular relaxation time measured by DTI are prolonged, r=0.59 and r=0.73, respectively (P<0.001). The 15 patients with diastolic heart failure were identified by MPWD or DTI but only 11 by AVPD with 8, 10 and 9 false-positive, respectively (P<0.01, P<0.05 and NS). CONCLUSIONS: Regional DTI show a consistent pattern in patients with left ventricular dysfunction and heart failure. Regional DTI has similar accuracy as MPWD in identifying diastolic heart failure patients and is superior to the AVPD technique. DTI may be a useful diagnostic tool in diastolic heart failure patients. PMID- 10856729 TI - Serum N-terminal atrial natriuretic peptide in adult patients late after surgical repair of atrial septal defect. AB - BACKGROUND: The purpose of surgical closure of atrial septal defect (ASD) is to relieve the cardiovascular system from a haemodynamic burden. Excessive amounts of atrial peptides are released in congestive heart failure, valvular diseases and congenital heart diseases. AIMS: To examine whether patients after surgical repair of ASD have higher concentrations of N-terminal atrial natriuretic peptide (ANP-N) than age-, sex- and body mass index (BMI)-matched control subjects. METHODS: Medical history, physical examination, standard 12-lead electrocardiogram, and ANP-N concentrations were obtained in 65 adult patients operated for ASD at the age of 21+/-13 years (mean+/-standard deviation), 21+/-6 years after surgical closure of ASD. Sixty-seven healthy subjects matched for age, sex and BMI served as controls. RESULTS: In the patients serum ANP-N was higher than in the control subjects 0.41+/-0.32 nmol/l, median 0.31 nmol/l, interquartile range (IQR) 0.21-0.49 nmol/l vs. 0.24+/-0.12 nmol/l, median 0.23 nmol/l, IQR 0.17-0.29 nmol/l, (P=0.0003). Patients with concomitant diseases had higher ANP-N concentrations (0.51+/-0.39 nmol/l, median 0.34, IQR 0.26-0.73 nmol/l) than ASD patients without any history or signs of disease (0.28+/-0.16 nmol/l, median 0.27, IQR 0.17-0.40 nmol/l, P=0.01). The 'healthy' ASD patients had higher hormone concentrations than age-, sex- and BMI-matched control subjects (0.28+/-0.16 median 0.27 nmol/l, IQR 0. 17-0.40 nmol/l and 0.21+/-0.07 nmol/l, median 0.20 nmol/l, IQR 0. 15-0.27 nmol/l, P=0.01). Multiple stepwise linear regression analysis showed that age at operation was strongly associated with the post-operative ANP-N concentration (r(2)=0.25, P=0.0002). CONCLUSION: ASD patients have higher ANP-N concentrations late after surgical repair. Hormone levels correlate with age at operation. Our finding supports the clinical praxis of operating on these patients in their childhood and adolescence. PMID- 10856730 TI - Negative inotropic effects of recombinant interleukin 2 in patients without left ventricular dysfunction. AB - Experimental data have shown that rIL2 has negative inotropic properties. This has not been investigated in humans with normal left ventricular function. Seventeen consecutive renal cell carcinoma patients who received rIL2 therapy because of dissemination were analyzed before and after treatment with a low dose of rIL2 subcutaneously. Left ventricular ejection fraction (echocardiography), heart rate variability parameters (24 h electrocardiography), and TNF alpha, IL1 beta and nitric oxide metabolites (NO(x)) were measured. LVEF decreased from 54+/ 7 to 50+/-6% (mean+/-S.D.; P=0.012), with a concomitant increase in heart rate from 87+/-13 to 94+/-13 beats/min (P=0.031). All frequency domain HRV parameters decreased: the total power from 18.0+/-7.9 to 14.0+/-5.0 ms (P=0.001), the low frequency from 10.3+/-5.4 to 8. 3+/-3.4 ms (P=0.001), and the high frequency from 6.3+/-2.6 to 4. 5+/-1.1 ms (P=0.001). There was no measurable effect on TNF alpha, IL1 beta concentrations. Plasma levels of nitrate (NO(x)) increased from 22.8+/-14.4 to 41.8+/-26.6 micromol/l (P=0.007). CONCLUSIONS: A low dose of rIL2 has a negative inotropic effect that may be mediated by increased NO concentrations. It also reduces sympathetic activity as reflected in HRV parameters. PMID- 10856731 TI - MIC trial: metoprolol in patients with mild to moderate heart failure: effects on ventricular function and cardiopulmonary exercise testing. AB - Beta-blocker therapy results in a functional benefit in patients with heart failure (CHF) due to idiopathic dilated cardiomyopathy (DCM). This study assessed if similar effects were observed in patients with ischemic heart disease (CAD), NYHA II-III after 6 months of therapy with metoprolol. METHODS AND RESULTS: Fifty two patients with CHF secondary to DCM (26 patients) and CAD (26 patients) and a left ventricular ejection fraction (EF)<40% were enrolled in the placebo controlled study. The study medication was titrated over 6 weeks, the mean final dosage was 135 mg/day. Three patients died due to cardiogenic shock, two received placebo and one metoprolol. Eight patients did not complete the study due to non compliance. Metoprolol significantly reduced heart rate at rest and after submaximal and maximal exercise. Vo(2)-max and Vo(2)-AT as well as the 6-min walk test improved significantly after metoprolol treatment. There was a significant increase in EF at rest (27.3-35. 2%), submaximal (28.5-37.7%) and maximal exercise (28.7-40.9%) in the metoprolol-treated patients. No differences were found between patients with CAD and DCM. We also observed reduced left ventricular volumes. CONCLUSION: The additional therapy with metoprolol improved cardiac function and the cardiopulmonary exercise capacity in patients with CHF. PMID- 10856732 TI - Improvement of cardiac output in patients with severe heart failure by use of ACE inhibitors combined with the AT1-antagonist eprosartan. AB - BACKGROUND: The efficacy of ACE-inhibitor therapy is well documented in the treatment of chronic heart failure. As pharmacological mechanisms of ACE inhibition and angiotensin II AT1-receptor-antagonists differ, an additional positive effect concerning left ventricular function can be expected in combining both classes of drugs. METHODS: Twenty patients (64.9+/-8.5 years) with advanced chronic heart failure (NYHA class III) receiving long-term medication with digitalis, diuretics and ACE-inhibitors were randomized to either eprosartan (540+/-96 mg/day) or placebo, according to a blinded protocol. Hemodynamic measurements by impedance cardiography were performed at baseline and after 8.85+/-1. 5 days of study medication treatment. RESULTS: Additional treatment with eprosartan resulted in a higher cardiac output than in the control group (P<0.05). While in the active treatment group cardiac output increased significantly from baseline (2.27-3.24 l/min, P=0. 039), there was no change in the control group. CONCLUSIONS: The additional treatment with the AT1-receptor antagonist eprosartan, given to severe heart failure patients, who received digitalis, diuretics and ACE-inhibitors, resulted in a beneficial effect by increasing cardiac output. This effect may be due to eprosartan's additional property of blocking the autocrine interaction of locally and not ACE-generated angiotensin II with their respective vascular and myocardial AT1-receptors as well as the influence on prejunctional AT1-receptors located on sympathetic nerve terminals. PMID- 10856733 TI - The effects of L-carnitine treatment on left ventricular function and erythrocyte superoxide dismutase activity in patients with ischemic cardiomyopathy. AB - We studied the effects of L-carnitine on left ventricular systolic function and the erythrocyte superoxide dismutase activity in 51 patients with ischemic cardiomyopathy. They all previously were under the treatment of angiotensin converting enzyme inhibitor, digitalis and diuretics. Patients were randomized into two groups. In group I (n=31), 2 g/day L-carnitine was added to therapy. L Carnitine was not given to the other 20 patients (Group II). In group I (mean age 64.3+/-7.8 years), 27 of the patients were men, and four were women. In group II (mean age 66.2+/-8.7 years), 17 of the patients were men, and three were women. Twenty age-matched healthy subjects (mean age: 60.1+/-5.3 years) constituted the control group. In each group, left ventricular ejection fraction (LVEF) by echocardiography and red cell superoxide dismutase activity by spectrophotometric method were measured initially and after 1 month of randomisation. Compared with normal healthy subjects (n=20), patients (n=51) had significantly higher red cell SOD activity (5633+/-1225 vs. 3202+/-373 U/g Hb, P<0.001). At the end of 1 month of L-carnitine therapy, red cell SOD activity showed an increase in group I (5918+/-1448 to 7218+/-1917 U/g Hb, P<0.05). In group II, red cell SOD activity showed no significant change after 1 month of randomisation (5190+/-545 to 5234+/ 487 U/g Hb, P=0. 256). One month after randomisation there was a significant increase in LVEF in both groups I and II (37.8-42.3%, P<0.001 in group I; 41. 5 43.8%, P<0.001 in group II). The improvement in LVEF was more significant in the L-carnitine group (4.5% vs. 2.3%, P<0.01). We conclude that, as a sign of increased free radical production, superoxide dismutase activity was further increased in patients with L-carnitine treatment. L-Carnitine treatment in combination with other traditional pharmacological therapy might have an additive effect for the improvement of left ventricular function in ischemic cardiomyopathy. PMID- 10856734 TI - Left ventricular-based pacing in patients with chronic heart failure: comparison of acute hemodynamic benefits according to underlying heart disease. AB - BACKGROUND: Acute left ventricular-based pacing has been shown to improve hemodynamics in patients with severe heart failure and left bundle branch block (LBBB). However, it is not known whether the cause of the underlying heart disease influences the potential effect of left ventricular-based pacing. OBJECTIVES: The aim of this study was to determine whether beneficial hemodynamic effects of acute left ventricular-based pacing in severe chronic heart failure are dependent on underlying heart disease. METHODS: After coronary angiography, patients with severe heart failure and LBBB were separated into two groups: dilated (25 patients; 20 male) and ischemic cardiomyopathy (21 patients; 20 male). Hemodynamic parameters were evaluated at baseline and during left ventricular-based pacing. RESULTS: Improvement in hemodynamic parameters were similar in both groups, during acute left ventricular pacing (changes expressed in percentage): pulmonary capillary wedge pressure, -16+/-15% vs. -14+/-10%; V wave amplitude, -25+/-18% vs. -21+/-17%; and biventricular pacing, -15+/-15% vs. 11+/-11% and -23+/-18% vs. -16+/-18%, respectively. CONCLUSION: Underlying heart disease does not influence the response to acute left ventricular-based pacing in patients with severe heart failure and LBBB. This finding provides support for including all patients with enlarged heart and heart failure in future studies evaluating left ventricular-based pacing. PMID- 10856735 TI - Angiotensin converting enzyme (ACE) inhibitors in the treatment of heart failure in general practice in north Cumbria. AB - BACKGROUND: A great deal of research has demonstrated the benefits of treating patients with chronic heart failure with Angiotensin Converting Enzyme (ACE) inhibitors. There is rather less research on the actual uptake of treatment in general practice, and in particular methods that might improve that uptake. AIM: To study the attitudes and practice of medical practitioners in North Cumbria in the treatment of heart failure. METHOD: Semi-structured interviews with 16 general practitioners and nine hospital physicians in the Carlisle area and an audit of general practice case notes. RESULTS: Two hundred and fifty-eight patients were identified with heart failure. Prevalence was 1.1%. Fifty percent were on an ACE inhibitor, the mean dose of which was less than half the typical research dose. Patients who had an echocardiogram were much more likely to be on an ACE inhibitor. General practitioners were enthusiastic to use ACE inhibitors, but felt that greater access to echocardiography was required. Hospital physicians were happy to improve access within an agreed protocol. CONCLUSION: Improved uptake of ACE inhibitors could be assisted by the development of a protocol for investigation and treatment. This protocol should be evidence-based and agreed between local GPs, hospital physicians and the Health Authority. PMID- 10856736 TI - Clinical trials update: OPTIME-CHF, PRAISE-2, ALL-HAT. AB - This is a summary of reports of presentation made at the American College of Cardiology 49th Scientific Sessions, Anaheim, 12-15 March 2000. Studies with a particular interest for heart failure physicians have been reviewed. OPTIME-CHF: Outcomes of a Prospective Trial of Intravenous Milrinone for Exacerbations of Chronic Heart Failure. OPTIME-CHF was a randomised-controlled trial comparing a 48-h infusion of Milrinone or standard therapy in 951 patients recruited over a 2 year period. Patients were excluded if the investigator believed their clinical condition mandated inotropic therapy. Patients were randomised within 48 h of admission for an acute exacerbation of chronic heart failure to receive Milrinone or placebo infision for 48 h. Of the patients 43% were diabetics, 70% were receiving an angiotensin converting enzyme inhibitor, 25% were already on a beta Blocker, and 34% had atrial fibrillation. There was no significant difference between the two groups in length of hospital stay during the index admission, subsequent readmissions and days in hospital over the following 60 days. Subjective clinical assessment scores were also no different. There was an average admission rate over the next year of one per patient in both groups. However, there was a significant increase in the incidence of sustained hypotension in the Milrinone group, which accounted for all of the increased adverse event rates for the active therapy. The 60-day mortality was 10% in both groups. This and previous trials of the oral formulation of Milrinone have now clearly demonstrated a lack of benefit with Milrinone in either during acute exacerbations of or in stable severe chronic heart failure [Packer M, Carver JR, Rodeheffer RJ et al. Effect of oral Milrinone on mortality in severe chronic heart failure. N Engl J Med 1991;325:1468-1475.]. Medium sized studies of Milrinone in patients with milder severities of heart failure also suggested an adverse impact on prognosis in the presence or absence of digoxin [DiBianco R, Shabetai R, Kostuk W, Moran J, Schlant RC, Wright R. A comparison of oral Milrinone, digoxin, and their combination in the treatment of patients with chronic heart failure. N Engl J Med 1989;320:677-683.]. Whether Milrinone even has a role for the management of a haemodyamic crisis requiring inotropic therapy must also be questioned. PMID- 10856738 TI - The management of patients with heart failure in France. AB - Heart failure is becoming, in France as in other countries, a major public health problem, occurring in elderly patients with concurrent disease. A general overview of the current care of heart failure patients in France is provided in this article, and some ideas about possible ways of improving the present management of heart failure. PMID- 10856739 TI - Young children are sensitive to how an object was created when deciding what to name it. AB - How do young children extend names for human-made artifacts, such as knife, toy, and painting? We addressed this issue by showing 3-year-olds, 5-year-olds, and adults a series of simple objects and asking them for each, 'What is this?' In one condition, the objects were described as purposefully created; in another, the objects were described as being created by accident. This manipulation had a significant effect on the participants' responses: even 3-year-olds were more likely to provide artifact names (e.g. 'a knife') when they believed the objects were intentionally created and material-based descriptions (e.g. 'plastic') when they believed the objects were accidentally created. This result supports a theory of artifact naming in which intuitions about intention play an important role. PMID- 10856740 TI - Caricaturing facial expressions. AB - The physical differences between facial expressions (e.g. fear) and a reference norm (e.g. a neutral expression) were altered to produce photographic-quality caricatures. In Experiment 1, participants rated caricatures of fear, happiness and sadness for their intensity of these three emotions; a second group of participants rated how 'face-like' the caricatures appeared. With increasing levels of exaggeration the caricatures were rated as more emotionally intense, but less 'face-like'. Experiment 2 demonstrated a similar relationship between emotional intensity and level of caricature for six different facial expressions. Experiments 3 and 4 compared intensity ratings of facial expression caricatures prepared relative to a selection of reference norms - a neutral expression, an average expression, or a different facial expression (e.g. anger caricatured relative to fear). Each norm produced a linear relationship between caricature and rated intensity of emotion; this finding is inconsistent with two-dimensional models of the perceptual representation of facial expression. An exemplar-based multidimensional model is proposed as an alternative account. PMID- 10856741 TI - Spoken word recognition and lexical representation in very young children. AB - Although children's knowledge of the sound patterns of words has been a focus of debate for many years, little is known about the lexical representations very young children use in word recognition. In particular, researchers have questioned the degree of specificity encoded in early lexical representations. The current study addressed this issue by presenting 18-23-month-olds with object labels that were either correctly pronounced, or mispronounced. Mispronunciations involved replacement of one segment with a similar segment, as in 'baby-vaby'. Children heard sentences containing these words while viewing two pictures, one of which was the referent of the sentence. Analyses of children's eye movements showed that children recognized the spoken words in both conditions, but that recognition was significantly poorer when words were mispronounced. The effects of mispronunciation on recognition were unrelated to age or to spoken vocabulary size. The results suggest that children's representations of familiar words are phonetically well-specified, and that this specification may not be a consequence of the need to differentiate similar words in production. PMID- 10856742 TI - Illusory inferences from a disjunction of conditionals: a new mental models account. AB - (Johnson-Laird, P.N., & Savary, F. (1999, Illusory inferences: a novel class of erroneous deductions. Cognition, 71, 191-229.) have recently presented a mental models account, based on the so-called principle of truth, for the occurrence of inferences that are compelling but invalid. This article presents an alternative account of the illusory inferences resulting from a disjunction of conditionals. In accordance with our modified theory of mental models of the conditional, we show that the way individuals represent conditionals leads them to misinterpret the locus of the disjunction and prevents them from drawing conclusions from a false conditional, thus accounting for the compelling character of the illusory inference. PMID- 10856743 TI - Illusions and models: a reply to Barrouillet and Lecas. PMID- 10856744 TI - Goal attribution in chimpanzees. AB - Does the chimpanzee attribute goals to others? Recent infant studies using the looking time measure have been interpreted as evidence that human infants attribute goals. An experiment modeled on these studies was carried out on chimpanzees, and the chimpanzees responded the way infants do. This indicates that chimpanzees also attribute goals and hence that this capacity is not distinctively human. PMID- 10856745 TI - Causal status effect in children's categorization. AB - The current study examined the causal status effect (weighing cause features more than effect features in categorization) in children. Adults (Study 1) and 7-9 year-old children (Study 2) learned descriptions of novel animals, in which one feature caused two other features. When asked to determine which transfer item was more likely to be an example of the animal they had learned, both adults and children preferred an animal with a cause feature and an effect feature rather than an animal with two effect features. This study is the first direct demonstration of the causal status effect in children. PMID- 10856746 TI - Semantic negative priming in picture categorization and naming. AB - Ignoring a particular stimulus can hamper subsequent attended processing of the same stimulus, a phenomenon known as 'negative priming'. Previous research has yielded partial support for the claim that the effect results from the attentional inhibition of central representations, as suggested by the fact that it appears to extend to semantic associates of ignored stimuli. In the current experiment, participants categorized or named pictorial stimuli with superimposed distractor pictures; the effect of unrelated, identical, or categorically related previously ignored pictures was investigated. Furthermore, visual similarity between categorically related picture pairs was manipulated in the naming task. The results indicated negative priming from identical ignored pictures irrespective of task. However, semantic negative priming was observed in the categorization task only. In the naming task, even visual overlap between categorically related picture pairs failed to induce negative priming. These findings argue against a central locus of the negative priming effect. PMID- 10856747 TI - Stability in the HIV vDNA pool in peripheral CD4+ T cells of untreated patients by single tube quantitative PCR. AB - HIV infection leads to loss of CD4 T cells and development of AIDS in most individuals without treatment. While disease progression during HIV infection correlates with the plasma viral load, much less is known about the levels of HIV vDNA. This paper describes the development and validation of a sensitive, quantitative PCR assay for the assessment of HIV vDNA. The system uses novel single tube, multiply competitive PCR technology, which allows five-point competitor competition in a single PCR reaction. The reproducibility and performance characteristics of the assay are extensively studied, which indicate that the system performs well in high DNA backgrounds. Using this assay system on a cohort of protease naive patients, HIV vDNA was assessed from PBMCs over an average follow-up period of 5 years. The data indicate that the HIV vDNA pool does not appreciably accumulate over the follow-up period, with many of the patients followed for up to 8 years. A reliable, quantitative assessment of vDNA pools will allow a better understanding of the dynamics of HIV pathogenesis. PMID- 10856748 TI - Isolation and identification of bluetongue virus. AB - Bluetongue virus (BTV) is an arthropod-borne orbivirus that infects sheep, wild ruminants and occasionally cattle. Detection and specific identification of BTV is a multistep process. The first step involves the isolation of the virus from the animal's blood or other tissues, followed by inoculation of embryonating chicken eggs (ECE). After the virus has been amplified in ECE, it is passaged into BHK-21 cell culture for subsequent replication and identification. The virus is then amplified further and identified in microtiter plates by the immunoperoxidase assay using a group specific monoclonal antibody. Finally, the viral isolate is typed by a virus neutralization test. PMID- 10856749 TI - A fast one-step reverse transcription and polymerase chain reaction (RT-PCR) amplification procedure providing highly specific complementary DNA from plant virus RNA. AB - Reverse transcription and polymerase chain reaction (RT-PCR) are being used increasingly for detection and typing RNA viruses. For this purpose, metal block thermal cyclers (MBTC) are considered to provide higher DNA yield, whereas air thermal cyclers (ATC) allow PCR amplification in a much shorter time. A fast ATC protocol (0 s denaturation, 0 s annealing, and 4-8 s elongation) was developed to amplify genomic segments from two RNA viruses, which allowed increasing the number of cycles without a parallel increase of non-specific DNA fragments. Under these conditions, 80-90 cycles with the ATC provided a DNA yield close to that of a standard 40-cycles MBTC protocol in about half the time. The DNA synthesised by the new procedure was highly specific and could be cloned readily. PMID- 10856750 TI - Rescue of infectious classical swine fever and foot-and-mouth disease virus by RNA transfection and virus detection by RT-PCR after extended storage of samples in Trizol. AB - A method for storing samples containing classical swine fever virus (CSFV) or foot-and-mouth disease virus (FMDV), respectively, was developed, which abolishes the infectivity of both plus strand RNA viruses, and allows storage of samples above 0 degrees C for an extended time, yet preserves the viral RNA in a state which allows its detection by reverse transcription-polymerase chain reaction (RT PCR), and even rescue of infectious virus after transfection of the extracted RNA into susceptible cells. Supernatants from infected cell cultures as well as organs from diseased animals were stored in Trizol(R) for 1-4 weeks at -20 degrees C, 4 degrees C, room temperature, or 37 degrees C. RNA was then extracted and used subsequently for RT-PCR, as well as transfection into susceptible cells to initiate the replication of progeny virus. Formaldehyde-fixed samples were also included in this study. Storage up to 4 weeks at 37 degrees C in Trizol(R) still yielded positive RT-PCR results and rescue of infectious virus upon RNA transfection. In contrast, formaldehyde fixation reduced drastically the detectability of viral RNA. This method represents a safe and inexpensive alternative to -70 degrees C (dry ice) storage or transport of samples, and abolishes the biosafety risks involved in shipping deep-frozen infectious materials. PMID- 10856751 TI - A gag gene heteroduplex mobility assay for subtyping HIV-1. AB - A heteroduplex mobility assay (HMA) using 753 and 446 base pair (bp) amplicons of the p17/p24 region of the gag gene of HIV-1 has been developed and validated with reference clones and clinical samples representative of subtypes A, B, C, D, E, G, and H. There was complete concordance between the gag HMA assigned subtype and the subtype known from gag or env sequence data or env HMA. The heteroduplexes from both amplicons can be clearly resolved on either MetaPhor XR agarose or MDE polyacrylamide gels. The MetaPhor XR gel system was the more convenient and is the preferred choice for routine HMA subtyping. This gag HMA provides a rapid, simple and inexpensive method for subtyping HIV-1 based on a genomic region other than the commonly used env gene target. The incorporation of gag HMA into subtype determination algorithms should allow the detection of gag/env recombinant strains of HIV-1. PMID- 10856752 TI - Characterization of a strain-specific monoclonal antibody to hepatitis delta virus antigen. AB - Sequences of the hepatitis delta virus (HDV) vary to different degrees among isolates. A monoclonal antibody, designated as HP6A1, against the antigen of HDV (HDAg) has been characterized for its specificity. HP6A1 bound to HDAg of isolate 25 (genotype I) that was used for immunization, but not to others of both genotypes I and II. The epitope recognized by HP6A1 was then determined by a phage library displaying various heptapeptides. A consensus peptide deduced has the best match with that of residues 4-10 of HDAg (isolate 25). To confirm the phage mapping result, Escherichia coli recombinant proteins containing different lengths and various segments of HDAg (isolate 25) were constructed. The shortest HDAg segment contained in the fusion protein that reacted with HP6A1 was residues 1-10. When this peptide was added to the N-terminus of a heterologous protein engineered for eucaryotic expression, the fusion protein was detected by HP6A1. It is concluded that HP6A1 recognizes an epitope located at the N-terminus of HDAg (isolate 25). Since viruses of quasi-species exist in natural infections, a question of how different viral strains interact in vivo remains to be explored. The highly specific MAb opens a possibility to examine the fate of one strain in the presence of other related species in a cell transfection system. PMID- 10856753 TI - Peptide based enzyme-linked immunoassays for detection of anti-HSV-2 IgG in human sera. AB - Glycoprotein G of HSV-2 (gG2) and a peptide, corresponding to a previously recognised immunodominant epitope spanning residues 561-578 of the protein, were compared directly for type-specific serodiagnosis of HSV-2. The protein was affinity purified and obtained in a commercially available EIA kit while the peptide, previously designated as peptide 55, was made as a multiple antigenic peptide. A panel of 100 characterised serum samples (60 HSV-2 positive, 20 HSV-1 positive and 20 HSV negative) was screened using the two antigens. The intact protein and peptide 55 showed the same sensitivity for antibodies in the serum of HSV-2 infected individuals, reacting with 96.7% (58/60) of the samples. The peptide did not react with any of the HSV-1 positive or HSV negative sera. In contrast, gG2 gave a number of false positive results, reacting with 20% (4/20) of the HSV-1 positive sera and 10% (2/20) of the HSV negative sera. The superior performance of peptide 55, together with the very much lower costs of its production, compared with gG2 suggest that the peptide will become the antigen of choice in enzyme immunoassays for type-specific serodiagnosis of HSV-2. PMID- 10856754 TI - A monoclonal blocking EIA for herpes simplex virus type 2 antibody: validation for seroepidemiological studies in Africa. AB - A competitive type-specific enzyme-linked immunosorbent assay (ELISA) for herpes simplex virus type 2 (HSV-2) antibody was developed using an infected cell antigen and a monoclonal antibody to glycoprotein G-2. This assay has been validated for use for epidemiological studies using a large panel of sera collected in rural Uganda and a panel of 143 sera characterised previously by Western blotting, the 'gold standard' for HSV type-specific serology. This evaluation was found to have a sensitivity of 96% and a specificity of 91% in comparison with Western blot on 143 sera from clinic patients. The ELISA had a sensitivity of 93% and a specificity of 91% in comparison with Western blot on 495 sera collected in Uganda. The assay showed good reproducibility and a low percentage of sera gave equivocal results, indicating its suitability for epidemiological studies. PMID- 10856755 TI - Differentiation of hepatitis B virus genotypes D and E by ELISA using monoclonal antibodies to epitopes on the preS2-region product. AB - An enzyme-linked immunosorbent assay (ELISA) has been described for serological determination of hepatitis B virus genotypes, using monoclonal antibodies (mAb) against seven distinct epitopes (b, m, k, s, u, f and g) on the preS2-region products of hepatitis B surface antigen (HBsAg). The usefulness of this method for serological detection of genotype E, however, was theoretical, because no HBsAg samples of this genotype were included in the original test panel. Moreover, the predicted serotype of genotype E (bksufg) closely resembled that of genotype D (bksu, bksuf or bksug). Four HBsAg samples of genotype E were tested by the original described ELISA. The epitope g, predicted to be present in these samples by amino acid sequences, was not detected when HBsAg of genotype E was captured on a solid phase by mAb to the common determinant 'a' of HBsAg and then reacted with mAb to g (5156) labeled with horseradish peroxidase. However, the four examples of HBsAg of genotype E were captured by mAb 5156 to g on a solid phase; they were then detected by labeled mAb to the common determinant 'a'. Since epitopes f and g co-occurred on HBsAg of genotype E, HBsAg samples of this genotype were also detected, by 'sandwiching' them between immobilized mAb to g and labeled mAb to f. By contrast, HBsAg of genotype D in 90 sera was not reactive when sandwiched between mAb to f and g. Thus, this modified ELISA enables the serological determination of all six genotypes of HBsAg and, by inference, of hepatitis B virus. PMID- 10856756 TI - Ultrasensitive in-house reverse transcription-competitive PCR for quantitation of HIV-1 RNA in plasma. AB - An ultrasensitive version of an 'in-house' reverse transcription-competitive polymerase chain reaction assay described previously for quantitation of human immunodeficiency virus type 1 (HIV-1) RNA in plasma was developed. The increase in sensitivity from 400 to 50 HIV-1 RNA copies/ml was achieved by pelleting virus particles from 1.8 ml plasma by centrifugation prior to RNA extraction, modifying competitor DNA structure and amounts, and redesigning primers. Quantitation of HIV-1 RNA in 130 samples tested previously by the standard assay showed that the two procedures yield comparable results (mean absolute difference, 0.26+/-0.20 log) and that the ultrasensitive version detects HIV-1 RNA below the threshold of sensitivity of the standard method. The ultrasensitive 'in-house assay' and the reference QUANTIPLEX HIV-1 RNA 3.0 had the same sensitivity and gave equivalent results (mean absolute difference, 0.19+/-0.11 log), as shown by parallel blinded testing of 47 plasma samples. Titration experiments with reconstructed plasma samples allowed the determination of a dynamic range of 50-500000 HIV-1 RNA copies/ml for the 'in-house' system. The interassay coefficient of variation for samples nominally containing 200, 4000 and 80000 HIV-1 RNA copies/ml were 33.4, 22.9 and 38.2%, respectively. The performance, turnaround time, and cost effectiveness of this system make it suitable for medium-scale clinical application. PMID- 10856757 TI - The detection of small round-structured viruses in water and environmental materials. AB - A procedure for concentrating small round-structured viruses (SRSVs) (Norwalk like viruses) from water and other environmental materials is described. Primers based on the helicase region of the SRSV genome were confirmed as specific by reaction with typed specimens, and used to detect virus in concentrates of unseeded and seeded samples. Virus was detected in estuarine recreational water polluted by untreated sewage, although not in seawater samples taken some distance from outfall discharges. It was also detected in river water downstream of a sewage treatment plant. Virus could be detected in all matrices when they were seeded with a positive stool extract, including sewage seeded with as little as 2 microl stool extract, thus confirming the suitability of the method for environmental monitoring. PMID- 10856758 TI - Competitive ELISA for detection of antibodies to porcine reproductive and respiratory syndrome virus using recombinant E. coli-expressed nucleocapsid protein as antigen. AB - The 15 kDa nucleocapsid (N) protein is the most abundant protein of the porcine reproductive and respiratory syndrome virus (PRRSV), and is highly antigenic, which therefore makes it a suitable candidate for the detection of virus-specific antibodies and diagnosis of the disease. In this study, complementary DNA corresponding to the entire N gene of the IAF-Klop strain of PRRSV was cloned into the pGEX-4T-1 vector, and the N protein was expressed in Escherichia coli fused to the glutathione S-transferase (GST) protein. The resulting GST-N recombinant fusion protein was purified by affinity chromatography and used as antigen for serological testing by indirect enzyme-linked immunosorbent assay (ELISA). Two anti-N specific monoclonal antibodies (MAbs) (IAF-K8 and IAF-2B4), obtained following fusion experiments with spleen cells of BAlb/c mice that were immunized with the purified virus, were used in a competitive assay to increase the specificity of the ELISA. Both MAbs were found to be directed against highly conserved conformational epitopes of North American isolates of PRRSV. Optimal concentration of GST-N protein was determined by checkerboard titration, using hyperimmune pig antiserum to the homologous PRRSV strain, and corresponded to a range of 0.1-0.5 microg protein per well. When tested on 95 sera from pigs that were experimentally infected with the IAF-Klop strain, the competitive ELISA (K8 ELISA) was capable of detecting anti-PRRSV antibodies in 86.7% (65/75) and 92.6% (63/68) of pig sera known to be seropositive by indirect immunofluorescence (antibody titers >16) and a currently used commercial ELISA (HerdCheck(R); Idexx), with specificity values of 100 and 96.2%, respectively. When tested on clinical samples (542 sera) from 28 positive and 28 negative pig herds, the K8 ELISA performed in a similar way to HerdCheck(R) and immunofluorescence (IF) tests as shown by kappa values of 0.762 and 0.803. The sensitivity and specificity of K8-ELISA were 100% on a herd basis, whereas sensitivity values of 80 and 82% with a specificity of 98.7% were determined on an individual basis in comparison with HerdCheck(R) and IF tests. PMID- 10856759 TI - Molecular methods to distinguish between classical rabies and the rabies-related European bat lyssaviruses. AB - A rapid and sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) assay for the detection of classical rabies virus (genotype 1) and the rabies related European bat lyssaviruses (EBLs) (genotypes 5 and 6) was developed. When combined with specific oligonucleotide probes and a PCR-enzyme linked immunosorbent assay (PCR-ELISA), genotype 5 and 6 viruses can be distinguished from each other and from genotype 1 viruses. Ninety-two isolates from the six established genotypes of rabies and rabies-related viruses were screened by RT PCR and PCR-ELISA to determine the specificity of the assays. All genotype 1, 5 and 6 viruses were detected by RT-PCR while none of the genotype 2, 3 and 4 viruses were detected. All the genotype 5 and 6 viruses were detected by the two PCR-ELISA probes when used in combination while none of the genotype 1-4 viruses were detected. When used individually, the PCR-ELISA probes also distinguished between the genotype 5 and 6 viruses. This new discriminatory test should allow the rapid genotyping of all lyssaviruses likely to be encountered in Europe and as such could provide useful epidemiological information in the event of an outbreak. PMID- 10856760 TI - Detection of human serum antibodies against type-specifically reactive peptides from the N-terminus of glycoprotein B of herpes simplex virus type 1 and type 2 by surface plasmon resonance. AB - A single-step surface plasmon resonance protocol for the detection of antibodies against herpes simplex virus type 1 and type 2 (HSV-1, HSV-2) in human sera was established using the BIAcore system. Two peptides from corresponding segments of the N-terminus of HSV-1 and HSV-2 glycoprotein B (gB), i.e. peptide gB-1 (60-73) (GAAPTGDPKPKKNK) and peptide gB-2 (55-68) (SPATTKARKRKTKK), were identified as immunogenic. Employing both peptides as diagnostic antigens in the surface plasmon resonance assay, a sensitivity for the detection of HSV-1 and HSV-2 type specific antibodies of 83 and 86%, respectively, was achieved as compared with immunoblotting as a reference method. Peptide gB-1 (60-73) allowed the discrimination between HSV-1 and HSV-2 type-specific antibodies with a specificity of 67%, whereas peptide gB-2 (55-68) reacted in a strictly HSV-2 type specific manner. It is concluded that peptides from the N-terminus of gB-1 and gB 2 are recognized predominantly by human sera in an HSV-specific manner. Peptide gB-2 (55-68) can be employed successfully for the determination of type-specific antibodies against HSV-2. PMID- 10856761 TI - Genetic grouping of classical swine fever virus by restriction fragment length polymorphism of the E2 gene. AB - A method for genetic grouping of classical swine fever viruses (CSFV) was developed based on the restriction fragment length polymorphism (RFLP) revealed by AvaII, BanII and PvuII digestion of RT-PCR amplified segments of the E2 gene. From inspection of the genetic sequences of Thai isolates and reference strains, the RFLP method was designed to be capable of differentiating all known genogroups and subgenogroups suggested by phylogenetic analysis of the CSFV E2 gene. The method was applied to 60 CSFV samples which included three genogroups and seven subgenogroups. Unlike previously described RFLP methods, the agarose gel patterns obtained from these samples were completely in agreement with the predicted RFLP patterns and enabled accurate genetic grouping of CSFV at the subgenogroup level. The simplicity of this method allows rapid CSFV genogrouping at diagnostic laboratories without sequencing facilities and provides a useful method for diagnosis as well as epidemiological investigation and control of classical swine fever outbreaks. PMID- 10856762 TI - Simultaneous quantitation of two orchid viruses by the TaqMan real-time RT-PCR. AB - Simultaneous quantitation of two orchid viruses, cymbidium mosaic potexvirus (CymMV) and odontoglossum ringspot tobamovirus (ORSV), were carried out using the TaqMan((R)) real-time RT-PCR, a novel detection technique that combines RT-PCR with the power of fluorescent detection. Four TaqMan((R)) probes were synthesized, targeting at the RNA-dependent RNA polymerase (RdRp) and coat protein (CP) genes of both viruses. The reporter dye FAM (6-carboxyfluorescein) was used to label the 5' terminus of probes specific to CymMV, while TET (tetrachloro-6-carboxyfluorescein) was used for the ORSV probes. TAMRA (6-carboxy tetramethyl-rhodamine), which was attached at the 3' terminus of each probe, was used as the universal quencher. With increasing amounts of standard RNA templates, the respective threshold cycle (C(T)) values were determined and a linear relationship was established between these C(T) values and the logarithm of initial template amounts. The amounts of starting templates in mixed-infected Oncidium flowers and leaves were estimated from the standard curves. As little as 10(4) copies or 5 fg each of CymMV and ORSV could be detected simultaneously with either the RdRp or CP gene as the target. This system offers a sensitive, high throughput and rapid method for plant virus detection. PMID- 10856763 TI - The heteroduplex mobility assay (HMA) as a pre-sequencing screen for Norwalk-like viruses. AB - Molecular epidemiological studies of Norwalk-like viruses (NLVs), previously known as small round structured viruses (SRSVs), are dependant currently on DNA sequencing of PCR amplicons, which is expensive and time consuming. The Heteroduplex Mobility assay (HMA) was evaluated as a method for identification of PCR amplicons from the commonly circulating NLV strains without DNA sequencing. The procedure was developed for use with two reference strains, a Mexico virus like strain (MXV-like; Hu?NLV?RBH?1993?UK) and the Grimsby virus strain (Hu?NLV?Gimsby?1995?UK), and was optimised with regards to the annealing and electrophoresis conditions and the electrophoresis gel matrix. Using the optimised conditions, amplicons of less than 90% sequence identity formed visible heteroduplexes, allowing the strains to be placed into three categories; Mexico like, Grimsby-like and non-Mexico virus/non-Grimsby virus strains. Outbreak strains 'genotyped' previously by DNA sequencing as Mexico virus or Grimsby virus were identified correctly by the heteroduplex mobility assay. The procedure was applied prospectively to strains from 130 outbreaks occurring in the UK between 1997 and 1998. Heteroduplex mobility assay was successful on 120 (92%) strains of which 68 (57%) were GRV-like strains, three (2.5%) were Mexico virus-like strains and 49 (41%) were categorised as non- Mexico/non-Grimsby virus strains. Amplicons from 50 of the 120 strains were sequenced and there was perfect correlation between the heteroduplex mobility assay categorisation and phylogenetic analysis. HMA offers a rapid, robust and far cheaper alternative to sequencing for the identification of prevalent Norwalk-like virus genotypes for molecular epidemiological studies. PMID- 10856764 TI - Amplification of human genomic sequences by human papillomaviruses universal consensus primers. AB - Detection of human papillomaviruses DNA (HPV) in tumour samples is often determined by a PCR based approach with standard universal consensus oligonucleotides. It is shown that these primers under the same amplification conditions can amplify a human genomic sequence of 1361 nucleotides in oral carcinomas and normal DNA samples. This sequence is detected more easily as the copy number of HPV DNA decreases. Therefore, in tumour samples that have a low copy number of HPV or that are contaminated by normal tissue there is a potential risk of misidentification of the presence of HPV if this observation is not taken into account. PMID- 10856765 TI - A non-radioisotopic quantitative competitive polymerase chain reaction method: application in measurement of human herpesvirus 7 load. AB - Quantitative-competitive polymerase chain reaction (QCPCR) is a well-optimised and objective methodology for the determination of viral load in clinical specimens. A major advantage of QCPCR is the ability to control for the differential modulation of the PCR process in the presence of potentially inhibitory material. QCPCR protocols were developed previously for CMV, HHV-6, HHV-7 and HHV-8 and relied upon radioactively labelled primers, followed by autoradiography of the separated and digested PCR products to quantify viral load. Whilst this approach offers high accuracy and dynamic range, non radioactive approaches would be attractive. Here, an alternative detection system is reported, based on simple ethidium bromide staining and computer analysis of the separated reaction products, which enables its adoption in the analysis of a large number of samples. In calibration experiments using cloned HHV-7 DNA, the ethidium bromide detection method showed an improved correlation with known copy number over that obtained with the isotopic method. In addition, 67 HHV-7 PCR positive blood samples, derived from immunocompromised patients, were quantified using both detection techniques. The results showed a highly significant correlation with no significant difference between the two methods. The applicability of the computerised densitometry method in the routine laboratory is discussed. PMID- 10856766 TI - Foreword. PMID- 10856767 TI - Lipid lowering improves endothelial functions. AB - The healthy endothelium usually provides an anticoagulant, vasodilatory and anti inflammatory spectrum of functions that are central in vascular homeostasis. Dysfunction of the endothelium is a common feature of all phases of atherosclerosis. Hypercholesterolemia provokes many aspects of endothelial dysfunction before and during the development of atheroma. For example, a high cholesterol diet leads to the formation of a fatty streak and the recruitment and binding of blood leukocytes to the artery wall. This process requires expression by the endothelial cells of adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1). In rabbits that are fed an atherogenic diet, the aortic endothelium, which usually expresses little VCAM-1, shows foci of VCAM-1 expression soon after initiating this diet. Furthermore, lowering plasma cholesterol by diet or drugs down-regulates the expression of VCAM-1 and reduces the density of inflammatory cells in the atherosclerotic plaque. Hypercholesterolemia also attenuates normal vasodilatation to several stimuli such as exercise and acetylcholine. In healthy subjects, the vascular endothelium produces the vasodilator nitric oxide. In atherosclerosis, however, nitric oxide bioavailability is impaired. As a result, atherosclerotic coronary arteries commonly display a vasoconstrictor response when challenged with acetylcholine. Lipid lowering appears to favorably influence endothelial vasomotor and inflammatory functions in ways that may benefit patients with coronary artery disease. Continued probing of the basic mechanisms of endothelial dysfunction and its treatment may lead to new therapies that offer clinical benefits in patients with atherosclerosis, including reductions in coronary events. PMID- 10856768 TI - Plaque stabilization: the role of lipid lowering. AB - The unstable atheroma characteristically has a thin, eccentric fibrous cap and a large necrotic core of lipid and cellular debris. This plaque configuration is particularly unstable because large mechanical stresses develop in the thinnest portions of the fibrosis cap over the lipid pool. Numerous recent observations tell us that critical biological factors other than lesion morphology also contribute to the ultimate fracture of the fibrous cap. Matrix-degrading enzymes, particularly members of the metalloproteinase family, are overexpressed in atherosclerotic tissue. The regions with the greatest amounts of these enzymes are the critical high stress regions, which are usually also infiltrated with inflammatory cells. At the cellular level a number of factors regulate expression of these enzymes by vascular smooth muscle cells. These factors include mechanical stimuli and cytokines known to be overexpressed in atheroma by both smooth muscle cells and macrophages. Low density lipoprotein cholesterol in the lesion plays a significant role in most or all of these processes, and our current understanding of the unstable atheroma is consistent with the dramatic clinical successes of lipid-lowering therapy. PMID- 10856769 TI - The role of small, dense low density lipoprotein (LDL): a new look. AB - Plasma low density lipoprotein (LDL) plays a central role in atherogenesis, and elevated levels of LDL are associated with an increased risk of coronary heart disease (CHD). Studies have now revealed that LDL is structurally heterogeneous, based on its size and density. Patients with combined hyperlipidemia exhibit a lipid profile - the so-called atherogenic lipoprotein phenotype - that is associated with elevated triglyceride levels, low levels of high density lipoprotein and a preponderance of atherogenic, small, dense LDL particles. Such individuals are at an increased risk of CHD events, regardless of their total LDL circulating mass. Evidence suggests that when plasma triglycerides exceed a critical threshold of approximately 133 mg/dl (1.5 mmol/l), this favours the formation of small, dense LDL from larger, less dense species. Lipid-lowering agents that are capable of lowering triglyceride levels below this threshold value will cause a shift to a less dense and, therefore, less atherogenic LDL profile. This effect has been demonstrated for the HMG-CoA reductase inhibitor atorvastatin which, in addition to its ability to markedly decrease the total LDL circulating mass, can also shift the LDL profile towards less dense, larger species. This suggests that atorvastatin may also affect the atherogenic lipoprotein phenotype found in patients with combined hyperlipidemia. PMID- 10856770 TI - The undertreatment of LDL-cholesterol: addressing the challenge. AB - Patients with dyslipidemia are at increased risk of coronary heart disease (CHD), while treatment to reduce low density lipoprotein cholesterol (LDL-C) concentrations lessens this risk. Consequently, the Lipid Treatment Assessment Project (L-TAP) was undertaken in the US to determine the extent to which primary care practitioners utilize lipid-lowering therapy and to evaluate the success of current therapeutic regimens, using the National Cholesterol Education Program (NCEP) guidelines as therapeutic targets. The L-TAP study, initiated in 1996 and completed in February 1997, recorded LDL-C levels in 4888 patients from 619 US practices. All patients had received lipid-lowering therapy for at least 3 months. The primary care practitioners involved in the study were questioned about the NCEP guidelines and the results confirmed that these physicians were representative of primary care practitioners in the USA. The 4888 patients were categorized according to risk: patients with <2 risk factors (RFs) but no CHD, those with >/=2 RFs but no CHD and patients with confirmed CHD. Overall, only 38% of the patients attained LDL-C target levels or had values lower than these goals. The greater the number of RFs, the lower the proportion of patients achieving target levels. LDL-C targets were less often attained in patients receiving dietary therapy only compared with those receiving lipid lowering drug treatment. However, there was good correlation between the success of treatment and both receipt of and compliance with dietary instruction. In conclusion, a large proportion of dyslipidemic patients who are being treated in primary care are not achieving NCEP target LDL-C levels. PMID- 10856771 TI - Development of an improved in vitro activity assay for medium chain length PHA polymerases based on CoenzymeA release measurements. AB - An improved activity assay for polyhydroxyalkanoate (PHA) polymerases from Pseudomonas oleovorans GPo1 was developed. The activity assay is based on the detection of released Coenzyme A (CoA) using 5,5'-dithiobis (2-nitrobenzoic acid) (DTNB), a compound which specifically reacts with thiol groups. The formed adduct was measured spectrophotometrically with high sensitivity and accuracy. The assay was used to study the effect of several additives on the activity of granule associated PHA polymerase. Mild non-ionic detergents such as Tween-20, Triton X 100, CHAPS and Hecameg all appeared to be strongly inhibitory. In contrast, bovine serum albumin (BSA) had a strong stimulatory effect on the activity and stability of the PHA polymerases. Using optimized conditions, activities up to 5.8 U/mg granule-bound polymerase have been measured. PMID- 10856772 TI - A comparison of 35S-SO(4)(2-) radiotracer techniques to determine sulphate reduction rates in laminated sediments. AB - In order to find a simple and efficient method to determine sulphate reduction rates in environmental samples, we tested different 35S-SO(4)(2-) radiotracer techniques. The methods varied in the application of 35S-SO(4)(2-) and subsequent extraction of reduced 35S-sulphur species. Samples were either incubated as sediment slurries mixed with the radiotracer, or as undisturbed sediment cores after core injection of the radiotracer. Reduced 35S-sulphur species were retrieved passively by diffusion or actively by reflux distillation. The methods were applied to surface sediments derived from three aquatic habitats situated in Germany: (1) a tideless brackish water, (2) a mining lake and (3) a natural freshwater lake. The best possible method was expected to yield the highest sulphate reduction rates, which were reproducible with respect to magnitude and depth distribution. At the same time, we aimed to keep the disturbance of samples as well as the expenditure of labour and equipment to a minimum. For all three types of aquatic habitats, the combination of core injection followed by diffusion was the most reliable and efficient method. This combination is therefore recommended for determination of sulphate reduction rates in laminated sediments. PMID- 10856773 TI - Simple detection method for distinguishing dead and living yeast colonies. AB - A rapid and simple assay was developed for detection of yeast colonies containing dying or dead cells. Methylene blue, phloxin B, rose bengal and trypan blue at concentrations of 5-10 micromol l(-1) were shown to stain non-viable cells in colonies of Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans and Filobasidium capsuligenum without staining or affecting the viability of living cells of the colonies. PMID- 10856774 TI - Adaptation of E. coli cell method for micro-scale nitrate measurement with the Griess reaction in culture media. AB - The E. coli cell method for nitrate measurement consists of two-steps: nitrate reduction by the E. coli cell usually under anaerobic conditions and subsequently nitrite measurement with the Griess reaction. It was found that the E. coli DSM 498k wildtype cell can reduce nitrate to nitrite under aerobic conditions. Therefore, the E. coli method for nitrate measurement was adapted to be performed under aerobic conditions in a microtiter plate. The adapted method is simpler than the original E. coli method and other nitrate methods such as those with inorganic reductants and with purified enzymes. Furthermore, it was found that for the Griess reaction the pH values of samples after addition of the Griess reagent A should be lower than 1.8 for a stable absorbance at 540 nm to be reached. It is important to add the two Griess reagents separately and to read the absorbance twice consecutively in a microtiter plate. The adapted E. coli method was successfully applied to measure the traces of nitrate in MRS and other medium components by measuring the standard curve of a dilution of each individual medium component. It was found that many organic medium components contain traces of nitrate, while none of them contain detectable nitrite. Among these, the extract of meat and yeast extract contain relatively high amounts of nitrate: 217 mg N/kg and 99 mg N/kg respectively. MRS broth contains nitrate from 0.3 to 0.6 mg N/l depending on the batch numbers of the product. The adapted E. coli can also be used for nitrate measurement in other matrices. PMID- 10856775 TI - Genotypic characterization of Salmonella typhi by amplified fragment length polymorphism fingerprinting provides increased discrimination as compared to pulsed-field gel electrophoresis and ribotyping. AB - Amplified fragment length polymorphism (AFLP) is a recently developed, PCR-based high resolution fingerprinting method that is able to generate complex banding patterns which can be used to delineate intraspecific genetic relationships among bacteria. In the present study, AFLP was evaluated for its usefulness in the molecular typing of Salmonella typhi in comparison to ribotyping and pulsed-field gel electrophoresis (PFGE). Six S. typhi isolates from diverse geographic areas (Malaysia, Indonesia, India, Chile, Papua New Guinea and Switzerland) gave unique, heterogeneous profiles when typed by AFLP, a result which was consistent with ribotyping and PFGE analysis. In a further study of selected S. typhi isolates from Papua New Guinea which caused fatal and non-fatal disease previously shown to be clonally related by PFGE, AFLP discriminated between these isolates but did not indicate a linkage between genotype with virulence. We conclude that AFLP (discriminatory index=0.88) has a higher discriminatory power for strain differentiation among S. typhi than ribotyping (DI=0.63) and PFGE (DI=0.74). PMID- 10856776 TI - Rapid detection of Mycoplasma pneumoniae in clinical samples by real-time PCR. AB - M. pneumoniae is a common causative agent of community-acquired pneumonia in children. The diagnosis of such infections is usually based on serology using complement fixation or, more recently, enzyme-immuno assays. PCR has been shown to be a promising alternative. We have evaluated a real-time PCR assay targeting the P1 adhesion protein gene and compared it to a conventional semi-nested PCR assay with the 16S rDNA as target. Comparison of 147 specimens from 48 patients showed an overall agreement of 97.4%. Real-time PCR proved to be of equal value on clinical specimens as conventional PCR regarding sensitivity and specificity, but is clearly advantageous regarding speed, handling and number of samples that can be analyzed per run. PMID- 10856777 TI - Evaluation of a simplified semi-quantitative protocol for the estimation of Vibrio vulnificus in bathing water using cellobiose-colistin agar: a collaborative study with 13 municipal food controlling units in Denmark. AB - A simplified semi-quantitative method using pre-enrichment in alkaline peptone water supplemented with polymyxin B and plating onto cellobiose-colistin (CC) agar for the estimation of Vibrio vulnificus in bathing water was evaluated. This protocol was tested in a collaborative study with 13 food controlling laboratories in Denmark during the 1999 bathing season in periods when water temperatures exceeded 20 degrees C. The average percentage of yellow colonies larger than 1 mm in diameter on CC agar that could be identified as V. vulnificus by colony hybridization with a species-specific DNA probe was 79%. This high percentage of specificity demonstrated that by using CC agar in estimating the level of V. vulnificus in bathing water, recognition of yellow colonies larger than 1 mm is sufficient for the identification of V. vulnificus with no further characterization needed. The simplified protocol may be included in the routine control of the microbial quality of bathing water done by the local food controlling laboratories, since it involves simple traditional and low-cost microbiological methods with no use of molecular skills or sophisticated equipment. PMID- 10856778 TI - Heteroduplex mobility assay detects DNA mutations for differentiation of closely related phytoplasma strains. AB - We present the first use of DNA heteroduplex mobility assay (HMA) to detect the point mutations including substitutions and deletions/insertions in 16S rDNA of aster yellows phytoplasma (AY27) and to differentiate phytoplasmas collected from field samples of clover proliferation (CP) and alfalfa witches'-broom (AWB). The phytoplasmal 16S rDNA fragment was amplified from AY27 by polymerase chain reaction (PCR) and cloned into a plasmid vector. The cloned DNA fragment was subjected to in vitro mutation to produce 1- to 4-base substitutions and 1- to 3 base deletions. The mutated 16S rDNA fragments were analyzed by HMA. The results showed that a single two-base substitution or a single-base deletion/insertion in the 529 bp DNA fragment was directly detected and that a DNA divergence at a level of as low as 0.2% was detectable by HMA. Heteroduplex mobilities were affected by the number and composition of the phytoplasma DNA bases in mismatches or gaps and were proportional to the degree of DNA divergences. Gaps caused greater retardation in heteroduplex mobility than mismatches did. HMA was highly sensitive in detecting the mixed infections of phytoplasmas. In analyses of CP and AWB field samples collected in Alberta, two CP and one AWB phytoplasma isolates were differentiated from others by HMA but not by restriction fragment length polymorphism (RFLP). Therefore, HMA provides a simple, rapid, highly sensitive and analytical method to detect and estimate the genetic divergence of phytoplasmas when other methods such as RFLP are not readily applicable. PMID- 10856779 TI - Real-time monitoring of nitrile biotransformations by mid-infrared spectroscopy. AB - In this study mid-infrared spectroscopy was used to follow the enzyme kinetics involved in nitrile biocatalysis using whole cell suspensions of the bacterium Rhodococcus rhodochrous LL100-21. The bacteria were grown on acetonitrile to induce a two-step enzymatic pathway. Acetonitrile was biotransformed to acetamide by a nitrile hydratase enzyme and subsequently to acetic acid (carboxylate ion) by an amidase enzyme. The bacteria were also grown on benzonitrile to induce a one-step enzymatic pathway. Benzonitrile was biotransformed directly to benzoic acid (carboxylate ion) by a nitrilase enzyme. These reactions were followed by React IR using a silicon probe and gave excellent quantitative and qualitative real-time data of both nitrile biocatalytic reactions. This study has shown that this novel technique has potentially useful applications in biocatalysis. PMID- 10856780 TI - Immuno-capture differential display method (IDDM) for the detection of environmentally induced promoters in rhizobacteria. AB - A rapid immunological method for trapping and selection of functionally regulated prokaryotic promoters is described. The method is based on application of a novel mini-Tn5 derived promoter probe (pUTTKZY-promoterless lacZY as a reporter and kanamycin resistance) to mutagenise a plant growth promoting fluorescent pseudomonad, Pseudomonas fluorescens 54/96. The transposon allows selection of operon fusion mutants (lacZY(+)) directly on media containing lactose as a sole carbon source as well as selection for kanamycin and lacZ (beta-galactosidase) expression on X-gal indicator media. We have extended the technique to target the surface expression of the induced lactose permease gene (lacY) from mutagenised libraries and the immuno-capture of bacteria with magnetic beads and anti-LacY monospecifc antisera. The benefits of the lacZY reporter are that a library can be rapidly generated and screened in vitro to isolate non-expressed mutants for further in situ screening. Here we demonstrate the development and utility of the technique and its potential as a differential display method for the isolation of promoters that direct regulated gene expression in the phytosphere, or under other imposed conditions. PMID- 10856781 TI - Effects of handling aids on calf behavior. AB - Effects of three different handling aids on calf behavior were determined. Group 1 calves were intensively-reared intact Holstein males (mean 180 days old); Group 2, extensively-reared beef-breed females (mean 230 days); Group 3, extensively reared castrated beef-breed males (mean 253 days). Calves in each group were assigned to one of three handling aid treatments (n=5 per treatment subgroup; total n=45): electric prod (Prod), oar with rattles (Oar), manual urging (Manual). Treatments were applied only as needed to encourage forward movement of calves through the length of a solid-sided semicircular chute system. Number of treatment applications, length of time required to move through the entire chute system, and behavior during movement through the chute were recorded. An approach test was conducted 1 day before and 1 day and 1 week after chute tests to evaluate changes in behavior due to handling aid application. During chute tests, Group 1 Prod calves required the fewest treatment applications (4.9) vs. 23.5 (Oar) or 13.5 (Manual), ran most often (1.40 times) vs. 0.20 times (Manual) or 0.33 times (Oar), and made contact with chute sides most often (1.8 times vs. 0.2 times (Manual) or 0.7 times (Oar), respectively (all P<0.05). Similar trends were observed for calves in Groups 2 and 3. There were no significant differences between behaviors observed during the approach tests conducted before and after handling aid treatments had been imposed. Regardless of treatment, intensively reared Group 1 calves appeared markedly less fearful of handlers during approach tests compared to extensively-reared calves in Groups 2 and 3, which demonstrated overt attempts to escape from the test facilities. One week after chute tests, 13 of 15 Prod calves from all three groups walked, rushed, or backed >1 m away from the handler when the prod was buzzed but not applied, suggesting that the buzzing sound alone may have sufficed to encourage movement by calves that had previously experienced both the sensation and sound associated with electric prodding. PMID- 10856782 TI - Strategies for the avoidance of faeces by grazing sheep. AB - Experiments were conducted to investigate which environmental cues were used by sheep when discriminating against patches of pasture contaminated with faeces. The influence of the spatial distribution of contaminated patches and the parasite infection status of sheep on avoidance of contaminated patches and ingestion of parasite larvae was also investigated. In experiment 1, sheep infected with the parasite Ostertagia circumcincta were given the opportunity to graze in uncontaminated or aggregated contaminated patches. Patch contamination comprised of either faeces from sheep infected with O. circumcincta larvae, faeces from uninfected sheep, or O. circumcincta larvae only. Infected sheep discriminated against faeces from parasite-infected animals and faeces from uninfected animals equally. Sheep did not discriminate against patches contaminated with parasite larvae only. In experiment 2, sheep infected with O. circumcincta and uninfected sheep grazed experimental plots with differing spatial patterns of faecal-contaminated patches, allowing animals the opportunity to forage in contaminated or uncontaminated patches of herbage. Plots were also grazed by infected and uninfected animals that were fistulated at the oesophagus to enable the collection of ingested herbage. Sheep spent a greater proportion of their time foraging in uncontaminated patches than in contaminated patches. Where patches were highly aggregated, infected animals spent a greater proportion of total grazing time in uncontaminated patches than did uninfected animals, and grazed uncontaminated patches for longer on each sampling occasion. On grazing plots where all patches were contaminated, the difference between the numbers of larvae isolated from pasture herbage and ingested herbage was greatest for infected animals. In this situation, infected animals avoided parasites most. On grazing plots consisting of both contaminated and uncontaminated patches, the difference between the numbers of larvae isolated from pasture herbage and ingested herbage was greatest for uninfected animals. In this situation, uninfected animals were most effective at parasite avoidance as they consumed fewer parasite larvae relative to what was available on pasture. PMID- 10856783 TI - Social hierarchy in the domestic goat: effect on food habits and production. AB - Outside the scientific world, the effect of social behaviour on production is little taken into account, but the importance of this relationship has been sufficiently proven in some animal species. Nevertheless, there are scarce works that emphasise the importance of behaviour in the production of the goat. The main objective of this paper is to determine if there is a stable hierarchy of dominance in a flock of goats fed in pasture, and if this hierarchy influences somehow the diet selected in the pasture and in its production of milk and meat. The study was carried out in a flock of goats in semi-extensive grazing management. The interactions observed in the pasture during the supplementary feeding and during the milking were written down. This allowed us to determine the dominance rank. The diet was determined in the pasture by the direct observation method. The production of milk was measured daily. The meat production consisted on the weight of the kids in their first day of life and after a month. Among the most prominent results, the following should be indicated: (a) Within the herd, a clearly established, quite stable and linear hierarchic order exists. (b) The most aggressive animals are those that occupy the highest positions within the social hierarchy. (c) Age, large size and horns seem to be the physical factors that most favor dominance. (d) When more forage becomes available, differences appear in the diet chosen by dominant and subordinate animals, that is, they become more selective. In the months of greater shortage, these differences in feeding disappear, and they become more generalist. (e) The production of animals is affected by dominance. However, contrary to what might otherwise be thought, it is the middle range of goats that are the most productive. PMID- 10856784 TI - Prevalence of behaviour problems reported by owners of dogs purchased from an animal rescue shelter. AB - This study examined the prevalence of behaviour problems exhibited by dogs within 4 weeks of acquisition from a rescue shelter in Northern Ireland. One thousand five hundred and forty-seven people who had purchased a dog from a rescue shelter in Northern Ireland were sent a postal questionnaire designed to collect information on the behaviours exhibited by their dog within the first month of acquisition. Five hundred and fifty-six people responded to the survey, representing a response rate of 37%. The majority of respondents (68.3%) reported that their dog exhibited a behaviour problem, the most common being fearfulness. Most of those respondents (89.7%) who returned their dog to the shelter did so because the animal exhibited behaviour that they considered undesirable. Male dogs showed more unacceptable behaviours than females, specifically inter-male aggression, sexual problems and straying tendencies. More stray dogs displayed undesirable behaviour than unwanteds, specifically straying tendencies. Puppies were less likely to exhibit unacceptable behaviours than juveniles or adults, particularly fearfulness, sexual problems and straying tendencies. More juvenile dogs showed excessive activity and excessive barking than puppies or adults. More adult dogs displayed aggression towards other dogs than juveniles or puppies. Findings indicate that dogs purchased from rescue shelters do exhibit behaviour problems that may lead to their return. The number of dogs admitted or returned to rescue shelters with behaviour problems may be reduced by raising public awareness regarding the value of behaviour therapy and introducing behaviour therapy schemes to rescue shelters. PMID- 10856785 TI - The effect of increasing visual horizons on stereotypic weaving: implications for the social housing of stabled horses. AB - Stabled horses commonly perform stereotypic patterns of weaving, where the horse shifts its weight from side to side often swinging its head. Ten warm-blood types, of which five were known to reliably weave, were housed in similar 12x12 ft wooden loose boxes in a single stable block surrounding a courtyard. Each horse was exposed to each of five stable designs. These were: the conventional front top-half of the door open only with a view of the stable courtyard (F); front half-door open and a similar half-door open at the back of the stable with a view to the surrounding fields (FB); back open only (B); front and one-side panel open with a view into the adjacent stable (FS); and front, back and both sides open (All4). During observation days, horses were brought in from the field at 0830 h, fed concentrate at 0930 h, fed haylage at 1005 h and turned out at 1600 h. Behaviour was recorded from 0900 to 1040 h, 1200 to 1300 h and 1500 to 1600 h. Weaving was most common prior to feeding in the morning and prior to putting out to pasture in the afternoon. There was a significant effect of stable design on weaving, with less weaving in the FS and All4 designs than the F treatment. There was also a significant effect of stable design on repetitive nodding, though in this case, FB, B, FS and All4 designs each reduced nodding compared with the F treatment. The effect of stable design can be explained in a number of ways. Firstly, it could be the novelty of the environmental change, though there was no evidence in this study of an increase in stereotypy with prolonged exposure to the new stable designs. Secondly, opening windows may increase opportunities for environmental interaction, and the expression of new activities may compete with stereotypic behaviour for the horse's time. Thirdly, the open windows may allow expression of specific activities such as environmental monitoring or social interaction that are denied by the conventional stable. PMID- 10856786 TI - Identification of T-helper cell epitopes in the hypervariable region of the nucleocapsid (N) protein of rinderpest virus (RPV) in cattle. AB - The proliferative responses to synthetic peptides by lymphocytes derived from rinderpest virus (RPV)-infected cattle, the natural host for RPV, were assayed by determining [3H]thymidine incorporation into the DNA. In eight out of twelve cattle tested, significant responses were detected to peptides representing amino acids 452-501 in the C-terminal hypervariable region of the virus nucleocapsid (N) protein. It appears that helper T-cell epitope(s) for cattle which can be broadly recognized within an MHC diverse population, exists in this region of the protein. PMID- 10856787 TI - Egg-adapted replication-restricted virus protects mice against lethal influenza. AB - Infection of mice with the egg-adapted (EA) strain of influenza virus was studied as a murine model of human live attenuated virus vaccine. The growth and spread of the EA virus in the mouse lungs were restricted, and only small inflammatory changes were detected in the respiratory tracts. Deletion and substitutions of amino acids were found in the hemagglutinin molecule of the EA virus, which were attributable to the reduced envelope fusion activity in virus multiplication cycles. Intranasal inoculation of mice with the EA virus induced specific IgG and IgA antibody production together with a specific cytotoxic T lymphocyte response. Immunized mice showed a distinct resistance to subsequent lethal challenge with the virulent influenza virus. These results indicate that the mutant virus loaded with a growth restriction in the respiratory tract is an appropriate candidate for a live attenuated vaccine. PMID- 10856788 TI - Early life humoral response of ducks to DNA immunization against hepadnavirus large envelope protein. AB - DNA vaccination may represent an interesting strategy for early life immunization. However, in some cases, this approach has been shown to induce a tolerance rather than immunity. We have compared the efficiency of neonatal DNA or protein immunization against hepadnavirus envelope protein using the duck hepatitis B virus (DHBV) model. Three-day-old ducklings were immunized with either a plasmid encoding the DHBV pre-S/S large envelope protein (L), or a recombinant preS protein, followed by sequential DNA or protein boosts at weeks 4 and 15. Our results showed that genetic immunization of duck neonates induced specific humoral response to DHBV L protein. Interestingly, an enhanced antibody response was elicited when animals received DNA priming-DNA boosting as compared to DNA priming-protein boosting. PMID- 10856789 TI - Induction of Th2-directed immune responses by IL-4-transduced dendritic cells in mice. AB - Dendritic cell (DC)-based vaccines have been used to generate Th1-mediated, protective immunity against cancers and infectious microorganisms. As an attempt to develop a new vaccine protocol for the induction of Th2-directed responses, we introduced an IL-4 plasmid vector into the XS106 DC line (derived from A/J mice). Although relatively small fractions of XS106 cells exhibited apparent intracellular deposition of IL-4, they secreted biologically relevant amounts of the cytokine. IL-4-transduced XS106 DC and control XS106 DC transfected with vector alone were pulsed with KLH and injected s.c. into A/J mice. The overall magnitude of KLH-specific cellular and humoral responses was comparable between the two animal groups. However, they differed in the isotype profile albeit only transiently, with the IL-4-transduced DC group showing higher IgE and lower IgG2a responses, and in the cytokine profile, with spleen cells isolated from the IL-4 transduced DC group producing higher IL-13 and lower IL-12. Thus, delivery of IL 4 gene to relatively small numbers of DC is sufficient to modify the immunological outcome of DC-based vaccines. PMID- 10856790 TI - Duration of rubella immunity induced by two-dose measles, mumps and rubella (MMR) vaccination. A 15-year follow-up in Finland. AB - A national two-dose vaccination program with a combined measles, mumps and rubella (MMR-II) vaccine was introduced in Finland, in 1982, immunizing children at the ages of 14-18 months and 6 years. Antibody levels were determined from serial samples from a group of originally 350 children during 15 years. The latest samples were taken 15.5 years after the first vaccination and 70% of the children could still be reached. The aim of this study was to determine the kinetics of rubella antibodies induced by the MMR-II vaccine in these individuals. Rubella antibodies were analyzed from three different cohorts: Group I seronegative children (n=166) vaccinated at 14-18 months and 6 years, Group II seronegative children (n=139) and Group III seropositive children (n=16) vaccinated at 6 and 11-13 years. Samples collected 0-9 years after vaccination were analyzed by hemolysis-in-gel (HIG) and later samples by enzyme immunoassay (EIA) techniques. The primary vaccination induced 100% seropositivity in vaccinees with a mean zone diameter of 10 (+/-1.3), 10.2 (+/-1.1) and 11.5 (+/ 0.9) mm, in Groups I, II and III, respectively. The seropositivity rate was still high at 15 years, 99%, 100% and 100% with the geometric mean titer 23, 46 and 105 IU/ml, respectively. At 15 years, antibody levels <15 IU/ml which is the suggested protective level, were found in 31, 9 and 0% of children in Groups I, II and III, respectively. Because almost a third of the individuals in Group I now, at the age of 17 years, had low levels of rubella antibodies, it is possible that rubella infections may re-emerge during pregnancy. A careful surveillance including serological follow-up is therefore very important. PMID- 10856791 TI - Immunization of rhesus monkeys with a recombinant of modified vaccinia virus Ankara expressing a truncated envelope glycoprotein of dengue type 2 virus induced resistance to dengue type 2 virus challenge. AB - Dengue epidemics increasingly pose a public health problem in most countries of the tropical and subtropical areas. Despite decades of research, development of a safe and effective live dengue virus vaccine is still at the experimental stage. To explore an alternative vaccine strategy, we employed the highly attenuated, replication-deficient modified vaccinia Ankara (MVA) as a vector to construct recombinants for expression of the major envelope glycoprotein of one or more dengue virus serotypes. MVA recombinants expressing the highly immunogenic C terminally truncated dengue type 2 virus (DEN2) or dengue type 4 virus (DEN4) envelope protein (E), approx. 80% of the full-length, were evaluated for their protective immunity in animal models. Each of these recombinants elicited an elevated antibody response to DEN2 or DEN4 E in mice following the booster inoculation, as detected by radio-immunoprecipitation. Recombinant MVA-DEN2 80%E, but not MVA-DEN4 80%E, induced a neutralizing antibody response. The MVA-DEN2 80%E recombinant was chosen to further evaluate its ability to induce resistance to wild type DEN2 challenge in monkeys. Monkeys immunized twice with recombinant MVA-DEN2 80%E developed a low to moderate antibody response and were partially protected against DEN2 challenge, as determined by the viremia pattern. Importantly, the subsequent study showed that all four monkeys immunized with the recombinant in a three dose schedule developed an increased level of antibodies and were completely protected against DEN2 challenge. The potential efficacy of recombinant MVA-DEN2 80%E to protect primates against dengue infection suggests that construction and evaluation of MVA recombinants expressing other serotypes of dengue virus E for use in a tetravalent vaccine strategy might be warranted. PMID- 10856792 TI - Chimeras between the human immunodeficiency virus (HIV-1) Env and vaccinia virus immunogenic proteins p14 and p39 generate in mice broadly reactive antibodies and specific activation of CD8+ T cell responses to Env. AB - A vaccine based on the envelope protein (Env) of the human immunodeficiency virus type 1 (HIV-1) that triggers widely reactive antibodies might be a desirable approach to control virus infection. To expose epitopes which could induce broadly reactive antibodies against HIV-1 Env, we have generated vaccinia virus (VV) recombinants that express Env fused at its N- or C-terminus with two major antigenic proteins of VV, p14 (A27L gene) and p39 (A4L gene). Biochemical analysis of the chimeric proteins in cell cultures revealed that, in all cases, recombinant viruses expressed the correct fusion proteins. When p14 or p39 are fused at the N-terminus of Env the chimeric proteins are poorly glycosylated but when p14 or p39 are fused at the C-terminus of Env, the chimeric proteins are fully glycosylated. In Balb/c mice, immunisation with the referred VV recombinants induced similar levels of CD8+ T cell specific responses to Env as immunisation with the entire Env protein. The humoral immune response triggered by the fusion proteins was broader than in animals immunised with VV expressing the entire Env (VVEnv1), and was directed to epitopes outside of the V3 loop (V1/V2, C1, C2, C4). One of the chimeric constructs induced a better neutralising antibody response than VVEnv1. We conclude that fusing VV proteins p14 or p39 to Env provides an effective means to induce broadly reactive antibodies and CD8+ T cell responses to Env. This approach might have utility against HIV and other pathogens. PMID- 10856793 TI - Comparison of immunogenicity and reactogenicity of a measles, mumps and rubella (MMR) vaccine in German children vaccinated at 9-11, 12-14 or 15-17 months of age. AB - Children aged 9-11, 12-14 or 15-17 months, respectively were vaccinated with a measles, mumps and rubella (MMR) vaccine and serum antibody responses and reactogenicity were compared. The data of 118 children could be analysed (group 1=9-11 months, n=46; group 2=12-14 months, n=29, group 3, 15-17 months, n=43). The only significant difference observed was for seroconversion against measles virus between group 1 and group 3 (84.8% vs 100%, p=0.012). No serious adverse events were reported. Local side reactions were mild, infrequent and independent of age. Immunisation against MMR is safe and effective even when administered before the currently recommended age of 12 months. PMID- 10856794 TI - Development of semisynthetic triterpenoid saponin derivatives with immune stimulating activity. AB - Aldehyde-containing triterpene saponins have adjuvant properties, but only those from Quillaja saponaria Molina stimulate the production of cytotoxic T lymphocytes (CTL) against exogenous antigens. Quillaja saponins have two normonoterpene ester moieties, linked linearly to their fucosyl residue, that play a critical role in the stimulation of CTL. These ester moieties are also responsible for these saponins' instability and toxicity. Based on the structure activity relationships for the different groups of Q. saponaria saponins, new semi-synthetic analogs were developed that have the adjuvanticity of quillaja saponins, yet with less toxicity and greater stability in aqueous solutions. The quillaja saponin analogs were prepared by replacing their hydrolytically unstable ester groups with another lipophilic chain linked by a stable amide bond on these saponins' glucuronic acid residue. One of these analogs, GPI-0100, is a dodecylamide saponin derivative that stimulates an antibody isotype profile that corresponds to a Th1 type immune response, as well as CTL production against exogenous antigens. PMID- 10856795 TI - Anti-major histocompatibility complex antibody responses in macaques via intradermal DNA immunizations. AB - In simian immunodeficiency virus (SIV) models, immunization of macaques with uninfected human cells or human major histocompatibility complex (MHC) proteins can induce xenogeneic immune responses which can protect the animals from subsequent SIV challenges. These studies suggest that the induction of anti-MHC immune responses can be a viable vaccine strategy against human immunodeficiency virus type 1 (HIV-1). We have previously shown in mouse studies that DNA immunization with class I and class II MHC-encoding plasmids can elicit both xenogeneic and allogeneic antibody responses against conformationally intact MHC molecules (Vaccine 17 (1999) 2479-92). Here we take these observations one step closer to human applications and report that intradermal needle immunizations of non-human primates with plasmid DNA encoding human MHC alleles can safely elicit xenogeneic anti-MHC antibody responses. Moreover, injecting macaques with DNA encoding a specific macaque allogeneic MHC induced anti-allogeneic MHC antibodies production. These studies show that DNA immunization with MHC-encoding vectors can indeed be used to induce specific anti-human xenogeneic, as well as anti macaque allogeneic MHC immunity in non-human primates. This strategy could thus be used to mobilize anti-MHC antibody response which may be useful as part of an anti-HIV-1 vaccination approach. PMID- 10856796 TI - A dengue virus serotype-1 DNA vaccine induces virus neutralizing antibodies and provides protection from viral challenge in Aotus monkeys. AB - A DNA vaccine that expresses the premembrane/membrane (prM) and envelope (E) genes of dengue virus serotype-1 was tested for immunogenicity and protection against dengue-1 virus challenge in Aotus nancymae monkeys. The vaccine, in 1 mg doses, was administered intradermally (i.d.) to three monkeys and intramuscularly (i.m.) to three others. For controls, a 1 mg dose of vector DNA was administered i.d. to two monkeys and i.m. to one. All animals were primed and then boosted at one and five months post priming. Sera were collected monthly and analyzed for dengue-1 antibodies by enzyme linked immunosorbent assay (ELISA) and plaque reduction neutralization test (PRNT). Dengue-1 antibodies were detectable in the sera from i.d. and i.m. vaccine inoculated animals one month after the first boost and peaked one month after the second boost. The antibody levels from sera of animals that received the vaccine via the i.d. route were twice those from sera of animals that received the vaccine via the i.m. route. Six months after the second boost all inoculated and two naive monkeys were challenged with 1.25x10(4) plaque forming units (PFU) of dengue-1 virus. Two vaccine immunized animals were protected from viremia while the others showed a reduction in viremia. The mean days of viremia were 1 and 1.3 for the animals that were immunized with the vaccine via the i.d. or i.m. route, respectively vs 4 and 2 mean days of viremia in the animals inoculated with control DNA. Naive animals were viremic for an average of 4 days. All of the three control monkeys that received control DNA inoculum by either the i.d. or i.m. route had an intermittent viremia pattern with one or more negative days interspersed between the positive days. This pattern was not observed in any of the vaccine recipients or the naive control monkeys. These results demonstrate that DNA immunization is a promising approach for the development of dengue vaccines and that A. nancymae monkeys are suitable for dengue vaccine trials. PMID- 10856797 TI - Ex vivo targeting of the macrophage mannose receptor generates anti-tumor CTL responses. AB - MUC1 is highly expressed in adenocarcinomas and is a possible target for immunotherapy. In mice, oxidized mannan linked to MUC1 (M-FP), given in vivo, induces potent MHC-restricted CTL and tumor protection. Because of the resistance of cancer patients to immunization, ex vivo immunization of macrophage/dendritic cells was examined using oxidized mannan MUC1 to target the mannose receptor and the MHC Class I antigen presentation pathway. Here, we show that murine mannose receptor (MR) bearing macrophages derived from peritoneal exudate cells (PEC) and cultured ex vivo with M-FP can, after adoptive transfer, efficiently present MUC1 to T cells, leading to the generation of high frequency of CTL and protection from tumor challenge. Mice immunized once with syngeneic PEC pulsed with M-FP elicit a similar CTLp frequency to that obtained with three in vivo immunizations. Targeting the MR is crucial to obtain high frequency CTL, and without oxidation the CTLp frequency was low. GM-CSF is important, as GM-CSF o/o mice gave reduced responses, a deficiency corrected by in vivo GM-CSF. In addition, the treatment of macrophages ex vivo with GM-CSF gave enhanced responses and treating mice with GM-CSF prior to M-FP immunizations also enhanced cellular responses. M-FP targets the MR and ensures rapid passage of peptides to Class I molecules, and can also directly stimulate in vitro IL-12 production by macrophages. While many studies are now focussing on dendritic cells, in this study the cells involved were adherent F4/80+ 33D1- macrophages. The findings could be of benefit for the immunization of patients with cancer. PMID- 10856798 TI - Analysis of latency in cattle after inoculation with a temperature sensitive mutant of bovine herpesvirus 1 (RLB106). AB - Calves were inoculated with the bovine herpes virus 1 (BHV-1) vaccine strain (RLB 106), which is a temperature sensitive mutant. The route of inoculation was intranasal instillation or intramuscular (i.m.) injection (flank or neck). As a control, five calves were given placebo by i.m. injection of the neck. Regardless of the infection route, clinical symptoms did not occur. However, BHV-1 neutralizing antibodies were detected after inoculation demonstrating that sero conversion occurred. At 60 days post-inoculation, dexamethasone was given by i.m. injection to attempt reactivation of RLB 106. Only those calves inoculated by the intranasal route shed virus leading to an increase in BHV-1 specific antibodies. As expected, viral DNA and the latency related-RNA were detected in trigeminal ganglia (TG) of calves inoculated by the intranasal route. In contrast, viral nucleic acid was not detected in TG of calves inoculated by the i.m. route or in calves inoculated with placebo. In cervical ganglia or sacral dorsal root ganglia, viral nucleic acid was not consistently detected. This study provides evidence that efficient latency and reactivation does not occur following i.m. inoculation. Since serum-neutralizing antibodies were detected in all inoculated calves, i.m. inoculation led to sero-conversion. PMID- 10856799 TI - Protective immunity induced by the full-length cDNA encoding paramyosin of Chinese Schistosoma japonicum. AB - The full-length cDNA encoding paramyosin of Chinese S. japonicum (Sjc97) has been cloned and sequenced for the first time. The homology of the nucleotide sequence of paramyosin and the deduced amino acid sequence between Chinese, Philippine, Japanese strains and S. mansoni (Sm97) were 99.7, 99.8 and 96% at amino acid level, respectively; 99.4, 99.2 and 91% at nucleotide level, respectively. The immunogenicity and protective efficacy of a DNA vaccine encoding Sjc97 was evaluated in C57BL/6 mice. Mice immunized intramuscularly with pCMV-Sjc97 resulted in predominantly IgG2a and IgG2b immune responses, and immune sera were able to mediate antibody-dependent macrophage mediated cytotoxicity in vitro. Cytokine profiles in immunized C57BL/6 mice demonstrated Th1 bias, with IFN-gamma and IL-2 production and lack of IL-4 and IL-5. Immunization with pCMV-Sjc97 conferred a significant level of protection against cercariae in C57BL/6 mice. These results demonstrate that the nucleic acid encoding Sjc97 was able to induce a Th1 type immune response and confer protective efficacy in C57BL/6 mice when administrated via the intramuscular route. PMID- 10856800 TI - Direct composite inlays versus conventional composite restorations: 5-year follow up. AB - OBJECTIVES: To determine at 5 year follow-up the failure rate, wear rates and other aspects of clinical performance of direct composite inlays compared with conventional composite restorations placed incrementally. METHODS: 100 matched pairs of restorations were originally entered into the trial. Each pair consisted of a direct composite inlay and a conventional composite restoration made from the same material. At 5 years it was possible to recall 65 pairs, of which 54 were complete. Clinical assessments were made using USPHS criteria (indirect measurements of occlusal wear were made using Ivoclar standard dies) and annual bite wing radiographs. RESULTS: There was a trend to more failure of inlays than conventional composites (17.4 c.f. 7.5%) but this was not significant. The clinical performance of both types of restoration was similar and compared favourably with the results of studies of other materials. Secondary decay was diagnosed in only one restoration. Between 3 and 5 years there was some deterioration in cavo-marginal discoloration, marginal adaptation (occlusally) and surface roughness (occlusally). There was no apparent deterioration in colour match, proximal contact, shim stock contacts and Gingival Index. Wear rates of both types of restoration showed no significant difference and were essentially linear with a mean of 33-34 microm per year. CONCLUSIONS: Both inlays and conventional composite restorations complied with ADA specification minimum requirements for posterior composite restorations. In this study the direct inlay technique gave no clinical advantage over conventional, incremental placement. PMID- 10856801 TI - Performance of tunnel restorations at 3-6 years. AB - OBJECTIVES: the purpose of this study was to evaluate the success of the tunnel restoration method in the Norwegian public dental service. METHODS: all patients from three age cohorts (born 1975-1977) who had received one or more tunnel restorations at least 3 years earlier, at the public dental clinic in Kongsberg and a neighboring clinic in Numedal, were examined clinically and radiographically by two calibrated dentists. Individuals with two or more filled surfaces per year were classified as "caries active". The statistical analyses consisted of non-parametric Kaplan-Meyer estimates of the survival function, and rank tests for associations to the longevity data and the background variables. RESULTS: a total of 182 restorations in 94 patients were studied. Sixty-five percent of the restorations were considered successful. A total of 118 restorations were censored within the 76-month observation period. The median survival time was estimated to be 55 months, with a 95% confidence interval of 51 61 months. About 90% survived 3 years, while only 35% survived 5 years. Both caries activity and operator had significant effects on the survival period. On the other hand, there was no difference between "wells" and "tunnels", tooth type, tooth surface or jaw with regard to success rate. CONCLUSIONS: The tunnel preparation filled with currently available glass-ionomer cement is not a generally favorable alternative in primary approximal lesions. However, in the hands of a well-trained, careful operator it may be chosen as a semi-permanent solution for patients with modest caries activity. PMID- 10856802 TI - Water fluoridation, poverty and tooth decay in 12-year-old children. AB - AIM: To examine the influence of water fluoridation, and socio-economic deprivation on tooth decay in the permanent dentition of 12 year old children. SETTING: The North of England, fluoridated Newcastle and non-fluoridated Liverpool. A total of 6,638 children were examined. OUTCOME MEASURES: Multiple Regression analysis of fluoride status, mean electoral ward DMFT in 1992/93 and ward Townsend Scores from the 1991 census. RESULTS: Social deprivation and tooth decay were significantly correlated in areas with and without water fluoridation. Multiple linear regression showed a statistically significant interaction between ward Townsend score, mean DMFT and water fluoridation, showing that the more deprived the area the greater the reduction in tooth decay. At a Townsend score of zero (the English average) there was a predicted 37% reduction in decay in 12 year-olds in fluoridated wards. CONCLUSIONS: Tooth decay is strongly associated with social deprivation. The findings confirm that the implementation of water fluoridation has markedly reduced tooth decay in 12-year-old children and that socio-economic dental health inequalities are reduced. PMID- 10856803 TI - Oral pH and drinking habit during ingestion of a carbonated drink in a group of adolescents with dental erosion. AB - OBJECTIVES: To investigate the relationship between dental erosion, oral pH and drinking habit in a group of adolescents. METHODS: Oral pH was measured simultaneously at the surface of four teeth in 11 patients, aged 10-16 years, with erosion and in 10 controls subjects without erosion using antimony electrodes. Measurements were made before, during and after drinking 330 ml of a carbonated drink. The method and timing of drinking the beverage, reported dietary intake of acidic foods and flow rate and buffering capacity of saliva were recorded. RESULTS: The erosion patients reported drinking more carbonated drinks (p<0.01) and drinking directly from a can more frequently than the controls (p<0.05). They also drank twice as quickly (p<0.05). The pH at the buccal surface of a molar remained lower for longer in the erosion patients than in the patients without erosion (p<0.01), whilst the labial surface of the upper central incisor had a longer exposure to low pH in the controls subjects (p<0.05). CONCLUSION: The pattern of oral pH differed between subjects with and without erosion after drinking an acidic beverage. This may be related to observed differences in drinking habit, which could have influenced the pattern of erosion in these subjects. PMID- 10856804 TI - Marginal integrity of large compomer Class II restorations with cervical margins in dentine. AB - OBJECTIVES: The aim of the present study was to investigate the marginal integrity of total-bond Dyract AP restorations in large Class II restorations with cervical margins in dentine. The efficacy of a new non-rinse conditioner and the effect of beveling the enamel margins were also studied. METHODS: Large MOD cavities with cervical margins located 1mm below the CEJ were prepared in 48 extracted human molars. Six groups (n=8) were filled using a total-bond technique with Spectrum TPH or Dyract AP or a sandwich technique with Dyract in combination with Spectrum TPH. For Dyract AP total-bond restorations, a new non-rinse conditioner was tested vs. a total-etch with 36% phosphoric acid in beveled and butt-joint cavities. After water storage for 21 days and thermocycling (2000x, 5 55 degrees C), replicas were produced for quantitative marginal analysis in the SEM. Afterwards, teeth were immersed in 0.5% basic fuchsin for 24h and dried. Percent dye penetration over the total margin length was analysed in three layers using a sequential grinding technique. Statistical analysis was performed with non-parametric tests and the Bonferroni correction for multiple comparisons at p<0.05. RESULTS: All restoration types showed microleakage. On cervical margins in dentine, Dyract AP restorations showed better marginal adaptation than Spectrum TPH total-bond restorations (marginal openings (MO), median 33.8 vs. 79.6%), but were inferior to Dyract/Spectrum TPH sandwich restorations (MO: 0. 0%). The non-rinse conditioner improved the marginal adaptation of Dyract AP restorations in dentine (MO: 4.7 vs. 38.4%, p=0.0206 for beveled cavities, 12.2 vs. 33.8%, p=0.0238, for butt-joint cavities) and yielded similar results in enamel margins provided that enamel margins were beveled. Beveling of enamel significantly reduced the occurrence of enamel microcracks. CONCLUSIONS: The use of a non-rinse conditioner in combination with Dyract AP may improve the marginal adaptation of Class II restorations compared to composite restorations. The sandwich technique with Dyract results in better marginal adaptation in cervical dentine compared to all other restorative techniques tested in this study. However, microleakage cannot predictably be prevented with the sandwich technique. PMID- 10856805 TI - Improving fissure sealant quality: mechanical preparation and filling level. AB - OBJECTIVE: To assess the effect of different modes of fissure preparation and filling level on the quality of pits and fissure sealants. METHODS: Various modes of fissure preparation in combination with two filling levels were examined. A total of 90 caries-free extracted human molar teeth were divided into three groups according to fissure preparation: (a) no mechanical preparation; (b) mechanical preparation with a round carbide bur; and (c) mechanical preparation with a tapered fissure diamond bur. All fissures were acid-etched and each group was subdivided according to filling level, either to the border or overfilled, using Helioseal (Vivadent, Schaan, Lichtenstein) as a sealant material. A modified microleakage assay was performed, combining occlusal loading and thermocycling, prior to staining with 0.5% Basic Fuchsin. After sectioning, teeth were examined and photographed in a reflected light microscope, sealant retention was observed and recorded. Microleakage and sealant penetration depths were measured for each section. Two-way ANOVA was used for statistical analysis. RESULTS: Sealant penetration and retention were significantly improved by mechanical preparation compared to non-prepared fissures (p<0.0001), and preparation with a tapered fissure diamond bur was superior to the round carbide bur. Overfilled fissures caused significantly higher levels of microleakage (p<0.004). CONCLUSIONS: Based on the results of this in vitro study, it is suggested that fissure sealant quality may be improved by mechanical preparation, preferably with a tapered diamond bur, and filling just to the border. Overfilling of fissures should be avoided. PMID- 10856806 TI - In vivo study of the pulp reaction to Fuji IX, a glass ionomer cement. AB - OBJECTIVE: Our aims were to investigate the pulp biocompatibility of Fuji IX, a glass ionomer cement (GIC) used as a restorative material in cavities prepared in rat's upper molars, and to assess the value of this in vivo model for testing dental biomaterials. METHOD: Half-moon class V-like cavities were drilled on the mesial aspect of 26 rat upper first molars. Half of the experimental rats whose molars were restored with the GIC were killed after 8days and the second half after 30days. They were compared with two control groups, also submitted to cavity preparation, but with cavities left unfilled. Again, half of the control rats were killed at 8 days and the second half after 30days. Following intracardiac perfusion with the fixative solution, the specimens were processed to histologic procedures. RESULTS: After 8 days, in both groups a few inflammatory cells were observed. The odontoblastic layer was disrupted and dilated blood vessels were seen in the pulp area related to the cut tubules. The experimental group displayed a moderate inflammatory reaction whereas only a slight reaction was detected in the control group. In few teeth, bacteria were visualized in dentine tubules beneath the GIC restoration. Such colonies were not observed in unfilled molars.After 30days, in both groups, the pulp tissue recovered and displayed a normal appearance. Disruptions of the odontoblast layer were not visible anymore. Bacteria penetration into dentine tubules was reduced compared with the 8-day situation. A thick layer of reparative osteodentine was formed. However no difference in thickness was detected between the experimental and control groups, supporting that the formation of reparative dentine is not impaired. Irregular mineralizations including calcospherites were induced by the GIC. CONCLUSIONS: This study demonstrates that, despite small alterations in the mineralization processes, the GIC Fuji IX has a good biocompatibility and does not induce any harmful effect on pulp cells. PMID- 10856807 TI - Microleakage and SEM interfacial micromorphology of amalgam restorations using three adhesive systems. AB - OBJECTIVE: This study evaluated the microleakage and interfacial micromorphology of Class V cervical amalgam restorations lined with OptiBond, AElitebond, or Panavia 21. METHODS: Unlined amalgams served as control. Cavities were treated with each dentin bonding system according to the manufacturers' instructions and restored with Tytin non-gamma 2 spherical amalgam. After one week of storage in tap water at 37 degrees C, the specimens were thermocycled (1000 cycles, 6-60 degrees C, 30s dwell time). Microleakage was assessed by means of basic fuchsin dye penetration and recorded according to an ordinal scale. RESULTS: None of the systems tested in this study completely eliminated microleakage. Kruskal-Wallis one-way ANOVA and Mann-Whitney U test found that on the occlusal wall, Panavia 21 and the control group had the least leakage (P<0.05). No statistically significant differences were found at dentin margins (P>0.05). Wilcoxon matched pairs signed rank test found that Panavia 21 and the control group had less leakage at the occlusal than at the dentin margins (P<0.05); when AElitebond and OptiBond groups were evaluated, microleakage at the enamel and at the dentin margins was similar for each group. With the adhesive systems, perhaps the hydrophilic bonding agents incorporated the dye during specimen immersion and/or sectioning. CONCLUSIONS: The use of adhesives may not be as worthy as resin cements for sealing and bonding amalgam restorations to enamel and dentin. PMID- 10856808 TI - Time-and-motion study on class II copy-milled ceramic inlays. AB - OBJECTIVES: To obtain an estimate of working times of Class II copy-milled ceramic inlays as an indication of their efficiency, and to analyse factors of influence. METHODS: In a controlled clinical trial, 173 MO/DO or MOD ceramic inlays of the Celay system were constructed in 101 patients. Treatment was carried out by seven dentists. The inlays were replacements of existing amalgam restorations in both molar and premolar teeth. Net-times needed for the treatment and for the laboratory stage were registered. Variables of influence on the working time were assessed by using ln-working times in ANOVA. Differences of working times were expressed as the relative difference between the upper and lower value of each variable (Delta). RESULTS: The mean clinical treatment time was 67(+/-22)min. The laboratory stage required 60(+/-17)min working time. 'Cavity modification technique' (composite basing/conventional, Delta=17%), 'clinical operator' (seven dentists, Delta=57%), and 'size of the restoration' (large/medium, Delta=16%) significantly influenced the clinical treatment time (p<0.001). Although the dentists were familiar with inlay construction, they showed a 25% decrease in working time towards the end of the study. CONCLUSIONS: Class II copy-milled ceramic inlays in this time-and-motion study required about 125min of working time. Working time increased by applying composite basing and making large restorations. The dentist influenced working times, while efficiency increased over time. Dentists needed more laboratory time to produce an inlay than the dental technician. PMID- 10856809 TI - Evaporation of solvent in one-bottle adhesives. AB - OBJECTIVE: To compare the evaporation of vehicles included in different single bottle adhesives as a function of time. METHODS: The following adhesives were used: Prime and Bond 2.1 (Dentsply); Single Bond (3M); PrimaBond 97' (BJM); Syntac Sprint (Vivadent); Optibond Solo (Kerr) and Syntac Single Component (Vivadent). Two primers that are used in multiple-component adhesives (PermaQuick Primer, Ultradent and Scotchbond Multipurpose Primer, 3M) as well as distilled water, 96% ethanol and acetone were also included in the experiment. Samples of these products were stored in small glass containers under controlled temperature and humidity conditions. Initials mass and mass after different periods of time were registered to calculate the percentage loss of mass. RESULTS: The analysis of variance of the results followed by orthogonal contrast comparisons revealed a significantly higher loss of mass in the single-bottle adhesives that contain organic vehicles (especially in those in which the vehicle was acetone). A somewhat lower loss of mass was found in a water-based product. CONCLUSION: There is a relationship among the evaporation possibilities of the vehicles used in single-bottle adhesive systems and their loss of mass during storage. PMID- 10856810 TI - Characterisation of tribochemically assisted bonding of composite resin to porcelain and metal. AB - OBJECTIVE: The fracture of bonded ceramic to metal restorations remains a problem in clinical dental practice. The use of resin based composites to repair such fractures is generally unsatisfactory. Tribochemical technology creates a surface layer of small silica particles fused to the surface substrate. Such a layer potentially improves adhesion of resin to both alloy and porcelain. Adhesion between two substrates is traditionally studied using shear or tensile bond strength tests. However, the highest stress at bond failure may not represent the real bonding characteristics correctly. An alternative method is to describe the bonding characteristics by determining the strain energy release rate for a given interface. This study compares the bonding characteristics of a resin to gold/porcelain interface using a tribochemical coating process with those of a control group using simple gritblasting. METHODS: Pre-cracked specimens were subjected to load-unload cycles using a simple four point bending test and the resultant strain energy release rates were calculated. RESULTS: Tribochemically pretreating the porcelain resulted in a significant increase in the resultant strain energy release rate from 42.72+/-3.65J/m(2) for the controls to 61.35+/ 6.26J/m(2). Likewise there was a significant improvement in the strain energy release rate for the gold/composite interface from 27.31+/-3.00J/m(2) to 42.13+/ 4.83J/m(2). CONCLUSIONS: Tribochemical technology offers significant potential advantages for clinical dental practice. PMID- 10856811 TI - The effect of postcuring on quantity of remaining double bonds, mechanical properties, and in vitro wear of two resin composites. AB - OBJECTIVES: To determine the effect of different postcuring methods on degree of conversion, mechanical properties, and in vitro wear of two resin composites (Z100 and Charisma). The postcuring methods involved devices for inlay curing as well as devices present for other purposes in many dental laboratories or dental offices. METHODS: Specimens of the resin composites were initially light cured and then postcured according to one of the following methods: Translux EC handheld curing unit (10min), Translux EC light box (10min), Triad II (10min), 40 degrees C (10min), 70 or 110 degrees C for 10min, 1, 6, or 24h. The properties were determined following storage of the specimens for 1week in water at 37 degrees C. The degree of conversion was determined using transmission IR. The mechanical properties tested were diametral tensile strength, flexural strength, and flexural modulus. In vitro wear was induced by a three-body wear simulator. RESULTS: Most postcuring methods increased degree of conversion of both materials. Postcuring increased the mechanical properties and in vitro wear resistance of Charisma, whereas no effect of postcuring was found on these properties of Z100. CONCLUSION: Postcuring with the use of devices readily available in the dental laboratory and dental office increased the degree of conversion of Z100 and Charisma as well as the mechanical properties and in vitro wear resistance of Charisma. A heat treatment at 110 degrees C for 10-60min was found to be the most promising postcuring method. PMID- 10856812 TI - Life cycle of Eimeria procera in experimentally infected grey partridges (Perdix perdix). AB - We examined exogenous and endogenous development of Eimeria procera in experimentally infected grey partridges (Perdix perdix). Our examination included data on morphology, localization, duration of schizogony and gametogony and morphology of sporulated oocysts. The endogenous stages of E. procera developed in large numbers within the epithelial cells of caecal crypts. The asexual development comprised three generations of schizonts. The first fully developed macrogametes and microgamonts were observed on Day 5 post-infection (p.i.) in histologic section. The patent period began on Day 6 p.i. and ended on Day 11 p.i. with peak production of oocysts on Days 7 and 8. Long oval oocysts of E. procera measured 25.78-28.13 microm in length and 14.06-15.24 microm in width, sporulation time ranged from 18 to 24h at 25 degrees C and from 36 to 48h at 20 degrees C. PMID- 10856813 TI - In vitro and in vivo efficacy of lasalocid for treatment of experimental cryptosporidiosis. AB - In vitro viability of purified Cryptosporidium parvum oocysts, exposed for 30, 60, 90 and 120min to 0.27mg/ml lasalocid suspension was evaluated by inclusion or exclusion of two fluorogenic vital dyes and an excystation technique. Continuously, preventive and curative efficacies at different doses (9, 6.75, 5.625 and 4.5mg/kg body weight) and regimens of lasalocid against cryptosporidial infection were evaluated on an experimental neonatal mice model. In vitro assays demonstrated a decrease in the oocyst viability related to an increase in exposure time for exposure to the lasalocid suspension. The infection was eradicated when the suspension was administered with a dose of > or = 6.75mg/kg body weight. No apparent toxic effects were observed. PMID- 10856814 TI - The impact of internal parasites on the productivity of young cattle organically reared on semi-natural pastures in Sweden. AB - A grazing experiment with young cattle was conducted over two consecutive (1997, 1998) grazing seasons on semi-natural pasturelands in central-eastern Sweden. Comparisons were made between groups of animals that were either untreated and set-stocked, ivermectin bolus treated and set-stocked or untreated but moved in mid-summer (mid-July) to ungrazed pasture. The whole experimental area had remained virtually free of cattle during the previous two seasons and the cattle had been raised indoors since birth. To introduce low-levels of parasite infection into the experimental system, each animal received a 'priming dose' of approximately 10, 000 infective trichostrongylid larvae at the time of turnout for both years. Results of the first year study showed that the level of parasitism was so low that it failed to induce any productivity losses in both groups of untreated cattle, which grew as well as those given boluses at turnout. In contrast, in 1998 both groups of untreated cattle suffered varying degrees of sub-clinical and clinical parasitism to result in an average of 30kg liveweight depression, compared with the bolus treated cattle, at the end of the season. The only major departure between the two years was that in the latter, the cattle in the untreated groups were exposed to infective larval pickup, which had overwintered on pasture. Cattle in the move treatment grazed in the same sequence on pastures used by similar classes of animals during the previous year. That is, their pastures at turnout had not been grazed since mid-summer of the previous year. Clearly this early season (1997) grazing by young cattle resulted in sufficient overwintered larvae at the start of the following year (1998) to cause productivity losses of the same magnitude as those recorded for young cattle grazing on pastures contaminated for the entire grazing season of the previous year. This was confirmed by tracer tests that were carried out on all treatments, at the time of turnout and the mid-summer move in 1999. These results have major significance to organic cattle producers in Sweden who have a much higher tendency to practice a variety of grazing management techniques aimed at controlling nematode parasite infections in young cattle, than their conventional farming colleagues. It has been identified that one of these strategies is to simply use summer/autumn saved pastures for young stock at turnout, which if grazed by young stock prior to this, could prove to be counter-productive. PMID- 10856815 TI - Cross-sectional serological survey on gastrointestinal and lung nematode infections in first and second-year replacement stock in the netherlands: relation with management practices and use of anthelmintics. AB - A cross-sectional survey was carried out on 86 farms randomly distributed in The Netherlands. After housing following the first and the second grazing season (FGS and SGS) serum samples were collected to determine IgG levels against Cooperia oncophora and Dictyocaulus viviparus, and the pepsinogen content. A questionnaire was used to inquire on grazing management practices and the use of anthelmintic drugs. On 80.7 and 60.2% of the farms FGS and SGS animals, respectively, were treated at least once with an anthelmintic drug. The percentage for the SGS animals indicates that the use of anthelmintic drugs in those animals has increased enormously over the last 10-15 years. Generally, parasitic nematode control in the FGS is good on most farms, but it can be characterised as being overprotective. There is a tendency that if anthelmintic drugs are used in the FGS they also are used more often in the SGS. On 12 farms (14%), no anthelmintic drugs were given in the FGS and the SGS. These farms did not differ from the others with respect to management practices in any obvious way. The serological results were in general very low, indicating low levels of exposure to gastrointestinal nematode infection in both FGS and SGS animals. This was not surprising in view of the good to high level of nematode control practices reported by the farmers. Although not statistically significant, a consistent result was that serological results for the SGS animals were more often positive or on average higher on those farms where FGS parasite control tended to be excessive. For D. viviparus, a prevalence rate of 41% positive farms was found. Following comparison with previous data, it is speculated that lungworm (sero )prevalence in replacement stock may be declining as a result of continuing high levels of parasite control in replacement stock. It is concluded that the results confirm previous surveys, lending support to the conclusion that parasitic nematode control on Dutch dairy farms, certainly in FGS calves, is good but tends to be overprotective. PMID- 10856816 TI - Differentiation by polymerase chain reaction - restriction fragment length polymorphism of some Oestridae larvae causing myiasis. AB - The cytochrome oxidase I (COI) gene of the most wide-spread Italian species of Oestridae larvae causing myiasis (Gasterophilus spp., Hypoderma bovis, Hypoderma lineatum, Oestrus ovis and Przhevalskiana silenus) was amplified by polymerase chain reaction (PCR) using conserved primers. Restriction fragment length polymorphism (RFLP) of amplicons was also carried out and their restriction profiles compared. A clear genetic difference between the Oestridae larvae examined was demonstrated by using Taq(alpha) I, Hinf I, Rsa I and Hpa II enzymes. No intra-specific variation in RFLPs was detected between the two species of Hypoderma. The results highlight the taxonomic and phylogenetic relationships among larvae belonging to the different subfamilies, and thus offer additional diagnostic and epidemiological instruments. PMID- 10856817 TI - Serum antibody to Sarcoptes scabiei and house dust mite prior to and during infestation with S. scabiei. AB - In this study, serum antibodies to Sarcoptes scabiei var. canis (SS), Dermatophagoides farinae (DF), and D. pteronyssinus (DP) were determined in 19 healthy, random-source dogs prior to infestation with scabies then again during a primary infestation, cure and challenge infestation with scabies. Prior to scabies infestation, serum of 11 dogs contained faintly detectable amounts of IgE and/or IgG to proteins in SS extract, probably resulting from sensitization to dust mites that share cross-reactive antigenic epitopes with SS. After becoming infested with scabies, the response to SS antigens became stronger with antibodies appearing to more antigens as the scabies infestation progressed. Three of the newly recognized proteins were 170, 155 and 142/133kD and could be used in a diagnostic test since antibodies to them appeared during the primary infestation. In addition, during the primary infestation, 14 of 15 dogs developed IgE to 1-11 new SS proteins in addition to an increase in IgE binding to those proteins recognized prior to infestation. Overall, the strongest antibody responses (IgE and IgG) were exhibited during cure of the first infestation, when dead mites were still present in the stratum corneum. As expected, the antibody response was strong and rapid during challenge when the infestation self-cured. The immunogenic SS proteins identified by serum antibody binding during challenge, when the hosts self-cured, are candidates for inclusion in a vaccine. These candidate proteins are 200, 185, 170, 155, 142/133, 112, 97, 74, 57, 45/42, 32 and 22kD. Some of the proteins in SS that exhibited new or increased antibody binding during the experiment also had IgE and IgG binding to proteins with similar molecular weights in DF and DP extracts. These results illustrate the difficulties involved in understanding and interpreting serum antibody for developing a serological test for the diagnosis of scabies, isolating relevant SS antigens that could be included in a vaccine for prevention of scabies, and for understanding the immune response mechanism to scabies. PMID- 10856818 TI - Efficacy of doramectin 0.5% w/v Pour-On for control of the horn fly, Haematobia irritans. AB - In 1998, three groups of cattle at three locations in Lousiana were treated with Dectomax((R)) (0.5% doramectin) Pour-On and horn fly populations were monitored. Acceptable levels (less than 50 flies per side) of horn fly control were observed from 4 to 8 weeks. Differences in the length of control among the three sites were most likely affected by immigration of adult flies from untreated groups. In 1999, acceptable horn fly control was obtained for 13 weeks by the use of two treatments of doramectin Pour-On. PMID- 10856819 TI - Ocular Onchocerca infections in two dogs in western United States. AB - Two dogs, one from California and one from Arizona, were found to have aberrant infections caused by filarial nematodes of the genus Onchocerca. In both cases, the parasites are localized in or near the eye. In one case the worm was located in the cornea and was surgically removed. In the second case, a very marked granulomatous reaction was induced in the retrobulbar space, mimicking an abscess. This eye was enucleated. The worms in both instances were female, and were gravid, i.e. contained microfilariae in utero, indicating that a male worm(s) had been present and mating had occurred. The exact identity of the species of Onchocerca responsible cannot be determined, although the features observed are most like Onchocerca lienalis of cattle. These cases represent the fourth and fifth such cases reported from the US, and are especially interesting because of the unusual location of the worms, the small number of recognized cases, and the similarity to a recent zoonotic human infection. PMID- 10856820 TI - Hypothalamic c-fos-like immunoreactivity in high-fat diet-induced obese and resistant mice. AB - Some C57Bl/6 mice become obese, whereas others remain lean when raised on a high fat diet. The mechanisms underlying this interindividual susceptibility to diet induced obesity remain unknown. Because hypothalamus plays a major role in the regulation of body weight, this study was conducted to identify the differences of hypothalamic neuronal activity between diet-induced obese and diet-resistant mice. Using c-fos as a marker, this study showed that diet-induced obese mice significantly increased c-fos-like immunoreactive neurons in the dorsal part of lateral hypothalamus (+183%) and dorsomedial hypothalamic nucleus (+87.5%) compared with diet-resistant mice. Furthermore, switching from high fat to low fat, or high n-3 polyunsaturated fatty acid diet, significantly decreased body weight gain (-35.7% and -31.0%), overall fat storage (-63.4% and -59.6%), and c fos-like immunoreactive neurons in the dorsal part of lateral hypothalamus ( 76.5% and -64.7%) and dorsomedial hypothalamic nucleus (-73.3% and -56.7%) in diet-induced obese mice, respectively. The present study also showed that the ratio of serum leptin/fat mass was threefold higher in the diet-resistant mice than in the diet-induced obese mice, which may be responsible for the less fat storage in the diet-resistant mice. The current data further confirm that the increased neuronal activity in the key autonomic regulatory centers may contribute to the excessive fat storage in diet-induced obese mice. Moreover, both high-fat diet-induced excessive fat storage and the altered hypothalamic neuronal activity may be largely corrected by reducing dietary fat content or replacing it with non-obesogenic fat. PMID- 10856821 TI - Sexually dimorphic cognitive style in rats emerges after puberty. AB - In a water maze (WM), rats employ different and sexually dimorphic behavioral strategies to solve a place-learning task, a test of cognitive/propositional ability. Puberty is the last step in brain development and marks an important phase with regard to sexually dimorphic cognitive performance and behavior. The present study assessed possible sex differences in cognitive style before and after puberty in a WM place-learning task. Since nitric oxide (NO) is implicated in spatial learning and hippocampal function, and since brain NO(-)(2) + NO(-)(3) levels (stable metabolites of NO) display region-specific sex differences in rat brain, NO(-)(2) + NO(-)(3) levels were determined after behavioral testing. The sex-related style difference emerged very clearly but only in the adult rats, which suggests that the female behavioral strategy in the WM place-learning task requires the presence of female sex hormones at puberty. Although NO(-)(2) + NO( )(3) levels were higher in the adult rats and males compared to prepubertal and female rats, respectively, no significant correlations emerged between brain NO and behavior. The fact that the behavioral sexually dimorphic cognitive-style effect observed here and in previous studies appears to emerge only after puberty suggests that awareness of such postpubertal sex differences may also be important in human educational and therapeutic contexts. PMID- 10856822 TI - Fos induction in the Japanese quail brain after expression of appetitive and consummatory aspects of male sexual behavior. AB - We investigated the expression of Fos, the protein product of the immediate early gene c-fos in the brain of male Japanese quail after they engaged in either appetitive or consummatory sexual behavior (i. e., copulation). For 1 h, castrated males treated with testosterone were either allowed to copulate with a female or to exhibit a learned social proximity response indicative of appetitive sexual behavior. Control birds were either left in their home cage or placed in the experimental chamber but did not exhibit the appetitive sexual behavior because they had never learned it. Fos expression was studied with an immunocytochemical procedure in two sets of adjacent sections through the entire forebrain. These sections were immunolabelled with 2 different antibodies raised against a synthetic fragment corresponding to the 21 carboxy-terminal residues of the chicken Fos sequence. Contrary to the results of a previous study in which gonadally intact birds were used, Fos induction was observed neither in the medial preoptic nucleus nor in the nucleus intercollicularis in birds that had interacted for 1 h with a female. This may be related to a lower frequency of copulation in the testosterone-implanted birds than in intact birds, or to differences in the time the brains were collected after the birds engaged in sexual behavior between the two studies (60 min in this study, 120 min in the previous study). The performance of copulation and/or appetitive sexual behavior increased the number of Fos-immunoreactive cells in the ventral hyperstriatum, medial archistriatum, and nucleus striae terminalis. These increases were observed using both antibodies, although each antibody produced minor differences in the number of Fos-immunoreactive cells observed. Using one of the antibodies, but not the other, increases in Fos immunoreactivity were also observed in the nucleus accumbens and hyperstriatum after either copulation or appetitive sexual behavior. These differences illustrate how minor technical variations in the Fos immunocytochemical procedure influence the results obtained. These differences also show that Fos induction in a number of brain regions is observed after performance of consummatory (copulation) as well as appetitive (looking at the female) sexual behavior. This induction is, therefore, not related solely to the control of copulatory acts but, presumably, also to the processing in a variety of telencephalic association areas of stimuli originating from the female. The observation that increased Fos immunoreactivity is present in birds that had learned the response indicative of appetitive sexual behavior, and not in those that had not learned the behavior, further indicates that it is not simply the sight of the female that results in this Fos induction, but the analysis of the relevant stimuli in a sexually explicit context. Conditioned neural activity resulting from a learned association between the stimulus female and the performance of copulatory behavior may also explain some aspects of the brain activation observed in birds viewing, but not allowed to interact with, the female. PMID- 10856823 TI - The effect of isoflurane on the release of [(3)H]-acetylcholine from rat brain cortical slices. AB - Volatile general anaesthetics are believed to affect synaptic transmission, but their actions in the central nervous system (CNS) remains unclear. Acetylcholine (ACh) is one of the most important neurotransmitter in the CNS and thus, it is possible that its release could be one of the targets for volatile anaesthetic action. However, the effects of these agents on the release of ACh are not yet fully understood. Rat brain cortical slices were loaded with [(3)H]-choline in order to study the effect of isoflurane on the release of [(3)H]-ACh from this preparation. Isoflurane (28, 43, 54, 95 and 182 nM) significantly increased the basal release of [(3)H]-ACh. This effect was independent of the extracellular sodium and calcium concentration but was decreased by tetracaine and dantrolene, inhibitors of Ca(2+-)release from intracellular stores. These findings indicate that isoflurane may cause a Ca(2+-)release from internal stores that increases [(3)H]-ACh release in rat brain cortical slices. PMID- 10856824 TI - Lateral parabrachial nucleus lesions in the rat: aversive and appetitive gustatory conditioning. AB - Previous research involving tests of innate preferences and aversions shows that bilateral ibotenic acid lesions of the visceral neurons located in the lateral parabrachial nucleus of the pons selectively disrupt consumption of those gustatory stimuli whose intake is augmented or restricted by their postoral consequences. The present study examined the performance of the same experimental subjects in learned preference and aversion tasks. The lesioned rats failed to develop a conditioned taste aversion (Experiment 1), a conditioned flavor preference (Experiment 2), and a conditioned aversion to the oral trigeminal stimulus, capsaicin (Experiment 3). The pattern of results from both types of taste-guided behaviors (innate and learned) suggests that excitotoxic lesions of the lateral parabrachial nucleus diminish sensitivity to gastrointestinal feedback which, in the present experiments, precludes aversive and appetitive associative learning. PMID- 10856825 TI - Metabotropic glutamate receptor subtypes involved in cardiovascular regulation in the rostral ventrolateral medulla of rats. AB - We have previously reported that metabotropic glutamate receptors (mGluR) participate in cardiovascular regulation in the rostral ventrolateral medulla (RVLM) of rats. In the present study, we have tried to elucidate which subtype of mGluR contributes to cardiovascular responses elicited by L-glutamate in the RVLM. Adult male Wistar rats (348 +/- 11 g, n = 21) were anesthetized and artificially ventilated. Microinjections of agonists and antagonists for each mGluR subtype were done into unilateral RVLM. Each of group I, II and III mGluR agonist (1 nmol/50 nl) produced significant increases in arterial pressure (18 +/ 2, 9 +/- 2 and 34 +/- 3 mmHg, respectively) and heart rate (18 +/- 4, 13 +/- 3 and 33 +/- 12 bpm, respectively). Microinjections of group I, II and III mGluR antagonists failed to inhibit the cardiovascular responses induced by subsequently injected agonists. However, group I antagonist [(RS)-1-aminoindan 1,5-dicarboxylic acid] elicited transient increases in arterial pressure and heart rate, followed by decreases in both variables (-19 +/- 4 mmHg and -22 +/- 4 bpm). These results suggest that all of three subtypes of mGluR participate in cardiovascular responses induced by L-glutamate, and group I mGluR may play an important role in the maintenance of arterial pressure. PMID- 10856826 TI - Previous exposure to footshock stress attenuates nicotine-induced serotonin release in rat striatum during the subsequent stress. AB - We have analyzed the effects of chronic or repeated footshock stress on the release of serotonin (5-hydroxytryptamine: 5-HT) in the striatum of rats that received nicotine by using a microdialysis technique. Neither local infusion of nicotine alone nor stress application alone changed 5-HT release. Local infusion of 1 mM nicotine to the striatum, however, significantly increased 5-HT release in the striatum to 145.9 +/- 30.8 pg/dialysate during simultaneous stress application. These increases of extracellular 5-HT release induced by the combination of nicotine and stress application were also observed in rats that had received daily chronic footshock. However, the previously administered footshock induced the reduced release of 5-HT from the striatum to 33.5 +/- 8. 6 (repeated footshock) and 10.0 +/- 3.6 pg/dialysate (daily footshock) when footshock was given together with nicotine infusion. These results suggest that previous exposure to stress attenuated the nicotine-induced 5-HT release in the striatum during the subsequent stress. PMID- 10856827 TI - An angiotensin system in the anterior hypothalamic area anterior is involved in the maintenance of hypertension in spontaneously hypertensive rats. AB - An overactive brain renin-angiotensin system is one of the factors contributing to the pathogenesis of hypertension in spontaneously hypertensive rats (SHR). We examined brain sites where enhanced activity of an angiotensin system is responsible for the pathogenesis of hypertension in SHR. The angiotensin receptor antagonist, losartan was injected into tissues around rostral parts of the third ventricle in conscious rats. Losartan (0.22 nmol) injected into the anterior hypothalamic area, anterior (AHA) produced a depressor response in SHR but not in Wistar Kyoto rats (WKY). Angiotensin II (0.091-0.91 pmol) injected into the AHA produced a pressor response in both WKY and SHR, and the pressor response to angiotensin II was greater in SHR than that of WKY. Carbachol (3 pmol) injected into the AHA also produced a pressor response in WKY and SHR, and the pressor response to carbachol was almost the same in both strains of rats. Release of angiotensin peptides in the AHA was greater in SHR than that of WKY. These findings suggest that an angiotensin system in the AHA is enhanced and this enhancement of angiotensin system is involved in the maintenance of hypertension in SHR. Both increased pressor reactivity to angiotensin II and increased release of angiotensin peptides in the AHA appear to be related to this enhancement of angiotensin system in SHR. PMID- 10856828 TI - Comparison of G-protein activation in the brain by mu-, delta-, and kappa-opioid receptor agonists in mu-opioid receptor knockout mice. AB - Mice lacking the mu-opioid receptor gene have been developed by a gene knockout procedure. In this study, the activity of opioid receptor coupled G-proteins was examined to investigate whether there is a change in the extent of coupling for mu, delta-, and kappa-opioid receptors in mu-opioid receptor knockout mice. Selective agonists of mu- (DAMGO), delta- (DPDPE), and kappa- (U-69,593) opioid receptors stimulated [(35)S]GTPgammaS binding in the caudate putamen and cortex of wild-type mice. In contrast, only U-69,593 stimulated [(35)S]GTPgammaS binding in these regions of mu-opioid receptor knockout mice. These results confirmed the absence of G-protein activation by a mu-opioid receptor agonist in mu-opioid receptor knockout mice, and demonstrated that coupling of the kappa-opioid receptor to G-proteins is preserved in these mice. However, G-protein activation by the delta-opioid receptor agonist, DPDPE, was reduced in the mu-opioid receptor knockout mice, at least in the brain regions studied using autoradiography. PMID- 10856829 TI - Luteinizing hormone-releasing hormone and oxytocin response to hyperosmotic stimulation: in vitro study. AB - It was shown previously that luteinizing hormone-releasing hormone (LHRH) affects the neurohypophysial oxytocin release in water-deprived rats. However, the detailed mechanisms by which LHRH modifies the oxytocin response to hyperosmotic stimulation have not been explained so far. Using the isolated hypothalamo neurohypophysial explants obtained from euhydrated rats, the effect of LHRH on the oxytocin secretion was studied under conditions of direct osmotic (i.e., Na(+)- evoked) as well as nonosmotic (i.e., K(+)-evoked) stimulation. Additionally, the oxytocin response to LHRH was investigated using the explants obtained from animals drinking 2% saline for eight days (systemic, i. e., both direct and indirect, osmotic stimulation). LHRH significantly enhanced Na(+)- and K(+)-evoked oxytocin release from explants taken from rats drinking tap water, indicating that LHRH could affect the Na(+)/K(+)-dependent depolarization of perikarya of oxytocin neurones. In contrast, LHRH significantly diminished the K(+)-stimulated hormone release when the neurohypophysial complex was obtained from previously salt-loaded rats, suggesting that peripheral osmotic stimulation somehow modifies the sensitivity of oxytocinergic neurones to LHRH (possible mechanisms are discussed). It is concluded that LHRH may participate in the regulation of oxytocin secretion via both direct and indirect impact on magnocellular oxytocinergic neurones depending on the current functional status of the hypothalamo-neurohypophysial complex. PMID- 10856830 TI - Poly(ADP-ribose) polymerase-1: what have we learned from the deficient mouse model? AB - Poly (ADP-ribose) polymerase (113 kDa; PARP-1) is a constitutive factor of the DNA damage surveillance network developed by the eukaryotic cell to cope with the numerous environmental and endogenous genotoxic agents. This enzyme recognizes and is activated by DNA strand breaks. This original property plays an essential role in the protection and processing of the DNA ends as they arise in DNA damage that triggers the base excision repair (BER) pathway. The generation, by homologous recombination, of three independent deficient mouse models have confirmed the caretaker function of PARP-1 in mammalian cells under genotoxic stress. Unexpectedly, the knockout strategy has revealed the instrumental role of PARP-1 in cell death after ischemia-reperfusion injury and in various inflammation process. Moreover, the residual PARP activity found in PARP-1 deficient cells has been recently attributed to a novel DNA damage-dependent poly ADP-ribose polymerase (62 kDa; PARP-2), another member of the expanding PARP family that, on the whole, appears to be involved in the genome protection. The present review summarizes the recent data obtained with the three PARP knockout mice in comparison with the chemical inhibitor approach. PMID- 10856831 TI - Retroviral expression of the hepatitis B virus x gene promotes liver cell susceptibility to carcinogen-induced site specific mutagenesis. AB - Mutational inactivation of the tumor suppressor gene p53 is common in hepatocellular carcinomas (HCC). AGG to AGT transversion in codon 249 of exon 7 of the p53 gene occurs in over 50% of HCC from endemic regions, where both chronic infection with the hepatitis B virus (HBV) and exposure to carcinogens such as aflatoxin B1 (AFB1) prevail. In this study, we report the effect of the HBV x protein (HBx) on carcinogen-induced cytotoxicity and AGG to AGT mutation in codon 249 of the p53 gene in the human liver cell line CCL13. Expression of HBx, as revealed by its transactivation function, results in enhanced cell susceptibility to cytotoxicity induced by the AFB1 active metabolite, AFB1-8,9 epoxide, and benzo(a)pyrene diol-epoxide. Under similar conditions, expression of HBx promotes apoptosis in a subset of cell population. Exposure to AFB1-8, 9 epoxide alone induces a low frequency of AGG to AGT mutation in codon 249 of the p53 gene, as determined by an allele-specific polymerase chain reaction (AS-PCR) assay. However, expression of HBx enhances the frequency of AFB1-epoxide-induced AGG to AGT mutation compared to control cells. In summary, this study demonstrates that expression of HBx enhances liver cell susceptibility to carcinogen-induced mutagenesis, possibly through alteration of the balance between DNA repair and apoptosis, two cellular defense mechanisms against genotoxic stress. PMID- 10856832 TI - Calmodulin antagonists and cAMP inhibit ionizing-radiation-enhancement of double strand-break repair in human cells. AB - Ionizing radiation (IR) enhances double-strand-break (DSB)-repair fidelity in plasmids processed in normal lymphoblasts but not in lymphoblasts from ataxia telangiectasia (A-T) patients. Putatively, signal-transduction pathways mediate this DNA-repair induction. Because IR inhibition of DNA synthesis is defective in A-T cells and is mediated by a calmodulin (caM)-dependent pathway, we evaluated the involvement of caM-dependent pathways in DSB-repair induction. Human lymphoblasts were gamma-irradiated with or without treatment with caM antagonists and the cells' abilities to repair shuttle pZ189 carrying a single DSB (linDNA) were assessed. In untreated controls, IR enhanced DSB-rejoining fidelity if transfection occurred promptly but diminished fidelity if transfection was delayed. Treatment with two caM antagonists, W-7 and W-13, prior to irradiation blocked this IR-enhancement of DSB-rejoining fidelity. Vinpocetine, a caM dependent phosphodiesterase inhibitor, and 8-bromo-cAMP also inhibited IR enhancement of repair fidelity, but caM-dependent protein kinase II inhibitor KN62 had no effect. Other protein kinase inhibitors, staurosporine and genistein, also did not inhibit IR enhancement of DSB repair fidelity. However, staurosporine blocked the twofold reduction in DSB-repair fidelity seen if linDNA transfection was delayed 2 h after irradiation. These findings point to the involvement of caM/cAMP-dependent pathway(s) in mediating IR-enhancement of DSB rejoining fidelity in mammalian cells. PMID- 10856833 TI - Identification of factors interacting with hMSH2 in the fetal liver utilizing the yeast two-hybrid system. In vivo interaction through the C-terminal domains of hEXO1 and hMSH2 and comparative expression analysis. AB - Mutations in DNA mismatch repair (MMR) genes have been shown to segregate with Hereditary Nonpolyposis Colorectal Cancer (HNPCC). However, because many HNPCC families fail to display mutations in known MMR genes, we argued that changes in other components of the MMR pathway may be responsible. The increasing number of proteins reported to interact in the MMR pathway suggests that larger complexes are formed, the composition of which may differ among cell types and tissues. In an attempt to identify tissue-specific MMR-associated factors, we employed the yeast two-hybrid system, using the human hMSH2 as bait and a human fetal liver library as prey. We demonstrate that hMSH2 interacts with a human 5'-3' exonuclease 1 (hEXO1/HEX1) and that this interaction is mediated through their C terminal domains. The hMSH6 protein does not interact with hEXO1 in the two hybrid system. Dot-blot analysis of multiple tissue RNA revealed that hMSH2 and hEXO1 are coexpressed at high levels in fetal liver as well as in adult testis and thymus. Northern blot analysis also revealed that hEXO1/HEX1 is highly expressed in several liver cancer cell lines as well as in colon and pancreas adenocarcinomas, but not in the corresponding non-neoplastic tissue. PMID- 10856834 TI - Levels of peripheral blood cell DNA damage in insulin dependent diabetes mellitus human subjects. AB - Increased production of reactive oxygen species (ROS) in vivo can lead to cellular biomolecule damage. Such damage has been suggested to contribute to the pathogenesis of insulin dependent diabetes mellitus (IDDM). In this study, we used the alkaline comet assay to measure DNA damage (single-stranded DNA breaks and alkali-labile sites) in freshly isolated whole blood, lymphocytes, monocytes, and neutrophils from 23 subjects with IDDM and 32 age- and sex-matched controls. Analysis of the results showed elevated levels of DNA damage (expressed as % comet tail DNA) in the lymphocyte (4.10+/-0. 47, 3.22+/-0.22), monocyte (4.28+/ 0.47, 3.49+/-0.18), and whole blood (4.93+/-0.51, 4.51+/-0.23) fractions from IDDM subjects compared to controls, respectively, but the increases observed were not statistically significant. However, we found significantly elevated basal levels of DNA damage in the neutrophil fraction (8. 38+/-0.64, 4.07+/-0.23; p<0.001, Mann-Whitney U test) in IDDM subjects compared to controls. Given these novel neutrophil findings, we extended the study to include a total of 50 IDDM subjects and 50 age- and sex-matched control subjects and determined basal levels of DNA damage in the neutrophils of all 100 subjects. We found significantly elevated mean levels of DNA damage (8.40+/-0.83, 4. 34+/-0.27; p<0.001, Mann Whitney U test) in the neutrophils from the IDDM subjects when compared to controls. Our results show that even with acceptable glycaemic control there is a significantly elevated level of DNA damage within diabetic neutrophils in vivo. PMID- 10856835 TI - Copper ions mediate the lethality induced by hydrogen peroxide in low iron conditions in Escherichia coli. AB - Iron ions mediate the formation of lethal DNA damage by hydrogen peroxide. However, when cells are depleted of iron ions by the treatment with iron chelators, DNA damage can still be detected. Here we show that the formation of such damage in low iron conditions is due to the participation of copper ions. Copper chelators can inhibit cell inactivation, DNA strand breakage and mutagenesis induced by hydrogen peroxide in cells pre-treated with iron chelators. The Fpg and UvrA proteins play an important role in the repair of DNA lesions formed in these conditions, as suggested by the great sensitivity of the uvrA and fpg mutant strains to the treatment when compared to the wild type strain. PMID- 10856837 TI - Online shopping for ambulatory surgery: let the buyer beware! PMID- 10856838 TI - The paradox of ambulatory surgery in the third world. PMID- 10856836 TI - Gamma-irradiation stimulates homology-directed DNA double-strand break repair in Drosophila embryo. AB - To test the DNA double-strand break (DSB) repair activities present in Drosophila early embryos, we have analyzed the circularization of a microinjected linear plasmid. In order to study repair by homologous recombination, the linear plasmid was injected with an homologous fragment encompassing the break. After extraction from embryos, repair products were analyzed directly by PCR and after their cloning into bacteria. We demonstrate, in addition to the repair by homologous recombination, the presence of an efficient end-joining activity in embryos. Plasmid circularization by end-joining was accompanied by short deletions frequently associated with non-random insertions. Most importantly, pre irradiation of embryos specifically enhanced the accurate repair by homologous recombination. Such a stimulation is described for the first time in the context of a whole higher organism. PMID- 10856839 TI - Patient satisfaction after ambulatory inguinal hernia repair in Hong Kong. AB - Ambulatory surgery was introduced to Asia in the 1990s. Acceptance of ambulatory surgery by oriental patients remains largely unknown. A telephone survey was conducted to evaluate the level of patient satisfaction following ambulatory inguinal hernia repair. A total of 157 patients (61%) completed the telephone survey. More than 90% of the respondents expressed satisfaction with regard to the pre-operative, operative and post-operative service. The majority of the respondents (>80%) preferred to undergo day surgery again in case of hernia recurrence. Our findings prove that ambulatory surgery has a high level of acceptance in Chinese patients and supports the expansion of a day surgery service in Hong Kong. PMID- 10856840 TI - Anxiety management: a distinct nursing role in day surgery. AB - Improved anaesthetic techniques and the increase in minimal access surgery over the past 20 years has had a considerable impact upon the pattern of nursing care required by the surgical patient. In order to adapt to these changes some day surgery nurses have opted for an extension to their role while the majority have expanded their remit and perform nursing interventions within a multi-skilled role. Amid these changing patterns, the nursing profession has been active in conducting research into best practice concerning day surgery. Consequently, a great deal of information is available regarding areas for possible growth. One possible growth area which is also a vitally important issue for patients prior to day surgery is anxiety management. A future nursing role could involve formal anxiety management implicit within a multi-skilled role and as part of an expanded role. The role possibilities are discussed together with an information provision plan as both are central to the effective handling of pre-operative fears. PMID- 10856841 TI - The pain of haemorrhoidectomy: a prospective study. AB - The efficacy of a multimodal analgesic approach for ligation excision haemorrhoidectomy was evaluated in a prospective series of 62 patients. Opioid was given as intravenous fentanyl intra-operatively, as part of a standardised general anaesthetic technique, followed by post-operative parenteral fentanyl or oral oxycodone as required. Pre-emptive local anaesthesia was provided via ischiorectal fossae and haemorrhoid pedicle infiltration. The non-steroidal anti inflammatory drug indomethacin was administered rectally at the end of surgery and regularly orally for 5 post-operative days. A wide range of pain scores was recorded post-operatively but all mean scores were between 2 and 3. Pain was highest at the time of the first bowel action but this was successfully managed in the patient's home. Patient satisfaction with their pain management was achieved in 95% of patients. We conclude that the multimodal analgesia technique combined with pre-operative patient education leads to successful pain control following haemorrhoidectomy. PMID- 10856842 TI - Ambulatory surgery for groin hernia: the Gilbert repair. AB - The aim of this study was to evaluate the results of the Gilbert repair for primary treatment of indirect inguinal hernias performed as day cases. From September 1996 to September 1998, 145 patients who were admitted for ambulatory surgery underwent Gilbert tension-free repair for treatment of unilateral inguinal hernia. Sex, age, the American Society of Anaesthesiologists (ASA) preoperative assessment score, type of anaesthesia, operating time, postoperative recovery, postoperative pain, morbidity, mortality, recurrence, return to work and the normal daily activities were assessed. The mean follow-up was 21 months (range 12-36). Gilbert's classification, type 2 and 3 hernias were the most common. Spinal anaesthesia was used in 73% of patients. Mortality was zero. Four patients developed postoperative haematomas, two urinary retention, three seromas, and two wound infections. During the follow-up period, only two recurrences of hernia were noted (1.4%). In conclusion, these data show that Gilbert repair is a safe operation, which is simple to learn. It can be performed on an outpatient basis, with a low complication rate, a low level of pain and a short recovery period. Although it seems to have a low risk of recurrence, a long term follow-up is needed. PMID- 10856843 TI - One-week recovery profiles after spinal, propofol, isoflurane and desflurane anaesthesia in ambulatory knee arthroscopy. AB - There are comprehensive findings on the immediate recovery of patients from different types of anaesthesia, but more information is needed on how patients manage at home after ambulatory surgery. One hundred and seventy-three elective knee arthroscopy patients were randomised into four different anaesthesia groups to receive either spinal anaesthesia (SA) with 5% lidocaine or general anaesthesia (GA) with propofol infusion, isoflurane inhalation or desflurane inhalation. The patients were interviewed over the phone on the next day and asked to complete a questionnaire after 1 week. One hundred and sixty-eight patients (97%) were reached by phone. The questionnaire was returned by 163 patients (94%). After 24 h, all the patients were satisfied with the type of anaesthesia they had received, but 2% of the SA patients would have chosen GA and 4.3% of the GA patients would have chosen SA for the next operation. Based on the questionnaires returned after 1 week, 8.3% of the SA patients would have wanted to have GA, and 4.7% of the GA patients would have wanted to have SA in the future. The incidence of nausea (4.2%) and vomiting (1.8%) was very low in the whole series, with no differences between the anaesthesia groups. Headache after 24 h was experienced by 15.7% of the SA and 10.3% of the GA patients. After 1 week, SA patients reported headache upon standing in 13.5% of the cases, backache in 36.5% and lower leg pain in 59.6%. The corresponding figures for GA patients were 4.5, 9.9 and 39.6% (P<0.05). In spite of the good immediate recovery profile in the all anaesthesia groups, the fact that SA patients reported a higher incidence of headache, backache and lower leg pain after 1 week may be signs of post spinal headache and transient neurologic symptoms (TNS). For overall patient comfort, GA might be a better anaesthetic choice in ambulatory surgery. PMID- 10856844 TI - Day surgery for older people (70+): selection versus outcome effects. AB - As length of Australian hospital stays decreased, concerns were raised about benefits of shorter stays for older people. We investigated personal characteristics, perceived health outcomes (SF-36) and service use of day-only and other patients aged 70+, at one and 12 weeks after hospital discharge. Day only patients were younger, had better self-reported health, were selected for orthopaedic, gastrointestinal and ophthalmic procedures and used similar levels of formal and informal services after discharge as people with longer stays. There was no evidence of ill effects of day surgery for older people, but improved selection and information giving procedures can improve outcomes. PMID- 10856845 TI - Premedication with EMLA cream for ambultory surgery in children. AB - Objectives: The objective of this study was to confirm the ability of EMLA(R) cream (Eutectic Mixture of Local Anaesthetics, Astra, Sweden) to provide effective dermal analgesia after topical application on the skin of the dorsum of the hand 1 h before venous cannulation for anaesthetic induction. Material and Methods: Prospective, randomized, double blind study. We included 100 children, ASA I-III, distributed into three groups: Group EMLA (E, n=34), Placebo (P, n=33) and Control (C, n=33). Results: The EMLA group of patients (E) refered an evaluation of pain (visual analogical scale (mean=2.34+/-2.41), significantly smaller than the other groups (Placebo=5.54+/-3.40, Control=6.03+/-2.77). Conclusion: EMLA cream, when topically applied 1 h before venous cannulation, provides effective dermal analgesia for venous cannulation. No general or local adverse reactions were observed. PMID- 10856846 TI - Quality indicators in ambulatory surgery. A prospective study. AB - Introduction: Ambulatory Surgery has emerged and developed in Spain in a similar way as in other occidental countries. Once established as a model for surgical care, results must be improved using objective quality indicators to assure minimal morbidity, efficient use of resources and satisfaction of patients and family. Objective: To provide a list of quality assessment indicators in Ambulatory Surgery and to discuss the necessity of obtaining a general consensus regarding the complexity of both the operations and the patients undergoing surgery. Design: Prospective and descriptive study. Patients and Methods: The study was performed on 833 ambulatory patients operated on in the Ambulatory Surgical Unit of the Can Misses Hospital of Ibiza. Surgical specialities involved were General Surgery, Ophthalmology, Orthopaedics, Urology, Gynaecology, and E.N.T. All diagnoses and proceedings were classified according to the CIE-9 and DGR systems, showing as an example the codes related to the General Surgery department. Quality Indicators evaluated were: (a) unplanned admission index; (b) re-admission index; (c) emergency department consultations; (d) cancellation index; (e) substitution index; and (f) postoperative pain evaluation. Results: Specialities involved were: Ophthalmology 277 patients (33.2%), General Surgery 189 (22.8%), Orthopaedics 142 (17%), Urology 90 (10.8%), E.N.T. 79 (9.5%) and Gynaecology 56 (6.7%). Unplanned Admission Index was 7.2%; Re-Admission Index 0.4%; Emergency department consultation 1.6%; Cancellation Index 3.3% and Substitution Index 70%. The reasons for unplanned admission were due to specific complications in 36 cases (60.1%), followed by those secondary to an inadequate selection of the patient in 22 cases (36.6%). Conclusions: It is necessary to apply a series of Quality Indicators in Ambulatory Surgery. Their systematic evaluation may help us define national standards in order to continuously improve our results. PMID- 10856847 TI - Outpatient surgical unit: critical review after the first year of operation. AB - Introduction: Outpatient surgery has been quickly accepted by many hospitals, showing a lot of advantages, both in the quality of health care and also in hospital management. Following this trend, our Hospital started operating an Outpatient Surgical Unit in May 1997. Objectives: Once we had reached our 'cruising speed', we thought it wise to make a critical review of our experience. Material and methods: We have reviewed 331 patients who underwent surgery during the first year of operation, with special attention to the acceptance of this type of surgery by the population in general, and to the changes in pre-operative and post-operative protocols that this process has caused. Results: The rate of admission was 3.02%, and all of them occurred after 24 h. We found 57 minor complications (spotty dressing, mild inflammatory signs) on the first day assessment, but only 16 patients had refered complications. 91.5% of the questionnaires sent back by the patients qualify the experience as good or excellent. Conclusions: Outpatient Surgery is a good solution for many surgical pathologies, as it is well accepted by the general population. Inclusion and exclusion parameters regarding age, architectural obstacles and post-operative care can become selectively more lax. PMID- 10856848 TI - Contribution of the outpatient surgery unit ITO the general surgery department of a district hospital. AB - Introduction: The creation of Outpatient Surgery (OPS) units to combine the quality of medical attention and rationalize costs allows for greater efficiency in the use of resources. Aim: To report our series of patients undergoing surgery at the OPS units integrated into our Hospital (Type II): Patients and method: Between May 1994 and March 1998, 832 outpatients, of a total of 5230, underwent surgery at our General Surgery Unit. The criteria for exclusion from the programme depended on the patient and the enviroment or resulted from the operation itself. Results: Mean patient age was 47.5 years; there were 420 males and 412 females. Surgery was performed for 229 inguinofemoral hernias, 47 umbilical-epigastric hernias, nine incisional hernias, 193 pilonidal sinuses, 156 mammary nodules, 65 varicose veins, 64 arteriovenous fistulae and 69 proctology operations. The most common anesthesia techniques performed were rachianesthesia and local anesthesia. Eight point seven percent of the patients required admission (OPS failure), the most frequent causes being excessive pain, orthostatic-syncopal hypotension, nausea and vomiting and urine retention. There was no morbidity or mortality. Conclusion: OPS is a highly efficient procedure for resolving the most common pathologies in General Surgery. The anesthesia technique was an important factor in the rate of failure. PMID- 10856849 TI - Treatment of the abdominal wall defects in an ambulatory surgical setting: our experience. AB - Introduction: The creation of Outpatient Surgery (OPS) units has allowed to reduce the costs and the waiting lists in an efficient fashion. We describe our series of patients operated on for abdominal wall defects, a pathology suitable for ambulatory surgery. Patients and methods: Between May 1994 and March 1998, 206 inguinal hernias, 23 femoral hernias, 47 umbilical-epigastric hernias and nine incisional hernias were operated on in an ambulatory surgical setting. The patients were selected following the selection criteria previously established (related to the patient, the environment and the surgical procedure). The average age was 45 years, and the distribution by sex, 210 men and 75 women. Spinal anesthesia was preferently performed. The surgical techniques employed were Lichtenstein's hernioplasty and Shouldice and Bassini procedures for inguinal hernias; Lichtenstein's plug technique for femoral hernias and simple closure or preperitoneal mesh for the middle line defects. Results: 44 patients needed readmitttance to hospital (failure of OPS), the most important causes being excessive pain, urinary retention and nausea/vomiting. There was no severe morbidity nor mortality. Conclusion: Surgery for abdominal wall defects constitutes a group of procedures suitable for efficient and low risk OPS programs. PMID- 10856850 TI - Reconstruction of the RVOT with valved biological conduits: 25 years experience with allografts and xenografts. AB - OBJECTIVE: The reconstruction of the RVOT in congenital heart disease often requires the implantation of a valved conduit. Although allografts are considered the conduit of choice their availability is limited and therefore xenografts are implanted as well. We compared the long-term durability of both grafts in the RVOT over a 25-year period. METHODS: Between January 1974 and August 1999, 505 patients (median age 4.0 years, range 2 days-31 years; median weight 14.5 kg, range 2.2-76.6 kg; median body length 103 cm, range 48-183 cm) with congenital malformations (PA 25.3%, TOF 14.5%, TOF+PA 2.4%, DORV 4.2%, TGA+PS 8.7%, TAC 24.8%, and other 20.2%) received their first valved conduit (174 xenografts: median diameter 14 mm, range 8-27 mm; 331 allografts: median diameter 19 mm, range 8-30 mm). RESULTS: Follow-up is 3017 patient-years. The 10-year survival probability for all patients. was 66% with a mean reoperation-free interval for conduit-exchange of 13.3 years (mean reoperation-free interval for allografts, 16.0 years; mean reoperation-free interval for xenograft, 10.3 years). One hundred and thirteen patients underwent a conduit-exchange, mostly due to conduit stenosis. Fourteen patients had a second exchange and three patients a third exchange. For patients with conduit diameters <18 mm (n=235: allograft n=116, xenograft n=119; median age 9 months, range 0-27.3 years), the mean reoperation free interval was 11.2 years (mean interval allograft, 13.1 years; mean interval xenograft, 8.6 years, P=0.03). For conduit diameters >/=18 mm (n=270: allograft n=215, xenograft n=55, median age 7.4 years, range 0-34.3 years) the mean interval from freedom of conduit exchange was 15.1 years (for allografts 14.1 years, for xenografts 12.5 years, P<0.01). Comparing xenografts to allografts, we found no difference in patient survival probability (P=0.62). There was no significant difference between antibiotic (n=198) preserved vs. cryopreserved (n=133) allografts (P=0.06). Blood group compatibility of allografts to recipients had no significant influence on allograft function (P=0.42). The donors allograft origin, whether aortic or pulmonary valve, had also no significant influence on allograft long-term function (P=0.15). CONCLUSION: For the reconstruction of the right ventricular outflow tract (RVOT) allografts show significantly better long-term durability than xenografts regardless of the age at implantation and the diameter. PMID- 10856851 TI - Tetralogy of Fallot: what operation, at which age. AB - BACKGROUND: The optimal management of tetralogy of Fallot is still under debate, particularly with respect to surgical approach and the age of operation. In recent times a transatrial-transpulmonary approach and primary repair in younger patients is favoured. The purpose of the present study was to analyze the result of our current surgical management by assessing the perioperative and intermediate term follow up in order to define the optimal strategy and timing of operation for our institution. METHODS: One hundred and thirty two patients with tetralogy of Fallot who underwent definitive repair between May 1993 and December 1998 were analyzed by reviewing their medical records and follow-up. Median age was 15. 5 (2.3-68.6) months and median weight was 8.8 (5-16) kg. Ten (7.57%) patients were under 6 months, 38 (28.78%) were between 6 and 12 months, 36 (27.27%) were between 12 and 18 months, 23 (17.42%) were between 18 and 24 months and 25 (18.93%) were more than 24 months age. During the study period there was a move to earlier surgery and differing methods of repair depending on the anatomy observed. Follow up was conducted by the referring cardiologist. Median follow up was 35.48 (8.07-74.93) months. RESULTS: Forty-two (31.8%) patients required a palliative procedure before total correction due to unfavourable anatomy. Subpulmonary infundibular obstruction with a fibrous component increased significantly with age (P<0.05). Operations were entirely transatrial in 14 (10.6%), transatrial and transpulmonary in 69 (52.2%), transatrial and transventricularly in 42 (31.8%) and a homograft conduit was used in seven (5.3%) patients. Younger patients had narrower pulmonary valves and required a transannular patch more frequently. All patients were in sinus rhythm, 28 (21.1%) showing right bundle branch block. Median hospital stay was 8 (5-54) days. No patient required reintervention during follow up and there was no early or late mortality. CONCLUSION: Correction of tetralogy of Fallot at younger age does not increase morbidity or mortality and has potential advantages. A surgical technique adapted to the anatomy of the right ventricular outflow tract, achieves the best results. PMID- 10856852 TI - Surgical management of complex and tunnel-like subaortic stenosis. AB - BACKGROUND: Relief of primary or secondary subaortic stenosis (SAS) remains a surgical challenge. Heart block, aortic valve regurgitation and recurrent obstruction have been persistent problems. METHODS: Forty six patients who underwent surgery for complex and tunnel-like SAS between January 1990 and November 1998 were reviewed. In 45 of the 46 patients SAS developed following repair of a primary congenital heart defect and only one patient presented with de novo tunnel-like SAS. Fifteen of the 45 patients had undergone repair of double-outlet right ventricle (DORV) and the remaining 30 had undergone repair of a variety of defects. The median age at the time of surgery was 5 years. The modified Konno procedure was performed in 15 patients, Konno procedure in three, Ross-Konno procedure in two and resection of the conal septum in 12 patients. Five patients with DORV underwent replacement of the intraventricular baffle and two patients underwent an aortic valve-preserving procedure in conjunction with mitral valve replacement. RESULTS: There were no deaths. None of the patients had an exacerbation of aortic regurgitation and none developed complete heart block. The median follow-up was 3 years (range 1 month-8.5 years). Two patients developed recurrent SAS defined as a gradient of 40 mmHg or greater diagnosed by transthoracic echocardiography. Freedom from SAS at 1, 3 and 5 years was 100, 94 and 86%, respectively. CONCLUSIONS: We favor the modified Konno procedure and conal resection to the Konno or the Ross procedure, since insertion of a prosthetic valve or homograft is avoided and aortic valve function is preserved. Excellent relief of tunnel-like SAS can be achieved without damage to the conduction tissue. PMID- 10856853 TI - Primary repair of aortic arch obstruction with ventricular septal defect in preterm and low birth weight infants. AB - OBJECTIVE: Previous reports have suggested that prematurity and low birth weight are risk factors for definitive surgical intervention in congenital cardiac malformations. The following data review our experience with primary repair of the complex malformation of aortic arch obstruction with ventricular septal defect (VSD) in this patient population. METHODS: Since 1988, 21 consecutive preterm (/=20% increase, which declined to preoperative values after 1 year; group C, (n=18) FeV1, 20-40% increase, sustaining at 1 year; group D, (n=21) FeV1>/=40% increase, sustaining at 1 year. The statistics comprised of analysis of variance (ANOVA) and chi-square testing, with values presented as means+/-SEM. RESULTS: No differences were found for lung function parameters (FeV1: 27.7+/-2.7, 26.0+/-2.5, 23. 9+/-2.2 and 23.9+/-1.9% predicted, in groups A, B, C and D, respectively). Arterial blood gas levels preoperatively revealed significant differences between the groups; the arterial pO(2) was 66.2+/-1.2 mmHg in groups A+B compared with 61.8+/-1.5 mmHg in groups C+D (P=0.030). The arterial pCO(2) was 39.2+/-1.1 mmHg in groups A+B compared with 43.3+/-1.5 mmHg in groups C+D (P=0.038). The morphometric data had a strong trend towards higher heterogeneity in groups C and D. Marked DHI was found in 59 and 81% of patients in groups A+B versus C+D, respectively (P=0.121). Marked DHG was present in 22 and 54% of patients in groups A+B versus C+D, respectively (P=0.010). CONCLUSION: Preoperative arterial pO(2) and pCO(2), and the DHG are predictors for long-term benefit after LVRS with regard to the FeV1, 1 year postoperatively. PMID- 10856858 TI - Risk analysis for thoracoscopic lung volume reduction: a multi-institutional experience. AB - OBJECTIVE: Most reports of thoracoscopic lung volume reduction (TLVR) are relatively small and early experiences from a single institution, factors which limit both the statistical validity and the applicability to the population at large. In order to address these shortcomings we undertook an analysis of the TLVR experience at five separate institutions to assess operative morbidity and identify predictors of mortality. METHODS: Questionnaires were sent to four groups of surgical investigators at five institutions actively performing TLVR. Data was requested regarding preoperative, operative and postoperative parameters. Twenty-five potential predictors of mortality were analyzed and seven proved to be at least marginally significant (P<0.10). These parameters were entered into a stepwise logistic regression analysis to identify independent predictors. RESULTS: The 682 patients (415 males, 267 females, mean age 64.0 years) underwent unilateral (410) or bilateral (272) TLVRs. Overall, operative mortality was 6% with half of the deaths resulting from respiratory causes. The remaining patients were discharged to home (88%), a rehabilitation facility (4%) or a ventilator facility (2%). There were 25 perioperative factors chosen representing clinically important indices such as spirometry, oxygenation, functional status, clinical and demographic variables. Univariate analysis identified seven variables as predictors of mortality (P<0.10) and these were entered into a stepwise logistic regression analysis. Only age, 6-min walk, gender (male 8%, female 3% mortality) and the procedure performed (unilateral 4.6%, bilateral 8%) were independent predictors while preoperative steroid therapy, preoperative oxygen administration, and time since smoking cessation dropped out of the model. The specific institution, learning curve (early vs. late experience), type of lung disease, spirometric indices and predicted maximum VO(2) were not significant predictors. CONCLUSION: This experience suggests that unilateral and bilateral lung volume reduction procedure can be performed with acceptable morbidity and mortality. Although age, gender, exercise capacity and the procedure performed are all independent predictors of mortality, the risk of operative death did not appear excessive in this fragile patient subset. PMID- 10856859 TI - Morphologic grading of emphysema is useful in the selection of candidates for unilateral or bilateral reduction pneumoplasty. AB - OBJECTIVE: Radiologic morphology of emphysema proves useful in the selection of candidates for bilateral reduction pneumoplasty. We developed a simple morphologic grading system capable of identifying subsets of patients who had maximal functional improvement after unilateral or bilateral operation. METHODS: Fifty-two patients who underwent unilateral (n=34) or bilateral (n=18) reduction pneumoplasty were evaluated. Emphysema morphology was visually scored by digital roentgenograms and high-resolution computed tomography. In each lung, severity of emphysema (ES), heterogeneity (DHT) and hyperinflation (DHF) degrees, were assessed. Asymmetric ratio of emphysema (ARE) between the lungs was expressed as: higher ES/lower ES scores. Morphometric data were correlated with absolute preoperative-postoperative FEV(1) change (DeltaFEV(1)). RESULTS: No difference was found between the unilateral and the bilateral group for ES and DHT. DHF was greater in the bilateral group (3.1 vs. 2.7, P=0.02) whereas ARE was greater in the unilateral group (1.29 vs. 1. 05, P=0.0001). Stepwise logistic regression extracted as best predictors of maximal DeltaFEV(1), ARE (odds ratio=238, Wald test P=0.04) in the unilateral group, and DHT (odds ratio=24, P=0.03) in the bilateral group. Unilateral group DeltaFEV(1) was greater in patients with ARE>/=1.3 (0.44 vs. 0.24 l, P=0.02). Bilateral group DeltaFEV(1) was greater in patients with DHT>1 (0.50 vs. 0.31 l, P=0. 03). No difference was found when comparing DeltaFEV(1) resulting from unilateral RP and ARE>/=1.3, and bilateral RP (0.44 vs. 0.41 l, not significant). CONCLUSIONS: This morphologic grading system identified subsets of patients who had maximal functional benefit from unilateral or bilateral reduction pneumoplasty and might be useful in the preoperative screening of candidates for either approach. PMID- 10856860 TI - Chronic sequels after thoracoscopic procedures for benign diseases. AB - OBJECTIVE: Chronic pains after lateral thoracotomy are present in up to 40% of cases. Chronic sequels after thoracoscopy are less common, but nevertheless, a cause for complaints by patients. Pain often reflects a recurrence of malign disease. For this reason, we only investigated patients with benign disease. METHODS: We retrospectively investigated the incidence of chronic sequels in a consecutive series of 161 patients who underwent thoracoscopy for benign disease and were not converted to an open procedure. The data from all 144 patients, contactable at the time of investigation, who were at least 2 months postsurgery, were analyzed. RESULTS: Chronic sequels were present in an overall of 31.4% of patients. Patients complained of chronic pain (20.1%), numbness distal to the incision sites (16.9%) and disaesthesia (8.3%). Painkillers are taken on a regular basis by 82.8% of patients with chronic pain. The use of Staplers, as well as the number of drains (1 vs. 2) used, were statistically significant (P>0.05) for chronic sequels. All other investigated factors, such as sex, age, and length of drainage, were not significantly different in the two groups. CONCLUSION: The thoracoscopic approach is not likely to impact on the prevalence of long-term postthoracotomy sequels, and therefore, further strengths are necessary to reduce this number. PMID- 10856861 TI - Ultra-thin needle thoracoscopic surgery for hyperhidrosis with excellent cosmetic effects. AB - BACKGROUND: In spite of its cosmetic benefits over open surgical techniques, endoscopic sympathectomy using 5 mm or larger instruments still has the problems of operative scar as well as pain on the trocar sites. Recently we have begun using 2 mm endoscopic instruments. The purpose of this study was to confirm the safety and feasibility of fine needle endoscopic instruments in thoracic sympathetic ablation. METHODS: We have exclusively used 2 mm endoscopic instruments since January 1997, and from that time to May 1999 417 patients were underwent surgical procedures for hyperhidrosis. T2 or T2/T3 sympathectomy was performed for the first 56 patients, after June 1997, in 361 patients the interconnecting sympathetic trunk was divided instead of ganglion resection, and this procedure was named sympathicotomy. Palmar hyperhidrosis was presented in 375 patients (89.9%) and facial in 28 (6.7%) and axillary in 14 (3.4%). The level of division or resection of the ganglion differed according to the patient's symptoms. RESULTS: Sympathicotomy and sympathectomy were successful and all patients were satisfied with immediate dryness of affected sites. There were not any cases of bleeding or reoperation or hospital mortality. A large endoscope was required to eliminate the pleural adhesion in fourteen cases (7.7%). Thoracotomy conversion was required in two pleural adhesion cases. Minor complications were occurred in 17 patients (4.1%); such as closed thoracostomy in ten cases, peripheral nerve injury in three, pulmonary parenchymal injury in two, Horner's syndrome in two and atrial fibrillation in one. We have five cases of recurrent symptoms (1.2%). CONCLUSION: Our experience indicates that, for the treatment of hyperhidrosis, 2 mm ultra-thin needle endoscopic instruments are safe and effective to operate on palmar and facial hyperhidrosis patients. PMID- 10856862 TI - 'Needlescopic' video-assisted thoracic surgery for palmar hyperhidrosis. AB - OBJECTIVE: The video-assisted thoracic surgery (VATS) approach for thoracodorsal sympathectomy has been well accepted. We report the use of ultra-fine thoracoscopic equipment for this procedure, based on the experience from two centers in Asia. MATERIALS AND METHODS: Thirty-eight patients with palmar hyperhidrosis underwent bilateral VATS thoracodorsal sympathectomy using 2-mm instruments exclusively. General anesthesia with selective one lung ventilation was used. Carbon dioxide insufflation was used when lung collapse was found to be inadequate. In 11 patients, the sympathetic chain was excised (T2-T3 for palmar hyperhidrosis alone, extending to T4 for axillary hyperhidrosis), and in 27 patients, the chain was cauterized. The choice of procedure reflects the surgeon's preference. No chest drains were left after the procedure and no stitching of the wound was necessary. RESULTS: There was no mortality or major complications. A small pneumothorax was found in the postoperative chest X-ray in three patients. They all resolved without further intervention. Twenty-seven patients were discharged on the same day of admission, and 11 patients were discharged on postoperative day one. After an average follow-up of 16 months (range 5-28), there has been no recurrence of symptoms. Compensatory truncal hyperhidrosis was encountered in two patients, but the symptoms were not severe enough to interfere with lifestyle, and this required no further treatment. CONCLUSION: Thoracodorsal sympathectomy using 2-mm instruments is technically feasible and is associated with an excellent clinical outcome. Limitations of the equipment, however, exist (narrow field of vision, lower resolution and difficulty in maintaining fine control), and we are currently restricting its use to relatively simple procedures. PMID- 10856863 TI - Esophagectomy and staged reconstruction. AB - OBJECTIVE: Esophageal resection with diversion and staged reconstruction of the upper gastrointestinal (GI) tract is an option in the management of complex problems. This study characterizes circumstances, indications, outcomes and their predictors for staged reconstruction, and estimates the optimal timing for reconstruction. METHODS: Between October 1981 and March 1999, 43 patients were identified with planned staged reconstruction. Twenty-six had esophageal cancer, and 17 had complications of benign disease. Primary diversion with esophageal resection was needed in 16 patients, and secondary diversion with takedown of previous esophageal reconstruction was needed in 27. Common indications were failed esophageal anastomosis and esophageal perforation. Death before and death after reconstruction were considered as competing risks. Multivariable analyses were used to estimate the optimal timing of reconstruction. RESULTS: The survival was 75, 21 and 9% at 3 months, 5 and 10 years, with survival only somewhat better (P=0. 06) among patients having benign versus malignant disease. A similar proportion of patients died before reconstruction as underwent reconstruction, resulting in only 17 reconstructions, typically 9 months after diversion. The risk factors for death included cancer and primary diversion. The survival was best for benign disease when reconstruction was early. The survival was poor after reconstruction in the few patients with malignant disease. CONCLUSIONS: Patients requiring staged esophageal reconstruction are heterogeneous, with malignant or benign disease, and primary or secondary diversion. The outcome is poor, and is influenced by the pathology and timing of diversion. Patients with benign disease should be reconstructed as early as feasible; reconstruction is rarely indicated for patients with cancer. PMID- 10856864 TI - Limited right anterolateral thoracotomy for mitral valve surgery. AB - OBJECTIVE: There has been great enthusiasm in recent years to perform mitral valve surgery through small multiple incisions with the use of the Port Access technique. The procedure is costly, involves a relatively long training curve and leaves the patient with multiple scars in the chest and groin. We used a mini thoracotomy technique for mitral valve patients and compared our results with the conventional technique. METHODS: We randomized 100 consecutive patients presenting to our practice for mitral valve surgery between two groups. The first group (test group) consisted of 50 patients in which mitral valve surgery was performed via mini-right anterolateral thoracotomy approach. The control group (50 patients) underwent classical mitral valve surgery through median sternotomy. Standard aortic and bicaval cannulation with antegrade blood cardioplegia was adopted in both groups. RESULTS: There was no statistical difference between the two groups preoperatively regarding their age, pathology, LV function and male/female ratio. Most of the patients had valve replacement except four in the test group and three in the control group. The incision in the test group was 12 15 cm long in the right submammary groove. Direct aortic cannulation, clamping and cardioplegia administration was achieved in all patients easily. The mean bypass time was slightly longer in the test group (64+/-12 min) when compared with the test group (59+/-11 min). The cross-clamp time was lower in the test group (27+/-8 min) when compared with the control group (31+/-9 min). There was no hospital mortality in both groups and there was one morbidity in the form of sternal infection in the control group. The mean hospital stay was similar for both groups (7+/-2 days). CONCLUSION: The cosmetic appearance in the test group was excellent and the patients' wounds were scarcely apparent in the female patients. The study demonstrates the efficacy and safety of this older technique, with excellent cosmetic results and no additional cost or risk to the patients. PMID- 10856865 TI - Do patients want minimally invasive aortic valve replacement? AB - OBJECTIVE: Access to aortic valve can be performed through small incisions. However, a considerable advantage of this approach has not been proven by randomized studies so far. We wanted to elucidate the opinion of patients when they are informed objectively about advantages and disadvantages of minimally invasive approach prior to operation. METHODS: This prospective study was performed with 27 patients undergoing isolated aortic valve replacement. These patients were informed prior to operation by the same resident concerning objective data. A photograph was shown illustrating a patient with postoperative wound after a standard- and a mini-incision, respectively. After the interview the patient could decide between full and partial sternotomy. RESULTS: After the interview 21/27 (78%) patients preferred to have a full sternotomy (group F) and 6/27 (22%) patients (group P) decided to have a partial sternotomy. Comments of group F: surgeon should have best exposure (n=15); cosmetics aspects unimportant (n=14); operation time as short as possible (n=7). Group P: cosmetic aspects important (n=6). Significant differences between groups (group F vs. group P): age (years), 69.1+/-1.5 vs. 49.2+/-7.3 (P=0.024); operation time (min), 142+/-7 vs. 189+/-15 (P=0.002); CK (IU/l), 111+/-11 vs. 374+/-114 (P=0.0007); CKMB (IU/l), 17+/-2 vs. 45+/-17 (P=0.006); ICU-stay (days), 2.6+/-0.2 vs. 3.2+/-0.2 (P=0.044). Pericardial effusion requiring drainage was observed in two patients of group P. One patient of group P suffered myocardial infarction. CONCLUSION: When patients are informed objectively about advantages and disadvantages of minimal invasive aortic valve surgery only a smaller number decides to have a mini incision. The patients preferring short incisions are significantly younger since cosmetic aspects are more important. Longer duration of operation may be due to longer hemostasis based on limited exposure. Air bubbles due to inadequate de-airing might be responsible for higher CK and CK-MB levels in group P. PMID- 10856866 TI - The left atrial appendage: our most lethal human attachment! Surgical implications. AB - OBJECTIVES: To prevent death from atrial fibrillation, a cardiac disease which kills by producing emboli. Atrial fibrillation causes about 25% of strokes and increases stroke rate by five times. Over 90% of these embolic strokes are from clots originating in the left atrial appendage. This study addresses the surgical feasibility of removing the appendage to prevent future deaths in two subcategories of patients. (1) Prophylactic removal during open-heart surgery to study its safety. Theoretically, as these patients age and some develop atrial fibrillation, protection from embolic strokes would already be present. (2) Therapeutic removal in chronic atrial fibrillation patients by means of a thorascopic approach. Its technical feasibility is demonstrated. Its actual stroke prevention potential awaits large studies. METHODS: Appendectomy has been evaluated three ways. (1) Experimentally, thorascopic appendage removal was performed on 20 goats with endoscopic approach. Late studies showed a cleanly healed atrial closure after stapling, and no puckering of tissue as seen with the purse-string approach. (2) Safety of human appendectomy was demonstrated in 437 patients (1995-1997). Routine appendectomy was performed during open-heart surgery. Forty-three appendages were stapled, 391 sutured off. (3) Thorascopic appendectomy in seven patients with chronic atrial fibrillation has been successfully accomplished as an isolated surgical procedure. Stapling or suture closure provides a much cleaner, non-puckered suture line than a purse string. RESULTS: In prophylactic removal, no acute bleeding occurred. No late problems have been identified. Endoscopic removal of the appendage has been successful in seven atrial fibrillation patients. CONCLUSIONS: The left atrial appendage is a lethal source of emboli in atrial fibrillation patients. As patients age and often develop atrial fibrillation, prophylactic appendage removal whenever the chest is open is suggested as a method to prevent future strokes. In chronic atrial fibrillation patients, appendectomy can be done with a mini-thorascopic approach. Further studies are planned to demonstrate the effectiveness of appendectomy in preventing strokes in the chronic fibrillating patients. PMID- 10856867 TI - Expression of adhesion molecules and cytokines after coronary artery bypass grafting during normothermic and hypothermic cardiac arrest. AB - OBJECTIVE: Cardiac surgery with cardiopulmonary bypass (CPB) results in vascular injury and tissue damage which involves leukocyte-endothelial interactions mediated by cytokines and adhesion molecules. This study was designed to demonstrate the effect of normothermic and hypothermic CPB to cytokine and soluble adhesion molecule levels in adults and to determine whether these levels correlate to the patients postoperative course. DESIGN AND PATIENTS: In 25 patients after normothermic and in 25 patients after hypothermic coronary artery bypass grafting with cardiopulmonary bypass (CPB), blood samples for cytokine and soluble adhesion molecule analysis were taken preoperatively, 24, 36, 48 h, and 6 days postoperatively. Soluble adhesion molecules (sE-selectin, sICAM-1) were measured by ELISA and cytokines (TNF-alpha, IL-6, IL-8) by chemilumenscent immunoassay. Clinical data were collected prospectively. RESULTS: Postoperatively, adhesion molecule and cytokine levels were significantly elevated after CPB. Mean plasma levels of sICAM-1 was 2.4-fold higher after 6 days. Mean plasma concentration of sE-selectin peaked after 48 h with a 2-fold increase compared to normothermic conditions. In the hypothermia group sICAM-1, sE-selectin, IL-6, and IL-8 showed significantly higher levels (P<0.0057, P<0.0012, P<0.0419, P<0.0145) after 24 h compared to the normothermia group. No clinical differences were seen. CONCLUSION: Adhesion molecules and cytokines are elevated after CPB. Patients after hypothermic CPB show significant higher sICAM 1, sE-selectin, IL-6, and IL-8 levels after 24 h compared to normothermic conditions. These results are mainly due to longer CPB and crossclamp times but do not alter the patient's postoperative course. PMID- 10856868 TI - Reduction of pro-inflammatory cytokine levels and cellular adhesion in CABG procedures with separated pulmonary and systemic extracorporeal circulation without an oxygenator. AB - OBJECTIVE: We have recently shown that a considerable amount of pro-inflammatory cytokines is released during pulmonary passage after aortic declamping in patients undergoing coronary artery bypass grafting. The present study was performed to investigate whether bilateral extracorporeal circulation with the lungs as oxygenators can reduce the inflammatory responses of the lungs. METHODS: Eighteen consecutive patients undergoing coronary artery bypass grafting were randomly assigned to routine extracorporeal circulation with cannulation of right atrium and aorta (routine circulation, ten patients) or to a bilateral extracorporeal circulation with additional cannulation of left atrium and pulmonary artery (bilateral circulation, eight patients). Blood was simultaneously drawn from right atrium and pulmonary vein at 1, 10 and 20 min reperfusion. The levels of interleukin (IL)-6 and IL-8 and the adhesion molecules CD41 and CD62 on platelets and CD11b and CD41 on leukocytes were determined. Because of considerable interindividual scatter, the pulmonary venous levels are normalized to percent of the respective right atrial value at each time point. RESULTS: At 1 min reperfusion pulmonary venous levels of IL-6 and IL-8 in routine circulation were +44+/-15% and +43+/-28% of the respective right atrial values. The respective values in bilateral circulation were -3+/-4% and -6+/-7% (P=0.02 and P=0.05 vs. respective right atrium). Similar increments were found after 10 and 20 min. Platelet-monocyte coaggregates were retained during pulmonary passage at 1 min reperfusion in routine circulation (-21+/-6%), but washed out in bilateral circulation (+5+/-8%, P=0. 007). At 20 min reperfusion, activated polymorphonuclear neutrophils (PMN) were retained in routine circulation (-16+/ 9%) but washed out in bilateral circulation (+19+/-29%, P=0.05; all data given as mean+/-SEM). CONCLUSIONS: Bilateral extracorporeal circulation without an artificial oxygenator significantly reduces the inflammatory responses during pulmonary passage after aortic declamping. PMID- 10856869 TI - Inflammatory response and myocardial injury following coronary artery bypass grafting with or without cardiopulmonary bypass. AB - OBJECTIVE: In coronary artery bypass grafting (CABG) without cardiopulmonary bypass (CPB) the inflammatory response is suggested to be minimized. Coronary anastomoses are performed during temporary coronary occlusion. Inflammatory response and myocardial ischaemia need to be studied in a randomized study comparing CABG in multivessel disease with versus without CPB. METHODS: Following randomization 30 consecutive patients received CABG either with (n=16) or without CPB (n=14). Primary study endpoints were parameters of the inflammatory response (interleukin (IL)-6, interleukin-10, ICAM-1, P-selectin) and of myocardial injury (myoglobin, creatine kinase-MB (CK-MB), troponin I) (intraoperatively, 4, 8, 16, 24 and 48 h after surgery). The secondary endpoint was clinical outcome. RESULTS: The incidence of major (death: CABG with CPB n=1, not significant (n.s.)) and minor adverse events (wound infection: with CPB n=2, without CPB n=1, n.s. ; atrial fibrillation: with CPB n=3, without CPB n=2, n.s.) was comparable between both groups. The release of IL-6 was comparable during 8 h of observation (n.s.). Immediately postoperatively IL-10 levels were higher in the operated group with CPB (211.7+/-181.9 ng/ml) than in operated patients without CPB (104.6+/-40.3 ng/ml, P=0.0017). Thereafter no differences were found between both groups. A similar pattern of release was observed in serial measures of ICAM-1 and P selectin, with no difference between both study groups (n.s.). Eight hours postoperatively the cumulative release of myoglobin was lower in operated patients without CPB (1829.7+/-1374. 5 microg/l) than in operated patients with CPB (4469.8+/-4525.7 microg/l, P=0.0152). Troponin I release was 300.7+/-470.5 microg/l (48 h postoperatively) in patients without CPB and 552.9+/-527.8 microg/l (P=0.0213). CK-MB mass release was 323.5+/-221.2 microg/l (24 h postoperatively) in operated patients without CPB and 1030. 4+/-1410.3 microg/l in operated patients with CPB (P=0.0003). CONCLUSIONS: This prospective randomized study suggests that in low-risk patients the impact of surgical access on inflammatory response may mimic the influence of long cross-clamp and perfusion times on inflammatory response. Our findings indicate that multiregional warm ischaemia, caused by snaring of the diseased coronary artery, causes considerably less myocardial injury than global cold ischaemia induced by cardioplegic cardiac arrest. PMID- 10856870 TI - Emergency reinstitution of cardiopulmonary bypass following cardiac surgery: outcome justifies the cost. AB - OBJECTIVE: Crash back on bypass (crash-BOB) is occasionally required in the resuscitation of patients developing life-threatening complications following cardiac surgery. This study aims to determine the incidence, aetiology and cost effectiveness of such intervention. METHODS: Retrospective review of all crash BOB patients over 5.5 years at one hospital. RESULTS: The incidence of crash-BOB was 0.8% and occurred at a mean of 7 h post-operatively (range 1 h-20 days). Pre operative Parsonnet scores were similar to the overall population of patients undergoing surgery in our institution (mean score 10; range 0-45). The original cardiac operations were coronary revascularization (39), valve surgery (12) and others (4). Indications for crash-BOB were cardiac arrest (23), bleeding (20), hypotension (7), ischaemia (1) and others (4). Of the 55 patients, 20 died on the operating table. Of the remaining 35, a further 12 died in hospital. Overall survival was therefore 42%. Where crash-BOB was for bleeding, 17 of 20 patients (85%) survived to leave theatre, of whom 11 patients (55%) left hospital alive. In the 35 non-bleeders, only 18 (51%) survived crash-BOB and 12 (34%) left hospital alive. Sixteen patients required a second period of aortic cross clamping of whom 13 (81%) survived to leave theatre, and 11 (69%) left hospital alive. Conversely, of nine patients in whom no specific diagnosis was found during crash-BOB, only two (22%) survived the procedure and none survived to hospital discharge. Multiple logistic regression identified pre-operative Parsonnet score (P=0.045) and the need for aortic cross-clamping to deal with an identified surgical problem (P=0.03) as significant predictors of hospital survival. Indication for crash-BOB (bleeder/non-bleeder) failed to reach significance (P=0.08). Age, sex, intra-aortic balloon pump use at the primary procedure, and time following the primary procedure to crash-BOB were not identified as predictors of hospital survival. Of the 23 hospital survivors, three patients suffered a stroke post-operatively and made a good functional recovery prior to discharge. Two patients developed sternal wound dehiscence requiring surgical rewiring. At follow-up (mean 3 years, range 1-6 years), 19 patients were in NYHA class I and four were in class II. Crash-BOB patients required an average of 8 extra intensive care days and 2 extra ward days. The total cost of these resources was pound164900 (including theatre time, cardiopulmonary bypass and intra-aortic balloon pump use). This was equivalent to pound7170 per life saved. CONCLUSIONS: Crash-BOB occurred in 0.8% of cases and was associated with a survival to discharge of 42%, and a justifiable cost of only pound7170 per life saved. Establishing an accurate diagnosis for the cause of clinical deterioration resulting in crash-BOB intervention was important, and the need for a further period of aortic cross-clamping did not preclude a favourable outcome. PMID- 10856871 TI - Clinical versus actual outcome in cardiac surgery: a post-mortem study. AB - BACKGROUND: Clinical attribution of the cause of death can be misleading, with the only true outcome measure being post-mortem analysis. Despite this there is very little published data on post-mortems following cardiac surgery. METHODS: Prospective consecutive post-mortem data were collected on 167 patients (84.4% of all in-hospital cardiac surgical deaths) in a single institution. Clinical diagnoses were compared with post-mortem findings. RESULTS: The mean age at death was 69.8 with 67.6% male. The proportion undergoing coronary artery bypass graft (CABG) alone was 52.1%, valve surgery 18.6%, valve+CABG 19.2% and other procedures 10.1%. The mean time to death was 7.9 days (range 0-87). The causes of death were cardiac 67.7%, gastrointestinal 9.6%, respiratory 8.4%, haemorrhage/technical failure 4.8%, stroke (cerebrovascular accident) 3.6%, multiorgan failure 3.0%, sepsis 1.8%, malignancy 0. 6% and trauma 0.6%. Post mortem revealed an unsuspected cause of death in 19 (11.4%). These were gastrointestinal (infarction nine, perforation two), cardiac three, adult respiratory distress syndrome two, technical two and pulmonary embolus one. In addition, an unsuspected lung cancer was found in 1 patient who died of cardiac causes. When cardiac deaths were compared with non-cardiac causes the Parsonnet score was higher 20.0 (+/-1.4) vs. 15.3 (+/-1.6), P=0. 07; and a greater proportion tended to have poor ejection fractions (34 vs. 15%), P=0.12. There was no significant difference between the groups in terms of age, sex, operation, hypertension, diabetes, creatinine and body mass. CONCLUSIONS: Post-mortem can determine unsuspected diagnoses in a significant proportion of cases. Pre operative risk factors do not correlate with eventual cause of death. Post-mortem still has an important role to play in cardiac surgery. PMID- 10856872 TI - Herniation of the heart due to traumatic rupture of the pericardium. PMID- 10856873 TI - Large thoracic duct cyst - a case report and review of the literature. AB - Large thoracic duct cysts are rare and standard lateral thoracotomy is usually used for resection. In the reported case the combination of an antero-lateral thoracotomy with a partial longitudinal median sternotomy (hemiclamshell approach) allowed an excellent visualization and dissection of a large thoracic duct cyst expanding in the anterior cervico-thoracic junction, and was associated with an uncomplicated recovery. PMID- 10856874 TI - Pseudoaneurysm in the mitral-aortic intervalvular fibrosa. A cause of mitral regurgitation. AB - Left ventricular outflow tract pseudoaneurysm is an uncommon but potentially catastrophic complication of aortic valve surgery, aortic valve endocarditis or chest trauma. We describe a case of a left ventricular outflow tract pseudoaneurysm 1 month after an aortic valve replacement that caused a systolic compression of mitral valve and a severe regurgitation. The diagnosis was confirmed using transoesophageal echocardiography, magnetic resonance image and intraoperative endoscopy. Surgical repair of the pseudoaneurysm corrected the mitral regurgitation. PMID- 10856875 TI - Undergoing cardiopulmonary bypass using enoxaparin only during a cardiac transplantation procedure. AB - The use of enoxaparin as a replacement drug to standard heparin, for anticoagulation during extracorporeal circulation, in patients with heparin induced thrombocytopenia, is still very limited. Enoxaparin significantly reduces thrombin formation and activity during cardiopulmonary bypass. The prolonged circulating rate, slow elimination rate and non-total reversion of enoxaparin by protamine can induce important postoperative bleeding. We are describing the first case of cardiac transplantation where enoxaparin was used as a replacement drug to standard heparin. PMID- 10856876 TI - Symptomatic calcific stenosis of a Toronto stentless porcine valve. AB - We describe the calcific structural failure of a Toronto stentless porcine valve (TSPV) which had been used to replace a calcified bicuspid aortic valve in a 46 year-old man. Against expectations, left ventricular hypertrophy persisted and the transvalvular pressure gradient rose to 125 mmHg by 6 years with the patient becoming symptomatic and requiring redo surgery. On removal the TSPV showed atypical calcification of the leaflet hinges and wall. To our knowledge this is the first case reported and it may have implications for long term durability and future surgery using this prosthesis. PMID- 10856877 TI - Multiple endocrine neoplasia type 1 (MEN1) gene mutations in a subset of patients with sporadic and familial primary hyperparathyroidism target the coding sequence but spare the promoter region. AB - Germ line mutations of the multiple endocrine neoplasia type 1 (MEN1) tumour suppressor gene cause MEN1, a rare familial tumour syndrome associated with parathyroid hyperplasia, adenoma and hyperparathyroidism (HP). Here we investigated the role of the MEN1 gene in isolated sporadic and familial HP. Using RT-PCR single-strand conformational polymorphism screening, somatic (but not germ line) mutations of the MEN1 coding sequence were identified in 6 of 31 (19.3%) adenomas from patients with sporadic primary HP, but none in patients (n=16) with secondary HP due to chronic renal failure. MEN1 mutations were accompanied by a loss of heterozygosity (LOH) for the MEN1 locus on chromosome 11q13 in the adenomas as detected by microsatellite analysis. No DNA sequence divergence within the 5' region of the MEN1 gene, containing the putative MEN1 promoter, was detectable in HP adenomas. Clinical characteristics were not different in HP patients with or without MEN1 mutation. Heterozygous MEN1 gene polymorphisms were identified in 9.6% and 25% of patients with primary and secondary HP respectively. In a large kindred with familial isolated familial HP, MEN1 germ line mutation 249 del4 and LOH was associated with the HP phenotype and a predisposition to non-endocrine malignancies. We suggest that the bi-allelic somatic loss of MEN1 wild-type gene expression is involved in the pathogenesis of a clinically yet undefined subset of sporadic primary HP adenomas. MEN1 genotyping may further help define the familial hyperparathyroidism-MEN1 disease complex, but it seems dispensable in sporadic primary HP. PMID- 10856878 TI - Characterization of parathyroid hormone/parathyroid hormone-related protein receptor and signaling in hypercalcemic Walker 256 tumor cells. AB - Parathyroid hormone (PTH)-related protein (PTHrP) is the main factor responsible for humoral hypercalcemia of malignancy. Both PTH and PTHrP bind to the common type I PTH/PTHrP receptor (PTHR), thereby activating phospholipase C and adenylate cyclase through various G proteins, in bone and renal cells. However, various normal and transformed cell types, including hypercalcemic Walker 256 (W256) tumor cells, do not produce cAMP after PTHrP stimulation. We characterized the PTHrP receptor and the signaling mechanism upon its activation in the latter cells. Scatchard analysis of PTHrP-binding data in W256 tumor cells revealed the presence of high affinity binding sites with an apparent K(d) of 17 nM, and a density of 90 000 sites/cell. In addition, W256 tumor cells immunostained with an anti-PTHR antibody, recognizing its extracellular domain. Furthermore, reverse transcription followed by PCR, using primers amplifying two different regions in the PTHR cDNA corresponding to the N- and C-terminal domains, yielded products from W256 tumor cell RNA which were identical to the corresponding products obtained from rat kidney RNA. Consistent with our previous findings on cAMP production, 1 microM PTHrP(1-34), in contrast to 10 microg/ml cholera toxin or 1 microM isoproterenol, failed to affect protein kinase A activity in W256 tumor cells. However, in these cells we found a functional PTHR coupling to G(alpha)(q/11), whose presence was demonstrated in these tumor cell membranes by Western blot analysis. Our findings indicate that W256 tumor cells express the PTHR, which seems to be coupled to G(alpha)(q/11). Taken together with previous data, these results support the hypothesis that a switch from the cAMP pathway to the phospholipase C-intracellular calcium pathway, associated with PTHR activation, occurs in malignant cells. PMID- 10856879 TI - Regulation of insulin-like growth factor-binding protein-3 ternary complex in feline diabetes mellitus. AB - The 140 kDa ternary complex of insulin-like growth factor-binding protein-3 (IGFBP-3), IGFs and an acid-labile subunit (ALS) has previously been shown to be decreased in diabetes mellitus in humans and rats. We have studied IGF-I levels and ternary complex formation in normal and diabetic cats. Total IGF-I concentrations, measured by RIA using des(1-3)-IGF-I as tracer were (+/-s.e.m.) 54+/-13 nmol/l in eight normal and 227+/-57 nmol/l in eight diabetic cats (P<0.01). The size-distribution of IGFBPs in the cat circulation was determined by incubation with (125)I-IGF-II and Superose 12 chromatography. In normal animals 26+/-2% of the (125)I-IGF-II were in a 140 kDa form compared with 48+/-5% in diabetic cats (P<0.01). When samples from normal and diabetic animals were co incubated 52+/-3% were at 140 kDa. A similar shift was seen when normal cat and normal human serum were co-incubated. A 2-fold increase in the 140 kDa form in diabetic cats was confirmed first by size-fractionating samples and then performing a ligand-binding assay with (125)I-IGF-I or -II and charcoal separation. SDS-PAGE and Western ligand blotting demonstrated a 45 kDa doublet (presumably IGFBP-3) and 30-35 kDa forms. There were no apparent differences between normal and diabetic profiles on SDS-PAGE, suggesting that a proportion of IGFBP-3 which circulates 'free' in normal cats forms a ternary complex in the diabetic circulation. We conclude that (i) in contrast to humans and rats, ALS is the limiting factor for ternary complex formation in normal cats, (ii) ALS concentrations increase in feline diabetes mellitus and, by promoting ternary complex formation, this leads to an increase in total IGF-I concentrations, and (iii) total IGF-I concentrations may not be reliable in the diagnosis of acromegaly in diabetic cats. PMID- 10856880 TI - Differential effects of IGF-binding proteins, IGFBP-3 and IGFBP-5, on IGF-I action and binding to cell membranes of immortalized human chondrocytes. AB - Insulin-like growth factor-I (IGF-I) is an important anabolic factor for cartilage tissue and its action is, in part, regulated by IGF-binding proteins (IGFBPs). The object of this study was to investigate the effects of IGFBPs on IGF-I action and on binding of IGF-I to cells using a reproducible immortalized human chondrocyte culture model. Treatment of the C-28/I2 cells with IGF-I or des(1-3)IGF-I in serum-free medium stimulated cell proliferation in a dose dependent manner. However, the effect of des(1-3)IGF-I was more potent, thereby suggesting that endogenously produced IGFBPs inhibited IGF action. The stimulatory effect of IGF-I was inhibited significantly by addition of IGFBP-3 but enhanced slightly by IGFBP-5. However, neither IGFBP-3 nor IGFBP-5 had an effect on basal cell growth. Binding of (125)I-labeled IGF-I to the cells was displaced by both IGFBP-3 and IGFBP-5, although higher concentrations of unlabeled IGFBP-5 were required to displace IGF-I to the same extent as IGFBP-3. Treatment of the cells with IGF-I increased the levels of IGFBP-5 protein measured by Western ligand blotting, and stimulated a corresponding increase in IGFBP-5 mRNA while increasing type II collagen mRNA. Our findings indicate that the balance between IGFBP-3 and IGFBP-5 influences IGF receptor binding and its action on chondrocyte proliferation, and may thereby modulate cartilage metabolism. PMID- 10856881 TI - The effect of opioid antagonism and environmental restriction on plasma oxytocin and vasopressin concentrations in parturient gilts. AB - Oxytocin plays an important role at parturition due to its involvement in uterine contractions, foetal expulsion and the onset of maternal behaviour. The role of the related neurohypophysial hormone, vasopressin, is less clear; however, there is some evidence that it is also involved in maternal behaviour and its role in osmotic regulation is well established. The aim of this study was to investigate the inhibitory effects of endogenous opioids on these hormones during the expulsive phase of parturition in the pig, and to examine how opioid restraint interacts with environmental restriction. The subjects of this study were 31 Large Whitex Landrace primiparous sows (gilts). An indwelling jugular catheter was implanted under general anaesthesia at 12 days before the expected parturition day (EPD). From 5 days before the EPD 15 of the gilts were individually housed in a restrictive parturition crate without straw and 16 were individually housed in a straw-bedded pen. Blood samples were taken with increasing frequency towards and during parturition through a catheter extension to reduce disturbance. At 7.5 min after the birth of the first piglet half of the gilts in each environment received a dose of the opioid receptor antagonist naloxone (1 mg/kg, i.v.) with the remaining gilts receiving saline as a control. Overall, there was no effect of environment on either circulating oxytocin or vasopressin. However, both oxytocin and vasopressin were inhibited by endogenous opioids during the expulsive phase. The inhibitory effects of opioids on these hormones did not appear to have any adverse effects on the progress of parturition as judged by cumulative piglet birth intervals. The regulation of the opioid inhibition of oxytocin and vasopressin during parturition is discussed in relation to other neurotransmitters and whether opioid inhibition of these neurohypophysial hormones is part of the 'normal' physiological response to parturition or whether it is stress-induced. PMID- 10856882 TI - Oxytocin receptor gene expression in rat uterus: regulation by ovarian steroids. AB - The present study was designed to investigate a possible role for ovarian steroids in the regulation of rat uterine oxytocin receptor (OTR) mRNA expression before labour. By using a competitive RT-PCR system, we have previously reported that parturition was associated with high levels of uterine OTR mRNA in all the animals examined. On the other hand, near term, some rats showed high OTR mRNA levels while others did not. We therefore examined the changes in OTR mRNA expression before and during prostaglandin F(2)(alpha) (PGF(2)(alpha))-induced parturition; a paradigm adopted to reduce the variation in the onset of parturition. Injection of PGF(2)(alpha) on day 18 of pregnancy significantly increased OTR mRNA expression in all the rats within 24 h of treatment, suggesting that the variation in OTR mRNA levels during spontaneous parturition may be due to the difference in the timing of the onset of parturition. The increase in OTR mRNA was significantly abolished by injection of the anti oestrogen compound, tamoxifen. The stimulatory action of oestrogen on OTR mRNA expression was then examined in the presence or absence of ovarian factors. Pregnant rats were ovariectomized (OVX) or sham-operated on day 18 of pregnancy and either oestrogen or vehicle was administered 6 h after the surgical operation. Oestrogen increased OTR mRNA significantly in OVX rats 18 h after administration compared with sham-operated animals. Moreover, ovariectomy alone on day 18 of pregnancy increased OTR mRNA expression to a level which reached statistical significance 24 h after the operation. In addition, oestrogen treatment increased OTR mRNA levels in OVX virgin rats in which progesterone tubes were implanted for 1 week and removed 6 h before oestrogen injection. The stimulatory effect of oestrogen was not observed in rats in which the progesterone tubes were implanted for 1 week and not removed. These results suggest that the decline of progesterone is necessary for the expression of the stimulatory effects of oestrogen on uterine OTR mRNA. PMID- 10856883 TI - Human umbilical vein endothelial cells express multiple prolactin isoforms. AB - Members of the prolactin (PRL) hormonal family have direct effects on endothelial cell proliferation, migration and tube formation. Moreover, isoforms of PRL may function as autocrine regulators of endothelial cells. Bovine brain capillary endothelial cells (BBCEC) express the PRL gene, while anti-PRL antibodies inhibit BBCEC proliferation. Here, we show the expression of the PRL gene into various PRL isoforms in endothelial cells from the human umbilical vein. Reverse transcription-polymerase chain reaction of total RNA from human umbilical vein endothelial cells (HUVEC) detected the full-length PRL mRNA as well as a 100 bp smaller PRL transcript similar to the one previously reported in BBCEC. HUVEC were positive to PRL immunocytochemistry. In addition, various PRL immunoreactive proteins were detected in HUVEC extracts and HUVEC conditioned media by metabolic labelling immunoprecipitation analysis. These PRL immunorelated proteins had apparent molecular masses of 60, 23, 21, 16 and 14 kDa. In contrast to previous findings in BBCEC, HUVEC conditioned media contained very little PRL bioactivity as determined by the selective bioassay of Nb2 cell proliferation. Moreover, some polyclonal or monoclonal antibodies directed against PRL stimulated HUVEC proliferation, in contrast to the inhibitory effect seen in BBCEC. The present findings extend the previous observations about the expression of PRL gene in endothelial cells from bovine brain capillaries to human cells of the umbilical vein, implicating that endothelium from different types of vessels and species share the expression of PRL gene but may differ in the putative autocrine role of the PRL isoforms expressed. PMID- 10856884 TI - Three novel paralogs of the rodent prolactin gene family. AB - The prolactin (PRL) family consists of a collection of genes expressed in the uterus, placenta and anterior pituitary. These cytokines/hormones participate in the control of maternal-fetal adaptations to pregnancy. In this report, we establish the presence of three new members of the PRL family. Novel expressed sequence tags (ESTs) with homology to PRL were isolated from embryonic and placental cDNA libraries. The cDNAs were sequenced and compared with those of other members of the PRL family. The three new cDNAs were assigned to the PRL family on the basis of sequence similarities and were referred to as PRL-like protein-J (PLP-J), PRL-like protein-K (PLP-K) and PRL-like protein-M (PLP-M). Both rat and mouse PLP-J cDNAs were identified. Rat PLP-J cDNA encodes for a predicted 211 amino acid protein containing a 29 amino acid signal peptide and two putative N-linked glycosylation sites, whereas the mouse PLP-J cDNA encodes for a 212 amino acid protein containing a 29 amino acid signal peptide with a single N-linked glycosylation site. Rat and mouse PLP-J proteins share approximately 79% and 70% nucleotide and amino acid sequence identity, respectively. A full-length rat PLP-K cDNA and a partial tentative mouse PLP-K cDNA were identified. The rat PLP-K cDNA encodes for a predicted 228 amino acid protein containing a 31 amino acid signal peptide and one putative N-linked glycosylation site; the mouse PLP-M cDNA encodes for a predicted 228 amino acid protein containing a 28 amino acid signal peptide and one putative N-linked glycosylation site. Genes for PLP-J, PLP-K and PLP-M are situated at the Prl family locus on mouse chromosome 13. PLP-J was exclusively expressed in decidual tissue from both the mouse and rat. PLP-K was expressed in trophoblast cells of the chorioallantoic placenta and showed an apparent species difference. In the mouse, virtually all trophoblast lineages expressed PLP-K, whereas in the rat, PLP-K expression was restricted to the labyrinthine trophoblast cells. Mouse PLP M expression was restricted to the junctional zone of the chorioallantoic placenta. In summary, we have identified three new members of the rodent PRL gene family that are expressed in uterine and placental structures. Future experimentation is needed to determine the specific roles of each of these ligands in the biology of pregnancy. PMID- 10856885 TI - Long-term testosterone treatment prevents gonadal and adrenal tumorigenesis of mice transgenic for the mouse inhibin-alpha subunit promoter/simian virus 40 T antigen fusion gene. AB - We have developed a transgenic (TG) mouse model for tumorigenesis of gonadal somatic cells using a 6 kb fragment of the mouse inhibin-alpha subunit promoter (Inh-alpha) fused with the simian virus 40 T-antigen (Tag) coding sequence. Gonadal tumors, of Leydig or granulosa cell origin, develop in the TG mice with 100% penetrance by the age of 5-8 months. Conspicuously, if the mice are gonadectomized, they develop adrenal tumors. Gonadal and adrenal tumorigenesis in these mice seem to be gonadotropin dependent. On the other hand, testosterone stimulates the proliferation of a cell line (C alpha 1) established from one of the adrenal tumors. The purpose of the present study was therefore to investigate further whether testosterone affects the growth of these gonadal and adrenal tumors in vivo. Two experimental models were used: (1) Tag TG/hypogonadotropic (hpg) double mutant mice and (2) castrated Tag TG mice. Both were treated between 1-2 and 7-8 months of age with Silastic rods (length 2 cm) containing testosterone. None of the control or testosterone-treated Tag/hpg mice developed gonadal or adrenal tumors. The castrated Tag TG mice displayed, upon microscopical examination, early stages of adrenal tumors, whereas those receiving testosterone did not show such changes. Testosterone increased the weights of gonads in the Tag/hpg mice, and those of uteri and seminal vesicles in both groups. In contrast, the adrenal weights were significantly reduced in both groups by testosterone treatment. Gonadal histology of the testosterone-treated mice showed hyperplasia of testicular Leydig cells and ovarian stroma. Spermatogenesis was induced by testosterone in the Tag/hpg mice. Adrenal histology of the testosterone-treated animals demonstrated the disappearance of the X-zone. Serum levels of FSH in testosterone-treated Tag/hpg mice were significantly increased, while those of serum LH were decreased. In conclusion, the present result indicate that the suppression of gonadotropins by testosterone implants in castrated Inh-alpha/Tag TG mice prevents the tumorigenesis of their adrenals. In intact Tag/hpg mice, testosterone implants were not able to induce gonadal or adrenal tumorigenesis. Although testosterone treatment was able to induce interstitial cell hyperplasia in gonads of the Inh-alpha/Tag mice, direct gonadotropin action is responsible for gonadal and adrenal tumorigenesis. PMID- 10856886 TI - Expression of proliferating cell nuclear antigen in luminal epithelium during the growth and regression of rat uterus. AB - Proliferating cell nuclear antigen (PCNA), a processivity factor of DNA synthesis, has often been used as a marker that reveals proliferating cells. However, it also plays a role other than in DNA replication. The aim of this study was to examine the relationship between the expression of PCNA and cell proliferation, and also its relation to cell death in the uterine epithelium under various hormonal conditions. Rats with regular estrous cycles were killed at various stages of the cycle, and their uteri were removed for the detection of PCNA and apoptosis by immunohistochemical and terminal deoxynucleotidyl transferase-mediated nick end-label staining respectively. There was an inverse relationship between the expression of PCNA and apoptosis in the uterine epithelium during the estrous cycle. From diestrus to proestrus, the expression of PCNA increased, and few apoptotic cells were detected in the luminal epithelium. However, at estrus, apoptosis occurred markedly, and the expression of PCNA disappeared. To study further the effects of estrogen on PCNA expression and cell growth in the uterus, rats were ovariectomized and then implanted s.c. with estrogen capsules 2 weeks later. In ovariectomized rats, only a few PCNA positive cells were observed in the uterine epithelium. After estrogen treatment, PCNA was expressed strongly in the luminal and glandular epithelia. In these rats, the removal of estrogen capsules resulted in apoptotic death and surprisingly strong PCNA expression in the cells of luminal epithelium. Our results demonstrate that PCNA is expressed not only in the estrogen-stimulated uterine growth, but also in the processes of regression induced by the withdrawal of estrogen. Although the expression of PCNA has been reported to represent cell proliferation, our results implicate functions other than cell replication for PCNA in the uterus. PMID- 10856887 TI - Expression of 3 beta-hydroxysteroid dehydrogenase/isomerase in the female rat pituitary. AB - 3 beta-Hydroxysteroid dehydrogenase/isomerase (3 beta-HSD) catalyses an essential step in the biosynthesis of steroid hormones and is widely distributed in peripheral steroid target organs. The present report describes for first time the expression of this enzyme in the pituitary of female rats. Immunohistochemistry at the light microscopic level was performed on pro-oestrous and ovariectomized rat pituitaries. Immunoreactive cells were scattered and randomly distributed throughout the anterior lobe, whereas cells located in the posterior lobe and pars intermedia were immunonegative. Differences were observed in cell morphology and in the number of 3 beta-HSD-immunopositive cells between ovariectomized and pro-oestrous female rat pituitaries, suggesting that steroidogenic activity is affected by ovarian endocrine function. Apart from adenohypophyseal immunoreactive cells, 3 beta-HSD immunopositivity was also noted in endothelial cells of almost all pituitary capillaries located in the anterior and posterior lobes. PMID- 10856888 TI - Accelerated telomere shortening and senescence in human pancreatic islet cells stimulated to divide in vitro. AB - Widespread application of beta-cell replacement strategies for diabetes is dependent upon the availability of an unlimited supply of cells exhibiting appropriate glucose-responsive insulin secretion. Therefore, a great deal of effort has been focused on understanding the factors that control beta-cell growth. Previously, we found that human beta-cell-enriched islet cultures can be stimulated to proliferate, but expansion was limited by growth arrest after 10-15 cell divisions. Here, we have investigated the mechanism behind the growth arrest. Our studies, including analyses of the expression of senescence associated beta-galactosidase, p16(INK4a) levels, and telomere lengths, indicate that cellular senescence is responsible for limiting the number of cell divisions that human beta-cells can undergo. The senescent phenotype was not prevented by retroviral transduction of the hTERT gene, although telomerase activity was induced. These results have implications for the use of primary human islet cells in cell transplantation therapies for diabetes. PMID- 10856889 TI - Glucose-induced insulin secretion from islets of fasted rats: modulation by alternate fuel and neurohumoral agonists. AB - Islets from fed and 24-h-fasted rats were studied immediately after collagenase isolation. (1) After a 24-h fast, the insulin secretory responses to 8 mM glucose measured during perifusion were reduced by more than 90% from islets of fasted donors. (2) Increasing glucose to 11 or 27.5 mM resulted in enhanced insulin secretion from islets of fasted animals. (3) Fasting did not reduce islet insulin content. (4) Responses to 8 or 27.5 mM glucose were not affected if fatty acid free albumin was used during the perifusion. (5) Inclusion of alpha ketoisocaproate (5 mM), monomethyl succinate (10 mM) or carbachol (10 microM) significantly amplified insulin release from fasted islets in the simultaneous presence of 8 mM glucose. (6) Phospholipase C activation by glucose, carbachol or their combination was not adversely affected by fasting. (7) The response to the protein kinase C activator, phorbol 12-myristate 13-acetate (500 nM), was reduced by about 60% after fasting. (8) Extending the fast to 48 h resulted in a severe decline in response to 11 mM glucose; however, the further addition of 10 microM carbachol still enhanced release from these islets. The results confirm that caloric restriction impairs islet sensitivity to glucose stimulation and that protein kinase C may be involved in the reduction of glucose-induced insulin release from these islets. The activation of phospholipase C by cholinergic stimulation may contribute to the maintenance of insulin secretion from calorically restricted animals. These results also demonstrate that free fatty acids are not essential for glucose to evoke secretion from isolated islets of fasted donors. PMID- 10856890 TI - Interleukin-6 increases insulin secretion and preproinsulin mRNA expression via Ca2+-dependent mechanism. AB - Interleukin (IL)-6, one of the cytokines released from inflammatory cells, stimulates insulin secretion in a physiological concentration (1-100 pg/ml), but the exact mechanism is still unknown. The present studies were undertaken to investigate the mechanism of IL-6-induced stimulation of insulin secretion in HIT T 15 cells. The effects of the addition of nifedipine on the IL-6 (100 pg/ml) induced stimulation of insulin secretion were investigated. We also examined the possibility that IL-6 (1-100 pg/ml) may stimulate insulin messenger ribonucleic acid (mRNA) expression, using the reverse transcription-polymerase chain reaction method. The addition of 100 and 1000 nM nifedipine significantly attenuated the stimulatory effects of 100 pg/ml IL-6 on insulin secretion. The addition of 1-100 pg/ml IL-6 dose-dependently increased preproinsulin mRNA expression relative to beta-actin mRNA. IL-6 increased insulin gene promoter activity of fragments A ( 2188 to +337 bp) and B (-1782 to +270 bp) but not fragments C (-1275 to +270 bp), D (-1138 to +270 bp), E (-880 to +236 bp) or F (-356 to +252 bp). The addition of 10 nM nifedipine completely abolished the stimulatory effect of 10-100 pg/ml IL-6 on relative preproinsulin mRNA expression. These data raised the possibility that IL-6 increased preproinsulin mRNA expression via the stimulation of Ca(2+) influx which enhances insulin gene expression. PMID- 10856891 TI - Adrenaline, insulin and glucagon do not have acute effects on plasma leptin levels in sheep: development and characterisation of an ovine leptin ELISA. AB - Leptin, a recently discovered hormone secreted mainly from adipose tissue, was first described as a regulator of adiposity, food intake and energy metabolism. It is now apparent that leptin physiology is much more complex and is likely to play an important role in many other systems including reproduction, haematopoiesis and immunity. Leptin levels have been shown to be well correlated with body fat in both humans and rodents, with administration of exogenous leptin to rats and mice resulting in loss of body fat. Leptin is, therefore, likely to be an important humoral signal to the central nervous system on body composition and regulation of food consumption. Due to the limited cross-reactivity of leptin from other species in the current assays for leptin, physiological research on leptin has, to a large extent, been restricted to rodents and humans. The aim of this study was to develop a leptin immunoassay suitable for use with sheep, enabling the investigation of the basic physiology of leptin in an animal larger than rats or mice, thus allowing repeated blood sampling. Using this assay we investigated the short-term effects of insulin, adrenaline and glucagon (all modulators of blood glucose) on plasma leptin levels. Antiserum to bovine recombinant leptin (brLeptin) raised in chickens was used to develop a competitive ELISA. Using brLeptin as standard, the assay has a sensitivity of 0. 5 ng/ml with inter- and intra-assay variation of 15% and 7% respectively. The cross-reactivity of human recombinant leptin was 36.5%, while mouse leptin showed no cross-reactivity. Plasma samples from ewes, male castrate animals and rams (n=4-5) diluted in parallel to the standard with mean leptin concentrations of 6.0+/-2. 9, 3.3+/-0.4 and 3.1+/-1.3 ng/ml respectively. Leptin levels in rams were significantly lower than in ewes. The non-significant difference in leptin levels between rams and male castrate animals suggests that testosterone may not be responsible for the lower levels of leptin. Four groups of 3-4 ewes were given intravenous insulin (1 iu/kg), adrenaline (65 microg/kg), glucagon (24 iu/kg) or saline. Blood samples were taken at 1, 3, 5, 10, 20, 30, 60, 90 and 120 min after injection. As expected, glucose levels declined within 10 min of the insulin injection and rose after 3 min following both adrenaline and glucagon injections. Leptin levels, however, remained relatively unchanged for the 2 h following the treatments. Finally, a bolus intravenous dose of glucose (240 mg/kg) was given and sequential blood samples taken. Despite plasma glucose levels rising to over 200 mg/dl, leptin levels did not significantly change over the three hours following treatment. These data indicate that plasma leptin levels in sheep, in contrast to rodents, are not responsive to short-term changes in blood glucose or insulin, as has been shown in humans. PMID- 10856892 TI - Thyrotropin-releasing hormone time-dependently influences thyrotropin microheterogeneity--an in vivo study in euthyroidism. AB - Thyrotropin (TSH) is secreted not as one distinct hormone, but rather as a group of isohormones which differ in their oligosaccharide composition. Although the mechanisms regulating TSH glycosylation are not fully understood, there is strong evidence that TRH plays an important role. The aim of our study was to determine the dynamic influence of TRH on TSH microheterogeneity. Sera were obtained from euthyroid volunteers (n=20) before and 30, 60, 120, 180 and 240 min after intravenous, nasal and oral administration of TRH in three independent runs (randomized order, at a time-interval of 3 weeks between each run). TSH was immuno-concentrated and analysed by isoelectric focusing (IEF) and lentil lectin affinity chromatography. TSH immunoreactivity was measured by an automated second generation TSH immunoassay. Overall, serum TSH concentrations reached maximal values 30 min after intravenous, 60 min after nasal and 180 min after oral TRH stimulation. IEF analysis revealed 63.3+/-3.3% of pituitary standard TSH (IRP 80/558) in the neutral pH range (8>pH>6). In contrast, 30 min after TRH stimulation 80.8+/-3.7% (P<0.001) and 60 min after TRH stimulation 44.9+/-2.2% (P<0.001) of the TSH of euthyroid probands were found in this pH range, whereas 180 min after TRH stimulation 58.4+/-2.3% (P<0.001) were detected in the acidic pH range (pH<6). This shift of TSH composition in euthyroidism after TRH stimulation was confirmed by lentil lectin analysis of TSH: core-fucose content of euthyroid TSH was 73.4+/-3.8% 30 min and 22.9+/-3.2% 120 min after TRH stimulation in contrast to basal (53.3+/-1.8%; P<0.001) and pituitary standard (IRP 80/558) TSH (63.0+/-0.9%; P<0.001). In conclusion, in euthyroidism, TRH stimulation time-dependently changes the distribution pattern of the TSH isoforms from an alkaline and neutral to a more acidic one. This corresponds to the secretion of isohormones with altered bioactivity which could influence the fine tuning of thyroid function. PMID- 10856893 TI - Glucocorticoid receptors in human preadipocytes: regional and gender differences. AB - Glucocorticoid excess causes visceral obesity and its accompanying insulin resistance, dyslipidemia and hypertension. Glucocorticoids enhance preadipocyte (PA) differentiation and increase their aromatase activity (oestrogen production) and there is regional variability in these PA processes. Therefore, we studied human PAs for the presence of, and any regional or gender differences in, glucocorticoid receptors (GRs). Confluent subcultured human subcutaneous (Sc) and visceral (Vis) PAs from both genders contained GRs as assessed by GR gene expression and specific glucocorticoid (dexamethasone) binding. The dissociation constant was similar to that of other human cells and there was no difference between Sc and Vis sites or between males and females. There was significantly less GR mRNA in Vis PAs compared with Sc PAs in females (P=0.008) but not in males. There was less glucocorticoid binding in Vis compared with Sc PAs in females, measured by maximal binding capacity (P=0.035) or single saturating dose glucocorticoid binding (Bssd) (P=0.019). There was no regional difference in specific glucocorticoid binding in males. There was a gender difference with fewer GRs in Vis PAs in females compared with males measured by Bssd (P=0.006). In summary, GRs are present in human PAs. There is a lower GR density in Vis compared with Sc PAs in females, and females have fewer GRs in Vis PAs compared with males. These differences are likely to affect regional aromatase activity and to contribute to the smaller visceral fat mass in females compared with males. PMID- 10856894 TI - Ageing, testicular tumours and the pituitary-testis axis in dogs. AB - Dogs of different ages without testicular diseases were evaluated to study possible age-related changes in hormone concentrations in serum. Dogs with testicular tumours were also investigated to study the relation between tumour type and hormone concentrations; in this study, dogs with Sertoli cell tumours, Leydig cell tumours and seminomas were included. We measured testosterone, oestradiol, LH, FSH and inhibin-like immunoreactivity concentrations in peripheral venous and testicular venous blood of these animals. In normal dogs there appeared to be no age-related changes in the concentrations of the investigated hormones, except for a significant age-related decrease in oestradiol concentrations in testicular venous blood (P<0.02). Dogs with a Sertoli cell tumour had greater oestradiol concentrations and inhibin-like immunoreactivity in both peripheral and testicular venous blood than did dogs without a neoplasm (P<0. 05). Testosterone concentrations were reduced in dogs with Sertoli cell tumours, as were FSH and LH. Feminisation occurred in eight of 13 dogs with a Sertoli cell tumour and in two of 14 dogs with a Leydig cell tumour; it was accompanied by a significantly greater oestradiol concentration than in normal dogs and in dogs with Sertoli cell tumours without signs of feminisation. Dogs with a Leydig cell tumour had greater concentrations of oestradiol and inhibin-like immunoreactivity in both peripheral venous and testicular venous blood than did dogs without a neoplasm (P<0.05). The testosterone concentration in testicular venous blood of these dogs was lower than that in dogs with normal testes. The concentration of LH in peripheral venous blood was also reduced (P<0. 05). Hormone concentrations in dogs with a seminoma were not different from those in normal dogs. It was concluded that seminomas are not endocrinologically active. In contrast, both Sertoli cell tumours and Leydig cell tumours can cause increased oestrogen production leading to signs of feminisation. These tumours also have considerable amounts of inhibin like immunoreactivity, but only in Sertoli cell tumours does this result in a reduction in FSH concentrations, suggesting that Sertoli cell tumours secrete dimeric inhibin, whereas Leydig cell tumours presumably produce loose alpha subunits that cross-react in the inhibin assay but are not biologically active. PMID- 10856895 TI - Diazepam-binding inhibitor/acyl-CoA-binding protein mRNA and peripheral benzodiazepine receptor mRNA in endocrine and immune tissues after prenatal diazepam exposure of male and female rats. AB - Peripheral benzodiazepine (BDZ) receptor (PBR) and diazepam-binding inhibitor/acyl-CoA-binding protein (DBI/ACBP) characterized as a ligand at central BDZ receptors, at PBR with involvement in the regulation of steroidogenesis, and as an intracellular acyl-CoA transporter, are both known to interact with BDZ in adult systems. We investigated their expression after prenatal exposure to BDZ. Diazepam (1.25 mg/kg per day s.c.) was administered to time-pregnant Long Evans rats from gestational day (GD) 14 to 20. Expression of mRNAs encoding for PBR and for DBI/ACBP was studied in the same animals with (33)P-labeled 60 mer oligonucleotides (oligos) by in situ hybridization at GD20, and with (32)P-labeled oligos by Northern blot in steroidogenic and immune organs at postnatal day (PN) 14 and in adult offspring. Prenatal diazepam increased DBI/ACBP mRNA expression in male fetal adrenal and in fetal and PN14 testis. Thymus exhibited increased DBI/ACBP mRNA in male fetuses and in adult female offspring, and reduced organ weight at PN14 in both sexes. In female spleen, an increase in DBI/ACBP mRNA and a decrease in PBR mRNA was seen at PN14. Apart from the finding in spleen, no drug-induced changes in PBR mRNA were observed. The effects of prenatal diazepam were superimposed on treatment-independent sex differences in DBI/ACBP mRNA and PBR mRNA expression. Our data indicate that expression of DBI/ACBP mRNA in steroidogenic and immune organs can be affected by exposure to BDZ during ontogeny, while PBR mRNA expression appears to be less sensitive. They further reveal marked sex differences in the developmental patterns of the two proteins during pre- and postpubertal ontogeny. PMID- 10856896 TI - Tri-iodothyronine induces proliferation in cultured bovine thyroid cells: evidence for the involvement of epidermal growth factor-associated tyrosine kinase activity. AB - The effects of the tri-iodothyronine (T(3)) secreted by thyroid cells on the growth of the thyrocyte are poorly known. In this study we analyzed the effects of T(3) on the proliferation of bovine thyroid follicles in primary culture previously depleted of endogenous T(3). Cellular deoxiribonucleic acid (DNA) synthesis, determined by [(3)H]thymidine incorporation, was stimulated by T(3) (0.1-5.0 nM) for 24 h in a concentration-dependent fashion with a maximal effect at 1.0 nM T(3) (P<0.01). This T(3) action was time-dependent when assayed from 12 to 72 h. The induction of mitogenic activity was corroborated by the increase in proliferating cell nuclear antigen (PCNA) measured by Western blot analysis. PCNA increased after treatment with T(3) (0.1-5.0 nM) in a concentration-dependent manner. Since T(3) modifies the activity of growth factors whose actions are mainly mediated by tyrosine kinase (TK) activation in diverse cellular types, we assayed the effects of genistein, a general TK inhibitor, and tyrphostin A25, a specific epidermal growth factor (EGF)-receptor (EGFR)-dependent TK activity inhibitor, on the proliferative effects of T(3). The T(3)-induced [(3)H]thymidine incorporation was inhibited by both agents in a concentration-dependent manner. A significant increase in the total TK activity measured in cellular protein extracts was induced by 0.5 and 1.0 nM T(3) (P<0.001). Tyrosine phosphorylation of the EGFR was also stimulated by T(3) (P<0.001) with no change in the EGFR expression as determined by Western blot analysis. Both, the T(3)-stimulated [(3)H]thymidine incorporation and the TK activity were inhibited by a anti-mouse EGF antibody. These results lead us to propose that T(3) could operate as a proliferative agent in bovine thyroid cells through a mechanism involving an autocrine/paracrine EGF/EGFR-dependent regulation. PMID- 10856897 TI - Direct modulation of basal and angiotensin II-stimulated aldosterone secretion by hydrogen ions. AB - Disturbances in acid-base balance in vivo are associated with changes in plasma aldosterone concentration, and in vitro changes in extracellular pH (pH(o)) influence the secretion of aldosterone by adrenocortical tissue or glomerulosa cells. There is considerable disparity, however, as to the direction of the effect. Furthermore, the mechanisms by which pH(o) independently affects aldosterone secretion or interacts with other secretagogues are not defined. Thus, bovine glomerulosa cells maintained in primary monolayer culture were used to examine the direct effects of pH(o) on cytosolic free calcium concentration ([Ca(2+)](i))( )and aldosterone secretion under basal and angiotensin II (AngII) stimulated conditions. pH(o) was varied from 7.0 to 7.8 (corresponding inversely to changes in extracellular H(+) concentration from 16 nM to 100 nM). Whereas an elevation of pH(o) from 7.4 to 7.8 had no consistent effect, reductions of pH(o) from 7.4 to 7.2 or 7.0 caused proportionate increases in aldosterone secretion that were accompanied by increases in transmembrane Ca(2+) fluxes and [Ca(2+)](i). These effects were abolished by removal of extracellular Ca(2+). A decrease in pH(o) from 7.4 to 7.0 also enhanced AngII-stimulated aldosterone secretion. This effect was more pronounced at low concentrations of AngII and was manifested as an increase in the magnitude of the secretory response with no effect on potency. In contrast to its effect on AngII-stimulated aldosterone secretion, a reduction of pH(o) from 7.4 to 7.0 inhibited the Ca(2+) signal elicited by low concentrations (Be) fast neutrons and 4 MeV photons. AB - Fast neutrons have been used in the clinical radiation therapy of tumors largely because of experimental evidence that their cytotoxic effects are much less dependent on oxygen levels than those of low-LET photons. The potential therapeutic advantage of fast neutrons based on hypoxia alone can be calculated as the "hypoxic gain factor", which is the ratio of the OERs for the fast-neutron compared to the photon beams. The hypoxic gain factor that is generally anticipated based on studies with established mammalian cell lines is about 1.6. However, surprisingly few studies have examined the influence of hypoxia on the fast-neutron radiosensitivity of human tumor cells of different histological types. For this reason, we have determined the OERs of five human tumor cell lines exposed to 62.5 MeV (p-->Be) cyclotron-generated fast neutrons or 4 MeV photons from a clinical linear accelerator. The OERs for four chemotherapy-naive cell lines, HT29/5, Hep2, HeLa and RT112, were invariably greater for photons than for neutrons, but all of these values were lower than expected on the basis of the previous literature. Despite their low OERs, these cell lines showed hypoxic gain factors that were within the range of 1.31-1.63, indicating that such effects cannot entirely explain the disappointing clinical results obtained with fast neutrons. In contrast, comparison of the surviving fractions at clinically relevant doses (1.6 Gy of neutrons and 2.0 Gy of photons) for these four tumor cell lines suggested that little benefit should result from neutron treatment. Only the cisplatin-resistant OAW42-CP line showed a significant hypoxic gain factor by this method of analysis. We conclude that, at the dose fractions used in clinical radiation therapy, there may not be a radiobiological precedent for higher local control rates after fast-neutron irradiation of hypoxic tumor cells. PMID- 10856967 TI - RAS-Mediated radiation resistance is not linked to MAP kinase activation in two bladder carcinoma cell lines. AB - The expression of activated RAS oncogenes has been shown to increase radioresistance in a number of cell lines. The pathways by which RAS leads to radioresistance, however, are unknown. RAS activates several signal transduction pathways, with the RAF-MAP2K-MAP kinase pathway perhaps the best studied. MAP kinase has also been shown to be activated by radiation through this pathway. Given the important role of MAP kinase in multiple signaling events, we asked if radioresistance induced by RAS was mediated through the activation of MAPK. Cells of two human bladder carcinoma cell lines were used, one with a mutated oncogenic HRAS (T24) and other with a wild-type HRAS (RT4). The surviving fraction after exposure to 2 Gy of radiation (SF2) for the T24 cell lines was found to be 0.62, whereas that for RT4 cells was 0.40. Treatment with the farnesyl transferase inhibitor (FTI) L744,832, which inhibits RAS processing and activity, decreased the SF2 of T24 cells to 0.29, whereas the SF2 of RT4 cells remained unchanged after FTI treatment, thus demonstrating the importance of RAS activation to the radiosensitivity of cells with mutated RAS. MAP kinase activation was found to be constitutive and dependent on RAS in T24 cells, while it was inducible by radiation and was independent of RAS in RT4 cells. Treatment of both cell lines with the MAP2K inhibitor PD98059 inhibited MAPK activation; however, inhibiting MAPK activation had no effect on radiation survival of T24 or RT4 cells. These data indicate that MAPK activation does not contribute to RAS-induced radioresistance in this system. PMID- 10856968 TI - Activation of the nitric oxide synthase 2 pathway in the response of bone marrow stromal cells to high doses of ionizing radiation. AB - Reverse transcription-polymerase chain reaction and immunofluorescence analysis of D2XRII murine bone marrow stromal cells showed that gamma irradiation with doses of 2-50 Gy from (137)Cs stimulated expression of nitric oxide synthase 2 (Nos2, also known as iNos). The activation of Nos2 was accompanied by an increase in the fluorescence of 4,5-diaminofluorescein diacetate, a nitric oxide trap, and accumulation of 3-nitrotyrosine within cellular proteins in a dose-dependent manner. These effects were inhibited by actinomycin D and by N-[3 (aminomethyl)benzyl]acetamidine dihydrochloride, a specific inhibitor of Nos2. The induction of Nos2 expression and Nos2-dependent release of nitric oxide in D2XRII cells was observed within 24 h after irradiation and was similar in magnitude to that observed in cultures incubated with Il1b and Tnf. We conducted (1) confocal fluorescence imaging of 3-nitrotyrosine in bone marrow cells of irradiated C57BL/6J mice and (2) 3-nitrotyrosine fluorescence imaging of FDC P1JL26 hematopoietic cells that were cocultured with previously irradiated D2XRII bone marrow stromal cells. Exposure to ionizing radiation increased the production of 3-nitrotyrosine in irradiated bone marrow cells in vivo and in nonirradiated FDC-P1JL26 cells cocultured with irradiated D2XRII cells for 1 or 4 h. We suggest that nitrative/oxidative stress to the transplanted multilineage hematopoietic cells due to exposure to nitric oxide released by host bone marrow stromal cells may contribute to the genotoxic events associated with malignant alterations in bone marrow tissue of transplant recipients who are prepared for engraftment by total-body irradiation. PMID- 10856969 TI - The kinetics of postirradiation chromatin restitution as revealed by chromosome aberrations detected by premature chromosome condensation and fluorescence in situ hybridization. AB - In a study of X-ray-induced chromosome aberrations in human G(0) lymphocytes irradiated with 4 Gy using premature chromosome condensation (PCC) and fluorescence in situ hybridization (FISH), the time-dependent pattern of chromosome fragments and interchromosomal exchanges involving chromosome 4 was recorded after postirradiation incubation times varying from 0.5 to 46.5 h. Unattached acentric fragments and incomplete interchromosomal exchanges have high initial yields, followed by an exponential decrease, while complete interchromosomal exchanges have almost zero initial yield with a subsequent increase in their number. Plateau values of all yields are reached after about 25 h. This temporal variation of aberration yields can consistently be explained by the competition of disruptive PCC stress with the progress of postirradiation structural restitution at the sites of radiation-induced chromatin instabilities. Details of the temporal pattern of incomplete exchanges reflect the different kinetics of the alpha and beta components of the yield of aberrations. The observed large difference between late-PCC and metaphase yields of unattached acentric fragments and the almost perfect conversion from incomplete prematurely condensed chromosomes into complete metaphase exchanges are explained by a difference in the magnitude of chromosome condensation stress between PCC and mitotic conditions. Chromatin sites prone to fragmentation and incompleteness under conditions of PCC can therefore persist as genetic instabilities hidden during mitosis. PMID- 10856970 TI - Radiomodfication of xanthine oxidoreductase system in the liver of mice by phenylmethylsulfonyl fluoride and dithiothreitol. AB - The widely distributed xanthine oxidoreductase (XOR) system has been shown to be modulated upon exposure of animals to ionizing radiation through the conversion of xanthine dehydrogenase (XDH) into xanthine oxidase (XO). In the present work, radiomodification of the XOR system by phenylmethylsulfonyl fluoride (PMSF) and dithiothreitol (DTT) was examined using female Swiss albino mice which were irradiated with gamma rays at a dose rate 0.023 Gy s(-1). PMSF, a serine protease inhibitor, and DTT, the sulfhydryl reagent, were administered intraperitoneally prior to irradiation. The specific activities of XDH and XO as well as the XDH/XO ratio and the total activity (XDH+XO) were determined in the liver of the mice. The inhibition of XO activity, restoration of XDH activity, and increase in the XDH/XO ratio upon administration of PMSF were suggestive of irreversible conversion of XDH into XO mediated through serine proteases. The biochemical events required for the conversion were probably initiated during the early phase of irradiation, as the treatment with PMSF immediately after irradiation did not have a modulatory effect. Interestingly, DTT was not effective in modulating radiation-induced changes in the XOR system or oxidative damage in the liver of mice. The DTT treatment resulted in inhibition of the release of lactate dehydrogenase. However, the protection appears to be unrelated to the formation of TBARS. On the other hand, the presence of PMSF during irradiation inhibited radiation-induced oxidative damage and radiation-induced increases in the specific activity of lactate dehydrogenase. These findings suggest that a major effect of ionizing radiation is irreversible conversion of xanthine to xanthine oxidase. PMID- 10856971 TI - Uptake and washout of borocaptate sodium and borono-phenylalanine in cultured melanoma cells: a multi-nuclear NMR study. AB - The cellular uptake and washout of the two principal boron neutron capture therapy (BNCT) agents, borocaptate sodium (BSH) and borono-phenylalanine (BPA), were monitored on-line, noninvasively, using nuclear magnetic resonance (NMR) spectroscopy. The uptake and washout of inorganic borate (B(i)) was also followed for comparison. M2R mouse melanoma cells grown on polystyrene microspheres were perfused inside the NMR sample tube. (11)B NMR was used to detect the presence of B(i), BSH and BPA, and (19)F NMR was applied to detect fluorinated BPA ((19)F BPA). The results revealed chemical modifications of BSH due to spontaneous formation of the borocaptate dimer, BSSB, in the culture medium. BPA readily formed a complex with glucose contained in the culture medium but was also converted in the cells to a yet unidentified compound in a reaction that probably involves the hydrolysis of BPA and the release of B(i). The cellular accumulation ratio for BPA was significantly higher than 1 and was also significantly higher than that for BSH. On the other hand, the cellular retention time observed for BSH was much longer than for BPA, indicating a strong trapping of BSH in cells. PMID- 10856972 TI - Absence of linkage between radiosensitivity and the predisposing atp7b gene mutation for heritable hepatitis in the LEC rat. AB - The LEC rat is known to be a mutant strain that spontaneously develops heritable hepatitis due to copper accumulation, caused by mutation of the copper transporting ATPase gene (Atp7b). Immunodeficiency and radiosensitivity have also been observed. Hayashi et al. extensively examined the radiosensitivity of the LEC rat and concluded that its hypersensitivity is controlled by a single autosomal gene. Furthermore, they suggested the possibility that it correlates to copper accumulation due to the Atp7b gene mutation, because ionizing radiation induced hydroxyl radicals might act in concert with copper-induced hydroxyl radicals. In the present experiment, we analyzed linkage between radiosensitivity and the mutation responsible for hepatitis in F(1) animals of a cross with the F344 rat. Our results clearly demonstrated an absence of any significant association. In addition, partial dominance for radiosensitivity was observed, and radiosensitive (F(1) x LEC) backcross rats were twice as numerous as their radioresistant counterparts, suggesting the possibility of control by two or more recessive genes. PMID- 10856973 TI - Endothelial cell function and thrombosis. AB - The endothelium is pivotal in the control of haemostasis and thrombosis because it is the primary source of many of the major haemostatic regulatory molecules. Healthy endothelial cells, unlike extravascular cells, are anticoagulant and antithrombotic. This is due to the regulated secretion of antiplatelet agents, including prostacyclin and nitric oxide. Following vessel injury, platelet adhesion to exposed matrix requires von Willebrand Factor, another endothelial cell product. Local generation of thrombin causes a series of receptor-mediated endothelial cell functional responses, while the surface of the endothelium is additionally the site for inactivation of thrombin by antithrombin, and its conversion to a coagulation inhibitor by interaction with thrombomodulin. Endothelial cells are also the source of circulating tissue-type plasminogen activator and its inhibitor, and Tissue Factor pathway inhibitor. In disease states, many of these endothelial cell properties are perturbed towards a more procoagulant and prothrombotic phenotype. PMID- 10856974 TI - Possible involvement of cytokines in diffuse intravascular coagulation and thrombosis. AB - Recently, basic and clinical advances have provided insights into the molecular events that link inflammation with blood coagulation and thrombosis. At least in cell culture, the inflammatory cytokines, especially tumour necrosis factor alpha (TNF) and interleukin 1-beta (IL-1), are major mediators that can elicit changes in cell phenotype. With respect to coagulation, one of the clot-promoting and one of the inhibitory pathways seem especially prone to modulation by these cytokines. Whenever Tissue Factor contacts the blood, coagulation is initiated rapidly. These cytokines can elicit Tissue Factor production on endothelium and monocytes. Thus, the cytokines elaborate Tissue Factor formation intravascularly. This contrasts with the normal situation in which Tissue Factor is located exclusively in the extravascular space, largely on fibroblasts, where it is expressed constitutively. Furthermore, cytokines, especially interleukin 6 (IL 6), can stimulate new platelet formation, and the new platelets responding to IL 6 have increased sensitivity to thrombin activation and increased procoagulant activity. Regulating the clotting process are a large number of anticoagulant and fibrinolytic mechanisms. The three major anticoagulant mechanisms appear to involve antithrombin-heparin, Tissue Factor pathway inhibitor (TFPI) and the Protein C pathway. Of these, the Protein C pathway appears to be the primary target for cytokine action. The Protein C pathway is initiated when thrombin binds to thrombomodulin (TM). TM is expressed constitutively on endothelium. In tissue culture, TNF, IL-1 or endotoxin lead to a slow loss of TM and endothelial cell Protein C receptor (EPCR) from the cell surface. In addition, Protein S levels decrease in patients with disseminated intravascular coagulation (DIC). Taken together, these results suggest that cytokines should elicit massive thrombotic responses when administered systemically. At near toxic levels, TNF fails to elicit an overt DIC or thrombotic response in patients, although sensitive markers of coagulation do detect changes in coagulation in response to TNF. In baboons, very high levels of TNF also fail to elicit fibrinogen or platelet consumption. However, if the Protein C pathway is blocked, these cytokines can elicit either DIC or deep-vein thrombosis, depending on the conditions. Thrombus formation is potently potentiated by impeding flow and/or by catheterization. DIC is facilitated by providing membrane surfaces, possibly mimicking complement mediated platelet activation/damage that occurs in shock. Thus, available evidence suggests important roles for inflammatory cytokines in DIC and thrombosis, but they seem insufficient by themselves to elicit overt thrombotic responses without secondary stimuli. Current data suggest that anti inflammatory drugs are a viable candidate to blocking DIC or thrombosis without impairing the haemostatic balance. PMID- 10856975 TI - Tissue factor pathway. AB - Blood coagulation is initiated in response to vessel damage in order to preserve the integrity of the mammalian vascular system. The coagulation cascade can also be initiated by mediators of the inflammatory response, and fibrin deposition has been noted in a variety of pathological states. The cascade of coagulation zymogen activations which leads to clot formation is initiated by exposure of flowing blood to Tissue Factor (TF), the cellular receptor and cofactor for Factor VII (FVII). FVII binds to the receptor in a I:I stoichiometric complex and is rapidly activated. FVIIa undergoes an active site transition upon binding TF in the presence of calcium which enhances the fundamental properties of the enzyme. This results in rapid autocatalytic activation of FVII to FVIIa, thereby amplifying the response by generating more TF-FVIIa complexes. The TF-FVIIa activates both FIX and FX. Further FXa generation by the FIXa-FVIIIa-Ca2+ phospholipid complex is required to sustain the coagulation mechanism, since the TF-FVIIa complex is rapidly inactivated by Tissue Factor pathway inhibitor (TFPI). TFPI circulates in plasma, is associated with vascular cell surface and is released from platelets following stimulation by thrombin. TFPI requires the formation of an active TF-FVIIa complex and FXa generation before inhibition can occur. TFPI prevents further participation of TF in the coagulation process by forming a stable quaternary complex, TF-FVIIa-FXa-TFPI. PMID- 10856976 TI - Endogenous mediators and thrombophilia. AB - Platelets are one of the most important components of primary haemostasis. Since they lack a nucleus, their functional characteristics are determined at the time of production. The role of platelets in thrombosis is further modified by the interaction with vascular mediators that are endogenously produced in response to a variety of stimuli. This chapter discusses the factors that influence platelet production, the interaction with endogenous mediators, and the potential therapeutic benefits achieved by modifying this interaction in the clinical setting. PMID- 10856977 TI - Activation markers of coagulation. AB - Advances in our understanding of the biochemistry of the haemostatic mechanism have led to the development of sensitive methods for measuring peptides, enzyme inhibitor complexes, and enzymes that are liberated with the activation of the coagulation system in vivo. Studies employing these markers have provided important mechanistic information regarding haemostatic mechanism function both under normal conditions and in response to pathogenic stimuli. While assays for particular components can denote the presence of a 'biochemical' hypercoagulable state prior to the appearance of overt thrombotic phenomena, most of these markers thus far have not been shown to be useful in managing individual patients. Properly designed prospective studies will be required to determine whether these assay techniques will aid in the identification of patients predisposed to thrombotic events or the monitoring of antithrombotic therapy. PMID- 10856978 TI - Recent insights into antiphospholipid antibody-mediated thrombosis. AB - The clinically relevant antiphospholipid antibodies (APA) include anticardiolipin antibodies and lupus anticoagulant. Most autoimmune APA require the presence of a cofactor for phospholipid binding, and the growing list of candidate cofactors has prompted redefinition of APA to 'antiphospholipid protein antibodies'. Current evidence favours beta2-glycoprotein I (beta2GPI) and prothrombin as the primary antigens for anticardiolipin antibodies and lupus anticoagulant respectively. Patients with APA show a predisposition for venous and arterial thromboembolism, recurrent fetal loss, thrombocytopenia and a number of neurological syndromes and miscellaneous conditions. The association between APA and thrombosis has been well documented, but a definite mechanism remains to be clarified. Proposed mechanisms have included disruption of endothelial regulatory processes, impairment of fibrinolysis, augmented platelet activation and/or adhesion, inhibition of antithrombin activity and negation of the anticoagulant effects of beta2GPI and annexin V. In this review we describe recent insights into the role of beta2GPI as a natural anticoagulant, the procoagulant effects of APA on the Protein C system, the interactions between APA and prothrombin resulting in augmentation of thrombin generation, and cellular expression of Tissue Factor in patients with APA. Cellular immunity to beta2GPI is also discussed. Elucidation of these pathophysiological mechanisms may shed further light on the association between APA and thrombosis. PMID- 10856979 TI - Fibrinolysis and thrombosis. AB - The fibrinolytic system generates plasmin, which dissolves fibrin in haemostatic plugs and in thrombi. It is often regarded simply as a secondary phenomenon responsive to the generation of thrombi but it is, rather, in dynamic balance with fibrin formation, such that abnormalities in either can lead to thrombosis. This chapter summarizes the fibrinolytic system and its regulation. It considers the components of the system in blood, both in plasma and in circulating cells, with emphasis on protease-inhibitor balance. It goes on to discuss local fibrinolytic potential in thrombi, both venous and arterial, and in the diseased vessel wall, presenting evidence that increased local inhibition of fibrinolysis by PAI-1, PAI-2 and alpha2-antiplasmin is intimately involved in thrombus stability and in the generation of fibrin-rich vessel wall lesions. Finally, it reviews the evidence that defective plasma fibrinolysis has a causal role in venous and arterial thrombosis. PMID- 10856980 TI - Rheological influences on thrombosis. AB - Blood flow plays important roles in the localization and morphology of thrombosis within the circulation. Blood flow properties (rheological variables) are associated with thrombotic risk factors and thrombotic risk; conversely their modification may reduce thrombotic risk. PMID- 10856981 TI - Hyperhomocysteinaemia. AB - Homocysteine is a sulphur-containing amino acid that is derived primarily from protein of animal origin. Classical homocystinuria is an inherited metabolic disorder that arises from defects in either the re-methylation or trans sulphuration pathways of homocysteine metabolism and leads to skeletal abnormalities, mental retardation and a high risk of vascular disease. In contrast, moderate hyperhomocysteinaemia is associated with an increased risk of both arterial and venous thrombotic disease but no other abnormalities. This increased risk appears to be independent of other conventional risk factors. Many cases of hyperhomocysteineaemia have been attributed to defects in the enzyme cystathionine-beta-synthase (CBS) but this accounts for less than 1.5% of cases. A thermolabile variant of the enzyme methylenetetrahydrofolate reductase (MTHFR) arises from a C --> T transition at nucleotide 677 in the MTHFR gene resulting in an alanine-to-valine substitution. While the mutation does not appear to be associated with an increased risk of vascular disease, it results in excessively high homocysteine levels in response to a low or low-normal serum folate. Supplementation of the diet with folate, B6 and B12 can reduce homocysteine levels and this is the mainstay of treatment. Supplementation of grain with folate is undertaken in the USA to reduce the risk of neural tube defects in pregnant women. However, by reducing plasma homocysteine levels, it is estimated that this will save up to 50,000 lives per annum. PMID- 10856982 TI - The molecular genetics of familial venous thrombosis. AB - In the past few years, important advances have been made in the identification of factors predisposing to familial thrombophilia. Particular attention has been paid to the characterization of known inherited defects and their genotype phenotype relationship, and to studying the interaction between single or multiple inherited conditions and acquired risk factors for venous thrombosis. The recent discovery of 'new' and very common genetic lesions predisposing to thrombosis has greatly expanded the interest in this field. Hereditary predisposition to venous thrombosis may be related to lesions in one or more of 10-15 genes encoding antithrombin, Protein C, Protein S, Factor V, prothrombin, enzymes of the homocysteine metabolic pathway, fibrinogen, heparin cofactor II, plasminogen and thrombomodulin. About 500 different gene lesions (substitutions, deletions, insertions) have so far been reported to affect these genes in patients with thrombotic disease. Because there are potentially multiple interactions between genetic and environmental factors, familial thrombophilia is now considered to be a multifactorial disease. The aim of this chapter is to review aspects of the molecular genetics of familial thrombophilia. In particular, those gene/protein defects for which there is convincing evidence of an association with familial thrombosis will be examined in detail. PMID- 10856983 TI - Genetic determinants of arterial thrombosis. AB - This chapter describes examples of genetic variation involved in the function or regulation of a number of haemostatic proteins involved in the thrombotic process. In each case, the data suggest associations between genotype and disease and, particularly in the case of fibrinogen, PAI-1, Factor VII and Factor XIII, there is interaction between genotype and environment in determination of the relevant plasma level, providing a possible explanation for the differential response of individuals to their environment. PMID- 10856984 TI - Deep-vein thrombosis of the lower limbs: diagnosis and management. AB - Acute deep venous thrombosis (DVT) of the lower extremities is a serious and potentially fatal disorder which often complicates the course of severely ill, hospitalized patients but may also affect ambulatory and otherwise healthy people. It is uncommon in young individuals and becomes more frequent with advancing age. The clinically important problems associated with DVT are death from pulmonary embolism (PE), morbidity resulting from the acute event, the post thrombotic syndrome, and the inconvenience and side-effects of investigations and treatment. Furthermore, an often underemphasized problem is the anxiety that may occur in those patients who have suffered a thrombotic episode. PMID- 10856985 TI - Lipoproteins and the haemostatic system in atherothrombotic disorders. AB - The remarkable extent to which interactions between the plasma lipoproteins, inflammatory factors and the haemostatic system contribute to the response to injury and growth of the plaque in atherosclerosis is being increasingly documented. High plasma concentrations of very-low density (VLDL) and low-density lipoproteins (LDL), together with oxidatively modified LDL and lipoprotein (a), can induce responses in vascular endothelial cells, smooth muscle cells, monocytes/macrophages, platelets, neutrophils and humoral factors that are in a variety of ways both procoagulant and antifibrinolytic. Plasma high-density lipoproteins appear to promote anticoagulant mechanisms. Post-prandial lipaemia is associated with transient changes in factor VII which may be indicative of temporary hypercoagulability. The cellular and humoral effects of LDL and VLDL on the haemostatic system appear to be largely reversible, which may help to explain the prompt improvement in the atherothrombotic state gained by correction of hyperlipidaemia. PMID- 10856986 TI - Haemostatic function, arterial disease and the prevention of arterial thrombosis. AB - Recent years have seen the expansion of information linking raised plasma levels of individual clotting factors and evidence of disturbances of fibrinolytic activity with the risk of thrombotic manifestations of arterial disease, both in community-based, apparently healthy populations and in patients with known atherosclerosis. Some of these prothrombotic changes in the haemostatic system may result partly from underlying chronic inflammation or acute infection and may, in turn, contribute substantially to the thrombotic risk which accompanies these underlying processes. The importance of the coagulation system in the pathogenesis of arterial thrombosis is further illustrated by the benefit in the Thrombosis Prevention Trial of low-intensity, dose-adjusted warfarin in the primary prevention of ischaemic heart disease. Clinical trials of bezafibrate, which is being used for its fibrinogen-lowering as well as lipid-modifying properties, are in progress. PMID- 10856987 TI - [Tuberculosis continues to grow!]. PMID- 10856988 TI - [The importance of the tuberculin test in adults with bacteriologically confirmed pulmonary tuberculosis]. AB - The tuberculin skin test is used worldwide to assess the tuberculosis infection in humans and animals. It is conventionally admitted that a reaction larger than 10 mm expresses a consumptive infection. It is possible to find a reaction smaller than 10 mm in patients with active disease, like bacteriological confirmed pulmonary tuberculosis, that is a negative reaction. 1719 patients with recently discovered bacteriological confirmed pulmonary tuberculosis were tested with PPD-IC65 solution 2 U/0.1 ml; 497 of them (28.9%) reacted less than 10 mm; 151 of these were retested with PPD solution 10 U/0.1 ml, in 11.5% of them the test was negative. PMID- 10856989 TI - [Group tuberculous contamination in preschoolers]. AB - The study of a new collective tuberculosis contamination is proper for us to report news about the antituberculosis fight. In 1997, an ill teacher with cavity produced 19 cases of tuberculin turnings in a class with 25 children, but none of them had tuberculosis. We tried to find an explanation. We studied another 3 epidemiological events. In 1970, a teacher from Holbav produced 80 tuberculin turnings with 42 ill children. In Sacele another teacher with cavity produced 39 tuberculin turnings with 5 ill children. In Prejmer, in 1988 a sick child with cavity made himself 28 tuberculin turnings with a single case of illness. We tried to explain these differences: similar sources (patients with cavities) and the same conditions of illness but different effects. Our epidemiological investigations showed that in Holbav only 3.6% of children had BCG vaccine; in Sacele were 18%; in Prejmer were 98%; in Codlea 96%. BCG vaccination prevents tuberculosis. But in this case is another problem, did all the tuberculin turnings signify a virulent infection? After a year when we tested again the children, we found that for 58% of tuberculin turnings the reaction diminished considerably. We consider that, this is the particularity of the evolution of the postvaccinal allergy. We conclude that hyperreactivity may be a transitory exacerbation of the postvaccinal allergy and not an evidence of the infection. BCG vaccination reduces the risk of tuberculosis even when the infection is important and virulent. PMID- 10856990 TI - [Radiological observations in relation to occupational exposure to silicon carbide (SiC)]. AB - There have been investigated radiologically 191 workers from a SiC factory, exposed to dust concentrations exceeding largely that TLV and with a content of non-reactive SiO2 less than 1%. Radiological examinations were performed systematically at a three years interval, beginning with 1988. Radiology interpreting was performed according to ILO 1980 classification for persistent radiological opacities due to mineral dusts. Our results sustain the idea of the pneumoconiogen potential of SiC: there is a high prevalence of radiological findings (16.2%) and the opacities have an extend from 2 to 6 areas and the profusion from 0/1 to 2/2. PMID- 10856991 TI - [Electrical impedance tomography, a new imaging method?]. PMID- 10856992 TI - [Pulmonary actinomycosis]. PMID- 10856993 TI - [The therapeutic position of nedocromil sodium (Tilade) in the treatment of bronchial asthma]. PMID- 10856994 TI - [Breast tuberculosis--a case report]. AB - The authors report a case of breast tuberculosis in a 58-year-old patient in which the diagnosis was made with difficulty, initially the case being interpreted as a neoplastic process. This diagnosis should be considered in such cases. In our area, in the present tuberculosis endemic, the cases of tuberculosis with extrapulmonary localisation are more and more frequent. PMID- 10856995 TI - [Tumor markers and genetic changes in lung cancer]. PMID- 10856996 TI - [Serum tumor markers in lung cancer]. PMID- 10856997 TI - [The reasons for noncompliance with antitubercular treatment and the ways to combat it]. PMID- 10856998 TI - [Smoking in teenagers]. PMID- 10856999 TI - Principles and paradigms used in human medical ethics can be used as models for the assessment of animal research. PMID- 10857000 TI - Percutaneous intravenous injection in neonatal mice. PMID- 10857001 TI - Marburg and Ebola virus infections in laboratory non-human primates: a literature review. AB - BACKGROUND AND PURPOSE: Several non-human primate species are used as laboratory animals for various types of studies. Although importation of monkeys may introduce different diseases, special attention has recently been drawn to Marburg and Ebola viruses. This review presented here discusses the potential risk of these viruses for persons working with non-human primates as laboratory animals by focusing on epidemiology, virology, symptoms, pathogenesis, natural reservoir, transmission, quarantine of non-human primates, therapy, and prevention. CONCLUSION: A total of 23 Marburg and Ebola virus outbreaks causing viral hemorrhagic fever has been reported among humans and monkeys since the first outbreak in Marburg, Germany in 1967. Most of the 1,100 human cases, with nearly 800 deaths, developed in Africa due mainly to direct and intimate contact with infected patients. Few human cases have developed after contact with non human primates used for various scientific purposes. However, adequate quarantine should be applied to prevent human infections not only due to Marburg and Ebola viruses, but also to other infective agents. By following proper guidelines, the filovirus infection risk for people working with non-human primates during quarantine exists, but is minimal. There seems to be little risk for filovirus infections after an adequate quarantine period. Therefore, non-human primates can be used as laboratory animals, with little risk of filovirus infections, provided adequate precautions are taken. PMID- 10857002 TI - Histologic, cytologic, and bacteriologic examinations of experimentally induced Salmonella typhimurium infection in Lewis rats. AB - BACKGROUND AND PURPOSE: Histopathologic changes, cellular composition, and bacterial spreading were studied in rat spleen after experimentally induced infection with Salmonella typhimurium. METHODS: Lewis rats were inoculated intraperitoneally with 10(6) bacteria. Spleen weight, cell numbers, and cell surface markers were studied together with histopathologic changes, and expression of inducible nitric oxide synthase (iNOS). The spread of bacteria to blood, spleen, liver, mesenteric lymph nodes, lung, and kidney was studied at 12 hours, and 1, 3, 7, 14, and 28 days after inoculation. RESULTS: Experimentally induced infection caused an increase in spleen weight and leukocyte numbers, and a decrease in CD49d, on postinoculation days (PID) 3 through 7. Numerous granulomas were disseminated throughout the splenic red pulp also on PID 3 through 7. From PID 14 on, clearance of cellular exudate and regeneration of tissue structure were observed. Massive expression of iNOS was seen on PID 3. Bacterial growth was observed in liver and spleen from 12 hours to 14 days after inoculation. Bacteria were detected in blood on PID 3 and mesenteric lymph nodes were infected from PID 3 through 14. CONCLUSIONS: Salmonella typhimurium was rapidly taken up by the reticuloendothelial system. The infection induced weight increase and reversible changes in the spleen, peaking on PID 3 with granuloma formation and infiltration with macrophages. On PID 3, extensive production of iNOS within the granulomas was observed, suggesting initial killing of phagocytosed bacteria, followed by bacterial clearance and tissue regeneration. Cell surface marker expression on CD4+ T cells indicated no change in their numbers; however, there was a time-dependent change in expression of CD49d. PMID- 10857003 TI - Histopathologic changes in the brain, heart, and skeletal muscle of rhesus macaques, ten days after exposure to soman (an organophosphorus nerve agent). AB - BACKGROUND AND PURPOSE: Soman, an organophosphorus, anticholinergic, chemical warfare nerve agent, is studied at few research facilities, and there have been few pathologic studies of soman-exposed primates. We describe the brain, heart, and skeletal muscle lesions, review lesions described in literature, and discuss possible pharmacologic mechanisms for soman-induced neuron necrosis. METHODS: In this retrospective, histopathologic study, records were obtained for 36 rhesus macaques (Macaca mulatta) that were euthanized 10 days after soman exposure, from a larger group of 103 monkeys that were exposed to soman and used for pharmacologic and lethality studies. RESULTS: Brain lesions were seen in 9 of 15 animals that convulsed and in only 1 of 21 that did not convulse. The brain lesions in our primates were limited to the hippocampus, amygdala, and thalamus (of one animal), and consisted of neuron necrosis and dropout, spongiosis, gliosis, astrocytosis, and vascularization. Heart lesions consisted of myocardial degeneration and necrosis. Three animals had brain and heart lesions, 7 had brain lesions only, and 3 had heart lesions only. Skeletal muscle lesions, although minimal to mild, were in most of the animals, whether they had convulsed, but most had muscular tremors. These lesions were in the biceps brachii (11 of 22 monkeys), anterior tibialis (8/22), biceps femoris (7/22), flexor carpi radialis (5/22), gastrocnemius (3/22), and diaphragm (1/22). The limited literature on soman lesions in primate brain and heart, and the limited information on skeletal muscle lesions, is reviewed. CONCLUSIONS: Brain lesions were not as wide-spread as reported in other studies of primates and rodents, and were significantly associated with convulsions. Unlike other studies using rodents, we observed poor correlation between heart and brain lesions; thus, a single hypothesis to explain the pathogenesis for the brain and heart lesions may be difficult to establish. PMID- 10857004 TI - Role of cytoprotectants and nitric oxide inhibition in nonsteroidal anti inflammatory drug-induced gastroduodenal injury in the rat. AB - BACKGROUND AND PURPOSE: Nonsteroidal anti-inflammatory drugs (NSAIDs) induce gastroduodenal injury and ulceration. The pathogenesis is uncertain, although reductions in cytoprotective prostaglandins and nitric oxide (NO) have been proposed. The effects of several cytoprotective agents on inhibition of gastroduodenal ulcerogenesis induced by CI-987, a novel NSAID, were evaluated in Wistar rats. METHODS: Male Wistar rats were given CI-987 orally (p.o.) at a dosage of 300 or 450 mg/kg of body weight or subcutaneously (s.c.) (3 x 50 mg/kg), alone or with misoprostol pretreatment (2 x 1 mg/kg, p.o.). In a second experiment, rats were pre-treated with 2 ml of gelusil p.o., 500 mg of sucralfate/kg, p.o., 100 mg of ranitidine/kg s.c., or 200 mg of N omega-nitro-L arginine methyl ester (L-NAME)/kg, s.c.. Duodenal injury was induced by administration of 450 mg of CI-987/kg, p.o., 3 x 50 mg of CI-987/kg, s.c., or 300 mg of cysteamine/kg, s.c. Animals were euthanized within 24 to 48 hours, and the gastrointestinal tract was examined for evidence of gross or microscopic change. RESULTS: The L-NAME significantly reduced the incidence and severity of gastroduodenal injury induced by CI-987 and cysteamine. Prostaglandin ameliorated duodenal lesions induced by CI-987 given s.c., and Gelusil, ranitidine, and sucralfate were without effect on duodenal lesions induced by NSAID. CONCLUSIONS: Preemptive blockade of NO synthase is important in preventing NSAID-induced duodenal injury in rats. Inhibition of cytoprotective prostaglandins and enhanced acid-induced damage are unlikely to be primary mechanisms underlying NSAID induced duodenal injury in rats. PMID- 10857005 TI - Morphologic and biochemical changes caused by experimentally induced dicroceliosis in hamsters (Mesocricetus auratus). AB - BACKGROUND AND PURPOSE: The aim of the study reported here was to investigate the pathomorphologic changes caused by experimentally induced dicroceliosis and their correlation with hepatobiliary function. METHODS: Studies were carried out at days 80 and 120 after oral inoculation of hamsters with 40 metacercariae of Dicrocoelium dendriticum. RESULTS: The parasite-induced pathologic changes were assessed by presence of fluke eggs in feces, increased plasma alanine transaminase and aspartate transaminase activities and morphologic alterations. Dicroceliosis was characterized by bile ductular proliferation and enlargement of the bile duct surface area caused by hyperplastic cholangitis in septal bile ducts. The liver from infected animals contained portal tracts infiltrated with small to moderate numbers of lymphocytes, macrophages, and eosinophils. Simultaneously, there was an increase in portal tract collagen that extended to the interlobular septa and caused pressure atrophy of the hepatic parenchyma. The concentration of thiobarbituric acid-reactive substances and the ratio of oxidized to reduced glutathione, measured as markers of oxidative stress, were significantly increased. CONCLUSIONS: The presence of oxidative alterations could be related to the morphologic evidence of chronic inflammatory response as well as to liver cellular injury indicated by cellular swelling, and increased presence of peroxisomes and lysosomes. PMID- 10857006 TI - Experimentally induced pneumonia in scid/beige mice, using a bovine isolate of Pasteurella haemolytica. AB - BACKGROUND AND PURPOSE: Intranasal challenge of immunocompetent mice with Pasteurella haemolytica results in little or no pulmonary inflammation. The study reported here was designed to investigate the inflammatory response in the lungs of immunodeficient scid/beige mice after similar challenge. METHODS: Fifty-five scid/beige mice were challenged intranasally with saline or one of three doses (2.8 x 10(6), 3.4 x 10(9), or 3.3 x 10(11) colony-forming units [CFU]/ml) of P. haemolytica. The lungs were examined for changes in weight, bacterial count, and presence of gross and microscopic lesions at 24, 48, or 96 hours after challenge. RESULTS: Intranasal challenge with concentrations > or = 3.4 x 10(9) CFU/ml of P. haemolytica induced significantly heavier lung weight, with severe pulmonary lesions, and development of suppurative and fibrinous bronchopneumonia in dose- and time-dependent manner 48 hours after challenge. Pasteurella haemolytica was consistently isolated from the lungs at 24 hours after challenge. CONCLUSIONS: Bronchopneumonia was induced by P. haemolytica in mice without manipulation of the mouse or the bacteria. The lesions were similar to those that develop in the lungs of cattle infected with P. haemolytica and indicate potential use of the model for the study of this host/bacterial interaction. PMID- 10857007 TI - Telemetric evaluation of body temperature and physical activity as predictors of mortality in a murine model of staphylococcal enterotoxic shock. AB - BACKGROUND AND PURPOSE: Hypothermia and death are used as experimental markers in murine models of staphylococcal enterotoxic shock. This study determined whether body temperature and physical activity, monitored telemetrically, could predict impending death and provide an earlier, more humane experimental endpoint. METHODS: The study consisted of two iterations (experiments 1 and 2) to determine reproducibility of the model. Each experiment consisted of 24 BALB/c mice surgically implanted with intra-abdominal telemetry transmitters and then injected intraperitoneally with sublethal or lethal doses of staphylococcal enterotoxin B (SEB) and/or lipopolysaccharide (LPS). Core body temperature and physical activity were continuously monitored in all mice for 10 days before, and 5 days after, injections. Additionally, in experiment 2, subcutaneous temperatures were compared with core body temperatures obtained by telemetry. RESULTS: Body temperature and physical activity were reduced in mice after administration of SEB and LPS, or LPS alone, but not SEB only. There was a significant (P < 0.05) correlation between mortality and body temperature (P = 0.0077), but not physical activity (P = 0.97). CONCLUSION: Body temperature proved to be an early indicator of mortality in this murine model of staphylococcal enterotoxic shock. PMID- 10857008 TI - Improved longevity and functionality of a canine model providing portal vein and multi-site intestinal access. AB - BACKGROUND AND PURPOSE: The canine intestinal and venous access port (IVAP) model is valuable for investigating hepatic elimination and region-specific intestinal absorption of pharmaceuticals. Previously, long-term functionality of this preparation has been variable. METHODS: Catheters of different construction were placed in the proximal and distal portions of the small intestine, colon, and portal vein of subject animals and were attached to separate subcutaneous access ports. Intraoperative, postoperative, and long-term maintenance techniques were developed, modified, and analyzed. RESULTS: Intestinal catheter infections and access site failures were associated with breakdown at the intestinal insertion site. The ileal catheter was prone to obstruction with ingesta. A modified Witzel technique, specialized port-catheter systems, scheduled port-flushing methods, and venous port infection treatment protocols improved the model's longevity. CONCLUSIONS: The canine IVAP model is a powerful tool for investigation of regional differences in intestinal absorption and hepatic elimination of drugs. Other researchers can derive increased longevity with the IVAP model by using the technical modifications detailed here: strict sterile technique, closed-end slit valve catheters, GPV ports, the Witzel tunnel technique, routine portal vein infection surveillance, 50% dextrose intestinal catheter infusion, rapid removal of infected intestinal catheters, and critical appraisal of their results. Longevity of the model continues to be improved. PMID- 10857009 TI - Body composition of growing and adult cats as measured by use of dual energy X ray absorptiometry. AB - BACKGROUND AND PURPOSE: Total body scans were performed on 89 domestic cats of various ages, using dual energy x-ray absorptiometry (DEXA) to determine body composition, including fat, lean, and bone mineral content. Bone mineral density results from scans also are presented. METHODS: This cross sectional study included data for cats from a closed colony and from privately owned cats. Data were grouped by age and were analyzed by sex and reproductive status to provide information as to the rate of growth of the individual components of body composition. RESULTS: The results indicate that the rate of accretion of bone mineral, fat, and lean tissue differs throughout maturation and by sex. Regressions are provided to highlight age- and sex-related differences. CONCLUSIONS: The results of this study emphasize the benefits of examining the growth of each component of body composition when studying the effects of nutrition, disease processes, or therapeutic interventions. PMID- 10857010 TI - Cortisol and prolactin concentrations during repeated blood sample collection from freely moving, mouse-sized mammals (Phodopus spp.). AB - BACKGROUND AND PURPOSE: Validation of a method for obtaining blood samples that does not change cortisol or prolactin concentrations yet allows serial blood samples to be collected from animals under anesthesia, without prior handling, from freely interacting social groups of small mammals. METHODS: Results from five experiments are reported. Male dwarf hamsters (Phodopus spp.) were housed in modified home cages under continuous flow of compressed air that could be switched to isoflurane in O2 vehicle without approaching the cages. RESULTS: Dwarf hamsters respond to manual restraint with behavioral distress and increase in the concentration of the dominant glucocorticoid, cortisol, and decrease in prolactin concentration. Both effects are evident within one minute. In contrast, when this new method was used, neither cortisol nor prolactin changed in response to repeated sample collection (up to 8 successive samples at 2 hour intervals), prolonged isoflurane exposure, or substantial blood volume reduction (30%). Prolactin concentration was suppressed and cortisol concentration was increased in response to stimuli from other hamsters tested without anesthesia. Suppression of prolactin concentration was graded in response to the degree of stress and equaled the pharmacologic reduction caused by bromocryptine mesylate (50 microg of CB154 x 3 days). CONCLUSIONS: The technique is superior to alternatives for studies of behavioral endocrinology of freely interacting small mammals. PMID- 10857011 TI - Outbreak of Orthoreovirus-induced meningoencephalomyelitis in baboons. AB - BACKGROUND AND PURPOSE: Spontaneous viral encephalitis is rare in the baboon; yet, during a 13-month period (1993-1994), eight juvenile baboons (Papio cynocephalus spp.) developed acute, progressive nonsuppurative meningoencephalomyelitis caused by an unknown agent. Clinical signs of disease included disorientation and truncal ataxia that rapidly progressed to hemiparesis or paraparesis. Clinicopathologic findings were not remarkable and appreciable gross lesions were not seen at necropsy. Microscopic examination revealed CNS lesions that were characterized by lymphoplasmacytic perivascular cuffing, microglial nodules, demyelination, axonal degeneration, vacuolization, and hemorrhage. Subsequently, a novel syncytium-inducing mammalian orthoreovirus was isolated from the brain tissue of five baboons with clinical signs of infection. METHODS: To confirm the etiologic role of the orthoreovirus, two juvenile baboons were inoculated with the virus, then were monitored for 6 weeks. RESULTS: Lesions similar to those seen in spontaneous cases were found in the CNS, and orthoreovirus was isolated from the brain of both animals. CONCLUSION: Analysis of the outbreak indicated juvenile baboons were most susceptible to disease and the virus had a possible incubation time of 46 to 66 days, but did not indicate a source of the virus or mode of transmission. PMID- 10857012 TI - Poor quality of oocytes from Xenopus laevis used in laboratory experiments: prevention by use of antiseptic surgical technique and antibiotic supplementation. AB - BACKGROUND AND PURPOSE: Episodic phases of continuous poor-quality oocytes obtained from South American Clawed Frogs (Xenopus laevis) often are observed. In publications dealing with the surgical technique of oocyte removal, the frogs' robust constitution and resistance against infections provided by magainins are pointed out. For this reason, clean rather than sterile conditions for the surgical procedure are mostly recommended. However, in most instances, antibiotics are added to the buffer medium when in vitro experiments are performed using oocytes. METHODS: After a long phase of poor oocyte quality at our facility, involving oocytes that had been obtained by use of a "clean" surgical procedure, we subsequently cultured oocytes in a buffer medium containing the three antibiotics: penicillin G, gentamicin, and streptomycin. RESULTS: During DNA injection experiments, the oocytes developed black spots on their surface by postoperative day two. Pure culture of the gram-negative non fermentative rod Pseudomonas fluorescens was obtained from the impaired oocytes; the isolate was resistant to the three antibiotics. By contrast, after aseptic surgical removal and culture of oocytes in buffer medium containing the antibiotics tetracycline and gentamicin, perfect oocytes without bacterial contamination were obtained. CONCLUSION: Whenever impaired oocyte quality is observed, microbial contamination should be considered as a possible cause. PMID- 10857014 TI - Rhesus monkeys with late-onset hydrocephalus differ from non-impaired animals during neonatal neurobehavioral assessments: six-year retrospective analysis. AB - BACKGROUND AND PURPOSE: A recent case study indicated that a hydrocephalic rhesus monkey had abnormal response patterns in a standardized neonatal primate assessment. We conducted a retrospective study to determine whether this assessment could also differentiate neonatal rhesus monkeys that appeared normal but developed signs of hydrocephalus later in life from neonates with normal development and no evidence of hydrocephalus. METHODS: One-hundred eighty-two rhesus monkeys were assessed on postnatal days 7, 14, 21, and 30. As neonates, clinical signs of hydrocephalus or other illnesses were not evident in any animal. Six monkeys developed signs of hydrocephalus between 5 months and 5 years of age, and each received confirmed diagnoses of hydrocephalus at necropsy. RESULTS: Compared with colony norms, the monkeys that developed hydrocephalus had diminished orientation abilities, more muscle tension, less behavioral evidence of distress, and more pronounced responses to some reflex-evoking stimuli, and difficulty in self-righting (day 7 only). Discriminant function analysis comparing the hydrocephalic animals with a matched control group provided a high probability of correct group assignment at days 7, 14, and 21. CONCLUSIONS: Some as yet undetermined factor may predispose some monkeys to develop hydrocephalus, which may also be reflected in different scores on neurodevelopmental test items during early infancy. PMID- 10857013 TI - Incidence of testicular lesions in a population of tree shrews (Tupaia belangeri). AB - BACKGROUND AND PURPOSE: The sexual activity of male tree shrews is socially influenced; therefore, the testicular lesions in adult male tree shrews were of interest. METHODS: The testes of 229 adult and 9 subadult male tree shrews were obtained during routine necropsy and were subjected to light microscopy. At one time, 138 animals were experimentally exposed to social conflicts. RESULTS: Hypospermatogenesis (testicular inactivity) was observed in social stress-exposed males up to two years of age. Seasonality of hypospermatogenesis could not be statistically supported. Testicular atrophy, observed in 21 animals, was neither stress- nor age-related; it developed unilaterally, with the left testis preferred. Testicular tumors developed in animals older than 2 years, with increasing frequency particularly of Leydig cell tumors in animals more than four year old. CONCLUSION: Testicular lesions were more frequently found in male tree shrews than they were observed in nonhuman primates kept at the German Primate Center. Connections to social stress were statistically supported, particularly with respect to hypospermatogenesis. Testicular tumors, in contrast, were distinctly age related. PMID- 10857015 TI - Atopic dermatitis in NC/Jic mice associated with Myobia musculi infestation. PMID- 10857016 TI - Ectoparasites in a pigeon colony. PMID- 10857017 TI - [Methods of searching for antigenic determinants for proteins with known primary structure]. AB - Theoretical and experimental methods for locating antigenic determinants of proteins with known amino acid sequences are discussed. These methods are systematized on the basis of the theoretical approaches applied, and the efficiency of various predictive methods is compared. Some examples of experimental epitope determination for a number of proteins are given. PMID- 10857018 TI - [Solid-phase synthesis of peptides containing arginine with an unprotected guanidine group]. AB - A new variant of the solid phase synthesis of arginine-containing peptides was proposed. The conditions for the attachment to the Wang polymer of N alpha-Fmoc arginine containing a protonated guanidine group were found. We demonstrated that this attachment is accompanied by neither racemization nor the attachment of the second Arg residue. Side reactions involving the guanidine group of arginine were studied, and methods for their prevention were proposed. The comparison of the carbodiimide method with a 1-hydroxybenzotriazole additive and a modified method with the use of Kastro's reagent for the introduction of N alpha-Fmoc-Arg residue with the unprotected guanidine group into the growing peptide chain demonstrated the advantages of the second method. Bradykinin and a peptide corresponding to the 584-591 sequence of the transmembrane gp41 from HIV-1 were synthesized by the method proposed here. PMID- 10857019 TI - [Pyridoxyl amino acid esters in peptide synthesis]. AB - Pyridoxyl residue was suggested to be used as a multifunctional protective and modifying group in peptide synthesis. The modification was carried out by introducing the pyridoxyl residue in free or partially protected peptides or by the addition of amino acid pyridoxyl esters by the methods of conventional peptide synthesis without the removal of the pyridoxyl group at the terminal stages of the synthesis (the second approach is more convenient). Pyridoxyl residue was also used as a spacer in solid phase peptide synthesis. It was attached to the polymer by the alkylation of the hydroxyl groups or of the pyridine ring of the pyridoxyl derivatives with the chloromethylated styrene divinylbenzene copolymer (the standard Merrifield resin). Potentials for the use of pyridoxyl derivatives in the synthesis of linear, multiplet, and cyclic peptides are discussed. PMID- 10857020 TI - [Point amino acid substitutions in Ca2+-binding centers of recoverin. III. Mutant with the fourth reconstructed Ca2+-binding site]. AB - Unlike wild type recoverin with only two (the second and the third) functioning Ca(2+)-binding sites out of four potential ones, the +EF4 mutant contains a third active Ca(2+)-binding site. This site was reconstructed from the fourth potential Ca(2+)-binding domain by the introduction of several amino acid substitutions in it by site-directed mutagenesis. The effect of these mutations in the fourth potential Ca(2+)-binding site of myristoylated recoverin on the structural features and conformational stability of the protein was studied by fluorimetry and circular dichroism. The apoform of the resulting mutant (free of Ca2+ ions) was shown to have a higher calcium capacity, significantly lower thermal stability, and noticeably different secondary and tertiary structures as compared with the apoform of wild type recoverin. PMID- 10857021 TI - [Conformational analysis of biologically active RGD-containing cyclopentapeptides]. AB - The theoretical conformational analysis of the biologically active RGD-containing pentapeptide cyclo(-Arg-Gly-Asp-Phe-DVal-), an inhibitor of laminin P1 interaction with its receptor, was performed. The space of permissible torsional angles of the backbone of the molecule was studied by the Monte Carlo method. From the large number of predicted low-energy conformers with various packings of the cyclic moiety of this pentapeptide, only those were selected that corresponded to stable structures of the model linear tripeptide Ac-Ala-Gly-Asp NHMe. This peptide simulated the spatial possibilities of the backbones of RGD containing fragments of laminin, vitronectin, and fibronectin. We selected several dozen structures that may be potential biologically active conformers, but only a few of them were capable of forming stable intramolecular hydrogen bonds. We assumed that a biologically active conformer of cyclo(-Arg-Gly-Asp-Phe DVal-) can be present in significant amounts in an equilibrium mixture in solution along with other conformers without necessarily dominating among them. PMID- 10857022 TI - [Binding sites for A2 and A24 monoclonal antibodies on the alpha-latrotoxin molecule]. AB - alpha-Latrotoxin (alpha-LTX) binding sites to functionally active monoclonal antibodies (MA) A4 and A24 were localized using three approaches: hydrolysis of the toxin followed by the N-terminal sequencing of immunoreactive peptides; the study of antibody interaction with several recombinant alpha-LTX fragments; and Western immunoblotting of synthetic overlapping peptides (6-8 aa) whose structures correspond to that of the immunoreactive alpha-LTX fragment. It was shown that the MA A4 epitope is located within the F234-M294 protein fragment and that of MA A24 interacts with the fragment 347FDKDIT352. PMID- 10857023 TI - [Chemical ligation and recombination of DNA fragments by formation (exchange) of disulfide bonds, located in the sugar-phosphate backbone]. AB - Effective methods of the directed introduction of diphosphoryl disulfide bridges into hairpin DNA duplexes in place of natural phosphodiester groups were developed using the H2O2-effected ligation of 3'- and 5'-thiophosphorylated oligonucleotides or the autoligation of a preactivated oligonucleotide derivative with a phosphorothioate-bearing oligomer. The postsynthetic recombination of the disulfide-linked oligonucleotide fragments was characterized. It was shown that, along with template-directed reactions, out-of-duplex formation and exchange of diphosphoryl disulfide bonds in the DNA sugar-phosphate backbone may occur. In modified hairpin DNA, a spontaneous exchange of disulfide-linked fragments virtually does not take place because of the intramolecular duplex formation. PMID- 10857024 TI - [Immunoenzyme method for detecting microbial producers of palytoxin]. AB - A competitive ELISA using the intact toxin as a coating antigen for detecting palytoxin was developed. This immunoassay allows palytoxin (PTX) to be determined in the range of 6-250 ng/ml. In sensitivity, this determination is comparable with RIA but is three times inferior to ELISA using monoclonal antibodies. Inhibition experiments using some toxins of marine invertebrates proved the serological specificity of the palytoxin binding to antibodies. Both the indirect and competitive ELISA were used to find PTX-producing bacteria among 420 isolates of sea bacteria. It was found that gram-negative bacteria Aeromonas sp. and Vibrio sp. associated with toxic samples of the soft coral Palythoa sp. produced compounds antigenically related to PTX. PMID- 10857025 TI - Financing veterinary medical education. PMID- 10857026 TI - Comments regarding the claimed effectiveness of bovine colostrum. PMID- 10857027 TI - Comments on anesthesia and analgesia. PMID- 10857028 TI - "Integrationalism"--the way of the future. PMID- 10857029 TI - An ethicist's commentary on euthanizing deaf Dalmatian puppies. PMID- 10857030 TI - Vertical and horizontal transmission of Neospora caninum in dairy herds in Quebec. AB - Neospora caninum is an important cause of abortion in dairy cattle. The objective of this observational study was to estimate the rate of vertical transmission of N. caninum in dairy herds in Quebec and to investigate horizontal transmission in the same herds. The genealogy of cows from 23 dairy herds were examined. Prevalence of seropositive animals in herds studied varied from 4.3% to 61.8% (average, 21.9%). The overall rate of vertical transmission was estimated to be 44.4%, varying from 0% to 85.7%. Seven cases of horizontal transmission were identified in 6 of the 23 herds studied. Estimated vertical transmission rate varied from herd to herd, but appeared to be higher in herds with a high prevalence of seropositive animals. Although horizontal transmission was identified in 6 herds, it does not appear to be the major route of infection for N. caninum. PMID- 10857031 TI - Equine glaucoma: a retrospective study of 13 cases presented at the Western College of Veterinary Medicine from 1992 to 1999. AB - The prevalence of equine glaucoma seen by the ophthalmology service at the Western College of Veterinary Medicine (WCVM) was 6.5%. The majority of cases (11/13) were associated with clinical manifestations of uveitis. Congenital glaucoma was documented in 1 case, and primary glaucoma was diagnosed in a 12 year-old quarter horse. There were no breed or sex predilections evident. Affected horses were middle-aged to old (average age = 9.5 years, ranging from 2 weeks to 23 years). The clinical manifestations of equine glaucoma included blindness, diffuse corneal edema, corneal vascularization, buphthalmia, corneal striae, recurrent secondary ulcerative keratitis, and less commonly, iris bombe, tapetal hyper-reflectivity, complete pupillary occlusion from posterior synechiae, and optic disc cupping. Elevated intraocular pressure confirmed the diagnosis (n = 10), while 3 cases were normotensive with signs of glaucoma including corneal striae and buphthalmia. Affected eyes were treated medically and/or surgically. Regardless of the therapy instituted, the visual outcome was poor. Most affected eyes were blind at presentation or became blind within a few weeks. PMID- 10857032 TI - A preliminary trial comparison of several anesthetic techniques in cats. AB - The aim of this study was to investigate the effect of several drug combinations (atropine, xylazine, romifidine, methotrimeprazine, midazolam, or fentanyl) with ketamine for short term anesthesia in cats. Twelve cats were anesthetized 6 times by using a cross-over Latin square protocol: methotrimeprazine was combined with midazolam, ketamine, and fentanyl; midazolam and ketamine; romifidine and ketamine; and xylazine and ketamine. Atropine was combined with romifidine and ketamine, and xylazine and ketamine. Temperature, heart rate, and respiratory rate decreased in all groups. Apnea occurred in 1 cat treated with methotrimeprazine, romifidine, and ketamine, suggesting that ventilatory support may be necessary when this protocol is used. Emesis occurred in some cats treated with alpha 2-adrenoceptor agonists, and this side effect should be considered when these drugs are used. PMID- 10857033 TI - Lymphosarcoma of the pharyngeal region in a 7-month-old beef steer. AB - A steer presented for retropharyngeal swelling and dyspnea. Biopsy of the swelling indicated lymphoblastic lymphosarcoma, and necropsy revealed involvement of regional veins and arteries. The tumor was classified as a non-T, non-B lymphoma based on CD3 polyclonal antibody stains. Lymphosarcoma is unusual in this location, but should be considered with retropharyngeal swelling. PMID- 10857034 TI - Abnormal hematologic findings in an African hedgehog (Atelerix albiventris) with gastrointestinal lymphosarcoma. AB - A 4-year-old African hedgehog (Atelerix albiventris) was examined for weight loss and hematochezia, and was subsequently diagnosed with gastrointestinal lymphosarcoma. Abnormal hematological findings included marked leukocytosis with lymphocytosis and atypical circulating lymphocytes. This report represents the first documentation of hemogram abnormalities associated with gastrointestinal lymphosarcoma in this species. PMID- 10857035 TI - Tyzzer's disease in an 11-day-old foal. AB - An 11-day-old pony became depressed, anorectic, and pyrexic 2 days after the topsoil of its paddock had been turned over. Rapid progression to colic and head pressing occurred, despite intensive therapy for Tyzzer's disease, and the foal died within 7 h of the appearance of central nervous system signs. PMID- 10857036 TI - Lelystad-like strain of porcine reproductive and respiratory syndrome virus (PRRSV) identified in Canadian swine. PMID- 10857037 TI - Canine leptospirosis: serology. PMID- 10857038 TI - Fees--demon or delight? PMID- 10857039 TI - Diagnostic ophthalmology. Right superficial corneal ulcer with mild secondary anterior uveitis and osseous choristoma in a guinea pig. PMID- 10857040 TI - Frequency of coexistence of cytomegalovirus and Chlamydia pneumoniae in atherosclerotic plaques. AB - BACKGROUND: Cytomegalovirus (CMV) and Chlamydia pneumoniae (C. pneumoniae) antigens and DNA sequences have been demonstrated in atherosclerotic plaques by several investigators. Most significantly, CMV DNA was found both in atherosclerotic lesions as well as in uninvolved areas of aortas and carotid artery, whereas C. pneumoniae was mostly detected in advanced carotid atherosclerotic lesions. METHODS AND RESULTS: Atherosclerotic plaques removed from seventeen patients during carotid endarterectomy were analysed for the simultaneous presence of CMV and C. pneumoniae DNA sequences using polymerase chain reaction (PCR). Of the seventeen samples, nine (53%) were positive for CMV DNA sequences and seven (41%) contained C. pneumoniae DNA sequences. Four samples (24%) were positive for both CMV and C. pneumoniae DNA. CMV DNA or C. pneumoniae DNA was detected in 12 (71%) of 17 carotid plaques and 2 additional patients had high titers of antibodies to CMV. CMV DNA and C. pneumoniae DNA were found in the same tissue specimens in 4 (24%) patients. CONCLUSIONS: These results present evidence that CMV DNA and/or C. pneumoniae DNA can be detected in 71% of carotid atherosclerotic plaques and in some instances DNA of both agents in the same tissue. The possible pathogenetic role of these agents in the initiation or promotion of the development of atherosclerotic plaques deserves increased attention. PMID- 10857041 TI - Tobacco control in the Czech Republic. AB - Main points of tobacco control measures in the country are mentioned: smoking prevalence among population and health professionals (both about one third), smoking cessation availability (about 70 smoking cessation clinics), education of health professionals, both pre- and post-gradual, public-oriented actions, advertising, tobacco prices, legislation, cigarette consumption, mortality. PMID- 10857042 TI - Dyslipoproteinemias in industrial workers: relationship with hypertension and overweight. AB - The aim of the investigation was to study the lipid profile of industrial workers with attention to the relationship of lipid indexes with hypertension and overweight. 139 hypertensive male blue-collar workers were investigated, compared to 107 normotensive ones. The overweight and obese workers in each group were compared with normal body mass ones. Cholesterol and triacylglycerols (TG) were assayed by enzymatic tests CHOD-PAP and GPO-PAP respectively and the lipoprotein fractions were separated by precipitation method. Our results show high rate of industrial workers with dyslipoproteinemias, especially among the hypertensive and overweight and obese ones. The hypertension is associated with total cholesterol (TC), low density lipoprotein cholesterol (LDL-C) and low density lipoprotein TG (LDL-TG), while overweight is related with TG and high density lipoprotein cholesterol (HDL-C), effecting significantly on very low density lipoprotein TG (VLDL-TG). The high rate of persons with concentrations of the examined lipid indexes indicating moderate and high cardiovascular risk motivates multifactorial preventive strategy towards coping the hypertension, optimizing the nutritional habits, increase in physical activity and reduction of body mass. PMID- 10857043 TI - Effect of new quaternary bisammonium compounds on the growth and cell surface hydrophobicity of Enterobacter cloacae. AB - The effect of new quaternary bisammonium salts (QBAS) of series A: 1, 10 [bis(alkyldimethyl-diammonium)]-4,7-dioxo-3,8-dioxoundecandi bromides and series B: alpha-omega[bis(dodecyldimethyl-diammonium)]-3,x-dioxo-4,y- dioxoalcandibromides on the growth and surface hydrophobicity of E. cloacae strain was studied. The compounds of series A possess changing side substituent and the inhibition of growth displayed evident dependence of effect on length of alkyl with optimum at C9-C11 (cut off effect). In series B are compounds with changing joining chain between two ammonium cations where the mentioned effect was not observed. The cell surface hydrophobicity of tested strain was determined by the method of adhesion to hydrocarbon--xylene (BATH) and in salt aggregation test of ammonium sulphate (SAT). The effect of 1/4 of the MIC compounds series A with nonyl, decyl and undecyl reduced the adhesion of E. cloacae cells below 50% against the control. These compounds at sub-MICs decreased salt-aggregative ability of E. cloacae cells. The influencing of cell surface hydrophobicity was not found after affecting of compounds at sub-MIC from series B with the exception of moderate decrease at compound with joining chain of C7. The length of chain connecting two symmetrical parts of molecule has slight impact on the effect of bisammonium salts. The influence of QBAS at sub-MICs on cell surface hydrophobicity can interfere with pathogenic potential of E. cloacae. PMID- 10857044 TI - Influence of positioning of infants on long-term changes of cephalic dimensions. AB - The submitted investigation describes long-term changes of 3 main cephalic dimensions (head circumference, maximal length and maximal width of the head) and analyses the possible influence of positioning of infants after birth (prone, supine and side sleeping position) on these changes. Information about children aged 6 months to 3.99 years, where the need of up-to-date data is greatest, were collected as part of an extensive anthropological survey implemented in 1995 to 1997 in the entire Czech Republic. The authors confirmed the trend of debrachycephalization, which is manifested by a statistically significant increase of the maximal length of the head and a statistically significant decrease of the maximal width of the head, as compared with children examined in the anthropological survey in 1956 to 1962 (1). These changes were established in the group of boys (200 boys) as well as in the group of girls (167 girls). The differences of the magnitude of long-term changes between boys and girls were not significant. Evaluation of the long-term changes of the head circumference in the entire group of 366 children aged 0.5-3.99 years (the head circumference of one girl was not measured) revealed a statistically significant increase of this dimension. During the period from 1956/62 till 1996, the influence of positioning on the magnitude of long-term changes of head circumference was not proved. We can say the same about the maximal length of the head of boys and girls and about the maximal width of the head of girls. Only between three differently positioned groups of boys (prone, side, supine) statistically significant differences in the magnitude of long-term changes of the maximal width of the head were found (p < 0.05). Highly significant changes of the maximum width and maximum length of the head occurred as compared with a reference group in all three groups of positioning of infants and in both sexes. The trend of debrachycephalization seems to be thus a more potent factor, which affects long-term changes in the shape of the head, then the predominating sleeping position during the first months after birth. This conclusion is supported by the persisting trend of debrachycephalization, although the supine position is now preferred. PMID- 10857045 TI - Use and evaluation of the Czech version of the SF-36 questionnaire self-reported health status of medical students. AB - SF-36 questionnaires were completed by 231 medical students of the Faculty of Medicine in Hradec Kralove (1997, 1998). Results of measurements of eight health dimensions are presented here. Significantly lower values for bodily pain were found in the group of overweight students. Students with some reported cured diseases have significantly lower values for bodily pain and general health dimensions in comparison with students without any reported disease. In our sample a high rate of non-smokers (86.4% men and 93.6% women) and low rate of students with BMI > 25 (18.4% men and 3.8% women) were found. About 30% of respondents reported one or more cured diseases. In addition to the SF-36 questionnaire, students in 1998 completed also a special one-page form (3). The one-page form enabled direct estimates of the eight dimensions of the health status on a scale from 0% to 100%. This study compares the results of measurement of the health status for both instruments. Differences found here are compared and discussed with similar comparisons in an American study (3). Results in both studies are similar but not the same. An indirect measurement of health status with specific questions in the SF-36 is more objective than a direct measurement with the one-page form. Nevertheless, the SF-36 is limited in the number of possible answers for some dimensions (RP, RE). In that case, our results indicate that a percentage scale from the one-page form seems better. Additionally this study compares the results of the SF-36 in Czech medical students with comparable samples from other three European countries. On average, the health dimensions of SF-36 in Czech medical students achieved the worst values in comparison with samples from Switzerland, Germany and Great Britain. PMID- 10857046 TI - A comparison between the lifestyles of men and women--parents of school age children. AB - Women live longer than men and experience lower overall and specific mortality resulting from various diseases, even when younger. The reasons for this have yet to be satisfactorily explained. However, biological differences on one hand and differing lifestyles on the other might be responsible. The purpose of the study was to examine to what extent the lifestyle of men and women differs within a relatively homogeneous population group. The lifestyles of 4,353 parents of school age children (58% women and 42% men) were examined using questionnaires. The results show considerable differences between the genders. Men had worse dietary habits--consuming significantly less vegetables, fruit and milk, but too much meat; they consumed more processed meat and fat-containing items within the food sub-categories; they preferred less low-fat milk products and consumed less wholemeal products. Men more often consumed alcohol, drank more of it and often crossed the limits hazardous for health. There were more smokers among the men, they smoked more cigarettes and the non-smokers more often indicated passive exposure to cigarette smoke. Overweight and obesity occurred more often among men. Relatively minor differences, rather to the benefit of men, occurred in the field of leisure-time physical sporting activities, where slightly more men pursued regular sporting activities but in significantly higher amounts than the women, whereas the men did less regular daily walking. Women, as opposed to men, displayed more interest in comprehensive primary preventive medical examinations. The results obtained suggest that women lived a generally more healthy lifestyle than men within the examined homogeneous group of parents of school age children, consisting mostly of pairs of partners. They support the assumption that the healthier lifestyle of women very significantly contributes to their lower mortality. PMID- 10857047 TI - Thickening of the roentgenometrical gastric fold in the elderly: relation to peptic ulcer incidence. AB - Radiographic upper gastrointestinal barium examination is commonly used to diagnose peptic ulcer. However, little attention has been paid to its thickening, except in Menetrier disease and gastric carcinoma of Borrman type IV. The present study was undertaken to investigate the relation of age to roentgenometrical gastric fold width and history of peptic ulcer. The subjects were 724 men (35-64 years old) who participated in a periodic medical health examination and underwent radiographic upper gastrointestinal barium examination. The gastric fold width of the anterior wall in the body was evaluated by air-contrast examination and expressed in millimeters. In the group with a history of peptic ulcer, the roentgenometrical gastric fold was significantly thicker than that in the group without it. The fold width was significantly greater in the elderly group (55-64 years old) than in the young (35-44 years old) and middle-aged (45 54 years old) groups. The fold width tended to increase with age in persons with peptic ulcer history, but not in those without it. When the subjects were divided into three groups by gastric fold width, the incidence of peptic ulcer was significantly higher in the upper third group compared with the lower third group. This relationship between the gastric fold width and the incidence of peptic ulcer tends to become stronger with aging. In the heavy smoker group (> or = 10 of cigarettes per day), the gastric fold width was significantly thicker than that of non-smokers or those who smoked less. This relationship also tended to grow stronger with aging. The mean incidence of peptic ulcers was significantly higher among the heavy smokers. However, daily alcohol drinkers did not show any significant difference in gastric fold width from the other subjects. The gastric fold seems to be thicker in persons with peptic ulcer history, and the incidence of peptic ulcer is higher in persons with thicker gastric fold. These relationships tend to grow stronger with aging.a PMID- 10857048 TI - History of the Public Health Institute of Semmelweis Medical University, Budapest. AB - The science of public health of the XVIIIth century named politia medica together with medicina forensis became an independent obligatory subject in 1793 at the Medical Faculty of the Hungarian Royal University of Science. The independent Public Health Institute of the Medical Faculty was established in 1874. The first professor of public health was Jozsef Fodor who attained international reputation during his professorship. He organized training for school physicians and health teachers first in Europe and he organized courses for medical officers and for military doctors. He held courses for law-, engineer- and architect-students. He promoted all fields of the public health. His research on the bactericide effect of serum places him among the founders of immunology. Fodor's successors at the Chair of Public Health were Leo Liebermann whose research activities included physico-chemistry, biochemistry, microbiology and social hygiene; Gusztav Rigler who focused on the epidemiology of communicable diseases, on the health effects of spa treatment and mineral waters. The next famous professor was Gyula Daranyi. His scientific field was public health bacteriology and public health chemistry. They were followed by Jozsef Melly and Laszlo Dabis (Scheff). After the Second World War fundamental changes took place in the life of the university. The Faculty of Medicine was separated from the University of Science on February 1, 1951 and became an independent university under the control of the Ministry of Health. In 1953 the Institute of Public Health was cut into two separate institutes: Institute of Public Health and Institute for the Organization of Health Service. The Institute of Public Health was transformed to Institute of Public Health and Epidemiology in 1973. The Institute for the Organization of Health Service was transformed into Institute of Social Medicine and History of Medicine in 1985 and later into Institute of History of Medicine and Social Medicine in 1991. The Institute of Public Health and Epidemiology and The Institute of History of Medicine and Social Medicine were reunited as Institute of Public Health in 1997. The Institute teaches public health to medical, dental and pharmacy students in Hungarian, in English and in German. PMID- 10857049 TI - Health risk factors and mortality in Pecs City, Hungary in the 1990s. AB - In the period of 1990-1994 an increase of all causes mortality for 35-74 years old males was observed both in Pecs and in all Hungary. From 1994 to 1997 the mortality decreased. Similar changes, but of smaller dimension, were observed in the female population. The increasing mortality of the early 1990s is attributed primarily to the extra psycho-social stress of this period. The data of the population survey at Pecs in 1995-96 were compared to the data of earlier surveys. The mean blood total cholesterol levels and the prevalence of smoking decreased from 1990 to 1996. The prevalence of hypertension and male obesity increased. Physical inactivity, unhealthy diet and lack of improvement of diet still represent significant health problems. High prevalence of increased gamma glutamyl transferase indicate high prevalence of excess alcohol consumption. The risk factor profile of 18-25 year old males is very unfavourable. Smoking prevalence in females aged 26-35 years exceeds that of males of the same age group. Preventive efforts should be focused to young males and females. PMID- 10857050 TI - Cyclicity in incidence variations of meningococcal infections in Bulgaria is similar to that of solar activity. AB - This is a retrospective study on meningococcal meningitis (MM) in Bulgaria that has, for the first time, reported results on non-linear temporal patterns of incidence and its variations. Methods of descriptive statistics, linear and non linear modelling as well as periodogram regression analysis have been applied. A non-linear decreasing trend in crude incidence rates per 100 persons over the years 1940-1990 has been described (p < 0.0001) and cyclic variations revealed (periods T = 8.00, 18.75, 24.75 and 33.50 years, p < 0.05-0.01). Above cycles have been detected after the reciprocal trend has been removed (y = 1.04 + 15.78/t). A similar cyclicity (periods T = 8.25 and 27.5 years) in the variations of solar activity (sunspot number Rz) over the same time interval of 51 years has been established after the main cycle of 10.5-13 years has been removed by a two step procedure. The results from this study have added to our previous findings on cyclic variations in mortality and lethality from meningococcal infections in Bulgaria (1, 2). Above similarity is also in accordance with earlier conclusions on relations of solar activity cycles with epidemics of cerebrospinal meningitis in New York and USA over the years 1800-1935 (3, 4). PMID- 10857051 TI - The concentration levels of Cd, Pb, Hg, Cu, Zn and Se in blood of the population in the Czech Republic. AB - Knowledge of normal levels of concentrations of trace elements (Cd, Pb, Hg, Cu, Zn, and Se) in the population serves, among others, in design of regulations concerning health protection, determination of exposure limits and prevention of diseases caused by deficiency of trace elements. Concentrations of the named elements in whole blood of the Czech population were determined by means of atomic absorption spectrometry. The blood was collected during 1996-1998 from 1,216 blood donors (896 males and 320 females, average age 33 years) and 758 children (397 boys and 361 girls, average age 9.9 years). Mineralisation in a microwave digestion system was used in sample preparation. The accuracy of results was checked by means of the Control material Seronorm Whole Blood 404107 and Seronorm Serum 704121, Nycomed. Values of concentrations of the trace elements in blood found for adult (medians) were 0.7 microgram Cd.l-1, 800 micrograms Cu.l-1, 0.78 microgram Hg.l-1, 41 micrograms Pb.l-1, 76 micrograms Se.l-1, and 5,800 micrograms Zn.l-1, respectively. Statistically significant differences between men and women have been found in the concentrations of Cu, Hg, Pb, and Zn. In the juvenile population following medians of concentrations have been found: 0.15 microgram Cd.l-1, 1,047 micrograms Cu.l-1, 0.46 microgram Hg.l-1, 34 micrograms Pb.l-1, 69 micrograms Se.l-1, and 8,180 micrograms Zn.l-1. Statistically significant differences between boys and girls were found only in Pb and Zn concentrations. Concentrations of the studied elements correspond to the published values concerning population not exposed professionally. PMID- 10857052 TI - Daily mortality and air pollution in northern Bohemia: different effects for men and women. AB - The association between short term changes in ambient air concentrations of TSP and SO2 and day to day fluctuations in mortality was analyzed in the highly polluted Northern Bohemia region inhabited by approximately 630,000 people. A logistic regression model was adjusted for long term trends, seasonal cycles, influenza epidemics and weather parameters. The pollution and mortality data were available from 1982 to 1994. When the association was evaluated regardless of gender and age, 100 micrograms/m3 increase of TSP, but not SO2, was associated with a 3.4% increase of daily mortality lagged by 2 days. Evident differences in this association have been found between men and women. A significant increase in daily total and CVD mortality was observed in men below 65 while in women of the same age we found no association or even significant decrease in daily mortality. For the population over 65 the differences between genders were again apparent. The mortality in women increased significantly while in men significant decrease was demonstrated. CONCLUSIONS: The evaluation with respect to gender might contribute to identifying susceptible subgroups. PMID- 10857053 TI - An overview of European Union tobacco control legislation. AB - The European Union (EU) has been active in tobacco control policy since 1985 when the Milan Council announced its intention to establish a Europe Against Cancer (EAC) Programme, although it had previously adopted several Directives on aspects of tobacco taxation prior to this date. Shortly after the establishment of the EAC first action plan the European Commission presented its first legislative proposals on tobacco control. Three of these proposals on labelling and maximum tar yields became Directives by 1992. The fourth on tobacco advertising finally became law in 1998 and is currently being transposed into national law in the 15 EU Member States. In 1996 the Commission published a Communication on the future of EU tobacco control and in 1999 at the 2nd European Conference on Tobacco or Health the Social Affairs Commissioner announced his intention to bring forward further legislative proposals to amend an consolidate existing EU legislation in this sector. This article is intended to present an overview of EU tobacco control legislation from 1970 until 1998 and to look at future options post year 2000. PMID- 10857054 TI - The ascent of parity-violation: exochirality in the solar system and beyond AB - We review recent developments stemming from Stephen Mason's work on the origin of homochirality, focusing in particular on the parity-violating energy difference (PVED) between enantiomers. We summarize results of calculations of the PVED- both our own and those of other groups--which show that the natural enantiomers are indeed favoured by the weak force in most cases. The PVED has become important not only to explain the selection of the L-amino acids in the origin of life, but also as a "molecular footprint" of fundamental physics, leading to proposals to derive values of the Weinberg angle from future spectroscopic measurements of the PVED. The new field of exochirality--chirality outside the Earth--is now taking off, with reports of excesses of L-amino acids in meteorites, and proposals to look for homochirality as a signature of life on other planets and even in other solar systems. If it was indeed the PVED that determined life's handedness, we would expect to find L-amino acids rather than D everywhere in the universe. PMID- 10857055 TI - Stereoisomerism in bridged octahedral dimeric complexes with tetradentate ligands. Diastereospecific formation of syn-delta delta-(cis-beta-L)TiO2 (L chiral salen type ligands) AB - The first two examples of the successful application of chiral salen ligands for diastereoselective syntheses of dinuclear Ti(IV), (L1Ti)2O2 and (PdI2L2Ti)2O2 species with defined chirality and geometry at the metal centre are presented. The structure of the complexes was established by X-ray structural analysis. Both complexes have similar structural features: (1) chiral salen ligands have cis beta configuration around Ti centres, (2) (R,R) chirality of ligands induces delta helicity of ligand, and (3) syn arrangement of ligand donor atoms in relation to the TiO2Ti plane. Stereoselective effects of the dimers are discussed engaging available literature data. A model has been proposed to rationalise high syn selectivity in the formation of dimers. PMID- 10857056 TI - Enantiomeric synthesis of (3R)-3-amino-5-methyl-2-oxo-hexan-1-ol hydrochloride from cyclohexylideneglyceraldehyde AB - The first enantioselective synthesis of the title compound, (3R)-3-amino-5-methyl 2-oxo-hexan-1-ol hydrochloride (9a), an inhibitor of metalloprotein aminopeptidases has been developed from an enantiomerically pure 3-alkylglycerol. The required 3-alkylglycerol was, in turn, prepared by simple Grignard addition to cyclohexylideneglyceraldehyde 1 followed by separation of the epimeric product carbinols by normal column chromatography. The other attractive features of the synthesis was the use of inexpensive reagents and operationally simple synthetic protocols. PMID- 10857057 TI - Chromatographic resolution and elution order of alkyl aryl and aryl benzyl sulfoxides on cellulose-based chiral stationary phases AB - Several alkyl aryl and aryl benzyl sulfoxides have been prepared in optically active form via enantioselective Ti-catalyzed oxidation of the corresponding sulfides. The absolute configuration was assigned on the basis of optical rotation while in the case of some new sulfoxides it was determined by the analysis of their circular dichroism spectra. The alkyl aryl sulfoxides have been efficiently resolved by CHIRALCEL OB chiral stationary phase (CSP) while the aryl benzyl sulfoxides were better separated on CHIRALCEL OJ CSP. In both cases the S enantiomer was always eluted first. This finding can then allow to determine the absolute configuration of alkyl aryl and aryl benzyl sulfoxides on the basis of their elution order on these CSPs. PMID- 10857058 TI - Asymmetric redox reactions in human liver stereoselective oxidation of optically active dihydrohaloperidols, dihydrobromoperidols and stereospecific reduction of haloperidol and bromoperidol. AB - The stereoselective oxidation of dihydrohaloperidols (3a, b) and dihydrobromoperidols (4a, b), which are the main metabolites of haloperidol (1) and bromoperidol (2) in humans, respectively, were pharmacokinetically investigated using human liver microsomes and human cytochrome P450(CYP) isoenzymes expressed in the human cell line. The oxidation rates of the (R) isomers (3a and 4a) in the human liver microsomes were faster than those of the (S)-isomers (3b and 4b), and the R/S enantiomeric ratios of 3 and 4 for intrinsic clearance (Vmax/Km) were 1.40 and 3.10, respectively, showing that stereoselective oxidation occurred in human liver. Concerning the involvement of the CYP isoenzymes in this oxidative pathway, the (R)-isomers (3a and 4a) were catalyzed by both CYP3A4 and CYP2D6, however, the (S)-isomers (3b and 4b) were catalyzed only by CYP3A4. PMID- 10857059 TI - Synthesis, circular dichroism, and absolute stereochemistry of a Fecht acid analog and related compounds AB - 2,6-Dimethylspiro[3.3]heptane-2,6-dicarboxylic acid (2), an analog of the Fecht acid (1), was enantioresolved by the method of (1S,2R,4R)-(-)-2,10-camphorsultam yielding enantiopure acid (+)-2, the S absolute configuration of which was unambiguously determined by X-ray crystallography of camphorsultam amide derivative (R)-(-)-12b and chemical correlation. To determine the absolute configuration of chiral spiro[3.3]heptane compounds by the circular dichroism (CD) exciton chirality method, acid (+)-2 was converted to (+)-2,6 bis(phenylacetylenyl)-2,6-dimethylspiro[3.3]heptane (3) and (+)-2,6-bis(4 methoxyphenylacetylenyl)-2,6-dimethylspiro[3.3]heptane (4). The CD spectra of (+) 3 and (+)-4 exhibit intense exciton split Cotton effects of positive chirality, from which their S absolute configurations were confirmed. PMID- 10857060 TI - [Proximal amputation injuries of the thumb including the radial half of the hand. Case reports]. AB - The primary treatment of proximal amputations of the thumb and radial half of the hand is of particular importance, if the primary functions of a basic hand are to be restored. It is of utmost importance, that such treatment be carried out by a highly experienced reconstructive surgeon, since the transplantation or transposition of blood vessels, nerves, tendons and skin and soft-tissue flaps may be necessary. Five case reports illustrate the fundamental principles of primary and secondary reconstruction of the hand. PMID- 10857061 TI - [Bone growth after finger replantation in childhood]. AB - Following replantation in childhood, growth disorders of the affected epiphysis can influence function and aesthetic appearance of the hand. In a retrospective study long-term results of replantation in the upper limb in childhood were analysed with respect to factors influencing further growth. 22 patients with 29 replanted fingers were reviewed clinically and radiologically after an average interval of ten years. An average bone growth of 93% compared to the contralateral non-injured side was found. With the epiphysis affected, bone growth was reduced to 86% of the contralateral side. Analysis of single phalanges showed a growth rate of 71 to 100% in phalanges distal to the amputation and also in phalanges initially severed. Potential factors influencing bone growth were assessed separately. Best results were achieved in straight injuries without epiphyseal affection. An influence of anoxia time or number of anastomosed vessels could not be found. Even though replantation in childhood affects growing bone, almost normal bone growth can be expected afterwards. PMID- 10857062 TI - [Fractures near the base of the first metacarpal bone--clinical outcome of 21 patients]. AB - Fractures of the base of the first metacarpal are particularly common. The present examination was carried out in order to find out a correlation between the clinical outcome and the type of the fracture, the quality of reduction, the surgical procedure, and the extent of osteoarthrosis. From February 1992 to August 1997, 24 patients with Bennett fracture-dislocation, Rolando fracture und extraarticular fracture--eight patients in each group--were treated operatively. After a median interval of 33 months, 21 patients were reviewed. The evaluation was done using a new score with subjective (pain and utility) and objective (strength and range of motion) parameters. The "DASH score" was also used. Excellent results were found in the intra- and the extraarticular fractures. The extent of palmar abduction was 98% and 86% of the contralateral side in Bennett and Rolando fractures, 90% in extraarticular fractures. Key pinch was 93%, 77%, and 90% of the contralateral side (Bennett, Rolando and extraarticular fractures). This means 97, 85, and 87 points in the new score and 100, 93, and 85 points in the DASH score. The results after intraarticular fractures did not correlate with the type of treatment. Osteoarthrosis was found to correlate with the quality of reduction of the fracture, but remained asymptomatic. Intraarticular fractures have no worse prognosis. Nevertheless, exact reduction, either by the open or closed method, should be the aim of treatment. PMID- 10857063 TI - [Osteosynthesis with resorbable hemi-cerclage in metacarpal fractures]. AB - The use and results of biodegradable hemicerclages for metacarpal fracture fixation were reviewed retrospectively. A total of 92 metacarpal fractures in 78 patients were treated with polyglycolic or polydioxanon sutures. Study parameters included time for bony union, duration of immobilisation, total active range of motion, and complications. The hemicerclage achieved rigid fracture fixation and permitted early mobilisation exercises without jeopardizing bony union. Immobilization of metacarpals was performed for a median of 3.4 (1.5 to 6) weeks. There were no complications of wound healing. Adequate bony union was achieved after a median of 4.5 weeks (3.5 to 7 weeks). In one case, premature loading of the fracture led to displacement and delayed union. At the end of treatment (6.1 [4 to 7.5] weeks), total active range of motion was 98 (85 to 100)%. Ideal indications are oblique or torsion fractures of the metacarpals. In these cases, immobilisation up to wound healing is sufficient. PMID- 10857064 TI - [Minimally invasive management of metacarpal I fractures with a mini-fixateur]. AB - Displaced perarticular fractures of the first metacarpal are mainly treated operatively due to the importance of the first ray for hand function. Open reduction and internal fixation as well as minimal-invasive techniques using minifixator systems may be employed. We use a minifixator, which allows pin positioning even in small perarticular fragments eliminating the necessity of bridging joints. The anatomical basis, operative technique, and data of seven patients treated according to this method and results are presented. Range of motion according to the neutral-zero-method, tendon gliding, five-second-holding power, and force grip were equal compared to the uninjured contralateral side. The use of a minifixator to stabilize perarticular fractures of the first metacarpal includes the advantages of a minimal invasive procedure, respects the advantages of conservative fracture treatment, and permits secure fracture fixation after reduction. PMID- 10857065 TI - [Osseous avulsion injury of the extensor carpi radialis brevis tendon from the base of the 3rd metacarpal bone]. AB - An uncommon case of a closed avulsion fracture to the base of the third metacarpal bone by the extensor carpi radialis brevis tendon is described. The dislocation of the small bony fragment was reduced by open reposition and internal fixation with two Leibinger mini-screws. PMID- 10857066 TI - [Does low intensity, pulsed ultrasound speed healing of scaphoid fractures?]. AB - Since pulsed low-intensity ultrasound (frequency: 1.5 MHz, pulsed by 1 kHz, signal burst width: 200 microseconds, intensity: 30 mW/cm2) has been proven to stimulate fracture healing both clinically and experimentally, our question was whether this therapy also accelerates healing of fresh stable scaphoid fractures. Addressing this question, we did the following prospective randomized clinical trial. Regarding the results of former clinical fresh fracture studies by Heckman and Kristiansen, we postulated that low intensity ultrasound accelerates healing by about 30%. Based on this thesis, we calculated that 30 patients divided into two groups would be necessary to show significant differences between the standard treatment (treated by casting) and an adjunctive ultrasound treatment (treated by casting and additional daily 20 minutes ultrasound treatment) if present. Diagnosis and healing was assessed by CT scans every two weeks. CT's were analyzed by two independent radiologists and one hand surgeon. Furthermore, areas of cancellous bone bridging in correlation to the diameter of the scaphoid was measured in each CT scan. The results showed ultrasounded fractures healing in 43.2 +/- 10.9 days versus 62 +/- 19.2 days in the control group (p < 0.01). Trabecular bridging six weeks after injury showed 81.2% +/- 10.4% healed in the ultrasound-stimulated fractures versus 54.6% +/- 29% in the control (p < 0.05). Our study results confirm those of Heckman and Kristiansen and show a similar acceleration of bone healing. Low intensity ultrasound is successful in accelerating the healing of fresh scaphoid fractures. PMID- 10857067 TI - [Aseptic necrosis of the capitate: a rare cause for wrist pain. Case report and review of the literature]. AB - Avascular necrosis of the capitate is a rare condition. Most cases involve direct or indirect trauma with or without fracture. Etiological conditions, symptoms, diagnosis, and treatment options are presented in a case report and a review of the literature. PMID- 10857068 TI - [Treatment of a radio-ulnar synostosis by resection and interposition of a septofascial flap--a case report]. AB - Posttraumatic bone formation between radius and ulna can limit forearm rotation considerably. Recurrence after resection of synostoses is likely to develop if the bony surfaces are not covered by gliding soft tissue that is well vascularised. The interposition of a fascial forearm flap pedicled on the septal vessels of the posterior interosseous artery is suitable for this particular purpose. We report on the case of a spontaneous radioulnar synostosis by cartilaginous exostoses and its treatment by resection and interposition of a vascularised fascial flap. Forearm rotation was restored to normal. Until two years postoperatively there has been no recurrence of the synostosis. PMID- 10857069 TI - [Prosthetic replacement of the first carpometacarpal joint with a cemented ball and socket prosthesis (de la Caffiniere)]. AB - We analysed the fate of 43 de la Caffiniere prostheses implanted for arthrosis of the first carpometacarpal joint after a follow-up of 63 months (range 45 to 81). Ten prostheses failed. Further eight patients were lost for follow up. In the remaining 25 patients the rate of tilting of the cups (92%) was extremely high and the survival rate disappointing: 66% at 68 months. The prostheses were implanted in cancellous bone and the ball and socket reduces the moving axes of the normal joint to one center of rotation. These factors might explain the higher rate of loosening of the cups (28%), compared to 15% for the stems. In contrast, the clinical results were comparable with other operative procedures. The advantage is the brief postoperative rehabilitation period. Accordingly, this prosthesis can not be universally recommended and should be reserved for elderly patients, who must not undertake strenuous work. PMID- 10857070 TI - [Soft tissue tumors with difficult to distinguish boundaries. From myxoma to myxofibrosarcoma: a case report]. AB - Myxofibrosarcoma is a slow-growing subcutaneous tumor found in the older patient. Because of its deceptive macroscopic and histologic appearance, it is often misinterpreted as a benign lesion. We report a case of this tumor in a young woman recurring five times before the final diagnosis was made. Since recurrence may lead to tumor progression and increases the risk of metastasis, accurate diagnosis and radical removal of the lesion are extremely important. Eight benign and malignant myxoid tumors that have to be considered as differential diagnosis are reviewed. PMID- 10857071 TI - [Assessment of hearing impairment in workers exposed to mixtures of organic solvents in the paint and lacquer industry]. AB - Clinical and experimental studies indicate a possible harmful effect of chemicals, especially organic solvents, on the hearing system. In combined exposure to noise and solvents, very common in industry, it is most likely that a synergetic action of these factors enhances the traumatising effect of exposure to noise. The aim of this study was to assess the incidence and the risk of hearing impairment in 117 paint and lacquer factory workers exposed to a mixture of organic solvents. An analysis of organic solvent mixtures reveals that xylene and ethyl acetate are their major components whose concentrations depend on individual workposts. The control group consisted of 76 workers exposed to noise exceeding, Threshold Limit Value and 125 healthy subjects exposed neither to noise nor to solvents in their occupational setting. Pure tone audiometry revealed the highest hearing thresholds in workers exposed to solvents, lower thresholds in those exposed to noise, and the lowest ones in the non-exposed individuals. Hearing loss was found in 30% of workers exposed to organic solvents, in 20% of noise-exposed subjects, and in only 6% of non-exposed subjects. The comparison of relative risk values also indicated significantly enhanced probability of hearing impairment in workers of the paint and lacquer factory (9.6; 3.2-25.6), which is even more strongly pronounced than in the group of subjects exposed to noise (4.2; 1.2-13.2). An analysis of hearing impairment risk in particular frequencies suggests that organic solvents may damage the inner ear in much greater extent than noise. The results of the study show that exposure to organic solvents may create a significant risk of hearing impairment. Therefore, further steps should be taken to include the exposed population into effective preventive programmes. PMID- 10857072 TI - [Evaluation of occupational exposure of copper welders]. AB - Exposure of MMA/Plasma/Cu welders to fumes and soluble and insoluble Cu was assessed. Mean weighted concentrations of dusts/fumes and copper compounds at the welders' breathing zone were: 3.1 mg/m3 (0.7-7.6 mg/m3) for fumes; 0.08 mg/m3 (0.02-0.17 mg/m3) for soluble Cu; and 0.39 mg/m3 (0.08-0.76 mg/m3) for insoluble Cu. Correlation was found between the concentration of Cu compounds and fume/dust concentrations (r = 0.55 MMA/Plasma/Cu; r = 0.93 MMA/Cu). The mean percentage of copper in fumes/dusts was 17 (9.5-28.5%). Our results show that copper welding may pose a hazard to the welders' health. The index of combined exposure to determined agents was 1.2 (0.3-2.6), including soluble Cu (0.8) and insoluble Cu (0.4). The proportion of respirable fraction of fumes and their components varied between 40-80%. The background concentrations of the components (about 6 times lower than those in the welders' breathing zone) did not exceed the MAC value. PMID- 10857073 TI - [Assessment of carcinogenic effect of aluminosilicate ceramic fibers produced in Poland. Animal experiments]. AB - The effect of aluminosilicate ceramic fibres produced in Poland was assessed. The experiment was performed on two animal species: Wistar rats and BALB/C mice. The animals were administered intraperitoneally the studied fibres and krokidolit UICC--in doses of 25 and 5 mg and left for survival. All dead and sacrificed animals were examined histopathologically. Carcinogenic properties of ceramic aluminosilicate fibres were found to be rather weak. Only in 1 (2.5%) of 39 rats under study benign mesothelioma of tunica vagiualis testis was diagnosed. Peritoneal mesothelioma was found in none of 50 mice studied. For comparison the effect of krokidolit UICC was assessed. Krokidolit UICC is characterised by strong carcinogenic properties. It induced peritoneal mesothelioma in 43 mice (44.2%) and in 29 (80.5%) of 36 rats under study. PMID- 10857074 TI - [Carcinogenic effects of diesel emission: an epidemiological review]. AB - The results of recent epidemiological studies and meta-analysis relating to carcinogenic effects of diesel emissions in exposed populations were reviewed. Statistical, but still not causal association between risk of lung cancer and occupational exposure to diesel emissions was found in a great number of studies under review. Long-term exposure to diesel exhausts (> 20 years) increases by 30 40% lung cancer risk in workers of the transport industry: truck drivers, diesel engine mechanics, locomotive engineers and brakesmen. The results are inconsistent among heavy equipment operators, bus drivers and miners. Relative risk of lung cancer among workers occupationally exposed to diesel emission may be comparable with that of environmental tobacco smoke. Further research is also needed in the area of carcinogenic mechanisms, and biomarkers of exposure should be developed and validated before reliable quantitative estimates of risk of harmful effects to the human health in occupational setting are made. PMID- 10857075 TI - [Baker's asthma: what remains still unknown about it?]. AB - In many countries baker's respiratory allergy is reported as one of the most common occupational diseases. The paper presents epidemiological data on and risk factors of sensitisation to bakery allergens. Different flour allergens, especially those of wheat and soya flour, and (alpha-amylase are also described, and problems occurring most frequently in diagnosing occupational allergy to flour discussed. It is stressed that skin prick tests should be included into the battery of preliminary examinations before starting vocational training. PMID- 10857076 TI - [The role of melatonin in the molecular mechanism of weak, static and extremely low frequency (50 Hz) magnetic fields (ELF)]. AB - Melatonin is a neurohormone produced by the pineal gland. It has been recently found that it is also an antioxidant and a free radical scavenger. Melatonin was documented to be a direct trap of hydroxyl and peroxyl radicals. Therefore, this hormone could protect cells, tissues and organs against oxidative (free radicals) damage (DNA, protein, lipids). It has been suggested that noxious effects of ELF exposure (cancer or immunological disturbances) could be due to increased the concentration of free radicals induced by magnetic field. This is also leading to a hypothesis that melatonin suppression (by electromagnetic fields) in humans may increase the probability of mutagenic and carcinogenic risks. The future experiments, in vitro and in vivo, should provide an answer to the question on what is the real role of melatonin in the molecular (free radicals) mechanisms of weak magnetic fields. PMID- 10857077 TI - [The impact of lead ions on metabolism of erythrocytes]. AB - Lead contamination of the environment constitutes one of the major ecological problems. The determination of blood lead concentration is essential for evaluating lead exposure. Lead ions result in changes in the activity of numerous enzymes, disturb metabolic pathways of cells (inhibition of glycolysis), and inhibit synthesis of haem and globin. However, the activity of lead via pathological formation of free radicals is thought to be one of the most important molecular mechanisms of lead toxicity. Based on available literature, the issues concerning the effect of lead on metabolism of erythrocytes is discussed in this paper. PMID- 10857078 TI - [Training in occupational medicine: accreditation issues]. AB - Accreditation of training is an essential element of establishing modern and efficient system of education in the area of occupational medicine that satisfies up-to-date standards. The authors present basic aims and objectives of the accreditation process, as well as problems faced by specialists trying to achieve common understanding of the conception itself which will lead to setting uniform criteria for training in occupational medicine in Europe. PMID- 10857079 TI - [Positive and negative effects of fluorine in the human organism. The source of fluorine in the environment]. AB - In Poland and in other countries around the world, a great number of epidemiological studies have provided evidence that fluorine contributes to decreasing the intensity of dental decay. Nevertheless, the negative effect of its overdose should not be ignored. At present, no data indicating the amount of fluorine uptake is available. A wide diversity of opinions on tolerable limits of fluorine, and its predicted dosage known as optimal, suggests a need to monitor constantly this element throughout the whole country, and not only in selected regions. PMID- 10857080 TI - An overview of affinity chromatography. PMID- 10857081 TI - Weak affinity chromatography. PMID- 10857082 TI - Fluidized-bed receptor-affinity chromatography. PMID- 10857083 TI - Site-specific immobilization of antibodies to protein G-derivatized solid supports. PMID- 10857084 TI - Affinity purification of monoclonal antibodies. PMID- 10857085 TI - Protein A mimetic (PAM) affinity chromatography. Immunoglobulins purification. PMID- 10857086 TI - Periodate oxidation of antibodies for site-selective immobilization in immunoaffinity chromatography. PMID- 10857087 TI - Mini-antibody affinity chromatography of lysozyme. PMID- 10857088 TI - Nitrocellulose-based immunoaffinity chromatography. PMID- 10857089 TI - Immunoaffinity chromatography. PMID- 10857090 TI - Affinity partitioning of proteins. PMID- 10857091 TI - Boronate affinity chromatography. PMID- 10857092 TI - Dye-ligand affinity chromatography for protein separation and purification. PMID- 10857093 TI - DNA affinity chromatography. PMID- 10857094 TI - Affinity chromatography of pyrogens. PMID- 10857095 TI - Western cross blot. PMID- 10857096 TI - Affinity perfusion chromatography. PMID- 10857097 TI - Phage display technology. Affinity selection by biopanning. PMID- 10857098 TI - Phage display technology. Identification of peptides as model ligands for affinity chromatography. PMID- 10857099 TI - The complement system: an overview. PMID- 10857100 TI - Purification of complement components, regulators, and receptors by classical methods. PMID- 10857101 TI - Immunoaffinity methods for purification of complement components and regulators. PMID- 10857102 TI - Measurement of complement hemolytic activity, generation of complement-depleted sera, and production of hemolytic intermediates. PMID- 10857103 TI - Measurement of complement lysis of nucleated cells. PMID- 10857104 TI - Functional assays for complement regulators. PMID- 10857105 TI - Immunochemical measurement of complement components and activation products. PMID- 10857106 TI - Complement deposition in tissues. PMID- 10857107 TI - Complement regulators and receptors in tissues. PMID- 10857108 TI - Measurement of C3 fragment deposition on cells. PMID- 10857109 TI - Screening for complement deficiency. PMID- 10857110 TI - C1-inhibitor: antigenic and functional analysis. PMID- 10857111 TI - Autoantibodies to complement components. PMID- 10857112 TI - Allotyping of complement components. PMID- 10857113 TI - Complement and immune complexes. PMID- 10857114 TI - Knocking out complement genes. PMID- 10857115 TI - Inherited complement deficiencies in animals. AB - Following the initial description of natural C5 deficiency in inbred mice, a growing number of complement component deficiencies in animals have been described, caused by a variety of genetic defects. Studies on such animals have contributed greatly to an understanding of the specific roles of classical, alternative and terminal pathway activation and inhibition in many infectious and inflammatory diseases. Further investigations in the more recently described models, in combination with the use of genetically engineered knockout mice (with targeted disruption of individual complement components and inhibitors), should continue to provide a fertile source of information regarding the role of complement in such experimental situations. This information is likely to have significant therapeutic implications for human disease. PMID- 10857116 TI - Pathophysiology of abnormal uterine bleeding. AB - Abnormal uterine bleeding is a frequent patient complaint. Recognition of the severity of the problem, appropriate and timely evaluation, and treatment with good outcomes is the goal. The physician must determine which method of diagnosis he or she is most comfortable with, carefully consider the economic impact, and offer treatment that is best suited for each patient. With this practice patients will obtain maximum benefit from the newer treatments in development. PMID- 10857117 TI - Endometrial sampling techniques in the diagnosis of abnormal uterine bleeding. AB - There have been many advances in sampling of the endometrium. Ideas and technologies have evolved, increasing our ability to gather adequate specimens that provide reliable information about uterine cavity pathologies. No technique surpasses the sensitivity and specificity of hysteroscopy with directed biopsy. Owing to its superior diagnostic potential, hysteroscopy, even when performed in the office with narrow scopes (not significantly larger in diameter than the Pipelle catheter), leads to precise diagnosis and appropriate management of intrauterine pathologic conditions. For physicians who are untrained or lacking the equipment to perform diagnostic hysteroscopy with directed biopsy, simple in office endometrial sampling techniques with no visual control provide a means to obtain reasonably reliable samples with negligible patient discomfort. PMID- 10857118 TI - Radiographic imaging techniques for the diagnosis of abnormal uterine bleeding. AB - The introduction of SIS has been a significant advance in TVUS evaluation of the endometrial cavity in the 1990s. SIS provides an unparalleled, clear, enhanced view of the endomyometrial complex that cannot be obtained with TVUS alone. Focal and global endometrial pathology can be differentiated with SIS. Saline infusion improves the sensitivity for the detection of endometrial abnormalities. The continuing challenge for gynecologists is to provide patients with cost effective, minimally invasive evaluation and directed therapy for menstrual dysfunction. SIS targets patients needing biopsy, directs the surgical approach, and minimizes office diagnostic hysteroscopy--all with a quick office procedure. For patients, the benefits include minimal and brief discomfort and a better understanding of intrauterine pathology through viewing the ultrasound monitor. Patients also appreciate the ease of scheduling, the minimal time away from work, and that no escort is needed after the procedure. SIS provides an extension of the pelvic gynecologic examination. SIS is the most important imaging modality for evaluating endometrial pathology. Although there is no perfect test to evaluate the endometrium, overall, SIS is superior to other imaging and diagnostic procedures. It is less expensive than D&C or hysteroscopy. It is a safe, efficient, convenient, and well-tolerated procedure. In some instances, however, neither TVUS nor SIS is definitive in determining the location of fibroids or able to discern adenomyosis. In these instances, MR imaging triage is helpful. MR imaging is gaining widespread acceptance and, in many instances, is a cost-effective tool in the evaluation of abnormal uterine bleeding. It is noninvasive, differentiates uterine anatomy in response to exogenous hormones or the normal menstrual cycle, and reliably localizes pelvic pathology and size of lesions. When uterine conservation is desired in women with fibroids and TVUS or SIS is indeterminate in localizing depth of myometrial involvement of a fibroid, MR imaging should be considered as a part of the clinical algorithm. The precision of MR imaging localization of submucosal fibroids can obviate the need for hysterectomy and permit a skilled surgeon to hysteroscopically resect the fibroids. If the clinical examination is suspicious for adenomyosis and the US is nondiagnostic, the clinician should consider MR imaging strongly. When the results of the imaging study would influence surgical route and planning, MR imaging should be considered in the preoperative evaluation. PMID- 10857119 TI - Diagnostic hysteroscopy to evaluate the cause of abnormal uterine bleeding. AB - Diagnostic hysteroscopy has become an important and valuable tool for the gynecologist in the evaluation of many conditions previously evaluated with blind and inaccurate techniques. The safety, ease of proficiency, and ability to see and diagnose in an office setting have taken much of the guesswork out of office diagnosis. This modality brings the evaluation of many pathologic conditions, including the cause for abnormal uterine bleeding, infertility, and recurrent pregnancy loss, back into a relaxed office environment. PMID- 10857120 TI - Medical management of abnormal uterine bleeding. AB - Abnormal uterine bleeding occurs secondary to a wide variety of functional and structural abnormalities. Although there is clearly a place for surgery, medical therapy has enormous potential for most women, especially those with dysfunctional uterine bleeding. To provide women with appropriate options for therapy, the clinician must be prepared to distinguish abnormal bleeding that is associated with ovulation from that which is anovulatory and to use appropriate ancillary tests to identify structural and endocrinologic anomalies or lifestyle factors that may explain the bleeding. In undertaking such an investigation, it is important for the clinician to be able to distinguish lesions that may be asymptomatic and unrelated to the bleeding from those that truly are the source of the problem. With this information, a rationally determined set of medical and, if appropriate, surgical therapeutic options may be presented to the woman. Among these medical treatment options are a number of treatment options that have not seen widespread use in North America but are inexpensive, effective, and well tolerated. It is clear that medical therapy is not for everyone. Women deserve the opportunity to relieve their symptoms with nonsurgical options. PMID- 10857121 TI - Instrumentation and distention media for the hysteroscopic treatment of abnormal uterine bleeding. AB - New hysteroscopes and resectoscopes with continuous-flow designs have greatly facilitated diagnostic and therapeutic hysteroscopy. Saline is the ideal distending medium for hysteroscopic procedures in which mechanical or bipolar instruments are used; 5% mannitol may be the safest medium for resectoscopic surgery. Regardless of the medium chosen, careful fluid monitoring is essential. PMID- 10857122 TI - Energy systems for operative hysteroscopy. AB - The use of energy to perform intrauterine surgery can be broken down into three fundamental elements: delivery, transmission, and tissue strike. Thermal delivery devices, including monopolar or bipolar electrodes and quartz fibers, transmit energy either directly or indirectly to tissue. Transmission is unaffected, facilitated, or deterred by the distention medium that fills the intrauterine cavity. The final targeted-tissue effects represent the complex summation of energy type and concentration; treatment time; tissue constituents and hydration; and the convective currents created by tissue vascularity and circulating distention medium. Efficiency, safety, and efficacy are inextricably linked to the thoughtful orchestration of all of these elements. The responsibilities to know how and why are only stepping stones to attaining the best surgical result. We are reminded that surgery transcends technique only when experience and judgment are coupled with fundamental knowledge. Only then can science become art in the hands of the hysteroscopic surgeon. PMID- 10857123 TI - Hysteroscopic treatment of the patient with intracavitary pathology (myomectomy/polypectomy). AB - Hysteroscopy is a simple and effective method for treating intrauterine leiomyomas and polyps. These lesions often cause abnormal uterine bleeding and infertility. Using the techniques in this article, most lesions can be removed in an office or outpatient setting. PMID- 10857124 TI - Treatment of the patient without intracavitary pathology. Comparison of traditional hysteroscopic techniques for endometrial ablation. AB - Endometrial ablation has become a necessary and useful procedure for the management of abnormal uterine bleeding in women desiring uterine conservation. Current ablation techniques are safer and more effective than earlier methods. This article explains the steps to perform laser and resectoscopic endometrial ablation and provides suggestions for making these processes more effective. PMID- 10857125 TI - Intraoperative and early postoperative complications of operative hysteroscopy. AB - With preoperative evaluation, meticulous technique, and vigilance for impending problems, intraoperative and early postoperative complications of operative hysteroscopy are largely preventable. Fluid overload is the most common serious complication. The hysteroscopist must understand the significant differences between hypotonic, electrolyte-free distention media and isotonic, electrolyte containing media and their respective sequelae. As new operative tools become available, hypotonic and electrolyte-free distention media may become obsolete. The physiology and management of air embolism, the most grave intraoperative complication, are essential to the knowledge base of any active hysteroscopist. Mechanical accidents, anesthetic complications, laser and electrical injury, and infections can be reduced by knowledge and preparation. Technologic advances, ongoing research, and postgraduate training in hysteroscopic technique continue to expand the safe and beneficial applications of hysteroscopy into the next century. PMID- 10857126 TI - Late complications of operative hysteroscopy. AB - The late complications of operative hysteroscopy result from either persistent endometrium after ablation or myometrial damage during surgery. Residual endometrium can become neoplastic, cause pain, or support a pregnancy. Myometrial damage can produce catastrophic consequences during a later pregnancy. These long term problems place the impetus on the operating physician to select patients carefully, prepare the endometrium, and operate in such a way as to minimize the likelihood of residual endometrium and unnecessary myometrial damage. The value of operative hysteroscopy for infertility secondary to adhesions and uterine septa is unequivocal. Hysteroscopic surgery offers increased fertility rates while avoiding the risks of open surgery. For the treatment of abnormal uterine bleeding, endometrial ablation can be performed safely, and the long-term benefits are durable. As more operative hysteroscopy is performed, more delayed complications will arise. Easy-to-perform global ablation techniques and multifunctional operative hysteroscopes have enticed more gynecologists to test the waters of endometrial ablation and operative hysteroscopy. Although they empower the hysteroscopist to offer more advanced and more valuable minimally invasive options to patients, these tools simultaneously can tempt the surgeon to forego meticulous preoperative evaluation. Evidence exists that too often women undergo surgery without complete diagnostic assessment. In one study, 50% of women underwent hysterectomy without any diagnostic evaluation of the endometrium. Hysterectomy possesses a saving grace in that it provides cover for many missed diagnoses. Conservative, nonextirpative procedures offer no such life raft. Meticulous diagnostic assessment and preoperative consideration of risk factors for residual endometrium and future pregnancy remain the keys to minimizing late complications. PMID- 10857127 TI - New developments in operative hysteroscopy. AB - Although there is a plethora of new devices currently being investigated that can ablate the endometrial cavity successfully, it is most certain that a strong place remains for operative hysteroscopy for the therapy of benign uterine conditions. New instruments that continue to make operative hysteroscopy a safer and easier procedure to perform should continue in their development. PMID- 10857128 TI - Global endometrial ablation technologies. AB - The value of extirpative surgery for excessive uterine bleeding is questionable. A number of technologies have been developed that destroy the endometrial lining while preserving the uterus. This article compares and contrasts multiple modalities of global endometrial ablation technology. PMID- 10857129 TI - Embolotherapy for myoma-induced menorrhagia. AB - Uterine fibroids are a common cause of abnormal uterine bleeding. Uterine artery embolization has proven to be highly effective in controlling fibroid related menorrhagia and triggering tumor degeneration. PMID- 10857130 TI - Myomectomy. Comparison of open and laparoscopic techniques. AB - It appears that both laparoscopic and laparotomy approaches to myomectomy are viable surgical procedures. Both surgeries can be performed with minimal risk. Pregnancy rates after either procedure are high. The advantage of laparotomy over laparoscopy is that it is technically easier. On the other hand, laparoscopy enables quicker and easier recovery. Moreover, the risk of adhesion formation is reduced. If the surgeon can rise to the technical challenge, a laparoscopic approach to myomectomy should be the intended surgery. PMID- 10857131 TI - Myoma coagulation (myolysis). AB - Myoma coagulation or myolysis by way of the laparoscope or hysteroscope is a valuable addition to the armamentarium of treatments for a problem that remains pervasive among women: uterine leiomyomata. Likewise, surgical techniques include the use of the Nd:YAG laser as well as the bipolar needle. The addition of myolysis to earlier uterine-sparing endometrial ablation or resection markedly improves the success rate of these minimally invasive alternatives to hysterectomy. Myoma coagulation when combined with endometrial ablation among women with symptomatic fibroids and bleeding also reduces all subsequent surgery rates compared with endometrial ablation alone. The continued goal for therapy of fibroids and debilitating menorrhagia must take into consideration the needs and desires of the patient in terms of her lifestyle (e.g., days lost from work because of symptoms) and childbearing plans. Hysterectomy continues to be costly in billions of dollars spent annually as well as in the more fundamental terms of morbidity and mortality when compared with the less invasive alternatives of myomectomy, ablation, and myolysis. PMID- 10857132 TI - Total laparoscopic hysterectomy. AB - Alternatives to hysterectomy can provide excellent treatment outcomes for many women. In general, these alternatives are underused. For some women, however, alternative treatments fail and hysterectomy provides the best approach. The goal of future research should be to define better this group of women. For women who require hysterectomy, the laparoscopic approach affords the benefit of less postoperative discomfort, shorter hospital stay, and quicker recovery. The surgical techniques and instruments for laparoscopically directed hysterectomy are still in development. Few randomized, prospective studies that involve large numbers of patients have compared the risks and benefits of this approach with standard hysterectomy. In addition, data on the effectiveness of the operation, as performed by large numbers of gynecologists, are still not clear. Although the potential for real benefit exists, it is hoped that further study will clarify the place of laparoscopic-assisted hysterectomy in the nonsurgical and surgical treatment offered to patients. PMID- 10857133 TI - Laparoscopic supracervical hysterectomy. AB - Laparoscopic supracervical hysterectomy (LSH) provides an efficient, effective method of uterine extirpation for individuals with appropriate indications for the procedure. This article gives a rationale for the procedures and provides a step-by-step methodology for performing LSH. Data from existing literature are discussed, and suggestions for improving morbidity and mortality are presented. PMID- 10857134 TI - [Testicular cancer and male fertility]. AB - Disorders of fertility after treatment of testicular cancer constitute important sequelae. Almost one half of patients initially present decreased fertility. Chemotherapy, radiotherapy and surgery each exert their specific harmful effects. Although the desire for subsequent paternity is often satisfied spontaneously, some patients remain permanently sterile. Systematic semen storage, designed to palliate this sterility, when necessary, must be systematically part of the therapeutic contract. It is best performed before or immediately following castration. It also plays a psychological role, projecting the patient into his future cure. The limits of freezing have been considerably reduced with progress in vitro fertilization and ICSI. In the case of radiotherapy or chemotherapy, effective contraception must be recommended for 2 years after starting treatment. The long-term teratogenicity of the various treatments used is still very poorly known. PMID- 10857135 TI - [Urethral instability]. AB - Urethral instability is an abnormal behaviour of the urethra associated with sudden reduction of urethral pressures. This disease is rarely described in view of its nonspecific clinical features and the abnormalities only detected on continuous urethral recording. Alteration of the voiding reflex is the most likely aetiopathogenic basis, although other urethral and neurological diseases have been observed. The numerous treatment options reflect the limited knowledge concerning aetiological factors. PMID- 10857136 TI - [Malignant adrenal cortical neoplasms. Report of 22 cases]. AB - OBJECTIVES: Analysis of the treatment of a series of patients with adrenal cortical carcinoma. MATERIAL AND METHODS: This series consists of 22 patients (15 women and 7 men) with a mean age of 35 years (range: 23 to 58 years). 55% of patients had a non-secreting tumour; 45% of tumours were associated with excessive hormonal secretion. The tumour was located on the right side in 12 cases and on the left side in 11 cases (the tumour was bilateral in 1 case). Ultrasound and CT were sufficient for localization of the tumour. The patients were classified as stage I: 3 cases, stage II: 2 cases, stage III.: 8 cases, stage IV: 9 cases, according to MacFarlane's classification. RESULTS: Adrenalectomy was performed in 18 patients. Two patients died during the operation and another 8 died within the first four years. Eight patients are currently alive, including 7 with no signs of recurrence; one patient presented a large local recurrence 39 months after adrenalectomy. CONCLUSION: Surgery remains the only effective treatment at the present time. The diagnosis of adrenal cortical carcinoma must be made as early as possible. PMID- 10857137 TI - [Treatment of female stress urinary incontinence with cystocele by Gore Tex colpofixation and Burch operation]. AB - OBJECTIVE: We treat female urinary stress incontinence (USI) with cystocele with or without associated genital prolapse by a combination of sacral colpopexy and Burch anterior colposuspension. We evaluated the results of a consecutive series of 77 patients completing a telephone interview after a mean follow-up of 40.6 months (range: 15 to 74 months). MATERIAL AND METHODS: From January 1991 to December 1995, 77 patients (mean age: 56.7 years) underwent Gore-Tex sacral colpopexy and Burch colposuspension for USI. Levator ani myorrhaphy was also performed in 53 severe cases. Incontinence was severe in every case, and associated with stage > or = 2 cystocele in 93% of cases. Urodynamic assessment revealed detrusor instability (DI) in 17.3% of cases and sphincter insufficiency (SI) < or = 35 cm H2O in 11.9% of cases. The main complications were: 4 haemorrhages requiring transfusion of one unit, 2 wound abscesses, one wall haematoma, and one small bowel obstruction at the 4th month, treated surgically. RESULTS: Six patients were lost to follow-up (good early results) and 2 files could not be analysed with sufficient follow-up (patients died from other diseases). Good results were defined by at least two of the following criteria: patient satisfied, no incontinence and no need for protective pads. We obtained 59 successes (85.5%) and 10 failures (14.5%). The results remained stable over time with 88.8% of success after a follow-up of more than 60 months (20 patients). Nine of the 10 failures occurred during the first year. They were demonstrated by the survey, while the post-operative follow-up at one month had been satisfactory. No specific treatment was therefore proposed. The two main factors of failures were DI and severe SI. Three times more failures were observed when the preoperative closure pressure were less than 30 cm H2O. CONCLUSION: Sacral colpopexy combined with Burch operation is a reliable solution for repair of USI with marked cystocele. It ensures good initial results which persist at one year and in the long-term. PMID- 10857138 TI - [Periurethral injection of silicone microparticles in the treatment of sphincter deficiency urinary incontinence]. AB - OBJECTIVE: To assess the mid-term efficacy of periurethral injection of silicone micro-implants in women with urinary incontinence due to intrinsic sphincter deficiency. MATERIAL AND METHODS: Between July 1992 and March 1999, 25 women (mean age 65.17 +/- 13.5 years) with intrinsic sphincter deficiency underwent periurethral injection of silicone micro-implants. Urodynamic investigations were performed prior to and after surgery. The subjective degree of continence was also assessed for 3 years. RESULTS: Subjective success rate were 80%, 72%, 65%, 60%, and 60% at 6 weeks, 3 months, 6 months, 1 year and 3 years, respectively. Post-operative maximum urethral closure pressure was significantly increased (post-op 32.35 +/- 18.04 cm H2O versus pre-op 23.68 +/- 9.4 cm H2O; P < 0.01) No serious operative or postoperative complication occurred. CONCLUSION: In spite of time-dependent decrease in success rate periurethral silicone micro-implants injection remains an effective and safe procedure for women with urinary incontinence due to intrinsic sphincter deficiency. PMID- 10857139 TI - [Study of urethral anatomy and pelvic floor using MRI with surface and endorectal coil]. AB - PURPOSE: Although high resolution MRI can play a critical role in the evaluation of diseases affecting the female urethra, normative values have not been established. In this retrospective study, the normal values for female urethral dimensions and its supportive structures were measured and compared using a body coil (BC) and endorectal coil (ERC), and correlated with age and menopause. MATERIAL AND METHODS: BC and/or ERC images of the pelvis in 20 patients (ages 27 82) with confined cervical cancer (stage IB or less) were reviewed. None of the patients had a history of urinary symptoms, pelvic prolapse, pelvic radiation, or prior bladder or urethral surgery. Images evaluated included axial and/or sagittal T2 weighted SE images of the lower pelvis before and/or after endorectal coil placement. Several measurements including urethral and bladder dimensions were obtained independently by two radiologists and compared statistically. Calculated urethral volume was correlated with the patients' age and menopausal status. The impact of calculated bladder volume on urethral dimensions was evaluated. Additional measurements of contiguous supporting structures were also correlated with age. RESULTS: Inter-rater reliability showed a strong intra-class correlation (95% CI) for urethral dimensions. A statistically significant difference between raters was only noted for the right pubovesical ligament measurement. Inter-technique reliability was also strong (95% CI) except for the distal transverse urethral dimension. Bladder volume did not effect measurement of urethral dimensions (p > .39). Lastly, calculated urethral volume utilizing the ERC technique showed an inverse correlation with age (p < 0.05) and with the BC a correlation with menopausal status (p < 0.05). CONCLUSIONS: Measurement of urethral dimensions by either ERC or BC MRI is reliably reproducible by independent radiologists. There is no need for standardization of bladder volumes during urethral MRI. Normative values for all measured angles and dimensions are presented. There is evidence of correlation of urethral volume with age and menopause, though a larger study is warranted. PMID- 10857140 TI - [Features of prostatic cancer in French individuals of African-Caribbean origin]. AB - OBJECTIVES: To evaluate the clinical, laboratory and histological characteristics of prostate cancer at the time of diagnosis and after radical treatment in various ethnically different patient groups. PATIENTS AND METHODS: Prostatic biopsies were performed in 466 consecutive patients because of an abnormal digital rectal examination and/or isolated elevation of PSA (greater than 3 ng/ml). In this series, 40 patients were Black and 426 were Caucasian. The other aspect of the study concerns 320 patients undergoing radical prostatectomy for stage T1 T2 prostatic tumour (25 Black, 295 Caucasian). In the biopsied group, we analysed mean age, mean PSA, mean cancer length on biopsies and mean Gleason score. In the operated group, we studied preoperative characteristics, histological stage, resection margin status, laboratory progression (PSA greater than 0.05 ng/ml) and time to progression. RESULTS: At the time of diagnosis, the mean age was 61.4 years (48-73) for Blacks and 65.2 years (42-87) for Caucasians (p < 0.05). The median Gleason score was 7 in the two groups. The PSA was 13.4 (1.7-105) ng/ml versus 14.4 (0.4-600) ng/ml, respectively. The mean percentage of invaded tissue on biopsies was 24% versus 18.8% and the mean percentage of positive biopsies was 53% versus 39%, respectively. In the operated group, capsular effraction rates were 39% in Blacks and 48.1% in Caucasians. Positive resection margin rates were 21.7% versus 36.6%, respectively. The laboratory progression rate with a mean follow-up of 33 months (6-126) was identical in the 2 groups (42.1% versus 41.1%), but the time to progression was shorter for Blacks (9 months versus 12.3 months). CONCLUSIONS: In this patient series, Black patients had the same laboratory profile as Caucasian patients at the time of diagnosis. However, they were younger at the time of discovery of the disease, had more positive biopsies and more tumour-invaded tissue on biopsies, and Black patients undergoing radical prostatectomy developed laboratory recurrence more rapidly. PMID- 10857141 TI - [Power Doppler and 3D vascular sonography of intraprostatic blood supply: assessment criteria and value for the diagnostic and clinical staging of prostatic cancer]. AB - OBJECTIVES: To compare the value of Power Doppler Sonography (PDS) and B mode sonography in the diagnosis of prostate cancer and to assess the value of PDS to assess cell differentiation as a function of the degree of blood supply and to specify capsular effraction of the cancer. PATIENTS: 133 patients, divided into 2 groups, were investigated: one group consisted of 41 patients with no suspicion of cancer (15 control subjects and 26 patients with acute prostatitis) and a second group consisted of 92 patient with suspected cancer (PSA > 4 ng/ml). METHODS: Power Doppler sonography with 3D reconstruction was used to calculate a graduated blood supply index from 1 to 3 for nodules of the peripheral zone. Three types of blood supply (A: regular avascular capsule, B: irregular avascular capsule, C: vessels crossing the capsule) were described as a function of the presumed stage of cancer (A: intraprostatic, B: undetermined, C: extraprostatic). Comparison with histology was performed on randomized biopsies (92 cases) and radical prostatectomy specimens (21 cases). RESULTS: A cancer was diagnosed in 57 of the 92 patients (62%) with suspected cancer. The overall sensitivity of PDS in the initial diagnosis of prostatic cancer was 94.7%, and its specificity was 77.1% (versus 93% and 42.8% for sonography alone, respectively). For a prevalence between 0.4 and 0.9, the PPV ranged from 73.4% to 97.4% and the NPV ranged from 95.6% to 62% (p = 0.02). The correlation between tumour blood supply and Gleason score showed that 20/40 tumours (50%) with a Gleason score > or = 7 had a blood supply index of 3 versus 6/17 (35%) of tumours with a Gleason score < 7 (r = 0.283, p = 0.033). The 3 vascular types, A, B, C, were evaluated prospectively in the detection of capsular effraction. Capsular effraction was detected in one of the 8 cases of type A cancer and in 6 of the 8 cases (75%) of type C cancer. CONCLUSION: PDS improves the reliability of sonography in the diagnosis and staging of prostate cancer. There is a correlation between tumour blood supply and Gleason score. PMID- 10857142 TI - [Efficiency and tolerance of terazosine in ambulatory patients with benign prostatic hypertrophy: comparative randomized and double-blind trial versus alfuzosin. The MG Terazosine Group]. AB - OBJECTIVE: To compare, in general practice, the efficacy and safety of terazosin (5 mg per day in single dose) versus alfuzosin (7.5 mg per day in 3 doses) in patients with symptomatic benign prostatic hyperplasia (BPH) treated for 16 weeks. MATERIAL AND METHODS: Thirteen investigators included patients over the age of 50 years presenting with BPH with an International Prostate Symptom Score (IPSS) greater than 12 and a post-voiding residual volume less than 300 ml. After a one-week observation period, these patients were randomized to receive either terazosin or alfuzosin for 16 weeks (112 days) under double-blind conditions. The primary endpoint was the percentage reduction of the IPSS score at 3 weeks and 16 weeks; the secondary endpoint was the IPSS quality of life score. Safety was evaluated by recording adverse events and monitoring blood pressure. RESULTS: Seventy four patients were included: 39 in the terazosin group, 35 in the alfuzosin group. The 2 groups were not significantly different before treatment. Improvement of the IPSS score was similar in the 2 treatment groups (p = 0.97 at 3 weeks, and p = 0.29 at 16 weeks), as was the improvement of the quality of life score (p = 0.47 at 3 weeks and p = 0.71 at 16 weeks). Treatment was considered to be "effective or very effective" in 31 patients (86%) in the terazosin group, and in 28 patients (82%) in the alfuzosin group. The IPSS score was greater than or equal to 12 before treatment for all patients included in the study. Twenty-five patients had a score < 12 at 3 weeks versus 56 at 16 weeks (p = 0.0001). Seven patients had a quality of life score less than 2 before treatment, versus 38 at 3 weeks, and 56 at 16 weeks (p = 0.0001). No significant difference was observed between the 2 groups in terms of the number of adverse events reported, or the course of blood pressure and prostate specific antigen. No patient dropped out of the study because of treatment-related adverse events. Two deaths were observed in the terazosin group (2 patients aged 86 and 93 years), but any relation to treatment was excluded. CONCLUSION: During this study, terazosin appeared to be as effective and as well tolerated as alfuzosin. PMID- 10857143 TI - [Patterns of use of terazosine in current medical practice in ambulatory patients with obstructive and irritative obstructive disorders of urination]. AB - OBJECTIVES: To evaluate the efficacy and to describe the modalities of use of terazosin hydrochloride dihydrate, prescribed under conditions of routine clinical practice to a vast population of patients with benign prostatic hyperplasia (BPH). MATERIAL AND METHODS: 1,624 patients suffering from BPH and requiring medical treatment were included in this multicentre open clinical trial performed by 983 general practitioners. After a one-week titration phase, terazosin was administered for 4 weeks at the dosage of 5 mg per day as a single dose in the evening at bedtime. The efficacy of treatment was assessed by the variation of the IPSS score between inclusion and the end of treatment, the treatment response rate (at least 3 point reduction of the IPSS score) and the course of the quality of life score. The safety of treatment was evaluated by clinical interview and physical examination at each visit and by analysis of all adverse events occurring during the trial. RESULTS: A mean 45.8% improvement of the IPSS score (corresponding to a mean decrease of 8.81 points) was demonstrated between D0 and D35 (p < 0.001). The treatment response rate was 91.8%. A mean improvement of 2.11 points (p < 0.001) of the quality of life score was obtained. Complementary subgroup analysis (moderate dysuria n = 775 and severe BPH n = 702) showed that the efficacy of treatment was independent of the initial severity of the voiding disorders. The clinical safety of treatment was considered to be good in 92.5% of cases by the investigators. The incidence of adverse events attributable to treatment and related to a fall in blood pressure was 2.6%. Less than half (20/43, i.e. 1.25% of the population) of the patients experiencing this type of adverse event had to discontinue treatment. The ease of use of treatment was considered to be good in 85.2% of cases. CONCLUSION: Terazosin constitutes an effective symptomatic treatment for BPH when surgery is not indicated, whether the initial disorders are moderate or severe. The safety of treatment appears to be perfectly satisfactory, in a patient series representative of the general practice outpatient population concerned by BPH (age, state of health, concomitant treatments). The one-week progressive dose regimen allows cautious introduction of treatment and generally does not raise any problems of acceptability. PMID- 10857144 TI - [Repair of urethrocutaneous fistula with double skin flap. Report of 32 cases]. AB - OBJECTIVES: Urethro-cutaneous fistula constitutes the main complication of urethroplasty for hypospadias. The frequency of this complication ranges from 12 to 90%. The multiplicity of fistula closure techniques, reflects the difficulty involved in repairing these lesions. The authors evaluated fistula repair by a double pedicle skin flap according to the Wise technique. MATERIAL AND METHODS: 32 patients were treated: 18 had already undergone 34 attempted repairs of urethro-cutaneous fistulas complicating urethroplasties (9 Mathieu, 17 Duplay and 6 Leveuf) for anterior penile (9), middle penile (14) and penoscrotal hypospadias (9). In 14 cases, the Wise technique was the first procedure used to repair the fistula. RESULTS: The authors obtained 29 successes. The 3 failures concerned fistulas after Duplay urethroplasties to correct penoscrotal hypospadias. The overall success rate of this technique in our series (90%) was therefore close to that reported by Wise (5 out of 6 successes), who described this technique in 1977. CONCLUSION: The Wise technique is a good technique for repair of large, recurrent urethro-cutaneous fistulas after several attempts of repair. PMID- 10857145 TI - [Spermatic cord torsion in adults]. AB - OBJECTIVES: The objective of this retrospective study, conducted between January 1988 and December 1998, was to demonstrate that physical examination is sufficient to manage patients with torsion of the spermatic cord. MATERIAL AND METHODS: 72 patients, admitted with an empirical diagnosis of torsion of the spermatic cord based on physical examination, underwent emergency surgery via a scrotal incision. Orchidopexy was performed when the surgical diagnosis was in favour of torsion. We classified our series into 2 groups according to the presence or absence of torsion and we defined two subgroups in the torsion group: the orchidectomy subgroup and the non-orchidectomy subgroup. RESULTS: The suspicion of torsion of the spermatic cord was confirmed in 70.8% of cases. No mortality or morbidity were observed for wrongly operated patients. The study of the sensitivity (Se) and positive predictive value (PPV) of clinical signs defined a group of men requiring emergency surgical exploration: men presenting with scrotal pain in the absence of any urinary signs (Se = 98%, PPV = 72%), negative urinary dip-stick (Se = 97%, PPV = 71%), with pain described as violent (Se = 80%, PPV = 76%), an ascended testis (Se = 62%, PPV = 86%) and a subacute stage (Se = 68%, PPV = 79%). The only pejorative factor detected in the presence of testicular necrosis was a delay before management greater than 6 hours. The length of hospital stay (p < 0.041) and the complication rate (p < 0.023) were greater in the orchidectomy subgroup compared to the non-orchidectomy subgroup. CONCLUSION: This retrospective study confirms that physical examination is sufficient to ensure good management of torsion of the spermatic cord in adults and that emergency surgical exploration is justified at the slightest doubt. PMID- 10857146 TI - [Penile-perineal-scrotal gangrene. Epidemiologic, diagnostic, and therapeutic features. Report of 32 cases]. AB - The authors analyse the epidemiological, diagnostic and therapeutic aspects of peno-perineo-scrotal gangrene in the urology department of a university hospital of a developing African country. The frequency of this disease is low, four cases per year, and most patients (about 72%) are between the ages of 40 to 70 years. The authors identified urethral stricture (31.25%) and idiopathic causes as the main predisposing factors, and many pathogenic bacteria, including P. mirabilis, P. aeruginosa, S. aureus, K. pneumoniae etc., as decisive factors. The treatment of peno-perineo-scrotal gangrene must be both medical and surgical. In addition to the medical treatment currently performed in the department, the authors would like to combine hyperbaric oxygen therapy and, as in Japan, the use of argatroban to accelerate wound healing. PMID- 10857147 TI - [Trauma of the renal pedicle in children. Report of 2 cases of late revascularization with endovascular prosthesis]. AB - OBJECTIVE: The diagnosis of renal pedicle rupture in children is difficult and often delayed. Ultrasound alone is insufficient in the assessment of all cases of renal contusion. This often leads to nephrectomy, due to the uncertain results of a late surgical revascularization procedure. A minimally invasive endovascular approach is therefore sometimes useful. CASE REPORTS: 1st case: a 6-year-old boy was admitted to the emergency department with contusion of the right flank caused by a road accident. The immediate test for haematuria was negative. Twenty-four hours after trauma, renal duplex ultrasound was performed due to the appearance of microscopic haematuria and it demonstrated trunkal thrombosis of the right renal artery, while the initial ultrasound was normal. 2nd case: a 15-year-old girl who jumped out of a window. 48 hours after the trauma, IVU was performed because of persistent microscopic haematuria and revealed a silent kidney, while the initial ultrasound was normal. Renal arteriography showed complete dissection of the right renal artery in both cases. Stenting was performed. The postoperative course was uneventful. In the first case, follow-up duplex ultrasound revealed a modification of the echostructure of the superior pole (absence of blood supply) while the lower pole had a normal interlobular blood supply. CT scan at the 2nd month confirmed normal excretion from the lower pole. In the first case, revascularization was satisfactory on the follow-up duplex ultrasound. After 20 months of follow-up, the kidney presented a normal functional and morphological appearance. CONCLUSION: The diagnosis of renal pedicle lesions remains difficult and is based on emergency CT angiography. Treatment by vascular stenting can be performed in children. In some cases of renal artery dissection, it can constitute an alternative to surgery. However, it raises the question of the medium- and long-term repercussions of renal artery stenosis on the child's growth. PMID- 10857148 TI - [Inverted papilloma of the pyelo-ureteral junction, current literature data]. AB - The authors report the fourth case of inverted papilloma of the ureteropelvic junction since the first description of this lesion in 1963 by Potts and Hirst. In view of the rarity of this lesion and its unusual presentation, they conducted an exhaustive review of the literature concerning inverted papillomas of the upper urinary tract. This proliferative lesion of the urothelium can precede, accompany or follow superficial or invasive urothelial tumours. Rigorous treatment and surveillance are therefore justified to detect an associated tumour. The controversy concerning the benign nature of inverted papilloma has still not been clarified. PMID- 10857149 TI - [Nephrogenic metaplasia of the ureter]. AB - The authors report a case of nephrogenic metaplasia of the ureter in a 35-year old patient presenting with left low back pain, haematuria and frequency. Physical examination was not contributive. IVU showed a filling defect in the left iliac ureter. Ureteroscopy demonstrated a tumour nodule in the left iliac ureter. Ureteroscopic biopsy with histological examination showed nephrogenic metaplasia of the ureteric mucosa. Treatment consisted of ureterotomy with resection of the ureteric polyp, followed by closure of the ureterotomy over a double J stent. The course was favourable and postoperative follow-up (IVU, cystoscopy) at 6 months and 12 months was normal. PMID- 10857150 TI - [Splenic lobulation and pseudo-cancer of the left kidney]. AB - Discovery of a mass in the spleno-reno-pancreatic region requires complementary investigations to exclude a false diagnosis of renal tumour. While resembling a renal tumour, such masses can actually correspond to splenic lobulation, a cyst of the tail of the pancreas or a splenic entity (accessory spleen or splenosis). PMID- 10857151 TI - [Migration of an intrauterine device to the bladder]. AB - The authors report the case of a patient presenting with recurrent urinary tract infection and deterioration of the general status. Ultrasonography, plain x-rays and especially cystoscopy demonstrated the presence in the bladder of an intrauterine contraceptive device inserted 4 years previously. Endoscopic removal combined with antibiotics ensured cure of this patient. PMID- 10857152 TI - [Report of a case of compartmental syndrome secondary to prolonged lithotomy position]. AB - The lithotomy position is widely used in urological surgery to obtain adequate exposure of the perineal plane. It is used, for instance, for stenosis of the posterior urethra. Fortunately, it rarely gives rise to complications although if the operation takes a long time the patient may suffer various adverse reactions; these range from simple peroneal nerve distress to thromboembolism [1, 2] and the much more serious "compartmental syndrome" [2, 3, 4]. There is still debate about the best therapeutic approach to a lesion caused by prolonged muscle compression. Some suggest immediate fasciotomy, whether others hold out for conservative treatment. We report here a case of compartmental syndrome arising in a patient who had to remain the the lithotomy position for a long time, which responded well to conservative treatment. PMID- 10857153 TI - [Report of 2 cases of prostatic abscess]. AB - Prostatic abscess is a rare disease. In the light of two cases, the authors discuss the diagnostic and therapeutic aspects of this disease. Two patients, aged 17 and 55 years, presented nonspecific clinical features. Medical imaging (US, CT) established the diagnosis by showing a loculated cystic prostatic mass. Treatment consisted of transurethral drainage and antibiotics with a favourable course in both cases. Prostatic abscess is a rare disease for which the diagnosis has been facilitated by progress in medical imaging. The treatment of choice remains transurethral endoscopic drainage. PMID- 10857154 TI - [Peroperative erection in lower urologic surgery]. AB - Erection is a rare event during lower urinary tract surgery. When it is extremely refractory, endoscopy is impossible and the lower urinary tract surgery may need to be deferred. The development of erection during spinal anaesthesia is due to manipulations before complete installation of sensory block or incomplete blockade of sacral segments (S2 to S4) supplying the nervi erigentes. The mechanism of erection during general anaesthesia remains poorly elucidated. General anaesthesia may suppress central or peripheral sympathetic control of flaccidity. The peripheral target could be the smooth muscle of the cavernous tissue, either by a direct action or via alpha-adrenergic receptors. Psychogenic and reflexogenic stimuli have also been proposed, possibly facilitated by amplification of sensations during stage II anaesthesia. The various treatments are based on experience acquired in the field of priapism. Systemic treatments such as ketamine an beta 2 mimetics or benzodiazepines are not always effective and are associated with considerable adverse effects. Intracavernous injections of alpha-adrenergic drugs constitute the treatment of choice. Phenylephrine and etilefrine are preferred because of their rapid efficacy and particularly their only moderate cardiovascular adverse effects. PMID- 10857155 TI - [Distal urethroplasty by ventral pediculated flap with penile skin: technical aspects]. AB - The authors present a distal urethroplasty technique (or urethromeatoplasty) which consists of reconstruction of the floor of the distal urethral (glandular urethra) by inversion of a pedicled ventral penile skin patch graft raised transversely. This technique allows treatment of recurrent stricture of the glandular urethra in circumcised patients with good aesthetic and functional results. PMID- 10857156 TI - [Male urinary incontinence and dynamic graciloplasty]. AB - Only a few surgical treatments are available for male urinary stress incontinence and artificial urinary sphincter remains the reference treatment. It is associated with a certain morbidity and specific technical limitations and can therefore not always be used in a given patient. Stimulated graciloplasty could constitute a useful alternative treatment in exceptional situations. The objective of this study was to review the technique, the indications already defined apart from urinary incontinence and the results of stimulated graciloplasty in urology. PMID- 10857157 TI - Efficacy of immunoscintigraphy for detection of lymph node metastases. AB - The size of a lymph node is not in principle a limitation for the detection of cancer by Nuclear Medicine techniques. A radioactive pinhead is detectable if it has enough radioactivity on it. The approach of Nuclear Medicine to the demonstration of impalpable lymph nodes or to those lymph nodes detected by radiological techniques that are under 1 cm as to whether or not they contain cancer, is to increase the activity attached to cancer cells in such a lymph node as much as possible and to use sophisticated image analysis techniques to distinguish such uptake from its environment. This may be undertaken using a non specific technique such as F-18 Deoxyglucose and Positron Emission Tomography which is highly sensitive and which has been successful. The alternative approach is to use a highly specific and sensitive agent, such as a radio-labelled peptide or a radio-labelled monoclonal antibody together with image analysis. This paper describes these approaches and in particular the use of Tc-99m SM3 monoclonal antibody in the detection of impalpable axillary nodes in patients with breast cancer before surgery, using a change detection analysis providing a probability map of the significance of uptake of this radiopharmaceutical. It is a robust approach, providing the patient and the surgeon with information as to the likely need for extensive axillary surgery well prior to operation. A negative study should be followed by a sentinel node evaluation at surgery. PMID- 10857158 TI - Sensitivity and specificity of positron emission tomography (PET) for the diagnosis of lymph node metastases. AB - The introduction of Positron Emission Tomography (PET) and the use of the glucose analog F-18-Deoxyglucose (FDG) can help to improve the sensitivity of the diagnosis of lymph node metastases. Sensitivity exceeding 90% can be achieved when advanced imaging protocols and image reconstruction methods are used for PET. Superior staging information is obtained with PET as compared to morphological imaging methods for the most frequent tumor types. The accuracy of N-staging can be significantly improved by adding PET to the pretherapeutic diagnostic procedures. Limitations exist with regard to false positive results. Acute or chronic inflammation as well as unspecific reactions following radiotherapy may mimic tumor tissue. PMID- 10857159 TI - Diagnosis and staging of lymph node metastasis. AB - The process of lymph node metastasis is not completely understood. Although we know some basic molecular mechanisms of metastasis, the exact procedure from the initiation of a primary tumour to overt lymph node metastasis remains obscure. A few morphological features of different primary tumours are known to correlate with the probability of lymph node metastasis, e.g., tumour histology (carcinoma vs. sarcoma), tumour size, T category, poor grade of differentiation and invasion of lymph vessels. Few attempts have been made to use markers of molecular differentiation such as nm23 as an additional indicator of lymph node metastasis. A drawback is the lack of an exact definition of lymph node metastasis or micrometastasis and how to include new findings such as the demonstration of cytokeratin-positive cells in "tumourfree" lymph nodes. Some of these aspects will be discussed within this review and proposals for classifications presented. PMID- 10857160 TI - Detection and relevance of immunohistochemically identifiable tumor cells in lymph nodes. AB - Lymph node metastasis is a well-known feature of poor prognosis in potentially resectable solid epithelial tumors. However, a significant number of apparently lymph node negative patients die early of metastatic disease. Therefore, it has to be assumed that in some patients an early tumor cell dissemination has occurred which is clearly underestimated by current staging procedures. Recently, it has been shown, that an early dissemination of individual carcinoma cells to regional lymph nodes can be detected by using sensitive immunocytochemical techniques with monoclonal antibodies against epithelium-specific proteins. The incidence of immunohistochemically positive patients varies between 12% and 70% depending on the type of primary tumor, the immunohistochemical staining procedure used, and especially on the primary monoclonal antibody. The detection of disseminated tumor cells in lymph nodes by immunocytochemistry is associated with a poorer prognosis in different types of epithelial tumors such as lung cancer or esophageal cancer. The immunocytochemical method might also be useful in the detection of occult tumor cells in sentinel lymph nodes. In conclusion, the immunohistochemical detection of disseminated tumor cells in lymph nodes can help to obtain a more exact identification of patients with an unfavorable prognosis. Whether the identified patients will gain from an adjuvant therapy has to be evaluated in further studies. PMID- 10857161 TI - Cell biology of lymphatic metastasis. The potential role of c-erbB oncogene signalling. AB - Lymphatic metastases are an important indicator of the malignancy of epithelial cancers. Empirical clinical observations associating specific genetic abnormalities with tumour progression, allied with basic laboratory investigations, are providing not only improved prognostic and diagnostic opportunities, but also a detailed understanding of the molecular machinery of metastasis. One such association--between the c-erbB oncogene family and metastasis--has proved particularly instructive. Functional links between over expression (and occasionally mutational activation) of c-erbB-1 (EGFR) and c-erbB 2 and specific phenotypes of metastatic cells have been elucidated. Activated c erbB oncogenes potentiate tumour cell adhesion to endothelial cells and upregulate VEGF, potentially facilitating angiogenesis and vascular invasion. In addition, cells over-expressing these oncogenes frequently show aberrant cell cell and cell-matrix interactions, mediated by changes in integrin and cadherin function. Thirdly, both EGFR and c-erbB-2 signalling can significantly upregulate specific matrix metalloproteinases, key enzymes involved in angiogenesis and invasion. Finally, c-erbB receptors linked to the actin cytoskeleton and highly expressed on invadopodia, are thought to assist cell migration. Taken together, these observations suggest that such receptors can act as "master switches" in metastasis, whose activation co-ordinately controls events normally utilised in development, now subverted by the metastatic cell. As such, they represent ideal targets for therapeutic intervention. PMID- 10857162 TI - The lymph node as a bridgehead in the metastatic dissemination of tumors. AB - The metastatic spread of tumors is not a random process. Distinct patterns of metastasis can be discerned which vary from tumor type to tumor type. A common pattern, particularly for carcinomas, is that regional lymph nodes are often the first organs to develop metastases. This pattern of metastasis is central to the utility of the sentinel lymphonodectomy surgical technique. However, not all tumors and tumor types metastasize first to the regional lymph nodes. The mechanisms which determine whether regional lymph nodes or other sites first develop metastases remain poorly understood. In this article I review the anatomical, cellular and molecular factors which play a role in metastatic dissemination and determine patterns of metastasis. I then explore the importance of tumor heterogeneity and the selection of metastatically competent tumor cells during systemic dissemination, and suggest that some secondary sites are more readily colonised by metastasizing cells than others. Metastases at these sites act as bridgeheads, constituting a reservoir of tumor cells which, because they have already successfully metastasized, possess many of the properties required for metastasis to further sites. These tumor cells are therefore more likely than cells in the primary tumor to acquire all of the properties required for metastasis to less favourable secondary sites. To illustrate the bridgehead concept, I argue that features of the design and function of the lymphatic system make it highly amenable to the entry of metastasizing tumor cells and the formation of lymph node metastases, and suggest that lymph node metastases form a bridgehead for further metastatic spread. PMID- 10857163 TI - Functional anatomy of lymphatic vessels under the aspect of tumor invasion. AB - In the human body there are about 800 lymph nodes, of which 300 are located in the head and neck area. The lymphatic system begins with initial finger-shaped lymphatic vessels consisting of valveless lymphatic capillaries and precollectors with valves. The precollectors become collectors and transport the lymph to the so-called lymph stems. These vessels lead into the right and left lymphatic duct and into the jugular trunk of the head and neck region. At the end they join into the blood circulation at the junction point of the jugular and subclavian veins. On the basis of the morphology of the initial lymphatics, these are probably the main port of entry of tumor cells. Investigations demonstrate the active invasion into the lymphatic system of melanoma cells. The preference of melanoma cells to adhere to the extracellular fibronectin of the endothelial cells may be a reason for the privileged invasion of the melanoma cells in the lymphatic system, because the extracellular matrix is not protected by a continuous basal lamina like blood vessels. Based on the directed movement of melanoma cells towards the fiber felt, there might be regulative mechanisms influencing the process of targeted adhesion. PMID- 10857165 TI - The concept of the sentinel lymph node. AB - Lymphangiograms performed via direct cannulation of lymphatic ducts demonstrate drainage of the lymph into a specific lymph node center, the so-called SLN. Contrast materials such as lipiodol, injected directly into the tissue (e.g., tongue) can demonstrate the SLN. As the neoplastic cells can be carried through the lymphatic ducts, the SLN is the first filter in the lymphatic pathway, and the SLN is indeed the most likely regional node to harbor metastatic carcinoma. The results of these efforts challenged the surgical community worldwide to recognize the importance of the concept of SLN. This concept needs to be inexpensive and easily applied in daily practice. Recently, brilliant investigators have found that using "blue dye" and or radioactive tracers are a resourceful way in identifying SLN and have applied the benefits in their daily practice. Morton [15] using the "blue dye" and Krag [1] using radioactive tracers are pioneers in the application of these concepts in other malignant diseases. The SLN concept today is feasible to apply in the investigation, diagnosis, staging and treatment of almost all solid tumors in human pathology. Numerous elegant reports have proved the validation of the concept [2, 7-9, 11, 12, 16, 17, 20, 21, 26-28]. PMID- 10857164 TI - Molecular diagnosis of head and neck cancer. AB - Patients with advanced stages of head and neck cancer frequently develop locoregional recurrence as well as distant metastases. These data indicate that traditional diagnostic methods such as histopathology and radiology are not sensitive enough to detect the small numbers of tumor cells which are left behind, defined as minimal residual disease (MRD). Sensitive diagnostic assays based on molecular markers appear to be powerful tools to improve the staging of these patients. At the DNA level, tumor-specific p53 mutations seem to have great potential for the detection of "occult" tumor cells at surgical margins and lymph nodes. At the RNA level HNSCC associated antigens like the E48 antigen, allow the detection of rare HNSCC cells in blood and bone marrow and, it is hoped, also in lymph nodes and lymph node aspirates. However, the molecular assays which are used to detect MRD are subject to certain (technical) problems which affect their sensitivity and specificity. In this paper we will present examples of molecular assays such as the plaque assay using p53 mutations and the E48 RT-PCR, and show their use for MRD detection in cervical lymph nodes. In addition, we will discuss the problems and pitfalls associated with these sensitive techniques. PMID- 10857166 TI - Sentinel node localization by lymphoscintigraphy: a reliable technique with widespread applications. AB - The concept of the sentinel lymph node (SN) represents an important contribution to guide appropriate surgery of cancer. Diagnostic non-invasive or minimally invasive procedures that provide accurate preoperative staging of the lymph node status are badly needed. The technique of SN biopsy, first developed with the purpose to select melanoma patients for regional node dissection, has been extended to other malignancies. Initial studies in breast carcinoma, conducted with vital blue dye, showed that the SN concept was biologically valid, although SN was missed in up to 30%-40% of cases. If a radioactive tracer is injected close to the tumor, then the SN can be identified by lymphoscintigraphy (LS), and a gamma ray detecting probe (GDP) can be used to locate the skin projection of SN and assist biopsy. These techniques are already used successfully in melanoma and breast carcinoma where the various parameters involved, such as the size of the radioactive particles, the injection site and injection volume, have recently been optimized. In a large series of breast cancer patients, the overall predictive value of the SNs biopsy guided by LS and GDP was 96.8%; in patients with small carcinomas (< 1.2 cm diameter), the concordance between SN and axillary status was 98.6%. In patients with melanoma, LS combined with GDP showed itself to be superior to the blue dye mapping. LS associated with GDP allowed the detection of SN in 98% of cases and 72 SNs in 54 basins were localized. Using blue dye instead, SN was stained only in 80% of patients (50 SNs in 40 basins). Lymphoscintigraphic techniques have shown promising results also in tumors such as vulva and tongue. In conclusion, LS is a simple nuclear medicine technique, relatively inexpensive and well accepted by patients. SN biopsy guided by a GDP is becoming widely adopted for a variety of neoplasms, contributing significantly to the search for less aggressive treatments in patients with early stages of cancer. PMID- 10857167 TI - Vital dye and radiolabelled colloids--complement or alternative? AB - Several different protocols for retrieval of the sentinel node (SN) have been described: gamma probe (GP) and/or dye guided biopsy, preceded by lymphoscintigraphy or not. Especially in American studies, predominantly executed by surgeons, dye or GP guidance only is used with good results. The disadvantages of applying dye only are: an extensive learning curve, lower retrieval rate of the SN and, especially in the learning phase, a higher rate of false negative biopsies. If only GP guidance is applied, the technique seems more simple to master. A recent multicentre study, however, revealed an unacceptably high false negative rate. It must be considered that most published studies were executed by highly experienced surgeons. In most European studies, scintigraphy is a standard part of the procedure. Lymphoscintigraphy provides the surgeon with a "road map", revealing the number and approximate location of the SNs in the lymphatic basin(s). Scintigraphy proves useful especially if an SN is situated close to the injection site (breast cancer), or if SNs are situated at unexpected locations (head-and-neck or trunk melanoma). A combination of all three available steps results in the highest number of successful procedures with the lowest false negative rate. This may prove to be especially important for general hospitals where the number of biopsy procedures is often smaller compared to specialized centres. PMID- 10857168 TI - Application of the sentinel node concept in urogenital cancer. AB - Pre and intra-operative lymphatic mapping and SLN biopsy is readily available in the armamentarium of Urologic Surgical Oncology. The concept of the SLN should be considered for diagnosis and therapy. Ongoing studies are investigating the impact of SLN biopsy on survival as well as the prognostic significance of micro metastasis founded by histopathology and immunohistological techniques. The concept is simple, the operatory technique is easy, and the blue dye and radioactive agents are available anywhere. The validation in penile carcinoma has demonstrated, based on experience, that metastasis does not occur in other lymph nodes if the SLN is free of metastasis. Its applicability in other areas of urology is still awaiting the enthusiasm of investigators. PMID- 10857169 TI - Intraoperative lymphatic mapping and sentinel node identification: gynecologic applications. AB - The status of regional lymph nodes is a powerful predictor of survival in patients with early cancers of the vulva, cervix, and uterus. Radical resection of vulvar and cervix cancers along with extensive lymphadenectomy remains the standard of care for these cancers. Intraoperative lymphatic mapping and sentinel node identification has the potential to improve the treatment of patients with gynecologic cancer with improved detection of lymph node metastases and reduced morbidity. This chapter will focus primarily on vulvar cancer and include a review of previous innovations in treatment and current experience with intraoperative lymphatic mapping in these patients. PMID- 10857170 TI - Sentinel node detection in malignant melanoma. AB - The initial application of intraoperative lymphatic mapping and sentinel lymphadenectomy followed by selective complete lymphadenectomy (LM/SL/SCLND) was in melanoma. This arose as a solution to the ongoing debate concerning immediate vs. delayed lymph node dissection. Acceptance of the concept and advances in nuclear medicine, surgery, and pathology aspects of the sentinel node procedure have brought it into widespread use for melanoma and have expanded its application for other solid tumors that progress through the lymphatic route. Although the diagnostic accuracy of the procedure has been demonstrated in multicenter trials, caution should be exercised regarding therapeutic aspects until definitive benefit can be shown from well-designed clinical trials. Current issues of active discussion and debate are reviewed including ideal nomenclature, clinical significance of occult metastatic disease, quality assurance, and the role of LM/SL/SCLND outside high-volume melanoma centers. PMID- 10857171 TI - Adjuvant therapy of malignant melanoma and the role of sentinel node mapping. AB - BACKGROUND: Controversy still exists about standard management of a primary melanoma. Over the last decades randomized phase III trials have addressed questions about the width of margin in relation to the Breslow thickness of the primary lesion, the role of prophylactic isolated limb perfusion, and the role of elective lymph node dissection. Overall these trials have demonstrated that less extensive surgery is as good as more extensive surgery. Wide excision margins, prophylactic isolated limb perfusions, or the elective lymph node dissection did not improve overall survival significantly in any of the phase III trials conducted. ADJUVANT THERAPY IN HIGH RISK MELANOMA: No standard systemic adjuvant therapy with confirmed impact on overall survival has been identified thus far for clinically node negative stage I-II (TxN0M0) patients after excision of the primary, nor for clinically node positive stage III (TxN1-2M0) patients after lymph node dissection for metastatic regional node involvement. Poor Staging in the Past. One of the main problems associated with the trials assessing systemic adjuvant treatments in management of high risk primary melanoma is the fact that in general patients were poorly staged. About 25%-30% of patients with primaries thicker than 1.5 mm have micro-metastatic disease in the regional lymph nodes and beyond. This population was usually submerged by the other 70%-75% of the patients with excellent prognosis, obscuring the potential benefit of the adjuvant surgical procedure (ELND) or a systemic adjuvant treatment. SENTINEL LYMPH NODE MAPPING: Sentinel lymph node (SLN) mapping is resolving many of the inadequacies of the past and has completely changed the management of primary melanoma. As a small procedure with low morbidity it identifies that part of the population which has microscopic involvement of regional lymph nodes with greater precision than an elective lymph node dissection. SLN-mapping allows for a detailed histopathologic evaluation involving multiple sections, H&E staining in combination with IHC (immunohistochemical staining) of the node with the highest chance of containing metastatic foci. Moreover in the near future it is most likely that RT-PCR on negative nodes will complete the diagnostic workup as a promising last step in the procedure to determine whether tumor cells are present in the sentinel node. Sentinel lymph node status has been shown recently to be by far the strongest independent prognostic factor of melanoma stage I-II patients. SLN-status is a much stronger prognostic factor than tumor thickness, which looses its prognostic relevance in SLN-positive patients. CONSEQUENCES FOR DEVELOPMENT AND/OR ALLOCATION OF ADJUVANT THERAPY: Thus we now have a procedure by which the melanoma stage I-II population can be dissected in a group at truly high risk for recurrence and a group with truly low risk of recurrence. The high risk group with a greater than 75% chance for systemic disease can then be selected for trial participation of various systemic adjuvant therapy regimens that may be allowed to be toxic, considering the very high risk for relapse in these patients. The node negative group of patients can be selected for participation in trials evaluating systemic adjuvant treatment of low toxicity considering the low chance for distant metastatic disease. PMID- 10857172 TI - Selective neck dissection and sentinel node biopsy in head and neck squamous cell carcinomas. AB - The Sentinel Node concept is now well established for HNSCC and gives us a strong basis to treat patients with N0 neck where the rate of occult node metastasis is high. At the present time, the most accurate method for staging N0 neck is pathologic examination of the neck content. In this way, sentinel node dissection (SND) and sentinel node biopsy (SNB) are complementary surgical procedures. SNB has limited indications in HNSCC because of the inaccessibility of most of the primary sites to local injection of Tc99m colloid. However it seems to be an encouraging approach for small tumors of the oral cavity. In other primary sites, except for small glottic tumors, patients must undergo an SND. Supraomohyoid neck dissection which removes levels I, II and III, is performed in oral cavity tumors. Lateral neck dissection which removes levels II, III and IV, is used by many authors for laryngeal, oropharyngeal and hypopharyngeal tumors. In our experience, SND could be limited to levels II and III for laryngeal and oropharyngeal tumors without more neck failures. SND is a reliable procedure, we report only 1.5% of skip nodal metastases in 464 patients who had this staging procedure. PMID- 10857173 TI - Sentinel lymph node dissection for thyroid malignancy. AB - Sentinel lymph node dissection (SLND) for melanoma and breast cancer has been validated as an accurate technique to assess the status of the lymph nodes in the regional drainage basin. The sentinel node concept has also been investigated in other solid tumors, and more recently, in thyroid carcinoma. SLND using a vital blue dye during thyroidectomy for suspected thyroid malignancy successfully identifies sentinel nodes, with minimal morbidity. Excised sentinel nodes can be examined for micrometastases, and if negative, then the rest of the cervical nodes are likely to be negative. The false negative rate of SLND for thyroid malignancy is unknown, however, because modified neck dissections have not accompanied all cases. The impact that lymph node metastasis in thyroid carcinoma has on prognosis is debatable, unlike breast cancer and melanoma, which therefore makes the utility of thyroid SLND less clear. The technique, results, and morbidity of SLND during thyroidectomy is presented, and its possible utility in well-differentiated and medullary thyroid carcinoma is discussed. PMID- 10857174 TI - Lymphoscintigraphy and ultrasound-guided fine needle aspiration cytology of sentinel lymph nodes in head and neck cancer patients. AB - Accurate staging of the regional lymph nodes is crucial for the appropriate management of patients with squamous cell carcinoma of the head and neck (HNSCC). However, the current diagnostic modalities have low accuracy for N0 neck, and even the most optimal procedure, ultrasound-guided fine needle aspiration cytology (USgFNAC), still has a sensitivity of only 42%-73%. In this study we evaluated whether the identification of the sentinel node might improve the selection of lymph nodes for USgFNAC. Twelve HNSCC patients received 3-4 peritumoral injections of 10-30 MBq 99mTc-labeled colloidal albumin, and the sentinel node was identified by dynamic scintigraphy and marked on the skin using a handheld probe, and/or by scintillation counting of the aspirates. After sentinel node identification USgFNAC was performed. Correct aspiration of the identified sentinel node(s) was confirmed by scintillation counting. In 11 out of 12 cases the sentinel node(s) could be visualized by dynamic planar imaging. In one case the sentinel node(s) were identified by scintillation counting only. In a number of patients different or supplementary lymph nodes were aspirated on the basis of sentinel node identification. These initial data strongly suggest that sentinel node identification might improve the staging of the neck by USgFNAC. PMID- 10857175 TI - Present and future of sentinel node lymphadenectomy in breast cancer. PMID- 10857176 TI - Specification of potential indications and contraindications of sentinel lymph node biopsy in breast cancer. AB - The promising results of various studies applying different methods of SN biopsy in heterogeneous patient subpopulations, show that sentinel lymph node biopsy is a reliable and minimally invasive method for the determination of the nodal status in breast cancer patients. While multicenter studies for the evaluation of the method's accuracy are still ongoing, future indications and contraindications are discussed. Based on our own experience, we try to give an actual overview of the potentials and problems of sentinel node biopsy in breast cancer. Sentinel node detection was performed in 146 patients with breast cancer stages I-III, consisting of 127pT1/2 tumors and 19pT3/4. All of them underwent standard axillary dissection after SN biopsy. Using the radionuclide method including preoperative lymphoscintigraphy and intraoperative gamma. Probe detection, the detection rate varied in relation to the tumor size between 94% for tumors with a diameter < 1 cm, 85% (1-3 cm), 70% (3-5 cm) and 63% (> 5 cm). The accuracy of the SN-biopsy in the prediction of the nodal status varied also with tumor diameter ranging from 100% for very small tumors (< 1 cm), over 97% (1-3 cm) and 88% (3-5 cm), to 67% (> 5 cm). In the subgroup of patients with pT1-2 tumors (n = 106), 57 patients (53%) showed true negative sentinel nodes, 38 (36%) revealed tumor cells in the H&E staining and an additional 7 patients (7%) solely in the immunohistochemical staining. 4 (4%) of these patients, all of them from the first half of the study period, underwent false-negative SN-biopsy, all of them showing lymphangiosis carcinomatosa and/or extensive infiltration of the metastatic lymph node(s). The results presented show, that in about 50% of early breast cancer patients surgical intervention could potentially be avoided after a negative SN-biopsy, and an additional 5-10% of conventionally nodal negative patients can be found by immunohistochemical examination of the sentinel node. The SN concept is not recommended in clinically nodal positive patients or advanced disease. Potential applications include the evaluation of parasternal lymph nodes and patients with recurrent tumor. Before clinical application, quality control of the medical center and the performing surgeon have to be established potentially including the performance of about 20 procedures under supervision for each surgeon, an individual accuracy of at least 93%, and the possibility of immunohistochemical staining as well as a regular follow-up. PMID- 10857177 TI - Potential and pitfalls of sentinel node detection in breast cancer. AB - The last decade has seen the development of a minimally invasive technique to identify representative nodes--sentinel nodes--that reflect the tumor status of nodes in the axillary lymphatic basin draining a primary breast carcinoma. Sentinel lymph node dissection (SLND), originally developed as an alternative to elective complete lymph node dissection in patients with primary cutaneous melanoma, has been applied successfully to the management of patients with breast cancer. SLND holds promise as a staging technique to replace formal level I and II axillary lymph node dissection in selected patients with breast carcinoma, thus avoiding an unnecessary procedure that has no role in many patients with tumor-free axillae. Under way are two large randomized trials examining the role of SLND for the management of patients with invasive breast carcinoma. Even when tumor is detected in the sentinel node, a focused examination of this node may indicate whether or not completion axillary lymph node dissection is necessary. However, although SLND has great potential, its successful widespread use requires more stringent definition of the sentinel node and standardized guidelines for lymphatic mapping. Each institution must carefully assess the accuracy and consistency of results obtained by its multidisciplinary SLND team. PMID- 10857178 TI - Gastric lymphography and detection of sentinel nodes. AB - The lymphatic drainage system of the stomach was studied using lymphography with various dyes, and several major routes have been shown. Gastric lymph channels are multidirectional and form complex networks. We conducted a retrospective study to know the first site of metastasis from small gastric cancers by examining 89 cases with only one lymph node metastasis. The perigastric nodal area close to the primary tumor was the first site of metastasis in only 62% of the cases. N2 metastasis without N1 involvement was seen in 13%. In order to identify sentinel nodes for local resection of gastric cancer, a novel method needs to be developed. PMID- 10857179 TI - Potential and futility of sentinel node detection for gastric cancer. AB - The technique and scientific background of sentinel node dissection has spread extremely rapidly over the surgical community. Following the addition of this technique to the tools of oncologic surgery for treatment of malignant melanoma and breast cancer, questions arise regarding the use of this method in gastric cancer also. While the lymphatic flow on the surface of the body can be defined easily, the lymphatic drainage of the stomach is much more complicated. Following rotation of the stomach during embryonic development, the lymphatic flow is not directed in a simple fashion. It is questionable whether a specific area of the stomach will drain into one lymph node echelon only. This is one of the essential obstacles for SLND in gastric cancer. Furthermore, skip metastasis seems to be quite common in cancer of the stomach. In gastric cancer, the value and the extent of classical lymph node dissection itself is still under scientific discussion. The rationale, aims, and extent of LA in gastric cancer are addressed. The scientific discussion on whether D1 or an extended lymphadenectomy are appropriate is not finally closed as yet. The possibilities and problems concerning an individualised indication for a selective lymphadenectomy in gastric cancer are discussed. PMID- 10857180 TI - Probe-guided surgery for colorectal cancer. AB - Anti-CEA-scintigraphy turned out to be very reliable in detecting primary and recurrent colorectal cancer, its overall accuracy being more than 90%. The intraoperative application of this technology should provide similar results when focussing at extrahepatic tumor deposits, for example in lymph nodes, thus allowing accurate staging of the underlying disease. To test this hypothesis we launched the following feasibility study the results of which are compared to those reported in the recent literature. We investigated 20 patients, six with rectum and 14 with colon cancer. 24 hours before surgery they were intravenously given 1 ml of a fab'-fragment-antibody to CEA, labeled with 25 mCi of 99mTc (CEA Scan). During surgery the radioactivity in lymph glands regional to the tumors was measured and compared to the much lower activity in healthy nodes. For this we used a scintillation probe (C-Trak, Care Wise, Inc., Morgan Hill, CA). All lymph nodes of interest were then excised and submitted to frozen section pathology. In 7 out of 20 cases scintimetry led to an up-staging of the disease. In addition we found metastatic spread to lymph nodes that were basically not regional to the primary tumor (retroperitoneum, renal hilum etc.). Scintimetry can precisely identify even very small tumor deposits. So it leads to accurate staging while surgery is still ongoing. In a further step the concept of sentinel node diagnosis, which is right now being clinically evaluated, may some day be applied in colorectal surgical oncology. PMID- 10857181 TI - Identification of lymph node metastases in recurrent colorectal cancer. AB - Lymph node metastases are an important prognostic prediction factor in patients with recurrent colorectal cancer, particularly those with liver metastasis. Fifty six patients with recurrent colorectal cancer were operated by us using the RIGS (radioimmunoguided surgery) technology. Patients were injected with 1 mg monoclonal antibody (MoAb) CC49 labeled with 2 mCi 125I. In surgery, traditional exploration was followed by survey with a gamma-detecting probe. Sixty of 151 patients enrolled in the Neo2-14 Phase III study for recurrent colorectal cancer were diagnosed with liver metastases based on preoperative CT. In 17/56 patients (30%), RIGS identified at least one tumor site confirmed by pathology (H&E). This resulted in 16 major changes in surgical plan. RIGS performance varied between lymphatic and non-lymphatic tissue, with positive predictive value (PPV) of 100% and negative predictive value (NPV) of 94% for non-lymphoid tissue, compared to PPV of 46.5% and NPV of 100% for the lymphoid tissue. Thirty-five out of 60 patients were considered resectable after traditional evaluation. RIGS identified occult tumor in 10 of these patients (28.5%). 7/10 occult patients expired (70%), while only 7/25 of the non-occult patients expired (28%) (P = 0.046). In localizing patients, no RIGS activity in lymph nodes signifies no tumor, while H&E confirmation is needed for decisions based on RIGS activity in the lymph nodes. RIGS provides important staging information, identifying patients for whom surgery may be done with curative intent. PMID- 10857182 TI - Fluorescence as a concept in colorectal lymph node diagnosis. AB - Fluorescence detection may constitute an appropriate means in gastrointestinal cancers to diagnose lymphatic tumor spread as opposed to gamma-scintillation methods. Photodiagnostic tracers have been shown to localize rapidly in malignant cells and may enable sensitive detection of small cell aggregates in lymph nodes. To reach a detection depth of several millimeters, a broad banded unspecific tissue autofluorescence may be controlled by so-called background subtracting techniques, generally based either on fluorescence observation at several wavelengths or on dual-wavelength fluorescence excitation. Using such comparative fluorescence detection techniques, some tumor entities can be differentiated soley based upon autofluorescence characteristics. Introducing a further enhancement in sensitivity for longer life-time fluorophores by time delayed fluorescence detection we ran a pilot trial comprising 174 lymph nodes from colorectal cancer specimen from 9 patients. Metastatically involved lymph nodes could be differentiated from all other palpable nodes in the mesenteric fat at a specificity of 85% with a sensitivity of 65%. Specific fluorescence features may be useful to preselect tissue samples for further histological analysis. PMID- 10857183 TI - From spontaneous to induced neurological mutations: a personal witness of the ascent of the mouse model. PMID- 10857184 TI - Mapping genes that modulate mouse brain development: a quantitative genetic approach. PMID- 10857185 TI - Genetic interactions during hindbrain segmentation in the mouse embryo. PMID- 10857186 TI - Neurogenetic compartments of the mouse diencephalon and some characteristic gene expression patterns. PMID- 10857187 TI - Neuronogenesis and the early events of neocortical histogenesis. PMID- 10857188 TI - Programmed cell death in mouse brain development. PMID- 10857189 TI - Neurotrophic factors: versatile signals for cell-cell communication in the nervous system. PMID- 10857190 TI - Growth factor influences on the production and migration of cortical neurons. PMID- 10857191 TI - Signalling from tyrosine kinases in the developing neurons and glia of the mammalian brain. PMID- 10857192 TI - The role of the p35/cdk5 kinase in cortical development. PMID- 10857193 TI - The Reelin-signaling pathway and mouse cortical development. PMID- 10857194 TI - The subpial granular layer in the developing cerebral cortex of rodents. PMID- 10857195 TI - Development of thalamocortical projections in normal and mutant mice. PMID- 10857196 TI - [The molecular genetic mapping of cereal crops]. AB - The application of modern methods of genetic mapping using RFLP and PCR technologies allowed to advance essentially in construction of rye genome genetic maps and mapping of some morphological and breeding-valuable genes. Genetic mapping of cereal genomes, such as rye, wheat, maize and rice using common set of DNA-probes permitted to reveal considerable evolutionary conservation in gene organization and localization. This allows to use more effectively method of comparative mapping for fast localization and tagging of genes in genomes of less investigated species. PMID- 10857197 TI - Complex structure of knobs and centromeric regions in maize chromosomes. AB - The recovery of maize (Zea mays L.) chromosome addition lines of oat (Avena sativa L.) from oat x maize crosses enables us to analyze the structure and composition of individual maize chromosomes via the isolation and characterization of chromosome-specific cosmid clones. Restriction fragment fingerprinting, sequencing, and in situ hybridization were applied to discover a new family of knob associated tandem repeats, the TR1, which are capable of forming fold-back DNA segments, as well as a new family of centromeric tandem repeats, CentC. Analysis of knob and centromeric DNA segments revealed a complex organization in which blocks of tandemly arranged repeating units are interrupted by insertions of other repeated DNA sequences, mostly represented by individual full size copies of retrotransposable elements. There is an obvious preference for the integration/association of certain retrotransposable elements into knobs or centromere regions as well as for integration of retrotransposable elements into certain sites (hot spots) of the 180-bp repeat. DNA hybridization to a blot panel of eight individual maize chromosome addition lines revealed that CentC, TR1, and 180-bp tandem repeats are found in each of these maize chromosomes, but the copy number of each can vary significantly from about 100 to 25,000. In situ hybridization revealed variation among the maize chromosomes in the size of centromeric tandem repeats as well as in the size and composition of knob regions. It was found that knobs may be composed of either 180-bp or TR1, or both repeats, and in addition to large knobs these repeated elements may form micro clusters which are detectable only with the help of in situ hybridization. The association of the fold-back elements with knobs, knob polymorphism and complex structure suggest that maize knob may be consider as megatransposable elements. The discovery of the interspersion of retrotransposable elements among blocks of tandem repeats in maize and some other organisms suggests that this pattern may be basic to heterochromatin organization for eukaryotes. PMID- 10857198 TI - [The chromosomal localization of the major genes for the quantitative traits (QTL) of wheat by using D chromosome gene markers]. AB - The possibility of biochemical and morphological marker genes application for QTL and determination of their association with definite D chromosomes was shown on F2 populations of bread wheat from crossing of hexaploids AABBDD with different D subgenomes and the same genome AB part. Algorithm for calculation of the theoretical means for evaluation of differences according t-criterion was proposed and examined. The results of biometrical genetic analysis of the quantitative traits obtained by joint scaling Cavalli test using 6-10 different generations were used for calculation of theoretical means. PMID- 10857199 TI - [The use of gliadin genetic markers in the selection of winter wheat in Krasnodar]. AB - Influence of gliadin components blocks on technological and baking grain qualities was investigated. It was shown that interaction of gliadin blocks affects on quality. The 1A4 and 1D4 blocks reduce negative action of the block 1B3 on baking quality. Combinations of gliadin components blocks of first three homeological chromosomes: 5.1.7., 10.1.4., 4.1.7., 5.1.3., 4.1.3 were the best in influence on gluten quality in Krasnodar conditions. It was managed to select lines without adverse Gld 1B3 effect. Good quality varieties: Spartanka, Skifyanka, Pobeda 50, Jirovka, Yuna, Hasarka, Verna, Ofelia, Leda, Rufa, Gorlitsa, Echo were created in the Institute. Evolutional approach, gradual development of quality traits, using of different complementary sources of quality in system of complex stepped hybridization allow to adjust "conveyor" for creation of new high-quality varieties, and gliadin markers method allow to identify them at early stages of selection. PMID- 10857200 TI - [The effect of natural selection on the allelic frequency of gliadin-coding loci and their associations in an artificially created hybrid population of winter wheat]. AB - Hybrid population with heterogeneity for five gliadin-coding loci was created by crossing of five winter common wheat cultivars (Belozerskaya 47, Ilyichevka, Kyyanka, Mironovskaya 808, Polesskaya 70) by the complete scheme. Frequencies of alleles at these loci and their associations were analyzed after two, four and ten years of resowing. The ratio of alleles at the studied loci was changed as reproduction occurred. Gli 1A5 allele displayed stable tendency to frequency increasing, Gli 1D1--a little tendency and Gli 6A3--very essential tendency. The share of Gli 6D2 allele increased only after ten years of population reproduction. Combination of alleles at two, three, and five loci was not accidental. The most common regularity was predominance of the frequency of allelic associations characteristic of the parents (except for Ilyichevka) over new combinations. Possible causes of this phenomenon are discussed. PMID- 10857201 TI - [The genetic systems of the type and rate of development in soft wheat]. AB - 25 years studying of particular genetics of wheat growth habit and rate development in Plant Breeding and Genetics Institute of UAAS was summed up. The main genetic systems involved in control of flowering response differences and possibility of application in breeding of genetic diversity of Vrn, Ppd genes, ear-lines per se and duration vernalization requirement were pointed out. PMID- 10857202 TI - [The inheritance of the traits of resistance to fungal diseases by distant hybrids of wheat with amphiploids]. AB - The inheritance of resistance to powdery mildew, leaf rust and septorios of hybrids from crosses of wheat--rye and two wheat--Elymus amphiploids with durum and bread wheat was studied. Spontaneous hybridization was revealed to be a determining factor of the new available trait formation in the wide crosses. Control of the leaf rust and heary leaf traits by the same alien chromosome of Elytricum fertile is confirmed. While in the wheat--Aegilops amphiploid H74/90 258 these traits are not linked and controlled by one and two genes, respectively. The alien resistance is easier introgessed into wheat genome in the cross of 8x-amphiploid with durum wheat, than with bread wheat. Constant homozygous strains with group resistance to the studied diseases have been revealed. PMID- 10857203 TI - [The polymorphism and inheritance of aspartate aminotransferase in apricot]. AB - Scheme of apricot AAT control by Aat-1, Aat-2, Aat-3 loci on the base of formal genetic analysis was created. Determination by three active (a, b, c) and null (n) alleles Aat-1 of interspecies polymorphism of fast migrating components was revealed. Alleles frequencies and heterozygosity of Aat-1 locus were determined in ecologic-geographical groups of apricot as well as varieties of hybrid origin. PMID- 10857204 TI - [The efficiency of inducing spring barley haploids depending on the method of haploid production and on the genotype of the initial diploids]. AB - The comparative assessment of the combining application of bulbosum technique and anther culture in vitro for spring barley haploid production has been presented. It has been shown that the yield of haploids depended on genotype of initial material and genotype and method interaction. The advantages of the use of these two techniques complemented each other in barley breeding program are discussed. PMID- 10857205 TI - [The segregation characteristics at the alleles of the gliadin-coding Gli-B1 locus in hybrid soft winter wheat]. AB - Segregation at the Gli-B1 locus was studied in F2 seeds of common wheat from crosses between near-isogenic lines with respect to this locus. Segregations differed from the expected ratio in hybrids involving the lines with the allele Gli-B1l (Gli-B1-3), which is a marker for the 1BL/1RS translocation, as well as in the hybrid between the lines with the alleles Gli-B1b (1) and Gli-B1e (4). Reduced transmission of the chromosome with the 1BL/1RS translocation through pollen was observed in the hybrids involving the line with this translocation. In the cross GLI-B1-1 x GLI-B1-4, the significantly lower frequency of female gametes with the allele Gli-B1e (4) was detected. This is due to linkage of the Gli-B1 locus to a factor responsible for segregation distortion in female gametes. We proposed to designate this locus Sd3. The line with the gliadin block Gli-B1e differs in alleles at the Sd3 locus from the lines with the blocks Gli B1b and Gli-B1o. PMID- 10857206 TI - [Genetically determined enzyme polymorphism in soy varieties (Glycine max) and in wild soy (Glycine soja)]. AB - Analysis of 22 genetic-biochemical systems (42 loci) in 18 varieties of domestic soybean (G. max) and in 3 population of wild soybean (G. soja) was carried out. The part of polymorphous loci (P), intraspecies genetic differentiation (genetic distances--DN) were higher in domestic plants in comparison with wild ones (P = 45%, 17%: DN = 0.038-0.269, 0.059-0129). The preferable polymorphism of loci, coding the enzymes of glycolysis and Kreb's cycle was revealed in wild species. Domestic soybean had more polymorphous enzyme loci, which did not participate in glucose metabolism in comparison with wild species. The presence of the specific part of the gene pool in ancestor species, which was involved in soybean domestication and forming of varieties was discussed. PMID- 10857207 TI - [The genetic interpretation of the electrophoregrams of the helianthin in F1 sunflower seeds]. AB - It has been shown, that in most cases the fertile plants of maternal line and F1 plants of respective hybrid are present among sterile plants of sunflower female lines in crossing plots. As a result, 16 various genotypes of seeds are ascertained in crossing plots at monogenic differences in marker trait. Only two classes are F1 seeds. In such cases specific share of 5 genotype groups may be distinguished by phenotypes of heliantin electrophoregrammes. Seeds of biological admixture may be attributed to the 6th distinguishing type. PMID- 10857208 TI - [Seed proteins as markers in resolving the problems of genetic plant resources, selection and seed production]. AB - The article discusses the main directions and prospects of usage of protein and DNA markers in decision applied and theoretical problems of plant genetic resources (gene banks), plant breeding, seed production and seed testing. The examples of usage of protein and DNA markers for solving important problems of applied botany, genetics and plant breeding were considered. PMID- 10857209 TI - [The high-molecular glutenins of the soft winter wheats from European countries and their relationship to the glutenin composition of the ancient and modern wheat varieties of Ukraine]. AB - The sources of high-quality components of HMW glutenines determining grain quality, as initial material for breeding in the conditions of Ukraine were revealed on the base of analysis of 75 literature sources data about composition of high-molecular weight (HMW) glutenin and pedigrees of 598 European wheats from 12 countries, bred in 1923-1997, including, 449 cultivars from West and 149 East Europe. Origin of these components was observed in varieties of Great Britain, France and Germany from ancient Ukrainian wheat Red Fife and it derivative spring wheats of Canada--Marquis, Garnet, Regent, Saunders, Selkirk and of USA--spring wheat Thatcher and winter wheats--Kanred and Oro--as directly as via cultivars of European countries and Australia; in wheats of East European countries from winter wheats Myronivs'ka 808 and Bezostaya 1 (derivative of Ukrainian cultivars Ukrainka and Krymka) and their descendants; in wheats of Austria and Italy--from the both genetical sources. PMID- 10857210 TI - Approval of mifepristone (RU 486) in Europe. AB - The discovery of mifepristone (ex-RU486) was the achievement of a large research programme on steroidal compounds with antihormone properties conducted at Roussel Uclaf in partnership with the INSERM unit of Pr. E. E. Baulieu. Mifepristone exhibited a strong affinity to the progesterone and the glucocorticoid receptors. Consequently, it exerted competitive antagonism to these hormones both in in vitro and animal experiments. The identification of the antiprogesterone activity led to propose mifepristone use for the termination of early human pregnancy. Mifepristone at the dose of 600 mg initially used alone, was then used with a subsequent low dose of prostaglandins that led to a success rate of 95% as a medical method for early termination of pregnancy. In parallel, other indications were extended to the cervical dilatation prior to surgical termination of pregnancy in the first trimester, in the therapeutic termination of pregnancy for medical reasons beyond the first trimester, and for labour induction in case of fetal death in utero. The efficacy and safety of that treatment have been confirmed on the basis of more than 10 years of use with close adherence to the approved recommendations. Besides the political and philosophical hurdles that delayed clinical research with this molecule, other potential indications either in gynecological or cancer areas should be further developed. PMID- 10857211 TI - [Surgical procedure after primary chemotherapy of breast carcinoma--an unresolved clinical problem]. AB - Neoadjuvant chemotherapy has more and more become clinical routine during the past years. Results from large randomized trials like NSABP-B18 show that survival parameters are not affected if sequence of therapy is changed. Survival parameters have been intensively studied, but surgical standards after primary chemotherapy are much less well defined. Results from the early trials comparing lumpectomy or quadrantectomy with mastectomy are generally transposed to the neoadjuvant situation. In this context the "result of downstaging" is surgically treated like otherwise the primary tumor would have been treated. Though local recurrence rates reported after primary chemotherapy are not increased within the whole population this may not be correct for subgroups. E.g. within the NSABP-B18 trial significantly higher local recurrence rates are reported for those patients who initially were proposed to have mastectomy and who actually received lumpectomy after effective primary chemotherapy. Another unresolved problem is surgery after complete remission, which as histopathology demonstrates corresponds often not to pathological complete remission. Therefore in most cases the initially involved area is resected, which may result in a more radical surgical approach to complete remission than to partial remission. Further standardisation of surgical approach to patients after neoadjuvant chemotherapy should be evaluated within phase III trials. PMID- 10857212 TI - [Anthracyclines in therapy of ovarian carcinoma: a systematic review of primary and 2nd-line therapy after platinum]. AB - Despite being the subject of clinical research for more than 20 years, the role of anthracyclines in the treatment of ovarian cancer is still undefined. This review summarizes and re-evaluates the published data with the anthracylines doxorubicin, epirubicin (4'-epi-doxorubicin), daunorubicin (idarubicin), aclarubicin, and pirarubicin. The studies were identified by MEDLINE-search and analysis of the references cited in the identified articles and reviews covering this subject. First-line therapy: Studies from the 70ies could demonstrate that doxorubicin was as effective as alkylating agents. The combination of anthracylines and alkylating agents provided superior results compared with alkylating agents alone. Further improvements were gained by the introduction of platinum-anthracyline combination regimens. Doxorubicin and epirubicin showed comparable activity when combined with platinum. The comparison between platinum anthracyline containing combination regimens with platinum based regimens without anthracylines did not provide definitive answers. The majority of studies (with relatively small patient numbers) did not reveal any significant benefit for the anthracyline containing regimens. In contrast, four meta-analyses combining several studies showed a significant improvement when anthracylines were added to platinum based regimens. This question came up again when standard treatment for ovarian cancer was modified by the introduction of paclitaxel in the mid 90ies. Currently, two randomized large intergroup trials evaluate the role of anthracyclines in combination with platinum and paclitaxel. Results from these studies will be available in 1-2 years and might help to describe the role for anthracyclines in ovarian cancer treatment more precisely. Until then, the use of anthracyclines in upfront chemotherapy of ovarian cancer outside of clinical trials cannot be recommended. 2n-line therapy: The anthracyclines doxorubicin and epirubicin are among the most active single agents for the treatment of platinum refractory ovarian cancer. Both agents achieved tumor responses in up to 25%. Preliminary data suggest, that the combination with paclitaxel could increase efficacy. No such effect was reported for any other combination. In contrast, single agent anthracyclines cannot be recommended for treatment of platinum sensitive tumors. However, combining anthracyclines with platinum resulted in remarkable response rates of more than 50% of patients with platinum sensitive tumors. Only few studies report the use of anthracylines after platinum and paclitaxel containing first-line chemotherapy. Preliminary data from small studies (40 patients) suggest that anthracyclines showed activity in this particular population. No final conclusions can be drawn. There is a need for developing effective 2nd-line therapies for patients failing platinum and paclitaxel combination regimens. Among others, anthracyclines should be evaluated in this clinical setting. PMID- 10857213 TI - [Long-term side-effects following cyproterone acetate containing therapy in gynecology]. AB - It has been suggested that cyproterone acetate (CPA) has a mutagenic potency. It has been postulated that a threshold dosage of CPA has mutagenic effects, but in the same way data have been published documenting that a continuous low dosage of cyproterone acetate leads to a reduction of mutagenic episodes. Despite published data about higher levels of DNA adduct creations due to CPA an international multicentre study analysing 2,506 patients with 7,971 patient-years that used CPA could not find any liver cell cancers, even if due to epidemiological data 6 liver cell cancers should have occurred upon this study group. The present study deals with the evaluation of 57 women which received CPA in combination with EE2 11-13 years before. The daily dosage was 2 mg CPA in combination with 35 mg or 50 mg EE2. In Germany these drugs were registered under the name of Diane 35 or Diane 50. Long-term follow-up evaluation concerning side effects, especially the appearance of liver cell carcinomas, were the aim of this study. With the records of 32% (18/57) of the above mentioned patient group the following long-term follow-up side effects could be observed: 1) weight gain, 2) headache, 3) migraine, 4) gastrointestinal disorders, 5) mood affections/depressions, 6) oedema of the legs, 7) skin affections, 8) mastodynia. No benign liver tumor or liver cell carcinoma was detected upon our group of investigated patients. In conclusion we can affirm that the use of CPA in a dosage of 2 mg per day does not lead to serious side effects under long-term follow-up observation conditions and that it's use does not correlate with a higher appearance of liver cell carcinomas. PMID- 10857214 TI - [Clinical behavior of serous and mucinous borderline tumors of the ovary with diploid DNA stem line. Follow-up studies after organ preserving and non-organ preserving therapy]. AB - Even today, the situation is still unclear with regard to the radicality of the operation and any adjuvant therapy required in treatment of borderline tumors of the ovary. In the last two decades, treatment of these tumors in the Department of Gynecology and Obstetrics at our hospital has depended on the stage of the disease and the age of the patient. It ranged from laparoscopic cyst extirpation to hysterectomy with bilateral adnectomy and omental resection and regional lymphnodectomy. From the end of 1985 to the end of 1992, 35 patients with borderline tumors of the ovaries were operated on. Histologically, 14 borderline tumors were mucinous, 21 were serous. A follow-up investigation was carried out five to 11 years after primary operation. In this period, no patient died of borderline tumor. Twenty-eight patients who were followed up were clinically free of recurrence, five patients are alive, but could not be followed up. Two patients have died of other diseases after five years. Only one patient received adjuvant chemotherapy. She could not be followed up. In the meantime, peritoneal implants were demonstrated at second-look laparoscopies in four out of 18 patients. Later, these could no longer be demonstrated (one patient) or did not affect the survival time (three patients) and thus ultimately was not pathologically relevant. Primarily, two of these four patient had stage Ia-Ic. The paraffin blocks of the preparations are still available from 26 patients, and additional investigations could be carried out. Nineteen percent of the borderline tumors showed micropapillary structures of an MPCS (= micropapillary serous carcinoma), which evidently did not have a negative effect on the prognosis. All borderline tumors showed diploid distributions in DNA cytometry. It is not possible to make a definitive treatment recommendation on the basis of this investigation because the number of patients followed up was too small. PMID- 10857215 TI - [The Menopause Rating Scale (MRS II): methodological standardization in the German population]. AB - The Menopause-Rating-Scale (MRS I) was used in clinical practice since 1992. The physician had to document the severity of 10 important estrogen-related climacteric symptoms. Practical experience and a critical methodological evaluation justified a revision: The MRS I should be converted into a self administrative rating scale, the wording somewhat optimized, the layout adjusted and one item added. The standardization of the new scale (MRS II) was performed in a representative sample of the German population aged 45-60 years. Three dimensions were extracted from the menopausal symptoms using multivariate statistical techniques: somato-vegetative, psychological, and urogenital complexes of symptoms. A simple evaluation scheme was developed for the MRS II by summing up scoring points. Reference values for the frequency of 4 levels of intensity of complaints in the population were defined and provided for purposes of comparison. The MRS II meets a high methodological standard as an instrument standardized in the population. Moreover, it is convenient to apply this instrument in daily practice in order to quantitate variation of menopausal complaints. PMID- 10857216 TI - [Testicular feminization--sequelae of chronic neglect of a neoplastic process]. AB - This paper reports on a 56-year-old patient with a history of "testicular feminization syndrome" who was admitted to hospital because of a rapid gain in her abdominal girth (106 cm). A benign cyst was removed laparotomically which, histologically, was equivalent to a cystadenofibroma. Based on this case report, the clinical significance of testicular feminization syndrome and the necessity for gonadectomy due to the risk of growth of a possibly malignant tumor are discussed. PMID- 10857217 TI - [Clinical data protection and the internet]. AB - The benefit of using the internet in health care is growing: Information retrieval, communication, and telemedical cooperation support the physician and serve the patient. On the other hand connecting a hospital to the internet leads to considerable security problems, exposes sensible data to danger, and corrupts the professional discretion, unless the connection is installed and managed with extreme care. The state of the art for an internet connection is a firewall system. Moreover the security in a networked environment depends on a cryptographic infrastructure. This will soon be provided in Germany by Health Professional Cards. PMID- 10857218 TI - [Selection of a hospital information system--experiences and critical aspects]. AB - Due to typical problems (heterogeneity, lack of clinical functionality, Y2K problems) the board of directors of the university hospital of Marburg decided in 1997 to replace major components of the existing system by commercially available software. The products available on the market were analyzed, and, under participation of different user groups, a comprehensive functional specification was generated. This was the basis for a Europe-wide vendor selection process. In this context, several key aspects were identified, which are critically important for realizing a HIS that fulfills the specified functional requirements. Among these key aspects are the integration of heterogeneous system components, the support of cross-departmental workflow, and flexibility as well as adaptability to specific clinical requirements. As a result, we found that with today's commercially available products and standards there is no single solution that fully meets all requirements. However, some "generalist" vendors are offering integrated systems with acceptable clinical functionality. Tools are emerging which enable the clinical user to generate forms for data input and data flow. Still, a hospital information system will consist of separate components that have to be integrated, but the role of integrated, component-based approaches is becoming more important. PMID- 10857219 TI - Effects of joint incongruity on articular pressure distribution and subchondral bone remodeling. PMID- 10857220 TI - Function and regulation of temperature-inducible bacterial proteins on the cellular metabolism. AB - Temperature is an important environmental factor which, when altered, requires adaptive responses from bacterial cells. While a sudden increase in the growth temperature induces a heat shock response, a decrease results in a cold shock response. Both responses involve a transient increase in a set of genes called heat and cold shock genes, respectively, and the transient enhanced synthesis of their proteins allows the stressed cells to adapt to the new situation. A sudden increase in the growth temperature results in the unfolding of proteins, and hydrophobic amino acid residues normally buried within the interior of the proteins become exposed on their surface. Via these hydrophobic residues which often form hydrophobic surfaces proteins can interact and form aggregates which may become life-threatening. Here, molecular chaperones bind to these exposed hydrophobic surfaces to prevent the formation of protein aggregates. Some chaperones, the foldases, allow refolding of these denatured proteins into their native conformation, while ATP-dependent proteases degrade these non-native proteins which fail to fold. Most chaperones and energy-dependent proteases are heat shock proteins, and their genes are either regulated by alternate sigma factors or by repressors. The cold shock response evokes two major threats to the cells, namely a drastic reduction in membrane fluidity and a transient complete stop of translation at least in E. coli. Membrane fluidity is restored by increasing the amount of unsaturated fatty acids and translation resumes after adaptation of the ribosomes to cold. Neither an alternative sigma factor nor a repressor seems to be involved in the regulation of the cold shock genes in E. coli, the only species studied so far in this respect. PMID- 10857221 TI - Particle stress in bioreactors. AB - In many biological processes, e.g. the fermentation of cells and sensitive microorganisms or bioconversion with immobilised enzymes, low shear stress is of crucial importance for the optimal course of processes. Starting with the causes of particle stress, the following report discussed the hydrodynamic principles of the most frequently used model reactors and bioreactors, which are required for an approximate calculation of stress. The main part of the report describes the results of systematic investigations into the hydrodynamic stress on particles in stirred tanks, reactors with dominating boundary-layer flow, shake flasks, viscosimeters, bubble columns and gas-operated loop reactors. These results for model and biological particle systems permit fundamental conclusions on particle stress and the dimensions and selection of suitable bioreactors according to the criterion of particle stress. PMID- 10857222 TI - The engineering effects of fluids flow on freely suspended biological macro materials and macromolecules. AB - The manufacture of many biotechnologically important products requires consideration of the physical breakage and biochemical degradation pathways at all stages during processing, storage and transportation. The engineering flow environment in most items of bioprocess equipment has long been recognised as a key factor in determining these pathways and is the focus of the present review. Because of its industrial significance, the detrimental effects of the engineering flow environment on freely suspended bioparticles have been the subject of many scientific investigations over the past few decades. There is a general consensus of opinion that fluid shear and elongational stresses are the two main breakage pathways of relevance to processing of most biomaterials. An additional degradation pathway has also been identified involving significant losses of biological activity of macromolecules at gas-liquid, gas-solid and liquid-liquid interfaces. In such cases, the engineering flow field is shown to have a secondary role in determining the kinetics of inactivation. An equally important consideration in the optimisation of the relevant unit operations is the biomechanical integrity of the flow sensitive material. The biomechanical and biorheological parameters that determine the integrity of biomaterials are poorly defined, their evaluations present future research challenges and are of immediate engineering significance. PMID- 10857223 TI - Influence of stress on adherent cells. AB - Stress is a broad term often used with animal cells. Frequently mechanical forces are meant using this term but chemical stress is also important cultivating animal cells. The chemical environment of the cell in a reactor have to be considered very carefully. The complexity of the medium requirements and the metabolic pathway cause very often growth limitations. Studying these limitations in order to find the reasons showed to be difficulty because of the complexity of the system. Nevertheless, glucose, glutamine, lactate and ammonia are found to be critical parameter as well as the osmotic pressure. The influence of mechanical forces on cell viability is of great importance when growing the cells in agitated systems. By far the greatest amount of work reported in the literature has been done on suspension cells but adherent cells also experience shear forces not only in bioreactors also in vivo. Therefore, most research has be done on endothelial cells but studies exists done on non-endothelial cells. The influence of shear forces on cell growth, morphology and productivity will be discussed as well as possibilities of making the cells more resistant. PMID- 10857224 TI - Effects of hydrodynamic and interfacial forces on plant cell suspension systems. AB - Plant cells are perceived to be sensitive to the hydrodynamic environment in conventional bioreactors. Heightened sensitivity, relative to most bacterial cultures, is frequently attributed to larger plant cell sizes, extensive vacuolization and aggregation patterns. Early studies of shear sensitivity focused on cell lysis and/or loss of viability. More recently, an extensive array of sub-lethal responses has been identified. A fuller understanding of these sub lytic effects may assist in the optimization of large-scale cultivation conditions. This paper reviews recent work on the hydrodynamic shear sensitivity of plant cell systems, under cultivation conditions and in purpose-built shearing devices. The relevance of different approaches to the characterization of the intensity of a given hydrodynamic environment is discussed. Indicators of cell response to hydrodynamic stress are evaluated. The potential significance of cellular defense mechanisms, observed in response to mechanical stimulants, is identified. PMID- 10857225 TI - [Do we use reasonably macrolides in primary care?]. PMID- 10857226 TI - [Benefits of treatment to eradicate Helicobacter pylori infection in patients with ulcer at a primary care center]. AB - OBJECTIVES: To evaluate the clinical evolution and the use of Primary Care health resources one year after treatment to eradicate Helicobacter pylori (Hp) infection in patients with peptic ulcers and Hp infection. DESIGN: Retrospective study on the effect of an intervention. SETTING: Urban, reformed primary care centre. PATIENTS: 102 patients with peptic ulcers and Hp infection. INTERVENTION: Treatment to eradicate Hp. MEASUREMENTS AND MAIN RESULTS: a) Total medical attendance; b) attendance for dyspepsia; c) number of ulcerous outbreaks; d) medicines taken to treat dyspepsia. 79.4% of the patients treated were male. Overall mean age was 47.8 +/- 12.4. After the intervention, total attendance (from 8.3 to 6.6, p < 0.001), attendance for dyspepsia (from 3.1 to 1.1, p < 0.00001), and ulcerous outbreaks (from 1.2 to 0.06, p < 0.00001) all dropped sharply. The mean number of medicines prescribed for dyspepsia per patient fell from 1.24 to 0.43, p < 0.0001. Ranitidine prescription fell from 72.7 to 13.8 days (p < 0.001); and omeprazol from 35.1 to 12.2 days (p < 0.03). Estimated total saving per patient was 26,792 pesetas at 1998 values. CONCLUSIONS: Treatment in primary care to eradicate Hp(+) in ulcerous patients reduced the needs of attendance and the prescription of drugs for ulcers. Just in the first year this supposed a clinical benefit for these patients and important economic savings for the public health service. PMID- 10857227 TI - [Effectiveness of preventive activities analyzed within the framework of health centers involved in the Program for Preventive Activities and Health Promotion (PAPPS) of the semFYC. Group for the Evaluation of PAPPS]. AB - OBJECTIVE: To find the effectiveness of a programme of preventive activities for adults (hypertension, tobacco and alcohol) measured by the number of cases identified, the evaluation of the initial interventions and the degree of control over the identified factors. DESIGN: Retrospective study: review of clinical records. SETTING: Primary care. Multi-centre study: health centres from the whole of Spain. PATIENTS: 7562 clinical records of patients over 20, who participated voluntarily and were extracted by systematic sampling from 378 care units (doctor and nurse) at 85 health centres committed to the Programme of Preventive Activities and Health Promotion (PAPPS) of the Spanish Society of Family and Community Medicine (semFYC). The study period was from May 1 1995 to April 30 1997. MEASUREMENTS AND MAIN RESULTS: Age, sex, data on tobacco habit, alcohol consumption and blood pressure were obtained through a questionnaire (recording the actions taken, date of diagnosis, initial assessment and subsequent monitoring). 28.3% were tobacco-dependent, 6.9% consumed too much alcohol, and 22.2% had hypertension. About 20% cases of each risk factor were detected during the study period. Giving up tobacco was recorded in 7.6% of smokers, and giving up alcohol in 19.7% of excess drinkers. 78.6% of hyperintense patients had acceptable-optimum control. CONCLUSIONS: The number of cases detected shows that the PAPPS programme performs acceptably. The tobacco and drink given up and the hypertension control attained due to the intervention are similar to the published trials. The PAPPS is an effective programme for controlling risk factors in primary care. PMID- 10857228 TI - [Antibodies against Helicobacter pylori in saliva. Study of their validity versus breath test and its agreement with serology]. AB - OBJECTIVES: To evaluate prospectively the validity of a new diagnostic method based on a saliva sample, taking as reference the breath test with 13C-marked urea, and to compare the results of this technique with another indirect method based on the detection of antibodies, "classical" serology using venous blood. METHODS: 48 individuals, 24 healthy volunteers and 24 consecutive patients with gastro-duodenal ulcer disease, were studied prospectively. Treatment during the previous month with gastro-erosive medication, antibiotics, proton-pump inhibitors or bismuth-derived drugs, prior treatment to eradicate H. pylori, gastric surgery and the presence of linked illnesses, were all considered criteria of exclusion from the study. For the diagnostic test in saliva a commercial enzyme-linked immunosorbent assay (ELISA, trademark Helisal) was used; and for blood serology, another commercial ELISA (Helico-G). The staff responsible for reading the saliva, serology and breath tests did not know the result of the other diagnostic methods. The result of the breath test with 13C urea (TAU-kit) was taken as the reference standard for H. pylori infection. RESULTS: The mean age of the healthy volunteers was 23 +/- 0.7 years; and of ulcer patients, 55 +/- 18. The prevalence of H. pylori infection, valued by the gold standard, was 79.2% in the ulcer patients and 54% in the volunteers. The sensitivity, specificity, positive predictive value, negative predictive value and diagnostic accuracy of the saliva test in the ulcer patients were, respectively: 100% (95% CI, 79-99), 60% (17-93), 90% (68-98), 100% (31-97) and 92% (71-98). In the volunteers these figures were: 46% (20-74), 73% (39-93), 67% (31-91), 53% (27-78) and 58% (37-77). The serology results were better, with 100% sensitivity in both groups and outstanding diagnostic accuracy (92% and 96% for ulcer patients and volunteers, respectively). Concordance between serology and the saliva test in ulcer patients was perfect (kappa, 1). However, in asymptomatic individuals concordance was deficient (kappa, 0.28), and the prevalence of infection diagnosed with the two tests was not homogeneous (McNemar, 2.8; p < 0.05). CONCLUSION: The diagnostic test for H. pylori infection in saliva is lacking in diagnostic accuracy in healthy individuals, which indicates that it cannot be used for screening infection in the asymptomatic population. The technique is more valuable in ulcer patients, although it does not reach the specificity desirable. For these reasons, the saliva test evaluated in this study cannot be recommended for diagnosis of H. pylori infection. PMID- 10857229 TI - [How many patients need treatment for hypercholesterolemia?]. AB - OBJECTIVE: To find out the percentage of individuals from a population who need pharmacologic treatment for their hypercholesterolemia according to different guidelines. DESIGN: Descriptive transversal study. SETTING: Population from 6 areas of Lleida (province) including the city of Lleida. METHOD: The study has been done on a randomized sample of 401 individuals. First the cardiovascular risk from the equation in the Framingham study was calculated, then the percentage of individuals who should be treated with lipid-lowering medication according to the US National Cholesterol Education Program, The European Society of Atherosclerosis and The Spanish Society of Atherosclerosis. RESULTS: The prevalence of individuals with cholesterolemia > 250 mg/dl has been 16.3% among men and 22.4% in women. According to NCEP, 20.5% of men and 17.6% of women from 383 individuals older then 6 years old need pharmacologic treatment for hypercholesterolemia. The Framingham equation was applied to 281 individuals (over 30 and under 75 years old) estimating a risk higher than 20% in 10 years for 13.7% of the population under study (23.9% in men and 3.5% in women). CONCLUSIONS: The percentage of patients who need pharmacologic treatment for hypercholesterolemia varies according to the different guidelines employed. We consider necessary the calculation of the cardiovascular risk with the Framingham equation to use lipid-lowering medication in order to achieve a better protection of the population at higher risk of developing a cardiovascular disease. PMID- 10857230 TI - [Prevalence of insomnia and use of psychodrugs among elderly in a basic health area in Cuenca]. AB - OBJECTIVES: Insomnia is the most common sleeping disorder, increasing as people get older, which therefore creates an increase in the use of hypnotics. The presence of insomnia in elderly people, according to different authors, ranges between 17 and 43% depending on the criteria of diagnosis used and the group of population studied. The objectives of this study are to determine the prevalence of insomnia in a population of 65 years and over in a Basic Health Area and the medical consumption related to it. DESIGN: A cross sectional study by means of an ad hoc questionnaire about sleeping habits given by medical staff, including social demographic variables, psychotropic medication consumption, cognitive assessment by means of Mini Mental Status Examination and a range of anxiety depression of Goldberg. Hartman and DSM-IV criteria were used for the insomnia diagnosis. SETTING: Cuenca I Primary Care Center, Cuenca (Spain). PATIENTS: A random sample of 343 patients of a population of 2253, 65 years and over. MEASUREMENTS AND MAIN RESULTS: The prevalence found was 13.6% (Hartman) and 30.7% (DSM-IV) more common amongst women (p < 0.005), sufferers of psychiatric illnesses (p < 0.01) and those at the top-half of the anxiety-depression scale (p < 0.001). A 46.1% suffer from daylight hypersomniac. A 19.1% takes some kind of medication to help them sleep and the 74.6% of them take it daily. Long and short plasma half-life benzodiazepines are the most consumed, with women and insomniacs being the majority consumers. CONCLUSIONS: The prevalence of insomnia in our population is slightly inferior to that of other studies and the consumption of sleep-enhancing medication although inappropriate is similar to that referred to in literature. PMID- 10857231 TI - [Chronic complications of type 2 diabetes mellitus. Clinical course after 5 years of follow-up]. AB - OBJECTIVE: To evaluate the modifications in the prevalence of complications and the incidence of end-points in diabetic patients observed for five years, and the effectiveness of a diabetes care protocol on the process indicators. DESIGN: Prospective observational study between 1991 and 1996. SETTING: Primary care centre. PARTICIPANTS: Diabetic patients monitored between 1991 and 1996. MEASUREMENTS AND MAIN RESULTS: Social and demographic variables, DM epidemiology variables, cardiovascular risk factors, complications and end-points were measured. 318 of the 352 patients selected in 1991 were followed. Average age was 68.6 (SD 11.2) and 39% were male. Mean observance was for 53 months (SD 10). There was an increase of insulin use (19%) and self-analysis (34.7%) both in men (chi 2 = 14.7, p < 0.001) and in women (chi 2 = 40.5, p < 0.001), independently of age (chi 2 = 37.77, p < 0.001). Complications increased: microvascular ones from 33.4% to 42.1%, macrovascular ones from 22.3% to 37.2%. The most common end points were CVA (7.8%) and angor (3.6%). 22 patients died (6.9%), with ischaemic cardiopathy (30%) and neoplasm (30%) the most common causes. Hypertension increased from 51.6% to 59.8% and hypercholesterolaemia from 42.6% to 47%. Obesity (42%) and tobacco dependency (28%) remained stable. Systolic blood pressure went down by 4.7 mmHg and diastolic pressure by 3.76 mmHg, and in women, over-65s and those who had had the illness longest (> 10 years). 6.5% (CI 1 12.9%) of patients improved their blood pressure (< 135/85 mmHg). HbA1c worsened independently of sex, age or years of evolution of diabetes. PMID- 10857232 TI - [Temporal disability caused by psychiatric pathology at a health center]. AB - OBJECTIVE: To find the incidence, duration, diagnosis and treatment of time off work for psychiatric reasons. DESIGN: Retrospective, descriptive study. SETTING: The Dr. Pujol i Capsada Health Centre in Baix Llobregat county, Barcelona. PARTICIPANTS: All the records of time off occurring between January 1992 and December 1996 were reviewed. Time off for psychiatric reasons, and its length, follow-up, relapses and treatment were analysed. MEASUREMENTS AND MAIN RESULTS: 71 patients had had time off on some occasion for psychiatric reasons (34% men, 66% women). The mean age was 38 (SD 10). The mean number of days off in the first period of time off (n = 71) was 155 days (SD = 302), in the second (n = 21) 209 (SD = 268), in the third (n = 8) 187 (SD = 327), and in the fourth (n = 3) 206 days (SD = 326). Total number of days off came to 16,894. 4% obtained invalidity status for psychiatric reasons. The most common initial reasons for the first period of time off were, by pathology: psychiatric (58%), osteo-muscular (17%), nervous system and sense organs (7%) and digestive system (6%). The psychiatric reasons for the first period off were: anxiety (48%), depression (38%), anxiety depression (10%), schizoid disorders (3%) and personality disorders (1%). Half the patients were referred and almost all were supervised by the general practitioner. The most commonly used drugs were the benzodiazepines (75%) and serotonin re-uptake inhibitors (38%). CONCLUSIONS: Time off for psychiatric reasons tends to last a long time and to recur. 42% of these periods of time off was not initially classified as for psychiatric reasons. Half the patients are shared and supervised jointly with the psychiatrist: thus, the need for good coordination between the two areas. PMID- 10857233 TI - [Consensus on care for the users of the Maintenance Program with Methadone at the health centers of the province of Cadiz]. AB - OBJECTIVES: To reach a consensus on primary care actions concerning transmissible illnesses in users of the methadone maintenance programme. To promote coordination between levels. To find the needs of primary care professionals. DESIGN: Qualitative study based on the nominal group technique. SETTING: Cadiz province, including primary care districts, NHS hospitals and county drug dependency centres. PARTICIPANTS: 76 health professionals: two doctors and a nurse from each health centre, selected for their experience and motivation; hospital specialists in preventive medicine and internal medicine also with experience and motivation, and with knowledge of primary care; a doctor from each county drug-dependency centre, chosen by the Provincial Drug-Dependency Management. INTERVENTIONS: The study had two phases: a) nominal group technique with homogeneous groups for each professional environment; b) modified nominal group with heterogeneous groups of professionals from all environments by hospital areas. MEASUREMENTS AND MAIN RESULTS: The final result of the study was the overall consensus in the four areas. The prioritized activities focused on "recruitment and study of patients and contacts", "educational and health promotion interventions", "immunisation programme", "monitoring therapy compliance", and "unified record of actions". What was needed to put into practice the interventions was agreed by consensus. CONCLUSIONS: Consensus techniques are a useful tool for planning primary care activities. A high degree of reproducibility can be reached if the participants are properly selected. Restrictions in being able to introduce some of the agreed interventions could be avoided with user participation and involvement of the health authorities. PMID- 10857234 TI - [Diagnosis and treatment of postmenopausal osteoporosis after a Colles' fracture]. AB - OBJECTIVES: To investigate whether the presence of Colles fracture leads to diagnostic studies or treatment of osteoporosis in postmenopausal women. DESIGN: Retrospective study with follow-up of incident cases. SETTING: Hospital General de Vic and Primary Care Centre Osona. Vic. Barcelona. PARTICIPANTS AND METHODS: We studied 80 postmenopausal women with Colles' fracture during 1995-1996. The clinical records were systematically reviewed and treatment with antiosteoporotic drugs before and after Colles' fracture were compared. MEASUREMENTS AND MAIN RESULTS: Diagnostic studies were found in 3 (3.8%) patients. 6 patients (7.5%) took antiosteoporotic drugs before the fracture while 21 (26.3%, p < 0.001) did so after it. CONCLUSIONS: A recent Colles' fracture induces few osteoporosis diagnostic studies. However, it leads to a significant increase in the use of antiosteoporotic drugs. PMID- 10857235 TI - [Physiopathologic basis of obesity]. PMID- 10857236 TI - [How to manage our bibliography: development and maintenance of a personal bibliographic file]. PMID- 10857237 TI - [Bioethics and family medicine (IV)]. PMID- 10857238 TI - [Safety of the new quinolones]. PMID- 10857239 TI - [Cholestasis secondary to antitubercular treatment]. PMID- 10857240 TI - [Peripheral neuropathy and amiodarone]. PMID- 10857241 TI - [Morbidity of caregivers of patients in their homes]. PMID- 10857242 TI - [Unusual case of non-compliance of treatment]. PMID- 10857243 TI - Introduction to the special issue: transitions in substance use across time, gender, and culture. PMID- 10857244 TI - The genetics of smoking initiation and quantity smoked in Dutch adolescent and young adult twins. AB - Not much is known about the genetic and environmental determinants of various aspects of substance use in adolescents. This study examined whether the inheritance of initiation of tobacco use in adolescents is independent of the inheritance of the number of cigarettes smoked. Alternative multifactorial threshold models were applied to data on tobacco use in 1676 Dutch adolescent twin pairs. The three models that were considered are (i) the single liability dimension model, (ii) the independent liability dimension model, and (iii) the combined model (CM). The results showed that there is not one underlying continuum of liability to smoking. The CM was the best-fitting model. This model postulates that there are separate initiation and quantity dimensions but allows for the possibility that there are some individuals who are so low on the liability to level of consumption that they are not using tobacco. There were no differences between males and females in the magnitude of the genetic and environmental influences on individual differences in smoking initiation and quantity smoked. Smoking initiation was influenced by genetic factors (39%) and shared environmental influences (54%). Once smoking is initiated genetic factors determine to a large extent (86%) the quantity that is smoked. PMID- 10857245 TI - Genetic and social determinants of initiation and age at onset of smoking in Australian twins. AB - Retrospective data on age at onset of smoking, reported by 3810 adult Australian twin pairs, were analyzed to determine the role of genetic and environmental factors in the onset of smoking. Results of nonmetric multidimensional scaling supported a two-process model in which different etiologic factors determined which individuals were at risk of becoming smokers and the age at onset of smoking in those who were at risk. Parametric model-fitting confirmed this difference. For female twins and younger male twins (aged 30 years or less), the onset of smoking was strongly influenced by genetic factors, with shared and nonshared environmental effects having a more modest impact. For older male twins, shared environmental influences on onset of smoking were very important, and the influence of genetic predisposition was slight. The age at which smoking onset occurred, however, was influenced by both genetic and nonshared environmental effects, but not by shared environmental effects, in both sexes and both cohorts. PMID- 10857246 TI - A twin study of drinking and smoking onset and latencies from first use to regular use. AB - The early onset of alcohol and tobacco use has been associated with increased risk for later substance abuse and dependence problems. This study investigated genetic and environmental influences on age at onset of alcohol and tobacco use by examining twin resemblance for several retrospectively reported onset milestones including age at first use, age at first alcohol intoxication experience, and age at regular use. In addition, we also examined the latency between age at first use and age at regular use of tobacco and alcohol. The subjects were a volunteer sample of older adult twins 50 to 96 years of age. MZ twin correlations for age at first alcohol use and age at first tobacco use were .57 and .44, respectively, compared to .45 and .37 for DZ same-sex twins. MZ twins correlated .30 and .26 for the latencies between first use and regular use of alcohol and of tobacco, while DZ correlations were -.01 and .05, respectively. Biometrical model-fitting results confirmed that familial resemblance for age at first use for both alcohol and tobacco was largely the result of shared environmental factors, while the latencies between first use and regular patterns of use were more genetically influenced. These findings add to a growing body of literature suggesting that initiation of substance use is influenced primarily by environmental rather than genetic factors. PMID- 10857247 TI - The genetics of smoking persistence in men and women: a multicultural study. AB - Using a correlated liability dimensions model, we examined the extent to which the same genetic and environmental factors influence both initiation of regular cigarette smoking and maintenance of the smoking habit in men and women. We analyzed questionnaire survey data obtained from large samples of male and female like-sexed twins from three countries, Australia (N = 1535 pairs), Sweden (N = 5916 pairs), and Finland (N = 4438 pairs), subdivided into three age bands (18 25, 26-35, and 36-46 years of age). We found that familial influences on risk for persistence in smoking cannot be entirely explained by the same factors responsible for risk of smoking initiation. Total genetic variance for smoking persistence varied little by age band and sex (range, 39-49% in women and 42-45% in men); however, even among twins in the youngest group (18-25 years of age), who on average have the fewest years of cigarette use, less than 40% of the total genetic variance in smoking persistence was accounted for by the same genetic factors that increased risk of smoking initiation, and this percentage decreased to less than 10% in the 36-46 year olds. PMID- 10857248 TI - Searching for interactive effects in the etiology of early-onset substance use. AB - This study sought to expand the modest literature investigating gene x environment interactions in the prediction of substance use. Our sample consisted of 591 male twins from the Minnesota Twin Family Study. Their relative genetic risk was estimated from their parents' substance-related diagnoses and their environmental risk from their affiliations at age 11 with social groups likely to either encourage or discourage substance use. At age 14, the boys' own substance use was assessed. We hypothesized both main effects and an interaction between our genetic- and environmental-risk variables in the prediction of substance use by this young age. We further theorized that the boys' inherited risk might take the form of temperament, specifically externalizing tendencies. Using regression analyses and biometrical modeling, we corroborated earlier research by finding evidence for a significant interactive effect in the etiology of substance use. Our results suggest that low levels of environmental risk may buffer against the potentially unfavorable effects of high familial risk; however, when environmental risk is high, the degree of familial risk is consequential. We were not able to support our second hypothesis; rather, temperament predicted substance use only through shared environmental factors. PMID- 10857249 TI - The influence of religion on alcohol use initiation: evidence for genotype X environment interaction. AB - We examined the possible role of religious upbringing as a mediator of the shared environmental influences and as a moderator of the genetic influences on the risk of alcohol use initiation in a large population-based sample of Dutch adolescent and young adult twins (1967 twin pairs). There was not a significant association between religious participation and alcohol use initiation among Dutch adolescents and young adults. We also hypothesized that the relative magnitude of the genetic influences on the risk of alcohol use initiation would be greater for those adolescents and young adults who were raised in a less religious environment compared to those adolescents and young adults who were raised in a more religious environment. We indeed found higher heritabilities for females without a religious upbringing compared to females with a religious upbringing. Genetic influences accounted for 40% of the variance in alcohol use initiation in nonreligious females, compared to 0% in religiously raised females. Shared environmental influences accounted for 54% of the variance for nonreligious females and 88% of the variance in religious females. For males, the genetic variance was also higher in the nonreligious group compared to the religious group, but this difference was not statistically significant. Whether or not they were raised religiously, the liability to alcohol use initiation in males was moderately influenced by genetic factors (30%) and substantially influenced by shared environmental factors (60%). PMID- 10857250 TI - Longitudinal analyses of the determinants of drinking and of drinking to intoxication in adolescent twins. AB - Genetic and environmental determinants of self-reported alcohol consumption were investigated in a sample of 2513 twin pairs who were first assessed at age 16 and were followed-up at age 17. At age 16, 77% of the sample was drinking, and 65% of drinkers reported drinking to intoxication. Both drinking and drinking to intoxication increased at the 1-year follow-up. Model fitting indicated that most of the variance in drinking initiation was due to shared environmental effects but that shared environmental effects were less important, and additive genetic effects were more important, in explaining frequency of drinking among subjects who had already initiated drinking. Similarly, shared environmental effects explained most of the variation in initiation of drinking to intoxication but were less important in explaining frequency of intoxication among subjects who had already initiated drinking to intoxication. The magnitude of genetic and environmental estimates for males and females did not differ significantly, but it was clear that either different genetic factors or different shared environmental factors were influencing males and females. For all drinking variables studied, shared environmental effects decreased from age 16 to age 17, while additive genetic effects increased from age 16 to age 17. PMID- 10857251 TI - An assessment of the genetic relationship between alcohol metabolism and alcoholism risk in Australian twins of European ancestry. AB - The present analyses examined genetic influences on alcohol metabolism and their possible relationship to risk of alcohol dependence. Subjects were 206 Australian twin pairs who participated in an alcohol challenge protocol in 1979-1981, in which they were given a 0.75 g/kg dose of alcohol; blood alcohol concentrations (BACs) measured at five times over a 3-hr period after alcohol ingestion were examined. Structural equation modeling, fitting a combined autoregressive and common factor model, indicated significant heritabilities for both men and women (h2 range = 0.19-0.71), with significant parameter heterogeneity as a function of gender. In 1992-1993, both twins from 159 of the alcohol challenge pairs completed a telephone-administered psychiatric diagnostic interview. Repeated measures MANOVAs were used to examine whether respondent's or cotwin's DSM-III-R alcohol dependence status, or parental history of alcohol problems, was associated with variation in alcohol metabolism. There was some evidence that individuals at increased genetic risk of alcohol dependence [with either a paternal history of alcohol problems (women) or an MZ male cotwin who reported a history of alcohol dependence by 1992-1993] showed lower initial BACs than other groups. However, this effect was not seen in those who themselves had a history of alcohol dependence by interview follow-up, perhaps because this relationship was already masked by a history of excessive drinking at baseline. PMID- 10857252 TI - Genetic and environmental influences on transitions in drug use. AB - Genetic and environmental factors influence drug abuse, but abuse represents the culmination of a sequence of events. Different levels of use may have different determinants and these determinants may differ across drug types. Approximately 3200 male-male twin pairs from the Vietnam Era Twin Registry were interviewed by telephone. Data were obtained regarding exposure to six categories of illicit drugs, initiation of use, continuation of use, regular usage, and diagnosis of drug abuse/dependence. Genetic, common environmental, and unique environmental influences on transitions of drug involvement, defined as movement from one level of drug use to the next, were investigated. Marijuana had the highest conditional probability for the transition from exposure to use, from use to use more than five times, and from use more than five times to regular use. The rate of transition to regular use of heroin was higher than the rate for amphetamine, cocaine, sedatives, and psychedelics. Cocaine had the highest conditional probability for the transition from regular use to abuse/dependence. Significant genetic influences were observed for a number of transitions in marijuana, amphetamine, and cocaine usage. PMID- 10857253 TI - Male and female gametophyte selection of barley for salt tolerance. AB - The contribution of the gametophyte in the inheritance of salt tolerance was studied by crossing F3 and BC1 hybrids of the tolerant cultivars Rannii 1 and Pirkka with the sensitive cultivar K-30356. The third generation shows that the progenies of heterozygous plants grown in saline conditions show elevated salinity tolerance. Again, a comparison of the BC1 hybrid progenies shows that the male and female gametophytes contribute to the inheritance of salt tolerance. Gametic selection is maximally efficient during the formation of the female gametophyte and the germination of pollen grains on the stigma. PMID- 10857254 TI - Assessing the allozyme variation in cultivars and Swedish landraces of rye (Secale cereale L.). AB - Genetic interpretation and diversity of 9 isozyme loci have been estimated in 7 improved varieties and 19 landraces from Sweden by means of starch gel electrophoresis. The isozyme systems were ACO, DIA, GPI, MDH, PGD and PGM. For the statistic analysis we used the following measures: average number of alleles per locus, percentage of polymorphic loci, average heterozygosity direct count and average heterozygosity Hardy-Weinberg expected unbiased estimate. The measures were made on species and population levels. The distribution of the total genetic diversity among populations was also calculated. To illustrate the genetic relationships among populations, genetic distances were measured and principal component analysis performed. As expected in a cross-pollinated crop we found high genetic diversity and a larger variation within than among the populations. Somewhat unexpectedly, however, we found that the currently used varieties have the same high level of heterozygosity as the landraces but in the dendrogram the two groups are separated. The dendrogram showed three main clusters. The large cluster included 21 populations and the two small clusters were clearly distinguishable from the rest. The landrace spring-type could not be separated from the landraces winter-type, but we did detect a difference between different spring types. A few populations had unique alleles for certain loci. PMID- 10857255 TI - Mating system parameters in species of genus Prosopis (Leguminosae). AB - The section Algarobia of genus Prosopis involves important natural resources in arid and semiarid regions of the world. Their rationale use requires a better knowledge of their biology, genetics and mating system. There are contradictory information about their mating system. Some authors claim they are protogynous and obligate outcrosser. However, some evidence have been shown indicating that they might not be protogynous and that they might be somewhat self-fertile. The current paper analyses genetic structure and mating system parameters in populations of seven species of this section from South and North America based on isozyme data. In all species a significant homozygote excess was found in the offspring population but not in mother plant genotypes. Multilocus and mean single locus outcrossing rates (tm, ts) indicated that about 15% selfing can occur in the studied populations. The heterogeneity between pollen and ovule allele frequencies was low suggesting population structuration, in agreement with the estimates of correlation of tm within progeny (rt) and correlation of outcrossed paternity (rp). The difference of FIS estimates between offspring and mother plants suggest some selection favouring heterozygotes between seedling and adult stages. PMID- 10857256 TI - Chromosomal location of powdery mildew resistance genes and cytogenetic analysis of meiosis in common wheat cultivar Meri. AB - Common wheat cv. Meri was crossed to a set of 21 Chinese Spring monosomic lines to characterize resistance to powdery mildew and to determine the chromosomal location of the gene(s). Monosomic F1 plants were allowed to self-pollinate and to produce F2 seeds. Seedlings of F2 and F3 plants and their parents were inoculated with isolates Ns 2 and 9 of Erysiphe graminis f. sp. tritici. Analysis of obtained data revealed that one major dominant gene conferring resistance is located on chromosome 1B of cv. Meri. The new gene is designated by symbol Pm28. On the basis of the trivalent configuration frequency (without univalent) at the 1st metaphase of meiosis it was found that two reciprocal translocations involving chromosomes 2A/5A and 5B/5D differentiate cv. Meri from cv. Chinese Spring. In the F1 monosomic hybrids, genes causing a decrease in pairing are found on chromosomes 4D and 6D, and genes enhancing pairing--on chromosomes 3A and 7B. PMID- 10857257 TI - MtDNA polymorphism in the Hungarians: comparison to three other Finno-Ugric speaking populations. AB - Mitochondrial DNA sequence variation as well as restriction site polymorphisms were examined in 437 individuals from four Finno-Ugric-speaking populations. These included the Hungarians (Budapest region and the Csangos from Hungary and Romania), the Finns and two Saami groups from northeastern Finland (Inari Saami and Skolt Saami), and the Erzas from central Russia. The mtDNA data obtained in this study were combined with our previous data on Y chromosomal variation for eight different loci in these populations. The genetic variation observed among the Hungarians resembled closely that found in other European populations. The Hungarians could not be distinguished from the neighboring populations (e.g., the Austrians) any more than from their Finno-Ugric linguistic relatives. PMID- 10857258 TI - The chloroplast genomes of azuki bean and its close relatives: a deletion mutation found in weed azuki bean. AB - A physical map of the azuki bean (Vigna angularis cv. Erimo-shozu) chloroplast DNA (cpDNA) was constructed by localising the cleavage sites of PstI, SalI, SmaI, SacI, KpnI, PvuII and XhoI. The resulting map is more similar to the cpDNA-maps of two Vigna species (mung bean, V. radiata, and V. nakashimae) than to common bean (Phaseolus vulgaris) cpDNA map. Azuki bean was originally classified in the genus Phaseolus, and the inclusion of this taxon in the genus Vigna is a recent taxonomic decision. Our result is thus in favour of the taxonomic placement of azuki bean in the same genus as V. nakashimae and mung bean. We also found that a weed-form accession of azuki bean has a 96-bp deletion relative to the cultivar Erimo-shozu. The 96-bp deletion is located between the trnS-UGA and psbC genes in the large single-copy region of the chloroplast genome. This deletion is flanked by imperfect 9-bp direct repeats, suggesting that the deletion was a result of intra-molecular recombination mediated by these direct repeats. PMID- 10857259 TI - Robertsonian translocations and B chromosomes in the Wellington tree weta, Hemideina crassidens (Orthoptera: Anostostomatidae). AB - Two karyotypes within the species Hemideina crassidens are described, 2n = 15 (XO) and 2n = 19 (XO). These two karyotypes have a NF of 28. The 19-karyotype was found exclusively in the southern part of the species range and the 15-karyotype was found in the north. The differences between the two karyotypes are interpreted as arising from two Robertsonian translocations (fission/fusion). Laboratory matings between weta with the two karyotypes produced viable offspring. During meiosis in F1 intraspecific hybrids metacentric and acrocentric autosomes aligned to form two trivalents, confirming homologies predicted by Robertsonian translocations. The subspecies H. c. crassicruris, (confined to Stephens Island) was found to be polymorphic for a metacentric B chromosome. An unusual association of sex and presence of B chromosome was observed in this island population with Bs found only in male weta. PMID- 10857260 TI - Chromosomal abnormalities in hypoprolific boars. AB - Four new chromosomal rearrangements are reported in the domestic pig: 3 reciprocal translocations, rcp(4;12)(p13;q13) in a crossbred boar, rcp(1;7)(q17;q26) in a Large White purebred boar, rcp(1;6)(q17;q35) in a purebred synthetic paternal line boar, and a pericentric inversion inv(2)(p13q11) in a crossbred boar. The 1/7 reciprocal translocation and the pericentric inversion were detected in animals that had sired small litters. The effect of the 1/7 translocation was accurately determined: -4.5 piglets born per litter, i.e. -36%. Both the 1/6 and 1/7 reciprocal translocations were of maternal origin. All the chromosomal rearrangements were highlighted using GTG and/or RBG banding techniques. Chromosome painting experiments were also carried out to confirm the proposed hypotheses for the three reciprocal translocations. PMID- 10857261 TI - Number and sites of rDNA loci of Guizotia abyssinica (L. f.) Cass. as determined by fluorescence in situ hybridization. AB - Fluorescence in situ hybridization (FISH) was employed on somatic chromosome preparations of Guizotia abyssinica using a DNA probe of the 18S-5.8S-26S rRNA genes including the transcribed and non-transcribed spacer sequences. A maximum of six major and two minor signal sites of rDNA was observed. However, the minor signals were not persistently detected. The positions of the FISH signals coincided with the sites of the nucleolar organizer regions and their adjacent C banded heterochromatin when present. PMID- 10857262 TI - Genomic organisation of the neural sex determination gene fruitless (fru) in the Hawaiian species Drosophila silvestris and the conservation of the fru BTB protein-protein-binding domain throughout evolution. AB - We report the cloning and sequencing of the fru gene from the Hawaiian picture wing species Drosophila silvestris. The fru gene has seven exons spanning 15-kb encoding two transcripts with ORFs of 841 and 695 amino acids. The protein encoded by the fruA transcript is well conserved with the D. melanogaster type A protein, particularly the BTB protein-protein-binding domain, which is encoded by exons I and II and is 100% conserved. The peptide encoded by exon III has several sequence differences but these are confined mostly to regions of repetitive sequence and exons IV to VI are well conserved. The peptide encoded by exon VII is semi-conserved for the 5' end and 100% conserved for the Zinc finger domains; the rest of the peptide is virtually unconserved. The FRUA protein has a BTB domain and two zinc finger domains whereas the FRUC protein only has the BTB domain. The genomic DNA sequence encoding the BTB domain of the fru gene has been cloned from 21 species of Diptera. The protein-coding sequence is highly conserved and the amino acid sequence is identical except for two changes in the Tephritidae. The intron sequences are completely unconserved except between very closely related species such as the Hawaiian Drosophila. The phylogeny produced using the BTB exon sequences suggests that the most closely related mainland Drosophila species to the Hawaiian clade is D. moriwakii of the melanica species group. The phylogeny also shows that the Scaptomyza are closely related to the Hawaiian Drosophila so supporting a Hawaiian origin for the Scaptomyza. The genus Zaprionus is placed in the subgenus Drosophila closely related to D. immigrans along with the genera Samoaia and Liodrosophila. PMID- 10857263 TI - Karyotypic characterization by mitosis, meiosis and C-banding of Eriopis connexa Mulsant (Coccinellidae: Coleoptera: Polyphaga), a predator of insect pests. AB - Eriopis connexa presents a chromosome number of 2n = 18 + XX for most females analyzed and a meioformula of n = 9 + Xyp for all males. A small metacentric B chromosome restricted to females occurred in 10% of our sample and, when submitted to C-banding, it was shown to be almost completely euchromatic. Chromosome pairs 2 and 3 had satellites and probably contained the nucleolar organizer regions (NORs). C-band analysis also revealed that the constitutive heterochromatin was localized in the centromeres of all chromosomes in the complement. PMID- 10857264 TI - Magnetic brain stimulation--a new therapeutic tool in psychiatry. PMID- 10857265 TI - Siblings of schizophrenic patients--a review. AB - The families of patients with schizophrenia carry an enormous emotional and social burden. This article is a historical review of the study of the well siblings of schizophrenic individuals. The early investigations (1950s-1970s) were based on the theory of familial transmission of schizophrenia and focused on siblings from the aspect of their susceptibility to the disease. Many claimed that even siblings who did not develop full blown schizophrenia still suffered from considerable psychiatric disorders that were attributable to pathological familial psychodynamics. Case control studies were used to explain how some well siblings "escaped" the pathological family network. With the introduction of the concept of family burden in the late 1960s, research shifted to the emotional impact of growing up with a schizophrenic brother or sister. This was accompanied by the emergence of self-help groups and published case histories of siblings themselves. In the early 1980s, the data were essentially descriptive. Investigators studied sibling shame, poor self-esteem and feelings of stigmatization. Different patterns of coping with the subjective burden were distinguished. More recently, greater efforts have been made to systematically define the variables associated with the burden experienced by siblings. To answer the many still outstanding questions, further studies are needed based on a longitudinal design and homogeneous samples. For example: Are sibling's personal relationships outside the family affected? Does the sibling place within the family affect his or her reaction to the sick brother or sister? Does guilt play a role? What type of intervention is needed? With the increasing role of the community in the management of the mentally ill, such research has become very important. PMID- 10857266 TI - A multifaceted mental health training program in reducing burnout among occupational social workers. AB - This article looks at ways of reducing stress through a multifaceted training program involving the acquisition of mental health knowledge and skills and experiential training in a group of occupational social workers. The training group of 13 occupational workers was compared to a group of occupational social workers not attending the program. Post-training revealed statistically significant increases in professional self-efficacy associated with awareness of psychological and psychopathological issues and professional social support. Reduction in cognitive weariness and listlessness associated with burnout was also found. Gaining competence in mental health issues relating to the occupational setting coupled with emotional sharing helped to reduce professional burnout. PMID- 10857267 TI - A comparison between demographic and clinical characteristics of younger and older elderly new referrals to an out-patient psychogeriatric service. AB - We compared the demographic and clinical characteristics of 37 subjects under and 41 subjects over 75 years of age who were consecutive new referrals to an out patient psychogeriatric service in order to examine if an age-related dichotomy emerges in this population. Even though the groups were similar in most demographic (gender, education, years in Israel, family status) and some clinical aspects (number of physical diagnoses, medications taken) they differed significantly in some other clinical variables. "Functional" disorders, independence in performing household activities and Activities of Daily Living (ADL) and more recommendations for ambulatory follow-up were more prominent in the younger group. This profile has much in common with elderly patients newly referred to the general mental health services. The older group had more "organic" disorders, significant need for assistance in household activities and ADL, and more recommendations for follow-up in day-care centers, a profile more characteristic of patients who are treated in memory/dementia clinics. Thus, from a services organizational point of view, it is possible to subdivide out-patient elderly individuals with cognitive and emotional disturbances into separate groups, each with its own characteristics and needs. PMID- 10857268 TI - Impairment in quality of life among patients seeking surgery for hyperhidrosis (excessive sweating): preliminary results. AB - BACKGROUND: The present paper describes the initial stages of the development and administration of a short, disease-specific, health related questionnaire to assess the impact of suffering from hyperhidrosis (excessive sweating) on the Quality of Life (QoL) of patients who are anticipating surgery for this disorder. METHOD: The study was performed in two stages: 1. The life domains in which the condition impairs QoL were assessed by in-depth interviews with 10 patients suffering from hyperhidrosis. 2. A questionnaire covering five life domains was built based on these interviews. 3. This questionnaire was administered to 48 patients, 30 females and 18 males between the ages 15 and 48. RESULTS: Results showed that subjective QoL was significantly lower among females in four of the five life areas and that duration of the condition correlates with a lower quality of life. A regression analysis showed that the subjective suffering of the patients was explained mainly by social aspects. CONCLUSIONS: The questionnaire is a novel attempt to assess QoL in a disorder with strong esthetic and social consequences and could improve communication between patients and their physicians. PMID- 10857269 TI - Lymphocytes, autoantibodies and psychosis--coincidence versus etiological factor: an update. AB - Several factors have been suggested in the etiopathology of schizophrenia, including autoimmune factors. Subgroup of schizophrenics have been found to have immunologic abnormalities. Evidence is presented of the role of lymphocytes and cytokine production in psychiatric disorders. Hypersecretion of IL-2 and IL-6 in acute exacerbation or in relapse-prone patients, decreased ratio of CD4+/CD8+, detection of antinuclear, anticytoplasmatic, antiphospholipid antibodies and others in chronic schizophrenic patients and lately the presence of antiphospholipid antibodies in unmedicated psychotic patients are examples of the immunologic abnormalities findings. These results suggest that schizophrenia may result from an immunologic disorder, which is mediated mainly (but probably not only) by lymphocyte dysfunction. PMID- 10857270 TI - Unusual combination: polydipsia with hypernatremia in a schizophrenic patient. AB - OBJECTIVE: The authors describe a patient with an unusual combination of polydipsia and isolated hypernatremia without any other changes of electrolytes. The patient had two attacks of hypernatremia which were successfully treated with clozapine. Some speculations about possible mechanisms of this unusual combination are discussed. PMID- 10857271 TI - Vitamin B12 levels are low in hospitalized psychiatric patients. AB - BACKGROUND: Deficiency of vitamin B12, a key component in the catabolism of monoamines, is associated with various neuropsychiatric disorders and may be more frequent in hospitalized patients. METHOD: We reviewed vitamin B12 assays performed in a laboratory of a large Israeli psychiatric hospital over a 23-month period to examine prevalence of low values and compared vitamin B12 deficient patients to those with normal levels on various parameters. In addition, vitamin levels in a random sample of in-patients whose nutritional intake was determined, were examined. RESULTS: 20% of 644 vitamin B12 assays were in the low (200 pg/ml) and 10% in the deficient (< 160 pg/ml) range. 24 selected vitamin B12 deficient patients (70.8% with diagnosis of schizophrenia) did not differ from controls (N = 35) in age, sex ratio, hemoglobin concentration, MCV, diagnostic distribution or number and length of hospitalizations, but had slightly lower (but normal) mean folate levels. Rates of vitamin B12 deficiency in the patient sample, whose nutritional intake was adequate, did not differ significantly from those in the laboratory survey. CONCLUSION: Vitamin B12 deficiency is common in chronically ill psychotic patients with adequate nutrition and is not readily detected by routine hematology tests. PMID- 10857272 TI - Cushing's syndrome presenting as a schizophrenia-like psychotic state. AB - Neuropsychiatric abnormalities, of which depression is the most common, are frequently associated with Cushing's syndrome. The following paper describes a 58 year-old woman in whom a prolonged psychotic state--schizophrenia-like--was the presenting sign of Cushing's syndrome. The psychiatric symptoms disappeared shortly after the cortisol level has been normalized. Due to the variety of symptoms, both organic and psychiatric, which may occur with Cushing's syndrome, a high suspicion index is advisable. Suspect clinical findings should always prompt the appropriate endocrine work-up. PMID- 10857273 TI - Patient treatment interaction: where are we and how do we proceed. AB - The need to maximize our treatment efforts and arrive at better therapy results has led to comparative therapy research. However, due to significant differences between patients, this approach has not been successful. Therefore, research into matching treatment to patient began. Unfortunately, single variable models predominated in the field, yielding meager and contradictory results. Two comprehensive models now exist, one offered by Beutler and Clarkin and the other by Anderson. It is hoped that both clinicians and researchers will begin to use these models in therapy as well as in research so that we will not continue behaving as Maslow commented "if you only have a hammer you treat everything like a nail." PMID- 10857274 TI - Story-telling ability in children with autism or Asperger syndrome: a window into the imagination. AB - BACKGROUND: Autism spectrum conditions are diagnosed on the basis of impaired imagination. The present study used a totally free story-telling method to assess if narratives produced by children with autism or Asperger Syndrome (AS) contained fewer imaginative events. METHOD: In Condition 1, children were offered an imaginary theme and asked to elaborate a story. In Condition 2, they were offered a reality-based theme with the same instructions. Comparison groups included 13 children with autism, 14 children with AS, 15 children with moderate learning difficulties (MLD), and 14 normally developing children. The non autistic controls had a verbal mental age (VMA) either equivalent or lower than the autism and AS groups. RESULTS: Both the children with autism and AS were less likely to introduce imaginary elements into their stories in Condition 2, though the children with AS were more able to produce imaginative narratives than children with autism in Condition 1. CONCLUSIONS: This study provides experimental evidence for imaginative impairments in story-telling in children with autism spectrum conditions. These are discussed in terms of two cognitive theories: executive dysfunction and theory of mind. LIMITATIONS: In this study it was not possible to match the children with autism and AS with each other on VMA, as the children with autism were not as high-functioning. Future research could examine VMA matched groups of autism and AS. PMID- 10857275 TI - [Indication for surgical correction of venous hemodynamics in the occlusion stage of post-thrombophlebitic syndrome]. AB - Complex examination of 73 patients with post-thrombophlebitic occlusion of iliac veins was performed. The regional venous hemodynamics indexes and the results of surgical treatment depended upon the disease duration. The indications for surgical correction of the pelvic venous hemodynamics disorders while presence of one-sided occlusion of iliac veins were substantiated. PMID- 10857276 TI - [The influence of sympathectomy of the superior mesenteric artery on the pancreatic blood flow]. AB - The results of performance of intraoperative rheopancreatography in 8 patients, operated on for an ulcer disease, were presented. The performance of selective proximal vagotomy and sympathectomy of a mesenterica superior had promoted the pancreatic blood flow improvement. It is substantiated in patients with secondary chronic pancreatitis, caused by the ulcer disease. PMID- 10857277 TI - [Aspects of pancreatic ductal system in the acute pancreatitis and its complications]. AB - While investigation of pancreatic gland on 133 specimen of the organ it was established, that in edematous pancreatitis the ductal system is hermetic and when the organ destruction occurs--its integrity is compromised. In the occurrence of purulent necrotic complications of an acute pancreatitis the contrast substance "exit" is characteristic; via the lowest ranking ducts in the para-pancreatic cellular tissue necrosis and via the highest ranking ducts--in retroperitoneal phlegmon. PMID- 10857278 TI - [Application of different kinds of internal biliary duct drainage in emergency surgery]. AB - The immediate and late results of treatment of 139 patients, operated on with formation of biliodigestive anastomosis, were analyzed. Six (4.32%) patients died. Of 39 patients, to whom choledochoduodenostomy was performed, in 18--good late follow-up result was noted and the fair one--in 21. Of 23 patients, in whom choledochojejunostomy in Roux-en-Y fashion was performed, good result was noted in 4, fair--in 18 and poor--in 1. PMID- 10857279 TI - [Acute postop ischemic hepatitis and hypotension]. AB - The significance of the pronounced durable systemic arterial hypotension (SAH) in the origin of an acute postoperative ischemic hepatitis (APIH) was established, basing on the analysis of 40 clinical observations. Its occurrence is promoted by hemorrhage with 30% and more the circulating blood volume (CBV) deficiency, chronic cardiovascular system and pulmonary diseases, liver cirrhosis, shock, massive infusions of the blood and its components, the abdominal aorta atherosclerosis with stenosis of tr. coeliacus, a. hepatica. Forgoing SAH, the presence of promoting factors, jaundice, the transpherase activity raising in 3-5 times, the level of blood coagulating factors reduction, stable intestinal paresis were diagnostically significant symptoms. Experimental model of an APIH was elaborated in dogs, which occurs due to hypotension, caused by CBV reduction by 40% during two hours. The refractoriness of a. hepatica propria to the blood reinfusion was established. In the APIH occurrence threat the perftoran application in the 20 ml/kg dosage is the prophylaxis method as well as the method of the curative tactics choice. PMID- 10857280 TI - [Electron microscopy of lymphocytes in patients with acute paralytic ileus]. AB - Characteristic changes of the cells were revealed while examination of 36 patients with an acute adhesional impassability of small intestine and of 14 healthy persons using electron-microscopic method of the peripheral blood lymphocytes studying. The authors consider the observed dystrophic nuclear changes, the nuclear membrane structure and mitochondrial architecture disorders, partial lysis of plasmatic membrane to be the morphological substrate of the intracellular power and the lymphocytes immunoactivity changes. PMID- 10857281 TI - [Strategies of surgical treatment of patients with small intestinal ileus depending on the stage of the disease]. AB - The results of treatment of 316 operated patients with an acute impassability of small intestine (AISI) were analyzed. Four stages of AISI were assigned: 1st stage--compensation, 2nd--sub-compensation, 3d--decompensation, 4th--polyorganic insufficiency. Such classification had permitted to unify the initial state estimation as well as of postoperative course, to differentiate the approach to complex treatment of patients. PMID- 10857282 TI - [The role of temperature in the preparation of cardioplegic solution for the heart open surgery]. AB - There was compared the occurrence frequency of the acute cardiac insufficiency, acute myocardium infarction, acute disorder of cerebral blood circulation, disorders of cardiac rhythm and conduction and other perioperative complications in patients of three groups, operated on using artificial blood circulation, in whom the cardioplegic solution, prepared according to different methods, was used. It is recommended not to permit the crystallization (freezing) of isotonic sodium chloride solution occurrence, used for cardioplegia and for flooding to the pericardium cavity also to cool the heart externally. PMID- 10857283 TI - [The application of regional lymphatic stimulation in the comprehensive treatment of purulent wounds]. AB - There were examined 72 patients with phlegmon, abscess and infected wound of the upper and lower extremities soft tissues. The efficacy of dalargin application for improvement of processes of the purulent wound clearance and reparation was shown using rheologic, biochemical, cytological and planimetric methods of control of the wound process course. PMID- 10857284 TI - [Strategies in the treatment of diabetic foot]. AB - The experience of treatment of 709 patients with diabetic foot was analysed. The tactics of treatment had consisted of the foot immobilization, local antiinflammatory treatment or the antiseptic bandage application, the diabetic phlegmon opening, necrectomy, resection of the affected portion of the foot in the healthy tissues borders; the extremity amputation on the level of foot or hip. Supportive function of the foot was preserved in 97.5% of patients. PMID- 10857285 TI - [Specifics of surgical correction of the oblique inguinal hernia depending on the size]. AB - The results of surgical treatment of 257 patients aged from 10 to 50 years with the oblique inguinal hernia were analyzed. The structure of inguinal channel was studied up using anatomical dissection and measurement on 35 male cadavers aged from 12 to 60 years old. There was substantiated the expediency of surgical correction of the intrachannel and not large external oblique inguinal hernia in the presence of the three-wall-fissure-like inguinal channel using the improved method of the hernial sac mobilization from its neck. PMID- 10857286 TI - [Specifics of preop preparation of patients with diabetes mellitus and destructive pulmonary tuberculosis]. AB - Peculiarities of preoperative preparation in 138 patients with diabetes mellitus and destructive pulmonary tuberculosis were adduced. Application of electrophoresis of the 6% sodium salicylate solution and 5000 U of heparin in the author's modification, ultraviolet irradiation of blood, electrostimulation of lower extremities and part of the back were added to the intensive chemotherapy, diet- and insulinotherapy, detoxication of organism, administration of immunostimulants and other pathogenetically substantiated measures. PMID- 10857287 TI - [Errors and complications in the treatment of the nerve and tendon injuries of the digital flexors]. AB - Retrospective analysis of mistakes and complications occurred during the treatment of 86 patients with the tendons injuries of 157 digital flexors was accomplished. The diagnosis of the tendons injury was established in 94.2% and of the nerves--in 19.6% of patients. In 88.4% of patients the flexing contracture of proximal interphalangeous joints occurred and in the majority of them an active flexing of interphalangeous joints did not restore. The majority of mistakes were caused by traumatic suturing of tendon and prolonged immobilization of the digit in flexing position. PMID- 10857288 TI - [Diagnostic-therapeutic possibilities of the intestinal fibroendoscopy in the young age children]. AB - In 994 children aged up to three years old the experience of endoscopic investigation and curative endoscopy was summarized. PMID- 10857289 TI - [Etiology, pathogenesis, clinical course and treatment of the special forms of acute purulent destructive pyelonephritis]. AB - Ethiopathogenetic peculiarities of an acute purulent obstructive pyelonephritis (PN) were studied in 280 patients. If urinogenic infectioning of kidney occurs the diffuse purulent inflammation which begins from the renal tub spreads on interstitial or the organ medullary substance. In untimely restoration of the urodynamics disorders together with urinogenic the hematogenic infectioning of the kidney had occurred with formation of foci of destruction and purulent processes in cortical substance of kidney. For destructive PN the open operative intervention was performed to patients, in presence of nondestructive purulent PN -transcutaneous nephrostomy. PMID- 10857290 TI - [The use of abdomino-supra-anal resection of rectal cancer]. AB - The results of surgical treatment of 193 patients with upper- and middle ampullar cancer recti, to whom the abdominal supra-anal rectal resection was performed, were summarized. The operation procedure performance was depicted. There were no intraoperative complications. Postoperative complications had occurred in 19 (9.8%) of patients, 3 (1.6%) died. Necrosis of the descended intestine was not noted. The anal sphincter function had restored in 1 mo after performance of operative intervention. PMID- 10857291 TI - [The results of surgical treatment of rectal cancer]. AB - The results of treatment of 948 patients radically operated for cancer recti during 1989-1998 period was studied up. The postoperative complications frequency had lowered as well as mortality from 8.05 down to 1.32% due to application of the improved surgical tactics and an adequate correcting therapy also. Simultaneously there was not noted the trustworthy enhancement of the local recurrencies occurrence frequency and lowering of the average life span of patients after performance of rectal abdomino-anal resection. PMID- 10857292 TI - [The treatment of patients with acute colonic ileus of tumoral genesis at the general surgery department]. AB - The results of treatment of 57 patients with an acute ileus (AI), caused by colonic cancer, were analyzed. The compensated (in 9 patients), sub-compensated (in 35) and decompensated (in 13) forms of AI were assigned. In the right-sided localization of tumor or left-sided one and compensated AI the one-stage radical operation was done. Radical colonic resection or hemicolectomy, like Hartmann's operation, was the operation of choice in sub- and decompensated AI and the left sided localization of tumor. Postoperative mortality had lowered from 33 down to 11% due to individualization of the treatment tactics. PMID- 10857293 TI - [Nonspecific active immunotherapy in surgical treatment of patients with pulmonary cancer]. AB - Application of nonspecific active immunotherapy in the treatment of 59 radically operated patients with pulmonary cancer had permitted in brief period of time to restore quantitative and functional indexes of cellular link of immunity, had promoted noncomplicated course of postoperative period, the reduction of the postoperative complications frequency and improvement of late follow-up results of surgical treatment of the patients in the absence of metastases in regional lymphatic nodes. PMID- 10857294 TI - [Prophylaxis and methods of correction of the parathyroid insufficiency in surgical treatment of the thyroid cancer]. AB - The state of calcium metabolism in 249 patients after performance of thyroidectomy was studied up. It was established that precisional technique of the surgical intervention performance directed on the parathyroid glands vascularization preservation, is the main prophylactic method for the parathyroid insufficiency occurrence. Indications for the parathyroid tissue autotransplantation application during the performance of thyroidectomy are substantiated. PMID- 10857295 TI - [The formation of large intestine anastomoses by experimental electrothermal adhesion]. AB - The method of colonic anastomoses formation by the tissues conjunction using alternating electric current (electric welding) was proposed. The procedure of the anastomosis formation in experiment is adduced. PMID- 10857296 TI - [Chronic duodenal ulcer hemorrhage in elderly and senile patients]. PMID- 10857297 TI - [Surgical treatment of thyroid cancer with laryngotracheal invasiveness]. PMID- 10857298 TI - [The tubing for enteral feeding and gastric and duodenal drainage]. PMID- 10857299 TI - [The evaluation of pancreatic blood flow in patients with gastroduodenal ulcer complicated by hemorrhage, according to rheopancreatography]. AB - The results of the pancreatic gland (PG) blood flow was studied up in 121 patients with hemorrhage of different severity using intraoperative rheopancreatography. Under the influence of hemorrhage in the PG the arterial blood flow is lowering, the arteriovenous shunting mechanisms are starting. PMID- 10857300 TI - [Clinical sequelae of blood remaining in the intestinal inlet after the hemorrhage]. AB - In experiment there were revealed significant changes of composition of aerobic and anaerobic microorganisms in the jejunum and ileum mucosa after hemorrhage of middle severity. While application of antibiotics these changes were more pronounced, demanding serious substantiation of its application. PMID- 10857301 TI - [Diagnosis and surgical treatment of gastroesophageal reflux disease with motor disorders]. AB - There were performed examination and surgical treatment of 93 patients with chronic gastroesophageal reflux disease (GERD) and motor disorders of gastroesophageal junction, esophageal and gastric bodies. In 49 (52.7%) of patients the total fundoplication was done, in 44 (47.3%)--incomplete one. After the operation all the patients had survived. In 5 years and more there were examined 71 (76.3%) of patients, in all of them good and fair results were obtained. PMID- 10857302 TI - [Resection of rectum and bladder for cancer]. AB - In 1970-1997 period 43 patients with cancer recti, invading bladder, were operated on. The base of surgical tactics was performance of primarily restorational operation as independent procedure and in the complex of treatment as well. There was noted the fair immediate and late follow-up result of treatment. PMID- 10857303 TI - [The use of endobiliary interventions for purulent cholangitis in patients with obturation jaundice]. AB - Endobiliary interventions--external cholangiostomy, external-internal drainage, the biliary ducts endoprosthesis--were done in 156 patients with obturational jaundice of various genesis, complicated by purulent cholangitis. The method of the dosed decompression after transcutaneous transhepatic cholangiostomy, which was performed one day before, being applied in 37 patients, was proposed. Application of the method in complex with antibacterial therapy and the extracorporeal detoxication methods had permitted to lower the mortality down to 12.6%, in patients with the abscesses formation cholangitis--to 28.5%. After the stabilization of general status occur 72 patients were operated on. PMID- 10857304 TI - [Minimally invasive interventions for concurrent diseases of extrahepatic biliary ducts and pancreas]. AB - The results of treatment of 689 patients with calculous cholecystitis and of 12 with cholecystogenic acute pancreatitis (AP), to whom laparoscopic interventions were performed, are adduced. In 50 patients the indexes of operative stress were studied up. The laparoscopic cholecystectomy efficacy was established in patients with cholecystogenic AP in the absence of obturational jaundice. PMID- 10857305 TI - [The dynamics of pancreatic blood flow ion various methods of pancreatic stump formation in patients operated for gastric cancer]. AB - The results of direct rheopancreatography and estimation of the central hemodynamics indexes in 22 patients, operated on for gastric cancer with the pancreatic gland (PG) resection performance, are adduced. The phase changes of pancreatic blood flow (hyperemia, ischemia and normalization), not depending on the method of the PG stump formation are noted. PMID- 10857306 TI - [The changes of bile lithogenecity in late postoperative stages after gastric resection]. AB - The bile lithogenicity change in late period after the gastric resection performance had witnessed the presence of heightened risk of pigmental cholelithiasis occurrence. PMID- 10857307 TI - [The immunity index changes in patients with acute adhesional ileus and possibilities for its correction]. AB - In patients with an acute adhesional small intestinal ileus (AASII) the essential change of the organism immunoreactivity was noted, especially of the immunity cell link, the degree of which had depended on duration of intestinal obstruction, the presence of peritonitis and the disease course severity. The autoimmune reactions studying may be used as the prognosis marker for the recurrent adhesions forming occurrence. The immune reactions of postoperative period are correlating with presence of complications. PMID- 10857308 TI - [The importance of radionuclide tests in postop monitoring of patients with differentiated thyroid cancer]. AB - The results of postoperative dynamic examination of 270 patients with differentiated cancer (DC) of thyroid gland (TG) using radionuclide method were analysed. In 74.7% of patients the residual TG tissue was revealed, in 21.8%--the cancer metastases in cervical and mediastinal lymph nodes, in 2.6%--the distant metastases. Inradicalicity of the 1st operation in 14.1% of patients had caused the necessity of the reoperation performance in 43.1% of them. The radioiodine therapy performance after the operation was indicated in 65.5% of patients with DCTG, the distance radiation therapy--in 1.1%. The necessity of performance of postoperative radionuclide monitoring in patients with DCTG was established. PMID- 10857309 TI - [Hirschsprung's disease complicated by severe colonic dilatation]. AB - The Hirschsprung's disease course, complicated by significant dilatation of colon, was considered in 37 patients. The late diagnosis constitutes the cause of the complication occurrence. The "giant" colon is revealed predominantly in senior children with short and ultrashort aganglyosis zone. Surgical treatment needs application of complex, frequently nonstandard methods because of inconcordance of the hypertrophic and dilated intestine size to the size of the anal canal, causing the trophic changes occurrence in the anastomosis region. PMID- 10857310 TI - [Risk factors of the lower extremity amputation and mortality of necrotic inflammation of the foot in patients with diabetes mellitus]. AB - The comparative analysis of the clinical examination, biochemical and morphological investigations data in 75 patients with diabetes mellitus (DM) of the 1st type and in 341 patients with DM of the 2nd type, complicated by necrotically-inflammatory damage of foot (NIDF), was done. Amputation of lower extremity (LE) on the hip level was performed in 21.1%, on the knee one--in 3.6% of patients. Twenty eight, or 6.7%, patients died. There were following risk factors for the high level of the LE amputation and mortality: the senior age of patients, long duration of the DM course, the presence of the metabolism and rheologic properties of blood disorders, progressing atherosclerosis of the main arteries of hip and crus, the anamnesis data concerning other DM complications (nephropathy, anemia), and an ischemic heart disease, myocardium infarction, disorders of cerebral blood supply as well. PMID- 10857311 TI - [Analgesia after orthopedic interventions]. AB - The efficacy of application of tramadol and naclofen for analgesia after performance of orthopedic operations was studied up. There were examined 112 patients, to whom were done total endoprosthesis of coxofemoral joint, 19--total endoprosthesis of the knee joint, 54--the knee joint arthroscopy. The analgesia efficacy was estimated according to visual-analogue scale and to the sensations index. High efficacy of the tramadol application for postoperative analgesia was established. In patients, to whom tramadol and naclofen were administered, maximal analgetic effect was noted. PMID- 10857312 TI - [Mechanisms of incidence and specifics of spondylolysis and spondylolisthesis course in vertebral osteochondropathy]. AB - There were studied up the morphofunctional changes of vertebral motor segments in 21 patients with vertebral osteochondropathy as well as with initial spondylolysis and spondylolisthesis. The patients were from 12 to 21 years old, the average follow-up was 4.5 years. In vertebral deformity in sagittal plane the spondylolystic spondylolisthesis was not revealed in patients, and in particular the leading clinical sign presented was the syndrome of spondyloarthrosis on the level of compensational hyperlordosis; in vertebral osteochondropathy with preserved or the smoothed physiologic vertebral curvatures--dysplastic spondylolisthesis with early occurrence of hyperplastic spondyloarthrosis as a consequence of the Schmorl's hernia in middle lumbar vertebral segments and of instability of the lower lumbar segments due to vertebral sliding shift with subsequent occurrence of hyperplastic spondyloarthrosis predominantly on these levels. The presence of hyperplastic spondyloarthrosis in lower lumbar vertebral segments while overextension causes the risk rise of the vertebral canal stenosis occurrence. PMID- 10857313 TI - [Methods of rehabilitation of the injured anterior cruciate ligament]. AB - In 83 patients in the term up to 3 weeks after the injury the restoration of the damaged anterior cruciform ligament (ACL) was done. In 62 patients there was applied the elaborated method of transosseous refixation of ACL, in 15 with intermediary laceration--suturing using functionally-stabilizing suture, in 6- the alloplasty by the method of the ACL stumps fixing to the implant. Late follow up result was studied up in 72 patients. In 68 of them there was achieved the restoration of the knee joint stability and functioning, in 4--its anterior instability of the 1st stage was noted. PMID- 10857314 TI - [Tissues for the closure of hepatic parenchyma]. AB - The review of literature, dedicated to application of modern materials for the renal parenchyma suturing, including one, based on usage of venous wall and the umbilicus canaliculus vein of man, is adduced. Original method of reflecting fixation using quilting with the help of continuous suture was proposed. PMID- 10857315 TI - [Antimicrobial action of ozone in the treatment of mandibular fracture]. AB - During experiment in vitro and in the clinical environment in patients with the mandibula fracture there were studied up antimicrobic action of the ozonized distilled water, the ozone concentration in which had constituted 0.3 mg/l. Ozonecontaining solution was used in the form of small baths and gargles instead of conventional antiseptic solutions. While local application the pronounced antimicrobic action of ozone was noted. Additionally there was established immunomodulating action of ozone on the local immunity factors in oral cavity, demonstrated by the rise of the secretory immunoglobulin A (SIgA) level and by lowering of the serum immunoglobulins in saliva. This had witnessed the rise of resistance and lowering of expression of the mucosal inflammatory changes. In patients, to whom the conventional treatment was done, the immunologic indexes dynamics was opposite, witnessing presence in them posttraumatic immunodepression. The comparison of data, obtained in the clinic and in experiment, permits to suggest, that antimicrobic action of ozone is mostly mediated via the local immunity activization. PMID- 10857316 TI - [The possibilities of application of iodinedicerinum in urgent surgery]. AB - Application of ioddicerynum while performance of an urgent operative intervention in 63 patients with an acute appendicitis, perforated gastroduodenal ulcer and an old postoperative abscess of anterior abdominal wall had permitted to lower the occurrence frequency for postoperative purulent complications by two times. PMID- 10857317 TI - [Interventional radiology: history, priorities, first experimental results]. PMID- 10857318 TI - [Modern aspects of surgical treatment of hepatic echinococcosis]. PMID- 10857320 TI - [A device for the closure of wounds]. PMID- 10857319 TI - [Acute small intestinal ileus as the symptom of increased intraabdominal pressure]. PMID- 10857321 TI - [Implants and devices for the formation of compressive anastomosis]. PMID- 10857322 TI - [Prophylaxis of purulent complications of acute cholangiogenic pancreatitis]. PMID- 10857323 TI - [Specifics of gastric pseudotumor course]. PMID- 10857324 TI - [The application of microlaparotomy access during the intervention for acute pancreatitis]. PMID- 10857325 TI - [The use of regional intraarterial therapy in the treatment of the lower extremities open fracture, complicated by purulent infection]. PMID- 10857326 TI - [In Process Citation] PMID- 10857327 TI - [The treatment of acute pancreatitis]. AB - Experience of surgical treatment of 721 patient with necrotic pancreatitis was summarized. Indications and tactics of treatment of patients with an acute pancreatitis (AP), methods of their conservative and operative treatment were substantiated. Classification of an AP was elaborated. PMID- 10857328 TI - [The use of greater omentum in revascularization of ischemia-affected organs]. AB - Anatomo--morphological peculiarities of greater omentum (GO) in 63 patients, in whom its flap was applied for revascularization of ischemized tissues of extremities, myocardium and the brain, were studied up. Microsurgical transplantation of the GO free flap was performed in 41 patient, omentocardiopexy without the break of the GO vascular pedicle--in 22. Possibilities of the GO application for the ischemized organs revascularization were estimated. PMID- 10857329 TI - [Possibilities of transcutaneous intervention under ultrasonographic control in comprehensive treatment of severe necrotizing pancreatitis]. AB - In 46 patients with an destructive acute pancreatitis (AP) the invasive intervention under control of ultrasonography, in 38 of them--in stage of purulent complications, was performed. It had promoted stabilization of state in 39 (89.8%) patients, normalization of clinic--laboratory indexes, that had permitted to realize radical operative intervention with minimum risk in perspective. Invasive intervention under control of ultrasonography--highly effective primary surgical manipulation in complex of treatment of the destructive AP severe forms. PMID- 10857330 TI - [Prophylaxis of acute postoperative pancreatitis in the course of treatment of the giant duodenal ulcer]. AB - The results of surgical treatment of 96 patients with giant duodenal ulcer, to whom selective proximal vagotomy (SPV), duodenoplasty (DP), vagotomy with pylorus destroying operation (PDO), gastric resection were performed, were analyzed. Prophylactic methods for an acute postoperative pancreatitis (APP) occurrence before, during and after the operation performance were elaborated. It was established, that the frequency of APP occurrence after SPV and DP is lower than after gastric resection and vagotomy with PDO. PMID- 10857331 TI - [Acute pancreatitis in pregnant women and women in childbirth]. AB - The result of treatment of 11 pregnant women with an acute pancreatitis in 1991 1998 period was analyzed. In all the patients biliary pancreatitis was diagnosed. The necessity of treatment of cholelithiasis in women before pregnancy as well as timely beginning of treatment of pregnant woman just from first signs of an acute pancreatitis occurrence were established. PMID- 10857332 TI - [Classification of biliary peritonitis]. AB - The authors had proposed new classification of biliary peritonitis. PMID- 10857333 TI - [Comprehensive treatment of calculous cholecystitis associated with surgery requiring diseases of other organs]. AB - Experience of performance of concurrent operations in 124 patients with chronic and acute calculous cholecistitis (CC) in presence of concurrent surgical diseases was summarized. Application of oblique minilaparotomic access in right subcostal region without m. rectus abdomini intersection had permitted after the cholecystectomy performance to broaden the indications for the concordant operative interventions performance using other surgical accesses. Inexpediency of doing of surgical interventions on stomach, n. vagi in CC and noncomplicated gastroduodenal ulcer disease in favor of medicinal therapy were substantiated. PMID- 10857334 TI - [Surgical treatment of bleeding duodenal ulcer in elderly and senile patients with intraoperative verification of the diagnosis]. AB - The method of duodenotomy performance for duodenal ulcer with hemorrhage was proposed. Duodenum was dissected longitudinally across the ampulla lower edge going all over medial edge of pars descendens, permitting not only doing the revision but revealing the hemorrhage source, securing homeostasis. The rules of local hemostasis performance were elaborated. Application of the above mentioned access prevents damage of nervi vagi duodenal anterior brunch, guaranteeing the duodenal motor-evacuation function preservation. PMID- 10857335 TI - [Diagnostic possibilities of laparoscopy in acute surgical diseases of the abdominal organs]. AB - The diagnostic combined laparoscopy was performed in 3.3% of patients, hospitalized with conjectural diagnosis of an acute disease of the abdominal cavity organs. In 655 of the total of 16,233 patients operated on in emergency, an acute disease was not revealed. In 37.3% of patients an urgent laparoscopy was performed for the abdominal trauma. Diagnosis, established before the laparoscopy performance, was confirmed in 36.6% of patients. Basing on the urgent laparoscopy data, performed for abdominal trauma, in 133 (49%) of patients the laparotomy conduction have been avoided. Conjectural diagnosis of an acute appendicitis according to the laparoscopy data was confirmed in 25.6% of observations, perforative ulcer--in 28.6%, an acute cholecystitis--in 54.3%, an acute pancreatitis--in 45.6%. PMID- 10857336 TI - [Risk factors in acute ileus]. AB - Results of treatment of 335 patients with an acute ileus (AI) was analyzed. Application of the SAPS system (Simplified Acute Physiology Score) had permitted to estimate the severity of the patient's state with AI and of degree of the homeostasis disorder in any moment of therapeutic-diagnosis process. Three stages of AI are delineated: compensated (in 41.8% of patients), subcompensated (in 38.8%), decompensated (in 19.4%), which demand different approach to diagnosis and treatment. PMID- 10857337 TI - [Thoraco-abdominal wound in gunshot multiple wound]. AB - Thoracoabdominal wounding in the shot gun polytrauma was revealed in 56 (25.6%) of injured persons, severe shock of III-IV degree--in 71.4%. Operative intervention on the abdominal cavity organs was done in all injured persons, on the thoracic one--in 92.9%, and on the other anatomical regions--in 55.4%. PMID- 10857338 TI - [The result of adjuvant chemotherapy of colorectal cancer]. AB - In 103 patients with colorectal cancer the efficacy of three schedules of chemotherapy using fluorouracil (FU), FU with doxorubicin, FU with levamizol (LEV) was studied up. The result of treatment in 3 and 5 years was compared with that in patients, to whom merely surgical treatment was performed. Most effective treatment was application of FU with LEV: the three-year survival index of patients had constituted 66.7%, five-year one--54.5% and in control group--46.8 and 41.3% accordingly. The adjuvant chemotherapy performance was most effective in cancer stage III and in nondifferentiated type of tumor. PMID- 10857339 TI - [The role of radical mastectomy in complex of treatment of patients with locally spread mammary gland cancer]. AB - Results of treatment of 270 patients with the mammary gland cancer (MGC) of III A -III B stage were analyzed. In complex of treatment of patients with MGC III A stage expediency of application of radical mastectomy was established. In patients with MGC III B stage the performance of mastectomy is palliative intervention. PMID- 10857340 TI - [Surgical treatment of abdominal hernia appearing after median laparotomy]. AB - The results of the surgical treatment of 456 patients with postoperative abdominal hernia (PAH) of various localization were analyzed. In 356 (78.5%) of patients PAH had occurred after performance of median laparotomy. In 1 to 10 years the PAH recurrence had occurred in 51 (16.1%) of patients: after hernioplasty according to Sapezhko method in 18, to Napalkov method--in 8, Mayo- in 4, Molodenkov--in 16, closure in layers with the aponeurousis duplicature forming--in 1, after the hernia defect closure at the joint with suture line immobilization--in 4. PMID- 10857341 TI - [The use of transpedicular system of stabilization in surgical treatment of vertebral column tumors]. AB - The experience of application of the titanium transpedicular systems in 23 patients with the vertebral column (VC) tumor was summarized. In all the patients good VC stability was noted after the operation, what had permitted to conduct their early activization. PMID- 10857342 TI - [The use of vobenzym in the comprehensive treatment of patients with digital flexor tendon injury]. AB - The results of treatment of 56 patients with tendons of digital flexors were analyzed. In 28 of them in complex of treatment vobenzim was included, and an early active mobilization as well. Considerable antiinflammatory, antioedematous, secondarily analgetic effect of preparation, its application in early period permitted to realize active movements, to reach high functional result of treatment of patients' were noted. PMID- 10857343 TI - [The use of perftoran in patients with burn shock]. AB - Basing on studying of the biogenic amines metabolism and indexes of cardiodynamics in 49 patients with the burn shock of middle severity it was established, that performance of single infusion of perftoran preparation in the first day of the burn trauma had stimulated the organism adaptation reconstruction due to demolition of the catecholamines metabolism dysbalance, prevalence of stress-limiting processes in the reply to stress structure, formation of moderate hyperdynamics of blood circulation. The double infusion of the preparation had permitted to guarantee the indexes stabilization during all acute period of the burn disease. PMID- 10857344 TI - [Instrumental methods of evaluation of the patient's status in acute mild closed head injury]. AB - Examination of patients in an acute period of mild craniocerebral trauma, using clinical and instrumental methods, was performed. The disorders of state of central and vegetative nervous system, mental and somatic spheres, pancerebral changes of biotocs, the signs of irritation, injury of median structures, hydrocephalia, changes of the vessels tone, lowering of the brain blood filling, difficulty of the venous outflow were revealed. PMID- 10857345 TI - [Specific aspects of the clinical course in some forms of acute purulent inflammatory kidney disease]. AB - Clinic course of an acute purulent-inflammatory disease of kidneys and perinephral corpus adiposum in 244 patients and serous formation of an acute obstructive pyelonephritis (PN)--in 80 was analyzed. Intoxication and cardio vascular system disorders were characteristic clinical features of transition of an acute PN in purulent stage. Local features of destruction and suppuration were less expressed than common symptoms. PMID- 10857346 TI - [The prospects of improving efficacy in the treatment of nephrolithiasis]. AB - In 83.7% of patients with nephrolithiasis the morpho-functional disorders of urinary ways were disclosed. The complex of pathogenetically substantiated treatment of such patients was proposed. Together with performing of sparing removing of calculi the operative correction of hemo- and urodynamics with disorders local homeostasis restoration is performed using application of specific inhibitors of urolithogenesis, basing on hyperbaric oxygenation. PMID- 10857347 TI - ["Reperfusion collapse" phenomenon during post-ischemic reperfusion in experiment]. AB - Postischemic changes of central and regional hemodynamic while various reperfusion regimens have been studied on animal experimental model. We had revealed that despite vasodilatation, peripheral vascular resistance in the postischemic extremity rises, as a result of reperfusion by the lowered perfusion volume. Such phenomenon has been named as "reperfusion-collapse". In spite of the microcirculation disorders progression there have been no significant rising of the peripheral vascular resistance as well as worsening of the regional hemodynamic indexes in the postischemic limb during reperfusion by either normal or increased perfusion volume. PMID- 10857348 TI - [The knee joint endoprosthesis]. PMID- 10857349 TI - [Purulent-septic complications, their prophylaxis and prognosis of its course in patients with acute pancreatitis]. PMID- 10857350 TI - [The need of surgical treatment in patients with pulmonary tuberculosis and diabetes mellitus]. PMID- 10857351 TI - [The use of silicone correctors in complex of treatment of neuropathic digital injury in patients with diabetes mellitus]. PMID- 10857352 TI - [Diagnosis of latent insufficiency of deep veins of lower extremities]. PMID- 10857353 TI - [Monitoring of neuromuscular conductivity during application of norkurone in laparoscopic surgery]. PMID- 10857354 TI - [The gastric ulcer perforation into the pleural cavity]. PMID- 10857355 TI - Managed care: the year in review. PMID- 10857356 TI - Medicare 2000. PMID- 10857357 TI - The Maryland year in review. PMID- 10857358 TI - Maryland's appeals and grievance statute at work. PMID- 10857359 TI - Maryland insurance law: a review. PMID- 10857360 TI - The aftermath of a denial. PMID- 10857361 TI - Towards a philosophy of general practice: a study of the virtuous practitioner. PMID- 10857362 TI - Photosensitized reduction of methyl viologen using eosin-Y in presence of a sacrificial electron donor in water-alcohol mixture AB - Photoreduction of methyl viologen (MV2+) by eosin-Y (EY2-) in the presence of triethanolamine (TEOA) has been investigated in water-methanol mixture by means of steady-state photolysis and laser-flash photolysis in the visible/near infrared regions. The complete conversion to the persistent methyl viologen radical cation (MV.+) was observed in the presence of lower concentrations of EY2 and excess TEOA. By laser-flash photolysis measurements, electron transfer was confirmed to occur from the triplet state of EY2- [3(EY2-)*] to MV2+ in the rate constants of ca 2.0 x 10(10) M-1 s-1. The rates and efficiencies of production of MV.+ were found to be dependent on solvent compositions and concentrations of MV2+, ionic salt and TEOA. The back electron transfer reaction from MV.+ to EY.- was retarded in the presence of TEOA, which supports that EY2- is reproduced by accepting an electron from TEOA. In the presence of excess TEOA, the indirect formation of MV.+ from EY.3-, which was produced by accepting an electron from TEOA, was confirmed. The contributions of both the oxidative and reductive routes of 3(EY2-)* for the MV.+ formation have been confirmed. PMID- 10857363 TI - Solar UV dose patterns in Italy. AB - Since 1992 solar ultraviolet (UV) spectral irradiance (290-325 nm) has been measured at two Italian stations of Rome (urban site) and Ispra (semirural site) using Brewer spectrophotometry. The data collected under all sky conditions, are compared with the output of a sophisticated radiative transfer model (System for Transfer of Atmospheric Radiation--STAR model). The STAR multiple scattering scheme is able to cope with all physical processes relevant to the UV transfer through the atmosphere. The experience so far acquired indicates that, in spite of the unavoidable uncertainties in the input parameters (ozone, aerosol, surface albedo, pressure, temperature, relative humidity, cloud cover), measured and computed clear sky iradiances are in reasonable agreement. The STAR model is applied to build up the solar UV geographic patterns in Italy: the daily dose in the range 290-325 nm is computed at about 70 sites where a thorough and homogeneous climatology is available. For each month the concept of an idealized "standard day" is introduced and the surface distribution of solar UV field determined. The map of solar UV patterns for Italy, available for the first time, meets the study requirements in the field of skin and eye epidemiology, as well as in other investigations dealing with the impact of UV on the biosphere. The results are interpreted in terms of atmospheric and meteorological parameters modulating UV radiation reaching the ground. PMID- 10857364 TI - Hydrogen peroxide formation and decay in iron-rich geothermal waters: the relative roles of abiotic and biotic mechanisms AB - Hydrogen peroxide (H2O2) is widely distributed in surface waters where the primary photochemical formation pathway involves the interaction between dissolved organic carbon (DOC) and ultraviolet radiation (UVR). In laboratory studies using iron-rich water from Yellow-stone's Chocolate Pots spring, H2O2 formation depended on sample treatment (unfiltered, < 0.2 micron filtered, autoclaved) prior to irradiation, suggesting several formation pathways. Similar H2O2 formation in filtered and unfiltered water indicates that it is primarily soluble material that is responsible for H2O2 formation. H2O2 formation with soluble material probably includes only photochemical reactions with DOC and/or metals. Greater H2O2 formation in unfiltered and filtered water than in autoclaved water suggests that the agent(s) involved in H2O2 formation is (are) not stable at high temperatures and pressures and degrade to nonphotoreactive species. Such unstable agents may include DOC and/or dissolved complexes of iron or other metals. UVR absorbance occurs across the UV spectrum and, though slightly greater in the UVA range (320-400 nm), is similar to that of other surface waters. Increased UVR absorbance after autoclaving suggested degradation or alteration of some components, which in turn affected H2O2 formation. The spectral region used for irradiation affected net formation and yield. H2O2 formation in water irradiated with UVA radiation was 2.5-3 times that formed in water irradiated with UVB radiation (280-320 nm) in experiments using artificial light sources. Apparent quantum yields comparable to those reported by others could not be calculated because the instrumental designs are not the same. However, approximate quantum yields were calculated for these experiments but should be viewed with caution. Quantum yields were higher in these experiments (0.0040 mol H2O2 per mol photon at 310 nm and 0.0012 mol H2O2 per mol photon at 350 nm) than values reported by other researchers (< 0.0007 mol H2O2 per mol photon at 300 nm and < 0.0005 H2O2 per mol photon at 340 nm; [Scully, N. M., D. R. S. Lean, D. J. McQueen and W. J. Cooper (1996) Limnol. Oceanogr. 41, 540 548]). In natural solar source experiments, H2O2 formation was greater in experiments with UVA and photosynthetically active radiation (PAR; 400-700 nm) than with PAR alone or with UVB, UVA and PAR. However, H2O2 capacity (nM H2O2 W-1 h-1 m2) was greatest with UVB radiation and lowest with PAR radiation. Source regions could not be studied separately. Dark decay of H2O2 occurred via two mechanisms. The main mechanism responsible for H2O2 decay involved particulate matter (probably microorganisms), whereas a secondary mechanism involved soluble matter (i.e. DOC, metal ions and other dissolved species involved in Fenton reactions). PMID- 10857365 TI - The effect of suntan parlor exposure on delayed and contact hypersensitivity. AB - Cutaneous and systemic immune function are believed to play an important role in cutaneous carcinogenesis. We therefore sought to determine whether the suntan parlor radiation sources commonly used in the United States cause measurable qualitative suppression of immune function and quantitative alterations in circulating T cell subpopulations. Subjects (n = 22) were recruited and randomly assigned to receive suntan parlor exposures (10 full-body UV exposures over a 2 week period, shielding only the right flexural arm) or no exposure. Baseline circulating T lymphocyte subpopulations (T helper lymphocyte, CD4; T suppressor/cytotoxic lymphocyte, CD8) were measured. Two weeks later (upon completion of UV exposures for those in this group), circulating T cell subpopulations were measured and dinitrochlorobenzene (DNCB) sensitization (in the UV group, on the UV-exposed buttock) was performed. Subsequent DNCB elicitation was performed in a bilateral fashion (in the UV group, on the right UV-shielded and the left UV-exposed upper arm). We found that subjects in the UV group demonstrated localized suppression of contact hypersensitivity sensitization and elicitation and also an increase in circulating CD8 cells when compared to the control group (P < or = 0.05). We conclude that suntan parlor exposures, as typically received in this country, suppress contact hypersensitivity and increase the circulating T suppressor/cytotoxic cell number quantity. PMID- 10857366 TI - Expression of psbA genes is regulated at multiple levels in the cyanobacterium Synechococcus sp. PCC 7942. AB - In cyanobacterium Synechococcus sp. PCC 7942 the photosystem II reaction-center protein D1 is encoded by three psbA genes. The psbAI gene encodes D1:1 protein, the form used for acclimated growth, and psbAII and psbAIII genes encode the stress-induced form, D1:2 protein. Strong light and low temperature have been shown to induce the expression of psbAII/III genes and down-regulate the expression of psbAI gene. Recently, we reported the involvement of reduced thiols in the up-regulation of psbAII/III genes. In this study, we have analyzed the regulation of psbA gene expression in Synechococcus further, at both the transcriptional and post-transcriptional levels. We show that the inhibitors of the photosynthetic electron-transfer chain, which have different effects on the redox state of the plastoquinone (PQ) pool, have similar effect on the transcription of psbA genes. The inhibitors 3-(3,4 dichlorophenyl)-1,1 dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) do not cause any changes in psbA gene expression when added under low-light conditions, but dramatically reduce the high-light induction of psbAII/III genes when added upon a high-light shift. Moreover, when the thiol reductant, dithiothreitol, is added to Synechococcus cells together with DCMU concomitant with the high-light shift, no inhibition of psbAII/III gene up-regulation takes place, indicating that the thiol redox state rather than the redox state of the PQ pool regulates psbA gene transcription. We also provide evidence for post transcriptional regulation of psbA gene expression, particularly, inhibition of translation of psbAI transcripts at high light, and demonstrate that the D1 protein synthesis and degradation processes are coregulated in Synechococcus. PMID- 10857367 TI - Effect of carotenoid biosynthesis inhibition on the chlorosome organization in Chlorobium phaeobacteroides strain CL1401. AB - We have studied the effect of the absence of carotenoids on the organization of bacteriochlorophylls (BChls) in chlorosomes of Chlorobium (Chl.) phaeobacteroides strain CL1401. Carotenoid-depleted chlorosomes were obtained by means of 2 hydroxybiphenyl-supplemented cultures. In the presence of the inhibitor, isorenieratene (Isr) and beta-Isr biosynthesis were inhibited to more than 95%, leading to an accumulation of the colorless precursor phytoene inside the chlorosomes. In addition, there was a 30-40% decrease in the baseplate BChl a content. The absorption spectrum of the carotenoid-depleted chlorosomes showed a 10 nm blue shift in the BChl e Qy absorption peak. Under reducing conditions, a decrease in the BChl a/BChl e fluorescence emission ratio was observed in carotenoid-depleted chlorosomes relative to that in control chlorosomes, caused mainly by the decrease in the BChl a content. The steady-state fluorescence emission anisotropy in the BChl e region dropped from approximately 0.24 for native chlorosomes to approximately 0.14 for carotenoid-depleted ones, indicating reorganization of BChl e. The circular dichroism (CD) signal of the carotenoid depleted chlorosomes was increased two times in the BChl e Qy region. A simple model based on the structure proposed was used to explain the observed effects. Carotenoids might affect the angle between the direction of the BChl e Qy transition and the axis of the rod. The orientation of BChl a in the baseplate remains unchanged in carotenoid-depleted chlorosomes, although there is a partial loss of BChl a as a consequence of a decrease in the baseplate size. The carotenoids are most likely rather close to the BChls and appear to be important for the aggregate structure in Chl. phaeobacteroides. PMID- 10857368 TI - Photodynamic therapy of superficial basal cell carcinoma with 5-aminolevulinic acid with dimethylsulfoxide and ethylendiaminetetraacetic acid: a comparison of two light sources. AB - The aim of this prospective randomized study was to compare the clinical and cosmetic outcome of superficial basal cell carcinomas (BCC), using either laser or broadband halogen light, in photodynamic therapy with topical 5-aminolevulinic acid (ALA). A total of 83 patients with 245 superficial BCC were included in the study. Standard treatment involved 15 min of local pretreatment with 99% dimethylsulfoxide (DMSO) before topical application of 20% ALA with DMSO (2%) and ethylendiaminetetraacetic acid (2%) as cofactors for 3 h before light exposure with either laser or a broadband lamp (BL). A complete response was achieved in 95 lesions (86%) in the laser group and 110 lesions (82%) in the BL group 6 months after treatment. Of these, 80 lesions (84%) in the laser group and 101 lesions (92%) in the lamp group were independently evaluated to have an excellent or good cosmetic post-treatment score. No serious adverse events were reported. This study shows that there is no statistical significant difference in cure the rate (P = 0.49) and the cosmetic outcome (P = 0.075) with topical application of a modified ALA-cream between light exposure from a simple BL with continuous spectrum (570-740 nm) or from a red-light laser (monochromatic 630 nm). Cost and safety are further elements in favor of the BL in this setting. PMID- 10857369 TI - Autofluorescence patterns in short-term cultures of normal cervical tissue. AB - Fluorescence spectroscopy has potential to improve cervical precancer detection. The relationship between tissue biochemistry and fluorescence is poorly understood. The goal of this study was to characterize normal cervical autofluorescence, using fresh tissue short-term tissue cultures and epithelial cell suspensions. Transverse, short-term tissue cultures were prepared from 31 cervical biopsies; autofluorescence images were obtained at 380 and 460 nm excitation. Fluorescence excitation-emission matrices were measured from normal, precancerous and cancerous cervical cell suspensions. Observed fluorescence patterns contrast those reported for frozen-thawed tissue, and were placed into groups with (1) bright epithelial and weak stromal fluorescence; (2) similar epithelial and stromal fluorescence; and (3) weak epithelial and bright stromal fluorescence. The average ages of women in the groups were 30.9, 38.0 and 49.2 years. Epithelial fluorescence intensity was similar in Groups 1 and 2, but weaker in Group 3. Stromal intensity was similar in Groups 2 and 3, but weaker in Group 1. The ratio of epithelial to stromal fluorescence intensity was significantly different for all groups. EEMs of cell suspensions showed peaks consistent with tryptophan, reduced form of nicotinamide adenine dinucleotide (phosphate) and flavin adenine dinucleotide. Short-term tissue cultures represent a novel, biologically appropriate model to understand cervical autofluorescence. Our results suggest a biological basis for the increased fluorescence seen in older, postmenopausal women. PMID- 10857370 TI - Multispectral imaging autofluorescence microscopy for the analysis of lymph-node tissues. AB - Although histochemical and immunohistochemical methods are the standard procedures in diagnosis of lymphoproliferative disorders, useful improvements in evidencing histopathologic manifestations can be obtained with the introduction of tissue autofluorescence analyses. We used microspectrofluorometry and a Multispectral Imaging Autofluorescence Microscopy (MIAM) technique to analyze lymph-node biopsies from patients with lymphoadenopathy of different origins. Images of tissue autofluorescence were obtained by excitation at 365 nm of lymph node sections and sequential detection with interference filters (50 nm bandwidth) peaked at 450, 550 and 658 nm. Monochrome images were combined together in a single red-green-blue color image. Most of the fluorescence was observed within the blue spectral band because of large contributions from extracellular collagen and elastin fibers as well as from reduced form of intracellular nicotinamide adenine dinucleotide (phosphate). Autofluorescence imaging shows morphological differences between neoplastic and non-neoplastic tissues. The reactive hyperplasia samples show the typical lymph-node organization with weak fluorescent follicles separated by high fluorescent connective trabeculae. In the neoplastic lymph nodes the loss of follicle organization is observed. Consequently, MIAM permits to discriminate between non neoplastic and neoplastic tissues on the basis of their autofluorescence pattern. Multispectral imaging of tissue autofluorescence may present some advantages with respect to standard histochemical microscopy since it (1) does not require any chemical manipulation of samples; (2) gives real-time results performing the analysis immediately upon specimen resection; and (3) supplies a representation of the biological structure organization linked to endogenous fluorophores. PMID- 10857371 TI - The Monodelphis melanoma model: initial report on large ultraviolet A exposures of suckling young. AB - The objective of this study was to determine whether exposure of early suckling young of the opossum Monodelphis domestica to ultraviolet A (UVA) radiation (320 400 nm) can lead to the development of melanocytic lesions similar to those induced after exposure to ultraviolet B (UVB) radiation (280-320 nm) to total doses as low as 380 J/m2. A total of 576 sucklings received nine exposures of 0.6, 2.6 or 15.5 kJ/m2 per dose (total doses approximately 6, 23 and 140 kJ/m2, respectively) from a Blak Ray lamp source with a narrow range emission at 365 nm. A further 280 sucklings were exposed in the same way to doses of 2.6 kJ/m2 per dose (total approximately 23 kJ/m2) broad-band UVA with visible wavelengths from a Dermalight lamp. Frequency of litter loss following all of the UVA-exposure protocols was similar to that within the same stocks in the colony at large. Only one of the 856 UVA-exposed individuals possessed a melanocytic lesion at the 5 month assessment point. No radiation-induced lesions of any type were evident on the skin of the other animals exposed as sucklings. The affected male was from a group of 70 individuals exposed to the highest total dose (140 kJ/m2) from the Blak Ray light source. The melanocytic hyperplasia was provisionally identified as a potential melanoma but it slowly regressed as the animal aged. We conclude that in the opossum suckling exposure system, the potency of UVA for melanoma induction is extremely low compared with that of UVB. Possible explanations, amenable to further investigations, are given for the low UVA sensitivity of the suckling model compared to the adult exposure model of Ley (Ley, R. D. [1997] Cancer Res. 57, 3682-3684). PMID- 10857372 TI - Comparison of photodynamic targets in a carcinoma cell line and its mitochondrial DNA-deficient derivative. AB - The relative contribution, to cell death, of photodynamic damage to respiratory proteins (known targets of photodynamic therapy with many photosensitizers) and other cellular sites was examined. The models were a human ovarian carcinoma cell line 2008, and its mitochondrial DNA-deficient derivative ET3, which lacks several key respiratory protein subunits. Phototoxicity was compared in the two cell lines with photosensitizers that localized to different cellular compartments. Photosensitizers included Victoria Blue BO (VBBO; mitochondria); Photofrin with a short incubation, (plasma membrane) or a long incubation (intracellular membranes including mitochondria); and Nile Blue A (NBA; lysosomes). Photosensitizer content and localization did not differ between the 2008 and ET3 cells. For sensitizers without a primary mitochondrial localization (NBA and Photofrin with a short incubation), there was no significant difference between 2008 and ET3 toxicity. Consistent with a mitochondrial localization of VBBO and independence from respiratory-chain damage, ET3 cells were less susceptible than 2008 to both dark- and light-activated VBBO-mediated damage. Statistical analysis of the data demonstrated minimal photobleaching of VBBO and a significant difference between the phototoxicity curves of ET3 and 2008. For Photofrin with a long incubation, dark- and phototoxicity effects were similar for both cell lines. Inhibition of respiratory enzymes is thus only a minor component of Photofrin-mediated (long incubation) phototoxicity in these cell lines and is overwhelmed by more significant damage elsewhere, whereas it is a major but not the exclusive element of death mediated by VBBO. PMID- 10857373 TI - Daily and circadian variation in survival from ultraviolet radiation in Chlamydomonas reinhardtii. AB - The survival of organisms depends on their ability to adapt to their environment, one important aspect of which is the daily cycle of day and night. During the day, organisms use a variety of strategies to protect themselves from deleterious ultraviolet (UV) wavelengths of sunlight. Among those strategies could be timing of UV-sensitive cellular processes to occur at night to avoid UV-induced damage. We tested whether the unicellular alga Chlamydomonas reinhardtii uses this strategy by measuring the survival of cells following exposure to UV radiation at different phases of the day. Chlamydomonas cells displayed a rhythm of survival from UV radiation where the most sensitive phases occurred during the end of the day and at the beginning of the night. This phase of sensitivity corresponds to the time of nuclear division. The rhythm continues in constant light indicating control by a circadian clock. The results presented here suggest a hypothesis of how circadian clocks may have evolved; a temporal program whereby light-sensitive processes are timed to avoid sunlight-induced damage would be advantageous and therefore selected. PMID- 10857374 TI - The effect of polarized versus nonpolarized light on melatonin regulation in humans. AB - The aim of this study was to compare the effects of polarized light versus nonpolarized light on melatonin secretion in healthy, humans (mean age, 25 years; N = 6). On separate evenings, each subject was exposed to four different light intensities (20, 40, 80 and 3200 lx) of both polarized and nonpolarized light, as well as to a control, dark exposure. Each evening experiment consisted of a 120 min dark exposure (0000-0200 h) followed by a 90 min light exposure (0200-0330 h). Subjects' pupils were dilated prior to exposures. Blood samples were drawn at the start and end of each light-exposure period and later assayed for melatonin by radioimmunoassay. When compared to control exposures, both polarized and nonpolarized light elicited significant suppression of plasma melatonin at each illuminance (P < 0.03 to P < 0.0001), There were no significant differences between the effects of polarized light and nonpolarized light at any illuminance. The two light stimuli modalities demonstrated very similar fluence-response relationships between illuminance and melatonin suppression. Thus, the human pineal gland is responsive to ocular exposure with polarized light in a dose dependent manner similar to that of nonpolarized light, although no significant differences were detected between polarized and nonpolarized light on melatonin regulation. PMID- 10857375 TI - The enhanced green fluorescent protein as a tool for the analysis of protein dynamics and localization: local fluorescence study at the single-molecule level. AB - The green fluorescent protein (GFP) has emerged, in recent years, as a powerful reporter molecule for monitoring gene expression, protein localization and protein-protein interaction. Several mutant variants are now available differing in absorption, emission spectra and quantum yield. Here we present a detailed study of the fluorescence properties of the Phe-64-->Leu, Ser-65-->Thr mutant down to the single molecule level in order to assess its use in quantitative fluorescence microscopy and single-protein trafficking. This enhanced GFP (EGFP) is being used extensively as it offers higher-intensity emission after blue-light excitation with respect to wild-type GFP. By means of fluorescence spectroscopy we demonstrate the absence of the neutral form of the chromophore and the lack of photobleaching recovery after ultraviolet light irradiation. Furthermore, we show that the EGFP spectral properties from isolated to densely packed molecules are highly conserved. From these experiments EGFP emerges as an ideal molecule for quantitative studies of intra and intercellular tagged-protein dynamics and fluorescence-activated cell sorting, but not for monitoring single-protein trafficking over extended periods of time. PMID- 10857376 TI - [Prognostic markers of sepsis and septic shock]. AB - The first stage of systemic inflammatory response during sepsis and septic shock is the massive production of proinflammatory cytokines. Numerous clinical trials were done investigating various agents that were thought to stop this reaction. The results, however, were disappointing. Then it was realised that massive production of antiinflammatory cytokines could also be delirious. Persistent immunosuppression in the course of sepsis increased the risk of death. Therefore the proper balance between pro- and antiinflammatory mediators is extremely important and the methodologies available for monitoring immunological status in patients with sepsis and septic shock are currently of great interest. PMID- 10857377 TI - [Repair of ionizing radiation induced DNA double strand breaks]. AB - DNA double-strand breaks (DSB) are created by ionizing radiation, an important environmental genotoxic agent. DSB are repaired by two mechanisms associated with recombination. In eukaryotic cells homologous recombination depends on genes belonging to the RAD52 epistatic group. Alternative pathway, DNA end-joining in non-homologous recombination involves DNA-dependent protein kinase (DNA-PK). PMID- 10857378 TI - [Strategies for hepatocyte transplantation]. AB - Hepatic parenchymal cell transplantation is a promising method for managing patients with the acute liver failure and may create a bridge to whole organ grafting. Elimination or reduction of the immunogenicity of the hepatocytes would permit long-term graft survival without the need of non-specific immunosuppression. The presented experimental evidence suggests that modulation of hepatocyte immunogenicity by purification and cryopreservation reduces the alloresponse to hepatocytes both in vitro and in vivo. Identification of inoculated cells facilitates long-term monitoring of their localization and metabolic activity. PMID- 10857379 TI - [Human gastrointestinal tract mucins encoded by the MUC gene family]. AB - Epithelial mucins, MUC gene products, are widely expressed in human organs such as airways, the urogenital and gastrointestinal tracts, and the eyes. MUC-type mucins have very large sizes and complex structures with very extensive O glycosylation and are regarded as protective molecules. The aim of this review is to discuss mucin glycoproteins structure, biosynthesis and functions in normal and pathologically altered epithelial mucosa of human gastrointestinal tract. PMID- 10857380 TI - [The biological role of the basement membranes. Filtering barrier of the glomerulus]. AB - This article presents the latest information about biological functions of basement membranes. We drew a special attention to glomerular basement membrane. Besides, this article contains information about filter barrier and interactions between basal membranes and tissues. PMID- 10857381 TI - [The role of neutrophil elastase in pathophysiology of obturative pulmonary diseases]. AB - The article is the review of the data from literature concerning the role of neutrophil elastase in pathophysiology of bronchial obturative diseases. The functions of elastase and its inhibitors in airways remodelling and in development of asthmatic responses are discussed. PMID- 10857382 TI - [Free radical concepts in the etiopathogenesis of Graves-Basedow disease]. AB - Increased metabolism due to hyperthyroidism leads to the dysfunction of the mitochondria respiratory chain, resulting in elevated formation of the reactive oxygen species in the course of Graves-Basedow disease. It has also been reported that excessive thyroid hormone level may induce oxidative tissue injury. Furthermore, Graves-Basedow disease is frequently associated with changes of the antioxidant defence system activity. The disturbed balance between oxidative and antioxidative processes may be of significant importance in the pathogenesis of Graves-Basedow disease. PMID- 10857383 TI - [Kynurenic pathway in health and disease]. AB - The paper gives a review about kynurenic pathway metabolism of tryptophan. The implications of its metabolites in pathomechanism of several dysfunctions of tissues and organs as well as possibilities of therapeutic interference are described. PMID- 10857384 TI - Caco-2 cell permeability vs human gastrointestinal absorption: QSPR analysis. AB - The aim of this study is to elucidate quantitative structure-permeability relationship (QSPR) of various organic molecules through Caco-2 cells, and to ascertain the relationship between gastrointestinal (GI) absorption in humans and Caco-2 cell permeability. Caco-2 cell permeability and human GI absorption data were obtained from the literature. The maximum hydrogen bond-forming capacity corrected for intra-molecular H-bonding (Hbc) and Lien's QSAR model were used in this study. The latest CQSAR software was utilized in calculating the logarithm of partition coefficient in octanol/water (Clog P) and in deriving all regression equations. For 51 compounds, a significant correlation was obtained between Caco 2 cell permeability (log Pcaco-2) and Hbc, octanol/PBS (phosphate buffered saline, pH 7.4) distribution coefficient (log Doct), log MW and an indicator variable (I) for the charge, with a correlation coefficient of 0.797. When these compounds were divided into three subgroups, namely neutral, cationic and anionic compounds, much better correlations (r = 0.968, 0.915 and 0.931, respectively) were obtained using different combinations of various physico-chemical parameters. A plot of human GI absorption vs. Caco-2 cell permeability obtained from different laboratories reveals that Caco-2 cell permeability cannot be used to precisely predict human GI absorption for compounds with Pcaco-2 below 5 x 10( 6) cm/s, due to interlaboratory and experimental variabilities, and the lack of a simple correlation between human GI absorption and Caco-2 cell permeability. Caco 2 cell permeability may be estimated from the structures of drug molecules using the above-mentioned physicochemical parameters. In general, for compounds with Pcaco-2 above 5 x 10(-6) cm/s, human GI absorption ranges from 50 to 100%. This is generally acceptable for development into oral dosage form. For the compounds with Pcaco-2 below 5 x 10(-6) cm/s, careful interpretation of caco-2 cell permeability and use of internal standard for comparison are recommended. Otherwise, good drug candidates may be excluded due to incorrectly predicted poor absorption. PMID- 10857385 TI - Pharmacology of appetite suppression. AB - Despite a rising worldwide epidemic of obesity there is currently only a very small number of anti-obesity drugs available to manage the problem. Large numbers of differing pharmacological agents reliably produce a reduction in food intake when administered acutely to animals, and when administered chronically they result in a significant decrease in body mass. Behavioural analysis of drug induced anorexia in animals demonstrates that various compounds profoundly effect feeding behaviour in differing ways. This indicates the variety of mechanisms by which pharmacological agents can induce changes in food intake, body weight and eventually body composition. Some of the same drugs produce decreases in food intake and weight loss in humans. Some of these drugs do so by modifying the functioning of the appetite system as measured by subjective changes in feelings of hunger and fullness (indices of satiety). Such drugs can be considered as "appetite suppressants" with clinical potential as anti-obesity agents. Other drugs induce changes in food intake and body weight through various physiological mechanisms inducing feelings of nausea or even by side effect related malaise. Of the drugs considered suitable candidates for appetite suppressants are agents which act via peripherally satiety peptide systems (such as CCK, Bombesin/GRP, Enterostatin and GLP-1), or alter the CNS levels of various hypothalamic neuropeptides (NPY, Galanin, Orexin and Melanocortins) or levels of the key CNS appetite monoamine neurotransmitters such as serotonin (5-HT) and noradrenaline (NA). Recently, the hormone leptin has been regarded as a hormonal signal linking adipose tissue status with a number of key central nervous system circuits. The peptide itself stimulates leptin receptors and it links with POMC and MC-4 receptors. These receptors may also provide drug targets for the control of appetite. Any changes induced by a potential appetite suppressant should be considered in terms of the (i) psychological experience and behavioural expression of appetite, (ii) metabolism and peripheral physiology, and (iii) functioning of CNS neural pathways. In humans, modulation of appetite may involve changes in total caloric consumption, subjective changes in feelings of hunger and fullness, preferences for specific food items, and general macronutrient preferences. These may be expressed behaviourally as changes in meal patterns, snacking behaviour and food choice. Within the next 20 years it is certain that clinicians will have a new range of anti-obesity compounds available to choose from. Such novel compounds may act on a single component of the appetite system or target a combination of these components detailed in this review. Such compounds used in combination with lifestyle changes and dietary intervention may be useful in dealing with the rising world epidemic of obesity. PMID- 10857386 TI - Serotonin, dopamine and norepinephrine transporters in the central nervous system and their inhibitors. AB - An overview is presented on progress made in the research on neuronal transporters of serotonin, dopamine and norepinephrine in the central nervous system. Tools developed by molecular biology, such as expression of cloned transporters, their mutants and chimera in non-neuronal cells offered the opportunity to study the putative domains for binding of substrates and uptake inhibitors and discover factors in the regulation of the transporter function. The study of the distribution of monoamine transporters in human brain became possible by the development of selective radiolabelled transport inhibitors. The relationships between the chemical structure of the uptake inhibitors and the affinity for the monoamine transporters is reported, and the (potential) therapeutic applications of the compounds are discussed. PMID- 10857387 TI - Neuropeptides in drug research. AB - Neuropeptides have been a subject of considerable interest in the pharmaceutical industry over the last 20 years or more. Many drug discovery teams have contributed to our understanding of neuropeptide biology but no significant drugs that act selectively upon neuropeptide receptors have yet emerged from the clinic. There are, however, a plethora of clinically useful drugs that act at other classes of neurotransmitter and neuromodulator receptors, many of them discovered over the last 20 years. Nevertheless, we think that the future for the discovery of novel drugs acting at neuropeptide receptors looks bright for two reasons: (1) there has been a substantial increase in our understanding of the function of neuropeptides; and (2) high-throughput screening (HTS) against neuropeptide receptors has now begun to yield many interesting drug-like molecules, rather than peptides, that have the potential to become clinically useful drugs. The objective of this review is to summarise our current understanding of specific areas of neuropeptide biology and pharmacology in the CNS as well as the PNS. We will also speculate on where we think the new generation of neuropeptide agonists and antagonists could emerge from the clinic. PMID- 10857388 TI - Regulation of NMDA receptors by ethanol. AB - NMDA receptors are glutamate-gated ion channels, mediating excitatory neurotransmission in the brain. These widely distributed receptors are known to play a role in neuronal development and synaptic plasticity, but over stimulation of these receptors can lead to neurotoxicity. In recent years, NMDA receptors have emerged as an important site of action of ethanol. It is believed that at least some of the deleterious effects of ethanol like alcohol dependence, development of tolerance to alcohol and alcohol withdrawal syndrome are mediated via NMDA receptors. The sensitivity of NMDA receptors to ethanol, however, varies regionally. This diversity of NMDA receptor sensitivity is believed to result, at least in part, from heterogeneity of receptor subunit composition. Ethanol's effects on NMDA receptors, including alteration in receptor function and number, probably result from interplay of multiple mechanisms some of which are discussed here. PMID- 10857389 TI - Troglitazone and emerging glitazones: new avenues for potential therapeutic benefits beyond glycemic control. AB - Insulin resistance is characterized as one of the major pathogeneses of type 2 diabetes and has been associated with these same cardiovascular risk factors. Troglitazone, rosiglitazone, and pioglitazone are a new class of oral antidiabetic agents which can ameliorate peripheral insulin resistance in type 2 diabetes. There is considerable evidence that trogliterazone may have beneficial effects on cardiovascular and metabolic abnormalities associated with insulin resistance. There is supportive evidence for positive effects of the other glitazones, but they have been less well studied. These potential benefits span effects ranging from molecular events in the arterial wall to amelioration and/or improvement in lipid parameters known to be associated with atherosclerosis. PMID- 10857390 TI - Applications of developmental biology to medicine and animal agriculture. AB - With the complete sequence of the human genome expected by winter 2001, genomic based drug discovery efforts of the pharmaceutical industry are focusing on finding the relatively few therapeutically useful genes from among the total gene set. Methods to rapidly elucidate gene function will have increasing value in these investigations. The use of model organisms in functional genomics has begun to be recognized and exploited and is one example of the emerging use of the tools of developmental biology in recent drug discovery efforts. The use of protein products expressed during embryo-genesis and the use of certain pluripotent cell populations (stem cells) as candidate therapeutics are other applications of developmental biology to the treatment of human diseases. These agents may be used to repair damaged or diseased tissues by inducing or directing developmental programs that recapitulate embryonic processes to replace specialized cells. The activation or silencing of embryonic genes in the disease state, particularly those encoding transcription factors, is another avenue of exploitation. Finally, the direct drug-induced manipulation of embryonic development is a unique application of developmental biology in animal agriculture. PMID- 10857391 TI - [Student anamnesis groups--help toward self-help]. PMID- 10857392 TI - [Classical theme. The autogenic training of I. H. Schultz]. PMID- 10857393 TI - [The question of the connection between obsessive-compulsive disorder and narcissism]. AB - Freud's psychodynamic concept of the anal-sadistic regression in obsessive compulsive neurosis is completed with the clinical observation of the structural ego deficit in connection with obsessive-compulsive disorder. Not being able to rely on one's own behaviour, described by Quint, for example, explains that the self-assessing function is lacking in the obsessive-compulsive patient. Such patients need the other person to acknowledge and confirm their behaviour as a self-object, i.e. as a part of his self. The aspect of the narcissistic bond of the obsessive-compulsive patient is emphasised here as a differentiation to the more superficial criteria of narcissism in the DSM-IV. The obsessive-compulsive symptomatology has the function to hold on to his self-object as no autonomy is available and loss of bond is experienced as loss of self. The relation of obsessive-compulsive disorder and narcissism is described on the basis of three case histories and supported by biographic and psychotherapeutic data. Furthermore attention is drawn to the consequences of this opinion for a modified psychodynamic therapeutical approach. PMID- 10857394 TI - [Psychosocial sequelae after severe head injury. Interviews with patients and their relatives in comparison]. AB - Head injury patients often suffer from psychosocial sequelae in the long-term. Since patient reports are regarded as little reliable due to lack of awareness, we compared the patients' to the relatives' view of such sequelae. By means of a semi-structured interview, 37 patients and their relatives were investigated on average 4.4 years after severe head injury. Psychosocial and neuropsychological effects were markedly severe in most cases. However, patients' compared to relatives' perspectives differed significantly only regarding report of aggressive-disinhibited behaviour of the patients, which was more often reported on by the relatives than by the patients themselves. This difference was also found in respect of pre-injury behaviour. Surprisingly, in six patients "positive" psychosocial sequelae were reported. For example, a patient said he had become more tender and more serious. Residual morphological findings in these six patients were either rather minor, or marked left-brain lesions. We conclude that the patients' significant other signs should be included in diagnostic and therapeutic efforts of rehabilitation procedures. Since besides neuropsychological factors psychosocial factors seem to influence outcome after brain injury, psychotherapeutic techniques should be more intensively studied in treatment of head injury patients. PMID- 10857395 TI - [Expectations and attitudes of medical students concerning work stress and consequences of their future medical profession]. AB - In the following cross-sectional study 245 medical students from two different terms (three years apart) were being questioned about work stress and the consequences of their future medical profession. Attitudes and expectations regarding future work stress, work satisfaction, coping strategies, gender specific problems, effects on private life and future quality of life were studied. As expected, students in the upper term were able to judge their professional and private future more realistically. The expectations of the advanced students are more realistic and as a consequence it is likely that a disillusioning process is taking place. The results contribute to a better understanding of the present study situation and well-being of medical students concerning their professional and private future which is also important for teaching and the teaching professionals. PMID- 10857396 TI - [School climate and psychological symptoms--correlations between school stress, sense of coherence and physical-psychological impairment in high school students]. AB - The research project "Health Promotion and School Culture" investigated associations between school climate, school stress, sense of coherence and physical/psychological symptoms in high school students. 565 students, aged 15 to 20 from two high schools (Gymnasium) in the canton of Zurich took part in the study. Assessment measures included questionnaires on school climate (FUK 7-10), school stress, sense of coherence (SOC-13), physical (GBB-24) and psychological symptomatology (SCL-90-R, ADS-K). 166 of the students also participated in a standardised psychiatric interview (DIA-X). Results revealed significant associations between 1. school climate and school stress, 2. biographical risk and protective factors and sense of coherence, 3. school stress and sense of coherence, and 4. school-stress and sense of coherence on the one side and symptomatology on the other. The results are discussed in terms of Karasek and Theorell's (1990) three dimensional stress model. Finally, the need for health promotion in school, especially in classes of high school, is addressed. PMID- 10857397 TI - K+ channels and colonic function. PMID- 10857398 TI - Mammalian G-protein function in vivo: new insights through altered gene expression. PMID- 10857399 TI - Structural determinants of scorpion toxin affinity: the charybdotoxin (alpha-KTX) family of K(+)-channel blocking peptides. PMID- 10857400 TI - [Just alarming]. PMID- 10857401 TI - [Effect of antiretroviral treatment and prophylaxis for opportunistic infections on survival of patients with AIDS]. AB - OBJECTIVE: To analyze the influence of anti-retroviral therapy (ART) and prophylaxis against opportunist disease on survival of patients with AIDS. PATIENTS AND METHODS: Study of AIDS patients diagnosed from January 1996 to October 1997 in a Madrid hospital. An analysis was made of demographic, clinical, and immunological data, as well as ART and prophylaxis against Pneumocystis carinii pneumonia (PCP) and tuberculosis (TB). Both univariate and multivariate analyses were performed, as well as Kaplan-Meier curves. RESULTS: A total of 205 patients were included in the study (83% male) with a median age of 34 years. ART, PCP prophylaxis, and TB prophylaxis were received by 147 (72%), 141 (69%) and 22 (11%) patients, respectively. Among individuals on ART, the likelihood of survival at 12 and 22 months since diagnosis of AIDS was made was 79% and 76%, respectively, and among non treated individuals 54% and 54%, respectively (p < 0.05). ART was associated with a 57% decrease in death risk, and regarding PCP prophylaxis, no benefit on survival was found. CONCLUSIONS: ART was significantly associated with a lower risk of death among AIDS patients. The survival rate was not increased with PCP prophylaxis. PMID- 10857402 TI - [Estimate of osteoporosis fracture risk with ultrasound bone assessment]. AB - BACKGROUND: Multiple studies suggest that ultrasound measurement of the bone can be a rapid, cheap, and radiation-free alternative to determine the fracture risk. In this paper the ultrasound measurement of the bone was performed among 288 postmenopausal women, and the influence of gynecologic history and factors related to lifestyle on the obtained values was examined. PATIENTS AND METHODS: One hundred nineteen healthy postmenopausal women and 169 women with previous osteoporotic fractures were included in the study. Both weight and height were determined and a clinical questionnaire was administered to assess factors related to bone mineral density. The values of broadband ultrasound attenuation (BUA) and speed of sound (SOS) were obtained with a contact ultrasound analyzer. RESULTS: Among women without fractures the mean BUA and SOS values (64.1 [14.9] and 1,601.1 [34.5], respectively) were significantly higher than mean BUA (48.8 [17.3]) and SOS (1,573 [57.8]) values among women with fractures (p < 0.001). Using the logistic regression analysis for predicting fracture risk, the model that suited best was that including BUA (OR = 0.668 [0.544-0.818]), age (OR = 1.102 [1.055-1.151]), age at postmenopause (OR = 0.794 [0.731-0.862]) and height (OR = 0.932 [0.883-0.983]). The area under the curve for this model was 0.871. CONCLUSIONS: BUA and SOS values are lower among women with osteoporotic fractures. The fracture risk can be predicted by means of a model including the variables BUA, age, postmenopausal age, and height. PMID- 10857403 TI - [Polymorphism of the gene for vitamin D receptor, bone mass, and bone turnover in women with postmenopausal osteoporosis]. AB - OBJECTIVE: To analyze which genotypes of vitamin D receptor (VDR) are associated with the loss or maintenance of bone mass among spanish postmenopausal women. MATERIAL AND METHODS: A total of 105 women with postmenopausal osteoporosis and 51 postmenopausal women without osteoporosis (control group), diagnosed with bone densitometry (DEXA) were included in the study. The genetic locus was analyzed with RFLP by using the restriction enzyme BsmI. The obtained polymorphism was correlated with clinical, densitometric and metabolic parameters (including bone turnover markers). RESULTS: Two alleles were obtained with three bands of different size: one 580-570 bp band corresponding to allele B, and two 405-400 and 175-170 bp bands for allele b. None of the three VDR genotypes (BB, bb, Bb) was significantly associated with a higher or lower bone mass among the studied populations. Likewise, in the group of patients with osteoporosis no genotype was associated with bone mineral density (BMD) values or with bone turnover parameters analyzed. No differences were found between subgroups with non-severe (n = 58) and severe (n = 47) osteoporosis regarding the association with any of the three VDR genotypes. CONCLUSIONS: None of the VDR genotypes was associated with BMD loss or with the activity of bone turnover parameters analyzed in a group of spanish women with postmenopausal osteoporosis. None of the VDR genotypes confers a higher susceptibility to suffer from severe osteoporosis. PMID- 10857404 TI - [Analysis of factors associated with anticoagulant treatment indication in chronic atrial fibrillation. Prospective study of 170 patients admitted at an internal medicine service]. AB - BACKGROUND: Anticoagulant therapy reduces the risk of stroke among patients with chronic atrial fibrillation. The objective of this study was to evaluate the use of anticoagulant therapy and to analyze the factors associated with the indication of anticoagulants in patients with chronic atrial fibrillation. PATIENTS AND METHODS: Prospective study of all patients with chronic atrial fibrillation admitted to our Department of Internal Medicine from February 1997 to September 1998. From each patient data related to the cause of atrial fibrillation, other associated vascular risk factors, use of anticoagulant and/or antiplatelet agents and contraindication to anticoagulants were recorded. RESULTS: A total of 170 patients with chronic atrial fibrillation were studied. The mean age of patients was 77 years (range: 49-94). One hundred and four patients (61%) were older than 75 years. Atrial fibrillation was the main cause for admission only in 11 patients (6.5%). One hundred and sixty-seven patients (98%) had indication for receiving anticoagulant therapy; however, it was indicated in only 67 patients (39%). In other 68 patients (40%), antiplatelet agents were used. Patients over 75 years received anticoagulants less frequently (p < 0.0001). Factors associated with the prescription of anticoagulants in the bivariate analysis included: diabetes mellitus (p = 0.046), high cholesterol level (p = 0.023), age < or = 75 years old (p < 0.0001), history of previous embolic events (p = 0.001) and valvular atrial fibrillation (p < 0.0001). The multivariate analysis showed that only two factors were indeed associated with the prescription of anticoagulants: age < or = 75 years (OR: 6.15) and valvular atrial fibrillation (OR: 4.24). CONCLUSIONS: Anticoagulant therapy is underused in patients with chronic atrial fibrillation, particularly in elderly patients. PMID- 10857405 TI - [Superior vena cava syndrome caused by multinodular goiter]. AB - Currently, the vena cava superior syndrome (VCSS) is mainly of oncologic origin. We report here four cases of this syndrome caused by intrathoracic multinodular goiter. All patients had compressive symptoms, particularly of the oesophagus and trachea. Axial CT was the imaging technique that delineated the intrathoracic multinodular goiter compressing brachiocephalic vessels. Surgery (total thyroidectomy) was used and all compressive symptoms resolved. PMID- 10857406 TI - [Should antioxidants be recommended for the prevention of cardiovascular diseases?]. PMID- 10857407 TI - [New criteria for the prophylaxis of infective diseases in patients infected by HIV]. PMID- 10857408 TI - [Progressive creeping eruption]. PMID- 10857409 TI - [Young male with cavitated image of the upper right lung lobe]. PMID- 10857410 TI - [Abdominal tumor and fever]. PMID- 10857411 TI - [Recurrent fever and knee pain in an 84-year-old male]. PMID- 10857412 TI - [Pancytopenia and splenic tumor in a young woman]. PMID- 10857413 TI - [Surgical treatment of multiresistant tuberculosis. Report of a case]. PMID- 10857414 TI - [Relation between preoperative plasma levels of hypercoagulation markers and acute phase reactants in patients with rheumatoid arthritis who are candidates for hip or knee arthroplasty]. PMID- 10857415 TI - [Crohn disease: a wide clinical spectrum]. PMID- 10857416 TI - [Sarcoidosis and hyperthyroidism caused by autoimmune thyroiditis]. PMID- 10857417 TI - [Antilipemic agents, hypothyroidism, and rhabdomyolysis]. PMID- 10857418 TI - [Shoshin beri-beri: reversible cause of multiple organ failure]. PMID- 10857419 TI - [Medical and nursing responsibility: interprofessional dialogue is required]. PMID- 10857420 TI - [Outpatient consultation for injuries--management and therapeutic standards]. PMID- 10857421 TI - [Subcutaneous infusion of fluids in geriatric care]. PMID- 10857422 TI - [Palliative medicine--a requirement in clinical practice, education and research]. PMID- 10857423 TI - [Arthrosis as a national disease]. PMID- 10857424 TI - [Goethe between health and disease]. PMID- 10857425 TI - [Drug addiction in adolescence]. PMID- 10857426 TI - [The polite young girl ... the drug addict and the nurse]. PMID- 10857427 TI - [Different drugs and their effects]. PMID- 10857428 TI - [A refuge for adolescents]. PMID- 10857429 TI - [Why a center for parents?]. PMID- 10857430 TI - [Taking care of newborn infants of drug-addicted mothers]. PMID- 10857431 TI - [Public health. Diseases and emerging risk factors in children]. PMID- 10857432 TI - [Caregivers and their secrets]. PMID- 10857433 TI - [Quality of health care and delivery of drugs]. PMID- 10857434 TI - [Prevention of complications from diabetic foot]. PMID- 10857435 TI - [Hepatitis C today]. PMID- 10857436 TI - [What patients say]. PMID- 10857438 TI - [Experience at the Val de Marne/Essonne network]. PMID- 10857437 TI - [The essential questions about hepatitis C]. PMID- 10857439 TI - [Hepatitis C and drug addiction]. PMID- 10857440 TI - [Accidental exposure to blood and hepatitis C]. PMID- 10857441 TI - [Hepatitis C surveillance. Coordination of action in the entire network]. PMID- 10857442 TI - Radioactivity of honeybee honey. PMID- 10857443 TI - Speciation of pulp mill derived resin acids in the Tarawera River, New Zealand. PMID- 10857444 TI - Survey on levels of PCDDs, PCDFs, and non-ortho Co-PCBs in soil and sediment from a high cancer area near a batch-type municipal solid waste incinerator in Japan. PMID- 10857445 TI - Pollution status of wetlands in urban Coimbatore, Tamilnadu, India. PMID- 10857446 TI - Behavior of "organic" and "synthetic" fertilizer nutrients when applied to irrigated, unsaturated soil. PMID- 10857447 TI - Dissipation and mobility of the pyrazol herbicide JV 485 in the soil of winter wheat crops. PMID- 10857448 TI - Adsorption of acid black 1 wastewater by basic oxygen furnace slag. PMID- 10857449 TI - Measuring dermal exposure to pesticide residues with attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. PMID- 10857450 TI - Area and personal airborne exposure during abatement of asbestos-containing roofing material. PMID- 10857451 TI - Uptake of phthalate esters, di(n-butyl)phthalate and di(2-ethylhexyl)phthalate, as environmental chemicals in monkeys in Japan. PMID- 10857452 TI - Total mercury in feathers of white-tailed eagle (Haliaeetus albicilla L.) from Northern Germany over 50 years. PMID- 10857453 TI - Hg biomagnification in the ichthyofauna of the Tapajos River Region, Amazonia, Brazil. PMID- 10857454 TI - Environmental factors and the succession of aquatic insects in a shallow Chinese lake. PMID- 10857455 TI - Acute toxicity of arsenic to three species of New Zealand Chironomids: Chironomus zealandicus, Chironomus sp. a and Polypedilum pavidus (Diptera, Chironomidae). PMID- 10857456 TI - Implications of chemical-based effluent regulations in assessing DNA damage in fathead minnows (Pimephales promelas) when exposed to metal plating wastewater. PMID- 10857457 TI - Toxicity of triphenyltin to Spirulina subsalsa. PMID- 10857458 TI - Effect of zinc chloro complexes to photoluminescent bacteria: dependence of toxicity on metal speciation. PMID- 10857459 TI - Comparison of the sensitivity of Brachionus calyciflorus and Brachionus patulus (Rotifera) to selected heavy metals under low and high food (Chlorella vulgaris) levels. PMID- 10857460 TI - Toxicity and bioaccumulation of copper, zinc, and cadmium in some aquatic organisms. PMID- 10857461 TI - Thermal tolerance of red shiner (Cyprinella lutrensis) after exposure to atrazine, terbufos, and their mixtures. PMID- 10857462 TI - Preferential reduction of Na+/K+ ATPase alpha3 by 17beta-estradiol influences contraction frequency in rat uteri. AB - One beta1 and two alpha (alpha1 and alpha3) isoforms of Na+/K+-ATPase exist in rat uteri. Previous immunocytochemistry studies have suggested that the alpha3 isoform may be involved in calcium regulation indirectly. Estrogens are known to both modulate Na+/K+-ATPase activities in non-uterine tissues and suppress spontaneous uterine contractions in rats. Thus the purpose of this study was to examine the correlation between estrogens-modulated uterine contraction and the expression of Na+/K+-ATPase alpha3 isoform in rats. After 1-, 2-, and 4- day treatments with 17beta-estradiol (E2, 5 microg/ml/kg, s.c., daily), the diameter of uterine horn was measured. The contraction force of uterine strips was measured by standard muscle bath apparatus. The protein abundance and enzyme activity of Na+/K+-ATPase in rat uteri were measured by Western blot analysis and ATPase assay, respectively. One day of E2 decreased both contraction frequency and alpha3-protein expression without the change in uterine diameter, enzyme activity or other isoforms. Two days of E2 reduced contraction frequency, the enzyme activity, as well as alpha3- and beta1- protein abundance but increased alpha1-protein and uterine diameter. Four days of E2 elicited similar effects as two days of E2, but did not affect alpha1-protein abundance. In conclusion, E2 elicits differential effects on isoform expression. After 1-day treatment with 17beta-estradiol, the decrease in the expression of alpha3 and beta1 without a change in Na+/K+-ATPase activity suggests that some isoform other than beta1 exist in rat uteri. The positive correlation between the reduction of alpha3-and the decrease of contraction frequency suggests the involvement of alpha3 isoform in uterine oscillation. PMID- 10857463 TI - Cardiac and pulmonary vagal neurons receive excitatory chemoreceptor input. AB - The effects of hypercapnia and hypocapnia on the activities of the cardiac and pulmonary vagal single fibers were examined in the decerebrated, unanesthetized, paralyzed, and vagotomized cats. The animals breathed 100% O2. Fractional end tidal CO2 concentration was raised to 9% by adding CO2 into the O2 inlet. Average discharge rate of efferent cardiac vagal units (n=10) increased from 1.0+/-0.3 to 2.2+/-0.3 Hz. Hypocapnia apnea was produced by hyperventilation. Activities of cardiac vagal units tested (n = 4) showed dramatic decrease (0.1+/-0.0 Hz). Mean arterial blood pressure did not change significantly under these conditions. In contrast, only instantaneous firing rate during inspiration was significantly increased for efferent pulmonary vagal units (n = 11) during hypercapnia. The activities of the 3 pulmonary vagal units tested with hypocapnia decreased significantly. We concluded that cardiac and pulmonary vagal neurons were excited by chemoreceptor input. PMID- 10857464 TI - Opposition of rapid baroreceptor resetting by prostanoids in rabbits. AB - Arterial baroreceptors reset rapidly within minutes during acute hypertension; baroreceptor pressure threshold (Pth) is increased and the pressure-baroreceptor activity relation is shifted to the right. The purpose of the present study was to determine if prostacyclin (PGI2) or other prostanoids, released during acute hypertension modulate the magnitude of baroreceptor resetting. Baroreceptor activity was recorded from the vascularly-isolated carotid sinus during distension of the sinus with slow pressure ramp in rabbits anesthetized with chloralose. Pressure-activity curves were generated after holding carotid sinus pressure for 10-15 min from 30 to 100 mmHg. In control, the elevation of holding pressure increased Pth from 44+/- to 65+/-5 mmHg (p < 0.05, n = 12). In the presence of PGI2 (20 microM), Pth averaged 43+/-4 and 45+/-3 mmHg (n = 12) after holding pressure at 30 and 100 mmHg, respectively. In the control group before exposing the carotid sinus to indomethacin, an elevation of holding pressure increased Pth from 49+/-2 to 71+/-3 mmHg (p < 0.05, n = 12). After inhibition of the endogenous formation of prostanoids with indomethacin (20 microM), Pth increased by a significantly greater extent from 61+/-2 to 90+/-3 mmHg (p < 0.05, n = 12) with the increase in holding pressure. The slope of the pressure-activity curve (baroreceptor gain) was not influenced by the change in holding pressure. It was increased significantly by PGI2, while decreased by indomethacin. Neither the change in holding pressure nor PGI2 affected the circumferential wall strain of carotid sinus over a wide range of pressure alteration. The results suggest that PGI2 or other prostanoids released during acute hypertension sensitizes baroreceptors and provides a negative feedback mechanism that opposes and limits the magnitude of rapid baroreceptor resetting. PMID- 10857465 TI - Arecoline excites rat locus coeruleus neurons by activating the M2-muscarinic receptor. AB - The action of arecoline on rat locus coeruleus neurons was studied by intracellular recording from the in vitro brain slice preparation. Superfusion of arecoline (0.1-100 microM) caused two dose-related effects, an increased firing rate and, in neurons previously hyperpolarized to a constant potential by passing a steady hyperpolarizing current across the membrane, depolarization. Both effects were associated with a reduction in membrane input resistance. Moreover, the arecoline-induced excitatory effects were antagonized by the muscarinic receptor antagonist, atropine, but not by the nicotinic receptor antagonist, hexamethonium. Methoctramine, a selective M2-muscarinic receptor antagonist, was also effective in reversing the arecoline-induced effects, with a dissociation equilibrium constant of 14.2+/-1.2 nM (n=6). These results therefore suggest that arecoline exerts its excitatory actions by binding to M2-muscarinic receptors on the cell membrane of neurons of the locus coeruleus. PMID- 10857466 TI - NPC-15199, a novel anti-inflammatory agent, mobilizes intracellular Ca2+ in bladder female transitional carcinoma (BFTC) cells. AB - This report demonstrates that NPC-15199 [(N-(9-fluorenylmethoxycarbonyl)L leucine)], a novel anti-inflammatory agent, increases intracellular Ca2+ concentration ([Ca2+]i) in human bladder female transitional cancer (BFTC) cells. Using fura-2 as a Ca2+ probe, NPC-15199 (0.1-2 mM) was found to increase [Ca2+]i concentration-dependently. The response saturated at 2-5 mM NPC-15199. The [Ca2+]i increase comprised an initial rise, a slow decay, and a plateau. Ca2+ removal partly inhibited the Ca2+ signals. In Ca2+-free medium, pretreatment with 1 mM NPC-15199 abolished the [Ca2+]i increase induced by 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor); and after pretreatment with thapsigargin, NPC-15199-induced Ca2+ release was dramatically inhibited. This indicates that NPC-15199 released internal Ca2+ mostly from the endoplasmic reticulum. Adding 3 mM Ca2+ increased [Ca2+]i in cells pretreated with 1 mM NPC 15199 in Ca2+-free medium. Together, the findings suggest that in BFTC bladder cancer cells, NPC-15199 induced Ca2+ release from the endoplasmic reticulum and activating Ca2+ entry. PMID- 10857467 TI - Startle responses to electric shocks: measurement of shock sensitivity and effects of morphine, buspirone and brain lesions. AB - The present study developed a new protocol to assess shock sensitivity in rats. Male Wistar rats were subjected to footshock stimuli ranging from 0 to 1.6 mA (0.1 s) in a startle apparatus and startle responses elicited by shocks were measured. Acoustic stimuli (95, 105, or 115 dB) were dispersed within the shock series serving as a control measurement of motor performance. Results indicated that the magnitude of shock startle responses significantly increased with the shock intensity in a linear trend. Morphine (8.0 mg/kg) and buspirone (1.0, 2.5, or 5.0 mg/kg), both of which possessing analgesic effects, depressed shock startle but had no such effect on acoustic startle. The effect of morphine was readily reversed by pretreatment of naloxone (1.0 mg/kg). To investigate the neural basis underlying this response, radio-frequency lesions of various structures implicated in processing of nociceptive or aversive information were undertaken. Lesions of the ventroposterior thalamic nucleus, insular cortex, or amygdala decreased startle reactivity to electric shocks but not to acoustic stimuli. Lesions of the anterior cingulate gyrus or medial prefrontal cortex, while altered the reactivity to acoustic stimuli, had no effect on the shock elicited startle. These results suggested that the amplitude of startle in response to electric shocks provide a quantitative measurement of shock sensitivity within an extended range of stimulus intensities. Performing this response may engage the the central nociceptive pathway. PMID- 10857468 TI - Origins of health services research. PMID- 10857469 TI - Access of vulnerable groups to antiretroviral therapy among persons in care for HIV disease in the United States. HCSUS Consortium. HIV Cost and Services Utilization Study. AB - OBJECTIVE: To employ the behavioral model of health services use in examining the extent to which predisposing, enabling, and need factors explain the treatment of the HIV-positive population in the United States with highly active antiretroviral therapy (HAART). DATA SOURCE: A national probability sample of 2,776 adults under treatment for human immunodeficiency virus (HIV) infection. STUDY DESIGN: The article uses data from the baseline and six-month follow-up surveys. The key independent variables describe vulnerable population groups including women, drug users, ethnic minorities, and the less educated. The dependent variable is whether or not a respondent received HAART by December 1996. DATA COLLECTION: All interviews were conducted using computer-assisted personal interview instruments designed for this study. Ninety-two percent of the baseline interviews were conducted in person and the remainder over the telephone. PRINCIPAL FINDINGS: A multistage logit regression shows that the predisposing factors that have previously described vulnerable groups in the general population with limited access to medical care also define HIV-positive groups who are less likely to gain early access to HAART including women, injection drug users, African Americans, and the least educated (odds ratios, controlling for need, ranged from 0.35 to 0.59). CONCLUSIONS: Those HIV-positive persons with the greatest need (defined by a low CD4 count) are most likely to have early access to HAART, which suggests equitable access. However, some predisposing and enabling variables continue to be important as well, suggesting inequitable access, especially for African Americans and lower-income groups. Policymakers and clinicians need to be sensitized to the continued problems of African Americans and other vulnerable populations in gaining access to such potentially beneficial therapies. Higher income, anonymous test sites, and same day appointments are important enabling resources. PMID- 10857470 TI - Children of working low-income families in California: does parental work benefit children's insurance status, access, and utilization of primary health care? AB - OBJECTIVE: To examine financial and nonfinancial access to care and utilization of primary health care services among children of working low-income families earning below 200 percent of the federal poverty level in California, and to compare them to children in nonworking low-income families and in families earning over 200 percent of poverty. DATA SOURCES/STUDY SETTING: The 1994 National Health Interview survey weighted to reflect population estimates for California. STUDY DESIGN: This cross-sectional study of 3,831 children under age 19 focuses on financial access, that is, the prevalence and continuity of health insurance coverage; structural access, including the presence of a usual source of care, the predominant care source, its responsiveness to patient's needs, and any indications of delayed or missed care; and utilization of health care measured by the presence of an outpatient doctor's visit and the mean number of visits relative to child health status. DATA COLLECTION: The study uses secondary analysis. FINDINGS: Compared to children of nonworking low-income parents and to nonpoor children, children of working low-income parents were more likely to be uninsured (32.1 percent versus 15.6 percent and 10.3 percent, p = .0001) and to experience disruptions in insurance coverage (p = .0009). These differences persisted after controlling for other covariates in multivariate analyses. Children of working low-income parents did not differ significantly from children of nonworking low-income parents on measures of structural access or utilization, after adjusting for other covariates. However, they differed significantly from nonpoor children on structural access and utilization, and these differences mostly persisted after adjusting for other covariates (odds ratios from 1.5 to 2.9). Similar patterns were observed when children of full-time, year-round working parents with low earnings were compared with the two reference populations. CONCLUSION: Children in working low-income families in California have some of the worst access problems. Even full attachment to the workforce does not guarantee health insurance benefits, access to care, or improved health care use for children of low-income parents. These children are not better off than other low-income children of nonworking parents and are much worse off than nonpoor children. Expansion of health insurance coverage through Healthy Families and Medi-Cal, and attention to nonfinancial barriers to care for working low income families may help to reduce these disparities. PMID- 10857471 TI - Medical and psychosocial services in drug abuse treatment: do stronger linkages promote client utilization? AB - OBJECTIVE: To examine the extent to which linkage mechanisms (on-site delivery, external arrangements, case management, and transportation assistance) are associated with increased utilization of medical and psychosocial services in outpatient drug abuse treatment units. DATA SOURCES: Survey of administrative directors and clinical supervisors from a nationally representative sample of 597 outpatient drug abuse treatment units in 1995. STUDY DESIGN: We generated separate two-stage multivariate generalized linear models to evaluate the correlation of on-site service delivery, formal external arrangements (joint program/venture or contract), referral agreements, case management, and transportation with the percentage of clients reported to have utilized eight services: physical examinations, routine medical care, tuberculosis screening, HIV treatment, mental health care, employment counseling, housing assistance, and financial counseling services. PRINCIPAL FINDINGS: On-site service delivery and transportation assistance were significantly associated with higher levels of client utilization of ancillary services. Referral agreements and formal external arrangements had no detectable relationship to most service utilization. On-site case management was related to increased clients' use of routine medical care, financial counseling, and housing assistance, but off-site case management was not correlated with utilization of most services. CONCLUSIONS: On-site service delivery appears to be the most reliable mechanism to link drug abuse treatment clients to ancillary services, while referral agreements and formal external mechanisms offer little detectable advantage over ad hoc referral. On-site case management might facilitate utilization of some services, but transportation seems a more important linkage mechanism overall. These findings imply that initiatives and policies to promote linkage of such clients to medical and psychosocial services should emphasize on-site service delivery, transportation and, for some services, on-site case management. PMID- 10857472 TI - Across time and space: variations in hospital use during Canadian health reform. AB - OBJECTIVES: To investigate change in hospital utilization in a population and to discuss analytical strategies using large administrative databases, focusing on variations in rates of different types of hospital utilization by income quintile neighborhoods. DATA SOURCES: Hospital discharge abstracts from Manitoba Health, used to study the changes in utilization rates over eight fiscal years (1989 1996). STUDY DESIGN: We test the hypotheses that health reform has changed utilization rates, that utilization rates differ significantly across income quintiles (defined by the relative affluence of neighborhood of residence), and that these variations have been maintained over time. Our approach uses generalized estimating equations to produce robust and consistent results for studying rates of recurrent and nonrecurrent events longitudinally. DATA EXTRACTION METHODS: Rates of individuals hospitalized, hospital discharges, days of hospitalization, and hospitalization for different types of medical conditions and surgical procedures are generated for the period April 1, 1989 through March 31, 1997 for residents of Winnipeg, Manitoba. Data are grouped according to the individual's age, gender, and neighborhood of residence on April 1 of each of the eight fiscal years for the rate calculations. Neighborhood of residence and the 1991 Canadian Census public use database are used to assign individuals to income quintiles. PRINCIPAL FINDINGS: The substitution of outpatient surgery for inhospital surgery accounted for much of the change in hospital utilization over the 1989-1996 period. Health care reform did not have a significant effect on the utilization gradient already observed across socioeconomic groups. Health reform markedly accelerated declines in in-hospital utilization. CONCLUSIONS: Grouping the data with key characteristics intact facilitates the statistical analysis of utilization measures previously difficult to study. Such analyses of variations across time and space based on parametric models allows adjustment for continuous covariates and is more efficient than the traditional nonparametric approach using standardized rates. PMID- 10857473 TI - A trauma resource allocation model for ambulances and hospitals. AB - OBJECTIVE: To develop a mathematical model for the location of trauma care resources. DATA SOURCES/STUDY SETTING: Severely injured patients queried from Maryland hospital discharge and vital statistics data. A spatial injury profile was created by parsing these patients into ZIP codes. STUDY DESIGN: The Trauma Resource Allocation Model for Ambulances and Hospitals (TRAMAH) was formulated using integer and heuristic programming. To maximize coverage of severely injured patients, trauma centers and aeromedical depots were simultaneously sited using TRAMAH. A severe injury was considered covered if at least one trauma center was sited within a time standard by ground, or if an aeromedical depot-trauma center pair was sited in such a way that the sum of the flying time from the aeromedical depot to the scene of injury plus the flying time from the scene of injury to the trauma center was within the same time standard. PRINCIPAL FINDINGS: From 1992 to 1994, 26,774 severe injuries were considered for coverage. Across Maryland, 94.8 percent of severely injured residents had access to trauma system resources within 30 minutes and 70.3 percent had access within 15 minutes. For the same number of resources as the existing Maryland Trauma System, TRAMAH achieved a coverage objective of 99.97 percent within 30 minutes. This translated into an additional 461 severely injured people covered each year. Holding in place the trauma centers of the existing system, approximately the same percentage of coverage as that of the existing system was achieved within 15 minutes by optimally locating six fewer aeromedical depots. CONCLUSIONS: TRAMAH will allow trauma systems planners to better locate their resources with respect to spatial needs and response times. PMID- 10857474 TI - Comparing the agreement among alternative models in evaluating HMO efficiency. AB - OBJECTIVE: To describe the efficiency of HMOs and to test the robustness of these findings across alternative models of efficiency. This study examines whether these models, when constructed in parallel to use the same information, provide researchers with the same insights and identify the same trends. DATA SOURCES: A data set containing 585 HMOs operating from 1985 through 1994. Variables include enrollment, utilization, and financial information compiled primarily from Health Care Investment Analysts, InterStudy HMO Census, and Group Health Association of America. STUDY DESIGN: We compute three estimates of efficiency for each HMO and compare the results in terms of individual performance and industry-wide trends. The estimates are then regressed against measures of case mix, quality, and other factors that may be related to the model estimates. PRINCIPAL FINDINGS: The three models identify similar trends for the HMO industry as a whole; however, they assess the relative technical efficiency of individual firms differently. Thus, these techniques are limited for either benchmarking or setting rates because the firms identified as efficient may be a consequence of model selection rather than actual performance. CONCLUSIONS: The estimation technique to evaluate efficient firms can affect the findings themselves. The implications are relevant not only for HMOs, but for efficiency analyses in general. Concurrence among techniques is no guarantee of accuracy, but it is reassuring; conversely, radically distinct inferences across models can be a warning to temper research conclusions. PMID- 10857475 TI - Effect of multiple-source entry on price competition after patent expiration in the pharmaceutical industry. AB - OBJECTIVE: To analyze the effect of multiple-source drug entry on price competition after patent expiration in the pharmaceutical industry. DATA SOURCES: Originators and their multiple-source drugs selected from the 35 chemical entities whose patents expired from 1984 through 1987. Data were obtained from various primary and secondary sources for the patents' expiration dates, sales volume and units sold, and characteristics of drugs in the sample markets. STUDY DESIGN: The study was designed to determine significant factors using the study model developed under the assumption that the off-patented market is an imperfectly segmented market. PRINCIPAL FINDINGS: After patent expiration, the originators' prices continued to increase, while the price of multiple-source drugs decreased significantly over time. By the fourth year after patent expiration, originators' sales had decreased 12 percent in dollars and 30 percent in quantity. Multiple-source drugs increased their sales twofold in dollars and threefold in quantity, and possessed about one-fourth (in dollars) and half (in quantity) of the total market three years after entry. CONCLUSION: After patent expiration, multiple-source drugs compete largely with other multiple-source drugs in the price-sensitive sector, but indirectly with the originator in the price-insensitive sector. Originators have first-mover advantages, and therefore have a market that is less price sensitive after multiple-source drugs enter. On the other hand, multiple-source drugs target the price-sensitive sector, using their lower-priced drugs. This trend may indicate that the off-patented market is imperfectly segmented between the price-sensitive and insensitive sector. Consumers as a whole can gain from the entry of multiple-source drugs because the average price of the market continually declines after patent expiration. PMID- 10857476 TI - A simple approach for the analysis of intracellular movement of oxidant-producing intracellular compartments in living human neutrophils. AB - In human neutrophils, superoxide is generated primarily within specialized oxidant-producing intracellular compartments. The present study employs a simple methodological approach to evaluate the intracellular movement of these structures in living human neutrophils. Using a CCD camera system, we monitored fluorescence in cells loaded with the succinimidyl ester of dichlorodihydrofluorescein diacetate, which is nonfluorescent until oxidized by reactive oxygen species. Fluorescence-positive intracellular compartments became detectable after neutrophils were stimulated with phorbol myristate acetate for 1 min. Further stimulation increased the intracellular compartments in both number and size in a time-dependent manner. Upon stimulation with phorbol myristate acetate, no fluorescence was seen in intracellular compartments of neutrophils isolated from patients with X-linked chronic granulomatous disease lacking gp91 phox, a membrane component of NADPH oxidase. The method enables tracking of the movement of a single oxidant-producing intracellular compartment following cell stimulation and visualization of the intracellular structures formed by fusion of oxidant-producing intracellular compartments with endocytotic vesicles and phagosomes. Therefore, it is considered to be an informative tool for evaluation of the intracellular dynamics of oxidant-producing intracellular compartments in living human neutrophils and may have a diagnostic value. PMID- 10857477 TI - Expression, characterization, and localization of Rab26, a low molecular weight GTP-binding protein, in the rat parotid gland. AB - We investigated the expression of the genes encoding Rab proteins, low molecular weight GTP-binding proteins, in the rat parotid gland by the use of reverse transcription-polymerase chain reaction, and detected cDNAs of Rab3D, Rab4, and Rab26. We further examined the characteristics and localization of Rab26 by western blotting and light and electron microscopic immunocytochemistry. Western blotting using an antibody against the Rab26-specific, C-terminal peptide detected the His-tagged Rab26 protein as a single 27-kDa band. This band also displayed binding to [alpha-32P]GTP. The fraction containing secretory granule membranes in an acinar cell homogenate was immunostained with the antibody. Supporting this, the immunocytochemical reaction for Rab26 was localized immediately around secretory granules in the acinar cells. The immunostaining disappeared from the acinar cells after treatment of rats with isoproterenol. These findings suggest that Rab26 participates in the regulated secretion of granules and functionally belongs to the Rab3 group. PMID- 10857478 TI - A preformed basal lamina alters the metabolism and distribution of hyaluronan in epidermal keratinocyte "organotypic" cultures grown on collagen matrices. AB - A rat epidermal keratinocyte (REK) line which exhibits histodifferentiation nearly identical to the native epidermis when cultured at an air-liquid interface was used to study the metabolism of hyaluronan, the major intercellular macromolecule present in basal and spinous cell layers. Two different support matrices were used: reconstituted collagen fibrils with and without a covering basal lamina previously deposited by canine kidney cells. REKs formed a stratified squamous, keratinized epithelium on both support matrices. Hyaluronan and its receptor, CD44, colocalized in the basal and spinous layers similar to their distribution in the native epidermis. Most (approximately 75%) of the hyaluronan was retained in the epithelium when a basal lamina was present while most (approximately 80%) diffused out of the epithelium in its absence. While REKs on the two matrices synthesized hyaluronan at essentially the same rate, catabolism of this macromolecule was much higher in the epithelium on the basal lamina (half-life approximately 1 day, similar to its half-life in native human epidermis). The formation of a true epidermal compartment in culture bounded by the cornified layer on the surface and the basal lamina subjacent to the basal cells provides a good model within which to study epidermal metabolism. PMID- 10857479 TI - The congenital chloride diarrhea gene is expressed in seminal vesicle, sweat gland, inflammatory colon epithelium, and in some dysplastic colon cells. AB - Congenital chloride diarrhea (CLD) is an autosomal recessive disorder of intestinal electrolyte transportation caused by mutations in the anion transporter protein encoded by the down-regulated in adenoma (DRA), or CLD, gene. In this study, in situ hybridization and immunohistochemistry were performed to investigate the expression of CLD in extraintestinal normal epithelia and in intestinal inflammatory and neoplastic epithelia. The expression of the closely related anion transporter diastrophic dysplasia sulfate transporter, DTDST, was also examined and compared with that of CLD in colon. The only extraintestinal tissues showing CLD expression were eccrine sweat glands and seminal vesicles. In inflammatory bowel disease and ischemic colitis, expression of CLD mRNA in colon epithelium was similar to histologically normal colon epithelium, but the protein was found deeper in crypts, including proliferative epithelial cells. In intestinal tumors, the expression pattern of CLD was dependent on the differentiation status of the tissue studied: epithelial polyps with no or minor dysplasia showed abundant expression, whereas adenocarcinomas were negative. The DTDST gene was abundantly expressed in the upper crypt epithelium of colonic mucosa. PMID- 10857480 TI - Enzyme histochemistry on human placental trophoblasts: the effect of fixation delay on enzyme activity. AB - We examined the effect of time delay in tissue fixation on enzyme histochemically detectable enzyme activity and its localization in term human placental trophoblasts. Four placental enzymes, alkaline phosphatase, acid phosphatase, glucose-6-phosphatase, and cytochrome c oxidase, were studied. A fixation delay of 15 min did not markedly alter the activity or distribution pattern of the four enzymes, excepted for a slight reduction in cytochrome c oxidase activity and the appearance of dilated endoplasmic reticula positive for glucose-6-phosphatase. A fixation delay of 60 min abolished cytochrome c oxidase activity, but the activities of the other three enzymes remained positive. When the placental tissue was stored at 4 degrees C without cutting for 24 h before fixation, cell degeneration occurred. However, alkaline phosphatase activity was still clearly demonstrable. In enzyme histochemistry, ,,immediate" fixation is superior, but even if this cannot be performed, the placentas, especially when they are from patients with rare disorders, should not be discarded. Observations made here will be useful for clinician's attempting enzyme histochemistry in organs other than the placenta. PMID- 10857481 TI - In situ detection of apoptosis in dental and periodontal tissues of the adult mouse using annexin-V-biotin. AB - An early event in apoptosis is exposure of phosphatidylserine, an aminophospholipid normally present in the inner leaflet of the plasma membranes, at the outer leaflet of the plasma membrane facing the extracellular space. Annexin V (Anx-V) is a 35-kDa protein with high affinity for phosphatidylserine, which can be applied to detect apoptosis. We injected biotin-labelled Anx-V intravenously in adult mice and examined the tissue distribution of Anx-V labelled cells in dental and periodontal tissues using ABC-peroxidase histochemistry. In the continuously erupting incisors, strong and frequent immunostaining was observed in transitional stage and late maturation stage ameloblasts with less frequent staining in preameloblasts. Frequency of staining in odontoblasts and pulp cells was low but increased slightly at older stages of dentinogenesis. Labelling was also seen in phagocytic or phagocytic-like cells in the enamel organ and pulp. A positive staining was furthermore found in fibroblasts of the periodontal ligament in continuously erupting incisors and in fully erupted molar teeth. Staining intensity and the number of positive cells were enhanced by antigen retrieval using high-pressure cooking. We conclude that Anx-V-biotin labels dental cells in early stages of cell death and indirectly cells that have ingested labelled apoptotic cells during the course of the experiment. The data confirm that during amelogenesis most cell death occurs in transitional stage and late maturation stage ameloblasts. Thus, labelling with Anx-V is a useful marker for studying cell death and the dynamics of clearance of apoptotic cells during tooth development. PMID- 10857482 TI - Immunological detection of polycystin-1 in human kidney. AB - Polycystin-1 is the protein product of the PKD1 gene. Mutations in this gene are responsible for most cases of polycystic kidney disease, but little is known about how these mutations lead to the development of cysts. Indeed, even less is known about the normal role of polycystin-1 in the kidney. The cellular localization of polycystin-1 has been the subject of intense investigation by many groups, including ours. In this report we describe our results and compare our data with those of others. We generated 14 different polyclonal antisera against fragments of the predicted 462-kDa polycystin-1 molecule to enable us to investigate the expression of polycystin-1 in cells and tissues by immunocytochemistry, western blotting, and immunoprecipitation. Our antibodies readily recognized a 134-kDa polycystin-1 fragment overexpressed in COS cells and stained the epithelial cells of fetal, adult, and cystic kidney sections with the same pattern as reported by others. However, further investigations revealed that this pattern was not specific for polycystin-1. We could not unequivocally detect polycystin-1 in vivo, either by immunoblotting or immunocytochemistry. Therefore our studies do not support the reported pattern of polycystin-1 expression in the kidney. PMID- 10857483 TI - Tissue transglutaminase in the small intestine of the mouse as a marker for apoptotic cells. Colocalization with DNA fragmentation. AB - Besides the morphological changes in cells undergoing apoptosis, such as chromatin condensation and cell shrinkage, histological demonstration of DNA fragmentation by in situ end labeling (ISEL) has been widely used for the demonstration of apoptotic cells in tissue sections. Although DNA fragmentation can be demonstrated in apoptotic cells and apoptotic bodies in most cases, there is no clear correlation of ISEL staining with apoptosis. It has often been demonstrated that, in many morphologically intact cells, nuclei with fragmented DNA can be found. Thus staining with ISEL for the detection of apoptosis is useful only in connection with other markers for apoptosis as, for example, characteristic morphological changes. Here we show that tissue transglutaminase protein is unequivocally expressed in apoptotic enterocytes as shown by DNA fragmentation and morphology. Tissue transglutaminase is not expressed in enterocytes with healthy morphology, although DNA fragmentation can be demonstrated in these cells. Thus the immunohistochemical demonstration of tissue transglutaminase may serve as a simple marker for apoptotic epithelial cells in tissue sections. PMID- 10857484 TI - Organization of desmin-containing intermediate filaments during differentiation of mouse decidual cells. AB - Decidualization of the mouse endometrium consists of a redifferentiation of the endometrial stromal fibroblasts. During decidualization these fibroblasts undergo growth, change of shape, multinucleation, and establishment of intercellular junctions. One feature of rodent decidual cells is the accumulation of intermediate filaments. In spite of the fact that fibroblasts normally have vimentin intermediate filaments, they acquire a large amount of desmin intermediate filaments while they undergo decidualization. The light and electron microscope immunocytochemical results of the present work show that during the initial stages of decidual transformation the desmin intermediate filaments accumulate around the nuclei, often forming caps around the nuclear envelope. As the decidual cells grow, the filaments form bundles and nets that radiate from the nuclei toward the cell surface. During the final stages of differentiation, on day 8 of pregnancy, staining of differentiated decidual cells decreases and most filaments accumulate under the cell surface. A role for intermediate filaments is suggested for decidualization of mouse endometrial cells. PMID- 10857485 TI - Carcinogenesis of primary liver malignancies. AB - Hepatocellular carcinoma (HCC) is one of the most common tumors in humans today, and its incidence has been rising over recent decades in Western countries. The main risk factors for the development of HCC are viral infections (hepatitis B and C), alcohol use, and the intake of mycotoxins in some areas of the world. In addition to these risk factors, genetic disorders such as hemochromatosis or alpha1-antitrypsin deficiency play a role. Regardless of the etiology, HCC generally develops on the basis of liver cirrhosis. Various pathomechanisms are effective during the neoplastic transformation of the hepatocyte. In chemical experimental models, hepatocarcinogenesis occurs in three stages: initiation, progression, and promotion. Molecular mechanisms of HCC development include the alteration in growth factor and tumor suppressor gene expression, dysregulation of apoptosis, and occurrence of reactive oxygen species generated during inflammation. PMID- 10857486 TI - Impact of virtual reality imaging on hepatic liver tumor resection: calculation of risk. AB - The risk involved in partial liver resections depends mainly on tumor localization, invasion of central vascular structures, and parenchymal function. The imaging techniques available today (computed tomography, magnetic resonance imaging) allow us to detect precisely the extent of tumor invasion and their relationship to central vessels. The various three-dimensional reconstruction techniques are helpful with regard to a virtual planning of liver resections. The calculation of remaining liver volumes subsequent to partial hepatectomies are considered to be an essential predictive parameter in terms for the development of postoperative liver failure. In a retrospective and a later consecutive, prospective clinical study we analyzed the postoperative risk in a series of 570 patients. In an univariate analysis 13 of 31 parameters showed significant values. In multivariate analysis only three parameters (partial hepatic resection rate, PHRR), gamma-glutamyltranspeptidase, and prothrombin activity) were independent parameters for predicting liver failure, generating the most significant values for the PHRR. In our experience the most comfortable and precise technique for evaluating PHRR is the b-spline technique. PMID- 10857487 TI - Hepatoma of the liver--resection or transplantation? AB - INTRODUCTION: Despite recent advances with techniques of in situ tumor ablation, surgical therapy remains at present the mainstay in the treatment of primary hepatic malignancies. DISCUSSION: After an initial endeavor to establish liver transplantation as a treatment option, especially for unresectable liver tumors, only a few indications, for example early hepatocellular carcinoma in cirrhosis, are currently agreed upon. Other indications, such as peripheral cholangiocarcinoma and hepatocellular carcinoma in non-cirrhotics, have largely been abandoned or are still under debate, as with fibrolamellar carcinoma. Selection of patients suffering from hepatocellular carcinoma in cirrhosis for liver transplantation is still based on tumor size and node number, because the current state of diagnostic imaging fails to reliably predict the most important prognostic parameter: vascular infiltration. Other selection criteria are under investigation. Studies on multimodal therapy are also underway but have not yet demonstrated a benefit. PMID- 10857488 TI - Ex-vivo resection techniques in tissue-preserving surgery for liver malignancies. AB - Some primary and secondary liver tumours are not absolutely irresectable, but cannot be resected using a conventional approach because of the limited warm ischaemia tolerance of the liver or poor accessibility of the tumour region. In such situations, the techniques of ex vivo liver surgery, pioneered by Rudolf Pichlmayr some 10 years ago, offer new chances for R0 resection. All the three different approaches, namely "in situ"-, "ante situm"-, and "ex situ" resection, require the use of measures originally developed for transplantation, such as hypothermic liver perfusion and veno-venous bypass. They differ mainly in the extent to which major vessels are divided in order to achieve optimal mobility of the organ. The results show that radical resection can be achieved accomplished in many cases. If necessary, complex vascular reconstructions can be performed. Although perioperative morbidity and mortality are high, there are a number of long-term survivors. Tumour recurrence, however, remains the main problem over the long term. In conclusion, ex vivo liver surgery is an important extension of surgical treatment possibilities. However, the procedure is suitable only for a small number of carefully selected patients and should be reserved for use in specialised centres. Furthermore, in view of the fact that the results are not yet optimal, additive and adjuvant treatment modalities are needed. PMID- 10857489 TI - Results after nonsurgical local treatment of primary liver malignancies. AB - Although it is generally agreed that surgical resection is the only curative treatment for liver malignancy, this is not possible in all cases. In situ methods for destruction of liver tumors are now available. We describe recently developed methods, with an up-date based on the literature and our own experience. Under ultrasound guidance chemical agents such as ethanol, hot saline, and acetic acid may be injected into the tumor, or thermally mediated techniques involving cryotherapy, microwaves, radiofrequency, or interstitial laser photocoagulation may be applied. Also, various ways of delivering radiotherapy to primary liver cancer have been developed. Although there are few randomized studies to compare these local, nonsurgical techniques, they have been shown to improve survival and in some cases to render tumors resectable. The long term results will show the real benefits of these newer treatments, particularly in high-risk patients. PMID- 10857490 TI - Surgical management of gastrointestinal stromal tumors of the stomach. AB - BACKGROUND: Gastrointestinal stromal tumors (GISTs) are rare tumors of the GI tract with varying degree of dignity and prognosis. Intramural or extragastral growth of gastric GISTs is associated with diagnostic difficulties and uncertainty about the type and extent of surgical therapy. Based on our experience, we tried to formulate management guidelines for gastric GISTs. METHODS: Five patients with gastric GIST (36-85 years old) underwent subserosal excision with subsequent B-II resection (1x), full-thickness partial gastric resection (2x), gastrotomy with submucosal excision (1x), or gastrectomy for carcinoma with an incidental finding of a leiomyoma (1x). RESULTS: Tumor size ranged from 2x2x1 cm to 9x6x4 cm. These tumors were classified as epithelioid leiomyosarcoma (1x), GIST (3x), or leiomyoma (1x). The prognosis of risk ranged from no risk (leiomyoma) to low-malignancy (leiomyosarcoma) depending on tumor size and mitotic index. No recurrent disease has been noted so far during follow up ranging from 3 months to 6 years. CONCLUSIONS: Staging of gastric disease should include the probability of gastric GIST. Surgical resection is the therapy of choice for potential malignant GISTs to ensure a local radical removal. Metachronic metastases should be resected if possible. Depending on tumor stage and prognostic parameters, an individual follow-up with endoscopic and radiologic examinations is recommended. Further studies should be undertaken to elaborate prognostic determinants and stage-adapted treatment. PMID- 10857491 TI - Evaluation of the effects of a pneumoperitoneum with carbon dioxide or helium in a porcine model of endotoxemia. AB - BACKGROUND: The expansion of the laparoscopic techniques to the critically ill patient is currently under debate. In order to evaluate the potential risks of performing laparoscopy in a body with signs of sepsis, the effects of the pneumoperitoneum were studied in a porcine model of mild endotoxemia. METHODS: Twenty-eight pigs were separated into four groups of seven animals: untreated control (C), 2 microg/kg/h endotoxin (E), endotoxin and a pneumoperitoneum (3 h, 15 mmHg) with CO2 (EC) or with helium (EH). Hemodynamic and homeostatic variables were studied for a period of 5.5 h. Primary endpoints were arterial and mucosal pH and the ATP content of the bowel wall. Statistical evaluation was performed using analysis of variance and the Bonferroni test. RESULTS: Endotoxin infusion induced characteristic symptoms of early sepsis: increase of arterial CO2, pulmonary arterial, portal venous, and pulmonary artery wedge pressure, and decrease of arterial pressure, cardiac output, arterial and mucosal pH. An additional pneumoperitoneum led to aggravation of all criteria with significant alterations in arterial and mucosal pH, arterial CO2, wedge and portal venous pressure. The most striking derangement of mean values was observed for mucosal pH (EC: 7.40, EH: 7.54) and arterial pH (EC: 7.15, EH: 7.18). In group EC, two animals died in septic shock. CONCLUSION: Applying a pneumoperitoneum during an ongoing sepsis significantly deteriorates hemodynamic and homeostatic variables, thus enhancing the risk of severe complications. PMID- 10857492 TI - Malignant hemangiopericytoma of the bone. AB - Hemangiopericytoma is a rare vascular tumor of pericyte origin with variable malignant potential. Very rarely, this tumor occurs as a primary bone lesion. We present a case of a highly malignant hemangiopericytoma of the proximal tibia. Current therapy consists of radical resection of the tumor with postoperative radiation therapy being recommended. Chemotherapy seems to be useful in disseminated disease. The prognosis correlates to the histological grading of the tumor. Early or late recurrence and distant metastases with fatal outcome, as shown in our case study, are not uncommon. PMID- 10857493 TI - High- or low-potassium solutions for the storage of abdominal and thoracic organs. AB - BACKGROUND: Clinically, intracellular type solutions are the most widely used solutions to preserve organs. The optimal ion composition of preservation solutions, however, is still unknown and extracellular-type solutions have frequently been superior to intracellular solutions in various experimental studies. MATERIALS AND METHODS: In this study, we measured extracellular (interstitial) electrolyte concentrations in rat livers, kidneys, hearts and lungs at 4 degrees C by means of microdialysis sampling. RESULTS: After 24 h cold ischaemia, [Na+]int and [K+]int were 104 +/- 25 mmol/l and 6.5 +/- 0.7 mmol/l in hearts, 92 +/- 12 mmol/l and 6.9 +/- 1.0 mmol/l in livers, 115 +/- 22 mmol/l and 6.3 +/- 0.9 mmol/l in kidneys and 87 +/- 17 mmol/l and 6.4 +/- 0.6 mmol/l in lungs. After preservation of organs in intracellular-type solutions, [Na+]int was significantly lower for each organ (range from 69 +/- 8 mmol/l to 73 +/- 20 mmol/l) and [K+]int was significantly higher (range from 8.0 +/- 1.7 mmol/l to 9.8 +/- 1.0 mmol/l). In no instance did the interstitial electrolyte concentration equilibrate with the intracellular electrolyte concentration. When the diffusion gradient from the vascular space to the interstitial space was calculated for Na+ and K+, a significantly higher barrier was found for K+ than for Na+ (P<0.001 and P<0.01 for hearts). CONCLUSIONS: These studies indicate that during cold storage of rat hearts, lungs, livers and kidneys, intra- and extracellular electrolytes do not equilibrate. Ion exchange stabilises at extracellular Na+ concentrations between 87 mmol/l and 115 mmol/l and K+ concentrations between 6.3 mmol/l and 6.9 mmol/l. Storage of organs in solutions with extracellular-type ion compositions might improve graft function and survival not only after lung and liver but also after heart and renal preservation. PMID- 10857494 TI - Enhanced cytolytic activity of intestinal intraepithelial lymphocytes in patients with Crohn's disease. AB - BACKGROUND AND AIMS: Dysfunction of the immune system with inappropriate responses of lymphocytes to various antigens has been implicated in the development of Crohn's disease. Therefore, the functional and phenotypic characteristics of intestinal intraepithelial lymphocytes (IEL) in comparison to peripheral blood lymphocytes (PBL) were analyzed in patients with and without Crohn's disease. PATIENTS AND METHODS: Six patients with Crohn's disease and six control patients were studied. Isolated IEL and PBL were tested for cytolytic activity against the human adenocarcinoma cells DLD-1 and the human leukemia cells K562 in a 51Cr-release assay. Two-color flow cytometry was performed for phenotype analysis of isolated lymphocytes. RESULTS: IEL from patients with Crohn's disease showed significantly increased cytolytic activity against epithelial-derived target cells when compared with IEL from control patients. In contrast, no functional changes were detectable among PBL from patients with Crohn's disease. IEL from patients with Crohn's disease contained a significantly higher percentage of CD8+ lymphocytes when compared with IEL from control patients, whereas no phenotypic changes were observed among PBL. CONCLUSIONS: In Crohn's disease, the functional and phenotypic changes of T cells are limited to lymphocytes of the intestinal mucosa. Furthermore, it is conceivable that the increased cytolytic activity of IEL contributes to the tissue damage in this disease. PMID- 10857495 TI - Tumor volume: a novel prognostic factor in patients who undergo curative resection for gastric cancer. AB - BACKGROUND AND AIMS: The present study evaluates the significance of tumor volume as a prognostic factor in gastric cancer. PATIENTS/METHODS: Tumor volume was measured from serial tissue sections of 101 patients who had undergone curative resection for solitary carcinoma of the stomach using a computer graphics analysis program. These patients were analyzed with respect to survival based on univariate and multivariate analyses of clinicopathological factors, including tumor volume, to determine an independent prognostic factor. RESULTS: Significant differences in survival were found with respect to depth of tumor invasion (P=0.001), status of lymph-node metastasis (P=0.018), tumor diameter (P=0.005), and tumor volume (P<0.0001) based on univariate analysis. However, multivariate analysis indicated only tumor volume as a valid factor in determining prognosis among the nine variables and was significantly associated with the prognosis (P=0.0005; relative risk 18.23; 95% confidence interval 3.52-94.37). CONCLUSION: The present findings indicate that tumor volume is an important prognostic factor in patients who undergo curative resection for gastric cancer and may be an alternative to conventional factors, thus providing a novel independent prognostic factor in gastric cancer. PMID- 10857496 TI - Duodenum-preserving pancreatic head resection in patients with benign and borderline tumors of the pancreatic head. AB - BACKGROUND: Benign tumors of the pancreas are rare, accounting for only 1-2% of primary pancreatic lesions. Up to now, partial duodenopancreatectomy is still one of the established forms of treatment of benign tumors of the pancreas. We applied duodenum-preserving pancreatic head resection in 12 patients with benign pancreatic tumors to evaluate the feasibility, morbidity and recurrence rates after this less aggressive method. METHODS: Between April 1984 and December 1999, 12 patients with benign and borderline tumors of the pancreatic head were operated on by duodenum-preserving pancreatic head resection. RESULTS: All five patients with serous cystadenoma are free of recurrence 4.4 years after primary resection. One of two patients with mucinous cystadenoma and one of three patients with intraductal papillary mucinous tumor developed recurrent tumor within the former pancreatic head 5 years and 6 years, respectively, after the primary operation. Both patients were resected a second time. One of two patients with gastrinoma still has elevated serum gastrin levels. There was no hospital or long-term mortality. CONCLUSION: For a symptomatic serous cystadenoma, duodenum preserving pancreatic head resection is a good alternative to partial duodenopancreatectomy. In borderline tumors with malignant potential, we would rather suggest a more radical duodenum-preserving segmental resection. A video clip (3 min) is attached demonstrating the basic steps of duodenum-preserving pancreatic head resection. PMID- 10857497 TI - Effects of interferon therapy on inhibition of hepatocellular carcinoma development in patients with chronic hepatitis C. PMID- 10857498 TI - An evaluation of tissue polypeptide antigen (TPA) in the two bronchoalveolar lavage fractions of lung cancer patients. AB - BACKGROUND: It has been proven that cytokeratins (CKs) are useful tumor markers for the follow-up, treatment monitoring and prognosis evaluation of lung cancer and among these, tissue polypeptide antigen (TPA) plays an important role. Nevertheless, only a small number of studies have been reported about their diagnostic capacity. Bronchoalveolar lavage (BAL) can be divided into two fractions: bronchiolar (BF) and alveolar (AF). For the above reasons, our aims were (1) to analyze the diagnostic usefulness of TPA in the BAL of lung cancer patients and (2) to observe if, in lung cancer patients, TPA levels in the two BAL fractions are different. This should mean that the study of tumor markers in the BAL should be carried out in both fractions to increase their diagnostic capacity. METHODS: We studied 289 BALs divided into two phases. In phase I, TPA was analyzed in the BAL of six groups of subjects (healthy persons, chronic bronchitis, asthma, respiratory infections, diffuse interstitial pulmonary diseases and lung cancer). In phase II, TPA was studied in both BAL fractions of a group of patients with lung cancer. RESULTS: We observed that TPA levels were significantly higher in the BAL of patients with bronchogenic neoplasias. In these patients, TPA was increased in the BF of the lavage in relation to the AF. In smoker patients with pulmonary carcinomas, TPA was higher in the AF of the BAL than in the lavage of non-smokers. This did not occur in the BF. We found no relation between the TPA concentrations and cancer histology. CONCLUSIONS: We believe that TPA is a useful tumor marker with diagnostic capacity and this capacity is increased when it is studied in the two BAL fractions. Smoking habit may play a role in the secretion of tumor markers by the tumor cells. PMID- 10857499 TI - Suicidal ideation in cancer patients with major depression. AB - BACKGROUND: Major depression is a well-documented risk factor for suicide in cancer patients as well as in the general population. However, there are no data explaining why some cancer patients suffering from major depression have suicidal ideation whereas others with the same disorder do not. METHODS: We investigated the background differences between cancer patients suffering from major depression with and without suicidal ideation by analyzing consultation data on patients referred to the Psychiatry Division, National Cancer Center Hospital, Japan. RESULTS: Univariate analysis showed that poor performance status, advanced stage and severe depression were the risk factors for suicidal ideation in these depressive cancer patients. Multivariate logistic regression analysis indicated that older age and severe depression are the final significant risk factors. CONCLUSION: These preliminary findings suggest that severe major depression may be an important indicator of suicidal ideation among cancer patients, that patients' age should be given greater attention and that closer monitoring and more intensive intervention against major depression may be needed. PMID- 10857500 TI - The relationship between technical parameters of external beam radiation therapy and complications for localized prostate cancer. AB - BACKGROUND: This study was performed to review retrospectively the clinical course of chronic rectal bleeding as a complication of external beam radiation therapy for localized prostate cancer and to analyze the relationship between technical parameters of radiation therapy and the complications. METHODS: Seventy one patients with stages A2, B and C were treated with local-field radiotherapy (total dose 52.5-66 Gy, daily dose 2.0-3.28 Gy, field area 30-81 cm2, number of fields 3-15 ports, planning simulations X-ray or CT-based) between 1989 and 1998 at three institutions. The protocols were consistent during this same period at these institutions. RESULTS: Multivariate analysis revealed pretreatment PSA and Gleason sum to be statistically significant predictors of 5 year prostatic specific antigen (PSA) relapse-free rates in a median follow-up period of 42 months (range 12-119 months). The significant risk factors for higher grading of acute morbidity were a biological equivalent dose, alpha/beta = 10(BED10) > or =65 Gy, dose per fraction > or =3.0 Gy, field area > or =42 cm2, fewer ports and X-ray planning simulation. However, no parameter was associated with higher grading of late morbidity. Eleven patients (15.4%) experienced a late GI complication: grade 1 (4.2%), grade 2 (9.8%), grade 3 (1.4%). The median time to occurrence of rectal bleeding was 12 months after radiotherapy and the mean duration of morbidity was 11 months. CONCLUSIONS: Higher total dose and dose per fraction, larger field area, fewer ports and X-ray simulation increased the grades of acute morbidity. A majority of chronic rectal bleedings were transient and responded to conservative treatment. PMID- 10857501 TI - Causes of interruption of radiotherapy in nasopharyngeal carcinoma patients in Taiwan. AB - BACKGROUND: Nasopharyngeal cancer (NPC) is now curable with early diagnosis and radiotherapy treatment. In the past several decades, few studies have investigated why some patients fail to complete the recommended full course of radiotherapy. METHODS: A total of 3273 nasopharyngeal carcinoma patients were treated at the Radiation Oncology Department of Linkou Chang Gung Memorial Hospital in a span of 18 years from 1979 to 1996. Among these patients, 276 did not complete the full course of treatment of radiation therapy. The medical records of these patients were reviewed to determine the factors contributing to treatment interruption. RESULTS: Of the 276 patients whose treatment was interrupted, 120 (43.5%) were unable to endure the acute side effects of radiation therapy and were afraid of the possible complications resulting from the treatment; 57 (20.7%) had doubts about the diagnosis or had the subjective perception that the treatment offered would be ineffective in view of the severity of their disease; 50 (18.1%) resorted to folk prescriptions; 17 (6.2%) were faced with socioeconomic problems; 15 (5.4%) sought treatment at another hospital owing to transport considerations; 10 (3.6%) stopped radiation therapy and switched to chemotherapy for palliative management; seven (2.5%) resorted to praying, god worshipping and taking incense powder and magic elixirs because their families were against any established therapy. CONCLUSIONS: The acute side effects and complications caused by radiation therapy were the major factors influencing patients' decisions to discontinue treatment. This finding suggests that more attention should be paid to providing care with regard to the acute side effects of radiotherapy and to reinforcing pretreatment education. PMID- 10857502 TI - A case of thyroid cancer involving the trachea: treatment by partial tracheal resection and repair with a latissimus dorsi musculocutaneous flap. AB - A 65 year-old man had undergone left thyroidectomy for thyroid cancer. The cancer had directly invaded the cervical esophagus and trachea and the patient was referred to our hospital for radical resection and reconstruction. Cervical computed tomography showed a mass at the left-posterior wall of the trachea. Cervical esophagectomy, resection of the left half of the trachea (6 x 3 cm) including seven rings and cervical lymph node dissection were performed. The tracheal defect was covered by a latissimus dorsi musculocutaneous flap. The patient did not lose vocal function and remains alive and well 3 years after surgery without any evidence of recurrence. Latissimus dorsi muscle flap coverage of tracheal defects seems to be a useful technique in the combined resection of the trachea. PMID- 10857503 TI - Spermatic cord metastases from gastric cancer with elevation of serum hCG-beta: a case report. AB - Spermatic cord metastases from gastric cancer are rare. We here document a case involving a gastric cancer that mimicked primary testicular tumor because of elevation of the serum human chorionic gonadotropin-beta (hCG-beta). The possibility of metastasis or recurrence of prior malignancies should therefore be considered when the clinical features described here are encountered, although elevation of hCG-beta is rare with tumors other than those in testis. PMID- 10857504 TI - Local recurrence of renal cell carcinoma causing massive gastrointestinal bleeding: a report of two patients who underwent surgical resection. AB - We report two cases of local recurrence of renal cell carcinoma, which invaded the gastrointestinal tract. The intervals to local recurrence after primary nephrectomies were 5 and 13 years. Both cases developed massive gastrointestinal bleeding. Blood transfusions and arterial embolization were performed; however, continuous melena prevented improvement in their general condition. Although distant metastases were observed in both cases, tumor resections combined with intestine invasion were performed. Although this surgical approach does not significantly improve the prognosis, even in patients who have remaining metastatic lesions, it may provide palliation and improve the quality of survival. PMID- 10857505 TI - The role of quality of life assessment in gastric cancer. PMID- 10857506 TI - Recent topics in breast cancer. PMID- 10857507 TI - Colorectal cancer mortality rates by prefectures in Japan. PMID- 10857508 TI - Early versus delayed inpatient stroke rehabilitation: a matched comparison conducted in Italy. AB - OBJECTIVE: To assess the specific influence of onset-admission interval (OAI) on rehabilitation results. DESIGN: A case-control study in consecutive stroke inpatients, enrolled in homogeneous subgroups, matched for age (within 1 year) and Barthel Index (BI) score at admission, and different for OAI to the rehabilitation ward. The short OAI group began rehabilitation treatment within the first 20 days from stroke, medium OAI group between days 21 and 40, and long OAI between days 41 and 60. SETTING: Rehabilitation hospital. PATIENTS: One hundred forty-five patients with sequelae of first stroke. MAIN OUTCOME MEASURES: Efficiency (average increase in BI per day), effectiveness (proportion of potential improvement achieved during rehabilitation) of treatment, and percentage of low- and high-response patients, calculated on BI, were evaluated. Odds ratios (ORs) of dropouts and of poor and excellent therapeutic response were also quantified. RESULTS: The short OAI subgroup had significantly higher effectiveness of treatment than did the medium (p < .05) and the long OAI groups (p < .005). Beginning treatment within the first 20 days was associated with a significantly high probability of excellent therapeutic response (OR = 6.11; 95% confidence interval [CI], 2.03-18.36), and beginning later was associated with a similar risk of poor response (OR = 5.18; 95% CI, 1.07-25.00). On the other hand, early intervention was associated with a five times greater risk of dropout than that of patients with delayed start of treatment (OR = 4.99; 95% CI, 1.38-18.03). The three subgroups were significantly (p < .05) different regarding the percentage of low and high responders. CONCLUSION: Our results showed a strong association between OAI and functional outcome. PMID- 10857509 TI - Driving skills in elderly persons with stroke: comparison of two new assessment options. AB - OBJECTIVE: To compare the effectiveness of two methods of assessing off-road driving skills that claim to predict on-road driving fitness of persons with stroke. METHOD: Fifty-six persons with stroke (age 44 to 82 yrs; mean, 60.2 yrs) completed the 2 off-road driving assessments along with standard clinical and on road driving tests. MAIN OUTCOME MEASURES: Linear stepwise regression on 4 variables of the Dynavision Performance Assessment Battery (DPAB), the Cognitive Behavioral Driver's Inventory (CBDI) variable (composite score), and the variables of age, gender, and lesion side. RESULTS: A 4-minute endurance subtest from the DPAB was superior to the CBDI in predicting success/failure in the on road driving test (75%). However, success on both the 4-minute endurance subtest from the DPAB and the CBDI tests significantly improved the prediction of on-road success. If participants passed the CBDI and the endurance test from the DPAB, they also passed the on-road assessment. CONCLUSION: Driving fitness of elderly persons with stroke can be assessed with reasonable accuracy using off-road tests, minimizing the expense and risk associated with on-road assessments in this population. PMID- 10857510 TI - Perceptual differences between stroke patients with cerebral infarction and intracerebral hemorrhage. AB - OBJECTIVE: To assess perceptual performances of patients with intracerebral hemorrhage (ICH) compared with those of ischemic patients early after stroke and to analyze the psychometric properties of three perceptual tests used in the study. DESIGN: Cross-sectional study. SETTING: A rehabilitation unit at a teaching hospital. PATIENTS: Twenty-two stroke patients with ICH and 22 demographically matched stroke patients with infarction. MAIN OUTCOME MEASURES: Loewenstein Occupational Therapy Cognitive Assessment (LOTCA), Rivermead Perceptual Assessment Battery (RPAB), and Motor-Free Visual Perception Test (MVPT). RESULTS: Stroke patients with ICH had significantly more severe deficits on a task of thinking operations than did patients with infarction. A significant lateralized effect of stroke existed in the ICH group, with patients with right hemisphere strokes scoring lower than patients with left-hemisphere strokes on the figure-ground discrimination subtest of the RPAB. A considerable overlap among the three instruments was found. Yet, the observed correlations between supposedly similar subtests from the tests proved to be moderate, indicating that to a certain extent these test measures tap different perceptual processes. Four factors were generated from a joint LOTCA-RPAB-MVPT factor analysis. They assessed different facets of perceptual functioning, including higher-level and lower-level perceptual skills, part/whole conceptual integration, and color perception. This factor pattern accounted for 75.5% of the variance. CONCLUSIONS: Higher-level perceptual functions tend to be relatively susceptible to ICH stroke pathology early in the course of the disease. This information has important clinical implications in the early treatment planning for the stroke patients with ICH, such that specific compensatory strategies for these deficiencies should be devised to facilitate a successful rehabilitation. Knowledge regarding the influences of specific deficits on the performance of daily activities may also be useful to the patients' family. PMID- 10857511 TI - Medicare's payment system: its effect on discharges to skilled nursing facilities from rehabilitation hospitals. AB - OBJECTIVE: To determine if Medicare's payment system for rehabilitation hospitals encourages discharges to skilled nursing facilities (SNFs). Medicare payments to hospitals are based on limits derived from a hospital's average allowable patient charge during a base year. Thereafter, payments are capped, but hospitals receive additional incentive payments if succeeding costs are reduced. It was a hypothesis of this study that discharges to SNFs would increase after the base year. In this way, rehabilitation hospitals would limit high-cost patients when under reimbursement limitation. METHODS: Medicare claims data for 162,239 discharges from 69 rehabilitation hospitals between 1987 and 1994 were analyzed. After controlling for patient and provider characteristics, we compared the odds of being discharged to a SNF before, during, and after the base year. RESULTS: Before and during the base year, 4.7% and 6.6% of patients were discharged to a SNF. After the base year, 9% of patients were sent to a SNF. After controlling for temporal and seasonal trends, as well as for patient and provider characteristics, those discharged after the base year were significantly more likely to be sent to a SNF than those discharged during the base year. These odds increased with increasing length of stay in the rehabilitation hospital. For those with a length of stay of 29 days (75th percentile) the odds increased by 11% (odds ratio, 1.11; 95% confidence interval, 1.04-1.18). CONCLUSIONS: The incentives of Medicare's reimbursement system may encourage an increase in the percentage of patients discharged to SNFs after the base year. These findings have significant implications regarding the structure of Medicare's prospective payment system currently planned for this class of hospital. PMID- 10857512 TI - Searching for rehabilitation articles on MEDLINE and EMBASE. An example with cross-over design. AB - OBJECTIVE: To analyze the usefulness of MEDLINE and EMBASE biomedical databases in rehabilitation and to identify descriptors and text words necessary to do a comprehensive search. METHODS: We looked for articles published since 1990 relating to neurologic, orthopedic, respiratory, urologic, and rheumatologic rehabilitation. We looked for all descriptors and text words pertinent to rehabilitation and linked them with "cross-over." RESULTS: We found 165 articles in MEDLINE and 159 in EMBASE with an overlap of only 17% of articles. Only 32% of the articles in MEDLINE and 35% in EMBASE were relevant. Of the 214 nonoverlapping articles, 84% were published in journals present in both databases, but were indexed differently. CONCLUSION: At least two databases must be used to ensure a comprehensive literature search. Searching in EMBASE after MEDLINE we gained 25 articles (32%). Bibliographic search in rehabilitation is particularly complex because of the heterogeneity of the subject matter. Cooperation between an information professional and a clinician is essential to ensure a comprehensive search. PMID- 10857513 TI - The Westmead Pediatric TBI Multidisciplinary Outcome study: use of functional outcomes data to determine resource prioritization. AB - OBJECTIVE: To measure functional outcome in the 2 years after traumatic brain injury (TBI) in 2 groups of children and to determine the usefulness of a TBI severity classification system for resource allocation. DESIGN: Prospective inception cohort study with 3 assessment points during the 2 years after trauma. SETTING: Tertiary pediatric trauma center in Sydney, Australia. PARTICIPANTS: Eighty-one consecutive admissions aged 0 to 14 years. Fifty-one were allocated to the Mild (n = 26) or Severe (n = 25) TBI groups, according to preset determinants of severity; 30 admissions with non-TBI trauma constituted the control group. MAIN OUTCOME MEASURES: Standardized psychometric and clinical assessments of cognition, communication and feeding ability, motor performance (ambulation, fine and gross motor), neurologic status, self-care independence, and school/academic performance. RESULTS: Those with Mild TBI severity had no significant deficits at the 2-year data point. In contrast, those in the Severe TBI group demonstrated continued problems with fine motor performance, neurologic status, self care, and school/academic performance. CONCLUSIONS: A classification system has been developed that may be useful in the allocation of children with a TBI, age younger than 15 years, to 1 of 2 severity groups early in their rehabilitation. This classification system may be useful in determining areas of high and low resource prioritization. PMID- 10857514 TI - Kinetic alterations independent of walking speed in elderly fallers. AB - OBJECTIVES: To determine if joint kinetic gait alterations in fallers persist when they attempt to walk at a faster speed that is more comparable with nonfallers' comfortable walking speed. DESIGN: Retrospective, case-control study. Stereophotogrammetric and force platform data were collected. SETTING: A gait laboratory. PARTICIPANTS: Sixteen elderly subjects who had at least 2 falls in the last 6 months from an unclear cause and 23 elderly subjects with no history of repeated falls. MAIN OUTCOME MEASURES: Differences in all major peak joint kinetic (moment and power) values during the gait cycle between elderly nonfallers walking at comfortable speed and elderly fallers walking at (1) comfortable and (2) fast speed. RESULTS: Statistically significant differences present at both comfortable and fast walking speeds were present in 4 sagittal plane parameters. There was an increase in peak external hip flexion moment in stance, a reduction in peak hip extension moment, a reduction in knee flexion moment in preswing, and a reduction in knee power absorption in preswing. CONCLUSION: The presence and persistence of 4 specific alterations in sagittal plane joint kinetics at both comfortable and fast walking speeds imply specific intrinsic pattern differences and allow for new insights into the mechanics of gait in elderly people who fall. The presence of these alterations also suggests they may serve as potential identifiable markers to detect those who may be at risk for falls. PMID- 10857515 TI - Prediction of walking function in stroke patients with initial lower extremity paralysis: the Copenhagen Stroke Study. AB - OBJECTIVES: The majority of stroke patients with initial leg paralysis do not regain independent walking. We characterize the minority who, despite initial leg paralysis, regained independent walking. DESIGN: Consecutive and community based. SETTING: A stroke unit receiving all stroke patients from a well-defined community. PATIENTS: A total of 859 acute stroke patients; 157 (15%) initially had leg paralysis. MAIN OUTCOME MEASURES: Scandinavian Stroke Scale (SSS) and Barthel index (BI) on admission and weekly during rehabilitation. Univariate and multivariate statistics were considered. RESULTS: Of the 157 patients with initial leg paralysis, 84 (60%) died; 73 (40%) survived. Fifteen (21%) survivors regained walking function (the walking group), and 58 (79%) did not (the nonwalking group). The BI on admission was the only factor of significant predictive value (p < .03). Mean admission BI was 50 in the walking group versus 3 in the nonwalking group (p < .001). Age, gender, lesion size, total SSS score, and comorbidity had no predictive value. Within the first week, the walking group gained 3.2 points in the SSS subscore for leg strength versus 0.5 points in the nonwalking group (p < .02). CONCLUSION: Only 10% of stroke patients with initial leg paralysis regained independent walking. In these patients, BI on admission was high and leg strength improved quickly in the first week. PMID- 10857516 TI - Multidisciplinary treatment of chronic pain patients: its efficacy in changing patient locus of control. AB - OBJECTIVE: To assess the efficacy of multidisciplinary treatment in altering chronic pain patient locus of control beliefs. DESIGN: A before-and-after treatment design including demographics. PARTICIPANTS: Seventy-three chronic nonmalignant pain patients who completed study questionnaires both before and after treatment. SETTING: Comprehensive, outpatient, multidisciplinary pain management program at a large Midwestern university medical center. MAIN OUTCOME MEASURES: Pain Locus of Control Scale and Survey of Pain Attitudes Control subscale. RESULTS: Patients' perceptions of personal control over pain increased from pretreatment to posttreatment, and patients' perceptions of external control over pain, such as fate or powerful others. decreased from pretreatment to posttreatment. CONCLUSIONS: This study supports the efficacy of chronic pain management centers in altering patient beliefs about pain. The ability to increase patients' self-efficacy in their control over pain and to decrease external attributions are essential to successful pain management. PMID- 10857517 TI - Therapeutic selective nerve root block in the nonsurgical treatment of atraumatic cervical spondylotic radicular pain: a retrospective analysis with independent clinical review. AB - OBJECTIVE: To investigate the outcomes resulting from the use of fluoroscopically guided therapeutic selective nerve root block (SNRB) in the nonsurgical treatment of atraumatic cervical spondylotic radicular pain. STUDY DESIGN: Retrospective study with independent clinical review. PARTICIPANTS: Twenty subjects (10 men, 10 women) with mean age 56.6 years. METHODS: Each patient met specific physical examination, radiographic, and electrodiagnostic criteria to confirm a level of cervical involvement. Those patients whose root level remained indeterminate were required to demonstrate a positive response to a fluoroscopically guided diagnostic SNRB prior to the initiation of treatment. Therapeutic injections were administered in conjunction with physical therapy. Data collection and analysis were performed by an independent clinical reviewer. MAIN OUTCOME MEASURES: Pain score, work status, medication usage, and patient satisfaction. RESULTS: Twenty patients with an average symptom duration of 5.8 months were included. An average of 2.2 therapeutic injections was administered. Follow-up data collection transpired at an average of 21.2 months following discharge from treatment. A significant reduction (p = .001) in pain score was observed at the time of follow up. Medication usage was also significantly improved (p = .005) at the time of follow-up. An overall good or excellent result was observed in 60%. Thirty percent of patients required surgery. Younger patients were more likely (p = .0047) to report the highest patient satisfaction rating following treatment. CONCLUSIONS: This study suggests that fluoroscopically guided therapeutic SNRB is a clinically effective intervention in the treatment of atraumatic cervical spondylotic radicular pain. PMID- 10857518 TI - Respiratory muscle weakness and respiratory muscle training in severely disabled multiple sclerosis patients. AB - OBJECTIVE: To evaluate the contribution of respiratory muscle weakness (part 1) and respiratory muscle training (part 2) to pulmonary function, cough efficacy, and functional status in patients with advanced multiple sclerosis (MS). DESIGN: Survey (part 1) and randomized controlled trial (part 2). SETTING: Rehabilitation center for MS. PATIENTS: Twenty-eight bedridden or wheelchair-bound MS patients (part 1); 18 patients were randomly assigned to a training group (n = 9) or a control group (n = 9) (part 2). INTERVENTION: The training group (part 2) performed three series of 15 contractions against an expiratory resistance (60% maximum expiratory pressure [PEmax]) two times a day, whereas the control group performed breathing exercises to enhance maximal inspirations. MAIN OUTCOME MEASURES: Forced vital capacity (FVC), inspiratory, and expiratory muscle strength (PImax and PEmax), neck flexion force (NFF), cough efficacy by means of the Pulmonary Index (PI), and functional status by means of the Extended Disability Status Scale (EDSS). RESULTS: Part 1 revealed a significantly reduced FVC (43% +/- 26% predicted), PEmax (18% +/- 8% predicted), and PImax (27% +/- 11% predicted), whereas NFF was only mildly reduced (93% +/- 26% predicted). The PI (median score, 10) and EDSS (median score, 8.5) were severely reduced. PEmax was significantly correlated to FVC, EDSS, and PI (r = .77, -.79, and -.47, respectively). In stepwise multiple regression analysis. PEmax was the only factor contributing to the explained variance in FVC (R2 = .60), whereas body weight (R2 = .41) was the only factor for the PI. In part 2, changes in PImax and PEmax tended to be higher in the training group (p = .06 and p = .07, respectively). The PI was significantly improved after 3 months of training compared with the control group (p < .05). After 6 months, the PI remained significantly better in the training group. CONCLUSIONS: Expiratory muscle strength was significantly reduced and related to FVC, cough efficacy, and functional status. Expiratory muscle training tended to enhance inspiratory and expiratory muscle strength. In addition, subjectively and objectively rated cough efficacy improved significantly and lasted for 3 months after training cessation. PMID- 10857519 TI - Resistive inspiratory muscle training: its effectiveness in patients with acute complete cervical cord injury. AB - OBJECTIVE: To evaluate if resistive inspiratory muscle training (RIMT) can improve lung function in patients with complete tetraplegia within half a year after trauma. DESIGN: A prospective study. The experimental patients received training with a Diemolding Healthcare Division inspiratory muscle trainer for 15 to 20 minutes per session, twice per day, 7 days a week for 6 weeks. SETTING: Hospital-based rehabilitation units. PATIENTS: Twenty patients who were in their first 6 months of complete cervical cord injury were randomly enrolled into RIMT (10 patients) and control (10 patients) groups. MAIN OUTCOME MEASURE: Spirometry, lung volume test, maximal inspiratory pressure, maximal expiratory pressure, and modified Borg scale measurements at rest were performed before training and at the end of 6 weeks of training. RESULTS: Most of the pulmonary parameters showed statistically significant improvements within the RIMT and control groups, but the improvements were greater in the RIMT group. In addition, the improvements in total lung capacity, total lung capacity predicted percentage, vital capacity, minute ventilation, forced expiratory volume in 1 second predicted percentage, and the resting Borg scale in the RIMT group showed significantly greater improvement. CONCLUSION: RIMT can improve ventilatory function, respiratory endurance, and the perceived difficulty of breathing in patients with complete cervical spinal cord injury within half a year after trauma. PMID- 10857520 TI - Pulmonary function in chronic spinal cord injury: a cross-sectional survey of 222 southern California adult outpatients. AB - OBJECTIVES: To evaluate risk factors for respiratory morbidity in chronic spinal cord injury (SCI). SETTING: Model SCI care system based at an urban public rehabilitation medical center. DESIGN: Case series with evaluation of pulmonary function by conventional spirometric testing. PARTICIPANTS: Two hundred twenty two adults with SCI of more than 1-year duration who were not chronically dependent on mechanical ventilation, including 98 with tetraplegia (62 with complete and 26 with incomplete motor lesions) and 124 with paraplegia (87 with complete and 37 with incomplete motor lesions). MAIN OUTCOME MEASURES: Forced vital capacity (FVC), forced expired volume in 1 second (FEV1), and peak expiratory flow rate (PEFR), all measured in the supine and erect seated positions and compared with predicted normal values for industrial workers. RESULTS: FVC and FEV1 were normal in persons with low-level paraplegia who had never smoked, but both decreased similarly with rising SCI level, more markedly in those with tetraplegia. PEFR decreased with rising SCI level. Incomplete lesions mitigated function loss in those with tetraplegia. In middle-aged individuals with tetraplegia, longer duration of injury was associated with greater function loss, independent of age. Current smokers showed excess function loss, except for those with high tetraplegia. Most people with complete tetraplegia showed FVC and FEV1 increases in the supine position relative to the erect position. CONCLUSIONS: Pulmonary function is compromised by most lesions of the spinal cord, even in those with paraplegia, and is affected relative to the level of lesion. Efforts to help SCI patients minimize respiratory complications in particular, assistance in smoking cessation-should be given high priority. PMID- 10857521 TI - Antimicrobial resistance in gram-negative bacteria isolated from the urinary tract in community-residing persons with spinal cord injury. AB - OBJECTIVE: To assess the epidemiology of antimicrobial resistance among community residing persons with spinal cord injury (SCI). DESIGN: Retrospective analysis of existing data. SETTING: Data were obtained from persons with SCI attending clinic for annual examinations. PARTICIPANTS: Two hundred eighty-seven SCI outpatients. INTERVENTION: None. MAIN OUTCOME MEASURE: Occurrence of bacteriuria with gram negative organisms demonstrating resistance to antimicrobial agents in 2 or more classes. RESULTS: There were 706 gram-negative isolates from 444 urine specimens. Resistance to drugs in 2 or more classes occurred in 33% of bacterial isolates, but did not significantly increase in frequency among those injured for longer periods or more severely. Significantly higher rates of multidrug-resistant bacteria occurred in specimens from males, younger age group (< or =45 yrs), and persons with indwelling and condom catheters. CONCLUSIONS: Antimicrobial resistance in outpatients with SCI is common and is related to widespread use of specific drugs, type of bladder management, and other host factors. PMID- 10857522 TI - Electrophysiologic evaluation of muscle fatigue development and recovery in late polio. AB - OBJECTIVES: To study aspects of fatigue in late-polio patients and healthy controls. We hypothesized that late-polio subjects would develop more peripheral fatigue, assessed with surface electromyography (EMG), and that no major differences would exist between the two groups in neuromuscular junction transmission. DESIGN: Case-control study. SETTING: University hospital laboratory. SUBJECTS: Ten patients with a history of polio (mean age, 54 yrs, SD = 5; mean time since polio onset, 49 yrs, SD = 7) and a matched control group (mean age, 52 yrs, SD = 8). METHODS: A protocol with a stepwise force increase up to 80% of maximal voluntary contraction ending with an 8-minute recovery period was performed twice, first with surface EMG and then with electrical stimulation and surface-recorded evoked M-response. MAIN OUTCOME MEASURES: Surface EMG analysis of voluntary activity and evoked M-response. RESULTS: No significant differences existed between groups in the relative decrease during the fatigue protocol. The recovery of force was slower in the late-polio subjects. A reduction in the root mean square (RMS) value during recovery was seen in the polio group, although a normalization of the mean power frequency (MPF) was seen in both groups. CONCLUSION: The weakness during the fatigue procedure was not caused by neuromuscular blockade, because electrical nerve stimulation evoked a normal response. The weakness after exercise was the result of a slow recovery that may reflect both central and peripheral fatigue. PMID- 10857523 TI - Exercise therapy effect on natural killer cell cytotoxic activity in stomach cancer patients after curative surgery. AB - OBJECTIVE: To evaluate the effect of early exercise therapy on the natural killer cell cytotoxic activity (NKCA) of patients who had undergone curative resection of stomach cancer. DESIGN: Prospective study. PATIENTS: Thirty-five stomach cancer patients who had undergone curative surgery were randomly divided into an exercise group (n = 17) and a control group (n = 18). INTERVENTION: From postoperative day 2, moderated exercise using arm and bicycle ergometers performed twice a day, 5 times a week, for 14 days. The intensity of exercise was 60% of maximal heart rate. Venous blood samples were obtained on postoperative days 1, 7, and 14. MAIN OUTCOME MEASURE: Mean sequential change of NKCA. RESULTS: The mean sequential change of NKCA decreased until postoperative day 7 and then increased. Mean NKCA of day 7 decreased in both groups, compared with that at postoperative day 1. At day 14, the mean NKCA of the exercise group demonstrated a significant increase compared with that of the control group (p < .05). CONCLUSION: This study suggests that early moderate exercise has a beneficial effect on the function of in vitro NK cells in stomach cancer patients after curative surgery. PMID- 10857524 TI - Postural sway characteristics in women with lower extremity arthritis before and after an aquatic exercise intervention. AB - OBJECTIVE: To examine the reliability of postural sway assessment in women with lower extremity arthritis and to ascertain the effects of an aquatic exercise intervention program on these measures. DESIGN: The reliability of postural sway measures was analyzed by within-subjects (Subject times Trial) analysis of variance (ANOVA). The effects of aquatic exercise were analyzed by repeated measures ANOVA using a planned comparison approach with an independent 2 x 2 (Group times Test) design. SETTING: Testing in a motor control research laboratory; aquatic exercise in a warm water pool at an area YMCA. PARTICIPANTS: Volunteer sample, 24 women with lower extremity arthritis (rheumatoid [RA] n = 11, osteo [OA] n = 13) randomly assigned into an aquatic exercise group (n = 14) or control group (n = 10). INTERVENTION: Postural sway measures under a two legged stance test on two separate test days: day 1, pretest; day 2, posttest, administered after a 6-week aquatic exercise program. RESULTS: Reliability correlation coefficients for postural sway measures ranged from .64 to .94 for both subject groups. Aquatic exercise subjects significantly reduced lateral sway and total sway area scores (by 18% to 30%) under both visual conditions after the 6-week intervention. Postural sway scores were significantly higher under the no vision condition than under the vision condition in each group for both test sessions. Both OA and RA groups had normal sagittal/lateral ratio scores. CONCLUSION: Women with lower extremity arthritis can be reliably assessed on postural sway measures on a stable two-legged stance test. Although they had normal sagittal/lateral sway ratio scores (ie, scores typical for nonarthritic peers), vision played an important role in their postural stability for this balance task. Aquatic exercise reduced postural sway in women with lower extremity arthritis, as demonstrated by a two-legged stance test, and this exercise program appears to be a viable treatment for increasing postural stability in this population. PMID- 10857525 TI - Patellar tendon repair: postoperative treatment. AB - OBJECTIVE: To compare results of patellar tendon repair after early and delayed postoperative mobilization. DESIGN: Two separate treatment groups, comparing 2 treatment alternatives at different time periods (before-after trial). PARTICIPANTS: Postoperative rehabilitation of 10 men who underwent patellar tendon repair. INTERVENTION: Delayed mobilization group: weight-bearing in a cast and isometric lower-extremity exercises for 6 weeks; active flexion and extension exercises thereafter. Early mobilization group: weight-bearing in an extension brace, isometric lower-extremity exercises, prone active knee flexion, and passive knee extension for 6 weeks; active flexion and extension exercises thereafter. Sixteen-month minimum follow-up. RESULTS: Clinical (physical) findings were: 2 excellent, 1 good, 2 fair in the delayed-mobilization group; 1 excellent, 3 good, 1 fair in early-mobilization group. Functional (pain and activity level) findings were: 2 good, 2 fair, 1 poor in the delayed-mobilization group; 3 good, 1 fair, 1 poor in the early-mobilization group. CONCLUSIONS: Clinical and functional results were similar for both treatment groups. Further study is required to determine any significant long-term differences between rehabilitation methods. PMID- 10857526 TI - The relation between lower extremity strength and shoulder overuse symptoms: a model based on polio survivors. AB - OBJECTIVE: To determine the relation between lower extremity weakness and shoulder overuse symptoms among polio survivors. We predicted that individuals with moderate weakness in their leg extensor muscles would use their arms to help compensate for this weakness and would be at high risk for developing symptoms of shoulder overuse. DESIGN: A cohort study of polio survivors recruited from the Einstein-Moss Postpolio Management Program (Philadelphia), the community, and the surrounding region. SETTING: A research laboratory at Moss Rehabilitation Research Institute, Philadelphia, PA. PARTICIPANTS: One hundred ninety-four polio survivors. Demographic and medical history data, symptom data, and strength data were obtained for each. MAIN OUTCOME MEASURES: Presence or absence of shoulder symptoms and ratings of pain by visual analogue scale were recorded. Strength was measured using a hand-held dynamometer and manual muscle testing. RESULTS: Shoulder symptoms could be grouped into two distinct clusters based on the type of testing used for assessment. Symptoms elicited by palpation were present in 26% of the subjects and were strongly related to knee extensor strength and weight. These symptoms were more common among women than men (42% and 10%, respectively). Symptoms elicited by resistance tests were present in 33% of the subjects and were seen with equal frequency in both sexes. These symptoms were also related to lower extremity strength, but the specific relationship was not as clear as for the palpation-related symptoms. CONCLUSIONS: Lower extremity weakness predisposes individuals to shoulder overuse symptoms. Sex and body weight are contributing factors. These results may be generalized to other populations with lower extremity weakness, including the elderly. PMID- 10857527 TI - Assessment of neuropsychologic impairments after head injury: interrater reliability and factorial and criterion validity of the Neurobehavioral Rating Scale-Revised. AB - OBJECTIVE: To study interrater reliability and factorial and criterion validity of the Neurobehavioral Rating Scale-Revised (NRS-R). DESIGN: Validity study on persons with traumatic brain injury (TBI) and test-retest reliability study on a randomly selected subset of patients. Factor analyses, kappa statistics, intraclass correlation coefficients, and Cronbach's alphas were used. SETTING: Inpatients from 15 French hospitals, mainly rehabilitation units. Other recruitment sites included a neurology hospital unit and a psychiatry hospital specifically devoted to TBI rehabilitation. PATIENTS: Two hundred eighty-six TBI patients ages 16 to 70 years (convenience sample). RESULTS: For the reliability study, the average of percentages of agreement among the items was 74.3% and the average of kappa statistics was .40. Factor analyses disclosed a maximum likelihood extraction of 5 correlated factors (F), explaining 42.2% of total variance: (F1) deficits in intentional behavior and in memory, (F2) lowering of emotional state, (F3) emotional and behavioral hyperactivation, (F4) lowering of arousal state and of attention, and (F5) language and speech problems. Results support the criterion validity of the factors. Reliability of the factor scores and internal consistencies of factors were very good. CONCLUSIONS: Results describe some important properties of the NRS-R and, through an understanding of its underlying structure and relationships with the patients' clinical characteristics, contribute to the conceptual framework of neuropsychologic impairments after TBI. PMID- 10857528 TI - Employment and social issues in adults with cerebral palsy. AB - OBJECTIVE: To assess the social and employment status of adults with cerebral palsy. DESIGN: Detailed medical history, physical examination, and functional rating in the PULTIBEC system were performed on all study participants; they also responded to a standardized social adaptation questionnaire. SETTING: Outpatient clinic. SUBJECTS: Volunteer participants (n = 101), all with cerebral palsy, between the ages of 27 and 74 years, living independently in the community. RESULTS: More than 80% wished that their physician knew more about cerebral palsy. The majority (84%) felt their parents overprotected them in childhood. More than 90% desired more sexual education. More than half (67%) lived independently, 34% with and 33% without attendant. Of the 53% who were competitively employed, 22% earned an income high enough that advancement would cause financial loss through termination of disability benefits. Speech deficits severely compromised functional verbal communication in 50%. Type of employment correlated more with adequate cognition than with physical or communicative impairments. CONCLUSIONS: Compared with earlier studies, the present study showed more adults with cerebral palsy achieving competitive employment and independent living, despite moderate to severe physical disability. Advances in rehabilitation technology, better home support services, and legal mandates in education and environmental access may have facilitated positive change for persons with cerebral palsy. Further studies are encouraged with emphasis on longitudinal designs. PMID- 10857529 TI - Important qualities in physiatrists: perceptions of rehabilitation team members and patients. AB - OBJECTIVE: To determine the personal qualities important in physiatrists as described by patients and rehabilitation team members. DESIGN: An oral survey. SETTING: Adult rehabilitation service at a teaching rehabilitation institution affiliated with an academic medical center. PARTICIPANTS: Convenience sample of 171 people including inpatients, rehabilitation nurses, occupational and physical therapists, and resident and full-time attending physicians. INTERVENTIONS: Two questions to elicit the personal qualities that were considered important in a physical medicine and rehabilitation physician. MAIN OUTCOME MEASURES: Subjects' responses grouped as personal qualities (personality), working qualities (professionalism), competence, caring, and collegiality. RESULTS: Caring and competence qualities were the most important to patients. Collegiality and caring were valued most by the rehabilitation staff. Personal qualities and competence were of relatively greater importance to the physiatrists. CONCLUSIONS: There was significant variability in the relative. value assigned to each of the five groups of traits among the respondents. Knowledge of the desires and expectations of the people involved in the rehabilitation process may benefit rehabilitation patients, physicians, and staff. PMID- 10857530 TI - Changes in spiritual beliefs after traumatic disability. AB - OBJECTIVES: To discover the effect of sudden-onset disability on spirituality, specifically, to investigate changes following the onset of disability in spiritual concepts and to outline a theoretical framework consisting of relationships with the self, others, the world, and a supreme power. STUDY DESIGN: The study used a cross-sectional, qualitative approach to understand changes in spirituality from the perspective of the disabled person. Intensive semistructured interviews were conducted with 16 participants, each of whom had either a spinal cord injury or brain injury, within the 2-year period after discharge from rehabilitation. Changes in spiritual concepts were explored in relation to 3 types of relationships (intrapersonal, interpersonal, and transpersonal) and 5 themes (awareness, closeness, trust, purpose, and vulnerability). RESULTS: Specific changes in spirituality described by sample members were: greater awareness of the self; a change in their view of their own independence; a sense of purpose in life that was not present before the onset of the disability; greater awareness of their own mortality and vulnerability; a new understanding of trust, especially when depending on others; loss of some significant relationships; greater appreciation and closeness with others and the world; and greater understanding of other disadvantaged groups. CONCLUSIONS: The interviews portrayed a significant ability to conceptualize issues in a spiritual context in the 2-year period after discharge from rehabilitation. Further, the changes reported suggest a positive effect of spirituality in the adjustment period following onset. PMID- 10857531 TI - Total knee replacement in an amputee patient: a case report. AB - Osteoarthritis is the most prevalent and more disabling of the rheumatic diseases. One of the most effective forms of treatment of severe osteoarthritis is total joint arthroplasty. Although studies suggest that the incidence of osteoarthritis is higher in prosthetic users, research supporting total joint arthroplasty as an option for treating amputee patients with advanced osteoarthritis is lacking. We report the case of a 76-year-old man with right transtibial amputation who had an excellent outcome after undergoing bilateral total knee replacements for advanced osteoarthritis. PMID- 10857532 TI - Shoulder pain as an unusual presentation of pneumonia in a stroke patient: a case report. AB - Etiologies of shoulder pain in the hemiplegic population, such as glenohumeral subluxation, frozen shoulder, and reflex sympathetic dystrophy (RSD), have been described extensively. We present an 89-year-old woman with right hemiparesis secondary to ischemic lacunar infarction who developed sudden onset of right shoulder pain on the fifth day of inpatient rehabilitation. The pain was severe, limiting range of motion (ROM) and participation in therapy. Extensive investigations to rule out subluxation, fracture, connective tissue disease, RSD, and pulmonary embolism were negative. Ultimately, her shoulder pain and decreased ROM completely resolved with antibiotic treatment for right lower lobe pneumonia. We conclude that her symptoms were possibly referred pain from diaphragmatic irritation transmitted via right C4 sensory axons in the phrenic nerve, which shares the same dermatome as the right acromion area. This case was an unusual presentation of pneumonia in an elderly woman with hemiplegia. We recommend that pneumonia be considered in the differential diagnoses of shoulder pain. PMID- 10857533 TI - Adrenal insufficiency masquerading as sepsis in a patient with tetraparesis: a case report. AB - Several endocrine changes have been reported in patients with tetraplegia after spinal cord injury (SCI). These changes should be considered when prescribing medications that influence the endocrine pathways. Megestrol acetate has gained acceptance as a way to promote weight gain in cachectic patients without significant adverse effects. We present a case of a 51-year-old man with C5-C6 tetraparesis who was only 67% of his ideal body weight and was placed on megestrol acetate 5 months before admission for a urologic procedure. Postoperatively, the patient had severe hypotension and tachycardia that was interpreted as a septic or cardiac event. Further workup revealed subnormal levels of 8AM cortisol. An adrenocorticotrophic hormone stimulation test demonstrated results consistent with adrenal suppression. Hydrocortisone supplementation was started, and 6 months later cortisol levels were within normal limits. Cachexia, hypotension, and mild tachycardia are not uncommon in patients with SCI. When severe hypotension and tachycardia are seen in patients with tetraplegia, the diagnosis of adrenal insufficiency should be considered. PMID- 10857534 TI - Paraneoplastic cerebellar degeneration as the first evidence of cancer: a case report. AB - Paraneoplastic cerebellar degeneration (PCD) is the most frequently seen paraneoplastic syndrome affecting the brain. PCD is most commonly associated with cancers of the ovary, breast, and lung. The anti-Purkinje cell antibodies (anti Yo) that specifically damage the Purkinje cells of the cerebellum are found in the patient's serum and cerebrospinal fluid. The typical presentation of PCD includes limb and truncal ataxia, often along with dysarthria. This report describes the case of a 47-year-old woman without significant medical history who developed new onset of unsteady gait, headache, and vertigo. The imaging studies suggested rhombencephalitis. The patient initially responded to corticosteroid treatment. Unfortunately, her gait ataxia worsened and she developed dysarthria, neither of which responded to increasing dosages of corticosteroids. Extensive imaging studies showed no evidence of tumor, but the patient was found to have positive anti-Yo antibodies and elevated cancer antigen 125 (CA-125). Pathology results from exploratory laparotomy revealed stage III C adenocarcinoma of the ovary. This case demonstrates that PCD may be the presenting symptom of an occult malignancy. The pathogenesis, diagnosis, and treatment of PCD, and its rehabilitation implications, are reviewed. PMID- 10857535 TI - A ganglion cyst causing lumbar radiculopathy in a baseball pitcher: a case report. AB - This report describes a case of a professional baseball pitcher who developed acute left lumbar radicular symptoms after a baseball game and was subsequently sidelined for the rest of the season. Physical examination revealed depressed reflexes in the left posterior tibialis and left medial hamstring muscles, mild weakness in the left extensor hallucis longus, and positive dural tension signs. Magnetic resonance imaging demonstrated an ovoid mass at the L4-L5 level, causing compression of the dura. Surgical resection of the mass resulted in resolution of his symptoms. Pathology revealed that the mass was a ganglion cyst. A ganglion cyst is a rare cause of lumbar radiculopathy and should be considered in the differential diagnosis if a patient with lumbar radiculopathy fails to respond to conservative treatment. PMID- 10857536 TI - Gait evaluation of a transfemoral prosthetic simulator. AB - OBJECTIVE: To test a prosthetic simulator developed to allow persons without amputation to walk like a person with a transfemoral (TF) amputation. PATIENTS: Five able-bodied subjects; comparison with data from the literature on persons with TF amputations. SETTING: Motion analysis laboratory. DESIGN: Two 45- to 60 minute gait training sessions before subjects walked along a 10-meter walkway. There were 6 trials: 3 walking with a cane, 3 without a cane. MAIN OBJECTIVE MEASURES: Sagittal plane kinematic and kinetic analysis of ankle, knee, and hip: angular velocity, joint moment, and power. RESULTS: Kinematic and kinetic analyses showed that joint mechanics during walking were similar between the test subjects and comparative results from persons with TF amputations (reported in the literature). Test subjects walked slower and moved their hip and knee joints faster (higher angular velocity values during the terminal swing) than the TF amputee subjects, although these results were not statistically significant (p < .05). These findings were consistent with new prosthetic users who are more tentative during gait training. However, a perfect simulation would show no difference in kinematic results. CONCLUSION: These results support the use of a TF prosthetic simulator to help health care professionals experience the process of fitting the prosthesis from the client's perspective. PMID- 10857537 TI - Growth of bifidobacteria in soymilk and their survival in the fermented soymilk drink during storage. AB - Growth of Bifidobacterium infantis CCRC 14633 and B. longum B6, in soymilk was investigated in the present study. It was found that soymilk could support the growth of both organisms tested. B. infantis grew better than B. longum in soymilk. Supplementation of bifitose (isomaltooligosccharie), glucose, lactose or galactose to soymilk increased the growth of B. infantis and B. longum as determined after 48 h of fermentation. On the other hand, addition of yeast extract, peptone, tryptone, casitone or N-Z-Case plus to soymilk enabled B. infantis to reach its maximum population in a shorter cultivation time of 24 h. Acid production by B. longum and B. infantis in soymilk was mainly non-growth associated, while in the yeast extract-supplemented soymilk, acid produced by B. infantis was found to be growth-associated. Populations of B. longum reduced more than did B. infantis in the prepared fermented soymilk drink during storage period. Viable population of both test organisms reduced less in the fermented drink held at 5 degrees C than at 25 degrees C. After a 10-day storage at 5 degrees C, viable B. infantis and B. longum reduced by 0.44 and 3.18 log CFU/ml, respectively, in the fermented drink. Addition of sucrose to the fermented drink resulted in an increased reduction of viable bifidobacteria during the storage period. This phenomenon was most prominent with B. infantis in the fermented drink held at 25 degrees C. PMID- 10857538 TI - Survival characteristics and the applicability of predictive mathematical modelling to Listeria monocytogenes growth in sous vide products. AB - Survival and growth of Listeria monocytogenes isolates during sous vide processing and storage, and the applicability of predictive modelling in determining the potential for growth of L. monocytogenes in broth models and in sous vide products was investigated. L. monocytogenes grew in anaerobic tryptose phosphate broth and in chicken and beef samples by 2 log cycles in 8 days at 3 degrees C and 4-5 log cycles in 6 days at 8 degrees C. However, heating to an internal temperature of 70 degrees C resulted in a 4-5 log reduction and 70 degrees C/2 min resulted in a reduction greater than 7 log cycles. Lowering the product pH to 5.0 was effective in inhibiting L. monocytogenes growth, whereas a sodium chloride concentration of 2% had a negligible effect on growth rates. The square root model (Ratkowsky et al., 1983) predicted L. monocytogenes growth rates at 0-25 degrees C with a coefficient of determination (R2 value) of 98.36 99.63% and a bias factor of 1.08 to 1.21 in beef, chicken and broth substrates of unmodified pH. In addition, the Response Surface Polynomial Model (Version 3.1, Buchanan et al., 1989) predicted generation times at 5-25 degrees C with a 0 17.4% difference between observed and expected generation times in tryptose phosphate broth at pH 7.3. There were however, large differences (25.5 vs. 5.3 h) between observed generation times at pH 5.6 (8 degrees C) and those predicted by the Pathogen Modelling Program in tryptose phosphate broth. A divergence from predicted values was also noted at lower temperatures (0-3.5 degrees C) in the square root model. PMID- 10857539 TI - The spoilage microflora of cured, cooked turkey breasts prepared commercially with or without smoking. AB - Lactobacillus sakei subsp. carnosus was predominant in the spoilage flora of sliced, vacuum-packed, smoked, oven-cooked turkey breast fillets which developed mild, sour spoilage flavors after 4 weeks storage at 4 degrees C. In contrast, Leuconostoc mesenteroides subsp. mesenteroides predominated in the spoilage flora of sliced, vacuum-packed, unsmoked, boiled turkey breast fillets from the same plant which were also stored at 4 degrees C. The spoilage flora of the unsmoked breasts grew faster than that of the smoked breasts and was more diverse. Lactobacillus sakei, Weissella viridescens and an atypical group of leuconostoc like bacteria were also members of the unsmoked turkey breasts flora. Consequently, the unsmoked breasts spoiled after 2 weeks at 4 degrees C: the packs swelled and the meat developed strong sour odors and flavors and abundant slime. Except for the unidentified leuconostocs, which apparently survived boiling of the unsmoked turkey, all the spoilage organisms contaminated the meats during the slicing and vacuum packaging operations. From their biochemical reactions and cellular fatty acid profiles, the atypical leuconostocs were more closely related to Leuconostoc carnosum than W. viridescens. Carnobacteria and Brochothrix thermosphacta were present in relatively large numbers on the raw turkey, but were not numerous in the spoilage flora of the cooked, vacuum-packed meat products. PMID- 10857540 TI - Effects of weak acid preservatives on the growth and thermal death of the yeast Pichia membranifaciens in a commercial apple juice. AB - Pichia membranifaciens exhibited a dissociative temperature profile (the temperature range of thermal death was distinct from the temperature range of growth) when incubation took place either in a commercial apple juice (AJ) or in a synthetic mineral medium with glucose and vitamins (MGV). In AJ the maximum temperature for growth (Tmax) was 38.6 degrees C, which decreased to 36 degrees C in the presence of either 1 mM sorbic or 1 mM benzoic acid. The minimum temperatures of thermal death (Tmind) were, respectively, 40 and 38 degrees C with either of the acids. The yeast could grow with up to 2 mM sorbic or 3 mM benzoic acid, at 25 degrees C, which is close to the optimum temperature for growth (Top). At temperatures slightly above Tmind, sorbic acid was an actual enhancer of death rather than benzoic, the latter conferring some protection. However, these effects were reversed at higher temperatures (above 43 degrees C), at which benzoic acid was the most operative, in contrast to sorbic which was highly protective of the yeast against thermal death. The addition of acetaldehyde to sulphur-dioxide-containing juice reduced the lag phase and increased the overall specific growth rates. Sporulated or stationary vegetative cultures were more heat-resistant than exponential cultures, particularly at temperatures above 45 degrees C. PMID- 10857541 TI - Development of a dose-response relationship for Escherichia coli O157:H7. AB - E. coli O157:H7 is an emerging food and waterborne pathogen. The development of acceptable guidelines for exposure to this organism based on quantitative microbial risk assessment requires a dose response curve. In this study, a prior animal study was used to develop a dose response relationship. The data was adequately fit by the beta-Poisson dose response relationship. This relationship was validated with reference to two outbreaks of this organism. It was found that the low dose extrapolation of the animal data using the beta-Poisson relationship provided estimates of risk concordant with those noted in the outbreaks. The fitted dose response relationship in conjunction with population estimates of the prevalence of E. coli O157:H7 illness indicates that the overall exposure is quite low in the US. PMID- 10857542 TI - Survival of osmotic and acid stress by Listeria monocytogenes strains of clinical or meat origin. AB - The ability of 30 Listeria monocytogenes strains, 15 of meat origin and 15 of clinical origin, to use carnitine as an osmoprotectant and to resist acid stress was determined. All strains examined were able to use carnitine as an osmoprotectant, indicating the importance of this characteristic to the survival of L. monocytogenes in natural environments. Clinical and meat strains, however, differed with respect to this characteristic. Specifically, 73% of meat strains reached a lower maximum cell density in the presence of carnitine with osmotic stress than in its absence with no stress. Only 33% of clinical strains displayed the same feature whereas the remaining clinical strains reached a higher maximum cell density in the presence of carnitine with osmotic stress than in its absence with no stress. The physiological reasons and advantage of this difference are unclear. When exposed to conditions of severe acid stress (pH 2.5) for 2 h, only two L. monocytogenes strains (L66 and L78), both of meat origin, displayed significant reductions (P < 0.05) in number (3.51 and 2.79 log cfu, respectively). Acid-sensitive strains were not found among the clinical isolates examined, highlighting the importance of acid stress resistance in the infection process. PMID- 10857543 TI - Combined PCR and slot blot assay for detection of Salmonella and Listeria monocytogenes. AB - Detection of Salmonella typhimurium and Listeria monocytogenes by the polymerase chain reaction (PCR) assay coupled with slot blot detection was investigated in this study. After being extracted from diluted bacterial culture with the extraction buffer, bacterial DNA was subjected to PCR. The slot blot assay was optimized and used to detect PCR products. The lowest detection level of this method was 10(3) cfu/ml in the original culture media for both pathogens, or 5 bacterial cells in the PCR reaction. Combined with immunomagnetic separation (IMS) to separate and concentrate bacteria from samples, the detection limit could be 40 cfu/ml of bacteria from milk samples. The whole detection procedure was completed within 7 h. After multiplex PCR (amplification of DNA from two different bacteria in the same PCR tube) and slot blot, a detection level of 10(3) cfu/ml was achieved in the simultaneous detection for both pathogens, which was similar to that of individual detection for each pathogen. The combination of PCR and slot-blot seems to be highly sensitive and time-efficient, and is therefore promising for routine use in the detection of Salmonella and L. monocytogenes in food samples such as milk. PMID- 10857544 TI - The use of a starter culture in the fermentation of cassava for the production of "kivunde", a traditional Tanzanian food product. AB - Three cassava fermentation methods (spontaneous fermentation, back-slopping and the use of starter culture) for the production of kivunde, executed in three trials at 30 degrees C, were compared in terms of cyanide level reduction, microbiology and product quality improvement. Among the isolates from spontaneously fermented cassava batches, four strains were selected on the basis of their enzymatic activities and acid production. All were identified as Lactobacillus plantarum and were used as starters in this study. Lowest residual cyanide levels were detected after 120 h fermentation time in samples fermented with the starter culture and were below the maximum value of 10 mg/kg recommended by the Codex/FAO for cassava flour. This finding seems to be related to the alpha glucosidase activity of the inoculated strains of which API-zyme (Bio-Merieux) tests showed activities of between 20 and > or = 40 nmol/4 h. The total residual cyanide levels of the spontaneous and back-slopping fermentations at 96 h were respectively 43.5 and 47.7 mg/kg dry weight of cassava. Extension of the fermentation period to 5 days, lead to further substantial reduction in the residual cyanide level in both these processes, but not below the recommended maximum value as in the case of starter culture fermented products. The spontaneous and back-slopping fermented cassava showed signs of deterioration after 3 days of fermentation. There was a sharp drop of pH and an increase of titratable acidity for all three batches during the first 48 h followed by a slow rise of pH and drop in titratable acidity towards the end of fermentation. The samples fermented with the starter culture had a smooth texture and pleasant fruity aroma, as opposed to the course and dull appearance and more complex flavour of the samples of spontaneous and back-slopping batches. During fermentation with starter culture, Enterobacteriaceae and yeasts and moulds could not be isolated throughout the period of fermentation (detection limit: 10 colony forming units/g). The present findings indicate the suitability of these Lb. plantarum strains as starter cultures for cassava fermentation in the kivunde process. The paper highlights the potential for the improvement of a traditional African fermented food (kivunde) through the use of a starter culture. PMID- 10857545 TI - Effects of salts on Debaryomyces hansenii and Saccharomyces cerevisiae under stress conditions. AB - The effect of Na+ and K+ on growth and thermal death of Debaryomyces hansenii and Saccharomyces cerevisiae were compared under stress conditions as those commonly found in food environments. At the supraoptimal temperature of 34 degrees C both cations at concentrations of 0.5 M stimulated growth of D. hansenii, while K+ had no effect and Na+ inhibited growth of S. cerevisiae. At 8 degrees C, close to the minimum temperature for growth in both species, both cations inhibited both yeasts, this effect being more pronounced with Na+ in S. cerevisiae. At extreme pH values (7.8 and 3.5) both cations at concentrations of 0.25 M stimulated D. hansenii while Na+ inhibited S. cerevisiae. K+ inhibited this yeast at pH 3.5. Thermal inactivation rates, measured at 38 degrees C in D. hansenii and at 48 degrees C in S. cerevisiae, decreased in the presence of both cations. This protective effect could be observed in a wider range of concentrations in D. hansenii. These results call the attention to the fact that not all yeasts have the same behaviour on what concerns synergy or antagonism of salt together with other stress factors and should be taken into consideration in the establishment of food preservation procedures. PMID- 10857546 TI - Kinetic analysis and modelling of combined high-pressure-temperature inactivation of the yeast Zygosaccharomyces bailii. AB - Eight foodborne yeasts were screened for sensitivity to high-pressure (HP) inactivation under a limited number of pressure-temperature combinations. The most resistant strains were Zygoascus hellenicus and Zygosaccharomyces bailii. The latter was taken for a detailed study of inactivation kinetics over a wide range of pressures (120-320 MPa) and temperatures (-5 to 45 degrees C). Isobaric and isothermal inactivation experiments were conducted in Tris-HCl buffer pH 6.5 for 48 different combinations of pressure and temperature. Inactivation was biphasic, with a first phase encompassing four to six decades and being described by first-order kinetics, followed by a tailing phase. Decimal reduction times (D) were calculated for the first-order inactivation phase and their temperature and pressure dependence was described. At constant temperature, D decreased with increasing pressure as expected. At constant pressure, D showed a maximum at around 20 degrees C, and decreased both at lower and at higher temperatures. A mathematical expression was developed to describe accurately the inactivation of Z. bailii as a function of pressure and temperature under the experimental conditions employed. A limited number of experiments in buffer at low pH (3-6) suggest that the model is, in principle, applicable at low pH. In apple and orange juice however, higher inactivation than predicted by the model was achieved. PMID- 10857547 TI - Differentiation of Tetragenococcus populations occurring in products and manufacturing processes of puffer fish ovaries fermented with rice-bran. AB - Tetragenococcus strains isolated from the manufacturing process of Japanese puffer fish ovaries fermented with rice-bran were characterized and differentiated phenotypically and genotypically. A total of 413 Tetragenococcus isolates were evaluated. On the basis of five representative substrates, the isolates were grouped into seven groups. An RFLP (restriction fragment length polymorphisms) analysis of the 16S rRNA gene of representative strains of major groups revealed that they could be grouped into two groups: one was identified as the most prominent halophilic lactic acid coccus, Tetragenococcus halophilus, and the other as T. muriaticus, which has recently been added to the genus Tetragenococcus as a new species. Physiologically, the major differences between the two groups were found in the ability to grow in medium not supplemented with NaCl and the fermentation of L-arabinose, sucrose and D-mannitol, and several other carbohydrates. PMID- 10857548 TI - High levels of background flora inhibits growth of Escherichia coli O157:H7 in ground beef. AB - The influence of natural background flora under aerobic and anaerobic incubation on the growth of Escherichia coli O157:H7 in ground beef was investigated. The background flora from eight different commercial ground beef were added to ground beef spiked with E. coli O157:H7 and stored either aerobically or anaerobically at 12 degrees C. The results showed that the presence of a large number of background bacteria in the ground meat inhibited the growth of E. coli O157:H7 both aerobically and anaerobically. Inhibition was more pronounced under anaerobic conditions. The background floras consisted mainly of lactic acid bacteria of which approximately 80% were Lactobacillus sakei. These results show the importance of the natural background flora in meat for inhibition of growth of E. coli O157:H7. PMID- 10857550 TI - A new rapid automated method for the detection of Listeria from environmental swabs and sponges. AB - Many food and meat processors test environmental swabs and sponges to confirm the absence of Listeria spp. Spectral pattern changes in a liquid growth medium, resulting from esculin hydrolysis by Listeria in contaminated swabs and sponges, were automatically monitored by the BioSys instrument in a semifluid layer (SFL). The blackening of SFL in modified MOX broth resulted in sharply declining curves, which were easily detected by the instrument. The instrument detected all nine strains of Listeria monocytogenes tested. None of the gram negative organisms (Proteus, Escherichia coli, Pseudomonas, Citrobacter and Yersinia) were detected by the system, nor were most gram positive organisms, including Bacillus, Streptococcus, and Lactobacillus strains, Staphylococcus aureus, Enterococcus faecium and E. faecalis, which hydrolyze esculin, produced black colonies on PALCAM and Oxford media and were also detected in the system. A total of 122 sponges and swabs collected at food processing plants were evaluated by this method. Of these, 99 were negative, and 11 were positive. L. innocua was the dominant Listeria species in these environmental samples. Good correlation was obtained between numbers of Listeria and detection times of esculin hydrolysis: 1000 CFU/swab were detected in 10-13 h, whereas 1-10 CFU/swab were detected in less than 22 h. The total assay time was 26 h. PMID- 10857549 TI - UV-induced Lactobacillus gasseri mutants resisting sodium chloride and sodium nitrite for meat fermentation. AB - Lactobacillus gasseri, one of the predominant lactobacilli in human intestinal tracts, is utilized for probiotics and dairy starter cultures. However, since L. gasseri is relatively sensitive to sodium chloride and sodium nitrite (essential compounds for meat products), it is difficult to utilize this species for conventional fermented meat products. In this study, efforts were directed to generate mutants of L. gasseri resisting sodium chloride and sodium nitrite. UV irradiation of the strain of L. gasseri JCM1131(T) generated several mutants resisting these compounds. A mutant strain 1131-M8 demonstrated satisfactory growth in meat containing 3.3% sodium chloride and 200 ppm sodium nitrite. Although proteins extracted from the cell surface of 1131-M8 were slightly different from those of the original strain, other biochemical characteristics of both strains were indistinguishable. These results suggest that the L. gasseri mutant obtained in this study could be utilized as a starter culture to develop probiotic meat products. PMID- 10857552 TI - Bibliography of food microbiology. PMID- 10857551 TI - Bacterial populations associated with bulk packaged beef supplemented with dietary vitamin E. AB - Five Simmentaler type calves were fed diets supplemented with 500 mg vitamin E per day and five fed control diets, Rump steaks from each carcass were PVC overwrapped and bulk packaged in 100% CO2 or 20% CO2:80% O2. Bulk packs were stored up to 42 days at 4 degrees C and PVC-overwrapped samples subsequently displayed up to 7 days at 4 degrees C. After display the Aerobic Plate Count (APC) of steaks was determined and four colonies were randomly selected from the highest dilution APC plates showing growth. A total of 627 colonies were obtained. Gram-reaction, catalase, oxidase, morphology and motility of the isolates were determined. The gram-negative and gram-positive isolates were then identified using a dichotomous identification key. Enterobacteriaceae, Pseudomonas spp. and Acinetobacter spp. predominated on rump steaks from both feeding treatments and in packaging treatments. After 42 days bulk storage Enterobacteriaceae, Pseudomonas spp., lactic acid bacteria and Acinetobacter spp. predominated in 20% CO2:80% O2 and 100% CO2 bulk packaging. Enterobacteriaceae, Pseudomonas spp. and Acinetobacter spp. predominated on rump steaks, from both feeding and packaging treatments, during the aerobic display period of 7 days. PMID- 10857553 TI - The effects of insulin-like growth factors on tumorigenesis and neoplastic growth. AB - Several decades of basic and clinical research have demonstrated that there is an association between the insulin-like growth factors (IGFs) and neoplasia. We begin with a brief discussion of the function and regulation of expression of the IGFs, their receptors and the IGF-binding proteins (IGFBPs). A number of investigational interventional strategies targeting the GH or IGFs are then reviewed. Finally, we have assembled the available scientific knowledge about this relationship for each of the major tumor types. The tumors have been grouped together by organ system and for each of the major tumors, various key elements of the relationship between IGFs and tumor growth are discussed. Specifically these include the presence or absence of autocrine IGF-I and IGF-II production; presence or absence of IGF-I and IGF-II receptor expression; the expression and functions of the IGFBPs; in vitro and in vivo experiments involving therapeutic interventions; and available results from clinical trials evaluating the effect of GH/IGF axis down-regulation in various malignancies. PMID- 10857554 TI - Congenital adrenal hyperplasia due to 21-hydroxylase deficiency. AB - More than 90% of cases of congenital adrenal hyperplasia (CAH, the inherited inability to synthesize cortisol) are caused by 21-hydroxylase deficiency. Females with severe, classic 21-hydroxylase deficiency are exposed to excess androgens prenatally and are born with virilized external genitalia. Most patients cannot synthesize sufficient aldosterone to maintain sodium balance and may develop potentially fatal "salt wasting" crises if not treated. The disease is caused by mutations in the CYP21 gene encoding the steroid 21-hydroxylase enzyme. More than 90% of these mutations result from intergenic recombinations between CYP21 and the closely linked CYP21P pseudogene. Approximately 20% are gene deletions due to unequal crossing over during meiosis, whereas the remainder are gene conversions--transfers to CYP21 of deleterious mutations normally present in CYP21P. The degree to which each mutation compromises enzymatic activity is strongly correlated with the clinical severity of the disease in patients carrying it. Prenatal diagnosis by direct mutation detection permits prenatal treatment of affected females to minimize genital virilization. Neonatal screening by hormonal methods identifies affected children before salt wasting crises develop, reducing mortality from this condition. Glucocorticoid and mineralocorticoid replacement are the mainstays of treatment, but more rational dosing and additional therapies are being developed. PMID- 10857555 TI - The roles of prolactin, growth hormone, insulin-like growth factor-I, and thyroid hormones in lymphocyte development and function: insights from genetic models of hormone and hormone receptor deficiency. AB - An extensive literature suggesting that PRL, GH, IGF-I, and thyroid hormones play an important role in immunity has evolved. Because the use of one or more of these hormones as immunostimulants in humans is being considered, it is of critical importance to resolve their precise role in immunity. This review addresses new experimental evidence from analysis of lymphocyte development and function in mice with genetic defects in expression of these hormones or their receptors that calls into question the presumed role played by some of these hormones and reveals unexpected effects of others. These recent findings from the mutant mouse models are integrated and placed in context of the wider literature on endocrine-immune system interactions. The hypothesis that will be developed is that, with the exception of a role for thyroid hormones in B cell development, PRL, GH, and IGF-I are not obligate immunoregulators. Instead, they apparently act as anabolic and stress-modulating hormones in most cells, including those of the immune system. PMID- 10857556 TI - Leukemia-inhibitory factor-neuroimmune modulator of endocrine function. AB - Leukemia-inhibitory factor (LIF) is a pleiotropic cytokine expressed by multiple tissue types. The LIF receptor shares a common gp130 receptor subunit with the IL 6 cytokine superfamily. LIF signaling is mediated mainly by JAK-STAT (janus kinase-signal transducer and activator of transcription) pathways and is abrogated by the SOCS (suppressor-of cytokine signaling) and PIAS (protein inhibitors of activated STAT) proteins. In addition to classic hematopoietic and neuronal actions, LIF plays a critical role in several endocrine functions including the utero-placental unit, the hypothalamo-pituitary-adrenal axis, bone cell metabolism, energy homeostasis, and hormonally responsive tumors. This paper reviews recent advances in our understanding of molecular mechanisms regulating LIF expression and action and also provides a systemic overview of LIF-mediated endocrine regulation. Local and systemic LIF serve to integrate multiple developmental and functional cell signals, culminating in maintaining appropriate hormonal and metabolic homeostasis. LIF thus functions as a critical molecular interface between the neuroimmune and endocrine systems. PMID- 10857557 TI - Spectrophotometric and HPLC determination of secnidazole in pharmaceutical tablets. AB - Simple and accurate spectrophotometric and HPLC methods were developed for the determination of secnidazole in tablets dosage form. The first spectrophotometric method depends on the reduction of secnidazole molecule with zinc dust and hydrochloric acid followed by condensation with either p dimethylaminobenzaldehyde or anisaldehyde to give colored chromogens having absorbance at 494 and 398 nm, respectively. The second method was based on the reaction of the drug with sodium nitroprusside in the presence or absence of hydroxylammonium hydrochloride. The formed colored chromogens were measured at 584 and 508 nm, respectively. The experimental conditions were optimized and Beer's law was obeyed over the applicable concentration ranges. The application of HPLC procedures depended on using either a conventional or microbore reverse phase (C18) column along with mobile phases consisting of water and methanol (30:70), at pH of 3.5. Both techniques were applied successfully for the analysis of secnidazole in tablets form. The results obtained from both procedures were statistically compared using the Student's-t and F-variance ratio tests. PMID- 10857558 TI - Uniform-sized molecularly imprinted polymer material for (S)-propranolol. AB - A uniform-sized molecularly imprinted polymer for (S)-propranolol has been prepared by a multi-step swelling and thermal polymerization method using methacrylic acid (MAA) and ethylene glycol dimethacrylate (EDMA) as a host functional monomer and cross-linker, respectively. The obtained (S)-propranolol imprinted MAA-EDMA materials were evaluated using aqueous-rich eluents by HPLC. The (S)-propranolol imprinted MAA-EDMA materials had specific recognition for (S) propranolol and moderate recognition for some structurally related beta adrenergic antagonists (especially, pindolol and alprenolol), but had no recognition for other basic compounds, and neutral and acidic compounds. Enantioseparations of propranolol and some structurally related beta-adrenergic antagonists were attained with the imprinted MAA-EDMA materials. Hydrophobic interaction of the naphthyloxy and isopropyl groups of propranolol, and ionic interaction of the isopropylamino group of propranolol with the (S)-propranolol imprinted MAA-EDMA materials could play an important role in enantioselective recognition of propranolol. PMID- 10857559 TI - Integrated pervaporation/detection for the determination of fluoride in pharmaceuticals. AB - A selective dynamic method for the determination of fluoride in pharmaceuticals based on the integration of pervaporation and potentiometric detection in a laboratory-made module is proposed. The analyte was continuously converted into volatile trimetylfluorosilane by reaction with hexamethyldisilazane, injected into a donor stream and accepted in a basic buffer solution after pervaporation. The method thus developed has a determination range between 1.5 and 200 mg l-1, precision (expressed as R.S.D.) of 3.4%, and has been applied to the determination of fluoride in different pharmaceutical products, with yields ranging between 90.6 and 100.3%. B.V. PMID- 10857560 TI - A comparison of matrix resolution method, ratio spectra derivative spectrophotometry and HPLC method for the determination of thiamine HCl and pyridoxine HCl in pharmaceutical preparation. AB - A comparison of two spectrophotometric methods and a HPLC method were described in this work for the analysis of pyridoxine hydrochloride and thiamine hydrochloride in a vitamin combination. In the first method, A(1)1 (1%, 1 cm) values of these two compounds were calculated using absorbances measured at 246.8 and 290.5 nm in zero-order spectra. The matrix was written for A(1)1 (1%, 1 cm) values and the concentration of both compounds were determined using 'Matlab' software. In the second method, the measurements in the derivative of the ratio spectra were made at 297.8 and 309.5 nm for pyridoxine hydrochloride and at 245.6 and 257.7 nm for thiamine hydrochloride. The calibration graphs were established in the range 8-40 microg/ml of both vitamins. In the HPLC method, the separation of these compounds was realized on a Nucleosil 100-5 C18 column with 0.1 M (NH4)2C03-water methanol (5:15:80 v/v) as the mobile phase. Results of spectrophotometric and HPLC procedures were compared. PMID- 10857561 TI - Voltammetric methods for analytical determination of fleroxacin in quinodis tablets. AB - The direct current (dc) and differential pulse (dp) polarographic reduction of fleroxacin was done in a wide pH range from 2.48 to 13.00. The appropriate buffer choice was made for its dp polarographic determination in a range from 1.845 to 16.926 microg /ml, at pH 8.50. The adsorptive properties of fleroxacin were investigated in order to achieve an increase in sensitivity and a possibility of fleroxacin determination by applying the adsorptive stripping voltammetric method. The adsorptive processes at the hanging mercury drop electrode were investigated in Britton-Robinson and borate buffers. Adsorptive preconcentration followed by differential-pulse cathodic stripping showed one wave at approximately - 1.1 V being the most sensitive for analytical determination of fleroxacin. Two linear ranges were obtained, the first one from 18.465 to 258.51 ng/ml, and the second one from 3.693 to 18.465 ng/ml. PMID- 10857562 TI - A fast and simple method for the determination of clavulanic acid in human plasma using derivatisation reaction kinetics. AB - A stopped-flow mixing technique is used to determine clavulanic acid in human plasma after plasma deproteinisation with acetonitrile and removal of the organic solvent by extraction with dichloromethane. The reaction kinetic profiles for the reaction between clavulanic acid and imidazole are determined by measuring the absorbance at 312 nm of the imidazole derivative. The reaction rates are proportional to the concentration of the clavulanic acid and by plotting reaction rates against clavulanic acid concentrations linear calibration curves could be constructed over the range 0.30-10.0 microg/ml. PMID- 10857563 TI - Quantification of gadodiamide as Gd in serum, peritoneal dialysate and faeces by inductively coupled plasma atomic emission spectroscopy and comparative analysis by high-performance liquid chromatography. AB - An inductively coupled plasma atomic emission spectroscopy (ICP-AES) method for determination of gadodiamide as Gd in serum, peritoneal dialysate and faeces was developed. The within-day and between-day precision for determination of Gd in serum and peritoneal dialysate were 0.60-2.9 and 1.8-4.4%, respectively, and the accuracy was 98.0-99.3%. The quantification limits in serum and peritoneal dialysate were 6.5 and 1.6 microM Gd, respectively. The within-day and between day precision determination of gadolinium in faeces were 1.0-5.3 and 2.2-7.9%, respectively, and the accuracy was 104-116%. The quantification limit was 11 nmol Gd/g dry weight. For the high-performance liquid chromatography (HPLC) method, the within-day precision in determination of gadodiamide in peritoneal dialysate was 1.2% and the accuracy was 103%. The quantification limit was 0.9 microM Gd. Comparative analysis of gadodiamide in serum and peritoneal dialysate from severely impaired renal patients by ICP-AES and HPLC revealed no metabolism of chelator or transmetallation of gadolinium, even in samples obtained as long as 7 days after dosing. Furthermore, the ICP-AES determination of Gd in faeces allows for the determination of faeces content of Gd corresponding to less than 0.1% of a clinical dosage of a Gd-based contrast medium. PMID- 10857564 TI - A validated LC method for the quantitative determination of celecoxib in pharmaceutical dosage forms and purity evaluation in bulk drugs. AB - A new reversed-phase, isocratic LC method was developed for the quantitative determination of COX-2 inhibitor celecoxib in bulk drugs and in pharmaceutical dosages. The proposed method is also applicable for the purity evaluation of celecoxib in bulk drugs. 5-Methyl 2-Nitro phenol has been used as internal standard for the quantitative determination of celecoxib. The method has been completely validated and proven to be rugged. The limit of detection (LOD) and limit of quantitation (LOQ) for celecoxib impurities namely, 4-hydrazino benzene sulfonamide (Intermediate I) and 1-(4-methyl phenyl)-4,4.4-trifluro butan-1,3 dione (Intermediate II) were found to be 32.0 and 97 ng. respectively. The active pharmaceutical ingredient was extracted from its finished dosage form (capsule) using methanol. The percentage recoveries ranged from 90.7 to 93.8. The stability studies were performed for celecoxib solution placed on laboratory bench and in refrigerator for hundred days. The samples were found to be stable for the study period. PMID- 10857565 TI - Characterization of bovine collagens using capillary electrophoresis. AB - A capillary electrophoresis method was developed and characterized for analyzing the spectrum of collagen subspecies in collagen preparations. The Bio-Rad CE-SDS protein kit was used for the dynamic sieving separation of collagen subspecies in this CE method (DSCE). The optimized method utilized a 36 cm (or 24 cm) x 50 microm uncoated capillary, electrophoretic injection at 10 kV for 10 s, a run voltage of 15 kV, a capillary temperature of 20 degrees C, and UV detection at 220 nm. A preliminary validation of the method was performed. The assay had good repeatability (RSDs for peaks were 15%), and responses were linear for assay solutions with collagen concentrations from 0.125 to 1.25 mg/ml. The DSCE electropherogram of bovine skin collagen provided a profile of subspecies similar in number and relative abundance to that generated by scanning of Coomassie stained SDS-PAGE gels. PMID- 10857566 TI - Simultaneous determination of zidovudine and lamivudine in human serum using HPLC with tandem mass spectrometry. AB - A method employing high performance liquid chromatography (HPLC) with tandem mass spectrometry (MS) has been developed and validated for the simultaneous determination of clinically relevant levels of zidovudine (AZT) and lamivudine (3TC) in human serum. The method incorporates a fully automated ultrafiltration sample preparation step that replaces the solid-phase extraction step typically used for HPLC with UV detection. The calibration range of the dual-analyte LC MS/MS method is 2.5-2,500 and 2.5-5,000 ng ml-1 for AZT and 3TC, respectively, using 0.25 ml of human serum. The lower limit of quantification was 2.5 ng ml-1 for each analyte, with a chromatographic run time of approximately 6 min. Overall accuracy, expressed as bias, and inter- and intra-assay precision are < +/- 7 and < 10% for AZT, and < +/- 5 and < 12.1% for 3TC over the full concentration ranges. A cross-validation study demonstrated that the LC-MS/MS method afforded equivalent results to established methods consisting of a radioimmuno-assay for AZT and an HPLC-UV method for 3TC. Moreover, the LC-MS/MS was more sensitive, allowed markedly higher-throughput, and required smaller sample volumes (for 3TC only). The validated method has been used to support post-marketing clinical studies for Combivir a combination tablet containing AZT and 3TC. PMID- 10857567 TI - Ranitidine hydrochloride X-ray assay using a neural network. AB - A simple X-ray powder diffractometric (XRD) method with artificial neural networks (ANNs) for data modelling was developed to recognize and quantify two crystal modifications of ranitidine HCl in mixtures and thus, provide information about the solid state of the bulk drug. The method was also used to quantify ranitidine HCl from tablets in the presence of other components. An ANN consisting of three layers of neurons was trained by using a back-propagation learning rule. A sigmoid output function was used in the hidden layer to facilitate non-linear fitting. Unlike other techniques the ANN method described here employed pattern recognition on the entire XRD pattern. Correct classification was mainly influenced by the XRD pattern resolution. It was shown that data transformations improved the quantitative performance when the XRD patterns were not contaminated by other components. Only smoothed X-ray diffractograms were required to distinguish between the two crystalline forms in a mixture. In the case of ranitidine-HCl quantification from tablets, where significant interference with tablet excipients was present, better results were obtained without data transformations. The trained ANN perfectly quantified ranitidine HCI polymorphic forms from mixtures (mean sum of squared error was less than 0.02%) and ranitidine HCl form 1 from tablets (recovery = 98.65). Excellent quantification performance of the ANN analysis. demonstrated in this study, serves as an indication of the broad potential of neural networks in pattern analysis. While the system described has been developed to interpret XRD patterns, peak detection has implications in every chemical application where the recognition of peak-shaped signals in analytical data is important. PMID- 10857568 TI - Use of nitric acid in sample pretreatment for determination of trace elements in various biological samples by ETAAS. AB - Trace elements in liquid biological samples may be determined by direct electrothermal atomic absorption spectrometry (ETAAS). In our previous work it was found that samples containing proteins or DNA may leak out of the graphite tube before the drying step, despite the addition of various modifiers. In order to keep the sample to the graphite tube, samples were diluted before analysis 1 + 1 with 32% v/v nitric acid, or 5 microl of 32% v/v nitric acid was added to the graphite tube before ETAAS determination. Applying the proposed procedure, the concentrations of lead in eluted fractions after gel chromatographic separation of human cerebellar nucleus dentatus supernatant and platinum in isolated DNA samples were determined. The use of nitric acid in sample pretreatment prevent sample leakage out of the graphite tube, provided for even drying and considerably reduced nonspecific absorption in lead determination. The repeatability of measurements was better than + 6%. The accuracy of the procedure was checked by spiking samples. The recoveries for both elements lay between 93- 104%. Nitric acid was found to be a better modifier than TRITON X-100. PMID- 10857569 TI - High-performance liquid chromatography with mass spectrometry detection for quantitating COL-3, a chemically modified tetracycline, in human plasma. AB - COL-3, 6-deoxy-6-demethyl-4-dedimethylamino-tetracycline, is a matrix metalloproteinase inhibitor. A specific and sensitive analytical method was necessary to quantitate the analyte in human plasma. High-performance liquid chromatography with atmospheric pressure chemical ionization mass spectrometry detection was utilized to quantitate COL-3 from 30 to 10,000 ng/ml in two calibration curves: 30-1,500 and 400-10,000 ng/ml. The sample preparation consisted of acetonitrile precipitation for all plasma samples. COL-3 is separated on a Waters Symmetry C-18 (2.1 x 150 mm) column with oxalic acid (0.01 M, pH 2.2)-acetonitrile mobile phase. The total run time was 23 min. Identification of COL-3 and the internal standard was through positive chemical ionization and selective ion monitoring. A quantifying and qualifying ion for COL 3 is used to verify the presence of COL-3 in patient samples. Inter- and intra run mean percent errors for all of the quality controls were less than 18.3', and relative standard deviations were all less than 14.9'% Recovery of COL-3 and the internal standard was approximately 55 and 72', respectively. Freeze thaw stability of COL-3 was variable. This method is suitable for quantifying COL-3 in patient samples and to further characterize the clinical pharmacology of this compound. PMID- 10857570 TI - Simultaneous analysis of methylprednisolone, methylprednisolone succinate, and endogenous corticosterone in rat plasma. AB - A reversed-phase HPLC method is reported for simultaneous quantitation of methylprednisolone (MP), MP succinate (MPS), and endogenous corticosterone (CST) in plasma of rats. Additionally, the 11-keto metabolite of MP (methylprednisone, MPN) is resolved from the other analytes. After addition of internal standard (triamcinolone acetonide: IS) and an initial clean up step, the analytes of interest are extracted into methylene chloride. The steroids are then resolved on a reversed-phase polymer column using a mobile phase of 0.1 M acetate buffer (pH 5.7): acetonitrile (77:23) which is pumped at a flow rate of 1.5 ml min-1. Sample detection was accomplished using an UV detector at a wavelength of 250 nm. All the five components (MPS, MP, MPN, CST and IS) were baseline resolved from each other and other components of plasma. Linear relationships were found between the steroids: IS peak area ratios and plasma concentrations in the range of 0.1-4 mircog ml-1 for MP and MPS and 0.1-1.0 microg ml-1 for MPN and CST. The assay is accurate as intra- and inter-run error values were < +/- 8% for all the components. Further, the intra- and inter-run CVs of the assay were < 16% at all the concentrations and for all the components. The application of the assay was demonstrated after the injection of a single 5 mg kg-1 (MP equivalent) dose of MPS or a macromolecular prodrug of MP to rats. PMID- 10857571 TI - The metabolism and excretion of [14c] 2- and 4-chlorobenzoic acids in the rat. AB - The metabolic fate of [14C]-labelled 2 and 4-chlorobenzoic acids (2- and 4-CBA) has been determined in the rat following intraperitoneal (i.p.) administration at 100 mg/kg to male rats. The major route of elimination for both 2-and 4-CBA was urine with > 80%, of the dose recovered in the initial 0-24 h after administration. Glycine conjugation was found to be the dominant metabolic fate for both [14C] 2- and 4-CBA however, the position of chloro substitution had a clear effect on the extent of metabolism via this route with ortho substitution reducing the extent of metabolism via this pathway. PMID- 10857572 TI - Monitoring of two intravenous immunoglobulin. Preparations for immunoglobulin G subclasses and specific antibodies to bacterial surface antigens and relation with their levels in treated immunodeficient patients. AB - Patients with antibody deficiency disorders are highly susceptible to bacterial infections. Replacement therapy with intravenous immunoglobulin preparations (IVIG) has been established in such patients for two decades. The efficacy of IVIG treatment depends on the amount of functional pathogen-specific antibodies provided. The present study was undertaken to determine the levels of immunoglobulin classes, IgG subclasses, and specific antibodies to bacterial surface antigens in two different IVIG preparations (Sandoglobulin and Gamimmune) and blood sera of IVIG-treated immunodeficient patients. The levels of IgG, IgA, IgM and IgG subclasses were determined in both IVIG preparations and in patients' sera and were compared with those of healthy individuals. Sandoglobulin contained significantly higher concentrations of IgA, IgG, and IgG4 than Gamimmune. The latter contained higher concentrations of IgG1. Patients treated with Gamimmune) had significantly lower concentration of IgG4 as compared with healthy individuals and Sandoglobulin-treated patients. This finding was related to the preparation's composition. Screening of 20 lots from each preparation for antibodies to frequent clinically isolated strains of Escherichia coli, Staphylococcus aureus, S. epidermidis. Klebsiella pneumoniae and Enterococci spp. showed a high lot-to-lot variability. In order to overcome the lot-to-lot variability and correlate the observed effects with each IVIG preparation, the administered IVIG lots were selected so that their titers were in the interval of mean value +/- S.D. for each pathogen. The two tested preparations showed significant differences in their content of specific antibodies that ultimately affected the levels of these antibodies in treated patients. More specifically, Sandoglobulin contained higher levels of antibodies to E. coli and S. epidermidis strains. Infusion of this preparation maintained the respective antibodies in the recipients significantly higher than those of healthy individuals. Gamimmune infusion led to similar and comparable levels. Both IVIG preparations had comparable antibody titers towards K. pneumoniae, provided high amounts of antibodies, and kept recipients' specific IgG at levels significantly higher than those of the healthy individuals. Enterococci spp. specific antibodies were significantly higher in Gamimmune, whereas titers of antibodies towards S. aureus were comparable. Levels of antibodies against both Enterococci spp. and S. epidermidis after administration of both preparations were close to those in healthy individuals. None of the patients developed infection during the time of the study. In conclusion, most of the lots of the two IVIG preparations studied, despite some quantitative differences, provide patients with sufficient amounts of antibodies to bacterial surface antigens that protect them against infections. PMID- 10857573 TI - A surface plasmon resonance method for detecting multiple modes of DNA-ligand interactions. AB - A simple and general surface plasmon resonance (SPR) based method has been developed to detect and quantitate binding of low molecular weight compounds (200 1,200 Da) to double stranded DNA. Several compounds were chosen to probe three different modes of binding interactions, intercalation, minor groove binding and electrostatic interactions. Ethidium bromide (MW 390 Da), a probe of intercalative binding, was tested by plotting the steady state SPR responses measured on a DNA modified surface versus ethidium bromide concentration. The best fit of the binding isotherm gave a Keq of 1.8 x 105 M-1. Co-solvents such as DMSO are often used in activity assays to increase the solubility of poorly water soluble drugs. The effect of DMSO on the ethidium bromide/DNA interaction was also tested by measuring binding in the presence of 0, 1 and 5%, DMSO. No effect on the measured Keq was observed at these DMSO concentrations. The binding of actinomycin (MW 1,255 Da), an antibiotic known to bind DNA through intercalation and minor groove binding, was also tested. The Keq estimated from the steady state responses on a DNA surface was 1.9 x 106 M-1. DAPI (MW 350 Da) (4',6 diamidino-2-phenylindole) a fluorescent probe which binds the minor groove of DNA was also tested and gave a Keq of 1.8 x 106 M-1 measured by SPR. Finally, spermine (MW 202) a compound known to bind DNA through ionic interactions gave the weakest Keq of 1.7 x 104 M-1. All the Keq values measured by SPR and reported for these compounds were in good agreement with literature values measured by other techniques. PMID- 10857574 TI - Anodic adsorptive voltammetric determination of the vitamin 1 (thiamine). PMID- 10857575 TI - Intensity fluctuations of ultrasonic scattering in a highly turbulent flow. AB - Aspects of ultrasound intensity fluctuations backscattered from additive microstructures in a turbulent flow have been investigated theoretically and experimentally for the conditions of a small insonified volume, a high sound frequency and strong turbulence. These conditions are typically found in high resolution Doppler sonar applications. An easily applicable expression for the auto-correlation of scattering intensity fluctuations is obtained by introducing open-channel turbulence theory, a semi-empirical scalar spectrum (including a Batchelor spectrum) and a Gaussian window function. Experiments carried out in a laboratory-clear water, open-channel flow for different turbulence levels verify the underlying assumptions. A good agreement is found with the predictions made with the above-derived expression. The feasibility of extracting flow information from the backscattered intensity fluctuations is discussed. PMID- 10857576 TI - The feasibility of constructing a cylindrical array with a plane rotating beam for interstitial thermal surgery. AB - Thermal surgery has been shown to be a useful therapeutic option when external ultrasound applicators cannot be used as their beam will not reach the target site. If plane transducers are used, the ultrasound beam has to be rotated in order to generate a sufficiently large volume of necrosis. However, rotating deep seated interstitial applicators and controlling their shooting direction presents major technical problems. The purpose of this study is to evaluate the feasibility of constructing a cylindrical array with a plane rotating beam for ablating esophageal tumors by interstitial hyperthermia. The feasibility of such an array has been initially evaluated using a plane array (which is easier to make from a technical point of view). This array was made with a new piezoelectric material because its mechanical properties make it ideal for the construction of a cylindrical array in the future. We showed that the beam of each array element is sufficiently divergent and that cross-coupling is small enough to generate a plane wave from a cylindrical array. In addition, power tests and electro-acoustic efficiency measurements demonstrated that the output was sufficient to induce tissue necrosis in the relevant conditions. PMID- 10857577 TI - Doppler power spectrum from a Gaussian sample volume. AB - A closed-form expression for the Doppler power spectrum due solely to the range of blood velocities passing through a Gaussian sample volume placed anywhere in a vessel under conditions of axisymmetric flow, uniform backscatter and negligible intrinsic spectral broadening has been derived. The formulation presented here allows the independent specification of the sample volume position and width, in the three dimensions, and enables simple estimations of spectral shape for pulsed wave Doppler systems. Simpler expressions were derived for the cases of symmetric sample volume projections onto the vessel cross-section and/or sample volumes centred in the vessel. Closed form expressions were derived for mean frequency and spectral width in the case of a symmetric sample volume projection centred in the vessel. The effects of sample volume size and position on the Doppler spectral width and mean frequency are shown for a range of velocity profiles. PMID- 10857578 TI - Ghosts in the Born images of a layer probed by acoustic waves. AB - The Born inversion of time records of acoustic pulses reflected from a layer object gives rise to the images with ghost-like artifacts that make it difficult to pick out the exact thickness and composition of the object. The origin of these ghosts is explained herein and a means is proposed for their suppression. PMID- 10857579 TI - Tissue structure study through ultrasonic forward scattering. AB - A procedure is demonstrated for characterization of biological tissues at small scattering angles. The power spectra of ultrasonic pulses transmitted through excised tissue samples were measured and compared to the spectra of signals transmitted through a water path. The specimens were examined in two spatial frequency bands by acquiring data at scattering angles of 10 degrees and 20 degrees using 2.25 MHz transducers. Peaks in the measured power spectra are interpreted using two signal models. The medium is modelled either as a periodic structure producing a single spectral peak, or by two discrete targets producing a periodic modulation of the spectrum. The periodic structure model appears to be the more promising method for interpretation of forward-scattered signals. Data acquired from hyperplastic spleen and atheromatous aorta specimens both exhibited increases in pulse-tissue interaction at low spatial frequencies compared to normal specimens of those tissues. This observation is tentatively linked to increases in the size or separation of distributed scattering structures resulting from those pathologies. PMID- 10857580 TI - Xenogeneic transplantation of human spermatogonia. AB - In the last 3 years, several studies have shown that xenogeneic transplantation of rodent spermatogonia is feasible. The treatment of infertile patients with spermatogenic arrest using the injection of immature germ cells has yielded only poor results. We attempted to establish a complete spermatogenetic line in the testes of mutant aspermatogenic (W/Wv) and severe combined immunodeficient mice (SCID) transplanted with germ cells from azoospermic men. Spermatogenic cells were obtained from testicular biopsy specimens of men (average age of 34.3 +/- 9 years) undergoing infertility treatment because of obstructive and non obstructive azoospermia. Testicular tissue was digested with collagenase to promote separation of individual spermatogenic cells. The germ cells were injected into mouse testicular seminiferous tubules using a microneedle (40 microm inner diameter) on a 10 ml syringe. To assess the penetration of the cell suspension into the tubules, trypan blue was used as an indicator. Mice were maintained for 50 to 150 days to allow time for germ cell colonisation and development prior to them being killed. Testes were then fixed for histological examination and approximately 100 cross-sectioned tubules were examined for human spermatogenic cells. A total of 26 testicular cell samples, 16 frozen and 10 fresh, were obtained from 24 men. The origin of the azoospermia was obstructive (OA) in 16 patients and non-obstructive (NOA) in 8 patients. The concentration of spermatogenic cells in the OA group was 6.6 x 10(6) cells/ml, and 1.3 x 10(6) cells/ml in the NOA group (p < 0.01). The different spermatogenic cell types were distributed equally in the OA samples, ranging from spermatogenia to fully developed spermatozoa, but in the NOA group the majority of cells were spermatogonia and spermatocytes. A total of 23 testes from 14 W/Wv mice and 24 testes from 12 SCID mice were injected successfully, as judged by the presence of spermatogenic cells in histological sections of testes removed immediately after the injection. However, sections from the remaining testes examined up to 150 days after injection showed tubules lined with Sertoli cells and xenogeneic germ cells were not found. The reason why the two strains of mouse used as recipients did not allow the implantation of human germ cells is probably due to interspecies specificity involving non-compatible cell adhesion molecules and/or immunological rejection. PMID- 10857581 TI - Acrosomal status and motility of guinea pig spermatozoa during in vitro penetration of the cumulus oophorus. AB - Previous studies have suggested that both acrosome-intact and acrosome-reacted guinea pig sperm are capable of binding to the zona pellucida of cumulus-free oocytes, but the acrosomal status of guinea pig sperm during penetration of the cumulus has not been reported. We made video recordings of the interaction between capacitated guinea pig sperm and cumulus-invested guinea pig oocytes. The videotapes were analysed to identify sperm with hyperactivated motility and to classify the acrosomal status of sperm during penetration of the cumulus and after binding to the zona pellucida. The resolution of the video recordings was not sufficient to recognise sperm with swollen acrosomes. However, sperm that had completed the acrosome reaction were easily identified. Acrosome-reacted sperm were found adherent to the outer boundary of the cumulus, but were never observed to penetrate the cumulus. The percentage of acrosome-intact, hyperactivated sperm was higher in the cumulus oophorus than in culture medium, suggesting that changes in motility were elicited in response to contact with the cumulus. Fully acrosome-reacted sperm were found adherent to the zona pellucida, and solubilised guinea pig zona pellucida was capable of inducing acrosome reactions in capacitated guinea pig sperm. Acrosome-intact sperm were also observed on the zona, but they were not tightly bound and did not have hyperactivated motility, suggesting that these sperm were not functionally capacitated. Our observations demonstrate that guinea pig sperm penetrate the cumulus matrix in an acrosome intact state. Although we did not observe sperm undergoing the acrosome reaction, our observations and experimental data suggest that the acrosome reaction of guinea pig sperm is completed on or near the surface of the zona pellucida. PMID- 10857582 TI - Localisation of phosphorylated MAP kinase during the transition from meiosis I to meiosis II in pig oocytes. AB - Mitogen-activated protein kinase (MAPK) has been reported to be involved in oocyte maturation in all animals so far examined. In the present study we investigate the expression and localisation of active phosphorylated MAPKs (p44ERK1/p42ERK2) during maturation of pig oocytes. In immunoblot analysis using anti-p44ERK1 antibody which recognised both active and inactive forms of p44ERK1 and p42ERK2, we confirmed that MAPKs were phosphorylated around the time of germinal vesicle breakdown (GVBD) and the active phosphorylated MAPKs (pMAKs) were maintained until metaphase II, as has been reported. On immunofluorescent confocal microscopy using anti-pMAPK antibody which recognised only phosphorylated forms of MAPKs, pMAPK was localised at the spindle poles in pig mitotic cells. On the other hand, in pig oocytes, no signal was detected during GV stage. After GVBD, the area around condensed chromosomes was preferentially stained at metaphase I although whole cytoplasm was faintly stained. At early anaphase I, the polar regions of the meiotic spindle were prominently stained. However, during the progression of anaphase I and telophase I pMAPK was detected at the mid-zone of the elongated spindle, gradually becoming concentrated at the centre. Finally, at the time of emission of the first polar body, pMAPK was detected as a ring-like structure between the condensed chromosomes and the first polar body, and the staining was maintained even after the metaphase II spindle was formed. The inhibition of MAPK activity with the MAPK kinase inhibitor U0126 during the meiosis I/meiosis II transition suppressed chromosome separation, first polar body emission and formation of the metaphase II spindle. From these results, we propose that the spindle-associated pMAPKs play an important role in the events occurring during the meiosis I/meiosis II transition, such as chromosome separation, spindle elongation and cleavage furrow formation in pig oocytes. PMID- 10857583 TI - Soybean trypsin inhibitor as a probe for the acrosome reaction in motile cynomolgus macaque sperm. AB - Soybean trypsin inhibitor (SBTI) inhibits the catalytic activity of serine proteases, and has been shown to bind to acrosin, an acrosomal hydrolase which is not exposed on the surface of macaque sperm until after the acrosome reaction. Following activation with caffeine and dibutyryl cAMP, cynomolgus macaque sperm were induced to acrosome react with calcium ionophore A23187 in the presence of SBTI and were fixed for ultrastructural observation. Transmission electron microscopy (TEM) revealed secondary labelling of anti-SBTI-IgG with colloidal gold in association with the acrosomal matrix and fused membranes of sperm undergoing the acrosome reaction, but gold labelling was not observed on acrosome intact sperm. When SBTI was conjugated with the fluorochrome Alexa 488, labelled (acrosome-reacted) sperm showed bright fluorescence that ranged from a patchy or punctate appearance to solid labelling over the region of the acrosomal cap. Following treatment with ionophore, the percentages of total acrosome-reacted sperm (motile and non-motile) as assessed with Alexa-SBTI, fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (FITC-PSA), and TEM were 54.6%, 51.6% and 61.5%, respectively. Measures of acrosomal status with FITC-PSA and Alexa-SBTI were highly correlated (r = 0.94; n = 3). Macaque zonae pellucidae were co-incubated with activated sperm for 1 min and then rinsed in medium containing Alexa-SBTI and immediately observed with epifluorescence microscopy. The mean percentage of Alexa-SBTI-labelled (acrosome-reacted) motile sperm bound to the zona was 45.7 +/- 14 (range: 22-80.4%; n = 4). Fewer than 1% of the motile sperm in suspension surrounding the zonae were acrosome-reacted. Alexa-SBTI had no effect on sperm motility, survival, or zona binding capability. PMID- 10857584 TI - Influence of oviductal cells and conditioned medium on porcine gametes. AB - The aim of this study was to optimise porcine in vitro fertilisation (IVF) with cryopreserved semen with the exploitation of the oviduct secretion. The oocytes were cultured in NCSU37 supplemented with db-cAMP (1 mM), porcine follicular fluid (pFF; 10%), cysteine (0.1 mg/ml) and beta-mercaptoethanol (25 microM) for 44 h (the first 20 h with 10 IU/ml hCG and PMSG). The oviductal epithelial cells (OEC) were cultured in TCM-199 medium (with 10% FCS, 0.2 mM pyruvate and 50 microg/ml gentamicin) for 48 h. To determine the effects of OEC and conditioned medium, oocytes were separated into five groups for the last 3 h of maturation and placed in: fresh maturation medium (controls), OEC-cNCSU with OEC in the maturation medium for 24 h; OEC-fNUSU with fresh OEC in maturation medium; cTCM with TCM-199 conditioned with OEC for 48 h; or fTCM with fresh TCM-199. Results indicate that OEC-cNCSU and OEC-fNCSU increase the number of oocytes reaching the two pronucleus (2PN) stage (p < 0.01) and decrease the polyspermy rate (p < 0.01) compared with controls. The rates are significantly lower than controls when cTCM and fTCM were used (p < 0.01). As regards blastocyst rates, an increase was observed in the OEC-cNCSU and cTCM groups (p < 0.05). For the second experiment, spermatozoa were incubated with OEC in IVF medium (mTBM medium supplemented with 0.1% BSA) without caffeine for 4 h prior to IVF. Results indicate that sperm treatment with OEC increases the 2PN rate (p < 0.01) compared with controls and reduces the polyspermy rate (p < 0.01). In conclusion, our study shows that co incubation of OEC with both oocytes and sperm before IVF reduces polyspermy rates and improves embryo development. PMID- 10857585 TI - Progression of mouse oocytes from metaphase I to metaphase II is inhibited by fusion with G2 cells. AB - We show that in contrast to metaphase II oocytes, metaphase I oocytes cannot be activated by fusion with the zygote. Fusion of metaphase I oocytes with G2 zygotes was followed by premature chromosome condensation, with 60% of the hybrids becoming arrested at metaphase I, the remainder progressing and arresting at metaphase II. Hybrids of metaphase I oocytes and M-phase zygotes underwent accelerated maturation, but all arrested at metaphase II. In both cases the arrest could be overcome by treatment with the parthenogenetic activators ethanol and cycloheximide. We discuss these findings in relation to the possibility that the metaphase I oocyte contains cytostatic factor activity that is activated by its zygotic partner. Alternatively, the G2 zygote may provide an inhibitor of anaphase, normally never present in the metaphase I oocyte and which is absent from the M-phase zygote. PMID- 10857586 TI - Degradation of pig cyclin B1 molecules precedes MAP kinase dephosphorylation during fertilisation of the oocytes. AB - Pig oocytes at metaphase II were activated by penetration of spermatozoa in cycloheximide-free and cycloheximide-containing fertilisation media. The precise nuclear stage, and the kinetics of degradation of cyclin B1 and dephosphorylation of MAP kinase were assessed after insemination. After maturation culture, 96% of oocytes reached metaphase II. At 6 h after insemination in cycloheximide-free medium, 68% of the oocytes were activated and had progressed to anaphase II or beyond. After 8 h, 89% of the oocytes were activated: a female pronucleus had formed and the heads of penetrating spermatozoa had enlarged and changed to male pronuclei. In the cycloheximide-containing medium, activation of oocytes started earlier than in cycloheximide-free medium. After 4 h, 43% of the oocytes were activated, and the percentage increased to 97% after 6 h. Pig cyclin B1 disappeared in the oocytes at 6 h after insemination in both cycloheximide containing and cycloheximide-free media. Pig oocytes at metaphase II contained two types of MAP kinase--ERK 1 and ERK 2--in their active phosphorylated forms. At 8 h after insemination ERK 2 changed to the fast-migrating inactive form in the oocytes cultured in both cycloheximide-containing and cycloheximide-free media, although the shift-down was not complete. The change was delayed by 2 h after the degradation of cyclin B1 molecules. These results demonstrate that degradation of pig cyclin B1 molecules corresponds to the transition of the oocytes from metaphase II arrest to anaphase II/telophase II and was followed by MAP kinase dephosphorylation. PMID- 10857587 TI - Inhibition of the synthesis of glycosphingolipid by a ceramide analogue (PPMP) in the gastrulation of Bufo arenarum. AB - In the present study the role of glycosphingolipids (GSL) in amphibian development was investigated. We analysed the de novo synthesis of neutral GSL and gangliosides through the initial stages of Bufo arenarum embryo development and their participation during gastrulation using 1-phenyl-2-palmitoyl-3 morpholino-1-propanol (PPMP), a potent inhibitor of glucosylceramide synthase. Ganglioside synthesis began at the blastula stage and reached a maximum during gastrulation (stages 10-12) while neutral GSL synthesis showed a slight gradual increase, the former being quantitatively more significant than the latter. Ganglioside synthesis was reduced by 90% while neutral GSL synthesis was inhibited by 65% when embryos at blastula stage were cultured for 24 h in 20 microM PPMP. The depletion of GSL from amphibian embryos induced an abnormal gastrulation in a dose-dependent manner. We found that PPMP had a pronounced effect on development since no embryos exhibited normal gastrulation; their developmental rate either slowed down or, more often, became totally arrested. Morphological analysis of arrested embryos revealed inhibition of the gastrulation morphogenetic movements. Analysis of mesodermal cell morphology in those embryos showed a severe decrease in the number and complexity of cellular extensions such as filopodia and lamellipodia. Mesodermal cells isolated from PPMP-treated embryos had very low adhesion percentages. Our results suggest that glycosphingolipids participate in Bufo arenarum gastrulation, probably through their involvement in cell adhesion events. PMID- 10857588 TI - Effect of gap junction uncoupling in full-grown Bufo arenarum ovarian follicles: participation of cAMP in meiotic arrest. AB - The aim of the present study was to determine the presence of the connexins Cx43, Cx32 and Cx26 in Bufo arenarum ovarian follicles during the breeding season as well as to analyse the possible alterations in the meiotic process when connexins are blocked by specific antibodies. Western blot analysis revealed that the Cx43 and Cx32 proteins were present but not Cx26. We demonstrated that the anti-Cx43 and anti-Cx32 antibodies produced the uncoupling of the gap junctions. When these junctions are blocked the maturation process is triggered in the oocytes. We determined that dbcAMP exerts an inhibitory effect on the maturation induced by the uncoupling of the gap junctions when the oocytes are injected or pretreated with this metabolite. We propose the idea that cAMP is the regulatory molecule in meiotic arrest in this amphibian species. PMID- 10857589 TI - Binding of porcine sperm plasma membrane proteins to sheep, hamster and mouse oocyte plasma membrane. AB - Four porcine sperm plasma membrane proteins were previously identified as putative ligands for the oocyte plasma membrane. The present study examined the binding of these proteins and two additional porcine sperm membrane proteins to oocytes from sheep, mice and hamsters as a first step in assessing potential conservation of these putative sperm ligands across species and across mammalian orders. Plasma membrane vesicles were isolated from porcine sperm, solubilised, and the proteins separated by one-dimensional gel electrophoresis. The 7, 27, 39 and 62 kDa porcine sperm protein bands demonstrating predominant binding of the porcine oocyte plasma membrane on ligand blots, a 90 kDa protein band demonstrating minor binding, and a 97 kDa protein band that did not bind the oocyte plasma membrane probe were electroeluted. Proteins were biotinylated, and incubated with zona-free oocytes. Bound biotinylated protein was labelled with fluorescent avidin and the oocytes examined with a confocal microscope. The 7 kDa, 27 kDa and the 39 kDa proteins bound to the sheep oocytes but not to a majority of the hamster or mouse oocytes. The 62 kDa protein bound to sheep oocytes and mouse oocytes but not to a majority of the hamster oocytes. The 90 kDa protein bound to oocytes from all three species. The 97 kDa protein, which did not recognise the porcine oocyte probe on a Western ligand blot, did not bind to oocytes from any species and served as a negative control. These observations are consistent with significant conservation of molecule and function among species within the same mammalian order. Hence, one species may be a good model for other species from the same order. Only limited conservation of binding activity of porcine sperm plasma membrane proteins to rodent oocytes was observed, suggesting a greater divergence either in molecular structure or in function among species from different orders. PMID- 10857590 TI - New methods for detecting Salmonella. PMID- 10857591 TI - Adding dimensions to MS with electron monochromators. PMID- 10857592 TI - Measuring insulin secretion from single cells. PMID- 10857593 TI - Sandia introduces lab-on-a-chip prototype. PMID- 10857594 TI - HPLC looks for orthogonal techniques. PMID- 10857595 TI - Is your T-shirt toxic? PMID- 10857596 TI - Academic quality management PMID- 10857597 TI - Problematic standards. PMID- 10857598 TI - Drugs on money. PMID- 10857599 TI - Biomolecular Interaction Analysis Mass Spectometry. BIA/MS can detect and characterize protiens in complex biological fluids at the low- to subfemtomole level. PMID- 10857600 TI - HPLC: past and present. PMID- 10857601 TI - E-noses keep an eye on the future. PMID- 10857602 TI - STAT: a saccharide topology analysis tool used in combination with tandem mass spectrometry. AB - Sequential stages of mass spectrometry (MSn) have the potential to provide a great deal of structural information in glycan analysis. The saccharide topology analysis tool (STAT) presented here is a Web-based computational program that can quickly extract sequence information from a set of MSn spectra for an oligosaccharide of up to 10 residues. After information such as precursor ion mass, possible monosaccharide moieties, charge carrier, and product ion mass has been input, all possible connectivities are generated and evaluated against the MSn data. The list of possible structures is given a rating based on the likelihood that it is the correct sequence. Examples are given to demonstrate the feasibility of applying STAT to MSn data generated from bacterial lipooligosaccharides and an N-linked glycan. The major advantage of STAT is that the list of possible structures is generated quickly and the rating system pushes the more likely structures to the top of the list. Combining the data generated by STAT with data on the branching patterns of the glycan serves to eliminate all but a handful of structures. These remaining structures could then be used to guide further structural analysis. PMID- 10857603 TI - De novo peptide sequencing by two-dimensional fragment correlation mass spectrometry. AB - A novel concept of two-dimensional fragment correlation mass spectrometry and its application to peptide sequencing is described. The daughter ion (MS2) spectrum of a peptide contains the sequence information of the peptide. However, deciphering the MS2 spectrum, and thus deriving the peptide sequence is complex because of the difficulty in distinguishing the N-terminal fragments (e.g., b series) from the C-terminal fragments (e.g., y series). By taking a granddaughter ion (MS3) spectrum of a particular daughter ion, all fragment ions of the opposite terminus are eliminated in the MS3 spectrum. However, some internal fragments of the peptide will appear in the MS3 spectrum. Because internal fragments are rarely present in the MS2 spectrum, the intersection (a spectrum containing peaks that are present in both spectra) of the MS2 and MS3 spectra should contain only fragments of the same terminal type. A two-dimensional plot of the MS2 spectrum versus the intersection spectra (2-D fragment correlation mass spectrum) often gives enough information to derive the complete sequence of a peptide. This paper describes this novel technique and its application in sequencing cytochrome c and apomyoglobin. For a tryptic digest of cytochrome c, approximately 78% of the protein sequence was determined. For the Glu-C/tryptic digest of apomyoglobin, approximately 66% of the protein sequence was determined. PMID- 10857604 TI - Attenuation of matrix effects in inductively coupled plasma mass spectrometry with a supplemental electron source inside the skimmer AB - Electrons are added from a heated filament at the base of the skimmer to reduce the space charge repulsion in the ion beam. This technique improves the analyte sensitivity moderately and also minimizes the matrix effects caused by other elements in the sample significantly. The suppression of signal for even the most troublesome combination of light analyte and heavy matrix elements can be attenuated from 90 to 99% to only 2-10% for 2 mM matrix solutions with an ultrasonic nebulizer. The supplemental electron current can be adjusted to "titrate" out the matrix effects as desired. PMID- 10857605 TI - Reduction of space charge effects in inductively coupled plasma mass spectrometry using a supplemental electron source inside the skimmer: ion transmission and mass spectral characteristics AB - An electron source consisting of a heated filament has been added to the skimmer to suppress space charge effects in inductively coupled plasma mass spectrometry (ICPMS). Electrons from this source can reduce the space charge repulsion between the positive ions in the ion beam. As a result, ion transmission efficiency and analyte ion sensitivities are significantly improved across the full mass range. MO+/M+ abundance ratios are not affected, M2+/M+ abundance ratios increase only slightly, and no new background ions are created by this electron injection technique. PMID- 10857606 TI - Laser activation voltammetry: selective removal of reduced forms of methyl viologen deposited on glassy carbon and boron-doped diamond electrodes AB - The effect of high-intensity laser pulses on the reduction of methyl viologen at glassy carbon electrodes in aqueous solution is investigated using laser activation voltammetry (LAV) under both channel flow and no-flow conditions and compared with the effect of conventional variable-temperature voltammetry. The reduction proceeds in two consecutive one-electron steps, and the neutral two electron-reduction product of methyl viologen is shown by voltammetry and in situ optical microscopy to form two types of deposits, amorphous and crystalline, on the electrode surface. Laser activation voltammetry using a 10 Hz pulsed Nd-YAG 532 nm laser is shown to remove the deposits from the electrode surface at different laser intensities: the amorphous material is more easily ablated than the crystalline deposit. By conventional variable-temperature voltammetry, it is shown that the two stripping peaks disappear as the temperature is increased. However, with conventional heating, the opposite ease of removal is detected compared to the case of laser activation voltammetry: the stripping response associated with the crystalline material disappears at lower temperatures compared to that for the amorphous material. In the presence of high-intensity laser pulses (>0.17 W cm(-2)), glassy carbon surfaces are damaged and the voltammetric characteristics become poor. It is shown that, by the employment of a thin-film boron-doped diamond electrode grown using a chemical vapor deposition procedure on a tungsten substrate, much higher laser intensities can be applied and well-defined LAV signals can be obtained without deactivation of the electrode. PMID- 10857607 TI - Heterogeneous immunosensing using antigen and antibody monolayers on gold surfaces with electrochemical and scanning probe detection. AB - We report the use of antibody and antigen monolayer immunosurfaces as detection elements in a competitive heterogeneous immunoassay employing either electrochemical or scanning probe detection. Antibody or antigen monolayers were prepared by covalent attachment of the desired immunoreagent to a two-component self-assembled monolayer via amide linkages. More specifically, mixed monolayers of a carboxylic acid-terminated thiol (thioctic acid) and a methyl-terminated thiol (butanethiol) were used to control the surface epitope density. The microscopic structure of the resulting antibody and antigen arrays was characterized by AFM (atomic force microscopy). Individual, surface-confined rabbit IgG antibodies could be directly imaged in contact mode. The average height of the capture antibodies was found to be 7.1 nm; the average antibody diameter, after correcting for tip convolution effects, was determined to be between 7 and 10 nm. The surface epitope density could be varied over approximately 2 orders of magnitude by changing the composition of the mixed monolayer. AFM was also used to characterize the antibody-antigen binding characteristics of these immunosurfaces, and an average binding efficiency of 22.8% was measured for rabbit IgG antibody arrays. In the second part of this study, the electrochemical detection scheme originally developed by Heineman and co-workers was adapted to our system. A calibration data set was measured, and the linear least-squares correlation coefficient (R2) was found to be 0.993. Finally, the electrochemical and scanning probe detection modes were directly compared. We find an excellent correlation between the surface density of antibody-antigen complexes measured by AFM and the electrochemical response of the same immunosurfaces. PMID- 10857608 TI - Ion-selective electrode for dodecyldimethylamine oxide: a "twice-Nernstian" slope for the determination of a neutral component AB - An electrode originally sensitive to dodecyltrimethylammonium (DTA+) was proven to be sensitive to dodecyldimethylamine oxide (DDAO), a surfactant with acidobasic properties. The response of the electrode was tested from pH 2 to 9.3. Its slope is Nernstian when the surfactant is entirely protonated. At a pH where the molecule is mainly under the neutral form, the electrode responds with a "twice-Nernstian" slope around 120 mV/decade. The validity of this electrode for measurements was checked by confronting the evolution of the critical micelle concentration of DDAO vs pH with data already published and by determining the complexation constant of DDAO and beta-cyclodextrin. A possible explanation of the "twice-Nernstian" slope, using a dimer of DDAO is proposed. PMID- 10857609 TI - Scanning electrochemical microscopy. Theory of the feedback mode for hemispherical ultramicroelectrodes: steady-state and transient behavior AB - This contribution represents the first comprehensive attempt to treat complex geometry configurations of the scanning electrochemical microscope (SECM) using the alternating direction implicit finite difference method (ADIFDM). Specifically, ADIFDM is used to simulate the steady-state as well as the transient (chronoamperometric) behavior of a hemispherical ultramicroelectrode (UME) tip of the SECM. The feedback effect in this configuration is less pronounced as compared with a disk-shaped UME system. The differences between the two systems are discussed. Analytical approximations for the steady-state behavior and for characteristic features of the transient behavior are suggested. Finally, experimental feedback currents measured above a conductor and an insulator are in excellent agreement with the theory. PMID- 10857610 TI - PVC-based 1,3,5-trithiane sensor for cerium(III) ions AB - A PVC membrane sensor for cerium(III) ions based on 1,3,5-trithiane as membrane carrier was prepared. The sensor has a linear dynamic range of 1.0 x 10(-1)-5.0 x 10(-5) M, with a Nernstian slope of 19.4+/-0.4 mV decade(-1), and a detection limit 3.0 x 10(-5) M. It has a fast response time of < 15 s and can be used for at least 5 months without any considerable divergence in potentials. The proposed sensor revealed comparatively good selectivities with respect to alkali, alkaline earth, and some transition and heavy metal ions and could be used in a pH range of 5.0-8.0. It was used as an indicator electrode in potentiometric titration of oxalate and fluoride ions and in determination of F- ion in some mouth wash preparations. PMID- 10857611 TI - Oxidation-state speciation of AB - The analytical utility of chemically modified microelectrodes for oxidation-state speciation of redox couples by cyclic voltammetry has been explored. [Re(I)(DMPE)3]+/[Re(II)(DMPE)3]2+, where DMPE = 1,2-bis(dimethylphosphino)ethane, was studied at carbon-fiber microelectrodes of approximately 5 microm in radius coated with Nafion-entrapped solgel-derived silica (Nafion-silica) composite. The results are compared with cyclic voltammetry of [Fe(CN)6]3-/[Fe(CN)6]4- at bare carbon-fiber microelectrodes. At both microelectrodes, the cathodic and anodic limiting currents are linearly proportional to the concentrations of the reducible and oxidizable species of a redox couple, respectively. The shape of the cyclic voltammogram and the magnitude of the steady-state limiting current are not affected by the potential at which the scan starts. Speciation of both forms of a redox couple could be achieved voltammetrically at the microelectrodes. However, a considerably slower scan rate was required to achieve steady state at the modified electrode because of the smaller diffusion coefficients of [Re(I)(DMPE)3]+ and [Re(II)(DMPE)3]2+ in the Nafion-silica composite. The detection limit at the modified electrode was considerably lower (5 x 10(-9) M for [Re(I)(DMPE)3]+) than at the bare electrode (6 x 10(-5) M for [Fe(CN)6]3- and [Fe(CN)6]4-) because of the substantial preconcentration of [Re(I)(DMPE)3]+ by the Nafion-silica composite. PMID- 10857612 TI - Quantitative analysis using steady-state free precession nuclear magnetic resonance AB - The use of steady-state free precession nuclear magnetic resonance (NMR) for quantitative analysis in low magnetic field is investigated and shown to exhibit substantial advantages compared to more conventional NMR methods. With only minor additional requirements, the technique permits a considerable increase in signal to-noise ratio for a given acquisition time. The experimental conditions needed for implementation and optimization of the acquisition parameters are explored and shown to be easily accessible with unsophisticated equipment. The applicability of the technique for quantitative analysis of samples with a range of relaxation rates is tested using various examples of practical interest. Highly reproducible results can be obtained much faster and without any special sample-dependent adjustments. PMID- 10857613 TI - Electrothermal atomic absorption spectrometric diagnosis of familial hypercholesterolemia. AB - We have developed a new nonradioactive assay to identify human low-density lipoprotein receptor defects. It is based on the incubation of cultured cells with colloidal gold-LDL conjugates and quantitation of the gold associated with the cells by electrothermal atomic absorption spectrometry. After an oxidative treatment with nitric and hydrochloric acids, the biological matrix interferes neither with the gold recovery nor with the gold measurements, which are linear, at least from 0.15 to 3 ng of gold. When cells expressing a functional LDL receptor are incubated with increasing amounts of colloidal-gold LDL conjugates, the obtained saturation curve parallels that described when [125I]LDL is used as ligand. Moreover, this new assay allows us to clearly distinguish among fibroblasts from normal subjects or from heterozygous or homozygous patients of familial hypercholesterolemia, a very common autosomal disease. The assay is easy to perform, is sensitive, and avoids the use of radioactive compounds. Therefore, it could be successfully employed in the clinical diagnosis of this disease. Furthermore, since the methodology developed here can be applied to quantify the association of other gold-conjugated ligands to cells, it could have a widespread use in a variety of clinical and basic research studies. PMID- 10857614 TI - Determination of motility forces of bovine sperm cells using an "optical funnel". AB - An optical funnel, a new technique for the evaluation of the force of a microorganism, was applied to the determination of the motility force of bovine sperm cells. In this approach, sperm cells, suspended in an aqueous solution, are introduced into a flow cell, to which radiation pressure is applied from the direction opposite to a medium flow. The sperm cell, which is moving in a stream, is captured by radiation pressure and forced to move to the position at which the force induced by the laser radiation is equal to the force induced by a medium flow. The sperm cell then escapes by its own power on the way to this equilibrium (entrapping) position. The radiation force increases with decreasing distance from the focal point, and as a result, the force of the sperm cell can be determined by measuring the position where the sperm cell escaped against the laser irradiation field. The motility force of the sperm cell was measured in aqueous solution at different pH values and potassium ion concentrations. It was possible to measure more than 250 sperm cells in 3 h. Thus, the optical funnel has potential for use as a rapid and repetitive means for the determination of the motility force of the sperm cell. PMID- 10857615 TI - Molecular imprinting of biotin derivatives and its application to competitive binding assay using nonisotopic labeled ligands. AB - Synthetic biotin-binding polymers were prepared by molecular imprinting. Methacrylic acid (MAA) was copolymerized with ethylene glycol dimethacrylate in the presence of biotin methyl ester (B-Me) in chloroform. Hydrogen-bonding-based complexation of B-Me with MAA generates the binding sites complementary to B-Me after extracting B-Me from the resulting copolymers. Data from NMR titration suggest a one-to-one prepolymerization complex formation of B-Me with MAA in chloroform. A possible complex structure was estimated by docking of the most stable conformers by intermolecular Monte Carlo conformational search under the assumption of a one-to-one association. The selectivity of the imprinted polymers was investigated and an imprinted polymer-based competitive binding assay for B Me was demonstrated using biotin p-nitrophenyl ester as a nonisotopic-labeled ligand. PMID- 10857616 TI - Chlorocatechol detection based on a clc operon/reporter gene system. AB - A sensitive and selective sensing system for chlorocatechols (3-chlorocatechol and 4-chlorocatechol) was developed based on Pseudomonas putida bacteria harboring the plasmid pSMM50R-B'. In this plasmid, the regulatory protein of the clc operon, ClcR, controls the expression of the reporter enzyme beta galactosidase. When bacteria containing components of the clc operon are grown in the presence of chlorocatechols, ClcR activates the clcA promoter, which is located upstream from the beta-galactosidase gene. Thus, the concentration of chlorocatechols can be related to the production of beta-galactosidase in the bacteria. The concentration of beta-galactosidase expressed in the bacteria was determined by measuring the chemiluminescence signal emitted with the use of a 1,2-dioxetane substrate. ClcR has a high specificity for chlorocatechols and provides the sensing system with high selectivity. This was demonstrated by evaluating several structurally related organic compounds as potential interfering agents. Both 3-chlorocatechol and 4-chlorocatechol can be detected with this sensing system at concentrations as low as 8 x 10(-10) and 2 x 10(-9) M, respectively, using a 2-h induction period. In the case of 3-chlorocatechol, a highly selective sensing system was developed that can detect this species at concentrations as low as 6 x 10(-8) M after a 5-min induction period; the presence of 4-chlorocatechol at concentrations as high as 2 x 10(-4) M did not interfere with this system. PMID- 10857617 TI - Electron monochromator mass spectrometry for the analysis of whole bacteria and bacterial spores. AB - Spores from a variety of Bacillus species were analyzed with direct probe mass spectrometry using an electron monochromator to select electrons of distinct energies for ionization. Electron energies were chosen to match the electron capture energies of taxonomically important compounds such as dipicolinic acid and fatty acids. Previous negative ion interferences were not observed when the monochromator was used, and the signal-to-noise ratio of targeted compounds was significantly enhanced using this approach. To demonstrate the selectivity of the technique, the monochromator was swept over a range of electron energies while monitoring the masses of compounds with known electron capture energies. Scanning the monochromator while the mass spectrometer was operated in single-ion mode enabled dipicolinic acid to be detected in 10(5) spores. The results presented here demonstrate the utility of the electron monochromator for selectively ionizing compounds directly in bacteria and bacterial spores. PMID- 10857618 TI - Evaluation of binding selectivities of caged crown ligands toward heavy metals by electrospray ionization/quadrupole ion trap mass spectrometry AB - Electrospray ionization (ESI)/quadrupole ion trap mass spectrometry is used to evaluate the heavy metal binding selectivities of five caged crown ethers and two polyether reference compounds in methanol solution. The binding preferences for Hg2+, Pb2+, Cd2+, and Cu2+ were analyzed by comparison of ESI mass spectral intensities with the aim of developing this method for the rapid screening of binding selectivities of new synthetic ligands. The cage compounds preferentially bind Hg2+, except for the cage cryptand derivative, which favors Pb2+. The preference for Hg2+ stems from the favorable positioning of the nitrogen or sulfur atoms for linear coordination of Hg2+, whereas the cryptand derivative favors Pb2+ because of its larger cavity size. The counterions of the metal salts influence the type of complexes observed in the ESI mass spectra because the strengths of the metal-anion bonds affect retention of the anion in the complexes. PMID- 10857619 TI - Bioanalytical high-throughput selected reaction monitoring-LC/MS determination of selected estrogen receptor modulators in human plasma: 2000 samples/day. AB - The high-throughput determination of small molecules in biological matrixes has become an important part of drug discovery. This work shows that increased throughput LC/MS/MS techniques can be used for the analysis of selected estrogen receptor modulators in human plasma where more than 2000 samples may be analyzed in a 24-h period. The compounds used to demonstrate the high-throughput methodology include tamoxifen, raloxifene, 4-hydroxytamoxifen, nafoxidine, and idoxifene. Tamoxifen and raloxifene are used in both breast cancer therapy and osteoporosis and have shown prophylactic potential for the reduction of the risk of breast cancer. The described strategy provides LC/MS/MS separation and quantitation for each of the five test articles in control human plasma. The method includes sample preparation employing liquid-liquid extraction in the 96 well format, an LC separation of the five compounds in less than 30 s, and selected reaction monitoring detection from low nano- to microgram per milliter levels. Precision and accuracy are determined where each 96-well plate is considered a typical "tray" having calibration standards and quality control (QC) samples dispersed through each plate. A concept is introduced where 24 96-well plates analyzed in 1 day is considered a "grand tray", and the method is cross validated with standards placed only at the beginning of the first plate and the end of the last plate. Using idoxifene-d5 as an internal standard, the results obtained for idoxifene and tamoxifen satisfy current bioanalytical method validation criteria on two separate days where 2112 and 2304 samples were run, respectively. Method validation included 24-h autosampler stability and one freeze-thaw cycle stability for the extracts. Idoxifene showed acceptable results with accuracy ranging from 0.3% for the high quality control (QC) to 15.4% for the low QC and precision of 3.6%-13.9% relative standard deviation. Tamoxifen showed accuracy ranging from 1.6% to 13.8% and precision from 7.8% to 15.2%. The linear dynamic range for these compounds was 3 orders of magnitude. The limit of quantification was 5 and 50 ng/ mL for tamoxifen and idoxifene, respectively. The other compounds in this study in general satisfy the more relaxed bioanalytical acceptance criteria for modern drug discovery. It is suggested that the quantification levels reported in this high-throughput analysis example are adequate for many drug discovery and related early pharmaceutical studies. PMID- 10857620 TI - Helium plasma source time-of-flight mass spectrometry: off-cone sampling for elemental analysis AB - In this paper, an atmospheric pressure, helium microwave-induced plasma (MIP) ion source coupled with an orthogonal acceleration time-of-flight mass spectrometer (TOFMS) is explored for elemental analysis. Studies of the relationship between ion signals and sampling distance of the MS reveal that background signals can be suppressed dramatically without sacrificing the signal intensities of analytes when the microwave plasma plume is off the tip of the sampler orifice. This "off cone" ion sampling mode provides a technique to obtain nearly "clean" background spectra and, thus, eliminates the spectral interference from entrainment air and the working-gas species, making it possible to sensitively determine isotopes that suffered from spectral interference in ICPMS and MIPMS (such as 40Ca, 52Cr, 55Mn, and 56Fe). On the other hand, since the high-temperature plasma is kept away from the sampler aperture, off-cone sampling places little demand on the cooling device and the lifetime of the sampler plate can be extended. The instrumental system can provide a fairly good mass resolution of 1100 (fwhm). The detection limits (3sigma) in the tens of picograms per milliliter level for the elements studied can be achieved with a digital oscilloscope. These detection limits can be easily improved with an advanced detection system, which is currently available in commercial markets. PMID- 10857621 TI - A multimicrospray nebulizer for microwave-induced plasma mass spectrometry AB - We have developed a nebulizer, called a multimicrospray nebulizer (MMSN), that efficiently introduces analytes for plasma mass spectrometry and plasma emission spectrometry. In this nebulizer, both the sample solution and the nebulizer gas are divided into several streams to produce a multispray. That is, the MMSN is a nebulizer that contains several micronebulization units, each unit including an orifice for passing the nebulizer gas and a capillary for introducing a sample solution. The microspray from each micronebulization unit can be operated at a microliter per minute sample uptake rate to achieve high nebulization efficiency. The multimicrospray nebulizer is capable of introducing more analyte to the plasma compared with a single-orifice micronebulizer, which has a very low sample uptake rate. In this work, an MMSN with three orifices was found to be suitable for microwave-induced plasma mass spectrometry (MIP-MS). The sample uptake rate can be varied within a range of 5-250 microL/min. Therefore, the nebulizer is unique in its ability to deal with various sample volumes and provide high nebulization efficiency. The sensitivity for all elements obtained with the MMSN was higher than that obtained with a conventional concentric nebulizer (CCN), which is difficult to achieve with other types of microintroduction nebulizers. For most elements, the MIP-MS sensitivity was improved about 2-fold at a sample uptake rate of 150 microL/min, a much lower rate than that for the CCN (usually 0.5-1.5 mL/min). The sensitivity for arsenic was improved by a factor of 5. The relative standard deviation was found to be less than 2.0%. PMID- 10857622 TI - Speciation of sodium nitrate and sodium nitrite using kiloelectronvolt energy atomic and polyatomic and megaelectronvolt energy atomic projectiles with secondary ion mass spectrometry AB - The negative-ion mass spectra produced by kiloelectronvolt energy (CsI)nCs+ (n = 0-2) and megaelectronvolt energy 252Cf fission fragment projectile impacts on NaNO3 and NaNO2 were collected and compared. The mass spectra generated by impacts of the kiloelectronvolt polyatomic primary ions on NaNO3 were markedly different from those derived from the fission fragment impacts, featuring higher relative intensities of nitrate (NO3-) specific secondary ions (those that reflect the sample stoichiometry). The most prominent secondary ion (SI) peaks produced from NaNO3 by the kiloelectronvolt energy projectiles were NO3- and Na(NO3)2-, both of which relate directly back to the chemical composition of the staring material. Likewise, the most prominent peaks produced by the kiloelectronvolt energy polyatomic projectile impacts on NaNO2 were NO2- and Na(NO2)2-. The fission fragment projectiles produced SI spectra from NaNO3 that were dominated by signals characteristic more of NaNO2, indicating that the megaelectronvolt energy ions induce considerable degradation of the nitrate solid. In addition, the fission fragment projectile produced relative negative SI intensity distributions that are remarkably similar to those reported in earlier studies of the use of laser desorption to produce SI signals from NaNO3. Of the projectiles examined in this study, the 20 keV (CsI)Cs+ projectile generated negative-ion mass spectra that best differentiated NaNO3 and NaNO2, primarily by producing a base peak in the NaNO3 spectrum that was unambiguously representative of the original sample stoichiometry. PMID- 10857623 TI - Characterization of microorganisms and biomarker development from global ESI MS/MS analyses of cell lysates. AB - The capability for sensitive and accurate identification of microorganisms has potential applications that include the monitoring of industrial bioprocessing operations, food safety analyses, disease diagnosis, and detection of potential biological hazards. Efforts based upon matrix-assisted laser desorption/ionization mass spectrometry to detect and identify specific microorganisms have been actively pursued for several years. We report a new method being developed to select useful biomarkers for the identification of microorganisms based upon electrospray ionization (ESI)-ion trap mass spectrometry. Crude cell lysates are processed using a recently developed dualmicrodialysis device and then directly infused into an ion trap MS. The low ESI flow rate and precursor ion accumulation capability of the ion trap MS enables high-sensitivity MS/MS analyses. Precursor ions are automatically selected and analyzed using tandem MS (MS/MS) to produce "global" MS/MS surveys and processed to yield two-dimensional MS/MS spectral displays. Such global MS/MS surveys are demonstrated for Escherichia coli lysates. The distinctive MS/MS spectral patterns can be used to identify mass spectrometric-detected species useful as biomarkers, which then provide a basis for confident microorganism identification. The results presented demonstrate the application of this method for the identification of microorganisms, as well as for detection of bacteriophage MS2 in the presence of a large excess of E. coli. PMID- 10857624 TI - ProFound: an expert system for protein identification using mass spectrometric peptide mapping information. AB - We describe the protein search engine "ProFound", which employs a Bayesian algorithm to identify proteins from protein databases using mass spectrometric peptide mapping data. The algorithm ranks protein candidates by taking into account individual properties of each protein in the database as well as other information relevant to the peptide mapping experiment. The program consistently identifies the correct protein(s) even when the data quality is relatively low or when the sample consists of a simple mixture of proteins. Illustrative examples of protein identifications are provided. PMID- 10857625 TI - Characterization of polyether and polyester polyurethane soft blocks using MALDI mass spectrometry AB - Selective degradation reactions combined with MALDI analysis have been applied for molecular weight (MW) determination of polyether and polyester polyurethane (PUR) soft blocks. Selective degradation allows recovery of the polyols, and direct observation of the soft block oligomer distribution is possible for the first time by using MALDI. Ethanolamine is applied for polyether PUR degradation. MALDI analysis indicates that the recovered polytetrahydrofuran (pTHF) MW distribution is nearly identical to the unreacted pTHF material. Reduction in the ethanolamine reaction time allows observation of oligomer ions containing the diisocyanate linkage, which provide identification of the diisocyanate. Ethanolamine is not used for polyester PUR's degradation because the ester bonds will be cleaved. Therefore, phenylisocyanate is applied for polyester PUR degradation. Polybutylene adipate (pBA) oligomers were directly observed in the MALDI spectra of the degraded pBA-PUR samples. Comparison of the degraded pBA-PUR oligomer distribution with the unreacted pBA material indicates that low-mass oligomers are less abundant in the degraded pBA-PURs. Oligomer ions containing the diisocyanate linkage are also observed in the spectrum, providing a means for identifying the diisocyanate used for PUR syntheses. Size-exclusion chromatography (SEC) was combined with MALDI to provide accurate MW determination. Narrow MW fractions of the degraded and unreacted polyols were collected and analyzed by MALDI. This method allows precise calibration of the SEC chromatogram. The SEC-MALDI results provide significantly larger Mw and PD values than MALDI alone. Using SEC-MALDI, it was determined that the PD indexes of the pTHF and pBA samples are larger than the assumed values, which are based on the polyol synthesis reactions. The combination of selective degradation with SEC-MALDI, using either ethanolamine or phenylisocyanate, is a viable method for polyurethane polyol characterization. PMID- 10857626 TI - High-throughput detection of unknown mutations by using multiplexed capillary electrophoresis with poly(vinylpyrrolidone) solution. AB - Single-nucleotide polymorphism detection has been the focus of much attention recently. Although many methods have been reported, low-cost, high-throughput, and high-detection-rate methods are still in demand. We present a fast and reliable mutation detection scheme based on temperature-gradient capillary electrophoresis. A large temperature gradient (10 degrees C) was applied with a precision of 0.02 degrees C and a temperature ramp of 0.7 degrees C/min. Multiple unlabeled samples from PCR were injected and analyzed. Ethidium bromide was used as the intercalating dye for laser-induced fluorescence detection. Mutations can be recognized by comparing the electrophoretic patterns of the heteroduplex with that of a homoduplex reference without prior knowledge of the exact type of mutation present. Mutations in all five test samples were successfully detected with high confidence. This scheme is demonstrated in 96-capillary array electrophoresis for screening single-point polymorphism in large numbers of samples prior to full sequencing of only the positive samples to identify the nature of the mutation. PMID- 10857627 TI - Pressure-tunable column selectivity for high-speed vacuum-outlet GC. AB - A pressure-tunable ensemble of two series-coupled capillary columns operated at subambient outlet pressure is described. The ensemble consists of a 4.5-m length of nonpolar dimethyl polysiloxane column followed by a 7.5-m length of polar trifluoropropylmethyl polysiloxane column. Air at an inlet pressure of 1.0 atm is used as carrier gas, and a vacuum pump is used to pull the carrier gas and injected samples through the column ensemble. Detection is provided by a photoionization detector operated at a pressure of 0.3 psia. Ensemble selectivity is controlled by means of an electronic pressure controller located at the junction point between the columns. The minimum pressure step size is 0.1 psi, and 50 different set-point pressures can be used, each one producing a different pattern of peaks eluting from the column ensemble. Measured ensemble retention factors for a set of target compounds produce straight lines when plotted versus the ratio of the calculated holdup time of the first column in the ensemble to the total ensemble holdup time. A component band trajectory model is used to describe the effects of ensemble junction-point pressure on the elution patterns generated by the ensemble. Ensemble retention times predicted by the model are in good agreement with values obtained from chromatograms. The use of on-the-fly set point pressure changes during a separation (selectivity programming) is demonstrated and used to improve the quality of the separation of a 19-component test mixture. PMID- 10857628 TI - Microfabricated electrophoresis chips for simultaneous bioassays of glucose, uric acid, ascorbic acid, and acetaminophen. AB - A micromachined capillary electrophoresis chip is described for simultaneous measurements of glucose, ascorbic acid, acetaminophen, and uric acid. Fluid control is used to mix the sample and enzyme glucose oxidase (GOx). The enzymatic reaction, a catalyzed aerobic oxidation of glucose to gluconic acid and hydrogen peroxide, occurs along the separation channel. The enzymatically liberated neutral peroxide species is separated electrophoretically from the anionic uric and ascorbic acids in the separation/reaction channel. The three oxidizable species are detected at the downstream gold-coated thick-film amperometric detector at different migration times. Glucose can be detected within less than 100 s, and detection of all electroactive constituents is carried out within 4 min. Measurements of glucose in the presence of acetaminophen, a neutral compound, are accomplished by comparing the responses in the presence and absence of GOx in the running buffer. The reproducibility of the on-chip glucose measurements is improved greatly by using uric acid as an internal standard. Factors influencing the performance, including the GOx concentration, field strength, and detection potential, are optimized. Such coupling of enzymatic assays with electrophoretic separations on a microchip platform holds great promise for rapid testing of metabolites (such as glucose or lactate), as well as for the introduction of high-speed clinical microanalyzers based on multichannel chips. PMID- 10857629 TI - Automated ultra-thin-layer SDS gel electrophoresis of proteins using noncovalent fluorescent labeling. AB - Ultra-thin-layer SDS gel electrophoresis in conjunction with automated laser induced fluorescence detection is a novel and powerful method for the analysis of fluorophore-labeled proteins. The technique described in this paper employs instant, noncovalent fluorophore labeling by the addition of a fluorescent staining dye to the sample proteins either during or immediately prior to the sample loading process. Thus, the method does not require time-consuming post- or preseparation staining/labeling. By combining the multilane format of SDS polyacrylamide slab gel electrophoresis and the high separation efficiency of capillary SDS gel electrophoresis, ultra-thin-layer SDS gel electrophoresis features rapid, high-throughput, and high-resolution analysis of proteins in the molecular mass range of 14-116 kDa. The good heat dissipation inherent to the ultrathin format enables the use of agarose and agarose-based composite separation matrixes, which can be easily replaced within the separation platform. Labeling efficiency as a function of the concentration of the staining dye, SDS, and proteins is thoroughly discussed. Detection sensitivity of the method was found to be at the low-femtomole level (1.25 ng/band), determined by analyzing a set of serial dilutions of standard proteins. Practical example of molecular mass determination and characterization of a complex protein mixture are also shown. PMID- 10857630 TI - Conditions for similitude between the fluid velocity and electric field in electroosmotic flow AB - Electroosmotic flow is fluid motion driven by an electric field acting on the net fluid charge produced by charge separation at a fluid-solid interface. Under many conditions of practical interest, the resulting fluid velocity is proportional to the local electric field, and the constant of proportionality is everywhere the same. Here we show that the main conditions necessary for this similitude are a steady electric field, uniform fluid and electric properties, an electric Debye layer that is thin compared to any physical dimension, and fluid velocities on all inlet and outlet boundaries that satisfy the Helmholtz-Smoluchowski relation normally applicable to fluid-solid boundaries. Under these conditions, the velocity field can be determined directly from the Laplace equation governing the electric potential, without solving either the continuity or momentum equations. Three important consequences of these conditions are that the fluid motion is everywhere irrotational, that fluid velocities in two-dimensional channels bounded by parallel planes are independent of the channel depth, and that such flows exhibit no dependence on the Reynolds number. Similitude is demonstrated by comparing measured and computed fluid streamlines with computed electric flux lines. PMID- 10857631 TI - On-line investigation of the generation of nonaqueous intermediate radical cations by electrochemistry/mass spectrometry AB - Anodic oxidation of triphenylamine (TPA) in acetonitrile was investigated by electrochemistry (EC) combined online with mass spectrometry (MS) through a particle beam (PB) interface, EC/PB/MS. Electrooxidation of TPA generates a TPA*+ radical cation (m/z 245) which dimerizes to tetraphenylbenzidine (TPB, MW 488). TPB is readily oxidized to TPB*+ (m/z 488) and TPB2+ (m/z 244) at the oxidation potential of TPA. In EC/PB/MS, direct monitoring of the oxidation of TPB to TPB*+ radical cation as a function of the electrode potential was achieved via selective ion monitoring of the ion peak at m/z 488. By using the relative intensity ratio of ions at m/z 244 (TPB2+) to 245 (TPA*+), the formation of TPB2+ as a function of the electrode potential was also monitored. EC/PB/MS showed a maximum rate of formation of TPB*+ at +1.2 Vvs Pd, while TPB2+ is generated at a maximum rate at +1.6 V vs Pd. The effect of spectral interference from the electron impact ionization of TPA, on EC/PB/MS results, is also discussed. Finally, a significant signal enhancement is observed in the presence of tetrabutylammonium perchlorate (TBAP) and is reported for the first time. Compatibility of coupling of EC with MS via PB interface for EC/MS studies in nonaqueous solvents is demonstrated. The observation of significant signal enhancement in the presence of TBAP may facilitate other applications of LC/MS. PMID- 10857632 TI - Capillary electrochromatography of cholesterol and its ester derivatives. AB - Separation of cholesterol and its ester derivatives using micellar electrokinetic chromatography is a challenge due to the extreme hydrophobicity of these compounds. In this work, an isocratic capillary electrochromatography (CEC) method has been developed to separate a complex mixture of cholesterol and its 12 ester derivatives. The proportions of mobile phase (tetrahydrofuran, acetonitrile, water), as well as the effects of acid modifiers, buffer concentrations, voltage, and temperature on the separation of cholesterol derivatives were investigated. Addition of a polymeric surfactant, poly(sodium N undecanoyl-L-glycinate), to the mobile phase reduced migration time and improved resolution of the analytes. The CEC method developed allows baseline separation of a complex mixture of cholesterol and 12 ester derivatives in less than 40 min. Finally, the method is applied to the characterization of cholesterol, cholesterol linoleate, and cholesterol oleate extracted from atherosclerotic plaque deposits in the arterial walls of a human aorta. PMID- 10857633 TI - A dc microplasma on a chip employed as an optical emission detector for gas chromatography AB - A micromachined plasma chip is coupled to a conventional gas chromatograph to investigate its performance as an optical emission detector. The device employs a 180-nL plasma chamber in which an atmospheric pressure dc glow discharge is generated in helium. Applied power is 9 mW (770 V, 12 microA) and helium flow rate 320 nL s(-1). A number of carbon-containing compounds are detected in the column effluent by recording the emission at 519 nm. For hexane, the detector has a linear dynamic range of over two decades and a minimum detectability of 10(-12) g s(-1) (800 ppb). The detector signal shows a marked peak broadening and tailing when compared with the signal of a flame ionization detector. This is mainly attributed to dead volumes and chromatographic processes introduced by the connecting tubing and the chip glass channels. The device was operated for more than 24 h without a significant change in performance. Operation is stable and instrumental requirements are simple. Future use of the detector chip in conventional gas chromatography or as an integrated detector in on-chip gas chromatography is discussed. PMID- 10857634 TI - Capillary electrophoresis with postcolumn infectivity assay for the analysis of different serotypes of human rhinovirus (common cold virus). AB - Differentiation of virus serotypes with capillary zone electrophoresis was demonstrated. For four serotypes of human rhinovirus (HRV2, HRV14, HRV16, HRV49), different electrophoretic mobility was achieved at pH 8.3 (borate/boric acid buffer, 100 mmol/L). Addition of detergent (Triton X-100-R, deoxycholate, and/or SDS) to the background electrolyte was required for reduction of wall adsorption and improvement of peak shape. A major nonviral contaminant, present in all virus samples, was best separated from the viral peaks with 10 mmol/L SDS as additive. The method allowed detecting serotypes HRV16 and HRV49 in crude, partially purified virus preparations. An infectivity assay carried out off-line with fractions collected at the capillary outlet enabled the sensitive and biospecific identification of the peaks of HRV2 and HRV14. PMID- 10857635 TI - H-point curve isolation method for coupled liquid chromatography and UV-visible spectrophotometry. AB - The H-point curve isolation method (HPCIM) for the detection of unknown interferences in chromatography is proposed. The method allows one to estimate the UV-vis spectra of interfering species in a sample as well as to test the purity of the chromatographic peaks. Besides the detection of the unknown interferences in a sample, this method allows one to calculate the concentration of an analyte in the presence of unknown compounds. To illustrate the reliability of the proposed method, samples of diuretics and amphetamines have been analyzed by normal- and reversed-phase high-performance chromatography. PMID- 10857636 TI - Determination of amino sugars in environmental samples with high salt content by high-performance anion-exchange chromatography and pulsed amperometric detection. AB - Amino sugars were determined in natural samples, including seawater, using high performance anion-exchange chromatography with pulsed amperometric detection and a new-off-line sample cleanup procedure. Samples were hydrolyzed with 3 M HCl for 5 h (100 degrees C) and neutralized with anion retardation resin. Before injection, salts and organic contaminants were removed with a strong cation exchanger in the Na+ form. Detection limits for amino sugars were between 1 and 4 nM (signal-to-noise ratio 3), allowing for the first time quantification of amino sugars in seawater without preconcentration. Precision was 2-11% at the 20 nM level. The relatively simple and rapid sample preparation makes it suitable for routine analyses. PMID- 10857637 TI - Use of relationships between retention behaviors and chemical structures in subcritical fluid chromatography with CO2/modifier mixtures for the identification of triglycerides. AB - Satisfactory separations of vegetable oil triglycerides (TG) differing in fatty acid composition are obtained by subcritical fluid chromatography (SubFC) with octadecyl packed columns and CO2/modifier mobile phases. However, the identification of TG can be sometimes difficult due to the small retention differences between the compounds. A method of TG identification in SubFC was achieved, which does not require calculation of retention pattern but uses the differences in retention behavior related to TG structure and to the nature of the subcritical mobile phases. These retention differences were produced by the variation of either outlet pressure or modifier percentage or of temperature. Whatever the column aging, this method allows the determination of the triglyceride total chain length and double bond number. Among numerous structures, these two criteria restrict the structural hypothesis at worst to three or four possibilities and sometimes only to one. The validity of this relative identification method was confirmed by electronic impact mass spectrometry of triglyceride fractions collected from the analysis of a peanut oil. The analysis of concentrated fractions is favored by the spontaneous elimination after the pressure regulator of carbon dioxide, the main fluid of the subcritical mobile phase. PMID- 10857638 TI - Experimental characterization of end-column electrochemical detection in conjunction with nonaqueous capillary electrophoresis AB - An end-column electrochemical detector arrangement for capillary electrophoresis (CE) based on a 75-microm-i.d. capillary and a 25-microm microdisk electrode is characterized. The investigations were carried out using a nonaqueous (acetonitrile-based) buffer and ferrocene model compounds which offer high reliability for voltammetric measurements. The positioning of the microdisk electrode relative to the capillary outlet is the most important parameter for optimization of detection performance as it determines the characteristics of mass transport toward the electrode and the effect of ohmic potential drop resulting from the electrophoretic current on the actual detection potential. On the basis of spatially resolved studies, it was concluded that for the detection system used the microdisk electrode should be placed in a central position relative to the capillary outlet at a distance within the range of 75-100 microm. The presence of a high-voltage electric field had no negative effect on baseline noise, which was demonstrated by comparison of capillary flow injection based on gravity flow and CE experiments. Even a faster stabilization of the baseline was observed by increasing the separation voltage. PMID- 10857639 TI - Creating addressable aqueous microcompartments above solid supported phospholipid bilayers using lithographically patterned poly(dimethylsiloxane) molds AB - Herein we report the use of microcontact displacement (microCD) to generate addressable arrays of aqueous solutions above fluid lipid bilayers by bringing a patterned PDMS mold into conformal contact with a phospholipid membrane on a solid supported substrate. Epifluorescence microscopy established that the bilayer material was displaced in regions where contact between the mold and substrate was made. Photobleaching experiments confirmed that the bilayer sectors remained individually fluid but completely separated. The microcompartments created by microCD could be individually injected with aqueous solutions that remained sealed from their neighbors. This procedure was then exploited for screening a small library of aqueous-phase molecules for their ability to inhibit binding between surface-bound ligands and soluble protein receptors. PMID- 10857640 TI - Development of binding assays in microfabricated picoliter vials: an assay for biotin. AB - A homogeneous binding assay for the detection of biotin in picoliter vials was developed using the photoprotein aequorin as the label. The binding assay was based on the competition of free biotin with biotinylated aequorin (AEQ-biotin) for avidin. A sequential protocol was used, and modification of the assay to reduce the number of steps was examined. Results showed that detection limits on the order of 10(-14) mol of biotin were possible. Reducing the number of steps provided similar detection limits but only if the amount of avidin used was decreased. These binding assays based on picoliter volumes have potential applications in a variety of fields, including microanalysis and single-cell analysis, where the amount of sample is limited. In addition, these assays are suitable for the high-throughput screening of biopharmaceuticals. PMID- 10857641 TI - Quantification of lanthanides in rocks using succinic acid-derivatized sorbents for on-line SPE-RP-ion-pair HPLC AB - The use of new poly(norbornene-block-7-oxanorborn-2ene-5,6-dicarboxylic acid) coated silica-based sorbents as well as of beaded polymers based on poly(norborn 2-ene-5,6-dicarboxylic acid-co-1,4,4a,5,8,8a-hexahydro-1,4,5,8-exo-endo dimethanonaphthalene) for the on-line preconcentration of lanthanides from rock digests and their subsequent RP-ion-pair HPLC separation is described. Block co polymers of norborn-2-ene and 7-oxanorborn-2-ene-5,6-dicarboxylate used for coating were prepared by ring-opening metathesis polymerization (ROMP) and poly(norborn-2-ene-5,6-dicarboxylic acid-co-1,4,4a,5,8,-8a-hexahydro-1,4,5,8-exo endo-dimethanonaphthalene)-based polymer beads were prepared by ring-opening metathesis precipitation polymerization. Both types of sorbents exhibit an extraordinarily good pH stability, are hydrophilic and therefore easily wetable by water alone, and show high extraction efficiencies for lanthanides within a pH of 3.5-5.5. The rare earth element (REE) content in the investigated rocks varied over 3 orders of magnitude (0.19-108 microg/g). REE concentrations prior to enrichment were typically in the range of 1-25 ng/mL; the total amount of each REE sorbed onto the precolumn was in the range of 8-270 ng. Extraction selectivities of the sorbent may be enhanced by adding 5-sulfosalicylic acid as a masking agent for iron and aluminum as well as methanol as an inhibitor for the precipitation of o-silicic acid. Gradient elution of the lanthanides from the precolumn and their subsequent separation on a RP-C18 column was achieved using hydroxyisobutyric acid (HIBA) and sodium octanesulfonate. Depending on the actual concentration of the lanthanides in the digests and in order to suppress interfering cations, UV detection was carried out with two different postderivatization reagents, 4-(2-pyridylazo)resorcinol (PAR) and arsenazo III. The high selectivity in enrichment as well as the complementary use of post derivatization reagents allows the fast, quantitative, and highly reproducible quantification of REEs present in rocks by complete removal or suppression of all other interfering components. Thus, recoveries were found to be within a range of 97-103% for most REEs with relative standard deviations of 2-5%. PMID- 10857642 TI - Low-energy ion--surface reactions of pyrazine with two classes of self-assembled monolayers: influence of alkyl chain orientation AB - Collisions of pyrazine with two classes of self-assembled monolayer (SAM) films are employed to determine whether surface confinement and the resulting alkyl chain orientation, influences low-energy ion-surface reactions. SAM films formed from n-alkanethiols (CH3(CH2)n-S-Au, n = 14-17) and 4-(4 alkoxyphenylbenzenethiols (4-(4-CH3(CH2)mOC6H4)-C6H4-S-Au, m = 14-17) chemisorbed onto Au (111) substrates are known to exhibit a chain-length-dependent odd-even effect that places the terminal C-C bond into different orientations. Ion-surface collisions (20 eV) of pyrazine molecular ion (M = m/z 80) with these surfaces yield reaction product ions corresponding to the addition of hydrogen atoms ([M + H]+ = m/z 81) and methyl groups ([M + CH3]+ = m/z 95) from the surface to the probe ion. Differences in the relative abundance of the reaction product ions are measured as a function of chain length for both classes of SAM film. SAM films with odd chain lengths (n, m = 14 and 16) have a consistently higher abundance of H addition product ions than SAM films with even chain lengths (n, m = 15 and 17). Alternating reactivity is also observed for the addition of CH3, with methyl addition occurring more readily on even-chain-length films. The variations are consistent with the well-characterized orientation differences known to exist for films of this type. Specifically, odd-chain-length films are oriented such that the last C-C bond is more parallel to the plane of the surface than it is for even-chain-length films. The critical element of the parallel orientation is that it leaves, on average, one hydrogen atom on the terminal methyl and both hydrogen atoms on the first underlying methylene in more reactive positions compared to even chain lengths. Conversely, the trend in the relative abundance of CH3 addition indicates that the orientation produced by an even-chain-length film, with the last C-C bond more perpendicular to the surface, allows the probe ion better access to the methyl carbon. Reflection absorption IR spectroscopy (RAIRS) data independently confirm the orientational disposition of the films. The RAIRS data show that the odd-even effect is less dramatic for the n-alkanethiols when compared to 4-(4-alkoxyphenyl)benzenethiols. A smaller difference in ion-surface reactivity is measured for n-alkanethiols, demonstrating that ion-surface reactions can distinguish subtle differences in average orientation. In short, we report that the extent of ion-surface reactions of pyrazine ion with two classes of SAM films is directed by the spatial orientation of the surface-confined species that participate in the reaction. PMID- 10857643 TI - X-ray Raman spectroscopy of carbon in asphaltene: light element characterization with bulk sensitivity. AB - X-ray Raman spectra of the carbon K-edge have been recorded using 6.461 keV radiation for a petroleum asphaltene. By comparison with coronene, graphite, and paraffin standards, the asphaltene spectrum is seen to be composed of contributions from saturated and aromatic carbon species. The information contained in the carbon K-edge was extracted with bulk (approximately 1 mm) sensitivity, because the Raman method used hard X-rays. This helps alleviate concerns about surface artifacts that frequently occur with soft X-ray spectroscopy of light elements. X-ray Raman spectroscopy shows great potential for characterization of light elements in fuels, catalysts, and other complex materials under chemically relevant conditions. PMID- 10857644 TI - Trace chemical characterization using monochromatic X-ray undulator radiation AB - An efficient Johansson-type X-ray fluorescence spectrometer has been developed for advanced X-ray spectroscopic analysis with third-generation synchrotron radiation. Kalpha and Kbeta X-ray fluorescence spectra for trace metals have been collected by a Ge(220) analyzing crystal with a Rowland radius of 150 mm, under monochromatic X-ray excitation at the undulator beamline at the SPring-8. The energy resolution is approximately 10 eV for most of the K lines for 3d transition metals. In light of the greatly improved efficiency, as well as the excellent signal-to-background ratio, the relative and absolute detection limits achieved are 1 ppm and 1.2 ng of copper in a carbon matrix, respectively. The energy resolution of the present spectrometer permits the observation of some chemical effects in Kbeta spectra. It has been demonstrated that the changes in Kbeta5 and Kbeta'' intensity for iron and cobalt compounds can be used for the analysis of chemical states. Resonant X-ray fluorescent spectra are another important application of monochromatic excitation. In view of trace chemical characterization, the present spectrometer can be a good alternative to a conventional Si(Li) detector system when combined with highly brilliant X-rays. PMID- 10857645 TI - SIMS of organic anions adsorbed onto an aminoethanethiol self-assembled monolayer: an approach for enhanced secondary ion emission AB - Secondary ion mass spectrometry (SIMS) was used to monitor the uptake of organic anions from solution by aminoethanethiol (AET) monolayers on Au substrates, as a test of the applicability of this monolayer as a substrate for organic SIMS analysis. Event-by-event bombardment and detection mode coupled with coincidence counting allowed the atomic and polyatomic projectile impacts on a particular sample surface to be compared simultaneously and under the same experimental conditions. The mass spectra produced from the monolayer surface and those from Au and Si blanks demonstrate that the AET monolayer is important to the uptake of the organic anion. The exchanged monolayer surfaces were used to measure secondary ion yields, defined as the number of secondary ions detected per incident primary ion, produced from ultrathin films by (CsI)nCs+ (n = 0-2) projectiles at the limit of single-ion impacts. The yield of a tetradecyl sulfate (IDS) anion was improved by a factor of 200 using the AET substrate instead of the thick salt target. The intact ion and fragment ion yield trends produced from the AET surface were measured as a function of number of atoms in the primary projectile and energy. We observed a yield increase for both the intact ion and the fragment ion with the projectile complexity and energy. The increase in yield per projectile atom was linear for the emission of intact TDS and intact dodecyl sulfate from the AET surfaces. A supralinear yield enhancement, however, was observed for the fragment ion SO3- when the three-atom (CsI)Cs+ cluster was used. The experiments demonstrate that the various organosulfate and suffonates are weakly bound to the AET surface and their adsorption to the AET monolayer is reversible. The utility of the AET monolayer on Au was also tested as a general substrate for the characterization of derivatized organic molecules with biological and industrial importance by TOF-SIMS. PMID- 10857646 TI - Mapping interfacial chemistry induced variations in protein adsorption with scanning force microscopy. AB - In this work, we demonstrate the sensitivity of scanning force microscopy (SFM), operated in friction force mode, to adsorbed protein conformation or orientation. We employ patterned films of methyl- and carboxylate-terminated alkanethiolate monolayers on gold as substrates for protein adsorption to observe the effect of each functional group in the same image. Infrared spectroscopic and SFM studies of bovine fibrinogen (BFG) adsorption to single-component monolayers indicate that complete films of BFG that are stable to imaging are formed at each functional group. After adsorption of BFG to a patterned monolayer, we observe a contrast in friction images due to differences in adsorbed BFG conformation or orientation induced by each functional group. We also observe frictional contrast in films of other proteins adsorbed on patterned monolayers. These observations lead to the conclusion that SFM-measured friction is sensitive to adsorbed protein state. PMID- 10857647 TI - Characterization of the "helix clamp" motif of HIV-1 reverse transcriptase using MALDI-TOF MS and surface plasmon resonance. AB - A helix-turn-helix motif in the crystal structure of human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) was proposed to be a conserved nucleic acid binding domain among several nucleotide polymerizing enzymes (Hermann, T.; Meier, T.; Gotte, M.; Heumann, H. Nucleic Acids Res. 1994, 22, 4625-4633). The sequence of this domain is homologous to 259KLVGKL-(X)16KLLR284 of HIV-1 RT, which acts as a "helix clamp" grasping the template-primer (T-P) complex. We characterized the helix clamp motif using MALDI-TOF MS and surface plasmon resonance (BIAcore). Our studies showed that the "helix clamp" has a nucleic acid binding function that may not be sequence specific. This evidence suggests that ionic interactions between the helix clamp and oligonucleotide backbone are not solely responsible for binding. Secondary and tertiary structures of the protein may also play a significant role in nucleic acid binding. The association and dissociation constants, ka and kd, for the binding of single-stranded oligonucleotide to the helix clamp were determined to be 7.03 x 10(3) M(-1) s(-1) and 1.22 x 10(3) s(-1), respectively. PMID- 10857648 TI - Electrochemically modulated liquid chromatography coupled on-line with electrospray mass spectrometry. AB - Electrochemically modulated liquid chromatography (EMLC) has been coupled to an electrospray mass spectrometer. This combination takes advantage of the ability of EMLC to manipulate retention and enhance separation efficiency solely through changes in the potential applied to a conductive stationary phase, thereby minimizing complications because of possible changes in analyte ionization efficiencies when gradient elution techniques are used. Three examples are presented that demonstrate the attributes of this EMLC/electrospray mass spectrometry (ES-MS) coupling. The first two examples involve the separation of mixtures of corticosteroids or of benzodiazepines, showing the general utility of the union for eluent identification and low-level detection. The ability to identify products from on-column redox transformations is also demonstrated using the benzodiazepine mixture. The third example investigates the electrooxidation of aniline by utilizing an EMLC column as an on-line electrochemical reactor and product separator and ES-MS for detection and product identification. PMID- 10857649 TI - Supersonic jet/multiphoton ionization/mass spectrometry of dioxins formed by the thermal reaction of phenols in the absence and presence of an FeCl3 catalyst. AB - Dioxins, which are thermally produced from several precursor molecules, were investigated by supersonic jet/multiphoton ionization/time-of-flight mass spectrometry (SSJ/MPI/TOF-MS). Dibenzofuran and dibenzo-p-dioxin were efficiently generated from o-chlorophenol and also from phenol after a chlorination reaction with FeCl3. The present technique was employed for the continuous monitoring of a specified isomer, e.g., m-chlorophenol, which is formed at relatively low temperatures by chlorination of phenol with FeCl3. A dimerization reaction that forms a dibenzo-p-dioxin, e.g., dichlorodibenzo-p-dioxin from 2,4-dichlorophenol, at relatively high temperatures was also investigated. The number of chlorine atoms in the dioxin molecule was largely correlated with the number of chlorine atoms in the precursor molecule. However, some unexpected compounds, which probably occur by dechlorination and rearrangement reactions, were also found. Thus, the SSJ/MPI/TOF-MS technique represents a sensitive, as well as selective, analytical method for monitoring thermally generated dioxins. PMID- 10857650 TI - A screening method for antigen-specific IgE using mast cells based on intracellular calcium signaling. AB - A simple screening method is presented for the measurement of antigen-specific IgEs in sera in which mast cells are used. This method is based on the intracellular calcium signal in mast cells induced by cross-linking the surface high-affinity Fc receptors (FcepsilonRIs) with IgEs and multivalent antigens. When a serum containing various antigen-specific IgEs is added to the mast cell suspension, various antigen-specific IgEs are captured by FcepsilonRIs on the cell surface. However, the required antigen-specific IgE can be specifically detected after the addition of the corresponding antigen. The resulting increase in intracellular calcium concentration ([Ca2+]i), monitored by Ca2+-fluorometry, was found to be an analytical measure for the screening of IgEs. Two kinds of rodent mast cells, cell-lined RBL2H3 cells and primary cultured BMMCs, were used as a representative model system of mast cells. A DNP hapten (DNP35-HSA) and ovalbumin (OVA) were chosen for illustrative antigens, and these antigen-specific IgEs (DNP-specific IgE, OVA-specific IgE) in the corresponding rodent sera were target antibodies. It was found that [Ca2+]i increased linearly with IgE concentrations ranging from 25 to 5000 ng/mL for DNP-specific IgE and from 5 to 50 ng/mL for OVA-specific IgE. For these dynamic ranges, optimum concentrations of antigens were found to be 10 ng/mL and 1 microg/mL for DNP35-HSA and OVA, respectively. It was concluded that by monitoring the increase of [Ca2+]i in mast cells, we could determine the antigen-specific IgEs. The present immunological assay based on the Ca2+ signal transduction in mast cells offers new possibilities for efficient screening of antigen-specific IgEs and the immunogenicity of IgE in sera. PMID- 10857651 TI - Lab-on-a-Cable for electrochemical monitoring of phenolic contaminants. AB - The "Lab-on-a-Cable" concept, based on scaling down an electrochemical flow system to a cable platform, is described. The system integrates the analyte collection and the sample handling, with the electrochemical detection of the reaction product in a sealed cylindrical unit, connected to a long shielded cable. An enzymatic assay, involving collection of a phenolic substrate, its mixing with an internally delivered tyrosinase solution, and amperometric detection of the liberated quinone product, is used for illustrating the operation of the flow probe and demonstrating its advantages over remote phenol sensors. The internal buffer solution ensures independence of sample conditions such as pH, ionic strength, or natural conditions, that commonly influences the performance of remote sensors. The "built-in" flow pulsation of the integrated micropump is exploited for a sensitive hydrodynamic-modulation voltammetric detection of hydrazine and peroxide pollutants. PMID- 10857652 TI - On-line D/H analysis for water, natural gas, and organic solvents by manganese reduction AB - A new technique for on-line sample preparation and D/H determination is described. The technique is suitable for the preparation of fresh and brine waters, as well as natural gases and organic solvents. A 5-microL sample of water or hydrogen equivalent is injected and reduced by means of hot manganese metal in a specially designed reaction tube surrounded by a tube furnace and attached directly to the mass spectrometer inlet without modification.The hydrogen gas flows directly into the MS to be analyzed by reference/sample comparison. The reproducibility varied between 0.7 and 1.8% for all liquid and gas samples. The accuracy of this technique is confirmed by analysis of IAEA standard waters V SMOW, GISP, and SLAP, as well as NGS-3 (IAEA methane intercomparison material). PMID- 10857653 TI - Thermal denaturation: a useful technique in peptide mass mapping. AB - The use of thermal denaturation of proteins prior to in-solution digestion and mass spectral peptide mass mapping is reported. Thermal denaturation is preferred over chemical denaturation because it does not require purification/concentration prior to mass spectral analysis. Enzymatic digestions of proteins that are resistant to proteolysis are significantly enhanced by thermal denaturation. Native proteins that are sensitive to proteolysis show similar or slightly lower digestion yields following thermal denaturation. Proteins that are resistant to digestion become more susceptible to digestion, independent of protein size, following thermal denaturation. For example, amino acid sequence coverage from digest fragments increases from 15 to 86% in myoglobin and from 0 to 43% in ovalbumin. This leads to more rapid and reliable protein identification by MALDI peptide mass mapping. Although some proteins aggregate upon thermal denaturation, the protein aggregates are easily digested by trypsin and generate sufficient numbers of digest fragments for protein identification. PMID- 10857654 TI - Thin-film glucose biosensor based on plasma-polymerized film: simple design for mass production. AB - We propose a simple thin-film glucose biosensor based on a plasma-polymerized film. The film is deposited directly onto the substrate under dry conditions. The resulting films are extreme thin, adhere well onto the substrate (electrode), and have a highly cross-linked network structure and functional groups, such as amino groups, which enable a large amount of enzyme to be immobilized. Since this design allows fabrication through a dry process, with the exception of the enzyme immobilization, which is the last stage of the process, the chip fabrication can be designed as a full-wafer process to achieve mass production compatibility. The resulting sensors produced using this film are more reproducible, exhibit lower noise, and reduce the effect of interference to a greater degree than sensors made using conventional immobilization methods, e.g., via 3 (aminopropyl)triethoxysilane. The obtained film is a good interfacial design between enzyme and electrode; enzyme two-dimensionally locates very close to the electrode in a manner that is quite reproducible. Therefore, a wide dynamic range (up to 60 mM) and rapid response time (11.5+/-0.8 s) were obtained. Because of its highly cross-linking network structure, the amperometric response due to interferences such as ascorbic acid and acetaminophen was reduced by size discrimination of plasma-polymerized films. PMID- 10857655 TI - Attention deficits: is there a right hemisphere specialization for simple reaction time, sustained attention, and phasic alertness? AB - Impairments of attentional focus often are claimed to be associated with lesions of the right hemisphere. Although some studies comparing right- and left-brain damaged patients have supported this idea, others have not found differences between these patients in various attention tasks. The present study was carried out in order to further investigate the putative role of the right hemisphere in a simple reaction time task, a sustained attention task, and a phasic alertness task. Subjects were 46 patients with right-hemisphere lesions and 37 patients with left-hemisphere lesions. Results showed no difference between right- and left-brain-damaged patients in simple reaction time, in speed of response over time (sustained attention), and in the capacity to prepare to an uncoming stimulus (phasic alertness). Future studies will have to use a more precise categorization of lesions than only right-versus left-brain damage. PMID- 10857656 TI - Development and decay of extra-linguistic communication. AB - Cognitive Pragmatics theory is concerned with analyzing the cognitive processes underlying communication. In previous works we have explained the emergence of communication in context, as revealed by very young children, and the communicative deficits shown by closed head injury patients. The aim of the present work is an extension of Cognitive Pragmatics to the emergence and the decay of extra-linguistic communication. In particular, we investigate the performance of 2- to 7-year-old children and that of Alzheimer's patients in standard and nonstandard (irony and deceit) pragmatic tasks. The predictions derived by Cognitive Pragmatics are confirmed. Comprehension of pragmatic phenomena which are more complex according to the theory emerges later in the development (Experiment 1), and their decay is most severe in Alzheimer's patients (Experiment 2). We conclude that the framework provided by Cognitive Pragmatics can accommodate both the development and the decay of extra-linguistic communication. PMID- 10857657 TI - Spatial learning on the Morris Water Maze Test after a short-term paradoxical sleep deprivation in the rat. AB - Twelve rats were deprived of paradoxical sleep (PS) for eight hours using the small platform method. PS-deprived and control rats then learned either the standard allocentric version (using external cues) of the Morris Water Maze (MWM) or a delayed alternation version (changing the platform location between trials: MWM(DA)). Overall, rats learning the MWM(DA) made more quadrant entries than rats learning the allocentric version. Compared to other rats, PS-deprived rats crossed more quadrants only in the MWM(DA). These results show that MWM(DA) is a more complex task to learn and is more vulnerable to PS deprivation than allocentric spatial orientation. Since delayed alternation is dependent upon frontal structures, we propose that tasks involving the frontal cortex are more sensitive to short-term PS deprivation than tasks related to hippocampal structures. PMID- 10857658 TI - Procedural learning in schizophrenia: further consideration on the deleterious effect of neuroleptics. AB - Deficits in procedural learning remain a controversial issue in schizophrenia. This may be related to the nature of the neuroleptic treatment of schizophrenic patients as conventional neuroleptics may be more deleterious than new atypical neuroleptics. However, there is no comparative study on the effect of specific neuroleptics on procedural learning. In this study, three groups of patients treated with different neuroleptics were compared to normal controls on two procedural learning tasks. In a visuo-motor task, patients and controls showed similar learning rates, although schizophrenic patients showed generally lower performances than normal controls. However, patients treated with a conventional neuroleptic, but not those treated with the atypical neuroleptics, showed many fluctuations during the initial learning phase. In a problem-solving task, all groups were comparable in performance and learning fluctuations, but learning rates were lower in patients treated with the neuroleptic showing the higher incidence of extrapyramidal symptoms. This suggests that procedural learning abilities may be significantly affected by neuroleptics in schizophrenia, although the effect may differ between tasks and the specific neuroleptics. Fluctuations in the initial learning phase of the visuo-motor task probably results from a frontal dysfunction while reduced learning rates, such as those observed in the problem solving task, may be attributed to a striatal dysfunction. This is concordant with the differential pharmacological actions of the conventional and atypical neuroleptics in these two cerebral areas. PMID- 10857659 TI - Neuromotor profiles: what are they and what can we learn from them? AB - Eye movements, alternating movements, rapid pointing movements, and various tremors were measured on patients with Parkinson's disease (n = 21), on Cree subjects exposed to methylmercury (n = 36), and on healthy control subjects (n = 30). Neuromotor profiles were created according to thirty characteristics extracted from test results of four subgroups matched for age and composed of six subjects each. Z scores were calculated with respect to the mean and standard deviation of the control group for each of the 30 characteristics. The subgroup with the lower methylmercury blood level had larger z scores than the control subgroup and with a few positive values above one standard deviation. The subgroup with the higher methylmercury blood level had several z scores above two standard deviations. Interestingly, the abnormal values for the subgroup with Parkinson's disease were mostly limited to static tremor recorded with no visual feedback and reached up to 5 standard deviations. These results indicate that neuromotor profiles can be used to summarize information extracted from different neuromotor tests and to differentiate neurological conditions. PMID- 10857660 TI - Intelligence profiles in children and adolescents with left temporal lobe epilepsy: relationship to language laterality. AB - Pre- and postoperative IQ profiles were examined in children and adolescents with left temporal-lobe epilepsy. Participants were grouped according to their pattern of speech dominance, as indicated by the intracarotid sodium amobarbital procedure. Seven participants with typical (left hemisphere) speech representation were matched for age and sex to seven participants with atypical (bilateral or right hemisphere) speech representation. The group with atypical speech representation tended to perform worse pre- and postoperatively on several verbal and nonverbal subtests. Unlike Strauss et al.'s (1990) sample, however, the groups did not differ significantly on pre- or postoperative Performance IQ. The results suggest that when individuals are matched closely for age and sex, there may only be limited support for a nonverbal "crowding" hypothesis. Atypical speech representation in children and adolescents with early temporal-lobe epilepsy appears to be associated with lowered performance on both verbal and nonverbal IQ subtests. PMID- 10857661 TI - Neuropsychological evaluation of neurologically asymptomatic HIV-infected children. AB - Forty-two children born to HIV positive mothers (29 infected at different stages of the disease, according to the Disease Control Classification Centers, and 13 noninfected) underwent evaluation using a battery of neuropsychological tests. Executive function impairments were present in all infected children, whereas memory and visuo-prassic deficits were evident only in those with full-blown AIDS. Language abilities and overall intelligence were spared. Performance of seroreverters was in the normal range. These findings suggest that even in neurologically asymptomatic children, neuropsychological evaluation can identify early impairment of specific cognitive functions. The findings are discussed in the light of the prognostic power of neuropsychological assessment for early signs of HIV neurological involvement. PMID- 10857662 TI - A componential analysis of visual object recognition deficits in patients with herpes simplex virus encephalitis. AB - Five patients with a diagnosis of Herpes Simplex Virus Encephalitis (HSVE) underwent neuropsychological assessment to explore the integrity of their visual perceptual abilities. Selective deficits affecting different levels of the recognition processing were found; impaired recognition abilities were also influenced by selective task requirements, which resulted either in facilitatory or constraining effects on patients' performance. A theoretical model of object recognition (Humphreys & Riddoch, 1987) was taken into account to explain patients' performance. Further, the role of specific components of visual processing was evidenced in explaining the performance of patients affected by HSVE. PMID- 10857663 TI - Two functional magnetic resonance imaging f(MRI) tasks that may replace the gold standard, Wada testing, for language lateralization while giving additional localization information. AB - Two tasks were used to lateralize and localize language functions noninvasively, using functional magnetic resonance (fMRI BOLD sequences). fMRI images produced during comprehension of the gist of a tale derived from an ordered series of inferential questions were used to lateralize and locate the center of and margins within language dominant hemisphere near posterior temporal-parietal occipital (TPO) cortical area(s), e.g., Wernicke's. A silent noun-generating task was used to lateralize and localize naming functions within and along superior temporal gyrus (STG) and/or the basal temporal language area (BTLA) in fusiform gyrus. Used within a series of tasks, their purpose was to investigate the reliability and validity of replacing the invasive gold standard for language lateralization, Wada test, with a noninvasive test, BOLD fMRI. PMID- 10857664 TI - A developmental analysis of the relationship between hand preference and performance: II. A performance-based method of measuring hand preference in children. AB - The present study describes a performance-based method of measuring hand preference in children. Three aspects of handedness were considered to be important in developing the paradigm (a) overall hand preference across a number of tasks, (b) consistency of hand use and, (c) the use of the preferred hand in a bimanual task. The new paradigm, termed the WatHand Box Test (WBT), requires participants to perform a variety of unimanual tasks such as, using a hammer, tossing a ball, and opening a lock with a key. To determine the validity of the WBT and examine the developmental trends in hand preference, eighty right-handed children and adults (ages 3-4, 6-7, 9-10, and 18-24 years) performed the WBT. First, the WBT was found to correlate significantly with scores on a standard hand preference questionnaire for the adults. As well, significant developmental trends were noted in hand preference as measured by the WBT. Most specifically, three- and four-year-olds had significantly lower scores on the WBT indicating a less stable pattern of hand preference than in the other three age groups. PMID- 10857665 TI - Phonological and semantic information in word and nonword reading in a deep dyslexic patient. AB - Deep dyslexia is diagnosed when brain-injured, previously literate adults make reading errors that include hallmark semantic paralexias (e.g., reading HEART as BLOOD) and are also impaired at reading nonwords (e.g., FRIP). The diversity of these symptoms have led most researchers to conclude that there are multiple sources of impairment in this syndrome and that one of the most critical is a failure to process phonological information at a sublexical level. The patient (SD) reported in this study fits the deep dyslexia profile to the extent that she makes several semantically related reading errors. She also shows the classic frequency and image ability effects of the syndrome. However, as we report, she does read some nonwords correctly and she shows a strong advantage for naming when phonemic cues are presented. We discuss the performance of SD, on these preliminary tasks, in terms of a phonological selection impairment. PMID- 10857666 TI - Age-related differences in semantic priming: evidence from event-related brain potentials. AB - Hasher and Zacks (1988) suggested that aging may affect processes involved in the inhibition of irrelevant information during language processing. Our experiment tested this hypothesis using the N400 event-related brain potential in a priming paradigm and assessed whether older subjects could benefit from the constraints of a sentence context. Twenty older (63 to 88 years) and 20 young (19 to 29 years) subjects read sentences and word pairs. Each final word varied on the degree of relatedness to the preceding context, with some being highly related (BC), moderately related (R), or unrelated (U). Younger subjects showed the expected N400 effect gradient (U > R > BC) in both the sentence and word-pair contexts, while older adults showed no discrimination between the conditions (U = R = BC) for the sentence and limited discrimination (U > BC) for the word pairs. These results support the inhibition-deficit hypothesis, whereby older adults fail to inhibit related items in working memory, and suggest that older adults do not benefit from the constraints of a sentence context. PMID- 10857667 TI - Interaction between lateralized systems: exploring the complexity of laterality. AB - To assess the extent of interaction between lateral biases in response systems at different levels of the neuraxis and detect the possible presence of different patterns of interaction related to population subgroups, we investigated laterality in hand reaching, whole-body turning and eye use in 20 bushbabies (Otolemur garnettii). Two subgroups were clearly identified: the STABLE group was composed of subjects, mainly females, that were consistent in hand preference and had correlation of hand/eye bias; the UNSTABLE group included subjects, mainly males, that showed instability in hand preference as a function of change in test conditions and had correlation of hand/turning bias. Results are interpreted to support the value of the study of interaction between lateral biases as a way of gaining a deeper understanding of the complexity of laterality. PMID- 10857668 TI - Foreign Accent Syndrome following a catastrophic second injury: MRI correlates, linguistic and voice pattern analyses. AB - A case study of Foreign Accent Syndrome (FAS) is presented with discussion of anatomical localization of injury and comparisons of pre- and postinjury linguistic, phonetic, and acoustic speech characteristics. Because the patient's injury and symptoms were unrelated to previously injured left frontal cortex, and in light of another case history (Moonis et al., 1996), we suggest that FAS has a primary subcortical involvement. We also show that this case is accompanied by a deficit in linguistic, but not affective, prosodic expression. We agree that the "foreign" quality of the FAS speech is a perceptual impression of the listener and not inherent in the patient's vocalization. Finally, we suggest a battery of tests for future FAS cases to further our study and understanding of the syndrome. PMID- 10857669 TI - Impulsivity and control of inhibition in Benign Focal Childhood Epilepsy (BFCE). AB - Childhood epilepsy represents abnormal brain functioning and may affect cognitive functions that depend on the late development of the frontal lobes. This study addresses the possible consequences of benign epilepsy on frontal functions, specifically action regulation and inhibition in the absence of explicit frontal neurological signs. Thirteen children (8 males; 6-12 years old) with benign epilepsy were matched to 13 controls. They performed tasks designed to measure impulsivity and control of inhibition: CPT, Stop Signal Paradigm, Stroop test, and Matching Familiar Figures Test (MFFT). Children with epilepsy made more errors on the MFFT (p < .02), made more errors in the interference condition on the Stroop test (p < .01), and had a longer response time to the Stop Signal (p < .05) than controls, with no differences on nonexecutive functions measures. Thus, children with benign epilepsy have a deficit on some measures of impulsivity and inhibition, which may reflect poor frontal lobe functioning. PMID- 10857670 TI - Imageability and word recognition in the left and right visual fields: a signal detection analysis. AB - Identifying the information processing constraints that determine whether or not imagery moderates visual field asymmetries is essential for constructing a dynamic model of hemispheric interaction during language processing. In this investigation, we manipulated the global experimental context in which imageable and nonimageable words were presented by contrasting mixed and blocked word lists using a lateralized lexical decision task. Signal detection analyses were employed to assess whether global stimulus context and imageability differentially affect word discriminability (d prime) and response bias (log beta) across visual fields. Both discriminability and response bias varied with imageability and stimulus context, but to a comparable extent across visual fields. This suggests that both hemispheres are sensitive to the global context in which words are presented, and can adjust processing based not only on semantic characteristics of the words themselves, but also on the variability of items in the stimulus environment. PMID- 10857671 TI - Distributional typicality: a new approach to estimating noun and verb usage from large scale text corpora. AB - This paper reports a new approach to estimating the extent to which words have predominant noun and verb usages which do not require human judgments about parts of speech. The Hyperspace Analog to Language model (HAL, Lund & Burgess, 1996) was used to computationally estimate noun vs verb usage based on the statistical regularities present in a large-scale electronic text corpus. This measure can be used to estimate the extent to which a given word occurs in typical noun or verb sentence contexts (i.e., its distributional typicality) in informal contemporary discourse. PMID- 10857672 TI - Correlation between the position of the egocentric reference and right neglect signs in left-brain-damaged patients. AB - Fourteen control subjects and thirty left-brain-damaged (LBD) patients with or without neglect performed a straight-ahead pointing task with their left hand while blindfolded. Results showed a significant correlation between the position of the egocentric reference and the presence of right neglect signs. We discuss here the egocentric hypotheses of spatial bias. PMID- 10857673 TI - Age effects in a computational model of memory. AB - Previous research demonstrates that semantic priming is relatively unaffected by age (Chiarello, Church, & Hoyer, 1985; Howard, 1988). To determine how age might affect representations in the HAL model of memory, the authors gathered text from older and younger adults and generated a global co-occurrence matrix for each. An analysis demonstrated that, as in humans, there was little difference in measures of semantic priming between memory matrices constructed from the two corpora. However, the authors discovered that semantic word neighborhoods generated by the older adult corpus were denser than corresponding neighborhoods derived from the younger adult corpus. This result has implications for changes in the semantic representation of words due to the aging process. PMID- 10857674 TI - Flexibility, inhibition, and planning: frontal dysfunctioning in schizophrenia. AB - Neuropsychological tests known to reveal abnormalities in patients with frontal lobe damage were used to explore cognitive function in 20 chronic schizophrenic patients. Eleven control subjects, matched on age and NLV-IQ (NLV is the Dutch version of the NART) were also tested. No impairments of planning ability were found on either the Action Program test or the Zoomap test, both subtests from the BADS (Behavioural Assessment of the Dysexecutive Syndrome). No abnormalities were apparent on tests of reactive flexibility, measured by task-switching and by the Rule Shift Cards test, also a subtest of the BADS. Patients with schizophrenia, however, had significantly greater difficulty in inhibiting irrelevant information and in generating words in a verbal fluency task, a measure of spontaneous flexibility. PMID- 10857675 TI - An evolutionary perspective on hemispheric asymmetries. AB - The evolution of hemispheric specialization of function has obvious benefits in terms of overall processing capacity, but it may also have associated costs to each hemisphere in isolation. These costs are not apparent in an intact brain since information can be readily transferred between the hemispheres via the cortical commissures. We suggest that the evolution of language in the left hemisphere may have occurred at the expense of some visuospatial functions. Because the right hemisphere was still capable of performing those functions, the relative left-hemisphere impairment for visuospatial processing would be invisible. If the cortical commissures are severed, as in callosotomy patients, the costs of specialization may become more apparent. We report data supporting the idea that the left hemisphere may have lost visuospatial abilities that it once possessed, and suggest that this process may also result in right hemisphere deficits in abilities related to linguistic processing. PMID- 10857676 TI - Passive and active processes in visuo-spatial memory: double dissociation in developmental learning disabilities. AB - The distinction between passive and active visuo-spatial memory has been useful to interpret various pattern of deficits reported in individual differences studies. However, this interpretation raises the issue of task difficulty, since active tasks could be failed simply because more complex and the corresponding deficit could reflect a reduced capacity of the system. We describe two children with Nonverbal Learning Disability whose performance provides evidence of a dissociation between passive and active memory processes. One of the children showed a selective impairment in passive tasks and performed flawlessly in active tasks, whereas the second child displayed the opposite pattern. These data suggest that a qualitative difference between passive and active processes does exist and that differences in performance do not reflect a lower/higher level of task difficulty. Further, these data underlie the importance of formulating theoretical models of visuo-spatial memory including both material-related (i.e., visual vs spatial) and process-related (i.e., passive vs active) distinctions. PMID- 10857677 TI - Writing impairment in schizophasia: two case studies. AB - Two case studies were conducted of patients suffering from paranoid schizophrenia with glossomanic schizophasia. Their dictation-taking performance was assessed via a comprehensive standardized neuropsychological test battery following a protocol specifically designed for the purpose (Lecours, 1996.). The patients' performance was compared with normative data from the general population. The results show that the patients committed graphemic paragraphias, that is, they replaced target graphemes with others representative of the corresponding phoneme (ph and f for/f/, in French) or with graphemes representative of a similar phoneme (v for/f/). These errors suggest that components of a cognitive model of writing are dysfunctional. PMID- 10857678 TI - Phonological awareness in French-speaking children at risk for reading disabilities. AB - We report the phonological awareness abilities of preliterate French-speaking children. The performance of a group of children identified At Risk (n = 26) for reading disabilities was compared to that of normally developing age-matched controls (n = 22) on a range of standardised and experimental tests. Results showed the At Risk children to have a selective impairment in expressive relative to receptive language, whereas Controls performed at equivalent levels on both measures. Although the children At Risk performed at a significantly lower level than Controls on all but one of the metaphonological tests, their pattern of performance was similar to that of Controls, suggesting a developmental delay. Interestingly, both groups showed a superiority of awareness for syllables over phonemes, reflecting the phonological structure of the French language. PMID- 10857679 TI - Signal-to-noise ratio sensitivity in ERPs to stimulus and task complexity: different effects for early and late components. AB - Traditionally, early and late ERP components are evaluated by the latency and amplitude of their components. We investigated their signal-to-noise (S/N) ratio: how well individual ERP trials correlate with their overall averaged ERP. High correlations indicate minimal noise or individual trial variability while low correlations reflect greater trial variability. Continuous EEG activity was recorded from 57 university students during single- and dual-task conditions in a within-subjects paradigm (easy and difficult tone discrimination; with and without verbal working memory task). For early components, the addition of the second task increased the correlations between the individual trials and the averaged ERPs. For the late components, both the addition of the second task and an increase in stimulus difficulty decreased individual trial correlations. Results suggest that the S/N ratio analysis is a useful method for highlighting differences between early and late components associated with stimulus processing manipulations. PMID- 10857680 TI - Language after hemispherectomy. AB - We studied the spoken language of 49 children who had undergone hemispherectomy as part of the UCLA Pediatric Epilepsy Surgery Research Program and analyzed, among a number of clinical factors, the relation between acquired vs developmental pathology and spoken language outcomes. In this paper we will briefly review the results of our study and attempt to explain (1) why "the early" is not always better, (2) why so many right hemispherectomies fail to develop language, and (3) why some left hemispherectomized children develop remarkably good language despite removal of the "language" hemisphere. This account will rest on the proposed model of brain maturation and progressive lateralization. PMID- 10857681 TI - Priming arithmetic reasoning in an amnesic patient. AB - The present study elucidates the role of implicit memory in problem solving by evaluating priming effects in a severely amnesic patient. An arithmetic series completion task was adopted to investigate the effects of lags, of items difficulty, as well as the influence of different distractor tasks on priming numerical reasoning. The results indicated that the priming effect was maximized in difficult trials and disappeared after long lags. On the other hand, the facilitation effect was not modulated by the nature of the distractor tasks. Critically, the experimental design controlled for peripheral facilitation of visual input and verbal output processes and a recognition task indicated that the effect may not be attributed to explicit memory. Thus, the facilitation must be located to a more central stage, possibly at the stage in which the abstract formula of the underlying algorithm are generated (Langdon & Warrington, 1997). PMID- 10857682 TI - Neuromotor microalterations in subjects with spasmophilia symptoms: a preliminary study. AB - Spasmophilia is a relatively unknown condition characterized by perturbations of the neuromuscular system. We hypothesized that spasmophilia may negatively affect neuromotor functions in subtle ways. Three tests including tremor, rapid pointing movements, and alternating movements were quantified in a group of subjects with spasmophilia symptoms (SS) (n = 10) and a healthy control group (n = 10). Most of the characteristics used to evaluate motor functions in these three tests revealed no significant differences between the two groups except for two characteristics in alternating movements and two characteristics in rapid pointing movements. In terms of variances, a dissociation between voluntary movements and involuntary movements was observed for the two groups. Control subjects had significantly higher variances in involuntary movements such as tremor, while subjects with SS had significantly higher variances in voluntary movements such as alternating and rapid pointing movements. A significant increase in asymmetry in hand laterality was also noted for some characteristics in subjects with SS. PMID- 10857683 TI - Differentiating the N3 and N4 electrophysiological semantic incongruity effects. AB - An event-related potential termed the N4 has been widely studied due to its sensitivity to semantic incongruity. A recent report (Nobre & McCarthy, 1994) indicates there is also an N3 component that is sensitive to semantic incongruity. To differentiate these two components, an existing data set with 65 electrode sites, 78 subjects, and 120 sentences was examined. Instead of the usual procedure of averaging over the stimuli within distinct categories for each subject, a new approach--averaging over subjects--was employed. In this item average approach, 120 averages (one per sentence) were produced. Correlational analyses indicate that the N3 is equally sensitive to cloze probability and sentential constraint. The N4, by contrast, is more sensitive to sentential constraint and less to cloze probability; it is also correlated with familiarity. We interpret these results as evidence that the N3 is more responsive to semantic fit whereas the N4 is more responsive to semantic expectancy. PMID- 10857684 TI - Superior single dimension relative to "exclusive or" categorization performance by a patient with category-specific visual agnosia: empirical data and an ALCOVE simulation. AB - ELM, a patient with category-specific visual agnosia, was tested on a single dimension categorization problem, and the "exclusive or" (XOR) categorization problem. Stimuli were computer-generated shapes in which exemplars within a shape set shared values across two visual dimensions (curvature and thickness). In single-dimension categorization only curvature was relevant, and ELM performed as well as normal participants. In the XOR problem, categorization depended on being able to extract from memory values on curvature AND thickness for each exemplar, and ELM was significantly impaired on this task. A computer simulation using ALCOVE (Kruschke, 1992) reproduced ELM's behavior by changing a single (specificity) parameter related to how easily proximate objects within a multidimensional shape space could be disambiguated. PMID- 10857685 TI - Musical instrument naming impairments: the crucial exception to the living/nonliving dichotomy in category-specific agnosia. AB - In category-specific agnosia (CSA) patients typically have more trouble naming animals, fruits, and vegetables than tools, furniture, and articles of clothing. A crucial exception to this living vs nonliving rule involves the category of musical instruments. Patients with problems naming living objects often repeatedly fail to name musical instruments. In CSA it is crucial to equate living and nonliving object lists on object name frequency, complexity, and familiarity. The present study shows, however, that even the most rigorously controlled object lists can lead to erroneous conclusions if nonliving stimuli contain an overrepresentation of musical instruments. Naming capabilities of a herpes encephalitis patient were assessed using matched lists of living and nonliving objects and showed no indication of category-specific deficits. When exemplars were separated into biological objects, musical instruments and man made artifacts, strong category-specificity emerged: artifact naming was flawless whereas musical instrument and biological object naming were both severely impaired. It is concluded that CSA is a veridical phenomenon but that our understanding of CSA is limited by adhering to the spurious living/nonliving distinction. PMID- 10857686 TI - Age, gesture span, and dissociations among component subsystems of working memory. AB - Working memory was examined in old and young adults using a series of span tasks, including the forward versions of the visual-spatial and digit span tasks from the Wechsler Memory Scale-Revised, and comparable hand gesture and visual design span tasks. The observation that the young participants performed significantly better on all the tasks except digit span suggested that aging has an impact on some component subsystems of working memory but not others. Analyses of intercorrelations in span performance supports the dissociation among three component subsystems, one for auditory verbal information (the articulatory loop), one for visual-spatial information (visual-spatial scratch-pad), and one for hand/body postural configuration. PMID- 10857687 TI - The use of reaction time measures to evaluate nonword reading in primary progressive aphasia. AB - We reported on a subject with nonfluent primary progressive aphasia (PPA), NL, who demonstrated an impaired ability to make rhyme judgments (Dowhaniuk, Dixon, Roy, Black, & Square, in press). Our hypothesis was that these deficits represent a precursor to phonological alexia. However, no definitive evidence supported the existence of a phonological reading impairment as NL made relatively few errors reading nonwords. To further evaluate NL's nonword reading, nonword and real word reaction times were compared. NL's reaction times were significantly longer for only nonwords compared to the slowest control subject. We then assessed the first two stages of processing involved in nonword reading (Coltheart, 1996). NL did not demonstrate deficits with graphemic parsing or phoneme assignment. His continuing problems with auditory rhyme judgments support the presence of a phonological processing deficit not specific to reading. We conclude that reaction time measures allow for the detection of subtle nonword reading deficits. PMID- 10857688 TI - Task dependent processing of visual information about target acceleration. AB - The objective of this study was to investigate the sensitivity of the perceptual and motor systems to target acceleration information using verbal magnitude estimations of target acceleration and manual interception of these targets. The results showed that in the perceptual task the participants were responding mainly to acceleration threshold values, which is acceleration as a function of initial, final, and average velocities, rather then to the absolute accelerations. When manually intercepting the targets the participants responded mainly to the absolute acceleration value and target initial velocity. Thus, these results suggest that target motion can be processed in the ventral (perception) and dorsal (action) visual streams however different motion characteristics are processed in these streams depending on the required output. PMID- 10857689 TI - Pantomime recognition impairment in Alzheimer's disease. AB - This study was designed to examine whether 10 AD patients with diffuse lesions and an early semantic deficit could comprehend the meaning of pantomimes and symbolic gestures. We especially wanted to determine the relationship between pantomime recognition and production. The tasks involved naming or showing an understanding (e.g., circumlocutions) of the pantomimes executed by the examiner and producing pantomimes to verbal command or to imitation. The results showed no significant relationship between pantomime comprehension and pantomime production. However, 60% of AD patients performed poorly compared to controls on the Gesture Recognition task. Gesture comprehension was highly correlated with a measure of global cognitive impairment (3MS). The data are consistent with the proposed separation of comprehension and action production systems in a cognitive model of praxis. Gesture comprehension may be diffusely distributed and thus preserved in the early stages of AD as in circumscribed lesions of the left parietal lobe. PMID- 10857690 TI - Linguistic lateralization and asymmetries in interhemispheric transmission time. AB - The Interhemispheric Conduction Delay (ICD) theory of cerebral lateralization claims that longer interhemispheric transfer times (IHTT's) should result in greater functional lateralization for time-critical tasks. IHTT was estimated in 40 normal participants using both a visual and an auditory version of the Poffenberger (1912) paradigm. There was a significant response-hand by side-of presentation interaction, and the length of IHTT for auditory information being transferred from the right to left hemisphere was significantly related to linguistic lateralization as assessed with a dichotic-listening task. These results support the ICD theory of cerebral lateralization. PMID- 10857691 TI - Understanding metaphoric sentences in the two cerebral hemispheres. AB - Previous studies indicate that the right hemisphere (RH) has a unique role in maintaining activation of metaphoric single word meanings. The present study investigated hemispheric asymmetries in comprehending metaphoric word meanings within a sentence context. Participants were presented with incomplete priming sentences followed by (literally) true, false, or metaphoric lateralized target words and were asked to decide whether each sentence is literally true or false. Results showed that responses to metaphoric sentences were slower and less accurate than to false sentences when target words were presented to the right visual field (RVF)-LH as well as to the left visual field (LVF)-RH. This suggests that the understanding of lexical metaphors within a sentence context involves LH as well as RH processing mechanisms and that the role of each hemisphere in processing nonliteral language is flexible and may depend on the linguistic task at hand. PMID- 10857692 TI - Controlling for stimulus dominance effects in the Fused Dichotic Words Test when predicting speech lateralization in children. AB - A log-linear analysis (Grimshaw et al., 1994) of Fused Dichotic Words Test data was examined in a sample of 28 children with epilepsy who had undergone the intracarotid amobarbital procedure to determine the nature of speech representation. The analysis yields a measure of ear dominance (lambda*) that controls for stimulus dominance confounds. Most patients with unilateral speech obtained statistically significant ear advantages, whereas most with bilateral speech displayed no significant ear advantage. Despite controlling for stimulus dominance confounds, some of the scores from patients with left-hemisphere speech overlapped with those from patients with bilateral speech representation. PMID- 10857693 TI - Effects of sleep deprivation on performance and EEG spectral analysis in young adults. AB - Nine totally sleep deprived (TSD) and nine control subjects were evaluated with a complete battery for attention and memory performance. Frontal and temporal EEGs (5 min, eyes closed) were also recorded before and after the night. TSD subjects exhibited three performance deficits: learning the Pursuit Rotor Task, implicit recall of paired words, and distractibility on the Brown-Peterson Test. Relative to evening recordings, control subjects showed decreased morning absolute powers in all electrodes for all frequencies except for Frontal delta; TSD subjects showed increased Frontal and Temporal theta and Frontal beta. These results show that motor procedural, implicit memory, and working memory are sensitive to one night of TSD, and that Frontal and Temporal theta spectral power seem to discriminate between a night with sleep from a night without. PMID- 10857694 TI - Pattern of semantic memory impairment in dementia of Alzheimer's type. AB - The specific pattern of semantic memory impairment in patients with dementia of Alzheimer's type (DAT) remains unclear. Specifically, the presence of a category specific deficit for biological concepts (reflected in anomia for these concepts) has been questioned. We studied 9 DAT patients using a semantic association judgement test in which they had to decide which of the two given words was most like a target word (e.g., lamb: goat, sheep). The 150 target words were drawn from 6 categories: animals, clothing and furniture, fruits and vegetables, tools, action verbs, and abstract nouns. Age- and education-matched control subjects performed equivalently (between 86 and 90% correct) in all categories. Compared to control subjects, DAT patients made significantly more errors in abstract and biological nouns, but not in verbs and man-made artifacts. This pattern of semantic memory impairment--a sparing of verbs and a selective deficit of nouns in the biological category--has been documented in patients with temporal lobe damage, suggesting a critical dysfunction in the temporal lobes of DAT. PMID- 10857695 TI - Laterality effects in processing tonal and atonal melodies with affective and nonaffective task instructions. AB - Right-handed university subjects were presented with monaural melodies that either conformed to the rules of the Western tonal system (tonal melodies) or that systematically deviated from it (atonal melodies) while containing similar contours and pitch skips. Subjects were tested under two different task instructions. One group was requested to judge whether each melody sounded correct or not (the nonaffective task); the other group had to judge whether each melody sounded pleasant or not (the affective task). The nonaffective task was found to elicit essentially no ear difference. In contrast, the affective instruction induced opposite and reliable laterality effects, depending on the valence of the response. The pleasant responses were indicative of a left hemisphere predominance and the unpleasant responses of a right hemisphere predominance. The results are consistent with the claim that the left hemisphere is biased toward positive emotions and the right to negative emotions. Moreover, the results suggest that affective appreciation of melodies is dissociable from their nonaffective judgment. PMID- 10857696 TI - Stimulus-response incompatibility effects on event-related potentials in children with attention-deficit hyperactivity disorder. AB - In order to test the hypothesis of a response choice deficit in attention-deficit hyperactivity disorder (ADHD) children, event-related potentials (ERPs) were recorded from 30 scalp electrodes in 21 ADHD and 21 normal boys during a spatial stimulus-response compatibility (SRC) task. ADHD children made fewer correct responses than control children, but did not show a larger incompatibility effect on response speed and accuracy. In ERPs, ADHD children had longer N1 latency and larger condition effect on the frontal N2, which would reflect a greater frontal involvement for the correct responses. The ADHD group who performed the SRC task first showed a larger condition effect on an early occipital P3 only, while the ADHD group who performed the SRC task second showed a larger condition effect on a later central P650 component and on a late parietal NSW, as compared with normal controls. These results suggest strategic differences in information processing in ADHD children, rather than a specific deficit. PMID- 10857697 TI - ERPs and behavioral inhibition in a Go/No-go task in children with attention deficit hyperactivity disorder. AB - In order to study the behavioral responses and the brain inhibition process in children with attention-deficit hyperactivity disorder (ADHD), Event-Related Potentials (ERPs) were recorded from 30 scalp electrodes in 21 ADHD and 21 normal boys during performing a Go/No-go task. ADHD children made fewer correct responses to both Go and No-go stimuli than normal controls. The frontal N2 amplitude was larger for No-go stimuli than Go stimuli in both groups, reflecting inhibition of responding. Smaller N2 amplitudes to No-go stimuli were found in ADHD children, but only when the Go/No-go task was performed after a first stimulus-response compatibility (SRC) task. In addition, the controls exhibited a prolonged N2 only when the Go/No-go task was performed second. However, the ADHD subjects exhibited this prolonged N2 when the task was first, but not when it was second. These results suggest an inhibitory regulation problem rather an inhibition deficit in ADHD children. PMID- 10857698 TI - The scripting of activities of daily living in normal aging: anticipation and shifting deficits with preservation of sequencing. AB - The purpose of this study was to elaborate upon Shallice's frontal lobe model by evaluating 20 elderly normal and 20 young adult controls, all women, with cognitive tests, a paper-pencil script generation task and a kitchenette meal preparation task. The elderly were significantly weaker on standardized tests of executive functions and on the paper-pencil script generation task. An anticipation and a sequencing impairment clearly emerged. Though anticipation and shifting were compromised in the kitchenette script task, sequencing was however preserved. This supports Shallice's notion according to which scripting is hierarchically organized with top level components being more frontal lobe dependent (supervisory attentional system or SAS) and the lower level components more tributary to subcortical circuits involving procedural learning (contention scheduling or CS). PMID- 10857699 TI - Writing and rewriting numerals: a dissociation within the transcoding processes. AB - In the present study we report the case of A.B., a patient with a specific, though not isolated, deficit in transcoding verbal to Arabic numerals. Despite perfect production of Arabic numerals in a writing to dictation task, she frequently produced syntactic errors when the input was in written verbal form (e.g., duecentotrenta [two hundred and thirty] --> 20030). In absence of problems in the verbal comprehension system, A.B.'s performance is difficult to accommodate within current models of number processing. In the attempt to interpret the present findings, we suggest that different numerical codes, i.e, spoken and written verbal numerals, activate with different efficiency the transcoding algorithm. PMID- 10857700 TI - Spatial and temporal organization of episodic and semantic processes revealed by event-related potentials to unfamiliar faces. AB - This study investigated whether the semantic and episodic processes underlying the old/new effect can be dissociated by analyzing the spatial and temporal characteristics of event-related potentials (ERPs) evoked by unfamiliar face stimuli during direct and indirect memory tests. The ERP old/new effects obtained included three components. First, the classical posterior component, present in both tasks, which was thus interpreted as reflecting changes in the automatic activation of semantic representation. Second, an earlier frontal component, present in the direct test only, which was seen a consequence of the building of a contextual representation of presented items formed of a conjunction of elements stored in semantic memory. Third, a late right-temporal component, in the direct test, which was related to the integration of this conjunction into a unitized representation of the presented stimuli. PMID- 10857701 TI - Tactile gap detection and language lateralization. AB - Recent work suggests that hemispheric language lateralization might be related to the fine-grained temporal discriminations that are required for linguistic processing (Nicholls, 1996). Studies concerning tactile processing have also shown a significant left-hemisphere (L-H) advantage for tactile gap detection (Nicholls & Whelan, 1997). We hypothesized that language and tactile processing are both preferentially processed by the left hemisphere because it is specialized for tasks requiring fine-grained temporal resolution. Thirty-two participants (16 right and 16 left handers) were tested for both linguistic processing (using the Fused Dichotic Words Test (FDWT)) and tactile gap detection. Both right and left handers showed a significant L-H advantage for both language processing and tactile gap detection. The present study supports recent claims that language lateralization is attributable to the left hemisphere's better suitability to process tasks that require fine-grained temporal resolution. PMID- 10857702 TI - Investigating visual knowledge in dementia of the Alzheimer type (DAT): evidence for a structured organization. AB - Given that certain types of semantic dysfunction in DAT have been attributed to difficulties in accessing visual knowledge, the issues of integrity of access to, and format of storage, within the "visual store" are particularly important. In these experiments, we tested 10 DAT patients and matched controls on the reality decision and part-whole matching tasks. Visual knowledge was found to be significantly impaired in the DAT group. Moreover, DAT patients displayed disproportionate deficits in accessing visual knowledge of items from nonliving categories. Results are interpreted within a framework of differential hierarchical access to visual representations of perceptually similar and perceptually distinct categories of items. PMID- 10857703 TI - Side preference in adults for holding infants: contributions of sex and handedness in a test of imagination. AB - Five hundred one right-handers (150 men, 351 women) and 53 left-handers (15 men, 38 women) were asked to imagine holding a young infant in their arms. Right handers reported significant left-side biases--in 68% of the men and 73% of the women. For left-handers, side preferences were weaker, the left-side bias dropping to 47% for men and 60% for women, with neither figure different from chance. The results are discussed in the context of theory and research on the functional neuroanatomy of attention, emotional arousal, and the generation, maintenance, and manipulation of mental images. PMID- 10857704 TI - The expression of manual asymmetries following extensive training of the nondominant hand: a kinematic perspective. AB - A single experiment involving 5600 discrete aiming trials with the left hand and 560 trials with the right hand designed to track the kinematic changes associated with left hand practice is reported. In general, performance data revealed that following practice movement time of the left hand achieved a level of performance similar to that of the right. However, the kinematic variables analyzed were instructive in identifying differential control processes associated with the right and left hand. Specifically, the right hand was more accurate during the ballistic phase of movement execution, while practice enabled the left hand to utilize response-produced feedback more efficiently. PMID- 10857705 TI - Double dissociations and neurophysiological expectations. AB - Double dissociations are among the most highly regarded and influential evidence for particular modular divisions of cognitive performance. We present here a method for obtaining double dissociations across any task, even within an avowedly indivisible and unmodular system. We argue that the existence of double dissociations is not necessarily evidential and suggest what sort of additional evidence would be necessary to support the desired conclusions. We suggest that the idea of expected behavior after injury is important and should receive more attention. PMID- 10857706 TI - Not only in the brain: Cabanis and the Montpellierian tradition of localization. AB - Antonio Damasio (1995) has recently presented evidence to the effect that we are perhaps wrong in thinking that it is only the brain that thinks. Rational decision making involves emotional reactions as a necessary condition and background. And since emotions involve bodily reactions which are not limited to the brain but which embrace the autonomous nervous system and the viscera, one could say that we actually think with our bodies and not merely with our brains. According to Damasio the incapacity of patients with frontal lobe pathology in decision making could be explained by a disturbance in emotional reactions involving the whole organism. Philosophical discussions concerning brains in a vat have completely forgotten these aspects of our mental life. Despite the fact that the idea that we think exclusively with our brains has during the modern age been a rather widely held "received view," there is a physiological and philosophical tradition which regarded mental functions as the result of the interaction of several organs, instead of seeing them as the result of the activity of the brain alone. PMID- 10857707 TI - Handedness and footedness in Korean college students. AB - This study was conducted to obtain normative data on foot preference and to compare footedness and handedness in a large sample (N = 866) of college students in Korea, where left-hand use for writing and other public acts is severely restricted (Kang & Harris, 1993). Based on scores from Korean-language versions of the Edinburgh Handedness Inventory (EHI; Oldfield, 1971) and the Waterloo Footedness Questionnaire Revised (WFQ-R; Elias, Bryden, & Bulman-Fleming, 1988), 11% of the subjects were left-footed but only 4.2% as left-handed. A significantly higher percentage of left-handers than right-handers showed crossed lateral preference, that is, for preference of the opposite-side foot. Of the left-handers with crossed preference, the majority were inconsistent left-handers (ILH; Peters & Servos, 1989), whereas most of those with uncrossed preference were consistent left-handers (CLH). Factor analysis of the EHI and WFQ-R revealed 2 handedness factors and 2 footedness factors. The footedness factors for skilled unipedal actions and for balancing-stabilizing varied in direction, strength, and relation to handedness in mixed-footers and left-handers, consistent with the possibility that the division of footedness into these categories might be neuropsychologically meaningful. PMID- 10857708 TI - The effects of nicotine on Parkinson's disease. AB - Post-mortem studies have demonstrated a substantial loss of nicotinic receptors in Parkinson's disease (PD), which may be at least partially responsible for some of the cognitive, motoric, and behavioral deficits seen in this disorder. Epidemiologic studies have suggested that cigarette smoking is a strong negative risk factor for the development of PD. We have previously shown that blockade of central nicotinic receptors produces cognitive impairment in areas of new learning, short-term memory, and psychomotor slowing with increasing dose sensitivity with age and disease. Studies of acute stimulation of nicotinic receptors in Alzheimer's disease with nicotine and the novel agonist ABT-418 in our laboratory and others have shown improvements in several measures of cognitive function. Prior studies of the effects of nicotine in PD have suggested some improvements in clinical symptomatology. We have begun quantitative studies of both acute and chronic nicotine in PD to assess both cognitive and motor effects. Fifteen (15) nondemented subjects (age 66 +/- 5.3; M/F = 11/4) with early to moderate PD (mean Hoehn-Yahr stage = 1.77; MMSE = 28.6) received a dose ranging study of intravenous nicotine up to 1.25 microg/kg/min, followed by chronic administration of nicotine by transdermal patch with doses ranging up to 14 mg per day for 2 weeks. Testing occurred both during drug administration and up to 2 months after drug cessation to look for prolonged effects. Preliminary analysis shows improvements after acute nicotine in several areas of cognitive performance, particularly measures such as reaction time, central processing speed, and decreased tracking error. Improvements in attention and semantic retrieval were not seen. After chronic nicotine, improvements were seen in several motor measures suggesting improved extrapyramidal functioning. This appeared to be sustained for up to 1 month after drug. The treatment was well tolerated. Nicotinic stimulation may have promise for improving both cognitive and motor aspects of Parkinson's disease. PMID- 10857709 TI - Dissociation of reading strategies: letter-by-letter reading in the native language and normal reading in the learned language. A case study. AB - In letter-by-letter reading, which is typically observed in Dejerine's (1892) "pure alexia," oral reading seems to be mediated by the naming of the constituent letters of the printed sequence: reading time rises abnormally as a function of the number of letters of the target item. We describe a patient with fluent aphasia who showed the unusual pattern of letter-by-letter reading together with surface dyslexia in her native language (French) and apparently normal reading in the second, learned language (English). Thus, letter-by-letter reading should be considered as a description of a symptom rather than the consequence of a unique type of functional impairment. PMID- 10857710 TI - Age of acquisition and name agreement as predictors of mean response latencies in picture naming of French adults. AB - We studied the influence of five variables on picture naming latencies in French adult subjects: frequency, age of acquisition and length of words, name agreement in oral naming, and visual complexity of pictures. 140 pictures were presented to 56 subjects with control of individual factors (age, educational level, gender). Naming latency was measured as the time from picture onset until keystroke corresponding to the begin of verbal production. The results of simple and multiple regression analyses show that only two variables make independent contributions to mean response latency: age of acquisition and name agreement. Such state of affairs confirms similar findings obtained with English speaking subjects. PMID- 10857711 TI - Do living and nonliving categories need further fractionation? A study of picture naming in a pathological sample. AB - A group of 51 patients affected by possible semantic memory deficit were given a picture naming task for the purposes of a comparison between six categories, three of Nonliving nature (tools, furniture, vehicles) and three of a Living nature (animals, fruits, and vegetables). A logistic regression analysis was used for a multiple single case study, where also the items' basic difficulty was included in the model. Besides some patients showing a dissociation between Living and Nonliving categories, other patients showed a finer selectivity on naming, differentiating animals from fruits and vegetables, tools from nonmanipulable objects, and even vehicles from furniture. These results are examined in the light of current theories of semantic category specificity. PMID- 10857712 TI - A possible approach for discrimination between normal and pathological signals using the WFLC algorithm. AB - The discrimination between normal and pathological tremor is difficult when amplitude is relatively small. The WFLC algorithm, a time domain adaptive Fourier transform, is designed to control physiological tremor and to improve precision during microsurgery. We added two iterative optimization processes to initialize the following parameters: initial frequency weight (omega0), amplitude adaptation rate (mu) and frequency adaptation rate (mu0). Then, we applied the methods on data sets recorded on patients with different tremors (control, parkinsonian, cerebellar, and essential) sampled at 200 Hz. After filtering the data, the WFLC algorithm tracked the time-varying dominant frequencies and amplitudes of the transformed data sets. Our results illustrate the potential of using this algorithm as an approach to discriminate between normal and pathological signals even when amplitude is not a significant discriminating factor. PMID- 10857713 TI - Modularity of object and place memory in children. AB - The present study investigated modularity of object and place memory in children with the reaction time/accuracy paradigm. The memory task was presented in two spatial arrays, a frame with landmarks and a grid. Modularity was tested using perceptual size judgment (what-interference) and movement direction judgment (where-interference). Latencies of object memory were increased by same-system what-interference in all age groups indicating limited capacity of an object memory system. With respect to accuracy of object and place memory, only young children's performance was susceptible to same-system interference of perceptual judgment. PMID- 10857714 TI - The association between stress, hemispheric specialization, and callosal interactions. AB - This study examines the effects of stress on hemispheric specialization and callosal transfer. Previous research suggests that the two cerebral hemispheres are asymmetric in their regulation of and response to stress. Further, studies have suggested that the activity of the corpus callosum may also be affected by stress. A group of 28 participants completed the Spielberger State-Trait Anxiety Inventory and performed a bilateral Stroop task. The bilateral Stroop task contains separate conditions which measure both hemispheric specialization and callosal transfer. A significant correlation was found between state anxiety and our measure of callosal transfer. Specifically, those individuals with greater state anxiety experience more efficient callosal transfer than did those with lower state anxiety. We conclude that state anxiety enhances callosal transfer without affecting hemispheric specialization. PMID- 10857715 TI - The analysis of drawing from memory performance in brain-damaged patients. AB - The drawing from memory task is frequently used in cognitive neuropsychology to investigate visual processing impairments. However, in surprising contrast to most other neuropsychological tests, the analyses of results on this task are most often based solely on qualitative judgements about the normality of a patient's performance. In most case reports, these judgements are not made with reference to normative data and are not made by individuals who are impartial with respect to the study (that is, using a blind rating procedure). There are several grounds for arguing that such analyses are inadequate. First, seemingly abnormal drawings made by a patient may well be within the range of performance shown by control subjects. Second, judgments that are not based on a blind rating procedure are likely to be influenced by knowledge about the patient's performance on other tasks. We describe an alternative assessment procedure that addresses both of these concerns. PMID- 10857716 TI - Context use by right-hemisphere-damaged individuals under a compressed speech condition. AB - The effect of increased processing demands on context use by RHD individuals was examined using a word-monitoring task. Subjects were required to monitor for a target word in sentences that were either normal, semantically anomalous, or both syntactically and semantically anomalous. Stimuli were presented at two rates of speech--normal and compressed to 70% of normal. Contrary to expectations, the RHD group performed similar to normals in demonstrating an effect of context at both rates of speech. Results are discussed relative to recent studies of normal brain functioning that suggest that the involvement of the LH versus the RH in context use depends upon the type of contextual information being processed. PMID- 10857717 TI - Syntactic comprehension deficits in agrammatism. AB - Eleven agrammatic and 16 fluent aphasic patients were given a comprehension task consisting of simple, active and passive reversible sentences. The purpose of the study is to reconsider the comprehension disorders in agrammatism, and particularly of passive reversible sentences, to test to what extent Grodzinsky's trace deletion hypothesis (TDH) is generalizable to other types of NP-movement, and finally to ascertain whether the pattern of impairment observed in agrammatism differs from that of fluent aphasic patients. The study confirms that trace analysis may be selectively impaired in agrammatism. However, this deficit is not the only mechanism underlying comprehension disorders and cannot be said to occur in all agrammatic patients. Comprehension disorders also involve the processing of clitic object pronouns which also underly NP-movement. Finally, the impairment found in fluent aphasic patients differs, both in type and severity, from that of agrammatic patients, thus confirming the peculiar aspects of the agrammatic comprehension deficit suggested by Grodzinsky's TDH. PMID- 10857718 TI - Effect of a directional cue on line bisection. AB - Normal participants display a leftward bias when bisecting horizontal lines. It has been proposed that bisection errors can be affected by representational biases. We investigated the possibility that this effect could also be explained as the result of perceptual cueing. Participants bisected horizontal lines with identical arrows arranged around the lines, all pointing to the same end. The arrows significantly affected bisection placement, although participants bisected significantly away from the direction the arrows pointed to. The findings favor the hypothesis that cueing, rather than representation, can affect bisection errors. PMID- 10857719 TI - Reducing neglect by introducing ipsilesional global cues. AB - Halligan and Marshall (1994) have proposed that unilateral neglect (UN) can be reduced by ipsilesional global cues which induce processing of the whole object, including contralesional features. This hypothesis has the potential to distinguish difficulties with disengaging attention after ipsilesional capture from the failure of contralesional stimuli to capture attention. To test the role of global cues, seven individuals with unilateral right-hemispheric lesions copied and verbally identified drawings adapted from Seron, Coyette, and Bruyer (1989). The level of meaningful information presented in the ipsilesional hemispace was varied across three levels: symmetrical objects, objects with an identifying left-side feature, and objects that required processing of the left side information for plausible identification. The results support the global processing hypothesis by demonstrating that the less the ipsilesional information, the less was the neglect of contralesional information. PMID- 10857720 TI - Language development in a child with left hemispherectomy. AB - A longitudinal study of a left hemispherectomized boy (AB) was conducted to document linguistic evolution and maturation and determine the extent to which right hemisphere processes allow development of language. Resection of the left hemisphere occurred at age 5 years 6 months, following intractable epilepsy. Tests of language comprehension (pointing, understanding of prepositions, understanding of narratives) and production (naming, repetition, lexical diversity, grammatical production) were administered at ages 6:2, 6:4, 6:6, and 6 years 9 months. Observations showed little progress, if any, in most aspects of linguistic performance. In contrast to studies with left-hemispherectomized children, AB showed only a modest expansion of the semantic lexicon and the phonological repertoire more than a year after the surgical intervention. These observations indirectly suggest (1) poor functional involvement of the right hemisphere in the development of adequate linguistic abilities, (2) the necessary integrity of the LH for adequate development of language, or (3) that variations in individual brain maturation rates may account for AB's linguistic progress. PMID- 10857721 TI - Structural influences on the resolution of lexical ambiguity: an analysis of hemispheric asymmetries. AB - Structural influences on lexical ambiguity resolution in the two cerebral hemispheres was investigated using a divided visual field procedure. Participants were presented with auditory Wh- sentences containing an ambiguous word, where the grammatical role of the word was apparent only at a sentence-final verb (e.g., "Which BANK did the woman see?"). Following a sentence, either immediately or after 600 ms, a target word was presented in either the right or left visual field. Targets were related to the ambiguous word's dominant meaning (MONEY), the subordinate meaning (RIVER), or were unrelated. With left visual field presentation, priming occurred for both dominant- and subordinate-related targets at a 0 ms delay, but only for dominant-related targets at 600 ms. With left visual field presentation, priming occurred for subordinate-related targets only at both delays. The results suggest that grammatical assignment triggers the selection of meaning in the left hemisphere, whereas processing in the right hemisphere operates independently of structural analyses. PMID- 10857722 TI - Is the hand to speech what speech is to the hand? AB - Interference between the manual and the verbal performance on two types of concurrent verbal-manual tasks was studied on a sample of 48 female right handers. The more complex verbal task (storytelling) affected both hands significantly, the less complex (essentially phonemic) task affected only the right hand, with insignificant negative influence on the left-hand performance. No significant reciprocal effects of the motor task on verbalization were found. PMID- 10857723 TI - Automatic and controlled processing in chronic tic disorders. AB - The study compared controlled and automatic processing in 17 people diagnosed with chronic tic disorder (CTD), 20 with habit disorder (HT), and 30 screened controls. Using a countermanding paradigm, time to initiate a response (GO-time) was compared with time taken to inhibit a response (STOP-time) under automated and controlled response conditions. The signals were 2 sets of traffic-light-like computer displays with ready, go, and stop lights. The automated response was a repetitive series of taps and the controlled response was a morse code pattern of taps. There were no group differences in GO time under any conditions. The control group but not the HD or CTD groups showed a significant practice effect over time. The CTD group was significantly slower to STOP the automated than the controlled response. The results suggest that the CTD group had specifically, greater difficulty inhibiting automated than controlled actions. PMID- 10857724 TI - Asynchronous language acquisition in developmental dysphasia. AB - A longitudinal study was conducted to document and compare evolution of children with linguistic acquisition impairment. To determine whether development of the analytic mechanisms underlying linguistic processing occured in similar fashion, two children with mixed developmental dysphasia were assessed from 4 to 5:6 years of age with psycholinguistic tests at 6-months interval. Spontaneous speech and language production (consonant repertory in initial word position, MLU, and lexical diversity) were investigated in a standardized symbolic play context. The phonologic and lexico-morphologic evolution analyses revealed a marked improvement in motor control of phonology and in the application of morphosyntaxic rules in child 1, whereas child 2 was still impaired in phonology and morphosyntax. The singular developmental changes in spontaneous speech results indicate dynamic relationships between various language production facets and variability in the kind of deficit and lexical automation presented by these children. These contrasts in the evolution of language production profiles between child 1 and child 2 also underline the importance of longitudinal studies in the analysis of the atypical linguistic processing paths used by children with developmental dysphasia. PMID- 10857725 TI - Atypical cognitive disorders in a man with developmental surface dyslexia. AB - The neuropsychological profile of a man with a developmental surface dyslexia is presented here. This case study is of interest because J.C. exhibited a pattern of cognitive disorders rarely documented in previous data. Results showed that JC's difficulties in reading comprehension were closely related to complex memory disorders and were also associated with cognitive slowness. The present observations do not support the visual memory failure hypothesis. The data rather suggest that the core difficulty primarily lies with the nonautomatization of grapheme-phoneme correspondence rules, which in turn dramatically contributed to lexicon weaknesses. The hypothesis of a timing mechanism in reading disorders is discussed. PMID- 10857726 TI - Ocular motility and visuo-spatial attention in children with periventricular leukomalacia. AB - Periventricular leukomalacia (PVL) is known to cause visual, motor, and cognitive impairments varying in their severity. Most studies focused on impairment at early ages. Little is known about the PVL's later outcome. Effects of neural plasticity are for instance insufficiently known to prognose precisely their behavioral outcome. It is even hard to determine the consequence of one defect on global development over time. Because of the established neural link between eye motion and space perception, 5-to-9 year-old PVL's were tested in visual detection, postural control, and space attention tasks. In this paper, discussion will focus on behavioral asymmetry, developmental outcome, and brain injury. PMID- 10857727 TI - Selective effects of prior motivational experience on current on-line control of attention. AB - This study examined the influence of prior motivational experience on the efficiency of executive attention control during performance of a task set reconfiguration task (Rogers & Monsell, 1995). Results revealed that motivation manipulations selectively affected attention switching mechanisms, but did not influence either inhibition of crosstalk from competing stimuli, or basic response execution time. This provides additional evidence for distinct attentional systems involved in resolution of response and perceptual competition. Speculations regarding the neural systems that mediate both motivation and attention switching are considered, pointing to a possible involvement of dopaminergic influences on the ventral striatum. PMID- 10857728 TI - A developmental analysis of the relationship between hand preference and performance: I. Preferential reaching into hemispace. AB - The present study describes a developmental performance measure of hand preference that considers task complexity and position in hemispace. Eighty right handed children and adults (ages 3-4, 6-7, 9-10, 18-24) were observed for hand selection responses to 2 unimanual tasks (simple vs complex) across positions in hemispace. Results revealed an age-related trend in the tendency to use the preferred hand in right and left hemispace. While the adult's and 3- to 4-year old's preferred hand use decreased as they moved into left hemispace, children between the ages of 6 and 10 years tended to use their preferred hands consistently throughout both regions of hemispace. The relationship between hand preference and skilled, cost-efficient performance throughout development are discussed. PMID- 10857729 TI - An EEG coherence test of the frontal dorsal versus ventral hypothesis in N-back working memory. AB - We examined Goldman-Rakic's conceptualization of working memory (WM) involving modality-specific reverbatory circuits using electrophysiological measures. An n back WM task, with either spatial information or object information to be remembered, was administered with EEG coherence used to provide a functional measure of corticocortical communication. Support was not found for modality specific WM systems in this task. Instead, results suggest that there is a circuit that connects dorsal frontal with dorsal parietal areas which is activated during all levels of the WM vigilance task, regardless of the type of stimuli involved. Data also suggest that left frontal areas are sensitive to the level of difficulty of the WM task (0,1,2, and 3-back), as well as familiarity of task demands. Overall, our results support a model that conceptualizes a unitary WM system rather than a system composed of separate WM circuits for different modalities. PMID- 10857730 TI - Visual aiming movements in Alzheimer's disease. AB - This study examined whether AD affects the control of pointing movements. Sixteen older adults with probable AD and 10 age-matched healthy adults pointed at targets varying in size (3 and 7 mm in diameter) and located at three positions (at the midline and 33 degrees to the left and right). Results revealed the patients exhibited longer movement time, lower peak velocity and more time in deceleration, although they exhibited effects of target size and location comparable to the healthy controls. AD, then, would appear to slow down movement without affecting the motor system's ability to respond to task demands. PMID- 10857731 TI - Proper names in the early stages of Alzheimer's disease. AB - A short proper name retrieval test was found to discriminate between unselected subjects and age- and education- matched patients affected by Alzheimer's type dementia at a stage where the Mini-Mental State Examination could not detect a difference between these groups. The proper name retrieval task also compared favorably with the 3MS, a more sensitive, modified version of the MMSE. These findings suggest that proper name retrieval could be used to an advantage and become a routine component of short batteries for the early detection of Alzheimer's disease. PMID- 10857732 TI - Memory for/and execution of future intentions: evidence from patients with Herpes Simplex Encephalitis. AB - Prospective remembering was studied in a group of patients who suffered from Herpes Simplex Encephalitis (HSE). All patients showed a marked deficit in executing intentions for future actions under all the given constraints. The deficit extended to both time- and event-based intentions. The analysis of errors showed somewhat different patterns and some dissociations with evidence for selective preservation or damage to specific components involved in prospective remembering. These patients, in fact, may fail because of a difficulty in taking into account all given constraints or in activating stored intentions and in forming intentions or in remembering the content of the actions. Time- and event based tasks seem to show different sources of errors also in the same patient. PMID- 10857733 TI - Lateralization of speech processing in the brain as indicated by mismatch negativity and dichotic listening. AB - The goal of the present study was to evaluate the differences between dichotic listening and mismatch negativity as measures of speech lateralization in the human brain. For this purpose, we recorded the magnetic equivalent of the mismatch negativity, elicited by consonant-vowel syllable change, and tested the same subjects in the dichotic listening procedure. The results showed that both methods indicated left-hemisphere dominance in speech processing. However, the mismatch negativity, as compared to the right-ear advantage, suggested slightly stronger left-hemisphere dominance in speech processing. No clear correlation was found between the laterality indexes of mismatch negativity and right-ear advantage calculated from dichotic listening results. The possible explanation for this finding would be that these two measures reflect different stages of speech processing in the human brain. PMID- 10857734 TI - Reading/writing vs handedness influences on line length estimation. AB - Left- and right-handed school children with differing reading/writing experiences (unidirectional left-to-right vs bidirectional) were asked to draw 3-cm lines from right-to-left or from left-to-right with each hand. With either hand, lines drawn from left to right were more accurate than those drawn from right to left, particularly for right-handed left-to-right users; bidirectional readers showed no directional bias. Moreover, bidirectional readers were more accurate than unidirectional readers. The findings support a greater influence of directional scanning effects than handedness on the task of line length estimation. PMID- 10857735 TI - Left anterior lobectomy and category-specific naming. AB - Damasio and colleagues (1996) have proposed that the left anterior temporal region supports knowledge pertaining to living objects, whereas more posterior temporal regions play a critical role in naming nonliving things. Accordingly, one might expect that left-sided anterior temporal lobectomy should have a more profound effect on the naming of living as opposed to nonliving things. As part of a multicenter collaborative project, seventy-nine patients (all left hemisphere speech dominant) were tested pre- and post-left-temporal lobectomy on a task that required naming of living and nonliving items equated for name frequency, familiarity, and visual complexity. Consistent with the proposals of Damasio et al. (1996), left temporal lobectomy impaired naming ability, particularly for living things. When individual outcomes were considered, twice as many patients showed a relative decline in naming living as opposed to nonliving things. PMID- 10857736 TI - A novel approach to the amplitude vs position control controversy. PMID- 10857737 TI - Pierre Jean Georges Cabanis (1757-1808): an early nineteenth century source for the concept of nervous energy in European behavioral neurosciences. AB - The nineteenth century witnessed many advances in neuroscientific concepts. Among the notable are Charles Bell's (1774-1842) and Francois Magendie's (1783-1855) identification of sensory and motor pathways, Thomas Henry Huxley's (1825-1895) elaboration of evolutionary theory in the context of comparative neuroanatomy, and Emile Du Bois-Reymond's (1818-1896) and Hermann von Helmholtz's (1821-1894) work in experimental neurophysiology and on the concept of nervous energy. In Germany, the idea that the nervous system consisted of two elements, one that generated nervous energy and another that conducted it throughout the body, had wide currency in mid-nineteenth century. In France, Pierre Jean Georges Cabanis (1757-1808), physician, philosopher, and one of the founders of modern psychophysiology, argued that the brain is the part of the body in which electricity is stored. In his Rapports du Physique et du Moral de l'Homme, published between 1796 and 1802 (translated into German under the title Verhaltnis der Seele zum Korper (1808)), Cabanis proposed new ideas on brain function, on the brain's own sensibility, on the concept of will, and on the chemical basis of nervous activity. In the Rapports Cabanis proposed a theory of how brain and nerves relate to thought and behavior. Foreshadowing later developments in neuropsychology, he suggested that different parts of the nervous system have separate functions. Despite the fact that Cabanis had many interesting ideas about brain function, he has been largely ignored by historians of neuroscience; e. g., he is mentioned briefly in Clark and Jacyna (1989), in only two footnotes in Neuburger (1897/1981), and not at all in Finger (1994). Cabanis's far-reaching theory of how the brain works helped shape understanding of the general notion of nervous energy in nineteenth-century European neuroscience. PMID- 10857738 TI - Reliability of a dichotic consonant-vowel pairs task using an ABX procedure. AB - The present study examined the reliability of a dichotic consonant-vowels (CV) pairs task using an ABX procedure. Twenty-four right-handed subjects (12 females, 12 males) completed a dichotic CV pairs task twice in a test-retest approach. The conventional free-recall or directed-attention procedure was replaced by an ABX procedure, in which participants were required to determine whether a CV heard binaurally was present in a dichotic pair presented on the same trial. Results showed the expected right-ear advantage in dichotic listening for verbal material. However, reliability was not as high as has been reported with a directed-attention procedure. Methods for improving the reliability of the ABX discrimination task are discussed. In addition, the discussion emphasizes the usefulness of the ABX approach for testing clinical populations. PMID- 10857739 TI - Posture and target location effects on manual preference. AB - Twenty-nine right-handed participants made unimanual movements to 5 target locations while standing and while balancing on a rocker platform. Manual preference was affected by target location (p < .001; manual preference tended to be concordant with target hemispace) and posture (p < .05; responses from the rocker platform were associated with a lower level of right-hand responding). A location by posture interaction (p < .05) indicated that the effect of posture was present only for targets in left hemispace; the right hand was used less frequently in the rocker condition than simple standing. Manual preference is viewed as a process of response selection motivated by an effort to minimize neural processing and mechanical costs. PMID- 10857740 TI - Oral and written discourse in adolescents with closed head injury. AB - Analysis of oral and written discourse suggested differing cognitive demands for modes of expression. Verbal samples were provided by 8 adolescents with closed head injury (CHI) and 8 controls. A generation task using a picture stimulus was the basis for discourse. Eight measures [productivity, efficiency, semantic ties (lexical, incomplete, elliptical), maze use, coherence (global, local)] were utilized. A covariate model consisting of group membership (CHI vs control), executive functioning and working memory helps to explain variance in the discourse skills of adolescents with CHI. PMID- 10857741 TI - Seasonal variation in human functional cerebral lateralization and free testosterone concentrations. AB - Men have previously been reported to exhibit seasonal fluctuations on specific types of cognitive performance. It has been speculated that this performance variability is a result of changes in cerebral asymmetry due to lowered testosterone concentrations in the spring relative to the fall. In the present study, functional cerebral lateralization was measured in a group of men and women in the spring and fall. Free testosterone concentrations were assessed for participants to determine what associations might exist between seasonal variability in lateralization and seasonal fluctuations in testosterone exposure. Men and women tested in the spring exhibited exaggerated patterns of asymmetry compared to participants tested in the fall. Lower testosterone concentrations were observed in the spring compared to the fall in women, but not men. No direct associations between testosterone and lateralization were detected for either sex at either season. These results illustrate that seasonal fluctuations in testosterone exposure do not directly influence seasonal changes in functional lateralization patterns. PMID- 10857742 TI - Hemispheric contributions to pragmatics. AB - Twenty-seven patients with right-hemisphere damage (RBD) and thirty-one patients with left-hemisphere damage (LBD) received a new pragmatics battery in Hebrew consisting of two parts: (1) comprehension and production of basic speech acts (BSAs), including tests of assertions, questions, requests, and commands, and (2) comprehension of implicatures, including implicatures of quantity, quality, relevance, and manner. Each test had a verbal and a nonverbal version. Patients also received Hebrew versions of the Western Aphasia Battery and of the Right Hemisphere Communication Battery. Both LBD and RBD patients were impaired relative to controls but did not differ from each other in their overall scores on BSAs and on Implicatures when scores were corrected by aphasia and neglect indices. There was a systematic localization of BSAs in the left hemisphere (LH) but not in the right hemisphere (RH). There was poor localization of Implicatures in either hemisphere. In LBD patients, BSAs were associated with language functions measured with the WAB, suggesting the radical possibility that the classic localization of language functions in aphasia is influenced by the localization of the BSAs required by aphasia language tests. Both BSAs and implicatures show greater functional independence from other pragmatic, language and cognitive functions in the RBD than in the LBD patients. Thus, the LH is more likely to contain an unmodular domain-nonspecific set of central cognitive mechanisms for applying means-ends rationality principles to intentional activity. PMID- 10857743 TI - A high-density single-nucleotide polymorphism map of Xq25-q28. AB - A high-density single-nucleotide polymorphism (SNP) map was developed for Xq25 q28 using a targeted approach to SNP discovery. This high-density map includes 217 new SNP markers, and 117 are informative in the CEPH parent population with >20% minor allele frequency. The average distance between SNP markers is 100 kb in the targeted regions. This is the densest genetic map of Xq25-q28 to date. The SNP markers are presented in order by their distance in megabases along the X chromosome, and the markers from the current genetic map are placed using the same scale to produce an integrated map of the region. PMID- 10857744 TI - A new submicroscopic deletion that refines the 9p region for sex reversal. AB - Male to female sex reversal has been described in patients with deletions of chromosome 9p, and a region critical for sex reversal has been localized to p24.3, at the tip of the chromosome (TD9). It was proposed that the sex reversal may arise by haploinsufficiency for a gene localized to the minimum deletion. The 9p24.3 genes DMRT1 and DMRT2 are the favorite TD9 candidates to date, in virtue of their sequence similarity to doublesex and mab-3, sexual regulators in Drosophila and Caenorhabditis elegans, respectively. The hypothesis of sex reversal by combined haploinsufficiency for the two genes was put forward to explain the lack of mutations in either gene in XY sex-reversed females. Here we describe a XY sex-reversed patient carrying a novel 9p deletion that extends over less than 700 kb of genomic DNA. This region defines the smallest interval for sex reversal found to date. DMRT1 and DMRT2 map outside this region. Our data do not support the hypothesis of combined haploinsufficiency for DMRT1 and DMRT2. Nevertheless, DMRT1 localizes very close to the deletion breakpoint and has a pattern of expression compatible with a role in sex determination. It therefore remains a candidate gene for 9p sex reversal. PMID- 10857745 TI - A transcript map of a 2-Mb BAC contig in the proximal portion of the mouse X chromosome and regional mapping of the scurfy mutation. AB - A physical clone contig has been constructed, spanning 2 Mb on the proximal mouse X chromosome containing the mouse scurfy (sf) and tattered (Td) mutations. Extensive transcript mapping in this interval has identified 37 potential transcription units, including a number of novel genes, and 4 pseudogenes. These genes have been ordered by STS content and restriction mapping. Comparison of the transcript map to the corresponding region in human Xp11.23-p11.22 shows extensive homology, with complete conservation of gene order for loci in common between the two maps. Further, using a novel method to identify simple sequence length polymorphisms, we have developed a number of genetic markers, which has enabled the region containing the sf mutation to be narrowed to <300 kb. This contig has already allowed the cloning of the Td gene using a candidate gene approach and now serves as a starting point for the cloning of the sf mutation. PMID- 10857746 TI - The human and mouse Period1 genes: five well-conserved E-boxes additively contribute to the enhancement of mPer1 transcription. AB - The clock gene, Period1, from human and mouse was sequenced and characterized. Both human PERIOD1 (human PER1) and mouse Period1 (mouse Per1) consisted of 23 exons spanning approximately 16 kb, and their structures showed strong similarity to each other. For example, six highly conserved regions were identified in the 5' upstream sequences. These conserved segments exhibited 77-88% identity and possessed several potential regulatory elements including five E-boxes (the binding site of the CLOCK-BMAL1 complex) and four cyclic AMP response elements. Transient transfection assays using a mPer1-luciferase fusion gene revealed that each of the conserved E-boxes additively functions as an enhancer for the transactivation of mPer1 by mCLOCK and mBMAL1. PMID- 10857747 TI - Alterations of H19 imprinting and IGF2 replication timing are infrequent in Beckwith-Wiedemann syndrome. AB - Beckwith-Wiedemann syndrome (BWS) is an overgrowth disorder resulting from dysregulation of multiple imprinted genes through a variety of distinct mechanisms. A frequent alteration in BWS involves changes in the imprinting status of the coordinately regulated IGF2 and H19 genes on 11p15. Patients have been categorized according to alterations in the imprinted expression, allele specific methylation, and regional replication timing of these genes. In this work, IGF2/H19 expression, H19 DNA methylation, and IGF2 regional replication timing were studied in nine karyotypically normal BWS fibroblasts and two BWS patients with maternally inherited 11p15 chromosomal rearrangements. Informative patients (9/9) maintained normal monoallelic H19 expression/methylation, despite biallelic IGF2 expression in 6/9. Replication timing studies revealed no changes in the pattern of asynchronous replication timing for both a patient with biallelic IGF2 expression and a patient carrying an 11p15 inversion. In contrast, a patient with a chromosome 11;22 translocation and normal H19 expression/methylation exhibited partial loss of asynchrony and a shift toward earlier replication times. These results indicate that in BWS, (1) H19 imprinting alterations are less frequent than previously estimated, (2) IGF2 imprinting and H19 imprinting are not necessarily coordinated, and (3) alterations in regional replication timing are generally not correlated with either chromosomal rearrangements or the imprinting status of IGF2 and H19. PMID- 10857748 TI - Identification of a glialblastoma cell differentiation factor-related gene mRNA in human microvascular endothelial cells. AB - Vascular endothelial cells (VEC) transduce mitogenic and chemoattractant signals in response to erythropoietin (Epo). An analysis of changes in gene expression in VEC would be helpful to understanding the molecular nature of mitogenic signals. An effective method for analysis of gene expression is through differential display. Using this approach, we obtained from Epo-treated human microvascular endothelial cells (HMVEC) a cDNA fragment with characteristics of the 3'end of mRNA. Using the cDNA fragment, we then isolated a full-length clone from a HMVEC cDNA library. The cDNA of interest encodes a protein consisting of 404 amino acids with a carboxy-terminal end sequence identical to glialblastoma cell differentiation factor-related protein (GBDR1). Northern blot analysis showed that GBDR1 mRNA was ubiquitously expressed in human tissues. In Southern blot analysis, GBDR1 cDNA identified a single gene on chromosome 9. Since analysis of the amino acid sequence revealed several putative phosphorylation sites for different protein kinases, the GBDR1 protein was expressed and purified from bacterial extracts and, as predicted, casein kinase II phosphorylated GBDR1 in vitro. Immunofluorescence and biochemical data revealed that the GBDR1 protein is not entirely localized in the cytosolic fraction, suggesting that it may interact with another protein(s). These findings demonstrate that GBDR1 is an intracellular signaling molecule that may play a role in the regulation of endothelial cell growth. PMID- 10857749 TI - Comparative sequence analysis of the VHL tumor suppressor gene. AB - Comparative genome analysis may provide novel insights into gene evolution and function. To investigate the von Hippel-Lindau (VHL) disease tumor suppressor gene, we sequenced the VHL gene in seven primate species. Comparative analysis was performed for human, primate, and rodent VHL genes and for a putative Caenorhabditis elegans VHL homologue identified by database analysis. The VHL gene has two translation initiation sites (at codons 1 and 54); however, the relative importance of the full-length translation product (pVHL30) and that translated from the second internal translation initiation site (pVHL19) is unclear. The N-terminal sequence of pVHL30 contains eight copies of a GXEEX acidic repeat motif in human and higher primates, but only three copies were present in the marmoset, and only one copy was present in rodent VHL genes. Evolutionary analysis suggested that the N-terminal repetitive sequence in pVHL30 was of less functional importance than those regions present in both pVHL30 and pVHL19. The VHL gene product is reported to form complexes with various proteins including elongin B, elongin C, VBP-1, fibronectin, Spl, CUL2, and HIF-1. Although most of the regions in pVHL that had been implicated in binding specific proteins demonstrated evolutionary conservation, the carboxy-terminal putative VBP-1 binding site was less well conserved, suggesting that VBP-1 binding may have less functional significance. Although an amino acid substitution (K171T) close to the pVHL elongin binding region was found in baboon, analysis of the structure of human pVHL suggested that this substitution would not interfere with pVHL/elongin C interaction. In general, there was a good correlation between the pVHL domains that demonstrated most evolutionary conservation and those that were most frequently mutated in tumors. Analysis of human/C. elegans conservation and human germline and somatic mutation patterns identified a highly conserved mutation cluster region between codons 74 and 90. However, this region is likely to be important for the structural integrity of pVHL rather than representing an additional protein binding domain. PMID- 10857750 TI - Identification of two novel proteins that interact with germ-cell-specific RNA binding proteins DAZ and DAZL1. AB - The human DAZ (deleted in azoospermia) gene family on the Y chromosome and an autosomal DAZ-like gene, DAZL1, encode RNA-binding proteins that are expressed exclusively in germ cells. Their role in spermatogenesis is supported by their homology with a Drosophila male infertility gene boule and sterility of Daz11 knock-out mice. While all mammals contain a DAZL1 homologue on their autosomes, DAZ homologues are present only on the Y chromosomes of great apes and Old World monkeys. The DAZ and DAZL1 proteins differ in the copy numbers of a DAZ repeat and the C-terminal sequences. We studied the interaction of DAZ and DAZL1 with other proteins as an approach to investigate functional similarity between these two proteins. Using DAZ as bait in a yeast two-hybrid system, we isolated two DAZAP (DAZ-associated protein) genes. DAZAP1 encodes a novel RNA-binding protein that is expressed most abundantly in the testis, and DAZAP2 encodes a ubiquitously expressed protein with no recognizable functional motif. DAZAP1 and DAZAP2 bind similarly to both DAZ and DAZL1 through the DAZ repeats. The DAZAP genes were mapped to chromosomal regions 19p13.3 and 2q33-q34, respectively, where no genetic diseases affecting spermatogenesis are known to map. PMID- 10857751 TI - Cloning and characterization of HARP/SMARCAL1: a prokaryotic HepA-related SNF2 helicase protein from human and mouse. AB - The SNF2 gene family consists of a large group of proteins involved in transcriptional regulation, maintenance of chromosome integrity, and various aspects of DNA repair. We cloned a novel SNF2 family human cDNA, with sequence identity to the Escherichia coli RNA polymerase-binding protein HepA and named the human hepA-related protein (HHARP/SMARCAL1). In addition, the mouse ortholog (Mharp/Smarcal1) was cloned, and the Caenorhabditis elegans ortholog (CEHARP) was identified in the GenBank database. Phylogenetic analysis indicates that the HARP proteins share a high level of sequence similarity to the seven motif helicase core region (SNF2 domain) with identifiable orthologs in other eukaryotic species, except for yeast. Purified His-tagged HARP/SMARCAL1 protein exhibits single-stranded DNA-dependent ATPase activity, consistent with it being a member of the SNF2 family of proteins. Both the human and the mouse genes consist of 17 exons and 16 introns. The human gene maps to chromosome 2q34-q36, and the mouse gene is localized to the syntenic region of chromosome 1 (between markers Gls and Acrg). HARP/SMARCAL1 transcripts are ubiquitously expressed in human and mouse tissues, with testis presenting the highest levels of mRNA expression in humans. PMID- 10857752 TI - Molecular cloning and chromosomal mapping of a candidate cytokine gene selectively expressed in human CD34+ cells. AB - A rare population of human bone marrow (BM) and cord blood (CB) mononuclear cells bearing the CD34 surface marker (CD34+) function as hematopoietic stem/progenitor cells. Cells lacking CD34 expression (CD34-) in BM and CB are largely mature hematopoietic cells of various lineages that are derived from the CD34+ cells. To elucidate molecular mechanisms governing functional differences between CD34+ and CD34- hematopoietic cells, we used representational difference analysis (RDA) based subtraction to identify genes that are specifically or preferentially expressed in CD34+ cells. Among the 73 RDA fragments initially sequenced, 30% are derived from the CD34 and c-kit genes that are preferentially expressed in CD34+ cells. An additional 27 (37%) are novel or homologous only to entries in expressed sequence tag databases. One (C17) was found four times and is expressed in CD34+ but not in CD34- cell populations from CB or BM. The cloned C17 cDNA encodes a novel polypeptide of 136 amino acids with a signal sequence. No homology to this peptide was found in the public databases. A secondary-structure analysis predicts that the C17 peptide contains four alpha-helices, a characteristic of hematopoietic cytokines and interleukins. This novel gene is mapped to human chromosome 4p15-p16. PMID- 10857753 TI - Isolation and initial characterization of a novel zinc finger gene, DNMT3L, on 21q22.3, related to the cytosine-5-methyltransferase 3 gene family. AB - We have isolated the DNMT3L gene that is related to the cytosine-5 methyltransferase 3 (DNMT3) family. The gene is located on chromosome 21q22.3 between the AIRE and the KIAA0653 genes and spans approximately 16 kb of genomic sequence. The encoded protein of 387 amino acids has a cysteine-rich region containing a novel-type zinc finger domain that is conserved in DNMT3A and DNMT3B but also in ATRX, a member of the SNF2 protein family. The novel domain, called an ADD (ATRX, DNMT3, DNMT3L)-type zinc finger, contains two subparts: a C2C2 and an imperfect PHD zinc finger. Expression of the DNMT3L mRNA was not detectable by Northern blotting; however, RT-PCR amplification revealed that it is expressed at low levels in several tissues including testis, ovary, and thymus. PMID- 10857754 TI - NABC1 (BCAS1): alternative splicing and downregulation in colorectal tumors. AB - We have identified a new splicing variant of the gene "novel amplified in breast cancer 1," NABC1 (HGMW-approved symbol BCAS1). This variant, which we call NABC1_5B, uses a previously unidentified 135-bp exon. Also in this report, we confirm that NABC1 is overexpressed in breast tumors and show that both NABC1 and NABC1_5B are downregulated in colorectal tumors. PMID- 10857755 TI - Expression of interleukin 1 alpha and beta, and interleukin 1 receptor antagonist mRNA in the rat central nervous system after peripheral administration of lipopolysaccharides. AB - Interleukin 1alpha (IL-1alpha) and IL-1beta, and the endogenous IL-1 receptor antagonist (IL-1ra) are known members of the IL-1 family. Using in situ hybridization histochemistry we demonstrated that following endotoxin injection (lipopolysaccharides, LPS, 2.0 mg/kg, i.p.) a time dependent expression and partly different expression patterns of the cytokines occurred within the rat brain and pituitary gland. All cytokines were observed in the choroid plexus. In addition, IL-1ra mRNA expressing cells were observed scattered in the brain parenchyma, whereas scattered IL-1beta mRNA expressing cells were restricted to central thalamic nuclei, the dorsal hypothalamus, and cortical regions, such as the parietal and frontal cortex. A strong IL-1beta mRNA expression was found in the circumventricular organs. In the pituitary gland, a low IL-1alpha and a high IL-1beta mRNA expression was observed, with the highest density of cytokine expressing cells seen in the posterior pituitary. The cell types expressing the mRNA's of IL-1alpha, IL-1beta and IL-1ra were identified as monocytes in the circumventricular organs and the pituitary gland, and as microglia in the brain parenchyma. In conclusion, the present findings revealed that cytokine production in response to a peripheral endotoxin challenge mainly occurs in peripherally derived monocytes in the circumventricular organs and the pituitary gland. IL 1beta is the predominant form expressed, whereas the expression of IL-1alpha mRNA and IL-1ra mRNA is lower. Our observations support the view that peripherally derived IL-1 may play a role in the induction of centrally mediated illness symptoms. PMID- 10857756 TI - Interleukin 4 and 13 participation in mycobacterial (type-1) and schistosomal (type-2) antigen-elicited pulmonary granuloma formation: multiparameter analysis of cellular recruitment, chemokine expression and cytokine networks. AB - The contribution of IL-4 and IL-13 to inflammation and cytokine responses was compared in mice with types-1 or -2 pulmonary granulomas (GR) elicited by beads bound to antigens of Mycobacteria bovis (PPD) or Schistosoma mansoni eggs (SEA). Type-2 SEA-GR produced the most IL-4 and IL-13. Type-1 PPD-GR produced detectable IL-13, but not IL-4. Mice were treated with anti-IL4 or anti-IL-13 Abs, then lesion size/composition, cytokine/chemokine mRNA and lymph node cytokines were measured. Type-1 GRs resisted individual Abs, but combined Abs augmented lesions by 20%. In contrast, anti-IL-4 abrogated type-2 GR by 30-40% and eosinophil recruitment by 60%. Anti-IL-13 abrogated type-2 GR by 20-30% with no effect on eosinophils. Combined depletion reduced lesion area by 60% and eosinophils by more than 80%. In type-1 GR lungs, anti-IL-4 and anti-IL-13 augmented IFNgamma and TNFalpha mRNA. In type 2 lungs, anti-IL-13 did likewise, but anti-IL-4 decreased TNFalpha without affecting IFNgamma mRNA. In both responses, IL-4 promoted MCP-1 and MCP-5 mRNA, but IL-13 inhibited chemokines in type-1 GR. In lymph nodes, anti-IL-4, but not anti-IL-13, abrogated type-2 cytokines. In fact, IL-13 down-regulated itself and other type-2 cytokines. In summary, IL-4 and IL 13 have common and disparate regulatory functions in types 1 and 2 responses. PMID- 10857757 TI - The role of protein kinase C and calcium in induction of human polymorphonuclear leukocyte IL-1 beta gene expression by GM-CSF. AB - At infection sites, synthesis of interleukin (IL-)1beta by polymorphonuclear leukocytes (PMNs) facilitates the recruitment of inflammatory cells and enhances the inflammatory response. We investigated the role of protein kinase C (PKC) and Ca2+ in the induction of PMN IL-1beta gene expression by GM-CSF. The PKC inhibitors chelerythrine and H7 blocked induction of IL-1beta mRNA expression in human PMNs. HA1004, an H7 analogue with little activity towards PKC, had no inhibitory effect. Similarly, H7 blocked IL-1beta transcription in nuclear run-on analysis, while HA1004 had little effect. The intracellular Ca2+ chelator BAPTA/AM inhibited induction of IL1beta mRNA accumulation and transcription by GM CSF. At concentrations similar to those used to inhibit IL-1beta gene expression, H7, chelerythrine, and BAPTA all inhibited substrate phosphorylation by PKC isolated from PMN lysates. Thus, PKC and Ca2+ are potential targets for modulating an important PMN immunoregulatory function. PMID- 10857758 TI - Regulation of no synthesis induced by inflammatory mediators in RAW264.7 cells: collagen prevents inhibition by osteopontin. AB - Osteopontin has been shown to inhibit the induction of inducible nitric oxide synthase (iNOS, or NOS2) by lipopolysaccharide and interferon-gamma in the RAW264.7 mouse monocyte/macrophage line and in primary mouse proximal tubule epithelial cells. However, the RAW264.7 cells become refractory to the action of OPN after several subcultures or under dilute culture conditions, possibly because of changes in the composition of the extracellular matrix. We make this suggestion because if the cells are plated on a collagen type I or collagen type IV substrate the inhibitory action of OPN is completely suppressed; this is not the case on substrates consisting of laminin, fibronectin, poly-D-lysine, or poly (2-hydroxyethylmethylacrylate). These observations imply that macrophages are sensitive to regulation by OPN only in certain physiological contexts. Both hyaluronate, which binds CD44, and rat IgGs are also able to inhibit the induction of NO synthesis by the inflammatory mediators. The similar actions of HA and OPN are consistent with the possibility that CD44 may be a receptor for OPN. PMID- 10857759 TI - Reduction in serum IL-6 after vacination of breast cancer patients with tumour associated antigens is related to estrogen receptor status. AB - Elevated serum IL-6 concentrations have been associated with poor prognosis in a variety of cancers, and decreases in serum IL-6 concentrations have been reported after chemotherapy. We have demonstrated that serum IL-6 concentrations are elevated in breast cancer patients [normal women 0.7 +/- 2.5 pg/ml (n=36), breast cancer patients 38.3 +/- 138.7 pg/ml (n = 111)]. After vaccination of breast cancer patients with a combination of tumour-associated antigens and biological adjuvants (IL-2 and GM-CSF), the concentration of IL-6 decreased significantly (P<0.05) to 8.1 +/- 14.6 pg/ml (n=85). Other studies have shown that oestrogen suppresses IL-6 production in oestrogen receptor positive breast cancer cells. We have demonstrated that the decrease in IL-6 associated with vaccination is related to the oestrogen receptor status of the tumours from breast cancer patients, as a decrease in IL-6 from 124.0 +/- 267.5 pg/ml (n=26) to 6.2 +/- 11.0 pg/ml (n=34) only occurs in patients with oestrogen receptor negative tumours. The IL-6 concentration in breast cancer patients with oestrogen receptor positive tumours remained unchanged (9.5 pg/ml before vaccination, and 9.3 pg/ml after vaccination). These results suggest that postmenopausal women with oestrogen receptor negative breast cancers, who do not respond well to either hormonal therapy with tamoxifen or adjuvant chemotherapy, may have a significant response to vaccination with autologous tumour-associated antigens. PMID- 10857760 TI - Glycosaminoglycans alter the conformation of interferon-gamma. AB - Interferon-gamma (IFN-gamma) is a potent immunomodulatory cytokine whose physiological roles are modulated by the extracellular matrix. Here, circular dichroism and fluorescence spectroscopic techniques are used to demonstrate that low molecular weight heparin and chondroitin sulfate cause significant changes in secondary and tertiary structure of IFN-gamma. The results suggest that heparin and chondroitin sulfate modulate IFN-gamma activity by causing structural changes in the IFN-gamma dimer. PMID- 10857761 TI - Pilot study of IFN-gamma-induced specific hyposensitization for house dust mites in atopic dermatitis: IFN-gamma-induced immune deviation as a new therapeutic concept for atopic dermatitis. AB - IFN-gamma/IL-4 imbalance is a central immunologic defect which is responsible for increased IgE antibody response in atopic dermatitis. Effects of hyposensitization were controversial in atopic dermatitis. Reversed IFN-gamma/IL 4 balance was induced using IFN-gamma in atopic dermatitis and specific hyposensitization with house dust mites (HDM) was tried in the status of IFN gamma-induced immune deviation. A total of 58 atopic dermatitis patients who had obvious allergy to HDM were selected in this study. IFN-gamma-induced hyposensitization for HDM was tried in 10 patients. Twenty-two patients received IFN-gamma therapy and six were treated by simple hyposensitization. Twenty were enrolled as control subjects. The clinical severity scores decreased effectively only by IFN-gamma-induced hyposensitization for HDM. Specific hyposensitization for HDM in the status of IFN-gamma-induced immune deviation successfully improved atopic dermatitis. HDM might play a pathogenic role in subpopulation of atopic dermatitis. PMID- 10857762 TI - Differential glycosylation of interleukin 2, the molecular basis for the NOD Idd3 type 1 diabetes gene? AB - The insulin-dependent diabetes (Idd) gene, Idd3, has been localised to a 0.35 cM region of chromosome 3 containing the structural gene for the cytokine interleukin 2 (IL-2). While variation of the N-terminal amino acid sequence of IL 2 has been shown to correlate with Idd3 allelic variation, differences in induction of proliferation by IL-2 allotypes have not been detected. In the current study, we examined the electrophoretic migration of IL-2 allotypes and have found two distinct patterns, consistent with differences in glycosylation, that correlate with diabetes-resistance and susceptibility. These findings strongly suggest that IL-2 variants may be functionally distinct. PMID- 10857763 TI - Regulation of IL-10 secretion after phagocytosis of Mycobacterium tuberculosis by human monocytic cells. AB - Downregulation of pro-inflammatory events in the immune response to Mycobacterium tuberculosis is critical to prevent host tissue injury. Interleukin (IL-)10 is an important anti-inflammatory cytokine secreted in human tuberculosis but little is known about the control of such IL-10 release. Using an established cellular model, we measured IL-10 secretion after phagocytosis of M. tuberculosis. Phagocytosis of M. tuberculosis but not of inert latex beads by human monocytic (THP-1) cells resulted in IL-10 secretion maximal at 24 h. The magnitude and kinetics of IL-10 secretion were distinct from IL-10 secretion after phagocytosis of yeast-derived zymosan and depended on transcriptional activity and protein synthesis in infected monocytes. IL-10 secretion was decreased in a dose dependent manner by specific inhibitors of tyrosine kinases, protein kinase (PK) C and PKA. Inhibition of more than one pathway did not result in further synergistic or additive reduction in IL-10 secretion. Finally, specific neutralising antibody directed against IL-10 demonstrated that IL-10 secreted by infected monocytic cells did not block autologous IL-8 secretion. PMID- 10857764 TI - The role of ceramide in the modulation of intracellular Ca2+ levels by interleukin 1 beta in rat cortical synaptosomes. AB - We recently demonstrated that the pro-inflammatory cytokine, interleukin 1beta (IL-1beta) elevates intracellular free Ca2+ levels ([Ca2+]i) in rat cortical synaptosomes in a manner involving activation of the IL-1 receptor and stimulation of p42 mitogen-activated protein (MAP) kinase. We now report that the effects of IL-1beta on [Ca2+]i are mimicked by the sphingolipid metabolite ceramide. In cortical synaptosomes ceramide elevates [Ca2+]i in a p42 MAP kinase dependent manner, and we conclude that the effects of IL-1beta on Ca2+ homeostasis involve ceramide as an upstream component of the p42 MAP kinase pathway. PMID- 10857765 TI - Beta2-adrenoceptor agonist suppresses tumour necrosis factor production in rat mesangial cells. AB - This study aimed to investigate the time-course of the effect of beta2 adrenoceptor stimulation with terbutaline on lipopolysaccharide (LPS)-induced tumour necrosis factor(TNF)-alpha production in rat mesangial cells. Cells were cultured from 0-24 h in the presence of LPS (1 microg/ml) and/or terbutaline (10( 7)-10(-8) mol/l). After 1 h of incubation, terbutaline inhibited TNF-alpha protein release as well as transcription and translation of TNF-alpha and mitogen activated protein kinase (MAPK, p42/p44) activity. At 3 h, terbutaline enhanced intracellular cAMP but suppressed TNF-alpha release and transcription. By 24 h, whereas terbutaline was no longer influencing transcription or translation, TNF alpha release remained depressed which correlated with an increase in supernatant interleukin (IL)-6. Terbutaline did not affect the LPS-induced IL-10 produced in the cell. These findings indicate that beta2-adrenoceptor stimulation during an LPS challenge prevented TNF-alpha production as a consequence of MAPK inhibition and enhanced cAMP generation, which at a later stage was associated with an anti inflammatory effect of IL-6. PMID- 10857766 TI - Suppression of interleukin 2 biosynthesis by three modes of oxidative cellular stress: selective prevention by N-acetyl cysteine. AB - Acute stress induced by reagent hydrogen peroxide suppressed interleukin (IL-)2 biosynthesis in a dose-dependent fashion, reaching almost complete abolishment at 200 microM. Cells exposed to longitudinal oxidative stress and irradiation did not exhibit complete suppression of IL-2 biosynthesis, probably because intensities high enough to achieve such response would be lethal. These results suggest that suppression of IL-2 biosynthesis is a sensitive measure of acute oxidative stress. N-acetyl cysteine (NAC) prevented oxidative stress-induced suppression of IL-2 biosynthesis, except for that induced by acute stress at 100 microM and above. NAC was very efficient in preventing longitudinal and irradiation-induced stresses. Therefore, NAC appears to be a promising candidate for providing defence to individuals exposed to environmental conditions in which reactive oxygen intermediates are generated. PMID- 10857767 TI - Effect of cytokines and lipid mediators on the synthesis of interleukin 1 beta by human bone marrow stromal cells. AB - This study investigates the production of interleukin (IL-)1beta by cultured human bone marrow stromal cells. RT-PCR experiments indicate that two-thirds of cultures constitutively express IL-1beta mRNA transcripts. Their cell-associated IL-1beta levels are elevated after stimulation with tumour necrosis factor (TNF )alpha but not with cytokines such as IL-1alpha, IL-3, IL-4, IL-6, IL-7, IL-10, SCF, G-CSF, M-CSF and TGF-beta or lipid mediators such as PGE2, LTB4, LXA4, LXB4, 12-HETE, 15-HETE and PAF. Addition of IL-4, but not IL-10 or TGF-beta, reduces the TNF-alpha-induced cell-associated IL-1beta. IL-1beta is never detected in bone marrow stromal cell supernatants whatever the stimulant added. In conclusion the pro-inflammatory molecule TNF-alpha stimulates bone marrow stromal cell associated IL-1beta levels while the anti-inflammatory cytokine IL-4 reduces the TNF-alpha-induced effect. These results strengthen the key regulatory role of IL 4 on the production of haematopoietic cytokines by human bone marrow stromal cells. PMID- 10857769 TI - In vivo intracellular cytokine production by leukocytes during haemodialysis. AB - Several chronic inflammatory changes undergone during chronic haemodialysis are associated with increased pro-inflammatory cytokine production. Although generation of anaphylatoxins has been incriminated in the untoward effects of haemodialysis, it is still debated whether anaphylatoxins stimulate monocyte secretion of TNF-alpha and IL-1. We demonstrate that peripheral mononuclear cells isolated from healthy controls and cultured with complement-activated autologous serum or recombinant C5a induced high levels of IL-1, IL-1ra, IL-8 and MCP-1, low levels of TNFalpha and sTNFRII but no IL-10 and MIP-1alpha. Cytokine production by leukocytes was investigated by FACS analysis in six patients dialysed consecutively with three equivalent low permeability membranes known to activate the complement to different degrees: polysulfone (F6HPS), cellulose acetate (CA) and cuprophane (CP). Percentage of leukocytes expressing IL-1, IL-1ra, TNF-alpha and IL-8 is increased in patients dialysed with CP. Moreover, we show for the first time that haemodialysis is associated with the production of cytokines by circulating neutrophils. Predialysis plasma levels of MCP-1 and TNFRII did not increase during the dialysis session at the time when anaphylatoxin generation was highest. Dialysis with membranes that activate the complement to a high extent induce activation of leukocytes which may explain chronic complications associated with dialysing with CP. PMID- 10857768 TI - Lymphokine release from human lymphomononuclear cells after co-culture with isolated pancreatic islets: effects of islet species, long-term culture, and monocyte-macrophage cell removal. AB - This study evaluated the release of Th1 and Th2 cytokines from human lymphomononuclear cells (LMC) in response to purified human (HI) or bovine (BI) islets, and the role of long-term (3-4 weeks) islet culture and removal of monocyte-macrophage cells. The results showed that HI and BI caused a similar increase of the release of gamma interferon (IFN), IL-2 and IL-6 from LMC, whereas BI had a more marked effect than HI on IL-10 release. Culturing the islets had possible positive effects (reduction of IFN and IL-2), but also potentially negative effects (increase of TNF). Removal of monocyte-macrophage cells determined a significant reduction of IL-6, IL-10 and TNF production in response to xeno-islets. PMID- 10857770 TI - Interferon-alpha acutely impairs sleep in healthy humans. AB - We investigated the effects of two low doses of interferon-alpha (IFN-alpha) on nocturnal sleep in 18 healthy men by means of polysomnographic sleep recordings. At 1900h, human recombinant IFN-alpha (1000 or 10000 U/kg body weight) or placebo was administered subcutaneously. Between 2300h and 0700h subjects were allowed to sleep. In general effects were stronger at the dose of 10000 than 1000 U/kg body weight of IFN-alpha. Although, after IFN-alpha subjects experienced increased fatigue, the cytokine impaired the quality of nocturnal sleep. The higher dose of IFN-alpha suppressed slow wave sleep (17.8 +/- 2.0% vs 25.2 +/- 2.6% following placebo, P<0.003) but increased time spent in shallow sleep (P<0.05) during the first half of sleep time. Rapid eye movement (REM) sleep latency was postponed (P<0.02) and time spent in REM sleep was significantly decreased after IFN-alpha (P<0.04). The impairing influence of IFN-alpha on sleep in humans is in contrast with findings of sleep promoting effects of this cytokine in animals. Our data suggest that endogenous IFN-alpha may be a factor responsible for alterations of sleep, e.g. in the course of viral infections. PMID- 10857771 TI - Effect of cytokines and growth factors on the macrophage colony-stimulating factor secretion by human bone marrow stromal cells. AB - We have investigated the effect of growth factors, inflammatory and anti inflammatory cytokines on the macrophage colony-stimulating factor (M-CSF) secretion by cultured human bone marrow stromal cells. Their production of M-CSF cultured in serum-free medium is enhanced in a time-dependent manner in response to tumour necrosis factor (TNF-)alpha and interleukin (IL-)4 but not to IL-1, IL 3, IL-6, IL-7, IL-10, SCF, granulocyte-macrophage colony-stimulating factor (GM CSF), G-CSF, bFGF and transforming growth factor (TGF-)beta. The co-addition of IL-4 and TNF-alpha has a greater than additive effect on the secretion of M-CSF suggesting that they act synergistically. The anti-inflammatory molecules IL-10 and TGF-beta have no effect on the TNF-alpha-induced M-CSF synthesis by marrow stromal cells. In conclusion TNF-alpha and IL-4 are potent stimulators of the M CSF synthesis by human bone marrow stromal cells, a result of importance regarding the role of M-CSF in the proliferation/differentiation of mononuclear phagocytic cells and the role of marrow stromal cells as regulators of marrow haematopoiesis. PMID- 10857772 TI - Tumour necrosis factor-alpha up-regulates the expression of BMP-4 mRNA but inhibits chondrogenesis in mouse clonal chondrogenic EC cells, ATDC5. AB - Tumour necrosis factor (TNF)-alpha causes the degradation of articular cartilage in arthritis via direct actions on chondrocytes. However, it remains unknown whether TNF-alpha affects chondrogenesis in chondroprogenitors. In the present study, we assessed the effects of TNF-alpha in vitro on chondrogenesis using mouse clonal chondrogenic EC cells, ATDC5. TNF-alpha (10 ng/ml) stimulated [3H] thymidine incorporation in undifferentiated ATDC5 cells, and suppressed cartilaginous nodule formation and the accumulation of cartilage-specific proteoglycan. We recently showed that undifferentiated ATDC5 cells express BMP-4 and that exogenously administered BMP-4 promotes chondrogenesis in these cells. Interestingly, TNF-alpha up-regulated the expression of BMP-4 mRNA in undifferentiated ATDC5 cells in time- and dose-dependent manners. However, exogenously administered BMP-4 was not capable of reversing the inhibitory action of TNF-alpha on chondrogenesis in ATDC5 cells. These results indicate that TNF alpha stimulates both cell proliferation and BMP-4 expression but inhibits chondrogenesis in chondroprogenitor-like ATDC5 cells. PMID- 10857773 TI - The diminished postoperative capacity of blood leukocytes to produce IL-6 is associated with high concentrations of IL-6 in the circulation. AB - Surgical trauma is followed both by a transient increase of interleukin 6 (IL-6) concentrations in the serum and impaired function of circulating leukocytes. Perioperatively, we investigated the relationship of IL-6 concentrations in the serum with lipopolysaccharide (LPS)-stimulated cytokine production in the whole blood of patients undergoing elective major abdominal operations. In 50 patients, we found a transient increase of IL-6 concentrations in the serum. Six hours after skin incision, in vitro stimulated production of IL-6 and TNFalpha was diminished by 72% (P<0.05). A significant increase in cytokine production was observed three days postoperatively, however this was 63% below the preoperative values. Patients with high concentrations of circulating IL-6 showed a significantly lower stimulated IL-6 production than patients with low serum concentrations of IL-6. We conclude, that two opposing effects are associated with surgery: an activation leading to IL-6-release into the circulation, and a prolonged hyporesponsiveness of circulating leukocytes. These reactions are positively related. PMID- 10857774 TI - Immunoexpression of tumour necrosis factor-alpha and its receptors 1 and 2 correlates with proliferation/apoptosis equilibrium in normal, hyperplasic and carcinomatous human prostate. AB - Immunohistochemical and semiquantitative study of TNF-alpha, its receptors types 1 (TNFR1) and 2 (TNFR2), cell proliferation (Ki-67 nuclear antigen), and apoptosis (Tunel method) was carried out in human prostates, in normal healthy conditions, as well as in benign prostatic hyperplasia (BPH) and prostatic carcinoma (PC). Cell proliferation was higher in BPH than in normal prostates, and even higher in PC, mainly in neoformations showing a microglandular pattern. The apoptotic index was similar in BPH and normal prostates, and increased significantly in PC with independence of the pattern. In BPH, immunoreaction to TNF-alpha decreased as compared with that of normal prostates, while immunoreactions to both TNF-alpha receptors increased. This suggests a feedback downregulation of the factor, and that the low TNF-alpha activity in BPH are compensated by the increased amount of receptors. In PC, immunoreaction to TNF alpha and its two receptors increased markedly, suggesting that the TNF-induced effects are also increased. Contrarily to cell proliferation immunoexpression, PC reaction to TNFR2 was stronger in the papillar pattern than in the micrograndular pattern, and this suggests an inverse correlation between TNFR2 expression and cell proliferation. PMID- 10857775 TI - Cysteine protease production by human osteosarcoma cells (MG63, SAOS2) and its modulation by soluble factors. AB - The production of cysteine protease by two human osteosarcoma cell lines (MG-63 and SaOS2) was analyzed, as well as their modulation by interleukin 1beta (hIL-1 beta), interleukin 6 (hIL-6), insulin growth factor-1 (hIGF-1), oncostatin M (hOSM), leukemia inhibitory factor (hLIF) and growth hormone (hGH). Cysteine protease activities were detected using a synthetic substrate. The protease activities (especially cathepsin L activity) of both cell lines were increased significantly in the presence of hIL-1 beta, hIL-6 and hOSM. In contrast, hIGF-1 and hGH decreased these activities, and no effect was detectable in the presence of hLIF. The addition of antibodies against the gp-130 chain of the hIL-6 and hOSM receptors totally inhibited the stimulating effect of these two cytokines on cysteine protease activities. In increasing collagen type I degradation, hIL 1beta, hIL-6 and hOSM could be involved in bone resorption, whereas the inhibitory action of hIGF-1 and hGH on collagen type I degradation suggest that this factor could play a role in bone formation. PMID- 10857776 TI - Seasonal variation in interleukin 4 in patients with hayfever. AB - This study was performed to investigate the value of interleukin 4 as a marker of activity in mild atopic disease. We compared IL-4 levels to eosinophil cationic protein (ECP), a suggested inflammatory marker in allergic disease, in patients with hayfever. Patients with hayfever were assessed during January and then in late June at the height of the grass pollen season, and their levels of serum ECP and IL-4 compared. Serum ECP was determined by radio-immunoassay and serum IL-4 by a high-sensitivity enzyme-linked immunosorbent assay. ECP was found to increase significantly in patients with hayfever during the grass pollen season (P<0.01). Conversely, serum levels of IL-4 were found to decrease significantly over the same period when compared with winter values. ECP and IL-4 were not seen to correlate significantly with each other. The fall in serum IL-4 seen during the grass pollen season in the hayfever patients may reflect allergen driven upregulation of membrane IL-4 receptor expression or sequestration of cytokine producing cells to inflammatory sites. These findings suggest that serum IL-4 is a poor indicator of inflammatory status in allergic disease. PMID- 10857777 TI - Contributions of eighteenth and nineteenth century French medicine to colon and rectal surgery. PMID- 10857778 TI - Intraarticular injection of hyaluronan as treatment for knee osteoarthritis: what is the evidence? PMID- 10857779 TI - Natural killer cells and natural killer T cells. AB - NK cells are important in protecting against viral infections, and they may regulate the immune response. They are activated by hematopoietic blasts and pose a barrier to bone marrow transplantation. They are also abundant in the pregnant uterine decidua, although their role there is unknown. NK cells are normally inhibited from responding to host cells by inhibitory receptors that recognize self class I MHC antigens. There is evidence that NK cells may be important in the regulation of autoimmunity, but there is even stronger evidence that NKT cells regulate autoimmunity. The mechanisms by which these cells are activated and by which they regulate other cells are now being understood at the molecular level. PMID- 10857780 TI - Lytic Epstein-Barr virus infection in the synovial tissue of patients with rheumatoid arthritis. AB - OBJECTIVE: To evaluate the existence of Epstein-Barr virus (EBV) infection in the synovial tissue of patients with rheumatoid arthritis (RA). METHODS: Synovial tissues were obtained at synovectomy or arthroplasty from 32 patients with RA and 30 control patients with osteoarthritis (OA). EBV DNA was detected by Southern blot hybridization and/or polymerase chain reaction (PCR) amplification. To localize the EBV-infected cells, tissue sections were studied by RNA in situ hybridization (ISH) for the EBV-encoded small RNA 1 (EBER-1), by DNA ISH for the Bam HI W region of EBV DNA, and by immunohistochemistry for EBV lytic proteins BZLF1 and gp350/220. RESULTS: EBV DNA was detected by PCR in 15 of the 32 samples from RA patients (47%), but in none of those from the 30 OA patients (P < 0.01). Of the 15 PCR-positive samples, 9 contained >1 EBV copy/1,000 cells (referred to as EBV 2+), and 6 contained 1 copy/1,000-5,000 cells (EBV 1+). Among the 9 EBV 2+ samples, 3 were also positive for EBV DNA by Southern blot hybridization, 5 were positive for EBER-1 by RNA ISH, and 3 were positive for EBV DNA by DNA ISH. Immunohistochemical analysis showed positive signals in all samples for BZLF1 and in 7 samples for gp350/ 220. In each examination, the positive signals were detected not only in lymphocytes, but also in synovial lining cells. CONCLUSION: EBV was frequently detected in the synovial tissue of RA patients. The infected cells were both lymphocytes and synovial cells, and expressed EBV proteins associated with virus replication. These findings suggest that EBV may play a role in the pathogenesis of RA. PMID- 10857781 TI - Differential expression pattern of membrane-type matrix metalloproteinases in rheumatoid arthritis. AB - OBJECTIVE: To study the expression of messenger RNA (mRNA) for different membrane type matrix metalloproteinases (MT-MMPs) and compare their expression pattern in rheumatoid arthritis (RA) and normal synovium. METHODS: Polymerase chain reaction (PCR) with specific primers was performed to analyze the presence of MT1-, MT2-, MT3-, and MT4-MMP in synovial tissue and synovial fibroblasts from 10 patients with RA and 4 subjects without arthritis. In addition, in situ hybridization with digoxigenin-labeled RNA probes was used to investigate the expression pattern of MT-MMPs in the synovium of these subjects. MT-MMP-expressing cells were characterized by immunohistochemical double labeling with anti-CD68 monoclonal antibodies. RESULTS: Reverse transcription-PCR revealed the expression of MT1-, MT2-, MT3-, and MT4-MMP mRNA in all tissues and cell cultures examined. However, in situ hybridization showed considerable differences in the expression pattern of the different MT-MMPs in RA synovium. MT1- and MT3-MMP mRNA were highly expressed in both the lining and the sublining layer, with more intense staining in the lining. Immunohistochemical double labeling demonstrated the presence of mRNA for MT1-MMP in fibroblasts and macrophages, as well as in osteoclast-like cells at sites of bone resorption. Expression of MT3-MMP mRNA was seen in fibroblasts and some macrophages. Expression of MT2- and MT4-MMP was characterized by staining of only a few CD68-negative fibroblasts, and no differences could be found between the lining and sublining. Normal synovial samples showed only limited staining for all MT-MMPs. CONCLUSION: Our results indicate a role for MT1-MMP not only in the matrix degradation by fibroblasts, but also in osteoclast-mediated bone resorption in RA. Given the ability of MT1 MMP to activate MMP-2 and MMP-13, the findings also point to a cooperation between fibroblasts and macrophages in degrading cartilage and bone. While MT3 MMP is also intensely expressed in RA synovium, MT2- and MT4-MMP appear not to be involved in rheumatoid joint destruction. PMID- 10857782 TI - Human cartilage gp-39+,CD16+ monocytes in peripheral blood and synovium: correlation with joint destruction in rheumatoid arthritis. AB - OBJECTIVE: To investigate the expression of human cartilage (HC) gp-39, a possible autoantigen in rheumatoid arthritis (RA), in peripheral blood and synovium, to characterize its cellular source, and to analyze correlations with clinical features. METHODS: The expression of HC gp-39 in synovium and peripheral blood mononuclear cells (PBMC) was assessed by immunohistochemistry and flow cytometry. Synthesis and secretion were investigated by both reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. RESULTS: PBMC expressing HC gp-39 were increased in RA patients compared with spondylarthropathy patients (P = 0.0029) and with healthy control subjects (P = 0.0013). HC gp-39+ cells were also slightly overrepresented in RA synovium (P = 0.01). In both blood and synovium, HC gp-39+ cells were identified as CD14dim,CD16+ monocytes, a phenotype which can differentiate from classic CD14++ monocytes by maturation in vitro. HC gp-39 messenger RNA was detected in RA synovium and PBMC, and PBMC secreted HC gp-39 in vitro. The number of HC gp-39+ PBMC correlated with serum levels of C-reactive protein (r = 0.39, P = 0.003) and HC gp-39 (r = 0.52, P = 0.014). HC gp-39 expression in RA synovial lining correlated with joint destruction (r = 0.77, P < 0.001). CONCLUSION: CD16+ monocytes, a cellular source of HC gp-39 in vivo, are overrepresented in both RA peripheral blood and synovial tissue. The presence of HC gp-39+ cells in RA synovium is correlated with the degree of joint destruction. These data support a role of these cells in the local autoimmune response that leads to chronic inflammation and joint destruction. PMID- 10857783 TI - Macrophage-derived cytokine and nuclear factor kappaB p65 expression in synovial membrane and skin of patients with psoriatic arthritis. AB - OBJECTIVE: Monocyte-derived cytokines are important mediators in synovitis and represent novel therapeutic targets. This study was undertaken to analyze their expression in synovial membrane (SM) of patients with psoriatic arthritis (PsA) compared with that in skin of patients with PsA and SM of patients with rheumatoid arthritis (RA). METHODS: Multiple synovial biopsy samples (24 from patients with PsA, 20 from patients with RA, 5 from patients with osteoarthritis [OA]) and skin biopsy samples (lesional and perilesional skin from 25 PsA patients) were obtained. Standard leukocyte antigens, cytokines (tumor necrosis factor alpha [TNFalpha], interleukin-1apha [IL-1alpha], IL-1beta, IL-15, and IL 10) and the transcription factor nuclear factor KB (NF-kappaB; active p65 subunit) were localized and quantified immunohistochemically by light microscopy and digital image analysis. RESULTS: Sublining cellular infiltration, lymphoid aggregation, and vascularity were similar in PsA and RA SM. Lining layer thickness was greater in RA SM, associated with more CD68+ macrophages. In PsA SM, TNFalpha, IL-1alpha, IL-1beta, IL-15, and IL-10 were primarily localized to lining layer and perivascular macrophages, as were cells expressing the active subunit of NF-kappaB (p65). TNFalpha, IL-1p, and IL-15 expression in PsA lining layer was less than that in RA lining layer, likely reflecting lower macrophage numbers. In sublining areas, levels of TNFalpha and IL-15 were lower in PsA patients than in RA patients, whereas IL-lalpha and IL-1beta expression was equivalent. IL-10 was identified at similar levels in RA and PsA SM lining layer and sublining. Expression of NF-kappaB (p65) was equal in lining layer from both patient groups, but lower in PsA than RA sublining. Histologic findings did not correlate with clinical parameters of disease. Cytokine expression in skin did not correlate directly with that in SM. Cytokine expression was greater in PsA and RA SM than in OA SM. CONCLUSION: This study shows, for the first time, that monocyte-derived cytokines are found in PsA SM and demonstrates the relative paucity of the antiinflammatory cytokine IL-10 in PsA skin and SM. Significant divergence from RA SM expression was observed, despite similar clinical and demographic features in the 2 patient groups. PMID- 10857784 TI - Dysregulated intracellular Ca2+ stores and Ca2+ signaling in synovial fluid T lymphocytes from patients with chronic inflammatory arthritis. AB - OBJECTIVE: Peripheral blood (PB) T cells from rheumatoid arthritis (RA) patients proliferate poorly to mitogen, a change that is related to decreased intracellular Ca2+ ([Ca2+]i) signaling after T cell receptor (TCR) stimulation. We hypothesized that this was, in part, due to the effect of mediators of inflammation and predicted that greater changes in [Ca2+]i signaling would be seen in synovial fluid (SF) T cells. We also examined the mechanisms underlying the altered [Ca2+]i signals. METHODS: Paired PB and SF T cells from patients with chronic inflammatory arthritis were stimulated with mitogen to assess the magnitude of the [Ca2+]i signal in cell populations by fluorometry, the pattern of the [Ca2+]i signal in individual cells in a single-cell ion-imaging system, and the spatial distribution of Ca2+ within intracellular organelles. RESULTS: There was a significantly smaller [Ca2+]i signal after phytohemagglutinin protein stimulation of SF T cells (peak rise in [Ca2+]i signal PB versus SF 200 nM versus 180 nM; P < 0.05). In single SF T cells, a change in the pattern of the [Ca2+]i signal and a reduction in the number of responding cells was seen. These changes were a magnification of those seen in RA PB compared with control PB T cells. The contribution of Ca2+ release from intracellular stores to the final [Ca2+]i signal in PB and SF T cells was equal, but there was a significant increase in the Ca2+ remaining in the endoplasmic reticulum (ER) in SF T cells after TCR activation (PB versus SF 6 nM versus 19 nM; P < 0.05). Non-ER Ca2+ stores were not similarly affected. CONCLUSION: We found abnormalities in the magnitude, pattern, and spatial distribution of [Ca2+]i signaling in T cells from SF of patients with chronic inflammatory arthritis. A reduction in the number of responding SF T cells may partly explain some of our observations. However, we propose that the observed redistribution of SF Ca2+ stores may underlie the altered [Ca2+]i signaling, thus making these cells hyporesponsive to mitogen. The inflammatory environment of the joint and the late stage of differentiation of SF T cells are both likely to contribute to these changes in [Ca2+]i signaling, resulting in aberrant T cell function and promotion of disease chronicity. PMID- 10857785 TI - Temporal expression of inflammatory cytokines and chemokines in rat adjuvant induced arthritis. AB - OBJECTIVE: To examine cytokine and chemokine production during the evolution of rat adjuvant-induced arthritis (AIA), a model of rheumatoid arthritis. METHODS: Clinical and laboratory assessment of the course of AIA was performed over a 47 day period. Levels of the cytokines tumor necrosis factor a (TNFalpha), interleukin-1beta (IL-1beta), and IL-6, as well as levels of the chemokines macrophage inflammatory protein 1alpha (MIP-1alpha) and JE, the murine homolog of monocyte chemoattractant protein 1, were determined by enzyme-linked immunosorbent assay in the sera and joints of AIA and control rats. Synovia from AIA rats were (immuno)histochemically analyzed. Results of cytokine and chemokine measurements were correlated with clinical and laboratory markers of inflammation and histology. RESULTS: Early (before day 14 post adjuvant injection) and later phases of AIA could be distinguished. Cytokine and chemokine production was increased in AIA versus control rats. The production of TNFalpha, IL-1beta, MIP 1alpha, and, as determined earlier, epithelial neutrophil-activating peptide 78 like protein was abundant prior to and during the course of AIA, while that of IL 6 and JE was elevated in the late phase of AIA. Cytokine and chemokine levels were correlated with the clinical symptoms of arthritis and blood neutrophil counts. Joint levels of IL-1beta showed correlation with synovial lining proliferation and neutrophil ingress into AIA synovium. CONCLUSION: Cytokines and chemokines are involved in the clinical, laboratory, and histologic changes underlying AIA. The production of these mediators may be temporally and spatially regulated. These findings may be important for the optimal timing of cytokine and chemokine targeting. PMID- 10857786 TI - Identification of four new quantitative trait loci regulating arthritis severity and one new quantitative trait locus regulating autoantibody production in rats with collagen-induced arthritis. AB - OBJECTIVE: Collagen-induced arthritis (CIA) is a polygenic model of experimentally induced autoimmunity and chronic joint inflammation. This study maps genetic loci that regulate CIA susceptibility in DA/Bkl (DA) and BN/SsNHsd (BN) rats. METHODS: Genome scans covering chromosomes 1-20 and interval mapping techniques using 159 simple sequence-length polymorphism markers were used to identify quantitative trait loci (QTLs) that regulate CIA in (DA x BN)F2 hybrids. Serum antibody titers to type II collagen were determined by enzyme-linked immunosorbent assay. RESULTS: DA rats were high responders to porcine type II collagen (PII) and developed severe CIA (100%). BN rats were low responders to PII and resistant to CIA (0%). BN genes strongly repressed PII-induced CIA. Only 12% of (DA x BN)F1 rats (7 of 60) and 31% of (DA x BN)F2 rats (307 of 1,004) developed CIA. Three new QTLs (Cia11, Cia12, and Cia13) with significant logarithm of odds (LOD) scores of 5.6, 4.6, and 4.5, respectively, plus a suggestive QTL (Cia14*, LOD 3.0) regulating arthritis severity were identified on chromosomes 3, 12, 4, and 19. A new QTL, Ciaa3, associating with anticollagen antibody titer (antibody to PII LOD 6.5; antibody to rat type II collagen LOD 5.2) mapped to chromosome 9. Of 10 CIA QTLs previously identified in (DA x F344) and (DA x ACI) rats, only Cia1 in the major histocompatibility complex and a region coincident to Cia5 on chromosome 10 (LOD >8.0) influenced CIA severity in (DA x BN)F2 rats. CONCLUSION: Since CIA exhibits many of the pathologic features of rheumatoid arthritis, the data indicate that the variety of genetic elements regulating human autoimmune and rheumatic diseases may be much larger and more varied than originally envisioned. PMID- 10857787 TI - Selective inhibition of inducible nitric oxide synthase reduces progression of experimental osteoarthritis in vivo: possible link with the reduction in chondrocyte apoptosis and caspase 3 level. AB - OBJECTIVE: To evaluate the in vivo therapeutic efficacy of N-iminoethyl-L-lysine (L-NIL), a selective inhibitor of inducible nitric oxide synthase, on the progression of structural lesions in the experimental canine model of osteoarthritis (OA), and to explore the effect of L-NIL on the level of chondrocyte apoptosis and of important proteins involved in the apoptotic phenomenon, i.e., caspase 3 (inducer) and Bcl-2 (inhibitor). METHODS: The OA model was created by sectioning the anterior cruciate ligament. Dogs were placed into 4 experimental groups: unoperated dogs that received no treatment (controls), operated (OA) dogs that received placebo treatment, OA dogs that received oral L-NIL at 10 mg/kg/day, and OA dogs that received oral L-NIL at 1.0 mg/kg/day. In both L-NIL groups, treatment started immediately after surgery. The OA dogs were killed at 12 weeks after surgery. RESULTS: OA dogs treated with L NIL showed a reduction in the size of osteophytes and a significant decrease in the severity of macroscopic and histologic cartilage lesions on both condyles and plateaus, compared with untreated OA dogs. L-NIL treatment also significantly decreased metalloprotease activity in cartilage. Immunohistochemical analysis revealed that the levels of chondrocyte apoptosis, caspase 3, and Bcl-2 were markedly increased in OA cartilage (P < 0.0001). A positive correlation between the levels of chondrocyte apoptosis and levels of caspase 3 was found (r = 0.54, P < 0.0001). OA dogs treated with the higher dosage L-NIL showed significantly reduced levels of chondrocyte apoptosis (P < 0.003) and caspase 3 (P < 0.04), but no effect on the increased level of Bcl-2 was demonstrated. CONCLUSION: This study shows that L-NIL reduces the progression of experimental OA. This effect could be related to a reduced level of chondrocyte apoptosis and is likely mediated by a decrease in the level of caspase 3 activity. A sparing effect of L NIL on the increased level of Bcl-2 may also be a contributing factor. PMID- 10857788 TI - Reduction of interleukin-17-induced inhibition of chondrocyte proteoglycan synthesis in intact murine articular cartilage by interleukin-4. AB - OBJECTIVE: To investigate the role of interleukin-4 (IL-4) and IL-10 in basal and IL-1- and IL-17-mediated inhibition of chondrocyte metabolism. METHODS: Cartilage explants of patellae from naive mice were incubated with IL-17 and/or IL-1 alone or were pretreated with IL-4 and IL-10. In addition, knee joints of naive mice were injected intraarticularly with IL-4 and IL-10 alone or were coinjected with IL-1. Chondrocyte proteoglycan (PG) synthesis was measured in intact murine articular cartilage. Levels of nitric oxide (NO) were measured using the Griess reagent. RESULTS: IL-17, a novel cytokine secreted by CD4+ activated memory T cells, inhibited chondrocyte PG synthesis in intact murine articular cartilage, although the suppressive effect was less potent than that of IL-1. The suppressive effect of IL-17 was completely abolished in the presence of L-NIO (L NS-[1-iminoethyl]ornithine), an inhibitor of NO metabolism, and IL-17 only slightly induced inhibition of PG synthesis in cartilage explants of patellae from iNOS (inducible NO synthase) knockout mice. This indicates that the suppressive effect of IL-17 was mediated by NO. Pretreatment with IL-4, but not IL-10, significantly reduced the inhibition of chondrocyte PG synthesis induced by IL-1 or IL-17. The IL-4-induced reduction in the inhibitory effects of IL-1 and IL-17 on chondrocyte PG synthesis was accompanied by decreased NO formation in the culture supernatants. CONCLUSION: IL-17 plays a role in the inhibition of chondrocyte PG synthesis. IL-4 and IL-10 have no effect on basal chondrocyte metabolism. However, IL-4-pretreated cartilage is less sensitive to the suppressive effect of IL-1 as well as IL-17. This suggests that IL-4 is protective in T cell-driven cartilage disturbances, probably through reduction of iNOS. PMID- 10857789 TI - Profile of glycosaminoglycan-degrading glycosidases and glycoside sulfatases secreted by human articular chondrocytes in homeostasis and inflammation. AB - OBJECTIVE: To determine enzymatic activities of the 8 key glycosaminoglycan degrading glycosidases and glycoside sulfatases in cultured human articular chondrocytes and in synovial fluid from patients with osteoarthritis. METHODS: The following enzymes were analyzed: hexosaminidase and its isoenzyme A, N-acetyl alpha-D-glucosaminidase, beta-galactosidase, beta-glucuronidase, alpha-L iduronidase, aryl sulfatase, and galactose-6-sulfate sulfatase. Activity of the selected enzymes was analyzed by fluorometry with the aid of 4 methylumbelliferryl derivatives of the appropriate monosaccharides. RESULTS: Hexosaminidase was found to be the dominant enzyme released by chondrocytes into the extracellular compartment. Stimulation of chondrocytes with interleukin-1beta resulted in a selective increase of the extracellular hexosaminidase activity and, to a lesser degree, of the extracellular beta-galactosidase activity, without significant changes in the activity of the other studied enzymes. Analysis of the pH dependency of the enzymatic activities revealed that even at neutral pH, hexosaminidase expressed a measurable activity, much higher than the activity of the other studied enzymes. Chondrocyte apoptosis did not result in increased extracellular glycosidase activities, including hexosaminidase activity. The spectrum of glycosidase and glycoside sulfatase activities in the synovial fluid from patients with osteoarthritis was similar to that in cultured human articular chondrocytes. CONCLUSION: These data support the concept that lysosomal glycosidases, in particular hexosaminidase, represent a distinct subset of cartilage matrix-degrading enzymes that are activated by proinflammatory stimuli. PMID- 10857790 TI - Stimulation of hyaluronan metabolism by interleukin-1alpha in human articular cartilage. AB - OBJECTIVE: To determine the effects of interleukin-1alpha (IL-1alpha) on the expression of hyaluronan synthase (HAS), CD44, and aggrecan in human articular chondrocytes, and to assess the net result of these metabolic changes on the accumulation of hyaluronan within articular cartilage. METHODS: Normal human articular cartilage slices, as well as isolated chondrocytes, were treated with IL-1alpha. Changes in the relative expression of messenger RNA (mRNA) for HAS-2, CD44, and aggrecan were determined by competitive, quantitative reverse transcriptase-polymerase chain reaction. Hyaluronan accumulation was characterized by staining with a hyaluronan-specific binding protein and by fluorophore-assisted carbohydrate electrophoresis, while proteoglycan content was determined by alcian blue and Safranin O staining, CD44 protein expression by immunohistochemistry, and aggrecan biosynthesis by 35S-sulfate incorporation. Changes in cell-associated matrix sizes were visualized by a particle exclusion assay. RESULTS: IL-1alpha stimulated the expression of HAS-2 and CD44 mRNA (3.5 fold and 3-fold, respectively), but inhibited the expression of aggrecan mRNA. In IL-1-treated chondrocytes, extracellular hyaluronan decreased, while intracellular accumulation of hyaluronan was enhanced. Together with the decrease in expression of aggrecan, a dramatic reduction in cell-associated matrix was observed. IL-1-treated cartilage slices displayed a prominent depletion of aggrecan as well as hyaluronan within the upper layers of the tissue. The regional loss of hyaluronan coincided with a regional up-regulation of CD44. CONCLUSION: These data demonstrate that IL-1alpha stimulates HAS-2 at the same time as it inhibits the expression of aggrecan. Although hyaluronan biosynthesis is up-regulated, so too is the expression of CD44 and the internalization/catabolism of hyaluronan. The net result is a loss of hyaluronan in areas of the articular cartilage where increases in CD44 expression are most prominent. This depletion of hyaluronan in the upper layers of the tissue likely facilitates the prominent loss of aggrecan from the tissue. PMID- 10857791 TI - Small nucleolar RNP scleroderma autoantigens associate with phosphorylated serine/arginine splicing factors during apoptosis. AB - OBJECTIVE: Proteins that are phosphorylated during apoptosis are commonly precipitated by autoantibodies found in the sera of patients with systemic lupus erythematosus. We sought to determine whether scleroderma autoantigens such as small nucleolar RNPs (snoRNP) also associate with phosphoproteins in response to various cellular stressors. METHODS: We screened a panel of monoclonal antibodies derived from mice exposed to mercury, a well-characterized murine model of the anti-snoRNP autoimmune response, for the ability to selectively precipitate phosphoproteins from radiolabeled lysates prepared from Jurkat T cells subjected to stressful stimuli. RESULTS: Monoclonal antibodies reactive with snoRNPs precipitated a phosphoprotein complex (pp42, pp34, and pp23) from lysates prepared from apoptotic cells. Several novel phosphoproteins (pp62 and pp18) were also observed. The phosphorylation and/or recruitment of these proteins to the snoRNP complex is induced by multiple apoptotic stimuli (e.g., Fas ligation, anisomycin, or ultraviolet irradiation), an effect that is blocked by overexpression of Bcl-2. We were unable to demonstrate an association of the phosphoprotein complex with snoRNPs in cells treated with the xenobiotic agent mercury. The snoRNP-associated phosphoprotein complex is composed of serine/arginine (SR) splicing factors, including SRp40. CONCLUSION: The association of phosphorylated SR proteins with snoRNPs in cells undergoing apoptosis suggests that the immune response to fibrillarin that characterizes a subset of patients with scleroderma may be related to cell death induced by apoptotic stimuli (e.g., Fas ligation, irradiation, or chemical toxins), or by exposure to mercury. PMID- 10857792 TI - Treatment of rolling neutrophils with antineutrophil cytoplasmic antibodies causes conversion to firm integrin-mediated adhesion. AB - OBJECTIVE: The vascular lesions associated with autoimmune small-vessel vasculitis may arise from activation of circulating neutrophils by antineutrophil cytoplasmic antibodies (ANCA), resulting in increased adhesion of these neutrophils to the vessel wall. The present study examined the effects of ANCA positive IgG (ANCA IgG), derived from patients with small-vessel vasculitis, on neutrophil adhesion. METHODS: An in vitro, flow-based adhesion assay was used to determine the effects of ANCA IgG on neutrophils rolling on P-selectin presented by a monolayer of activated platelets. The platelets act as a surrogate vessel wall and can also support beta2 integrin-mediated immobilization of neutrophils if they are purposefully activated (e.g., by FMLP). RESULTS: In the absence of any added agents, neutrophils rolled continuously over the platelet monolayer. Superfusion of ANCA IgG over rolling cells resulted in conversion to stationary adhesion accompanied by shape change. The ANCA-mediated response was transient, peaking at 5-6 minutes and returning to baseline by 15 minutes, even in the continued presence of ANCA. In contrast, normal (ANCA-negative) IgG and ANCA F(ab')2 fragments caused minimal conversion to stationary adhesion. Pretreatment of neutrophils with blocking antibodies directed toward Fc gamma receptor type IIA or the integrin chain CD11b completely inhibited the ANCA-mediated conversion, confirming that ANCA-mediated activation occurred through Fc gamma receptors and that neutrophil immobilization was mediated by the activated beta2 integrin (CD11b/CD18). CONCLUSION: These findings support the concept that ANCA can directly activate neutrophils to become firmly adherent to vessel walls, where they may obstruct flow, initiate tissue damage, and contribute to pathogenesis of vasculitis. PMID- 10857793 TI - Successful treatment of active ankylosing spondylitis with the anti-tumor necrosis factor alpha monoclonal antibody infliximab. AB - OBJECTIVE: Tumor necrosis factor alpha (TNFalpha) has been detected in sacroiliac joint biopsy specimens from patients with spondylarthropathy. The present open pilot study was undertaken to test the efficacy of the anti-TNFalpha monoclonal antibody infliximab in the treatment of active ankylosing spondylitis (AS). METHODS: Eleven patients with AS of short duration (median 5 years, range 0.5-13 years) that had been active for at least 3 months (range 3-72 months) were treated with 3 infusions of infliximab (at weeks 0, 2, and 6), in a dosage of 5 mg/kg. Ten of the 11 patients had elevated C-reactive protein (CRP) levels (>6 mg/liter) before treatment; these elevations were known to have had persisted > 1 year in at least 3 patients. The Bath AS Disease Activity Index (BASDAI), the Bath AS Functional Index (BASFI), pain as measured on a visual analog scale, and the Bath AS Metrology Index (BASMI) were assessed. Quality of life was assessed using the Short Form 36 instrument. Laboratory markers of disease activity, including interleukin-6 (IL-6) levels, were determined. Dynamic magnetic resonance imaging (MRI) of the spine was performed in 5 patients. RESULTS: One patient withdrew from the study due to the occurrence of urticarial xanthoma 8 days after the first infusion. At study enrollment, 3 of 5 patients had evidence of spinal inflammation (spondylitis and spondylodiscitis) as detected by MRI; followup MRI 2-6 weeks after the third infusion revealed improvement in 2. Improvement of > or = 50% in activity, function, and pain scores was documented in 9 of 10 patients; the median improvement in the BASDAI after 4 weeks was 70% (range 41-94%). This clear-cut benefit lasted for 6 weeks after the third infusion in 8 of 10 patients. The median CRP level decreased from 15.5 mg/liter (range <6-90.8) to normal, and the median IL-6 level from 12.4 mg/liter (range 0 28.4) to normal (<5). There was improvement in all 9 SF-36 concepts; the improvement was significant for 6 concepts. CONCLUSION: These data suggest that anti-TNFalpha therapy is very effective for several weeks in AS. Whether this therapy, in addition to its antiinflammatory effect, prevents ankylosis remains to be determined. PMID- 10857794 TI - The X-chromosome and susceptibility to ankylosing spondylitis. AB - OBJECTIVE: Ankylosing spondylitis (AS) affects 0.25-1.0% of the population, and its etiology is incompletely understood. Susceptibility to this highly familial disease (lambda(s) = 58) is primarily genetically determined. There is a significant sex bias in AS, and there are differences in recurrence risk to the offspring of affected mothers and fathers, suggesting that there may be an X linked recessive effect. We undertook an X-chromosome linkage study to determine any contribution of the X-chromosome to AS susceptibility. METHODS: A linkage study of the X-chromosome using 234 affected sibling pairs was performed to investigate this hypothesis. RESULTS: No linkage of the X-chromosome with susceptibility to AS was found. Model-free multipoint linkage analysis strongly excluded any significant genetic contribution (lambda > or = 1.5) to AS susceptibility encoded on the X-chromosome (logarithm of odds [LOD] <-2.0). Smaller genetic effects (lambda > or = 1.3) were also found to be unlikely (LOD < 1.0). CONCLUSION: The sex bias in AS is not explained by X-chromosome-encoded genetic effects. The disease model best explaining the sex bias in occurrence and transmission of AS is a polygenic model with a higher susceptibility threshold in females. PMID- 10857795 TI - The familial form of spondylarthropathy: a clinical study of 115 multiplex families. Groupe Francais d'Etude Genetique des Spondylarthropathies. AB - OBJECTIVE: To investigate the interrelationships among different phenotypes, and their relationship to the HLA-Blocus, in multiplex families with spondylarthropathy (SpA). METHODS: We recruited 115 white French families, each of which had at least 2 members with SpA. Pedigrees were established. Clinical data and pelvic radiographs were collected. The HLA-B27 status of all patients was determined. Analysis was performed to determine the prevalence of SpA manifestations according to sex, disease duration, and HLA-B status, and to examine clustering of specific manifestations in subsets of families. RESULTS: We identified 329 SpA patients. Mean +/-SD age at onset was 24+/-9.4 years. The male:female ratio was 186:143, or 1.3, with few sex differences in disease expression. Axial manifestations and HLA-B27 were each present in 97% of the patients. Inflammatory bowel disease and HLA-B35 were overrepresented in the 7 families containing HLA-B27-negative patients. The frequency of radiographic sacroiliitis increased in parallel with disease duration. Peripheral enthesitis, radiographic sacroiliitis, and psoriasis were evenly distributed in the families. Clustering independent of age was only observed for peripheral arthritis, suggesting that specific factors may predispose individuals to this manifestation. CONCLUSION: Familial SpA appears to be homogeneous, based on the high frequencies of axial skeletal involvement and HLA-B27. The lack of clustering of most manifestations in families suggests that a predominant shared component, including HLA-B27, predisposes individuals to all forms of familial SpA, and that ubiquitous genetic or environmental factors contribute to phenotype diversity. PMID- 10857796 TI - Primary association of tumor necrosis factor-region genetic markers with susceptibility to rheumatoid arthritis. AB - OBJECTIVE: To determine whether tumor necrosis factor (TNF) polymorphisms are associated with susceptibility to rheumatoid arthritis (RA) independently of the HLA-DR shared epitope. METHODS: Fifty-two Spanish families with one or more affected members were typed for HLA-DRB1, TNF promoter polymorphisms, and TNFa and TNFb microsatellites. We performed an association analysis comparing transmitted versus not transmitted haplotypes, with or without shared epitope, to determine whether there is an independent effect of TNF genetic markers on RA susceptibility. RESULTS: TNFa6,b5 was significantly associated with susceptibility to RA. The haplotypes containing these markers were preferentially transmitted to the affected offspring, even if these haplotypes lacked the HLA-DR shared epitope. TNF promoter polymorphisms were not associated with susceptibility to RA. CONCLUSION: The data suggest that TNFa/b is an independent marker of RA susceptibility, pointing to a genetic role of the TNF region in the pathogenesis of RA. PMID- 10857797 TI - Antineutrophil cytoplasmic antibodies in patients with early rheumatoid arthritis: an early marker of progressive erosive disease. AB - OBJECTIVE: To evaluate the clinical associations of antineutrophil cytoplasmic antibodies (ANCA) in patients with early rheumatoid arthritis (RA), as well as the possible predictive role of ANCA. We also assessed the overlap of ANCA with other specific serologic markers of RA. METHODS: Eighty-two RA patients with symptoms for < or = 12 months were studied for the presence of ANCA by immunofluorescence and specific enzyme immunoassays. ANCA were determined and clinical, radiographic, and laboratory data were collected at study entry and later at 12, 36, 60, and 84 months. RESULTS: In 2 patients, the first serum samples (obtained at study entry) were no longer available for the determination of ANCA. Perinuclear ANCA (pANCA) were found in 40 patients (50%), and atypical cytoplasmic ANCA were found in 3 patients (4%) at study entry. Perinuclear ANCA positive patients were significantly more frequently positive for rheumatoid factor (78%) than were ANCA-negative patients (54%) (P = 0.0297). Fifty-five percent of pANCA-positive patients and 22% of ANCA-negative patients were positive for antiperinuclear factor (P = 0.0044). Similarly, pANCA-positive patients had antikeratin antibodies more frequently than did ANCA-negative patients (35% versus 20%). During a 7-year followup, the progress of radiographic joint destruction, assessed with Larsen scores, was significantly more rapid in patients who were pANCA positive at study entry than in those who were ANCA negative (P = 0.0015). Also, the mean titer of pANCA at study entry was significantly higher in those patients who subsequently had advanced radiographic joint destruction at 60 and 84 months. The association of pANCA with rapid radiographic destruction in patients with early RA was further corroborated by a logistic regression analysis that selected pANCA positivity as an independent and statistically significant predictor of rapid radiographic joint destruction. CONCLUSION: In patients with early RA, pANCA are associated with specific serologic markers of RA and predict rapid radiographic joint destruction. PMID- 10857798 TI - Computer-based methods for measuring joint space and estimating erosion volume in the finger and wrist joints of patients with rheumatoid arthritis. AB - OBJECTIVE: This study was conducted 1) to determine the feasibility of using computer programs to measure radiographic joint space width and estimate erosion volume in the hands of patients with rheumatoid arthritis (RA), and 2) to compare the new computer-based methods with established scoring methods. METHODS: To measure the joint space width in the finger and wrist joints of RA patients, hand and wrist radiographic films were scanned using a tabletop scanner and analyzed with programs written using the "macro" capabilities of NIH Image software. Estimation of erosion volume was determined by utilizing gray-scale intensity to calibrate bone density units per mm3, which made possible comparisons between the erosions and noneroded, anatomically similar sites. RESULTS: In 3 sets of duplicate measurements of joint space width on 79, 48, and 48 finger and wrist joints, the mean absolute deviation from the mean of the 2 measurements was 0.036 mm (SD 0.034), 0.032 mm (SD 0.049), and 0.021 mm (SD 0.016), respectively. Joint space measurements and scoring of joint space narrowing both demonstrated a difference between active treatment and placebo in an old trial set on gold therapy (P = 0.03 and P = 0.01, respectively). Two repeated measurements of bone density units in the bones of 3 different hands differed from the mean of the measurement by 2.29-4.04%. In 2 experiments, estimates of erosion volume showed a greater difference between gold therapy and placebo than did erosion scores in the trial set (P = 0.049 and P = 0.016 versus P = 0.27). CONCLUSION: Computer based methods for measuring finger and wrist joint spaces and estimating erosion volume in patients with RA agree with the results of radiographic scoring of erosions and joint space narrowing. PMID- 10857799 TI - Social, emotional, and behavioral functioning of children with juvenile rheumatoid arthritis. AB - OBJECTIVE: To investigate the hypothesis that children with juvenile rheumatoid arthritis (JRA) would have more social and emotional problems than case-control classmates. METHODS: Using a case-control design, children with JRA (n = 74), ages 8-14, were compared with case-control classmates (n = 74). Peer relationships, emotional well-being, and behavior, based on peer-, teacher-, parent-, and self-report scores on common measures, were compared using analysis of variance. RESULTS: Relative to case-control classmates, children with JRA were similar on all measures of social functioning and behavior. Mothers reported more internalizing symptoms in the child with JRA, but child self reports and father reports showed no differences. Scores on all standardized measures were in the normal range for both the JRA and the case-control groups. CONCLUSION: Children with JRA were remarkably similar to case-control children on measures of social functioning, emotional well-being, and behavior. These findings are not supportive of disability/stress models of chronic illness in childhood and suggest considerable psychological hardiness among children with JRA. PMID- 10857800 TI - Polymorphism at NRAMP1 and D2S1471 loci associated with juvenile rheumatoid arthritis. AB - OBJECTIVE: To examine the role of NRAMP1 in susceptibility to juvenile rheumatoid arthritis (JRA). METHODS: DNA from 119 JRA patients (72 pauciarticular, 47 polyarticular) and 111 healthy controls from Latvia was genotyped for a functional repeat polymorphism in the promoter of NRAMP1 and a linked (<150 kb) microsatellite D2S1471. The findings were compared with those from HLA-DQ alleles typed previously. Chi-square analyses were performed using the Mantel-Haenszel test and stratification according to pure Latvian or pure Russian descent. Haplotype analysis was performed using the Associate program to implement the expectation-maximization algorithm based on the gene-counting technique. RESULTS: Allele 3 at NRAMP1 conferred increased risk (odds ratios [ORs] 2.26, 2.31, and 2.19; P = 0.0006, 0.003, and 0.019) of disease in the JRA, pauciarticular, and polyarticular patient groups, respectively. Allele 2 conferred protection (OR 0.44, 0.43, and 0.46). Alleles at D2S1471 that conferred susceptibility (6 and 12) or protection (11) did so only when on a haplotype with alleles 3 or 2, respectively, at NRAMP1. Allele 3 at NRAMP1 was additive with HLA-DQ7 for susceptibility (OR 3.71, 3.71, and 4.02), and allele 2 at NRAMP1 was additive with HLA-DQ5 for protection (OR 0.19, 0.08, and 0.12). CONCLUSION: The NRAMP1 allele conferring susceptibility to JRA drives high levels of NRAMP1 expression, while the allele associated with protection drives low levels. These 2 alleles are inversely associated with susceptibility to infectious disease, consistent with their maintenance in populations through balancing selection. PMID- 10857801 TI - Nephrotic-range proteinuria, the major risk factor for early atherosclerosis in juvenile-onset systemic lupus erythematosus. AB - OBJECTIVE: To determine the presence of early carotid atherosclerosis and associated risk factors in patients with juvenile-onset systemic lupus erythematosus (SLE). METHODS: The carotid intima-media wall thickness (IMT) was measured by B-mode ultrasound in patients with SLE onset before the age of 16 years and in sex- and age-matched healthy control subjects. Risk factors for atherosclerosis were determined at the time of the ultrasound scan and included traditional cardiovascular and SLE-related risk factors. RESULTS: Twenty-six patients with juvenile-onset SLE and 26 healthy controls were studied. The mean (+/- SD) IMT of the SLE patients was significantly higher than that of the control group (0.57+/-0.05 mm and 0.54+/-0.03 mm, respectively; P = 0.006). The results of IMT measurement were not correlated with the patients' age, disease duration, SLE Disease Activity Index (SLEDAI) score, Systemic Lupus International Collaborating Clinics/American College of Rheumatology (SLICC/ACR) Damage Index (DI) score, laboratory indicators of lupus activity, or cumulative prednisone dose. Patients with nephrotic-range (NR) proteinuria (> or = 3.5 gm/24 hours; n = 6) had a significantly higher IMT than did those without (n = 20) (P = 0.02). Patients with NR proteinuria also had significantly higher SLEDAI scores, SLICC/ACR DI scores, and systolic and diastolic blood pressures, and significantly higher levels of total cholesterol, low-density lipoprotein cholesterol, apolipoprotein B, and fibrinogen. No difference in any of the above variables, including the IMT, was observed when SLE patients without NR proteinuria were compared with healthy controls. CONCLUSION: These patients with juvenile-onset SLE had ultrasonographic evidence of premature atherosclerosis. The risk of early atherosclerosis may be higher in patients with NR proteinuria. PMID- 10857802 TI - The use of alternative medical therapies in patients with systemic lupus erythematosus. Trination Study Group. AB - OBJECTIVE: As part of an ongoing study of health resource utilization and diminished productivity in patients with systemic lupus erythematosus (SLE), the use of alternative medical therapies was assessed. METHODS: A cohort of 707 patients with SLE from 3 countries completed questionnaires on demographics, social support, health status (using the Short Form 36 health survey), satisfaction with health care, health resource utilization (conventional resources and alternative therapies), and time losses in labor market and non labor market activities. Annual direct and indirect costs (1997 Canadian dollars) were calculated and compared for users and nonusers of alternative medical therapies. RESULTS: Among the 707 patients, 352 (49.8%) were found to use alternative therapies and at similar rates across Canada, the United States, and the United Kingdom. Users were younger and better educated than nonusers, exhibited poorer levels of self-rated health status and satisfaction with medical care, and had minimal to no objective evidence of worse disease (according to the revised Systemic Lupus Activity Measure instrument). The mean of log direct medical costs for conventional resources was higher for users of select alternative therapies compared with nonusers. In a logistic regression, neither the number of alternative therapies used nor the individual therapy increased the probability of incurring indirect costs. CONCLUSION: The use of alternative medical therapies is common in patients with SLE. Users of many alternative medical therapies accrue greater conventional medical costs compared with nonusers. The use of alternative medical therapy may be a marker for care-seeking behavior associated with higher consumption of conventional medical resources in the absence of demonstrable additional morbidity and should be considered in future cost analyses of patients with SLE. PMID- 10857803 TI - "G" stands for gout. PMID- 10857804 TI - Val58Ile polymorphism of the neutrophil chemoattractant LECT2 and rheumatoid arthritis in the Japanese population. PMID- 10857805 TI - Nail shedding (Beau's lines) after severe gold dermatitis. PMID- 10857806 TI - Poly(ADP)-ribose polymerase and susceptibility to systemic lupus erythematosus and primary antiphospholipid syndrome: comment on the article by Delrieu et al. PMID- 10857807 TI - Analysis of the association of the matrillin-1 gene (CRTM) with osteoarthritis: comment on the article by Meulenbelt et al. PMID- 10857808 TI - Diagnosis of osteoarthritis in relation to molecular processes in cartilage: comment on the article by Clark et al. PMID- 10857809 TI - Trends in the inflammatory response in biopsy-proven giant cell arteritis: comment on the article by Cid et al, and the letters by Nesher and Sonnenblick and Liozon et al. PMID- 10857810 TI - Macular drusen quantitation. PMID- 10857811 TI - Wavelength considerations for laser prophylaxis in AMD. PMID- 10857812 TI - Excimer laser PRK and nerve fiber layer thickness measurements. PMID- 10857813 TI - To reconstruct or not. PMID- 10857814 TI - Complications of small clear zone radial keratotomy. PMID- 10857815 TI - Development of the foveal avascular zone. PMID- 10857816 TI - Glaucoma and myopia. PMID- 10857817 TI - Drinking water and contact lenses. PMID- 10857818 TI - Effect of heparin-surface-modified intraocular lenses on postoperative inflammation after phacoemulsification: a randomized trial in a United States patient population. Heparin-Surface-Modified Lens Study Group. AB - OBJECTIVE: To compare postoperative inflammation occurring with heparin-surface modified (HSM) versus non-HSM polymethyl methacrylate intraocular lenses (IOLs) after phacoemulsification. DESIGN: Randomized, double-masked, multicenter, parallel trial. PARTICIPANTS: A total of 367 patients, consisting of routine (n = 220), glaucoma (n = 58), and diabetes (n = 89) patients, from eight US medical centers. METHODS: Patients were observed for 1 year after phacoemulsification and lens implantation (week 1, months 1, 3, 6, 12). MAIN OUTCOME MEASURES: Primary measures of postoperative inflammation defined as the presence of giant cells on the lens surface via specular micrography and cellular deposits via slit-lamp examination. RESULTS: The cross-sectional analyses showed that consistently fewer routine patients with HSM lens implants had giant cells on the IOL than those with non-HSM lens implants across all follow-up visits. The statistical significance (P < 0.05) was observed at all visits except month 12 for routine patients. The diabetes patients also demonstrated the same giant cell difference, and the statistical significance was observed at all visits including month 12. A similar trend was also observed in the glaucoma patients, with statistical significance only at the 3-month visit. For cell deposits, significant differences in favor of the HSM lens (P < 0.05) were observed at 3 months among routine and diabetes patients, and at 3 and 6 months among glaucoma patients. A longitudinal data analysis using the generalized estimating equation approach indicated statistically significant treatment effect of HSM lenses in reducing inflammation in all patients except for cellular deposits in diabetes patients. In all patient groups, sight-threatening complications were not reported either more frequently or with more severity than normally expected for patients who have undergone cataract extraction and IOL implantation. CONCLUSIONS: The present study, the only one to have used phacoemulsification in virtually all patients (211/220 [96%] routine, 57/58 [98%] glaucoma, and 84/89 [94%] diabetes) is the largest to evaluate and compare concurrently routine, glaucoma, and diabetes patients. It is also the first US patient population study to document that heparin surface modification reduces postoperative inflammatory responses, as measured by specular micrography and slit-lamp examination, especially in the early postoperative period. PMID- 10857819 TI - Epidemic Bacillus endophthalmitis after cataract surgery II: chronic and recurrent presentation and outcome. AB - OBJECTIVE: To report the clinical outcome of chronic Bacillus endophthalmitis after cataract surgery. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Five eyes of five patients with late-onset or recurrent inflammation after exposure to bacteria-contaminated viscoelastic material were studied. INTERVENTION: Repeated vitrectomies, wide excision of the remnant posterior capsule, and intravitreal injections of antibiotics in five patients. Eventual explantation of the intraocular lens in four patients. MAIN OUTCOME MEASURES: Final visual acuities and results of microbiologic studies of aqueous and vitreous specimens as well as pathologic studies using hematoxylin-eosin, Gram, and periodic acid-Schiff (PAS) stain of explanted capsular remnants were obtained. RESULTS: Final visual acuity of 20/40 or better was obtained in three patients. Bacillus species were grown from two cases. PAS- and Gram-positive microorganisms were identified in the capsular tissue in three of four patients who had explantation of the intraocular lens. CONCLUSIONS: A chronic form of Bacillus endophthalmitis is described for the first time. The clinical outcome is similar to chronic endophthalmitis caused by other organisms. PMID- 10857820 TI - In vitro adherence of Staphylococcus epidermidis to polymethyl methacrylate and acrysof intraocular lenses. AB - PURPOSE: To investigate the adherence of one clinically relevant ocular isolate of Staphylococcus epidermidis to polymethyl methacrylate (PMMA) and Acrysof (Alcon Surgical, Fort Worth, TX) intraocular lenses (IOLs). DESIGN: Experimental study. PARTICIPANTS: The authors examined the in vitro adherence of one clinically relevant ocular isolate of S. epidermidis. Adherence was tested on 12 PMMA IOLs and 12 Acrysof IOLs. METHODS: Six IOLs (three of each type) were placed in different test tubes containing bacterial suspension (10(8) cfu/ml) and incubated at 37 degrees C. At different times (3 minutes, 30 minutes, and 90 minutes), each IOL type was removed from the test tube, rinsed in sterile phosphate-buffered saline, and transferred into sterile brain-heart infusion broth. The broth with the IOL was sonicated on low power for 3 minutes to remove adhered bacteria. Two serial 10-fold dilutions of the broth containing the dislodged bacteria were plated on mannitol agar plates. The plates were incubated overnight at 37 degrees C and then bacterial colonies were counted. All assays were performed in triplicate. Additional lenses (three of each type) were incubated with S. epidermidis for different times (3 minutes, 30 minutes, and 90 minutes) and then examined with scanning electron microscopy. MAIN OUTCOME MEASURES: The number of adhered bacteria per area (mm ) of IOL optic was calculated. Statistical analysis included calculation of arithmetic means and 95% confidence intervals (t test). RESULTS: Direct counting of viable adherent bacteria released by sonication showed that the amount of adhered bacteria per area of IOL optic after 3, 30, and 90 minutes' incubation in S. epidermidis suspension was 0.1 x 10(2)/mm2, 3.6 x 10(2)/mm2, and 11 x 10(2)/mm2 (PMMA IOLs), and 4.4 x 10(2)/mm2, 3.1 x 10(2)/mm2, and 2.3 x 10(2)/mm2 (Acrysof IOLs). Direct counting of adherent bacteria in scanning electron microscopy photographs revealed that the total amount of adhered bacteria per area of IOL optic after 3, 30, and 90 minutes' incubation in S. epidermidis suspension was 1.1 x 10(2)/mm2, 4.4 x 10(2)/mm2, and 5.5 x 10(2)/mm2 (PMMA IOLs) and 13 x 10(2)/mm2, 33.9 x 10(2)/mm2, and 72 x 10(2)/mm2 (Acrysof IOLs). CONCLUSIONS: Results suggest that in vitro adherence of S. epidermidis to IOLs is influenced by IOL materials. After 3 minutes' incubation, Acrysof IOLs appeared to be more permissive to S. epidermidis adherence than PMMA IOLs. The difference was statistically significant (P < 0.05). However, at 90 minutes, Acrysof IOLs had a lower viable bacterial count than did the PMMA IOLs. Bacterial adherence appeared to be greater in areas with surface irregularities. Adherence of S. epidermidis to IOLs may play a role in the pathogenesis of some forms of endophthalmitis after cataract surgery. PMID- 10857821 TI - Improved detection of microorganisms by polymerase chain reaction in delayed endophthalmitis after cataract surgery. AB - OBJECTIVE: To evaluate whether the use of polymerase chain reaction (PCR) improves the identification of the causative pathogen in eyes developing delayed endophthalmitis after cataract surgery. DESIGN: Prospective, noncomparative case series. PARTICIPANTS: Consecutive series of 25 eyes with the clinical diagnosis of delayed endophthalmitis after cataract. MAIN OUTCOME MEASURE: Presence of bacterial or fungal DNA in aqueous humor and vitreous samples. RESULTS: In the aqueous humor the causative pathogen was identified in 84% (n = 21) of the eyes by PCR compared with 0% by diagnostic culture and 0% by microscopy. In the vitreous samples the pathogen was identified in 92% (n = 23) of the eyes by PCR compared with 24% by diagnostic culture (n = 6) and 0% by microscopy. CONCLUSIONS: PCR is useful for the identification of the causative pathogen in delayed endophthalmitis and had a higher rate of positive identification of the causative organism than microscopy or diagnostic culture. PMID- 10857822 TI - Pilot study on erbium laser phacoemulsification. AB - PURPOSE: To investigate the suitability of erbium laser phacoemulsification for cataract surgery using a prospective pilot study. DESIGN: Prospective, single center, nonrandomized clinical trial. PATIENTS AND METHODS: Slit-lamp microscopy, keratometry, best-corrected visual acuity, refraction, pachymetric corneal thickness, endothelial cell density, and intraocular pressure were assessed before surgery and on the first, fourth, fourteenth, and the sixtieth day after surgery in 40 eyes of 34 patients with senile cataract (17 males with a mean age of 67.3 years; 17 females with a mean age of 73.2 years). All operations were performed by one surgeon (HH) using the MCL-29 erbium laser (Aesculap-Meditec, Jena, Germany). The nuclear sclerosis grade ranged from 0 to 4. MAIN OUTCOME MEASURES: Primary outcome measures were defined as the ability to emulsify the lens nucleus under clinical conditions and the occurrence of side effects. Secondary outcome measures included the change in visual acuity, refraction, intraocular pressure, corneal thickness, and endothelial cell density. RESULTS: Complete emulsification of the lens nucleus using the erbium laser was achieved in 36 of 40 eyes (90%; nuclear sclerosis grade, 0-3). Partial emulsification of the nucleus was possible in two cases with grade 3 nuclear sclerosis, in one case of grade 2 nuclear sclerosis, and in one case of cataract with grade 4 nuclear sclerosis. Mean phacoemulsification time was 3 minutes, and the total applied energy equaled 38.5 J. The postoperative changes in visual acuity, spherical and astigmatic refraction, and intraocular pressure were found to be the same as with ultrasonic phacoemulsification. The decrease in the density of endothelial cells (by 0.96%) was not statistically significant. Posterior capsule ruptures occurred in three eyes early in the series. There were no further vision-threatening complications. CONCLUSIONS: Erbium laser phacoemulsification is effective for lenses with mild to moderate nuclear sclerosis. For higher grades of nuclear sclerosis, further improvements in technical and surgery-related parameters are required. PMID- 10857823 TI - Demarcation laser photocoagulation of selected macula-sparing rhegmatogenous retinal detachments. AB - OBJECTIVE: To report a series of macula-sparing rhegmatogenous retinal detachments (MSRRDs) treated with demarcation laser photocoagulation (DLP). DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Thirty-one patients (34 eyes) with primary or recurrent MSRRDs without associated visual field loss, necrotizing retinitis, or proliferative vitreoretinopathy (PVR), managed with DLP from November 1992 through May 1999. INTERVENTION: Demarcation laser photocoagulation consisting of a triple row of confluent laser burns. MAIN OUTCOME MEASURES: Best corrected postoperative visual acuity and MSRRD progression or recurrence. RESULTS: Thirty-four primary and recurrent MSRRDs were treated by DLP, which consisted of a triple row of confluent laser burns. Macula sparing rhegmatogenous retinal detachments were located in all quadrants and affected 10% to 45% of the retina. Findings associated with MSRRDs included lattice degeneration (12 eyes), vitreous hemorrhage (4 eyes), and demarcation line (9 eyes). Symptoms (photopsias or floaters) were associated with 14 MSRRDs. Eight eyes were myopic and 11 were pseudophakic. Thirty-two MSRRDs were shallow, two were dome shaped, and all were smooth without corrugations. Follow-up ranged from 1.5 to 80 months (mean, 15.8 months; median, 17 months). Thirty-three of 34 detachments remained stable after DLP. Three flattened spontaneously. One eye was managed with scleral buckle 6 weeks after DLP. Progression was attributed to incomplete laser treatment. Best corrected postoperative visual acuity was the same or improved in all but one eye, in which a cataract developed. CONCLUSIONS: Demarcation laser photocoagulation is an effective method to manage acute or chronic, primary or recurrent MSRRDs without associated PVR that are shallow and smooth without corrugations. Demarcation laser photocoagulation is an alternative to both observation and surgical repair for these select MSRRDs. PMID- 10857824 TI - Visual acuity outcomes with and without surgery in patients with persistent fetal vasculature. AB - PURPOSE: To investigate visual acuity outcomes in patients with persistent fetal vasculature (PFV) left untreated or treated with vitreoretinal surgical techniques and to investigate clinical features associated with prognosis. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: All patients with PFV examined at the Bascom Palmer Eye Institute from January 1, 1983 through December 31, 1998. INTERVENTION: All patients in the study had unilateral PFV. Of 42 PFV patients identified, 30 patients underwent vitreoretinal surgery. Indications for surgery included media opacity (e.g., cataract), vitreoretinal traction, and retinal detachment. MAIN OUTCOME MEASURES: Final best postoperative visual acuity, prognostic ocular clinical features, and surgical complications. RESULTS: In the surgical group of patients, median age at diagnosis was 8 weeks, and median length of follow-up was 32 months, with all patients having at least 1 year of follow-up. Two patients had clinical and echographic findings consistent with anterior PFV, 2 patients had strictly posterior PFV, and the remaining 26 patients had components of both anterior and posterior PFV. Fourteen eyes (47%) achieved a final visual acuity of 20/400 or better at last follow-up. Risk factors for a poor visual acuity outcome (<20/400) included microphthalmia (28% of patients with microphthalmia versus 67% of patients with normal axial length achieved a final vision of 20/400 or better; P = 0.061) and preoperative retinal detachment or retinal or optic nerve abnormalities, or both, such as hypoplasia, folds, or indistinct macula with hypopigmentation (25% of patients with any of these anomalies versus 61 % of patients without these findings achieved a final vision of 20/400 or better; P = 0.072). After surgery, retinal detachment developed in three eyes, chronic hypotony in two other eyes, and neovascular glaucoma in one eye. In the nonsurgical group there were 6 male and 6 female patients. Two patients with posterior PFV had minimal disease and were not considered surgical candidates, whereas 10 patients with combined anterior and posterior PFV had advanced pathologic features, and it was believed that surgery would not offer significant visual improvement; median age at diagnosis was 9.5 months, and median length of follow-up was 36 months, with all patients having at least 1 year of follow-up. At last follow-up, 3 eyes (25%) had a final visual acuity of 20/400 or better. During follow-up, retinal detachment developed in 2 eyes and chronic hypotony in an additional 2 eyes. CONCLUSIONS: The current study indicates that approximately 50% of patients undergoing surgery for PFV will achieve useful vision. Visual acuity outcomes in patients with PFV are correlated with the nature and extent of ocular risk factors. Some patients may not be candidates for surgery because of either minimal changes or advanced disease that limit the potential of visual improvement. PMID- 10857825 TI - Surgical treatment of macular holes with multiple recurrences. AB - PURPOSE: To evaluate the results of a third macular hole surgery in eyes with recurrent macular holes and two prior macular hole surgeries. DESIGN: Retrospective consecutive noncomparative case series. PARTICIPANTS: Sixteen eyes of sixteen patients with two prior macular hole surgeries with recurrent macular hole. INTERVENTION: A third vitreous surgery was performed in each eye using a long-acting gas bubble. MAIN OUTCOME MEASURE: Closure of the macular hole and change in visual acuity. RESULTS: The macular hole was closed in 12 of 16 eyes (75%) at 3 months after the third surgery. Visual acuity improved 2 or more Snellen lines in 9 of 16 eyes (56%), and 5 of 16 eyes (31%) achieved 20/40 or better vision. Six eyes (37.5%) had cataract surgery after the third macular hole surgery, and visual acuity results were similar in eyes with or without cataract surgery. Successful closure of the macular hole improved the visual acuity from 20/80 -1 to 20/50 +1 (P < 0.001). Eyes in which one of the previous surgeries had been temporarily successful in closing the macular hole improved from a mean of 20/80 to 20/40 (P = 0.003). Eyes in which both prior macular hole surgeries had been primary failures had minimal benefit with a preoperative visual acuity of 20/100 +1 and a postoperative visual acuity of 20/100 +2 (P = 0.67). CONCLUSIONS: Repeat macular hole surgery should be considered in eyes with recurrent macular holes and two prior surgeries when the macular hole was temporarily closed by at least one of the two previous surgeries. Successful closure of a macular hole in such cases usually results in significant visual acuity improvement. PMID- 10857826 TI - Perfluoroperhydrophenanthrene versus perfluoro-n-octane in vitreoretinal surgery. AB - PURPOSE: To compare two perfluorocarbon liquids, perfluoroperhydrophenanthrene (Vitreon) and perfluoro-n-octane (PFO), regarding (1) ease of removal at the time of surgery and (2) efficacy and safety. DESIGN: Retrospective, nonrandomized, comparative trial. PARTICIPANTS: Two hundred sixty-four patients. METHODS: All medical records of patients who participated in either the Vitreon (n = 142) or the PFO (n = 122) multicenter trials at the Wilmer Eye Institute were reviewed. Patients in the two groups were compared with respect to presence and amount of postoperative perfluorocarbon. Safety and efficacy of the two perfluorocarbon liquids were evaluated in the subset of patients undergoing surgery for retinal detachment with PVR grade C or more, who had at least 6 months of postoperative follow-up (n = 46 in the Vitreon group and n = 55 in the PFO group). Best corrected visual acuity, intraocular pressure, and status of the cornea, lens, and retina after 3, 6, and 12 months of follow-up were compared in the two groups. MAIN OUTCOME MEASURES: Percentage of eyes with retained perfluorocarbon, final visual acuity, final retinal attachment rate. RESULTS: At the 6-month postoperative visit, retained Vitreon or PFO was detected in 8.4% and 4% of the eyes, respectively (P < 0.05). No statistically significant difference was found between the two perfluorocarbons in attachment rate or visual acuity after 6 and 12 months. After 6 months, the retina was attached in 71.7% of the eyes in which Vitreon was used and in 78.3% of the eyes in which PFO was used (P = 0.81). Visual acuity improved or remained stable in 80% of the Vitreon group eyes and in 74.5% of the PFO group eyes (P = 0.75). No statistically significant difference was found in final visual acuity or intraocular pressure between the two groups after 6 or 12 months. There was a tendency at the 12-month follow-up examination for the cornea to remain clearer in the Vitreon group (P = 0.01). CONCLUSIONS: Although PFO was easier to visualize and remove completely from within the eye at the time of surgery, the efficacy in terms of retinal attachment and final visual outcome was similar between the Vitreon and PFO groups. PMID- 10857827 TI - Vitreoretinal surgery through multifocal intraocular lenses compared with monofocal intraocular lenses in fluid-filled and air-filled rabbit eyes. AB - PURPOSE: To compare vitrectomy procedures and visualization of posterior segment structures through multifocal silicone intraocular lenses (IOLs) with the same procedures through monofocal silicone IOLs in rabbit eyes. DESIGN: Experimental study. PARTICIPANTS: Twelve eyes of six rabbits. METHODS: Each rabbit eye underwent phacoemulsification of the lens and posterior chamber implantation of a silicone multifocal or silicone monofocal IOL. The type of IOL (monofocal vs. multifocal) implanted in the first eye of each rabbit was randomly decided. The fellow eye then received the other IOL type. Vitrectomy procedures were performed through the IOLs by using a flat contact lens (part 1) or wide-angled contact lens (part 2) for visualization through fluid-filled (parts 1 and 2) and air filled (part 2) eyes. MAIN OUTCOME MEASURES: Image quality, stereopsis, and contrast were subjectively graded on a scale of 0 (none) to 4 (excellent) for each eye by each surgeon. RESULTS: In part 1, image quality averaged 4 for the monofocal IOL and 3.6 for the multifocal IOL. Stereopsis averaged 4 for the monofocal IOL and 4 for the multifocal IOL. Contrast averaged 4 for the monofocal IOL and 3.9 for the multifocal IOL. Vitrectomy with retinal surface maneuvers was successfully performed in both pigmented and nonpigmented rabbit eyes through both IOL types. In part 2, image quality, stereopsis, and contrast were rated as 4 for both multifocal and monofocal silicone IOLs. Air-fluid exchange was performed without difficulty. Image quality, stereopsis, and contrast were rated as 4 for air-filled eyes. CONCLUSIONS: Visualization of posterior segment structures through multifocal silicone IOLs was sufficient for retinal surface maneuvers during vitrectomy procedures in both fluid-filled and air-filled rabbit eyes. PMID- 10857828 TI - Evaluation of functional defects in branch retinal vein occlusion before and after laser treatment with scanning laser perimetry. AB - OBJECTIVE: This study was aimed at localizing and quantifying retinal defects in patients with branch retinal vein occlusion by means of scanning laser perimetry and analyzing the mechanism involved in the functional recovery after laser therapy. DESIGN: Prospective nonrandomized clinical trial with concurrent comparison group. PARTICIPANTS: Fifty-eight patients with isolated branch retinal vein occlusion. Thirty-nine eyes received laser treatment; 19 eyes were observed without treatment. INTERVENTION: Argon laser photocoagulation was performed according to the Branch Vein Occlusion Study recommendations. MAIN OUTCOME MEASURES: Retinal functional deficits were evaluated with scanning laser perimetry and fluorescein angiography first at baseline, at 3-month follow-up visits and 3 months after laser treatment. RESULTS: After laser treatment, the scotoma encroached on foveal fixation in 31% of eyes, remained stable in 36%, and regressed from the foveal avascular zone in 33%. Improvement in vision was correlated with increasing scotoma distance from fixation. Total scotoma size increased in 50% of eyes after treatment. Depth of scotoma and degree of angiographic leakage showed no direct correlation. CONCLUSIONS: Stabilization and increase in visual acuity after laser treatment did not correlate with an overall decrease in scotoma size. Improved central visual function seen in 25% of treated eyes appeared to be due to withdrawal of scotoma from the fovea. PMID- 10857829 TI - Unusual immunogammopathy maculopathy. AB - PURPOSE: To describe an unusual maculopathy in patients with serum immunogammopathies. DESIGN: Retrospective observational small case series. PARTICIPANTS: Three patients derived from the clinical retina practices of the authors were noted to have unusual maculopathy. METHODS: Each patient underwent fluorescein angiography and serum laboratory evaluation. MAIN OUTCOME MEASURE: Findings on fluorescein angiography. RESULTS: An unusual and atypical macular detachment with or without subretinal precipitates or fundus signs of serum hyperviscosity, such as retinal hemorrhages and dilated retinal veins, may be observed in patients with immunogammopathies such as multiple myeloma, Waldenstrom's macroglobulinemia, and benign polyclonal gammopathy. Fluorescein angiography shows macular hypofluorescence with no evidence of retinal vascular or retinal pigment epithelial leakage within the macular elevation. CONCLUSIONS: Patients with atypical unilateral or bilateral macular detachment may be afflicted with a serum immunogammopathy such as multiple myeloma or Waldenstrom's macroglobulinemia. Diagnostic serum protein electrophoresis and hematology consultation should be considered. PMID- 10857830 TI - Clinical and electroretinographic findings of female carriers and affected males in a progressive X-linked cone-rod dystrophy (COD-1) pedigree. AB - OBJECTIVE: To study the clinical and electroretinographic findings of affected males and female carriers in a family with X-linked cone-rod dystrophy (COD-1). DESIGN: Observational case series. PARTICIPANTS: Twenty-five members of a five generation pedigree were examined. METHODS: A history of visual impairment including age at onset, loss of acuity, color vision abnormalities, photophobia, and nyctalopia was obtained. A complete ophthalmologic examination was performed, including kinetic perimetry with a Goldmann perimeter, FM 100-hue testing, and standardized Ganzfeld electroretinography following the ISCEV protocol. MAIN OUTCOME MEASURES: Patients were classified as affected or unaffected on the basis of the clinical examination. All carrier females had affected sons. RESULTS: Nine affected males and seven female carriers were identified. Affected males noted decreased visual acuity and poor color vision within the first two decades of life. Early in the disease, macular retinal pigment epithelial (RPE) changes were found that progressed to an atrophic macular scar by the fifth decade. Evidence of progression from macular pigment mottling to an atrophic macular lesion over a 13-year period was identified in one patient. The photopic, single-flash, b-wave amplitude was low in all affected males and declined with age. The 30-Hz flicker b-wave implicit times were abnormally prolonged in all affected males. Female carriers were asymptomatic although three had slightly abnormal color vision and small paracentral field defects and subtle RPE defects were found in three carriers. Carriers demonstrated prolongation of the 30-Hz flicker b-wave implicit time and interocular asymmetry. Five of seven carriers and two affected males demonstrated reduced oscillatory potentials and an abnormal-appearing flattened photopic a-wave. Five men and two women demonstrated a characteristic tapetal like retinal sheen. CONCLUSIONS: Affected patients in this pedigree demonstrate early loss of visual acuity and poor cone function with late rod involvement. Female carriers may appear clinically normal or may be identified by subtle color vision defects, fundus abnormalities, prolongation of the 30-Hz flicker implicit time with interocular asymmetry, or an abnormal flattened photopic a-wave. Genetic linkage analysis of this family was recently reported and the disease causing gene has been mapped to an approximately 1-Mb interval on chromosome Xp11.4. PMID- 10857831 TI - Keratopathy in pseudoexfoliation syndrome as a cause of corneal endothelial decompensation: a clinicopathologic study. AB - PURPOSE: To provide clinical and histopathologic evidence of a distinct keratopathy as a potential cause of corneal edema in patients with pseudoexfoliation syndrome. DESIGN: Retrospective observational case series. PARTICIPANTS: Twenty-two patients with clinically diagnosed pseudoexfoliation syndrome undergoing penetrating keratoplasty for irreversible corneal endothelial decompensation. METHODS: The clinical and histopathologic findings of the corneal buttons are described compared with classic Fuchs' endothelial dystrophy. RESULTS: Clinically, the patients showed diffuse corneal edema, a pleomorphic and numerically reduced corneal endothelium, and retrocorneal flakes of pseudoexfoliation material in three cases. Histopathologically, all corneal buttons showed an abnormal diffuse, irregular thickening of Descemet's membrane and focal accumulations of locally produced pseudoexfoliation material onto or within Descemet's membrane in seven cases. The absence of typical guttata, a higher degree of fibroblastic transformation and melanin phagocytosis of endothelial cells, and a more pronounced endothelial cell loss distinguished the pseudoexfoliation specimens from specimens with classical Fuchs' dystrophy even in the absence of the pathognomonic pseudoexfoliation material. CONCLUSIONS: In patients with pseudoexfoliation syndrome, a distinct type of corneal endotheliopathy may occur, which can lead to an early corneal endothelial decompensation and which might have been previously misdiagnosed as an "atypical nonguttata Fuchs' endothelial dystrophy." This pseudoexfoliation keratopathy may potentiate the known complications in pseudoexfoliation eyes. PMID- 10857832 TI - Penetrating keratoplasty for keratoconus: visual outcome and success. AB - OBJECTIVE: To determine the long-term effect on vision of penetrating keratoplasty performed for keratoconus. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: All patients with keratoconus who received a corneal graft and who remained in our center for follow-up and visual rehabilitation during the study period. INTERVENTION: Penetrating keratoplasty was performed in 93 eyes of 78 patients. MAIN OUTCOME MEASURES: Graft survival, visual acuity, and astigmatism. RESULTS: One (1.08%) graft failure was encountered over a mean follow-up of 46 months. Mean preoperative (best corrected) and postoperative visual acuity is (best-tolerated correction) were 0.9 (20/160) and 0.24 (20/80) logMAR, respectively. Visual acuity in 86% of eyes was 0.3 logMAR (20/40) or better at the latest follow-up, with 67% of eyes being corrected with spectacles. Mean preoperative corneal power by keratometry was more than 52 diopters (D) in 83% of eyes; mean postoperative corneal power was 45 +/- 2 D. No significant predictors of postgraft astigmatism were found. Mean preoperative and postoperative best-eye acuities of the better eye were 0.32 (20/40-1) and 0.18 (20/32+1) logMAR, respectively (P < 0.001). CONCLUSIONS: Graft survival was excellent. A corrected visual acuity of 20/40 or better was obtained in 86% of eyes. Astigmatism could not be predicted from preoperative factors. Visual acuity measured in the better eye improved by 0.14 logMAR (1.4 lines), implying an overall functional gain for the patient. PMID- 10857833 TI - Hyperopic laser in situ keratomileusis with the Nidek EC-5000 excimer laser. AB - OBJECTIVE: To evaluate the efficacy, safety, and predictability of hyperopic laser in situ keratomileusis (H-LASIK) using modified software. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: The first 72 consecutive eyes of 44 patients with up to +5.00 diopters (D) hyperopia. INTERVENTION: Hyperopic LASIK using the Automatic Corneal Shaper (ACS; Chiron Vision, Claremont, CA) and the Nidek EC-5000 excimer laser (Nidek, Tokyo, Japan). MAIN OUTCOME MEASURES: Uncorrected visual acuity, manifest spherical equivalent (MSE), best-corrected visual acuity (BCVA), and complications were studied. RESULTS: At 6 months, in the low hyperopia group (<3.00 D), mean MSE was +0.30 +/- 0.71 D, with 88.9% eyes within 1 D of emmetropia compared with +1.09 +/- 0.92 D and 51.8% within 1 D of emmetropia in the moderate hyperopia group (> or =3.00 D; P = 0.003). Uncorrected visual acuity was 20/40 or better in 43 of 45 eyes (95.6%) and in 21 of 27 (77.8%) eyes in the low and moderate hyperopia groups, respectively. Only one eye (1.4%) from the moderate hyperopia group lost two lines of BCVA. Eighteen eyes (25%) required retreatment to correct residual hyperopia, 9 eyes (20.0%) in the low hyperopia group and 9 eyes (33.3%) in the moderate hyperopia group. Retreatments resulted in an MSE of +0.02 +/- 0.45 D and +0.04 +/- 0.73 D in the low and moderate hyperopia groups, respectively. No flap related complications were seen. CONCLUSIONS: Hyperopic LASIK with the ACS and the Nidek EC-5000 excimer laser using our modified software is a safe, effective, and predictable procedure for low hyperopia. Results are satisfactory up to moderate hyperopia. Significant regression can occur for low and moderate hyperopia. Retreatment can be performed safely and effectively to improve the visual and refractive results. PMID- 10857834 TI - Topographically-guided laser in situ keratomileusis to treat corneal irregularities. AB - OBJECTIVE: To evaluate the predictability and safety of topographically guided laser in situ keratomileusis (LASIK) to treat corneal irregularities. DESIGN: Prospective, noncomparative interventional case series. PARTICIPANTS: Twenty seven patients (29 eyes) with postsurgical corneal irregularities, divided into four subgroups (postkeratoplasty, 6 eyes; posttrauma, 6 eyes; postphotorefractive keratectomy (PRK)/LASIK with decentered or small ablations, 11 eyes; post PRK/LASIK with central islands, 6 eyes). INTERVENTION: LASIK was performed using the Automatic Corneal Shaper and the Keracor 117 C spot-scanning excimer laser (Bausch & Lomb Surgical Technolas, Munich, Germany). Individual ablation patterns were calculated on the basis of axial radii of curvature data obtained with the Corneal Analysis System (EyeSys Premier, Irvine, CA). MAIN OUTCOME MEASURES: Change of corneal topography pattern, patient satisfaction, manifest spectacle refraction, and visual acuity at 12 months after surgery. RESULTS: Corneal topography showed improved corneal regularity in 66% of eyes in the postkeratoplasty group, whereas 34% remained irregular. In the posttrauma group, 83% improved and 17% remained irregular. In the decentered/small optical zone group, 91 % improved and 9% remained irregular. In the central islands group, 50% improved and 50% remained irregular. Refractive cylinder decreased from 5.83 +/- 1.25 diopters (D) to 2.96 +/- 1.23 D in the postkeratoplasty group (P = 0.01), from 2.21 +/- 1.35 D to 0.50 +/- 0.84 D in the posttrauma group (P = 0.001), from 0.73 +/- 0.71 D to 0.36 +/- 1.05 D in the decentered/small optical zone group (NS), and from 1.42 +/- 1.13 D to 0.50 +/- 0.84 D in the central island group (P = 0.01). Uncorrected visual acuity improved from 20/200 +/- 0.07 to 20/50 +/- 0.17 in the postkeratoplasty group (P = 0.01), from 20/83 +/- 0.12 to 20/50 +/- 0.28 in the posttrauma group (P = 0.01), from 20/60 +/- 0.16 to 20/50 +/- 0.29 in the decentered/small optical zone group (NS), and from 20/71 +/- 0.12 to 20/60 +/ 0.24 in the central island group (NS). CONCLUSIONS: The topographically-guided LASIK method used in this study resulted in a significant reduction of refractive cylinder, a significant increase of uncorrected visual acuity, and improved corneal regularity in a large percentage of patients with severe corneal irregularities such as decentered/small optical zones after LASIK or irregular astigmatism after keratoplasty or trauma. With small irregularities such as central islands, results were sufficiently poor to advise against the use of our technique in these patients. PMID- 10857835 TI - Ocular surface changes after excimer laser phototherapeutic keratectomy. AB - PURPOSE: To study the ocular surface disorder in patients with Avellino, granular, and lattice dystrophy, band keratopathy, and corneal leukoma before and after excimer laser phototherapeutic keratectomy. DESIGN: A prospective case controlled study. PARTICIPANTS: A total of 45 eyes of 33 patients with superficial corneal opacities seen at Kobe Kaisei Hospital, Department of Ophthalmology, and 40 eyes of 20 normal control subjects were studied. INTERVENTION: The subjects underwent routine ophthalmic examinations, corneal sensitivity measurements, tear film break up time (BUT), Schirmer test, tear film lipid layer interferometry, and conjunctival impression cytology. MAIN OUTCOME MEASURES: The patients and the control subjects were compared for prephototherapeutic keratectomy (PTK) tear function parameters, tear film lipid layer interferometry grade, goblet cell density, and conjunctival squamous metaplasia grade. Alterations of these parameters within 3 months after PTK were also looked for. RESULTS: The average pre-PTK corneal sensitivity and tear film break up time were lower in patients compared with control subjects before PTK. Tear film lipid layer interferometry grade and conjunctival squamous metaplasia grades were higher in the patients than the controls before PTK. All these parameters improved gradually and significantly after PTK. Goblet cell density was significantly lower in the patients compared with controls before PTK. Schirmer test results and goblet cell density did not show any significant alterations after PTK. CONCLUSIONS: Concurrent improvements in corneal sensitivity, tear film break up time, lipid layer interference grades, and conjunctival squamous metaplasia grades all point to the favorable effects of PTK on the ocular surface by improving the stability of the tear film and ocular surface health through attainment of a regular corneal surface and probably inducing qualitatively/quantitatively better mucin production by a healthier epithelium after PTK. PMID- 10857836 TI - Prenatal diagnosis of aniridia. AB - OBJECTIVE: To use molecular genetic techniques to prenatally screen for aniridia. DESIGN: Case report. METHODS: DNA was extracted from cultured fibroblasts obtained through amniocentesis. Two mutation detection methods, Ava1 restriction digestion and single-strand conformational polymorphism electrophoresis, were used to screen the PAX6 gene. MAIN OUTCOME MEASURES: The results from the amniocentesis sample were compared with DNA obtained from the affected father, firstborn infant, and unaffected mother to determine whether the fetus carried the PAX6 mutation. RESULTS: DNA from the fetus demonstrated the same banding pattern as the affected father and firstborn infant. CONCLUSIONS: The fetus carried the mutated PAX6 allele and was predicted to develop aniridia. This was later confirmed when the child was born. This case report illustrates an important use of genetic mutation screening in the clinical setting. PMID- 10857837 TI - Vernal keratoconjunctivitis revisited: a case series of 195 patients with long term followup. AB - OBJECTIVE: This study aimed at revisiting vernal keratoconjunctivitis (VKC) on the basis of anamnestic, clinical, immunologic, histopathologic, and followup data of 195 patients. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: One hundred and ninety-five patients with VKC. METHODS: Clinical evaluation and outcome in 151 of 195 patients with a median followup of 47 months. Evaluation was by telephone survey in 69 patients. MAIN OUTCOME MEASURES: (1) Demographic, clinical, and immunologic features of VKC and their influence on the course of the disease; (2) conjunctival and corneal complications and efficacy of treatment observed during the followup period. RESULTS: VKC is a chronic disease. More than 60% of patients had repeated recurrences all year round. Males had an earlier presentation of symptoms than females and the male/female ratio decreased with age. Major (greater than 80%) and minor (up to 80%) diagnostic criteria were defined for clinical signs and symptoms of the disease. Negative skin test or radioallergosorbent test was present in approximately 50% of patients, whereas eosinophil infiltration was a constant histopathologic finding. A marked conjunctival sensitivity to nonspecific stimuli was noted in more than one third of patients. In 6% of cases, a reduction of visual acuity resulted from corneal scarring, and in 2% of patients, steroid induced glaucoma was observed. The large size of giant papillae indicates poor prognosis for the persistence of the disease and its evolution into a chronic, perennial condition. CONCLUSIONS: VKC is a chronic eosinophilic disease of the ocular surface involving IgE, non IgE-mediated mechanisms, and age-sex-related influences. Although the disease has a good prognosis, severe visual impairments may result from long-standing inflammation. PMID- 10857838 TI - Anterior uveitis with sectoral iris atrophy in the absence of keratitis: a distinct clinical entity among herpetic eye diseases. AB - OBJECTIVE: To determine the cause and describe the clinical features of unilateral anterior uveitis with sectoral atrophy of the iris in the absence of associated keratitis. DESIGN: Retrospective, observational case series. PARTICIPANTS: Thirty-one patients with unilateral anterior uveitis with sectoral iris atrophy and without (previous) keratitis. METHODS: The patients were selected from our database of 592 patients with anterior uveitis. MAIN OUTCOME MEASURES: We reviewed the clinical data on the 31 patients and the results of diagnostic anterior chamber fluid analysis for 24 of the 31 patients. Specifically, production of local antibodies against herpes simplex virus (HSV) and varicella zoster virus (VZV) was determined and the polymerase chain reaction was performed to demonstrate the DNA of HSV, VZV, and cytomegalovirus (CMV) in the aqueous samples. RESULTS: Main clinical characteristics of anterior uveitis with iris atrophy included unilateral involvement with a prolonged course and recurrent exacerbations in all cases. Elevated intraocular pressure during intraocular inflammation occurred in 90% of patients (28 of 31). Visual outcome was favorable because 29 of 31 patients (94%) retained a visual acuity of 20/32 or more. The causal agent was identified as HSV in 83% (20 of 24) and VZV in 13% (3 of 24) and was inconclusive in one case. The patients with HSV uveitis were younger than those with VZV uveitis (mean age at onset 34 and 65 years, respectively; P = 0.0056). CONCLUSIONS: Unilateral anterior uveitis with sectoral atrophy of the iris without associated (previous) keratitis is a distinct entity among herpetic eye diseases. Recurrent unilateral anterior uveitis with iris atrophy and/or elevated intraocular pressure has most likely been caused by HSV. PMID- 10857839 TI - Effects of systemic beta-blocker therapy on the efficacy and safety of topical brimonidine and timolol. Brimonidine Study Groups 1 and 2. AB - PURPOSE: To determine the impact of coadminstration of systemic beta-blockers on the ocular hypotensive efficacy and safety of topical timolol, a nonselective, beta-blocker, and that of brimonidine, an alpha2-selective adrenergic agonist, in patients with glaucoma or ocular hypertension. DESIGN: Post hoc evaluation of data collected from two prospective, multicenter, randomized, double-masked, parallel-group, actively-controlled, 12-month clinical trials. PARTICIPANTS: Of the 926 subjects with ocular hypertension or glaucoma that were enrolled in the two prospective trials, 66 (7.1%) were concurrently maintained on systemic beta blocker therapy. Of these patients, 34 had been assigned to the brimonidine group and 32 to the timolol group. METHODS: Subjects instilled into each eye either 1 drop of brimonidine 0.2% or timolol 0.5% twice daily for 1 year. Study subjects within medication treatment groups were classified as to their use or nonuse of concurrent systemic beta-blockers, and mean intraocular pressure (IOP) reduction, adverse events, heart rate, and blood pressure were compared. MAIN OUTCOME MEASURES: Mean IOP reduction from baseline was the primary efficacy variable. Adverse events and mean changes in heart rate and blood pressure from baseline were the primary safety variables. RESULTS: Timolol-treated subjects concurrently taking systemic beta-blockers had smaller decreases in IOP, a greater mean change in systolic (at week 2, months 1, 2, 6, and 9; P < or = 0.001) and diastolic blood pressure (months 2 and 6; P < or = 0.02), and a significantly greater mean decrease in heart rate (month 6; P = 0.004) compared with timolol subjects not taking systemic beta-blockers. By contrast, there was a modest enhancement of IOP lowering efficacy at trough and no effect on blood pressure or heart rate in brimonidine-treated subjects who were concurrently receiving systemic beta blocker therapy compared with brimonidine subjects not receiving systemic beta blockers. CONCLUSIONS: Concurrent systemic beta-blocker therapy had no deleterious effect on ocular hypotensive efficacy and no impact on systemic safety parameters with topical brimonidine, whereas efficacy was reduced and systemic safety parameters were impacted with topical timolol. Ocular hypotensive agents other than beta-blockers, such as the alpha2 agonist brimonidine, may be a more appropriate first-line therapy for ocular hypertension and glaucoma patients concurrently taking systemic beta-blockers. PMID- 10857840 TI - Comparison of the intraocular pressure-lowering effect of latanoprost and timolol in patients with chronic angle closure glaucoma: a preliminary study. AB - OBJECTIVE: To compare the intraocular pressure (IOP)-reducing effect and side effects of 0.005% latanoprost once daily to 0.5% timolol twice daily in patients with primary chronic angle closure glaucoma (CACG). DESIGN: Randomized, double masked two-center clinical trial. PARTICIPANTS: Thirty-two Asian patients with CACG, defined as glaucomatous optic neuropathy with a compatible visual field defect and at least 6 clock hours of synechial angle closure on gonioscopy were recruited. All patients had previous peripheral iridotomy (PI) with IOP >21 mmHg after PI and were thereafter controlled (IOP <22 mmHg) with one or two pressure reducing drugs. INTERVENTION: After a washout period, the patients were randomized to a 2-week treatment period with either placebo in the morning and 0.005% latanoprost in the evening or 0.5% timolol twice daily. MAIN OUTCOME MEASURES: The short-term IOP reduction of latanoprost and timolol in patients with CACG. IOP was measured at baseline, and after 2, 7, and 14 days of treatment. In addition, the short-term ocular and systemic adverse events of the two drugs were evaluated. RESULTS: Thirty patients completed the study. Two patients in the timolol group were withdrawn because of inadequate IOP control. Compared with baseline, the IOP after 2 weeks of treatment was statistically significantly reduced by 8.8 +/- 1.1 mmHg (mean +/- SEM, P < 0.001) in the latanoprost group, and by 5.7 +/- 0.9 mmHg (P < 0.001) in the timolol group. The difference in IOP reduction between the two treatment groups was 3.1 +/- 1.5 mm Hg in favor of latanoprost (P = 0.04). The main ocular adverse events reported in both treatment groups were conjunctival hyperemia and discomfort. CONCLUSIONS: In this preliminary study, a significantly greater IOP reduction was achieved with 0.005% latanoprost once daily compared with 0.5% timolol twice daily in patients with CACG. The results suggest that latanoprost may be a therapeutic choice for the medical treatment of primary CACG. PMID- 10857841 TI - Low-dose 5-fluorouracil trabeculectomy as initial surgery in uncomplicated glaucoma: long-term followup. AB - OBJECTIVE: To compare the effectiveness of trabeculectomy with adjunctive, low dose, subconjunctival 5-fluorouracil (5-FU) to trabeculectomy alone in patients with uncomplicated glaucoma undergoing their first incisional surgical procedure. DESIGN: Retrospective, nonrandomized comparative trial. PARTICIPANTS: Consecutive series of 52 patients and 74 control subjects. INTERVENTION: Trabeculectomy was performed in all patients. Study patients received adjunctive, subconjunctival injections of 5-FU up to 14 days from the date of surgery. MAIN OUTCOME MEASURES: Intraocular pressure, number of postoperative antiglaucoma medications, interventions, and complications were evaluated. RESULTS: Mean followup for all patients was 58.1 +/- 44.1 months (range, 1.1-159.9 months). Mean followup for successful eyes was 55.9 +/- 47.1 months (range, 7.6-159.9 months). The cumulative 5-year success (intraocular pressure [IOP] < or = 21 mmHg) was 77.8% in the 5-FU group and 62.2% in the control group (P = 0.02, Wilcoxon test. Complete success (IOP < or = 21 mmHg without medications) at 5 years was lower in both the 5-FU group (72.3%) and the control group (51.3%). Postoperative mean IOP at 5 years for all successful patients was lower in eyes receiving 5-FU (10.7 +/- 3.6 mmHg vs. 16.0 +/- 6.1 mmHg [P = 0.02, t-test]). For those patients considered to be complete successes, there was no difference in IOP between the two groups of patients at any evaluated time interval. Patients in the 5-FU group were using 0.7 +/- 1.1 medications at final followup compared with 1.8 +/- 1.4 medications in the control group (P = < 0.0001, t test). Bleb-related ocular infection occurred in 6.3% of patients and was more common in patients receiving 5-FU than controls (6 of 52 vs 2 of 74, respectively; P = 0.05, Fischer's exact test). CONCLUSIONS: Adjunctive, low-dose 5-FU at the time of initial surgery in uncomplicated glaucoma improves long-term IOP control and reduces the need for postoperative, antiglaucoma therapy. Eyes receiving 5-FU are at greater risk of developing late bleb-related ocular infection. PMID- 10857842 TI - Assessment of the diurnal variation in central corneal thickness and intraocular pressure for patients with suspected glaucoma. AB - OBJECTIVE: To assess whether a single daily measurement using ultrasonic pachymetry gives a representative assessment of mean central corneal thickness (CCT) in patients with suspected glaucoma and whether diurnal changes in CCT are related to diurnal variations in intraocular pressure (IOP). DESIGN: Cross sectional study. METHOD: Central CCT and IOP were measured by a single observer in 56 eyes of 28 patients with suspected glaucoma using an ultrasonic pachymeter and a Goldmann tonometer. Four measurements were made over a 24-hour period: at 8:00 AM, 12:00 PM, 4:00 PM, and 8:00 PM. MAIN OUTCOME MEASURES: Intraocular pressure and pachymetry. RESULTS: Mean IOP was 19.80 mmHg at 8:00 AM (95% confidence interval [CI], 18.95-20.66 mmHg), 20.38 mmHg at 12:00 PM (95% CI, 19.49-21.26 mmHg), 19.91 mmHg at 4:00 PM (95% CI, 19.99-21.83 mmHg), and 19.23 mmHg at 8:00 PM (95% CI, 18.35-20.11 mmHg). Mean CCT was 569.4 microm (95% CI, 560.2-578.7 microm), 567.6 microm (95% CI, 558.4-576.7 microm), 569.1 microm (95% CI, 559.5-578.6 microm), and 567.2 microm (95% CI, 557.9-576.4 microm) at the four respective time points. There was no significant correlation between IOP and CCT in any patient (Pearson rank correlation coefficient); nor was there any significant correlation between the mean diurnal variations of IOP and CCT. CONCLUSIONS: In this group of patients with suspected glaucoma, there was no significant variation in CCT. Therefore, a single measurement of CCT is sufficient when assessing patients with suspected glaucoma. There was no correlation between change of IOP and change of CCT. PMID- 10857843 TI - Concordance of parapapillary chorioretinal atrophy in ocular hypertension with visual field defects that accompany glaucoma development. AB - OBJECTIVE: To determine whether the extent and location of progressive parapapillary chorioretinal atrophy noted in some patients with ocular hypertension are correlated with the extent and location of visual field defects that occur with progression to glaucoma. STUDY DESIGN: Retrospective cohort study. PARTICIPANTS: Thirty patients with ocular hypertension who had progressive changes of parapapillary atrophy develop before clinically detectable optic disc or visual field damage. MAIN OUTCOME MEASURES: Assessment of changes in the parapapillary atrophy and visual field parameters. METHODS: Baseline and follow up optic disc photographs and visual field test results were retrospectively analyzed. The relationship between the extent of parapapillary atrophy observed during the ocular hypertension period and initial visual field abnormalities detected after glaucoma development, as well as their spatial relationship, was statistically analyzed. RESULTS: The extent of progressive changes of the parapapillary atrophy detected during the ocular hypertension period was correlated with the extent of changes in the visual field parameters, including corrected pattern standard deviation and mean deviation measured after glaucoma development (Mantel-Haenszel chi-square test, P = 0.026, P = 0.037, respectively). In addition, the visual field abnormalities occurred in the corresponding quadrants of the progressive parapapillary atrophy. Analysis of the spatial relationship revealed that the location of progressive changes of the parapapillary atrophy was concordant with the location of visual field abnormalities in 78% of the quadrants (94 of 120 quadrants) (chi-square test, P = 0.001). CONCLUSIONS: The extent and location of visual field abnormalities that develop in ocular hypertensive eyes with progression to glaucoma exhibit a concordance with the extent and location of progressive parapapillary atrophy noted in the ocular hypertension period. This suggests the importance of detailed examination of the parapapillary area in ocular hypertensive eyes. PMID- 10857844 TI - Ultrasound biomicroscopic patterns after glaucoma surgery in congenital glaucoma. AB - OBJECTIVE: This study aimed to demonstrate specific morphologic patterns in congenital glaucoma after various surgical procedures by means of ultrasound biomicroscopy (UBM) and to investigate correlations between UBM morphology and the effectiveness of glaucoma surgery in reducing intraocular pressure. DESIGN: Observational case series. PARTICIPANTS AND INTERVENTIONS: Thirty four eyes of 18 consecutive patients, not older than 18 years, with congenital glaucoma and with a history of previous antiglaucomatous surgery underwent UBM examination of the anterior chamber angle in the treatment area and in an untreated region. MAIN OUTCOME MEASURES: The morphology of the anterior chamber angle region and the tissue reflectivity were analyzed. RESULTS: Specific UBM patterns of the anterior chamber angle in congenital glaucoma were observed after goniotomy, trabeculotomy, trabeculectomy, deep sclerectomy, and cyclodialysis. In the first months after surgery, a limited correlation was found between morphology and the success of filtering surgery. Adhesions of the iris or the ciliary processes to the trabeculectomy cleft were detected in 19 of 25 eyes after filtering procedures. CONCLUSIONS: In cases of cloudy cornea and unknown previous glaucoma surgery, UBM can be used to identify the type and localization of previous surgery in congenital glaucoma, thus assisting surgical planning for subsequent glaucoma management. The correlation between UBM morphology and the effectiveness of filtering surgery is less convincing than previously demonstrated in adults, possibly underlining the importance of individual nonsurgical factors for prognosis in congenital glaucoma. PMID- 10857845 TI - Endoscopic conjunctivodacryocystorhinostomy with Jones tube placement. AB - OBJECTIVE: Conjunctivodacryocystorhinostomy (CDCR) with Jones tube placement as described by Jones has traditionally been performed as an "open" or external procedure by means of medial canthal incision. Application of endoscopic technique for CDCR with Jones tube placement has not been well described in the peer-reviewed literature. DESIGN: Retrospective nonrandomized comparative trial. PARTICIPANTS: Ten patients with epiphora secondary to canalicular stenosis. METHODS: A total of 13 consecutive CDCR with Jones tube procedures were reviewed. Five procedures (performed predominantly in the early study period) were done by means of a traditional external approach with a medial canthal incision. Eight procedures were performed with an intranasal endoscopic approach and instrumentation with Jones tube placement under direct endoscopic visualization. MAIN OUTCOME MEASURES: Total operative time, estimated blood lost, intraoperative, and postoperative complications and need for secondary surgery were evaluated. RESULTS: All procedures were successfully completed with no intraoperative complications. Average operative time was 59 minutes in the endoscopic group and 74 minutes in the external group. Average blood loss was 3.5 ml and 4.4 ml in the endoscopic and external groups, respectively. Postoperative adjustment of tube size or position (performed as an office procedure with topical/local anesthesia) was common: five of eight endoscopic and three of five external approach. Two patients in the endoscopic group required secondary surgery for anatomic reasons. Ultimately, all cases in both groups demonstrated patent, retained Jones tubes and relief of epiphora. CONCLUSION: Endoscopic technique appears to be a reasonable approach for CDCR with Jones tube placement. Operative time and blood loss were comparable in the two groups, with the endoscopic group being slightly lower for each variable. Endoscopic Jones tube placement can be accomplished with readily available instrumentation. In this series, we did not find it necessary to use laser, radiofrequency, or monopolar devices for intranasal hemostasis. PMID- 10857846 TI - Interleukin-16. AB - Interleukin 16 (IL-16) was initially described in 1982 as the first T cell chemoattractant. Through interaction with CD4, IL-16 has now been characterized as a chemoattractant for a variety of CD4+ immune cells. Recent in vivo studies have more fully characterized IL-16 as an immunomodulatory cytokine that contributes to the regulatory process of CD4+ cell recruitment and activation at sites of inflammation in association with asthma and several autoimmune diseases. Since its cloning in 1994, IL-16 structure and function have been studied extensively. This review addresses the current data regarding IL-16 protein and gene structure; the expanding list of cells capable of generating IL-16; the direct interaction of IL-16 with its receptor, CD4; and the functional bioactivities of IL-16 as they relate to inflammation and HIV-1 infection. In addition, potential therapeutic modalities for IL-16 relating to inflammation and immune reconstitution in HIV-1 infection are also discussed. PMID- 10857847 TI - Toll receptors: an expanding role in our understanding of human disease. AB - Toll receptor proteins in Drosophila are involved in establishing the dorsal ventral axis in embryogenesis as well as participating in the innate immune response to invading pathogens. The basic mediators of this response show striking similarities in plants, insects, and vertebrates. The cytoplasmic signaling cascade is exemplified by the human interleukin-1 receptor complex (IL 1R), resulting in transcriptional activation of effector proteins through nuclear factor-kappaB (NF-kappaB). Six mammalian/human Toll-like receptors (TLR) have been described to date. The TLRs share the IL-1R cytoplasmic signaling cascade but are distinguished by their extracellular leucine-rich repeat (LRR) structure. The LRR superfamily comprises a diverse group of proteins, including a cohort involved in transmembrane signaling. Two of the human TLRs (TLR2, TLR4) have been shown to be involved in the innate response to bacterial pathogens and appear to provide a link between the innate and adaptive immune response. A better understanding of this response may provide improved therapeutic modalities in the treatment of bacterial and fungal sepsis, which continues to be a significant source of morbidity and mortality worldwide. In addition, similar to Drosophila, Toll receptors and related proteins in the LRR superfamily may also be involved in human development, as well as in noninfectious human disease. PMID- 10857848 TI - Suppression of T cell function: a potential role for transcriptional repressor ICER. AB - In this article, we review the inducible cAMP early repressor (ICER) and its possible critical involvement in modulation of T cell responsiveness by its capacity to transcriptionally attenuate interleukin-2 (IL-2) gene expression. It seems clear that the failure to produce the IL-2 is an important determinant of anergy induction. It is important that the CD28-responsive element (CD28RE), a composite DNA binding element consisting of NFAT and cyclic AMP-responsive (CRE) like motifs in position of -160 of IL-2 promoter has the high affinity for ICER binding as well as NFAT/ICER complex formation. Moreover, CD28RE with adjacent DNA sequences was also shown to be essential for conferring anergy in T lymphocytes. Because ICER does not possess a transactivation domain required for the recruitment of CBP/p300, the binding of ICER to CD28RE and/or composite motifs containing CRE-like DNA motifs may lead to uncoupling of CBP/p300 thus extinguishing IL-2 expression as well as expression of numerous other cytokines and chemokines. PMID- 10857849 TI - Transient infiltration of neutrophils into the thymus in association with apoptosis induced by whole-body X-irradiation. AB - Generally, the process of apoptosis does not cause leakage of noxious cytosolic contents and is therefore non-inflammatory. However, as previously shown, macrophages ingesting apoptotic CTLL-2 cells produced pro-inflammatory cytokines, particularly interleukin-8 (IL-8) and macrophage inflammatory protein-2 (MIP-2), a murine IL-8 homolog. This predicted that rapid and massive apoptosis may induce neutrophil accumulation in vivo. In this study, we tested this prediction by inducing apoptosis by whole-body X-irradiation in mice. After exposure to 4 Gy X ray irradiation, mice exhibited considerable apoptosis of thymic cells, which was associated with transient infiltration of neutrophils as well as MIP-2 mRNA expression. In contrast, in p53-deficient mice in which irradiation-induced apoptosis was suppressed, as has been reported, infiltration of neutrophils into the thymus was less than that found in p53+/+ mice. Taken together, these results suggest that massive and rapid apoptosis can result in infiltration of neutrophils. PMID- 10857850 TI - Hypoxemia modifies circulating and exudate neutrophil number and functional responses in carrageenin-induced pleurisy in the rat. AB - To assess the effect of hypoxemia on the responses of polymorphonuclear neutrophils (PMN) during an inflammatory response, rats were maintained in a low F1O2 atmosphere (9% O2) or room air for 12 h before intrathoracic injection of carrageenin or intradermal injections of agonists. After 4 h, hypoxemic rats had 50% more circulating PMN in blood and 25% less PMN in pleural exudate, whereas the number of PMN in skin biopsies did not differ from controls. Following hypoxemia, basal adhesion of blood PMN to serum-coated plastic wells was unchanged, whereas fMLP-stimulated adhesion was 50% greater. In contrast, basal adhesion of exudate PMN was 72% greater. In hypoxemic rats, exudate PMN produced 64% more PMA-stimulated superoxide than blood PMN; furthermore, blood and exudate PMN produced 4.5- and 2-fold more LPS-stimulated nitric oxide than controls, respectively. These results show that a moderate level of hypoxemia may trigger mechanisms that will interfere with PMN emigration yet prime these cells for enhanced responses upon stimulation. PMID- 10857851 TI - Oral feeding of an immunodominant MHC donor-derived synthetic class I peptide prolongs graft survival of heterotopic cardiac allografts in a high-responder rat strain combination. AB - The efficacy of two synthetic major histocompatibility complex (MHC)-derived DA (RT1.Aa) 25-mer peptides (residues 56-80 and 96-120) to modulate alloreactivity was tested in Lewis (RT1.A1) responder animals. The DA peptide 56-80, but not peptide 96-120, induced delayed-type hypersensitivity (DTH). DTH was significantly reduced by oral feeding of peptide 56-80, P = 0.004. In addition, oral feeding of this peptide in combination with a short course of cyclosporin A (CsA) prolonged graft survival of 60% of heterotope transplanted DA cardiac allografts in Lewis recipient rats. Long-term survivors developed low levels of allo-antibodies against donor tissue as compared to rejecting animals and increased levels of interleukin-4 (IL-4) within the allograft. Similarly, IL-4 secreting splenocytes were identified by flow cytometry in these animals, indicating a Th2-type cytokine pattern. However, graft survival was particularly limited to cardiac allografts because donor-type skin grafts were acutely rejected in tolerant animals. It is interesting that residue alignment of peptide 56-80 to the motif of the RT1.A1 molecule showed a preferred class I motif within this sequence, suggesting indirect presentation of this peptide to recipient T cells. Thus, peptide 56-80 appears to represent a dominant epitope that can be exploited for establishing tolerance in this transplantation strain combination. PMID- 10857852 TI - Dexamethasone promotes phagocytosis and bacterial killing by human monocytes/macrophages in vitro. AB - One of the actions of glucocorticoids (GC) in multiple sclerosis (MS) is an inhibitory effect on demyelination. This can be caused by a reduction in the number of infiltrating macrophages and/or by an effect on the phagocytosis of myelin. Here we investigate the effect of GC on the phagocytosis of myelin. Contrary to what was expected, we found that incubation of human monocytes with dexamethasone (DEX) for 48 h augmented (approximately threefold) the phagocytosis of myelin. This enhancement of phagocytosis by human monocytes was not restricted to myelin. Phagocytosis of various particles mediated by different macrophage receptors was increased by DEX. We found that not only the phagocytosis of Staphylococcus aureus bacteria was augmented, but also the killing of these bacteria was at least twice as effective after culture with DEX. Tumor necrosis factor alpha production of human monocyte-derived macrophages induced by lipopolysaccharide and S. aureus was suppressed by DEX. Together our results show that DEX promotes the phagocytosis of particles by human monocytes and thereby may contribute to tissue repair after immune-mediated tissue damage or infection. These data shed a new light on the clinical application of GC. PMID- 10857853 TI - Exoenzyme S from Pseudomonas aeruginosa induces apoptosis in T lymphocytes. AB - Exoenzyme S from Pseudomonas aeruginosa is a unique T cell mitogen; it is a powerful immunostimulus that activates a large proportion of T cells, but results in delayed and reduced lymphocyte proliferation. This study was performed to explain the discrepancy between early T cell activation and subsequent proliferation. Studies revealed that exoenzyme S induced rapid and unsustained surface expression of CD69, but could not induce interleukin-2 receptor alpha (IL 2R alpha) up-regulation on T cells. IL-2 was undetectable in supernatants and addition of rIL-2 could not reverse the unresponsiveness, indicating that anergy was not involved. Exoenzyme S induced membrane phosphatidylserine translocation, DNA hypodiploidy, and DNA fragmentation, implicating apoptosis as the mechanism for the unresponsiveness. Exoenzyme S-induced apoptosis shows features of both propriocidal and death by neglect, suggesting shared characteristics of an intermediate pathway. Thus, a Pseudomonas exoproduct induces T cell apoptosis, which may contribute to the pathogenesis of Pseudomonas infections in diseases such as cystic fibrosis. PMID- 10857854 TI - Alloantigenic stimulation bypasses CD28-B7 costimulatory blockade by an interleukin-2-dependent mechanism. AB - Allogeneic leukocytes have been used as biological adjuvants for T cell-specific responses to tumor and recall antigens, but the mechanisms underlying this effect have not been fully understood. The present study investigates whether alloantigen stimulation of human T cells would bypass an in vitro T cell costimulatory dysfunction induced by CTLA4Ig blockage of CD28-B7 interaction. Here, we demonstrate that costimulation with intact allogeneic leukocytes plus viral antigen circumvented the inhibition of this costimulatory pathway via interleukin-2 (IL-2) production, resulting in the generation of influenza specific cytotoxic T lymphocytes (CTL). The alloantigen-induced help for influenza-specific CTL generation did not require cell-to-cell contact between responding and allogeneic stimulator cells. These results suggest that alloantigens can be used to bypass defects in the CD28-B7 costimulatory pathway and, therefore, may contribute to understanding the mechanisms of alloantigen induced restoration of T cell-mediated immunity. PMID- 10857855 TI - Regulation of chemokine-induced transendothelial migration of T lymphocytes by endothelial activation: differential effects on naive and memory T cells. AB - Human T lymphocyte transendothelial migration (TEM) was examined in response to chemokines across cytokine-activated endothelium. Monocyte chemotactic protein-1 (MCP-1), RANTES, and macrophage inflammatory protein-1alpha (MIP-1alpha) induced TEM by memory T cells, while stromal cell-derived factor-1 (SDF-1) induced TEM by both naive and memory T cells. Tumor necrosis factor alpha (TNF-alpha) and interleukin-1 (IL-1) increased endothelial adhesion molecule (CAM) expression, whereas interferon-gamma (IFN-gamma) induced little up-regulation of CAM. However, both TNF-alpha and IFN-gamma strongly facilitated T cell migration, which was completely inhibited by pertussis toxin and both greatly increased TEM to RANTES, MIP-1alpha, and SDF-1 selectively of memory but not naive T cells. Thus, the dual selective effect on memory T cells of endothelial activation and these chemokines promotes the preferential recruitment of memory T cells to inflammatory sites. However, the enhanced chemokine-induced migration by memory T cells across activated endothelium appears to be independent of the increase in endothelial CAM expression. G-protein-linked stimuli may play an important part in T cell TEM across cytokine-activated endothelium. PMID- 10857856 TI - Involvement of glutamine, arginine, and polyamines in the action of ornithine alpha-ketoglutarate on macrophage functions in stressed rats. AB - The ability of ornithine alpha-ketoglutarate (OKG) to enhance macrophage cytotoxicity in stress situations has been described, but the mechanisms involved remain unclear. It is known that OKG administration generates glutamine (GLN), arginine (ARG), and polyamines. This study will (1) evaluate the effect of OKG on tumor necrosis factor alpha (TNF-alpha) secretion and nitric oxide (NO*) production in macrophages from glucocorticoid (DEX)-treated rats, and determine whether these effects can be reproduced by GLN or ARG supplementations, and (2) use in vivo metabolic inhibitors methionine sulfoximine (inhibitor of GLN synthetase), S-methylthiourea (inhibitor of inducible nitric oxide synthase), and difluoromethylornithine (inhibitor of ornithine decarboxylase) to assess the roles of GLN, ARG, and polyamines in OKG action. Controls received a mixture of nonessential amino acids (NEAA). GLN, ARG, and OKG all restored TNF-alpha secretion by macrophages of glucocorticoid-treated rats. The same results were obtained with GLN and ARG supplementation. However, the use of inhibitors clearly showed that OKG does not modulate TNF-alpha secretion by GLN, ARG, or polyamine pathways. We also observed that OKG enhanced NO* release by stimulated macrophages (DEX-OKG, 1.77 +/- 0.64 vs. DEX-NEAA, 0.29 +/- 0.29 nmol/ 10(6) cells, P < 0.05). Using inhibitors, it appears that this action of OKG is probably mediated via polyamine synthesis and GLN. However, an oral administration of an equimolar amount of GLN failed to reproduce the OKG-mediated effect, possibly because OKG generates more GLN in the systemic circulation than GLN itself when these substances are given orally. Our results underline the complexity of the mechanism of action of OKG, which can differ according to the functions of even a single cell type. PMID- 10857857 TI - Role of mast cell leukotrienes in neutrophil recruitment and bacterial clearance in infectious peritonitis. AB - Stimulated mast cells release a variety of chemotactic factors such as tumor necrosis factor alpha (TNF-alpha) and leukotriene B4. Recent studies have shown that mast cell-derived TNF-alpha plays a critical role in host defense against Gram negative bacterial infections by the recruitment of neutrophils to the sites of infection. In the present study, we sought to investigate if mast cells release leukotriene (LT) B4 in response to bacteria and, if so, to establish its in vivo relevance. We show that mast cells release significant amounts of LTB4 and LTC4 in response to exposure to FimH-expressing type 1 fimbriated Escherichia coli in vitro. To test the functional significance of mast cell-derived LTs during an E. coli infection in vivo, we examined the effect of a LT-synthesis inhibitor, A-63162, on bacterial clearance and neutrophil influx in an infectious peritonitis model in mast cell-deficient mice (WBB6F1-W/WV) and their normal congenic control (WBB6F1-+/+) mice. Our results show that a treatment with A 63162 reduced neutrophil influx and bacterial clearance in the peritoneal cavities of mast cell-sufficient but not -deficient mice. Thus, mast cell-derived LTs contribute to host defense by mediating early neutrophil influx and bacterial clearance at sites of infection. PMID- 10857858 TI - Characterization of activated lymphocyte-tumor cell adhesion. AB - This study demonstrates the variable expression of ICAM-1 and leukocyte function antigen-3 (LFA-3) on four tumor cell lines (COLO526, K562, Daudi, and HT-29). In addition, phorbol ester (PMA) activation of lymphocytes modulated LFA-1 from a uniform to a clustered surface distribution; whereas after treatment with high levels of Mg2+ ions, the unique epitope for high-affinity LFA-1 was identified using clone Mab24. Using a flow cytometric adhesion assay it was demonstrated that PMA-activated lymphocytes formed conjugates with COLO526 and Daudi, and that these conjugates were inhibited by anti-CD2 with varying inhibition by LFA-1 clones MHM24 and 25.3.1. When lymphocytes were induced to express the high affinity form of LFA-1, conjugates were identified with COLO526, K562, and Daudi and these conjugates were sensitive to the presence of both CD2 and LFA-1 antibodies. Further studies using confocal microscopy confirmed significant adhesion between peripheral blood lymphocytes pretreated with either PMA or high levels of Mg2+ and the adherent cell line COLO526. In conclusion, this unique study has demonstrated for the first time the important role of the active form of LFA-1 on the lymphocyte cell surface for conjugate formation with an ICAM-1 expressing tumor cell; also, two pathways of cell signaling were identified for conjugate formation to occur. PMID- 10857859 TI - Inhibition of TNF-alpha processing and TACE-mediated ectodomain shedding by ethanol. AB - Alcohol (EtOH) is a well-documented immunosuppressant. Acute EtOH-induced immunosuppression is partially due to suppression of tumor necrosis factor alpha (TNF-alpha) secretion. We investigated the mechanism of acute EtOH-induced TNF alpha suppression in two monocytic cell lines, Mono Mac 6 and DRM. EtOH inhibited TNF-alpha secretion in a dose-dependent manner. However, TNF-alpha transcription was not affected by EtOH. Enzyme-linked immunosorbent assay and confocal microscopy showed that EtOH treatment increased cell-associated TNF-alpha. Ectodomain shedding of TNF-alpha from the cell surface is mediated by TNF-alpha converting enzyme (TACE). In contrast with TNF-alpha, EtOH did not inhibit interleukin-8 (IL-8) secretion, which does not require shedding. Furthermore, TNF p75 receptor shedding, a biomarker for TACE activity, was inhibited by EtOH in both cell lines. EtOH also inhibited TNF p75 receptor shedding in TACE reconstituted fibroblasts, suggesting that EtOH inhibits the shedding process. These data show that acute EtOH exposure can posttranscriptionally suppress TNF alpha production, resulting in specific defects in immune defense. PMID- 10857860 TI - Murine macrophages differentially produce proinflammatory cytokines after infection with virulent vs. avirulent Legionella pneumophila. AB - Virulent Legionella pneumophila replicate readily in thioglycollate-elicited peritoneal macrophages from genetically permissive A/J mice, but avirulent L. pneumophila do not. The production of cytokines by macrophages infected with L. pneumophila has been studied, but the correlation of bacterial virulence with immune responses of macrophages, such as proinflammatory cytokine production, is not well understood. In this regard, production of the cytokines tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-1alpha, IL-1beta, and IL-6 were examined in macrophage cultures infected in vitro with virulent vs. avirulent L. pneumophila. Infection of macrophages from A/J mice with the virulent L. pneumophila up-regulated mRNA expression for these cytokines, whereas avirulent bacteria resulted in only a slight or no detectable increase in cytokine mRNA. Similarly, virulent L. pneumophila induced the macrophages to produce relatively high levels of TNF-alpha, IL-1alpha, IL-1beta, and IL-6 proteins as measured by enzyme-linked immunosorbent assays, whereas avirulent bacteria induced only low or often undetectable amounts of these cytokines. Thus, these results show the murine macrophages from susceptible A/J mice are readily infected with virulent L. pneumophila in vitro and stimulated to produce the proinflammatory acute-phase cytokines TNF-alpha, IL-1alpha, IL-1beta, and IL-6, but avirulent L. pneumophila did not. Such differences in induction of these proinflammatory cytokines by macrophages in response to virulent vs. avirulent L. pneumophila infections may be an important factor in the pathogenesis induced by these intracellular bacteria. PMID- 10857861 TI - Serum-induced monocyte differentiation and monocyte chemotaxis are regulated by the p38 MAP kinase signal transduction pathway. AB - Regulation by the p38 mitogen-activated protein (MAP) kinase signaling pathway of monocytic inflammatory functions was evaluated using L-790,070, a potent and selective inhibitor of p38 MAP kinase. Three major functions of monocytes were investigated: differentiation, chemotaxis, and phagocytosis. L-790,070 inhibited serum-induced monocyte differentiation with an IC50 of 0.5 nM. Monocyte chemotaxis induced by RANTES, macrophage inflammatory protein-1alpha (MIP 1alpha), monocyte chemotactic protein- (MCP-1), and fMLP were all sensitive to L 790,070. When titrated, L-790,070 inhibited MCP-1-induced chemotaxis in a concentration-dependent manner with an IC50 of 0.3 nM. However, the ability of serum-derived macrophages to phagocytose apoptotic neutrophils was unaffected by L-790,070. The concentration with which L-790,070 inhibited both differentiation and chemotaxis was similar to that necessary to inhibit p38 MAP kinase activation of MAPKAP kinase (0.3 nM) in response to stimulation by lipopolysaccharide. Therefore, the data in this report suggest that the mechanism by which L-790,070 blocked monocyte differentiation and prevented chemotaxis was by inhibiting p38 MAP kinase activity. PMID- 10857862 TI - Dexamethasone lowers cytosolic pH in macrophages by altering alkalinizing pH regulatory mechanisms. AB - The effect of dexamethasone on cytosolic pH (pHc) in resident mouse peritoneal macrophages was investigated using the fluorescent probe 2',7'-bis(carboxyethyl) 5(6)-carboxyfluorescein tetra-acetoxymethyl ester (BCECF-AM). Dexamethasone was found to significantly lower pHc and this reduction of pHc evolved gradually with time, was near maximal at 10 nM dexamethasone, and could be prevented by the glucocorticoid receptor antagonist RU-38486. The lower pHc of dexamethasone treated cells was neither due to a reduction of cellular buffer capacity nor to an altered regulation of pHc by Na+/H+-exchange or by acidifying Na+-independent Cl-/HCO3- exchange, as assessed by studies of pH recovery after acute acid and alkali loads, respectively. Instead, an impaired pHc recovery by both the H+ ATPase and the alkalinizing Na+-dependent Cl-/HCO3- exchange was observed. This impairment was most likely not caused by an altered expression or localization of the 39-kDa subunit of the proton pump. Dexamethasone treatment caused a reduction of pHc also in a HCO3--containing solution, suggesting that acid extrusion by both the H+-ATPase and Na+-dependent Cl-/HCO3- exchange is important for maintenance and regulation of macrophage resting pHc. The lowering of macrophage pHc might be one mechanism whereby glucocorticoids exert their anti-inflammatory effects. PMID- 10857863 TI - Activation of TNF-alpha transcription utilizes distinct MAP kinase pathways in different macrophage populations. AB - Stimulation of macrophages by lipopolysaccharide (LPS) leads to the rapid activation of MAP kinases (MAPK) and the subsequent induction of cytokine gene expression. We sought to determine whether LPS-inducible cytokine genes were differentially regulated in macrophages derived from different tissues. Our studies revealed that PD98059, an inhibitor of the extracellular-regulated kinase (ERK) pathway, blocked LPS-induced activation of tumor necrosis factor alpha (TNF alpha) gene expression in a murine cell line derived from alveolar macrophages but not in a nonpulmonary macrophage cell line. These findings were confirmed using primary murine alveolar and peritoneal macrophages. This suggests that the TNF-alpha promoter contains MAPK-dependent and -independent regulatory elements that are used in a cell type-specific manner. We also found that differences in MAPK-regulated signaling were not mediated by NF-KB, LITAF, Egr-1, CREB, or ATF2/ c-Jun. Together, these studies demonstrate that transcriptional activation of the TNF-alpha gene requires the ERK signaling cascade in selected macrophage populations. PMID- 10857864 TI - cAMP induces CD14 expression in murine macrophages via increased transcription. AB - CD14, a glycoprotein that binds bacterial lipopolysaccharide, plays a critical role in the inflammatory response to infection by gram-negative bacteria. Studies were undertaken to determine whether cyclic adenosine monophosphate (cAMP) regulates CD14 expression in macrophages. Incubation of RAW 264.7 cells with 8-Br cAMP resulted in a significant increase in steady-state CD14 mRNA levels. The increase in mRNA levels was also associated with both cell-associated and soluble CD14 protein. H89 completely blocked the 8-Br-cAMP-induced CD14 mRNA up regulation. There was no change in CD 14 mRNA half-life in the presence of 8-Br cAMP. The CD14 gene transcription rate was increased about twofold after exposure to 8-Br-cAMP. cAMP-dependent increases in CD14 mRNA were also observed in rat peritoneal macrophages, demonstrating that this is an authentic response of mature macrophages. This study provides evidence that cAMP and protein kinase A are important regulators of CD14 expression in macrophages. PMID- 10857865 TI - Registry based perinatal research. PMID- 10857866 TI - The Medical Birth Registry of Norway. Epidemiological research and surveillance throughout 30 years. PMID- 10857867 TI - Birthweight by gestational age in Norway. AB - OBJECTIVE: To describe birthweight by gestational age in Norway for the period 1967-1998, evaluate secular trends and provide new standards for small for gestational age for 16 to 44 weeks of gestation. SUBJECTS AND METHODS: The analyses were based on more than 1.8 million singleton births, covering all births in Norway for a 32 year period. Percentiles for birthweight by gestational age were estimated using smoothed means and standard deviations. In the preterm weeks, means and standard deviations were carefully screened for birthweight gestational age consistency, adapting a method of Wilcox and Russell. Differences in birthweight by gestational age for stillbirths and livebirths in extremely preterm weeks (16-28) are presented, and the effects of cesarean section are evaluated. We observed a clear increase in birthweight by gestational age for all term weeks, but a decrease for most of the preterm weeks over the same period. This decrease was related to the increase in deliveries by cesarean section. CONCLUSIONS: Percentiles for birthweight by gestational age are presented for clinical use, based on a current period 1987-98, covering 20-44 completed gestational weeks. In the final standards we excluded stillbirths, infants born with malformations and cesarean sections. Birthweights in the Scandinavian populations are high and standards from other populations may not be representative, especially for the term weeks. Also, the secular changes demonstrated in this study indicate that old birthweight by gestational age standards need revision, especially due to changes in obstetrical routines influencing preterm data. PMID- 10857868 TI - Birthweight percentiles by gestational age in multiple births. A population-based study of Norwegian twins and triplets. AB - OBJECTIVE: To assess secular trends for birthweight by gestational age in twins in Norway and to develop current national birthweight standards by gestational age for twin and triplet births using population-based data. MATERIAL AND METHODS: The analysis of secular trends for birthweight and gestational age in twins was based on 32,379 twin livebirths (1967-95). Taking into account the observed secular trends in birthweight for 35-40 weeks of gestation, data on twins born during 1987-95 only were included in the calculation of birthweight percentiles for 35-40 weeks, while for lower and upper weeks, data on twins born during 1967-95 were used. The construction of birthweight-for-gestation curves for triplets was based on the data on 690 triplets. RESULTS: Whereas the overall mean birthweight and gestational age decreased in 1987-95 compared with the previous years, the mean birthweights by gestational age for the 35-40 weeks of gestation was significantly higher in 1987-95. Male twins weighed more than female twins throughout the gestation with consistent and significant differences from 27 to 42 weeks of gestation. Smoothed curves for birthweight-by-gestational age percentiles of male and female twins are plotted. The birthweight-by gestational-age curves of triplets were almost identical with twin curves before 30 weeks of gestation, starting to diverge from them progressively thereafter. The intrauterine growth of twin births also starts to differ markedly from singletons at approximately 30 weeks of gestation. CONCLUSION: This study shows that plurality-specific birthweight-by-gestation standards should be used for assessment of fetal growth in multiple births rather than singleton standards. PMID- 10857869 TI - Infants' length at birth: an independent effect on perinatal mortality. AB - AIM: To investigate whether variations in birth length (crown-heel-length) were associated with perinatal mortality rate independent of birth weight. MATERIAL: The study population was singleton live- and stillbirths from 16 weeks of gestation compiled in the Medical Birth Registry of Norway from 1967 to 1997, totaling 1,705,652 births. METHOD: The total population was analyzed using z scores for length at birth, birth weight and gestational age. Variation in perinatal mortality by length at birth was studied within birth weight strata (250 g) by logistic regression. RESULTS: Perinatal mortality varied more by birth length than by birth weight or gestational age, especially for values above the population means. Within birth weight strata, the association between perinatal mortality and length was similar in all 250 g birth weight categories above 1,500 grams: mortality was lowest at birth lengths 0-2 cm below average, with mortality rates increasing exponentially in either direction. CONCLUSION: Within all birth weight strata, and adjusted for gestational age, long infants had the higher risk of perinatal death, suggesting that length at birth may be a valuable predictor when assessing the risk of perinatal mortality. PMID- 10857870 TI - Birth defects and paternal occupational exposure. Hypotheses tested in a record linkage based dataset. AB - MAIN QUESTION: To test previously established hypotheses on associations of birth defects with paternal occupation on the basis of a Norwegian registry material. METHODS: The study comprised all births in Norway 1970 -1993 for which linkage with population censuses 1970, -80 and -90 on parents' job title could be obtained--about 1 million births (75% all births). The reference population was offspring of the group that did not belong to the actual occupation. RESULTS: Vehicle mechanics had an association with hypospadias--OR 5.19 (CI 1.31-14.24), painters had a non-significant association with spina bifida--OR 2.03 (CI 0.99 3.75) and printers with club foot--OR 1.61 (CI 0.89-2.90). Associations observed previously in off-spring of fathers in large occupational groups such as teachers, drivers, electricity related occupations, sales related occupations and agricultural workers were not confirmed in this dataset. CONCLUSIONS: The study gave further evidence of cause effect relationships in the confirmed positive associations, though without any clarification of possible mechanisms involved. Possible false negative findings might be caused by low statistical power due to small occupational groups or non-differential misclassification of exposure. PMID- 10857871 TI - Maternal reproductive history: a registry based comparison of previous pregnancy data derived from maternal recall and data obtained during the actual pregnancy. AB - OBJECTIVE: To compare the quality of data on previous pregnancies, based on maternal recall, to that of actual data in previous birth records, and to quantify to which extent recall based data bias results in epidemiologic studies on adverse reproductive outcomes. METHODS: Through linkage between agricultural censuses and the Medical Birth Registry, we identified 87,113 Norwegian female farmers with 174,764 single births in 1967-1991, including 533 late abortions (16 27 weeks). Data were linked into sibship records. In routine birth records, mothers provide information at each birth on previous fetal loss. Thus, we had information both based on recall and on the actual record. RESULTS: Among all mothers with a loss documented in previous records, 73.5% had a recall of previous loss (sensitivity), while 74.3% of all mothers reporting a previous loss also had a loss in previous sibship records (positive predictive value). Late abortion after a previous loss according to the sibship record was more frequent than after a previous loss based on maternal recall (18.3 vs. 10.8 per 1,000). There was a particular lack of recall for previous losses of short gestational age or when the current birth was a late abortion; both conditions tended to deflate the association between a previous and a current adverse outcome. Grain farming was associated with late abortion in mothers with previous loss in sibship records (odds ratio, 2.6) but not among mothers without previous loss (odds ratio, 1.0). When information on previous loss was based on maternal recall the odds ratios were 1.4 and 1.3, respectively. CONCLUSIONS: Consecutive sibship records give more valid information than data derived from maternal recall. PMID- 10857872 TI - Assessing quality of obstetric care for low-risk deliveries; methodological problems in the use of population based mortality data. AB - BACKGROUND: Studies evaluating safety of different birth settings for low-risk deliveries are often difficult to interpret because of great methodological problems. OBJECTIVE: To assess potential bias in comparisons of mortality between maternity institutions with different size and level of care, particularly when using various definitions of low-risk delivery and when studying stillbirth rates. DESIGN: Population-based study. POPULATION: The population of 1.74 million births in Norway from 1967 to 1996 recorded in The Medical Birth Registry of Norway. METHODS: First we explored the problems of properly identifying low-risk deliveries from population-based data and calculated adjusted perinatal mortality rates in sub-populations by excluding different risk factors. Then we measured the difference in apparent low-risk deliveries between institutions of different size and level of care. Finally we explored bias by using stillbirths and discuss the loss of statistical power by studying only livebirths. RESULTS: The occurrence of a whole spectrum of risk factors differed between small and large institutions, even after adjustment for birthweight. Although the majority of births were from low-risk deliveries, only 1/10th of all perinatal deaths occurred in this group after admission to a maternity unit. There was a systematic difference in the reporting of time of death for stillbirths between types of institutions; the rate of stillbirths occurring during delivery was higher among small institutions, while large institutions were more often uncertain in classifying time of death for stillbirths. CONCLUSIONS: Adjustments for a large number of different risk factors, large sample-sizes and caution in including stillbirth as outcome measure are needed when comparisons of safety between different sizes of delivery units are made for low-risk pregnancies. PMID- 10857873 TI - Maternal smoking and birthweight: effect modification of period, maternal age and paternal smoking. AB - OBJECTIVE: To study the effect on birthweight of maternal smoking, and its modification by study period, maternal age and paternal smoking. DESIGN: A retrospective questionnaire based national survey comprising a random sample (n=34,799) of all mothers giving birth in Norway 1970-91. Variables studied were parental smoking during pregnancy, birthweight, maternal age and infant's year of birth. RESULTS: The overall difference in mean birthweight between non-smoking and smoking mothers was 197 g. The difference in birthweight between non-smoking and smoking mothers increased with maternal age from 182 g (<20 years of age) to 232 g (35+ years of age). There was no significant effect of paternal smoking on birthweight when the mother was a non-smoker. When the mother was a smoker and the father was a non-smoker, the birthweight, adjusted for maternal age, was reduced by 153 g (p<0.005). However, when both parents smoked, the birthweight, adjusted for maternal age, was reduced by 201 g (p<0.0005). Even though the prevalence of paternal smoking decreased by 38% during the study period, there was no significant increase in overall mean birthweight. IMPLICATION AND RELEVANCE OF RESULTS: The negative effect of maternal smoking on birthweight appears to increase with maternal age. For a non-smoking pregnant woman to live with a smoking partner has little, if any, effect on birthweight. The negative effect of paternal smoking was only observed when the mother was smoking and might reflect two possible mechanisms: (1) that a smoking mother has a greater cigarette consumption when the partner also smokes, and (2) that a smoking mother is less concerned about passive smoking than a non-smoking mother. PMID- 10857874 TI - Pregnancy complications and delivery practice in women with connective tissue disease and inflammatory rheumatic disease in Norway. AB - OBJECTIVE: To assess possible associations between inflammatory rheumatic disease and pregnancy complications/delivery practice. METHODS: In a population based study proportions were compared of obstetrical complications and interventions at delivery notified to the Medical Birth Registry of Norway during the years 1967 95 in women with (3,403) and without (671,221) rheumatic disease. RESULTS: Women with rheumatic disease had significantly higher rates of preeclampsia and cesarean section. The relative risk of preeclampsia was particularly high in women with connective tissue disease in the years 1977-86. In women with inflammatory arthritides, the relative risk of preeclampsia was particularly high during 1987-95. The relative risk of cesarean section was high in all patient groups throughout the observation period and particularly in women with connective tissue disease. CONCLUSION: High rates of preeclampsia and cesarean section in connective tissue disease pregnancies documented in a population based study emphasize the importance of monitoring and obstetrical interventions. PMID- 10857875 TI - Outcome of pregnancies subsequent to placental abruption: a risk assessment. AB - OBJECTIVE: To assess the risk of small for gestational age (SGA), preterm birth, pregnancy induced hypertension (PIH), and perinatal death in the pregnancy immediate subsequent to a placental abruption (PA) in the same mother. DESIGN: A cohort study based on the Medical Birth Registry of Norway. RESULTS: Odds ratios of SGA in subsequent PA- and non-PA deliveries were 2.8 (absolute risk = 18.5%) and 2.0 (13.9%), respectively, compared with non-PA deliveries without a history of previous PA among siblings (7.5%) after exclusion of cases with SGA in the immediate previous birth. After exclusion of cases with spontaneous preterm birth in the immediate previous delivery, odds ratios of spontaneous preterm birth in subsequent PA- and non-PA deliveries were 17.0 (36.3%) and 2.1 (6.6%), compared with non-PA deliveries without a history of previous PA among siblings (3.2%). After exclusion of cases with PIH in the immediate previous pregnancy, odds ratios of PIH in subsequent PA- and non-PA pregnancies were 2.9 (6.3%) and 1.6 (3.4%), compared with non-PA deliveries without a history of previous PA among siblings (2.3%). After adjustment for demographic variables and obstetrical complications, the increased risks persisted. CONCLUSION: A pregnancy following a PA must be considered a high risk pregnancy, not only in terms of excess risk of recurrence, but also due to excess risk of SGA, preterm birth, and PIH irrespective of recurrence of PA. Consequently, all pregnancies following a pregnancy with PA should be offered close antenatal surveillance and care. PMID- 10857876 TI - Obstetric history and the risk of placenta previa. AB - OBJECTIVE: To evaluate secular trends in the occurrence of placenta previa and whether placenta previa is associated with the outcome of previous pregnancies, cesarean section, and sociodemographic factors. DESIGN: A cohort study based on the Medical Birth Registry of Norway. Placenta previa in the second pregnancy was investigated for associations with outcomes in the first pregnancy and sociodemographic factors. RESULTS: In birth orders 1 and 2 the occurrence of placenta previa was 1.2 and 2.2 per 1,000, respectively, with no secular trend. The occurrence increased with maternal age and was lowest in women aged 20-29 years. The recurrence rate was 23 per 1,000 (adjusted odds ratio (OR) of recurrence=9.7). In women with prior delivery at < or =25 gestational weeks the risk of placenta previa was 6.7 per 1,000 (adjusted OR=3.0). In women with prior placental abruption the risk was 5.8 per 1,000 (OR=2.6). In women with prior perinatal death the risk was 4.4 per 1,000 (adjusted OR= 1.8). No independent relationship emerged with socio-economic factors, previous birthweight, and a history of pregnancy induced hypertension. Cesarean section was associated with subsequent development of placenta previa (adjusted OR= 1.3). CONCLUSIONS: We found no secular trends in the occurrence of placenta previa. Placenta previa is associated with previously described risk factors for placental abruption. The increased risk of placenta previa subsequent to placental abruption supports the theory of a shared etiologic factor. However, placenta previa and placental abruption do not share a common etiology in relation to a history of pregnancy induced hypertension, fetal growth retardation, and socio-economic factors. PMID- 10857877 TI - Secular trends in peri- and neonatal mortality in breech presentation; Norway 1967-1994. AB - OBJECTIVES: To assess secular trends in mortality rates in breech presentation in Norway and the effects of gestational age, birth defects and delivery method. MATERIAL AND METHODS: The Medical Birth Registry of Norway 1967-1994, with 45,579 breech presentation births from 24 weeks of gestation onwards, with mortality rate comprising all stillbirths from 24 completed weeks of gestation and all neonatal deaths (extended peri- and neonatal mortality) as main outcome variable. RESULTS: The extended peri- and neonatal mortality rate in breech presentation births declined during the study period from 9.2% in 1967-76 to 5.5% in 1977-86 and to 3.0% in 1987-94. The highest relative risk of mortality in breech presentation versus the total birth population was observed in intrapartum death and in mortality less than 24 hours after delivery. Stillbirth represented about half of the extended peri- and neonatal mortality throughout the study period. Also in infants with birth defects, the survival increased during the study period. The extended peri- and neonatal mortality was highest in vaginal deliveries, but decreased during the period, irrespective of delivery method. CONCLUSIONS: Probably due to improved obstetrical and neonatal care, mortality associated with breech presentation has substantially decreased. Increased focus especially on stillbirth, might be instrumental in further reducing the mortality associated with breech presentation. PMID- 10857878 TI - Knowledge and attitudes of folate, and use of dietary supplements among women of reproductive age in Norway 1998. AB - BACKGROUND: Authorities in many countries, including Norway from March 1998, recommend that women consume supplemental folate before and early in pregnancy to prevent neural tube defects. The aim of this survey was to establish Norwegian baseline data on knowledge, use and attitudes of folate and dietary supplements before implementing national campaigns on folate and pregnancy. METHOD: A telephone survey was carried out in late 1998 among 1,146 Norwegian women of reproductive age. The women were recruited from a nationally representative stratified random sample. RESULTS: Among the women aged 18-45 years, 50.4% had heard about folate, 32.9% knew about its role in pregnancy and 9.5% that it may prevent a malformation. Only 4.0% of the women knew that the critical period for folate supplementation to prevent a neural tube defect is before and early in pregnancy. The strongest determinants of knowledge were closeness to a pregnancy and educational level. Dietary supplements were used daily or almost daily by 53.3% of the women. The most commonly used types were multivitamin supplements and cod liver oil while only 0.9% of the women reported current use of supplemental folate. The women were also asked about use of folate and dietary supplements before or early in their last pregnancy: 44.3% reported that they had used a dietary supplement and 2.4% had used folate. Among the few women who had been pregnant within the last year of the interview, 10.3% reported use of a folate supplement. Overall, 56.0% of the women stated that they would use a folate supplement in a future pregnancy and 66.7% that they wanted more information about folate. CONCLUSIONS: Although about half of Norwegian women had heard about folate in 1998, just below 10% knew that it could prevent a malformation. Use of folate supplements was low in 1998, but more than half of the women stated that they would use folate supplements in a future pregnancy. PMID- 10857879 TI - Protection of privacy against protection of health. PMID- 10857880 TI - Regional node failure in patients with four or more positive lymph nodes submitted to conservative surgery followed by radiotherapy to the breast. AB - A retrospective analysis was conducted to evaluate the incidence of nodal failure in a subgroup of patients who had T1-T2 breast cancer and four or more positive nodes. Sixty-four 5 patients ranging in age from 29 to 73 years (median, 51) received conservative surgery followed by radiotherapy to the breast between November 1980 and May 1995. Adjuvant chemotherapy was administered to 56 patients, 27 of whom were also treated with tamoxifen, which was used alone in 5 patients. Three patients received no adjuvant treatment. Sixty-two patients are evaluable for regional node failure. There were 10 nodal failures, 4 in the axillary and 6 in the supraclavicular regions, in 9 patients, at a median of 56.5 and 27 months, respectively. There was no internal mammary node failure. Median follow-up was 72.6 months. The 10-year probability of developing axillary and supraclavicular failure is 13.9 +/- 7.7% and 10.5 +/- 4.1%, respectively. Prognosis was better for patients with axillary and breast recurrence and worse when relapse was in the supraclavicular region. On the basis of our results and data already published in premenopausal patients, we believe that radiotherapy to the supraclavicular region should be considered in patients with four or more positive axillary nodes, after a complete dissection. PMID- 10857881 TI - Low grade gliomas treated with adjuvant radiation therapy in the modern imaging era. AB - The purpose of this study is to evaluate tumor control and failure patterns in patients with low grade gliomas treated with surgery and conventional adjuvant radiation therapy. Twenty-eight patients with low grade gliomas (7 grade I, 21 grade II) were retrospectively evaluated. Extent of resection was gross total (3), subtotal (17), and biopsy alone (8). All grade I tumors underwent subtotal resection. Median radiation therapy dose was 54 Gy delivered to localized fields. Tumor control and patterns of failure were determined from follow-up computed tomography and/or magnetic resonance scans. Median follow-up was 86 months (range, 2.4-177 months). Thirteen patients (46%) (four grade I, nine grade II) developed tumor progression. The 5-year actuarial progression-free survival rates for grade I and grade II patients were 86% and 51%, respectively. Corresponding 5 year actuarial survival rates were 100% and 70%. All recurrences were within the treated volume. Our results reveal that conventional adjuvant radiation therapy is associated with high rates of local tumor progression in both grade II and incompletely resected grade I low grade gliomas. Alternative strategies need to be explored in these patients in an effort to improve tumor control and outcome. PMID- 10857882 TI - Recurrent malignant chondroid syringoma of the foot: a case report and review of the literature. AB - Malignant chondroid syringoma, or mixed tumor of the skin, salivary gland type, is an uncommon neoplasm believed to originate in sweat glands. This neoplasm occurs mostly in women and is typically seen in the extremities and torso. A case of recurrent malignant chondroid syringoma of the right foot in a man aged 34 years is described with a review of pertinent literature. The surgically excised neoplasm was evaluated by routine histology, immunohistochemistry, and transmission electron microscopy. The malignant chondroid syringoma showed microscopic dermal satellite tumor nodules. Immunohistochemical staining was positive for keratin and S100 and negative for actin and p53. Ki-67 showed <10% positive staining. Ultrastructurally, the neoplasm was composed of epithelial cells with tonofilaments, cell junctions, and electron-dense amorphous keratin like substance in the intercellular spaces. No evidence of myoepithelial differentiation was noted. Given the tumoral size, acral location, and histologic findings, the neoplasm was classified as a malignant chondroid syringoma. After reviewing the literature, it became apparent that wide surgical excision, adjuvant radiation therapy as well as patient education are critical in facilitating long-term survival. PMID- 10857883 TI - Induction chemotherapy followed by concurrent chemotherapy and high-dose radiotherapy for locally advanced squamous cell carcinoma of the upper-thoracic and midthoracic esophagus. AB - The purpose of this study was to evaluate the efficacy and toxicity of an induction chemotherapy schedule followed by high-dose radiotherapy and concurrent chemotherapy for locally advanced squamous cell carcinomas of the upper and midthoracic esophagus. Patients were treated with three courses of fluorouracil, leucovorin, etoposide, and cisplatin-containing induction chemotherapy followed by high-dose external beam radiotherapy to 65 Gy in 6 weeks for T4 and obstructing T3 tumors. Transversable T3 tumors received 60 Gy in 6 weeks by external radiotherapy, followed by two high-dose-rate esophageal brachytherapy fractions of 4 Gy in 5-mm tissue depth. Concurrent to radiotherapy, cisplatin and etoposide were given. Long-term survival of 22 patients was 41% and 31% at 2 and 3 years, respectively, with a median follow-up of 39 months. The probability of locoregional tumor recurrence was 60% at 3 years for all patients and 30% for those with a partial or complete response to induction chemotherapy. Acute toxicity of this schedule was moderate. Long-term survivors had a good swallowing function. This schedule offers a considerable chance of long-term survival for patients with locally advanced squamous cell carcinomas of the upper and midthoracic esophagus. Local in-field recurrences are the main risk after definitive radiochemotherapy. Dose escalation of radiotherapy is possible because of the observed low late toxicity. PMID- 10857884 TI - Primary granulocytic sarcoma of the ovary. AB - Granulocytic sarcomas are rare extramedullary tumors of malignant myeloid precursor cells. Exceedingly rare in childhood, it commonly involves skin, lymph nodes, bone, and the spine. Ovarian involvement is rare. It can arise de novo, precede the development of acute nonlymphocytic leukemia, or be the sole manifestation of relapse. We describe a 26-year-old woman with granulocytic sarcoma of the ovary without any hematologic disorder. PMID- 10857885 TI - Cytokeratin fragment 19 and squamous cell carcinoma antigen for early prediction of recurrence of squamous cell lung carcinoma. AB - Sixty patients with squamous cell carcinoma (SCC) of the lung, including 25 cases with recurrence and 35 cases without recurrence 1 year after operation, were enrolled in this study. The serial serum levels of cytokeratin fragment 19 (CYFRA 21-1) and SCC antigen were measured before operation and 1 week, 1 month, 3 months, 6 months, 9 months, and 12 months after operation for early detection of recurrence. The results revealed that 1) mean serum values of CYFRA 21-1 were significantly higher at early and any times after operation in 25 patients with recurrent SCC when compared with 35 patients without recurrent SCC; and 2) mean serum values of SCC antigen were significantly higher until 9 and 12 months after operation, in 25 patients with recurrent SCC when compared with 35 patients without recurrent SCC. We conclude that CYFRA 21-1 is a better marker than SCC antigen for early prediction of SCC recurrence in the lung. PMID- 10857886 TI - Treatment of malignant ovarian germ cell tumors with preservation of fertility: reproductive performance after persistent remission. AB - To describe our experience with malignant ovarian germ cell tumors with special reference to reproductive performance after remission, medical records of 31 patients were reviewed. The mean age at diagnosis was 18.6 years. Tumor by stage was I in 16 cases, II in 5, III in 5, IV in 3, and recurrence in 2. Histology was dysgerminoma in 7 cases, yolk sac tumor in 10, immature teratoma in 7, choriocarcinoma in 1, and mixed-type tumor in 6. Conservative surgery for fertility preservation was performed in 21 cases. Postoperative chemotherapy was given to all cases except two with stage Ia dysgerminoma. Of 31 cases, 4 including one fertility-preserved case died of disease. The other 27 cases including 20 fertility-preserved cases were successfully treated. Twenty-five cases (92.6%) have been followed longer than 60 months and 13 cases (48.1%) longer than 120 months. By the last follow-up, 8 of the 20 fertility-preserved cases delivered a total of 9 normal babies. Of the remaining 12 nonpregnant cases, 3 married, 9 have had regular menses, and 3 have had menstrual problems. Two of the latter three cases have been in hypergonadotropic anovulatory cycles. One patient has been diagnosed with tubal infertility caused by peritubal adhesion. Thus, management of the disease with fertility preservation is safe and the majority of patients can attain or retain normal ovarian function and reproductive potential. PMID- 10857887 TI - Standard off-cord lung oblique fields do not include the entire mediastinum: a computed tomography simulator study. AB - The routinely recommended target volume for off-cord lung oblique fields in the treatment of postoperative bronchogenic carcinoma includes the entire mediastinum, as defined by coverage of the contralateral mainstem bronchus and subcarinal space. However, this may be difficult to accomplish with the field angles of 20 degrees to 40 degrees, recommended in the recently completed Intergroup Trial (Radiation Therapy Oncology Group 91-05). This project was undertaken to define the oblique angle necessary to encompass the entire mediastinum as determined by computerized tomography simulator verification. Axial computerized tomography simulation images of 25 patients with non-small cell lung cancer were used in this study. Ten patients had prior lobectomy or pneumonectomy as part of their management. The contralateral mainstem bronchus, subcarinal space (SS), and the spinal cord were each contoured as separate volumes. The length of the contralateral mainstem bronchus was defined as extending from the carina to the bifurcation of the lobar bronchi. The subcarinal space was defined as a triangular space (in a coronal plane) with the carina at the apex, the mainstem bronchi superiorly, and a horizontal line 5 cm below the carina as the base of the triangle. The minimal angle to encompass the contralateral mainstem bronchus and subcarinal space, and to exclude the spinal cord was determined for each patient. The contoured volumes did not have additional margin added. The position of the carina was scored as "midline" if located in the midsagittal plane, or "off-midline" if deviated to either side from midline. Midline deviation was determined at the level of the carina to evaluate possible anatomical distortion relating to the tumor or prior surgery, and its effect on the minimal angle was assessed. The median minimal angle measured was 45 degrees (range: 28-65 degrees) for the entire group, and in 64% of those evaluated, this oblique angle was significantly greater than the 40 degrees recommended in Radiation Therapy Oncology Group guidelines (p = 0.017). In patients without midline deviation (n = 17), the median minimal angle was 45 degrees (range: 28-60 degrees), and in patients with midline deviation (n = 8), it was determined to be 44 degrees (range: 27-65 degrees), with no statistical difference noted between the two groups (p = NS). Although midline deviation was present in 4 of 10 patients previously resected, the above relationship remained unchanged. Based on computerized tomography simulation verification, off-cord oblique field angles of 20 degrees to 40 degrees do not adequately cover the entire mediastinum in most patients. To adequately encompass the entire mediastinum as defined in the Intergroup Trial (Radiation Therapy Oncology Group 91-05) with off-cord oblique fields, treatment angles greater than 40 degrees are necessary. Whether the potential increase in lung volume exposed to radiation from these larger angles results in a poorer therapeutic ratio requires further investigation. PMID- 10857888 TI - Control of cisplatin-induced emesis with intravenous ondansetron plus intravenous dexamethasone: a crossover study of triple 8-mg dose of ondansetron. AB - Two hundred seventy-five patients were enrolled in one of two arms in a crossover fashion. Arm A: three 8-mg doses of ondansetron intravenous (IV) were given at 4 hour intervals plus dexamethasone 20 mg IV from the start of chemotherapy followed by dexamethasone 5 mg IV every 12 hours. Arm B: as in arm A but with three 8-mg doses of ondansetron IV were given at 24-hour intervals substituted for ondansetron IV given at 4-hour intervals. There were 237 patients in arm A and 223 patients in arm B. Complete protection from acute and delayed vomiting/nausea obtained in arm A was 94.5%/90.3% and 71.3%/57.8%, respectively; protection obtained in arm B was 92.7%/91.0% and 71.7%/60.5%, respectively. No differences were observed in control of acute emesis after the addition of dexamethasone to ondansetron, given as either a triple 8-mg dose at 4-hour intervals or a single 8-mg dose. The triple dose of ondansetron given at 24-hour intervals was also not more effective than ondansetron given at 4-hour intervals in preventing delayed emesis when dexamethasone was added. However, the former improved control of delayed nausea on day 2. Adverse events tended to be minor, with constipation and hiccup the most common. PMID- 10857889 TI - 21-day oral etoposide for metastatic breast cancer: a phase II study and review of the literature. AB - Previous studies of etoposide for metastatic breast cancer commonly used bolus regimens given over a short period of time and included heavily pretreated patients. Results were poor. Chronic oral regimens would be expected to be superior to bolus doses based on pharmacologic studies and patients with less previous chemotherapy would be expected to have higher response rates. We studied the efficacy of oral etoposide at a dose of 50 mg/m2/day for 21 days of a 28-day cycle in good-risk patients with metastatic breast cancer. Healthy patients (Eastern Cooperative Oncology Group performance status 0, 1, or 2) who had not received chemotherapy for at least 1 year before study entry were selected for therapy. Thirty-four patients were entered; three patients were ineligible and one was cancelled. Thirty patients were available for analysis of response. One complete response and eight partial responses were documented (response rate, 30%; 95% confidence interval, 15-49%). A higher response rate was observed in those patients who never received chemotherapy compared with those who had received prior chemotherapy (57 vs. 6%, p = 0.004). There were two treatment related deaths, both owing to myelosuppression and infection. We found long-term administration of oral etoposide to have a reasonable response rate for metastatic breast cancer (30%). Our response rate was comparable to those of other published studies of long-term oral etoposide regimens for metastatic breast cancer. Response rates in single-arm studies have generally been higher for long-term oral regimens than those for bolus regimens. We also found the regimen to be significantly toxic, an observation that may be underemphasized in the earlier literature. PMID- 10857890 TI - Preoperative elevation of serum C-reactive protein is related to impaired immunity in patients with colorectal cancer. AB - The significance of a preoperative elevation of serum C-reactive protein (CRP) as an indicator of the malignant potential and prognosis in colorectal cancer is reported. The reduction of circulating lymphocytes reflects the immunosuppressive conditions of patients with neoplasms. The aim of the current study was to elucidate the significance of a preoperative elevation of serum CRP as an indicator of the impaired immunity of the patients with colorectal cancer. The subjects were 155 consecutive patients with colorectal cancer who were treated with surgical resection. The preoperative serum CRP level and the proportion of circulating lymphocytes in peripheral blood were measured and the relationship between these values was investigated. The mean value of lymphocytes percentages in patients with the preoperative elevation of serum CRP was 25.2 +/- 8.7%, which was significantly lower than that (33.4 +/- 9.3%) in patients without the preoperative elevation of serum CRP (p < 0.01). In summary, preoperative elevation of serum CRP was significantly related to the reduction of lymphocyte percentages in peripheral blood, and it can be an indicator of impaired immunity in the patients with colorectal cancer. PMID- 10857891 TI - Are the results obtained in randomized clinical trials on antiemetics sufficiently reproducible in clinical practice? AB - To evaluate whether the incidence of emesis in patients undergoing cisplatin chemotherapy and receiving the standard antiemetic prophylaxis during daily clinical practice was similar to that obtained in antiemetic trials, a prospective study was carried out, adopting very wide eligibility criteria. In the first cycle of chemotherapy, 308 consecutive adult patients were evaluated, 112 in the second, and 89 in the third cycle. Results were compared with those obtained in three published randomized clinical trials. In the first cycle of chemotherapy, complete protection from acute vomiting/nausea was obtained by 78.9% (243/308) and 71.8% (221/308) of patients. These results were quite similar to those obtained in the three randomized studies: 79.7%/72.1%, 78.3%/71.4%, and 78.7%/77.2%. No significantly different results among these studies were obtained, even in the second and third cycles of chemotherapy. In conclusion, in patients undergoing cisplatin chemotherapy, the effectiveness of the same standard antiemetic prophylaxis is similar to the efficacy found in randomized clinical trials, regardless of the eligibility/exclusion criteria and the setting of the study. PMID- 10857892 TI - Phase II trail of didemnin B in previously treated non-Hodgkin's lymphoma: an Eastern Cooperative Oncology Group (ECOG) Study. AB - Patients with non-Hodgkin's lymphoma (NHL) who fail initial therapy have a poor prognosis. We conducted a phase II study to determine the efficacy and toxicity of didemnin B, a non-myelosuppressive marine compound, in patients with NHL who relapsed or progressed after receiving one or two previous chemotherapy regimens. Fifty-one eligible patients were registered on this phase II study. Twenty-nine patients had intermediate or high grade (IG/HG) disease and 22 patients had low grade (LG) disease. Twenty-five patients received didemnin B at a dose of 6.3 mg/m2 and the remainder received 5.6 mg/m2, administered intravenously every 28 days. The patients had an Eastern Cooperative Oncology Group (ECOG) performance status of 0-2 and biopsy-proven relapsed disease. Objective responses were observed in two (7%) patients (one complete remission [CR] and one partial remission [PR]) with IG/HG disease and five (23%) patients (one CR and four PR) with LG disease. Patients with IG/HG disease had a median time to treatment failure (TTF) of 1.6 months and a median survival of 8.0 months. In contrast, the group with LG disease had a median TTF of 4.6 months and a median survival of 2.7 years. There were five grade V, 12 grade IV, and 57 grade III toxicities. Didemnin B appears to have modest activity in low grade NHL. However, the drug has considerable toxicity in this population of patients. PMID- 10857893 TI - Prophylactic administration of granulocyte colony-stimulating factor when monocytopenia appears lessens neutropenia caused by chemotherapy for lung cancer. AB - In a retrospective study, we showed that a monocyte count of <150/microl on days 6 to 8 might be a predictor of grade III or IV neutropenia during cancer chemotherapy given at 3- or 4-week intervals. In the present study, we investigated whether the administration of granulocyte colony-stimulating factor (G-CSF) when monocytopenia appears lessens neutropenia during chemotherapy for lung cancer. Between June 1997 and August 1998, 60 patients who received chemotherapy at 3- or 4-week intervals for unresectable lung cancer were randomized to receive G-CSF (2 microg/kg or 50 microg/m2) when monocytopenia (<150/microl) appeared on days 6 to 8 after chemotherapy (group A) or when neutropenia (<1,000/microl) or leukopenia (<2,000/ microl) appeared after chemotherapy (group B). The administration of G-CSF was stopped when the leukocyte or neutrophil counts reached > 10,000/microl or 5,000/microl, respectively. The blood cells counts were examined three times a week and the degree, duration, and frequency of chemotherapy-induced neutropenia of the two groups were compared. One patient in group A was excluded because whole brain irradiation during chemotherapy was required. Twenty-nine and 30 patients in groups A and B, respectively, received platinum-based chemotherapy and their chemotherapy-induced hematologic toxicities were analyzed. The mean neutrophil count nadir of group A (1,558 +/- 1,771/microl) was significantly higher than that of group B (810 +/- 639/microl, p = 0.032). The duration of grade III neutropenia in group A (1.4 +/- 1.7 days) was significantly shorter than that in group B (2.9 +/- 1.9 days, p = 0.004), and the frequency of grade III neutropenia in group A (48%) was significantly lower than that in group B (83%, p = 0.002). Infectious episodes occurred in five and eight patients in groups A and B, respectively. The durations of G-CSF therapy required by group A and B patients (4.8 +/- 3.1 vs. 4.7 +/- 2.7 days) were not significantly different. Prophylactic administration of G-CSF did not exacerbate anemia or thrombocytopenia induced by chemotherapy. We conclude that the prophylactic administration of G-CSF when monocytopenia appears can lessen neutropenia caused by chemotherapy for lung cancer without increasing the total G-CSF dose. PMID- 10857894 TI - Recruitment for a pilot case control study of oxidative DNA damage and breast cancer risk. AB - Oxidative DNA damage (ODD) can result from numerous endogenous metabolic processes as well as from exposure to environmental and dietary oxidants. One important type of ODD that may have a role in carcinogenesis is the formation of hydroxylated DNA bases. Our major purpose was to determine the potential for subject accrual for a multisite case-control study of ODD and breast cancer risk within a large urban university medical center. We examined the levels of a hydroxylated thymine residue, 5-hydroxymethyl-2'-deoxyuridine in DNA obtained from the peripheral blood of 26 women with breast cancer and an age-matched group of 29 control women without breast cancer. The isolated DNA was analyzed for levels of 5-hydroxymethyl-2'-deoxyuridine by gas chromatography with mass spectral detection. Our recruitment methods resulted in a relatively high yield of eligible cases (72%) and a lower yield of controls (46%). We evaluated the dose-response relationship of ODD level to breast cancer risk, using quartiles of ODD. The covariate-adjusted odds ratio of breast cancer exceeded 2.0 for women in the highest quartile of ODD (compared with the lowest quartile), although this result was not statistically significant. ODD levels were significantly more variable among African-American controls (SD = 224.1) than among white controls (SD = 57.5), p < 0.001. Overall, these results suggest a possible slight increase in breast cancer risk among women in the highest ODD quartile, after adjusting for race, menopausal status, and family history of breast cancer. PMID- 10857895 TI - Oxaliplatin, 5-fluorouracil, and folinic acid (Folfox) in patients with metastatic renal cell carcinoma: results of a pilot study. AB - We report the results of a chemotherapy regimen combining oxaliplatin, 5 fluorouracil, and folinic acid in patients with metastatic renal cell carcinoma. The objective of this pilot study was to define the potential efficacy of this second-line combination in patients previously treated with interleukin-2 alone or in combination with interferon alpha. Fourteen patients with metastatic renal cell carcinoma in failure after immunotherapy were included in this trial. During treatment, patients received six chemotherapy courses (Folfox regimen) administered every 2 weeks. Each cycle combined oxaliplatin day (D) D1 and folinic acid plus 5-fluorouracil D1 and D2. At completion of treatment, no objective response was observed and two patients presented stable disease. This chemotherapy schedule in patients with metastatic renal cell carcinoma previously treated with immunotherapy does not seem to be effective. PMID- 10857896 TI - Renal relapse in bilateral synchronous testicular lymphoma. AB - Bilateral synchronous testicular lymphoma is an uncommon presentation of non Hodgkin's lymphoma. A young man with non-Hodgkin's lymphoma of both testes synchronously is described here. He underwent bilateral orchiectomy, received chemotherapy, and was in complete remission when there was a relapse in the kidneys. Salvage chemotherapy was given. The patient developed progression of the residual lesion subsequently and was started on palliation with chlorambucil and prednisolone. He was alive at 34 months after diagnosis with disease. PMID- 10857897 TI - A phase III study of high-dose intensification without hematopoietic progenitor cells support for patients with high-risk primary breast carcinoma. AB - Patients with more than nine ipsilateral lymph node involvement or inflammatory breast cancer have a 5-year survival rate of approximately 50%. We studied the efficacy of high-dose intensification, comparing it with the standard dose chemotherapy for patients with high-risk primary breast cancer. Patients with inflammatory breast cancer or more than nine ipsilateral lymph node involvement without evidence of distant metastasis were randomized to receive either standard dose 5-fluorouracil, doxorubicin, and cyclophosphamide (FAC) every 3 weeks for nine courses (control) or six courses of FAC followed by two courses of cyclophosphamide (5.25 g/m2), etoposide (1,500 mg/m2), and cisplatin (165 mg/m2) (HDCVP). The study was terminated in 1998 because of slow accrual of patients. Forty-six patients were entered in the study. At 4 years, the overall survival was 72.8% (SE 11.9%) and 61.7% (SE 12.4%), and disease-free survival were 45.5% (SE 12.3%) and 33.7% (SE 11.9%) for the control and HDCVP groups, respectively (p = 0.757 and 0.720). With the small number of patients in our study, a small overall survival benefit of high-dose intensification compared with the standard therapy cannot be excluded. However, any substantial benefit is unlikely. PMID- 10857898 TI - A phase I study of liposomal doxorubicin (Doxil) with topotecan. AB - New therapies are needed for patients with advanced ovarian cancer who relapse after initial treatment with platinum and/or paclitaxel-based regimens. This study sought to determine the toxicities of combined liposomal doxorubicin (Doxil) and topotecan, and to determine a regimen for future phase II testing in ovarian cancer. Nine patients with advanced malignancies were treated with topotecan 1.0 mg/m2/day X 5 days followed by liposomal doxorubicin at a starting dose of 30 mg/m2 on day 5. Cycles were repeated every 28 days. A total of 13 cycles of therapy were administered. Grade IV neutropenia and grade IV thrombocytopenia developed in both of the two patients treated at the first dose level. Subsequent patients received only 20 mg/m2 liposomal doxorubicin. At that dose level, three patients experienced dose-limiting toxicity (one grade IV neutropenia, two grade IV neutropenia and thrombocytopenia). No responses were observed. These data indicate that the described regimen of liposomal doxorubicin and topotecan is not feasible because of excessive hematologic toxicity. Escalation to doses of liposomal doxorubicin or topotecan that have previously demonstrated antitumor activity was not possible. Future strategies to minimize such toxicity may include limiting eligibility to patients with minimal prior therapy, reducing the number of days of topotecan administration, or use of oral topotecan. PMID- 10857899 TI - Neoadjuvant chemotherapy and radiation for inoperable carcinoma of the maxillary antrum: a matched-control study. AB - A matched-control study comparing standard radiotherapy versus neoadjuvant chemotherapy and radiation was undertaken to clarify the effects of neoadjuvant systemic chemotherapy for locally advanced squamous cell carcinoma of the maxillary antrum. Thirty-four patients with inoperable maxillary cancer were treated with neoadjuvant chemotherapy and radiotherapy (Group II). Before starting radiotherapy, all patients in Group II received two or three cycles of neoadjuvant chemotherapy consisting of cisplatin and a 5-day continuous infusion of 5-fluorouracil with or without intravenous injection of vinblastine. Radiation doses ranged from 66 Gy to 75 Gy (median, 70 Gy). The response rate, patterns of failure, toxicity, and survival for Group II were compared with those for 34 stage-matched patients treated with radiation alone (Group I). Despite a higher response rate to neoadjuvant chemotherapy, the recurrence rate and patterns of treatment failure were not influenced by the addition of neoadjuvant chemotherapy. In most cases, neoadjuvant chemotherapy did not interfere with subsequent radiotherapy, and radiation-induced late complications occurred equally in both treatment groups. After a median follow-up of 48 months, there was no significant difference in 5-year actuarial survival or disease-free survival between the two treatment groups. Radiation alone for inoperable maxillary cancer was clearly suboptimal for improving local control and survival rate, but neoadjuvant chemotherapy in addition to standard radiotherapy failed to demonstrate any therapeutic advantage over radiation alone. PMID- 10857900 TI - Successful conservative treatment of neutropenic enterocolitis complicating taxane-based chemotherapy: a report of five cases. AB - Five cases of acute neutropenic enterocolitis complicating taxane-based chemotherapy are described. During a 34-month period, our department administered 4,600 courses of taxane-based (paclitaxel and docetaxel) chemotherapy to 800 cancer patients. Seven to 10 days postchemotherapy in five patients (0.1% of the given courses), neutropenic fever, abdominal pain, rebound tenderness, and grade II-IV diarrhea (bloody in two cases) developed. Two patients had oral candidiasis, and in two others septic shock developed. Computed tomography scans of the abdomen revealed in all patients thickening of the colon wall and pericolic edema, and a pericolic abscess was revealed in three of them. Both clinical and radiologic findings supported the diagnosis of acute neutropenic enterocolitis. All patients were successfully treated with broad-spectrum antibiotics and recombinant human granulocyte colony-stimulating factor. In conclusion, acute neutropenic enterocolitis is a severe complication of taxane based chemotherapy. Early diagnosis and appropriate conservative treatment leads to complete recovery. Although rare, this infection is less often associated with other chemotherapeutic regimens. PMID- 10857901 TI - Modulation of fluorouracil by methotrexate, leucovorin, and cisplatin (M-FLP) in the treatment of advanced pancreatic cancer: a phase II study of the Italian Oncology Group for Clinical Research (GOIRC). AB - The objective of this trial was to evaluate the activity and tolerability of biomodulation of 5-fluorouracil by leucovorin, methotrexate, and platinum in patients with advanced measurable disease. Thirty-five patients with histologically or cytologically proven adenocarcinoma of the pancreas were treated with methotrexate (100 mg/m2 in 500 ml 5% dextrose in a 2-hour infusion, day 1), 5-fluorouracil (800 mg/m2/day, i.v. in continuous infusion from days 2 to 5) plus 1-leucovorin (7.5 mg/m2 given per os every 6 hours, from days 2 to 5) and platinum (60 mg/m2 i.v., day 2), every 28 days. Four partial responses (12%; exact 95% confidence interval: 1-23%) were obtained in 34 evaluable patients with a median survival time of 49 weeks (range, 20-77 weeks). Ten (29%) of 34 patients had stable disease. Median time to treatment failure from the beginning of therapy was 11 weeks (range, 4-59 weeks) and median survival time was 20 weeks (range, 4-77 weeks). The most common grade III-IV toxicities were diarrhea (15%), stomatitis (41%), and vomiting (17%). Hematologic toxicity was mild. There were no therapy-related deaths. In conclusion, this trial did not report an increase or improvement in response rate and survival rates, and this regimen cannot be recommended as effective therapy for advanced pancreatic cancer. PMID- 10857902 TI - Durable clinical response of refractory hepatocellular carcinoma to orally administered thalidomide. AB - Thalidomide, a sedative agent previously associated with severe fetal malformations, has anti-angiogenic activity. We describe the antitumor effects of thalidomide in a patient with hepatocellular carcinoma that was refractory to systemic and intraarterial therapy. PMID- 10857903 TI - Stereotactic irradiation for brain metastases from ovarian carcinoma. PMID- 10857904 TI - A case of primary adenocarcinoma of the vermiform appendix. PMID- 10857905 TI - Chemical and enzymatic probing of effector-mediated changes in the conformation of a maxizyme. AB - The protein encoded by chimeric BCR-ABL mRNA causes chronic myelogenous leukemia (CML). We showed previously that a novel allosterically controllable ribozyme, of the type known as a maxizyme, can cleave this mRNA, with high specificity and high-level activity in vivo. We designed the maxizyme in such a way that it was able to form an active core with which to capture the catalytically indispensable Mg2+ ions only in the presence of the BCR-ABL mRNA junction. In order to probe the putative conformational changes, we used a weakly alkaline solution (pH 9.2) in the presence of 25 mM Mg2+ ions to hydrolyze differentially phosphodiester bonds that were located in different environments. Phosphodiester bonds in single stranded regions were clearly more susceptible to attack by alkali than those within a double-stranded helix. As indicated by earlier data obtained in vivo, our results demonstrated that the active conformation was achieved only in the presence of the junction within the chimeric BCR-ABL mRNA. Moreover, we demonstrated that the use of mild alkaline solutions to probe RNA structures is very informative. PMID- 10857906 TI - Zinc tetraruthenated porphyrin binding and photoinduced oxidation of calf-thymus DNA. AB - The photooxidation of calf-thymus DNA has been investigated in the presence of a supramolecular tetraruthenated zincporphyrin (ZnTRP) sensitizer. A strong interaction of ZnTRP with DNA has been observed, exhibiting a gradual transition from a non-specific electrostatic binding mode to a more specific one at high DNA concentrations. Formation of O2(1delta(g)) has been detected from its near infrared emission, after the excitation of ZnTRP in dioxygen-containing solutions. In the presence of DNA and dioxygen, ZnTRP promotes efficient photocatalytic oxidation of the 2'-deoxyguanosine sites, via their direct reaction with O2(1delta(g)), as in a previous work on the ZnTRP-photoinduced oxidation of the free nucleosides. PMID- 10857907 TI - Histochemical characterization of silver-induced metallothionein in rat kidney. AB - Histochemical characterizations of Ag-induced metallothionein (MT) in the kidney of the rat have been reported. Ag, Cu and Zn contents increased in kidney and liver after Ag injection. In particular, the Cu content in kidneys increased dramatically after three injections of Ag. Sephadex G-75 elution profiles of the renal cytosol of rats injected with Ag revealed that the accumulated Cu in the kidney was bound to MT as were Ag and Zn. In addition, localization of Cu- and Ag MT in the kidney was studied using autofluorescent signals, which are dependent on Cu- or Ag-thiol clusters, and immunohistochemistry. Although the MT induced by Ag was predominantly observed in the cortex of the kidney, some MT signals were also detected in the outer stripe of the outer medulla, as well as in the kidneys of LEC rats, an animal model of Wilson disease (a hereditary disorder of Cu metabolism). In these LEC rats, the Cu-MT also accumulated in the outer stripe of the outer medulla of the kidney. From these results, one possibility could explain that the Cu-MT detected in the outer stripe of the outer medulla in the kidney of Ag-injected rat was associated with the Cu transporter affected by Ag. PMID- 10857908 TI - Can the 1,5-disubstituted tetrazole ring modify the co-ordinating ability and biological activity of opiate-like peptides? AB - The copper(II) complexing ability and the biological activity of beta-casomorphin 7 tetrazole analogues have been investigated. Potentiometric and spectroscopic (UV-Vis, CD and EPR) studies have been used to establish the thermodynamic stability, speciation and structure of Cu(II) complexes with YP-psi(CN4)-FPGPI NH2 (1), YPF-psi(CN4)-AGPI-NH2 (2) and YPFP-psi(CN4)-GPI-NH2 (3). Comparison of the binding ability of the tetrazole analogues reveals that the most effective ligand for copper(II) is YPF-psi(CN4)-AGPI-NH2. The effectiveness of this ligand comes from its particular conformation suited for the Cu(II) 2N co-ordination mode in the physiological pH region. The ability of casomorphin tetrazole analogues to activate rat mast cells to histamine release in vitro in the presence of copper(II) has been studied. PMID- 10857909 TI - Synthesis, characterization and DNA binding of ruthenium(II) complexes containing the atatp ligand. AB - Acenaphtheno[1,2-b]-1,4,8,9-tetraazatriphenylene (atatp) and its complexes [Ru(L)2atatp](ClO4)2 x nH2O (L = 2,2'-bipyridine (bpy), n=2 (1); 1,10 phenanthroline (phen), n=2 (2); and 2,9-dimethyl-1,10-phenanthroline (dmp), n=1 (3)) have been synthesized and characterized by elemental analyses and 1H NMR. The spectral and electrochemical properties of these complexes are also examined. Complexes 1 and 2 display bright luminescence in acetonitrile but very weak luminescence in water solution. However, complex 3 is not luminescent in either solvent. The interaction of the complexes with calf thymus DNA (CT-DNA) has been studied by absorption, emission and viscosity measurements. The intrinsic binding constants of complexes 1 and 2 are 7.6 x 10(4) and 8.8 x 10(4) M(-1) respectively. The relatively low affinities of complexes 1 and 2 with DNA may arise from the atatp ligand, indicating that the size and shape of the intercalated ligand have a marked effect on the strength of interaction. Complexes 1 and 2 bind with CT-DNA in an intercalative mode but complex 3 in a non-intercalative one, showing that changing the ancillary ligand affects not only the binding magnitude, but also the binding mode of the interaction. PMID- 10857910 TI - Cis- and trans-conformations for peroxynitrite anions. AB - The biochemically important anion, peroxynitrite, is commonly thought to have a cis-conformation which is stable. This is thought to have a cyclic structure, which blocks the formation of the trans-conformation. Furthermore, it is often suggested that the latter structure isomerises to the far more stable nitrate ion. Here I suggest that the cis-structure has comparable stability to the trans structure, and they are in rapid equilibrium at room temperature. This is supported by the very large linewidths for certain vibrational bands at room temperature, which become narrow, well-separated bands at 4 K. Lifetimes are about 10(-12) s for each isomer. PMID- 10857911 TI - Synthesis, chelating properties towards gallium and biological evaluation of two N-substituted 3-hydroxy-4-pyridinones. AB - Two N-substituted 3-hydroxy-4-pyridinones (1-(3'-aminopropyl)-3-hydroxy-2-methyl 4-pyridinone (L1) and 1-(2'-carboxyethyl)-3-hydroxy-2-methyl-4-pyridinone (L2)) were prepared through one- and three-step reactions, respectively. The pKa values of the ligands and the stability constants of their Ga(III) complexes have been determined. Both the complexes are strongly coordinated to three (O,O) hydroxypyridonate moieties. There is a clear effect of the N-substituents in the lipophilic-hydrophilic balance and in the Ga(III) binding interaction; the acid derivative (L2) has lower lipophilicity but higher chelating strength than the amine derivative (L1). Both chelators are shown to interfere in the typical biological behavior of 67Ga-citrate in mice: L1 enhanced the urinary excretion leading to an increased 67Ga removal from the soft tissue, while L2 induced a lower blood clearance with a pronounced bone uptake mainly at 48 h after injection, thus suggesting that the 67Ga-L2 complex could have potential interest as a bone imaging agent. PMID- 10857912 TI - A 1H NMR study of the oligonucleotide binding of [(en)Pt(mu-dpzm)2Pt(en)]Cl4. AB - The non-covalent binding of [(en)Pt(mu-dpzm)2Pt(en)]4+ to the dodecanucleotides d(CGCGAATTCGCG)2 and d(CAATCCGGATTG)2 has been studied by 1H NMR spectroscopy in order to gain a greater understanding of the pre-covalent binding association of cationic dinuclear platinum(II) anti-cancer drugs. NOESY experiments showed that the metal complex bound in the minor groove at the A/T rich regions of both dodecanucleotides. The metal complex did not induce any major DNA conformational changes. However, given the relative dimensions of the DNA minor groove and the metal complex, it is reasonable to expect that the metal complex binding significantly widens the minor groove at the A/T rich binding sites. The results of this study suggest that although dinuclear platinum(II) anti-cancer drugs covalently bind at GC sequences in the DNA major groove, they will preferentially associate with AT sequences in the minor groove before the covalent binding. PMID- 10857913 TI - Synthesis, X-ray structure, and anti-leukemic activity of oxovanadium(IV) complexes. AB - In a systematic effort to identify and develop effective anticancer agents, four oxovanadium(IV) complexes with 1,10-phenanthroline (Phen) or 4,7-dimethyl-1,10 phenanthroline (Me2-Phen) as ligand(s) were synthesized and characterized. Among the four oxovanadium(IV) complexes synthesized, the crystal structure of the bis(phenanthroline)oxovanadium(IV) complex bis(1,10 phenanthroline)sulfatooxovanadium(IV) ([VO(SO4)(Phen)2], compound 1) has been determined. Compound 1 crystallized in the space group P2(1)/n with unit cell parameters a = 14.2125(17) A, b = 10.8628(13) A, c = 20.143(2) A, alpha = 90 degrees, beta = 102.569(2) degrees, gamma = 90 degrees, V = 3035.3(6) A3, and Z = 4. The refinement of compound 1 by full-matrix least-squares techniques gave an R factor of 0.0785 for 4356 independent reflections. The structure contains two enantiomorphous molecules, lambda and delta, which are related by an inversion center. Compound 1 exhibited 3.5-fold more potent cytotoxic activity against NALM 6 human leukemia cells than the mono(phenanthroline)oxovanadium(IV) complex (diaqua)(1,10-phenanthroline)sulfatooxovanadium(IV) ([VO(SO4)(Phen)(H2O)2], compound 2) (IC50 values: 0.97+/-0.10 microM versus 3.40+/-0.20 microM: P=0.0004). Methyl substitution in the phenanthroline ligand enhanced the anti leukemic activity of the mono(phenanthroline)oxovanadium(IV) complex 4.4-fold (IC50 values: 0.78+/-0.10 microM, compound 4, versus 3.40+/-0.20 microM, compound 2; P=0.0003) and the anti-leukemic activity of the bis(phenanthroline)oxovanadium(IV) complex 5.7-fold (IC50 values: 0.17+/-0.02 microM, compound 3, versus 0.97+/-0.10 microM, compound 1; P=0.001). The leading oxovanadium compound, bis(4,7-dimethyl-1,10-phenanthroline)sulfatooxovanadium(IV) ([VO(SO4)(Me2-Phen)2], compound 3) triggered the production of reactive oxygen species (ROS) in human leukemia cells, caused G1-arrest and inhibited clonogenic growth at nanomolar concentrations. PMID- 10857914 TI - Model studies of the precipitation of silica in the presence of aluminium; implications for biology and industry. AB - The unique chemical affinity between the oxides of silicon and aluminium has been cited as a potential route for the amelioration of the detrimental effects of aluminium in the environment and in biological systems. A greater understanding of silicon-aluminium interactions may assist in this endeavour and also provide a means of overcoming silica fouling problems encountered by industry which are exacerbated by the presence of aluminium. It is also conceivable that this increased knowledge may demonstrate a positive use for aluminium in the processing of the silicon dioxide phase. In this study we report the effect of aluminium ions, derived from aluminium chloride, on silicic acid species obtained from potassium catecholato complexes of silicon at circumneutral pH at the molar ratios 1000Si:Al, 100Si:Al and 50Si:Al. Silica and low levels of aluminium-rich silica materials were formed with Si:Al ratios of about 3.5:1 comparable with the element ratios detected in senile plaques and aluminium-rich scale. A kinetic study showed that aluminium in the reaction medium slowed down the rate of formation of one of the silica species formed early in the condensation process, e.g. trimers, but increased the rate at which silicic acid was removed from sub 1 nm diameter particles. The materials precipitated in the presence of aluminium were composed of smaller particles and aggregates with smaller pores (Si100:Al and Si50:Al systems) or larger pores (Si1000:Al) compared to the control. The nature of the interactions responsible for these differences is discussed. The effects described here demonstrate the ability of silica and aluminium to interact under conditions such as those found in biological systems. That silica reacts with aluminium in the presence of catechol supports the protective role assigned to silicon. PMID- 10857915 TI - Studies on the interactions between human serum albumin and trans-indazolium (bisindazole) tetrachlororuthenate(III). AB - The interactions between HInd[RuInd2Cl4] and human serum albumin have been investigated through UV-Vis, circular dichroism (CD), fluorescence spectroscopy and the inductively coupled plasma-atomic emission spectroscopy (ICP(AES)) method. Binding of Ru(III)-indazole species to albumin has strong impact on protein structure and it influences considerably albumin binding of other molecules like warfarin, heme or metal ions. The metal complex-human serum albumin (HAS) interactions cause conformational changes with loss of helical stability of the protein and local perturbation in the domain IIA binding pocket. The relative fluorescence intensity of the ruthenium-bound HSA decreased, suggesting that perturbation around the Trp 214 residue took place. This was confirmed by the destabilization of the warfarin-binding site, which includes Trp 214, observed in the metal-bound HSA. PMID- 10857916 TI - Synthesis, crystal structure, spectral properties and cytotoxic activity of platinum(II) complexes of 2-acetyl pyridine and pyridine-2-carbaldehyde N(4) ethyl-thiosemicarbazones. AB - The reactions of Na2PtCl4 with pyridine-2-carbaldehyde and 2-acetyl pyridine N(4) ethyl-thiosemicarbazones, HFo4Et and HAc4Et respectively, afforded the complexes [Pt(Fo4Et)Cl], [Pt(HFo4Et)2]Cl2, [Pt(Fo4Et)2] and [Pt(Ac4Et)Cl], [Pt(HAc4Et)2]Cl2 x 2H2O, [Pt(Ac4Et)2]. The new complexes have been characterized by elemental analyses and spectroscopic studies. The crystal structure of the complex [Pt(Ac4Et)Cl] has been solved. The anion of Ac4E coordinates in a planar conformation to the central platinum(II) through the pyridyl N, azomethine N and thiolato S atoms. Intermolecular hydrogen, non-hydrogen bonds, pi-pi and weak Pt pi contacts lead to aggregation and a supramolecular assembly. The cytotoxic activity for the platinum(II) complexes in comparison to that of cisplatin and thiosemicarbazones was evaluated in a pair of cisplatin-sensitive and -resistant ovarian cancer cell lines A2780 and A2780/Cp8. The platinum(II) complexes showed a cytotoxic potency in a very low micromolar range and were found able to overcome the cisplatin resistance of A2780/Cp8 cells. PMID- 10857917 TI - Metal(II) binding ability of a novel N-protected amino acid. A solution-state investigation on binary and ternary complexes with 2,2'-bipyridine. AB - The binary and ternary systems 2,2'-bipyridine (bpy)-M(II)-NO2psglyH2 (M(II) = Mn(II), Co(II), Ni(II), Cu(II), Zn(II), Cd(II), Pb(II); NO2psglyH2 = N-(2 nitrophenylsulfonyl)glycine) were investigated in aqueous solution by means of potentiometry and electron spectroscopy in order to identify the type, number and stability of the complex species as a function of pH and metal-to-ligand molar ratio. The aim is to evaluate the effect of a substituent on the phenyl ring of the N-sulfonyl amino acids on their coordination properties. The prevailing species in the binary systems is the [ML] (M = Co(II), Ni(II), Cu(II), Cd(II), Pb(II)) where the amino acid molecule is in the dianionic form and coordinates the metal ion through both carboxylic oxygen and deprotonated sulfonamidic nitrogen, while in the Mn(II)- and Zn(II)-containing binary system the only complex species found are those with the amino acid in the monoanionic form. In the ternary 2,2'-bipyridine-containing systems the chelating coordination mode of the dianionic amino acid is maintained with M(II) = Co(II), Ni(II), Cu(II), Cd(II), Pb(II) and the addition of the aromatic base also enables the Zn(II) ion to substitute for the sulfonamide nitrogen-bound hydrogen of NO2psglyH2. PMID- 10857918 TI - Synthesis, crystal structure and antimicrobial activities of two isomeric gold(I) complexes with nitrogen-containing heterocycle and triphenylphosphine ligands, [Au(L)(PPh3)] (HL = pyrazole and imidazole). AB - Two isomeric gold(I)-triphenylphosphine complexes with nitrogen-containing heterocycles, [Au(L)(PPh3) (HL = pyrazole (1), imidazole (2)) were isolated as colorless cubic crystals for 1 and colorless plate crystals for 2, respectively. The crystal structures of 1 and 2 were determined by single-crystal X-ray diffraction. These complexes were also fully characterized by complete elemental analyses, thermogravimetric/differential thermal analyses (TG/DTA) and FT-IR in the solid state and by solution NMR (31P, 1H and 13C) spectroscopy and molecular weight measurements in acetone solution. These complexes consisted of a monomeric 2-coordinate AuNP core both in the solid state and in solution. The molecular structures of 1 and 2 were compared with those of related gold(I) complexes, [Au(1,2,3-triz)(PPh3)] (3, Htriz = triazole), [Au(1,2,4-triz)(PPh3)]2 (4) as a dimer through a gold(I)-gold(I) bond in the solid state, and [Au(tetz)(PPh3)] (5, Htetz = tetrazole). Selective and effective antimicrobial activities against two gram-positive bacteria (B. subtilis, S. aureus) and modest activities against one yeast (C. albicans) found in these gold(I) complexes 1-4 are noteworthy, in contrast to poor activities observed in the corresponding silver(I) complexes. PMID- 10857919 TI - Reactions of Azotobacter vinelandii nitrogenase using Ti(III) as reductant. AB - Nitrogenase-catalyzed reactions using Ti(III) were examined under a wide variety of conditions to determine the suitability of Ti(III) to serve as a general nitrogenase reductant. Solutions prepared from H2-reduced TiCl3, aluminum-reduced TiCl3, TiCl2, evaporated TiCl3 from an HCl, solution, and TiF3 were evaluated as reductants. Three general types of reactivity were observed. The first showed that, below Ti(III) concentrations of about 0.50 mM, nitrogenase catalysis utilized Ti(III) in a first-order reaction. The second showed that, above 0.50 mM, the rate of nitrogenase catalysis was zero order in Ti(III), indicating the enzyme was saturated with this reductant. Above 2.0-5.0 mM, nitrogenase catalysis was inhibited by Ti(III) depending on the titanium source used for solution preparation. This inhibition was investigated and found to be independent of the buffer type and pH, while high salt and citrate concentrations caused moderate inhibition. [Ti(IV)] above 2.0-3.0 mM and [Ti(III)] above about 5.0 mM were inhibitory. ATP/2e values were 4-5 for [Ti(III)] at or below 1.0-2.0 mM, 2.0 from 5.0 to 7.0 mM Ti(III) where nitrogenase is not inhibited, and 2.0 above 7.0 mM Ti(III) where severe inhibition occurs. For nitrogenase-catalyzed reactions using Ti(III) as reductant, the potential of the solution changes with time as the Ti(III)/Ti(IV) ratio changes. From the change in the rate of product formation (Ti(III) disappearance) with change in solution potential, the rate of nitrogenase catalysis was determined as a function of solution potential. From such experiments, a midpoint turnover potential of -480 mV was determined for nitrogenase catalysis with an associated n = 2 value. PMID- 10857920 TI - Metalloporphyrin mediated DNA cleavage by a low concentration of HaeIII restriction enzyme. AB - The plasmid DNA scission by the restriction enzyme HaeIII was investigated in the presence of tetrakis(1-methylpyridinium-4-yl)porphyrin (H2TMPyP) and its manganese(III), iron(III), nickel(II), cobalt(III) and zinc(II) derivatives. The effect of metalloporphyrins on plasmid DNA cleavage was ascertained by gel electrophoresis. UV-Vis absorption spectroscopy and circular dichroism (CD) spectroscopy. In the absence of the metalloporphyrins, plasmid DNA scission did not occur in the presence of a low concentration of HaeIII (0.2 units microL(-1)) at 37 degrees C after 1 h incubation. However, DNA cleavage occurred in the presence of the metalloporphyrins and HaeIII (0.2 units microL(-1)) at 37 degrees C after 1 h incubation. Gel electrophoresis results indicate the catalytic effect of metalloporphyrins (Mn(III)-, Fe(III)-, Co(III)- and Zn(II)TMPyP) by binding to both DNA and the enzyme through electrostatic interaction, which was confirmed by the change in UV-Vis and CD spectra. A mechanism for the enhanced DNA cleavage is proposed. PMID- 10857921 TI - Antimicrobial effects of Finnish plant extracts containing flavonoids and other phenolic compounds. AB - Plant phenolics, especially dietary flavonoids, are currently of growing interest owing to their supposed functional properties in promoting human health. Antimicrobial screening of 13 phenolic substances and 29 extracts prepared from Finnish plant materials against selected microbes was conducted in this study. The tests were carried out using diffusion methods with four to nine microbial species (Aspergillus niger, Bacillus subtilis, Candida albicans, Escherichia coli, Micrococcus luteus, Pseudomonas aeruginosa, Saccharomyces cerevisiae, Staphylococcus aureus and Staphylococcus epidermidis). Flavone, quercetin and naringenin were effective in inhibiting the growth of the organisms. The most active plant extracts were purple loosestrife (Lythrum salicaria L.) against Candida albicans, meadowsweet (Filipendula ulmaria (L.) Maxim.), willow herb (Epilobium angustifolium L.), cloudberry (Rubus chamaemorus L.) and raspberry (Rubus idaeus L.) against bacteria, and white birch (Betula pubescens Ehrh.), pine (Pinus sylvestris L.) and potato (Solanum tuberosum. L.) against gram positive Staphylococcus aureus. PMID- 10857922 TI - Efficacy of allyl isothiocyanate in killing enterohemorrhagic Escherichia coli O157:H7 on alfalfa seeds. AB - Volatile compounds occurring in the essential oil of plants were tested for their efficacy in killing Escherichia coli O157:H7. Experiments using an agar disk assay revealed that exposure of the pathogen to 50 microl of eugenol, carvacrol, linalool, or methyl jasmonate in a 950-cc jar at 20, 37 or 47 degrees C for up to 48 h failed to inhibit colony formation. However, exposure to 8 microl of allyl isothiocyanate (AIT) (equivalent to 8.4 ppm in the air within the jar, if completely volatilized) resulted in more than a 7-log10 reduction in population of E. coli O157:H7 at 37 degrees C within 48 h; significant (P < or = 0.05) reduction in populations also occurred in the presence of 4 microl of AIT compared to 2 microl, which had no lethal affect. At 20 degrees C, the lethality of AIT was substantially less, although significant reduction occurred when disks were exposed to 8 or 10 microl of AIT compared to 4 or 6 microl and when exposed to 4 or 6 microl compared to 2 microl. Treatment with 10 microl of AIT for 5 h at 47 degrees C resulted in death of 6 log10 of E. coli O157:H7. The efficacy of AIT in killing E. coli O157:H7 on dry and wet alfalfa seeds was investigated. The pathogen, at an initial population of 2.7 log10 cfu/g of seed, was not recovered by direct plating (< 0.7 log10 cfu/g) or enrichment of wet seeds exposed to 50 microl of AIT/950-cc jar for 24 h at 37 or 47 degrees C. Exposure of dry seeds containing 2.9 log10 cfu of E. coli O157:H7 per g to an atmosphere containing 100 microl of AIT/950-cc jar (ca. 105 ppm AIT if completely volatilized) for 24 h at 47 degrees C did not eliminate viable E. coli O157:H7 cells. Unfortunately, the enhanced effectiveness of AIT in killing the pathogen on wet alfalfa seeds is offset by a dramatic reduction in seed viability. Nevertheless, the use of AIT as an alternative to chlorine for the purpose of killing E. coli O157:H7 and perhaps other pathogens on alfalfa seed holds promise. Factors that may influence conditions rendering increased sensitivity of E. coli O157:H7 to AIT without compromising seed viability should be investigated. PMID- 10857923 TI - Amplified fragment length polymorphism (AFLP) analysis of Clostridium perfringens for epidemiological typing. AB - Thirty-five Clostridium perfringens isolates from patients and foods implicated in seven outbreaks of suspected Cl. perfringens food poisoning together with five unrelated incidents were analysed by serotyping and amplified fragment length polymorphism (AFLP). Despite minor band differences, AFLP was found to be highly reproducible and 16 different profiles (each unique to the 12 incidents) were recognised. The results from both serotyping and AFLP analysis identified exactly the same groups of related cultures. It is concluded that AFLP can provide a rapid, sensitive and reproducible method for the typing of Cl. perfringens for outbreak investigation. PMID- 10857924 TI - Mathematical modelling of the growth rate and lag time for Listeria monocytogenes. AB - Growth data for Listeria monocytogenes were collected from the literature and a global model built with existing secondary models describing independently the effects of environmental factors on the growth rate and lag time was based on these data. The growth rates calculated with this model were consistent with the published ones but the fit was poor near the limits of growth of the micro organism. The model was also less accurate to describe the lag time. It seems then that reliable predictions of the growth rate of L. monocytogenes could be obtained in a wide range of growth conditions, but models should take into account interactions between environmental factors. Furthermore, it is necessary to better model the lag phase duration and particularly to model the effect of the history of the inoculum on the subsequent lag time. PMID- 10857925 TI - Modelling the growth rate of Listeria monocytogenes with a multiplicative type model including interactions between environmental factors. AB - A multiplicative secondary model previously published to describe independently the effects of environmental factors on the growth rate of Listeria monocytogenes (Augustin and Carlier, 2000) was improved by taking into account interactions between these environmental factors. The proposed model allowed to decrease the rate of fail-safe growth predicted from 13.5% to 12.1% and the rate of fail dangerous no growth predicted from 16.1% to 7.1%. PMID- 10857926 TI - Microgradients in bacterial colonies: use of fluorescence ratio imaging, a non invasive technique. AB - Fluorescence ratio imaging is a non-invasive technique for studying the formation of microgradients in immobilised bacterial colonies. These gradients can be quantified easily when combined with the gel cassette system designed at the Institute of Food Research, Norwich, UK. Colonies of Lactobacillus curvatus were observed using this technique and relevant pH gradients were present when the colonies reached a diameter of about 100 microm. These pH gradients were due to production of lactic acid by L. curvatus cells in the colonies. The spatial resolution of the images was about 1.5 microm (scale of bacterial cells) and therefore very suitable for observing local effects in colonies which ranged in sizes from 1 to 500 microm. PMID- 10857927 TI - Potentiation of the lethal effect of peroxygen on Bacillus cereus spores by alkali and enzyme wash. AB - Bacillus cereus present in pipes and heat-exchangers represents a potential quality problem for dairy industry. The peroxygen-containing disinfectants investigated had only negligible sporicidal effect when applied at the recommended in-use temperature and concentration. However, cleaning agents used before disinfection potentiated their lethal activity. Pre-exposure of B. cereus spores to 1% sodium hydroxide at temperatures over 40 degrees C increased the sporicidal effect of the peroxygen-containing disinfectant. The effect was dependent on the alkali concentration and the temperature. Also, a significant potentiating activity of an enzyme-based cleaning agent was obtained, but the effect was smaller than for alkali treatment. The results indicated that disinfectants based on peroxygen can be used to eliminate B. cereus spores at non corrosive temperatures and concentrations if the surfaces are cleaned with alkali or enzyme-based disinfectants prior to disinfection. PMID- 10857928 TI - Safety assessment of potential probiotic lactic acid bacterial strains Lactobacillus rhamnosus HN001, Lb. acidophilus HN017, and Bifidobacterium lactis HN019 in BALB/c mice. AB - The general safety of immune-enhancing lactic acid bacteria (LAB) strains Lactobacillus rhamnosus HN001 (DR20), Lb. acidophilus HN017, and Bifidobacterium lactis HN019 (DR10) was investigated in a feeding trial. Groups of BALB/c mice were orally administered test LAB strains or the commercial reference strain Lb. acidophilus LA-1 at 2.5 x 10(9), 5 x 10(10) or 2.5 x 10(12) colony forming units (CFU)/kg body weight/day for 4 weeks. Throughout this time, their feed intake, water intake, and live body weight were monitored. At the end of the 4 week observation period, samples of blood, liver, spleen, kidney, mesenteric lymph nodes, and gut tissues (ileum, caecum, and colon) were collected to determine: haematological parameters (red blood cell and platelet counts, haemoglobin concentration, mean corpuscular volume, mean corpuscular haemoglobin, and mean corpuscular haemoglobin concentration); differential leukocyte counts; blood biochemistry (plasma total protein, albumin, cholesterol, and glucose); mucosal histology (epithelial cell height, mucosal thickness, and villus height); and bacterial translocation to extra-gut tissues (blood, liver, spleen, kidney and mesenteric lymph nodes). DNA finger printing techniques were used to identify any viable bacterial strains recovered from these tissues. The results demonstrated that 4 weeks consumption of these LAB strains had no adverse effects on animals' general health status, haematology, blood biochemistry, gut mucosal histology parameters, or the incidence of bacterial translocation. A few viable LAB cells were recovered from the tissues of animals in both control and test groups, but DNA fingerprinting did not identify any of these as the inoculated strains. The results obtained in this study suggest that the potentially probiotic LAB strains HN001, HN017, and HN019 are non-toxic for mice and are therefore likely to be safe for human use. PMID- 10857929 TI - The use of reverse transcription-polymerase chain reaction (RT-PCR) for monitoring aflatoxin production in Aspergillus parasiticus 439. AB - A detection system based on reverse transcription PCR (RT-PCR) has been developed to monitor aflatoxin gene expression in Aspergillus parasiticus. Total RNAs of aflatoxigenic A. parasiticus 439 grown in aflatoxin permissive and non-permissive media were amplified and monitored over time by RT-PCR with specific primers designed from two genes of the aflatoxin biosynthetic pathway. Gene transcription in both media was assessed by monitoring the house keeping beta-tubulin gene and aflatoxin production was correlated with transcription by thin layer chromatography. This RT-PCR technique has the potential to be employed as a tool to investigate the effects of a variety of physiological factors on the transcription of the aflatoxin genes. PMID- 10857930 TI - Ochratoxin formation in Aspergillus ochraceus with particular reference to spoilage of coffee. AB - Production of ochratoxin on media by eight isolates of Aspergillus ochraceus from coffee or its processing environment in India, Indonesia, Kenya, and Brazil, and seven Brazilian isolates from other commodities, has been compared with yields in shaken fermentation on shredded wheat and coffee (Coffea arabica). Shredded wheat most consistently allowed expression of biosynthesis of ochratoxins A and B in yields up to 3.5% of the dry product. Culture on artificial media was an unreliable predictor of ochratoxin yield on both shredded wheat and coffee. Coffee was a relatively poor substrate for ochratoxin production particularly when sterilised. Notably, two Asian coffee isolates produced 400 mg kg(-1) ochratoxin A on unsterilised ground green coffee, showing this to be a preferred substrate for further experimentation. The study focused on isolates of A. ochraceus, which from evidence of culture on media would not be expected to be suitable fungi for future studies to establish both the fact of spoilage of coffee by A. ochraceus and the dynamics of ochratoxin formation by isolates of this species. PMID- 10857931 TI - Increasing incidence of type 2 diabetes in the third millennium: is abdominal fat the central issue? PMID- 10857932 TI - Meeting American Diabetes Association guidelines in endocrinologist practice. AB - OBJECTIVE: To determine whether American Diabetes Association (ADA) guidelines can be met in the context of routine endocrinology practice. RESEARCH DESIGN AND METHODS: Charts were reviewed for a group of patients who were examined in 1998, followed for > or = 1 year, and had two or more visits during that year. Process measures and metabolic outcomes were studied for patients with type 2 diabetes, and glycemic control was assessed for patients with type 1 diabetes. RESULTS: A total of 121 patients with type 2 diabetes had a mean age of 63 years, a mean BMI of 31 kg/m2, and a mean duration of diabetes of 12 years. Many had comorbidities or complications: 80% had hypertension, 64% had hyperlipidemia, 78% had neuropathy, 22% had retinopathy, and 21% had albuminuria. Management of type 2 diabetic patients was complex: 38% used oral hypoglycemic agents alone (54% of these were using two or more agents), 31% used oral hypoglycemic agents and insulin, and 26% used insulin alone; 42% of patients taking insulin therapy injected insulin three or more times per day. Within 12 months, 74% of patients had dilated eye examinations, 70% had lipid profiles, and 55% had urine albumin screening. Of the patients, 87% had a foot examination at their last visit. Blood pressure levels averaged 133/72 mmHg, cholesterol levels averaged 4.63 mmol/l, triglyceride levels averaged 1.99 mmol/l, HDL cholesterol levels averaged 1.24 mmol/l, and LDL cholesterol levels averaged 2.61 mmol/l. Random blood glucose levels averaged 8.0 mmol/l, and HbAlc levels averaged 6.9 +/- 0.1%. A total of 87% of patients had HbAlc levels < or = 8.0%. A total of 30 patients with type 1 diabetes had mean age of 44 years, a mean BMI of 26 kg/m2, and a mean duration of diabetes of 20 years. All type 1 diabetic patients used insulin and averaged 3.4 injections a day; their average HbAlc level was 7.1 +/- 0.2%, and 80% had HbAlc levels < or = 8.0%. CONCLUSIONS: Although endocrinologists must manage patients with multifaceted problems, complex treatment regimens yield glycemic control levels comparable with the Diabetes Control and Complications Trial and allow ADA guidelines to be met in a routine practice setting. PMID- 10857933 TI - Moderate-intensity physical activity and fasting insulin levels in women: the Cross-Cultural Activity Participation Study. AB - OBJECTIVE: The purpose of this study was to determine the association between moderate-intensity physical activity (PA) and fasting insulin levels among African-American (n = 47), Native American (n = 46), and Caucasian women (n = 49), aged 40-83 years, enrolled in the Cross-Cultural Activity Participation Study. Associations by race/ethnicity, levels of central obesity, and cardiorespiratory fitness were also examined. RESEARCH DESIGN AND METHODS: Physical activity scores were obtained from detailed PA records that included all PA performed during two consecutive 4-day periods scheduled 1 month apart. Using MET intensity (the associated metabolic rate for a specific activity divided by a standard resting metabolic rate), PA was expressed as MET-min (the product of the minutes for each activity times the MET intensity level) per day of energy expended in moderate (3-6 METs) and moderate/vigorous (> or = 3 METs) PA. Fasting insulin levels were determined by radioimmunoassay. Data were analyzed by multiple linear regression analysis. RESULTS: After adjusting for race/ethnicity, age, educational attainment, and site, an increase of 30 min of moderate intensity PA was associated with a 6.6% lower fasting insulin level (P < 0.05). The association was similar among races/ethnicities, centrally lean and centrally obese women, and women with low and high cardiorespiratory fitness levels. CONCLUSIONS: These findings lend support to the 1995 Centers for Disease Control and Prevention and American College of Sports Medicine recommendations for an accumulation of 30 min/day in moderate-intensity PA. They also contribute to the growing literature suggesting that moderate amounts of PA have a significant role in reducing the burden of hyperinsulinemia and diabetes among ethnic populations at highest risk for these conditions. PMID- 10857934 TI - Influence of caffeine on the frequency and perception of hypoglycemia in free living patients with type 1 diabetes. AB - OBJECTIVE: To examine the influence of caffeine on the frequency and perception of hypoglycemia in "free-living" patients with type 1 diabetes. RESEARCH DESIGN AND METHODS: A total of 34 patients with type 1 diabetes were recruited for a prospective randomized placebo-controlled double-blind study. After a lead-in phase and while adhering to a low-caffeine diet, subjects were randomized to capsules containing either 200 mg caffeine or matched placebo with crossover at 3 months. Hypoglycemic episodes were monitored throughout with capillary blood glucose readings and a symptom questionnaire. During the study, measurements of blood pressure, middle cerebral artery blood velocity (a surrogate measure of cerebral blood flow), cognitive function (via a four-choice reaction time test), HbAlc levels, and lipid profiles were taken at the beginning and end of each phase. RESULTS: Throughout the study, no changes were evident regarding glycemic control or lipid profile. The number of symptomatic episodes was greater with caffeine (1.3 vs. 0.9 episodes/week; P < 0.03) and was associated with more intense warning symptoms (29 vs. 26 total symptom score; P < 0.05). For women, caffeine ingestion caused a modest pressor response (115 vs. 110 mmHg; P < 0.01). Four-choice reaction time improved slightly with caffeine supplementation (P < 0.05). CONCLUSIONS: Ingestion of modest amounts of caffeine enhances the intensity of hypoglycemia warning symptoms in patients with type 1 diabetes without altering the prevailing standard of glycemic control or increasing the incidence of severe hypoglycemic episodes. PMID- 10857935 TI - Dietary variables and glucose tolerance in pregnancy. AB - OBJECTIVE: To investigate relationships between dietary macronutrient intakes and glucose tolerance in pregnancy RESEARCH DESIGN AND METHODS: Nulliparous pregnant Chinese women diagnosed with gestational diabetes mellitus (GDM) (n = 56) were compared to age-, gestational age-, height-, and parity-matched groups with normal glucose tolerance (n = 77) and glucose intolerance (IGT) (n = 38) based on the results of an oral glucose tolerance test (National Diabetes Data Group criteria), performed between 24 and 28 weeks of pregnancy. A 24-h recall dietary assessment was also obtained at the time of screening. RESULTS: Subjects with IGT and GDM were significantly heavier (66.1 +/- 1.4 and 68.6 +/- 1.2 kg, respectively, mean +/- SEM) (P < 0.0001) than the normal group (61.2 +/- 1.8 kg) and had a higher BMI. Overall energy intake was similar between groups, as were the intakes of each macronutrient (%kcal). However, there was a highly significant reduction in polyunsaturated fat intake in the IGT and GDM groups whether expressed as %kcal, % of total fat, or fat kcal. This effect was independent of body weight or BMI whether assessed by ordinal logistic regression or by analysis of a weight- and BMI-matched subgroup of the subjects (P = 0.002 for %kcal; n = 47 normal, 26 IGT, and 43 GDM subjects). In logistic regression analysis of the complete data set, increased body weight (P < 0.0001) and decreased polyunsaturated fat intake (P = 0.0014) were both independent predictors of glucose intolerance (IGT and GDM), as were increased body weight and a low dietary polyunsaturated to saturated fat ratio. CONCLUSIONS: Increased polyunsaturated fat intake is associated with a reduced incidence of glucose intolerance during pregnancy. This finding may have major implications for dietary management of women with or at risk of developing GDM. PMID- 10857936 TI - Visceral adiposity and risk of type 2 diabetes: a prospective study among Japanese Americans. AB - OBJECTIVE: We conducted a prospective study among Japanese Americans of diabetes incidence in relation to visceral and regional adiposity, fasting insulin and C peptide, and a measure of insulin secretion, because little prospective data exist on these associations. RESEARCH DESIGN AND METHODS: Baseline variables included plasma glucose, C-peptide, and insulin measured after an overnight fast and 30 and 120 min after a 75-g oral glucose tolerance test; abdominal, thoracic, and thigh fat areas by computed tomography (CT); BMI (kg/m2); and insulin secretion (incremental insulin response [IIR]). RESULTS: Study subjects included 290 second-generation (nisei) and 230 third-generation (sansei) Japanese Americans without diabetes, of whom 65 and 13, respectively, developed diabetes. Among nisei, significant predictors of diabetes risk for a 1 SD increase in continuous variables included intra-abdominal fat area (IAFA) (odds ratio, 95% CI) (1.6, 1.1-2.3), fasting plasma C-peptide (1.4, 1.1-1.8), and the IIR (0.5, 0.3-0.9) after adjusting for age, sex, impaired glucose tolerance, family diabetes history, and CT-measured fat areas other than intra-abdominal. Intra abdominal fat area remained a significant predictor of diabetes incidence even after adjustment for BMI, total body fat area, and subcutaneous fat area, although no measure of regional or total adiposity was related to development of diabetes. Among sansei, all adiposity measures were related to diabetes incidence, but, in adjusted models, only IAFA remained significantly associated with higher risk (2.7, 1.4-5.4, BMI-adjusted). CONCLUSIONS: Greater visceral adiposity precedes the development of type 2 diabetes in Japanese Americans and demonstrates an effect independent of fasting insulin, insulin secretion, glycemia, total and regional adiposity, and family history of diabetes. PMID- 10857937 TI - The association of diabetes specialist care with health care practices and glycemic control in patients with type 1 diabetes: a cross-sectional analysis from the Pittsburgh epidemiology of diabetes complications study. AB - OBJECTIVE: To determine whether diabetes care characteristics and glycemic control differ by use of specialist care in a representative cohort of patients with type 1 diabetes. RESEARCH DESIGN AND METHODS: Health care, sociodemographic characteristics, and glycemic control were compared between participants in the Pittsburgh Epidemiology of Diabetes Complications Study who reported receiving specialist care (n = 212) and those who did not (n = 217). Specialist care was defined as having received care from an endocrinologist or diabetologist or diabetes clinic attendance during the last year. RESULTS: Patients who reported receiving specialist care were more likely to be female, to have an education level beyond high school, to have an annual household income >$20,000, and to have health insurance. Additionally, patients receiving specialist care were more likely to have received diabetes education during the previous 3 years, to have knowledge of HbAlc testing and to have received that test during the previous 6 months, to have knowledge of the Diabetes Control and Complications Trial results, to self-monitor blood glucose, and to inject insulin more than twice daily. A lower HbA1 level was associated with specialist care versus generalist care (9.7 vs. 10.3%; P = 0.0006) as were higher education and income levels. Multivariate analyses suggest that the lower HbA1 levels observed in patients receiving specialist care were restricted to patients with an annual income >$20,000. CONCLUSIONS: Specialist care was associated with higher levels of participation in diabetes self-care practices and a lower HbA1 level. Future efforts should research and address the failure of patients with low incomes to benefit from specialist care. PMID- 10857938 TI - Self-monitoring of blood glucose: language and financial barriers in a managed care population with diabetes. AB - OBJECTIVE: Self-monitoring of blood glucose (SMBG) is a cornerstone of diabetes care, but little is known about barriers to this self-care practice. RESEARCH DESIGN AND METHODS: This cross-sectional study examines SMBG practice patterns and barriers in 44,181 adults with pharmacologically treated diabetes from the Kaiser Permanente Northern California Region who responded to a health survey (83% response rate). The primary outcome is self-reported frequency of SMBG. RESULTS: Although most patients reported some level of SMBG monitoring, 60% of those with type 1 diabetes and 67% of those with type 2 diabetes reported practicing SMBG less frequently than recommended by the American Diabetes Association (three to four times daily for type 1 diabetes, and once daily for type 2 diabetes treated pharmacologically). Significant independent predictors of nonadherent practice of SMBG included longer time since diagnosis, less intensive therapy, male sex, age, belonging to an ethnic minority, having a lower education and neighborhood income, difficulty communicating in English, higher out-of pocket costs for glucometer strips (especially for subjects with lower incomes), smoking, and excessive alcohol consumption. CONCLUSIONS: Considerable gaps persist between actual and recommended SMBG practices in this large managed care organization. A somewhat reduced SMBG frequency in subjects with linguistic barriers, some ethnic minorities, and subjects with lower education levels suggests the potential for targeted, culturally sensitive, multilingual health education. The somewhat lower frequency of SMBG among subjects paying higher out of-pocket expenditures for strips suggests that removal of financial barriers by providing more comprehensive coverage for these costs may enhance adherence to recommendations for SMBG. PMID- 10857939 TI - Prevalence and correlates of preventive care among adults with diabetes in Kansas. AB - OBJECTIVE: To assess the prevalence and correlates of recommended preventive care among adults with diabetes in Kansas. RESEARCH DESIGN AND METHODS: A cross sectional telephone survey was conducted among a sample of adults (> or = 18 years of age) with self-reported diabetes. Recommended preventive care was defined based on four criteria: number of health-care provider (HCP) visits per year (> or = 4 for insulin users and > or = 2 for nonusers), number of foot examinations per year (> or = 4 for insulin users and > or = 2 for nonusers), an annual dilated eye examination, and a blood pressure measurement in the past 6 months. RESULTS: The mean age of the 640 respondents was 61 years, 58% were women, and 86% were white. In the preceding year, 62% of respondents reported the appropriate number of visits to a HCP 27% the appropriate number of foot examinations, 65% an annual dilated eye examination, and 89% a blood pressure measurement in the preceding 6 months. Only 17% (95% CI 14-20) met all four criteria for recommended care. The adjusted odds of receiving recommended care were higher for males than for females (odds ratio [OR] 1.6; 95% CI 1.1-2.5), higher for people whose HCP scheduled follow-up appointments than for those who self-initiated follow-up (OR 2.7; 95% CI 1.6-4.8), and higher for former smokers than for current smokers (OR 3.1; 95% CI 1.6-6.9). CONCLUSIONS: Preventive care for people with diabetes is not being delivered in compliance with current guidelines, especially for women and current smokers. Scheduling follow-up visits for patients, targeting certain high-risk populations, and developing protocols to improve foot care may be effective in improving care. PMID- 10857940 TI - Testosterone, sex hormone-binding globulin, and the development of type 2 diabetes in middle-aged men: prospective results from the Massachusetts male aging study. AB - OBJECTIVE: The objective was to examine prospectively the association between low testosterone and sex hormone-binding globulin (SHBG) levels and the subsequent development of type 2 diabetes in men. RESEARCH DESIGN AND METHODS: Analyses were conducted on the cohort of the Massachusetts Male Aging Study, a population-based random sample of men aged 40-70. Of the 1,709 men enrolled in 1987-1989 (T1), 1,156 were followed up 7-10 years later (T2). Testosterone and SHBG levels at T1 were used to predict new cases of diabetes between T1 and T2. RESULTS: After controlling for potential confounders, diabetes at follow-up was predicted jointly and independently by lower baseline levels of free testosterone and SHBG. The odds ratio for future diabetes was 1.58 for a decrease of 1SD in free testosterone (4 ng/dl) and 1.89 for a 1SD decrease in SHBG (16 nmol/l), both significant at P < 0.02. CONCLUSIONS: Our prospective findings are consistent with previous, mainly cross-sectional reports, suggesting that low levels of testosterone and SHBG play some role in the development of insulin resistance and subsequent type 2 diabetes. PMID- 10857941 TI - Nationwide program for improving the care of diabetic patients in Israeli primary care centers. AB - OBJECTIVE: To improve the effectiveness of primary care providers in Israel to monitor and control glycemic levels of diabetic patients. RESEARCH DESIGN AND METHODS: We designed a 2-year program to improve the effectiveness of primary care providers to administer diabetes care. The program was conducted by the largest Israeli health maintenance organization, which insures 60% of the population. Interventions included continuing medical education and establishing guidelines and diabetes registers in every clinic. A retrospective cohort study was conducted from 1995 to 1997 to evaluate the project's effect on the care of diabetic patients. One patient was randomly chosen for review from each of the physicians' updated diabetes registers. The same indicators and variables were collected for each year. RESULTS: The response rate was 72.7%. Nationwide, 876 physicians participated in the review. From 1995 to 1997, there was a statistically significant improvement in the prevalence of performing all of the parameters for monitoring the primary care of diabetic patients. The process parameters showed a considerable improvement: the prevalence of recording weight increased from 35% of the diabetic patients in 1995 to 60% in 1997: the prevalence of conducting foot inspections increased from 40 to 63%; the prevalence of conducting fundus examinations increased from 38.5 to 68.3%; and the prevalence of measuring HbAlc values increased from 30.6 to 69.9%. As a result, metabolic control significantly improved: the percentage of diabetic patients with HbAlc concentration >9% decreased from 33.2% in 1995 to 22.5% in 1997; the percentage of diabetic patients with HbAlc concentration <7.4% increased from 45.1 to 50.5%. CONCLUSIONS: A major intervention plan based on quality assurance principles can improve physicians' performance on a national scale without the use of punitive administrative measures. PMID- 10857942 TI - Evaluation of an electrochemical sensor for measuring blood ketones. AB - OBJECTIVE: To evaluate the performance of a hand-held ketone sensor that is able to measure blood beta-hydroxybutyrate (beta-HBA) concentrations within 30 s in patients with diabetic ketoacidosis (DKA) and patients who attend a weight management clinic. RESEARCH DESIGN AND METHODS: Two groups of patients were studied: 19 patients admitted with DKA and 156 patients attending a weight management clinic. Paired capillary and venous whole blood samples were measured using the ketone sensor and also using an enzymatic laboratory reference method. RESULTS: The ketone sensor accurately measured beta-HBA concentrations in patients with DKA (limits of agreement -0.9 to + 1.0 mmol/l) or starvation induced ketonemia (limits of agreement -0.5 to +0.5 mmol/l). CONCLUSIONS: This ketone sensor accurately measures whole blood beta-HBA concentrations within 30 s. PMID- 10857943 TI - The efficacy of octreotide in the therapy of severe nonproliferative and early proliferative diabetic retinopathy: a randomized controlled study. AB - OBJECTIVE: The pilot study examined the ability of octreotide to retard progression of diabetic retinopathy (DR) and delay the need for panretinal photocoagulation (PRP) in patients with advanced stages of retinal disease. RESEARCH DESIGN AND METHODS: Patients with severe nonproliferative DR (NPDR) or early non-high-risk proliferative DR (PDR) were randomly assigned to conventional diabetes management (control group, 12 patients) or to treatment with maximally tolerated doses of octreotide (200-5,000 microg/day subcutaneously; 11 patients). Ocular changes in each eye were assessed at a minimum of every 3 months for 15 months or until disease progressed to high-risk PDR requiring laser surgery. Endocrine assessments occurred at 3-month intervals during the study RESULTS: Only 1 of 22 eyes from patients treated with octreotide reached high-risk PDR requiring PRP, compared with control patients, in whom 9 of 24 eyes required PRP. The decreased incidence of progression requiring laser surgery was statistically significant if events were considered independently (P < 0.006). The incidence of ocular disease progression was only 27% in patients treated with octreotide compared with 42% in patients with conventional diabetes management. This treatment effect on whether the retina worsened approached statistical significance using repeated measures analysis (P = 0.0605). Endocrine management was similar between treatment groups. Thyroxine replacement therapy was administered to maintain a euthyroid state for all octreotide-treated patients and 7 of 12 control patients. CONCLUSIONS: Our results suggest that octreotide treatment in euthyroid patients may retard progression of advanced DR and may delay the time to laser surgery. PMID- 10857944 TI - Patterns of quantitative sensation testing of hypoesthesia and hyperalgesia are predictive of diabetic polyneuropathy: a study of three cohorts. Nerve growth factor study group. AB - OBJECTIVE: To test quantitative sensation testing (QST) patterns of hypoesthesia and hyperalgesia as indicators of diabetic polyneuropathy (DPN) and its severity RESEARCH DESIGN AND METHODS: We used Computer-Assisted Sensory Examination IV; characterized the QST results of the foot of each patient in three diabetic cohorts (approximately 1,500 patients) as hyperesthetic (< or = 2.5th percentile), low-normal (2.5th-50th percentiles), high-normal (50th-97.5th percentiles), or hypoesthetic (> or = 97.5th percentile); and tested associations with symptoms, impairments, and test abnormalities. RESULTS: Overall neuropathic impairment was most severe in the pancreas-renal transplant and nerve growth factor cohorts, but it was much less severe in the population-based Rochester Diabetic Neuropathy Study (RDNS) cohort. The frequency distribution of sensory abnormalities mirrored this difference. When the QST spectra of diabetic cohorts were compared with those of the control subject cohort for vibration and cooling sensations, the only abnormality observed was hypoesthesia, which was expressed as an increased number of subjects with values at or above the 97.5th percentile or by an increased percentage of cases with high-normal values. Symptoms and impairments of DPN were significantly more frequent in the subjects with values at or above the 97.5th percentile than in the subjects whose values were between the 50th and 97.5th percentiles. For heat pain (HP) sensation thresholds (intermediate pain severity [HP:5], pain threshold [HP:0.5], and pain-stimulus response slope [HP:5-0.5]), an increased frequency of both hypoalgesia and hyperalgesia was observed (especially in the RDNS cohort). Steeper pain-stimulus response slopes were significantly associated with sensory symptoms, including severity of pain. CONCLUSIONS: 1) Decreased vibratory sensation (hypoesthesia) appears to be characteristic of mild DPN, whereas panmodality hypoesthesia is characteristic of severe DPN. 2) A shift of vibratory and cold detection thresholds (and also of attributes of nerve conduction and a measure of autonomic dysfunction) from low-normal (2.5th-50th percentiles) to high-normal (50th-97.5th percentiles) appears to precede overt expression of DPN and to thereby provide evidence of subclinical abnormality 3) Heat stimulus-induced hyperesthesia (low thresholds) occurs especially in mild DPN, and, because it correlates with DPN symptoms and impairments, it must be attributed to hyperalgesia rather than to supersensitivity Therefore, hypoalgesia or hyperalgesia may be an indicator of early DPN. PMID- 10857945 TI - Increased prandial insulin secretion after administration of a single preprandial oral dose of repaglinide in patients with type 2 diabetes. AB - OBJECTIVE: To examine the dose-related pharmacodynamics and pharmacokinetics of a single preprandial oral dose of repaglinide in patients with type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 16 Caucasian men with type 2 diabetes participated in two placebo-controlled double-blind randomized cross-over studies. Patients were randomized to receive a single oral dose of repaglinide (0.5, 1.0, and 2.0 mg in study 1 and 4.0 mg in study 2) or placebo (both studies) administered 15 min before the first of two sequential identical standard meals (breakfast and lunch) that were 4 h apart. During each of the study days, which were 1 week apart, blood samples were taken at frequent intervals over a period of approximately 8 h for measurement of plasma glucose, insulin, C-peptide, and repaglinide concentrations. RESULTS: During the first meal period (0-240 min), administration of repaglinide reduced significantly the area under the curve (AUC) for glucose concentration and significantly increased the AUC for insulin levels, C-peptide levels, and the insulin secretion rate. These results, compared with those of administering placebo, were dose dependent and log linear. The effect of repaglinide administration on insulin secretion was most pronounced in the early prandial period. Within 30 min, it caused a relative increase in insulin secretion of up to 150%. During the second meal period (240-480 min), there was no difference between repaglinide and placebo administration in the AUC for glucose concentration, C-peptide concentration, and the estimated insulin secretion rate. CONCLUSIONS: A single dose of repaglinide (0.5-4.0 mg) before breakfast improves insulin secretion and reduces prandial hyperglycemia dose dependently Administration of repaglinide had no effect on insulin secretion with the second meal, which was consumed 4 h after breakfast. PMID- 10857946 TI - Cigarette smoking and plasma total homocysteine levels in young adults with type 1 diabetes. AB - OBJECTIVE: The purposes of this study were to compare plasma total homocysteine (tHcy) levels, a recognized cardiovascular risk factor, in nondiabetic subjects and type 1 diabetic patients, and to evaluate whether chronic cigarette smoking had a deleterious effect on plasma tHcy levels in type 1 diabetic patients. RESEARCH DESIGN AND METHODS: Plasma tHcy concentrations were measured in 60 young type 1 diabetic patients without clinical evidence of macroangiopathy and in 30 healthy control subjects who were matched for age, sex, BMI, and smoking habit. RESULTS: Plasma tHcy levels were significantly higher in type 1 diabetic patients than in control subjects (12.5 +/- 4.8 vs. 10.3 +/- 2.2 micromol/l, P = 0.01). After stratification by smoking status, diabetic smokers had values for age, sex, BMI, lipids, creatinine, blood pressure, glycometabolic control, diabetes duration, and microvascular complications that were superimposable on their nonsmoking counterparts. Nevertheless, plasma tHcy levels were markedly elevated in diabetic smokers versus nonsmokers (15.5 +/- 5.7 vs. 10.6 +/- 3 pmol/l, P < 0.0001) in a dose-dependent fashion (P < 0.0001, by analysis of variance when subjects were categorized for the number of cigarettes smoked daily). CONCLUSIONS: Chronic cigarette smoking seems to adversely affect plasma tHcy levels in young adults with type 1 diabetes. PMID- 10857947 TI - Assessing impaired hypoglycemia awareness in type 1 diabetes: agreement of self report but not of field study data with the autonomic symptom threshold during experimental hypoglycemia. AB - OBJECTIVE: The aim of our study was to determine the agreement of two noninvasive methods, a self-report and a field study method, for the assessment of impaired hypoglycemia awareness with a gold standard criterion of hypoglycemia awareness, the autonomic symptom threshold during experimental hypoglycemia. RESEARCH DESIGN AND METHODS: A total of 19 type 1 diabetic patients completed a standardized questionnaire to assess impaired hypoglycemia awareness and performed a hand-held computer (HHC) study to assess their recognition of hypoglycemic episodes occurring during 24 weeks. Patients subsequently underwent a stepped hypoglycemic clamp to study responses to standardized hypoglycemia. Diagnoses of impaired hypoglycemia awareness were based on the separate self-report questions, a composite self-report score, and three different cutoff levels for the percentage of accurately recognized hypoglycemic episodes during the field study Agreement of these noninvasive measures with the hypoglycemic clamp measure were tested by calculating kappa values, sensitivity, and specificity. RESULTS: The composite self-report score agreed reasonably well with the hypoglycemic clamp measure (kappa 0.49, sensitivity 66.7%, and specificity 85.7%) and showed a better agreement than the separate self-report questions. The HHC criterion of impaired hypoglycemia awareness did not agree with the hypoglycemic clamp criterion at any of the cutoff levels tested. CONCLUSIONS: The composite self-report tested in this study is a reasonably reliable assessment method for the diagnosis of impaired hypoglycemia awareness, using the physiological definition of an absence of autonomic symptoms at a blood glucose level of 3 mmol/l. In contrast, the recognition of hypoglycemic events in everyday life as measured using the HHC method is not related to the hypoglycemic clamp criterion. PMID- 10857948 TI - Ambulatory blood pressure, microalbuminuria, and autonomic neuropathy in adolescents with type 1 diabetes. AB - OBJECTIVE: To examine the relationship between 24-h blood pressure (BP) measurements, urinary albumin excretion rates, and autonomic neuropathy (AN) in adolescents with type 1 diabetes. RESEARCH DESIGN AND METHODS: A total of 31 patients with microalbuminuria (MA), 20 patients with intermittent MA (I-MA) and 11 patients with persistent MA (P-MA) were identified from the diabetes clinics at two major Australian tertiary care pediatric hospitals. Two control groups were used; one consisted of 19 age-, sex-, and diabetes duration-matched adolescents with normoalbuminuria (NA), and the other consisted of 46 age- and sex-matched nondiabetic control subjects. A medical history and physical examination were followed by a series of noninvasive tests of cardiovascular and pupillary autonomic function and then by 24-h ambulatory blood pressure monitoring (ABPM). RESULTS: ABPM showed an incremental increase in all BP parameters from nondiabetic control subjects through subjects with NA. A parallel incremental increase in diurnal and nocturnal ambulatory heart rates was also evident. Subjects with MA had significantly reduced pupillary adaptation to darkness compared with nondiabetic subjects and subjects with NA. The above results paralleled an incremental increase in HbAlc levels in adolescents with type 1 diabetes from subjects with NA to subjects with P-MA. CONCLUSIONS: Higher 24-h BP values and evidence of subclinical signs of AN are present before P-MA develops and may have important implications for timing the introduction of treatments designed to prevent or retard the microvascular complications of type 1 diabetes in adolescents. PMID- 10857949 TI - Prevalence of renal artery stenosis in subjects with type 2 diabetes and coexistent hypertension. AB - OBJECTIVE: To assess the prevalence of renal artery stenosis (RAS) in subjects with type 2 diabetes and coexistent hypertension by using magnetic resonance angiography (MRA) of the renal arteries, to assess clinical and biochemical predictors of RAS, and to assess the hemodynamic significance of RAS, by using the captopril test (a measure of the response of plasma renin activity to a single oral dose of captopril). RESEARCH DESIGN AND METHODS: A total of 117 subjects with type 2 diabetes and coexistent hypertension between 40 and 70 years of age and with creatinine concentrations < 150 micromol/l were recruited from two inner-city general diabetes clinics. All subjects underwent MRA of the renal arteries. In a subgroup of 85 subjects, data concerning possible clinical and biochemical predictors of RAS were collected, and the captopril test was performed. For comparison of a continuous variable between subjects with a positive MRA and those with a negative MRA, the Mann-Whitney test was used. For comparison of a discrete variable between subjects with a positive MRA and those with a negative MRA, Fisher's exact test was used. RESULTS: The prevalence of RAS detected by using MRA in 117 hypertensive type 2 diabetic subjects was 17%; 19 subjects had unilateral RAS, and only 1 subject had bilateral RAS. A femoral bruit was significantly more common in subjects with a positive MRA versus subjects with a negative MRA (21 vs. 0%; Fisher's exact test P < 0.005); however, other clinical features of atherosclerotic disease were not statistically associated. Greater duration of hypertension and treatment with statins were features of subjects with RAS (P < 0.05). The captopril test was negative in all subjects, although the antihypertensive response to oral captopril was significantly greater in subjects with RAS detected by MRA. CONCLUSIONS: RAS is common in hypertensive type 2 diabetic subjects. The presence of a femoral bruit is a useful predictive clinical marker. The captopril test is not useful in predicting the hemodynamic significance of RAS in this patient group. PMID- 10857950 TI - Influence of insertion/deletion polymorphism in the ACE-I gene on the progression of diabetic glomerulopathy in type 1 diabetic patients with microalbuminuria. AB - OBJECTIVE: To investigate the influence of the insertion/deletion polymorphism of the ACE gene on the progression of early diabetic glomerulopathy in patients with and without antihypertensive treatment (AHT). RESEARCH DESIGN AND METHODS: There were 30 microalbuminuric patients with >5 years of type 1 diabetes who had renal biopsies taken at baseline and after 26-48 months of follow-up. Of the 30 patients, 13 (4 with II genotype and 9 with ID and DD genotypes) were randomized to AHT (enalapril or metoprolol) during the study. The ACE genotype was determined by a polymerase chain reaction. Glomerular structural changes were measured by stereological methods. RESULTS: Of the patients, 8 had the II genotype, 19 had ID genotype, and 3 had DD genotype. During the study, basement membrane thickness, matrix star volume, and the overall diabetic glomerulopathy index were increased in patients with ID and DD genotypes only (P < 0.001, P = 0.01, P < 0.001, respectively). Among those with ID and DD genotypes, progression of basement membrane thickening and diabetic glomerulopathy index were increased in those without AHT, as compared with the antihypertensive treated patients (P < 0.001, P = 0.02, respectively). In multivariate analysis, the ACE genotype had an independent influence on the progression of basement membrane thickening (P = 0.01), when AHT (P < 0.001) and the mean HbAlc during the study (P < 0.001) were also taken into account. ACE genotype tended to be independently associated with the diabetic glomerulopathy index (P = 0.05). CONCLUSIONS: Microalbuminuric type 1 diabetic patients carrying the D-allele have an increased progression of diabetic glomerulopathy. Presence of this allele and no AHT seems to enhance this process. Larger studies are needed to confirm the clinical significance of our findings. PMID- 10857951 TI - American Diabetes Association annual meeting, 1999: nephropathy and neuropathy. PMID- 10857952 TI - A comparison in a clinical setting of the efficacy and side effects of three thiazolidinediones. PMID- 10857953 TI - Nocturnal glucose control and free insulin levels in children with type 1 diabetes by use of the long-acting insulin HOE 901 as part of a three-injection regimen. PMID- 10857954 TI - Increase in serum ceruloplasmin levels is correlated with a decrease of serum nitric oxide levels in type 2 diabetes. PMID- 10857955 TI - CCTTT-repeat polymorphism in the human NOS2-promoter confers low risk of diabetic nephropathy in type 1 diabetic patients. PMID- 10857956 TI - Capillary blood volume and pain intensity depend on lancet penetration. PMID- 10857957 TI - Screening for silent myocardial ischemia in diabetic patients. PMID- 10857958 TI - Link between diabetes and osteoporosis. PMID- 10857959 TI - Effects of oral hypoglycemic agents and diet on protein metabolism in type 2 diabetes. AB - OBJECTIVE: We tested whether oral hypoglycemic agents (OHA), gliclazide with or without metformin, during an isoenergetic (ISO) and then a low-energy diet (LED) improve the altered kinetics of whole-body protein metabolism in type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 13 type 2 diabetic patients (aged 51+/-2 years, weight 110+/-5 kg, BMI 41+/-1 kg/m2, fasting glucose [FSG] 11.5+/-0.9 mmol/l) (means+/-SEM) and 10 obese control subjects (48+/-3 years, 98+/-6 kg, 37+/-2 kg/m2, FSG 5.5+/-0.3 mmol/l) consumed an ISO, 1.5 g x kg(-1) x day(-1) protein for a body weight corresponding to a BMI of 25 (BMI25), a formula diet (7 days for obese control subjects, 15 days for diabetic patients), and then a 28 day LED with 50% of the energy of ISO but the same protein intake (101+/-2 g/day). OHAs were given during ISO (days 8-15) and LED. On days 6-8 (and 12-14 for diabetic subjects) of ISO and 26-28 of LED, the 60-h oral 15N-glycine method was used to obtain nitrogen flux (Q), synthesis (S), and breakdown (B). Muscle protein catabolism was estimated from N(tau)-methylhistidine (3MH) excretion. RESULTS: During ISO with hyperglycemia, Q, and B adjusted for fat-free mass, sex, and age were higher and nitrogen balance and net endogenous protein synthesis (S B) lower than in control subjects (P<0.05). OHA decreased FSG (9+/-1 mmol/l) and 3MH and increased plasma insulin-to-glucose ratio, nitrogen retention, and S-B to levels in control subjects. The change in S-B correlated with that in FSG (r = 0.845, P = 0.001) and in fasting plasma C-peptide (r = 0.852, P = 0.0005). With LED and OHA, weight decreased 6.3 kg, glycemia reached near-normal levels, and nitrogen equilibrium was maintained; Q decreased by 7%, S and B by 11% (P<0.05) to values found in control subjects. CONCLUSIONS: OHA during ISO corrected protein turnover in relation to glycemia and plasma C-peptide. The LED maintained protein homeostasis in obese control subjects and, in diabetes patients with OHA, normalized protein metabolism. These findings have implications for diet and OHA prescription. PMID- 10857960 TI - Beneficial effects of viscous dietary fiber from Konjac-mannan in subjects with the insulin resistance syndrome: results of a controlled metabolic trial. AB - OBJECTIVE: Dietary fiber has recently received recognition for reducing the risk of developing diabetes and heart disease. The implication is that it may have therapeutic benefit in prediabetic metabolic conditions. To test this hypothesis, we investigated the effect of supplementing a high-carbohydrate diet with fiber from Konjac-mannan (KJM) on metabolic control in subjects with the insulin resistance syndrome. RESEARCH DESIGN AND METHODS: We screened 278 free-living subjects between the ages of 45 and 65 years from the Canadian-Maltese Diabetes Study. A total of 11 (age 55+/-4 years, BMI 28+/-1.5 kg/m2) were recruited who satisfied the inclusion criteria: impaired glucose tolerance, reduced HDL cholesterol, elevated serum triglycerides, and moderate hypertension. After an 8 week baseline, they were randomly assigned to take either KJM fiber-enriched test biscuits (0.5 g of glucomannan per 100 kcal of dietary intake or 8-13 g/day) or wheat bran fiber (WB) control biscuits for two 3-week treatment periods separated by a 2-week washout. The diets were isoenergetic, metabolically controlled, and conformed to National Cholesterol Education Program Step 2 guidelines. Serum lipids, glycemic control, and blood pressure were the outcome measures. RESULTS: Decreases in serum cholesterol (total, 12.4+/-3.1%, P<0.004; LDL, 22+/-3.9%, P<0.002; total/HDL ratio, 15.2+/-3.4%, P<0.003; and LDL/HDL ratio, 22.2+/-4.1%, P< 0.002), apolipoprotein (apo) B (15.1+/-4.3%, P<0.0004), apo B/A-1 ratio (13.1+/-3.4%, P< 0.0003), and serum fructosamine (5.2+/-1.4%, P<0.002) were observed during KJM treatment compared with WB-control. Fasting blood glucose, insulin, triglycerides, HDL cholesterol, and body weight remained unchanged. CONCLUSIONS: A diet rich in high-viscosity KJM improves glycemic control and lipid profile, suggesting a therapeutic potential in the treatment of the insulin resistance syndrome. PMID- 10857961 TI - Gestational diabetes: is a higher cesarean section rate inevitable? AB - OBJECTIVE: To determine the rate of and indication for cesarean section for women with gestational diabetes mellitus (GDM) compared with glucose-tolerant women. RESEARCH DESIGN AND METHODS: From a consecutive series of women with GDM seen over a 9-year period for medical management, women who had had a cesarean section were identified and the reason for the section determined from a review of the medical record. A control group of women who had had a section were obtained from an existing database of glucose-tolerant women. RESULTS: The section rate for women with GDM was higher at 19.8% than the 15.6% for glucose-tolerant women. However, after adjustment for age and parity, no significant differences were found. There were also no differences found for the primary indication for section. CONCLUSIONS: In our health area of New South Wales, Australia, women with GDM do not have a higher section rate compared with glucose-tolerant women. Concerns about the diagnosis of GDM leading to an increased rate of obstetric intervention should not be generalized. PMID- 10857962 TI - Alcohol intake and incidence of type 2 diabetes in men. AB - OBJECTIVE: To evaluate the relation between alcohol intake and incidence of type 2 diabetes. RESEARCH DESIGN AND METHODS: This prospective study included 8,663 men with fasting plasma glucose measurements from at least two medical examinations. Alcohol intake was classified into five groups: nondrinkers and four quartiles (Qs) of drinkers according to the amount of alcohol intake. Type 2 diabetes was diagnosed by 1997 American Diabetes Association criteria. RESULTS: There were 149 incident cases of type 2 diabetes during 52,588 person-years of follow-up. There was a U-shaped association between alcohol intake and diabetes, with the lowest incidence of diabetes at Q2 (61.9-122.7 g/week). As compared with Q2, men in Q3 and Q4 had a 2.2- (95% CI 1.2-3.9, P = 0.01) and 2.4-fold (1.4-4.4, P<0.01) risk of developing diabetes, while nondrinkers and men in Q1 had 1.8- (1.0-3.3, P<0.05) and 1.4-fold (0.7-2.6, P = 0.34) higher risk of diabetes, respectively. These associations persisted after adjustment for age, fasting plasma glucose, smoking, BMI, blood pressure, serum triglyceride concentration, cardiorespiratory fitness, HDL cholesterol, waist circumference, and parental diabetes. CONCLUSIONS: We observed an elevated risk of developing type 2 diabetes in nondrinkers and men with high alcohol intakes, when compared with men who reported moderate alcohol intake. Men with a high alcohol intake may be able to reduce their risk of developing type 2 diabetes if they drink less. PMID- 10857963 TI - Depressive symptoms and metabolic control in African-Americans with type 2 diabetes. AB - OBJECTIVE: To determine the prevalence of depressive symptoms and the relationship between depressive symptoms and metabolic control. RESEARCH DESIGN AND METHODS: We conducted a cross-sectional study of 183 African-American adults aged 35-75 years with type 2 diabetes who were recruited from two primary care clinics in East Baltimore, Maryland. Depressive symptoms, using the Center for Epidemiological Studies Depression Scale (CES-D), HbA1c, fasting lipid profile, BMI, and blood pressure, were measured on each participant. Diabetes-related health behaviors were assessed by questionnaire. RESULTS: The prevalence of depressive symptoms (CES-D > or =22) was 30%. After adjustment for age, sex, income, social support, and duration of diabetes in linear regression models, there were significant graded relationships between greater depressive symptoms and higher serum levels of cholesterol and triglycerides (P<0.050). Similar, albeit less statistically significant, relationships were found with higher levels of HbA1c (P = 0.104), diastolic blood pressure (P = 0.073), and LDL cholesterol (P = 0.176). Unexpectedly, individuals who reported more depressive symptoms also had higher serum levels of HDL cholesterol (P = 0.047). The associations were not explained by differences in diabetes-related health behaviors. CONCLUSIONS: Depressive symptoms are marginally associated with suboptimal levels of HbA1c, diastolic blood pressure, and LDL cholesterol, and significantly associated with suboptimal levels of total cholesterol and triglyceride levels. Prospective studies are required to determine whether improved identification and management of depressive symptoms would enhance metabolic control in this population. PMID- 10857964 TI - Cardiovascular morbidity and early mortality cluster in parents of type 1 diabetic patients with diabetic nephropathy. AB - OBJECTIVE: A familial predisposition was proposed to be a determinant of the increased morbidity and mortality from cardiovascular disease in type 1 diabetic patients with diabetic nephropathy. The insertion allele of an insertion/deletion polymorphism in the ACE (ACE/ID) gene seems to protect against coronary heart disease in nondiabetic and diabetic subjects. The aim of the present study was to evaluate these hypotheses in parents of a large group of type 1 diabetic patients with and without diabetic nephropathy. RESEARCH DESIGN AND METHODS: We investigated cardiovascular morbidity and mortality of parents of 163 type 1 diabetic patients with nephropathy and parents of 163 sex- and age-matched normoalbuminuric patients with type 1 diabetes. RESULTS: Kaplan-Meier curves showed that total parental mortality was significantly increased in parents of type 1 diabetic patients with nephropathy (121 of 244 [ approximately 50%] ) as compared with parents of normoalbuminuric type 1 diabetic patients (119 of 269 [approximately 44%]) (P = 0.008 [log-rank test]) partially due to an increase in cardiovascular deaths (48 of 244 [approximately 20%] vs. 42 of 269 [approximately 16%], P<0.05). In addition, more patients with nephropathy, as compared with the normoalbuminuric group, had at least one parent with fatal/nonfatal cardiovascular disease (46% [95% CI 38-54] vs. 36% [28-44], P = 0.05). Fathers of patients homozygous for the I-allele of the ACE/ID polymorphism had significantly less myocardial infarction as compared with other genotypes (P = 0.03), regardless of the nephropathic state of the offspring. CONCLUSIONS: Cardiovascular morbidity and early mortality clusters in parents of type 1 diabetic patients with diabetic nephropathy The ACE/ID polymorphism helps explain the increased morbidity from cardiovascular disease. PMID- 10857965 TI - Impaired fasting glucose: how low should it go? AB - OBJECTIVE: Impaired fasting glucose (IFG) has been recently introduced as a stage of abnormal carbohydrate metabolism, but the evidence on which its glucose limits (fasting plasma glucose [FPG] 6.1-6.9 mmol/l) are based is not strong. The aim of this study was to determine if 6.1 mmol/l represents a clear cutoff in terms of the risk of future diabetes and in terms of elevated cardiovascular risk factor levels, and to examine the use of other lower limits of IFG. RESEARCH DESIGN AND METHODS: A population-based survey of the island of Mauritius was undertaken in 1987, with a follow-up survey 5 years later. On both occasions, an oral glucose tolerance test was performed and cardiovascular risk factors were measured. RESULTS: Data were available from 4,721 nondiabetic people at baseline, and from 3,542 at follow-up. At baseline, blood pressure, lipids, and obesity increased in a linear fashion with increasing FPG, with no evidence of a threshold effect. The risk of developing hypertension at follow-up was greater for those people with baseline FPG > or =6.1 mmol/l (P<0.001). The risk of developing diabetes at follow-up increased with increasing baseline FPG, but there was little evidence of a threshold near 6.1 mmol/l. CONCLUSIONS: Cardiovascular risk and risk of future diabetes increase continually with increasing FPG, and there is no threshold value on which to base a definition of IFG. If a lower limit of approximately 5.8 mmol/l is used, the category defines a group more similar to the group with impaired glucose tolerance, with regard to total prevalence and the risk of subsequent diabetes. PMID- 10857966 TI - Similar 9-year mortality risks and reproducibility for the World Health Organization and American Diabetes Association glucose tolerance categories: the Hoorn Study. AB - OBJECTIVE: To compare the risks of all-cause and cardiovascular disease (CVD) mortality in the American Diabetes Association (ADA) and World Health Organization (WHO) glucose tolerance categories after 9 years of follow-up in the Hoorn Study and to study the test-retest reproducibility of those categories. RESEARCH DESIGN AND METHODS: In this population-based cohort study of 2,468 elderly men and women, subjects were classified according to both the WHO and the ADA criteria. Causes of death were extracted from the medical records. Age- and sex-adjusted relative risks were estimated by Cox's proportional hazards model. Reproducibility of the diagnostic criteria was assessed in a sample of 1,109 subjects with duplicate oral glucose tolerance tests. RESULTS: Subjects with known diabetes had a four to five times higher risk of all-cause and CVD mortality compared with normal subjects (P<0.05). The relative risks of all-cause mortality were 1.67 (95% CI 1.09-2.57) and 1.56 (1.00-2.43) for newly diagnosed diabetic subjects according to the WHO and ADA criteria, respectively. The WHO and ADA criteria had similar levels of reproducibility The overall K was 0.59 (0.54-0.64) for WHO criteria and 0.61 (0.56-0.66) for ADA criteria. For the category of newly diagnosed diabetes according to WHO or ADA, the percentages of agreement for the second test compared with the first test were 77% (85/110) and 74% (74/100), respectively. CONCLUSIONS: Both sets of diagnostic criteria identify criteria-specific diabetic subjects with an increased mortality risk compared with normal subjects, and the reproducibility of both criteria is similar. PMID- 10857967 TI - Fasting plasma glucose variability predicts 10-year survival of type 2 diabetic patients: the Verona Diabetes Study. AB - OBJECTIVE: In the present study, we evaluated whether the coefficient of variation (CV) of fasting plasma glucose (FPG) over a 3-year period was a significant predictor of mortality in type 2 diabetic patients aged 56-74 years. RESEARCH DESIGN AND METHODS: All type 2 diabetic patients (n = 1,409) aged 56-74 years attending the Verona Diabetes Clinic and having at least two FPG determinations in each of the years 1984-1986 were followed for 10 years (1987 1996) to assess total and cause-specific mortality Patients were grouped into tertiles of mean and CV of FPG during 1984-1986. These parameters as well as sex, age, diabetes duration, insulin treatment, smoking, hypertension, and hypercholesterolemia were included in multivariate survival analyses. RESULTS: During the follow-up, 468 patients died. The CV of FPG was an independent predictor of total, cardiovascular, and cancer mortality. Mean FPG was a predictor of total mortality only when the CV of FPG was not included in the analyses. CONCLUSIONS: Long-term variability of fasting glucose is an independent predictor of mortality in patients with type 2 diabetes. The CV of FPG might be considered a useful additional parameter in the management of these patients. PMID- 10857968 TI - Trends in persistent proteinuria in adult-onset diabetes: a population-based study. AB - OBJECTIVE: This study investigates temporal trends in the prevalence and incidence of persistent proteinuria among people with adult-onset diabetes (age > or =40 years). RESEARCH DESIGN AND METHODS: The complete community-based medical records of all Rochester, Minnesota, residents with a diagnosis of diabetes or diabetes-like condition from 1945 through 1989 were reviewed to determine whether they met National Diabetes Data Group (NDDG) criteria. All confirmed diabetes cases residing in Rochester on 1 January 1970 (n = 446), 1980 (n = 647), and/or 1990 (n = 940) were identified. The medical records of these prevalence cases were reviewed from the time of the first laboratory urinalysis value to the last visit, death, or 1 April 1992 (whichever came first) for evidence of persistent proteinuria (two consecutive urinalyses positive for protein, with no subsequent negative values). Similarly, the medical records of all 1970-1989 diabetes incidence cases (n = 1,252) were reviewed to investigate temporal changes in 1) the likelihood of having persistent proteinuria before the date NDDG criteria was met, i.e., baseline; 2) the risk of persistent proteinuria after baseline; and 3) the relative risk of mortality associated with persistent proteinuria. RESULTS: The proportion of diabetes prevalence cases with persistent proteinuria on or before the prevalence date declined from 20% in 1970 to 11% in 1980 and 8% in 1990. Among the 1970-1989 diabetes incidence cases, 77 (6%) had persistent proteinuria on or before baseline; the adjusted odds declined by 50% with each 10 year increase in baseline calendar year (P<0.001). Among individuals free of persistent proteinuria at baseline, 136 subsequently developed persistent proteinuria; the estimated 20-year cumulative incidence was 41% (95% CI 31-59); the adjusted risk did not differ as a function of baseline calendar year. Survival of individuals with persistent proteinuria relative to those without was reduced but did not differ by baseline calendar year. CONCLUSIONS: The prevalence of persistent proteinuria among people with adult-onset diabetes in Rochester, Minnesota, declined 60% between 1970 and 1990. The decline appears because of a decrease in the proportion of diabetes incidence cases with persistent proteinuria before baseline rather than secular declines in the risk of persistent proteinuria after baseline or secular increases in the risk of mortality associated with persistent proteinuria. Similarity over time in age and fasting glucose at baseline, and at prevalence dates, is evidence that earlier detection of diabetes is not the sole explanation for the decline. PMID- 10857969 TI - Homeostasis model assessment closely mirrors the glucose clamp technique in the assessment of insulin sensitivity: studies in subjects with various degrees of glucose tolerance and insulin sensitivity. AB - OBJECTIVE: To evaluate whether the homeostasis model assessment (HOMA) is a reliable surrogate measure of in vivo insulin sensitivity in humans. RESEARCH DESIGN AND METHODS: In the present study, we compared insulin sensitivity as assessed by a 4-h euglycemic (approximately 5 mmol/l) hyperinsulinemic (approximately 300 pmol/l) clamp with HOMA in 115 subjects with various degrees of glucose tolerance and insulin sensitivity. RESULTS: We found a strong correlation between clamp-measured total glucose disposal and HOMA-estimated insulin sensitivity (r = -0.820, P<0.0001), with no substantial differences between men (r = -0.800) and women (r = -0.796), younger (aged <50 years, r = 0.832) and older (r = -0.800) subjects, nonobese (BMI <27 kg/m2, r = -0.800) and obese (r = -0.765) subjects, nondiabetic (r = -0.754) and diabetic (r = -0.695) subjects, and normotensive ( r = -0.786) and hypertensive (r = -0.762) subjects. Also, we found good agreement between the two methods in the categorization of subjects according to insulin sensitivity (weighted k = 0.63). CONCLUSIONS: We conclude that the HOMA can be reliably used in large-scale or epidemiological studies in which only a fasting blood sample is available to assess insulin sensitivity PMID- 10857970 TI - Metabolic effects of troglitazone therapy in type 2 diabetic, obese, and lean normal subjects. AB - OBJECTIVE: To characterize metabolic effects of troglitazone in type 2 diabetic, obese, and lean subjects, and examine the effects of troglitazone 2-3 weeks after discontinuation. RESEARCH DESIGN AND METHODS: Nine type 2 diabetic, nine obese, and nine lean subjects underwent baseline metabolic studies including an 8-h meal tolerance test (MTT) and a 5-h glucose clamp. Subjects then received troglitazone (600 mg/day) for 12 weeks and subsequently had repeat metabolic studies. Diabetic subjects remained off hypoglycemic agents for 2-3 weeks and then underwent a 5-h glucose clamp. RESULTS: In diabetic subjects, fasting plasma glucose was reduced (P<0.05) and insulin-stimulated glucose disposal (Rd) was enhanced by treatment (P<0.02). The area under the MTT 8-h plasma glucose curve declined with therapy (P<0.001), and its change was positively correlated with the improvement in Rd (r = 0.75, P<0.05). There was also a positive correlation between the change in fasting hepatic glucose output (HGO) and the change in fasting plasma glucose with treatment (r = 0.92, P<0.001). Discontinuation of therapy for 2-3 weeks did not significantly affect fasting plasma glucose or insulin-stimulated glucose Rd. In obese subjects, insulin-stimulated glucose Rd improved with therapy (P<0.001), allowing for maintenance of euglycemia by lower plasma insulin concentrations (P<0.05). In lean subjects, an increase in fasting HGO (P<0.001) and glucose clearance (P<0.01) was observed. CONCLUSIONS: Troglitazone lowers fasting and postprandial plasma glucose in type 2 diabetes by affecting both fasting HGO and peripheral insulin sensitivity. Its effects are evident 2-3 weeks after discontinuation. In obese subjects, its insulin sensitizing effects suggest a role for its use in the primary prevention of type 2 diabetes. PMID- 10857971 TI - Sulfonylurea receptor 1 gene variants are associated with gestational diabetes and type 2 diabetes but not with altered secretion of insulin. AB - OBJECTIVE: To investigate the possible association of the variants in the nucleotide binding fold regions of the sulfonylurea receptor 1 (SUR1) gene with gestational diabetes mellitus (GDM), type 2 diabetes, and altered insulin secretion in Finnish subjects. RESEARCH DESIGN AND METHODS: The nucleotide binding fold regions of the SUR1 gene were amplified with polymerase chain reaction and screened by the single-strand conformational polymorphism analysis in 42 subjects with GDM and 40 subjects with type 2 diabetes. Detected variants were further investigated in 377 normoglycemic subjects by restriction fragment length polymorphism analysis. The effect of the variants of the SUR1 gene on first-phase insulin secretion was studied in 295 normoglycemic subjects. RESULTS: In subjects with GDM or type 2 diabetes, one amino acid change (S1369A), four silent substitutions (R1273R, L829L, T759T, and K649K), and three intron variants were identified in the nucleotide binding fold regions of the SUR1 gene. A tagGCC allele of exon 16 splice acceptor site was more frequent in subjects with GDM (0.55 allele frequency, n = 42) and type 2 diabetes (0.60, n = 40) than in normoglycemic subjects (0.43, n = 377) (P1 = 0.024 and P2 = 0.009, respectively). Similarly, an AGG allele of the R1273R polymorphism was more common in subjects with GDM (0.87) and type 2 diabetes (0.87) than in normoglycemic subjects (0.74) (P1 = 0.009 and P2 = 0.001, respectively). However, the S1369A, R1273R, and cagGCC-->tagGCC variants of the SUR1 gene were not associated with altered first phase insulin secretion in 295 normoglycemic subjects. CONCLUSIONS: These results suggest that a functional variant that contributes to the risk of GDM and type 2 diabetes may locate close to the SUR1 gene. PMID- 10857972 TI - Association of hyperandrogenemia and hyperestrogenemia with type 2 diabetes in Hispanic postmenopausal women. AB - OBJECTIVE: Accumulating evidence suggests that hyperandrogenemia may be a risk factor for coronary heart disease (CHD) in women. The present study was carried out to test the hypothesis that hyperandrogenemia is associated with type 2 diabetes in women and thus may contribute to the increased risk of CHD in women with type 2 diabetes. RESEARCH DESIGN AND METHODS: Sex hormones, sex hormone binding globulin (SHBG), and risk factors for CHD were measured in 20 postmenopausal women with type 2 diabetes and in 29 control subjects. All of the diabetic and control subjects were Hispanic women aged >55 years who were not taking hormone replacement therapy lipid-lowering drugs, or insulin and who were otherwise randomly chosen from a cohort of stroke-free subjects from the Northern Manhattan Stroke Study RESULTS: Mean age, BMI, total cholesterol, LDL cholesterol, HDL cholesterol, triglycerides, blood pressure, and smoking were not significantly different between cases and control subjects, but waist-to-hip ratio (WHR) was significantly higher in the diabetic subjects (P = 0.01). The mean levels of free testosterone (FT) (P = 0.01), dehydroepiandrosterone sulfate (P<0.04), and estradiol (P = 0.01) (controlled for WHR) were significantly higher in the diabetic subjects; with the statistical outliers removed, the testosterone (P = 0.05) and androstenedione (P = 0.002) levels (controlled for WHR) were also significantly higher in the diabetic subjects. The mean levels of estrone, cortisol, and SHBG were not significantly different. The results were similar in the 10 diabetic subjects treated with diet only Significant positive correlations (controlled for age and BMI) were observed between FT or testosterone and cholesterol, LDL cholesterol, and blood pressure. CONCLUSIONS: Postmenopausal Hispanic women with type 2 diabetes had both hyperandrogenemia and hyperestrogenemia, and testosterone or FT correlated positively with risk factors for CHD. Hyperandrogenemia may be a link between diabetes and CHD in women. PMID- 10857973 TI - Vitreous levels of IGF-I, IGF binding protein 1, and IGF binding protein 3 in proliferative diabetic retinopathy: a case-control study. AB - OBJECTIVE: To evaluate vitreous levels of IGF-I and its binding proteins IGFBP-1 and IGFBP-3 in patients with proliferative diabetic retinopathy (PDR). Because intravitreal proteins are elevated in patients with PDR due to the disruption of the blood-retinal barrier, we have corrected vitreal IGF-I and IGFBPs by total vitreal proteins to avoid this confounding factor. RESEARCH DESIGN AND METHODS: We compared 21 diabetic patients with proliferative retinopathy (group A) and 13 nondiabetic patients (group B) in whom a vitrectomy was performed. Both groups were matched by age, serum IGF-I, IGFBP-1, and IGFBP-3 levels. Serum and vitreous levels of IGF-I, IGFBP-1, and IGFBP-3 were measured by immunological methods. Vitreal proteins were assessed by turbidimetric method. RESULTS: Vitreal levels of IGF-I were elevated in group A (median 1.35 ng/ml [range 0.3-8.7]) in comparison with group B (median 0.25 ng/ml [range 0-1.38]), P<0.001. After adjusting by vitreal proteins [ratio IGF-I (ng/ml)/protein (mg/ml)], the differences remain significant (P<0.005). Vitreal levels of IGFBP-1 and IGFBP-3 were also elevated in diabetic patients (IGFBP-1: group A, median 1.6 ng/ml [range 0.6-20.7]; group B, median 0.4 ng/ml [range 0.3-1.9], P<0.001. IGFBP-3: group A, median 102.6 ng/ml [range 53.9-350.8]; group B, median 29.0 ng/ml [range 3.2-87.8], P<0.001). However, when the ratio IGFBP/protein was considered, the differences were not significant. CONCLUSIONS: Intraocular synthesis contributes to elevated vitreous concentrations of IGF-I found in PDR. By contrast, unspecific increase of intravitreal proteins is the main factor explaining the elevated vitreous levels of IGFBP-1 and IGFBP-3 found in diabetic patients. PMID- 10857974 TI - Total homocysteine in patients with type 1 diabetes. AB - OBJECTIVE: Our aim was to study the presence of moderate hyperhomocysteinemia, a risk factor for premature cardiovascular disease, its modifying vitamin factors (folates, vitamins B12 and B6), and lipid risk factors in juvenile type 1 diabetes. RESEARCH DESIGN AND METHODS: A total of 91 patients with type 1 diabetes (46 girls and 45 boys) were studied, with ages ranging from 11 to 18 years, a duration of diabetes from 1 to 15 years, and in pubertal development (stages III, IV, V). In all patients, cholesterol, triglycerides, HDL and LDL cholesterol, lipoprotein(a), folates, cobalamin, vitamin B6, and total homocysteine were determined by specific assays. Microalbuminuria, defined as a ratio of albumin/creatinine >3 mg/mmol creatinine, was analyzed in the first morning specimen. RESULTS: Plasma total homocysteine (tHcy) concentrations were not different in the 91 diabetic children (median [range]) (11-15 years, 6.1 micromol/l [3.2-9.6]; 16-18 years, 7.3 micromol/l [3.9-12]) compared with the control group (11-15 years, 6.6 micromol/l [4.4-10.8]; 16-18 years, 8.1 micromol/l [4.6-11.3]). No significant differences were found in tHcy values in relation to the metabolic control of the disease as assessed by glycohemoglobin values, the duration of disease, alterations in fundus oculi, or presence of lymphocytic thyroiditis. A positive correlation was found between tHcy and plasma creatinine in type 1 diabetic patients that might be related with the increase in muscle mass. There was a negative correlation between tHcy and serum folate (P<0.001) and vitamin B12 (P<0.05), but not with vitamin B6 levels. No significant correlations were found between tHcy and the lipid parameters. CONCLUSIONS: Hyperhomocysteinemia was not detected in adolescents with type 1 diabetes. PMID- 10857975 TI - Increased plasminogen activator inhibitor-1 activity in offspring of type 2 diabetic patients: lack of association with plasma insulin levels. AB - OBJECTIVE: To determine whether a dysregulation of the fibrinolytic system exists in normal glucose tolerant offspring of type 2 diabetic patients. RESEARCH DESIGN AND METHODS: In this cross-sectional study, 32 offspring of type 2 diabetic patients and 26 subjects with no family history of diabetes were studied. With respect to the metabolic parameters, plasma fasting and 2-h postload (75 g glucose) glucose and insulin levels, total cholesterol, triglycerides, and HDL cholesterol concentrations were determined. To evaluate the status of hemostatic factors, fibrinogen, tissue plasminogen activator (tPA) antigen level, plasminogen activator inhibitor-1 (PAI-1) antigen level, and PAI-1 activity were assessed. The statistical analyses included the Mann-Whitney U test to check the significance of differences between variables in the two groups and Spearman's rank correlation tests to check the interrelationships between the hemostatic and metabolic parameters in the offspring group. RESULTS: All subjects had normal glucose tolerance according to the American Diabetes Association criteria. Plasma fasting and postload insulin concentrations were significantly higher in offspring compared with control group (P<0.00001 and P<0.01, respectively). Plasma fasting and postload glucose, fibrinogen, tPA antigen, total cholesterol, and BMI were comparable between the groups. The offspring had significantly higher waist-to-hip ratio (WHR) (P = 0.03), higher triglycerides (P = 0.01), and lower HDL cholesterol (P<0.01) compared with the control group. PAI-1 antigen level and PAI-1 activity were higher in the offspring (P = 0.05 and P = 0.04, respectively). In the offspring group, PAI-1 activity was correlated with plasma PAI-1 antigen level (r = 0.40, P = 0.02), fibrinogen (r = 0.45, P = 0.01), and HDL cholesterol (r = -0.36, P = 0.04). However, tPA antigen level, fasting and postload plasma glucose and insulin, total cholesterol, triglycerides, WHR, and BMI did not correlate with PAI-1 activity. CONCLUSIONS: These data suggest that normal glucose tolerant offspring of type 2 diabetic subjects have elevated PAI-1 activity indicating to hypofibrinolysis in this group. The elevated PAI-1 activity has no association with plasma insulin concentration. PMID- 10857976 TI - Smoking and diabetes: American Diabetes Association. PMID- 10857977 TI - Use of influenza and pneumococcal vaccines in people with diabetes. PMID- 10857978 TI - Immunization and the prevention of influenza and pneumococcal disease in people with diabetes: American Diabetes Association. PMID- 10857979 TI - Pancreas and islet transplantation for patients with diabetes. PMID- 10857980 TI - Pancreas transplantation for patients with type 1 diabetes: American Diabetes Association. PMID- 10857981 TI - American Diabetes Association Annual Meeting, 1999: diabetes and obesity. PMID- 10857982 TI - Diabetes screening practices among individuals aged 45 years and older. PMID- 10857983 TI - HbA1c is not recommended as a screening test for diabetes in cystic fibrosis. PMID- 10857984 TI - Pro115Gln peroxisome proliferator-activated receptor-gamma and obesity. PMID- 10857985 TI - Insulin secretion, insulin sensitivity, and glucose effectiveness in nonobese individuals with varying degrees of glucose tolerance. PMID- 10857986 TI - Late-onset troglitazone-induced hepatic dysfunction. PMID- 10857987 TI - Glyburide-induced hemolysis in myelodysplastic syndrome. PMID- 10857988 TI - Effects of exposure at an altitude of 3,000 m on performance of glucose meters. PMID- 10857989 TI - Deterioration of glycemic control after long-term treatment with troglitazone in nonobese type 2 diabetic patients. PMID- 10857990 TI - UCN-01 enhances the in vitro toxicity of clinical agents in human tumor cell lines. AB - UCN-01 is undergoing Phase I evaluation and is a candidate for combination strategies in the clinic. UCN-01 has been shown to have a variety of effects on cellular targets and the cell cycle. It has also been reported to sensitize cells to several clinical drugs in vitro, possibly in a manner related to p53 status. Thus, combinations of UCN-01 with a series of clinical agents in variety of cell lines have been investigated in vitro. Certain cell lines demonstrated synergistic interactions with combinations of UCN-01 (20-150 nM) and thiotepa, mitomycin C, cisplatin, melphalan, topotecan, gemcitabine, fludarabine or 5 fluorouracil. In contrast, UCN-01 combinations with the antimitotic agents, paclitaxel and vincristine, or topoisomerase II inhibitors, adriamycin and etoposide, did not result in synergy, only in additive toxicity. Cells with non functional p53 were significantly more susceptible to the supra-additive effects of certain DNA-damaging agents and UCN-01 combinations, than cells expressing functional p53 activity. In contrast, there was no significant relationship between p53 status and susceptibility to synergy between antimetabolites and UCN 01. The mechanism behind the observed synergy appeared unrelated to effects on protein kinase C or abrogation of the cell cycle in G2. Moreover, increased apoptosis did not fully explain the supradditive response. These data indicate that UCN-01 sensitizes a variety of cell lines to certain DNA-damaging agents (frequently covalent DNA-binding drugs) and antimetabolites in vitro, but the mechanism underlying this interaction remains undefined. PMID- 10857991 TI - Proteasome inhibition: a new strategy in cancer treatment. AB - The ubiquitin proteasome pathway is a highly conserved intracellular pathway for the degradation of proteins. Many of the short-lived regulatory proteins which govern cell division, growth, activation, signaling and transcription are substrates that are temporally degraded by the proteasome. In recent years, new and selective inhibitors of the proteasome have been employed in cell culture systems to examine the anti-tumor potential of these agents. This review covers the chemistry of selected proteasome inhibitors, possible mechanisms of action in cell culture and the in vivo examination of proteasome inhibitors in murine and human xenograft tumor models in mice. One inhibitor, PS-341, has recently entered Phase I clinical trials in cancer patients with advanced disease to further test the potential of this approach. PMID- 10857992 TI - DNA G-quadruplexes, telomere-specific proteins and telomere-associated enzymes as potential targets for new anticancer drugs. AB - Telomeres and telomerase have been subjects to a tremendous attention from scientists and oncologists during the past 5 years. This interest has been motivated by the potential of telomerase as a tumor marker for the diagnosis and the prognosis of cancer. The possible use of telomerase or telomeres as new targets for anticancer drugs also triggered investigations. The expression of telomerase was found in overall 85% of cancers. Telomerase is early expressed during oncogenesis with a gradient indicating that a high level of telomerase expression could be associated with a bad prognosis. Therefore, drugs targeting telomerase and telomeres might be useful in many human tumors with little restrictions regarding the tumor type or on the stage of the disease. Moreover, since telomerase is not or slightly expressed in normal cells, it has been postulated that drugs targeting telomerase would induce low toxicity. The race for the discovery of telomerase inhibitors has started while the identification of the components controlling telomerase, telomeres, cell survival, senescence, and apoptosis was still in progress. The recent identification of components regulating telomere length and telomerase expression (TRF1, TRF2, and tankyrase) opened a variety of new opportunities to control telomerase/telomere interactions. Meanwhile, a proof of principle was provided that changing telomere interactions with telomere binding proteins by chemical or biological means can induce cancer cell death. Interestingly, recent data challenge the old paradigm which suggested that a long exposure to telomerase and telomere inhibitors is necessary to induce anticancer effects. In this paper, we review the most recent information concerning the regulation of telomere length and telomerase expression, with emphasis on mechanisms that might translate into new drug discovery. PMID- 10857993 TI - Cytotoxic effects of topotecan combined with various active G2/M-phase anticancer drugs in human tumor-derived cell lines. AB - Topotecan (TPT) is a DNA-Topoisomerase I poison that exhibits antitumor activity. TPT, like other DNA-damaging agents, arrests or delays cell cycle progression during S- and G2-phase in a wide variety of tumor-derived cell lines. Particularly, the G2-arrest gives time for the cell to repair its DNA lesions prior to starting a new cell cycle. Based on these observations, we assessed the interaction between TPT and G2/M-active agents in p53-mutated cell lines of diverse origin in order to achieve cell toxicity. Two short-term sequential schedules were administered (TPT --> G2/M-active drug at the interval of greatest TPT-induced G2/M-phase cell arrest, and G2/M-active drug --> TPT), in three human tumor-derived cell lines with proven sensitivity to the following drugs: Bleomycin in HEp-2 (squamous larynx carcinoma); Docetaxel in SKBr-3 (breast adenocarcinoma); Etoposide in NCI-H23 (non-small-cell lung cancer). Our results show that: 1) Sequential TPT --> G2/M-active drugs are synergistic when administration overlapped the maximum percentage of TPT-induced G2/M-phase cell arrest interval in all three mutated p53 cell lines; 2) the reverse sequential schedule (G2/M-active drug --> TPT) was antagonistic, and being only additive for Etoposide --> TPT association. In conclusion, our findings further support the potential cytotoxic role of TPT in combination with other active drugs when the correct schedule of administration is applied. In addition, they provide a rationale for new applications in clinical trials using short-term sequential TPT --> G2/M-active drugs. PMID- 10857994 TI - Cdc25 as a potential target of anticancer agents. AB - Ever since its discovery in yeast more than a decade ago [1], Cdc25 has continued to surprise and intrigue researchers. This dual-specificity protein tyrosine phosphatase (dsPTPase) and other members of the protein tyrosine phosphatase family (PTPases) have only recently joined the protease and kinase enzyme families in drug discovery efforts. The role of phosphatases in tumourigenesis was reviewed recently by Parsons [2]. He is arguing that the phosphatase family of enzymes is involved in a variety of cancers and thus poses both a challenge and an opportunity for new therapeutics. The general biology and biochemistry of Cdc25 were recently reviewed [3]. Here I shall first summarize the recent literature on the role of Cdc25 in disease, as well as on new insights into the regulation of this family of proteins. In the second part, I will review current knowledge of the Cdc25 protein structure and the chemical structures and activities of published Cdc25 inhibitors. PMID- 10857995 TI - Chemotherapy in non-small cell lung cancer. PMID- 10857996 TI - CI-980 in advanced melanoma and hormone refractory prostate cancer. AB - INTRODUCTION: CI-980 is a novel chemotherapeutic agent that inhibits polymerization of tubulin. Preclinical studies have indicated a high level activity of this agent against various tumor cell lines. METHODS: 13 malignant melanoma patients who had failed prior chemotherapy and/or immunotherapy and 13 hormone refractory prostate cancer patients, including 4 who had received prior chemotherapy, were treated in 2 separate NCI-supported clinical trials. Subjects received a recommended phase II dose of CI-980 of 4.5 mg/m2/day by continuous infusion for 72 hours every 3 weeks. RESULTS: No activity was seen in either study. Toxicity was tolerable with neutropenia being the most common, significant toxicity. Among the melanoma patients, 15% and 31% developed grade 3 and grade 4 neutropenia, while 7% and 38% of the prostate patients developed grade 3 and grade 4 neutropenia, respectively. CONCLUSIONS: CI-980 at this dose and schedule is ineffective against malignant melanoma and hormone refractory prostate cancer. PMID- 10857997 TI - A phase II pilot study of KW-2189 in patients with advanced renal cell carcinoma. AB - BACKGROUND: KW-2189 is a semi-synthetic, water-soluble analog of duocarmycin B2, a new class of potent antitumor antibiotics produced by streptomyces, with improved in vitro antitumor potency. PATIENTS AND METHODS: Forty patients with pathologically confirmed metastatic renal cell carcinoma were treated in this multicenter, open-label phase II trial. All patients received 0.4 mg/m2 KW-2189 as an i.v. infusion for Cycle I. Cycles were repeated every 5 to 6 weeks with escalations to 0.5 mg/m2 in the absence of significant toxicity or disease progression. RESULTS: No patient had an objective response. The most common drug related toxicity was hematological-delayed neutropenia and thrombocytopenia, with recovery by week 6. Non-hematologic toxicity consisted of mild to moderate fatigue, nausea and vomiting, and anorexia that was generally manageable. CONCLUSIONS: KW-2189 in this dose and schedule has a predictable safety profile of reversible myelosuppression. No activity in metastatic renal cell carcinoma was demonstrated. PMID- 10857998 TI - A phase II trial of topotecan in esophageal carcinoma: a Southwest Oncology Group study (SWOG 9339). AB - BACKGROUND: Chemotherapeutic treatments containing topoisomerase I inhibitors have shown antitumor activity against a number of solid tumors. Responses have been seen in Phase I trials using topotecan in ovarian, lung, and esophageal cancer. A phase II trial using continuous infusion topotecan was completed to assess activity in esophagus cancer. METHODS: Forty-five eligible patients with locally-advanced or metastatic squamous cell carcinoma or adenocarcinoma of the esophagus received a regimen consisting of 24-hour continuous infusion topotecan at 1.5 mg/m2/day on Days 1, 8, 15, 22 (of 42-day cycle). Patients continued on treatment until evidence of disease progression or unacceptable toxicity. RESULTS: Partial response was demonstrated in 1 patient (2% confirmed response rate). Thirty-six patients progressed during the first cycle of treatment. The median survival was 3 months, and the median progression-free survival was 1 month. Toxicity was mild with only one Grade 4 toxicity reported. CONCLUSIONS: This phase II trial indicates no significant anti-neoplastic activity for topotecan administered in the dose and schedule to patients with squamous cell or adenocarcinoma of the esophagus. PMID- 10857999 TI - Masking of tinnitus through a cochlear implant. AB - The relief of tinnitus has been effected in a cochlear-implant patient by presenting masking sounds to the microphone of the implant. Oddly enough, the presence of the masking sound improved the clarity of speech and music. PMID- 10858000 TI - Evaluation and treatment of severe hyperacusis. AB - A 52-year-old male was evaluated by the authors after initially reporting fullness in his left ear while traveling on an airplane. A unique feature of the patient's complaint was the development of severe bilateral hyperacusis (loudness discomfort levels of between 20-34 dB HL) in spite of the fact that the hearing loss was initially reported in the left ear. To achieve loudness comfort, the patient was initially fit with ER-25 musician earplugs that proved to be unsuccessful. The patient next purchased earplugs and earmuffs from a gun shop in order to obtain relief from the pain and discomfort caused by his exposure to everyday environmental sounds. This paper describes the use of hearing devices that proved to be effective in providing attenuation sufficient that the patient rarely needs to rely on earplugs and earmuffs for relief from his hyperacusis. PMID- 10858001 TI - Catastrophic progressive hearing loss in childhood. AB - We present a report on a 5-year-old child with a complex medical and audiologic history who exhibited catastrophic progression in hearing loss. Hearing loss was initially attributed to bacterial meningitis at age 3 months; progression was apparently related to perilymph fistula at age 8 years. Etiologies associated with progressive hearing loss in children as well as signs of progression and monitoring protocols for children at risk for progressive hearing loss are discussed. PMID- 10858002 TI - Cochlear implantation of auditory neuropathy. AB - Auditory neuropathy (AN) is a hearing disorder that presents with a grossly abnormal or absent neural response as measured by evoked potentials in the presence of normal outer hair cell function evidenced by present otoacoustic emissions or cochlear microphonics. Rehabilitation for patients with AN is challenging due to abnormal temporal encoding at the auditory nerve leading to severely impaired speech perception. Although patients with AN may demonstrate improvement in thresholds with amplification, temporal encoding dysfunction, and consequently speech perception degradation, is not alleviated by amplification. Another issue is the heterogeneity of the AN population in terms of audiologic and neurologic findings, in addition to uncertain etiology and pathophysiology. For children with prelingual onset of AN, development of auditory and oral communication skills is particularly compromised. All children with hearing loss in the severe-to-profound range who do not benefit from conventional amplification can be considered candidates for a cochlear implant (CI). This paper presents a case study of a child with AN who received a CI. Whereas no synchronous neural response auditory brainstem response could be elicited to acoustic stimuli, an electrically evoked auditory nerve action potential was evident following implantation, suggesting restoration to some degree of neural synchrony. Significant improvement in speech perception was found post-CI. Recommendation to implant all patients with AN would be premature, but these findings suggest that electrical stimulation in some cases of auditory neuropathy can be a viable option. PMID- 10858003 TI - Adaptation to loudness and environmental stimuli in three newly fitted hearing aid users. AB - Hearing aid fitting strategies can be categorized according to whether a loudness normalization or a loudness equalization rationale is employed. Regardless of the underlying rationale, the amount of patient participation in determining the initial hearing aid settings will vary when an audiologist-driven (AD) versus a patient-driven protocol is employed. When an AD protocol is used, few changes are made during the initial fitting session based on user feedback. It is assumed that the patient will adapt to the loudness and/or sound quality provided by the hearing aids if not immediately acceptable. The following three case reports document varying degrees of adaptation to hearing aid settings derived using an AD approach. Clinical implications will be discussed. PMID- 10858004 TI - Cisplatin ototoxicity, increased DPOAE amplitudes, and magnesium deficiency. Distortion product otoacoustic emissions. AB - Outer hair cell (OHC) metabolism is blocked by cisplatin. Concurrent changes in the renal handling of magnesium occur because of the damage cisplatin causes to the renal proximal tubule cells within the thick ascending loop of Henle. Although there is no evidence of cisplatin within the OHCs, there are significant levels of intracellular calcium, the antagonist to magnesium at the cell membrane. The OHC motile response is dependent on intracellular calcium. When the calcium current is suppressed by an antagonist, the extracellular OHC microphonic potential decreases. Magnesium deficiency is known to produce hyperexcitability within the central nervous system, including fatal audiogenic seizures. In addition, increases in the amplitude of the auditory brainstem response wave V occur with aminoglycoside therapy and magnesium deficiency. This paper illustrates the amplitude growth of distortion product otoacoustic emissions in two patients treated with cisplatin and explores the possible underlying reasons why this may be related to magnesium metabolism. PMID- 10858005 TI - Just noticeable and objectionable group delays in digital hearing aids. AB - Group delay in a digital signal processing (DSP) hearing aid may be perceived as an echo in the sound heard by a wearer listening to his or her own voice, due to a combination of unprocessed sound received at the ear through head and air pathways and delayed sound reaching the eardrum through the hearing aid. Depending on the amount, this delay may be audible or objectionable and can even result in auditory confusion. This study presents results from 18 subjects listening to their own voices through a DSP hearing aid with a variable group delay. The subjects varied the length of the delay and determined the amounts that were noticeable and objectionable as compared to the undelayed condition, while listening to their own amplified voices. Results indicated that a delay of 3 to 5 msec was noticeable to the listeners in 76 percent of the trials, and a delay of longer than 10 msec was objectionable 90 percent of the time. PMID- 10858006 TI - Auditory neuropathy: case study with hyperbilirubinemia. AB - Auditory neuropathy (AN) has been described in the literature as presenting with a combination of audiometric findings that include elevated behavioral audiometric thresholds, auditory brainstem response findings that are not consistent with audiometric findings, poor speech recognition, and present otoacoustic emissions (OAEs) and/or cochlear microphonics. Since the availability of clinical OAE testing, AN has come to be identified with increasing frequency; however, incidence and prevalence figures are unavailable. There is a great deal of discussion about the accurate diagnosis of AN, its characteristics, and its treatment. Some of this discussion is occurring on the Internet and over the telephones. The need to continue to provide information in accessible peer reviewed journals is paramount. Following a review of the literature, a case study is presented of a boy who was diagnosed with AN as a newborn. He experienced hyperbilirubinemia and other neonatal health complications. His educational intervention was managed elsewhere until recently. Information is presented about the progression of the case over a 5-year period that includes audiologic data and communication development results. PMID- 10858007 TI - More on CAPD. Central Auditory Processing Disorders. PMID- 10858008 TI - HIV-related tuberculosis in a transgender network--Baltimore, Maryland, and New York City area, 1998-2000. AB - During June-August 1998, the Tuberculosis (TB) Control Program of the Baltimore City Health Department (BCHD) identified four cases of TB among young black men. Three of these men also had human immunodeficiency virus (HIV) infection. The four reported belonging to a social network of transgender persons (i.e., persons who identify with or express a gender and/or sex different from their biologic sex) (1). By October 1998, test results on Mycobacterium tuberculosis isolates from the four men demonstrated a matching 11-band DNA fingerprint pattern (2), suggesting that these case-patients were epidemiologically linked. This report describes the public health investigation of these TB case-patients to identify contacts in Baltimore and the New York City area (NYC); the findings suggest that an interstate outbreak of TB has occurred within a social network that includes transgender persons. PMID- 10858009 TI - Escherichia coli O111:H8 outbreak among teenage campers--Texas, 1999. AB - In June 1999, the Tarrant County Health Department reported to the Texas Department of Health (TDH) that a group of teenagers attending a cheerleading camp during June 9-11 became ill with nausea, vomiting, severe abdominal cramps, and diarrhea, some of which was bloody. Two teenagers were hospitalized with hemolytic uremic syndrome (HUS), and two others underwent appendectomies. Routine stool cultures from eight ill persons failed to yield a pathogen. Stools subsequently were sent to laboratories at the Texas Department of Health and CDC, where Escherichia coli O111:H8 was isolated from two specimens. This report summarizes the investigation of this outbreak. PMID- 10858010 TI - Public health aspects of the Rainbow Family of Living Light annual gathering- Allegheny National Forest, Pennsylvania, 1999. AB - The Rainbow Family of Living Light (RFLL) is a loosely organized group that developed out of the late 1960s counterculture movement. RFLL has had a 2-week "Gathering for World Peace and the Healing of the Earth" in a different national forest each summer since 1972. For the June 21-July 10, 1999, gathering, RFLL selected the Allegheny National Forest in Pennsylvania. The site was not accessible by vehicle and was an hour's walk to the nearest road. No sanitary facilities were available, and water from streams was consumed without treatment. Approximately 20,000 persons attended from the United States and several foreign countries. The state health department requested federal assistance to establish and maintain public health surveillance and to advise on outbreak prevention and control. This report describes the public health aspects of the gathering and presents recommendations for the management of health risks at large outdoor events. PMID- 10858011 TI - Prevalence of leisure-time physical activity among overweight adults--United States, 1998. AB - In the United States, overweight and obesity have reached epidemic proportions among all segments of the population and regions of the country (1). Obesity is a risk factor for numerous chronic health conditions and weight loss can reduce risk factors for these conditions (2). National guidelines recommend that weight reduction should involve reducing calorie intake and increasing physical activity (3). The National Heart, Lung, and Blood Institute's clinical guidelines (3) and the federal dietary guidelines for Americans (4) recommend at least 30 minutes of physical activity on most days of the week for all healthy adults. To assess patterns of physical activity among overweight U.S. adults trying to lose weight, and to estimate the proportion who engage in leisure-time physical activity (LTPA) from selected demographic groups, CDC analyzed data from the 1998 Behavioral Risk Factor Surveillance System (BRFSS). This report summarizes the results of that analysis, which indicate that two thirds of overweight persons trying to lose weight reported using physical activity as a strategy for weight loss; however, only one fifth reported being active at recommended levels. PMID- 10858012 TI - Inducible cyclooxygenase-2 gene expression in the human thyroid epithelial cell line Nthy-ori3-1. AB - OBJECTIVE: To investigate whether the genes encoding Cyclooxygenase-1 and -2 are expressed in thyroid epithelial cells, in vitro. MATERIALS AND METHODS: COX-1/-2 gene expression was examined in the thyroid epithelial cell line Nthy-ori3-1 using semi-quantitative RT-PCR and Western blot analysis. ELISAs were employed to assess whole cell COX-enzyme activity, PGE2 and IL-6 formation. RESULTS: In response to IL-1beta and TNF-alpha combined cells of the thyroid epithelial cell line Nthy-ori3-1 secreted marked amounts of PGE2 in a time-dependent fashion. This is attributed to increased levels of COX-2 specific mRNA, increased amounts of COX-2 protein and COX enzyme activity in the absence of detectable COX-1 protein. The inhibition of the induced COX enzyme activity by the selective COX-2 inhibitor NS-398 demonstrated the presence of COX-2 pharmacologically. The expression of the COX-2 gene was also accompanied by a marked induction of IL-6 formation, a well described inflammatory response of thyroid epithelial cells. CONCLUSIONS: Our observation presents first evidence that COX-2 gene expression is inducible in thyroid epithelial cells, in vitro, upon stimulation with the pro inflammatory cytokines IL-1beta and TNF-alpha. This finding may indicate that thyroid epithelial cells could play an inflammatory controlling role perhaps during auto-immune thyroid diseases. PMID- 10858013 TI - ONO-4817, an orally active matrix metalloproteinase inhibitor, prevents lipopolysaccharide-induced proteoglycan release from the joint cartilage in guinea pigs. AB - OBJECTIVE AND DESIGN: To evaluate the effect of a newly developed inhibitor of matrix metalloproteinases (MMPs), ONO-4817, on the degradation of cartilage in the guinea pig arthritis model. MATERIALS: 42 guinea pigs were used in the arthritis model. TREATMENT: Lipopolysaccharide (LPS) was injected into guinea pig knee joints. The content of proteoglycan released in synovial cavity was measured as a marker of cartilage degradation. ONO-4817, dexamethasone or indomethacin were administered orally to the animals. RESULTS: ONO-4817 showed a broad inhibitory activity against MMPs except MMP-1 and MMP-7. The oral administration of ONO-4817 dose-dependently suppressed the release of proteoglycan from the cartilage of the knee joints. CONCLUSION: This study suggests the possibility that a novel MMP inhibitor, ONO-4817 may have a therapeutic utility for MMP related diseases. PMID- 10858014 TI - Clinical consequences of mast cell heterogeneity. AB - The heterogeneous morphological, biochemical and functional characteristics of mast cells from different species and from different tissue sites in the same species have been described for over 30 years. Far from being mere histochemical or pharmacological curiosities these differences have far reaching implications for therapeutic practice. This review concentrates on two important areas affected by mast cell heterogeneity, those of adverse reactions to therapeutic agents and the efficacy of anti-allergy therapy. In vitro studies of preformed and de novo synthesised mediator release have demonstrated a wide variability in the response of basophils and isolated mast cells to anti-allergy drugs and therapeutic agents such as radiographic contrast media, general anaesthetics, opioids and muscle relaxants. This heterogeneity is not limited to the mast cell's tissue of origin as there is also variability in the response of basophils and mast cells from different donors to the same drug or agent. These data have considerable clinical implications for the study of adverse drug reactions and the design of novel anti-allergic drugs. PMID- 10858015 TI - The suppression of rat collagen-induced arthritis and inhibition of macrophage derived mediator release by liposomal methotrexate formulations. AB - OBJECTIVE AND DESIGN: This study was designed to determine whether liposomes are suitable vehicles for the delivery of methotrexate (MTX-gamma-DMPE) for arthritis therapy. MATERIAL OR SUBJECTS: Liposomal formulations containing either egg lecithin (EPC), cholesterol (CHOL) and phosphatidic acid (PA) (MTX-EPC) or distearoylphosphatidylcholine (DSPC), CHOL and distearoylphosphatidylethanolamine conjugated to polyethyleneglycol (PEG) (MTX-PEG) were employed. Rat peritoneal macrophages (rPM phi) were used to test the mechanism of action of these liposomes in vitro, whilst, the rat collagen-induced arthritis (CIA) model was used to evaluate the in vivo efficacy of MTX-EPC and MTX-PEG. TREATMENT: In vitro, rPM phi were incubated with liposomal MTX concentrations ranging from 0 to 15 microg/well. In vivo, rats were given 4 daily intravenous injections of liposomal MTX (2.5 mg/Kg). METHODS: IL-1beta and prostaglandin-E2 (PGE2) release from rPM phi were quantified by immunoradiometric assay. Arthritis progression, in vivo, was measured by serial clinical score and hind paw diameter measurements. RESULTS: MTX-EPC and MTX-PEG respectively (15 microg of MTX and 0.15 mg of lipid) were powerful inhibitors of both IL-1beta (77 +/- 2.3%; 79 +/- 4.0%) and PGE2 (75.5 +/- 4.9%; 68.5 +/- 2.3%) release (mean +/- SEM % inhibition) from lipopolysaccaride stimulated rPM phi. In vivo, only MTX-EPC exerted an anti inflammatory effect, clinical score (p < 0.001) and paw diameter (p < 0.001) measurements being significantly lower than in control rats, after 2 days treatment. CONCLUSIONS: MTX-EPC and MTX-PEG are potent inhibitors of pro inflammatory mediators in vitro, but liposomes with long circulation times do not appear to have therapeutic potential for treating arthritis in vivo. PMID- 10858016 TI - Proinflammatory cytokines trigger MUC gene expression and mucin release in the intestinal cancer cell line LS180. AB - OBJECTIVES AND DESIGN: Proinflammatory cytokines and a defective mucus layer are involved in the pathogenesis of colitis. Therefore, we determined cytokine effects on MUC gene expression and mucin secretion. MATERIALS AND METHODS: LS180 cells were characterized by light and electron microscopy and subsequently exposed to interleukin 1 (IL-1, 1 ng/ml), interleukin 6 (IL-6, 10 ng/ml), or tumor necrosis factor-alpha (TNFalpha, 10 ng/ml). MUC gene (MUC2, MUC5AC, MUC5B, MUC6) mRNA expression was assessed by RT-PCR, the encoded proteins were identified by immunocytochemistry and Western blotting, and the released mucins were isolated and chromatographically characterized. RESULTS: Thirty to 40% of the cells contained intracellular mucin granules. Incubation with IL-1 transiently stimulated the mRNA expression of MUC2 and MUC5AC, whereas IL-6 induced an early response of MUC2, MUC5B and MUC6. TNFalpha upregulated the expression of MUC2 and MUC5B for 3 hours, and had no effect on the expression of MUC 5AC and MUC6. Immunocytochemistry and Western blotting confirmed TNFalpha effects on MUC2 and MUC5AC on the protein levels. All cytokines stimulated the release of less glycosylated mucins and considerably modulated their carbohydrate composition. CONCLUSION: Our data demonstrate differential cytokine effects on mucin synthesis, secretion and composition. These alterations may contribute to the defective mucus layer in colitis. PMID- 10858017 TI - Participation of endogenous endothelins in delayed eosinophil and neutrophil recruitment in mouse pleurisy. AB - OBJECTIVE: Investigate the role of endothelins in leukocyte recruitment in allergic and non allergic inflammation. METHODS: Pleurisy was induced in mice by intrathoracic injection of ovalbumin (OVA; in sensitized animals), E. coli LPS, carrageenan, Mycobacterium bovis (BCG) or zymosan. Animals were treated with BQ 123 or BQ-788 (1.5-150 pmol/cavity), or intravenously with bosentan (30 mg/kg). RESULTS: None of the ET receptor antagonists modified early neutrophil recruitment (at 4 h) induced by OVA, LPS, carrageenan, BCG or zymosan or plasma leakage caused by carrageenan or zymosan. Mononuclear and eosinophil accumulation triggered by OVA were reduced by BQ-123 (150 pmol/cavity) or bosentan (68 and 43% inhibition of eosinophilia), but unaffected by BQ-788. BQ-123 and bosentan also inhibited LPS increases in neutrophil (by 67 and 40%) and eosinophil (by 63 and 74%) at 24 h. CONCLUSIONS: Endothelins, acting via ETA receptors, play a role in late eosinophil and neutrophil accumulation (24 h), but not in the acute (4 h) neutrophilic response. PMID- 10858018 TI - Induction of the activation-related antigen CD69 on human eosinophils by type IIA phospholipase A2. AB - OBJECTIVE: High levels of human type IIA phospholipase A2 (PLA2-IIA) have been found in the eosinophil-mediated inflammation sites, although the pathophysiological role of PLA2-IIA in the eosinophil activation has remained poorly understood. We investigated the effects of PLA2-IIA on eosinophil activation. METHODS: Eosinophils were incubated with recombinant human PLA2-IIA or other stimuli, and then eosinophil peroxidase (EPO) (by colorimetric assay) and leukotriene C4 (by enzyme immunoassay) released in the incubation buffer were measured. Expression of CD11b and CD69 on the cell surface was also measured by flow cytometry (by mean fluorescence intensity (MFI)). EPO, LTC4, and CD11b are thought to be markers for early phase activation (occurred in an hour after stimulation), and CD69 is to be a marker for late phase activation (occurred after several hours). RESULTS: While PLA2-IIA (5 microg/ml) did not induce any early phase activation, it induced significant expression of an activation related antigen, CD69, on human blood eosinophils. The PLA2-IIA, when enzymatically inactivated by either p-bromophenacyl bromide or EDTA, lost its effect on the CD69 induction. Similarly to PLA2-IIA, several lysophospholipids (1 microg/ml) also induced CD69 on eosinophils significantly (control, 0.71 +/- 0.11; PLA2-IIA, 3.29 +/- 0.37*; lysophosphatidic acid, 2.57 +/- 0.43*; specific MFI +/- S.E.M., n = 4, * indicate p < 0.05 vs. control). CONCLUSIONS: PLA2-IIA induces CD69 expression on the eosinophils through its catalytic activity at least partly via the enzymatic products such as several lysophospholipids from the eosinophil membrane phospholipids. PLA2-IIA may contribute to the eosinophilic inflammation synergistically with other factors. PMID- 10858019 TI - New cyclopentenone fatty acids formed from linoleic and linolenic acids in potato. AB - [1-14C]Linoleic acid was incubated with a whole homogenate preparation from potato stolons. The reaction product contained four major labeled compounds, i.e., the alpha-ketol 9-hydroxy-10-oxo-12(Z)-octadecenoic acid (59%), the epoxy alcohol 10(S),11(S)-epoxy-9(S)-hydroxy-12(Z)-octadecenoic acid (19%), the divinyl ether colneleic acid (3%), and a new cyclopentenone (13%). The structure of the last-mentioned compound was determined by chemical and spectral methods to be 2 oxo-5-pentyl-3-cyclopentene-1-octanoic acid (trivial name, 10-oxo-11-phytoenoic acid). Steric analysis demonstrated that the relative configuration of the two side chains attached to the five-membered ring was cis, and that the compound was a racemate comprising equal parts of the 9(R),13(R) and 9(S),13(S) enantiomers. Experiments in which specific trapping products of the two intermediates 9(S) hydroperoxy-10(E),12(Z)-octadecadienoic acid and 9(S),10-epoxy-10,12(Z) octadecadienoic acid were isolated and characterized demonstrated the presence of 9-lipoxygenase and allene oxide synthase activities in the tissue preparation used. The allene oxide generated from linoleic acid by action of these enzymes was further converted into the cyclopentenone and alpha-ketol products by cyclization and hydrolysis, respectively. Incubation of [1-14C]linolenic acid with the preparation of potato stolons afforded 2-oxo-5-[2'(Z)-pentenyl]-3 cyclopentene-1-octanoic acid (trivial name, 10-oxo-11,15(Z)-phytodienoic acid), i.e., an isomer of the jasmonate precursor 12-oxo-10,15(Z)-phytodienoic acid. Quantitative determination of 10-oxo-11-phytoenoic acid in linoleic acid-supplied homogenates of different parts of the potato plant showed high levels in roots and stolons, lower levels in developing tubers, and no detectable levels in leaves. PMID- 10858021 TI - Biased distribution of the branched-chain fatty acids in ceramides of vernix caseosa. AB - The compositions of ester- and amide-linked fatty acids from ceramides of human vernix caseosa were described with emphasis on the distribution of the branched chain fatty acid (BCFA). Two novel ceramides were isolated from vernix caseosa in the course of this study: the acylated type of esterified alpha-OH hydroxyacid/sphingosine ceramide (Cer[EAS]) and nonacylated type of non-OH fatty acid/hydroxysphingosine ceramide (Cer[NH]). Their chemical structures were identified by nuclear magnetic resonance and chemical procedure. The Cer[EAS] was an acylceramide and consisted of the highest concentrations of ester- and amide linked BCFA (62 and 67%, respectively). The iso- or anteiso-branching structures of the aliphatic chains were confirmed by the mass spectra of their picolinyl or pyrrolidide derivatives. As a whole, amide-linked fatty acids of ceramides 1-7 and Cer[NH] were normal types of straight-chain fatty acids with or without alpha or omega-hydroxylation. The BCFA concentrations of amide-linked fatty acids in these ceramides (ceramides 1-7 and Cer[NH]) were low and less than 10%. The BCFA thus occurred exclusively in a novel acylceramide of Cer[EAS] in the vernix caseosa. PMID- 10858020 TI - Fatty acid-specific, regiospecific, and stereospecific hydroxylation by cytochrome P450 (CYP152B1) from Sphingomonas paucimobilis: substrate structure required for alpha-hydroxylation. AB - Fatty acid alpha-hydroxylase from Sphingomonas paucimobilis is an unusual cytochrome P450 enzyme that hydroxylates the alpha-carbon of fatty acids in the presence of H2O2. Herein, we describe our investigation concerning the utilization of various substrates and the optical configuration of the alpha hydroxyl product using a recombinant form of this enzyme. This enzyme can metabolize saturated fatty acids with carbon chain lengths of more than 10. The Km value for pentadecanoic acid (C15) was the smallest among the saturated fatty acids tested (C10-C18) and that for myristic acid (C14) showed similar enzyme kinetics to those seen for C15. As shorter or longer carbon chain lengths were used, Km values increased. The turnover numbers for fatty acids with carbon chain lengths of more than 11 were of the same order of magnitude (10(3) min(-1)), but the turnover number for undecanoic acid (C11) was less. Dicarboxylic fatty acids and methyl myristate were not metabolized, but monomethyl hexadecanedioate and omega-hydroxypalmitic acid were metabolized, though with lower turnover values. Arachidonic acid was a good substrate, comparable to C14 or C15. The metabolite of arachidonic acid was only alpha-hydroxyarachidonic acid. Alkanes, fatty alcohols, and fatty aldehydes were not utilized as substrates. Analysis of the optical configurations of the alpha-hydroxylated products demonstrated that the products were S-enantiomers (more than 98% enantiomerically pure). These results suggested that this P450 enzyme is strictly responsible for fatty acids and catalyzes highly stereo- and regioselective hydroxylation, where structure of omega-carbon and carboxyl carbon as well as carbon chain length of fatty acids are important for substrate-enzyme interaction. PMID- 10858022 TI - Utilization of docosahexaenoic acid from intravenous egg yolk phospholipid. AB - Docosahexaenoic acid (DHA, 22:6n-3) is provided directly to human premature infants during parenteral nutrition from the egg yolk fraction of an intravenous fat emulsion. This study aimed to determine whether the high egg yolk phospholipid content of Intralipid 10% (IL 10%, Pharmacia, Uppsala, Sweden) relative to the standard emulsion Intralipid 20% (IL 20%, Pharmacia) could be a strategy to increase the delivery of DHA to the developing brain. Male, Large White piglets were randomly selected from sows 3 d after birth. Piglets were assigned to receive a 9-d continuous intravenous infusion commencing 5 d after birth of either Intralipid (IL) 10%, IL 20%, or Lipofundin S 20% (LFS; B. Braun, Melsungen, Germany). There were four piglets in each treatment group. IL 10% provides twice as much DHA as IL 20%, while LFS provides no DHA. Protein and other nutrients were provided enterally using a low-fat milk formula. After 9 d, animals were killed, and the fatty acid compositions of blood, liver, and cerebral cortex were analyzed. IL 10% infusion approximately doubled the amount of plasma phospholipid DHA (microg/mL of plasma) in comparison to IL 20%. However, red blood cells, liver, and cerebral cortex phospholipid DHA levels were indistinguishable between these two groups. LFS was associated with reduced levels of DHA in plasma, red blood cell and liver phospholipids in comparison to IL 20%. We conclude that infusion of additional phospholipid is an ineffective strategy for increasing DHA delivery to piglet tissues. This may be due to the formation of inert phospholipid particles in plasma. The data do not support the concept of using IL 10% as a means of providing additional DHA to premature human infants. PMID- 10858023 TI - High dietary 18:3n-3 increases the 18:3n-3 but not the 22:6n-3 content in the whole body, brain, skin, epididymal fat pads, and muscles of suckling rat pups. AB - The objective of this study was to test the hypothesis that increasing maternal dietary 18:3n-3 by decreasing the 18:2n-6/18:3n-3 ratio will increase the 18:3n-3 and 22:6n-3 content of the whole body, liver, skin (epidermis, dermis, and subcutaneous tissue), epididymal fat pads, and muscles (arms and legs) of 2-wk old rat pups. Sprague-Dawley dams at parturition were fed semipurified diets containing either a low (18:2n-6 to 18:3n-3 ratio of 24.7:1) or a high (1 8:2n-6 to 18:3n-3 ratio of 1.0:1) 18:3n-3 fatty acid content. During the first 2 wk of life, rat pups received only their dams' milk. Fatty acid composition of the pups' stomach contents (dams' milk), whole body, brain, liver, skin, epididymal fat pads, and muscles was determined. The stomach fatty acid composition of 18:3n 3 reflected the dams' diet. The content of 18:3n-3 in whole body, brain, liver, skin, epididymal fat pads, and muscles was significantly (P< 0.05) greater in rat pups fed the high compared with the low 18:3n-3 fatty acid diet. The 22:6n-3 content of the whole body, brain, skin, epididymal fat pads, and muscles was not quantitatively different in rat pups fed either the low or high 18:3n-3 fatty acid diet. The 20:5n-3 and 22:5n-3 content of the whole body, skin, and epididymal fat pads was significantly increased in rat pups fed the high compared with the low 18:3n-3 fatty acid diet. High content of 18:3n-3 was found in the skin of rat pups fed either a low or high 18:3n-3 fatty acid diet. These findings demonstrate that high maternal dietary 18:3n-3 significantly increases the 18:3n 3 but not the 22:6n-3 content of the whole body, brain, skin, epididymal fat pads, and muscles with approximately 39 and 41% of the whole body 18:3n-3 content being deposited in the skin of suckling rat pups fed either the low or high 18:3n 3 diet, respectively. PMID- 10858024 TI - Increased alpha-linolenic acid intake increases tissue alpha-linolenic acid content and apparent oxidation with little effect on tissue docosahexaenoic acid in the guinea pig. AB - The essential fatty acids do not have identical roles in nutrition. Linoleic acid (LA) accumulates throughout the body of most mammals, whereas alpha-linolenic acid (ALA) is rarely found in tissue lipids to the same extent as LA. It has been argued that this is the result of metabolism of ALA to docosahexaenoic acid (DHA) or that ALA is rapidly beta-oxidized to acetyl CoA and CO2. In this study, we consider the effect of high and low ALA levels on the tissue distribution of ALA and other n-3 polyunsaturated fatty acids (PUFA) in all tissues. Guinea pigs were fed one of two defined diets for 3 wk from weaning with both diets containing 1.8% (by weight) of LA and either 1.7% ALA or 0.03% ALA. The high ALA diet was associated with significantly increased ALA levels in all tissues except the brain and significantly increased levels of long-chain n-3 PUFA in all tissues except intestines, brain, carcass, and skin. The long-chain n-3 PUFA content of the whole body was less than 5% of that of the ALA content in both diet groups, and the major long-chain n-3 PUFA (>66% of total) in the body was 22:5n-3. The brain was the only tissue where the DHA content exceeded that of 22:5n-3. On the low ALA diet, there appeared to be conservation of ALA based on a comparison of the ratio of LA to ALA in the tissues compared with that in the diet. On the high ALA diet there was a loss of ALA relative to LA in the tissues compared with the diet. These studies suggest that the low levels of tissue ALA in the guinea pig are likely the result of beta-oxidation or excretion via the skin and fur rather than metabolism to DHA. PMID- 10858025 TI - Effect of n-3 polyunsaturated fatty acid supplementation on lipid peroxidation of rat organs. AB - The present study was undertaken in order to reexamine the effect of n-3 polyunsaturated fatty acid (PUFA)-rich diet supplementation on lipid peroxidation and vitamin E status of rat organs. Male Wistar rats were fed a diet containing safflower or fish oil at 50 g/kg diet and an equal amount of vitamin E at 59 mg/kg diet (1.18 g/kg oil; and 1.5 g/kg PUFA in safflower oil diet, and 4.3 g/kg PUFA in fish oil diet) for 6 wk. Fatty acid composition of total lipids of brain, liver, heart, and lung of rats fed fish oil was rich in n-3 PUFA, whereas that of each organ of rats fed safflower oil was rich in n-6 PUFA. The vitamin E levels in liver, stomach, and testis of the fish oil diet group were slightly lower than those of the safflower oil diet group, but the levels in brain, heart, lung, kidney, and spleen were not different between the two diet groups. The levels of phospholipid hydroperoxides were determined by the high-performance liquid chromatography-chemiluminescence method and the levels of thiobarbituric acid reactive substances (TBARS) were determined at pH 3.5 in the presence of butylated hydroxytoluene with or without EDTA. Levels of phospholipid hydroperoxides and TBARS in the brain, liver, heart, lung, kidney, spleen, stomach and testis of the fish oil diet group were similar to those of the safflower oil diet group. The results indicate that high fish oil intake does not induce increased levels of phospholipid hydroperoxides and TBARS in rat organs. PMID- 10858026 TI - Influence of sources of dietary oils on the life span of stroke-prone spontaneously hypertensive rats. AB - In recent studies, the life span of stroke-prone spontaneously hypertensive (SHRSP) rats was altered by a variety of dietary fats. It was relatively shorter in rats fed canola oil as the sole source of fat. The present study was performed to find out whether the fatty acid profile and the high content of sulfur compounds in canola oil could modulate the life span of SHRSP rats. SHRSP rats (47 d old, n = 23/group) were matched by body weight and systolic blood pressure and fed semipurified diets containing 10% canola oil, high-palmitic canola oil, low-sulfur canola oil, soybean oil, high-oleic safflower oil, a fat blend that mimicked the fatty acid composition of canola oil, or a fat blend high in saturated fatty acids. A 1% sodium chloride solution was used as drinking water to induce hypertension. After consuming the diets for 37 d, five rats from each dietary group were killed for collection of blood and tissue samples for biochemical analysis. The 18 remaining animals from each group were used for determining their life span. The mean survival time of SHRSP rats fed canola oil (87.4+/-4.0 d) was not significantly different (P > 0.05) from those fed low sulfur canola oil (89.7+/-8.5 d), suggesting that content of sulfur in canola oil has no effect on the life span of SHRSP rats. The SHRSP rats fed the noncanola oil-based diets lived longer (mean survival time difference was 6-13 d, P < 0.05) than those fed canola and low-sulfur canola oils. No marked differences in the survival times were observed among the noncanola oil-based groups. The fatty acid composition of the dietary oils and of red blood cells and liver of SHRSP rats killed after 37 d of treatment showed no relationship with the survival times. These results suggest that the fatty acid profile of vegetable oils plays no important role on the life span of SHRSP rat. However, phytosterols in the dietary oils and in liver and brain were inversely correlated with the mean survival times,indicating that the differential effects of vegetable oils might be ascribed, at least partly, to their different phytosterol contents. PMID- 10858027 TI - Dietary fat-induced suppression of lipogenic enzymes in B/B rats during the development of diabetes. AB - This study was designed to determine the level of inhibition of gene transcription by the reduction in insulin levels upon the onset of diabetes in spontaneously diabetic B/B rats and if reducing the level of polyunsaturated fatty acids (PUFA) in the diet will increase lipogenic enzyme activity. Control (eight animals per group) and spontaneously diabetic B/B male weanling rats (25 animals per group) were fed semipurified diets containing 20% (w/w) fat of either low (0.25) or high (1.0) polyunsaturated to saturated (P/S) fatty acid ratio. Rats were killed at the onset of diabetes [blood glucose level of approximately/= 100 mg/dL (5.55 mM)] and as they became highly diabetic [blood glucose level of approximately/= 400 mg/dL (22.22 mM)]. Total RNA was extracted from liver, and the relative amount of mRNA coding for fatty acid synthase (FAS), acetyl-CoA carboxylase, malic enzyme, pyruvate kinase, and phosphoenolpyruvate carboxykinase was determined. Liver enzyme activities were also measured. The mRNA levels for FAS, acetyl-CoA carboxylase, and malic enzyme decreased compared to control animals. The mRNA level for pyruvate kinase decreased at the onset of diabetes as compared to control animals. Feeding animals the low P/S diet treatment elevated the level of mRNA and lipogenic enzyme activity compared to animals fed the high P/S diet treatment, suggesting that the effect of PUFA on lipogenic enzymes is through a direct effect on gene expression. PMID- 10858028 TI - Effects of dietary phenolic compounds on tocopherol, cholesterol, and fatty acids in rats. AB - The effects of the phenolic compounds butylated hydroxytoluene (BHT), sesamin (S), curcumin (CU), and ferulic acid (FA) on plasma, liver, and lung concentrations of alpha- and gamma-tocopherols (T), on plasma and liver cholesterol, and on the fatty acid composition of liver lipids were studied in male Sprague-Dawley rats. Test compounds were given to rats ad libitum for 4 wk at 4 g/kg diet, in a diet low but adequate in vitamin E (36 mg/kg of gamma-T and 25 mg/kg of alpha-T) and containing 2 g/kg of cholesterol. BHT significantly reduced feed intake (P < 0.05) and body weight and increased feed conversion ratio; S and BHT caused a significant enlargement of the liver (P < 0.001), whereas CU and FA did not affect any of these parameters. The amount of liver lipids was significantly lowered by BHT (P < 0.01) while the other substances reduced liver lipid concentrations but not significantly. Regarding effects on tocopherol levels, (i) feeding of BHT resulted in a significant elevation (P< 0.001) of alpha-T in plasma, liver, and lung, while gamma-T values remained unchanged; (ii) rats provided with the S diet had substantially higher gamma-T levels (P < 0.001) in plasma, liver, and lung, whereas alpha-T levels were not affected; (iii) administration of CU raised the concentration of alpha-T in the lung (P < 0.01) but did not affect the plasma or liver values of any of the tocopherols; and (iv) FA had no effect on the levels of either homolog in the plasma, liver, or lung. The level of an unknown substance in the liver was significantly reduced by dietary BHT (P < 0.001). BHT was the only compound that tended to increase total cholesterol (TC) in plasma, due to an elevation of cholesterol in the very low density lipoprotein + low density lipoprotein (VLDL + LDL) fraction. S and FA tended to lower plasma total and VLDL + LDL cholesterol concentrations, but the effect for CU was statistically significant (P < 0.05). FA increased plasma high density lipoprotein cholesterol while the other compounds reduced it numerically, but not significantly. BHT, CU, and S reduced cholesterol levels in the liver TC (P < 0.001) and percentages of TC in liver lipids (P < 0.05). With regard to the fatty acid composition of liver lipids, S increased the n-6/n-3 and the 18:3/20:5 polyunsaturated fatty acids (PUFA) ratios, and BHT lowered total monounsaturated fatty acids and increased total PUFA (n-6 + n-3). The effects of CU and FA on fatty acids were not highly significant. These results suggest some in vivo interactions between these phenolic compounds and tocopherols that may increase the bioavailability of vitamin E and decrease cholesterol in rats. PMID- 10858029 TI - Cholesterol-lowering effects of guar gum: changes in bile acid pools and intestinal reabsorption. AB - Soluble fibers such as guar gum (GG) may exert cholesterol-lowering effects. It is generally accepted that bile acid (BA) reabsorption in portal blood is reduced, thus limiting the capacity of BA to down-regulate liver cholesterol 7alpha-hydroxylase, the rate-limiting enzyme of BA synthesis. In the present work, rats were adapted to fiber-free (FF) or 5% GG diets (supplemented or not with 0.25% cholesterol), to investigate various aspects of enterohepatic BA cycling. GG in the diet at a level of 5% elicited a significant lowering of plasma cholesterol during the absorptive period, in cholesterol-free (-13%) or 0.25% cholesterol (-20%) diet conditions. In rats adapted to the GG diets, the small intestinal and cecal BA pools and the ileal vein-artery difference for BA were markedly enhanced; reabsorption in the cecal vein was also enhanced in these rats. [14C]Taurocholate absorption, determined in perfused ileal segments, was not significantly different in rats adapted to the FF or GG diet, suggesting that a greater flux of BA in the ileum might support a greater ileal BA reabsorption in rats adapted to the GG diet. In contrast, capacities for [14C]cholate absorption from the cecum at pH 6.5 were higher in rats adapted to the GG diet than to the FF diet. Acidification of the bulk medium in isolated cecum (from pH 7.1 down to pH 6.5 or 5.8) or addition of 100 mM volatile fatty acids was also found to stimulate cecal [14C]cholate absorption. These factors could contribute to accelerated cecal BA absorption in rats fed the GG diet. The effects of GG on steroid fecal excretion thus appear to accompany a greater intestinal BA absorption and portal flux to the liver. These results suggest that some mechanisms invoked to explain cholesterol-lowering effect of fibers should be reconsidered. PMID- 10858030 TI - Changes in liver fatty acid unsaturation after partial hepatectomy in the rat. AB - The purpose of this study was to assess changes in the degree of fatty acid unsaturation in rat liver after partial hepatectomy. This is the first study in which liver fatty acid unsaturation has been analyzed over a long period of regeneration until day 28 after operation. The relationship between changes in unsaturation and fatty acid composition in the regenerating liver were also investigated in this study. Proton nuclear magnetic resonance spectroscopy revealed significantly elevated levels of unsaturation with a maximum on day 5 after partial hepatectomy, compared with untreated controls (11.72+/-0.55 vs. 11.05+/-0.26%, P < 0.05). No significant changes in unsaturation were found in day 1 regenerating liver, which is rich in absolute amounts of fatty acids. Based on gas-liquid chromatography, the relative amounts of oleic acid (18:1n-9) and linoleic acid (LA; 18:2n-6) were increased, while polyunsaturated fatty acids such as arachidonic acid (20:4n-6) and docosahexaenoic acid (DHA; 22:6n-3) were decreased on day 1. On the other hand, on day 5 of regeneration, while most fatty acids were returning to their preoperative control levels, only DHA was higher than the control value (7.69+/-0.58 vs. 5.57+/-0.37%, P < 0.001). The high levels of unsaturation on day 5 were found to be partly due to the increase in DHA. The findings suggest that some significant signals are transmitted during the regeneration process owing to alterations in the membrane structure by the high levels of fatty acid unsaturation and the increase in DHA levels on day 5 after partial hepatectomy. PMID- 10858031 TI - Lipid composition of erythrocytes and thrombocytes of a subantarctic seabird, the king penguin. AB - Phospholipid (PL) compositions and fatty acid (FA) patterns of PL were determined in the erythrocytes and blood thrombocytes of a seabird, the king penguin, living in the subantarctic area and feeding on prey rich in n-3 polyunsaturated FA. Results were compared between birds in three different physiological states (breeding and molting adults, chicks) to those reported for other birds. In erythrocytes, the ratios of cholesterol to PL and of sphingomyelin to phosphatidylcholine (PC) were lower than in other birds. The PL distribution was similar to those previously reported in the hen and pigeon. In contrast to other birds, cardiolipin levels were unexpectedly high (4%). Very long chain n-3 FA were abundant (13-27%) in phosphatidylethanolamine (PE), phosphatidylserine and PC, probably in relation to the natural diet of these birds. Among n-3 FA, 22:6n 3 was the most abundant in all PL (2-20%), whereas the highest levels of arachidonic acid were observed in PE (14%). In thrombocytes, the PL distribution and FA composition of the main PL (PC, PE) differed from those of erythrocytes, and in particular, levels of n-3 FA (9-12%) were 1.5-2 times lower. The highest levels of arachidonic acid were found in phosphatidylinositol (24%). The lipid profile of penguin erythrocytes could contribute to the efficiency of blood circulation and oxygen delivery in microvascular beds, thus favoring diving capacity of these animals. Our observations do not support the hypothesis of a common origin of avian thrombocytes and erythrocytes. PMID- 10858032 TI - Esterification of polyunsaturated fatty acids by various forms of immobilized lipase from Rhizomucor miehei. AB - Ethyl esterification specificity of a lipase from Rhizomucor miehei for polyunsaturated fatty acids (PUFA) was compared at 1 and 100 mM to study molecular recognition of PUFA. The chemical shift of methylene adjacent to carboxyl groups in the nuclear magnetic resonance spectrum of docosahexaenoic acid (DHA) in ethanol moved to a lower magnetic field as the concentration of DHA increased, suggesting that the degree of dissociation of DHA decreased. Specificity constants or apparent second-order rate constants (Vmax/Km or catalytic power) for 1 mM esterification by immobilized lipases were higher than the native lipase. Immobilized hydrophobic carrier of low mass transfer resistance for the esterification substrate may improve maximal velocity and affinity for the substrate. Higher specificity constants for 1 mM substrates were observed using immobilized lipases fixed on an anion exchange resin with glutaraldehyde and on a cation exchange carrier with carbodiimide. Activity yields measured with 1 mM PUFA substrate were high. For the substrates at a concentration of 100 mM, higher specific constants with these bifunctional reagents were not observed but higher activity yields were found. PMID- 10858033 TI - Cold light, heat burn. PMID- 10858034 TI - Re: Accuracy of burn size estimation and subsequent fluid resuscitation prior to arrival at the Yorkshire Regional Burns Unit. A three year retrospective study. PMID- 10858035 TI - Development and function of B-1 cells. AB - Results from immunoglobulin-transgenic mice and BCR-mutant mice have been widely interpreted in recent years as supporting a simple 'activation' model for the origin of CD5+/B-1 B cells. However cell transfer experiments over 10 years ago and recent work investigating pre-BCR signaling suggest striking differences between B cell development in fetal liver and adult bone marrow, lending support for a 'lineage' model that we favor. Recent progress has been made relating to the development and function of the CD5+/B-1 B cell subpopulation in mice; the data can be viewed in the context of the generation of this subpopulation by a distinctive fetal B cell developmental process. PMID- 10858036 TI - Optimal screening of surface-displayed polypeptide libraries. AB - Cell surface display of polypeptide libraries combined with flow cytometric cell sorting presents remarkable potential for enhancement of protein-ligand recognition properties. To maximize the utility of this approach, screening and purification conditions must be optimized to take full advantage of the quantitative feature of this technique. In particular, discrimination of improved library mutants from an excess of wild-type polypeptides is dependent upon an effective screening methodology. Fluorescence discrimination profiles for improved library mutants were derived from a mathematical model of expected cell fluorescence intensities for polypeptide libraries screened with fluorescent ligand. Profiles for surface-displayed libraries under equilibrium or kinetic screening conditions demonstrate distinct discrimination optima from which optimal equilibrium and kinetic screening parameters were derived. In addition, a statistical model of low cytometrically analyzed cell populations indicates the importance of low-stringency sorting followed by amplification through regrowth and resorting at increased stringency. This analysis further yields quantitative recommendations for cell-sorting stringency. PMID- 10858037 TI - Poor reversal of low molecular weight heparin by protamine. PMID- 10858038 TI - Isochromosome 12p in mediastinal centroblastic lymphoma. PMID- 10858039 TI - Prevalence of the prothrombin gene G20210A mutation in Azerbaijan. PMID- 10858040 TI - The effect of a high water intake on the kidney's ability to concentrate the urine in man. 1957. PMID- 10858041 TI - Gentle microinjection for myeloid cells using SLAM. PMID- 10858042 TI - Proceedings of the Workshop on Biomathematical Models of Circadian Rhythmicity, Sleep Regulation and Neurobehavioral Function in Humans held May 18-21, 1999, in Dedham, Massachusetts. PMID- 10858043 TI - Reply to technical note: nonlinear interactions between circadian and homeostatic processes: models or metrics? PMID- 10858044 TI - [Record no. 34: hSNF5/INI1 (SMARCB1)]. PMID- 10858045 TI - [Cannabinoids, among others, send malignant glial tumors to nirvana. . ]. PMID- 10858046 TI - [Importance of thalidomide in the treatment of cancer]. PMID- 10858047 TI - [The Bezwoda affair]. PMID- 10858048 TI - Commentary: patients, preferences, and evidence. PMID- 10858049 TI - Childhood obesity. Aim should be weight maintenance, not loss. PMID- 10858050 TI - Childhood obesity. Breast feeding is important. PMID- 10858051 TI - Childhood obesity. Opportunity for physical activity has been lost. PMID- 10858052 TI - Childhood obesity. Health schools approach is needed. PMID- 10858053 TI - Childhood obesity. Interventions should be critically evaluated. PMID- 10858054 TI - Visual field defect associated with vigabatrin. Many more patients may be affected than were found in study. PMID- 10858055 TI - Visual field defect associated with vigabatrin. Method of estimating prevalence was inappropriate. PMID- 10858056 TI - Visual field defect associated with vigabatrin. Means of selecting patients was misleading. PMID- 10858057 TI - Guideline for prescribing vigabatrin in children has been revised. Vigabatrin Paediatric Advisory Group. PMID- 10858058 TI - Association between SSRIs and upper gastrointestinal bleeding. Coprescription of antiulcer drugs with SSRIs is fairly common. PMID- 10858059 TI - Association between SSRIs and upper gastrointestinal bleeding. SSRIs are no more likely than other drugs to cause such bleeding. PMID- 10858060 TI - Association between SSRIs and upper gastrointestinal bleeding. Self treatment with non-steroidal drugs may be confounding factor. PMID- 10858061 TI - Heartburn treatment in primary care. Prescribing omeprazole would conflict with desire to control prescribing costs. PMID- 10858062 TI - Heartburn treatment in primary care. Step up approach to management is best. PMID- 10858063 TI - Heartburn treatment in primary care. Study's results seem to be promotional rather than evidence based. PMID- 10858064 TI - Incidence of congenital rubella in Greece has decreased. PMID- 10858065 TI - Points for reading as part of continuing medical education. e-Journal club allows doctors to gain points. PMID- 10858066 TI - Points for reading as part of continuing medical education. How about earning points for continuing medical entertainment? PMID- 10858067 TI - Is physical therapy effective? Catch-22 of meta-analysis. PMID- 10858068 TI - Is concurrent twice-daily thoracic radiotherapy with chemotherapy the reference treatment for small cell lung cancer? PMID- 10858069 TI - [Cisplatinum polyneuropathy]. PMID- 10858070 TI - Mechanisms and biological significance of pulsatile hormone secretion. Symposium proceedings. London, United Kingdom, 2-4 March 1999. PMID- 10858071 TI - Antibody mAb33 from transduction laboratories detects human CD95L in ELISA but not in immunoblots. PMID- 10858072 TI - mAb33 from transduction laboratories specifically binds human CD95-L. PMID- 10858073 TI - Apoptosis and autoimmunity. PMID- 10858074 TI - Effect of saline irrigation flow rate on temperature profile during cooled radiofrequency ablation. AB - INTRODUCTION: Cooled radiofrequency ablation has been developed clinically for the treatment of ventricular tachycardia. Although clinical studies employ a constant saline flow rate for cooling, we hypothesized that varying the flow rates might optimize the temperature profile at depth. METHODS: In excised ovine left ventricle, we compared the temperature profile from a catheter tip electrode thermocouple to those placed at depths of 0.0 mm, 1.0 mm, and 2.0 mm. We compared the following settings: 20 Watts without flow, 20 Watts with 0.3 cc/sec flow, 20 Watts with 0.5cc/sec flow, and 70C surface temperature without flow (temperature control). RESULTS: The temperatures decreased from 77.5 +/-10.5 degrees C, 91.7+/ 6.3 degrees C, 65.5 +/- 11.8 degrees C, and 52.5 +/- 11.8 degrees C at 20W without saline irrigation at the tip, 0.0mm, 1.0mm, and 2.0 mm, respectively, to 33.0+/-1.4 degrees C, 63.4 +/- 7.0 degrees C, 57.1+/-5.8 degrees C, 49.9+/-5.8 degrees C+ at 20W with 0.5 ml/sec flow (*p<0.01, +p = 0.09). The lesion volumes were 79.6mm3 for 20W without flow, 64.1 mm3 for 20W with 0.3 ml/sec flow, 47.5 mm3 for 20W with 0.5 ml/sec flow, and 28.6 mm3 for temperature control. CONCLUSIONS: We conclude that 1) the temperature profile greatly depends upon the rate of saline flow for cooling; 2) at high flow rates, the 0.0 mm and 1.0 mm temperatures are similar; 3) even at high irrigation rates, lesion size is greater than for temperature control; 4) the tip temperature significantly underestimates the surface temperature and improved methods of measuring temperature are needed. PMID- 10858075 TI - Sonoluminescence: 1996-1998. PMID- 10858076 TI - Electrochemiluminescence: 1996-1998. PMID- 10858077 TI - The 4th International AIDS Malignancy Conference. Bethesda, Maryland, USA. May 16 18, 2000. Abstracts. PMID- 10858078 TI - Publishing in the journals of the APS: why are authors charged fees? PMID- 10858079 TI - Publishing in the journals of the APS: why are authors charged fees? PMID- 10858080 TI - Determination of plasma adrenomedullin concentrations with commercial radioimmunoassay kits: a note of caution. PMID- 10858081 TI - Assessing virologic efficacy of combination lamivudine and low-dose hepatitis B immune globulin posttransplantation with the ultrasensitive digene hybrid capture II assay. PMID- 10858082 TI - Whither living donor liver transplantation? PMID- 10858083 TI - EMBO workshop--"Molecular and Cellular Gerontology": Sepiano/Switzerland, September 18-22, 1999. AB - The EMBO Workshop "Molecular and Cellular Gerontology" provided a comprehensive analysis of recent developments in experimental gerontology. Various animal models for aging were presented. The focus of the conference was on the role of oxidative stress in aging at the cellular and organismal level. Evolution of aging was another topic discussed in the course of the meeting. Another focal point of the meeting was on age-related changes in cellular repair systems, in particular DNA repair and protein repair. Emphasis was also placed on the role of mitochondrial function and mitochondrial DNA mutations in the aging process. These molecular approaches were complemented by a session on age-related diseases, including aging of the immune system, cardiovascular disease and Alzheimer's disease. PMID- 10858084 TI - Which routes do Plasmodium sporozoites use for successful infections of vertebrates? PMID- 10858085 TI - Paradoxical response to radiotherapy in malignant spinal cord compression. PMID- 10858086 TI - Gastrointestinal bleeding in advanced cancer patients. PMID- 10858087 TI - Laxative effects of fresh baker's yeast. PMID- 10858088 TI - 'Thumb localizing test' for detecting a lesion in the posterior column-medical lemniscal system. PMID- 10858089 TI - Publishing in the Journals of the APS: Why Are Authors Charged Fees? PMID- 10858091 TI - Hospice care. PMID- 10858090 TI - [Otorhinolaryngological disorders due to psychological stress in children]. PMID- 10858092 TI - International Symposium on Gait Disorders. Prague, Czech Republic. September 4-6, 1999. Abstracts. PMID- 10858093 TI - Proceedings of the 5th Congress of the European Confederation of Medical Mycology and the 33rd Scientific Meeting of the Deutschsprachige Mykologische Gesellschaft. Dresden, Germany, June 3-6, 1999. Abstracts. Issue dedicated to Professor Edouard Drouhet. PMID- 10858094 TI - Influenza and hospitalizations in children. PMID- 10858095 TI - Distribution of research awards from the National Institutes of Health among medical schools. PMID- 10858096 TI - Distribution of research awards from the National Institutes of Health among medical schools. PMID- 10858097 TI - Rethinking the role of tube feeding in patients with advanced dementia. PMID- 10858098 TI - Rethinking the role of tube feeding in patients with advanced dementia. PMID- 10858099 TI - Rethinking the role of tube feeding in patients with advanced dementia. PMID- 10858100 TI - Rethinking the role of tube feeding in patients with advanced dementia. PMID- 10858101 TI - The problems with punitive damages in lawsuits against managed-care organizations. PMID- 10858102 TI - The problems with punitive damages in lawsuits against managed-care organizations. PMID- 10858103 TI - The problems with punitive damages in lawsuits against managed-care organizations. PMID- 10858104 TI - The problems with punitive damages in lawsuits against managed-care organizations. PMID- 10858105 TI - Antiretroviral therapy in patients with dual infection with human immunodeficiency virus types 1 and 2. PMID- 10858106 TI - Centrioles go for a stroll. PMID- 10858108 TI - [Needle threader for tension bound laminoplasty]. PMID- 10858107 TI - [Third-generation pill]. PMID- 10858109 TI - [Problem of hair-shaving in neurosurgery]. PMID- 10858110 TI - Professional misconduct. PMID- 10858111 TI - Images in medicine: alcoholic cardiomyopathy a possible cause. PMID- 10858112 TI - Medical restrictions to driving: the awareness of patients and doctors. PMID- 10858113 TI - Fish odour syndrome. PMID- 10858114 TI - Drug induced syndrome of inappropriate antidiuretic hormone secretion. PMID- 10858115 TI - [Proton therapy of uveal melanomas. Interview by H. Allouch]. PMID- 10858116 TI - [Memory: a function that ages well!]. PMID- 10858117 TI - [Response to psychiatric problems of adolescents--a survey on the responses to adolescent children at university hospitals and special facilities]. PMID- 10858118 TI - [Inpatient treatment of children and adolescents--current status, problems, and overview]. PMID- 10858119 TI - [Mental health problems at schools]. PMID- 10858120 TI - [The voice of argument in the case of "Polish paranoia"]. PMID- 10858121 TI - [Preventive mental health services program and modernization of psychiatry in the context of new system of health care financing in Poland]. PMID- 10858122 TI - Environmental dangers: asbestos and tuberculosis. PMID- 10858123 TI - On the edge of facts and hypotheses. PMID- 10858124 TI - Hard metal particles and lung disease: coincidence or causality? PMID- 10858125 TI - Biomedical ethics. Penn report, agency heads home in on clinical research. PMID- 10858126 TI - Development. Brain cells reveal surprising versatility. PMID- 10858127 TI - Vaccine development. Radical steps urged to help underserved. PMID- 10858128 TI - Women's health. Reports see progress, problems, in trials. PMID- 10858129 TI - Biomedical research. Patients help track down disease gene. PMID- 10858130 TI - Alternative medicine. Stephen Straus's impossible job. PMID- 10858131 TI - Alternative medicine. Bastions of tradition adapt to alternative medicine. PMID- 10858132 TI - Mouse genetics. Australian 'ranch' gears up to mass-produce mutant mice. PMID- 10858133 TI - Marine census. Grants kick off ambitious count of all ocean life. PMID- 10858134 TI - Nazi research. Reopening the darkest chapter in German science. PMID- 10858135 TI - Ecology. Mount St. Helens, revisited. PMID- 10858136 TI - Annotation of the human genome. PMID- 10858137 TI - Determining the 3D structure of HIV-1 protease. PMID- 10858138 TI - A brief history of polio vaccines. PMID- 10858139 TI - Flower arranging in Arabidopsis. PMID- 10858140 TI - Archaeology. The cradle of agriculture. PMID- 10858141 TI - Doctor migration -- Daniel in the lions' den. PMID- 10858142 TI - Doctor migration -- Daniel in the lion's den. PMID- 10858143 TI - Doctor migration -- Daniel in the lions' den. PMID- 10858144 TI - Doctor migration -- Daniel in the lions' den. PMID- 10858145 TI - [Gastroesophageal reflux in children]. PMID- 10858146 TI - [Business problem: sick newly milked cows]. PMID- 10858147 TI - Introduction. Neoral use in organ transplantation. PMID- 10858148 TI - Missing the meeting: technologic innovation and work demand versus payment initiatives. PMID- 10858149 TI - Efficacy of unilateral versus bilateral temporal artery biopsies for the diagnosis of giant cell arteritis. PMID- 10858150 TI - Vitrectomy for chronic pseudophakic cystoid macular edema. PMID- 10858151 TI - Dystrophia myotonica and succinylcholine. PMID- 10858152 TI - Dr. Elizabeth Helly. PMID- 10858153 TI - Nightmare from the sixties. PMID- 10858154 TI - Management of fibromyalgia. PMID- 10858155 TI - Management of fibromyalgia. PMID- 10858156 TI - Management of fibromyalgia. PMID- 10858157 TI - Management of fibromyalgia. PMID- 10858158 TI - Management of fibromyalgia. PMID- 10858159 TI - Time and medicine. PMID- 10858160 TI - Auscultation through a shirt. PMID- 10858161 TI - Hemolytic uremic syndrome in a patient treated with clopidogrel. PMID- 10858162 TI - Cholangiocarcinoma and AIDS-related sclerosing cholangitis. PMID- 10858163 TI - Isotretinoin in respiratory papillomatosis. PMID- 10858164 TI - The cadaveric option. PMID- 10858165 TI - Interruption of the inferior vena cava in a patient with Hirschsprung disease. PMID- 10858166 TI - Eugenic sterilization and a Nazi analogy. PMID- 10858167 TI - Richard Lower: anatomist and physiologist. PMID- 10858168 TI - Epidemiologic relation between HIV and invasive pneumococcal disease in San Francisco County, California. PMID- 10858169 TI - Epidemiologic relation between HIV and invasive pneumococcal disease in San Francisco County, California. PMID- 10858170 TI - Chronic dizziness among older adults. PMID- 10858171 TI - Cocaine-induced acute cytologic hepatitis in HIV-infected patients with nonactive viral hepatitis. PMID- 10858172 TI - Parvovirus B19-related anemia in an HIV-infected patient: rapid control after production of neutralizing antibodies during highly active antiretroviral therapy. PMID- 10858173 TI - Consequences of blowing the whistle in medical research. PMID- 10858174 TI - Medicine and MBAs. PMID- 10858175 TI - The cost-effectiveness of sildenafil. AB - BACKGROUND: Coverage of sildenafil by health insurance plans is a contentious issue. OBJECTIVE: To evaluate the cost-effectiveness of sildenafil treatment for erectile dysfunction. DESIGN: A Markov decision model to estimate the incremental cost-effectiveness of sildenafil compared with no drug therapy. DATA SOURCES: Values for the efficacy and safety of sildenafil and quality-of-life utilities were obtained from the published medical literature. Base-case values were chosen to bias against sildenafil use. TARGET POPULATION: Men 60 years of age with erectile dysfunction. TIME HORIZON: Lifetime. PERSPECTIVE: Societal and third party payer. INTERVENTION: Sildenafil or no treatment in identical hypothetical cohorts. OUTCOME MEASURES: Cost per quality-adjusted life-year (QALY) gained. RESULTS OF BASE-CASE ANALYSIS: The cost per QALY gained for sildenafil treatment compared with no therapy was $11,290 from the societal perspective and $11,230 from the third-party payer perspective. RESULTS OF SENSITIVITY ANALYSIS: From the societal perspective, the cost per QALY gained associated with sildenafil was less than $50,000 if treatment-related morbidity was less than 0.8% per year, mortality was less than 0.55% per year, treatment was successful in more than 40.2% of patients, or sildenafil cost less than $244 per month. The results were sensitive to variation of erectile dysfunction utilities, but cost per QALY gained was less than $50,000 if successful treatment increased utility values by 0.05 or more on a scale of 0 (death) to 1 (perfect health). CONCLUSIONS: In an analysis biased against use of sildenafil, the cost-effectiveness of sildenafil treatment compared favorably with that of accepted therapies for other medical conditions. PMID- 10858176 TI - Self-study from web-based and printed guideline materials. A randomized, controlled trial among resident physicians. AB - BACKGROUND: On-line physician education is increasing, but its efficacy in comparison with existing self-study methods is unknown. OBJECTIVE: To compare knowledge, learning efficiency, and learner satisfaction produced by self-study of World Wide Web-based and print-based guidelines for care after acute myocardial infarction. DESIGN: Randomized, controlled trial. SETTING: 12 family medicine and internal medicine residency programs at four universities. PARTICIPANTS: 162 residents. INTERVENTIONS: In proctored sessions, participants were randomly assigned to study from printed materials or from SAGE (Self-Study Acceleration with Graphic Evidence), a Web-based tutorial system. Both methods used identical self-assessment questions and answers and guideline text, but SAGE featured hyperlinks to specific guideline passages and graphic evidence animations. MEASUREMENTS: Scores on multiple-choice knowledge tests, score gain per unit of study time, and ratings on a learner satisfaction scale. RESULTS: Immediate post-test scores on a 20-point scale were similar in the SAGE and control groups (median score, 15.0 compared with 14.5; P>0.2), but SAGE users spent less time studying (median, 27.0 compared with 38.5 minutes; P<0.001) and therefore had greater learning efficiency (median score gain, 8.6 compared with 6.7 points per hour; P = 0.04). On a scale of 5 to 20, SAGE users were more satisfied with learning (median rating, 17.0 compared with 15.0; P<0.001). After 4 to 6 months, knowledge had decreased to the same extent in the SAGE and control groups (median score, 12.0 compared with 11.0; P = 0.12). CONCLUSIONS: On-line tutorials may produce greater learning efficiency and satisfaction than print materials do, but one self-study exposure may be insufficient for long-term knowledge retention. Further research is needed to identify instructional features that motivate greater final learning and retention. PMID- 10858177 TI - Cyclophosphamide is associated with pulmonary function and survival benefit in patients with scleroderma and alveolitis. AB - BACKGROUND: Lung inflammation (alveolitis) may cause lung fibrosis in scleroderma. OBJECTIVE: To determine whether cyclophosphamide treatment is associated with retention of lung function and improved survival in scleroderma patients with alveolitis. DESIGN: Retrospective cohort study. SETTING: Johns Hopkins and University of Maryland Scleroderma Center. PATIENTS: 103 patients with scleroderma who had bronchoalveolar lavage or lung biopsy. INTERVENTION: Cyclophosphamide therapy. MEASUREMENTS: 1) Serial measurement of forced vital capacity (FVC) and carbon monoxide diffusing capacity and 2) survival. RESULTS: During a median follow-up of 13 months after bronchoalveolar lavage or biopsy, patients with alveolitis who did not receive cyclophosphamide therapy experienced a decrease in FVC (mean difference, -0.28 L [95% Cl, -0.41 to -0.16 L] and -7.1% of the predicted value [Cl, -10.9% to -4.0%]). Carbon monoxide diffusing capacity also decreased in these patients (mean difference, -3.3 x mmol min(-1) x kPa(-1) [Cl, -4.6 to -2.1 mmol x min(-1) x kPa(-1)] and -9.6% of the predicted value [Cl, -16.7% to -2.4%]). During a median follow-up of 16 months, patients with alveolitis who received cyclophosphamide were more likely to have a good outcome (stabilization or improvement) in FVC (relative risk, 2.5 [Cl, 1.5 to 4.1]) and diffusing capacity (relative risk, 1.5 [Cl, 1.0 to 2.2]). These patients also had improved survival; the median survival rate was 89% (25th, 75th percentiles, 84%, 94%) compared with 71% (25th, 75th percentiles, 55%, 86%) in untreated patients (P = 0.01, log-rank test). CONCLUSIONS: The presence of lung inflammation identifies patients with scleroderma who are more likely to have worsening lung function. Lung function outcomes and survival are improved in patients with alveolitis who receive cyclophosphamide. PMID- 10858178 TI - Percutaneous coronary revascularization in elderly patients: impact on functional status and quality of life. AB - BACKGROUND: Percutaneous coronary intervention (PCI) is frequently performed in elderly patients, but little is known about its impact on overall health and quality of life. OBJECTIVE: To examine changes in health-related quality of life among elderly patients after PCI. DESIGN: Observational study. SETTING: 75 U.S. hospitals. PATIENTS: Participants in two clinical trials of PCI. MEASUREMENTS: Health-related quality of life was assessed by using the Medical Outcomes Study Short Form (SF-36) survey and the Seattle Angina Questionnaire at baseline, 6 months, and 1 year. RESULTS: Serial data on health-related quality of life were available for 295 elderly (> or =70 years) and 1150 nonelderly (<70 years) patients. At 6 months, physical health had improved in 51% of elderly patients and mental health had improved in 29%. Cardiovascular-specific health status had improved in 58% to 75% of elderly patients. Improvement did not significantly differ between elderly and non-elderly patients at 6 months or 1 year. CONCLUSIONS: Elderly patients selected for participation in a trial of PCI had substantial improvements in health-related quality of life after PCI that were similar to those in younger patients. PMID- 10858179 TI - Effect of hepatitis G virus infection on progression of HIV infection in patients with hemophilia. Multicenter Hemophilia Cohort Study. AB - BACKGROUND: Infection with hepatitis G virus (HGV), also known as GB virus C, is prevalent but is not known to be associated with any chronic disease. Infection with HGV may affect the risk for AIDS in HIV-infected persons. OBJECTIVE: To compare AIDS-free survival in patients with and those without HGV infection during 16 years of follow-up after HIV seroconversion. DESIGN: Subanalysis of a prospective cohort study. SETTING: Comprehensive hemophilia treatment centers in the United States and Europe. PATIENTS: 131 patients with hemophilia who became HIV-positive between 1978 and 1985. MEASUREMENTS: Age, CCR5 genotype, HIV and HCV viral loads, CD4+ and CD8+ lymphocyte counts, and 12-year AIDS-free survival by HGV positivity (viremia [RNA] or anti-E2 antibodies). RESULTS: Compared with HGV negative patients, the 60 HGV-positive patients (46%), including 22 who were positive for HGV RNA, had higher CD4+ lymphocyte counts (difference, 211 cells/mm3 [95% Cl, 88 to 333 cells/mm3]) and 12-year AIDS-free survival rates (68% compared with 40%; rate difference, 1.9 per 100 person-years [Cl, -0.3 to 4.2 per 100 person-years]), despite similar ages and HIV viral loads. In multivariate proportional hazards models, risk for AIDS was 40% lower for HGV positive patients independent of age, HIV and HCV viral loads, CD4+ and CD8+ lymphocyte counts, and CCR5 genotype. CONCLUSIONS: Patients with past or current HGV infection have higher CD4+ lymphocyte counts and better AIDS-free survival rates. The mechanism of this association is unknown. PMID- 10858180 TI - The quality of reporting in published cost-utility analyses, 1976-1997. AB - PURPOSE: Cost-utility analysis is a type of cost-effectiveness analysis in which health effects are measured in terms of quality-adjusted life-years (QALYs) gained. Such analyses have become popular for examining the health and economic consequences of health and medical interventions, and they have been recommended by leaders in the field. These recommendations emphasize the importance of good reporting practices. This study determined 1) the quality of reporting in published cost-utility analyses through 1997 and 2) whether reporting practices have improved over time. We examined quality of reporting by journal type and number of cost-utility analyses a journal has published. DATA SOURCES: Computerized databases were searched through 1997 for the Medical Subject Headings or text keywords quality-adjusted, QALY, and cost-utility analysis. Published bibliographies of the field were also searched. STUDY SELECTION: Original cost-utility analyses written in English were included. Cost effectiveness analyses that measured health effects in units other than QALYs and review, editorial, or methodologic articles were excluded. DATA EXTRACTION: Each of the 228 articles found was audited independently by two trained readers who used a standard data collection form to determine the quality of reporting in several categories: disclosure of funding, framing, reporting of costs, reporting of preference weights, reporting of results, and discussion. RESULTS: The number of cost-utility analyses in the medical literature increased greatly between 1976 and 1997. Analyses covered a wide range of diseases and interventions. Most studies listed modeling assumptions (82%), described the comparator intervention (83%), reported sensitivity analysis (89%), and noted study limitations (84%). Only 52% clearly stated the study perspective; 34% did not disclose the funding source. Methods of reporting costs and preference weights varied widely. The quality of published analyses improved slightly over time and was higher in general clinical journals and in journals that published more of these analyses. CONCLUSIONS: The study results reveal an active and evolving field but also underscore the need for more consistency and clarity in reporting. Better peer review and independent, third-party audits may help in this regard. Future investigations should examine the quality of clinical and economic assumptions used in cost-utility analyses, in addition to whether analysts followed recommended protocols for performance and reporting. PMID- 10858181 TI - Evaluation and management of infertility in women: the internists' role. AB - Interventions for infertility have greatly increased in number and sophistication. Women with multiple medical problems and women near or beyond menopause are now able to conceive. The internist will be called on to assess the risk that infertility interventions pose and to counsel patients accordingly. Knowledge of the medical illnesses associated with infertility, the types of infertility treatments available, and the medical complications of these interventions are required to properly assess this risk. Medical complications of infertility interventions can be direct effects of related drugs and technologies and indirect consequences of the induced pregnancy, multiple gestation, or associated medical conditions. This article reviews the definitions and scope of infertility, the interventions used for treatment of infertility, the medical complications of these interventions, the potential risks of fertility treatment in women unable to conceive spontaneously, and important issues for preconception counseling. PMID- 10858182 TI - A multiyear prospective study of the risk factors for and incidence of diarrheal illness in a cohort of Peace Corps volunteers in Guatemala. AB - BACKGROUND: Diarrheal illness is the most common medical disorder among travelers from developed to developing countries and is common among expatriate residents in developing countries. OBJECTIVE: To assess the risk factors for and incidence of diarrheal illness among Americans living in a developing country. DESIGN: Prospective longitudinal study. SETTING: Rural Guatemala. PATIENTS: Cohort of 36 Peace Corps volunteers. MEASUREMENTS: Collection of daily dietary and symptom data for more than 2 years; identification by multivariate Poisson regression analyses of risk factors for clinically defined episodes of diarrheal illness. RESULTS: The 36 Peace Corps volunteers in this study had 307 diarrheal episodes (median, 7 per person), which lasted a median of 4 days (range, 1 to 112) and a total of 10.1% of the 23 689 person-days in the study. The incidence density (episodes per person-year) was 4.7 for the study as a whole, 6.1 for the first 6 month period, 5.2 for the second 6-month period, and 3.6 thereafter. Statistically significant risk factors for diarrheal illness included drinking water whose source (for example, the tap) and, therefore, quality, was unknown to the person; eating food prepared by a Guatemalan friend or family; eating food at a small, working-class restaurant; eating fruit peeled by someone other than a Peace Corps volunteer; drinking an iced beverage; and eating ice cream, ice milk, or flavored ices. The relative risks comparing the presence of these exposures during the first 6-month period overseas with their absence during the second year of residence ranged from 1.90 to 2.67, and the summary attributable risk percentage (that is, the percentage of diarrheal episodes that could be ascribed to the exposures) was 75.4%. Exposures generally were riskier if they occurred during travel elsewhere in Guatemala rather than in the person's usual work area. CONCLUSIONS: Diarrheal illness of mild-to-moderate severity continued to occur throughout Peace Corps service but decreased in incidence as length of stay increased. Various dietary behaviors increased the risk for diarrheal illness, which suggests that avoidance of potentially risky foods and beverages is beneficial. PMID- 10858183 TI - The importance of diagnosing the polycystic ovary syndrome. AB - The polycystic ovary syndrome (PCOS) is an extremely common disorder that occurs in 4% to 7% of women of reproductive age. Although PCOS is known to be associated with reproductive morbidity and increased risk for endometrial cancer, diagnosis is especially important because PCOS is now thought to increase metabolic and cardiovascular risks. These risks are strongly linked to insulin resistance and are compounded by the common occurrence of obesity, although insulin resistance and its associated risks are also present in nonobese women with PCOS. Women with PCOS are at increased risk for impaired glucose tolerance, type 2 diabetes mellitus, and hypertension. Cardiovascular disease is believed to be more prevalent in women with PCOS, and it has been estimated that such women also have a significantly increased risk for myocardial infarction. Many lipid abnormalities (most notably low high-density lipoprotein cholesterol levels and elevated triglyceride levels) and impaired fibrinolysis are seen in women with PCOS. Early diagnosis of the syndrome and close long-term follow-up and screening for diabetes and cardiovascular disease are warranted. An opportunity exists for preventive therapy, which should improve the reproductive, metabolic, and cardiovascular risks. PMID- 10858184 TI - Tough choices: the cost-effectiveness of sildenafil. PMID- 10858185 TI - The informationist: a new health profession? PMID- 10858186 TI - Functional analysis of genes in the rfb locus of Leptospira borgpetersenii serovar Hardjo subtype Hardjobovis. AB - Lipopolysaccharide (LPS) is a key antigen in immunity to leptospirosis. Its biosynthesis requires enzymes for the biosynthesis and polymerization of nucleotide sugars and the transport through and attachment to the bacterial membrane. The genes encoding these functions are commonly clustered into loci; for Leptospira borgpetersenii serovar Hardjo subtype Hardjobovis, this locus, named rfb, spans 36.7 kb and contains 31 open reading frames, of which 28 have been assigned putative functions on the basis of sequence similarity. Characterization of the function of these genes is hindered by the fact that it is not possible to construct isogenic mutant strains in Leptospira. We used two approaches to circumvent this problem. The first was to clone the entire locus into a heterologous host system and determine if a "recombinant" LPS or polysaccharide was synthesized in the new host. The second approach used putative functions to identify mutants in other bacterial species whose mutations might be complemented by genes on the leptospiral rfb locus. This approach was used to investigate the function of three genes in the leptospiral rfb locus and demonstrated function for orfH10, which complemented a wbpM strain of Pseudomonas aeruginosa, and orfH13, which complemented an rfbW strain of Vibrio cholerae. However, despite the similarity of OrfH11 to WecC, a wecC strain of E. coli was not complemented by orfH11. The predicted protein encoded by orfH8 is similar to GalE from a number of organisms. A Salmonella enterica serovar Typhimurium strain producing no GalE was used as a background in which orfH8 produced detectable GalE enzyme activity. PMID- 10858187 TI - Identification of three essential regulatory gene loci governing expression of Staphylococcus epidermidis polysaccharide intercellular adhesin and biofilm formation. AB - The formation of adherent multilayered biofilms embedded into a glycocalyx represents an essential factor in the pathogenesis of Staphylococcus epidermidis biomaterial-related infections. Using biofilm-producing S. epidermidis 1457 and transposon Tn917 carried on plasmid pTV1ts, we isolated nine isogenic biofilm negative transposon mutants. Transduction by S. epidermidis phage 71 was used to prove the genetic linkage of transposon insertions and altered phenotypes. Mapping of the different transposon insertions by Southern hybridization and pulsed-field gel electrophoresis indicated that these were inserted in four unlinked genetic loci. According to their phenotypes, including quantitative differences in biofilm production in different growth media, in the amount of the polysaccharide intercellular adhesin (PIA) produced, in the hemagglutination titers, and in the altered colony morphology, the mutants could be separated into four phenotypic classes corresponding with the genetic classes. Synthesis of PIA was not detectable with class I and II mutants, whereas the amount of PIA produced reflected the residual degree of biofilm production of class III and IV mutants in different growth media. Chromosomal DNA flanking the transposon insertions of five class I mutants was cloned and sequenced, and the insertions were mapped to different locations of icaADBC, representing the synthetic genes for PIA. Expression of icaADBC from a xylose-dependent promoter in the different isogenic mutant classes reconstituted biofilm production in all mutants. In a Northern blot analysis no icaADBC-specific transcripts were observed in RNA isolated from mutants of classes II, III, and IV. Apparently, in addition to icaADBC, three other gene loci have a direct or indirect regulatory influence on expression of the synthetic genes for PIA on the level of transcription. PMID- 10858188 TI - Ruminant gastrointestinal cell proliferation and clearance of Escherichia coli O157:H7. AB - Human infections with Escherichia coli O157:H7 cause hemorrhagic colitis that can progress to a life-threatening sequelae. The most common mode of disease transmission is ingestion of contaminated bovine food products, and it is well established that E. coli O157:H7 is a transient member of the bovine microbiota. However, the conditions that induce acquisition and subsequent clearance of this bacterium from the ruminant gastrointestinal tract (GIT) are not understood. Evidence that the rates of epithelial cell proliferation in the lower GIT of cattle are associated with the duration animals remained E. coli O157:H7 culture positive is presented. Cattle with slower rates of intestinal cell proliferation in the cecum and the distal colon were culture positive significantly longer than cohort cattle with faster cell proliferation rates. Cell death rates (apoptotic indices) between the short- and long-term culture-positive animals were not different. Typical grain-based finishing diets and forage-based growing diets did not effect GIT cell proliferation or the duration animals remained E. coli O157:H7 culture positive. To identify a dietary intervention that would effect GIT cell proliferation, we used sheep as a model ruminant. A fasting-refeeding regime that increased the rate of GIT cell proliferation was developed. The fasting-refeeding protocol was used in cattle to test the hypothesis that feeding interventions that increase the rate of GIT cell proliferation induce the clearance of E. coli O157:H7 from the bovine GIT. PMID- 10858189 TI - Coxiella burnetii exhibits morphological change and delays phagolysosomal fusion after internalization by J774A.1 cells. AB - Coxiella burnetii, the etiological agent of Q fever, is an obligate intracellular bacterium proliferating within the harsh environment of the phagolysosome. Mechanisms controlling trafficking to, and survival of pathogens within, the phagolysosome are unknown. Two distinct morphological variants have been implicated as playing a role in C. burnetii survival. The dormant small-cell variant (SCV) is resistant to extracellular stresses and the more metabolically active large-cell variant (LCV) is sensitive to environmental stresses. To document changes in the ratio of SCVs to LCVs in response to environment, a protein specific to SCV, ScvA, was quantitated. During the first 2 h after internalization of C. burnetii by J774A.1 cells, the level of ScvA decreased, indicating a change from a population containing primarily SCVs to one containing primarily LCVs. In vitro experiments showed that 2 h of incubation at pH 5.5 caused a significant decrease in ScvA in contrast to incubation at pH 4.5. Measuring in vitro internalization of [(35)S]methionine-[(35)S]cysteine in response to pH, we found the uptake to be optimal at pH 5.5. To explore the possibility that after uptake C. burnetii was able to delay phagolysosomal fusion, we used thorium dioxide and acid phosphatase to label phagolysosomes during infection of J774A.1 cells. We determined that viable C. burnetii was able to delay phagolysosomal fusion. This is the first time that a delay in phagolysosomal fusion has been shown to be a part of the infection process of this pathogenic microorganism. PMID- 10858190 TI - Delayed mortality and attenuated thrombocytopenia associated with severe malaria in urokinase- and urokinase receptor-deficient mice. AB - We explored the role of urokinase and tissue-type plasminogen activators (uPA and tPA), as well as the uPA receptor (uPAR; CD87) in mouse severe malaria (SM), using genetically deficient (-/-) mice. The mortality resulting from Plasmodium berghei ANKA infection was delayed in uPA(-/-) and uPAR(-/-) mice but was similar to that of the wild type (+/+) in tPA(-/-) mice. Parasitemia levels were similar in uPA(-/-), uPAR(-/-), and +/+ mice. Production of tumor necrosis factor, as judged from the plasma level and the mRNA levels in brain and lung, was markedly increased by infection in both +/+ and uPAR(-/-) mice. Breakdown of the blood brain barrier, as evidenced by the leakage of Evans Blue, was similar in +/+ and uPAR(-/-) mice. SM was associated with a profound thrombocytopenia, which was attenuated in uPA(-/-) and uPAR(-/-) mice. Administration of aprotinin, a plasmin antagonist, also delayed mortality and attenuated thrombocytopenia. Platelet trapping in cerebral venules or alveolar capillaries was evident in +/+ mice but absent in uPAR(-/-) mice. In contrast, macrophage sequestration in cerebral venules or alveolar capillaries was evident in both +/+ and uPAR(-/-) mice. Polymorphonuclear leukocyte sequestration in alveolar capillaries was similar in +/+ and uPAR(-/-) mice. These results demonstrate that the uPAR deficiency attenuates the severity of SM, probably by its important role in platelet kinetics and trapping. These results therefore suggest that platelet sequestration contributes to the pathogenesis of SM. PMID- 10858191 TI - Correlations between antibody immune responses at different mucosal effector sites are controlled by antigen type and dosage. AB - Monitoring specific secretory immunoglobulin A (IgA) responses in the intestines after mucosal immunization or infection is impeded by the fact that sampling of small intestinal secretions requires invasive methods not feasible for routine diagnostics. Since IgA plasma cells generated after intragastric immunization are known to populate remote mucosal sites as well, secretory IgA responses at other mucosal surfaces may correlate to those in the intestines and could serve as proxy measures for IgA secretion in the gut. To evaluate the practicability of this approach, mice were immunized intragastrically with 0.2, 2, and 20 mg of ovalbumin plus 10 microg of cholera toxin, and the antigen-specific local secretory IgA responses in duodenal, ileal, jejunal, rectal, and vaginal secretions, saliva, urine, and feces, as well as serum IgG and IgA responses were analyzed by enzyme-linked immunosorbent assay. Correlation analysis revealed significant relationships between serum IgG and IgA, urinary IgA, salivary IgA, and secretory IgA in duodenal, jejunal, ileal, and rectal secretions for the 0.2 mg but not for the 20-mg ovalbumin dose. Fecal samples were poor predictors for intestinal antiovalbumin IgA responses, and no correlations could be established for cholera toxin, neither between local anti-cholera toxin levels nor to the antiovalbumin responses. Thus, specific IgA in serum, saliva, or urine can serve as a predictor of the release of specific IgA at intestinal surfaces after intragastric immunization, but the lack of correlations for high ovalbumin doses and for cholera toxin indicates a strong dependency on antigen type and dosage for these relationships. PMID- 10858192 TI - Measurement of T-cell-derived antigen binding molecules and immunoglobulin G specific to Candida albicans mannan in sera of patients with recurrent vulvovaginal candidiasis. AB - Immunoglobulin G (IgG) and T-cell-derived antigen binding molecules (TABM) specific to whole Candida extract and to Candida-derived mannans prepared by both the cetryltrimethylammonium bromide (CTAB) and alkaline degradation (PEAT) methods were measured in the sera of women with vulvovaginal candidiasis and controls. In the patients there were significantly higher levels of IgG to both CTAB and PEAT mannans and of TABM to CTAB mannan. TABM specific to CTAB mannan was purified from the serum of a patient with a high titer of this TABM. The purified TABM bound specifically to CTAB mannan and to other yeast and mold extracts. This TABM preparation was associated with transforming growth factor beta2 (TGF-beta2), and on specific binding to mannan there was a marked increase in the level of detectable TGF-beta2. This increase in TGF-beta2 level was critically dependent on the relative concentrations of the purified TABM and mannan, being smallest when either was in excess. The TABM specific to CTAB mannan was also shown to inhibit Candida-stimulated gamma interferon production. The results suggest that CTAB mannan-specific TABM may increase susceptibility to vulvovaginal candidiasis in association with a shift in the immune response to the Th2 type. PMID- 10858193 TI - Clostridium perfringens iota-toxin requires activation of both binding and enzymatic components for cytopathic activity. AB - Iota-toxin is produced by Clostridium perfringens type E strains and consists of two independent components, the enzymatic and binding components, referred to as Ia and Ib, respectively. A recombinant C. perfringens strain, strain 667/pMRP147, produced processed Ia and partially processed Ib, while a recombinant C. perfringens type A strain, strain TS133/pMRP147, in which the VirR-VirS two component system is inactivated, produced only precursor forms of Ia and Ib. This suggests that iota-toxin is processed by a VirR-VirS-responsive protease, although not completely in the recombinant type A strain. The precursor forms of Ia and Ib were purified from cultures of the latter strain, and their proteolytic activation was examined. Treatment with proteases cleaved off small peptides (9 to 13 amino acid residues) and a 20-kDa peptide from the N termini of the Ia and Ib precursors, respectively, leading to their active forms. They were activated efficiently by alpha-chymotrypsin, pepsin, proteinase K, subtilisin, and thermolysin but only weakly by trypsin, as demonstrated by the cell-rounding assay. lambda-Protease from the C. perfringens type E strain, which was found to be a zinc-dependent protease related to thermolysin, activated iota-toxin as efficiently as did alpha-chymotrypsin. These results suggest that lambda-protease is most responsible for the activation of iota-toxin in type E strains. PMID- 10858194 TI - Tuberculosis DNA vaccine encoding Ag85A is immunogenic and protective when administered by intramuscular needle injection but not by epidermal gene gun bombardment. AB - Immunogenicity and protective efficacy of a DNA vaccine encoding Ag85A from Mycobacterium tuberculosis were compared in BALB/c and C57BL (B6 and B10) mice immunized by intramuscular (i.m.) needle injection or epidermal gene gun (gg) bombardment. In BALB/c mice, gg immunization could induce elevated antibody and cytotoxic T lymphocyte responses with plasmid doses 50-fold lower than those required for i.m. immunization. Interleukin-2 (IL-2) and gamma interferon (IFN gamma) secretion, however, was much lower in gg-immunized than in i.m.-immunized BALB/c mice. On the other hand, C57BL mice reacted only very weakly to gg immunization, whereas elevated Ag85A-specific antibody, IL-2, and IFN-gamma responses (significantly higher than in BALB/c mice) were detected following vaccination by the i.m. route. Antibody isotypes were indicative of Th2 activation following gg injection of BALB/c and of Th1 activation following i.m. injection of C57BL mice. Finally, C57BL but not BALB/c mice were protected by i.m. Ag85A DNA immunization against intravenous M. tuberculosis challenge, as measured by reduced numbers of CFU in spleen and lungs, compared to animals vaccinated with control DNA. Gene gun immunization was not effective in either BALB/c or C57BL mice. These results indicate that i.m. DNA vaccination is the method of choice for the induction of protective Th1 type immune responses with the Ag85A tuberculosis DNA vaccine. PMID- 10858195 TI - Regulation of Brucella abortus catalase. AB - All aerobic organisms have mechanisms that protect against oxidative compounds. Catalase, peroxidase, superoxide dismutase, glutathione, and thioredoxin are widely distributed in many taxa and constitute elements of a nearly ubiquitous antioxidant metabolic strategy. Interestingly, the regulatory mechanisms that control these elements are rather different depending on the nature of the oxidative stress and the organism. Catalase is well documented to play an important role in protecting cells from oxidative stress. In particular, pathogenic bacteria seem to use this enzyme as a defensive tool against attack by the host. To investigate the significance of catalase in hostile environments, we made catalase deletion mutations in two different B. abortus strains and used two dimensional gel analysis, survival tests, and adaptation experiments to explore the behavior and role of catalase under several oxidative stress conditions. These studies show that B. abortus strains that do not express catalase activity exhibit increased sensitivity to hydrogen peroxide. We also demonstrate that catalase expression is regulated in this species, and that preexposure to a sublethal concentration of hydrogen peroxide allows B. abortus to adapt so as to survive subsequent exposure to higher concentrations of hydrogen peroxide. PMID- 10858196 TI - Induction of a polarized Th1 response by insertion of multiple copies of a viral T-cell epitope into adenylate cyclase of Bordetella pertussis. AB - The adenylate cyclase (CyaA) of Bordetella pertussis delivers the N-terminal catalytic domain into the cytosol of a large number of eukaryotic cells, in particular, professional antigen-presenting cells. This allows the delivery of CD8(+) T-cell epitopes to the major histocompatibility complex class I presentation pathway. We have previously shown that immunization of mice with CyaA carrying a single CD8(+) T-cell epitope leads to antiviral protection as well as to protective and therapeutic antitumor immunity associated with the induction of specific cytotoxic T-lymphocyte (CTL) responses. Here, we evaluated the capacity of CyaA carrying one to four copies of the CD8(+) CD4(+) T-cell epitope from the nucleoprotein of the lymphocytic choriomeningitis virus to induce T-cell responses. Both CTL and Th1-like specific responses were detected in mice immunized with recombinant CyaA with or without adjuvant. Although the insertion of the larger peptides resulted in partial loss of the invasive capacity of recombinant CyaA, insertion of several copies of the same epitope led to a strong enhancement of Th1 responses and, to a lesser degree, CTL responses. These results underscore the potency of CyaA for vaccine design with a new impact on diseases in which the Th1 response has been described to have a beneficial effect. PMID- 10858197 TI - Antigen-specific responses to diphtheria-tetanus-acellular pertussis vaccine in human infants are initially Th2 polarized. AB - Immune responses to exogenous antigens in infant experimental animals display various degrees of Th2 polarization. Preliminary evidence from small human studies suggest a similar age-dependent response pattern to vaccines, but detailed investigations on vaccine immunity during infancy have not yet been undertaken. We report below the results of a comprehensive prospective study on responses to the tetanus component of the diphtheria, tetanus, acellular pertussis (DTaP) vaccine in a cohort of 55 healthy children, employing peripheral blood mononuclear cells (PBMC) collected at the 2-, 4-, and 6-month vaccinations and at 12 months. Antigen-specific production of interleukin-4 (IL-4), IL-5, IL 6, IL-9, IL-10, IL-13, and gamma interferon (IFN-gamma) was determined at each sample point, in parallel with polyclonal (phytohemagglutinin PHA-induced) cytokine responses. Our results indicate early and persistent Th2 responses to the vaccine, in contrast to a more delayed and transient pattern of IFN-gamma production. This initial disparity between the Th1 and Th2 components of the vaccine response was mirrored by patterns of polyclonally induced cytokine production, suggesting that the delayed maturation of the Th1 component of the vaccine response during infancy is secondary to developmental processes occurring within the overall Th cell system. PMID- 10858198 TI - Use of RNA arbitrarily primed-PCR fingerprinting to identify Vibrio cholerae genes differentially expressed in the host following infection. AB - Evidence suggests that a repertoire of Vibrio cholerae genes are differentially expressed in vivo, and regulation of virulence factors in vivo may follow a different pathway. Our work was aimed at characterization of in vivo-grown bacteria and identification of genes that are differentially expressed following infection by RNA arbitrarily primed (RAP)-PCR fingerprinting. The ligated rabbit ileal loop model was used. The motility of in vivo-grown bacteria increased by 350% over that of in vitro-grown bacteria. Also, the in vivo-grown cells were more resistant to killing by human serum. By using the RAP-PCR strategy, five differentially expressed transcripts were identified. Two in vitro-induced transcripts encoded polypeptides for the leucine tRNA synthatase and the 50S ribosomal protein, and the three in vivo-induced transcripts encoded the SucA and MurE proteins and a polypeptide of unknown function. MurE is a protein involved in the peptidoglycan biosynthetic pathway. The lytic profiles of in vivo- and in vitro-grown cells suspended in distilled water were compared; the former was found to be slightly less sensitive to lysis. Ultrathin sections of both cells observed under the transmission electron microscope revealed that in contrast to the usual wavy discontinuous membrane structure of the in vitro-grown cells, in vivo-grown cells had a more rigid, clearly visible double-layered structure. The V. cholerae murE gene was cloned and sequenced. The sequence contained an open reading frame of 1,488 nucleotides with its own ribosome-binding site. A plasmid containing the murE gene of V. cholerae was transformed into V. cholerae 569B, and a transformed strain, 569BME, containing the plasmid was obtained. Ultrathin sections of 569BME viewed under a transmission electron microscope revealed a slightly more rigid cell wall than that of wild-type 569B. When V. cholerae 569B and 569BME cells were injected separately into ligated rabbit ileal loops, the transformed cells had a preference for growth in the ileal loops versus laboratory conditions. PMID- 10858199 TI - The CXC chemokines gamma interferon (IFN-gamma)-inducible protein 10 and monokine induced by IFN-gamma are released during severe melioidosis. AB - Gamma interferon (IFN-gamma)-inducible protein 10 (IP-10) and monokine induced by IFN-gamma (Mig) are related CXC chemokines which bind to the CXCR3 receptor and specifically target activated T lymphocytes and natural killer (NK) cells. The production of IP-10 and Mig by various cell types in vitro is strongly dependent on IFN-gamma. To determine whether IP-10 and Mig are released during bacterial infection in humans, we measured plasma levels of IP-10 and Mig in patients with melioidosis, a severe gram-negative infection caused by Burkholderia pseudomallei. IP-10 and Mig were markedly elevated in patients with melioidosis on admission, particularly in blood culture-positive patients, and remained elevated during the 72-h study period. Levels of IP-10 and Mig showed a positive correlation with IFN-gamma concentrations and also correlated with clinical outcome. In whole blood stimulated with heat-killed B. pseudomallei, neutralization of IFN-gamma and tumor necrosis factor alpha (TNF-alpha) partly attenuated IP-10 and Mig release, while anti-interleukin-12 (IL-12) and anti-IL 18 had a synergistic effect. Stimulation with other bacteria or endotoxin also induced strong secretion of IP-10 and Mig. These data suggest that IP-10 and Mig are part of the innate immune response to bacterial infection. IP-10 and Mig may contribute to host defense in Th1-mediated host defense during infections by attracting CXCR3(+) Th1 cells to the site of inflammation. PMID- 10858200 TI - The lipopolysaccharide structures of Salmonella enterica serovar Typhimurium and Neisseria gonorrhoeae determine the attachment of human mannose-binding lectin to intact organisms. AB - Mannose-binding lectin (MBL) is an important component of the innate immune system. It binds to the arrays of sugars commonly presented by microorganisms and activates the complement system independently of antibody. Despite detailed knowledge of the stereochemical basis of MBL binding, relatively little is known about how bacterial surface structures influence binding of the lectin. Using flow cytometry, we have measured the binding of MBL to a range of mutants of Salmonella enterica serovar Typhimurium and Neisseria gonorrhoeae which differ in the structure of expressed lipopolysaccharide (LPS). For both organisms, the possession of core LPS structures led to avid binding of MBL, which was abrogated by the addition of O antigen (Salmonella serovar Typhimurium) or sialic acid (N. gonorrhoeae). Truncation of the LPS within the core led to lower levels of MBL binding. It was not possible to predict the magnitude of MBL binding from the identity of the LPS terminal sugar alone, indicating that the three-dimensional disposition of LPS molecules is probably also of importance in determining MBL attachment. These results further support the hypothesis that LPS structure is a major determinant of MBL binding. PMID- 10858201 TI - The relapsing fever spirochete Borrelia hermsii contains multiple, antigen encoding circular plasmids that are homologous to the cp32 plasmids of Lyme disease spirochetes. AB - Borrelia hermsii, an agent of tick-borne relapsing fever, was found to contain multiple circular plasmids approximately 30 kb in size. Sequencing of a DNA library constructed from circular plasmid fragments enabled assembly of a composite DNA sequence that is homologous to the cp32 plasmid family of the Lyme disease spirochete, B. burgdorferi. Analysis of another relapsing fever bacterium, B. parkeri, indicated that it contains linear homologs of the B. hermsii and B. burgdorferi cp32 plasmids. The B. hermsii cp32 plasmids encode homologs of the B. burgdorferi Mlp and Bdr antigenic proteins and BlyA/BlyB putative hemolysins, but homologs of B. burgdorferi erp genes were absent. Immunoblot analyses demonstrated that relapsing fever patients produced antibodies to Mlp proteins, indicating that those proteins are synthesized by the spirochetes during human infection. Conservation of cp32-encoded genes in different Borrelia species suggests that their protein products serve functions essential to both relapsing fever and Lyme disease spirochetes. Relapsing fever borreliae replicate to high levels in the blood of infected animals, permitting direct detection and possible functional studies of Mlp, Bdr, BlyA/BlyB, and other cp32-encoded proteins in vivo. PMID- 10858202 TI - Low interleukin-12 activity in severe Plasmodium falciparum malaria. AB - We compared interleukin-12 (IL-12) and other cytokine activities during and after an acute clinical episode in a matched-pair case-control study of young African children who presented with either mild or severe Plasmodium falciparum malaria. The acute-phase, pretreatment plasma IL-12 and alpha interferon (IFN-alpha) levels, as well as the acute-phase mitogen-stimulated whole-blood production capacity of IL-12, were significantly lower in children with severe rather than mild malaria. IL-12 levels, in addition, showed strong inverse correlations both with parasitemia and with the numbers of circulating malaria pigment-containing neutrophils. Acute-phase plasma tumor necrosis factor (TNF) and IL-10 levels were significantly higher in those with severe malaria, and the concentrations of both of these cytokines were positively correlated both with parasitemia and with the numbers of pigment-containing phagocytes in the blood. Children with severe anemia had the highest levels of TNF in plasma. In all the children, the levels in plasma and production capacities of all cytokines normalized when they were healthy and parasite free. The results indicate that severe but not mild P. falciparum malaria in young, nonimmune African children is characterized by down regulated IL-12 activity, contrasting markedly with the up-regulation of both TNF and IL-10 in the same children. A combination of disturbed phagocyte functions resulting from hemozoin consumption, along with reduced IFN-gamma responses, may contribute to these differential effects. PMID- 10858203 TI - Inactivation of Pasteurella (Mannheimia) haemolytica leukotoxin causes partial attenuation of virulence in a calf challenge model. AB - The leukotoxin of Pasteurella (Mannheimia) haemolytica is believed to play a significant role in pathogenesis, causing cell lysis and apoptosis that lead to the lung pathology characteristic of bovine shipping fever. Using a system for Cre-lox recombination, a nonpolar mutation within the lktC transacylase gene of the leukotoxin operon was created. The lktC locus was insertionally inactivated using a loxP-aph3-loxP cassette, and then the aph3 marker was excised from the chromosome by Cre recombinase expressed from a P. haemolytica plasmid. The resulting lktC strain (SH2099) secretes inactive leukotoxin and carries no known antibiotic resistance genes. Strain SH2099 was tested for virulence in a calf challenge model. We inoculated 3 x 10(8) or 3 x 10(9) CFU of wild-type or mutant bacteria into the lungs of healthy, colostrum-deprived calves via transthoracic injection. Animals were observed for clinical signs and for nasal colonization for 4 days, after which they were euthanized and necropsied. The lower inoculum (3 x 10(8) CFU) caused significantly fewer deaths and allowed lung pathology to be scored and compared, while the 3 x 10(9) CFU dose of either the wild-type or mutant was lethal to >/=50% of the calves. The estimated 50% lethal dose of SH2099 was four times higher than that of the wild-type strain. Lung lesion scores were reduced twofold in animals inoculated with the mutant, while clinical scores were nearly equivalent for both strains. The wild-type and mutant strains were equally capable of colonizing the upper respiratory tracts of the calves. In this study, the P. haemolytica lktC mutant was shown to be less virulent than the parent strain. PMID- 10858204 TI - Identification of glycosaminoglycan binding domains in Plasmodium falciparum erythrocyte membrane protein 1 of a chondroitin sulfate A-adherent parasite. AB - Accumulation of Plasmodium falciparum-infected erythrocytes in the placenta is a key feature of maternal malaria. This process is mediated in part by the parasite ligand P. falciparum erythrocyte membrane protein 1 (PfEMP1) at the surface of the infected erythrocyte interacting with the host receptor chondroitin sulfate A (CSA) on the placental lining. We have localized CSA binding activity to two adjacent domains in PfEMP1 of an adherent parasite line and shown the presence of at least three active glycosaminoglycan binding sites. A putative CSA binding sequence was identified in one domain, but nonlinear binding motifs are also likely to be present, since binding activity in the region was shown to be dependent on conformation. Characterization of this binding region provides an opportunity to investigate further its potential as a target for antiadhesion therapy. PMID- 10858205 TI - Complementation of Brucella abortus RB51 with a functional wboA gene results in O antigen synthesis and enhanced vaccine efficacy but no change in rough phenotype and attenuation. AB - Brucella abortus RB51 is a stable rough, attenuated mutant vaccine strain derived from the virulent strain 2308. Recently, we demonstrated that the wboA gene in RB51 is disrupted by an IS711 element (R. Vemulapalli, J. R. McQuiston, G. G. Schurig, N. Srirauganathan, S. M. Halling, and S. M. Boyle, Clin. Diagn. Lab. Immunol. 6:760-764, 1999). Disruption of the wboA gene in smooth, virulent B. abortus, Brucella melitensis, and Brucella suis results in rough, attenuated mutants which fail to produce the O polysaccharide (O antigen). In this study, we explored whether the wboA gene disruption is responsible for the rough phenotype of RB51. We complemented RB51 with a functional wboA gene, and the resulting strain was designated RB51WboA. Colony and Western blot analyses indicated that RB51WboA expressed the O antigen; immunoelectron microscopy revealed that the O antigen was present in the cytoplasm. Crystal violet staining, acryflavin agglutination, and polymyxin B sensitivity studies indicated that RB51WboA had rough phenotypic characteristics similar to those of RB51. Bacterial clearance studies of BALB/c mice indicated no increase in the survival ability of RB51WboA in vivo compared to that of RB51. Vaccination of mice with live RB51WboA induced antibodies to the O antigen which were predominantly of the immunoglobulin G2a (IgG2a) and IgG3 isotypes. After in vitro stimulation of splenocytes with killed bacterial cells, quantitation of gamma interferon in the culture supernatants indicated that RB51WboA immunization induced higher levels of gamma interferon than immunization with RB51. Mice vaccinated with RB51WboA were better protected against a challenge infection with the virulent strain 2308 than those vaccinated with RB51. These studies indicate that in addition to the disruption of the wboA gene there is at least one other mutation in RB51 responsible for its rough phenotype. These studies also suggest that the expressed O antigen in RB51WboA is responsible either directly or indirectly for the observed enhancement in the T cell response. PMID- 10858206 TI - Identification and HLA restriction of naturally derived Th1-cell epitopes from the secreted Mycobacterium tuberculosis antigen 85B recognized by antigen specific human CD4(+) T-cell lines. AB - Antigen 85B (Ag85B/MPT59) is a major secreted protein from Mycobacterium tuberculosis which is a promising candidate antigen for inclusion in novel subunit vaccines against tuberculosis (TB). The present study was undertaken to map naturally derived T-cell epitopes from M. tuberculosis Ag85B in relation to major histocompatibility complex (MHC) class II restriction. Antigen-specific CD4(+) T-cell lines were established from HLA-typed TB patients and Mycobacterium bovis BCG vaccinees by stimulation of peripheral blood mononuclear cells with purified Ag85B in vitro. The established T-cell lines were then tested for proliferation and gamma interferon (IFN-gamma) secretion in response to 31 overlapping synthetic peptides (18-mers) covering the entire sequence of the mature protein. The results showed that the epitopes recognized by T-cell lines from TB patients were scattered throughout the Ag85B sequence whereas the epitopes recognized by T-cell lines from BCG vaccinees were located toward the N terminal part of the antigen. The T-cell epitopes represented by peptides p2 (amino acids [aa] 10 to 27), p3 (aa 19 to 36), and p11 (aa 91 to 108) were frequently recognized by antigen-specific T-cell lines from BCG vaccinees in both proliferation and IFN-gamma assays. MHC restriction analysis demonstrated that individual T-cell lines specifically recognized the complete Ag85B either in association with one of the self HLA-DRB1, DRB3, or DRB4 gene products or nonspecifically in a promiscuous manner. At the epitope level, panel studies showed that peptides p2, p3, and p11 were presented to T cells by HLA-DR-matched as well as mismatched allogeneic antigen-presenting cells, thus representing promiscuous epitopes. The identification of naturally derived peptide epitopes from the M. tuberculosis Ag85B presented to Th1 cells in the context of multiple HLA-DR molecules strongly supports the relevance of this antigen to subunit vaccine design. PMID- 10858207 TI - Molecular cloning and expression of Cu/Zn-containing superoxide dismutase from Fasciola hepatica. AB - The cytosolic superoxide dismutase (SOD) of Fasciola hepatica, a causative agent of fascioliasis, was purified and characterized. The enzyme consists of two identical subunits, each with an apparent molecular mass of 17.5 kDa. An analysis of the enzyme's primary structure and inhibition studies revealed that the enzyme is a copper/zinc-containing SOD (Cu/Zn-SOD). The enzyme activity was relatively stable in a broad pH range, from pH 7.0 to 10.0, and the enzyme showed maximum activity at pH 7.5. This enzyme also displayed strong antigenicity against sera of bovine and human subjects with fascioliasis. The SOD gene fragment was amplified by PCR with degenerate oligonucleotide primers derived from amino acid sequences conserved in the Cu/Zn-SODs of other organisms. An F. hepatica cDNA library was screened with the SOD gene fragment as a probe. As a result, a complete gene encoding the Cu/Zn-SOD was identified, and its nucleotide sequence was determined. The gene had an open reading frame of 438 bp and 146 deduced amino acids. Comparison of the deduced amino acid sequence of the enzyme with previously reported Cu/Zn-SOD amino acid sequences revealed considerably high homologies. The coding region of the F. hepatica Cu/Zn-SOD was cloned and expressed in Escherichia coli. Staining of native polyacrylamide gel for SOD activity of the expressed protein revealed SOD activity that was inactivated by potassium cyanide and hydrogen peroxide but not by sodium azide. This means that the presence of the recombinant fusion protein is indicative of Cu/Zn-SOD. The expressed protein also reacted with sera of bovine and human subjects with fascioliasis, but it did not react with sera of uninfected bovine and human subjects. PMID- 10858208 TI - Selection of recombinant antibodies specific for pathogenic Streptococcus suis by subtractive phage display. AB - A semisynthetic antibody phage display library was used to select recombinant antibodies directed against surface components of a pathogenic strain of Streptococcus suis serotype 2 and against extracellular factor (EF), a protein known to be exclusively associated with pathogenic S. suis serotype 2 strains. Three distinct monoclonal phage antibodies directed against conformational epitopes of surface protein components of S. suis were selected. In addition, three different monoclonal phage antibodies were isolated that recognized EF. To isolate antibody fragments that recognize epitopes specific for a pathogenic S. suis serotype 2 strain, compared to a nonpathogenic serotype 2 strain, we applied a subtractive selection procedure. With this procedure, only one distinct phage antibody was found, and it was shown to be directed against EF. This demonstrates the selectivity of the applied procedure and confirms that EF is indeed differentially expressed by pathogenic and nonpathogenic strains. It also shows that EF is a very dominant antigen in phage antibody selections. PMID- 10858209 TI - Molecular characterization of the Mycoplasma gallisepticum pvpA gene which encodes a putative variable cytadhesin protein. AB - A putative cytadhesin-related protein (PvpA) undergoing variation in its expression was identified in the avian pathogen Mycoplasma gallisepticum. The pvpA gene was cloned, expressed in Escherichia coli, and sequenced. It exhibits 54 and 52% homology with the P30 and P32 cytadhesin proteins of the human pathogens Mycoplasma pneumoniae and Mycoplasma genitalium, respectively. In addition, 50% homology was found with the MGC2 cytadhesin of M. gallisepticum and 49% homology was found with a stretch of 205 amino acids of the cytadherence accessory protein HMW3 of M. pneumoniae. The PvpA molecule possesses a proline rich carboxy-terminal region (28%) containing two identical directly repeated sequences of 52 amino acids and a tetrapeptide motif (Pro-Arg-Pro-X) which is repeated 14 times. Genetic analysis of several clonal isolates representing different expression states of the PvpA product ruled out chromosomal rearrangement as the mechanism for PvpA phase variation. The molecular basis of PvpA variation was revealed in a short tract of repeated GAA codons, encoding five successive glutamate resides, located in the N-terminal region and subject to frequent mutation generating an in-frame UAA stop codon. Size variation of the PvpA protein was observed among M. gallisepticum strains, ranging from 48 to 55 kDa and caused by several types of deletions occurring at the PvpA C-terminal end and within the two directly repeated sequences. By immunoelectron microscopy, the PvpA protein was localized on the mycoplasma cell surface, in particular on the terminal tip structure. Collectively, these findings suggest that PvpA is a newly identified variable surface cytadhesin protein of M. gallisepticum. PMID- 10858210 TI - Peptidoglycan and lipoteichoic acid from Staphylococcus aureus induce tumor necrosis factor alpha, interleukin 6 (IL-6), and IL-10 production in both T cells and monocytes in a human whole blood model. AB - We have examined the ability of peptidoglycan (PepG) and lipoteichoic acid (LTA) isolated from Staphylococcus aureus to induce the release of tumor necrosis factor alpha (TNF-alpha), interleukin-6 (IL-6), and IL-10 in whole human blood and identified the cellular origins of these cytokines. Both PepG and LTA induced transient increases in TNF-alpha and IL-10 in plasma, with peak values at 6 and 12 h, respectively. IL-6 values increased throughout the experimental period (24 h). The TNF-alpha, IL-6, and IL-10 release induced by PepG and LTA was dose dependent. Only PepG was a potent inducer of TNF-alpha secretion. After stimulation of whole blood with PepG or LTA, very pure populations of monocytes (CD14 positive), T cells (CD2 positive), B cells (CD19 positive), and granulocytes (CD15 positive) were isolated by immunomagnetic separation and analyzed by reverse transcription-PCR for mRNA transcripts encoding TNF-alpha, IL 6, and IL-10. The TNF-alpha mRNA results were inconclusive. In contrast, PepG induced IL-6 and IL-10 mRNA accumulation in both T cells and monocytes. LTA, as well as lipopolysaccharide, induced IL-6 and IL-10 mRNA production in monocytes and possibly in T cells. Whether granulocytes and B cells produce cytokines in response to bacterial stimuli remains obscure. Blockade of the CD14 receptors with monoclonal antibodies (18D11) had no influence on the PepG-induced release of TNF-alpha but attenuated the LTA-induced release of the same cytokine. In conclusion, our data indicate that circulating T cells and monocytes contribute to cytokine production in sepsis caused by gram-positive bacteria. PMID- 10858211 TI - Identification and subcellular localization of the Legionella pneumophila IcmX protein: a factor essential for establishment of a replicative organelle in eukaryotic host cells. AB - The gram-negative respiratory pathogen Legionella pneumophila infects and grows within mammalian macrophages and protozoan host cells. Upon uptake into macrophages, L. pneumophila establishes a replicative organelle that avoids fusion with endocytic vesicles. There are 24 dot/icm genes on the L. pneumophila chromosome required for biogenesis of this vacuole. Many of the Dot/Icm proteins are predicted to be components of a membrane-bound secretion apparatus similar to type IV conjugal transfer systems. We have been investigating the function of L. pneumophila dot/icm gene products that do not have obvious orthologs in other type IV transfer systems, since these determinants could govern processes unique to phagosome biogenesis. The icmX gene product falls into this category. To understand the role of the IcmX protein in pathogenesis, we have detailed interactions between an L. pneumophila icmX deletion mutant and murine bone marrow-derived macrophages. These data demonstrate that icmX is required for biogenesis of the L. pneumophila replicative organelle. Immunoblot analysis indicates that the icmX gene product is a polypeptide with an estimated molecular mass of 50 kDa. The IcmX protein was localized to the bacterial periplasm, and periplasmic translocation was mediated by an N-terminal sec-dependent leader peptide. A truncated IcmX product was secreted into culture supernatants by wild type L. pneumophila growing extracellularly in liquid media; however, transport of the IcmX protein into eukaryotic host cells was not detected. Proteins similar in molecular weight to IcmX were identified in other Legionella species by immunoblot analysis using a monoclonal antibody specific for L. pneumophila IcmX protein. From these data, we conclude that the IcmX protein is an essential component of the dot/icm secretion apparatus, and that a conserved mechanism of host cell parasitism exists for members of the Legionellaceae family. PMID- 10858212 TI - Prevalence of virulence genes and clonality in Escherichia coli strains that cause bacteremia in cancer patients. AB - Phenotypic analysis of Escherichia coli strains causing bacteremia in cancer patients suggests that they possess specific virulence properties. To investigate this hypothesis, we compared the frequency of the virulence-related genes cnf1, cnf2, papC, hlyC, and iut in 155 E. coli strains isolated from hospitalized cancer patients with epidemiologically unrelated cases of bacteremia to their frequency in 70 E. coli strains isolated from the feces of healthy unrelated volunteers. Of the blood isolates, 24, 37, and 26% were positive for cnf1, papC, and hlyC, respectively, versus only 6, 17, and 6% of the fecal isolates (P < 0.05 in all instances). By contrast, 47% of both isolates carried the iut gene. The patients' clinical characteristics did not significantly influence these frequencies. The presence on various pathogenicity islands (PAIs) of a combination of the cnf1, papC, and hlyC genes on the chromosome was strongly suggested by Southern blotting of pulsed-field gel electrophoresis (PFGE) patterns with specific DNA probes. The phylogenetic relatedness among 60 strains carrying three, two, one, or no virulence genes and 6 ECOR strains included as references was determined by neighbor joining, the unweighted pair-group method with arithmetic mean, and Wagner analysis of the randomly amplified polymorphic DNA (RAPD) patterns generated by 11 primers. Identification of a major cluster including 96.4% of the strains carrying the cnf1, papC, and hlyC genes and ECOR subgroup B2 strains suggested that the virulent E. coli strains causing bacteremia in cancer patients are closely related to ECOR B2 strains. The presence in the E. coli population surveyed of a strong linkage disequilibrium, and especially of a highly significant correlation between PFGE and RAPD genetic distances, confirms that clonal propagation has a major impact on the E. coli population structure. Nevertheless, low bootstrap values in the phylogenetic tree suggested that frequent genetic exchange inhibits the individualization of discrete genetic lineages, which are stable on an evolutionary scale. PMID- 10858213 TI - Inhibitory and bactericidal effects of hydrogen peroxide production by Streptococcus pneumoniae on other inhabitants of the upper respiratory tract. AB - An inverse correlation between colonization of the human nasopharynx by Streptococcus pneumoniae and Haemophilus influenzae, both common upper respiratory pathogens, has been reported. Studies were undertaken to determine if either of these organisms produces substances which inhibit growth of the other. Culture supernatants from S. pneumoniae inhibited growth of H. influenzae, whereas culture supernatants from H. influenzae had no effect on the growth of S. pneumoniae. Moreover, coculture of S. pneumoniae and H. influenzae led to a rapid decrease in viable counts of H. influenzae. The addition of purified catalase prevented killing of H. influenzae in coculture experiments, suggesting that hydrogen peroxide may be responsible for this bactericidal activity. H. influenzae was killed by concentrations of hydrogen peroxide similar to that produced by S. pneumoniae. Hydrogen peroxide is produced by the pneumococcus through the action of pyruvate oxidase (SpxB) under conditions of aerobic growth. Both an spxB mutant and a naturally occurring variant of S. pneumoniae, which is downregulated in SpxB expression, were unable to kill H. influenzae. A catalase reversible inhibitory effect of S. pneumoniae on the growth of the respiratory tract pathogens Moraxella catarrhalis and Neisseria meningitidis was also observed. Elevated hydrogen peroxide production, therefore, may be a means by which S. pneumoniae is able to inhibit a variety of competing organisms in the aerobic environment of the upper respiratory tract. PMID- 10858214 TI - Acquisition of expression of the Pseudomonas aeruginosa ExoU cytotoxin leads to increased bacterial virulence in a murine model of acute pneumonia and systemic spread. AB - Pseudomonas aeruginosa is the nosocomial bacterial pathogen most commonly isolated from the respiratory tract. Animal models of this infection are extremely valuable for studies of virulence and immunity. We thus evaluated the utility of a simple model of acute pneumonia for analyzing P. aeruginosa virulence by characterizing the course of bacterial infection in BALB/c mice following application of bacteria to the nares of anesthetized animals. Bacterial aspiration into the lungs was rapid, and 67 to 100% of the inoculum could be recovered within minutes from the lungs, with 0.1 to 1% of the inoculum found intracellularly shortly after infection. At later time points up to 10% of the bacteria were intracellular, as revealed by gentamicin exclusion assays on single cell suspensions of infected lungs. Expression of exoenzyme U (ExoU) by P. aeruginosa is associated with a cytotoxic effect on epithelial cells in vitro and virulence in animal models. Insertional mutations in the exoU gene confer a noncytotoxic phenotype on mutant strains and decrease virulence for animals. We used the model of acute pneumonia to determine whether introduction of the exoU gene into noncytotoxic strains of P. aeruginosa lacking this gene affected virulence. Seven phenotypically noncytotoxic P. aeruginosa strains were transformed with pUCP19exoUspcU which carries the exoU gene and its associated chaperone. Three of these strains became cytotoxic to cultured epithelial cells in vitro. These strains all secreted ExoU, as confirmed by detection of the ExoU protein with specific antisera. The 50% lethal dose of exoU-expressing strains was significantly lower for all three P. aeruginosa isolates carrying plasmid pUCP19exoUspcU than for the isogenic exoU-negative strains. mRNA specific for ExoU was readily detected in the lungs of animals infected with the transformed P. aeruginosa strains. Introduction of the exoU gene confers a cytotoxic phenotype on some, but not all, otherwise-noncytotoxic P. aeruginosa strains and, for recombinant strains that could express ExoU, there was markedly increased virulence in a murine model of acute pneumonia and systemic spread. PMID- 10858215 TI - Toxoplasma gondii uses sulfated proteoglycans for substrate and host cell attachment. AB - Toxoplasma gondii is an obligate intracellular parasite that actively invades a wide variety of vertebrate cells, although the basis of this pervasive cell recognition is not understood. We demonstrate here that binding to the substratum and to host cells is partially mediated by interaction with sulfated glycosaminoglycans (GAGs). Addition of excess soluble GAGs blocked parasite attachment to serum-coated glass, thereby preventing gliding motility of extracellular parasites. Similarly, excess soluble GAGs decreased the attachment of parasites to human host cells from a variety of lineages, including monocytic, fibroblast, endothelial, epithelial, and macrophage cells. The inhibition of parasite attachment by GAGs was observed with heparin and heparan sulfate and also with chondroitin sulfates, indicating that the ligands for attachment are capable of recognizing a broad range of GAGs. The importance of sulfated proteoglycan recognition was further supported by the demonstration that GAG deficient mutant host cells, and wild-type cells treated enzymatically to remove GAGs, were partially resistant to parasite invasion. Collectively, these studies reveal that sulfated proteoglycans are one determinant used for substrate and cell recognition by Toxoplasma. The widespread distribution of these receptors may contribute to the broad host and tissue ranges of this highly successful intracellular parasite. PMID- 10858216 TI - Identification of ragAB as a temperature-regulated operon of Porphyromonas gingivalis W50 using differential display of randomly primed RNA. AB - Porphyromonas gingivalis is a gram-negative, black-pigmented anaerobe that has been associated with advanced periodontal disease. The genome of P. gingivalis has the potential to produce a number of virulence determinants including proteases, hemagglutinins, hemolysin, invasion-associated proteins, and products of the pathogenicity island ragAB; however, little is known about how their expression is controlled. Periodontal pockets experience a higher temperature during inflammation, and this elevated temperature may influence the pathogenicity of P. gingivalis by changing its patterns of gene expression. In this study, RNA has been isolated from cells of P. gingivalis grown to steady state at temperatures of 37, 39, and 41 degrees C under hemin excess conditions (pH 7.0) in a chemostat. The RNA was subjected to PCR amplification following reverse transcription, using various combinations of randomly selected oligonucleotide primers. Reproducible RNA fingerprints have been obtained; however, differences were demonstrated in the RNA profiles of cells grown at the three temperatures, indicating differences in gene expression. Several PCR fragments were isolated that appeared to represent temperature-regulated genes. The nucleotide sequence of one of these has been identified as part of the ragAB locus, which codes for both a 55-kDa immunodominant antigen (RagB) and a homologue of the family of TonB-linked outer membrane receptors (RagA). These data indicate that expression of ragAB may be modulated in response to changes in temperature and that this may suggest a mechanism of evading the host response in the inflamed periodontal pocket. PMID- 10858217 TI - Natural history of Streptococcus sanguinis in the oral cavity of infants: evidence for a discrete window of infectivity. AB - The heterogeneous group of oral bacteria within the sanguinis (sanguis) streptococci comprise members of the indigenous biota of the human oral cavity. While the association of Streptococcus sanguinis with bacterial endocarditis is well described in the literature, S. sanguinis is thought to play a benign, if not a beneficial, role in the oral cavity. Little is known, however, about the natural history of S. sanguinis and its specific relationship with other oral bacteria. As part of a longitudinal study concerning the transmission and acquisition of oral bacteria within mother-infant pairs, we examined the initial acquisition of S. sanguinis and described its colonization relative to tooth emergence and its proportions in plaque and saliva as a function of other biological events, including subsequent colonization with mutans streptococci. A second cohort of infants was recruited to define the taxonomic affiliation of S. sanguinis. We found that the colonization of the S. sanguinis occurs during a discrete "window of infectivity" at a median age of 9 months in the infants. Its colonization is tooth dependent and correlated to the time of tooth emergence; its proportions in saliva increase as new teeth emerge. In addition, early colonization of S. sanguinis and its elevated levels in the oral cavity were correlated to a significant delay in the colonization of mutans streptococci. Underpinning this apparent antagonism between S. sanguinis and mutans streptococci is the observation that after mutans streptococci colonize the infant, the levels of S. sanguinis decrease. Children who do not harbor detectable levels of mutans streptococci have significantly higher levels of S. sanguinis in their saliva than do children colonized with mutans streptococci. Collectively, these findings suggest that the colonization of S. sanguinis may influence the subsequent colonization of mutans streptococci, and this in turn may suggest several ecological approaches toward controlling dental caries. PMID- 10858218 TI - Expression of cytokine genes during pneumococcal and nontypeable Haemophilus influenzae acute otitis media in the rat. AB - Acute otitis media (AOM) elicits potent inflammatory responses from the cells of the middle ear mucosa as well as from infiltrating leukocytes. To explore host responses during experimental AOM induced by Streptococcus pneumoniae type 3 and nontypeable Haemophilus influenzae (NTHi), otomicroscopy findings and expression of cytokine genes in the middle ear were monitored up to 1 month postinoculation. The mucosa and infiltrating cells responded rapidly to the bacterial challenge. Otomicroscopically, AOM appeared 1 day after NTHi inoculation and 3 days after pneumococcus inoculation. Pneumococcal AOM was more severe than NTHi otitis, but in general, lower transcript levels were detected in pneumococcus-infected than in NTHi-infected animals. Interleukin-6 (IL-6) mRNA levels peaked at 3 to 6 h for both pneumococcus-infected and NTHi-infected animals. IL-1alpha, tumor necrosis factor alpha, and IL-10 mRNA levels peaked at 6 h for NTHi otitis and 1 to 3 days for pneumococcal otitis. Comparing otomicroscopy with expression profiles, it would appear that the majority of cytokine mRNAs had passed their peak before the AOM diagnosis could be made clinically. Only transforming growth factor beta mRNA followed a slower time course, peaking very late and continuing expression even after the AOM was otomicroscopically resolved. IL-2 and IL-4 mRNAs were not detected in any animal at any time. Most of the investigated cytokines are very early markers for AOM and may be involved in initiation of inflammation, but they would be poor targets for pharmacological manipulation since their levels decline before clinical signs appear. PMID- 10858219 TI - Beryllium, an adjuvant that promotes gamma interferon production. AB - Beryllium is associated with a human pulmonary granulomatosis characterized by an accumulation of CD4(+) T cells in the lungs and a heightened specific lymphocyte proliferative response to beryllium (Be) with gamma interferon (IFN-gamma) release (i.e., a T helper 1 [Th1] response). While an animal model of Be sensitization is not currently available, Be has exhibited adjuvant effects in animals. The effects of Be on BALB/c mice immunized with soluble leishmanial antigens (SLA) were investigated to determine if Be had adjuvant activity for IFN gamma production, an indicator of the Th1 response. In this strain of Leishmania susceptible BALB/c mice, a Th2 response is normally observed after in vivo SLA sensitization and in vitro restimulation with SLA. If interleukin-12 (IL-12) is given during in vivo sensitization with SLA, markedly increased IFN-gamma production and decreased IL-4 production are detected. We show here that when beryllium sulfate (BeSO(4)) was added during in vivo sensitization of BALB/c mice with SLA and IL-12, significantly increased IFN-gamma production and decreased IL 4 production from lymph node and spleen cells were detected upon in vitro SLA restimulation. No specific responses were observed to Be alone. Lymph node and spleen cells from all mice proliferated strongly and comparably upon in vitro restimulation with SLA and with SLA plus Be; no differences were noted among groups of mice that received different immunization regimens. In vivo, when Be was added to SLA and IL-12 for sensitization of BALB/c mice, more effective control of Leishmania infection was achieved. This finding has implications for understanding not only the development of granulomatous reactions but also the potential for developing Be as a vaccine adjuvant. PMID- 10858220 TI - Investigation of the role of type IV Aeromonas pilus (Tap) in the pathogenesis of Aeromonas gastrointestinal infection. AB - Although there is substantial evidence that type IV pili purified from diarrhea associated Aeromonas species (designated Bfp for bundle-forming pilus) are intestinal colonization factors (S. M. Kirov, L. A. O'Donovan, and K. Sanderson, Infect. Immun. 67:5447-5454, 1999), nothing is known regarding the function of a second family of Aeromonas type IV pili (designated Tap for type IV Aeromonas pilus), identified following the cloning of a pilus biogenesis gene cluster tapABCD. Related pilus gene clusters are widely conserved among gram-negative bacteria, but their significance for virulence has been controversial. To investigate the role of Tap pili in Aeromonas pathogenesis, mutants of Aeromonas strains (a fish isolate of A. hydrophila and a human dysenteric isolate of A. veronii bv. sobria) were prepared by insertional inactivation of the tapA gene which encodes the type IV pilus subunit protein, TapA. Exotoxic activities were unaffected by the mutation in tapA. Inactivation of tapA had no effect on the bacterial adherence of these two isolates to HEp-2 cells. For the A. veronii bv. sobria isolate, adhesion to Henle 407 intestinal cells and to human intestinal tissue was also unaffected. There was no significant effect on the duration of colonization or incidence of diarrhea when the A. veronii bv. sobria strain was tested in the removable intestinal tie adult rabbit diarrhea model or on its ability to colonize infant mice. Evidence was obtained that demonstrated that TapA was expressed by both Aeromonas species and was present on the cell surface, although if assembled into pili this pilus type appears to be an uncommon one under standard bacterial growth conditions. Further studies into factors which may influence Tap expression are required, but the present study suggests that Tap pili may not be as significant as Bfp pili for Aeromonas intestinal colonization. PMID- 10858221 TI - Importance of holotoxin assembly in Ptl-mediated secretion of pertussis toxin from Bordetella pertussis. AB - We examined the structural components of pertussis toxin that are required for efficient export from Bordetella pertussis via the Ptl system, a member of the type IV family of macromolecular transporters. First, we constructed a strain of B. pertussis that contains a functional Ptl system but does not produce pertussis toxin. Plasmids which express either the S1 subunit or the B oligomer were then introduced into this strain. We found that the B oligomer of the toxin is not secreted in the absence of the S1 subunit. Conversely, the S1 subunit is also not secreted by a Ptl-mediated mechanism in the absence of the B oligomer. Thus, an assembled holotoxin is required for Ptl-mediated export of pertussis toxin from B. pertussis. PMID- 10858222 TI - RgpA-Kgp peptide-based immunogens provide protection against Porphyromonas gingivalis challenge in a murine lesion model. AB - Porphyromonas gingivalis, a gram-negative bacterium, has been linked to the onset and progression of periodontitis, a chronic inflammatory disease of the supporting tissues of the teeth. A major virulence factor of P. gingivalis is an extracellular complex of Arg- and Lys-specific proteinases and adhesins designated the RgpA-Kgp complex (formerly the PrtR-PrtK complex). In this study we show that the RgpA-Kgp complex, when used as an immunogen with incomplete Freund adjuvant (IFA), protects against challenge with invasive and noninvasive strains of P. gingivalis in the murine lesion model. We identified a variety of peptide vaccine candidates from the RgpA and Kgp polyprotein sequences that involved the putative active site histidine of both proteinases and five repeat motifs in the adhesin domains of both polyproteins implicated in aggregation and binding to host substrates, designated adhesin-binding motif (ABM) peptides. These peptides were synthesized using standard, solid-phase protocols for 9 fluorenylmethoxy carbonyl chemistry with S-acetylmercaptoacetic acid (SAMA) as the N-terminal residue. The SAMA-peptides were then conjugated to diphtheria toxoid and used with IFA to immunize BALB/c mice. Both active-site peptides and three of the five ABM peptides gave protection (P < 0.005) against challenge with P. gingivalis in the murine lesion model. The three ABM peptide sequences that conferred protection exist within a 100-residue span in the RgpA44 and Kgp39 adhesins of the RgpA-Kgp complex. Protective anti-RgpA-Kgp complex mouse antisera recognized the RgpA27, Kgp39, and RgpA44 adhesins in an immunoblot. Epitope mapping of the RgpA27 adhesin using the protective anti-RgpA-Kgp antisera identified a major protective epitope that mapped immediately N terminal to one of the protective ABM peptides in the 100-residue span in RgpA44 and Kgp39. This identified protective epitope contains clusters of basic residues spatially surrounded by hydrophobic amino acids, a finding which is characteristic of a heparin binding motif. PMID- 10858223 TI - Full capacity of recombinant Escherichia coli heat-stable enterotoxin fusion proteins for extracellular secretion, antigenicity, disulfide bond formation, and activity. AB - We have successfully used the major subunit ClpG of Escherichia coli CS31A fimbriae as an antigenic and immunogenic exposure-delivery vector for various heterologous peptides varying in nature and length. However, the ability of ClpG as a carrier to maintain in vitro and in vivo the native biological properties of passenger peptide has not yet been reported. To address this possibility, we genetically fused peptides containing all or part of the E. coli human heat stable enterotoxin (STh) sequence to the amino or carboxyl ends of ClpG. Using antibodies to the ClpG and STh portions for detecting the hybrids; AMS (4 acetamido-4'-maleimidylstilbene-2, 2'-disulfonate), a potent free thiol-trapping reagent, for determining the redox state of STh in the fusion; and the suckling mouse assay for enterotoxicity, we demonstrated that all ClpG-STh proteins were secreted in vitro and in vivo outside the E. coli cells in a heat-stable active oxidized (disulfide-bonded) form. Indeed, in contrast to many earlier studies, blocking the natural NH(2) or COOH extremities of STh had, in all cases, no drastic effect on cell release and toxin activity. Only antigenicity of STh C terminally extended with ClpG was strongly affected in a conformation-dependent manner. These results suggest that the STh activity was not altered by the chimeric structure, and therefore that, like the natural toxin, STh in the fusion had a spatial structure flexible enough to be compatible with secretion and enterotoxicity (folding and STh receptor recognition). Our study also indicates that disulfide bonds were essential for enterotoxicity but not for release, that spontaneous oxidation by molecular oxygen occurred in vitro in the medium, and that the E. coli cell-bound toxin activity in vivo resulted from an effective export processing of hybrids and not cell lysis. None of the ClpG-STh subunits formed hybrid CS31A-STh fimbriae at the cell surface of E. coli, and a strong decrease in the toxin activity was observed in the absence of CS31A helper proteins. In fact, chimeras translocated across the outer membrane as a free folded monomer once they were guided into the periplasm by the ClpG leader peptide through the CS31A-dependent secretory pathway. In summary, ClpG appears highly attractive as a carrier reporter protein for basic and applied research through the engineering of E. coli for culture supernatant delivery of an active cysteine-containing protein, such as the heat-stable enterotoxin. PMID- 10858224 TI - Tumor necrosis factor alpha-mediated toxic shock in Trypanosoma cruzi-infected interleukin 10-deficient mice. AB - Using interleukin-10 (IL-10)-deficient (IL-10(-/-)) mice, previous studies revealed a pathological immune response after infection with Trypanosoma cruzi that is associated with CD4(+) T cells and overproduction of proinflammatory cytokines. In this study we further investigate the pathology and potential mediators for the mortality in infected animals. T. cruzi-infected IL-10(-/-) mice showed reduced parasitemia accompanied by increased systemic release of gamma interferon (IFN-gamma), IL-12, and reactive nitrogen intermediates and overproduction of tumor necrosis factor alpha (TNF-alpha). Despite this early resistance, IL-10(-/-) mice died within the third week of infection, whereas all control mice survived acute infection. The clinical manifestation with weight loss, hypothermia, hypoglycemia, hyperkalemia, and increased liver-derived enzymes in the blood together with hepatic necrosis and intravascular coagulation in moribund mice indicated a toxic shock-like syndrome, possibly mediated by the systemic TNF-alpha overproduction. Indeed, high production of systemic TNF-alpha significantly correlated with mortality, and moribund mice died with critically high TNF-alpha concentrations in the blood. Consequent treatment with anti-TNF alpha antiserum attenuated pathological changes in T. cruzi-infected IL-10(-/-) mice and significantly prolonged survival; the mice died during the fourth week postinfection, again with a striking correlation between regaining high systemic TNF-alpha concentrations and the time of death. Since elevated serum IL-12 and IFN-gamma concentrations were not affected by the administration of antiserum, these studies suggest that TNF-alpha is the direct mediator of this toxic shock syndrome. In conclusion, induction of endogenous IL-10 during experimentally induced Chagas' disease seems to be crucial for counterregulating an overshooting proinflammatory cytokine response resulting in TNF-alpha-mediated toxic shock. PMID- 10858225 TI - High intracellular level of guanosine tetraphosphate in Mycobacterium smegmatis changes the morphology of the bacterium. AB - Almost one-third of the world population today harbors the tubercle bacillus asymptomatically. It is postulated that the morphology and staining pattern of the long-term persistors are different from those of actively growing culture. Interestingly, it has been found that the morphology and staining pattern of the starved in vitro population of mycobacteria is similar to the persistors obtained from the lung lesions. In order to delineate the biochemical characteristics of starved mycobacteria, Mycobacteria smegmatis was grown in 0.2% glucose as a sole carbon source along with an enriched culture in 2% glucose. Accumulation of the stringent factor guanosine tetraphosphate (ppGpp) with a concomitant change in morphology was observed for M. smegmatis under carbon-deprived conditions. In addition, M. smegmatis assumed a coccoid morphology when ppGpp was ectopically produced by overexpressing Escherichia coli relA, even in an enriched medium. The Mycobacterium tuberculosis relA and spoT homologue, when induced in M. smegmatis, also resulted in the overproduction of ppGpp with a change in the bacterium's growth characteristics. PMID- 10858226 TI - Detection of phase variation in expression of proteins involved in hemoglobin and hemoglobin-haptoglobin binding by nontypeable Haemophilus influenzae. AB - Haemophilus influenzae can utilize different protein-bound forms of heme for growth in vitro. A previous study (I. Maciver, J. L. Latimer, H. H. Liem, U. Muller-Eberhard, Z. Hrkal, and E. J. Hansen. Infect. Immun. 64:3703-3712, 1996) indicated that nontypeable H. influenzae (NTHI) strain TN106 expressed a protein that bound hemoglobin-haptoglobin and was encoded by an open reading frame (ORF) that contained a CCAA nucleotide repeat. Southern blot analysis revealed that several NTHI strains contained between three and five chromosomal DNA fragments that bound an oligonucleotide probe for CCAA repeats. Three ORFs containing CCAA repeats were identified in NTHI strain N182; two of these ORFs were arranged in tandem. The use of translational fusions involving these three ORFs and the beta lactamase gene from pBR322 revealed that these three ORFs, designated hgbA, hgbB, and hgbC, encoded proteins that could bind hemoglobin, hemoglobin-haptoglobin, or both compounds. Monoclonal antibodies (MAbs) specific for the HgbA, HgbB, and HgbC proteins were produced by immunizing mice with synthetic peptides unique to each protein. Both HgbA and HgbB were readily detected by Western blot analysis in N182 cells grown in the presence of hemoglobin as the sole source of heme, whereas expression of HgbC was found to be much less abundant than that of HgbA and HgbB. The use of these MAbs in a colony blot radioimmunoassay analysis revealed that expression of both HgbA and HgbB was subject to phase variation. PCR and nucleotide sequence analysis were used in conjunction with Western blot analyses to demonstrate that this phase variation involved the CCAA repeats in the hgbA and hgbB ORFs. PMID- 10858227 TI - Identification of genes required for chronic persistence of Brucella abortus in mice. AB - The genetic basis for chronic persistence of Brucella abortus in lymphoid organs of mice, cows, and humans is currently unknown. We identified B. abortus genes involved in chronic infection, by assessing the ability of 178 signature-tagged mutants to establish and maintain persistent infection in mice. Each mutant was screened for its ability to colonize the spleens of mice at 2 and 8 weeks after inoculation. Comparison of the results from both time points identified two groups of mutants attenuated for chronic infection in mice. The first group was not recovered at either 2 or 8 weeks postinfection and was therefore defective in establishing infection. Mutants in this group carried transposon insertions in genes involved in lipopolysaccharide biosynthesis (wbkA), in aromatic amino acid biosynthesis, and in type IV secretion (virB1 and virB10). The second group, which was recovered at wild-type levels 2 weeks postinfection but not 8 weeks postinfection was able to establish infection but was unable to maintain chronic infection. One mutant in this group carried a transposon insertion in a gene with homology to gcvB of Mycobacterium tuberculosis, encoding glycine dehydrogenase, an enzyme whose activity is increased during the state of nonreplicating persistence. These results suggest that some mechanisms for long-term persistence may be shared among chronic intracellular pathogens. Furthermore, identification of two groups of genes, those required for initiating infection and those required only for long-term persistence, suggests that B. abortus uses distinct sets of virulence determinants to establish and maintain chronic infection in mice. PMID- 10858228 TI - Molecular cloning and expression of a gene encoding Cryptosporidium parvum glycoproteins gp40 and gp15. AB - Cryptosporidium parvum is a significant cause of diarrheal disease worldwide. The specific molecules that mediate C. parvum-host cell interactions and the molecular mechanisms involved in the pathogenesis of cryptosporidiosis are unknown. In this study we have shown that gp40, a mucin-like glycoprotein, is localized to the surface and apical region of invasive stages of the parasite and is shed from its surface. gp40-specific antibodies neutralize infection in vitro, and native gp40 binds specifically to host cells, implicating this glycoprotein in C. parvum attachment to and invasion of host cells. We have cloned and sequenced a gene designated Cpgp40/15 that encodes gp40 as well as gp15, an antigenically distinct, surface glycoprotein also implicated in C. parvum-host cell interactions. Analysis of the deduced amino acid sequence of the 981-bp Cpgp40/15 revealed the presence of an N-terminal signal peptide, a polyserine domain, multiple predicted O-glycosylation sites, a single potential N glycosylation site, and a hydrophobic region at the C terminus, a finding consistent with what is required for the addition of a GPI anchor. There is a single copy of Cpgp40/15 in the C. parvum genome, and this gene does not contain introns. Our data indicate that the two Cpgp40/15-encoded proteins, gp40 and gp15, are products of proteolytic cleavage of a 49-kDa precursor protein which is expressed in intracellular stages of the parasite. The surface localization of gp40 and gp15 and their involvement in the host-parasite interaction suggest that either or both of these glycoproteins may serve as effective targets for specific preventive or therapeutic measures for cryptosporidiosis. PMID- 10858229 TI - Cloning and sequence analysis of a highly polymorphic Cryptosporidium parvum gene encoding a 60-kilodalton glycoprotein and characterization of its 15- and 45 kilodalton zoite surface antigen products. AB - The apicomplexan parasite Cryptosporidium parvum is a major cause of serious diarrheal disease in both humans and animals. No efficacious chemo- or immunotherapies have been identified for cryptosporidiosis, but certain antibodies directed against zoite surface antigens and/or proteins shed by gliding zoites have been shown to neutralize infectivity in vitro and/or to passively protect against, or ameliorate, disease in vivo. We previously used monoclonal antibody 11A5 to identify a 15-kDa surface glycoprotein that was shed behind motile sporozoites and was recognized by several lectins that neutralized parasite infectivity for cultured epithelial cells. Here we report the cloning and sequence analysis of the gene encoding this 11A5 antigen. Surprisingly, the gene encoded a 330-amino-acid, mucin-like glycoprotein that was predicted to contain an N-terminal signal peptide, a homopolymeric tract of serine residues, 36 sites of O-linked glycosylation, and a hydrophobic C-terminal peptide specifying attachment of a glycosylphosphatidylinositol anchor. The single-copy gene lacked introns and was expressed during merogony to produce a 60-kDa precursor which was proteolytically cleaved to 15- and 45-kDa glycoprotein products that both localized to the surface of sporozoites and merozoites. The gp15/45/60 gene displayed a very high degree of sequence diversity among C. parvum isolates, and the numerous single-nucleotide and single-amino-acid polymorphisms defined five to six allelic classes, each characterized by additional intra-allelic sequence variation. The gp15/45/60 single-nucleotide polymorphisms will prove useful for haplotyping and fingerprinting isolates and for establishing meaningful relationships between C. parvum genotype and phenotype. PMID- 10858230 TI - Plasmodium chabaudi-infected erythrocytes adhere to CD36 and bind to microvascular endothelial cells in an organ-specific way. AB - Adherence of erythrocytes infected with Plasmodium falciparum to microvascular endothelial cells (sequestration) is considered to play an important role in parasite virulence and pathogenesis. However, the real importance of sequestration for infection and disease has never been fully assessed. The absence of an appropriate in vivo model for sequestration has been a major barrier. We have examined the rodent malaria parasite Plasmodium chabaudi chabaudi AS in mice as a potential model. Erythrocytes infected with this parasite adhere in vitro to purified CD36, a critical endothelium receptor for binding P. falciparum-infected erythrocytes. P. c. chabaudi-infected erythrocytes adhere in vitro to endothelial cells in a gamma interferon-dependent manner, suggesting the involvement of additional adhesion molecules in the binding process, as is also the case with P. falciparum-infected cells. Furthermore, plasma or sera from infected and hyperimmune mice, respectively, have the ability to block binding of infected erythrocytes to endothelial cells. In vivo, erythrocytes containing mature P. c. chabaudi parasites are sequestered from the peripheral circulation. Sequestration is organ specific, occurring primarily in the liver, although intimate contact between infected erythrocytes and endothelial cells is also observed in the spleen and brain. The results are discussed in the context of the use of this model to study (i) the relationship between endothelial cell activation and the level of sequestration and (ii) the primary function of sequestration in malaria infection. PMID- 10858231 TI - Relationship between the Tsh autotransporter and pathogenicity of avian Escherichia coli and localization and analysis of the Tsh genetic region. AB - The temperature-sensitive hemagglutinin Tsh is a member of the autotransporter group of proteins and was first identified in avian-pathogenic Escherichia coli (APEC) strain chi7122. The prevalence of tsh was investigated in 300 E. coli isolates of avian origin and characterized for virulence in a 1-day-old chick lethality test. Results indicate that among the tsh-positive APEC isolates, 90.6% belonged to the highest virulence class. Experimental inoculation of chickens with chi7122 and an isogenic tsh mutant demonstrated that Tsh may contribute to the development of lesions within the air sacs of birds but is not required for subsequent generalized infection manifesting as perihepatitis, pericarditis, and septicemia. Conjugation and hybridization experiments revealed that the tsh gene is located on a ColV-type plasmid in many of the APEC strains studied, including strain chi7122, near the colicin V genes in most of these strains. DNA sequences flanking the tsh gene of strain chi7122 include complete and partial insertion sequences and phage-related DNA sequences, some of which were also found on virulence plasmids and pathogenicity islands present in various E. coli pathotypes and other pathogenic members of the Enterobacteriaceae. These results demonstrate that the tsh gene is frequently located on the ColV virulence plasmid in APEC and suggest a possible role of Tsh in the pathogenicity of E. coli for chickens in the early stages of infection. PMID- 10858232 TI - Comparative genomics of Helicobacter pylori: analysis of the outer membrane protein families. AB - The two complete genomic sequences of Helicobacter pylori J99 and 26695 were used to compare the paralogous families (related genes within one genome, likely to have related function) of genes predicted to encode outer membrane proteins which were present in each strain. We identified five paralogous gene families ranging in size from 3 to 33 members; two of these families contained members specific for either H. pylori J99 or H. pylori 26695. Most orthologous protein pairs (equivalent genes between two genomes, same function) shared considerable identity between the two strains. The unusual set of outer membrane proteins and the specialized outer membrane may be a reflection of the adaptation of H. pylori to the unique gastric environment where it is found. One subfamily of proteins, which contains both channel-forming and adhesin molecules, is extremely highly related at the sequence level and has likely arisen due to ancestral gene duplication. In addition, the largest paralogous family contained two essentially identical pairs of genes in both strains. The presence and genomic organization of these two pairs of duplicated genes were analyzed in a panel of independent H. pylori isolates. While one pair was present in every strain examined, one allele of the other pair appeared partially deleted in several isolates. PMID- 10858233 TI - Lyme arthritis resolution with antiserum to a 37-kilodalton Borrelia burgdorferi protein. AB - A 37-kDa protein from Borrelia burgdorferi (the agent of Lyme disease) was identified as a target for immune-mediated resolution of Lyme arthritis. Studies in a mouse model have shown that arthritis resolution can be mediated by antibodies (against unknown target antigens) within immune sera from actively infected mice. Immune sera from infected mice were therefore used to screen a B. burgdorferi genomic expression library. A gene was identified whose native product is a putative lipoprotein of approximately 37 kDa, referred to here as arthritis-related protein (Arp). Active and passive immunization of mice with recombinant Arp or Arp antiserum, respectively, did not protect mice from challenge inoculation. However, when Arp antiserum was administered to severe combined immunodeficient (SCID) mice with established infections and with ongoing arthritis and carditis, treatment selectively induced arthritis resolution without affecting the status of carditis or influencing the status of infection, including spirochetemia. The selective arthritis-resolving effect of Arp antiserum mimics the activity of immune serum from immunocompetent mice when such serum is transferred into SCID mice with established infections. The arp gene could not be amplified from unrelated B. burgdorferi isolates but hybridized with those isolates only under very-low-stringency conditions. Arp antiserum reacted against proteins of similar size in a wide range of B. burgdorferi isolates. PMID- 10858234 TI - The abundant larval transcript-1 and -2 genes of Brugia malayi encode stage specific candidate vaccine antigens for filariasis. AB - Lymphatic filariasis is a major tropical disease caused by the mosquito-borne nematodes Brugia and Wuchereria. About 120 million people are infected and at risk of lymphatic pathology such as acute lymphangitis and elephantiasis. Vaccines against filariasis must generate immunity to the infective mosquito derived third-stage larva (L3) without accentuating immunopathogenic responses to lymphatic-dwelling adult parasites. We have identified two highly expressed genes, designated abundant larval transcript-1 and -2 (alt-1 and alt-2), from each of which mRNAs account for >1% of L3 cDNAs. ALT-1 and ALT-2 share 79% amino acid identity across 125 residues, including a putative signal sequence and a prominent acidic tract. Expression of alt-1 and alt-2 is initiated midway through development in the mosquito, peaking in the infective larva and declining sharply following entry into the host. Humans exposed to Brugia malayi show a high frequency of immunoglobulin G1 (IgG1) and IgG3 antibodies to ALT-1 and -2, distinguishing them from adult-stage antigens, which are targeted by the IgG4 isotype. Immunization of susceptible rodents (jirds) with ALT-1 elicited a 76% reduction in parasite survival, the highest reported for a single antigen from any filarial parasite. ALT-1 and the closely related ALT-2 are therefore strong candidates for a future vaccine against human filariasis. PMID- 10858235 TI - The putative proteinase maturation protein A of Streptococcus pneumoniae is a conserved surface protein with potential to elicit protective immune responses. AB - Surface-exposed proteins often play an important role in the interaction between pathogenic bacteria and their host. We isolated a pool of hydrophobic, surface associated proteins of Streptococcus pneumoniae. The opsonophagocytic activity of hyperimmune serum raised against this protein fraction was high and species specific. Moreover, the opsonophagocytic activity was independent of the capsular type and chromosomal genotype of the pneumococcus. Since the opsonophagocytic activity is presumed to correlate with in vivo protection, these data indicate that the protein fraction has the potential to elicit species-specific immune protection with cross-protection against various pneumococcal strains. Individual proteins in the extract were purified by two-dimensional gel electrophoresis. Antibodies raised against three distinct proteins contributed to the opsonophagocytic activity of the serum. The proteins were identified by mass spectrometry and N-terminal amino acid sequencing. Two proteins were the previously characterized pneumococcal surface protein A and oligopeptide-binding lipoprotein AmiA. The third protein was the recently identified putative proteinase maturation protein A (PpmA), which showed homology to members of the family of peptidyl-prolyl cis/trans isomerases. Immunoelectron microscopy demonstrated that PpmA was associated with the pneumococcal surface. In addition, PpmA was shown to elicit species-specific opsonophagocytic antibodies that were cross-reactive with various pneumococcal strains. This antibody cross-reactivity was in line with the limited sequence variation of ppmA. The importance of PpmA in pneumococcal pathogenesis was demonstrated in a mouse pneumonia model. Pneumococcal ppmA-deficient mutants showed reduced virulence. The properties of PpmA reported here indicate its potential for inclusion in multicomponent protein vaccines. PMID- 10858236 TI - Comparison of protection in rabbits against host-adapted and cultivated Borrelia burgdorferi following infection-derived immunity or immunization with outer membrane vesicles or outer surface protein A. AB - In this study, infection-derived immunity in the rabbit model of Lyme disease was compared to immunity following immunization with purified outer membrane vesicles (OMV) isolated from Borrelia burgdorferi and recombinant outer surface protein A (OspA). Immunization of rabbits with OMV isolated from virulent strain B31 and its avirulent derivative B313 (lacking OspA and DbpA) conferred highly significant protection against intradermal injection with 6 x 10(4) in vitro cultivated virulent B. burgdorferi. This is the first demonstration of protective immunogenicity induced by OMV. While immunization with OspA and avirulent B31 OMV provided far less protection against this challenge, rabbits with infection derived immunity were completely protected. Protection against host-adapted B. burgdorferi was assessed by implantation of skin biopsies taken from rabbit erythema migrans (a uniquely rich source of B. burgdorferi in vertebrate tissue) containing up to 10(8) spirochetes. While all of the OMV- and OspA-immunized rabbits were fully susceptible to skin and disseminated infection, rabbits with infection-derived immunity were completely protected. Analysis of the antibody responses to outer membrane proteins, including DbpA, OspA, and OspC, suggests that the remarkable protection exhibited by the infection-immune rabbits is due to antibodies directed at antigens unique to or markedly up-regulated in host adapted B. burgdorferi. PMID- 10858237 TI - Cytopathogenic effect of Trichomonas vaginalis on human vaginal epithelial cells cultured in vitro. AB - In this study we established human vaginal epithelial cells (hVECs) in culture and evaluated their interaction with Trichomonas vaginalis parasites to complement previous studies using other cell types. Primary cultures of hVECs were established. Contaminating fibroblasts were separated from epithelial cells by differential trypsinization. Specific antibody staining revealed that over 92% of cells in hVEC monolayers were epithelial cells. T. vaginalis adhered to hVECs and produced severe cytotoxic effects resulting in obliteration of the monolayer within 24 h. Adherence and cytotoxicity were not observed when T. vaginalis was exposed to human vaginal fibroblasts or bovine vaginal epithelial cells. Likewise, the bovine parasite Tritrichomonas foetus had no cytotoxic effects on hVECs. We concluded that the interaction between T. vaginalis and hVECs is both cell specific (limited to epithelial cells and not vaginal fibroblasts) and species specific (limited to human vaginal cells and not bovine cells). Pretreatment of T. vaginalis with metronidazole or periodate abolished the adhesion of parasites to cell monolayers and the cytotoxic effect, suggesting involvement of carbohydrate-containing molecules in these processes. Different clinical isolates of T. vaginalis caused damage to cultured cells at different rates. Parasites separated from the vaginal cell monolayer by a permeable membrane did not produce a cytopathic effect, suggesting contact-dependent cytotoxicity. PMID- 10858238 TI - Effects of estradiol and progesterone on susceptibility and early immune responses to Chlamydia trachomatis infection in the female reproductive tract. AB - We have used a previously described rodent model to examine the influence of hormonal environment on susceptibility and immune responses to genital Chlamydia infection. Ovariectomized rats were administered estradiol, progesterone, or a combination of both, infected with Chlamydia trachomatis via the intrauterine route, and sacrificed 5 days later. Histopathological examination showed severe inflammation in the uteri and vaginae of progesterone-treated animals, whereas animals receiving estradiol or a combination of both hormones showed no inflammation. Large numbers of chlamydiae were found in vaginal secretions of progesterone-treated and combination-treated animals, while estradiol-treated animals had none. Tissue localization showed that numerous chlamydial inclusions were present in the uterine epithelium of the progesterone group and the cervicovaginal epithelium of the combination group. Examination of the acute immune responses of the infected animals showed that maximum activation was present in the draining lymph node cells from the progesterone-treated group, and these cells were producing large amounts of interleukin-10 and gamma interferon compared to other hormone-treated groups. In contrast, spleen cell proliferation was suppressed in progesterone-treated animals compared to other hormone-treated groups. We conclude that progesterone increases and estradiol decreases susceptibility to intrauterine chlamydial infection in this rat model. Our data demonstrate that hormone environment, at the time of infection, has a profound effect on the outcome of microbial infection in the female reproductive tract. PMID- 10858239 TI - The most abundant glycoprotein of amebic cyst walls (Jacob) is a lectin with five Cys-rich, chitin-binding domains. AB - The infectious stage of amebae is the chitin-walled cyst, which is resistant to stomach acids. In this study an extraordinarily abundant, encystation-specific glycoprotein (Jacob) was identified on two-dimensional protein gels of cyst walls purified from Entamoeba invadens. Jacob, which was acidic and had an apparent molecular mass of approximately 100 kDa, contained sugars that bound to concanavalin A and ricin. The jacob gene encoded a 45-kDa protein with a ladder like series of five Cys-rich domains. These Cys-rich domains were reminiscent of but not homologous to the Cys-rich chitin-binding domains of insect chitinases and peritrophic matrix proteins that surround the food bolus in the insect gut. Jacob bound purified chitin and chitin remaining in sodium dodecyl sulfate treated cyst walls. Conversely, the E. histolytica plasma membrane Gal/GalNAc lectin bound sugars of intact cyst walls and purified Jacob. In the presence of galactose, E. invadens formed wall-less cysts, which were quadranucleate and contained Jacob and chitinase (another encystation-specific protein) in secretory vesicles. A galactose lectin was found to be present on the surface of wall-less cysts, which phagocytosed bacteria and mucin-coated beads. These results suggest that the E. invadens cyst wall forms when the plasma membrane galactose lectin binds sugars on Jacob, which in turn binds chitin via its five chitin-binding domains. PMID- 10858240 TI - Cryptococcus neoformans is a facultative intracellular pathogen in murine pulmonary infection. AB - To produce chronic infection, microbial pathogens must escape host immune defenses. Infection with the human pathogenic fungus Cryptococcus neoformans is typically chronic. To understand the mechanism by which C. neoformans survives in tissue after the infection of immunocompetent hosts, we systematically studied the course of pulmonary infection in mice by electron microscopy. The macrophage was the primary phagocytic cell at all times of infection, but neutrophils also ingested yeast. Alveolar macrophages rapidly internalized yeast cells after intratracheal infection, and intracellular yeast cells were noted at all times of infection from 2 h through 28 days. However, the proportion of yeast cells in the intracellular and extracellular spaces varied with the time of infection. Early in infection, yeast cells were found predominantly in the intracellular compartment. A shift toward extracellular predominance occurred by 24 h that was accompanied by macrophage cytotoxicity and disruption. Later in infection, intracellular persistence in vivo was associated with replication, residence in a membrane-bound phagosome, polysaccharide accumulation inside cells, and cytotoxicity to macrophages, despite phagolysosomal fusion. Many phagocytic vacuoles with intracellular yeast had discontinuous membranes. Macrophage infection resulted in cells with a distinctive appearance characterized by large numbers of vacuoles filled with polysaccharide antigen. Similar results were observed in vitro using a macrophage-like cell line. Our results show that C. neoformans is a facultative intracellular pathogen in vivo. Furthermore, our observations suggest that C. neoformans occupies a unique niche among the intracellular pathogens whereby survival in phagocytic cells is accompanied by intracellular polysaccharide production. PMID- 10858241 TI - In vitro cell invasion of Mycoplasma gallisepticum. AB - The ability of the widespread avian pathogen Mycoplasma gallisepticum to invade cultured human epithelial cells (HeLa-229) and chicken embryo fibroblasts (CEF) was investigated by using the gentamicin invasion assay and a double immunofluorescence microscopic technique for accurate localization of cell associated mycoplasmas. The presence of intracellular mycoplasmas in both cell lines was clearly demonstrated, with organisms entering the eukaryotic cells within 20 min. Internalized mycoplasmas have the ability to leave the cell, but also to survive within the intracellular space over a 48-h period. Frequencies of invasion were shown to differ between the two cell lines, but were also considerably dependent on the mycoplasma input population. Of the prototype strain R, a low-passage population in artificial medium, R(low), was capable of active cell invasion, while a high-passage population, R(high), showed adherence to but nearly no uptake into HeLa-229 and CEF. By passaging R(low) and R(high) multiple times through HeLa-229 cells, the invasion frequency was significantly increased. Taken together, these findings demonstrate that M. gallisepticum has the capability of entering nonphagocytic host cells that may provide this pathogen with the opportunity for resisting host defenses and selective antibiotic therapy, establishing chronic infections, and passing through the respiratory mucosal barrier to cause systemic infections. PMID- 10858242 TI - Genetic locus for streptolysin S production by group A streptococcus. AB - Group A streptococcus (GAS) is an important human pathogen that causes pharyngitis and invasive infections, including necrotizing fasciitis. Streptolysin S (SLS) is the cytolytic factor that creates the zone of beta hemolysis surrounding GAS colonies grown on blood agar. We recently reported the discovery of a potential genetic determinant involved in SLS production, sagA, encoding a small peptide of 53 amino acids (S. D. Betschel, S. M. Borgia, N. L. Barg, D. E. Low, and J. C. De Azavedo, Infect. Immun. 66:1671-1679, 1998). Using transposon mutagenesis, chromosomal walking steps, and data from the GAS genome sequencing project (www.genome.ou.edu/strep. html), we have now identified a contiguous nine-gene locus (sagA to sagI) involved in SLS production. The sag locus is conserved among GAS strains regardless of M protein type. Targeted plasmid integrational mutagenesis of each gene in the sag operon resulted in an SLS-negative phenotype. Targeted integrations (i) upstream of the sagA promoter and (ii) downstream of a terminator sequence after sagI did not affect SLS production, establishing the functional boundaries of the operon. A rho independent terminator sequence between sagA and sagB appears to regulate the amount of sagA transcript produced versus transcript for the entire operon. Reintroduction of the nine-gene sag locus on a plasmid vector restored SLS activity to the nonhemolytic sagA knockout mutant. Finally, heterologous expression of the intact sag operon conferred the SLS beta-hemolytic phenotype to the nonhemolytic Lactococcus lactis. We conclude that gene products of the GAS sag operon are both necessary and sufficient for SLS production. Sequence homologies of sag operon gene products suggest that SLS is related to the bacteriocin family of microbial toxins. PMID- 10858243 TI - Intracellular trafficking of Brucella abortus in J774 macrophages. AB - Brucella abortus is a facultative intracellular bacterium capable of surviving inside professional and nonprofessional phagocytes. The microorganism remains in membrane-bound compartments that in several cell types resemble modified endoplasmic reticulum structures. To monitor the intracellular transport of B. abortus in macrophages, the kinetics of fusion of phagosomes with preformed lysosomes labeled with colloidal gold particles was observed by electron microscopy. The results indicated that phagosomes containing live B. abortus were reluctant to fuse with lysosomes. Furthermore, newly endocytosed material was not incorporated into these phagosomes. These observations indicate that the bacteria strongly affect the normal maturation process of macrophage phagosomes. However, after overnight incubation, a significant percentage of the microorganisms were found in large phagosomes containing gold particles, resembling phagolysosomes. Most of the Brucella bacteria present in phagolysosomes were not morphologically altered, suggesting that they can also resist the harsh conditions prevalent in this compartment. About 50% colocalization of B. abortus with LysoSensor, a weak base that accumulates in acidic compartments, was observed, indicating that the B. abortus bacteria do not prevent phagosome acidification. In contrast to what has been described for HeLa cells, only a minor percentage of the microorganisms were found in compartments labeled with monodansylcadaverine, a marker for autophagosomes, and with DiOC6 (3,3'-dihexyloxacarbocyanine iodide), a marker for the endoplasmic reticulum. These results indicate that B. abortus bacteria alter phagosome maturation in macrophages. However, acidification does occur in these phagosomes, and some of them can eventually mature to phagolysosomes. PMID- 10858244 TI - Ineffective cellular immune response associated with T-cell apoptosis in susceptible Mycobacterium bovis BCG-infected mice. AB - It has previously been reported that inhibition of delayed-type hypersensitivity mediating functions of T cells during mycobacterial infection in mice is haplotype dependent. In the present study, we show that Mycobacterium bovis BCG infection induced, in susceptible C57BL/6 and BALB/c mice but not in resistant C3H/HeJ and DBA/2 mice, an important splenomegaly. An in vitro defect in T-cell proliferation in response to T-cell receptor (TCR) stimulation with mitogens or anti-CD3 antibodies was associated with enhanced levels of CD4(+) and CD8(+) T cell apoptosis in susceptible but not in resistant mice 2 weeks after infection. Further investigations of C57BL/6 and C3H/HeJ mice revealed that in vivo splenomegaly was associated with destruction of the lymphoid tissue architecture, liver cellular infiltrates, and increased numbers of apoptotic cells in both spleen and liver tissue sections. Infection of C57BL/6 mice but not of C3H/HeJ mice induced massive production of tumor necrosis factor alpha (TNF-alpha) in serum, as well as an increase in Fas and Fas ligand (FasL) expression in T cells. In vitro addition of neutralizing anti-TNF-alpha antibodies led to a significant reduction in CD3-induced T-cell apoptosis of both CD4(+) and CD8(+) T cells of C57BL/6 mice, while the blockade of Fas-FasL interactions reduced apoptosis only in CD4(+) but not in CD8(+) T cells. Together, these results suggest that TNF alpha and Fas-FasL interactions play a role in the activation-induced cell death (AICD) process associated with a defect in T-cell proliferation of the susceptible C57BL/6 mice. T-cell death by apoptosis may represent one of the important components of the ineffective immune response against mycobacterium induced immunopathology in susceptible hosts. PMID- 10858245 TI - Influence of beta(2)-integrin adhesion molecule expression and pulmonary infection with Pasteurella haemolytica on cytokine gene expression in cattle. AB - beta(2)-Integrins are leukocyte adhesion molecules composed of alpha (CD11a, -b, c, or -d) and beta (CD18) subunit heterodimers. Genetic CD18 deficiency results in impaired neutrophil egress into tissues that varies between conducting airways and alveoli of the lung. In this study, we investigated whether CD18 deficiency in cattle affects proinflammatory cytokine (PIC) expression in pulmonary tissue after respiratory infection with Pasteurella haemolytica. Cattle were infected with P. haemolytica via fiberoptic deposition of organisms into the posterior part of the right cranial lung lobe. Animals were euthanized at 2 or 4 h postinoculation (p.i.), and tissues were collected to assess PIC gene expression using antisense RNA probes specific for bovine interleukin-1alpha (IL-1alpha), IL 1beta, IL-6, gamma interferon (IFN-gamma), and tumor necrosis factor alpha (TNF alpha) along with the beta-actin (beta-Act) housekeeping gene. Expression of PIC was induced at 2 h p.i. in P. haemolytica-infected cattle and continued to 4 h p.i. At 2 h p.i., induction of gene expression and increase of cells that expressed PIC were observed both in CD18(+) and CD18(-) cattle after inoculation of P. haemolytica. The induction of gene expression with P. haemolytica inoculation was more prominent in CD18(-) cattle than in CD18(+) cattle by comparison to pyrogen-free saline (PFS)-inoculated control animals. At 4 h p.i., however, the induction of PIC, especially IL-1alpha, IL-6, and IFN-gamma, in the lungs of CD18(+) cattle inoculated with P. haemolytica was greater than that in lungs of the CD18(-) cattle. IFN-gamma and TNF-alpha genes were not increased in P. haemolytica-inoculated CD18(-) cattle lungs compared to the PFS-inoculated control lungs at 4 h p.i. In PFS-inoculated lungs, we generally observed a higher percentage of cells and higher level of gene expression in the lungs of CD18(-) cattle than in the lungs of CD18(+) cattle, especially at 4 h p.i. The rate of neutrophil infiltration into the lungs of CD18(-) cattle at 2 h p.i. was significantly higher than that of CD18(+) cattle; at 4 h p.i., there was no difference between the two groups. These data suggest that beta(2)-integrins may contribute to the induction of expression of some PIC genes, as a consequence of P. haemolytica infection. PMID- 10858246 TI - Requirement for NF-kappaB in transcriptional activation of monocyte chemotactic protein 1 by Chlamydia pneumoniae in human endothelial cells. AB - Infection with Chlamydia pneumoniae, a causative agent of acute and chronic respiratory diseases, has recently been implicated as a potential risk factor in atherosclerosis. Atherosclerotic lesions are characterized by monocyte infiltration, which may be regulated by the chemokine monocyte chemotactic protein 1 (MCP-1). We have previously shown that C. pneumoniae infection stimulates MCP-1 production in human endothelial cells, an event which may be specific to this species of Chlamydia, since Chlamydia trachomatis infection fails to induce this response. To examine the underlying mechanisms by which C. pneumoniae infection induces MCP-1 production in endothelial cells, the present study investigated the role of transcription factor NF-kappaB in MCP-1 mRNA expression. Human umbilical vein endothelial cells (HUVEC) were infected with the coronary isolate C. pneumoniae A-03 or with C. trachomatis L2, and MCP-1 mRNA expression was assessed after different periods of infection by reverse transcription-PCR. Expression of MCP-1 mRNA in C. pneumoniae-infected HUVEC was significantly elevated as early as 1 h postinfection and increased dramatically by 12 and 24 h compared to baseline controls. Nuclear translocation of NF-kappaB occurred by 30 min of infection, as determined by electrophoretic mobility shift assays and immunofluorescence staining. Treatment of C. pneumoniae-infected HUVEC with parthenolide, a specific inhibitor of NF-kappaB activation, suppressed MCP-1 mRNA expression. In contrast, infection with C. trachomatis L2 did not induce MCP 1 mRNA in infected HUVEC and failed to activate NF-kappaB. Results from this study demonstrate a requirement for NF-kappaB activation in stimulation of MCP-1 gene expression by C. pneumoniae in human endothelial cells. Furthermore, the data suggest that, within the genus Chlamydia, functionally distinct signaling pathways leading to NF-kappaB activation are utilized by C. pneumoniae in endothelial cells during infection. PMID- 10858247 TI - CXC chemokine receptor CXCR2 is essential for protective innate host response in murine Pseudomonas aeruginosa pneumonia. AB - Pulmonary infection due to Pseudomonas aeruginosa has emerged as a leading cause of mortality. A vigorous host response is required to effectively clear the organisms from the lungs. This host defense is dependent on the recruitment and activation of neutrophils and macrophages. A family of chemotactic cytokines (chemokines) has been shown to participate in this protective response. In this study, we assessed the role of the ELR(+) (glutamic acid-leucine-arginine motif positive) CXC chemokines and their CXC chemokine receptor (CXCR2) in lung antibacterial host defense. The intratracheal administration of Pseudomonas to mice resulted in the time-dependent influx of neutrophils to the lung, peaking at 12 to 24 h after inoculation. The influx of neutrophils was associated with a similar time-dependent expression of the ELR(+) CXC chemokines, KC, macrophage inflammatory protein 2 (MIP-2), and lipopolysaccharide-induced CXC chemokine (LIX). Selective neutralization of MIP-2 or KC resulted in modest changes in neutrophil influx but no change in bacterial clearance or survival. However, neutralization of CXCR2 resulted in a striking increase in mortality, which was associated with a marked decrease in neutrophil recruitment and bacterial clearance. Conversely, the site-specific transgenic expression of KC resulted in enhanced clearance of bacteria after Pseudomonas challenge. This study indicates that ELR(+) CXC chemokines are critical mediators of neutrophil-mediated host defense in Pseudomonas pneumonia. PMID- 10858248 TI - The human cationic antimicrobial protein (hCAP-18) is expressed in the epithelium of human epididymis, is present in seminal plasma at high concentrations, and is attached to spermatozoa. AB - Innate immunity is important for the integrity of the host against potentially invasive pathogenic microorganisms in the environment. Antibiotic peptides with broad antimicrobial activity are part of the innate immune system. We investigated the presence of the cathelicidin, human cationic antimicrobial protein (hCAP-18), in the male reproductive system. We found strong expression of the hCAP-18 gene by in situ hybridization and hCAP-18 protein, as detected by immunohistochemistry, in the epithelium of the epididymis, but not in the testis. The highest expression in the epididymis was in the caudal part. Western blotting showed a doublet band, the upper part corresponding to the size of hCAP-18 in plasma and neutrophils. Using a specific enzyme-linked immunosorbent assay (ELISA), levels of 86.5 +/- 37.8 microg/ml (mean +/- standard deviation; range, 41.8 to 142.8 microg/ml; n = 10) were detected in seminal plasma from healthy donors, which is 70-fold higher than the level in blood plasma. Flow cytometry and immunocytochemistry revealed the presence of hCAP-18 on spermatozoa. ELISA measurement showed levels of 196 ng/10(6) spermatozoa, corresponding to 6.6 x 10(6) molecules of hCAP-18 per spermatozoon. Our results suggest a key role for hCAP-18 in the antibacterial integrity of the male reproductive system. The attachment of hCAP-18 to spermatozoa may implicate a role for hCAP-18 in conception. PMID- 10858249 TI - Helicobacter pylori modulates lymphoepithelial cell interactions leading to epithelial cell damage through Fas/Fas ligand interactions. AB - Helicobacter pylori causes a common chronic infection of humans that leads to epithelial cell damage. Studies have shown that apoptosis of the gastric epithelium is increased during infection and this response is associated with an expansion of gastric T-helper type 1 (Th1) cells. We report that gastric T cells contribute to apoptosis of the epithelium by a Fas/Fas ligand (FasL) interaction. Fas receptor expression was detected on freshly isolated gastric epithelial cells by flow cytometry and immunohistochemistry, and this level of expression was increased during infection with H. pylori. The expression of Fas receptor on three gastric epithelial cell lines was increased by H. pylori, either alone or in combination with gamma interferon or tumor necrosis factor alpha. The role of Fas in apoptosis of gastric epithelial cell lines was evidenced by DNA fragmentation after cross-linking of Fas with specific antibodies. FasL expression was detected by immunohistochemistry on mononuclear cells in gastric biopsy specimens of infected but not uninfected subjects. Gastric T-cell lines were also shown to express FasL, as evidenced by reverse transcription-PCR and killing of target cells expressing Fas receptor. Moreover, these T-cell lines were capable of killing cultured gastric epithelial target cells and antibodies that block the interaction between Fas receptor and FasL inhibited this cytotoxic activity. These observations demonstrate that local Th1 cells may contribute to the pathogenesis of gastric disease during H. pylori infection by increasing the expression of Fas on gastric epithelial cells and inducing apoptosis through Fas/FasL interactions. PMID- 10858250 TI - Antibodies against ribosomal phosphoprotein P0 of Plasmodium falciparum protect mice against challenge with Plasmodium yoelii. AB - Antibodies against the Plasmodium falciparum P0 ribosomal phosphoprotein (PfP0) have been detected exclusively but extensively in malaria-immune persons. Polyclonal rabbit and mice sera were raised against two recombinant polypeptides of P. falciparum P0 protein, PfP0N and PfP0C, covering amino acids 17 to 61 and the remaining amino acids 61 to 316, respectively. Sera against both these domains detected a 35-kDa protein from Plasmodium yoelii subsp. yoelii, a rodent malarial parasite, and stained the surface of merozoites in immunofluorescence assays. Total immunoglobulin G (IgG) purified from rabbit and mouse anti-PfP0 sera by ammonium sulfate and DEAE-cellulose chromatography was used for passive transfer experiments in mice. Mice passively immunized with both anti-PfP0N and anti-PfP0C showed distinctly lower levels of parasitemia than control mice. With immunizations on days -1, 0, 1, 3, and 5, about 50% of both sets of mice receiving anti-PfP0N and anti-PfP0C cleared the lethal 17XL strain of P. yoelii and revived by day 25. All the control mice died by day 10. By extending the immunization schedule, the survival period of the mice could be extended for every mouse that received anti-PfP0 IgG. These data demonstrate the cross protection of the anti-PfP0 IgG and establish parasite P0 protein as a target for invasion-blocking antibodies. PMID- 10858251 TI - The Entamoeba histolytica mitochondrion-derived organelle (crypton) contains double-stranded DNA and appears to be bound by a double membrane. AB - Amebae have an Hsp60-associated, mitochondrion-derived organelle (crypton). In this study, the crypton was stained with multiple DNA-binding fluorochromes and a monoclonal anti-double-stranded DNA antibody. Transmission microscopy of partially purified cryptons revealed organelles bound by a double membrane. PMID- 10858252 TI - Motility is required to initiate host cell invasion by Yersinia enterocolitica. AB - Invasin-mediated invasion of host cells by the pathogen Yersinia enterocolitica was shown to be affected by flagellar-dependent motility. Motility appears to be required to ensure the bacterium migrates to and contacts the host cell. Nonmotile strains of Y. enterocolitica were less invasive than motile strains, but the reduction in invasion could be overcome by artificially bringing the bacteria into host cell contact by centrifugation. Mutations in known regulatory genes of the flagellar regulon, flhDC and fliA, resulted in less inv expression but did not have a significant effect on invasin levels. However, invasin levels were reduced for strains that harbored flhDC on a multicopy plasmid, apparently as a result of increased proteolysis of invasin. PMID- 10858253 TI - Binding and activation of human plasminogen by Mycobacterium tuberculosis. AB - The first evidence of the interaction of Mycobacterium tuberculosis with the plasminogen system is herein reported. By FACScan analysis and affinity blotting, lysine-dependent binding of plasminogen to M. tuberculosis was demonstrated. The binding molecules were 30-, 60-, and 66-kDa proteins present in cell wall and soluble protein extracts. The activation of plasminogen, which occurred only in presence of fibrin and was not inhibited by the host serpin, alpha(2) antiplasmin, was also demonstrated. PMID- 10858254 TI - Pseudomonas aeruginosa cell-to-cell signaling is required for virulence in a model of acute pulmonary infection. AB - Cell-to-cell signaling controls many virulence genes in Pseudomonas aeruginosa. We tested the virulence of las and rhl quorum-sensing mutants in neonatal mice. A lasI rhlI double mutant was nearly avirulent, and the respective single mutant strains were reduced in virulence compared with the wild-type strain. Quorum sensing plays a role in P. aeruginosa pneumonia in neonatal mice. PMID- 10858255 TI - Adherence of isogenic flagellum-negative mutants of Helicobacter pylori and Helicobacter mustelae to human and ferret gastric epithelial cells. AB - Isogenic flagellum-negative mutants of Helicobacter pylori and Helicobacter mustelae were screened for their ability to adhere to primary human and ferret gastric epithelial cells, respectively. We also evaluated the adherence of an H. pylori strain with a mutation in the flbA gene, a homologue of the flbF/lcrD family of genes known to be involved in the regulation of H. pylori flagellar biosynthesis. H. pylori and H. mustelae mutants deficient in production of FlaA or FlaB and mutants deficient in the production of both FlaA and FlaB showed no reduction in adherence to primary human or ferret gastric epithelial cells compared with the wild-type parental strains. However, adherence of the H. pylori flbA mutant to human gastric cells was significantly reduced compared to the adherence of the wild-type strain. These results show that flagella do not play a direct role in promoting adherence of H. pylori or H. mustelae to gastric epithelial cells. However, genes involved in the regulation of H. pylori flagellar biosynthesis may also regulate the production of an adhesin. PMID- 10858256 TI - Host resistance to Listeria monocytogenes infection is enhanced but resistance to Staphylococcus aureus infection is reduced in acute graft-versus-host disease in mice. AB - Acute graft-versus-host disease (GVHD) is characterized by the production of high levels of T helper 1 (Th1)-type cytokines. Bone marrow transplantation from allogeneic C57BL/6 cells to CBF(1) mice produced acute GVHD. Host resistance to Th1-driven Listeria monocytogenes was enhanced, whereas host resistance to Th2 driven Staphylococcus aureus was reduced during acute GVHD. These results suggest that opposite host responses are observed between Th1-driven and Th2-driven bacterial infections in acute GVHD. PMID- 10858257 TI - Mechanical fractionation reveals structural requirements for enteropathogenic Escherichia coli Tir insertion into host membranes. AB - Enteropathogenic Escherichia coli (EPEC) inserts its receptor for intimate adherence (Tir) into host cell membranes by using a type III secretion system. Detergents are frequently used to fractionate infected host cells to investigate bacterial protein delivery into mammalian cells. In this study, we found that the Triton X-100-soluble membrane fraction from EPEC-infected HeLa cells was contaminated with bacterial proteins. We therefore applied a mechanical method of cell lysis and ultracentrifugation to fractionate infected HeLa cells to investigate the biology and biochemistry of Tir delivery and translocation. This method demonstrates that the translocation of Tir into the host cell membrane requires its transmembrane domains, but not tyrosine phosphorylation or binding to Tir's ligand, intimin. PMID- 10858258 TI - Adjuvant activity of a nontoxic mutant of Escherichia coli heat-labile enterotoxin on systemic and mucosal immune responses elicited against a heterologous antigen carried by a live Salmonella enterica serovar Typhimurium vaccine strain. AB - Systemic and mucosal antibody responses against both the major subunit of colonization factor antigen I (CFA/I) of enterotoxigenic Escherichia coli (ETEC) and the somatic lipopolysaccharide expressed by recombinant bivalent Salmonella vaccine strains were significantly enhanced by coadministration of a detoxified derivative with preserved adjuvant effects of the ETEC heat-labile toxin, LT((R192G)). The results further support the adjuvant effects of LT((R192G)) and represent a simple alternative to improve responses against passenger antigens expressed by orally delivered Salmonella vaccine strains. PMID- 10858259 TI - Mutational analysis of the Helicobacter pylori vacuolating toxin amino terminus: identification of amino acids essential for cellular vacuolation. AB - The functional importance of the amino terminus of the Helicobacter pylori vacuolating cytotoxin (VacA) was investigated by analyzing the relative levels of vacuolation of HeLa cells transfected with plasmids encoding wild-type and mutant forms of the toxin. Notably, VacA's intracellular activity was found to be sensitive to small truncations and internal deletions at the toxin's amino terminus. Moreover, alanine-scanning mutagenesis revealed the first VacA point mutations (at proline 9 or glycine 14) that completely abolish the toxin's intracellular activity. PMID- 10858260 TI - Host and tissue specificity of Trichomonas vaginalis is not mediated by its known adhesion proteins. AB - Adhesion of Trichomonas vaginalis is believed to be dependent on four adhesion proteins, which are thought to bind to vaginal epithelial cells in a specific manner with a ligand-receptor type of interaction. However, the specific receptors on the host cell have not yet been identified. In this work, the ability of the T. vaginalis adhesins to bind to cells of different histologic derivations and from different species has been studied. HeLa, CHO, and Vero cell lines; erythrocytes from different species; and a prokaryote without a cell wall, Mycoplasma hominis, were employed in order to investigate the cell specificity of the T. vaginalis adhesins. We observed that the T. vaginalis adhesins are able to bind to the different cell types to the same extent, suggesting that the host and tissue specificity of T. vaginalis adhesion should not be due to specificity of the parasite adhesins. Our results suggest that the data published to date on the subject are probably artifactual and that the experiments reported in the literature are not appropriate for identification of protozoan adhesins. PMID- 10858261 TI - Granulocytic ehrlichiosis in mice deficient in phagocyte oxidase or inducible nitric oxide synthase. AB - Mice deficient in phox (gp91(phox-/-)) or NOS2 (NOS2(-/-)) were infected with the agent of human granulocytic ehrlichiosis (HGE) to evaluate the importance of these pathways in the eradication of HGE bacteria. NOS2(-/-) mice had delayed clearance of the HGE agent in comparison to control or gp91(phox-/-) mice, suggesting that reactive nitrogen intermediates play a role in the early control of HGE. PMID- 10858262 TI - Cytocidal and apoptotic effects of the ClyA protein from Escherichia coli on primary and cultured monocytes and macrophages. AB - Cytolysin A (ClyA) is a newly discovered cytolytic protein of Escherichia coli K 12 that mediates a hemolytic phenotype. We show here that highly purified ClyA and ClyA-expressing E. coli were cytotoxic and apoptogenic to fresh as well as cultured human and murine monocytes/macrophages. PMID- 10858263 TI - Expression of AniA, the major anaerobically induced outer membrane protein of Neisseria gonorrhoeae, provides protection against killing by normal human sera. AB - Anaerobically grown Neisseria gonorrhoeae has previously been shown to have elevated serum resistance in the absence of exogenous CMP-N-acetylneuraminic acid or detectable sialylation. We hypothesized that the anaerobically induced gonococcal outer membrane protein AniA might have a role in this phenomenon, as it is the only known gonococcal protein that is absent under aerobic conditions. An N. gonorrhoeae F62 derivative, RUG7035, in which aniA is under control of the tac promoter, was used to examine the effect of AniA expression on serum resistance. In this study, we found that expression of AniA enhanced the serum resistance of N. gonorrhoeae and may account for these earlier observations. PMID- 10858264 TI - Long-term immunological memory induced by recombinant oral Salmonella vaccine vectors. AB - We have previously shown that Salmonella enterica serovar Typhimurium expressing the hagB hemagglutinin gene from Porphyromonas gingivalis can induce primary and recall immune responses in serum and secretions in mice; however, the longevity of memory induced by oral Salmonella carriers has not been adequately demonstrated. In this study, we examined the capacity of mice to mount a recall response 52 weeks after primary immunization. Recall responses were seen in serum immunoglobulin G (IgG) and IgA following boosting at week 52, and in most cases, they were equal to or greater than the primary responses. Significant mucosal IgA recall responses in saliva and vaginal wash were also detected following boosting at week 52. In addition, there was a considerable residual response in secretions at week 51, prior to boosting. These results indicate that oral Salmonella vectors can induce long-term memory to recombinant HagB and are particularly effective at inducing long-lasting mucosal responses as well as at inducing the capacity for mucosal recall responses. PMID- 10858265 TI - Applications of plastic embedding to electron microscopic immunocytochemistry and in situ hybridization in observations of production and secretion of peptide hormones. AB - Plastic embedding has been used to localize various antigens in conjunction with immunohistochemistry. Peptide hormones have been among the antigens that have been studied extensively. Recent application of water-soluble plastics such as LR White and Lowicryl has extended the ranges of detectable antigens and enabled the observation of antigen-antigen or mRNA-antigen combinations. This review article deals with technical aspects, procedures, and applications in endocrine cells. PMID- 10858266 TI - Calculation of maximal hybridization capacity (Hmax) for quantitative in situ hybridization: a case study for multiple calmodulin mRNAs. AB - In estimations of mRNA copy numbers, quantitative in situ hybridization (ISH) is expected to be performed at saturating probe concentration. In practice, however, this condition can rarely be fulfilled when medium to high amounts of transcripts exist and/or in large-scale experiments. To resolve this problem, we developed and tested a double-step procedure involving the use of calmodulin (CaM) I, II, and III [(35)S]-cRNA probes on adult rat brain sections; the hybridization signals were detected with a phosphorimager. By means of hybridization at increasing probe concentrations for a time sufficient for saturation, saturation curves were created for and maximal hybridization capacity (Hmax) values were assigned to selected brain areas. The Kd values of these various brain areas did not differ significantly, which allowed the creation and use of one calibration graph of Hmax vs hybridized [(35)S]-cRNA values for all brain areas for a given probe concentration. Large-scale ISH experiments involving a subsaturating probe concentration were performed to estimate Hmax values for multiple CaM mRNAs. A calibration graph corresponding to this probe concentration was created and Hmax values (expressed in ISH copy no/mm(2) units) were calculated for several brain regions via the calibration. The value of the method was demonstrated by simultaneous quantification of the total accessible multiple CaM mRNA contents of several brain areas in a precise and economical way. PMID- 10858267 TI - Spatiotemporal expression, distribution, and processing of POMC and POMC-derived peptides in murine skin. AB - In murine skin, after depilation-induced anagen, there was a differential spatial and temporal expression of pro-opiomelanocortin (POMC) mRNA, of the POMC-derived peptides beta-endorphin, ACTH, beta-MSH, and alpha-MSH, and of the prohormone convertases PC1 and PC2 in epidermal and hair follicle keratinocytes and in the cells of sebaceous units. Using a combination of in situ hybridization histochemistry and immunohistochemistry, we found cell-specific variations in the expression of POMC mRNA that were consistent with immunoreactivities for POMC derived peptides. Cells that contained POMC peptide immunoreactivity (IR) also expressed POMC mRNA, and where the IR increased there was a parallel increase in mRNA. The levels of PC1-IR and PC2-IR also showed cell-specific variations and were present in the same cells that contained the POMC peptides. Based on the cleavage specificities of these convertases and on the spatial and temporal expression of the convertases and of ACTH, beta-endorphin, beta-MSH, and alpha MSH, we can infer that the activities of PC1 and PC2 are responsible for the cell specific differential processing of POMC in murine skin. PMID- 10858268 TI - Placental expression and chromosomal localization of the human Gcm 1 gene. AB - Although gcm was first recognized for its role in specifying glial cell fate in Drosophila melanogaster, its mammalian counterparts are expressed predominantly in non-neural tissues. Here we demonstrate expression of the mouse and human GCM 1 proteins in placenta. We have prepared a highly specific antibody that recognizes the GCM 1 protein and have used it to assess the temporal and spatial expression profile of the protein. In both mouse and human placenta, the protein is associated with cells that are involved with exchange between maternal and fetal blood supplies: the labyrinthine cells of the mouse placenta and the syncytio- and cytotrophoblasts of the human placenta. Using the full-length hGcm 1 cDNA as a probe, we have mapped the gene on human chromosome 6p12 by fluorescent in situ hybridization. PMID- 10858269 TI - Lectin histochemistry of the spleen: a new lectin visualizes the stromal architecture of white pulp and the sinuses of red pulp. AB - The subcompartmentalization of the white pulp in the spleen is the result of interactions of specific resident stromal cells and migrating subtypes of lymphocytes. Because carbohydrate residues of cell membranes and extracellular matrices are involved in cell-cell and cell-matrix interactions, they were investigated in rat spleen by a broad panel of lectins. Splenic macrophages, which were also demonstrated by Perls' Prussian blue reaction, were labeled selectively by most mannose-specific lectins and gave the characteristic distribution patterns in all splenic (sub)compartments. One recently isolated lectin, Chelidonium majus agglutinin (CMA), visualized predominantly central arterioles, the reticular meshwork (RM) in the periarteriolar lymphatic sheaths (PALS), the circumferential reticulum cells limiting PALS and follicles, and some follicular dendritic cells (FDCs) in white pulp. The endothelial cells of venous sinuses in red pulp were also labeled by CMA and, if frozen sections were used, CMA also labeled the macrophages of the red pulp. Compared to CMA, the monoclonal antibody CD11, which can be used only in frozen sections, stained almost solely the fibrous (extracellular) component of the RM. Because CMA stains the reticulum cells in particular, it is better suited to visualize the stromal architecture of splenic white pulp than the monoclonal antibody. Because CMA can be applied to paraffin-embedded material, it is a particularly useful tool to study the splenic stromal architecture in archival material. PMID- 10858270 TI - Signal amplification in immunohistochemistry at the light microscopic level using biotinylated tyramide and nanogold-silver staining. AB - Signal amplification techniques greatly enhance the sensitivity of immunohistochemical (IHC) and in situ hybridization (ISH) methods. In particular, catalyzed signal amplification (CSA) using labeled tyramide or Nanogold-silver staining is an important signal amplification tool. We have applied a combination of both techniques, as has been introduced for ISH, for a further increase in sensitivity of an IHC method to detect cathepsin B. This lysosomal proteinase can also be expressed extracellularly, particularly in relation to cancer metastasis. Higher sensitivity of the IHC method was needed because existing methods failed to demonstrate cathepsin B protein where cathepsin B activity was found with a fluorescence enzyme histochemical method. Combined CSA and Nanogold-silver staining provided the sensitivity that was required. Moreover, this signal amplification method enabled the use of a 10-fold lower concentration of primary antibody (1 microg/ml). Nonspecific background staining was low provided that endogenous biotin, avidin, and peroxidase were completely blocked. The method was reproducible when all steps, and particularly the silver enhancement step, were rigidly controlled. The method resulted in localization patterns of cathepsin B protein that were in agreement with those of cathepsin B activity in serial sections of rat liver containing colon cancer metastases. We concluded that combined application of CSA and Nanogold-silver staining provides high sensitivity for immunohistochemical methods and that activity localization by an enzyme histochemical method is a very attractive alternative to IHC localization of an enzyme because it is at least as sensitive, it is rapid and simple, and it provides direct information on the function of an enzyme. PMID- 10858271 TI - Immunohistochemical localization of histamine N-methyltransferase in guinea pig tissues. AB - Histamine plays important roles in gastric acid secretion, inflammation, and allergic response. Histamine N-methyltransferase (HMT; EC 2.1.1.8) is crucial to the inactivation of histamine in tissues. In this study we investigated the immunohistochemical localization of this enzyme in guinea pig tissues using a rabbit polyclonal antibody against bovine HMT. The specificity of the antibody for guinea pig HMT was confirmed by Western blotting and the lack of any staining using antiserum preabsorbed with purified HMT. There was strong HMT-like immunoreactivity (HMT-LI) in the epithelial cells in the gastrointestinal tract, especially in the gastric body, duodenum, and jejunum. The columnar epithelium in the gallbladder was also strongly positive. Almost all the myenteric plexus from the stomach to the colon was stained whereas the submucous plexus was not. Other strongly immunoreactive cells included the ciliated cells in the trachea and the transitional epithelium of the bladder. Intermediately immunoreactive cells included islets of Langerhans, epidermal cells of the skin, alveolar cells in the lung, urinary tubules in the kidney, and epithelium of semiferous tubules. HMT-LI was present in specific structures in the guinea pig tissues. The widespread distribution of HMT-LI suggests that histamine has several roles in different tissues. PMID- 10858272 TI - An artifactual in situ hybridization signal associated with apoptosis in rat embryo. AB - We report an artifactual in situ hybridization (ISH) labeling pattern in embryonic rat tissues. It is caused by a short multiple cloning site-derived sequence incorporated into the RNA probes by in vitro transcription of templates cloned into pBluescript or its descendants. The artifact was seen in tissues in which programmed cell death (apoptosis) takes place during embryogenesis, i.e., in the mesonephric area, developing nervous system, interdigital mesenchyme of the hand plate, and permanent kidney. Labeling of the radioactive ISH with TUNEL verified the co-localization of the artifactual hybridization signal with cells at early stages of apoptosis. Even though the identity of the hybridization target in apoptotic cells remains unknown, it might be highly species-specific, because this artifact was never observed in mouse tissues. PMID- 10858273 TI - Protease-elicited TUNEL positivity of non-apoptotic fixed cells. AB - The appearance of free DNA ends in the chromatin is usually considered an indication of advanced apoptosis. Unexpectedly, the nuclei of non-apoptotic cells derived from mouse thymuses could be specifically labeled by terminal transferase after proteinase K treatment of the fixed, cytocentrifuged samples. Artifactual mechanical or contaminating nucleolytic factors have been ruled out as players in the generation of free DNA ends. The phenomenon was detected in both formaldehyde and ethanol-fixed specimens, in agarose-embedded fixed cells, and in chromatin spreads. By urea-agarose gel electrophoresis, the average single-strand size of the DNA molecules carrying the free ends was found between 50 and 250 kb. We suggest that ss discontinuities preexisting in the fixed normal cells are unmasked by protease treatment eliciting TUNEL (terminal transferase-mediated nick end-labeling) positivity. PMID- 10858274 TI - Posttranslational regulation of glucose-6-phosphate dehydrogenase activity in tongue epithelium. AB - Expression of glucose-6-phosphate dehydrogenase (G6PD) activity is high in tongue epithelium, but its exact function is still unknown. It may be related either to the high proliferation rate of this tissue or to protection against oxidative stress. To elucidate its exact role, we localized quantitatively G6PD activity, protein and mRNA using image analysis in tongue epithelium of rat and rabbit, two species with different diets. Distribution patterns of G6PD activity were largely similar in rat and rabbit but the activities were twofold lower in rabbit. Activity was two to three times higher in upper cell layers of epithelium than in basal cell layers, whereas basal layers, where proliferation takes place, contained twice as much G6PD protein and 40% more mRNA than upper layers. Our findings show that G6PD is synthetized mainly in basal cell layers of tongue epithelium and that it is posttranslationally activated when cells move to upper layers. Therefore, we conclude that the major function of G6PD activity in tongue epithelium is the formation of NADPH for protection against oxidative stress and that diet affects enzyme expression in this tissue. PMID- 10858275 TI - Epithelial rests of Malassez express immunoreactivity of TrkA and its distribution is regulated by sensory nerve innervation. AB - The periodontal ligament is the connective tissue that fills the space between the tooth and its bony socket. It is abundantly innervated by the sensory and sympathetic nerves. We first investigated the immunoreactivity of TrkA, which is a high-affinity receptor of nerve growth factor (NGF), in the periodontal ligament of rats. Immunoreactivity was observed at the epithelial cells in the cervical and furcation regions of the molars. These epithelial cells, which gather together to form clusters or networks, are known as the epithelial rests of Malassez. Immunoreactivity was not observed in other non-neuronal cells, such as osteoblasts, fibroblasts, odontoblasts, cementoblasts, endothelial cells, and/or osteoclasts. On the basis of these findings, we investigated the possible involvement of sensory nerve innervation in the immunoreactivity of the epithelial cells. Denervation of the inferior alveolar nerve resulted in a marked decrease in the distribution area and size of the clusters of immunoreactive cells compared with those of sham-operated rats. These findings suggest that sensory nerve innervation may have a regulatory role in maintenance of the epithelial rests of Malassez expressing TrkA in the periodontal ligament. PMID- 10858276 TI - Immunolocalization of tenascin-C, alpha9 integrin subunit, and alphavbeta6 integrin during wound healing in human oral mucosa. AB - Tenascin-C (TN-C) and its isoforms are multidomain extracellular matrix (ECM) proteins that are believed to be involved in the regulation of stromal-epithelial interactions. Some of the interactions between TN-C and cells are mediated by integrins. In this study we analyzed the expression of TN-C and its large molecular weight splice isoform (TN-C(L)) and the putative TN-C-binding alpha9 and alphavbeta6 integrins during human wound repair. In 3-day-old oral mucosal wounds, immunoreactivity for alpha9 integrin localized abundantly at the migrating basal wound epithelial cells. TN-C and TN-C(L) were localized in the matrix between and underneath alpha9-expressing epithelial cells. In parallel with gradual downregulation of alpha9 integrin immunoreactivity in 7-day and older wounds, the expression of alphavbeta6 integrin was temporarily induced. Integrin alphavbeta6 co-localized in the same area as TN-C and TN-C(L) immunoreactivity at the cell-cell contacts of the basal and suprabasal cell layers of the wound epithelium. During granulation tissue formation and reorganization from 7 to 28 days after wounding, TN-C and TN-C(L) were abundantly localized in the granulation tissue. The findings show that TN-C(L) is expressed under the migrating epithelial front and in the granulation tissue during matrix deposition in wound repair. Preferential localization of alpha9 integrin in migrating epithelial cells and of alphavbeta6 integrin in epithelium after wound closure suggests different functions for these integrins in wound repair. PMID- 10858277 TI - Cellular localization of gene expression for progranulin. AB - Granulins, also called epithelins, are 6-kD peptides with growth modulatory effects on a variety of cells. The granulin/epithelin precursor supports tumorigenesis in appropriate cell models and is the only growth factor able to overcome the cell cycle block that occurs in murine fibroblasts after deletion of a functional IGF-1 receptor. However, little is known of the role of granulin/epithelin gene products in vivo. To understand the physiological role of granulins it is essential to know the cell types and conditions in which it is expressed. We examined granulin/epithelin gene expression in adult rodents by in situ hybridization. The granulin/epithelin precursor is constitutively expressed in a number of epithelia, particularly in the skin, GI tract, and reproductive system. Other epithelia express the gene less strongly. Progranulin is expressed in immune cells in vivo and in specific neurons in the brain, including Purkinje cells, pyramidal cells of the hippocampus, and some cerebral cortical neurons. Little expression was detected in muscle cell, connective tissue, or endothelium. Cumulatively, these results define the basal gene expression of a new growth factor system and suggest that the progranulin/epithelin gene is multifunctional, with important constitutive roles in epithelial homeostasis, reproductive, immunological, and neuronal function. PMID- 10858278 TI - Unaltered distribution of laminins, fibronectin, and tenascin in celiac intestinal mucosa. AB - Extensive remodeling of the extracellular matrix (ECM) occurs in inflammatory tissues. The celiac lesion in the small intestine is characterized by inflammation accompanied by profound morphological alterations. We used immunohistochemistry to determine the distribution of laminin, fibronectin, and tenascin isoforms in small intestinal biopsies of untreated patients with celiac disease. In normal mucosa, the distribution of laminin isoforms defines three epithelial basement membrane (BM) zones. We found that the organization of these zones was maintained in the celiac mucosa. Thus, components of laminin-5 (alpha3 and beta3) were found in the surface epithelial BM, laminin alpha2 chain was found selectively at crypt bottoms, and laminin alpha5 chain was the sole alpha type chain in middle crypt BMs. Likewise, the distribution of fibronectin and tenascin resembled that of the normal gut. The organization of pericryptal fibroblasts and lamina propria smooth muscle strands, as defined by immunostaining for alpha-smooth muscle actin, also remained unchanged in the celiac mucosa. Unexpectedly, major ECM changes were not detected in the celiac lesion. PMID- 10858279 TI - Structural similarities between MutT and the C-terminal domain of MutY. AB - One of the functions of MutY from Escherchia coli is removal of adenine mispaired with 7,8-dihydro-8-oxoguanine (8-oxoG), a common lesion in oxidatively damaged DNA. MutY is composed of two domains: the larger N-terminal domain (p26) contains the catalytic properties of the enzyme while the C-terminal domain (p13) affects substrate recognition and enzyme turnover. On the basis of sequence analyses, it has been recently suggested that the C-terminal domain is distantly related to MutT, a dNTPase which hydrolyzes 8-oxo-dGTP [Noll et al. (1999) Biochemistry 38, 6374-6379]. We have studied the solution structure of the C-terminal domain of MutY by NMR and find striking similarity with the reported solution structure of MutT. Despite low sequence identity between the two proteins, they have similar secondary structure and topology. The C-terminal domain of MutY is composed of two alpha-helices and five beta-strands. The NOESY data indicate that the protein has two beta-sheets. MutT is also a mixed alpha/beta protein with two helices and two beta-sheets composed of five strands. The secondary structure elements are similarly arranged in the two proteins. PMID- 10858280 TI - Domains I and III of the human copper chaperone for superoxide dismutase interact via a cysteine-bridged Dicopper(I) cluster. AB - Copper binding to the human copper chaperone for superoxide dismutase (hCCS) has been investigated by X-ray absorption spectroscopy. Stoichiometry measurements on the dialyzed, as-isolated protein indicated that up to 3.5 Cu ions bound per hCCS molecule. Reduction with either sodium dithionite or dithiothreitol decreased the copper binding ratio to 2 coppers per hCCS monomer. Analysis of the as-isolated EXAFS data indicated coordination of Cu by a mixture of S and N backscatterers, suggestive of heterogeneous binding of copper between Cu-cysteine binding sites of domain I or III and copper-histidine SOD1-like metal binding sites of domain II. The best fit was obtained with 1.6 Cu-S (cysteine) at 2.24 A (2sigma(2) = 0.011 A(2)) and 1.1 N (histidine) at 1.98 A (2sigma(2) = 0.005 A(2)). A peak of variable intensity in the Fourier transform (FT) of the as-isolated protein at 2.7 A was suggestive of the presence of a heavy atom scatterer such as Cu. Analysis of the dithionite- and DTT-reduced derivatives indicated that copper was lost from the histidine coordinating sites, resulting in a S-only environment with copper coordinated to three S backscatterers at 2. 26 A. The heavy atom scatterer peak was now prominent in the FT and could be well fit by a Cu-Cu interaction at 2.72 A. The data were best interpreted by a dinuclear mu(2)() bridged cluster with doubly bridging cysteine ligands similar to the cluster proposed to exist in the cytochrome c oxidase chaperone COX17. Analysis of primary sequence and X-ray structural information on yeast CCS strongly suggests that this cluster bridges between domains I and III in hCCS. A mechanism for copper translocation is briefly discussed. PMID- 10858281 TI - Ligand-induced changes in the structure and dynamics of a human class Mu glutathione S-transferase. AB - Glutathione transferases are detoxification enzymes that catalyze the addition of glutathione (GSH) to a wide variety of hydrophobic compounds. Although this group of enzymes has been extensively characterized by crystallographic studies, little is known about their dynamic properties. This study investigates the role of protein dynamics in the mechanism of a human class mu enzyme (GSTM2-2) by characterizing the motional properties of the unliganded enzyme, the enzyme substrate (GSH) complex, an enzyme-product complex [S-(2,4 dinitrobenzyl)glutathione, GSDNB], and an enzyme-inhibitor complex (S-1 hexylglutathione, GSHEX). The kinetic on- and off-rates for these ligands are 10 20-fold lower than the diffusion limit, suggesting dynamic conformational heterogeneity of the active site. The off-rate of GSDNB is similar to the turnover number for its enzymatic formation, suggesting that product release is rate-limiting when 1-chloro-2,4-dinitrobenzene is the substrate. The dynamic properties of GSTM2-2 were investigated over a wide range of time scales using (15)N nuclear spin relaxation, residual dipolar couplings, and amide hydrogen deuterium exchange rates. These data show that the majority of the protein backbone is rigid on the nanosecond to picosecond time scale for all forms of the enzyme. The presence of motion on the millisecond to microsecond time scale was detected for a small number of residues within the active site. These motions are likely to play a role in facilitating substrate binding and product release. The residual dipolar couplings also show that the conformation of the active site region is more open in solution than in the crystalline environment, further enhancing ligand accessibility to the active site. Amide hydrogen-deuterium exchange rates indicate a reduction in the dynamic properties of several residues near the active site due to the binding of ligand. GSH binding reduces the exchange rate of a number of residues in proximity to its binding site, while GSHEX causes a reduction in amide-exchange rates throughout the entire active site region. The location of the dinitrobenzene (DNB) ring in the GSDNB-GSTM2-2 complex was modeled using chemical shift changes that occur when GSDNB binds to the enzyme. The DNB ring makes a number of contacts with hydrophobic residues in the active site, including Met108. Replacement of Met108 with Ala increases the turnover number of the enzyme by a factor of 1.7. PMID- 10858282 TI - Self-assembly of the binuclear metal center of phosphotriesterase. AB - The active site of the bacterial phosphotriesterase (PTE) from Pseudomonas diminuta contains two divalent metal ions and a carboxylated lysine residue. The native enzyme contains two Zn(2+) ions, which can be replaced with Co(2+), Cd(2+), Ni(2+), or Mn(2+) without loss of catalytic activity. Carbon dioxide reacts with the side chain of lysine-169 to form a carbamate functional group within the active site, which then serves as a bridging ligand to the two metal ions. The activation of apo-PTE using variable concentrations of divalent metal ions and bicarbonate was measured in order to establish the mechanism by which the active site of PTE is self-assembled. The time courses for the activation of apo-PTE are pseudo-first-order, and the observed rate constants are directly proportional to the concentration of bicarbonate. In contrast, the apparent rate constants for the activation of apo-PTE decrease as the concentrations of the divalent cations are increased and then become constant at higher concentrations of the divalent metal ions. These results are consistent with a largely ordered kinetic mechanism for the assembly of the binuclear metal center where CO(2)/bicarbonate reacts with the apo-PTE prior to the binding of the two metal ions. When apo-PTE is titrated with 0-8 equiv of Co(2+), Cd(2+), or Zn(2+), the concentration of activated enzyme increases linearly until 2 equiv of metal ion is added and then remains constant at elevated levels of the divalent cations. These results are consistent with the synergistic binding of the two metal ions to the active site, and thus the second metal ion binds more tightly to the protein than does the first metal ion. Measurement of the mean dissociation constant indicates that metal binding to the binuclear metal center is strong [(K(alpha)K(beta))(1/2) = 6.0 x 10(-)(11) M and k(off) = 1.5 x 10(-)(3) min(-)(1) for Zn(2+)]. The removal of the carbamate bridge through the mutagenesis of Lys 169 demonstrates that the carbamate bridge is required for both efficient catalysis and overall stability of the metal center. PMID- 10858283 TI - Functional characterization of DNase X, a novel endonuclease expressed in muscle cells. AB - The activation of endonucleases resulting in the degradation of genomic DNA is one of the most characteristic changes in apoptosis. Here, we report the characterization of a novel endonuclease, termed DNase X due to its X-chromosomal localization. The active nuclease is a 35 kDa protein with 39% identity to DNase I. When incubated with isolated nuclei, recombinant DNase X was capable of triggering DNA degradation at internucleosomal sites. Similarly to DNase I, the nuclease activity of DNase X was dependent on Ca(2+) and Mg(2+) and inhibited by Zn(2+) ions or chelators of bivalent cations. Overexpression of DNase X caused internucleosomal DNA degradation and induction of cell death associated with increased caspase activation. Despite the presence of two potential caspase cleavage sites, DNase X was processed neither in vitro nor in vivo by different caspases. Interestingly, after initiation of apoptosis DNase X was translocated from the cytoplasm to the nuclear compartment and aggregated as a detergent insoluble complex. Abundant expression of DNase X mRNA was detected in heart and skeletal muscle cells, suggesting that DNase X may be involved in apoptotic or other biological events in muscle tissues. PMID- 10858284 TI - Two substrate binding sites in ascorbate peroxidase: the role of arginine 172. AB - Site-directed mutagenesis has been used to probe the role of Arg172 in ascorbate utilization by ascorbate peroxidase. Arg172 was changed to lysine, glutamine, and asparagine. Although each of these variants retains the ability to utilize guaiacol as a reductant, they exhibit large decreases in their steady-state rates of ascorbate utilization. Spectroscopic, steady-state, and transient-state experiments indicate that these variant proteins are capable of reacting with hydrogen peroxide to form Compound I, but their ability to oxidize ascorbate to form Compound II, and subsequently the resting state, is severely impeded. Results are presented which highlight the importance of Arg172, and a model is proposed to explain its role in ascorbate utilization. PMID- 10858285 TI - The tissue factor region that interacts with substrates factor IX and Factor X. AB - The enzymatic activity of coagulation factor VIIa is controlled by its cellular cofactor tissue factor (TF). TF binds factor VIIa with high affinity and, in addition, participates in substrate interaction through its C-terminal fibronectin type III domain. We analyzed surface-exposed residues in the C terminal TF domain to more fully determine the area on TF important for substrate activation. Soluble TF (sTF) mutants were expressed in E. coli, and their ability to support factor VIIa-dependent substrate activation was measured in the presence of phospholipid vesicles or SW-13 cell membranes. The results showed that factor IX and factor X interacted with the same TF region located proximal to the putative phospholipid surface. According to the degree of activity loss of the sTF mutants, this TF region can be divided into a main region (residues Tyr157, Lys159, Ser163, Gly164, Lys165, Lys166, Tyr185) forming a solvent-exposed patch of 488 A(2) and an extended region which comprises an additional 7-8 residues, including the distally positioned Asn199, Arg200, and Asp204. Some of the identified TF residues, such as Trp158 and those within the loop Lys159 Lys165, are near the factor VIIa gamma-carboxyglutamic acid (Gla) domain, suggesting that the factor VIIa Gla-domain may also participate in substrate interaction. Moreover, the surface identified as important for substrate interaction carries a net positive charge, suggesting that charge interactions may significantly contribute to TF-substrate binding. The calculated surface exposed area of this substrate interaction region is about 1100 A(2), which is approximately half the size of the TF area that is in contact with factor VIIa. Therefore, a substantial portion of the TF surface (3000 A(2)) is engaged in protein-protein interactions during substrate catalysis. PMID- 10858286 TI - A quantitative study of the in vitro binding of the C-terminal domain of p21 to PCNA: affinity, stoichiometry, and thermodynamics. AB - Proliferating cell nuclear antigen (PCNA) plays an essential role in DNA replication, repair, and control of cell proliferation, and its activity can be modulated by interaction with p21(Waf1/Cip1) [Cox, L. S., (1997) Trends Cell Biol. 7, 493-497]. This protein-protein interaction provides a particularly good model target for designing therapeutic agents to treat proliferative disorders such as cancer. In this study, the formation of complexes between PCNA and peptides derived from the C-terminus of p21 has been investigated at the molecular level and quantified using a competitive PCNA binding assay and isothermal titration calorimetry (ITC). The affinity constant for the interaction between p21 (141-160) peptide and PCNA has been determined to be 1.14 x 10(7) M( )(1), corresponding to a K(d) of 87.7 nM. Measurement of the interaction of truncation and substitution analogues based on the p21 (141-160) sequence with PCNA revealed that the N-terminal part (residues 141-152) of the above peptide is the minimum recognition motif, required for PCNA binding. Truncation of the C terminal region p21 (153-160), though, inhibited significantly the ability of the peptides to compete with the full-length p21 (141-160) for binding to PCNA. Alanine mutation of Met 147 or Asp 149 completely abolished or significantly decreased, respectively, the level of the PCNA binding and the inhibition of SV40 DNA replication. Comparison of the data obtained by the competitive PCNA binding assay and the ITC measurements demonstrated the usefulness of this assay for screening for compounds that could modulate the PCNA-p21 interaction. Using this assay, we have screened rationally designed peptides for binding to PCNA and interruption of the PCNA-p21 (141-160) complex. As a result of this screening, we have identified a 16-residue peptide (consensus motif 1 peptide) with the following sequence: SAVLQKKITDYFHPKK. Consensus motif 1 peptide and p21 (141-160) have similar affinities for binding PCNA and abilities to inhibit in vitro replication of DNA originated from SV40. Such peptides could prove useful in assessing p21-mimetic strategies for cancer treatment. PMID- 10858287 TI - Insertion loop 256-268 in coagulation factor IX restricts enzymatic activity in the absence but not in the presence of factor VIII. AB - Insertions in surface loops bordering the substrate-binding groove have been shown to play a major role in the interaction of serine proteases with their cognate inhibitors and substrates. In the present study, we investigated the functional role of factor IX insertion loop 256-268, and in particular of residues Asn(264) and Lys(265) therein. To this end, the purified and activated mutants des-(N264,K265)-FIX and FIX-K265A were compared to normal factor IXa with regard to a number of functional properties. The catalytic efficiency of des (N264,K265)-FIXa and FIXa-K265A toward the amide substrate CH(3)SO(2)-Leu-Gly-Arg pNA was 2-3-fold increased relative to that of normal factor IXa. Comparison of the activities of normal and mutant factor IXa toward a series of closely related amide substrates indicates that mutation of residues Asn(264)-Lys(265) influences the interactions in the S2-binding site. The mutations in loop 256-268 also increased the susceptibility of factor IXa to antithrombin inhibition by approximately 3-fold. Factor X activation experiments in the absence of factor VIIIa revealed that the catalytic efficiency of des-(N264,K265)-FIXa and FIXa K265A was about 20 times higher than that of normal factor IXa. In the presence of factor VIIIa, however, the activity toward factor X was similar to that of normal factor IXa. The reduced sensitivity of the factor IXa mutants to factor VIIIa was neither due to an increase in factor IXa-dependent inactivation of factor VIIIa, nor to a lower affinity for this cofactor. Overall, these data demonstrate that loop 256-268 restricts the activity of factor IXa toward both synthetic and natural substrates. Complex formation with factor VIIIa alleviates the inhibitory effect of this insertion loop on the activation of FX. PMID- 10858288 TI - Conformation of the isolated cepsilon3 domain of IgE and its complex with the high-affinity receptor, FcepsilonRI. AB - Immunoglobulin E (IgE) exhibits a uniquely high affinity for its receptor, FcepsilonRI, on the surface of mast cells and basophils. Previous work has implicated the third domain of the constant region of the epsilon-heavy chain (Cepsilon3) in binding to FcepsilonRI, but the smallest fragment of IgE that is known to bind with full affinity is a covalent dimer of the Cepsilon3 and Cepsilon4 domains. We have expressed the isolated Cepsilon3 in Escherichia coli, measured its affinity for FcepsilonRI, and examined its conformation alone and in the complex with FcepsilonRI. Sedimentation equilibrium in the analytical centrifuge reveals that this product is a monomer. The kinetics of binding to an immobilized fragment of the FcepsilonRI alpha-chain, measured by surface plasmon resonance, yields an affinity constant K(a) = 5 x 10(6) M(-)(1), as compared with 4 x 10(9) M(-)(1) for IgE. The circular dichroism spectrum and measurements of fluorescence as a function of the concentration of a denaturant do not reveal any recognizable secondary structure or hydrophobic core. On binding to the FcepsilonRI alpha-chain fragment, there is no change in the circular dichroism spectrum, indicating that the conformation of Cepsilon3 is unchanged in the complex. Thus the isolated Cepsilon3 domain is sufficient for binding to FcepsilonRI, but with lower affinity than IgE. This may be due to the loss of its native immunoglobulin domain structure or to the requirement for two Cepsilon3 domains to constitute the complete binding site for FcepsilonRI or to a combination of these factors. PMID- 10858289 TI - Tetranectin-binding site on plasminogen kringle 4 involves the lysine-binding pocket and at least one additional amino acid residue. AB - Kringle domains are found in a number of proteins where they govern protein protein interactions. These interactions are often sensitive to lysine and lysine analogues, and the kringle-lysine interaction has been used as a model system for investigating kringle-protein interactions. In this study, we analyze the interaction of wild-type and six single-residue mutants of recombinant plasminogen kringle 4 expressed in Escherichia coli with the recombinant C-type lectin domain of tetranectin and trans-aminomethyl-cyclohexanoic acid (t-AMCHA) using isothermal titration calorimetry. We find that all amino acid residues of plasminogen kringle 4 found to be involved in t-AMCHA binding are also involved in binding tetranectin. Notably, one amino acid residue of plasminogen kringle 4, Arg 32, not involved in binding t-AMCHA, is critical for binding tetranectin. We also find that Asp 57 and Asp 55 of plasminogen kringle 4, which both were found to interact with the low molecular weight ligand with an almost identical geometry in the crystal of the complex, are not of equal functional importance in t-AMCHA binding. Mutating Asp 57 to an Asn totally eliminates binding, whereas the Asp 55 to Asn, like the Arg 71 to Gln mutation, was found only to decrease affinity. PMID- 10858290 TI - Fibrillar amyloid beta-protein binds protease nexin-2/amyloid beta-protein precursor: stimulation of its inhibition of coagulation factor XIa. AB - Cerebrovascular deposition of fibrillar 39-42 amino acid amyloid beta-protein (Abeta), a condition known as cerebral amyloid angiopathy (CAA), is a key pathological feature of Alzheimer's disease and related disorders including hereditary cerebral hemorrhage with amyloidosis-Dutch type (HCHWA-D). Severe cases of CAA, particularly in HCHWA-D, lead to recurrent and often fatal hemorrhagic strokes. Although the reasons for this pathological consequence remain unclear, alterations in proteolytic hemostasis mechanisms have been implicated. For example, the Abeta parent molecule protease nexin-2/amyloid beta protein precursor (PN-2/AbetaPP), which is elevated in HCHWA-D cerebral vessels with Abeta deposits, is a potent inhibitor of coagulation factor XIa (FXIa). Here we show that fibrillar HCHWA-D Abeta binds PN-2/AbetaPP, but not its isolated Kunitz-type proteinase inhibitor (KPI) domain, in a saturable, dose-dependent manner with a K(d) of approximately 28 nM. Neither PN-2/AbetaPP nor its KPI domain bound to nonfibrillar HCHWA-D Abeta. The fibrillar Abeta binding domain on PN-2/AbetaPP was localized to residues 18-119. PN-2/AbetaPP that bound to fibrillar HCHWA-D Abeta immobilized either in plastic wells or on the surface of cultured cerebrovascular smooth muscle cells was active in inhibiting FXIa. Quantitative kinetic measurements revealed that fibrillar HCHWA-D Abeta caused a >5-fold enhancement of FXIa inhibition by PN-2/AbetaPP. Similar stimulatory effects on FXIa inhibition by PN-2/AbetaPP were also observed with fibrillar wild type Abeta. However, fibrillar Abeta had no effect on the inhibition of trypsin by PN-2/AbetaPP. These findings suggest that fibrillar Abeta deposits in cerebral vessels can effectively localize and enhance the anticoagulant functions of PN 2/AbetaPP, thereby contributing to a microenvironment conducive to hemorrhaging. PMID- 10858291 TI - Regulation of amyloid precursor protein (APP) secretion by protein kinase calpha in human ntera 2 neurons (NT2N). AB - Cleavage of amyloid precursor protein (APP) by beta-secretase generates beta amyloid (Abeta), the major component of senile plaques in Alzheimer's disease. Cleavage of APP by alpha-secretase prevents Abeta formation, producing nonamyloidogenic APP products. Protein kinase C (PKC) has been shown to regulate APPs secretion, and PKCalpha and PKCepsilon have been implicated in APPs secretion in fibroblasts. This study examined the PKC isoform involved in regulated APPs secretion in human NT2N neurons and in CHO cells stably expressing APP(695). Inhibition of PMA-induced APPs secretion with the PKC inhibitors Calphostin C and GF109203X demonstrated that PKC is involved in PMA-regulated APPs secretion in NT2N cells. The specific PKC isoforms present in NT2N and CHO695 cells were identified, and PKCalpha and PKCepsilon were found to translocate from cytosol to membranes in NT2N and CHO695 cells. Translocation of PKC to the membrane allows for activation of the enzyme, as well as for positioning of the enzyme close to its substrate. Long-term PMA treatment led to complete downregulation of PKCalpha in NT2N cells and to downregulation of PKCalpha and PKCepsilon in CHO695 cells. PKCalpha downregulation in the NT2N cells resulted in loss of PMA-regulated APPs secretion and a substantial reduction in constitutive APPs secretion. Downregulation of PKCalpha and PKCepsilon in CHO695 cells resulted in loss of PMA-regulated APPs secretion; however, constitutive APPs secretion was unaffected. These findings suggest that PKCalpha is involved in PMA-regulated APPs secretion in NT2N cells and PKCalpha and/or PKCepsilon is involved in PMA-regulated APPs secretion in CHO695 cells. PMID- 10858292 TI - Specific mutagenesis of the rieske iron-sulfur protein in Rhodobacter sphaeroides shows that both the thermodynamic gradient and the pK of the oxidized form determine the rate of quinol oxidation by the bc(1) complex. AB - In the Rieske iron-sulfur protein (ISP) of the ubiquinol:cytochrome c(2) oxidoreductase (bc(1) complex) of Rhodobacter sphaeroides, residue Tyr 156 is located close to the iron-sulfur cluster. Previous studies of the equivalent residue in both Saccharomyces cerevisiae [Denke, E., Merbitz-Zahradnik, T., Hatzfeld, O. M., Snyder, C. H., Link, T. A., and Trumpower, B. L. (1998) J. Biol. Chem. 273, 9085-9093] and Paracoccus denitrificans [Schroter, T., Hatzfeld, O. M., Gemeinhardt, S., Korn, M., Friedrich, T., Ludwig, B. , and Link, T. A. (1998) Eur. J. Biochem. 255, 100-106] have indicated that mutations at this site can lead to modifications in the redox potential of the ISP. To study the effect of similar modifications on the thermodynamic behavior and kinetics of partial reactions of the bc(1) complex upon flash activation, we have constructed four mutant strains of Rb. sphaeroides where Tyr 156 was mutated to His, Leu, Phe, or Trp. The bc(1) complex was assembled and able to support photosynthetic growth in all mutants. Three substitutions (Leu, Phe, Trp) led to alteration of the midpoint potential (E(m)) of the ISP and a slowing in rate of quinol oxidation, suggesting that electron transfer from quinol to the oxidized ISP controls the overall rate and that this step includes the high activation barrier. The Trp mutation led to an increase of approximately 1 pH unit in the pK value of the oxidized ISP. The pH dependence of the rate of quinol oxidation in this mutant was also shifted up by approximately 1 pH unit, showing the importance of the protonation state of the ISP for quinol oxidation. This provides support for a model in which the dissociated form of the oxidized ISP is required for formation of the enzyme-substrate complex [Ugulava, N., and Crofts, A. R. (1998) FEBS Lett. 440, 409-413]. PMID- 10858293 TI - -deltaG(AB) and pH dependence of the electron transfer from P(+)Q(A)(-)Q(B) toP(+)Q(A)Q(B)(-) in Rhodobacter sphaeroides reaction centers. AB - The electron transfer from the reduced primary quinone (Q(A)(-)) to the secondary quinone (Q(B)) can occur in two phases with a well-characterized 100 micros component (tau(2)) and a faster process occurring in less than 10 micros (tau(1)). The fast reaction is clearly seen when the native ubiquinone-10 at Q(A) is replaced with naphthoquinones. The dependence of tau(1) on the free-energy difference between the P(+)Q(A)(-)Q(B) and P(+)Q(A)Q(B)(-) states (-) and on the pH was measured using naphthoquinones with different electrochemical midpoint potentials as Q(A) in Rhodobacter sphaeroides reaction centers (RCs) and in RCs where - is changed by mutation of M265 in the Q(A) site from Ile to Thr (M265IT). Q(B) was ubiquinone (UQ(B)) in all cases. Electron transfer was measured by using the absorption differences of the naphthosemiquinone at Q(A) and the ubisemiquinone at Q(B) between 390 and 500 nm. As - was changed from -90 to -250 meV tau(1) decreased from 29 to 0.2 micros. The free-energy dependence of tau(1) provides a reorganization energy of 850 +/- 100 meV for the electron transfer from Q(A)(-) to Q(B). The slower reaction at tau(2) is free-energy independent, so processes other than electron transfer determine the observed rate. The fraction of the reaction at tau(1) increases with increasing driving force and is 100% of the reaction when - is approximately 100 meV more favorable than in the native RCs with ubiquinone as Q(A). The fast phase, tau(1), is pH independent from pH 6 to 11 while tau(2) slows above pH 9. As the Q(A) isoprene tail length is increased from 2 to 10 isoprene units the fraction at tau(1) decreases. However, tau(1), tau(2), and the fraction of the reaction in each phase are independent of the tail length of UQ(B). PMID- 10858294 TI - Reversible carbon monoxide binding and inhibition at the active site of the Fe only hydrogenase. AB - Carbon monoxide binding and inhibition have been investigated by electron paramagnetic resonance (EPR) spectroscopy in solution and in crystals of structurally described states of the Fe-only hydrogenase (CpI) from Clostridium pasteurianum. Simulation of the EPR spectrum of the as-isolated state indicates that the main component of the EPR spectrum consists of the oxidized state of the "H cluster" and components due to reduced accessory FeS clusters. Addition of carbon monoxide to CpI in the presence of dithionite results in the inhibition of hydrogen evolution activity, and a characteristic axial EPR signal [g(eff(1)), g(eff(2)), and g(eff(3)) = 2.0725, 2.0061, and 2.0061, respectively] was observed. Hydrogen evolution activity was restored by successive sparging with hydrogen and argon and resulted in samples that exhibited the native oxidized EPR signature that could be converted to the reduced form upon addition of sodium dithionite and hydrogen. To examine the relationship between the spectroscopically defined states of CpI and those observed structurally by X-ray crystallography, we have examined the CpI crystals using EPR spectroscopy. EPR spectra of the crystals in the CO-bound state exhibit the previously described axial signal associated with CO binding. The results indicate that the addition of carbon monoxide to CpI results in a single reversible carbon monoxide-bound species characterized by loss of enzyme activity and the distinctive axial EPR signal. PMID- 10858295 TI - Examination of the activity of carboxyl-terminal chimeric constructs of human and yeast ferrochelatases. AB - Insertion of ferrous iron into protoporphyrin IX is catalyzed by ferrochelatase (EC 4.99.1.1). Human and Schizosaccharomyces pombe forms of ferrochelatase contain a [2Fe-2S] cluster with three of the four coordinating cysteine ligands located within the 30 carboxyl-terminal residues. Saccharomyces cerevisiae ferrochelatase contains no cluster, but has comparable activity. Truncation mutants of S. cerevisiae lacking either the last 37 or 16 amino acids have no enzyme activity. Chimeric mutants of human, S. cerevisiae, and Sc. pombe ferrochelatase have been created by switching the terminal 10% of the carboxy end of the enzyme. Site-directed mutagenesis has been used to introduce the fourth cysteinyl ligand into chimeric mutants that are 90% S. cerevisiae. Activity was assessed by rescue of Deltahem H, a ferrochelatase deficient strain of Escherichia coli, and by enzyme assays. UV-visible and EPR spectroscopy were used to investigate the presence or absence of the [2Fe-2S] cluster. Only 2 of the 13 chimeric mutants that were constructed produced active enzymes. HYB, which is predominately human with the last 40 amino acids being that of S. cerevisiae, is an active protein which does not contain a [2Fe-2S] cluster. The other active chimeric mutant, HSp, is predominately human ferrochelatase with the last 38 amino acids being that of Sc. pombe ferrochelatase. This active mutant contains a [2Fe-2S] cluster, as verified by UV-visible and EPR spectroscopic techniques. No other chimeric proteins had detectable enzyme activity or a [2Fe-2S] cluster. The data are discussed in terms of structural requirements for cluster stability and the role that the cluster plays for ferrochelatase. PMID- 10858296 TI - Structural examination of the nickel site in chromatium vinosum hydrogenase: redox state oscillations and structural changes accompanying reductive activation and CO binding. AB - An X-ray absorption spectroscopic study of structural changes occurring at the Ni site of Chromatium vinosum hydrogenase during reductive activation, CO binding, and photolysis is presented. Structural details of the Ni sites for the ready silent intermediate state, SI(r), and the carbon monoxide complex, SI-CO, are presented for the first time in any hydrogenase. Analysis of nickel K-edge energy shifts in redox-related samples reveals that reductive activation is accompanied by an oscillation in the electron density of the Ni site involving formally Ni(III) and Ni(II), where all the EPR-active states (forms A, B, and C) are formally Ni(III), and the EPR-silent states are formally Ni(II). Analysis of XANES shows that the Ni site undergoes changes in the coordination number and geometry that are consistent with five-coordinate Ni sites in forms A, B, and SI(u); distorted four-coordinate sites in SI(r) and R; and a six-coordinate Ni site in form C. EXAFS analysis reveals that the loss of a short Ni-O bond accounts for the change in coordination number from five to four that accompanies formation of SI(r). A shortening of the Ni-Fe distance from 2.85(5) A in form B to 2.60(5) A also occurs at the SI level and is thus associated with the loss of the bridging O-donor ligand in the active site. Multiple-scattering analysis of the EXAFS data for the SI-CO complex reveals the presence of Ni-CO ligation, where the CO is bound in a linear fashion appropriate for a terminal ligand. The putative role of form C in binding H(2) or H(-) was examined by comparing the XAS data from form C with that of its photoproduct, form L. The data rule out the suggestion that the increase in charge density on the NiFe active site that accompanies the photoprocess results in a two-electron reduction of the Ni site [Ni(III) --> Ni(I)] [Happe, R. P., Roseboom, W., and Albracht, S. P. J. (1999) Eur. J. Biochem. 259, 602-608]; only subtle structural differences between the Ni sites were observed. PMID- 10858297 TI - Effects of Mg(2+) on the pre-steady-state kinetics of the biotin carboxylation reaction of pyruvate carboxylase. AB - The effects of Mg(2+) concentration on the kinetics of both ATP cleavage and carboxyenzyme formation in the approach to steady state of the biotin carboxylation reaction of pyruvate carboxylase have been studied. It was found that the enzyme underwent dilution inactivation at low Mg(2+) concentrations and that this occurred at higher enzyme concentrations than had been previously observed. At 10 mM Mg(2+), dilution inactivation was prevented and activation of the enzyme also occurred. When the enzyme was mixed with an ATP solution to initiate the carboxylation reaction, dilution inactivation was reversed and further enzyme activation was induced to a final level that was dependent on Mg(2+) concentration. With the exception of the reaction at 10 mM Mg(2+) in the presence of acetyl CoA, the experimental data could be adequately described as first-order exponential approaches to steady state. At 10 mM Mg(2+) in the presence of acetyl CoA, both ATP cleavage and carboxyenzyme formation data were best described as a biexponential process, in which there was little ATP turnover at steady state. Modeling studies have been performed which produced simulated data that were similar to the experimental data, using a reaction scheme modified from one proposed previously [Legge, G. B., et al. (1996) Biochemistry 35, 3849 3856]. These studies indicate that the major foci of action of Mg(2+) are in the decarboxylation of the enzyme-carboxybiotin complex, the return of the biotin to the site of the biotin carboxylation reaction, and the coupling of ATP cleavage to biotin carboxylation. PMID- 10858298 TI - Role of tyrosine 65 in the mechanism of serine hydroxymethyltransferase. AB - Crystal structures of human and rabbit cytosolic serine hydroxymethyltransferase have shown that Tyr65 is likely to be a key residue in the mechanism of the enzyme. In the ternary complex of Escherichia coli serine hydroxymethyltransferase with glycine and 5-formyltetrahydrofolate, the hydroxyl of Tyr65 is one of four enzyme side chains within hydrogen-bonding distance of the carboxylate group of the substrate glycine. To probe the role of Tyr65 it was changed by site-directed mutagenesis to Phe65. The three-dimensional structure of the Y65F site mutant was determined and shown to be isomorphous with the wild type enzyme except for the missing Tyr hydroxyl group. The kinetic properties of this mutant enzyme in catalyzing reactions with serine, glycine, allothreonine, D and L-alanine, and 5,10-methenyltetrahydrofolate substrates were determined. The properties of the enzyme with D- and L-alanine, glycine in the absence of tetrahydrofolate, and 5, 10-methenyltetrahydrofolate were not significantly changed. However, catalytic activity was greatly decreased for serine and allothreonine cleavage and for the solvent alpha-proton exchange of glycine in the presence of tetrahydrofolate. The decreased catalytic activity for these reactions could be explained by a greater than 2 orders of magnitude increase in affinity of Y65F mutant serine hydroxymethyltransferase for these amino acids bound as the external aldimine. These data are consistent with a role for the Tyr65 hydroxyl group in the conversion of a closed active site to an open structure. PMID- 10858299 TI - Inhibition of p-hydroxyphenylpyruvate dioxygenase by the diketonitrile of isoxaflutole: a case of half-site reactivity. AB - p-Hydroxyphenylpyruvate dioxygenase (HPPD) catalyzes the formation of homogentisate from p-hydroxyphenylpyruvate and molecular oxygen. In plants, this enzyme is the molecular target of new families of very active bleaching herbicides. In the study presented here, we report for the first time on the purification to homogeneity of a plant enzyme, as obtained from recombinant Escherichia coli cells expressing a cDNA encoding carrot HPPD. The purified enzyme allowed us to carry out a detailed characterization of the inhibitory properties of a diketonitile (DKN), the active inhibitor formed from the benzoylisoxazole herbicide isoxaflutole. Inhibition kinetic analyses confirmed that DKN exerts a slow and tight-binding inhibition of HPPD, competitive with respect to the p-hydroxyphenylpyruvate substrate. The stoichiometry of DKN binding to HPPD determined by kinetic analyses or by direct binding of [(14)C]DKN revealed a half-site reactivity of DKN. PMID- 10858300 TI - Kinetics of the initial steps of rabbit psoas myofibrillar ATPases studied by tryptophan and pyrene fluorescence stopped-flow and rapid flow-quench. Evidence that cross-bridge detachment is slower than ATP binding. AB - The kinetics of the tryptophan fluorescence enhancement that occurs when myofibrils (rabbit psoas) are mixed with Mg-ATP were studied by stopped-flow in different solvents (water, 40% ethylene glycol, 20% methanol) at 4 degrees C. Under relaxing conditions (low Ca(2+)) in water (mu = 0.16 M, pH 7.4) and at high ATP concentrations, the transient was biphasic, giving a k(fast)(max) of 230 s( )(1) and a k(slow)(max) of 15 s(-)(1). The kinetics of the two phases were compared with those obtained by chemical sampling using [gamma-(32)P]ATP and quenching in acid (P(i) burst experiments: these give unambiguously the ATP cleavage kinetics), or cold Mg-ATP (cold ATP chase: ATP binding kinetics). k(slow) is due to ATP cleavage, as with S1. Interestingly, k(fast) is slower than the ATP binding kinetics. Instead, this constant appears to report ATP-induced cross-bridge detachment from actin because (1) it was identical to the fluorescence transient obtained on addition of ATP to pyrene-labeled myofibrils; (2) when the initial filament overlap in the myofibrils was decreased, the amplitude of the fast phase decreased; (3) there was no fluorescent enhancement upon the addition of ADP to myofibrils. This is different from the situation with S1 or actoS1 where there was also a fast fluorescent ATP-induced transient but whose kinetics were identical to those of the tight ATP binding. To increase the time resolution and to confirm our results, we also carried out transient kinetics in ethylene glycol and methanol. We interpret our results by a scheme in which a rapid equilibrium between attached (AM.ATP) and detached (M.ATP) states is modulated by the fraction of myosin heads in rigor (AM) during the time of experiment. PMID- 10858301 TI - In vivo regulation of protein synthesis by phosphorylation of the alpha subunit of wheat eukaryotic initiation factor 2. AB - The regulation of protein synthesis is a critical component in the maintenance of cellular homeostasis. A major mechanism of translational control in response to diverse abiotic and biotic stress signals involves the phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha). The pathway has been demonstrated in all eukaryotes except plants, although components of a putative plant pathway have been characterized. To evaluate the in vivo capability of plant eIF2alpha to participate in the translation pathway, we have used vaccinia virus recombinants that constitutively express wheat eIF2alpha and inducibly express the eIF2alpha dsRNA-stimulated protein kinase, PKR, in BSC-40 cells. Activation of PKR in cells expressing wild-type wheat eIF2alpha resulted in an inhibition of cellular and viral protein synthesis and an induction of cellular apoptosis correlating with phosphorylation of eIF2alpha on serine 51. Expression of a nonphosphorylatable mutant (51A) of plant eIF2alpha reversed the PKR mediated translational block as well as the PKR-induced apoptosis. A direct interaction of the plant proteins with the mammalian translational initiation apparatus is supported by coimmunoprecipitation of wild-type plant eIF2alpha and the 51A mutant with mammalian eIF2gamma and the localization of the plant proteins in ribosome fractions. These findings suggest that plant eIF2alpha is capable of interacting with the guanine nucleotide exchange factor eIF2B within the context of the eIF2 holoenzyme and provide direct evidence for its ability to participate in phosphorylation-mediated translational control in vivo. PMID- 10858302 TI - Identification of lipid-accessible sites on the nephrops 16-kDa proteolipid incorporated into a hybrid vacuolar H(+)-ATPase: site-directed labeling with N-(1 Pyrenyl)cyclohexylcarbodiimide and fluorescence quenching analysis. AB - Proton translocation by the vacuolar H(+)-ATPase is mediated by a multicopy transmembrane protein, the 16-kDa proteolipid. It is proposed to assemble in the membrane as a hexameric complex, with each polypeptide comprising four transmembrane helices. The fourth helix of the proteolipid contains an intramembrane acidic residue (Glu140) which is essential for proton translocation and is reactive toward N,N'-dicyclohexylcarbodiimide (DCCD). Current theoretical models of proton translocation by the vacuolar ATPase require that Glu140 should be protonated and in contact with the membrane lipid. In this study we present direct support for this hypothesis. Modification with the fluorescent DCCD analogue N-(1-pyrenyl)cyclohexylcarbodiimide, coupled to fluorescence quenching studies and bilayer depth measurements using the parallax method, was used to probe the position of Glu140 with respect to the bilayer. Glutamate residues were also introduced mutagenically as targets for the fluorescent probe in order to map additional lipid-accessible sites on the 16-kDa proteolipid. These data are consistent with a structural model of the 16-kDa proteolipid oligomer in which the key functional residue Glu140 and discrete faces of the second and third transmembrane helices of the 16-kDa proteolipid are exposed at the lipid-protein interface. PMID- 10858303 TI - Folding character of cytochrome c studied by o-nitrobenzyl modification of methionine 65 and subsequent ultraviolet light irradiation. AB - The protein folding character of cyt c was studied with the use of a photocleavable o-nitrobenzyl derivative of Met65 (NBz-Met65). For the NBz-Met65 cyt c, the Soret absorption band slightly blue shifted compared with the unlabeled cyt c, the 695 nm absorption band related to the Met80 sulfur ligation to the heme iron disappeared, and its resonance Raman spectrum was characteristic of a six-coordinate low-spin species, all characters demonstrating coordination of a non-native ligand, probably a histidine, instead of Met80 to the heme iron. The far-UV circular dichroism (CD) spectrum of cyt c was altered, and the transition midpoint concentration value of guanidine hydrochloride (GdnHCl) for unfolding the protein decreased by 0.9 M by the modification, which showed perturbation of the structure and decrease in protein stability, respectively. With irradiation of 308 nm laser pulses on the NBz-Met65 cyt c, the Soret absorption band slightly red shifted, the 695 nm absorption band appeared, and the CD spectrum shifted toward that of the native protein, which demonstrated recovery of the methionine heme coordination and the native protein structure, due to reconversion of NBz-Met65 to unlabeled methionine. A fast phase was detected as a change in Soret absorbance with a rate constant of 21 000 +/- 4000 s(-)(1) during refolding of cyt c initiated by irradiation of a 308 nm pulse on the NBz-Met65 cyt c in the presence of 2 M GdnHCl. The observed rate constant corresponded well with that reported by the tryptophan fluorescence study [Shastry, M. C. R. S., and Roder, H. (1998) Nat. Struct. Biol. 5, 385-392]. The intermediate decayed with a rate constant of 90 +/- 15, followed by another phase with a rate constant of 13 +/- 3 s(-)(1), and was not seen in the absence of GdnHCl. PMID- 10858304 TI - 13C and (15)N kinetic isotope effects on the reaction of aspartate aminotransferase and the tyrosine-225 to phenylalanine mutant. AB - Heavy atom isotope effects at C-2, C-3, and the amino nitrogen of aspartate were determined for the reaction of porcine heart cytosolic aspartate aminotransferase and the tyrosine-225 to phenylalanine mutant of Escherichia coli aspartate aminotransferase. The effects of deuteration at C-2 of aspartate and of D(2)O on the observed heavy atom isotope effects were determined. The multiple isotope effects support the contribution of C(alpha)-H cleavage, ketimine hydrolysis, and oxaloacetate dissociation to the rate limitation with the wild-type enzyme. The existence of a quinonoid intermediate could not be determined due to the kinetic complexity of the enzyme. For the tyrosine-225 to phenylalanine mutant, we are able to conclude that ketimine hydrolysis is the major rate-determining step. PMID- 10858305 TI - PSF/p54(nrb) stimulates "jumping" of DNA topoisomerase I between separate DNA helices. AB - We have previously shown [Straub et al. (1998) J. Biol. Chem. 273, 26261] that the pyrimidine tract binding protein associated splicing factor PSF/p54(nrb) binds and stimulates DNA topoisomerase I. Here we show that cleavage and religation half-reactions of topoisomerase I are unaffected by PSF/p54(nrb), whereas the propensity of the enzyme to jump between separate DNA helices is stimulated. To demonstrate such an effect, topoisomerase I was first captured in suicidal cleavage of an oligonucleotide substrate. Subsequently, a cleavage/ligation equilibrium was established by adding a ligation donor under conditions allowing recleavage of the ligated substrate. Finally, a second oligonucleotide was added to the mixture, which also allowed suicidal cleavage by topoisomerase I, but did not accommodate the ligation donor of the first oligonucleotide. Thus, topoisomerase I was given the choice to engage in repeated cleavage/ligation cycles of the first oligonucleotide or to jump to the second suicide substrate and get trapped. PSF/p54(nrb) enhanced the cleavage rate of the second oligonucleotide (11-fold), suggesting that it stimulates the dissociation of topoisomerase I after ligation. Thus, stimulation of topoisomerase I catalysis by PSF/p54(nrb) seems to be affected by mobilization of the enzyme. PMID- 10858306 TI - Base excision repair is impaired in mammalian cells lacking Poly(ADP-ribose) polymerase-1. AB - In mammalian cells, damaged bases in DNA are corrected by the base excision repair pathway which is divided into two distinct pathways depending on the length of the resynthesized patch, replacement of one nucleotide for short-patch repair, and resynthesis of several nucleotides for long-patch repair. The involvement of poly(ADP-ribose) polymerase-1 (PARP-1) in both pathways has been investigated by using PARP-1-deficient cell extracts to repair single abasic sites derived from uracil or 8-oxoguanine located in a double-stranded circular plasmid. For both lesions, PARP-1-deficient cell extracts were about half as efficient as wild-type cells at the polymerization step of the short-patch repair synthesis, but were highly inefficient at the long-patch repair. We provided evidence that PARP-1 constitutively interacts with DNA polymerase beta. Using cell-free extracts from mouse embryonic cells deficient in DNA polymerase beta, we demonstrated that DNA polymerase beta is involved in the repair of uracil derived AP sites via both the short and the long-patch repair pathways. When both PARP-1 and DNA polymerase beta were absent, the two repair pathways were dramatically affected, indicating that base excision repair was highly inefficient. These results show that PARP-1 is an active player in DNA base excision repair. PMID- 10858307 TI - Characterization of sequence-specific DNA binding by the transcription factor Oct 1. AB - The DNA-binding domain of the Oct-1 transcription factor, POU, recognizes a defined DNA sequence known as the octamer element to regulate the expression of both general and cell-type-specific genes. The two-part DNA-binding domain partially encircles the DNA to recognize the eight base pairs of the octamer element. We have characterized the binding of Oct-1/POU to an octamer element using isothermal titration calorimetry. As found for other cognate protein/DNA complexes, the formation of the Oct-1 POU/DNA complex is associated with a large negative heat capacity change, DeltaC(p)()(, obs). However, the observed change is much greater than expected by empirical relationships with buried surface area. Supported by data from proteolysis studies on the free and DNA-bound protein, we propose that the discrepancy in heat capacity arises principally from the partial folding of the Oct-1 POU protein upon complex formation. Formation of the Oct-1 POU/DNA complex is strongly dependent on ionic strength, and the detailed quantification of this relationship suggests that six charged contacts are made between the protein and the phosphate groups of the DNA. This agrees with observations from the crystal structure of an Oct-1 POU/DNA complex. PMID- 10858308 TI - Inhibition of carboxypeptidase A by D-penicillamine: mechanism and implications for drug design. AB - Zinc metalloprotease inhibitors are usually designed to inactivate the enzyme by forming a stable ternary complex with the enzyme and active-site zinc. D-Cysteine inhibits carboxypeptidase, ZnCPD, by forming such a complex, with a K(i) of 2.3 microM. In contrast, the antiarthritis drug D-penicillamine, D-PEN, which differs from D-Cys only by the presence of two methyl groups on the beta-carbon, inhibits ZnCPD by promoting the release of the active-site zinc. We have given the name catalytic chelator to such inhibitors. Inhibition is a two-step process characterized by formation of a complex with the enzyme (K(i(initial)) = 1.2 mM) followed by release of the active-site zinc at rates up to 420-fold faster than the spontaneous release. The initial rate of substrate hydrolysis at completion of the second step also depends on D-PEN concentration, reflecting formation of a thermodynamic equilibrium governed by the stability constants of chelator and apocarboxypeptidase for zinc (K(i(final)) = 0.25 mM). The interaction of D-PEN and D-Cys with the active-site metal has been examined by replacing the active site zinc by a chromophoric cobalt atom. Both inhibitors perturb the d-d transitions of CoCPD in the 500-600 nm region within milliseconds of mixing but only the CoCPD.D-Cys complex displays a strong S --> Co(II) charge-transfer band at 340 nm indicative of a metal-sulfur bond. While the D-Cys complex is stable, the CoCPD.D-PEN complex breaks down to apoenzyme and Co(D-PEN)(2) with a half life of 0.5 s. D-PEN is the first drug found to inhibit a metalloprotease by increasing the dissociation rate constant of the active-site metal. The ability of D-PEN to catalyze metal removal from carboxypeptidase A and other zinc proteases suggests a possible mechanism of action in arthritis and Wilson's disease and may also underlie complications associated with its clinical use. PMID- 10858309 TI - Reassessment of the active site quino-cofactor proposed to occur in the Aspergillus niger amine oxidase AO-I from the properties of model compounds. AB - Quino-cofactors have been found in a wide variety of prokaryotic and eukaryotic organisms. Two variants have, thus far, been demonstrated to derive from tyrosine precursors: these are the 2,4, 5-trihydroxyphenylalanine quinone (topa quinone or TPQ) [Janes, S. M. , et al. (1990) Science 248, 98] and an o-quinone analogue containing the side chain of a lysine residue (lysyltyrosine quinone or LTQ) [Wang, S. Z., et al. (1996) Science 273, 1078]. Additionally, a third variant of the family of tyrosine-derived cofactors has been reported to exist in an Aspergillus niger amine oxidase AO-I. This was described as an o-quinone cross linked to the side chain of a glutamate residue [Frebort, I. (1996) Biochim. Biophys. Acta 1295, 59]. We have synthesized model compounds related to the proposed structure. Characterization of the redox properties for the model compound and spectral properties of its 4-nitrophenylhydrazine derivative lead us to conclude that the cofactor in A. niger amine oxidase AO-I has been misidentified. A TPQ carboxylate ester is considered an unlikely candidate for a biologically functional quino-cofactor. PMID- 10858310 TI - A novel delta(3),delta(2)-enoyl-CoA isomerase involved in the biosynthesis of the cyclohexanecarboxylic acid-derived moiety of the polyketide ansatrienin A. AB - The side chain of the antifungal polyketide ansatrienin A produced by Streptomyces collinus contains a cyclohexanecarboxylic acid (CHC) derived moiety. This CHC in the coenzyme A activated form (CHC-CoA) is derived from shikimic acid via a pathway in which the penultimate step is the isomerization of 2 cyclohexenylcarbonyl-CoA to 1-cyclohexenylcarbonyl-CoA. We have purified a 28 kDa 2-cyclohexenylcarbonyl-CoA isomerase (ChcB) from S. collinus and cloned and sequenced the corresponding chcB gene. The predicted amino acid sequence of ChcB showed moderate sequence identity to members of the hydratase/isomerase superfamily of enzymes. The recombinant ChcB was overexpressed in Escherichia coli and purified to homogeneity using metal chelate chromatography. Kinetic analysis demonstrated that recombinant ChcB had wide substrate specificity and could catalyze a double bond isomerization using 2-cyclohexenylcarbonyl-CoA (K(m) 116 +/- 68 microM, k(cat)( )()3.7 +/- 1.0 min(-)(1)), trans-3-hexenyl-CoA (K(m) 39 +/- 10 microM, k(cat)( )()12.8 +/- 1 min(-)(1)), and vinylacetyl-CoA (K(m) 156 +/- 34 microM, k(cat)( )()29 +/- 3 min(-)(1)) as substrates. ChcB activity in cell extracts of S. collinus SP1, an insertionally disrupted chcB mutant, was shown to decrease by more than 99% (as compared to the wild-type strain) using all three of these substrates. The S. collinus SP1 strain, unlike the wild-type strain, could not produce omega-cyclohexyl fatty acids but was still able to grow efficiently on methyl oleate as a sole carbon source. These observations demonstrate that the S. collinus ChcB is required for catalyzing the isomerization of 2-cyclohexenylcarbonyl-CoA to 1-cyclohexenylcarbonyl-CoA during CHC-CoA biosynthesis but not for degradation of unsaturated fatty acids. The chcB gene does not appear to be associated with the ansatrienin biosynthetic gene cluster, which has previously been shown to contain at least one gene known to be essential for CHC-CoA biosynthesis. This finding represents a notable exception to the general rule regarding the clustering of polyketide biosynthetic pathway genes. PMID- 10858311 TI - DNA bending is a determinant of calicheamicin target recognition. AB - Calicheamicin is a hydrophobic enediyne antibiotic that binds noncovalently to DNA and causes sequence-selective oxidation of deoxyribose. While the drug makes several base contacts along the minor groove, the diversity of binding-site sequences and the effects of DNA conformation on calicheamicin-induced DNA cleavage suggest that sequence recognition per se is not the primary determinant of target selection. We now present evidence that calicheamicin bends its DNA targets. Using a gel mobility assay, we observed that polymers of oligonucleotide constructs containing AGGA and ACAA binding sites for calicheamicin did not possess intrinsic curvature. Binding of calicheamicin epsilon, the aromatized form of the parent calicheamicin gamma(1)(I), to oligonucleotide constructs containing binding sites in phase with the helical repeat caused a shift to smaller circle sizes in T4 ligase-mediated circle formation assays, with a much smaller shift observed with constructs containing out-of-phase binding sites. It was also observed that binding of calicheamicin epsilon to a 273 bp construct with phased binding sites caused an increase in the molar cyclization factor, J, from 8 x 10(-8) to 9 x 10(-6) M. These results are consistent with DNA bending as part of an induced-fit mechanism of DNA target recognition and with the hypothesis that the preferred targets of calicheamicin, the 3' ends of oligopurine tracts, are characterized by unique conformational properties. PMID- 10858312 TI - Multiple structures for the 2-aminopurine-cytosine mispair. AB - The base pair formed between 2-aminopurine (2AP) and cytosine (C) is an intermediate in transition mutations generated by 2AP. To date, several structures have been proposed for the 2AP-C mispair, including those involving a rare tautomer, a protonated base pair, and a neutral wobble structure. In this paper, we describe a series of UV, fluorescence, and NMR studies which demonstrate that an equilibrium exists between the neutral wobble and the protonated Watson-Crick structures. The apparent pK value for the transition between the structures is 5.9-6.0. Formation of a Watson-Crick base pair is accomplished predominantly by protonation of the 2AP residue as indicated by UV spectral changes, fluorescence quenching, and changes in proton chemical shifts. Rapid transfer of the shared proton between the 2AP and cytosine residues is indicated by the rapid exchange of the cytosine amino protons of the protonated Watson-Crick configuration. The relative contribution of the neutral wobble and protonated Watson-Crick configurations to 2AP-induced transition mutations is discussed. PMID- 10858313 TI - Hydration changes implicated in the remarkable temperature-dependent membrane permeation of cyclosporin A. AB - Cyclosporin A is a cyclic peptide believed to exist as multiple conformers in aqueous solution. Two major conformations, distinguished by a single cis-trans isomerization and the presence of four either intramolecular or intermolecular hydrogen bonds, have been confirmed depending on whether CsA is characterized in organic solvents or bound in aqueous complex with cyclophilin. The relationship between CsA conformation and its ability to penetrate biological membranes is currently unknown. Using Caco-2 cell monolayers, we documented a remarkable increase (more than 2 orders of magnitude) in the membrane permeation of the peptide as temperature was increased from 5 to 37 degrees C. The solubility of CsA was 72 microM at 5 degrees C, but decreased by more than an order of magnitude at 37 degrees C. Moreover, CsA partitioned into non-hydrogen bond donating solvents linearly as a function of increasing temperature, suggestive of a significant conformational change. However, while NMR spectra of CsA confirmed the previously predicted presence of multiple conformers in aqueous solution, the equilibrium between the two major species was not affected by changes in temperature. These NMR data indicated that the observed temperature-dependent changes in the membrane permeability of CsA do not originate from changes in the peptide backbone conformation. Sedimentation equilibrium analysis revealed that CsA behaves in a highly nonideal manner over the temperature range tested. We interpret this behavior as a change in the hydration state with a smaller (or weaker) hydration shell surrounding the peptide at higher temperatures. Such a change would result in lower peptide desolvation energy, thereby promoting partitioning into cellular membranes. We contend that changes in membrane penetration result from alterations in the hydration state of CsA and are not related to the interconversion of the defined conformations. PMID- 10858314 TI - p50(cdc37) is a nonexclusive Hsp90 cohort which participates intimately in Hsp90 mediated folding of immature kinase molecules. AB - Hsp90 and p50(cdc37) provide a poorly understood biochemical function essential to certain protein kinases, and recent models describe p50(cdc37) as an exclusive hsp90 cohort which links hsp90 machinery to client kinases. We describe here the recovery of p50(cdc37) in immunoadsorptions directed against the hsp90 cohorts FKBP52, cyp40, p60HOP, hsp70, and p23. Additionally, monoclonal antibodies against FKBP52 coadsorb maturation intermediates of the hsp90-dependent kinases p56(lck) and HRI, and the presence of these maturation intermediates significantly increases the representation of p50(cdc37) and hsp90 on FKPB52 machinery. Although the native heterocomplex between hsp90 and p50(cdc37) is salt labile, their dynamic interactions with kinase substrates produce kinase chaperone heterocomplexes which are highly salt-resistant. The hsp90 inhibitor geldanamycin does not directly disrupt the native association of hsp90 with p50(cdc37) per se, but does result in the formation of salt-labile hsp90-kinase heterocomplexes which lack the p50(cdc37) cohort. We conclude that p50(cdc37) does not simply serve as a passive structural bridge between hsp90 and its kinase substrates; instead, p50(cdc37) is a nonexclusive hsp90 cohort which responds to hsp90's nucleotide-regulated conformational switching during the generation of high-affinity interactions within the hsp90-kinase-p50(cdc37) heterocomplex. PMID- 10858315 TI - Predicted exocyclic amino group alkylation of 2'-deoxyadenosine and 2' deoxyguanosine by the isopropyl cation. AB - Diisopropyltriazene in aqueous 10% acetonitrile (pH 7.0 +/- 0.4) yields N(6) isopropyl-2'-deoxyAdo as the predominant product and N(2)-isopropyl-2'-deoxyGuo in yields comparable with the O(6) adduct in reactions with 2'-deoxyAdo and 2' deoxyGuo, respectively. These observations are inconsistent with what is expected on the basis of the regnant hypothesis concerning factors that determine atom site selectivity in diazonium ion-mediated alkylations. An alternative explanation based on the fleeting existence of the reactive intermediates involved is consistent with these observations. PMID- 10858316 TI - A quantitative structure--activity relationships model for the acute toxicity of substituted benzenes to Tetrahymena pyriformis using Bayesian-regularized neural networks. AB - We have used a new, robust structure-activity mapping technique, a Bayesian regularized neural network, to develop a quantitative structure-activity relationships (QSAR) model for the toxicity of 278 substituted benzenes toward Tetrahymena pyriformis. The independent variables used in the modeling were derived solely from the molecular structure, and the model was tested on 20% of the data set selected from the whole set by cluster analysis and which had not been used in training the network. The results show that the method is robust and reliable and give results for mixed class compounds which are comparable to earlier QSAR work on single-chemical class subsets of the 278 compounds and which employed measured physicochemical parameters as independent variables. Comparisons of Bayesian neural net models with those derived by classical PLS analysis showed the superiority of our method. The method appears to be able to model more diverse chemical classes and more than one mechanism of toxicity. PMID- 10858317 TI - Quantitative structure-activity analysis of the algae toxicity of nitroaromatic compounds. AB - Proliferation toxicity toward the algae Scenedesmus vacuolatus in a 24 h one generation reproduction assay was determined for nitrobenzene and 18 derivatives, including two phenols. The resultant EC(50) values covering more than 4 orders of magnitude were subjected to a quantitative structure-activity analysis (QSAR) using hydrophobicity in terms of the octanol/water partition coefficient in logarithmic form, log K(ow), and 16 quantum chemical descriptors of molecular reactivity that were calculated with the AM1 scheme. For 13 mononitro derivatives and the highly hydrophobic trifluralin, a narcotic-type mode of action can explain most of the toxicity variation. Correction of log K(ow) for ionization for the phenols and quantification of the molecular susceptibility for one electron reduction as apparently rate-determining biotransformation step by the energy of the lowest unoccupied molecular orbital, E(LUMO), yields a highly significant QSAR for all 19 compounds (r(adj)(2) = 0.90), which can be further improved when adding the maximum net atomic charge at the nitro nitrogen, q(nitro)(-)(N), as the third descriptor (r(adj)(2) = 0.93). Comparison of the energy of the singly occupied molecular orbital, E(SOMO), of the radical anions as initial metabolites with the E(SOMO) of known redox cyclers suggests that dinitrobenzenes and TFM as well as multiply chlorinated nitrobenzenes may also exert oxidative stress. This is based on an E(SOMO) window of -0.30 to 0. 55 eV as a tentative criterion for molecular structures to have the potential for redox cycling, derived from a set of eight known redox cyclers. The discussion includes a detailed analysis of apparently relevant metabolic pathways and associated modes of toxic action of nitroaromatics. PMID- 10858318 TI - Carbon dioxide but not bicarbonate inhibits N-nitrosation of secondary amines. Evidence for amine carbamates as protecting entities. AB - Hydrogen carbonate (bicarbonate, HCO(3)(-)) has been proposed to accelerate the decomposition of N(2)O(3) because N-nitrosation of morpholine via a nitric oxide/oxygen mixture ((*)NO/O(2)) was inhibited by the addition of HCO(3)(-) at pH 8.9 [Caulfield, J. L., Singh, S. P., Wishnok, J. S., Deen, W. M., and Tannenbaum, S. R. (1996) J. Biol. Chem. 271, 25859-25863]. In the study presented here, it is shown that carbon dioxide (CO(2)) is responsible for this kind of protective effect because of formation of amine carbamates, whereas an inhibitory function of HCO(3)(-) is excluded. N-Nitrosation of morpholine (1-10 mM) at pH 7.4-7.5 by the (*)NO-donor compounds PAPA NONOate and MAMA NONOate (0.5 mM each) was not affected by the presence of large amounts of HCO(3)(-) (up to 100 mM) in aerated aqueous solution. Similar results were obtained by replacing the (*)NO donor compounds with authentic (*)NO (900 microM). In agreement with data from the study cited above, (*)NO/O(2)-mediated formation of N-nitrosomorpholine (NO Mor) was indeed inhibited by about 45% in the presence of 50 mM HCO(3)(-) at pH 8.9. However, 500 MHz (13)C NMR analysis with (13)C-enriched bicarbonate revealed that significant amounts of morpholine carbamate are formed from reaction of equilibrated CO(2) with morpholine (1-100 mM) at pH 8.9, but only to a minor extent at pH 7. 5. The protective effect of morpholine carbamate formation is explained by a significantly reduced charge density at nitrogen. This view is supported by the results of density functional theory/natural population analysis, i.e., quantumchemical calculations for morpholine and morpholine carbamate. In agreement with its lower pK(a), another secondary amine, piperazine, had already produced significant amounts of piperazine carbamate at pH 7. 4 as shown by (13)C NMR spectrometry. Consequently, and in contrast to morpholine, N-nitrosation of piperazine (2 mM) by both (*)NO/O(2) (PAPA NONOate, 0.5 mM) and the (*)NO/O(2)(-)(*)-releasing compound SIN-1 (1 mM) was inhibited by about 66% in the presence of 200 mM HCO(3)(-). PMID- 10858319 TI - Conformation and proton configuration of pyrimidine deoxynucleoside oxidation damage products in water. AB - Emerging data strongly suggest that the oxidation of DNA bases can contribute to genomic instability. Structural changes to DNA, induced by base oxidation, may reduce the fidelity of DNA replication and interfere with sequence-specific DNA protein interactions. We have examined the structures of a series of pyrimidine deoxynucleoside oxidation damage products in aqueous solution. The modified nucleosides studied include the deoxynucleoside derivatives of 5-hydroxyuracil, 5 hydroxycytosine, 5-(hydroxymethyl)uracil, 5-(hydroxymethyl)cytosine, 5 formyluracil, and 5-formylcytosine. The influence of base oxidation on ionization constants, sugar conformation, and tautomeric configuration has been determined on the basis of UV, proton, and nitrogen NMR spectra of the (15)N-enriched derivatives. The potential biological consequences of the structural perturbations resulting from base oxidation are discussed. PMID- 10858320 TI - Chemometric models for toxicity classification based on NMR spectra of biofluids. AB - 1H NMR spectroscopic and pattern recognition (PR)-based methods were used to investigate the biochemical variability in urine obtained from control rats and from rats treated with a hydrazine (a model hepatotoxin) or HgCl(2) (a model renal cortical toxin). The 600 MHz (1)H NMR spectra of urine samples obtained from vehicle- or toxin-treated Han-Wistar (HW) and Sprague-Dawley (SD) rats were acquired, and principal components analysis (PCA) and soft independent modeling of class analogy (SIMCA) analysis were used to investigate the (1)H NMR spectral data. Variation and strain differences in the biochemical composition of control urine samples were assessed. Control urine (1)H NMR spectra obtained from the two rat strains appeared visually similar. However, chemometric analysis of the control urine spectra indicated that HW rat urine contained relatively higher concentrations of lactate, acetate, and taurine and lower concentrations of hippurate than SD rat urine. Having established the extent of biochemical variation in the two populations of control rats, PCA was used to evaluate the metabolic effects of hydrazine and HgCl(2) toxicity. Urinary biomarkers of each class of toxicity were elucidated from the PC loadings and included organic acids, amino acids, and sugars in the case of mercury, while levels of taurine, beta-alanine, creatine, and 2-aminoadipate were elevated after hydrazine treatment. SIMCA analysis of the data was used to build predictive models (from a training set of 416 samples) for the classification of toxicity type and strain of rat, and the models were tested using an independent set of urine samples (n = 124). Using models constructed from the first three PCs, 98% of the test samples were correctly classified as originating from control, hydrazine-treated, or HgCl(2)-treated rats. Furthermore, this method was sensitive enough to predict the correct strain of the control samples for 79% of the data, based upon the class of best fit. Incorporation of these chemometric methods into automated NMR based metabonomics analysis will enable on-line toxicological assessment of biofluids and will provide a tool for probing the mechanistic basis of organ toxicity. PMID- 10858321 TI - Structure of adduct X, the last unknown of the six major DNA adducts of mitomycin C formed in EMT6 mouse mammary tumor cells. AB - Treatment of EMT6 mouse mammary tumor cells with mitomycin C (MC) results in the formation of six major MC-DNA adducts. We identified the last unknown of these ("adduct X") as a guanine N(2) adduct of 2, 7-diaminomitosene (2,7-DAM), in which the mitosene is linked at its C-10 position to guanine N(2). The assigned structure is based on UV and mass spectra of adduct X isolated directly from the cells, as well as on its difference UV, second-derivative UV, and circular dichroism spectra, synthesis from [8-(3)H]deoxyguanosine, and observation of its heat stability. These tests were carried out using 17 microg of synthetic material altogether. The mechanism of formation of adduct X involves reductive metabolism of MC to 2,7-DAM, which undergoes a second round of reductive activation to alkylate DNA, yielding adduct X and another 2,7-DAM-guanine adduct (adduct Y), which is linked at guanine N7 to the mitosene. Adduct Y has been described previously. Adduct X is formed preferentially at GpC, while adduct Y favors the GpG sequence. In contrast to MC-DNA adducts, the 2,7-DAM-DNA adducts are not cytotoxic. PMID- 10858322 TI - Pure-silica zeolites (Porosils) as model solids for the evaluation of the physicochemical features determining silica toxicity to macrophages. AB - The interaction between inhaled particles and alveolar macrophages plays a key role in silica-related diseases. It has been previously shown [Fubini, B., et al. (1999) Chem. Res. Toxicol. 12, 737-745] that a monocyte-macrophage cell line (J774) may be employed in the evaluation of the degree of cytotoxicity to alveolar macrophages of various silica dusts. In this paper, pure-silica zeolites (porosils) in microcrystalline form have been employed as "model solids" in an effort to show which physicochemical properties of the silica particle are playing a major role in the toxicity to macrophages. The samples employed covered four different porosil crystal structures (MFI, FAU, TON, and MTT) and also include a synthetic rodlike cristobalite (CRIS-rd). When compared at equal weight, the samples cover a wide range of cytotoxicity from inert to toxic as unheated mineral cristobalite [Fubini, B., et al. (1999) Chem. Res. Toxicol. 12, 737-745]. Mild grinding did not affect cytotoxicity. Calcined (open pores) and uncalcined (pore filled with template) TON exhibited the same cytotoxicity, indicating that only the outer surface is implied. The hydrophobic and/or hydrophilic character of TON, evaluated by adsorption calorimetry, is close to what has been previously found for silicalite and is consistent with a hydrophilic outer surface and hydrophobic pore walls. The potential for generating hydroxyl radicals from hydrogen peroxide varies among the various porosils that have been studied. A model is proposed for the correlation between inhibition of growth on proliferating cells and physicochemical properties varying from one to the other sample. The extent of external surface and the aspect ratio were related to the intensity of the cytotoxic effect, while the level of radical release was not. This suggests, on one hand, that comparison of toxicity among various dusts should be made at equal particle surface and, on the other, that in the model studied, free radical release does not play a crucial role in the primary event of toxicity to alveolar macrophages. PMID- 10858323 TI - Modification of bovine serum albumin structure following reaction with 4,5(E) epoxy-2(E)-heptenal. AB - Bovine serum albumin (BSA) was incubated for different periods of time and in the presence of several concentrations of 4, 5(E)-epoxy-2(E)-heptenal, at pH 7.4 and 37 degrees C, in an effort to analyze the changes produced in its structure as a consequence of its reaction with this product of lipid oxidation. The epoxyalkenal modified the primary structure of BSA as determined by lysine losses and formation of oxidative stress product epsilon-N-pyrrolylnorleucine (Pnl), which depended on the concentration of the aldehyde and the incubation time. These changes also modified secondary and tertiary structures of the protein, which were determined by studying protein denaturation and polymerization. In addition, all these modifications were parallel to the development of color and fluorescence, which were produced as a consequence of the formation and polymerization of pyrrole amino acid residues. The above results indicated that epoxyalkenals modify the protein structure and develop color and fluorescence. A failure in the degradation of these modified proteins might induce their accumulation and, thus, participation in lipofuscin or age pigments formation. PMID- 10858324 TI - Mass spectrometric measurement of formaldehyde generated in breast cancer cells upon treatment with anthracycline antitumor drugs. AB - Selected ion flow tube-chemical ionization mass spectrometry was used to measure formaldehyde levels in human breast cancer cells in comparison with levels in cells treated with the antitumor drugs doxorubicin (DOX) and daunorubicin (DAU) and the daunorubicin-formaldehyde conjugate Daunoform (DAUF). The measurement was performed on cell lysates and showed only background levels of formaldehyde in untreated cells and drug-treated resistant cells (MCF-7/Adr cells) but levels above background in DOX- and DAU-treated sensitive cells (MCF-7 cells). The level of formaldehyde above background was a function of drug concentration (0.5-50 microM), treatment time (3-24 h), cell density (0.3 x 10(6) to 7 x 10(6) cells/mL), and cell viability (0-100%). Higher levels of formaldehyde were observed in lysates of MCF-7 cells treated at higher drug levels, unless the treatment resulted in low cell viability. Elevated levels were directly related to cell density and were observed even with 0.5 microM drug. A lower limit for excess formaldehyde in MCF-7 cells treated with 0.5 microM DAU for 24 h is 0.3 mM. Control experiments showed that formaldehyde was not produced after cell lysis. Lysates of sensitive and resistant cells treated with 0.5 micromolar equiv of the formaldehyde conjugate (DAUF) for 3 h showed only background levels of formaldehyde. The results support a mechanism for drug cytotoxicity which involves drug induction of metabolic processes leading to formaldehyde production followed by drug utilization of formaldehyde to virtually cross-link DNA. PMID- 10858325 TI - Acrylamide: a cooking carcinogen? AB - Exposure to acrylamide (AA) has been monitored by mass spectrometric detection of the adduct, N-(2-carbamoylethyl)valine (CEV), to the N-termini of hemoglobin (Hb), according to the N-alkyl Edman method. In these studies, a conspicuous background level, about 40 pmol/g of globin, of apparently the same adduct was regularly observed in Hb from persons without known exposure to AA. For testing of the hypothesis that this adduct originates from AA formed in cooking, rats were fed fried animal standard diet for 1 or 2 months. These animals exhibited a strong increase of the level of the studied Hb adduct, compared to control rats fed unfried diet. By gas chromatography/tandem mass spectrometry, the identity with CEV was confirmed by the concordance of the product ion spectrum of the studied adduct with that of a verified standard and by interpretation of the fragment ions. Further support of the chemical structure, at the same time pinpointing AA as the causative reactive factor, was obtained through the demonstration that AA is formed in the heating of the feed and that the level of AA in the fried feed is compatible with the measured levels of the CEV adduct. The raised CEV adduct levels observed in experimental animals are of a magnitude that is similar to the background level in nonsmoking humans. These data render it likely that cooking of food is a major source of the background dose of AA also in humans. An evaluation of cancer tests of AA and available data for its metabolism leads to the estimation that the background dose of AA is associated with a considerable cancer risk. PMID- 10858326 TI - Mutagenicity of the 1-nitropyrene-DNA adduct N-(deoxyguanosin-8-yl)-1-aminopyrene in Escherichia coli located in a nonrepetitive CGC sequence. AB - 1-Nitropyrene, a common environmental pollutant, forms a major DNA adduct, N (deoxyguanosin-8-yl)-1-aminopyrene (dG(AP)). Mutational spectra of randomly introduced dG(AP) in Escherichia coli included many different types of mutations. However, a prior site-specific study in a CGCG(AP)CG sequence showed only CpG deletions and +1 frame shifts. To further explore the context effects of dG(AP) in mutagenesis, in this work this adduct was incorporated into a nonrepetitive CGC sequence in single-stranded M13mp7L2 DNA. Upon replication of this construct in repair-competent E. coli, one-base deletions and base substitutions were detected. The -1 frame shifts, whose frequency increased 3-6-fold with SOS (to an average frequency of 1.5%), involved deletion of the adjacent C residues. The base substitutions ( approximately 2.2%) included targeted G-to-T and G-to-C transversions, whose frequencies did not increase with SOS. This suggests that dG(AP) mutagenesis is highly dependent on the local DNA sequence. PMID- 10858327 TI - Rat liver cytosol catalyzes a reaction involving activated N-nitrosodimethylamine and a carbohydrate from the pentose phosphate pathway volume 13, number 2, february 2000, pp 126-133 PMID- 10858328 TI - The problems of using the transmission/disequilibrium test to infer tight linkage. AB - Family-based association methods such as the transmission/disequilibrium test (TDT) have become very popular during the past few years, often being preferred to case-control studies because family-based approaches avoid the difficulties of ascertainment of appropriate populations of cases and controls for case-control studies. Significant TDT results indicate both linkage and allelic association. However, significant TDT results are often interpreted as implying tight linkage of marker and disease locus, and we shall argue here that, in general, this interpretation is not justified. PMID- 10858329 TI - Mutation frequencies and antibiotic resistance. PMID- 10858330 TI - Stenotrophomonas maltophilia D457R contains a cluster of genes from gram-positive bacteria involved in antibiotic and heavy metal resistance. AB - A cluster of genes involved in antibiotic and heavy metal resistance has been characterized from a clinical isolate of the gram-negative bacterium Stenotrophomonas maltophilia. These genes include a macrolide phosphotransferase (mphBM) and a cadmium efflux determinant (cadA), together with the gene cadC coding for its transcriptional regulator. The cadC cadA region is flanked by a truncated IS257 sequence and a region coding for a bin3 invertase. Despite their presence in a gram-negative bacterium, these genetic elements share a common gram positive origin. The possible origin of these determinants as a remnant composite transposon as well as the role of gene transfer between gram-positive and gram negative bacteria for the acquisition of antibiotic resistance determinants in chronic, mixed infections is discussed. PMID- 10858331 TI - In vitro selection of mutations in the human immunodeficiency virus type 1 reverse transcriptase that decrease susceptibility to (-)-beta-D-dioxolane guanosine and suppress resistance to 3'-azido-3'-deoxythymidine. AB - Human immunodeficiency virus type 1 (HIV-1) isolates resistant to (-)-beta-D dioxolane-guanosine (DXG), a potent and selective nucleoside analog HIV-1 reverse transcriptase (RT) inhibitor, were selected by serial passage of HIV-1(LAI) in increasing drug concentrations (maximum concentration, 30 microM). Two independent selection experiments were performed. Viral isolates for which the DXG median effective concentrations (EC(50)s) increased 7.3- and 12.2-fold were isolated after 13 and 14 passages, respectively. Cloning and DNA sequencing of the RT region from the first resistant isolate identified a K65R mutation (AAA to AGA) in 10 of 10 clones. The role of this mutation in DXG resistance was confirmed by site-specific mutagenesis of HIV-1(LAI). The K65R mutation also conferred greater than threefold cross-resistance to 2',3'-dideoxycytidine, 2', 3'-dideoxyinosine, 2',3'-dideoxy-3'-thiacytidine, 9-(2 phosphonylmethoxyethyl)adenine, 2-amino-6-chloropurine dioxolane, dioxolanyl-5 fluorocytosine, and diaminopurine dioxolane but had only marginal effects on 3' azido-3'-deoxthymidine (AZT) susceptibility. However, when introduced into a genetic background for AZT resistance (D67N, K70R, T215Y, T219Q), the K65R mutation reversed the AZT resistance. DNA sequencing of RT clones derived from the second resistant isolate identified the L74V mutation, previously reported to cause ddI resistance. The L74V mutation also decreased the AZT resistance when the mutation was introduced into a genetic background for AZT resistance (D67N, K70R, T215Y, T219Q) but to a lesser degree than the K65R mutation did. These findings indicate that DXG and certain 2',3'-dideoxy compounds (e.g., ddI) can select for the same resistance mutations and thus may not be optimal for use in combination. However, the combination of AZT with DXG or its orally bioavailable prodrug (-)-beta-D-2, 6-diaminopurine-dioxolane should be explored because of the suppressive effects of the K65R and L74V mutations on AZT resistance. PMID- 10858332 TI - Quinupristin-dalfopristin combined with beta-lactams for treatment of experimental endocarditis due to Staphylococcus aureus constitutively resistant to macrolide-lincosamide-streptogramin B antibiotics. AB - Quinupristin-dalfopristin (Q-D) is an injectable streptogramin active against most gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA). In experimental endocarditis, however, Q-D was less efficacious against MRSA isolates constitutively resistant to macrolide-lincosamide streptogram B (C-MLS(B)) than against MLS(B)-susceptible isolates. To circumvent this problem, we used the checkerboard method to screen drug combinations that would increase the efficacy of Q-D against such bacteria. beta-Lactams consistently exhibited additive or synergistic activity with Q-D. Glycopeptides, quinolones, and aminoglycosides were indifferent. No drugs were antagonistic. The positive Q-D-beta-lactam interaction was independent of MLS(B) or beta-lactam resistance. Moreover, addition of Q-D at one-fourth the MIC to flucloxacillin containing plates decreased the flucloxacillin MIC for MRSA from 500 to 1,000 mg/liter to 30 to 60 mg/liter. Yet, Q-D-beta-lactam combinations were not synergistic in bactericidal tests. Rats with aortic vegetations were infected with two C-MLS(B)-resistant MRSA isolates (isolates AW7 and P8) and were treated for 3 or 5 days with drug dosages simulating the following treatments in humans: (i) Q-D at 7 mg/kg two times a day (b.i.d.) (a relatively low dosage purposely used to help detect positive drug interactions), (ii) cefamandole at constant levels in serum of 30 mg/liter, (iii) cefepime at 2 g b.i.d., (iv) Q-D combined with either cefamandole or cefepime. Any of the drugs used alone resulted in treatment failure. In contrast, Q-D plus either cefamandole or cefepime significantly decreased valve infection compared to the levels of infection for both untreated controls and those that received monotherapy (P < 0.05). Importantly, Q-D prevented the growth of highly beta-lactam-resistant MRSA in vivo. The mechanism of this beneficial drug interaction is unknown. However, Q-D beta-lactam combinations might be useful for the treatment of complicated infections caused by multiple organisms, including MRSA. PMID- 10858333 TI - Clinical efficacy of intravenous followed by oral azithromycin monotherapy in hospitalized patients with community-acquired pneumonia. The Azithromycin Intravenous Clinical Trials Group. AB - The purpose of this study was to evaluate intravenous (i.v.) azithromycin followed by oral azithromycin as a monotherapeutic regimen for community-acquired pneumonia (CAP). Two trials of i.v. azithromycin used as initial monotherapy in hospitalized CAP patients are summarized. Clinical efficacy is reported from an open-label randomized trial of azithromycin compared to cefuroxime with or without erythromycin. Bacteriologic and clinical efficacy results are also presented from a noncomparative trial of i.v. azithromycin that was designed to give additional clinical experience with a larger number of pathogens. Azithromycin was administered to 414 patients: 202 and 212 in the comparative and noncomparative trials, respectively. The comparator regimen was used as treatment for 201 patients; 105 were treated with cefuroxime alone and 96 were given cefuroxime plus erythromycin. In the comparative trial, clinical outcome data were available for 268 evaluable patients with confirmed CAP at the 10- to 14-day visit, with 106 (77%) of the azithromycin patients cured or improved and 97 (74%) of the comparator patients cured or improved. Mean i.v. treatment duration and mean total treatment duration (i.v. and oral) for the clinically evaluable patients were significantly (P < 0.05) shorter for the azithromycin group (3.6 days for the i.v. group and 8.6 days for the i.v. and oral group) than for the evaluable patients given cefuroxime plus erythromycin (4.0 days for the i.v. group and 10.3 days for the i.v. and oral group). The present comparative study demonstrates that initial therapy with i.v. azithromycin for hospitalized patients with CAP is associated with fewer side effects and is equal in efficacy to a 1993 American Thoracic Society-suggested regimen of cefuroxime plus erythromycin when the erythromycin is deemed necessary by clinicians. PMID- 10858334 TI - IB-367, a protegrin peptide with in vitro and in vivo activities against the microflora associated with oral mucositis. AB - Although the microflora associated with oral mucositis initiated by cytotoxic therapy is not well characterized, several studies suggest that reduction of the microbial load in the oral cavity has some clinical benefit. The MICs of IB-367, a synthetic protegrin analog, ranged from 0.13 to 64 microgram/ml for gram positive bacteria (Streptococcus mitis, Streptococcus sanguis, Streptococcus salivarius, and Staphylococcus aureus) and from 0.06 to 8 microgram/ml for gram negative species (Klebsiella, Escherichia, and Pseudomonas). IB-367 exhibited rapid, microbicidal activity against both log- and stationary-phase cultures of methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa. At concentrations near the MICs for these two organisms (4 and 2 microgram/ml, respectively), IB-367 reduced viability by more than 3 logs in less than 16 min. Similarly, IB-367 effected a 4-log reduction of the endogenous microflora in pooled human saliva within 2 min at 250 microgram/ml, a concentration readily attained by local delivery. After nine serial transfers at 0.5x the MIC, the MIC of IB-367 for MRSA and P. aeruginosa increased only two to four times. In a phase I clinical study with healthy volunteers, IB-367 was well tolerated, with no detectable systemic absorption. One hour after treatment with 9 mg of IB-367, the prevalence of gram-negative bacteria and yeast was reduced, and the density of the predominant gram-positive oral flora was decreased 1,000 times. IB-367's properties (speed of killing, breadth of spectrum, and lack of resistance) make the compound a strong candidate for the prophylaxis of oral mucositis. Phase II clinical trials with IB-367 are under way for this indication in immunocompromised subjects. PMID- 10858335 TI - Genetic localization and molecular characterization of the nonS gene required for macrotetrolide biosynthesis in Streptomyces griseus DSM40695. AB - The macrotetrolides are a family of cyclic polyethers derived from tetramerization, in a stereospecific fashion, of the enantiomeric nonactic acid (NA) and its homologs. Isotope labeling experiments established that NA is of polyketide origin, and biochemical investigations demonstrated that 2-methyl-6,8 dihydroxynon-2E-enoic acid can be converted into NA by a cell-free preparation from Streptomyces lividans that expresses nonS. These results lead to the hypothesis that macrotetrolide biosynthesis involves a pair of enantiospecific polyketide pathways. In this work, a 55-kb contiguous DNA region was cloned from Streptomyces griseus DSM40695, a 6.3-kb fragment of which was sequenced to reveal five open reading frames, including the previously reported nonR and nonS genes. Inactivation of nonS in vivo completely abolished macrotetrolide production. Complementation of the nonS mutant by the expression of nonS in trans fully restored its macrotetrolide production ability, with a distribution of individual macrotetrolides similar to that for the wild-type producer. In contrast, fermentation of the nonS mutant in the presence of exogenous (+/-)-NA resulted in the production of nonactin, monactin, and dinactin but not in the production of trinactin and tetranactin. These results prove the direct involvement of nonS in macrotetrolide biosynthesis. The difference in macrotetrolide production between in vivo complementation of the nonS mutant by the plasmid-borne nonS gene and fermentation of the nonS mutant in the presence of exogenously added (+/-)-NA suggests that NonS catalyzes the formation of (-)-NA and its homologs, supporting the existence of a pair of enantiospecific polyketide pathways for macrotetrolide biosynthesis in S. griseus. The latter should provide a model that can be used to study the mechanism by which polyketide synthase controls stereochemistry during polyketide biosynthesis. PMID- 10858336 TI - Role of antibiotic penetration limitation in Klebsiella pneumoniae biofilm resistance to ampicillin and ciprofloxacin. AB - The penetration of two antibiotics, ampicillin and ciprofloxacin, through biofilms developed in an in vitro model system was investigated. The susceptibilities of biofilms and corresponding freely suspended bacteria to killing by the antibiotics were also measured. Biofilms of Klebsiella pneumoniae were developed on microporous membranes resting on agar nutrient medium. The susceptibilities of planktonic cultures and biofilms to 10 times the MIC were determined. Antibiotic penetration through biofilms was measured by assaying the concentration of antibiotic that diffused through the biofilm to an overlying filter disk. Parallel experiments were performed with a mutant K. pneumoniae strain in which beta-lactamase activity was eliminated. For wild-type K. pneumoniae grown in suspension culture, ampicillin and ciprofloxacin MICs were 500 and 0.18 microgram/ml, respectively. The log reductions in the number of CFU of planktonic wild-type bacteria after 4 h of treatment at 10 times the MIC were 4.43 +/- 0.33 and 4.14 +/- 0.33 for ampicillin and ciprofloxacin, respectively. Biofilms of the same strain were much less susceptible, yielding log reductions in the number of CFU of -0.06 +/- 0.06 and 1.02 +/- 0.04 for ampicillin and ciprofloxacin, respectively, for the same treatment. The number of CFU in the biofilms after 24 h of antibiotic exposure was not statistically different from the number after 4 h of treatment. Ampicillin did not penetrate wild-type K. pneumoniae biofilms, whereas ciprofloxacin and a nonreactive tracer (chloride ion) penetrated the biofilms quickly. The concentration of ciprofloxacin reached the MIC throughout the biofilm within 20 min. Ampicillin penetrated biofilms formed by a beta-lactamase-deficient mutant. However, the biofilms formed by this mutant were resistant to ampicillin treatment, exhibiting a 0.18 +/- 0.07 log reduction in the number of CFU after 4 h of exposure and a 1.64 +/- 0.33 log reduction in the number of CFU after 24 h of exposure. Poor penetration contributed to wild-type biofilm resistance to ampicillin but not to ciprofloxacin. The increased resistance of the wild-type strain to ciprofloxacin and the mutant strain to ampicillin and ciprofloxacin could not be accounted for by antibiotic inactivation or slow diffusion since these antibiotics fully penetrated the biofilms. These results suggest that some other resistance mechanism is involved for both agents. PMID- 10858337 TI - Peptide deformylase in Staphylococcus aureus: resistance to inhibition is mediated by mutations in the formyltransferase gene. AB - Peptide deformylase, a bacterial enzyme, represents a novel target for antibiotic discovery. Two deformylase homologs, defA and defB, were identified in Staphylococcus aureus. The defA homolog, located upstream of the transformylase gene, was identified by genomic analysis and was cloned from chromosomal DNA by PCR. A distinct homolog, defB, was cloned from an S. aureus genomic library by complementation of the arabinose-dependent phenotype of a P(BAD)-def Escherichia coli strain grown under arabinose-limiting conditions. Overexpression in E. coli of defB, but not defA, correlated to increased deformylase activity and decreased susceptibility to actinonin, a deformylase-specific inhibitor. The defB gene could not be disrupted in wild-type S. aureus, suggesting that this gene, which encodes a functional deformylase, is essential. In contrast, the defA gene could be inactivated; the function of this gene is unknown. Actinonin-resistant mutants grew slowly in vitro and did not show cross-resistance to other classes of antibiotics. When compared to the parent, an actinonin-resistant strain produced an attenuated infection in a murine abscess model, indicating that this strain also has a growth disadvantage in vivo. Sequence analysis of the actinonin resistant mutants revealed that each harbors a loss-of-function mutation in the fmt gene. Susceptibility to actinonin was restored when the wild-type fmt gene was introduced into these mutant strains. An S. aureus Deltafmt strain was also resistant to actinonin, suggesting that a functional deformylase activity is not required in a strain that lacks formyltransferase activity. Accordingly, the defB gene could be disrupted in an fmt mutant. PMID- 10858338 TI - A population pharmacokinetic analysis of nelfinavir mesylate in human immunodeficiency virus-infected patients enrolled in a phase III clinical trial. AB - A population pharmacokinetic analysis was conducted on nelfinavir in patients infected with human immunodeficiency virus (HIV) who were enrolled in a phase III clinical trial. The data consisted of 509 plasma concentrations from 174 patients who received nelfinavir at a dose of 500 or 750 mg three times a day. The analysis was performed using nonlinear mixed-effect modeling as implemented in NONMEM (version 4.0; double precision). A one-compartment model with first-order absorption best described the data. The timing and small number of early postdose blood levels did not allow accurate estimation of volume of distribution (V/F) and the absorption rate constant (k(a)). As a result, two models were used to analyze the data: model 1, in which oral clearance (CL/F), V/F, and k(a) were estimated, and model 2, in which V/F and k(a) were fixed to known values and only CL/F was estimated. Estimates of CL/F ranged from 41. 9 to 45.1 liters/h, values in close agreement with previous studies. Neither body weight, age, sex, race, dose level, baseline viral load, metabolite-to-parent drug plasma concentration ratio, history of liver disease, nor elevated results of liver function tests appeared to be significant covariates for clearance. The only significant covariate-parameter relationship was concomitant use of fluconazole on CL/F, which was associated with a modest reduction in interindividual variability of CL/F. Patients who received concomitant therapy with fluconazole had a statistically significant reduction in nelfinavir CL/F of 26 to 30%. Since serious dose-limiting toxicity and concentration-related toxicities are not apparent for nelfinavir, this effect of fluconazole is unlikely to be of clinical significance. PMID- 10858339 TI - Comparative in vitro activities of linezolid, quinupristin-dalfopristin, moxifloxacin, and trovafloxacin against erythromycin-susceptible and -resistant streptococci. AB - The in vitro activities of the new agents linezolid, quinupristin-dalfopristin, moxifloxacin, and trovafloxacin were determined and compared with those of penicillin, clindamycin, and four macrolides against 53 erythromycin-resistant Streptococcus pneumoniae, 117 S. pyogenes (64 erythromycin-susceptible and 53 resistant), and 101 S. agalactiae (53 erythromycin-susceptible and 48 -resistant) isolates. Differentiation of macrolide resistance phenotypes was performed by the double-disk method. The genetic basis for macrolide resistance in 52 strains was also determined. The M phenotype was found in 84.9, 6.3, and 1.9% of S. pyogenes, S. agalactiae, and S. pneumoniae isolates, respectively. These strains were susceptible to miocamycin and clindamycin. Strains with the inducible phenotype accounted for 27.1% of S. agalactiae isolates and 9.4% each of S. pyogenes and S. pneumoniae isolates. All erythromycin-resistant isolates were also resistant to the 14- and 15-membered macrolides tested. Strains with all three phenotypes were susceptible to 0.5 x 10(9) neutrophils/liter) or up to a maximum of 3 days following the end of neutropenia, unless a systemic fungal infection was documented or suspected. The maximum treatment duration was 56 days. In the intent-to-treat population, invasive aspergillosis was noted in 5 (1.8%) of the 281 patients assigned to itraconazole oral solution and in 9 (3.3%) of the 276 patients assigned to oral amphotericin B; of these, 1 and 4 patients died, respectively. Proven systemic fungal infection (including invasive aspergillosis) occurred in 8 patients (2.8%) who received itraconazole, compared with 13 (4.7%) who received oral amphotericin B. Itraconazole significantly reduced the incidence of superficial fungal infections as compared to oral amphotericin B (2 [1%] versus 13 [5%]; P = 0.004). Although the incidences of suspected fungal infection (including fever of unknown origin) were not different between the groups, fewer patients were administered intravenous systemic antifungals (mainly intravenous amphotericin B) in the group receiving itraconazole than in the group receiving oral amphotericin B (114 [41%] versus 132 [48%]; P = 0.066). Adequate plasma itraconazole levels were achieved in about 80% of the patients from 1 week after the start of treatment. In both groups, the trial medication was safe and well tolerated. Prophylactic administration of itraconazole oral solution significantly reduces superficial fungal infection in patients with hematological malignancies and neutropenia. The incidence of proven systemic fungal infections, the number of deaths due to deep fungal infections, and the use of systemic antifungals tended to be lower in the itraconazole-treated group than in the amphotericin B-treated group, without statistical significance. Itraconazole oral solution is a broad-spectrum systemic antifungal agent with prophylactic activity in neutropenic patients, especially for those at high risk of prolonged neutropenia. PMID- 10858350 TI - Antipneumococcal activity of ABT-773 compared to those of 10 other agents. AB - MICs, time-kills, and postantibiotic effects (PAEs) of ABT-773 (a new ketolide) and 10 other agents were determined against 226 pneumococci. Against 78 ermB- and 44 mefE-containing strains, ABT-773 MICs at which 50% of the isolates tested were inhibited (MIC(50)s) and MIC(90)s were 0.016 to 0.03 and 0.125 microgram/ml, respectively. Clindamycin was active only against macrolide-resistant strains containing mefE (MIC(50), 0.06 microgram/ml; MIC(90), 0.125 microgram/ml). Activities of pristinamycin (MIC(90), 0.5 microgram/ml) and vancomycin (MIC(90), 0.25 microgram/ml) were unaffected by macrolide or penicillin resistance, while beta-lactam MICs rose with those of penicillin G. Against 19 strains with L4 ribosomal protein mutations and two strains with mutations in domain V of 23S rRNA, ABT-773 MICs were 0.03 to 0.25 microgram/ml, while macrolide and azalide MICs were all >/=16.0 microgram/ml. ABT-773 was bactericidal at twice the MIC after 24 h for 8 of 12 strains (including three strains with erythromycin MICs greater than or equal to 64.0 microgram/ml). Kill kinetics of erythromycin, azithromycin, clarithromycin, and roxithromycin against macrolide-susceptible strains were slower than those of ABT-773. ABT-773 had longer PAEs than macrolides, azithromycin, clindamycin, or beta-lactams, including against ermB containing strains. ABT-773, therefore, shows promising in vitro activity against macrolide-susceptible as well as -resistant pneumococci. PMID- 10858351 TI - Effect of zidovudine on the primary cytolytic T-lymphocyte response and T-cell effector function. AB - Azidothymidine (AZT) and other nucleoside analogues, used to treat AIDS, can cause severe clinical side effects and are suspected of suppressing immune cell proliferation and effector immune cell function. The purpose of the present study was to quantitatively measure the effects of AZT on cytotoxic T-lymphocyte (CTL) priming and to determine if the major histocompatibility complex-restricted CTL killing was affected by AZT exposure. For this purpose, we employed a murine alloantigen model and limiting-dilution analysis (LDA) to estimate cytotoxic effector cell frequencies of alloreactive splenocytes treated with drug during antigen sensitization. This noninfectious model was chosen to avoid analysis of a virus-compromised immune system. Exposure of splenocytes to therapeutic concentrations of AZT (2 to 10 microM) caused a two- to threefold dose-dependent reduction in CLT precursor frequency. This reduction was caused by decreased proliferation of alloantigen-specific CTLs rather than loss of function, because full cytolytic function could be restored by adjusting the AZT-treated effector/target cell ratios to that of untreated cells. In addition, when AZT was added to the assay system at various times during antigen sensitization there was a time-related loss of the suppressive effect on the generation of cytolytic effector function, suggesting that functional CTLs are not affected by even high doses of AZT. Taken together, the data indicate that the reduction of CTL function associated with AZT treatment is due to a quantitative decrease of effector cell precursor frequency rather than to direct drug cytotoxicity or interference with mediation of cytolysis. Furthermore, antigen-naive immune cells were most sensitive to this effect during the first few days following antigen encounter. PMID- 10858352 TI - Genetic organization of the downstream region of the mecA element in methicillin resistant Staphylococcus aureus isolates carrying different polymorphisms of this region. AB - We describe here the genetic organization of the mec element downstream of the mecA gene in 34 different methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates carrying 13 of the most frequent polymorphisms of mecA and representing the major epidemic clones of MRSA. All polymorphisms carried three common genetic elements: the hypervariable region, a copy of IS431, and a unique 2-kb sequence (downstream constant segment, or dcs) for which no homologous sequences are found in data banks. Polymorphisms of the downstream region were shown to be caused by the presence of linearized plasmids flanked by insertion sequences (pUB110, pT181, and pI258) and the autonomous insertion sequence IS256. PMID- 10858353 TI - Amphotericin B resistance and membrane fluidity in Kluyveromyces lactis strains. AB - The membrane fluidity of reduced-amphotericin B (AmB)-sensitivity Kluyveromyces lactis mutant strain is higher than that of the wild-type K. lactis strain. After culture of the K. lactis and K. lactis mutant cells in the presence of subinhibitory doses of AmB (10 and 125 mg/liter, respectively), the plasma membranes of both yeast strains also showed a higher fluidity than did those of control cells. High membrane fluidity was associated with changes in the structural properties of the membranes. Culture of the K. lactis and K. lactis mutant cells in the presence of AmB induced changes in membrane lipid contents. In particular, phospholipid contents were increased in both strains treated with AmB, compared with their corresponding counterparts. As a result, the sterol/phospholipid ratio decreased. The relative proportion of monounsaturated fatty acids also increased after AmB treatment. The saturated fatty acid/monounsaturated fatty acid ratio decreased in K. lactis and K. lactis mutant cells treated with AmB but also in K. lactis mutant control cells compared to that in the K. lactis wild strain. These changes in lipid composition explain the higher fluidity, which could represent a process of metabolic resistance of the yeasts to AmB. PMID- 10858354 TI - Evaluation of voriconazole pharmacodynamics using time-kill methodology. AB - Voriconazole is an investigational azole antifungal agent with activity against a variety of fungal species, including fluconazole-susceptible and -resistant Candida species and Cryptococcus neoformans. In this study, we employed in vitro time-kill methods to characterize the relationship between concentrations of voriconazole and its fungistatic activity against Candida albicans, Candida glabrata, Candida tropicalis, and C. neoformans. Isolates were exposed to voriconazole concentrations ranging from 0.0625 to 16 times the MIC, and the viable colony counts were determined over time. The 50 and 90% effective concentrations (EC(50) and EC(90), respectively) were determined at 8, 12, and 24 h following the addition of voriconazole. At each time point, near-maximal fungistatic activity, as indicated by the EC(90), was noted at a drug concentration of approximately three times the MIC. Additionally, EC(50) and EC(90) did not change over time, thus suggesting that the rate of activity was not improved by increasing concentrations. Voriconazole exhibits non concentration-dependent pharmacodynamic characteristics in vitro. PMID- 10858355 TI - Comparison of a rabbit model of bacterial endocarditis and an in vitro infection model with simulated endocardial vegetations. AB - Animal models are commonly used to determine the efficacy of various antimicrobial agents for treatment of bacterial endocarditis. Previously we have utilized an in vitro infection model, which incorporates simulated endocardial vegetations (SEVs) to evaluate the pharmacodynamics of various antibiotics. In the present study, we compared four experimental rabbit endocarditis protocols to an in vitro infection model in an effort to determine if these models are comparable. We have evaluated the activity of clinafloxacin, trovafloxacin, sparfloxacin, and ciprofloxacin in rabbit models against Staphylococcus aureus and Enterococcus spp. In vitro models were performed simulating the antibiotic pharmacokinetics obtained in the in vivo studies. Models were dosed the same as rabbit models, and SEVs were evaluated at the same time the rabbit vegetations were examined. Clinafloxacin and trovafloxacin were evaluated against methicillin susceptible (MSSA1199) and -resistant (MRSA494) strains of S. aureus. Ciprofloxacin was studied against MSSA1199 and MSSA487. Sparfloxacin and clinafloxacin were evaluated against Enterococcus faecium SF2149 and Enterococcus faecalis WH245, respectively. We found that reductions in SEV bacterial density obtained in the in vitro model were similar to those obtained in rabbit vegetations, indicating that the SEV model may be a valuable tool for assessing antibiotic potential in the treatment of bacterial endocarditis. PMID- 10858356 TI - In vitro activities of daptomycin, arbekacin, vancomycin, and gentamicin alone and/or in combination against glycopeptide intermediate-resistant Staphylococcus aureus in an infection model. AB - Daptomycin, a lipopeptide antibiotic, has broad activity against gram-positive organisms, similar to vancomycin; however, its mechanism of action differs, resulting in interference with cell membrane transport and a more rapid bactericidal activity. In light of increasing need for alternative treatments against intermediate-resistant Staphylococcus aureus, there is revitalized interest in this antibiotic. We, therefore, evaluated the activity of daptomycin alone or in combination in an in vitro infection model against two glycopeptide intermediate-resistant S. aureus (GISA) isolates. Newly designed regimens of daptomycin at 4 and 6 mg/kg of body weight every 24 h (q24h) were compared to the previous regimen of 3 mg/kg q12h. Daptomycin MICs and minimal bactericidal concentrations (MBCs) (MIC/MBC) for Mu-50, HIP5836 (992), and MRSA-67 were 0.5/1.0, 0.5/1.0, and 0.125/0.5 microgram/ml, respectively. MICs and MBCs of arbekacin for the three strains were 2.0/8.0, 0. 125/0.5, and 0.125/0.25 microgram/ml, respectively. Vancomycin and gentamicin MICs and MBCs for the three strains were 8.0/8.0, 8.0/8.0, and 0.5/1.0 microgram/ml and 128/128, 0.5/1.0, and 0.25/0.5 microgram/ml, respectively. Our experience with daptomycin in an in vitro infection model has shown significant kill against the two GISA strains (Mu 50 and 992) (P < 0.03). We also noted that kill was related to a total dose effect for 992, in which simulated daptomycin in vivo dosages of 6 mg/kg q24h and 3 mg/kg q12h produced similar kill and 4 mg/kg q24h resulted in significant regrowth (P 0.75. Scores describing limited physical function were strikingly low. Specifically, activity limited to bed to chair movement and the need for complete assistance with activities of daily living were all assigned utility scores <0.2. Twenty-four hour oxygen dependence was scored at 0.33. Presence or absence of pulmonary illness did not predict scores for any outcome. CONCLUSIONS: Whether patients suffer from chronic lung disease or not, they do not regard the postoperative outcomes reported in the lung surgery literature as sufficiently morbid to forego important surgery. However, physical debility is perceived as extremely undesirable, and anticipation of its occurrence could deter surgery. Therefore, identification of preoperative predictors of postoperative physical debility would be invaluable for counseling patients who face difficult decisions about lung resection. PMID- 10858383 TI - Lung volume reduction surgery: a survey on the European experience. AB - STUDY OBJECTIVE: To evaluate the activity and evolution in the field of lung volume reduction surgery (LVRS) performed at surgical centers in Europe. BACKGROUND: LVRS is a novel surgical therapy with the potential to improve lung function, exercise performance, and quality of life in selected patients suffering from severe pulmonary emphysema. METHODS: Questionnaire addressed to 75 European thoracic surgical centers presumed to perform LVRS, and review of the literature. RESULTS: Of 45 responding centers, 42 centers in 17 countries covering a population of 423 million reported performing LVRS. Until the end of 1998, 1,120 patients were reported to have undergone LVRS, corresponding to 2.6 patients/million inhabitants. Thirty-one of 40 centers (78%) perform the operation bilaterally. Most centers (83%) evaluate their activity prospectively. The average perioperative mortality rate of 4.1% is moderate. The most commonly utilized technique is video-assisted thoracoscopy, which is most frequently performed bilaterally. Two thirds of the centers treat patients with alpha(1) antitrypsin deficiency, and half of the centers will consider patients with homogenous morphology of emphysema on CT scan for LVRS. Half of the centers also perform lung transplantation. The five largest centers have operated on 49% of all LVRS patients assessed by this survey. CONCLUSIONS: LVRS is performed at few thoracic surgical centers throughout Europe, with a large variation in the operative activity between different regions. Half of the centers also perform lung transplantation. Between 1995 and 1997, the number of LVRS procedures performed per year nearly tripled but has reached a plateau since then. As five centers perform nearly half the total number of operations, an optimal exchange of knowledge with smaller centers seems important. PMID- 10858384 TI - Correlation of tumor size and survival in patients with stage IA non-small cell lung cancer. AB - OBJECTIVE: The purpose of this study was to determine the relationship between tumor size and survival in patients with stage IA non-small cell lung cancer (non small cell lung cancer; ie, lesions < 3 cm). METHOD: Five hundred ten patients with pathologic stage IA (T1N0M0) non-small cell lung cancer were identified from our tumor registry over an 18-year period (from 1981 to 1999). There were 285 men and 225 women, with a mean age of 63 years (range, 31 to 90 years). The Cox proportional model was used to examine the effect on survival. Tumor size was incorporated into the model as a linear effect and as categorical variables. The Kaplan-Meier product limit estimator was used to graphically display the relationship between the tumor size and survival. RESULTS: The Cox proportional hazards model did not show a statistically significant relationship between tumor size and survival (p = 0.701) as a linear effect. Tumor size was then categorized into quartiles, and again there was no statistically significant difference in survival between groups (p = 0.597). Tumor size was also categorized into deciles, and there was no statistical relationship between tumor size and survival (p = 0.674). CONCLUSIONS: This study confirms stratifying patients with stage IA non-small cell lung cancer in the same TNM classification, given no apparent difference in survival. Unfortunately, these data caution that improved small nodule detection with screening CT may not significantly improve lung cancer mortality. The appropriate prospective randomized trial appears warranted. PMID- 10858385 TI - Outcome of bronchial carcinoma in situ. AB - INTRODUCTION: The proportion of patients with carcinoma in situ in whom invasive cancer will develop is not known. It is important for clinical decision making to know the outcome of these lesions. The same applies for studies assessing the effectiveness of chemoprevention treatment or endobronchial therapy. METHODS: The records of patients with a bronchial carcinoma in situ who had undergone autofluorescence bronchoscopic examinations at regular intervals during a follow up period for at least 6 months were reviewed. Data were examined for the outcome of carcinoma in situ, and for the detection, course, and bronchoscopic findings of neoplastic lesions at other bronchial sites. RESULTS: Progression to carcinoma occurred in five of nine patients (56%) with a carcinoma in situ. Eight neoplastic lesions were detected at other sites in four of the nine patients (44%). In earlier biopsy specimens of two sites that later showed a severe dysplasia and a carcinoma, only normal epithelium was found. Biopsies had been performed at these sites because they were assessed as suspicious during autofluorescence bronchoscopy. CONCLUSION: The majority of sites showing a carcinoma in situ progressed to invasive carcinoma. A considerable portion of the patients had neoplastic lesions at other bronchial sites. The fluorescence pattern of the bronchial mucosa may reflect early changes that are not found at histopathologic examination, but which may progress to neoplastic growth. PMID- 10858386 TI - Clinical predictors of N2 disease in non-small cell lung cancer. AB - OBJECTIVES: To identify clinical or radiologic predictors of mediastinal lymph node involvement in patients with non-small cell lung cancer, and to define the indications of preoperative mediastinoscopy. METHODS: From August 1992 through April 1997, 387 patients with lung cancer (290 adenocarcinoma and 97 squamous cell carcinoma) underwent surgical resection. We retrospectively measured all mediastinal lymph node sizes both in the shortest and longest axes on contrast enhanced CT scan to determine the optimal size criteria. Using multivariate logistic regression analysis, we identified clinical or radiologic predictors of N2 disease. RESULTS: We could not identify reliable size criteria for nodal involvement. We found two significant predictive factors of N2 disease on the basis of multivariable analysis: maximum tumor dimension and serum carcinoembryonic antigen (CEA) concentrations. The lymph node size did not prove to be a significant factor. Among 50 patients with serum CEA concentrations < 5.0 ng/mL and maximum tumor dimension < 20 mm, pathologic N2 disease was proven only in three patients (6%), regardless of the lymph node size on CT scan. Among 140 patients with serum CEA concentrations > or = 5.0 ng/mL and maximum tumor dimension > or = 20 mm, approximately one third (n = 46) showed N2 disease. CONCLUSION: Serum CEA concentrations and maximum tumor dimension were more valuable in predicting N2 disease than the lymph node size on CT scan. Mediastinoscopy is indicated in patients with serum CEA concentrations > or = 5.0 ng/mL and maximum tumor dimension > or = 20 mm, and not indicated in patients with serum CEA concentrations < 5.0 ng/mL and maximum tumor dimension < 20 mm. PMID- 10858387 TI - A multicenter phase II trial of vinorelbine plus gemcitabine in previously untreated inoperable (Stage IIIB/IV) non-small cell lung cancer. AB - STUDY OBJECTIVE: Vinorelbine and gemcitabine are two active single agents used in the treatment of non-small cell lung cancer (NSCLC). A clinical trial was conducted to evaluate the efficacy and toxicity of vinorelbine plus gemcitabine in patients with inoperable (stage IIIB or IV) NSCLC. DESIGN: A multicenter phase II study. Vinorelbine, 20 mg/m(2), was given as a 10-min IV infusion, followed by a 30-min IV infusion of gemcitabine, 800 mg/m(2), on days 1, 8, and 15 of each 28 day cycle. PATIENTS AND MEASUREMENTS: From March 1998 to August 1998, 40 patients were enrolled in the study. The efficacy and toxicity of the treatment were recorded. RESULTS: All patients are evaluable for treatment response and toxicity profile. Two patients achieved a complete response, and 27 patients achieved a partial response, with an overall response rate of 72.5% (95% confidence interval, 58.7 to 86.3%). Median survival time was 11 months. The significant (World Health Organization grade, 3/4) toxicities were myelosuppression, including leukopenia (47.5% of patients), anemia (17.5% of patients), and thrombocytopenia (12.5% of patients). However, febrile neutropenia occurred in three patients and accounted for one treatment-related death. Fatigue, or flu like syndrome, occurred in 17 patients, and the symptoms were reversed spontaneously 1 to 2 days after injection in 10 patients. Another seven patients needed dose reduction to ameliorate symptoms. Interstitial pneumonitis occurred in six patients who recovered after steroid treatment. No patient suffered from grade 3 or 4 nausea/vomiting. CONCLUSION: The combination of vinorelbine and gemcitabine in patients with advanced NSCLC is a highly active non-cisplatin containing regimen with an acceptable toxicity profile. PMID- 10858388 TI - Changes in airway responsiveness following mantle radiotherapy for Hodgkin's disease. AB - STUDY OBJECTIVES To investigate whether mantle radiotherapy (MRT) for the lung, through its proinflammatory effects, can induce an increase in airway responsiveness. DESIGN: Follow-up of the changes in lung function and methacholine responsiveness in patients 1, 6, 12, and 24 months after they underwent MRT. PATIENTS: Thirteen nonasthmatic patients with bulky Hodgkin's lymphoma who were scheduled for MRT. MEASUREMENTS AND RESULTS: Chest radiographs, lung function tests, methacholine thresholds of the bronchi (the provocative dose of methacholine causing a 10% fall in FEV(1) [PD(10)]) and central airway (the provocative dose of methacholine causing a 25% fall in the maximal mid inspiratory flow [PD(25)MIF(50)]), and the provocative dose of methacholine causing five or more coughs (PDcough) were serially assessed. One month after patients underwent MRT, there were significant decreases in PD(10) (mean [+/- SEM], 2,583 +/- 414 microg to 1,512 +/- 422 microg, respectively; p < 0.05), PD(25)MIF(50) (mean 2,898 +/- 372 microg to 1,340 +/- 356 microg, respectively; p < 0.05), and PDcough (mean 3,127 +/- 415 microg to 1,751 +/- 447 microg; p < 0.05), which were independent of the decrease in FEV(1) and reversed within 6 months in all patients but three. Six months after undergoing MRT, four patients showed radiation-induced lung injury (RI) on chest radiographs, which subsequently evolved into fibrosis. These patients had greater decreases in vital capacity, FEV(1), MIF(50), and methacholine thresholds than those without RI, and this persisted up to 2 years after they had undergone MRT. One year after the patients underwent MRT, a close relationship was found overall between the change in FEV(1) and those in both PD(10) (r = 0.733; p = 0.004) and PD(25)MIF(50) (r = 0.712; p = 0.006). CONCLUSIONS: : MRT triggers an early transient increase in airway responsiveness, which reverses spontaneously. In patients with RI, the persistence of airway dysfunction long after undergoing MRT may depend on airway remodeling from radiation fibrosis. PMID- 10858389 TI - Sleep-disordered breathing and myocardial ischemia in patients with coronary artery disease. AB - STUDY OBJECTIVES: To examine the occurrence of nocturnal myocardial ischemia and its relationship to sleep-disordered breathing (apneas and oxygen desaturations) in randomly selected men and women undergoing coronary angiography because of angina pectoris. DESIGN: An observational study using an overnight sleep study and Holter recording to examine disordered breathing (oxyhemoglobin desaturations > or = 4% and apnea-hypopneas), heart rates, and ST-segment depressions (> or = 1 mm, > or = 1 min). SETTING: University Hospital, Umea, a teaching hospital in northern Sweden. PATIENTS: One hundred thirty-two men and 94 women referred for consideration of coronary intervention were randomly included, by lot. RESULTS: ST-segment depressions occurred in 59% (134 of 226) of the patients, and nocturnal ST-segment depressions occurred in 31% (69 of 226). A ST-segment depression occurred within 2 min after an apnea-hypopnea or desaturation in 12% (27 of 226) of patients. This temporal association was seen in 19% of nocturnal ST-segment depressions (71 of 366), more frequently in men (p < 0.01) and in more severely disordered breathing (p < 0.001). Most of these ST-segment depressions were preceded by a series of breathing events: three or more apnea-hypopneas or desaturations or both in 70% (50 of 71). CONCLUSION: Episodes of nocturnal myocardial ischemia are common in patients with angina pectoris. However, a temporal relationship between sleep-disordered breathing and myocardial ischemia is present only in a minority of the patients, but occurs more frequently in men and in more severely disordered breathing. PMID- 10858390 TI - Sleep medicine practices, training, and attitudes: a wake-up call for pulmonologists. AB - STUDY OBJECTIVES: To determine attitudes and knowledge about sleep medicine among chest physicians. DESIGN: : Interactive survey of self-selected respondents. SETTING: Interactive session at the 1998 American College of Chest Physicians (ACCP) annual meeting. PARTICIPANTS: Approximately 60 chest physicians. INTERVENTIONS: Interactive questions about the knowledge, training, attitudes, and practice of sleep medicine. MEASUREMENTS AND RESULTS: Response rates demonstrated that 65% of respondents directed or were on the staff of a sleep laboratory, 18% had American Board of Sleep Medicine (ABSM) certification, and only 3% had completed formal sleep medicine training, and performance on test questions about sleep-disordered breathing was better than that on questions about "nonpulmonary" sleep disorders. We polled approximately 60 participants in an interactive session called "Issues in Sleep Medicine Education and Practice" at the ACCP annual meeting in October 1998. The group was well-credentialed, with about one third of participants being board-certified in pulmonary medicine and critical care medicine, and about 17% having passed the ABSM examination. About two thirds of the group spent < or = 25% of their time in the practice of sleep medicine, but > 30% directed sleep laboratories. Respondents thought that sleep training was better addressed in pulmonary fellowship training than in medical school or other postgraduate training experiences. Forty-three percent of the group had received training in sleep medicine as part of a pulmonary fellowship. About half of the sample thought that formal training should be required for eligibility to take the ABSM examination. When presented with two "nonpulmonary" sleep disorder cases, this well-trained and self-selected group did not perform very well. The findings suggest that pulmonologists are actively involved in the practice of sleep medicine and that they both need and desire formal training in sleep disorders during pulmonary fellowship training. CONCLUSIONS: Participants were actively involved in the practice of sleep medicine, most had trained informally, and performance on questions about nonpulmonary sleep disorders was not good. PMID- 10858391 TI - Nasal continuous positive airway pressure use in children with obstructive sleep apnea younger than 2 years of age. AB - STUDY OBJECTIVES: To assess the efficacy of continuous positive airway pressure (CPAP) in obstructive sleep apnea (OSA) patients who are < 2 years of age. DESIGN: A retrospective chart review of 18 patients from 1992 to 1999 who had OSA confirmed by polysomnography. All patients in this study also completed a separate night of CPAP polysomnography to determine the effectiveness of CPAP in the correction of OSA. Nasal CPAP compliance data were gathered via clinical follow-up examination, telephone interview, or mailed questionnaire. SETTING: All patients were studied in the Sleep Disorders Center at Loma Linda University Children's Hospital in Loma Linda, CA. PATIENTS: All patients were < 2 years old. INTERVENTION: After OSA was confirmed by the results of technician-attended nocturnal polysomnography, separate technician-attended nocturnal CPAP polysomnography was completed. On CPAP nights, CPAP pressure was titrated to ameliorate OSA and snoring. CPAP pressure was increased by 2-cm H(2)O or 1-cm H(2)O increments. RESULTS: Data were analyzed by dependent groups t test at p < 0.05 level of significance. CPAP statistically improved respiratory parameters significantly when compared to baseline polysomnography. The following four patient subgroups emerged from the analysis: group 1 consisted of six patients who had tracheostomies prior to the CPAP trial, with two patients using CPAP as an alternative to tracheostomy; group 2 consisted of two patients who had previous unsuccessful adenostonsillectomies and who used CPAP successfully, with both having OSA resolution over time; group 3 consisted of four patients who did not tolerate CPAP on the study night; and group 4 consisted of six patients who used CPAP nightly, had OSA resolution over time, and therefore, no longer needed CPAP therapy. Thus, 10 of 18 patients used CPAP either on an interim basis for corrective therapy or as a primary treatment modality for OSA. CONCLUSIONS: These data show that children < 2 years of age can tolerate and use CPAP effectively. In several cases, CPAP treatment could be discontinued as OSA resolved over time. The reasons for this are discussed in the text. PMID- 10858392 TI - Validation of the POLY-MESAM seven-channel ambulatory recording unit. AB - STUDY OBJECTIVES: To determine the recording capabilities of the POLY-MESAM (PM) unit (MAP; Martinsried, Germany), an American Sleep Disorders Association level III system, and to compare it with simultaneous 12-channel polysomnography in the sleep laboratory. MEASUREMENTS AND RESULTS: Fifty-three patients (49 men and 4 women) with obstructive sleep-related breathing disorders of varying severity were included. The apnea-hypopnea indexes (AHIs) obtained using the two methods differed significantly from each other, although the correlation was close. The PM unit produced false-negative results in patients with mild to moderate obstructive sleep apnea (OSA). The sensitivity of the PM unit in detecting patients with an AHI > 10 was 92%, while the specificity was 96.3%. CONCLUSIONS: The correlation of AHIs obtained with polysomnography and with the PM unit is close. However, in some cases, the PM may underestimate OSA parameters. The PM unit produces false-negative results in patients with mild to moderate OSA. While inpatient polysomnography remains the "gold standard," the PM unit may provide an inexpensive alternative in some special cases. PMID- 10858393 TI - Cyclophosphamide in the treatment of idiopathic pulmonary fibrosis: a prospective study in patients who failed to respond to corticosteroids. AB - STUDY OBJECTIVES: To prospectively examine the role of cyclophosphamide in patients with idiopathic pulmonary fibrosis that is unresponsive to or intolerant of high-dose steroid treatment. DESIGN: Prospective study. SETTING: Tertiary referral center. PATIENTS: Nineteen patients with biopsy specimen-proven usual interstitial pneumonia who failed to respond (n = 16) or experienced adverse effects (n = 3) from corticosteroid treatment (1 mg/kg/d for 3 months). INTERVENTION: Steroid therapy was tapered quickly, and oral cyclophosphamide, 2 mg/kg/d, was prescribed (mean duration of treatment, 6.0 +/- 0.9 months). MEASUREMENTS AND RESULTS: In 10 patients, response to therapy was determined by pretreatment and posttreatment clinical (dyspnea), radiographic (chest radiograph), and physiologic (pulmonary function, including exercise saturation) scores (CRP). Response was defined as a > 10-point drop in CRP; stable as +/- 10 point change in CRP; and nonresponders as > 10-point rise in CRP. In nine patients, physiologic criteria were used to assess response; significant changes in pulmonary function were defined as follows: total lung capacity, +/- 10% of baseline value; FVC, +/- 10% of baseline value, diffusion capacity of the lung for carbon monoxide, +/- 20% of baseline value; and resting pulse oximetry, +/- 4% of baseline value. Patients who died while receiving or shortly after discontinuing cyclophosphamide were classified as nonresponders (n = 2). Among 19 patients treated with cyclophosphamide, only 1 patient demonstrated sustained response; 7 patients remained stable and 11 deteriorated while receiving the drug. Toxicity associated with cyclophosphamide was substantial; more than two thirds of the patients developed drug-related adverse effects, and almost half discontinued the drug prematurely due to side effects. In the remaining patients, cyclophosphamide therapy was discontinued due to lack of improvement or progressive deterioration. CONCLUSIONS: Cyclophosphamide therapy is of limited efficacy in patients with idiopathic pulmonary fibrosis who fail to respond or who experience adverse effects from corticosteroid treatment, and adverse effects often complicate its use. PMID- 10858394 TI - Evaluation of the short-form 36-item questionnaire to measure health-related quality of life in patients with idiopathic pulmonary fibrosis. AB - OBJECTIVE: To validate the use of the 36-item short-form questionnaire (SF-36) for measuring health-related quality of life (HRQL) in patients with idiopathic pulmonary fibrosis (IPF). DESIGN: : Observational data at a single point in time. SETTING: : A specialized outpatient respiratory clinic. PARTICIPANTS: Thirty-four patients (mean +/- SE age, 58.29 +/- 1.87 years) with IPF and no significant comorbidity. A matched control group for HRQL measurements was composed of 34 normal subjects (mean age, 58.00 +/- 1.89 years). MEASUREMENTS AND RESULTS: Dyspnea was measured by the baseline dyspnea index (BDI). Respiratory function evaluation included FVC, FEV(1), and resting arterial blood gases. IPF patients showed a mean BDI score of 5.21 +/- 0.46. The mean FVC and FEV(1) values were 62.41 +/- 2.96% and 66.41 +/- 3.33%, respectively. The mean PaO(2) was 67 +/- 2.51 mm Hg, and the mean PaCO(2) was 37 +/- 1. 05 mm Hg. Patients scored significantly worse than control subjects with respect to the SF-36 domains of physical functioning, physical role, general health perceptions, vitality, social functioning, emotional role, and mental health index. BDI scores were significantly correlated with five SF-36 components, and FVC and FEV(1) were significantly correlated with two SF-36 components. Significant negative correlations were found between arterial pH and four SF-36 domains. CONCLUSIONS: Patients with IPF have a significant impairment of HRQL in both physical and psychological functioning. Dyspnea is the most important factor influencing the quality of life in these subjects. The SF-36 questionnaire is a valid instrument to evaluate HRQL in IPF patients. PMID- 10858395 TI - Inhaled corticosteroids in stable COPD patients: do they have effects on cells and molecular mediators of airway inflammation? AB - STUDY OBJECTIVE: To investigate possible changes in cells and molecular mediators of airway inflammation following inhaled steroid treatment of stable COPD patients. DESIGN: Six-week open preliminary prospective study. SETTING: A university respiratory disease clinic. PATIENTS: : Stable COPD patients with mild disease. INTERVENTION: Six-week treatment with inhaled beclomethasone (1.5 mg die). MEASUREMENTS: The levels of interleukin (IL)-8, myeloperoxidase, eosinophilic cationic protein and tryptase, and cell numbers in bronchial lavage specimens were determined, and the symptom score, the endoscopic bronchitis index, and functional parameters were recorded. RESULTS: After treatment there were significant reductions in the lavage levels of IL-8 ([mean +/- SEM] 1,603.4 +/- 331.2 vs 1,119.2 +/- 265.3 pg/mL, respectively; p = 0. 01) and myeloperoxidase (1,614.5 +/- 682.3 vs 511.2 +/- 144.2 microg/L, respectively; p = 0.05), in cell numbers (250.6 +/- 27.7 vs 186.3 +/- 11.5 cells x 10(3)/mL, respectively; p = 0.04), neutrophil proportion (59.7 +/- 14.3% vs 31.5 +/- 10.1%; p = 0.01), symptom score (4.5 +/- 0.6 vs 1.4 +/- 0.5; p = 0.01), and bronchitis index (8.5 +/- 0.8 vs 5.5 +/- 0.7; p = 0.007). CONCLUSIONS: In stable patients with COPD, inhaled steroid treatment may induce changes on some cellular and molecular parameters of airway inflammation. PMID- 10858396 TI - Relationship of sputum color to nature and outpatient management of acute exacerbations of COPD. AB - STUDY OBJECTIVES: To stratify COPD patients presenting with an acute exacerbation on the basis of sputum color and to relate this to the isolation and viable numbers of bacteria recovered on culture. DESIGN: Open, longitudinal study of sputum characteristics and acute-phase proteins. SETTING: Patients presenting to primary-care physicians in the United Kingdom. Patients were followed up as outpatients in specialist clinic. PATIENTS: One hundred twenty-one patients with acute exacerbations of COPD were assessed together with a single sputum sample on the day of presentation (89 of whom produced a satisfactory sputum sample for analysis). One hundred nine patients were assessed 2 months later when they had returned to their stable clinical state. INTERVENTIONS: The expectoration of green, purulent sputum was taken as the primary indication for antibiotic therapy, whereas white or clear sputum was not considered representative of a bacterial episode and the need for antibiotic therapy. RESULTS: A positive bacterial culture was obtained from 84% of patients sputum if it was purulent on presentation compared with only 38% if it was mucoid (p < 0.0001). When restudied in the stable clinical state, the incidence of a positive bacterial culture was similar for both groups (38% and 41%, respectively). C-reactive protein concentrations were significantly raised (p < 0.0001) if the sputum was purulent (median, 4.5 mg/L; interquartile range [IQR], 6. 2 to 35.8). In the stable clinical state, sputum color improved significantly in the group who presented with purulent sputum from a median color number of 4.0 (IQR, 4.0 to 5.0) to 3.0 (IQR, 2.0 to 4. 0; p < 0.0001), and this was associated with a fall in median C reactive protein level to 2.7 mg/L (IQR, 1.0 to 6.6; p < 0.0001). CONCLUSIONS: The presence of green (purulent) sputum was 94.4% sensitive and 77.0% specific for the yield of a high bacterial load and indicates a clear subset of patient episodes identified at presentation that is likely to benefit most from antibiotic therapy. All patients who produced white (mucoid) sputum during the acute exacerbation improved without antibiotic therapy, and sputum characteristics remained the same even when the patients had returned to their stable clinical state. PMID- 10858397 TI - Interactions of regional respiratory mechanics and pulmonary ventilatory impairment in pulmonary emphysema: assessment with dynamic MRI and xenon-133 single-photon emission CT. AB - STUDY OBJECTIVES: Dynamic MRI and (133)Xe single-photon emission CT (SPECT) were used to directly evaluate the interaction of regional respiratory mechanics and lung ventilatory function in pulmonary emphysema. METHODS: Respiratory diaphragmatic and chest wall (D/CW) motions were analyzed by sequential MRI of fast-gradient echo pulse sequences during two to three respiratory cycles in 28 patients with pulmonary emphysema, including 9 patients undergoing lung volume reduction surgery (LVRS). The extent of air trapping in the regional lung was quantified by the (133)Xe retention index (RI) on three-dimensional (133)Xe SPECT displays. RESULTS: By contrast to healthy subjects (n = 6) with regular, synchronous D/CW motions, pulmonary emphysema patients showed reduced, irregular, or asynchronous motions in the hemithorax or location with greater (133)Xe retention, with significant decreases in the maximal amplitude of D/CW motions (MADM and MACWM; p < 0.0001 and p < 0.05, respectively). The removal of (133)Xe retention sites by LVRS effectively and regionally improved D/CW motions in nine patients, with significant increases in MADM and MACWM (p < 0.01 and p < 0.001, respectively). In a total of 40 studies of the 28 patients including post-LVRS studies, normalized MADM and MACWM correlated with percent predicted FEV(1) (r = 0.881, p < 0.0001; and r = 0.906, p < 0.0001, respectively), and also with (133)Xe RI in each hemithorax (r = -0.871, p < 0 0.0001; and r = -0.901, p < 0 0.0001, respectively.) CONCLUSIONS: This direct comparison of regional respiratory mechanics with lung ventilation demonstrated a close interaction between these impairments in pulmonary emphysema. The present techniques provide additional sensitivity for evaluating pathophysiologic compromises in pulmonary emphysema, and may also be useful for selecting resection targets for LVRS and for monitoring the effects. PMID- 10858398 TI - International comparison of median age at death from cystic fibrosis. AB - STUDY OBJECTIVES: To compare international trends in mortality from cystic fibrosis. DESIGN: Comparison of trends in median age at death using national mortality data. SETTING: Data from 10 countries in North America, Europe, and Australasia. PARTICIPANTS: All persons registered as having died of cystic fibrosis in specified years from 1980 to 1994. INTERVENTIONS: Comparison of relative odds of death at the international median age at death for the year of death between countries for two periods of time; from 1980 to 1987 (10 countries) and from 1980 to 1994 (7 countries). MEASUREMENTS AND RESULTS: The international median age at death increased from 8 years in 1974 to 21 years in 1994. Median age at death also increased within all countries, was consistently highest in the United States, and varied significantly by a factor of > twofold between countries. Women were significantly more likely to die at a younger age than the median age at death than men. CONCLUSIONS: Median age at death from cystic fibrosis is increasing, but our findings imply that clinically significant differences in survival with cystic fibrosis persist between countries. PMID- 10858399 TI - Misidentification of Burkholderia cepacia in US cystic fibrosis treatment centers: an analysis of 1,051 recent sputum isolates. AB - BACKGROUND: Burkholderia cepacia remains a significant pathogen in persons with cystic fibrosis (CF). The medical and psychosocial consequences of pulmonary colonization with this bacterium are enormous. However, B cepacia may be frequently misidentified from CF sputum culture. STUDY OBJECTIVES: To determine the rate of misidentification of B cepacia recently recovered from CF sputum culture of persons receiving care in US treatment centers. DESIGN: Bacterial isolates cultured from CF sputum and putatively identified as B cepacia or other related nonlactose-fermenting Gram-negative species were referred from participating treatment centers. Isolates underwent polyphasic analyses employing phenotypic (selective media and biochemical testing) and genotypic (polymerase chain reaction) assays to determine species identification. Taxonomic evaluations were performed by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins and amplified-fragment length polymorphism analysis. MEASUREMENTS AND RESULTS: A total of 1,051 isolates recovered from 608 patients were received from 115 treatment centers in 91 US cities. Among the isolates identified as B cepacia by referring laboratories, 11% could not be confirmed as B cepacia by polyphasic analyses. In addition, 36% of isolates not specifically identified by the referring laboratory or identified as a species other than B cepacia were, in fact, found to be members of the B cepacia complex. CONCLUSIONS: Rates of misidentification of B cepacia remain unacceptably high among US treatment centers. These data suggest the need for increased awareness of this problem among CF centers and their affiliated laboratories, better adherence to recommended protocols for evaluation of CF sputum, and greater use of reference laboratories equipped to provide advanced analyses. PMID- 10858400 TI - Reliability, repeatability, and sensitivity of the modified shuttle test in adult cystic fibrosis. AB - STUDY OBJECTIVES: The purpose of this study was to investigate the test-retest reliability, repeatability, and sensitivity of the modified shuttle test (MST) in adult patients with cystic fibrosis (CF). DESIGN: : Prospective study. SETTING: Adult CF Unit, Belfast City Hospital. PATIENTS: : Adult patients with CF. INTERVENTIONS: Test-retest reliability-none; sensitivity-inpatient IV antibiotic therapy for an acute exacerbation of respiratory disease. MEASUREMENTS: The test retest reliability and repeatability of the MST was assessed by comparing performance on two consecutive MSTs performed in 12 patients with CF and stable disease. The sensitivity of the MST was assessed by measuring the change in MST performance after 2 weeks of IV antibiotic therapy in 24 patients admitted to hospital with acute exacerbations of their respiratory disease. RESULTS: In the assessment of test-retest reliability and repeatability (n = 12), there was a significant and strong correlation between trials for distance completed (Pearson's r = 0. 99; p < 0.01), peak heart rate (Pearson's r = 0.99; p < 0.01), peak arterial oxygen saturation (SaO(2); Pearson's r = 0.99; p < 0.01), and peak Borg rating of perceived breathlessness (Pearson's r = 0. 99; p < 0.01). The coefficients of repeatability for these variables were small (coefficient of repeatability: distance completed, 4 shuttles; peak heart rate, 6 beats/min; peak SaO(2), 4%; and peak Borg rating of perceived breathlessness, 0.9). In the assessment of sensitivity (n = 24), the standardized response mean (SRM) for distance completed on MST (SRM = 1.18) was the SRMs for spirometric measures of lung function (FEV(1), SRM = 0.96; FEV(1) percent predicted, SRM = 0.88). CONCLUSIONS: This study demonstrates that the MST is a reliable, repeatable, and sensitive measure of exercise capacity in adult CF. The MST may be of value in determining prognosis, evaluation for lung transplantation, exercise prescription, and establishing the impact of new treatments on the disability associated with CF. PMID- 10858401 TI - Fungal empyema thoracis: an emerging clinical entity. AB - STUDY OBJECTIVES: To analyze the clinical spectra, pathogenesis, treatment, outcome, and prognostic factors of fungal empyema thoracis. DESIGN: The medical records of patients with positive fungal cultures from pleural effusions were retrospectively analyzed. SETTING: A university-based tertiary care hospital in Taipei, Taiwan. PATIENTS AND METHODS: From January 1990 through December 1997, patients diagnosed with fungal empyema were included in this study. The criteria for diagnosis of fungal empyema thoracis were as follows: (1) isolation of a fungal species from the pleural effusion; (2) significant signs of infection, such as fever (body temperature > 38.3 degrees C) and leukocytosis (white blood cell > 10,000/microL); and (3) isolation of the same mold species from pleural effusion on more than one occasion, or from pleural effusion and other specimens such as blood, sputum, or surgical wounds that showed evidence of tissue invasion. RESULTS: Sixty-seven patients with fungal empyema thoracis were included. Their mean age was 54 years (range, 2 weeks to 93 years), and 64% (43 patients) were men. Fifty-seven patients (85%) had various underlying diseases, and 18 (27%) had more than one immunocompromising condition. A total of 73 fungal isolates were recovered from pleural effusion; the most commonly encountered were Candida species (47 isolates, 64%), Torulopsis glabrata (13 isolates, 18%), and Aspergillus species (9 isolates, 12%). Candida albicans (28 isolates) was the most common Candida species, followed by Candida tropicalis (13 isolates). Six patients (9%) had two fungal strains isolated, and 16 (24%) had concomitant bacterial empyema thoracis. Eighteen patients (27%) had concurrent fungemia. Most (56 patients, 84%) cases of fungal empyema thoracis were nosocomial, and many case (43 patients, 64%) were acquired in ICUs. Abdominal disease (20 patients, 30%), especially previous abdominal surgery and GI perforation (12% and 10%, respectively), was the most common cause of fungal empyema thoracis, followed by bronchopulmonary infection (15 patients, 22%) and chest surgery (12 patients, 18%). Forty-nine patients (73%) received systemic antifungal therapy, and 38 (57%) underwent closed drainage therapy. Eleven patients (16%) underwent pleural irrigation with normal saline solution, povidone-iodine solution, or antifungal agents. Six patients (9%) finally received decortication. All patients receiving surgery or pleural irrigation with antifungal agents survived. Despite the aforementioned management, the crude mortality was high (73%). Multivariate analysis showed a significantly increased risk of death in immunocompromised patients (relative risk, 1.58; p < 0.005) and those with respiratory failure (relative risk, 2.31; p < 0.001). Systemic antifungal therapy was associated with a significantly lower risk of death (relative risk, 0.69; p < 0.05). CONCLUSION: These data imply an increasing incidence of fungal empyema thoracis in recent years and the necessity for aggressive treatment of patients with this disease. PMID- 10858402 TI - Decreased apoptosis and increased activation of alveolar neutrophils in bacterial pneumonia. AB - STUDY OBJECTIVES: The central role of apoptosis in the regulation of lung inflammation is increasingly recognized. The aim of this study was to determine the parameters of cell activation and apoptosis on neutrophils from the circulation and the pulmonary compartment in patients with community-acquired pneumonia (CAP), and to assess the role of the Fas system and of complement regulating molecules in this context. DESIGN AND METHODS: The study population consisted of nine patients with CAP (group 1) and six age-matched control patients without evidence of bronchopulmonary inflammation (group 2). Apoptosis rate and expression of CD11b, CD16, CD55, CD59, CD95, and CD114 surface molecules on systemic and bronchoalveolar neutrophils were assessed ex vivo using fluorescence-activated cell sorter analysis. RESULTS: In patients with CAP, we found a significant decrease of the mean apoptosis rate in pulmonary neutrophils compared to systemic neutrophils, without concomitant changes in Fas expression. In contrast, cell activation markers were significantly increased on pulmonary cells (CD11b, 288 +/- 98.2 relative mean fluorescence intensity [rMFI] vs 53.8 +/ 10.8 rMFI on peripheral cells), and similar changes were observed with respect to the expression of complement-regulating molecules. Pulmonary polymorphonuclear neutrophils of the control group showed analogous changes, compared to systemic neutrophils, but a significantly higher rate of apoptosis and a lower increase of activation-marker expression were found, compared to pulmonary neutrophils of patients with pneumonia. CONCLUSIONS: Pulmonary neutrophils from patients with CAP show a decreased rate of apoptosis and increased activation status in the alveolar compartment, which may be important for effective control of pulmonary inflammation. PMID- 10858403 TI - A 10-year experience with bacteriology of acute thoracic empyema: emphasis on Klebsiella pneumoniae in patients with diabetes mellitus. AB - STUDY OBJECTIVES: To provide an updated evaluation of the bacteriology of acute thoracic empyema for more efficacious treatment. DESIGN: : The medical and microbiological records of all patients who received a diagnosis of acute thoracic empyema were reviewed. Based on the bacteria isolated from the pleural fluid, the patients were classified into the following four groups: aerobic or facultative Gram-positive; aerobic Gram-negative; anaerobic; and mixed. SETTING: A university-affiliated tertiary medical center. PATIENTS AND METHODS: From January 1989 to December 1998, 171 patients with a diagnosis of acute thoracic empyema were treated. A comparative analysis of the isolates from pleural effusions, the mean length of hospital stay, the mean duration of chest tube drainage, the mean duration between the onset of symptoms and the establishment of diagnosis, treatment efficacy, and the need for subsequent intervention was performed. RESULTS: A total of 163 microorganisms were isolated from the pleural fluid of 139 patients. These patients were classified according to the following types of isolates: aerobic or facultative Gram-positive (n = 47); aerobic Gram negative (n = 59); anaerobic (n = 14); and mixed (n = 19). Klebsiella pneumoniae was the most commonly isolated pathogen (24. 4%) and was strongly associated with a diagnosis of diabetes mellitus. The mortality rate of patients with aerobic Gram-negative bacilli isolated was the highest (22.0%), followed by those with mixed pathogens isolated (15.7%), aerobic or facultative Gram-positive (6.4%), and anaerobic (0%). CONCLUSIONS: The increasing incidence of acute thoracic empyema caused by Gram-negative bacilli, especially by K pneumoniae, has become an increasing problem. The isolation of aerobic Gram-negative bacilli or multiple pathogens from pleural fluid is associated with a poor prognosis and indicates a need for more aggressive antimicrobial chemotherapy. PMID- 10858404 TI - Prospective randomized trial comparing pressure-controlled ventilation and volume controlled ventilation in ARDS. For the Spanish Lung Failure Collaborative Group. AB - STUDY OBJECTIVES: To compare in-hospital mortality of patients with ARDS ventilated with either pressure-controlled ventilation (PCV) or volume-controlled ventilation (VCV) with a square-wave inspiratory flow. DESIGN: : Multicenter and randomized trial. SETTING: Twelve medical-surgical ICUs located in tertiary-care hospitals. PATIENTS: Seventy-nine patients having ARDS, as defined by the American-European Consensus Conference. INTERVENTIONS: Patients were randomly assigned to be ventilated with either PCV (n = 37) or VCV (n = 42). In both instances, inspiratory plateau pressure was limited to < or = 35 cm H(2)O. MEASUREMENTS AND RESULTS: There were no significant differences among the studied groups at the moment of randomization, although there was a trend toward greater renal failure in patients assigned to VCV. Ventilatory settings and blood gases did not significantly differ over time between the two groups. Patients in the VCV group had both a significantly higher in-hospital mortality rate than those in the PCV group (78% vs 51%, respectively) and a higher number of extrapulmonary organ failures (median, 4 vs 2, respectively). The development of renal failure during the study period was also significantly more frequent among VCV patients (64% vs 32%, respectively). Multivariate analysis showed that factors independently associated with an increased mortality rate were the presence of two or more extrapulmonary organ failures (odds ratio [OR], 4.61; 95% confidence interval [CI], 1.38 to 15.40) and acute renal failure (OR, 3.96; 95% CI, 1.10 to 14.28) but not the ventilatory mode used. CONCLUSIONS: The increased number of extrapulmonary organ failures developed in patients of the VCV group was strongly associated with a higher mortality rate. The development of organ failures was probably not related to the ventilatory mode. PMID- 10858405 TI - Efficacy of chest CT in a pediatric ICU: a prospective study. AB - STUDY OBJECTIVES: (1) To determine whether chest CT provides additional information compared with chest radiography regarding the nature of intrathoracic disease in critically ill children, (2) to determine whether such information alters clinical management, (3) to assess the role of a low-dose high-resolution CT (HRCT) protocol in pediatric ICU (PICU) patients. DESIGN: Prospective study. SETTING: Specialized PICU in a teaching hospital serving London and the south of England. PATIENTS: Twenty children (age range, 3 weeks to 12 years; median, 11 months) underwent chest CT during a 33-month period. Inclusion criteria were (1) inconclusive diagnosis from chest radiograph (CXR) or (2) CXR appearances inconsistent with high oxygenation or ventilatory requirements (PaO(2) to fraction of inspired oxygen ratio < 30 or mean airway pressure > 15 cm H(2)O). INTERVENTIONS: Low-dose HRCT scans (50 mA, 2-mm slice thickness at intervals of 10 or 15 mm) were performed on 12 patients, and helical CT (50 to 250 mA; pitch, 1 to 1.5) performed on 8 patients. MEASUREMENTS AND RESULTS: CT provided additional information regarding the nature of intrathoracic disease in 17 of 20 patients (85%) and resulted in changes to subsequent clinical management in 12 of 20 patients (60%). CONCLUSIONS: Chest CT can add to the accuracy of intrathoracic diagnosis provided by the CXR and may directly influence the acute management of critically ill children. The CT protocol should be tailored to the clinical and radiologic question posed for each individual patient. Noncontiguous HRCT can often provide accurate assessment of pulmonary parenchymal and pleural disease at a reduced radiation dose compared with helical CT. PMID- 10858406 TI - Preoperative and postoperative endotoxemia in children with congenital heart disease. AB - STUDY OBJECTIVES: Recent data indicate that increases in inflammatory cytokines are seen in patients with diverse cardiac diseases. However, the primary stimulus for cytokine secretion during cardiac illness remains unknown. Since bacterial endotoxin is a potent inducer of cytokines, we determined the incidence, magnitude, and clinical relevance of endotoxemia in children with congenital heart disease before and after surgical repair. DESIGN: A prospective, observational study. SETTING: A large, urban, university-affiliated, tertiary care children's hospital. PATIENTS: Thirty children with a variety of congenital heart defects (median age, 59 days; median weight, 4.0 kg) were sequentially enrolled. INTERVENTIONS: Blood was sampled prior to surgery, and at 1, 8, 24, 48, and 72 h following cardiopulmonary bypass. Assays included plasma endotoxin, lipopolysaccharide-binding protein (LBP), and interleukin-6 (IL-6). MEASUREMENTS AND RESULTS: Twenty-nine of 30 patients (96%) had evidence of endotoxemia during the study period. Twelve of the 30 patients (40%) were significantly endotoxemic prior to surgery. LBP, a plasma marker that responds to bacteria and endotoxin, rose significantly following cardiopulmonary bypass, as did the plasma levels of IL-6. Fifteen of 30 patients met prospectively defined criteria for experiencing a severe hemodynamic disturbance in their postoperative course. These patients had significantly higher preoperative plasma LBP (p < 0.02) and plasma endotoxin levels (p < 0.05), compared to patients with less-severely disturbed hemodynamics. Mortality was 25% in patients with preoperative endotoxemia, compared with no mortality in patients who were not endotoxemic before surgery (p = 0.05). CONCLUSIONS: These data demonstrate that endotoxemia in children with congenital heart disease is more common than previously suspected, and is associated with clinical outcomes. We conclude that clinical trials targeting endotoxin will be necessary to determine if endotoxin is a causal, etiologic agent in the disease process. PMID- 10858407 TI - Prognostic value of hemodynamic vs big endothelin measurements during long-term IV therapy in advanced heart failure patients. AB - STUDY OBJECTIVE: To compare hemodynamics and plasma big endothelin levels in patients awaiting heart transplantation who are receiving continuous IV therapy, and to establish their respective potency for predicting future cardiac events. DESIGN: A randomized, prospective trial of ambulatory continuous treatment with IV prostaglandin E(1) (PGE(1)) vs dobutamine. A subanalysis was conducted of all patients who completed 4 weeks of follow-up in regard to treatment effects on hemodynamics and big endothelin plasma levels. PATIENTS: Thirty-two listed heart transplant candidates who were refractory to oral treatment, 21 patients who were receiving PGE(1), and 11 patients receiving dobutamine. MEASUREMENTS AND RESULTS: Hemodynamics and plasma big endothelin levels were measured at baseline and after 4 weeks. The cardiac index increased significantly (PGE(1) group, 1.7 +/- 0.4 vs 2.5 +/- 0.6 L/min/m(2); dobutamine group, 1.8 +/- 0.3 vs 2.3 +/- 0.6 L/min/m(2); p < 0.05), whereas the systemic vascular resistance index (SVRI) decreased significantly only in the PGE(1) group (3,352 +/- 954 vs 2,178 +/- 519 dyne. s. cm(-5)/m(2); p < 0. 05). The plasma big endothelin level decreased significantly (PGE(1) group, 7.6 +/- 3.1 vs 4.7 +/- 2.6 fmol/mL; dobutamine group, 6.5 +/- 3.7 vs 5.0 +/- 2.6 fmol/mL; p < 0.01 for the time effect). Plasma big endothelin (beta = 0.393; chi(2) = 10.8; p = 0.001) and SVRI (beta = 0.003; chi(2) = 6.9; p < 0.01), both measured after 4 weeks of continuous treatment, were the only independent predictors of future outcome. CONCLUSION: Continuous treatment over 4 weeks with either PGE(1) or dobutamine in patients awaiting heart transplantation yields an improved hemodynamic state accompanied by a reduction of increased big endothelin levels. Plasma big endothelin measured after 4 weeks of continuous therapy provides prognostic information about future outcome. PMID- 10858408 TI - Effects of eicosapentaenoic and gamma-linolenic acids (dietary lipids) on pulmonary surfactant composition and function during porcine endotoxemia. AB - STUDY OBJECTIVES: To investigate whether a diet enriched with fish and borage oils, with their high polyunsaturated fatty acid (PUFA) content, alters surfactant composition and function during endotoxemia. DESIGN: Prospective, randomized, blinded, controlled animal study. SETTING: Research laboratory at a medical center. PARTICIPANTS: Thirty-six 15- to 25-kg, disease-free, castrated male pigs. DIETS AND MEASUREMENTS: Three groups of pigs (n = 12 per group) were fed for 8 days diets containing either omega-6 fatty acids (FAs) (corn oil; diet A), or omega-3 FAs (fish oil; diet B), or a combination of omega-6 and omega-3 FAs (borage and fish oils; diet C). Eight of 12 pigs in each group received a 0.1 mg/kg bolus of Escherichia coli endotoxin followed by a continuous infusion (0. 075 mg/kg/h). One lung was subsequently isolated ex vivo, and pressure-volume curves were measured. The contralateral lung was lavaged, and surfactant was analyzed for total and individual phospholipids and FA composition. Minimum and maximum surface tension was measured by bubble surfactometry. RESULTS: Pigs fed either diet B or C had increased oleic acid (C(18:1) omega-9), eicosapentaenoic acid (EPA; C(20:5) omega-3), docosahexaenoic acid (C(22:6) omega-3), and total omega-3 and monounsaturated FAs in their surfactant PUFA pools. The relative percentage of linoleic acid (C(18:2) omega-6) and total omega-6 FAs were significantly lower from pigs fed diets B and C compared with diet A. Palmitic acid (C(16:0)) concentrations, the primary FA in surfactant, had a tendency to be lower in pigs fed diets B and C. There were no demonstrable effects on surfactant function or pulmonary compliance. CONCLUSIONS: Diets containing EPA or EPA and gamma-linolenic acid altered the PUFA composition of pulmonary surfactant, but without demonstrable effects on surfactant function during porcine endotoxemia. PMID- 10858409 TI - Effects of streptokinase and deoxyribonuclease on viscosity of human surgical and empyema pus. AB - STUDY OBJECTIVE: To investigate the effects of streptokinase and deoxyribonuclease (DNase) on the viscosity of pus to assess whether the DNase in the old preparation of streptokinase-streptodornase used intrapleurally to treat empyema was contributing to easier drainage of pus compared with purified streptokinase. DESIGN: In vitro measurement of pus viscosity. PATIENTS: Pus from three patients with surgically drained soft tissue abscesses and from six patients with empyema thoracis of varying etiology was studied. INTERVENTIONS: Pus samples were incubated with saline solution as control and with streptokinase, streptokinase-streptodornase, human recombinant DNase, and a mixture of streptokinase and DNase in concentrations approximating those achieved in clinical practice. RESULTS: Purified streptokinase had little effect on pus viscosity, with a mean reduction of 11.1% in the surgical specimens and 1.7% in the empyema samples. Streptokinase-streptodornase reduced viscosity by a mean of 52.8% in the surgical samples and 94.8% in the empyema samples. Human recombinant DNase reduced viscosity by a mean of 32. 79% in surgical samples and 93.4% in empyema samples. Adding streptokinase to human recombinant DNase produced no further reduction in viscosity. Final viscosities in samples treated with DNase were very similar whatever the starting viscosity. CONCLUSIONS: DNase significantly reduces pus viscosity, whereas streptokinase has little or no effect, and in empyema may work simply by breaking down loculations. Clinical studies should be undertaken to see if these in vitro changes produce clinical benefits. The simple viscometer devised for these experiments may also prove useful in other contexts. PMID- 10858410 TI - Antibiotic levels in empyemic pleural fluid. AB - OBJECTIVE: To determine the degree to which bioactive penicillin, metronidazole, ceftriaxone, clindamycin, vancomycin, and gentamicin penetrate into empyemic pleural fluid using our new rabbit model of empyema. METHODS: An empyema was created via the intrapleural injection of 10(8)()Pasteurella multocida bacteria into the pleural space of New Zealand white rabbits. After an empyema was verified by thoracentesis and pleural fluid analysis, penicillin, 24,000 U/kg; metronidazole, 37 mg/kg; ceftriaxone, 30 mg/kg; clindamycin, 9 mg/kg; vancomycin, 15 mg/kg; or gentamicin, 1 mg/kg, were administered IV. Antibiotic levels in samples of pleural fluid and serum, collected serially for up to 480 min, were then determined using a bioassay. RESULTS: The degree to which the different antibiotics penetrated into the infected pleural space was highly variable. Penicillin penetrated most easily, followed by metronidazole, ceftriaxone, clindamycin, vancomycin, and gentamicin. Of the antibiotics tested, penicillin and metronidazole equilibrated the most rapidly with the infected pleural fluid. Penicillin levels remained elevated in pleural fluid even after serum levels had decreased. CONCLUSIONS: Using this rabbit model of empyema, there was marked variation in the penetration of antibiotics into the empyemic fluid. Although there are species differences between rabbit and human pleura, the variance in degree of penetration of antibiotics into the pleural space should be considered when antibiotics are selected for the treatment of patients with empyema. PMID- 10858411 TI - Congenital malformations of the right atrium and the coronary sinus: an analysis based on 103 cases reported in the literature and two additional cases. AB - STUDY OBJECTIVES: Congenital malformations of the right atrium (RA) and the coronary sinus (CS) are rare, and only sporadic cases have been reported. Little is known about the clinical relevance of this disorder. We report on two patients, one with a giant RA diverticulum, the other with a diverticulum of the CS, and review 103 cases of such malformations that have been reported previously. DESIGN: A MEDLINE search was performed to collect all cases of congenital malformations of the RA and the CS reported in the literature between 1955 and 1998. Cases were classified into the following categories: (1) congenital enlargement of the RA; (2) single diverticulum of the RA; (3) multiple diverticula of the RA; and (4) diverticulum of the CS. Clinical presentation and outcome of the different types of malformations were analyzed. RESULTS: The patients most frequently presenting with symptoms were those with diverticula of the CS (n = 28) followed by those with single diverticula of the RA (n = 13), multiple diverticula (n = 4), and congenital enlargements of the RA (n = 60). The percentages of symptomatic patients were 93, 84, 75, and 53%, respectively. Symptoms were frequently caused by arrhythmias. Supraventricular tachycardia (SVT) was found in 42 of the patients (40%) and was most common in patients with diverticula of the CS (24 of 28 patients) and multiple atrial diverticula (3 of 4 patients). Sudden cardiac death was reported more frequently in patients with diverticula of the CS (18%) compared to those with congenital enlargement of the RA (5%) or single or multiple diverticula of the RA (6%). All seven patients with diverticula of the CS who were not treated with catheter or surgical ablation eventually died. CONCLUSION: Congenital malformations of the RA and the CS frequently are associated with arrhythmias. SVT and sudden cardiac death have been reported in a significant percentage of patients with diverticula of the CS. PMID- 10858412 TI - Negative fluid balance predicts survival in patients with septic shock: a retrospective pilot study. AB - OBJECTIVE: We hypothesized that patients with septic shock who achieve negative fluid balance (< or =-500 mL) on any day in the first 3 days of management are more likely to survive than those who do not. DESIGN: Retrospective chart review. PATIENTS: Thirty-six patients admitted with the diagnosis of septic shock. SETTING: Twelve-bed medical ICU of a 300-bed community teaching hospital. METHODS: Medical records of 36 patients admitted to our medical ICU over a 21 month period were examined. Patients with septic shock who required dialysis prior to hospitalization were not included. A number of demographic and physiologic variables were extracted from the medical records. Admission APACHE (acute physiology and chronic health evaluation) II and daily sequential organ failure assessment (SOFA) scores were computed from the extracted data. Variables were compared between survivors and nonsurvivors and in patients who did vs those who did not achieve negative (< or = 500 mL) fluid balance in > or = 1 day of the first 3 days of management. Survival risk ratios (RRs) were used as the measure of association between negative fluid balance and survival. RRs were adjusted for age, APACHE II scores, SOFA scores on the first and third days, and the need for mechanical ventilation, by stratified analyses. RESULTS: Patients ranged in age from 16 to 85 years with a mean (+/- SE) age of 67.4 +/- 3.3 years. The mean admission APACHE II score was 25.4 +/- 1.4, and the day 1 SOFA score was 9.0 +/- 0.8. Twenty patients did not survive; nonsurvivors had higher mean APACHE II scores than survivors (29.8 vs 20.4, respectively) and higher first day SOFA scores than survivors (10.8 vs 6.9, respectively), and they were more likely to require vasopressors and mechanical ventilation compared to patients who survived. Whereas all 11 patients who achieved a negative balance of > 500 mL on > or = 1 of the first 3 days of treatment survived, only 5 of 25 patient who failed to achieve a negative fluid balance of > 500 mL by the third day of treatment survived (RR, 5.0; 95% CI, 2.3 to 10.9; p = 0.00001). At least 1 day of net negative fluid balance in the first 3 days of treatment strongly predicted survival across the strata of age, APACHE II scores, first- and third-day SOFA scores, the need for mechanical ventilation, and creatinine levels measured at admission. CONCLUSION: These results suggest that at least 1 day of negative fluid balance (< or = -500 mL) achieved by the third day of treatment may be a good independent predictor of survival in patients with septic shock. These findings suggest the hypothesis "that negative fluid balance achieved in any of the first 3 days of septic shock portends a good prognosis," for a larger prospective cohort study. PMID- 10858413 TI - A European view on the North American fifth consensus on antithrombotic therapy. AB - An American-Canadian group of experts have, in the November 1998 issue of CHEST, published for the fifth time their recommendations for antithrombotic therapy. This remarkable consensus document was the result of an extensive review of the literature by an interdisciplinary group. Considering the impact of this document on medical practice, also outside North America, a group of European experts reviewed in detail the fifth report, particularly the sections on clinical indications of antithrombotic treatment. The aim was not to indicate the many areas of agreement and to quote literature that has become available since publication of the last consensus documents, but rather to refer to the gray zones of uncertainty and limited number of divergent opinions. PMID- 10858414 TI - Lung infections: role of apoptosis in host defense and pathogenesis of disease. AB - Apoptosis is a form of cell death that has gained enormous attention during the past few years, and its mechanisms, important to biology and medicine, are being unraveled at an accelerating pace. Apoptosis of lung cells occurs during lung infections and may be either a host defense mechanism or reflect the pathogenesis of the infection. In the first part of this review, the biochemistry and physiology of apoptotic pathways and its regulators are discussed. This is followed by an overview of apoptotic mechanisms in selected lung infections. The implications of apoptosis in host immunity, pathogenesis, and treatment of pulmonary infections will be discussed in this context. PMID- 10858415 TI - Pneumomediastinum in a 63-year-old woman with asthma exacerbation. PMID- 10858416 TI - A 66-year-old man with dyspnea, left lower lobe infiltrate, and abnormal imaging. PMID- 10858417 TI - Diagnosis and treatment of mediastinal tumors by thoracoscopy. AB - OBJECTIVES: Thoracoscopic management of mediastinal tumors is still subject to analysis. Seventy-three patients underwent thoracoscopy for treatment of mediastinal masses and were analyzed retrospectively in order to evaluate the effectiveness and complications of the procedure. METHODS: Between 1983 and 1999, 21 conventional thoracoscopies and 52 video-assisted thoracic surgeries were performed (33 for diagnostic purposes and 40 for therapy). Patient ages ranged from 2 to 81 years (mean, 43.8 years) with a slight predominance of girls and women over men and boys (41 vs 32, respectively). All patients underwent general anesthesia using simple intubation (22 patients) or double-lumen intubation (51 patients). RESULTS: The histologic type of tumors was obtained in all patients. For therapeutic purposes, a change of procedure to thoracotomy was necessary in nine patients. The reasons for this change were tumor size, tumor invasion of nearby structures, difficulties in continuing the dissection, the performance of an upper lobectomy, and suturing the iatrogenic lesion of the diaphragm. Four patients died during the first 30 postoperative days as a consequence of their primary pathology. CONCLUSION: Thoracoscopy was confirmed as an effective diagnostic and therapeutic alternative for the treatment of mediastinal disorders. PMID- 10858418 TI - Dermatomyositis as a presentation of pulmonary inflammatory pseudotumor (Myofibroblastic tumor). AB - Inflammatory pseudotumor (IPT) is a rare pulmonary tumor of uncertain etiology that usually presents as an asymptomatic radiographic finding. We describe a case of pulmonary IPT presenting as dermatomyositis with complete resolution following surgical resection. PMID- 10858419 TI - Laryngotracheobronchial involvement in a patient with nonendemic rhinoscleroma. AB - We report the first case of rhinoscleroma in an Israeli citizen, a former sailor with a transatlantic shipping company. Characteristic histologic changes from a tracheal biopsy and isolation of Klebsiella rhinoscleromatis from a blood culture after diagnostic bronchoscopy confirmed the diagnosis. Extreme delay in the diagnosis, a not uncommon feature in nonendemic areas, was associated with severe advanced laryngotracheobronchial disease. Treatment with quinolones was followed by significant improvement, but the patient died 1 month after presentation, apparently from upper airway obstruction. PMID- 10858420 TI - The acute effects of transvenous biventricular pacing in a patient with congestive heart failure. AB - The management of congestive heart failure remains an issue of great interest. Encouraging data emerged over the last 2 years supporting the use of multisite pacing in patients with severe congestive heart failure and intraventricular conduction delay. We present a case of acute biventricular pacing in a 81-year old man with dilated cardiomyopathy and symptomatic congestive heart failure. This novel form of pacemaker treatment resulted in a rapid hemodynamic and clinical improvement. PMID- 10858421 TI - A patient with syncope, only "vagally" related to the heart. AB - Syncope due to atrioventricular block may occur as a result of a cardiac vasodepressor reflex. This article reports a case of syncope in a 58-year-old man with high-grade atrioventricular block documented by ambulatory ECG monitoring at home. What makes this case unusual is that the patient's principal diagnosis was noncardiac. PMID- 10858422 TI - Symptomatic pericarditis after influenza vaccination: report of two cases. AB - The authors report two cases of benign acute pericarditis after the patients received vaccinations against influenza virus. The diagnoses were confirmed by serologic changes and by the findings of 12-lead electrocardiogram and echocardiography. Symptoms and clinical status improved on aspirin therapy. The authors underline the possible mechanisms of this rare complication of influenza vaccination. PMID- 10858423 TI - Single-breath measurements of pulmonary oxygen uptake and gas flow rates for ventilator management in ARDS. AB - Monitoring data in critical care and anesthesiology should be displayed to present a rapid and easily comprehensible definition of the patient's clinical status. A graphic computer display of the analog output of gas flow rates and the O(2) and CO(2) concentrations of respiratory gases profiles the expired breath for an estimation of pulmonary function and gas exchange. An estimate of pulmonary perfusion, cardiac output, and the general adequacy of cardiovascular circulation is obtained from the computer calculation of O(2) uptake and CO(2) elimination, dead space, and alveolar ventilation. Adjunctive data from the spirometric measurements of airway pressures, volumes, and compliance, supplemented by hemodynamic monitoring, aids in the diagnosis of physiologic changes. For > 10 years, we have used this system to monitor patients who are anesthetized, sedated, and receiving mechanical ventilation during anesthesia and surgery, and recently have extended the technique to intensive care areas. Our experience has shown good correlation of changes in the computer-assisted expired breath analysis with coinciding clinical events, including upper airway obstruction, bronchospasm, and alveolar volume/pulmonary capillary blood flow impairment. To demonstrate the use of this system, we describe the ventilator management for a patient with severe ARDS. In this patient, changes in ventilator management, including pressure control ventilation, improved pulmonary O(2) uptake (mean, 18.7 vs 8.5 mL/breath), CO(2) elimination (mean, 17 vs 13 mL/breath), and compliance (mean, 29.7 vs 19.0 mL/cm H(2)O), were compared with intermittent mandatory ventilation. PMID- 10858424 TI - Persistent pneumomediastinum in interstitial fibrosis associated with rheumatoid arthritis: treatment with high-concentration oxygen. AB - We present a case of persistent spontaneous pneumomediastinum precipitated by an upper respiratory infection in a patient with interstitial fibrosis associated with rheumatoid arthritis who was receiving chronic corticosteroid treatment. The persistent nature of the mediastinal emphysema over 2 months eventually required treatment with high concentrations of inhaled oxygen that resulted in rapid resolution of the pneumomediastinum without recurrence over 6 months of follow up. This case, along with others in the medical literature, emphasizes the need for early use of high-concentration inhaled oxygen in the treatment of pneumomediastinum in high-risk patients, such as those with connective tissue disorders. PMID- 10858425 TI - Pulmonary alveolar proteinosis in association with household exposure to fibrous insulation material. AB - We report the case of a 35-year-old woman who developed pulmonary alveolar proteinosis requiring multiple lavage treatments, in association with household exposure to ventilation system dust comprised at least partially by a cellulose fire-resistant fibrous insulation material. Scanning electron microscopy with energy-dispersive x-ray analysis documented the presence of spectral peaks consistent with the insulation material in transbronchial biopsy tissue. The patient showed symptomatic improvement once exposure to the insulation material had ceased. We believe that this case demonstrates an unusual association with pulmonary alveolar proteinosis. This case emphasizes the broad differential diagnosis for this histologic injury pattern and the need to thoroughly investigate environmental exposures in patients with unexplained pulmonary disease. PMID- 10858426 TI - Pathophysiology of a fall in arterial oxygen saturation during sputum induction. PMID- 10858427 TI - Angiotensin-converting enzyme inhibitors, angiotensin II receptor antagonists, and symptomless dysphagia. PMID- 10858428 TI - Electrocautery in endobronchial therapy. PMID- 10858429 TI - Corticosteroid therapy in acute asthma. PMID- 10858430 TI - IV epoprostenol in Gaucher's disease. PMID- 10858431 TI - Infusion phlebitis in patients in a general internal medicine service. PMID- 10858432 TI - Differences in percutaneous dilational tracheostomy kits. PMID- 10858433 TI - Metabolite profiling of sesquiterpene lactones from Lactuca species. Major latex components are novel oxalate and sulfate conjugates of lactucin and its derivatives. AB - Wounding leaves or stems of Lactuca species releases a milky latex onto the plant surface. We have examined the constituents of latex from Lactuca sativa (lettuce) cv. Diana. The major components were shown to be novel 15-oxalyl and 8-sulfate conjugates of the guaianolide sesquiterpene lactones, lactucin, deoxylactucin, and lactucopicrin. The oxalates were unstable, reverting to the parent sesquiterpene lactone on hydrolysis. Oxalyl derivatives have been reported rarely from natural sources. The sulfates were stable and are the first reported sesquiterpene sulfates from plants. Unusual tannins based on 4 hydroxyphenylacetyl conjugates of glucose were also identified. Significant qualitative and quantitative variation was found in sesquiterpene lactone profiles in different lettuce varieties and in other Lactuca spp. The proportions of each conjugate in latex also changed depending on the stage of plant development. A similar profile was found in chicory, in which oxalyl conjugates were identified, but the 8-sulfate conjugates were notably absent. The presence of the constitutive sesquiterpene lactones was not correlated with resistance to pathogens but may have a significant bearing on the molecular basis of the bitter taste of lettuce and related species. The induced sesquiterpene lactone phytoalexin, lettucenin A, was found in the Lactuca spp. but not in chicory. PMID- 10858434 TI - Dynamic interaction of human vasopressin/oxytocin receptor subtypes with G protein-coupled receptor kinases and protein kinase C after agonist stimulation. AB - Examination of the structure of [Arg(8)]-vasopressin receptors (AVPRs) and oxytocin receptors (OTRs) suggests that G protein-coupled receptor kinases (GRKs) and protein kinase C (PKC) are involved in their signal transduction. To explore the physical association of AVPRs and OTRs with GRKs and PKC, wild types and mutated forms of these receptor subtypes were stably expressed as green fluorescent protein fusion proteins and analyzed by fluorescence, immunoprecipitation, and immunoblotting. Addition of a C-terminal GFP tag did not interfere with ligand binding, internalization, and signal transduction. After agonist stimulation, PKC dissociated from the V(1)R, did not associate with the V(2)R, but associated with the V(3)R and the OTR. After AVP stimulation, only GRK5 briefly associated with AVPRs following a time course that varied with the receptor subtype. No GRK associated with the OTR. Exchanging the V(1)R and V(2)R C termini altered the time course of PKC and GRK5 association. Deletion of the V(1)R C terminus resulted in no PKC association and a ligand-independent sustained association of GRK5 with the receptor. Deletion of the GRK motif prevented association and reduced receptor phosphorylation. Thus, agonist stimulation of AVP/OT receptors leads to receptor subtype-specific interactions with GRK and PKC through specific motifs present in the C termini of the receptors. PMID- 10858435 TI - Macrophage lipoprotein lipase promotes foam cell formation and atherosclerosis in low density lipoprotein receptor-deficient mice. AB - The role of macrophage lipoprotein lipase (LPL) expression in atherosclerotic lesion formation was examined in low density lipoprotein receptor (LDLR(-/-)) mice using dietary conditions designed to induce either fatty streak lesions or complex atherosclerotic lesions. First, LDLR(-/-) mice chimeric for macrophage LPL expression were created by transplantation of lethally irradiated female LDLR(-/-) mice with LPL(-/-) (n = 12) or LPL(+/+) (n = 14) fetal liver cells as a source of hematopoietic cells. To induce fatty streak lesions, these mice were fed a Western diet for 8 weeks, resulting in severe hypercholesterolemia. There were no differences in plasma post-heparin LPL activity, serum lipid levels, or lipoprotein distribution between these two groups. The mean lesion area in the proximal aorta in LPL(-/-) --> LDLR(-/-) mice was significantly reduced by 33% compared with LPL(+/+) --> LDLR(-/-) mice, and a similar reduction (38%) in lesion area was found by en face analysis of the aortae. To induce complex atherosclerotic lesions, female LDLR(-/-) mice were lethally irradiated, transplanted with LPL(-/-) (n = 14), LPL(+/-) (n = 13), or LPL(+/+) (n = 14) fetal liver cells, and fed the Western diet for 19 weeks. Serum cholesterol and triglyceride levels did not differ between the three groups. After 19 weeks of diet, the lesions in the proximal aorta were complex with relatively few macrophages expressing LPL protein and mRNA in LPL(+/+) --> LDLR(-/-) mice. Analysis of cross-sections of the proximal aorta demonstrated no differences in the extent of lesion area between the groups, whereas en face analysis of the aortae revealed a dose-dependent effect of macrophage LPL on mean aortic lesion area in LPL(-/-) --> LDLR(-/-), LPL(-/+) --> LDLR(-/-), and LPL(+/+) --> LDLR(-/ ) mice (1.8 +/- 0. 2%, 3.5 +/- 0.5% and 5.9 +/- 0.8%, respectively). Taken together, these data indicate that macrophage LPL expression in the artery wall promotes atherogenesis during foam cell lesion formation, but this impact may be limited to macrophage-rich lesions. PMID- 10858436 TI - A key point mutation (V156E) affects the structure and functions of human apolipoprotein A-I. AB - A naturally occurring point mutant of human apolipoprotein A-I (apoA-I), V156E, which is associated with extremely low plasma apoA-I and high density lipoprotein (HDL) levels, and coronary artery disease (Huang, W., Sasaki, J., Matsunaga, A., Nanimatsu, H., Moriyama, K., Han, H. Kugi, M., Koga, T., Yamaguchi, K., and Arakawa, K. (1998) Arterioscler. Throm. Vasc. Biol. 18, 389-396), was produced in an Escherichia coli expression system. The purified recombinant proapoA-I V156E mutant was examined in its structural and functional properties, both, in the lipid-free and lipid-bound states. In the lipid-free form the mutant protein exhibited small changes in conformation, but was more stable, and quite resistant to self-association, compared with control apoA-I. The V156E mutant was able to interact with phospholipid (PL) at high PL:protein ratios (95:1, mol/mol), but was inefficient in forming reconstituted HDL (rHDL) complexes at lower PL:protein ratios (40:1). In the lipid-bound, rHDL state, the mutant protein was somewhat more alpha-helical and formed a larger complex (110 A) than control apoA-I (97 A). Furthermore, the rHDL particles containing the V156E mutant did not rearrange to smaller particles in the presence of low density lipoproteins, and had minimal reactivity with lecithin-cholesterol acyltransferase (LCAT), compared with rHDL particles made with control apoA-I. These results suggest a key role for Val-156, or the adjacent central region of apoA-I in the modulation of apoA-I conformation, stability, and self-association in solution, and in the formation of small HDL, the conformational adaptability of apoA-I leading to structural rearrangements of HDL, and the activation of LCAT. PMID- 10858437 TI - Collagen, convulxin, and thrombin stimulate aggregation-independent tyrosine phosphorylation of CD31 in platelets. Evidence for the involvement of Src family kinases. AB - Platelet endothelial cell adhesion molecule-1 (CD31) is a 130-kDa glycoprotein receptor present on the surface of platelets, neutrophils, monocytes, certain T lymphocytes, and vascular endothelial cells. CD31 is involved in adhesion and signal transduction and is implicated in the regulation of a number of cellular processes. These include transendothelial migration of leukocytes, integrin regulation, and T-cell function, although its function in platelets remains unclear. In this study, we demonstrate the ability of the platelet agonists collagen, convulxin, and thrombin to induce tyrosine phosphorylation of CD31. Furthermore, we show that this event is independent of platelet aggregation and secretion and is accompanied by an increase in surface expression of CD31. A kinase capable of phosphorylating CD31 was detected in CD31 immunoprecipitates, and its activity was increased following activation of platelets. CD31 tyrosine phosphorylation was reduced or abolished by the Src family kinase inhibitor PP2, suggesting a role for these enzymes. In accordance with this, each of the Src family members expressed in platelets, namely Fyn, Lyn, Src, Yes, and Hck, was shown to co-immunoprecipitate with CD31. The involvement of Src family kinases in this process was confirmed through the study of mouse platelets deficient in Fyn. PMID- 10858438 TI - Sterol-dependent transactivation of the ABC1 promoter by the liver X receptor/retinoid X receptor. AB - Tangier disease, a condition characterized by low levels of high density lipoprotein and cholesterol accumulation in macrophages, is caused by mutations in the ATP-binding cassette transporter ABC1. In cultured macrophages, ABC1 mRNA was induced in an additive fashion by 22(R)-hydroxycholesterol and 9-cis-retinoic acid (9CRA), suggesting induction by nuclear hormone receptors of the liver X receptor (LXR) and retinoid X receptor (RXR) family. We cloned the 5'-end of the human ABC1 transcript from cholesterol-loaded THP1 macrophages. When transfected into RAW macrophages, the upstream promoter was induced 7-fold by 22(R) hydroxycholesterol, 8-fold by 9CRA, and 37-fold by 9CRA and 22(R) hydroxycholesterol. Furthermore, promoter activity was increased in a sterol responsive fashion when cotransfected with LXRalpha/RXR or LXRbeta/RXR. Further experiments identified a direct repeat spaced by four nucleotides (from -70 to 55 base pairs) as a binding site for LXRalpha/RXR or LXRbeta/RXR. Mutations in this element abolished the sterol-mediated activation of the promoter. The results show sterol-dependent transactivation of the ABC1 promoter by LXR/RXR and suggest that small molecule agonists of LXR could be useful drugs to reverse foam cell formation and atherogenesis. PMID- 10858440 TI - Stimulation of leukemia inhibitory factor receptor degradation by extracellular signal-regulated kinase. AB - Leukemia inhibitory factor (LIF) signals via the heterodimeric receptor complex comprising the LIF receptor alpha subunit (LIFRalpha) and the common signal transducing subunit for interleukin-6 cytokine receptors, gp130. This study demonstrates that in different cell types, the level of LIFRalpha decreases during treatment with LIF or the closely related cytokine oncostatin M (OSM). Moreover, insulin and epidermal growth factor induce a similar LIFRalpha down regulation. The regulated loss of LIFRalpha is specific since neither gp130 nor OSM receptor beta shows a comparable change in turnover. LIFRalpha down regulation correlates with reduced cell responsiveness to LIF. Using protein kinase inhibitors and point mutations in LIFRalpha, we demonstrate that LIFRalpha down-regulation depends on activation of extracellular signal-regulated kinase 1/2 and phosphorylation of the cytoplasmic domain of LIFRalpha at serine 185. This modification appears to promote the endosomal/lysosomal pathway of the LIFRalpha. These results suggest that extracellular signal-regulated kinase activating factors like OSM and growth factors have the potential to lower specifically LIF responsiveness in vivo by regulating LIFRalpha half-life. PMID- 10858439 TI - Cytokine signaling through Stat3 activates integrins, promotes adhesion, and induces growth arrest in the myeloid cell line 32D. AB - Hematopoietic cell development and function is dependent on cytokines and on intercellular interactions with the microenvironment. Although the intracellular signaling pathways stimulated by cytokine receptors are well described, little is known about the mechanisms through which these pathways modulate hematopoietic cell adhesion events in the microenvironment. Here we show that cytokine activated Stat3 stimulates the expression and function of cell surface adhesion molecules in the myeloid progenitor cell line 32D. We generated an erythropoietin receptor (EpoR) isoform (ER343/401-S3) that activates Stat3 rather than Stat5 by substituting the Stat3 binding/activation sequence motif from gp130 for the sequences surrounding tyrosines 343 and 401 in the receptor cytoplasmic region. Activation of Stat3 leads to homotypic cell aggregation, increased expression of intercellular adhesion molecule 1 (ICAM-1), CD18, and CD11b, and activation of signaling through CD18-containing integrins. Unlike the wild type EpoR, ER343/401 S3 is unable to support long term Epo-dependent proliferation in 32D cells. Instead, Epo-treated ER343/401-S3 cells undergo G(1) arrest and express elevated levels of the cyclin-dependent kinase inhibitor p27(Kip1). Sustained activation of Stat3 in these cells is required for their altered morphology and growth properties since constitutive SOCS3 expression abrogates homotypic cell aggregation, signaling through CD18-containing integrins, G(1) arrest, and accumulation of p27(Kip1). Collectively, our results demonstrate that cytokine activated Stat3 stimulates the expression and function of cell surface adhesion molecules, indicating that a role for Stat3 is to regulate intercellular contacts in myeloid cells. PMID- 10858441 TI - Activase region on chloroplast ribulose-1,5-bisphosphate carboxylase/oxygenase. Nonconservative substitution in the large subunit alters species specificity of protein interaction. AB - In the active form of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC ), a carbamate at lysine 201 binds Mg(2+), which then interacts with the carboxylation transition state. Rubisco activase facilitates this spontaneous carbamylation/metal-binding process by removing phosphorylated inhibitors from the Rubisco active site. Activase from Solanaceae plants (e.g. tobacco) fails to activate Rubisco from non-Solanaceae plants (e.g. spinach and Chlamydomonas reinhardtii), and non-Solanaceae activase fails to activate Solanaceae Rubisco. Directed mutagenesis and chloroplast transformation previously showed that a proline 89 to arginine substitution on the surface of the large subunit of Chlamydomonas Rubisco switched its specificity from non-Solanaceae to Solanaceae activase activation. To define the size and function of this putative activase binding region, substitutions were created at positions flanking residue 89. As in the past, these substitutions changed the identities of Chlamydomonas residues to those of tobacco. Whereas an aspartate 86 to arginine substitution had little effect, aspartate 94 to lysine Rubisco was only partially activated by spinach activase but now fully activated by tobacco activase. In an attempt to eliminate the activase/Rubisco interaction, proline 89 was changed to alanine, which is not present in either non-Solanaceae or Solanaceae Rubisco. This substitution also caused reversal of activase specificity, indicating that amino acid identity alone does not determine the specificity of the interaction. PMID- 10858442 TI - Molecular characterization of a novel beta-glucuronidase from Scutellaria baicalensis georgi. AB - We cloned a gene encoding Scutellaria beta-glucuronidase (sGUS) that is involved in the initiation of H(2)O(2) metabolism in skullcap (Scutellaria baicalensis). This gene consists of a 1581-nucleotide open reading frame, the deduced amino acid sequence of which contains an ATP/GTP binding site and a leucine zipper motif. sGUS has apparent similarity to the heparan sulfate-metabolizing beta glucuronidase heparanase but no homology to family 2 beta-glucuronidases. In addition, neither the family 2 glycosylhydrolase signature nor family 2 acid-base catalyst was found in this enzyme. These results suggested that sGUS does not belong to the family 2 beta-glucuronidases. We modified several residues predicted to act as the acid-base or nucleophilic residue of sGUS by site directed mutagenesis. Mutations at Glu(212) or Glu(329) resulted in much lower k(cat)/K(m) values in the mutants as compared with the wild-type enzyme, indicating that these are the acid-base and nucleophilic residues of the active site, respectively. Moreover, similar site-directed mutagenesis confirmed that Tyr(281) is also involved in the beta-glucuronidase activity. The amino acid sequences of small regions containing these active site residues were conserved in heparanases. As sGUS has various structural characteristics in common with heparanase, we concluded that sGUS and heparanase belong to the same new family. PMID- 10858443 TI - Transcription elongation factor S-II confers yeast resistance to 6-azauracil by enhancing expression of the SSM1 gene. AB - Loss of function of S-II makes yeast sensitive to 6-azauracil. Here, we identified a multi-copy suppressor gene of this phenotype, termed SSM1 (suppressor of 6-azauracil sensitivity of the S-II null mutant 1), that encodes a novel protein consisting of 280 amino acid residues. Although both the SSM1 null mutant and the S-II/SSM1 double null mutant were viable under normal growth conditions, they resembled the S-II null mutant in being sensitive to 6 azauracil. Expression of the SSM1 gene was found to be repressed in the S-II null mutant but was restored by overexpression of chimeric S-II molecules that were able to stimulate transcription elongation by RNA polymerase II in vitro. Furthermore, we identified two transcription arrest sites within the transcription unit of the SSM1 gene in vitro that could be relieved by S-II. These results indicate that S-II confers yeast resistance to 6-azauracil by stimulating transcription elongation of the SSM1 gene. PMID- 10858444 TI - A thermodynamic coupling mechanism for the disaggregation of a model peptide substrate by chaperone secB. AB - Molecular chaperones prevent protein aggregation in vivo and in vitro. In a few cases, multichaperone systems are capable of dissociating aggregated state(s) of substrate proteins, although little is known of the mechanism of the process. SecB is a cytosolic chaperone, which forms part of the precursor protein translocation machinery in Escherichia coli. We have investigated the interaction of the B-chain of insulin with chaperone SecB by light scattering, pyrene excimer fluorescence, and electron spin resonance spectroscopy. We show that SecB prevents aggregation of the B-chain of insulin. We show that SecB is capable of dissociating soluble B-chain aggregates as monitored by pyrene fluorescence spectroscopy. The kinetics of dissociation of the B-chain aggregate by SecB has been investigated to understand the mechanism of dissociation. The data suggests that SecB does not act as a catalyst in dissociation of the aggregate to individual B-chains, rather it binds the small population of free B-chains with high affinity, thereby shifting the equilibrium from the ensemble of the aggregate toward the individual B-chains. Thus SecB can rescue aggregated, partially folded/misfolded states of target proteins by a thermodynamic coupling mechanism when the free energy of binding to SecB is greater than the stability of the aggregate. Pyrene excimer fluorescence and ESR methods have been used to gain insights on the bound state conformation of the B-chain to chaperone SecB. The data suggests that the B-chain is bound to SecB in a flexible extended state in a hydrophobic cleft on SecB and that the binding site accommodates approximately 10 residues of substrate. PMID- 10858445 TI - Interaction of nucleolin with an evolutionarily conserved pre-ribosomal RNA sequence is required for the assembly of the primary processing complex. AB - The first processing event of the precursor ribosomal RNA (pre-rRNA) takes place within the 5' external transcribed spacer. This primary processing requires conserved cis-acting RNA sequence downstream from the cleavage site and several nucleic acids (small nucleolar RNAs) and proteins trans-acting factors including nucleolin, a major nucleolar protein. The specific interaction of nucleolin with the pre-rRNA is required for processing in vitro. Xenopus laevis and hamster nucleolin interact with the same pre-rRNA site and stimulate the processing activity of a mouse cell extract. A highly conserved 11-nucleotide sequence located 5-6 nucleotides after the processing site is required for the interaction of nucleolin and processing. In vitro selection experiments with nucleolin have identified an RNA sequence that contains the UCGA motif present in the 11 nucleotide conserved sequence. The interaction of nucleolin with pre-rRNA is required for the formation of an active processing complex. Our findings demonstrate that nucleolin is a key factor for the assembly and maturation of pre ribosomal ribonucleoparticles. PMID- 10858446 TI - Structural characterization of the O-antigenic polysaccharide of the lipopolysaccharide from Rhizobium etli strain CE3. A unique O-acetylated glycan of discrete size, containing 3-O-methyl-6-deoxy-L-talose and 2,3,4-tri-O-,methyl l fucose. AB - The O-antigenic polysaccharide of the Rhizobium etli CE3 lipopolysaccharide (LPS) was structurally characterized using chemical degradations (Smith degradation and beta-elimination of uronosyl residues) in combination with alkylation analysis, electrospray, and matrix-assisted laser desorption ionization-time of flight mass spectrometry, tandem mass spectrometry, and (1)H COSY and TOCSY nuclear magnetic resonance spectroscopy analyses of the native polysaccharide and the derived oligosaccharides. The polysaccharide was found to be a unique, relatively low molecular weight glycan having a fairly discrete size, with surprisingly little variation in the number of repeating units (degree of polymerization = 5). The polysaccharide is O-acetylated and contains a variety of O-methylated glycosyl residues, rendering the native glycan somewhat hydrophobic. The molecular mass of the major de-O-acetylated species, including the reducing end 3-deoxy-d-manno-2 octulosonic acid (Kdo) residue, is 3330 Da. The polysaccharide is comprised of a trisaccharide repeating unit having the structure -->4)-alpha-d-GlcpA-(1-->4) [alpha-3-O-Me-6-deoxy-Talp-(1--> 3)]-alpha -l-Fucp-(1-->. The nonreducing end of the glycan is terminated with the capping sequence alpha-2,3, 4-tri-O-Me-Fucp-(1- >4)-alpha-d-GlcpA-(1-->, and the reducing end of the molecule consists of the non repeating sequence -->3)-alpha-l-Fucp-(1-->3)-beta-d-Manp-(1-->3)-beta-QuiNA cp (1-->4)-a lpha-Kdop-(2-->, where QuiNAc is N-acetylquinovosamine (2-N-acetamido 2,6-dideoxyglucose). The reducing end Kdo residue links the O-chain polysaccharide to the core region oligosaccharide, resulting in a unique location for a Kdo residue in LPS, removed four residues distally from the lipid A moiety. Structural heterogeneity in the O-chain arises mainly from the O-acetyl and O methyl substitution. Methylation analysis using trideuteriomethyl iodide indicates that a portion of the 2,3,4-tri-O-methylfucosyl capping residues, typically 15%, are replaced with 2-O-methyl- and/or 2,3-di-O-methylfucosyl residues. In addition, approximately 25% of the 3,4-linked branching fucosyl residues and 10% of the 3-linked fucosyl residues are 2-O-methylated. A majority of the glucuronosyl residues are methyl-esterified at C-6. These unique structural features may be significant in the infection process. PMID- 10858447 TI - Binding and cross-linking studies show that scavenger receptor BI interacts with multiple sites in apolipoprotein A-I and identify the class A amphipathic alpha helix as a recognition motif. AB - Scavenger receptor, class B, type I (SR-BI) mediates the selective uptake of high density lipoprotein (HDL) cholesteryl ester without the uptake and degradation of the particle. In transfected cells SR-BI recognizes HDL, low density lipoprotein (LDL) and modified LDL, protein-free lipid vesicles containing anionic phospholipids, and recombinant lipoproteins containing apolipoprotein (apo) A-I, apoA-II, apoE, or apoCIII. The molecular basis for the recognition of such diverse ligands by SR-BI is unknown. We have used direct binding analysis and chemical cross-linking to examine the interaction of murine (m) SR-BI with apoA I, the major protein of HDL. The results show that apoA-I in apoA-I/palmitoyl oleoylphosphatidylcholine discs, HDL(3), or in a lipid-free state binds to mSR-BI with high affinity (K(d) congruent with 5-8 microgram/ml). ApoA-I in each of these forms was efficiently cross-linked to cell surface mSR-BI, indicating that direct protein-protein contacts are the predominant feature that drives the interaction between HDL and mSR-BI. When complexed with dimyristoylphosphatidylcholine, the N-terminal and C-terminal CNBr fragments of apoA-I each bound to SR-BI in a saturable, high affinity manner, and each cross linked efficiently to mSR-BI. Thus, mSR-BI recognizes multiple sites in apoA-I. A model class A amphipathic alpha-helix, 37pA, also showed high affinity binding and cross-linking to mSR-BI. These studies identify the amphipathic alpha-helix as a recognition motif for SR-BI and lead to the hypothesis that mSR-BI interacts with HDL via the amphipathic alpha-helical repeat units of apoA-I. This hypothesis explains the interaction of SR-BI with a wide variety of apolipoproteins via a specific secondary structure, the class A amphipathic alpha helix, that is a common structural motif in the apolipoproteins of HDL, as well as LDL. PMID- 10858448 TI - Casein kinase I is anchored on axonemal doublet microtubules and regulates flagellar dynein phosphorylation and activity. AB - Flagellar dynein activity is regulated by phosphorylation. One critical phosphoprotein substrate in Chlamydomonas is the 138-kDa intermediate chain (IC138) of the inner arm dyneins (Habermacher, G., and Sale, W. S. (1997) J. Cell Biol. 136, 167-176). In this study, several approaches were used to determine that casein kinase I (CKI) is physically anchored in the flagellar axoneme and regulates IC138 phosphorylation and dynein activity. First, using a videomicroscopic motility assay, selective CKI inhibitors rescued dynein-driven microtubule sliding in axonemes isolated from paralyzed flagellar mutants lacking radial spokes. Rescue of dynein activity failed in axonemes isolated from these mutant cells lacking IC138. Second, CKI was unequivocally identified in salt extracts from isolated axonemes, whereas casein kinase II was excluded from the flagellar compartment. Third, Western blots indicate that within flagella, CKI is anchored exclusively to the axoneme. Analysis of multiple Chlamydomonas motility mutants suggests that the axonemal CKI is located on the outer doublet microtubules. Finally, CKI inhibitors that rescued dynein activity blocked phosphorylation of IC138. We propose that CKI is anchored on the outer doublet microtubules in position to regulate flagellar dynein. PMID- 10858449 TI - Cloning and characterization of the gene for phosphatidylcholine synthase. AB - Phosphatidylcholine (PC) is the major membrane-forming phospholipid in eukaryotes and can be synthesized by either of two pathways, the CDP-choline pathway or the methylation pathway. In prokaryotes only the methylation pathway was thought to occur. Recently, however, we could demonstrate (de Rudder, K. E. E., Sohlenkamp, C., and Geiger, O. (1999) J. Biol. Chem. 274, 20011-20016) that a second pathway for phosphatidylcholine biosynthesis exists in Sinorhizobium (Rhizobium) meliloti involving a novel enzymatic activity, phosphatidylcholine synthase, that condenses choline and CDP-diacylglyceride in one step to form PC and CMP. Using a colony autoradiography method we have isolated mutants of S. meliloti deficient in phosphatidylcholine synthase and which are no longer able to incorporate radiolabeled choline into PC. Complementation of such mutants with a sinorhizobial cosmid gene bank, subcloning of the complementing fragment, and sequencing of the subclone led to the identification of a gene coding for a presumptive CDP-alcohol phosphatidyltransferase. Amplification of this gene and its expression in Escherichia coli demonstrates that it codes for phosphatidylcholine synthase. Genomes of some pathogens (Pseudomonas aeruginosa and Borrelia burgdorferi) contain genes similar to the sinorhizobial gene (pcs) for phosphatidylcholine synthase. Although pcs-deficient S. meliloti knock-out mutants show wild type-like growth and lipid composition, they are unable to perform rapid PC biosynthesis that normally is achieved via the phosphatidylcholine synthase pathway in S. meliloti wild type. PMID- 10858450 TI - Free energy requirement for domain movement of an enzyme. AB - Domain movement is sometimes essential for substrate recognition by an enzyme. X ray crystallography of aminotransferase with a series of aliphatic substrates showed that the domain movement of aspartate aminotransferase was changed dramatically from an open to a closed form by the addition of only one CH(2) to the side chain of the C4 substrate CH(3)(CH(2))C((alpha))H(NH(3)(+))COO(-). These crystallographic results and reaction kinetics (Kawaguchi, S., Nobe, Y., Yasuoka, J., Wakamiya, T., Kusumoto, S., and Kuramitsu, S. (1997) J. Biochem. (Tokyo) 122, 55-63; Kawaguchi, S. and Kuramitsu, S. (1998) J. Biol. Chem. 273, 18353-18364) enabled us to estimate the free energy required for the domain movement. PMID- 10858451 TI - Studies on the specificity of the tetrapyrrole substrate for human biliverdin IXalpha reductase and biliverdin-IXbeta reductase. Structure-activity relationships define models for both active sites. AB - A comparison of the initial rate kinetics for human biliverdin-IXalpha reductase and biliverdin-IXbeta reductase with a series of synthetic biliverdins with propionate side chains "moving" from a bridging position across the central methene bridge (alpha isomers) to a "gamma-configuration" reveals characteristic behavior that allows us to propose distinct models for the two active sites. For human biliverdin-IXalpha reductase, as previously discussed for the rat and ox enzymes, it appears that at least one "bridging propionate" is necessary for optimal binding and catalytic activity, whereas two are preferred. All other configurations studied were substrates for human biliverdin-IXalpha reductase, albeit poor ones. In the case of mesobiliverdin-XIIIalpha, extending the propionate side chains to hexanoate resulted in a significant loss of activity, whereas the butyrate derivative retained high activity. For human biliverdin IXalpha reductase, we suggest that a pair of positively charged side chains play a key role in optimally binding the IXalpha isomers. In the case of human biliverdin-IXbeta reductase, the enzyme cannot tolerate even one propionate in the bridging position, suggesting that two negatively charged residues on the enzyme surface may preclude productive binding in this case. The flavin reductase activity of biliverdin-IXbeta reductase is potently inhibited by mesobiliverdin XIIIalpha and protohemin, which is consistent with the hypothesis that the tetrapyrrole and flavin substrate bind at a common site. PMID- 10858452 TI - First apyrase splice variants have different enzymatic properties. AB - LALP70 is a novel lysosomal membrane protein belonging to the apyrase protein family. The apyrase protein family comprises enzymes capable of cleaving nucleotide tri- and diphosphates in a calcium- or magnesium-dependent manner, not being altered by P-type, F-type, or V-type NTPase inhibitors. In this study we have cloned and sequenced the human LALP70 gene to determine the genomic structure. The gene is organized in 11 introns and 12 exons covering a genomic region of approximately 16 kilobase pairs. By fluorescence in situ hybridization analysis, the hLALP70 gene was mapped to the human chromosome 8p21.1-p21.3. We further show that there is at least one alternatively spliced variant, hLALP70v, which can be generated via an alternative splice side at the 3'-end of exon 7, leading to a protein variant differing in 8 amino acids (VSFASSQQ). This is the first splice variant that has been described in the apyrase protein family. Reverse transcriptase polymerase chain reaction analysis showed an ubiquitous expression of both variants, with different relative mRNA expression levels in different tissues. Comparison of the enzymatic properties of the splice variants revealed a broader substrate specificity for hLALP70v with CTP, UDP, CDP, GTP, and GDP as preferred substrates, while hLALP70 utilized UTP and TTP preferentially. Furthermore, enzyme activity of hLALP70v was equally dependent on Ca(2+) and Mg(2+), being saturated already at 1 mm concentration. In contrast, hLALP70 enzymatic activity were unsaturated up to 10 mm Ca(2+), while Mg(2+) showed a saturation at already 1 mm concentration with 2-3-fold lower enzymatic activity as observed with Ca(2+). Our data suggest that the presence or absence of the 8-amino acid motif VSFASSQQ provoke differences in substrate specificity and divalent cation dependence of hLALP70/hLALP70v. PMID- 10858453 TI - Gravin-mediated formation of signaling complexes in beta 2-adrenergic receptor desensitization and resensitization. AB - Agonist-induced desensitization and resensitization of G-protein-linked receptors involve the interaction of receptors with protein kinases, phosphatases, beta arrestin, and clathrin organized by at least one scaffold protein. The dynamic composition of the signaling complexes and the role of the scaffold protein AKAP250 (gravin) in agonist-induced attenuation and recovery of beta-adrenergic receptors were explored by co-immunoprecipitation of target elements, antisense suppression, and confocal microscopy. Gravin associated with unstimulated receptor, and the association was increased significantly after agonist stimulation for up to 60 min. Agonist stimulation also induced a robust association of the receptor-gravin complex with protein kinases A and C, G protein-linked receptor kinase-2, beta-arrestin, and clathrin. Confocal microscopy of the green fluorescence protein-tagged beta(2)-adrenergic receptor showed that the receptor underwent sequestration after agonist stimulation. Suppression of gravin expression via antisense oligodeoxynucleotides disrupted agonist-induced association of the receptor with G-protein-linked receptor kinase 2, beta-arrestin, and clathrin as well as receptor recovery from desensitization. Gravin deficiency also inhibited agonist-induced sequestration. These data reveal that gravin-mediated formation of signaling complexes with protein kinases/phosphatases, beta-arrestin, and clathrin is essential in agonist-induced internalization and resensitization of G-protein-linked receptors. PMID- 10858454 TI - Effect of protein kinase A activity on the association of ADP-ribosylation factor 1 to golgi membranes. AB - The small GTP-binding protein ADP-ribosylation factor 1 (ARF1) is an essential component of the molecular machinery that catalyzes the formation of membrane bound transport intermediates. By using an in vitro assay that reproduces recruitment of cytosolic proteins onto purified, high salt-washed Golgi membranes, we have analyzed the role of cAMP-dependent protein kinase A (PKA) on ARF1 incorporation. Addition to this assay of either pure catalytic subunits of PKA (C-PKA) or cAMP increased ARF1 binding. By contrast, ARF1 association was inhibited following C-PKA inactivation with either PKA inhibitory peptide or RIIalpha as well as after cytosol depletion of C-PKA. C-PKA also stimulated recruitment and activation of a recombinant form of human ARF1 in the absence of additional cytosolic components. The binding step could be dissociated from the activation reaction and found to be independent of guanine nucleotides and saturable. This step was stimulated by C-PKA in an ATP-dependent manner. Dephosphorylated Golgi membranes exhibited a decreased ability to recruit ARF1, and this effect was reverted by addition of C-PKA. Following an increase in the intracellular level of cAMP, ARF proteins redistributed from cytosol to the perinuclear Golgi region of intact cells. Collectively, the results show that PKA exerts a key regulatory role in the recruitment of ARF1 onto Golgi membranes. In contrast, PKA modulators did not affect recruitment of beta-COP onto Golgi membranes containing prebound ARF1. PMID- 10858455 TI - Generation of structurally and functionally distinct factors from the basic helix loop-helix gene Hes3 by alternative first exons. AB - The basic helix-loop-helix gene Hes3 is expressed by differentiating and mature Purkinje cells in the cerebellum and by neural precursor cells in the embryonal nervous system. We here found that the transcript of cerebellar Hes3, designated Hes3a, has a distinct 5'-terminal structure from that of embryonal Hes3, designated Hes3b, and that the two types of Hes3 transcripts are generated from different first exons. Hes3a lacks the amino-terminal half of the basic region and thus does not bind to the DNA, whereas Hes3b contains a complete basic region and binds to the N box sequence with a high affinity like Hes1, another member of the Hes family. Both types of Hes3 proteins functionally antagonize the neuronal determination factor Mash1, but only Hes3b represses transcription from the N box containing promoter like Hes1. Furthermore, misexpression of Hes3b with retrovirus in neural precursor cells inhibits neuronal differentiation like Hes1, whereas Hes3a does not. Thus, alternative promoters and first exons that are differentially utilized during neural development generate structurally and functionally distinct proteins from a single Hes3 gene locus. PMID- 10858456 TI - Copper(II)-induced conformational changes and protease resistance in recombinant and cellular PrP. Effect of protein age and deamidation. AB - While PrP(C) rearranges in the area of codons 104-113 to form PrP(Sc) during prion infections, the events that initiate sporadic Creutzfeldt-Jakob disease are undefined. As Cu(II) is a putative ligand for PrP(C) and has been implicated in the pathogenesis of Creutzfeldt-Jakob disease and other neurodegenerative diseases, we investigated the structural effects of binding. Incubation of brain microsomes with Cu(II) generated approximately 30-kDa proteinase K-resistant PrP. Cu(II) had little effect on fresh recombinant PrP23-231, but aged protein characterized by conversion of Asn-107 to Asp decreased alpha-helical content by approximately 30%, increased beta-sheet content 100%, formed aggregates, and acquired proteinase K resistance in the presence of Cu(II). These transitions took place without need for acid pH, organic solvents, denaturants, or reducing agents. Since conversion of Asn to Asp proceeds by a spontaneous pathway involving deamidation, our data suggest that covalent variants of PrP(C) arising in this manner may, in concert with Cu(II), generate PrP(Sc)-like species capable of initiating sporadic prion disease. PMID- 10858457 TI - The mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episode syndrome-associated human mitochondrial tRNALeu(UUR) mutation causes aminoacylation deficiency and concomitant reduced association of mRNA with ribosomes. AB - The pathogenetic mechanism of the mitochondrial tRNA(Leu(UUR)) A3243G transition associated with the mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome has been investigated in transmitochondrial cell lines constructed by transfer of mutant mitochondrial DNA (mtDNA)-carrying mitochondria from three genetically unrelated MELAS patients or of isogenic wild type mtDNA-carrying organelles into human mtDNA-less cells. An in vivo footprinting analysis of the mtDNA segment within the tRNA(Leu(UUR)) gene that binds the transcription termination factor failed to reveal any difference in occupancy of sites or qualitative interaction with the protein between mutant and wild-type mtDNAs. Cell lines nearly homoplasmic for the mutation exhibited a strong (70-75%) reduction in the level of aminoacylated tRNA(Leu(UUR)) and a decrease in mitochondrial protein synthesis rate. The latter, however, did not show any significant correlation between synthesis defect of the individual polypeptides and number or proportion of UUR codons in their mRNAs, suggesting that another step, other than elongation, may be affected. Sedimentation analysis in sucrose gradient showed a reduction in size of the mitochondrial polysomes, while the distribution of the two rRNA components and of the mRNAs revealed decreased association of mRNA with ribosomes and, in the most affected cell line, pronounced degradation of the mRNA associated with slowly sedimenting structures. Therefore, several lines of evidence indicate that the protein synthesis defect in A3243G MELAS mutation-carrying cells is mainly due to a reduced association of mRNA with ribosomes, possibly as a consequence of the tRNA(Leu(UUR)) aminoacylation defect. PMID- 10858458 TI - The glioma-associated protein SETA interacts with AIP1/Alix and ALG-2 and modulates apoptosis in astrocytes. AB - Expression of the src homology 3 (SH3) domain-containing expressed in tumorigenic astrocytes (SETA) gene is associated with the tumorigenic state in astrocytes. SETA encodes a variety of adapter proteins containing either one or two SH3 domains, as suggested by the sequence heterogeneity of isolated cDNAs. Using both SH3 domains in a yeast two-hybrid screen of a glial progenitor cell cDNA library, we isolated the rat homolog of the ALG-2-interacting protein 1 or ALG-2 interacting protein X (AIP1/Alix). In vitro confrontation experiments showed that the SH3-N domain of SETA interacted with the proline-rich C terminus of AIP1. In co-immunoprecipitation experiments, SETA and AIP1 interacted and could form a complex with apoptosis-linked gene 2 protein. Endogenous SETA and AIP1 proteins showed similar patterns of staining in primary rat astrocytes. Misexpression of a variety of SETA protein isoforms in these astrocytes revealed that they localized to the actin cytoskeleton. Furthermore, SETA proteins containing the SH3-N domain were able to sensitize astrocytes to apoptosis induced by UV irradiation. Expression of the isolated SH3-N domain had the greatest effect in these experiments, indicating that interference in the interaction between endogenous SETA and AIP1 sensitizes astrocytes to apoptosis in response to DNA damage. PMID- 10858459 TI - Abrogation of nerve growth factor-induced terminal differentiation by ret oncogene involves perturbation of nuclear translocation of ERK. AB - Oncogenic variants of the receptor tyrosine kinase, Ret, cause formation of tumors of neuroendocrine derivation in the multiple endocrine neoplasia type 2 and, thus, likely interfere with antiproliferative and/or differentiative extracellular signals. Here we took advantage of two rat pheochromocytoma-derived cell lines (PC12/MEN2A and PC12/MEN2B) to investigate whether Ret-induced nerve growth factor (NGF) unresponsiveness might involve impairment of ERK signaling. In fact, these cells, stably transfected with distinct forms of the active ret oncogene, fail to block proliferation, even upon NGF stimulation. In these cells we show the presence of both chronic ERKs activity and high expression levels of MKP-3, an ERK-specific phosphatase. Despite the presence of MKP-3, ERK activity can be further stimulated by NGF, but it fails to translocate into the nucleus and consequently to induce immediate-early gene transcription. Because of the presence of MKP-3, our results suggest the existence of a negative regulatory feedback acting on ERKs as a mechanism responsible for the abrogation of NGF induced terminal differentiation. Indeed, MKP-3 seems to be implicated in the persistence of ERKs in cell cytoplasm. This interpretation is further supported by the observation that in ret-transfected cells, forced expression of an active form of MEK-1 may overcome this block; it restores transcription from the c-fos promoter, induces translocation of ERKs into the nucleus, and inhibits cell proliferation. PMID- 10858460 TI - Alternative translation initiation of human fibroblast growth factor 2 mRNA controlled by its 3'-untranslated region involves a Poly(A) switch and a translational enhancer. AB - Five fibroblast growth factor 2 (FGF-2) isoforms are synthesized from human FGF-2 mRNA by a process of alternative initiation of translation. The regulation of FGF 2 isoform expression by the mRNA 5823-nucleotide-long 3'-untranslated region containing eight alternative polyadenylation sites was examined. Because previous studies had shown that FGF-2 expression was regulated in primary cells but not in transformed cells, primary human skin fibroblasts were used in this study. Using an approach of cell transfection with synthetic reporter mRNAs, a novel translational enhancer (3'-TE) was identified in the 1370-nucleotide mRNA segment located upstream from the eighth poly(A) site. Deletion mutagenesis showed that the 3'-TE was composed of two domains with additive effects. The 3'-TE exhibited the unique feature of modulating the use of FGF-2 alternative initiation codons, which favored the relative expression of CUG-initiated isoforms. Interestingly, the use of an alternative polydenylation site removing the 3'-TE was detected in skin fibroblasts in response to heat shock and cell density variations. At high cell densities, 3'-TE removal was correlated with a loss of CUG-initiated FGF-2 expression. These data show that the FGF-2 mRNA 3'-untranslated region is able to modulate FGF-2 isoform expression by the coupled processes of translation activation and alternative polyadenylation. PMID- 10858461 TI - Molecular cloning of endo-beta -galactosidase C and its application in removing alpha -galactosyl xenoantigen from blood vessels in the pig kidney. AB - Galalpha1-3Gal is the major xenoantigenic epitope responsible for hyperacute rejection upon pig to human xenotransplantation. Endo-beta-galactosidase C from Clostridium perfringens destroys the antigenic epitope by cleaving the beta galactosidic linkage in the Galalpha1-3Galbeta1-4GlcNAc structure. Based on partial peptide sequences of the enzyme, we molecularly cloned the enzyme gene, which encodes a protein with a predicted molecular mass of about 93 kDa. The deduced protein sequence of the enzyme has limited homology in the C-terminal half with endo-beta-galactosidase from Flavobacterium keratolyticus and beta-1,3 glucanases. The enzyme expressed in Escherichia coli removed the alpha-galactosyl epitope nearly completely from pig erythrocytes and from pig aortic endothelial cells. The enzyme-treated endothelial cells in culture were greatly reduced in cell surface antigens, which were recognized by IgM, IgG, or IgA in human sera, and became much less susceptible to complement-mediated cytotoxicity caused by human sera. When the pig kidney was perfused with the enzyme, the vascular endothelial cells became virtually devoid of the alpha-galactosyl epitope, with concomitant decrease in binding to IgM in human plasma. These results demonstrated that the recombinant endo-beta-galactosidase C is a valuable aid in xenotransplantation. PMID- 10858462 TI - Innovative technologies. The Information Age and the BioIntelligence Age. PMID- 10858463 TI - Laparoscopic ventral hernia repair: a report of 100 consecutive cases. AB - INTRODUCTION: Effective surgical therapy for ventral and incisional hernias is problematic. Recurrence rates following primary repair range as high as 25-49%, and breakdown following conventional treatment of recurrent hernias can exceed 50%. As an alternative, laparoscopic techniques offer the potential benefits of decreased pain and a shorter hospital stay. This study evaluates the efficacy of the laparoscopic approach for ventral herniorrhaphy. METHODS: A retrospective review was performed for 100 consecutive patients with ventral hernias who underwent laparoscopic repair at our institutions between November 1995 and May 1998. All patients who presented during this period and were candidates for a mesh hernia repair were treated via an endoscopic approach. RESULTS: One hundred patients underwent a laparoscopic ventral hernia repair. There were 48 men and 52 women. The patients were typically obese, with a mean body mass index (BMI) of 31 kg/m(2). Each had undergone an average of 2.5 (range; 0-8) previous laparotomies. Forty-nine repairs were performed for recurrent hernias. An average of two patients (range; 1-7) had previously failed open herniorhaphies; in 20 cases, intraabdominal polypropylene mesh was present. There were no conversions to open operation. The mean size of the defects was large at 87 cm(2) (range; 1-480). In all cases, the mesh (average, 287 cm(2)) was secured with transabdominal sutures and metal tacks or staples. Operative time and estimated blood loss averaged 88 min (range; 18-270) and 30 cc (range; 10-150). Length of stay averaged 1. 6 days (range; 0-4). There were 12 minor and (two) major complications: cellulitis of the trocar site (two), seroma lasting >4 weeks (three), postoperative ileus (two), suture site pain > 2 weeks (two), urinary retention (one), respiratory distress (one), serosal bowel injury (one), and skin breakdown (one) and bowel injury (one). Both of the latter complications required mesh removal. With an average follow-up of 22.5 months (range; 7-37), there have been (three) recurrences. CONCLUSION: The laparoscopic approach to the repair of both primary and recurrent ventral hernias offers a low conversion rate, a short hospital stay, and few complications. At 23 months of follow-up, the recurrence rate has been 3%. Laparoscopic repair should be considered a viable option for any ventral hernia. PMID- 10858464 TI - Operative techniques and strategies for minimally invasive fetoscopic fetal cardiac interventions in sheep. AB - BACKGROUND: Recent efforts to develop procedures for fetoscopic fetal cardiac interventions have been prompted by the development of severe secondary damage to the fetal heart due to semilunar valvar obstructions and the poor outcome of therapy-refractory fetal arrhythmias. The purpose of our manuscript is to analyze and share our experience with the creation of an operative setup for these procedures in sheep. METHODS: We studied a total of 48 fetal sheep between 81 and 106 days of gestation (term, 145 days). After entering the amniotic cavity by a percutaneous approach, we performed various fetoscopic fetal cardiac procedures. We analyzed the success of percutaneous fetal access, methods of trocar support, the incidence and management of trocar dislodgement or accidental insertion into the chorioamniotic space, problems related to amniotic insufflation and trocar placement, as well as techniques for fetal posturing and uterine closure. RESULTS: Percutaneous fetal access was achieved in all sheep. The use of resterilizable trocars substantially decreased the costs of our procedures. Utilizing a percutaneous transuterine purse-string suture for trocar support helped to minimize the number of nonabsorbable T-fasteners remaining inside the uterus postoperatively. As complications such as trocar dislodgement, insertion of the trocar into the chorioamniotic space, and problems with intraamniotic insufflation and gas loss were mastered, conversion to an open operative approach was never required. A novel strategy that we devised for percutaneous fetal posturing permitted adequate fetal posturing with ease and minimal trauma to the fetal skin. CONCLUSION: As operative techniques have become more refined, the feasibility of performing fetoscopic fetal cardiac interventions in human fetuses now depends mainly on technical improvements in imaging and interventional catheters, as well as advances in pacemaker equipment. PMID- 10858465 TI - Quantitative evaluation of surgical task performance by remote-access endoscopic telemanipulation. AB - BACKGROUND: The performance limitations inherent in minimally invasive surgery may be overcome by using an interface that provides intuitive orientation for video display and tool manipulation. A prototype remote-access endoscopic telemanipulator was designed to fulfill these requirements and used for a surgical anastomosis task. METHODS: A remote-access telemanipulator system, employing remote center-of-motion geometry, was used to complete distant in vitro tubular anastomoses. The performance of four surgeons using this system was compared with that achieved in completing the same anastomosis task in an open environment using open surgical techniques and in a minimally invasive environment using standard laparoscopic methods. RESULTS: The average performance times for completion of the anastomosis task was 1448 +/- 130 s using the telemanipulator system compared with 2108 +/- 291 s with laparoscopic instruments and 296 +/- 25 s with conventional techniques. Leakage rates from the tubular anastomoses were 5.2 +/- 1.4 ml/s in the telemanipulator group, 6.9 +/- 2.0 ml/s in the laparoscopic group, and 3.2 +/- 0.9 ml/s in the conventional methods group. All experimental subjects were able to complete the assigned task in each experimental condition successfully without complications. CONCLUSIONS: Our results in this pilot study suggest that remote-access endoscopic telemanipulation can execute complex three-dimensional manipulations, and that the intuitive orientation of the surgeon's workstation may contribute to easier task completion. PMID- 10858466 TI - Recent experience with percutaneous endoscopic gastrostomy/jejunostomy (PEG/J) for enteral nutrition. AB - BACKGROUND: Enteral feeding is the preferred means of nutritional support in patients unable to eat orally. Jejunal-placed feeding tubes are often considered optimal for this purpose. Successful administration of such tube feedings depends on the method of placement and the size of the tube. Herein we review our experience with endoscopically placed jejunal feeding tubes. METHODS: Thirteen percutaneous endoscopic gastrostomy/jejunostomy (PEG/J) tubes were placed in 13 patients at the Emory University hospital by one surgeon. Indications for jejunal placement included aspiration in five patients and suspicion of increased reflux susceptibility in eight patients. Insertion of an 8.5-Fr nasobiliary tube was attempted in nine patients using the technique described by Coates and MacFadyen. A 12-Fr tube was placed in four patients using a technique that took advantage of previously placed PEG tubes. RESULTS: Initial placement was successful in all but one patient. Nine tube-related complications occurred in seven patients. These included six tube occlusions, one tube site infection, one peristomal leak, and one tube perforation that required replacement. Five of six tube occlusions (83%) occurred in the smaller 8.5-Fr. tubes. There was one non-tube-related death. CONCLUSIONS: PEG/J insertion can be performed successfully and safely in most patients. Long-term tube patency is, however, dependent on the use of tubes with a large diameter; thus, modalities that enable placement of larger-sized tubes are preferable. Further technical developments are needed to facilitate the endoscopic insertion of larger jejunostomy tubes. PMID- 10858467 TI - The effect of immune enhancement and suppression on the development of laparoscopic port site metastases. AB - BACKGROUND: Recent clinical case reports and experimental studies have suggested that laparoscopic cancer surgery is associated with an increased risk of tumor spread to abdominal wall wounds. While the etiology of this problem was initially believed to be related to mechanical contamination of wounds, it is now recognized that there are other contributory factors, including disturbed immune function within the peritoneal cavity. To investigate this question further, we evaluated the effect of immune modulation within an established laparoscopic cancer model. METHODS: Eighteen immune-competent syngeneic rats underwent modulation of their immune system, followed 18 h later by laparoscopy with the introduction of a suspension of adenocarcinoma cells into the peritoneal cavity. Rats were randomly allocated to receive either systemic cyclosporin (immune suppressor), intraperitoneal endotoxin (immune enhancer), or no agent (controls). Seven days later, all rats were killed and their peritoneal cavity was inspected for tumor implantation and port site metastases. RESULTS: Cyclosporin did not influence the study outcome, but tumor growth (p = 0.008) and port site metastases (p < 0.0001) were less common following the administration of intraperitoneal endotoxin. CONCLUSION: The results of this study suggest that the immune system plays a role in the genesis of port site metastases. A preventive role for endotoxin in patients undergoing laparoscopic cancer surgery, however, remains speculative. PMID- 10858468 TI - The impact of a full-time director of minimally invasive surgery: clinical practice, education, and research. AB - BACKGROUND: One of the biggest challenges of the laparoscopic surgery revolution is resident training. To enhance resident training, some programs have hired an experienced laparoscopic surgeon. This study documents the impact of this addition to our training program. METHODS: The number and types of laparoscopic cases, the number of laparoscopic training sessions, and the number of minimally invasive research projects were tabulated for 12-month periods before (period 1) and after (period 2) the arrival of the laparoscopic surgeon. RESULTS: Laparoscopic procedures increased from 524 (period 1) to 1,077 (period 2). Advanced procedures increased from 213 to 629. Laparoscopic training sessions increased from 2 to 11, and approved minimally invasive research projects increased from 0 to 7. CONCLUSIONS: The addition of an experienced laparoscopic surgeon in a resident training program increased laparoscopic cases in which the residents participate by more than 100%. Laparoscopic training sessions and minimally invasive research projects also increased measurably. PMID- 10858469 TI - Hepatic tumor spread of colorectal cancer in a laparoscopic animal model. AB - BACKGROUND: We devised a standardized animal model to study the impact of laparoscopic colorectal surgery on intrahepatic tumor cell growth. METHODS: The technique of laparoscopic surgery in the rat was extended by endoscopic inoculation of colon cancer cells (CC531) into the portal vein (1 x 10(4), 5 x 10(4), 1 x 10(5), 3 x 10(5), 5 x 10(5) cells/ml) of WAG/Rij rats (n = 25). As controls, five animals underwent laparotomy and open intraportal inoculation of 5 x 10(4) cells/ml. RESULTS: Hepatic tumor growth occurred after inoculation of 5 x 10(4), 1 x 10(5), 3 x 10(5), and 5 x 10(5) cells/ml. Extrahepatic tumor and conflating hepatic tumor was observed after the inoculation of 1 x 10(5), 3 x 10(5), and 5 x 10(5) cells/ml. Concentrations of 5 x 10(4) cells/ml injected either laparoscopically or via an open technique led to single hepatic tumor nodules. No tumor growth was seen after inoculation of 1 x 10(4) cells/ml. CONCLUSIONS: Laparoscopic intraportal tumor cell inoculation is a feasible technique to create hepatic metastases. The inoculation of 5 x 10(4) CC531 cells leads to reliable cell growth that can be used to investigate the impact of various laparoscopic techniques on tumor spread. PMID- 10858470 TI - Fluorescence diagnosis of endometriosis using 5-aminolevulinic acid. AB - BACKGROUND: The diagnosis of nonpigmented endometrial lesions by simple laparoscopic visualization is difficult and often inaccurate. We therefore sought to establish a new and more accurate method to visualize these nonpigmented peritoneal changes caused by endometriosis. METHODS: A total of 37 patients received 30 mg 5-aminolevulinic acid/kg body weight 10 to 14 hs prior to surgery. Laparoscopy was then performed using a D-light system (Storz, Tuttlingen, Germany). The findings were evaluated first in the white-light mode; the D-light system was then activated, and all areas of fluorescence were documented. Multiple specimens were obtained by biopsy. RESULTS: The sensitivity of the fluorescence diagnosis in detecting endometriosis in nonpigmented areas and normal-looking peritoneum is 100%, with a specificity of 75%. Diagnosis by simple visualization under white illumination has a sensitivity of only 69% and a specificity of 70%. Occult areas of endometriosis were discovered using fluorescence diagnosis. CONCLUSIONS: Our findings suggest that fluorescence diagnosis using 5-aminolevulinic acid is feasible and can improve the diagnosis of endometriosis in nonpigmented and occult endometrial lesions. Fluorescence diagnosis is a promising new tool in the diagnosis of endometriosis. PMID- 10858471 TI - Laparoscopic surgery preserves monocyte-mediated tumor cell killing in contrast to the conventional approach. AB - BACKGROUND: Experimental animal research shows that immunologic defenses against tumor cells are disturbed by surgical trauma, resulting in an increased rate of tumor implantation and the growth of subsequent metastases. Minimally invasive surgery is associated with a preservation of postoperative immunologic functions and, in animal models, with decreased tumor growth. The objective was to study the influence of several surgical procedures, approached conventionally and laparoscopically, on interleukin-6 (IL-6) and monocyte-mediated cytotoxicity (MMC). METHODS: Five groups of five patients each were included in this prospective study: laparoscopic cholecystectomy (minor trauma) group, Nissen fundoplication (laparoscopic and conventional as moderate trauma) groups, and sigmoid colectomy (laparoscopic and conventional as major trauma) groups. Preoperatively, 1 and 4 days after surgery, IL-6 and MMC against SW948 colon cancer cell line were determined. RESULTS: The IL-6 levels differed significantly between the three laparoscopic procedures (p = 0.004) and increased according to the degree of trauma. There was no significant difference in MMC between the three laparoscopic procedures. However, MMC was suppressed after conventional procedures and preserved after laparoscopic procedures (p = 0.001). There was no correlation between IL-6 levels and changes in MMC. CONCLUSIONS: More extensive laparoscopic procedures induce increased levels of IL-6, reflecting higher levels of trauma. Conventional surgical procedures result in depressed MMC in the postoperative period. After laparoscopic procedures, MMC is preserved. These findings may be of importance in preventing implantation and growth of cancer cells spread by surgical manipulation. PMID- 10858472 TI - Minilaparoscopically assisted placement of ventriculoperitoneal shunts. AB - BACKGROUND: Ventriculoperitoneal (VP) shunting remains the preferred treatment for hydrocephalus. Laparoscopic techniques to aid in the placement of the peritoneal portion of the catheter have been reported previously. We describe a minilaparoscopic VP shunt (MLVPS) insertion technique that facilitates directed placement of the peritoneal portion of the catheter in most patients, including those with obese abdomens previously subjected to surgery. In this study we review our experience with MLVPS placement. METHODS: All cases of MLVPS insertions at the University of Kentucky Medical Center and Lexington VA Hospital performed between February 1998 and March 1999 were reviewed retrospectively. A total of 27 patients (13 males and 14 females) ranging in age from 4 to 81 years (mean, 41 years) underwent VP shunting. The MLVPS insertion was performed via a 2 mm laparoscope and a separate 2-mm incision for catheter insertion using a venous introducer kit. In patients who had prior abdominal surgery, a 5-mm direct-view trocar was used. RESULTS: The MLVPS procedure was successful in 27 patients (100%). The mean number of prior shunts was 2 (range, 0-28). Of the 27 patients, 16 (59%) had undergone previous abdominal surgery. The mean operative time was 76 min (range, 19-155 min). There were no intra- or postoperative complications, and no mortalities. The follow-up period extended from 1 to 12 months. CONCLUSIONS: Findings show MLVPS placement to be safe and feasible. It allows accurate, directed placement of the VP shunt with a 2-mm laparoscope and a second 2-mm incision for shunt insertion. The procedure is associated with reduced trauma to the abdominal wall and minimal postoperative ileus. Long-term follow-up assessment of shunt function is planned. PMID- 10858473 TI - The preoperative predictability of the short esophagus in patients with stricture or paraesophageal hernia. AB - BACKGROUND: Esophageal shortening is a known complication of advanced gastroesophageal reflux disease that may preclude a tension-free antireflux procedure. A retrospective analysis was performed to test the accuracy of preoperative testing. METHODS: From September 1993 to December 1998, 39 patients underwent esophageal mobilization with intraoperative length assessment. Patients were selected on the basis of irreducible hiatal hernia, stricture formation, or both. Patients in the upright position with a fixed hiatal hernia larger than 5 cm on an esophagram were considered to have a short esophagus. Manometric length two standard deviations below the mean for height was considered abnormally short. RESULTS: In 31 patients, intraoperative mobilization was sufficient to allow the gastroesophageal junction to lie 2 cm below the diaphragmatic crus, so no esophageal-lengthening procedure was required. Eight patients with a short esophagus required an esophageal-lengthening procedure after complete mobilization. Two patients subsequently underwent intrathoracic migration of the gastroesophageal junction (GEJ), with recurrence of symptoms and required gastroplasty during the second surgery. An esophagram had a sensitivity of 66% and a positive predictive value of 37%, whereas manometric length had a sensitivity of 43% and a positive predictive value of 25% for the diagnosis of short esophagus. The preoperative endoscopic finding of either a stricture or Barrett's esophagus was the most sensitive test for predicting the need for a lengthening procedure. CONCLUSIONS: Manometry and esophagraphy are not reliable predictors of the short esophagus. Additional tests and/or tests combined with other parameters are needed. PMID- 10858474 TI - Laparoscopic nephrectomy in children. AB - BACKGROUND: Laparoscopic nephrectomy in the adult population is reported with increased frequency. We present our initial experience with laparoscopic nephrectomy in children. METHODS: Over a 2-year period, 11 nephrectomies were performed in nine children aged 16 months to 16 years (mean, 6.5 years). All patients were referred due to complications of a nonfunctioning kidney. Seven patients had recurrent urinary tract infections, and two had refractory hypertension. Two patients underwent bilateral laparoscopic nephrectomy. The operation was performed using four access ports measuring 3.5 to 10 mm. RESULTS: All kidneys were removed successfully using a laparoscopic technique. The average length of the operation was 163 min per kidney (range, 90-420). The estimated blood loss was <10-150 ml (mean, 45). No patient required transfusion. Seven patients were discharged home by postoperative day 2. The two patients with the longest operating times were discharged home on postoperative days 4 and 5 due to delay in return of bowel function. Narcotic use was minimal, and all patients enjoyed a rapid return to full activity. CONCLUSION: Laparoscopic nephrectomy is a viable alternative to open nephrectomy in children. Further experience with this technique is required to establish its efficacy and reduce the operating time PMID- 10858475 TI - A comparison of laparoscopic Nissen fundoplication and Rossetti's modification in 239 patients. AB - BACKGROUND: Laparoscopic Nissen fundoplication and the Rossetti modification represent two different surgical approaches to resolving gastroesophageal reflux disease (GERD). Concerns have arisen that the Rossetti modification results in increased postoperative dysphagia. In this study, we compared a group of patients who underwent a laparoscopic Nissen fundoplication with a group who had undergone the Rossetti modification to determine if there was a significant difference in postoperative dysphagia. Additionally, we wanted to confirm that the Nissen procedure performed laparoscopically could resolve GERD as successfully as the Rossetti modification, with no difference in operative complications. METHODS: We prospectively collected data on 101 patients who underwent laparoscopic Nissen fundoplication and compared outcomes with those of 138 patients who had undergone the laparoscopic Rossetti modification in a previous series. RESULTS: All patients experienced resolution of reflux symptoms. No statistically significant differences were found between the groups in terms of intraoperative or postoperative complications, conversions to open procedure, or length of hospitalization. Paradoxically, there was a significant difference in operating time between the Rossetti and the Nissen groups (70.6 min vs 45.6 min, p = 0.006). Postoperative dysphagia requiring dilation was significantly higher in the Rossetti group (21.7% vs 8.9%, p = 0.008). However, there was a significantly higher percentage of patients in the Rossetti group who had had esophagitis preoperatively (95.7% vs 86.1%, p = 0.009), although the proportion of patients having Barrett's esophagus was higher in the Nissen group (9.4% vs 24.8%, p = 0.001). CONCLUSIONS: Both approaches resolved reflux symptoms without significant differences in complications, conversions, or length of stay. Preoperative differences between groups, as well as the method of sequentially comparing the two different procedures, prevent us from attributing greater postoperative dysphagia in the Rossetti group solely to the choice of surgical approach. Prospective randomized studies are needed to control for variables, such as surgical team experience and patient differences. PMID- 10858476 TI - Laparoscopic management of mechanical small bowel obstruction: are there predictors of success or failure? AB - BACKGROUND: Laparoscopy is used increasingly for the management of acute abdominal conditions. For many years, previous abdominal surgery and intestinal obstruction have been regarded as contraindications to laparoscopy because there is an increased risk of iatrogenic bowel perforation. The role of laparoscopy in acute small bowel obstruction remains unclear. METHODS: Since 1995, data from patients undergoing laparoscopic surgery have been entered prospectively into a database. Patients who underwent surgery before 1995 were added retrospectively to the same database. The charts of all patients treated surgically for mechanical small bowel obstruction were reviewed. Univariate analysis was performed to identify factors associated with success or failure, especially intraoperative complications, conversion, and postoperative morbidity. Stepwise logistic regression was used to assess for independent variables. RESULTS: This study included 83 patients (56 women and 27 men) with a mean age of 56 years (range, 17-91 years). Conversion was necessary in 36 cases (43%). Laparoscopy alone was successful in 47 patients (57%). Intraoperative complications were noted in 16% and postoperative complications in 31% of the patients. Eight reoperations (9%) were necessary. Mortality was 2.4%. Duration of surgery (p < 0.001) and a bowel diameter exceeding 4 cm (p = 0. 02) were predictors of conversion. No risk factor for intraoperative complication was identified. Accidental bowel perforation (p = 0. 008) and the need for conversion (p = 0.009) were the only independent factors associated with an increased risk of postoperative complications. CONCLUSIONS: Laparoscopic management of small bowel obstruction is possible in roughly 60% of the patients selected for this approach. Morbidity is lower, resumption of a normal diet is faster, and hospital stay is shorter than with patients requiring conversion. No clear predictor of success or failure was identified, but intraoperative complications must be avoided. If the surgeon is widely experienced in advanced laparoscopic surgery and there is a liberal conversion policy, laparoscopy is a valuable alternative to conventional surgery in the management of acute small bowel obstruction. PMID- 10858477 TI - Emergency laparoscopy: a community hospital experience. AB - BACKGROUND: By now, laparoscopic surgery has achieved widespread acceptance among surgeons and, generally speaking, by the public. Therefore, we set out to evaluate whether this technique is a feasible method of treating patients with abdominal emergencies, traumatic or not. To assess the routine use of emergency laparoscopy in a community hospital setting, we undertook a retrospective analysis of an unrandomized experience (presence or absence of a surgeon with laparoscopic experience). METHODS: Between January 1993 and October 1998, 575 emergency abdominal surgical procedures were done in our department. In all, 365 (63.4%) were diagnostic and operative laparoscopy procedures (acute small bowel obstruction: 23 cases; hernia disease: one case; gastroduodenal ulcer disease: 15 cases; biliary system disease: 89 cases; pelvic disease: 237 cases). These cases represent almost 56% of all laparoscopic procedures done during the same period at our institution. Laparoscopy was not performed in patients with a history of a previous abdominal approach to malignant disease, a history of more than two major abdominal surgeries, or massive bowel distension; nor was it used in patients whose general conditions contraindicate this approach. RESULTS: The conversion rate was 6.8%. The morbidity and mortality rates were, respectively, 4.1% and 0.8%. A definitive diagnosis was provided in 95.3% of cases, with the possibility to treat 88.2% of them by laparoscopy. CONCLUSIONS: We consider the laparoscopic approach in patients with abdominal emergencies to be feasible and safe in experienced hands. It provides diagnostic accuracy as well as therapeutic capabilities. Sparing patients laparotomy reduces postoperative pain, improves recovery of GI function, reduces hospitalization, cuts health care costs, and improves cosmetic results. This approach promises to play a significant role in emergency abdominal situations and will certainly become increasingly important in today's health care environment. PMID- 10858478 TI - Laparoscopy in the evaluation of the incarcerated mass in groin hernia. AB - BACKGROUND: In hernia patients, the preoperative diagnosis of strangulation is difficult. In this prospective study, we investigated the usefulness of an exploratory laparoscopy to evaluate the viability of a viscus incarcerated in a groin hernia. METHODS: Twenty-seven patients with an acute irreducible inguinal mass underwent exploratory laparoscopy. The hernia was reduced, and the viability of the incarcerated viscus was judged laparoscopically on the basis of color, congestion, and contractility. RESULTS: Twenty-four hernias were found. In sixteen patients, the contents of the hernia were viable. In five patients, a necrotic bowel segment was found, and a laparotomy and resection were done. In three patients, no hernias were found. The cause of inguinal pain was spermatic cord hematoma in one patient and inguinal abscess in another; however, the cause of pain in the third patient remained unclear. After laparotomy, one patient developed a fascial rupture that required reoperation. There were no other complications. CONCLUSION: At laparoscopy, the judgment of the viability of the contents of the hernia is similar to that at laparotomy. The early use of laparoscopy can prevent many unnecessary laparotomies. PMID- 10858479 TI - A serum-soluble factor(s) stimulates tumor growth following laparotomy in a murine model. AB - BACKGROUND: Our laboratory and others have previously demonstrated that tumors grow larger and are more easily established following laparotomy than after CO(2) pneumoperitoneum. The etiology of increased tumor growth after surgery is unknown. We hypothesized that, following laparotomy, a serum soluble factor(s) is generated that causes tumors to proliferate more rapidly. The purpose of the current study was to determine if in vitro tumor cells proliferate faster when incubated with serum from laparotomized mice than cells incubated with sera from mice who have undergone CO(2) pneumoperitoneum or anesthesia alone. METHODS: In the first experiment, female Balb/C mice (n = 84) were randomly divided into the following three groups: (a) control (AC), (b) CO(2) insufflation (INS), and (c) laparotomy (OPEN). The AC mice underwent no procedure. The INS group underwent CO(2) pneumoperitoneum at 4-6 mmHg for 20 min. The OPEN group had a midline incision from xiphoid to pubis. The serum of seven mice from each group were collected on postoperative days (POD) 1, 2, 4, and 7 via a cardiac puncture. The sera at each time point for each group were pooled. Twenty thousand C-26 colon cancer cells were incubated separately in growth media containing 10% mouse serum from each group (seven determinations/group) at each time point. In the second experiment, female Balb/C (n = 30) mice were divided into AC and OPEN groups. On POD4, sera were collected and pooled. Three separate studies were performed for the second experiment. In the first study, tumor cells were incubated with 10% AC sera or varying concentrations of OPEN mice sera (4-10%). In the second study, aliquots of sera from the OPEN group mice were then heated at 100 degrees C for 1 or 5 min. Tumors were then incubated separately in media with 10% AC, OPEN, or heated OPEN group sera. In the third study, aliquots of sera from the OPEN group mice were dialyzed against PBS through a 3.5-kD or an 8-kD dialysis membrane tubing for 24 h. Tumors were then incubated separately in media with 10% AC, OPEN, or dialyzed OPEN group sera. For both experiments, tumor proliferation was determined and compared between groups after 72 h of incubation. RESULTS: Tumor cells incubated with POD2 and POD4 sera from OPEN group mice proliferated twice as fast as those incubated with sera from either AC or INS group mice. The difference in proliferation was maximal on POD4 and started to decline by POD7. Proliferative activity from the OPEN group sera decreased significantly when heated for 1 min and was completely ablated after 5 min of heating. Proliferative activity from the OPEN group sera was completely ablated after dialysis. CONCLUSIONS: We conclude that there is a serum-soluble factor(s) present postoperatively that stimulates tumors to grow significantly faster after laparotomy. The mitogenic effect of laparotomized mice sera is dilutable. It is uncertain whether the factor is heat labile, since heating most likely destroys other necessary proteins in the sera. The size of the factor is undeterminable using the dialysis method. Further efforts to identify these factors are currently underway. PMID- 10858480 TI - The role of laparoscopy in preoperative staging of esophageal cancer. AB - BACKGROUND: Diagnostic laparoscopy has been used to determine resectability and to prevent unnecessary laparotomy in patients with advanced esophageal cancer. The objective of this prospective study was to evaluate the role of laparoscopy in conjunction with computed tomography (CT) scan in staging patients with esophageal cancer. METHODS: From March 1995 to October 1998, 59 patients with biopsy-proven esophageal cancer underwent diagnostic laparoscopy with concurrent vascular access device and feeding jejunostomy tube placement. RESULTS: Laparoscopy changed the treatment plan in 10 of 59 patients (17%). Of the patients with normal-appearing regional or celiac nodes, 78% were confirmed by biopsy to be tumor free, whereas 76% of patients with abnormal-appearing nodes were confirmed by biopsy to have node-positive disease. CONCLUSIONS: Diagnostic laparoscopy is useful for detecting and confirming nodal involvement and distant metastatic disease that potentially would alter treatment and prognosis in patients with esophageal cancer. PMID- 10858482 TI - A simple laparoscopic method to provide access to the gastroesophageal junction in obese patients. AB - By now, the feasibility of laparoscopic surgery in obese patients is well established; a conversion rate of 1.4-4.3% has been reported [1, 2]. The main reason for conversion in these cases is the difficulty encountered in exposing the gastroesophageal junction due to a huge fatty liver that covers the entire upper abdomen ("the invisible stomach" [1]). We report here a simple method that allows easy access to the upper stomach in such cases. This technique involves the exposure of the gastroesophageal junction using a laparoscopic suprahepatic route. PMID- 10858483 TI - Laparoscopic intragastric surgery using a radially expandable sleeve. AB - Laparoscopic intragastric surgery (LIGS) has become more widely established in Japan as a therapy for early gastric cancers and some types of submucosal gastric tumors. However, there have been some technical difficulties with the original method of LIGS. Certain complicated procedures to access the gastric lumen are required, along with repair of the gastric wall after endoluminal procedures. Using a 5-mm radially expandable sleeve (RES) for the working ports in LIGS, it becomes easier to establish access to the gastric lumen, and repair of the port sites on the gastric wall is not required. Using RES makes LIGS a simpler, less invasive procedure. PMID- 10858484 TI - Left side thoracoscopically assisted gastroplasty: a new technique for managing the shortened esophagus. AB - Laparoscopic antireflux surgery is the procedure of choice for gastroesophageal reflux disease (GERD). However, many clinicians have reservations about its application in patients with complicated GERD, notably those with esophageal shortening. In this report, we present our experience with the laparoscopic management of the shortened esophagus. A total of 235 patients with primary GERD underwent laparoscopic antireflux procedures, 38 of whom were suspected preoperatively to have a shortened esophagus. Of the 235 patients, 8 (3.4%) needed a left thoracoscopically assisted gastroplasty in addition to laparoscopic Toupet repair (n = 4) or Nissen fundoplication (n = 4). Complications included pleural effusion (n = 1), pneumothorax (n = 2), and minor atelectasis (n = 1). The average hospital stay was 3 days. Results were satisfactory in 7 of 8 patients, with a mean follow-up of 20.2 months (range, 9-34 months). The surgical management of the shortened esophagus is difficult. However, the role of minimally invasive techniques is justified. Early results are appealing, with less morbidity, satisfactory control of GERD related symptoms, and a shortened hospital stay. PMID- 10858487 TI - Bridging the treatment gap. AB - The leading cause of death and disability in the United States today is cardiovascular disease (CVD). The main risk factor, hypercholesterolemia, is grossly undertreated, although it is widely appreciated that lowering cholesterol levels is key to reducing the incidence of CVD. Cholesterol-lowering therapy decreases total mortality, cardiovascular events, the need for revascularization procedures, and hospitalization costs. A 25-35% reduction of low-density lipoprotein (LDL) indicates significant benefits with regard to morbidity and mortality. Unfortunately, most patients who are candidates for cholesterol lowering treatment do not receive it. Wider use of lipid-lowering agents could, in fact, make a significant difference in patient outcomes. There is strong interest on the part of consumers in over-the-counter (OTC) cholesterol-lowering products that may help them reduce their risk of developing heart disease and live healthier lives. Surveys estimate that half of all patients with high cholesterol would like to have an OTC statin product made available. Increased availability of cholesterol-lowering therapies as well as changes in physician prescribing practices could benefit a broad spectrum of the population that is currently untreated or undertreated. Current prescribing practices and guidelines have not resulted in widespread use of these therapies; therefore, outcomes for CVD prevention remains suboptimal. The proposed advantages of making statins available over-the-counter include their known efficacy, dose consistency, and proven safety profile. PMID- 10858488 TI - Insights on treating an over-the-counter-type subgroup: data from the Air Force/Texas Coronary Atherosclerosis Prevention Study Population. AB - The expansion of therapeutic options for management of dyslipidemia is a potentially valuable avenue for the optimal treatment of most patients at low-to moderate risk for coronary artery disease (CAD). In primary prevention, this population is closely approximated by that of the landmark Air Force/Texas Coronary Atherosclerosis Prevention Study (AFCAPS/TexCAPS). In AFCAPS/TexCAPS, 6,605 men and women without evidence of CAD and with average total cholesterol (180-264 mg/dL) and low-density lipoprotein (LDL)-cholesterol (130-190 mg/dL) concentrations and low high-density lipoprotein (HDL)-cholesterol levels (< or =45 mg/dL for men, < or =47 mg/dL for women) were treated with either lovastatin or placebo for a mean of 5.2 years. With few exceptions, the characteristics of the AFCAPS/TexCAPS cohort were similar to the profile of the majority of people in the United States and that of a potential over-the-counter (OTC)-type subgroup. The dosage of lovastatin used was 20-40 mg/day, titrated to achieve an LDL-cholesterol target of < or =110 mg/dL. Treatment reduced the combined incidence of fatal and nonfatal myocardial infarction, unstable angina, and sudden cardiac death by 37% (p<0.001). The risk for fatal and nonfatal heart attack was reduced by 40% (p<0.002), and the need for coronary revascularization procedures was reduced by 33% (p = 0.01). Post hoc analysis of data from a subgroup of the AFCAPS/TexCAPS cohort resembling those in the general population who may benefit from OTC statins indicates similar benefits. The results have important implications for the identification and treatment of persons at risk for coronary disease. PMID- 10858489 TI - Defining patient risks from expanded preventive therapies. AB - In clinical trials, all lipid-lowering agents have been associated with mild, asymptomatic elevations of alanine aminotransferase (ALT) and asparate aminotransferase enzymes. This, along with the fact that 3-hydroxy-3 methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors are hepatotoxic in some animals, led the US Food and Drug Administration (FDA) to recommend monitoring of liver enzymes for all lipid-lowering agents, except the bile acid sequestrants. Because the drugs act by different mechanisms, ALT elevations may be a pharmacodynamic effect related to lipid lowering, rather than a direct effect of the drug. Animal studies support this assumption. ALT elevations of 3 times the upper limit of normal occur in <3% of patients in clinical trials of lipid lowering drugs. The elevations are transient and often dose-related, and they usually revert to normal while continuing therapy and have never been associated with hepatotoxicity. Confounding factors include alcohol, acetaminophen, and pre existing liver disease, such as chronic hepatitis C and type II diabetes with fatty liver, which are both associated with mild, intermittent elevations of ALT. The more important issue is whether or not lipid-lowering agents are hepatotoxic. There are case reports of hepatotoxicity (cholestasis, jaundice, hepatitis, chronic active hepatitis, fatty liver, cirrhosis and acute liver failure) with all of the drugs, except cholestyramine. To date there are just 5 cases of documented liver failure linked to lovastatin. There is no evidence that monitoring reduces the rate of hepatotoxicity. Mild elevations of ALT that occur with many drugs, including HMG-CoA reductase inhibitors, do not predict hepatotoxicity. Liver enzyme elevations appear to be a class characteristic of lipid-lowering agents. Hepatotoxicity is a rare idiosyncratic reaction, occurring only with sustained released nicotinic acid. PMID- 10858490 TI - Population benefits of cholesterol reduction: epidemiology, economics, and ethics. AB - Cardiovascular disease mortality-rate reductions have slowed in the United States in the last decade, suggesting that additional strategies are needed to reduce rates further. Population-wide cholesterol reduction is a promising approach. Selection of a particular strategy is less an issue of efficacy, which has been proven through numerous studies, than it is an issue of epidemiology, economics, and ethics. These 3 imperatives constitute the foundation of renewed efforts to reduce the US population's cholesterol levels. Epidemiologic imperatives include risk reduction in low-to-moderate risk individuals, who comprise approximately 30% of the population and one third of incident cases of coronary disease. Any cholesterol-lowering strategy must address the challenge of reducing the incidence of coronary disease; to do otherwise will result in an increasing prevalence of disease, with the attendant cost and disability burdens. Economic imperatives include the extension of preventive coverage to the low-to-moderate risk segment of the population, which currently is not included in any risk reduction programs. Although cholesterol reduction with pharmacologic agents may not meet current standards for cost-effectiveness, over-the-counter (OTC) agents are under the rubric of individual, not societal, costs. Finally, current and proposed options for nonprescription cholesterol-lowering drugs raise a number of ethical issues such as beneficence, nonmaleficence, justice, and autonomy. Population-wide cholesterol reduction must be a mainstay for any strategy to reduce the burden of cardiovascular disease. PMID- 10858491 TI - VIP-derived sequences modified by N-terminal stearyl moiety induce cell death: the human keratinocyte as a model. AB - Vasoactive intestinal peptide (VIP) is a recognized growth factor affecting many cell types. We have previously developed a series of lipophilic VIP analogues containing an N-terminal covalently attached stearyl moiety. The current studies identified stearyl-Nle(17)-VIP and stearyl-Nle(17)-neurotensin(6-11)VIP(7-28), acting at microM concentrations, as cytotoxic to human keratinocytes. The core C terminal active VIP-derived peptide, stearyl-Lys-Lys-Tyr-Leu-NH(2) (St-KKYL NH(2)), was identified as being responsible for the observed cytotoxicity. Cytotoxicity coincided with marked reduction in intracellular cyclic GMP and was abolished by co-treatment with the endonuclease inhibitor, aurine-tricarboxylic acid, suggesting apoptotic mechanisms. Stearyl-VIP derivatives thus offer lead compounds for future drug development against hyperproliferative skin conditions. PMID- 10858492 TI - VIP and the potent analog, stearyl-Nle(17)-VIP, induce proliferation of keratinocytes. AB - Vasoactive intestinal polypeptide (VIP) exhibits effects on cell proliferation. Here, VIP, as well as the related peptide, pituitary adenylate cyclase activating peptide (PACAP), promoted human keratinocyte division. Stearyl-Nle(17)-VIP (SNV) was identified as a superior mitogen for the keratinocytic cell line, HaCaT, both in potency (fM-nM concentrations) and efficacy. Reverse transcription-polymerase chain reaction detected in keratinocytes only PACAP mRNA and the relevant type 1 (VPAC(1)R) and type 2 (VPAC(2)R) receptors, while VIP and the third receptor (PAC(1)) transcripts were absent. Upon serum deprivation of HaCaT, the VPAC(1)R mRNA was apparently increased, while the VPAC(2)R transcript remained constant. Incubation of HaCaT with VIP or SNV increased nitric oxide and cGMP formation. In contrast to VIP, SNV did not augment cAMP. Thus, the paracrine VIP, and autocrine PACAP, related pathways leading to keratinocyte proliferation may involve VPAC(1)R/VPAC(2)R and nitric oxide/cGMP production. PMID- 10858493 TI - Characterisation of oxidative phosphorylation in skeletal muscle mitochondria subpopulations in pig: a study using top-down elasticity analysis. AB - In skeletal muscle, two mitochondrial populations are present which, on the basis of their localisation, are termed intermyofibrillar and subsarcolemmal mitochondria (IMF and SS, respectively). These two populations have different biochemical characteristics and show different responses to physiological stimuli. In this paper, we characterise the oxidative phosphorylation of SS and IMF using 'top-down' elasticity analysis. We excluded the possibility that their different characteristics can be attributed to a different degree of breakage of the two types of mitochondria due to the different isolation procedures used in their preparation. The higher respiration rate and higher respiratory control ratio shown by IMF compared with those shown by SS are principally due to the higher activities of the reactions involved in substrate oxidation as confirmed by the measurement of cytochrome oxidase activity. There is no difference in the leak of protons across the inner mitochondrial membrane between IMF and SS; a faster rate of ATP synthesis and turnover is driven by the lower membrane potential in SS compared with in IMF. PMID- 10858494 TI - Protein disulfide isomerase mediates integrin-dependent adhesion. AB - Cell adhesion is mediated by the integrin adhesion receptors. Receptor-ligand interaction involves conformational changes in the receptor, but the underlying mechanism remains unclear. Our earlier work implied a role for sulfhydryls in integrin response to ligand binding in the intact blood platelet. We now show that non-penetrating blockers of free sulfhydryls inhibit beta(1) and beta(3) integrin-mediated platelet adhesion regardless of the affinity state of the integrin. Removal of the inhibitors prior to adhesion fully restores adhesion despite the irreversible nature of inhibitor-thiol interaction, indicating sulfhydryl exposure in response to adhesion. We further show that blocking protein disulfide isomerase (PDI) inhibits adhesion. These data indicate that: (a) ecto-sulfhydryls are necessary for integrin-mediated platelet adhesion; (b) disulfide exchange takes place during this process; (c) surface PDI is involved in integrin-mediated adhesion. PMID- 10858495 TI - A new route to peroxynitrite: a role for xanthine oxidoreductase. AB - Peroxynitrite, a potent oxidising, nitrating and hydroxylating agent, results from the reaction of nitric oxide with superoxide. We show that peroxynitrite can be produced by the action of a single enzyme, xanthine oxidoreductase (XOR), in the presence of inorganic nitrite, molecular oxygen and a reducing agent, such as pterin. The effects of oxygen concentration on peroxynitrite production have been examined. The physiologically predominant dehydrogenase form of the enzyme is more effective than the oxidase form under aerobic conditions. It is proposed that XOR-derived peroxynitrite fulfils a bactericidal role in milk and in the digestive tract. PMID- 10858496 TI - A novel gene derived from developing spinal cords, SCDGF, is a unique member of the PDGF/VEGF family. AB - We isolated a novel gene designated spinal cord-derived growth factor (SCDGF). Its expression was increased in chick spinal cords with embryonic development and decreased after hatching. The amino acid sequences of chick and human SCDGFs revealed a putative signal sequence followed by a CUB domain and a region homologous to the members of the platelet-derived growth factor/vascular endothelial growth factor family. Furthermore, human SCDGF secreted from the cells showed a mitogenic activity for 10T1/2 cells in vitro. These results led us to speculate that SCDGF plays an important role in the development of the spinal cord. PMID- 10858497 TI - Delayed fluorescence emitted from light harvesting complex II and photosystem II of higher plants in the 100 ns-5 micros time domain. AB - This study presents the first report on delayed fluorescence (DF) emitted from spinach thylakoids, D1/D2/Cytb-559 preparations and solubilized light harvesting complex II (LHCII) in the ns time domain after excitation with saturating laser flashes. The use of a new commercially available multichannel plate with rapid gating permitted a sufficient suppression of detector distortions due to the strong prompt fluorescence. The following results were obtained: (a) in dark adapted thylakoids, the DF amplitudes at 100 ns and 5 micros after each flash of a train of saturating actinic pulses exhibit characteristic period four oscillations of opposite sign: the DF amplitudes at 100 ns oscillate in the same manner as the quantum yield of prompt fluorescence, whereas those at 5 micros resemble the oscillation of the micros kinetics of P680(.) reduction in samples with an intact water oxidizing complex, (b) the quantum yield of total DF emission in the range up to a few micros is estimated to be <10(-4) for thylakoids, (c) the DF of D1/D2/Cytb-559 exhibits a monophasic decay with tau approximately 50 ns, (d) DF emission is also observed in isolated LHCII with biphasic decay kinetics characterized by tau values of 65 ns and about 800 ns, (e) in contrast to thylakoids, the amplitudes of DF in D1/D2/Cytb-559 preparations and solubilized LHCII do not exhibit any oscillation pattern and (f) all spectra of DF from the different sample types are characteristic for emission from the lowest excited singlet state of chlorophyll a. The implications of these findings and problems to be addressed in future research are briefly discussed. PMID- 10858498 TI - Identification of alternative splicing variants of the beta subunit of human Ca(2+)/calmodulin-dependent protein kinase II with different activities. AB - The beta subunit of human Ca(2+)/calmodulin-dependent protein kinase II (beta CaMKII) was identified by searching through an expressed sequence tag database and rapid amplification of cDNA 5'-ends and was assigned to chromosome 7. Reverse transcription-polymerase chain reaction and sequencing analysis identified at least five alternative splicing variants of beta CaMKII (beta, beta6, betae, beta'e, and beta7) in brain and two of them (beta6 and beta7) were first detected in any species. When expressed in HEK 293 cells, the Ca(2+)/calmodulin-dependent kinase activity of beta7, the shortest variant, was much lower than that of either beta (the longest one) or betae (the medium one), suggesting possible regulation of beta CaMKII activity by alternative splicing. PMID- 10858499 TI - The mnn2 mutant of Saccharomyces cerevisiae is affected in phosphorylation of N linked oligosaccharides. AB - We studied the phosphorylation of the inner core region of N-linked oligosaccharides in the mannan defective mutant Saccharomyces cerevisiae mnn2 which was described as unable to synthesize branches on the outer chain. We performed structural studies of the N-oligosaccharides synthesized by the strains mnn2, mnn1mnn2mnn9 and mnn1mnn9ldb8, and the results are compared with previously published structural data of mnn1mnn2mnn10 and mnn1mnn9 [Hernandez, L.M., Ballou, L., Alvarado, E., Tsai, P.-K. and Ballou, C.E. (1989) J. Biol. Chem. 264, 13648 13659]. We conclude that the mnn2/ldb8 mutation is responsible for the inhibition of incorporation of phosphate to mannose A(3) (see below), a particular phosphorylation site of the inner core, while phosphorylation at the other possible site (mannose C(1)) is allowed, although it is also reduced. *Phosphorylation sites in mnn1mnn9. (see structure below) PMID- 10858500 TI - Distinctive properties of the catalase B of Aspergillus nidulans. AB - Aspergillus nidulans catalase B (CatB) was purified to homogeneity and characterized as a hydroperoxidase which resembles typical catalases in some physicochemical characteristics: (1) it has an apparent molecular weight of 360000 and is composed of four glycosylated subunits, (2) it has hydrophobic properties as revealed by extractability in ethanol/chloroform and binding to phenyl-Superose, and (3) it has an acidic isoelectric point at pH 3. 5. Also CatB exhibits some distinctive properties, e.g. it is not inhibited by the presence of 2% sodium dodecyl sulfate, 9 M urea or reducing agents. Furthermore, even though CatB does not exhibit any residual peroxidase activity, it is able to retain up to 38% of its initial catalase activity after incubation with the typical catalase inhibitor 3-amino-1,2,4-triazole. PMID- 10858501 TI - Estimation of H2O2 gradients across biomembranes. AB - When cells are exposed to an external source of H2O2, the rapid enzymatic consumption of H2O2 inside the cell provides the driving force for the formation of the gradient across the plasma and other subcellular membranes. By using the concepts of enzyme latency, the following gradients - formed after a few seconds following the exposure to H2O2 - were estimated in Jurkat T-cells: [H2O2](cytosol)/[H2O2](peroxisomes)=3; [H2O2](extracellular)/[H2O2](cytosol)=7. The procedure presented in this work can easily be applied to other cell lines and provides a quantitative framework to interpret the data obtained when cells are exposed to an external source of H2O2. PMID- 10858502 TI - Absorption and metabolism of genistin in the isolated rat small intestine. AB - Uptake and intestinal metabolism of physiologically active genistin were studied in an ex vivo intestinal perfusion model; luminally applied concentrations were 5.9, 12.0, and 23.8 micromol/l. The intestinal absorption of genistin was 14.9% (+/-2.3, n=9), irrespective of the amounts applied. The majority of the absorbed genistin appeared as genistein glucuronide (11.6%), also recovered as the main metabolite on the luminal side (19.5%). Minor amounts of genistin (1.3%) and genistein (1.9%) were found on the vascular side, whereas 15.4% of applied genistin was luminally cleaved to yield genistein. Sulfate derivatives of genistein or genistin were not observed. PMID- 10858503 TI - A novel Ser O-glucuronidation in acidic proline-rich proteins identified by tandem mass spectrometry. AB - Human acidic proline-rich salivary protein PRP-1 and its C-terminally truncated form PRP-3 were analyzed by electrospray tandem mass spectrometry. Post translational modifications were detected and characterized. A pyroglutamic acid residue was demonstrated at the N-terminus, Ser-8 and Ser-22 were shown to be phosphorylated and an O-linked glucuronic acid conjugation was identified. The latter modification was located to Ser-17 and found to be present in approximately 40% of the polypeptides. PMID- 10858504 TI - 1H, 15N and (13)C assignments and secondary structure of the EGF-like module pair 3-4 from vitamin K-dependent protein S. AB - Vitamin K-dependent protein S, which is a cofactor for activated protein C and thus important for down-regulation of the coagulation cascade, contains several Ca(2+)-binding sites with unusually high affinity. The 89 amino acid fragment constituting the third and fourth epidermal growth factor-like (EGF) modules of protein S is the smallest fragment that retains high-affinity Ca(2+) binding and is therefore useful for investigating the structural basis of this property. Heteronuclear multidimensional nuclear magnetic resonance experiments were used to obtain extensive assignments of the (1)H, 15N and (13)C resonances of the module pair with one Ca(2+) bound in EGF 4. In addition, nearly complete assignments of the (1)H resonances of the isolated Ca(2+)-free EGF 3 module were obtained. The assignment process was complicated by broadening of several resonances, spectral heterogeneity caused by cis-trans isomerisation of the peptide bond preceding Pro-168, and dimerisation. Analysis of weighted average secondary chemical shifts, (3)J(HNHalpha) coupling constants, and NOE connectivities suggest that both EGF modules in this fragment adhere to the classical secondary structure of EGF modules, consisting of one major and one minor anti-parallel beta-sheet. PMID- 10858505 TI - The presence and subcellular localization of caspase 3-like proteinases in plant cells. AB - Caspases play a very important role in initiating and executing apoptotic processes in animal cells. In this study we show that plant mitochondria were able to initiate the activation of caspase 3 in a Xenopus cell free system. Caspase 3-like activity was found to be present in plant cells and could only be inhibited by the specific caspase 3 inhibitor N-acetyl-Asp-Glu-Val-Asp fluoromethylketone (Ac-DEVD-fmk) and not by cysteine protease inhibitors. By micro-injection of the caspase 3 substrate in living Chara cells we showed that caspase 3-like activity was mainly present in the cytosol rather than in the vacuole. This is the first time that in vivo caspase 3-like activity has been demonstrated in plants. PMID- 10858506 TI - In vivo measurements of control coefficients for hexokinase and glucose-6 phosphate dehydrogenase in Xenopus laevis oocytes. AB - Hexokinase and glucose-6-phosphate dehydrogenase activities were increased in Xenopus laevis oocytes by microinjection of commercial pure enzymes. The effect of increased fractional activities on glycogen synthesis or on the production of 14CO(2) (the oxidative portion of the pentose phosphate pathway) was investigated by microinjection of [1-(14)C]glucose and measurements of the radioactivity in glycogen and CO(2). Control coefficients calculated from the data show that hexokinase plays an important role in the control of glycogen synthesis (control coefficient=0.7) but its influence on the control of the pentose phosphate pathway is almost nil (control coefficient=-0.01). Glucose-6-phosphate dehydrogenase injections did not affect the production of 14CO(2) by the pentose phosphate pathway, indicating that other factors control the operation of this pathway. In addition, an almost null control of this enzyme on glycogen synthesis flux was observed. PMID- 10858507 TI - Characterization of NPY receptors controlling lipolysis and leptin secretion in human adipocytes. AB - In order to characterize neuropeptide Y (NPY) receptors present in human adipocytes, we used selective ligands together with specific molecular probes able to recognize the different NPY receptor subtypes. RT-PCR experiments revealed the presence of Y(1) receptor transcripts with Y(4) and Y(5) and absence of Y(2) signals. Binding studies, using selective radioiodinated ligands, detected a high number (B(max)=497+/-124 fmol/mg protein) of a high affinity binding site only with [(125)I]peptide YY (PYY) and [(125)I](Leu(31), Pro(34))PYY. These sites exhibited a typical Y(1) profile as indicated by the rank order of affinity of NPY analogs and the high affinity of two selective NPY receptor antagonists, SR120819A and BIBP3226. In [(35)S]GTPgammaS binding experiments, PYY activation was totally inhibited by SR120819A and BIBP3226. Both compounds antagonized, with similar efficiency, the antilipolytic effect exerted by NPY in isolated adipocytes. Finally, PYY and Y(1) ligands enhanced adipocyte leptin secretion, an effect totally prevented by SR120819A. Thus, highly expressed in human adipocytes, the Y(1) receptor sustains the strong antilipolytic effect of NPY and exerts a positive action on leptin secretion. PMID- 10858508 TI - Nine novel precursors of Buthus martensii scorpion alpha-toxin homologues. AB - The cDNAs encoding nine novel alpha-toxin homologues were isolated from the venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch (BmK). They are rich in AAAA and TTTT elements at the 5' UTRs. The flanking region of the translation initiation codon ATG is AAAATGAA, which is highly conserved in scorpion Na(+), K(+) and Cl(-) channel toxin genes. These putative scorpion alpha toxins shared 45.5-98.4% homology with the characterized BmK alpha-toxins, and were completely conserved in the positions of all eight cysteines. This showed, together with higher homology at nucleotide level than that at amino acid level, that these toxins may originate from a common ancestor. The discovery of a series of homologues of scorpion alpha-toxin with a different degree of natural mutation in the primary structure will provide us with a valuable system for studying the structure-function relationship of scorpion toxins. PMID- 10858509 TI - Characterization of venom (Duvernoy's secretion) from twelve species of colubrid snakes and partial sequence of four venom proteins. AB - R.E. Hill and S.P. Mackessy. Characterization of venom (Duvernoy's secretion) from twelve species of colubrid snakes and partial sequence of four venom proteins. Toxicon XX, xx-yy, 2000. - Venomous colubrids, which include more than 700 snake species worldwide, represent a vast potential source of novel biological compounds. The present study characterized venom (Duvernoy's gland secretion) collected from twelve species of opisthoglyphous (rear-fanged) colubrid snakes, an extremely diverse assemblage of non-venomous to highly venomous snakes. Most venoms displayed proteolytic activity (casein), though activity levels varied considerably. Low phosphodiesterase activity was detected in several venoms (Amphiesma stolata, Diadophis punctatus, Heterodon nasicus kennerlyi, H. n. nasicus and Thamnophis elegans vagrans), and acetylcholinesterase was found in Boiga irregularis saliva and venom, but no venoms displayed hyaluronidase, thrombin-like or kallikrein-like activities. High phospholipase A(2) (PLA(2)) activity was found in Trimorphodon biscutatus lambda venom, and moderate levels were detected in Boiga dendrophila and D. p. regalis venoms as well as B. dendrophila and H. n. nasicus salivas. Non-reducing SDS-PAGE revealed 7-20 protein bands (3.5 to over 200 kD, depending on species) for all venoms analyzed, and electrophoretic profiles of venoms were typically quite distinct from saliva profiles. Components from A. stolata, Hydrodynastes gigas, Tantilla nigriceps and T. e. vagrans venoms showed protease activity when run on gelatin zymogram gels. N-terminal protein sequences for three 26 kD venom components of three species (H. gigas, H. torquata, T. biscutatus) and one 3.5 kD component (T. nigriceps) were also obtained, and the 3.5 kD peptide showed apparent sequence homology with human vascular endothelial growth factor; these data represent the first sequences of colubrid venom components. Protease, phosphodiesterase and PLA(2) activities are also common to elapid and viperid snake venoms, but it is apparent that numerous other (as yet undescribed) components make up the majority of colubrid venom proteins. The complex nature of venoms produced by most species surveyed, and the high levels of protease or phospholipase A(2) activity of some venoms, suggest that many colubrids could become an important source of human health concern as encounters with these snakes increase. PMID- 10858510 TI - Paralytic shellfish toxins in the freshwater cyanobacterium Aphanizomenon flos aquae, isolated from Montargil reservoir, Portugal. AB - Montargil reservoir, located in a dry flat area in the centre of Portugal, was filled in 1958 to fulfil agricultural, electric and industrial requirements. In May 1996, an intensive bloom of phytoplankton was detected. The algal community was strongly dominated by cyanobacteria with predominance of Aphanizomenon flos aquae from May to June and Microcystis aeruginosa from July to August. Extracts of samples collected during the bloom period showed high toxicity by mouse bioassay. During the M. aeruginosa predominance period, the toxicity was ascribed to the presence of hepatotoxins, but clear symptoms of paralytic shellfish poison were observed when A. flos-aquae was the dominant species. In order to confirm the production of neurotoxins a strain of A. flos-aquae was isolated and established in culture. In this manuscript, we show the morphological characteristics and confirm paralytic shellfish toxins production by the strain isolated and maintained in culture. Identification of the saxitoxin analogs was achieved using high performance liquid chromatography with postcolumn fluorescence derivatization (HPLC-FLD) and liquid chromatographic mass spectrometry technique (LC-MS). The toxins found in the culture extract were GTX5 (64.5 mol%), neoSTX (23.0 mol%), dcSTX (6.1 mol%), STX (5.4 mol%) and GTX6 (1.1 mol%). This is, to our knowledge, the first report of unambiguous evidence of paralytic shellfish toxins produced by freshwater cyanobacteria in Portugal. The toxin profile is rather different from the previously reported PSP producing A. flos-aquae and demonstrates its diversity in terms of toxin production. PMID- 10858511 TI - Characterisation of the biochemical and biological variations from the venom of the death adder species (Acanthophis antarcticus, A. praelongus and A. pyrrhus). AB - We report on species variation in the venoms of the three species of death adder; the Common death adder (Acanthophis antarcticus), the Northern death adder (Acanthophis praelongus) and the Desert death adder (Acanthophis pyrrhus). The venoms were found to vary in their biochemical (chromatography) and biological (PLA(2) activity, anticoagulant activity and reactivity with commercial death adder antivenom) properties. Each species produced significant differences in the profile and distribution of PLA(2) activity, when whole venom was applied to a cation-exchange Mono-S column. PLA(2) enzymes were purified from each venom and termed acanthoxin B (from A. praelongus), acanthoxin C (from A. pyrrhus) and the previously characterised acanthoxin A (from A. antarcticus). Acanthoxin B and C showed lower enzymatic activities than acanthoxin A (4.0, 13.7 and 23.9 micromol of phospholipid hydrolyzed/min/mg protein, respectively). N-terminal sequencing revealed acanthoxin B to share highest homology with the numerous PLA(2) isozymes (Pa-12C, Pa-1G, Pa-12A) from the King brown snake (Pseudechis australis) and Acanthin I from the Common death adder. Similar to acanthoxin A, acanthoxin C showed highest homology with Acanthin I/II, and pseudexin A-chain from the Red bellied black snake (Pseudechis porphyriacus). Whole venom from A. antarcticus, A. praelongus and A. pyrrhus each showed weak anticoagulant activity (being able to prolong coagulation of the plasma for 107, 220 and 195 s, respectively). By immunodiffusion, each venom produced precipitation bands against commercial death adder antivenom. PMID- 10858512 TI - The envenomation syndrome caused by the Australian Red-bellied Black Snake Pseudechis porphyriacus. AB - The Australian elapids inject venom which is characteristic of each species; and which cause characteristic and specific envenomation syndromes in human victims of snakebite. Because many of the medically significant Australian elapids look similar, when glimpsed in the field by snakebite victims, defining human envenomation syndromes with secure species identification has been a slow process. Correlations between securely identified species and the human envenomation syndromes which they produce are still evolving. The genus Pseudechis is the most widespread in Australia of the dangerous Australian elapid genera; and P. porphyriacus, the Red-bellied Black Snake, was the first terrestrial Australian elapid to be described and illustrated and the first to be the subject of experimental study. We present here five previously unreported cases of human envenomation in which the species diagnosis is secure. From these and with the perspective of a selected literature review, we describe the full envenomation syndrome of this species. Until the development of the Commonwealth Serum Laboratories' Venom Detection Kit in 1979 and the occasional case report of victims of securely identified species, envenomation syndromes for most Australian snake species have remained indeterminate, because of the lack of professional expertise in the identification of the species involved. Symptoms of the P. porphyriacus envenomation syndrome include those of bite-site pain, nausea and vomiting, generalised pruritus, chest pain, prostration and abnormalities of taste and smell. Signs include local necrosis and scarring of tissue at the bite site, gross inflammation of surrounding tissues and, at least in one case, epilepsy. Although envenomation by the Red-bellied Black Snake is not lethal in adults, the correct therapy is Tiger Snake antivenom, administered with judgement, taking into account knowledge of the specific envenomation syndrome of this species and the clinical status of the victim. PMID- 10858513 TI - Toxin production in cyanobacterial mats from ponds on the McMurdo ice shelf, Antarctica. AB - Cyanobacteria are known to produce hepatotoxic substances, the functional and ecological role of these toxins, however, remains largely unclear. Toxic properties of cyanobacteria collected in Antarctica were investigated to determine whether toxin-producing species can also be found under these environmental conditions. Samples were collected from meltwater ponds on the McMurdo Ice Shelf, Antarctica in the summers of 1997 to 1999. These ponds are colonized by benthic algae and cyanobacterial mats. Oscillatoriales, Nodularia sp., and Nostoc sp. constituted the major taxa in freshwater ponds, while Nostoc sp. was missing from brackish and saline ponds. Samples were taken from either floating, submerged or benthic mats, and extracted for in vitro toxicity testing. The presence of toxins was determined by the phosphatase-inhibition assay and by high performance liquid chromatography. The cytotoxic properties of the extracts were investigated in hepatocytes determining 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl-tetrazolium bromide metabolism and trypan blue dye exclusion. The results show that all cyanobacterial extracts display phosphatase-inhibiting activity, of which approximately half had significantly greater than 50% inhibiting activity. The presence of nodularin and microcystin-LR was established by high performance liquid chromatography. Cytotoxic properties, independent of the phosphatase inhibiting activity, were also detected. Toxic strains of cyanobacteria can therefore also be found in Antarctica and this finding may lead to further insight into potential ecological roles of cyanobacterial phosphatase inhibiting toxins. PMID- 10858514 TI - On the kaliotoxin and dendrotoxin binding sites on rat brain synaptosomes. AB - The toxic polypeptides alpha-, beta-, gamma- and delta-dendrotoxin (DTX), known to be potent blockers of voltage-dependent potassium channels of the Kv1 family, were purified from the venom of the green mamba Dendroaspis angusticeps. Their binding behaviour to synaptosomal membranes of rat brain was analysed and compared with that of kaliotoxin (KTX), in a competition assay using [(125)I] KTX. alpha-DTX and delta-DTX were found to compete with radioiodinated-KTX (IC(50) of 8 pM and 0.2 nM respectively), whereas gamma-DTX did not. Several minor components that competed with radioiodinated-KTX binding were identified and characterised chemically and biologically. PMID- 10858515 TI - Isolation and characterization of microcystins from a river nile strain of Oscillatoria tenuis Agardh ex Gomont. AB - The River Nile is the major source of drinking water in Egypt, however, increased eutrophication due to agricultural, municipal and industrial runoff has contributed to the growth of toxin producing cyanobacteria. This study describes the isolation and characterization of microcystins (MCYSTs), cyclic heptapeptide hepatotoxins, from a rare strain of Oscillatoria tenuis, isolated from the River Nile at Sohag province in July 1995. The MCYST concentration of laboratory cultured O. tenuis strain E6 was found to be 0.3 mg/g freeze-dried weight determined by enzyme-linked immunosorbent assay (ELISA). Two microcystins, 1 and 2, were isolated from lyophilized cells using solid phase extraction and reversed phase high performance liquid chromatography (HPLC). Structures were assigned based upon their amino acid analyses, electrospray ionization mass spectrometry (ESIMS, ESIMS-CID-MS), high resolution fast atom bombardment mass spectrometry, and nuclear magnetic resonance data ((1)H and (1)H COSY NMR). Toxin 1 was identified as MCYST-LR, and toxin 2, a new MCYST, as MCYST-LHArg ([L homoarginine(4)]). Previous studies indicate that Oscillatoria agardhii strains produce demethylated MCYSTs (containing D-Asp and/or dehydroalanine). This is the first report of a toxic O. tenuis, strain E6, one which produces a fully methylated MCYST, MCYST-LR and a new L-homoarginine containing MCYST, MCYST LHArg. PMID- 10858516 TI - Leucocyte recruitment induced by type II phospholipases A(2) into the rat pleural cavity. AB - Bothropstoxin-I (BthTX-I) and bothropstoxin-II (BthTX-II) are Lys-49 and Asp-49 phospholipases A(2) (PLA(2)s), respectively, isolated from Bothrops jararacussu venom. Piratoxin-I (PrTX-I) is a Lys-49 PLA(2) isolated from Bothrops pirajai venom. In this study, the ability of BthTX-I, BthTX-II and PrTX-I to recruit leucocytes into the rat pleural cavity and potential mechanisms underlying this effect were investigated. Intrapleural injection of either BthTX-I or PrTX-I (10 100 microg/cavity each) caused a significant leucocyte infiltration at 12 h after injection. The maximal cell migration was observed with the dose of 30 microg/cavity (14.9+/-15.5 and 17.6+/-1. 6x10(6) cells/cavity, respectively). Leucocyte counts consisted mainly of mononuclear cells, but significant amounts of neutrophils and eosinophils were also observed. Intrapleural injection of BthTX-II (10-100 microg/cavity) caused a marked leucocyte infiltration at 6 and 12 h after injection. The maximal response was observed with the dose of 100 microg/cavity (57.3+/-3.4x10(6) cells/cavity, 6 h). The leucocyte counts were mainly composed of neutrophils and mononuclear cells. The treatment of either BthTX-I (30 microg/cavity, 12 h) or BthTX-II (30 microg/cavity, 6 h) with the PLA(2) inhibitor p-bromophenacyl bromide (p-BPB) had no effect on the total and differential leucocyte counts induced by these proteins. The same treatment partially reduced the PrTX-I-induced pleural leucocyte infiltration. In rats depleted of the histamine and 5-hydroxytryptamine (5-HT) stores by chronic treatment with compound 48/80, the total leucocyte counts in response to BthTX-I, BthTX-II and PrTX-I was not significantly affected compared to control animals. In addition, BthTX-I, BthTX-II and PrTX-I (100 microg/ml each) significantly degranulated pleural mast cells in vitro leading to the release of [(14)C]5 hydroxytryptamine ([(14)C]5-HT). p-BPB and heparin (50 IU/ml) significantly reduced the [(14)C]5-HT release induced by these PLA(2)s. Our results demonstrate that BthTX-I, BthTX-II and PrTX-I recruit leucocyte into the pleural cavity of the rat by mechanisms unrelated to enzymatic activity and pleural mast cell degranulation. PMID- 10858517 TI - The effects of lignocaine on actions of the venom from the yellow scorpion "Leiurus quinquestriatus" in vivo and in vitro. AB - Many toxins from scorpion venoms activate sodium channels, thereby enhancing neurotransmitter release. The aim of the present work was to determine if the in vivo and in vitro effects of Leiurus quinquestriatus venom (LQQ) could be ameliorated by lignocaine, a sodium channel blocker. In urethane anaesthetised rabbits, LQQ venom (0.5 mg kg(-1), i.v.) caused initial hypotension and bradycardia followed by hypertension, pulmonary oedema, electrocardiographic changes indicating conduction defects, ischaemia, infarction, and then hypotension and death. Lignocaine (1 mg kg(-1) i.v. bolus initially, followed by i.v. infusion of 50 microg kg(-1) min(-1)) significantly attenuated the majority of the venom-evoked effects and reduced mortality. Addition of LQQ venom (1, 3 and 10 microg ml(-1)) to chick biventer cervicis, guinea pig ileum, and rat vas deferens preparations, increased the height of electrically-induced twitches, elevated resting tension, and caused autorhythmic oscillations. Lignocaine (3 x 10(-4)-1.2 x 10(-3) M) greatly attenuated these venom-evoked actions in the three preparations. Antagonists of appropriate neurotransmitters were also tested to determine the contribution of released transmitters to LQQ effects. Atropine significantly decreased the venom-elicited effects on guinea pig ileum preparations, while prazosin and guanethidine significantly reduced the venom's actions on rat vas deferens. In chick biventer cervicis preparations, tubocurarine and hexamethonium significantly attenuated the venom-induced effects. This study supports the hypothesis that many effects of LQQ venom involve the release of neurotransmitters and may be ameliorated by treatment with lignocaine. PMID- 10858518 TI - Giant hornet (Vespa mandarinia) venomous phospholipases. The purification, characterization and inhibitory properties by biscoclaurine alkaloids. AB - Two species of giant hornet phospholipase B (PLB), alpha and beta, were purified from the venom of Vespa mandarinia. The purification procedure was simplified by two steps of column chromatographies, Sephadex G-100 and SP-Sepharose. The molecular sizes of PLB alpha and beta were 29.5 and 26.0 kDa, respectively. The isoelectric point of alpha and beta enzymes were pH 10.6 and 10.7, respectively. The temperature optimum for egg yolk lecithin was a broad peak at 40-60 degrees C for both enzymes. Amino acid compositions of both enzymes were high contents of aspartic acid, glycine, leucine, lysine and other aliphatic amino acids. Cystine was similar amounts to other species of phospholipases (PLs). The K(m) values of alpha and beta enzymes were 8.29 and 7.53 mg/ml for egg yolk lecithin, respectively. In the catalytic specificity for L-alpha-phosphatidylcholine-beta oleoil-gamma-palmitoil, the K(m) values of alpha enzyme for gamma-palmitoil and beta-oleoil residues were 0.528 and 1.392 mM, respectively. While the K(m) values of beta enzyme for gamma-palmitoil and beta-oleoil residues were 7.91 and 2. 68 mM, respectively. Both alpha and beta enzymes were inhibited strongly by cepharanthine. The lecithin hydrolysis of alpha enzyme was competitively inhibited, but beta enzyme was uncompetitive. Cepharanthine also inhibited noncompetitively PLA(2)s of bovine pancreas, bee venom and Naja mossambica mossambica. PMID- 10858519 TI - The isolation and purification of two peptides from the venom of Buthus martensii Karsch. AB - Two peptides that extensively prolong action potentials (APs) in rat and frog nerves have been isolated and purified from the venom of the scorpion Buthus martensii Karsch (BMK). The peptides were purified using gel filtration, ion exchange, FPLC, and HPLC chromatography. Action potentials recorded in the presence of nanomolar concentrations of the peptides were extensively prolonged without much attenuation in their heights. The N-terminal sequences of both the peptides, BMK 9(3)-1 and BMK 9(3)-2, were determined. The N-terminal sequences of BMK 9(3)-1 and BMK 9(3)-2 were found to be: GRDAYIADSEN-PYF-GANPN and GRDAYIADSEN PYT-ALNP. Sequence similarity comparisons to other alpha-scorpion toxins suggest that the two blanks in each of the sequences are cysteines. The first 20 residues of the two BMK peptides differ by only three amino acid substitutions. The molecular weight (MW) of BMK 9(3)-1 and BMK 9(3)-2 were determined by LC/MS/MS to be 7020 and 7037 Da. Since both of the peptides prolong APs when both K(+) and Ca(++) channels are blocked and show sequence similarity to other alpha neurotoxins, it appears likely that BMK 9(3)-1 and BMK 9(3)-2 act to alter Na channel inactivation to produce their effects. The first 20 residues of BMK 9(3) 2 are identical to those observed for makatoxin I, a toxin isolated from Buthus martensii Karsch venom, that alters nitric oxide transmitter release. Since the two toxins also have very similar molecular weights, BMK 9(3)-2 may be identical to makatoxin I; however, BMK 9(3)-2 acts to alter Na channels to exert its effect, thus the two toxins may differ, or if they are identical, they can exert effects on both neural transmission and AP propagation. PMID- 10858520 TI - Characterization of 9H-(1,3-dichlor-9, 9-dimethylacridin-2-ona-7-yl)-phosphate (DDAO) as substrate of PP-2A in a fluorimetric microplate assay for diarrhetic shellfish toxins (DSP). AB - Specific inhibition of protein-phosphatases by diarrhetic shellfish toxins (DSP) of the okadaic acid group, has led to the development of a fluorescent enzyme inhibition assay for these toxins using protein-phosphatase 2A (PP-2A) and fluorogenic substrates of the enzyme. Two different substrates of PP-2A have been previously used in this microplate assay: 4-methylumbelliferyl phosphate and fluorescein diphosphate (FDP). In this report, we present the results obtained using a new fluorogenic substrate of PP-2A, the compound dimethylacridinone phosphate (DDAO). A linear relationship between PP-2A concentration and DDAO induced fluorescence was observed. Okadaic acid (0.0157-9.43 nM)-dependent inhibition of phosphatase activity showed similar results using FDP and DDAO. Recovery percentages obtained with FDP and DDAO in spiked mussel samples (both raw and canned) were very similar and reproducible. Comparative analysis of DSP contaminated mussel samples by HPLC and FDP/DDAO-PP-2A showed a good correlation among all methods, thus demonstrating that DDAO can be used as a fluorogenic substrate to quantify okadaic acid and related toxins in bivalve molluscs with optimum reliability. PMID- 10858521 TI - Isolation of subunits, alpha, beta and gamma of the complex taipoxin from the venom of Australian taipan snake (Oxyuranus s. scutellatus): characterization of beta taipoxin as a potent mitogen. AB - The venom of Australian taipan snake (Oxyuranus s. scutellatus) is extremely potent due to the presence of taipoxin. The intact complex molecule of taipoxin having molecular weight 45.6 kDa is composed of alpha, beta and gamma subunits. This report describes the high pressure liquid chromatography (HPLC) separation of alpha, beta (beta-1 and beta-2) and gamma subunits from taipan crude venom. The fractions containing the taipoxin subunits were further purified to obtain homogeneous proteins. The toxicity in mice showed the alpha subunit as most toxic, the gamma subunit as moderately toxic and the beta-1 and beta-2 subunits were nontoxic. The proteins beta-1 and beta-2 were found to be mitogenic having neurotrophic activity on PC12 cells in culture similar to nerve growth factor. Immunologically, alpha, beta-1, beta-2 and gamma subunits were found to be different, showing cross reactivity, and beta-1 and beta-2 were found to be identical for biological properties and molecular weight. Further characterization of unexpected mitogenic activity of beta subunits is underway. PMID- 10858522 TI - Neutralisation of the clotting activity of Australian snake venoms by snake plasma. AB - Plasmas from Pseudonaja textilis and Notechis scutatus were tested in vitro for their ability to neutralise the procoagulant activity, in human plasma, of nine elapid venoms. Pseudonaja textilis plasma inhibited the procoagulant activity of all Pseudonaja species and in one taipan (Oxyuranus scutellatus). However there was no inhibitory activity against any from the Notechis species. Plasma from Notechis scutatus exhibited no inhibitory activity against any Notechis species, including self, only weak inhibition against the Pseudonaja species and, again, total inhibition of Oxyuranus scutellatus. Thus, protection of a species from the effects of its own venom does not appear to be universal. PMID- 10858523 TI - Neutralizing potency of horse antibothropic Brazilian antivenom against Bothrops snake venoms from the Amazonian rain forest. AB - Neutralization of lethal toxicity (50% effective dose; ED(50)), hemorrhagic (minimum hemorrhagic dose; MHD) and hemolytic activity (PLA(2)) and levels of antibodies, measured by enzyme-linked immunosorbent assay (ELISA), were investigated to test the potency of horse antibothropic serum (ABS) against Bothrops venoms from the Amazonian rain forest. ABS neutralized the lethal activity with a potency (mg of venom neutralized per 1 ml of antivenom) of 5.5, 3.7, 1.6, 1.3 and 6.5, respectively, for B. jararaca (reference venom for assessing the ABS potency in Brazil), B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms. The volume of antivenom (microl) that neutralized one MHD of B. jararaca, B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms was 5, 7.71, 7.76, 8.3 and 5, respectively. ABS neutralized the PLA(2) activity with a potency of 6.2, 3.2, 1.4, 2.6 and 5 respectively, for B. jararaca, B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms. ELISA reactivity of ABS against the separate venoms was found to be quite variable. The reactivity against B. jararaca venom was higher than against other Bothrops venoms. In conclusion, the assays described here suggest that Brazilian Bothrops polyspecific antivenom is not very efficient in neutralizing the effects of venom from some Amazonian Bothrops species. PMID- 10858524 TI - Cantharidin poisoning due to "Blister beetle" ingestion. AB - Cantharidin, the active ingredient of "Spanish Fly", is contained in a number of insects collectively called blister beetles and is a well known toxin and vesicant. We report on a case of ingestion of Mylabris dicincta ("Blister beetle") in Zimbabwe by a 4 year old girl. The ingested beetles were probably mistaken for the edible Eulepida mashona. She presented with many of the classic signs and symptoms of cantharidin poisoning including haematuria and abdominal pains. This was recognised only after consultation with the drug information centre. She was managed conservatively, recovered and was discharged after 9 days. A overview of the clinical effects of cantharidin toxicity and its treatment is presented. PMID- 10858525 TI - Saxitoxin and neosaxitoxin as toxic principles of Alexandrium andersoni (Dinophyceae) from the Gulf of Naples, Italy. AB - A clonal culture of Alexandrium andersoni, obtained from germination of a resting cyst, collected from the Gulf of Naples, was found positive for PSP toxicity by mouse bioassay. The toxicity profile of this dinoflagellate consists mainly of toxins belonging to the saxitoxin class, in particular of Saxitoxin (STX) and Neosaxitoxin (NEO), as determined by a wide MS and (1)H NMR analysis. This represents the first report of the presence of A. andersoni in the Mediterranean Sea, as well as of its toxicity. PMID- 10858526 TI - Plant glycine-rich proteins: a family or just proteins with a common motif? AB - Twelve years ago a set of glycine-rich proteins (GRP) of plants were characterized and since then a wealth of new GRPs have been identified. The highly specific but diverse expression pattern of grp genes, taken together with the distinct sub-cellular localisation of some GRP groups, clearly indicate that these proteins are implicated in several independent physiological processes. Notwithstanding the absence of a clear definition of the role of GRPs in plant cells, studies conducted with these proteins have provided new and interesting insights on the molecular and cell biology of plants. Complex regulated promoters and distinct mechanisms of gene expression regulation have been demonstrated. New protein targeting pathways, as well as the exportation of GRPs from different cell types have been discovered. These data show that GRPs can be useful as markers and/or models to understand distinct aspects of plant biology. In this review, the structural and functional features of this family of plant proteins will be summarised. Special emphasis will be given to the gene expression regulation of GRPs isolated from different plant species, as it can help to unravel their possible biological functions. PMID- 10858528 TI - Cell type specificity and mechanism of control of a gene may be reverted in different strains of Dictyostelium discoideum. AB - Twelve genes which are expressed exclusively in pre-spore cells of Dictyostelium strain AX3 are expressed exclusively in pre-stalk cells of strain AX2. One gene has the opposite behavior: it is expressed in pre-stalk cells in AX3 and in pre spore cells in AX2. The change in cell type specificity involves a change in the mechanism of control of gene expression. When they are expressed in pre-stalk cells, genes are controlled at the level of transcription, whilst in pre-spore cells, they are controlled at the level of mRNA stability. Genes expressed in pre stalk cells in strain AX2, fused with an AX2 pre-spore specific promoter, become regulated at the level of mRNA stability. These findings indicate that at least a group of pre-stalk mRNAs possess the cis-destabilizing element typical of pre spore mRNAs, though they are not destabilized in disaggregated cells. This is due to the fact that ribosomal protein S6, phosphorylation of which is responsible for controlling the stability of pre-spore mRNAs, is not dephosphorylated in disaggregated pre-stalk cells. These cells lack an S6 phosphatase activity which has been purified from disaggregated pre-spore cells. PMID- 10858531 TI - Identification of domains within the epsilon-subunit of the translation initiation factor eIF2B that are necessary for guanine nucleotide exchange activity and eIF2B holoprotein formation. AB - Eukaryotic translation initiation factor, eIF2B, is a guanine nucleotide exchange factor (GEF) composed of five dissimilar subunits. eIF2B is important for regenerating GTP-bound eIF2 during the initiation process. This event is obligatory for eIF2 to bind initiator methionyl-tRNA, forming the ternary initiation complex. In the current investigation, deletion mutants of the catalytic subunit, eIF2B epsilon, were constructed to identify regions that are necessary for eIF2B catalytic activity and formation of the holoprotein. We used the baculovirus expression system to coexpress wild-type and truncated forms of the epsilon-subunit of mammalian eIF2B (eIF2B epsilon) with the other four subunits (alpha, beta, gamma, delta) of the protein in Sf9 cells. Removal of either the N- or the C-terminal conserved domains of eIF2B epsilon resulted in a significant loss of GEF activity and reduced or abolished interaction with the alpha-, gamma- and delta-subunits of eIF2B. Removal of the C-terminal 552 amino acids of eIF2B epsilon markedly reduced its interaction with the beta-subunit of eIF2 whereas loss of the N-terminal 431 amino acids did not. The results suggest that intact eIF2B epsilon is required for full catalytic activity and formation of the eIF2B holoprotein. In contrast, the C-terminal domain of eIF2B epsilon is sufficient alone for binding the beta-subunit of its substrate, eIF2, in vitro. PMID- 10858535 TI - K(+) and Mg(2+) ions promote the self-splicing of the td intron RNA inhibited by spectinomycin. AB - The effects of Mg(2+) and K(+) ions on the self-splicing inhibition of the td (thymidylate synthase gene) intron RNA by spectinomycin were investigated. The maximum splicing activity occurred at 20 mM KCl. The K(m) and V(max) values for GTP in the presence of 5 mM Mg(2+) are 2.25 microM and 0.55 min(-1), whereas those for GTP both in the presence of 5 mM Mg(2+) and 5 mM K(+) are 1.23 microM and 0. 46 min(-1), respectively. Spectinomycin at 10 mM concentration inhibited the splicing by about 10%, but at 20 mM concentration, the splicing rate was inhibited by about 63%. The splicing inhibition by the low concentration of spectinomycin was overcome markedly as the concentration of Mg(2+) ion was raised. At 30 mM spectinomycin, however, the splicing inhibition was not significantly affected by increasing the concentration of Mg(2+). A similar activation of the splicing rate was observed as the concentration of K(+) ion was increased. The concentration of K(+) ion required for the normal recovery of the splicing was much higher than that of Mg(2+) ion. Unlike Mg(2+) ion, 30 mM K(+) ion effectively alleviated the splicing inhibition by spectinomycin at its high concentration. The results indicate that K(+) and Mg(2+) ions may show mechanistically different interactions with spectinomycin in the self-splicing reaction of the td intron RNA. PMID- 10858536 TI - Gene, stimulus and cell-type specific regulation of activator protein-1 in mesangial cells by lipopolysaccharide and cytokines. AB - Activator protein-1 (AP-1) plays an important role in the regulation of gene expression in mesangial cells (MC) during the pathogenesis of glomerular inflammatory disease. The precise regulation of the AP-1 family by agents that are known to activate MC is, however, poorly understood. The action of platelet derived growth factor (PDGF) and, for the first time, lipopolysaccharide (LPS), interleukin-6 (IL-6), interferon-gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha) on AP-1 gene expression in MC was therefore studied. Whilst the expression of JunD was not affected by any of the mediators, the mRNA levels of c fos and JunB were induced by LPS, IL-6, IFN-gamma, PDGF and TNF-alpha, and that of c-jun by LPS, IFN-gamma, PDGF and TNF-alpha. Electrophoretic mobility shift assays showed a time-dependent increase in AP-1 DNA binding activity with JunB representing the major mediator-inducible member involved in DNA-protein interactions. However, stimulus-specific changes in the kinetics and magnitude of AP-1 mRNA expression and DNA binding activity were identified and, additionally, the results showed the potential existence of cell-type-specific mechanisms in the regulation of the AP-1 family. These studies provide novel insights into the mediator-specific modulation of AP-1-regulated gene expression and the activation of MC in renal diseases. PMID- 10858537 TI - The hnRNP 2H9 gene, which is involved in the splicing reaction, is a multiply spliced gene. AB - The hnRNP 2H9 gene products are involved in the splicing process and participate in early heat shock-induced splicing arrest. By combining low/high stringency hybridisation, database search, Northern and Western blotting it is shown that the gene is alternatively spliced into at least six transcripts: hnRNPs 2H9, 2H9A, 2H9B, 2H9C, 2H9D and 2H9E predicting proteins containing 346, 331, 297, 215, 145 and 139 amino acids, respectively. The hnRNP 2H9A cDNA sequence was used to obtain a genomic BAC clone and the structure of the hnRNP 2H9 gene was revealed by sequencing two subclones together spanning about 6.7 kb. The six transcripts are processed from at least 10, 10, 8, 7, 5 and 4 exons, respectively, with all intron/exon junctions obeying the 'GT-AG' rule. The hnRNP 2H9 and 2H9A proteins contain two RNA recognition motifs of the quasi-RRM type found in the two C-terminal qRRMs of the hnRNPs H, H' and F proteins. The hnRNP 2H9B protein has a partially deleted N-terminal qRRM, which is completely deleted in hnRNP 2H9C. hnRNPs 2H9D and 2H9E contain only one slightly modified C-terminal qRRM. Furthermore, the six proteins vary in their auxiliary domains outside the qRRMs. Western blotting indicates that the alternatively spliced transcripts give rise to different sets and levels of proteins expressed among various human cells and tissues. Due to their great structural variations the different proteins may thus possess different functions in the splicing reaction. PMID- 10858540 TI - Early steps in termination of the immortalization state in Burkitt lymphoma: induction of genes involved in signal transduction, transcription, and trafficking by the ganglioside IV(3)NeuAc-nLcOse(4)Cer. AB - Stimulation by the ganglioside IV(3)NeuAc-nLcOse(4)Cer leads to growth arrest in the Burkitt lymphoma cell line Raji. In order to analyze the primary response of Raji cells to that stimulus, a cDNA array screen and a suppression subtractive hybridization-PCR approach were performed. Twenty-four genes with assigned functions were confirmed to be induced by the ganglioside in reverse Northern blot experiments covering e.g. protein kinase B, phospholipase C, the MAP-kinase ERK3, the transcription factors YY1, DR1 and NSEP, the membrane traffic protein TAP, and the nuclear export protein CRM1. Most of the genes identified are involved in signal transduction, transcription, and cell trafficking. For selected genes, the induction of expression was quantified by semiquantitative RT PCR. PMID- 10858542 TI - Pyrimidine motif triplexes containing polypurine RNA or DNA with oligo 2'-O methyl or DNA triplex forming oligonucleotides. AB - Triplex forming oligonucleotides (TFOs) are potentially useful in targeting RNA for antisense therapeutic applications. To determine the feasibility of targeting polypurine RNA with nuclease-resistant oligonucleotides, TFOs containing 2'-deoxy or 2'-O-methyl (2'-OMe) backbones, designed to form pyrimidine motif triplexes with RNA, were synthesized. TFOs were made which can form trimolecular triplexes, or bimolecular, 'clamp' triplexes with polypurine RNA and DNA. It was found that the relative stabilities of the triplexes formed followed the order: M.DM(clamp)>>>D.DD approximately M.DD>M. RM>D.DM>M.RD approximately M.DM, where M is a 2'-OMe, D is a DNA and R is an RNA backbone. The third strand is listed first, separated by a dot from the purine strand of the Watson-Crick duplex, followed by the pyrimidine strand of the duplex. The results described here provide insight into the feasibility of using TFOs containing a 2'-OMe backbone as antisense agents. PMID- 10858544 TI - Myc and YY1 mediate activation of the Surf-1 promoter in response to serum growth factors. AB - The human Surf-1 and Surf-2 genes are divergently transcribed and share a single bi-directional promoter. The addition of serum growth factors to serum-starved cells activates transcription in the Surf-1 direction, but has no effect on transcription in the Surf-2 direction. Mutations that block the binding of YY1 to a site immediately downstream of the major Surf-1 transcription start point abolish this response to serum factors. Here we show that over-expression of mitogen-activated protein (MAP) kinase phosphatase MKP-1, an inhibitor of the MAP kinase cascade, also blocks the response the Surf-1 promoter to serum factors. YY1 has previously been shown to interact with several transcription factors including Myc. We show that although the Surf-1/Surf-2 promoter does not contain Myc binding sites (E-boxes), Myc over-expression, or the activation of a Myc oestrogen receptor fusion protein, activates transcription in the Surf-1 direction and that this response to Myc requires a functional YY1 binding site. Our data suggest that the MAP kinase cascade is required for the stimulation of Surf-1 promoter activity and that the Myc-YY1 interaction mediates this response. PMID- 10858549 TI - Identification of a novel FGF, FGF-21, preferentially expressed in the liver. AB - We isolated cDNA encoding a novel FGF (210 amino acids) from mouse embryos. As this is the 21st documented FGF, we tentatively term it FGF-21. FGF-21 has a hydrophobic amino terminus ( approximately 30 amino acids), which is a typical signal sequence, and appears to be a secreted protein. The expression of FGF-21 mRNA in mouse adult tissues was examined by Northern blotting analysis. FGF-21 mRNA was most abundantly expressed in the liver, and also expressed in the thymus at lower levels. We also isolated human cDNA encoding FGF-21 (209 amino acids). Human FGF-21 is highly identical ( approximately 75% amino acid identity) to mouse FGF-21. Among human FGF family members, FGF-21 is most similar ( approximately 35% amino acid identity) to FGF-19. PMID- 10858550 TI - Cloning, expression and functional characterization of rat napsin. AB - A full-length cDNA clone coding for rat napsin was identified by homology search of the ZooSeq rat EST database (Incyte). Northern blot analysis revealed high expression of napsin mRNA transcripts in kidney, lung and spleen. Western blot analysis showed that rat napsin is expressed in kidney as a 50-kDa, highly glycosylated, monomeric protein. Lysates prepared from human embryonic kidney cells (HEK293) transfected with rat napsin showed increased enzymatic activity which was inhibited by pepstatin. PMID- 10858556 TI - The human SOX18 gene: cDNA cloning and high resolution mapping. AB - SOX genes comprise a family of genes that are related to the mammalian sex determining gene SRY and these genes play key roles during animal development. We report here cloning and characterisation of the human SOX18 gene. SOX18 gene is expressed in foetal brain as well as in a wide range of foetal and adult tissues indicating its function is not restricted to early development. Mapping analysis has revealed that SOX18 gene is located on human chromosome 20q13.3, 27.29 cR distal from the marker D20S173. PMID- 10858562 TI - Extraordinarily high density of unrelated genes showing overlapping and intraintronic transcription units. AB - The cloning of pyrroline 5-carboxylate reductase from Drosophila melanogaster was accomplished by cDNA complementation of an Escherichia coli proline auxotroph. The corresponding P5cr gene is tightly clustered with three other expressed coding regions. A bidirectional promoter, an overlapping 3'UTR and an intraintronic sequence may all be found in only 4.3 kb, making this the most densely clustered region of unrelated genes in any eukaryote. PMID- 10858564 TI - cDNA cloning of cytoplasmic ribosomal protein S7 of winter rye (Secale cereale) and its expression in low-temperature-treated leaves. AB - A cDNA clone (ScRPS7) encoding the cytoplasmic ribosomal protein S7 was isolated from a rye leaf cDNA library and sequenced. The deduced protein of 192 amino acids with M(r) 22189 shows identity of 52% or 47%, respectively, relative to S7 proteins of human or yeast. A RPS7 mRNA accumulation is higher in the meristematic zone at the leaf base than in the non-meristematic middle and upper section of leaves. Short periods of cold stress sharply reduce the mRNA level while leaves of cold hardened plants contain normal levels of ScRPS7 transcripts. PMID- 10858565 TI - Identification of a new gene (rat TM6P1) encoding a fasting-inducible, integral membrane protein with six transmembrane domains. AB - We describe the isolation of a new gene that encodes a membrane-integrated protein with six transmembrane domains, termed TM6P1. A 403-bp expressed sequence tag was isolated from fasted rat liver subtracted cDNA library, and its full length cDNA is 1482 bp long. It contains an open reading frame of 816 bp and is predicted to encode a 271-amino acid protein with a deduced mass of 29520 Da. A sequence homology search failed to show significant correspondence to any known protein in the databank. TM6P1 has six highly hydrophobic domains that are predicted to be transmembrane helices. Consistent with this prediction, the TM6P1 EGFP fusion protein was shown to localize to the plasma membrane. TM6P1 mRNA is widely expressed in rat tissues, with placenta and liver being the most abundant sites. Fasting increased TM6P1 mRNA nearly two-fold in liver. Taken together, our data suggest that TM6P1 is a unique new membrane integral protein that might have a function important during fasting-induced catabolism. PMID- 10858567 TI - Mouse type-2 retinaldehyde dehydrogenase (RALDH2): genomic organization, tissue dependent expression, chromosome assignment and comparison to other types. AB - Retinaldehyde dehydrogenase (RALDH) isozymes catalyze the formation of an essential developmental modulator, retinoic acid. We determined the structural organization of mouse type-2 Raldh2 by isolation of overlapping genomic DNA clones from a phage library. The gene consists of 14 exons spanning more than 70 kb of genomic DNA. It was localized to mouse chromosome 6. Northern blot analysis revealed testis-specific expression. The RALDH genes belong to the aldehyde dehydrogenase (ALDH) multi-gene family. Three types of RALDH genes (e.g. human ALDH1/mouse Ahd2/rat RalDH(I), human ALDH11/mouse Raldh2/rat RalDH(II) and human ALDH6) are highly conserved during evolution, sharing about 70% identity at the amino acid level between any two gene types and 90% identity between any two mammalian genes of the same type. Different RALDH types show specific tissue and developmental expression patterns, suggesting (i) a regulatory mechanism of retinoic acid synthesis via different promoters of RALDH genes, and (ii) distinctive biological roles of different isozymes in embryogenesis and organogenesis. PMID- 10858568 TI - Prolonged reduction of salivary immunoglobulin A (sIgA) after a major academic exam. AB - OBJECTIVE: In a previous study we observed a continuous reduction of salivary IgA concentration ([sIgA]) during a period of academic stress. This reduction of sIgA concentration exceeded the stress period by at least 1 week. The present study aimed to replicate and extend our previous finding. In particular, we wanted to examine the time of recovery of [sIgA] alterations associated with academic stress. METHOD: Twenty-seven participants in a major medical exam and 27 controls not participating in any exam during the study provided daily saliva samples (immediately after awakening), from the 6th day prior to their last exam until the 14th day afterwards, for analysis of salivary IgA. Data were averaged for the last weeks of exams and the first and second week after exams, respectively. RESULTS: A prolonged reduction of sIgA in exam students as compared to controls was observed. Fourteen days post-stress sIgA concentrations of exam students were still significantly lower than control levels (P=0.004). No recovery was observable. At the same time exam students and controls did not differ in terms of self-reported stress and recovery. CONCLUSIONS: Psychological and immunological stress effects may be dissociated, the latter considerably exceeding the stress period. A closer look at the temporal dynamics of stress induced immune alterations might increase our understanding of psychoimmuno relationships. PMID- 10858569 TI - Trait anxiety and prior exposure to non-stressful stimuli: effects on psychophysiological arousal and anxiety. AB - Arousal and anxiety responses to stressful stimulation are products of multiple factors that may include the physiological, behavioral, cognitive, affective, trait, and state components of our six-system model, as well as mediational and non-mediational perspectives. Within the context of this model, the present experiment examined the effects of prior exposure to neutral stimuli on arousal and anxiety responses to stress. High and low trait-anxious participants were randomly assigned to one of three prior exposure conditions in which they were exposed to stressful stimuli following exposure to: (a) neutral, non-stressful stimulation in the same modality (the Intramodality Prior Exposure or IPE experimental condition); (b) neutral stimuli in a different modality (the Cross modality Prior Exposure or CPE control condition); or (c) a rest period (the Stress Only or SO control condition). Results showed that low trait anxious subjects had consistently larger arousal and anxiety responses to stress than did highs and that prior exposure to certain same-modality neutral stimuli reduced responses to subsequent stressors. In addition, arousal was greater in response to cognitive than to affective and in response to non-mediational than mediational conditions. Results are discussed in terms of the inverted-U arousal function and of the six-system model and its implications for understanding anxiety and arousal. PMID- 10858570 TI - Effects of inter-item lag on recognition memory in seizure patients preceding temporal lobe resection: evidence from event-related potentials. AB - The electrophysiological correlates of word recognition are well characterized. Repeated 'old' words evoke a more positive-going waveform starting at approximately 300 ms compared with first-presented, 'new' words. The old/new effect is thought to be generated, in part, by structures within the medial temporal lobe. In the present study, event-related potentials were recorded during a continuous verbal recognition memory task in unoperated patients with either left (L) or right (R) unilateral temporal lobe epilepsy (Epil) and neurologically intact controls. To manipulate the difficulty of the memory task, the lag between the initial and subsequent presentation of the repeated words was varied from one, four to 16 items. In the controls, ERPs to old words were more positive going than new words from approximately 350-650 ms. The old/new effect diminished as the inter-item lag increased. Patient old/new effects showed a later onset (450 ms) and resolution (750 ms) compared with the controls. Furthermore, the late component of the old/new effect was significantly reduced in the L Epil. Although patient behavioral performance did not differ significantly from that of the controls, neuropsychological testing revealed impaired verbal memory function in the L Epil patients. It is concluded that the reduced old/new effect in the L Epil patients provides evidence that medial temporal lobe structures contribute to the scalp-recorded old/new effect. PMID- 10858571 TI - Adrenocorticotropin responses to interpersonal stress: effects of overt anger expression style and defensiveness. AB - This study evaluated the influence of overt anger expression style and defensiveness on the hypothalamic-pituitary-adrenocortical (HPA) responses to acute psychological stress. These personality traits are thought to modulate the stress cardiovascular response and influence disease risk, however, little is known about their influence on HPA responses. Forty-six young, healthy male volunteers worked on counterbalanced extended public-speaking and mental arithmetic. The sample was dichotomitized into groups low vs. high in anger-out, using Spielberger's Anger-Expression Inventory, and in defensiveness, using the Marlowe-Crown Social Desirability Scale. Serum cortisol and adrenocorticotropic hormone (ACTH) concentrations were measured before and after performing each task. Heart rate (HR) and blood pressures (BP) were obtained continuously in 2 min intervals before, during and after the tasks. Public speaking produced greater adrenocortical and cardiovascular stress responses than mental arithmetic, and the greatest increases in ACTH occurred in subjects high in anger out and defensiveness. These preliminary findings provide evidence that a mismatch between traits of preferred anger expression style and defensive style produces pronounced adrenocorticotropic responses during socially salient stress. PMID- 10858572 TI - Cardiovascular reactivity during public speaking as a function of personality variables. AB - An experiment was conducted to assess the effects of a real-life stressor (public speaking) upon cardiovascular reactivity (CVR). Changes in blood pressure and heart rate from baseline to task were measured in a sample of 86 men and women. The purpose was to examine the effects of individual differences (Type A personality, hostility and gender) on CVR. Participants gave a 6-min oral presentation during which they were evaluated by their professor and with classmates as the audience. Results indicated that all participants had marked CVR during public speaking. There were differences in reactivity patterns between men and women, but personality did not play a role except for high hostile men. It is suggested that intense stressors may result in high levels of CVR independent of personality variables that moderate reactivity at lower levels of stress. PMID- 10858573 TI - The time course of emotional and attentional modulation of the startle eyeblink reflex during imagery. AB - Two studies were conducted to examine the time course of attentional and emotional processing using the startle eyeblink reflex. Forty-eight participants listened to a series of 1500-ms tones that occurred every 6 s. The tones signaled participants to generate emotional images that were positive or negative in valence and high or low in arousal. Auditory startle probes occurred 120 ms, 1400 ms, or 4000 ms after tone onset. Startle inhibition was seen 120 ms after tone onset and startle facilitation was found at 1400 ms, compared to startles elicited 4000 ms after tone onset. Startle inhibition was greater at 120 ms when the tone signaled imagery, indicating an attentional effect. A second experiment found that this effect was not caused by the comparative rarity of the tones signaling imagery. Startle magnitude was also smaller at 1400 ms when the tone signaled imagery compared to no-imagery tones. The type of imagery did not modulate the startle response 120 ms after tone onset, but negative valence imagery enhanced startle magnitude at 1400 ms and 4000 ms after tone onset, and high arousal also enhanced startle magnitude at 4000 ms. Thus, attention and emotion followed different time courses in affecting the startle reflex response during imagery. PMID- 10858574 TI - The relationship between subjective sleep estimation and objective sleep variables in depressed patients. AB - INTRODUCTION: To our knowledge there is no evidence in the literature about the relationship between subjective sleep estimation and objective sleep variables in depression. It is not known whether the subjective estimation of sleep quality and sleep duration is directly related to any objective sleep variable in depressed patients. METHODS: Thirty patients with major depression and 10 healthy subjects have been investigated in our sleep laboratory during 1 or 2 consecutive nights after 1 night for adaptation. Every subject, after final awakening in the laboratory, answered questions concerning the subjective feelings about sleep duration, number of awakenings and sleep depth. We compared the sleep estimation in both groups and calculated the correlation between objective and subjective sleep variables in depressed patients. RESULTS: The degree of a wrong sleep estimation in depressed patients is larger than in healthy subjects. Slow wave sleep (SWS) in depressed patients correlates positively with the subjective estimation of sleep duration. Eye movement density in REM sleep correlates with the subjective estimation of the number of awakenings. CONCLUSION: SWS in depression has a positive influence on the subjective feeling of sleep duration while phasic REM sleep activity has a negative influence. PMID- 10858575 TI - Stress, social support and cardiovascular activity over the working day. AB - The influence of stress on ambulatory blood pressure monitored over the working day, and the potential buffering effect of social support, was assessed in 104 school teachers (37 men and 67 women). Blood pressure and heart rate were measured every 20 min and energy expenditure was assessed using accelerometers. Participants rated the degree of stress they were experiencing at the time of each measurement on a seven-point scale. Episodes of both high and low stress during the working day were reported by 62 participants. They were divided by median split into high and low social support groups on the Interpersonal Support Evaluation List. After controlling for body mass and concomitant energy expenditure, high stress was associated with increased systolic blood pressure, diastolic blood pressure and heart rate. However, the impact of stress was buffered by social support, with no significant increase in blood pressure or heart rate with stress in the high support group. The accelerometers were also shown effectively to discriminate between blood pressure readings taken in the seated and standing positions in terms of energy expenditure. The results corroborate laboratory studies, in showing that social support buffers the impact of episodic stress on cardiovascular activity under naturalistic conditions during the working day. PMID- 10858576 TI - Event-related brain potentials to attended and ignored olfactory and trigeminal stimuli. AB - Event-related brain potentials (ERPs) were recorded from 26 young adults, with equal numbers of male and female subjects, using attended and ignored, olfactory and trigeminal stimuli. The amplitudes and latencies of the N1, P2, and P3 components were recorded using a single-stimulus paradigm, with an inter-stimulus interval of 60 s, employing amyl acetate as the olfactory stimulus and ammonia as the trigeminal stimulus. Subjects estimated stimulus intensity in the attend condition or continued with a visual tracking task in the ignore condition. Results indicate that olfactory information is processed 30-70 ms faster than trigeminal information for the N1 and P2 potential and 100 ms faster for the P3 ERP component. N1/P2 interpeak amplitude was greater for the trigeminal than the olfactory stimuli, and greater in the attended than ignored condition. P3 amplitude was greater in the attend than ignore condition for olfactory information processing and equivalent for trigeminal information processing. These findings suggest that neuronal resource allocation is greatest for attended stimuli and that a painful stimulus demands neuronal resources even when ignored. PMID- 10858577 TI - Phototrophs in high iron microbial mats: microstructure of mats in iron depositing hot springs. AB - Chocolate Pots Hot Springs in Yellowstone National Park are high in ferrous iron, silica and bicarbonate. The springs are contributing to the active development of an iron formation. The microstructure of photosynthetic microbial mats in these springs was studied with conventional optical microscopy, confocal laser scanning microscopy and transmission electron microscopy. The dominant mats at the highest temperatures (48-54 degrees C) were composed of Synechococcus and Chloroflexus or Pseudanabaena and Mastigocladus. At lower temperatures (36-45 degrees C), a narrow Oscillatoria dominated olive green cyanobacterial mats covering most of the iron deposit. Vertically oriented cyanobacterial filaments were abundant in the top 0.5 mm of the mats. Mineral deposits accumulated beneath this surface layer. The filamentous microstructure and gliding motility may contribute to binding the iron minerals. These activities and heavy mineral encrustation of cyanobacteria may contribute to the growth of the iron deposit. Chocolate Pots Hot Springs provide a model for studying the potential role of photosynthetic prokaryotes in the origin of Precambrian iron formations. PMID- 10858578 TI - Carbofuran degradation mediated by three related plasmid systems. AB - Two carbofuran-metabolizing Sphingomonas strains, TA and CD, were isolated from soils with differing histories of exposure to carbofuran. These strains were compared with a previously described strain, Sphingomonas sp. CFO6, with regard to growth rate, formation of metabolites, and plasmid content and structure. Extensive regions of similarity were observed between the three different plasmid systems as evidenced by cross hybridization. In addition, all three systems harbor IS1412, an insertion sequence (IS) element involved in heat-induced loss of carbofuran phenotype in CFO6, and heat-induced carbofuran deficient mutants of all three strains correlated with loss of IS1412. A carbofuran deficient mutant of TA generated by induction of IS elements was complemented by reintroduction of the wild-type plasmid, confirming the presence of genes required for carbofuran metabolism on this plasmid. Carbofuran metabolism in these three strains is clearly linked via plasmids of different numbers and sizes that share extensive common regions, and carbofuran-degrading genes may be associated with active IS elements. PMID- 10858579 TI - Influence of growth rate and starvation on fluorescent in situ hybridization of Rhodopseudomonas palustris. AB - In situ hybridization with a fluorescently labeled 16S rRNA-targeted probe was examined using Rhodopseudomonas palustris as a model organism, which had been grown at different rates and under different conditions of growth and starvation. The specific growth rate did not affect the percentage of hybridized cells in aerobically grown R. palustris cultures. However, significant changes in the percentage of hybridized cells occurred during extended periods of starvation. These changes were observed both in batch cultures grown and starved aerobically in the dark, and in cultures grown phototrophically and starved anaerobically in the dark. Aerobic growth in batch culture and subsequent starvation resulted in a complete lack of detectable hybridization after 20 days of starvation. In contrast, even after 30 days of starvation, 50% of all cells were still detectable in cultures grown aerobically at growth rates <0.06 h(-1) and then starved aerobically in the dark. The same was true for phototrophically grown cells that were starved anaerobically in the light. During starvation there was a clear, though non-linear, positive correlation between the percentage of hybridized cells and the RNA content. In contrast, no direct correlation was observed between the number of hybridized cells in a culture and the viability of this culture. Thus, in habitats with growing, non-growing, and starving bacteria, data on quantitative detection of populations based on 16S rRNA-targeted probing should be used with extreme caution as the detectability of the individual cells is strongly influenced by their physiological history and current physiological state. PMID- 10858580 TI - Behavior of sulfate reducing bacteria under oligotrophic conditions and oxygen stress in particle-free systems related to drinking water. AB - The response of sulfate reducing bacteria (SRB) to oxygen stress under oligotrophic conditions in particle-free systems was studied in (i) sterile Berlin drinking water; (ii) mineral medium; and (iii) in coculture experiments with aerobic bacteria. Using a polyphasic approach including anaerobic cultivation, fluorescent in situ hybridization (FISH) and digital image analysis, the behavior of the strains zt3l and zt10e, isolated from Berlin groundwater and affiliated to the family Desulfovibrionaceae, was compared to the type strains Desulfomicrobium baculatum and Desulfovibrio desulfuricans. Anaerobic deep agar dilution series were performed for the determination of cell culturability. FISH and subsequent digital image analysis of probe-conferred fluorescence intensities were used for the assessment of metabolic activity. For the in situ identification of both isolates in coculture tests, two strain-specific oligonucleotides were developed and evaluated. The total cell counts of stressed SRB in drinking water decreased during the course of the assay dependent on the strain. Both environmental isolates could be cultured for a longer period than cells of D. baculatum and D. desulfuricans, respectively. The FISH intensities showed a strain-specific behavior. When exposed to simultaneous oxygen stress and carbon limitation in mineral medium, total cell counts of all four strains remained constant throughout a period of 72 days. The rate of culturability differed between the investigated strains. The decrease of metabolic activity as assessed by FISH was a strain-specific property. Exposure of SRB to oxygen stress and carbon starvation in coculture experiments with Aquabacterium commune resulted in strain dependent prolonged culturability and a delayed decrease of the metabolic activity compared to pure culture tests for all strains tested. Total cell counts of SRB were constant throughout the whole experiment. PMID- 10858581 TI - The genetic diversity of predominant Escherichia coli strains isolated from cattle fed various amounts of hay and grain(1). AB - When the 16S rDNA of predominant Escherichia coli strains from cattle was digested with HhaI and HaeIII, the strains could be sub-divided into four operational taxonomic units. When genomic DNA was digested with XbaI, strains could be grouped into 24 pulsed-field gel electrophoresis genotypes (>95% Dice similarity) and five clades (>20% Dice similarity). Diet (hay versus grain) and gastrointestinal compartment (rumen versus colon) did not have a large impact on diversity. However, both analyses indicated that the cows (n=2) had different E. coli populations. When all 22 colonic strains were inoculated into a maltose limited chemostat, only a single genotype persisted. Based on these results, the genetic diversity of E. coli in the cattle is very great and this bacterium can occupy different niches. PMID- 10858582 TI - Hypersaline waters in salterns - natural ecological niches for halophilic black yeasts. AB - Hypersaline waters in salterns have so far been considered to be populated only with halophilic algae and bacteria and completely lacking halophilic fungi. In this paper we present population dynamics of polymorphic black yeasts, isolated from hypersaline waters (3-30% NaCl) of a saltern, in relation to different physicochemical parameters. Hortaea werneckii, Phaeotheca triangularis, Trimmatostroma salinum, Aureobasidium pullulans and Cladosporium spp. were detected with the highest frequency just before the peak of halite (NaCl) concentration. Since H. werneckii, P. triangularis and T. salinum are not known outside saline environments, these results suggest that hypersaline water is their natural ecological niche. PMID- 10858583 TI - Use of the T-RFLP technique to assess spatial and temporal changes in the bacterial community structure within an agricultural soil planted with transgenic and non-transgenic potato plants. AB - The aim of this study was to examine whether the terminal restriction fragment length polymorphism (T-RFLP) analysis represents an appropriate technique for monitoring highly diverse soil bacterial communities, i.e. to assess spatial and/or temporal effects on bacterial community structure. The T-RFLP method, a recently described fingerprinting technique, is based on terminal restriction fragment length polymorphisms between distinct small-subunit rRNA gene sequence types. This technique permits an automated quantification of the fluorescence signal intensities of the individual terminal restriction fragments (T-RFs) in a given community fingerprint pattern. The indigenous bacterial communities of three soil plots located within an agricultural field of 110 m(2) were compared. The first site was planted with non-transgenic potato plants, while the other two were planted with transgenic GUS and Barnase/Barstar potato plants, respectively. Once prior to planting and three times after planting, seven parallel samples were taken from each of the three soil plots. The T-RFLP analysis resulted in very complex but highly reproducible community fingerprint patterns. The percentage abundance values of defined T-RFs were calculated for the seven parallel samples of the respective soil plot. A multivariate analysis of variance was used to test T-RFLP data sets for significant differences. The statistical treatments clearly revealed spatial and temporal effects, as well as spacextime interaction effects, on the structural composition of the bacterial communities. T-RFs which showed the highest correlations to the discriminant factors were not those T-RFs which showed the largest single variations between the seven-sample means of individual plots. In summary, the T-RFLP technique, although a polymerase chain reaction-based method, proved to be a suitable technique for monitoring highly diverse soil microbial communities for changes over space and/or time. PMID- 10858584 TI - High incidence of halotolerant bacteria in Pacific hydrothermal-vent and pelagic environments. AB - The abundance of halotolerant microorganisms in hydrothermal-vent and pelagic waters in the North and South Pacific was estimated by the most probable number (MPN) technique using a heterotrophic 16% NaCl medium incubated at 20-24 degrees C. Based on these MPNs and direct counts with epifluorescence microscopy to enumerate the total microbial population, salt-tolerant microbes comprised from <0.01 to >28% of the total microbial community. Fourteen isolates from these MPN enrichments were identified by sequencing a portion of the 16S rRNA gene, and all were found to belong to the genera Halomonas and Marinobacter. The response to salt of mesophilic hydrothermal-vent microbial isolates obtained without selecting for salt tolerance was also examined. Forty-one of 65 strains cultured from hydrothermal plume waters, low-temperature hydrothermal fluids, sulfide rock and an animal specimen at approximately 2000-2200 m depth from the Endeavour Segment of the Juan de Fuca Ridge were subjected to increasing concentrations of NaCl, and over half grew at a NaCl concentration that is lethal to many commonly isolated marine bacteria. At least 36 of the 65 isolates (>/=55%) grew in the enrichment medium supplemented with 10% NaCl; at least 30 of 65 (>/=46%) grew with 16% NaCl; at least 20 of 65 (>/=31%) tolerated 22% NaCl. Based on phylogenetic analysis of the 16S rRNA gene in nine of these 65 isolates, four belonged to the genus Halomonas. These Halomonas strains tolerated 22-27% NaCl. It is possible that a majority of the other 16 isolates which grew with 22% NaCl are also Halomonas based on their degree of halotolerance, morphology, and apparent abundance as revealed by MPN enrichments. The four Halomonas strains obtained without selecting for halotolerance were further characterized physiologically and metabolically. Overall, they grew between -1 degrees C and 40 degrees C, were facultative aerobes, oxidized between 49 and 70 organic compounds according to Biolog plate substrate utilization matrices, grew with oligotrophic quantities of carbon (0.002% yeast extract) in liquid media, reduced nitrate to nitrite, and tolerated up to 0.05-3 mM Cd(2+). Halomonas is one of the most abundant culturable organisms in the ocean, and its success may be attributed to its metabolic and physiological versatility. PMID- 10858585 TI - Effect of soil aggregate size on methanogenesis and archaeal community structure in anoxic rice field soil. AB - In anoxically incubated slurries of Italian rice field soil, CH(4) production is initiated after a lag phase during which ferric iron and sulfate are reduced. The production of CH(4) was affected by the size of soil aggregates used for the preparation of the soil slurry. Rates of CH(4) production were lowest with small aggregates (<50 and 50-100 um), were highest with aggregates of 200-2000 um size and were intermediate with aggregates of 2000-15000 um size. The different amounts of CH(4) accumulated were positively correlated to the concentrations of acetate, propionate and caproate that transiently accumulated in the slurries prepared from different aggregate sizes and also to the organic carbon content. The addition of organic debris that was collected from large-size aggregates to the aggregate size fractions <200 and <50 um resulted in an increase of CH(4) production to amounts that were comparable to those measured in unamended aggregates of 200-2000 um size, indicating that CH(4) production in the different aggregate size fractions was limited by substrate. The distribution of archaeal small-subunit rRNA genes in the different soil aggregate fractions was analyzed by terminal restriction fragment length polymorphism which allowed seven different archaeal ribotypes to be distinguished. Ribotype-182 (consisting of members of the Methanosarcinaceae and rice cluster VI), ribotype-389 (rice cluster I and II) and ribotype-820 (undigested DNA, rice cluster IV and members of the Methanosarcinaceae) accounted for >20, >30 and >10% of the total, respectively. The other ribotypes accounted for <10% of the total. The relative quantity of the individual ribotypes changed only slightly with incubation time and was almost the same among the different soil aggregate fractions. Ribotype 389, for example, slightly decreased with time, whereas ribotype-182 slightly increased. At the end of incubation, the relative quantity of ribotype-182 seemed to be slightly higher in soil fractions with larger than with smaller aggregates, whereas it was the opposite with ribotype-80 (Methanomicrobiaceae) and ribotype 88 (Methanobacteriaceae). Ribotype-280 (Methanosaetaceae and rice cluster V), ribotype-375 (rice cluster III), ribotype-389 and ribotype-820, on the other hand, were not much different among the different soil aggregate size fractions. However, the differences were not significant relative to the errors encountered during the extraction of polymerase chain reaction (PCR)-amplifiable DNA from soil. In conclusion, soil aggregate size and incubation time showed a strong effect on the function but only a small effect on the structure of the methanogenic microbial community. PMID- 10858587 TI - Causes or consequences of inflammation and pathological signs of Alzheimer disease. PMID- 10858588 TI - Neuroinflammation and Alzheimer's disease: critical roles for cytokine/Abeta induced glial activation, NF-kappaB, and apolipoprotein E. PMID- 10858589 TI - Alzheimer's disease: an inflammatory disease? PMID- 10858590 TI - Inflammation in Alzheimer's disease: a view from the periphery. PMID- 10858591 TI - Inflammatory hypotheses: novel mechanisms of Alzheimer's neurodegeneration and new therapeutic targets? PMID- 10858592 TI - Anti-inflammatory therapy for Alzheimer's disease. PMID- 10858593 TI - Anti-inflammatory drugs and Alzheimer's disease. PMID- 10858594 TI - Key issues in Alzheimer's disease inflammation. PMID- 10858595 TI - A selective defect of cytochrome c oxidase is present in brain of Alzheimer disease patients. AB - To assess mitochondrial function and test the hypothesis of an underlying oxidative phosphorylation defect in Alzheimer disease (AD), we evaluated the activities of mitochondrial respiratory chain enzyme complexes I+III, complexes II+III, complex IV (cytochrome c oxidase, COX), succinate dehydrogenase, and citrate synthase in the frontal cortex, temporal cortex, hippocampus, and cerebellum of 23 AD patients and 13 normal human brains. The major finding was a significant decrease in COX activity in AD temporal cortex and hippocampus, both whether activities were expressed per noncollagen protein content (49 +/-4.6 versus 78+/-10.8 nmol/min/mg NCP, P = 0.006; 23+/-1.9 versus 48.6+/-8.1 nmol/min/mg NCP, p = 0.003) or corrected for citrate synthase activity (1.6+/-0.2 versus 3+/-0.4, P = 0.001; 0.76+/-0.1 versus 1.76+/-0.26, P = 0.0009). There were no significant differences in the activities of complexes I+III, II+III, and of succinate dehydrogenase in any of the brain regions examined. Our results suggest a specific defect of COX in the AD brain versus the normal human brain, which may contribute to impaired energy generation. Biochemically, the defect is confined to selected brain regions, suggesting anatomic specificity. PMID- 10858596 TI - Chemotactic-like receptors and Abeta peptide induced responses in Alzheimer's disease. AB - Evidence suggests that beta-amyloid (Abeta) has chemokine-like properties and may act through formyl chemotactic receptors (FPR) to induce pathophysiologically important functional changes in Alzheimer's disease (AD) microglia. We have shown that Abeta 1-42, fibrillar Abeta 1-40, and Abeta 25-35 potentiate the release of interleukin-1beta (IL-1beta) from LPS activated human THP-1 monocytes [26] and LPS primed rat microglia. Moreover, Abeta-stimulated IL-1beta secretion seems to be receptor mediated because it is calcium dependent and requires activation of specific G-proteins [27]. Thus, we have evaluated the ability of Abeta 1-42 to mimic formyl chemotactic peptides in stimulating IL-1beta release from THP-1 monocytes. Several of the formyl chemotactic peptides and Abeta 1-42 significantly enhanced IL-1beta production in THP-1 monocytes. In contrast, a formyl chemotactic receptor antagonist inhibited Abeta 1-42-induced IL-1beta release from both human THP-1 monocytes and primary rat microglia. Further, primary rat microglia grown in culture expressed FPR as demonstrated by immunocytochemistry. Given the multiple pathophysiologic roles IL-1beta may play in AD, agents that block Abeta interactions with formyl chemotactic receptors on microglia might be important antiinflammatory therapeutic targets. PMID- 10858586 TI - Inflammation and Alzheimer's disease. AB - Inflammation clearly occurs in pathologically vulnerable regions of the Alzheimer's disease (AD) brain, and it does so with the full complexity of local peripheral inflammatory responses. In the periphery, degenerating tissue and the deposition of highly insoluble abnormal materials are classical stimulants of inflammation. Likewise, in the AD brain damaged neurons and neurites and highly insoluble amyloid beta peptide deposits and neurofibrillary tangles provide obvious stimuli for inflammation. Because these stimuli are discrete, microlocalized, and present from early preclinical to terminal stages of AD, local upregulation of complement, cytokines, acute phase reactants, and other inflammatory mediators is also discrete, microlocalized, and chronic. Cumulated over many years, direct and bystander damage from AD inflammatory mechanisms is likely to significantly exacerbate the very pathogenic processes that gave rise to it. Thus, animal models and clinical studies, although still in their infancy, strongly suggest that AD inflammation significantly contributes to AD pathogenesis. By better understanding AD inflammatory and immunoregulatory processes, it should be possible to develop anti-inflammatory approaches that may not cure AD but will likely help slow the progression or delay the onset of this devastating disorder. PMID- 10858597 TI - The women's health initiative estrogen replacement therapy is neurotrophic and neuroprotective. AB - The current study investigated the neurotrophic and neuroprotective action of the complex formulation of conjugated equine estrogens (CEEs), the most frequently prescribed estrogen replacement therapy in the United States and the estrogen replacement therapy of the Women's Health Initiative. Morphologic analyses demonstrated that CEEs significantly increased neuronal outgrowth in hippocampal, basal forebrain, occipital, parietal and frontal cortex neurons. Dose-response analyses indicated that the lowest effective concentration of CEEs exerted the maximal neurotrophic effect with greatest potency occurring in hippocampal and occipital cortex neurons. CEES induced highly significant neuroprotection against beta amyloid(25-35), hydrogen peroxide and glutamate-induced toxicity. Rank order of potency and magnitude of CEE-induced neuroprotection in the brain regions investigated was hippocampal neurons > basal forebrain neurons > cortical neurons. In hippocampal neurons pre-exposed to beta amyloid(25-35), CEEs halted Abeta(25-35)-induced cell death and protected surviving neurons from further cell death induced by Abeta(25-35). Because CEEs are the estrogen replacement therapy of the Women's Health Initiative, results of the current study could provide cellular mechanisms for understanding effects of CEEs on cognitive function and risk of Alzheimer's disease derived from this prospective clinical trial. PMID- 10858598 TI - The EVA spectral descriptor. AB - The EVA descriptor is derived from fundamental IR and Raman range molecular vibrational frequencies. EVA is sensitive to 3-D structure, but has an advantage over field-based 3-D QSAR methods inasmuch as it is invariant to both translation and rotation of the structures concerned and thus structural superposition is not required. The latter property and the demonstration of the effectiveness of the descriptor for QSAR means that EVA has been the subject of a great deal of interest from the modelling community. This review describes the derivation of the descriptor, details its main parameters and how to apply them, and provides an overview of the validation that has been done with the descriptor. A recent enhancement to the technique is described which involves the localised adjustment of variance in such a way that enhanced internal and external predictability may be obtained. Despite the statistical quality of EVA QSAR models, the main draw back to the descriptor at present is the difficulty associated with back-tracking from a PLS model to an EVA pharmacophore. Brief comment is made on the use of the EVA descriptor for diversity studies and the similarity searching of chemical structure databases. PMID- 10858599 TI - EPC syntheses and structure-activity relationships of hypoglycaemic semicyclic amidines. AB - A series of homochiral sterically hindered mono- and bicyclic amidines was prepared as hypoglycaemic agents by lethargic reaction of O-methylcaprolactim and 3-ethoxy-2-azabicyclo[2.2.2]oct-2-ene, respectively, with homochiral cis-2 substituted cyclopentane amines provided by asymmetrical reductive amination of racemic 2-substituted cyclopentanones. All compounds, except the cyclohexylmethyl isoquinuclidone derivative which inhibited secretion at 100 microM, significantly stimulated insulin secretion 2-8-fold at 10 microM and 100 microM in INS-1 cells. The most potent activator was the 2-cyclopentyl-substituted caprolactam derivative 5e. The stimulatory effects on secretion increased with rising steric hindrance of both the amidine alpha-carbon and the bicyclic amidine moiety itself. Enantiomeric discrimination was observed for the 2-?(cis-2 bulkysubstituted cyclopentyl)iminohexahydroazepine halides 5e and 5f and for the 3-?(cis-2-substituted cyclopentyl)imino-2-azabicyclo?2.2.2octane halides 6a and 6c. The amidines depolarized INS-1 cells and generated action potentials, accompanied by a decrease of membrane conductance. Simultaneously [Ca(2+)](i) increased, probably due to Ca(2+)-entry through voltage-dependent Ca(2+) channels. At high concentrations, where inhibition of secretion was observed, ?Ca(2+)(i) still rose upon application of the amidines, indicating an additional inhibitory pathway downstream to the elevation of ?Ca(2+)(i). Even at high concentrations (100 microM), the amidines had no toxic effects on insulin secreting INS-1 cells. PMID- 10858600 TI - Dependence of fungistatic activity of 2, 4-dihydroxythiobenzanilideson the structure and lipophilic nature of the compounds. AB - The quantitative dependencies of in vitro fungistatic action on the physico chemical parameters connected with the structure of 2, 4 dihydroxythiobenzanilides were investigated. It was stated that the action of these compounds depends on lipophilicity determined by substitution of the N-aryl moiety and on electron properties of molecules. The lipophilicity expressed by R(Mw) values was determined in the reversed-phase system (HPTLC). The changes in the nature of the thioamide bond were interpreted on the basis of UV and EI-MS spectra. PMID- 10858601 TI - Depsides as non-redox inhibitors of leukotriene B(4) biosynthesis and HaCaT cell growth, 2. Novel analogues of obtusatic acid. AB - Aseries of obtusatic acid analogues has been synthesized and evaluated as inhibitors of leukotriene B(4) (LTB(4)) biosynthesis and as antiproliferative agents. The 4-O-benzylated and the 4-O-demethylated congeners were the most potent inhibitors of LTB(4) production of the depside class of compounds, with IC(50) values in the submicromolar range. Furthermore, these compounds do not function as redox-based inhibitors because they were not reactive against a stable free radical, 2,2-diphenyl-1-picrylhydrazyl, and did not produce appreciable amounts of deoxyribose degradation as a measure of their potency to generate hydroxyl radicals. Some obtusatic acid congeners were also potent inhibitors of keratinocyte growth. Growth inhibition was not mediated by damage to the cell membrane, as the activity of lactate dehydrogenase released from the cytoplasm was in the control range. PMID- 10858602 TI - Pyrrolo[3,4-c]-beta-carboline-diones as a novel class of inhibitors of the platelet-derived growth factor receptor kinase. AB - Members of the structurally diverse family of beta-carbolines have previously been shown to exhibit a wide range of biological activities. A novel synthetic strategy for generation of beta-carbolines was developed, allowing imido-beta carbolines to be created in three steps from known compounds. The compounds were screened for inhibition of platelet-derived growth factor (PDGF)-stimulated tyrosine phosphorylation in Swiss 3T3 fibroblasts. A number of the newly synthesized beta-carbolines with moderate to potent inhibitory activity were revealed. The most active derivative, 2,3-dihydro-8,9-dimethoxy-5-(2 methylphenyl)-1H,6H-pyrrolo[3, 4-c]pyrido?3,4-bindole-1,3-dione 2ee, inhibited purified PDGF receptor kinase and PDGF-receptor autophosphorylation in intact cells with IC(50) values of 0.4 and 2.6 microM, respectively. Dione 2ee also inhibited PDGF-stimulated DNA synthesis in Swiss 3T3 fibroblasts with an IC(50) of 3.2 microM. The compound had no effect on Src or epidermal growth factor (EGF) receptor kinase activity and a six-seven-fold higher IC(50) for inhibition of basic fibroblast growth factor (bFGF)-stimulated tyrosine phosphorylation or Kit/stem cell factor (SCF) receptor autophosphorylation, indicating a reasonable extent of kinase specificity. Thus, beta-carbolines present a new lead of tyrosine kinase inhibitors with the capacity to selectively interfere with PDGF receptor signal transduction and PDGF-dependent cell growth. PMID- 10858603 TI - Plasma stability and cytotoxicity of lipophilic daunorubicin derivatives incorporated into low density lipoproteins. AB - The selective targeting of antineoplastic drugs to tumours by incorporation in low density lipoproteins (LDL) is an attractive possibility if the drug-LDL complex remains stable in the circulation and is taken up by the tumour. In previous studies we have shown that vincristine- and N-trifluoroacetyladriamycin 14-valerate-LDL complexes were unstable in vivo. We synthesized five N substituted lipophilic derivatives of daunorubicin and studied their incorporation into LDL. Three out of five daunorubicin derivatives incorporated successfully into LDL. In vitro these complexes were more cytotoxic towards LDL receptor positive Chinese hamster ovary cells than LDL receptor negative cells. Non-specific cytotoxicity was explained by slow dissociation of the drug-LDL complex in plasma. Our results underline the importance of careful studies of plasma stability when investigating lipoproteins and other carriers in drug targeting. PMID- 10858604 TI - Synthesis and pharmacological evaluation of new 4-2-(7-heterocyclemethoxynaftalen 2-ylmethoxy)ethylbenzoic acids as LTD(4)-antagonists. AB - A group of new 4-[2-(7-heterocyclemethoxynaftalen-2-ylmethoxy)ethyl]benzoic acids have been synthesized and pharmacologically evaluated as LTD(4)-antagonists. Thiazole derivatives, especially 4-[2-[7-(4-cyclobutylthiazole-2 ylmethoxyl)naphthalen- 2-ylmetho-xy]et hyl]benzoic acid, present considerable activity and improved pharmacokinetic profiles in comparison with our quinoline containing lead molecule confirming the interest of our compounds as potentially oral antiasthmatics and that the 4-alkylthiazole system can be considered as bioisosteric of the quinoline ring at least in our series of compounds. PMID- 10858605 TI - Synthesis of benzylideneacetophenones and their inhibition of lipid peroxidation. AB - A series of benzylideneacetophenone derivatives have been synthesized and found to show potent inhibition of the lipid peroxidation (LPO) in rat liver microsomes. All 19 compounds prepared in this series are LPO inhibitors. The highest activity was found in para hydroxy derivatives with two meta tert-butyl substituents. PMID- 10858606 TI - Vesicle number does not predict postsynaptic measures of miniature synaptic activity frequency in cultured cortical neurons. AB - We tested the hypothesis that heterogeneity in the frequency of miniature synaptic activity reflects differences in the number of vesicles present in presynaptic terminals. Using imaging techniques, we measured dendritic miniature synaptic calcium transients attributed to the spontaneous release of single transmitter quanta. Following imaging, the identified neurons were processed for serial transmission electron microscopy. At sites of quantal Ca(2+) transients mediated by N-methyl-D-aspartate receptors, we confirmed the presence of excitatory synapses and measured the total number of vesicles and the number of docked vesicles. We observed no correlation between the frequency of spontaneous miniature activity and either the total vesicle number or the number of docked vesicles. We conclude that the presynaptic vesicle complement as measured by ultrastructural analysis does not necessarily determine the frequency of spontaneous activity at synapses mediated by N-methyl-D-aspartate receptors. PMID- 10858607 TI - Synaptic transmission in the neocortex during reversible cooling. AB - We studied the effects of reversible cooling on synaptic transmission in slices of rat visual cortex. Cooling had marked monotonic effects on the temporal properties of synaptic transmission. It increased the latency of excitatory postsynaptic potentials and prolonged their time-course. Effects were non monotonic on other properties, such as amplitude of excitatory postsynaptic potentials and generation of spikes. The amplitude of excitatory postsynaptic potentials increased, decreased, or remain unchanged while cooling down to about 20 degrees C, but thereafter it declined gradually in all cells studied. The effect of moderate cooling on spike generation was increased excitability, most probably due to the ease with which a depolarized membrane potential could be brought to spike threshold by a sufficiently strong excitatory postsynaptic potential. Stimuli that were subthreshold above 30 degrees C could readily generate spikes at room temperature. Only at well below 10 degrees C could action potentials be completely suppressed. Paired-pulse facilitation was less at lower temperatures, indicating that synaptic dynamics are different at room temperature as compared with physiological temperatures. These results have important implications for extrapolating in vitro data obtained at room temperatures to higher temperatures. The data also emphasize that inactivation by cooling might be a useful tool for studying interactions between brain regions, but the data recorded within the cooled area do not allow reliable conclusions to be drawn about neural operations at normal temperatures. PMID- 10858608 TI - Nerve growth factor but not neurotrophin-3 is synthesized by hippocampal GABAergic neurons that project to the medial septum. AB - Conventional uptake of neurotrophins takes place at axon terminals via specific receptors, and is followed by retrograde transport. Recent studies demonstrated that, with the exception of nerve growth factor, other neurotrophins may be delivered anterogradely to the region containing the receptor expressing neurons. In this study we used a triple labeling method that combines retrograde tract tracing, in situ hybridization and immunocytochemistry to examine whether non principal cells projecting from the hippocampus to the septum synthesize nerve growth factor. Our results show that, on average, 59% of the horseradish peroxidase-labeled hippocamposeptal nonpyramidal neurons also display nerve growth factor messenger RNA hybridization signal. The ratio was slightly higher in the CA1 stratum oriens and the hilus of the dentate gyrus (64 and 62%, respectively) compared to stratum oriens of the CA3 region (58%). In addition, we demonstrated that many nerve growth factor-positive septally projecting neurons also contain the calcium-binding protein calbindin D-28K, whereas nerve growth factor-negative projecting cells mostly lack this neurochemical marker. In contrast to nerve growth factor, neurotrophin-3 has never been found in hippocamposeptal cells. Hippocamposeptal GABAergic cells are reciprocally connected with the medial septum, thus they are in a key position to regulate nerve growth factor release as a function of the activity level in the septohippocampal system. Furthermore, our results raise the intriguing possibility that nerve growth factor may be transported also in an anterograde manner. Regardless of the direction of transport, the presence of nerve growth factor in hippocamposeptal cells suggests that long distance fast synaptic mechanisms and slow neurotrophin action are coupled in these neurons. PMID- 10858609 TI - Electroconvulsive stimuli enhance both neuropeptide Y receptor Y1 and Y2 messenger RNA expression and levels of binding in the rat hippocampus. AB - Repeated electroconvulsive stimulations and other seizure modalities produce an increase in neuropeptide Y synthesis and local release in the rat hippocampus, and perhaps as a consequence, a change in the concentration of neuropeptide Y binding sites in the same region. The aim of the present study was to determine possible changes in the expression of neuropeptide Y receptor subtypes affected by repeated stimulations in the hippocampus. Rats were exposed to 14 daily stimulations, and the brains were removed 24h after the last stimulation. For in vitro receptor autoradiography and in situ hybridisation histochemistry, the brains were frozen, sectioned, and levels of neuropeptide Y binding sites and messenger RNA expressions were determined quantitatively on sections from the same animals. In order to determine the contribution of different neuropeptide Y receptor subtypes, serial sections were incubated with either 125I-labelled peptide YY alone or the same radio-labelled peptide mixed with an excess of a number of displacing compounds with affinity for either neuropeptide Y receptor subtype Y1, Y2, or both. Binding studies revealed that the majority of peptide YY binding sites was represented by Y2, and that electroconvulsive stimulations reduced the binding capacity or the concentration of this receptor. A prominent reduction of Y1-preferring binding sites was determined in the dentate gyrus, and to a lesser extent in the CA1 and CA3 regions. Similarly, the treatment produced a significant reduction of Y2-preferring binding sites in the CA1 and CA3 region, but not in the granular cell layer of the dentate gyrus. Using semi-quantitative in situ hybridization, Y1 receptor messenger RNA level in the granular cell layer of the dentate increased by the stimulations. In the same region, Y2 receptor messenger RNA was expressed in low to undetectable amounts, but after the repeated stimulations, this transcript was found in moderate to high levels. These data suggest that the neuropeptide Yergic system in the dentate gyrus and the pyramidal cell layer are affected by the treatment, and that this includes both Y1 and Y2 receptor subtypes. Because levels of messenger RNA and binding are distinctly regulated, the turnover of both Y1 and Y2 molecules is strongly increased under electroconvulsive stimulations, suggesting that the intrahippocampal neuropeptide Yergic neurotransmission is also increased under the stimulations. PMID- 10858610 TI - Kainic acid-induced seizures produce necrotic, not apoptotic, neurons with internucleosomal DNA cleavage: implications for programmed cell death mechanisms. AB - Prolonged seizures (status epilepticus) induced by kainic acid activate programmed cell death mechanisms, and it is believed that kainic acid-induced status epilepticus induces neuronal apoptosis. In order to test this hypothesis, adult rats were subjected to 3-h kainic acid-induced seizures, with 24- or 72-h recovery periods. Neuronal death was assessed by light microscopy with the Hematoxylin and Eosin stain and with in situ terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL stain), by electron microscopy, and by agarose gel electrophoresis of DNA extracted from five vulnerable brain regions. Spontaneous and MK-801-induced apoptotic neurons from retrosplenial cortex of neonatal rats, evaluated by light and electron microscopy, were used as positive controls for apoptosis. Surprisingly, the large chromatin clumps of apoptotic neurons were TUNEL negative, whereas the cytoplasm showed light-to-moderate TUNEL staining, consistent with a lack of identifiable nuclear membranes ultrastructurally, and with intermingling of nuclear and cytoplasmic contents. Ultrastructurally, the acidophilic neurons produced by kainic acid-induced status epilepticus, identified with Hematoxylin and Eosin stain, were dark, shrunken and necrotic, with pyknotic nuclei containing small, dispersed chromatin clumps, and with cytoplasmic vacuoles, some of which were swollen, disrupted mitochondria. No apoptotic cells were seen. Acidophilic neurons were found in up to 20 of 23 brain regions examined and comprised 10-25% of the total number of neurons examined. A subset of these neurons (<10% of the total number of neurons in five of 23 regions) had TUNEL-positive nuclei 72h but not 24h after status epilepticus. Internucleosomal DNA cleavage (DNA "laddering") occurred in the four most damaged brain regions examined by electron microscopy 24h after SE and the three most damaged regions 72h after status epilepticus. Our results demonstrate that kainic acid-induced status epilepticus produces neuronal necrosis and not apoptosis in adult rats. The necrotic neurons show nuclear pyknosis, chromatin condensation and DNA laddering. Programmed cell death mechanisms activated by kainic acid induced status epilepticus occur in neurons which become necrotic and could contribute to necrotic, as well as apoptotic, neuronal death. PMID- 10858611 TI - Amygdaloid N-methyl-D-aspartate and gamma-aminobutyric acid(A) receptors regulate sensorimotor gating in a dopamine-dependent way in rats. AB - Sensorimotor gating can be measured as prepulse inhibition of the startle response in humans and rats. Since prepulse inhibition is impaired in schizophrenics there is considerable interest in understanding the neuronal basis of prepulse inhibition. Neuropathological findings indicate a dysfunction of the glutamatergic and GABAergic system in cortico-limbic areas in schizophrenics. We tested whether blockade of N-methyl-D-aspartate or GABA(A) receptors in the basolateral amygdala affects prepulse inhibition in rats. Local infusion of the N methyl-D-aspartate receptor antagonist dizocilpine (0, 6.25 microg/0.5 microl), or of the GABA(A) receptor antagonist picrotoxin (0, 5.0, 10.0 ng/0.5 microl) reduced prepulse inhibition. The prepulse inhibition-disrupting effect of 6.25 microg dizocilpine or 10.0 ng picrotoxin was reversed by systemic co administration of the dopamine antagonist haloperidol (0.1mg/kg i.p.). These data indicate that sensorimotor gating is regulated in a dopamine-dependent way by N methyl-D-aspartate and GABA(A) receptors in the basolateral amygdala. Our findings are discussed with respect to neuropathological findings in schizophrenics. PMID- 10858612 TI - Effects of oligonucleotide antisense to dopamine D(1A) receptor messenger RNA in a rodent model of levodopa-induced dyskinesia. AB - Dyskinesias are abnormal involuntary movements which develop as a side-effect of long-term treatment with levodopa in patients with Parkinson's disease. The pathophysiology underlying these dyskinesias remains unclear, although, it has been suggested that heightened activity of dopamine D(1) receptor-bearing striatonigral neurons may play a key role. Chronic pulsatile levodopa administration to hemiparkinsonian rats results in sensitization of rotational responses to apomorphine. This sensitization is thought to be analogous to levodopa-induced dyskinesias in humans. In these studies, we further clarify the role of the dopamine D(1A) receptor in this rodent model of levodopa-induced dyskinesias using an in vivo oligonucleotide antisense approach. Hemiparkinsonian rats received twice daily injections of levodopa for three weeks followed by intrastriatal infusion of dopamine D(1A) receptor antisense (7nmol/day, three days), a scrambled missense control sequence, or saline. Those animals treated with antisense displayed significantly fewer apomorphine-induced rotations than saline- or missense-treated controls.By reducing dopamine D(1A) receptor expression, we were able to attenuate sensitization of the response to apomorphine resulting from chronic pulsatile levodopa treatment. Thus, the dopamine D(1A) receptor appears to play a significant role in levodopa-induced dyskinesias and warrants further examination. These findings may have important implications for the development of selective treatment strategies designed to alleviate parkinsonian symptoms, while minimizing motor complications. PMID- 10858613 TI - Changes in CArG-binding protein A expression levels following injection(s) of the D1-dopamine agonist SKF-82958 in the intact and 6-hydroxydopamine-lesioned rat. AB - We recently characterized the rat brain homolog of mouse muscle CArG-binding protein A initially identified in C2 myogenic cells and showed an inverse temporal correlation between increased expression levels of this messenger RNA, c fos and zif268 messenger RNA levels following the addition of nerve growth factor to PC12 cells. In addition, we found an inverse correlation between c-Fos protein and CArG-binding protein A messenger RNA levels in the lateral caudate-putamen of rats treated acutely and chronically with the D2 receptor antagonist fluphenazine (phenothiozine typical psychotic). To determine whether D1 receptor stimulation is also capable of inducing CArG-binding protein A up-regulation, drug naive or dopamine-depleted (i.e. 6-hydroxydopamine-lesioned) D1 hypersensitized rats (i.e. rats given repeated daily injections of SKF-82958 for 14days) were acutely injected with the D1 agonist SKF-82958 and examined using a combination of in situ hybridization for CArG binding protein A and immunocytochemistry for c-Fos. Both acutely treated animals and dopamine-depleted hypersensitized animals showed increases in CArG-binding protein A. Moderate increases were found in the medial caudate-putamen and nucleus accumbens core and shell regions following acute treatment whereas large increases in CArG-binding protein A expression levels were found in the medial and lateral caudate-putamen and the shell and core of the nucleus accumbens following hypersensitization. No change in CArG-binding protein A expression level was found in the dopamine-depleted, drug naive animals relative to controls. Regions of the basal ganglia where increases in CArG binding protein A were detected following each treatment correlated perfectly with c-Fos protein induction. The results demonstrate that CArG-binding protein A responds to SKF-82958 and that the changes in CArG-binding protein A match perfectly with the pattern of c-Fos induction induced by the D1 agonist. PMID- 10858614 TI - Nicotine binding in human striatum: elevation in schizophrenia and reductions in dementia with Lewy bodies, Parkinson's disease and Alzheimer's disease and in relation to neuroleptic medication. AB - Striatal nicotinic acetylcholine receptors with high affinity for nicotinic agonists are involved with the release of a number of neurotransmitters, including dopamine. Previous findings as to whether these receptors are changed in Parkinson's disease and Alzheimer's disease are inconsistent and no previous investigations have focused on these receptors in dementia with Lewy bodies and schizophrenia, which are also associated with disorders of movement. The present autoradiographic study of striatal [3H]nicotine binding in Alzheimer's and Parkinson's diseases, dementia with Lewy bodies and schizophrenia was conducted with particular reference to the potentially confounding variables of tobacco use and neuroleptic medication. [3H]Nicotine binding in both dorsal and ventral caudate and putamen was significantly reduced in Parkinson's disease (43-67%, n=13), Alzheimer's disease (29-37%, n=13) and dementia with Lewy bodies (50-61%, n=20) compared to age-matched controls (n=42). Although tobacco use in the control group was associated with increased [3H]nicotine binding (21-38%), and neuroleptic treatment in dementia with Lewy bodies and Alzheimer's disease was associated with reduced [3H]nicotine binding (up to 29%), differences between neurodegenerative disease groups and controls persisted in subgroups of Alzheimer's disease cases (26-33%, n=6, in the ventral striatum) and dementia with Lewy body cases (30-49%, n=7, in both dorsal and ventral striatum) who had received no neuroleptic medication compared to controls who had not smoked (n=10). In contrast, striatal [3H]nicotine binding in a group of elderly (56-85 years) chronically medicated individuals with schizophrenia (n=6) was elevated compared with the entire control group (48-78%, n=42) and with a subgroup that had smoked (24-49%, n=8). The changes observed in [3H]nicotine binding are likely to reflect the presence of these receptors on multiple sites within the striatum, which may be differentially modulated in the different diseases. Further study is warranted to explore which nicotinic receptor subunits and which neuronal compartments are involved in the changes in [3H]nicotine binding reported, to aid development of potential nicotinic receptor therapy. PMID- 10858615 TI - Neurturin protects striatal projection neurons but not interneurons in a rat model of Huntington's disease. AB - Glial cell line-derived neurotrophic factor and neurturin are neurotrophic factors expressed in the striatum during development and in the adult rat. Both molecules act as target-derived neurotrophic factors for nigrostriatal dopaminergic neurons. While glial cell line-derived neurotrophic factor has also been described to have local trophic effects on striatal neurons, the effects of neurturin in the striatum have not yet been described. Here we examine whether neurturin protects striatal projection neurons (calbindin-positive) and interneurons (parvalbumin- or choline acetyltransferase-positive) in an animal model of Huntington's disease. A fibroblast cell line engineered to over-express neurturin was grafted into adult rat striatum 24h before quinolinate injection. In animals grafted with a control cell line, intrastriatal quinolinate injection reduced the number of calbindin-, parvalbumin- and choline acetyltransferase positive neurons, seven days post-lesion. Intrastriatal grafting of neurturin secreting cells protected striatal projection neurons, but not interneurons, from quinolinate excitotoxicity. This effect was much more robust than that reported previously for a glial cell line-derived neurotrophic factor-secreting cell line on striatal calbindin-positive neurons. However, intrastriatal grafting of glial cell line-derived neurotrophic factor- but not neurturin-secreting cells prevented the decrease in choline acetyltransferase activity induced by quinolinate injection. Taken together, our results show that neurturin- and glial cell line-derived neurotrophic factor-secreting cell lines have clearly differential effects on striatal neurons. Grafting of the neurturin-secreting cell line showed a more specific and efficient trophic effect on striatal projection neurons, the neuronal population most affected in Huntington's disease. Therefore, our results suggest that neurturin is a good candidate for the treatment of this neurodegenerative disorder. PMID- 10858616 TI - Organization of somatic motor inputs from the frontal lobe to the pedunculopontine tegmental nucleus in the macaque monkey. AB - To reveal the somatotopy of the pedunculopontine tegmental nucleus that functions as a brainstem motor center, we examined the distribution patterns of corticotegmental inputs from the somatic motor areas of the frontal lobe in the macaque monkey. Based on the somatotopical map prepared by intracortical microstimulation, injections of the anterograde tracers, biotinylated dextran amine and wheat germ agglutinin-conjugated horseradish peroxidase, were made into the following motor-related areas: the primary motor cortex, the supplementary and presupplementary motor areas, the dorsal and ventral divisions of the premotor cortex, and the frontal eye field. Data obtained from the present experiments were as follows: (i) Corticotegmental inputs from orofacial, forelimb, and hindlimb representations of the primary motor cortex tended to be arranged orderly from medial to lateral in the pedunculopontine tegmental nucleus. However, the distribution areas of these inputs considerably overlapped; (ii) The major input zones from distal representations of the forelimb and hindlimb regions of the primary motor cortex were located medial to those from their proximal representations, although there was a substantial overlap between the distribution areas of distal versus proximal limb inputs; (iii) The main terminal zones from the forelimb regions of the primary motor cortex, the supplementary and presupplementary motor areas, and the dorsal and ventral divisions of the premotor cortex appeared to overlap largely in the mediolaterally middle aspect of the pedunculopontine tegmental nucleus; and (iv) Corticotegmental input from the frontal eye field was scattered over the pedunculopontine tegmental nucleus.Thus, the present results indicate that the pedunculopontine tegmental nucleus is likely to receive partly separate but essentially convergent cortical inputs not only from multiple motor-related areas representing the same body part, but also from multiple regions representing diverse body parts. This suggests that somatotopical representations are intermingled rather than segregated in the pedunculopontine tegmental nucleus. PMID- 10858617 TI - Backpropagation of the delta oscillation and the retinal excitatory postsynaptic potential in a multi-compartment model of thalamocortical neurons. AB - Uniform and non-uniform somato-dendritic distributions of the ion channels carrying the low-threshold Ca(2+) current (I(T)), the hyperpolarization-activated inward current (I(h)), the fast Na(+) current (I(Na)) and the delayed rectifier current (I(K)) were investigated in a multi-compartment model of a thalamocortical neuron for their suitability to reproduce the delta oscillation and the retinal excitatory post-synaptic potential recorded in vitro from the soma of thalamocortical neurons. The backpropagation of these simulated activities along the dendritic tree was also studied. A uniform somato-dendritic distribution of the maximal conductance of I(T) and I(K) (g(T) and g(K), respectively) was sufficient to simulate with acceptable accuracy: (i) the delta oscillation, and its phase resetting by somatically injected current pulses; as well as (ii) the retinal excitatory postsynaptic potential, and its alpha-amino-3 hydroxy-5-methyl-4-isoxazole proprionate and/or N-methyl-D-aspartate components. In addition, simulations where the dendritic g(T) and g(K) were either reduced (both by up to 34%) or increased (both by up to 15%) of their respective value on the soma still admitted a successful reproduction of the experimental activity. When the dendritic distributions were non-uniform, models where the proximal and distal dendritic g(T) was up to 1.8- and 1. 2-fold larger, respectively, than g(T(s)) produced accurate simulations of the delta oscillation (and its phase resetting curves) as well as the synaptic potentials without need of a concomitant increase in proximal or distal dendritic g(K). Furthermore, an increase in proximal dendritic g(T) and g(K) of up to fourfold their respective value on the soma resulted in acceptable simulation results. Addition of dendritic Na(+) channels to the uniformly or non-uniformly distributed somato dendritic T-type Ca(2+) and K(+) channels did not further improve the overall qualitative and quantitative accuracy of the simulations, except for increasing the number of action potentials in bursts elicited by low-threshold Ca(2+) potentials. Dendritic I(h) failed to produce a marked effect on the simulated delta oscillation and the excitatory postsynaptic potential. In the presence of uniform and non-uniform dendritic g(T) and g(K), the delta oscillation propagated from the soma to the distal dendrites with no change in frequency and voltage dependence, though the dendritic action potential amplitude was gradually reduced towards the distal dendrites. The amplitude and rising time of the simulated retinal excitatory postsynaptic potential were only slightly decreased during their propagation from their proximal dendritic site of origin to the soma or the distal dendrites. These results indicate that a multi-compartment model with passive dendrites cannot fully reproduce the experimental activity of thalamocortical neurons, while both uniform and non-uniform somato-dendritic g(T) and g(K) distributions are compatible with the properties of the delta oscillation and the retinal excitatory postsynaptic potential recorded in vitro from the soma of these neurons. Furthermore, by predicting the existence of backpropagation of low-threshold Ca(2+) potentials and retinal postsynaptic potentials up to the distal dendrites, our findings suggest a putative role for the delta oscillation in the dendritic processing of neuronal activity, and support previous hypotheses on the interaction between retinal and cortical excitatory postsynaptic potentials on thalamocortical neuron dendrites. PMID- 10858618 TI - Presynaptic GABA(B) receptors modulate organum vasculosum lamina terminalis evoked postsynaptic currents in rat hypothalamic supraoptic neurons. AB - Whole-cell patch-clamp recordings obtained from 36 hypothalamic supraoptic nucleus neurons in explant preparations evaluated a role for GABA(B) receptors in modulating postsynaptic inhibitory and excitatory currents evoked by electrical stimulation in the organum vasculosum of the lamina terminalis. At a holding current of -65 mV, application of baclofen (1-10 microM) induced a dose-dependent reduction in the amplitude of pharmacologically isolated inhibitory and excitatory postsynaptic currents, converted paired-pulse depression in inhibitory postsynaptic currents to paired-pulse facilitation, and enhanced paired-pulse ratios for excitatory postsynaptic currents. In media containing 2 hydroxysaclofen (200-400 microM), baclofen-associated events were blocked and paired-pulse depression in evoked inhibitory postsynaptic currents was abolished. In addition, a progressive increase in the amplitude of inhibitory postsynaptic currents implied that GABA was endogenously active at presynaptic GABA(B) receptors. In contrast, no paired-pulse depression was observed for inhibitory postsynaptic currents evoked in six non-magnocellular neurons. Neither baclofen nor 2-hydroxysaclofen altered holding currents or input resistances in supraoptic neurons, or altered the kinetics of the evoked responses.These observations imply that the terminals of both inhibitory (GABAergic) and excitatory (glutamatergic) afferents to supraoptic nucleus neurons from organum vasculosum lamina terminalis neurons are subject to modulation by presynaptic GABA(B) receptors, and that this modulation is preferentially directed to the inhibitory inputs. PMID- 10858619 TI - Inflammatory mediators release calcitonin gene-related peptide from dorsal root ganglion neurons of the rat. AB - The interactions between the inflammatory mediators bradykinin, serotonin, prostaglandin E(2) and acid pH were studied in rat dorsal root ganglion neurons in culture. For this purpose, the cultures were stimulated by inflammatory mediators (bradykinin, serotonin, prostaglandin E(2), 10(-5)M each) or acid solution (pH 6.1) for 5 min and the content of calcitonin gene-related peptide was determined in the supernatant before, during and after stimulation, using an enzyme immunoassay. Acid solution resulted in a threefold increase of the basal calcitonin gene-related peptide release which was entirely dependent on the presence of extracellular calcium. The release could not be blocked by the addition of the capsaicin antagonist capsazepine (10(-5)M). Bradykinin (10(-5)M) caused a 50% increase of the basal calcitonin gene-related peptide release which was again dependent on the presence of extracellular calcium, whereas serotonin and prostaglandin E(2) were each ineffective at 10(-5)M concentration. The combination of bradykinin, serotonin and prostaglandin E(2) led to a fivefold increase of the calcitonin gene-related peptide release which could not be further enhanced by acidification. The competitive capsaicin receptor antagonist capsazepine (10(-5)M) significantly reduced the release induced by the combination of bradykinin, serotonin and prostaglandin E(2). It is suggested that the inflammatory mediators co-operate and together may act as endogenous agonists at the capsaicin receptor to cause calcium influx and consecutive neuropeptide release. PMID- 10858620 TI - Induction of homosynaptic long-term depression at spinal synapses of sensory a delta-fibers requires activation of metabotropic glutamate receptors. AB - The synaptic strength between primary afferent Adelta-fibers, many of which convey pain-related information, and second order neurons in the spinal dorsal horn can be depressed for prolonged periods of time in a use- and N-methyl-D aspartate receptor-dependent fashion. Here, we have used a transverse spinal cord slice-dorsal root preparation of young rat to characterize the nature of this form of long-term depression and the role of metabotropic glutamate receptors. Dorsal roots were bisected and intracellular recordings were made from lamina II neurons with independent excitatory synaptic inputs from both dorsal root halves. Conditioning stimulation of one dorsal root half (1 Hz, 900 pulses) induced long term depression that was specific for the stimulated pathway, i. e. homosynaptic in nature. The induction of long-term depression was prevented by non-selective group I and group II mGluR antagonist (S)-alpha-methyl-4-carboxyphenylglycine, by selective group I receptor antagonist (S)-4-carboxyphenylglycine and by selective group II mGluR antagonist (RS)-alpha-methylserine-O-phosphate monophenyl ester. Group III mGluR antagonist (RS)-alpha-methylserine-O-phosphate was ineffective. Short-term depression was not affected by any of these antagonists.Thus, a homosynaptic form of long-term depression exists at putative nociceptive synapses in the spinal dorsal horn and its induction requires the activation of both group I and II metabotropic glutamate receptors. PMID- 10858621 TI - Decreased levels of proteasome activity and proteasome expression in aging spinal cord. AB - Neuron death and neuron degeneration occur in the CNS during the course of aging. Although multiple cellular alterations transpire during the aging process, those that mediate age-associated neuron death have not been identified. Recent evidence implicates oxidative stress as a possible means of neuron death and neuron degeneration during aging. In the present study, we demonstrate a marked decrease in multicatalytic proteasome activity in the spinal cord of Fisher 344 rats at 12, 24 and 28 months, compared with spinal cord tissue from 3-week- and 3 month-old animals. Application of oxidative injury (FeSO(4)) or the lipid peroxidation product 4-hydroxynonenal decreases multicatalytic proteasome activity in a time- and dose-dependent manner in a motor neuron cell line. Loss of multicatalytic proteasome activity occurs before the loss of multicatalytic proteasome immunoreactivity, with FeSO(4)- and 4-hydroxynonenal-mediated decreases ameliorated by the application of a cell permeable form of the antioxidant glutathione. Application of multicatalytic proteasome inhibitors, but not inhibitors of lysosomal proteases, induced neuron death that was attenuated by the caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp-(O-methyl) fluoromethyl ketone or N-acetyl-Asp-Glu-Val-Asp-Cho (aldehyde). Together, these data suggest that multicatalytic proteasome inhibition occurs during aging of the spinal cord, possibly as the result of oxidative stress, and that multicatalytic proteasome inhibition may be causally related to neuron death. PMID- 10858622 TI - Temperature dependence of membrane sealing following transection in mammalian spinal cord axons. AB - Using an in vitro sucrose-gap recording chamber, sealing of cut axons in isolated strips of white matter from guinea pig spinal cord was measured by recording the "compound membrane potential". This functional sealing was found to correlate well with anatomical resealing, measured by a horseradish peroxidase uptake assay. Near-complete functional and anatomical recovery of the axonal membrane occurred routinely within 60 min following transection at 37 degrees C in regular Krebs' solution. The rate of membrane potential recovery is exponential, with a time-constant of 20+/-5 min. The sealing process at 31 degrees C was similar to that at 37 degrees C, and was effectively blocked at 25 degrees C, under which condition most axons continued to take up horseradish peroxidase for more than 1h, and failed to substantially recover their membrane potential. Seventy-five percent of the cords transected at 40 degrees C had similar sealing behavior to those at 37 degrees C and 31 degrees C. The balance failed to seal the cut end. Two-dimensional morphometric analysis has shown that raising the temperature from 25 degrees C to above 31 degrees C significantly decreases axonal permeabilization to horseradish peroxidase (increases the sealing of transected ends) across all areas of a transverse section of spinal cord. Moreover, this enhancement of sealing exists across all axon calibers. Since severe cooling compromises membrane resealing, caution needs to be taken when hypothermic treatment (below 25 degrees C) is applied within the first 60 min following mechanical injury. In summary, we have found that at normal temperature (37 degrees C), nerve fibers repair their damaged membrane following physical injury with an hour. This is similar at mildly lower (31 degrees C) and relatively higher (40 degrees C) temperature, although some fibers tend to collapse under this febrile temperature. Moreover, severely low temperature (25 degrees C) hindered the repair of damaged membranes. Based on our study, caution is needed in treating spinal cord injury with low temperatures. PMID- 10858623 TI - Fos immunoreactivity in the lamina terminalis of adrenalectomized rats and effects of angiotension II type 1 receptor blockade or deoxycorticosterone. AB - Neural activity, as measured immunohistochemically by the presence of Fos protein, was determined in the lamina terminalis, a thin strip of tissue forming the anterior wall of the third brain ventricle, after adrenalectomy. Several weeks after surgery, the adrenalectomized rats were maintained with access to water and a low sodium diet for five days. In addition, hypertonic (0.5M) NaCl solution was available for the entire five-day period (sodium available) or only during the first four days (sodium unavailable). The number of neurons expressing Fos, determined at the end of the fifth day, was increased in the adrenalectomized rats with or without NaCl solution to drink. Fos activity in the median preoptic nucleus was increased only in adrenalectomized rats without access to NaCl solution. Treatment of adrenalectomized rats with the sodium retaining mineralocorticoid hormone, deoxycorticosterone, at the end of the fourth day, decreased Fos expression in the subfornical organ and the organum vasculosum of the lamina terminalis when NaCl solution was available but not when the NaCl solution was unavailable. In the adrenalectomized rats with NaCl solution available, mineralocorticoid treatment decreased both urinary sodium excretion and daily sodium intake. Brain nuclei in the lamina terminalis also became activated in intact rats made sodium deplete by treatment with the diuretic, furosemide. Relative to sodium-deplete intact rats, however, sodium deplete adrenalectomized rats had a greater number of neurons expressing Fos in the organum vasculosum. Treatment of sodium-deplete rats, adrenalectomized or intact, with the angiotensin II-type 1 receptor antagonist, ZD7155, decreased sodium intake and Fos expression in the subfornical organ but not in the organum vasculosum of the lamina terminalis or median preoptic nucleus. In conclusion, the results demonstrated that activation of the brain nuclei located in the lamina terminalis of adrenalectomized rats was primarily related to sodium deficit and not to the absence of the mineralocorticoid hormones, although the adrenal hormones may have a role in limiting the activation of organum vasculosum of the lamina terminalis during sodium depletion. Furthermore, the results obtained with the administration of the angiotensin receptor antagonist are consistent with the proposal that sodium appetite of the sodium-deplete rat, adrenalectomized or intact, is mediated by circulating angiotensin II acting in the subfornical organ. PMID- 10858624 TI - Modulation of odor-induced increases in [Ca(2+)](i) by inhibitors of protein kinases A and C in rat and human olfactory receptor neurons. AB - Protein kinases A and C have been postulated to exert multiple effects on different elements of signal transduction pathways in olfactory receptor neurons. However, little is known about the modulation of olfactory responses by protein kinases in intact olfactory receptor neurons. To further elucidate the details of the modulation of odorant responsiveness by these protein kinases, we investigated the action of two protein kinase inhibitors: H89, an inhibitor of protein kinase A, and N-myristoylated EGF receptor, an inhibitor of protein kinase C, on odorant responsiveness in intact olfactory neurons. We isolated individual olfactory neurons from the adult human and rat olfactory epithelium and measured responses of the isolated cells to odorants or biochemical activators that have been shown to initiate cyclic AMP or inositol 1,4,5 trisphospate production in biochemical preparations. We employed calcium imaging techniques to measure odor-elicited changes in intracellular calcium that occur over several seconds. In human olfactory receptor neurons, the protein kinase A and C inhibitors affected the responses to different sets of odorants. In rats, however, the protein kinase C inhibitor affected responses to all odorants, while the protein kinase A inhibitor had no effect. In both species, the effect of inhibition of protein kinases was to enhance the elevation and block termination of intracellular calcium levels elicited by odorants. Our results show that protein kinases A and C may modulate odorant responses of olfactory neurons by regulating calcium fluxes that occur several seconds after odorant stimulation. The effects of protein kinase C inhibition are different in rat and human olfactory neurons, indicating that species differences are an important consideration when applying data from animal studies to apply to humans. PMID- 10858625 TI - Beta-chemokines and human immunodeficiency virus type-1 proteins evoke intracellular calcium increases in human microglia. AB - Activation of beta-chemokine receptors, co-receptors for human immunodeficiency virus type-1 (HIV-1), stimulates movement and secretion in microglia, possibly through a Ca(2+)-dependent mechanism. We studied chemokine activation of Ca(2+) signaling processes in microglia. Human fetal microglia were grown in primary culture and chemokine-induced increases in intracellular calcium concentration ([Ca(2+)](i)) were measured in single cells using indo-1-based microfluorimetry. Application of 50 ng/ml regulated on activation, normal T expressed and secreted (RANTES; 120 s) evoked responses in 26% of the microglia (187/719 cells). [Ca(2+)](i) increased from a basal level of 66+/-6 nM to peak at 268+/-23 nM (n=187). Chemokine-evoked responses rapidly desensitized as indicated by the rapid return to basal [Ca(2+)](i) levels in the maintained presence of RANTES. The removal of extracellular Ca(2+) or stimulation in the presence of Ni(2+) (2mM) or La(3+) (100 microM) blocked the RANTES-elicited [Ca(2+)](i) increase. The L-type calcium channel antagonist nimodipine (10 microM) inhibited the RANTES mediated increase in [Ca(2+)](i) by 80+/-16%. Thus, the RANTES-evoked calcium transient appears to result from Ca(2+) influx with little if any release from intracellular stores. Application of gp120(clade) (E) and gp120(CM235) (50 ng/ml) neither mimicked nor antagonized the RANTES-evoked response. Application of 50 ng/ml eotaxin (120 s) evoked an increase in [Ca(2+)](i) in 13% of the human microglia in culture (61/469 cells). The HIV-1 regulatory protein Tat (50 ng/ml) increased the [Ca(2+)](i) in a subset of eotaxin-responsive cells (16/30). The L type calcium channel antagonist nimodipine (3 microM) inhibited eotaxin- and Tat mediated increases in [Ca(2+)](i) by 88+/-6% and 93+/-6%, respectively. Thus, activation of CCR3 appears to evoke Ca(2+) influx through L-type Ca(2+) channels.These results indicate that beta-chemokines, RANTES and eotaxin, activate a nimodipine sensitive Ca(2+) influx pathway in human fetal microglia. HIV-1 Tat protein mimicked chemokine-mediated Ca(2+) signaling and may modulate the migratory and secretory responses of microglia. PMID- 10858626 TI - Cognitive behavioral psychotherapy for schizophrenia: a review of recent empirical studies. AB - A set of cognitive behavioral psychotherapies (CBT) has been developed for schizophrenia. These interventions have been used for the treatment of both recent onset patients and those with treatment-refractory symptoms. This article reviews clinical trials of CBT for schizophrenia since 1990. The CBT interventions appear to be beneficial in reducing overall symptom levels, especially the severity of delusions. The relative efficacy of CBT is more evident when CBT is compared with routine care than when it is compared with other therapies matched for therapist attention. Further studies are needed to objectively determine the active ingredients of CBT and to better identify the interactions of CBT with concurrent psychosocial and medication treatments. PMID- 10858627 TI - Decreased S100-beta protein in schizophrenia: preliminary evidence. AB - The S100 proteins are a family of calcium-binding proteins found in the central and peripheral nervous systems of vertebrates. S100beta, the most abundant member of this family in the CNS, mediates calcium signal transduction, and shows neurotrophic, gliotrophic and mitogenic actions that influence the development and maintenance of the nervous system. Another member of the S100 family (S100A10) was found to modulate phospholipid turnover by inhibiting the activity of enzyme phospholipase A2 (PLA2). We determined the concentration of S100beta protein in the plasma of 23 medicated schizophrenic patients and 23 healthy controls. S100beta protein accounts for 96% of the total S100 in the brain. Schizophrenic patients showed reduced S100beta concentrations (p=0.003), and this finding was not related to clinical variables or to intake of antipsychotic medication. Decreased S100beta could be related to the findings of increased PLA2 activity and to brain maldevelopment in schizophrenia. These results are discussed further with respect to the role of adenosine in S100beta release. PMID- 10858628 TI - Reduced anterior cingulate gyrus volume correlates with executive dysfunction in men with first-episode schizophrenia. AB - Although frontal lobe structural and functional abnormalities have been identified in schizophrenia, their relationship remains elusive. Because the frontal lobes are both structurally and functionally heterogeneous, it is possible that some measures of frontal lobe structure may not have accurately identified relevant frontal lobe subregions. The authors hypothesized that the volumes of two dorsal, 'archicortical' subregions (i.e. superior frontal gyrus and anterior cingulate gyrus), but not a ventral, 'paleocortical' subregion (i.e. orbital frontal region) would be significantly and selectively correlated with executive and motor dysfunction in patients with schizophrenia as previously reported for the anterior hippocampal region. Volumes of these frontal lobe subregions were measured from magnetic resonance images based on sulcal anatomy in 20 men and 15 women with first-episode schizophrenia. All patients completed a comprehensive neuropsychological test battery while clinically stabilized that encompassed six domains of functioning: attention, executive, motor, visuospatial, memory and language. Findings indicated that reduced anterior cingulate gyrus volume was significantly correlated with worse executive functioning in men; among women, there were no significant correlations. Among men, anterior cingulate gyrus volume was significantly more strongly correlated with executive functioning than with attention, visuospatial, memory, language and general intellectual functioning. Neither executive nor motor functioning was significantly more strongly correlated with the dorsal 'archicortical' volumes than with orbital frontal volume. These findings suggest a link between executive deficits and dysfunction of the dorsal 'archicortical' system and implicate sex differences in their relationship in first-episode schizophrenia. PMID- 10858629 TI - The specificity of neurological signs in schizophrenia: a review. AB - This review examines the extent to which neurological signs are more prevalent in schizophrenia patients, compared to mood-disorder patients and healthy subjects, and whether there is a pattern in any of the differences that may be found. We included 17 studies and calculated the weighted mean prevalence of 30 neurological signs. The prevalence of most signs appears to be significantly different between schizophrenia patients and normal controls, but there are fewer differences between schizophrenia and mood-disorder patients. Several signs - poor stereognosis and rhythm tapping - are even more prevalent in mood-disorder patients than in schizophrenia patients. Only lack of extinction, dysdiadochokinesia, poor tandem walk, finger-thumb-opposition and articulation are significantly more prevalent in schizophrenia compared to mood-disorder patients. Impaired motor coordination seems most specific to schizophrenia. The discriminating power of motor sequencing still needs to be studied. So far, there is no evidence of a clearly interpretable pattern of neurological signs distinguishing schizophrenia patients from mood-disorder patients. PMID- 10858630 TI - Obstetric factors, urbanization and psychosis. AB - BACKGROUND: Epidemiologic evidence as early as the 1930s has suggested urbanization is linked to schizophrenia, either by place of admission, place of upbringing, or, more recently, place of birth. In the past decade, obstetric complications have been implicated in the etiology of schizophrenia. METHODS: With appropriate protections for anonymity, the files of the Danish Medical Birth Register were linked with the files of the Danish Psychiatric Case Register. The linkage produced 132 cases of schizophrenia and 69 cases of affective psychosis, who were born in 1973 or later, who entered a Danish psychiatric hospital before 1994. Controls were drawn from a 10% sample of the Medical Birth Register. Analysis was by logistic regression. RESULTS: The risk of hospitalization for schizophrenia was 4.20 times higher (95% CI=2.4-7.4) for those born in Copenhagen versus those born in rural areas of Denmark, and a linear relationship was demonstrated between urbanization of birthplace and risk. There was no difference in risk of hospitalization for affective psychosis for those born in Copenhagen versus rural areas. Obstetric complications had a moderate sized relationship to schizophrenia, but the relationship of urban birth to schizophrenia was unaffected by adjustment for obstetric complications. CONCLUSION: Urban birth is a strong risk factor for schizophrenia, not mediated by obstetric complications, which deserves further exploration. PMID- 10858631 TI - Developmental instability and schizotypy. AB - INTRODUCTION: It has been suggested that evidence of developmental disturbance of cognition and lateralisation in schizophrenia can be best understood from the perspective of developmental stability (DS), an indicator of the extent to which an individual develops according to a specified ontogenic programme in the presence of environmental noise. Higher levels of fluctuating asymmetry (FA; the difference between right and left side of a quantitative morphological trait such as dermatoglyphics) are thought to reflect less DS. We examined this issue for dimensions of schizotypy. METHODS: Associations between FA, measures of laterality and cognitive function on the one hand, and negative and positive dimensions of schizotypy on the other, were examined in a sample of 260 healthy adolescents aged 11.9-15.6years. FA was measured as a-b ridge count right-left differences. Neuropsychological measures yielded a general cognitive ability score and a frontal function score. Laterality was assessed with the Annett scale. RESULTS: Measures of psychosis proneness were normally distributed. Negative schizotypy was associated with more FA and lower general cognitive ability in a dose-response fashion. The association with FA was more apparent in boys. No associations existed with laterality or frontal function. CONCLUSION: The negative dimension of schizotypy may be associated with early developmental instability, resembling the pattern seen in the negative symptom dimension of schizophrenia. Measures of fluctuating asymmetry may be more sensitive with regard to the schizotypy phenotype than measures of laterality. PMID- 10858632 TI - Comparative evaluation of conventional and novel antipsychotic drugs with reference to their subjective tolerability, side-effect profile and impact on quality of life. AB - This study compared the effectiveness of conventional and novel antipsychotic drugs from a patient's perspective. Five comparable groups of schizophrenic patients (n=230) clinically stabilized on conventional antipsychotic drugs, risperidone, olanzepine, quetiapine or clozapine for a period of 6months or longer were cross-sectionally evaluated. Patients' clinical symptom profile, subjective responses and attitudes toward drugs, prevalence of dysphoria, akathisia, abnormal involuntary movements and Parkinsonian symptoms, and quality of life were ascertained using standardized rating scales. Between-group differences were examined with analysis of variance and chi-square tests. Patients receiving novel antipsychotic drugs experienced fewer side-effects, reported positive subjective responses and favourable attitudes toward their treatment, and revealed a lower prevalence of neuroleptic dysphoria. The differences were statistically significant (p<0.05) with the risperidone, olanzepine and quetiapine groups. Self-rated quality of life, measured with the sickness impact profile, was also significantly better among patients receiving novel antipsychotic drugs. These perceived benefits, however, were not reflected in the clinician rated (objective) measures of psychosocial functioning and quality of life. These findings substantiate the general notion that novel antipsychotic medications are uniformly better tolerated as indicated by the measures of subjective responses, side-effects and self rated quality of life. PMID- 10858633 TI - Hopelessness and its impact on rehabilitation outcome in schizophrenia -an exploratory study. AB - The primary focus in contemporary psychiatry on symptoms and their neurobiological basis, although fundamentally important, is nevertheless incomplete. The long-term course and outcome of schizophrenia are determined not only by the disorder, but also by the interaction between the person and the disorder. Not only psychopathological symptoms but also cognitive variables such as negative self-concepts, low expectations and external loci of control can influence the patient's coping strategies and may lead to hopelessness and chronicity. Hopelessness here refers to a cognitive-affective state in which the patient perceives the disorder and its consequences to be beyond his control, feels helpless and has given up expecting to influence its course positively, thereby abandoning responsibility and active coping strategies. In a prospective study, we examined these relationships by using logistic regression in data from 46 schizophrenic outpatients who were participating in a vocational rehabilitation program. Negative self-concepts, external loci of control, and depression correlated to a higher extent with depressive-resigned coping strategies than did schizophrenic symptoms. Thus, poor rehabilitation outcome may be predicted to a high degree by the presence of external loci of control, pessimistic outcome expectancies, negative symptoms, and depressive-resigned coping strategies. After having eliminated the influence of negative symptoms, external control beliefs still had significant predictive value for the outcome. Rehabilitation outcome in schizophrenic patients can be only partially predicted by negative symptoms; the other predictive factor is whether the patient has already given up or not. PMID- 10858634 TI - Striatal D(2) dopamine receptor density and psychotic symptoms in schizophrenia: a longitudinal study. PMID- 10858636 TI - Community provision of hearing aids and related audiology services. PMID- 10858637 TI - Costs and benefits of community postnatal support workers: a randomised controlled trial. AB - OBJECTIVES: This study aimed to measure the effect and the total cost per woman of providing postnatal support at home, based on a Dutch model. The research hypothesis was furnished by some existing evidence that postnatal support could reduce the risk of postnatal depression and encourage breastfeeding. DESIGN: The randomised controlled trial aimed to measure differences in health status in a group of women who were offered postnatal support from a community midwifery support worker (SW) compared with a control group of women who were not offered this support. Women were followed-up by postal questionnaire at 6 weeks and 6 months postnatally. SETTING AND SUBJECTS: All women who delivered a baby at the recruiting hospital were eligible to take part in the trial if they lived within the study area, were aged 17 years or over, and could understand English. INTERVENTION: The intervention consisted of the SW offering practical and emotional support and to help women rest and recover after childbirth. The SW offered ten visits in the first 28 days postnatally, for up to 3 hours per day. The SW's activities included housework, talking with the mother, and care for the baby or other siblings. The service was provided in addition to routine visits by the community midwife. MAIN OUTCOME MEASURES: The primary outcome was the general health perception domain of the Short Form-36 at 6 weeks. Secondary outcomes were mean Edinburgh Postnatal Depression Scale (EPDS), Duke Functional Social Support (DUFSS) scores and breastfeeding rates. RESULTS: The 623 randomised women were well-matched by group with a good response to follow-up. At 6 weeks there was no evidence of a significant difference between the two groups for the primary outcome. There was a non-significant trend for the control group to have better mean DUFSS and EPDS scores at 6 weeks. Breastfeeding rates were not significantly different at follow-up. At 6 months, both groups had similar health status. Satisfaction with the service was higher than for all other services received. The incremental cost of introducing the service comprised setting up and running the service. There were no differences between the groups in other resource use (general practitioner contacts, hospital services, prescriptions or medicines bought for mothers and babies) to 6-month follow-up. The total mean NHS cost to 6 month follow-up for the intervention group was pound180 per woman greater than for the control group (confidence interval, pound79.60, pound272.40). CONCLUSIONS: Although women valued the service, there was no evidence of any health benefit at the 6-week or 6-month follow-up, no difference in use of NHS services, and the additional cost of the service provision would be around pound 180 per woman. PMID- 10858638 TI - Ten thousand years of head lice infection. PMID- 10858639 TI - Trypanosome evolution under the microscope. PMID- 10858640 TI - Can roll back malaria achieve its goal? A challenge. PMID- 10858643 TI - Good worms or bad worms: do worm infections affect the epidemiological patterns of other diseases? PMID- 10858644 TI - Horse racing, sesame seeds and hyenas on the Web PMID- 10858645 TI - The topoisomerases of protozoan parasites. AB - Protozoan parasites are responsible for a wide range of debilitating and fatal diseases that are proving notoriously difficult to treat. Many of the standard chemotherapies in use today are expensive, have toxic side effects and, in some cases have marginal efficacy because of the emergence of drug-resistant parasites. In the search for more effective treatments, protozoan topoisomerases are now being considered as potential drug targets, building on the clinical success of anticancer and antibacterial agents that target human and bacterial topoisomerases. In this review, Sandra Cheesman explores progress in this relatively new but potentially important field of research. PMID- 10858646 TI - Functional diversity in the trans-sialidase and mucin families in Trypanosoma cruzi. AB - Trypanosomes are unable to synthesize the monosaccharide sialic acid, but some African trypanosomes and the American Trypanosoma cruzi can incorporate sialic acid derived from the host. To do so, T. cruzi expresses a trans-sialidase, an enzyme that catalyzes the transfer of sialic acid from host glycoconjugates to mucin-like molecules located on the parasite surface membrane. The importance of the process is indicated by the fact that T. cruzi has hundreds of genes encoding trans-sialidase, trans-sialidase-like proteins and mucin core proteins. Sequence divergence of members of these families has resulted in some molecules having functions unrelated to the acquisition of sialic acid. In this article, Alberto Frasch reviews the structure and possible function of the proteins making up these families. PMID- 10858647 TI - Cryptosporidium systematics and implications for public health. AB - There is controversy in the taxonomy of Cryptosporidium parasites and the public health significance of Cryptosporidium isolates from various animals. Recent advances in molecular characterization of Cryptosporidium parasites have allowed the re-examination of species structure of the genus Cryptosporidium. Non-parvum Cryptosporidium spp and new C. parvum genotypes in immunocompromised humans can now be clearly detected. In this article, Lihua Xiao and colleagues summarize the current biological and molecular evidence for different Cryptosporidium spp, and the public health importance of these species and new C. parvum genotypes. PMID- 10858648 TI - Leishmania-host-cell interaction: complexities and alternative views. AB - Leishmania are protozoan parasites that infect various mammalian species, including humans. It is generally thought that random attachment of the flagellated promastigotes to mononuclear phagocytes initiates their uptake via circumferential pseudopods. Intracellularly, the promastigotes become located in phagolysosomes in which they transform to and survive as 'aflagellated' amastigotes that hide their shortened flagellum within the flagellar pocket. Unrestricted replication of these amastigotes is assumed to cause the eventual burst of the host cell, thereby releasing the infectious parasites. Here, Mike Rittig and Christian Bogdan review a large body of literature containing potentially important but poorly appreciated findings, which together with recent results, argue for Leishmania-host-cell interactions that are much more complex than generally thought. PMID- 10858649 TI - Effectiveness of annual ivermectin treatment for Wuchereria bancrofti infection. AB - Using estimates for the anthelmintic efficacy of a single dose of ivermectin in the treatment of lymphatic filariasis patients, Anton Plaisier, Wilma Stolk, Gerrit van Oortmarssen and Dik Habbema here present and discuss model predictions of the impact of a five-year programme of annual community treatment on the intensity of infection. They show that the effectiveness of such programmes in terms of reductions in the microfilarial density depends critically on the treatment coverage and the pattern of attendance at repeated mass administrations. Improving these factors will possibly be more important than improving the efficacy of ivermectin by increasing its dosage or by adding other drugs. PMID- 10858650 TI - Towards an atlas of human helminth infection in sub-Saharan Africa: the use of geographical information systems (GIS). AB - The value of a geographical perspective to infectious disease epidemiology and control has long been recognized. However, the labour required to produce maps, and keep them up to date, has inhibited the development of this area, and very little is currently known about the spatial distribution of parasitic infections other than malaria, trypanosomiasis and onchocerciasis. A recent initiative by an international group of collaborators is attempting to redress the absence of detailed spatial information on the major helminth infections of humans. In this article, Simon Brooker and colleagues describe progress made by this initiative in mapping helminth infections in sub-Saharan Africa, highlighting the value as well as the limitations of this empirical mapping approach. PMID- 10858651 TI - Microsatellite markers and genetic mapping in Plasmodium falciparum. AB - Whole-genome methods are changing the scope of biological questions that can be addressed in malaria research. In the rich context provided by Plasmodium falciparum genome sequencing, genetic mapping is a powerful tool for identifying genes involved in parasite development, invasion, transmission and drug resistance. The recent development of a high-resolution P. falciparum linkage map consisting of hundreds of microsatellite markers will facilitate an integrated genomic approach to understanding the relationship between genetic variations and biological phenotypes. Here, Michael Ferdig and Xin-zhuan Su provide an overview for applying microsatellite markers and genetic maps to gene mapping, parasite typing and studies of parasite population changes. PMID- 10858652 TI - Good worms or bad worms: do worm infections affect the epidemiological patterns of other diseases? PMID- 10858653 TI - Innate immune response: friend and foe. PMID- 10858654 TI - The Wnt/Ca2+ pathway: a new vertebrate Wnt signaling pathway takes shape. AB - Members of the vertebrate Wnt family have been subdivided into two functional classes according to their biological activities. Some Wnts signal through the canonical Wnt-1/wingless pathway by stabilizing cytoplasmic beta-catenin. By contrast other Wnts stimulate intracellular Ca2+ release and activate two kinases, CamKII and PKC, in a G-protein-dependent manner. Moreover, putative Wnt receptors belonging to the Frizzled gene family have been identified that preferentially couple to the two prospective pathways in the absence of ectopic Wnt ligand and that might account for the signaling specificity of the Wnt pathways. As Ca2+ release was the first described feature of the noncanonical pathway, and as Ca2+ probably plays a key role in the activation of CamKII and PKC, we have named this Wnt pathway the Wnt/Ca2+ pathway. PMID- 10858655 TI - Mouse as the measure of man? AB - The humble house mouse's cohabitation with humans has been noted since the birth of agriculture, about 10 000 years ago, in the fertile flood plains of the Middle East. In recent times, however, the mouse has been elevated from pest to model for the study of human health and disease. Recent genomics and genetics initiatives will ensure the continued growth of the house mouse as a disease model. PMID- 10858656 TI - tRNA gene number and codon usage in the C. elegans genome are co-adapted for optimal translation of highly expressed genes. PMID- 10858657 TI - Unified nomenclature for the COP9 signalosome and its subunits: an essential regulator of development PMID- 10858658 TI - How low can toll go? PMID- 10858659 TI - Adjusting the focus on human variation. AB - Studies of nuclear sequence variation are accumulating, such that we can expect a good description of the structure of human variation across populations and genomic regions in the near future. This description will help to elucidate the evolutionary forces that shape patterns of variability. Such an understanding will be of general biological interest, but could also facilitate the design and interpretation of disease-mapping studies. Here, we integrate the results from surveys of nuclear sequence variation. When nuclear sequences are considered together with mtDNA and microsatellites, it becomes clear that neither the standard neutral model, nor a simple long-term exponential growth model, can account for all the available human variation data. A possible explanation is that a subset of loci are not evolving neutrally; even so, more-complex models of effective population size and structure might be necessary to explain the data. PMID- 10858660 TI - The EGF-CFC gene family in vertebrate development. AB - EGF-CFC genes encode extracellular proteins that play key roles in intercellular signaling pathways during vertebrate embryogenesis. Mutations in zebrafish and mouse EGF-CFC genes lead to defects in germ-layer formation, anterior-posterior axis orientation and left-right axis specification. In addition, members of the EGF-CFC family have been implicated in carcinogenesis. Although formerly regarded as signaling molecules that are distant relatives of epidermal growth factor (EGF), recent findings indicate that EGF-CFC proteins act as essential cofactors for Nodal, a member of the transforming growth factor beta (TGF-beta) family. Here, we review molecular genetic evidence from mouse and zebrafish on biological and biochemical roles of the EGF-CFC family, and discuss differing models for EGF CFC protein function. PMID- 10858661 TI - Developmental regulation of eukaryotic gene loci: which cis-regulatory information is required? AB - It is becoming increasingly accepted that gene loci comprise an extensive cis regulatory system that encodes different layers of regulatory information, all of which are necessary to achieve and maintain tissue-specific gene expression in ontogeny. To gain a detailed understanding of developmental processes, it is clearly necessary to unravel the molecular basis behind the different regulatory processes that control gene expression. This information is also of utmost importance for any practical application that uses gene transfer technology. PMID- 10858662 TI - Organellar genes: why do they end up in the nucleus? AB - Many mitochondrial and plastid proteins are derived from their bacterial endosymbiotic ancestors, but their genes now reside on nuclear chromosomes instead of remaining within the organelle. To become an active nuclear gene and return to the organelle as a functional protein, an organellar gene must first be assimilated into the nuclear genome. The gene must then be transcribed and acquire a transit sequence for targeting the protein back to the organelle. On reaching the organelle, the protein must be properly folded and modified, and in many cases assembled in an orderly manner into a larger protein complex. Finally, the nuclear copy must be properly regulated to achieve a fitness level comparable with the organellar gene. Given the complexity in establishing a nuclear copy, why do organellar genes end up in the nucleus? Recent data suggest that these genes are worse off than their nuclear and free-living counterparts because of a reduction in the efficiency of natural selection, but do these population-genetic processes drive the movement of genes to the nucleus? We are now at a stage where we can begin to discriminate between competing hypotheses using a combination of experimental, natural population, bioinformatic and theoretical approaches. PMID- 10858663 TI - GHOST: a gene homology online search tool. PMID- 10858664 TI - Maitotoxin stimulates Cd influx in Madin-Darby kidney cells by activating Ca permeable cation channels. AB - This study examined the role of calcium channels for the uptake of cadmium (Cd) into Madin-Darby canine kidney (MDCK) cells. Maitotoxin, an activator of different types of calcium channels, increased accumulation of 109Cd and 45Ca in MDCK cells. We found that maitotoxin increased accumulation by stimulating 109Cd influx because it did not affect efflux. An inhibitor of store-operated Ca channels, SKF96365, partially blocked 45Ca influx but did not affect 109Cd influx. Ni and Mn, and loperamide and proadifen (SKF 525a), inhibited 45Ca and 109Cd influx in cells stimulated with maitotoxin, but La and nifedipine did not. Overnight treatment with phorbol 12, 13-ibutyrate (PDBu) to activate protein kinase C resulted in a decrease in the concentration of maitotoxin needed to stimulate 45Ca and 109Cd influx. The effect of PDBu was blocked by treating cells with the protein kinase C inhibitor GF109203X. Additionally, the effect of PDBu was lost in cells treated with an inhibitor of RNA synthesis actinomycin D. These results suggest that a Ca permeable cation channel different from voltage dependent and store-operated Ca channels mediates the uptake of Cd in MDCK cells. The expression of this channel is regulated by protein kinase C. PMID- 10858665 TI - Modes of propagation of Ca(2+)-induced Ca2+ release in bullfrog sympathetic ganglion cells. AB - How depolarization-induced Ca2+ entry or caffeine activates Ca(2+)-induced Ca2+ release (CICR) in the cytoplasm and nucleoplasm was studied by recording intracellular Ca2+ ([Ca2+]i) with a confocal microscope in cultured bullfrog sympathetic ganglion cells. The amplitude and propagation speed of voltage pulse induced rises in [Ca2+]i were greater in the submembrane (< 5 microns depth) region than in the core region, and delayed and smaller, but significant, in the nucleus. Ryanodine and dantrolene reduced the rises in [Ca2+]i in both the cytoplasm and nucleus. A rapid application of high K+ solution induced global rises in [Ca2+]i in both the cytoplasm and nucleoplasm, which were decreased by dantrolene. Caffeine produced a slow, small rise in [Ca2+]i which grew into a global, regenerative rise both in the cytoplasm and nucleoplasm with some inward gradient in the cytoplasm. Each of the high [Ca2+]i phases during caffeine induced [Ca2+]i oscillation began in the submembrane region, while low [Ca2+]i phases started in the core region. These results suggest that CICR activated by Ca2+ entry or caffeine occurs predominantly in the submembrane region causing an inwardly spreading Ca2+ wave or [Ca2+]i oscillations, and that the nuclear envelope can cause CICR in the nucleoplasm, which is delayed due to Ca2+ diffusion barrier at the nuclear pores. PMID- 10858666 TI - Activation of purinergic P2X receptors inhibits P2Y-mediated Ca2+ influx in human microglia. AB - Purinoceptor (P2X and P2Y) mediated Ca2+ signaling in cultured human microglia was studied using Ca2+ sensitive fluorescence microscopy. ATP (at 100 microM) induced a transient increase in [Ca2+]i in both normal and Ca(2+)-free solution suggesting a primary contribution by release from intracellular stores. This conclusion was further supported by the failure of ATP to cause a divalent cationic influx in Mn2+ quenching experiments. However, when fluorescence quenching was repeated after removal of extracellular Na+, ATP induced a large influx of Mn2+, indicating that inward Na+ current through a non-selective P2X coupled channel may normally suppress divalent cation influx. Inhibition of Mn2+ entry was also found when microglia were depolarized using elevated external K+ in Na(+)-free solutions. The possibility of P2X inhibition of Ca2+ influx was then investigated by minimizing P2X contributions of purinergic responses using either the specific P2Y agonist, ADP-beta-S in the absence of ATP or using ATP combined with PPADS, a specific inhibitor of P2X receptors. In quenching studies both procedures resulted in large increases in Mn2+ influx in contrast to the lack of effect observed with ATP. In addition, perfusion of either ATP plus PPADS or ADP-beta-S alone caused a significantly enhanced duration (about 200%) of the [Ca2+]i response relative to that induced by ATP. These results show that depolarization induced by P2X-mediated Na+ influx inhibits store-operated Ca2+ entry resulting from P2Y activation, thereby modulating purinergic signaling in human microglia. PMID- 10858667 TI - Metabolic, cationic and secretory response to D-glucose in depolarized and Ca(2+) deprived rat islets exposed to diazoxide. AB - D-glucose stimulates insulin release from islets exposed to both diazoxide, to activate ATP-responsive K+ channels, and a high concentration of K+, to cause depolarization of the B-cell plasma membrane. Under these conditions, the insulinotropic action of D-glucose is claimed to occur despite unaltered cytosolic Ca2+ concentration, but no information is so far available on the changes in Ca2+ fluxes possibly caused by the hexose. In the present experiments, we investigated the effect of D-glucose upon 45Ca efflux from islets exposed to both diazoxide and high K+ concentrations. In the presence of diazoxide and at normal extracellular Ca2+ concentration, D-glucose (16.7 mmol/l) inhibited insulin release at 5 mmol/l K+, but stimulated insulin release of 90 mmol/l K+. In both cases, the hexose inhibited 45Ca outflow. In the presence of diazoxide, but absence of Ca2+, D-glucose (8.3 to 25.0 mmol/l) first caused a rapid decrease in insulin output followed by a progressive increase in secretory rate. This phenomenon was observed both at 5 mmol/l or higher concentrations (30, 60 and 90 mmol/l) of extracellular K+. It coincided with a monophasic decrease in 45Ca efflux and either a transient (at 5 mmol/l K+) or sustained (at 90 mmol/l K+) decrease in overall cytosolic Ca2+ concentration. The decrease in 45Ca efflux could be due to inhibition of Na(+)-Ca2+ countertransport with resulting localized Ca2+ accumulation in the cell web of insulin-producing cells. A comparable process may be involved in the secretory response to D-glucose in islets exposed to diazoxide and a high concentration of K+ in the presence of extracellular Ca2+. PMID- 10858668 TI - Calcium uptake, release and ryanodine binding in melanosomes from retinal pigment epithelium. AB - High levels of calcium have been reported in pigmented tissues of the vertebrate eye, such as retinal pigment epithelium (RPE). Melanin granules also have high calcium concentrations, suggesting that melanin granules may be a calcium reservoir. Here we characterized the uptake and release of calcium in a pure melanosomal fraction obtained from frog RPE. Melanosomes take up 45Ca by a saturable system with an apparent KM of 0.5 mM. About 40% of 45Ca accumulation was insensitive to low temperature. 45Ca uptake was not affected by verapamil, nifedipine, dantrolene, vanadate, thapsigargin or cyclopiazonic acid, but it was reduced by 50% by ruthenium red, and increased by the ionophore A23187 and nigericin. Release of 45Ca-loaded was stimulated by caffeine and inositol 1,4,5 trisphosphate (IP3). Caffeine stimulated release of calcium was blocked by either ryanodine or ruthenium red, but calcium released by IP3 was not affected by heparin. No binding of 3H-IP3 was observed. The 3H-ryanodine binding sites exhibited a KB of 1.3 nM and a Bmax of 12.1 fmol/mg protein. Thus, our results suggest that melanosomes may function as intracellular organelles that regulate calcium concentration in RPE. PMID- 10858669 TI - Expression of multiple plasma membrane Ca(2+)-ATPases in rat pancreatic islet cells. AB - When stimulated by glucose, the pancreatic beta-cell displays large oscillations of intracellular free Ca2+ concentration ([Ca2+]i). To control [Ca2+]i, the beta cell must be equipped with potent mechanisms for Ca2+ extrusion. We studied the expression of the plasma membrane Ca(2+)-ATPases (PMCA) in three insulin secreting preparations (a pure beta-cell preparation, RINm5F cells and pancreatic islet cells), using reverse-transcribed PCR, RNase protection assay and Western blotting. The four main isoforms, PMCA1, PMCA2, PMCA3 and PMCA4 were expressed in the three preparations. Six alternative splice mRNA variants, characterized at splice sites A, B and C were detected in the three preparations (rPMCA1xb, 2yb, 2wb, 3za, 3zc, 4xb), plus two additional variants in pancreatic islet cells (PMCA4za, 1xkb). The latter variant corresponded to a novel variant of rat PMCA1 gene lacking the exon coding for the 10th transmembrane segment, at splice site B. At the mRNA and protein level, five variants predominated (1xb, 2wb, 3za, 3zc, 4xb), whilst one additional isoform (4za), predominated at the protein level only. This provides the first evidence for the presence of PMCA2 and PMCA3 isoforms at the protein level in non-neuronal tissue. Hence, the pancreatic beta cell is equipped with multiple PMCA isoforms with possible differential regulation, providing a full range of PMCAs for [Ca2+]i regulation. PMID- 10858670 TI - Medical errors--a costs burden on society. PMID- 10858671 TI - NHS litigation. PMID- 10858672 TI - The natural history of breast cancer--can anything be done about it? PMID- 10858673 TI - Damage awards increase. PMID- 10858674 TI - Thyroid aspiration cytology in Newcastle: a six year cytology/histology correlation study. AB - BACKGROUND: Fine needle aspiration cytology (FNAC) is a well-established technique for pre-operative investigation of thyroid nodule(s). Thyroid FNAC was introduced in the teaching hospitals of Newcastle upon Tyne in 1981, initially with a small group of clinicians as aspirators. Audit results for 1981-1986 inclusive showed an unsatisfactory rate of 25.3% and prediction of malignancy with a sensitivity of 93.5%. FNAC has become more popular locally for the investigation of thyroid disease and the number of clinicians performing aspirates has increased. The results for recent years have, therefore, been audited. METHODS: Medical records were reviewed for 239 patients with a dominant thyroid nodule who had FNAC carried out in the 6 year period 1990-1995 and subsequent partial or complete thyroidectomy. RESULTS: Histology of thyroid specimens showed 60 follicular adenomas and 34 malignant lesions (including 19 papillary, 10 follicular and 3 medullary carcinomas, one lymphoma and one follicular neoplasm with indeterminate malignant potential). A total of 302 FNAC had been carried out on these 239 patients. On cytological grounds the unsatisfactory sample (AC0 and AC1) rate was 43.1% on initial aspiration which was reduced to 32.2% on repeated aspiration. FNAC predicted neoplasia (AC3, AC4 and AC5) with a sensitivity of 86.8%, a specificity of 67.0%, a negative predictive value of 87.5% and a positive predictive value of 65.5%. Malignancy was predicted by FNAC (AC3, AC4 and AC5) with a sensitivity of 88.9%. A FNAC report of AC5 had a positive predictive value for malignancy of 100%. CONCLUSIONS: FNAC is an invaluable and minimally invasive procedure for the pre operative assessment of patients with a dominant thyroid nodule. It is, however, important that the number of aspirators and cytopathologists be kept small to maintain expertise and also that the results of FNAC be subjected to ongoing audit. PMID- 10858675 TI - Medullary thyroid carcinoma in Northern Ireland, 1967-1997. AB - BACKGROUND: The experience of managing medullary thyroid carcinoma (MTC) in a specialist endocrine surgery unit was reviewed. METHODS: The case records of 38 patients (19 male, 19 female) treated over a 30 year period were studied. RESULTS: There were 23 (60.5%) patients with sporadic MTC while the remainder had familial MTC--12 multiple endocrine neoplasia (MEN) type 2A, two MEN type 2B, one non-MEN familial medullary thyroid carcinoma (FMTC). Sporadic MTC patients were significantly older at presentation (median 56 years, interquartile range 41.5 61.3 years) compared to MEN 2A patients (median 26 years interquartile range 17.5 34 years) and had more advanced stage of disease. Survival of MTC patients was significantly worse in sporadic disease than in those with MEN 2A (P < 0.0001). All familial cases had bilateral multifocal tumour whereas in sporadic patients only unilateral disease was seen. The availability of genetic testing now allows early identification of affected members of familial MTC kindreds. This has led to total thyroidectomy being performed on the basis of positive genetic screening alone in three patients (two MEN 2A, one FMTC), in all of whom widespread C-cell hyperplasia and microscopic multifocal invasive MTC were identified histologically. CONCLUSIONS: The management of MTC has changed during the study period with total thyroidectomy recommended as the primary procedure of choice for all patients. In the familial setting, positive genetic testing now allows thyroidectomy to be performed at an early pre-clinical stage, with the hope of permanent cure. PMID- 10858676 TI - Management of isolated sternal fractures: determining the risk of blunt cardiac injury. AB - A review of the management of isolated sternal fractures in a regional cardiothoracic unit reveals that, in a 2 year period, 37 consecutive patients were admitted for observation and further investigation, including echocardiography and cardiac enzyme measurements to exclude blunt cardiac injury. Minor blunt cardiac injury was detected in only one patient, and was associated with an acutely abnormal electrocardiogram (ECG). ECG showed acute changes in 8 further patients, whilst 3 patients had an abnormal chest X-ray (CXR) due to widening of the mediastinum (1 patient had abnormal CXR and ECG), but none had evidence of cardiac injury. CXR and ECG were both normal in 23 patients, and were predictive of the absence of significant complications. A survey of 22 other cardiothoracic units around the UK confirms that the management of patients with isolated sternal fractures varies considerably from hospital to hospital. As suggested by previous reports, we believe that patients, who are otherwise fit and have normal ECG and CXR on presentation, can be safely discharged home on oral analgesics. The routine use of echocardiography and creatinine kinase (CK) assays in the assessment of isolated sternal fractures is not indicated. The introduction of these guidelines has resulted in a dramatic reduction in the number of patients admitted with isolated sternal fractures to our unit. PMID- 10858677 TI - 'Fast track' carotid duplex scanning in a district general hospital. AB - 'Fast track' carotid scanning is designed to rapidly identify patients with significant symptomatic carotid stenosis and, thereby, allow prompt surgery. We review the outcome of patients referred to our open-access scanning service over 3 years and 6 months. A total of 807 cases (62% males and 38% females with a mean age of 64 years) were referred. The main presenting symptoms were TIA in 69%, amaurosis fugax in 11% and minor CVA in 8.3%. The mean time between referral and scan was 17 days. In 80% of the cases, the scan showed no significant disease and the patients were not seen in the clinic. Significant abnormality (stenosis > 70% or occlusion) was found in 20% of the patients. Of the total, 12% were reviewed in the out-patient clinic following which no action was taken, 2% had angiography but no surgery, while 5% had angiography and surgery. 1% were lost to follow-up. The mean delay from scan to operation was 36 days. CONCLUSION: Fast track scanning has led to early detection of surgically relevant carotid lesions and avoidance of delay in surgical intervention. It is an efficient and cost effective practice. PMID- 10858678 TI - Clinical examination of varicose veins--a validation study. AB - The aim of this study was to determine the accuracy of clinical tests compared to colour duplex imaging in patients with primary varicose veins using a prospective, blinded comparison study. A total of 44 patients (70 limbs) with primary, previously untreated varicose veins presenting to the vascular laboratory of a university teaching hospital were studied. The patients underwent physical examination using the cough test, the tap test, Trendelenbergs' test, Perthes' test and hand-held Doppler (HHD) assessment prior to undergoing colour duplex scanning. Reflux was detected on duplex scanning, at the sapheno-femoral junction in 39/70 limbs (54%), the long saphenous vein in 47/70 limbs (64%) and the sapheno-popliteal junction in 9/70 limbs (13%). The cough test had low sensitivity (0.59) and specificity (0.67). The tap test had low sensitivity (0.18) and high specificity (0.92). The Trendelenberg test had high sensitivity (0.91) but low specificity (0.15). Perthes' test had a high sensitivity (0.97) but low specificity (0.20). Hand-held Doppler assessment of reflux at the sapheno femoral junction, in the long saphenous vein and at the sapheno-popliteal junction had high sensitivity (0.97, 0.82, and 0.80, respectively) and specificity (0.73, 0.92, and 0.90, respectively) of detecting reflux. Clinical tests used in the examination of patients with primary varicose veins are inaccurate. Assessment using hand-held Doppler is more accurate. Courses and clinical textbooks should be revised to replace these tests with instruction in how to use hand-held Doppler in the clinical examination of patients with varicose veins. PMID- 10858679 TI - A strategy for vascular services--testing the 600,000 population model. AB - BACKGROUND: Vascular services' delivery has been criticised, and re-organisation based on a 600,000 population model suggested. We assessed the feasibility of this model in three geographically disparate English regions. METHODS: Surgical arterial activity by Trust was analyzed using 1994/95 data from Hospital Episode Statistics. A postal survey of acute Trusts was used to assess vascular facilities and personnel. Distances between hospitals and enumeration districts were mapped using a Geographical Information System. MAIN OUTCOME MEASURES: Number (proportion) of Trusts performing over 100 arterial procedures a year. Number (proportion) of Trusts with a vascular on-call rota. Proportion of population likely to live more than 40 km away (equivalent to 1 h blue-light ambulance travel time) from a vascular unit under the proposed model. RESULTS: Twelve of the 32 Trusts (38%) performed over 100 arterial procedures annually; 23 Trusts completed the survey. Of these, five (22%) had a vascular on-call rota. Under the 600,000 model, in East Anglia a further 16.5% of the population would live > 40 km from a vascular unit. In Wessex, a further 0.4% of the population would live > 40 km from a vascular unit. Impact on access in North West Thames was negligible. CONCLUSIONS: A 600,000 population model could be feasible in urbanized regions, but not in geographically remote ones. PMID- 10858680 TI - Pseudoaneurysm of the medial inferior genicular artery following anterior cruciate ligament reconstruction. AB - Pseudoaneurysm is a rare complication of surgery or trauma around the knee. A 30 year-old man presented 10 days following anterior cruciate ligament repair with a 2 cm pulsatile swelling on the medial side of the knee. Angiography demonstrated a pseudoaneurysm of the medial inferior genicular artery. Surgical exploration and ligation of the feeding vessel to the aneurysm was performed and the patient made a full recovery. Vascular injury must be suspected in patients presenting with a haemarthrosis or pulsatile swelling following surgery on the knee. PMID- 10858681 TI - Vertical transtalar Steinmann pin fixation for unstable ankle fractures. AB - Ankle fractures are common injuries and can usually be managed by either cast immobilisation or open reduction and internal fixation. Occasionally, both of these methods are contra-indicated. In such cases, one solution is the use of a vertical transtalar Steinmann pin to stabilise the fracture. Over the last 7 years, we have managed 8 patients with severe ankle fractures using vertical transtalar Steinmann pin fixation. The median age of these patients was 76 years (range, 35-94 years). This method was used in two patients with open contaminated wounds, in 5 with atrophic and blistered skin, and in one following failure of internal fixation in a patient with atrophic skin. In all patients, a satisfactory reduction was achieved and maintained after removal of the pin. Although all patients began to develop osteo-arthritic changes secondary to their original fractures, no complications were directly attributable to the use of the pin. In unstable ankle fractures where damage and contamination of soft tissues would preclude internal fixation, we recommend the use of vertical transtalar Steinmann pin fixation. PMID- 10858682 TI - Circumcision using bipolar diathermy scissors: a simple, safe and acceptable new technique. AB - BACKGROUND: Circumcision may be performed by a variety of techniques. Postoperative haemorrhage is a recognised complication of the procedure. METHOD: A method of circumcision using bipolar diathermy scissors is described. Foreskin vessels are coagulated as the foreskin is cut away. RESULTS: Thirty patients underwent this procedure without complication. CONCLUSION: Circumcision using bipolar scissors can be a safe and simple operation. PMID- 10858683 TI - A prospective evaluation of the combined use of the modified Alvarado score with selective laparoscopy in adult females in the management of suspected appendicitis. AB - BACKGROUND: A diagnostic scoring system such as the modified Alvarado score, combined with selective laparoscopy in adult females, can be used in the assessment of acute abdominal pain suggestive of appendicitis. METHOD: A total of 84 consecutive patients presenting to our surgical team with suspected appendicitis were assessed prospectively using the modified Alvarado score. The definitive management of this study group was instigated according to a set algorithm based on the score. The algorithm included the use of diagnostic laparoscopy in adult female patients with scores suggestive of appendicitis. A negative appendicectomy rate was obtained from those undergoing appendicectomy using this approach and compared to that obtained from the 97 patients that had undergone appendicectomy under the care of the other surgical teams in our unit during the study period. RESULTS: The rate of negative appendicectomy in the study group was 0% compared to 18% in the control group (P < 0.05); 10% of adult female women had negative diagnostic laparoscopies for appendicitis thus saving this group an unnecessary appendicectomy. This was achieved without an increase in total in-patient stay. CONCLUSION: An algorithm combining the modified Alvarado score with selective laparoscopy is recommended for widespread use in the management of suspected acute appendicitis. PMID- 10858684 TI - Multicystic peritoneal inclusion cysts: the use of CT guided drainage for symptom control. AB - Our case study is that of a teenage male presenting with multilocular peritoneal inclusion cystic disease that is now managed symptomatically with a minimally invasive, repeatable technique. Between admissions he leads a relatively normal life. Symptomatic control in MPIC is possible using repeated CT guided aspirations. PMID- 10858685 TI - Intestinal failure after surgery for complicated radiation enteritis. AB - Between 1983 and 1997, a total of 16 patients were referred to a tertiary Intestinal Failure Unit (IFU) following surgery elsewhere for complications of radiation enteritis. Eleven were female with a mean age of 43 years (range 21-71 years) and the most common primary site of malignancy was genitourinary (n = 13). Patients had undergone an average of two laparotomies (range 1-7 laparotomies) for complications of radiation enteritis prior to transfer to the IFU. On admission, the principal problem in eight patients was persisting intestinal fistulation, four patients had continuing intestinal obstruction and four had the short bowel syndrome after extensive intestinal resection. Only one patient had evidence of residual malignancy; this patient with short bowel syndrome was allowed home without invasive therapy. Of the remaining 15 patients, 12 required an abdominal surgical procedure, while three were discharged without further surgery after training for home parenteral nutrition (HPN). Following abdominal surgery, five patients died in hospital, but the remaining seven patients went home alive--including two further patients on HPN. Overall, of the 15 patients referred with intestinal failure after surgery for complications of radiation enteritis and actively treated, one-third died in hospital and a further third required institution of HPN before being able to be discharged home. PMID- 10858686 TI - Large bowel impaction by the BioEnterics Intragastric Balloon (BIB) necessitating surgical intervention. AB - A case of large bowel impaction caused by migration of a BioEnterics Intragastric Balloon (BIB) is presented. The literature is reviewed regarding both the use and the complications inherent in such balloon devices. This is the first reported case of an intragastric balloon impacted in the colon 9 months after insertion. PMID- 10858687 TI - Demographic factors associated with knowledge of colorectal cancer symptoms in a UK population-based survey. AB - Greater public awareness of colorectal cancer symptoms might result in earlier presentation with improved cure by available treatments, but little is known about the extent of public knowledge of colorectal cancer symptoms. We asked a sample of the general population about knowledge of colorectal cancer symptoms and assessed demographic characteristics associated with differences in knowledge. A population-based telephone enquiry into knowledge of colorectal cancer-associated symptoms was conducted by a commercial telephone market research company. Persons called were asked if they knew any symptom of colorectal cancer. Those who answered 'yes' were asked to state any symptom. The ability to state a colorectal cancer-associated symptom was correlated with demographic characteristics, and the strength of the relation between stating a colorectal cancer associated and unassociated symptom (discrimination) was also assessed. 1019 persons (respondents) from an age, sex, and social class-balanced general population sample were questioned. 328 (31%) of respondents stated a colorectal cancer-associated symptom. Regression analysis identified older age, female sex and higher social class as being significantly associated with ability to state a colorectal cancer-associated symptom. There was a weak trend (P = 0.054) towards a relationship between ability to state an associated symptom and stating fewer unassociated symptoms. These results reveal a total ignorance of colorectal cancer symptoms in the majority of respondents and suggest a need for improved public education about colorectal cancer. Public education should be directed at population groups with higher risk and the least knowledge of colorectal cancer. PMID- 10858688 TI - Improving the view in the rectal clinic: a randomised control trial. AB - BACKGROUND: Rigid sigmoidoscopy forms an integral part of the out-patient assessment of patients with colorectal symptoms. However, the value of this of this examination is often diminished by faecal loading of the rectum. This trial aimed to determine the ability of a single self-administered glycerine suppository to clear the rectum in preparation for rigid sigmoidoscopy, and considered patient acceptability of this practice. METHODS: Consecutive patients were randomly allocated to receive suppository or no suppository prior to out patient rigid sigmoidoscopy. Assessment was made of patient compliance, the effectiveness of rectal examination, and the depth to which the sigmoidoscope was inserted. RESULTS: 131 patients were randomised into suppository (n = 66) or control groups (n = 65). The number of patients deemed to have good views of the rectum (> 75% of rectal mucosa seen) was significantly greater in suppository than control groups (79% versus 26.2%, P < 0.05 Chi square test), whilst that of poor examinations (< 50% of rectal mucosa seen) was significantly greater in control than suppository groups (44.6% versus 4%, P < 0.05). The depth of insertion of the sigmoidoscope was significantly greater in those receiving suppositories (54.5% versus 21.5% undergoing evaluation to 18 cm or more, P < 0.05). Compliance amongst those who received suppositories was high with only 3 of 53 (4.5%) patients in the suppository group evaluated by questionnaire reporting difficulty or concerns over their use. CONCLUSION: Self-administered suppositories are acceptable to patients and significantly improve the efficiency of outpatient rigid sigmoidoscopy. Their usage should become routine. PMID- 10858689 TI - Consent to surgery in a high risk specialty: a prospective audit. AB - A prospective audit was performed to assess how well patients were being consented for neurosurgery. Sixty patients with various neurosurgical conditions were included in the study. Audit was performed firstly by means of a questionnaire to examine the type of information given to patients, and their understanding of such information. Secondly, the patient's medical notes were reviewed to analyse any written evidence by the consenting doctor for the consenting procedure. 100% of the patients felt that they had been informed satisfactorily about the nature of their condition and the nature of the operation. 92% understood the specific risks of their proposed operation. However, only 25% were informed about the general risks of surgery and anaesthesia. Only 33% felt that they were informed fully about alternative treatment options. 97% of the patients felt that they had reached an informed decision regarding surgery. 67% of the case notes contained information on the nature and specific risks of the operation, while information on general risks of surgery and anaesthesia was documented in only 17% of the case notes. 33% of the case notes contained no information for the consenting procedure. Our audit showed that the patients had a good understanding of the nature and aim of the operation and the specific risks. Areas that require improvement are explaining the general risks of surgery and alternative treatment. For the consenting doctor, there should be more documentation in the notes, and there should be mention of the doctor's satisfaction that the patient was deemed to be competent and had made an informed decision. PMID- 10858690 TI - The NHS patient information lottery: it is whom you see rather than what you need. AB - OBJECTIVE: To examine the current scale of provision of patient information materials by consultant surgeons in UK NHS and private sector hospitals. DESIGN: Secondary analysis of the responses of 12,555 surgical patients to surveys evaluating surgical services provided by specific consultants. SETTING: 7 NHS Trusts and one private sector hospital distributed throughout the UK. MAIN OUTCOME MEASURES: Provision of information materials by hospital, surgeon, and case-mix. Comparison of this service with patients' evaluations of surgeons' verbal communication in the outpatient clinic. RESULTS: Great variation exists between surgeons of the same specialty, and between hospital surgical directorates as a whole in the routine provision of supportive information materials to patients undergoing surgery. This variation cannot be explained solely by clinical need. Patients treated in private hospitals were less likely to receive information materials compared to patients treated within the NHS. CONCLUSIONS: Provision of printed information materials to patients by clinicians appears to be arbitrary. With the prospect of national performance frameworks in the foreseeable future, it is reasonable to assume that not only will the content of patient information be determined by quality standards but, in addition, its availability will be decided by clinical need rather than the clinician's preferences or interests. PMID- 10858691 TI - Nutrition and behaviour: the role of n-3 fatty acids in cognitive function. PMID- 10858692 TI - Validation of dietary intakes measured by diet history against 24 h urinary nitrogen excretion and energy expenditure measured by the doubly-labelled water method in middle-aged women. AB - A diet history method for estimating energy and N intakes was validated against 24 h urinary N excretion and energy expenditure measured by the doubly-labelled water (DLW) method. Forty-eight women aged 50-65 years were studied over 1 year. Weighed diet records from 4 d and two 24 h urine collections, for measurement of urinary N excretion, were obtained in each of four seasons. At the end of the year, a diet history was obtained, BMR was measured by whole-body calorimetry, and, in sixteen women, total energy expenditure (EE) was measured by DLW. Energy intake (EI) and N intake (NI) were calculated using food tables. Using weighed records and diet history respectively mean NI were 11.21 (SD 2.09) g and 11.47 (SD 2.40) g (NS) and EI were 8.08 (SD 1.54) MJ and 8.20 (SD 1.86) MJ (NS). Mean urine N:NI and EI:BMR values indicated bias to under-reporting by weighed record and diet history techniques in some individuals, but there was no significant difference between these measures at the group level. The Pearson correlation coefficient (r) for urine N v. NI was 0.81 for the weighed record and 0.38 for the diet history. The correlation of EE v. EI was r 0.48 for weighed record and r 0.11 for diet history. In this study the diet history gave the same estimate of mean intake, but the weighed record appeared to perform better in ranking individuals. PMID- 10858693 TI - Daily energy expenditure and its main components as measured by whole-body indirect calorimetry in athletic and non-athletic adolescents. AB - The objectives of the present study were to determine whether differences in usual physical activity affect BMR, sleeping energy expenditure (EE), and EE during seated activities between athletic and non-athletic adolescents, and to establish individual relationships between heart rate and EE. Adolescents (n 49, four groups of eleven to fifteen boys or girls aged 16-19 years) participated in the study. Body composition was measured by the skinfold-thickness method and maximum O2 consumption (VO2max) by a direct method (respiratory gas exchange) on a cycloergometer. The subjects each spent 36 h in one of two large whole-body calorimeters. They followed a standardized activity programme including two periods of exercise simulating their mean weekly physical activities. Fat-free mass (FFM), VO2max, daily EE and EE during sleep and seated activities were significantly higher in athletic than in non-athletic subjects of both sexes. VO2max, daily EE and EE during exercise adjusted for FFM were higher in athletic than in non-athletic adolescents (P < 0.001), whereas sleeping EE, BMR and EE during seated activities and adjusted for FFM were not significantly different between athletic and non-athletic adolescents. However, sex differences in EE remained significant. Thus, differences in EE between athletic and non-athletic adolescents resulted mainly from differences in FFM and physical exercise. Usual activity did not significantly affect energy utilization of substrates. Finally, individual relationships were computed between heart rate and EE with activity programmes simulating the usual activities of athletic and non-athletic adolescents with the goal of predicting EE of the same subjects in free-living conditions. PMID- 10858694 TI - Improved reporting of habitual food intake after confrontation with earlier results on food reporting. AB - The aim of the present study was to improve the reporting of food intake by confronting subjects with their way of reporting food intake, e.g. under recording and/or under-eating. To minimize portion size errors, eighteen female dietitians were recruited as subjects. Energy- and water intake were measured for 1 week with a weighed dietary record. Resting metabolic rate was measured with an open-circuit ventilated-hood indirect calorimeter, and physical activity was measured with an accelerometer for movement registration. Water loss was estimated with 2H-labelled water. Energy balance was checked for by measuring empty body-weight 1 week before the start, at the start and at the end of the recording week. In the first part of the study, the change in body weight in the non-recording week was 0.14 kg and in the recording week -0.45 kg (P 0.02), indicating 12% under-eating. Total water intake closely matched measured water loss, indicating a high recording precision. There was under-reporting of habitual food intake that could be fully explained by under-eating. In the second part of the study, subjects were confronted with these results and the protocol was repeated. This time there was no significant change in body weight in the recording week, indicating no under-eating. The reporting of habitual food intake had been improved. In conclusion, in the studied group of highly motivated lean women, under-reporting of habitual food intake (here due to under-eating) could be eliminated by confrontation with the results of this phenomenon. PMID- 10858695 TI - Study of the effects of dietary fish intake on serum lipids and lipoproteins in two populations with different dietary habits. AB - Increased concentrations of n-3 polyunsaturated fatty acids (PUFA), namely eicosapentaenoic acid (20:5; EPA) and docosahexaenoic acid (22:6; DHA), have been shown to be beneficial in coronary artery disease (CAD). In the present study, the relationships between fish intake and concentrations of serum EPA and DHA and the effects of these fatty acids on serum lipids and lipoproteins were investigated. Two groups of men, one living in a fishing village and the other in a farming village, participated in this study. The daily fish consumption was ten times greater in the fishing village group than in the rural village group and the mortality from IHD in the rural village was four times higher. Serum concentrations of EPA and DHA were significantly higher in the fishing village group (P < 0.001). In this group, the serum concentration of arachidonic acid (20:4; AA), was significantly lower (P < 0.001), and the ratio EPA:AA was twice that of the rural village (P < 0.001). Moreover, in the fishing village group, the serum triacylglycerol and total cholesterol levels were significantly lower than those observed in the rural village (P < 0.01 and P < 0.05 respectively). In the fishing village group the serum LDL-cholesterol concentration was also lower, although the difference was not significant. Our results reinforce the hypothesis that a high intake of n-3 PUFA provides protection against CAD. PMID- 10858696 TI - Phyto-oestrogen content of berries, and plasma concentrations and urinary excretion of enterolactone after a single strawberry-meal in human subjects. AB - Quantitative data on phyto-oestrogen, particularly lignan, content in edible plants are insufficient. We, therefore, measured isoflavonoids and lignans in nine edible berries using an isotope dilution gas chromatography-mass spectrometry method for foods and found substantial concentrations of the lignan secoisolariciresinol (1.39-37.18 mg/kg DM), low amounts of matairesinol (0-0.78 mg/kg DM) and no isoflavones. To determine pharmacokinetics and urinary excretion pattern of the mammalian lignan enterolactone derived from plant lignans, a study with human subjects was conducted. Five healthy women and two men consumed, after a 72 h period of a phyto-oestrogen-free regimen, a single strawberry-meal containing known amounts of plant lignans. Basal and post-meal blood and urine samples were collected at short intervals. The samples were analysed using time resolved fluoroimmunoassay of enterolactone. The meal increased plasma concentration of enterolactone after 8-24 h and in urine in the 13-24 h and 25-36 h urine collections. High individual variability of the metabolic response was observed. Enterolactone excreted in the urine collected throughout the 48 h post meal yielded on average 114% of the plant lignans consumed. It is concluded that berries containing relatively high concentrations of plant lignans contribute to plasma and urinary levels of mammalian enterolactone in human subjects. PMID- 10858697 TI - Compilation of a provisional UK database for the phylloquinone (vitamin K1) content of foods. AB - This paper reports the compilation of a food composition database for phylloquinone (vitamin K1) derived from the direct analysis of foods, recipe calculation and the assignment of values based on food similarities. All the basic and other food items used in these calculations had been analysed by HPLC and about 170 of the items had been obtained and assayed in the UK. Recipe calculations took account of the cooking method and changes in water and fat content. Currently, approximately 1501 food items with Royal Society of Chemistry/Ministry of Agriculture, Fisheries and Food food codes have been allocated a vitamin K1 value, and a further 282 new recipe codes are included in the database. Representative values from each food group are reported together with an indication of the potential variation. Detailed examples of some recipe calculations are included, and also the impact of changing the type of fat in recipes. Vitamin K1 is associated with, and most abundant in, photosynthetic tissues of plants. Accordingly, the highest concentrations (3000-6000 micrograms/kg) are found in dark-green leafy vegetables and herbs, such as kale, parsley, spinach and green cabbage. Intermediate concentrations (1000-2000 micrograms/kg) are found in plants with paler leaves such as white cabbage and lettuce or in green, non-leafy vegetables such as broccoli and brussel sprouts. Fats and oils contain variable amounts of vitamin K1 with the highest concentrations (300-1300 micrograms/kg) in soyabean, rapeseed and olive oils and the margarines based on them. Other foods such as dairy products, meat dishes and cereal-based foods (bread, biscuits, cakes, desserts etc.), although not in themselves particularly rich in vitamin K1 (< 200 micrograms/kg), may contribute significantly to intakes when consumption of green vegetables is poor. Within the scope of this present study, it has not been possible to address issues such as inter-sample variability, losses during storage or the bioavailability from different foods and further work on these aspects is needed. PMID- 10858698 TI - Transsulfuration, protein synthesis rate and follicle mRNA in the skin of young Merino lambs in response to infusions of methionine and serine. AB - Methionine (Met) is usually the first limiting amino acid for sheep and supplements of Met may increase production of wool and meat. The wool response may be due to an increased supply of cysteine (Cys) from transsulfuration (TS) of Met. Met is catabolized through homocysteine to form Cys when the S from Met is transferred to serine (Ser). We hypothesized that providing additional Met would create a deficiency of Ser and that by simultaneously providing Met and Ser, TS and wool growth could be increased more than by providing Met alone. The effects of i.v. infusions of Met and Ser to young Merino lambs on TS, fractional synthesis rate (FSR) of protein in skin, follicle mRNA and wool growth were examined. Following 4 d of constant i.v. infusion of 3 g Met/d, or 10 g Ser/d or both, the isotope tracers: L-[3-(13)C]Cys, L-[ring-d5]phenylalanine (Phe) and L [2,3,3-d3]Ser were infused over 8 h to allow for measurements of irreversible loss rate (ILR), and TS in whole body and skin. Skin biopsies were taken for measurement of FSR. Wool growth rate was measured using autoradiography. An infusion of Met significantly (P < 0.05) improved wool growth rate and increased skin FSR, Cys supply from TS and enhanced levels of follicle mRNA (from the K2.10 intermediate filament gene and three gene families encoding keratin associated proteins KAP1, KAP4 and KAP12). The extra Met lowered Ser ILR. The infusion of Ser doubled Ser ILR in the body and increased skin FSR calculated using the Cys tracer in plasma (P < 0.05). However, there were no significant (P > 0.05) changes in TS, skin FSR calculated using the Phe and Ser tracers, follicle mRNA or wool growth rate as a result of Ser infusion. While there were trends towards increased TS and FSR with Ser infusion, the overall lack of significant changes indicates a high capacity for the de novo synthesis of Ser. PMID- 10858699 TI - Cholesterol crystallization in gall-bladder bile of pigs given cholesterol-beta cyclodextrin-enriched diets with either casein or soyabean concentrate as protein sources. AB - Cholesterol precipitation from supersaturated bile is the earliest and determinant step in the formation of cholesterol gallstones, which is thought to be diet-dependent. Bile composition, appearance and growth of cholesterol crystals were studied in fresh gall-bladder biles from pigs adapted to four different protein-containing diets over 3 weeks: 160 g dietary protein/kg as casein (C16; n 6), or as soyabean-protein concentrate (S16; n 6), or a mixture of both protein sources (casein-soyabean protein, 70:30, w/w) (CS16; n 6), or 320 g of the mixed protein/kg (CS32; n 6). Moreover, all four diets contained 3 g cholesterol/kg and 50 g beta-cyclodextrin/kg as modifiers of bile composition towards cholesterol pro-crystallization. Cholesterol precipitation was most active after the high-protein diet, CS32, and the casein diet, C16, and lowest after the soyabean-protein diet, S16. It was intermediate after the mixed diet, CS16, but still much lower than in the former two groups. These diet-induced variations were suggested to be mediated through modifications in the biliary profile of bile acids, whereas all other biliary constituents studied were essentially unchanged. The fasting level of plasma cholesterol was lowest in both 160 g protein/kg diets containing soyabean protein (S16 and CS16), highest for the high-protein diet CS32, and intermediate for the C16 diet. These results should encourage clinical studies on the effect of soyabean protein, or other vegetable proteins, for primary or recurrence prevention of cholelithiasis at its earliest stage. PMID- 10858700 TI - The effect of dietary peptide concentration on endogenous ileal amino acid loss in the growing pig. AB - The aim of the present study was to determine whether dietary peptide concentration had an effect on endogenous ileal amino acid flow in the growing pig. Eight 33 kg live weight entire male pigs had post-valve T-caecum (PVTC) cannulas surgically implanted for the collection of ileal digesta. The pigs were fed twice daily at 100 g/kg metabolic body weight per d and were given diets containing enzyme-hydrolysed casein (EHC) at 0, 50, 100 and 200 g/kg in a Latin square design. A basal casein-based diet was fed to the pigs for 6 d periods between receiving the experimental diets. The pigs received the experimental diets for 8 d periods, with continuous collection of digesta for 24 h on each of the fifth and eighth days. The endogenous ileal amino acid flows were determined with reference to recovery of the marker, Cr, directly for pigs receiving the protein-free diet or after centrifugation and ultrafiltration (10,000 Da molecular mass cut-off) for pigs on the EHC-based diets. Mean endogenous ileal N flows were 1753, 1948, 2851 and 5743 micrograms/g DM intake when the pigs received diets containing 0, 50, 100 and 200 g EHC/kg respectively. There was a significant (P < 0.05) effect of dietary peptide concentration on the endogenous ileal flows of N and all of the amino acids, with an increase in endogenous ileal amino flow with increasing dietary EHC concentration. PMID- 10858701 TI - The type of dietary fat alters the hepatic uptake and biliary excretion of cholesterol from chylomicron remnants. AB - The consumption of fat-enriched diets may alter the uptake and metabolism of chylomicron remnant cholesterol by the liver. To test this hypothesis, [3H]cholesterol-labelled chylomicron remnants derived from different dietary fats were studied in perfused livers both from rats fed on diets enriched in the corresponding fats and from rats fed on a low-fat diet. Livers from rats fed on each of the fat-enriched diets removed similar amounts (34-40%) of the [3H]cholesterol-labelled remnants added, whereas livers from rats fed on the low fat diet removed significantly more labelled fish-oil and butter-fat remnants than olive-, maize- or palm-oil remnants. Significantly more remnant [3H]cholesterol was secreted into the perfusate HDL by livers from rats fed on the olive-oil, fish-oil and butter-fat diets when compared with those from rats fed on the low-fat diet or the maize-oil diet. Furthermore, the excretion of remnant [3H]cholesterol via the bile acid was increased by the olive-, maize-, palm- or fish-oil diets, and decreased by the butter-fat diet when compared with the low-fat diet, although the [3H]bile acid excreted remained less on saturated fat diets. This investigation shows that the hepatic uptake and subsequent metabolism of cholesterol from chylomicron remnants is influenced by the type of fat in the diet as well as the fatty acid composition of the particles themselves, and may help to explain some of the hyper- and hypocholesterolaemic effects of saturated and unsaturated fatty acids. PMID- 10858702 TI - Diets containing long-chain n-3 polyunsaturated fatty acids affect behaviour differently during development than ageing in mice. AB - The effect of a standard diet providing essential fatty acids enriched in fish oil or palm oil was studied in young, mature and old mice. Two groups of pregnant and lactating OF1 mice were fed on diets with or without high levels of long chain n-3 polyunsaturated fatty acids. Offspring were maintained on these diets after weaning. The litter size did not differ. The weight increased more quickly in fish-oil-fed mice than palm-oil-fed mice. The fish-oil diet induced a significant increase in exploratory activity in young mice which was not found in mature and old mice. The level of locomotor activity was significantly higher in young, no different in mature, and lower in old fish-oil-fed mice than in controls. Habituation, the simpler form of learning, occurred to the same extent in the two diet groups. For the place learning protocol of the Morris water maze there was no difference between the two diet groups; however, in the probe trial, the mature fish-oil-fed mice remembered the situation well compared with the control mice. In the active avoidance test, on the first day of acquisition the young fish-oil-fed mice made more avoidances than control mice, whereas in contrast, mature and old-fish-fed mice made less avoidances than control mice. These results suggest a positive effect on arousal and learning ability of a diet enriched in long chain n-3 polyunsaturated fatty acids in young mice and a detrimental effect in old mice. PMID- 10858703 TI - Effects of dietary essential amino acid deficiencies on immunological variables in broiler chickens. AB - Two experiments were conducted to determine the effects of essential amino acid deficiencies on several immunological variables in male broiler chickens. Essential amino acids were classified into five groups as follows: S-containing amino acids (SAA; methionine + cysteine), aromatic amino acids (AAA; phenylalanine + tyrosine), branched-chain amino acids (BCAA; isoleucine + leucine + valine), arginine plus lysine (Arg + Lys), and other essential amino acids (OEAA; glycine + serine + histidine + threonine + tryptophan). Chickens were fed ad libitum from 10 to 24 d of age on a control diet or amino-acid-deficient diets formulated to contain each amino acid group at 50% and 16% (Expt 1) at 50% (Expt 2) of the recommended requirements (National Research Council, 1984). Effects of feed consumption on immune responses were also considered by setting pair-feeding (Expt 1) or restricted-feeding (Expt 2) groups fed on the control diet. In Expt 1, changes in lymphoid organ weights varied with the type and degree of deficiency of amino acid groups, with BCAA deficiency markedly decreasing weights. The haemagglutinin titres against sheep erythrocytes did not change in any amino-acid-deficient chickens except that the titres were lower in chickens fed on the 50%- and 16%-BCAA diets as compared with their pair-fed counterparts. In Expt 2, the splenocyte proliferative response to concanavalin A was higher in the chickens fed on the BCAA- and Arg + Lys-deficient diets and lower in chickens fed on the SAA- and AAA-deficient diets than the control chickens, independent of feed consumption. These results suggest that the effects of specific amino acid deficiencies on immune responses cannot be generalized, and that BCAA have the greatest potential to modulate immune responses among the amino acids in chickens. PMID- 10858704 TI - Chancroid in the United Kingdom. PMID- 10858705 TI - The COPE report 1999. PMID- 10858706 TI - Syphilis in pregnancy. AB - Syphilis can seriously complicate pregnancy and result in spontaneous abortion, stillbirth, non-immune hydrops, intrauterine growth restriction, and perinatal death, as well as serious sequelae in liveborn infected children. While appropriate treatment of pregnant women often prevents such complications, the major deterrent has been inability to identify the infected women and get them to undergo treatment. Screening in the first trimester with non-treponemal tests such as rapid plasma reagin (RPR) or venereal disease research laboratory (VDRL) test combined with confirmation of reactive individuals with treponemal tests such as the fluorescent treponemal antibody absorption (FTA-ABS) assay is a cost effective strategy. Those at risk should be retested in the third trimester. Treatment during pregnancy should be with penicillin. In determining a penicillin regimen, the clinician must consider the stage of the maternal infection and the HIV status of the mother. Patients who are allergic to penicillin should be desensitized before treatment. Despite appropriate treatment, as many as 14% will have a fetal death or deliver infected infants. Treatment may further be complicated by the Jarich-Herxheimer reaction, a complex allergic response to antigens released from dead micro-organisms, which can cause fetal distress and uterine contractions. Thanks to effective intervention strategies and inexpensive penicillin, syphilis rarely complicates pregnancy in the Western world today. In parts of the world where the traditional sexually transmitted diseases have not been controlled, the magnitude of problems associated with syphilis during pregnancy is reminiscent of that faced by the West during the early 1900's. PMID- 10858707 TI - Pelvic inflammatory disease epidemiology: what do we know and what do we need to know? PMID- 10858708 TI - The repertoire of human efforts to avoid sexually transmissible diseases: past and present. Part 2: Strategies used during or after sex. AB - BACKGROUND/OBJECTIVE: Despite the focus by public health programmes on condoms, chastity, or monogamy, people use a much wider variety of strategies to minimize their personal risk of sexually transmissible disease (STD). The objective of this study was to compile a comprehensive list of personal and societal STD avoidance strategies. METHODS: Data from clinical and research observations, computer searches, and historical texts were pooled. RESULTS: A variety of behaviours during or after sex, other than condoms, were identified that have been perceived to alter STD risk. STD avoidance strategies were often poorly documented and difficult to disentangle from other drives such as aesthetics, sexual variety, and contraception. They also varied in popularity in time and place. Some examples were douching; systemic and topical prophylactic antimicrobials; non-penetrative sexual practices, post-coital urination; and examining sexual partners' genitalia. Interest in some practices has been recently revived--for example, vaginal microbicides and post-exposure chemoprophylaxis, while others--for example, withdrawal and non-penetrative sexual practices, receive scant attention but may be much more widely used. CONCLUSION: The full spectrum of STD avoidance strategies warrants further study because some are ubiquitous across cultures and because they have the potential to complement or undermine safer sex programmes. Because of their greater acceptability, some less efficacious strategies may have greater public health importance than less popular but more efficacious strategies such as condoms. PMID- 10858709 TI - Are adolescents being screened for sexually transmitted diseases? A study of low income African American adolescents in San Francisco. AB - OBJECTIVES: To determine the proportion of sexually experienced African American adolescents who report having been screened for sexually transmitted diseased (STDs), and to determine the proportion who report having been screened for STDs among those adolescents who have had a preventive primary healthcare visit in the past 2 years. METHODS: A telephone survey of a population based sample of African American 12-17 year old adolescents residing in a low income San Francisco neighbourhood with a high prevalence of STDs. RESULTS: Of the 302 adolescents surveyed, 118 (39%) reported a history of sexual intercourse. Of these, 26% of the males and 59% of the females had been screened for an STD in the previous 12 months. 31% of the males and 63% of the females had been screened for an STD in the previous 24 months. Of the 93 participants who had had a preventive primary care visit since their first episode of sexual intercourse, 26% of the males and 60% of the females had been screened for an STD in the previous 24 months. CONCLUSIONS: Sexually experienced African American adolescents in San Francisco are being screened for STDs at rates well below that recommended by current clinical guidelines. A low rate of screening was found even in those adolescents who had been seen for a preventive primary care visit since they first had sex. This suggests that the preventive primary care visit is not being used to its full potential as an opportunity to screen and treat adolescents for STDs. Capitalizing on this opportunity to screen may increase the number of STDs diagnosed and, thus, decrease rates of STDs in this population. PMID- 10858710 TI - Sexual behaviour and early coitarche in a national sample of 17 year old Swedish girls. AB - OBJECTIVE: To evaluate sexual behaviour in 17 year old girls, using data from a national survey on adolescent sexuality. METHOD: The study was based on two samples of 17 year olds, comprising 2% of the population born in 1973 and carried out in 1990. A school sample and a sample of school non-attenders were recruited in a two step procedure. Data were collected by anonymous self administered questionnaires. 2583 questionnaires were distributed. Response rates from students was 92%, for school non-attenders 44%. 1121 female students and 118 female school dropouts responded. RESULTS: 64% of the student girls had experienced their first intercourse; 16% were "early starters" with coitarche before age 15. STD and pregnancy were reported by 15% of early starters and pregnancy by 14%, p < 0.001 and 0.002 respectively when compared with later starters. The number of coital partners, experience of first date intercourse, and of oral and anal sex was higher in the early starters, p < 0.001. Early starters reported menarche at age 11 or earlier more often than the later starters (OR 2.30, 95% CI 1.48-3.56), as well as a perceived social age exceeding the chronological by 2 years (OR 1.94, 95% CI 1.34-2.80). Sexual abuse was reported by 20% of the early and 11% of the later starters, p = 0.002. Among school non-attenders no significant differences were found with regard to age for coitarche. A majority of 83% of the girls had experienced voluntary intercourse, and 49% were early starters. Five girls were mothers. STD was reported by 19% and induced abortion by 14%. Sexual abuse was alleged by 28%. CONCLUSION: Coitarche before age 15 is related to early menarche and high perceived social age. High number of partners and first date intercourse make early starters at increased risk for STD and unintended pregnancy. Sexual abuse is alleged more often by early starters. PMID- 10858711 TI - Sociodemography of genital Chlamydia trachomatis in Coventry, UK, 1992-6. AB - OBJECTIVE: To describe the sociodemographic and geographic risk factors for incident Chlamydia trachomatis genital infection. DESIGN: Cross sectional retrospective study of cases diagnosed in local genitourinary clinics. SETTING: Coventry, West Midlands, from 1992 to 1996. SUBJECTS: 582 female and 620 male Coventry residents aged 15-64 years diagnosed with one or more episodes of genital Chlamydia trachomatis infection by enzyme immunoassay. Subjects were assigned a Townsend deprivation score based on residence. The denominator population aged 15-64 years was derived from 1991 census data. RESULTS: The mean annual incidence of genital chlamydia was 151 episodes (95% CI 140-163) per 100,000 population in men and 138 episodes (95% CI 128-149) per 100,000 population in women. Highest subgroup incidence was observed in 15-19 year old black women (2367 (95% CI 1370-4560) per 100,000), and 20-24 year old black men (1951 (95% CI 1158-3220) per 100,000). In univariate analyses, the most important risk factor for chlamydia infection in males was being black (incidence 1377 (95% CI 1137-1652) per 100,000 for black v 133 (95% CI 122-145) per 100,000 for white; RR 10.4, p < 0.0001) and for women was young age (incidence 475 (95% CI 415-540) per 100,000 for age group 15-19 years v 52 (95% CI 45-60) per 100,000 for age group 25-64 years; RR 9.1, p < 0.0001). In Poisson regression models of first episodes of genital chlamydia, for both males and females the effect of ethnic group could not be fully explained by socioeconomic confounding. There were significant interactions between age and ethnic group for both sexes and between age and level of deprivation for men. Geographical analysis revealed a high incidence of genital chlamydia in estates on the edge of the city as well as the urban core. CONCLUSIONS: There is a complex interaction between geographical location, age, ethnic group, and social deprivation on the risk of acquiring genital Chlamydia trachomatis in Coventry. Better population based data are needed. PMID- 10858712 TI - Trends in gonorrhoea in nine western European countries, 1991-6. European Study Group. AB - OBJECTIVE: To present, describe, and assess trends in gonorrhoea in western Europe between 1991 and 1996. METHODS: A European Union concerted action was initiated in 1990 to monitor the prevalence of HIV among patients with a sexually transmitted infection in sentinel networks in western Europe. Data from this concerted action were used to assess trends in gonorrhoea between 1991 and 1996. Where possible, the trends were validated by comparing them with national laboratory reports or data from more extensive sexually transmitted infection surveillance networks. RESULTS: 7192 episodes of gonorrhoea were recorded at 38 sentinel sites in nine countries between 1991 and 1996. In most networks, there was a decline in the number of cases of gonorrhoea among heterosexual men and women. The decline was most marked in the Scandinavian countries. Decreases were also observed among men having sex with men, but in some networks--England and Wales, Netherlands, and Scotland--an increase was observed in more recent years. This increase was mainly the result of an increase in cases among the older age group (25 years and above). The trends observed in six of the sentinel networks were confirmed by trends in national laboratory reports or data from more extensive sexual transmitted infection surveillance systems. CONCLUSIONS: These data indicate that, overall, there was a decline in the number of gonorrhoea cases in western Europe between 1991 and 1996. The results, however, also indicate that in more recent years there was an increase in the number of gonorrhoea cases among men having sex with men in some countries. Further investigations are necessary to determine if this observation is due to an increase in risky sexual behaviours in this population group. PMID- 10858713 TI - Effect of a syphilis control programme on pregnancy outcome in Nairobi, Kenya. AB - OBJECTIVES: To assess the impact of a syphilis control programme of pregnant women on pregnancy outcome in Kenya. METHOD: Women who came to deliver to Pumwani Maternity Hospital (PMH) between April 1997 and March 1998 were tested for syphilis. Reactive rapid plasma reagin (RPR) tests were titrated and confirmed with treponema haemagglutination test (TPHA). Equal numbers of RPR and TPHA negative women were enrolled. Antenatal syphilis screening and treatment history were examined from the antenatal cards. RESULTS: Of 22,466 women giving birth, 12,414 (55%) were tested for syphilis. Out of these, 377 (3%) were RPR reactive of whom 296 were confirmed by TPHA. Syphilis sero-reactive women had a more risky sexual behaviour and coexistent HIV antibody positivity; 26% were HIV seropositive compared with 11% among syphilis negative mothers. The incidence of adverse obstetric outcome defined as low birth weight and stillbirth, was 9.5%. Syphilis seropositive women had a higher risk for adverse obstetric outcome (OR 4.1, 95% CI 2.4-7.2). Antenatal treatment of RPR reactive women significantly improved pregnancy outcome but the risk of adverse outcome remained 2.5-fold higher than the risk observed in uninfected mothers. CONCLUSIONS: These data confirm the adverse effect of syphilis on pregnancy outcome. This study also shows the efficacy of antenatal testing and prompt treatment of RPR reactive mothers on pregnancy outcome. PMID- 10858714 TI - Pleural effusions in patients with AIDS. AB - OBJECTIVE: To describe the range of pathology causing pleural effusions in HIV infected patients with acute respiratory episodes and to attempt to identify whether any associated radiological abnormalities enabled aetiological discrimination. METHODS: Prospective study of chest radiographs of 58 consecutive HIV infected patients with pleural effusion and their microbiological, cytological, and histopathological diagnoses. RESULTS: A specific diagnosis was made in all cases. Diagnoses were Kaposi's sarcoma, 19 patients; para-pneumonic effusion, 16 patients; tuberculosis, eight patients; Pneumocystis carinii pneumonia, six patients; lymphoma, four patients; pulmonary embolus, two patients; and heart failure, aspergillus/leishmaniasis, and Cryptococcus neoformans, one case each. Most effusions (50/58) were small. Bilateral effusions were commoner in Kaposi's sarcoma (12/19) and lymphoma (3/4) than in para pneumonic effusion (3/16). Concomitant interstitial parenchymal shadowing did not aid discrimination. A combination of bilateral effusions, focal air space consolidation, intrapulmonary nodules, and/or hilar lymphadenopathy suggests Kaposi's sarcoma. Unilateral effusion with focal air space consolidation suggests para-pneumonic effusion if intrapulmonary nodules are absent: if miliary nodules and/or mediastinal lymphadenopathy are detected, this suggests tuberculosis. CONCLUSIONS: A wide variety of infectious and malignant conditions cause pleural effusions in HIV infected patients, the most common cause in this group was Kaposi's sarcoma. The presence of additional radiological abnormalities such as focal air space consolidation, intrapulmonary nodules, and mediastinal lymphadenopathy aids aetiological discrimination. PMID- 10858715 TI - A phase I study of a novel potential intravaginal microbicide, PRO 2000, in healthy sexually inactive women. AB - BACKGROUND: Although the male condom provides a reliable means of preventing HIV transmission, a broader choice of methods is required particularly in circumstances where the negotiation of condom use is difficult. Development of new products that may be effective as topical vaginal microbicides is the focus of a great deal of research activity currently. The novel agent PRO 2000, a naphthalene sulphonate derivative with in vitro activity against HIV and other sexually transmissible pathogens, is one such compound. We have studied the local and systemic safety and tolerance of a vaginal gel formulation of this agent at two concentrations (0.5% and 4%) over a 2 week period of daily exposure in two cohorts of healthy sexually abstinent women (one in London, UK, and the other in Antwerp, Belgium). METHODS: This was a randomised, placebo controlled, double blind, three arm clinical trial conducted on two sites. Macroscopic evidence of genital epithelial changes was sought using colposcopy and evidence of microscopic inflammation was acquired using high vaginal biopsy from predetermined sites (UK cohort only). Blood levels of PRO 2000 were measured and laboratory safety tests, including coagulation screens, were performed. The impact on vaginal ecology was also assessed. RESULTS: 73 women were enrolled across both sites (36 UK, 37 Belgium); 24, 24, 25 in the 4%, 0.5%, and placebo groups respectively. Of these, 70 completed 2 weeks' exposure to the study gel. Three (all in the 4% group) withdrew owing to adverse events which were possibly or probably gel related. Cervicovaginal abrasion was seen colposcopically in three subjects after 14 days of gel use (two in the 4% group and one in the placebo group). Genital ulceration was not seen during gel use in any of the subjects who completed the study. Histological evaluation of vaginal biopsy samples (36 women only) showed evidence of increased inflammatory signs in one participant of the 4.0% group. One volunteer in the placebo group had moderate inflammation at screening and at follow up. Severe inflammation was not seen among any of the subjects tested. Plasma levels of PRO 2000 and laboratory safety tests showed no evidence of systemic absorption. No impact was seen on normal vaginal ecology in the UK cohort where samples were taken 12 hours after the last gel application. CONCLUSION: In this phase I study PRO 2000 gel was found to be generally well tolerated with promising local and systemic safety profiles. The 0.5% gel was better tolerated than the 4% gel as fewer genital epithelial adverse events were seen in the former. Phase II studies are about to begin in sexually active women. PMID- 10858716 TI - Passive sentinel surveillance system for sexually transmitted diseases in primary healthcare sites in Ethiopia, 1991-3. AB - BACKGROUND: In 1989 the ministry of health of Ethiopia launched an STD control programme to strengthen the STD case management capabilities at public health centres and hospitals. The programme included the introduction of a syndrome based system for notification of STD cases. We here report the data originated by the syndromic case reporting system under programme conditions. METHODS: 35 (17%) of the total 225 hospitals and public health centres of Ethiopia were included in the programme. Information relevant to the years 1991 to 1993 was analysed at mid 1994. RESULTS: 32 clinical sites (91% of the total) provided at least one monthly report. The proportion of monthly reports received was 65% of those due, ranging from 51% in 1991 to 73% in 1992 and 42% in 1993. A total of 77,294 consultations for STD related symptoms were recorded, including 70,200 new cases, 6588 repeated consultations, and 506 partners of STD patients. Among first attendant patients 38,459 (52.7%) were males with a male to female ratio of 1:1. Urethral discharge and vaginal discharge were the leading cause of consultation among males (58%) and females (64%) respectively. The frequency of genital ulcer diseases was 26% among males and 15% among females. Inguinal adenopathy in the absence of genital ulcers was also frequent, accounting for 10% of consulting males and 5% of females. Based on Gram stain, gonorrhoea was identified in 64% of the cases of urethral discharge, while trichomoniasis and candidiasis were identified by wet mount in 28% and 16% of the cases of vaginal discharge respectively. CONCLUSIONS: STDs are a common cause of consultation at public health centre sites in Ethiopia. A syndromic case reporting system proved to be efficient and produced valuable information to initiate assessment of the problem and to set up bases for monitoring trends of STD morbidity. PMID- 10858717 TI - A colorimetric detection system for Calymmatobacterium granulomatis. AB - OBJECTIVE: To incorporate the first polymerase chain reaction (PCR) assay for Calymmatobacterium granulomatis into a colorimetric detection system for use in routine diagnostic laboratories. METHODS: A capture oligonucleotide specific for the Klebsiella phoE gene was covalently linked to tosyl activated magnetic beads. Biotinylated phoE PCR products obtained from 14 positive specimens from patients with donovanosis and isolates of Klebsiella pneumoniae, K rhinoscleromatis, and K ozaenae were cleaved with HaeIII for the purpose of differentiation, captured by the prepared beads, and subjected to standard EIA detection methodology. Eight samples from unrelated genital conditions underwent the same procedure. It was anticipated from the sequence data that the biotinylated fragment would be cleaved from the capture oligonucleotide target region in the three Klebsiella phoE products (that is, a negative colorimetric result) while the entire fragment of interest would remain intact in the positive C granulomatis phoE products (that is, a positive colorimetric result). RESULTS: All 14 positive specimens from patients with donovanosis gave strong colorimetric readings with this detection system. Isolates of K pneumoniae, K rhinoscleromatis, K ozaenae, and the eight specimens from unrelated genital conditions were negative. CONCLUSION: The successful development of a colorimetric detection system for C granulomatis incorporating two levels of specificity enables the molecular diagnosis of this condition to be undertaken by routine diagnostic laboratories. This should have an important role in the Australian government's campaign to eradicate donovanosis by 2003 though the test still needs to undergo trials and be validated using a larger number of samples from geographically diverse parts of the world in order to ascertain the generalisability of the methodology. PMID- 10858718 TI - Diagnostic tests for chancroid. PMID- 10858719 TI - Fortune's child. PMID- 10858720 TI - Successful treatment of recalcitrant condyloma with topical cidofovir. PMID- 10858721 TI - Bladder carcinoma presenting to genitourinary medicine departments. PMID- 10858722 TI - Atrial myxoma and HIV infection. PMID- 10858723 TI - The association between receptive cunnilingus and bacterial vaginosis. PMID- 10858724 TI - Is partner notification in the public interest? PMID- 10858725 TI - Sexual partner reduction and HIV infection. PMID- 10858726 TI - Features of AIDS and AIDS defining diseases during the highly active antiretroviral therapy (HAART) era, compared with the pre-HAART period: a case control study. PMID- 10858727 TI - Cardiac screening before non-cardiac operations. PMID- 10858728 TI - [Study of the onset and progression of peripheral neuropathy and hypertension in NIDDM]. AB - BACKGROUND: Diabetic neuropathy is the most common pathology affecting the peripheral nervous system. In prognostic terms, it is the most devastating complication of diabetes. About 50% of diabetics suffer from neuropathy between 25-30 years after the diagnosis of diabetes, even if over the past few decades there has been a considerable improvement in the diagnostic methods and criteria used to classify peripheral neuropathies, many of which are related to the development of neurophysiology. However, we still do not know enough about the incidence, prevalence and natural history of peripheral neuropathy diagnosed using clinical and electrophysiological criteria in non-insulin dependent diabetic patients. METHODS: The authors carried out a randomized study of the relationship between glucose intolerance, hyperglycemia, hyperinsulinemia, hypertension and early and manifest forms of peripheral neuropathy in 32 patients with NIDDM (aged 41-72 years old, duration of diabetes 1-27 years) over a 24 month period. In 11 patients diabetes was almost at onset (Group 1): 8 cases with diabetes for 1-2.5 years (4 hypertensives and 4 normotensives) and 3 cases with diabetes for 4 years (all normotensive). Twenty-one patients (Group 2) had had diabetes for longer (5-27 years): 5 were hypertensive and 16 normotensive. A full longitudinal neurophysiological study (EMG and ENG) was performed. In 11 NIDDM in Group 1, at basal conditions carpal tunnel syndrome (right CTS) was revealed in 1 case, right CTS with diffuse radiculopathy in 1 case, diffuse radiculopathy in 2 cases, lumbosacral radiculopathy in 1 case, and 1 right CTS with "mixed" symptoms. EMG-ENG were normal in 2 patients. RESULTS: The following developments were noted during the follow-up: rapid deterioration due to the onset of motor sensitive polyneuropathy (MSPN) in 1 patient, 3 cases of chronic neurogenic disorder with active denervation, 2 cases of "mixed" type symptoms. The results were only comparable in 2 cases. In 3 NIDDM with diabetes for 4 years, 1 patient presented MSPN and 2 were affected by chronic neurogenic disorders; during the follow-up the conduction of MSPN and active denervation deteriorated into chronic neurogenic syndrome. Moreover, 6 initially normotensive NIDDM developed hypertension. In 21 NIDDM of Group 2, 7 of the 16 who were initially normotensive became hypertensive. Three new cases of polyneuropathy were also reported in this group, and 5 already had MSPN but showed a deterioration of conduction during the follow-up in 1 case. One patient presented active denervation in chronic neurogenic symptoms and chronic neurogenic symptom was comparable in 1 case. One patient presented a normal EMG-ENG at both the start and end of the study. "Mixed" type of symptoms were recorded at the basal level in 11 patients (defined as the presence on the EMG of muscular areas with multiphase potentials of brief duration and low amplitude, first recruited under slight voluntary effort, isolated or mixed with areas of neurogenic potentials). Over the course of 12-24 months, eight patients deteriorated with chronic neurogenic symptoms without active denervation in 5 and present in 2 cases. One case also showed a deterioration of carpal tunnel syndrome. CONCLUSIONS: These results show that 1) metabolic control and a complete neurophysiological examination are essential for preventing and identifying the onset and progress of neuromuscular damage; 2) the onset or deterioration of these two complications mainly had a less well known common cause which was less studied and described. PMID- 10858729 TI - [Prognostic factors in the treatment of advanced cervical cancer. A retrospective study]. AB - BACKGROUND: The value of prognostic factors in patients with advanced cervix carcinoma treated by radiotherapy was assessed in a retrospective study. METHODS: From January 1977 through December 1990, 261 patients (average age 60 years) were treated at the Radiotherapy Department of the University of Turin. Distribution by stage was: 142 T2b (54%), 8 T3a (3%), 98 T3b (38%) and 13 T4 (5%). 83% of the patients underwent radiotherapy alone; the total dose was 45-88 Gy in 91 patients (42%) with poor clinical conditions, 60-75 Gy in 121 (56%) and 75-80 Gy in 5 cases. 17% of the patients was treated by surgery combined with radiotherapy. The median follow-up was 50 months (minimum 2, maximum 177 months). RESULTS: The 5 year NED survival and local control were 42% (52% for T2b, 33% for T3 and 15% for T4). The severe (G3-G4) complication rate was very low (1.9%). CONCLUSION: In our series, the prognostic factors which significantly influenced survival in the uni variate analysis were: advanced T stage, contemporary infiltration of parametrium and vagina, nodal status, non squamous neoplasm, younger age and the absence of brachytherapy in the radiotherapy alone protocol. PMID- 10858730 TI - [Takayasu's arteritis]. AB - A brief review of Takayasu's Arteritis (TA), a chronic granulomatous arteritis that mainly affects the aorta and its major branches, is made. The various ethiopathogenetic mechanisms which may give origin to the vascular damage and its pathologic pattern are described. TA is a more widespread disease than previously stated, it is not exclusive of young women of Japanese origin but it is actually present worldwide irrespective of age and with variegated patterns of clinical and angiographic presentation. However, two main forms may be identified: the Japanese form in which prevails the aortic arch involvement and the Indian form in which vasculitis is present in abdominal aorta and its branches (above all the renal arteries) with an upright extension to the thoracic aorta and a protean clinical picture with a more systemic spectrum. The new clinical and angiographic criteria for TA definition are reviewed and stressed. PMID- 10858731 TI - [Clinical contribution to the knowledge of Lyme disease. Case reports]. AB - Lyme disease (LD) is a well-recognized multi-system disorder, caused by the spirochaeta Borrelia burgdorferi. It involves the skin, nervous system, joints and heart. Our group reported the first case of Lyme Borreliosis in Italy in 1985. Italian Group for Lyme Borreliosis has recently reported the epidemiology of Lyme disease in Italy. The epidemiological data seem to underestimate the prevalence of Lyme arthritis in Italy. The pattern of articular involvement is most often mono- or oligoarthritis with polyarticular arthralgias and frequently needs laboratory confirmation. In this study, some clinical observations of Lyme Borreliosis occurred in the last year are reported. Analysis of the B. burgdorferi immune response by ELISA and Western Blot techniques has the potential to support or exclude a diagnosis of early stage as well as active LD infection was associated with Polymerase Chain Reaction assay based on amplification of the DNA of B. burgdorferi. When the direct evidence of spirochaeta in tissues is lacking, this approach permits the best evaluation of Lyme borreliosis. PMID- 10858732 TI - [With regard to the disorders of the musculoskeletal system]. PMID- 10858733 TI - Epilepsy management at primary health care level. South African Medical Association-Neurological Association of South Africa Epilepsy Working Group. PMID- 10858734 TI - Harmonization of dental education and curricula in Europe. AB - The need to harmonize undergraduate dental education and the recognition of dental degrees throughout the member countries of the European Union was recognised early on and was subject of EEC Directives in 1978. Growth of the EEC into the EU with the addition of more member nations has lead to continuing debate about the appropriate standards and measures for dental education. Attention has been given to defining both the 'input' measures of stipulating curricula and the 'output' measures of testing the competency of graduating dentists. PMID- 10858735 TI - Caries prevalence in 12-year-old schoolchildren in Senegal in 1989 and 1994. AB - Senegal, in middlewest Africa, has an estimated area of 197,000 km2 with a population of about 8.3 million of whom 45 per cent are less than 14-years of age. Apart from densely populated areas around the capital, Dakar, and other large towns, the population is scattered sparsely in rural areas. In the absence of any recent and representative oral health data, two cross-sectional studies on caries prevalence were conducted in 1989 and 1994 respectively to assess and compare the caries status among 12-year-old schoolchildren living in the urban and rural areas of Senegal. The results of the study are presented in this paper. PMID- 10858736 TI - Dentistry in Taiwan, Republic of China: national health insurance reforms, illegal dentistry and plans for peer review quality control. AB - The dental health care system in Taiwan, Republic of China is described in terms of demographics, structure, context of treatment and historical development of the dental health care payment system. A notable characteristic of the system is the existence of trade dentists, who operate without licensure. Their popularity and price advantage has maintained a political base that affects policy decisions. Health care reforms of March, 1995 with a comprehensive national health insurance, as well as ambitious plans for systematic peer review quality control of dentists' work are unique health care developments worthy of the attention of health care policy makers in other countries who are studying health care reform processes. PMID- 10858737 TI - Results of oral health and hygiene education in an institution for multiple handicapped children in Indonesia. AB - In a collaboration between the Dutch and Indonesian Dental Associations a system for oral health care for multiple handicapped children was initiated in a large care and rehabilitation institution in Jakarta, Indonesia. Part of the project was to develop a programme for oral health and hygiene education (OHE), with a specific plaque control component. This programme was aimed not only at the handicapped children but also at their parents and the teaching and (para) medical staff of the institution. A study to investigate the feasibility, acceptability and effectiveness of the OHE programme was carried out over a period of 2 1/2 years. The programme was well accepted, effective and of clinical significance. It is probably one of the first of its kind in a developing country. PMID- 10858738 TI - Dental anxiety among patients prior to different dental treatments. AB - The purpose of the present study was to evaluate dental anxiety among patients anticipating various dental treatments. One hundred and eighty patients who were scheduled for specific dental treatments at the School of Dental Medicine, the Hebrew University--Hadassah, Jerusalem, Israel participated in the study. Patients were anticipating one of the following dental procedures: scaling, filling, root canal therapy, preparation for crown, periodontal surgery or extraction. For each of the six dental treatments, 30 patients were selected at random. Dental Anxiety Scale (DAS) questionnaires were completed by the patients while waiting for their treatments. The results indicated that extraction caused the highest score, followed by scaling (though not significantly). Patients in the 35 to 49 year age group showed the highest total DAS scores. Women demonstrated higher total DAS scores than men. The relative influence of gender, type of treatment and age on the anxiety scores is demonstrated by using logistic regression, which revealed gender to have the strongest impact on the DAS scores, followed by the type of treatment and age. PMID- 10858739 TI - The intra-oral camera, dental health communication and oral hygiene. AB - This study aimed to determine the effectiveness of oral-hygiene instruction in improving oral health in 100 patients following oral hygiene instruction, with and without use of an intra-oral camera. The two groups of 50 patients were similar in age and sex distributions, frequency of caries, plaque accumulation and gingival bleeding. Prospective improvements in oral hygiene and compliance were measured by means of plaque levels and gingival bleeding at baseline and four weeks later. While both groups showed a clear reduction in plaque accumulation, the test group benefited from the use of the intra-oral camera. A majority of patients (88 per cent) thought that the extra information provided by the camera was helpful and desirable. This study demonstrates that the intra-oral camera can effectively augment oral-hygiene instruction and help create improvements in patient compliance. PMID- 10858740 TI - Relationship between oral tori and temporomandibular disorders. AB - The aim of the study was to compare the presence of oral tori and parafunctional activity (clenching, grinding teeth and/or bruxism) between temporomandibular disorder (TMD) patients and control subjects. Fifty-nine TMD and 353 control subjects were included. The groups were similar in sex and age distribution. There was no significant difference in prevalence of torus palatinus (TP) between TMD and control group (P = 0.2), while torus mandibularis (TM) was more common in TMD than in control group (P < 0.0005). Parafunctional habit was more common in TMD patients than in the control group (P < 0.0005). There was no significant association between size of TM and Helkimo's dysfunction index (P = 0.4). The results show that the prevalence of TM and parafunctional activity was higher in TMD than control patients. TM might be useful as an indicator of increased risk of TMD in some patients. PMID- 10858741 TI - Frequent use of sugar products by schoolchildren in 20 European countries, Israel and Canada in 1993/1994. AB - The aim of this study was to describe the daily use of certain between-meal sugar products (soft drinks and sweets) of schoolchildren in 20 European countries, Israel and Canada as a part of the Cross-National Survey on Health Behaviour in School-Aged Children--a WHO Collaborative Study. The data were collected using standardised anonymous questionnaires in school classrooms during the 1993/1994 school year. In each country nationally or regionally representative samples of about 1,300 schoolchildren (450 in Greenland) were targeted. Use of sugar products was analysed according to sex, age, country, self-reported school performance and self-reported family economy. One third to one half of the children (30-48 per cent) drank coke or other soft drinks more than once a day in Israel, Northern Ireland, Scotland, the Slovak Republic and Flemish-speaking Belgium. Use of soft drinks was very uncommon in Finland, Sweden, Norway, Denmark, Latvia and Estonia. The strongest association (odds ratios) was between the use of soft drinks and good family economy; in Russia (20.3), in Lithuania (11.3), in Latvia (10.0), in Poland (8.5) and in Estonia (8.0). In Israel, Scotland, Northern Ireland, Russia and French-speaking Belgium 41-29 per cent of the children ate sweets more than once a day. Boys drank soft drinks and ate sweets slightly more often than girls did. In conclusion, large differences were found between the different countries in the frequency of use of soft drinks and sweets. This should be considered when developing the content of oral health promotion programmes. PMID- 10858742 TI - Knowledge and reported behaviour concerning the prevention of caries in children: a questionnaire survey of Japanese parents resident in London, UK. AB - The objective of this study was to assess the knowledge and self-reported behaviours of a sample of Japanese parents resident in London, UK with respect to the prevention of caries in their children. Japanese parents were asked to complete a structured questionnaire which was distributed through the school authorities to seventeen primary schools. Data was collected on: socio demographics, dental service use in their children, knowledge and reported behaviours with respect to the prevention of dental caries in their children. After one reminder the questionnaire response rate was 69 per cent (n = 111). The mean age of the children was 6.75 years (range 3-12 years) and 73 per cent had been born in Japan. Those respondents with children born in the UK were more likely to take their children to the dentist regularly and more likely to correctly identify the cause and prevention of dental caries. Whilst a majority of respondents reported that their children brushed their teeth twice daily, toothpaste was not used by one quarter of the children. Barriers to dental attendance were identified. Whilst there were some similarities in levels of knowledge and reported behaviours, the parents of Japanese born children were disadvantaged with respect to knowledge about the causes and prevention of dental caries and the use of UK primary dental care services. These parents should be offered information about these issues. PMID- 10858743 TI - Implant identification system. AB - Osseointegrated oral implantology has become a widespread option of dental care. A universal system of implant identification is required to enable dentists, patients and participating third parties to accurately identify a particular implant and historically record and follow its bio-clinical status. A simple system, based on the existing FDI two-digit tooth identification system is presented. PMID- 10858744 TI - A review of atraumatic restorative treatment (ART). AB - The purpose of this paper was to critically analyse the results obtained with the Atraumatic Restorative Treatment (ART) technique. The ART approach involves the excavation of cavitated carious lesions with hand instruments and restoration of the cavities and associated pits and fissures with a glass ionomer restorative material. The clinical trial outcomes involving ART include retention rates, cost effectiveness, operative sensitivity, and the effect of personnel with different educational backgrounds involved in this alternative operative treatment. Comparative studies involving permanent and deciduous teeth using amalgam and glass ionomer sealants have also been included in the same projects. Specially defined clinical criteria have been used to evaluate the results. ART offers an opportunity for restorative dental treatment under field conditions where no electricity is available. Three-year data have been published, but long term studies using relevant comparison alternatives are lacking. ART has so far been largely employed on populations with a low DMFT. The technique should also be applied to high risk patients with rampant caries before the maximal benefit of the treatment can be ascertained. PMID- 10858745 TI - The impact of the ART approach on the treatment profile in a mobile dental system (MDS) in South Africa. AB - The changing profile of oral care rendered through the Mobile Dental System (MDS), after the introduction of the Atraumatic Restorative Treatment (ART) approach is described. During the first year of introduction of ART, the percentages of amalgam restorations and tooth extractions decreased significantly (P < 0.0001). This is partly ascribed to a change in choice of treatment by dental operators in favour of ART and also due to an increase in acceptance by patients because of the reduced fear, and the patient-friendly nature of the ART approach. The reduction in amalgam restorations was 16.0 per cent for permanent and 1.4 per cent for primary posterior teeth. Extraction of posterior teeth was reduced by 17.4 per cent in the permanent and 35.7 per cent in the primary dentitions. The restorative component of oral care increased by 33.4 per cent in the permanent and 37.1 per cent in primary posterior teeth. The one-year survival of one-surface ART restorations using Fuji IX and KetacMolar was 93.6 per cent. Full and partial (more than 90 per cent) retention of the sealant part of the ART restoration was obtained in 75 per cent of the cases after one-year. During the one-year period, infection control was made more simple and this facilitated easier maintenance of mobile dental equipment. The introduction of the ART approach reduced extraction, restored more teeth and made oral care in the MDS more preventive, less threatening and thus more patient-friendly. PMID- 10858746 TI - The aetiology of the non-carious cervical lesion. AB - Erosion and abrasion have been widely reported as causes of non-carious cervical lesions (NCCL). However, more recently, tooth flexure has been implicated in the formation of these lesions generating renewed interest in the pathogenesis of the non-carious loss of cervical tooth substance. This paper considers the causes of erosion and abrasion, related to modern lifestyles, and reviews the literature concerning tooth flexure as a cause of NCCL. A description of different types of NCCL is given, as an aid to determining the aetiology, yet at the same time accepting that the causation and pathogenesis of NCCL is probably multi-factorial resulting in many different clinical presentations. Consideration is given to the indications for treatment of NCCL and to the selection of materials for restoring such defects. PMID- 10858747 TI - Measuring work stress among Dutch dentists. AB - Individual differences among dentists determine to a large degree what is experienced as work stress, but assessment of specific areas is necessary to be able to act preventively. The aim of this study was to develop an instrument to be used to monitor the experience of work stress in detail, to measure its levels, and to relate those levels to job (dis-) satisfaction. A questionnaire was developed, the Dentists' Experienced Work Stress Scale (DEWSS), covering widespread aspects of dental work, which was completed by 709 dentists, forming a highly representative sample of Dutch general dental practitioners. Factorial, correlational, and reliability analyses were conducted, after which seven areas of stress emerged: Work Pressure, Financial Aspects, Patient Contacts, Work Contents, Career Aspects, Team Aspects, and Professional and Private Life. Of these, Patient Contacts and Work Contents showed highest mean scores, as did the specific items: 'defaulters', 'governmental instructions', and 'unreasonable or demanding patients'. A strong inverse relationship was found between work stress, in particular lack of career perspective, and job satisfaction. The questionnaire is a valuable instrument to monitor pressure at work as felt by Dutch general dental practitioners. Patient contacts, work pressure, and career perspective clearly need preventive attention in the Dutch situation. PMID- 10858748 TI - HIV infection, dental treatment demands and needs among patients seeking dental services at the Muhimbili Medical Centre in Dar-es-Salaam, Tanzania. AB - The objectives of this cross-sectional study were to determine the frequency of HIV infection among dental patients attending the three dental facilities at Muhimbili Medical Centre (MMC) in Dar-es-Salaam, Tanzania, and to compare the dental treatment demands and needs of the patients found to be HIV-infected with those of their HIV-seronegative counterparts. The facilities were; the dental outpatient department (DOPD) clinic, the dental minor surgery department, and the dental ward. This study which was conducted between March and April, 1996 enrolled a total of 460 patients. The investigations involved detection of anti HIV IgG antibodies in saliva, examination of oral and peri-oral tissues, and assessment of dental and periodontal status. The overall HIV frequency among the dental patients was 10.9 per cent. The frequencies of HIV infection among patients attending the dental OPD clinic, minor surgery, and those admitted in the dental ward were 9.4 per cent, 26.3 per cent, and 25.0 per cent, respectively. The dental treatment demands and needs of HIV-seropositive patients were not different from that of HIV-seronegative patients. The high frequency of HIV infection calls for institution of infection control measures in the dental clinics. However, such measures need to be tailored for the poor countries, with potentially high frequency of HIV infection and minimal resources, in order to make them relevant. PMID- 10858749 TI - Oral health status of children and adults in the Republic of Niger, Africa. AB - The present study was undertaken in order to describe the oral health status of children and adults in the Republic of Niger and to provide baseline data for the organisation and evaluation of systematic oral health promotion programmes in the country. The WHO pathfinder sampling procedures were applied to obtain representative samples of the following age groups: 6 years (n = 373); 12 years (n = 400); 18 years (n = 300) and 35-44 years (n = 400). Data were collected in 1997 according to the WHO methods, including information on dental caries and CPITN. In 6-year-olds, 56 per cent had caries and a mean score of 1.3 DMFT was observed among the 12-year-olds; the 35-44-year-olds had an average score of 5.7 DMFT. Differences in dental caries prevalence were found according to sex, province and urbanisation. Ninety-nine per cent of individuals at age 18, and 87 per cent at age 35-44 had maximum CPITN score 2 (calculus). Where 6- and 35-44 year-olds are concerned, the data may indicate increasing levels of dental caries. The implementation of primary prevention and community-based oral health education is therefore a matter of urgency. PMID- 10858750 TI - Restorative treatment decisions on approximal caries in Norway. AB - A random sample of dentists in Norway were asked which radiographic criterion for assessing the initiation of restorative treatment of approximal caries they would use, and which type of cavity preparation and filling material they would prefer for a distal lesion on an upper second premolar. Only 19 per cent stated that they would treat approximal lesions confined to enamel, with 81 per cent opting to wait until lesions had reached dentine, compared with 66 per cent in a similar study performed in 1983. The tunnel preparation was cited most often as the preparation of choice (47.3 per cent), while 28.2 per cent preferred traditional class II preparations and 24.3 per cent a saucer shaped preparation. Only 15.5 per cent of the dentists chose amalgam as the restorative, 15.8 per cent composite, 22.3 per cent a conventional glass ionomer cement, 7.2 per cent a resin modified glass ionomer cement and 22.4 per cent a combination of glass ionomer and composite. There has been a shift in operative treatment criteria among the majority of dentists in Norway from 1983 to 1995, with most now waiting until the lesion is diagnosed in dentine radiographically before restoring. Most dentists prefer new preparation techniques for approximal caries using tooth coloured materials. Only every fifth dentist has amalgam as a first choice for approximal restorations in the posterior region. PMID- 10858751 TI - Present state of dental health knowledge, attitudes/behaviour and perceived oral health of Japanese employees. AB - The aim of this survey was to assess the present state of dental health knowledge, attitudes/behaviour and perceived oral health of Japanese employees. A 60-item questionnaire was used in a dental health project in the work place. The subjects comprised 77,845 employees, 76 per cent of whom reported delaying a dental visit until they had toothache, with about 60 per cent delaying even when they discovered a decayed tooth. The majority did not regard decayed teeth as a disease and only a minority reported regular dental visits. About three quarters reported bleeding gums on brushing, although more than half had never been taught professionally how to clean their teeth and less than 5 per cent flossed daily. More than half believed that false teeth were inevitable in old age, and that their teeth were getting worse despite daily brushing. About 70 per cent of the employees thought that it was impossible to prevent gum disease with toothbrushing alone, and nearly half believed a toothpaste with fluoride was effective in preventing periodontal disease. Reorientation of oral health care in Japan, therefore, is urgently needed and dental services have to be provided for the implementation of systematic oral health promotion for employees in the workplace. PMID- 10858752 TI - Periodontal conditions in 65-74 year old adults in France, 1995. AB - In 1995, a study was undertaken in France to assess the periodontal health status of 603 noninstitutionalized elderly subjects aged 65-74 years. Thirty areas were identified in the Rhone-Alpes region, with a sampling method based on stratified quotas according to sex, place of residence and socio-economic group (S-EG). The CPITN index was used. The total prevalence of healthy dentate adults (n = 483) was 16.5 per cent, whilst 16.3 per cent of the adults were edentulous. The prevalence of CPITN code 1 + 2 (low) was 50.7. The higher S-EG having fewer codes 1 and 2 (45.8 per cent) than the lower S-EG (49.7 per cent) and the medium S-EG (55.7 per cent). The total prevalence of periodontal disease (code 3 + 4) was 31.5. The prevalence of periodontal disease was lower in adults of medium socio economic status and was also lower in adults living in urban residences. Overall, 66.9 per cent of the entire population needed oral hygiene instruction, 56.6 per cent scaling and 2.3 per cent complex periodontal treatment. PMID- 10858753 TI - Guidelines for the use of antimicrobial agents to minimise development of resistance. AB - There is currently worldwide concern about the problems of antimicrobial resistance. A number of important bodies such as the World Health Organisation and the British House of Lords have identified the reasons for the emergence of resistance to antimicrobial agents and the preventive measures which need to be urgently implemented to curb the spread of resistant organisms. The reasons for the emergence of resistant organisms are not difficult to find. During the past half-century, since the discovery of penicillin by Fleming, people in both the developing and the developed world have accepted antimicrobial agents as a fundamental right, not only to demand at the first sign of a trivial infection but also to self prescribe with readily available, cheap antimicrobial agents. Such unbridled abuse of antimicrobial agents not only in man but also in animals could lead down a slippery slope to an era where the microbe may rule supreme once again. Indeed some authorities are forecasting a 'post-antibiotic era' (as opposed to the pre-antibiotic era before the discovery of penicillin) in the foreseeable future when many infectious diseases will once again be almost impossible to treat. PMID- 10858754 TI - Dental caries, socio-economic development and national oral health policies. AB - A relationship between a population's level of socio-economic development and dental caries has often been assumed. Proxy measures such as sugar consumption have been used to reflect this. This study tests the hypothesis that there is a relationship between dental caries and the level of socio-economic development, using recent international data. It goes on to explore the implications of this relationship for the development of national oral health policies. Dental caries data was obtained from the WHO, Global Oral Epidemiology Data Bank for the period 1981-1996. Socio-economic data was obtained from the United Nations Development Programme (UNDP). Countries were ranked according to the Human Development Index (HDI) and their GNP. The study confirms the existence of a relationship between dental caries and development. Caries is a good proxy measure for socio-economic development. Countries in the throes of socio-economic transition have the highest DMFT scores. PMID- 10858755 TI - Malocclusion and orthodontic treatment need of secondary school students in Nigeria according to the dental aesthetic index (DAI). AB - The aims of this study were to measure the distribution, prevalence and the severity of malocclusion and treatment need amongst randomly selected (n = 703) rural and urban Nigerian children aged 12-18 years (mean 14.0 +/- 1.84) using the dental aesthetic index (DAI), and to assess whether malocclusion was affected by age, gender and socio-economic background. Data were collected according to the method recommended by WHO. Most of the children (77.4 per cent) had a dental appearance which required no orthodontic treatment. Over 13 per cent fell into the group where treatment for malocclusion is considered to be 'elective'. However, a substantial proportion (9.2 per cent) of the population had severe to handicapping malocclusion where treatment is 'highly desirable' or 'mandatory'. There were no statistically significant differences (P > 0.05) in DAI scores between age groups, gender and socio-economic background. This study also found that Nigerian adolescents had better dental appearance and less orthodontic treatment need compared with the Caucasian and Oriental populations. PMID- 10858756 TI - The prevalence of dental erosion in the maxillary incisors of 14-year-old schoolchildren living in Tower Hamlets and Hackney, London, UK. AB - The purpose of this study was to assess the prevalence of dental erosion in the maxillary incisors of a sample of 14-year-old schoolchildren and to explore the aetiological factors responsible for that erosion. The cross-sectionally design study took place in secondary schools in inner-city London, UK and involved 525, 14-year-old schoolchildren selected at random in a clinical examination and a self-completed questionnaire. The outcomes measures for dental erosion were; the prevalence, the area and depth of lesions and the risk factors. The prevalence of labial and palatal erosion was 16.9 per cent and 12 per cent respectively. Risk factors and behaviours including daily frequency of ingestion of acidic fruits and drinks, food vomiting, toothbrushing frequency, and swimming habits were not shown to have any relationship with the presence of erosion. It was concluded that the prevalence of erosion in the maxillary incisors of this sample was higher labially and lower palatally than in previously reported national figures. The risk factors which were investigated were not shown to have any relationship with the presence of erosion. Further investigations of these issues are necessary to establish whether or not dental erosion is a public health problem in the UK. PMID- 10858757 TI - Fluoride release and uptake by aged resin-modified glass ionomers and a polyacid modified resin composite. AB - Little has been reported of the relationship of fluoride release and weight loss, and the effects of use of different fluoride agents on restorations, for the new generation of glass ionomer cements. The objectives of this study were to compare fluoride release of fresh and aged specimens of a polyacid-modified resin composite (Dyract), and of three resin-modified glass ionomer cements (Fuji II LC, Photac-Fil, Vitremer); and to correlate fluoride release and weight loss of aged specimens after recharging with three different fluoride agents. All materials showed high initial fluoride release immediately after uptake when using the agents. However, the levels of fluoride release dropped rapidly soon afterwards. Although initial fluoride release was significantly different between Dyract and the three resin-modified glass ionomers, when different fluoride agents were used on aged specimens after recharging, no significant differences were found after the first few hours. Linear regression analyses also showed no correlation between cumulative fluoride release and weight loss. Possible beneficial oral health effects may only be expected by frequent exposure of these materials to fluoride agents. PMID- 10858758 TI - Carbamide-containing polyol chewing gum and prevention of dental caries in schoolchildren in Madagascar. AB - The objective of this investigation was to evaluate the effect on dental caries experience of using carbamide polyol chewing gum as a supplement to standard oral hygiene procedures for schoolchildren in a developing country (Madagascar). In 1994, grades 1 and 4 children of demonstration schools were allocated to experimental and control groups; all children participated in a school-based oral health education programme, including daily toothbrushing supervised by the classroom teacher. At grade 1, the test group (n = 125) also used chewing gum (V6: 55.5 per cent sorbitol, 4.3 per cent xylitol, 2 per cent carbamide) three times a day. At grade 4, one test group (n = 177) had chewing gum three times a day and an additional test group (n = 74) had chewing gum five times a day. The control groups included 117 children at grade 1 and 209 at grade 4. Dental caries was registered in 1994 and 1997 according to the Recording System for the Danish Child Dental Services. In grade 1 children, the preventive effect of the total DMFS was not statistically significant except for occlusal caries (-0.65 DMFS, P < 0.01). In the grade 4 test groups, non-significant reductions of dental caries experience were found when compared with controls. The present community trial indicates that the use of polyol chewing gum may be considered a supplement to the control of occlusal dental caries in young primary schoolchildren, particularly in developing countries with limited resources for dental care. PMID- 10858759 TI - The influence of oral bacteria on the surfaces of resin-based dental restorative materials--an in vitro study. AB - Three tooth-coloured, resin-based restorative materials (Charisma, Dyract, and Pertac) were exposed to typical oral bacteria (S. mutans, S. oralis and A. naeslundii) over a period of up to 35 days. The three strains of bacteria all colonised the resin-based materials within a few hours and formed thick bacterial films. Determination of the bacterial glucose consumption and lactate production during the incubation period showed no difference from the controls which contained no resin samples. Following the experimental exposure, the materials were examined by scanning electron microscopy (SEM) for possible surface damage and roughness was measured in a perthometer. Little damage to the resin-based composite material surfaces (Charisma, Pertac) could be observed, whereas the polyacid-modified composite material (Dyract) showed greater damage. There was a significant difference in the resin surface roughness after exposure to S. mutans and to A. naeslundii. The study clearly showed that the bacteria used strongly adhered to the resin-based restorative materials. As a consequence of bacterial colonisation and/or poor oral hygiene, damage to the restorative materials might develop. This suggests the need for dentists to evaluate personal oral hygiene, along with general indications and economic factors, in selecting materials for restorations, since the known anti-bacterial properties of amalgam are considerable. PMID- 10858760 TI - Integrating oral health into primary health care--experiences in Bangladesh, Indonesia, Nepal and Tanzania. AB - When primary health care (PHC) was developed and implemented in developing countries, oral health was not included. The present consequences are marked disparities in the distribution of oral health care, since conventional dentistry can only serve relatively few people and at high costs. Oral health care is virtually non-existent in rural areas of most developing countries where more than 80 per cent of the population live. More recently, community based oral health programmes have been initiated in some countries to fill the gap. These programmes give more emphasis on oral health promotion and on the prevention of oral diseases than on treatment of its consequences, since history has shown that the latter is ineffective in preventing oral diseases. Unfortunately, most of these oral health programmes have been implemented next to the existing PHC system and hence they face enormous management, logistic and financial problems, which seriously threaten their sustainability. This paper presents a proposal to counteract the problems that many countries face in developing an adequate primary oral health care (POHC) service. PMID- 10858761 TI - Clinical management of selected oral fungal and viral infections during HIV disease. AB - The purpose of this review is to describe current possibilities of management of selected fungal and viral oral opportunistic infections including oral candidiasis, herpes simplex type 1 and 2-related lesions (HSV1,2), oral hairy leukoplakia (OHL) and oral lesions associated with human papilloma viruses (HPV). Less common diseases such as cytomegalovirus infection or human herpes virus type 8 associated with Kaposi's sarcoma and others are not considered. In a number of instances lifelong therapy or prophylaxis has to be instituted. Antiretroviral combination therapy, also called highly active antiretroviral therapy (HAART), has considerably changed the frequency of oral lesions caused by opportunistic agents. A short description of the antiretroviral agents available including respective side-effects is presented. PMID- 10858762 TI - Infection control practices and compliance to national recommendations among dentists in Romania. AB - The aim of this study was to measure dental office compliance with current Romanian infection control regulations. A questionnaire was completed and returned from 61 randomly selected offices (32 private and 29 public with 94 dentists), where the sterilizers were also biologically monitored. Results indicated that with few exceptions, infection control practices in public and private offices were the same, with compliance on sterilising reusable instruments. Private offices monitored their sterilizers more frequently and had much newer equipment. Gowns were universally worn, but use of gloves, masks and protective eye-wear showed non-compliance with less than 10 per cent of the offices using personal protective equipment for all patients. Cost was the deciding factor. Predominant environmental disinfectants were alcohol and bleach. Offices were in compliance as to the use of disposable anaesthetic needles and carpules. Dentists reported reluctance to be vaccinated against hepatitis B even when offered free immunizations (6.4 per cent) and only 18.1 per cent of dentists had received any infection control training in the last three years. Results indicate that offices were in compliance for most national regulations. However, there are no recently published standards for dentistry in Romania concerning disinfectants or continuing education. Comparison with the literature indicates comparable compliance with recommended national infection control procedures for Romanian dentists as for dentists in other countries. PMID- 10858763 TI - Dental knowledge and attitudes among Arab schoolteachers in northern Israel. AB - A representative random sample of 597 Arab school-teachers in northern Israel, was surveyed regarding sources and levels of knowledge and attitudes about dental caries prevention. Data were measured according to a self-administered questionnaire from a 91.4 per cent response rate. When ranking the effectiveness of different caries preventive measures teachers on average listed optimal water fluoridation as a lower priority compared to toothbrushing, dental visits, fluoride mouthrinses and eating fewer sweet products. Placing of fissure sealants was ranked as the second least effective caries preventive measure, with 39.6 per cent not knowing the effectiveness. Only 68.5 per cent of the school-teachers were aware of the anti-bacterial role of fluoride, and only a small minority knew of fluoride's potential in healing incipient caries. Teachers seemed less motivated to being involved in dental health school programmes which involved dedicating school time and their active involvement, such as fissure sealant programmes at school, supervision of brushing and flossing, and school mouthrinsing programmes. They revealed positive attitudes towards: informing parents about the importance of oral hygiene and teaching children about preventive dentistry. Teachers' main reported source of knowledge was the dental office. It is the responsibility of the dental profession to ensure that updated knowledge is correctly conveyed to schoolteachers, who are an important and potentially influential sector of dental health consumers and health education agents. PMID- 10858764 TI - Experimental intervention study about recognition of erythroplakia by undergraduate dental students. AB - The aim of this study was to assess the efficacy of an examiners' training programme applied to identification of erythroplakia, which undergraduate dental students have found particularly difficult. An experimental group of 5th year undergraduate dental students received a pictorial handout with diagnostic criteria for oral red lesions and two, one hour-long diagnostic seminars in which oral erythroplakia was discussed, together with other red lesions with similar clinical appearance. Three months later a set of 16 photographic slides depicting previously pathologically diagnosed red lesions were projected. Sensitivity, specificity and agreement were all higher for the experimental group compared to a control group. It is suggested that teaching procedures using slides could be useful for training future examiners at recognizing oral erythroplakias. PMID- 10858765 TI - Evaluation of bioartificial pancreas function made from sodium alginate and poly L-lisine microcapsules settled with viable pancreatic islets. AB - The possibility of transplantation of immuno-isolated viable pancreatic islets is a promising alternative for treatment diabetic patients. To establish the value of the artificial prosthesis made from settled with viable pancreatic islets microcapsulated using sodium alginate and poly-L-lisine, the glucose plasma levels were analyzed versus time after implantation into peritoneal cavity of rats or mice with alloxan-induced diabetes. The proper function of the bio artificial pancreas manifested in normalization of glucose plasma level has been observed from 14 to 21 days after implantation in diabetic mice and rats. PMID- 10858766 TI - Investigation of antistaphylococcal activity of collagen dressing containing bacitracin. AB - In the present paper the anti-bacterial effect of bacitracin released from the collagen-based dressing and penetrating to the Staphylococcus aureus-colonised polyurethane sponge was assessed. The bacteria-colonised sponge, regarded as the "in vitro model of an infected wound", was used as a modification of the previously utilised by us analogous experimental systems. In addition, kinetics of the penetration of bacitracin from the collagen dressing to the sponge was estimated. The results indicate that in spite of the fact that the amount of bacitracin in the polyurethane sponge exceeded the minimal bactericidal concentration (MBC), not all bacteria from the S. aureus species colonising the sponge could be killed. Moreover, it appeared that in order to efficiently eradicate the sponge-adherent staphylococci the concentration of bacitracin 50 fold higher than the MBC value must be used. PMID- 10858767 TI - The influence of new polyester block copolymer on morphology of hepatocytes of rats liver. AB - The liver is a complex organ with metabolic, excretory, and defence functions, so the effect of toxic polymers is often find in disorganisation of hepatic structure and may induce cell damage. In this study it is estimated the influence of recently synthesised multiblock copolymers based on poly/butylene terepthalate) and a dimerized fatty acid. The films of polymers were implanted intra-abdominally in 45 rats of Wistar breed covering the right lobe of liver. The specimens of liver were collected in 1, 3 and 6 day of experiment in order to be evaluated under microscope. The morphology of lobules, size and shape of hepatocytes, size and shape of nuclei's of hepatocytes, any infiltration of plasma cell were taken into account during examination. There have been no difference in morphology between specimens of liver from rats with implanted polymer and control group which could implicate their toxicity. PMID- 10858768 TI - [Comparative evaluation of biomaterials use in surgical treatment of periodontitis]. AB - Clinical and radiological evaluation of three biomaterials--HA-Biocer, Bio-Gran and Bio-Oss was carried out two years after their grafting in parodontium. The tests were carried out for 91 patients with parodontis. Comparative clinical results (remission of the inflammatory state of parodontium tissues, essential reduction of gingival pockets and rebuilding of alveolar process bone on aimed X ray pictures) were noted. The carried out observations show that biomaterials application for filling vertical alveoral process bone defects for patients with parodontis gives satisfactory clinical results. PMID- 10858769 TI - Reducing cataract surgery-related complications. PMID- 10858770 TI - Herpes simplex keratitis. AB - Chronic infection of the cornea by Herpes simplex virus (HSV) continues to be an important cause of unilateral blindness. Despite considerable progress in the understanding of the virus at cellular and molecular levels, the prospect of prevention still appears to be long way off. The development of non-toxic topical antiviral agents has been an important step forward in management. However, correct diagnosis and treatment, in particular, the avoidance of inappropriate use of topical steroid remains as important as ever. This article reviews the virological and clinical aspects of HSV keratitis including the current concepts of pathogenesis and management. PMID- 10858771 TI - Excimer laser phototherapeutic keratectomy: indications, results and its role in the Indian scenario. AB - PURPOSE: To report indications, technique, and results of excimer phototherapeutic keratectomy (PTK), and describe possible reasons for the small numbers of such procedures performed in a referral institute in India. METHODS: Retrospective review of case records of 10 patients (11 eyes) who underwent excimer PTK at our institute between February 1994 and September 1997. RESULTS: Corneal scars were the most common indication for treatment. Best-corrected visual acuity (BCVA) improved in 6 eyes (mean: 2 lines of Snellen acuity). All eyes had BCVA > or = 6/12 after treatment. None of the patients experienced loss of BCVA after treatment. Unaided visual acuity improved in 3 eyes and decreased in 2 eyes. Change in spherical equivalent refraction > or = 1 diopter occurred in 77.8% of eyes after treatment. Treating central corneal scars resulted in a significant hyperopic shift in refraction. CONCLUSIONS: Excimer PTK is a safe and effective procedure for the treatment of superficial corneal opacities. Post treatment ametropia may require further correction with optical aids. Inappropriate referrals, deep corneal scars, and cost of the procedure could have contributed to the small numbers of PTK performed at our institute. Improved understanding of procedural strengths and limitations could lead to increased use of this procedure, with satisfying results in selected patients. PMID- 10858772 TI - Incidence and management of posteriorly dislocated nuclear fragments following phacoemulsification. AB - PURPOSE: To report the incidence, management and complications of nucleus dislocation into the vitreous during phacoemulsification. METHODS: Retrospective review of 1250 consecutive phacoemulsification performed by consultants and residents in a teaching hospital. RESULTS: The incidence of nucleus drops was 0.8% (10 out of 1250). Loss of nuclear fragments occurred during phacoemulsification in 9 patients. In one, the dislocation was caused by hydro dissection. All except one patient (who refused further intervention) underwent pars plana vitrectomy with removal of nuclear fragments. Eight of them had intraocular lens (IOL) inserted at the time of cataract surgery or at vitrectomy; one patient was scheduled for a secondary IOL. Postoperative best corrected visual acuity ranged from 6/24-6/6; 8 patients achieved a vision of 6/12 or better. Complications included cystoid macular oedema (5 patients), retinal break (1 patient) and retinal detachment (1 patient). CONCLUSION: Appropriate management of posteriorly dislocated nucleus can restore good visual acuity. The use of phacoemulsification mandates availability of referral facilities for management of complications. PMID- 10858773 TI - Subconjunctival cysts following silicone oil injection: a clinicopathological study of five cases. AB - PURPOSE: To study the occurrence, risk factors and management of subconjunctival cysts formed following the use of intraocular silicone oil as a tamponade. METHODS: We analyzed 5 cases of single and multioculated subconjunctival oil cysts between 1986 and 1996. RESULTS: Cysts were observed 15 days to 4 months following silicone oil injection. Clinically they showed minimal inflammatory signs but histopathology of removed cysts showed emulsified silicone oil globules with chronic inflammatory cellular infiltration. CONCLUSION: Though silicone oil is considered to be nontoxic, it can cause chronic inflammation when spilled into the subconjunctival space. PMID- 10858774 TI - Management of ocular perforations resulting from peribulbar anaesthesia. AB - PURPOSE: To analyze the clinical presentation and outcome of treatment for globe perforation secondary to peri-bulbar anaesthesia. METHODS: Eight patients (3 females and 5 males) aged 66-84 years were included in the study. Ocular perforations were suspected in 3 cases before or during surgery, in 4 cases diagnosis was established within one week and in one case at 3 weeks. Three patients underwent indirect argon laser photocoagulation to seal the retinal break, one patient had cryotherapy, 3 patients underwent a pars plana vitrectomy with fluid gas exchange and endo-laser; and one patient refused any further treatment. RESULTS: The final visual acuity after a mean follow up of 14 months was better than 6/9 in 2 patients, between 6/9-6/12 in 4 patients, and perception of light in 2 patients. CONCLUSION: If diagnosed early and treated adequately, a majority of patients with globe perforation during periocular anaesthetic could be saved. PMID- 10858775 TI - Bacterial contamination of anterior chamber during IOL surgery. AB - PURPOSE: To study the nature and frequency of bacterial contamination during cataract surgery. METHODS: The preoperative smears from the conjunctiva and anterior chamber (AC) fluid aspirates during extra-capsular cataract surgery (ECCE) with posterior chamber intraocular lens (PCIOL) implantation in 40 eyes were analysed for aerobic and anaerobic bacteria. Any change in the bacterial strains isolated before and after cataract surgery was also studied. RESULTS: AC fluid aspirates were positive for bacteria in 15 eyes (37.5%). Coagulase-negative Staphylococcus was the most common aerobe (39.4%) and Propionibacterium acnes the most common anaerobe. Of the 15 cases with positive AC fluid cultures, 6 showed an organism in the AC aspirate different from the conjunctival smear. CONCLUSION: Clinically there was no endophthalmitis in any of the eyes. Factors such as preoperative antibiotic use, the antibacterial properties of aqueous, or low inoculum size could explain this. The preoperative conjunctival smear may not be useful in predicting the AC fluid contamination or outcome of cataract surgery. PMID- 10858776 TI - Cortical blindness: an unusual sequela of snake bite. AB - Several ophthalmic effects may follow snake bite; this report describes an instance of cortical blindness that resulted from snake bite. PMID- 10858777 TI - Lipaemia retinalis in a case of juvenile diabetic ketoacidosis. AB - A rare case of diabetic retinal lipaemia is described in a 5-year-old child. PMID- 10858778 TI - Blindness due to firearm eye injuries in rural western Uttar Pradesh. AB - In a retrospective analysis of 440 cases of firearm injuries, 104 patients had ocular injuries. Following treatment only 14 patients (13.5%) could regain visual acuity of > or = 6/60. PMID- 10858779 TI - Bilateral frosted branch angiitis and cytomegalovirus retinitis in acquired immunodeficiency syndrome. AB - This report describes a case of frosted branch angiitis associated with AIDS. PMID- 10858780 TI - Congenital retinal arterial loops and spontaneous vitreous haemorrhage: a case report. AB - This report describes a case of unilateral spontaneous vitreous haemorrhage associated with congenital retinal arterial loops. PMID- 10858781 TI - Yavatmal District Blindness Control Society: a case study. AB - PURPOSE: To retrospectively study the records and reports available at the District Blindness Control Society (DBCS), Yavatmal in terms of target fixation, performance and utilisation of manpower and equipment. METHODS: All the available records, reports, correspondence, and proceedings of meetings from 1981-98 were scrutinized and analyzed. RESULTS: The performance records and reports showed that over the last 10 years the target achievement of DBCS is close to 100%. However, the fixed facility (District hospital/Tertiary hospital where cataract surgeries are being performed under strict aseptic conditions) performance does not match the targets. The district mobile unit camp performance achieved 35-40% of the target in the last quarter of the financial year. CONCLUSION: The target fixation is irrational and needs improvement, and it is necessary for the program managers in the district to undertake analysis of the available data to ensure performance improvement. PMID- 10858782 TI - An alternative approach to a posteriorly dislocated intraocular lens. PMID- 10858783 TI - [Fifty years of plastic surgery in the Netherlands. I. Seemingly minor accidents: major hand injuries]. AB - In patients with hand injury, careful clinical examination is necessary, even in cases of seemingly trivial lesions such as small puncture wounds. Four patients, two women aged 18 and 51 years and two men aged 16 and 28 years, presented with a small volar cut caused by a glass fragment (three patients) and a saw. They suffered from loss of motor function, loss of sensibility or loss of circulation in one finger. They were treated by plastic surgery. Lesions of tendons and peripheral nerves can be diagnosed by relatively simple tests. It is important to recognize them at an early stage because adequate treatment may result in recovery of the sensibility and the motor function. Even a 'dead' finger may be salvaged. PMID- 10858784 TI - [Fifty years of plastic surgery in the Netherlands. II. Introduction]. AB - Plastic surgery is a medical specialism that was born out of was surgery. The first famous Dutch plastic surgeon, Esser, operated in Austria and Hungary during World War I. Koch, trained in England during World War II, and companions founded the Dutch Society for Plastic Surgery on October 7th 1950. During the last 50 years plastic surgery has changed into a very divers specialism, which focuses on craniofacial, esthetic, and reconstructive surgery, micro- and hand surgery, trans-sexuality and surgery of congenital abnormalities, especially of the face and hands. Each plastic operation aims at improvement of form and/or function. There have been many developments in all the fields of interest; these will be highlighted in a short series of articles. PMID- 10858785 TI - [Fifty years of plastic surgery in the Netherlands. III. Structural surgery of the genitalia]. AB - Plastic surgeons have contributed to the constructive and reconstructive surgery of the external genitals. This development followed the increase of surgical technical possibilities. Whereas in the past the possibilities were limited to transplantation of skin grafts and pedicled flaps, they have now been supplemented by free transplantation using microsurgical techniques. Other specialists have adopted the techniques developed by plastic surgeons. In future, in the structive surgery of genitals use will be made of the growing possibilities of transplantation of donor organs and the results of investigation of tissue cultures and biomaterials. The demand for aesthetical corrections of the external genitals will increase. PMID- 10858786 TI - [Fifty years of plastic surgery in the Netherlands. IV. Treatment of children with cleft lip and palate]. AB - During the last decades treatment of patients with cleft lip and palate exchanged a solitary for a multidisciplinary approach. This warrants a balanced development of cranio-maxillofacial growth yielding good results both in restoring facial contour and correcting functional defects. Nowadays, refinements in indications for treatment and in techniques, based upon scientific evidence are the goals. In the future more basic studies in the genetical and ecological field may provide tools for patient tailored counselling and effective prevention. PMID- 10858787 TI - [Prognostic benefits evaluated: new guidelines for systemic adjuvant therapy for axillary node-negative breast cancer vs. implementation of sentinel node biopsy procedure]. AB - The new guidelines for adjuvant systemic therapy in patients with resectable breast cancer adopted by the Dutch National Breast Cancer Platform (NABON) and the Dutch Society for Medical Oncology (NVMO) have as a prerequisite that adjuvant treatment must increase the expected absolute number of patients with 10 year survival by 5% or more. It is therefore apparently considered justifiable to treat 20 patients in order to obtain 10-year survival for one extra of these. This view appears to be in contrast to the view held by adversaries of implementation of the sentinel node biopsy procedure who consider it justifiable to submit 650 patients to a mutilating complete axillary lymph node dissection for disease-staging purposes in order to gain only one extra patient with a 10 year survival. PMID- 10858788 TI - [Adjuvant systemic therapy for patients with resectable breast cancer: guideline from the Dutch National Breast Cancer Platform and the Dutch Society for Medical Oncology]. AB - There is an abundance of evidence that adjuvant systemic therapy with chemotherapy or endocrine therapy results in better survival for all patients with resectable breast cancer. The absolute 10-year survival advantage however varies for the different patient groups. Therefore, for each individual patient the choice of adjuvant therapy must take into account the potential benefits and the possible side effects. A group of medical oncologists from the Dutch National Breast Cancer Platform (NABON) and the Dutch Society for Medical Oncology (NVMO) prepared a guideline for the treatment of patients with early resectable breast cancer. The criterium for choosing adjuvant systemic therapy for the individual patient is an expected increase in 10-year survival of 5% or more. In the guideline a difference is made between patients with and without axillary lymph node metastasis. In patients with axillary lymph node metastasis the choice for adjuvant systemic therapy depends on the following prognostic factors: menopausal status, age, and the presence of estrogen and progesterone receptors in the tumour. In patients without axillary lymph node metastasis the choice depends also on the following prognostic factors: the size of the tumour, the mitotic activity index, or the histopathologic grade of differentiation. PMID- 10858789 TI - [Long QT-interval syndrome and investigation of heritability: psychological reactions in three generations in one family]. AB - DNA diagnostics were carried out in a family after the long QT interval syndrome had been diagnosed in one of its members. The psychic reactions to this testing were different from those seen in other hereditary diseases such as Huntington's disease. This was probably due to the sudden and unexpected occurrence of the arrhythmia. The family members in whom clinical and DNA diagnostics gave purely negative findings were not relieved, owing to solidarity with the affected relatives. Their partners did not understand this response. The anxiety and concern of the gene carriers had nothing to do with their own health status but with that of their carrier children. These parents were in need of educational counselling and advice. The results of clinical and DNA diagnostics affected the family relationships: in carriers the feeling of closeness grew, while the non carriers were afraid of loss of family closeness. PMID- 10858790 TI - [Fifty years of plastic surgery in the Netherlands. V. History]. AB - For many centuries plastic surgical operations have been practised incidentally by general surgeons. Both World Wars combined with a multidisciplinary approach have been instrumental to the development of plastic surgery as a specialty. The Great War experience of maxillofacial surgery was for some surgeons in Europe a reason to dedicate their lives solely towards plastic surgery. A prominent Dutch surgeon in this field was J.F.S. Esser. With his systematic study and clinical application of arterial flaps in plastic surgery he proved to be far ahead of his time. Plastic surgery as such was established in the Netherlands after World War II in 1950 by the triumvirate Koch, Raadsveld and Honig. In the beginning, plastic surgery was strongly British influenced, while now, 50 years later, European cooperation on harmonization of the training in plastic surgery of a good quality is in progress. PMID- 10858791 TI - [Assistance in hunger strikes: legal guidelines]. AB - Hunger strikes raise ethical and legal issues, in addition to societal and medical ones. The World Medical Association adopted resolutions in 1975 (Declaration of Tokyo) and 1991 (Declaration of Malta) in which respect for the decision to refuse food was confirmed. A survey of the relevant international and national standards shows that in the Netherlands law and policy are more supportive of respect for food refusal (and against forced feeding) than would seem to be the case at the international level. However, respect for the decision of the hunger striker requires that it is well-considered, informed, and free from group coercion. The existence of an unambiguous legal framework will not save the advising physician from difficult dilemmas which will in particular occur in case of a protracted hunger strike. In anticipation of expected loss of judgement capacities in protracted hunger strikers it is advisable that the wishes of the striker and the professional policy that the physician will adopt are written down. In case of hunger strike legal standards cannot fully replace psychological insight, professional ethics and conscience, however. PMID- 10858792 TI - [Deep venous thrombosis of the arm: etiology, diagnosis and therapy]. PMID- 10858793 TI - [Deep venous thrombosis of the arm: etiology, diagnosis and therapy]. PMID- 10858794 TI - [Good results from a multidisciplinary and behavioral program for chronic back pain]. PMID- 10858795 TI - [Risk of inducing resistance upon stopping and restarting lithium after long-term usage]. PMID- 10858796 TI - Surgical treatment of epilepsy resistant to medical therapy. PMID- 10858797 TI - Understanding the GMC. PMID- 10858798 TI - The value of ultrasound scanning in breast disease. AB - Ultrasound is the modality of choice for imaging breast masses. Most benign and malignant lesions can be accurately categorized as a result of improvements in grey scale imaging and the development of sensitive colour Doppler vascular mapping. Ultrasound contrast agents, harmonic imaging, elasticity imaging and other advances will further enhance the utility of breast ultrasound. PMID- 10858799 TI - Developments in cardiac ultrasound. AB - This article gives an overview of recent developments in cardiac ultrasound for the general hospital physician. It discusses contrast echocardiography, harmonic imaging, three-dimensional echocardiography, Doppler tissue imaging and perfusion imaging and give an outlook on future perspectives. PMID- 10858800 TI - Surgical correction of congenital uterine anomalies. AB - Congenital uterine abnormalities have been associated with poor reproductive outcome. Anatomical corrections utilizing open or endoscopic surgery has been recommended to improve these outcomes. This article assesses the available evidence. PMID- 10858801 TI - Wegener's granulomatosis: unusual presentations. AB - Wegener's granulomatosis is a necrotizing vasculitis characterized by respiratory tract involvement and focal glomerulonephritis. Rare presentations include abdominal pain as a result of gut involvement, pericarditis, cardiac arteritis and blindness. Detection of antineutrophil cytoplasmic antibodies should not be used as a substitute for a histological diagnosis. PMID- 10858802 TI - Update on antiendotoxin therapies. AB - Endotoxin has been implicated in the processes that can lead to organ failure and death after surgery and critical illness. While there are no currently available commercial therapies directed against endotoxin, many have been tried or are in an experimental stage. In this article we outline past, present and future approaches to anti-endotoxin therapy. PMID- 10858803 TI - Weakness of the pelvic floor: urological consequences. AB - The pelvic floor comprises three compartments: anterior, posterior and middle. Weakness of the pelvic floor can lead to prolapse, urinary or faecal incontinence. This article deals with the defects in the anterior compartment which lead to urological consequences. The anatomy and management of stress incontinence are discussed. PMID- 10858804 TI - The role of entacapone in the management of Parkinson's disease. AB - Catechol-O-methyltransferase (COMT) inhibition is an important advance in the treatment of Parkinson's disease. This consensus statement provides guidelines for the optimal use of the only currently available COMT inhibitor, entacapone (Comtess, Orion Pharma (UK) Ltd, Newbury, Berkshire). PMID- 10858805 TI - Can doctors self-manage stress? AB - The NHS has expressed concern about stress in its workforce and is taking steps to reduce it. This paper reviews the factors associated with stress and burnout throughout a medical career. Clear messages emerge for all doctors and their employers. PMID- 10858806 TI - Disaster medicine: an emerging specialty. AB - The diverse nature of natural and man-made disasters demands a wide range of medical skills and resources from any attendant clinician. Working in a hostile environment may also require armed protection. This article considers the types of disasters, their medical requirements and the training of doctors wishing to be involved in this field of medicine. It includes a description of the Diploma in the Medical Care of Catastrophes. PMID- 10858807 TI - Bias in case-control studies. AB - Case-control studies are largely used to explore differences between groups of individuals. They can identify potential risk factors associated with disease, or they can investigate patient behaviour, such as why some people do not attend for services. As such, case-control studies are often used to generate or test hypotheses about causal factors. Nonetheless, bias is always a danger in case control studies, arising especially from the way in which study samples are selected or from the collection of retrospective data. Confounding also remains a problem. This short paper explores ways in which such flaws can be uncovered in published studies. PMID- 10858808 TI - Continuous ambulatory peritoneal dialysis and uterovaginal prolapse. PMID- 10858809 TI - Coronary arteritis in Wegener's granulomatosis causing fatal myocardial infarction. PMID- 10858810 TI - Not waving but drowning? PMID- 10858811 TI - Intestinal perforation as a presentation of Wegener's granulomatosis. PMID- 10858812 TI - Guidelines for MRSA control. PMID- 10858813 TI - Bisoprolol in heart failure. PMID- 10858814 TI - Photodynamic therapy. PMID- 10858815 TI - Photodynamic therapy. PMID- 10858816 TI - Radiology on the Internet. PMID- 10858817 TI - Knee arthroscopy in the day surgery unit. PMID- 10858818 TI - Differential processing of neuropeptides influences Drosophila heart rate. AB - Peptides that play critical physiological roles are often encoded in precursors that contain several structurally-related gene products. Differential processing of a precursor by cell-specific processing enzymes can yield multiple messengers with diverse distributions and activities. We have reported the isolation of SDNFMRFamide, DPKQDFMRFamide, and TPAEDFMRFamide from adult Drosophila melanogaster. The peptides are encoded in the FMRFamide gene and have a common C terminal FMRFamide but different N-terminal extensions. In order to investigate the processing of the FMRFamide polypeptide protein precursor, we generated antisera to distinguish among the structurally-related neuropeptides. Utilizing a triple-label immunofluorescent protocol, we mapped the distribution of the peptides. Each peptide has a unique, non-overlapping cellular expression pattern in neural tissue suggesting that the precursor is differentially processed. In order to identify a biological activity of the peptides, we established an in vivo heart rate assay. SDNFMRFamide decreases heart rate but DPKQDFMRFamide and TPAEDFMRFamide do not, indicating that the N-terminal residues are critical for this activity. SDNFMRFamide immunoreactivity is present in the aorta, implying that SDNFMRFamide acts locally to affect heart rate; DPKQDFMRFamide and TPAEDFMRFamide antisera do not stain cardiac tissue. Our data support the conclusion that Drosophila contains cell-specific proteolytic enzymes to differentially process a polypeptide protein precursor resulting in unique expression patterns of structurally-related, yet functionally distinct neuropeptides. PMID- 10858819 TI - Abnormal courtship conditioning in males mutant for the RI regulatory subunit of Drosophila protein kinase A. AB - The previously described site-selected P-element mutagenesis of a Drosophila gene encoding the regulatory subunit of cAMP-dependent protein kinase generates mutants that have defective behavior in the olfactory learning test. Here we describe the effect of the same mutations in a courtship conditioning assay. Wild type males can distinguish between virgin females (which they court vigorously), and fertilized females (which they court less vigorously). After exposure to fertilized females, wild-type males modify their behavior by decreasing courtship to subsequent target virgins, an effect that may last for many hours. Like wild type males, PKA-RI mutant males are also able to distinguish between virgin and fertilized females. PKA-RI males also modify their behavior towards virgin females after prior exposure to a fertilized female, but such an effect is short lived, suggesting a defect in memory rather than learning. We also show that under these conditions the behavior of PKA-RI males is similar to that of amnesiac, dunce and rutabaga males. PMID- 10858820 TI - The UNC-119 family of neural proteins is functionally conserved between humans, Drosophila and C. elegans. AB - C. elegans animals mutant for the unc-119 gene exhibit movement, sensory and behavioral abnormalities. Consistent with a nervous system role, unc-119 reporter genes are expressed throughout the C. elegans nervous system. The UNC-119 protein has strong sequence similarity to the predicted protein from a human gene, HRG4/HsUNC-119, whose transcript is abundant in the retina. Using these similarities, we have identified a Drosphila homolog, DmUNC-119, which is expressed in the Drosophila nervous system. The predicted C. elegans, human and Drosophila gene products are conserved across two domains. Expression of portions of HRG4/HsUNC-119 or DmUNC-119, directed by the unc-119 promoter, can fully rescue the C. elegans unc-119 mutant phenotype. We tested the ability of portions of HRG4/HsUNC-119 to rescue, and found that its function in C. elegans requires the conserved carboxyl terminus, while the dissimilar amino terminus is dispensable. UNC-119, HRG4 and DmUNC-119 constitute members of a new class of neural genes whose common function has been maintained through metazoan evolution. PMID- 10858821 TI - Fruitless is in the regulatory pathway by which ectopic mini-white and transformer induce bisexual courtship in Drosophila. AB - Bisexual courtship in male Drosophila melanogaster may be induced in some circumstances. These include ectopic expression of the transformer (tra) gene, ectopic expression of the mini-white (mw) gene, and the homozygous presence of mutant alleles of the fruitless (fru) gene. Experiments were performed to determine if ectopic mw and fru, as well as ectopic tra and fru, acted in the same pathway to control courtship. Male flies homozygous for the frusat allele court females little if at all and males at a low level. When homozygous, the frusat allele suppresses the bisexual courtship induced by both ectopic mw and ectopic tra, indicating that the fru wild-type function is necessary for expression of the ectopic mw and ectopic tra effect. This demonstrates that fru shares a pathway controlling courtship behavior with these ectopically expressed genes. PMID- 10858822 TI - Synaptic localization and restricted diffusion of a Drosophila neuronal synaptobrevin--green fluorescent protein chimera in vivo. AB - Fluorescent markers for subcellular compartments in Drosophila neurons should allow one to combine genetic mutant analysis with visualization of subcellular structures in vivo. Here we describe an analysis of two markers which may be used to observe different compartments of live Drosophila synapses. Soluble jellyfish green fluorescent protein (GFP) expressed at high levels in neurons diffuses freely in the neuronal cytosol as evidenced by confocal microscopy and fluorescence recovery from photobleaching experiments. Thus, the distribution pattern of soluble GFP in motor axons and larval motor terminals indicates the expected distribution for diffusible presynaptic molecules. In contrast to GFP, a neurally expressed neuronal synaptobrevin-GFP chimera (n-syb GFP) is transported down axons and specifically localized to nerve terminals. We demonstrate that n syb GFP labels synaptic-vesicle membrane at larval motor terminals by documenting its restriction to presynaptic varicosities, its colocalization with synaptic vesicle antigens, and its redistribution in Drosophila shits1 mutant nerve terminals transiently depleted of synaptic vesicles. Surprisingly, n-syb GFP expressed in muscle is concentrated at the subsynaptic reticulum (SSR), postsynaptic infoldings of muscle plasma membrane. We suggest, using different membrane markers, that this apparent postsynaptic enrichment simply reflects a concentration of plasma membrane in the SSR, rather than a selective targeting of n-syb GFP to postsynaptic sites. Utilities and implications of these studies are demonstrated or discussed. PMID- 10858823 TI - Caenorhabditis elegans degenerins and vertebrate ENaC ion channels contain an extracellular domain related to venom neurotoxins. AB - The DEG/ENaC (DEGenerin/Epithelial Na+ Channel) superfamily includes closely related ion channel subunits from divergent species ranging from the simple nematode Caenorhabditis elegans to humans. Members of this protein group play roles in several important processes including transduction of mechanical stimuli, sodium re-absorption and blood pressure regulation. Structure/function relationships in members of this superfamily are just beginning to be elaborated. Using a bio-informatics approach, we identified a novel structural element in the extracellular region of DEG/ENaC proteins that exhibits significant similarity to venom neurotoxins. Since venom neurotoxins bind to sodium channels at high affinity, we suggest that the related domain embedded in DEG/ENaC channels may interact with other regions of the channel or channel complex to modulate channel function. PMID- 10858824 TI - Clinical governance. PMID- 10858825 TI - Intravenous morphine for emergency treatment of cancer pain. AB - Despite the wide use of the World Health Organization (WHO) analgesic ladder for the relief of cancer pain, it is not uncommon to find patients presenting with severe pain to palliative care centres. This is more so in the developing world, where facilities for pain relief are few and the health care system is not well organized. It has been the practice in a pain and palliative care clinic in south India to give repeated boluses of 1.5 mg of morphine intravenously every 10 min to patients presenting with severe pain. An audit of the procedure was undertaken by a retrospective study of 793 case notes. Seventy-nine per cent of patients had total relief of their pain with intravenous morphine. Three per cent of patients experienced side-effects during the procedure. These included nausea and vomiting, itching, giddiness, restlessness, dyspnoea, chest pain, disorientation and a feeling of uneasiness. Thirty-two per cent of patients had drowsiness, which was one of the end-points of the procedure. It is concluded that intravenous morphine in repeated boluses of 1.5 mg every 10 min is a safe and effective method of managing cancer pain emergencies in a clinical setting in a developing country. PMID- 10858826 TI - Does spirometry predict dyspnoea in advanced cancer? AB - This study explores the similarities and differences between subjective assessments of dyspnoea and objective spirometric indices of respiratory function in advanced cancer. Of 155 patients investigated, 71 (46%) were dyspnoeic and 108 (70%) had spirometry (94 post-salbutamol). Of the 94, 84 had height and weight measured to calculate predicted spirometry. Average dyspnoea levels over 24 h were measured by patient visual analogue scales (VASMe 24). Forced expiratory volume after 1 s (FEV1) and forced vital capacity (FVC) were almost always lower than predicted, indicating frequent impaired respiratory function. Mean spirometric increase post-salbutamol was 21% for FEV1 and 12% for FVC. Correlations between VAS dyspnoea scores and spirometry were low; hence, the latter cannot be relied upon as a measure of the former. Respiratory impairment tended to be obstructive (mean FEV1/FVC = 65%). PMID- 10858827 TI - A comparison of artificial saliva and chewing gum in the management of xerostomia in patients with advanced cancer. AB - This was a prospective, randomized, open, crossover study comparing a mucin-based artificial saliva (Saliva Orthana) with a low-tack, sugar-free chewing gum (Freedent) in the management of xerostomia in patients with advanced cancer. The conclusions of this study were that both Saliva Orthana and Freedent are effective in the management of xerostomia in patients with advanced cancer, that both Saliva Orthana and Freedent cause some side-effects in this group of patients, and that patients with cancer think that chewing gum is an acceptable treatment. Sixty-nine per cent of the patients preferred the chewing gum to the artificial saliva. Furthermore, the chewing gum scored better than the artificial saliva on every measure of efficacy. However, none of these results reached statistical significance. PMID- 10858828 TI - The compatibility and stability of octreotide acetate in the presence of diamorphine hydrochloride in polypropylene syringes. AB - Varying concentrations of octreotide acetate (Sandostatin) and diamorphine hydrochloride were prepared and stored in polypropylene syringes at 37 degrees C in the dark. The solutions were analysed for octreotide acetate content using a validated HPLC method at regular intervals over a 48-h period. The results indicate that octreotide acetate remains stable in the presence of diamorphine hydrochloride at 37 degrees C for 24 h. In addition, the solutions prepared maintained their clarity, with no signs of precipitation upon visual examination under normal light conditions. PMID- 10858829 TI - Learning to care: a medical perspective. AB - The development of palliative care as a recognizable specialty has been supported by an acknowledgment of palliative medicine as a discrete discipline within the medical profession. While the knowledge and skills required for training in palliative medicine are well defined, there are elements of the medical care of people at the end of life that are more difficult to outline. Nursing practitioners and academics in particular have made important contributions in defining caring as an entity, and published work in the field of nursing, bio ethics and philosophy has encouraged an understanding of what caring is and how it is practised. However, it has rarely been addressed specifically in the medical literature. Undergraduate and postgraduate curricula outline some of the attitudes required to practise palliative medicine but the way in which doctors learn to care and indeed should care has not been clearly detailed. This paper reviews some of the literature pertinent to this aspect of palliative care, with particular reference to some of the elements that may influence how and why doctors learn to care in the way that they do in their practice of medicine. PMID- 10858830 TI - Wernicke's encephalopathy and terminal cancer: case report. PMID- 10858831 TI - Is the Hospital Anxiety and Depression Scale (HADS) useful in assessing depression in palliative care? PMID- 10858832 TI - Audit of three antimuscarinic drugs for managing retained secretions. PMID- 10858833 TI - GP views on developments in palliative care services. PMID- 10858834 TI - Zimbabwe. PMID- 10858835 TI - Palliative care needs of hospital inpatients. PMID- 10858836 TI - [Neuroplasticity and chronic pain]. AB - In the seventies and eighties spinal mechanisms inhibiting pain processing were discovered in animal studies leading to new therapeutic regimens such the use of spinal opioids. During the last decade additional studies revealed an increased sensibility of the spinal cord upon severe, long lasting pain perception, a mechanism called wind-up. Hyperalgesia is accompanied by persisting genetic changes of spinal cord cells, which may contribute to the chronification of pain. The severity and duration of acute pain apparently contributes to the possibility of chronic pain development. Although not all the consequences of these findings are clear, they may influence our way of performing anaesthesia and treating postoperative or acute pain situations, e.g. pain during herpes zoster or pain after trauma and amputation. In general, analgetic measures should be potent enough to prevent spinal sensiblisation, which can be best achieved with spinal blockade by local anesthetics. Another way of counteracting pain-induced spinal plasticity is by blocking or antagonizing its pathways with specific transmitters or their equivalents. All these spinally mediated regimens should be performed prior to later predominating mechanisms of supraspinal plasticity involving psychic changes due to persisting pain, which seem to evolve with delay to spinal processes. PMID- 10858837 TI - [Local anesthetics from ester to isomer]. AB - The use of chemical substances to prevent or treat local pain had its origin in South America. It was known that central nervous system stimulation occurred among the natives of Peru who chewed the leaves of an indigenous plant (Erythroxylon coca). Circumoral numbness was believed to have occurred as a by product of this custom. Attempts to isolate the active principle from these leaves finally resulted in the isolation of the alkaloid, cocaine, by Niemann in 1860. The clinical usefulness of cocaine was not appreciated until 1884, when Koller reported upon topical anesthesia of the eye. The chemical identification of cocaine as a benzoic acid ester led to the synthesis of numerous compounds which were basically benzoic ester derivates. In 1905, Einhorn reported the synthesis of procaine. Tetracaine, the most potent ester of the benzoic acid series appeared in 1930. A major breakthrough in the chemistry of local anesthetic agents occurred in 1943 when Loefgren synthesized lidocaine, since it was not an ester but an amide derivate of diethylamino acetic acid. Concerning structure-activity relationships, local anesthetic agents, in general, possess the chemical arrangement of: aromatic portion--intermediate chain--amine portion. Changes in the aromatic or amine portion of a local anesthetic substance will alter its lipid/water distribution coefficient and its protein-binding characteristics which, in turn, will markedly alter the anesthetic profile. The toxic effects of long-acting local anesthetics on brain and heart, firstly reported by Albright, provided the initial stimulus to develop new amide-type local anesthetics. The first of these drugs, which has come into clinical practice was ropivacaine, the S-enantiomer of two possible optical isomers. It is structurally related to bupivacaine and mepivacaine, exerting a different pharmacodynamic profile, specifically on cardiac electrophysiology (less arrhythmogenic than bupivacaine). Studies on the anesthetic activities and toxicity of the individual enantiomers of bupivacaine and mepivacaine generally indicate, that the S-enantiomers are longer-acting and less toxic than the R enantiomers. PMID- 10858838 TI - [Sympathoadrenergic, hemodynamic and stress response during coinduction with propofol and midazolam]. AB - OBJECTIVE: This study was undertaken to investigate simultaneously the influence of coinduction with propofol and midazolam on sympathoadrenergic and hemodynamic reactions and stress response during the extended induction period of TIVA. METHODS: 2 x 20 patients over the 60th year of life with major visceral surgery were investigated in a prospective randomized design. All patients received about 0.1 mg/kg BW midazolam for oral premedication. In the midazolam-group, induction of TIVA was undertaken with 0.05 mg/kg BW midazolam, 2.5 micrograms/kg BW fentanyl and 1.0 mg/kg BW propofol (1 min later). Controls received 2.5 micrograms/kg BW fentanyl and 2.0 mg/kg BW propofol. After muscle relaxation with 0.1 mg/kg BW vecuronium, TIVA was conducted in both groups with 8-5 mg/kg BW/h propofol and bolus injections of fentanyl with respect to clinical demands. Beyond consumption of anesthetics and recovery, sympathoadrenergic and hemodynamic reactions and SEF90 were investigated at 6 time points before induction of anesthesia and at skin incision. Plasma levels of ADH, ACTH and cortisol were measured twice before induction and at skin incision. RESULTS: Biometric data of both groups were comparable, and mean age was over 65 years. In the midazolam-group, co-induction with a mean dosage of 3.8 mg midazolam halved the induction dose of propofol and reduced the propofol demand during the following course of anesthesia as well, achieving a significant over all reduction of 200 mg propofol (p = 0.04). With respect to recovery from anesthesia more than 90 min later, no significant differences were found. Plasma concentrations of noradrenaline and adrenaline dropped significantly in both groups (p < 0.0001), as well as mean arterial pressure and heart rate (Fig. 1 and 2). For ADH, ACTH, cortisol and SEF90, no statistical differences were observed. Plasma-concentrations of midazolam were significantly higher in the midazolam group. CONCLUSION: In elderly patients, co-induction with 0.05 mg/kg BW midazolam halved the induction dose of propofol and led to a further dose reduction with maintained hypnotic potency. However, the halvage of propofol induction dose had no effect on the reduction of the sympathoadrenergic tone with decrease of blood pressure and heart rate. The overall moderation of the stress response was comparable as well. Thus, both induction regimens investigated in this study cannot be recommended in patients with severe hemodynamic disorders like hemorrhagic or cardiac shock. The propofol reduction should be primarily considered under economic aspects. PMID- 10858839 TI - Economic aspects of different muscle relaxant regimens. AB - OBJECTIVE: At a time of cost reduction in medical care efforts to manage the ever increasing costs of new pharmaceutical drugs become increasingly important. Costs of four different muscle relaxant regimens including the new intermediate-acting neuromuscular blocking drugs (NMBD) cisatracurium and rocuronium will be analyzed. METHODS: Eighty patients undergoing laparoscopic cholecystectomy were prospectively studied. All patients received standardized general anaesthesia with desflurane/fentanyl. Muscle relaxation was achieved with atracurium, cisatracurium, vecuronium, or rocuronium with 20 patients in each group. Intraoperatively muscle relaxants were added to maintain two twitches of the train-of-four (TOF) assessment. RESULTS: There were no differences among the four groups regarding biometric data, duration of surgery and anesthesia, number of patients with reversal of neuromuscular blockade, and time of extubation. Length of stay in the postanesthesia care unit (PACU) and the incidence of side effects were similar in all groups. Total costs of used drugs were significantly lowest in the atracurium-treated patients (per patient: 18.27 Euro) and significantly highest in the cisatracurium group (26.71 Euro) compared with the other groups (vecuronium: 22.61 Euro; rocuronium: 22.63 Euro). CONCLUSION: It is summarized that the use of cisatracurium was associated with higher costs compared to a standard muscle relaxant regimen using atracurium, whereas patient outcome was the same in all study groups. The routine use of the newer NMBDs can only be justified economically, if considerable improvements to clinical practice can be demonstrated. PMID- 10858840 TI - [Ventilation according to the "open lung" concept of multiple trauma patients]. AB - OBJECTIVE: The ventilation mode clearly influences the course of patients with multiple trauma on the ICU. Ventilation according the "open lung" approach rapidly opens up atelectatic lung regions. Generation of an adequate intrinsic PEEP enables to keep the lung open. We studied the consequences of the "open lung" approach on the lung function and monitored its side effects on patients with multiple trauma. METHODS: 18 consecutive patients with multiple trauma and additional thoracic trauma were routinely ventilated according the "open lung" approach between May and November 1999. We were mainly interested in data of lung mechanics, oxygenation and ventilation. Side effects on other organ systems and consequence for the infection rate were monitored. RESULTS: Ventilation according the "open lung" approach enables early sufficient oxygenation and ventilation of patients with severe multiple trauma and accompanying thoracic trauma. The ventilation mode helps to prevent baro-, volu- and atelectrauma and thus fulfils the requirements for a present-day ventilation mode. An immediate complete healing of the lung damages was not found. Nevertheless, as a trend the length of ventilation support seems short. Even extensive osteosynthesis at multiple fractures was possible without delay. Side effects of the high opening pressure on the lung or other organs as well as sequels of the high intrinsic PEEP on liver, kidney or intestine were not found. The infection rate was low, therapeutic doses of antibiotics were necessary only in less than half of the ICU stay. CONCLUSION: Ventilation according the "open lung" approach is a very effective and safe way to ventilate patients after severe multiple trauma with accompanying thoracic trauma. PMID- 10858841 TI - [The effect of midazolam and flunitrazepam on the liberation of lysozyme and beta glucuronidase from neutrophil granulocytes in vitro]. AB - OBJECTIVE: Polymorphonuclear neutrophile leucocytes (PMNL) play an important role in the defence against bacterial infections. It is known that some anaesthetics are able to disturb PMNL functions. We examined the influence of midazolam and flunitrazepam on the activity of the bactericidal enzymes lysozyme and beta glucuronidase released from PMNL in vitro. METHODS: As described before [4], PMNL were isolated from venous blood samples obtained from 10 healthy male volunteers. PMNL stimulation and measurement of lysozyme and beta-glucuronidase activities were conducted according to the description by Metcalf et al. [5]. The BIOMED system [8] was used for statistical evaluation. RESULTS: Neither midazolam nor flunitrazepam caused any statistically important alteration of lysozyme activity. However, clinically relevant concentrations of both benzodiazepines significantly enhanced beta-glucuronidase activity. The additives of flunitrazepam did not play any role. CONCLUSION: Surprisingly enough, midazolam and flunitrazepam increased the activity of beta-glucuronidase released from PMNL in vitro. At present, this result can neither be explained nor can its importance be estimated. On the other hand, the benzodiazepines did not relevantly influence lysozyme activity. PMID- 10858842 TI - [Ethical conflicts in emergency situations]. AB - Emergency medicine is a symbol of the fight against sudden death. By increasing medical and technical ability we come more and more in the position of acting against life threatening. The more we are able to do the more ethical problems must be considered by the emergency team, too. Therefore ethical tools are as necessary as qualification in pharmacological and medical ability. The analysis of case reports is one way to obtain knowledge of ethics. Ethical questions are shown by some emergency situations and criterions which have to be considered are named. PMID- 10858843 TI - [Quality of whole blood as a result of storage and preparation (inline-leukocyte depletion). Evidence for autologous predeposit]. AB - AIM OF THE STUDY: Investigation of various laboratory parameters in stored whole blood, with respect to duration of storage and kind of product. METHODS: Whole blood was donated by 12 healthy volunteers using CPDA1 stabilisator. Six units were filtered with the Leukotrap A1-System (PALL Comp., Dreieich, Germany) for leukocyte depletion. The twelve units were stored for 49 days. Several hematological, biochemical and coagulatory parameters were analysed during storage. RESULTS: There was an adequate reduction of lycocytes by filtration (< 3 x 10(6) white cells per unit). ATP decreased during storage to 45% of initial value at the 49th day, without any influence of the kind of preparation. The course of other parameters such as lactate and free haemoglobin (increase), PH value (decrease), antithrombin III (decrease), prothrombin, protein C, thrombin antithrombin-complex, alpha-2-Anti-plasmin (decrease or indifferent) did not show any influence of the kind of preparation. Coagulation factors V and VIII decreased in both preparations, which was significantly less pronounced in whole blood with leukocyte depletion. In contrast parameters of activated coagulation such as D-Dimere and fibrinmonomeric did not change during storage after leukocyte reduction but increased at the end of storage time in CPDA1-blood. CONCLUSIONS: Several parameters indicating quality of stored blood were constant in whole blood independent of the kind of preparation during a storage of 49 days. This is in contrast to the main part of specific scientific communications. A beneficial influence of leukocyte depletion was observed for some coagulation parameters whereas increasing characteristics of activated coagulation in CPDA1 stored whole blood at the end of storage time had to be observed. The preparation of whole CPDA1-blood is recommended for autologous predonation, if storage time does not exceed 30 days. Storage time of > 40 days seems to be possible for autologous whole blood after filtration for leucocyte depletion. PMID- 10858844 TI - [S-(+)-ketamine versus ketamine racemic mixture: hemodynamic studies]. AB - OBJECTIVE: Evaluation of hemodynamic effects of S-(+)-ketamine versus ketamine racemic mixture during induction of anesthesia, during steady-state of a fentanyl midazolam-anesthesia and in the period of aortic cross-clamping during extracorporeal circulation. METHODS: PATIENTS: 80 patients scheduled for coronary revascularization. STUDY DESIGN: double-blind, randomized. STUDY 1: Induction of anesthesia with ketamine-racemic mixture (3 mg/kg) or S-(+)-ketamine (1.5 mg/kg) plus midazolam 0.15 mg/kg. PARAMETERS: invasive hemodynamic monitoring including right ventricular volumes and pressure. STUDY 2: Bolus of ketamine-racemic mixture (3 mg/kg), S-(+)-ketamine (1.5 mg/kg) or placebo during steady-state anesthesia with fentanyl and midazolam. PARAMETERS: see study 1, additionally left ventricular systolic and end-diastolic pressure and maximum speed of left ventricular pressure increase (dp/dt). STUDY 3: Bolus of ketamine racemic mixture (3 mg/kg), S-(+)-ketamine (1.5 mg/kg) or placebo in the period of aortic cross clamping during ECC. PARAMETERS: mean systemic pressure, central venous pressure, reservoir volume. RESULTS: STUDY 1: Heart rate and systemic blond pressure remained unchanged until intubation, which caused significant increases of these parameters. Stroke volume Index and cardiac index decreased in the S-(+)-group compared with racemic mixture, right- and left ventricular filling parameters remained unchanged throughout the study. STUDY 2: There were no significant hemodynamic changes with time or between the groups. STUDY 3: Significant arterial vasodilation was observed in the racemic mixture group, venous parameters remained unchanged. CONCLUSION: There were no major differences in the hemodynamic profiles of S-(+)-ketamine and the racemic mixture. S(+) ketamine did not provide hemodynamic advantages. The use of both preparations should be limited to selected clinical situations in patients with reduced coronary reserve. PMID- 10858845 TI - [Hypothermia for cardiac surgery with cessation of circulation--start of open heart surgery in Germany]. AB - The first surface hypothermia with cessation of the circulation in Europe has been performed in Duesseldorf on February 9, 1955 for the direct suture of an atrial septal defect. Risks and insufficient basic knowledge are mentioned and the procedure is described with a protocol of anesthesia, cooling in ice water and rewarming in a water mattress. Eighth weeks later, the first series of 7 patients with closure of an atrial septal defect were presented at the congress of the German Society of Surgery. One patient died on the 13th postoperative day due to a cerebral thrombosis, unrelated to hypothermia. After 4 years a pump oxygenator became available and hypothermia was reduced to about half of the open heart procedures for 5 years and decreased further. Until 1978 with more than 1800 hypothermias for open heart surgery and some aneurysms of the aorta and cerebral arteries without the aid of a pump oxygenator the largest series in the world could be attained. As most important success was regarded that in none of our patients with cessation of circulation, which was repeated up to three times, a lasting cerebral damage occurred. PMID- 10858846 TI - [Long-term results of anatomic repair of transposition of great vessels]. AB - The long-term complications after anatomical repair of transposition of the great arteries (TGA) were analysed in a prospective study of 30 successive patients, from August 1996 to October 1999, who were presumed asymptomatic and investigated 10 years after surgery. All underwent clinical examination, ECG, stress Thallium 201 myocardial scintigraphy, Doppler echocardiography, Holter ECG, pulmonary perfusion scintigraphy, right and left cardiac catheterization with selective coronary angiography. Five patients had coronary lesions (4 thromboses and 1 coronary-pulmonary artery fistula). The other abnormalities observed were mild bilateral stenosis of the two pulmonary arteries (1 case), grade 1 aortic regurgitation (6 cases), including 1 case of aortic root dilatation. Type B to E coronary circulations (Yacoub classification) were not significantly correlated with coronary artery disease in this series (p = 0.06). For the diagnosis of these lesions, myocardial scintigraphy and Doppler echocardiographic detection of wall motion abnormalities had a sensitivity of 50% and respective specificities of 88% and 35%. Long-term results after anatomical repair of TGA are satisfactory. However, the high incidence of coronary lesions makes regular follow-up and systematic coronary angiography necessary in all children. PMID- 10858847 TI - [Dobutamine echocardiography in children after heart transplantation]. AB - Coronary disease of the transplanted heart is the principal cause limiting long term survival of patients and grafts. In view of the invasive nature of coronary angiography, dobutamine echocardiography has been proposed as a non-invasive diagnostic method for this disease. The aim of this study was to determine the feasibility and reliability of this investigation in transplanted children. Twenty-one echoes were performed with dobutamine infusions in 17 patients transplanted at 10 months to 16.9 years of age (average 8.4 years), and followed up 1.1 to 10.1 years (average 4.4 years): 4 were on antihypertensive drugs but none were treated by betablockers. Dobutamine echocardiography was performed according to the standard protocol used in adults. The maximal level was attained in all cases. No major side effects were observed. The maximal heart rate attained 57 to 89% of the theoretical maximal rate, an increase of 44 to 184% compared with the basal heart rate. The maximal systolic blood pressure rose to 120 to 194 mmHg, an increase of 8 to 109% compared with resting values. The contractility scores and segmental contractile index were normal in 18 cases, abnormal at the maximal level in 2 cases (hypokinesia of segments 8 and 9 and akinesia of segments 10 and 16 with an index of 1.2), abnormal at the lowest levels (hypokinesia of segment 7 with an index of 1.1) and maximal level (hypokinesia of segments 1 and 7 with an index of 1.2) in one case. These results were concordant with coronary angiography performed within 2 to 8 days of echocardiography, and considered as the diagnostic investigation of reference (sensitivity 75%, specificity 100%, positive predictive value 100% and negative predictive value 93%). The authors conclude that dobutamine echocardiography is a non-invasive method easily performed with low risk in transplanted children but its diagnostic performance in coronary disease of the transplanted heart should be confirmed in larger studies. PMID- 10858848 TI - [Follow-up of patients treated by conduits between the right ventricle and the pulmonary artery]. AB - Between 1981 and 1998, 77 right ventricle-pulmonary artery conduits were implanted in 67 patients (37 boys, 30 girls, average age 6.3 years; range: 3 months to 17 years). The diagnoses were transposition of the great arteries with ventricular septal defect and obstruction of the pulmonary outflow tract (N = 22), tetralogy of Fallot (N = 16), truncus arteriosus (N = 9), double outlet right ventricle with pulmonary stenosis or atresia (N = 8) and agenesis of the pulmonary valve with pulmonary stenosis (N = 2). The implanted conduits were homografts in 50 cases (43 aortic and 7 pulmonary), 11 valved Dacron grafts, 4 valved polystans grafts and 2 non-valved conduits. The average follow-up period was 3.6 years (range: 1 month to 17 years). Early death was observed in 8 patients (12%) and late death in 6 patients (9%). The 5, 10 and 15 year survival rates were 78.4%, 65.3%, and 65.3%, respectively. The conduits had to be replaced in 10 patients (15%). The non-replacement rate of all conduits at 5, 10 and 15 years was 81.4%; 40.7% and 40.7%, respectively. The causes of replacement were pure stenosis (54.5%), pure regurgitation (9%) and mixed stenosis and regurgitation (27.2%). These results are comparable to other published series showing a 15 year survival rate of 65% and a 59% reoperation rate at 15 years. Homografts have a slightly longer life compared with valved Dacron conduits but the difference is not statistically significant. PMID- 10858849 TI - [Percutaneous closure of patent ductus arteriosus with the Amplatzer duct occluder]. AB - BACKGROUND AND METHODS: Percutaneous closure of a moderate-to-large sized patent ductus arteriosus using multiple coils or a Rashkind double-umbrella may be technically demanding and raises a significant rate of embolization and residual shunting. This is why we tried a new self-expandable device called Amplatzer Duct Occluder to close transvenously large ductuses in eighteen patients at a median age of 2.8 years (range: 0.7 to 34). They had a patent ductus arteriosus with a minimal diameter > 3 mm (mean: 5 +/- 2 mm). The device had a mean diameter of 9 +/- 2 mm and was delivered through a 6F venous sheath. Aortography was done 5 to 10 minutes after the release and follow-up evaluation was performed with color Doppler echocardiography within 24 h, 1 and 3 to 6 months after the procedure. RESULTS: Placement was successful in all but the first patient (95%). Complete closure was achieved immediately in 4/17 patients (24%) and in 13/17 (76%) 24 h later. Two patients were lost for follow-up. Complete closure was confirmed in 14 of the 15 (93%) reviewed patients at 1 and 3 months. In all patients with a ductal diameter < or = 7.5 mm the closure was complete at 1 month. Embolization occurred in one patient with a 9.7 mm duct and surgery was undertaken. A mild and transient hematuria was observed in one patient. No pulmonary artery stenosis nor aortic obstruction were observed on follow-up. CONCLUSION: The Amplatzer Duct Occluder device offers a safe and effective option to treat patients with a ductus arteriosus which minimal diameter is up to 7.5 mm. PMID- 10858850 TI - [Three-dimensional reconstruction by transesophageal echocardiography of Amplatzer and CardioSEAL prosthetic devices after percutaneous closure and atrial septal defects]. AB - OBJECTIVE: The Amplatzer and cardioSEAL devices are currently used for percutaneous closure of atrial septal defects. The in vivo morphology of these devices is unknown and this is why we sought to describe their geometric profile by three-dimensional echocardiography. METHODS: Thirteen patients, aged 6 to 18, underwent transcatheter closure of a secundum type atrial septal defect with either the CardioSEAL (n = 8) or the Amplatzer (n = 5) device. Three-dimensional reconstructions of the defect were obtained from transoesophageal bidimensional echocardiographic views. RESULTS: In the Amplatzer group, the stretched diameter of the defect was larger than in the CardioSEAL group (22 +/- 2 vs 18 +/- 1 mm, p = 0.003), but ehocardiographic surfaces were the same (3.3 +/- 1 vs 3.6 +/- 1 cm2, p = 0.3). The surface of the CardioSEAL device was measured at 4.8 +/- 2 cm2 whereas the Amplatzer's surface was 6.8 +/- 2 cm2 (p = 0.03). The Amplatzer device had a volume of 9 +/- 1 cm3 while the CardioSeal's volume was 3.1 +/- 1 cm3 (p < 0.0001). CONCLUSIONS: The CardioSEAL device occupies a small volume but it covers less of the septal defect and this may result in a higher incidence of residual shunting. The Amplatzer device almost always occludes the septum, but it tends to bulge into the atria and may cause mechanical complications. PMID- 10858851 TI - [Cardiac pacing in children with breath-holding spells]. AB - Breath-holding spells are common and usually benign. However, the authors chose to implant a pacemaker in children presenting with severe symptoms. Over the last 15 years, 11 children with severe breath-holding spells were paced. All had reflex spells with loss of consciousness, spontaneously or after minor trauma, and 6 had seizures. All had a normal ECG with marked bradycardic responses to ocular pressure. The 24 hour ECG showed pauses (12-25 s) in 4 patients, sudden bradycardia (< 30/min) in 3 patients, and sinus arrhythmia in the remaining 4 patients. Medical treatment has been unsuccessful. Pacemaker implantation was decided because of the severity and/or the frequency of the episodes in 10 children, and because of intolerable familial anxiety in the other one. Age at implantation ranged from 14 months to 5.5 years (mean: 16.5 +/- 20 months). The device was implanted by an epicardial (7) or from an endocardial (4) approach. All had a ventricular demand device, except for one who was paced from the atrium. The results were spectacular, with disappearance of spells and restoration of normal activities. Holter monitoring showed normal function of the pacemakers. Recurrences were observed in 3 patients, either due to loss of capture (2 cases) or to the need for explantation because of cutaneous erosion. Follow-up ranged from 10 months to 14 years (mean: 7.9 +/- 4.2 years); 2 patients were lost to follow-up; 4 patients totally recovered and only 5 are still vagotonic. Two pacemakers have been changed at 13 and 15 years respectively. The authors conclude that although psycho-social factors play a part in breath holding spells, pacemaker implantation is very effective in suppressing symptoms in severely affected children. PMID- 10858852 TI - [Therapy and prognosis of infectious complete atrioventricular block in children]. AB - From 1983 to 1997, we have studied ten children with complete atrioventricular block likely due to myocarditis in order to assess its prognosis and to define a therapeutic strategy. Their age ranged from 6 days to 16 years (median: 4.1 years). All were admitted for sudden complete block, with symptoms in seven: syncope or fainting, seizures, collapse. Three had an asymptomatic bradycardia which was detected on routine auscultation in children with fever or already hospitalized; fever was present in 5. The disease was related to infection on biological data in 4 cases (1 listeriosis and 3 seroconversions for Epstein Barr or cytomegalic or Coxsackie B viruses), on a myocardial biposy in 1 case and on scintigraphic data in 1 case. In the remaining 4, indirect arguments were considered such as infectious context, normal recent ECG, favourable outcome. Five children were given intravenous isoprenalin with ventricular tachycardia in 3. Five were treated with steroids and 3 with specific antiviral agents. Seven patients were paced temporarily. One child died, 6 recovered totally and 3 have a permanent block with a definitive pacemaker implanted in 2. In conclusion, sudden acquired complete atrioventricular blocks are often ill-tolerated in children and have to be treated with transient pacing. Recovery occurs as a rule but some of these blocks may be definitive. Infective myocarditis is likely to be the cause of the disease even if the pathogen agent cannot always be identified. PMID- 10858853 TI - [Angelman syndrome and severe vagal hypertonia. Three pediatric case reports]. AB - Angelman's syndrome is an association of severe mental retardation with absence of language, ataxia, convulsions and hyperactive, joyful behaviour with frequent bouts of laughing. Genetic diagnosis is possible in about 80% of cases. No cardiovascular abnormalities have been described in this syndrome to date. The authors report the cases of three children with Angelman's syndrome who presented with severe malaise due to increased vagal tone. The age of onset of symptoms was between 20 months and 8 years. One of the children had malaises triggered by bouts of laughing. The diagnosis was confirmed in all three cases by the results of Holter 24 hour ECG recording and oculo-cardiac reflex. The treatment chosen was Diphemanil (Prantal) in the two patients under 2 years of age (after failure of a trial of betablockers in one case) and Disopyramide for the oldest child with excellent results in all cases. However, one child died suddenly at the age of 6, two years after stopping diphemanil. Based on these observations, the authors suggest that all malaises in patients with Angelman's syndrome should be investigated by Holter ECG and oculo-cardiac reflex (or tilt test). In view of the potential gravity of the syncopal attacks, long-term medical treatment seems to be justified. PMID- 10858854 TI - [Heart malformations and vascular complications associated with Turner's syndrome. Prospective study of 26 patients]. AB - Turner's syndrome is associated with congenital heart disease in a third of cases. Several reports of aortic dilatation and of death by dissection or rupture of the aorta have been published. The authors undertook a prospective study to assess the incidence of cardiac malformations and aortic dilatation in genetically confirmed Turner's syndrome. Twenty-six out of 34 patients recalled (76%), aged 7 to 30 years (average 17 +/- 6 years) accepted their inclusion in this study and underwent clinical examination, ECG, chest X-ray and echocardiography. Thirteen patients had a monosomy 45X and 13 a mosaic or structural abnormality. Six had a history of cardiovascular disease (operated coarctation: 2 cases, kinking: 2 cases, Hypertension: 2 cases). Eight patients (30%) had one or several anatomical cardiovascular abnormalities: bicuspid aortic valve (19.2%), abnormalities of the aortic isthmus (kinking or coarctation) (15.4%), aortic regurgitation (7.7%), mitral stenosis (3.8%), partial anomalous venous drainage (3.8%), patent ductus arteriosus (3.8%) and left superior vena cava (11.5%). Systematic evaluation of the aorta resulted in the diagnosis of dilatation of the ascending aorta in 1 case and dilatation of the sinus of Valsalva in 2 other cases. The authors conclude that echocardiographic evaluation is essential after the diagnosis of Turner's syndrome. It should be repeated regularly to detect dilatation of the aorta which carries the risk of serious complications, such as rupture or dissection of the aorta. PMID- 10858855 TI - [Acute myocarditis in children. Study of 11 clinical cases]. AB - The diagnosis of acute myocarditis in children is based on histological criteria. Often viral in origin, it results in acute left ventricular dysfunction, the clinical manifestations of which are very variable. The potential severity of the disease is maximal in its initial phase, justifying rapid and intensive treatment. Long-term outcome is relatively good although there is a risk of chronic left ventricular dysfunction. This retrospective study is based on 11 cases of acute myocarditis admitted to the paediatric unit of Clermont-Ferrand University Hospital between February 1989 and March 1999. The initial symptoms were non-specific. Echocardiography was the key diagnostic procedure. Half of the patients had severe cardiac failure requiring admission to the intensive care unit. Four cases presented with a severe complication: two embolic events, one syncopal atrioventricular block and one cardiac arrest. The cardiac treatment was classical (digitalis, diuretics, angiotensin converting enzyme inhibitors, anticoagulants). The aetiology was established in 3 cases (toxoplasmosis, haemolytic and uraemic syndrome, Kawasaki) and a viral cause was suspected in 6 other cases (adenovirus in 3 cases, herpes virus, RSV and enterovirus in 1 case). There were no deaths in the acute phase. The long-term outcome was globally good: complete regression in 8 cases, 1 chronic left ventricular dysfunction and 2 late deaths due to intractable cardiac failure. This short series illustrates the often misleading presentation of acute myocarditis in childhood, the value of systematic investigation in the hope of a specific treatment becoming available in the near future for the often viral aetiology. PMID- 10858856 TI - [Prenatal diagnosis of conotruncal heart diseases. Results in 337 cases]. AB - Prenatal diagnosis of conotroncal disease has developed by examination of the great arteries during morphological foetal ultrasonic scanning in the second term of pregnancy. This study reports the results of a series of 337 of these malformations diagnosed in the foetus over five and a half years. Their incidence with respect to all foetal cardiac malformations is 16.2%. Tetralogy of Fallot and pulmonary atresia with ventricular septal defect make up 56% of these cases, vascular malposition 16%, coarctation with or without interruption of the aortic arch 14%, truncus arteriosus 9% and agenesis of the pulmonary valves 5%. A karyotypic anomaly was found in 28 cases (8.2%) and, in the foetus with a normal standard karyotype, deletion of chromosome 22q11 was identified in 54 out of 237 cases. There was an associated polymalformative syndrome in 29 cases (8.3%). The training of obstetricians has increased the number of antenatal diagnoses of conotroncal abnormalities. Prenatal counselling should be given by a pluridisciplinary team because the association of these malformations with chromosomal abnormalities or with syndromes difficult to diagnose is common. PMID- 10858857 TI - [Clinical spectrum of prenatal tetralogy of Fallot]. AB - The aim of this study of 44 cases of tetralogy of Fallot was to assess the echocardiographic aspects and the prognosis with respect to associated abnormalities and the potential evolution in utero. Group I, tetralogy of Fallot with other abnormalities (N = 27: 2 valvular agenesis, 26.5 5.3 weeks), had genetic anomalies in 18 of the foetus (10 trisomies including 5 trisomy 21, 5 structural abnormalities including 2 micro-deletions 22q11 in the two cases of valvular agenesis, and one deletion of chromosome 8p23.1, 3 mendelian syndromes) and other abnormalities in 9 cases. Hypoplasia of the pulmonary artery was present in 60% of cases with a non-dilated aorta in 72%, infundibular hypertrophy in 33% and 2 evolutions to pulmonary atresia. Aspect of "isolated" ventricular septal defect were observed in 20% of cases. Survival was 10%. In Group II, tetralogy of Fallot was isolated (N = 17, including 2 pulmonary valve agenesis, 31 +/- 6 weeks) (p < 0.01 versus Group I). Pulmonary artery hypoplasia was observed in 50% of cases with dilatation of the aorta and infundibular hypertrophy in all and in one a postnatal progression towards pulmonary atresia. A correlation between growth of the pulmonary artery and gestational age was found in 5 foetus out of 9 studied sequentially (p between 0.03 and 0.007) and between age at first surgery and size of the pulmonary artery (r = 0.80, p = 0.001). Survival was 84%. The risk of malformation (61%) and the prenatal potential evolution of this disease justifies continuous follow-up of all cases of tetralogy of Fallot, high resolution karyotyping and postnatal evaluation in a specialized centre. PMID- 10858858 TI - [Cardiogenetics in the year 2000]. AB - This review article presents the up-to-date of knowledge progression during the past decade in the genetics of cardiopathies. The cardiopathies presented here have all a mendelian type of inheritance but this list is not exhaustive. PMID- 10858859 TI - [Pregnancy and anticoagulant therapy. Indications and strategies]. AB - Anticoagulant therapy is sometimes required during pregnancy either for the prevention of thromboembolic disease, for patients already on long-term antithrombotic treatment (for valvular prostheses) or for the prevention of complications of risk factors such as hereditary or acquired thrombophilia. Pregnancy is in itself a hyper-coagulable condition and the risk of thromboembolic complications is raised. Anticoagulation is a risk to the mother and to the foetus, and the management (heparin or vitamin K antagonists, respective doses) must be adapted to the underlying pathology and the stage of pregnancy. Mechanical valve prostheses are the most difficult problem and different strategies are proposed. The use of low molecular weight heparin may improve the outcome of these patients, but further trials are necessary. PMID- 10858860 TI - [Acute intravascular hemolysis after complete occlusion of a patent ductus arteriosus by detachable coils]. AB - A case of acute intravascular haemolysis after complete occlusion of a patent ductus arteriosus by detachable coils in a 10 months old child is reported. The child had a patent ductus arteriosus, a small secundum atrial defect, mild valvular pulmonary stenosis, and stenosis of branches of the pulmonary artery not diagnosed before closure of the ductus. Haemolysis developed several hours after complete occlusion of the ductus by two detachable coils. The mechanism of the haemolysis was thought to be the presence of metallic spirals in the left pulmonary artery, just beyond stenosis situated at the origin of this artery. Simple balloon dilatation of the left pulmonary artery stenosis resulted in the complete regression of haemolysis. PMID- 10858861 TI - [Hemolysis, a rare complication after percutaneous closure of an atrial septal defect]. AB - Haemolysis after closure of an atrial septal defect with an Amplatzer device is a rare complication. The suggested mechanism is the association of a residual shunt with flow of tricuspid regurgitation, impinging on the inferior edge of the prosthesis, factors which should be taken into consideration during the implantation of this type of device. PMID- 10858862 TI - [Severe arterial hypertension and pheochromocytoma in childhood. Case report and review of the literature]. AB - Pheochromocytoma was diagnosed in a 14 year old boy twenty-four months after the onset of symptoms. The diagnosis was made during a severe hypertensive crisis. Previously, the child had been followed up for delayed growth and functional psychiatric symptoms. After investigation, the tumour was removed by a trained surgical team. Pheochromocytoma usually arises from the adrenal gland and secretes catecholamines. It is less common in children than in adults. The presentation is very variable with hypertension as the main sign. The diagnosis is based on the finding of raised urinary catecholamines and CT scanning with scintigraphy for localising the tumour. These cases should be managed by teams familiar with this pathology because of the risk of catecholamine discharge, the need for complementary investigation of associated disease, the medical preoperative preparation and the specificity of the surgical intervention. PMID- 10858863 TI - [Osler endocarditis of a ventricular septal defect in a 21-month old child]. AB - The authors report a case of streptococcus mitis endocarditis of a ventricular septal defect in a 21 months old girl admitted for necrotic purpura of the lower limbs and a history of general ill health and pyrexia for two months. The severity of this case of endocarditis was illustrated by the importance of the vasculitis, the biological signs of infection (disseminated intravascular coagulation), and the size of the vegetation. The vegetation extended from the tricuspid valve as far as the pulmonary orifice which was partially obstructed, causing signs of right ventricular failure. The portal of entry was not found. Surgical ablation of the vegetation with tricuspid valvuloplasty was necessary after 48 hours of antibiotherapy. The outcome after one year's follow-up is good. The diagnosis of bacterial endocarditis must be considered even in very young children, especially those with congenital heart disease, and, in particular, ventricular septal defect. PMID- 10858864 TI - [Bundle of His tachycardia and chronic reciprocating rhythm: rare forms of prenatal tachycardia]. AB - In cases of permanent tachycardia, ante-natal diagnosis of chronic reciprocating rhythms with long RP' intervals or His bundle tachycardias is difficult. The authors report two cases of permanent foetal tachycardia with 1/1 atrioventricular conduction. In one case, the tachycardia rate was 170/min with anasarca treated by amiodarone in view of a family history of His bundle tachycardia. In the other case, the tachycardia rate was 200/min but with no signs of cardiac failure and was, therefore, not treated. The ECG at birth confirmed the diagnosis of His bundle tachycardia in the first case and identified a chronic reciprocating rhythm in the other. PMID- 10858865 TI - [Autosomal dominant Mendelian midline complex. Secundum atrial septal defect associated with cardiac and facial-thoracic defects. A familial case]. AB - The kindred of 38 individuals reported here have various anomalies: 1. facio thoracic malformations: hypertelorism, nasal deviation, cleft lip and palate, upper-incisors diastema and pectus excavatum; 2. cardiac anomalies: sinus node bradycardia, atrial fibrillation, nodal rhythm, atrial septal defect. Wolff Parkinson-White syndrome, low insertion of the septal tricuspid valve corresponding to an Ebstein syndrome, pulmonic "en dome" valve stenosis, aortic valve stenosis, long QT, and intraventricular conduction blocks. Almost all these defects are septal or para-septal. Mitral stenosis is probably rheumatoid. Such median varied pathology has not been yet reported. All the extra-cardiac anomalies are situated along the vertical upper half-body midline. All cardiac anomalies are in the septal or para-septal region. It is an autosomal dominant trait that implies the early embryonic development of the midline of cardiac and extra-cardiac structures. PMID- 10858866 TI - [Pseudo-inherited form of left heart obstructive defects revealing maternal phenylketonuria]. AB - If an adequate diet is not given to mothers with phenylketonuria, their offsprings often exhibit intra-uterine growth retardation with associated microcephaly and various malformations. Here, we report two families in whom we observed recurrent left heart malformations associated with microcephaly masquerading as a mendelian condition and revealing a maternal phenylketonuria. These observations suggest that when confronted to recurrent heart malformations with extra-cardiac defects that are not due either to an inherited chromosomal anomaly or to a well characterized mendelian disease, a maternal teratogen should be identified and more particularly maternal hyperphenylalaninemia if an intra uterine growth retardation or a microcephaly is part of the syndrome. PMID- 10858867 TI - [Causes of preoperative mortality in transposition of great vessels. 2 cases]. AB - The prognosis of transposition of the great arteries improved tremendously with the development of an early medico-surgical strategy including balloon atrioseptostomy, prostaglandin infusion and the arterial switch operation within the first days of life. Nevertheless, some patients still die preoperatively. We report on two newborn infants whose fatal outcome was promoted by an inadequate intercirculatory mixing. Since the diagnosis was not immediately made, the restrictive foramen ovale resulted very quickly in deep metabolic acidosis and balloon atrioseptostomy performed yet in the first hours of life could not prevent death. We emphasize the importance of prenatal echographic detection of this defect, only way to plan a balloon septostomy immediately after delivery in those infants suffering from inadequate atrial mixing. PMID- 10858868 TI - Biochemical activities of extracts from Hypericum perforatum L. 5th communication: dopamine-beta-hydroxylase-product quantification by HPLC and inhibition by hypericins and flavonoids. AB - Extracts from the herb "St. John's wort" (Hypericum perforatum L.) exhibit beneficial effects on patients suffering from mental depressions. Lack of catecholamine neurotransmitters may be one biochemical mechanism for this problem under discussion. It has been recently reported that alcoholic extracts from Hypericum perforatum inhibit dopamine-beta-hydroxylase (D-beta-H) with an I50 of 0.1 mumol/l on the basis of total hypericin content and with an I50 of 21 mumol/l with pure commercial hypericin. As test system polarographic determination of oxygen uptake with tyramine as a substrate analogue was used. In the present paper the quantification of the enzymatic activity and the potential influence of inhibitors are reported using dopamine as substrate and product (noradrenaline) quantification by HPLC. With this test system it could be shown that D-beta-H is strongly inhibited by pseudohypericin (I50 = approx. 3 mumol/l) and hypericin (I50 = approx. 5 mumol/l), whereas the I50-values of various flavonoids (quercitrin, isoquercitrin, hyperoside, rutin, quercetin, amentoflavone, kaempferol) are in the range of 50 mumol/l or higher. PMID- 10858869 TI - Evaluation of modulations in heart rate variability caused by a composition of herbal extracts. AB - The purpose of this prospective, placebo-controlled, randomized double-blind study was the examination of changes of the basic vegetative rhythms due to Cardiodoron, a composition of extracts of blossoms from Primula officinalis and Onopordon acanthium and from the herbs of Hyoscyamus niger. In its clinical use it is known as a modulating medicine in the treatment of functional disturbances of the cardiovascular system. With use of Holter monitoring, 24-h ECG recordings were obtained from 100 healthy subjects of whom 50 took the composition and 50 a placebo. Heart rate variability was evaluated from the 24-h ECGs by means of a power spectral analysis based on the Fast Fourier Transformation (FFT). Regulative influences on the rhythmic system due to the medicine were found. After four weeks of medication half of the verum group showed a tendency to an increased variability in the low and high frequency range at night (LFn: 0.04 0.15 Hz, HFn: 0.15-0.4 Hz) in contrast to the placebo group. The mean heart rate at night (HRn) showed a tendency towards a normalization in the verum group: in subjects with a low HRn the heart rate was increased and in subjects with a high HRn the heart rate was decreased. This effect could not be observed in the placebo group. After two further weeks without any medication this difference between verum and placebo was abolished. PMID- 10858870 TI - Effects of the oral antidiabetic repaglinide on the reproduction of rats. AB - Repaglinide (CAS 135062-02-1), a biguanide, is an orally available insulin secretagogue (beta-cell stimulant) and has been developed for the treatment of Type II diabetes. The developmental toxicity of the compound was investigated in rats. The effects on fertility, on embryo- and fetogenesis and on peri- and postnatal development were investigated. In a 'fertility study' 24 males were treated with 0, 1, 30, 300 mg/kg for 10 weeks prior to mating and during mating, and 24 females with 0, 1, 30, 80 mg/kg for 4 weeks prior to mating until gestation day 22 (hysterectomy group) or through gestation day 22 until postnatal day, i.e. lactation day 21 (littering group). In a 'teratogenicity study' with a hysterectomy group and a littering group included, 36 females were treated with 0, 0.5, 5 and 80 mg/kg from gestation day 7-16. In a 'peri-postnatal study' with a cross-foster group included 23 females were treated in the core study with 0, 0.5, 5, 30 and 80 mg/kg from gestation day 16 to lactation day 21 and in the cross-foster group 18 females with 0 and 80 mg/kg from gestation day 16 to lactation day 21. In a 'supplementary study' with five treatment windows 13 females each were treated with 80/30 mg/kg from gestation day 1-5 (W 1), gestation day 6-16 (W 2), gestation day 17-22 (W 3), lactation day 1-14 (W 4) and lactation day 15-21 (W 5). Effects of repaglinide on fertility, implantation, early and late embryogenesis, fetogenesis, birth and postnatal development including fertility of the offspring were investigated. In the 'fertility study' the NOTEL (no toxic effect level) for males was estimated to be 1 mg/kg and for females 30 mg/kg. In the 'teratogenicity study' the maternal NOTEL was 0.5 mg/kg as it was in the 'peri-postnatal study' 5 mg/kg. Food consumption and body weight gain of females were significantly reduced at the beginning of the respective treatment periods of the 'supplementary study' indicating a strong reaction of the dams to the treatment underlined by the death of individual animals (W 3, W 4 and W 5). Offspring survival during the last trimester of pregnancy and during lactation was affected in the 'fertility study' and in the 'peri-postnatal study' after 30 and 80 mg/kg and in the 'supplementary study' slightly in W 3 and more pronounced in W 4 and W 5. Changes of the skeleton in the extremities of the offspring were observed in all studies where the animals were exposed to repaglinide during late pregnancy (i.e. after completion of organogenesis) and/or lactation. At radiography the skeletal alterations comprised deformities of the coracoid process and acromion process, of the proximal humeral epiphysis, and of the epiphysis distalis and the condylus distalis of the femur. Deltoids of the humerus showed a slight increase of height and a length reduction. The radius and ulna were slightly bent. The most marked effects and the highest incidence were induced during the first half of lactation (W 4). As age of offspring increased the changes were more pronounced and occurred with a higher incidence. Correspondingly, ash weight, calcium, magnesium and phosphorus content of bones were reduced, but the proportions remained constant. Histopathological examination (supplementary study) showed that small fibrotic foci were formed in the area of dislocation of parts of the epiphyseal plate and that the remaining hyaline cartilage was thinner than normal (W 3, W 4 and W 5). Additionally, the longitudinal axis of the diaphysis juxtaposed to the growth zone was markedly bent, becoming convex to the lateral side. The studies clearly demonstrated that long bone development was not impaired during embryogenesis and early fetogenesis but after completition of organogenesis exclusively, indicating that repaglinide was not teratogenic. Effects of repaglinide were clearly effects on growth. The effects seen in all studies only occurred at excessively high plasma concentrations which will not be reached at human therapeutic dos PMID- 10858871 TI - Preclinical pharmacology of desloratadine, a selective and nonsedating histamine H1 receptor antagonist. 2nd communication: lack of central nervous system and cardiovascular effects. AB - Desloratadine (descarboethoxyloratadine, CAS 100643-71-8) is a selective histamine H1 antagonist that exhibits qualitatively similar pharmacodynamic activity to its parent, loratadine (CAS 79794-75-5), but is 2.5-4 times more potent orally. In studies of central nervous system (CNS) effects that might lead to sedation, desloratadine had no behavioral, neurological or autonomic effects in the conscious mouse and rat. At large multiples of the antihistaminic dose in the mouse, it did not inhibit convulsions caused by electroconvulsive shock and inhibited acetic acid-induced writhing only at a dose approximately 1,000 times the antihistaminic dose in the mouse. Desloratadine had no effects on blood pressure, heart rate or electrocardiographic parameters in the rat or guinea pig or on electrocardiographic parameters in the monkey. Notably, there was no effect on the corrected Q-wave to T-wave (QTc) interval. Desloratadine did not inhibit IKr channel human ether-a-go-go-related gene (HERG)-induced current in a study in which HERG was expressed in Xenopus oocytes. In the rat, desloratadine did not cause effects in urine volume, electrolytes or creatinine, or inhibit gastric emptying or intestinal transit, or cause any harmful effects on gastric mucosa. The results of these preclinical studies provide evidence that desloratadine is a safe antihistamine without CNS or cardiovascular effects. PMID- 10858872 TI - Modulation of nitric oxide synthesis in inflammation. Relationship to oxygen derived free radicals and prostaglandin synthesis. AB - The role of nitric oxide (NO) derived from constitutive (cNOS) and inducible (iNOS) nitric oxide synthases and its relationship to oxygen-derived free radicals and prostaglandins was investigated in two models of inflammation, namely, carrageenan granuloma air pouch (acute model) and Freund's adjuvant induced arthritis (chronic model) in rats. Inflammation was assessed by measurement of NO and prostaglandin E2 (PGE2) levels and the lysosomal leakage of the enzyme N-acetyl-B-D-glucosaminidase (NAG) into the exudate of the granuloma pouch 4 h after carrageenan injection. Evaluation of paw volume and determination of serum NO, lipid peroxide (LP), and PGE2 levels were used for the assessment of adjuvant-induced arthritis after either 4 days (early phase) or 16 days (late phase) of adjuvant injection. Results of the study showed that the administration of either NG-nitro-L-arginine methyl ester (L-NAME, non-selective cNOS/iNOS inhibitor) or aminoguanidine (AG, selective iNOS inhibitor), prior to carrageenan injection or during development of adjuvant arthritis, caused a significant reduction in NO and PGE2 levels and in the NAG activity of the granuloma inflammatory exudate, whereas decreases in paw volume and in serum NO level were noticed in the adjuvant model as related to untreated rats. Similar treatment with L-arginine failed to elaborate a significant change in the parameters measured. Other observations included: no noticeable differences between the results of early and late phases of adjuvant arthritis; no clear correlation between NO, LP and PGE2 levels in the adjuvant arthritis inflammation and inability of the NOS inhibitors to modify the levels of serum LP that is increased during adjuvant-induced arthritis. The data give further evidence that NO is implicated in the development of both acute and chronic inflammation and that NOS inhibitors have potential antiinflammatory activity. Further studies are required to unravel the mechanisms by which NO interacts with other mediators of inflammation. PMID- 10858873 TI - Responses of isolated normal human detrusor muscle to various spasmolytic drugs commonly used in the treatment of the overactive bladder. AB - The spasmolytic activity of flavoxate (CAS 15301-69-6) and the anticholinergic agents oxybutynin (CAS 5633-20-5), tolterodine (CAS 124937-51-5) and trospium chloride (CAS 10405-02-4), all of which are commonly utilized in the treatment of urinary incontinence, on muscarinic tension and electrically evoked contractions of isolated human detrusor smooth muscle strips was studied using the organ bath technique. Within the concentration ranges tested (trospium chloride 10(-11)-10( 6) mol/l, flavoxate and oxybutynin 10(-9)-10(-5) mol/l, tolterodine 10(-10)-10( 5) mol/l), each drug caused a concentration-dependent relaxation of the tension elicited by muscarinic stimulation and dose-dependently attenuated the contractions induced by electrical field stimulation (EFS). The effects of trospium chloride and tolterodine on carbachol-induced muscarinic tension were more pronounced than those of oxybutynin, while trospium chloride and oxybutynin were most effective in inhibiting the contractions induced by EFS. Flavoxate was significantly less effective than all other drugs tested. Regardless the individual drug concentrations needed for maximal efficacy, the potency of oxybutynin and flavoxate to reverse muscarinic tension and attenuate EFS-evoked contractions was almost comparable while tolterodine and trospium chloride were more effective in relaxing the muscarinic tension of the detrusor strip preparations than causing inhibition of EFS-induced contractions. Our results again underline the ratio for the use of nortropane analogues (trospium chloride) and phenylpropylamine cresols (tolterodine) in the treatment of frequency, urgency and urge incontinence secondary to an overactive bladder. PMID- 10858874 TI - Design, synthesis, and biological evaluation of aromatic ester prodrugs of 1-(3' hydroxypropyl)-2-methyl-3-hydroxypyridin-4-one (CP41) as orally active iron chelators. AB - In order to improve chelation efficacy and to minimise toxicity, eleven aromatic ester prodrugs of 1-(3'-hydroxypropyl)-2-methyl-3-hydroxypyridin-4-one (CP41) have been synthesised. The distribution coefficients of these ester prodrugs between 1-octanol and MOPS buffer pH 7.4 were measured together with their rates of hydrolysis at pH 2 and pH 7.4, in rat blood and liver homogenate. The biliary metabolic profiles of selected ester prodrugs were investigated in rats. The in vivo iron mobilisation efficacy of these ester prodrugs has been compared with that of the parent drug using a 59Fe-ferritin loaded rat model. The hydrolytic rates of these esters vary appreciably, esters with heteroaromatic acid moieties being less stable than the corresponding benzoyl analogues. Many prodrugs were found to enhance the ability of the parent hydroxypyridinone to facilitate 59Fe excretion, the optimal effect being observed with the 4-methylbenzoyl ester derivative 8d. However, not all prodrugs provide an increased efficacy, indicating that lipophilicity is not the only factor which influences drug efficacy. Furthermore no clear correlation between lipophilicity, susceptibility towards hydrolysis and efficacy was detected. PMID- 10858875 TI - Modulation of mistletoe (Viscum album L.) lectins cytotoxicity by carbohydrates and serum glycoproteins. AB - Three D-galactose and/or N-acetyl-D-galactosamine specific mistletoe lectins, ML I, ML II and ML III, were purified by affinity chromatography followed by cation exchange chromatography. These lectins were toxic for Molt 4 cells in culture at concentrations in the pg/ml range, ML III being the most cytotoxic. Carbohydrates able to bind to the B-chain of these lectins inhibited their toxic activity. The digalactosides Gal beta 1,2Gal beta-allyl and Gal beta 1,3Gal beta-allyl were 60 and 30 times, respectively, more potent than D-galactose in their ability to protect the cells from the ML I cytotoxicity. N-acetyl-D-galactosamine and rho nitrophenyl N-acetylgalactosamine protected mainly from the toxic effects of ML II and III. Protection from cytotoxicity varied in the same order as the affinity of the tested carbohydrates for lectins. Serum glycoproteins particularly haptoglobin, but also alpha 1-acid glycoprotein and transferrin, notably inhibited the cytotoxicity of the lectins. This effect was due to the binding of lectin to the sugar moiety of the glycoprotein because deglycosylated haptoglobin did not have a protective activity on Molt 4 cells. Inhibition of the cytotoxicity of lectins by serum glycoproteins explains why mistletoe extracts containing lectins can be administered to cancer patients without harmful effects. PMID- 10858876 TI - Synthesis and antileishmanial activity of new 1-(pyridin-2-yl)imidazolidin-2-ones derived from 2-amino-4,6-dimethylpyridine. AB - Derivatives of 2-amino-4,6-dimethylpyridine resulting from the integration of the amino function into a 2-imidazolidinone were synthetized via the corresponding 2 chloroethylurea. N3-benzylation, acylation or sulfonylation afforded the target compounds 6-14 which were evaluated for their in vitro and in vivo antileishmanial activity. Two compounds, the N3-benzyl derivative 7 and the N3 tolylsulfonyl derivative 14, exhibited potent inhibition against cultured extracellular promastigotes of Leishmania mexicana with IC50 comparable to that of the previously studied N-(4,6-dimethylpyridin-2-yl) furan-2-carboxamide 2: 32.4, 46 and 69 mumol/l, respectively. Experimentation of their activity against mice macrophage amastigotes pointed out that IC50 of imidazolidones 7 and 14 were 7 and 13-fold lower than that of amide 2: 13.7 and 89 mumol/l. In vivo evaluation in Balb/c mice, intradermally infested with Leishmania mexicana, confirmed that, in the lesion site, compound 14 was able to significantly reduce the parasite burden at a daily i.p. dose of 10 mg/kg. It was demonstrated that these N pyridinylimidazolidinones could act by interference with the parasite PLA2 activity. PMID- 10858877 TI - Subchronic toxicity of the new quinolone antibacterial agent irloxacin in beagle dogs. AB - The subchronic oral toxicity of the new quinolone antibacterial agent irloxacin (6-fluorine-7-(pyrrol-1-yl)-1-ethyl-1,4-dihydro-4-oxo-quinolone- 3-carboxylic acid, CAS 91524-15-1) in Beagle dogs was investigated in studies of 4 and 29 weeks of duration. In both studies animals received dosages of 10, 120 and 1400 mg/kg/d. Pale coloured faeces were seen on animals receiving 1400 mg/kg/d. Animals receiving 1400 mg/kg/d for 29 weeks showed an increased incidence of wax in the ears during the latter half of the treatment period, and one male and one female experienced transitory locomotive difficulties at the end of the first week of treatment. The liver was identified as the target organ for toxicity with presence of lipofuscin in the hepatocytes of animals receiving 120 or 1400 mg/kg/d for 29 weeks. Slight increases in liver weights were observed in animals receiving 120 or 1400 mg/kg/d for 4 weeks, and in all groups receiving irloxacin for 29 weeks. However, no histopathological findings were observed in the liver of animals receiving irloxacin for 4 weeks or those receiving 10 mg/kg/d for 29 weeks. Other relevant findings observed in the 29 week study were increased triglyceride, phospholipid and cholesterol levels in males receiving 120 mg/kg/d and animals receiving 1400 mg/kg/d, increased albumin and decreased beta-globulin concentrations in females receiving 1400 mg/kg/d, and prolonged activated partial thromboplastin time in animals receiving 1400 mg/kg/d. On the basis of the results obtained it is concluded that 10 mg/kg/d can be considered as the non toxic dose after 29 week oral administration of irloxacin in dogs. PMID- 10858878 TI - [Establishment and evaluation of microdilution assays for the in vitro sensitivity testing of Aspergillus fumigatus]. AB - For the in vitro sensitivity assessment of moulds different methods of micro- and macro-dilution assays as well as agar diffusion assays in different media formulations have been described so far. Until now, no standardized method has been preferred. The aim of the present study is to establish and to evaluate a microdilution assay for the in vitro chemosensitivity testing of Aspergillus fumigatus. This method should show a late onset of superficial growth and sporulation as well as a relative independency of the test inoculum. As reference substances, amphotericin B (CAS 1397-89-3) and 5-fluorocytosine (CAS 2022-85-7), which are both used for therapy of systemic mycoses, have been tested. Both substances show an excellent water solubility in liquid media. Fifty clinical isolates of A. fumigatus were used for the testings. The strains showed a better growth in Isosensitest (ISO)-medium than in Yeast-Nitrogen-Glucose (YNG)-medium. Growth was submers++ during the observation time, adherent to the bottom of the microtiter plate. Superficial growth with sporulation only occurred in a late stage of growth. Only a weak inoculum effect could be observed and the MIC (minimal inhibitory concentration) values determined macroscopically as well as using an ELISA reader showed to be almost comparable with a clearly defined breakpoint for the MIC discrimination. For amphotericin B, lower MIC values were observed in ISO-medium (0.25-2 micrograms/ml; median M = 1 microgram/ml) than in YNG-medium (0.25 microgram/ml; M = 1 microgram/ml). For 5-fluorocytosine, in both media MIC-values of > or = 32 micrograms/ml with a median of 64 micrograms/ml in ISO-medium and 128 micrograms/ml in YNG-medium were observed, respectively. With this test procedure, a standardized chemosensitivity testing can be performed for strains of A. fumigatus by a test system which is relatively easy to handle. These in vitro sensitivity data have to be correlated with clinical data as well as data obtained from animal studies. Only after this assessment final therapeutic consequences should be drawn from these in vitro findings. PMID- 10858879 TI - Ethical issues in enhancement: an introduction. PMID- 10858880 TI - Normal functioning and the treatment-enhancement distinction. PMID- 10858881 TI - Grand dreams of perfect people. PMID- 10858882 TI - Gene therapies and the pursuit of a better human. PMID- 10858883 TI - Are "genetic enhancements" really enhancements? PMID- 10858884 TI - Cosmetic surgery and the internal morality of medicine. PMID- 10858885 TI - The moral significance of the therapy-enhancement distinction in human genetics. PMID- 10858886 TI - CQ sources/bibliography. PMID- 10858887 TI - Consultation and discussion with other physicians in cases of requests for euthanasia and assisted suicide refused by family physicians. PMID- 10858888 TI - Patient autonomy and social fairness. PMID- 10858889 TI - Response to "Giving 'moral distress' a voice: ethical concerns among neonatal intensive care unit personnel" by Pam Hefferman and Steve Heilig and "Neonatal viability in the 1990s: held hostage by technology" by Jonathan Muraskas et al. (CQ Vol 8, no 2). Continuing the dialogue: resuscitation of marginally viable neonates. PMID- 10858890 TI - Telemedicine: a proposal for an ethical code. PMID- 10858891 TI - Physician-assisted suicide: where to draw the line? PMID- 10858892 TI - Post-exposure prophylaxis for HIV following possible sexual transmission: an ethical evaluation. PMID- 10858893 TI - Forming professional bioethicists: the program at the University of Tennessee, Knoxville. PMID- 10858894 TI - Hospital ethics committees in Paris. PMID- 10858895 TI - Ram Dass on being a patient. Interview by Steve Heilig. PMID- 10858896 TI - Pediatric reconstructive surgery. AB - The field of reconstructive pediatric urologic surgery is constantly changing. Recent changes in pediatric urologic reconstructive surgery are discussed in the present review. Surgical techniques for treating patients with hypospadias, exstrophy, incontinence, and ambiguous or variant genitalia are also discussed. PMID- 10858897 TI - Biomaterials in functional reconstruction. AB - Recent initiatives in the development of biomaterials for functional reconstruction involve the alloplasts, the biological and the bioengineered biomaterials. Anti-infective alloplastic biomaterials (Foley catheters coated with rifampicin/minocycline bonded to silicone or ciprofloxacin liposome containing hydrogel) allow a reduction in the rate of bacterial contamination, but the risk of future bacterial resistance is a matter for concern. New generations of biologic collagen-based tissue-matrix grafts are derived from bladder (bladder acellular matrix graft and bladder submucosa collagen matrix), ureter or small intestine (subintestinal submucosa). There are high hopes that these materials may have applications in augmentation cystoplasty. Using tissue engineering (autologous cells expanded in vitro and grafted onto biodegradable matrix), biocompatible malleable penile prostheses have been obtained experimentally. Most of the results obtained with these new biomaterials are exclusively experimental, but they offer great hope for future functional reconstruction of the urinary tract. PMID- 10858898 TI - Principles of ureteric reconstruction. AB - The principles of ureteric reconstruction are not different from those of reconstructive urology in the rest of the urinary system. The importance of ensuring good vascular supply, complete excision of pathological lesions, good drainage and a wide spatulated and tension-free anastomosis of mucosa to mucosa remain paramount. Although time of diagnosis is the most single most adverse factor affecting outcome, the majority of ureteric injuries still present postoperatively, and delays in diagnosis are the rule rather than the exception. Successful management requires early and definitive intervention using endoscopic means or percutaneous drainage and stenting where possible. Failing this, a number of open surgical options to foreshorten the course of the ureter should be implemented. Most ureteric injuries below the pelvic brim can be treated easily with a ureteroneocystostomy using a bladder elongation procedure or a Boari flap. Mid and upper ureteric injuries above the pelvic brim, however, can be repaired with a spatulated ureteroureterostomy if the defect is small. In those with extensive ureteral loss, measures such as mobilizing the kidney, transureteroureterostomy, renal autotransplantation and ureteral substitution using small bowel may be required. Artificial ureteral substitutes may be an alternative in selected cases. PMID- 10858899 TI - Bladder reconstruction: a critical reappraisal. AB - The operative management of muscle invasive bladder cancer has been dramatically advanced by the advent of orthotopic reconstruction. Several studies reported during the past year have further demonstrated the utility of this form of urinary diversion. The long-term safety and efficacy of bladder replacement with respect to both surgical and metabolic complications has been demonstrated. As series include more patients with sufficient follow up, we are gaining a better appreciation of the results of treatment that patients and urologists can expect in terms of function and risk of complications. Taken together, published studies have made the following important points. Exenterative surgery as currently performed alters pelvic floor/urethral physiology. Early reports of complications in studies with short periods of follow up are not meaningful. When basic principles and complication rates are established for a procedure in the long term, sufficient follow up is required before it can be established that a modification to that procedure really is better. The occurrence of retention in a female patient with orthotopic bladder continues to be poorly understood. PMID- 10858900 TI - The contemporary role of the artificial urinary sphincter. AB - The artificial urinary sphincter has been in use for more than 25 years as a treatment for urinary incontinence due to intrinsic sphincter deficiency. Recent clinical studies have increased our knowledge concerning its use in children and in adult patient populations. Particularly, more long-term data and data on the combination of the artificial urinary sphincter with reconstructive techniques that utilize the bowel have become available. Some interesting new ideas regarding changes in the design of the artificial urinary sphincter have been advanced. PMID- 10858901 TI - Time for a neoadjuvant chemotherapy strategy in prostate cancer? PMID- 10858902 TI - Salvage radical prostatectomy for radio-recurrent prostate cancer: is this a viable option? AB - The role of salvage prostatectomy for radio-recurrent prostate cancer remains unclear. Recurrent prostate cancer after radiation therapy is in many cases biologically aggressive. It is unclear whether the biologic aggressiveness of radio-recurrent prostate cancer is due to time-dependent cancer clonal evolution (potentially induced by radiation damage), or is due to an innately aggressive tumor secondary to overexpression or mutation of apoptotic inhibitors that render these tumors resistant to radiation. Recent studies examined the role of DNA ploidy, p53 and bcl-2 expression, proliferative indices and glutathione S transferase-pi in predicting response to radiation therapy or salvage prostatectomy. Because of the potential for significant morbidity after salvage prostatectomy, preoperative parameters that aid in the identification of the patients who are most likely to benefit from surgery are needed. PMID- 10858903 TI - Endoscopic management of urologic disease with the holmium laser. AB - The efficiency and safety profile of the holmium laser have made this tool a versatile multipurpose instrument for use in the endoscopic treatment of a wide variety of urologic disorders. Herein are reviewed holmium laser physics and current endourologic applications, as well as the performance of new low-power holmium lasers. PMID- 10858904 TI - Intraoperative cavernous nerve stimulation during nerve sparing radical prostatectomy: how and when? AB - Although nerve-sparing prostatectomy is widely practiced, the results with respect to potency preservation often do not meet expectations. The concept of intraoperative cavernous nerve stimulation is rational. Recent data that link the response to sildenafil after prostatectomy to bilateral nerve sparing has increased the importance of optimizing nerve sparing. The cavernous nerves are often difficult to visualize and may have a variable course. A tumescent response to nerve stimulation can be consistently demonstrated. The response may be subtle, and characterized by a minimal increase in penile circumference and blood flow. Immediately after prostectomy, proximal nerve stimulation identifies whether neural continuity has been maintained, and is predictive of recovery of erectile function. A new device, the Cavermap, has been developed to permit intraoperative nerve stimulation with tumescence monitoring. An initial phase 2 and subsequent phase 3 single blinded, randomized, multicenter study that compared Cavermap-assisted prostatectomy with conventional nerve sparing demonstrated a significant benefit in terms of the duration of nocturnal tumescence by Rigiscan (Timm Medical Technologies, Eden Prairie, Minnesota) at 1 year. Other approaches are being explored, including sural nerve grafting, use of nerve stimulation during cystectomy or abdominal-perineal resection, and direct corpus cavernosum pressure monitoring during nerve stimulation. These approaches warrant further evaluation. PMID- 10858905 TI - Surgical management of intrinsic sphincter deficiency in women. AB - The past decade has witnessed significant changes not only in our understanding of intrinsic sphincter deficiency, but also in our surgical approach to this problem. It became evident that the patient's medical condition, expectations, and degree of incontinence should define the approach in order to make the greatest impact on quality of life. The present review describes the current concepts and surgical approaches to intrinsic sphincter deficiency, namely slings, injectables, and artificial sphincters. PMID- 10858906 TI - Bibliography. Current world literature. Functional reconstruction and trauma. PMID- 10858907 TI - Bibliography. Current world literature. Urological surgery, surgical techniques, technology. PMID- 10858908 TI - [50 years lithium treatment]. AB - Answers have been sought to the following questions of current interest: Does lithium have a prophylactic action? The claim that such an action has never been demonstrated was based on wrong assumptions, biased selection of references, and unjustified generalizations. What effect does long-term lithium treatment have on the patients' suicidal behavior? There is a close association between long-term lithium treatment, on the one hand, and lowered mortality and reduced suicidal behavior, on the other; no such association has been demonstrated for other mood stabilizers. What are the effects of lithium when given during pregnancy and lactation? The risk/benefit ratio of lithium is lower than that of the anticonvulsant drugs. Are new drugs about to oust lithium? Among contending drugs the anticonvulsants carbamazepine and valproate have attracted particular interest, but the evidence for a prophylactic action in typical bipolar disorder is weak or missing. What are the requirements of the designs of future comparative trials? In order to prove new drugs prophylactically superior to lithium, trial designs must be such that they provide valid information; some proposals are outlined here. Which drug or drugs should be used for prophylactic treatment? In addition to prophylactic superiority over lithium the new drugs must be as good as or better than lithium as regards a number of other properties, including risk/benefit ratio during pregnancy and lactation, and a close association between long-term treatment and lowered suicidal behavior. The most urgent problem at the present time is to compare in a just and balanced manner the prophylactic efficacy and other relevant properties of new drugs with those of lithium. Many manic-depressive patients' health and sometimes life depend on whether at any time they are given the best prophylactic drug available. PMID- 10858909 TI - [Evaluating the quality of life of 42 heart transplant patients and candidates: a cross-sectional study]. AB - OBJECTIVE: Although evaluation of heart transplant candidates and recipients is usually based on objective clinical variables, self-assessment has been proved to be an important component of treatment evaluation. Quality of life is a multidimensional concept, a mix of objective and subjective measures, that could reflect the adjustment to the illness and its treatment. Few studies have reported on quality of life in heart transplantation candidates. This exploratory study, conducted in Bordeaux (France), was designed to assess both objectively and subjectively the quality of life in heart transplant candidates and recipients and to determine the relationships between subjective and objective variables. METHOD: The assessment was cross-sectional; 21 candidates evaluated at an average of 10 (Sd 21.4) months into the waiting period, were matched with 21 recipients at 29.5 months post operative. Subjective evaluation of the quality of life was self-assessed by the Tableau d'evaluation assistee de la qualite de la vie (TEAQV) and the Nottingham Health Profile (NHP). A semi structured psychiatric interview, and the NYHA (New York Heart Association) cardiac insufficiency score provided objective measurements. RESULTS: The NHP and TEAQV mean scores were not not significantly different between the two groups: candidates (C) and recipients (R) reported similar subjective data regarding positive quality of life experience. The objective data indicated significant disadvantages for the candidate group: the cardiac insufficiency score was worse in the candidates [(NYHA mean score: (C) = 2.7 Sd 0.56 vs (R) = 0.7 Sd 0.8, t de Student p < 0.01)] and the DSM III-R axis 1 diagnoses were more frequent in the candidates [(C) = 16/21 vs (R) = 9/21 Chi2 p < 0.05)]. There was a prevalence of adjustment disorders in the candidates. Significant correlations were found between NYHA and NHP mean scores (r = 0.6, p < 0.01) and NYHA and physical and psychological dimensions of the TEAQV (r = -0.65 and r = -0.55, p < 0.01) in the recipient group. In the candidate group, no correlation was found between these scores. CONCLUSION: In the recipient group, objective and subjective assessment showed greater concordance than in the candidate group. Despite more objective physical and moderate yet frequent psychiatric complications, the candidate group reported as positively as did the recipients upon the quality of their life experience. This could be the result of psychological adaptation to the stressful situation. These data were in accordance to several earlier reports. However, the literature has remained controversial upon the evaluation of the quality of life of the candidates. The results of this study, limited by some methodological bias (the small number of patients assessed), need to be confirmed in a prospective study. PMID- 10858910 TI - [Cognitive inhibition and psychopathology: toward a less simplistic conceptualization]. AB - "Cognitive inhibition" is a concept that has found a firm place in the interpretation of performance by normal subjects on tasks involving adherence to a plan and suppression of incorrect responses to distractors. The presence of "negative priming" is the classical indicator of cognitive inhibition. Negative priming occurs when, in a sequence of stimuli each of which is composed of a target and a distractor, the distractor of the first stimulus becomes the target of the second stimulus: reaction time to the second stimulus is slowed because of the inhibition applied to the distractor of the first stimulus. The concept has been extended to the interpretation of pathological behavior and symptoms. Pathological subjects have been found to show deficient negative priming. Thus, negative ideation in depression as well as intrusive paranoid associations in schizophrenia have been related to a deficit in the capacity to inhibit inappropriate representations. In this paper, we briefly review some of the experimental evidence from normal subjects that has contributed to the acceptance of cognitive inhibition as a key process in the control of normal cognition, as well as more recent evidence that has led to a revision of the concept. Negative priming in normal subjects has been found to be dependent upon characteristics of the experimental situation as perceived by the subject. In particular, priming is observed when the subject anticipates difficulty in determining the response and proceeds with caution. Thus, inhibition is not an automatic "brake" applied to irrelevant material, but rather the product of strategic considerations within the experimental situation. This revision of the cognitive inhibition hypothesis leads to a re-interpretation of the apparently deficient cognitive inhibition seen in depressed or schizophrenic subjects. According to this more recent interpretation, deficient cognitive inhibition in pathological subjects can be seen as a less adaptive strategic adjustment to the task. The pathology seems to touch higher-level executive functions rather than a deficient inhibitory "brake". In depressed subjects, abnormal performance in selective attention tasks could be related to the underlying pathology in two ways: some depressed subjects show a marked lack of energy and a psychomotor slowing: these subjects do not exhibit normal negative priming, probably because of a reduction of cognitive resources. Other depressed subjects show abnormal performance as reflected by negative priming greater than normal: this result could be related to an exaggerated tendency to verify a correct response. Schizophrenic subjects show a lack of negative priming that seems most plausibly to be related to an ineffectual integration of the experimental instructions concerning both speed and accuracy in the response. This re-interpretation of the cognitive deficiency in pathological patients provides a better fit with recent experimental results from normal subjects, and with cognitive deficits measured in pathological subjects. PMID- 10858911 TI - [Psychiatric disorders induced by drug dependence other than alcohol]. AB - Most of psychoactive substances abuse or dependence disorders are associated to another psychiatric disorders. Depression, anxiety and psychotic disorders are the more frequent comorbid disorders. Psychiatric comorbidity is induced by acute consumption of psychoactive agents, chronic consumption or withdrawal. Psychiatric disorders are more frequent when patient are assessed immediately after the withdrawal. Main biological factors implicated in the pathophysiology of psychiatric disorders associated to dependence disorders are: increase in norepinephrine activity, during withdrawal, activation of locus coeruleus, kinding induced by repeated withdrawals. Psychotic disorders in opiate dependent patients can be induced by withdrawals. These psychotic disorders are more often described after methadone discontinuation. Consumption of cocaine can provocate paranoid delusions. Phenylcyclidine provocates sensorial distortion or delusive disorders resembling schizophrenia. Flash backs, following withdrawal realized brief and transient psychotic disorders. They can occur up to one year after the end of the intoxication. The occurrence of depression in dependent patients is frequent. Depressed patients are at risk for suicide. Retrospective studies showed that near of 40% of the subjects died from suicide have presented alcohol or drug abuse or dependence. Withdrawal from opiates provocates depression. Clinical picture included apathy, blunting of the affects, sadness and loss of interest. Cocaine consumption provocates manic-like disinhibition at the beginning of the intoxication. Long term consumption and withdrawal increase the risk of depression. PMID- 10858912 TI - [Arterial hypertension and anxiety in primary care]. AB - Various forms of hypertension and anxiety are two very frequent reasons for consultations in primary care. Prevalence of comorbidity HTA/anxiety is unknown in France. Practically, general practitioners meet sometime the association of both typical pathologies. More often, they wonder about the interpretation of high blood pressure figures with an anxious patient, and inversely about the responsibility of the hypertension (with the suite of phantasms) with the apparition of anxious symptoms. Both these last clinical situations are the more frequent in general practice. The global clinical approach, the listening, the time use as help for diagnosis, the use of personal patient's characteristics and his/her environment more often allow to distinguish and to propose an individual therapeutic strategy. PMID- 10858913 TI - [Diagnosis and management of panic disorder in psychiatry (PANDA Study)]. AB - Panic disorder is a genuine public health problem given by their frequency and the various and repeated consultations that they involve. PD is underdiagnosed in primary care and in medical specialist. A public campaign might lead to improved diagnosis and better treatment of panic disorder, with a beneficial effect on medico-economic indicators. Intervention by the psychiatrist is of key importance, although it has not been evaluated to any great extend. The objective of the PANDA study was to look at the prevalence and diagnostic of panic disorder, the conditions of access to and use of care, as well as the method of treatment. Four hundred and twenty three psychiatrists participated in the study and 8,137 patients seen consecutively were included. The prevalence of actual panic disorder evaluated using the Mini International Neuropsychiatric Interview (MINI) systematic is 9%. In two third of cases coexisted agoraphobia and in one third a depression. Eighty six percent of patients with actual panic disorder were treated by the psychiatrists. The diagnosis and suitable treatment of panic disorder would appear to be a significant objective in term of public health, leading to a reduction in medical and social cost of this disorder. PMID- 10858914 TI - [Early signs of autism and family films: a new study by informed evaluators and those unaware of the diagnosis]. AB - Since the initial individualization of infantile autism by Kanner, subsequent work has attempted to define the age at which disorders appear, their symptomatology and their specificity. Initially, retrospective studies based on questionnaires and interviews with parents were conducted in order to determine the age of at which the first signs appeared. Combined with interviews, clinical observations have provided incontestable aid for describing the early signs of autism. The study of home movies taken by parents before their infant's disorders were recognized has led to a new approach to the initials signs of autism. Our study is a continuation of work in our Child Psychiatry Unit under way since 1984. The aim of this work is the symptomatological and comparative analysis of home movies of 14 autistic and 10 normal infants, during the first two years of their life. Each film was scored for the 0-8, 9-17 and 18-24 month periods. Based on the use of the Infant Behavioral Summarized Evaluation (IBSE), this study confirms prior data and also shows the emergence of very early disorders, perceptible within the first few months even by blind evaluators: a docile baby, showing no overt manifestations, not seeking contact, with an absence of pre language. Even so, the results require caution when interpreting for methodological reasons which are discussed. PMID- 10858915 TI - [Methylphenidate, tics and compulsions]. AB - Obsessive compulsive (OC) symptoms following methylphenidate (MPH) administration are seldom reported and usually not even mentioned among its adverse effects. We report here a case of MPH-induced OC symptoms which began ten months after the treatment was initiated and that were exacerbated 14 months later. This delay and the symptoms fluctuations might lead out of the right diagnostic though tic occurrence could then correct. Cross-sensitization between stress and psychostimulants might as well explain the long delayed onset of obsessive compulsive symptoms as their fluctuation both along time and contexts. PMID- 10858916 TI - [Vulnerability to schizophrenia. III: Importance and limits of the Identical Pairs Continuous Performance Test]. AB - Since several investigations have shown attentional deficits both in patients with schizophrenia and in subjects at high risk for schizophrenia, these deficits could be valuable vulnerability markers for schizophrenia. The aim of this study was to investigate wether non psychotic relatives of schizophrenic probands have deficits in sustained attention as measured by the Continuous Performance Test, Identical Pairs (CPT-IP) version. The study subjects were 25 schizophrenic probands, 50 of their first-degree relatives and 46 normal controls. For each subject, attention was assessed during 6 experimental conditions (2 standard, 2 slow, 2 easy conditions) of visual stimuli (digit-numbers and shapes). In each of the six conditions, the value of the sensitive index d' in the first-degree relative group was at an intermediate level between the patient and control groups. Moreover, in the standard shape condition, the d' value was significantly lower in the schizophrenic and in the relative groups than in the control group. This deficit was all the more interesting since it was not explained by a deficit in general intellectual abilities or by psychopathology such as anxiety or depression. Furthermore, the schizophrenic patients made more random errors in the standard and in the slow number conditions than both other groups. All groups improved their performance when the stimulus duration increased and when the processing load decreased. As a conclusion, this investigation seems: 1) to confirm the existence of an attentional deficit in the first-degree relatives of patients with schizophrenia; 2) to demonstrate the interest of the CPT-IP to detect this deficit. It must be emphasized that in order to detect the deficit, one only needs to explore the standard shape condition and that under such circumstances, the CPT-IP test has the advantage of being less time consuming than tests used in previous studies. PMID- 10858917 TI - [Deficit in suppression of interference in visual information processing by schizophrenic subjects]. AB - Although many studies have indicated information processing deficits in schizophrenic patients, the precise nature and underlying causes of these deficits remain largely uncertain. One prominent hypothesis is that these patients show insufficient attentional inhibition. This deficit to inhibition has been linked to certain cognitive disorders in schizophrenic patients, including attention deficits, as well as to some clinical symptoms, especially those involving delusional thought, hallucinations,and poor contact with reality. The hypothesis of deficient attentional inhibition, although attractive in some ways, is difficult to work with, because it is not easy to directly measure "attentional inhibition". Several studies involving normal subjects have linked attentional inhibition with performance on a task demanding the suppression of distracting information: the presumption is that efficient attentional inhibition will permit rapid responses because the distracting information will be quickly suppressed, allowing undistracted processing of the target information. The present study measures schizophrenic patients' performance on a task demanding suppression of rapidly-presented visual information. An important methodological feature of this study is that performance is measured in terms of "percent correct responses" rather than the reaction time measures typically used in tasks demanding distractor suppression, such as Stroop-like selective attention tasks. Since reaction times are not considered, the results cannot be interpreted in terms of deficient response organization and execution. Schizophrenic (18) and normal (18) subjects underwent trials in which a visual target was the second of two stimuli presented in rapid succession. Interference produced by a non-target significantly impaired perception of the target for schizophrenic patients. This effect persisted longer in the schizophrenic subjects possibly because of deficient attentional inhibition. PMID- 10858918 TI - [Addictions and action systems]. AB - Generalizing from some previous analyses of addiction, and introducing the concept of an action system which governs all actions which are focussed on what Brown (1988) calls "hedonic management", we argue that addictions of every kind involve an action system that displays high salience, low variety and low vicariance. Addictions also involve what Apter (1982) calls the "paratelic state". A study was carried out comparing 31 drug addicts with 29 control subjects in terms of action system variables. To measure these variables, we constructed a new instrument, the Activity-System Drawing Test, and also used the Telic Dominance Scale to measure frequency of paratelic states. Dysphoria was measured by means of the BATE (anxiety), IDA-13 (depression), SEI (self-esteem), and TAS-20 (alexithymia) instruments. Strongly significant differences were found between groups for both action system variables and dysphoria, and there were also strong correlations between both groups of variables. This supports the idea that addictions emerge from systemic properties of the action system. PMID- 10858919 TI - [Effects of sulbutiamine (Arcalion 200) on psycho-behavioral inhibition in major depressive episodes]. AB - Psycho-behavioural inhibition is characteristic of major depressive disorder and frequently recedes after the other depressive symptoms. This may induce an important psychosocial impairment which could be a risk factor for relapse. METHODS: The aim of this eight weeks, multicentric, randomized, double blind, placebo controlled trial was to assess the efficacy and safety of sulbutiamine (Arcalion) [600 mg p.d.] on the symptoms of psycho-behavioural inhibition of inpatients with DSM III-R defined Major Depressive Episode (MDE) treated by adjusted doses of clomipramine [75 to 150 mg pd]. Moderate doses of hypnotics and anxiolytics without potential activity on the mood were authorized during the trial. The MDE was assessed with the MADRS, HAM-A and CGI scales. Patients who did not respond adequately to the antidepressant treatment were prematurely withdrawn from the trial. The three Sheehan Disability Scales (SDS), the Norris Visual Analogue Scale (VAS) and the Depressive Psychomotor Retardation Scale (ERD) were used to monitor psycho-behavioural inhibition. RESULTS: The mean intake scores were, as expected, fairly high: MADRS (32), HAMA-A (23), CGI (5) and ERD (27). The SDS and EVA scores showed that the patients felt severely handicapped in their social, professional and family life functioning as well as in their emotional, affective, cognitive and behavioural performances. At four weeks the MADRS, HAM-A and CGI scores indicated that the global improvement of the MDE was comparable in both treatment groups. However, the scores at the EVA and SDS scales showed that the patients treated with sulbutiamine were significantly less incapacitated than the placebo group in all of the various facets (affective, cognitive, emotional, behavioural) of psycho-behavioural inhibition. Furthermore, the safety data shows that both treatment groups were comparable and in particular that sulbutiamine had not induced any inappropriate behaviour, including suicide attempts, or mania. CONCLUSIONS: Sulbutiamine has no antidepressive effect but it can hasten the resorption of psycho-behavioural inhibition occurring during major depressive disorder and thereby facilitate the rehabilitation of patients in their social, professional and family life functioning. PMID- 10858920 TI - [Follow-up study of 3 years group therapy with lithium treatment]. AB - Group psycho-education for bipolar patients has been shown to increase patient self-esteem and to cut down on the number of re-admissions. For the purposes of this study, a comparison was made between a 10-sessions group programme, without additional written information (26 patients) and a highly structured 5-sessions group programme which did include written information (15 patients). Measurements were done before and after group therapy over a 3-year follow-up period. In the 10-sessions programme there was evidence of improved patient self-esteem; patients in both programmes reported that they had benefitted from them and that subsequently they had fewer psychosocial problems. In the 3-year follow-up period, there was a significant decrease in the non-compliance (from 54 and 47 percent to 31 and 33 respectively) and hospitalization ratio in both programme groups. The ratio of hospitalizations in the first and second programmes was: 0.33 and 0.25 a year previously; 0.25 and 0.26 respectively whilst taking lithium; and 0.08 and 0.09 at the end of the group education of 3-year follow-up period. PMID- 10858921 TI - [Schizophrenic with obsessive-compulsive disorder or symptoms]. AB - The concept of pseudo-obsessive schizophrenia has been often used in the past. Clinically, severe obsessive-compulsive disorders (OCD) are closed from psychotic symptoms and ask questions about differential diagnosis with schizophrenia. Moreover some characterized schizophrenia may present in some cases obsessive compulsive symptoms (OCS). Finally, schizophrenia treated by atypical antipsychotics can be complicated by obsessive compulsive symptoms following the onset of the drug. Until now, there have been no control trials on this specific topic. Reviewing data of studies exploring the prevalence of OCD in schizophrenia this current article summarizes the different pharmaceutical approaches used in treating this disorder. In addition, a review about antipsychotics causing either emergence or exacerbation of OCS is presented. PMID- 10858922 TI - [New therapeutic strategies in schizophrenia: apropos of acute decompensation treated with olanzapine]. AB - Mr B is a 32-year-old patient with a disorganized type of schizophrenia and his treatment, started when he was 20, has always been difficult and his social integration has become progressively worse. In 1998, treatment with olanzapine was started when he suffered acute decompensation on top of his disorganized condition which had an adverse effect on his symptoms. In a short space of time, a very satisfactory improvement was seen, not only in his symptoms but also in his social integration. This improvement has been sustained throughout two and a half years of follow-up. The perspectives offered by the new antipsychotic drugs in the management of acute episodes of schizophrenia are discussed in the light of recent data from the literature and lead to a reconsideration of our therapeutic practices. PMID- 10858923 TI - [Depression and neurology]. PMID- 10858924 TI - [Unusual disorder: CADASIL]. PMID- 10858925 TI - [Migraine and depression]. PMID- 10858926 TI - [Is there a "migraine personality"?]. PMID- 10858927 TI - [Depression after cerebrovascular accident (CVA)]. PMID- 10858928 TI - [Depression and Parkinson disease]. PMID- 10858929 TI - [The symptoms of Parkinson disease]. PMID- 10858930 TI - [Asymptomatic "child" of the parent affected by Huntington disease]. PMID- 10858931 TI - [Alzheimer's disease and general medicine: what are the conditions for the early diagnosis and efficient treatment?]. PMID- 10858932 TI - [Epilepsy and depression]. PMID- 10858933 TI - [Depression, cognition and brain imaging]. PMID- 10858934 TI - [Systemic view of Alzheimer's disease: the wholeness as the indicator of the disorder]. PMID- 10858935 TI - Understanding ethnic variation in pregnancy-related care in rural Guatemala. AB - OBJECTIVES: This study examines the relatively low use of modern pregnancy related care in Guatemala, especially among indigenous women, and explores the role of socioeconomic status, social and cultural variables, and access to biomedical health facilities in accounting for ethnic differences in care. METHODS: The data for the analysis come from the Guatemalan Survey of Family Health--a population-based survey of rural women that contains detailed data on care received during pregnancy and delivery along with extensive background information. Binomial and multinomial logit models are used to identify the variables that affect the likelihood of receiving different types of care during pregnancy and delivering in a medical facility and the extent to which sociocultural factors and measures of access account for the observed ethnic differences. RESULTS: The estimates not only confirm previous findings of a large ethnic difference in the use of modern pregnancy-related care, but also extend them by identifying a gradient within the indigenous population. The analysis demonstrates that, in general, sociocultural variables are more strongly associated with modern pregnancy-related care than are measures of access and that the former variables explain more of the ethnic variation in care than the latter. The results also demonstrate that pregnant women, especially indigenous women, are more likely to seek biomedical care in conjunction with traditional midwifery care rather than to rely solely on the former. CONCLUSION: The findings suggest that midwives are likely to continue to be key providers of pregnancy related care in the future, even as access to modern health facilities improves. Current efforts directed toward the training and integration of midwives into the formal health system are likely to be much more effective at improving pregnancy related care than the replacement of midwives with biomedical providers. PMID- 10858937 TI - The new 2001 Census question set on cultural characteristics: is it useful for the monitoring of the health status of people from ethnic groups in Britain? AB - The health of minority ethnic groups has been accorded a priority position in the British government's strategy to improve the population's health, including the provision of the necessary information to address inequalities. The independent Acheson Inquiry has also called for improvements in the capacity to monitor inequalities in health of ethnic groups which requires the use of appropriate ethnic group categories. The new 2001 Census question set includes a substantially revised ethnic group question and a new question on religion which address some of the shortcomings in the 1991 Census question. However, the token breakdown of the white group and its unsatisfactory capture of those of Irish origin, the use of Indian subcontinent groups that ignore ethno-religious differences and have little saliency amongst those being described, and the omission in Scotland of a subdivided white group and the religion question, are important deficiencies. The use of a few pan-ethnic racial groups in the proposed census tables is a major drawback. It is important that these changes are widely debated. PMID- 10858936 TI - Dental health and access to dental care for ethnic minorities in Sweden. AB - OBJECTIVE: To describe access to dental care in a population-based sample of foreign-born Swedish residents in relation to dental health. DESIGN: The study was based on data from the Immigrant Survey of Living Conditions in four minority study groups consisting of a total of 1,898 Swedish residents born in Poland, Chile, Turkey and Iran aged 27-60. An age-matched study group of 2,477 Swedish born residents from the Survey of Living Conditions of 1996 was added as a comparison group. The study also included 2,228 children aged 3-15 years in the minority households and 2,892 children in the households of the Swedish-born study group. RESULTS: The risk of poor dental health was higher in all four minority study groups than for the Swedish-born study group after adjusting for socio-economic variables. In the adult minority study groups the adjusted odds ratios (ORs) for having prostheses and problems with chewing was 6.3 (4.3-9.1) and 2.7 (1.8-4.3), respectively, for the Polish-born, 4.8 (3.3-7.1) and 3.2 (2.1 4.9) for the Chilean-born, 4.6 (3.1-6.9) and 4.8 (3.6-7.2) for the Turkish-born, and 2.7 (1.5-4.8) and 6.5 (4.1-10.3) for the Iranian-born compared with the Swedish-born. In the child study group all four minority groups had an increased risk of caries ranging from OR 1.6 (1.3-2.1) in the Chilean group to 2.5 (2.0 3.0) in the Turkish group compared with the children with Swedish-born parents. The adults in all four minority study groups more often lacked regular treatment by a dentist than Swedish-born residents. The OR for not having been treated by a dentist during the 2 years preceding the interview ranged from 1.9 (1.4-2.6) in the Polish-born study group to 3.0 (2.3-4.0) in the Chilean-born study group after adjustment for socio-economic factors and general health. CONCLUSION: This study demonstrates that adults in minority populations in Sweden use less dental care despite having greater needs of dental treatment than the majority population. This inequity calls for action in health policy and preventive dental health programmes. PMID- 10858938 TI - Limiting long-term illness among black Caribbeans, black Africans, Indians, Pakistanis, Bangladeshis and Chinese born in the UK. AB - AIM: To examine limiting long-term illness among ethnic groups born in the UK. DESIGN: Study members in the ONS Longitudinal Study, a representative 1% sample of England and Wales, present at the 1991 Census were examined. Socio-economic position was measured using housing tenure and access to cars. MAIN RESULTS: Black Caribbeans, Black Africans, Indians, Pakistanis and Bangladeshis reported more limiting long-term illness than whites, and the Chinese less. The level was higher among Black Africans than Black Caribbeans, even after adjustment for socio-economic position. Corresponding patterns were seen across the generations for all groups except Black Africans. Black Africans born in the UK reported more limiting long-term illness than did those born in Africa. Adjusting for socio economic position lowered the risks in every group except Indians. The effect was strongest for Black Caribbeans, among whom adjustment removed the higher risks, and Bangladeshis, among whom the higher risks were no longer significant. CONCLUSION: For South Asians and Black Caribbeans poor health persisted across generations, and for Black Africans health worsened. There are over 1.5 million second- and third-generation migrants in these ethnic groups, but little is known about the consequences of having a multigenerational identity. PMID- 10858939 TI - The impact of diabetes mellitus among the Metis of western Canada. AB - OBJECTIVES: To examine the impact of diabetes mellitus on the lives of the Metis of western Canada, and to determine the extent of co-morbidity among Metis with diabetes. DESIGN: The source of data was the Aboriginal Peoples Survey (APS), conducted by Statistics Canada in 1991. The survey was administered to a representative sample of Aboriginal peoples throughout Canada. Analysis was completed on self-identified Metis participants from the Canadian provinces of Manitoba, Saskatchewan and Alberta. RESULTS: Metis participants with diabetes were more likely than those without diabetes to report their health status as poor. Significantly greater numbers of Metis with diabetes reported activity limitations at work, at home and in leisure activities, the need for assistance with activities of daily living and difficulties with ambulation than did those without diabetes. The extent of co-morbidity was also significant. Metis with diabetes were almost three times more likely to report hypertension and heart problems and twice as likely to report sight impairments than were those without diabetes. CONCLUSIONS: This research represents the first account of the effects of diabetes on the lives of the Metis. The APS data have provided a clear picture of the impairments in physical functioning experienced by the Metis with diabetes and the impact upon quality of life. In addition, the strong associations between diabetes and hypertension, heart problems and sight impairments suggest profound morbidity in this population that warrants prompt attention. PMID- 10858940 TI - Investigation of validity of closed questions in a survey of British South Asian and white populations. AB - OBJECTIVES: The paper concerns the validity of closed-format questions in an interview-based population survey and focuses on a comparison of South Asian and white respondents. METHOD: A two-part interview consisting of open, respondent centred questions followed by closed questions taken from a large-scale interview based health survey was carried out with 15 persons from white and 14 from South Asian communities resident in the UK. Interviewees' views of their stays in hospital was the focus. RESULTS: The two-part interview was found to provide a broadly satisfactory method for the investigation of validity although a limitation was recognised. Twenty-nine per cent of the variation in response to a closed question on 'overall satisfaction' was predictable from views expressed in the open interview. The views of both groups, as expressed in the open interview, were inadequately represented by the closed questions. This appeared to be especially true for the South Asian sample. The validity of the closed question appeared greater for white than for South Asian interviewees. CONCLUSIONS: It is concluded that when designing and piloting health surveys consideration should be given to members of all communities to be included in the survey. PMID- 10858941 TI - Cross-cultural ranking of IADL skills. AB - Although causal theories of aging differ considerably across disciplines, the common denominator in published papers on the health and function of the aged is to be found in the methods section. The instrumental activities of daily living (IADL), used to determine the independence of the aged living in the community, are utilized both at a clinical and population level. While there has been a cross-disciplinary need for a scale to measure sub-populations, many scholars have been reticent to use the existing IADL items across geographical backgrounds. The results of this study, which surveys 236 physicians from 11 different cultures on the importance of individual IADL items, suggests the IADL may be useful as a universal measure of function. PMID- 10858942 TI - A study of body weight concerns and weight control practices of 4th and 7th grade adolescents. AB - OBJECTIVE: The purpose of this study was to assess grade, race, socioeconomic status and gender differences in perceptions of body size, weight concerns, and weight control practices between 4th and 7th grade students in South Carolina. DESIGN: Two random samples, consisting of a total of 1,597 children (53.1% white, 51.97% female, 44.9% 4th graders) participated in two questionnaire surveys. Both surveys included a series of seven female and seven male body size drawings, body image and weight concern questions, and questions pertaining to weight control practices. Responses to the questionnaire were analyzed using chi-square analysis and the General Linear Model. RESULTS: Using socioeconomic status (SES), race, gender, and grade as independent variables, differences in ideal adult body size, opposite gender ideal adult body size, weight concerns, perceptions of family/peer weight concerns, perceptions of own body size and weight control practices were studied. Analyses revealed that 4th grade males select a larger ideal adult body size and opposite gender ideal adult body size when compared to 7th grade males (p < 0.0001 and p = 0.0078, respectively). Fourth grade females indicated less personal concern about their weight than 7th grade females (p = 0.0009). Fourth graders also perceived less family/peer concern about weight than 7th graders (p = 0.0027) and 7th graders described themselves as being more overweight than 4th graders (p = 0.0039). Blacks selected a significantly larger body size than white children for ideal adult (p = 0.0287) and ideal opposite gender adult body size (p < 0.0001 for males, p = 0.0030 for females). Blacks also perceived less personal and family/peer concern about weight when compared to whites (p = 0.0083). More whites reported that they were trying to lose weight, as compared to blacks (p = 0.0010). Males also selected significantly larger body size silhouettes than females for ideal adult body size (p = 0.0012). Males expressed less personal concern about weight (p < 0.0001), perceived less family/peer concern about weight (p < 0.0001), and were less likely than females to be engaged in weight loss (p < 0.0001). Females in the high SES category selected a significantly smaller ideal male adult body size than females in the low SES group (p = 0.0124) and more females in the high SES category were trying to lose weight when compared to females in the low SES group (p = 0.0055). CONCLUSION: This study indicates that early in a child's sociocultural development, grade level, gender, race, and SES are influential in the perception of ideal adult body size and opposite gender ideal adult body size. These factors are also influential in determining concerns about weight and weight control practices. The findings of this study support the need to begin health and wellness education efforts early in childhood while taking into account racial, gender, age, and SES disparities. This knowledge can also be useful in targeting interventions for both obesity and eating disorders. PMID- 10858943 TI - [What has shown itself to be reliable in treatment of autistic disorders?]. PMID- 10858944 TI - [Methodological problems of longitudinal studies on schizophrenia]. AB - Longitudinal studies are a key to understanding schizophrenia. They are the more informative, the longer the periods covered. Hence, good studies into the course of schizophrenia almost exclusively involve a lot of effort and cost. In practice, however, time-consuming methods and design variables must be avoided. The pitfalls this constraint produces are instructive of the difficulties longitudinal studies are faced with in striving for valid results. For reasons of research economy, requirements must be adjusted to study objectives. Studies into the short term course are less time-consuming, but because of the rapid changes in the illness course study intervals should be defined clearly and observed strictly. In long-term studies, too, one source of error lies in the highly varying lengths of illness of the patients studied. Even some of the classic long term studies are marred by this error. The beginning of the follow-up period should be comparable across the study cohort and as close to illness onset as possible. To obtain generally valid results the probands must be representative of all the illness cases in the general population not only at the outset, but also all the later stages of the study. Besides the efforts to avoid attrition in the study cohort, ways must be found for correcting and estimating data for an acceptable proportion of drop-outs. In the analysis of course and outcome the indicators chosen must be apt to the traditional subtypes as well as to a theoretical symptom patterns and empirical symptom structures. In the context of typical design variables of longitudinal studies the assets and weaknesses of two retrospective and one prospective design will be discussed. Concerning the social course, importance of disease-independent factors, such as age, sex and level of social development at illness onset, as well as of control groups will be demonstrated. Predictor models will be discussed with reference to the direct and indirect influences involved. Examples of such analyses will be given. PMID- 10858945 TI - [Collaboration of the general practitioner and the psychiatrist with the psychiatric hospital. A literature review]. AB - Co-operation of physicians in private practice with psychiatric hospitals was investigated in Germany scarcely until now, although evaluation of consumer satisfaction is of great importance to quality assurance in psychiatry. In this paper, findings from previous studies are presented together with data from interviews with general practitioners and psychiatrists, evaluating their expectations regarding psychiatric hospitals. Substantial problem area in collaboration is referral to the psychiatric hospital. Apart from sociodemographic and disease-related variables, referral practice depends on referring physician's attitudes and competence in psychiatry, and provider influences like delay of admission, communication with referring physician, and competence of the hospital. As conclusion, constructive collaboration must be developed at the interface of outpatient and inpatient care. On account of increasing diversification of psychiatric services, functional network should be an ongoing goal to improve treatment continuity of patients with mental disorders. PMID- 10858946 TI - [Improvement of agitation and anxiety in dementia patients after psychoeducative training of their caregivers]. AB - It has been convincingly demonstrated that in dementia psycho-educative training of caregivers positively impacts on motivation for care and satisfaction of the caregivers. It has, however, been neglected to examine the effect of psycho educative training on the behavioural and psychological symptoms of dementia sufferers. In a three-month, expert-based and conceptualized group intervention with caregiving relatives of demented patients we investigated, whether functional impairment and behavioural and psychological symptoms may improve, which of a set of independent variables may predict improvement, and how the group intervention will be appreciated by the caregivers. The group intervention yielded a significant improvement of memory-related functions in daily living and a significant decrease of agitation and anxiety of the demented patients. The presence of an additional somatic disease predicted worse outcome of the intervention with respect to the impairment of memory-related functions in daily living and of agitation. Anonymous inquiry of the caregivers with respect to their judgement of the intervention revealed high acceptance and appreciation. This study demonstrated that a psycho-educative group intervention with caregiving relatives of dementia sufferers is helpful for both the caregivers and the demented patient. This evidence of a positive mediate effect of the group intervention on the functional and behavioural impairment of the demented patients underscores the importance of nonpharmacological strategies in the treatment plan of dementia. PMID- 10858948 TI - [6th gerontopsychiatric specialist discussion, Dusseldorf, November 26-27, 1999]. PMID- 10858947 TI - [Treatment of cocaine dependence. Intoxication, withdrawal and prevention of relapse]. AB - The aim of this review article is to evaluate the treatment of cocaine withdrawal, cocaine-intoxication and long-term relapse prevention of cocaine addicts. Some 25% of police recognized first time drug users in Germany consume cocaine. However, there is an increasing number of cocaine-abusers and -addicts in the USA. The withdrawal of cocaine can be divided into three phases dominated mainly by psychiatric symptoms. Life-threatening condition can occur in cocaine intoxication mainly in combination with other drug-use. A high risk of relapse is seen in follow-up trials of cocaine-addicts. Intensive craving, high cocaine- and substance-abuse is reported regularly in cocaine-addicts after detoxification therapy. Recommendations in the treatment of cocaine-intoxication, withdrawal and long-term relapse prevention are made. The use of antidepressives, anticonvulsants, dopaminergic and serotonergic medications as well as behavioural, psychoanalytical and combined therapies and their efficacy in clinical and trails is evaluated. A short review of new experimental therapies in the treatment of cocaine-dependence is shown. PMID- 10858949 TI - [Comment on "Dementia of the Alzheimer type in women." J. C. Nedoschill. Forschr Neurol Psychiat 67: 441-117, 1999]. PMID- 10858950 TI - Cell cycle control and cancer. PMID- 10858951 TI - Implication of the G2 checkpoint in the maintenance of genome integrity. AB - Checkpoints are surveillance mechanisms that block cell cycle transitions, for instance in response to DNA damage. We summarise here recent progress in the molecular characterisation of the G2 checkpoint which controls the entry into mitosis, and review new evidence which implicates deregulated expression of checkpoint proteins and proteins involved in DNA damage repair in cancer development. There now exists good evidence that individuals who have inherited mutations in genes involved in G2 checkpoint and DNA damage repair are predisposed to the development of various types of cancer, their cells having a strong tendency to accumulate additional mutations. However, the occurrence of mutations of most of these genes in sporadic tumors has yet to be analysed more accurately. PMID- 10858952 TI - [Role of CDC25 phosphatases in the control of proliferation]. AB - Progression in cell cycle is controlled by CDKs (cyclin dependent kinases) and their cyclins regulatory subunits. In mammalian cells, three dual specificity phosphatases called CDC25 activate CDKs/cyclin complexes. The activity of CDC25 is regulated by phosphorylation and dephosphorylation events. CDC25 phosphatases also participate in cell cycle checkpoints activated in response to DNA damage. Two members of this family, CDC25 A and CDC25 B, have oncogenic properties, and their overexpression has been detected in various types of tumors. PMID- 10858953 TI - The functions of the cdk-cyclin kinase inhibitor p21WAF1. AB - p21WAF1 plays a critical role in regulating cell growth and the cell response to DNA damage. The primary targets of p21WAF1 (hereafter referred to as p21) are the cdk-cyclins which regulate the progression of eukaryotic cells through the cell cycle, and proliferating cell nuclear antigen (PCNA), an accessory protein of DNA polymerase delta. p21 forms complexes with a class of cdk-cyclins to inhibit their kinase activity and with PCNA to inhibit DNA synthesis. These distinct properties map to the N-terminal and the C-terminal regions of p21, respectively. Cell cycle arrest in G-1 (G-1 checkpoint) following DNA damage is mediated by p53 and is deficient in p21 null cells. p53 thus upregulates p21 expression in response to DNA damage, which in turn inhibits cdk2-associated kinase activity. Retinoblastoma protein is regulated by cdk-cyclin kinases, and acts as a downstream target of p21 in DNA damage-induced G-1 arrest. Furthermore, accumulating evidence indicates that p21 may play a role in maintaining G-2 arrest after DNA damage. Transcriptional control of p21 by factors other than p53 is critical for growth arrest and for cell differentiation in many instances. PMID- 10858954 TI - p27: a pleiotropic regulator of cellular phenotype and a target for cell cycle dysregulation in cancer. AB - The eukaryotic cell cycle is regulated by the sequential activation of cyclin dependent kinases (CDKs). CDK activation is regulated by phosphorylation of the catalytic subunit, by binding to activating (cyclins) and inactivating subunits (cyclin-dependent kinase inhibitor). In this review, we will focus on the role of the cyclin-dependent kinase inhibitor p27 which has been recently the subject of extensive work. This negative regulator of cell growth indeed illustrates the pleiotropic biological effects of such molecules in both normal and cancer cells and the complexity of the regulatory mechanisms involved. PMID- 10858955 TI - Erythroid cell development and leukemic transformation: interplay between signal transduction, cell cycle control and oncogenes. AB - Studies using genetically modified mice and ex vivo tissue culture of erythroid progenitors converge to show that generation of mature erythroid cells depends on the interplay between specific transcriptional regulators and intracellular signals controlled by cytokines and growth factors. These studies also show that terminal differentiation in the erythroid lineage is unusual since the acquisition of the phenotypic traits of mature cells occurs while the cells are still actively dividing. Furthermore, under specific stress conditions, a massive and sustained self-renewal of committed erythroid progenitors can take place to replenish the pool of terminally differentiated cells. We review here how the erythroid genetic program and its interplay with specific cytokines, growth factors and hormones controls survival, proliferation and differentiation of erythroid progenitors both in normal and stress conditions. Special emphasis is laid on our present understanding of the differences in cell cycle control, which result either in self-renewal of erythroid progenitors or in the particular cell divisions which accompany terminal differentiation. Finally, we discuss how deregulation of the various aspects of the physiological control of erythroid progenitor survival, proliferation and differentiation can lead to erythroblast transformation and erythroleukemia. PMID- 10858956 TI - Signaling to p53: breaking the posttranslational modification code. AB - In unstressed cells, the tumor suppressor protein p53, a tetrameric transcription factor, is present in a latent state and is maintained at low levels through targeted degradation. A variety of cellular stresses including DNA damage, hypoxia, nucleotide depletion, viral infection, and cytokine-activated signaling pathways that transiently stabilize the p53 protein, cause it to accumulate in the nucleus, and activate it as a transcription factor. Activation leads either to growth arrest at the G1/S or G2/M transitions of the cell cycle or to apoptosis. The molecular mechanisms by which stabilization and activation occur are incompletely understood, but accumulating evidence points to roles for multiple posttranslational modifications in mediating these events through several potentially interacting but distinct pathways. Both the approximately 100 amino acid N-terminal and approximately 90 amino acid C-terminal domains are highly modified by phosphorylation and acetylation, whereas modifications to the central sequence-specific DNA binding domain have not been reported. Seven serines and one threonine in the first 46 residues of the transactivation domain and four to five serines in the carboxyl-terminal domain are now known to be phosphorylated, and Lys320 and Lys382 in the carboxyl-terminal domain (human p53) can be acetylated. Antibodies that recognize p53 only when it has been modified at specific sites have been developed by several laboratories, and studies with these have shown that most of the known posttranslational modifications are induced when cells are exposed to DNA-damaging agents. Exceptions are Ser378, which is reported to be constitutively phosphorylated, and Ser376, which is dephosphorylated in response to DNA damage. These recent results, coupled with biochemical and genetic studies, suggest that several amino-terminal phosphorylations can be important in stabilizing p53 in response to DNA damage and in directing acetylation at C-terminal sites. DNA damage-induced modifications to the C-terminus inhibit the ability of this domain to negatively regulate sequence-specific DNA binding either by inducing a conformational change in the protein or by inhibiting non-sequence-specific DNA binding by the C terminus. C-terminal modifications also modulate the oligomerization state of p53, and may modulate nuclear import/export. Modifications in response to DNA damage to other components that interact with p53 may also be important. In most cases, clear roles for specific modifications, interactions among individual modifications, and the enzymes responsible for each modification remain to be defined. Nevertheless, the field appears poised for major advances in the understanding of the molecular mechanisms that regulate p53 function. PMID- 10858957 TI - The role of p53 in the response to mitotic spindle damage. AB - The p53 tumour suppressor protein has defined roles in G1/S and G2/M cell cycle checkpoints in response to a range of cellular stresses including DNA damage, dominant oncogene expression, hypoxia, metabolic changes and viral infection. In addition to these responses, p53 can also be activated when damage occurs to the mitotic spindle. Initially, spindle damage activates a p53-independent checkpoint which functions at the metaphase-anaphase transition and prevents cells from progressing through mitosis until the completion of spindle formation. Cells eventually escape from this block (a process termed 'mitotic slippage'), and an aberrant mitosis ensues in which sister chromatids fail to segregate properly. After a delay period, p53 responds to this mitotic failure by instituting a G1 like growth arrest, with an intact nucleus containing 4N DNA, but without the cells undergoing division. Cells lacking wild-type p53 are still able to arrest transiently at mitosis, and also fail to undergo division, underscoring that the delay in mitosis is p53-independent. However, these cells are not prevented from re-entering the cell cycle and can reduplicate their DNA unchecked, leading to polyploidy. Additionally, p53-null cells which experience spindle failure often show the appearance of micronuclei arising from poorly segregated chromosomes which have decondensed and been enclosed in a nuclear envelope. The ability of p53 to prevent their formation suggests an additional G2 involvement which prevents nuclear breakdown prior to mitosis. The molecular mechanism by which p53 is able to sense mitotic failure is still unknown, but may be linked to the ability of p53 to regulate duplication of the centrosome, the organelle which nucleates spindle formation. PMID- 10858958 TI - p53 and cell-cycle control: a finger in every pie. AB - The p53 protein plays diverse, complementary roles in the regulation of cell proliferation, genetic integrity and survival after exposure to various forms of genotoxic and non-genotoxic stresses. While there is considerable in vivo evidence that induction of apoptosis in response to DNA damage is an important biological response mediated by p53, the physiological role of p53 in the control of the cell cycle is still poorly understood. In this review, we first discuss evidence showing that p53 expression, protein level and activity are regulated in a cell-cycle dependent manner (by transcriptional control, control of protein stability, and changes in phosphorylation, acetylation and protein conformation). We then summarize the data available on the direct or indirect involvement of p53 in the control of cell-cycle progression in G1, S, G2 and M phases. Based on recent in vitro and in vivo data, we suggest that permanent cell-cycle arrest may represent a response to stress in cells with limited proliferative potential, which allows both genetic and tissular integrity to be preserved. We also discuss the way in which cell-cycle regulation may contribute to control p53 protein levels and activity during the normal cell cycle. PMID- 10858959 TI - Apoptosis and cell cycle: distinct checkpoints with overlapping upstream control. AB - The question as to whether apoptosis (programmed cell death) is controlled by one or few checkpoints is still unresolved. A growing body of evidence suggests that (one of) the decisive event(s) of cell death consists in the permeabilization of mitochondrial membranes. Indeed, multiple pro-apopotic signal transduction pathways converge on the proteins of the Bcl-2/Bax family which, in concert with the so-called permeability transition pore complex (PTPC), regulate mitochondrial membrane barrier function. Mitochondrial permeabilization causes the release of soluble intermembrane proteins, some of which are involved in the activation of apoptotic proteases and nucleases. Thus, the putative checkpoint determining the death/life decision is clearly different from the known checkpoints of cell cycle progression. Prominent oncogenes (e.g., c-Myc, Ras, Raf, Bcl-2) and tumor suppressor genes (e.g., p53, Bax) have been shown to modulate apoptosis via a direct or indirect effect on mitochondrial membranes. All these oncoproteins and tumor suppressor proteins may simultaneously influence the cell cycle and the propensity to undergo apoptosis. Several cell cycle regulatory proteins (e.g., cyclins, cdk, etc.) can induce or inhibit apoptosis via yet unknown pathways. PMID- 10858960 TI - sHsp as novel regulators of programmed cell death and tumorigenicity. AB - sHsp (small stress proteins) are molecular chaperones involved in cellular defence mechanisms against several different types of aggressions. These proteins also participate in essential physiological processes, such as regulation of cell cycle, differentiation, programmed cell death and tumorigenicity. For example, sHsp are transiently expressed during the cell division to differentiation transition and this phenomenon prevents differentiating cells from undergoing apoptosis. sHsp also protect against apoptosis induced by different conditions or agents, particularly anti-cancer drugs. Of interest, tumor cells usually express high levels of sHsp, and anti-cancer drugs, such as cisplatin, trigger the accumulation of sHsp. These proteins are also known to interfere with programmed cell death induced by TNF alpha and Fas ligand. Moreover, they enhance the growth of tumors in vivo. Taken together, these observations suggest that sHsp can allow cancerous cells to escape the immunosurveillance mediated by death ligands and can render these cells resistant to therapy. Hence, sHsp represent prime targets for therapeutic interventions. This review is focused on the role of sHsp in different aspects of the life and death of mammalian cells and on the role of these survival proteins in cancer. PMID- 10858961 TI - Lytic infection by double-strand DNA viruses and cell cycle alterations. AB - Polyomaviruses, papillomaviruses, adenoviruses and herpesviruses are double stranded DNA viruses that replicate in the nucleus of the cells they infect and have evolved various strategies to create a cellular environment that is optimally conducive to their replication. One of these strategies consists of activating cellular genes, mostly S-phase genes that are required for the replication of the viral genome. Concomitantly, they encode one or several proteins that negatively regulate the response of the cell to viral infection, notably cell cycle arrest and/or apoptosis. As a result, these viruses profoundly alter the biochemical pathways that normally control cellular growth, and may thus promote uncontrolled cell proliferation. This review describes some well known mechanisms of cell cycle alteration induced by these viruses. PMID- 10858962 TI - Replicative senescence: mechanisms and implications for human cancer. AB - The proliferative lifespan of most normal human cells, even in ideal growth conditions, is limited by intrinsic inhibitory signals which induce cell cycle arrest after a preset number of cell divisions. This process of 'replicative senescence' is activated in many cell types by the progressive erosion of the specialised ends of chromosomes--telomeres--which act as a molecular 'clock'. Although many details are still to be elucidated, one major signal pathway linking telomere shortening to growth arrest operates via activation of the tumour suppressor gene (TSG) product, p53, which in turn induces the cell-cycle inhibitor p21WAF1, and at least in some cell types wild-type p53 function is an absolute requirement for normal senescence. Given the evidence that replicative senescence represents a natural obstacle to tumour progression, the need to escape p53-mediated senescence may therefore represent a major selection pressure for loss of p53 function in many human cancers. PMID- 10858963 TI - [The ARF-p53 pathway: a line of defence against oncogenic signals]. AB - The MTS1 (Multiple Tumor Suppressor 1) locus is a very original one as its organization results in the expression of two alternative transcripts that encode two structurally and functionally different proteins: INK4a and ARF (also designated p19ARF in mouse and p14ARF in man). Recent findings indicate that the latter is a major component of a regulatory pathway of oncogenic signals culminating in p53 activation by stabilisation of the protein. While the importance of this pathway has been overtly established in animal experimental oncology, it still has to be further documented in human oncology in order for this gene to acquire its full biological significance. PMID- 10858964 TI - Small GTPases, adhesion, cell cycle control and proliferation. AB - In many cells, proliferation is under the coordinated control of growth factors and the extracellular matrix (ECM). Autocriny and anchorage-independent growth are observed in many transformed cells where this balance is often altered. The outside-in and inside-out exchanges are mediated by integrins, acting as molecular bridges between the ECM and the cytoskeleton. Integrins, as the major receptors for components of the ECM, are important not only for the physical aspects of cell adhesion, but also for two key aspects of cell fate: cell cycle progression, and apoptosis. The latter aspect will not be covered in the present analysis, which will rather focus on the data accumulated on Ras and Ras-related small GTPases such as Rho, Rac and Cdc42. PMID- 10858965 TI - [Integrins and cell cycle control by the environment]. AB - Integrins insure cell adhesion to extra-cellular matrix components; they are thus involved in tissue architecture. They also can insure intercellular adhesions by binding to surface molecules from the immunoglobulin superfamily. Integrins binding to their ligands induce cytoskeleton reorganisation and, consequently, they gather into focal adhesion contacts. This greatly strenghthens mechanical forces. Nevertheless, integrins can also participate in cell locomotion and, moreover, tranduce within cells signals that can extensively influence cell metabolism, cell cycle and apoptosis. Doing so, they can interact with signals from other cellular receptors, such as soluble growth factors. They are therefore key molecules to integrate intrinsic and extrinsic events of the cellular behavior. They profoundly influence oncogenesis and the metastatic process. PMID- 10858966 TI - [Chemical inhibitors of cyclic-dependent kinases: preclinical and clinical study]. AB - In the past decade, the use of a large variety of cellular models and cell biology, biochemistry and molecular biology techniques has led to the discovery of key proteins that are intimately involved in the regulation of tumor growth. In particular, it has been shown that cyclin-dependent kinases (CDKs) are key regulators of the cell-division cycle. Their frequent deregulation in human tumors make them attractive targets for the identification of new antineoplasic agents. Intensive screening has led in the past few years to the identification of a series of selective and potent chemical inhibitors of CDKs. Drugs representing new lead structures like flavopiridol, indirubin and staurosporine++ derivatives have already been used in clinical evaluation for cancer treatment (clinical trials, phase I and II). Anticancer drug development is being pursued to reduce their toxic side effects, to improve their pharmacokinetic properties and to increase their anti-tumor activity. In this context, traditional drug screening methods in biological test systems have led to the discovery of new compounds such as purine derivatives and paullones, which display remarkable selectivity and efficiency. These novels drugs may result in substantial progress in cancer treatment in the near future. PMID- 10858967 TI - Morning sickness: a mechanism for protecting mother and embryo. AB - Approximately two-thirds of women experience nausea or vomiting during the first trimester of pregnancy. These symptoms are commonly known as morning sickness. Hook (1976) and Profet (1988) hypothesized that morning sickness protects the embryo by causing pregnant women to physically expel and subsequently avoid foods that contain teratogenic and abortifacient chemicals, especially toxic chemicals in strong-tasting vegetables, caffeinated beverages and alcohol. We examined this hypothesis by comprehensively reviewing the relevant medical, psychological and anthropological literature. In its support, (i) symptoms peak when embryonic organogenesis is most susceptible to chemical disruption (weeks 6-18), (ii) women who experience morning sickness are significantly less likely to miscarry than women who do not (9 of 9 studies), (iii) women who vomit suffer fewer miscarriages than those who experience nausea alone, and (iv) many pregnant women have aversions to alcoholic and nonalcoholic (mostly caffeinated) beverages and strong-tasting vegetables, especially during the first trimester. Surprisingly, however, the greatest aversions are to meats, fish, poultry, and eggs. A cross cultural analysis using the Human Relations Area Files revealed 20 traditional societies in which morning sickness has been observed and seven in which it has never been observed. The latter were significantly less likely to have animal products as dietary staples and significantly more likely to have only plants (primarily corn) as staples than the 20 societies in which morning sickness occurred. Animal products may be dangerous to pregnant women and their embryos because they often contain parasites and pathogens, especially when stored at room temperatures in warm climates. Avoiding foodborne microorganisms is particularly important to pregnant women because they are immunosuppressed, presumably to reduce the chances of rejecting tissues of their own offspring (Haig 1993). As a result, pregnant women are more vulnerable to serious, often deadly infections. We hypothesize that morning sickness causes women to avoid foods that might be dangerous to themselves or their embryos, especially foods that, prior to widespread refrigeration, were likely to be heavily laden with microorganisms and their toxins. The alternative hypotheses that morning sickness is (i) an epiphenomenon of mother-offspring genetic conflict or hormones associated with viable pregnancies, or (ii) an indicator to potential sexual partners and kin that the woman is pregnant, resulting in reduced sexual behavior and increased nepotistic aid, were not well supported. Available data are most consistent with the hypothesis that morning sickness serves an adaptive, prophylactic function. PMID- 10858968 TI - Scientific animal breeding in Moravia before and after the rediscovery of Mendel's theory. AB - Leading Moravian sheep breeders, who joined with university professors and other educated citizens to form a Sheep Breeders' Society in 1814, looked to science to provide a reliable basis for breeding. Their activities reached a climax in the 1830s, when they defined and focused on heredity as the central research goal. Among the members taking part was Abbot Cyrill F Napp, who in 1843 would accept Mendel into the monastery. The contributions of Abbot Napp to the sheep breeders' view of heredity are here described. After 1900, when Moravian animal breeding sought to embrace Mendelism, in competition with other theories, a major influence was exerted by Jaroslav Krizenecky (1896-1964). In 1963, Krizenecky accepted responsibility for establishing the Mendel Museum (Mendelianum) in Brno as a vehicle for historical research into the origin and essence of Mendel's discovery. PMID- 10858969 TI - Mechanisms of protein degradation: what do the rat studies tell us. AB - Uremia induces substantial changes in protein metabolism. The branched-chain amino acids serve as useful markers of these changes and their catabolism is increased in uremia, particularly in the presence of metabolic acidosis. Glucocorticoids also are involved in accelerating protein degradation, and the negative nitrogen balance which results in loss of lean body mass. Cellular mechanisms accounting for these changes include an up-regulation of the ubiquitin proteasome pathway and branched-chain ketoacid dehydrogenase activity in muscle. A low insulin level also appears to play a permissive role in causing increased catabolism. These findings have important clinical implications because correction of the metabolic acidosis with alkali blunts these responses and improves nutritional status. PMID- 10858970 TI - The vascular endothelium in chronic renal failure. AB - Chronic renal failure (CRF) is associated with a 20-fold increased risk of cardiovascular death, two principal mechanisms being: sudden, arrhythmic death associated with left ventricular hypertrophy, and ischaemic heart disease, associated with accelerated atherosclerosis. In recent years, the vascular endothelium has been recognised as a large and complex endocrine organ, with many important physiological functions including the control of vascular tone. Endothelial dysfunction, commonly characterised by reduced production of the vasodilator nitric oxide (NO), is thought to be a key initial event in the development of atherosclerosis and is present in patients with hypertension and hyperlipidaemia. While these cardiovascular risk factors are also prevalent in CRF, other factors more specific to uraemia such as accumulation of homocysteine and asymmetric dimethylarginine (endogenous inhibitor of NO synthase) may impair endothelial function. Modulation of endothelial function in CRF may offer a novel strategy to reduce cardiovascular disease. PMID- 10858971 TI - 3 R study: renal outcome in renal ischemia: revascularisation or medical treatment. AB - Ischemic nephropathy refers to the kidney damage following stenosis or an obstructive lesion in the main kidney arteries. This disorder has been overlooked in the past and a more rational and specific use of clinical criteria, and the development of not very invasive techniques with a good diagnostic accuracy such as spiral CT angiography, NMR angiography and echo-colour-Doppler have improved our ability to identify these patients. It is therefore likely that, in the next few years, we will find ourselves treating an increasing number of patients with renovascular ischemic disorders. Transluminal angioplasty and, more recently, the use of endovascular stents, have led to a marked improvement in the treatment of stenoses and, together with vascular surgery, allow to treat almost all patients with this disorder. There is, however, a lack of prospective and controlled studies, which demonstrate the long term benefit of revascularization treatment, as compared with optimum conservative treatment in reducing cardiovascular mortality, cardiovascular events and preserving renal function. The Ischemic Nephropathy Study Group of the Italian Society of Nephrology has organized a prospective, controlled study over a period of three years, aimed at comparing the effect of revascularization versus medical therapy in 300 patients with renal artery stenosis, ranging between 50 and 90 per cent, who will be randomly assigned to the two treatments. End point will be cardiovascular mortality and morbidity and need for renal replacement therapy. PMID- 10858972 TI - Variability of renal echo-Doppler measurements in healthy adults. AB - BACKGROUND: We examined the most widely used echo-Doppler variables in healthy adults to define their normal distribution and variability in relation to age, sex, body surface area and the right and left kidney. METHODS: Ninety healthy subjects were selected, stratified for sex and age (range 20-65 years). We also examined 8 subjects with a congenital solitary kidney and 15 surgically nephrectomized patients. Variables studied were the diameters and volume of the kidneys, renal blood flow (RBF) and resistive index (RI) measured on the renal, interlobar and cortical arteries. RESULTS: The mean length was greater in the left kidney (p<0.01) and width in the right one (p<0.02). Volume was no different on the two sides. RBF showed an age-dependent reduction (p<0.0001), while interlobar (p<0.0001) and cortical (p<0.0001) RI showed a selective age-dependent increase. RI were higher in females and diameters, volume and RBF, after correction for body surface area, were not different in the two sexes. Cortical RI was lower than the interlobar and renal RI (p<0.0001). In the group with congenital single kidney, length, volume and RBF were all greater. In the mononephrectomized patients, length and volume were greater but RBF was the same as in a normal single kidney. CONCLUSIONS: The ultrasonographic and Doppler variables studied did not show any differences from normal distribution but they were influenced differently by some demographic parameters; however, the integrated use of these measurements offers precision and repeatibility, and could help evaluating diffuse or localized abnormalities. PMID- 10858973 TI - Routine immunofluorescence and light microscopy processing with a single renal biopsy specimen: 18 years' experience in a single centre. AB - To obtain a correct histological renal diagnosis, two adequate cortical biopsy cylinders are routinely taken for light microscopy and immunohistology in nephrology units. We describe our simple method for light microscopy (LM) study using the same frozen renal material used for immunofluorescence (IF). Of over 2,000 biopsies processed with this method, only three showed thawing artifacts and all occurred when the room temperature was above 30 degrees C. Our 18 years' experience and the large number of biopsies indicate that this method offers an interesting new way to do LM and IF with one biopsy core and could help stimulate etiopathogenic and diagnostic studies in nephrology. The other renal specimen can be used for histological investigation, applying recent molecular biology techniques (PCR, RT-PCR, in situ hybridization). PMID- 10858974 TI - Transtubular potassium concentration gradient (TTKG) and urine ammonium in differential diagnosis of hypokalemia. AB - BACKGROUND: Hypokalemia is a common and sometimes serious clinical problem, whose etiological diagnosis can frequently be based on the patient's history and the clinical setting. Measurement of urinary indices such as excretory rate of K+, random urine K+ concentrations and blood acid-base parameters have been employed in the pathophysiological diagnosis, though with some pitfalls. METHODS: To investigate the diagnostic usefulness of the transtubular potassium concentration gradient (TTKG) and urine ammonium in the differentiation of hypokalemia, we measured serum K+ and osmolality, random urine electrolytes, osmolality and ammonium, the urinary [Na]/[K] ratio (U(Na)/K), plasma aldosterone and TTKG in 7 patients with diarrhea, 6 with vomiting, 7 with mineralocorticoid excess, 6 with diuretic usage, and compared them with those of 7 overnight fasted and acid loaded healthy volunteers. RESULTS: The urine K+ concentrations did not reflect urinary loss of potassium according to the subjects' hydration status. U(Na)/k in the hypokalemic patients with mineralocorticoid excess (1.4 +/- 0.5) was lower than in normal subjects (2.3 +/- 0.4) (p<0.05). TTKG was higher in hypokalemic patients with mineralocorticoid excess (13.3 +/- 4.4) and diuretic usage (8.6 +/- 1.3) and lower in those with diarrhea (1.6 +/- 0.3) than in the normal controls (5.0 +/- 0.7) (p<0.5). TTKG in the patients with vomiting (3.5 +/- 0.6) was the same as in normal controls. TTKG was stronger correlated with the plasma aldosterone levels in the hypokalemic patients due to renal potassium loss. Urine ammonium concentrations of the acid-loaded normal subjects (73.3 +/- 5.0 mEq/L), patients with diarrhea (74.4 +/- 2.0 mEq/L) and patients with mineralocorticoid excess (68.7 +/- 6.9 mEq/L) were higher than in overnight-fasted normal subjects (31.3 +/- 4.9 mEq/L). CONCLUSION: TTKG and random urine ammonium were useful in the pathophysiological differential diagnosis of hypokalemia. PMID- 10858975 TI - Effect of valsartan on renal handling of uric acid in healthy subjects. AB - OBJECTIVE: To evaluate the effect of valsartan on renal handling of uric acid in healthy subjects. METHODS: A randomized, single-blind, placebo-controlled clinical trial was performed in twelve healthy volunteers. Six received a morning dose of valsartan 80 mg orally during 14 to 21 days and the other six received placebo. Before and after valsartan or placebo, clearance, fractional excretion and excretion uric acid rates were calculated. Presecretory reabsorption, tubular secretion and postsecretory reabsorption of uric acid were assessed by pharmacological tests using pyrazinamide or probenecid. RESULTS: There were no differences in clearance, fractional excretion and excretion uric acid rates with valsartan and placebo. Valsartan did not affect presecretory reabsorption (p = 0.92), tubular secretion (p = 0.62) or postsecretory reabsorption (p = 0.52) of uric acid. CONCLUSION: Valsartan did not significantly affect renal handling of uric acid in healthy subjects. PMID- 10858976 TI - Ultrastructural localization of advanced glycation end products and beta2 microglobulin in dialysis amyloidosis. AB - BACKGROUND: beta2-microglobulin (beta2m) is considered to be the amyloidogenic precursor in dialysis-related amyloidosis (DRA, Abeta2M amyloidosis). beta2m modified with advanced glycation end products (AGE) may be an important factor in the pathogenesis of DRA. The presence of AGE in beta2m-positive amyloid deposits and surrounding macrophages has been demonstrated by immunohistochemical techniques in light microscopy. METHODS: In order to better define the localization of beta2m and AGE in amyloid deposits and in cells, carpal tunnel connective tissues obtained from surgical specimens in six patients with DRA were studied by immunohistochemistry and electron microscopy, using the avidine biotine complex and immunogold staining procedures, respectively. A polyclonal rabbit anti-human beta2m and two monoclonal mouse anti-AGE antibodies [AG-1 anti imidazolone and AG-10 anti-N(epsilon)-carboxymethyl-lysine] enabled us to label their respective antigens at the optical and ultrastructural level. RESULTS: with both techniques, extracellular amyloid deposits strongly reacted with anti-beta2m and anti-AGE antibodies, although the immunoreactivity of beta2m was more intense. Macrophage-like synovial cells (CD-68 positive) surrounding amyloid deposits were also immunoreactive for beta2m and AGE, which were detected in lysosomes and in intracellular fibrillar material. Anti-AGE reactivity was also evident in collagenous structures in the absence of beta2m or amyloid deposits, supporting the proposal that AGE modification of collagen might have pathogenic relevance in the development of DRA. CONCLUSIONS: The co-localization of AGE and beta2m, both intra- and extra-cellularly, in amyloid fibrils was confirmed by immunoelectron microscopy; however, the positivity of collagen to anti-AGE antibodies and a different pattern of intracellular localization suggest that molecules other than beta2m may also be modified by AGE and may be involved in the pathogenesis of DRA. PMID- 10858977 TI - Deciding not to start dialysis--a one year prospective study in Teesside. AB - OBJECTIVES: To study the number and type of patients in whom it was decided not to commence dialysis for presumed irreversible renal failure, and the reasons for those decisions. DESIGN: A prospective twelve-month observational study. Information, regarding the decision making process, was obtained by proforma from the doctors involved. Patient details were retrieved from case notes. SETTING: The South Cleveland Hospital Renal Unit, which is the sole provider of chronic dialysis for a catchment population of 900,000 in Teesside and parts of South Durham and North Yorkshire. SUBJECTS: Patients referred for consideration for dialysis for presumed irreversible renal failure in whom it was decided not to commence dialysis. Data are included regarding all patients who did commence renal replacement therapy for end-stage renal failure over the same time period for demographic comparison. RESULTS: Eleven patients did not start dialysis, compared with 88 commencing RRT. In the undialysed patients, age range was 56 to 90, but six were 80 or over. Only one had previously attended a renal clinic, none had a specific renal diagnosis. Only 3 had no immediate indications for dialysis. Six patients were thought likely to die soon with or without dialysis. All 11 had significant co-morbidity, and 6 were recorded as housebound. Four patients were totally incapable of making a decision regarding dialysis; only one of the other seven was offered the choice. During the study period, both the acceptance rate and the early death rate for dialysis were higher than in previous years. CONCLUSIONS: A small number of patients with multiple medical and social problems are not selected for dialysis. Although some would have had their life prolonged by dialysis, about half were thought to be about to die from non renal causes. The patients were rarely involved in the decision making process. The process of being studied may have altered either decision making or classification of patients being offered dialysis. PMID- 10858978 TI - Physical function, employment and quality of life in end-stage renal disease. AB - INTRODUCTION: The impact of end-stage renal disease (ESRD) on quality of life (QoL) can be measured in terms of physical, psychological and social consequences, including the ability to work. SUBJECTS AND METHODS: This multi center, cross-sectional study explored relationships between QoL, employment status and physical function in ESRD patients aged 18-65 years, via a customised interviewer-administered questionnaire, which included the SF-36 health survey. The International Labour Office method was applied to describe employment rate. RESULTS: 144 patients (85 male, 49 female), comprising 49 haemodialysis (HD), 35 peritoneal dialysis (PD) and 60 renal transplant (TX) patients were studied. Mean age was 44 +/- 12 years. 32 were voluntarily not working, leaving 112 in the labour force. Of the latter, 49% were unemployed, in contrast with the concurrent national rate of 10%. QoL in the ESRD group was reduced in the SF-36 physical and social dimensions compared to population norms. Unemployed ESRD patients scored significantly lower than those employed in physical function, role physical, bodily pain, general health, vitality and role emotional scales. Logistic regression demonstrated that multiple comorbidities (p<0.005), a premorbid physical occupation (p<0.05) and poor physical function (p<0.05) predicted unemployment in ESRD independent of all other variables. Multiple regression showed that age (p<0.05), female sex (p<0.05) and a diagnosis of musculoskeletal disease (p<0.005) were independent predictors of poor physical function. CONCLUSIONS: These findings suggest that vocational rehabilitation of ESRD patients must consider physical function and occupational demands as well as co morbidity and that musculoskeletal disease is key factor in impaired physical function. PMID- 10858979 TI - Fate of kidneys from the same donor grafted into different recipients: do they behave similarly and are they influenced by donor-related factors? AB - BACKGROUND: The fate of paired kidneys might be similar and could therefore reflect the influence of donor-related factors on graft outcome. PATIENTS AND METHODS: To verify whether two kidneys retrieved from a single donor and grafted into different recipients have similar short, and middle-term outcomes we investigated the clinical outcome of 103 pairs of cadaveric kidneys grafted into 206 recipients. We evaluated the influence of donor-related factors such as age, sex and cause of death, and of the storage solution and method of harvesting. The incidence of delayed graft function was considered as the short-term outcome and serum creatinine levels at two years as the middle-term outcome. We evaluated the difference from expected frequencies in the incidence of delayed graft function and the incidence of similar serum creatinine levels in each pair of recipients. Univariate analysis of possible risk factors was made by the t-test, chi2 test and Fisher test, as appropriate. Multivariate analysis was done by logistic regression analysis with a forward stepwise variable selection. RESULTS: Delayed graft function was seen in both recipients from the same donor 2.5 times more than the expected frequency (p<0.001). Serum creatinine levels were similar in both recipients with a higher frequency than expected (p<0.01). Multivariate analysis showed that donor-related factors on graft function were age, cause of death and storage solution. CONCLUSIONS: Paired kidneys have similar performances in both the short and the long term. Major donor-related factors in delayed graft function are age and the storage solution. Major donor-related factors in graft function are age and cause of death. PMID- 10858980 TI - Bilateral emphysematous pyelonephritis with perirenal abscess cured by conservative therapy. AB - Emphysematous pyelonephritis is a rare life-threatening infection of the renal parenchyma. It usually affects unilateral kidney and occurs mostly in diabetic patients. It is characterized by the presence of gas within the renal parenchyma and requires prompt diagnosis and early aggressive therapy. Bilateral emphysematous pyelonephritis is even more rare and is associated with high mortality. We describe a case of a 62-year-old diabetic woman who presented with nonketotic hyperosmolar coma and bilateral emphysematous pyelonephritis caused by Klebsiella pneumoniae. Diagnosis of bilateral emphysematous pyelonephritis was confirmed by an abdominal computed tomographic scan and microbiologic studies. Our patient was successfully treated using percutaneous catheter drainage and long-term antibiotic therapy. PMID- 10858981 TI - Peanut and soy allergy: a clinical and therapeutic dilemma. PMID- 10858982 TI - Mechanisms of allergen-specific immunotherapy. PMID- 10858983 TI - Comparison of allergen-induced changes in bronchial hyperresponsiveness and airway inflammation between mildly allergic asthma patients and allergic rhinitis patients. AB - Bronchial eosinophilic inflammation and bronchial hyperresponsiveness (BHR) are the main features of allergic asthma (AA), but they have also been demonstrated in allergic rhinitis (AR), suggesting a continuity between both diseases. In spite of not fully reproducing natural allergenic exposure, the allergen bronchial provocation test (A-BPT) has provided important knowledge of the pathophysiology of AA. Our aim was to verify the existence of a behavior of AA and AR airways different from the allergen bronchial challenge-induced airway eosinophilic inflammation and BHR changes. We studied a group of 31 mild and short-evolution AA and 15 AR patients, sensitized to Dermatophagoides pteronyssinus. The A-BPT was performed with a partially biologically standardized D. pteronyssinus extract, and known quantities of Der p 1 were inhaled. Peripheral blood (eosinophils and ECP) and induced sputum (percentage cell counts, ECP, albumin, tryptase, and interleukin [IL]-5) were analyzed, before and 24 h after A-BPT. Methacholine BHR, assessed before and 32 h after the A-BPT, was defined by M-PD20 values and, when possible, by maximal response plateau (MRP). The A-BPT was well tolerated by all the patients. AA presented a lower Der p 1 PD20 and a higher occurrence of late-phase responses (LPR). M-PD20 values decreased in AA, but not in AR, patients. MRP values increased in both groups. Eosinophils numbers and ECP levels increased in blood and sputum from both AA and AR, but only the absolute increment of sputum ECP levels was higher in AA than AR patients (P = 0.025). The A-BPT induced no change in sputum albumin, tryptase, or IL-5 values. We conclude as follows: 1) In spite of presenting a lower degree of bronchial sensitivity to allergen, AR patients responded to allergen inhalation with an eosinophilic inflammation enhancement very similar to that observed among AA. 2) MRP levels increased in both AA and AR patients after allergen challenge; however, M-PD20 values significantly changed only in the AA group, suggesting that the components of the airway response to methacholine were controlled by different mechanisms. 3) It is possible that the differences between AR and AA lie only in the quantitative bronchial response to allergen inhalation. PMID- 10858984 TI - Quality of life and capsaicin sensitivity in patients with sensory airway hyperreactivity. AB - BACKGROUND: A group of patients with asthma-like symptoms and sensitivity to chemical irritants has shown an increased cough sensitivity to inhaled capsaicin compared to patients with asthma and to healthy controls. The condition is called sensory hyperreactivity (SHR), and the patients often feel that they are socially handicapped because of the risk of exposure to chemical irritants in daily life. METHODS: Twenty-six patients with asthma-like symptoms after exposure to nonspecific irritating stimuli, but without IgE-mediated allergy or demonstrable bronchial obstruction, were selected for a study of the response to a capsaicin test and measurement of quality of life by a general health profile (the Nottingham Health Profile [NHP]). We also investigated whether there was a correlation between quality of life and sensitivity to capsaicin. RESULTS: The patients demonstrated a dose-dependent response to the capsaicin provocation, with coughing and respiratory and other symptoms, that significantly differed from 12 healthy controls. The health profile showed that patients with SHR had a significantly reduced quality of life compared to reference values, and there was a significant correlation between the health profile and sensitivity to capsaicin. CONCLUSIONS: Patients with asthma-like symptoms verified by the capsaicin inhalation test for sensory hyperreactivity have a poor quality of life. The correlation between quality of life and sensitivity to capsaicin objectively demonstrates the validity of this general health profile study. PMID- 10858985 TI - Cough provocation with capsaicin is an objective way to test sensory hyperreactivity in patients with asthma-like symptoms. AB - BACKGROUND: A group of patients with asthma-like symptoms and sensitivity to chemical irritants, but without bronchial obstruction, has been found among subjects referred for suspected asthma. They have no well-defined diagnosis, and no objective diagnostic method has previously been available. These patients are more sensitive to inhaled capsaicin than are patients with asthma or healthy controls. The aim was to study cough and other capsaicin-induced symptoms and to test the effect of a drug (lidocaine) that inhibits nerve transmission in sensory nerves. METHODS: Twelve patients were provoked with three different concentrations of inhaled capsaicin solutions in a randomized, double-blind order. They all had asthma-like symptoms and were sensitive to chemical irritants, but had no IgE-mediated allergy or demonstrable bronchial obstruction. Before the provocations, the patients inhaled lidocaine or placebo (saline), also in a double-blind, randomized order. The results were expressed as the number of coughs and scores of various symptoms. RESULTS: The patients reacted in a dose dependent way with cough, airway, and eye symptoms, which were significantly reduced after preinhalation of lidocaine. CONCLUSIONS: A drug that inhibits transmission in sensory nerves successfully blocked the number of coughs and other symptoms provoked by inhalation of capsaicin. This indicates that the mechanisms underlying chemical sensitivity in these patients may originate in the sensory nervous system, and we call this condition "sensory hyperreactivity". PMID- 10858986 TI - Severe atopic dermatitis is associated with sensitization to staphylococcal enterotoxin B (SEB). AB - BACKGROUND: Staphylococcus aureus has been identified as a possible trigger factor in atopic dermatitis (AD). Some 30-60% of S. aureus strains isolated from patients with AD are able to produce exotoxins with superantigenic properties, mostly staphylococcal enterotoxins A, B, C, and D (SEA-D) and toxic shock syndrome toxin-1 (TSST-1). Recently, it was demonstrated that the presence of IgE antibodies to SEA and SEB is correlated with the severity of skin lesions in children with AD. To determine the relevance of staphylococcal enterotoxins in adult patients with AD, we investigated the relationship between the severity of skin lesions and sensitization to SEA and SEB. METHODS: Clinical severity was determined by the SCORAD index. Circulating IgE antibodies to SEA and SEB, serum eosinophil cationic protein (ECP) levels, and urine eosinophil protein X (EPX) levels were measured. RESULTS: The skin condition was significantly worse in patients sensitized to SEB than in unsensitized patients. Serum ECP and urine EPX levels were found to be significantly higher in SEB-sensitized patients, confirming the higher degree of cutaneous inflammation. CONCLUSIONS: Our results demonstrate a relationship between severity of skin lesions and sensitization to SEB in adult patients with AD, but a relationship between disease activity and sensitization to SEA could not be shown. PMID- 10858987 TI - 2S methionine-rich protein (SSA) from sunflower seed is an IgE-binding protein. AB - BACKGROUND: Sunflower seed contains 2S albumins that in other crops have been associated with allergenicity. The sunflower seed methionine-rich 2S albumin (SSA) may be an IgE-binding protein responsible for anaphylactic reactions in some sunflower seed-sensitive subjects. The objective was to demonstrate that SSA is an IgE-binding protein. METHODS: SSA was purified and the amino-acid sequence determined. The degree of purity of SSA was evaluated by silver staining, and its IgE-binding capacity by immunoblotting with serum from a subject with a convincing clinical history of anaphylaxis to sunflower seed. RESULTS: The amino acid sequence confirmed that the purified protein was the mature form of the methionine-rich storage protein SSA from sunflower seed (Helianthus annuus). The SSA was specifically recognized by IgE from the serum of the sunflower seed allergic subject. CONCLUSIONS: SSA is an IgE-binding protein, and subjects allergic to sunflower seed whose IgE binds to SSA are at risk of developing allergic reactions if they consume SSA. PMID- 10858988 TI - A double-blind, placebo-controlled oral challenge study with lyophilized larvae and antigen of the fish parasite, Anisakis simplex. AB - BACKGROUND: The third-stage larvae of Anisakis simplex may be a hidden source of allergens in fish. The objective was to determine whether the ingestion of lyophilized A. simplex larvae, or antigen, induces clinical symptoms in a group of A. simplex-sensitized patients. METHODS: Double-blind, placebo-controlled oral challenges were conducted in 11 individuals who had experienced allergic reactions after eating fish. Another patient had chronic urticaria unrelated to the ingestion of fish. All patients had positive skin tests and specific IgE determinations for A. simplex and negative skin tests to a battery of fish species. Conjunctival tests with A. simplex extracts were conducted in all patients and in five controls. The 12 patients received capsules containing either lactose or one, five, or 25 lyophilized larvae of A. simplex at 2-h intervals in a double-blind fashion. The highest single dose was 100 larvae. ECP and tryptase levels in serum were measured before and after the last oral challenge. Lyophilized antigen was also given to five patients. RESULTS: None of the 12 patients experienced a positive reaction after the ingestion of the placebo, the lyophilized larvae, or the antigen. Tryptase and ECP levels before and after challenges did not change significantly. Conjunctival provocation tests were positive in 11 out of the 12 patients and in none of the controls. CONCLUSIONS: The ingestion of 100 lyophilized A. simplex larvae, or its equivalent in antigen, does not induce clinical symptoms in individuals with a clinical history and laboratory findings of hypersensitivity to A. simplex. The data suggest that only the ingestion of live larvae may be capable of inducing allergic manifestations. PMID- 10858989 TI - IgE-binding activity to enzyme-digested ovomucoid distinguishes between patients with contact urticaria to egg with and without overt symptoms on ingestion. AB - BACKGROUND: We occasionally see egg-allergic children who develop contact urticaria to hen's egg despite the absence of the overt symptoms on ingestion. The mechanisms remain to be elucidated. METHODS: Twenty-one subjects with positive reactions to 20-min patch tests for egg-white antigens were divided into subgroups with positive (n = 10) and negative (n = 11) results to oral challenge tests by the same antigens. We measured IgE antibody for egg white and its components, and IgE-binding activities to digestive enzyme-treated ovomucoid by RAST inhibition. RESULTS: There were no significant differences in IgE antibody titers to egg white (positive vs negative: 30.3% vs 15.3%, P=0.130), ovomucoid (21.5% vs 10.2%, P= 0.078), ovotransferrin (9.9% vs 3.7%, P = 0.105), and lysozyme (3.4% vs 2.9%, P=0.944), except ovalbumin (16.8% vs 5.6%, P=0.024), between the positive and negative subjects in the provocation tests. In contrast, the concentration (1.93 microg/ml) of pepsin-treated ovomucoid needed for 50% RAST inhibition in the challenge-positive subjects was significantly (P=0.0003) lower than that (114.9 microg/ml) of negative subjects. Similar but less significant differences were obtained when ovomucoid fragments treated with chymotrypsin (0.91 microg/ml vs 6.86 microg/ml, P=0.014) and trypsin (0.75 microg/ml vs 4.67 microg/ml, P= 0.041) were used as inhibitors. CONCLUSIONS: We suggest that IgE antibodies from subjects showing contact urticaria despite the absence of reactions to the ingestion of egg white recognize the epitope(s) unstable to digestive enzymes. PMID- 10858990 TI - Fel d 1 production in the cat skin varies according to anatomical sites. AB - BACKGROUND: Fel d 1 is the major cat allergen, inducing asthma in sensitized individuals. It is produced by the skin and lies on fur. Recently, it was established that the amount of Fel d 1 on fur varies among anatomical sites. However, it is not known whether the allergen production by skin varies in parallel. The objective was to compare the Fel d 1 production by male cat skin in two anatomical sites, the face and the chest, in order to correlate it with Fel d 1 amounts on fur, and to assess the reaccumulation of Fel d 1 after washing. METHODS: Ten intact male cats were shaved under general anesthesia at both areas, and the fur was collected. The skin was washed and the washing fluid collected for Fel d 1 assays. Fel d 1 levels were measured in microg/g fur and ng/cm2 skin by ELISA before and after washing and 48 h later. RESULTS: In skin washing, the mean Fel d 1 level was significantly higher in the face (1015.2 +/- 821.6 ng/cm2) than the chest (115.2 +/- 66.8 ng/cm2). In the fur, the respective levels were 63.6 +/- 34 and 29.6 +/- 13.6 microg/g. In the skin sample taken after skin washing, the level of Fel d 1 dropped to 25.1 +/- 25.7 ng/cm2 on the face and to 22 +/- 17.4 ng/cm2 on the chest. After 2 days, skin Fel d 1 levels returned to basal values, with higher values on the face than the chest. CONCLUSIONS: This study shows that Fel d 1 levels on the skin are dramatically higher on the facial area than chest. This anatomical variation is concordant with the levels of Fel d 1 found on fur. Washing reduces levels of major allergen on cat skin and fur, but the accumulation on skin is restored within 2 days. PMID- 10858991 TI - Evidence of very delayed clinical reactions to cow's milk in cow's milk intolerant patients. AB - BACKGROUND: In patients with cow's milk protein intolerance (CMPI), delayed clinical reactions to cow's milk (CM) ingestion may be misdiagnosed if the clinical symptoms are not "classical" and there is a long time lapse between ingestion of CM and the clinical reaction. The aim was to evaluate the clinical outcome of CMPI in a cohort of CM-intolerant children, with particular attention to the occurrence of clinical manifestations beyond 72 h after CM challenge. METHODS: Eighty-six consecutive patients (44 boys, 42 girls) with new CMPI diagnoses were enrolled; median age at diagnosis was 4 months. Patients were followed up for a mean period of 40 months. In all patients, CMPI diagnosis was made on the observation of symptoms, their disappearance after elimination diet, and their reappearance on double-blind CM challenge. At CMPI diagnosis, immunologic tests to demonstrate IgE-mediated hypersensitivity were performed. After 12 months of CM-free diet, CM tolerance was re-evaluated with a CM challenge continued at home for up to 30 days, according to a double-blind, placebo-controlled method. Patients who did not achieve CM tolerance continued a CM-free diet and subsequently underwent yearly CM challenge. RESULTS: The percentages of CMPI patients who became CM-tolerant after 1, 2, and 3 years of CM free diet were 30%, 54.5%, and 70%, respectively. At the end of the follow-up period, 26/86 subjects showed persistent CMPI; these patients had a higher percentage of positivity of total serum IgE (P<0.05), RAST (P<0.01), and cutaneous prick tests for CM antigens (P<0.001) than all the others. At CMPI diagnosis, all patients had a clinical reaction within 72 h from the beginning of the CM challenge; at the subsequent "cure" challenges, we observed patients who first reacted to CM more than 72 h after ingestion. In total, 10 out of 86 patients showed "very delayed reactions"; in these patients, the mean time between the beginning of CM challenge and the onset of a clinical symptom was 13.3 days (range 4-26 days). The number of "very late reactors" increased from the first to the third of the "cure" CM challenges, performed at yearly intervals. The "very delayed" CMPI manifestations in these subjects were constipation (five cases), wheezing (two cases), dermatitis plus constipation (two cases), and dermatitis alone (one case); in 6/10 patients, the symptoms observed at the "cure challenge" were different from those at CMPI onset. CONCLUSIONS: Very delayed clinical reactions to reintroduction of CM in the diet can occur in CMPI patients; thus, accurate follow-up and frequent outpatient observation in patients with a long history of CMPI are probably more useful and safer than prolonged CM challenge. PMID- 10858992 TI - Hypogammaglobulinemia resembling BOOP. PMID- 10858993 TI - Hypersensitivity reaction to ioversol. PMID- 10858994 TI - Ultrarush SIT in venom allergy. PMID- 10858995 TI - Urticaria developing into mastocytosis. PMID- 10858996 TI - Food allergy to frog. PMID- 10858997 TI - Exercise-induced anaphylaxis. PMID- 10858998 TI - Unusual sensitization to Anisakis simplex. PMID- 10858999 TI - Latex and amoxicillin-induced asthma. PMID- 10859000 TI - Ductal carcinoma in situ of the breast--a review of diagnosis, treatment and outcome in a hospital-based Norwegian series. AB - Between 1980 and 1994, 71 women with histologically proven ductal carcinoma in situ (DCIS) were diagnosed at Ulleval Hospital; bilateral tumours were found in two patients. Surgical treatment was mastectomy (42 lesions) or local excision (31 lesions). Median follow-up time was 7.2 years. Ten patients experienced a local recurrence, seven of which were invasive carcinomas. The actuarial 5-year local recurrence rate was 22% after local excision. A multivariate analysis found that tumour size was the only factor that predicted local recurrence after local excision. An analysis of relative survival in a nation-wide material of 832 DCIS patients in the period 1980 to 1994 demonstrates that relative survival after a DCIS diagnosis is almost 100%, irrespective of surgical treatment of the initial lesion. PMID- 10859001 TI - Primary breast lymphomas--a retrospective analysis of twelve cases. AB - This study was undertaken to define the natural history and treatment results of patients with primary breast non-Hodgkin's lymphoma (NHL). Twelve female patients who had been followed at Hacettepe University Hospital between 1973 and 1997 were retrospectively evaluated. All patients presented with breast masses (6 in the right breast and 6 in the left) that had recently enlarged. The most common histologic subtype was diffuse, small cleaved-cell lymphoma. Chemotherapy regimens were employed in 9 patients. Radiotherapy was delivered to the breast and its lymphatics in 8 patients. Lumpectomy, simple or modified radical mastectomy was performed in 5 cases. An objective response was attained with surgery, chemotherapy, or radiotherapy alone in 2, 1, and 1 cases, respectively. Combined modality treatment including either two or three modalities was successful in 7 cases. The median progression-free and overall survival times were 49 and 56 months, respectively. Although primary NHL of the breast is a rare disease compared to carcinoma, it should be considered in the differential diagnosis of breast masses. PMID- 10859002 TI - Metallothione in levels in cell fractions from breast cancer tissues. AB - Metallothioneins (MTs) protect the cell against reactive forms of oxygen, ionizing radiation, pharmacological agents and mutagens. Metallothioneins are also responsible for neoplastic cell resistance to cytostatic drugs. The aim of this study was to determine the MT level in cell fractions as well as to determine whether there is any change in the concentration of these proteins in a neoplastic cell, and in which cell fractions this change takes place. The neoplastic tissue examined was histopathologically ductal carcinoma invasive, and the control tissue was mastopathic tissue with proliferated connective and glandular tissue. The level of MTs was determined using cadmium isotope (109Cd). It was determined that there was an increase in MT level in the neoplastic tissue, the highest level being found in the mitochondrial fraction obtained from the control and mastopathic tissues. The greatest changes in MT concentrations in breast carcinoma were observed in the nuclear and cytosol fractions. In the nuclear fraction in the breast carcinoma tissue, the MT level was almost three times that of the control group. PMID- 10859003 TI - Increased urinary albumin excretion rate in breast cancer patients. AB - A high frequency of slightly increased urinary albumin excretion (UAE) has been reported in patients with malignancies. Earlier studies have indicated a prognostic significance of UAE in some malignant diseases. We measured urinary albumin in 24-h urine samples in 44 patients with newly diagnosed early breast cancer and in 22 patients with relapse of metastatic breast cancer disease. The prevalence of microalbuminuria ( > or = 20 microg/min) was 20.5% in patients with early breast cancer and 54.5% in patients with metastatic disease. Median UAE was significantly higher in patients with metastatic breast cancer compared with the early breast cancer group (20.5 microg/min vs. 9.2 microg/min; p < 0.01). In patients with metastatic breast cancer, univariate survival analysis revealed a significantly lower survival rate in patients with microalbuminuria compared with the normoalbuminuric group (p <0.001). The present study demonstrates a high frequency of microalbuminuria in patients with breast cancer. Increased UAE was most prevalent in patients with metastatic disease. Our results also suggest that UAE may be a prognostic marker in metastatic breast cancer. Further prospective studies with a larger number of patients and controls are needed to test the validity of these observations. PMID- 10859004 TI - Apoptosis and the pathogenesis of lymphoma. AB - The pathogenesis of follicular lymphoma and chronic lymphocytic leukemia (CLL) involves the dysregulation of the bcl-2 protein. This protein is a critical regulator of apoptosis and interferes with the caspase cascade, which is the backbone of the apoptotic machinery. The multiple interactions of caspases with bcl-2 and its related proteins which comprise the bcl-2 family of proteins have become more clear over the past few years and its importance in carcinogenesis is increasingly appreciated. The network of proteins of both the apoptotic pathway and the cell cycle regulators and their interactions and mutual regulations are in the process of being elucidated. It is hoped that this will soon be translated to clinical benefit for patients with lymphomas. PMID- 10859005 TI - Dose-volume analysis of different stereotactic radiotherapy mono-isocentric techniques. AB - Several stereotactic irradiation techniques, using Linacs with the patient in lying and sitting position and a Gamma Knife Unit, were compared with regard to mono-isocentric three-dimensional dose distributions. Three types of target volumes, a sphere and two ellipsoids, were used for the comparisons. All three targets were centered on a real head, reconstructed from transversal CT scans. The ARTEMIS 3D Treatment Planning System, developed by the Tenon Hospital, Paris, was used for the dosimetry and the dose-volume histogram (DVH) calculation. For the comparative study, several quantitative parameters were used, derived from the dose-volume histogram calculation. Differential DVHs were plotted for each target volume and beam arrangement. Irradiation techniques were compared by deriving quantitative parameters from the DVHs such as mean and integral dose delivered to the target and normal tissue irradiated, as well as by the relative volume of the examined areas. All techniques used in this study produced very similar dose distributions. The small differences confirm the capability of the studied techniques to produce the same irradiation effects. By changing from the spherical target shape to a more elliptical shape, more of the normal tissue was irradiated with higher doses. For elliptical cases we therefore identified a need for more conformal stereotactic planning. PMID- 10859006 TI - Comparative analysis of dose volume histogram reduction algorithms for normal tissue complication probability calculations. AB - A model for estimating radiotherapy treatment outcome through the probability of damage to normal tissue and the probability of tumour control is a useful tool for treatment plan optimization, dose escalation strategies and other currently used procedures in radiation oncology. Normal tissue complication estimation (NTCP) is here analysed from the point of view of the reliability and internal consistency of the most popular model. Five different dose volume histogram (DVH) reduction algorithms, applied to the Lyman model for NTCP calculation. were analysed and compared. The study was carried out for sets of parameters corresponding to quite different expected dose-response relationships. In particular, we discussed the dependence of the models on the parameters and on the dose bin size in the DVH. The sensitivity of the different reduction schemes to dose inhomogeneities was analysed, using a set of simple DVHs representing typical situations of radiation therapy routine. Significant differences were substantiated between the various reduction methods regarding the sensitivity to the degree of irradiation homogeneity, to the model parameters and to the dose bin size. Structural aspects of the reduction formalism allowed an explanation for these differences. This work shows that DVH reduction for NTCP calculation has still to be considered as a very delicate field and used with extreme care, especially for clinical applications, at least until the actual formulations are tuned against strong clinical data. PMID- 10859007 TI - Health-related quality of life and occurrence of intestinal side effects after pelvic radiotherapy--evaluation of long-term effects of diagnosis and treatment. AB - Health-related quality of life (HRQOL) and occurrence of late intestinal side effects were assessed 3-4 years after pelvic radiotherapy for carcinoma of the endometrium and cervix. During 1988-1990, 143 women were included in a clinical trial to evaluate the effect of a low fat, low lactose diet on radiation-induced diarrhoea. Of 94 survivors, 79 (84%) answered the request. HRQOL was assessed by the EORTC QLQ-C36 and compared with population-based norms. The women scored lower than the general population on role functioning (81.5 versus 90.6 (p < 0.01)) and higher on diarrhoea (23.8 versus 9.5 (p < 0.01)). Compared with pre treatment conditions, an increase in cases with pain in the lower back, hips and thighs was seen. Substantial pain and diarrhoea were associated with deterioration in HRQOL. In conclusion, few treatment and/or disease-related effects were detected 3-4 years after radiotherapy, with the exception of increased bowel frequency and pain in the lower back, hips and thighs. PMID- 10859008 TI - Time course of radiological lung density changes after postmastectomy radiotherapy. AB - The purpose of this study was to quantify the time course of radiological lung changes in patients after postmastectomy radiotherapy assessed from routine follow-up chest x-rays. Radiological density changes in the apex of the irradiated lung were quantified by a recent lung densitometry assay. Lung changes were expressed as the so-called Relative change in Equivalent Absorber Thickness (REAT). The clinical series comprised 329 patients treated with postmastectomy radiotherapy between 1978 and 1982. Of these patients 100 were treated with chemotherapy (CTX or CMF) and 41 were given tamoxifen as an additional adjuvant treatment; 290 patients (88.2%) had pretreatment x-rays and at least one x-ray after completion of radiotherapy and these were included in the study. A total of 2,209 chest x-rays was taken during follow-up, and among these 1921 x-rays (86.9%) were judged to be assessable. Late changes were defined as changes occurring more than a year after radiotherapy. A total of 280 patients had at least one chest x-ray taken more than one year after radiotherapy and these were evaluable with respect to late changes. There were 1,390 follow-up x-rays in these patients and 207 patients (73.9%) had three or more follow-up x-rays. Linear regression of REAT vs. observation time was used to identify three patterns of time changes: progressive, regressive, and stable. The results were as follows. Two phases of lung changes were observed. The early phase peaks around 6 months and the density changes may subsequently resolve, completely or in part. In most cases (173 patients or 84%), the lung density changes reached a stable level 12 months after irradiation. Yet, in 16 patients (7.7%) the lung changes progressed for five or more years. Regression of the density changes was seen in 18 patients (8.7%), and in some cases there were signs that this apparent regression was caused by contraction of the fibrotic tissue. We conclude that two phases of lung response can be distinguished and can be graded according to severity using this assay. Early lung changes reach a maximum around 6 months after RT. Late reactions reach a plateau in most patients after one year, but progress in some cases for five or more years. PMID- 10859009 TI - Health, life and disability insurance and hereditary risk for breast or colorectal cancer. AB - Fear of insurance discrimination affecting the insurance-seeker and family has been reported as the singlemost important reason why individuals choose not to undergo genetic testing. The eleven health insurers operating on the Norwegian market were mailed a questionnaire asking them to list their insurance products and evaluate two individuals' requests for insurance. The requests were constructed in order to illustrate a high genetic risk for (a) colorectal (HNPCC) and (b) breast cancer (BRCAI/BRCA2), respectively. Nine out of 11 insurers responded. While no restriction was documented concerning risk of BRCA1/BRCA2 and life insurance or disability pension, the premium paid by persons with susceptibility to HNPCC varied between the different insurers from standard to raised premiums. The product 'critical disease' insurance was refused or obtained at normal or raised premiums in both cases, depending on the insurer in question. On examining personal indemnity insurance, we found that the BRCA1/BRCA2-risk individual was offered insurance at the standard premium, whereas HNPCC-risk individuals were offered a standard or raised premium. Only the major Norwegian insurer is in fact diverging in its policies. PMID- 10859010 TI - Value of transsternal core biopsy in patients with a newly diagnosed mediastinal mass. AB - Histopathologic analysis of an anterior mediastinal mass of unknown origin is essential for treatment decision. Mediastinoscopy is the most common procedure performed to obtain biopsies, but general anaesthesia and hospitalization are necessary. The aim of this study was to evaluate whether transsternal core biopsies, an easy outpatient biopsy technique, could be an alternative to mediastinoscopy. A biopsy instrument that makes it possible to reach tumours hidden behind bone was used for transsternal CT-guided core biopsies in 21 patients with a newly diagnosed anterior mediastinal mass. No severe side effects were observed. In 19/21 (90%) patients the biopsies were diagnostic. In 2/21 patients additional biopsy techniques had to be used. In these two patients Hodgkin's disease was suspected in the first biopsy procedures. The diagnosis was confirmed by new core biopsies, from other parts of the tumour, not using a transsternal approach (transclavicular and parasternal, respectively). In addition, one mediastinoscopy was performed in a patient who was diagnosed with a non-Hodgkin's lymphoma but where more material was needed for lymphoma subclassification. It is concluded that CT-guided transsternal core biopsy is a clinically valuable method in patients with a newly diagnosed anterior mediastinal mass. PMID- 10859011 TI - Effect of nitroimidazole sensitizers on in vitro glycolytic metabolism of hypoxic squamous cell carcinoma. AB - Two nitroimidazole compounds, misonidazole (MISO) and nimorazole (NIMO), were evaluated for their potential to modify uptake of [5,6-3H] 2-fluoro-2-deoxy-D glucose (3H-FDG) in the human squamous carcinoma cell line UT-SCC-5 exposed to increasing levels of hypoxia. UT-SCC-5 cells were incubated with 0-10 mM of MISO or NIMO under normal or reduced oxygen concentrations of 20%, 1.5%, or 0% with 5% CO2 for 6 h, after which 74 KBq of 3H-FDG was added in media for 1 h. In the presence of normal concentrations of O2, both sensitizers increased 3H-FDG uptake by up to 178% (MISO) or 84% (NIMO) when compared with untreated cells. In anoxia, MISO decreased 3H-FDG uptake to 35% of that of control whereas NIMO-treated cells showed a respective decrease in tracer uptake to 62%. Clonogenic assays clearly indicated that MISO was toxic and NIMO moderately toxic for hypoxic cells, whereas both sensitizers exerted only a very modest effect on survival of fully oxygenated cells. Our findings indicate that nitroimidazole treatment consistently increases 3H-FDG uptake into UT-SCC-5 cells under normal oxygen concentrations. In hypoxia, the observed decrease in tracer uptake is dependent on both the level of ambient oxygen and drug concentration and may reflect both direct toxicity and inhibition of glycolysis. The observations may be useful for further applications of 18F-FDG positron emission tomography (PET) to monitor effects of hypoxic cell radiosensitizers on tumor metabolism in vivo. PMID- 10859012 TI - High intratumoral accumulation of stealth liposomal doxorubicin in sarcomas- rationale for combination with radiotherapy. AB - Sarcomas are radioresistant tumors, the only curative therapy being radical surgical resection. Stealth liposomal doxorubicin (Caelyx) is a novel drug formulation that allows prolonged circulation and high intratumoral concentration. This study investigates the concurrent use of radiotherapy with Caelyx in a cohort of 7 patients with locally advanced or recurrent sarcoma. Radiotherapy was given as a standard fractionation regimen to a total dose of 70 Gy. Caelyx was given as a 30-min infusion at a dose of 25 mg/m2 every 2 weeks. Scintigraphic imaging with Caelyx-99mTc-DTPA showed an increased (2.8 +/- 0.9 times higher) intratumoral drug accumulation compared to the surrounding healthy tissue. The regimen was well tolerated without any severe hematological or systemic toxicity. 'In field' radiation toxicity was not increased. Complete response was observed in 4/7 cases. It is concluded that combined chemo radiotherapy with stealth liposomal doxorubicin for locally advanced sarcomas is feasible and promising, the benefit expected from the unique ability of the stealth liposomes to accumulate selectively in the tumoral tissue. PMID- 10859013 TI - Symptoms of laryngeal carcinoma and their prognostic significance. AB - Symptoms of 301 patients with laryngeal squamous cell carcinoma were evaluated. Tumour site affected the symptom profile significantly. Hoarseness was more common among patients with glottic and subglottic tumours, but was also the leading symptom in supraglottic patients. Other symptoms were mainly associated with supraglottic tumours and more advanced glottic lesions. There was a positive correlation between the number of symptoms and stage, regardless of tumour site. Patients with a supraglottic tumour had significantly more symptoms (median 2) than those with a glottic lesion (median 1). The median duration of the symptoms was significantly longer in stages III-IV (4.7 months) than in stages I-II (3.8 months). In the multivariate analysis, tumour stage was the only variable showing prognostic significance. PMID- 10859014 TI - Results of prophylactic irradiation in patients with resected keloids--a retrospective analysis. AB - The data of 139 patients with 166 keloids treated postoperatively between 1962 and 1996 were evaluated for prognostic factors and outcomes. Treatment commenced within 48 h after surgery. Radiotherapy was carried out as brachytherapy, using an integrated radionuclide 90 Sr-90Y surface applicator. The median dose delivered to the subcutis amounted to 14 Gy (range 7.5-28.5 Gy). The overall recurrence-free response rate was calculated to be 80% for all keloids. Response rates differed significantly (p < 0.001) between the different anatomical regions. The recurrence rate was lowest (2%) with keloids of the face and neck and highest with keloids of the thorax (49%). Outcome also differed significantly, depending on the etiology. Keloids following burns had a poorer success rate than those developing after surgery or mechanical trauma (p < 0.001). We were unable to demonstrate any significance in outcome related to gender, age or size. No direct correlation was found between total doses and response rates. In our patients there were no signs of secondary malignancies in the irradiation area within a median follow-up period of 12 years. Two new prognostic factors have been identified: keloid etiology and localization of the disorder. PMID- 10859015 TI - Efficacy and tolerability of tropisetron in the prevention of cisplatin-induced nausea and vomiting in advanced non-small cell lung cancer. AB - The efficacy and tolerability of tropisetron were studied in an open trial comprising a total of 30 patients with advanced non-small cell lung cancer undergoing high-dose, cisplatin-based chemotherapy (cisplatin dosage 100 mg/m2). Patients received tropisetron 5 mg intravenous infusions for 15 min on day 1. followed by 5 mg tropisetron taken orally in the morning on days 2 6. All treated patients were assessed during the entire treatment period (6 days). Acute nausea and vomiting were evaluated during the 24 h after chemotherapy. Delayed nausea and vomiting were evaluated during days 2-6 after chemotherapy. Response to tropisetron was graded as: complete control, major control, minor control and failure for nausea or vomiting. Rates for complete plus major control of acute nausea and vomiting in cycles 1-5 were 77%, 81%, 86%, 67% and 75%, respectively. Rates for complete plus major control of delayed nausea and vomiting in cycles 1 5 were 87%, 76%, 86%, 78% and 75%, respectively. Adverse reactions were mainly headache and diarrhea, but both reactions were mild and are common in most patients treated with this type of antiemetic agent. It is concluded that tropisetron is an effective drug for the prevention of side effects of highly emetogenic drugs such as cisplatin. The dosage and schedule of tropisetron reported here can prevent both acute and delayed nausea and vomiting. PMID- 10859016 TI - Phase II trial of docetaxel in Asian patients with inoperable stage III non-small cell lung cancer. AB - Docetaxel has a response rate of greater than 30% in first-line treatment of Western patients with advanced non-small cell lung cancer (NSCLC). The goal of this open-label. phase II study was to evaluate the activity and safety profile of docetaxel in Asian patients with inoperable untreated stage III NSCLC. Docetaxel was given at 100 mg/m2 as a 1-h infusion every 3 weeks. Prophylactic dexamethasone was given to reduce hypersensitivity reactions and edema. Thirty five patients were enrolled in the study. The response rate was 34% (95% CI, 19% 50%) according to intent-to-treat analysis. No complete response was observed. Twenty-four patients (69%) had grade 3 or 4 neutropenia in cycle 1, and febrile neutropenia was seen in 12 patients. Six patients (17%) experienced mild fluid retention. Docetaxel is an active agent in first-line treatment of Asian patients with locally advanced NSCLC, with the main toxicity being neutropenia. Fluid retention was a minor problem in this study. PMID- 10859017 TI - Esthesioneuroblastoma--what is the optimal treatment? AB - A retrospective review was conducted on 13 patients with esthesioneuroblastoma (ENB), treated at our institution from 1977 to 1997. According to the Kadish classification, one patient was in stage A, 5 patients were classified as stage B and 7 patients were in stage C. Five-year disease-specific survival was found to be 51%. Forty-six percent of the patients experienced relapse and despite intensive salvage therapy, median survival after recurrences was only 12 months. This indicates the need for good primary control in local as well as distant disease. The role of pre- versus postoperative radiotherapy to secure good local control is discussed and compared with the literature, and treatment guidelines are proposed. The tumours were graded according to the Hyams' classification and its importance as a prognostic factor is briefly discussed. PMID- 10859018 TI - Testicular seminoma and diabetes in twins. PMID- 10859019 TI - Seizures in a patient with disseminated testicular cancer due to cisplatin induced hypomagnesaemia. PMID- 10859020 TI - Hypereosinophilic syndrome in acute lymphoblastic leukaemia--case report and literature review. PMID- 10859021 TI - A young female with an endodermal sinus tumor in a pericardial localized cyst. PMID- 10859022 TI - Hypoxia, normoxia and hyperoxia--terminology for medical in vitro cell biology. PMID- 10859023 TI - The Joslin Diabetes Center. PMID- 10859024 TI - HIV infections: the global epidemiology and goals for vaccine research. PMID- 10859025 TI - Differentiation of human embryonic stem cells into embryoid bodies compromising the three embryonic germ layers. AB - BACKGROUND: Embryonic stem (ES) cells are lines of cells that are isolated from blastocysts. The murine ES cells were demonstrated to be true pluripotent cells as they differentiate into all embryonic lineages. Yet, in vitro differentiation of rhesus ES cells was somewhat inconsistent and disorganized. The recent isolation of human ES cells calls for exploring their pluripotential nature. MATERIALS AND METHODS: Human ES cells were grown in suspension to induce their differentiation into embryoid bodies (EBs). The differentiation status of the human ES cells and EBs was analyzed by following the expression pattern of several lineage-specific molecular markers using reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization. RESULTS: Here we report the induction in vitro of cystic embryoid bodies from human ES cells. Our findings demonstrate induction of expression of cell-specific genes during differentiation of the human ES cells into EBs. In the human EBs, we could show a characteristic regional expression of embryonic markers specific to different cellular lineages, namely, zeta-globin (mesoderm), neurofilament 68Kd (ectoderm), and alpha fetoprotein (endoderm). Moreover, we present a synchronously pulsing embryoid body that expresses the myocardium marker alpha-cardiac actin. In addition, dissociating the embryoid bodies and plating the cells as monolayers results in multiple morphologies, among them cells with neuronal appearance that express neurofilament 68Kd chain. CONCLUSION: Human ES cells can reproducibly differentiate in vitro into EBs comprising the three embryonic germ layers. The ability to induce formation of human embryoid bodies that contain cells of neuronal, hematopoietic and cardiac origins will be useful in studying early human embryonic development as well as in transplantation medicine. PMID- 10859026 TI - Intravenous injection of an adenovirus encoding hepatocyte growth factor results in liver growth and has a protective effect against apoptosis. AB - BACKGROUND: Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic cytokine with mitogenic, motogenic and morphogenic effects for a wide variety of cells. Previous studies have reported that the in vivo infusion in normal, untreated mice of recombinant HGF results in low levels of DNA synthesis and liver proliferation. In this study, we examined whether liver regeneration could be obtained by the in vivo injection of a recombinant adenoviral vector encoding human HGF (Ad.CMV.rhHGF) in normal, intact mice. MATERIALS AND METHODS: C57BL/6 mice were infused intravenously with doses increasing from 1 to 4 x 1011 particles of the recombinant human HGF (rhHGF) adenoviral vector or with a control virus encoding Escherichia coli beta-galactosidase (Ad.CMV.lacZ). At day 5, mice were sacrificed and evaluated for the presence of hepatocyte mitogenesis and liver regeneration (5-bromo-2'-deoxyuridine (BrdU) assays and liver weight determination) and for the presence of liver damage (serum alanine amino transferase (ALT) measurements and TUNEL assays). RESULTS: In vivo administration of rhHGF stimulated DNA synthesis of hepatocytes and liver weight in a dose dependent fashion. The maximal effect was seen after the infusion of 3 x 1011 particles which resulted at day 5 in >130% increase in relative liver mass with little cytopathic effect. In contrast, administration of the lacZ adenoviral vector caused little hepatocyte replication, but induced high levels of serum ALT (approximately 3 times higher than the rhHGF vector) and significant apoptotic cell death. CONCLUSIONS: This study shows that a single injection of Ad.CMV.rhHGF alone is able to induce in vivo and in a very short period of time, robust DNA synthesis and liver proliferation in normal mice without liver injury or partial hepatectomy. This recombinant adenoviral vector has a lower toxicity than the control lacZ adenovirus. This suggests that HGF may have a protective effect against adenovirus-induced pathology. PMID- 10859027 TI - X-chromosome inactivation and mutation pattern in the Bruton's tyrosine kinase gene in patients with X-linked agammaglobulinemia. Italian XLA Collaborative Group. AB - BACKGROUND: The diagnosis of X-linked agammaglobulinemia (XLA) is not always clearcut. Not all XLA conform to the classic phenotype and less than 50% of affected boys have a family history of immunodeficiency. Mutations in the gene for Bruton's tyrosine kinase (BTK) are responsible for the majority of agammaglobulinemia cases. However, a certain proportion of patients may have mutations involving other genes, although they show with an XLA phenotype. We performed BTK gene mutation analysis in 37 males with presumed XLA and analyzed the pattern of X-chromosome inactivation (XCI) in 31 mothers to evaluate the relevance of these approaches to diagnosis and genetic counseling. MATERIALS AND METHODS: Twenty affected males with a sporadic occurrence and 17 familial cases belonging to nine families were enrolled within the framework of the Italian Multicenter Clinical Study on XLA. We used non-isotopic RNase cleavage assay (NIRCA), followed by cDNA sequence determination to screen for BTK mutations and X-chromosome inactivation analysis for carrier detection. RESULTS: Using the cDNA based approach, the identification of BTK gene abnormalities confirmed the clinical diagnosis of XLA in 31 of 37 affected infants. Missense was the most frequent mutational event and the kinase domain was mostly involved. In addition, nine novel mutations were identified. In sporadic cases, BTK gene abnormalities were identified in 9 out of 10 patients whose mothers had a nonrandom pattern of XCI and in 5 out of 6 patients whose mother had a random pattern of XCI. With the exception of one family, all patients with a familial occurrence and born to mothers with a nonrandom pattern of XCI had mutations of the BTK gene. CONCLUSIONS: Our findings indicate that in sporadic cases BTK gene sequencing is the only reliable tool for a definitive diagnosis of XLA and support XCI as the first diagnostic tool in the mothers of affected males in multiple generations. Furthermore, our molecular analysis confirms that 10-20% of BTK-unaltered patients have disorders caused by defects in other genes. PMID- 10859028 TI - Immunological evidence that non-carboxymethyllysine advanced glycation end products are produced from short chain sugars and dicarbonyl compounds in vivo. AB - BACKGROUND: The Maillard reaction that leads to the formation of advanced glycation end-products (AGE) plays an important role in the pathogenesis of angiopathy in diabetic patients and in the aging process. Recently, it was proposed that AGE were not only created by glucose, but also by dicarbonyl compounds derived from the Maillard reaction, autoxidation of sugars and other metabolic pathways of glucose. In this study, we developed four types of non carboxymethyllysine (CML) anti-AGE antibodies that recognized proteins modified by incubation with short chain sugars and dicarbonyl compounds. MATERIALS AND METHODS: AGE-modified serum albumins were prepared by incubation of rabbit serum albumin with glyceraldehyde, glycolaldehyde, methylglyoxal or glyoxal. After immunization of rabbits, four types of AGE-specific antisera were obtained that were specific for the AGE modification. To separate non-CML AGE antibodies (Ab) (non-CML AGE-Ab-2, -3, -4, and -5), these anti-AGE antisera were subjected to affinity chromatography on a matrix coupled with four kinds of AGE bovine serum albumin (BSA) or CML-BSA. These non-CML AGE antibodies were used to investigate the AGE content of serum obtained from diabetic patients on hemodialysis. RESULTS: Characterization of the four types of non-CML AGE antibodies obtained by immunoaffinity chromatography was performed by competitive ELISA and immunoblot analysis. Non-CML AGE-Ab-2 crossreacted with the protein modified by glyceraldehyde or glycolaldehyde. Non-CML AGE-Ab-3 and -Ab-4 specifically cross reacted with protein modified by glycolaldehyde and methylglyoxal, respectively. NonCML AGE-Ab-5 cross-reacted with protein modified with glyoxal as well as methylglyoxal and glycolaldehyde. Three kinds of non-CML AGE (AGE-2, -4, and -5) were detected in diabetic serum as three peaks with apparent molecular weights of 200, 1.15, and 0.85 kD; whereas, AGE-3 was detected as two peaks with apparent molecular weights of 200 and 0.85 kD. CONCLUSION: We propose that various types of non-CML AGE are formed by the Maillard reaction, sugar autoxidation and sugar metabolism. These antibodies enable us to identify such compounds created by the Maillard reaction in vivo. PMID- 10859029 TI - A secreted tumor-suppressor, mac25, with activin-binding activity. AB - BACKGROUND: mac25 is a follistatin (FS)-like protein that has a growth suppressing effect on a p53-deficient osteosarcoma cell line (Saos-2). The protein exhibits a strong homology to FS, an activin-binding protein, and part of its sequence includes the consensus sequence of the member of the Kazal serine protease inhibitor family. MATERIALS AND METHODS: Localization of mac25 protein was analyzed using mac25 protein fused with green fluorescent protein (GFP). Recombinant mac25 protein was expressed in E. coli and purified. The recombinant mac25 protein was added in culture medium for analysis of growth suppression and cell cycle analysis. Binding of mac25 protein to activin A was studied by immunoprecipitation and Western blots analysis. RESULTS: mac25 protein was localized in the cytoplasm and secreted into culture medium. Addition of recombinant mac25 protein (10-7 M) into the culture medium induced significant suppression of the growth of human cervical carcinoma cells (HeLa) and murine embryonic carcinoma cells (P19), as well as osteosarcoma cells (Saos-2). mac25 protein was co-immunoprecipitated with activin A, a result that suggests that mac25 may be a secreted tumor-suppressor that binds activin A. CONCLUSION: mac25 exhibits homology to insulin-like growth factor-binding proteins (IGF-BPs) and to fibroblast growth factor receptor. The multi-functional nature of mac25 protein may be important for growth-suppression and/or cellular senescence. PMID- 10859030 TI - A manipulated dichotomy in global health policy. PMID- 10859031 TI - Search for prognostic markers for acute pancreatitis. PMID- 10859032 TI - Neuroprotection in acute ischaemic stroke: a tale of for whom the bell tolls? PMID- 10859033 TI - Heparins in management of acute coronary syndromes without ST-segment elevation. PMID- 10859034 TI - Variation in couple fecundity and time to pregnancy, an essential concept in human reproduction. PMID- 10859035 TI - Working off low back pain. PMID- 10859036 TI - Varicella vaccine in renal failure. PMID- 10859037 TI - Early use of non-invasive ventilation for acute exacerbations of chronic obstructive pulmonary disease on general respiratory wards: a multicentre randomised controlled trial. AB - BACKGROUND: Within the intensive-care unit, non-invasive ventilation (NIV) can prevent the need for intubation and the mortality associated with severe episodes of chronic obstructive pulmonary disease (COPD). The aim of this study was to find whether the introduction of NIV, early after the admission on a general respiratory ward, was effective at reducing the need for intubation and the mortality associated with acute exacerbations of COPD. METHODS: We did a prospective multicentre randomised controlled study comparing NIV with standard therapy in patients with mild to moderate acidosis. NIV was administered on the ward with a simple non-invasive ventilator and a standardised predefined protocol. Patients were recruited from 14 UK hospitals over 22 months. FINDINGS: 236 patients were recruited, 118 received standard therapy alone and 118 additional NIV. The two groups had similar characteristics at enrolment. The use of NIV significantly reduced the need for intubation as defined by the failure criteria. 32/118 (27%) of the standard group failed compared with 18/118 (15%) of the NIV group (p=0.02). In-hospital mortality was also reduced by NIV, 24/118 (20%) died in the standard group compared with 12/118 (10%) in the NIV group (p=0.05). In both groups pH, PaCO2, and respiratory rate improved at 4 h (p<0.01). However, NIV led to a more rapid improvement in pH in the first hour (p=0.02) and a greater fall in respiratory rate at 4 h (p=0.035). The duration of breathlessness was also reduced by NIV (p=0.025). INTERPRETATION: The early use of NIV for mildly and moderately acidotic patients with COPD in the general ward setting leads to more rapid improvement of physiological variables, a reduction in the need for invasive mechanical ventilation (with objective criteria), and a reduction in in-hospital mortality. PMID- 10859038 TI - Unfractionated heparin and low-molecular-weight heparin in acute coronary syndrome without ST elevation: a meta-analysis. AB - BACKGROUND: In acute coronary syndrome without ST elevation, the role of unfractionated and low-molecular-weight heparin in aspirin-treated patients remains unclear, and there is conflicting evidence regarding the efficacy and safety of low-molecular-weight heparin (LMWH) relative to unfractionated heparin. We did a systematic overview of the randomised trials to assess the effect of unfractionated heparin and LMWH on death, myocardial infarction, and major bleeding. METHODS: Randomised trials comparing unfractionated heparin or LMWH with placebo or untreated control, or comparing unfractionated heparin with LMWH, for the short-term and long-term management of patients with acute coronary syndrome without ST elevation, were identified by electronic and manual searches and through contact with experts and industry representatives. Odds ratios for death, myocardial infarction, and major bleeding were calculated for each trial, and results for the individual trials were combined by a modification of the Mantel-Haenszel method. FINDINGS: 12 trials, involving a total of 17157 patients, were included. The summary odds ratio (OR) for myocardial infarction or death during short-term (up to 7 days) unfractionated heparin or LMWH compared with placebo or untreated control was 0.53 (95% CI 0.38-0.73; p=0.0001) or 29 events prevented per 1000 patients treated; during short-term LMWH compared with unfractionated heparin was 0.88 (0.69-1.12; p=0.34); and during long-term LMWH (up to 3 months) compared with placebo or untreated control was 0.98 (0.81-1.17; p=0.80). Long-term LMWH was associated with a significantly increased risk of major bleeding (OR 2.26, [95% CI 1.63-3.14], p<0.0001), which is equivalent to 12 major bleeds per 1000 patients treated. INTERPRETATION: In aspirin-treated patients with acute coronary syndrome without ST elevation, short-term unfractionated heparin or LMWH halves the risk of myocardial infarction or death. There is no convincing difference in efficacy or safety between LMWH and unfractionated heparin. Long-term LMWH has not been proven to confer benefit additional to aspirin and there is no evidence to support its use after the first 7 days. PMID- 10859039 TI - Review of regional measles surveillance data in the Americas, 1996-99. AB - BACKGROUND: In 1994, ministers of health of countries of North and South America established the goal of measles eradication from the western hemisphere by 2000. To accomplish this goal, the Pan American Health Organization (PAHO) developed an enhanced measles vaccination strategy. METHODS: PAHO's measles eradication vaccination strategy has evolved into three principal components; a catch-up measles vaccination campaign, maintenance of high vaccination coverage (keep-up), and periodic follow-up measles vaccination campaigns. To monitor progress towards measles eradication, measles surveillance has been strengthened, including the laboratory investigation of suspected measles cases. FINDINGS: Both the catch-up and follow-up mass campaigns achieved high vaccination coverages in the respective targeted age groups. In 1996, only 2109 confirmed measles cases were reported in the Americas. In 1997, there was a resurgence of measles in the Americas, mostly as a result of a large measles outbreak with over 42000 cases, which occurred mainly among unvaccinated young adults in Sao Paulo State, Brazil. By 1998, there was a reduction in the number of reported confirmed measles cases, with a total of 14474 cases. Reduction of cases continued to the end of 1999, with a total of only 2828 confirmed cases. INTERPRETATION: PAHO's measles eradication strategy has been effective in interrupting transmission and maintaining the absence of measles virus circulation in most parts of the Americas. The PAHO experience provides strong evidence that with full implementation of an appropriate vaccination strategy, measles transmission can be effectively interrupted. PMID- 10859040 TI - Glycine antagonist (gavestinel) in neuroprotection (GAIN International) in patients with acute stroke: a randomised controlled trial. GAIN International Investigators. AB - BACKGROUND: Early treatment may improve acute ischaemic stroke outcome. Gavestinel is a selective antagonist at the glycine site of the N-methyl-D aspartate (NMDA) receptor, and is neuroprotective in animal models of ischaemic stroke. METHODS: We did a randomised, double-blind, placebo-controlled trial to test whether gavestinel could improve functional outcome after acute stroke in human beings. Conscious patients with stroke involving limb weakness received either gavestinel at an intravenous loading dose of 800 mg followed by 200 mg every 12 h for five doses, or matching placebo, within 6 h of stroke onset. Stratification variables were age and stroke severity. A computed tomography brain scan within 18 h of stroke onset identified the primary efficacy population with ischaemic stroke. Outcome was assessed by an independent observer with the Barthel index at 3 months. Three outcome categories were applied: good (Barthel index 95-100), moderate (60-90), and poor (0-55 or dead). Analysis was by intention to treat. FINDINGS: Of 1804 patients randomised, 16 received no treatment, and 333 had primary intracranial haemorrhage. 891 patients received gavestinel and 897 received placebo. Outcome in 721 patients who received gavestinel and were analysed for the primary endpoint at 3 months was good in 246 (34.1%), moderate in 136 (18.8%), and poor in 339 (47.0%), compared with 256 (34.9%), 133 (18.1%), and 345 (47.0%), respectively, of 734 patients who received placebo (p=0.8). Mortality at 3 months was 147 (20.4%) in the gavestinel group and 138 (18.8%) in the placebo group. Outcomes within preplanned subgroup and secondary analyses were also neutral. There were no significant differences in serious side-effects between the groups. INTERPRETATION: Treatment with gavestinel within 6 h of acute ischaemic stroke did not improve outcome. PMID- 10859041 TI - Early prediction of severity in acute pancreatitis by urinary trypsinogen activation peptide: a multicentre study. AB - BACKGROUND: There is a pressing clinical requirement for an early simple test of severity in acute pancreatitis. We investigated the use of an assay of trypsinogen activation peptide (TAP). METHODS: We undertook a multicentre study in 246 patients (172 with acute pancreatitis [35 with severe disease], 74 controls). We assessed the predictive value of urinary TAP concentrations measured by a validated competitive immunoassay. We compared the results with those for plasma C-reactive protein and three clinicobiochemical scoring systems. TAP and C-reactive protein concentrations were analysed at set times after symptom onset and compared with the clinicobiochemical systems scores at key times during hospital stay. FINDINGS: At 24 h after symptom onset, the median urinary TAP concentration was 37 nmol/L (IQR 17-110) for severe and 15 nmol/L (5 35) for mild disease (p<0.001). The respective values for plasma C-reactive protein were 24 mg/L (3-34) and 25 mg/L (6-75; p=0.208). The sensitivity, specificity, positive predictive, and negative predictive values of the test to show severe acute pancreatitis compared with mild acute pancreatitis at 24 h were: for TAP (>35 nmol/L), 58%, 73%, 39%, and 86%, respectively, and for C reactive protein (>150 mg/L), 0%, 90%, 0%, and 75%. 48 h after admission the values for the clinicobiochemical scoring systems were: APACHE II (> or =8), 56%, 64%, 30%, and 85%; Ranson score (> or =3), 89%, 64%, 38%, and 96%; and Glasgow score (> or =3), 77%, 75%, 44%, and 93%. At 48 h, the values for C-reactive protein were 86%, 61%, 37%, and 94% and for TAP were 83%, 72%, 44%, and 94%. Combined testing of C-reactive protein and TAP was not superior to TAP alone for accuracy. INTERPRETATION: Urinary TAP provided accurate severity prediction 24 h after onset of symptoms. This single marker of severity in acute pancreatitis deserves routine clinical application. PMID- 10859042 TI - Time trends in biological fertility in Britain. AB - BACKGROUND: There is evidence of a decline in semen quality in some countries, including Britain, in recent decades. This retrospective cohort study examined the hypothesis that biological fertility had also declined. The trend in couple fertility was assessed by means of time to pregnancy (TTP--a sensitive and validated measure of fertility. METHODS: A representative sample of the British population aged 16-59 years was surveyed. TTP was obtained for all births conceived after unprotected intercourse that began during 1961-93, excluding contraceptive failures. The sample size was 1540. FINDINGS: In contrast to the original hypothesis, this study found that fertility has increased; the rising trend was accompanied by slight dips during 1976-80 and 1986-90. These results were consistent between male and female respondents, and undiminished by adjustment for possible confounding factors. A stronger and more consistent relation was found with the year when unprotected intercourse started (a period effect) than with the year of birth of either partner (a birth cohort effect). INTERPRETATION: The findings could not be explained by trends in age at first birth, increased treatment of subfertility, or changes in oral contraceptive use. If a decline in male fertility has occurred, it has been more than compensated for by a countervailing increase in couple fertility. PMID- 10859043 TI - Atypical angina. PMID- 10859044 TI - Osteoporosis and osteopenia in men with eating disorders. AB - The occurrence of eating disorders and related deficiencies in bone mineral density are well established in women. However, we provide evidence that eating disorders are as common in men as in women, and are perhaps more severe. PMID- 10859045 TI - Hospital admission and mortality rates for venous thromboembolism in Oxford region, UK, 1975-98. AB - Multiple-cause-coded mortality rates for thromboembolism in the Oxford region of the UK showed a steady decline fom 1975 to 1998. The trend for admission rates was less clear-cut, but recent changes were not specific to women of ages associated with use of oral contraceptives or hormone-replacement therapy. PMID- 10859046 TI - Association between brain size and abstinence from alcohol. AB - Brain shrinkage with chronic alcoholism is well acknowledged. We have shown, with quantitative analysis of serial scans, an increase in hippocampal, cerebral, and cerebellar volume after abstinence from alcohol. PMID- 10859047 TI - Factor XIII-mediated inhibition of fibrinolysis and venous leg ulcers. AB - Densitometric analysis shows an accelerated healing rate and a significantly diminished lesional fibrinolytic activity in patients with venous leg ulcers treated topically with the fibrin-stabilising factor XIII compared with controls. PMID- 10859048 TI - Effect of pregnancy on exposure to malaria mosquitoes. AB - Pregnant women attracted twice the number of Anopheles gambiae complex--the predominant African malaria-carrying mosquito--than did their non-pregnant counterparts. We postulate that physiological and behavioural changes that occur during pregnancy are responsible for increased attractiveness, which could be important in intervention strategies aimed at protecting this high-risk group against malaria. PMID- 10859049 TI - Laser thrombolysis: safe and rapid removal of clots? PMID- 10859050 TI - Stricter regulation proposed for US gene-therapy trials. PMID- 10859051 TI - WHO staff members express concerns over internal restructuring. PMID- 10859052 TI - Kazakhstan's medical community considers new strategies for its workforce. PMID- 10859053 TI - "False patients" assess health standards in Spain. PMID- 10859054 TI - Control of sexually transmitted diseases for HIV-1 prevention: understanding the implications of the Mwanza and Rakai trials. AB - Two randomised controlled trials of sexually transmitted disease (STD) treatment for the prevention of HIV-1 Infection, in Mwanza, Tanzania, and Rakai, Uganda, unexpectedly produced contrasting results. A decrease in population HIV-1 incidence was associated with improved STD case management in Mwanza, but was not associated with STD mass treatment in Rakai. Some reductions in curable STDs were seen in both studies. These trials tested different interventions in different HIV-1 epidemic settings and used different evaluation methods; the divergent results may be complementary rather than contradictory. Possible explanations include: differences in stage of the HIV-1 epidemic, which can influence exposure to HIV-1 and the distribution of viral load in the infected population; potential differences in the prevalence of Incurable STDs (such as genital herpes); perhaps greater Importance of symptomatic than symptomless STDs for HIV-1 transmission; and possibly greater effectiveness of continuously available services than of intermittent mass treatment to control rapid STD reinfection. Implications of the trials for policy and future research agenda are discussed. PMID- 10859055 TI - The illness and death of Eva Peron: cancer, politics, and secrecy. PMID- 10859056 TI - Exertional heat illness. PMID- 10859057 TI - Exertional heat illness. PMID- 10859058 TI - Exertional heat illness. PMID- 10859059 TI - Exertional heat illness. PMID- 10859060 TI - Exertional heat illness. PMID- 10859061 TI - Endomyocardial fibrosis syndrome in Uganda. PMID- 10859062 TI - How NG-nitro-L-arginine methyl ester may alter airway responsiveness. PMID- 10859063 TI - Assessing goitre prevalence. PMID- 10859064 TI - Assessing goitre prevalence. PMID- 10859065 TI - Critical-care beds: the numbers. PMID- 10859066 TI - Helicobacter pylori among siblings. PMID- 10859067 TI - Helicobacter pylori among siblings. PMID- 10859068 TI - A check on limiting multiple births. PMID- 10859069 TI - Prudence and isolation. PMID- 10859070 TI - Internet mirror sites. PMID- 10859071 TI - The Nobel chronicles. 1992: Edmond H Fischer (b 1920) and Edwin G Krebs (b 1918). PMID- 10859072 TI - Safety and efficacy of dynamic graciloplasty for fecal incontinence: report of a prospective, multicenter trial. Dynamic Graciloplasty Therapy Study Group. AB - PURPOSE: Dynamic graciloplasty has been used for intractable fecal incontinence, and good results have been reported. The aim of this study was to assess prospectively the safety and efficacy of dynamic graciloplasty for intractable fecal incontinence in a prospective, multicenter trial. METHODS: A total of 123 adults were treated with dynamic graciloplasty at 20 institutions. Continence was assessed preoperatively and postoperatively by use of 14-day diaries. RESULTS: There was one treatment-related death. One hundred eighty-nine adverse events occurred in 91 patients (74 percent). Forty-nine patients (40 percent) required one or more operations to treat complications. One hundred seventy (90 percent) events were resolved. Sixty-three percent of patients without pre-existing stomas recorded a 50 percent or greater decrease in incontinent events 12 months after dynamic graciloplasty, and an additional 11 percent experienced lesser degrees of improvement. Twenty-six percent were not improved, worsened, or exited. In patients with pre-existing stomas, 33 percent achieved successful outcomes at 12 months. This number increased to 60 percent at 18 months. Seventy-eight percent of patients had increased enema retention time, and mean anal canal pressures improved significantly at 12 months. Significant changes in quality of life were also observed. CONCLUSIONS: Objective improvement can be demonstrated in the majority of patients with end-stage fecal incontinence treated with dynamic graciloplasty. Reduction in incontinence episodes can be correlated with improved quality of life. Adverse events are frequently encountered, but most resolve with treatment. PMID- 10859073 TI - Functional and quality-of-life outcomes in patients with rectal cancer after combined modality therapy, intraoperative radiation therapy, and sphincter preservation. AB - PURPOSE: Locally advanced primary and recurrent rectal cancers treated with external beam radiation therapy, intraoperative radiation therapy, and chemotherapy represent a complex group of patients in the setting of extensive pelvic surgery and sphincter preservation. We sought to define functional outcome and quality of life in this subset of patients. METHODS: We retrospectively reviewed our experience with locally advanced primary and recurrent rectal cancer patients who underwent intraoperative radiation therapy with either low anterior resection (n = 12) or coloanal anastomosis (n = 6) between 1991 and 1998. Current functional outcome and quality of life were evaluated by a detailed questionnaire. RESULTS: Median time from operation to assessment was 24 (range, 6 93) months. Using a standardized Sphincter Function Scale, incorporating the number of bowel movements per day and degree of incontinence, patients were graded as poor, fair, good, or excellent function. Of all patients, 56 percent reported unfavorable (poor or fair) function. Of the subset of patients with coloanal anastomosis or very low low anterior resection, 88 percent had unfavorable function as compared with 30 percent with standard low anterior resection. (P = 0.02; Fisher's exact probability test). A quality-of-life satisfaction score based on social, professional, and recreational restrictions demonstrated 56 percent of patients to be dissatisfied with their bowel function. CONCLUSIONS: The majority of patients with advanced rectal cancers who require external beam radiation therapy, extensive pelvic surgery, and intraoperative radiation therapy report unfavorable functional and quality-of-life outcomes after sphincter preservation. In this setting patients being considered for coloanal anastomosis or very low anterior resection may be better served by permanent diversion. PMID- 10859074 TI - Usefulness of FDG-PET scan in the assessment of suspected metastatic or recurrent adenocarcinoma of the colon and rectum. AB - PURPOSE: The purpose of this study was to evaluate the clinical efficacy of positron emission tomography with 2-[18F] fluoro-2-deoxy-D-glucose compared with computed tomography plus other conventional diagnostic studies in patients suspected of having metastatic or recurrent colorectal adenocarcinoma. METHODS: The records of 105 patients who underwent 101 computed tomography and 109 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography scans for suspected metastatic or recurrent colorectal adenocarcinoma were reviewed. Clinical correlation was confirmed at time of operation, histopathologically, or by clinical course. RESULTS: The overall sensitivity and specificity of 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography in detection of clinically relevant tumor were higher (87 and 68 percent) than for computed tomography plus other conventional diagnostic studies (66 and 59 percent). The sensitivity of 2 [18F] fluoro-2-deoxy-D-glucose positron emission tomography in detecting mucinous cancer was lower (58 percent; n = 16) than for nonmucinous cancer (92 percent; n = 93). The sensitivity of 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography in detecting locoregional recurrence (n = 70) was higher than for computed tomography plus colonoscopy (90 vs. 71 percent, respectively). The sensitivity of 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography in detecting hepatic metastasis (n = 101) was higher than for computed tomography (89 vs. 71 percent). The sensitivity of 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography in detecting extrahepatic metastases exclusive of locoregional recurrence (n = 101) was higher than for computed tomography plus other conventional diagnostic studies (94 vs. 67 percent). 2-[18F] fluoro-2-deoxy D-glucose positron emission tomography altered clinical management in a beneficial manner in 26 percent of cases (26/101) when compared with evaluation by computed tomography plus other conventional diagnostic studies. CONCLUSION: 2 [18F] fluoro-2-deoxy-D-glucose positron emission tomography is more sensitive than computed tomography for the detection of metastatic or recurrent colorectal cancer and may improve clinical management in one-quarter of cases. However, 2 [18F] fluoro-2-deoxy-D-glucose positron emission tomography is not as sensitive in detecting mucinous adenocarcinoma, possibly because of the relative hypocellularity of these tumors. PMID- 10859075 TI - Re-establish pneumoperitoneum in laparoscopic-assisted sigmoid resection? Randomized trial. AB - PURPOSE: Operating room time and anastomosis-related morbidity of laparoscopic assisted sigmoid resection with anastomosis performed in an open fashion through a horizontal suprapubic incision or laparoscopically after re-establishing pneumoperitoneum were compared. METHODS: A randomized trial was performed on patients with recurrent uncomplicated diverticulitis of the sigmoid colon during a 14-month period. Inclusion criteria were persistence of symptoms despite medical treatment and two previous admissions. Exclusion criteria included complicated diverticulitis, suspected cancer, and previous extensive abdominal surgery. Because skin incisions were similar and patients were randomly assigned in the operating room, the trial was performed as double blind. RESULTS: There were no deaths. Two patients were excluded before randomization. Three patients were not treated as allocated because of conversion to open surgery. Aside from previous abdominal-surgery rates, 16 patients with laparoscopic-assisted sigmoid resections after re-establishing pneumoperitoneum and 15 patients with laparoscopic-assisted sigmoid resections with anastomosis performed in an open fashion through a horizontal suprapubic incision were well-matched for age, gender, weight, American Society of Anesthesiology class, previous admissions, skin-incision length, size of circular stapler, and mobilization of splenic flexure. There were no significant differences in morbidity rates (3/16 vs. 3/15), complete doughnuts (16/16 vs. 15/15), blood loss (300 vs. 200 ml), flatus (4 vs. 4 days), solid-food resumption (5 vs. 6 days), stay (8.5 vs. 9 days) in laparoscopic-assisted sigmoid resection after re-establishing pneumoperitoneum and laparoscopic-assisted sigmoid resection with anastomosis performed in an open fashion through a horizontal suprapubic incision groups, respectively. Patients with laparoscopic-assisted sigmoid resection after re-establishing pneumoperitoneum had statistically longer operating room time (295 vs. 190 minutes; P < 0.01). Median follow-up was 12 and 10 months in 10 patients with laparoscopic-assisted sigmoid resection after re-establishing pneumoperitoneum and 11 patients with laparoscopic-assisted sigmoid resection with anastomosis performed in an open fashion through a horizontal suprapubic incision, respectively. One patient with laparoscopic-assisted sigmoid resection with anastomosis performed in an open fashion through a horizontal suprapubic incision had an anastomotic stenosis endoscopically dilated. CONCLUSIONS: Nonrestoration of pneumoperitoneum after laparoscopic-assisted sigmoid resection allows a decrease in operating room time and a similar outcome. PMID- 10859076 TI - Pathologic significance of tumor progression in locally recurrent rectal cancer: different nature from primary cancer. AB - PURPOSE: It has recently been demonstrated that the tumor growth rate is a stronger determinant of survival than the extent of the growth in local recurrence of rectal cancer. We studied which factors controlled the tumor growth rate using modern immunohistochemical methods. METHODS: In 51 patients who underwent extended resection for this condition, paraffin-embedded specimens were examined for 1) tumor angiogenesis by CD31 staining and microvessel counting, 2) apoptosis by terminal deoxynucleotide transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling staining, and 3) cellular proliferative activity using anti-proliferative cell nuclear antigen antibody. The results were compared with carcinoembryonic antigen doubling time and survival. RESULTS: The five-year survival rate was 20 percent. The postoperative carcinoembryonic antigen doubling time, which was the strongest predictor of survival, correlated highly with proliferative cell nuclear antigen labeling index, but did not correlate with the apoptotic index or microvessel counts. CONCLUSION: Our study shows that cancer cell proliferation rather than apoptosis or angiogenesis is a major determinant of tumor growth rate and survival in patients with locally recurrent rectal cancer. PMID- 10859077 TI - Flat adenoma of the large bowel: re-evaluation with special reference to central depression. AB - PURPOSE: The aim of this study was to re-evaluate the clinicopathologic features of flat adenomas with special reference to the role of central depression found in flat adenomas. METHODS: Clinicopathologic features, such as grade of atypia by size, site, central depression, coexisting lesions, gender, and family history of cancer, were evaluated in 236 flat adenomas from 183 patients selected retrospectively and prospectively. RESULTS: Of the 236 flat adenomas, 175 had mild, 33 had moderate, and 28 had severe (i.e., intramucosal carcinoma) atypia. The frequency of severe atypia correlated positively with size. Severe atypia were significantly more frequent in females (21 percent in females and 10 percent in males; P < 0.05). Sixteen percent of flat adenomas in patients with coexisting cancer showed severe atypia, which was significantly more than those without coexisting cancer (P < 0.05). The frequency of multiple flat adenomas in patients with two or more family members with cancer was 53 percent, which was significantly higher than in patients with one or no family members with cancer (P < 0.05). Central depression was observed in 19 percent of flat adenomas. The rate of severe atypia (22 percent) of flat adenomas with central depression was significantly higher than that (9 percent) of flat adenoma without central depression (P < 0.05). Patients who had flat adenomas with central depression had a higher incidence of coexisting cancer and cancer in the family (P < 0.05 and P < 0.01). CONCLUSION: At colonoscopic examination consideration should be given for the increased potential to become malignant in flat adenomas, especially those with central depression and in patients with a history of malignancy or with a family history of malignancy. PMID- 10859078 TI - Biofeedback for fecal incontinence using transanal ultrasonography: novel approach. AB - PURPOSE: Neosphincter procedures may prove to be the treatment of choice for patients with neuropathic fecal incontinence but are rarely proposed for milder forms of the disease. Biofeedback may prove beneficial to these patients but is yet unproven. The objectives of this study were to develop a method of performing biofeedback using transanal ultrasound to teach the patient to contract repetitively and to determine biologic measures of sphincter function using transanal ultrasound in healthy and incontinent patients. METHODS: Initial uncontrolled studies were performed to determine the compliance, normal values, biologic measures of external sphincter strength (isotonic and isometric fatigue times), and early efficacy data using continence scores and visual analog scale scores. RESULTS: Forty-four patients were assessed during three months, with relative improvements in continence scores (St. Mark's Hospital, 40 percent; Pescatori, 20 percent) and patient and investigator visual analog scale scores (38 percent for both) and measurable increase in biologic fatigue times measured by transanal ultrasound. CONCLUSIONS: Transanal ultrasound seems to be a method of teaching external sphincter contraction and measuring sphincter strength with good initial compliance. Clinically and statistically significant improvements in incontinence scores, visual analog scale scores, and biologic strength of the external sphincter were detected in the short-term follow-up with uncontrolled data. The randomized, controlled trial that we have begun will either confirm or refute these results. PMID- 10859079 TI - Ambulatory manometry in patients with colonic J-pouch and straight coloanal anastomoses: randomized, controlled trial. AB - PURPOSE: Bowel function after ultralow anterior resection may be improved by a colonic J-pouch. The aim of this study was to compare the bowel function and ambulatory manometry in patients randomly assigned to straight coloanal anastomosis or colonic J-pouch. METHODS: Forty-seven consecutive patients underwent ultralow anterior resection for adenocarcinoma. The colonic J-pouch was constructed with 6-cm limbs. A bowel function questionnaire was administered at one year after surgery. Ambulatory manometry was performed before and at one year after surgery. RESULTS: Values are expressed below as mean and (standard error of the mean). Patients with colonic J-pouch were found to have less frequent stools (4.6 (0.3) vs. 7.1 (0.9) stools/day; P < 0.05) and stool clustering (35 vs. 63.2 percent; P < 0.05) and were less unlikely to soil when passing flatus (85 vs. 35.3 percent; P < 0.05). The ambulatory anorectal pressure gradient was better preserved in the colonic J-pouch group (30.3 (3.7) vs. 18 (2.6) mmHg; P < 0.05). Stool frequency was predicted by the mean rectal pressures (t = 3.368; P = 0.003). However, higher mean rectal pressures were tolerated by the colonic J pouch for each daily bowel movement (6.7 (0.6) vs. 4.4 (0.5) mmHg/stool; P = 0.008). Anal sampling episodes and slow wave activity were impaired postoperatively in both groups. The minimal anal pressures were lower in patients unable pass flatus without soiling (12.4 (5.3) vs. 26 (2.3) mmHg; P = 0.004). Large contraction waves were not seen, and this may be related to the absence of severe defecation problems with 6-cm colonic J-pouches. CONCLUSIONS: A colonic J pouch resulted in better bowel function and more favorable ambulatory manometric findings at one year of follow-up. PMID- 10859080 TI - Alpha-1 adrenoceptor blockade: potential new treatment for anal fissures. AB - PURPOSE: Patients with chronic anal fissures are known to have high resting anal pressures that return to normal after successful surgical treatment. Internal anal sphincter activity is increased by sympathetic excitatory innervation via alpha adrenoceptors. The objective of this study was to determine the effect of alpha-1 adrenoceptor blockade on anal sphincter pressure in patients with and without chronic anal fissures. METHODS: The effect on the anal canal pressure profile of a single oral 20 mg dose of indoramin, an alpha-1 adrenoceptor antagonist, on seven patients with chronic anal fissure and six healthy patients without a fissure was investigated. RESULTS: Indoramin reduced anal resting pressures in those with anal fissure by a mean of 35.8 percent, from 106.9 +/- 22.15 cm H2O to 68.6 +/- 20.35 cm H2O, and in those without anal fissure by a mean of 39.9 percent, from 84.17 +/- 27.46 cm H2O to 52.17 +/- 24.78 cm H2O, after one hour. This pressure reduction persisted at three hours, and its magnitude is comparable to that obtained after internal sphincterotomy. The pressure reduction occurred over the whole length of the anal canal. CONCLUSION: It is proposed that alpha-1 adrenoceptor antagonists could be a suitable treatment for chronic anal fissure and other painful conditions where reduction in anal pressure is warranted. PMID- 10859081 TI - Ketorolac improves recovery after outpatient anorectal surgery. AB - PURPOSE: The purpose of this study was to evaluate the effectiveness of ketorolac combined with local anesthetics for anorectal surgery. METHODS: From June 1998 through March 1999, 123 outpatients undergoing anorectal surgery were entered into a prospective, randomized, double-blinded study involving three treatment groups. All patients received intravenous sedation consisting of fentanyl and a propofol infusion, with a local anesthesia mixture of lidocaine, bupivacaine, and bicarbonate. Group A (41 patients) received placebo (saline) injections. Group B (41 patients) received 60 mg of intravenous ketorolac at the onset of the procedure, and Group C (41 patients) received 60 mg of ketorolac mixed with the local anesthetic. Data were analyzed using analysis of variance and chi-squared tests. RESULTS: All groups had similar demographic characteristics and operative procedures. Twenty-nine of the 123 patients were human immunodeficiency virus positive. There was no difference in operative or anesthesia time. Anesthesia and fluids given were similar in across groups. A significantly higher percentage of Group A patients had pain (34 percent) and required additional oral analgesia (20 percent) in the Day Surgery Unit. Only 5 percent of Group B and Group C patients complained of pain, with oral analgesics given to 2 percent of Group B and none in Group C. Voiding difficulties were more common in Group A patients, one patient requiring catheterization. CONCLUSION: The addition of ketorolac (60 mg), either intravenous or injected with local anesthetics, reduces voiding problems and significantly decreases postoperative analgesic requirements in outpatients undergoing anorectal surgery. PMID- 10859082 TI - Hemorrhoidectomy with posterior perineal block: experience with 400 cases. AB - PURPOSE: The aim of this study was to evaluate the advantages and feasibility of hemorrhoidectomy using regional anesthesia (posterior perineal block). METHODS: From March 1994 to December 1998 we performed 400 hemorrhoidectomies with regional anesthesia in an overnight-stay regimen in our department (Colo-Rectal Unit). Posterior perineal block involves anesthesia of the deep plains (infiltration of the inferior hemorrhoidal nerves, the posterior branch of the internal pudendal nerves, and the anococcygeal nerves) and anesthesia of the superficial plains (block of the inferior gluteal nerves and of perineal branches of minor nerves from the sacral plexus). RESULTS: Posterior perineal block was always effective; optimal to satisfactory intraoperative analgesia was obtained in 379 patients (95.2 percent), whereas in 17 cases (4.2 percent) intravenous analgesic drugs were administered. No conversion to general anesthesia was needed. Urinary retention was 7.8 percent. In our study most of patients (70 percent) reported no pain at all for five to ten hours. Ninety-two percent of patients were discharged in the first 24 hours. CONCLUSIONS: Posterior perineal block allows the surgeon to perform radical hemorrhoidectomies in an overnight stay regimen with safe and effective intraoperative and postoperative analgesia, sphincter relaxation, and low incidence of urinary retention. Experience of the surgeon combined with careful surgical handling are of great importance for success in this technique. PMID- 10859083 TI - Results of sphincteroplasty in 86 patients with anal incontinence. AB - PURPOSE: This study was designed to analyze critically the short-term and long term outcome of sphincteroplasty and to identify high-risk factors. METHODS: Eighty-six patients with fecal incontinence associated with an ultrasound defect of the external anal sphincter were treated by anal sphincteroplasty. Clinical and physiologic assessment was made before surgery, and clinical evaluation was made three months and an average of 40 months after surgery. RESULTS: The evaluation of 86 patients three months after surgery showed that 42 patients were totally continent (49 percent), 28 were incontinent for gas (33 percent), and 16 still had fecal incontinence (19 percent). Seventy-four patients (86 percent) were contacted 40 months after surgery. Twenty-one patients (28 percent) were totally continent, 17 were incontinent to gas (23 percent), and 36 were incontinent to feces (49 percent). Forty-six percent of patients felt they were clearly improved after surgery. Poor results were associated with an internal anal sphincter defect. CONCLUSIONS: Our study suggests that in the long term, one third of patients are totally continent after sphincteroplasty. One-half of patients are satisfied, but only if their incontinence to feces has totally disappeared. Results of sphincteroplasty deteriorate with time. One factor in poor prognosis is the presence of an associated defect of the internal anal sphincter. PMID- 10859084 TI - Nitric oxide production is diminished in colonic circular muscle from acquired megacolon. AB - PURPOSE: Nitric oxide modulates human colonic smooth muscle function. To determine whether nitric oxide production is altered in colon from acquired megacolon, we measured cholinergic nerve-mediated contractions in vitro before and after inhibition of nitric oxide synthase. METHODS: Intramural nerves in circular smooth muscle from histologically normal colon (n = 12) and acquired megacolon (n = 3) were activated by electrical field stimulation. RESULTS: In controls blockade of nitric oxide synthase by N(G)-Nitro-L-Arginine induced increases (P < 0.05) in amplitude of contractions; these increases in amplitudes were blocked by L-Arginine (analysis of variance; P < 0.05). By contrast, blockade of nitric oxide synthase did not increase amplitudes of contractions with circular smooth muscle from acquired megacolon. An immediate phasic contraction was blocked by atropine sulfate. CONCLUSIONS: The results support the concept that nitric oxide production modulates cholinergic nerve-mediated contractions in normal colonic circular muscle, whereas acquired megacolon is associated with altered release of this inhibitory neurochemical. Potential explanations include depletion of tissue L-Arginine, decreased capacity to recycle citrulline to arginine, or decreased release of vasoactive intestinal peptide from circular smooth muscle in acquired megacolon. PMID- 10859085 TI - Does better functional result equate with better quality of life? Implications for surgical treatment in familial adenomatous polyposis. AB - PURPOSE: The main impetus for a patient with familial adenomatous polyposis to choose colectomy with ileorectal anastomosis over ileal pouch-anal anastomosis is the better functional result. However, does better functional result necessarily translate into better overall quality of life? Previous studies of other diseases have demonstrated no such correlation. This study was performed to determine whether any relationship exists between functional result and quality of life in patients with familial adenomatous polyposis after ileorectal anastomosis and ileal pouch-anal anastomosis. METHODS: All patients with familial adenomatous polyposis who underwent colectomy with ileorectal anastomosis or proctocolectomy with ileal pouch-anal anastomosis from 1980 to 1998 were studied. Functional data were obtained by questionnaire. Health-related quality of life was assessed by two validated instruments, the SF-36 Physical and Mental Health Summary Scales and the SF-36 Health Survey, which measure physical and mental functioning and eight separate health-quality dimensions, including health perception, physical and social functioning, physical and emotional role limitations, mental health, bodily pain, and energy or fatigue. RESULTS: Data were obtained in 44 of 68 patients, 14 with ileorectal anastomosis and 30 with ileal pouch-anal anastomosis. No differences were demonstrated between the two groups for patient age, mean follow-up time, and mean patient age at operation. Functional results were worse for the ileal pouch-anal anastomosis group vs. the ileorectal anastomosis group in number of bowel movements per day (7.5 vs. 5.2; P < 0.05), leakage (43 vs. 0 percent; P < 0.01), pad usage (17 vs. 0 percent; P < 0.01), perianal skin problems (33 vs. 7 percent; P < 0.01), food avoidance (80 vs. 43 percent; P < 0.01), and inability to distinguish gas (37 vs. 7 percent; P < 0.01). Results of the health-related quality-of-life surveys, however, demonstrated no difference between the ileal pouch-anal anastomosis and ileorectal anastomosis groups. The Physical and Mental summary scales for the ileal pouch-anal anastomosis and ileorectal anastomosis groups were not significantly different (Physical Health Scale, 50.3 vs. 50.9; Mental Health Scale, 51.7 vs. 49.6), and none of the eight dimensions of the SF-36 health survey demonstrated statistical differences between the ileal pouch-anal anastomosis and ileorectal anastomosis groups. CONCLUSION: Better functional results were not equated with better quality of life in this pilot study. Although patients with the ileorectal anastomosis have better functional results than those with ileal pouch-anal anastomosis, the measured health-related quality of life as determined by a validated generic health-related quality-of-life instrument is the same for both groups. These results suggest that all patients with familial adenomatous polyposis might be optimally treated with an ileal pouch-anal anastomosis. More importantly, this study suggests that health-related quality of life should play a greater role in the evaluation of care and treatment in colon rectal surgery. PMID- 10859086 TI - What is the benefit of preoperative sperm preservation for patients who undergo restorative proctocolectomy for benign diseases? AB - PURPOSE: In patients with benign colorectal diseases undergoing a restorative proctocolectomy with an ileal pouch-anal anastomosis, semen cryopreservation seems rational to enable the possibility of procreation in case surgery leads to sexual disorders or impotence. The aim of this study was to determine the preoperative and postoperative semen quality in patients undergoing ileal pouch anal anastomosis. In addition, the study sought to determine the incidence of surgery-induced sexual dysfunction to evaluate the economic efficiency of semen cryopreservation as compared with alternatives such as microsurgical epididymal sperm aspiration. METHODS: Preoperative and postoperative semen analyses were offered to 97 patients with ileal pouch-anal anastomosis with benign colorectal diseases since 1989. The direct costs of the semen cryopreservation program were determined and compared with those of alternatives. RESULTS: In 34 of 40 consecutive patients with ileal pouch-anal anastomosis who made use of preoperative semen preservation, normal sperm concentrations, motility, and morphology were found. Mean semen characteristics of all 23 patients who returned for postoperative analysis were not different from preoperative values, but they were for total sperm number. Two patients developed temporary retrograde ejaculation postoperatively. None of the preserved semen samples was used, thus semen cryopreservation benefited none of these patients. The total costs of semen cryopreservation are between 2.2 and 5 times higher than the costs for one microsurgical epididymal sperm aspiration procedure. CONCLUSIONS: Preoperative semen cryopreservation in patients undergoing ileal pouch-anal anastomosis because of benign colorectal diseases is quite feasible. However, most likely because of improved surgical techniques and the increasing number of effective alternatives, preoperative semen cryopreservation in patients with ileal pouch anal anastomosis is no longer cost effective. PMID- 10859087 TI - Long-term results of low anterior resection with intersphincteric anastomosis in carcinoma of the lower one-third of the rectum: analysis of 31 patients. AB - INTRODUCTION: Between 1985 and 1996, 190 patients underwent a low anterior rectal resection with coloanal anastomosis for adenocarcinoma of the lower one-third of the rectum. METHODS: This article reports on 31 (17 males) of these patients with a very low localization of the tumor (distal tumor margin 1.3 +/- 0.9 cm above the dentate line). If the function of the sphincter was acceptable and we could exclude tumor infiltration into the sphincter through endosonography, we relocated the resection plane distally into the intersphincteric region to attain an acceptable margin of safety. In all of these cases, it was impossible for us to perform the usual surgical procedure of a mechanical anastomosis by means of a circular stapler. After intersphincteric rectal resection, the anastomosis was handsewn, using interrupted sutures from the perineal approach, 2.5 to 3 cm above the anal verge, implementing Parks' retractor. A protective stoma was performed in all cases. All data were documented prospectively. RESULTS: COMPLICATIONS: Postoperative mortality was 0 percent. Postoperatively, none of the patients showed an indication for relaparotomy. The leakage rate was 48 percent. Only 16 percent later needed additional surgery for anastomotic strictures or for rectovaginal fistulas. Long-term observations showed that the anastomosis healed well in 27 patients (87.1 percent). Four patients (12.9 percent) decided to have a terminal colostomy performed (anastomotic stricture, 3 patients; anorectal incontinence, 1 patient). FOLLOW-UP: During the follow-up period of 6.8 +/- 3.7 years, six patients (19.4 percent) developed a tumor progression (9.7 percent local recurrences and 12.9 percent distant spread). The five-year survival rate was 79 percent (Dukes A, 100 percent (n = 18); Dukes B, 67 percent (n = 4); and Dukes C, 44 percent (n = 9)). Continence: One-third of patients developed anorectal incontinence for liquid (29.6 percent) or solid stool (3.7 percent). Average stool frequency was 3.3 times per day. Resting pressure decreased significantly by 29 percent (preoperative, 105 +/- 37 cm H2O and postoperative, 75 +/- 19 cm H2O; P < 0.05), whereas squeeze pressure did not change. CONCLUSION: In selected patients with tumors close to the dentate line, an intersphincteric resection of the rectum may help to avoid an abdominoperineal excision of the rectum with a terminal stoma, without any curtailment of oncologic standards. A protective stoma for three months is advantageous. PMID- 10859088 TI - Idiopathic slow-transit constipation is not associated with mutations of the RET proto-oncogene or GDNF. AB - PURPOSE: Idiopathic slow-transit constipation is a severe disorder of unknown cause. The onset in early childhood and history of constipation or Hirschsprung's disease in close family relatives suggest that slow-transit constipation could have a genetic basis. Several germline mutations have been described in Hirschsprung's disease, including mutations of RET and the gene encoding its ligand glial cell-derived neurotrophic factor. The aim of this study was to screen a panel of 16 cases of familial idiopathic slow-transit constipation, including 4 families in which there were relatives with Hirschsprung's disease, for RET and glial cell-derived neurotrophic factor mutations previously identified in Hirschsprung's disease. METHODS: Genomic DNA from 16 patients with slow-transit constipation and four relatives with Hirschsprung's disease was analyzed using single strand and heteroduplex conformation polymorphism analysis at two conditions and by direct DNA sequencing using the fluorescent dideoxy terminator method. RESULTS: Although common sequence polymorphisms were demonstrated with a frequency comparable with published data, no published or new mutation was seen in any of the exons of RET or glial cell-derived neurotrophic factor. CONCLUSIONS: Mutation of RET or glial cell-derived neurotrophic factor is not a frequent cause of idiopathic slow-transit constipation. PMID- 10859089 TI - Enterochromaffin and serotonin cells are abnormal for patients with colonic inertia. AB - PURPOSE: In recent studies, serotonin and several gut peptides have been shown to serve as regulators of colonic transit. Thus, the distribution, density, and intensity of cells secreting serotonin or certain gut peptides could be abnormal in patients with colonic inertia. The aim of this study was to evaluate the distribution, density, and staining intensity of enterochromaffin and serotonin cells in the colonic mucosa of patients with colonic inertia compared with a control group. METHODS: Between 1993 and 1998 tissue blocks from the right and left side of the colon were obtained in 19 consecutive patients (18 females; mean age, 43.7 +/- 11.5 years) who underwent subtotal colectomy for colonic inertia. The control group consisted of colonoscopic biopsies from the right and left colon of 15 patients (all females; mean age, 52.7 +/- 16.5 years) for indications other then constipation, inflammatory bowel diseases, or carcinoma. Immunocytochemical staining of enterochromaffin and serotonin cells were performed on 4 microm tissue sections with the primary rabbit antibody against chromogranin A or serotonin, and the biotinylated secondary antibody and enzyme labeled-streptavidin. The average cell number per microscopic field (x200) was calculated and the proportion of cells with various staining distribution was expressed as the percentage of the entire positive cell population as low, moderate, and high intensity. Student's t-test and chi-squared test were used for statistical analysis, with significance level set at P < 0.05. RESULTS: The quantity of both enterochromaffin cells (16.8 +/- 10.2) and serotonin cells (12.1 +/- 6.4) in the mucosa of the left colon in patients with colonic inertia was significantly higher when compared with the right side of the colon (enterochromaffin cells, 9.4 +/- 6.0; serotonin cells, 7.8 +/- 3.6; P < 0.01). The percentage of both types of cells with low staining intensity was increased, whereas the cells with high and moderate staining intensity were decreased (P < 0.01) in the left colon as compared with the right. The number of enterochromaffin cells in left-sided colonic mucosa was significantly higher in the colonic inertia group than in the control group (16.8 +/- 10.1 vs. 10.4 +/- 6.0; P < 0.05). Moreover, the numbers of serotonin cells in both the right and left colon was also significantly higher in the colonic inertia group than in the control group (right, 7.8 +/- 3.6 vs. 4.1 +/- 2.4; left, 12.1 +/- 6.4 vs. 5.8 +/- 3.7; P < 0.01). In both sides of the colon, the percentage of enterochromaffin and serotonin cells with low staining was significantly higher, whereas percentage of those cells with high or moderate staining was significantly lower in the colonic inertia group than in the control group. In the colonic inertia group there was a significantly positive correlation between numbers of enterochromaffin and serotonin cells (right side, P < 0.01; left side, P < 0.05). CONCLUSION: In patients with colonic inertia, the number of both enterochromaffin and serotonin cells are significantly increased in the colonic mucosa, especially in the left colon. As indicated by staining distribution, enterochromaffin and serotonin cells contain significantly less hormone than do the same cells in the control group. PMID- 10859090 TI - Colonic adenocarcinoma occurring in an Indiana pouch: report of a case and review of the literature. AB - Colonic adenocarcinoma has been frequently reported after ureterosigmoidostomy based on carcinogenic substances created when feces and urine are mixed. However, colonic adenocarcinoma has never been reported arising in an Indiana pouch after cystectomy. We report a case of adenocarcinoma arising in a urinary pouch after cystectomy for transitional cell cancer. We believe this to be caused by hereditary nonpolyposis colon cancer (an autosomal dominant syndrome that puts individuals at risk for genitourinary, colonic, and several other cancers) rather than carcinogenic effects of urine on colonic mucosa. When planning urinary reconstruction after cystectomy for malignancy, it is important to consider the possibility that an individual may suffer from hereditary nonpolyposis colon cancer before selecting the colon as a urinary reservoir. PMID- 10859091 TI - Continuous intra-arterial 5-FU chemotherapy in a patient with a repeated recurrence of rectal cancer: report of a case. AB - PURPOSE: We report a patient with a recurrent pelvic tumor after abdominoperineal resection of a rectal carcinoma who was treated sufficiently by repeated intra arterial infusions of 5-fluorouracil. METHODS: A continuous, 24-hour 5 fluorouracil administration was made through the bilateral internal iliac artery at a dosage of 250 mg/m2/day by the subcutaneous reservoir located at both upper legs using a Baxter infusor. RESULTS: In this patient pain in the hip and pelvis was relieved. A complete regression in the infused field of pelvic tumor was observed not only with computed tomography and magnetic resonance imaging but also confirmed by operative findings at the seventh month after the intra arterial infusion. The abnormal serum level of carcinoembryonic antigen and carbohydrate antigen 19-9 was decreased to within the normal range at the 19th and 3rd week respectively. When the repeated recurrence was suspected in follow up, normalization of the re-elevated carcinoembryonic antigen and carbohydrate antigen 19-9 levels was also obtained by repeating the same treatment. The side effects and complications were tolerable, consisting of local skin erosion on the hips and lower extremity neuropathy caused by the 5-fluorouracil. CONCLUSIONS: Clinical local regression of a pelvic recurrence was observed in a patient with rectal recurrent tumor who received continuous intra-arterial chemotherapy. Local recurrence of rectal cancer may be controlled effectively and safely by repeating long-term, continuous, intra-arterial 5-fluorouracil infusion. PMID- 10859092 TI - Rare presentation of actinomycosis as an abdominal mass: report of a case. AB - PURPOSE: The purpose of this article was to report an unusual presentation of abdominal actinomycosis masquerading as a tumor. METHODS: The patient was a 54 year-old male who presented with vague abdominal discomfort and a palpable left lower quadrant mass defined on CT scan. Multiple intraoperative core biopsies were nondiagnostic, and he underwent en bloc resection of the mass and adjacent organs for a presumed tumor. RESULTS: Examination of tissue from deep within the excised specimen revealed sulfur granules diagnostic for actinomycosis. CONCLUSION: Abdominal actinomycosis is an extremely rare infection that can mimic multiple disease processes and requires accurate diagnosis for successful therapy. This novel presentation and a review of the literature are reported. PMID- 10859093 TI - Ileal pouch and trauma. PMID- 10859094 TI - Stapled rectal prolapsectomy. PMID- 10859095 TI - Treatment of ruptures of the lateral ankle ligaments: a meta-analysis. AB - BACKGROUND: Ruptures of the lateral ankle ligaments are very common; however, treatment remains controversial. The aim of the current study was to perform a meta-analysis of randomized, controlled clinical trials of existing treatment strategies for acute ruptures of the lateral ankle ligaments. METHODS: Randomized, controlled trials reported between 1966 and 1998 were included if they involved acute ruptures of the lateral ankle ligaments. Randomized, controlled trials are defined as comparative studies with an intervention group and a control group in which the assignment of participants to a group is determined by the formal procedure of randomization. Summary measures of effectiveness were expressed as relative risks with use of random effects modeling. RESULTS: When analyzing the trials, we searched for comparable outcome measures in both short and long-term follow-up studies (studies with six months to 3.8 years of follow-up). This resulted in the analyses of three outcome measures: time lost from work, residual pain, and giving-way. This report summarizes the results of twenty-seven trials. With respect to giving-way, a significant difference was noted between operative treatment and functional treatment (relative risk, 0.23; 95 percent confidence interval, 0.17 to 0.31) in favor of operative treatment and a significant difference was also noted between functional treatment and treatment with a cast for six weeks (relative risk, 0.69; 95 percent confidence interval, 0.50 to 0.94) in favor of functional treatment. With respect to residual pain, no significant difference was found between operative and functional treatment and a significant difference was found between functional treatment and treatment with a cast for six weeks (relative risk, 0.67; 95 percent confidence interval, 0.50 to 0.90). We found minimal or no treatment to result in more residual pain (relative risk, 0.53; 95 percent confidence interval, 0.27 to 1.02) and giving-way (relative risk, 0.34; 95 percent confidence interval, 0.17 to 0.71) than did functional treatment. CONCLUSIONS: We concluded that a no-treatment strategy for ruptures of the lateral ankle ligaments leads to more residual symptoms. Operative treatment leads to better results than functional treatment, and functional treatment leads to better results than cast immobilization for six weeks. PMID- 10859096 TI - Nonoperative treatment of ipsilateral fractures of the scapula and clavicle. AB - BACKGROUND: Internal fixation of one or both bones is the recommended treatment for floating shoulder injuries (ipsilateral fractures of the scapula and clavicle). Perceived risks of nonoperative treatment include abduction weakness, decreased range of motion, chronic pain, malunion, and nonunion. None of these problems, however, have been confirmed by clinical studies. The purpose of this retrospective study was to analyze the clinical and radiographic results of nonoperative treatment of floating shoulder injuries. METHODS: Twenty patients with a floating shoulder injury were treated with either a sling or a shoulder immobilizer. Eleven clavicular fractures were displaced ten millimeters or more, and five scapular fractures were displaced more than five millimeters. Physical therapy was begun three days to two weeks after the injury. Patients were evaluated with three separate scoring systems: those of Herscovici et al., Rowe, and Constant and Murley. Shoulder abduction and flexion were measured, and abduction strength was evaluated by clinical examination and comparison with the uninjured extremity. The duration of follow-up averaged twenty-eight months (range, nine to seventy-nine months). RESULTS: Nineteen of the twenty pairs of fractures united uneventfully. One clavicular nonunion occurred secondary to segmental bone loss from a gunshot wound. On the basis of the Herscovici rating system, seventeen patients had an excellent result and three had a good result. According to the Rowe system, eighteen patients had an excellent result, one had a good result, and one had a fair result. The average Rowe score was 95. The average Constant score was 96. In all twenty patients, the strength of the injured extremity was equal to that of the uninjured extremity. Eighteen patients had a full, symmetrical range of shoulder motion, one lost 15 degrees of flexion, and one lost 20 degrees of abduction. CONCLUSIONS: Nonoperative treatment of floating shoulder injuries, especially those with less than five millimeters of fracture displacement, can achieve satisfactory results that are probably equal or superior to those reported after operative treatment, without the risk of operative complications. PMID- 10859097 TI - Conversion of external fixation to intramedullary nailing for fractures of the shaft of the femur in multiply injured patients. AB - BACKGROUND: From 1989 to 1997, 1507 fractures of the shaft of the femur were treated with intramedullary nailing at The R Adams Cowley Shock Trauma Center. Fifty-nine (4 percent) of those fractures were treated with early external fixation followed by planned conversion to intramedullary nail fixation. This two stage stabilization protocol was selected for patients who were critically ill and poor candidates for an immediate intramedullary procedure or who required expedient femoral fixation followed by repair of an ipsilateral vascular injury. The purpose of the current investigation was to determine whether this protocol is an appropriate alternative for the management of fractures of the femur in patients who are poor candidates for immediate intramedullary nailing. METHODS: Fifty-four multiply injured patients with a total of fifty-nine fractures of the shaft of the femur treated with external fixation followed by planned conversion to intramedullary nail fixation were evaluated in a retrospective review to gather demographic, injury, management, and fracture-healing data for analysis. RESULTS: The average Injury Severity Score for the fifty-four patients was 29 (range, 13 to 43); the average Glasgow Coma Scale score was 11 (range, 3 to 15). Most patients (forty-four) had additional orthopaedic injuries (average, three; range, zero to eight), and associated injuries such as severe brain injury, solid organ rupture, chest trauma, and aortic tears were common. Forty fractures were closed, and nineteen fractures were open. According to the system of Gustilo and Anderson, three of the open fractures were type II, eight were type IIIA, and eight were type IIIC. Intramedullary nailing was delayed secondary to medical instability in forty-six patients and secondary to vascular injury in eight. All fractures of the shaft of the femur were stabilized with a unilateral external fixator within the first twenty-four hours after the injury; the average duration of the procedure was thirty minutes. The duration of external fixation averaged seven days (range, one to forty-nine days) before the fixation with the static interlocked intramedullary nail. Forty-nine of the nailing procedures were antegrade, and ten were retrograde. For fifty-five of the fifty-nine fractures, the external fixation was converted to intramedullary nail fixation in a one stage procedure. The other four fractures were associated with draining pin sites, and skeletal traction to allow pin-site healing was used for an average of ten days (range, eight to fifteen days) after fixator removal and before intramedullary nailing. Follow-up averaged twelve months (range, six to eighty seven months). Of the fifty-eight fractures available for follow-up until union, fifty-six (97 percent) healed within six months. There were three major complications: one patient died from a pulmonary embolism before union, one patient had a refractory infected nonunion, and one patient had a nonunion with nail failure, which was successfully treated with retrograde exchange nailing. The infection rate was 1.7 percent. Four other patients required a minor reoperation: two were managed with manipulation under anesthesia because of knee stiffness, and two underwent derotation and relocking of the nail because of rotational malalignment. The rate of unplanned reoperations was 11 percent. The average range of motion of the knee was 107 degrees (range, 60 to 140 degrees). CONCLUSIONS: We concluded that immediate external fixation followed by early closed intramedullary nailing is a safe treatment method for fractures of the shaft of the femur in selected multiply injured patients. PMID- 10859098 TI - Total hip arthroplasty with use of the Metasul metal-on-metal articulation. Four to seven-year results. AB - BACKGROUND: Total hip replacements with a metal-on-metal articulation were commonly used until the mid-1970s; most were then abandoned in favor of hip replacement with a metal-on-polyethylene articulation. The reason for this change was primarily early cup loosening, which was more prevalent with these metal-on metal designs than it was with metal-on-polyethylene designs. In the late 1980s, a metal-on-metal design with improved clearance (adequate space between the femoral head and the acetabular articulation surface to allow fluid film lubrication and clearance of any debris from within this joint), metal hardness, and reproducible surfaces was introduced by Sulzer Orthopedics in Switzerland. Orthopaedic surgeons were interested in this Metasul articulation because the contribution of polyethylene wear particles to the failure of total hip replacements had become evident. This study was undertaken to review the clinical performance of this implant and to determine if early acetabular loosening or revision and wear and osteolysis were prevalent. METHODS: Between 1991 and 1994, seventy patients (seventy hips) had a total hip replacement with the Metasul metal-on-metal articulation and a cemented Weber cup. Nine patients died less than four years after the replacement; none of these deaths were related to the operation. Five patients were not available for radiographic evaluation, but they were contacted and it was known that the hip was not painful and had not been revised. Fifty-six patients (fifty-six hips) had complete clinical and radiographic data four to 6.8 years after the operation, and they made up the study group. The patients were evaluated with use of the Harris hip score, a patient-self-assessment form, and radiographs. RESULTS: At an average of 5.2 years (range, four to 6.8 years) after the operation, the average total Harris hip score for the fifty-three patients who did not have a revision was 89.6 points (range, 62 to 100 points). The average Harris pain score was 41.0 points (range, 30 to 44 points), and the average Harris limp score was 9.4 points (range, 5 to 11 points). One patient had revision of a loose cup, but there were no other loose acetabular components in the series. Two patients had revision of the acetabular component because of dislocation. No patient had a loose or revised femoral component. Therefore, the mechanical failure rate was one (2 percent) of fifty-six patients. Thirty-six of forty-seven patients who completed the patient-self-assessment form rated their result as excellent; seven, as very good; two, as good; one, as fair; and one, as poor. Wear could not be measured on radiographs because of the metal-on-metal articulation. No hip had radiographic evidence of acetabular osteolysis and two hips had calcar resorption, but there was no other radiographic evidence of focal osteolysis. CONCLUSIONS: Our four to seven-year experience with this articulation surface indicates that the clinical results are similar to those of total hip replacements with a metal-on polyethylene articulation. We believe that the Metasul articulation may have a role in reducing the wear that occurs with total hip replacement. The Metasul articulation appears to be particularly indicated for more active patients. A historical comparison with the reports in the literature of which we are aware indicated that the hips in our study had a lower rate of acetabular revision and loosening than did those with previous metal-on-metal designs and that they had no more acetabular loosening or osteolysis than did those with metal-on polyethylene articulations followed for an average of five years. PMID- 10859099 TI - Distraction osteogenesis after acute limb-shortening for segmental tibial defects. Comparison of a monofocal and a bifocal technique in rabbits. AB - BACKGROUND: Segmental bone defects can be treated with immediate limb-shortening followed by monofocal or bifocal distraction osteogenesis. In the present study, the efficacy of monofocal distraction osteogenesis was compared with that of bifocal distraction osteogenesis in a rabbit model. METHODS: Twenty-four skeletally mature New Zealand White rabbits were divided into two equal groups: one group had monofocal distraction osteosynthesis, and the other had bifocal distraction osteosynthesis. In both groups, a one-centimeter-long segment of bone was resected from the midpart of the tibial shaft. In the monofocal reconstruction group, the limb was immediately shortened to close the segmental defect and the defect was allowed to heal for ten days. Lengthening was then begun at this site, with use of a specially designed external fixator, at a rate of 0.5 millimeter per twelve hours. In the bifocal reconstruction group, the segmental defect was closed immediately and the fragments were fixed with microplates. A subperiosteal osteotomy was performed proximal to the tibiofibular junction, and lengthening was performed at the site of the osteotomy. The animals in both groups were killed twenty days after the lengthening was completed. New bone formation then was evaluated with use of radiographs, densitometry, biomechanical testing, and histological and histomorphometric analysis. RESULTS: Osseous consolidation occurred in all but one of the animals. Biomechanical testing demonstrated that the tibiae that had been treated with use of the simple monofocal reconstruction technique tended to have greater torsional stiffness (p = 0.14) and strength (p = 0.09). Follow-up radiographs revealed that both groups had a significant decrease in radiolucent area (p < 0.05), which occurred at essentially the same rate after lengthening. No significant differences were found between the groups with respect to new-bone mineral density, new-bone area, or the amount of callus. Thus, after resection of a diaphyseal bone segment comprising 10 percent of the original length of the tibia and acute shortening, limb reconstruction was completed successfully through distraction osteogenesis with use of either a monofocal or a bifocal technique in rabbits. CONCLUSIONS: In the present study, both monofocal and bifocal techniques of shortening and distraction osteogenesis were effective for the reconstruction of segmental bone defects. Under some conditions, the monofocal method may provide a simpler means of treating such defects. CLINICAL RELEVANCE: Damage to the soft-tissue envelope as well as venous and lymphatic stasis impose limits on the amount of limb shortening that can be achieved with use of the monofocal method and also influence the indications for this procedure in the clinical setting. PMID- 10859100 TI - Anatomical considerations regarding the posterior interosseous nerve during posterolateral approaches to the proximal part of the radius. AB - BACKGROUND: The purpose of our study was to quantify the dimensions of a surgically safe zone along the proximal part of the radius, from the posterolateral aspect. METHODS: The posterolateral approach between the anconeus and the extensor carpi ulnaris was performed in thirty-two cadaveric specimens, and the posterior interosseous nerve was exposed. Forearms were measured from the radial styloid process to the radiocapitellar joint. The distance from the capitellum to the point where the posterior interosseous nerve crossed the radial shaft and the angle between the nerve and the shaft were measured with forearms in pronation and supination. RESULTS: Pronation of the forearm allowed safe exposure of at least the proximal thirty-eight millimeters of the lateral aspect of the radius, with an average proximal safe zone of 52.0 +/- 7.8 millimeters. Supination decreased this proximal safe zone to as little as twenty-two millimeters and an average of 33.4 +/- 5.7 millimeters. The angle formed by the posterior interosseous nerve and the radial shaft in supination averaged 47.4 +/- 6.8 degrees; this decreased to 27.8 +/- 6.7 degrees with pronation. CONCLUSIONS: Approaching the lateral aspect of the proximal part of the radius is safest in pronation. PMID- 10859101 TI - Type-B-IIIa hip rotationplasty: an alternative operation for the treatment of malignant tumors of the femur in early childhood. AB - BACKGROUND: The biological plasticity of the cartilaginous proximal part of the tibia in children makes it possible to use the tibia to reconstruct the lower extremity after excision of a sarcoma of the thigh. A type-B-IIIa rotationplasty is an alternative to prosthetic replacement in very young children who have a malignant tumor of the femur that requires extensive resection. METHODS: A type-B IIIa rotationplasty was done in eight patients who had a femoral tumor: four had a Ewing sarcoma; three, an osteosarcoma; and one, a primitive neuroectodermal tumor. The ages ranged from two years and eight months to ten years and six months at the time of the procedure. RESULTS: All eight patients were able to bear full weight and had a good range of motion of the hip joint at a median of five years and one month (range, two years and four months to eight years) postoperatively. They also were able to participate in sports activities. Radiographs and magnetic resonance imaging studies confirmed that the lateral part of the tibial plateau had remodeled to form a structure that resembled a developing femoral head. Seven patients were operated on only once, and a second hospital stay was not necessary. The remaining patient had a prolonged hospitalization for revision of the wound. CONCLUSIONS: As an alternative to amputation or an extendable tumor prosthesis, a type-B-IIIa rotationplasty offers not only a better functional result but also biological reconstruction. Placement of the cartilaginous head of the tibia into the acetabulum permits development of a new femoral head. Thus, not only is the foot preserved as a functional knee joint but a newly formed hip joint develops as well. PMID- 10859102 TI - The reliability and validity of the self-reported patient-specific index for total hip arthroplasty. AB - BACKGROUND: The Patient-Specific Index is unique in that it reflects how individual patients weigh concerns in rating the outcome of total hip arthroplasty. The Patient-Specific Index was originally administered by an interviewer, which is not always feasible and can be costly. The purposes of the present study were (1) to create a self-reported version of the Patient-Specific Index, (2) to determine the reliability of this new self-reported version, and (3) to determine the relationship between the scores on the new self-reported version and those on the original interviewer-administered version. METHODS: A self-reported version of the Patient-Specific Index was developed, and a pilot test was performed on ten patients. Patients who were scheduled for a total hip arthroplasty or who had recently had a total hip arthroplasty were eligible for the reliability and validity testing. A copy of the new self-reported Patient Specific Index was mailed to the patients, and they completed it independently. The patients' ratings of the importance and severity of twenty-four concerns prior to total hip arthroplasty were added together to create a summary Patient Specific Index score. To determine test-retest reliability, patients completed the self-reported Patient-Specific Index a second time, two weeks later. To determine criterion validity, participants also completed the interviewer administered Patient-Specific Index. RESULTS: Fifty-five patients completed the study. The random-effects intraclass correlation test-retest coefficient was 0.79 (greater than 0.75 represents excellent reliability). The mean Patient-Specific Index scores on the self-reported version and on the interviewer-administered version were 173 and 165 points, respectively (Student t test, p = 0.45). The self-reported Patient-Specific Index was concordant with the interviewer administered Patient-Specific Index (intraclass correlation coefficient, 0.78). CONCLUSIONS: We concluded that a self-reported version of the Patient-Specific Index, which focuses on the concerns of individuals, is reliable and has criterion validity compared with an interviewer-administered version. PMID- 10859103 TI - Challenges in evaluating patients lost to follow-up in clinical studies of rotator cuff tears. AB - BACKGROUND: Long-term follow-up studies are necessary to critically evaluate the outcome of a treatment intervention for a specific disorder. However, patients may cease participating in a long-term study and become lost to follow-up; thus, their current condition is unknown. The underlying characteristics that predispose a patient to become lost to follow-up are difficult to identify and control. Patients who are lost to follow-up may be contacted by telephone; however, the effect of administering a functional assessment questionnaire by telephone compared with that of mailing a questionnaire is unknown. The purpose of this study was to compare patients who continued to respond to requests for follow-up with those who did not. A second purpose was to compare responses obtained by mail with those obtained by telephone interview. METHODS: Two hundred and twenty-four patients with a rotator cuff tear were enrolled in an ongoing study of shoulder function and general health. Self-assessment questionnaires were mailed to every patient at six-month intervals. Sixty-seven patients (30 percent) regularly responded to mailings (identified as responders in this study), fifty-five patients (25 percent) responded occasionally (these patients were not included in the analysis), and 102 patients (46 percent) ceased to respond and became lost to follow-up (identified as nonresponders in this study). This investigation was performed to determine: (1) the characteristics of nonresponders compared with those of responders, (2) the functional status of nonresponders as assessed with a questionnaire over the telephone, and (3) the effect of administering a self-assessment functional questionnaire by telephone compared with that of sending the same questionnaire by mail. RESULTS: Nonresponders tended to have lower initial scores for the mental health summary (p = 0.03) and for social function (p = 0.01), were less likely to have had surgery (p = 0.009), and were less likely to consume alcohol (p = 0.03). At the last known time when they completed the mailed questionnaire, nonresponders reported significantly worse shoulder function than responders (p = 0.0001). However, on telephone questioning the mean number of shoulder functions that the nonresponders indicated that they could perform was greater than the mean number documented on their last mailed questionnaire (p < 0.0001). In a random subgroup of responders, the mean number of functions that the patients indicated that they could perform when interviewed by telephone was significantly greater than the number indicated on their most recent mailed questionnaire (p < 0.01). The results obtained by telephone from this random subgroup of responders were similar to those obtained by telephone from the nonresponders. CONCLUSIONS: There are differences between patients who continue to participate in a study and those who become lost to follow-up. Functional assessment questionnaires administered by telephone yield different results than the same questionnaires sent by mail. These considerations are relevant to the design, implementation, and interpretation of clinical studies in which functional questionnaires are used. PMID- 10859104 TI - Competence of the deltoid ligament in bimalleolar ankle fractures after medial malleolar fixation. AB - BACKGROUND: The stability of the ankle joint is provided by the medial and lateral malleoli and ligaments. Recent studies of cadaveric ankles have demonstrated that injury to the medial structures of the ankle is necessary to allow lateral subluxation of the talus after fracture. However, cadaveric models are limited by the fracture pattern chosen for the model. We sought to investigate the competency of the deltoid ligament in vivo in patients with an operatively treated bimalleolar ankle fracture. METHODS: Twenty-seven patients with a bimalleolar ankle fracture were evaluated. In each patient, the medial malleolus was anatomically reduced and fixed. A radiograph of the ankle was then made with application of an external rotation load to the joint. All lateral malleolar injuries were then reduced and fixed. The radiographs were evaluated for restoration of the competence of the deltoid ligament according to established criteria. RESULTS: Seven (26 percent) of the twenty-seven patients had radiographically evident incompetence of the deltoid ligament after medial malleolar fixation. This finding was associated with a small medial malleolar fragment. CONCLUSIONS: In bimalleolar fractures, the medial injury may be an osseous avulsion, leaving the deltoid intact on the displaced fragment, or it may be a combination of ligamentous and osseous injury with disruption of the deep portion of the deltoid ligament. PMID- 10859105 TI - Dynamic glenohumeral stability provided by the rotator cuff muscles in the mid range and end-range of motion. A study in cadavera. AB - BACKGROUND: Both static and dynamic factors are responsible for glenohumeral joint stability. We hypothesized that dynamic factors could potentially operate throughout the entire range of glenohumeral motion, although capsuloligamentous restraints (a static factor) have been thought to be primarily responsible for stability in the end-range of motion. The purpose of this study was to quantitatively compare the dynamic glenohumeral joint stability in the end-range of motion (the position of anterior instability) with that in the mid-range by investigating the force components generated by the rotator cuff muscles. METHODS: Ten fresh-frozen shoulders from human cadavera were obtained, and all soft tissues except the rotator cuff were removed. The glenohumeral capsule was resected after the rotator cuff muscles had been released from the scapula. A specially designed frame positioned the humerus in 60 degrees of abduction and 45 degrees of extension with respect to the scapula. The compressive and shear components on the glenoid were measured before and after a constant force was applied individually to each muscle with the humerus in five different positions (from neutral to 90 degrees of external rotation). The dynamic stability index, a new biomechanical parameter reflecting these force components and the concavity compression mechanism, was calculated. The higher the dynamic stability index, the greater the dynamic glenohumeral stability. RESULTS: In the mid-range of motion, the supraspinatus and subscapularis provided higher dynamic stability indices than did the other muscles (p < 0.05). On the other hand, when the position of anterior instability was simulated in the end-range of motion, the subscapularis, infraspinatus, and teres minor provided significantly higher dynamic stability indices than did the supraspinatus (p < 0.005). CONCLUSIONS: The rotator cuff provided substantial anterior dynamic stability to the glenohumeral joint in the end-range of motion as well as in the mid-range. CLINICAL RELEVANCE: A glenohumeral joint with a lax capsule and ligaments might be stabilized dynamically in the end-range of motion if the glenoid concavity is maintained and the function of the external and internal rotators, which are efficient stabilizers in this position, is enhanced. PMID- 10859106 TI - Spontaneous osteonecrosis of the knee: the result of subchondral insufficiency fracture. AB - BACKGROUND: Spontaneous osteonecrosis of the knee is a superficial subchondral lesion classically seen in the medial femoral condyle; in general, it is markedly different in its clinicopathological presentation from the classic wedge-shaped subchondral osteonecrotic lesions seen in the hip, knee, and other joints. Recent reports on subchondral insufficiency fracture of the femoral head, which has marked morphological similarities with spontaneous osteonecrosis of the knee, led us to reevaluate a series of patients who had had operative treatment because of a clinical and pathological diagnosis of spontaneous osteonecrosis of the knee. METHODS: We reviewed the cases of fourteen patients who had had operative treatment of spontaneous osteonecrosis of the knee in order to reevaluate the gross and histological morphology of this lesion. The patients included eight women and six men who ranged in age from fifty-nine to eighty-eight years. In all patients, the diagnosis of spontaneous osteonecrosis of the knee had been based on clinical presentation, imaging studies, and pathological findings. The appearance of the lesion on plain radiographs was categorized into four stages, which corresponded to the gross and histological findings. In stage 1, the radiographic appearance is normal; in stage 2, a radiolucent oval area is seen subchondrally or there is slight flattening of the convexity of the condyle, or both; in stage 3, the radiolucent area is expanded and is surrounded by a sclerotic halo; and in stage 4, secondary osteoarthritic changes are apparent. RESULTS: No patient had a stage-1 lesion. Three patients, all of whom had a stage 2 lesion, were considered to have a subchondral insufficiency fracture of the medial femoral condyle. Another six patients, all of whom had a stage-3 lesion, were considered to have a subchondral fracture and associated focal osteonecrosis that was confined to the area between the fracture line and the articular surface. The remaining five patients, three of whom had a stage-3 lesion and two of whom had a stage-4 lesion, had indeterminate findings because the lesion had become detached from the condyle. CONCLUSIONS: Our histopathological findings suggest that the primary event leading to spontaneous osteonecrosis of the knee is a subchondral insufficiency fracture and that the localized osteonecrosis seen in association with this disease is the result of a fracture. PMID- 10859107 TI - Chronic recurrent posterior dislocation of the hip after a Pipkin fracture treated with intertrochanteric osteotomy and acetabuloplasty. A case report. PMID- 10859108 TI - Vitamin C reduced the incidence of reflex sympathetic dystrophy after wrist fracture. PMID- 10859109 TI - Botulinum toxin type A improved ankle function in children with cerebral palsy and dynamic equinus foot deformity. PMID- 10859110 TI - Orthopaedic information mastery: applying evidence-based information tools to improve patient outcomes while saving orthopaedists' time. PMID- 10859111 TI - Orthopaedic injuries in children secondary to airbag deployment. PMID- 10859112 TI - Wear rate of polyethylene bearing components. PMID- 10859113 TI - Analysis of blood management. PMID- 10859114 TI - Classification of thoracic adolescent idiopathic scoliosis. PMID- 10859115 TI - Collective bargaining. PMID- 10859116 TI - Occupational orthopaedics. PMID- 10859117 TI - Supplement 1. American Diabetes Association: clinical practice recommendations 2000. PMID- 10859118 TI - Measurement of a Peak in the Cosmic Microwave Background Power Spectrum from the North American Test Flight of Boomerang. AB - We describe a measurement of the angular power spectrum of anisotropies in the cosmic microwave background (CMB) at scales of 0&fdg;3 to 5 degrees from the North American test flight of the Boomerang experiment. Boomerang is a balloon borne telescope with a bolometric receiver designed to map CMB anisotropies on a long-duration balloon flight. During a 6 hr test flight of a prototype system in 1997, we mapped more than 200 deg(2) at high Galactic latitudes in two bands centered at 90 and 150 GHz with a resolution of 26&arcmin; and 16&farcm;5 FWHM, respectively. Analysis of the maps gives a power spectrum with a peak at angular scales of 1 degrees with an amplitude 70 uK(CMB). PMID- 10859119 TI - A Measurement of Omega from the North American Test Flight of Boomerang. AB - We use the angular power spectrum of the cosmic microwave background, measured during the North American test flight of the Boomerang experiment, to constrain the geometry of the universe. Within the class of cold dark matter models, we find that the overall fractional energy density of the universe Omega is constrained to be 0.85/=100 h(-1) Mpc can be probed down to the amplitude of less than 200 km s(-1) with the MAP data and down to only approximately 30 km s(-1) with the Planck mission. PMID- 10859121 TI - On the Role of Minor Galaxy Mergers in the Formation of Active Galactic Nuclei. AB - The large-scale ( approximately 100 kpc) environments of Seyfert galaxies are not significantly different from those of non-Seyfert galaxies. In the context of the interaction model of the formation of active galactic nuclei (AGNs), it has thus been proposed that AGNs form via "minor mergers" of large disk galaxies with smaller companions. We test this hypothesis by comparing the nuclear spectra of 105 bright nearby galaxies with measurements of their R- or r-band morphological asymmetries at three successive radii. We find no significant differences between these asymmetries among the 13 Seyfert galaxies in the sample and galaxies having other nuclear spectral types (absorption, H ii region-like, LINER), nor is there strong qualitative evidence that such mergers have occurred among any of the Seyfert galaxies or LINERs. Thus, either any minor mergers began greater, similar1 Gyr ago and are essentially complete, or they did not occur at all, and AGNs form independently of any type of interaction. Support for the latter interpretation is provided by the growing evidence that supermassive black holes exist in the cores of most elliptical and early-type spiral galaxies, which in turn suggests that nuclear activity represents a normal phase in the evolution of the bulges of massive galaxies. Galaxy mergers may increase the luminosity of Seyfert nuclei to the level of QSOs, which could explain why the latter objects appear to be found in rich environments and in interacting systems. PMID- 10859122 TI - The Mass Assembly and Star Formation Characteristics of Field Galaxies of Known Morphology. AB - We discuss a new method for inferring the stellar mass of a distant galaxy of known redshift based on the combination of a near-IR luminosity and multiband optical photometry. The typical uncertainty for field galaxies with I<22 in the redshift range 0/=56) elements for each of the three stars is well matched by a scaled solar system r-process abundance distribution. Second, a weighted mean-observed Th/Eu ratio for the stars implies an age for the neutron-capture material in M15 stars of 14+/-3 Gyr, in reasonable agreement with other recent age estimates for Galactic globular clusters. PMID- 10859125 TI - Near-Infrared Imaging Polarimetry of the GG Tauri Circumbinary Ring. AB - We present 1 um Hubble Space Telescope/near-infrared camera and multiobject spectrometer resolved imaging polarimetry of the GG Tau circumbinary ring. We find that the ring displays east-west asymmetries in surface brightness as well as several pronounced irregularities but is smoother than suggested by ground based adaptive optics observations. The data are consistent with a 37 degrees system inclination and a projected rotational axis at a position angle of 7 degrees east of north, determined from millimeter imaging. The ring is strongly polarized, up to approximately 50%, which is indicative of Rayleigh-like scattering from submicron dust grains. Although the polarization pattern is broadly centrosymmetric and clearly results from illumination of the ring by the central stars, departures from true centrosymmetry and the irregular flux suggest that binary illumination, scattering through unresolved circumstellar disks, and shading by these disks may all be factors influencing the observed morphology. We confirm a approximately 0&farcs;25 shift between the inner edges of the near infrared and millimeter images and find that the global morphology of the ring and the polarimetry provide strong evidence for a geometrically thick ring. A simple Monte Carlo scattering simulation is presented that reproduces these features and supports the thick-ring hypothesis. We cannot confirm filamentary streaming from the binary to the ring, also observed in the ground-based images, although it is possible that there is material inside the dynamically cleared region that might contribute to filamentary deconvolution artifacts. Finally, we find a faint fifth point source in the GG Tau field that, if it is associated with the system, is almost certainly a brown dwarf. PMID- 10859126 TI - A Theoretical Light-Curve Model for the 1985 Outburst of RS Ophiuchi. AB - A theoretical light-curve model of the 1985 outburst of RS Ophiuchi based on a thermonuclear runaway model is presented. The system consists of a very massive white dwarf (WD) with an accretion disk and a red giant. The early phase of the V light curve is well reproduced only by the bloated WD photosphere of the thermonuclear runaway model on a 1.35+/-0.01 M middle dot in circle WD, while the later phase is dominated both by the irradiated accretion disk and by the irradiated red giant underfilling the inner critical Roche lobe. The UV light curve is also well reproduced by the same model with a distance of 0.6 kpc to RS Oph. The envelope mass at the optical peak is estimated to be 2x10-6 M( middle dot in circle), indicating a rather high mass accretion rate of 1.2x10-7 M( middle dot in circle) yr(-1) between the 1967 and 1985 outbursts. About 90% of the envelope mass is blown off in the outburst wind, while the residual 10% (2x10 7 M( middle dot in circle)) has been left and added to the helium layer of the WD. The net increasing rate of the WD mass is 1.2x10-8 M( middle dot in circle) yr(-1). Thus, RS Oph is certainly a strong candidate for a Type Ia supernova progenitor. PMID- 10859127 TI - Detection of Lead in the Carbon-rich, Very Metal-poor Star LP 625-44: A Strong Constraint on s-Process Nucleosynthesis at Low Metallicity. AB - We report the detection of the Pb i lambda4057.8 line in the very metal-poor (&sqbl0;Fe&solm0;H&sqbr0;=-2.7), carbon-rich star, LP 625-44. We determine the abundance of Pb (&sqbl0;Pb&solm0;Fe&sqbr0;=2.65) and 15 other neutron-capture elements. The abundance pattern between Ba and Pb agrees well with a scaled solar system s-process component, while the lighter elements (Sr-Zr) are less abundant than Ba. The enhancement of s-process elements is interpreted as a result of mass transfer in a binary system from a previous asymptotic giant branch (AGB) companion, an interpretation strongly supported by radial velocity variations of this system. The detection of Pb makes it possible, for the first time, to compare model predictions of s-process nucleosynthesis in AGB stars with observations of elements between Sr and Pb. The Pb abundance is significantly lower than the prediction of recent models (e.g., Gallino et al.), which succeeded in explaining the metallicity dependence of the abundance ratios of light s-elements (Sr-Zr) to heavy ones (Ba-Dy) found in previously observed s process-enhanced stars. This suggests that one should either (1) reconsider the underlying assumptions concerning the (13)C-rich s-processing site ((13)C pocket) in the present models or (2) investigate alternative sites of s-process nucleosynthesis in very metal-poor AGB stars. PMID- 10859128 TI - Possible Rapid Gas Giant Planet Formation in the Solar Nebula and Other Protoplanetary Disks. AB - Gas giant planets have been detected in orbit around an increasing number of nearby stars. Two theories have been advanced for the formation of such planets: core accretion and disk instability. Core accretion, the generally accepted mechanism, requires several million years or more to form a gas giant planet in a protoplanetary disk like the solar nebula. Disk instability, on the other hand, can form a gas giant protoplanet in a few hundred years. However, disk instability has previously been thought to be important only in relatively massive disks. New three-dimensional, "locally isothermal," hydrodynamical models without velocity damping show that a disk instability can form Jupiter-mass clumps, even in a disk with a mass (0.091 M middle dot in circle within 20 AU) low enough to be in the range inferred for the solar nebula. The clumps form with initially eccentric orbits, and their survival will depend on their ability to contract to higher densities before they can be tidally disrupted at successive periastrons. Because the disk mass in these models is comparable to that apparently required for the core accretion mechanism to operate, the models imply that disk instability could obviate the core accretion mechanism in the solar nebula and elsewhere. PMID- 10859129 TI - Fine Structure in Solar Flares. AB - We present observations of several large two-ribbon flares observed with both the Transition Region and Coronal Explorer (TRACE) and the soft X-ray telescope on Yohkoh. The high spatial resolution TRACE observations show that solar flare plasma is generally not confined to a single loop or even a few isolated loops but to a multitude of fine coronal structures. These observations also suggest that the high-temperature flare plasma generally appears diffuse while the cooler ( less, similar2 MK) postflare plasma is looplike. We conjecture that the diffuse appearance of the high-temperature flare emission seen with TRACE is due to a combination of the emission measure structure of these flares and the instrumental temperature response and does not reflect fundamental differences in plasma morphology at the different temperatures. PMID- 10859130 TI - The Origin of the Solar Flare Waiting-Time Distribution. AB - It was recently pointed out that the distribution of times between solar flares (the flare waiting-time distribution) follows a power law for long waiting times. Based on 25 years of soft X-ray flares observed by Geostationary Operational Environmental Satellite instruments, it is shown that (1) the waiting-time distribution of flares is consistent with a time-dependent Poisson process and (2) the fraction of time the Sun spends with different flaring rates approximately follows an exponential distribution. The second result is a new phenomenological law for flares. It is shown analytically how the observed power law behavior of the waiting times originates in the exponential distribution of flaring rates. These results are argued to be consistent with a nonstationary avalanche model for flares. PMID- 10859131 TI - Interferometric Measurement of Resonance Transition Wavelengths in C iv, Si iv, Al iii, Al ii, and Si ii. AB - We have made the first interferomeric measurements of the wavelengths of the important ultraviolet diagnostic lines in the spectra C iv near 155 nm and Si iv near 139 nm with a vacuum ultraviolet Fourier transform spectrometer and high current discharge sources. The wavelength uncertainties were reduced by 1 order of magnitude for the C iv lines and by 2 orders of magnitude for the Si iv lines. Our measurements also provide accurate wavelengths for resonance transitions in Al iii, Al ii, and Si ii. PMID- 10859132 TI - Neuronal interactions related to working memory processes in the primate prefrontal cortex revealed by cross-correlation analysis. AB - To understand neuronal mechanisms for manipulating and/or integrating information in working memory processes, we examined functional interactions among prefrontal neurons exhibiting various task-related activities by cross-correlation analysis. Among 168 neuron pairs isolated, 84 (50%) had significant peaks in cross correlograms (CCGs); 30 had excitatory central peaks at time 0, 38 had excitatory peaks displaced from time 0, 13 had inhibitory central peaks, and three had both excitatory and inhibitory peaks displaced from time 0. Although significant interactions were observed among prefrontal neurons having various task-related activities, the information flow is present from prefrontal neurons having cue period activity to neurons having oculomotor activity through neurons having delay-period activity. In addition, neuron pairs both having delay-period activity tended to have significant excitatory peaks in CCGs. Further, neuron pairs that had excitatory central peaks in CCGs tended to have similar directional preferences in task-related activities, and this similarity was the highest in neuron pairs both having cue-period activity. Neuron pairs that had displaced peaks in CCGs also showed similarity in directional preferences in task related activities, and this similarity was also higher in neuron pairs both having cue-period activity. Interactions between neurons exhibiting task-related activity with different directional preferences increase as the temporal sequence of the task progresses. These results suggest that these interactions play an important role for manipulating and integrating information. PMID- 10859133 TI - An event-related functional MRI study of the stroop color word interference task. AB - In this study we have attempted to define the neural circuits differentially activated by cognitive interference. We used event-related functional magnetic resonance imaging (fMRI) to identify areas of the brain that are activated by the Stroop word-color task in two experiments. In the first experiment, we used infrequent, incongruent colored word stimuli to elicit strong Stroop interference (the 'conventional Stroop' paradigm). In the second experiment, we used infrequent, congruent colored words (the 'inverse Stroop' paradigm) to confirm that the regions identified in the first experiment were in fact specifically related to the Stroop effect and not to nonspecific oddball effects associated with the use of infrequent stimuli. Performance of the conventional Stroop specifically activated the anterior cingulate, insula, premotor and inferior frontal regions. These activated regions in the current experiment are consistent with those activated in fMRI experiments that use a more traditional block design. Finally, analysis of the time course of fMRI signal changes demonstrated differential onset and offset of signal changes in these activated regions. The time course results suggest that the action of various brain areas can be temporally dissociated. PMID- 10859134 TI - Developmental changes in cell calcium homeostasis during neurogenesis of the embryonic rat cerebral cortex. AB - We quantified cytoplasmic Ca(2+) (Ca(2+)(c)) levels in cells dissociated from the embryonic (E) rat cortex during neurogenesis. Dual-recordings by flow cytometry using calcium and voltage-sensitive dyes revealed that, at the beginning of cortical development (E11-12), precursor cells exhibited either low (<100 nM), moderate (approximately 250 nM) or high (>1 microM) resting Ca(2+)(c) levels and well-polarized (-70 mV) or less-polarized (-40 mV) resting membrane potentials which reflected postmitotic or proliferative stages of the cell cycle. Ca(2+)(c) levels of all cells included a Ca(2+)(o) entry component, which was also Mn(2+) permeant in actively proliferating precursors. Postmitotic, but not premitotic, precursors exhibited thapsigargin-sensitive intracellular Ca(2+) (Ca(2+)(i)) stores, which had similar capacities throughout neuronal lineage development. Differentiating neurons, but not precursors expressed Ca(2+)(i) stores with ryanodine and caffeine sensitivity and baseline Ca(2+)(c) levels that depended on Na(+)-Ca(2+) exchange activity. Voltage-dependent Ca(2+)(o) entry was not detected in precursors, but emerged during neuronal differentiation, with most of the neurons expressing functional L-type Ca(2+) channels. Ca(2+) imaging of individually immunoidentified cells acutely recovered in culture confirmed that precursors differentiate into neurons which stereotypically exhibit Ca(2+)(o) entry at the level of the membrane with increased Ca(2+)(i) release mechanisms on Ca(2+)(i) stores, Na(+)-Ca(2+) exchange activity and expression of voltage dependent Ca(2+) channels. PMID- 10859135 TI - Binaural interactions in primary auditory cortex of the awake macaque. AB - The functional organization of primary auditory cortex in non-primates is generally modeled as a tonotopic gradient with an orthogonal representation of independently mapped binaural interaction columns along the isofrequency contours. Little information is available regarding the validity of this model in the primate brain, despite the importance of binaural cues for sound localization and auditory scene analysis. Binaural and monaural responses of A1 to pure tone stimulation were studied using auditory evoked potentials, current source density and multiunit activity. Key findings include: (i) differential distribution of binaural responses with respect to best frequency, such that 74% of the sites exhibiting binaural summation had best frequencies below 2000 Hz; (ii) the pattern of binaural responses was variable with respect to cortical depth, with binaural summation often observed in the supragranular laminae of sites showing binaural suppression in thalamorecipient laminae; and (iii) dissociation of binaural responses between the initial and sustained action potential firing of neuronal ensembles in A1. These data support earlier findings regarding the temporal and spatial complexity of responses in A1 in the awake state, and are inconsistent with a simple orthogonal arrangement of binaural interaction columns and best frequency in A1 of the awake primate. PMID- 10859136 TI - A parametric manipulation of central executive functioning. AB - The central executive is both an important and poorly understood construct that is invoked in current theoretical models of human cognition and in various dysexecutive clinical syndromes. We report a task designed to isolate one elementary executive function, namely the allocation of attentional resources within working memory. The frequency with which attention was switched between items in working memory was varied across different trials, while storage and rehearsal demands were held constant. Functional magnetic resonance imaging revealed widespread areas, both prefrontal and more posterior, that differentially activated as a function of a trial's executive demands. Furthermore, areas that differed as a function of executive demands tended to lie adjacent to areas that were activated during the task but that did not so differ. Together, these data suggest that a distributed neuroanatomy, rather than a specific and unique locus, underlies this attention switching executive function. PMID- 10859137 TI - Arrangement of orientation pinwheel centers around area 17/18 transition zone in cat visual cortex. AB - In the primary visual cortex of higher mammals, orientation preferences are represented continuously except for singular points, so-called pinwheel centers. In spite of the uniqueness of orientation pinwheel centers, very little is known about the pattern of their arrangement. In this study we examined the arrangement of orientation pinwheel centers in the cat visual cortex by optical imaging of intrinsic signals. Our results demonstrate that orientation pinwheel centers are arranged in a unique geometric pattern around the area 17/18 transition zone: pinwheel centers of the same type are arranged in rows parallel to the transition zone, and rows of clockwise and counterclockwise pinwheel centers are arranged alternately. We suggest that the areal border imposes a strong restriction on the pattern formation of orientation preference maps in the visual cortex. PMID- 10859138 TI - Disruption of neuronal migration and radial glia in the developing cerebral cortex following ablation of Cajal-Retzius cells. AB - Cortical neurons are generated within the proliferative layer and follow a strict 'inside-out' gradient of migration and positioning, which determines the characteristic layering and pattern of neural connections in the adult cerebral cortex. Thus, directional migration of postmitotic neuroblasts towards layer I and regulation of the radial glia phenotype subserving cortical migration are central issues in corticogenesis. Recent studies showing that the gene disrupted in the reeler mutation--reelin--is expressed in Cajal-Retzius cells have indicated a role for these pioneer neurons in cortical migration. We show here that ablation of Cajal-Retzius cells in layer I by local application of domoic acid in newborn mice arrests migration of the late-generated neurons, destined to cortical layers II-III, that have been labeled by 5-bromodeoxyuridine injections administered at E16. In addition, degeneration of Cajal-Retzius cells in newborn mice dramatically decreases the number of radial glial apical processes identified by nestin-immunostaining, but increases the number of maturing glial fibrillary acidic protein-positive astrocytes. These findings support an essential role for Cajal-Retzius cells in neuronal migration and corticogenesis, by regulating the identity and function of radial glia and the radial glia-to astrocyte transformation. PMID- 10859139 TI - Depotentiation in the dentate gyrus of freely moving rats is modulated by D1/D5 dopamine receptors. AB - Hippocampal depotentiation comprises a reversal of tetanization- induced long term potentiation (LTP) which occurs following low-frequency stimulation. In the CA1 region, it has been reported that agonist activation of D1/D5 dopamine receptors enhances LTP expression and inhibits depotentiation. The role of these receptors in synaptic plasticity in the dentate gyrus (DG) has not been characterized. This study therefore investigated the role of D1/D5 receptors in LTP and depotentiation in the DG of freely moving rats. Male Wistar rats underwent chronic implantation of a recording electrode in the DG granule cell layer, a bipolar stimulating electrode in the medial perforant path and a cannula in the ipsilateral cerebral ventricle (to enable drug administration). The D1/D5 agonist Chloro-PB dose-dependently inhibited depotentation in the DG. This effect was prevented by the D1/D5 antagonist SCH 23390. Neither D1/D5 agonist nor antagonist had an effect on LTP expression or basal synaptic transmission. These results highlight differences between D1/D5 receptor-involvement in LTP and depotentiation in the CA1 region and DG, and indicate that whereas D1/D5 receptor activation may not be a critical factor in LTP induction in the DG, a differential role for these receptors in the expression of depotentiation, in this hippocampal subfield, may exist. PMID- 10859140 TI - Colocalization of glutamate ionotropic receptor subunits in the human temporal neocortex. AB - alpha-Amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA), kainate and N-methyl D-aspartate (NMDA) receptors represent major classes of glutamate receptors (GluR) which play fundamental roles in normal excitatory synaptic activity and, probably, in the etiology of several brain diseases. These receptors are composed of multiple receptor subunit proteins, and the differential expression of these subunits in cortical neurons is considered to be one of the substrates for the functional diversity of cortical excitatory circuitry. In the monkey neocortex, different subpopulations of neurons have been identified on the basis of immunocytochemical colocalization studies using subunit-specific antibodies, but no comparable investigations have been made in the human neocortex. The aim of the present study was to determine quantitatively GluR subunit combinations in the human temporal neocortex by double-labeling immunocyto- chemical experiments. We quantified the neuronal populations expressing different receptor subtypes with fluorescent tags visualizing them with confocal laser microscopy. We studied AMPA, kainate- and NMDA-receptor subunits, using antibodies against GluR1, GluR2, GluR2/3, GluR2/4, GluR5/6/7 and NMDAR1 subunits. A high degree of colocalization (93-100%) using combinations of antibodies against GluR2 with GluR2/3, GluR2/3 with GluR2/4, and GluR2 or GluR2/4 with NMDAR1 was found, whereas for other combinations the degree of colocalization varied between 38% and 88%. Some of the percentages reported here are similar to those found in the monkey cortex, whereas others differ considerably. These results emphasize the diversity of excitatory circuits in the human neocortex, and suggest species differences with regard to some of these GluR-mediated circuits. PMID- 10859141 TI - Selective and potent inhibition of human CYP2C19 activity by a conformationally targeted antipeptide antibody. AB - A conformationally targeted anti-peptide antibody was produced by immunizing a rabbit with a cyclized peptide corresponding to a loop region of human CYP2C19 (residues 250-261). In an enzyme-linked immunosorbent assay, the antibody bound strongly to recombinant CYP2C19 and poorly to recombinant CYP2C8, CYP2C9, and CYP2C18. In immunoblotting studies, the antibody bound strongly to recombinant CYP2C19 and weakly to recombinant CYP2C8. No binding to recombinant CYP1A2, CYP2C9, CYP2C18, CYP2D6, CYP2E1, and CYP3A4 was detected. In immunoinhibition experiments, the anti-peptide antibody targeted against CYP2C19 potently inhibited (S)-mephenytoin 4'-hydroxylase activity of human hepatic microsomal fraction (>90%). It had no appreciable effect on ethoxyresorufin O-deethylase (CYP1A2), tolbutamide methyl-hydroxylase (CYP2C9), dextromethorphan O-demethylase (CYP2D6), 4-nitrophenol hydroxylase (CYP2E1), or testosterone 6beta-hydroxylase (CYP3A4) activity of human hepatic microsomal fraction. However, large amounts of purified IgG fractions were able to inhibit up to 35% of paclitaxel 6alpha hydroxylase (CYP2C8) activity. In conclusion, we have demonstrated that an anti peptide antibody targeted against residues 250 to 261 of human CYP2C19 selectively and potently inhibited CYP2C19 activity of human hepatic microsomal fraction. PMID- 10859143 TI - Metabolism of ticlopidine by activated neutrophils: implications for ticlopidine induced agranulocytosis. AB - Ticlopidine is associated with a relatively high incidence of agranulocytosis and aplastic anemia. We have shown that other drugs associated with agranulocytosis are metabolized to reactive metabolites by activated human neutrophils or by HOCl, which is the major oxidant produced by activated neutrophils. We set out to test the hypothesis that ticlopidine also fits this pattern and is oxidized to a reactive intermediate by activated neutrophils and HOCl. As much as 8% ticlopidine was metabolized by activated human neutrophils to a dehydro ticlopidine; however, this product did not account for all of the decrease in ticlopidine concentration. The oxidation products of ticlopidine by the combination of myeloperoxidase and hydrogen peroxide were the same as those by HOCl: dehydrogenated ticlopidine and 2-chloroticlopidine. A neutrophil-derived reactive metabolite of ticlopidine was trapped with GSH and the same ticlopidine GSH conjugate was found in both the myeloperoxidase and HOCl systems. Evidence for the identity of the reactive metabolite was obtained by reaction of ticlopidine with HOCl in a flow reaction system coupled to a mass spectrometer. The mass spectra suggested that the reactive metabolite was a thiophene-S chloride. We conclude that ticlopidine follows the same pattern of reactive metabolite formation by activated neutrophils as other drugs associated with a high incidence of agranulocytosis, and the putative thiophene-S-chloride formed by activated neutrophils may be responsible for ticlopidine-induced agranulocytosis. PMID- 10859142 TI - Structure-function analysis of human CYP3A4 using a specific proinhibitory antipeptide antibody. AB - An anti-peptide antibody targeted against residues 253 to 269 of human CYP3A4 was produced that specifically and potently inhibited its activity in human hepatic microsomal fraction (>90%). The function of this region in P450 catalysis was investigated. Antibody binding to CYP3A4 was unable to affect the magnitude of the Type I spectrum on addition of testosterone. It also had no effect on the K(m) of the enzyme for testosterone, but it did cause a marked decrease in V(max) (>90%) of testosterone 6 beta-hydroxylation. There was no change in the ability of the antibody-bound CYP3A4 to form the steady-state level of the enzymatically or chemically reduced P450-CO complex or even the steady-state level of the dioxy ferrous complex during testosterone metabolism, but the oxidation of NADPH by CYP3A4 in the presence of antibody was 60% that of CYP3A4 in the absence of antibody. The binding of the antibody also resulted in potent inhibition of cumene hydroperoxide-supported testosterone 6 beta-hydroxylase activity of human liver microsomal fraction (>90%). Our conclusion is that the loop region targeted in CYP3A4 is not involved in substrate binding, in reductase binding, in the transfer of the first or second electron from the reductase to CYP3A4, or in the binding of molecular oxygen. We speculate that antibody binding to CYP3A4 inhibits enzyme activity by destabilizing the ternary hydroperoxo complex, by interfering with the second proton transfer, and/or by interfering with the conformational changes that are suggested to be induced by substrate binding. PMID- 10859144 TI - Pharmacokinetics and hepatic disposition of bis[1-(ethoxycarbonyl)propyl]5 acetylamino-2,4,6-triiodoisophthalate in rats and isolated perfused rat livers. AB - Bis[1-(Ethoxycarbonyl)propyl]5-acetylamino-2,4,6- triiodoisophthalate+ (NC 68183) was designed as a new computed tomography imaging agent. The purpose of this study was to determine the pharmacokinetics and metabolism of NC 68183 in conscious rats and in the isolated perfused rat liver. Animals were i.v. dosed at 69 and 690 mg of iodine/kg. Blood samples were collected at 5, 15, 30, and 60 min, and 7 days after dosing. Tissue samples (liver, kidney, and spleen) were taken at 60 min and 7 days after dosing. NC 68183 was cleared from blood in first order kinetics following an i.v. administration of 69 mg I/kg. The volume of distribution (Vss) at steady state and elimination half-life (t(1/2)) were estimated as 24 ml and 11 min. The clearance of NC 68183 from blood was changed to zero-order kinetics following administration of 690 mg/kg, and its elimination rate was 16 microg I/ml.min. The liver and spleen were the only tissues to have the nanoparticle residue at day 7 following administration. NC 68183 (75 mg of agent, 35 mg of I) was injected into the isolated perfused rat liver system. Bile flow increased from 1.0 to 1.3 microl/min/g liver following administration. The biliary excretion rate maximum was estimated as 11 microg/min/g liver. The metabolite was identified using liquid chromatography/mass spectrometry as a monocarboxylic acid product, which exclusively excreted into the bile in a soluble iodinated metabolite. Pharmacokinetics data suggested that NC 68183 primarily resides in the blood pool following an i.v. administration with a plasma half-life appropriate for blood pool imaging. PMID- 10859145 TI - Metabolism of the influenza neuraminidase inhibitor prodrug oseltamivir in the rat. AB - The metabolism of [2-acetyl-(14)C]oseltamivir (GS4104, Ro 64-0796), the prodrug of the novel influenza neuraminidase inhibitor GS4071 (Ro 64-0802), was examined in rats after oral dosing. Intact oseltamivir was observed only in lung and urine, accounting for 37 and 15% of the total radioactivity in these samples, respectively. GS4071 was the major metabolite in plasma, tissues, and urine, and accounted for 32 to 56% of the radioactivity present in these samples. The second most abundant peak in these samples (13-24% of radioactivity) was a novel metabolite (M3). This metabolite was purified from urine of rats dosed orally with oseltamivir and was identified by liquid chromatography-mass spectrometry and NMR as the (R)-omega-carboxylic acid metabolite of oseltamivir. The omega carboxylic acid metabolite of oseltamivir could not be produced in vitro. However, omega-hydroxylated products of oseltamivir were produced by rat liver microsomes. Both the (R)- and (S)-omega-hydroxylated products were observed, but formation of the (R)-isomer predominated. These data indicated that in the rat, oseltamivir was primarily metabolized to the active influenza neuraminidase inhibitor GS4071 and, to a lesser extent, to an (R)-omega-carboxylic acid metabolite. PMID- 10859146 TI - The effect of rate of drug administration on the extent and time course of phencyclidine distribution in rat brain, testis, and serum. AB - The goal of these studies was to examine the relationship between the rate of phencyclidine (PCP) administration and PCP tissue distribution. The time course of PCP distribution in serum, brain, and testis after rapid (i.v.) and slow (s.c.) administration was studied. Brain and serum PCP concentrations after an i.v. bolus dose (1 mg/kg at 900 microg/min) were highest at 30 s and decreased biphasically, with serum concentrations decreasing 30 times faster than brain concentrations during the early phase. Consequently, the brain-to-serum PCP concentration ratio increased from 8:1 at 30 s to 14:1 at 20 min before equilibrating at a ratio of 3:1 that remained constant from 1 to 8 h. In contrast, the testis-to-serum ratio increased slowly from 1:1 to 12:1 over 4 h, and then remained constant. In a separate group of animals, an s.c. infusion of PCP (18 mg/kg/day or 3.6 microg/min) produced a brain-to-serum ratio (6:1) that remained constant throughout the 96-h infusion. Testis-to-serum ratios increased from 4:1 at 1 h to 12:1 at 8 h and then remained constant for 96 h. Steady-state infusion of a pharmacologically inactive dose (2.5 mg/kg/day) produced a brain-to serum ratio (3:1) that was significantly lower than the ratio (6:1) after infusion of the three pharmacologically active doses (10-25 mg/kg/day). The temporary high brain PCP concentrations and the dynamic disequilibrium between brain and serum concentrations after rapid i.v. administration could provide a better understanding of the preference of the human drug abuser for rapid rates (e.g., i.v. or smoking) of drug administration. PMID- 10859147 TI - Metabolism of fluroxypyr, fluroxypyr methyl ester, and the herbicide fluroxypyr methylheptyl ester. I: during percutaneous absorption through fresh rat and human skin in vitro. AB - Percutaneous absorption of pesticides is a major determinant for risk assessment. Furthermore, cutaneous metabolism plays a role in penetration of certain chemicals. Therefore, the aim of these studies was to determine the transdermal metabolism of three related compounds [the herbicide, fluroxypyr methylheptyl ester (FPMH), fluroxypyr methyl ester (FPM), and fluroxypyr (FP)] during penetration through human and rat skin in vitro. The data presented in this article show that both FPM and FPMH were completely metabolized during their passage through human and rat skin in vitro. The only metabolite produced was that of the hydrolysis product, FP, with no parent ester penetrating through the skin. The extent of FP formation within the skin was directly correlated to the degree of stratum corneum reservoir formation. The larger the stratum corneum reservoir, the lower the levels of FP recovered from within the skin. This suggests that as the ester partitioned out of the SC it was immediately hydrolyzed to FP, which could then pass freely through the remainder of the epidermis and dermis. Similar metabolic profiles were observed for the transdermal metabolism of FPM and FPMH in previously frozen rat skin, indicating the robust nature of the esterase enzymes involved. In conclusion, systemic exposure after skin contact with FPM or FPMH is likely to be to the acid metabolite, FP, only and not to the parent ester. In addition, the rate and extent of percutaneous absorption will be a major determinant of cutaneous metabolism. PMID- 10859148 TI - Metabolism of fluroxypyr, fluroxypyr methyl ester, and the herbicide fluroxypyr methylheptyl ester. II: in rat skin homogenates. AB - Fluroxypyr methyl ester (FPM) and the herbicide fluroxypyr methylheptyl ester (FPMH) are completely hydrolyzed during penetration through human and rat skin in vitro to the acid metabolite, fluroxypyr (FP) (). This article presents additional studies to determine the enzyme kinetics (K(m) and V(max)) of this ester hydrolysis, using crude rat whole-skin homogenate. Both FPM and FPMH were extensively metabolized in rat skin homogenates to the acid metabolite, FP. In no instance were any other metabolites detected. FPM was essentially hydrolyzed completely within 1 h. In FPMH incubations, there was still parent ester present after 24 h at all concentrations tested. The kinetics of hydrolysis of the two esters were different: V(max) was approximately 3-fold greater for FPM than FPMH (1400 and 490 micromol FP/min/g of tissue, respectively); however, K(m) values were very similar, 251 and 256 microM, respectively. Preliminary inhibitory studies suggest that FPM and FPMH are hydrolyzed by a carboxylesterase, because this reaction was inhibited by bis-p-nitrophenyl phosphate. Mercuric chloride (an inhibitor of A-esterase and arylesterase) and eserine (a cholinesterase inhibitor) had no inhibitory effect on the hydrolysis of FPM or FPMH. Taken together with the data presented by, it can be concluded that no parent ester will pass through the skin in vivo, only the metabolite, FP. Therefore, first pass metabolism will be complete before these compounds reach the systemic circulation. PMID- 10859149 TI - Verapamil metabolite exposure in older and younger men during steady-state oral verapamil administration. AB - To determine the effect of age on exposure to the circulating major verapamil metabolites norverapamil, N-dealkylverapamil (D-617), and N-dealkylnorverapamil (D-620), plasma concentrations of verapamil and the three metabolites were determined during the last dose interval of a 14-day administration period of 240 mg of sustained release verapamil once daily in 11 older (aged 65-75 years) and 8 younger (20-28 years) healthy male volunteers. Area under the plasma concentration time curve (AUC) was greater for verapamil (mean +/- S. D.) (2815 +/- 733 older versus 1639 +/- 466 ng/ml.h(-1) young; P <. 0007) and norverapamil (2927 +/- 655 versus 2143 +/- 471 ng/ml. h(-1); P <.007); however, it was not significantly different for D-617 [2386 +/- 772 versus 1894 +/- 418 ng/ml.h(-1); not significantly different (NS)] and N-dealkylnorverapamil (897 +/- 366 versus 757 +/- 104 ng/ml.h(-1); NS) in older as compared with young subjects. These data indicate that impaired verapamil oral clearance previously described in older men does not result in decreased exposure to the formed major metabolites, rather there is increased exposure to norverapamil and the same or a trend toward greater exposure to D-617 as well. This suggests that in addition to the impaired clearance mechanisms for verapamil, which are thought to be primarily mediated by CYP3A, biotransformation processes distal to the formation of norverapamil and D 617 are impaired as well. PMID- 10859150 TI - Role of furanocoumarin derivatives on grapefruit juice-mediated inhibition of human CYP3A activity. AB - With juices of grapefruit and related fruits, possible relationships between contents of six different furanocoumarins and extents of inhibition of microsomal CYP3A activity have been studied in vitro. Microsomal CYP3A-mediated testosterone 6beta-hydroxylation was inhibited by the addition of a fruit juice (2.5%, v/v) from eight different grapefruit sources, two sweeties, three pomelos, and one sour orange, whereas no clear inhibition was observed with two sweet orange juices. The inhibitory component in grapefruit juice resides mainly in the precipitate rather than in the supernatant after centrifugation. Higher amounts of (R)-6',7'-dihydroxybergamottin (DHB) were distributed in the supernatant, whereas GF-I-1, GF-I-2, GF-I-4, and the newly isolated GF-I-5 and GF-I-6 were detected predominantly in the precipitate. Mixing of five representative furanocoumarins at their detectable levels in grapefruit juice reproduced roughly the inhibitory potencies of grapefruit juice, but omission of any of the components resulted in decreased potencies. These results suggested that all the major furanocoumarins contributed to the CYP3A inhibitory properties of grapefruit juice. Furthermore, all six furanocoumarins showed stronger CYP3A inhibitory potencies after preincubation in the presence of NADPH, suggesting that both competitive and mechanism-based inhibition occur in a grapefruit juices drug interaction. PMID- 10859151 TI - Absorption, disposition, and metabolism of rosiglitazone, a potent thiazolidinedione insulin sensitizer, in humans. AB - Rosiglitazone is a potent peroxisome proliferator-activated receptor gamma agonist that decreases hyperglycemia by reducing insulin resistance in patients with type 2 diabetes mellitus. The disposition of (14)C-labeled rosiglitazone was determined after oral and i.v. dosing of rosiglitazone solution, and the disposition of nonradiolabeled rosiglitazone was determined after oral dosing of tablets in this open-label, three-part, semirandomized, crossover study. The absorption of rosiglitazone was rapid and essentially complete, with absolute bioavailability estimated to be approximately 99% after oral tablet dosing and approximately 95% after oral solution dosing, and clearance was primarily metabolic. The time to maximal concentration of radioactivity and the elimination half-life for two metabolites in plasma were significantly longer than for rosiglitazone itself (4-6 h versus 0. 5-1 h, and ca. 5 days versus 3-7 h). Radioactivity was excreted primarily via the urine ( approximately 65%) and was excreted similarly after oral and i.v. dosing. The major routes of metabolism were N-demethylation and hydroxylation with subsequent conjugation, of which neither was affected by the route of drug administration. The major metabolites, those of intermediate importance, and nearly all of the trace metabolites in humans have been identified previously in preclinical studies. Rosiglitazone was well tolerated in all formulations. PMID- 10859152 TI - Measurement of cytochrome P450 gene induction in human hepatocytes using quantitative real-time reverse transcriptase-polymerase chain reaction. AB - Drug-induced changes in expression of cytochrome (CYP) P450 genes are a key cause of drug-drug interactions. Consequently, preclinical prediction of these changes by novel compounds is an integral component of drug development. To date, in vitro models of CYP induction have used mRNA measurement, immunodetection, and substrate metabolism as reporters. Here, we describe the application of quantitative real-time reverse transcriptase polymerase chain reaction to study CYP1A1 and CYP3A4 gene induction in 5-day-old cultures of human hepatocytes by known CYP inducers. After 5 days in culture, CYP1A1 expression was significantly elevated (5.1- to 26-fold; P <.01) in all four livers studied. In direct contrast, CYP3A4 mRNA levels consistently decreased during culture (80- to 300 fold; P <.001). In three independent experiments, a 48-h exposure to 3 methylcholanthrene, omeprazole, and lansoprazole significantly induced CYP1A1 expression in comparison to untreated cultures (P <.05). Rifampicin and solvent were without effect on CYP1A1 expression. Under identical experimental conditions, rifampicin and lansoprazole significantly elevated CYP3A4 mRNA expression (P <.05), whereas 3-methylcholanthrene, omeprazole, and dimethyl sulfoxide were without significant effect. These data demonstrate the applicability of quantitative reverse transcriptase polymerase chain reaction to the determination of gene dynamics in human hepatocytes. This offers a highly specific alternative to quantification of drug effects on CYP expression using immunodetection and substrate metabolism. PMID- 10859153 TI - Interaction of cisapride with the human cytochrome P450 system: metabolism and inhibition studies. AB - Using human liver microsomes (HLMs) and recombinant cytochrome P450s (CYP450s), we characterized the CYP450 isoforms involved in the primary metabolic pathways of cisapride and documented the ability of cisapride to inhibit the CYP450 system. In HLMs, cisapride was N-dealkylated to norcisapride (NORCIS) and hydroxylated to 3-fluoro-4-hydroxycisapride (3-F-4-OHCIS) and to 4-fluoro-2 hydroxycisapride (4-F-2-OHCIS). Formation of NORCIS, 3-F-4-OHCIS, and 4-F-2-OHCIS in HLMs exhibited Michaelis-Menten kinetics (K(m): 23.4 +/- 8.6, 32 +/- 11, and 31 +/- 23 microM; V(max): 155 +/- 91, 52 +/- 23, and 31 +/- 23 pmol/min/mg of protein, respectively). The average in vitro intrinsic clearance (V(max)/K(m)) revealed that the formation of NORCIS was 3.9- to 5. 9-fold higher than that of the two hydroxylated metabolites. Formation rate of NORCIS from 10 microM cisapride in 14 HLMs was highly variable (range, 4.9-133.6 pmol/min/mg of protein) and significantly correlated with the activities of CYP3A (r = 0.86, P =. 0001), CYP2C19, and 1A2. Of isoform-specific inhibitors, 1 microM ketoconazole and 50 microM troleandomycin were potent inhibitors of NORCIS formation from 10 microM cisapride (by 51 +/- 9 and 44 +/- 17%, respectively), whereas the effect of other inhibitors was minimal. Of 10 recombinant human CYP450s tested, CYP3A4 formed NORCIS from 10 microM cisapride at the highest rate (V = 0.56 +/- 0. 13 pmol/min/pmol of P450) followed by CYP2C8 (V = 0.29 +/- 0.08 pmol/min/pmol of P450) and CYP2B6 (0.15 +/- 0.04 pmol/min/pmol of P450). The formation of 3-F-4 OHCIS was mainly catalyzed by CYP2C8 (V = 0.71 +/- 0.24 pmol/min/pmol of P450) and that of 4-F-2-OHCIS by CYP3A4 (0.16 +/- 0.03 pmol/min/pmol of P450). Clearly, recombinant CYP2C8 participates in cisapride metabolism, but when the in vitro intrinsic clearances obtained were corrected for abundance of each CYP450 in the liver, CYP3A4 is the dominant isoform. Cisapride was a relatively potent inhibitor of CYP2D6, with no significant effect on other isoforms tested, but the K(i) value derived (14 +/- 16 microM) was much higher than the clinically expected concentration of cisapride (<1 microM). Our data suggest that CYP3A is the main isoform involved in the overall metabolic clearance of cisapride. Cisapride metabolism is likely to be subject to interindividual variability in CYP3A expression and to drug interactions involving this isoform. PMID- 10859154 TI - Uptake of enalapril and expression of organic anion transporting polypeptide 1 in zonal, isolated rat hepatocytes. AB - Sinusoidal entry is the first obligatory process preceding intracellular drug removal in liver. Transport of the angiotensin converting enzyme inhibitor enalapril (1-750 microM with [(3)H]enalapril), a substrate of Oatp1, the sodium independent organic anion transporting polypeptide 1 cloned from rat liver, was studied in rat hepatocytes isolated from all zones of the liver (homogeneous) and from enriched periportal (PP) and perivenous (PV) hepatocytes prepared by collagenase perfusion and zone-selective destruction with digitonin, respectively. Uptake was linear over 1 min and was concentration-dependent. Transport by the homogeneous hepatocytes (in the presence and absence of Na(+)) and PP and PV cells was described by single saturable components of similar kinetic constants (K(m) values of 344-461 microM and V(max) values of 9.5-11.6 nmol/min/10(6) cells; P >.05, ANOVA). The K(m) value for enalapril uptake in hepatocytes was of the same order of magnitude compared with that for Oatp1 expressed in HeLa cells transfected with cDNA-Oatp1 and Western blot analysis revealed similar levels of immunoreactive Oatp1 expression in PP and PV hepatocytes. However, enalapril was not taken up by Oatp2 nor by the human OATP expressed in recombinant vaccinia systems. PMID- 10859155 TI - Effect of zonal transport and metabolism on hepatic removal: enalapril hydrolysis in zonal, isolated rat hepatocytes in vitro and correlation with perfusion data. AB - Previous studies showed that the transport of enalapril occurred homogeneously among zonal rat hepatocytes. However, the metabolism of hepatic arterially delivered enalapril, swept into the rat liver by the portal or hepatic venous flow (HAPV and HAHV perfusion), was more abundant in the perivenous (PV) than the periportal (PP) region. Hence, metabolic activities toward enalapril in 9000g supernatant (S9) fractions of enriched rat PP and PV hepatocytes were examined. Although Michaelis-Menten kinetics were invariably observed, the metabolic activity toward enalapril (intrinsic clearance or V(max)(met)/K(m)(met) of 0.008 ml/min/mg of S9 protein, V(max)(met) of 21 +/- 6 nmol/min/mg of S9 protein, and K(m)(met) of 2612 +/- 236 microM) was greater in PV than in PP (V(max)(met) of 5.5 +/- 3.1 nmol/min/mg of S9 protein and K(m)(met) of 1049 +/- 335 microM; intrinsic clearance of 0.0052 ml/min/mg of S9 protein) hepatocytes. These metabolic intrinsic clearances were much lower than the sinusoidal influx clearances observed from previous transport studies, revealing metabolism as the rate-limiting step. Substitution of the scaled-up transport and metabolic intrinsic clearances into the "well stirred", "parallel-tube", and "dispersion" models predicted higher steady-state extraction ratios for HAHV perfusion. By contrast, integration of the scaled-up in vitro parameters on zonal metabolism and homogeneous transport into a "zonal-compartment" model of three zonal subcompartments (1, 2, and 3) provided an improved description of the extraction ratios during HAPV and HAHV. Zonal factors are important for the scale-up of data in vitro to the whole organ. PMID- 10859156 TI - A mechanistic approach to understanding species differences in felbamate bioactivation: relevance to drug-induced idiosyncratic reactions. AB - In an attempt to understand the species-selective toxicity of felbamate (2-phenyl 1,3-propanediol dicarbamate, FBM), which is thought to result from bioactivation to 2-phenylpropenal, FBM metabolism was evaluated in rats and humans. The formation of 2-phenylpropenal was monitored by the amount of its mercapturates excreted in urine. The data show a relative 5-fold increase in mercapturate excretion in patient urine as a result of differences in metabolism through P450 , esterase-, and aldehyde dehydrogenase-mediated pathways. To compensate for the significant species differences in FBM metabolism, and to produce toxic levels of 2-phenylpropenal in rat comparable to humans levels, monocarbamate felbamate (2 phenyl-1,3-propanediol monocarbamate, MCF), was administered to rats in the hopes of eliciting a toxic response. The desired result, an increase in mercapturate excretion, was not observed in MCF-treated rats due to the identification of a new FBM metabolite, 2-phenyl-1,3-propanediol monocarbamate-alpha-D-glucuronic acid (MCF-glucuronide). Formation of MCF-glucuronide is significant and represents about 80% of MCF metabolites in MCF-dosed rats, 3% of FBM metabolites in FBM-dosed rats, and about 11% of FBM metabolites in FBM patients. To overcome the protective effect of glucuronidation, uridine diphosphoglucuronosyltransferase (UGT)-deficient Gunn rats were treated with FBM and MCF, which surprisingly had no effect on the amount of MCF-glucuronide formed. Given the known UGT polymorphisms and the fact that MCF glucuronidation contributes to the elimination of a 2-phenylpropenal precursor, the correlation between poor UGT activity and an increase in mercapturates excretion was evaluated in patients. The result of the first 34 patients screened suggests that a patient with poor UGT activity is not necessarily at risk for FBM toxicity. PMID- 10859157 TI - Identification and characterization of N-acetylcysteine conjugates of valproic acid in humans and animals. AB - Reactive and hepatotoxic metabolites formed from the biotransformation of valproic acid (VPA) are normally detoxified by conjugating with GSH and followed by mercapturic acid metabolism to produce their respective N-acetylcysteine (NAC) conjugates. Hence, the levels of NAC conjugates of VPA in human urine are an indirect measure of exposure of the liver toward reactive metabolites of the anticonvulsant drug. We report here the synthesis, identification, and characterization of a second NAC conjugate of (E)-2-propyl-2, 4-pentadienoic acid in the urine samples (n = 39) of humans on VPA therapy, namely, (E)-5-(N acetylcystein-S-yl)-2-ene VPA by gas chromatography/mass spectrometry and liquid chromatography with tandem mass spectrometry. In this study, we were able to separate the diastereomers of (E)-5-(N-acetylcystein-S-yl)-3-ene VPA by HPLC. The NAC conjugate of 4,5-epoxy VPA, namely, 5-NAC-4-OH-VPA gamma-lactone, previously identified in rats treated with 2-propyl-4-pentenoic acid (4-ene VPA), was not detected in any of the human urine samples studied. This suggests that in humans, the P-450 metabolism of 4-ene VPA to the reactive epoxide is not a significant pathway. The excretion of the NAC conjugate of (E)-2, 4-diene VPA glucuronide in the urine of seven patients on VPA was also examined and was not detected. The limit of detection of 5-NAC-3-keto VPA and its decarboxylated product, 1-NAC-3 heptanone, was estimated at 25 ng (signal to noise ratio > 3). Neither 5-NAC-3 keto VPA nor 1-NAC-3-heptanone was detected in the urine of patients on VPA therapy or 4-ene VPA-treated guinea pigs, but 1-NAC-3-heptanone was detected in the urine of 4-ene VPA-treated rats. PMID- 10859158 TI - Role of C-5 chiral center in R-(+)-pulegone-mediated hepatotoxicity: metabolic disposition and toxicity of 5, 5-dimethyl-2-(1-Methylethylidene)-cyclohexanone in rats. AB - Metabolic disposition of 5, 5-dimethyl-2-(1-methylethylidene)-cyclohexanone (I) was examined in rats. Compound (I) was administered orally (250 mg/kg of body weight/day) to rats for 5 days. The following urinary metabolites were isolated and identified: 4,5,6,7-tetrahydro-3,6, 6-trimethylbenzofuran (III), 3,3 dimethylcyclohexanone (VI), 5, 5-dimethyl-3-hydroxy-2-(1-methylethylidene) cyclohexanone (X), 5, 5-dimethyl-2-(1-hydroxymethylethyl)-cyclohexanone (IX), 3 hydroxy-5-hydroxymethyl-5-methyl-2-(1-methylethylidene)-cyclo hexano ne (XI), 5,6 dihydro-3,6,6-trimethyl-2(4H)-benzofuranone (VIII), and 5,5-dimethyl-3-hydroxy-2 (1-carboxy ethylidene)-cyclohexanone (XIII). Incubation of compound (I) with phenobarbital (PB)-induced rat liver microsomes in the presence of NADPH resulted in the formation of a metabolite, tentatively identified as a furanoterpene (III) based on proton magnetic resonance, gas chromatography, and gas chromatography mass spectroscopy analyses. The formation of III was inhibited to a significant extent by carbon monoxide, metyrapone, SKF 525-A, and cytochrome c, suggesting the participation of PB-induced microsomal cytochrome P-450 system in the conversion of I to III. Compound I gave type I spectral change in the PB-induced liver microsomes and the dissociation constant (Ks) for I was 38.5 microM. Intraperitoneal administration of a single dose (250 mg/kg) of I to rats resulted in 26, 23, and 41% decreases in the levels of cytochrome P-450, glucose-6 phosphatase, and aminopyrine N-demethylase, respectively, at the end of 24 h. During this period, a 11-fold increase in serum glutamate pyruvate transaminase level was also observed. However, a decrease in the level of cytochrome P-450 and glucose-6-phosphatase, and an increase in serum glutamate pyruvate transaminase values were comparatively more pronounced when R-(+)-pulegone (250 mg/kg) or CCl(4) (0.6 ml/kg) was administered to rats. Pretreatment of rats with PB potentiated the hepatotoxicity caused by I, whereas pretreatment with 3 methylcholanthrene protected from it. This suggests that PB-induced cytochrome P 450-catalyzed reactive metabolites may be responsible for the toxic effects caused by I. PMID- 10859159 TI - Disposition of valproic acid in maternal, fetal, and newborn sheep. I: placental transfer, plasma protein binding, and clearance. AB - Separate 24-h maternal and fetal infusions of valproic acid (VPA) were administered to five pregnant sheep at 125 to 138 days gestation (term approximately 145 days) to determine maternal-fetal disposition. The pharmacokinetics of VPA were also investigated in five newborn 1-day-old lambs after a 6-h drug infusion. Plasma, urine, and amniotic and fetal tracheal fluid samples were analyzed for VPA using gas chromatography-mass spectrometry. During maternal drug infusion, the average steady-state fetal/maternal unbound VPA plasma concentration ratio was 0.81 +/- 0.09. Unbound maternal-to-fetal VPA placental clearance (69.0 +/- 20.2 ml/min/kg) was similar to that in the other direction (61.9 +/- 24.2 ml/min/kg); this indicates passive placental diffusion and intermediate placental permeability of VPA in sheep. Newborn unbound VPA clearance (0.66 +/- 0.28 ml/min/kg) was much lower than in the mother (5.4 +/- 2.7 ml/min/kg) or the fetus (62.1 +/- 22.4 ml/min/kg), and exhibited pronounced Michaelis-Menten characteristics. The elimination half-life of the drug was much longer in the newborn (18.6 +/- 2.6 h) relative to the mother (5.6 +/- 1.4 h) and the fetus (4.6 +/- 1.9 h). Thus, VPA elimination in newborn lambs is much slower as compared with adult sheep, a situation similar to that in humans. Plasma protein binding of VPA was saturable, with similar VPA binding capacities and affinities in maternal and fetal plasma. VPA was extensively displaced from binding sites in the newborn lamb during the first 1 to 2 days of life, possibly because of increased plasma free fatty acid concentrations at birth. Thereafter, newborn plasma appeared to have a similar VPA binding capacity but lower affinity compared with the mother and the fetus. PMID- 10859160 TI - Disposition of valproic acid in maternal, fetal, and newborn sheep. II: metabolism and renal elimination. AB - Metabolism and renal excretion of valproic acid (VPA) were examined in maternal, fetal, and newborn sheep to identify the underlying reasons for the previously observed reduced VPA clearance in newborn lambs. Plasma and urine from VPA infusion studies in maternal, fetal, and newborn sheep were analyzed for VPA and its metabolites [VPA-glucuronide; beta-oxidation products: (E)-2-ene, (E)-3-ene, and 3-keto VPA; hydroxylated metabolites: 3-hydroxy, 4-hydroxy, and 5-hydroxy VPA (5-OH VPA); and 4-ene VPA, 4-keto VPA, 2-propylglutaric acid, and 2 propylsuccinic acid] using gas chromatography-mass spectrometry. All measured metabolites were detectable in maternal and fetal plasma, with 3-keto and 5-OH VPA being at higher concentrations in the fetus. Plasma concentrations of (E)-2 ene, (E)-3-ene, 3-keto, and 5-OH VPA were higher in the newborn compared with the mother, whereas those of the other metabolites were similar. A smaller percentage of the dose was excreted as VPA-glucuronide in newborn lamb urine (28.3 +/- 12.0%) compared with the mother (77.0 +/- 7.8%). Similarly, a lower fraction of the dose was excreted unchanged in newborn urine (11.0 +/- 5.8%) relative to the urine of the mother (19.3 +/- 5.8%); however, significantly larger percentages were excreted as (E)-2-ene (0.11 +/- 0.04 versus 0.02 +/- 0.01%), 3-keto (11.6 +/ 3.5 versus 1. 6 +/- 0.8%), 4-hydroxy (6.1 +/- 3.2 versus 2.3 +/- 1.3%), and 5-OH VPA (2.2 +/- 0.6 versus. 0.8 +/- 0.6%). The major reason for the reduced VPA elimination in newborn lambs appears to be impaired renal excretion and glucuronidation capacity. As a result, a larger fraction of the dose is channeled to beta-oxidation and hydroxylation pathways. The beta-oxidation activities are high at birth; this may explain the high plasma concentrations of (E)-2-ene and 3 keto VPA observed in newborn lambs and human newborns exposed to VPA. PMID- 10859161 TI - RNA polymerase II and the integration of nuclear events. PMID- 10859162 TI - De novo nucleosome assembly: new pieces in an old puzzle. PMID- 10859163 TI - Aberrant cell cycle checkpoint function and early embryonic death in Chk1(-/-) mice. AB - The recent discovery of checkpoint kinases has suggested the conservation of checkpoint mechanisms between yeast and mammals. In yeast, the protein kinase Chk1 is thought to mediate signaling associated with the DNA damage checkpoint of the cell cycle. However, the function of Chk1 in mammals has remained unknown. Targeted disruption of Chk1 in mice showed that Chk1(-/-) embryos exhibit gross morphologic abnormalities in nuclei as early as the blastocyst stage. In culture, Chk1(-/-) blastocysts showed a severe defect in outgrowth of the inner cell mass and died of apoptosis. DNA replication block and DNA damage failed to arrest the cell cycle before initiation of mitosis in Chk1(-/-) embryos. These results may indicate that Chk1 is indispensable for cell proliferation and survival through maintaining the G(2) checkpoint in mammals. PMID- 10859164 TI - Chk1 is an essential kinase that is regulated by Atr and required for the G(2)/M DNA damage checkpoint. AB - Chk1, an evolutionarily conserved protein kinase, has been implicated in cell cycle checkpoint control in lower eukaryotes. By gene disruption, we show that CHK1 deficiency results in a severe proliferation defect and death in embryonic stem (ES) cells, and peri-implantation embryonic lethality in mice. Through analysis of a conditional CHK1-deficient cell line, we demonstrate that ES cells lacking Chk1 have a defective G(2)/M DNA damage checkpoint in response to gamma irradiation (IR). CHK1 heterozygosity modestly enhances the tumorigenesis phenotype of WNT-1 transgenic mice. We show that in human cells, Chk1 is phosphorylated on serine 345 (S345) in response to UV, IR, and hydroxyurea (HU). Overexpression of wild-type Atr enhances, whereas overexpression of the kinase defective mutant Atr inhibits S345 phosphorylation of Chk1 induced by UV treatment. Taken together, these data indicate that Chk1 plays an essential role in the mammalian DNA damage checkpoint, embryonic development, and tumor suppression, and that Atr regulates Chk1. PMID- 10859165 TI - Chaperone hsp27 inhibits translation during heat shock by binding eIF4G and facilitating dissociation of cap-initiation complexes. AB - Inhibition of protein synthesis during heat shock limits accumulation of unfolded proteins that might damage eukaryotic cells. We demonstrate that chaperone Hsp27 is a heat shock-induced inhibitor of cellular protein synthesis. Translation of most mRNAs requires formation of a cap-binding initiation complex known as eIF4F, consisting of factors eIF4E, eIF4A, eIF4E kinase Mnk1, poly(A)-binding protein, and adaptor protein eIF4G. Hsp27 specifically bound eIF4G during heat shock, preventing assembly of the cap-initiation/eIF4F complex and trapping eIF4G in insoluble heat shock granules. eIF4G is a specific target of Hsp27, as eIF4E, eIF4A, Mnk1, poly(A)-binding protein, eIF4B, and eIF3 were not bound by Hsp27 and were not recruited into insoluble complexes. Dissociation of eIF4F was enhanced during heat shock by ectopic overexpression of Hsp25, the murine homolog of human Hsp27. Overexpression of Hsc70, a constitutive homolog of Hsp70, prevented loss of cap-initiation complexes and maintained eIF4G solubility. Purified Hsp27 specifically bound purified eIF4G in vitro, prevented in vitro translation, eliminated eIF4G interaction with protein binding factors, and promoted eIF4G insolubilization. These results therefore demonstrate that Hsp27 is a heat induced inhibitor of eIF4F-dependent mRNA translation. PMID- 10859166 TI - FACKEL is a sterol C-14 reductase required for organized cell division and expansion in Arabidopsis embryogenesis. AB - In flowering plants, the developing embryo consists of growing populations of cells whose fates are determined in a position-dependent manner to form the adult organism. Mutations in the FACKEL (FK) gene affect body organization of the Arabidopsis seedling. We report that FK is required for cell division and expansion and is involved in proper organization of the embryo. We isolated FK by positional cloning. Expression analysis in embryos revealed that FK mRNA becomes localized to meristematic zones. FK encodes a predicted integral membrane protein related to the vertebrate lamin B receptor and sterol reductases across species, including yeast sterol C-14 reductase ERG24. We provide functional evidence that FK encodes a sterol C-14 reductase by complementation of erg24. GC/MS analysis confirmed that fk mutations lead to accumulation of intermediates in the biosynthetic pathway preceding the C-14 reductase step. Although fk represents a sterol biosynthetic mutant, the phenotype was not rescued by feeding with brassinosteroids (BRs), the only plant sterol signaling molecules known so far. We propose that synthesis of sterol signals in addition to BRs is important in mediating regulated cell growth and organization during embryonic development. Our results indicate a novel role for sterols in the embryogenesis of plants. PMID- 10859167 TI - A critical role of sterols in embryonic patterning and meristem programming revealed by the fackel mutants of Arabidopsis thaliana. AB - Here we report a novel Arabidopsis dwarf mutant, fackel-J79, whose adult morphology resembles that of brassinosteroid-deficient mutants but also displays distorted embryos, supernumerary cotyledons, multiple shoot meristems, and stunted roots. We cloned the FACKEL gene and found that it encodes a protein with sequence similarity to both the human sterol reductase family and yeast C-14 sterol reductase and is preferentially expressed in actively growing cells. Biochemical analysis indicates that the fk-J79 mutation results in deficient C-14 sterol reductase activity, abnormal sterol composition, and reduction of brassinosteroids (BRs). Unlike other BR-deficient mutants, the defect of hypocotyl elongation in fk-J79 cannot be corrected by exogenous BRs. The unique phenotypes and sterol composition in fk-J79 indicate crucial roles of sterol regulation and signaling in cell division and cell expansion in embryonic and post-embryonic development in plants. PMID- 10859168 TI - Drosophila SNS, a member of the immunoglobulin superfamily that is essential for myoblast fusion. AB - The Drosophila sticks-and-stones (sns) locus was identified on the basis of its mutant phenotype, the complete absence of body wall muscles and corresponding presence of unfused myoblasts. The genetic location of the mutation responsible for this apparent defect in myoblast fusion was determined by recombination and deficiency mapping, and the corresponding wild-type gene was isolated in a molecular walk. Identification of the SNS coding sequence revealed a putative member of the immunoglobulin superfamily (IgSF) of cell adhesion molecules. As anticipated from this homology, SNS is enriched at the membrane and clusters at discrete sites, coincident with the occurrence of myoblast fusion. Both the sns transcript and the encoded protein are expressed in precursors of the somatic and visceral musculature of the embryo. Within the presumptive somatic musculature, SNS expression is restricted to the putative fusion-competent cells and is not detected in unfused founder cells. Thus, SNS represents the first known marker for this subgroup of myoblasts, and provides an opportunity to identify pathways specifying this cell type as well as transcriptional regulators of fusion specific genes. To these ends, we demonstrate that the presence of SNS-expressing cells is absolutely dependent on Notch, and that expression of SNS does not require the myogenic regulatory protein MEF2. PMID- 10859169 TI - daf-12 encodes a nuclear receptor that regulates the dauer diapause and developmental age in C. elegans. AB - The daf-12 gene acts at the convergence of pathways regulating larval diapause, developmental age, and adult longevity in Caenorhabditis elegans. It encodes a nuclear receptor most closely related to two C. elegans receptors, NHR-8 and NHR 48, Drosophila DHR96, and vertebrate vitamin D and pregnane-X receptors. daf-12 has three predicted protein isoforms, two of which contain DNA- and ligand binding domains, and one of which contains the ligand-binding domain only. Mutations cluster in DNA- and ligand-binding domains, but correspond to distinct phenotypic classes. DAF-12 is expressed widely in target tissues from embryo to adult, but is upregulated during midlarval stages. In the adult, expression persists in nervous system and somatic gonad, two tissues that regulate adult longevity. We propose that DAF-12 integrates hormonal signals in cellular targets to coordinate major life history traits. PMID- 10859170 TI - Xenopus cdc7 function is dependent on licensing but not on XORC, XCdc6, or CDK activity and is required for XCdc45 loading. AB - The assembly and disassembly of protein complexes at replication origins play a crucial role in the regulation of chromosomal DNA replication. The sequential binding of the origin recognition complex (ORC), Cdc6, and the minichromosome maintenance (MCM/P1) proteins produces a licensed replication origin. Before the initiation of replication can occur, each licensed origin must be acted upon by S phase-inducing CDKs and the Cdc7 protein kinase. In the present report we describe the role of Xenopus Cdc7 (XCdc7) in DNA replication using cell-free extracts of Xenopus eggs. We show that XCdc7 binds to chromatin during G(1) and S phase. XCdc7 associates with chromatin only once origins have been licensed, but this association does not require the continued presence of XORC or XCdc6 once they have fulfilled their essential role in licensing. Moreover, XCdc7 is required for the subsequent CDK-dependent loading of XCdc45 but is not required for the destabilization of origins that occurs once licensing is complete. Finally, we show that CDK activity is not necessary for XCdc7 to associate with chromatin, induce MCM/P1 phosphorylation, or perform its essential replicative function. From these results we suggest a simple model for the assembly of functional initiation complexes in the Xenopus system. PMID- 10859171 TI - Rearrangement of chromatin domains during development in Xenopus. AB - A dynamic change in the organization of different gene domains transcribed by RNA polymerase I, II, or III occurs during the progression from quiescent [pre midblastula transition (pre-MBT)] to active (post-MBT) embryos during Xenopus development. In the rDNA, c-myc, and somatic 5S gene domains, a transition from random to specific anchorage to the nuclear matrix occurs when chromatin domains become active. The keratin gene domain was also randomly associated to the nuclear matrix before MBT, whereas a defined attachment site was found in keratinocytes. In agreement with this specification, ligation-mediated (LM)-PCR genomic footprinting carried out on the subpopulation of 5S domains specifically attached to the matrix reveals the hallmarks of determined chromatin after the midblastula transition. In contrast, the same analysis performed on the total 5S gene population does not reveal specific chromatin organization, validating the use of nuclear matrix fractionation to unveil active chromatin domains. These data provide a means for the determination of active chromosomal territories in the embryo and emphasize the role of nuclear architecture in regulated gene expression during development. PMID- 10859172 TI - National Science Foundation-Sponsored Workshop Report: "The 2010 Project" functional genomics and the virtual plant. A blueprint for understanding how plants are built and how to improve them. PMID- 10859173 TI - Genetic repair of mutations in plant cell-free extracts directed by specific chimeric oligonucleotides. AB - Chimeric oligonucleotides are synthetic molecules comprised of RNA and DNA bases assembled in a double hairpin conformation. These molecules have been shown to direct gene conversion events in mammalian cells and animals through a process involving at least one protein from the DNA mismatch repair pathway. The mechanism of action for gene repair in mammalian cells has been partially elucidated through the use of a cell-free extract system. Recent experiments have expanded the utility of chimeric oligonucleotides to plants and have demonstrated genotypic and phenotypic conversion, as well as Mendelian transmission. Although these experiments showed correction of point and frameshift mutations, the biochemical and mechanistic aspects of the process were not addressed. In this paper, we describe the establishment of cell-free extract systems from maize (Zea mays), banana (Musa acuminata cv Rasthali), and tobacco (Nicotiana tabacum). Using a genetic readout system in bacteria and chimeric oligonucleotides designed to direct the conversion of mutations in antibiotic-resistant genes, we demonstrate gene repair of point and frameshift mutations. Whereas extracts from banana and maize catalyzed repair of mutations in a precise fashion, cell-free extracts prepared from tobacco exhibited either partial repair or non-targeted nucleotide conversion. In addition, an all-DNA hairpin molecule also mediated repair albeit in an imprecise fashion in all cell-free extracts tested. This system enables the mechanistic study of gene repair in plants and may facilitate the identification of DNA repair proteins operating in plant cells. PMID- 10859174 TI - Targeting induced local lesions IN genomes (TILLING) for plant functional genomics. PMID- 10859175 TI - Ion changes in legume root hairs responding to Nod factors. PMID- 10859176 TI - Dirigent proteins and dirigent sites explain the mystery of specificity of radical precursor coupling in lignan and lignin biosynthesis. PMID- 10859177 TI - Subcellular localization of expansin mRNA in xylem cells. AB - Terminal differentiation of many vascular cells involves cell wall changes. Cells first elongate their primary wall, then lay down a lignified secondary wall, which is often followed by digestion of the primary wall. Expansins are wall proteins that regulate wall changes, but little is known about the specific functions of the many individual expansin isoforms. An in vitro cell culture of synchronously differentiating tracheary elements was used to identify three new expansins and to compare their expression kinetics with the timing of wall changes. The genes encoding these expansins from zinnia (Zinnia elegans), designated ZeExp1, ZeExp2, ZeExp3, are expressed during cell elongation. ZeExp1 and ZeExp2 mRNA decrease at the early stage of secondary wall formation, whereas ZeExp3 does not. In planta, all three ZeExp mRNAs are found predominantly in a single flank of cells adjacent to protoxylem and metaxylem vessels and in cells roughly at the radial position of the fasicular and interfasicular cambium. Furthermore, within these cells, Exp mRNA is localized exclusively either to the apical or basipetal end of cells depending on the expansin gene and organ, providing the first evidence for polar localization of mRNA in plant cells. ZeExp1 and ZeExp3 mRNA are localized at the apical tip, whereas ZeExp2 mRNA is found in the basal tip. These observations indicate that these three expansins are xylem cell specific and possibly involved in the intrusive growth of the primary walls of differentiating xylem cells. PMID- 10859178 TI - Cell wall and membrane-associated exo-beta-D-glucanases from developing maize seedlings. AB - A beta-D-glucan exohydrolase was purified from the cell walls of developing maize (Zea mays L.) shoots. The cell wall enzyme preferentially hydrolyzes the non reducing terminal glucosyl residue from (1-->3)-beta-D-glucans, but also hydrolyzes (1-->2)-, (1-->6)-, and (1-->4)-beta-D-glucosyl units in decreasing order of activity. Polyclonal antisera raised against the purified exo-beta-D glucanase (ExGase) were used to select partial-length cDNA clones, and the complete sequence of 622 amino acid residues was deduced from the nucleotide sequences of the cDNA and a full-length genomic clone. Northern gel-blot analysis revealed what appeared to be a single transcript, but three distinct polypeptides were detected in immunogel-blot analyses of the ExGases extracted from growing coleoptiles. Two polypeptides appear in the cell wall, where one polypeptide is constitutive, and the second appears at the time of the maximum rate of elongation and reaches peak activity after elongation has ceased. The appearance of the second polypeptide coincides with the disappearance of the mixed-linkage (1-->3), (1-->4)-beta-D-glucan, whose accumulation is associated with cell elongation in grasses. The third polypeptide of the ExGase is an extrinsic protein associated with the exterior surface of the plasma membrane. Although the activity of the membrane-associated ExGase is highest against (1-->3)-beta-D glucans, the activity against (1-->4)-beta-D-glucan linkages is severely attenuated and, therefore, the enzyme is unlikely to be involved with turnover of the (1-->3), (1-->4)-beta-D-glucan. We propose three potential functions for this novel ExGase at the membrane-wall interface. PMID- 10859179 TI - Ozone sensitivity in hybrid poplar correlates with insensitivity to both salicylic acid and jasmonic acid. The role of programmed cell death in lesion formation. AB - Our earlier studies demonstrated that the ozone-sensitive hybrid poplar clone NE 388 displays an attenuated level of ozone-, wound-, and phytopathogen-induced defense gene expression. To determine if this reduced gene activation involves signal transduction pathways dependent on salicylic acid (SA) and/or jasmonic acid (JA), we compared the responses of NE-388 and an ozone-tolerant clone, NE 245, to these signal molecules. JA levels increased in both clones in response to ozone, but only minimal increases in SA levels were measured for either clone. Treatment with SA and methyl jasmonate induced defense gene expression only in NE 245, indicating that NE-388 is insensitive to these signal molecules. DNA fragmentation, an indicator of programmed cell death (PCD), was detected in NE 245 treated with either ozone or an avirulent phytopathogen, but was not detected in NE-388. We conclude that these clones undergo two distinct mechanisms of ozone induced lesion formation. In NE-388, lesions appear to be due to toxic cell death resulting from a limited ability to perceive and subsequently activate SA- and/or JA-mediated antioxidant defense responses. In NE-245, SA-dependent PCD precedes lesion formation via a process related to the PCD pathway activated by phytopathogenic bacteria. These results support the hypothesis that ozone triggers a hypersensitive response. PMID- 10859180 TI - The role of pyruvate dehydrogenase and acetyl-coenzyme A synthetase in fatty acid synthesis in developing Arabidopsis seeds. AB - Acetyl-coenzyme A (acetyl-CoA) formed within the plastid is the precursor for the biosynthesis of fatty acids and, through them, a range of important biomolecules. The source of acetyl-CoA in the plastid is not known, but two enzymes are thought to be involved: acetyl-CoA synthetase and plastidic pyruvate dehydrogenase. To determine the importance of these two enzymes in synthesizing acetyl-CoA during lipid accumulation in developing Arabidopsis seeds, we isolated cDNA clones for acetyl-CoA synthetase and for the ptE1alpha- and ptE1beta-subunits of plastidic pyruvate dehydrogenase. To our knowledge, this is the first reported acetyl-CoA synthetase sequence from a plant source. The Arabidopsis acetyl-CoA synthetase preprotein has a calculated mass of 76,678 D, an apparent plastid targeting sequence, and the mature protein is a monomer of 70 to 72 kD. During silique development, the spatial and temporal patterns of the ptE1beta mRNA level are very similar to those of the mRNAs for the plastidic heteromeric acetyl-CoA carboxylase subunits. The pattern of ptE1beta mRNA accumulation strongly correlates with the formation of lipid within the developing embryo. In contrast, the level of mRNA for acetyl-CoA synthetase does not correlate in time and space with lipid accumulation. The highest level of accumulation of the mRNA for acetyl CoA synthetase during silique development is within the funiculus. These mRNA data suggest a predominant role for plastidic pyruvate dehydrogenase in acetyl CoA formation during lipid synthesis in seeds. PMID- 10859181 TI - The role of the Arabidopsis ELD1 gene in cell development and photomorphogenesis in darkness. AB - Because cell growth and differentiation are regulated by complex interactions among different signaling pathways, a growth defect affects subsequent differentiation. We report on a growth-defective mutant of Arabidopsis, called eld1 (elongation defective 1). Cell elongation was impaired in every organ examined. Later characteristics of the eld1 phenotype include defective vascular tissue differentiation, the inability to grow in soil, ectopic deposition of suberin around twisted vascular bundles, the de-etiolation phenotype, and continuation of shoot development and flowering in the dark. The dwarf phenotype of eld1 could not be rescued by treatment with exogenous growth regulators. Because defective cell elongation is the earliest and most universal feature detected in eld1 mutants, control of or activity in cell elongation may be the primary function of the ELD1 gene. The impaired cell growth results in pleiotropic effects on cell proliferation and differentiation, and the retardation in hypocotyl elongation enables growth and development in darkness. PMID- 10859182 TI - Immunolocalization of a cysteine protease in vacuoles, vesicles, and symbiosomes of pea nodule cells. AB - PsCYP15A is a cysteine protease from pea (Pisum sativum L.). It was first recognized as an up-regulated transcript in wilted shoots and subsequently in root nodules containing Rhizobium. Proteolytic activity of PsCYP15A in nodule extracts is now reported following immunopurification with polyclonal antiserum raised against recombinant antigen. Western-blot analysis indicated two forms of PsCYP15A, a pro-form (approximately 38 kD) and a mature form (approximately 30 kD). Both forms were present in most tissue samples, but only the mature form was isolated from cell-fractionated symbiosomes containing nitrogen-fixing bacteroids. Immunolabeling of nodule sections showed localization of PsCYP15A antigen in large vacuolar bodies, cytoplasmic vesicles, and the perisymbiont space. Immunolabeling of tissue sections from wilted shoots also indicated the presence of PsCYP15A in vacuoles and cytoplasmic vesicles. This protease may be involved in the adaptation to changes in cell turgor, both in wilted shoots and in nodule tissue. Additionally, the protease may be involved in protein turnover in the symbiosome compartment. PMID- 10859183 TI - Cambial-region-specific expression of the Agrobacterium iaa genes in transgenic aspen visualized by a linked uidA reporter gene. AB - The level of indole-3-acetic acid (IAA) was locally modified in cambial tissues of transgenic aspen (Populus tremula L. x Populus tremuloides Michx.). We also demonstrate the use of a linked reporter gene to visualize the expression of the iaa genes. The rate-limiting bacterial IAA-biosynthetic gene iaaM and the reporter gene for beta-glucuronidase (GUS), uidA, were each fused to the cambial region-specific Agrobacterium rhizogenes rolC promoter and linked on the same T DNA. In situ hybridization of the iaaM gene confirmed that histochemical analysis of GUS activity could be used to predict iaaM gene expression. Moreover, quantitative fluorometric analysis of GUS activity allowed estimation of the level of de novo production of IAA in transgenic lines carrying a single-copy insert of the iaaM, uidA T-DNA. Microscale analysis of the IAA concentration across the cambial region tissues showed an increase in IAA concentration of about 35% to 40% in the two transgenic lines, but no changes in the radial distribution pattern of IAA compared with wild-type plants. This increase did not result in any changes in the developmental pattern of cambial derivatives or the cambial growth rate, which emphasizes the importance of the radial distribution pattern of IAA in controlling the development of secondary xylem, and suggests that a moderate increase in IAA concentration does not necessarily stimulate growth. PMID- 10859184 TI - Aluminum accumulation at nuclei of cells in the root tip. Fluorescence detection using lumogallion and confocal laser scanning microscopy. AB - The mechanistic basis for Al toxicity effects on root growth is still a matter of speculation, but it almost certainly involves decreased cell division at the root apex. In this series of experiments, we attempt to determine whether Al enters meristematic cells and binds to nuclei when roots are exposed to a low Al(3+) activity in solution. The methodology involved the use of the Al-sensitive stain lumogallion (3-[2,4 dihydroxyphenylazo]-2-hydroxy-5-chlorobenzene sulfonic acid), the DNA stain 4',6-diamino-phenylindole, and confocal laser scanning microscopy. Soybean (Glycine max L. Merr.) cv Young (Al-sensitive) and PI 416937 (Al tolerant) genotypes were exposed to 1.45 microM Al(3+) for periods ranging from 30 min to 72 h, and then washed with 10 mM citrate to remove apoplastic Al. Fluorescence images show that within 30 min Al entered cells of the sensitive genotype and accumulated at nuclei in the meristematic region of the root tip. Substantial Al also was present at the cell periphery. The images indicated that the Al-tolerant genotype accumulated lower amounts of Al in meristematic and differentiating cells of the root tip and their cell walls. Collectively, the results support an important role for exclusion in Al tolerance. PMID- 10859185 TI - A stress-inducible gene for 9-cis-epoxycarotenoid dioxygenase involved in abscisic acid biosynthesis under water stress in drought-tolerant cowpea. AB - Four cDNA clones named CPRD (cowpea responsive to dehydration) corresponding to genes that are responsive to dehydration were isolated using differential screening of a cDNA library prepared from 10-h dehydrated drought-tolerant cowpea (Vigna unguiculata) plants. One of the cDNA clones has a homology to 9-cis epoxycarotenoid dioxygenase (named VuNCED1), which is supposed to be involved in abscisic acid (ABA) biosynthesis. The GST (glutathione S-transferase)-fused protein indicates a 9-cis-epoxycarotenoid dioxygenase activity, which catalyzes the cleavage of 9-cis-epoxycarotenoid. The N-terminal region of the VuNCED1 protein directed the fused sGFP (synthetic green-fluorescent protein) into the plastids of the protoplasts, indicating that the N-terminal sequence acts as a transit peptide. Both the accumulation of ABA and expression of VuNCED1 were strongly induced by drought stress in the 8-d-old cowpea plant, whereas drought stress did not trigger the expression of VuABA1 (accession no. AB030295) gene that encodes zeaxanthin epoxidase. These results indicate that the VuNCED1 cDNA encodes a 9-cis-epoxycarotenoid dioxygenase and that its product has a key role in the synthesis of ABA under drought stress. PMID- 10859186 TI - AXR2 encodes a member of the Aux/IAA protein family. AB - The dominant gain-of-function axr2-1 mutation of Arabidopsis causes agravitropic root and shoot growth, a short hypocotyl and stem, and auxin-resistant root growth. We have cloned the AXR2 gene using a map-based approach, and find that it is the same as IAA7, a member of the IAA (indole-3-acetic acid) family of auxin inducible genes. The axr2-1 mutation changes a single amino acid in conserved domain II of AXR2/IAA7. We isolated loss-of-function mutations in AXR2/IAA7 as intragenic suppressors of axr2-1 or in a screen for insertion mutations in IAA genes. A null mutant has a slightly longer hypocotyl than wild-type plants, indicating that AXR2/IAA7 controls development in light-grown seedlings, perhaps in concert with other gene products. Dark-grown axr2-1 mutant plants have short hypocotyls and make leaves, suggesting that activation of AXR2/IAA7 is sufficient to induce morphological responses normally elicited by light. Previously described semidominant mutations in two other Arabidopsis IAA genes cause some of the same phenotypes as axr2-1, but also cause distinct phenotypes. These results illustrate functional differences among members of the Arabidopsis IAA gene family. PMID- 10859187 TI - Isolation of Arabidopsis mutants lacking components of acquired thermotolerance. AB - Acquired thermotolerance is a complex physiological phenomenon that enables plants to survive normally lethal temperatures. This study characterizes the temperature sensitivity of Arabidopsis using a chlorophyll accumulation bioassay, describes a procedure for selection of acquired thermotolerance mutants, and provides the physiological characterization of one mutant (AtTS02) isolated by this procedure. Exposure of etiolated Arabidopsis seedlings to 48 degrees C or 50 degrees C for 30 min blocks subsequent chlorophyll accumulation and is eventually lethal. Arabidopsis seedlings can be protected against the effects of a 50 degrees C, 30-min challenge by a 4-h pre-incubation at 38 degrees C. By the use of the milder challenge, 44 degrees C for 30 min, and protective pretreatment, mutants lacking components of the acquired thermotolerance system were isolated. Putative mutants isolated by this procedure exhibited chlorophyll accumulation levels (our measure of acquired thermotolerance) ranging from 10% to 98% of control seedling levels following pre-incubation at 38 degrees C and challenge at 50 degrees C. The induction temperatures for maximum acquired thermotolerance prior to a high temperature challenge were the same in AtTS02 and RLD seedlings, although the absolute level of chlorophyll accumulation was reduced in the mutant. Genetic analysis showed that the loss of acquired thermotolerance in AtTS02 was a recessive trait. The pattern of proteins synthesized at 25 degrees C and 38 degrees C in the RLD and AtTS02 revealed the reduction in the level of a 27-kD heat shock protein in AtTS02. Genetic analysis showed that the reduction of this protein level was correlated with the acquired thermotolerance phenotype. PMID- 10859188 TI - Characterization of auxin conjugates in Arabidopsis. Low steady-state levels of indole-3-acetyl-aspartate, indole-3-acetyl-glutamate, and indole-3-acetyl glucose. AB - Amide-linked indole-3-acetic acid (IAA) conjugates constitute approximately 90% of the IAA pool in the dicot Arabidopsis, whereas ester-linked conjugates and free IAA account for approximately 10% and 1%, respectively when whole seedlings are measured. We show here that IAA-aspartate Asp, IAA-glutamate (Glu), and IAA glucose (Glc) are present at low levels in Arabidopsis. Nine-day-old wild-type Arabidopsis seedlings yielded 17.4 +/- 4.6 ng g(-1) fresh weight IAA-Asp and 3.5 +/- 1.6 ng g(-1) fresh weight IAA-Glu, and IAA-Glc was present at 7 to 17 ng g( 1) fresh weight in 12-d-old wild-type seedlings. Total IAA content in 9-d-old Arabidopsis seedlings was 1, 200 +/- 178 ng g(-1) fresh weight, so these three IAA conjugates together made up only 3% of the conjugate pool throughout the whole plant. We detected less than wild-type levels of IAA-Asp and IAA-Glu (7.8 +/- 0.4 ng g(-1) fresh weight and 1.8 +/- 0.3 ng g(-1) fresh weight, respectively) in an Arabidopsis mutant that accumulates conjugated IAA. Our results are consistent with IAA-Asp, IAA-Glu, and IAA-Glc being either minor, transient, or specifically localized IAA metabolites under normal growth conditions and bring into question the physiological relevance of IAA-Asp accumulation in response to high concentrations of exogenous IAA. PMID- 10859189 TI - Cucumber mosaic virus infection affects sugar transport in melon plants. AB - Viral infection often affects carbon assimilation and metabolism in host plants. To better understand the effect of cucumber mosaic virus (CMV) infection on sugar transport, carbohydrate levels and the amounts of the various sugars in the phloem sap were determined in infected melon (Cucumis melo L.) plants. Source leaves infected with CMV were characterized by high concentrations of reducing sugars and relatively low starch levels. The altered level of carbohydrates was accompanied by increased respiration and decreased net photosynthetic rates in the infected leaves. Although stachyose was the predominant sugar in phloem sap collected from petioles of control leaves, sucrose (Suc) was a major sugar in the phloem sap of infected leaves. Moreover, analyses of the newly fixed (14)CO(2) revealed a high proportion of radioactive Suc in the phloem sap of infected leaves 60 min post-labeling. The alteration in phloem sap sugar composition was found in source, but not old, leaves. Moreover, elevations in Suc concentration were also evident in source leaves that did not exhibit symptoms or contain detectable amounts of virus particles. The mode by which CMV infection may cause alterations in sugar transport is discussed in terms of the mechanism by which sugars are loaded into the phloem of cucurbit plants. PMID- 10859190 TI - The mechanic state of "inner tissue" in the growing zone of sunflower hypocotyls and the regulation of its growth rate following excision. AB - Spontaneous growth of isolated inner tissue from the etiolated sunflower (Helianthus annuus L.) hypocotyl growing zone was investigated. A new preparation technique allowed measurements starting 3 s after excision. Elongation with respect to the turgescent and plasmolized state was quantified in terms of relative growth rates, facilitating comparison to growth in situ. Turgor and turgor-induced strain were determined. Overall longitudinal strain in inner tissues in situ was positive, indicating that compressive forces exerted by peripheral tissues are outweighed by turgor-dependent tensile stress. Inner tissue expansion following isolation depended on water uptake. Extreme plastic extension rates occurred immediately after excision, suggesting that mechanical parameters of inner tissue in situ cannot be extrapolated from the mechanics of excised sections. In the long term, excised inner tissue autonomously established values of turgor, turgor-induced strain, and relative growth rates similar to values in the living plant. These results support historic models of tissue cooperation during organ growth, in which inner tissues actively participate in the control of growth rates. PMID- 10859191 TI - The structure and expression of the wheat starch synthase III gene. Motifs in the expressed gene define the lineage of the starch synthase III gene family. AB - The endosperm of hexaploid wheat (Triticum aestivum [L.]) was shown to contain a high molecular weight starch synthase (SS) analogous to the product of the maize du1 gene, starch synthase III (SSIII; DU1). cDNA and genomic DNA sequences encoding wheat SSIII were isolated and characterized. The wheat SSIII cDNA is 5,346 bp long and contains an open reading frame that encodes a 1,628-amino acid polypeptide. A putative N-terminal transit peptide, a 436-amino acid C-terminal catalytic domain, and a central 470-amino acid SSIII-specific domain containing three regions of repeated amino acid similarity were identified in the wheat gene. A fourth region between the transit peptide and the SSIII-specific domain contains repeat motifs that are variable with respect to motif sequence and repeat number between wheat and maize. In dicots, this N-terminal region does not contain repeat motifs and is truncated. The gene encoding wheat SSIII, designated ss3, consists of 16 exons extending over 10 kb, and is located on wheat chromosome I. Expression of ss3 mRNA in wheat was detected in leaves, pre anthesis florets, and from very early to middle stage of endosperm development. The entire N-terminal variable repeat region and the majority of the SSIII specific domain are encoded on a single 2,703-bp exon. A gene encoding a class III SS from the Arabidopsis genome sequencing project shows a strongly conserved exon structure to the wheat ss3 gene, with the exception of the N-terminal region. The evolutionary relationships of the genes encoding monocot and dicot class III SSs are discussed. PMID- 10859192 TI - The photoreduction of H(2)O(2) by Synechococcus sp. PCC 7942 and UTEX 625. AB - It has been claimed that the sole H(2)O(2)-scavenging system in the cyanobacterium Synechococcus sp. PCC 7942 is a cytosolic catalase-peroxidase. We have measured in vivo activity of a light-dependent peroxidase in Synechococcus sp. PCC 7942 and UTEX 625. The addition of small amounts of H(2)O(2) (2.5 microM) to illuminated cells caused photochemical quenching (qP) of chlorophyll fluorescence that was relieved as the H(2)O(2) was consumed. The qP was maximal at about 50 microM H(2)O(2) with a Michaelis constant of about 7 microM. The H(2)O(2)-dependent qP strongly indicates that photoreduction can be involved in H(2)O(2) decomposition. Catalase-peroxidase activity was found to be almost completely inhibited by 10 microM NH(2)OH with no inhibition of the H(2)O(2) dependent qP, which actually increased, presumably due to the light-dependent reaction now being the only route for H(2)O(2)-decomposition. When (18)O-labeled H(2)O(2) was presented to cells in the light there was an evolution of (16)O(2), indicative of H(2)(16)O oxidation by PS 2 and formation of photoreductant. In the dark (18)O(2) was evolved from added H(2)(18)O(2) as expected for decomposition by the catalase-peroxidase. This evolution was completely blocked by NH(2)OH, whereas the light-dependent evolution of (16)O(2) during H(2)(18)O(2) decomposition was unaffected. PMID- 10859193 TI - Accumulation of palmitate in Arabidopsis mediated by the acyl-acyl carrier protein thioesterase FATB1. AB - The acyl-acyl carrier protein thioesterase B1 from Arabidopsis (AtFATB1) was previously shown to exhibit in vitro hydrolytic activity for long chain acyl-acyl carrier proteins (P. Dormann, T.A. Voelker, J.B. Ohlrogge [1995] Arch Biochem Biophys 316: 612-618). In this study, we address the question of which role in fatty acid biosynthesis this enzyme plays within the plant. Over-expression of the AtFATB1 cDNA under a seed-specific promoter resulted in accumulation of high amounts of palmitate (16:0) in seeds. RNA and protein-blot analysis in Arabidopsis and rapeseed (Brassica napus) showed that the endogenous AtFATB1 expression was highest in flowers and lower in leaves. All floral tissues of wild type plants contained elevated amounts of 16:0, and in the polar lipid fraction of flowers close to 50 mol % of the fatty acids were 16:0. Therefore, flowers contain polar lipids with an unusually high amount of saturated fatty acids as compared to all other plant tissues. Antisense expression of the AtFATB1 cDNA under the cauliflower mosaic virus 35S promoter resulted in a reduction of seed and flower 16:0 content, but no changes in leaf fatty acids. We conclude that the AtFATB1 thioesterase contributes to 16:0 production particularly in flowers, but that additional factors are involved in leaves. PMID- 10859194 TI - Calcium-calmodulin suppresses the filamentous actin-binding activity of a 135 kilodalton actin-bundling protein isolated from lily pollen tubes. AB - We have isolated a 135-kD actin-bundling protein (P-135-ABP) from lily (Lilium longiflorum) pollen tubes and have shown that this protein is responsible for bundling actin filaments in lily pollen tubes (E. Yokota, K. Takahara, T. Shimmen [1998] Plant Physiol 116: 1421-1429). However, only a few thin actin-filament bundles are present in random orientation in the tip region of pollen tubes, where high concentrations of Ca(2+) have also been found. To elucidate the molecular mechanism for the temporal and spatial regulation of actin-filament organization in the tip region of pollen tubes, we explored the possible presence of factors modulating the filamentous actin (F-actin)-binding activity of P-135 ABP. The F-actin-binding activity of P-135-ABP in vitro was appreciably reduced by Ca(2+) and calmodulin (CaM), although neither Ca(2+) alone nor CaM in the presence of low concentrations of Ca(2+) affects the activity of P-135-ABP. A micromolar order of Ca(2+) and CaM were needed to induce the inhibition of the binding activity of P-135-ABP to F-actin. An antagonist for CaM, W-7, cancelled this inhibition. W-5 also alleviated the inhibition effect of Ca(2+)-CaM, however, more weakly than W-7. These results suggest the specific interaction of P-135-ABP with Ca(2+)-CaM. In the presence of both Ca(2+) and CaM, P-135-ABP organized F-actin into thin bundles, instead of the thick bundles observed in the absence of CaM. These results suggest that the inhibition of the P-135-ABP activity by Ca(2+)-CaM is an important regulatory mechanism for organizing actin filaments in the tip region of lily pollen tubes. PMID- 10859195 TI - Purification and characterization of bifunctional lysine-ketoglutarate reductase/saccharopine dehydrogenase from developing soybean seeds. AB - Both in mammals and plants, excess lysine (Lys) is catabolized via saccharopine into alpha-amino adipic semialdehyde and glutamate by two consecutive enzymes, Lys-ketoglutarate reductase (LKR) and saccharopine dehydrogenase (SDH), which are linked on a single bifunctional polypeptide. To study the control of metabolite flux via this bifunctional enzyme, we have purified it from developing soybean (Glycine max) seeds. LKR activity of the bifunctional LKR/SDH possessed relatively high K(m) for its substrates, Lys and alpha-ketoglutarate, suggesting that this activity may serve as a rate-limiting step in Lys catabolism. Despite their linkage, the LKR and SDH enzymes possessed significantly different pH optima, suggesting that SDH activity of the bifunctional enzyme may also be rate limiting in vivo. We have previously shown that Arabidopsis plants contain both a bifunctional LKR/SDH and a monofunctional SDH enzymes (G. Tang, D. Miron, J.X. Zhu-Shimoni, G. Galili [1997] Plant Cell 9: 1-13). In the present study, we found no evidence for the presence of such a monofunctional SDH enzyme in soybean seeds. These results may provide a plausible regulatory explanation as to why various plant species accumulate different catabolic products of Lys. PMID- 10859196 TI - Kinetics of constant gravitropic stimulus responses in Arabidopsis roots using a feedback system. AB - The study of gravitropism is hindered by the fact that as a root responds, the gravitational stimulus changes. Using a feedback system to connect a rotating stage platform to a video digitizer system, we were able to maintain a constant angle of gravistimulation to Arabidopsis roots for long time periods. The rate of curvature approximated the sine rule for angles of stimulation between 20 degrees and 120 degrees. For a given angle of stimulation, the rate of curvature also remained constant, with no observed diminishment of the response. Although previous reports of Arabidopsis root gravitropism suggest latent periods of approximately 30 min, using a smooth mechanical stage to reorient the root, we observed a mean time lag of approximately 10 min. This more rapid onset of curvature can, in part, be explained by reduced mechanical perturbation during the process of gravistimulation. This suggests that mechanical stimulation associated with rapid root re-orientation may confound investigations of early gravitropic events. PMID- 10859197 TI - Variation in the oxygen isotope ratio of phloem sap sucrose from castor bean. Evidence in support of the Peclet effect. AB - Theory suggests that the level of enrichment of (18)O above source water in plant organic material (Delta) may provide an integrative indicator of control of water loss. However, there are still gaps in our understanding of the processes affecting Delta. One such gap is the observed discrepancy between modeled enrichment of water at the sites of evaporation within the leaf and measured enrichment of the leaf water as a whole (Delta(L)). Farquhar and Lloyd (1993) suggested that this may be caused by a Peclet effect. It is also unclear whether organic material formed in the leaf reflects enrichment of water at the sites of evaporation within the leaf or Delta(L). To investigate this question castor bean (Ricinus communis L.) leaves, still attached to the plant, were sealed into a controlled-environment gas exchange chamber and subjected to a step change in leaf-to-air vapor pressure difference. Sucrose was collected from a cut on the petiole of the leaf in the chamber under equilibrium conditions and every hour for 6 h after the change in leaf-to-air vapor pressure difference. Oxygen isotope composition of sucrose in the phloem sap (Delta(suc)) reflected modeled Delta(L). A model is presented describing Delta(suc) at isotopic steady state, and accounts for 96% of variation in measured Delta(suc). The data strongly support the Peclet effect theory. PMID- 10859198 TI - A possible role for pyrophosphate in the coordination of cytosolic and plastidial carbon metabolism within the potato tuber. AB - The early stages of tuber development are characterized by cell division, high metabolic activity, and the predominance of invertase as the sucrose (Suc) cleaving activity. However, during the subsequent phase of starch accumulation the cleavage of Suc occurs primarily by the action of Suc synthase. The mechanism that is responsible for this switch in Suc cleaving activities is currently unknown. One striking difference between the invertase and Suc synthase mediated cleavage of Suc is the direct involvement of inorganic pyrophosphate (PPi) in the latter case. There is presently no convincing explanation of how the PPi required to support this process is generated in potato (Solanum tuberosum) tubers. The major site of PPi production in a maturing potato tubers is likely to be the reaction catalyzed by ADP-glucose pyrophosphorylase, the first committed step of starch biosynthesis in amyloplasts. We present data based on the analysis of the PPi levels in various transgenic plants altered in starch and Suc metabolism that support the hypothesis that PPi produced in the plastid is used to support cytosolic Suc breakdown and that PPi is an important coordinator of cytosolic and plastidial metabolism in potato tubers. PMID- 10859199 TI - Auxin inhibition of decapitation-induced branching is dependent on graft transmissible signals regulated by genes Rms1 and Rms2. AB - Decapitation-induced axillary bud outgrowth is a vital mechanism whereby shoots are able to continue normal growth and development. In many plants, including wild-type garden pea (Pisum sativum L.), this process can be inhibited by exogenous auxin. Using the ramosus (rms) increased branching mutants of pea, we present evidence that this response to auxin is dependent on graft-transmissible substance(s) regulated by the genes Rms1 and Rms2. The response to exogenous auxin is massively diminished in decapitated rms1 and rms2 mutant plants. However, basipetal auxin transport is not reduced in intact or decapitated mutants. Grafting rms1 or rms2 shoots onto wild-type rootstocks restored the auxin response, indicating that Rms1 and Rms2 gene action in the rootstock is sufficient to enable an auxin response in mutant shoots. We conclude that Rms1 and Rms2 act in the rootstock and shoot to control levels of mobile substance(s) that interact with exogenous auxin in the inhibition of bud outgrowth after decapitation. At least for rms1, the reduced auxin response is unlikely to be due to an inability of auxin to decrease xylem sap cytokinin content, as this is already low in intact rms1 plants. Consequently, we have genetic evidence that auxin action in decapitated plants depends on at least one novel long-distance signal. PMID- 10859200 TI - Isolation and characterization of cDNAs expressed in the early stages of flavonol induced pollen germination in petunia. AB - Petunia (Petunia hybrida) pollen requires flavonols (Fl) to germinate. Adding kaempferol to Fl-deficient pollen causes rapid and synchronous germination and tube outgrowth. We exploited this system to identify genes responsive to Fls and to examine the changes in gene expression that occur during the first 0.5 h of pollen germination. We used a subtracted library and differential screening to identify 22 petunia germinating pollen clones. All but two were expressed exclusively in pollen and half of the clones were rare or low abundance cDNAs. RNA gel-blot analysis showed that the steady-state transcript levels of all the clones were increased in response to kaempferol. The sequences showing the greatest response to kaempferol encode proteins that have regulatory or signaling functions and include S/D4, a leucine-rich repeat protein, S/D1, a LIM-domain protein, and D14, a putative Zn finger protein with a heme-binding site. Eight of the clones were novel including S/D10, a cDNA only expressed very late in pollen development and highly up-regulated during the first 0.5 h of germination. The translation product of the S/D3 cDNA shares some features with a neuropeptide that regulates guidance and growth in the tips of extending axons. This study confirmed that the bulk of pollen mRNA accumulates well before germination, but that specific sequences are transcribed during the earliest moments of Fl-induced pollen germination. PMID- 10859201 TI - Cytochrome P450-dependent metabolism of oxylipins in tomato. Cloning and expression of allene oxide synthase and fatty acid hydroperoxide lyase. AB - Allene oxide synthase (AOS) and fatty acid hydroperoxide lyase (HPL) are plant specific cytochrome P450s that commit fatty acid hydroperoxides to different branches of oxylipin metabolism. Here we report the cloning and characterization of AOS (LeAOS) and HPL (LeHPL) cDNAs from tomato (Lycopersicon esculentum). Functional expression of the cDNAs in Escherichia coli showed that LeAOS and LeHPL encode enzymes that metabolize 13- but not 9-hydroperoxide derivatives of C(18) fatty acids. LeAOS was active against both 13S-hydroperoxy-9(Z),11(E),15(Z) octadecatrienoic acid (13-HPOT) and 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid, whereas LeHPL showed a strong preference for 13-HPOT. These results suggest a role for LeAOS and LeHPL in the metabolism of 13-HPOT to jasmonic acid and hexenal/traumatin, respectively. LeAOS expression was detected in all organs of the plant. In contrast, LeHPL expression was predominant in leaves and flowers. Damage inflicted to leaves by chewing insect larvae led to an increase in the local and systemic expression of both genes, with LeAOS showing the strongest induction. Wound-induced expression of LeAOS also occurred in the def-1 mutant that is deficient in octadecanoid-based signaling of defensive proteinase inhibitor genes. These results demonstrate that tomato uses genetically distinct signaling pathways for the regulation of different classes of wound responsive genes. PMID- 10859202 TI - Inhibition of plant asparagine synthetase by monoterpene cineoles. AB - Asparagine (Asn) synthetase (AS) is the key enzyme in Asn biosynthesis and plays an important role in nitrogen mobilization. Despite its important physiological function, little research has been done documenting inhibitors of plant AS. Plant growth inhibition caused by the natural monoterpene 1,4-cineole and its structurally related herbicide cinmethylin was reversed 65% and 55%, respectively, by providing 100 microM Asn exogenously. Reversion of the phytotoxic effect was dependent on the concentration of Asn. The presence of either 1,4-cineole or cinmethylin stimulated root uptake of [(14)C]Asn by lettuce (Lactuca sativa) seedlings. Although the physiological responses suggested that both compounds affected Asn biosynthesis, biochemical analysis of AS activity showed that the natural monoterpene was a potent inhibitor (I(50) = approximately 0. 5 microM) of the enzyme, whereas the commercial product was not inhibitory up to levels of 10 mM. Analysis of the putative metabolite, 2-hydroxy-1,4-cineole, showed that the cis-enantiomer was much more active than the trans-enantiomer, suggesting that the hydroxyl group was involved in the specific ligand/active site interaction. This is the first report that AS is a suitable herbicide target site, and that cinmethylin is apparently a proherbicide that requires metabolic bioactivation via cleavage of the benzyl-ether side chain. PMID- 10859203 TI - ACX3, a novel medium-chain acyl-coenzyme A oxidase from Arabidopsis. AB - In a database search for homologs of acyl-coenzyme A oxidases (ACX) in Arabidopsis, we identified a partial genomic sequence encoding an apparently novel member of this gene family. Using this sequence information we then isolated the corresponding full-length cDNA from etiolated Arabidopsis cotyledons and have characterized the encoded recombinant protein. The polypeptide contains 675 amino acids. The 34 residues at the amino terminus have sequence similarity to the peroxisomal targeting signal 2 of glyoxysomal proteins, including the R [I/Q/L]-X5-HL-XL-X15-22-C consensus sequence, suggesting a possible microsomal localization. Affinity purification of the encoded recombinant protein expressed in Escherichia coli followed by enzymatic assay, showed that this enzyme is active on C8:0- to C14:0-coenzyme A with maximal activity on C12:0-coenzyme A, indicating that it has medium-chain-specific activity. These data indicate that the protein reported here is different from previously characterized classes of ACX1, ACX2, and short-chain ACX (SACX), both in sequence and substrate chain length specificity profile. We therefore, designate this new gene AtACX3. The temporal and spatial expression patterns of AtACX3 during development and in various tissues were similar to those of the AtSACX and other genes expressed in glyoxysomes. Currently available database information indicates that AtACX3 is present as a single copy gene. PMID- 10859204 TI - Recombinant expression of molybdenum reductase fragments of plant nitrate reductase at high levels in Pichia pastoris. AB - Mo reductase (MoR; formerly cytochrome c reductase) fragments of NADH:NO(3) reductase (NR; EC1.6.6.1) were cytosolically expressed in Pichia pastoris, a methylotrophic yeast, using spinach (Spinacia oleracea) and corn (Zea maize) cDNAs. In fermenter cultures, spinach MoR was expressed at 420 mg L(-1), corn MoR at 32 mg L(-1), and corn MoR plus with putative NR interface domain N terminus (MoR+) at 17 mg L(-1). Constitutively expressed MoR+ was structurally stable while it was degraded when expressed by methanol induction, which suggests methanol growth produces more proteinase. Methanol-induced expression yielded more target protein. All three MoR were purified to homogeneity and their polypeptides were approximately 41 (MoR) and approximately 66 (MoR+) kD. MoR was monomeric and MoR+ dimeric, confirming the predicted role for dimer interface domain of NR. MoR+, although differing in quaternary structure from MoR, has similar kinetic properties for ferricyanide and cytochrome c reductase activities and visible spectra, which were like NR. Redox potentials of MoR and MoR+ were similar for flavin, whereas MoR+ had a more negative potential for heme-iron. Reaction schemes for MoR catalyzed reactions were proposed based on fast-reaction rapid-scan stopped-flow kinetic analysis of MoR. P. pastoris is an excellent system for producing the large amounts of NR fragments needed for detailed biochemical studies. PMID- 10859205 TI - Deletion of the nitrate reductase N-terminal domain still allows binding of 14-3 3 proteins but affects their inhibitory properties. AB - Nitrate reductase (NR) is post-translationally regulated by phosphorylation and binding of 14-3-3 proteins. Deletion of 56 amino acids in the amino-terminal domain of NR was previously shown to impair this type of regulation in tobacco (Nicotiana plumbaginifolia) (L. Nussaume, M. Vincentez, C. Meyer, J.-P. Boutin, M. Caboche [1995] Plant Cell 7: 611-621), although both full-length NR and deleted NR (DeltaNR) appeared to be phosphorylated in darkness (C. Lillo, S. Kazazaic, P. Ruoff, C. Meyer [1997] Plant Physiol 114: 1377-1383). We show here that in the presence of Mg(2+) and phosphatase inhibitors, NR and endogenous 14-3 3 proteins copurify through affinity chromatography. Assay of NR activity and western blots showed that endogenous 14-3-3 proteins copurified with both NR and DeltaNR. Electron transport in the heme-binding domain of DeltaNR was inhibited by Mg(2+)/14-3-3, whereas this was not the case for NR. This may indicate a different way of binding for 14-3-3 in the DeltaNR compared with NR. The DeltaNR was more labile than NR, in vitro. Lability was ascribed to the molybdopterin binding domain, and apparently an important function of the 56 amino acids is stabilization of this domain. PMID- 10859206 TI - Interaction between gravitropism and phototropism in sporangiophores of Phycomyces blakesleeanus. AB - The interaction between gravitropism and phototropism was analyzed for sporangiophores of Phycomyces blakesleeanus. Fluence rate-response curves for phototropism were generated under three different conditions: (a) for stationary sporangiophores, which reached photogravitropic equilibrium; (b) for sporangiophores, which were clinostated head-over during phototropic stimulation; and (c) for sporangiophores, which were subjected to centrifugal accelerations of 2.3g to 8.4g. For blue light (454 nm), clinostating caused an increase of the slope of the fluence rate-response curves and an increase of the maximal bending angles at saturating fluence rates. The absolute threshold remained, however, practically unaffected. In contrast to the results obtained with blue light, no increase of the slope of the fluence rate-response curves was obtained with near ultraviolet light at 369 nm. Bilateral irradiation with near-ultraviolet or blue light enhanced gravitropism, whereas symmetric gravitropic stimulation caused a partial suppression of phototropism. Gravitropism and phototropism appear to be tightly linked by a tonic feedback loop that allows the respective transduction chains a mutual influence over each other. The use of tropism mutants allowed conclusions to be drawn about the tonic feedback loop with the gravitropic and phototropic transduction chains. The results from clinostating mutants that lack octahedral crystals (implicated as statoliths) showed that these crystals are not involved in the tonic feedback loop. At elevated centrifugal accelerations, the fluence-rate-response curves for photogravitropic equilibrium were displaced to higher fluence rates and the slope decreased. The results indicate that light transduction possesses a logarithmic transducer, whereas gravi-transduction uses a linear one. PMID- 10859207 TI - Hypersensitivity of an Arabidopsis sugar signaling mutant toward exogenous proline application. AB - In transgenic Arabidopsis a patatin class I promoter from potato is regulated by sugars and proline (Pro), thus integrating signals derived from carbon and nitrogen metabolism. In both cases a signaling cascade involving protein phosphatases is involved in induction. Other endogenous genes are also regulated by both Pro and carbohydrates. Chalcone synthase (CHS) gene expression is induced by both, whereas the Pro biosynthetic Delta(1)-pyrroline-5-carboxylate synthetase (P5CS) is induced by high Suc concentrations but repressed by Pro, and Pro dehydrogenase (ProDH) is inversely regulated. The mutant rsr1-1, impaired in sugar dependent induction of the patatin promoter, is hypersensitive to low levels of external Pro and develops autofluorescence and necroses. Toxicity of Pro can be ameliorated by salt stress and exogenously supplied metabolizable carbohydrates. The rsr1-1 mutant shows a reduced response regarding sugar induction of CHS and P5CS expression. ProDH expression is de-repressed in the mutant but still down-regulated by sugar. Pro toxicity seems to be mediated by the degradation intermediate Delta(1)-pyrroline-5-carboxylate. Induction of the patatin promoter by carbohydrates and Pro, together with the Pro hypersensitivity of the mutant rsr1-1, demonstrate a new link between carbon/nitrogen and stress responses. PMID- 10859208 TI - False-negative results in screening programmes: systematic review of impact and implications. AB - BACKGROUND: When assessing whether a screening programme is appropriate, there is a particular obligation to ensure that the harms as well as the benefits are considered. Among these harms is the likelihood that false-negative results will occur. In some cases, the consequences of these can be difficult to assess, although false reassurance leading to diagnostic delay and subsequent treatment has been suggested. However, no test is totally accurate (with 100% sensitivity and specificity), and false-negative results are inherent in any screening programme that does not have 100% sensitivity. This review was carried out to assess the medical, psychological, economic and legal consequences of false negative results that occur in national screening programmes. OBJECTIVES: (1) to determine the consequences of false-negative findings; (2) to investigate how their adverse effects can be minimised; to assess their implications for the NHS, including the impact of false-negatives on public confidence in screening programmes; to identify relevant theoretical perspectives that may be potentially useful when considering the implications of false-negative results. METHODS: A systematic literature review was carried out. This included a search of 18 electronic databases, various bibliographies and contact with experts to identify relevant literature and perspectives. Outcomes included in the review fell into four categories: medical outcomes (morbidity and mortality); psychological outcomes (distress, false reassurance, loss of confidence in services); economic outcomes (such as costs to the NHS); legal outcomes (such as litigation). Other outcomes, such as the impact of false-negatives on public confidence in screening programmes, were also included. The participants included individuals taking part in screening programmes, healthcare professionals and organisations responsible for screening programmes. Methodological details of the review are provided in the full report. RESULTS: A total of 6660 abstracts were screened, and 420 potentially relevant papers were identified. Most of the studies that were identified presented only anecdotal evidence. (1) Medical outcomes: In all, 13 papers presented quantitative information relevant to the medical consequences of false-negative results; seven of these were primary studies, and the remaining studies were literature reviews or models examining the likely impact of false negative results. (2) Psychological outcomes: A total of eight published studies presented information on the psychological consequences of negative results in general; only one study, on antenatal screening, provided direct evidence of the psychological consequences of false-negative results, where they were associated with lower parental acceptance of the affected child and with blaming others for this outcome. (3) Economic outcomes: Only two studies presented information on the economic consequences. The strength of evidence from most of the primary studies was low. There is some evidence that false-negative results may have a large legal impact. For example, in cervical screening they have led to legal action and its associated costs, including payment of compensation; this is based on reports of events in both the UK and US health systems. There also seems to be a consensus in the literature that false-negatives may have a negative impact on public confidence in screening; evidence is again limited however. CONCLUSIONS: False-negatives are evident in all screening programmes, even when the quality of the service provided is high. They may have the potential to delay the detection of breast and cervical cancer, but there is little evidence to help assess their psychological consequences in these or other screening programmes. False negatives are likely to lead to legal action being taken by those individuals affected, and potentially may reduce public confidence in screening. (ABSTRACT TRUNCATED) PMID- 10859209 TI - The accuracy of statistical methods for estimation of haplotype frequencies: an example from the CD4 locus. AB - Haplotype analysis has become increasingly important for the study of human disease as well as for reconstruction of human population histories. Computer programs have been developed to estimate haplotype frequencies statistically from marker phenotypes in unrelated individuals. However, there currently are few empirical reports on the accuracy of statistical estimates that must infer linkage phase. We have analyzed haplotypes at the CD4 locus on chromosome 12 that consist of a short tandem-repeat polymorphism and an Alu insertion/deletion polymorphism located 9.8 kb apart, in 398 individuals from 10 geographically diverse sub-Saharan African populations. Haplotype frequency estimates obtained using gene counting based on molecularly haplotyped (phase-known) data were compared with haplotype frequency estimates obtained using the expectation maximization algorithm. We show that the estimated frequencies of common haplotypes do not differ significantly with the use of phase-known versus phase unknown data. However, rare haplotypes are occasionally miscalled when their presence/absence must be inferred. Thus, for those research questions for which the common haplotypes are most important, frequency estimates based on the phase unknown marker-typing results from unrelated individuals will be sufficient. However, in cases where knowledge of rare haplotypes is critical, molecular haplotyping will be necessary to determine linkage phase unambiguously. PMID- 10859210 TI - Lessons from genetically engineered animal models. XII. IL-10-deficient (IL-10(-/ ) mice and intestinal inflammation. AB - Interleukin (IL)-10(-/-) mice spontaneously develop intestinal inflammation characterized by discontinuous transmural lesions affecting the small and large intestine and by dysregulated production of proinflammatory cytokines. The uncontrolled generation of IFN-gamma-producing CD4(+) T cells (Th1 type) has been shown to play a causal role in the development of enterocolitis affecting these mutants. This article discusses studies of IL-10(-/-) mice that have investigated the role of enteric organisms in triggering intestinal disease, the mediators responsible for initiating and maintaining intestinal disease, the role IL-10 plays in the generation and/or function of regulatory cells, and the results of IL-10 therapy in experimental animal models of inflammatory bowel disease (IBD) and human patients with IBD. PMID- 10859211 TI - Pathobiology of visceral pain: molecular mechanisms and therapeutic implications IV. Visceral afferent contributions to the pathobiology of visceral pain. AB - Functional bowel and other visceral disorders exhibit multiple characteristics that suggest the presence of visceral hyperalgesia. The discomfort, pain, and altered sensations (e.g., to intraluminal contents) that define the hyperalgesia typically arise in the absence of tissue insult or inflammation. Visceral hyperalgesia thus differs from somatic hyperalgesia, which is commonly associated with tissue injury and inflammation. Hyperalgesia could develop and be maintained by either peripheral or central mechanisms; the altered sensations associated with functional visceral disorders are contributed to by both peripheral and central mechanisms. The relative contributions of peripheral and central mechanisms are not well understood, and the focus in this Themes article is on potential peripheral contributions: sensitization of visceral receptors, nerve injury, and ion channels. PMID- 10859212 TI - Resistance to apoptosis is a mechanism of adaptation of rat stomach to aspirin. AB - Adaptation of the gastric mucosa to nonsteroidal anti-inflammatory drug-induced injury is a well-documented phenomenon, but the mechanisms are not known. We investigated whether changes in stress protein expression and apoptosis play roles in adaptation of rat stomach to aspirin. RT-PCR and Western blotting techniques were used to analyze mRNA and protein expression of HSP72 and HSP90 and cleavage of caspase 3 protein. Apoptosis was detected by the TUNEL method and quantified. HSP72 mRNA and protein expression was unchanged in adapted mucosa, whereas HSP90 mRNA and protein levels decreased. Caspase 3 protein was activated, and the number of apoptotic cells increased in mucosa after one aspirin dose. However, in adapted mucosa after aspirin, activated caspase 3 and the number of apoptotic cells had returned to basal levels. Induction of the stress response was found not to be a mechanism of mucosal adaptation against multiple doses of aspirin. Our results lead us to propose instead that resistance to aspirin induced apoptosis plays a role in the protective phenomenon of adaptation. PMID- 10859213 TI - Chronic stress impairs rat growth and jejunal epithelial barrier function: role of mast cells. AB - We examined the impact of chronic stress on rat growth rate and intestinal epithelial physiology and the role of mast cells in these responses. Mast cell deficient (Ws/Ws) rats and +/+ littermate controls were submitted to water avoidance stress or sham stress, 1 h/day, for 5 days. Seven hours after the last sham or stress session, jejunal segments were mounted in Ussing chambers, in which secretion and permeability were measured. Body weight (as a growth index) and food intake were determined daily. Stress increased baseline jejunal epithelial ion secretion (indicated by short-circuit current), ionic permeability (conductance), and macromolecular permeability (horseradish peroxidase flux) in +/+ rats, but not in Ws/Ws rats, compared with nonstressed controls. Stress induced weight loss and reduced food intake similarly in the groups. In +/+ rats, these parameters remained altered 24-72 h after the cessation of stress. Modulation of stress-induced mucosal mast cell activation may help in the management of certain intestinal conditions involving epithelial pathophysiology. PMID- 10859214 TI - Induction of transcriptional activity of AP-1 and NF-kappaB in the gastric mucosa during aging. AB - Although aging enhances expression and tyrosine kinase activity of epidermal growth factor receptor (EGFR) in the gastric mucosa, there is no information about EGFR signaling cascades. We examined the age-related changes in mitogen activated protein kinases (MAPKs) [extracellular signal-related kinases (ERKs), c Jun NH(2)-terminal kinases (JNKs), and p38], an EGFR-induced signaling cascade, and activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB) transcriptional activity in the gastric mucosa of 4- to 6-, 12- to 14-, and 22- to 24-mo-old Fischer 344 rats. AP-1 and NF-kappaB transcriptional activity in the gastric mucosa rose steadily with advancing age. This can be further induced by transforming growth factor-alpha. The age-related activation of AP-1 and NF kappaB in the gastric mucosa was associated with increased levels of c-Jun, c Fos, and p52, but not p50 or p65. Total and phosphorylated IkappaBalpha levels in the gastric mucosa were unaffected by aging. Aging was also associated with marked activation of ERKs (p42/p44) and JNK1. In contrast, aging decreased p38 MAPK activity in the gastric mucosa. Our observation of increased activation of ERKs and JNK1 in the gastric mucosa of aged rats suggests a role for these MAPKs in regulating AP-1 and NF-kappaB transcriptional activity. These events may be responsible for the age-related rise in gastric mucosal proliferative activity. PMID- 10859215 TI - Slowing of intestinal transit by fat depends on naloxone-blockable efferent, opioid pathway. AB - Slowing of transit through the proximal small intestine by fat in the distal gut is termed the ileal brake. Intravenous naloxone, an opioid receptor antagonist, abolished the fat-induced ileal brake, suggesting that an endogenous opioid pathway may be involved in this response. To test the hypothesis that slowing of intestinal transit by fat in the distal half of the gut depends on an opioid pathway located on the efferent limb of this response, we compared intestinal transit in dogs equipped with duodenal and midgut fistulas while naloxone was either compartmentalized with oleate to the distal half of the gut or with buffer to the proximal half of the gut. We found that intestinal transit depended on the perfusion conditions (P<0.00001). Specifically, compared with ileal brake (marker recovery of 35.7+/-7.4%), intestinal transit was accelerated when naloxone was delivered into the proximal half of the gut (76.2+/-5.2%) (P<0.005) but not the distal half of the gut (29.4+/-5.4%). We conclude that slowing of intestinal transit by fat in the distal half of the gut depends on an opioid pathway located on the efferent limb of the ileal brake. PMID- 10859216 TI - Salivary epidermal growth factor and intestinal adaptation in male and female mice. AB - Salivary epidermal growth factor (sEGF) levels are increased in male mice after small bowel resection (SBR) and may be important during intestinal adaptation. Since males have greater sEGF than females, the influence of sex on postresection adaptation was tested. Females had lower sEGF; however, sEGF substantially increased in both sexes after a massive (50%) SBR. Adaptive increases in DNA and protein content, villus height, and crypt depth, as well as crypt cell proliferation rates in the remnant ileum, were not different between males and females. Although significant postresection increases in sEGF were identified, EGF mRNA and protein did not change within the submandibular gland. Glandular kallikrein-13 and ileal EGF receptor expression were greater after SBR in female mice. Intestinal adaptation is equivalent in female and male mice after SBR. Despite lower sEGF, females demonstrated increased expression of a kallikrein responsible for sEGF precursor cleavage as well as amplified ileal EGF receptor expression. These results endorse an important differential response between sexes regarding sEGF mobilization and intestinal receptor availability during adaptation. PMID- 10859217 TI - Regulation of E-selectin expression in postischemic intestinal microvasculature. AB - Monolayers of cultured endothelial cells exposed to hypoxia-reoxygenation exhibit a transcription-dependent increase in E-selectin expression and E-selectin dependent neutrophil-endothelial cell adhesion. The overall objectives of this study were 1) to determine whether ischemia-reperfusion (I/R) promotes upregulation of E-selectin in vivo; 2) if so, to define the mediators of this response; and 3) to assess the contribution of E-selectin to I/R-induced neutrophil recruitment. The dual-radiolabeled monoclonal antibody (MAb) technique was used to measure E-selectin expression in the intestinal vasculature. Ischemia was induced by complete occlusion (30-60 min) of the superior mesenteric artery followed by 3-24 h of reperfusion. Increasing durations of ischemia elicited progressively increasing (2- to 5-fold) levels of E-selectin expression, with the peak response noted after 45 min of ischemia and 5 h of reperfusion. Subsequent experiments revealed that I/R-induced increase in E-selectin expression (at 5 h) is significantly blunted in transgenic mice that overexpress Cu,Zn-superoxide dismutase or by treatment of wild-type mice with either a blocking antibody against tumor necrosis factor (TNF)-alpha or an inhibitor of nuclear factor kappaB (NF-kappaB) activation (PS341). Administration of an E-selectin-specific MAb dramatically reduced I/R-induced recruitment of neutrophils in the intestine. These findings suggest that superoxide and TNF-alpha mediate gut I/R-induced E selectin expression via an NF-kappaB-dependent mechanism; this upregulation of E selectin contributes significantly to I/R-induced neutrophil recruitment. PMID- 10859218 TI - Intraepithelial lymphocytes coinduce nitric oxide synthase in intestinal epithelial cells. AB - The study of mucosal immunity has revealed that complex reciprocal interactions occur between intestinal intraepithelial lymphocytes (IEL) and intestinal epithelial cells (IEC). The present study focuses on the induction of inducible nitric oxide (NO) synthase in cocultures of freshly isolated rat IEL and the rat epithelial cell line IEC-18 after the addition of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha, or lipopolysaccharide. When IEL and IEC were separated using Transwell chambers, NO synthesis was not induced, indicating that cell-cell contact was required. Culture of IEC-18 with IEL, even in the absence of inflammatory stimuli such as IL-1beta, resulted in upregulation of class I and II antigens on IEC-18, due to the interferon-gamma (IFN-gamma) that is constitutively produced by IEL. Addition of anti-IFN-gamma antibody to the NO producing cocultures resulted in inhibition of NO synthesis as well as the upregulation of class I and II antigen expression. These data indicate that IFN gamma production by IEL conditions IEC for the expression of other components of the inflammatory process. PMID- 10859220 TI - Receptor-induced Ca(2+) signaling in cultured myenteric neurons. AB - We studied the effect of excitatory neurotransmitters (10(-5) M) on the intracellular Ca(2+) concentration ([Ca(2+)](i)) of cultured myenteric neurons. ACh evoked a response in 48.6% of the neurons. This response consisted of a fast and a slow component, respectively mediated by nicotinic and muscarinic receptors, as revealed by specific agonists and antagonists. Substance P evoked a [Ca(2+)](i) rise in 68.2% of the neurons, which was highly dependent on Ca(2+) release from intracellular stores, since after thapsigargin (5 microM) pretreatment only 8% responded. The responses to serotonin, present in 90.7%, were completely blocked by ondansetron (10(-5) M), a 5-HT(3) receptor antagonist. Specific agonists of other serotonin receptors were not able to induce a [Ca(2+)](i) rise. Removing extracellular Ca(2+) abolished all serotonin and fast ACh responses, whereas substance P and slow ACh responses were more persistent. We conclude that ACh-induced signaling involves both nicotinic and muscarinic receptors responsible for a fast and a more delayed component, respectively. Substance P-induced signaling requires functional intracellular Ca(2+) stores, and the 5-HT(3) receptor mediates the serotonin-induced Ca(2+) signaling in cultured myenteric neurons. PMID- 10859219 TI - Heterogeneity of detergent-insoluble membranes from human intestine containing caveolin-1 and ganglioside G(M1). AB - In intestinal epithelia, cholera and related toxins elicit a cAMP-dependent chloride secretory response fundamental to the pathogenesis of toxigenic diarrhea. We recently proposed that specificity of cholera toxin (CT) action in model intestinal epithelia may depend on the toxin's cell surface receptor ganglioside G(M1). Binding G(M1) enabled the toxin to elicit a response, but forcing the toxin to enter the cell by binding the closely related ganglioside G(D1a) rendered the toxin inactive. The specificity of ganglioside function correlated with the ability of G(M1) to partition CT into detergent-insoluble glycosphingolipid-rich membranes (DIGs). To test the biological plausibility of these hypotheses, we examined native human intestinal epithelia. We show that human small intestinal epithelia contain DIGs that distinguish between toxin bound to G(M1) and G(D1a), thus providing a possible mechanism for enterotoxicity associated with CT. We find direct evidence for the presence of caveolin-1 in DIGs from human intestinal epithelia but find that these membranes are heterogeneous and that caveolin-1 is not a structural component of apical membrane DIGs that contain CT. PMID- 10859221 TI - Accessibility of glycolipid and oligosaccharide epitopes on rabbit villus and follicle-associated epithelium. AB - The initial step in many mucosal infections is pathogen attachment to glycoconjugates on the apical surfaces of intestinal epithelial cells. We examined the ability of virus-sized (120-nm) and bacterium-sized (1-microm) particles to adhere to specific glycolipids and protein-linked oligosaccharides on the apical surfaces of rabbit Peyer's patch villus enterocytes, follicle associated enterocytes, and M cells. Particles coated with the B subunit of cholera toxin, which binds the ubiquitous glycolipid GM1, were unable to adhere to enterocytes or M cells. This confirms that both the filamentous brush border glycocalyx on enterocytes and the thin glycoprotein coat on M cells can function as size-selective barriers. Oligosaccharides containing terminal beta(1,4)-linked galactose were accessible to soluble lectin Ricinus communis type I on all epithelial cells but were not accessible to lectin immobilized on beads. Oligosaccharides containing alpha(2, 3)-linked sialic acid were recognized on all epithelial cells by soluble Maackia amurensis lectin II (Mal II). Mal II coated 120-nm (but not 1-microm) particles adhered to follicle-associated enterocytes and M cells but not to villus enterocytes. The differences in receptor availability observed may explain in part the selective attachment of viruses and bacteria to specific cell types in the intestinal mucosa. PMID- 10859222 TI - Glucagon-like peptide (GLP-1) is involved in the central modulation of fecal output in rats. AB - In addition to its insulinotropic action, exogenously administered glucagon-like peptide (GLP-1) inhibits gastropancreatic motility and secretion via central pathways. The aims of the present study were to evaluate the effects of exogenous GLP-1-(7-36) amide on fecal output and to investigate the role of endogenous GLP 1 on stress-induced colonic activity. With the use of a stereotaxic instrument, adult male Sprague-Dawley rats weighing 200-250 g were fitted with stainless steel cerebroventricular guide cannulas under ketamine anesthesia. A group of rats were placed in Bollman-type cages to induce restraint stress. Fecal output monitored for 2 h was increased significantly by intracerebroventricular GLP-1 to 500, 1, 000, and 3,000 pmol/rat (P < 0.05-0.01), whereas intraperitoneal GLP-1 had no effect. Intracerebroventricular administration of the GLP-1 receptor antagonist exendin-(9-39) (10 nmol/rat) reversed the increases induced by GLP-1 (500 pmol/rat; P<0.01). Similar results were also observed with the injection of corticotropin-releasing factor receptor antagonist astressin (10 microg/rat icv). The significant increase in fecal pellet output induced by restraint stress was also decreased by both intracerebroventricular exendin (10 nmol/rat) and astressin (10 microg/rat; P<0.01-0.001). These results suggest that GLP-1 participates in the central, but not peripheral, regulation of colonic motility via its own receptor and that GLP-1 is likely to be a candidate brain-gut peptide that acts as a physiological modulator of stress-induced colonic motility. PMID- 10859223 TI - Gene expression of divalent metal transporter 1 and transferrin receptor in duodenum of Belgrade rats. AB - Regulation of iron absorption is thought to be mediated by the amount of iron taken up by duodenal crypt cells via the transferrin receptor (TfR)-transferrin cycle and the activity of the divalent metal transporter (DMT1), although DMT1 cannot be detected morphologically in crypt cells. We investigated the uptake of transferrin-bound iron by duodenal enterocytes in Wistar rats fed different levels of iron and Belgrade (b/b) rats in which iron uptake by the transferrin cycle is defective because of a mutation in DMT1. We showed that DMT1 in our colony of b/b rats contains the G185R mutation, which in enterocytes results in reduced cellular iron content and increased DMT1 gene expression similar to levels in iron deficiency of normal rats. In all groups the uptake of transferrin bound iron by crypt cells was directly proportional to plasma iron concentration, being highest in iron-loaded Wistar rats and b/b rats. We conclude that the uptake of transferrin-bound iron by developing enterocytes is largely independent of DMT1. PMID- 10859224 TI - Luminal dietary protein, not amino acids, induces pancreatic protease via CCK in pancreaticobiliary-diverted rats. AB - We determined whether pancreatic adaptation to a high-protein diet depends on ingested protein in the intestinal lumen and whether such adaptation depends on a CCK or capsaicin-sensitive vagal afferent pathway in pancreaticobiliary-diverted (PBD) rats. Feeding a high-casein (60%) diet but not a high-amino acid diet to PBD rats increased pancreatic trypsin and chymotrypsin activities compared with those after feeding a 25% casein diet. In contrast, feeding both the high nitrogen diets induced pancreatic hypertrophy in PBD rats. These pancreatic changes by the diets were abolished by treatment with devazepide, a CCK-A receptor antagonist. Protease zymogen mRNA abundance in the PBD rat was not increased by feeding the high-casein diet and was decreased by devazepide. Perivagal capsaicin treatment did not influence the values of any pancreatic variables in PBD rats fed the normal or high-casein diet. We concluded that luminal protein or peptides were responsible for the bile pancreatic juice independent induction of pancreatic proteases on feeding a high-protein diet. The induction was found to be dependent on the direct action of CCK on the pancreas. Pancreatic growth induced by high-protein feeding in PBD rats may depend at least partly on absorbed amino acids. PMID- 10859225 TI - Pramlintide, an amylin analog, selectively delays gastric emptying: potential role of vagal inhibition. AB - The amylin analog pramlintide delays gastric emptying in type I diabetics. The effects of multiple doses of pramlintide and the mechanism of action in non amylin-deficient humans are unknown. We investigated the effects of pramlintide on gastrointestinal and colonic transit and on the plasma pancreatic polypeptide response to the meal in a parallel-group dose-response study with subjects randomized to placebo, or 30 or 60 microg (tid, sc) of pramlintide. Pramlintide delayed gastric emptying [half-time (t(1/2)): 112 min (SE 8.7 min), 169 min (SE 12 min), or 177 min (SE 25 min) after placebo or 30- or 60-microg pramlintide treatment, respectively; P = 0.033]. Pramlintide did not significantly affect small bowel or colonic transit. Pancreatic polypeptide concentrations in the first postprandial hour were lower with pramlintide than with placebo (P<0.01 for drug effect). An inverse correlation was observed between mean pancreatic polypeptide concentrations in the first postprandial hour and gastric emptying t(1/2) [Spearman correlation coefficient (R(s)) = 0.48; P = 0.044]. Pramlintide at 30 and 60 microg delays gastric emptying in healthy humans without affecting small bowel or colonic transit. Vagal inhibition is a potential mechanism of the effects of pramlintide on gastric emptying. PMID- 10859226 TI - Human caco-2 motility redistributes FAK and paxillin and activates p38 MAPK in a matrix-dependent manner. AB - The signals involved in restitution during mucosal healing are poorly understood. We compared focal adhesion kinase (FAK) and paxillin protein and phosphorylation, extracellular signal-regulated kinase (ERK) 1, ERK2, and p38 activation, as well as FAK and paxillin organization in static and migrating human intestinal Caco-2 cells on matrix proteins and anionically derivatized polystyrene dishes (tissue culture plastic). We also studied effects of FAK, ERK, and p38 blockade in a monolayer-wounding model. Compared with static cells, cells migrating across matrix proteins matrix-dependently decreased membrane/cytoskeletal FAK and paxillin and cytosolic FAK. Tyrosine phosphorylated FAK and paxillin changed proportionately to FAK and paxillin protein. Conversely, cells migrating on plastic increased FAK and paxillin protein and phosphorylation. Migration matrix dependently activated p38 and inactivated ERK1 and ERK2. Total p38, ERK1, and ERK2 did not change. Caco-2 motility was inhibited by transfection of FRNK (the COOH-terminal region of FAK) and PD-98059, a mitogen-activated protein kinase-ERK kinase inhibitor, but not by SB-203580, a p38 inhibitor, suggesting that FAK and ERK modulate Caco-2 migration. In contrast to adhesion-induced phosphorylation, matrix may regulate motile intestinal epithelial cells by altering amounts and distribution of focal adhesion plaque proteins available for phosphorylation as well as by p38 activation and ERK inactivation. Motility across plastic differs from migration across matrix. PMID- 10859227 TI - Lipolysis and lipid oxidation in cirrhosis and after liver transplantation. AB - On the basis of the finding that plasma glycerol concentration is not controlled by clearance in healthy humans, it has been proposed that elevated plasma free fatty acid (FFA) and glycerol concentrations in cirrhotic subjects are caused by accelerated lipolysis. This proposal has not been validated. We infused 10 volunteers, 10 cirrhotic subjects, and 10 patients after orthotopic liver transplantation (OLT) with [1-(13)C]palmitate and [(2)H(5)]glycerol to compare fluxes (R(a)) and FFA oxidation. Cirrhotic subjects had higher plasma palmitate (52%) and glycerol (33%) concentrations than controls. Palmitate R(a) was faster (1.45+/-0.18 vs. 0.85+/-0.17 micromol x kg(-1) x min(-1)) but glycerol R(a) and clearance slower (1.20+/-0.09 vs. 1.90+/-0.24 micromol x kg(-1) x min(-1) and 21.2+/-1.2 vs. 44.7+/- 4.9 ml x kg(-) x h(-1), respectively) than in controls. After OLT, plasma palmitate and glycerol concentrations and palmitate R(a) did not differ, but glycerol R(a) (1.16+/-0.11 micromol x kg(-1) x min(-1)) and clearance (26.7+/-2.4 ml x kg(-1) x h(-1)) were slower than in controls. We conclude that 1) impaired reesterification, not accelerated lipolysis, elevates FFA in cirrhotic subjects; 2) normalized FFA after OLT masks impaired reesterification; and 3) plasma glycerol concentration poorly reflects lipolytic rate in cirrhosis and after OLT. PMID- 10859228 TI - G(i-1)/G(i-2)-dependent signaling by single-transmembrane natriuretic peptide clearance receptor. AB - Single-transmembrane natriuretic peptide clearance receptor (NPR-C), which is devoid of a cytoplasmic guanylyl cyclase domain, interacts with pertussis toxin (PTx)-sensitive G proteins to activate endothelial nitric oxide synthase (eNOS) expressed in gastrointestinal smooth muscle cells. We examined the ability of NPR C to activate other effector enzymes in eNOS-deficient tenia coli smooth muscle cells; these cells expressed NPR-C and NPR-B but not NPR-A. Atrial natriuretic peptide (ANP), the selective NPR-C ligand cANP-(4-23), and vasoactive intestinal peptide (VIP) inhibited (125)I-ANP and (125)I-VIP binding to muscle membranes in a pattern indicating high-affinity binding to NPR-C. Interaction of VIP with NPR C was confirmed by its ability to inhibit (125)I-ANP binding to membranes of NPR C-transfected COS-1 cells. In tenia muscle cells, all ligands selectively activated G(i-1) and G(i-2); VIP also activated G(s) via VIP(2) receptors. All ligands stimulated phosphoinositide hydrolysis, which was inhibited by ANP-(1 11), PTx, and antibodies to phospholipase C-beta3 (PLC-beta3) and Gbeta. cANP-(4 23) contracted tenia muscle cells; contraction was blocked by U-73122 and PTx and by antibodies to PLC-beta3 and Gbeta in intact and permeabilized muscle cells, respectively. VIP and ANP contracted muscle cells only after inhibition of cAMP- and cGMP-dependent protein kinases. ANP and cANP-(4-23) inhibited forskolin stimulated cAMP in a PTx-sensitive fashion. We conclude that NPR-C is coupled to activation of PLC-beta3 via betagamma-subunits of G(i-1) and G(i-2) and to inhibition of adenylyl cyclase via alpha-subunits. PMID- 10859229 TI - Inhibition of human gastric H(+)-K(+)-ATPase alpha-subunit gene expression by Helicobacter pylori. AB - Clinical studies and in vitro data from isolated parietal cells suggest that acute Helicobacter pylori infection inhibits acid secretion. Gastric acidification is mediated by H(+)-K(+)-ATPase, an integral protein of parietal cell apical membranes. To test the hypothesis that H. pylori downregulates H(+) K(+)-ATPase alpha-subunit (HKalpha) gene expression and to identify potential intracellular signaling pathways mediating such regulation, we transfected human gastric adenocarcinoma (AGS) cells with human and rat HKalpha 5'-flanking DNA fused to a luciferase reporter plasmid. Histamine caused dose-dependent, cimetidine-sensitive (10(-4) M) increases in cAMP, free intracellular Ca(2+), and HKalpha promoter activation in AGS cells. H. pylori infection of transfected AGS cells dose dependently inhibited basal and histamine-stimulated HKalpha promoter activity by 80% and 66%, respectively. H. pylori dose dependently inhibited phorbol myristate acetate-induced (10(-7) M) and staurosporine- (10(-7) M) and calphostin C-sensitive (5 x 10(-8) M) activation of HKalpha promoter. Also, H. pylori inhibited epidermal growth factor (EGF) (10(-8) M), genistein-sensitive (5 x 10(-5) M) activation of HKalpha promoter, reducing activity to 60% of basal level. These data suggest that H. pylori inhibits HKalpha gene expression via intracellular pathways involving protein kinases A and C and protein tyrosine kinase, AGS cells have functional histamine H(2) and EGF receptors, and transiently transfected AGS cells are a useful model for studying regulation of HKalpha gene expression. PMID- 10859230 TI - Bile salts mediate hepatocyte apoptosis by increasing cell surface trafficking of Fas. AB - Toxic bile salts induce hepatocyte apoptosis by a Fas-dependent, Fas ligand independent mechanism. To account for this observation, we formulated the hypothesis that toxic bile salts induce apoptosis by effecting translocation of cytoplasmic Fas to the cell surface, resulting in transduction of Fas death signals. In McNtcp.24 cells the majority of Fas was cytoplasmic, as assessed by cell fractionation and immunofluorescence studies. However, cell surface Fas increased sixfold after treatment with the toxic bile salt glycochenodeoxycholate (GCDC) in the absence of increased Fas protein expression. Moreover, in cells transfected with Fas-green fluorescence protein, cell surface fluorescence also increased in GCDC-treated cells, directly demonstrating Fas translocation to the plasma membrane. Both brefeldin A, a Golgi-disrupting agent, and nocodazole, a microtubule inhibitor, prevented the GCDC-induced increase in cell surface Fas and apoptosis. In conclusion, toxic bile salts appear to induce apoptosis by promoting cytoplasmic transport of Fas to the cell surface by a Golgi- and microtubule-dependent pathway. PMID- 10859232 TI - Editor's preface PMID- 10859231 TI - Bile salts induce or blunt cell proliferation in Barrett's esophagus in an acid dependent fashion. AB - Barrett's esophagus (BE) results from acid and bile reflux and predisposes to cancer. We investigated the effect of bile salts, with or without acid, on cell proliferation in BE and assessed mechanism(s) involved. To mimic physiological conditions, biopsies of esophagus, BE, and duodenum were exposed to a bile salt mixture, either continuously or as a 1-h pulse, and were compared with control media without bile salts (pH 7.4) for < or =24 h. Similar experiments were also performed with acidified media (pH 3.5) combined with the bile salt mixture as a 1-h pulse. Cell proliferation was assessed by a [(3)H]thymidine incorporation assay with or without bisindolylmaleimide (BIM), a selective protein kinase C inhibitor. Bile salt pulses enhanced cell proliferation in BE without affecting cell proliferation in esophageal or duodenal epithelia. In the presence of BIM, there was complete obliteration of the bile salt-induced BE hyperproliferation. In contrast, 1-h pulses of bile salts in combination with acid significantly inhibited proliferation in BE but had no effect on esophagus or duodenum. We conclude that in BE explants, brief exposure to bile salts, in the absence of acid, increases proliferation, whereas exposure to a combination of bile salts and acid together inhibits proliferation. PMID- 10859233 TI - EDITOR'S PREFACE PMID- 10859234 TI - Cellular and molecular regulation of luteal formation, function, and regression: An introduction. PMID- 10859235 TI - Models of luteinization. AB - Luteinization is essential to the success of early gestation. It is the process by which elements of the ovarian follicle, usually including both theca interna and granulosa cells, are provoked by the ovulatory stimulus to develop into the corpus luteum. Although there are significant species differences in luteinization, some elements pervade, including the morphological and functional differentiation to produce and secrete progesterone. There is evidence that luteinization results in granulosa cell exit from the cell cycle. The mechanisms that appear to control luteinization include intracellular signalling pathways, cell adhesion factors, intracellular cholesterol and oxysterols, and perhaps progesterone itself as a paracrine or intracrine regulator. Cell models of luteinization, along with some of the conflicting observations on the luteinization process, are discussed in this review. PMID- 10859236 TI - Phosphorylation of myristoylated alanine-rich C kinase substrate (MARCKS) protein is associated with bovine luteal oxytocin exocytosis. AB - The ruminant corpus luteum, in addition to producing progesterone, synthesizes and secretes oxytocin (OT) during the estrous cycle. Secretion of oxytocin occurs by exocytosis of membrane-encapsulated granules of this hormone. Exocytosis of oxytocin involves transport of granules through a cytoskeletal matrix including an actin cortex closely associated with the plasma membrane (PM). Actin filaments crosslinked by various proteins give rise to the structural integrity of the cortex. Myristoylated alanine-rich C kinase substrate (MARCKS), a protein specifically phosphorylated by protein kinase C (PKC), crosslinks actin filaments and anchors the actin network to the inner leaflet of the PM. There is evidence that the intact actin cortex may serve as a barrier, precluding fusion of transport vesicles with the PM. In some secretory cells, phosphorylation of MARCKS has resulted in its translocation from the PM to the cytoplasm with an associated disassembly of the actin cortex. Prostaglandin F(2alpha) (PGF(2alpha)) stimulation of the bovine corpus luteum during the midluteal phase of the estrous cycle activates PKC, which is associated with an increase in OT secretion in vivo and in vitro. Data are presented demonstrating that stimulation of bovine luteal cells with PGF(2alpha) on Day 8 of the cycle promotes rapid phosphorylation of MARCKS protein and causes its translocation from the PM to the cytoplasm and concomitant, enhanced exocytosis of OT. These data are consistent with the premise that MARCKS plays a role in the exocytotic process. PMID- 10859237 TI - Increased expression of insulin-like growth factor binding protein-1 during induced regression of bovine corpora lutea. AB - Three experiments were conducted to examine gene expression during induced luteal regression in the cow; the initial purpose was the identification of potential embryotoxins. In experiment 1, changes in gene expression in the corpus luteum (CL) were identified by differential display reverse transcription-polymerase chain reaction (DD-PCR) during the first 72 h of luteal regression in cows treated with prostaglandin F(2alpha) (PGF(2alpha)) on Days 4-7 after estrus. Expression of insulin-like growth factor-binding protein-1 (IGFBP-1) was up regulated, with greatest expression at 24 h (P < 0.05) after treatment with PGF(2alpha) began. In experiment. 2, IGFBP-1 and its mRNA were quantified in CL collected 24 or 48 h after treatment with PGF(2alpha) on Day 4 or 10 after estrus. Because local mechanisms for exchange of hormones between the ovary and uterus are known in ruminants, uterine flushings were assayed for IGFBP-1 to seek evidence of local transfer of luteal IGFBP-1 to the uterus. IGFBP-1 mRNA was increased (P < 0.05) in CL 24 h after treatment when PGF(2alpha) that began on Day 10, and by 48 h after treatment that began on Day 4. Concentrations of IGFBP 1 increased (P < 0.05) in a pattern similar to mRNA, by 24 h on Day 10, and by 48 h on Day 4. Concentrations of IGFBP-1 in uterine flushings did not change on either day. Concentrations of progesterone decreased (P < 0.05) by 8 h after treatment with PGF(2alpha) that began on Day 10, but not until 24 h after treatment that began on Day 4. In experiment 3, cows received either saline or PGF(2alpha) and CL were collected 2 or 10 h after a single treatment, or 2 h after a second treatment that was given 8 h after the first. Expression of IGFBP 1 was increased by 2 h after treatment with PGF(2alpha) on both Days 4 and 10 after estrus. In conclusion, secretion of IGFBP-1 is increased during luteolysis, and may inhibit the steroidogenic effects of insulin-like growth factor-I (IGF I), but no evidence was found to implicate IGFBP-1 in the embryotoxic effect of regressing CL. PMID- 10859238 TI - Purification, characterization, and biological compartmentalization of rat fetal antigen 1. AB - This study has established the rat as an animal model for the analysis of the biological role of fetal antigen 1 (FA1), a protein previously described in humans and mice. FA1 was purified from rat amniotic fluid by immunospecific affinity chromatography. Immunochemical identity between mouse and rat FA1 was established by crossed tandem immunoelectrophoresis. Molecular size was analyzed by mass spectrometry (33 kDa). The amino acid composition was determined, and the amino acid sequence was analyzed. The overall amino acid composition and sequence of the 28 first N-terminal amino acids were identical to the corresponding parts of rat preadipocyte factor 1 and rat adrenal zona glomerulosa protein. Extensive sequence similarity was found between rat and mouse FA1 (86%) and between rat and human FA1 (82%). The concentration of FA1 in fetal serum, maternal serum, urine, and amniotic fluid in rats was determined using an ELISA. The highest concentrations were found in fetal serum and amniotic fluid around Day 18 of pregnancy. This is the first report on the physicochemical characteristics and compartmentalization of rat FA1. PMID- 10859239 TI - Estradiol-17beta inhibits nitric oxide synthase (NOS)-II and stimulates NOS-III gene expression in the rat uterus. AB - Nitric oxide (NO) is synthesized by NO synthases (NOS) from L-arginine in a variety of tissues, including rat uterus. Progesterone was shown to be required for maintaining elevated NOS II expression in pregnant rat uterus. However, effects of estrogens on uterine NOS II expression remains unclear. In the present study, we examined whether 17beta-estradiol regulates NO production and NOS II expression in the rat uterus during pregnancy and in nonpregnant rats treated with lipopolysaccharide (LPS). Rats on Day 18 of pregnancy received 17beta estradiol (0.5 or 5 microgram/rat). Groups of ovariectomized (ovx) rats received 17beta-estradiol (5 microgram/rat) or LPS (1 mg/rat) or a combination of the two or received vehicle only. All rats were sacrificed 24 h after treatments. Nitrite concentrations in uterine cultures were measured by Greiss reaction. Uterine NOS II and NOS III proteins and mRNA levels were determined by Western blotting and reverse transcription polymerase chain reaction, respectively. In the pregnant rat, estradiol administration caused inhibition in total NO production, suppression of both mRNA and protein levels of NOS II enzyme, and increase in NOS III mRNA and protein levels in the uterus in a dose-dependent manner. The data indicate that estradiol inhibits NOS II and total NO generation and stimulates NOS III expression. In ovx rats, LPS stimulated NOS II mRNA and NO production by the uterus. Coadministration of 5 microgram estradiol profoundly suppressed NOS II mRNA and NO generation but elevated NOS III mRNA. Thus, estradiol inhibited LPS-induced increases in NOS II mRNA. Estradiol inhibits NO production by NOS II through the inhibition of NOS II expression in the rat uterus. This inhibition of NOS II expression occurs whether NOS II expression is constitutive (pregnancy) or induced (LPS-treated nonpregnant). Estradiol inhibition of NOS II expression occurs in the presence (pregnancy) or absence (ovx) of progesterone. Estradiol may play a role in regulating NOS II expression and NO production and uterine contractility during pregnancy and labor. PMID- 10859240 TI - Trypsin/acrosin inhibitor activity of rat and guinea pig caltrin proteins. Structural and functional studies. AB - Dramatic inhibition of trypsin activity by rat caltrin and guinea pig caltrin I was spectrophotometrically demonstrated using the artificial substrate benzoylarginyl ethyl ester. Approximately 6% and 21% of residual proteolytic activity was recorded after preincubating the enzyme with 0.22 and 0.27 microM rat caltrin and guinea pig caltrin I, respectively. Reduction and carboxymethylation of the cysteine residues abolished the inhibitor activity of both caltrin proteins. Rat caltrin and guinea pig caltrin I show structural homology with secretory trypsin/acrosin inhibitor proteins isolated from boar and human seminal plasma and mouse seminal vesicle secretion and share a fragment of 13 amino acids of almost identical sequence (DPVCGTDGH/K/ITYG/AN), which is also present in the structure of Kazal-type trypsin inhibitor proteins from different mammalian tissues. Bovine, mouse, and guinea pig caltrin II, three caltrin proteins that have no structural homology with rat caltrin or guinea pig caltrin I, lack trypsin inhibitor activity. Rat caltrin, guinea pig caltrin I, and the mouse seminal vesicle trypsin inhibitor protein P12, which also inhibits Ca(2+) uptake into epididymal spermatozoa (mouse caltrin I), bound specifically to the sperm head, on the acrosomal region, as detected by indirect immunofluorescence. They also inhibited the acrosin activity in the gelatin film assay. Caltrin I may play an important role in the control of sperm functions such as Ca(2+) influx in the acrosome reaction and activation of acrosin and other serine-proteases at the proper site and proper time to ensure successful fertilization. PMID- 10859241 TI - Responsiveness of mouse corpora luteal cells to Fas antigen (CD95)-mediated apoptosis. AB - Regression of the corpus luteum (CL) occurs by apoptosis. The Fas antigen (Fas) is a cell surface receptor that induces apoptosis in sensitive cells when bound to Fas ligand or agonistic anti-Fas monoclonal antibodies (Fas mAb). A potential role for Fas to induce apoptosis in dispersed CL cell preparations was tested in cells isolated from mice on Days 2-4 of pseudopregnancy. Total CL dispersates, containing steroidogenic luteal cells, fibroblasts, and endothelial cells, were cultured. The effect of pretreatment of cultures with cytokines interferon gamma (IFN) and tumor necrosis factor alpha (TNF) was examined because these cytokines demonstrated effects on Fas-mediated apoptosis in other cell types. Fas mAb had no effect on viability of CL cells cultured in 5% fetal bovine serum (FBS) and pretreated with or without IFN or TNF, but Fas mAb did kill 23% of the cells in cultures pretreated with IFN + TNF. Fas mRNA was detectable in cultured CL cells and was increased 2.1-, 2. 0-, and 11.8-fold by treatment with TNF, IFN, or IFN + TNF, respectively. CL cells treated with the protein synthesis inhibitor cycloheximide (CX) were killed by Fas mAb in the absence of cytokine pretreatment (34%); pretreatment with IFN or IFN + TNF further potentiated killing (62% and 96%, respectively), whereas pretreatment with TNF had no effect (42%). Cells cultured in medium supplemented with insulin, transferrin, and selenium instead of FBS were killed by Fas mAb in the presence of IFN (23%) or IFN + TNF (29%) but not in the presence of TNF. Cells derived from the mouse CL have a functional Fas pathway that is inhibited by FBS and activated by treatment with CX, IFN, and IFN + TNF. PMID- 10859242 TI - Steroidal sigma receptor ligands affect signaling pathways in human spermatozoa. AB - In human spermatozoa, Ca(2+) entry is stimulated by progesterone or prostaglandin E(1) (PGE(1)). The regulation of cation currents by progestins involves sigma receptors, and sigma binding sites are abundant in testis. We examined the effects of sigma ligands on human spermatozoa. Ca(2+) entry induced by progesterone or PGE(1) was not altered by the sigma ligands haloperidol and ditolylguanidine. However, the steroidal sigma ligands RU 3117 and RU 1968 had distinct effects. Stimulation by RU 3117 resulted in activation and homologous desensitization of the sperm progesterone receptor but not of the PGE(1) receptor. Because haloperidol and ditolylguanidine did not affect RU 3117 and progesterone actions in spermatozoa, we conclude that sigma receptors are not involved. However, RU 1968 potently inhibited both the progesterone- and PGE(1) induced Ca(2+) entry and acrosome reaction. At higher concentrations, RU 1968 also inhibited hormonal Ca(2+) signaling in fibroblasts. Despite suppression of Ca(2+) mobilization, inhibition of phospholipase C by RU 1968 was not observed. Furthermore, RU 1968 did not impair the binding of inositol-1,4,5-trisphosphate to its endoplasmic reticulum receptor. Because RU 1968 preferentially inhibits signaling pathways in spermatozoa, the future development of more selective drugs structurally related to RU 1968 may be a novel approach for pharmacological contraception. PMID- 10859243 TI - Effects of X chromosome number and parental origin on X-linked gene expression in preimplantation mouse embryos. AB - Diploid androgenetic mouse embryos, possessing two sets of paternally inherited chromosomes, and control fertilized embryos were used to examine the relative effects of X chromosome number and parental chromosome origin on androgenone viability and X-linked gene expression. A significant difference in efficiency of blastocyst formation was observed between XX and XY androgenones in some experiments, but this difference was not uniformly observed. Significant effects of both X chromosome number and parental origin on X-linked gene expression were observed. Male and female control embryos expressed the XIST: RNA initially. This expression was followed by a preferential reduction in XIST: RNA abundance in male embryos, indicating that dosage compensation for the X chromosome may normally require the downregulation of XIST: RNA expression in male embryos, in conjunction with the production of stable XIST: transcripts in female embryos. By the late blastocyst stage, XX control embryos expressed significantly more XIST: RNA than did XY embryos. Unlike their normal counterparts, XX androgenones did not express significantly more XIST: RNA than did XY androgenones at the late blastocyst stage. Androgenones exhibited severe repression of the Pgk1 gene, but during development to the late blastocyst stage Pgk1 mRNA expression increased in XX androgenones and decreased in XY androgenones. Thus, the initial repression of the Pgk1 gene in XX androgenones was lost as the XIST: RNA declined in abundance, and this loss was correlated with a failure of XX androgenones to express significantly more XIST: RNA than did XY androgenones. These results indicate that androgenones may lack a factor that is expressed from the maternal genome and required for dosage compensation in preimplantation embryos. The results also indicate that early dosage compensation in preimplantation embryos may normally be reversible, thus providing flexibility to meet different developmental requirements of the embryonic and extraembryonic lineages. PMID- 10859244 TI - Paternal exposure to cyclophosphamide alters cell-cell contacts and activation of embryonic transcription in the preimplantation rat embryo. AB - Paternal exposure to chronic low doses of cyclophosphamide, an anticancer agent, results in aberrant embryonic development of the progeny. We hypothesized that paternal exposure to cyclophosphamide disturbs zygotic gene activity regulating proper progression through preimplantation development and that this disturbance results in improper cell-cell interactions. To test this hypothesis, we analyzed cell-cell interactions and the expression of cytoskeletal elements in preimplantation embryos sired by male rats gavaged with saline or 6 mg kg(-1) day(-1) cyclophosphamide for 5 wk. Embryos from control litters had 4-12 cells on Day 2 of gestation; cell-cell contacts were observed consistently. Embryos from litters sired by cyclophosphamide-treated males were frequently abnormal and had lower cell numbers and decreased cell-cell contacts. Steady state concentrations of the mRNAs for cell adhesion molecules (cadherins and connexin 43) and structural proteins (beta-actin, collagen, and vimentin) were low in two- and four-cell control embryos; expression increased dramatically by the eight-cell stage. In contrast, embryos sired by cyclophosphamide-treated males displayed the highest expression of most trancripts at the two-cell stage. In parallel with the mRNA profiles, E-cadherin immmunoreactivity was nearly absent in two-cell control embryos and was strong by the eight-cell stage; immunoreactivity in embryos sired by drug-treated fathers was strong at the two-cell stage but absent at later stages. Thus, drug exposure of the paternal genome led to dysregulated expression of structural elements and decreased cell interactions during preimplantation embryonic development. PMID- 10859245 TI - Pulsatile stimulation with recombinant single chain human luteinizing hormone elicits precocious sertoli cell proliferation in the juvenile male rhesus monkey (Macaca mulatta). AB - In this study, we determined the relative role of LH and FSH in initiating the pubertal proliferation of Sertoli cells in primates. Sixteen juvenile male rhesus monkeys (Macaca mulatta) bearing venous catheters received intermittent intravenous infusions of single chain human LH (schLH) or recombinant human FSH (rhFSH) or a combination of both for 11 days. The schLH infusion elicited a physiological testosterone response. On Day 11, monkeys were castrated, and one half of a testis was fixed in Bouin's fluid. Infusion of the gonadotropins, either alone or in combination, effected a significant increase in testicular weight, seminiferous cord diameter, and the number of Sertoli cells per testis (schLH, 295 +/- 46 x 10(6); rhFSH, 342 +/- 64 x 10(6); LH+FSH, 298 +/- 26 x 10(6) versus vehicle, 204 +/- 26 x 10(6)). The latter finding indicated that LH, in addition to FSH, plays a critical role in the initiation of the pubertal proliferation of Sertoli cells in primates. Moreover, combined gonadotropin treatment led to the appearance of germ cells as mature as early primary spermatocytes, indicating that initiation of spermatogenesis had been set in motion. Because the duration of hormone stimulation was only 11 days, the latter result suggests that Leydig and Sertoli cells of the juvenile monkey testis can immediately transduce a gonadotropin signal to the germ cell. PMID- 10859246 TI - Vesicular traffic and golgi apparatus dynamics during mammalian spermatogenesis: implications for acrosome architecture. AB - Vesicular membrane trafficking during acrosome biogenesis in bull and rhesus monkey spermatogenesis differs from the somatic cell paradigm as imaged dynamically using the Golgi apparatus probes beta-COP, giantin, Golgin-97, and Golgin-95/GM130. In particular, sorting and delivery of proteins seemed less precise during spermatogenesis. In early stages of spermiogenesis, many Golgi resident proteins and specific acrosomal markers were present in the acrosome. Trafficking in both round and elongating spermatids was similar to what has been described for somatic cells, as judged by the kinetics of Golgi protein incorporation into endoplasmic reticulum-like structures after brefeldin A treatment. These Golgi components were retrieved from the acrosome at later stages of differentiation and were completely devoid of immature spermatozoa. Our data suggest that active anterograde and retrograde vesicular transport trafficking pathways, involving both beta-COP- and clathrin-coated vesicles, are involved in retrieving Golgi proteins missorted to the acrosome and in controlling the growth and shape of this organelle. PMID- 10859247 TI - Sertoli cell ectoplasmic specializations in the seminiferous epithelium of the testosterone-suppressed adult rat. AB - The Sertoli cell ectoplasmic specialization is a unique junctional structure involved in the interaction between elongating spermatids and Sertoli cells. We have previously shown that suppression of testicular testosterone in adult rats by low-dose testosterone and estradiol (TE) treatment causes the premature detachment of step 8 round spermatids from the Sertoli cell. Because these detaching round spermatids would normally associate with the Sertoli cell via the ectoplasmic specialization, we hypothesized that ectoplasmic specializations would be absent in the seminiferous epithelium of TE-treated rats, and the lack of this junction would cause round spermatids to detach. In this study, we investigated Sertoli cell ectoplasmic specializations in normal and TE-treated rat testis using electron microscopy and localization of known ectoplasmic specialization-associated proteins (espin, actin, and vinculin) by immunocytochemistry and confocal microscopy. In TE-treated rats where round spermatid detachment was occurring, ectoplasmic specializations of normal morphology were observed opposite the remaining step 8 spermatids in the epithelium and, importantly, in the adluminal Sertoli cell cytoplasm during and after round spermatid detachment. When higher doses of testosterone were administered to promote the reattachment of all step 8 round spermatids, newly elongating spermatids associated with ectoplasmic specialization proteins within 2 days. We concluded that the Sertoli cell ectoplasmic specialization structure is qualitatively normal in TE-treated rats, and thus the absence of this structure is unlikely to be the cause of round spermatid detachment. We suggest that defects in adhesion molecules between round spermatids and Sertoli cells are likely to be involved in the testosterone-dependent detachment of round spermatids from the seminiferous epithelium. PMID- 10859248 TI - Development of in vivo-matured porcine oocytes following intracytoplasmic sperm injection. AB - The objective of this study was to assess the development of porcine ova fertilized by intracytoplasmic sperm injection (ICSI). Allyl trenbolone (Regumate) was used to synchronize estrus in 13 postpuberal gilts. Gilts were superovulated with pregnant mare serum gonadotropin and hCG. Ova were aspirated from 5- to 8-mm follicles at 36 h after hCG. Cumulus cells were removed by blunt dissection and pipetting in Beltsville embryo culture medium (BECM) supplemented with 0.1% hyaluronidase. Sperm were washed and resuspended in BECM + 8% polyvinylpyrrolidone. Ova (n = 237) that exhibited a polar body were centrifuged at 15 000 x g for 6 min and injected with a single spermatozoon. One hundred fifty-four ova were cultured in NCSU-23 medium in a 5% CO(2) in air environment for 168 h. Ova were fixed in acetic acid/ethanol and stained with 1% orcein. Sixty-nine ICSI ova were cultured for 24 h and transferred (mean = 23) to three recipients. Eighty-one ova (69%) that survived ICSI cleaved within 48 h. Thirty eight percent (31/81) of these ova became blastocysts (mean +/- SEM = 24.7 +/- 1.1 cells). One recipient gave birth to three pigs. These results demonstrate that porcine embryos derived from ICSI can develop into live pigs. PMID- 10859249 TI - Characterization of intracellular Ca(2+) increase in response to progesterone and cyclic nucleotides in mouse spermatozoa. AB - Rises in intracellular Ca(2+) concentration ([Ca(2+)](i)) caused by progesterone, an inducer of the acrosome reaction, or by cyclic nucleotides, possible second messengers, were investigated by Ca(2+) imaging of the head of individual mouse sperm. Progesterone induced a [Ca(2+)](i) rise in a dose-dependent manner (4-40 microM), primarily in the postacrosomal region. For 20-microM progesterone, Ca(2+) responses occurred in 42% of sperm, separated into two types: transient type (60% of responding cells; duration, 1-1.5 min; mean amplitude, 335 nM) and prolonged type (40%; >3 min; 730 nM). Prolonged responses required higher doses of progesterone, and their occurrence was enhanced significantly by preincubation for 2-4 h as compared with transient responses. 8-Bromo-cGMP (0.3-3 mM) induced a [Ca(2+)](i) rise more effectively than did 8-bromo-cAMP. For 1-mM 8-bromo-cGMP, 90% of cells exhibited transient Ca(2+) responses (approximately 1 min; 220 nM), independently of the preincubation time. In Ca(2+)-free medium, most sperm showed no Ca(2+) response to progesterone and 8-bromo-cGMP. Pimozide, a Ca(2+) channel blocker, completely blocked prolonged responses and partially inhibited transient responses. These results suggest that progesterone activates at least two distinct Ca(2+) influx pathways, with fast or slow inactivation kinetics, and some sperm show both types of response. A cyclic nucleotide-mediated process could participate in the progesterone-induced [Ca(2+)](i) rise. PMID- 10859250 TI - Production and regulation of cytokine-induced neutrophil chemoattractant in rat ovulation. AB - A cytokine-induced neutrophil chemoattractant (CINC/gro), which belongs to the interleukin (IL)-8 family, acts as a functional chemoattractant for neutrophils in rats. In the present study, we examined whether CINC/gro contributes to the ovulation process in the rat ovulation system. In rat ovaries, CINC/gro was immunohistochemically recognized in the theca layer of the antral follicle but not in the granulosa cells. To clarify the role of CINC/gro in the ovulation process, CINC/gro protein and mRNA were examined during pregnant mare serum gonadotropin (PMSG)-hCG treatment. CINC/gro protein did not increase as a result of PMSG injection. However, it increased rapidly after hCG injection and peaked at 6 h after hCG. CINC/gro mRNA was also strongly expressed after hCG injection. The increase of CINC/gro protein followed increases in IL-1beta and tumor necrosis factor alpha (TNFalpha). In the whole ovarian dispersate culture, FSH, hCG, IL-1beta, and TNFalpha stimulated the production of CINC/gro protein in a dose-dependent manner. In particular, the stimulatory effects of IL-1beta and TNFalpha were stronger than those of gonadotropins. These results suggest that CINC/gro plays an important role in the rat ovulation process by attracting neutrophils. CINC/gro increased just prior to ovulation, and it may be regulated directly by cytokines such as IL-1beta and TNFalpha and indirectly by gonadotropins. PMID- 10859251 TI - Leptin gene expression, circulating leptin, and luteinizing hormone pulsatility are acutely responsive to short-term fasting in prepubertal heifers: relationships to circulating insulin and insulin-like growth factor I(1). AB - In the present study, we tested the hypothesis that short-term fasting would reduce leptin gene expression, circulating leptin, and LH pulsatility in prepubertal heifers in association with a decrease in circulating concentrations of insulin and insulin-like growth factor I (IGF-I). Twelve prepubertal crossbred heifers (mean +/- SD = 315 +/- 5 kg body weight) were assigned randomly to one of two treatments in two replicates: 1) control; normal feed consumption (n = 6) and 2) fasted; 48 h of total feed restriction (n = 6). Blood samples were collected at 15-min intervals for 8 h on Days 0 and 2 of the experiment and twice on Day 1. Subcutaneous fat samples were collected before treatment onset (Day -1) and at the end of the intensive blood sampling on Day 2. Acute feed restriction markedly reduced leptin mRNA in adipose tissue (P < 0.01) and circulating concentrations of leptin (P < 0.05), IGF-I (P < 0.01), and insulin (P = 0.05) as compared with controls on Day 2. Moreover, the treatment x day interaction (P < 0.076) and within-day contrasts (expressed as a percentage of Day 0 values) revealed that the mean frequency of LH pulses in the fasted group was lower (P < 0.06) than in controls on Day 2. Neither mean concentrations of growth hormone (GH) nor GH secretory dynamics were affected by acute feed restriction. Fasting-mediated decreases in leptin gene expression and circulating leptin, in association with reductions in secretion of IGF-I, insulin, and LH, provide a basis for investigating leptin as a hormone signaling energy status to the central reproductive axis in cattle. PMID- 10859252 TI - Hyaluronic acid as an anti-angiogenic shield in the preovulatory rat follicle. AB - Angiogenesis in the preovulatory follicle is confined to the theca cell layers, and penetration of capillaries through the basement membrane into the granulosa cell layers does not occur until after ovulation. However, elevated expression of the angiogenic growth factor (VEGF) has been reported in the cumulus cells surrounding the oocyte, which are expelled from the follicle during ovulation. This spatial and temporal discrepancy between VEGF expression and angiogenesis was studied here in the rat ovarian follicle, and we showed that cumulus cells secrete to the follicular fluid, in addition to VEGF, material with antiangiogenic activity that blocks endothelial cell proliferation, migration, and capillary formation in vitro. Hyaluronic acid produced by the cumulus cells can account for this antiangiogenic activity. Degradation of hyaluronic acid by hyaluronidase restored proliferation and migration of endothelial cells directed toward the cumulus. Inhibition of hyaluronic acid synthesis with 6-diazo-5-oxo-1 norleucine restored endothelial proliferation and migration in vitro, and it also resulted in early penetration of capillaries across the follicular basement membrane in vivo. These results support the role of hyaluronic acid produced by the cumulus cells as a high-molecular-weight, antiangiogenic shield that prevents premature vascularization of the preovulatory follicle by blocking endothelial cell migration and proliferation. PMID- 10859253 TI - Regulation of nitric oxide synthase to promote cytostasis in ovarian follicular development. AB - Our own recent studies have demonstrated that inducible nitric oxide synthase (iNOS) is predominantly localized in granulosa cells of healthy immature follicles in the rat ovary, whereas granulosa cells of either healthy mature follicles or follicles destined to be atretic are devoid of iNOS. These findings suggest that iNOS is pivotal for immature follicles to remain dormant. To test this hypothesis, we examined the effects of a GnRH agonist (buserelin), a proapoptotic substance, and epidermal growth factor (EGF), a mitogenic and, consequently, antiapoptotic factor, on the amount of iNOS mRNA in rat granulosa cells. Administration of buserelin in immature female rats transiently diminished iNOS mRNA levels in the ovaries as determined by Northern blot analysis. In cultured rat granulosa cells, buserelin and EGF increased the incidence of apoptosis and DNA synthesis, respectively, whereas both reduced iNOS mRNA levels as determined by reverse transcription-coupled polymerase chain reaction. The concomitant addition of S-nitroso-N-acetyl-DL-penicillamine, an NO donor, together with buserelin or EGF eliminated the observed effects of these substances (i.e., induction of apoptosis and stimulation of DNA synthesis, respectively). These results suggest that the changes in developmental status of immature follicles either into development or atresia are associated with reduced iNOS levels in granulosa cells, thus reinforcing the notion of NO as a cytostatic factor in ovarian follicles. PMID- 10859254 TI - Role of sperm sulfogalactosylglycerolipid in mouse sperm-zona pellucida binding. AB - Sulfogalactosylglycerolipid (SGG) is the major sulfoglycolipid of mammalian male germ cells. Like other sulfoglycolipids, SGG is believed to be involved in cell cell/extracellular matrix adhesion. Specifically, we investigated whether sperm SGG played a role in sperm-egg interaction. Initially, we produced an affinity purified, rabbit polyclonal immunoglobulin (Ig) G antibody that specifically recognized SGG (anti-SGG). Indirect immunofluorescence using anti-SGG IgG localized SGG to the convex and concave ridges and the postacrosome of the mouse sperm head. Pretreatment of sperm with anti-SGG IgG/Fab inhibited sperm-zona pellucida (ZP) binding in vitro in a concentration-dependent manner (to a maximum of 62%). This inhibition was observed at the level of primary binding. Sperm treated with anti-SGG IgG underwent the spontaneous and ZP-induced acrosome reaction at the same rate as control sperm treated with preimmune rabbit serum IgG. Fluorescently labeled SGG liposomes were shown to associate specifically with the egg ZP, whereas fluorescently labeled liposomes of galactosylglycerolipid (SGG's parental lipid) and phosphatidylserine (negatively charged like SGG) did not. Furthermore, coincubation of SGG liposomes with sperm and isolated ZP inhibited sperm-ZP binding in a concentration-dependent manner. These results strongly suggest an involvement of sperm SGG in direct binding to the ZP. PMID- 10859255 TI - Expression of RUSH transcription factors in developing and adult rabbit gonads. AB - The RUSH transcription factors 1alpha and 1beta bind to the Rabbit Uteroglobin promoter and are members of the SWI/SNF complex that facilitates transcription by remodeling chromatin (Helicase). To characterize gonadal expression of RUSH, a cRNA probe that recognizes both isoforms was used for in situ hybridization studies. We found RUSH mRNA to be abundant in Sertoli cells from embryonic, neonatal, prepubertal, and pubertal rabbit testes. In adults, RUSH mRNA was detected in tubules with preleptotene spermatocytes and mature spermatids lining the lumen. However, RUSH was undetectable in tubules that contained leptotene spermatocytes and that lacked mature spermatids. In females, RUSH was expressed in presumptive granulosa cells of embryonic and neonatal ovaries before follicle organization. Abundant RUSH mRNA was detected in granulosa and theca cells surrounding preantral follicles of prepubertal and adult ovaries. Expression of RUSH remained high in granulosa cells of antral follicles in mature ovaries but was negligible in late-stage atretic follicles and in corpora lutea. Western blot analysis confirmed the RUSH-1alpha isoform predominated in both testicular and ovarian tissues. The expression pattern of RUSH indicates transcriptional activity in Sertoli cells and during multiple stages of differentiating granulosa cells, especially those of primordial follicles, which heretofore were considered to be dormant. PMID- 10859256 TI - Studies on the onset of Leydig precursor cell differentiation in the prepubertal rat testis. AB - Leydig cells of the adult rat testis differentiate postnatally from spindle shaped cells in the testis interstitium during the neonatal-prepubertal period. Which spindle-shaped cell types are the precursor for Leydig cells and the stimulus for initiation of their differentiation are, however, two unresolved issues. In the present study, our objectives were to identify unequivocally which spindle-shaped cells are the precursors to Leydig cells and to test whether the initiation of their differentiation into Leydig cells depends on LH. Testes from fifteen groups of Sprague-Dawley rats (n = 4 per group) from 7-21 days of age were fixed in Bouin solution and embedded in paraffin. Immunoexpression of 3beta hydroxysteroid dehydrogenase (3betaHSD), cytochrome P450 side-chain cleavage (P450(scc)), 17alpha-hydroxylase cytochrome P450 (P450(c17)), and LH receptors (LHR) in interstitial cells (other than fetal Leydig cells) was observed using the avidin biotin method. Of all spindle-shaped cell types in the testis interstitium, only the peritubular mesenchymal cells showed positive immunolabeling for all three steroidogenic enzymes, beginning from the 11th postnatal day. All three enzymes were expressed simultaneously in these cells, and their numbers increased significantly thereafter. Immunoexpression of LHR in a few of these cells was just evident for the first time on postnatal Day 12 (i.e., after acquiring the steroidogenic enzyme activity). Their numbers gradually increased with time. The number of immunolabeled cells per 1000 interstitial cells (excluding fetal Leydig cells and capillary endothelial cells) was not significantly different for the three steroidogenic enzymes tested at all ages; however, a lower value was observed for LHR at each time-point. Based on these observations, we suggest that 1) the precursor cell type for the adult generation of Leydig cells in the postnatal rat testis is the peritubular mesenchymal cells, 2) precursor cells acquire 3beta-HSD, P450(scc), and P450(c17) enzyme activity simultaneously during Leydig cell differentiation, and 3) onset of precursor cell differentiation during Leydig cell development does not depend on LH. PMID- 10859257 TI - Molecular cloning and characterization of functional domains of a human testis specific isoform of calpastatin. AB - Human serum containing sperm-agglutinating antibodies was used to screen a testis cDNA expression library to identify the cognate antigens that may be responsible for this biological effect. The longest positive phage clone (1.9 kb) was sequenced and found to be a testis-specific isoform of calpastatin (tCAST). The testis-specific segment of tCAST is encoded by a single exon within intron 14 of the calpastatin gene. A unique protein isoform is produced that differs in domain structure from the somatic calpastatins (sCAST). Human sCAST most commonly has an N-terminal domain L plus the four functional calpain inhibitory domains. Human tCAST consists of a 40-amino-acid N-terminal T domain plus a part of domain II and all of domains III and IV from the somatic isoform. Our data show that the T domain can target cytosolic localization and membrane association of tCAST, whereas domain I of sCAST exhibits a nuclear localization function. Calpastatin is the endogenous inhibitor of calpain. The calpain/calpastatin system is involved in membrane fusion events for several cell types, and calpain has been localized to the sperm acrosome. We detected tCAST in human sperm and testes extracts by Western blotting with specific antisera. These observations suggest that tCAST may modulate calpain in the calcium-mediated acrosome reaction that is required for fertilization. PMID- 10859258 TI - Age-dependent changes in sperm production, semen quality, and testicular volume in the black-footed ferret (Mustela nigripes). AB - The black-footed ferret (Mustela nigripes), which was extirpated from its native North American prairie habitat during the 1980s, is being reintroduced to the wild because of a successful captive-breeding program. To enhance propagation, the reproductive biology of this endangered species is being studied intensively. The typical life span of the black-footed ferret is approximately 7 yr. Female fecundity declines after 3 yr of age, but the influence of age on male reproduction is unknown. In this study, testis volume, seminal traits, sperm morphology, and serum testosterone were compared in 116 males from 1 to 7 yr of age living in captivity. Results demonstrated that testes volume during the peak breeding season was similar (P > 0.05) among males 1 to 5 yr of age, reduced (P < 0.05) among males 6 yr of age, and further reduced (P < 0.05) among males 7 yr of age. Motile sperm/ejaculate was similar in males 1 to 6 yr of age but diminished (P < 0.05) in those 7 yr of age. Males at 6 and 7 yr of age produced fewer (P < 0.05) structurally normal sperm than younger counterparts; however, serum testosterone concentrations were not reduced (P > 0.05) in older males. Histological comparison of testicular/epididymal tissue from 5- and 7-yr-old black-footed ferrets confirmed that the interval between these two ages may represent a transitional period to reproductive senescence. In summary, functional reproductive capacity of male black-footed ferrets exceeds that of females by at least 2 yr. Testes and seminal quality are indistinguishable among males 1 to 5 yr of age, with progressive reproductive aging occurring thereafter. PMID- 10859259 TI - Regression of the decidualized mesometrium and decidual cell apoptosis are associated with a shift in expression of Bcl2 family members. AB - The purpose of this study was to determine whether regression of the decidua basalis (DB), which begins on Day 14 of pregnancy in the rat, results from an intrinsic program of apoptosis regulated by Bax and Bcl2. Expression of Bax and Bcl2 and the incidence of apoptosis were evaluated throughout gestation by Western blot analysis and detection of DNA fragments. Antiprogestin (RU486) was also administered during proliferation of DB to study progesterone regulation of Bax/Bcl2 balance. Bax, the pro-apoptotic protein, was expressed at a low level throughout pregnancy, whereas Bcl2, the pro-survival partner, was most abundantly expressed on Days 8 and 10, which are a time of proliferation and decidualization, and declined to barely detectable levels thereafter. These changes resulted in a 12-fold increase in the Bax:Bcl2 ratio on Day 17 as compared with Day 8 of pregnancy (P < 0.05). DNA laddering and in situ staining of DNA fragments first became visible on Day 14 and involved 2% of cells by Days 17 and 21 (P < 0.05). Treatment with RU486 on Day 9 enhanced Bax and suppressed Bcl2 within 6 h, increasing the Bax:Bcl2 ratio sixfold (P < 0.05). Apoptosis was minimal at 6 h and increased to 9% of cells by 24 h (P < 0.05). Thus, progesterone appears to regulate the apoptotic threshold of stromal cells by modulating Bax and Bcl2 expression. PMID- 10859260 TI - Novel thiourea compounds as dual-function microbicides. AB - Sexually active women represent the fastest growing human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome risk group. In an effort to develop a vaginal microbicidal contraceptive potentially capable of preventing HIV transmission as well as providing fertility control, we previously reported the synthesis of novel nonnucleoside inhibitors (NNIs) of HIV-1 reverse transcriptase with sperm-immobilizing activity (SIA). To gain further insight into the structure-function relationship controlling these two properties of NNIs, we have rationally designed and synthesized 30 novel thiourea compounds and examined them for dual-function, anti-HIV and spermicidal activity. Twelve of the 30 thiourea compounds exhibited potent anti-HIV activity in the nanomolar range (IC(50) = <1 9 nM). Nine of the 30 thiourea derivatives exhibited both anti-HIV and spermicidal activity. Among the phenyl ring-containing thioureas, the 2-fluoro (HI-240) -substituted and 2-chloro (HI-253) -substituted derivatives exhibited potent anti-HIV activity (IC(50) = <1 nM) with SIA (EC(50) = 70 microM and 147 microM). Among the alicyclic ring-containing thioureas, the 5-bromo (HI-346) and 5-chloro (HI-445) functionalized cyclohexenyl ring-substituted thioureas were the most potent dual-function spermicides (EC(50) = 42 and 57 microM), with anti-HIV activity at nanomolar range (IC(50) = 3 nM). Unlike nonoxynol-9 (N-9), none of the potent dual-function thiourea compounds were cytotoxic to normal human vaginal, ectocervical, and endocervical epithelial cells at spermicidal concentrations. We conclude that as potent anti-HIV agents with SIA and reduced cytotoxicity when compared with N-9, the phenyl-substituted and cyclohexenyl substituted thiourea derivatives, especially compounds HI-253 (N-[2-(2 chlorophenethyl)]-N'-[2-(5-bromopyridyl)-thiourea), HI-346 (N-[2-(5 bromopyridinyl)]-N'-[2-(1-cyclohexenyl)ethyl-thiourea), and HI-445 (N-[2-(5 chloropyridinyl)]-N'-[2-(1-cyclohexenyl)ethyl-thiourea) show unique clinical potential to become the active ingredients of a vaginal contraceptive for women who are at high risk for acquiring HIV by heterosexual vaginal transmission. PMID- 10859261 TI - Pituitary adenylate cyclase-activating peptide stimulates acute progesterone production in rat granulosa/Lutein cells via two receptor subtypes. AB - Pituitary adenylate cyclase-activating peptide (PACAP) is transiently expressed in ovarian granulosa/lutein cells from eCG/hCG-treated rats, and in vitro immunoneutralization of endogenously released PACAP inhibits acute progesterone secretion and subsequent luteinization in such cells. This suggests that PACAP mediates locally some of the effects of the LH surge, but the putative PACAP receptor(s) involved in such an auto or paracrine activity is presently unknown. Reverse-transcription polymerase chain reaction with specific primers to the three cloned PACAP-binding receptors called PAC(1), VPAC(1), and VPAC(2) demonstrated both PAC(1) and VPAC(2) mRNA in extracts from preovulatory follicular cells. Radioligand-binding assays revealed the presence of high affinity binding sites with characteristics of these two receptors on the intact cells, and autoradiography demonstrated that the binding was restricted to a minor proportion of the follicular cells as well as the oocytes. Pituitary adenylate cyclase-activating peptide and vasoactive intestinal peptide (VIP) dose dependently stimulated cAMP accumulation and acute progesterone accumulation. Forskolin and db-cAMP also stimulated acute progesterone accumulation, and the protein kinase A inhibitor H89 dose-dependently inhibited peptide induced acute progesterone accumulation, suggesting involvement of cAMP and the protein kinase A pathway in the process. In conclusion, two of the three PACAP binding receptors are present on preovulatory follicular cells and are involved in the effects of PACAP on acute progesterone production. The data provide further evidence to establish PACAP as an auto- or paracrine regulator of LH-induced acute progesterone production in rat preovulatory follicles. PMID- 10859262 TI - Measurement of dimeric inhibins and effects of active immunization against inhibin alpha-subunit on plasma hormones and testis morphology in the developing cockerel. AB - Inhibins and activins are implicated as endocrine regulators of follicle stimulating hormone production and of testicular steroidogenesis and spermatogenesis in mammals. The potential involvement of these proteins in cockerels was investigated by measurement of circulating inhibin A, inhibin B, total inhibin alpha-subunit immunoreactivity (ir-alpha), activin A, LH, FSH, and testosterone from the juvenile state through to sexual maturity. Plasma inhibin A remained low between 6 to 12 wk of age and increased approximately threefold (P < 0.05) to a prepubertal peak between Weeks 14 to 18, followed by a gradual decline to the end of the study (Week 24). Although plasma FSH levels were not correlated to inhibin A before Week 16 (r = -0.17), they were negatively correlated from Week 18 (r = -0.49; P < 0.005). Inhibin B levels were below the assay detection limit until 16 wk of age but thereafter rose steadily in parallel with FSH (r = 0.27; P < 0.02) and testosterone (r = 0.35; P < 0.005). Thus, inhibins A and B showed divergent profiles during sexual maturation. Plasma ir-alpha levels were much higher than dimeric inhibin levels throughout, although the relative difference varied with age. Plasma activin A levels were below the assay detection at all times. Juvenile cockerels were actively immunized against a synthetic chicken inhibin alpha-subunit peptide conjugate to determine effects on plasma hormones and on testicular weight, morphology, and activin A content. Immunization generated circulating antibodies that bound (125)I-bovine 32-kDa inhibin but did not affect plasma FSH or testosterone levels at any stage of development. However, immunization reduced postpubertal plasma LH levels (P < 0.05) and promoted increased testicular weight (24%; P < 0.01) and total testicular activin A content (42%; P < 0.001) at 24 wk. Testis weight of immunized birds was positively correlated with inhibin antibody titer (r = 0.61; P < 0.05). Live weight gain was not affected by immunization. Morphometric analysis of testis sections showed that inhibin immunization had no effect on the fractional volume of the seminiferous tubule wall, seminiferous tubule lumen, or interstitial tissue area. Likewise, seminiferous tubule surface area and surface area:volume ratios were not different from controls. These findings support differential roles for inhibins A and B in regulating the pituitary-testicular axis during sexual maturation in the cockerel but highlight the need for more detailed studies to distinguish between potential endocrine and local intragonadal roles of inhibin-related peptides and to elucidate the mechanism by which immunization against inhibin alpha-subunit promotes testis enlargement without raising plasma FSH. PMID- 10859263 TI - Phylogenetic analysis of the vertebrate glycoprotein hormone family including new sequences of sturgeon (Acipenser baeri) beta subunits of the two gonadotropins and the thyroid-stimulating hormone. AB - The beta subunits of the two gonadotropins (GTH1 and GTH2) and of the thyroid stimulating hormone (TSH) of a chondrostean fish, Acipenser baeri, were cloned. These new sequences and selected representative members of beta subunits of vertebrate glycoprotein hormones, including tetrapod follicle-stimulating hormones (FSH) and luteinizing hormones (LH), allowed us to infer the phylogenetic relationships within this family. Both distance matrix and maximum parsimony methods were used on both nucleotide and amino acid sequences, with bootstrapping evaluation over 1000 replicates. The four trees obtained had highly similar topologies. In each case, three monophylogenetic lineages, TSH, GTH1-FSH, and GTH2-LH were clearly identified. The three monophylogenetic lineages were supported by 21-23 specific characters at the amino acid level, out of a total of 121 characters. The resolved topologies within each monophyletic hormone cluster were congruent with the known phylogenetic relationships between the related species. The inferred parental relationships within gonadotropins are in agreement with data concerning their biological functions. The present study demonstrates that GTH1 and GTH2 are the actinopterygian homologues of tetrapod FSH and LH, respectively. PMID- 10859264 TI - Insulin-like growth factor receptors and their ligands in gonads of a hermaphroditic species, the gilthead seabream (Sparus aurata): expression and cellular localization. AB - Expression of insulin-like growth factor (IGF)-I, IGF-II, and IGF type I receptor (IGF-1R) genes was studied in gonads at different developmental stages of the protandrous hermaphroditic species the gilthead seabream (Sparus aurata) by reverse transcription-polymerase chain reaction and Northern blot analysis. Both IGF-I and IGF-II mRNA levels were highest in bisexual gonads and decreased during gonadal development. Regardless of the stage of gametogenesis, IGF-II mRNA levels exceeded those of IGF-I. Transcripts for IGF-1R RNA were detected in gonads at all stages studied. A major transcript of 11 kb was found in gonads and in gill arch and brain, but it was not found in liver and muscle. Distribution of the two types of IGF-1R and IGF-I in gonads was studied by immunohistochemistry. Immunoreactive IGF-I was found in the granulosa and theca cells of follicles at different vitellogenic stages and in oocytes at the chromatin-nucleolus and perinucleolus stage. In the testis, immunoreactive IGF-I was found in somatic cells of the cyst wall, interstitial cells, and spermatogonia A. In addition, IGF 1R was detected in the membrane of previtellogenic oocytes and in the theca and granulosa cells of vitellogenic and late vitellogenic follicles. In the testis, a positive reaction was identified in spermatogonia A and spermatids for the germ cells and in somatic cells of the cyst walls and interstitial cells. Local expression and production of IGFs and their receptors in fish gonads support a role for the IGF system in fish gonadal physiology. PMID- 10859265 TI - Effects of the porcine oviduct-specific glycoprotein on fertilization, polyspermy, and embryonic development in vitro. AB - This study evaluated the effects of porcine oviduct-specific glycoprotein (pOSP) on in vitro fertilization (IVF), polyspermy, and development to blastocyst. Experiment 1 evaluated the effects of various concentrations (0-100 microgram/ml) of purified pOSP on fertilization parameters, including penetration, polyspermy, male pronuclear formation, and mean number of sperm penetrated per oocyte. Experiment 2 examined the ability of an anti-pOSP immunoglobulin G to inhibit the observed effects of pOSP on fertilization parameters. Experiments 3 and 4 examined various concentrations of pOSP (0-100 microgram/ml) on zona pellucida solubility and sperm binding, respectively. Lastly, experiment 5 assessed the effects of various concentrations of pOSP (0-100 microgram/ml) on the in vitro embryo cleavage rate and development to blastocyst. Pig oocytes matured and fertilized in vitro were used for all experiments. An effect of treatment (P < 0.05) was detected for pOSP on penetration, polyspermy, and mean number of sperm per oocyte. Concentrations for pOSP of 0-50 microgram/ml had no effect on sperm penetration rates; however, compared with the control, 100 microgram/ml significantly decreased the penetration rate (74% vs. 41%). Addition of 10-100 microgram/ml significantly reduced the polyspermy rate compared with the control (61% vs. 24-29%). The decrease in polyspermy achieved by addition of pOSP during preincubation and IVF was blocked with a specific antibody to pOSP. No effect of treatment was observed on zona digestion time relative to the control; however, the number of sperm bound to the zona pellucida was significantly decreased by treatment (P < 0.05). Compared with the control, all concentrations of pOSP examined reduced the number of sperm bound per oocyte (45 vs. 19-34). A treatment effect (P < 0.05) was observed for pOSP on embryo development to blastocyst but not on cleavage rates. Addition of pOSP during preincubation and fertilization significantly increased postcleavage development to blastocyst, but a synergistic stimulation on development was not detected when pOSP was included during in vitro culture. These results indicate that exposure to pOSP before and during fertilization reduces the incidence of polyspermy in pig oocytes, reduces the number of bound sperm, and increases postcleavage development to blastocyst. PMID- 10859266 TI - Increased birefringence in the meiotic spindle provides a new marker for the onset of activation in living oocytes. AB - The newly developed Pol-Scope allows imaging of spindle retardance, which is an optical property of organized macromolecular structures that can be observed in living cells without fixation or staining. Experiments were undertaken to examine changes in meiotic spindles during the initial stages of activation of living mouse oocytes using the Pol-Scope. Parthenogenetic activation of oocytes treated with calcium ionophore evoked a dynamic increase in meiotic spindle retardance, particularly of the midregion, before spindle rotation and second polar body extrusion. The pronounced increase in spindle retardance, which could, for the first time to our knowledge, be quantified in living oocytes, was maintained during polar body extrusion. Spindle retardance of newly in vivo fertilized oocytes was significantly higher than that of ovulated, metaphase II oocytes. Pol Scope imaging of fertilized oocytes did not affect subsequent development. These results establish that increased spindle retardance precedes polar body extrusion and pronuclear formation. The increased birefringence in the spindle provides an early indicator of oocyte activation. Thus, noninvasive, quantitative imaging of the onset of activation in living oocytes might improve the efficiency of assisted fertilization and other embryo technologies. PMID- 10859267 TI - Inhibition of rainbow trout (Oncorhynchus mykiss) estrogen receptor activity by cadmium. AB - This study was conducted to determine if the cadmium-mediated inhibition of vitellogenesis observed in fish collected from contaminated areas or undergoing experimental exposure to cadmium correlated with modification in the transcriptional activity of the estrogen receptor. A recombinant yeast system expressing rainbow trout (Oncorhynchus mykiss) estradiol receptor or human estradiol receptor was used to evaluate the direct effect of cadmium exposure on estradiol receptor transcriptional activity. In recombinant yeast, cadmium reduced the estradiol-stimulated transcription of an estrogen-responsive reporter gene. In vitro-binding assays indicated that cadmium did not affect ligand binding to the receptor. Yeast one- and two-hybrid assays showed that estradiol induced conformational changes and receptor dimerization were not affected by cadmium; conversely, DNA binding of the estradiol receptor to its cognate element was dramatically reduced in gel retardation assay. This study provides mechanistic data supporting the idea that cadmium is an important endocrine disrupter through a direct effect on estradiol receptor transcriptional activity and may affect a number of estrogen signaling pathways. PMID- 10859268 TI - Culture of bovine preantral follicles in a serum-free system: markers for assessment of growth and development. AB - Satisfactory development of bovine follicles in vitro remains elusive. This study used a serum-free system to evaluate the effects of insulin-like growth factor-1 (IGF-1) on bovine preantral follicles in culture and to identify the activity of gelatinase matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) in vitro to assess their potential as markers of development. Preantral follicles were cultured for 6 days in serum-free medium containing insulin and IGF-1 (10 ng/ml). No difference was observed in follicular growth, health, or antrum formation between IGF-1-treated follicles and controls. However, IGF-1 had a negative effect (P < 0.01) on oocyte size and granulosa cell proliferation. When MMP-9 was secreted, the probability of follicles having healthy granulosa or theca cells at the end of the culture period was 0.85 and 0.60, respectively. If TIMP-1 was released, the probability of follicles having healthy somatic cells was 0.79. When TIMP-2 was detected, the probability of granulosa and theca cell health was 0. 78 and 0.67, respectively. These results demonstrate no positive effects of IGF-1 on bovine follicles in this system. Furthermore, MMP-9 and TIMPs are related to follicular health and, therefore, can be used as markers of follicular development. PMID- 10859269 TI - Steroid-dependent up-regulation of adipose leptin secretion in vitro during pregnancy in mice. AB - Circulating leptin levels are elevated during the later stages of pregnancy in mammals, suggesting that maternal leptin may play a role in maintenance of pregnancy and/or preparation for parturition and lactation. The regulation and source of circulating leptin during pregnancy remains undetermined, but leptin mRNA levels increase in adipose tissue during this time in some species. Considerable controversy exists whether placenta is also a leptin-secreting tissue during pregnancy. Here, we directly demonstrate that leptin secretion rates from mouse adipose tissue in vitro are decreased during early pregnancy and up-regulated during late pregnancy and lactation. Changes in leptin secretion rates in vitro paralleled those of circulating leptin in vivo during gestation. Subcutaneous implants of estradiol or corticosterone into lactating mice for 48 h stimulated adipose leptin secretion rates in vitro to the level of that in pregnant mice. However, corticosterone, but not estradiol, increased leptin secretion when added to isolated adipose tissue in vitro. Placentae obtained at two stages of pregnancy did not secrete leptin in vitro, either when acutely isolated or when dissociated into cells for long-term cultures. Placental tissue (or cells) secreted progesterone, however, demonstrating placental viability. We conclude that hyperleptinemia during late pregnancy in mice primarily results from corticosterone-dependent up-regulation of leptin secretion from adipose tissue, and that the placenta does not contribute to leptin secretion. The initial decrease in leptin secretory rates from adipose tissue during early pregnancy may facilitate energy storage for the subsequent, increased metabolic demands of later pregnancy and lactation. PMID- 10859270 TI - Amino acids and preimplantation development of the mouse in protein-free potassium simplex optimized medium. AB - Development of outbred CF1 mouse zygotes in vitro was studied in a chemically defined, protein-free medium both with and without amino acids. The addition of amino acids to protein-free potassium simplex optimized medium (KSOM) had little effect on the proportion of embryos that developed at least to the zona-enclosed blastocyst stage. In contrast, amino acids stimulated very significantly, in a dilution-dependent way, the proportion of blastocysts that at least partially or completely hatched. Amino acids also stimulated cell proliferation in both the trophectoderm and inner cell mass (ICM) cells, at rates that favored proliferation of cells in the ICM; had no effect on the incidence of cell death (oncosis or apoptosis); and improved development of the basement membranes, which form on the blastocoelic surface of the trophectoderm and between the primitive endoderm and the primitive ectoderm. Thus, KSOM, supplemented with amino acids but containing no protein supplements, supports development of a newly fertilized ovum to the late blastocyst stage, in which its normal, three-dimensional structure is preserved and in which the ICM has been partitioned into the primitive ectoderm and primitive endoderm. PMID- 10859271 TI - Episialin acts as an antiadhesive factor in an in vitro model of human endometrial-blastocyst attachment. AB - Episialin, which is found on the apical membrane of human endometrial epithelium, has been postulated to act as an antiadhesive factor through the steric hindrance generated by its extensively glycosylated structure. The present studies were designed to test this hypothesis in an in vitro model of endometrial-blastocyst attachment. Episialin was expressed in human endometrial carcinoma cells (HEC-1A > RL95-2), and attachment of JAr choriocarcinoma cells to the endometrial cell monolayers was inversely related to episialin expression. Treatment of endometrial monolayers with type III sialidase increased JAr binding, and this increase was suppressed by HMFG1, a monoclonal antibody specific for episialin. The effects of sialidase appear to have resulted from a contaminant protease rather than from a loss of sialic acid residues, because sialidase preparations other than type III were ineffective. After sialidase treatment, conditioned medium from cells treated with type III sialidase contained more episialin than medium from cells treated with other sialidase preparations. Similar attachment assay results were obtained using O-sialoglycoprotein endopeptidase; after treatment, the increase in JAr binding (>50%) was suppressed by the antiepisialin antibody. These results demonstrate for the first time that episialin acts as an antiadhesive agent in a model of human endometrial-blastocyst attachment. PMID- 10859272 TI - Changes in uterine expression of leukemia inhibitory factor receptor gene during pregnancy and its up-regulation by prolactin in the western spotted skunk. AB - The multifunctional cytokine leukemia inhibitory factor (LIF) is presumed to participate in preparing the uterus for blastocyst implantation. Increased production of LIF is positively correlated with termination of embryonic diapause and preparation for implantation in the spotted skunk. This study examined changes in the expression, localization, and hormonal regulation of LIF receptor (LIFRbeta) gene expression in the uterus of the skunk. Changes in the uterine concentration of LIFRbeta mRNA during pregnancy or in response to hormones after ovariectomy were determined by Northern hybridization analysis and reverse transcriptase polymerase chain reaction. The skunk uterus produces two LIFRbeta transcripts, the levels of which increase in concentration when the blastocysts resume their development but then decline somewhat during the latter stage of blastocyst activation. Ovariectomy significantly reduced uterine LIFRbeta expression. Estradiol and/or progesterone failed to significantly elevate LIFRbeta mRNA levels in ovariectomized animals. Prolactin significantly increased uterine concentrations of LIFRbeta mRNA to greater than those of ovariectomized controls, but these levels were not comparable to those observed during preimplantation. The LIFRbeta mRNA was predominately localized to stromal cells surrounding the uterine glands and in yolk sac endoderm, syncytiotrophoblast, and cytotrophoblast of postimplantation embryos. PMID- 10859273 TI - Unique expression of gonadotropin-I and -II subunit genes in male and female red seabream (Pagrus major) during sexual maturation. AB - Two distinct gonadotropins (GTHs) have been demonstrated in a number of teleost fishes. Although the physiological roles of GTHs have been extensively studied in salmonids, little is known about their biological functions in nonsalmonid fishes. In this study, to elucidate the role of GTH-I and GTH-II in reproduction, we cloned the alpha-glycoprotein subunit (alphaGSU) and gonadotropin beta subunits (Ibeta and IIbeta) of red seabream using the 5'- and 3'-RACE methods and used these cDNA probes to reveal changes in mRNA levels of each subunit during sexual maturation of both male and female red seabream. The nucleotide sequences of alphaGSU, Ibeta, and IIbeta are 629, 531, and 557 base pairs long, encoding peptides of 117, 120, and 146 amino acids, respectively. The deduced amino acid sequence of each mature subunit showed high homology with those of other teleosts. Northern blot analysis showed that Ibeta mRNA levels of males increase in association with gonadal development, whereas those of females remain low throughout sexual maturation, indicating sexual dimorphism in the expression pattern of Ibeta. In contrast, IIbeta mRNA levels of both sexes are maintained at high levels from the beginning of gametogenesis to spawning season. These results are different than those of salmonids and suggest that GTH-I may have important roles in male, but not female, gametogenesis. GTH-II may be involved in regulation of early and late gametogenesis in both male and female red seabream. PMID- 10859274 TI - Luteinizing hormone has a stage-limited effect on preantral follicle development in vitro. AB - Although it is known that LH receptors are present from the time of thecal differentiation, the role of LH during early follicle development is not yet clear. The effect of LH on preantral follicle development has therefore been investigated in vitro using a culture system that supports the development of intact follicles. We have previously shown that although preantral follicles 150 micrometer in diameter (2-3 granulosa cell layers) do not require LH to proceed through antral development, smaller follicles (1-2 granulosa cell layers, 85-110 micrometer in diameter) do not develop beyond the large preantral stage in the presence of only FSH and 5% mouse serum. Follicles of this size were therefore used to determine the effects of LH and serum on their development in vitro. The results showed that although FSH must be continuously present, a low concentration of LH together with a slight increase in serum concentration was necessary, specifically during the primary stage of follicle development (from 85 micrometer in diameter until the follicles had reached 150 micrometer in diameter) to induce the capacity for subsequent LH-independent rapid growth and antral development. The in vitro development of maturable oocytes with normal spindle and chromatin morphology was also supported. These results indicate that LH probably induces changes in the early differentiating thecal cells, which are critical for the completion of subsequent follicular and oocyte development. PMID- 10859275 TI - Postnatal growth and behavioral development of mice cloned from adult cumulus cells. AB - Since the first successful cloning of mammals from adult somatic cells, there has been no examination of the learning or behavior of cloned offspring. The possibility of adverse effects on animals produced through adult somatic cell cloning is high because many natural biological processes are bypassed and DNA from adult cells, which presumably contain mutations, are used. In this study, we compared cloned mice produced by microinjection transfer of cumulus cell nuclei into enucleated oocytes, to control mice that were specifically generated to eliminate confounding factors that are unique to our cloning procedure. Postnatal weight gain of clones was significantly greater than that of controls. Preweaning development observations revealed that first appearance or performance of 3 out of 10 measures was delayed in cloned mice; however, results of subsequent tests of learning and memory, activity level, and motor skills were comparable for both groups. Together, these data suggest that nuclear transfer of adult somatic cell nuclei to produce cloned mice may delay the appearance of a few developmental milestones but it does not adversely affect the overall postnatal behavior of mice. In addition, this procedure may cause late onset of significantly increased body weight in cloned offspring, the cause or causes of which are being further examined. PMID- 10859276 TI - Photoperiodic versus metabolic signals as determinants of seasonal anestrus in the mare. AB - The objectives of this study were to compare the timing and mechanisms controlling the onset of anestrus in young and mature mares treated either continuously with melatonin and in those that remained untreated. Changes in body weight, subcutaneous body fat measured to provide an estimate of total body fat, and circulating concentrations of leptin were compared throughout the 1-yr experimental period. The results demonstrate that in young mares the timing of anestrus occurs significantly earlier in the year than in mature mares and that mature mares are more likely to exhibit continuous reproductive activity during the nonbreeding season. The propensity of mature mares to exhibit this phenomenon is not modified by continuous treatment with melatonin but is associated with higher mean circulating concentrations of leptin, body weight, and estimated percent of body fat. In both young and mature mares, body weight, percent of body fat, and circulating concentrations of leptin are higher during summer than winter months. We conclude that, in the mare, the reproductive response to a decrease in photoperiod or a presumptive inhibitory melatonin signal is modified by energy availability, which may be signaled to the hypothalamus-pituitary axis via a change in the circulating concentration of leptin. An additional observation confirmed that the prolactin axis is responsive to continuous treatment with melatonin but that a suppression of prolactin secretion is limited to the spring months. PMID- 10859277 TI - Sonication per se is not as deleterious to sperm chromosomes as previously inferred. AB - Although sonication is a simple way to immobilize ("kill") spermatozoa prior to injection into oocytes, this has been thought to be destructive to sperm chromosomes. Mouse and human spermatozoa were immobilized by sonication and kept in various media for up to 2 h, then their nuclei were individually injected into mouse oocytes for the analysis of chromosomes at the first cleavage metaphase. In both the mouse and human, incidence of structural chromosome aberrations was much higher in the spermatozoa sonicated and stored in Biggers-Whitten-Whittingham medium for 2 h at 37.5 degrees C than in those stored for 5 min in the same medium. We concluded, therefore, that it is not sonication per se but a prolonged exposure of sperm nuclei to extracellular milieu that is detrimental to sperm chromosomes. The incidence of structural chromosome aberrations of mouse and human spermatozoa was significantly reduced when the spermatozoa were sonicated and stored in K(+)-rich nucleus isolation medium containing EDTA. This suggests that sperm chromosome degradation following sperm immobilization by sonication is partly due to detrimental effects of a Na(+)-rich medium and of DNase on sperm chromatin. Ideally, it should be possible to prepare artificial media that maintain the integrity of sperm chromosomes for many hours after immobilization. PMID- 10859279 TI - June 20, 2000 PMID- 10859278 TI - Immunization of male mice with luteinizing hormone-releasing hormone fusion proteins reduces testicular and accessory sex gland function. AB - Genes for ovalbumin-luteinizing hormone-releasing hormone 7 (LHRH-7) and thioredoxin-LHRH-7 fusion proteins (containing seven LHRH inserts) were constructed by cassette and mismatch mutagenesis and expressed in Escherichia coli. In experiment 1, 10 microgram of either ovalbumin-LHRH-7 or thioredoxin LHRH-7 were suspended in Z-max adjuvant and injected three times at 4-wk intervals into postpubertal male BALB/c mice. In experiment 2, the fusion proteins were suspended in Immumax adjuvant and administered in equimolar quantities (0.4 nmol per injection) to postpubertal male BALB/c mice. In addition to injection of these two proteins alone, the proteins were also administered in different sequences or together in a mixture. Both LHRH fusion proteins induced significant antibody titers, which resulted in a significant decrease in vesicular gland and anterior prostate weight (measure of biological response) in both experiments. Vesicular gland and anterior prostate weight and LHRH antibody titers were significantly correlated in experiments 1 (r = -0.64) and 2 (r = 0.53). Percentage of animals responding to treatment varied from 40-60% in experiment 1 and from 11-89% in experiment 2, with the highest responses in treatments that used a combination of both fusion proteins. The variation in responders and nonresponders was evaluated by estimating antibody K(D) from displacement curves. Part, but not all, of the high antibody nonresponders can be explained by antibody affinity. PMID- 10859280 TI - Gas exchange efficiency in congestive heart failure. PMID- 10859281 TI - Cardiovascular disease mortality in familial forms of hypertriglyceridemia: A 20 year prospective study. AB - BACKGROUND: Familial combined hyperlipidemia (FCHL) and familial hypertriglyceridemia (FHTG) are 2 of the most common familial forms of hyperlipidemia. There is a paucity of prospective data concerning the risk of cardiovascular disease (CVD) in such families. The purposes of this study were to estimate 20-year total and CVD mortality risk among relatives in these families and to evaluate plasma triglyceride as a predictor of death. METHODS AND RESULTS: The study was based on lipid and medical history data from 101 families ascertained in 2 studies conducted in the early 1970s. Vital status and cause of death was determined during 1993 to 1997 for 685 family members, including first degree relatives of the probands and spouse control subjects. Compared with spouse control subjects, 20-year CVD mortality risk was increased among siblings and offspring in FCHL (relative risk 1.7, P=0.02) after adjustment for baseline covariates. In FHTG families, the relative risk was also 1.7 but was not statistically significant (P=0.39). Baseline triglyceride was associated with increased CVD mortality risk independent of total cholesterol among relatives in FHTG families (relative risk 2.7, P=0.02) but not in FCHL families (relative risk 1.5, P=0.16) after adjustment for baseline covariates. CONCLUSIONS: This prospective study establishes that relatives in FCHL families are at increased risk for CVD mortality and illustrates the need for effective prevention strategies in this group. Baseline triglyceride level predicted subsequent CVD mortality among relatives in FHTG families, adding to the growing evidence for the importance of hypertriglyceridemia as a risk factor for CVD. PMID- 10859282 TI - Association of a T29-->C polymorphism of the transforming growth factor-beta1 gene with genetic susceptibility to myocardial infarction in Japanese. AB - BACKGROUND: Transforming growth factor-beta (TGF-beta) is an important regulator of vascular remodeling and is involved in the pathogenesis of atherosclerosis. A T-->C transition at nucleotide 29 of the TGF-beta1 gene results in a Leu-->Pro substitution at amino acid 10 of the signal peptide. We have now examined a possible association of TGF-beta1 genotype with myocardial infarction (MI) in a Japanese population. METHODS AND RESULTS: TGF-beta1 genotype was determined in 315 Japanese patients (234 men and 81 women) with MI and 591 control subjects (289 men and 302 women). We found that age, body mass index, and incidence of habitual smoking, hypertension, diabetes mellitus, and hypercholesterolemia did not differ between the 2 groups for either men or women. Multivariable logistic regression analysis, however, demonstrated the frequency of the T allele to be significantly higher in male subjects with MI than in controls (TT + TC versus CC; P<0.0001, odds ratio 3.5, 95% CI 2.0 to 6.3). In contrast, the T allele was not associated with the prevalence of MI in women. In both male MI patients and controls, the serum concentration of TGF-beta1 was significantly higher in individuals with the CC genotype than in subjects with the TT or TC genotype. CONCLUSIONS: Findings suggest that the T allele at nucleotide 29 in the TGF-beta1 gene is a risk factor for genetic susceptibility to MI, at least in middle-aged Japanese men. PMID- 10859283 TI - Trial of abciximab with and without low-dose reteplase for acute myocardial infarction. Strategies for Patency Enhancement in the Emergency Department (SPEED) Group. AB - BACKGROUND: Low-dose alteplase with standard-dose abciximab enhances reperfusion 90 minutes after acute myocardial infarction (MI). We combined standard-dose abciximab with low-dose reteplase for acute MI in 2 phases. Two heparin doses were also explored. METHODS AND RESULTS: Phase A patients were randomized 4:1 to receive an abciximab bolus with infusion alone (n=63) or with 5 U, 7.5 U, 10 U, 5 U+2.5 U, or 5 U+5 U of reteplase (total n=241). Phase B tested the best phase A strategy (abciximab plus 5 U+5 U reteplase, expressed as abciximab-reteplase 5+5 U; n=115) against 10 U+10 U reteplase alone (n=109). The primary end point was Thrombolysis In Myocardial Infarction (TIMI) grade 3 flow at 60 to 90 minutes. In phase A, 62% of the abciximab-reteplase 5+5 U group had TIMI grade 3 flow versus 27% of the abciximab-only patients (P=0.001). In phase B, 54% of the abciximab reteplase 5+5 U group had grade 3 flow versus 47% of the reteplase-only patients (P=0.32). Grade 3 flow rates were 61% for a 60 U/kg heparin bolus and abciximab reteplase 5+5 U, 51% for a 40 U/kg heparin bolus and abciximab-reteplase 5+5 U (P=0.22), and 47% for reteplase alone (P=0.05 versus the 60 U/kg heparin group). Major bleeding rates in phase A were 3.3% for abciximab alone and 5.3% for abciximab-reteplase 5+5 U; rates in phase B were 9.8% for abciximab-reteplase 5+5 U and 3.7% for reteplase alone. Major bleeding was similar with standard- or low dose heparin (6.3% versus 10.5%, P=0.30). CONCLUSIONS: In this phase II trial, adding reteplase to abciximab treatment of acute MI versus reteplase alone enhanced the incidence of early complete reperfusion after the initiation of therapy in the emergency department. PMID- 10859284 TI - Long-term clinical outcome in the Bypass Angioplasty Revascularization Investigation Registry: comparison with the randomized trial. BARI Investigators. AB - BACKGROUND: The Bypass Angioplasty Revascularization Investigation (BARI) included 4039 patients with multivessel coronary artery disease; 1829 consented to randomization, and 2010 did not but were followed up in a registry. Thus, we can evaluate the outcome of physician-guided versus random assignment of percutaneous transluminal coronary angioplasty (PTCA) versus coronary artery bypass graft surgery (CABG). METHODS AND RESULTS: We compared the baseline features and outcomes for PTCA and CABG in the overall registry and its predesignated subgroups. We assessed the impact of treatment by choice versus random assignment by comparing the results in the registry with those of the randomized trial. Statistical adjustments for differences in baseline characteristics were made. Within the registry, nearly twice as many patients were selected for PTCA (1189) as CABG (625); mortality at 7 years was similar for PTCA (13.9%) and CABG (14.2%) (P=0.66) before and after adjustment for baseline differences between patients selected for PTCA versus CABG (adjusted RR, 1.02; P=0.86). In contrast to the randomized trial, the 7-year mortality rate of treated diabetics in the registry was equally high (26%) with PTCA or CABG. Seven year mortality was higher for patients undergoing PTCA in the randomized trial than in the registry (19.1% versus 13.9%, P<0.01) but not for those undergoing CABG (15.6% versus 14.2%, P=0.57). The adjusted relative mortality risk for PTCA in the randomized versus registry population was 1.17 (P=0.16). CONCLUSIONS: BARI physicians were able to select PTCA rather than CABG for 65% of registry patients who underwent revascularization without compromising long-term survival either in the overall population or in treated diabetics. PMID- 10859285 TI - Impairment of ventilatory efficiency in heart failure: prognostic impact. AB - BACKGROUND: Impairment of ventilatory efficiency in congestive heart failure (CHF) correlates well with symptomatology and contributes importantly to dyspnea. METHODS AND RESULTS: We investigated 142 CHF patients (mean NYHA class, 2.6; mean maximum oxygen consumption [VO(2)max], 15.3 mL O(2) x kg(-1) x min(-1); mean left ventricular ejection fraction [LVEF], 27%). Patients were compared with 101 healthy control subjects. Cardiopulmonary exercise testing was performed, and ventilatory efficiency was defined as the slope of the linear relationship of V(CO(2)) and ventilation (VE). Results are presented in percent of age- and sex adjusted mean values. Forty-four events (37 deaths and 7 instances of heart transplantation, cardiomyoplasty, or left ventricular assist device implantation) occurred. Among VO(2)max, NYHA class, LVEF, total lung capacity, and age, the most powerful predictor of event-free survival was the VE versus V(CO(2)) slope; patients with a slope 130% (54.7%; P<0.001). CONCLUSIONS: The VE versus V(CO(2)) slope is an excellent prognostic parameter. It is easier to obtain than parameters of maximal exercise capacity and is of higher prognostic importance than VO(2)max. PMID- 10859286 TI - QTL influencing blood pressure maps to the region of PPH1 on chromosome 2q31-34 in Old Order Amish. AB - BACKGROUND: Hypertension is a major risk factor for coronary heart disease, stroke, congestive heart failure, renal insufficiency, and peripheral vascular disease. Although the genetic contribution to variation in blood pressure is well recognized, the specific genes involved are mostly unknown. We carried out a genome-wide scan to identify loci influencing blood pressure in the Old Order Amish population of Lancaster County, Pennsylvania. METHODS AND RESULTS: Blood pressures were measured in 694 adult participants from families recruited without regard to blood pressure. We performed a quantitative linkage analysis by using 357 microsatellite markers. In multipoint analysis, strong evidence for linkage was observed with both diastolic (lod=3.36; P=0.00004) and to a lesser extent systolic (lod=1.64; P=0.003) blood pressure in the region of chromosome 2q31-34. Peak evidence for linkage occurred at map positions 217 and 221 cM from pter for diastolic and systolic blood pressure, respectively. CONCLUSIONS: A gene linked to familial primary pulmonary hypertension has recently been mapped to this same region, suggesting the intriguing hypothesis that other (attenuated) mutations in this same gene may influence variation in systolic and diastolic blood pressure in this population. PMID- 10859287 TI - Short-term clinical outcome of patients with acute pulmonary embolism, normal blood pressure, and echocardiographic right ventricular dysfunction. AB - BACKGROUND: The role of echocardiographic right ventricular (RV) dysfunction in predicting clinical outcome in clinically stable patients with pulmonary embolism (PE) is undefined. In this study, we assessed the prevalence and clinical outcome of normotensive patients with RV dysfunction among a broad spectrum of PE patients. METHODS AND RESULTS: This prospective clinical outcome study included cohort of 209 consecutive patients (age, 65+/-15 years) with documented PE. Acute RV dysfunction was diagnosed in the presence of >/=1 of the following: RV dilatation (without hypertrophy), paradox septal systolic motion, and Doppler evidence of pulmonary hypertension. Four groups were identified: 28 patients presenting with shock or cardiac arrest (13%), 19 hypotensive patients without shock (9%), 65 normotensive patients with echocardiographic RV dysfunction (31%), and 97 normotensive patients without RV dysfunction (47%). Among normotensive patients with RV dysfunction, 6 (10%) developed PE-related shock after admission: 3 of these patients died, and 3 were successfully treated with thrombolytic agents. In comparison, none of the 97 normotensive patients without RV dysfunction developed shock or died as a result of PE. CONCLUSIONS: A significant proportion (31%) of normotensive patients with acute PE presents with RV dysfunction; these patients with latent hemodynamic impairment have a 10% rate of PE-related shock and 5% in-hospital mortality and may require aggressive therapeutic strategies. Conversely, normotensive patients without echocardiographic RV dysfunction have a benign short-term prognosis. Thus, early detection of echocardiographic RV dysfunction is of major importance in the risk stratification of normotensive patients with acute PE. PMID- 10859288 TI - Early potent antithrombotic effect with combined aspirin and a loading dose of clopidogrel on experimental arterial thrombogenesis in humans. AB - BACKGROUND: We conducted a double-blind, randomized, crossover study to assess the antithrombotic effects of the combination of aspirin (acetylsalicylic acid, ASA) and clopidogrel, with or without a loading dose, versus ASA alone in a model of arterial thrombosis in humans. METHODS AND RESULTS: Eighteen male volunteers received the following 3 regimens for 10 days separated by a 1-month period: (1) 325 mg ASA daily, (2) 325 mg ASA+75 mg clopidogrel daily, (3) 325 mg ASA daily+300-mg clopidogrel loading dose on day 1 and +75 mg clopidogrel per day on days 2 to 10. The antithrombotic effect was measured 1.5, 6, and 24 hours after drug intake on day 1 and 6 hours after drug intake on day 10. Arterial thrombus formation was induced ex vivo by exposing a collagen-coated coverslip in a parallel-plate perfusion chamber to native blood for 3 minutes at an arterial wall shear rate. Without a loading dose, clopidogrel+ASA developed an antithrombotic effect within 6 hours after the first intake. It was superior to that produced by ASA, but it was moderate (P90%) exhibit mild hyperhomocysteinemia refractory to treatment with either regimen. This treatment refractoriness is not related to defects in folate absorption or circulating plasma and tissue distribution. PMID- 10859290 TI - Cardiovascular responses to the isoprostanes iPF(2alpha)-III and iPE(2)-III are mediated via the thromboxane A(2) receptor in vivo. AB - BACKGROUND: Isoprostanes (iPs) are free radical-catalyzed products of arachidonic acid that reflect lipid peroxidation in vivo. Several iPs exert biological effects in vitro and may contribute to the functional consequences of oxidant stress. For example, iPF(2alpha)-III (8-iso PGF(2alpha)) and iPE(2)-III modulate platelet function and vascular tone. Although these effects are blocked by antagonists of the receptor (TP) for the cyclooxygenase product thromboxane A(2), it has been speculated that the iPs may activate a receptor related to, but distinct from, the TP. METHODS AND RESULTS: Transgenic mice (TPOEs) were generated in which the TP-beta isoform was under the control of the preproendothelin promoter. They overexpressed TP-beta in the vasculature but not in platelets and exhibited an exaggerated pressor response to infused iPF(2alpha) III compared with wild-type mice. This was blocked by TP antagonism. The platelet response to the iP was unaltered in TPOEs compared with wild-type mice. By contrast, both the pressor response to iPF(2alpha)-III and its effects on platelet function were abolished in mice lacking the TP gene. This was also true of the effects of infused iPE(2)-III on mean arterial pressure and platelet aggregation. CONCLUSIONS: Both iPF(2alpha)-III and iPE(2)-III exert their effects on platelet function and vascular tone in vivo by acting as incidental ligands at membrane TPs rather than via a distinct iP receptor. Activation of TPs by iPs may be of importance in syndromes in which cyclooxygenase activation and oxidant stress coincide, such as in atherosclerosis and reperfusion after tissue ischemia. PMID- 10859291 TI - Adenosine A(1) receptor activation induces delayed preconditioning in rats mediated by manganese superoxide dismutase. AB - BACKGROUND: We have previously described a second window of protection against infarction in rabbits 24 to 72 hours after adenosine A(1) receptor (A(1)R) activation. In this study, we examined the potential role of the mitochondrial antioxidant manganese superoxide dismutase (Mn-SOD) as a potential end effector in mediating this protection. METHODS AND RESULTS: Rats were treated with an intravenous bolus of the A(1)R agonist 2-chloro-N(6)-cyclopentyladenosine (CCPA, 75 microg/kg) or saline vehicle. They were also given a 5 mg/kg IV infusion of a 22-mer phosphorothioate oligodeoxynucleotide (ODN) with sequence antisense to the initiation site of rat Mn-SOD mRNA. Sense ODN and scrambled ODN were used as controls. Twenty-four hours later, hearts were isolated and perfused with buffer at constant pressure and subjected to 35 minutes of regional ischemia and 2 hours of reperfusion. Treatment with CCPA compared with saline vehicle (control) significantly reduced infarct size, expressed as percentage of myocardium at risk (22.3+/-3.3% versus 42.1+/-3.8%, respectively; P=0.001). This protection was completely abolished by prior treatment with antisense ODN, which had no effect on its own. Neither sense ODN nor scrambled ODN had an effect on the CCPA-induced delayed cardioprotection. In separate animals, 24 hours after the same treatment, hearts were assayed for Mn-SOD content and activity. CCPA treatment induced a significant increase in myocardial Mn-SOD content and activity compared with the control condition; this increase was abolished by pretreatment with antisense ODN. CONCLUSIONS: This is the first study to show that transient A(1)R activation induces delayed cardioprotection in the rat. These results strongly suggest an important role for mitochondrial Mn-SOD as a potential end effector of this protection. PMID- 10859292 TI - Long-term endothelin receptor antagonist administration improves alterations in expression of various cardiac genes in failing myocardium of rats with heart failure. AB - BACKGROUND: We reported that long-term (3-month) treatment with the endothelin (ET) type A (ET(A)) receptor antagonist BQ-123 markedly improved survival in rats with chronic heart failure (CHF). However, it is not known whether long-term treatment with an ET receptor antagonist improves alterations in the expression of cardiac genes in failing hearts. METHODS AND RESULTS: CHF rats and control sham-operated rats were treated with BQ-123, SB209670 (ET(A/B) dual receptor antagonist), or saline (vehicle) for 3 months. The survival of CHF rats was markedly higher in the BQ-123 or SB209670 treatment group than in the saline treatment group. The changes in the gene expression of classic molecular markers for failing hearts (mRNA levels of atrial natriuretic peptide and beta-myosin heavy chain) were greatly inhibited by BQ-123 or SB209670 treatment in CHF rats. Long-term BQ-123 treatment also normalized the alterations in the expression of functional molecular markers in failing hearts (eg, mRNA levels of ryanodine receptor, sarcoplasmic reticulum Ca(2+)-ATPase, angiotensin-converting enzyme, angiotensin II type 1 receptor, and prepro-ET-1). CONCLUSIONS: We demonstrated for the first time that long-term (3-month) treatment with an ET receptor antagonist improves the alterations in the expression of various cardiac genes of classic molecular markers (eg, mRNA in atrial natriuretic peptide and beta-myosin heavy chain) and of functional molecular markers (eg, mRNA levels of ryanodine receptor, sarcoplasmic reticulum Ca(2+)-ATPase, angiotensin-converting enzyme, angiotensin II type 1 receptor, and prepro-ET-1) in the failing hearts of CHF rats, suggesting that the great improvement of survival in CHF rats by an ET blocker is partly attributed to the prevention of molecular changes in failing hearts. PMID- 10859293 TI - Left ventricular hypertrophy in ascending aortic stenosis mice: anoikis and the progression to early failure. AB - BACKGROUND: To determine potential mechanisms of the transition from hypertrophy to very early failure, we examined apoptosis in a model of ascending aortic stenosis (AS) in male FVB/n mice. METHODS AND RESULTS: Compared with age-matched controls, 4-week and 7-week AS animals (n=12 to 16 per group) had increased ratios of left ventricular weight to body weight (4.7+/-0.7 versus 3.1+/-0.2 and 5. 7+/-0.4 versus 2.7+/-0.1 mg/g, respectively, P<0.05) with similar body weights. Myocyte width was also increased in 4-week and 7-week AS mice compared with controls (19.0+/-0.8 and 25.2+/-1.8 versus 14. 1+/-0.5 microm, respectively, P<0.01). By 7 weeks, AS myocytes displayed branching with distinct differences in intercalated disk size and staining for beta(1)-integrin on both cell surface and adjacent extracellular matrix. In vivo left ventricular systolic developed pressure per gram as well as endocardial fractional shortening were similar in 4 week AS and controls but depressed in 7-week AS mice. Myocyte apoptosis estimated by in situ nick end-labeling (TUNEL) was extremely rare in 4-week AS and control mice; however, a low prevalence of TUNEL-positive myocytes and DNA laddering were detected in 7-week AS mice. The specificity of TUNEL labeling was confirmed by in situ ligation of hairpin oligonucleotides. CONCLUSIONS: Our findings indicate that myocyte apoptosis develops during the transition from hypertrophy to early failure in mice with chronic biomechanical stress and support the hypothesis that the disruption of normal myocyte anchorage to adjacent extracellular matrix and cells, a process called anoikis, may signal apoptosis. PMID- 10859294 TI - Cardiac hypertrophy is not a required compensatory response to short-term pressure overload. AB - BACKGROUND: Cardiac hypertrophy is considered a necessary compensatory response to sustained elevations of left ventricular (LV) wall stress. METHODS AND RESULTS: To test this, we inhibited calcineurin with cyclosporine (CsA) in the setting of surgically induced pressure overload in mice and examined in vivo parameters of ventricular volume and function using echocardiography. Normalized heart mass increased 45% by 5 weeks after thoracic aortic banding (TAB; heart weight/body weight, 8.3+/-0.9 mg/g [mean+/-SEM] versus 5. 7+/-0.1 mg/g unbanded, P<0.05). Similar increases were documented in the cell-surface area of isolated LV myocytes. In mice subjected to TAB+CsA treatment, we observed complete inhibition of hypertrophy (heart weight/body weight, 5.2+/-0.3 mg/g at 5 weeks) and myocyte surface area (endocardial and epicardial fractions). The mice tolerated abolition of hypertrophy with no signs of cardiovascular compromise, and 5-week mortality was not different from that of banded mice injected with vehicle (TAB+Veh). Despite abolition of hypertrophy by CsA (LV mass by echo, 83+/ 5 mg versus 83+/-2 mg unbanded), chamber size (end-diastolic volume, 33+/-6 microL versus 37+/-1 microL unbanded), and systolic ejection performance (ejection fraction, 97+/-2% versus 97+/-1% unbanded) were normal. LV mass differed significantly in TAB+Veh animals (103+/-5 mg, P<0.05), but chamber volume (end-diastolic volume, 44+/-6 microL), ejection fraction (92+/-2%), and transstenotic pressure gradients (70+/-14 mm Hg in TAB+Veh versus 77+/-11 mm Hg in TAB+CsA) were not different. CONCLUSIONS: In this experimental setting, calcineurin blockade with CsA prevented LV hypertrophy due to pressure overload. TAB mice treated with CsA maintain normal LV size and systolic function. PMID- 10859295 TI - Images in cardiovascular medicine. "Switched" precordial leads. PMID- 10859296 TI - MRI of Uhl's anomaly. PMID- 10859297 TI - First US Implantation of DeBakey Ventricular Assist Device. PMID- 10859298 TI - Evidence for ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50) self association through PDZ-PDZ interactions. AB - Ezrin-radixin-moesin (ERM)-binding phosphoprotein 50 (EBP50) is a versatile membrane-cytoskeleton linking protein that binds to the COOH-tail of specific integral membrane proteins through its two PDZ domains. These EBP50 binding interactions have been implicated in sequestering interactive sets of proteins into common microdomains, regulating the activity of interacting proteins, and modulating membrane protein trafficking. With only two PDZ domains, it is unclear how EBP50 forms multiprotein complexes. Other PDZ proteins increase their breadth and diversity of protein interactions through oligomerization. Hypothesizing that EBP50 self-associates to amplify its functional capacity, far-Western blotting of cholangiocyte epithelial cell proteins with EBP50 fusion protein revealed that EBP50 binds to a 50-kDa protein. Far-Western blotting of EBP50 isolated by two dimensional gel electrophoresis or immunoprecipitation demonstrates that the 50 kDa binding partner is itself EBP50. Further, co-transfection/co-precipitation studies show the self-association can occur in an intracellular environment. In vitro analysis of the EBP50-EBP50 binding interaction indicates it is both saturable and of relatively high affinity. Analysis of truncated EBP50 proteins indicates EBP50 self-association is mediated through its PDZ domains. The ability to self-associate provides a mechanism for EBP50 to expand its capacity to form multiprotein complexes and regulate membrane transport events. PMID- 10859299 TI - Oncogenic signals of HER-2/neu in regulating the stability of the cyclin dependent kinase inhibitor p27. AB - Overexpression and activation of HER-2/neu, a proto-oncogene, play a pivotal role in cancer formation. Strong expression of HER-2/neu in cancers has been associated with poor prognosis. Reduced expression of p27(Kip1), a cyclin dependent kinase inhibitor, correlates with poor clinical outcome in many types of carcinomas. Because many cancers with the overexpression of HER-2/neu overlap with those affected by reduced p27 expression, we studied the link between HER 2/neu oncogenic signals and p27 regulation. We found that down-regulation of p27 correlates with HER-2/neu overexpression. To address the molecular mechanism of this inverse correlation, we found that reduction of p27 is caused by enhanced ubiquitin-mediated degradation, and the HER-2/Grb2/MAPK pathway is involved in the decrease of p27 stability. Also, HER-2/neu activity causes mislocation of p27 and Jun activation domain-binding protein 1 (JAB1), an exporter of p27, into the cytoplasm, thereby facilitating p27 degradation. These results reveal that HER 2/neu signals reduce p27 stability and thus present potential points for therapeutic intervention in HER-2/neu-associated cancers. PMID- 10859300 TI - Identification of two pairs of spatially approximated residues within the carboxyl terminus of secretin and its receptor. AB - The carboxyl-terminal domains of secretin family peptides have been shown to contain key determinants for high affinity binding to their receptors. In this work, we have examined the interaction between carboxyl-terminal residues within secretin and the prototypic secretin receptor. We previously utilized photoaffinity labeling to demonstrate spatial approximation between secretin residue 22 and the receptor domain that includes the first 30 residues of the amino terminus (Dong, M., Wang, Y., Pinon, D. I., Hadac, E. M., and Miller, L. J. (1999) J. Biol. Chem. 274, 903-909). Here, we further refined the site of labeling with the p-benzoyl-phenylalanine (Bpa(22)) probe to receptor residue Leu(17) using progressive cleavage of wild type and mutant secretin receptors (V13M and V16M) and sequence analysis. We also developed a new probe incorporating a photolabile Bpa at position 26 of secretin, closer to its carboxyl terminus. This analogue was also a potent agonist (EC(50) = 72 +/- 6 pm) and bound to the secretin receptor specifically and with high affinity (K(i) = 10.3 +/- 2.4 nm). It covalently labeled the secretin receptor at a single site saturably and specifically. This was localized to the segment between residues Gly(34) and Ala(41) using chemical and enzymatic cleavage of labeled wild type and A41M mutant receptor constructs and immunoprecipitation of epitope-tagged receptor fragments. Radiochemical sequencing identified the site of covalent attachment as residue Leu(36). These new insights, along with our recent report of contact between residue 6 within the amino-terminal half of secretin and this same amino-terminal region of this receptor (Dong, M., Wang, Y., Hadac, E. M., Pinon, D. I., Holicky, E. L., and Miller, L. J. (1999) J. Biol. Chem. 274, 19161 19167), support a key role for this region, making the molecular details of this interaction of major interest. PMID- 10859301 TI - Amino acid identity and/or position determines the proteasomal cleavage of the HLA-A*0201-restricted peptide tumor antigen MAGE-3271-279. AB - The proteasome plays a crucial role in the proteolytic processing of antigens presented to T cells in the context of major histocompatibility complex class I molecules. However, the rules governing the specificity of cleavage sites are still largely unknown. We have previously shown that a cytolytic T lymphocyte defined antigenic peptide derived from the MAGE-3 tumor-associated antigen (MAGE 3(271-279), FLWGPRALV in one-letter code) is not presented at the surface of melanoma cell lines expressing the MAGE-3 protein. By using purified proteasome and MAGE-3(271-279) peptides extended at the C terminus by 6 amino acids, we identified predominant cleavages after residues 278 and 280 but no detectable cleavage after residue Val(279), the C terminus of the antigenic peptide. In the present study, we have investigated the influence of Pro(275), Leu(278), and Glu(280) on the proteasomal digestion of MAGE-3(271-285) substituted at these positions. We show that positions 278 and 280 are major proteasomal cleavage sites because they tolerate most amino acid substitutions. In contrast, the peptide bond after Val(279) is a minor cleavage site, influenced by both distal and proximal amino acid residues. PMID- 10859302 TI - GAKIN, a novel kinesin-like protein associates with the human homologue of the Drosophila discs large tumor suppressor in T lymphocytes. AB - Reorganization of the cortical cytoskeleton is a hallmark of T lymphocyte activation. Upon binding to antigen presenting cells, the T cells rapidly undergo cytoskeletal re-organization thus forming a cap at the cell-cell contact site leading to receptor clustering, protein segregation, and cellular polarization. Previously, we reported cloning of the human lymphocyte homologue of the Drosophila Discs Large tumor suppressor protein (hDlg). Here we show that a novel protein termed GAKIN binds to the guanylate kinase-like domain of hDlg. Affinity protein purification, peptide sequencing, and cloning of GAKIN cDNA from Jurkat J77 lymphocytes identified GAKIN as a novel member of the kinesin superfamily of motor proteins. GAKIN mRNA is ubiquitously expressed, and the predicted amino acid sequence shares significant sequence similarity with the Drosophila kinesin 73 motor protein. GAKIN sequence contains a motor domain at the NH(2) terminus, a central stalk domain, and a putative microtubule-interacting sequence called the CAP-Gly domain at the COOH terminus. Among the MAGUK superfamily of proteins examined, GAKIN binds to the guanylate kinase-like domain of PSD-95 but not of p55. The hDlg and GAKIN are localized mainly in the cytoplasm of resting T lymphocytes, however, upon CD2 receptor cross-linking the hDlg can translocate to the lymphocyte cap. We propose that the GAKIN-hDlg interaction lays the foundation for a general paradigm of coupling MAGUKs to the microtubule-based cytoskeleton, and that this interaction may be functionally important for the intracellular trafficking of MAGUKs and associated protein complexes in vivo. PMID- 10859303 TI - Involvement of p38 mitogen-activated protein kinase signaling pathway in osteoclastogenesis mediated by receptor activator of NF-kappa B ligand (RANKL). AB - The receptor activator of NF-kappaB ligand (RANKL) induces osteoclast differentiation from bone marrow cells in the presence of macrophage colony stimulating factor. We found that treatment of bone marrow cells with SB203580 inhibited osteoclast differentiation via inhibition of the RANKL-mediated signaling pathway. To elucidate the role of p38 mitogen-activated protein (MAP) kinase pathway in osteoclastogenesis, we employed RAW264 cells which could differentiate into osteoclast-like cells following treatment with RANKL. In a dose-dependent manner, SB203580 but not PD98059, inhibited RANKL-induced differentiation. Among three MAP kinase families tested, this inhibition profile coincided only with the activation of p38 MAP kinase. Expression in RAW264 cells of the dominant negative form of either p38alpha MAP kinase or MAP kinase kinase (MKK) 6 significantly inhibited RANKL-induced differentiation of the cells. These results indicate that activation of the p38 MAP kinase pathway plays an important role in RANKL-induced osteoclast differentiation of precursor bone marrow cells. PMID- 10859304 TI - Radiolytic studies of trimethylamine dehydrogenase. Spectral deconvolution of the neutral and anionic flavin semiquinone, and determination of rate constants for electron transfer in the one-electron reduced enzyme. AB - Trimethylamine dehydrogenase from the pseudomonad Methylophilus methylotrophus has been examined using the technique of pulse radiolysis to rapidly introduce a single reducing equivalent into the enzyme. Using enzyme that has had its iron sulfur center rendered redox-inert by prior reaction with ferricenium hexafluorophosphate, we determined the spectral change associated with formation of both the anionic and neutral forms that were generated at high and low pH, respectively, of the unique 6-cysteinyl-FMN of the enzyme. With native enzyme, electron transfer was observed within the radiolytically generated one-electron reduced enzyme but only at low pH (6.0). The kinetics and thermodynamics of this electron transfer in one-electron reduced enzyme may be compared with that studied previously in the two-electron reduced enzyme. In contrast to previous studies with two-electron reduced enzyme in which a pK(a) of approximately 8 was determined for the flavin semiquinone, in the one-electron reduced enzyme the semiquinone was not substantially protonated even at pH 6. 0. These results indicate that reduction of the iron-sulfur center of the enzyme significantly decreases the pK(a) of the flavin semiquinone of the active site. This provides further evidence, in conjunction with the strong magnetic interaction known to exist between the centers in the two-electron reduced enzyme, that the two redox active centers in trimethylamine dehydrogenase are in intimate contact with one another in the active site of the enzyme. PMID- 10859305 TI - Functional antagonism between Msx2 and CCAAT/enhancer-binding protein alpha in regulating the mouse amelogenin gene expression is mediated by protein-protein interaction. AB - Ameloblast-specific amelogenin gene expression is spatiotemporally regulated during tooth development. In a previous study, the CCAAT/enhancer-binding protein alpha (C/EBPalpha) was identified as a transcriptional activator of the mouse amelogenin gene in a cell type-specific manner. Here, Msx2 is shown to repress the promoter activity of amelogenin-promoter reporter constructs independent of its intrinsic DNA binding activity. In transient cotransfection assays, Msx2 and C/EBPalpha antagonize each other in regulating the expression of the mouse amelogenin gene. Electrophoresis mobility shift assays demonstrate that Msx2 interferes with the binding of C/EBPalpha to its cognate site in the mouse amelogenin minimal promoter, although Msx2 itself does not bind to the same promoter fragment. Protein-protein interaction between Msx2 and C/EBPalpha is identified with co-immunoprecipitation analyses. Functional antagonism between Msx2 and C/EBPalpha is also observed on the stably transfected 2.2-kilobase mouse amelogenin promoter in ameloblast-like LS8 cells. Furthermore, the carboxyl terminal residues 183-267 of Msx2 are required for protein-protein interaction, whereas the amino-terminal residues 2-97 of Msx2 play a less critical role. Among three family members tested (C/EBPalpha, -beta, and -gamma), Msx2 preferentially interacts with C/EBPalpha. Taken together, these data indicate that protein protein interaction rather than competition for overlapping binding sites results in the functional antagonism between Msx2 and C/EBPalpha in regulating the mouse amelogenin gene expression. PMID- 10859306 TI - A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast. AB - The mammalian inducer of apoptosis Bax is lethal when expressed in yeast and plant cells. To identify potential inhibitors of Bax in plants we transformed yeast cells expressing Bax with a tomato cDNA library and we selected for cells surviving after the induction of Bax. This genetic screen allows for the identification of plant genes, which inhibit either directly or indirectly the lethal phenotype of Bax. Using this method a number of cDNA clones were isolated, the more potent of which encodes a protein homologous to the class theta glutathione S-transferases. This Bax-inhibiting (BI) protein was expressed in Escherichia coli and found to possess glutathione S-transferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glutathione, causes a substantial reduction of total cellular phospholipids, diminishes the mitochondrial membrane potential, and alters the intracellular redox potential. Co-expression of the BI-GST/GPX protein brought the total glutathione levels back to normal and re-established the mitochondrial membrane potential but had no effect on the phospholipid alterations. Moreover, expression of BI-GST/GPX in yeast was found to significantly enhance resistance to H(2)O(2)-induced stress. These results underline the relationship between oxidative stress and Bax-induced death in yeast cells and demonstrate that the yeast-based genetic strategy described here is a powerful tool for the isolation of novel antioxidant and antiapoptotic genes. PMID- 10859307 TI - sn-1,2-diacylglycerol levels in the fungus Neurospora crassa display circadian rhythmicity. AB - The fungus Neurospora crassa is a model organism for investigating the biochemical mechanism of circadian (daily) rhythmicity. When a choline-requiring strain (chol-1) is depleted of choline, the period of the conidiation rhythm lengthens. We have found that the levels of sn-1,2-diacylglycerol (DAG) increase in proportion to the increase in period. Other clock mutations that change the period do not affect the levels of DAG. Membrane-permeant DAGs and inhibitors of DAG kinase were found to further lengthen the period of choline-depleted cultures. The level of DAG oscillates with a period comparable to the rhythm of conidiation in wild-type strains, choline-depleted cultures, and frq mutants, including a null frq strain. The DAG rhythm is present at the growing margin and also persists in older areas that have completed development. The phase of the DAG rhythm can be set by the light-to-dark transition, but the level of DAG is not immediately affected by light. Our results indicate that rhythms in DAG levels in Neurospora are driven by a light-sensitive circadian oscillator that does not require the frq gene product. High levels of DAG may feed back on that oscillator to lengthen its period. PMID- 10859308 TI - Identification and characterization of a 315-base pair enhancer, located more than 55 kilobases 5' of the apolipoprotein B gene, that confers expression in the intestine. AB - We recently reported that an 8-kilobase (kb) region, spanning from -54 to -62 kb 5' of the human apolipoprotein B (apoB) gene, contains intestine-specific regulatory elements that control apoB expression in the intestines of transgenic mice. In this study, we further localized the apoB intestinal control region to a 3-kb segment (-54 to -57 kb). DNaseI hypersensitivity studies uncovered a prominent DNaseI hypersensitivity site, located within a 315-base pair (bp) fragment at the 5'-end of the 3-kb segment, in transcriptionally active CaCo-2 cells but not in transcriptionally inactive HeLa cells. Transient transfection experiments with CaCo-2 and HepG2 cells indicated that the 315-bp fragment contained an intestine-specific enhancer, and analysis of the DNA sequence revealed putative binding sites for the tissue-specific transcription factors hepatocyte nuclear factor 3beta, hepatocyte nuclear factor 4, and CAAT enhancer binding protein beta. Binding of these factors to the 315-bp enhancer was demonstrated in gel retardation experiments. Transfection of deletion mutants of the 315-bp enhancer revealed the relative contributions of these transcription factors in the activity of the apoB intestinal enhancer. The corresponding segment of the mouse apoB gene (located -40 to -83 kb 5' of the structural gene) exhibited a high degree of sequence conservation in the binding sites for the key transcriptional activators and also exhibited enhancer activity in transient transfection assays with CaCo-2 cells. In transgenic mouse expression studies, the 315-bp enhancer conferred intestinal expression to human apoB transgenes. PMID- 10859309 TI - Nucleotide-dependent binding of the GTPase domain of the signal recognition particle receptor beta-subunit to the alpha-subunit. AB - The signal recognition particle (SRP) receptor (SR) is a heterodimer of two polypeptides (SRalpha and SRbeta) that each contain a GTP-binding domain. The GTP binding domain in the peripheral membrane SRalpha subunit has a well defined role in regulating targeting of SRP-ribosome-nascent chain complexes to the translocon. The only well established function for the transmembrane SRbeta subunit is anchoring SRalpha on the endoplasmic reticulum membrane. Deletion of the amino-terminal transmembrane domain of SRbeta did not affect receptor dimerization, but revealed a cryptic translocation signal that overlaps the GTPase domain. We demonstrate that the domain of SRalpha that binds SRbeta does so by binding directly to the nucleotide-bound form of the GTPase domain of SRbeta. An SRbeta mutant containing an amino acid substitution that allows the GTPase domain to bind XTP dimerized with SRalpha most efficiently in the presence of XTP or XDP, but not ATP. Our results suggest an additional level of regulation of SRP receptor function based on regulated dissociation of the receptor subunits. PMID- 10859310 TI - Role of the lipase-specific foldase of Burkholderia glumae as a steric chaperone. AB - Most lipases of Gram-negative bacteria require a lipase-specific foldase (Lif) in order to fold in the periplasm into their active, protease-resistant conformation prior to their secretion. The periplasmic domain of the Lif (amino acids 44-353) of Burkholderia glumae was purified as a His-tagged protein, and its function in the folding of lipase was studied in vitro. Refolding of the denatured lipase into its active conformation was dependent on the presence of the Lif. Circular dichroism revealed that the lipase refolded in the absence of Lif into a form with a native-like conformation, which was more stable against heat-induced denaturation than the native form, but was enzymatically inactive. This form of the protein could be activated by adding Lif after several hours, which demonstrates that the function of this chaperone is to help lipase to overcome an energetic barrier in the productive folding pathway rather than to prevent it from entering a non-productive pathway. The Lif was shown to interact with the native lipase in protease-protection experiments as well as by affinity chromatography, consistent with a role of the Lif late in the folding process. These results demonstrate that the Lif functions in a way analogous to the propeptides of many bacterial proteases and indicate that the amino acid sequence of the lipase does not contain all the information required for the protein to adopt its three-dimensional structure. PMID- 10859311 TI - The functional unit of the renal type IIa Na+/Pi cotransporter is a monomer. AB - The composition of the functional unit of the rat renal type IIa Na(+)/P(i) cotransporter (NaPi-IIa) was investigated by using two approaches based on the differential sensitivities of the wild type (WT) and mutant S460C proteins to 2 aminoethylmethanethiosulfonate hydrobromide (MTSEA), a charged cysteine modifier. Transport activity of S460C is completely blocked after incubation in MTSEA, whereas that of the WT remains unaffected. First, Xenopus laevis oocytes were coinjected with cRNAs coding for the WT and S460C in different proportions, and the transport inhibition after MTSEA incubation was assayed by electrophysiology. The relationship between MTSEA inhibition and proportion of cRNA was consistent with that for a functional monomer. Second, concatameric proteins were constructed that either comprised two WT proteins (WT-WT), two S460C mutants (S460C-S460C), or one of each (WT-S460C). Western blots of oocytes injected with fusion protein cRNA showed bands at approximately 200 kDa, whereas a main band at approximately 90 kDa was obtained for the WT cRNA alone. The kinetic properties of concatamers were the same as for the single proteins. Transport activity of the WT-WT concatamer was unaffected by MTSEA incubation, fully inhibited for S460C-S460C, but 50% inhibited for WT-S460C. This behavior was also consistent with NaPi-IIa being a functional monomer. PMID- 10859312 TI - Protein-tyrosine phosphatase PTPepsilon C inhibits Jak-STAT signaling and differentiation induced by interleukin-6 and leukemia inhibitory factor in M1 leukemia cells. AB - We engineered and expressed both a wild-type and mutant cytosolic isoform of PTPepsilon (PTPepsilonC) in murine M1 leukemic cells, which can be induced to growth arrest and monocytic differentiation by interleukin (IL)-6 and leukemia inhibitory factor (LIF). Forced expression of PTPepsilonC inhibited IL-6- and LIF induced monocytic differentiation and apoptosis in M1 cells, whereas expression of PTPepsilonM, a transmembrane isoform of PTPepsilon, did not. PTPepsilonC expression resulted in lower levels of IL-6-induced tyrosine phosphorylation of Jak1, Tyk2, gp130, and Stat3 compared with parent cells. In M1 transfectants expressing an inactive mutant of PTPepsilonC, both tyrosine phosphorylation and apoptosis induced by IL-6 and LIF were potentiated rather than inhibited. These results suggest an important role for PTPepsilonC in negative regulation of IL-6- and LIF-induced Jak-STAT signaling. PMID- 10859313 TI - Biochemical characterization of Rab3-GTPase-activating protein reveals a mechanism similar to that of Ras-GAP. AB - Small G proteins of the Rab family are regulators of intracellular vesicle traffic. Their intrinsic rate of GTP hydrolysis is very low but is enhanced by specific GTPase-activating proteins (GAPs) that switch G proteins to their inactive form. We have characterized the activity of recombinant Rab3-GAP on Rab3A in solution. The K(m) and K(d) values (75 microm) indicate a low affinity of Rab3-GAP for its substrate. The affinity is higher for the transition state analog Rab3A:GDP:AlF(x) (15 microm). The k(cat) (1 s(-)(1)) is within the range of values reported for other GAPs. A mutation in the switch I region of Rab3A disrupted the interaction with Rab3-GAP. Furthermore, Rabphilin, a putative target of Rab3, inhibited the activity of Rab3-GAP on Rab3. Therefore, the Rab3 GAP-binding site involves the switch I region of Rab3 and overlaps with the Rabphilin-binding domain. Substitution of a single arginine residue (Arg-728) of Rab3-GAP disrupted its catalytic activity but not its interaction with Rab3A. We propose that Rab3-GAP, like Ras- and Rho-GAPs, stabilizes the transition state of Rab3 and provides a critical arginine residue to accelerate the GTPase reaction. PMID- 10859314 TI - Bioengineering of human thyrotropin superactive analogs by site-directed "lysine scanning" mutagenesis. Cooperative effects between peripheral loops. AB - We have previously engineered the first superactive analogs of human thyrotropin (hTSH) by using a novel design strategy. In this study, we have applied homology comparisons focusing on the alphaL3 loop of the common alpha-subunit of human glycoprotein hormones. Seven highly variable amino acid residues were identified, and charge-scanning mutagenesis revealed three previously unrecognized modification permissive domains and four gain-of-function lysine substitutions. Such gain-of-function mutations were hormone- and receptor-specific and dependent on location and basic charge. Cooperativity of individual substitutions was established in double and triple lysine mutants. In combinations of the most potent alphaL3 loop analog with two previously characterized loop analogs, a higher degree of cooperativity for the alphaL3 loop analog compared with both the alphaL1 loop analog and the hTSH-betaL3 loop analog was observed. We demonstrated that spatially distinct regions of the common alpha-subunit contribute differentially to the interaction of hTSH with its receptor and that combinations of two modified loops on the same and on opposite sides of the hTSH molecule display similar increases in in vitro biopotency. In addition, combination of all three superactive loops showed cooperativity in receptor binding and activation resulting in the most potent hTSH superactive analog described to date. PMID- 10859315 TI - Characterization of binding between the chemokine eotaxin and peptides derived from the chemokine receptor CCR3. AB - The CC chemokine eotaxin plays a predominant role in eosinophil trafficking in vivo by specifically activating the chemokine receptor CCR3. We have screened a series of synthetic peptides corresponding to extracellular regions of CCR3 for their ability to bind eotaxin. A peptide corresponding to the N terminus of CCR3 (CCR3-(1-35)) bound to eotaxin with a dissociation constant of 80 +/- 38 micrometer. However, linear or cyclic peptides derived from the first and third extracellular loops of CCR3 did not bind to eotaxin. Linear and cyclic peptides derived from the second extracellular loop precipitated upon addition of eotaxin. (1)H-(15)N correlation NMR spectroscopy indicated that an extended groove in the eotaxin surface, whose edges are defined by the N-loop, 3(10)-helical turn, and beta(2)-beta(3) hairpin, is the most likely binding surface for CCR3-(1-35). NMR assignments for CCR3-(1-35) were obtained using two-dimensional and three dimensional homonuclear NMR experiments. (15)N-Filtered TOCSY spectra indicated that the central region of CCR3-(1-35), surrounding the DDYY sequence, is involved in the interaction with eotaxin. This was supported by the observation that a truncated N-terminal peptide (CCR3-(8-23)) binds to eotaxin with a dissociation constant of 136 +/- 23 micrometer, only slightly weaker than the full-length N-terminal peptide. Taken together with previous studies, these results suggest that interactions between the N-loop/beta(3) regions of chemokines and the N-terminal regions of their receptors may be a conserved feature of chemokine-receptor complexes across the CC, CXC, and C chemokine subfamilies. However, the low affinity of the interactions observed in these studies suggests the existence of additional binding regions in both the chemokines and the receptors. PMID- 10859316 TI - Interactions between the tetratricopeptide repeat-containing transcription factor TFIIIC131 and its ligand, TFIIIB70. Evidence for a conformational change in the complex. AB - In the transcription of tRNA and 5 S genes by RNA polymerase III, recruitment of the transcription factor (TF)IIIB is mediated by the promoter-bound assembly factor TFIIIC. A critical limiting step in this process is the interaction between the tetratricopeptide repeat (TPR)-containing subunit of TFIIIC (TFIIIC131) and the TFIIB-related factor Brf1p/TFIIIB70. To facilitate biochemical studies of this interaction, we expressed a fragment of TFIIIC131, TFIIIC131-(1-580), that includes the minimal TFIIIB70 interaction domain defined by two-hybrid studies together with adjacent sequences, up to the end of TPR9, implicated in the assembly reaction. TFIIIC131-(1-580) interacts with TFIIIB70 in solution and inhibits the formation of TFIIIB70.TFIIIC.DNA complexes. In a coupled equilibrium binding assay, the formation of TFIIIC131-(1-580).TFIIIB70 complexes was adequately described by a single-site binding model and yielded an apparent equilibrium dissociation constant of 334 +/- 23 nm. CD spectroscopy and limited proteolysis experiments defined a well structured and largely protease resistant core in TFIIIC131-(1-580) comprising part of the hydrophilic amino terminus, TPR1-5, the intervening non-TPR region, and TPR6-8. CD spectra showed that trifluoroethanol induced significant alpha-helical structure in TFIIIC131-(1 580). A more modest monovalent ion-dependent CD difference was observed in mixtures of TFIIIC131-(1-580) and TFIIIB70, suggesting that formation of the binary complex may proceed with the acquisition of alpha-helicity. PMID- 10859317 TI - The tissue-dependent keratin 19 gene transcription is regulated by GKLF/KLF4 and Sp1. AB - Keratins play critical roles in cellular differentiation and cytoskeletal organization. Keratin 19 (K19) is unique because it has been implicated as a marker of stem cells in some tissues, such as the hair follicle in the skin. It is also associated with malignant transformation in esophageal and pancreatic cancers. Here, we show that the K19 promoter is active in a subset of gastrointestinal cancer cells derived from esophageal and pancreas but inactive in other contexts. This activity was mapped to a short region containing an overlapping binding site for gut-enriched Kruppel-like factor (GKLF/KLF4) and Sp1. GKLF has a higher binding affinity and is the predominant binding factor in cells with low Sp-1 protein levels. Pancreatic acinar cells normally do not express K19, but overexpression of GKLF and Sp1 in these cells leads to aberrant expression, similar to what is observed in pancreatic cancer. These results demonstrate the functional interaction of ubiquitous and tissue-restricted transcription factors in determining tissue- and neoplasm-specific patterns of gene expression. PMID- 10859318 TI - A selective requirement for elevated calcium in DNA degradation, but not early events in anti-Fas-induced apoptosis. AB - Jurkat cells undergo apoptosis in response to anti-Fas antibody through a caspase dependent death cascade in which calcium signaling has been implicated. We have now evaluated the role of calcium during this death cascade at the single cell level in real time utilizing flow cytometric analysis and confocal microscopy. Fluo-3 and propidium iodide were employed to evaluate calcium fluxes and to discriminate between viable and non-viable cells, respectively. Anti-Fas treatment of Jurkat cells resulted in a sustained increase in intracellular calcium commencing between 1 and 2 h after treatment and persisting until subsequent loss of cell membrane integrity. The significance of this rise in calcium was evaluated by buffering intracellular calcium with BAPTA and/or removing calcium from the extracellular medium and monitoring the effects of these manipulations on calcium signaling and components of the apoptotic process. Complete inhibition of the anti-Fas induced rise in intracellular calcium required both chelation of [Ca(2+)](i) and removal of extracellular calcium. Interestingly, this condition did not abrogate several events in Fas-induced apoptosis including cell shrinkage, mitochondrial depolarization, annexin binding, caspase activation, and nuclear poly(A)DP-ribose polymerase cleavage. Furthermore, calcium-free conditions in the absence of anti-Fas antibody weakly induced these apoptotic components. In marked contrast, calcium depletion did not induce DNA degradation in control cells, and inhibited apoptotic DNA degradation in response to anti-Fas. These data support the concept that the rise in intracellular calcium is not a necessary component for the early signal transduction pathways in anti-Fas-induced apoptosis in Jurkat cells, but rather is necessary for the final degradation of chromatin via nuclease activation. PMID- 10859319 TI - Matrix metalloproteinases cleave tissue factor pathway inhibitor. Effects on coagulation. AB - The capacity of inflammatory cell-derived matrix metalloproteinases (MMPs) to cleave tissue factor pathway inhibitor (TFPI) and alter its activity was investigated. MMP-7 (matrilysin) rapidly cleaved TFPI to a major 35-kDa product. In contrast, MMP-1 (collagenase-1), MMP-9 (gelatinase B), and MMP-12 (macrophage elastase) cleaved TFPI into several fragments including the 35-kDa band. However, rates of cleavage were most rapid for MMP-7 and MMP-9. NH(2)-terminal amino acid sequencing revealed that MMP-12 cleaved TFPI at Lys(20)-Leu(21)(close to Kunitz I domain and producing a 35-kDa band), Arg(83)-Ile(84) (between Kunitz I and II domains), and Ser(174)-Thr(175) (between Kunitz II and III domains). MMP-7 and MMP-9 cleaved TFPI at Lys(20)-Leu(21) with additional COOH-terminal processing. These MMPs did not cleave tissue factor (TF), factor VII, and factor Xa. Proteolytic cleavage by MMP-1, MMP-7, MMP-9, and MMP-12 resulted in considerable loss of TFPI activity. These observations indicate specific cleavage of TFPI by MMPs, which broadens their substrate profile. Co-localization of MMPs, TF, and TFPI in atherosclerotic tissues suggests that release of MMPs from inflammatory cell leukocytes may effect TF-mediated coagulation. PMID- 10859320 TI - Actin-latrunculin A structure and function. Differential modulation of actin binding protein function by latrunculin A. AB - Latrunculin A is used extensively as an agent to sequester monomeric actin in living cells. We hypothesize that additional activities of latrunculin A may be important for its biological activity. Our data are consistent with the formation of a 1:1 stoichiometric complex with an equilibrium dissociation constant of 0.2 to 0.4 micrometer and provide no evidence that the actin-latrunculin A complex participates in the elongation of actin filaments. Profilin and latrunculin A bind independently to actin, whereas binding of thymosin beta(4) to actin is inhibited by latrunculin A. Potential implications of this differential effect on actin-binding proteins are discussed. From a structural perspective, if latrunculin A binds to actin at a site that sterically influences binding by thymosin beta(4), then the observation that latrunculin A inhibits nucleotide exchange on actin implies an allosteric effect on the nucleotide binding cleft. Alternatively, if, as previously postulated, latrunculin A binds in the nucleotide cleft of actin, then its ability to inhibit binding by thymosin beta(4) is a surprising result that suggests that significant allosteric changes affect the thymosin beta(4) binding site. We show that latrunculin A and actin form a crystalline structure with orthorhombic space group P2(1)2(1)2(1) and diffraction to 3.10 A. A high resolution structure with optimized crystallization conditions should provide insight regarding these remarkable allosteric properties. PMID- 10859321 TI - Crystal structure of a thermophilic cytochrome P450 from the archaeon Sulfolobus solfataricus. AB - The structure of the first P450 identified in Archaea, CYP119 from Sulfolobus solfataricus, has been solved in two different crystal forms that differ by the ligand (imidazole or 4-phenylimidazole) coordinated to the heme iron. A comparison of the two structures reveals an unprecedented rearrangement of the active site to adapt to the different size and shape of ligands bound to the heme iron. These changes involve unraveling of the F helix C-terminal segment to extend a loop structure connecting the F and G helices, allowing the longer loop to dip down into the active site and interact with the smaller imidazole ligand. A comparison of CYP119 with P450cam and P450eryF indicates an extensive clustering of aromatic residues may provide the structural basis for the enhanced thermal stability of CYP119. An additional feature of the 4-phenylimidazole-bound structure is a zinc ion tetrahedrally bound by symmetry-related His and Glu residues. PMID- 10859322 TI - Identification that KfiA, a protein essential for the biosynthesis of the Escherichia coli K5 capsular polysaccharide, is an alpha -UDP-GlcNAc glycosyltransferase. The formation of a membrane-associated K5 biosynthetic complex requires KfiA, KfiB, and KfiC. AB - The Escherichia coli K5 capsular polysaccharide consists of the repeat structure 4)GlcA-beta(1,4)-GlcNAc-alpha(1- and requires the KfiA, KfiB, KfiC, and KfiD proteins for its synthesis. Previously, the KfiC protein was shown to be a beta UDP-GlcA glycosyltransferase, and KfiD was shown to be a UDP-Glc dehydrogenase. Here, we demonstrate that KfiA is an alpha-UDP-GlcNAc glycosyltransferase and that biosynthesis of the K5 polysaccharide involves the concerted action of the KfiA and KfiC proteins. By site-directed mutagenesis, we determined that the acidic motif of DDD, which is conserved between the C family of glycosyltransferases, is essential for the enzymatic activity of KfiA. In addition, by Western blot analysis, we determined that association of KfiA with the cytoplasmic membrane requires KfiC but not KfiB, whereas the interaction of KfiC with the cytoplasmic membrane was dependent on both KfiA and KfiB. Likewise, KfiB was only detectable in cytoplasmic membrane fractions when both KfiA and KfiC were present. These data suggest that the interaction between the KfiA, KfiB, and KfiC proteins is essential for the stable association of these proteins with the cytoplasmic membrane and the biosynthesis of the K5 polysaccharide. PMID- 10859323 TI - Steroid-binding specificity of human sex hormone-binding globulin is influenced by occupancy of a zinc-binding site. AB - One calcium-binding site (site I) and a second poorly defined metal-binding site (site II) have been observed previously within the amino-terminal laminin G-like domain (G domain) of human sex hormone-binding globulin (SHBG). By soaking crystals of this structure in 2.5 mm ZnCl(2), site II and a new metal-binding site (site III) were found to bind Zn(2+). Site II is located close to the steroid-binding site, and Zn(2+) is coordinated by the side chains of His(83) and His(136) and the carboxylate group of Asp(65). In this site, Zn(2+) prevents Asp(65) from interacting with the steroid 17beta-hydroxy group and alters the conformations of His(83) and His(136), as well as a disordered region over the steroid-binding site. Site III is formed by the side chains of His(101) and the carboxylate group of Asp(117), and the distance between them (2.7 A) is increased to 3.7 A in the presence of Zn(2+). The affinity of SHBG for estradiol is reduced in the presence of 0. 1-1 mm Zn(2+), whereas its affinity for androgens is unchanged, and chemically-related metal ions (Cd(2+) and Hg(2+)) have similar but less pronounced effects. This is not observed when Zn(2+) coordination at site II is modified by substituting Gln for His(136). An alteration in the steroid binding specificity of human SHBG by Zn(2+) occupancy of site II may be relevant in male reproductive tissues where zinc concentrations are very high. PMID- 10859324 TI - Identification of sites of incorporation in the nicotinic acetylcholine receptor of a photoactivatible general anesthetic. AB - Most general anesthetics including long chain aliphatic alcohols act as noncompetitive antagonists of the nicotinic acetylcholine receptor (nAChR). To locate the sites of interaction of a long chain alcohol with the Torpedo nAChR, we have used the photoactivatible alcohol 3-[(3)H]azioctanol, which inhibits the nAChR and photoincorporates into nAChR subunits. At 1 and 275 microm, 3 [(3)H]azioctanol photoincorporated into nAChR subunits with increased incorporation in the alpha-subunit in the desensitized state. The incorporation into the alpha-subunit was mapped to two large proteolytic fragments. One fragment of approximately 20 kDa (alpha V8-20), containing the M1, M2, and M3 transmembrane segments, showed enhanced incorporation in the presence of agonist whereas the other of approximately 10 kDa (alpha V8-10), containing the M4 transmembrane segment, did not show agonist-induced incorporation of label. Within alpha V8-20, the primary site of incorporation was alpha Glu-262 at the C terminal end of alpha M2, labeled preferentially in the desensitized state. The incorporation at alpha Glu-262 approached saturation between 1 microm, with approximately 6% labeled, and 275 microm, with approximately 30% labeled. Low level incorporation was seen in residues at the agonist binding site and the protein-lipid interface at approximately 1% of the levels in alpha Glu-262. Therefore, the primary binding site of 3-azioctanol is within the ion channel with additional lower affinity interactions within the agonist binding site and at the protein-lipid interface. PMID- 10859325 TI - Cellular gangliosides promote growth factor-induced proliferation of fibroblasts. AB - Cell surface gangliosides have been proposed as modulators of transmembrane signaling. In this study, we used two complementary approaches to investigate the function of cellular gangliosides in the response of mammalian fibroblasts to growth factors. First, inhibition of glucosyl ceramide synthase by a new specific inhibitor of d-l-threo-1-phenyl-2-hexadecanoylamino-3 -pyrrolidino-1-propanol-HC l (glucosylceramide synthase), which depletes cellular gangliosides at a concentration of 1 microm without causing an increase in ceramide levels, blocked epidermal growth factor-stimulated proliferation of fibroblasts. Similarly, responses to several other growth factors that activate receptor tyrosine kinases, including fibroblast growth factor, insulin-like growth factor-I, and platelet-derived growth factor, were inhibited by 50-100%. Conversely, enrichment of cellular gangliosides by preincubation of the mouse and human fibroblasts with exogenously added gangliosides enhanced growth factor-elicited cell proliferation. Novel findings of this study, distinguishing it from previous similar studies, include differential effects of preincubation versus continuous incubation of cells with gangliosides on growth factor-dependent cell proliferation and the growth factor-like action of NeuNAc alpha 2-3Gal beta 1 3GalNAc beta 1-4(NeuNAc alpha 2-3)Gal beta 1-4Glc beta 1-1Cer when cells are pretreated with the ganglioside. PMID- 10859326 TI - Novel structural determinants of mu-conotoxin (GIIIB) block in rat skeletal muscle (mu1) Na+ channels. AB - mu-Conotoxin (mu-CTX) specifically occludes the pore of voltage-dependent Na(+) channels. In the rat skeletal muscle Na(+) channel (mu1), we examined the contribution of charged residues between the P loops and S6 in all four domains to mu-CTX block. Conversion of the negatively charged domain II (DII) residues Asp-762 and Glu-765 to cysteine increased the IC(50) for mu-CTX block by approximately 100-fold (wild-type = 22.3 +/- 7.0 nm; D762C = 2558 +/- 250 nm; E765C = 2020 +/- 379 nm). Restoration or reversal of charge by external modification of the cysteine-substituted channels with methanethiosulfonate reagents (methanethiosulfonate ethylsulfonate (MTSES) and methanethiosulfonate ethylammonium (MTSEA)) did not affect mu-CTX block (D762C: IC(50, MTSEA+) = 2165.1 +/- 250 nm; IC(50, MTSES-) = 2753.5 +/- 456.9 nm; E765C: IC(50, MTSEA+) = 2200.1 +/- 550.3 nm; IC(50, MTSES-) = 3248.1 +/- 2011.9 nm) compared with their unmodified counterparts. In contrast, the charge-conserving mutations D762E (IC(50) = 21.9 +/- 4.3 nm) and E765D (IC(50) = 22.0 +/- 7.0 nm) preserved wild type blocking behavior, whereas the charge reversal mutants D762K (IC(50) = 4139.9 +/- 687.9 nm) and E765K (IC(50) = 4202.7 +/- 1088.0 nm) destabilized mu CTX block even further, suggesting a prominent electrostatic component of the interactions between these DII residues and mu-CTX. Kinetic analysis of mu-CTX block reveals that the changes in toxin sensitivity are largely due to accelerated toxin dissociation (k(off)) rates with little changes in association (k(on)) rates. We conclude that the acidic residues at positions 762 and 765 are key determinants of mu-CTX block, primarily by virtue of their negative charge. The inability of the bulky MTSES or MTSEA side chain to modify mu-CTX sensitivity places steric constraints on the sites of toxin interaction. PMID- 10859327 TI - Complex contribution of the 3'-untranslated region to the expressional regulation of the human inducible nitric-oxide synthase gene. Involvement of the RNA-binding protein HuR. AB - Cytokine stimulation of human DLD-1 cells resulted in a marked expression of nitric-oxide synthase (NOS) II mRNA and protein accompanied by only a moderate increase in transcriptional activity. Also, there was a basal transcription of the NOS II gene, which did not result in measurable NOS II expression. The 3' untranslated region (3'-UTR) of the NOS II mRNA contains four AUUUA motifs and one AUUUUA motif, known to destabilize the mRNAs of proto-oncogenes, nuclear transcription factors, and cytokines. Luciferase reporter gene constructs containing the NOS II 3'-UTR showed a significantly reduced luciferase activity. The embryonic lethal abnormal vision (ELAV)-like protein HuR was found to bind with high affinity to the adenylate/uridylate-rich elements of the NOS II 3'-UTR. Inhibition of HuR with antisense constructs reduced the cytokine-induced NOS II mRNA, whereas overexpression of HuR potentiated the cytokine-induced NOS II expression. This provides evidence that NOS II expression is regulated at the transcriptional and post-transcriptional level. Binding of HuR to the 3'-UTR of the NOS II mRNA seems to play an essential role in the stabilization of this mRNA. PMID- 10859328 TI - Expression of bcl-X(L) restores cell survival, but not proliferation off effector differentiation, in CD28-deficient T lymphocytes. AB - Lymphocytes deficient in the T cell costimulatory molecule CD28 exhibit defects in cell survival, clonal expansion, and differentiation into effector cells. It is known that CD28-mediated signaling results in the upregulation of the Bcl family member Bcl-X(L). To investigate the role that Bcl-X(L) plays in the various functions of CD28, we expressed Bcl-X(L) in CD28-deficient primary T lymphocytes using retrovirus-mediated gene transfer. T cells were activated in vitro and infected with Bcl-X(L) or control retroviruses; this method allows gene expression in activated, cycling cells. Expression of Bcl-X(L) in naive T cells was achieved by reconstitution of the immune system of lethally irradiated recipient mice with retrovirus-infected purified bone marrow stem cells from CD28(-/)- or wild-type donor mice. Our studies demonstrate that Bcl-X(L) prolongs the survival of CD28(-/)- T cells but does not restore normal proliferation or effector cell development. These results indicate that the various functions of CD28 can be dissociated, and provide an experimental approach for testing the roles of downstream signals in the functions of cellular receptors such as CD28. PMID- 10859329 TI - Coupling of peripheral tolerance to endogenous interleukin 10 promotes effective modulation of myelin-activated T cells and ameliorates experimental allergic encephalomyelitis. AB - Several immune-based approaches are being considered for modulation of inflammatory T cells and amelioration of autoimmune diseases. The most recent strategies include simulation of peripheral self-tolerance by injection of adjuvant free antigen, local delivery of cytokines by genetically altered T cells, and interference with the function of costimulatory molecules. Although promising results have been obtained from these studies that define mechanisms of T cell modulation, efficacy, practicality, and toxicity, concerns remain unsolved, thereby justifying further investigations to define alternatives for effective downregulation of aggressive T cells. In prior studies, we demonstrated that an immunoglobulin (Ig) chimera carrying the encephalitogenic proteolipid protein (PLP)1 peptide corresponding to amino acid sequence 139-151 of PLP, Ig PLP1, is presented to T cells approximately 100-fold better than free PLP1. Here, we demonstrate that aggregation endows Ig-PLP1 with an additional feature, namely, induction of interleukin (IL)-10 production by macrophages and dendritic cells, both of which are antigen-presenting cells (APCs). These functions synergize in vivo and drive effective modulation of autoimmunity. Indeed, it is shown that animals with ongoing active experimental allergic encephalomyelitis dramatically reduce the severity of their paralysis when treated with adjuvant free aggregated Ig-PLP1. Moreover, IL-10 displays bystander antagonism on unrelated autoreactive T cells, allowing for reversal of disease involving multiple epitopes. Therefore, aggregated Ig-PLP1 likely brings together a peripheral T cell tolerance mechanism emanating from peptide presentation by APCs expressing suboptimal costimulatory molecules and IL-10 bystander suppression to drive a dual-modal T cell modulation system effective for reversal of autoimmunity involving several epitopes and diverse T cell specificities. PMID- 10859330 TI - Abrogation of experimental colitis correlates with increased apoptosis in mice deficient for CD44 variant exon 7 (CD44v7). AB - Experimental colitis in mice is characterized by infiltration of activated T helper (Th) cells and macrophages into the lamina propria. Particularly, these cells expressed CD44 variant exon 7 (CD44v7)-containing isoforms. Upregulation of CD44v7 isoforms was induced by CD40 ligation, an inflammation-driving interaction between activated Th cells and macrophages. To define the role of CD44v7 in colitis, mice bearing a targeted deletion for exon v7 were generated. In trinitrobenzene sulfonic acid-induced colitis, wild-type mice developed severe signs of persistent inflammation. Mice lacking CD44v7 initially showed unspecific inflammation, then recovered completely. The pathogenic origin was shown to reside in bone marrow-derived CD44v7(+) cells, because adoptive transfer experiments demonstrated an absolute requirement for CD44v7 on hematopoietic cells for maintenance of colitis. Interleukin (IL)-10-deficient mice, which develop a chronic Th1-driven enterocolitis, were crossbred with CD44v6/v7 null mice. In IL-10 x CD44v6/v7 double deficient mice, intestinal inflammation developed only weakly and at an older age. Analysis of cell death in the inflamed lesions revealed that mononuclear cells in the CD44v7 null infiltrates had higher rates of apoptosis than those from wild-type mice. Thus, the region encoded by CD44v7 appears to be essential for survival of effector lymphocytes, resulting in persistence of inflammation. PMID- 10859331 TI - Alteration at a single amino acid residue in the T cell receptor alpha chain complementarity determining region 2 changes the differentiation of naive CD4 T cells in response to antigen from T helper cell type 1 (Th1) to Th2. AB - To study whether changes in the structure of a T cell receptor (TCR) at a single peptide-contacting residue could affect T cell priming with antigenic peptide, we made transgenic mice with a point mutation in the TCR alpha chain of the D10.G4.1 (D10) TCR and bred them to D10 beta chain transgenic mice. The mutation consisted of a leucine to serine substitution at position 51 (L51S), which we had already established contacted the second amino acid of the peptide such that the response to the reference peptide was reduced by approximately 100-fold. A mutation in the reference peptide CA134-146 (CA-WT) from the arginine at peptide position 2 to glycine (R2G) restored full response to this altered TCR. When we examined in vitro priming of naive CD4 T cells, we observed that the response to doses of CA WT that induced T helper cell type 1 (Th1) responses in naive CD4 T cells from mice transgenic for the D10 TCR gave only Th2 responses in naive CD4 T cells derived from the L51S. However, when we primed the same T cells with the R2G peptide, we observed Th1 priming in both D10 and L51S naive CD4 T cells. We conclude from these data that a mutation in the TCR at a key position that contacts major histocompatibility complex-bound peptide is associated with a shift in T cell differentiation from Th1 to Th2. PMID- 10859332 TI - Protein kinase D. A selective target for antigen receptors and a downstream target for protein kinase C in lymphocytes. AB - Protein kinase Cs (PKCs) are activated by antigen receptors in lymphocytes, but little is known about proximal targets for PKCs in antigen receptor-mediated responses. In this report, we define a role for diacylglycerol-regulated PKC isoforms in controlling the activity of the serine/threonine kinase protein kinase D (PKD; also known as PKC mu) in T cells, B cells, and mast cells. Antigen receptor activation of PKD is a rapid and sustained response that can be seen in T cells activated via the T cell antigen receptor, B cells activated via the B cell antigen receptor, and in mast cells triggered via the high-affinity receptor for IgE (FcepsilonR1). Herein, we show that antigen receptor activation of PKD requires the activity of classical/novel PKCs. Moreover, PKC activity is sufficient to bypass the requirement for antigen receptor signals in the induction of PKD activity. These biochemical and genetic studies establish a role for antigen receptor-regulated PKC enzymes in the control of PKD activity. Regulation of PKD activity through upstream PKCs reveals a signaling network that exists between different members of the PKC superfamily of kinases that can operate to amplify and disseminate antigen receptor signals generated at the plasma membrane. PMID- 10859333 TI - An N-acetylated natural ligand of human histocompatibility leukocyte antigen (HLA)-B39. Classical major histocompatibility complex class I proteins bind peptides with a blocked NH(2) terminus in vivo. AB - Sequence-independent interactions involving the free peptidic NH(2) terminus are thought to be an essential feature of peptide binding to classical major histocompatibility complex (MHC) class I proteins. Challenging this paradigm, a natural Nalpha-acetylated ligand of human histocompatibility leukocyte antigen (HLA)-B39 was identified in this study. It matched the NH(2)-terminal sequence of two human helicases, was resistant to aminopeptidase M, and was produced with high yield from a synthetic 30 mer with the sequence of the putative parental protein by the 20S proteasome. This is the first reported natural ligand of classical MHC class I antigens that has a blocked NH(2) terminus. PMID- 10859334 TI - Antiidiotype antibody against platelet anti-GPIIIa contributes to the regulation of thrombocytopenia in HIV-1-ITP patients. AB - Patients with human immunodeficiency virus 1-associated immunological thrombocytopenia (HIV-1-ITP) have markedly elevated platelet-bound immunoglobulin (Ig)G, IgM, and C3C4, as well as serum circulating immune complexes (CICs) composed of the same. Affinity purification of IgGs from their CICs with fixed platelets reveals high-affinity antibody (Ab) against platelet glycoprotein (GP)IIIa 49-66, which correlates inversely with their platelet count. However, sera from these patients have little to no anti-GPIIIa activity. To investigate this, we assayed serum, purified serum IgG, and CIC-Ig from these patients. This revealed approximately 150-fold greater Ab activity in purified serum IgG, and approximately 4,000-fold greater reactivity in CIC-IgG. This was shown to be associated with the presence of antiidiotype Ab2 (both IgG and IgM) sequestered in the CIC-IgG. The IgM antiidiotype was predominantly blocking Ab, as demonstrated by specificity for F(ab')(2) fragments of anti-GPIIIa 49-66 of HIV-1 ITP patients and inhibition of reactivity with peptide GPIIIa 49-66, not with a control peptide. The IgM antiidiotype was not polyreactive. Similar measurements were made in nonthrombocytopenic HIV-1-infected patients. Their serum reactivity was not measurable, but serum Ig and CIC-IgG against platelet GPIIIa 49-66 was present, although considerably lower than that found in HIV-1-ITP patients (26- and 35-fold lower, respectively). In addition, their IgM antiidiotype reactivity was 12-fold greater than that found in HIV-1-ITP patients. The IgM antiidiotype Ab titer of both cohorts correlated with in vivo platelet count (r = 0.7, P = 0. 0001, n = 32). To test the in vivo effectiveness of the IgM antiidiotype, thrombocytopenia was induced in mice with 25 microgram of affinity-purified anti GPIIIa 49-66 (mouse GPIIIa has 83% homology with human GPIIIa and Fc receptors for human IgG1). Maximum effect was obtained at 4-6 h after intraperitoneal injection into Balb/c mice with a platelet count of approximately 30% baseline value. Preincubation of the anti-GPIIIa Ab with control IgM at molar ratios of IgM/IgG of 1:7 before intraperitoneal injection had no effect on the in vivo platelet count, whereas preincubation with patient IgM antiidiotype improved the platelet count to 50-80% of normal. Thrombocytopenia could be reversed after addition of IgM antiidiotype 4 h after induction of thrombocytopenia. Thus, CICs of HIV-1-infected patients contain IgM antiidiotype Ab against anti-GPIIIa, which appears to regulate their serum reactivity in vitro and their level of thrombocytopenia in vivo. PMID- 10859335 TI - The structural basis of repertoire shift in an immune response to phosphocholine. AB - The immune response to phosphocholine (PC)-protein is characterized by a shift in antibody repertoire as the response progresses. This change in expressed gene combinations is accompanied by a shift in fine specificity toward the carrier, resulting in high affinity to PC-protein. The somatically mutated memory hybridoma, M3C65, possesses high affinity for PC-protein and the phenyl-hapten analogue, p-nitrophenyl phosphocholine (NPPC). Affinity measurements using related PC-phenyl analogues, including peptides of varying lengths, demonstrate that carrier determinants contribute to binding affinity and that somatic mutations alter this recognition. The crystal structure of an M3C65-NPPC complex at 2.35-A resolution allows evaluation of the three light chain mutations that confer high-affinity binding to NPPC. Only one of the mutations involves a contact residue, whereas the other two have indirect effects on the shape of the combining site. Comparison of the M3C65 structure to that of T15, an antibody dominating the primary response, provides clear structural evidence for the role of carrier determinants in promoting repertoire shift. These two antibodies express unrelated variable region heavy and light chain genes and represent a classic example of the effect of repertoire shift on maturation of the immune response. PMID- 10859336 TI - Immunization and infection change the number of recombination activating gene (RAG)-expressing B cells in the periphery by altering immature lymphocyte production. AB - Recombination activating gene (RAG) expression in peripheral B cells increases after immunization with (4-hydroxy-3-nitrophenyl) acetyl coupled to chicken gamma globulin (NP-CGG) in alum. This increase could result from reinduction of RAG expression or, alternatively, from accumulation of RAG-expressing immature B cells in the periphery. We have used mice that carry a green fluorescent protein (GFP) RAG indicator transgene (RAG2-GFP) to characterize the RAG-expressing B cells in immunized spleens. Most of the RAG2-GFP-expressing B cells in unimmunized spleen are immature B cells. Injection with NP-CGG in alum initially suppresses lymphopoiesis in the bone marrow and decreases the number of immature RAG2-GFP-expressing B cells in the spleen. Recovery of lymphopoiesis in the bone marrow coincides with accumulation of RAG-expressing immature B cells in the spleen. Most of the RAG-expressing cells that accumulate in the spleen after immunization do not proliferate and they are not germinal center cells. Neither the initial suppression of lymphopoiesis nor the subsequent accumulation of RAG expressing cells in the spleen is antigen dependent, since similar changes are seen with alum alone. Furthermore, such changes in the numbers of developing and circulating immature lymphoid cells are seen after injection with complete Freund's adjuvant or malaria infection. Our experiments suggest that adjuvants and infectious agents cause previously unappreciated alterations in lymphopoiesis resulting in the accumulation of RAG-expressing immature B cells in the spleen. PMID- 10859337 TI - Fibroblasts as host cells in latent leishmaniosis. AB - Intracellular parasites are known to persist lifelong in mammalian hosts after the clinical cure of the disease, but the mechanisms of persistence are poorly understood. Here, we show by confocal laser microscopy that in the draining lymph nodes of mice that had healed a cutaneous infection with Leishmania major, 40% of the persisting parasites were associated with fibroblasts forming the reticular meshwork of the lymph nodes. In vitro, both promastigotes and amastigotes of L. major infected primary skin or lymph node fibroblasts. Compared with macrophages, cytokine-activated fibroblasts had a reduced ability to express type 2 nitric oxide synthase and to kill intracellular L. major. These data identify fibroblasts as an important host cell for Leishmania during the chronic phase of infection and suggest that they might serve as safe targets for the parasites in clinically latent disease. PMID- 10859338 TI - Interferon regulatory factor (IRF)-1 and IRF-2 regulate interferon gamma dependent cyclooxygenase 2 expression. AB - Cyclooxygenases (Cox) are rate-limiting enzymes that initiate the conversion of arachidonic acid to prostanoids. Cox-2 is the inducible isoform that is upregulated by proinflammatory agents, initiating many prostanoid-mediated pathological aspects of inflammation. In this study, we demonstrate that interferon (IFN)-gamma alone or in synergy with lipopolysaccharide (LPS) or interleukin 1alpha induces Cox-2 expression in mouse peritoneal macrophages, which is paralleled by changes in Cox-2 protein levels and prostaglandin E(2) (PGE(2)) release. Induction of Cox-2 was abrogated in macrophages that lack IFN regulatory factor (IRF)-1, consistent with an attenuated hepatic mRNA response in IRF-1(-/-) mice injected with LPS. Conversely, the absence of IRF-2 in macrophages resulted in a significant increase in both basal and inducible Cox-2 gene and protein expression as well as IFN-gamma-stimulated PGE(2) release, identifying IRF-2 as negative regulator of this promoter. Two IFN stimulation response elements were identified in the mouse Cox-2 promoter that were highly conserved in the human Cox-2 gene. Both bind endogenous IRF-1 and IRF-2 and regulate transcription in an IRF-1/2-dependent manner. Our data demonstrate conclusively the importance of IFN-gamma as a direct activator and coactivator of the Cox-2 gene, and the central role of IRF-1/2 family members in this process. PMID- 10859339 TI - Downmodulation of the inflammatory response to bacterial infection by gammadelta T cells cytotoxic for activated macrophages. AB - Although gammadelta T cells are involved in the regulation of inflammation after infection, their precise function is not known. Intraperitoneal infection of T cell receptor (TCR)-delta(-/-) mice with the intracellular bacterium Listeria monocytogenes resulted in the development of necrotic foci in the livers. In contrast, the peritoneal cavities of infected TCR-delta(-/-) mice contained an accumulation of low density activated macrophages and a reduced percentage of macrophages undergoing apoptosis. gammadelta T cell hybridomas derived from mice infected with Listeria were preferentially stimulated by low density macrophages from peritoneal exudates of infected mice. Furthermore, primary splenic gammadelta T cells isolated from Listeria-infected mice were cytotoxic for low density macrophages in vitro, and cytotoxicity was inhibited in the presence of antibodies to the gammadelta TCR. These results demonstrate a novel interaction between gammadelta T cells and activated macrophages in which gammadelta T cells are stimulated by terminally differentiated macrophages to acquire cytotoxic activity and which, in turn, induce macrophage cell death. This interaction suggests that gammadelta T cells regulate the inflammatory response to infection with intracellular pathogens by eliminating activated macrophages at the termination of the response. PMID- 10859340 TI - CD4(+) T cell subsets during virus infection. Protective capacity depends on effector cytokine secretion and on migratory capability. AB - To analyze the antiviral protective capacities of CD4(+) T helper (Th) cell subsets, we used transgenic T cells expressing an I-A(b)-restricted T cell receptor specific for an epitope of vesicular stomatitis virus glycoprotein (VSV G). After polarization into Th1 or Th2 effectors and adoptive transfer into T cell-deficient recipients, protective capacities were assessed after infection with different types of viruses expressing the VSV-G. Both Th1 and Th2 CD4(+) T cells could transfer protection against systemic VSV infection, by stimulating the production of neutralizing immunoglobulin G antibodies. However, only Th1 CD4(+) T cells were able to mediate protection against infection with recombinant vaccinia virus expressing the VSV-G (Vacc-IND-G). Similarly, only Th1 CD4(+) T cells were able to rapidly eradicate Vacc-IND-G from peripheral organs, to mediate delayed-type hypersensitivity responses against VSV-G and to protect against lethal intranasal infection with VSV. Protective capacity correlated with the ability of Th1 CD4(+) T cells to rapidly migrate to peripheral inflammatory sites in vivo and to respond to inflammatory chemokines that were induced after virus infection of peripheral tissues. Therefore, the antiviral protective capacity of a given CD4(+) T cell is governed by the effector cytokines it produces and by its migratory capability. PMID- 10859341 TI - The N-glycans determine the differential blood clearance and hepatic uptake of human immunoglobulin (Ig)A1 and IgA2 isotypes. AB - Human immunoglobulin (Ig)A exists in blood as two isotypes, IgA1 and IgA2, with IgA2 present as three allotypes: IgA2m(1), IgA2m(2), and IgA2m(n). We now demonstrate that recombinant, chimeric IgA1 and IgA2 differ in their pharmacokinetic properties. The major pathway for the clearance of all IgA2 allotypes is the liver. Liver-mediated uptake is through the asialoglycoprotein receptor (ASGR), since clearance can be blocked by injection of excess galactose Ficoll ligand and suppressed in ASGR-deficient mice. In contrast, only a small percentage of IgA1 is cleared through this pathway. The clearance of IgA1 lacking the hinge region with its associated O-linked carbohydrate was more rapid than that of wild-type IgA1. IgA1 and IgA2 that are not rapidly eliminated by the ASGR are both removed through an undefined ASGR-independent pathway with half-lives of 14 and 10 h, respectively. The rapid clearance of IgA2 but not IgA1 through the liver may in part explain why the serum levels of IgA1 are greater than those of IgA2. In addition, dysfunction of the ASGR or altered N-linked glycosylation, but not O-glycans, that affects recognition by this receptor may account for the elevated serum IgA seen in liver disease and IgA nephropathy. PMID- 10859342 TI - Deficiencies of human complement component C4A and C4B and heterozygosity in length variants of RP-C4-CYP21-TNX (RCCX) modules in caucasians. The load of RCCX genetic diversity on major histocompatibility complex-associated disease. AB - The complement component C4 genes located in the major histocompatibility complex (MHC) class III region exhibit an unusually complex pattern of variations in gene number, gene size, and nucleotide polymorphism. Duplication or deletion of a C4 gene always concurs with its neighboring genes serine/threonine nuclear protein kinase RP, steroid 21-hydroxylase (CYP21), and tenascin (TNX), which together form a genetic unit termed the RCCX module. A detailed molecular genetic analysis of C4A and C4B and RCCX modular arrangements was correlated with immunochemical studies of C4A and C4B protein polymorphism in 150 normal Caucasians. The results show that bimodular RCCX has a frequency of 69%, whereas monomodular and trimodular RCCX structures account for 17.0 and 14.0%, respectively. Three quarters of C4 genes harbor the endogenous retrovirus HERV-K(C4). Partial deficiencies of C4A and C4B, primarily due to gene deletions and homoexpression of C4A proteins, have a combined frequency of 31.6%. This is probably the most common variation of gene dosage and gene size in human genomes. The seven RCCX physical variants create a great repertoire of haplotypes and diploid combinations, and a heterozygosity frequency of 69.4%. This phenomenon promotes the exchange of genetic information among RCCX constituents that is important in homogenizing the structural and functional diversities of C4A and C4B proteins. However, such length variants may cause unequal, interchromosomal crossovers leading to MHC-associated diseases. An analyses of the RCCX structures in 22 salt losing, congenital adrenal hyperplasia patients revealed a significant increase in the monomodular structure with a long C4 gene linked to the pseudogene CYP21A, and bimodular structures with two CYP21A, which are likely generated by recombinations between heterozygous RCCX length variants. PMID- 10859343 TI - Mitochondrial basis for immune deficiency. Evidence from purine nucleoside phosphorylase-deficient mice. AB - We generated purine nucleoside phosphorylase (PNP)-deficient mice to gain insight into the mechanism of immune deficiency disease associated with PNP deficiency in humans. Similar to the human disease, PNP deficiency in mice causes an immunodeficiency that affects T lymphocytes more severely than B lymphocytes. PNP knockout mice exhibit impaired thymocyte differentiation, reduced mitogenic and allogeneic responses, and decreased numbers of maturing thymocytes and peripheral T cells. T lymphocytes of PNP-deficient mice exhibit increased apoptosis in vivo and higher sensitivity to gamma irradiation in vitro. We propose that the immune deficiency in PNP deficiency is a result of inhibition of mitochondrial DNA repair due to the accumulation of dGTP in the mitochondria. The end result is increased sensitivity of T cells to spontaneous mitochondrial DNA damage, leading to T cell depletion by apoptosis. PMID- 10859344 TI - Analysis of immunoglobulin (Ig) isotype diversity and IgM/D memory in the response to phenyl-oxazolone. AB - The distribution of immunoglobulin (Ig) isotypes within specific B cell clones in vivo after immunization is not well defined. Using an IgV(H)/CDR3- and isotype specific reverse transcription polymerase chain reaction method, we have carried out a survey of the diversification of the isotype in a splenic response to phenyl-oxazolone (phOx) on a chicken serum albumin carrier. The phOx-specific V(H) (V(H)Ox-1 with specific CDR3 motif) is associated with all of the heavy chains (mu, delta, alpha, gamma, and epsilon) after simple immunization with antigen in alum. The kinetics of expression of each isotype are distinct and reproducible. Focusing mainly on the expression of secretory Ig transcripts, IgM, IgG1, and IgE are found after priming, whereas IgD and IgA appear after boosting. Secretory IgD transcripts are found reproducibly at moderate levels and may, therefore, contribute significantly to the secreted Ig response in mice. Most crucially, we find enhanced levels of secretory IgM/V(H)Ox-1 transcripts (with 'phOx-specific' CDR3) after boosting, strongly indicating the existence of IgM memory cells that give rise to an enhanced specific IgM secretion in the secondary response. PMID- 10859345 TI - CPP1, a DNA-binding protein involved in the expression of a soybean leghemoglobin c3 gene. AB - Nodulin genes are specifically expressed in the nitrogen-fixing root nodules. We have identified a novel type of DNA-binding protein (CPP1) interacting with the promoter of the soybean leghemoglobin gene Gmlbc3. The DNA-binding domain of CPP1 contains two similar Cys-rich domains with 9 and 10 Cys, respectively. Genes encoding similar domains have been identified in Arabidopsis thaliana, Caenorhabditis elegans, the mouse, and human. The domains also have some homology to a Cys-rich region present in some polycomb proteins. The cpp1 gene is induced late in nodule development and the expression is confined to the distal part of the central infected tissue of the nodule. A constitutively expressed cpp1 gene reduces the expression of a Gmlbc3 promoter-gusA reporter construct in Vicia hirsuta roots. These data therefore suggest that CPP1 might be involved in the regulation of the leghemoglobin genes in the symbiotic root nodule. PMID- 10859346 TI - Acid potentiation of the capsaicin receptor determined by a key extracellular site. AB - The capsaicin (vanilloid) receptor, VR1, is a sensory neuron-specific ion channel that serves as a polymodal detector of pain-producing chemical and physical stimuli. The response of VR1 to capsaicin or noxious heat is dynamically potentiated by extracellular protons within a pH range encountered during tissue acidosis, such as that associated with arthritis, infarction, tumor growth, and other forms of injury. A molecular determinant for this important physiological activity was localized to an extracellular Glu residue (E600) in the region linking the fifth transmembrane domain with the putative pore-forming region of the channel. We suggest that this residue serves as a key regulatory site of the receptor by setting sensitivity to other noxious stimuli in response to changes in extracellular proton concentration. We also demonstrate that protons, vanilloids, and heat promote channel opening through distinct pathways, because mutations at a second site (E648) selectively abrogate proton-evoked channel activation without diminishing responses to other noxious stimuli. Our findings provide molecular evidence for stimulus-specific steps in VR1 activation and offer strategies for the development of novel analgesic agents. PMID- 10859347 TI - The origin of intermediary metabolism. AB - The core of intermediary metabolism in autotrophs is the citric acid cycle. In a certain group of chemoautotrophs, the reductive citric acid cycle is an engine of synthesis, taking in CO(2) and synthesizing the molecules of the cycle. We have examined the chemistry of a model system of C, H, and O that starts with carbon dioxide and reductants and uses redox couples as the energy source. To inquire into the reaction networks that might emerge, we start with the largest available database of organic molecules, Beilstein on-line, and prune by a set of physical and chemical constraints applicable to the model system. From the 3.5 million entries in Beilstein we emerge with 153 molecules that contain all 11 members of the reductive citric acid cycle. A small number of selection rules generates a very constrained subset, suggesting that this is the type of reaction model that will prove useful in the study of biogenesis. The model indicates that the metabolism shown in the universal chart of pathways may be central to the origin of life, is emergent from organic chemistry, and may be unique. PMID- 10859348 TI - A psychophysical dissection of the brain sites involved in color-generating comparisons. AB - We have used simple psychophysical methods to determine the sites of color generating mechanisms in the brain. In our first experiment, subjects viewed an abstract multicolored "Mondrian" display through one eye and an isolated patch from the display through the other. With normal binocular/monocular viewing, the patch has a different color when viewed on its own (void mode) or as part of the Mondrian display (natural mode) [Land, E. H. (1974) Proc. R. Inst. G. B. 49, 23 58]. When the two stimuli were viewed dichoptically, with the patch occupying the position that it would occupy in the Mondrian complex under normal viewing, the patch always appeared in its void color. In a second experiment, when subjects viewed multicolored displays through a different narrow-band filter placed over each eye, the information from the two eyes was combined to result in new colors, which were not seen through either of the two eyes alone. Taken together, these results dissect color-generating mechanisms into two stages, located at different sites of the brain: The first occurs before the appearance of binocular neurons in the cortex and compares wavelength information across space, whereas the second occurs after the convergence of the input from the two eyes and synthetically combines the results of the first. PMID- 10859349 TI - Hypoalgesia and altered inflammatory responses in mice lacking kinin B1 receptors. AB - Kinins are important mediators in cardiovascular homeostasis, inflammation, and nociception. Two kinin receptors have been described, B1 and B2. The B2 receptor is constitutively expressed, and its targeted disruption leads to salt-sensitive hypertension and altered nociception. The B1 receptor is a heptahelical receptor distinct from the B2 receptor in that it is highly inducible by inflammatory mediators such as bacterial lipopolysaccharide and interleukins. To clarify its physiological function, we have generated mice with a targeted deletion of the gene for the B1 receptor. B1 receptor-deficient animals are healthy, fertile, and normotensive. In these mice, bacterial lipopolysaccharide-induced hypotension is blunted, and there is a reduced accumulation of polymorphonuclear leukocytes in inflamed tissue. Moreover, under normal noninflamed conditions, they are analgesic in behavioral tests of chemical and thermal nociception. Using whole cell patch-clamp recordings, we show that the B1 receptor was not necessary for regulating the noxious heat sensitivity of isolated nociceptors. However, by using an in vitro preparation, we could show that functional B1 receptors are present in the spinal cord, and their activation can facilitate a nociceptive reflex. Furthermore, in B1 receptor-deficient mice, we observed a reduction in the activity-dependent facilitation (wind-up) of a nociceptive spinal reflex. Thus, the kinin B1 receptor plays an essential physiological role in the initiation of inflammatory responses and the modulation of spinal cord plasticity that underlies the central component of pain. The B1 receptor therefore represents a useful pharmacological target especially for the treatment of inflammatory disorders and pain. PMID- 10859350 TI - Long-term prevention of renal insufficiency, excess matrix gene expression, and glomerular mesangial matrix expansion by treatment with monoclonal antitransforming growth factor-beta antibody in db/db diabetic mice. AB - Emerging evidence suggests that transforming growth factor-beta (TGF-beta) is an important mediator of diabetic nephropathy. We showed previously that short-term treatment with a neutralizing monoclonal anti-TGF-beta antibody (alphaT) in streptozotocin-diabetic mice prevents early changes of renal hypertrophy and increased matrix mRNA. To establish that overactivity of the renal TGF-beta system mediates the functional and structural changes of the more advanced stages of nephropathy, we tested whether chronic administration of alphaT prevents renal insufficiency and glomerulosclerosis in the db/db mouse, a model of type 2 diabetes that develops overt nephropathy. Diabetic db/db mice and nondiabetic db/m littermates were treated intraperitoneally with alphaT or control IgG, 300 microgram three times per week for 8 wk. Treatment with alphaT, but not with IgG, significantly decreased the plasma TGF-beta1 concentration without decreasing the plasma glucose concentration. The IgG-treated db/db mice developed albuminuria, renal insufficiency, and glomerular mesangial matrix expansion associated with increased renal mRNAs encoding alpha1(IV) collagen and fibronectin. On the other hand, treatment with alphaT completely prevented the increase in plasma creatinine concentration, the decrease in urinary creatinine clearance, and the expansion of mesangial matrix in db/db mice. The increase in renal matrix mRNAs was substantially attenuated, but the excretion of urinary albumin factored for creatinine clearance was not significantly affected by alphaT treatment. We conclude that chronic inhibition of the biologic actions of TGF-beta with a neutralizing monoclonal antibody in db/db mice prevents the glomerulosclerosis and renal insufficiency resulting from type 2 diabetes. PMID- 10859351 TI - Molecular cloning and characterization of a lipid phosphohydrolase that degrades sphingosine-1- phosphate and induces cell death. AB - Sphingosine and sphingosine-1-phosphate (SPP) are interconvertible sphingolipid metabolites with opposing effects on cell growth and apoptosis. Based on sequence homology with LBP1, a lipid phosphohydrolase that regulates the levels of phosphorylated sphingoid bases in yeast, we report here the cloning, identification, and characterization of a mammalian SPP phosphatase (mSPP1). This hydrophobic enzyme, which contains the type 2 lipid phosphohydrolase conserved sequence motif, shows substrate specificity for SPP. Partially purified Myc tagged mSPP1 was also highly active at dephosphorylating SPP. When expressed in yeast, mSPP1 can partially substitute for the function of LBP1. Membrane fractions from human embryonic kidney HEK293 cells transfected with mSPP1 markedly degraded SPP but not lysophosphatidic acid, phosphatidic acid, or ceramide-1-phosphate. Enforced expression of mSPP1 in NIH 3T3 fibroblasts not only decreased SPP and enhanced ceramide levels, it also markedly diminished survival and induced the characteristic traits of apoptosis. Collectively, our results suggest that SPP phosphohydrolase may regulate the dynamic balance between sphingolipid metabolite levels in mammalian cells and consequently influence cell fate. PMID- 10859352 TI - Polymerase eta deficiency in the xeroderma pigmentosum variant uncovers an overlap between the S phase checkpoint and double-strand break repair. AB - The xeroderma pigmentosum variant (XPV) is a genetic disease involving high levels of solar-induced cancer that has normal excision repair but shows defective DNA replication after UV irradiation because of mutations in the damage specific polymerase hRAD30. We previously found that the induction of sister chromatid exchanges by UV irradiation was greatly enhanced in transformed XPV cells, indicating the activation of a recombination pathway. We now have identified that XPV cells make use of a homologous recombination pathway involving the hMre11/hRad50/Nbs1 protein complex, but not the Rad51 recombination pathway. The hMre11 complexes form at arrested replication forks, in association with proliferating cell nuclear antigen. In x-ray-damaged cells, in contrast, there is no association between hMre11 and proliferating cell nuclear antigen. This recombination pathway assumes greater importance in transformed XPV cells that lack a functional p53 pathway and can be detected at lower frequencies in excision-defective XPA fibroblasts and normal cells. DNA replication arrest after UV damage, and the associated S phase checkpoint, is therefore a complex process that can recruit a recombination pathway that has a primary role in repair of double-strand breaks from x-rays. The symptoms of elevated solar carcinogenesis in XPV patients therefore may be associated with increased genomic rearrangements that result from double-strand breakage and rejoining in cells of the skin in which p53 is inactivated by UV-induced mutations. PMID- 10859353 TI - PapD-like chaperones provide the missing information for folding of pilin proteins. AB - A fundamental question in molecular biology is how proteins fold into domains that can serve as assembly modules for building up large macromolecular structures. The biogenesis of pili on the surface of Gram-negative bacteria requires the orchestration of a complex process that includes protein synthesis, folding via small chaperones, secretion, and assembly. The results presented here support the hypothesis that pilus subunit folding and biogenesis proceed via mechanisms termed donor strand complementation and donor strand exchange. Here we show that the steric information necessary for pilus subunit folding is not contained in one polypeptide sequence. Rather, the missing information is transiently donated by a strand of a small chaperone to allow folding. Providing the missing information for folding, via a 13-amino acid peptide extension to the C-terminal end of a pilus subunit, resulted in the production of a protein that no longer required the chaperone to fold. This mechanism of small periplasmic chaperone function described here deviates from classical hsp60 chaperone assisted folding. PMID- 10859354 TI - Exploring the sequence space for tetracycline-dependent transcriptional activators: novel mutations yield expanded range and sensitivity. AB - Regulatory elements that control tetracycline resistance in Escherichia coli were previously converted into highly specific transcription regulation systems that function in a wide variety of eukaryotic cells. One tetracycline repressor (TetR) mutant gave rise to rtTA, a tetracycline-controlled transactivator that requires doxycycline (Dox) for binding to tet operators and thus for the activation of P(tet) promoters. Despite the intriguing properties of rtTA, its use was limited, particularly in transgenic animals, because of its relatively inefficient inducibility by doxycycline in some organs, its instability, and its residual affinity to tetO in absence of Dox, leading to elevated background activities of the target promoter. To remove these limitations, we have mutagenized tTA DNA and selected in Saccharomyces cerevisiae for rtTA mutants with reduced basal activity and increased Dox sensitivity. Five new rtTAs were identified, of which two have greatly improved properties. The most promising new transactivator, rtTA2(S)-M2, functions at a 10-fold lower Dox concentration than rtTA, is more stable in eukaryotic cells, and causes no background expression in the absence of Dox. The coding sequences of the new reverse TetR mutants fused to minimal activation domains were optimized for expression in human cells and synthesized. The resulting transactivators allow stringent regulation of target genes over a range of 4 to 5 orders of magnitude in stably transfected HeLa cells. These rtTA versions combine tightness of expression control with a broad regulatory range, as previously shown for the widely applied tTA. PMID- 10859355 TI - Amplification of the human dihydrofolate reductase gene via double minutes is initiated by chromosome breaks. AB - DNA sequence amplification is one of the most frequent manifestations of genomic instability in human tumors. We have shown previously that amplification of the dihydrofolate reductase (DHFR) gene in Chinese hamster cells is initiated by chromosome breaks, followed by bridge-breakage-fusion cycles that generate large intrachromosomal repeats; these are ultimately trimmed by an unknown process to smaller, more homogenous units manifested as homogenously staining chromosome regions (HSRs). However, in most human tumor cells, amplified DNA sequences are borne on unstable, extrachromosomal double minutes (DMs), which suggests the operation of a different amplification mechanism. In this study, we have isolated a large number of independent methotrexate-resistant human cell lines, all of which contained DHFR-bearing DMs. Surprisingly, all but one of these also had suffered partial or complete loss of one of the parental DHFR-bearing chromosomes. Cells in a few populations displayed what could be transient intermediates in the amplification process, including an initial HSR, its subsequent breakage, the appearance of DHFR-containing fragments, and, finally, DMs. Our studies suggest that HSRs and DMs both are initiated by chromosome breaks, but that cell types differ in how the extra sequences ultimately are processed and/or maintained. PMID- 10859356 TI - Differences in the polar clustering of the high- and low-abundance chemoreceptors of Escherichia coli. AB - The chemosensory complexes in Escherichia coli are localized predominantly in large aggregates at one or both of the cell poles, however, neither the role of the polar localization nor the role of the clustering is understood. In E. coli, the two classes of chemoreceptors or transducers, high- and low-abundance, differ in their ability to support chemotaxis when expressed as the sole chemoreceptor type in the cell. In this study, we examined both the contribution of individual chemoreceptors to polar clustering and the ability of each chemoreceptor type to cluster in the absence of all others. We found that polar clustering of methyl accepting chemotaxis proteins (MCPs) is not dependent on any one chemoreceptor type. Remarkably, when expressed individually at similar levels, the chemoreceptors display differential clustering abilities. The high-abundance transducers cluster at the cell pole almost as well as do the MCPs in cells expressing all four species, whereas the low-abundance transducers, although polar, are not particularly clustered. CheA and CheW distributions in strains expressing only one chemoreceptor type coincide with MCP localization, indicating that the low-abundance chemoreceptors are competent for ternary complex formation but are defective in aggregation. These studies reveal that, in contrast to our previous model, polarity of the chemoreceptors is independent of clustering, suggesting that the polar localization of the chemoreceptors is not simply caused by diffusion limitations on large protein aggregates. PMID- 10859357 TI - Identification and characterization of a newly recognized population of high-Na+, low-K+, low-density sickle and normal red cells. AB - We describe a population of sickle cell anemia red cells (SS RBCs) ( approximately 4%) and a smaller fraction of normal RBCs (<0.03%) that fail to dehydrate when permeabilized to K(+) with either valinomycin or elevated internal Ca(2+). The nonshrinking, valinomycin-resistant (val-res) fractions, first detected by flow cytometry of density-fractionated SS RBCs, constituted up to 60% of the lightest, reticulocyte-rich (R1) cell fraction, and progressively smaller portions of the slightly denser R2 cells and discocytes. R1 val-res RBCs had a mean cell hemoglobin concentration of approximately 21 g of Hb per dl, and many had an elongated shape like "irreversibly sickled cells," suggesting a dense SS cell origin. Of three possible explanations for val-res cells, failure of valinomycin to K(+)-permeabilize the cells, low co-ion permeability, or reduced driving K(+) gradient, the latter proved responsible: Both SS and normal val-res RBCs were consistently high-Na(+) and low-K(+), even when processed entirely in Na-free media. Ca(2+) + A23187-induced K(+)-permeabilization of SS R1 fractions revealed a similar fraction of cal-res cells, whose (86)Rb uptake showed both high Na/K pump and leak fluxes. val-res/cal-res RBCs might represent either a distinct erythroid genealogy, or an "end-stage" of normal and SS RBCs. This paper focuses on the discovery, basic characterization, and exclusion of artifactual origin of this RBC fraction. Many future studies will be needed to clarify their mechanism of generation and full pathophysiological significance. PMID- 10859358 TI - Dynamic spike threshold reveals a mechanism for synaptic coincidence detection in cortical neurons in vivo. AB - Cortical neurons are sensitive to the timing of their synaptic inputs. They can synchronize their firing on a millisecond time scale and follow rapid stimulus fluctuations with high temporal precision. These findings suggest that cortical neurons have an enhanced sensitivity to synchronous synaptic inputs that lead to rapid rates of depolarization. The voltage-gated currents underlying action potential generation may provide one mechanism to amplify rapid depolarizations. We have tested this hypothesis by analyzing the relations between membrane potential fluctuations and spike threshold in cat visual cortical neurons recorded intracellularly in vivo. We find that visual stimuli evoke broad variations in spike threshold that are caused in large part by an inverse relation between spike threshold and the rate of membrane depolarization preceding a spike. We also find that spike threshold is inversely related to the rate of rise of the action potential upstroke, suggesting that increases in spike threshold result from a decrease in the availability of Na(+) channels. By using a simple neuronal model, we show that voltage-gated Na(+) and K(+) conductances endow cortical neurons with an enhanced sensitivity to rapid depolarizations that arise from synchronous excitatory synaptic inputs. Thus, the basic mechanism responsible for action potential generation also enhances the sensitivity of cortical neurons to coincident synaptic inputs. PMID- 10859359 TI - Criteria for the control of drug-resistant tuberculosis. AB - Antibiotic resistance is a growing impediment to the control of infectious diseases worldwide, tuberculosis (TB) being among them. TB kills two million people each year and foci of multidrug-resistant TB (MDR-TB) have been identified in Eastern Europe, Africa, Asia, and Latin America. A critical question for health policy is whether standardized short-course chemotherapy for TB, based on cheap first-line drugs, can prevent and reverse the spread of drug resistance. Here we use mathematical modeling, in conjunction with treatment results from six countries, to show that best-practice short-course chemotherapy is highly likely to bring strains resistant to either of the two key drugs isoniazid and rifampicin under control and to prevent the emergence of MDR-TB. However, it is not certain to contain MDR-TB once it has emerged, partly because cure rates are too low. We estimate that approximately 70% of prevalent, infectious MDR-TB cases should be detected and treated each year, and at least 80% of these cases should be cured, in order to prevent outbreaks of MDR-TB. Poor control programs should aim to increase case detection and cure rates together for three reasons: (i) these variables act synergistically; (ii) when either is low, the other cannot succeed alone; and (iii) the second-line drugs needed to raise MDR-TB cure rates are few and extremely costly. We discuss the implications of these results for World Health Organization policy on the management of antibiotic resistance. PMID- 10859360 TI - Positive cooperativity without domains or subunits in a monomeric membrane channel. AB - The monomeric VDAC channel shows an accelerated pH titration of its transport properties with a Hill coefficient of about 2. This manifests itself as a sharp peak in conductance noise as well as a fast change in channel selectivity with pH. On the basis of the known structure of this channel, we propose that this cooperativity arises from a mechanically linked mobile pair of ionizable groups. Concerted movement of these groups between two states changes the distance from nearby electrostatic charge to influence the pK of the groups. This model of pH dependent motion produces positive cooperative behavior that fits the observations without need for subunits or identifiable domains within the protein. The mathematical formalism has never required such domains, but these are generally considered an essential part of cooperative behavior in proteins. The present proposal reduces the size of a cooperative unit to a minimum, extending the limits of what is perceived to be possible. Together with large scale conformational transitions, these subtle cooperative structural changes may allow proteins to adapt, with high sensitivity, to changes in their environment. They might also be relatively easy to engineer into a protein. PMID- 10859361 TI - Prediction of cytomegalovirus load and resistance patterns after antiviral chemotherapy. AB - Cytomegalovirus (CMV) replicates rapidly in its human host with a doubling time of approximately 1 day. Using simple mathematical models we show that the efficacy of the anti-CMV drug ganciclovir (GCV) against wild-type strains is 91.5% [95% confidence interval (CI) 89-94%] when given i.v. (5 mg/kg twice a day) but only 46.5% (95% CI 45-47.5%) when given orally (1 g three times a day) whereas the corresponding figures for a typical GCV-resistant virus are 62% (95% CI 57-66%) and 35% (95% CI 33-37%), respectively. During prolonged periods of GCV therapy we show that the apparent sudden appearance of GCV resistance is explained by the combination of two exponentially increasing populations (wild type and mutant) at doses of GCV that do not completely inhibit CMV replication. Cell culture methods to assess CMV drug resistance in vivo will underestimate its prevalence because of the fitness loss of resistant virus in the absence of therapy. The parameters determined from these models then were used to predict the likely viral load and resistance patterns in patients on prolonged therapy with GCV. The modeled and experimental data showed excellent agreement over extended time periods (up to 270 days of therapy) and provide a framework to predict the virologic course of patients at therapeutic initiation. PMID- 10859362 TI - Kaposi's sarcoma-associated herpesvirus encodes two proteins that block cell surface display of MHC class I chains by enhancing their endocytosis. AB - Down-regulation of the cell surface display of class I MHC proteins is an important mechanism of immune evasion by human and animal viruses. Herpesviruses in particular encode a variety of proteins that function to lower MHC I display by several mechanisms. These include binding and retention of MHC I chains in the endoplasmic reticulum, dislocation of class I chains from the ER, inhibition of the peptide transporter (TAP) involved in antigen presentation, and shunting of newly assembled chains to lysosomes. Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is a human herpesvirus strongly linked to the development of KS and to certain AIDS-associated lymphoproliferative disorders. Here we show that KSHV encodes two distinctive gene products that function to dramatically reduce cell surface MHC I expression. These viral proteins are localized predominantly to the ER. However, unlike previously described MHC I inhibitors, they do not interfere with the synthesis, translocation, or assembly of class I chains, nor do they retain them in the ER. Rather, they act to enhance endocytosis of MHC I from the cell surface; internalized class I chains are delivered to endolysosomal vesicles, where they undergo degradation. These KSHV proteins define a mechanism of class I down-regulation distinct from the mechanisms of other herpesviruses and are likely to contribute importantly to immune evasion during viral infection. PMID- 10859363 TI - Amino acid transport system A resembles system N in sequence but differs in mechanism. AB - Classical amino acid transport System A accounts for most of the Na(+)-dependent neutral amino acid uptake by mammalian cells. System A has also provided a paradigm for short- and long-term regulation by physiological stimuli. We now report the isolation of a cDNA encoding System A that shows close similarity to the recently identified System N transporter (SN1). The System A transporter (SA1) and SN1 share many functional characteristics, including a marked sensitivity to low pH, but, unlike SN1, SA1 does not mediate proton exchange. Transport mediated by SA1 is also electrogenic. Amino acid transport Systems A and N thus appear closely related in function as well as structure, but exhibit important differences in ionic coupling. PMID- 10859364 TI - Genetic susceptibility to tuberculosis in Africans: a genome-wide scan. AB - Human genetic variation is an important determinant of the outcome of infection with Mycobacterium tuberculosis. We have conducted a two-stage genome-wide linkage study to search for regions of the human genome containing tuberculosis susceptibility genes. This approach uses sibpair families that contain two full siblings who have both been affected by clinical tuberculosis. For any chromosomal region containing a major tuberculosis-susceptibility gene, affected sibpairs inherit the same parental alleles more often than expected by chance. In the first round of the screen, 299 highly informative genetic markers, spanning the entire human genome, were typed in 92 sibpairs from The Gambia and South Africa. Seven chromosomal regions that showed provisional evidence of coinheritance with clinical tuberculosis were identified. To identify whether any of these regions contained a potential tuberculosis-susceptibility gene, 22 markers from these regions were genotyped in a second set of 81 sibpairs from the same countries. Markers on chromosomes 15q and Xq showed suggestive evidence of linkage (lod = 2.00 and 1.77, respectively) to tuberculosis. The potential identification of susceptibility loci on both chromosomes 15q and Xq was supported by an independent analysis designated common ancestry using microsatellite mapping. These results indicate that genome-wide linkage analysis can contribute to the mapping and identification of major genes for multifactorial infectious diseases of humans. An X chromosome susceptibility gene may contribute to the excess of males with tuberculosis observed in many different populations. PMID- 10859365 TI - Hsp70 and hsp40 chaperones can inhibit self-assembly of polyglutamine proteins into amyloid-like fibrils. AB - The deposition of protein aggregates in neurons is a hallmark of neurodegenerative diseases caused by polyglutamine (polyQ) proteins. We analyzed the effects of the heat shock protein (Hsp) 70 chaperone system on the aggregation of fragments of huntingtin (htt) with expanded polyQ tracts. In vitro, Hsp70 and its cochaperone Hsp40 suppressed the assembly of htt into detergent-insoluble amyloid-like fibrils in an ATP-dependent manner and caused the formation of amorphous, detergent-soluble aggregates. The chaperones were most active in preventing fibrillization when added during the lag phase of the polymerization reaction. Similarly, coexpression of Hsp70 or Hsp40 with htt in yeast inhibited the formation of large, detergent-insoluble polyQ aggregates, resulting in the accumulation of detergent-soluble inclusions. Thus, the recently established potency of Hsp70 and Hsp40 to repress polyQ-induced neurodegeneration may be based on the ability of these chaperones to shield toxic forms of polyQ proteins and to direct them into nontoxic aggregates. PMID- 10859366 TI - A mutation in Rab27a causes the vesicle transport defects observed in ashen mice. AB - The dilute (d), leaden (ln), and ashen (ash) mutations provide a unique model system for studying vesicle transport in mammals. All three mutations produce a lightened coat color because of defects in pigment granule transport. In addition, all three mutations are suppressed by the semidominant dilute suppressor (dsu), providing genetic evidence that these mutations function in the same or overlapping transport pathways. Previous studies showed that d encodes a major vesicle transport motor, myosin-VA, which is mutated in Griscelli syndrome patients. Here, using positional cloning and bacterial artificial chromosome rescue, we show that ash encodes Rab27a. Rab GTPases represent the largest branch of the p21 Ras superfamily and are recognized as key players in vesicular transport and organelle dynamics in eukaryotic cells. We also show that ash mice have platelet defects resulting in increased bleeding times and a reduction in the number of platelet dense granules. These defects have not been reported for d and ln mice. Collectively, our studies identify Rab27a as a critical gene for organelle-specific protein trafficking in melanocytes and platelets and suggest that Rab27a functions in both MyoVa dependent and independent pathways. PMID- 10859367 TI - Genome-wide scan for body composition in pigs reveals important role of imprinting. AB - The role of imprinting in body composition was investigated in an experimental cross between Chinese Meishan pigs and commercial Dutch pigs. A whole-genome scan revealed significant evidence for five quantitative trait loci (QTL) affecting body composition, of which four were imprinted. Imprinting was tested with a statistical model that separated the expression of paternally and maternally inherited alleles. For back fat thickness, a paternally expressed QTL was found on Sus scrofa chromosome 2 (SSC2), and a Mendelian-expressed QTL was found on SSC7. In the same region of SSC7, a maternally expressed QTL affecting muscle depth was found. Chromosome 6 harbored a maternally expressed QTL on the short arm and a paternally expressed QTL on the long arm, both affecting intramuscular fat content. The individual QTL explained from 2% up to 10% of the phenotypic variance. The known homologies to human and mouse did not reveal positional candidate genes. This study demonstrates that testing for imprinting should become a standard procedure to unravel the genetic control of multifactorial traits. PMID- 10859368 TI - Replication of hepatitis C virus. PMID- 10859369 TI - Mutational analysis of hepatitis C virus NS3-associated helicase. AB - Nonstructural protein 3 (NS3) of hepatitis C virus contains a bipartite structure consisting of an N-terminal serine protease and a C-terminal DEXH box helicase. To investigate the roles of individual amino acid residues in the overall mechanism of unwinding, a mutational-functional analysis was performed based on a molecular model of the NS3 helicase domain bound to ssDNA, which has largely been confirmed by a recently published crystal structure of the NS3 helicase-ssDNA complex. Three full-length mutated NS3 proteins containing Tyr(392)Ala, Val(432)Gly and Trp(501)Ala single substitutions, respectively, together with a Tyr(392)Ala/Trp(501)Ala double-substituted protein were expressed in Escherichia coli and purified to homogeneity. All individually mutated forms showed a reduction in duplex unwinding activity, single-stranded polynucleotide binding capacity and polynucleotide-stimulated ATPase activity compared to wild-type, though to different extents. Simultaneous replacement of both Tyr(392) and Trp(501) with Ala completely abolished all these enzymatic functions. On the other hand, the introduced amino acid substitutions had no influence on NS3 intrinsic ATPase activity and proteolytic efficiency. The results obtained with Trp(501)Ala and Val(432)Gly single-substituted enzymes are in agreement with a recently proposed model for NS3 unwinding activity. The mutant phenotype of the Tyr(392)Ala and Tyr(392)Ala/Trp(501)Ala enzymes, however, represents a completely novel finding. PMID- 10859370 TI - Dengue virus type 1 DNA vaccine induces protective immune responses in rhesus macaques. AB - A candidate DNA vaccine expressing dengue virus type 1 pre-membrane and envelope proteins was used to immunize rhesus macaques. Monkeys were immunized intramuscularly (i.m.) or intradermally (i.d.) by three or four 1 mg doses of vaccine, respectively. Monkeys that were inoculated i.m. seroconverted more quickly and had higher antibody levels than those that were inoculated i.d. The sera exhibited virus-neutralizing activity, which declined over time. Four of the eight i.m.-inoculated monkeys were protected completely from developing viraemia when challenged 4 months after the last dose with homologous dengue virus. The other four monkeys had reduced viraemia compared with the control immunized monkeys. The i.d. -inoculated monkeys showed no reduction in viraemia when challenged with the virus. All vaccinated monkeys showed an anamnestic antibody response, indicating that they had established immunological memory. Vaccine induced antibody had an avidity index similar to that of antibody induced by virus infection; however, no clear correlation was apparent between antibody avidity and virus neutralization titres. PMID- 10859371 TI - Germinal centre localization of bovine viral diarrhoea virus in persistently infected animals. AB - Immunohistochemical analysis of peripheral lymph nodes from gnotobiotic calves persistently infected with bovine viral diarrhoea virus (BVDV) revealed extensive deposition of E(rns) and localization of the viral genome in the light zone of germinal centres. Viral antigen co-localized with immunoglobulin in the germinal centres and was shown to be extracellular. Despite the presence of viral antigen in germinal centres, circulating anti-BVDV antibody was not detected. These findings provide evidence that calves persistently infected with BVDV, in the absence of adventitious infection, can generate a B cell response to the persisting virus. The nature of the tolerance in calves persistently infected with BVDV is discussed in light of these findings. PMID- 10859372 TI - The complete sequence of hepatitis E virus genotype 4 reveals an alternative strategy for translation of open reading frames 2 and 3. AB - Isolates of hepatitis E virus (HEV) have recently been described from China that are distinct from Burmese, Mexican and US viruses and constitute a novel genotype (genotype 4). Here, the complete genomic sequence of a representative isolate of genotype 4 HEV, amplified directly from the stool of an acutely infected patient, is presented. Analysis of the entire sequence confirms our previous conclusion, based upon partial sequence data, that these Chinese isolates belong to a novel genotype. Typical of genetic variation in HEV, most nucleotide substitutions occur in the third base of the codon and do not affect the amino acid sequence. The genotype 4 virus is unusual in that a single nucleotide insertion in the ORF 3 region changes the initiation of ORF 3, and perhaps also ORF 2. The consequences of these changes are discussed. PMID- 10859373 TI - Expression of reporter genes from the defective RNA CD-61 of the coronavirus infectious bronchitis virus. AB - The defective RNA (D-RNA) CD-61, derived from the Beaudette strain of the avian coronavirus infectious bronchitis virus (IBV), was used as an RNA vector for the expression of two reporter genes, luciferase and chloramphenicol acetyltransferase (CAT). D-RNAs expressing the CAT gene were demonstrated to be capable of producing CAT protein in a helper-dependent expression system to about 1.6 microgram per 10(6) cells. The reporter genes were expressed from two different sites within the CD-61 sequence and expression was not affected by interruption of the CD-61-specific ORF. Expression of the reporter genes was under the control of a transcription-associated sequence (TAS) derived from the Beaudette gene 5, normally used for the transcription of IBV subgenomic mRNA 5. The Beaudette gene 5 TAS is composed of two tandem repeats of the IBV canonical consensus sequence involved in the acquisition of a leader sequence during the discontinuous transcription of IBV subgenomic mRNAs. It is demonstrated that only one canonical sequence is required for expression of mRNA 5 or for the expression of an mRNA from a D-RNA and that either sequence can function as an acceptor site for acquisition of the leader sequence. PMID- 10859374 TI - Replication-competent foot-and-mouth disease virus RNAs lacking capsid coding sequences. AB - RNA transcripts were prepared from plasmids encoding an infectious cDNA of foot and-mouth disease virus (FMDV) or derivatives in which the leader (Lab and Lb) and capsid protein coding sequences were deleted or replaced by sequences encoding chloramphenicol acetyltransferase (CAT). The transcripts were electroporated into BHK cells and the expression of CAT and the FMDV 3C protease was monitored. Detection of CAT and 3C was dependent on the ability of the transcript to replicate. All of the Lb coding sequence and 94% of P1 (the capsid protein precursor) coding sequence could be deleted without any apparent effect on the ability of the RNA to replicate. Thus, no cis-acting replication element is present within this region of the FMDV genome. TRANS:-encapsidation of these FMDV replicons was very inefficient, which may explain the lack of production of defective-interfering particles in FMDV-infected cells. PMID- 10859375 TI - Caspases are not involved in the cleavage of translation initiation factor eIF4GI during picornavirus infection. AB - Infection of cells by many picornaviruses results in the rapid inhibition of cellular protein synthesis due to cleavage of the translation initiation factor eIF4G. The poliovirus (PV) 2A and foot-and-mouth disease virus (FMDV) L proteases are each sufficient to mediate this cleavage, but the cleavage mechanism may be indirect, involving an unidentified cellular protease(s). eIF4G is also targetted for cleavage by caspase-3 during apoptosis. Here, it is shown that caspase inhibitors do not inhibit the cleavage of eIF4GI during PV or FMDV infection. Similarly, in transient-expression studies, the cleavage of eIF4GI induced by PV 2A or FMDV L was unaffected by these inhibitors. Furthermore, the cleavage of eIF4GI was observed in PV-infected MCF-7 cells lacking caspase-3. These data, and the fact that induction of apoptosis yields different eIF4GI cleavage fragments, indicate that caspases do not have a major role in the cleavage of eIF4GI during PV or FMDV infection. PMID- 10859376 TI - Antigenic properties of human parechovirus 1. AB - Human parechoviruses 1 and 2 (HPEV1 and HPEV2, respectively), formerly known as echoviruses 22 and 23, have been assigned to a novel picornavirus genus on the basis of their distinct molecular and biological properties. To study the immunological characteristics of HPEV1 capsid proteins, antigenic analysis was carried out by a peptide scanning technique, which can be used to identify the immunogenic peptide sequences of a protein. Partially overlapping peptides, representing the capsid of HPEV1, were synthesized using a 12 aa window in a three residue shift and reactivity of rabbit and murine HPEV1 antisera against these peptides were tested. Using this method, an antigenic site in the VP0 polypeptide, recognized by both rabbit and murine antisera, was identified. The sequence of this region was conserved among HPEV1 clinical isolates obtained from Finland and the United States. Antiserum against this peptide region showed neutralizing activity against HPEV1 in cell culture. Because the C-terminal region of HPEV1 VP1 contains a functional RGD motif, the antigenicity of this region was also tested. By using the corresponding peptide antiserum, neutralization of HPEV1 was observed. Cross-neutralization between HPEV1 and coxsackievirus A9, an enterovirus with a similar RGD motif in VP1, was also detected. PMID- 10859377 TI - The rate of progression to AIDS is independent of virus dose in simian immunodeficiency virus-infected macaques. AB - Of the viral factors that are proposed to influence the rate of progression to AIDS, the role of infectious dose remains unresolved. Intravenous infection of outbred Macaca mulatta with various doses of simian immunodeficiency virus isolate 8980 (SIV(8980)) revealed an endpoint from which an infectious dose 50 (ID(50)) was defined. In the six infected animals, the time to develop AIDS was variable with a spectrum of rapid, intermediate and slow progressors. High and sustained plasma viraemia with marked loss of CD4(+) T-cells was a distinguishing feature between rapid versus intermediate and slow progressors. Animals that received the highest doses did not develop the highest sustained viral loads, nor did they progress more rapidly to disease. Similarly, animals infected with lower doses did not uniformly develop lower viral loads or progress more slowly to AIDS. Furthermore, compiled data from more than 21 animals infected with different doses of the same virus administered by the same route failed to reveal any correlation of infectious dose with survival. Indeed, host factors of these outbred animals, rather than dose of the initial inoculum, were probably an important factor influencing the rate of disease progression in each individual animal. Comparison of animals infected with SIV(B670), from which SIV(8980) was derived, revealed marked differences in disease progression. Clearly, although dose did not influence viral loads nor disease progression, the virulence of the initial inoculum was a major determinant of the rate of progression to AIDS. PMID- 10859378 TI - The strong positive correlation between effective affinity and infectivity neutralization of highly cross-reactive monoclonal antibody IIB4, which recognizes antigenic site B on influenza A virus haemagglutinin. AB - Monoclonal antibody (MAb) IIB4 displays a rare combination of virus neutralization (VN) activity and broad cross-reactivity with influenza A virus strains of the H3 subtype isolated in a period from 1973 to 1988. The epitope of this antibody has been identified as around HA1 residues 198, 199 and 201. Here we report that residues 155, 159, 188, 189 and 193 also influence the binding of this antibody. We have used this antibody to study the relationship between antibody affinity and VN activity. Using one MAb and a single epitope on the haemagglutinin (HA) of different influenza viruses we found a strong positive correlation between effective affinity and VN activity of MAb IIB4. A 10-fold increase in effective affinity corresponded to the 2000-fold increase in VN titre. It follows from the law of mass action that for an effective affinity K=9x10(8) l/mol, 50% VN was achieved at approx. 10% occupation of HA spikes with antibody. In contrast, for an effective affinity K=6x10(7) l/mol, to achieve 50% VN, occupation of up to 98% of HA spikes was required. An effective affinity about K=6x10(7) l/mol thus represents the limiting value for VN because a further decrease in the affinity cannot be compensated by a higher concentration of antibody. PMID- 10859379 TI - Plasmid DNA encoding influenza virus haemagglutinin induces Th1 cells and protection against respiratory infection despite its limited ability to generate antibody responses. AB - Direct intramuscular injection of plasmid DNA can generate immune responses against encoded antigens. However, the relative ability of DNA vaccines to induce cellular and humoral immunity after a single or booster immunization and the persistence of this response have not been fully elucidated. In this study, induction and maintenance of antibody and T cell subtypes with different doses of naked DNA encoding the haemagglutinin (HA) gene of influenza virus were examined and compared to the immune responses and protection induced by respiratory tract infection and immunization with a killed virus vaccine. Like natural infection, immunization with HA DNA induced potent Th1 responses. Spleen cells from mice immunized once with HA DNA in the dose range 10 ng to 100 microgram secreted significant levels of IFN-gamma, but low or undetectable IL-5, in response to influenza virus in vitro. Furthermore, CD4(+) HA-specific Th1 clones were generated from spleens of immunized mice. Although T cell responses waned 12 weeks after a single immunization, antigen-specific Th1 cells persisted in the spleen for at least 6 months after two booster immunizations. In contrast, influenza virus-specific ELISA IgG titres were low after a single immunization and required two booster immunizations to reach significant levels. Furthermore, haemagglutination inhibition (HI) antibodies were weak or undetectable after two immunizations. Nevertheless, two doses of HA DNA conferred almost complete protection against respiratory challenge with live virus. Thus, despite the limited ability to induce antibodies, DNA vaccines confer protective immunity against influenza virus infection, which appears to be mediated by Th1 cells. PMID- 10859380 TI - Genetic analysis of wild-type Dobrava hantavirus in Slovenia: co-existence of two distinct genetic lineages within the same natural focus. AB - Genetic analysis was performed of wild-type (wt) Dobrava hantavirus (DOB) strains from Slovenia, the country where the virus was first discovered and where it was found to cause haemorrhagic fever with renal syndrome (HFRS), with a fatality rate of 12%. Two hundred and sixty mice of the genus APODEMUS:, trapped in five natural foci of DOB-associated HFRS during 1990-1996, were screened for the presence of anti-hantavirus antibodies and 49 APODEMUS: flavicollis and four APODEMUS: agrarius were found to be positive. RT-PCR was used to recover partial sequences of the wt-DOB medium (M) and small (S) genome segments from nine A. flavicollis and one A. agrarius. Sequence comparison and phylogenetic analysis of the Slovenian wt-DOB strains revealed close relatedness of all A. flavicollis derived virus sequences (nucleotide diversity up to 6% for the M segment and 5% for the S segment) and the geographical clustering of genetic variants. In contrast, the strain harboured by A. agrarius showed a high level of genetic diversity from other Slovenian DOB strains (14%) and clustered together on phylogenetic trees with other DOB strains harboured by A. agrarius from Russia, Estonia and Slovakia. These findings suggest that the DOB variants carried by the two species of APODEMUS: in Europerepresent two distinct genetic lineages. PMID- 10859381 TI - Neither phosphorylation nor the amino-terminal part of rabies virus phosphoprotein is required for its oligomerization. AB - Rabies virus (PV strain) phosphoprotein (P) was expressed in bacteria. This recombinant protein binds specifically to the nucleoprotein-RNA complex purified from infected cells. Chemical cross-linking and gel-filtration studies indicated that the P protein forms oligomers. Analytical centrifugation data demonstrated the co-existence of monomeric and oligomeric forms of rabies virus P protein and suggested that there is an equilibrium between these species. As P expressed in bacteria is not phosphorylated, this result indicates that P phosphorylation is not required for its oligomerization. Although an alignment of several rhabdovirus P sequences revealed that the amino-terminal domain of P has a conserved predicted propensity to form helical coiled coils, an amino-terminally truncated form of P protein, lacking the first 52 residues, was also shown to be oligomeric. Therefore, the amino-terminal domain of rabies virus P is not necessary for its oligomerization. PMID- 10859382 TI - The role of the UL41 gene of herpes simplex virus type 1 in evasion of non specific host defence mechanisms during primary infection. AB - The UL41 gene product (vhs) of herpes simplex virus (HSV) is packaged in the virion, and mediates host protein synthesis shutoff at the early stage of the virus replication cycle. In order to clarify the role of vhs in virus replication and virulence, we isolated a completely UL41-deficient mutant (the VRDelta41 strain) and its revertant (the VRDelta41R strain). In the mouse encephalitis model, the replication of strain VRDelta41 was inhibited after 2 days post infection, resulting in low virulence, by gamma-ray-sensitive cells such as lymphocytes and/or neutrophils. The result suggested that some cytokines, produced in VRDelta41-inoculated brains, activate and induce the migration of gamma-ray-sensitive cells to the infection site. Therefore, cytokines produced by HSV-1-infected human cells were screened, and potent inductions of interleukin (IL)-1beta, IL-8 and macrophage inflammatory protein-1alpha by VRDelta41 infection were observed. Moreover, the VRDelta41 strain showed 20- and 5-fold higher sensitivity to interferon-alpha and -beta compared to the wild-type strain, respectively. These results indicate that one important role of vhs in vivo is evasion from non-specific host defence mechanisms during primary infection through suppression of cytokine production in HSV-infected cells and reduction of the anti-HSV activity of interferon-alpha and -beta. PMID- 10859383 TI - Cross-reactivity of the anti-PML antibody PG-M3 with the herpes simplex virus type 1 immediate early protein ICP4. AB - The PML protein is one of the components of ND10, nuclear matrix-associated structures which undergo rapid disintegration at the onset of herpes simplex virus type 1 (HSV-1) infection. This disruption event has been frequently visualized in immunofluorescence assays using the anti-PML mouse monoclonal antibody PG-M3. This antibody was surprisingly found to also stain nuclear virus replication compartments when employed at higher concentrations. This was shown to be due to an unexpected cross-reactivity of the PG-M3 antibody with the HSV-1 immediate early protein ICP4, a known component of replication compartments. The sequences of ICP4 recognized by PG-M3 were found to map to the extreme amino terminal end of the protein, which includes a 21 amino acid segment that is partially homologous to the peptide of PML that was used to make PG-M3. These results suggest that PG-M3 may no longer represent an appropriate antibody for use in visualizing the fate of PML and ND10 during HSV-1 infection. PMID- 10859384 TI - Human cytomegalovirus UL37 immediate-early regulatory proteins traffic through the secretory apparatus and to mitochondria. AB - The human cytomegalovirus (HCMV) UL36-38 immediate-early (IE) locus encodes the UL37 exon 1 (pUL37x1) and UL37 (gpUL37) regulatory proteins, which have anti apoptotic activities. pUL37x1 shares its entire sequence, including a hydrophobic leader and an acidic domain, with the exception of one residue, with the amino terminus of gpUL37. gpUL37 has, in addition, unique N-linked glycosylation, transmembrane and cytosolic domains. A rabbit polyvalent antiserum was generated against residues 27-40 in the shared amino-terminal domain and a mouse polyvalent antiserum was generated against the full-length protein to study trafficking of individual UL37 proteins in human cells that transiently expressed gpUL37 or pUL37x1. Co-localization studies by confocal laser scanning microscopy detected trafficking of gpUL37 and pUL37x1 from the endoplasmic reticulum to the Golgi apparatus in permissive U373 cells and in human diploid fibroblasts (HFF). Trafficking of gpUL37 to the cellular plasma membrane was detected in unfixed HFF cells. FLAG-tagged gpUL37 trafficked similarly through the secretory apparatus to the plasma membrane. By using confocal microscopy and immunoblotting of fractionated cells, gpUL37 and pUL37x1 were found to co-localize with mitochondria in human cells. This unconventional dual trafficking pattern through the secretory apparatus and to mitochondria is novel for herpesvirus IE regulatory proteins. PMID- 10859385 TI - Characterization of the epstein-barr virus BRRF1 gene, located between early genes BZLF1 and BRLF1. AB - The switch from latency to a productive cycle in Epstein-Barr virus (EBV) infected B cells proliferating in vitro is thought to be due to the transcriptional activation of two viral genes, BZLF1 and BRLF1, encoding two transcription factors called EB1 and R respectively. However, a third gene, BRRF1 is contained in the BZLF1/BRLF1 locus, overlapping with BRLF1 but in inverse orientation. We have characterized the 5' end of the BRRF1 mRNA and the promoter, PNa, at which BRRF1 pre-mRNA is initiated. We show that although a single BRRF1 mRNA species is induced by 12-O-tetradecanoylphorbol 13-acetate/sodium butyrate in several EBV-infected B cell lines, in Akata cells treated with anti-IgG two BRRF1 mRNAs can be detected. Transcription initiated at the BRRF1 promoter was activated by EB1 but not by R, and EB1-binding sites which contribute to the EB1 activated transcription have been mapped to between positions -469 and +1. A 34 kDa protein could be translated from the BRRF1 mRNA both in vitro and in vivo, and was found predominantly in the nucleus of HeLa cells transfected with a BRRF1 expression vector. Thus there are three promoters in the region of the EBV chromatin containing the BZLF1/BRLF1 genes, two of which, PZ and PNa, potentially share regulatory elements. PMID- 10859386 TI - Herpesvirus papio encodes a functional homologue of the Epstein-Barr virus apoptosis suppressor, BHRF1. AB - The human tumour virus Epstein-Barr virus (EBV) encodes a 17 kDa protein, BHRF1, which is a member of the BCL:-2 family and has been shown to suppress apoptosis. The role of this gene in the life-cycle of EBV has not been fully elucidated. In order to identify motifs conserved in herpesviruses and possibly shed light on its function we isolated a BHRF1 homologue from herpesvirus papio (cercopithecine herpesvirus-12) a closely related gammaherpesvirus of baboons. The gene, hvpBHRF1, also encodes a 17 kDa protein which shares 64% identity and 79% similarity with EBV BHRF1 at the amino acid level. In biological assays, hvpBHRF1 and BHRF1 conferred similar levels of protection on human keratinocytes induced to apoptose with cis-platin. PMID- 10859387 TI - Establishment of a cell line persistently infected with bovine herpesvirus-4 by use of a recombinant virus. AB - Bovine herpesvirus-4 (BHV-4), a gammaherpesvirus lacking a clear disease association, productively infects multiple cell lines of various species and causes cell death. A human rhabdomyosarcoma cell line, RD-4, infected with BHV-4 produced low levels of early and late viral RNAs and infectious virus, but exhibited no cytopathic effect. Using a recombinant BHV-4 containing a neomycin resistance gene, we established RD-4-derived cell lines persistently infected with BHV-4. The viral genome in these cells was predominantly circular. Because of drug selection, every cell contained a viral genome. In addition, all cells stained with a BHV-4-specific antiserum. Therefore, these cell lines are not carrier cultures. These cells produced infectious virus at all passages tested. Even though cells were selected and maintained at a concentration of geneticin at least 2.5 times that necessary to kill uninfected RD-4 cells, selected cells contained only approximately one viral genome per diploid host cell genome. Persistently infected cells grew more slowly than uninfected cells, even in the absence of drug. The slower growth of these cells suggests that any growth advantage conferred by multiple copies of the neomycin-gene-carrying viral genome might be offset by the detrimental effects of viral gene expression. This situation contrasts with other gammaherpesviruses, which are able to growth transform cells. PMID- 10859388 TI - Differential recognition of ORF2 protein from type 1 and type 2 porcine circoviruses and identification of immunorelevant epitopes. AB - Two types of porcine circovirus (PCV) have been isolated and are referred to as PCV1 and PCV2. PCV1 represents an apathogenic virus, whereas PCV2 is associated with post-weaning multisystemic wasting syndrome. The two PCVs are related, since they display about 70% identity based on nucleotide sequences. In order to discriminate between common and type-specific antigens, an immunocytological approach was used following transfections with cloned circovirus DNAs, as well as recombinant proteins expressed by either baculovirus or plasmid vectors. The ORF1 encoded proteins in the two viruses were shown to be antigenically related, whereas the ORF2 proteins were recognized differentially by polyclonal anti-PCV2 antibodies. Furthermore, PEPSCAN analysis performed on overlapping fragments of the genes encoding part of ORF1 and the entire ORF2 and ORF3 led to the identification of five dominant immunoreactive areas, one located on ORF1 and four on ORF2. However, only some ORF2 peptides proved to be immunorelevant epitopes for virus type discrimination. The potential use of ORF2-derived antigens as diagnostic tools is demonstrated. PMID- 10859389 TI - Inverse relationship between the expression of the human papillomavirus type 16 transcription factor E2 and virus DNA copy number during the progression of cervical intraepithelial neoplasia. AB - The human papillomavirus type 16 (HPV-16) status of 43 cervical biopsies, which had been characterized histologically as normal, various grades of cervical intraepithelial neoplasia (CIN) and invasive squamous cell carcinoma, was examined by using (i) a novel antibody against the HPV-16 E2 protein, (ii) sensitive HPV-16 DNA in situ hybridization and (iii) microdissection/PCR for the E2 ORF. The data indicate that E2 protein expression is highest in koilocytes in lower-grade CIN (I), but decreases with increasing grade, whereas the detection of HPV DNA is delayed until CIN I/II, rising to the highest levels in carcinoma cells. Co-localization of E2 with HPV-16 DNA-positive cells was most commonly observed in koilocytes in CIN II lesions. PCR analyses of microdissected epithelium from the same or serial sections indicated that E2 ORFs were retained in an intact form in a number of higher-grade CIN lesions and invasive carcinomas. PMID- 10859390 TI - The complete nucleotide sequence of fowl adenovirus type 8. AB - The fowl adenovirus type 8 (FAdV-8) genome was sequenced and found to be 45063 nucleotides in length, the longest adenovirus (AdV) genome for which the complete nucleotide sequence has been determined so far. No regions homologous to early regions 1, 3 and 4 (E1, E3 and E4) of mastadenoviruses were recognized. Gene homologues for early region 2 (E2) proteins, intermediate protein IVa2 and late proteins were found by their similarities to protein sequences from other AdVs. However, sequences homologous to intermediate protein IX and late protein V could not be identified. Sequences for virus-associated RNA could also not be recognized. Two regions of repeated sequences were found on the FAdV-8 genome. The shorter repeat region contained five identical and contiguous direct repeats that were each 33 bp long, while the longer repeat region was made of 13 identical and contiguous, 135 bp long repeated subunits. PMID- 10859391 TI - Multiple infection, recombination and genome relationships among begomovirus isolates found in cotton and other plants in Pakistan. AB - Begomoviruses occur in many plant species in Pakistan and are associated with an epidemic of cotton leaf curl disease that has developed since 1985. PCR analysis with primer pairs specific for each of four already sequenced types of DNA-A of cotton leaf curl virus (CLCuV-PK types a, 26, 72b and 804a), or for okra yellow vein mosaic virus (OYVMV), indicated that many individual naturally infected plants of cotton and other malvaceous species contained two or three begomovirus sequences. Similarly, sequence differences among overlapping fragments of begomovirus DNA-A, amplified from individual naturally infected plants, indicated much multiple infection in malvaceous and non-malvaceous species. Some cotton plants contained DNA-A sequences typical of begomoviruses from non-malvaceous species, and some non-malvaceous plants contained sequences typical of CLCuV-PK. Some DNA-A sequences were chimaeric; they each included elements typical of different types of CLCuV-PK, or of different malvaceous and/or non-malvaceous begomoviruses. Often an apparent recombination site occurred at the origin of replication. No complete CLCuV-PK DNA-A sequence was found in malvaceous or non malvaceous species collected in Pakistan outside the area of the cotton leaf curl epidemic but chimaeric sequences, including a part that was typical of CLCuV-PK DNA-A, did occur there. We suggest that recombination among such pre-existing sequences was crucial for the emergence of CLCuV-PK. Recombination, following multiple infection, could also explain the network of relationships among many of the begomoviruses found in the Indian subcontinent, and their evolutionary divergence, as a group, from begomoviruses causing similar diseases in other geographical regions. PMID- 10859392 TI - Limitations on the use of fused green fluorescent protein to investigate structure-function relationships for the cauliflower mosaic virus movement protein. AB - To investigate the process of tubule formation for the cauliflower mosaic virus movement protein (CaMV MP), the green fluorescent protein (GFP) was fused to the MP to provide a vital marker for MP location after expression in insect cells. In contrast to the long tubular structures seen previously following baculovirus based expression of the wild-type MP, the fusion protein produced only aggregates of fluorescing material in the cytoplasm. However, by co-expressing wild-type MP and GFP-MP, or by engineering their co-accumulation by introducing a foot-and mouth disease virus 2A cleavage sequence between GFP and MP, long GFP-fluorescing tubules were formed. The experiments suggest that the presence of GFP at the N or C terminus of the tubule-forming domain of the CaMV MP places steric constraints upon the aggregation of the MP into a tubule but that this can be overcome by providing wild-type protein for inclusion in the aggregate. PMID- 10859393 TI - Potato leafroll virus protein P1 contains a serine proteinase domain. AB - The multi-domain potato leafroll virus replicase protein P1 was expressed in insect cells from the polyhedrin promoter of Autographa californica nucleopolyhedrovirus. Using antisera raised against P1, it was shown that P1 was cleaved near the VPg in insect cells in a manner similar to that in plant cells, to produce a approximately 27 kDa C-terminal fragment. Furthermore, it was shown that the proposed serine proteinase-like domain within P1 is responsible for this processing and that this can occur in a trans (intermolecular) reaction. Four conserved residues within the serine proteinase domain that are essential for catalysis have been identified, consistent with the proposal that this domain comprises a serine proteinase. PMID- 10859394 TI - Sugarcane yellow leaf virus: a novel member of the Luteoviridae that probably arose by inter-species recombination. AB - The 5895 nucleotide long single-stranded RNA genome of Sugarcane yellow leaf virus Florida isolate (SCYLV-F) includes six major ORFs. All but the first of these are homologous to genes of known function encoded by viruses of the three newly defined genera in the LUTEOVIRIDAE: ('luteovirids'), i.e. poleroviruses, luccccteoviruses and the enamoviruses. SCYLV-F ORFs 1 and 2 are most closely related to their polerovirus counterparts, whereas SCYLV-F ORFs 3 and 4 are most closely related to counterparts in luteovirus genomes, and SCYLV-F ORF5 is most closely related to the read-through protein gene of the only known enamovirus. These differences in affinity result from inter-species recombination. Two recombination sites in the genome of SCYLV-F map to the same genomic locations as previously described recombinations involving other luteovirids. A fourth type of luteovirid, Soybean dwarf virus, has already been described. Our analyses indicate that SCYLV-F represents a distinct fifth type. PMID- 10859395 TI - Expression of a Tranosema rostrale polydnavirus gene in the spruce budworm, Choristoneura fumiferana. AB - The endoparasitic wasp Tranosema rostrale (Ichneumonidae) transmits a polydnavirus (PDV) to its host, Choristoneura fumiferana, during oviposition. Unlike most other PDVs examined, the virus of T. rostrale (TrPDV) does not appear to play an important role in suppressing the host cellular immune response. However, it inhibits host metamorphosis. In the present study, TrPDV gene expression was examined in parasitized and virus-injected last-instar caterpillars. Northern analysis with viral DNA as a probe revealed only one detectable mRNA, of about 650 bp. The corresponding cDNA, termed TrV1, was cloned and sequenced and found to encode a protein of 103 amino acids which, following cleavage of the putative signal peptide, has a predicted molecular mass of 9.3 kDa. This protein displays limited similarity to the VHv1.4 cysteine-rich protein from the PDV of Campoletis sonorensis, mostly within the signal peptide region. By using a TrV1-specific probe, the TrV1 gene was localized to segment G of the TrPDV genome. The cuticle and fat body were identified as the principal sites of TrV1 transcription, with little transcription observed in haemocytes and midgut. Western analysis of proteins extracted from selected tissues of parasitized insects suggested that the TrV1 protein is secreted in the haemolymph. As observed for other PDVs, injection of TrPDV did not suppress transcription of the gene that encodes juvenile hormone esterase, the activity of which is inhibited by the virus. We speculate that the TrV1 protein may play a role in the inhibition of C. fumiferana metamorphosis. PMID- 10859396 TI - Filamentous actin is required for lepidopteran nucleopolyhedrovirus progeny production. AB - Autographa californica M nucleopolyhedrovirus (AcMNPV) is the prototypical member of the NUCLEOPOLYHEDROSIS: genus of the BACULOVIRIDAE:, a family of large, double stranded DNA viruses that are highly diverse. Nucleocapsid morphogenesis of AcMNPV and others in the NUCLEOPOLYHEDROVIRUS: genus takes place within the nuclei of infected host cells. Previously, we showed that filamentous actin (F actin) is essential for this process to occur in AcMNPV-infected cells, an unprecedented finding for a DNA virus that replicates within the nucleus. Because of the fundamental importance of this requirement to our understanding of virus host interactions, and because of the diversity of viruses included within the Nucleopolyhedrovirus genus, we were compelled to determine whether the replication of other nucleopolyhedroviruses was also F-actin dependent. We report here that progeny virus production of six other lepidopteran nucleopolyhedroviruses, representing both phylogenetic groups I and II within the genus, is also F-actin dependent. The six viruses studied (Spodoptera frugiperda MNPV, Bombyx mori NPV, Orgyia pseudotsugata MNPV, Lymantria dispar MNPV, Anticarsia gemmatalis MNPV and Helicoverpa zea SNPV) were unable to produce progeny in the presence of either cytochalasin D or latrunculin A, two actin binding agents that interfere with F-actin-dependent processes but differ in their modes of action. F-actin-dependent progeny morphogenesis, therefore, appears to be a characteristic common among viruses in this genus that have lepidopteran hosts. PMID- 10859397 TI - Dangerous liaisons? PMID- 10859398 TI - Home mechanical ventilation for end-stage chronic obstructive pulmonary disease (for all)? PMID- 10859399 TI - Welcome to Toronto. Welcome to the CIHR. PMID- 10859400 TI - Tuberculosis in aboriginal Canadians. AB - Endemic tuberculosis (TB) was almost certainly present in Canadian aboriginal people (aboriginal Canadians denotes status Indians, Inuit, nonstatus Indians and metis as reported by Statistics Canada) before the Old World traders arrived. However, the social changes that resulted from contact with these traders created the conditions that converted endemic TB into epidemic TB. The incidence of TB varied inversely with the time interval from this cultural collision, which began on the east coast in the 16th century and ended in the Northern Territories in the 20th century. This relatively recent epidemic explains why the disease is more frequent in aboriginal children than in Canadian-born nonaboriginal people. Treatment plans must account for the socioeconomic conditions and cultural characteristics of the aboriginal people, especially healing models and language. Prevention includes bacillus Calmette-Guerin vaccination and chemoprophylaxis, and must account for community conditions, such as rates of suicide, which have exceeded the rate of TB. The control of TB requires a centralized program with specifically directed funding. It must include a program that works in partnership with aboriginal communities. PMID- 10859402 TI - Predictors for multiple hospital admissions in children with asthma. AB - BACKGROUND: Asthma is one of the most frequent causes of preventable hospital admissions among children. OBJECTIVES: To identify preventable risk factors for future hospital admissions. Some of the environmental and patient characteristics of children who were hospitalized more than once with an acute asthma attack were investigated. DESIGN, SETTING AND PATIENTS: An analysis was performed of 140 children with asthma, aged three to 15 years, admitted to the Department of Pediatrics at Dicle University Hospital, Diyarbakir, Turkey, over three years, followed for a maximum of 48 months. Associations between hospital admissions and probable predictors (clinical findings, laboratory studies and a detailed case history) were analyzed. RESULTS: The asthma admissions were higher in boys than in girls (male to female ratio 1.86). Of 140 children, 30 (21.4%) had multiple admissions, defined as a mean of more than one admission per year during the follow-up period. The following factors were associated significantly with the frequency of hospital admission for asthma attacks when analyzed using a Chi2 test: indoor cigarette smoking (odds ratio [OR] 2.55), maternal smoking (OR 4.05), symptoms of dermal atopy (OR 2.96), symptoms of allergic conjunctivitis (OR 2.68), age less than five years (OR 5.12) and use of inhaled corticosteroids during the follow-up (OR 0.37). With multi-variate logistic regression analysis, among other factors, only maternal smoking (r=0.29, P=0.017) and age less than five years (r=-0.32, P=0.012) were significant effective factors for the multiple hospital admissions. No significant association with the frequency of hospital admission was found for sex, serum total immunoglobulin E, history of frequent upper respiratory tract infections or number of family members. CONCLUSIONS: Prevention of indoor smoking, especially maternal smoking, may be effective in decreasing hospital admissions in children with asthma. Children less than five years of age are more likely to be hospitalized than are children five years of age or greater. PMID- 10859401 TI - Tuberculosis among aboriginal and nonaboriginal persons in British Columbia. AB - OBJECTIVE: To compare cases of tuberculosis (TB) diagnosed among aboriginal persons with a random sample of nonaboriginal persons diagnosed with TB, and evaluate the trends in rates of disease between both groups during the same period. DESIGN: A case-control study. SETTING: A provincial TB control program. PATIENTS AND METHODS: All patients with TB diagnosed among aboriginal persons in British Columbia between 1992 and 1996 were compared with control patients diagnosed during the same period. For each patient a control patient was identified. INTERVENTION: The demographic details, type of disease, bacteriology, risk factors for TB, therapy received as well as mode of administration were documented. The number of contacts identified for each patient as well as the number of patients completing chemoprophylaxis were identified. The rates of disease during the same period were also documented. RESULTS: During the study, 202 patients with TB were diagnosed among aboriginal persons and 201 controls were chosen. Apart from age at diagnosis (35.1+/-20 years versus 45.7+/-19.7), differences in the prevalence of lymphadenopathy (5.9% versus 16.4%, P=0.0008) and pleural disease (21.3% versus 16.4%, P=0.00008), there were no differences in presentation between aboriginal and nonaboriginal people. Aboriginal people were more likely to have a history of contact with a patient with TB (53% versus 17.9%, P<0.05), to have received directly observed therapy (55% versus 33.8%, P=0.00002) and to have contacts who were purified protein derivative (PPD) positive (4+/-9 versus 2+/-3, P=0.002). These contacts were more likely to start isoniazid (2+/-3 versus 1+/-1, P=0.002). Overall, there was a significant decline in rates of TB among aboriginal persons compared with the general population, but there was a small increase in rates among all subjects in the final year of the study. CONCLUSIONS: In the present study, significant variations in rates of TB among different population groups in British Columbia were found. During the study period, there was a greater decline in the rates of TB among aboriginal persons. A greater use of directly observed therapy and greater use of chemoprophylaxis occurred among aboriginal persons, which may have contributed to this decline, or alternatively, it simply reflects the natural evolution of the TB epidemic. PMID- 10859403 TI - Gastroesophageal reflux and asthma: can the paradox be explained? AB - BACKGROUND AND OBJECTIVE: The reported effects of asthma on gastroesophageal reflux (GER), effects of GER on asthma and the effects of antireflux therapy on asthma are conflicting. The purpose of this paper is to review the evidence for a relationship between the two conditions. DESIGN: A search of the MEDLINE 1966 to 1999 database, combining the terms GER and asthma, was used to identify studies of the effects of acid perfusion of the esophagus, the physiological equivalent of GER and the effects of both medical and surgical antireflux therapy on asthma. Bibliographies of the identified papers were also reviewed. MAIN RESULTS: The collected evidence suggests that GER causes asthma symptoms but has minimal effects on pulmonary function. Both medical and surgical antireflux therapy can improve asthma symptoms and asthma medication requirements without improving pulmonary function. The paradox of GER causing symptoms without affecting pulmonary function may be because of the retrosternal discomfort that accompanies GER increases minute ventilation and respiratory sensation. CONCLUSIONS: Despite an extensive body of literature, many questions remain about the relationship between GER and asthma. A review of the data suggests a strong association between the two conditions, and that GER worsens asthma symptoms without affecting pulmonary function. Asymptomatic GER does not worsen asthma. Antireflux therapy may have a role in asthma patients with symptomatic GER, possibly being most beneficial for those with reflux-associated respiratory symptoms. Unfortunately, many studies contain flaws such as a lack of controls and small sample sizes. Further properly designed controlled trials, including ones that measure the effects of GER and antireflux therapy on quality of life, are needed to understand better the role of GER in asthma. PMID- 10859404 TI - Allergen-induced increase in nonallergic airway responsiveness: a citation classic revisited. AB - BACKGROUND: The present paper revisits the 1977 paper by DW Cockcroft, RE Ruffin, the late J Dolovich and FE Hargreave entitled "Allergen-induced increase in nonallergic bronchial reactivity" (Clin Allergy 1977;7:503-13) that became a citation classic. Although clinical types of asthma were recognized at the time, there was a poor understanding regarding the role of allergic reactions in causing increases in airway hyperresponsiveness. The objective was to study formally Dr Altounyan's observation that patients with asthma showed increases in airway responsiveness at the times of natural allergen exposure during pollen season. Thirteen atopic patients with asthma were studied over two days, following inhalation of diluent (control) and following doubling amounts of an allergen solution at 10-min intervals until forced expiration volume in 1 s fell by 20%. Methacholine and histamine challenges were performed before, at 8 h, at 32 h and seven days following the inhalations. A significant reduction (reduction of at least one doubling concentration) in the provocative concentration that causes a 20% fall in forced expiration volume in 1 s occurred in seven of 13 patients, and more often in subjects with a late bronchoconstrictor response to allergen challenge. IMPORTANCE: The study showed that large changes in airway responsiveness could occur in patients with asthma and suggested that allergens could cause, rather than trigger, asthma. The study also led to the concept of asthma inducers and inciters - inducers causing airway inflammation and inciters provoking bronchospasm. The results led to a series of observations that have now implicated immunoglobulin E-mediated airway inflammation as perhaps the most important cause of airway hyperresponsiveness in asthma. PMID- 10859405 TI - An infected mediastinal cyst. AB - The authors describe a 43-year-old patient who had a mediastinal mass that became infected after a transbronchial needle aspirate biopsy. A paraspinal, extrapleural window with a saline-lidocaine mixture was created that allowed the placement of a percutaneous drainage catheter into the infected lesion. This procedure resulted in an excellent clinical outcome, and obviated the need for a thoracotomy and more invasive surgical management. PMID- 10859406 TI - Amiodarone pulmonary, neuromuscular and ophthalmological toxicity. AB - Amiodarone is an iodinated benzofuran derivative class III antiarrhythmic that is highly effective in suppressing ventricular and supraventricular arrhythmias. It is also associated with an imposing side effect profile, which often limits its use. Numerous adverse effects have been documented including skin discolouration, photosensitivity, hepatitis, thyroid dysfunction, corneal deposits, pulmonary fibrosis, bone marrow suppression and drug interactions. These side effects are thought to be correlated with the total cumulative dose of amiodarone, but idiopathic reactions have been reported. The majority of adverse reactions resolve with discontinuation of the drug; however, rapid progression may occur, which may be fatal. The present report documents a patient who had a combination of serious amiodarone toxicities that, once recognized, were treated and eventually resulted in a good outcome. PMID- 10859407 TI - Mismatch negativity: clinical and other applications. AB - The perspectives of application of the mismatch negativity (MMN), generated by the brain's automatic response to change in auditory stimulation, are discussed. In light of the fact that the MMN (and its magnetic equivalent MMNm) currently provides the only objective measure of the accuracy of the central auditory function, these perspectives appear very promising. The MMN can be measured in the absence of attention and task requirements, which makes it particularly suitable for testing different clinical populations and infants. Furthermore, the MMN enables one to evaluate the accuracy of auditory discrimination separately for any acoustic feature, such as frequency, intensity and duration, and for learned categories, such as the phonemes of a particular language. In addition, by measuring the decay of the MMN amplitude as a function of the interstimulus interval, it is possible to estimate the duration of sensory (echoic) memory. PMID- 10859408 TI - Mismatch negativity: different water in the same river. AB - The mismatch negativity (MMN) is a frontal negative deflection in the human event related potential that typically occurs when a repeating auditory stimulus changes in some manner. The MMN can be elicited by many kinds of stimulus change, varying from simple changes in a single stimulus feature to abstract changes in the relationship between stimuli. The main intracerebral sources for the MMN are located in the auditory cortices of the temporal lobe. Since it occurs whether or not stimuli are being attended, the MMN represents an automatic cerebral process for detecting change. The MMN is clinically helpful in terms of demonstrating disordered sensory processing or disordered memory in groups of patients. Improvements in the techniques for measuring the MMN and in the paradigms for eliciting it will be needed before the MMN can become clinically useful as an objective measurement of such disorders in individual patients. PMID- 10859409 TI - Speech sound representation in the brain. AB - Biologic processes underlying speech sound perception and learning have been addressed using the mismatch negativity (MMN) evoked response. First is a consideration of how the acoustic properties of the signal affect the neural mechanisms and brain regions engaged. Because the MMN differs depending on the acoustic characteristics of the stimuli used to elicit the response, it has been used to probe mechanisms underlying the neural representation of stimuli along the auditory pathway. Second is a consideration of neurophysiologic correlates of speech sound perception and learning. Detailed is a 'behavioral-neurophysiologic, acoustic-phonetic approach', used to link perception with underlying physiologic processes in humans. The focus here is on children and what has been learned about normal maturation of speech sound perception and its disruption in certain children with learning disorders. The last topic is a consideration of central nervous system changes with perceptual learning. This includes long-term experience with one's native language and short-term auditory training in the laboratory. Limitations and future challenges are discussed. PMID- 10859410 TI - Involuntary attention and distractibility as evaluated with event-related brain potentials. AB - This article reviews recent event-related brain potential (ERP) studies of involuntary attention and distractibility in response to novelty and change in the acoustic environment. These studies show that the mismatch negativity, N(1) and P(3a) ERP components elicited by deviant or novel sounds in an unattended sequence of repetitive stimuli index different processes along the course to involuntary attention switch to distracting stimuli. These studies used new auditory-auditory and auditory-visual distraction paradigms, which enable one to assess objectively abnormal distractibility in several clinical patient groups, such as those suffering from closed-head injuries or chronic alcoholism. PMID- 10859411 TI - Maturation of the mismatch negativity: effects of profound deafness and cochlear implant use. AB - The use of cochlear implants to restore auditory sensation in deaf children is increasing, with a trend toward earlier implantation. However, little is known about how auditory deprivation and subsequent cochlear implant use affect the maturing human central auditory system. Our previous studies have demonstrated that the obligatory auditory evoked potentials (AEPs) of implanted children are very different from those of normal-hearing children. Unlike the obligatory potentials, which primarily reflect neural responses to stimulus onset, the mismatch negativity (MMN) provides a neurophysiological measure of auditory short term memory and discrimination processes. The purpose of this investigation is to review our studies of the effects of auditory deprivation due to profound deafness and cochlear implant use on the maturation of the MMN in children, placed in the context of overall age-related changes in the AEPs. The development and application of a statistical technique to assess the MMN in individuals is also reviewed. Results show that although the morphology of the obligatory AEPs is substantially altered by the absence of a normal N(1) peak, the MMN is robustly present in a group of implanted children who have good spoken language perception through their device. Differences exist in the scalp distribution of the MMN between implanted and normal-hearing children. Specifically, the MMN appears to be more symmetrical in amplitude over both hemispheres, whereas it is initially much larger over the contralateral hemisphere in normal-hearing children. These findings suggest that, compared to N(1), the MMN is a better measure of basic auditory processes necessary for the development of spoken language perception skills in profoundly deaf children and adults who use a cochlear implant. PMID- 10859412 TI - Coma outcome prediction using event-related potentials: P(3) and mismatch negativity. AB - The prediction of the outcome from coma is of considerable importance to the patients, their relatives and attendant medical staff, and yet current clinical methods lack sensitivity and specificity. Objective investigations can enhance the accuracy of such predictions and are an important adjuvant when reaching decisions to continue or terminate life support. Of the neurophysiological methods available, electroencephalography and short-latency somatosensory evoked potentials have proved the most useful in the clinical setting. These tests are good predictors of an adverse outcome; however, they tell us only about the ongoing cerebral activity and integrity of the primary somatosensory pathways, respectively. The presence of long-latency event-related potentials has been shown to be a useful predictor of a favourable neurological outcome, and thus their use complements other neurophysiological techniques. Their potential application in clinical practice is reviewed. PMID- 10859413 TI - Mismatch negativity and N100 in comatose patients. AB - Mismatch negativity (MMN) and N100 auditory evoked potential were recorded in 52 healthy subjects and in 128 severely comatose patients. The MMN was present in 33/128 patients and N100 in 84/128. A ratio of 30/33 patients with MMN and 70/84 with N100 regained consciousness in a mean time of 6.3 +/- 4 days after the recording session. Thus, in terms of predicting return to consciousness, the MMN was more specific (90.9%) than the N100 (57.6%), but its sensitivity was lower (31.6% for MMN and 73.7% for N100, respectively). The amplitudes of MMN and N100 in comatose patients were smaller than those of healthy subjects. It is concluded that MMN and N100 can be very useful in predicting whether or not a comatose patient will regain consciousness. PMID- 10859414 TI - Mismatch negativity and N100 monitoring: potential clinical value and methodological advances. AB - Continuous long-term recording of brainstem (BAEPs), middle-latency (MLAEPs) and long-latency auditory evoked potentials, including the mismatch negativity (MMN), brings additional information on the immediate functional state and the outcome of patients in coma or recovering after surgery, in relation with clinical observations and therapeutics. A recently designed monitoring system is introduced, aimed at the continuous recording of late auditory potentials (N100 and MMN) as well as BAEPs and MLAEPs. Specific methodological aspects are emphasized. Long-term monitoring data from one patient recorded in the recovery room after surgery are displayed, allowing an illustration of the techniques used and of the problems raised. PMID- 10859415 TI - Intracortical mechanisms of mismatch negativity dysfunction in schizophrenia. AB - Event-related potentials provide an objective index of neurocognitive dysfunction in schizophrenia. Schizophrenia subjects show a decreased mismatch negativity (MMN) amplitude relative to age- and sex-matched controls, along with a characteristic pattern of MMN dysfunction across conditions. Deficits in MMN generation are accompanied by (1) impaired precision of auditory sensory memory performance and (2) an interstimulus-interval-dependent deficit in auditory N(1) generation. Similar deficits are observed following systemic or local infusion of N-methyl-D-aspartate (NMDA) antagonists, supporting glutamatergic and phencyclidine/NMDA models of the disorder. Deficits in MMN generation may also be seen following focal cortical damage, especially to the dorsolateral prefrontal cortex. MMN thus provides a useful tool for investigating mechanisms underlying brain dysfunction in schizophrenia. PMID- 10859416 TI - Mismatch negativity in aging and in Alzheimer's and Parkinson's diseases. AB - Mismatch negativity (MMN) is an auditory event-related potential (ERP) that reflects automatic stimulus discrimination in the human auditory system. By varying the interstimulus intervals (ISIs), the MMN can be used as an index of auditory sensory memory. This paper focuses on MMN findings in aging and in Alzheimer's (AD) and Parkinson's diseases (PD). The accumulated data suggest that MMN to duration deviance, unlike MMN to frequency deviance, is reduced in amplitude in aging at short ISIs. The attenuated MMN to frequency deviance observed at long ISIs in elderly subjects seems to be caused by age-related memory trace decay. Existing results suggest that automatic discrimination for the frequency change is not affected in the early phase of AD, whereas the memory trace seems to decay faster in AD patients. The present findings on PD are not as conclusive, although they tentatively suggest deteriorated automatic change detection. The MMN appears to offer an objective tool for studying auditory processing and memory trace decay in different neurological disorders. PMID- 10859417 TI - Unilateral neglect after right-hemisphere damage: contributions from event related potentials. AB - Unilateral neglect is a frequent sequel of right-hemisphere damage. Patients suffering from neglect may fail to detect, orient to, acknowledge or respond to stimuli on their contralesional side, even in the absence of primary sensory or motor loss. Despite the major clinical significance of the phenomenon and its potential implications for our understanding of human cognition, the underlying cognitive deficits are not well understood. We review the relatively few event related potential studies that attempted to assess the different parts of the cognitive system in neglect patients. We suggest that theories of neglect, based primarily on performance data, may be refined by incorporating these results, and that this line of research may provide information that is not available using traditional performance measures. PMID- 10859418 TI - Towards optimal recording and analysis of the mismatch negativity. AB - In this paper, the conceptual and practical issues related to the measurement of mismatch negativity (MMN) are discussed from the viewpoint of cost-efficiency. First, various criteria for efficiency or optimality of measurements are described, including reliability and signal-to-noise ratio. Then a critical look is taken at some currently used concepts and data analysis methods. Practical guidelines for the measurement and analysis of MMN are given, complementing the earlier reviews on the subject. Finally, reliability studies on MMN are critically reviewed. PMID- 10859419 TI - Oxidative modification of low-density lipoprotein (LDL) in HD patients: role in electronegative LDL formation. AB - High cardiovascular mortality in patients on hemodialysis (HD) is largely attributed to oxidative stress and altered lipoprotein profiles. Markedly increased levels of mildly modified LDL subfractions, such as dense LDL and electronegatively charged LDL (LDL(-)), are present in the blood of HD patients and may be markers of atherosclerosis risk. LDL(-), characterized by modified protein content and elevated levels of lipid peroxidation products, is representative of multiple oxidative processes acting on plasma lipoproteins that prevail during HD. In this review, we discussed known mechanisms leading to that may account for oxidative protein modification and/or LDL(-) formation in the context of specific conditions associated with HD. PMID- 10859420 TI - Homocysteine as a cardiovascular risk factor. AB - Hyperhomocysteinemia (HH), a known risk factor for vascular diseases, is a frequent condition in hemodialysis (HD) patients. HH induces an oxidant stress to the vascular endothelium, causing a failure of vasodilation and an impairment of the antithrombotic properties. Vitamins B(6), B(12) and folic acid are important cofactors for the enzymes in the catabolism of homocysteine (Hcy). Failure of Hcy catabolism forces the cell to export Hcy into the plasma. The kidney is an important metabolic site for removal (up to 70%) of plasma Hcy (P-Hcy). HD lowers the P-Hcy concentration by 29 and 41% with cellulosic and noncellulosic membranes, respectively, yet values return to normal in only a few patients. Clearly, we must decrease the dangerous high levels of Hcy in different ways. Vitamin Supplementation: Vitamins B(6), B(12) and folic acid decreased the basal level of Hcy by about 40%, starting from the sixth month. Membranes: Some membranes performed better than the others. TECHNIQUES: On the chronic basis, in our 1-year experience, paired filtration dialyis led to the best results, when compared to bicarbonate dialysis and acetate-free biofiltration. Finally, as in HD patients no one type of treatment can normalize the P-Hcy concentration, we should try other, different strategies such as absorption, the use of liposomes and new types of supplementation. PMID- 10859421 TI - C-reactive protein as a marker of chronic inflammation in uremic patients. AB - Cardiovascular complications caused by an accelerated atherosclerotic disease represent the largest single cause of mortality in chronic renal failure patients. The rapidly developing atherosclerosis of the uremic syndrome appears to be caused by a synergism of different mechanisms, such as malnutrition, oxidative stress and genetic factors. Recent studies provide evidence that chronic inflammation plays an important role in the pathogenesis of cardiovascular diseases. Elevated serum levels of plasma C-reactive protein (CRP) are associated with an increased risk of experiencing myocardial infarction and sudden cardiac death in apparently healthy subjects. Several recently published papers have confirmed this strong association between CRP and the extent and severity of the atherosclerotic processes. In patients affected by predialytic renal failure, increased levels of CRP and interleukin (IL)-6 were recorded in 25% of our population; CRP and IL-6 were inversely related with renal function. These data suggest the activation - even in the predialytic phase of renal failure - of mechanisms known to contribute to the enhanced cardiovascular morbidity and mortality of the uremic syndrome. In recent years we have investigated the hypothesis that the chronic inflammatory state of the uremic patient could at least in part be due to the dialytic technique. We provide evidence suggesting that the increase of CRP in stable dialytic patients may be due to the stimulation of monocyte/macrophage by backfiltration of dialysate contaminants. PMID- 10859422 TI - Why hemodialysis patients are in a prooxidant state? What could be done to correct the pro/antioxidant imbalance. AB - Oxidative stress which results from an imbalance between reactive oxygen species production and antioxidant defense mechanisms is now well recognized in hemodialysis (HD) patients and could be involved in dialysis-related pathologies such as accelerated atherosclerosis, amyloidosis and anemia. In order to evaluate the rationale for preventive intervention against oxidative damage during HD, we review the factors that are implied and may be responsible for the imbalance between pro- and antioxidative mechanisms. The inflammatory state mainly due to hemobioincompatibility of the dialysis system plays a critical role in the production of free oxygen radical species contributing by this way to worsen the prooxidant status of uremic patients. Two factors largely contribute to the stimulation of the NADPH oxidase: hemoreactivity of the membrane and trace amounts of endotoxins. The antioxidant system is severely impaired in uremic patients and gradually altered with the degree of renal failure. HD could further impair this antioxidant system mainly by losses of (a) hydrophilic unbound small molecular-weight substances such as vitamin C, (b) trace elements and (c) enzyme regulatory compounds. Two main axes may be proposed in order to prevent and/or to decrease oxidative stress in HD patients. One consists in improving the hemocompatibility of the dialysis system mainly by using a dialyzer with low hemoreactivity and ultrapure, sterile, nonpyrogenic dialysate. The other consists in supplementing the deficiency patients with antioxidants. This could be achieved by oral or perdialytic supplementation. Vitamin E could be bound on dialyzer membrane. Alternatively, hemolipodialysis consists in loading HD patients with vitamin C or E via an ancillary circuit made of vitamin E-rich liposomes. The presence of liposomes could also facilitate the removal of hydrophobic prooxidative substances. PMID- 10859423 TI - Microbiologic purity of dialysate: rationale and technical aspects. AB - Dialysate purity has become a major concern in hemodialysis since it has been shown that microbial-derived products were stimulating the production and the release of proinflammatory cytokines in hemodialysis patients. This chronic microinflammatory state induced by hemodialysis has been putatively implicated in the development of dialysis-related pathology. In order to prevent risk related to these offenders and to reduce patient/dialysis interaction, it appears highly desirable to use ultrapure dialysis fluid aiming at sterility and apyrogenicity on a regular basis. Ultrapure dialysate results from a complex chain of production where purity grade relies on the weaker link of this chain. Technical aspects and pitfalls in the production of ultrapure dialysate are summarized in this paper. Production of ultrapure dialysate may be achieved on a routine basis, provided adequate components are used, and hygienic handling is regularly ensured. It includes the use of ultrapure water, clean and or sterile electrolytic concentrates (liquid or powder), implementation of ultrafilters on hemodialysis machines, microbiologic monitoring and hygienic handling of the chain with frequent disinfection. Safety and reliability of ultrapure dialysate production relies on a continuous quality assurance process, where results are coupled to corrective action in a feedback loop process. PMID- 10859424 TI - Chronic inflammation in hemodialysis: the role of contaminated dialysate. AB - Routine sodium bicarbonate-buffered dialysate is contaminated with predominantly gram-negative micro-organisms. These bacteria release pyrogenic substances such as endotoxins, peptidoglycans, exotoxins and fragments thereof. Pyrogens derived from contaminated dialysate either alone or in costimulation with activated complement components are the most important activators of circulating mononuclear cells in patients on chronic intermittent hemodialysis. Activated mononuclear cells release proinflammatory cytokines which are key mediators in acute and chronic inflammatory diseases associated with long-term hemodialysis therapy. Recent experimental and clinical data suggest that the use of pyrogen free dialysate prevents activation of mononuclear cells and improves the state of chronic inflammation, as indicated by decreased plasma levels of C-reactive protein in chronic hemodialysis patients. Future clinical studies have to prove whether the use of pyrogen-free dialysate in combination with biocompatible dialyzer membranes and tubings reduces the incidence and severity of chronic inflammatory diseases (beta(2)-microglobulin amyloidosis, muscle protein wasting, atherosclerosis) in long-term hemodialysis patients. PMID- 10859425 TI - Biofilms invade nephrology: effects in hemodialysis. AB - Bacteria attach to surfaces and aggregate in a biopolymer matrix to form biofilm. Studies on biofilm have shown its presence in many prosthetic devices used in nephrology as well as in fluid pathways of hemodialysis plants and monitors. Once present, this community of bacteria increases resistance to biocide due to slime production and, as a result, chemical products for dialysis monitor disinfection and descaling procedures do not result in an effective treatment. Ultrapure dialysate is a goal in modern hemodialysis, and ultrafiltration is used to obtain sterile and apyrogen fluids. Microbial colonisation of ultrafilters may occur if, due to inadequate disinfection protocols, membrane is exposed to persistent bacterial contamination, and biofilm is allowed to form and to grow. As more and more data link final dialysate microbial contamination to clinical effects of bioincompatibility from chronic inflammation in dialysis patients, attention has to be focused on possibilities of biofilm avoidance. PMID- 10859426 TI - Hemodiafiltration with on-line endogenous reinfusion. AB - Paired filtration dialysis is a modified form of hemodiafiltration with a double chamber hollow fiber. Convection is separated from diffusion, eliminating the potential risk of backfiltration (which can contain endotoxin or cytokine inducing substances). The regeneration of high volumes of plasma ultrafiltrate obtained in the first filter allows a large plasma volume to be treated, and at the same time enables the return of many beneficial substances such as hormones, small peptides and many vitamins. Ultrafiltrate is regenerated with a charcoal resin device and reinfused to the patient. Hemodiafiltration with on-line endogenous reinfusion is an easy and safe procedure. In addition, the method avoids risks associated with exogenous fluid infusion (endotoxin, pyrogens), allows exchange at no extra costs of large volumes of fluids and reduction in storage of fluid bags. Clinical advantages also include infusion of physiological fluid containing bicarbonate and calcium, good clinical tolerance and cardiovascular stability. PMID- 10859427 TI - Double-chamber on-line hemodiafiltration: a novel technique with intra-treatment monitoring of dialysate ultrafilter integrity. AB - On-line hemodiafiltration is a technique that relies on the re-injection of pyrogen-free substitution fluid obtained by cold filtration of dialysate. Therefore, safety of this treatment modality depends on the quality of dialysate and, mainly, on the integrity of the ultrafilter(s) employed. Double-chamber on line hemodiafiltration is a new technique where re-infusion takes place inside the dialyser by means of dialysate backfiltration. The peculiar geometry of the dialyser allows intra-treatment assessment of its fibre integrity. In this paper, we tested feasibility and safety of this new modality of on-line treatment. The extracorporeal blood and infusate pressure values resulted well inside the safety range. Blood urea clearances and beta(2) removal were consistent with the figures usually found in standard hemodiafiltration. Whole blood production of cytokines was similar when blood was exposed to saline or infusate, both values being comparable to the spontaneous whole blood cytokine release. The on-line dialyser fibre integrity check showed a great sensitivity even for minimal dialyser damage. We conclude that double-chamber on-line hemodiafiltration is a feasible and safe procedure. Our preliminary results encourage the undertaking of multicentre, prospective, randomised studies. PMID- 10859428 TI - In vitro activity of ceftazidime, cefepime and imipenem on 1,005 Pseudomonas aeruginosa clinical isolates either susceptible or resistant to beta-lactams. AB - BACKGROUND: Recently, new 'fourth-generation' cephalosporins, such as cefepime and cefpirome, were introduced into antibacterial chemotherapy. METHODS: In order to explore whether these new cephalosporins offer real advantages against Pseudomonas aeruginosa, we matched the in vitro activity of cefepime with that of ceftazidime and imipenem as reference compounds. RESULTS: Among the 1,005 clinical isolates tested, 86.6% were susceptible to ceftazidime, whereas 80.7 and 76.9% were susceptible to imipenem and cefepime, respectively. Furthermore, the activity of the three compounds against a significant number of clinical isolates of P. aeruginosa expressing different resistance mechanisms to beta-lactam antibiotics was investigated. Among these isolates, 62.5% were still susceptible to ceftazidime, and 52.1 and 38.7% were inhibited by imipenem and cefepime, respectively. CONCLUSION: Ceftazidime and imipenem retained their activity against the majority of clinical P. aeruginosa isolates collected in Italy. Cefepime did not offer competitive advantages in terms of in vitro activity. PMID- 10859429 TI - Comparative in vitro antifungal activity of amphotericin B lipid complex, amphotericin B and fluconazole. AB - Amphotericin B (AMB) is considered the gold standard in the treatment of serious systemic mycoses in spite of its nephrotoxicity and adverse effects. Association with lipids enables larger doses of AMB to be given with a longer t((1/2)) and C(max), without the toxic effects at lower concentrations. Liposome-encapsulated AMB shows a lower affinity for mammalian cells and improves V(d), thus decreasing toxicity. Amphotericin B lipid complex (ABLC) is an AMB formulation associated with a biodegradable phospholipid matrix (5% molar) from which the drug is released by cell phospholipases. ABLC is recommended for serious mycoses refractory to conventional antifungal therapy or when AMB is contraindicated. We compared the in vitro antifungal activity of ABLC, AMB and fluconazole (FLZ) against 328 strains of clinically significant opportunistic fungi using a microdilution method (NCCLS, M-27A). 64.9% of the yeasts were inhibited by MIC of ABLC /= WHO degree II reported in 1 out of 46 cycles only. CONCLUSION: Intensive i.v. AMB prophylaxis reduced invasive fungal infections and led to a reduction in fungal microabscesses in the liver or spleen, as well as pulmonary infiltrates, in patients treated for AL. The need for escalation to empirical i.v. AMB treatment was significantly reduced. Intensive AMB prophylaxis was feasible, with moderate adverse effects. PMID- 10859436 TI - Medical therapy for benign prostatic hyperplasia: a review of the literature. AB - OBJECTIVE: To review the existing evidence regarding the efficacy and safety of medical therapy for lower urinary tract symptoms (LUTS) indicative of benign prostatic hyperplasia (BPH). To assess randomised controlled trials investigating the six alpha-adrenergic receptor antagonists (alpha-blockers), prazosin, alfuzosin, indoramin, terazosin, doxazosin, and tamsulosin, that benefit patients by relaxing prostatic smooth muscle, and the anti-androgen, finasteride, that mediates its more long-term benefits by reducing prostate size. RESULTS: This review suggests that both classes of drug offer significant improvement in criteria used to evaluate symptomatic BPH and can be effective whilst being acceptably safe. Furthermore, the therapeutic efficacy of all contemporary alpha blockers appear similar, both in terms of symptom relief and urodynamic improvements. Randomised controlled trials have additionally demonstrated that finasteride therapy can provide improvement in terms of quality of life indices, prostate volume, and risks of progressing to acute urinary retention or prostatic surgery. While alpha-blockers have a rapid onset of action, likely to produce a therapeutic result within weeks, regardless of whether prostatic enlargement or bladder outlet obstruction is present, finasteride appears to be effective for more long-term therapy for up to 4 years, but only in alleviating symptoms when they are associated with a significantly large prostate. Neither finasteride nor the alpha(1a)-receptor-selective blocker, tamsulosin, are associated with the lowering of blood pressure and incidence of cardiovascular side effects that are apparent with other less selective alpha-blocker therapies such as dizziness and postural hypertension. They are, however, both associated with an increased risk of sexual dysfunction, albeit less than those associated with surgical intervention. Whereas tamsulosin is associated only with ejaculatory dysfunction, finasteride is additionally linked to decreased libido and impotence. PMID- 10859437 TI - Anatomy and physiology of female sexual function and dysfunction: classification, evaluation and treatment options. AB - Female sexual dysfunction is a significant age-related, progressive and highly prevalent problem that affects a substantial number of women in the United States. The female sexual response cycle is initiated by neurotransmitter mediated vascular and nonvascular smooth muscle relaxation resulting in increased pelvic blood flow, vaginal lubrication, and clitoral and labial engorgement. These mechanisms are mediated by a combination of neuromuscular and vasocongestive events. Physiological impairments that interfere with the normal female sexual response bring about complaints associated with diminished sexual arousal, libido, vaginal lubrication, genital sensation, and ability to achieve orgasm. Therapy aimed at restoring hormone levels as well as genital blood flow will be discussed. PMID- 10859438 TI - Initial results utilizing combination therapy for patients with a suboptimal response to either alprostadil or sildenafil monotherapy. AB - OBJECTIVE: Intraurethral alprostadil and oral sildenafil are useful in selected patients. However, there continues to be a significant treatment failure rate. Since their mechanisms of action are different, we wanted to evaluate the effectiveness of combination therapy. MATERIALS AND METHODS: Of 214 patients treated for erectile dysfunction (ED), 65 were not fully satisfied with the firmness of their erections via monotherapy. Responses were evaluated using the International Index for Erectile Function (IIEF) questionnaire before and after treatment. Group I consisted of 33 patients who tried maximal dose intraurethral alprostadil monotherapy initially, followed by the maximal dose of sildenafil monotherapy, and were still unsatisfied. Group II consisted of 32 patients who tried the maximal dose sildenafil monotherapy initially, followed by the maximal dose of alprostadil monotherapy, and were also unsatisfied. There 65 patients then underwent combination therapy. RESULTS: 60 out of the 65 patients stated they were satisfied with combination therapy. Questionnaire scores for erectile function were 23.1+/-2.0 (114%) for combination therapy vs. 19.2+/-1.8 (77%) and 15.2+/-1.6 (41%) for sildenafil and alprostadil monotherapies (p<0.05). There were no significant differences in responses between the two groups. The men also reported improvement in intercourse and overall satisfaction. CONCLUSIONS: Combination therapy may be an option for motivated patients who have a suboptimal response from monotherapy. PMID- 10859439 TI - Metallic stents in gynecologic cancer: an approach to treat extrinsic ureteral obstruction. AB - OBJECTIVE: We report on our experience with the use of self-expandable metal stents for the treatment of extramural ureteral obstruction in patients with gynecologic cancer to restore ureteral patency and to alleviate the ureterectasis and hydronephrosis proximal to the ureteral narrowing. METHODS: Fourteen women (mean age 48 years) with obstructive uropathy secondary to gynecologic malignancies were treated successfully by placement of Wallstent self-expandable intraureteral metallic stents. The patients were followed for a mean period of 15 (range 9-24) months. RESULTS: Obstructive uropathy was resolved in all cases. In 1 patient placement of an additional, totally coaxial, stent was considered necessary because of tumor ingrowth, occurring 6 months after the procedure. In another patient, tumor overgrowth invading the borderline area between the proximal ureteric end and the metallic prosthesis was seen 12 months after stent placement causing obstruction. Thus, an additional Wallstent was implanted overlapping the initially placed stent. Patency was achieved in all the remaining ureters, during the follow-up period, without any need for further intervention. CONCLUSION: Implantation of self-expandable metal stents is a safe and effective method for bypassing ureteral obstruction due to gynecologic malignancies. PMID- 10859440 TI - Late complications of ureteral stents. AB - OBJECTIVE: To review morbidity and late complications of ureteral stent insertion and to specifically evaluate hydronephrosis as a radiologic finding of obstruction in the presence of an indwelling ureteral stent. METHODS: In this prospective study, we evaluated 110 stented kidneys in a group of 90 patients. Of 110 stents, 52 were left in place for 3 months, 23 for 6 months, 11 for 9 months, and 24 for up to 12 months. With the stent in place, patients were followed by plain abdominal X-ray 1 and 30 days after stenting. Further follow-up was performed through ultrasound and plain film every 3 months until scheduled date for stent removal or the appearance of complications. RESULTS: In 11 of 110 cases (10%) there was stent fragmentation and in 9 (8.2%) stent migration. In 10 cases (9.1%), there was no change in the severity of the hydronephrosis, but because of flank pain or urinary tract infection with fever, the stents had to be removed. In 6 cases (5.4%) hydronephrosis developed or worsened after stenting. Of the 110 ureteral stents, 32.7% had to be removed because of late complications. CONCLUSIONS: Although ureteral stenting is undoubtedly an important procedure for the release of ureteral obstruction, the indications for stent insertion should be carefully considered in each patient. Late complications of ureteral stents are frequent and appear in one third of the patients. Close follow-up of stented patients is valuable in early detection of morbidity or complications, and in such cases the stent should be removed or exchanged as soon as possible. PMID- 10859441 TI - Analysis and reliability of data from 24-hour frequency-volume charts in men with lower urinary tract symptoms due to benign prostatic hyperplasia. AB - OBJECTIVES: The aims of this study were to analyse the data from frequency-volume charts and to study the reliability of these charts in men with lower urinary tract symptoms (LUTS) due to benign prostatic hyperplasia (BPH). METHODS: Males with LUTS due to BPH were consecutively included in the study if they met the criteria of the International Consensus Committee on BPH, voided more than 150 ml during uroflowmetry, residual volume and prostate size were estimated and frequency-volume charts were completed correctly. From the frequency-volume charts, voiding habits and fluid intake were evaluated. RESULTS: 160 patients could be included. Another 28 patients who met all other criteria did not complete the frequency-volume charts correctly. Agreement exists between reported voided volumes in the literature and those found by us. We found a significant correlation (p<0.001) between nycturia and score on symptom question 7, and between diuria and score on symptom question 2 of the AUA symptom index. The difference between results obtained from frequency-volume charts completed during 24 h and those obtained from charts completed during three or more 24-hour periods was negligible with respect to the variation of data at an individual level. CONCLUSION: Frequency-volume charts are reliable in the investigation of patients with LUTS due to BPH. Reporting on frequency-volume charts during just 24 h is sufficient to gain insight into their voiding habits during normal daily life. PMID- 10859442 TI - Sexual function in 131 patients with benign prostatic hyperplasia before prostatectomy. AB - OBJECTIVE: The aim of the present study was to ascertain the frequency of sexual dysfunction in patients with benign prostatic hyperplasia (BPH) before prostatectomy. METHODS: The study population included 131 patients aged 55-74 years (mean 61.4+/-2.7) with BPH. The patients had been complaining of daytime urgency and nocturia for the last 1-14 years (average 4.5+/-1.5 years). The diagnosis of BPH was based on the anamnestic data, the International Prostate Symptom Score (IPSS(0-35)) and quality of life assessment (L(0-6)), and results of digital rectal examination, transrectal ultrasound, and uroflowmetry. Sexual dysfunction was determined by retrospective analysis of the psychosexual history (with and separately from the spouse), penile brachial index (PBI), nocturnal penile tumescence (NPT), and blood hormone levels (testosterone). RESULTS: The patients were divided into two groups by severity of the urinary disorder: group I, severe (IPSS(0-35)-L(6); n = 70); and group II, mild relative to group I (IPSS(32-34)-L(4-5); n = 61). In group I, 15 patients (21.4%) performed normal coitus, 24 (34.2%) had weak coitus with incomplete penetration, and 21 (44.2%) had unsuccessful coitus because of a weak erection. In group II, 28 patients (45.9%) had normal coitus, 25 (40.9%) incomplete penetration, and 8 (13.1%) unsuccessful coitus. There was no significant correlation between sexual function and the patients' general health condition. The quantitative assessment of sexual dysfunction yielded the following results in groups 1 and 2, respectively: negative NPT in 32.4+/-1.8 and 24.4+/-2.1% (p<0.05); PBI <0.6 in 33.4+/-1.7 and 22.3+/-1.2% (p<0.001); and testosterone decrease to <12 nmol/l in 36.4+/-1.2 and 28.5+/-1.2% (p<0.05), respectively. The differences between the groups were significant (p<0.05) for all three parameters. CONCLUSIONS: Considering the 44. 2% rate of unsuccessful coitus in the patients with a severe urinary dysfunction compared to only 13.1% in those with a milder dysfunction and the significant correlation between severe urinary dysfunction and measures of sexual dysfunction, we suggest that BPH may be a risk factor for sexual dysfunction. PMID- 10859443 TI - Tolerability of 3.5 versus 2.5 high-energy transurethral microwave thermotherapy. AB - PURPOSE: To evaluate the tolerability of high-energy microwave thermotherapy in patients with benign prostatic hyperplasia (BPH) using two different treatment protocols (Prostasoft 2.5 and Prostasoft 3.5). MATERIALS AND METHODS: Pain and discomfort during treatment was evaluated using a visual analog scale in 39 patients undergoing 60-min Prostasoft 2.5 treatment and 41 patients undergoing 30 min Prostasoft 3.5 treatment. The duration of transurethral microwave thermotherapy (TUMT) 3.5 treatment is significantly shorter than TUMT 2.5 treatment. RESULTS: The pain level is significantly higher at the beginning of the Prostasoft 3.5 treatment compared to the Prostasoft 2.5 treatment. The reported pain level becomes similar 10 min into treatment, and remains similar to the end of the 3.5 treatment (at 30 min), when the pain level returns to baseline. The 2.5 protocol patients experience continuously increasing pain until the end of the treatment at 60 min. One minute following termination of treatment, the pain level drops back to the baseline level. No correlation between the level of pain and the baseline subjective or objective voiding parameters was observed. A correlation is also absent between the pain level, age and catheterization time. There only seems to be a weak correlation between the pain level and TUMT energy in the Prostasoft 2.5 treatment group. CONCLUSIONS: Both TUMT 2.5 and TUMT 3.5 are well tolerated. Even though patients undergoing TUMT 3.5 treatment experience more discomfort initially, the ultimate discomfort is similar to the TUMT 2.5 treatment, during the first 30 min. Shortening of treatment time significantly reduces the pain and discomfort experienced by the patient. Pretreatment parameters are not predictors of the pain level experienced. PMID- 10859444 TI - Modified extrafascial radical retropubic prostatectomy technique decreases frequency of positive surgical margins in T2 cancers <2 cm(3). AB - OBJECTIVES: In an effort to decrease the frequency of postoperative positive surgical margins (+SM), a modified extrafascial radical prostatectomy technique was developed and evaluated. METHODS: 402 consecutive radical prostatectomy specimens removed for clinical stage T2 cancers from 1987 to 1994 were histologically examined prospectively for tumor volume, extraprostatic extension and +SM. Surgical technique modification was introduced in 1990. We compared the histologic status and biological outcome of the prostatectomy cases in 1987-1989 (n = 166) to those treated from 1990 to 1994 (n = 236). RESULTS: The two series were comparable in (1) clinical stage and preoperative (PSA, (2) tumor volume, grade and location, and (3) capsular penetration, seminal vesicle and lymph node status. +SM fell from 32 to 25% overall, but for 146 (36%) prostates with a tumor volume <2 cm(3), +SM fell from 21 to 6% which was statistically significant. Outcome measured by biological progression showed a decrease from 33% for +SM to 13% for -SM for cases with a tumor volume <2 cm(3). For cancer volumes >2 cm(3), the incidence of +SM did not vary significantly. We describe the anatomic details necessary for exposure of periprostatic fascias and extrafascial dissection at (1) the prostatourethral junction which ensures wide excision of the anterior and apical aspect of the prostate, (2) the posterior and apical area (development of the prerectal space), lateral and posterior areas at the base of the prostate which ensures wide excision of the rectoprostatic fascia (Denonvilliers's fascia) and lateral prostatic fascia. CONCLUSIONS: Differences in surgical technique probably accounted for the significant decrease in +SM for those T2 cancers with volumes < or =2 cm(3) which represents 36% of the T2 cancers in our series. Recent screening with PSA (T1c cancers) increases the incidence of these cancers < or =2cm(3). This modified uni- or bilateral anatomic extrafascial prostatectomy with improved +SM and biological progression rates for T2 cases should be evaluated for T1c cases. PMID- 10859445 TI - Stage migration in clinically localized prostate cancer. AB - OBJECTIVES: To determine whether migration of pathological tumor stages in patients with clinically localized prostate cancer exists and whether this is due to an increasing frequency of treating patients with clinically insignificant cancer. METHODS: 1,063 radical retropubic prostatectomies were performed in patients with clinically localized prostate cancer in one institution within 7.5 years (from 1992 until June 1999). All specimens were prospectively processed according to the Stanford protocol. These were then analyzed regarding the migration of pathological tumor stages and cancer volumes. RESULTS: Within the observation period, the annual rate of radical retropubic prostatectomies increased by 225% from 69 to 224 cases. The authors noted a decline of advanced tumor stages (from 65 to 40%) and an increase in pathological T2 tumors (from 30 to 55%). The rate of small cancers (<0.5 cm(3)) remained stable between 2 and 5% over the last 5 years. CONCLUSION: The data confirm trends which were observed in large US centers with increasing detection and treatment of localized prostate cancer without unnecessary treatment of clinically insignificant cancers. PMID- 10859446 TI - Is microvascular invasion on radical prostatectomy specimens a useful predictor of PSA recurrence for prostate cancer patients? AB - OBJECTIVE: Angiogenesis is believed to play an important role in tumor progression and metastasis. The goal of this study was to investigate the clinical utility of vascular invasion in prostate cancer patients treated by radical prostatectomy as a predictor of PSA recurrence. METHODS: Between 1993 and 1998, 241 patients underwent radical prostatectomy at our institution and had routine analysis of vascular and/or lymphatic invasion (V/LI). V/LI was correlated with preoperative parameters including digital rectal examination (DRE), Gleason score (GS) on biopsy and serum PSA, with the pathological findings and with biochemical recurrence. RESULTS: V/LI incidence was 12.4% (30 of 241 patients). Of the 30 patients with V/LI, 28 (93%) had GS > or =7 (67%) had a pT3 disease and 7 had SV invasion (23%). V/LI was not associated with DRE and GS on prostate needle biopsy. However, V/LI was correlated with the worst pathological findings including pT3 disease, seminal invasion, positive surgical margins and GS on prostate specimen > or = 7. Biochemical recurrence-free survival was 92.5% for the patients without V/LI as compared to 30.1% for patients with V/LI on prostate specimen examination (p = 0.0001). Multivariate analysis showed that preoperative serum PSA, Stage and V/LI were independent predictors of PSA recurrence. Patients with pT2 disease without V/LI had a biochemical recurrence free survival of 99 vs. 31% in patients with V/LI (p = 0.0001). CONCLUSION: This study demonstrated that V/LI is strongly associated with biochemical recurrence after radical prostatectomy. The routine analysis of V/LI should be considered as a routine evaluation of the radical prostatectomy specimen. PMID- 10859447 TI - Prostate-specific antigen complexed to alpha(1)-antichymotrypsin in the early detection of prostate cancer. AB - OBJECTIVES: To study the usefulness of the complexed-to-total (C:T) prostate specific antigen (PSA) ratio in the early detection of prostate cancer in patients with a total PSA value <4.0 ng/ml. PATIENTS AND METHODS: Total PSA and PSA complexed to alpha(1)-antichymotrypsin were measured in plasma from 193 men with benign prostatic hyperplasia (BPH) and 34 with prostate cancer. The diagnosis was confirmed in 28 BPH and 16 prostate cancer patients by biopsy and in 165 BPH and 18 prostate cancer patients by histological study following transurethral prostatectomy or open prostatectomy. RESULTS: The area under the receiver operating characteristic (ROC) curve was significantly greater for the C:T PSA ratio (0.908) than for total PSA (0.692) (p<0.001). Using a cut-off point of 0.83 for the C:T PSA ratio and regardless of the digital rectal examination (DRE) finding, 20 of the 34 prostate cancer patients would have been given a correct diagnosis (59% sensitivity) and in only 8 of the 193 BPH patients would a biopsy have been necessary (96% specificity). With a cut-off of 0.79, the sensitivity increased to 85% with a specificity of 92%. When the analysis was restricted to the 44 patients with abnormal DRE, the area under the ROC curve for the C:T PSA ratio was 0.919, and a cut-off point of 0. 78 gave a sensitivity of 87% and a specificity of 93%. Using a cut-off of 0.63, all prostate cancers were detected (100% sensitivity) and 54% of the negative biopsies would have been eliminated. For the 183 patients diagnosed following surgery, a cut-off of 0.82 gave a sensitivity of 72% and a specificity of 94%. CONCLUSION: Our results show that the C:T PSA ratio significantly improves the clinical utility of the PSA assay for detecting prostate cancer in patients with total PSA < 4 ng/ml, increasing the sensitivity without significantly increasing the number of biopsies. PMID- 10859448 TI - Prediction of prostate volume based on total and free serum prostate-specific antigen: is it reliable? AB - OBJECTIVE: To analyze the utility of total/free prostate-specific antigen (PSA) and age as predictors of the prostate volume in men with symptomatic benign prostatic hyperplasia (BPH) and no evidence of prostate cancer. PATIENTS AND METHODS: Total and free serum PSA were determined in 681 patients with normal digital rectal examination and symptomatic BPH using the Hybritech method. Prostate volume was measured by transrectal ultrasound (TRUS). TRUS-guided biopsy was performed in 459 (67.4%) of the patients with a serum PSA >4.0 ng/ml. RESULTS: The relationship with prostate volume was best described in a log linear fashion by free PSA (R(2) = 0.367), total PSA (R(2) = 0.264) and age (R(2) = 0.017). Multiple linear regression demonstrates no significant influence of age. Free PSA was able to predict the individual TRUS prostate volume +/-10% in 67% of the patients and +/-20% in 91.2% and total PSA in 63 and 90. 9%, respectively. CONCLUSION: Prostate volume is strongly related with free and total PSA. Both determinations would be able to predict the TRUS prostate volume +/-20% in more than 90% of the patients. PMID- 10859449 TI - Total bilateral ureteral replacement for stenosing ureteritis in Henoch-Schonlein purpura. AB - Stenosing and calcified ureteritis occurring in Henoch-Schonlein purpura remains a rare urological complication. The authors describe their own experience of the diagnosis and treatment of this rare pathology along with a review of the literature. Recognition and early surgical management may prevent serious renal outcome. In cases of ureteral replacement, different procedures using ileal segment have been proposed. We report the results of the Cockett and Goodwin procedure with a follow-up of more than 25 years. PMID- 10859450 TI - Continent ileal pouch using the serous-lined principle. AB - OBJECTIVES: The flap valve principle is not easily applicable to an ileal pouch since a submucosal tunnel is difficult to create. We attempted to construct an ileal pouch with an umbilical stoma applying the serous-lined principle for both ureteral implantation and construction of a continent valve. METHODS: In 9 patients, a continent pouch with an umbilical stoma was created entirely from an ileal segment. Adopting the serous-lined principle, a continent valve was created by appendix in 2 cases, tapered ileum in 3 cases, and reconfigured ileum in 4 cases. These were implanted into the anterior suture line of the pouch and embedded into the serous-lined tunnel formed by the pouch wall. RESULTS: Patients can catheterize the pouch easily with a 14-Fr catheter. Postoperatively, pouch capacity is over 400 ml with complete continence. CONCLUSION: This technique can provide a simple and effective continent ileal pouch facilitating umbilical anastomosis. As a continent valve, the reconfigured ileal segment seems most useful for application of the serous-lined principle. PMID- 10859451 TI - Use of water jet resection in renal surgery: early clinical experiences. AB - OBJECTIVE: Various surgical techniques have been developed to reduce the risk of bleeding during dissection of renal tissue. Water jet technology has been designed for a gentle transection of parenchymal organs. We report about first experiences with a new water jet resection device in kidney surgery. METHODS: 7 patients underwent open surgery because of renal cell carcinoma or nephrolithiasis. In tumor patients a partial nephrectomy, in 1 stone patient a nephrolithotomy, and in the other 1 a lower pole resection were performed. RESULTS: By means of water jet resection tissue was dissected effectively avoiding damage to the vascular structures, which were then ligated selectively. Resection time was between 14 and 30 min with a minimal intraoperative blood loss. No significant problem was seen postoperatively. CONCLUSIONS: The results demonstrate that water jet technology is an applicable tool for transection of renal parenchyma. It has been shown to reduce the risk of blood loss significantly compared to other techniques. PMID- 10859452 TI - Gene expression of the phosphodiesterases 3A and 5A in human corpus cavernosum penis. AB - OBJECTIVE: The following study was performed to evaluate the importance of phosphodiesterases 3A (PDE3A) and 5A (PDE5A) for the regulation of penile smooth muscle tone. Furthermore, indications of side effects of specific inhibitors in certain tissues as well as of a possible relation between the level of PDE3A and PDE5A gene expression and erectile dysfunction should have been deduced from the results obtained. METHODS: Total ribonucleic acid was isolated from different human tissues (urogenital tract, gastrointestinal tract, cardiovascular system and central nervous system) and subjected to RTPCR analysis and Northern blotting using primers and probes specific for PDE3A and PDE5A. RESULTS: As shown by RT PCR and Northern blotting, PDE3A and PDE5A mRNAs exhibit a distinct distribution throughout the tissues examined but were 2-fold higher in cavernous tissue than in all other tissues investigated. However, there were no significant differences in the levels of gene expression between the two subgroups of patients. CONCLUSIONS: Very high expression levels of PDE3A and PDE5A in human cavernous tissue underscore the physiological importance of these enzymes for the regulation of penile erection, emphasizing their therapeutical and pharmacological relevance. The distribution pattern of the mRNA for the isoenzymes PDE3A and PDE5A may explain the pharmacological effects as well as the side effects of milrinone and sidenafil. PMID- 10859453 TI - Abrogation of the negative influence of opioids on IL-2 immunotherapy of renal cell cancer by melatonin. AB - IL-2 immunotherapy has been proven to be effective in the treatment of metastatic renal cell cancer (RCC). However, several drugs commonly used in the palliative therapy of cancer may potentially influence IL-2 efficacy, since the anticancer immunity has appeared to depend on complex interactions between immune system and psychoneuroimmunomodulation. In particular, experimental studies and preliminary clinical investigations have shown that the opioid substances, namely morphine, may suppress the anticancer immunity and the efficacy of IL-2 itself. In contrast, other neuroactive substances, in particular the pineal hormone melatonin (MLT), have been proven to stimulate the immune response, including the anticancer immunity, and to abrogate opioid-induced immunosuppression. On this basis, a study was planned to evaluate the effect of a concomitant MLT administration on the efficacy of IL-2 immunotherapy in advanced cancer patients chronically treated with morphine for cancer-related pain. The study was carried out in 30 metastatic RCC patients under chronic therapy with morphine at oral doses ranging from 60 to 120 mg/day. Patients were randomized to receive morphine alone or morphine plus MLT (20 mg/day orally in the evening). The immunotherapeutic cycle consisted of IL-2 subcutaneous administration at a dose of 6 million IU/day for 6 days/week for 4 consecutive weeks. In nonprogressing patients, a second cycle was planned after a 21-day rest period. The percent of partial responses achieved in patients treated with morphine alone was significantly lower than that observed in patients concomitantly treated with MLT (1/16 vs. 4/14, p<0.05). Moreover, the 3-year percent of survival was significantly higher in patients concomitantly treated with MLT (p<0.01). In contrast, no diminished analgesic efficacy of morphine occurred in patients concomitantly treated with MLT. This preliminary study seems to suggest that the negative influence of morphine therapy for cancer-related pain on the clinical efficacy of IL-2 cancer immunotherapy may be abrogated by the concomitant administration of the immunomodulating pineal neurohormone MLT. PMID- 10859454 TI - Genetic susceptibility to prostate cancer PMID- 10859455 TI - Lipid peroxidation and nitrite plus nitrate levels in brain tissue from patients with Alzheimer's disease. AB - BACKGROUND: Although oxidative stress has been implicated in the pathogenesis of neurodegenerative diseases, data found in the literature are still a matter of controversy. OBJECTIVE: To study lipid peroxidation and nitrite plus nitrate levels in brain specimens (frontal, parietal, temporal, and occipital cortices) from 8 patients with Alzheimer's disease (AD) in comparison to 6 young and 7 age matched controls. METHODS: Lipid peroxidation was estimated by the thiobarbiturate test for malonaldehyde and nitrate/nitrite using the Griess method. RESULTS: Our results show a clear malonaldehyde increase tendency in all brain areas from AD cases in relation to older individuals and younger controls. In temporal cortex there was no difference between AD cases and age-matched controls. We suggest that this may be due to incipient temporal cortex damage in control individuals and that this area could be more susceptible to age-related changes. We showed that nitrite plus nitrate levels significantly decrease in brain areas of AD cases in relation to young controls. Except for the frontal cortex, there was a trend for a decreased concentration of nitrite/nitrate in the aged group in all brain areas studied as compared with young controls. CONCLUSION: Our results support a major role for lipid peroxidation alterations in cerebral cortex of AD patients and showed not only a decrease in nitrite/nitrate levels in frontal cortex of AD cases in relation to young and old individuals, but also a reduction in these nitric oxide metabolites in the other cortical areas as related to young controls. PMID- 10859456 TI - Increased soluble tumor necrosis factor receptor levels in the serum of elderly people. AB - BACKGROUND: Soluble (s) forms of tumor necrosis factor (TNF) receptors are the only natural molecules known to interfere with TNF activity by competing for TNF binding with receptors on target cells. In a variety of pathologic situations, the concentrations of sTNF receptors (R) increase. OBJECTIVE: To discuss possible causes of increased risks for infectious disease and cancer seen in the elderly. METHODS: The participants were healthy subjects (n = 48) of three age groups (young, middle-aged, and elderly). Patients with senile dementia of Alzheimer type (n = 25) were also studied. For detection of cytokines, interleukin (IL) 1alpha, IL-1beta, IL-6, macrophage colony-stimulating factor (M-CSF), granulocyte colony-stimulating factor, and TNF-alpha were measured in serum by enzyme-linked immunosorbent assays, as were soluble (s) IL-1 receptor antagonist (IL-1ra), sIL 6R, p55sTNF-R, and p75sTNF-R. RESULTS: IL-1alpha, IL-1beta, IL-6, and TNF-alpha were not detected, and sIL-6R and IL-1ra concentrations were not significantly different between the three age groups. However, sTNF-R and M-CSF were increased in sera from the elderly, both healthy and demented. A significant correlation was seen between sTNF-R and M-CSF concentrations. CONCLUSIONS: Increased sTNF-R levels may oppose the physiologic and protective effects of TNF by interference with its receptor binding. This interaction may contribute to the susceptibility of the elderly to infectious and neoplastic diseases. PMID- 10859457 TI - Body composition and osteoporosis in elderly women. AB - OBJECTIVES: To study body composition in elderly osteoporotic women to determine the relationship of body weight, body fat mass and lean mass to bone mineral density (BMD), and to investigate the association between one-leg balance, osteoporosis and sarcopenia. DESIGN AND SETTING: A cross-sectional study of a community-based population in Toulouse, France. METHODS: For each participant, whole body composition and BMD were estimated using a dual-energy x-ray absorptiometry scanner. We investigated balance using a one-leg balance test. PARTICIPANTS: 129 healthy women aged 75-89 years, volunteers, ambulatory and living at home. RESULTS: Total fat mass and appendicular skeletal muscle mass (ASM) were significantly lower in osteoporotic women than in the age- and sex matched non-osteoporotic controls [18.7 +/- 4.6 vs. 22.2 +/- 6.6 for total fat mass (p < 0.01); 13.1 +/- 1.6 vs. 13.8 +/- 2.2 for ASM (p < 0. 05)]. We did not find a positive association between osteoporosis and sarcopenia (OR = 0.75, CI 0.3-1.84), osteoporosis and one-leg balance (OR = 1.27, CI 0.51-3.17), or sarcopenia and one-leg balance (OR = 1.31, CI 0.52-3.36). There were significant positive correlations between BMD in all areas and body measurements (weight, fat mass, lean tissue mass), but fat mass accounted for more of the variance in total body and femoral BMD than lean tissue mass. Total fat mass alone, in a multivariate model, was correlated with whole body BMD, whereas femoral BMD was associated with both fat mass and lean tissue mass. CONCLUSION: Higher values of fat mass and lean tissue mass may have a protective effect on femoral bone density. Sarcopenia and osteoporosis are not necessarily linked with balance. PMID- 10859458 TI - Prevalence of antibodies against hepatitis C virus in the elderly: a seroepidemiological study in a nursing home and in an open population. The Collaborative Group. AB - BACKGROUND: The prevalence of antibodies against hepatitis C virus (anti-HCV) increases in the general population with advancing age. Several discrepancies exist in the epidemiology of HCV, however, when selected elderly population groups are tested. OBJECTIVE: To evaluate the HCV prevalence in two groups of elderly people living in the same geopgraphical area of northeast Italy, i.e., one including residents of a nursing home, the other including subjects living at home. METHODS: The overall sample included 496 subjects (mean age 79.31 +/- 8.9 years); 288 were in a nursing home, and 208 were living at home. Enrollment in the latter group was based on all subjects over 65 years old listed under the public health service in the same district. The overall rate of adhesion to the study was 90%. Each subject was administered an anonymous questionnaire testing sociodemographic data and risk factors for HCV infection. Serological tests included: anti-HCV and hepatitis B virus serum markers. Multiple logistic regression analysis was performed to evaluate risk factors for anti-HCV positivity. RESULTS: Anti-HCV positivity was found in 34 of 288 (11.8%) elderly in the nursing home and in 23 of 208 (11.1%) in the open population. When the total population was considered, females exhibited a significantly a higher prevalence of anti-HCV than males (13.4 vs. 7.5%, p < 0.05). In both males and females, the highers rate of anti-HCV prevalence was found among the 75- to 79 year-old subjects. A decline in anti-HCV prevalence was observed in the very old subjects (over 80 years of age). None of the anti-HCV-positive subjects was found to be coinfected with hepatitis B surface antigen. However, multiple logistic regression analysis identified the age group between 70 and 79 years, female gender, and positivity for antihepatitis B surface antigen and/or antihepatitis B core antigen as independent variables significantly associated with HCV prevalence. CONCLUSIONS: The prevalence of anti-HCV proved identical among elderly people living in the nursing home or at home, suggesting that nursing homes do not represent a risk factor for HCV infections; the significant association between HCV prevalence and antihepatitis B surface antigen and/or antihepatitis B core antigen positivity supports a common route of transmission of the two viruses; these findings would suggest that there was an epidemic of HCV infection during the Second World War and in the years immediately afterwards. PMID- 10859459 TI - Audiometric and epidemiological analysis of elderly in the Veneto region. AB - BACKGROUND: Presbyacusis, or age-related hearing loss, has become a problem of increasing social interest due to the rise in the mean age of the population. Investigations performed to date have generally been carried out with the aid of self-reporting questionnaires, without the support of objective findings. OBJECTIVE: The purpose of this study was to analyze an extensive series of elderly people from different areas of the Veneto region to obtain an epidemiological descriptive analysis, as detailed as possible, of their presbyacusis, considering their hearing thresholds at various frequencies, distinguishing findings according to sex and age in classes and the geographical area where the survey was performed. METHODS: The survey was carried out collecting information from the audiometric reports on 13,710 subjects of both sexes aged 60 years and over, with a proportion of males (M/F x 100) of 92.02%, referred spontaneously for hearing examination to the ENT departments of eight hospitals between 1986 and 1994. The catchment area includes three provincial capitals and five mainly rural municipalities. RESULTS: The results show that the hearing threshold rises progressively with age in both sexes. The hearing loss is milder in women than in men, especially at the higher frequencies. No important differences emerged among findings recorded in their 80s and in their 90s or among findings in the different geographical areas considered. CONCLUSIONS: Statistical descriptive analysis confirms the typical trend of the audiometric curve in presbyacusis, tracing the typical audiometric curve of old age described in the literature. The mean values of the audiometric curve reveal no significant differences between people residing in the country and people living in the cities. PMID- 10859460 TI - Pulmonary embolism in the elderly: clinical, instrumental and laboratory aspects. AB - OBJECTIVE: To focus on diagnostic and therapeutic problems of pulmonary embolism in the elderly. METHODS: Retrospective analysis of 5 years of clinical, instrumental, and laboratory data (collected at the time of hospital admission) for patients 65 years and older with pulmonary embolism proven by a high probability scintigraphic lung scan or necropsy. Sixty-eight patients, 46 females and 22 males, 78.61 +/- (SD) 7.71 years old, were enrolled in the study. RESULTS: Dyspnea, chest pain, tachycardia, and tachypnea were the most common symptoms and signs; they were present alone or in combination in all patients. Bed rest over 4 days was found in 65% of the patients and deep vein thrombosis in the leg in 35%. Only 7 patients were on anticoagulant therapy which was likely to reduce the incidence of pulmonary embolism. The mortality was 29.5%. Major bleeding due to anticoagulant therapy was observed in 4.4% of the patients; 1 case was fatal. Sinus tachycardia, ST segment and T wave abnormalities in anterior leads, and incomplete bundle branch block were the most frequent electrocardiographic findings. Chest X-ray was normal in 19.5% of the patients and compatible with pulmonary embolism in 10%. A transthoracic two-dimensional echocardiogram was abnormal in 74% of the cases, with involvement of the right ventricle in the majority of them. Many patients had laboratory parameters within the normal range. The value of the latex agglutination D-dimer assay was less than the cutoff value of 500 microg/l in 16% of the patients. Hypoxemia and a high alveolar-arterial oxygen gradient were the most frequent aspects of the arterial blood gas analysis. Respiratory alkalosis was observed in only one third of the patients. CONCLUSIONS: Pulmonary embolism is often underdiagnosed in the elderly. Clinical, instrumental, and laboratory findings are nonspecific. Only acute suspicion can increase the number of diagnoses, reduce the time to diagnosis, and improve the prognosis. PMID- 10859461 TI - Life shortening and physician assistance in dying: euthanasia from the viewpoint of German legal medicine. AB - BACKGROUND: Around the world heated debates have broken out on the topic of active euthanasia. Specialists in the field of 'forensic medicine' have taken full part in these discussions. OBJECTIVE: The present survey from the point of view of forensic medicine begins with a look at current terminology and at the laws pertaining to euthanasia in Germany. These laws are then contrasted with actual practice, including a description of the increasing acceptance of active euthanasia by the German population and its legalization in Holland. The main argument against active euthanasia is that its formal acceptance in law would cause the dam of restraint to burst, culminating in widespread misuse, as already seen in recent serial killings by nurses in hospitals and homes for the elderly around the world. CONCLUSIONS: Contrasted to this are the arguments for taking active steps at the end of life, including emotional considerations such as the revulsion against mechanized medicine and the fear of pain and rational arguments such as the necessity to end a 'life unworthy of life', to save medical costs, and obtaining prior consent in 'living wills'. Such considerations have put in jeopardy the moral integrity of the medical profession - and thus the layperson's trust in physicians - around the world. In Germany especially the history of mass killing during the Nazi era constitutes a fundamental argument against active euthanasia. As a consequence, in Germany active euthanasia will not receive legal sanction, although recommendations on rendering dying more bearable are permitted. PMID- 10859462 TI - Is depression a risk factor for dementia or cognitive decline? A review. AB - BACKGROUND: It is generally accepted that depression can be associated with significant cognitive deficits and that depression can be comorbid with dementia. OBJECTIVE: This review seeks to go further and ask whether depression earlier in life can be a risk factor for subsequent dementia or for cognitive decline. METHODS: A review was made of the epidemiological evidence from case-control and prospective studies that depression is a risk factor. The literature was also reviewed in relation to six hypotheses that might explain an association: (1) depression treatments are a risk factor for dementia, (2) dementia and depression share common risk factors, (3) depression is a prodrome of dementia, (4) depression is an early reaction to cognitive decline, (5) depression affects the threshold for manifesting dementia, and (6) depression is a causal factor in dementia. RESULTS: A meta-analysis found that depression was associated with an increased risk of subsequent dementia in both case-control studies (95% CI for relative risk: 1.16-3.50) and prospective studies (95% CI: 1.08-3.20). There was little support for hypotheses 1 and 2. The other hypotheses have limited support, but warrant further research. CONCLUSION: There is sufficient evidence to take seriously the possibility that depression is a risk factor for dementia and cognitive decline. Further work is needed to examine depression as a prodrome of vascular dementia, depression as an early reaction to perceived cognitive decline, the effects of depression on the threshold for manifesting dementia, and depression as a source of hippocampal damage through a glucocorticoid cascade. PMID- 10859463 TI - News from the IAG. PMID- 10859464 TI - Antigen recognition by human gammadelta T lymphocytes. AB - Mature T lymphocytes carrying the conventional alphabeta T cell receptor (TCR) recognize peptide antigens in the context of MHC class I (CD8+) and MHC class II (CD4+) antigens, respectively. In striking contrast, human gammadelta T lymphocytes recognize unconventional antigens via their heterodimeric TCR in a non-MHC-restricted fashion. In this brief review, we discuss recent progress in the identification of ligands that are specifically recognized by human gammadelta T cells. It appears that gammadelta T cells have evolved to supplement the cellular immune response towards antigens that are not seen by alphabeta T lymphocytes. PMID- 10859465 TI - The immune response in adenoids and tonsils. AB - The adenoid and tonsils are lymphoid tissues located in the pharynx that play an important role in host defense against invading antigens of the upper respiratory tract. Histologically, these structures consist of four well-defined microcompartments which all participate in the immune response: the cryptepithelium, the follicular germinal center with the mantle zone and interfollicular area. With the uptake of antigen by M-cells present in the cryptepithelium a process is initiated which ultimately results in the generation and dissemination of antigen-specific memory and mainly dimeric IgA-producing effector B-lymphocytes. This process requires successful cognate interactions between antigen-presenting cells and lymphocytes and mutually between lymphocytes, which depend not only on antigen-specific signals but also on the expression of various complementary adhesion and costimulatory molecules. PMID- 10859466 TI - Lipid transfer protein: a pan-allergen in plant-derived foods that is highly resistant to pepsin digestion. AB - BACKGROUND: Lipid transfer proteins (LTPs) are small molecules of approximately 10 kD that demonstrate high stability. They have recently been identified as allergens in the Rosaceae subfamilies of the Prunoideae (peach, apricot, plum) and of the Pomoideae (apple). They belong to a family of structurally highly conserved proteins that are also present in non-Rosaceae vegetable foods. OBJECTIVE: The aim of this study was to investigate the cross-reactivity to non Rosaceae LTPs, and to study the role of protein stability in allergenicity. METHODS: Thirty-eight patients with a positive SPT to Rosaceae fruit extracts enriched for LTP were characterized by interview and SPT. To investigate IgE cross-reactivity between Rosaceae and non-Rosaceae LTPs, RAST and RAST inhibition as well as ELISA and ELISA inhibition were performed, using whole food extracts and purified LTPs. Both purified natural LTPs (peach, carrot and broccoli) and Pichia pastoris recombinant LTPs (carrot and wheat) were included. Pepsin digestion was used to address the role of stability in the allergenicity of LTPs. RESULTS: IgE antibodies to Rosaceae LTPs reacted to a broad range of vegetable foods, including Gramineae (cereals), Leguminosae (peanut), Juglandaceae (walnut), Anacardiaceae (pistachio), Brassicaceae (broccoli), Umbelliferae (carrot, celery), Solanaceae (tomato), Cucurbitaceae (melon), and Actinidiaceae (kiwi). Binding and inhibition studies with purified natural and recombinant LTPs confirmed their role in this cross-reactivity. Many of these cross-reactivities were accompanied by clinical food allergy, frequently including systemic reactions. Antibody binding to LTP was shown to be resistant to pepsin treatment of whole extract or purified LTP. CONCLUSION: LTP is a pan-allergen with a degree of cross-reactivity comparable to profilin. Due to its extreme resistance to pepsin digestion, LTP is a potentially severe food allergen. PMID- 10859467 TI - Dietary nucleotides can up-regulate antigen-specific Th1 immune responses and suppress antigen-specific IgE responses in mice. AB - BACKGROUND: It has been reported that dietary nucleotides enhance T helper cell activities. In this study, we have determined the effects of dietary nucleotides on antigen-specific Th1 and Th2 responses and IgE responses. METHODS: Ovalbumin (OVA)-specific T cell receptor (TCR) transgenic (OVA-TCR Tg) mice, 3 weeks old, were fed a nucleotide-free diet (NT(-) diet) or the NT(-) diet supplemented with dietary nucleotides (NT(+) diet) for 4 weeks. Cytokine production by spleen cells and macrophages obtained from these mice was measured in vitro. BALB/c mice, 3 weeks old, immunized intraperitoneally with OVA adsorbed onto alum, were fed the NT(-) diet or the NT(+) diet for 4 weeks. Serum levels of antigen-specific antibodies in the BALB/c mice were determined by ELISA. RESULTS: The level of production of antigen-specific interferon-gamma by spleen cells was significantly higher in the OVA-TCR Tg mice fed the NT(+) diet than in the control mice. The levels of secretion of bioactive IL-12 by spleen cells and peritoneal macrophages were also significantly increased in the NT(+) diet group. The serum OVA-specific IgE level was significantly decreased in BALB/c mice fed the NT(+) diet compared with those fed the NT(-) diet. CONCLUSION: These results show that dietary nucleotides up-regulate the antigen-specific Th1 immune response through the enhancement of IL-12 production and suppress the antigen-specific IgE response. PMID- 10859468 TI - An amplified ELISA inhibition method for the measurement of airborne soybean allergens. AB - BACKGROUND: Measurement of the soybean aeroallergen in Barcelona and other cities where soybean is unloaded is of increasing importance in controlling population exposure and evaluating the influence of such exposure on the persistence of asthma symptoms. OBJECTIVE: The aims of the study were: (1) to standardize an amplified ELISA inhibition method for the quantification of soybean aeroallergen and (2) to compare this method to a previously described RAST inhibition method. METHODS AND RESULTS: An amplified competitive ELISA inhibition method with a biotin-streptavidin system was carried out using a pool of sera from soybean sensitized patients. The results were expressed as U/ml using a low-molecular mass soybean allergen as reference standard. Reproducibility was calculated by statistically comparing the slope of the regression lines of the standard curve of 4 consecutive assays and by determining the coefficient of variation (CV) of the percent inhibition data for each point of several independent standard curves, each from the same assay (intra-assay) and also from a separate assay (inter-assay). No significant differences in the slopes were obtained by analysis of covariance (ANCOVA) F = 1.04. The CV between assays varied between 4 and 22% (for the assay range used in the reference standard) and was greater than the CV within assays (5-10%). Only values with a CV(%) smaller than 20% were considered acceptable. 78.5% of the samples satisfied this criterion. The RAST inhibition and ELISA inhibition methods were compared by difference plots from the values of 338 air filter eluates. The intraclass correlation coefficient was 0.456 (p < 0.001). After the results of both methods were classified as lower and higher than 165 U/m(3), the kappa index was 0.46 (p < 0.001). CONCLUSIONS: The data obtained in the present study are comparable to those reported from other similar immunoassays. Moreover, despite the difficulty in comparing air-sampling values from different laboratories, the kappa index may be taken to represent fairly good agreement beyond chance between both methods. All these data demonstrate that the present immunoassay is useful for measuring airborne soybean aeroallergens and can also be applied to evaluate the relationship between exposure and the development of asthma symptoms. PMID- 10859469 TI - Development of a novel enzyme-linked immunosorbent assay for blood and urinary eosinophil-derived neurotoxin: a preliminary study in patients with bronchial asthma. AB - BACKGROUND: Eosinophil-derived neurotoxin (EDN), also called eosinophil protein X (EPX), has been suggested to be a useful marker of eosinophilic inflammation. However, no commercial enzyme-linked immunosorbent assay (ELISA) kit for EDN is available yet. METHODS: EDN was purified from pooled urine from healthy male volunteers. Polyclonal and monoclonal anti-EDN antibodies were subsequently raised, and a sandwich ELISA for EDN was established. EDN levels in serum, plasma and urine from asymptomatic patients with bronchial asthma were measured by the ELISA method. Some of the blood samples were also measured by a commercial radioimmunoassay (RIA) kit. RESULTS: The ELISA method detected human EDN with a minimum detection limit of less than 0.62 ng/ml and did not cross-react with other eosinophil granule cationic proteins including eosinophil cationic protein. The intra- and interassay coefficients of variation of the ELISA method ranged from 2.6 to 3.6% and from 6.5 to 9.4%, respectively. Good linearity was observed with serially diluted different samples, and the recoveries of the purified EDN added to serum samples ranged from 85 to 110%. Median EDN concentrations in serum (36.9 vs. 19.1 ng/ml), plasma (23.0 vs. 14.5 ng/ml) and urine (118.2 vs. 72.1 microg/mmol Cr) were significantly raised in asymptomatic asthmatic patients compared with healthy control subjects. EDN levels in serum, plasma and urine from the patients significantly correlated with the number of peripheral blood eosinophils, but not total serum IgE levels. A significant relationship between EDN values measured by the EPX-RIA kit and the EDN-ELISA method was observed. CONCLUSIONS: We have developed a novel efficient ELISA method to specifically measure blood and urinary EDN, which may be useful to study the role of eosinophils in allergic diseases including bronchial asthma. PMID- 10859470 TI - Inhibitory effects of human neutrophil functions by the 45-kD glycoprotein derived from the parasitic nematode Trichinella spiralis. AB - AIM: We evaluated the effect of the 45-kD protein of Trichinella spiralis (gp45), purified by affinity chromatography, on random migration and chemotaxis, the oxidative metabolism of human neutrophils and on the CD11b upregulation induced by formyl-methionyl-leucyl-phenylalanine (f-MLP). METHODS: Donor neutrophils incubated with different amounts of gp45 (0.5, 1, 1.5, 2 microg/ml) or buffer and the random migration and chemotaxis, evaluated by means of a special technique of image analysis, and the chemiluminescence response to f-MLP or phorbol myristate acetate (PMA) were analyzed. The effect on CD11b upregulation was assessed incubating cells with the protein, when activating them with f-MLP. RESULTS: The results showed that gp45 inhibited both random and stimulated migrations, and reduced the response to f-MLP and PMA. Furthermore, gp45 significantly reduced the upregulation of the CD11b induced by f-MLP. CONCLUSION: The results show that gp45 inhibits PMN in different functions, suggesting an anti-inflammatory action. PMID- 10859471 TI - Isolation of a lactose-binding protein with monocyte/macrophage chemotactic activity. Biological and physicochemical characteristics. AB - BACKGROUND: We established a T cell line, STO-5, which constitutively produced monocyte/macrophage chemotactic activity via human T cell lymphoma-leukemia-virus induced transformation of normal human T cells. METHODS: We isolated and purified a lactose-binding protein, MCF-pl5-L (MW of about 50 kD, pl of about 5) from a conditioned medium of STO-5. By using highly purified MCF-pl5-L, its biological and physicochemical properties were elucidated in comparison with C5a and MCP-1. RESULTS: MCF-pl5-L exhibited an evident dose-dependent monocyte chemotactic activity (MCA). MCF-pl5-L had no or little affinity for heparin unlike chemokines such as MCP-1. We further found that MCF-pl5-L exhibited potent chemotactic activity not only for monocytes but also for alveolar macrophages. In contrast, C5a and MCP-1 failed to show evident chemotactic activity for alveolar macrophages though they did show MCA. MCF-pl5-L failed to exhibit evident eosinophil and neutrophil chemotactic activities, indicating its chemotactic activity is selective for monocytes/macrophages. Regarding the biological functions of MCF-pl5-L other than MCA and chemotactic activity for alveolar macrophages, we found that MCF-pl5-L but not C5a and MCP-1 could prolong the life span of alveolar macrophages, probably by inhibiting apoptosis of macrophages, and stimulate the production of TNF-alpha from macrophages. CONCLUSIONS: These results suggest that MCF-pl5-L plays a role as an immune modulator for monocytes/macrophages in the site. PMID- 10859472 TI - Dexamethasone and cyclosporin A affect the maturation of monocyte-derived dendritic cells differently. AB - In contrast to the confirmed effects of glucocorticoids (GCs) and cyclosporin A (CyA) on T cells, the effects of both agents on antigen-presenting cells (APCs), especially on dendritic cells (DCs), are still poorly understood. In this study, we cultured monocyte-derived DCs (MoDCs) under a variety of stimulations in the presence or absence of these immunosuppressants and compared their effects on the activation of MoDCs by these stimulations. The stimulations used were the following: three bacterial toxins, including lipopolysaccharide (LPS), staphylococcal enterotoxin A (SEA) and streptococcal pyrogenic exotoxin A (SPEA), the combination of IL-1beta and TNF-alpha, and an agonistic anti-CD40 antibody. All of these stimulations increased the expression of CD54, CD83, CD86, and HLA DR antigen, and the production of TNF-alpha in MoDCs. When MoDCs were treated with dexamethasone (Dex) during the stimulation, Dex significantly suppressed the augmentation of CD86 expression and TNF-alpha production induced by all of these stimulations. In contrast, when MoDCs were treated with CyA, it inhibited only the effects induced by the superantigens, SEA and SPEA, but not that induced by LPS, the combination of cytokines, or anti-CD40 antibody. The augmentation of CD54 or HLA-DR antigen expression was not significantly suppressed by either Dex or by CyA. When we used MoDCs pretreated with each of these stimulations + Dex or + CyA as APCs, however, significant suppression of T cell proliferation was observed only in the case of the pretreatment with IL-1beta/TNF-alpha + Dex. The allogeneic T cell stimulation by MoDCs pretreated with the other combinations did not significantly differ from that treated with the stimulation alone. Our present study succeeded in demonstrating a clear difference between Dex and CyA in the activation of MoDCs. These differences may induce a significant difference in their final immunological responses. PMID- 10859473 TI - Interaction between wall shear stress and circumferential strain affects endothelial cell biochemical production. AB - The wall shear stress (WSS) of flowing blood and the circumferential strain (CS) driven by the pressure pulse interact to impose a dynamic force pattern on endothelial cells (ECs) which can be characterized by the temporal phase angle between WSS and CS, a quantity which varies significantly throughout the circulation. To study the interaction of WSS and CS on endothelial production of vasodilators (prostacyclin and nitric oxide) and a vasoconstrictor (endothelin 1), bovine aortic ECs were cultured on the inner surface of compliant tubes and subjected to various flow conditions: steady shear (10 dyn/cm(2)), oscillatory shear (10 +/- 10 dyn/cm(2), rigid tube), and oscillatory shear (10 +/- 10 dyn/cm(2)) with CS (8%) either in or out of phase with shear. The 4-hour production rates of vasoactive agents show that steady shear stimulates the highest production of vasodilators whereas oscillatory shear stimulates the highest vasoconstrictor production. The addition of CS in concert with oscillatory shear enhances the production of vasodilators and inhibits the production of vasoconstrictors, and this effect is modulated by the phase angle between WSS and CS. These data suggest that the interactions of WSS and CS are important in vascular regulation and remodeling. PMID- 10859474 TI - Differential expression of the alpha1 type VIII collagen gene by smooth muscle cells from atherosclerotic plaques of apolipoprotein-E-deficient mice. AB - We have investigated the morphology, growth and gene expression of smooth muscle cell (SMC) cultures derived from advanced atherosclerotic plaques and from non plaque-containing aorta of individual apolipoprotein-E-deficient mice. The initial outgrowth of cells was faster from plaques (P) than from non-plaque segments (NP), but the cells in P cultures divided more slowly than NP cells in subcultures. By the 6th passage, the general growth pattern, morphology, ploidy and response to mitogenic factors of the cells were no longer consistently different in P and NP cultures. However, by the use of differential display, several transcripts were identified that were differentially expressed in three independent pairs of P and NP cultures. One of the transcripts, from a type VIII collagen gene, was elevated in all the P cultures compared to their NP counterparts even at the 40th passage. The alpha1 type VIII collagen transcripts were also readily detectable by RT-PCR in freshly dissected plaques, but not in the normal parts of aortas from the apolipoprotein-E-deficient mice. In situ hybridization showed that the transcripts were limited to the fibrous cap of plaques. Thus, SMCs from atherosclerotic plaques produce type VIII collagen and this differential expression continues when the cells are maintained in tissue culture. PMID- 10859475 TI - Differential transmural distribution of gene expression for collagen types I and III proximal to aortic coarctation in the rabbit. AB - To assess the effects on the biosynthesis of collagen types I and III associated with an acute increase in blood pressure, we established a mid-thoracic aortic coarctation in the rabbit and studied gene expression and protein accumulation of these collagen types proximal to the stenosis 1, 3 and 7 days and 2, 4 and 8 weeks after coarctation. The mRNA level of type I collagen pro-alpha2(I) was maximal at 3 days and returned to normal at 4 weeks. mRNA of pro-alpha2(I) was localized mainly in the outer media, adventitia and intima. Accumulation of type I collagen and its precursors was increased by 3 days, peaked at 4 weeks, and decreased toward normal by 8 weeks, corresponding to the distribution of pro alpha2(I) mRNA. Gene expression for pro-alpha1(III) was similar to that of pro alpha2(I) but was distributed throughout the media. We conclude that the mechanical stresses associated with an acutely induced alteration in pressure initiate rapid gene expression for collagen types I and III in the aortic wall. The response for collagen type I, predominantly in the outer media and adventitia, suggests that these regions play an immediate role in the resistance to excessive dilatation of the aorta. The diffuse response for collagen type III in the media suggests participation in a more extensive remodeling response associated with the reinforcement and reorganization of the musculo-elastic fascicles. PMID- 10859476 TI - Gender differences in cutaneous laser doppler flow response to local direct and contralateral cooling. AB - To investigate the role of sympathetic neurovascular reactivity in the gender differences of cutaneous, cold-induced vasomotor response, we compared direct (at the site of cooling) and indirect (at a site remote from the cooling site) response measured by laser Doppler (LD) flowmetry in 12 healthy males and 12 healthy females. The females underwent testing twice, once in the follicular and once in the luteal phase of the menstrual cycle. We measured LD flow before and during local cooling of one hand at 15 degrees C. We found that local cooling evokes a significantly greater decrease in cutaneous LD flow in females than in males in direct as well as in indirect response conditions (p < 0.05). Comparing the response in females in the different phases of menstrual cycle, we obtained a significantly greater direct response to local cooling in the luteal phase than in the follicular phase (p < 0.05). In contrast, there was no menstrual-cycle dependent difference in the indirect response to cold. The results of our study strongly indicate that sympathetic neural reactivity, as assessed by way of an indirect response to a cold stimulus, significantly contributes to gender differences in the response to local cooling. In contrast, the variation in microvascular responsiveness to cold exposure due to the menstrual cycle is most probably caused by local vascular mechanisms rather than by variation in sympathetic neural reactivity to local cooling. PMID- 10859477 TI - Low-dose cerivastatin inhibits spontaneous atherogenesis in heterozygous watanabe hyper lipidemic rabbits. AB - The aim of this study was to investigate whether cerivastatin (BAYw6228), a new potent 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor, was able to prevent atherogenesis in heterozygous Watanabe heritable-hyperlipidemic (WHHL) rabbits, a model never tested before using this HMG-CoA reductase inhibitor. The heterozygous WHHL rabbits of our breeding developed mild hypercholesterolemia along with focal atherosclerotic lesions in the thoracic aorta. A 9-week treatment with cerivastatin at doses comparable to those used in humans (50 microg/kg/day) reduced serum total cholesterol levels (from 94.4 +/- 10.9 to 43.6 +/- 10.5 mg/dl, p < 0.005) and prevented aortic lesion development (intima/media ratio: 0.058 +/- 0.032 vs 0.946 +/- 0.282 in the placebo group, p < 0.0005). Using a panel of monoclonal antibodies specific to macrophages and able to recognize different smooth muscle cell (SMC) phenotypes, we observed that cerivastatin treatment affected the differentiation properties of SMCs and drastically reduced SMC and macrophage accumulation in the intima of the thoracic aorta. These data show that in the presence of moderate atherosclerotic lesions, such as those of heterozygous WHHL rabbits, low doses of cerivastatin exert an antiatherogenic effect. PMID- 10859478 TI - Can the green laser doppler measure skin-nutritive perfusion in patients with peripheral vascular disease? AB - The recently developed green laser (GL; wavelength 543 nm) is thought to measure perfusion derived from a more superficial skin layer than does the standard near infrared laser (RL; wavelength 780 nm). These lasers were used to investigate the disturbances in the different layers of skin perfusion in ischaemic legs before and after treatment and compared with capillary microscopy. Eighteen patients (20 legs) with different stages of leg ischaemia scheduled for a vascular intervention (11 males, 7 females; median age 73, range: 52-81 years; Fontaine stages II-IV) were investigated by means of capillary microscopy, visualising the nail fold capillary perfusion, and a laser Doppler, equipped with a special dual probe conducting both GL and RL. The probe was attached to the pulp and the dorsum of the big toe to assess skin perfusion at rest and during reactive hyperaemia, while sitting and while supine. Resting and hyperaemic perfusion using GL was low and significantly lower (p < 0.01) than with RL in both areas and positions. Laser Doppler perfusion was higher in the pulp than on the dorsum with both wavelengths (p < 0.05). The hyperaemia response was highest using GL and differed among the three techniques. Postural reduction of capillary and RL flow was reduced, but not with GL. After treatment, skin capillary perfusion improved more clearly than did the laser Doppler perfusion with either wavelength, while postural vasoconstriction improved only when measured with the capillary microscope. The differences found between RL and GL Doppler perfusion, but also between GL and capillary microscopy measurements suggest that the GL does measure the more superficial, but not exclusively the nutritive skin perfusion. Clinically, the use of the green laser in its present form in patients with leg ischaemia offers no advantage over the red laser. PMID- 10859479 TI - The mitogenic effect of 17beta-estradiol on in vitro endothelial cell proliferation and on in vivo reendothelialization are both dependent on vascular endothelial growth factor. AB - In addition to their actions on reproductive function, estrogens have important effects on endothelial cells. The present study was designed to evaluate the mechanism(s) by which 17beta-estradiol (E2) promotes endothelial cell proliferation. The potential involvement of vascular endothelial growth factor (VEGF) was investigated by the coadministration of polyclonal anti-VEGF antibody. First, the effect of E2 on the proliferation of cultured foetal bovine aortic endothelial cells (FBAEC) was studied. E2 stimulated this proliferation with an EC50 between 10(-11) and 10(-10) M and this effect was inhibited by the anti-VEGF antibody. The effect of a physiological dose of E2 was then studied in the rat model of carotid injury. After deendothelializing balloon injury, reendothelialization of the denuded surface may influence the growth of the underlying smooth muscle cells. Male Sprague-Dawley rats were castrated and then received E2 from subcutaneously implanted pellets that released 3.2 microg/kg/day. Endothelial regrowth (Evans blue staining) and neointimal thickening were evaluated 2 weeks after the carotid injury. In comparison to the placebo group, E2 increased the extent of reendothelialization (p = 0.0002) and reduced neointimal thickening (p = 0.0007). Anti-VEGF antibody abolished the effect of E2 on reendothelialization as well as on neointimal thickening. Thoracic aorta VEGF content was increased in E2-treated rats compared to control rats. In conclusion, the present study demonstrates that E2 increases endothelial cell proliferation in vitro and reendothelialization in vivo by means of a mechanism dependent on endogenous VEGF. This effect could contribute to the antiatherogenic effect of a physiological dose of E2. PMID- 10859480 TI - Thrombospondin-1 induces endothelial cell apoptosis and inhibits angiogenesis by activating the caspase death pathway. AB - Thrombospondin-1 (TSP1) is a potent natural inhibitor of angiogenesis. Although TSP1 has been reported to induce endothelial cell apoptosis in vitro and to downregulate neovascularization in vivo, the molecular mechanisms that link these two processes have yet to be established. Here we report that TSP1 mediates endothelial cell apoptosis and inhibits angiogenesis in association with increased expression of Bax, decreased expression of Bcl-2, and processing of caspase-3 into smaller proapoptotic forms. The ability of TSP1 to induce both endothelial cell apoptosis in vitro and to suppress angiogenesis in vivo was blocked by the caspase-3 inhibitor z-DEVD-FMK. TSP1 also attenuated VEGF-mediated Bcl-2 expression in endothelial cells in vitro and angiogenesis in vivo. Furthermore, TSP1 induced endothelial cell apoptosis and inhibited neovascularization in sponge implants in SCID mice. We conclude that TSP1 induces endothelial cell apoptosis and inhibits neovascularization by altering the profile of survival gene expression and activating caspase-3. PMID- 10859481 TI - Modulation of IL-18 production in the adrenal cortex following acute ACTH or chronic corticosterone treatment. AB - Interleukin (IL)-18 is a proinflammatory cytokine and a stimulator of cell mediated immune responses. We have previously reported that acute stress stimulates the production of IL-18 mRNA in the glucocorticoid (GC)-producing cells of the adrenal cortex. In order to investigate the mechanisms governing the expression of IL-18 in the adrenal cortex, the effects of acute ACTH or chronic corticosterone treatment on the levels of IL-18 mRNA and protein were examined by in situ hybridization and Northern and Western blot assays. Adult male Sprague Dawley rats received a subcutaneous injection of ACTH or subcutaneous implantation of slow-release corticosterone pellets followed by an injection of saline or ACTH. After 4 h, ACTH induced a 4-fold increase in IL-18 mRNA levels and elevated the content of pro-IL-18 peptide. Six days of chronic corticosterone treatment did not alter the basal levels of IL-18 mRNA and reduced those of pro IL-18. Finally, ACTH treatment of animals under the corticosterone regimen induced a 2-fold increase in IL-18 mRNA and elevated the levels of the pro-IL-18 protein. The levels of the precursor, p45, and the active subunit p10 peptides of the IL-18-processing enzyme, IL-1beta-converting enzyme (ICE), showed no substantial differences in all the conditions tested. IL-1beta was not detected under these experimental conditions. These data demonstrate that the production of IL-18 in the adrenal cortex is stimulated by ACTH treatment and is not inhibited by the direct action of corticosterone. In contrast to the anti inflammatory action of GCs, IL-18 may have an immunostimulatory role during acute stress. PMID- 10859482 TI - Effect of interleukin-11 on body temperature in afebrile and febrile rats. AB - Interleukin (IL)-11 is a member of the gp130 family of cytokines. In contrast to IL-6 (another gp130 cytokine), IL-11 does not induce fever in humans. In the present study, the effect of recombinant human IL-11 (hrIL-11) injected intracerebroventricularly on body temperature of afebrile and febrile rats was studied. Results showed that: (i) hrIL-11 in doses of 5, 50 and 500 ng injected into the cerebral ventricles does not alter body temperature in rats; (ii) febrile response induced by intraperitoneal injection of E. coli endotoxin (50 microg/kg) was initiated more rapidly in rats injected with 500 ng of hrIL-11 in the cerebral ventricles, and (iii) the enhancement of the initial phase of fever induced by hrIL-11 was not accompanied by changes in plasma concentrations of IL 6 and tumor necrosis factor (TNF). These results indicate that hrIL-11 is not pyrogenic when administered into the brain ventricles. The data obtained also demonstrate that central application of hrIL-11 alters body temperature in conditions of pyrogenic stimulation, but that this effect is not due to the alterations in plasma concentrations of IL-6 or TNF. These data suggest that during the development of the systemic inflammatory response, activation of gp130 subunit becomes effective in altering body temperature. PMID- 10859483 TI - Effect of orchidectomy on the age-related modulation of IL-1beta and IL-1 receptors following lipopolysaccharide treatment in the mouse. AB - OBJECTIVE: To compare the effect of orchidectomy (ODX) in 7- and 24-week-old C57BL/6 mice on the age-related responses of the cytokine interleukin (IL)-1beta and its receptor to intraperitoneal injection of the bacterial endotoxin, lipopolysaccharide (LPS). METHODS: We measured IL-1beta concentrations in the plasma, hippocampus, hypothalamus and adrenal gland using ELISA and iodine-125 labeled recombinant human IL-1alpha ([(125)I]IL-1alpha) binding in the hippocampus following the intraperitoneal administration of saline or LPS. RESULTS: There were no significant differences in the concentrations of IL-1beta and its receptors in the brain and peripheral tissues between sham-operated and ODX mice in both age groups injected with saline. LPS induced significantly higher IL-1beta production in the plasma and hippocampus in sham-operated 24-week old mice than in 7-week-old mice. Coincident with the heightened IL-1beta response to LPS, hippocampal [(125)I]IL-1alpha binding was lower in 24-week-old mice than in 7-week-old mice after LPS injection in the sham-operated group. The age-related differences in the IL-1beta concentrations in the plasma and hippocampus and [(125)I]IL-1alpha binding in the hippocampus in response to LPS administration were abolished by ODX. Although LPS dramatically increased IL 1beta levels in the hypothalamus, no significant age-related differences in IL 1beta concentrations were seen, and ODX did not affect IL-1beta levels. In contrast, there were no significant differences between saline- and LPS-injected 7-week-old mice in relation to concentrations in the adrenal gland. Moreover, although the adrenal IL-1beta concentrations in 24-week-old mice were significantly higher than those in 7-week-old mice, ODX did not abolish these age related differences in concentrations in the adrenal gland. CONCLUSIONS: Our data suggest the involvement of testosterone (or gonadal product) in plasma and hippocampal IL-1beta regulation in relation to age, and demonstrate the importance of the gonadal development in mediating the effects of infectious challenge on the brain and immune function. PMID- 10859484 TI - Interleukin-2: structural and biological relatedness to opioid peptides. AB - Interleukin (IL)-2 is not only an immunoregulatory factor, but also an analgesic molecule. There are distinct domains of immune and analgesic functions in the IL 2 molecule. The analgesic domain is located around the 45th Tyr residue of human IL-2 in tertiary structure. Antiopioid (beta-endorphin, Leu-enkephalin, Met enkephalin and dynorphin A1-13) sera partially neutralized the analgesic activity of IL-2. Monoclonal antibody against the IL-2 receptor alpha subunit (Tac) could not block the analgesic activity of IL-2. There existed cross-reactivity between IL-2 and antiopioid sera by indirect ELISA. These studies show strong structural and biological similarities between IL-2 and opioid peptides. The tertiary structure around the 45th residue of IL-2 composes the analgesic domain that is similar to that of endogenous opioids. These results are consistent with the hypothesis that multiple domains of cytokines serve as the structural bases for the immunoregulatory and neuroregulatory effects of cytokines. PMID- 10859485 TI - Somatostatin and substance P induced in vivo by lipopolysaccharide and in peritoneal macrophages stimulated with lipopolysaccharide or interferon-gamma have differential effects on murine cytokine production. AB - We have investigated whether lipopolysaccharide (LPS) induces substance P (SP) and somatostatin (SOM) in popliteal lymph nodes in vivo and whether macrophages are a source of SP and SOM in vitro. We have also investigated the effect of SP and SOM treatment on the production of cytokines. SP reached a maximum 3 days after injection of LPS (100 microg/footpad) and then declined. SOM expression after LPS injection reached a maximum at 5-7 days. Stimulation of thioglycolate elicited peritoneal macrophages with LPS (20 microg/ml), recombinant interferon gamma (rIFN-gamma, 100 U/ml), and LPS plus rIFN-gamma induced SOM and SP. Thioglycolate-elicited, unstimulated peritoneal macrophages also synthesized these peptides. SOM (10(-12)-10(-8) M) significantly inhibited IL-6 and IFN-gamma production, whereas SP at those concentrations enhanced cytokine production by activated lymphocytes and macrophages. These findings suggest that neuropeptides which originate from macrophages and nerve fibers act as immunomodulators to mediate changes in the pattern of cytokine production. PMID- 10859486 TI - Modulation of early immune responses and suppression of Trypanosoma brucei brucei infections by surgical denervation of the spleen. AB - OBJECTIVE: To examine critical interactions between the nervous system and the immune system during experimental African trypanosomiasis. METHODS AND RESULTS: Inoculation of Trypanosoma brucei brucei resulted in early interferon (IFN)-gamma production, elevated corticosterone and prostaglandin E(2) (PGE(2)) levels and increased splenocyte proliferation, as measured by enzyme-linked immunospot assay, radioimmunoassay and thymidine incorporation assay, respectively. Splenic denervation suppressed IFN-gamma, corticosterone and PGE(2) production, enhanced splenocyte proliferation, and significantly reduced parasitemia and prolonged rat survival. CONCLUSIONS: Our data show substantial effects of the nervous system on early immune responses that may influence the outcome of this disease. These effects were not dependent on cytokine inhibitory mediators such as prostaglandins or stress hormones. More investigations are required to understand the evident neural control over the immune system during infectious challenges, which may assist in novel therapeutic approaches. PMID- 10859487 TI - Testosterone suppresses the response of the hypothalamic-pituitary-adrenal axis to interleukin-6. AB - OBJECTIVES: Endotoxin and the inflammatory cytokines interleukin (IL)-1 and IL-6 are potent activators of the hypothalamic-pituitary-adrenal (HPA) axis. Previous studies in the rodent and in the primate have shown that the responses of the HPA axis to endotoxin and to IL-1 were enhanced by gonadectomy and attenuated by testosterone or estradiol replacement. The mechanisms underlying these observations are unclear, but there is evidence that gonadal steroids have direct inhibitory effects on IL-6 synthesis and release. Since endotoxin and IL-1 both stimulate IL-6, the question arises as to whether the sex-steroid-induced suppression of the HPA response to endotoxin and IL-1 results solely from decreased IL-6 release, or whether other mediators are involved. METHODS: We have therefore examined the ACTH and corticosterone responses to IL-6 in intact and castrated male rats with and without testosterone replacement. Animals were castrated 2 weeks prior to study; testosterone was replaced by subcutaneous Silastic capsules. Four days prior to study, an indwelling right atrial catheter was implanted. Blood samples for ACTH and corticosterone radioimmunoassays were collected through the catheter 0, 20, 40, 60, 120 and 180 min after intravenous injection of recombinant human IL-6 (500 ng). RESULTS: IL-6 stimulated ACTH and corticosterone release in all groups, with peak stimulation occurring within the first hour. The release of both ACTH and corticosterone was significantly attenuated in the intact (n = 9) and testosterone-replaced (n = 5) animals compared to the castrated animals without replacement (n = 7). Peak ACTH levels were 340 +/- 58 and 133 +/- 41 pg/ml in the intact and testosterone-replaced animals versus 678 +/- 170 pg/ml in the castrated animals (p < 0.02). Peak corticosterone levels were 29 +/- 4.7 and 30 +/- 4.2 microg/dl in the intact and testosterone-replaced animals versus 47 +/- 5.8 microg/dl in the castrated animals (p < 0.05). CONCLUSIONS: We conclude that testosterone attenuates the response of the HPA axis to IL-6 in the rat. This would indicate that other mechanisms, in addition to the inhibition of IL-6 release, are responsible for restraining the HPA response to inflammatory stimuli in the presence of gonadal steroids. PMID- 10859488 TI - Conditioned immunosuppressive effect of cyclophosphamide on delayed-type hypersensitivity response and a preliminary analysis of its mechanism. AB - In the present study, camphor odor and intraperitoneal (i.p.) injection of cyclophosphamide (CY) were used as conditioned stimulus (CS) and unconditioned stimulus (US), respectively. In the unconditioned group, mice were exposed to camphor odor for 1 h followed by an i.p. injection of CY (75 mg/kg). On the next day, the above CS/US association trial session was repeated followed by smearing dinitrochlorobenzene (DNCB) on mouse abdominal skin for sensitizing the animal for delayed-type hypersensitivity (DTH) response. Five days after DNCB sensitization, mice were exposed to camphor odor (1 h), followed by an i.p. injection of CY, and then DNCB was smeared on the left ear of mice for the challenge of DTH response. Both the left/right ear weight ratio and the activity of leukocyte migration inhibitory factor (LMIF) were used as the index of DTH response, which was done 24 h after DNCB challenge. In the conditioned group, the treatment was the same as that in the unconditioned group, except that normal saline was injected on day 5 instead of CY. Furthermore, in order to analyze the mechanism of the conditioned response (CR), the mouse serum from the conditioned group (CR serum) was injected into normal mice 6 h prior to DNCB challenge. Results showed that in the conditioned group, left/right ear weight ratio and LMIF activity were statistically lower than that in the DTH group, and there was no difference between conditioned and unconditioned groups. Thus, an animal model of conditioned immunosuppressive response had been established. The results also showed that after CR serum was injected into normal mice, DTH response was also significantly suppressed. However, if CR serum was treated with dialysis (10,000 molecular weight cut-off), the suppressive effect of CR serum on DTH response disappeared. Taken together, the data suggested that a chemical compound(s) in serum, with a molecular weight less than 10,000, was important in mediating the conditioned immunosuppressive response. This may be a very important molecule(s) that could be very critical to our understanding of the interaction between the central nervous system and immune function. PMID- 10859489 TI - Transcriptional properties of Ptx1 and Ptx2 isoforms. AB - The Ptx (Pitx) family of homeobox transcription factors comprises Ptx1, Ptx2 and Ptx3. Ptx1 and Ptx2 are expressed in the stomodeum and its derivatives including the pituitary, as well as in mesodermal derivatives, whereas Ptx3 is expressed in one neuronal lineage of the brain and in the eyes. A large set of downstream target genes have been identified for Ptx1 in the pituitary gland where it acts as a pan-pituitary regulator of transcription. In particular, Ptx1 contributes to promoter- and lineage-specific transcription by interaction with cell-restricted factors such as SF-1, Egr-1, Pit1, and the basic helix-loop-helix heterodimer NeuroD1/Pan1. We describe the cloning from pituitary cells and the characterization of a Ptx1 isoform, named Ptx1b, generated by alternative promoter usage. The two Ptx1 and two Ptx2 isoforms have similar in vitro DNA binding specificities and they all activate transcription driven by a panel of pituitary promoters, including those for proopiomelanocortin, alphaGSU, LHbeta, FSHbeta, GnRH-R, TSHbeta, PRL, and GH. Also like Ptx1, the Ptx1b, Ptx2a, and Ptx2b transcription factors synergize with the structurally unrelated factors SF 1, Egr-1, Pit1, and NeuroD1/Pan1 to activate promoter-specific transcription. In conclusion, the pituitary transcriptional activities of the four Ptx isoforms do not appear to be dependent on the variant N-termini of these factors. PMID- 10859490 TI - Ontogeny of immunoreactive prostaglandin endoperoxide synthase isoforms in ovine fetal pituitary, hypothalamus and brainstem. AB - Parturition is initiated in sheep by an increase in the activity of the fetal hypothalamus-pituitary-adrenal (HPA) axis. Prostaglandins, known to augment the activity of this endocrine axis, have long been proposed as involved in the initiation of paturition. We have previously demonstrated that endogenously produced prostanoids augment the activity of the HPA axis, and we have proposed that the increased production of prostanoids within the fetal brain or pituitary at the end of gestation might be involved in the initiation of parturition. An important regulatory step in the biosynthesis of prostanoids is the activity of prostaglandin endoperoxide synthase (PGHS). The present study was designed to test the hypothesis that the abundance of one or both isoforms of PGHS (PGHS-1 and PGHS-2) increase in brain and/or pituitary at the end of gestation. We used immunoblot analysis to measure the abundance of immunoreactive PGHS-1 and PGHS-2 in pituitary, hypothalamus and brainstem collected from fetuses of known gestational ages. We found that the abundance of PGHS-1 was weakly but significantly increased at the end of gestation in the pituitary and brainstem. The abundance of PGHS-2, on the other hand, increased exponentially in the pituitary and hypothalamus with highest concentrations found in term fetuses. We conclude that these enzymes are developmentally regulated in pituitary and in brain regions important for HPA axis control. We speculate that the increased enzyme's abundance results in increased prostanoid biosynthesis near term, and is a link in the chain of events which initiates parturition. PMID- 10859491 TI - Regulation of C-fos protein in gonadotrope cells by oxytocin and gonadotropin releasing hormone. AB - The integrated regulation of luteinizing hormone (LH) from the anterior pituitary gland is vital to the functioning of the ovulatory cycle in the female and consists of several components acting at different time points. The best-studied is the rapid release of LH elicited by gonadotropin-releasing hormone (GnRH). The so-called primary (immediate early) response genes (PRGs), including c-fos, regulate relatively long-term activities, such as mitosis, protein synthesis, protein release and cell differentiation. Regular ovulatory cycles occur as a result of interaction of several peptide factors including the primary factor, GnRH and oxytocin, although GnRH and oxytocin do not have identical activities. We wished to determine whether oxytocin could mediate changes in expression of c fos protein and compare its effects with those of GnRH. Anterior pituitary glands were collected from female rats at proestrus and a single-cell suspension prepared. Cells were incubated with oxytocin or GnRH at selected concentrations for various times. C-fos protein was extracted and submitted to Western blot analysis. Other cells were stained immunohistochemically for c-fos and LH following incubation with the peptides and fixation. There was an increase in c fos protein from 15 to 60 min in Western blots of cells from all incubations. After immunohistochemistry, it was observed that both oxytocin (100 nM) and GnRH (100 nM) increased the percentage of cells that expressed c-fos protein (p < 0.001) and of cells that expressed LH (p < 0.001). The responses to the peptides were concentration dependent. We found that neither all LH-containing cells expressed c-fos, nor all c-fos-containing cells immunostained for LH. The effects of the peptides were not the same. High concentrations of GnRH (1 microM) induced the appearance of a higher percent of LH-containing cells having c-fos than did 10 nM GnRH (p < 0.01), whereas a lower percent of LH-containing cells with c-fos were observed when the oxytocin concentration was raised from 10 nM to 1 microM (p < 0.02). It appears, therefore, that the two peptides have different regulatory effects on LH-containing cells, indicating the possibility of specialized function. The results emphasize the suggestion that stimulation of LH secretion is not the sole index of gonadotrope-directed activity by a peptide. Collectively, these results indicate that the peptides oxytocin and GnRH are able to modulate processes that are associated with longer-term activities of gonadotropes and also demonstrate that specific subpopulations of LH-containing gonadotropes are stimulated to express c-fos. PMID- 10859492 TI - Blockage of N-methyl-D-aspartate receptors decreases testosterone levels and enhances postnatal neuronal apoptosis in the preoptic area of male rats. AB - Sexual dimorphism has been found in the preoptic area of the hypothalamus (POA), a major site of glutamate actions via N-methyl-D-aspartate (NMDA) receptors. The sexually dimorphic nucleus of the preoptic area (SDN-POA) of male rats exhibits about seven-fold greater nuclear volume than that of females. A naturally occurring neonatal neuronal apoptosis, that can be prevented by testosterone, may contribute to this sexual difference in SDN-POA nuclear volume. Since activation of NMDA receptors in the POA induces GnRH secretion, it may be involved in both elevation of serum testosterone and prevention of neuronal death in the SDN-POA. In the present study, protein expression of NMDA receptors in the POA of male and female fetuses was quantified on the day preceding the fetal testosterone peak (embryonic day 16; ED 16). Rats were then distributed in four groups: (1) untreated males, (2) untreated females, (3) males pretreated with MK-801 (a noncompetitive NMDA receptor antagonist), and (4) females pretreated with MK-801. Serum levels of testosterone were estimated on the afternoon of ED 18. Expression of Bcl-2 and Bax, as well as neuronal apoptosis in SDN-POA, were observed on postnatal day 8. The results showed that (1) expression of NMDA receptors in the POA of male fetuses was higher than that of females on ED 16; (2) levels of testosterone were lower in MK-801 pretreated male fetuses than in intact males on ED 18; (3) expression of Bcl-2 in the POA of MK-801 pretreated male rats was significantly less than that of control males; (4) the apoptotic incidence in the SDN-POA of MK-801 pretreated male rats was significantly greater than in control males, while there was no significant difference in apoptotic incidence in the SDN-POA between MK-801 pretreated and intact females. These results suggest that the NMDA receptor is highly expressed in prenatal male fetuses, and that it might play an important role in the elevation of testosterone levels. Moreover, activation of NMDA receptors may protect SDN-POA neurons from naturally occurring neuronal death, by modulating testosterone and/or Bcl-2 expression. PMID- 10859493 TI - Effect of suckling on prolactin receptor immunoreactivity in the hypothalamus of the rat. AB - Previous studies showed that expression of prolactin (PRL) receptor is increased in numerous hypothalamic nuclei in mid-lactating rats. The increase in PRL receptor expression could be initated by neurohormonal changes during proestrus or pregnancy, or by the suckling stimulus during lactation. The present study investigated whether the PRL receptor expression in numerous hypothalamic nuclei is altered by the suckling stimulus. Three groups (n = 4) of rats on lactation day 10 were used: a continuously suckled group, a nonsuckled group (pups removed for 12 h) and a resuckled group (pups removed for 12 and then resuckled for 9 h). Animals were perfused with 2% paraformaldehyde and brains were sectioned (20 microm) for the immunofluorescence study. Immunoreactivity was semiquantitatively analyzed by counting the immunoreactive cells and measuring the immunostaining intensity in a specific area. Neurons expressing PRL receptors were observed in numerous hypothalamic areas with the highest number being in the arcuate, paraventricular and supraoptic nuclei. The PRL receptor immunofluorescence in several nuclei was significantly decreased in the nonsuckled group, and recovered in the resuckled group. These areas included the ventromedial preoptic, ventrolateral preoptic, lateroanterior hypothalamic, ventrolateral hypothalamic and ventromedial hypothalamic nuclei. PRL receptor immunoreactivity in other areas was not significantly altered by the suckling stimulus. These results demonstrate that expression of PRL receptor in hypothalamic nuclei was differentially affected by the suckling stimulus. PRL receptors in those nuclei which were significantly altered by suckling stimulus may play more critical roles during lactation than those areas which were less sensitive to the suckling stimulus. PMID- 10859494 TI - Induction of pituitary adenylate cyclase-activating polypeptide mRNA in the medial parvocellular part of the paraventricular nucleus of rats following kainic acid-induced seizure. AB - We examined the effects of kainic acid (KA)-induced seizure on the expression of the pituitary adenylate cyclase-activating polypeptide (PACAP) gene in the paraventricular nucleus (PVN) of rats using in situ hybridization histochemistry. Subcutaneous administration of KA (12 mg/kg) in adult male Sprague-Dawley rats caused a progressive development of seizure behavior. An induction of the PACAP gene expression in the medial parvocellular part of the PVN (mpPVN) was observed 3, 6, 12, 24 and 48 h after subcutaneous administration of KA. From a nearly undetectable level, PACAP gene expression increased in the mpPVN and reached maximum 12 h after subcutaneous administration of KA. PACAP gene expression returned to near basal level 48 h after stimulation with KA. Using a specific monoclonal PACAP antibody, PACAP immunoreactivity (-IR) gradually increased during the following 24 h after KA administration. In controls, PACAP-IR was located exclusively in nerve fibers of the mpPVN, whereas KA administration induced PACAP-IR in cell bodies of the mpPVN, and a dense accumulation of PACAP IR nerve fibers in the external zone of the median eminence was observed. Induction of the PACAP gene expression following KA-induced seizure was significantly reduced by pretreatment with diazepam or MK-801 (nonselective N methly-D-aspartate receptor antagonist). These results suggest that PACAP in the hypothalamo-adenohypophysial system may have a hypophysiotropic role during KA induced seizure. PMID- 10859495 TI - Calcitonin inhibition of prolactin secretion in lactating rats: mechanism of action. AB - The effects of intracerebroventricular (10 ng/rat) or intravenous (10 or 40 microg/15 min/rat) administration of salmon calcitonin (sCT) on the prolactin (PRL) response to suckling and the activity of tyrosine hydroxylase (TH) were examined in lactating rats. Plasma concentration of PRL increased dramatically in control rats after the onset of the suckling stimulus, while administration of sCT resulted in inhibition of PRL response to suckling. The action of sCT was much more effective with intracerebroventricular administration, which totally blocked PRL release, compared to intravenous administration. The intracerebroventricular administration of sCT increased TH activity of tuberoinfundibular dopamine neuron (TIDA) in the stalk-median eminence, as measured by DOPA accumulation, while completely suppressing the PRL response to suckling. Injection of alpha-methyl-p-tyrosine (alpha-MT; 50 mg/kg), an inhibitor of TH and thus dopamine synthesis, increased PRL levels, and suckling caused a further increase in plasma concentrations of PRL. Injection of sCT (intracerebroventricularly) did not inhibit the PRL response to suckling in the presence of a depletion of dopamine. These results suggest that sCT inhibition of PRL secretion in lactating rats is mediated mainly by TIDA neurons without involvement of other neuroendocrine mechanisms. PMID- 10859496 TI - Seasonal birth patterns of neurological disorders. AB - Existing seasonal birth studies were reviewed for multiple sclerosis (MS), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), epilepsy, cerebral palsy, congenital malformations of the central nervous system and mental retardation. Epilepsy appears to have the most consistent pattern, with an excess of births in winter and a deficit in September. MS, ALS and possibly Parkinson's disease appear to have an excess of spring births. Studies of cerebral palsy are not conclusive, although there are suggestions that there may be an excess of summer births. The findings for Alzheimer's disease, congenital malformations of the central nervous system, and mental retardation are contradictory and insufficient to draw any conclusions. PMID- 10859497 TI - Inter-rater reliability of the diagnosis of vascular cognitive impairment at a memory clinic. AB - Consensus criteria for the diagnosis of vascular dementia (VaD) are gradually being replaced with data-based criteria. We report the inter-rater reliability of a new set of empirically-derived criteria for vascular cognitive impairment (VCI). Stratified sampling, with optimal allocation, was employed to randomly select 36 patients from the Queen Elizabeth II Health Science Centre's Memory Disability Clinic. Chart reviews were conducted independently by 4 physicians. Each physician classified the patients as having either: no cognitive impairment, VCI or Alzheimer's disease (AD). VCI was further classified both clinically (VCI without dementia, VaD or AD with a vascular component) and radiographically (infarcts, white matter changes, single strategic stroke). The intraclass correlation coefficient (ICC) for the diagnosis by physicians of VCI or otherwise was based on a repeated-measures analysis of variance with raters as the independent variable. A significant coefficient of reliability (average ICC = 0.88, 95% CI = 0.80-0.93) was obtained (H(o): rho /=65 years, n = 3,728) were screened at home with the Chinese version of the MMSE. Persons who screened positive for dementia, using different cutoff scores based on degree of literacy, were further evaluated using the criteria of DSM-III-R and ICD-10. Among subjects who screened positive in phase I, 134 were diagnosed as having dementia in phase II. The prevalence ratios of dementia were 2.68% in the population aged 60 years and older, and 3.49% in the population aged 65 years and older. The prevalence rates among those aged 65 years and older were 1.85% for Alzheimer's disease, 1.37% for vascular dementia and 0.27% for other dementia (including mixed dementia). The prevalence of all dementia and AD increased steeply with advancing age and was consistently higher in women, but it was not obviously higher for VaD in women. Alzheimer's disease was the commonest type of dementia. Our prevalence figures for dementia and AD are similar to those previously reported for China. PMID- 10859499 TI - A nationwide cohort study on Creutzfeldt-Jakob disease among human growth hormone recipients. AB - OBJECTIVE: In 1991, a Dutch patient who had been treated from 1963 to 1969 with human-derived growth hormone died of Creutzfeldt-Jakob disease (CJD). This study was performed to investigate whether among other Dutch human growth hormone recipients there were clinically suspected cases of iatrogenic CJD. METHODS: In a retrospective cohort study, all patients (n = 564) treated with human-derived growth hormone before May 1985 and recorded in the Dutch National Growth Registry were followed up until January 1995 for a clinical diagnosis of CJD. For this purpose, all human growth hormone recipients were linked to a database of the Foundation for Health Care Information comprising hospital discharges with a clinical diagnosis of iatrogenic CJD. Linkage of the two databases was performed on the basis of date of birth and gender. Subsequently, verification of patient's name and initials of the positively matched pairs took place. RESULTS: Linkage provided 37 positively matched pairs concerning 29 individual patients. After verification, no name from the hospital discharge records corresponded to the names of the human growth hormone recipients. CONCLUSIONS The follow-up of 564 Dutch human growth hormone recipients, who had been treated with human growth hormone until 1985 did not establish any clinically suspected case of iatrogenic CJD. Future cases, however, can still emerge due to the potentially long incubation period of prion diseases. PMID- 10859500 TI - Prevalence of Parkinson's disease in Bulgarian Gypsies. AB - We studied the prevalence of Parkinson's disease (PD) in Gypsies and Caucasians in Bulgaria. Cases were ascertained by examination in a clinic, a screening questionnaire and a door-to-door survey in a region of Sofia, Bulgaria. The prevalence of PD in the Gypsies was found to be 16/100,000 based on 1 case compared to 137/100,000 for Caucasians with 119 cases. It is concluded that PD is rare in Bulgarian Gypsies. PMID- 10859501 TI - Dietary intake of calcium, magnesium and antioxidants in relation to risk of amyotrophic lateral sclerosis. AB - Dietary factors have long been suspected of being risk factors for amyotrophic lateral sclerosis (ALS), but few human studies have been reported. To address several of the dietary hypotheses, a case-control study of risk factors for ALS conducted in New England in 1993-1996 included an abbreviated food frequency questionnaire. We examined the dietary intake of calcium, magnesium and antioxidants among 107 ALS cases and 262 community controls. Overall, these dietary factors were not related to risk of ALS, though modestly protective associations were suggested for magnesium and lycopene. PMID- 10859502 TI - Population attributable fraction of stroke incidence in middle-aged and elderly people: contributions of hypertension, smoking and atrial fibrillation. AB - We determined the population attributable fraction (PAF) of stroke due to hypertension (HT), atrial fibrillation (Af) and smoking in a Japanese community. Residents of Shibata (n = 2,302) who were surveyed initially in 1977 were followed until 1997. Two hundred and thirteen first strokes occurred. Among those 40-64 years of age, the risk ratio (RR) of Af was 11.24, followed by untreated HT (3.61), uncontrolled HT (3.69) and smoking (1.84). The PAFs, however, were 14.9% for smoking, 13.5% for untreated HT, 8.6% for uncontrolled HT and 3.6% for Af. Among those over 65 years, only Af was significant (RR 3.89) and the PAF was 6.0%. Determination of PAFs is also essential for designing effective stroke prevention programs in communities. PMID- 10859503 TI - Risk factors for intracerebral and subarachnoid hemorrhage among Hispanics and non-Hispanic whites in a New Mexico community. AB - BACKGROUND AND OBJECTIVE: A higher incidence of spontaneous intracerebral and subarachnoid hemorrhage among Hispanics than non-Hispanic whites has been measured in Bernalillo County, New Mexico. In an attempt to explain these differences, we compared historical vascular risk factors between Hispanics and non-Hispanic whites living in this community. METHODS: An ongoing telephone survey, the Behavioral Risk Factor Surveillance System, collected annual data about vascular risk factors among non-institutionalized, randomly selected adults. Data covering 6 years, 1988-1993, were analyzed. RESULTS: There were 843 Hispanic and 1,635 non-Hispanic white residents of Bernalillo County, New Mexico, who participated in this survey. Because Hispanics were significantly younger than non-Hispanic whites (37.7 vs. 43.4 years, p < 0.001), all other comparisons were adjusted for age. Prevalence of hypertension was similar between these ethnic groups (15-17%). Prevalence of alcohol drinking considered risky for abuse was similar between these ethnic groups (5-6%), but was significantly higher among Hispanic men than women (8.5 vs. 1.6%, p < 0.001). The quantity of alcohol consumption among those at risk for abuse was similar between these ethnic groups. Prevalence of current cigarette smoking was similar between these ethnic groups (22-23%), but Hispanics smoked significantly less than non-Hispanic whites (11.4 vs. 15.2 cigarettes per day, p < 0.001) and among non-Hispanic whites, men smoked significantly more than women (17.0 vs. 13.4 cigarettes per day, p = 0.001). CONCLUSION: The vascular risk factors which we compared between Hispanics and non-Hispanic whites do not help to explain the higher incidence of hemorrhagic strokes among the Hispanics in Bernalillo County, New Mexico. Additional risk factors for hemorrhagic strokes in these two ethnic groups should be studied. PMID- 10859504 TI - Ritleng intubation set: a new system for lacrimal pathway intubation. AB - Silicone tubing of the lacrimal pathway is used after lacrimal pathway injuries or canalicular stenosis. This procedure is often complicated by the so-called accordeon phenomenon. Far the last 3 years we have used a new system for lacrimal intubation: the Ritleng intubation set. It consists of a silcone tube which is connected to a polypropylene tube and a specially designed stainless probe for lacrimal insertion. Due to the smooth connection of the polypropylene tube and the silicone tube intubation can be performed easily and without the so-called accordeon phenomenon. In case of congenital or acquired lacrimal stenosis probing and silicone intubation can be performed in one step. We used this set in 19 lacrimal lacerations of the lacrimal pathway and in 7 cases of congenital and 12 cases of acquired lacrimal stenosis. With this specially designed lacrimal intubation set lacerated canaliculi can be quickly reapproximated. It allows probing of the lacrimal system and intubation in one procedure. In this report the technique and indications will be described and discussed. PMID- 10859505 TI - Indocyanine green angiography in Fundus flavimaculatus. AB - PURPOSE: To describe the characteristic findings of fundus flavimaculatus (Stargardt disease) as seen on indocyanine green angiography. METHODS: Twelve eyes of 6 consecutive patients with fundus flavimaculatus were studied by fundus color photographs, fluorescein angiography and indocyanine green angiography. RESULTS: Indocyanine green angiography allowed visualization of small, clearly demarcated areas in which hypofluorescence increased over time, leading eventually to a large reticular pattern with small areas of normal-appearing choroid encircled by a well-defined network of hypofluorescent curvilinear lesions. These hypofluorescent flecks were present in all 12 eyes but corresponded only partially to the yellow flecks visible on biomicroscopy of the fundus. The peripapillary area was well preserved on indocyanine green angiography and the periphery did not show any visible abnormalities. CONCLUSIONS: The hypofluorescent curvilinear areas visualized on indocyanine green angiography form a reticular pattern that is similar to the polygonal shape of the watershed zones between terminal choroidal arterioles, which supply the choriocapillaris. These dark areas may reflect choriocapillaris defects secondary to lysis of lipofuscin-engorged retinal pigment epithelial cells. The typical lesions of fundus flavimaculatus thus seem to be situated in areas of least vascular supply. Their absence in the peripapillary area, which benefits from anastomotic vascular connections, would support this hypothesis. PMID- 10859506 TI - Indocyanine green angiographic findings in choroidal hemangiomas: A study of 75 cases. AB - Indocyanine green (ICG) angiography is a new diagnostic modality that was suggested, in small series, to provide a typical angiographic pattern in cases of choroidal hemangioma. Our study, through an exceptionally large series of 75 patients, assessed in a prospective way whether a typical ICG pattern of choroidal hemangioma exists and what would be its possible variations. The most constant feature is the sequence of the different ICG angiographic phases. The arterial phase demonstrates the filling of intratumoral vessels on a hypofluorescent tumoral background. During the venous phase, the hemangioma reaches a stage of maximal ICG-A fluorescence, with superimposed hyper- and hypofluorescent spots. Sturge-Weber cases have also extratumoral hyperfluorescent spots. The late phase shows a hypofluorescent lesion with residual hyperfluorescent caverns and a well-delineated, but complex border structure. PMID- 10859507 TI - Ultrasound biomicroscopic analysis of the human ciliary body after 1 and 2% pilocarpine instillation. AB - We examined ciliary body thickness (CBT) of 11 human eyes after 1% followed by 2% pilocarpine instillation using ultrasound biomicroscopy. The examination revealed that CBT decreased in 2 eyes after 1% pilocarpine treatment but increased after 2% pilocarpine. In the eyes with decreased CBT by 1% pilocarpine, the pretreatment ciliary bodies were thicker and the changing rate of CBT induced by 2% pilocarpine, although not decreased, was also smaller than that in CBT increased eyes. These findings showed that the ciliary body was under relatively strong stimulus of endogenous acetylcholine and has subsensitivity to both 1 and 2% pilocarpine in CBT-decreased eyes. Configurative changes in CBT-decreased eyes may be explained by the localization of different muscarinic receptor subtypes in different portions of the ciliary muscle. PMID- 10859508 TI - Extracapsular cataract extraction with posterior chamber lens implantation in capsular glaucoma. AB - PURPOSE: To evaluate intraocular pressure (IOP) control after extracapsular cataract extraction (ECCE) with posterior chamber intraocular lens (PCIOL) implantation in patients with capsular glaucoma (CG) and coexisting cataract. METHODS: This prospective study included 20 patients (20 eyes) having CG and cataract whose IOPs were under 22 mm Hg and controlled with antiglaucoma medication. All patients had ECCE with PCIOL implantation and the follow-up period was at least 18 months. IOP was measured postoperatively at 3, 6, 9, 12 and 18 months and compared with preoperative IOP. RESULTS: Following the cataract extraction, PCIOL implantation produced a statistically significant reduction in IOP at all time points compared with the preoperative IOP during the study period (p < 0. 001). The mean preoperative IOP was 18.25 +/- 1.83 mm Hg; postoperatively at 1 month, it was 13.45 +/- 2.06 mm Hg; at 3 months 14.80 +/- 2.50 mm Hg; at 6 months 15.35 +/- 1.27 mm Hg, at 12 months 14.85 +/- 1.87 mm Hg and at 18 months 15.15 +/- 1.42 mm Hg (p < 0. 0008). The mean reduction in IOP was 16.98% from baseline at 18 months postoperatively. The mean number of antiglaucoma medication was reduced from 1.35/eye preoperatively to 0.60/eye postoperatively at 18 months (p < 0.0007). CONCLUSION: The result of our study revealed that ECCE with PCIOL implantation may be a reliable choice in controlling IOP in patients with CG. PMID- 10859509 TI - Development of glycine-accumulating neurons in retinal transplants. AB - Previous studies have shown that the fetal retina not only survives transplantation but also continues to develop and differentiate in the host eye. Several structural and functional proteins have been demonstrated in the transplanted retinas, and the presence of such proteins has been taken as evidence for the capability of retinal transplants to function. Glycine is an important inhibitory neurotransmitter and is found in a large number of the retinal neurons. Uptake of glycine rather than de novo synthesis is the main source of glycine in glycinergic neurons. The present study examined whether glycine-accumulating neurons develop normally in rabbit retina transplants. Embryonic day (E) 15 rabbit retinas were transplanted into the eyes of adult rabbits of the same strain. Transplants were allowed to survive for various times so that the grafts attained the equivalent ages of (donor age + survival time) E 19, 22 and 29 and postnatal days (PN) 2, 5, 9, 12, 19 and 58. On formaldehyde fixed cryostat sections of these transplants, glycine-accumulating neurons were demonstrated by immunohistochemistry by using an antibody against one of the glycine transporters: GLYT1. Immunoreactivity was first detected 2 days before birth and increased with age until it reached its mature level at PN 19. The immunoreactivity was found in cells belonging to the inner retinal layers, and in plexiform layers of the transplant equivalent to the normal inner and the outer plexiform layers. In places these cells integrated well with similar cells in the host. In the host retina, the immunoreactivity was found in proximal cell layers of the inner nuclear layer, in certain bipolar cells, and in the inner and the outer plexiform layers. The immunoreactivity was preserved even in the degenerated retina overlying the retinal graft. In conclusion, the present study demonstrates that glycine-accumulating neurons develop, integrate and survive in retinal transplants. PMID- 10859510 TI - Melanoma-associated retinopathy versus abnormal retinal function due to interferon-alpha/Isotretinoin therapy in cutaneous malignant melanoma. AB - PURPOSE: To analyze whether an abnormal retinal function in patients with a cutaneous malignant melanoma was due to paraneoplastic retinopathy or due to isotretinoin or interferon-alpha. METHODS: We studied 15 patients with malignant melanoma in stage IIa and IIb who are all participants in a randomized, multicentered, double-blind placebo-controlled clinical trial comparing interferon-alpha/isotretinoin versus interferon-alpha/placebo performed by the Department of Dermatology, University of Graz. Our assessment included a full ophthalmic history and examination, electrophysiological testing (ERG, EOG), dark adaption, color vision and visual field testing. RESULTS: The most prevalent ocular symptom patients complained about was ocular dryness (8 patients). Electrophysiological as well as psychophysical testings showed no abnormalities in 12 patients. In 1 patient the therapy was stopped because of electrophysiological and psychophysiological pathology. This patient suffered from severe reduction of night vision and visual disturbances. Another patient had had night blindness since childhood which remained stable. CONCLUSIONS: We postulate that in 1 of 15 patients, visual complaints are caused with a high probability by melanoma-associated retinopathy although, in the literature, isotretinoin is described to show similar effects on retinal function. PMID- 10859511 TI - A survey of trachoma: the histopathology and the mechanism of progressive cicatrization of eyelid tissues. AB - The aim of this study is to demonstrate the spectrum of conditions encompassed by the term 'trachomatous cicatrization of eyelid tissue', to discuss the mechanisms of scar tissue formation and to describe sequelae in this potentially blinding condition. Specimens of eyelid tissues were taken from 27 upper eyelids of 21 patients with entropion who underwent surgical procedures and 2 post-mortem upper eyelids with severe trachomatous entropion. Upper palpebral conjunctival swabs and biopsy specimens were taken from 5 patients with active trachoma and they were examined by fluorescence microscopy and routine histopathological methods. Conjunctival impression cytology samples were collected in all patients. In specimens taken from patients with active trachoma the inflammatory infiltrate was organized as lymphoid follicles in the underlying stroma and impression cytology showed cytoplasmic elementary bodies. In specimens taken from patients with scarring trachoma light microscopy studies showed subepithelial fibrous membrane formation, squamous metaplasia and loss of goblet cells, pseudogland formation in conjunctiva, degeneration of orbicularis oculi muscle fibres, subepithelial vascular dilatation, localized perivascular amyloidosis and subepithelial lymphocytic infiltration. Accessory lachrymal glands and the ducts of glands were compromised by subepithelial infiltration and scarring. The contraction of the subepithelial fibrous tissue formed by collagen fibres and anterior surface drying are the main factors contributing to the chronic cicatrization and entropion formation. PMID- 10859512 TI - Stevens-Johnson syndrome in India - risk factors, ocular manifestations and management. AB - We conducted a study to analyse the presentation, risk factors, ocular manifestations and ophthalmic management results of Stevens-Johnson syndrome (SJS) in Indian patients. A total of 20 patients with SJS with ocular involvement were studied. Female predominance (70%) was observed. The age ranged from 10 to 30 years (mean +/- SD: 16.85 +/- 5.96). The commonest precipitating risk factor was oral drug intake (80%), sulphonamide (30%; sulphamethoxazole-trimethoprim) being the commonest. One patient had reaction to ciprofloxacin which has not been reported previously. Four patients (20%) had spontaneous onset with no identifiable risk factor. None of the patients had specific infection per se as a risk factor. Conjunctival involvement and its sequelae were the major ocular manifestations. At presentation, the majority of the eyes (68%) had visual acuity less than 3/60. Despite appropriate medical treatment and surgical interventions (11 eyes; 28%) vision continued to deteriorate. In 1 eye, the vision improved after stem cell transplantation. Therapeutic penetrating keratoplasty could preserve ocular integrity in 1 patient. PMID- 10859513 TI - The apolipoprotein E epsilon4 allele is unlikely to be a major risk factor of age related macular degeneration in Chinese. AB - Apolipoprotein E (ApoE) is a major transporter of lipids and cholesterol in the nervous system. Age-related macular degeneration (ARMD), characterized by drusen containing lipids, was reported to show a lower frequency of the ApoE epsilon4 allele than control subjects. We sought to examine the association of this polymorphism with ARMD in Hong Kong Chinese. Among 98 ARMD subjects, the frequency of epsilon4 carriers showed a trend toward a decrease compared to controls, but it was not significant (11.2 vs. 15.0%, p < 0.52). The association of epsilon4 with an apparent reduced risk of ARMD was reported previously in the exudative form of the disease, however among 39 exudative ARMD patients there was also no significant difference in epsilon4 frequency (12.8%, p < 0.93). The lack of a statistically significant effect of epsilon4 may be due to the lower frequency of epsilon4 in Chinese than Europeans. Thus we cannot exclude a possible effect of this allele on ARMD risk, but we can conclude that this allele is likely not a major factor influencing ARMD risk in the Chinese. PMID- 10859514 TI - Immediate increase in aqueous humour endothelin 1 concentration and intra-ocular pressure after argon laser trabeculoplasty in the rabbit. AB - PURPOSE: To investigate the immediate changes of aqueous humour endothelin 1 (ET 1) concentration and intra-ocular pressure (IOP) after argon laser trabeculoplasty (ALT) in the rabbit. METHODS: Standard ALT was performed in one eye of 11 pigmented rabbits. IOP was measured with a Tono-Pen-2 tonometer before treatment, under general anaesthesia. Postlaser IOP measurements followed by aqueous humour aspiration were performed under general anaesthesia 30 and 60 min after treatment in 6 and in 5 animals, respectively. RESULTS: The aqueous humour ET-1 concentration was significantly higher (55.0 pg/ml) after ALT than in the contralateral eyes without laser treatment (8.2 pg/ml, p = 0.001). IOP increased significantly after ALT (p = 0.007) but remained unaltered (p = 0.10) in the contralateral eyes without treatment. No statistically significant difference was found either in postlaser IOP elevation or in interocular ET-1 difference between the groups. CONCLUSIONS: The results suggest that in the rabbit the increase in aqueous humour ET-1 concentration after ALT is immediate, detectable even 30 min after the laser treatment, followed by no further increase during the second 30 min period after ALT, and associated with an immediate postlaser IOP elevation within 1 h after ALT. These findings may suggest a similar response to ALT in the human eye, since a similar immediate IOP elevation is frequently observed after ALT in the clinical practice. PMID- 10859515 TI - Antiproliferative Wirkung von Genistein auf kultivierte retinale Pigmentepithelzellen vom Schwein. AB - The isoflavonoid genistein is known as a potent inhibitor of proliferation in endothelial cells and in some tumor cell lines in vitro and in vivo. Cell growth arrest is mediated by inhbitor of the tyrosine-kinase receptor, a target for many growth factors such as fibroblast growth factor, platelet-derived growth factor, epidermal growth-factor and vascular endothelial growth factor, which may play a role in the development of proliferative vitreoretinopathy (PVR). In this study we report on the antiproliferative effect of genistein on retinal pigment epithelial cells (RPE), which is the major cell type involved in PVR. METHODS: Cell culture experiments using porcine RPE treated with different concentrations (5 up to 100 microgram/ml) of genistein in minimum essential medium +10% fetal calf serum. Inhibition of proliferation was demonstrated by two different methods: (1) uptake of 5-bromo-2-deoxyuridin (BrdU) in S phase after different exposure times (2) expression of proliferating cell nuclear antigen 72 h after scratching confluent RPE. RESULTS: Exposure of RPE to genistein resulted in a concentration-dependent inhibition of cell proliferation at concentrations over 25 microgram/ml geinistein. BrdU incorporation was reduced to 65% after 24- and 48-hour treatment with 100 microgram/ml genistein in comparison with the untreated cells. Proliferation of RPE growing into the artificial wound was inhibited when cells were exposed for 72 h to more than 25 microgram/ml genistein. CONCLUSIONS: Genistein at concentrations over 25 microgram/ml inhibits RPE proliferation. Further studies will be required to determine the applicability of genistein or other phytoestrogens in order to prevent uncontrolled wound healing processes such as PVR. PMID- 10859516 TI - Bilateral serous retinal detachment associated with Alport's syndrome. AB - We report a 14-year-old girl with Alport's syndrome who developed bilateral exudative retinal detachment in the macula. With the development of chronic renal failure, bilateral serous retinal detachment appeared which had the retinal flecks characteristic of Alport's syndrome. The serous detachment was resolved and vision recovered following intensive hemodialysis. As far as we know this is the first case with documentation of the onset and resolution of serous retinal detachment in Alport's syndrome. PMID- 10859517 TI - Occlusions of branch retinal arterioles following amniotic fluid embolism. AB - Amniotic fluid embolism is a serious complication of pregnancy with a high mortality. We present a 28-year-old healthy woman who underwent dilatation and curettage for an elective abortion, followed by the sudden loss of vision in her left eye. Occlusion of one branch retinal arteriole was the initial finding of her left fundus, and two occlusions developed consecutively on the color fundus photographs. Fluorescein angiography demonstrated occlusions in three retinal arterioles among seven retinal arterioles originating from the optic disc. These findings suggest that possible mechanisms of amniotic fluid embolism are the unusual cause in retinal arteriolar occlusions. Here clinical course and ophthalmic findings are reviewed, and the relationship between amniotic fluid embolism and retinal arteriolar occlusions is discussed. PMID- 10859518 TI - The biology of tumor invasion, angiogenesis and lymph node metastasis. AB - It is now well established that the development of cervical metastases, in particular those with extranodal extension of tumor, negatively impacts both regional control and survival of patients with laryngeal carcinoma. This chapter will begin with an introduction of the important molecular events associated with the transition of the squamous epithelium of the upper aerodigestive tract to metastatic squamous cell carcinoma. We will then review the critical cellular events identified as the tumor progresses from an in situ to invasive and finally a metastatic head and neck squamous cell carcinoma. Finally we will review data from our own and other laboratories which are studying the process of new blood vessel growth (angiogenesis) induced by tumor-derived growth factors. As we develop a better understanding of the cellular and molecular mechanisms of metastasis in head and neck squamous cell carcinoma, new therapies effective at preventing the development of secondary tumors can be realized ultimately increasing the patient's survival. PMID- 10859519 TI - The new imaging-based classification for describing the location of lymph nodes in the neck with particular regard to cervical lymph nodes in relation to cancer of the larynx. AB - For over five decades, the principle landmarks used in cervical nodal classification were clinical and defined either by palpation or found at the operative table. However during the past two decades, sectional imaging has consistently improved its quality and resolution and it has been shown that imaging can identify deep structures and adenopathy not amenable to palpation. Such disease can alter planned operative or radiation fields. In the April 1999 issue of the Archives of Otolaryngology-Head Neck Surgery, for the first time an imaging-based classification was published that gave precise anatomic landmarks for use in classifying metastatic cervical adenopathy. This classification was developed in consultation with head and neck surgeons so that the nodal levels classified by this imaging-based system would correspond closely with the nodal levels determined by utilizing the most commonly employed clinically-based classifications. This article describes this imaging-based classification and demonstrates its use with axial diagrams. PMID- 10859520 TI - Diagnostic procedures for detection of lymph node metastases in cancer of the larynx. AB - Squamous cell carcinoma is the most common malignant neoplasm of the larynx. One of the most important influences on prognosis is the presence of metastases to the cervical lymph nodes. Accurate determination of lymph node involvement is therefore a prerequisite for individualized therapy in patients with squamous cell carcinoma of the larynx. Clinical palpation of the neck is not very accurate and the role of imaging techniques such as ultrasound, ultrasound-guided fine needle aspiration cytology, color Doppler ultrasound, computed tomography, magnetic resonance imaging and positron emission tomography is being applied in order to improve upon the results of clinical investigation alone. According to our investigations and review of the literature, the accuracy of computed tomography scanning (84.9%) and magnetic resonance imaging (85%) was superior to palpation (69.7%) and ultrasound (72.7%). Ultrasound-guided fine needle aspiration cytology showed an accuracy of 89% and was in the same range with positron emission tomography (90.5%). PMID- 10859521 TI - The pathology of neck dissection in cancer of the larynx. AB - Cancer of the larynx is a common problem in a head and neck oncological surgical practice; as such, pathology departments supporting such surgical practices will examine cervical lymph node dissection specimens with some frequency. Issues to be settled among pathologists and surgeons include--How precise an anatomic dissection of the specimen is called for? What histological features of the specimen will be of most use to the clinicians who are devising a course of postoperative therapy for the patient? What sorts of methods are needed to identify the maximum number of micrometastases which may be lurking within the lymph nodes of the specimen? Is there a role for routine application of special techniques--such as immunohistochemistry or molecular biology--in the analysis of these specimens? While the answers to these questions are likely to vary somewhat from one center to another, patients are best served when these questions are discussed amongst the respective physicians before surgical procedures are undertaken, rather than after the fact. PMID- 10859522 TI - Classification and terminology of neck dissections. AB - With the proliferation of operations designed to treat cervical metastatic nodal disease, it has become ever apparent for the need to more clearly and precisely communicate the location of the metastatic cervical nodes and the specific surgery performed. To this end, this paper reviews the variety of operations and the resultant confusing terminology that has emerged over the past five decades. It is suggested that a simplified technology be used that specifically describes the nodal levels dissected, the relevant nonlymphatic structures removed, and those structures that are preserved. It is also suggested that the new imaging based nodal classification be used to standardize the definition of the nodal levels. It is hoped that this approach will eliminate many of the often confusing and nondescriptive terms and there by facilitate better inter-physician and inter institutional communication. PMID- 10859523 TI - Surgical treatment of the neck in cancer of the larynx. AB - Current concepts in management of the clinically negative and clinically positive neck in laryngeal cancer are reviewed. Occult disease in the neck not detected by physical and radiographic examination may also be difficult to identify on routine histologic examination. Immunohistochemistry or molecular analysis may detect metastatic involvement not apparent by light microscopy. The surgeon should be aware of the relatively high incidence of micrometastases in patients with laryngeal cancer to establish optimal treatment approaches. Elective treatment of the neck is recommended for supraglottic tumors staged T2 or higher, and glottic or subglottic tumors staged T3 or higher. The neck may be treated electively by either surgery or irradiation, but irradiation is best reserved for cases where that modality is employed for the primary tumor. Elective neck dissection provides important information for prognostic purposes and therapeutic decisions, by establishing the presence, number, location and nature of occult lymph node metastases. The selective lateral neck dissection (levels II, III and IV), unilateral or bilateral, is the procedure of choice for elective treatment. Paratracheal nodes (level VI) should be dissected in cases of advanced glottic and subglottic cancer. Complete radical or functional neck dissections are excessive in extent, as levels I and V are almost never involved. Sentinel lymph node biopsy may fail to detect tumor on frozen section examination or may not reveal 'skip' metastases. The clinically involved neck is usually treated by complete radical or functional neck dissection of levels I through V. Selective neck dissection has been employed successfully in selected cases, particularly for N1 or occasionally N2 nodal involvement. The selective neck dissection can be extended to include structures at risk. More advanced disease has been treated in this manner often in association with adjuvant chemotherapy and/or irradiation. While the benefit of adjuvant treatment is difficult to assess, it appears most useful in cases with extranodal spread of disease, a factor associated with the worst prognosis. PMID- 10859524 TI - Nonsurgical treatment of advanced metastatic cervical disease in cancer of the larynx. AB - Historically, patients with advanced cervical adenopathy (N2 or N3) have between a 20 and 30% chance of surviving their disease at 5 years from treatment. Despite attempts at more aggressive surgical resection, including resection and reconstruction of the carotid artery, patients with advanced cervical adenopathy remain at the highest risk for the development of local recurrences and distant metastases. This chapter will review the current limitations of surgical resectability for advanced neck disease, discuss the evolution of combined chemoradiation therapy for these patients, and finally present promising recent technological advances in radiation oncology which will have significant impact on the treatment of these patients. PMID- 10859525 TI - Bronchus-associated lymphoid tissue (BALT) is not present in the normal adult lung but in different diseases. AB - Bronchus-associated lymphoid tissue (BALT) was first described in the lungs of rabbits and differs greatly between species. It is part of the integrated mucosal immune system. This review clarifies its morphological definition and focuses on the situation in humans. The frequency of BALT at different ages, after chronic stimulation and in different diseases is described. In healthy humans, BALT can only be found in the lungs of children and adolescents. The role of BALT in lung transplantation and in the development of low-grade malignant lymphomas in the airways is also discussed. Furthermore, questions concerning the inducibility of BALT as an entry site for vaccines, and the regulation of its activity for future therapeutic interventions in pulmonary immune reactions are addressed. PMID- 10859526 TI - Microdissection of tissue sections: application to the molecular genetic characterisation of premalignant lesions. AB - The characterisation of the early molecular genetic events of tumor development depends on the selective procurement of histopathologically defined small cell populations from premalignant tissue. In order to obtain high-quality DNA, mRNA and proteins from these small tissue samples and even from single cells, tissue microdissection is one of the most useful techniques, becoming increasingly important for molecular pathologists. Using different microdissection techniques which allow the isolation of morphologically defined cell populations under direct visualisation, it is now feasible to study molecular genetic events that drive the multistep evolution in tumours. This review aims to present the current techniques of tissue microdissection and these techniques are discussed in the light of their ability to isolate premalignant cell populations in particular. Furthermore, we describe the subsequent application of several multiplex molecular analyses for characterising the microdissected premalignant cells. Applying these advanced techniques, alterations in the cellular DNA or the fluctuation of expressed genes that correlate with a particular stage of carcinogenesis can ultimately be compared within or between individual patients. Thus, these new technologies will have an enormous impact on molecular pathology with several diagnostic, prognostic and therapeutic implications. PMID- 10859527 TI - Intimal thickness and layering, and smooth muscle cell phenotypes in aorta of youth. AB - Proneness to the lesions of atherosclerosis varies along the length and circumferential topography of the aorta. Smooth muscle cells, in particular those of the 'modulated' synthetic phenotype which are able to proliferate and synthesize matrix proteins, are considered to play an important role in lesion progression. We report on a study of the aortic intima at a lesion-prone site from abdominal aorta and a lesion-resistant site from thoracic aorta in young humans to determine (1) whether the histologic structure and the smooth muscle cell composition show quantitative differences between lesion-prone and lesion resistant aortic sites; (2) whether there are gender differences, and (3) whether any differences increase in degree with increasing age in this young population. Material for this study was obtained as part of the NIH-funded multicenter study on Pathobiological Determinants of Atherosclerosis in Youth (PDAY) from autopsies of male and female subjects between the ages of 15 and 34, victims of unexpected sudden death, usually from trauma. The samples consisted of strips of abdominal and thoracic aorta, all derived from the same anatomical sites standardized in the PDAY studies. The thickness of total intima (TI) and its elastic hyperplastic (EH) layer was measured. Smooth muscle cells of all types (SMC) and separately those of the synthetic phenotype (SynSMC) were quantified in each site using immunohistochemical procedures in replicate sections of uniform thickness. The intima of the atherosclerotic lesion-prone dorsal half of the abdominal aorta (AD) shows significant differences from the lesion-resistant ventral half of thoracic aorta (TV) in that (1) its EH layer is significantly thicker; (2) its EH layer has a comparatively higher number of both total SMC and SynSMC, even when adjusted for intimal thickness, and (3) the age-related increase in thickness of both TI and EH layer of AD is much greater than that of TV. PMID- 10859528 TI - Different metabolism of hexose sugars and sucrose in wound fluid and in fibroblast cultures derived from granulation tissue, hypertrophic scar and keloid. AB - Viscose cellulose sponge implants in the rat and fibroblast cultures established from granulation tissue, hypertrophic scar, or keloid were treated with different concentrations of glucose, fructose, galactose, mannose, and sucrose. The concentrations of the above-mentioned sugars in wound fluid and cell culture medium were examined at the termination of experiments by liquid chromatography. Results showed that glucose was present in wound fluid in relatively low levels. In addition to glucose, only mannose was found in wound fluid. On the other hand, cell culture studies showed that virtually all the added sugars were found in cell culture medium. The most prominent exception was the decreased concentration of mannose in keloid fibroblast cultures. In addition, glucose concentration in culture medium of keloid fibroblasts was constantly very low except in mannose treated cultures where the consumption of glucose was dose-dependently decreased compared to increased mannose concentration. Similarly, increased concentrations of galactose and mannose resulted in dose-dependent lowered consumption of glucose in granulation tissue and hypertrophic scar fibroblasts. These findings suggest that the sugar metabolism may differ in various fibroblast cultures. Further, at least in our wound model, only glucose and mannose are present in wound fluid, and excess sugar is rapidly cleared from wound fluid. PMID- 10859529 TI - Numerical density of cardiomyocytes in chronic nitric oxide synthesis inhibition. AB - OBJECTIVE: The myocardial effect on the normalization of arterial blood pressure in animals with hypertension previously induced by nitric oxide synthesis (NOS) inhibition is still unknown. The purpose of this study was to estimate the numerical density of cardiomyocytes that is able to show cardiomyocyte loss as a consequence of NOS inhibition. METHODS: Sixty rats were divided into the following groups: control for 25 days (C25), control for 40 days (C40) control for 80 days (C80) and three other groups in which the rats received the NOS inhibitor N(G)-nitro-L-arginine-methyl-ester hydrochloride L-NAME; 50 mg/kg/day for 25 days (L25), 40 days (L40) and 40 days, respectively, the latter group having another 40 days of only water and food ad libitum (without L-NAME; L80 group). The detection of apoptotic cells was performed using a monoclonal antibody. RESULTS: In the L25, L40 and L80 groups, blood pressure was 74.5, 90.2 and 56.3% higher than the respective age-matched control rats, the myocardium had hypertrophy of the cardiomyocytes and scattered areas of fibrosis, and apoptosis occurred in isolated cells. Compared to the controls, the heart mass/body mass ratio was significantly greater in the L-NAME groups L25, L40 and L80, i.e. 31, 26 and 21%, respectively, the numerical density of cardiomyocytes in L-NAME rats was 32.7, 48.8 and 41.7% lower and the mean volume of cardiomyocytes was 33, 53 and 48% greater. CONCLUSION: The cardiomyocyte loss in NOS inhibition seems to be mainly due to necrosis, although apoptosis is also present. PMID- 10859530 TI - Changes in brain neurofilament and beta-tubulin proteins after cerebral hypoxia ischemia in rabbits. AB - Neurofilaments (NF) and tubulin are highly phosphorylated proteins that are important in neuronal structure and function. Changes in phosphorylation alter their antigenicity, and previous studies examining the effect of ischemia on these proteins failed to consider this factor. Using phosphate-independent antibodies and a quantitative immunoassay, we examined whether the amount of NF 68-kD (NF 68), NF 160-kD (NF 160), NF 200-kD (NF 200) and class III beta-tubulin proteins in the brain are reduced after cerebral injury. Rabbits were subjected to 8 min of hypoxia and then to 8 min of ischemia. After 4 h of reperfusion, NF 68 decreased from an overall group mean (+/- SEM) of 17.5+/-2.3 ng NF 68 microg/total protein in the noninjured controls (n = 8) to 12.9+/-1.2 ng/microg total protein in the hypoxic-ischemic group (n = 9). Conversely, NF 200 increased from 31.6+/-3.3 ng/microg in controls to 47.7+/-3.2 ng/microg. The amount of NF 160 and beta-tubulin was unchanged. The response of the NF proteins to brain injury is more complicated than described previously. Additional studies examining the regulation and metabolism of the NF are warranted, especially regarding the role of phosphorylation. PMID- 10859531 TI - Several selective protein kinase C inhibitors including procyanidins promote hair growth. AB - We have previously reported that procyanidin oligomers selectively promote growth of murine hair epithelial cells in vitro and stimulate anagen induction in vivo. We report here the possible relationship between the protein kinase C-inhibiting activity of procyanidins and their hair-growing activity. Of the procyanidins, procyanidin B-2 and procyanidin C-1, which selectively inhibit protein kinase C, intensively promote hair epithelial cell proliferation in vitro and stimulate anagen induction in vivo. On the other hand, procyanidins, which inhibit both protein kinase C and A, showed relatively low activity in in vitro and in vivo evaluations. We also found that calphostin C, which is a selective inhibitor of protein kinase C, possesses hair epithelial cell growth-promoting activity in vitro and anagen phase-inducing hair-growing activity in vivo. Other selective protein kinase C inhibitors, such as hexadecylphosphocholine, palmitoyl-DL carnitine chloride, and polymyxin B sulfate, also show marked anagen phase inducing hair-growing activity in vivo. Nonselective protein kinase inhibitors, such as staurosporine and K252a, inhibit the growth of hair epithelial cells. 1,2 Dioctanoyl-sn-glycerol, a protein kinase C activator, dose-dependently decreases the growth of hair epithelial cells. Forskolin, an adenylate cyclase activator, promotes hair epithelial cell growth and boosts the growth-promoting effect of procyanidin B-2. It is speculated that the hair-growing activity of procyanidins is related to their protein kinase C-inhibiting activity. PMID- 10859532 TI - Biomelanin antioxidants in cosmetics: assessment based on inhibition of lipid peroxidation. AB - Acute adverse effects of ultraviolet (UV) radiation in humans include sunburn, photosensitivity reactions and immunological suppression. Chronic exposure to UV light, particularly the UVB (290-320 nm) component of the UV radiation, and certain environmental chemicals increase the risk of nonmelanoma skin cancer and play a major role in cutaneous aging. The lipid peroxidation (LPO) of biomembranes, mediated by reactive oxygen species and free radicals, is one of the major causes of cellular damage induced by UV radiation and toxins. Antioxidants, such as vitamin E, vitamin C and melanins, are reactive oxygen and radical scavengers, thereby minimizing the light- and toxin-induced tissue destruction. We examined the influence of 8 biotechnically produced polyphenolic melanins on the LPO of microsomal membranes in comparison with alpha-tocopherol, ascorbate and synthetic melanin. All biomelanins showed better inhibition of peroxidative damage than synthetic melanin. Three of the 8 tested drugs inhibited the LPO at least as effectively as vitamin C and vitamin E. The combination of the most effective biomelanin with both vitamin C and vitamin E resulted in greater LPO inhibition than caused by each agent alone. Our data show that biomelanins are potent inhibitors of the peroxidative destruction of biomembranes, indicating that these compounds may be useful antioxidative agents in cosmetic preparations. PMID- 10859533 TI - Differential modulation of transforming growth factor-beta by betamethasone-17- valerate and isotretinoin: corticosteroid decreases and isotretinoin increases the level of transforming growth factor-beta in suction blister fluid. AB - Retinoids and glucocorticoids are known to have a potential to modulate the expression of transforming growth factor-beta (TGF-beta). We investigated the effect of oral isotretinoin (13-cis-retinoic acid) on the expression of two distinct isoforms of TGF-beta, TGF-beta1 and TGF-beta2, in suction blister fluid and serum in acne patients. We also investigated the effect of topical glucocorticoid (betamethasone-17-valerate) and age on suction blister fluid TGF beta1 in healthy volunteers. Six weeks of isotretinoin treatment caused a statistically significant 19% increase in suction blister fluid TGF-beta1. The suction blister fluid TGF-beta2 level remained below the sensitivity level of the immunoassay in many cases. Isotretinoin did not affect the serum TGF-beta1 or TGF beta2 level. Betamethasone-17-valerate pretreatment for 3 days twice a day caused a statistically significant 17% decrease in suction blister fluid TGF-beta1. The active form of TGF-beta1 represented 5% of the total TGF-beta1 in suction blister fluid. Our diffusion calculations suggest that all TGF-beta1 and TGF-beta2 detected in suction blister fluid have diffused from systemic circulation. The increase in suction blister fluid TGF-beta1 after isotretinoin treatment seems to be of local origin, while the decrease in suction blister fluid TGF-beta1 after glucocorticoid pretreatment seems to be due to glucocorticoid-induced vasoconstriction resulting in decreased diffusion of TGF-beta1 from the circulation. Modulation of local interstitial fluid TGF-beta1 concentration may be one mechanism by which isotretinoin and glucocorticoids mediate their effects in skin. PMID- 10859534 TI - Use of infrared attenuated total reflectance spectroscopy for the in vivo measurement of hydration level and silicone distribution in the stratum corneum following skin coverage by polymeric dressings. AB - The ability of wound dressings to hydrate the stratum corneum has been tested experimentally using attenuated total reflectance (ATR) infrared spectroscopy. A silicone gel dressing was able to hydrate the stratum corneum to the extent that its water content within the sampled volume was >60% after 5 h of contact compared with about 15% of normal exposed skin and about 30% for a highly permeable hydrophilic polyurethane dressing. An ATR method was devised to determine the presence and depth of penetration of silicone in the stratum corneum as a result of skin coverage by the polymeric dressing. PMID- 10859535 TI - Evaluation of dermal irritancy potential of benzanthrone-derived dye analogs: structure activity relationship. AB - The twelve structural analogs of benzanthrone-derived dyes of commercial use were screened for their dermal irritation potential response using the Draize occlusive patch test. The test dyes, dissolved in DMSO as vehicle, were topically applied on the skin of the male Druckery rats (160 +/- 10 g) according to the OECD protocol. The potential dermal hazard was assessed in terms of the primary cutaneous irritation (PCI) index and irritancy. Irritancy was evaluated according to the AFNOR scale. In terms of irritancy, the twelve benzanthrone dyes qualified as moderately irritant (3.0-5.0) according to the above scale. In decreasing order, PCI index of the various dyes was: Navy Blue R (4.5); Jade Green XBN (4.25); 16, 17-dihydroxydibenzanthrone (3.84); Black NB (3.75), Jade Green 2G (3. 75); 3-bromobenzanthrone (3.58); Brilliant Purple 4R (3.58); Olive D (3.50); Dark Blue 2R (3.41); Olive Green B (3.33); isodibenzanthrone (3.33), and benzanthrone (3.16). These results indicate that benzanthrone-derived dyes/dye intermediates caused dermal toxicity which appears to be influenced by the number of carbonyl and amino-anthraquinone groups as well as by the presence of functional groups like halogen, nitro, hydroxy and methoxy in the parent molecule, benzanthrone. PMID- 10859536 TI - Biophysical characteristics of healthy skin and nonlesional skin in atopic dermatitis: short-term effects of ultraviolet A and B irradiation. AB - The present study was designed to evaluate basic differences in surface structure and viscoelastic properties of nonatopic versus atopic skin and facultative acute changes following ultraviolet irradiation. Therefore, biophysical measurements by means of profilometry and cutometry were carried out on sun-protected unaffected gluteal skin areas in both groups before and 24 h after single UVA and UVB irradiations. The results indicate that the clinically unaffected skin of patients with atopic eczema differs from normal skin in terms of increased roughness parameters, but not concerning depth of furrows or viscoelastic properties (viscosity and biological elasticity, cutometrically calculated). Single UVA irradiation with 50 J/cm(2) induced neither measurable changes in the skin's surface structure nor in its viscoelastic properties in both groups after 24 h. However, irradiation with a single erythemogenic dose of 1 MED UVB was followed by a short-term significant increase in the depths of furrows and decrease in biological elasticity in normal and atopic skin, accompanied by an increase in viscosity in normal skin. PMID- 10859537 TI - The inherent capacitance of moisturising creams: A source of false positive results? AB - The capacitance is a frequently used measurement of skin hydration in vivo, therefore any potential for false capacitance results is of importance. This in vitro study is to investigate the capacities of moisturizing creams to cause false positive results. Adding different volumes of demineralised water and saline on dry clean plastic-backed tissue paper and measuring the capacitance values, the standard relation curve of capacitance value and volume of water was established. Four moisturizing creams, petrolatum, a protective cream and a gel were then applied to the same kind of tissue paper in constant volumes, and were measured for weights and capacitance values over a 1-hour period allowing the gradual evaporation of water and volatile elements. A positive linear relationship was found between volume of water and capacitance value. All the creams had inherent capacitance that correlated with their water content. The inherent capacitance of creams decreased over time and ultimately reached comparable low values. The decrease in capacitance correlated significantly with weight reduction, except for petrolatum, which showed negligible inherent capacitance and weight reduction. The rates of decrease in capacitance value versus cream weight were significantly different for different creams. The results assured that capacitance value correlates to water content, and the inherent capacitance of creams may contribute to false positive skin capacitance in vivo if measured before water and other volatile elements of creams evaporate from the skin surface. In addition, the inherent capacitance and its durability may be relevant characteristics for further comparative studies of moisturizers and their clinical efficacy. PMID- 10859538 TI - Effect of internal wool lipid liposomes on skin repair. AB - Intercellular lipids of the stratum corneum (SC) play a crucial role in keeping an optimal skin barrier function and in regulating the water-holding capacity. Recently, the internal lipids have been extracted from wool fibre. This lipid extract has a composition similar to that of the SC lipids. Two parameters were used to test the effect of topically applied internal wool lipids (IWL), structured as liposomes, on the water barrier functions of disturbed and intact skin: transepidermal water loss and skin capacitance. Liposomes made up of lipids that simulate the composition of the SC were also applied for comparison. The single application of the IWL liposomes on disturbed skin resulted in an accelerated recovery of water barrier functions. Daily application of these liposomes on intact skin for 5 days reinforced the skin barrier and increased the water-holding capacity. The repairing effect of the IWL enhances their suitability in the treatment, prevention and care of skin. PMID- 10859539 TI - Biannual meeting of the European Retinoid Research Group (ERRG). Strasbourg, September 26-30, 1999. PMID- 10859540 TI - Endothelin A receptor antagonist reduces renal vasoconstriction induced by mechanical manipulation of renal artery. AB - The purpose of this study was to make a model of renal vasoconstriction induced by mechanical manipulation of the renal artery and to investigate the influence of endothelin (ET) in renal vasoconstriction. Renal arteries of anesthetized rats were stretched with a 30-gram weight for 5 min, and released. Renal blood flow (RBF) was monitored with a laser flowmeter. To determine the role of ET, a selective ET(A) receptor antagonist, BQ-123 (0.1 and 1 mg/kg) was administered into the aorta. The manipulation was invariably followed by a flow reduction of about 57% after temporary hyperemia. RBF returned to the basal level 5 min after the release. BQ-123 partially, but significantly, inhibited the mechanical manipulation-induced reduction of RBF in a dose-dependent manner. These results suggest that endogeneous ET may play an important role in renal vasoconstriction induced by mechanical manipulation of the renal artery. PMID- 10859541 TI - No generalized skin phototoxicity after intravesical application of 5 aminolevulinic acid for fluorescence diagnosis of superficial bladder cancer. AB - INTRODUCTION: A prospective monocenter open study was carried out to evaluate if generalized phototoxic skin reactions occur after intravesical application of 5 aminolevulinic acid (ALA) for fluorescence diagnosis of superficial bladder carcinomas. PATIENTS AND METHODS: On 21 patients, skin phototoxicity was determined prior to as well as 4, 8 and 28 h after intravesical instillation of a 3% ALA solution by exposing small skin areas to a progressively graded series of defined UVA-light doses (n = 9; 5-80 J/cm(2)). RESULTS: Prior to ALA instillation, erythema or pigmentation as signs of cutaneous phototoxicity occurred at an UVA-light dose of 28 +/- 1.5 J/cm(2) (mean +/- SEM). A reduction of the minimal phototoxic dose (MPD) was not detected 4 (28 +/- 1.9 J/cm(2)), 8 (28 +/- 1.6 J/cm(2)) and 28 h (28 +/- 1.5 J/cm(2)) after ALA instillation. Consequently phototoxic reactions were not observed. CONCLUSIONS: A reduction of MPD for UVA was not detected. Therefore, it is not necessary to protect the skin of patients from ambient or daylight after intravesical instillation of ALA for fluorescence diagnosis. PMID- 10859542 TI - Significance of carbohydrate antigen sialyl-Lewis X, sialyl-Lewis A, and possible unknown ligands to adhesion of human urothelial cancer cells to activated endothelium. AB - OBJECTIVE: To assess the contribution of the carbohydrate antigens, sialyl-Lewis X (sLe(x)) and sialyl-Lewis A (sLe(a)), which are known to be ligands for E selectin, to the adhesion between human urothelial cancer cells and cytokine activated human endothelial cells. MATERIALS AND METHODS: We studied the expression of sLe(x) and sLe(a) antigens of three bladder cancer cell lines (JTC 30, JTC 32, and T24) by flow cytometry and the adherence to interleukin 1beta activated human umbilical vein endothelial cells (HUVEC). RESULTS: JTC 30 and JTC 32 cells expressed both sLe(x) and sLe(a) antigens, and showed adhesion to activated HUVEC, which was completely abolished by anti-E-selectin antibody. T24 cells expressed neither sLe(x) nor sLe(a) antigen, and did not adhere to activated HUVEC. Each of anti-sLe(a) or anti-sLe(x) antibody partially blocked the attachment of JTC 30 cells to activated HUVEC, and combination of these antibodies almost completely blocked the adhesion. The combination of antibodies did not significantly influence the adhesion of JTC 32 cells. CONCLUSION: These results indicate that both sLe(a) and sLe(x) carbohydrate antigens are involved in E-selectin-mediated adhesion of some urothelial cancers, and that there might be unknown ligands for E-selectin on urothelial cancer cells. PMID- 10859543 TI - A germline homozygote deletion of the glutathione-S-transferase Mu1 gene predisposes to bladder cancer. AB - INTRODUCTION AND OBJECTIVES: Numerous studies have shown smoking and specific occupational exposures to be risk factors for bladder cancer. The risk of bladder cancer may be modified by the activity of carcinogen metabolizing enzymes. The glutathione-S-transferase Mu1 enzyme (GSTM1) detoxifies arylepoxides which are formed after exposure to certain polycyclic aromatic hydrocarbons and possibly aromatic amines. Approximately 40% of Caucasians lack GSTM1 activity due to a homozygous deletion of the GSTM1 locus on chromosome 1p13 (GSTM1 0/0 genotype). The aim of this study was to evaluate the combined effect of smoking and GSTM1 genotype on the risk of bladder cancer. MATERIALS AND METHODS: Sixty-one patients with transitional cell carcinoma of the bladder and 69 controls matched for age and sex were enrolled from the outpatient clinic. Lifestyle information was collected with a standardized questionnaire. DNA was extracted from white blood cells. The GSTM1 genotype was determined by a PCR-based method. RESULTS: 92% of the 61 patients had a history of smoking compared with 81% of the controls. There was a significant dose-response relationship for pack-years of smoking (trend test: p = 0.003). The proportion of GSTM1 0/0 genotype among patients was 62% compared with 43% among controls (odds ratio = 2.1; 95% CI 1.1-4. 3). The expected interaction between smoking and GSTM1 genotype was not observed. CONCLUSIONS: This study confirms the findings that a germline homozygous deletion of the GSTM1 gene predisposes to bladder cancer. An interaction with smoking was not found. PMID- 10859544 TI - No change with age in semen volume, sperm count and sperm motility in individual men consulting an infertility clinic. AB - In the human male, Leydig cell function of the testis declines slowly but significantly with age. A similar decline in the spermatogenic activity, however, is not well proven by consistent data. Cross-section studies quoted in the literature mostly describe only minor changes in different sperm parameters with age. In our database we identified 253 men who had attended our department since 1989 for subsequent semen analyses at intervals of more than 1 year. The semen analyses were performed according to the guidelines of the WHO with the exception that motility parameters were measured by CASA. The mean interval between the two analyses was 967 days with a range of 341-2,731 days. None of the parameters evaluated showed a significant increase or decrease with time. The mean seminal volume was identical in the two analyses. Mean sperm count increased during the observation period by 12. 10(6) spermatozoa/year. In 177 patients an increase and in 73 patients a decrease occurred. Also a corresponding increase in total sperm count was observed. Sperm motility increased in 123 patients, and it decreased in 123 patients. Only slight differences in the increase or decrease of the two parameters between the groups of patients with and without a history of genital diseases and/or varicocele were found. The results of our study do not support the suggestion that the sperm parameters decrease with age. PMID- 10859545 TI - Relation of sexual dysfunction to hormone levels, diseases and drugs used in andrological patients. AB - In 169 patients visiting our department complaining of sexual dysfunction, the medical history was taken using a semistructured interview. A clinical investigation and a hormone analysis were added. The age of patients, hormone values, and items of the interview were collected into a common database. The items were categorized as either dichotomous (yes/no) or ordinal. Statistical analysis was performed using regression analysis with the aim to generate hypotheses of relations. An increase of FSH levels and a decrease of testosterone levels with age occurred. None of the relations of hormone levels or diseases to symptoms of sexual dysfunction produced odds ratios (OR) statistically significant different from 1. However, the risk of having a reduced libido and reduced morning erections was lower in psychoneurological diseases, the risk of reduced arousal and libido was lower in men with diabetes mellitus, but the risk of reduced morning erections was higher in these men. The testosterone levels were not associated with the risk of having reduced penile rigidity, duration of erection, arousal and sexual libido, reduced morning erections and the ability to masturbate. Smoking was not associated with reduced arousal, libido and morning erections. However, a significant increase of testosterone levels with number of cigarettes used was observed. We conclude that sexual dysfunction in patients visiting an andrological department for diagnosis and treatment is mostly not associated to any single evaluable factor. PMID- 10859546 TI - Effect of N-(biphenylyl-methyl)imidazole, a type 1 angiotensin II receptor inhibitor, on the contractile function of the rat corpus cavernosum. AB - The effect of N-(biphenylyl-methyl)imidazole, losartan potassium, a newly developed antihypertensive type 1 angiotensin II receptor antagonist on the rat erectile function, was studied. Sexually active 9-week-old male spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats were given losartan 60 mg/kg intraperitoneal injections. Mean blood pressure (MBP) dropped significantly in both SHR and WKY rats (for SHR: from 140 +/- 8 to 114 +/- 5 mm Hg, p < 0.05, n = 8; for WKY: from 113 +/- 7 to 79 +/- 9 mm Hg, p < 0.05, n = 8). On the contrary, the intracavernous pressure (ICP) of SHR and WKY rats did not differ significantly from that of the corresponding controls receiving saline injections (p > 0.05, n = 8 for each group). For the chronic study, the rats were fed with losartan 30 mg/kg/day for 30 days. MBP decreased significantly in SHR but not in WKY rats (for SHR: from 137 +/- 7 to 113 +/- 5 mm Hg, p < 0.05, n = 8; for WKY: from 110 +/- 6 to 107 +/- 5 mm Hg, p > 0.05, n = 8). The ICP of the losartan-treated rats was not significantly different from that of control rats (p > 0.05, n = 8 for each group). In contrast, WKY rats receiving guanethidine 1 mg/kg/day for 30 days showed significantly decreased ICP. Angiotensin II (10(-9) 10(-5) M) and losartan (10(-9)- 10(-5) M) did not induce significant contractile responses of the cavernosal strip when tested in vitro. On the other hand, methoxamine 10(-4) M induced good contractile responses. In conclusion, the present study demonstrated that angiotensin II did not cause significant change in the contractile status of rat corpus cavernosum. Correspondingly, the type 1 angiotensin II inhibitor effectively lowered blood pressure but did not affect cavernosal contractile function, thus is useful clinically in the treatment of hypertensive disorders without significant detrimental effects on male sexual function. PMID- 10859547 TI - Assessment of a three-dimensional operating system with skill tests in a pelvic trainer. AB - OBJECTIVES: To compare the performance of laparoscopic skill assisted by a traditional two-dimensional (2D) and a three-dimensional (3D) endoscopic video system in a pelvic trainer. MATERIALS AND METHODS: The 3D imaging system (DeepVision((R)), Automated Medical Products Corp.) consists of a traditional single lens optic laparoscope, a light source, an endoscopic camera (Stryker), a DeepVision processor and a DeepVision monitor. The 2D images could be obtained with the same system without turning on the DeepVision processor. Thirty-four medical personnel with no laparoscopic surgical experience were enrolled to perform two skill tests, the object-pick-up and spatial orientation test in a trainer box. They were randomly divided into two groups, one group performed the test under 2D conditions first and 3D later, and another group performed the test under 3D conditions first and 2D later. The duration needed to complete the skill tests was recorded and the differences on performance time under 2D and 3D conditions were calculated for each participant. Two-way ANOVA was used to analyze the statistic difference on the performance time in two conditions. RESULTS: The duration needed to complete the initial skill tests was similar among 2D and 3D conditions. For both tests, the average performance time decreased significantly for the second attempt regardless of 2D or 3D conditions. Statistic analysis disclosed significant difference for learning factor (p < 0.001 for object-pick-up test and p < 0.01 for spatial orientation test), but no significant difference between 2D and 3D conditions (p = 0.276 for object-pick-up test and p = 0.327 for spatial orientation test). CONCLUSION: A significant decrease of the performance time at the second attempt reflected the importance of a learning process in laparoscopic surgery. It appears that no significant benefits were obtained by this 3D operating system for surgeons without laparoscopic surgical experience. PMID- 10859548 TI - Identification of the origin of a vesical mass occurring after cadaveric renal transplantation using short tandem repeat markers. AB - We report a case of polypoid cystitis in a 54-year-old female occurring 4 years after cadaveric kidney transplantation. Endoscopic exploration revealed a polypoid tumor near the stoma opened for the transplanted ureter. The diagnosis of polypoid cystitis was confirmed histopathologically. Genotyping of cells from the tumor with polymorphic short tandem repeat (STR) and amelogenin loci revealed that the tumor contained alleles from both the donor and recipient. Molecular genetic analysis provided strong evidence that the tumor cells arose from the donor tissue. PMID- 10859549 TI - Eosinophilic cystitis in a case presenting with an initial diagnosis of invasive bladder tumor. AB - Eosinophilic cystitis is a rare condition of the bladder that presents with hematuria, dysuria and suprapubic tenderness. A case of eosinophilic cystitis presenting as an invasive bladder tumor is reported. PMID- 10859550 TI - Emphysematous cystitis following brain contusion. AB - Emphysematous cystitis is a rare disorder most commonly seen in patients with urinary tract infection and diabetes mellitus. We present a nondiabetic case of a 46-year-old woman with this entity following brain contusion. PMID- 10859551 TI - Intravesical migration of AMS 800 artificial urinary sphincter and stone formation in a patient who underwent radical prostatectomy. AB - A patient who underwent placement of AMS 800 urinary sphincter for incontinence after salvage prostatectomy after radiation therapy failure, experienced urethral erosion of the cuff after 54 months. He was treated with cuff removal and prosthesis deactivation. Thirteen months later, a large bladder stone was removed transvesically, and it contained the reservoir. The remaining sphincter components were also removed. Patients with previous radiotherapy are particularly at risk for sphincter erosion, but intravesical displacement of the reservoir is a very uncommon complication. PMID- 10859552 TI - Nephron-sparing surgery in a case of giant renal hydatid cyst. AB - We report a case of a centrally located giant renal hydatid cyst managed successfully by excision of the cyst alone and preserving the renal parenchyma after clamping the main renal artery. Follow-up IVP showed good function. PMID- 10859553 TI - Localized fibrosis of the corpus cavernosum: an example of fibrosis arising from the vascular smooth muscle cells. Report of a case with histogenetic considerations. AB - We report a rare case of a localized fibrosis of the corpus cavernosum (CC) presenting as a nodular mass in the dorsal region of the penis without abnormal curvature of the organ. Histological and immunohistochemical examination revealed a fibrotic process, involving CC but not tunica albuginea, arising from the smooth muscle cells of the vascular channels. Two different morphological phases were identified in these vascular lesions: proliferative and residual phase. The proliferative phase was characterized by hyperplasia and moderate fibrous thickness of the smooth muscle layer of the vascular channels of CC, resulting with the formation of concentric perivascular cuffs and nodules. The residual phase was characterized by a progressive nodular fibrotic transformation with narrowing of the original vascular channels and replacement of the normal CC structures. The cells of this phase exhibited an immunophenotype of both fibroblasts (vimentin+; alpha-smooth muscle actin-) and myofibroblasts (vimentin+; alpha-smooth muscle actin+). The clinicopathological features of the lesion and histogenetic considerations are discussed. PMID- 10859554 TI - Sonographic detection of a foreign body in the urethra and urinary bladder. AB - Reported herein is a 41-year-old male who presented for ultrasound due to two episodes of lower urinary tract infections within a period of 5 months. The sonographic examination revealed an elongated foreign body in the urethra extending into the urinary bladder. Self-insertion of foreign bodies into the urethra is usually done for erotic stimulation. However, foreign bodies can be inserted by children due to curiosity and by mentally retarded people, patients with psychiatric disorders as well as by intoxicated patients and in confusional states. Due to embarrassment, the patients seek medical help only when they are symptomatic and hence some of the foreign bodies are removed only several months after insertion. Detection might be either by plain abdominal films when the foreign bodies are radiopaque or by the use of contrast media. In the case presented by us, this was done by sonography. Endoscopic removal of these foreign bodies is considered the treatment of choice. Recurrent or chronic unexplained urinary tract infections should raise a high index of suspicion to the possible existence of a foreign body in the urethra and/or urinary bladder. PMID- 10859555 TI - Patent and be damned: the new slogan for human genetics? PMID- 10859556 TI - Bone-marrow analysis predicts breast-cancer recurrence. PMID- 10859557 TI - Telomerase: the search for a universal cancer vaccine. PMID- 10859558 TI - Structural genomics. PMID- 10859559 TI - Plasma oxysterols: reliable indicators of heart disease? PMID- 10859560 TI - Home HIV testing - a possibility? PMID- 10859561 TI - Rapid update PMID- 10859562 TI - The role of nonhuman primate models in AIDS vaccine development. AB - Although animal models have been useful in guiding vaccine development, HIV/AIDS models have not yielded a clear correlate of immunity nor given consistent results on the potential efficacy of various vaccine approaches. Further development and improved uniformity in the use of animal models would maximize their potential to meet the urgent worldwide need for a safe and effective HIV vaccine. PMID- 10859563 TI - Mass spectrometry for genotyping: an emerging tool for molecular medicine. AB - Recent technological innovations have made proteins and nucleic acids accessible to mass spectrometric analysis. As a result of their inherently high specificity, accuracy and throughput, there is considerable interest in developing mass spectrometric methods for genotype analysis in clinical diagnostic and research applications. This review outlines some of the most promising genotyping methods developed using electrospray and matrix-assisted laser-desorption-ionization mass spectrometry. PMID- 10859564 TI - Lissencephaly and subcortical band heterotopia: molecular basis and diagnosis. AB - Magnetic resonance imaging is now used routinely in the evaluation of developmental and neurological disorders and provides exquisite images of the living human brain. Consequently, it is evident that cortical malformations are more common than previously thought. Among the most severe is classical lissencephaly, in which the cortex lacks the complex folding that characterizes the normal human brain. Lissencephaly includes agyria and pachygyria, and merges with subcortical band heterotopia. Current molecular genetic techniques combined with the identification of affected patients have enabled the detection of two of the genes responsible: LIS1 (PAFAH1B1) on chromosome 17 and DCX (doublecortin) on the X chromosome. This review highlights the discovery of these genes and discusses the advances made in understanding the molecular basis of cortical development and improvements in diagnosis and genetic counseling. PMID- 10859565 TI - Gene therapy for peripheral arterial disease. AB - Our understanding of the molecular biology of vascular disease is rapidly expanding, and this scientific growth has brought with it new opportunities for therapeutic intervention at the molecular and genetic levels. Although our tools for genetic manipulation in vivo and our knowledge of potential molecular targets are still crude and incomplete, the early application of these concepts to clinical problems is already underway, both in the pre-clinical and clinical arenas. The treatment of peripheral vascular disease, although greatly improved over recent decades by surgical and minimally-invasive techniques, remains limited by vascular proliferative lesions and by our inability to modulate the progression of native disease. This review explores some of the evolving concepts of therapeutic gene manipulation and their initial application in the peripheral circulation. PMID- 10859567 TI - Evolution and risky decisions. PMID- 10859566 TI - Prions, peptides and protein misfolding. PMID- 10859568 TI - Theory of mind and mirroring neurons. PMID- 10859569 TI - Reply to Schulkin. PMID- 10859570 TI - The evolution of speech: a comparative review. AB - The evolution of speech can be studied independently of the evolution of language, with the advantage that most aspects of speech acoustics, physiology and neural control are shared with animals, and thus open to empirical investigation. At least two changes were necessary prerequisites for modern human speech abilities: (1) modification of vocal tract morphology, and (2) development of vocal imitative ability. Despite an extensive literature, attempts to pinpoint the timing of these changes using fossil data have proven inconclusive. However, recent comparative data from nonhuman primates have shed light on the ancestral use of formants (a crucial cue in human speech) to identify individuals and gauge body size. Second, comparative analysis of the diverse vertebrates that have evolved vocal imitation (humans, cetaceans, seals and birds) provides several distinct, testable hypotheses about the adaptive function of vocal mimicry. These developments suggest that, for understanding the evolution of speech, comparative analysis of living species provides a viable alternative to fossil data. However, the neural basis for vocal mimicry and for mimesis in general remains unknown. PMID- 10859571 TI - Social perception from visual cues: role of the STS region. AB - Social perception refers to initial stages in the processing of information that culminates in the accurate analysis of the dispositions and intentions of other individuals. Single-cell recordings in monkeys, and neurophysiological and neuroimaging studies in humans, reveal that cerebral cortex in and near the superior temporal sulcus (STS) region is an important component of this perceptual system. In monkeys and humans, the STS region is activated by movements of the eyes, mouth, hands and body, suggesting that it is involved in analysis of biological motion. However, it is also activated by static images of the face and body, suggesting that it is sensitive to implied motion and more generally to stimuli that signal the actions of another individual. Subsequent analysis of socially relevant stimuli is carried out in the amygdala and orbitofrontal cortex, which supports a three-structure model proposed by Brothers. The homology of human and monkey areas involved in social perception, and the functional interrelationships between the STS region and the ventral face area, are unresolved issues. PMID- 10859572 TI - Aspects of body self-calibration. AB - The representation of body orientation and configuration is dependent on multiple sources of afferent and efferent information about ongoing and intended patterns of movement and posture. Under normal terrestrial conditions, we feel virtually weightless and we do not perceive the actual forces associated with movement and support of our body. It is during exposure to unusual forces and patterns of sensory feedback during locomotion that computations and mechanisms underlying the ongoing calibration of our body dimensions and movements are revealed. This review discusses the normal mechanisms of our position sense and calibration of our kinaesthetic, visual and auditory sensory systems, and then explores the adaptations that take place to transient Coriolis forces generated during passive body rotation. The latter are very rapid adaptations that allow body movements to become accurate again, even in the absence of visual feedback. Muscle spindle activity interpreted in relation to motor commands and internally modeled reafference is an important component in permitting this adaptation. During voluntary rotary movements of the body, the central nervous system automatically compensates for the Coriolis forces generated by limb movements. This allows accurate control to be maintained without our perceiving the forces generated. PMID- 10859573 TI - Gonadal hormones affect neuronal vulnerability to excitotoxin-induced degeneration. AB - The role of endogenous gonadal secretions in neuroprotection has been assessed in a model of hippocampal degeneration induced by the systemic administration of kainic acid to adult male and female rats. A low dose of kainic acid (7 mg/Kg b.w.) induced a significant loss of hilar dentate neurons in castrated males and did not affect hilar neurons in intact males. The effect of kainic acid on hilar neurons in female rats was different depending on the day of the estrous cycle in which the neurotoxin was administered; while no significant effect of kainic acid was observed when it was injected in the morning of estrus, there was a significant loss of hilar neurons when it was injected in the morning of proestrus as well as when it was injected into ovariectomized rats. Estradiol or estradiol plus progesterone prevented hilar neuronal loss when injected simultaneously with kainic acid in ovariectomized rat. Progesterone by itself did not prevent neuronal loss induced by kainic acid and estogen was only effective when it was injected either 24 h before or simultaneously with kainic acid and not when it was injected 24 h after the administration of the toxin. These findings indicate that endogenous gonadal hormones protect hippocampal hilar neurons from excitotoxic degeneration. In addition, the timing of exposure to ovarian hormones and the natural fluctuation of ovarian hormones during the estrous cycle may influence the vulnerability of hilar neurons to excitotoxicity. These findings are relevant to possible modifications in neurodegenerative risk in humans as endogenous levels of gonadal hormones change during the menstrual cycle and during aging. PMID- 10859574 TI - Immunogold study of regional differences in the distribution of glucose transporter (GLUT-1) in mouse brain associated with physiological and accelerated aging and scrapie infection. AB - Distribution of glucose transporter (GLUT-1) in brain microvascular endothelia, representing the anatomic site of the blood-brain barrier (BBB), was studied in adult, physiologically aged, senescence-accelerated prone (SAMP8) and in scrapie infected mice. Sections of tissue samples obtained from four brain regions (cerebral cortex, hippocampus, cerebellum, and olfactory bulb) and embedded in Lowicryl K4M were exposed to anti-GLUT-1 antiserum followed by gold-labeled secondary antibody. Labelling density was recorded over luminal and abluminal plasma membranes of the microvascular endothelial cells. We found that the density of immunosignals for GLUT-1 in the cerebral cortex showed a tendency toward insignificant diminution according to the following gradation-adult > SAMP8 > scrapie > aged mice-whereas in the hippocampus, this gradation was slightly different: adult > aged > scrapie > SAMP8 mice. In the cerebellum, immunolabelling was insignificantly diminished in aged mice, whereas it was significantly decreased in scrapie-infected and SAMP8 mice. The intensity of labelling of the vascular endothelium in the olfactory bulb was significantly lower than that in other brain regions, showing a slight decrease in the following sequence: adult > aged > scrapie > SAMP8 mice. These findings suggest that the process of aging as well as of related neurodegenerative disease affects unequally the distribution of GLUT-1 in the vasculature of different brain regions. PMID- 10859575 TI - Optic nerve regeneration after intravitreal peripheral nerve implants: trajectories of axons regrowing through the optic chiasm into the optic tracts. AB - We have studied axon regeneration through the optic chiasm of adult rats 30 days after prechiasmatic intracranial optic nerve crush and serial intravitreal sciatic nerve grafting on day 0 and 14 post-lesion. The experiments comprised three groups of treated rats and three groups of controls. All treated animals received intravitreal grafts either into the left eye after both left sided (unilateral) and bilateral optic nerve transection, or into both eyes after bilateral optic nerve transection. Control eyes were all sham grafted on day 0 and 14 post-lesion, and the optic nerves either unlesioned, or crushed unilaterally or bilaterally. No regeneration through the chiasm was seen in any of the lesioned control optic nerves. In all experimental groups, large numbers of axons regenerated across the optic nerve lesions ipsilateral to the grafted eyes, traversed the short distal segment of the optic nerve and invaded the chiasm without deflection. Regeneration was correlated with the absence of the mesodermal components in the scar. In all cases, axon regrowth through the chiasm appeared to establish a major crossed and a minor uncrossed projection into both optic tracts, with some aberrant growth into the contralateral optic nerve. Axons preferentially regenerated within the degenerating trajectories from their own eye, through fragmented myelin and axonal debris, and reactive astrocytes, oligodendrocytes, microglia and macrophages. In bilaterally lesioned animals, no regeneration was detected in the optic nerve of the unimplanted eye. Although astrocytes became reactive and their processes proliferated, the architecture of their intrafascicular processes was little perturbed after optic nerve transection within either the distal optic nerve segment or the chiasm. The re establishment of a comparatively normal pattern of passage through the chiasm by regenerating axons in the adult might therefore be organised by this relatively immutable scaffold of astrocyte processes. Binocular interactions between regenerating axons from both nerves (after bilateral optic nerve transection and intravitreal grafting), and between regenerating axons and the intact transchiasmatic projections from the unlesioned eye (after unilateral optic nerve lesions and after ipsilateral grafting) may not be important in establishing the divergent trajectories, since regenerating axons behave similarly in the presence and absence of an intact projection from the other eye. PMID- 10859576 TI - Structural basis of sympathetic-sensory coupling in rat and human dorsal root ganglia following peripheral nerve injury. AB - Tyrosine hydroxylase immunocytochemistry was used to reveal the sympathetic postganglionic axons that sprout to form basket-like skeins around the somata of some primary sensory neurons in dorsal root ganglia (DRGs) following sciatic nerve injury. Ultrastructural observations in rats revealed that these sprouts grow on the surface of glial lamellae that form on the neurons. Sciatic nerve injury triggers glial cell proliferation in the DRG, and the formation of multilamellar pericellular onion bulb sheaths, primarily around large diameter DRG neurons. We infer that these glia participate in the sprouting process by releasing neurotrophins and expressing growth supportive cell surface molecules. Many DRG cell somata, and their axons in intact nerves and nerve end neuromas, express alpha2A adrenoreceptors intracytoplasmically and on their membrane surface. However, sympathetic axons never make direct contacts with the soma membrane. The functional coupling known to occur between sympathetic efferents and DRG neurons must therefore be mediated by the diffusion of neurotransmitter molecules in the extracellular space. Sympathetic basket-skeins were observed in DRGs removed from human neuropathic pain patients, but the possibility of a functional relation between these structures and sensory symptoms remains speculative. PMID- 10859577 TI - Overexpression of rapsyn inhibits agrin-induced acetylcholine receptor clustering in muscle cells. AB - Rapsyn is a protein on the cytoplasmic face of the postsynaptic membrane of skeletal muscle that is essential for clustering acetylcholine receptors (AChR). Here we show that transfection of rapsyn cDNA can restore AChR clustering function to muscle cells cultured from rapsyn deficient (KORAP) mice. KORAP myotubes displayed no AChR aggregates before or after treatment with neural agrin. After transfection with rapsyn expression plasmid, some KORAP myotubes expressed rapsyn at physiological levels. These formed large AChR-rapsyn clusters in response to agrin, just like wild-type myotubes. KORAP myotubes that overexpressed rapsyn formed only scattered AChR-rapsyn microaggregates, irrespective of agrin treatment. KORAP cells were then transfected with mutant forms of rapsyn. A deletion mutant lacking residues 16-254 formed rapsyn microaggregates, but failed to aggregate AChRs. Substitution mutation to the C terminal serine phosphorylation site of rapsyn (M43(D405,D406)) did not impair the response to agrin, showing that differential phosphorylation of this site is unlikely to mediate agrin-induced clustering. The results indicate that rapsyn expression is essential for agrin-induced AChR clustering but that its overexpression inhibits this pathway. The approach of using rapsyn-deficient muscle cells opens the way for defining the role of rapsyn in agrin-induced AChR clustering. PMID- 10859579 TI - Community impact of anticoagulation services: rationale and design of the Managing Anticoagulation Services Trial (MAST). AB - We describe the design of the Managing Anti-coagulation Services Trial (MAST), a practice-improvement trial testing whether anticoagulation services are a preferred method of managing anticoagulation for stroke prevention among patients with atrial fibrillation. Most randomized trials within the health care environment are designed as efficacy studies to determine what works under ideal conditions or ideal clinical practice. In contrast, effectiveness trials seek to generalize the results of efficacy studies by determining what works under more typical practice conditions. Practice-improvement trials are effectiveness trials that examine the management of a clinical problem in the context in which care is usually given. Noteworthy features of the MAST include defining the intervention in functional terms and collaboration with managed care organizations. PMID- 10859580 TI - Anticoagulation clinics and patient self-testing for patients on chronic warfarin therapy: A cost-effectiveness analysis. AB - This study was intended to evaluate the cost-effectiveness of anticoagulation clinic care and self-testing for the management of patients on chronic warfarin therapy. Using a 5-year Markov model, we evaluated the health and economic outcomes associated with each of three different anticoagulation management approaches: (1) usual care, (2) anticoagulation clinic testing with a capillary monitor, and (3) patient self-testing with a capillary monitor. Data available in the published literature and data from a large health system were used to develop model assumptions. Model results indicate that over a 5-year period, compared with usual care, anticoagulation clinic testing results in a total of 1.7 fewer thromboembolic events and 2.0 less hemorrhagic events per 100 patients. Another 4.0 thromboembolic events and 0.8 hemorrhagic events are avoided with patient self-testing compared with anticoagulation clinic testing. In addition to the health advantages of these strategies, both also have cost advantages. When the costs incurred by provider organizations and patients are considered, patient self-testing is the most cost-effective alternative, resulting in an overall cost saving. PMID- 10859581 TI - Low molecular weight heparin in the treatment of acute deep vein thrombosis and pulmonary embolism: A paradigm change in care. PMID- 10859582 TI - Heparin-induced thrombocytopenia and its treatment. PMID- 10859583 TI - Medicare coverage policies: A macro and micro analysis. PMID- 10859584 TI - INR self-management following mechanical heart valve replacement. AB - INR self-management can reduce severe thromboembolic and hemorrhagic complications following mechanical heart valve replacement. Beginning anticoagulation therapy immediately in the postoperative period further reduces anticoagulant-induced complications. Data were collected from the first 600 surviving patients (from a total study sample of 1200 patients) who completed follow-up of at least 2 years. Patients were randomly divided into a self management group and a control group. INR self-management reduced severe hemorrhagic and thromboembolic complications (P=0.018). Nearly 80% of INR values recorded by patients themselves, regardless of educational level, were within the target therapeutic range of INR 2.5-4.5, compared with 62% of INR values monitored by family practitioners. Only 8.3% of patients trained in self management immediately after surgery were unable to continue with INR self management. The results differed slightly between patient groups with different levels of education. We conclude that all patients for whom anticoagulation is indicated are candidates for INR self-management regardless of education level. PMID- 10859585 TI - Anticoagulation therapy for atrial fibrillation and coronary disease. PMID- 10859586 TI - Workshop: Implementation of home treatment programs for deep vein thrombosis with low molecular weight heparin. PMID- 10859587 TI - Workshop: Developing a business plan for an anticoagulation management service. PMID- 10859588 TI - Workshop: Internet delivery of an anticoagulation therapy management certificate program. PMID- 10859589 TI - Workshop: Anticoagulation disease state management utilizing the Internet. PMID- 10859591 TI - Purinergic responses in HT29 colonic epithelial cells are mediated by G protein alpha -subunits. AB - Using Fura-2 to measure changes in intracellular calcium ([Ca(2+)](i)), we show that P(2U)receptors in HT29 cells trigger an increase in [Ca(2+)](i)by pertussis toxin-insensitive G proteins. We then use replication-deficient adenoviruses expressing wild-type and dominant negative mutants of G(alpha q)and G(alpha i2), antisense directed against G(alpha q)or the C-terminal fragment of beta adrenergic receptor kinase (beta ARK-CT) to identify these G proteins. We find the [Ca(2+)](i)response to UTP is not affected by increased expression of the wild-type G(alpha q), wild-type G(alpha i2)or beta ARK-CT, while it is blocked by over-expression of dominant negative G(alpha q). The timecourse of the UTP response is, however, altered by wild-type G(alpha q)and is only weakly inhibited by antisense G(alpha q). This suggests that the P(2U)response is mediated, at least partially, by a G protein distinct from G(alpha q). In contrast, the M(3)muscarinic response is inhibited by over-expression of antisense against G(alpha q), or over-expression of beta ARK-CT, a finding in agreement with our previous observation that the muscarinic response in HT29 cells is mediated by the beta gamma-subunits of G(q). We also find that P(2U)and M(3)receptors do not control identical Ca(2+)stores, suggesting that differential activation of G proteins can lead to Ca(2+)release from distinct stores. PMID- 10859592 TI - Differential modulation of inositol 1,4,5-trisphosphate receptor type 1 and type 3 by ATP. AB - Binding of ATP to the inositol 1,4,5-trisphosphate receptor (IP(3)R) results in a more pronounced Ca(2+)release in the presence of inositol 1,4,5-trisphosphate (IP(3)). Two recently published studies demonstrated a different ATP sensitivity of IP(3)-induced Ca(2+)release in cell types expressing different IP(3)R isoforms. Cell types expressing mainly IP(3)R3 were less sensitive to ATP than cell types expressing mainly IP(3)R1 (Missiaen L, Parys JB, Sienaert I et al. Functional properties of the type 3 InsP(3)receptor in 16HBE14o- bronchial mucosal cells. J Biol Chem 1998;273: 8983-8986; Miyakawa T, Maeda A, Yamazawa T et al. Encoding of Ca(2+)signals by differential expression of IP(3)receptor subtypes. EMBO J 1999;18: 1303-1308). In order to investigate the difference in ATP sensitivity between IP(3)R isoforms at the molecular level, microsomes of Sf9 insect cells expressing full-size IP(3)R1 or IP(3)R3 were covalently labeled with ATP by using the photoaffinity label 8-azido[alpha-(32)P]ATP. ATP labeling of the IP(3)R was measured after immunoprecipitation of IP(3)Rs with isoform-specific antibodies, SDS-PAGE and Phosphorimaging. Unlabeled ATP inhibited covalent linking of 8-azido[alpha-(32)P]ATP to the recombinant IP(3)R1 and IP(3)R3 with an IC(50)of 1.6 microM and 177 microM, respectively. MgATP was as effective as ATP in displacing 8-azido[alpha-(32)P]ATP from the ATP-binding sites on IP(3)R1 and IP(3)R3, and in stimulating IP(3)-induced Ca(2+)release from permeabilized A7r5 and 16HBE14o- cells. The interaction of ATP with the ATP-binding sites on IP(3)R1 and IP(3)R3 was different from its interaction with the IP(3)-binding domains, since ATP inhibited IP(3)binding to the N-terminal 581 amino acids of IP(3)R1 and IP(3)R3 with an IC(50)of 353 microM and 4.0 mM, respectively. The ATP-binding sites of IP(3)R1 bound much better ATP than ADP, AMP and particularly GTP, while IP(3)R3 displayed a much broader nucleotide specificity. These results therefore provide molecular evidence for a differential regulation of IP(3)R1 and IP(3)R3 by ATP. PMID- 10859593 TI - Diverse effects of sphingosine on calcium mobilization and influx in differentiated HL-60 cells. AB - Sphingosine induces a biphasic increase in cytosolic-free Ca(2+)([Ca(2+)](i)) with an initial peak followed by a sustained increase in HL-60 cells differentiated into neutrophil-like cells. The initial peak is not affected by the presence of ethylene glycol bis (beta-aminoethyl ether) N, N, N', N tetraacetic acid (EGTA) in the buffer and appears to be dependent on conversion of sphingosine to sphingosine -1-phosphate (S1P) by sphingosine kinase, since it is blocked by the presence of N, N-dimethylsphingosine (DMS), which, like sphingosine, causes a sustained increase itself. The sustained increase that is elicited by sphingosine or DMS is abolished by the presence of EGTA in the buffer. The sustained sphingosine-induced Ca(2+)influx does not appear due to Ca(2+)influx through store-operated Ca(2+)(SOC) channels, since the influx is not inhibited by SKF 96365, nor is it augmented by loperamide. In addition, sphingosine and DMS attenuate the Ca(2+)influx through SOC channels that occurs after depletion of intracellular stores by ATP or thapsigargin. Both the initial peak and the sustained increase in [Ca(2+)](i)elicited by sphingosine can be blocked by phorbol 12-myristate 13-acetate (PMA)-elicited activation of protein kinase C. Thus, in HL-60 cells sphingosine causes a mobilization of Ca(2+)from intracellular Ca(2+)stores, which requires conversion to S1P, while both sphingosine and DMS elicit a Ca(2+)influx through an undefined Ca(2+)channel and cause a blockade of SOC channels. PMID- 10859594 TI - Simultaneous measurement of intracellular Ca(2+) and nitric oxide: a new method. AB - Different methods to measure the unstable radical nitric oxide (NO) have been established. We are going to present a new method to measure intracellular calcium and NO simultaneously in endothelial cells. A new fluorescent dye (DAF-2) has been developed recently which binds NO resulting in an enhanced fluorescence. We loaded porcine aortic endothelial cells with Fura-2, a fluorescent dye commonly used to measure intracellular calcium, and DAF-2 simultaneously (cell permeable dyes). Using excitation wavelengths of lambda 340 nm (Fura-2) and lambda 485 nm (DAF-2) we could show that thrombin induces an intracellular calcium increase and simultaneously a NO formation in endothelial cells which could be blocked by a NO synthase inhibitor. This new method of a simultaneous measurement of intracellular calcium and NO provides the possibility to follow intracellular calcium and NO distributions online, and is sensitive enough to monitor changes of NO formed by the constitutive endothelial NO-synthase. PMID- 10859595 TI - Synchronization of Ca(2+)-signals within insulin-secreting pseudoislets: effects of gap-junctional uncouplers. AB - The secretory response of the intact islet is greater than the response of individual beta-cells in isolation, and functional coupling between cells is critical in insulin release. The changes in intracellular Ca(2+)([Ca(2+)](i)) which initiate insulin secretory responses are synchronized between groups of cells within the islet, and gap-junctions are thought to play a central role in coordinating signalling events. We have used the MIN6 insulin-secreting cell line, to examine whether uncoupling gap-junctions alters the synchronicity of nutrient- and non-nutrient-evoked Ca(2+)oscillations, or affects insulin secretion. MIN6 cells express mRNA species that can be amplified using PCR primers for connexin 36. A commonly used gap-junctional inhibitor, heptanol, inhibited glucose- and tolbutamide-induced Ca(2+)-oscillations to basal levels in MIN6 cell clusters at concentrations of 0.5 mM and greater, and it had similar effects in pseudoislets when used at 2.5 mM. Lower heptanol concentrations altered the frequency of Ca(2+)transients without affecting their synchronicity, in both monolayers and pseudoislets. Heptanol also had effects on insulin secretion from MIN6 pseudoislets such that 1 mM enhanced secretion while 2.5 mM was inhibitory. These data suggest that heptanol has multiple effects in pancreatic beta-cells, none of which appears to be related to uncoupling of synchronicity of Ca(2+)signalling between cells. A second gap-junction uncoupler, 18 alpha-glycyrrhetinic acid, also failed to uncouple synchronized Ca(2+) oscillations, and it had no effect on insulin secretion. These data provide evidence that Ca(2+)signalling events occur simultaneously across the bulk mass of the pseudoislet, and suggest that gap-junctions are not required to coordinate the synchronicity of these events, nor is communication via gap junctions essential for integrated insulin secretory responses. PMID- 10859596 TI - Subcellular organization of agonist-evoked Ca(2+) waves in the blowfly salivary gland. AB - We have studied the subcellular organization of intra- and intercellular Ca(2+)waves elicited by the neurohormone 5-hydroxytryptamine (5-HT) in intact blowfly salivary glands by using Ca(2+)-sensitive fluorescent probes and confocal microscopy. 5-HT (3 nM) elicited repetitive Ca(2+)waves (1) that were initiated at Ca(2+)-release sites close to the basal plasma membrane, (2) that sequentially spread to the cell apex and (3) that, after a delay of 0.7 +/- 0.20 s at the cell boundaries, spread into adjacent cells. [Ca(2+)](i)increases in the adjacent cells were first detectable at those portions of the lateral plasma membrane that faced a previously activated cell. Electron microscopy revealed that the sites of Ca(2+)wave transmission between the cells are correlated with the distribution of gap junctions that cluster in the basal cell portions. The ensuing intracellular Ca(2+)wave propagated at constant velocity (27 +/- 7.3 microm/s) in the lateral cell plane. Moreover, a basally to apically propagating wavefront was detectable at the cell membrane that bordered on the neighbor that provided the excitatory signal, whereas [Ca(2+)](i)increased simultaneously both apically and basally at the opposite lateral cell border. Overall, the subcellular patterns of Ca(2+)wave propagation differed from the patterns observed in mammalian secretory epithelial cells. The findings impose some constraints on the functional significance of intra- and intercellular Ca(2+)waves and potential mechanisms underlying 5-HT evoked fluid secretion. PMID- 10859597 TI - Humanity and science. PMID- 10859598 TI - Complementary medicine and general practice: an urban perspective. AB - BACKGROUND: Complementary medicine appears to be an increasingly popular option amongst both doctors and patients. General practitioners in more affluent parts of Britain have showed considerable interest in its use. OBJECTIVES: To ascertain use of and attitudes towards complementary medicine, amongst general practitioners working in a socioeconomically deprived urban area. METHODS: A postal questionnaire survey of all general practice principles in Liverpool, using freepost envelopes and one reminder after 3 weeks. With respect to eight common complementary therapies, respondents were asked whether they treat with, refer to or endorse each therapy; for their views on NHS funding, effectiveness, adverse reactions, training needs, and theoretical validity, for each therapy. RESULTS: The response rate was 131/252 (52%), higher amongst women and doctors aged under 40. During the previous week 74 (56%) of respondents had been involved in complementary medical activity with their patients: 13% had treated directly, 31% had referred to and 38% had endorsed one or more complementary therapies. Acupuncture was most popular as an NHS option, and along with osteopathy and chiropractic was the therapy most highly regarded by respondents in terms of effectiveness. Homeopathy and hypnotherapy received a mixed reaction, while medical herbalism, aromatherapy and reflexology were viewed more sceptically. Sixty-two per cent of respondents reported successful outcomes of complementary treatments, compared with 21% reporting adverse reactions. Knowledge and training desires were highest for homeopathy and acupuncture. Respondents were generally uncertain about the theoretical validity of these therapies: 50% though acupuncture had a valid basis, compared with only 23% for homeopathy and 8% for reflexology. CONCLUSIONS: The degree of support for complementary medicine therapies amongst general practitioners in this socioeconomically deprived urban area was similar to that found elsewhere in Britain. These general practitioners appeared to tolerate high levels of clinical uncertainty, endorsing a wide range of therapies, despite little knowledge of their content or conviction of their validity. PMID- 10859599 TI - Efficacy of a potentized homoeopathic drug (Arsenicum-album-30) in reducing cytotoxic effects produced by arsenic trioxide in mice: III. Enzymatic changes and recovery of tissue damage in liver. AB - OBJECTIVE: To determine whether the potentized homoeopathic drug Arsenicum Album 30 can induce enzymatic and some other biochemical changes to repair tissue damage caused by the injection of arsenic trioxide in mice. DESIGN: Controlled laboratory study. METHODS: Mice injected with arsenic trioxide and then orally administered the homoeopathic drug were compared with control animals who either received saline only, or injections of arsenic trioxide, or injections of arsenic trioxide followed by orally administered dilute alcohol. Activities of the enzymes acid and alkaline phosphatases, lipid peroxidation and reduced glutathione, which are used as 'marker' enzymes for cytotoxicity levels, were assessed by standard methods. Histopathological slide preparations of liver were made by routine microtechnique method of tissue sectioning and staining with haematoxylin- eosin for histological examination. RESULTS: The mice fed homoeopathic drug showed positive results of tissue recovery both in terms of enzymatic and histological changes, compared to controls. CONCLUSIONS: The homoeopathic drug is capable of preventing or repairing liver damage induced by arsenic trioxide and the positive changes were also confirmed by the activities of the enzymatic markers. PMID- 10859600 TI - How the public classify complementary medicine: a factor analytic study. AB - OBJECTIVES: To see how lay people group or classify various CAM therapies. DESIGN: Nearly 600 adults rated 39 relatively familiar branches of complementary medicine on four dimensions: whether they had heard of it, whether they think they know how it works; whether they had tried it; and a rating of efficacy on a 10-point scale. RESULTS: As predicted those most heard of were acupuncture, aromatherapy, herbal medicine, hypnosis, massage and yoga while those with lowest ratings were autogenic training, ayurveda, biochemic tissue salts, chelation cell therapy and ozone therapy. A number of multivariate statistical techniques were used to attempt to investigate the perceived dimensional structure of the different therapies. Slightly different structures emerged depending on the question asked and the analysis computed. CONCLUSION: The 'bottom-up' empirically derived taxonomization of therapies was interpretable and showed 10 different factors. The issue of classifying or taxonomizing complementary medicines is discussed. PMID- 10859601 TI - The prevalence of complementary and alternative medicine use among the general population: a systematic review of the literature. AB - OBJECTIVE: To conduct a systematic review of published research investigating the prevalence of complementary and alternative medicine (CAM) use in the general population. DESIGN: A protocol was developed for a systematic review of survey literature identified using two bibliographic databases and citation tracking. The protocol specified criteria for: 1) database searches; 2) selection of studies for review; and 3) description of methodological and substantive aspects of the studies. RESULTS: Twelve studies were reviewed. These estimated the prevalence of CAM use in Australia, Canada, Finland, Israel, the UK, and the USA. The most rigorous studies, conducted in Australia and the USA, showed that a high proportion of the population was using CAM. There was evidence from the USA that CAM use increased significantly among the general population during the 1990s. CONCLUSION: CAM is used by substantial proportions of the general population of a number of countries, but differences in study design and methodological limitations make it difficult to compare prevalence estimates both within and between countries. PMID- 10859602 TI - Complementary and alternative therapies for treating multiple sclerosis symptoms: a systematic review. AB - Multiple sclerosis (MS) is a chronic disease of the central nervous system without a known cure. Thus the role of complementary and alternative therapies (CATs) for the management of symptoms lies in palliative care and this is borne out by the popularity of these treatments amongst MS sufferers. This review is aimed at determining whether this use is supported by evidence of effectiveness from rigorous clinical trials. Database literature searches were performed and papers were extracted in a pre-defined manner. Twelve randomized controlled trials were located that investigated a CAT for MS: nutritional therapy (4), massage (1), Feldenkrais bodywork (1), reflexology (1), magnetic field therapy (2), neural therapy (1) and psychological counselling (2). The evidence is not compelling for any of these therapies, with many trials suffering from significant methodological flaws. There is evidence to suggest some benefit of nutritional therapy for the physical symptoms of MS. Magnetic field therapy and neural therapy appear to have a short-term beneficial effect on the physical symptoms of MS. Massage/bodywork and psychological counselling seem to improve depression, anxiety and self-esteem. The effectiveness for other CATs is unproven at this time. In all the CATs examined further investigations are needed in the form of rigorous large-scale trials. PMID- 10859603 TI - Autogenic training for stress and anxiety: a systematic review. AB - OBJECTIVE: The aim of this systematic review was to evaluate all controlled trials of autogenic training (AT) as a means of reducing stress and anxiety levels in human subjects. METHOD: A search for all published and unpublished controlled trials was carried out in the four major databases, specifically CISCOM, Medline, PsychLit and CINAHL. RESULTS: Eight such trials were located, all of which are included here. The majority of trials were methodologically flawed. A range of outcome measures were used, with Spielberger's State-Trait Anxiety Inventory being the most popular. Deviations from the accepted technique of AT were conspicuous and trials using the classical AT were in the minority. Seven trials reported positive effects of AT in reducing stress. One study showed no such benefit. Since one trial had used AT in combination with another technique, visual imagery, no conclusion can be drawn about the effect of AT in this case. CONCLUSION: No firm conclusions could be drawn from this systematic review. AT, properly applied, remains to be tested in controlled trials that are appropriately planned and executed. PMID- 10859604 TI - Economic analysis of complementary medicine: a systematic review. AB - OBJECTIVE: To review systematically all reports of economic analysis of complementary and alternative medicine. METHOD: Searches were performed in Medline, Embase and AMED for reports of cost description, cost comparison, cost effectiveness, or cost benefit studies. Prospective studies that investigated comparative groups were considered to be of higher quality. RESULTS: A total of 34 reports was included. Retrospective studies in which a range of therapies are provided in primary care suggest that these may reduce referral and treatment costs, but prospective studies suggest that complementary medicine is an additional expense and does not substitute for orthodox care. For individual therapies, one thorough but retrospective study suggests that carefully targeted acupuncture may reduce referral costs for musculoskeletal problems. One large pragmatic study of spinal manipulative therapy suggests that this treatment may reduce the societal costs of back pain, but four controlled trials found that manipulative therapy does not reduce the costs incurred by the back pain patients themselves or by their health insurance provider. CONCLUSION: Spinal manipulative therapy for back pain may offer cost savings to society, but it does not save money for the purchaser. There is a paucity of rigorous studies that could provide conclusive evidence of differences in costs and outcomes between other complementary therapies and orthodox medicine. The evidence from methodologically flawed studies is contradicted by more rigorous studies, and there is a need for high quality investigations of the costs and benefits of complementary medicine. PMID- 10859605 TI - CAM evaluation comes into the mainstream: NIH specialized Centers of research and the University of Maryland Center for Alternative Medicine Research in Arthritis. AB - In September of 1999 the National Institutes of Health (NIH) announced the funding of five Specialized Centers of Research in complementary and alternative medicine (CAM), bringing the total number of centers being supported to nine. The NIH center grant model provides a tremendous boost to the scientific investigation of CAM, nurturing an emerging field through the support of a step wise research program of clinical and pre-clinical trials and developmental and feasibility studies; the building of infrastructure; and the training of a new cadre of scientific investigators in the field. This article explains the overall objectives of the NIH Specialized Centers program and focuses on one of the oldest CAM research centers in the USA, exploring some of the challenges faced in conducting CAM research while developing a center, and some of the goals and activities of the center. PMID- 10859607 TI - Chiropractic peripheral joint technique PMID- 10859606 TI - The evidence for evidence-based medicine. PMID- 10859608 TI - Herbal medicine in primary care PMID- 10859609 TI - COMPLEMENTARY/ALTERNATIVE MEDICINE: AN EVIDENCE-BASED APPROACH PMID- 10859610 TI - Abc of complementary medicine PMID- 10859611 TI - Complementary medicine on the world wide web PMID- 10859613 TI - EVIDENCE-BASED COMPLEMENTARY MEDICINE: INTERNATIONAL CONGRESS ON CLINICAL RESEARCH AND QUALITY MANAGEMENT IN COMPLEMENTARY MEDICINE MUNICH GERMANY 6Eth 8, APRIL 2000 PMID- 10859612 TI - The millennium dome--reflections for mind and body. PMID- 10859614 TI - Advancing research in complementary medicine london 10 march 2000 the wellcome trust PMID- 10859615 TI - Complementar therapies in cancer care a report produced for macmillan cancer relief by dr michelle kohn PMID- 10859616 TI - Reflections on CAM research in the UK. PMID- 10859617 TI - Recently published research AB - This feature presents a personal selection of some significant articles that were listed in Medline from July to September 1999. Readers need to be aware that abstracts written by authors themselves may sometimes give a positive 'slant' to the paper which is not strictly based on the findings. Where authors' addresses are included in the database, we have given them here so that readers who wish for a reprint can request it directly. Alternatively, a number of journals now publish on the world wide web and some articles may be downloaded in pdf versions. Copyright 2000 Harcourt Publishers Ltd. PMID- 10859619 TI - Similia PMID- 10859618 TI - Events PMID- 10859620 TI - Nurse-doctor relationships: conflict, competition or collaboration. PMID- 10859621 TI - News and Issues. PMID- 10859622 TI - Discharge from intensive care: a view from the ward. AB - Relocation stress is a common phenomenon in patients discharged from an intensive care unit (ICU) to a ward. A variety of nursing interventions, initiated by intensive care nurses, have been introduced following research in this area. Ward nurses are ideally situated to minimize stress in this patient population, yet their contribution has not been considered. The aim of this study was to identify the experience of the ward nursing staff when receiving a patient from the ICU. An exploratory pilot study was conducted over a 6-month period. The sample group comprised nursing staff in two wards, who regularly received ICU patients. Data collection methods were triangulated and involved the use of open-ended questionnaires and semi-structured interviews. Thirty-six questionnaires were sent, yielding a 36.1% (n = 13) response rate. Seven staff of various grades were interviewed. Data analysis was undertaken using Burnard's (1991) Thematic Content Analysis. Four major categories were identified in the analysis of the data. These were emotions; problems; communication; and interventions. However, the experience of ward staff receiving patients from intensive care differed according to grade. PMID- 10859623 TI - Use of telephone follow-up for post-cardiac surgery patients. AB - Telephone follow-up has been used in a variety of settings as a means of supporting patients post-discharge. This paper describes the implementation of a telephone follow-up service for cardiac surgical patients, both to monitor their progress and to bridge the gap between home and hospital. Surgical unit nurses called 1594 patients between May 1995 and October 1997. These calls were made a month after each patient had been discharged and a specially designed form was used as a guideline for the calls. This covered three main aspects of their recovery: discharge monitoring; medical problems; and convalescence problems. As well as providing a framework for the calls, the forms were then used for data collection. The forms were audited on a monthly basis with the results being communicated to the ward staff. Data gathered from the calls have highlighted areas where patient education needs to be improved, e.g. pain control, and this has been addressed. In October 1996, an evaluation of the service was performed. One hundred patients were sent a postal questionnaire asking for their views on the telephone service. Eighty-two of these were returned and the results of this survey are also contained in this report. Over time, the number of patients called each month has increased; information regarding support post-discharge has improved and the form used for calls has been revised to make the questions more appropriate. PMID- 10859624 TI - Intensive care nurses' experiences of caring. Part 2: Research findings. AB - In considering the concept of caring, this second paper presents the research findings obtained from a phenomenological study conducted to answer the question 'what is caring for nurses working in intensive care?'. Unstructured, in-depth interviews were conducted to collect data on the experiences of nine staff nurses, who volunteered to participate in the study. Three major themes were drawn from the data using Colaizzi's procedure of inductive reduction. These were 'being involved', 'sustaining' and 'having frustrations'. A resultant description of caring identified that these nurses participated in delivering three types of caring involving physical, technical and emotional labour. These were 'being close', 'being there' and 'doing to'. Implications for nursing practice, education and further research are suggested. PMID- 10859625 TI - Building a framework for getting evidence into critical care education and practice. AB - One challenge for nurse educators is how best to enhance the integration of theory and practice elements in relation to critical care nursing. Practice should be evidence-based, i.e. the best available empirical evidence, including recent research findings, should be applied in practice in order to aid clinical decision-making. Barriers to the implementation of research exist at many levels including the individual practitioner, the clinical team, the practice setting and wider organizational factors. The authors propose that clinical guidelines can provide a vital link between theory and practice. At varying levels the use of care protocols, clinical pathways and algorithmic guidelines (provided they are rigorously reviewed and evidence-based) can help infuse research into practice, thereby promoting quality and standardization of care. The purpose of this paper is to discuss the value and use of these frameworks in promoting and raising awareness of the need for and use of evidence-based approaches to critical care education and practice. In this paper, we present outline information relating to an assessment method, adopted for continuing education courses in critical care within our department. This approach is designed to combine the best available evidence with reflective practice through the assessment process. PMID- 10859626 TI - Nursing care for raised intra-abdominal pressure and abdominal decompression in the critically ill. AB - Abdominal assessment is one of a number of continuous assessments that critical care nurses undertake. Since 1988 in the Department of Critical Care Medicine (DCCM), the technique of abdominal decompression has become another therapy for severe critical illness. The critical care nurse requires to have an understanding of raised intra-abdominal pressure assessment, pressure measurement and the care of abdominal polypropylene mesh insertion in the critical care setting. Our experience has been that the use of polypropylene mesh insertion halved since 1993. A retrospective study (Torrie et al. 1996) of 68 occasions (64 patients) of polypropylene mesh insertion, showed that seven patients developed fistulas and 32 patients died. There was no dehiscence of the mesh from the fascia. Forty-two wounds had primary fascial closure (28 with primary skin closure, 3 with secondary skin closure, 11 left to granulate) and 3 of them later dehisced. At follow-up (27 patients, median 7.5 months), 6 had stitch sinuses, and 5 had incisional hernias. Care of patients with polypropylene mesh inserted requires vigilant nursing practice but decompression of raised intra-abdominal pressure can be life-saving and complications are manageable. PMID- 10859627 TI - Predictors of rural critical care nurses' willingness to care for people with AIDS. AB - The purpose of this study was to examine the relationship between rural critical care nurses' attitudes about acquired immunedeficiency syndrome (AIDS) and people with AIDS (PWAs), and their willingness to provide care to AIDS patients. Sixty one critical care nurses in nine rural counties in the northeastern USA completed a mailed questionnaire as part of a larger study of 957 rural nurses. A bivariate logistic regression analysis revealed a relationship between willingness to provide care and positive attitudes about homosexuality, nursing care concerns, and professional-societal concerns. However, a multivariate logistic regression indicated that the most significant factors influencing rural critical care nurses' willingness to care were their feelings of not being prepared to care for people with AIDS, and their anxiety and fears about contracting the disease from their patients. These findings add insight into the care of critically ill AIDS patients and support the need for continuing educational efforts in rural areas of the USA to address critical care nurses' concerns. PMID- 10859629 TI - Chest X-ray quiz. PMID- 10859628 TI - The standard of suction for patients undergoing endotracheal intubation. AB - The purpose of this study was to determine whether using a standard method of endotracheal suctioning, to ensure consistent use of available knowledge, had any impact on patient care. Using experimental study design, the results of two different methods of suctioning in a cardiovascular surgery intensive care unit were compared. One method was the suctioning procedure applied by the nurses working in the intensive care unit. The other one, standard suctioning procedure, was developed based on the related literature and applied to the patients assigned to the experimental group by the researcher herself. Mean arterial blood pressure (MAP), heart rate (HR), and arterial blood gases (ABGs) were measured before the procedure, immediately after, 5 and 15 minutes after the procedures for both control and experimental group. The majority of the nurses suctioning the control group did not evaluate the ABGs after endotracheal suctioning, none of these patients was given oxygen both before and after the suctioning, and suctioning took longer time than recommended. To compare the results of the two different methods, the values of MAP, HR, PO2 (arterial oxygenation), PCO2 (arterial carbondioxide), and HCO3- (hydrogen carbonate) 15 minutes after the procedure were used, and the differences between the two methods were statistically significant (P < 0.05). PMID- 10859630 TI - Rofecoxib (Vioxx)--the first of a new generation of NSAIDs. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most frequently prescribed medicines in practice and, although of undoubted benefit for many, they are potentially toxic for some. Does rofecoxib, and the others which will inevitably follow, hold out the promise of safe and effective NSAID therapy in the future? PMID- 10859633 TI - Interleukin-1 receptor antagonist as a treatment of HIV infection. AB - The human immunodeficiency virus (HIV) apparently utilizes human chromosome 2, interleukin-1 (IL-1), glucocorticoid hormones, and viral Tat protein to accelerate its replication and the synthesis of all HIV proteins. HIV Tat protein binds to the long terminal repeat (LTR) ribonucleic acid, including the trans acting responsive (TAR) sequence and the promoter region to increase HIV replication. Tat-TAR transactivation requires a factor encoded on the long arm of chromosome 2. The interaction of HIV with chromosome 2 may also cause the observed inhibition of interleukin-1 receptor antagonist (IL-1RA), thus increasing the production of IL-1. IL-1, in turn, stimulates the HIV-1 enhancer region of the LTR, thus increasing HIV gene expression and replication. IL-1 also induces glucocorticoid hormone synthesis which stimulates HIV in the virion infectivity factor (Vif) region, thus increasing HIV infectivity. It is, thus, proposed that IL-1RA not only may serve to inhibit HIV-induced IL-1, but may be the unidentified human chorionic gonadotropin-associated factor recently found to have anti-HIV and anti-Kaposi's sarcoma activity. PMID- 10859634 TI - Is carbonyl detoxification an important anti-aging process during sleep? AB - Organisms living on the earth may undergo inevitable toxification by biological 'garbage', a variety of bio-metabolites. Such garbage includes a particularly large number of toxic carbonyls, such as alpha,beta-unsaturated carbonyls created by free radicals, glycation, and other post-translational side-reactions during various stresses and diseases. The accumulation of these toxic substances and their crosslinking products leads to the formation of different age pigments, such as lipofuscin, lens cataracts, and crosslinked collagen. The diurnal fluctuation in the concentration of the pineal gland hormone, melatonin, may be responsible for the 'cleaning activities' that reverse the covalently-bound semi toxified proteins and nucleic acids. This toxification-cleaning cycle may explain the biochemical necessity for sleep of human and animals during aging. PMID- 10859635 TI - Aboulia: neurobehavioural dysfunction of dopaminergic system? AB - Recent advances in the understanding of the neural substrates of goal-directed behaviour have created new interest in unlocking the mystery behind those disorders that are characterized by poverty of thought and action. In this review, various studies will be considered which proffer converging evidence that the dopaminergic brain circuitry running from ventral tegmental areas in the midbrain, via nucleus accumbens in the forebrain, to the frontal cortex, tends to produce aboulia when its restitutive function fails. Such aboulic deficits occur in various neurological and psychiatric disorders in which they have profound implications for the patients' management, rehabilitation and social interactions. We begin by examining the consequences of dopamine agonism and antagonism in pre-clinical studies and draw on the inferences that can be made from studies in humans. We then go on to discuss aboulic features in neuropsychiatric conditions, focusing on clinical manifestation, animal models, abnormal dopamine activity and pharmacological interventions. PMID- 10859636 TI - Tumor-specific immunotherapy based on dominant models of natural tolerance. AB - The assumption that cancer immunotherapy may be based on the existence of autoreactive lymphocytes recognizing self-antigens on cancer cells, obviously opens a new opportunity. Nevertheless this analysis, relying on a recessive model of natural tolerance, limits the approach to try to activate peripheral lymphocytes, by increasing co-stimulatory signals or using modified self-antigens for immunization. Here we hypothesize that, based on emerging dominant tolerance notions in autoimmunity, it would be possible to induce a specific autoimmunity against tumor cells and arrest their growth following the removal of regulatory T cells. These immunoregulatory cells suppress available immunocompetent autoreactive cells capable of destroying tumor cells. Therefore, in order to reach a complete tumor-specific autoimmunity it is necessary to combine the T cell immunosuppression which abrogates the regulatory cells, with the cancer vaccines, which induces extensive proliferation of lymphoid cells directed towards specificities on tumor cells. PMID- 10859637 TI - Schizotypy and leadership: a contrasting model for deficit symptoms, and a possible therapeutic role for sex hormones. AB - Associational loosening, slow and faulty information processing, poor gating of irrelevant stimuli, poor ability to shift attention, poor working memory, passivity, ambivalence, anhedonia, and impaired motor coordination are cardinal features of schizophrenia but, unlike delusions and hallucinations, they are related more to negative/deficit symptoms. As summarized by Bass, numerous studies have correlated leadership with 'ambition, initiative and persistence' (opposite of passivity), 'speed and accuracy of thought', 'finality of decision,' or decisiveness (the opposite of ambivalence), 'mood control, optimism and sense of humor' (opposite of anhedonia), etc. Andreasen et al postulate that a disruption in the circuitry among nodes located in the prefrontal regions, the thalamic nuclei, and the cerebellum produces 'cognitive dysmetria', meaning difficulty in prioritizing, coordinating, and responding to information, and that it can account for the broad diversity of symptoms of schizophrenia. A relationship between cognitive processes and cerebellar and basal ganglia functions, and a role of neocerebellum in rapidly shifting attention, have been demonstrated. The cognitive styles, including a proficiency to quickly shift attention, of several famous leaders are used as examples of this contrasting model. Julius Caesar and Napoleon, for instance, could dictate to up to six secretaries simultaneously, using their exceptional working memories, and proficiency in quickly and effortlessly shifting attention while flawlessly gating irrelevant external and internal stimuli. It is suggested that specific brain imaging studies could illustrate this contrast. Gray et al noted positive correlations between 'dominance', an important leadership trait, and serum levels of dehydroepiandrosterone (DHEA) and testosterone (T), but not of more potent dihydrotestosterone (DHT), in over 1700 older men. Though not scientifically rigorous, the author noted positive correlations (P = 0.0162) between the self rated ratings of voice depth (promoted by T) and of leadership, but none between those of body hair (DHT dependent) and of leadership in 47 male US National Academy of Sciences members. And 43 male US Senators had deeper voices than 36 male House members (P<0.01) who, in turn, had deeper voices than either of two groups (numbers 102 and 72) of male scientists (P<0.01). Therapeutically, before chlorpromazine, DHEA had been used in young schizophrenics with modest success in improving deficit symptoms. DHEA, or other sex hormones, or some of their natural and synthetic derivatives may prove to be valuable to treat deficit symptoms of schizophrenia in both sexes. PMID- 10859638 TI - Limitations of clinical and biological histology. AB - Histology, including histochemistry, histopathology, electron microscopy and immunocytochemistry, can be considered as two disciplines--clinical and biological. The former is used to make a clinical diagnosis, based on empirical comparisons between, on the one hand, the clinical health of a normal subject with the histological appearances of the organs, and on the other hand, the clinical disease or syndrome of a patient, with the different histological appearances of the organs believed by the clinicians to be affected. It is intended that biological histology should be the most accurate description possible of the structure and chemistry of the living tissue in the intact healthy organism. PMID- 10859639 TI - Does the mechanism responsible for TNF-mediated insulin resistance involve the proteasome? AB - Recent studies have demonstrated that in many pathological states there is an overproduction of tumour necrosis factor-alpha (TNF). Interestingly, TNF also seems to be responsible for the insulin resistance associated with these pathological states, since decreases the tyrosine kinase activity of the insulin receptor. Our group has demonstrated that TNF is able to activate the proteasome mediated ubiquitin-dependent proteolysis. Since this proteolytic system is involved in the control of receptor-associated tyrosine kinase activity (i.e. insulin receptor), it is postulated here that the mechanism of TNF-induced insulin resistance is mediated by the activation of the proteasomic, ubiquitin dependent proteolysis. PMID- 10859640 TI - Gallstones, the choledochoduodenal junction and initiation of acute pancreatitis: are two stones the culprits rather than one stone? AB - Reflux of biliary secretions into the pancreatic duct following gallstone obstruction of the common biliary pancreatic ampulla has been implicated as a cause of acute pancreatitis. However, the pancreatic duct pressure is higher than the biliary pressure and, therefore, the simple obstruction of the choledochoduodenal junction by one gallstone does not result in biliary pancreatic reflux. We propose a mechanism whereby simultaneous migration and sequential impaction above and below the common biliary pancreatic ampulla of two gallstones allows for the creation of a toxic bile-pancreatic juice mixture in the common bile duct, subsequent reversal of the pressure gradient and reflux of the toxic secretions into the pancreatic duct. PMID- 10859641 TI - The role of dysregulated amygdalic emotion in borderline personality disorder. AB - Borderline personality disorder (BPD), is a condition that has a high mortality and is associated with much distress for the sufferers as well as with difficult management problems for health professionals. Taking emotional dysregulation as the core feature of BPD, the authors propose that the disorder arises from impaired modulation of subcortical inputs to consciousness. We hypothesize that the amygdaloid complex, and its connections with thalamus, cingulate cortex and insular cortex are critical in the development and maintenance of the disorder. If this is the case, peptides such as galanin, somatostatin and cholecystokinin will be the most important neurotransmitters, thus explaining the relative lack of efficacy of standard antipsychotic and antidepressant drugs. PMID- 10859642 TI - Improvement of cystic fibrosis using antitumoral drugs: a hypothesis. AB - The improvement of cystic fibrosis using antitumoral drugs has been reported. The hypothesis of a somatic reduction to heterozygosity of the CFTR gene mutations by homologous recombination in lung epithelium is proposed. PMID- 10859643 TI - Reasons for the cooperative effects in the haemoglobin case. AB - The cooperative question in haemoglobin is faced, and the use of an analogy to an electronic circuit is hypothesized. On this basis, the Hb-oxygen uptake and release mechanism is discussed, and a new explanation of the cooperative effect is proposed. PMID- 10859644 TI - Dysbaric osteonecrosis: a reassessment and hypothesis. AB - Dysbaric osteonecrosis is associated with exposure to large ambient pressure changes, and comprises necrotic lesions in the fatty marrow-containing shafts of the long bones, and the ball and socket joints (hips and shoulders). The fundamental causes are still in question and the illness remains a significant health hazard. Radiological and pathological features of both dysbaric and non dysbaric osteonecrosis are indistinguishable and both are characterized by intramedullary venous stasis, ischemia and necrosis of bone. It has been generally accepted that gas bubbles (probably by initiating intramedullary venous stasis) are the prime cause of dysbaric osteonecrosis, as well as being responsible for Type 1 Decompression Sickness or 'the bends'. Importantly, however, not all series have found a correlation between dysbaric osteonecrosis and 'the bends'. Thus even though it is likely that gas bubbles remain the prime cause of dysbaric osteonecrosis, workers have proposed that in some cases there is another etiological factor which may exaggerate the pathologic effects of gas bubbles, making the bone more susceptible to necrosis. It is proposed that rapid compression by impeding venous drainage from bone initiates intramedullary venous stasis. In the presence of intramedullary gas bubbles, this may progress to thrombosis, ischemia and bone necrosis. The review offers an explanation for total sparing of the knee joint in dysbaric osteonecrosis, and sole involvement of the hip and shoulder (in terms of sub-articular lesions and subsequent joint collapse). In addition to continued observance of proper decompression procedures, a slower rate of compression may further reduce the incidence of dysbaric osteonecrosis. Bone death or osteonecrosis is a concept which Hippocrates put forward in antiquity (1), but it was not until 1794 that James Russell of Edinburgh wrote the first modern-day descriptions. In these cases infection was the predominant etiology (1,2). In 1888 Konig described necrosis of the adult femoral head without infection (3) (aseptic necrosis of bone) and in the same year Twynam reported a case of osteonecrosis in a caisson worker (4) in which there was still a significant infective component. In 1911 Bornstein and Plate, followed later and independently by Bassoe in 1913, presented radiological confirmation of aseptic necrosis of bone in compressed air workers (5). The first report of aseptic necrosis in an underwater diver subsequently appeared in 1936 (6). The condition of aseptic necrosis of bone in association with exposure to raised ambient pressure (previously referred to as caisson disease, pressure induced osteoarthropathy (7), 'bone rot' (8) and other synonyms (6)) is now generally known as dysbaric osteonecrosis (6). Despite detailed examination of this problem by many authorities, dysbaric osteonecrosis still remains a significant occupational hazard with serious medico-legal consequences (5-13). This suggests that preventative measures are being based upon an incomplete understanding of the pathophysiology of the disease, and that other etiological factors are perhaps being overlooked. PMID- 10859645 TI - Implications of early apoptosis of infected cells as an important host defense. AB - Apoptosis is widely recognized as being a host defense against viral infections, since viruses require live cells. There has been increasing acceptance of the view that apoptosis is also a defense against other intracellular pathogens and even against pathogens that adhere to host cells. An implication of apoptosis being a host defense is a need to reassess to what extent the cell death at infection sites may constitute a protective host response. A concept stressed here is that infected cells are a hazard to other cells and to the individual, so the benefits of early apoptosis are emphasized. Therefore, promoting the survival of infected cells, even though still functional, may carry risks. A further consideration is the possibility that the apoptotic stimulus of nutrient restriction may be acting in infection-induced anorexia to promote apoptosis of infected cells, thereby serving as a non-specific host defense. PMID- 10859646 TI - Why infection-induced anorexia? The case for enhanced apoptosis of infected cells. AB - A medically important paradox is why the body's own cytokines lead to reduced appetite and apparently inefficient metabolism as part of the acute-phase response. This self-induced nutrient restriction occurs just when the body must maintain a fever and other defensive functions. This paradox is often ignored or considered a metabolic derangement. Others, recognizing it to be a programmed response which must have net beneficial effects, consider the nutrient restriction to be an attempt to deny resources to infectious organisms. However, this explanation fails to address how the pathogen can be harmed more than the host. The hypothesis presented here offers an explanation. Apoptosis, or cell suicide, is becoming recognized as a useful defense against intracellular parasites, and nutrient restriction promotes apoptosis. Thus, nutrient restriction may encourage apoptosis of infected cells. Nutrient restriction can thereby offer protection by simultaneously limiting nutrients to both the host cells and the infectious organisms. PMID- 10859647 TI - Can the pentitol-hexitol theory explain the clinical observations made with xylitol? AB - The natural dietary carbohydrate xylitol has been used as a source of energy in infusion therapy and found to act curatively in certain clinical situations. Xylitol has also been used as a sweetener in the diabetic diet and as a non- or anticariogenic agent. Xylitol is a sugar alcohol (polyhydric alcohol) of the pentitol type. The various advantageous clinical effects associated with enteral and parenteral administration of xylitol can be considered to result from the five-carbon (pentitol) nature of the molecule and from the molecule's special configuration even when compared with other pentitols. Such effects may be regarded as simple consequences of evolutionary expediency in a situation where human nutrition and man's significant energy-yielding metabolic pathways are associated with the six-carbon nature of D-glucose and the close derivatives and polymers of D-glucose and related sugars, and the physiologic involvement of the five-carbon xylitol in several ancillary pathways. Consequently, most clinical effects occasioned by xylitol cannot be expected to be caused by six-carbon hexitols such as D-mannitol and D-glucitol. A simple pentitol-hexitol theory seems to explain most of the clinical effects associated with the administration of xylitol. This theory is in congruence with the general evolutionary development in which the metabolism of C(6)-based carbohydrates is often inhibited by C(5)-based ones (as manifested in certain bacterial infections in man), or where the presence of the C(5)-based xylitol forwards therapeutically significant metabolic pathways (as observed in parenteral nutrition and treatment of certain enzyme deficiencies). The validity of the theory can be verified in controlled clinical trials. PMID- 10859648 TI - A relationship between autoimmune thyroiditis and benign paroxysmal positional vertigo? AB - Benign paroxysmal positional vertigo (BPPV) is a labyrinthine disorder with a typical behavior: intense crises of rotational vertigo induced by postural changes of the head, with short duration and usually good responsiveness to rehabilitative maneuvers. This phenomenon is thought to be subsequent to the movement of floating particles in the labyrinthine fluids, which can provoke gravitational stimulations. In order to conduct a metabolic and autoimmune examination, 70 patients affected by BPPV were examined. In 34 cases (48.5%) autoimmune alterations were found: in 19 cases (27.1%) the level of anti-thyroid antibodies far exceeded the normal values with a significant incidence in comparison with a control group (P<0.01). No other 'risk factors' were present. It can be hypothesized that the diffusion of immune-complexes in the inner ear could change the composition of the endolymphatic fluid exerting a mechanical stimulation of the receptors and provoking the typical vertigo. PMID- 10859649 TI - Statistico-mechanics of chromosomal insertions in human cancers. AB - The paper discusses a mechanism for segmental insertions (ins) between two chromatids. In human cancers, such rearrangements are significantly less frequent than reciprocal translocations (rt). Regarding the clinical incidence of ins versus rt, the proposed dynamics is more satisfactory than the random mutation hypothesis. PMID- 10859650 TI - New aspects in genotoxic risk assessment of styrene exposure--a working hypothesis. AB - Styrene is one of the most important plastic monomers worldwide. Styrene-7,8 oxide (SO), the major in-vivo metabolite of styrene, is classified as probably carcinogenic to humans and carcinogenic in rodents. Biological monitoring of exposure to styrene is usually carried out by determination of mandelic acid and phenylglyoxylic acid, the two main styrene metabolites in urine. SO binds covalently to human plasma protein and haemoglobin. The ability of SO to induce DNA adducts and DNA strand-breaks has been well documented. Recently in-vitro results showed that SO may disrupt the pre-existing oxidative status in white blood cells. This disruption would alter the balance between oxidants and antioxidants in cells. Styrene exposure can also result in oxidative DNA damage. A significant increase of 8-hydroxy-2;-deoxyguanosine (8-OHdG) has been found in white blood cells of styrene-exposed workers. According to these findings we propose a new hypothesis for the genotoxic risk assessment of styrene. Depletion of glutathione and increase in lipid peroxidation, similarity in the decrease of high molecular weight (HMW) DNA fragments after SO exposure compared to hydrogen peroxide (H(2)O(2)) exposure, oxidative DNA damage (increased amounts of 8-OHdG and an increased level of DNA strand-breaks) following styrene or SO exposure are due to oxidative stress which can be a result of the imbalance between oxidants and antioxidants. Formation of protein-, RNA- and DNA-adducts, changes in DNA repair capacity and styrene metabolism following styrene exposure could cause this imbalance between oxidants and antioxidants. Oxidative stress seems to be the basis for genotoxic risk assessment of styrene. PMID- 10859651 TI - New insights into the annealing behaviour of DNA. AB - In mutagenesis experiments with megaprimers of length 450-770 bp optimal annealing of the megaprimer to the template-DNA was shown to occur at unexpectedly low temperatures under conditions suboptimal for amplification of the elongated megaprimer-product by the short flanking primers, although very high annealing temperatures would have been expected to give best results. This finding contradicts the rule that annealing temperatures should be just below the melting temperature of a given primer, and leads to the conclusion that the curves of melting and annealing temperatures diverge rather than running in parallel. PMID- 10859652 TI - Effect of angiotensin-converting enzyme inhibitors on non-diseased myocardium of experimental animals: potential clinical implications. AB - Angiotensin-converting enzyme (ACE) inhibitors protect the hearts of patients with different levels of cardiac disorder. The greatest benefit seems to be achieved in subjects with most severe heart failure. Moreover, ACE inhibition is protective also in patients without manifested heart failure but with severe systolic left ventricular dysfunction. Data are presented that ACE inhibitors can alter the composition of the myocardium also in control: healthy animals. In rats and rabbits with non-diseased heart, chronic ACE inhibition reduced fibrotic tissue concentration in the left ventricle. We speculate that if this were applied to humans, ACE inhibition may prove to be of potential benefit in subjects with normal systolic function but with a trend to left ventricular filling abnormalities caused by increased ventricular stiffness. In these patients reduction of myocardial fibrotic tissue might prevent deterioration of diastolic function. PMID- 10859653 TI - Memory creation and the treatment of psychiatric disorders. AB - False-memory syndrome describes the 'recovery' of memories of traumatic events which did not take place. 'Recovered' memories are created as a result of suggestion or other psychological maneuvers. This means that memories can be implanted. The author suggests that the possibility of use of implantation of 'good' memories for the treatment of certain psychiatric disorders should be explored and that implantation of 'good' memories may be called 'positive memory creation therapy'. The author also suggests that memory creation treatment should be used under strict ethical and legal control. PMID- 10859654 TI - The effects of natural and man-made electromagnetic fields on mood and behavior: the role of sleep disturbances. AB - Natural and man-made electromagnetic fields influence the mood and behavior of healthy and sick people. Considerable evidence suggests that electromagnetic fields affect sleep. The author suggests that electromagnetic field-induced changes in sleep may mediate the effects of electromagnetic fields on mood, behavior, and cognitive abilities. The author further suggests that the development of sleep abnormalities in persons exposed to artificial electromagnetic fields may predict the onset of a psychiatric disorder at a later time and that early intervention may prevent the onset of a psychiatric disease. PMID- 10859655 TI - Plausibility of homeopathy and conventional chemical therapy: the systemic memory resonance hypothesis. AB - The controversy surrounding clinical observations and double-blind studies on homeopathic treatments is lessened when modern dynamical systems analysis is applied to high-dilution therapies. The logic of recurrent feedback loops, which applies to all dynamical network systems, inexorably leads to the systemic memory hypothesis - that complex patterns of emergent information and energy are stored to various degrees in physical, chemical, and biological systems. The addition of resonance, a dynamic pattern recognition process, explains many classic observations using high-dilution therapies. The systemic memory resonance hypothesis potentially provides a plausible biophysical mechanism for explaining not only how high-dilution therapies contribute to healing, but by extension, how information and energy in low-dilution and chemical therapies contribute to healing as well. PMID- 10859656 TI - The sequestration hypothesis: an explanation for the sensitivity of malaria parasites to nitric oxide-mediated immune effector function in vivo. AB - While the effect of reactive nitrogen intermediates (RNI) against macrophage dwelling protozoa, such as Leishmania and Toxoplasma has become established, the possible antiparasitic function of nitric oxide (NO) and RNI against the intracellular blood stages of malaria and babesia has, until recently, been less well accepted. This was, at least in part, due to the long-standing notion that haemoglobin (Hb) universally scavenges NO and thus that erythrocytes act as a permanent sink for this molecule. It is now known that NO can be released as well as scavenged by Hb, and that the less oxygenated the blood the lower the affinity of Hb for NO. As a consequence, NO is preferentially released by venous erythrocytes. Based on the increased sensitivity of Plasmodium falciparum infected erythrocytes to RNI in venous blood that was recently demonstrated, it is proposed that the noted greater susceptibility of mature intra-erythrocytic forms of malaria, late-stage schizonts, is coincidental with their peripheral blood withdrawal by sequestration to deep-tissue capillaries. This environment is non NO-scavenging in nature and one which would bring schizonts and macrophages into intimate proximity, providing diffusion distances sufficiently short for RNI to be effective. Given its short half-life, this hypothesis explains the potential for NO to be toxic for malaria parasites in vivo, and suggests that sequestration, a mechanism adopted by the parasite to supposedly avoid immune surveillance, may in fact have a partially counteractive effect. PMID- 10859657 TI - A continuum approach to modelling cognitive disorders. AB - At the present time, the dominant conceptual framework in neuroscience views neurons as discrete information processing units. While this framework has undoubtedly been successful in explaining phenomena at the single-cell level, it has had less success in explaining large-scale neural phenomena such as cognitive disorders. An alternative conceptualization of brain function is in terms of an energy continuum, where energy is an abstract variable modelling the flow of electrochemical activity between neurons. Using this energy (E) model, large scale phenomena may be incorporated within a single conceptual framework. This framework can in turn be related to the underlying biochemistry. The E model also generates testable predictions, and should prove quantifiable using differential equations. These claims are illustrated with simple examples from the spectrum of cognitive disorders. PMID- 10859658 TI - Role of the pineal gland in hibernators: a concept proposed to clarify why hibernators have to leave torpor and sleep. AB - The contention that torpor during hibernation might not be a kind of sleep but a kind of sleep deprivation may be correct, as it can be conceptually explained. Sleep is part of life and cannot be part of the torpor state, which has no reactions. The state of torpor has a time limit and, if this is not observed, death follows by freezing. The evolutional differences particularly in pineal gland physiology may explain the differences between the blueprints governing vital reflexes in humans and hibernators. PMID- 10859659 TI - Moon phase at the dates of birth and decease of anthroposophic pioneers. AB - Early adherents of Rudolf Steiner, the founder of the anthroposophical movement, tend to be born and to die during the dark half of the lunar month. There is significant correlation (P = 0.03) between the distributions of the lunar elongation at birth and at decease. However, this correlation does not operate at the level of individuals, suggesting that the effects of birth date and death date are statistically independent. PMID- 10859661 TI - Ketosis with enhanced GABAergic tone promotes physiological changes in transcendental meditation. AB - Transcendental meditation (TM) is a stylized form of physical and mental relaxation which is associated with changes in the secretion and release of several pituitary hormones. The hormonal changes induced by TM mimic the effects of the inhibitory neurotransmitter gamma aminobutyric acid (GABA). It is hypothesized that TM produces changes in pituitary hormone secretion by enhancing hypothalamic GABAergic tone as a result of TM associated ketosis. Ketosis enhances the entry of glutamate, the amino acid substrate of GABA into synaptosomes, making more glutamate available for conversion to GABA through the glutamate decarboxylase pathway. PMID- 10859660 TI - The phenomenon of claimed memories of previous lives: possible interpretations and importance. AB - Several disorders or abnormalities observed in medicine and psychology are not explicable (or not fully explicable) by genetics and environmental influences, either alone or together. These include phobias and philias observed in early infancy, unusual play in childhood, homosexuality, gender identity disorder, a child's idea of having parents other than its own, differences in temperament manifested soon after birth, unusual birthmarks and their correspondence with wounds on a deceased person, unusual birth defects, and differences (physical and behavioral) between monozygotic twins. The hypothesis of previous lives can contribute to the further understanding of these phenomena. PMID- 10859662 TI - A theoretical model based upon mast cells and histamine to explain the recently proclaimed sensitivity to electric and/or magnetic fields in humans. AB - The relationship between exposure to electromagnetic fields (EMFs) and human health is more and more in focus. This is mainly because of the rapid increasing use of such EMFs within our modern society. Exposure to EMFs has been linked to different cancer forms, e.g. leukemia, brain tumors, neurological diseases, such as Alzheimer's disease, asthma and allergy, and recently to the phenomena of 'electrosupersensitivity' and 'screen dermatitis'. There is an increasing number of reports about cutaneous problems as well as symptoms from internal organs, such as the heart, in people exposed to video display terminals (VDTs). These people suffer from subjective and objective skin and mucosa-related symptoms, such as itch, heat sensation, pain, erythema, papules and pustules. In severe cases, people can not, for instance, use VDTs or artificial light at all, or be close to mobile telephones. Mast cells (MCs), when activated, release a spectrum of mediators, among them histamine, which is involved in a variety of biological effects with clinical relevance, e.g. allergic hypersensitivity, itch, edema, local erythema and many types of dermatoses. From the results of recent studies, it is clear that EMFs affect the MC, and also the dendritic cell, population and may degranulate these cells. The release of inflammatory substances, such as histamine, from MCs in the skin results in a local erythema, edema and sensation of itch and pain, and the release of somatostatin from the dendritic cells may give rise to subjective sensations of on-going inflammation and sensitivity to ordinary light. These are, as mentioned, the common symptoms reported from patients suffering from 'electrosupersensitivity'/'screen dermatitis'. MCs are also present in the heart tissue and their localization is of particular relevance to their function. Data from studies made on interactions of EMFs with the cardiac function have demonstrated that highly interesting changes are present in the heart after exposure to EMFs. One could speculate that the cardiac MCs are responsible for these changes due to degranulation after exposure to EMFs. However, it is still not known how, and through which mechanisms, all these different cells are affected by EMFs. In this article, we present a theoretical model, based upon observations on EMFs and their cellular effects, to explain the proclaimed sensitivity to electric and/or magnetic fields in humans. PMID- 10859663 TI - Does human betaA4 exert a protective function against oxidative stress in Alzheimer's disease? AB - The hypothesis is advanced that human betaA4--as opposed to rodent betaA4--may exert a protective function against the iron-induced oxidative stress associated with neurological diseases (notably Alzheimer's disease). Subsequent to its release by the host in response to oxidative injury, human betaA4 would interact with Cu(2+)ions whose level is correlatively elevated, adopting the 'aggregated' structure recently characterized by Atwood et al.(15). Then, depending on the oxidative state--hence the pH--of the medium, it might either return to its original structure if physiological pH is restored, or undergo site-specific copper-mediated oxidation and, finally, degradation. In this context, betaA4 pathogenicity could be due to an interfering mechanism preventing the degradation of the oxidized peptide, making its aggregation irreversible and inducing its final deposition. Coordination of side group oxygen donors of the oxidized peptide with 'hard' metal ions occurring in the physiological medium (notably Al(3+)) might be at the origin of this interference. PMID- 10859664 TI - Mental malfunction and memory maintenance mechanisms. AB - Dreams appear to be generated in the process of reinforcing memory circuits of the brain, as circuits are activated by self-generated electrical slow waves, with dream contents reflecting information stored in activated circuits. Illusory dreams and other healthy delirious states appear to occur when activated memory circuits are incompetent, containing synapses whose efficacies deviate from their 'dedicated' values. Organic delirium and some other mental disorders may have their basis in brain pathologies that alter reinforcing slow waves, causing synaptic efficacies to depart from dedicated values. Activation of these incompetent circuits leads to recall of faulty memories--a substrate for delirium. In treatment of organic delirium by electroconvulsive therapy (ECT), the electric shock temporarily suppresses abnormal slow-wave regimes, allowing remedial reinforcement regimes to resume. These restore dedicated synaptic efficacies, temporarily alleviating the delirium. The action of ECT shocks appears to parallel closely that of cardiac defibrillating shocks. Greater than normal amounts of circuit reinforcement protect sensory circuitry in fatal familial insomnia, and cognitive circuitry in encephalitis lethargica. PMID- 10859665 TI - Biological effects of low-level environmental agents. AB - We compare three similar but different biological effects: provocation neutralisation treatment of non-antibody-mediated hypersensitivities, hormesis and low-level effects in radiation biology. All three have not yet been fully explained but share some common and interesting properties: non-linear concentration dependence, typical stress pattern and typical immune response. We try to make a generalisation of the three phenomena in terms of the informational properties of the low concentrations, and imply the possible common mechanism. PMID- 10859666 TI - Does mRNA translation starting from an alternative initiation site contribute to the pathology of Huntington's disease? AB - Huntington's disease is associated with an expanded and unstable trinucleotide repeat (CAG)(n). Various possibilities have been suggested to explain the significance of poly-(CAG) length in HD, including changes in the structure of the product (huntingtin) which result in the protein acquiring deleterious properties. We have looked at the nucleotide sequence coding for huntingtin and find that another possibility may exist for the correlation between the occurrence of HD and poly-CAG length. We have noted an alternative reading frame that includes the trinucleotide repeat, now read as (GCA)(n). Upon close examination of this alternative gene product, we observe features that suggest it can likewise have deleterious properties. PMID- 10859667 TI - Antidromic sequences from replica chromatid disentanglement by flip-rotations. AB - This paper discusses a mechanism whereby a sister replica unit separates from its template via a flip-rotation rather than by eversion. Such an unusual process engenders a permanent antidromic sequence, as well as two (evanescent) kinks only on the stack of the replicated chromatid. PMID- 10859668 TI - NMDA receptor delayed maturation and schizophrenia. AB - This paper presents the hypothesis that NMDA receptor delayed maturation (NRDM) may lead to the pathogenesis of schizophrenic psychotic symptoms. This hypothesis is further analyzed in the language of a neural modeling formulation. This formulation points to a possible chain of pathological events, leading from molecular-level NRDM to over-increased synaptic plasticity, and to the formation of pathological attractors, a putative macroscopic-level correlate of schizophrenic positive symptoms. The relations of the NRDM hypothesis to other alterations which are assumed to take place in schizophrenia are discussed, together with possible ways to test this hypothesis. PMID- 10859669 TI - Pitfalls in prion research. AB - Prion research seems to get increasingly enigmatic since the protein only hypothesis has been established as almost the only working hypothesis. This may indicate that the hypothesis could be wrong, and should prompt the search for potential faults in past experiments. In fact some problematic experiments can be pinpointed, for example determination of the N-terminal cleavage site of the prion protein PrP, of the structure of PrP as determined by NMR, some conclusions concerning the function of PrP from gene knockout experiments including potential evidence against the protein only hypothesis, and some aspects of the prion purification procedure. PMID- 10859670 TI - Possible connection between milk and coronary heart disease: the calcium hypothesis. AB - Excessive milk consumption may adversely affect the circulation on account of the high calcium content of milk and because lactose promotes the intestinal absorption of calcium. Excessive calcium intake may cause calcification and rigidification of the large elastic arteries, which could be an important factor in causing myocardial ischaemia. The calcium hypothesis can throw light on some puzzling peculiarities of arterial disease, for instance the changing ratio of male and female mortality rates in various age groups, the apparently beneficial effect of a warm environment and the entirely different worldwide distribution of coronary heart disease and strokes. PMID- 10859671 TI - The role of genetic factors in the etiology of seasonality and seasonal affective disorder: an evolutionary approach. AB - The degree to which seasonal changes affect mood, energy, sleep, appetite, food preference, or the wish to socialize with other people has been called seasonality. Seasonal affective disorder (SAD), a condition where depressions in fall and winter alternate with non-depressed periods in spring and summer, is the most marked form of seasonality. Several lines of evidence suggest that genetic factors play an important role in the etiology of seasonality and SAD. Millions of years of evolution and adaptation have optimized human biochemical and physiological systems for function and survival under equatorial environmental conditions. Modern humans began their migration out of Africa only about 150 000 years ago. Little change in our 'equatorial' systems might have been expected over this relatively short evolutionary time-span. The author suggests that a genetic susceptibility to seasonal changes in mood and behavior is a genetic predisposition to an insufficient adaptation to temperate and high latitudes. PMID- 10859672 TI - Possible relevance of abnormal fatty acid metabolism in undernutrition: the relationship between oleic acid and growth. AB - Energy, protein, essential fatty acids, vitamins and minerals are all necessary for normal growth and development of children. In undernutrition, there is evidence for abnormal fatty acid metabolism. High levels of oleic acid and low levels of docosahexaenoic acid in plasma phosphatidylcholine have been associated with weight-for-age Z-score shifts to the left (increased prevalence of underweight) and vice versa (reduced prevalence of underweight). An alternative hypothesis is proposed, which describes a possible physiological mechanism whereby omega3 fatty acids contribute to growth. High blood oleic acid levels under conditions of undernutrition are proposed to be an adaptation to conserve glucose in the form of glycogen. Replacement with docosahexaenoic acid under conditions of adequate nutrition enhances membrane functioning so that glucose and energy become available for muscle formation. PMID- 10859673 TI - Colorectal cancer--time as the most important carcinogen: a risky hypothesis about risk. AB - The likelihood of having cancer increases proportionally with a constant power of age, beginning at 25 years old. An equation has been proposed that reproduces this incidence-age relationship, but until now the explanation remains unknown. In this paper, a hypothesis is presented in which tumoral development in human colon and rectum would be the consequence of having exceeded the 'limit number' of mitoses that every stem cell undertakes, from its origin until exhausting the security mechanisms that control the correct replication of genome. The age of cancer appearance would depend on the epithelial turnover time which basically responds to a genetic control. PMID- 10859674 TI - Vascular steal model of human temporal lobe epileptogenicity: the relationship between electrocorticographic interhemispheric propagation time and cerebral blood flow. AB - Human temporal lobe epileptogenicity (i.e. seizure frequency) depends on epileptic and non-epileptic cerebral blood flow (CBF). Increasing non-epileptic cortical CBF is associated with reduction in epileptic cortical CBF. Seizure frequency increases logarithmically with non-epileptic cortical CBF increase and epileptic cortical CBF reduction. A model of human temporal lobe epileptogenicity is derived from the mathematical equivalence to the logarithmic function of seizure frequency of (a) epileptic and non-epileptic CBF differential and (b) electrocorticographic (ECoG) interhemispheric propagation time (IHPT). The vascular steal model of human temporal lobe epileptogenicity suggests that a small CBF redistribution from non-epileptic to epileptic cortex should produce substantial reduction in temporal lobe seizure frequency in association with prolongation of IHPT. The equivalence of these CBF and ECoG parameters to the logarithmic function of seizure frequency suggests that the interhemispheric temporal lobe perfusion gradient and ECoG propagation time may be involved in the fundamental perturbation responsible for human temporal lobe epileptogenicity. PMID- 10859675 TI - Pharmacodynamic principles of homeopathy. AB - Homeopathy was already known to Hippocrates and further studied by Hahnemann. However, since the discovery of the medical effects of digitalis by William Withering around 1785, and the first synthesis of an organic molecule, urea, by Friedrich Wohler in 1828, and through the further rapid evolution of modern pharmacological chemistry and molecular biology, it has gradually been abandoned as a serious therapeutic alternative to allopathy by most practitioners of scientifically founded medicine. Because a credible scientific explanation for its mode of action has been lacking, homeopathy is regarded by many medical researchers and scientists as, at best, placebo therapy, in spite of the fact that for centuries hosts of patients have testified to its effects. It is suggested that the gulf between homeopathy and allopathy can be reconciled if one takes into consideration modern knowledge of physiology, biochemistry and the physical properties of the water used in the potentiation process. A description of the mechanisms occurring during potentiation, both inside and outside a live mammalian organism, is presented. PMID- 10859676 TI - Reactive arthritis: the result of an anti-idiotypic immune response to a bacterial lipopolysaccharide antigen where the idiotype has the immunological appearance of a synovial antigen. AB - The term reactive arthritis (ReA) was first used in 1969 to describe sterile joint disease that follows infection elsewhere in the body. This is an attempt to explain the immunological basis of this disease, give a rationale for the presence of a single bacterial antigen in the involved joints, explain why the class I MHC molecule HLA-B27 is necessary and to suggest possible therapy. This paper proposes an anti-idiotypic (anti-id) model for this disease where a bacterial lipopolysaccharide (LPS) epitope is recognized by idiotypic (Id) T cell receptors and antibody Fab immune recognition surfaces (IRS) which have the immunologic appearance of an antigen on the synovial surface. These Id immune effectors utilize an HLA-B27 molecule to present their IRS on their surface, which results in an anti-id response that can also target the synovial antigen. The anti-id IRS have the immunologic appearance of LPS and their detection in the arthritic joint falsely suggests the presence of bacterial LPS. Evidence is presented which supports this reactive arthritis model in which there is a synovial antigen that is attacked by an anti-id response against the LPS of arthritogenic bacteria. Therapeutic vaccination is supported by this hypothesis. PMID- 10859677 TI - Potential cytotoxic effect of chronic administration of creatine, a nutrition supplement to augment athletic performance. AB - Creatine is alleged to be an ergogenic aid to enhance sports performance and recently became a popular sports nutrition supplement. Although short-term supplementation of creatine has not been associated with major health risks, the safety of prolonged use has caused some concern. The present study demonstrates that creatine is metabolized to methylamine, which is further converted to formaldehyde by semicarbazide-sensitive amine oxidase (SSAO). Formaldehyde is well known to cross-link proteins and DNAs, and known to be a major environmental risk factor. SSAO-mediated production of toxic aldehydes has been recently proposed to be related to pathological conditions such as vascular damage, diabetic complications, nephropathy, etc. Chronic administration of a large quantity of creatine can increase the production of formaldehyde, which may potentially cause serious unwanted side-effects. PMID- 10859678 TI - Neonatal sensitization to latex. AB - Babies born in delivery rooms of hospitals are exposed to latex through skin and mucous membrane contact with prepowdered latex gloves worn by midwives and doctors, and through the inhalation of latex-bound starch powder in the air of the delivery room. This paper examines the hypothesis that they are at risk for latex sensitization, and that part of the sharp increase of childhood asthma, eczema and anaphylaxis in the past 30-40 years may be linked. These possibilities seem hitherto unsuspected. In over 700 papers on latex allergy no mention of neonatal exposure to latex has been found. Even obstetric papers discussing the risks for an atopic mother (atopy - a tendency to develop allergies) do not seem to anticipate any risk for the baby, who might also be atopic. Latex allergy is primarily regarded as an occupational hazard. This paper suggests that it is a hazard for every baby handled by latex gloves at birth. PMID- 10859679 TI - Chemical sensitivity and fatigue syndromes from hypoxia/hypercapnia. AB - The multiple chemical sensitivities syndrome (MCS) and other chronic syndromes causing fatigue, headache and other protean CNS symptoms without observable signs, are proposed to result from hypoxia/hypercapnia (H/H) due to disturbed breathing. The concept is explained in terms of sleep apnea (SA), although H/H could result from causes other than SA. Reasons for considering this etiologic linkage are as follows: 1. MCS symptoms resemble those of SA. 2. The only physical signs associated with MCS (upper airway inflammation and obstruction) can aggravate SA. 3. The only neuropsychiatric finding common among MCS symptomatics, reduced verbal recall, is associated with SA. 4. Many MCS symptomatics attribute onset of their condition to a pesticide or solvent exposure. Solvent neurotoxicity may cause cacosmia, a symptom of MCS and SA. 5. Improved upper airway patency, a first-line therapy in SA, may improve symptoms in some MCS-like conditions. Implications for diagnosis and treatment of MCS are discussed. PMID- 10859680 TI - Can the glyoxylate pathway contribute to fat-induced hepatic insulin resistance? AB - Recent studies have shown that increased hepatic gluconeogenesis is the predominant contributor to fasting hyperglycemia - the hallmark of type 2 diabetes. Although it has been known for a long time that over-supply of fat is able to stimulate gluconeogenesis both in-vitro and in-vivo, neither the leading substrate nor the mechanism responsible for this phenomenon have been fully identified. Recent observations that the glyoxylate pathway may exist in animals has shed light on this question. The glyoxylate pathway is able to convert fatty acid into glucose but has been thought to be absent in animals. Although further evidence is needed, current available data does suggest a possible mechanism which, by integrating both glucose and lipid metabolism together rather than interpreting them separately, may explain the role of fatty acids in hepatic insulin resistance. This hypothesis is based on current understanding of insulin resistance and supported by many laboratory observations. PMID- 10859681 TI - Another integrated form of the Michaelis-Menten equation, its analogy to electrical circuit model and implications for active transporters. AB - The Michaelis-Menten equation is integrated versus concentration to study active transporters. The integrated equation is analogous to compartmental and electrical circuit models. Simulation of the integrated equation suggests that an active transporter can run backwards and produce energy. PMID- 10859682 TI - A new look at the challenging world of tandem repeats. AB - Recent research has shown a correlation between some genetic diseases and genomic sequences tandemly repeated a variable and excessive number of times. The excessive number of tandem repeats is usually caused by a progressive expansion, generally considered as purely harmful. We put forward a number of hypotheses: the main one is that the number of repeats has normally a specific significance, and that there exist purposive mechanisms having as a primary function the management of tandem repeats length; such a function is generally useful and only rarely may it become harmful, because of some malfunctioning. These hypotheses are suggested by plausibility arguments, and are supported by a number of recent experimental results. They could provide a simple and unifying explanation of many pathological and non-pathological phenomena replacing many ad hoc assumptions. We finally propose to call the study of the above tandem repeat managing mechanisms 'dynamical genetics'. PMID- 10859683 TI - The evolution of sleep: a reconsideration of the development of the quiet sleep/active sleep cycle. AB - Quiet sleep (QS) is usually assumed to have evolved before active sleep (AS), because early studies of the primitive mammal echidna indicated that it experiences QS only. Theories designed to account for the development of AS therefore usually focused on the adaptive advantages of a QS/AS cycle over QS alone. There are several conceptual and empirical problems with those theories, however. Moreover, recent data indicate that a QS/AS cycle is extant in all mammals and birds. Empirical and conceptual evidence supporting the notion that AS developed earlier than previously thought, and that AS could have preceded QS in evolution, are discussed here. PMID- 10859684 TI - Is quantum brain dynamics involved in some neuropsychiatric disorders? AB - In this paper we review the evidence for quantum phenomena underlying neuropsychiatric disorders. Existing data can be explained only by resorting to non-local correlations within brain activity, such as the ones predicted by quantum theory. Such a situation suggests that perhaps a quantum description may be the best description of interrelationships between neural and cognitive phenomena. PMID- 10859685 TI - Alzheimer's disease revisited. AB - In a previous paper, it was suggested that a relative deficiency of essential fatty acids might play a role in the etiology of sporadic or non-familial Alzheimer's disease. A recent article regarding dementia in the Rotterdam Study reinforces this suggestion. It is also hypothesized that this relative deficiency could facilitate passage of aluminum into the brain, aluminum being increasingly suggested as one of the possible pathogenic factors in AD. It is further suggested that hypomethylation caused by a deficiency of S-adenosylmethionine might also play a role in the etiology of this disease and perhaps even of Parkinson's disease. PMID- 10859686 TI - Importance of immune deviation toward Th1 in the early immunopathogenesis of human T-lymphotropic virus type I-associated myelopathy. AB - Although the principal neuropathological feature of human T-lymphotropic virus type I (HTLV-I)-associated myelopathy (HAM) is chronic inflammation of the spinal cord, characterized by perivascular cuffing of mononuclear cells accompanied by parenchymal lymphocytic infiltration, the precise mechanisms by which HTLV-I infection causes chronic inflammation of the spinal cord are still obscure. In patients with HAM, peripheral blood CD4(+)T lymphocytes, particularly HTLV-I infected CD4(+)T lymphocytes, have increased adherent activity to endothelial cells and transmigrating activity through basement membranes. In addition, the profile of cytokine expression suggests increased numbers of Th1 cells in peripheral blood CD4(+)T lymphocytes of patients with HAM. These findings strongly suggest that immune deviation toward Th1, which might be based on high viral load of HTLV-I, plays an important role in tissue damage in the central nervous system of patients with HAM. We herein emphasize the importance of activated Th1 cells as the first trigger in the immunopathogenesis of HAM. PMID- 10859687 TI - Effect of earth's orbital chirality on elementary particles and unification of chiral asymmetries in life on different levels. AB - Life is chirally asymmetric at all scales from microscopic elementary particles to molecular and macroscopic levels. How these chiral asymmetries in life on different levels are unified remains unanswered. It has been demonstrated that both the biomolecular homochirality and biological rhythms can be caused by the right-handed helical force-field of the Earth's orbital chirality (EOC). Similar to the helical biomolecules (1), it is here suggested that the right-handed EOC force-field could make the right-handed elementary particles more stable than their left-handed enantiomers to result in the symmetry violation of elementary particles, and the EOC could also cause the macroscopic predominant selection of right-handed asymmetries of living objects (e.g. the helical seashells and plants). Our studies indicated that the weak force in weak interaction may only be a form of the EOC force-field at the microscopic particle level, and the chiral asymmetries in life on various levels could be unified by the natural right-handed EOC force-field. Moreover, the chiral and quantum effects, time, mass, rhythms and relativity could also be unified by the interaction of the EOC force-field with chiral motions and structures under certain conditions. PMID- 10859688 TI - Toward practical prevention of type 2 diabetes. AB - Even in individuals who are unwilling to make prudent changes in their diets and sedentary habits, the administration of certain nutrients and/or drugs may help to prevent or postpone the onset of type 2 diabetes. The evident ability of fiber rich cereal products to decrease diabetes risk, as documented in prospective epidemiological studies, may be mediated primarily by the superior magnesium content of such foods. High-magnesium diets have preventive (though not curative) activity in certain rodent models of diabetes; conversely, magnesium depletion provokes insulin resistance. Epidemiology also strongly suggests that regular moderate alcohol consumption has a major favorable impact on diabetes risk, particularly in women; this may reflect a direct insulin-sensitizing effect on muscle and, in women, a reduced risk for obesity. Chromium picolinate can also aid muscle insulin sensitivity, and initial reports suggest that it is an effective therapy for type 2 diabetes. High-dose biotin has shown therapeutic activity in diabetic rats and in limited clinical experience; increased expression of glucokinase in hepatocytes may mediate this benefit. Other nutrients that might prove to aid diabetic glycemic control, and thus have potential for prevention, include coenzyme Q and conjugated linoleic acids (CLA). Since the nutrients cited here - including ethanol in moderation - appear to be quite safe and (with the exception of CLA) quite affordable, supplementation with these nutrients may prove to be a practical strategy for diabetes prevention. Drugs such as metformin and troglitazone, which are expensive and require regular physician monitoring to avoid potentially dangerous side-effects, would appear to be less practical options from cost-effectiveness, convenience and safety standpoints, given the fact that the population at-risk for diabetes is huge. PMID- 10859689 TI - The insulin-sensitizing activity of moderate alcohol consumption may promote leanness in women. AB - Cross-sectional epidemiology reveals that women who drink alcohol regularly and moderately, on average, tend to have a decidedly lower body-mass index (BMI) than non-drinking women, despite slightly higher caloric intakes. In men, moderate drinkers are no heavier than non-drinkers, yet they consume considerably more calories. The thermogenic effect which this implies is not explained by the modest acute thermic effect of ethanol ingestion. However, there is indirect evidence that regular alcohol consumption has an insulin-sensitizing effect on skeletal muscle that down-regulates insulin secretion. Decreased insulin activity on adipocytes and the liver may discourage fat storage and promote hepatic mechanisms of ketogenesis, gluconeogenesis, and associated thermogenesis, thus possibly accounting for the relative leanness of female drinkers. The possibility that prescribing moderate alcohol intake could aid weight control in non-drinking overweight females should receive clinical evaluation. The impact of moderate drinking on risk for diabetes in women appears to be quite dramatic. PMID- 10859690 TI - Sulfated glycosaminoglycans and glucosamine may synergize in promoting synovial hyaluronic acid synthesis. AB - High-molecular-weight hyaluronic acid (HA) produced by the synovium may function physiologically to aid preservation of cartilage structure and prevent arthritic pain; both the size and concentration of HA in synovial fluid are diminished in osteoarthritis (OA). Glucosamine therapy for OA can be expected to increase synovial HA production by providing rate-limiting substrate. In addition, certain sulfated glycosaminoglycans and polysaccharides - including chondroitin sulfate (CS), dermatan sulfate, and pentosan polysulfate - stimulate synovial HA production, apparently owing to a hormone-like effect triggered by the binding of these polymers to membrane proteins of synovial cells. Surprisingly, a significant proportion of orally administered CS is absorbed as intact polymers - apparently by pinocytosis. These considerations may rationalize clinical studies concluding that oral CS provides slow-onset but durable pain relief and functional improvement in OA. The possibility that oral glucosamine and CS may interact in a complementary or synergistic fashion to improve synovial fluid HA content in OA should be assessed in clinical studies, and the potential of adjunctive CS administration to improve the clinical response achievable with optimal intakes of glucosamine should likewise be evaluated. In light of the fact that the synovium virtually functions as a 'placenta' for cartilage, focusing on synovium as the target for therapeutic intervention in OA may be a rational strategy. PMID- 10859691 TI - High-dose pyridoxine as an 'anti-stress' strategy. AB - Pyridoxine nutritional status has a significant and selective modulatory impact on central production of both serotonin and GABA - neurotransmitters which control depression, pain perception, and anxiety - owing to the fact that the decarboxylases which produce these neurotransmitters have a relatively low affinity for pyridoxal phosphate (PLP). Pyridoxine deficiency leads to increased sympathetic outflow and hypertension in rodents, possibly reflecting decreased central production of these neurotransmitters; conversely, supplemental pyridoxine lowers blood pressure in many animal models of hypertension, and there is preliminary evidence for antihypertensive activity in humans as well. Additionally, physiological levels of PLP interact with glucocorticoid receptors to down-regulate their activity. Thus, high-dose pyridoxine, by amplifying tissue levels of PLP, may be expected to have a favorable impact on certain dysphoric mental states, while diminishing sympathetic output and acting peripherally to blunt the physiological impact of corticosteroids. In light of growing evidence that chronic dysphoria, particularly when accompanied by hopelessness or cynicism, has a major negative impact on morbidity and mortality from a wide range of disorders, high intakes of pyridoxine may have the potential to improve prognosis in many individuals. With respect to cardiovascular health, reduction of homocysteine levels should contribute to this benefit. These predictions are consistent with recent epidemiology correlating plasma PLP levels with risk for vascular events and overall survival. PMID- 10859692 TI - Prenatal high-dose pyridoxine may prevent hypertension and syndrome X in-utero by protecting the fetus from excess glucocorticoid activity. AB - The increased risk for hypertension, insulin resistance syndrome, and coronary events associated with small-for-gestational-age birth, has plausibly been attributed to excessive prenatal exposure to glucocorticoids; this may up regulate glucocorticoid activity throughout life by permanently decreasing expression of hippocampal glucocorticoid receptors crucial for feedback control of cortisol secretion. Since pyridoxal phosphate is a safe physiological antagonist of glucocorticoid activity, it is proposed that prenatal supplementation with high-dose pyridoxine may counteract the adverse impact of glucocorticoids on fetal growth, as well as on subsequent cardiovascular risk. PMID- 10859693 TI - Etiology, pathogenesis, and experimental treatment of retinitis pigmentosa. AB - The paper provides an interdisciplinary evaluation of the etiology, pathogenesis, and experimental treatments of retinitis pigmentosa (RP). It addresses a 10-year controversy concerning the rate of progression of RP. One laboratory has estimated remaining visual field to be lost at a rate of 4.6% per year, whereas another laboratory estimates loss at 16-18%. This large discrepancy and lack of consensus needs resolution, since they pose serious statistical and operational problems for evaluating experimental treatment approaches to RP. The resolution of the controversy offered in the paper is based on a model of RP in which the initial rate of loss of visual field (the induction phase) is much slower than the subsequent logarithmic first-order rate of loss. The rationale for this kinetic model is that loss of mitochondrial function, possibly due to RP genetically-related radical processes, has to reach a critical threshold value before the mitochondrial trigger of programmed cell death or apoptosis (i.e., the release of mitochondrial cytochrome c by the opening of the permeability transition pore, PTP) can be activated by an encounter with a second, but kinetically constant causative stress factor - most likely a light-stress-related factor. In its essential (two-causal) aspects, this kinetic model for RP is identical to the kinetic theories that have been proposed for the Gombertz human mortality plot. The described kinetic model for RP provides a solution to the visual field-loss controversy, since the first study was performed with a population containing a greater number of patients in the slow stage of RP than the second. Another objective of the investigation was to identify possible mechanisms of how the numerous genetic mutations in the rods of RP patients could give rise to damaging free-radical reactions capable of triggering apoptosis through their adverse effects on mitochondrial function. Another reason for focusing on radical reactions in RP was to provide a rationale for the proposed use of an extensive array of antioxidants and nutritional supplements for stemming progression of RP. In particular, the investigation focuses on saving cone-dependent central vision, i.e. on saving cells not affected by the genetic problems of the rods, but cells which can become lethally damaged by a spill-over of radicals and related harmful chemical reactions occurring in the rods.The third objective deals with the development of a rationale for a new strategy for retarding RP. This involves the use of desmethyldeprenyl, a metabolite of the anti-Parkinson's drug, deprenyl. The rationale is, in part, based on an observation that desmethyldeprenyl exerts antiapoptotic activities in a variety of neurodegenerative disorders. The protective mechanism involves the overexpression of the anti-apoptotic bcl-2 gene, leading to higher concentrations of bcl-2 proteins, which by binding to mitochondria inhibits the trigger mechanism of apoptosis - the opening of PTP and release of cytochrome C. At the same time, desmethyldeprenyl causes the underexpression of the pro-apoptotic bax gene, which via bax proteins facilitates the opening of the PTP. Both the anti apoptotic and pro-apoptotic mechanisms appear to be mediated by the binding of desmethyldeprenyl to glyceraldehyde-3-phosphate dehydrogenase. Antiapoptotic effects can also be generated by the parent compound, deprenyl, when this is used daily in low concentrations of 1-2 mg/100 kg body weight. Under these conditions, it appears that the anti-apoptotic metabolite, desmethyldeprenyl, predominates over the pro-apoptotic metabolites of deprenyl, l -methamphetamine and l amphetamine. Methamphetamine is not formed if desmethyldeprenyl is administered directly and thus could give desmethyldeprenyl a pharmacokinetic advantage over deprenyl. (ABSTRACT TRUNCATED) PMID- 10859694 TI - Non-communicable diseases: is their emergence in industrialized societies related to changes in neuroendocrine function? AB - This hypothesis suggests that industrialization alters the human neuroendocrine system. The neuroendocrine changes come about because of changes in environmental stimuli. It is further proposed that changes in neuroendocrine function can account for the contrasting pattern of non-communicable diseases in traditional and industrialized societies. The hypothesis is based on subtle clinical differences in traditional and industrialized societies, and the evolving concept of neuroendocrine regulation of physiological processes. Compared to traditional societies, individuals from industrialized communities tend to have lower pain tolerance, slower gastrointestinal transit-time, and a greater chance of having a calcified pineal gland. These changes parallel the increasing incidence of non communicable diseases in industrialized societies. There is sufficient reason to suspect the variations in pain tolerance, gastrointestinal transit-time and pineal gland calcification represent changes in neuroendocrine function. Programming of the neuroendocrine system by environmental events early in life is one possible mechanism whereby these changes might be effected. Understanding the physiological changes that occur with industrialization, and how environmental stimuli interact with the developing neuroendocrine system might lead to new strategies for the prevention and treatment of non-communicable diseases. PMID- 10859695 TI - Acid-dependent dismutation of nitrogen oxides may be a critical source of nitric oxide in human macrophages. AB - The cytotoxic activity of the macrophage relies greatly on the secretion of a number of reactant intermediates, including superoxide (O(2)(-)), hydroxyl radical (OH(-)) and nitric oxide (NO). The latter, synthesized via cytokine mediated induction of inducible NO-synthase (iNOS), is readily observed in murine macrophages. However, a poorly reproducible or minimal response to cytokine stimulation in the human macrophage has questioned the presence or significance of this important pathway in man. Nevertheless, iNOS is present in other human phagocytic cells, e.g. neutrophils, while the NO metabolites, nitrite (NO(2)(-)) and nitrate (NO(3)(-)), are raised in human serum during infection. Low phagolysosomal pH is critical for the macrophage to destroy the engulfed pathogen. This acidic environment may allow synthesis of NO independently of iNOS via dismutation of NO(2)(-)to NO. Should this mechanism be active, assay for iNOS and NO by determination of NO(2)(-)could be misleading. In human macrophages, acid-induced conversion of imported nitrogen oxides (NOx) may take precedence over iNOS-mediated NO synthesis and should be investigated as a source of NO in these cells. PMID- 10859696 TI - Synergistic approach to cancer therapy: exploiting interactions between anti estrogens, retinoids, monoterpenes and tyrosine kinase inhibitors. AB - Non-responsiveness and toxicity are large problems encountered during cancer treatment. Utilization of compounds that synergize should increase treatment efficacy while avoiding problems of toxicity. This review explores interactions between classes of compounds, including anti-estrogens, retinoids, monoterpenes and tyrosine kinase inhibitors, that are effective independent, and how their synergistic interaction could be exploited in cancer treatment. The effects of these compounds on insulin-like growth factors (IGF) and transforming-growth factor-beta (TGF-beta) will also be examined. PMID- 10859697 TI - The marine toxin domoic acid may affect the developing brain by activation of neonatal brain microglia and subsequent neurotoxic mediator generation. AB - Amnesic shellfish poisoning, one of the shellfish poisoning syndromes, is caused by the marine diatom toxin domoic acid (DOM). While in adult rats, mice, monkeys and humans DOM poorly penetrates the blood-brain barrier, DOM has been shown to be very toxic to fetal in newborn mice, because the blood-brain barrier is incomplete during neurodevelopment. This fact may explain why neonates show a higher sensitivity to neurotoxins like DOM as compared to adult animals. Mechanistic studies on DOM's neurotoxicity have mainly concentrated on the investigation of DOM's effect on neuronal tissue. Recent studies have shown that glia is also involved in DOM's neurotoxicity to the adult as well as the developing nervous system. The scientific literature strongly supports the hypothesis that the microglia may play a critical role in mediating DOM's neurotoxic effects. However, the effect of DOM on microglia has not been systematically investigated. The literature supporting our hypothesis is presented and discussed. PMID- 10859698 TI - Hepatic osteodystrophy: the influence of liver disease and portal hypertension on cytokine activation. AB - Chronic liver disease is frequently complicated by bone disease. The mechanisms leading to bone-loss are unknown but are probably not related to abnormal vitamin D metabolism. The effects of portal hypertension and porto-systemic shunting on bone-loss have not been studied. It is postulated that liver disease, or the complications of liver disease (portal hypertension or porto-systemic shunting), might be responsible for the activation of cytokines. It is further postulated that these cytokines might be the final common pathway leading to bone-loss in both parenchymal and cholestatic liver disorders by modifying osteoblast or osteoclast function. PMID- 10859699 TI - Do NSAIDs contribute to acute fatty liver of pregnancy? AB - Acute fatty liver of pregnancy (AFLP) and the childhood encephalopathy known as Reye's syndrome are both characterised by microvesicular steatosis. Mothers with AFLP are frequently heterozygous for a mutation which reduces the activity of the trifunctional protein (TP) of fatty-acid oxidation. Several lines of evidence suggest that blockade of fatty-acid oxidation may also be the underlying cause of Reye's syndrome, and epidemiological studies have identified aspirin taken during a viral illness as a contributing factor to the development of the disease. The hypotheses are presented:* that children with Reye's syndrome may also be heterozygous for TP mutation, and* that inhibition of the residual long-chain fatty-acid oxidation by NSAIDs including aspirin precipitates the similar symptoms observed in patients with Reye's syndrome and AFLP. Identification of NSAIDs as candidates for the unidentified factor which precipitates AFLP suggests that avoidance of NSAIDs during pregnancy may lead to a reduction in the incidence of this life-threatening disease. PMID- 10859700 TI - Myosin regulatory light chain as a critical substrate of cell death: a hypothesis. AB - As known, gamma-interferon-induced cell death in HeLa cells can be mediated via the recently identified protein kinase, death associated protein (DAP) kinase which is localized to the microfilament system of the cytoskeleton. However, the downstream or upstream effectors of DAP kinase remain uncertain. In the present work, we hypothesize that the most probable substrate for DAP kinase is regulatory light chain of myosin II, by phosphorylating which the kinase can transduct death signals. PMID- 10859701 TI - Chronic fatigue syndrome: a matter of enzyme deficiencies? AB - The etiology of chronic fatigue syndrome (CFS) remains an enigma. But literature concerning chronic fatigue which does not focus on CFS points to all sorts of enzyme deficiencies as possible causes. The deficiencies are probably dismissed as causes of CFS because other characteristic symptoms are lacking in CFS patients. But these symptoms are often also lacking in patients with a deficiency. Symptom patterns in enzyme deficiencies are extremely variable. Therefore, patients with CFS should be screened systematically for enzyme deficiencies. PMID- 10859703 TI - Long term follow-up of neonates. PMID- 10859702 TI - The ratio of 2nd to 4th digit length: a new predictor of disease predisposition? AB - The ratio between the length of the 2nd and 4th digits is: (a) fixed in utero; (b) lower in men than in women; (c) negatively related to testosterone and sperm counts; and (d) positively related to oestrogen concentrations. Prenatal levels of testosterone and oestrogen have been implicated in infertility, autism, dyslexia, migraine, stammering, immune dysfunction, myocardial infarction and breast cancer. We suggest that 2D:4D ratio is predictive of these diseases and may be used in diagnosis, prognosis and in early life-style interventions which may delay the onset of disease or facilitate its early detection. PMID- 10859704 TI - Outcomes of children of extremely low birthweight and gestational age in the 1990s. AB - Advances in perinatal care have improved the chances for survival of extremely low birthweight (<800 grams) and gestational age (<26 weeks) infants. A review of the world literature reveals that among regional populations, survival at 23 weeks' gestation ranges from 2 to 35%, at 24 weeks' gestation 17 to 62% and at 25 weeks' gestation 35 to 72%. These wide variations may be accounted for by differences in population descriptors, in the criteria used for starting or withdrawing treatment, in the reported duration of survival and differences in care. Major neonatal morbidity increases with decreasing gestational age and birthweight. At 23 weeks' gestation, chronic lung disease occurs in 57 to 86% of survivors, at 24 weeks in 33 to 89% and at 25 weeks' gestation in 16 to 71% of survivors. The rates of severe cerebral ultrasound abnormality range from 10 to 83% at 23 weeks' gestation, 9 to 64% at 24 weeks and 7 to 22% at 25 weeks' gestation Of 77 survivors at 23 weeks' gestation, 26 (34%) have severe disability (defined as subnormal cognitive function, cerebral palsy, blindness and/or deafness). At 24 weeks' gestation, the rates of severe neurodevelopmental disability range from 22 to 45%, and at 25 weeks' gestation 12 to 35%. When compared with children born prior to the 1990s, the rates of neurodevelopmental disability have, in general, remained unchanged. We conclude that, with current methods of care, the limits of viability have been reached. The continuing toll of major neonatal morbidity and neurodevelopmental handicap are of serious concern. PMID- 10859705 TI - Follow-up of very low birthweight babies to adolescence. AB - Reports on the school-age outcomes and behavioural difficulties at adolescence of infants who were very low birthweight (VLBW) are only just emerging. Studies which compare VLBW with same age controls consistently show significantly poorer performance, with average scores between 8 and 13 points lower. Even children with no neurological impairments have scores which are significantly lower on cognitive and achievement measures. The extremely low birthweight (ELBW) adolescents fare worse on all measures and perform particularly poorly in mathematics. A high proportion of VLBW adolescents (15%-20%), and an even higher proportion of ELBW adolescents (30%-50%), are receiving remedial assistance and/or have failed a grade. There are conflicting reports on whether the behavioural problems increase or improve with age, but most studies show that at adolescence the problems are still significantly greater in the VLBW cohort than in their peers. Methodologically rigorous studies of the current survivors to school-age should be conducted to determine whether the technological innovations in the 1990s have contributed to a reduction in psychoeducational and behavioural difficulties. Future research should also be directed towards early identification of school difficulties and development of intervention strategies targeted to the most vulnerable infants. PMID- 10859706 TI - Measuring respiratory outcome. AB - Chronic respiratory morbidity is a common outcome of very premature birth. Infants who are chronically oxygen dependent with an abnormal chest radiograph are described as suffering from chronic lung disease (CLD), and those with the worst abnormalities diagnosed as having bronchopulmonary dysplasia. CLD infants are very likely to be readmitted to hospital during infancy, particularly during a respiratory syncytial virus (RSV) epidemic. Very low birthweight, prematurity and CLD are associated with recurrent respiratory symptoms and lung function abnormalities during the preschool years. These problems are detected even in adolescents who were chronically oxygen dependent after premature birth. Further research to identify effective preventative strategies is urgently required. PMID- 10859707 TI - Outcome after intrapartum asphyxia in term infants. AB - Investigatory techniques, particularly magnetic resonance (MR) imaging and spectroscopy, performed in the early neonatal period on infants suspected of intrapartum asphyxia i.e. abnormal fetal heart recording, poor cord gases, low Apgar scores and the need for resuscitation, or with neonatal encephalopathy or seizures, have allowed a much better understanding of the patterns of brain injury and the biochemical processes that follow these events. It is usually possible to distinguish these patterns from those seen in other, often confounding, diagnoses. This has allowed far more precision about the timing of insults and in the prediction of particularly motor, feeding and visual outcome and to some extent intellectual outcome. Long-term neurological and psychometric follow-up of infants in whom detailed perinatal clinical histories and examination, haematological and biochemical investigation and MR brain scans are obtained will allow even more accurate prediction of outcome in the future. Such studies also help to validate standardized neonatal and infant clinical neurological examinations, making them useful tools to predict outcome. PMID- 10859708 TI - Methodological issues in randomized controlled trials. AB - There is increasing emphasis on the need to practise evidence-based medicine and the strongest evidence comes from well designed and well-conducted randomized controlled trials. Every component is important for the success of a clinical trial; if the design or sample size is inappropriate, then the results of the study will be unreliable, however well the study is conducted. Conversely a well designed study may founder because of poor outcome measurement or unacceptably high subject loss. The advantages of a well-designed trial apply equally to studies with short term outcomes and to those requiring long-term follow up. This paper therefore focuses on general methodological issues with a discussion, where appropriate, of the special considerations associated with long-term follow-up. This emphasis is motivated by the belief that a trial with methodological weaknesses is both a waste of resources and unethical. Anyone planning to undertake a randomized controlled trial should consult a more comprehensive text [1-4]. Here, some selected issues are highlighted with the choice of topics reflecting the experience and interests of the authors. PMID- 10859709 TI - Measurement of health status and quality of life in neonatal follow-up studies. AB - Mortality and neuro-developmental outcome can be precisely measured but in order to interpret the significance of changes in them, further information is needed about health status, the effect of health status on lifestyle of survivors and their families, the quality of life experienced by survivors and the value placed by the public and survivors on different health states. The ability to measure such aspects of health now allows more relevant follow-up studies to be designed. This article discusses concepts of health status and quality of life and the problems in applying these to children. There follows a critique of modern instruments for measuring health status and quality of life and their application in neonatal follow-up studies. We recommend that only a small number of well established instruments should be considered so that results are valid and can be compared with other studies. PMID- 10859710 TI - Economic issues in the follow-up of neonates. AB - Children with conditions requiring neonatal intensive care impose a financial burden on health services, on families and carers, and on society generally. A systematic review of the literature identified 81 studies that conducted primary research on the cost of services as a result of conditions requiring neonatal intensive care. The majority of studies estimated costs incurred during the initial hospital stay. Relatively few studies considered health service costs following discharge from the neonatal intensive care unit, costs to other sectors of the economy or costs to families and carers. It is important that these costs are considered more fully. PMID- 10859711 TI - Parent concerns in long-term follow-up. AB - Consideration of the long-term follow-up of at-risk infants must take into account the role of parents and family contexts. Cognitive and social-emotional adaptation for a premature low birthweight infant is a product of complex transactions between biological and environmental risk and ameliorative factors that operate within powerful family and cultural contexts. Parental behaviors, psychosocial functioning, and social cognitions are particularly important in order to understand long-term developmental outcomes for infants as well as other family members. Interventions for high-risk infants have shown that these can be effective in reducing anxiety and concerns among parents and optimizing parent child relationships. PMID- 10859712 TI - Primary stenting for acute myocardial infarction via the transradial approach: a safe and useful alternative to the transfemoral approach. AB - BACKGROUND: Primary stenting in acute myocardial infarction (AMI) has been demonstrated to reduce recurrent ischemic events. However, transradial stenting in AMI has not been well established. Therefore, we sought to investigate the feasibility and utility of transradial coronary stenting in patients with AMI. METHODS: From April 1998 to April 1999, 56 patients (43 male; mean age of 57 years) who arrived within 6 hours of pain onset with culprit vessel size > 2.5 mm constituted this study. The transradial approach (Group 1) was used in 30 patients with hemodynamically stable and palpable right radial pulse. The transfemoral approach (Group 2) was used for vascular access in the remainder of patients (26) who might have required a second vascular access site for intraaortic balloon pumping (in cardiogenic shock) and/or a transvenous temporary pacemaker. RESULTS: Overall success rate was achieved in 54 of 56 patients (96%). The success rate was 90% (27/30) in Group 1 and 96% (25/26) in Group 2. The cannulation time (from patient arrival at the catheterization room to the time of arterial cannulation) and the total procedure time (from patient arrival at the catheterization room to the completion of the procedure) were not significantly different between Group 1 and Group 2 (9.2+/-5.3 versus 8.9+/-5. 8 minutes, p>0.05; 53.7+/-19.4 versus 57.5 +/-26.8 minutes, p>0.05, respectively). In the Group 1 patients, there was no forearm ischemia or loss of radial pulse during the 30-day follow-up period. CONCLUSION: Primary coronary stenting for acute myocardial infarction via the transradial approach is a safe and feasible alternative to the conventional transfemoral approach, and is especially useful for hemodynamically stable patients who do not require a second vascular access site. PMID- 10859713 TI - Percutaneous radial approach: isn't it time we put it in its place? PMID- 10859714 TI - Left internal mammary artery intervention: the left radial approach with a new guide catheter. AB - Coronary intervention involving left internal mammary bypass grafts is an increasingly common challenge for the interventionalist. Although successful in a high percentage of patients, the femoral approach may be technically challenging. The shorter, more direct approach from the left radial artery has potential advantages in these cases. The present study evaluated our experience using a new left transradial internal mammary guide catheter. Angiography alone was successfully performed with the catheter in 40 patients. In an additional 10 patients, internal mammary artery interventional procedures were performed. Angioplasty alone was performed in five patients. Five patients underwent coronary stenting within the internal mammary artery or native left anterior descending artery. Guide catheter backup was satisfactory and angioplasty catheters and/or stents could be advanced to the target lesion with minimal difficulty. No procedural complications occurred. The left radial artery access site is an excellent approach for left internal mammary intervention. PMID- 10859715 TI - The association between institutional primary angioplasty procedure volume and outcome in elderly Americans. AB - BACKGROUND: The association between greater procedure volume and improved patient outcome in cardiac procedures has been established in percutaneous transluminal coronary angioplasty (PTCA), coronary stent placement and coronary bypass surgery. The association between primary angioplasty volume and outcome has not been evaluated. METHODS: We evaluated the association between the volume of primary angioplasty procedures with short- and long-term outcome in 6,124 patients with documented acute myocardial infarction. Patients without shock on presentation treated with primary coronary angioplasty within 12 hours of hospital admission were selected from consecutive infarct patients included in the Cooperative Cardiovascular Project database. Patients were divided into quartiles based on the volume of primary PTCA procedures performed at their admitting hospital. RESULTS: The majority of United States (US) hospitals performed less than three primary PTCA procedures per month. Patients admitted to hospitals in the lowest volume quartile of primary PTCA had 31% higher 30-day mortality than those admitted to the highest volume quartile. After adjustment for baseline differences in patient characteristics, there was an association between admission to higher volume primary PTCA hospitals and lower 30-day mortality (odds ratio per volume quartile = 0.91; 95% confidence interval = 0.83 0.99). CONCLUSION: Eighty-two percent of US hospitals perform less than three primary PTCA procedures per month. In elderly Americans treated with primary PTCA, we observed an association between admission to higher volume hospitals and lower short- and long-term mortality. This association was independent of total PTCA volumes. PMID- 10859716 TI - Expertise in primary angioplasty--implications for optimal patient care. PMID- 10859717 TI - Angiographic and clinical follow-up of percutaneous revascularization for transplant coronary artery disease. AB - BACKGROUND: There are limited data on the use of percutaneous revascularization techniques for transplant coronary artery disease (CAD). METHODS: Medical records and angiographic results for cardiac transplant patients undergoing percutaneous revascularization at Emory University Hospital were reviewed. Procedural results, results of angiography 4Eth 6 months after intervention, and clinical follow-up were recorded. RESULTS: Nineteen patients underwent 51 interventions. Thirty eight lesions (75%) were de novo and 13 (25%) were restenotic. All patients had hypertension, 37% had diabetes, 79% had elevated lipid levels, and 53% had at least one episode of moderate to severe allograft rejection (grade 3A or greater). The primary procedural success rate was 100% with no major complications. Six-month restenosis rate (defined as > 50%) was 49%. At 23+/-17 months follow-up, 6 patients were dead or retransplanted (31%). Thirteen patients were alive without retransplantation (9 New York Heart Association class I, 3 class II, 1 class III). CONCLUSION: Percutaneous revascularization is safe and has a high initial procedural success rate in patients with transplant CAD. However, the restenosis rate in this population remains higher than reported for atherosclerotic coronary disease and the long-term prognosis remains poor. PMID- 10859718 TI - To intervene or not in transplanted coronary artery disease. PMID- 10859719 TI - What have We learned from ESPRIT? What will we learn from TARGET? PMID- 10859720 TI - Conservative management for an extensive type A aortic dissection complicating coronary angioplasty. AB - Aortic dissection is a recognized, though rare complication of percutaneous revascularization procedures. We report a case of an extensive type A dissection that occurred during an attempt to recanalize a chronic total occlusion of a right coronary artery. The patient was treated conservatively and was followed for 36 months, during which he remained well. We conclude that, even though surgery remains the preferred option, conservative management could also be considered in certain patients. PMID- 10859721 TI - Transient obstruction of the mitral valve by a massive thrombus complicating percutaneous valvuloplasty. AB - Percutaneous transseptal mitral valvuloplasty was attempted in a patient with severe rheumatic mitral stenosis. Introduction of the balloon into the left atrium resulted in thrombotic occlusion of the mitral valve. Circulatory arrest ensued. The thrombus was disrupted during cardiopulmonary resuscitation. Thrombolysis was administered and the patient recovered uneventfully. PMID- 10859722 TI - Severe, resistant spasm of the left main coronary artery--a serious pitfall. AB - A case of severe, resistant spasm of the left main coronary artery, which was not relieved even after 600 micrograms of intracoronary nitroglycerine over 30 minutes, is described. The case was mistakenly taken for fixed stenosis and would have been subjected to percutaneous transluminal coronary angioplasty with stenting at the same sitting, had not the case been fortuitously deferred. On repeat angiography after one week, the left main was found to be normal. Some guidelines to avoid such a situation are suggested. PMID- 10859723 TI - The SCRIPPS trial--catheter-based radiotherapy to inhibit coronary restenosis. AB - The SCRIPPS trial is a randomized, double-blind, placebo-controlled trial evaluating the impact of gamma radiation to inhibit in-stent restenosis. Fifty five patients were enrolled; twenty-six were assigned to receive catheter-based radiation with Ir-192 and 29 were treated with placebo. Angiographic restenosis in the Ir-192-treated patients was significantly reduced at six months (17% vs. 54%; p = 0.01), with the results sustained at 3-year follow-up (33.3% vs. 63. 6%; p<0.05). Likewise, the composite clinical endpoint of death, myocardial infarction or target lesion revascularization was also commensurately lower in the treated group as compared to the placebo group (23.1% vs. 55.2%; p = 0.01). Late angiography revealed no perforation, aneurysm or pseudoaneurysm or special safety issues unique to radiotherapy. PMID- 10859724 TI - Aorto-uni-iliac conversion of the bifurcated AneuRx device: a technical note. PMID- 10859725 TI - Preventive health care, 1999 update: prevention of oral cancer mortality. The Canadian Task Force on Preventive Health Care. AB - BACKGROUND: Approximately 3,000 new cases of oral cancer are diagnosed each year in Canada. Most of these cases occur among older adults with a history of tobacco use or excessive alcohol consumption. Preventive interventions for oral cancer include counselling of patients to modify risk factors and screening to identify precancerous and early-stage lesions. This report presents evidence-based guidelines on the prevention of oral cancer and precancer among asymptomatic patients. METHODS: Literature searches of the 1966-1999 MEDLINE and CANCERLIT databases were completed using the major MeSH heading mouth neoplasms. References from articles and recommendations of organizations were also reviewed. The evidence-based methods of the Canadian Task Force on Preventive Health Care were used to assess evidence and to develop guidelines. Advice from experts and other recommendations were taken into consideration. RESULTS: In cohort and case control studies, smoking cessation decreased the risk of oral cancer and precancer. Randomized controlled trials (RCTs) indicate counselling by trained health care professionals is effective in promoting smoking cessation. Although counselling has been effective for the reduction of excessive alcohol consumption in RCTs, no studies have examined whether alcohol reduction reduces the risk of oral cancer or precancer. The usefulness of general population screening is limited by the low prevalence and incidence of the disease, the potential for false-positive diagnoses and the poor compliance with screening and referral. There is no evidence that screening of the general population or high-risk groups leads to a reduction in mortality or morbidity from oral cancer. INTERPRETATION: There is good evidence to specifically consider smoking cessation counselling in a periodic health examination (grade A recommendation). For population screening, there is fair evidence to specifically exclude screening for oral cancer (grade D recommendation). For opportunistic screening during periodic examinations, there is insufficient evidence to recommend inclusion or exclusion of screening for oral cancer (grade C recommendation). For patients at high risk, annual examination by physician or dentist should be considered. Risk factors include tobacco use and excessive consumption of alcohol. These recommendations are similar to those made by the Canadian Task Force on the Periodic Health Examination in 1994 and by the U.S. Preventive Services Task Force in 1996. PMID- 10859726 TI - Clinical management of avulsed permanent incisors using Emdogain: initial report of an investigation. AB - The enamel matrix derivative Emdogain was recently approved for clinical use in a number of countries, including Canada. It has been shown to stimulate regeneration of periodontal ligament following periodontal surgery in adults. This paper reviews pertinent clinical and laboratory studies of Emdogain and describes the protocol and methods used for a longitudinal outcome study of replantation of avulsed permanent incisors in children and adolescents. Application of these methods is described in an illustrative case report of Emdogain use. This paper is meant to inform clinicians and guide those who are instituting similar investigations. PMID- 10859727 TI - Acquired tufted angioma of the lower lip mucosa. AB - The acquired tufted angioma is a unique, dusky red, vascular proliferation previously reported in the skin, usually developing in childhood or in young adults, which exhibits a distinctive microscopic appearance. Clinically, the condition enlarges at a variable rate, becomes stable and may regress spontaneously. A small, long-standing, vascular lesion of the mucosa of the lower lip, exhibiting microscopic and immunohistochemical features resembling those of acquired tufted angioma, is reported. The differential diagnosis, including pyogenic granuloma, capillary hemangioma and hemangiopericytoma, is discussed. PMID- 10859728 TI - Influence of composite inlay/onlay thickness on hardening of dual-cured resin cements. AB - This investigation evaluated the effect of resin composite inlay/onlay thickness on the hardness of a group of eight dual-cure resin-based cements. Fourteen disc specimens measuring 6 mm in diameter and 2.5 mm thick were prepared from each of eight dual-cure cements: Adherence, Choice, Duolink, Enforce, Lute-It, Nexus, Resinomer and Variolink. Two specimens from each material were directly light cured while the remainder of the specimens were light-cured through resin composite spacers varying in thickness from 1 mm to 6 mm. Curing through the spacers always resulted in a decrease in the Knoop hardness number. For some cements, hardness values were reduced by 50% or more when the resin composite spacer thickness was 4 mm or greater even when measurements were made one week after dual-curing. Low hardness values indicate the presence of a weak chemical curing mechanism that may compromise cement quality in areas of the cavity not readily accessible to the curing light. PMID- 10859729 TI - An evaluation of sampling and laboratory procedures for determination of heterotrophic plate counts in dental unit waterlines. AB - BACKGROUND: The high numbers of heterotrophic microorganisms that have been cultured from dental unit waterlines (DUWs) have raised concern that this water may exceed suggested limits for heterotrophic plate counts (HPCs). The main purpose of this investigation was to examine HPC variability in DUWs and to examine in detail the effect of laboratory processing of water samples on HPC values. METHODS: Water samples were collected from dental offices either at the beginning of or during the clinic day and were transported to the laboratory, where they were analyzed. RESULTS: Measuring HPC levels within an office would involve testing all units, because significant differences were found between units connected to the same municipal water supply. Within a unit, the average microbial count from high-speed lines was approximately twice the average count from air/water lines. The laboratory processing of water samples significantly affected the numbers of heterotrophic microorganisms that were recovered. Incubation temperature, time and media, as well as neutralization of residual chlorine, all had significant effects on the HPC values. However, no significant differences in microbial counts were found between samples plated with the spread plate method on R2A agar and those plated with the pour plate method with Plate Count Agar. CONCLUSIONS: Dental organizations have suggested target limits in terms of numbers of heterotrophic microorganisms recovered in water from dental units, but standards for laboratory handling must be established as well. A protocol for sample collection and laboratory handling is proposed. PMID- 10859730 TI - Linezolid (Zyvox). PMID- 10859731 TI - Meloxicam (Mobic) for osteoarthritis. PMID- 10859732 TI - Androgel. PMID- 10859733 TI - Ciclopirox (Penlac) nail lacquer for onychomycosis. PMID- 10859734 TI - The Atkins diet. PMID- 10859735 TI - [Plasma immunoadsorption treatment of malignant multiple sclerosis with severe and prolonged relapses]. AB - INTRODUCTION: The treatment of prolonged, severe relapses in multiple sclerosis (MS) patients who respond poorly to high-dose intravenous corticosteroids is not yet established. Plasma immunoadsortion (IA) removes immunoglobulins (IgG), immune complexes, and complement from plasma. It may bear some advantages compared to plasmapheresis, a nonselective procedure that requires substitution of patient plasma by colloids solutions or plasma, which may carry a potential risk for viral infections. PATIENTS AND METHODS: Three relapsing-remitting MS patients with a malignant course received IA. All they were experiencing a prolonged relapse unresponsive to high-dose intravenous corticosteroids, causing a locked-in state in two of the patients and severe pseudobulbar impairment in the third one. Five to six IA consecutive sessions were administered along a 7-10 days course. RESULTS: IA was followed by a prompt and unequivocal clinical response in all three patients, which paralleled a decrease in IgG, fibrinogen, and C3 complement serum levels. IA administration was followed by immunosuppressor therapy, either with cyclophosphamide and intravenous ACTH (2 cases) or mitoxantrone (1 case). Improvement has been sustained along a mean follow-up of 7.6 years (range: 7-8.5 years), only one of the patients suffering two mild clinical relapses. CONCLUSION: We believe that IA may be useful, either as a coadyuvant or alternative treatment in severe relapses in MS patients that do not respond to high-dose intravenous corticosteroid therapy. PMID- 10859736 TI - [Lack of correlation between plasma homocysteine levels and cerebral microangiopathy in patients wtih transient ischemic attack]. AB - OBJECTIVE: The aim of this study is to evaluate cerebral MRI findings in patients with atherothrombotic transient ischemic attacks (TIA) and its correlation with plasma homocysteine (Hcy) levels. PATIENTS AND METHODS: A total of 62 consecutive patients with the diagnosis of TIA of atherothrombotic origin were studied. MRI examinations were performed in all patients for the evaluation of the presence of infarct and/or white matter hyperintensities (WMHI). Plasma Hcy levels were determined according to the method described by Smolin and Schneider modified. RESULTS: Plasma Hcy levels were significantly (p < 0.036) higher in patients with MRI-detected infarcts (9.69 +/- 2.06 mumol/l) compared with patients without infarcts (8.65 +/- 1.7 mumol/l. There was no correlation (p < 0.33) between plasma Hcy levels and the presence or absence of WMHI seen on MRI. CONCLUSIONS: In TIA patients, plasma Hcy levels were significantly higher in patients with cerebral infarcts, but did not correlate with the presence of WMHI. Our results suggest that mild hyperhomocysteinemia would be associated with large-medium vessel rather than with small vessel disease. PMID- 10859737 TI - [Single fiber electromyography in the diagnosis of myasthenia gravis]. AB - INTRODUCTION: The electrophysiological exam complements the clinical diagnosis in patients with suspected myasthenia gravis. The most common used electrophysiological test are the repetitive nerve stimulation test (RNST) and the single fiber electromyography (SFEMG); this last one is more sensitive than the first one. OBJECTIVE: To describe the usefulness of SFEMG in a group of myasthenic patients. PATIENTS AND METHODS: We studied 52 patients with a clinical diagnosis of myasthenia gravis (female: 35, male: 17; age range: 10-48 years). They were classified according to Osserman (type I, IIa, IIb, III and IV) and correlated to the electrophysiological findings. RNST was carried out in abductor digiti minimi, trapezius and orbicularis oculi; and SFEMG was carried out in the extensor digiturum communis and frontalis muscles. It was analyzed the sensitiveness of each technique detecting neuromuscular transmission abnormalities, and the correspondence between the results of both techniques. RESULTS: The most frequent clinical form observed was the type I (22 patients), followed by 18 as type IIa, 9 as type IIb, and 3 as type IV; no one as type III. RNST with positive decrement was observed in 48% of patients, but neuromuscular jitter calculated by means of SFEMG was abnormal in the 100% of them, being statistically significant the difference (chi 2 = 79.72; p = 0.000). The highest positivity index in the SFEMG was observed in the frontalis muscle. Patients from groups IV, IIb and I were the most electrophysiologically affected (t = 8.23211; p < 0.05). CONCLUSION: SFEMG is a very sensitive electrophysiological tool, detecting abnormalities in neuromuscular jitter in the 100% of myasthenic patients, with an excellent clinical correlation. PMID- 10859738 TI - [Dyslexia as a disfunction in successive processing]. AB - INTRODUCTION: We present a study on reading and writing difficulties after normal instruction during a year. OBJECTIVE: Verifying if these patients showed a specific pattern of PASS (Planning, Attention, Sequential and Simultaneous) cognitive processing; if so, it allows us a rapid diagnosis and a useful cognitive remediation according to the PASS theory of intelligence. PATIENTS AND METHODS: Thirty patients were selected from neuropediatric patients because of learning disability. They were selected according to their performance on several tests of phonological aware and a test of writing to discover errors in spelling. Patients with verbal language problems, as in dysphasia, and patients with learning difficulty not determined by reading or writing were ruled out. A control group of 300 scholars was used. The translated DN:CAS battery was administered to the study group and the control group for assessing the PASS cognitive processing. Statistical factorial analysis of the control group was performed as a validity confirmation to discriminate the four PASS cognitive processes. Cluster analysis of the study group was performed to discriminate its homogeneity. Differences between means were tested with the t-Student. RESULTS: The four PASS cognitive processes were identified in the control group. The study group scored less than minus 1 SD in successive processing, the rest of the processes being clearly higher than minus 1 SD, and the mean of study group was inferior to control group (p = 0.001). CONCLUSION: A kind of dyslexia may be defined by disfunction in PASS successive processing. PMID- 10859739 TI - [Severe myoclonic epilepsy in childhood. Epidemiologic analytical study]. AB - INTRODUCTION AND OBJECTIVE: Severe myoclonic epilepsy in infancy (SMEI) is a relatively rare disorder which etiology is unknown. The descriptive epidemiologic study tries to find the exposure agents or other circumstances present in affected children. Were have studied this agents and circumstances in 28 children with SMEI. PATIENTS AND METHODS: The study included 28 children, 17 girls and 11 boys, diagnosed of SMEI. Patients were selected according to the criteria established by Dravet et al in 1984. For each patient, the following features were studied: sex, parents age, prenatal antecedents, perinatal antecedents, new born stage, psychomotor development until the first convulsion, country/urban social class, family history of epilepsy and for febrile seizures, and cause of the first seizure. RESULTS: All parents are young adults, between 25 and 40 years at the birth of the child. The number of the fratry is between the first and the fourth. There is no pre and perinatal antecedents. The birth's weight is between 1,800 g and 4,600 g (X: 3,580 g). In 16 children (57.15%), the first seizure appeared after any dose DTP vaccination, frequently the second. Familial antecedents existed for epilepsy in nine cases and for febrile convulsions in six cases. CONCLUSIONS: The more interesting findings are the high percentage of cases that have the first convulsions after any dose of vaccination DTP and the high percentage of cases with family history of epilepsy and/or febrile seizures. It suggests that in pathogenesis of the SMEI there is a constitutional factor or predisposition genetically conditioned in relation to immunological disorders or to a multifocal cortical microdysgenesis, possibly triggered by toxic-allergic factors. PMID- 10859740 TI - [The contribution of an intraarterial electroencephalographic study in patients with deep epileptogenic foci]. AB - OBJECTIVES: To evaluate the diagnostic efficacy of an intra-arterial electroencephalographic recording and determine which patients obtain most benefit from this technique, to compare the results obtained using other recording techniques and to establish a standard for recording. PATIENTS AND METHODS: We made 64 intra-arterial recordings in 30 patients from one of three groups: persons with drug-resistant temporal epilepsy; patients with epileptic seizures of any type who required cerebral arteriography and patients whose illness required selective anteriography for any reason. We used a Seeker 10 guide-wire, the end of which acted as an electrode and a 2 minute recording was made. The position of the electrode varied depending on the site of the patients disorder. Activity was simultaneously measured with surface electrodes. Using the chi squared non-parametric test, we analysed the efficacy of the test. The paired t test was used to establish the concordance between observers. We compared the results obtained from the intra-arterial EEG with the simultaneous surface recording. RESULTS: We found three types of electroencephalographic patterns. The commonest was defined by the presence of high-voltage multi-spiked acute waves. The sensitivity of the test was 93.33%, the specificity was 80% and the overall value of the test was 86.66%. The chi squared test showed its reliability in the diagnosis of deep epileptogenic foci. There was high concordance between the observers in the study. No complications were seen in the patients in this study. CONCLUSION: The intra-arterial EEG recording is a semiinvasive test which may be useful in a selected group of patients and has high sensitivity and specificity. PMID- 10859741 TI - [A study made on a data base of 2,471 patients with Parkinson disease and disorders of movement in Health District 1 of the autonomous region of Madrid. Observed demographic changes over 8 year-period]. AB - INTRODUCTION: A specialised unit may influence favourably primary levels of health system by alerting physicians and patients on the existence of those pathologies most commonly seen. OBJECTIVES: To determine demographic changes and referral patterns observed in a hospital-based monographic outpatient clinic on Parkinson's disease and movement disorders along an eight-year period (1991 1998). PATIENTS AND METHODS: Database analysis of 2,471 patients attending a specialised unit covering an urban and rural population of 630,000 people in South-East Madrid city and county. RESULTS: We observed a slight, albeit non significant reduction in the incidence of secondary parkinsonisms (40% of all parkinsonisms observed). This occurred despite a drastic reduction in drug induced parkinsonisms, particularly those caused by cinnarizine/flunarizine and flupentixol. The reduction was balanced by a relative increase in neurodegenerative parkinsonisms, particularly Lewy body dementia. There was a sustained decrease in the time elapsed between onset of symptoms and identification of parkinsonisms as well as identification of its primary or symptomatic nature. The incidence of tardive dystonia (46.7%) of all symptomatic dystonias) remained unchanged. Essential tremor referrals markedly increased along the study period. CONCLUSIONS: Demographics features of patients attending a Parkinson's disease and movement disorder unit are submitted to changes over time. These probably reflect better awareness about these conditions by patients and physicians, ready access to hospital-based specialised units, and a greater demand for specialised care by the aged. PMID- 10859742 TI - [Cerebrovascular reactivity by means of the breath holding index (voluntary apnea): the reliability of early repetition]. AB - INTRODUCTION: The Breath Holding Index (BHI) is a non-invasive method for evaluation of cerebrovascular reactivity, the results of which correlate with those obtained by using intravenous acetazolomide and CO2 inhalation, so that it can be used as an alternative method to these in cooperative patients. However, the technique is not completely defined. Therefore Markus et al make a second measurement of the BHI 2-3 minutes later and take the arithmetical mean of the two readings as the final result. OBJECTIVES: To assess the intra-observer concordance between two BHI obtained with less than two minutes difference, and therefore whether the second index obtained may be used as an indicator of cerebrovascular reactivity. PATIENTS AND METHODS: Our study included 18 patients (7 men and 11 women; average age: 28.8 years; limits 17-50 years) with primary headache and no known cerebrovascular disease, diabetes mellitus or arterial hypertension. Using transcranial Doppler 28 measurements of the BHI (BHI1) were made in the middle cerebral arteries (right and/or left) in a similar way to that described by Markus. Once the average arterial velocity returned to basal levels the procedure was repeated in the first two minutes (BHI2) after the first index. Statistical analysis was done using the Student's t test and the kappa index between the two BHI after taking 0.8 as the cut-off point. RESULTS: The BHI2 (average +/- DT: 0.79 +/- 0.34) was significantly lower than that of BHI1 (average +/- DT: 1.04 +/- 0.44) (p < 0.005, t = 3.683). The kappa index between the two indices was very low: 0.058 +/- 0.158. CONCLUSION: The BHI2 is not a reliable index of cerebrovascular reactivity since it under-estimates it in relation to the previous index, even when the average arterial velocity has returned to basal levels. PMID- 10859743 TI - [Genitourinary changes in multiple sclerosis: the need for a urodynamic study]. AB - OBJECTIVE: In this study we have evaluated the urinary and sexual alterations of patients with multiple sclerosis (MS) by means of neurourological and neuroandrological studies. PATIENTS AND METHODS: In 41 patients with MS we took a clinical history and made a neurourological physical examination. We also did a full urodynamic study including measurement of flow and postmictional residue, cystomanometry and test of the detrusor pressure/mictional flow. Selective electromyography of the periurethral sphincter and a cystourethrographic study were also done. In patients complaining of erection dysfunction this was also studied by means of an erection test with intracavernosal vasoactive drugs and special neurophysiological techniques including the obtention of S2-S4 evoked potentials in the bulbo-cavernosal muscle and the somatosensorial potentials of the pudendal nerve. RESULTS: In patients with MS there is a predominance of mixed and irritative urinary symptoms rather than obstructive symptoms. The vesico urethral dysfunction of upper motor neurone type was the most frequent urodynamic diagnosis. There were no statistically significant differences between the urinary symptoms of the different urodynamic diagnoses. Ninety percent of the patients with erectile dysfunction had associated vesico-urethral dysfunction of upper motor neurone type and 100% of the patient with erectile dysfunction had dysfunction of the sphincter of the urinary bladder. CONCLUSIONS: In patients with MS there is no correlation between the clinical symptoms and urodynamic diagnosis. Therefore urodynamic diagnosis is essential not only for diagnosis but also for development of suitable treatment. The urodynamic diagnosis of dysfunction of the bladder sphincter is a risk factor in patients with MS. In our series 100% of the cases with erectile dysfunction also had dysfunction of the bladder sphincter. PMID- 10859744 TI - [Lhermitte's sign in three oncological patients]. AB - INTRODUCTION: Lhermitte's sign was first described by Pierre Marie and Chatelin in 1917. Lhermitte published his report in 1920 and reviewed this in 1924. This phenomenon is characterized by the occurrence of an electric shock-like sensation going along the spine in a cervico-caudal direction with flexion of the neck, and may also be felt in the upper and lower limbs. Clinical cases. Case 1. A 49 year old woman diagnosed as having breast cancer and being treated with cisplatin presented with Lhermitte's sign. On physical examination the osteotendinous reflexes were absent but the abdominal cutaneous reflexes were present. There was reduced sensitivity to vibration. Case 2. An 80 year-old man, previously operated on for adenocarcinoma of the colon, without further treatment, presented with progressive weakness of all four limbs and Lhermitte's syndrome. On examination there was obvious weakness of all four limbs, with the sensory level at C3. A cervical MR scan showed a metastasis in the vertebral body of C3 with epidural involvement and compression of the spinal cord. Case 3. A 54 year-old man was being treated by radiotherapy for cancer of the larynx. He presented at the onset of Lhermitte's sign but MR and physical examination were normal. CONCLUSION: Lhermitte's sign is non-specific, although in oncological patients a detailed clinical history and examination should be done seeking data regarding chemotherapy, radiotherapy and spinal compression. PMID- 10859745 TI - [Spinal cord infarctions as a complication of coronary angiography]. AB - INTRODUCTION: Spinal cord infarction is the commonest vascular disorder of the spinal cord, but its incidence is low and difficulty in diagnosis means that often it is not recognized. Although cases of spinal cord infarction have been reported as complicating angiographic procedures, it rarely occurs nowadays. CLINICAL CASE: We describe a case of spinal cord infarction following coronariography. The patient was a 61 year-old man with ischemic cardiopathy who was admitted to our hospital for coronariographic study. Immediately after the study had been done, the patient complained of acute, intense lumbar pain with paraesthesia and progressive weakness of his legs, developing paraparesia and sphincter disorders. Magnetic resonance of the spine was done and in the spinal cord an oval lesion was seen in the medullar cone which was compatible with infarction. Six months later this finding was unchanged. CONCLUSION: A spinal infarction as a complication of invasive vascular studies, such as angiographies, is exceptional nowadays, but should be remembered as a possible adverse effect. It may be suspected in cases of acute lumbar pain with motor and sensory defects of the legs and may be confirmed on magnetic resonance studies of the spinal cord. PMID- 10859746 TI - [Spinal cord compression due to the vertebral hydatid cyst]. AB - INTRODUCTION: Osteomuscular hydatidosis is an uncommon disease in which the spine is involved in 50% of the cases. Preoperative diagnosis is difficult in our setting because of its low incidence and the absence of specific radiological criteria. CLINICAL CASE: We present the case of a 72 year-old woman who consulted for slowly progressive paraparesia and was diagnosed, using CT and MR, as having a vertebral lesion of T3. The patient was operated on and many vesicular lesions of hydatidosis were removed surgically. CONCLUSION: We review the different forms of presentation of spinal hydatid cysts and the possible therapeutic options. PMID- 10859747 TI - [Functional anatomy of the cerebral cortex implicated in visual processing]. AB - INTRODUCTION AND OBJECTIVE: The functional anatomy of the cerebral cortex implicated in vision is reviewed with the purpose of illustrating the complexity of the cortical processing in visual perception. DEVELOPMENT: A brief analysis of the retinogeniculocortical pathway is presented and a summary of the main findings of the physiological and anatomical studies of Hubel and Wiesel in the adult brain is also illustrated. Special attention is paid in showing the visual pathways for object and face recognition as well as spatial cognition. CONCLUSIONS: Visual cortical areas in primates and man can be divided roughly into a ventral stream in relation with the temporal lobe responsible for the processing and storage of information about the identity of objects, their shape, color, and other salient physical features; and a dorsal stream in relation with the parietal lobe and concerned with the analysis of the locations of objects, and their movements in space. These two cortical pathways are not totally segregated and both are related to the prefrontal cortex as well as other subcortical structures such as the thalamus, the basal ganglia, the superior colliculus and the cerebellum through the pontine nuclei. PMID- 10859748 TI - [The current state of citicholine in cerebral ischemia]. AB - INTRODUCTION: Citicoline has a neuroprotector effect since it reduces the lesions of nerve membranes, by increase in the synthesis of phospholipids, and reduces the levels of free fatty acids. In this study we review the existing data on the efficacy of citicoline in the treatment of acute ischemic cerebrovascular disease and its sequelae, both in animals and in clinical trials involving patients. DEVELOPMENT: In various animal models citicoline reduces the volume of cerebral infarction and neurological sequelae and also potentiates the effects of other neuroprotector drugs. On analysis of the literature, we found six randomised clinical trials, double-blind and placebo-controlled in which the effect of citicoline was evaluated in patients who had acute ischemic cerebrovascular accidents. In all of these it was found that citicoline reduced the neurological sequelae. The finding of a broad therapeutic window (24-48 hours) gives this an advantage over the fibrinolytic agents, which have to be given within the first three to six hours. In further controlled trials carried out in patients with sequelae of cerebrovascular disease different degrees of neurological improvement were found, although studies of greater duration are required to see improvement in a longer term. CONCLUSIONS: With the data available, we may affirm that citicoline is a safe, effective drug, although clinical trials are still underway in larger populations of patients. In these trials not only the neurological state but also the area of infarction are assessed to confirm their efficacy. PMID- 10859749 TI - [Are statins indicated in the prevention of cerebral infarct?]. AB - INTRODUCTION: Some studies of ischemic cardiopathy have shown that when pravastatin is used for the prevention of strokes, these are reduced. Whilst we await suitable clinical trials, we discuss the possible role played by these drugs in this subgroup of patients. DEVELOPMENT: A panel of experts from different specialties assess the data published on dislipemias in the epidemiology of strokes, the possible effect of statins in the prevention of cerebral infarcts in patients with atheromatous stenosis of the carotid artery and their mode of action. CONCLUSIONS: Pravastatin is indicated in all patients with ictus of atheromatous origin as primary prevention of ischemic cardiopathy, in patients with strokes and hypercholesterolemia, and in patients with symptomatic or asymptomatic carotid stenosis while we wait for more specific clinical trials. PMID- 10859750 TI - [Thalamic infarct and pseudomedullary level]. PMID- 10859752 TI - [Is the fundus striati a new limbic-motor interphase?]. PMID- 10859751 TI - [Acute unilateral cerebellar ataxia: a case report]. PMID- 10859753 TI - [Torticollis secondary to intraparenchymatous hemorrhage]. PMID- 10859754 TI - [Functional evaluation of intraoral reconstructive surgery. A valuable tool: articulatory evaluation of the acoustic signal]. AB - Functional tests are needed to assess the quality of reconstructive surgery after treatment of intraoral cancers. Quality of Life tests are subjective and Cinefluoroscopy is a demanding and non-comparative procedure. We develop here a method to test the capacity of patients to maximize use of their articulatory space. We recorded a corpus of sounds. These sounds were analyzed with classical signal processing procedures. By comparison with a non-distorded sound database, it was possible to evaluate speech disorders, localize the defect, and provide a guide for rehabilitation. This method is an objective, reproductible, and comparative measurement tool. PMID- 10859755 TI - [Orthognathic surgery and stereolithographic models. A new technic of dental occlusion transfer]. AB - Use of stereolithographic models in orthognathic surgery is limited by the difficult in considering the facial osteotomies and the dental occlusion at the same time. Different techniques allow the surgeon to perform the simulation using composite prototypes after including the dental casts on the models. These techniques require complex "stereotaxic" systems or a surgical approach before CT scanning in order to insert the reference screws. They cannot overcome the problem of mandibular movement during the CT session. Our technique is a simple way to include the dental casts in the stereolithographic model with high precision. This can easily be done in a maxillo-facial environment and does not require any further special knowledge other than that which can be aquired in a classical dental laboratory. The occlusion transfer is achieved with a silicon cast of the teeth and the bony structures of the sterolithographic model on which we include the plaster dental casts. The silicone cast of the dental occlusion can also be used to decrease the mandibular movement during CT scanning. PMID- 10859756 TI - [On the article "Orthognathic surgery and stereolithographic models: a new technic of dental occlusion transfer"]. PMID- 10859757 TI - [The valuation of surgical procedures using PMSI: various remarks on the article by F. Boutault et al]. PMID- 10859758 TI - [Various observations concerning the article: "Circumscribed mandibular osteitis in children due to hematogenous dissemination. Apropos of 5 clinical cases"]. PMID- 10859759 TI - [Diagnostic difficulties of chondrosarcoma of the jaw. Apropos of a case and review of the literature]. AB - Chondrosarcoma is an uncommon malignant mesenchymal tumor characterized by the production of cartilaginous tissue and the absence of production of bone tissue. Maxillary and mandibular localizations are extremely rare and have a poor prognosis. The clinical and radiographic findings are similar to those seen in other tumors of the jaw, often delaying diagnosis and treatment and subsequently jeopardizing prognosis. In the literature, the mean delay from first clinical signs to diagnosis is about 8 months. As in all tumoral diseases, pathology confirms the diagnosis. We report a case of chondrosarcoma of the maxillary bone and review the literature, focusing on the difficulty in establishing the diagnosis, even at the pathology examination. We propose wide surgical excision, which, in agreement with other reports in the literature, is the only therapeutic possibility. We observed no sign of recurrence at 5-year follow-up. Radiotherapy and chemotherapy may be useful in certain cases. PMID- 10859760 TI - [Giant cell lesions of the maxilla disclosing primary hyperparathyroidism]. AB - We report 2 cases of giant-cell lesions of the maxilla in two female patients aged 42 and 22 years. The initial diagnosis was giant-cell tumor and giant-cell repair granuloma. The diagnosis of brown tumor was established at discovery of primary hyperparathyroidism. We emphasize the importance of searching for hyperparathyroidism in patients with a giant-cell lesion of the maxilla. PMID- 10859761 TI - [Bilateral carotid paraganglioma]. AB - We report an unusual case of bilateral paraganglioma of the carotid. The tumor had been neglected for a long time and increased in volume during pregnancy, inducing compression and requiring surgical treatment. We focus on the therapeutic strategy: devascularization followed by percutaneous sclerosis before total surgical removal in a two-step procedure. PMID- 10859762 TI - [Proteins of the inter-alpha trypsin inhibitor (ITI) family. A major role in the biology of the extracellular matrix]. AB - A glycoprotein with a high inhibitory activity against trypsin was isolated in 1961 from human plasma and named inter-alpha trypsin inhibitor (ITI). Since then, several other proteins that share antigenic and structural similarities with ITI have been identified and classified as members of the ITI protein family. These glycoproteins built up from different combinations of four polypeptides HC1, HC2, HC3 and bikunin are encoded by four genes H1, H2, H3, L on three chromosomes. Bikunin has two proteinase inhibitor domains and belongs to the Kunitz-type protease inhibitor family; it displays an inhibitory activity against trypsin, leukocyte elastase and plasmin. The heavy chains do not have any protease inhibitory properties but have the capacity to interact in vitro and in vivo with hyaluronic acid. This binding promotes the stability of the extra-cellular matrix. Consequently, the ITI protein family is suspected of playing a key role in the extra-cellular matrix biology. Isolation of free heavy chains in bronchial secretions and the recent emphasis about the bikunin role in tumoral invasion should enhance the interest about ITI protein family in the pathophysiology of chronic bronchopulmonary diseases or lung cancer progression. PMID- 10859763 TI - [The respiratory muscles in emphysema. The effects of thoracic distension]. AB - Besides increasing the work of ventilation, emphysema increases lung volume which in itself has a deleterious effect on the inspiratory muscles. We review here the effects of an acute change in lung volume on the configuration of the rib cage and muscle function. We also discuss the effects of the chronic distension associated with emphysema. The effects produced by changes in muscle length and configuration on the mechanical force and action of inspiratory muscles is detailed with particular focus on the diaphragm and its structural adaptations to experimental emphysema. We also analyze the activation pattern of inspiratory and expiratory muscles during the breathing process in patients with emphysema. Finally, we discuss the effects of single-lung transplantation and reduction surgery on chest distension and improved inspiratory muscle function. PMID- 10859764 TI - [Titration of an effective positive pressure level in the treatment of obstructive sleep apnea syndromes using continuous positive pressure ventilation]. AB - The efficacy of continuous airway positive pressure ventilation with a nasal mask mainly depends on the appropriateness of the effective positive pressure level. Conventionally, this level is determined from polysomnographic recordings using progressively increasing pressure levels to determine the point where episodes of apnea, hypopnea and snoring regress in all sleep phases and in all body positions. Since the first description of this method, many other titration methods have been proposed. Some use sophisticated signals such as the analysis of respiratory exertion level or limitation of inspiratory flow to provide a more precise titration which is particularly useful in cases where the classical titration method is insufficiently effective. Inversely, simplified methods have been examined from an economic point of view. These methods do not require the presence of a technician in a specialized laboratory where the waiting list for diagnostic tests is often long. Recordings during naps or short nights have been proposed for the more severe cases. Likewise titrations during simple polygraphy recordings or with an autoCPAP device have been shown to be effective in one night laboratory recordings. Much work remains to be done to determine the effectiveness of these methods when used in the patient's home as well the long term effects. PMID- 10859765 TI - [Results of 248 patients with sleep apnea syndrome treated by continuous positive pressure ventilation between 1990 and 1995. A study of compliance and outcome of the apnea-hypopnea index]. AB - Between 1990 and 1995, 369 patients were investigated for obstructive sleep apnea syndrome (OSAS) by polysomnography. Among them, 248 patients with a mean Apnea Hyponea index (AHI) of 37.7 per hour were treated by nasal continuous positive airway pressure (n-CPAP). Mean follow up was 39.5 +/- 20.4 months. In this group, 23 patients (9.2%) refused nCPAP immediately or after the first night and 39 (15.7%) gave up later. 15 patients (6%) died during the period of the study. The cumulative compliance reached 70% at 72 months. Non compliant patients usually gave up n-CPAP before the end of the first year. We compared the group of 150 patients always treated at the date of 31/12/95 with the group of 62 patients who refused nCPAP initially or gave up later. There was no difference in clinical parameters or polysomnographic data between the two groups. In 94 patients treated by nCPAP for more than a year we evaluated the outcome of AHI by a polysomnography performed after 72 hours of nCPAP cessation. Mean AHI of the group at this time was 38.2 +/- 20.3/h and was well correlated with the initial index (r = 0.41, p < 0.0001). However for 28 patients (29.7%) we observed, at the time of this second AHI determination, a variation (plus or minus) of at least 50% of the index. 6 patients, without any significative weigth loss, had an AHI below 5/h at this second determination. In this small group nCPAP was interrupted for 6 to 12 months, then another polysomnography was performed. At this time mean AHI was 42.4/h and clinical symptoms had reappeared in all patients. This study demonstrated that compliance to nCPAP in OSAS patients is good. No clinical or polysomnographic factors allow to predict non compliance. AHI is not modified by long term treatment with nCPAP. PMID- 10859766 TI - [Simultaneous resistance to rifampicin and isoniazid in patients with pulmonary tuberculosis]. AB - Percentages of primary and acquired resistance to anti-tuberculosis drugs provide an epidemiological indicator useful for assessing national anti-tuberculosis programs. Rifampicin and isoniazide are widely used in countries with a high prevalence of tuberculosis. In tropical Africa, these drugs are the mainstay treatment for tuberculosis, used both in the initial and long-term regimens. Simultaneous resistance to these two antibiotics would seriously jeopardize therapeutic efficacy. We studied simultaneous rifampicin and isoniazide resistance in patients hospitalized for tuberculosis in the respiratory disease unit of the Treichville University hospital in Abidjan, Ivory Coast. Mycobacterium tuberculosis was isolated in 8 patients. All the strains isolated were resistant to streptomycin. History taking revealed that resistance was observed at the initial prescription in 6 cases. A notion of contagion was present in 4 cases. Six patients were HIV-positive. Surveillance of resistance to anti-tuberculosis drugs is helpful in detecting early changes which could compromise the efficacy of the therapeutic scheme. PMID- 10859767 TI - [Lung disease due to Mycobacterium xenopi excluding AIDS. Apropos of 8 cases]. AB - Between 1994 and early 1999, Mycobacterium xenopi was isolated in 11 HIV-negative patients seen at the Respiratory Disease Department of the Dijon University Hospital. Eight of these patients met the criteria of lung infection. Clinical and radiological features simulated pulmonary tuberculosis which delayed diagnosis until the germ was identified. Treatment is considered to be mandatory though it is difficult to manage and often disappointing. In spite of long-term medical care, sometimes associated with surgery, outcome is currently determined by the underlying disease rather than by Mycobacterium xenopi infection. PMID- 10859768 TI - [Digestive disorders and Legionnaires' lung disease. Accompanying signs or visceral location?]. AB - Digestive disorders in Legionella pneumophila pneumonia such as nausea, vomiting, diarrhoea, are common; they are clinical arguments to suspect this bacteria to be responsible for this pneumonia. In this case-report, a patient with pneumonia due to Legionella pneumophila serogroup I presented in the follow-up with signs of enteritis with ascites. We looked ahead in literature who made us discover the multiple organ involvement that may happen in Legionnaires' disease. Diagnostic procedures consist in simple tests as ultrasonography, abdominal computerised tomography, that show inflammatory disease signs and sometimes ascites. Exceptionally, Legionella pneumophila has been demonstrated with direct immunofluorescent microscopic study, in inflammatory colitis pieces with haemorrhagic necrosis in different stage processes. Pathogenesis could be explained by the systemic spread of the organism and formation at distance of necrotising enteritis focus. It is initiated by necrotising factors of bacterial origin and hypersensitivity reactions (type I and III). PMID- 10859770 TI - [Primary biphasic synovial sarcoma of the pleura]. AB - A 36-year-old man presented with a pleural tumor. The first pathologic analysis diagnosed biphasic pleural malignant mesothelioma. However, the atypical clinical course, the early development of lung metastases and a new reading of histologic documents led to the diagnosis of primary pleural synovial sarcoma. The literature review is limited, as only nine other cases have been reported to date. Chest pain is the only constant clinical feature. Misleading interpretation of histologic material is frequent (6 of 10 cases). Only a complete immuno histochemical study confronted with the clinical course can lead to the correct diagnosis. Because the efficacy of chemotherapy and/or radiotherapy is poor, surgery remains the basis of treatment, whenever possible. Evolution is mainly intra-thoracic, with multiple local recurrences and lung metastases. Prognostic is poor, a survival rate is similar to that of primary pulmonary sarcomas. PMID- 10859769 TI - [Plasmocytic pleural effusion disclosing multiple myeloma]. AB - Pleural effusion caused by plasma cell involvement in multiple myeloma has been reported unfrequently, and has been described at a frequency below 1% of multiple myeloma. In this study, we report an observation with pleural effusion as first symptom of multiple myeloma. The analysis of the pleural liquid showed plasma cells with a monoclonal IgG Kappa immunoglobulin. In addition, there was a bone marrow infiltration by plasma cells, a serum monoclonal immuno-globulin of the same type and osteolytic lesions. Our patient has received one course of chemotherapy with: vincristine, melphalan, cyclophosphamide and prednisone. The patient did not respond to the therapy and died one month later. Pleural effusion seems to be an expression of aggressive myeloma. Survival exceeds rarely 4 months. PMID- 10859771 TI - [Traumatic pulmonary pseudocysts. Mechanisms of formation]. AB - Pulmonary pseudocysts (PPC) classically relate to chest trauma. It is a rare entity in adults, with multiple differential diagnosis. PPC most often evolve favorably. The clinical diagnosis is difficult to assess due to the poor and non specific clinical data. Chest radiographs are usually unsufficient for the diagnosis and the imaging modality of choice is computed tomography (CT). CT patterns of PPC relate to single or numerous cavities surrounded by air space consolidations. The physiopathological mechanisms of PPC remains uncertain. The histological study of this reported case affords some worth data to highlight the pathogenesis of this acquired abnormality. PMID- 10859772 TI - [Vocal cord dysfunction simulating severe corticoid-dependent asthma]. AB - We report a case of severe asthma initially considered as cortico-resistant. Clinical analysis of dyspneic attacks demonstrated they were atypical, sometimes associated with dysphonia and syncopes. Severity of clinical presentation was discordant with lung function tests. The diagnosis of vocal cord dysfunction was confirmed by ENT specialized examination. It showed paradoxal inspiratory adduction of the vocal cords triggered by exercise. Treatment remained however difficult, based on speech therapy, relaxation and psychotherapy. This observation underlines the influence of searching a vocal cord dysfunction in cortico-dependent asthma, especially if clinical presentation is atypical. Treatment of this condition may allow to decrease steroid treatment in such patients. PMID- 10859773 TI - [Opacity of the left upper mediastinum and tracheal deviation]. PMID- 10859775 TI - "Combining probability distributions from experts in risk analysis" PMID- 10859774 TI - [In response to the article by L. Portel]. PMID- 10859776 TI - Risk management across the globe: insights from a comparative look at Sweden, Japan, and the United States. AB - In light of the present day risk controversies such as global warming and hormones in beef, partially caused by a more globalized world, national differences and similarities in how to manage risks become increasingly important. In this brief "perspective" we focus on how risks are managed in three nations, namely Japan, Sweden, and the United States, specifically focusing on the roles of deliberation, risk analysis, and the importance of cultural factors. PMID- 10859777 TI - Chemical and radiation environmental risk management: differences, commonalities, and challenges. AB - Driven by differing statutory mandates and programmatic separation of regulatory responsibilities between federal, state, and tribal agencies, distinct chemical and radiation risk management strategies have evolved. In the field this separation poses real challenges since many of the major environmental risk management decisions we face today require the evaluation of both types of risks. Over the last decade, federal, state, and tribal agencies have continued to discuss their different approaches and explore areas where their activities could be harmonized. The current framework for managing public exposures to chemical carcinogens has been referred to as a "bottom up approach." Risk between 10(-4) and 10(-6) is established as an upper bound goal. In contrast, a "top down" approach that sets an upper bound dose limit and couples with site specific As Low As Reasonably Achievable Principle (ALARA), is in place to manage individual exposure to radiation. While radiation risk are typically managed on a cumulative basis, exposure to chemicals is generally managed on a chemical-by-chemical, medium-by-medium basis. There are also differences in the nature and size of sites where chemical and radiation contamination is found. Such differences result in divergent management concerns. In spite of these differences, there are several common and practical concerns among radiation and chemical risk managers. They include 1) the issue of cost for site redevelopment and long-term stewardship, 2) public acceptance and involvement, and 3) the need for flexible risk management framework to address the first two issues. This article attempts to synthesize key differences, opportunities for harmonization, and challenges ahead. PMID- 10859778 TI - Generic assessment endpoints are needed for ecological risk assessment. AB - This article presents arguments for the development of generic assessment endpoints for ecological risk assessment. Generic assessment endpoints would be ecological entities and attributes that are assumed to be worthy of protection in most contexts. The existence of generic assessment endpoints would neither create a requirement that they be used in every assessment nor preclude the use of other assessment endpoints. They would simply be a starting point in the process of identifying the assessment endpoints for a particular assessment. They are needed to meet legal mandates, to provide a floor for environmental degradation, to provide some consistency in environmental regulation, as exemplars for site- or project-specific assessment endpoints, to allow development of methods and models, to give risk managers the courage to act, for screening and site independent assessments, to support environmental monitoring, to facilitate communication, and to avoid paralysis by analysis. Generic assessment endpoints should include not only a list of entities and attributes, but also explanations of each endpoint, guidance on their use and interpretation, and measures and models that could be used to estimate them. PMID- 10859779 TI - Expert and layperson perceptions of ecosystem risk. AB - This research examines and compares perceptions held by laypeople and ecologists about risks to ecosystems, particularly risk from global climate change (GCC). A survey elicited perceptions of 31 risk characteristics for 13 GCC and 12 non-GCC risks to ecosystems. Factor analysis was used to examine the structure of layperson and expert risk perceptions. Both experts and laypeople tend to perceive GCC risks to ecosystems as less avoidable and more acceptable than risks from other causes. Compared to laypeople's perceptions, though, experts perceived GCC risks to have slightly lower impacts, be less avoidable, more acceptable, and less understandable than non-GCC risks to ecosystems. These findings may help guide efforts to communicate with laypeople about ecological risks from climate change. PMID- 10859780 TI - The influence of trust and perceptions of risks and benefits on the acceptance of gene technology. AB - A causal model explaining acceptance of gene technology was tested. It was hypothesized that trust in institutions using gene technology or using modified products has a positive impact on perceived benefit and a negative influence on perceived risk of this technology. Furthermore, perceived benefit and perceived risk determine acceptance of biotechnology. In other words, trust has an indirect influence on the acceptance of the technology. The postulated model was tested using structural equation modeling procedures and data from a random quota sample of 1001 Swiss citizens between 18 and 74 years old. Results indicated that the proposed model fits the data very well. The same causal model explains females' and males' acceptance of gene technology. Gender differences were found for the latent variables trust, perceived benefit, and acceptance of gene technology. Females indicated more trust, perceived less benefit, and demonstrated less acceptance than did males. No significant difference was observed for perceived risk. The implications of the results are discussed. PMID- 10859781 TI - Distributions of total job tenure for men and women in selected industries and occupations in the United States, February 1996. AB - This article develops and fits probability distributions for the variability in projected (total) job tenure for adult men and women in 31 industries and 22 occupations based on data reported by the U.S. Department of Labor's Bureau of Labor Statistics. It extends previously published results and updates those results from January 1987 to February 1996. The model provides probability distributions for the variability in projected (total) job tenures within the time range of the data, and it extrapolates the distributions beyond the time range of the data, i.e., beyond 25 years. PMID- 10859782 TI - Modeling ship transportation risk AB - This article presents results from the Commission of the European Communities (CEC) project "Safety of Shipping in Coastal Waters" (SAFECO). The project was performed by ten European partners during the period 1995-1998. The principal aim of the SAFECO project was to determine the influences that could increase the safety of shipping in coastal waters by analyzing the underlying factors contributing to the marine accident risk level. The work reported here focuses on the Marine Accident Risk Calculation System (MARCS) that was further developed during the SAFECO project. This paper presents the methods used by MARCS, as well as data and results from a "demonstration of concept" case study covering the North Sea area. The estimated accident frequencies (number of accidents per year) were compared with historical accident data, to demonstrate the validity of the modeling approach. Reasonable (within a factor of 5) to good (within a factor of 2) agreement between calculated accident frequencies and observed accident statistics was generally obtained. However, significant discrepancies were identified for some ship types and accident categories. The risk model has particular problems with estimating the accident frequency for drift grounding in general and powered grounding for ferries. It was concluded that these discrepancies are related to uncertainties in several areas, specifically in the risk model algorithms, the traffic data, the error and failure probability data, and the historical accident statistics. PMID- 10859783 TI - Percentiles of the product of uncertainty factors for establishing probabilistic reference doses. AB - Exposure guidelines for potentially toxic substances are often based on a reference dose (RfD) that is determined by dividing a no-observed-adverse-effect level (NOAEL), lowest-observed-adverse-effect-level (LOAEL), or benchmark dose (BD) corresponding to a low level of risk, by a product of uncertainty factors. The uncertainty factors for animal to human extrapolation, variable sensitivities among humans, extrapolation from measured subchronic effects to unknown results for chronic exposures, and extrapolation from a LOAEL to a NOAEL can be thought of as random variables that vary from chemical to chemical. Selected databases are examined that provide distributions across chemicals of inter- and intraspecies effects, ratios of LOAELs to NOAELs, and differences in acute and chronic effects, to illustrate the determination of percentiles for uncertainty factors. The distributions of uncertainty factors tend to be approximately lognormally distributed. The logarithm of the product of independent uncertainty factors is approximately distributed as the sum of normally distributed variables, making it possible to estimate percentiles for the product. Hence, the size of the products of uncertainty factors can be selected to provide adequate safety for a large percentage (e.g., approximately 95%) of RfDs. For the databases used to describe the distributions of uncertainty factors, using values of 10 appear to be reasonable and conservative. For the databases examined the following simple "Rule of 3s" is suggested that exceeds the estimated 95th percentile of the product of uncertainty factors: If only a single uncertainty factor is required use 33, for any two uncertainty factors use 3 x 33 approximately 100, for any three uncertainty factors use a combined factor of 3 x 100 = 300, and if all four uncertainty factors are needed use a total factor of 3 x 300 = 900. If near the 99th percentile is desired use another factor of 3. An additional factor may be needed for inadequate data or a modifying factor for other uncertainties (e.g., different routes of exposure) not covered above. PMID- 10859784 TI - Probabilistic analysis of the inadvertent reentry of the Cassini spacecraft's radioisotope thermoelectric generators. AB - As part of the launch approval process, the Interagency Nuclear Safety Review Panel provides an independent safety assessment of space missions--such as the Cassini mission--that carry a significant amount of nuclear materials. This survey article describes potential accident scenarios that might lead to release of fuel from an accidental reentry during an Earth swingby maneuver, the probabilities of such scenarios, and their consequences. To illustrate the nature of calculations used in this area, examples are presented of probabilistic models to obtain both the probability of scenario events and the resultant source terms of such scenarios. Because of large extrapolations from the current knowledge base, the analysis emphasizes treatment of uncertainties. PMID- 10859785 TI - Fitting second-order finite mixture models to data with many censored values using maximum likelihood estimation. AB - Finite mixture models, that is, weighted averages of parametric distributions, provide a powerful way to extend parametric families of distributions to fit data sets not adequately fit by a single parametric distribution. First-order finite mixture models have been widely used in the physical, chemical, biological, and social sciences for over 100 years. Using maximum likelihood estimation, we demonstrate how a first-order finite mixture model can represent the large variability in data collected by the U.S. Environmental Protection Agency for the concentration of Radon 222 in drinking water supplied from ground water, even when 28% of the data fall at or below the minimum reporting level. Extending the use of maximum likelihood, we also illustrate how a second-order finite mixture model can separate and represent both the variability and the uncertainty in the data set. PMID- 10859786 TI - Air toxics and health risks in California: the public health implications of outdoor concentrations. AB - Of the 188 hazardous air pollutants (HAPs) listed in the Clean Air Act, only a handful have information on human health effects, derived primarily from animal and occupational studies. Lack of consistent monitoring data on ambient air toxics makes it difficult to assess the extent of low-level, chronic, ambient exposures to HAPs that could affect human health, and limits attempts to prioritize and evaluate policy initiatives for emissions reduction. Modeled outdoor HAP concentration estimates from the U.S. Environmental Protection Agency's Cumulative Exposure Project were used to characterize the extent of the air toxics problem in California for the base year of 1990. These air toxics concentration estimates were used with chronic toxicity data to estimate cancer and noncancer hazards for individual HAPs and the risks posed by multiple pollutants. Although hazardous air pollutants are ubiquitous in the environment, potential cancer and noncancer health hazards posed by ambient exposures are geographically concentrated in three urbanized areas and in a few rural counties. This analysis estimated a median excess individual cancer risk of 2.7E-4 for all air toxics concentrations and 8600 excess lifetime cancer cases, 70% of which were attributable to four pollutants: polycyclic organic matter, 1,3 butadiene, formaldehyde, and benzene. For noncancer effects, the analysis estimated a total hazard index representing the combined effect of all HAPs considered. Each pollutant contributes to the index a ratio of estimated concentration to reference concentration. The median value of the index across census tracts was 17, due primarily to acrolein and chromium concentration estimates. On average, HAP concentrations and cancer and noncancer health risks originate mostly from area and mobile source emissions, although there are several locations in the state where point sources account for a large portion of estimated concentrations and health risks. Risk estimates from this study can provide guidance for prioritizing research, monitoring, and regulatory intervention activities to reduce potential hazards to the general population. Improved ambient monitoring efforts can help clarify uncertainties inherent in this analysis. PMID- 10859787 TI - [Stretch and relaxation training with isokinetic training apparatus]. PMID- 10859788 TI - [Dangers and risks of black market anabolic steroid abuse in sports --gas chromatography-mass spectrometry analyses]. AB - Anabolic steroids have become increasingly popular among athletes even at subcompetitive or recreational level instead of extensive doping tests, educational campaigns and lethal incidents. Nowadays, the fitness boom has also produced a population of steroid users at high school level and also under non sports practicing children. After opening the borders to East Europe an explosion of the black-market for anabolic steroids occurred. Beside the well-known side effects of anabolic steroids new problems and risks occurred due to fake drugs from the black market. This review ist subdivided into two parts: We provide a detailed review of the literature an anabolic steroids to the reader the information needed to make an informed decision an the relative risks and benefits of anabolic steroids. Secondly, we evaluated 40 "anabolic steroids" obtained from the black market using mass spectrometry or gas chromatography analysis to evaluate the real pharmacological compounds. As the results of this analysis, we found that 15 (37.5%) these drugs contained different or any pharmacological compounds as labeled. From the external packing, a differentiation between original and the fake drugs was impossible. Therefore, a large information and credibility gap concerning anabolic steroids particular those from the black market exists between the athletes and the medical and scientific communities. We believe that this gap can only be closed if both groups are be better informed about anabolic steroids. PMID- 10859789 TI - [Coordination training after anterior cruciate ligament surgery]. AB - Many studies have shown that ACL-surgery leads to a decrease of proprioception abilities. In spite of the resulting high importance of coordinative exercise, there is nothing like a phase specific concept for coordinative training after ACL-operations going along with rehabilitation. Following a model, including all coordinative abilities, and a current after-care concept of ACL-surgery, the presented method is based on different categories of coordinative demands. PMID- 10859790 TI - [Injury profile in modern competitive karate--analysis of 1999 WKC-Karate World Championship Games in Bochum]. AB - Competitions in Karate are either carried out as Shobu sanbon (with fist padding) or Shobu Ippon (without fist padding). Aim of this study was to gain current data on injuries in modern competitive Karate and to compare the two different competitive systems. During the WKC-Karate-World-Championships held from June 12 13 1999 at Bochum, 392 bouts were carried out. Every injury that was seen by the tournament doctor was registered. 142 competitors sustained 168 injuries: 141 mostly minor contusions of the head and throat, 12 facial lacerations, 3 knock outs (mild brain injury), 3 thoracal contusions, 1 midfoot fracture and 9 other blunt injuries. We saw more injuries in Shobu Sanbon (146/302 bouts) than in Shobu Ippon (23/90). Most of the injuries (152) were caused by punches. In Shobu Sanbon, kicking techniques led to 17 injuries only. The injury pattern shown here is comparable to earlier studies. Severe injuries in competitive Karate are rare. The higher number of injuries in Shobu sanbon may be due to the longer fighting time and higher scoring. Fist pads used in Shobu Sanbon might also lead to a loss of control. Therefore, prophylactic fist padding to avoid injuries in competitive Karate has to be seen critically. PMID- 10859791 TI - [Injuries sustained in dinghy-sailing by beginners: an analysis]. AB - In the sailing season of 1998 536 students have been assigned to a study investigating injuries that happened during beginners courses of dinghy-sailing. They were advised to document all injuries even minor ones and their circumstances while being on the boat. Injuries of the upper extremities were found in 39.5%, the head was affected in 32.4%, the lower extremities was injured in 26.5% and 1.6% had injuries of neck and/or trunk. Various types of injuries were observed: contusions in 55.1%, graze of skin in 17.2%, cuts and tears in 14.3%, bruises in 6.3%, tender spots, blisters and callus in 4.6%, lacerations in 1.7%, pulled muscles in 0.4% and fractures were seen in 0.4%. Typical situations in which injuries occurred were the collision with the main boom (31.1%), the rigging up and down (13%), capsizing (10.50%), docking and casting off (9.7%), the handling of the sheets (9.2%), the conditions in the harbour (8.8%) and slipping while onboard (6.8%). It becomes apparent that contusions and gazes of the head are caused by the standing rigging in contrast to skin-graze, lacerations, cuts and bruises which are rather caused by the running rigging. There is a higher risk of injuries for the helmsman than for the foredeckhand. However the incidence is evenly distributed among the sexes. Moreover there are significant differences in the accident rate concerning the direction and the strength of wind. Stronger and onshore winds make the sailor more accident-prone. PMID- 10859792 TI - [Paget-Schroetter syndrome caused by wrestling]. AB - Upper extremity thrombosis (Paget-Schroetter syndrome) hints already at physical strains with regard to the synonymous term "effort thrombosis". We report two cases of upper extremity thrombosis caused by wrestling. Wrestling as an example of martial arts activity leads to traumatic venous intima lesions with resulting local activation of coagulation as a main reason in pathogenesis of these thromboses. The possibility of a Paget-Schroetter syndrome should be considered in differential diagnosis if upper extremity complaints of unknown origin occur after wrestling matches. This kind of subclavian vein thrombosis should be reported to the accident insurance. PMID- 10859793 TI - [Management of simple midfacial fractures, particularly in professional soccer players]. AB - Uncomplicated midfacial fractures represent a frequent and typical injury of soccer-players in oral and maxillofacial surgery. The treatment of these fractures in professional players requires special treatment modalities, especially concerning quick rehabilitation. The examples of a nasal bone fracture, a malar bone fracture and a zygomatic arch fracture in 3 professional soccer players demonstrate the therapeutic principles of these most common uncomplicated midfacial fractures. In this context the fabrication of individual facial masks is presented, which allow an improved postoperative protection and therefore an earlier participation of the player in training and matches. PMID- 10859794 TI - Prognostic significance of advanced neuromonitoring after traumatic brain injury using neural networks. AB - While the therapeutic impact of tissue oxygenation (PtiO2) supplementing ICP monitoring is proven by several clinical studies, its prognostic value is not well studied. In the following study artificial neural networks (ANN) were used to analyze the accuracy of outcome prediction after traumatic brain injury (TBI) for different combinations of clinical data and parameters derived from neuromonitoring. The total group included 95 patients suffering from TBI. For all patients clinical data (age, GCS, pupillary response etc.) were recorded and outcome was classified using Glasgow outcome scale after 6 months. In a first step a subgroup of 60 patients was chosen to train a neural network to predict outcome based only on clinical data. In a second step the resting 35 patients all having continuous neuromonitoring with automatic data storage of ICP and PtiO2 were chosen. Different network models were composed using the former clinical model plus up to three additional input units for the following parameters: (a) relative number of ICP > 40 mmHg, (b) relative number of PtiO2 < 5 mmHg and (c) relative number of ICP > 30 mmHg with simultaneous PtiO2 < 10 mmHg. For each model the following time periods were analyzed: day 1-2, day 1-3 and day 1-4 after trauma and additionally day 1-4 after trauma plus last day of neuromonitoring. Pure clinical data allowed to predict outcome with 74.3% accuracy. A combination of clinical data with ICP (a) significantly increased the confidence levels of outcome prediction in all time periods (p < 0.05) with accuracy rates rising up to 82.9% for the longer time periods. The combination of clinical data and ICP & PtiO2 (c) lead to comparable results. In contrast, no significant increases were observed in the early time periods when combining clinical data with PtiO2 (b) while accuracy rates rose up to 80% for extended time periods after trauma. A combination of all parameters lead to results lying between the above results. The results indicate that prediction of outcome can be improved by combining clinical and neuromonitoring data. The prognostic value of ICP might be superior to that of PtiO2. PMID- 10859795 TI - [Value of transcranial doppler ultrasonography compared with scintigraphic techniques and EEG in brain death]. AB - Since 1991 transcranial doppler sonography has been accepted in Germany as a technical confirmatory test for the assessment of a cerebral circulatory arrest in patients fulfilling the clinical criteria of brain death. This study correlated transcranial doppler findings to established scintigraphic methods such as planar scintigraphy, 99mTc-HMPAO SPECT and EEG patterns. 21 patients (15 males/6 females, mean age 15-69 yrs.) fulfilled all clinical criteria of brain death. They suffered from head injuries and spontaneous bleedings. All clinical and technical investigations were performed within 60-90 minutes. In 14/21 patients clinical findings and all confirmatory tests were consistent with brain death. Planar scintigraphy and SPECT gave completely corresponding results in all 21 patients. 7 patients showed not corresponding results. In two head-injured patients with skull defects TCD yielded an oscillating flow in the MCA but SPECT/planar scintigraphy gave a residual perfusion in the related brain areas. A corresponding residual EEG pattern was seen in one case. A patient with osteoclastic skull defect showed a collateral flow from the external carotid artery and another case a secondary reperfusion in depth of a regular expected MCA signal 12 hours after definitely verification of systolic spikes in the Circle of Willis. No cerebral perfusion was detectable in the scintigraphique techniques. In the three remainder with rest activity in EEG, TCD and radionuclide methods showed no intracranial perfusion. In the presence of open skull fractures, external liquor drainages and osteoclastic craniotomies oscillating flow in TCD does not constantly represent a cerebral circulatory arrest. Awaiting of systolic spikes is absolutely necessary, if no radionuclide method is available. Determination of brain death by TCD should be carried out by an experienced investigator since unexpected collateral flow signals can be misinterpreted. PMID- 10859796 TI - [Endoscopic use of the water jet dissector in the cerebral ventricle system--an experimental study]. AB - Endoscopic surgery in the cerebral ventricle is limited by the small number of suitable instruments and bleedings obscuring the visibility. The water jet cutter allows the dissection of tissue, generally leaving vessels intact. Therefore it could be an additional instrument for neuroendoscopic purposes. In this preclinical study the employment of the water jet dissector under endoscopic conditions was evaluated. Incision-experiments (n = 80) on the ventricular system of fresh porcine brain were carried out under endoscopic view. To achieve similarity to the ventricular system, specimens were dissected in liquid medium (Ringer's-lactate, 37 degrees C). The depths of incision were measured digitally and histological examination of the tissue was performed. Depending on the applied pressure, distance from cutting nozzle to tissue and cutting velocity, the depths of incision ranged from 0.12 mm up to 2.4 mm. The water jet dissector is good to handle under endoscopic conditions and vessels are preserved. Based on these factors, this instrument is predestinated for further neuroendoscopic application. Prior to clinical application our results will have to be tested on living, perfused cerebral tissue. PMID- 10859797 TI - [Epidemiology of disk surgery in Germany]. AB - Actually, there are no valid epidemiologic data concerning intervertebral disk surgery available for Germany. Using resources of official statistical information we estimated the incidence of inpatient and outpatient disk surgery in Germany during last 6 years: The incidence for degenerative intervertebral disk surgery in Germany is about 87 operations per 100,000 inhabitants and year. There is no significant increase in the last five years, but there is a tendency to more outpatient disk surgery. Presently 16% of disk surgery is done in a outpatient procedure. Comparing with other European countries there are similar frequencies of disk surgery, contradictory in the United States there was a huge increase of the incidence for disk surgery since 1980. PMID- 10859799 TI - [Recommendations for early care of patients with skull and brain trauma in multiple injuries (prepared by the German Interdisciplinary Union for Intensive and Trauma Care (DIVI) on November 5, 1999]. PMID- 10859798 TI - Solitary plasmacytoma of the calvarium. AB - A solitary plasmacytoma of the calvarium was removed radically without postoperative radiotherapy. A 69-year-old female patient was operated for a giant solitary plasmacytoma in the frontal region and the follow-up over 10 years revealed no recurrence. The authors discuss that solitary plasmacytoma of the calvarium may have a good prognosis if radically removed and the radiotherapy may not be necessary. PMID- 10859800 TI - The management of postnatal depression. AB - At least one in ten women experiences depression in the weeks or months after the birth of a baby. While many recover spontaneously within a few months, one-third to one-half still have features of depression 6 months after delivery, and some go on to develop a chronic or recurrent mood disorder. Depression in the early postnatal months can have important effects on the mother and her baby, and on other family relationships. Here, we discuss the management of postnatal depression, concentrating on the treatment of women with non-psychotic illness. PMID- 10859801 TI - Early management of bleeding oesophageal varices. AB - Bleeding from oesophageal varices is a medical emergency with high mortality. In this article, we discuss its early management, focusing on the use of endoscopic therapy and drug treatment. PMID- 10859802 TI - An alternative style of clinic. PMID- 10859803 TI - How I treat ... a woman with a hatred of children. PMID- 10859804 TI - When does heavy flow merit treatment? PMID- 10859805 TI - Premenstrual syndrome is real and treatable. PMID- 10859806 TI - PID: a chance to make a difference. PMID- 10859807 TI - When a patient requests an abortion. PMID- 10859808 TI - Advances in women's health. PMID- 10859809 TI - A GP guide to managing vulval pain. PMID- 10859810 TI - Better patient compliance in psoriasis. PMID- 10859811 TI - Breaking the bad news. PMID- 10859812 TI - Assessing and treating the depressed child. PMID- 10859813 TI - Shoulder injections in primary care. PMID- 10859814 TI - Hoarse voice: an early symptom of many conditions. PMID- 10859815 TI - A campaign for success. PMID- 10859816 TI - [Treatment of bladder tumors]. PMID- 10859817 TI - [Neurology and incontinence in the year 1999]. PMID- 10859818 TI - [Surgical treatment of stress incontinence in women under local anesthesia]. PMID- 10859819 TI - [Future of surgical treatment of stress incontinence in women: classical surgery, laparoscopy, preperitoneal endoscopy or TVT? Our experience]. PMID- 10859820 TI - [Requirements for accreditation or retention of accreditation]. PMID- 10859821 TI - [Let's be sensible when we recommend an adequate calcium intake]. PMID- 10859822 TI - [Assessment of the appropriate utilization of an emergency department observation unit with the Appropriateness Evaluation Protocol: analysis of 4,700 cases]. AB - BACKGROUND: The Emergency Department Observation Units (EDO) constitutes an alternative of conventional hospitalization. The admissions and stays' appropriateness may influence the efficient use of bed resource in an acute care hospital. METHOD: Prospective evaluation of EDO admissions in a 18 month period, using the original criteria of Appropriateness Evaluation Protocol (A.E.P.), which were adapted to the area characteristics. RESULTS: We evaluate 4,700 admissions (55.1% male, 44.9% female; mean age: 64.9 +/- 14.9 years old). Average length of stay was 23.8 (+/- 18.3) hours. 35.5% patients were discharged at home and 62.0% were admitted in hospital. In 98.1% patients we obtained clinical stabilization in 48 hours. 85.0% of admissions were explained by 35 DRG categories, the most prevalent being chronic obstructive pulmonary disease (COPD) (411 admissions, 9.0%). A total of 258 (5.5%) admissions were qualified as inappropriate, primary due to diagnostic and therapeutical services that could have been provided in an outpatient basis (3.4%). A total of 797 (17.0%) stays were considered inappropriate, mainly due to hospital bed occupancy (13.0%) and discharge delays because of conservative medical management of patients (3.4%), social problems representing only 0.6%. A daily average of 4.2 beds (total of 19) were inappropriate used as assessed by modified AEP criteria. DISCUSSION: EDO represents an alternative for conventional hospitalization, obtaining patient clinical stabilization in 48 hours and saving unnecessary hospital admissions. AEP application lets know the impact of the efficient use of this area in the emergency department and the hospital. PMID- 10859823 TI - [Functional deterioration of basic daily living activities after an emergency service consultation]. AB - OBJECTIVE: To determine the incidence of functional decline of elderly patients discharged from an emergency department and to analized functional impairment as a risk of readmission. METHODS: A prospective cohort aged 75 or older were followed up after discharge from an emergency department between 01-02-95 and 01 04-95. The study protocol included sociodemografics, clinicals, functionals and mentalsoutcomes. We studied the incidence of functional decline in basic activities of daily living, with Barthel Index, and association with the risk of readmission. RESULTS: The sample was composed by 125 elders (mean aged 81.9 +/- 4.6 years and 60.8% were women). The incidence of functional decline in basic activities of daily living at the visit to emergency department was 20.8% and one moth after discharge was 18.4%. Both activities with more functional impairment were bathing, dressing and movility activities. Functional decline was associated with the risk of readmission at emergency department (Odds Ratio = 4.1 [1.4 11.8]) CONCLUSIONS: 20% of patients who are discharged of emergency department present a new functional impairment in basics activities of daily living. Functional decline is associated with the risk of readmission one moth after discharged. PMID- 10859824 TI - [Barometric pressure and respiratory quotient for estimating the alveolar arterial oxygen gradient]. AB - OBJECTIVE: To evaluate how they influence in the calculation of the alveolar arterial oxygen difference (AaPO2) the measurement of the barometric pressure (Pb) and respiratory quotient (R), in patients with chronic respiratory disease. METHOD: Prospective study of 39 patients with chronic respiratory disease in stable situation. We calculate: 1. AaPO2 (PAO2 - PaO2) through the equation of the alveolar air [PAO2 = PIO2 - PaCO2 [FiO2 + (1 - FiO2)/R)], assuming Pb = 760 mmHg and R = 0.8 and 2. Real AaPO2, with the figures measured of Pb and R. RESULTS: The values of AaPO2 and real AaPO2 were (mean +/- SD) 27.85 +/- 9.32 mmHg and 34.23 +/- 11.17 mmHg, respectively (p = 0.000). The figure of real AaPO2 is greater than AaPO2 in 34 patients, of those which in 21 cases (53.8%) is > or = 5 mm Hg and in 8 cases (20.5%) is > or = 10 mmHg. CONCLUSIONS: The traditional calculation of AaPO2 minimizes the severity of the pulmonary disease and it can contribute, in part, to the mistakes in the physiopathologic classification of the origin of the respiratory insufficiency (hipoxemia caused by alveolar hypoventilation and hipoxemia caused by alveolar hipoventilation complicated by other abnormalities of gas exchange). PMID- 10859825 TI - [Risk factors of non-ICU nosocomial pneumonia]. AB - OBJECTIVE: To know the incidence and risk's factors of nosocomial pneumonia (NN) in patient entered in hospital with age more of 14 years-old. The study was not performed in patients of Unit Care. Also, morbility and mortality are showed and the relation between bacterial agent and inmunologic answer of the patients. By other hand, we reported about specific clinical predictors for NN. MATERIALS AND METHODS: During a period of twenty-two months, 103 cases of NN were diagnosed what supposed 0.34% of the patients entered in this period of time. The study has been carried out in the University Arnau of Vilanova Hospital in Lleida (HUAV), with a capacity of 435 beds. Factors of the patients' risk of NN were valued. Multivariate analysis has been carried out by means of the pattern Cox's regression for proportional risks. RESULTS: About 66.99% of the patients had followed previous antibiotic treatment to the pneumonia episode, but only were significants those that had been treated with amoxicillin-clavulanic acid (28.15%, p = 0.0191). Blocking of the channels of the calcium were administered to 14.56% (p = 0.0030), and treatment with sedative drugs in 27.18% (p = 0.0397). Aerosol therapy was performed in 29 patients (28.15%, p = 0.0030). Nasogastric tube was performed in 19.41% of the patients (p = 0.0132). Creatinine was elevated in 31.42% (p < 0.05). The attributable mortality to the NN was of 33.9%, having a rate of higher mortality that the patients without NN. The agent etiologic was isolated in 23.3% of cases. The NN was polymicrobial in 8 cases with P. aeruginosa and S. aureus as the most frequents. CONCLUSION: In patients with several pathologies and treatments could be changed the immunologic answer and these patients are more predisposed to suffer NN. The mortality is associate to the age and others illness of the patient. PMID- 10859826 TI - [Diabetic ketoacidosis in patients with soft-tissue necrotizing infections]. AB - Necrotizing soft tissue infection is an uncommon and severe infection of the skin, subcutaneous tissue and superficial fascia which is usually accompanied by severe systemic toxicity. These infections occur more frequently in diabetics and are associated with higher morbidity and mortality rate. The prognosis of necrotizing fasciitis is known to be dependent on early recognition and treatment. Therefore, clinical awareness is important to avoid fatal outcome in patients with diabetes mellffus. We present three patients with undiagnosed type 2 diabetes in whom hyperglycaemia may have facilffated me development of necrotizing tissue infection, which in tum may have precipitated diabetic ketoacidosis in patients who rarely develop this metabolic complication. PMID- 10859827 TI - [Pentachlorophenol poisoning. Report of 4 cases and review of the literature]. AB - Pentachlorophenol (PCP) was, and still is, one of the most frequently used fungicides and pesticides. The use of PCP has been more restricted during the last few years. PCP's most important industrial application is as a wood preservative. The pentachlorophenol can be absorbed into the body by all the routes of occupational exposure. Some epidemiological observations suggest that exposure to PCP solutions may result in an increased risk for certain diseases in humans, e.g., immunodeficiency, blood disorders, malignancies, congenital anomalies. Chronic poisoning is difficult to detect since symptoms are often vague. Acute poisoning is due to interference with oxidative phosphorylation and can present itself as an unexpected case of sudden death. Four cases of PCP poisoning, including one fatalitie, occurred in two small wood preservative plants. All cases presented with increased serum creatinine phosphokinasa. The clinical finding are reported, and treatment modalities commented. At present there is no antidote for PCP. The basis for treatment is intensive supportive care to maintain vital bodily function. In one patient plasmapheresis was used and rapid recover was obtained. It is suggested that such therapy may be lifesaving in such intoxications. PMID- 10859828 TI - [Sudden death as presentation form of papillary fibroelastoma of mitral valve]. AB - We describe the case of a 35-year-old male, who presented with sudden death, secondary to anterior miocardial infarction due to embolism. Echocardiography, transthoracic and transesophageal, revealed a tumor in the anterior mitral leaflet. Tumor, histopathologically an papillary fibroelastoma, was excised and the mitral valve was replaced with a mechanical prosthetic valve. We revised the current literature on intracardiac papillary fibroelastoma. PMID- 10859829 TI - [Femoral osteolytic lesions with soft tissue tumors and hypercalcemia as presentation form of a B-cell lymphoma]. AB - Hypercalcemia associated with haematological neoplasms account for 15 to 20% of hipercalcemia in malignancy, and occurs usually in patients with multiple myeloma. However, its incidence in patients with linfoma is low, and it is observed usually in T-cell linfomas. Bone affectation is also uncommon in patients with non-Hodgkin linfoma. It usually is seen as a late manifestation of the disease, and its occurrence as the form of presentation is exceptional. We hereby report a patient with a B-cell non-Hodgkin linfoma presenting with hypercalcemia and femoral osteolytic lesions. PMID- 10859830 TI - [Bone mastocytosis. Diagnostic approach]. AB - The mastocytosis form a group of complaints characterized by the accumulation of mast cell. Systemic mastocytosis is diagnoses when there are one or more organ or tissues affected, independent of their localization in the skin. Bone mastocytosis affects between 70-90% of the patients suffering from systemic mastocytosis and lacks common diagnostic criteria. The most characteristic finds of bone mastocytosis, collected from 213 cases published in the universal literature over the last 20 years will be discussed. The results which, were found in relation to clinical, analytical, radiological and anatomophatological variables, aimed at the diagnosis of systemic mastocytosis and indispensable for the diagnosis of bone mastocytosis will be commented upon. Compiling this data the publications about mastocytosis between 1977 and 1997 have been revised, choosing those in which complaints of radiologic bone could be found. PMID- 10859831 TI - [Acute pancreatitis as presentation form of Crohn's disease]. PMID- 10859832 TI - [Acute brucellosis and human immunodeficiency virus infection. Current status]. PMID- 10859833 TI - [Syncope as first manifestation of primary amyloidosis]. PMID- 10859834 TI - [Carbon monoxide poisoning caused by a coal grill]. PMID- 10859835 TI - [Fistulas and ulcerative colitis: report of a case]. PMID- 10859836 TI - [Delayed assistance in acute myocardial infarction at a regional hospital]. PMID- 10859837 TI - [Transient cerebral ischemia and cerebral infarction in an HIV-positive patient with protein S deficiency]. PMID- 10859838 TI - [Severe bronchospasm associated with smoked cocaine]. PMID- 10859839 TI - [Multiple cranial neuropathy as presentation form of gastric tumor]. PMID- 10859840 TI - [Hemicranial headache caused by orbital cellulitis successfully treated with sumatriptan]. PMID- 10859842 TI - Use of abbreviations in records. PMID- 10859841 TI - Now you see it, now you don't... PMID- 10859843 TI - Sensationalist claims must be stopped. PMID- 10859844 TI - To tea or not to tea... PMID- 10859845 TI - Mouth protection in sport in Scotland--a review. AB - The oral health strategy for Scotland, which was published in 1995, recommends that dentists promote the use of mouth protection in sport to reduce the risk of injury. There is compulsory mouthguard use in some sports including ice-hockey, fencing, boxing, lacrosse and some forms of autocycling. In cricket, face protection appears to be compulsory for batsmen only. The use of mouth protection in the martial arts is compulsory at international level but, in the UK, the rule does not seem to be always enforced at club level. Players of contact sports, such as rugby and hockey, are considered to be more at risk of dentoalveolar injury and the governing bodies of these sports recommend that players at all levels wear mouth protection but have not made it mandatory. PMID- 10859846 TI - Dentine caries excavation: a review of current clinical techniques. AB - Since the invention and application of rotary instruments, the operative treatment of carious lesions has often resulted in considerable removal of tooth structure. More recently, newer techniques for removal of carious dentine have been developed in an attempt to minimise this excessive tissue loss. The following article reviews and discusses some of the techniques available to excavate demineralised dentine clinically. These methods can be classified as mechanical and non-mechanical, rotary and non-rotary and include: dental handpieces/burs, manual excavators, air-abrasion, air-polishing, ultrasonication, sono-abrasion, chemo-mechanical methods, lasers and enzymes. The advantages and disadvantages of each technique are discussed. PMID- 10859847 TI - Impression making. PMID- 10859848 TI - How much do the general public want to be involved in decisions on implementing water fluoridation? AB - OBJECTIVE: Test general public view about how much they wish to be involved in water fluoridation implementation decisions. DESIGN: Qualitative research using focus group discussions led by an experienced moderator. SETTING: Among the general public living in three non-fluoridated areas in England. SUBJECTS: Members of the public more than 20 years of age and by social class. RESULTS: General public wish to be informed of water fluoridation plans but do not see themselves as being the appropriate implementation arbiters. CONCLUSIONS: In the public's eyes, it would be inappropriate and unwelcome to delegate to them the decision about whether to fluoridate the public water supply. PMID- 10859849 TI - The emotional effects of tooth loss: a preliminary quantitative study. AB - AIM: To establish how widespread the emotional effects of tooth loss are. METHOD: A questionnaire, distributed to 100 edentulous people undergoing routine prosthetic care in the Department of Prosthetic Dentistry at Guy's, King's and St Thomas' Dental Institute (GKT), was used to explore the emotional effects of tooth loss. RESULTS: Ninety four people completed the questionnaire of whom 42 stated that they had experienced difficulties in accepting the loss of their teeth. In comparison with people who had no difficulties in accepting the situation, these people were: more likely to feel less confident about themselves; more likely to feel inhibited in carrying out everyday activities; and less able to accept the inevitable change in facial shape which occurs following the loss of teeth. Additionally, they took longer to come to terms with their tooth loss (All these differences were statistically significant). Just over three-quarters of the people who were unprepared for the loss of their teeth, felt that an explanation from the dentist prior to dental extractions would have helped. CONCLUSION: The impact that tooth loss can have on people and their lives should not be underestimated. In this study it affected 45% of the participants. PMID- 10859850 TI - Appropriateness of a Resuscitation Council (UK) advanced life support course for primary care dentists. AB - OBJECTIVES: To investigate whether a standard Resuscitation Council (UK) ALS course is appropriate for primary care dentists or whether a course should be specifically designed for dentists. DESIGN: Opinions canvassed by pre-course expectation and post-course evaluation questionnaires. SUBJECTS: 23 West Pennine primary care dentists providing a general anaesthetic or conscious sedation service who attended an ALS course. RESULTS: Knowledge and skills were rated on a 5-point scale from 1 (not important at all) to 5 (extremely important). Basic airway management (mean = 5) and anaphylaxis (mean = 4.9) scored the highest on the 'expectation' questionnaire. Rhythm recognition (P < 0.001), defibrillation (P = 0.007) and arrest algorithms (P = 0.047) were rated as significantly more important after the course than before. Knowledge about rhythm disorder management, cardiac pacing, post-resuscitation care, blood gas interpretation and bereavement were not considered to be so important either before or after the course. CONCLUSIONS: Despite rating some aspects as unimportant, all dentists stated that this course had been appropriate. They did not want a specially designed ALS course for dentistry. Taking exactly the same recognised course and assessments as other healthcare professionals and gaining the same certification was felt to be important to this group of dentists. PMID- 10859851 TI - Separate lives, different interests: male and female reproduction in the Gambia. AB - We report the initial findings of a research programme on the fertility and reproductive health of both men and women in rural Gambia. The reproductive experiences of men and women in the population studied were very different. During the period 1993-97, the total fertility rates were 12.0 for men and 6.8 for women. For men fertility began later, reached higher levels and continued into older ages than for women. Through serial and polygynous marriages, men were able to extend their reproduction beyond what would be possible with one woman. Of the married men interviewed, 40% were married polygynously. Men's fertility preferences indicated that they recognized their reproductive potentials to be greater than those of their individual wives. On average, married men desired 15.2 children for themselves and 7.3 for each wife. In this polygynous population the means available for attaining reproductive goals were different for the two sexes, depending on the separate lives and different interests of men and women. PMID- 10859852 TI - Making abortions safe: a matter of good public health policy and practice. AB - Globally, abortion mortality accounts for at least 13% of all maternal mortality. Unsafe abortion procedures, untrained abortion providers, restrictive abortion laws and high mortality and morbidity from abortion tend to occur together. Preventing mortality and morbidity from abortion in countries where these remain high is a matter of good public health policy and medical practice, and constitutes an important part of safe motherhood initiatives. This article examines the changes in policy and health service provision required to make abortions safe. It is based on a wide-ranging review of published and unpublished sources. In order to be effective, public health measures must take into account the reasons why women have abortions, the kind of abortion services required and at what stages of pregnancy, the types of abortion service providers needed, and training, cost and counselling issues. The transition from unsafe to safe abortions demands the following: changes at national policy level; abortion training for service providers and the provision of services at the appropriate primary level health service delivery points; and ensuring that women access these services instead of those of untrained providers. Public awareness that abortion services are available is a crucial element of this transition, particularly among adolescent and single women, who tend to have less access to reproductive health services generally. PMID- 10859853 TI - Severe maternal morbidity from direct obstetric causes in West Africa: incidence and case fatality rates. AB - Data on maternal morbidity make it possible to assess how many women are likely to need essential obstetric care, and permit the organization, monitoring and evaluation of safe motherhood programmes. In the present paper we propose operational definitions of severe maternal morbidity and report the frequency of such morbidity as revealed in a population-based survey of a cohort of 20,326 pregnant women in six West African countries. The methodology and questionnaires were the same in all areas. Each pregnant woman had four contacts with the obstetric survey team: at inclusion, between 32 and 36 weeks of amenorrhoea, during delivery and 60 days postpartum. Direct obstetric causes of severe morbidity were observed in 1215 women (6.17 cases per 100 live births). This ratio varied significantly between areas, from 3.01% in Bamako to 9.05% in Saint Louis. The main direct causes of severe maternal morbidity were: haemorrhage (3.05 per 100 live births); obstructed labour (2.05 per 100), 23 cases of which involved uterine rupture (0.12 per 100); hypertensive disorders of pregnancy (0.64 per 100), 38 cases of which involved eclampsia (0.19 per 100); and sepsis (0.09 per 100). Other direct obstetric causes accounted for 12.2% of cases. Case fatality rates were very high for sepsis (33.3%), uterine rupture (30.4%) and eclampsia (18.4%); those for haemorrhage varied from 1.9% for antepartum or peripartum haemorrhage to 3.7% for abruptio placentae. Thus at least 3-9% of pregnant women required essential obstetric care. The high case fatality rates of several complications reflected a poor quality of obstetric care. PMID- 10859854 TI - Maternal mortality in rural Gambia: levels, causes and contributing factors. AB - A demographic study carried out in a rural area of the Gambia between January 1993 and December 1998 recorded 74 deaths among women aged 15-49 years. Reported here is an estimation of maternal mortality among these 74 deaths based on a survey of reproductive age mortality, which identified 18 maternal deaths by verbal autopsy. Over the same period there were 4245 live births in the study area, giving a maternal mortality ratio of 424 per 100,000 live births. This maternal mortality estimate is substantially lower than estimates made in the 1980s, which ranged from 1005 to 2362 per 100,000 live births, in the same area. A total of 9 of the 18 deaths had a direct obstetric cause--haemorrhage (6 deaths), early pregnancy (2), and obstructed labour (1). Indirect causes of obstetric deaths were anaemia (4 deaths), hepatitis (1), and undetermined (4). Low standards of health care for obstetric referrals, failure to recognize the severity of the problem at the community level, delays in starting the decision making process to seek health care, lack of transport, and substandard primary health care were identified more than once as probable or possible contributing factors to these maternal deaths. PMID- 10859855 TI - Criteria for clinical audit of the quality of hospital-based obstetric care in developing countries. AB - Improving the quality of obstetric care is an urgent priority in developing countries, where maternal mortality remains high. The feasibility of criterion based clinical audit of the assessment and management of five major obstetric complications is being studied in Ghana and Jamaica. In order to establish case definitions and clinical audit criteria, a systematic review of the literature was followed by three expert panel meetings. A modified nominal group technique was used to develop consensus among experts on a final set of case definitions and criteria. Five main obstetric complications were selected and definitions were agreed. The literature review led to the identification of 67 criteria, and the panel meetings resulted in the modification and approval of 37 of these for the next stage of audit. Criterion-based audit, which has been devised and tested primarily in industrialized countries, can be adapted and applied where resources are poorer. The selection of audit criteria for such settings requires local expert opinion to be considered in addition to research evidence, so as to ensure that the criteria are realistic in relation to conditions in the field. Practical methods for achieving this are described in the present paper. PMID- 10859856 TI - Perinatal mortality attributable to complications of childbirth in Matlab, Bangladesh. AB - Very few population-based studies of perinatal mortality in developing countries have examined the role of intrapartum risk factors. In the present study, the proportion of perinatal deaths that are attributable to complications during childbirth in Matlab, Bangladesh, was assessed using community-based data from a home-based programme led by professional midwives between 1987 and 1993. Complications during labour and delivery--such as prolonged or obstructed labour, abnormal fetal position, and hypertensive diseases of pregnancy--increased the risk of perinatal mortality fivefold and accounted for 30% of perinatal deaths. Premature labour, which occurred in 20% of pregnancies, accounted for 27% of perinatal mortality. Better care by qualified staff during delivery and improved care of newborns should substantially reduce perinatal mortality in this study population. PMID- 10859857 TI - Integration of prevention and care of sexually transmitted infections with family planning services: what is the evidence for public health benefits? AB - It has been widely believed that, by combining the services for preventing and treating sexually transmitted infections (STI) with those for family planning (FP), STI coverage would increase and the combined service would be of higher quality and more responsive to the needs of women. So far, there is little concrete evidence that integration has had such an impact. Besides the absence of documentation, a clear definition of integration is lacking. We therefore carried out a comprehensive review of concrete experiences with integrated services, and present a summary of our findings in this article. The results indicate that the tasks of STI prevention, such as education for risk reduction and counselling, have been integrated into family planning services much more frequently than the tasks of STI diagnosis and treatment. Some STI/FP integration efforts appear to have been beneficial, for instance when the integration of STI/HIV prevention had a positive impact on client satisfaction, and on the acceptance of family planning. Less clear is whether STI prevention, when concentrated among traditional FP clients, is having a positive impact on STI risk behaviours or condom use. A few projects have reported increases in STI caseloads following integration. In some projects, FP providers were trained in STI case management, but few clients were subsequently treated. PMID- 10859858 TI - Measuring reproductive health: review of community-based approaches to assessing morbidity. AB - This article begins by reviewing selected past approaches to estimating the prevalence of a range of morbidities through the use of household or community based interview surveys in developed and developing countries. Subsequently, it reviews epidemiological studies that have used a range of methods to estimate the prevalence of reproductive morbidities. A detailed review of recent community or hospital based health interview validation studies that compare self-reported, clinical and laboratory measures is presented. Studies from Bangladesh, Bolivia, China, Egypt, India, Indonesia, Nigeria, Philippines and Turkey provide empirical evidence that self-reported morbidity and observed morbidity measure different phenomena and therefore different aspects of reproductive health and illness. Rather than estimating the prevalence of morbidity, interview-based surveys may provide useful information about the disability or burden associated with reproductive health and illness. PMID- 10859859 TI - Assessing the burden of sexual and reproductive ill-health: questions regarding the use of disability-adjusted life years. AB - The use of the disability-adjusted life year (DALY) as the unit in which to calculate the burden of disease associated with reproductive ill-health has given rise to considerable debate. Criticisms include the failure to address the problem of missing and inadequate epidemiological data, inability to deal adequately with co-morbidities, and lack of transparency in the process of ascribing disability weights to sexual and reproductive health conditions. Many of these criticisms could be addressed within the current DALY framework and a number of suggestions to do so are made. These suggestions include: (1) developing an international research strategy to determine the incidence and prevalence of reproductive ill-health and diseases, including the risk of long term complications; (2) undertaking a research strategy using case studies, population-based surveillance data and longitudinal studies to identify, evaluate and utilize more of the existing national data sources on sexual and reproductive health; (3) comprehensively mapping the natural history of sexual and reproductive health conditions--in males and in females--and their sequelae, whether physical or psychological; (4) developing valuation instruments that are adaptable for both chronic and acute health states, that reflect a range of severity for each health state and can be modified to reflect prognosis; (5) undertaking a full review of the DALY methodology to determine what changes may be made to reduce sources of methodological and gender bias. Despite the many criticisms of the DALY as a measurement unit, it represents a major conceptual advance since it permits the combination of life expectancy and levels of dysfunction into a single measure. Measuring reproductive ill-health by counting deaths alone is inadequate for a proper understanding of the dimensions of the problem because of the young age of many of the deaths associated with reproductive ill-health and the large component of years lived with disability from many of the associated conditions. PMID- 10859860 TI - Health sector reform and reproductive health in Latin America and the Caribbean: strengthening the links. AB - Many countries in Latin America and the Caribbean (LAC) are currently reforming their national health sectors and also implementing a comprehensive approach to reproductive health care. Three regional workshops to explore how health sector reform could improve reproductive health services have revealed the inherently complex, competing, and political nature of health sector reform and reproductive health. The objectives of reproductive health care can run parallel to those of health sector reform in that both are concerned with promoting equitable access to high quality care by means of integrated approaches to primary health care, and by the involvement of the public in setting health sector priorities. However, there is a serious risk that health reforms will be driven mainly by financial and/or political considerations and not by the need to improve the quality of health services as a basic human right. With only limited changes to the health systems in many Latin American and Caribbean countries and a handful of examples of positive progress resulting from reforms, the gap between rhetoric and practice remains wide. PMID- 10859861 TI - When medicine rediscovered its social roots. PMID- 10859862 TI - Preventive medicine in obstetrics. 1952. PMID- 10859863 TI - Insecticide resistance in Anopheles sacharovi Favre in southern Turkey. AB - We report the resistance to 12 insecticides of specimens of Anopheles sacharovi, both in laboratory cultures and those collected in the malarious areas of Adana, Adiyaman, Antalya, Aydin, and Mugla in southern Turkey. Mortality was higher 24 h after exposure than immediately after exposure but was unaffected by temperature (24 degrees C or 29 degrees C) or the position of the test kit (horizontal or vertical). In Adana, Adiyaman and Antalya, A. sacharovi was susceptible only to malathion and pirimiphos-methyl. In Aydin it was susceptible to both these insecticides as well as to dieldrin, lambda-cyhalothrin, and etofenprox; and in Mugla it was susceptible to dieldrin, fenitrothion, lambda-cyhalothrin, cyfluthrin and etofenprox, as well as to malathion and pirimiphos-methyl. PMID- 10859864 TI - Antibody response of patients after postexposure rabies vaccination with small intradermal doses of purified chick embryo cell vaccine or purified Vero cell rabies vaccine. AB - Although the introduction of tissue culture vaccines for rabies has dramatically improved the immunogenicity and safety of rabies vaccines, they are often prohibitively expensive for developing countries. To examine whether smaller doses of these vaccines could be used, we tested the safety and immunogenicity of purified chick embryo cell vaccine (PCECV) on 211 patients in Thailand with World Health Organization (WHO) category II and III exposures to rabies. The patients presented at two Thai hospitals and were randomized into three groups. Patients in Group 1 received 0.1 ml PCECV intradermally at two sites on days 0, 3, 7, and at one site on days 30 and 90. Group 2 was treated similarly, except that purified Vero cell rabies vaccine (PVRV) was used instead of PCECV. Group 3 received 1.0 ml PCECV intramuscularly on days 0, 3, 7, 14, 30 and 90. After 0, 3, 7, 14, 30 and 90 days serum was collected from the subjects and the geometric mean titres (GMTs) of rabies virus neutralizing antibody determined. After 14 days the GMT of 59 patients vaccinated intradermally with PCECV was equivalent to that of patients who received PVRV. Adverse reactions were more frequent in patients who received vaccines intradermally, indicating the reactions were associated with the route of injection, rather than the vaccine per se. We conclude that PCECV is a safe and highly immunogenic vaccine for postexposure rabies vaccination when administered intradermally in 0.1-ml doses using the two site method ("2,2,2,0,1,1") recommended by WHO. PMID- 10859865 TI - Global public-private partnerships: Part II--What are the health issues for global governance? AB - This is the second of a two-part review of global public-private partnerships (GPPPs) for health development. Part I was published in the April issue of the Bulletin (Vol. 78, No. 4). The recent emergence of GPPPs is rapidly reconfiguring the international health landscape. While most multilateral and bilateral agencies are currently grappling with how to proceed, there is little information in the public domain concerning how individual partnerships work and to date very little consideration of the many implications of this trend. This paper differentiates between product-based, product development-based and issues/systems-based GPPPs and describes a number of examples of each type in the health sector. The benefits of these initiatives, not least the major resources which they harness for specific health problems, are identified. The final section of the paper explores the implications and dilemmas posed by GPPPs. It discusses whether or not shared goals can transcend conflicting values and mandates and how governance of partnership arrangements may transform and undermine certain attributes of multilateral organizations. The paper concludes that the current climate of goodwill between public and private sectors offers an opportunity that should not be missed: it can be used not only to foster new partnership but to ensure that partnership is truly in the interests of international public health. PMID- 10859866 TI - Japan launches nursing insurance scheme for the elderly. PMID- 10859867 TI - Guidelines on health databases must consider developing countries. PMID- 10859868 TI - EU adopts legislation to promote drug development for rare diseases. PMID- 10859869 TI - Users' satisfaction with psychiatry services. PMID- 10859870 TI - Assessing quality of life in people living with psychosis. PMID- 10859871 TI - Why it is so difficult for persons with schizophrenia living in the community to achieve an adequate quality of life. PMID- 10859872 TI - [The impact on the international literature of the scientific production of Italian researchers in the disciplines "psychiatry" and "psychology". A bibliometric evaluation]. AB - OBJECTIVE: The aim of the study was to present the results of a citation analysis concerned with the impact of Italian researchers and institutions in psychiatry and psychology upon the international scientific community. METHOD: The analysis has been performed using a database of the Institute for Scientific Information (ISI): All scientific papers which were published between 1981 and 1998 in psychiatric and psychological journals included in the Science Citation Index (SCI) and the Social Sciences Citation Index (SSCI) were considered. The most cited Italian papers, authors and institutions are reported, as well the most frequently utilised journals. RESULTS: Publications concerned with neuropsychology, psychopharmacology and biological psychiatry were the most cited. This prevalence also affected the ranking of the most cited authors, even though, in this case, research groups in disciplines such as clinical psychology and epidemiological psychiatry appeared to be strong. The four most productive Italian Universities were characterized by the presence of both a School of Medicine and a School of Psychology. The Journal of Neurology, Neurosurgery and Psychiatry and Psychopharmacology were the most frequent vehicles of scientific communication. CONCLUSIONS: The results entail important implications for Italian research in psychology and psychiatry. On a general level, these analyses appear to be helpful for monitoring scientific production by granting agencies and for comparing different individual researchers. On a more specific level the analysis has identified the leading trends in research of Italian psychiatry and psychology. PMID- 10859873 TI - [Construction and validation of a new screnning questionnaires for eating disorders: the inventory for the screening of eating disorders/(ISED)]. AB - OBJECTIVE: The study aims to construct and validate a new screening questionnaire for the identification of cases at risk for eating disorders. SETTING AND SAMPLE: We assessed 218 female adolescent students recruited in a vocational school in Mestre (VE) and 88 patients consecutively referred to the Eating Disorders Unit of the University of Padova. PROCEDURE AND MAIN OUTCOME MEASURES: All subjects completed the new questionnaire, the Inventory for the Screening of Eating Disorders (ISED), and the Eating Attitudes Test (EAT). Sixty-two percent of female students and all the patients underwent a structured diagnostic interview (SCID for DSM-IV) for the diagnosis of eating disorders. The reliability, validity and screening ability of the questionnaire have been assessed. RESULTS: The reliability of the ISED, measured by Cronbach's alpha, is good (alpha = 0.87), as is the convergent validity (correlations with EAT). In order to improve the screening ability, the questionnaire has been further divided in two subscales: one for the identification of cases at risk for anorexia nervosa (ISED AN) and the other for cases at risk for bulimia nervosa (ISED-BN). The screening ability of the two subscales is greater than that of EAT, although the difference is not statistically significant. The ISED cutoff points have greater positive predictive values than those of EAT in the screening of both anorexia nervosa and bulimia nervosa. CONCLUSIONS: The ISED appears to be a valid and reliable questionnaire among female adolescents. Its use might improve the identification of cases at risk for eating disorders and in particular those at risk for anorexia nervosa. Other studies are needed to confirm the validity and performance of the questionnaire among samples of different ages and socio economic status. PMID- 10859874 TI - [Personality disorders prevalence study among inpatients with mood disorders, psychoactive use disorders and anxiety disorders]. AB - OBJECTIVE: The aim of this study was to asses type and prevalence of Personality Disorders (PDs) and their patterns of comorbidity with Axis I disorders in a sample of psychiatric inpatients. SETTING: The sample consisted of 300 subjects admitted to a psychiatric unit on a voluntary bases for an index episode. The study was conducted over a period of 12 months, from 1.11.1997 to 31.10.1998. MAIN OUTCOME MEASURES: The Italian version of SCID-II-PQ (Structured Clinical Interview for DSM-III-R personality disorders, with Personality Questionnaire--PQ -a self report questionnaire). RESULTS: More than half the patients had at least one personality disorder. The mean of disorders per patient was 2.83 +/- 1.93 (+/ SD). The most prevalent Axis II disorders were Borderline PD (30.7%), Obsessive compulsive PD (30.7%) and Avoidant PD (25.3%). Women were significantly more likely than men to meet criteria for Dependent PD and Avoidant PD. Man showed significantly more frequently than women Antisocial PD. Significant associations (p < 0.05) were found for comorbidity of Mood Disorders and Avoidant PD, and for Psicoactive Use Disorders and Antisocial PD. CONCLUSIONS: Our study confirms the high prevalence of PDs in psychiatric inpatients and showe some interesting associations between Axis I and Axis II disorders. These results can't be generalized to outpatients because our clinical sample involved mainly severely ill inpatients, but they raise questions about the exact nature of PDs and of the relationship with Axis I disorders. Further research involving outpatients and general population is needed to examine factors that could affect development and course of Personality Disorders. PMID- 10859875 TI - [Quality of life assessment: validation of the Italian version of the WHOQOL Brief]. AB - OBJECTIVE: To test the psychometric properties of the Italian version of the WHOQOL-BRIEF (e.g., construct and internal validity, concurrent validity with the MOS SF-36 and test-retest reliability). The WHOQOL-BRIEF is a 26-items self report instrument which assesses four domains assumed to represent the Quality Of Life (QOL) construct: physical domain, psychological domain, social relationships domain and environment domain, plus two facets for assessing overall QOL and general health. METHODS: Data have been collected in three sites (Bologna, Modena and Padua), located in the North of Italy, in the framework of the international WHOQOL project. According to the study design, the sample had to include about 50% males and 50% females, 50% of subjects below and 50% above the age of 45, all in contact with various health services. A subsample has been re-interviewed after 2-3 weeks in order to study test-retest reliability. After the WHOQOL BRIEF, most subjects have also been administered the MOS-SF36 in order to test the concurrent validity between these two instruments. RESULTS: The instrument was administered to 379 subjects (1/6 healthy and 1/6 sick), chosen to be representative of a variety of different medical conditions. Seventy patients, who displayed stable health conditions, have been reassessed after 2-3 weeks to study test-retest reliability. The WHOQOL-BRIEF domains has shown good internal consistency, ranging from 0.65 for the social relationships domain to 0.80 for the physical domain; it has been able to discriminate between in- and out patients and between the two age groups considered in the present study (< 45, > or = 45 years). Only physical and psychological domains were found to discriminate between healthy and ill subjects. No gender differences in the mean scores for the four domains were found. Concurrent validity between the WHOQOL Brief and the MOS-SF-36 was satisfactory, and specific for the physical and psychological health domains. Test-retest reliability values were also good, ranging from 0.76 for the environment domain to 0.93 for the psychological domain. CONCLUSIONS: This study shows that the WHOQOL-BRIEF is psychometrically valid and reliable, and that it is also potentially useful in discriminating between subjects with different health conditions in clinical settings. PMID- 10859876 TI - [Output standard in the mental health services of Reggio Emilia, Italy. Methodological issues]. AB - OBJECTIVE: The project Output Standards of the Mental Health Department (MHD) of Reggio Emilia is set out to define outputs and quality standards and to guarantee transparency and to facilitate organizational improvement. METHODS: The MHD started an interprofessional working group that defined the MHD outputs as long as process, quality peculiarities, indicators and standards for each output. The MHD Director validated the group results. RESULTS: The MHD defined 9 outputs and its indicators and standards and consequently modified its data registration system, the way to supply free and partially charged services and budget indicators. As a result, a new instrument for management and quality control has been provided. CONCLUSIONS: The A. maintains that to define outputs, indicators and standards will allow to compare several services of the Department, get them omogeneous and guarantee and improve quality. PMID- 10859877 TI - Inventory for the Screening of Eating Disorders (ISED). PMID- 10859878 TI - Structural changes of mitochondria during free radical-induced apoptosis. AB - The initial proposal for apoptosis stressed nuclear change (condensation of chromatin) and the intactness of intracellular organelles, including mitochondria, based on light and electron microscopic observations. However, data have accumulated to demonstrate that the opening of megachannels of mitochondrial membranes, resulting in the swelling of the organelles, notably by Ca2+ and free radicals, is the crucial step in the apoptotic processes of the cell. Application of fluorescent dyes to mitochondria, combined with flow cytometry, has made it possible to detect subtle changes in the structure and function of the organelles related to apoptosis. The present article overviews structural aspects of mitochondria related to apoptosis, including the free radical-induced formation of megamitochondria. PMID- 10859879 TI - The connections of the endopiriform nucleus with the insular claustrum in the rat and rabbit. AB - The connections between two parts of the claustrum in the rat and rabbit were studied using the highly fluorescent lipophilic carbocyanine dye (Dil). After the application of Dil crystal into the endopiriform nucleus, labeled fibers in the insular claustrum were observed in its part directly neighboring the insular cortex and capsula externa. Additionally, numerous projections into the piriform, insular and entorhinal cortices were present. The presence of connections between the endopiriform nucleus and insular claustrum suggests its role concerned with the processes taking part in the allocortical regions as well as in the limbic system. PMID- 10859880 TI - Types of neurons of the subthalamic nucleus and zona incerta in the guinea pig- Nissl and Golgi study. AB - The studies were carried out on the subthalamus of adult guinea pigs. Golgi impregnation, Nissl and Kluver-Barrera methods were used for the study. In Nissl stained sections the subthalamic neuronal population consists of multipolar, fusiform, oval and pear-shaped perikarya. In two studied areas: nucleus subthalamicus (STN) and zona incerta (ZI) three types of neurons were distinguished. Type I, multipolar neurons with quadrangular, triangular or oval perikarya. They have 3-6 primary dendrites which run slightly wavy and spread out in all directions. Type II, bipolar neurons with fusiform or semilunar perikarya, they have two primary dendrites. Type III, pear-shaped neurons with 1-2 dendritic trunks arising from one pole of the neuron. In all types of neurons axon emerges from the perikaryon or initial segment of a dendritic trunk and can be followed at a maximum distance of about 50 microns. PMID- 10859881 TI - The effects of short-time caffeine administration on skeleton development in Wistar rats. AB - The aim of the study was to evaluate the influence of caffeine on skeleton ossification in rats. Caffeine was administered in Tween 80 solution, once daily, in oral bolus, during the whole second trimester, in three doses: C1--0.7 mg/kg, C2--7.0 mg/kg, C3--70.0 mg/kg. On the 21st day of gestation the pumps were delivered. The fetuses were fixed in Bouin's solution and subsequently observed for external and internal malformation or in alcohol for skeleton malformation. The skeletons were stained with alizarin red-S. The examination showed an insignificant (P < 0.05) number of skeleton malformations, external haematomas and any internal malformations. PMID- 10859882 TI - Connection types between portal vein and portal sinus during foetal life. AB - We examined 27 resin casts of foetal liver veins and found three types of connection between portal vein and portal sinus. The most frequent connection was endo-lateral (66.7%), when the end of the portal vein joins with the lateral wall of the portal sinus. The next type was latero-lateral (14.8%). In this type, the lateral walls of the portal sinus and the portal vein join together. The last type was latero-lateral through short vessel (18.5%), which resembles the previous one, but there is short vessel between the lateral walls of the portal vein and sinus. PMID- 10859883 TI - Surgical anatomy of anterior petrosectomy. AB - Neurosurgical procedures in the region of the petroclival region of the skull base require unique knowledge of the local anatomy. The measurements of this region considering the visible anatomical landmarks are helpful both during surgery and while planning the general schemes for the approach. We have evaluated the anatomy of the anterior surface of the petrous bone and of the middle fossa taking into consideration the surgical removal of part of the petrous bone--the anterior petrosectomy. We have measured the distances and angles between the chosen structures in this region. The measurements were taken on 10 skulls, on both sides. The results enrich the algorithm of the anterior petrosectomy. PMID- 10859884 TI - The neuronal structure of the mamillary nuclei in guinea pig: Nissl, Kluver Barrera and Golgi studies. AB - The neurons of the mamillary body of adult guinea pigs were classified into four types: Type 1--unidendritic cells with rounded perikarya (7-16 microns) and one thick primary dendrite, mostly dividing into tortuous secondary branches; Type 2- bipolar cells: curly or simple ones with fusiform perikarya (13-22 microns); the curly-bipolar neurons possess 2 primary dendrites which may divide, even into tertiary dendrites, but each of them runs in screw-like or bending patterns; the simple-bipolar neurons have slender dendrites following a more straight route; Type 3--multipolar cells with cap-like perikarya (10-20 microns) and 2-3 dendritic trunks originating from the base of the perikaryon and running in a wavy pattern; sometimes their dendrites possess spiny-like protrusions; Type 4- multipolar cells with triangular or quadrangular perikarya (13-28 microns) and 3 4 dendritic trunks, poorly ramified, having a rather rectilinear course. In all types of neurons, dendritic spines are absent or rare. The majority of these neurons have a short impregnated axon originating from the perikaryon or primary dendrite. PMID- 10859885 TI - Qualitative and quantitative differences in the motor and somatosensory cortical projections of the rat claustrum--combined retrograde transport and stereological studies. AB - Using axonal retrograde tracing, combined with morphometric analysis, we compared the distribution and number of claustral neurons projecting to the motor and somatosensory cortical areas in the Wistar rat. Comparable volumes of the retrograde tracer Fluoro-Gold, were injected into the motor or somatosensory cortices. Injections into these areas resulted in labeling of neurons along the entire length of the claustrum. Neurons retrogradely labeled after injection into the motor cortex prevailed in the anterior part of the claustrum, whereas those projecting to the somatosensory cortex predominated in the central part. The mean number of claustral neurons retrogradely labeled after tracer injections into the motor cortex significantly outnumbered that from the somatosensory cortical area (p < 0.01). Similarly, the mean value of the numerical density of the retrogradely labeled neurons was significantly higher for the motor projection zone in the claustrum, than for the somatosensory projection zone (p < 0.001). The contralateral claustral projections, both into the motor and somatosensory cortices, were considerably lower in number than the ipsilateral ones. These findings indicate that: (1) the claustral projections to the various cortical regions seem to be differentiated (2) the distribution of claustral neurons projecting to the motor and somatosensory neocortical areas shows an anteroposterior gradient, (3) the claustrum of the rat appears to be more closely related to the motor than to the somatosensory system, (4) the rat claustrum seems to function more as a satellite than a relay structure in relationship to the cerebral cortex. PMID- 10859886 TI - Hippocampal capillaries in different age groups of chronically ethanol intoxicated rats. Morphometrical studies. AB - The influence of chronic ethanol intoxication on the terminal vascularization of particular hippocampal fields and layers was investigated in different age groups of rats. Thirty-six male Wistar rats aged 6 weeks were used in the study. For twelve months 24 of them drank only 25% ethanol--12 starting at 6-week-age and 12 at 3-month-age. The control group of 12 rats drank only water. As an effect of long-term ethanol exposure on hippocampal capillaries we observed the increase in the terminal vessel diameter and the decrease in microvascular length, surface, and volume densities. These changes varied between different age groups and between particular hippocampal regions. The observed age and regional differentiation of ethanol-related microvascular changes did not correlate well with the damaging effects of alcohol on corresponding neuronal elements, which emphasizes the very complicated pathogenesis of ethanol-induced injuries. PMID- 10859887 TI - Influence of diet and fluoride on dentin and enamel deposition and maturation in rats. AB - Diet and fluoride can modify tooth-germ development. In many morphological and biochemical studies malnutrition was shown to impair odontogenesis. However, quantitative observations of the morphological changes implemented by underfeeding and fluoride are still scanty. The aim of the study was to assess stereologically the enamel and dentin deposition in tooth-germ of 14-day-old rat pups derived from dams fed with deficient diet and given water without or with low (10 mg/l) and high (110 mg/l) doses of natrium fluoride, starting from the 13th day of pregnancy. The volume fractions of ameloblasts, enamel, dentin and odontoblasts in histological sections were estimated by the point counting method. The lack of fluoride in drinking water in rats maintained on low-protein diet changed the proportions of the deposited dental mineralised tissues as compared to the control animals: it substantially increased deposition of enamel (by 48%), and significantly decreased dentin production (by 28%). The supplementation of drinking water with fluoride in rats fed with deficient diet partially reversed these effects towards values found in the control rats maintained on standard diet that drank water with trace amount of fluoride. The possible toxic activity of high doses of fluoride can only be conferred to the decreased volume fraction of ameloblasts. Our findings suggests an important role of the fluoride ion in the maintenance of the proper enamel and dentin relation in the developing teeth of rats fed with low-protein, low-fat, high-carbohydrate diet. PMID- 10859888 TI - [Andrology is booming in 2000]. PMID- 10859889 TI - [Relationship between human immunodeficiency virus and sperm--implications in medically assisted procreation]. AB - The number of HIV-sero-discordant couples (man HIV+, woman HIV-) asking for assisted reproductive technologies (ART) has been increasing more and more since the efficiency of antiretroviral therapy was clinically proven. Long-term survey and amelioration of life quality in treated HIV-seropositive patients have induced in these couples a strong wish to conceive but they expected the most reduced risk of viral contamination. Epidemiologic data concerning HIV transmission during episodic unprotected sexual acts showed an elevated annual seroconversion rate which justifies that since 1992, European biologists specialized in human reproduction have proposed to carry out ART using intrauterine insemination (IUI) with prepared sperm in the population of couples where the man is HIV-seropositive. In spite of adapted technologies of sperm preparation, presence of HIV nucleic acids was demonstrated in purified spermatozoon (SPZ) fractions, resulting from residual free virus or virus linked to SPZ or residual infected cells, not completely eliminated. However, approximatively 2000 IUI were carried out with an HIV-controlled sperm treatment and no female and newborn seroconversions were reported. Even if the total lack of risk is impossible to obtain, a strict method of infected sperm preparation associated with sensitive virological techniques should permit us to obtain a minimal risk of contamination of women after IUI. In vitro fertilization (IVF) with or without microinjection allowed us to obtain the same results but they should be confirmed by further studies to be more relevant. These European workings, associated to a clear legal regulation in France, permit us to considerate that carrying out ART in HIV-sero-discordant couples in which the man is HIV-seropositive is allowable regarding both the viral problem and eventual sterility. PMID- 10859890 TI - [Are there indications for cryogenically preserving ovarian tissue in gynecologic oncology?]. AB - One of the essential goals of ovarian tissue cryopreservation is to preserve the fertility of young women who will be undergoing sterilizing anticancer treatments, as cryopreservation of sperm serves in men. However, maturation of gametes after thawing has not yet been realized in humans and pregnancies have only been obtained in animal models. It is nonetheless possible today to cryopreserve the ovarian tissue of patients who have nothing to lose as their follicular reserve would in any case be destroyed or severely depleted by cancer treatment. In gynecologic oncology, ovarian tissue cryopreservation may be performed when curative or prophylactic ovariectomy must be undergone, or when chemotherapy with high-dose alkylating agents is planned, particularly in cases requiring chemotherapy combined with pelvic radiation. These clinical situations may be encountered in young women and especially in girls with early-onset forms of ovarian, cervical, uterine, or vaginal cancers. These situations, nevertheless, remain rare as gynecological cancers occur most often in the post menopausal woman. PMID- 10859891 TI - [Trouble within compassion: psychological guidance during prenatal diagnosis, from indication to decision]. AB - This paper outlines the importance of psychological guidance throughout the process of prenatal diagnosis, according to the author's everyday experience in a multidisciplinary fetal medicine department. Consequences of the indication of a fetal disease are described for both sides (doctor and patient). The psychological mechanisms involved lead to the discomfort in the obstetrician's compassion and explain the intensity of the mother-to-be's reaction in the devastating situation. Being guided from the indication of a fetal malformation to her final decision aims to allow the mother to think through her first often impulsive and spontaneous decision and to consider different possibilities concerning the outcome of her pregnancy. A protocol is proposed. Guidance throughout the process of medical termination of pregnancy is developed. The psychiatrist's role should complement and follow through the psychological medical guidance. Its aim is to back up the technical and medical teams, to work on the psychoanalytic understanding of what the mother and the couple go through and have experienced, and also the consequences and the significance for the family. This is a preventive approach which deals with the feeling of guilt. PMID- 10859892 TI - [Endometrial thermocoagulation via balloon: technique, mechanism of action, and evaluation]. AB - Thermal balloon endometrial ablation is indicated for women suffering from menorrhagia. This easy technique can be performed safely, without using operative hysteroscopy. The rate of minor complications is low (< 4%). Thermotherapy treats the endometrium and superficial myometrium tissues without risk of injury to the bladder and bowels. The success rate of this operation is 90% in selected patients of studies published between 1996 and 1998, with a follow-up period of 12-24 months. However, long-term randomized controlled studies are needed to compare thermotherapy and other endometrial ablation procedures. PMID- 10859893 TI - [Anthropological studies of the representations of menopause in a population of women invited to a mass screening for breast cancer in Bouches-du-Rhone and Charente]. AB - This is an anthropological study of a target population (breast screening women) in the Bouches-du-Rhone and Charente regions of France. The occurrence of menopause is comparable in these two departments and depends on providing medical care which leads to breast screening. Menopause is natural for these women and is considered as a sign of ageing. Psychosomatic symptoms vary with sociocultural groups. Hormonal replacement therapy furthers breast screening. In the popular imagination, there is a deficit between nature (non-HRT) and culture (HRT). They take estrogen in the form of soy to offset this inadequacy, which creates a new cultural syncretism. PMID- 10859894 TI - [Improving oral contraception compliance. The "ringing card": memory aid or new ritual?]. AB - Forgetting to take pills is frequent and induces an avoidable risk of unwanted pregnancy. The integration of the daily use of the pill into a ritual allows to improve compliance. Nine hundred and seventy-five women were retrospectively asked by 180 gynecologists about missed pills in the last three and six months. More than nine out of ten women declare having forgotten at least one pill in the last six months. In 39% of the cases the pill was missed during the first week of 'cycle' in which the risk of pregnancy is theoretically increased. In this survey, 485 women used the compliance card for an average time of 3.5 months. The compliance card is a device that reminds the user to take the pill daily. It is the size of a credit card and can be programmed to ring daily at the same time 21 days out of 28. The efficacy of this device is attested by the great number of women who think that it allowed them avoid forgetting at least one pill in the last three months. Regardless of the age of the women, 91% of the users of the compliance card acknowledged that it allowed them to decrease the number of missed pills. Eighty-four percent think avoided forgetting at least one pill in the last three months, 34% between two and three pills and 17% more than three pills. In practice, 41% of the compliance card users didn't have any failure in taking the pill in the last three months versus 19% among nonusers (P = 0.001). Although women aware of their poor compliance more often think that they benefit from the compliance card, 83% of women who declare themselves as compliant share this opinion. The number of avoided missed pills by the compliance card is greater among women who often fail to take their pill. The mean number of missed pill during the three months preceding the use of the compliance card was 1.6 +/- 1.7 versus 0.9 +/- 1.3 during the three months of use. Among users of the compliance card, 98% think that it is easy to use and 97% like the way it works. The compliance card is an easy and reliable device that improves the compliance of women using oral contraceptives by helping them to establish a ritual. PMID- 10859895 TI - [Role of ultrasonography in diagnosing female infertility]. PMID- 10859896 TI - [Stimulation via GnRH antagonists for in vitro fertilization: something still to be defined]. PMID- 10859897 TI - [GnRH antagonists: yes or no?]. PMID- 10859898 TI - [Evaluation of lumbar canal stenosis: decubitus imaging methods versus flexion extension myelography and surface measurements versus the diameter of the dural sac]. AB - Though CT and MRI are presently the most frequently required noninvasive methods for the diagnosis of lumbar spinal stenosis (LSS), imaging of a supine patient may not demonstrate the maximal spinal stenosis shown by upright flexion extension myelography (FEM). Our prospective study tries to assess the averaging discrepancies between the supine CT-myelograms and the upright FEM in 50 patients. Considering all L2-L3 to L4-L5 vertebral levels, a mean underestimation of 16% of the diameter of the dural sac is found when and CT-myelograms are compared with extension myelography. Meaningful clinical discrepancies of 30% and more are found in 15.5% of these levels, the L5-S1 level remaining rather stable. Marked variations--but of less critical diagnostic interest--are also found between flexion and extension with, for flexion, a mean underestimation of 20% and discrepancies of 30% and more in 18.33% of L2-L3 to L4-L5 levels. Paradoxical results--CT diameters inferior to extension diameters--concern 22% of all studied levels but are of little clinical significance; only small discrepancies of 8.2% are found in a majority of non stenotic levels--with a maximal intensity for the L5-S1 level rarely implicated in LSS-. Measuring the mean cross-sectional surface occupied by the neural elements in the dural sac on CT-myelograms (189 evaluations), our study also empirically confirms a 60 to 80 mm2 are++ being the landmark of absolute stenosis. Finally, measurements of the cross-sectional area of the dural sac-109 L2-L5 levels inferior to 8.5 mm on CT myelograms or CT studies--show a large dispersion of areas for diameters superior to 6.5 mm and confirm cross-sectional area of the dural sac to be a much reliable parameter of LSS than diameter of the dural sac. We conclude that upright FEM--while not a first-line imaging modality for LSS--should be performed to exclude functional or dynamic position-dependent LSS in the patients whose symptoms are not explained by routine cross-sectional imaging, as long as no other upright technology is available. PMID- 10859899 TI - Metastatic breast cancer presenting with diabetes insipidus. AB - We present a case of acute-onset diabetes insipidus in a 60-year-old woman who had been treated for breast cancer. MR images showed a thickened and enhancing pituitary stalk and an asymmetrical hypophysis. The clinical diagnosis of a pituitary metastasis of the breast carcinoma was made. PMID- 10859900 TI - Giant intraluminal duodenal diverticulum: conventional barium study and computed tomography findings. AB - A case is reported of an asymptomatic intraluminal duodenal diverticulum (IDD) in a 21-year-old male patient with associated congenital abnormalities. During endoscopy for anemia an ostium in the duodenum was visualized, presumed to be the entry to an extraduodenal diverticulum. Upper gastrointestinal (UGI) barium examination showed, however, findings compatible with IDD. This diagnosis was supported by an abdominal computed tomographic (CT) examination. Surgical resection revealed a web in D2 with coexistent large IDD. PMID- 10859901 TI - Osler-Rendu-Weber disease with liver involvement. PMID- 10859902 TI - Breast imaging. An update. PMID- 10859903 TI - Recent advances in breast sonography. AB - Breast sonography has come to maturity during the past decade. Recent and ongoing technological developments include higher-frequency wideband transducers, 2-D array phased-array technology, extended-field-of-view imaging, three-dimensional sonography, higher-sensitivity color and power Doppler imaging, contrast agents, and harmonic imaging. Applications of breast sonography have thus expanded, with more reliable tissue characterization and differentiation between benign and malignant diseases and improved staging of cancer patients. Sonography has become the guidance technique of choice not only for percutaneous needle biopsy but also for many other interventional procedures involving nonpalpable breast masses. In selected cases, sonography can even be used to biopsy or localize calcifications that can be clearly identified on sonography. A growing field of research is ultrasound-guided percutaneous ablation of breast lesions using cryotherapy or radiofrequency. PMID- 10859904 TI - MR imaging of the breast--present indications. AB - MRI of the breast has proved its role in the diagnosis of breast cancer. Despite the amount of studies with MRI, the technique has not been standardised yet. This is one of the reasons for the huge difference in specificity the reported in the different studies (from 30% to 95%). Because the many pitfalls of the technique, the specificity of MRI of the breast is correlated to the experience of the investigator. When the established indications are respected, specificity increases. Presently accepted indications are: differentiation between postoperative fibrosis and tumor recurrence, multifocality or contralateral breast carcinoma, evaluation of prosthesis, discrepancy between the different diagnostic techniques, high risk patients with dense breasts. Less frequently but nevertheless a good indication is proven axillary lymph node metastases from an unknown primary tumor. Microcalcifications still are not a good indication for MRI of the breast. MRI is still inferior to mammography in detecting ductal in situ carcinoma or very small carcinomas because the neo-angiogenesis of these tumors is too faint to be detected by contrast-enhanced MRI. MRI instead of mammography or to solve a problem resulting from a bad mammographical examination is certainly not advised. New sequences, including diffusion and perfusion, and new contrast agents are keenly awaited. PMID- 10859905 TI - Digital detectors in mammography. A technological overview. AB - At a first glance the future of digital mammography seems very bright considering the wealth of offerings for new detectors, made by companies on the market or in journals. However technically spoken, digital mammography is one of the most demanding applications in the spectrum of radiology along with a high degree of cost consciousness in the mammography screening programs. The functional requirements are so high for digital mammography that only the best and most expensive components are good enough to compete with the current screen-film systems. Next difficulties are the lack of quality standards for digital detectors, not at all existing in the past and still under discussion, and the non-existence or changing of approval procedures. This is discouraging the industries to enter the field and is slowing down the phase-in time. After these obstacles are removed, there will come the discussions on acceptance of digital mammography within the professional community, due to the 'look' of the images and also the need to gain confidence with these images. PMID- 10859906 TI - Egg-shape cortical avulsion of the distal fibula: a frequent underestimated lesion. PMID- 10859907 TI - [Radical surgical treatment of hepatic echinococcosis]. AB - The results of surgical treatment of 42 patients with hepatic echinococcosis were analyzed. Solitary cyst was revealed in 28 (66.7%) of patients, complicated forms of the disease--in 5 (11.9%). Radical surgical intervention included the hepatic resection (in 18 observations) and pericystectomy (in 24). Specific postoperative complications didn't occur, all the patients survived. In the late follow up period the recurrences were not noted. PMID- 10859908 TI - [Surgical strategies in focal hepatic lesions]. AB - There were examined 124 patients with nidal affection of liver. In 52.4% of them the cyst was revealed, in 31.5%--the benign tumor, in 16.1%--malignancy. In the treatment of cyst of all sizes the puncture method with injection into her cavity of 96% ethanol solution was applied. Benign tumor was verified using the puncture biopsy. If the tumor less than 5 cm in diameter was present, the atypical hepatic resection was performed, if bigger one--hemihepatectomy. Mortality had constituted 3-8%. Surgical tactics is substantiated. PMID- 10859909 TI - [Surgical strategies in the treatment of complicated forms of chronic pancreatitis]. AB - For the period from 1992 to 1998 years 554 patients with complicated chronic pancreatitis were operated on. In 248 patients the longitudinal pancreatojejunostomy was performed, in 113--pancreatojejunostomy in 75--external drainage of the cyst, of them in 12--under the ultrasonic investigation control. In 21 patients pancreatoduodenal resection was conducted, pancreatic gland (PG) distal resection--in 41. In 4 patients with the PG head calcinosis the pancreatic resection according to Beger was done, in 3--the Frey operation. In the total PG calcinosis in 9 patients the modified Frey's operation was performed in conjunction with platitude PG resection along posterior wall of her duct. The indirect operation on PG were done in 40 patients. Nine (1.6%) of patients died. PMID- 10859910 TI - [Changes in free amino acid plasma levels in patients with acute pancreatitis and their correction with early parenteral feeding]. AB - The lowering of general pool of free amino acids in the blood plasma by 54.9%, comparing with their content in practically healthy persons, was observed in 17 patients with an acute pancreatitis. And by this the content of the majority of amino acids was lowered, excluding glutamate, phenylalanine and methionine, the level of which had raised. Under the conventional treatment influence the amino acids concentration had enhanced and constituted 60% from such in the control group, while in application of parenteral nutrition (intravenous infusion of the infezol 40 solution)--85.9%. By that the formation of the polyorganic insufficiency syndrome was not noted and the level of each amino acid had normalized. PMID- 10859911 TI - [Regional blood flow in stomach in duodenal ulcer disease complicated by under compensated or decompensated gastric exit stenosis]. AB - The state of the gastric mucosa regional blood flow and microcirculation in 148 patients with duodenal ulcer disease, complicated by sub-compensated or decompensated stenosis of gastric outlet, was studied. The lowering of indexes in various parts of stomach, along curvature minor especially, was revealed. These date substantiate the necessity of application of the organ-preserving operation with the ulcer excision and the dilating pyloroduodenoplasty performance in late stage of stenosis. If is recommended application of preparations, promoting normalization of the blood rheologic properties and mucosal microcirculation, for prophylaxis and the regional blood flow restoration in the early postoperative period. PMID- 10859912 TI - [The formation of the transplant for its descending to perineum with the sphincter-salvaging surgery in patients with disseminated sigmoid colon blood supply]. AB - The methods of the transplant formation in the loose type of the sigmoyd colon blood supply and insufficiency of the over vessel, applied in patients during sphincter-preserving operation performance for cancer recti, was proposed. Satisfactory immediate and late follow up result was noted. PMID- 10859913 TI - [Mediastinal drainage in patients with purulent mediastinitis]. AB - The results of treatment of 51 patients with mediastinitis using mediastinal drain were summarized. Postoperative mortality was 15.7%. PMID- 10859914 TI - [Choice of surgical strategies in aorto-femoral transplant infections]. AB - Reconstructive operation was performed in 4926 patients for occlusive affection of the abdominal aorta, of them in 132 (2.7%) the infectioning of aorto-femoral transplant was noted. Original methods of reoperation were presented, depending on the prevalence of the infection. Preventive performance of reoperation before the erosive hemorrhage occurred, had permitted to lower postoperative mortality from 52.3% to 13.5% and to achieve the lower extremities revascularization in 77.5% of patients. PMID- 10859915 TI - [Surgical strategies in obliterating disease of brachiocephalic trunk and subclavian arteries]. AB - Forty patients were operated on for obliterating disease of truncus brachiocephalicus and subclavian arteries. In 83.3% of patients the atherosclerosis was the cause of the arteries affection and in 16.7%--non specific aortoarteritis. Application of modern complex of the examination methods have permitted to accomplish differential choice of operative treatment and to achieve positive result in 87% of observations. PMID- 10859916 TI - [The experience of fraxiparin application in urgent surgery]. AB - Leading place among all postoperative complications are occupied by thrombosis and embolism as the cause of mortality in elderly and senile patients. Application of existing anticoagulants of direct (heparin) and indirect (fenilin, neodicumarin, sincumar) action is not always effective. If is necessary to introduce in clinical practice the medicinal preparations, which possess more effective and selective antithrombotic activity. To these requests corresponds lower molecular heparin fraxiparin (nadroparin calcium). For the thrombosis and embolism prophylaxis fraxiparin was used in 0.3 ml dose once a day subcutaneously during 3-5 days after operation. Investigation of the blood coagulation system before and after application of fraxiparin trusted the hemostasis normalization comparing with initial tenseness of the coagulation system. Comparative estimation of the normalization degree of the homeostasis system in patients with high risk of the thrombi formation under the influence of heparin and fraxiparin high efficacy of lower molecular heparin, fraxiparin, was established. So far as fraxiparin practically do not influence the homeostasis system negatively, it is necessary to consider him the preparation of choice in patients with an acute surgical illness and to use in the clinic of urgent surgery. The way of fraxiparin injection performance, the lack of necessity in frequent control of the blood coagulating and anticoagulation systems allow to apply the preparation in surgical clinics of various ranking and rigging. PMID- 10859917 TI - [Intensive therapy of patients with infected necrotic pancreatitis]. AB - In 70 patients the course of infected forms of necrotic pancreatitis was analyzed. Severity degree of the patients conditions was estimated according to APACHE II scale. Practical recommendations about management of such patients were elaborated. PMID- 10859918 TI - [The application of perilesional bioelectrical stimulation in mechanical treatment of wounds]. AB - Clinico-experimental substantiation of the around-wound electrostimulation in the apparatus method of the wounds treatment was performed. The electrostimulation influence, while in active electrodes situated in the around-wound tissues, was studied. The method efficiency was confirmed by the results obtained while treatment of patients with purulent process in epithelial coccygial trench. The proposed method application have promoted the reduce of postoperative complications occurrence as well as the patients stationary treatment duration. PMID- 10859919 TI - [Individualization and optimization of surgical treatment of the splenic traumatic injuries]. AB - The frequency of organ-preservation was raised up to 24%, the frequency of the purulent-inflammatory complications was lowered to 15.7% as well as mortality--to 11.7% during performance of operations on spleen due to application of the proposed individualized staged approach and new methods of hemostasis. PMID- 10859920 TI - [Precision surgery of tuberculoma in patients with diabetes mellitus]. AB - Indications and contraindications for precisional excision of tuberculoma in patients with diabetes mellitus were adduced, the original method of its performance was elaborated. Organ-preserving operation was done in 25 patients. The precisional excision of tuberculoma have permitted to preserve healthy pulmonary parenchyma (the part--in 18 patients and the whole lung--in 3). In all the patients the ability to work was restored. PMID- 10859921 TI - [Purulent complications in surgical treatment of severe forms of scoliosis]. AB - The results of treatment of 2719 patients, in whom the purulent complications have occurred in the region of metallic distractors due to the haematoma formation, were summarized. An active drainage of the distractor bed was applied for the purulent process and fistula formation prophylactics. It is necessary to model the gypseous corset in the region of osseous protuberances thoroughly for the bedsores prophylactics, especially if myelopathy due to scoliosis is present, and to use wadded paddings to reduce the pressure. PMID- 10859922 TI - [Prolonged course of intestinal agangliosis in children]. AB - The experience of diagnosis and surgical treatment of 24 children with typical subtotal form and 3--with total form of the Hirschsprung's disease was summarized. PMID- 10859923 TI - [Diagnosis and treatment of secondary tracheomalacia in intrathoracic compression of respiratory tracts caused by vascular ring anomaly in children]. AB - There were 48 children examined, aged from 1,5 mo to 14 yrs with the respiratory ways intrathoracic compression, caused by the vascular ring anomalies. In 21 (43.7%) patients the secondary tracheomalacia (ST) was revealed. In diagnosis of ST the data of morphological and histological investigation were used. Methods of surgical and conservative treatment of ST was elaborated. PMID- 10859924 TI - [Surgical treatment of benign tumors of thyroid gland in children and adolescents]. AB - In 1981-1998 years 552 children and juveniles were operated on for benign tumors of thyroid gland (TG). The enhancement of the revelation frequency of solitary and polynodous goiter and lowering of follicular adenoma and cystadenoma was noted. The tactics of children and juvenile patients with benign TG tumors was proposed, which included an early morphological verification of diagnosis, the puncture biopsy performance as well as intraoperative express-histological investigation. While solitary node in TG existence and preoperatively established its benign essence, extrafascial hemithyroidectomy, using original method, is the operative of choice. PMID- 10859925 TI - [The analysis of clinical management of polytrauma in the last 14 years]. PMID- 10859926 TI - [The role of cytokines in pathogenesis of acute pancreatitis]. PMID- 10859927 TI - [Hemorrhage of chromic duodenal ulcer in elderly and senile patients]. PMID- 10859928 TI - [The conjunction of biological tissues by electric welding]. PMID- 10859929 TI - [Rare observation of "forgotten" foreign body in the small intestine]. PMID- 10859930 TI - [Splenic ectopy extending to the right pleural cavity]. PMID- 10859931 TI - [Local treatment of purulent necrotic complications of acute pancreatitis]. PMID- 10859932 TI - [The use of unisept-3 for the local treatment of purulent wounds]. PMID- 10859933 TI - [Observation of successful treatment of listeria-related osteomyelitis in a newborn child]. PMID- 10859934 TI - [Science of evaluating the characteristics, quality and safety of biotechnological products: RDNA-derived products, cell culture technology-derived products, gene therapy products, cellular therapy products, and transgenic animal derived protein products and cellular products]. AB - Recent progress in biotechnology, including recombinant DNA (rDNA) technology and cell culture technology, has enabled us to produce new medically useful agents intended for human use. These agents include therapeutic peptides and proteins derived from rDNA-modified cell substrates, continuous cell lines, diploid cell lines, and hybridoma cell lines, gene therapy products, cellular therapy products, and therapeutic protein products and cellular products derived from transgenic animals. To enable these products to be of use in human therapy, it is essential that suitable measures be taken by manufacturers and regulatory authorities to assure their quality, efficacy, and safety. This article describes points based on the latest sound scientific principles to be considered when producing, testing, evaluating and controlling biotechnology products for human therapy, especially with respect to their characteristics, quality, and safety. PMID- 10859935 TI - [Geobotanical studies on the island of Tanegashima (affinity of world property island Yakushima)]. AB - A list was drawn up of wild plants growing on Tanegashiama island that were identified in our field work, and the list was compared with the flora of the rest of Japan and the flora of Taiwan. There were 166 families and 1,218 species consisting of 23 families and 159 species of Pteridophyta, 4 families and 7 species of Gymnosperma, 113 families and 700 species of the dicotyledous Angiosperma, and 26 families and 353 species of monocotyledous Angiosperma. There are 229 families and 5,500 species of plants in Japan, 196 families and 3,019 species in Kyushu, and 228 families and 3,477 species in Taiwan. There are 11 species of endemic plants on Tanegashima and Yakushima, and the best known of them is Pinus armandii Francht. var. amamiana Hatsushima. There are 181 species of flora of flora limited to the northern element, including several important medicinal plants, such as Akebia quinata Decaisne and Zanthoxylum piperitum DC. The 69 species of flora limited to the southern element include several important tropical plants, such as Messerschmidia argentea Johnston and Clerodendrum inerme Gaertn. Most of these plants are distributed on both island, but some of are distributed only Tanegashima. We concluded that one of the temperate borderlines of Japanese flora in the temperate zone is the islands of Tokara. The flora of Tanegashima and Yakushima are having a closely affinity of plant species and having the rich plant species. PMID- 10859936 TI - [Biological activity of photoexcited fullerene]. AB - Fullerene (C60, C70, etc.) is a third carbon allotrope discovered in 1985, and a great deal of attention has been focused on its physical and chemical properties in recent years. We are very interested in its biological properties for use fullerene as a pharmacophore. We first developed a method of solubilizing fullerene itself in water to perform in vitro biological screening. The concentrations of aqueous C60 and C70 solution with 5% poly(vinylpyrorridone) (PVP) are 400 and 200 micrograms/mL, respectively. By using aqueous fullerene solutions prepared in this manner, we have clarified a series of biological activities of fullerene, consisting of DNA-cleavage, hemolysis, cancer initiation, and cell-toxicity under photoirradiation, and chondrogenesis and inhibition of glutathione S-transferase activity without photoirradiation. The biological activity of photo-excited fullerene was found to be promising, because fullerene is a highly efficient photo-sensitizer. We synthesized a C60 derivative with an acridine moiety as a DNA-chelating function and assessed its effective DNA-cleaving activity. What kind of active species is involved in the biological action of photo-excited fullerene is our next concerns. Two pathways have been reported for the photo-excitation of fullerene. The so-called Type II energy transfer pathway generates singlet oxygen (1O2), while the Type I electron transfer pathway gives a fullerene radical anion (C60.-, C70.-). In order to clarify the effective oxygen species actually responsible for the biological action of photo-excited fullerene, we performed DNA-cleaving tests and EPR spectroscopic analyses under several conditions. The results showed that the photo-induced biological activity of fullerene is not caused by 1O2, but by reduced oxygen species (O2.-, .OH) generated by the electron transfer reaction of C60.-, with molecular oxygen. Its specificity is thought to be mainly attributed to the high-reducible property of fullerene. Since the reductive activation of molecular oxygen by photo-excited fullerene was observed at physiological concentrations of NADH as the reductant, fullerene can be classified as an oxyl radical-generating photosensitizer. Pharmaceutical application of fullerene to cancer photo-dynamic therapy appears promising. PMID- 10859937 TI - [Diagnosis of hepatic enzyme activities of drug metabolizing enzymes-phenotyping and genotyping]. AB - Drug metabolizing enzymes (DMEs) play an important role in the biotransformation of various xenobiotics, generally by detoxifying and eliminating substrates by converting them to more hydrophilic derivatives, or in some cases, activating substrates by conversion to intermediates that are highly reactive with biological macromolecules. It is widely accepted that the enzymatic activities of hepatic DMEs are one of the most important determinants of the concentration of drugs at their site of action or in the blood. Wide inter-individual variability in drug concentrations in the blood has been demonstrated even after the administration of the same doses on the basis of body weight, and in many cases the wide differences in plasma drug concentrations have been attributed to the individual differences in the enzymatic activities of DMEs. Attempts have therefore been made to estimate the hepatic DME activities of each individual before administration of drugs clinically. Three different approaches were taken to estimate patients' hepatic DME activities: 1) the use of probe drugs (phenotyping); 2) molecular diagnosis of genetic DME deficiency (genotyping); 3) examination of DME levels/activities in peripheral blood leukocytes/lymphocytes on the assumption that their activities in leukocytes/lymphocytes are well correlated with hepatic enzyme activities. A great number of data have been accumulated concerning the specificity of certain DME for candidate probe drugs, and searches have been made for mutated alleles of DME genes that indicate whether an individual is deficient in DMEs. Gene expression is measured by sensitive methods such as the reverse-transcriptase polymerase chain reaction and/or immunochemical methods in peripheral blood leukocytes/lymphocytes, which are less invasive tissues. Knowledge is being accumulated that will allow the development of useful methods of predicting the activities of hepatic DMEs that affect individual pharmacokinetics/pharmacodynamics. In this article, various reported methods of assessing hepatic DME activities are reviewed for the purpose of maximizing the efficacy and safety of pharmacotherapy. PMID- 10859938 TI - Studies on karyotype evolution in higher primates in relation to human chromosome 14 and 9 by comparative mapping of immunoglobulin C epsilon genes with fluorescence in situ hybridization. AB - Karyotypic homologies in relation to human chromosome 14 and 9 were studied through comparative mapping of the immunoglobulin C epsilon genes in higher primates by fluorescence in situ hybridization (FISH) technique. The C epsilon genes will be suitable probes for the analysis of evolutionary rearrangements due to that the multiple recombinational events such as gene duplications and deletions have occurred repeatedly in the immunoglobulin CH gene family (IGH@) during the course of primate evolution. IGH@ locating on the terminal region of human chromosome 14 (HSA14), at band HSA14q32.33, has generated multiple pseudogenes and among subclasses of IGH@ the C epsilon genes have shown most dynamic changes with generating both truncated type (C epsilon 2) and processed type (C epsilon 3) pseudogenes. In this study, chromosomal homologies and rearrangements on HSA14 (C epsilon 1) and HSA9 (C epsilon 3) in relation to the evolutionary genesis of their primate homologous chromosomes in speciation were investigated by comparative mapping with FISH and chromosome painting (ZOO-FISH) techniques. Comparative mapping of the C epsilon 1 gene at HSA14q32.33 was carried out in seven species of nonhuman primates: common chimpanzee (PTR), pygmy chimpanzee (PPA), gorilla (GGO), orangutan (PPY), white-handed gibbon (HLA), agile gibbon (HAG), and Japanese macaque (MFU). The C epsilon 1 gene was assigned to the telomeric region of HSA14 homologues in each species, namely, PTR15q32, PPA15q32, GGO18q16, PPY15q32, HLA17qter, HAG17qter, and MFU7q29, respectively. These results suggested that HSA14 has high degree of syntenic organization with its primate homologues confirmed by ZOO-FISH. Concerning HSA9, comparative mapping of the C epsilon 3 gene at HSA9p24.2-->p24.1 was performed. The mapped positions indicated the HSA9 homologous regions detected by ZOO-FISH in each species, namely, PTR11q34, PPA11q34, GGO13q22, PPY13q16, HLA8qter, HAG8qter, and MFU14q22, respectively, suggesting that several dynamic chromosomal rearrangements including at least twice pericentric inversions have occurred during the course of hominoid evolution. The comparison of syntenic groups and painting results has provided a hypothesis of the evolutionary genesis of HSA9 and its homologues with defined breakpoints on the present chromosomes. Likewise, studies on karyotype evolution will be promoted by combining comparative mapping with ZOO-FISH that can more clearly define the chromosomal rearrangements among species. PMID- 10859939 TI - [A 13-week subchronic oral toxicity study of haematococcus color in F344 rats]. AB - A 13-week oral repeated dose toxicity study of haematococcus color, a food additive mainly composed of astaxanthin, was conducted in male and female F344 rats. Rats were randomly divided into 4 groups each consisting of 10 males and 10 females and given CRF-1 powder diet containing 0%, 0.025%, 0.075%, and 0.25% haematococcus color, correspond to 0%, 0.5%, 1.5%, and 5% as the product. None of the animals died during the administration period. There were no exposure-related changes in body weight gain or food consumptions. Serum biochemical examinations showed dose-related increase in cholesterol, but the differences were slight and not defined as an adverse effect. No effects related to treatment were noted in hematological examinations and organ weights, and no abnormalities that could be ascribed to exposure to heamatococcus color were observed in histopathological examinations. In conclusion, ingestion of haematococcus color in the diet for 13 weeks does not cause any toxicological changes in F344 rats. PMID- 10859940 TI - [Teratogenicity study of sodium chlorite in rats by oral administration]. AB - The teratogenicity of sodium chlorite (NaClO2) was assessed in Wistar rats (Crj: Wistar). Sodium chlorite dissolved in distilled water was given to pregnant Wistar rats by gavage once a day from day 6 through 15 of pregnancy at doses of 0, 25, 50 and 100 mg/kg/day. The pregnant rats were sacrificed on day 20 of pregnancy, and their fetuses were examined for malformations. Sodium chlorite caused decreased food consumption, anemia, sedation, hematuria, and death in the pregnant rats at 100 mg/kg, but no fetal effects, such as malformations or growth retardation, were observed even at 100 mg/kg. It was concluded that sodium chlorite has no teratogenicity in rats when administered orally. The no-observed adverse-effect level was 50 mg/kg/day for pregnant rats and 100 mg/kg/day or more for rat fetuses. PMID- 10859941 TI - [A 13-week subchronic oral toxicity study of Perilla extracts in F344 rats]. AB - A 13-week subchronic oral toxicity study of Perilla extracts in drinking water containing 0%, 2.5%, 5% and 10% extracts was performed in both sexes of F344 rats. Rats were randomly divided into 4 groups each consisting of 10 males and 10 females. No animals died during the period of administration. There were no treatment-related changes in body weight gain or in hematological or blood biochemistry values. Nor were any treatment-related histopathological changes observed in the highest dose group. These findings indicate that ingestion of 10% Perilla extracts in drinking water for 13-week does not cause any toxicological changes in rats. PMID- 10859942 TI - [13-week subchronic oral toxicity study of ammonium sulfate in rats]. AB - A 13-week subchronic oral toxicity study of ammonium sulfate was performed in both sexes of F344 rats by feeding them a CRF-1 powder diet containing concentrations of 0%, 0.38%, 0.75%, 1.5%, and 3.0% of the substance. Rats were randomly divided into 5 groups each consisting of 10 males and 10 females. Male animals in the 3% group exhibited diarrhea during the administration period. No changes indicating obvious ammonium sulfate toxicity were observed in the body weights, organ weights, hematological, serum biochemical, or histopathological examinations. Based on these results, the NOEL (no-observed-effect level) of ammonium sulfate for F344 rats was judged to be 1.5% in males (886 mg/kg/day) and 3% in females (1975 mg/kg/day), and the MTD (maximally tolerated dose) for 2-year carcinogenicity studies in F344 rats was concluded to be 3.0% or more in the diet. PMID- 10859943 TI - [A 13-week subchronic toxicity study of D-xylose in F344 rats]. AB - A 13-week subchronic toxicity study of D-xylose was performed in male and female F344 rats at dose levels of 0%, 0.2%, 0.6%, 1.7%, and 5% D-xylose in the CRF-1 powder diet to determine the maximum tolerable dose (MTD) for subsequent investigation of carcinogenicity. Rats were randomly allocated to 5 groups each consisting of 10 males and 10 females. Rats were randomly allocated to 5 groups each consisting of 10 males and 10 females. No treated groups showed changes in body weight gain or food intake, and all animals survived until the end of the experiment. Hematological examination revealed significant increases in RBC, Hb, and Ht in the male groups treated with 0.6% and 5% concentrations, whereas these values decreased significantly in all of the female groups treated with D-xylose. However, no clear dose-response effect was observed in the hematological data in either males or females given D-xylose. Serum biochemistry studies revealed decreases in AsT in the 0.2% and 5% D-xylose group male and 0.2%, 1.7%, and 5% group female, compared to the control value. However, the changes were not considered specific because of the lack of any clear dose-response effect. In addition, no histopathological changes indicating obvious toxicity of D-xylose were observed in the livers of either sex treated with D-xylose. Based on these data, the MTD of D-xylose in F344 rats of both sexes is judged to be 5% or more in the diet. PMID- 10859944 TI - [A 90-day subchronic oral toxicity study of Bacillus subtilis gum in F344 rats]. AB - A 90-day subchronic toxicity study of Bacillus subtilis gum was performed in both sexes of F344 rats by feeding of CRF-1 pellet diet containing 0%, 0.18%, 0.55%, 1.66% and 5%. Rats were randomly allocated to 5 groups, each consisting of 10 males and females. No animals died during the administration period and no differences in body weights and food intakes were found among groups of either sex. Kidney weight was significantly increased in both sexes in the groups given concentrations of 1.66% or more. However, the increases of kidney weight were slight in themselves and other data on serum biochemistry and histopathology did not show any apparent toxicological signs including renal toxicity. These findings indicate that the treatment of Bacillus subtilis gum in the diet for 90 days does not exert serious toxicity in rats even at the highest dose. PMID- 10859945 TI - [A 13-week subchronic oral toxicity study of orange color in F344 rats]. AB - A 13-week subchronic toxicity study of orange color was performed in both sexes of F344 rats by feeding them a CRF-1 powder diet containing 0%, 0.18%, 0.55%, 1.66%, and 5% concentrations of the substance. No animals died during the administration period, and no changes in body weight or food intake were found in any of the dosage groups. There were significant increases in serum cholesterol in males given 1.66% or higher concentrations of orange color and in females given 0.55% or higher concentrations, and significant increases in alkaline phosphatase in males given 1.66% or higher concentrations, possibly due to the high-fat composition of the orange color diets. In addition, some hematological, serum biochemical, and histopathological changes were observed in the groups given greater than 0.55% concentrations, but they did not suggest obvious toxicity. These findings indicate that under these experimental conditions the no observed-effect level (NOEL) of orange color in the diet for 13 weeks is 0.18% and the no-observed-adverse-effect level (NOAEL) is 5%. PMID- 10859946 TI - [A 13-week subchronic toxicity study of chitin in F344 rats]. AB - A subchronic toxicity study of chitin, a natural structural component of crustacean shells, was performed in F344 rats by feeding of the powdered diet containing 5%, 1.7%, 0.6%, 0.2%, and 0% concentrations of the substance. Each group consisted of 10 males and 10 females. All animals survived until the end of the experiment. There were no changes indicating obvious toxicity of chitin in the clinical signs, body weight, food intake, hematology, serum biochemistry, or histopathological findings, except a slight decrease in body weight gain in the 5% chitin-treated males. Although the mechanism is unclear, the suppression of body weight gain may be due to the slight decrease in caloric content of the food in the 5% chitin-treated animals, a change unrelated to toxicity. Thus, there was no obvious toxicity of chitin in F344 rats at concentrations up to 5% in the diet for 13 weeks. PMID- 10859947 TI - [Evaluation of molecular weight of hyaluronate preparations by size-exclusion chromatography]. AB - Hyaluronate (HA), a glycosaminoglycan polysaccharide, has been used as a biomedical polymer to treat osteoarthritis by intra-articular injections and in ophthalmic surgery, such as anterior segment surgery. In this study, the molecular weight (MW) of HA preparations was estimated by size-exclusion chromatography (SEC), using HA and pullulan fractions as molecular weight standards, and the MW values obtained were compared to those obtained with a low angle laser light scattering detector (LALLS). The results showed that the universal calibration with pullulan as the standard is useful for HA preparations. The conditions of SEC for HA were also investigated, and the results suggested that a high ionic strength and low flow rate of the eluent were preferable for high molecular weight HA preparations. PMID- 10859948 TI - Somatic embryogenesis and ginsenoside production of Panax ginseng in phytohormone free medium. AB - Embryogenic cultures of Panax ginseng were established without using phytohormones. Somatic embryos developed from the roots of an in vitro seedling and from excised leaf and petiole segments cultured in half-macro-salt strength Murashige and Skoog medium. Excised leaf and petiole segments were obtained from in vitro germinated seedlings. Plantlets were subsequently obtained from developing somatic embryos in phytohormone-free media. Shoot formation from somatic embryos was influenced by light intensity. The rate of growth and frequency of embryogenesis were improved when cut-up embryogenic tissues were inoculated into liquid media in the dark. The ginsenoside contents of a 4 year old field-cultivated root, seedlings from zygotic embryos, somatic embryos and embryogenic tissues were determined and compared. Somatic embryos contained 1.7 times the amount of ginsenoside Rb1 and 2.3 times the amount of ginsenoside Re compared to seedlings from zygotic embryos. Ginsenoside Rd, which was absent in the seedlings derived from zygotic embryos, was detected in somatic embryos. Higher ginsenosides Rd and Rg1 levels were found in embryogenic tissues grown on solid media than in tissues grown in liquid media. The total ginsenoside yields, including the ginsenosides Rb1 and Rg1 levels, of cut-up embryogenic tissues, were higher than those of clump tissues. PMID- 10859949 TI - [Triamcinolone Acetonide Reference Standard (Control 981) of National Institute of Health Sciences]. AB - The raw material of triamcinolone acetonide was examined for preparation of the "Triamcinolone Acetonide Reference Standard (Control 981)". The analytical data obtained were: melting point, 289 degrees C (decomposition); UV spectrum, lambda max of 238 nm; IR spectrum, same as that of the Triamcinolone Acetonide Reference Standard (Control 834); optical rotation, [alpha]D20 = +106.8 degrees; thin-layer chromatography, no impurities detected; high-performance liquid chromatography, total amount of impurities less than 0.4%; loss on drying, 1.3%; assay by HPLC, 100.1%. Based on the above results, the raw material was authorized as the Triamcinolone Acetonide Reference Standard (Control 981) of the National Institute of Health Sciences. PMID- 10859950 TI - [Triamcinolone Reference Standard (Control 981) of National Institute of Health Sciences]. AB - The raw material of triamcinolone was examined for preparation of the "Triamcinolone Reference Standard (Control 981)". The analytical data obtained were: melting point, 246 degrees C (decomposition); UV spectrum, lambda max of 239 nm and specific absorbance in methanol at 289 nm of 394; IR spectrum, specific absorptions at 3462, 1716, 1659, 1615, 1604, 1132 and 1061 cm-1; optical rotation, [alpha]D20 = +69.7 degrees; high-performance liquid chromatography, five impurities detected and amount of each impurity estimated to be less than 0.6% and total amount of impurities less than 1.4%; loss on drying, 0.24%. Based on the above results, the raw material was authorized as the Triamcinolone Reference Standard (Control 981) of the National Institute of Health Sciences. PMID- 10859951 TI - [Laparoscopic cholecystectomy. Our experience]. AB - BACKGROUND AND AIMS: The authors analyse their first two years of experience regarding the use of laparoscopic cholecystectomy and report the results of this series and their observations. METHODS: A total of approximately 200 cholecystectomies have been performed using a laparoscopic technique at S. Paolo Hospital in Naples (Department of Surgery; Head: Prof. R.A. Caliendo) since October 1996. Initially patients were rigorously selected in accordance with international criteria. At present, after the good results obtained, the real contraindications to laparoscopic cholecystectomy are considered to be: a) the patient's refusal; b) general conditions that do not allow the use of general anesthesia in surgery. RESULTS: None of these 200 patients was converted to open surgery, although six complications occurred: four of these were cases of bleeding in which it was decided to choose a carefully monitored wait-and-see approach. Mean postoperative hospitalisation was two days. No major postoperative complications were observed and it was not necessary to keep any patient for more than five days. CONCLUSIONS: On the basis of this experience, the authors underline the value of laparoscopic surgery versus open cholecystectomy (reduced morbidity, shorter hospitalisation with marked social and economic advantages). Furthermore, the authors emphasise that, when laparoscopy is first introduced, it is important for the same operating team always to be present in order to ensure a good learning curve and increasing familiarity with the technique. PMID- 10859952 TI - [Gasless laparoscopic cholecystectomy. Our experience with 130 cases compared with 450 cases treated with the CO2 technique]. AB - Alongside the technique based on the creation of an abdominal cavity for surgery following the introduction of gas (usually CO2) into the peritoneal cavity, a new method has been developed. This involves the use of an atraumatic mechanical lifting device connected to the same abdominal wall (gasless laparoscopy). The authors report a technique that uses an inflatable cushion inserted into the abdomen through a periumbilical incision. The cushion is connected to an external motorized hydraulic jack fixed to the operating table, fitted with an electric motor and friction gear. Between May 1991 and June 1998, 580 patients underwent laparoscopic cholecystectomy. Since December 1995 a total of 130 patients have undergone surgery using gasless laparoscopy. Shoulder pain and pain in the upper abdominal quadrant were no longer reported; pain was present in 70% of the patients operated using the CO2 technique. There was also a marked reduction in the anesthesiological risks, above all in elderly patients with cardiopulmonary insufficiency. Surgical manoeuvres are made easier owing to the possibility of using traditional surgical instruments. Washing and continuous aspiration allow a good control of intraoperative hemostasis, and reduce the phenomenon of lens misting without the risk of losing pneumoperitoneum. Less visibility of the surgical field was reported, particularly in obese patients, above all because of the reduced diaphragmatic distension and the lack of displacement of the intestinal loops. In the authors' opinion the gasless technique is suitable above all in patients affected by cardiopulmonary disorders in whom hypercapnia might represent a significant operating risk. PMID- 10859953 TI - [Peritoneal innervation and post-laparoscopic course. Role of CO2]. AB - BACKGROUND: 30-67% of patients undergoing laparoscopic surgery reports shoulder pain. Besides, post-surgical course of patients undergoing converted laparoscopic procedures is similar to the course of patients who received a completely laparoscopic procedure. It is supposed that there is a temporary neurotoxic damage of the peritoneal sensitive nervous fibres defined by CO2. METHODS: A prospective review has been carried out by histologically analyzing 38 peritoneal biopsies from 10 selected patients, during different laparoscopic surgical procedures (6 cholecystectomies, 2 appendectomies, 1 selective bilateral ligature of the spermatic vessles) and at different times during each operation. Patients whom anamnesis, clinical or local conditions were suggestive for peritoneal flogosis were excluded from the study: therefore only 29 biopsies from 8 patients have been considered useful to the study. RESULTS: Histological analysis has been carried out with different methods of coloration (hematoxylin eosin, argentic staining) and at different magnifications (30x, 60x, 100x), without electronical microscopy or immunohistochemical studies. No biopsy showed signs of damage of the nervous structures. CONCLUSIONS: Certainly, the realization of a pneumoperitoneum at CO2 doesn't cause damages of the peritoneal sensitive fibres. It has been demonstrated that the abdominal introduction of CO2 causes a "relative peritoneal acidosis", directly depending from the percentage of CO2 employed: the peritoneal pH decreases to 6.9 after 15 min of pneumoperitoneum with CO2 at 100% and to 7.35% with CO2 at 5% of air. Probably this condition causes a temporary biochemical change that defines reduction of the nervous impulses and, therefore, the "peritoneization" of the patient subjected to laparoscopic procedure. The "biochemical hypoesthesia", based on a change of the peritoneal homeostasis, would translate itself in a beneficial effect for the patient, persisting also when converted to laparotomic operation due the impossibility to proceed under laparoscopy, held up by the residual pneumoperitoneum. PMID- 10859954 TI - [Preliminary results of biliopancreatic diversion in the treatment of morbid obesity. Clinical considerations on 69 patients with a 3-year follow-up]. AB - BACKGROUND AND AIMS: The aim of this study was to evaluate the results obtained using Scopinaro's biliopancreatic diversion technique (AHS-BPD) in the surgical treatment of morbid obesity. METHODS: A retrospective study was carried out in 69 patients with a follow-up ranging between 6-44 months. All patients were operated and monitored by the Obesity Surgery Centre operating in Sardinia since february 1995 at the Department of Emergency Surgery of Sassari University. All the patients were severely obese with a mean BMI of 51.58 and, in the majority of cases, presented associated metabolic diseases with the following incidence: type 2 diabetes in 40.57%, arterial hypertension in 36.23%, severe alteration of lipid status in 52.17%; in overall terms, a plurimetabolic syndrome was present in 24.63% of cases. All patients underwent biliopancreatic diversion using Scopinaro's classic technique (AHS-BPD). Controls were carried out at set intervals (1-3-6-12-18 and 24 months) to evaluate weight loss and the metabolic effects of surgery in terms of the lipid, glucose and protein status. RESULTS: Results were good, as confirmed by the marked weight loss (BMI after 24 months: 30) and the normalisation of cholesterol and glycemia. No major reductions were observed in proteinemia and albuminemia levels. CONCLUSIONS: In the light of these results, the authors affirm that Scopinaro's technique is a valid solution for the treatment of morbid obesity. Its relatively invasive nature is justified by the results obtained in terms of weight control and its effect on associated metabolic diseases. PMID- 10859955 TI - [Severe acute pancreatitis. Clinical, diagnostic, and therapeutic features]. AB - BACKGROUND: The aim of this study is to define the actual role of surgical therapy in severe acute necrotizing pancreatitis. METHODS: A retrospective analysis has been carried out on the surgical treatment of severe acute pancreatitis at the Institute of General Surgery and Surgical Specialties, University of Siena (Italy). From January 1980 to December 1997, 230 patients affected by acute pancreatitis were admitted to institution: 24 patients affected by severe disease (necrotizing pancreatitis: clinical and radiological diagnosis, by CT-scan) was choosen for this study. Of 24 patients, 15 were males and 9 females, with mean age of 55 years (range 30-80). In all cases, surgical procedure consisted in pancreatic necrosectomy, multiple abdominal and retroperitoneal drainage and closed management; operated patients with biliary pancreatitis underwent colecystectomy and, if necessary, common biliary duct drainage. RESULTS: The patients underwent surgical procedure, but the remaining 14 were treated by intensive medical care: mortality in these two groups was respectively 40% (4 cases) and 21% (3 cases). CONCLUSION: The conclusion is drawn that intensive medical care is the first therapeutic approach in severe acute pancreatitis, reserving surgery only to selected cases, as those affected by pancreatic infectes necrosis or those who get worse despite of conservative therapy. As to surgical technique, closed procedures vs open or semiopen, and conservative surgery (necrosectomy, multiple drainage and abdominal washing) vs anatomical resection are preferred. PMID- 10859956 TI - [Surgery of large ventral hernias. Personal experience in 1990-1997]. AB - BACKGROUND: The surgical treatment of large wall defects conventionally defined as an extension over 10 cm is discussed. The difficulty to contain the bowels that have lost law of domicile in the abdominal hollow, constitutes motive for notable increase of the endo-abdominal Pressure with serious consequences in the postoperative course and this leads to the use of prothesis meshes that allow the closing of the abdominal hollow with the Tension-Free technique. METHODS: Personal experience embraces 45 patients, with large wall defects, divided into 21 patients with overumbilical location, 14 with umbilical location, 10 with periumbelical location; a simple suture has been used in 7 cases, the reconstruction of the wall according to Stoppa in 36 cases and the apposition of Goretex net internally and Marlex net externally in 2 cases. RESULTS: There have been neither mortality, neither recidivists of illness, but only some complications: 9 cases of superficial infection, 1 case of intestinal occlusion and 2 of subcutaneous seroma. CONCLUSIONS: According to their experience and wide literature review, the authors draw some conclusions: an accurate toilet and a careful evaluation of the respiratory functionality are fundamental; it's necessary to postpone surgical intervention in presence of local inflammation and, where this is improrogable it's opportune to avoid the use of prothesis meshes or refold on readsorbible prothesis; special care must be taken to the hemostasis and an aspirative drain for 24-48 hrs preserves from the risk of postoperative hematomas and following local infections. The submuscular mesh permits a Tension-Free suture and for this reason it would have nowdays a more extensive use. Finally it's pointed out the choice of a PTFEe mesh in contact with the intestinal skein. PMID- 10859957 TI - [Treatment of inguinal hernia. Case reports]. AB - BACKGROUND: The aim of this study was to evaluate the extent to which inguinal hernia surgery has changed over the past few years. As confirmed by the bibliography, the authors draw attention to the fact that the use of more resistant and well tolerated heterologous graft materials has led to the rapid spread of "tension-free" techniques which have now replaced conventional methods with good long-term results. The development of these techniques has been flanked by the growing use of day hospitals, leading to a marked reduction in hospitalization costs. METHODS: The authors describe the series of hernia operations performed by a day hospital linked to a general surgery ward over the past six years. The series refers to the 1st division of general surgery at Ospedale Nuovo Martini in Turin and covers the period from January 1993 to July 1998 with a total of 1387 patients. RESULTS: Over this short period, the percentage of inguinal hernia operations in inpatients fell from 92.9 to 22.4% while those undergoing surgery in the day hospital rose from 7.1 to 77.6%. This was flanked by a move away from conventional plastic surgery (Bassini, Shouldice) in favour of Lichtenstein's technique which was used in over 76% of monolateral hernia and over 55% of bilateral cases. Local anesthesia induced by the surgeon was used in the majority of cases (98%). Mersilene prostheses were initially used but were soon replaced by prolene grafts, both of which were well tolerated in over 84% of cases. Short-term antibiotic prophylaxis was administered in all cases, whereas the use of postoperative painkillers was limited to minor analgesics. CONCLUSIONS: The surgical treatment of inguinal hernia has changed drastically over a relatively short period with regard to both operating techniques, as is shown by the widespread use of prostheses, and the indications for day hospital surgery which have gradually increased owing to the reliability and tolerability of hernia mesh and the use of local anesthesia. PMID- 10859958 TI - [Duodenal and pancreatic injuries]. AB - BACKGROUND: Pancreatic and duodenal injuries occur rather infrequently and the incidence ranges between 1% and 12% of all abdominal injuries. The high rate of mortality and morbidity (10-40%) depends on associated complication rate of all intra-abdominal organs (90%). METHODS: Twenty-five cases of pancreatic and duodenal injuries observed between 1987 and 1997, with an incidence of 0.7% of all abdominal injuries, are reported. In 16 cases the cause was penetrating injury (gunshot) and in 9 cases it was blunt abdominal trauma. Only two patients presented an isolated pancreatic lesion, all the others had at least an associated lesion. In all the cases the patients were male and they were submitted to emergency laparotomy. RESULTS: The mortality rate was 20%, the morbidity was 24%. CONCLUSIONS: The relatively low incidence of these injuries and the high rate of associated lesions cause a difficult diagnostic and therapeutic approach, the absence of a unified method to follow and the unsatisfactory results observed. PMID- 10859959 TI - [Our experience with the Hartmann's operation in colorectal emergencies]. AB - BACKGROUND: The role of Hartmann's operation has been revised during the past few years in the context of emergency colorectal surgery: it represents an obligatory choice that enables the simultaneous treatment of the primary disorder and the complication. This study aims to emphasise the importance of this unique surgical choice and to stress that surgeons should not underestimate it. METHODS: The authors review the literature on the subject and make a retrospective analysis of 228 cases of colorectal surgery from 1988 to 1997 in which Hartmann's operation was performed in 16 patients with the following indications: Hinchey's stage III and IV peritonitis secondary to perforating diverticulitis of the sigma (elective indication) or occlusion of the left colon when preparation could not be accomplished in spite of intraoperative washout. RESULTS: Post-Hartmann recanalisation was successfully performed in 14 patients. CONCLUSIONS: The authors' experience and these results concord with the general view that this operation should be reserved for selected cases, in particular colorectal emergencies of a perforating nature; it is less appropriate for intestinal occlusion, although it is always preferable to be too prudent by resorting to Hartmann's operation or protective colostomy rather than risk anastomotic dehiscence. PMID- 10859960 TI - [Hemorrhoid disease. Physiopathology, etiopathology and surgical approach]. AB - The authors report a number of cases of hemorrhoid disease and describe the therapeutic iter followed with particular reference to the surgical approach used. After a description of the physiopathological aspects linked to the disease, bearing in mind the use of electromanometry and electromyography in diagnosis, the authors underline the contemporary presence of varices in the lower limbs and hemorrhoid disease, as well as the frequent finding of hemorrhoids in a syndrome of portal hypertension. They then affirm how it is impossible to establish the causes of this pathology with any certainty and how a single standardised treatment plan is untenable. The authors then indicate the guidelines used to choose the most appropriate form of surgery rather than pharmacological treatment, based on the ideal cases and conditions for surgery. The ultimate goals of surgery are also outlined. The study compares four possible surgical techniques, providing synthetic information regarding their adaptability to the various cases treated and the characteristics of their use. This means that, once decided, surgery must successfully resolve the patient's problems. In conclusion, once hemorrhoid disease has been diagnosed, it is important to intervene with appropriate medical treatment to control the evolution of the pathology; if this is not sufficient, surgery becomes an inevitable choice. PMID- 10859961 TI - [Diabetic foot. Physiopathology, clinical aspects, and recent therapeutic approaches]. AB - Diabetic foot is a complication of diabetes mellitus occurring in 15% of patients that is of specific surgical interest. Over the past few years, preventive measures and the use of new therapeutic resources has reduced the number of patients undergoing demolitive surgery. The authors present a concise but at the same time sufficiently detailed picture of modern knowledge of the physiopathology, clinical aspects and current therapeutic guidelines for diabetic foot. In particular, they analyse the validity of various forms of complementary treatment to surgery, including techniques to stimulate tissue repair processes, hyperbaric oxygen therapy and laser therapy, and they underline the importance of using a multidisciplinary approach to this pathology. To this end, they review all the articles on the subject reported on Medline from 1992 to June 1998, presenting and commenting on the most significant results. PMID- 10859962 TI - [Bilateral Morgagni-Larrey hernia. Report of a clinical case]. AB - A rare case of bilateral diaphragmatic hernia, containing a long segment of transverse colon in the left breach and most of the omentum and a short segment of colon in the right one, is described. Diaphragmatic hernia can occur at any age in both males and females and it is often asymptomatic and diagnosed by chance. The epidemiological, etiopathogenic, clinic, diagnostic and therapeutic aspects of the disease are discussed. The only treatment, which is advisable in asymptomatic patients too, is the closure of the hernial orifice. This closure can be done directly or through the insertion of prosthetic material. The conventional access can be thoracic or abdominal, but laparoscopy is going to replace both of them successfully in the future. PMID- 10859963 TI - [Acute diverticulitis of the right colon. Report of 3 cases]. AB - Three cases of acute diverticulitis of the right colon are described and a review of the international literature about this rare disease is made. Diverticulitis of the ascending colon is an uncommon disease which mimics appendicitis or perforated neoplasm. The correct diagnosis is rarely made, but can be suggested by signs and symptoms and confirmed by the barium enema. It should be treated conservatively, like left-colon diverticulitis. Isolated diverticula can be managed by simple excision and primary closure. Local resection may be necessary if the involved area is too large. Right hemicolectomy is performed when carcinoma is strongly suspected. PMID- 10859964 TI - [Essential varicocele. Which treatment?]. AB - Purpose of the paper is to sum up the problem of surgery of idiopathic varicocele according to the present possibilities, both surgical and sclerotic. A wide review of the literature underlines a high rate of relapses and persistence of the disease (with a percentage from 10 to and 10%) following the two most used technique: retrograde sclerotic therapy under radioscopic control and surgical retroperitoneal or inguinal ligature of the internal spermatic vein; this technique was preferred by us until 1997. The percentage of failures, high with reference to the benign form of the disease (over 10% in our series of more than 100 patients submitted to clinical and flow-meter examinations) let the authors suggest a combined and simultaneous operation of ligature both internal and external of the spermatic vein at the level of the internal inguinal ring. Anatomical reasons confirm the opportunity of this procedure since the involvement of the system of the external spermatic vein is present in about the 20% of the cases of idiopathic varicocele in accordance with various flebografic studies. The possibility of escape through the external spermatic vein is eliminated in the case in which such vessel is preserved, and it seems frequent above all in 3rd degree idiopathic varicocele where many anastomotic vessels between the two systems are present. This procedure can be made both in general or local anaesthesia, it doesn't involve postoperative hospital stay and present the same acceptable postoperative complication of other proposed operations. The laparoscopic treatment even if easily performed at the level of the internal inguinal ring, doesn't seem justified for the higher cost and equal compliance for the patient. Besides, it is not possible to proceed laparoscopically under local anaesthesia. PMID- 10859965 TI - [Subarachnoid hemorrhage in ruptured aneurysm in the year 2000: multidisciplinary approach]. PMID- 10859966 TI - [Subarachnoid hemorrhage imaging]. PMID- 10859967 TI - [Timing of surgery in subarachnoid hemorrhage]. PMID- 10859968 TI - [Surgery of intracranial aneurysm: innovations]. PMID- 10859969 TI - [Endovascular occlusion and percutaneous treatment of vasospasm]. AB - Current available minimal invasive endovascular technology allows for percutaneous treatment of cerebral aneurysms and of vasospasm, the associated disease fo cerebral arteries secondary to subarachnoid hemorrhage. Multicentric evaluation of standardized treatment techniques using platinum coils have shown that a high success rate is now obtained for certain forms of aneurysms, mostly those which have a small base (4 mm or less) of implantation of the vessel. Treatment of arterial vasospasm by techniques including angioplasty using soft silicon balloons, or intraarterial application of spasmolytics, or both may be very successful, when performed prior to establishment of irreversible ischemic damage to the brain tissue. An intensive patient monitoring during the posthemorrhagic period is therefore critical for the correct timing of such a procedures. PMID- 10859970 TI - [Subarachnoid hemorrhage: physiopathologic features and preoperative optimization]. PMID- 10859971 TI - [Peroperative anesthesiologic management and neuro-monitoring of patients with subarachnoid hemorrhage]. PMID- 10859972 TI - [Peroperative risks in the surgical treatment of cerebral aneurysm]. PMID- 10859973 TI - [Prevention of vasospasm secondary to subarachnoid hemorrhage caused by ruptured aneurysm: management]. PMID- 10859974 TI - [Indications for angiography in vasospasm]. PMID- 10859975 TI - [Vasospasm treatment in intensive care]. AB - Vasopasm is a dreadful complication of SAH associated with an important mortality and morbidity. Therapy begins with adequate monitoring and lines, and prevention of secondary brain injuries. 3-H therapy (hypervolemia--hypertension- hemodilution--hyperdynamism) aims to increase perfusion in ischemia areas. 3-H therapy is associated with systemic complication precluding it's prophylactic use. Calcium antagonists, in particular nimodipine, improve outcome and parenteral route is better than oral administration. Tirilazad seems to improve outcome of severe grades. Numerous experiments are performed with drugs interfering with the biochemical cascade leading to vasospasm, but up to today no drug is used in current clinical practice. Intraaortic balloon is still considered as experimental and may have a role in patients presenting with concomitant cardiac failure. Invasive radiology must be considered in vasospasm not improving with standard therapies. Vasopasm is a dire complication after SAH. Support and specific therapies allow a 3-fold reduction in morbidity associated with vasospasm. Vasospasm is a vital emergency, and intervention has to be quick and aggressive. PMID- 10859976 TI - [Neuroradiologic treatment of cerebral vasospasm]. PMID- 10859977 TI - [The voice]. PMID- 10859978 TI - [Alcoholism: short interventions for consumers at risk]. PMID- 10859979 TI - [Unemployment and health]. AB - Unemployment is a severe problem in Europe since 1980-1990, which is not only economic but also social and medical. European studies show that morbidity and mortality rates of unemployed persons and their families are consequently higher than in the general population. The longest the unemployment period, the more you troubles in physical and psychological health. Depression and psychosomatic trouble are the most frequently observed, secondary to stress, emotional perturbations and psychological burn-out. It is necessary to develop information and prevention strategies to identify sick unemployed persons, to treat them early to help them to recover their health and to try to reduce socioeconomics costs, pain and sicknesses in this specific fragile population. We should also try to inform public opinion and reach all the professionals concerned by unemployment to work together and look for the best solutions to this medical, psychological, social and economical problem. PMID- 10859980 TI - [Type A botulinum toxin: a new methods in treating focal hyperhidrosis. A summary of various possibilities in hyperhidrosis therapy with special emphasis type A botulinum toxin injections]. AB - Hyperhidrosis is defined as an excess of sweating over the amount necessary for thermoregulation. Essential focal hyperhidrosis is a overactivity of the sweat glands of the axilla, palms and soles probably due to a disorder of the sympathetic nervous system. The therapy is difficult, even though there are many therapeutic options. Beside the effort to treat the psychovegetative disorder (autogenic training or acupuncture), there are efforts to seal the lumen of terminal sweat ducts using aluminiumchlorhydroxide application or iontophoresis. Surgery has the aim to eliminate sweat glands either by excision or by denervation. It is also possible to use chemical denervation with systemic anticholinergics. Only recently the local chemodenervation with injections of botulinum toxin (BTX) was added to the therapeutic tools of focal hyperhidrosis. We present an overview of several therapeutic options in consideration of BTX. PMID- 10859981 TI - [Senile epilepsy]. AB - Epilepsy is an important factor of neurologic morbidity in the elderly population. The incidence is higher in the age group over 65 years than in children and adolescents. Cerebrovascular disturbances are the most frequent cause of seizures in the elderly. The so-called idiopathic epilepsies on the other hand are rather rare. The frequent co-morbidity and co-medications have to be taken in consideration when evaluating and treating such patients. There is a number of efficacious antiepileptic drugs at disposition. The selection of the drugs and their dosage have to be adapted to the needs of the particular age group. PMID- 10859982 TI - [Recurrent severe lower gastrointestinal hemorrhage. Acute recurrent diverticulum hemorrhage in the cecum]. AB - This 83-year-old patient was admitted to the emergency room after a sudden and abundant rectal bleeding without any other abdominal symptoms. The physical examination showed normal abdominal findings. However, digital rectal examination revealed dark red blood. Upper and lower endoscopic examination did not show an active bleeding source. A diagnosis of an acute lower gastrointestinal bleeding probably originating from a diverticulum was made. The patient was dismissed two days later without any signs of further bleeding. However, the same day he was readmitted again because of acute rebleeding. Emergency colonoscopy was not conclusive because of massive hemorrhage. Angiography of the abdominal arteries was performed which showed marked active bleeding in the coecal region. Ileocoecal resection was performed and histopathological examination showed a solitary coecal diverticulum. PMID- 10859983 TI - [Acute necrotizing hepatitis: an unusual side effect of oral anticoagulants]. AB - We report the case of a severe relapsing phenprocoumon-induced hepatitis. The first episode of hepatitis was thought to be caused by another drug (Verapamil). The anticoagulation with Phenprocoumon was therefore continued after healing of liver inflammation. The relapse typically developed after a shorter exposition time supporting the hypothesis of an allergic etiology. Fortunately we didn't find any cross-reaction between Phenprocoumon and Acenocoumarol. The patient could thus be anticoagulated orally without complications. If long term anticoagulation is absolutely essential, it is reasonable to prescribe a different Coumarin-derivate. In the case of a cross-reaction, the therapy should be continued with low-molecular weight heparin. PMID- 10859984 TI - [Traumatic perforation of the small intestine--a rare complication of inguinal hernia]. AB - Groin hernias are among the most common surgical diseases. Traumatic rupture of the intestine is a rare complication compared to incarceration. We report on a 90 year old male with a left-sided hernia and a traumatic rupture of the small bowel. The treatment consisted of an emergency laparoscopy with simple closure of the perforation, repair of the hernia by herniorrhaphy and drainage of the abdomen. PMID- 10859985 TI - [Macular exanthema in HIV infection]. PMID- 10859986 TI - [Epidemiology of heart failure]. AB - During the past decades incidence and prevalence of heart failure has markedly increased. This contrasts with most other cardiovascular diseases, which have declined. Among people aged 80 years and over prevalence may be as high as several percent of the population. According to the high prevalence of the disease, the high frequency of outpatient consultations, but in particular because of a high rate of hospitalizations with long average duration, costs for the health care system are very high. Patients with heart failure suffer from a reduction in quality of life exceeding that of most other chronic diseases, and mortality after diagnosis of heart failure remains high. PMID- 10859987 TI - [Pathophysiological basis of heart failure]. AB - The progression of heart failure is related to local and systemic neuroendocrine activation. On the level of the myocardium, neuroendocrine activation (angiotensin II, endothelin, aldosterone, norepinephrine) as well as mediators of inflammation and free oxygen radicals contribute to hypertrophy, dilation and remodeling of the ventricles. In addition, vascular endothelial dysfunction and alterations of skeletal muscle contribute to clinical symptoms of heart failure patients. Changes in ventricular geometry during the progression of cardiac disease are associated with specific subcellular alterations on the level of the myocytes. Especially, disturbed intracellular Ca2+ handling resulting in altered excitation contraction coupling may lead to impaired systolic and diastolic function. Disturbed Ca2+ homeostasis has been associated with reduced re-uptake capacity of the sarcoplasmic reticulum for Ca2+ and an enhanced activity of the sarcolemmal Na+/Ca(2+)-exchanger. In consequence, alterations in force-frequency behavior were attributed to a decline in intracellular Ca2+ transients at higher stimulation rates. The reduced expression and desensitization of myocardial beta adrenoceptors and alterations on the level of the G-proteins result in a reduced basal and catecholamine-stimulated activity of adenylate cyclase and a reduction in intracellular cAMP content. In consequence, reduced phosphorylation of intracellular functional proteins in the failing human heart contributes to altered Ca2+ handling. The Frank-Starling mechanism seems to be unaltered in isolated human myocardium from failing hearts. Endothelin and angiotensin may contribute to the regulation of myocardial contractility in the human heart, but their functional relevance in the regulation of myocardial contractility under clinical conditions remains to be evaluated. PMID- 10859988 TI - [Clinical diagnosis of heart failure]. AB - The clinical diagnosis of heart failure is based upon history and physical examination. Careful questioning and examination requires understanding of the pathophysiology of this systemic disorder. Symptoms and signs of congestive heart failure need to be differentiated from the manifestations of the underlying cardiovascular disorder. Only then will the specific signs and symptoms be unraveled. Symptoms arise from pulmonary congestion and peripheral or organ underperfusion. Findings related to congestion can be found over the lungs (rales, pleural effusion), or at the jugular veins displaying either frank central venous pressure elevation or paradoxic inspiratory venous pressure rise (Kussmaul sign), or the more discrete sign of right, left of biventricular failure, the hepatojugular reflux. Dilatation and hypertrophy of the cardiac chambers can clinically easily and reliably be assessed by careful palpation. Galopprhythm, right and/or left ventricular in origin, is a particularly reliable sign of a failing ventricle. While a presystolic, atrial sound indicates merely elevated resistance to ventricular filling, i.e. the presence diastolic dysfunction or increased chamber filling, is the ventricular diastolic galopp a reliable sign of ventricular failure. Especially the appearance of a quadruple rhythm or a summation galopp can be considered both highly specific as well as prognostically dubious. Relative mitral and/or tricuspid insufficiency as a sign of ventricular dilatation needs to be differentiated from organic valve disease. This requires often echocardiography. Oedema of cardiac origin is symmetric and more pronounced in the evening. It arises both from left and from right ventricular failure. History and physical examination are both reliable tools in the initial diagnosis, as well as during follow-up and for control of therapeutic measures. Technical methods, such as chest x-ray, echocardiography or else are used for quantification and documentation. Properly applied and utilized they allow the physician to sharpen his clinical acumen, thus allowing for both a reliable diagnosis and a semi-quantitative estimation of ventricular size, enddiastolic and atrial, as well as pulmonary pressures and valve function. PMID- 10859989 TI - [Non-invasive diagnostic procedures for heart failure: echocardiography, myocardial perfusion scintigraphy and radionuclide ventriculography]. AB - Even though the diagnosis of heart failure can often be made at bedside from the patient's history and a thorough physical examination, some mechanical examinations are needed to objectify the clinical symptoms. Due to the chronic and progressive character of heart failure with repeated periods of clinical treatment, non-invasive and reproducible methods are of great importance. Thanks to technical progress in data-acquisition and -processing, echocardiography has become one of the most important diagnostic tools in clinical cardiology. Many questions associated with the diagnosis and treatment of heart failure can be answered with the aid of the wide spectrum of echocardiography, including morphological description in conventional M- or B-Mode and functional analysis based on Dopplertechnique. On the other hand well-established nuclear techniques like myocardial perfusion scintigraphy and radionuclide ventriculography profit from recent technical development. In addition, the modification, specification and standardisation of examination protocols improve the diagnostic reliability of nuclear medical techniques. Performed by a specialist, both principles of examination--ultrasound and scintigraphy--are valid in the diagnosis of heart failure and in therapy-management. PMID- 10859990 TI - [Atrial natriuretic peptides: diagnostic and therapeutic potential]. AB - The natriuretic family consists out of three molecules that share significant amino acid sequence homologies and a looped motif. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are similar in their abilities to promote natriuresis and diuresis, to inhibit the renin-angiotensin aldosteroneaxis and to act as vasodilators. The understanding concerning the actions of the C-type natriuretic peptide is incomplete, but this new family member acts as a vasodilator. Plasma levels of ANP and BNP are elevated in patients with unstable angina, acute myocardial infarction, and with congestive heart failure. BNP may be superior to ANP as a prognosticator for risk stratification after myocardial infarction and is independent of left ventricular ejection fraction. ANP and BNP have little therapeutic potential while experimental work as well as clinical trials suggest that the inhibition of the catabolism of natriuretic peptides in particular in combination with ACE inhibitors may be clinically beneficial. PMID- 10859991 TI - [Therapy for heart failure]. AB - The treatment of congestive heart failure focuses on three steps: 1. Elimination of the precipitating cause or mechanism, and/or treatment of the underlying disease respectively. 2. Treatment of the failing heart syndrome itself. We shall concern ourselves with pharmacotherapy, omitting technical and surgical aspects. 3. Prophylactic treatment of complications, such as thromboembolism and arrhythmias. Drugs for the treatment of heart failure can be classified as follows: 1. Diuretics 2. Vasodilators 3. Neurohumoral Inhibitors 4. Inotropic drugs. Diuretics improve symptoms and exercise capacity and probably survival. They are the drug of first choice in acute and chronic heart failure. Potassium supplementation is necessary. Renal function needs to be monitored. The aldosterone antagonist spironolactone has probably important effects upon the myocardium. It retards fibrous tissue development and improves prognosis. Vasodilators unload the heart and improve contractile geometry and hemodynamics, thereby lessening symptoms. Prognosis, however, is not affected. They are indispensable in acute heart failure. In longterm treatment only the combination of nitrates with hydralazin has been shown to be effective. Angiotensin converting enzyme inhibitors combine vasodilation with neurohumoral inhibition. They are most effective in improving symptoms, exercise capacity and surviving chronic heart failure. If side effects (cough, allergy) prevent their use, then angiotensin II receptor antagonists can be used with equal benefit. However, both groups of drugs impair renal function and cannot be given in advanced renal failure or renal artery stenosis. Beta-receptor antagonists, previously considered contraindicated in heart failure are today amongst the most important drugs in heart failure. They improve survival and retard the need for cardiac transplantation in advanced failure. Their use, however, is rather difficult requiring extremely slow dose titration beginning with very low doses. Inotropic drugs are today mainly used in acute failure and cardiogenic shock. In longterm treatment only the digitalisglycosides have been shown to be effective in improving symptoms, exercise capacity and the general clinical course. Often antiarrhythmic treatment is necessary. Here amiodarone is the drug of choice today if beta blockers do not suffice. Prophylactic anticoagulation is indicated in all cases NYHA III and IV, with large hearts already in II. Future developments may include new inotropes, the ANP-system, and cytokines, as well as gene therapy for correction of myocardial phenotype change. PMID- 10859992 TI - [Ischemic vs nonischemic heart failure--does etiology matter?]. AB - In most large scale trials the prognosis of ischemic heart failure is worse than in patients with non-ischemic etiology. The therapeutic effect of essential drugs such as ACE-inhibitors, betablockers and diuretics is similar, but response to some other drugs (amiodarone, amlodipine, digoxin, growth hormone) is better in non-ischemic heart failure. Of great practical importance is the recognition of hibernating myocardium in coronary artery disease, since revascularisation may significantly improve left ventricular function. Specific therapeutic interventions are possible in hypertensive heart disease, alcoholic cardiomyopathy and LV-dysfunction to tachyarrhythmias. The etiology of heart failure should therefore be cleared in all patients. PMID- 10859993 TI - [Antiarrhythmic therapy in patients with heart failure]. AB - In patients with severe chronic heart failure, many deaths are sudden due to life threatening ventricular arrhythmias. Supraventricular arrhythmias such as paroxysmal or chronic atrial fibrillation may also cause serious complications in those patients due to acute loss of atrial contraction, pump failure during rapid ventricular response and embolic events. Two therapeutic strategies are currently available for therapy and prevention of malignant ventricular arrhythmias and subsequent sudden arrhythmic death: antiarrhythmic drug therapy and implantable defibrillators. However, selection of the most beneficial strategy for the individual patient to reduce the risk of sudden death remains a major challenge in cardiology. Betablockers exert a favorable antiarrhythmic action without increasing proarrhythmia, thus betablockers may serve as a basic medication in patients at risk for sudden death. However, the general use of antiarrhythmic drug therapy for symptomatic ventricular arrhythmias is not recommended, as these drugs have been shown to increase mortality in patients with severe congestive heart failure due to proarrhythmic or negative inotropic effects (e.g. class Ia antiarrhythmics). Even class III antiarrhythmic drugs such as amiodarone, which has been studied sufficiently in patients with left ventricular dysfunction, is not effective enough for significant reduction of cardiac mortality in patients with symptomatic ventricular arrhythmias and depressed ventricular function (e.g. EMIAT, CAMIAT). But as a positive result of available studies, amiodarone does not increase mortality in those patients. Dofetilide has also not been shown to prolong life significantly by suppressing malignant ventricular arrhythmias (DIAMOND-Study). In patients with symptomatic ventricular arrhythmias or aborted sudden death, ICD therapy has been proven to be superior to antiarrhythmic drug therapy in cardiac mortality reduction as a secondary prevention strategy (e.g. AVID, CASH, CIDS). For primary prevention of sudden arrhythmic death in high risk patients, 2 studies (MADIT, MUSST) have already demonstrated favorable results, decreasing mortality by ICD therapy in selected patient populations with partly reduced ventricular function and unsustained but inducible ventricular tachycardias. This topic is, however, undergoing further evaluation by ongoing trials (e.g. MADIT II, SCD-HeFT). From available data, antiarrhythmic drug therapy in high risk patients is not justified on a routine basis, whereas ICD therapy as a secondary and perhaps primary prevention strategy will significantly reduce cardiac mortality in patients with severe heart failure. Sotalol, a class III antiarrhythmic agent, has recently been shown to reduce ICD-shock delivery which indicates that concomitant drug therapy in patients with an ICD device already implanted may be beneficial in terms of reducing ICD discharges due to ventricular and supraventricular tachycardias. In patients with paroxysmal atrial fibrillation and congestive heart failure, restitution of sinus rhythm is the primary therapeutic goal which can be safely achieved by amiodarone and dofetilide (DIAMOND). In the latter, continuous monitoring of the patient is mandatory because of increased risk of torsade de pointes arrhythmias during the first days of drug administration. In patients with chronic atrial fibrillation rate control and anticoagulation with warfarin is the primary therapeutic option, which can be achieved with either drug treatment (Digoxin, betablockers, amiodarone) or by His bundle ablation with subsequent pacemaker insertion. PMID- 10859994 TI - [Heart transplantation: indications and results]. AB - Heart transplantation has been established as therapy in patients with terminal heart failure who remain severely symptomatic (NYHA III-IV) despite optimal drug therapy and surgical interventions other than transplantation. In addition to symptoms, various objective criteria are used to determine the patients most suitable for transplantation. Of these, peak oxygen consumption below 12-14 ml/kg/min, no irreversibly elevated pulmonary resistance, and no major concomitant disease are most important. However, conventional therapy of advanced heart failure has considerably improved over the last decade. Thus, heart transplantation may be avoided or at least postponed in many patients. Nevertheless, continuous treatment at a heart failure/heart transplantation clinic in collaboration with the general practitioner is essential in these patients. This may allow to closely monitor these severely ill patients and to select the optimal point of time for transplantation. Prognosis after heart transplantation is relatively good with a 10-year survival of 63%. However, graft coronary artery disease and lymphomas are still unresolved problems which limit the success of heart transplantation. Competent clinical monitoring, aggressive therapy of conventional risk factors and good co-operation of patients and doctors are the basis for a successful outcome. PMID- 10859995 TI - [Pharmacologic and biologic basis of drug allergies]. AB - Drugs are able to activate the immune system, which may generate hypersensitivity states in individuals. This article first deals with the critical processes that are involved in drug sensitizations: what are the specific features of drugs as immunogens; how are drugs recognized as non-self by activated immune cells; what is the spontaneous outcome of an immune response to drugs in individuals; does a genetic predisposition regulate the immune response to drug antigens; how much are biotransformation processes involved in the immunogenicity of drugs? The second part is mostly devoted to the biological investigation of drug sensitizations: what are we looking for and why; are all the available methods equally suitable for routine diagnosis; what are the major methological problems that we have to deal with in investigating patients who have presented with symptoms which are clinically suspected to be of immuno-allergic origin? PMID- 10859996 TI - [Immunoallergic reactions of drug origin: epidemiologic and clinical data]. AB - Allergic and pseudoallergic reactions frequently occur in hospitalized patients and represent up to one-third of adverse drug reactions. Allergic reactions are unpredictable reactions, related to immunologic mechanisms. Pseudoallergic reactions mimic allergic reactions but no drug-specific antibody or T-cell proliferation can be demonstrated. Clinical presentations are numerous and heterogeneous, from a mild urticaria to a dramatic anaphylactic shock and an extensive bullous skin disease. A true diagnosis is rarely set up and the tools for it are lacking. In this review, we will focus on some epidemiological data concerning these reactions, including data on incidence, mortality and cost. PMID- 10859997 TI - [Clinical data on COX-1 and COX-2 inhibitors: what possible alerts in pharmacovigilance?]. AB - Adverse effects of NSAIDs are serious, mainly related to gastrointestinal bleeding, and throughout the world cause about 260,000 hospitalizations and 26,000 deaths a year, but each day at least thirty million patients take NSAIDs. Selective COX-2 inhibitors (i.e. celecoxib, rofecoxib) have demonstrated in clinical trials better gastrointestinal tolerability but their safety in patients with active ulcer, cardiovascular or renal disease has still to be further investigated. When their long-term safety has been established by pharmacovigilance studies they could be prescribed in the at-risk population or for other indications, including pre-term labour, colorectal cancer and Alzheimer's disease, provided they have shown efficacy and safety in controlled trials. PMID- 10859998 TI - [In vitro study of of the effects of cysteinyl leukotrienes on human vascular preparations]. AB - Leukotrienes are 5-lipoxygenase-derived arachidonic acid metabolites. In addition to their bronchoconstrictor effects, leukotrienes are also important modulators of the vascular tone which may exert paradoxical effects. Indeed, depending on the vascular tone (in either the basal or norepinephrine-precontracted state), leukotrienes are capable of inducing either contraction or relaxation. These paradoxical effects of leukotrienes depend on the vascular bed and the species investigated. Since urinary LTE4 excretion is increased in various cardiovascular diseases, including arterial pulmonary hypertension or cardiac ischaemia, the study of the effects of leukotrienes on human vascular preparations is of interest. This article reviews the in vitro evidence linking cysteinyl leukotrienes to the modulation of the vascular tone on human vascular preparations. PMID- 10859999 TI - Culture of vascular cells in tridimensional (3-D) collagen: a methodological review. AB - Experimental models based on the culture of cells within tridimensional (3-D) gels add a 3-D organization to the classical 2-D culture of vascular cells. They allow the study of cell structure in an environment which is more representative of the in vivo situation and the investigation of cellular functions which cannot be studied using the basic 2-D models. This review shows examples of the use of cultures of vascular cells (endothelial cells, smooth muscle cells as well as fibroblasts) in 3-D collagen matrices for the study of cellular functions as diverse as angiogenesis, extracellular matrix reorganization, migration through 3 D collagen gels or phenotype modulation. It also describes recent advances in the in vitro reconstruction of biological blood vessels by bioengineering. A method for the preparation of 3-D collagen gels is described. PMID- 10860000 TI - [Inhibitors of HMG CoA reductase: new modes of action, new indications?]. AB - 3-hydroxy-3-methylglutaryl Coenzyme A reductase inhibitors, or statins, are hypocholesterolaemic agents used in the treatment of hypercholesterolaemia and cardiovascular diseases. Their hypocholesterolaemic action results from the inhibition of the intracellular synthesis of cholesterol via the mevalonate pathway. Recent studies have demonstrated that cholesterol is not the only intracellular target of statins. Indeed, statins have been shown to inhibit protein isoprenylation, a post-translational modification involved in membrane localization and protein activity. By inhibiting isoprenylation of Ras protein, statins induce the inhibition of the Ras-AP1 pathway and the decrease in cell proliferation. Similarly, inhibition of Rho protein insoprenylation leads to the disorganization of the cytoskeleton and the induction of the fibrinolytic system. All these effects need to be confirmed in vivo and in the human, and open new areas for these therapeutics agents. PMID- 10860001 TI - Development of a functional human bronchial model of mucin secretion. AB - In an attempt to study the functional aspects of respiratory mucin secretion and the effects of mediators of inflammation on the release of M1/MUC5AC mucins in airways diseases, an ex vivo human bronchial model of mucin secretion was developed. Anti-M1 mucin monoclonal antibodies raised against the peptidic core of ovarian cyst M1 mucins were used. PAS and Alcian blue stainings of sections of bronchial rings revealed the presence of mucins in epithelial goblet cells as well as in glandular mucous cells. Immunohistochemical labelling of these sections with anti-M1 monoclonal antibodies revealed a preferential localization of M1/MUC5AC mucins in epithelial goblet cells. Functional studies were performed on this bronchial model using various secretagogues (methacholine, leukotrienes D4 and anti-human immunoglobulin E antibodies). No statistical difference of M1/MUC5AC mucin secretion was observed after a one-hour stimulation of bronchial rings with these agents. The development of an ex vivo functional human bronchial model of mucin secretion and the use of specific anti-M1 antibodies are essential tools in studying the regulation of the M1/MUC5AC mucin release from human airways. PMID- 10860002 TI - [Safety pharmacology: central nervous system]. AB - Some of the results formerly included in the dossiers under the heading of 'General Pharmacology: Central Nervous System' were intended to document most of the information now covered by the CNS Safety Pharmacology. This paper allows to point out some general experimental conditions relating to the choice of (1) suitable animal species whose use should be advocated, (2) the route by which the effects of the drugs should be studied and (3) the controls of the experimental context (surroundings, temperature, schedules of the trials, interference with food-intake episodes, influence of noises and smells, expertise of the experimental staff) likely to optimize the predictive value of the trials. Besides the core of investigations recommended by most of the regulatory authorities (changes in gross behaviour, motor coordination, locomotor activity, interaction with hypnotics, convulsant potential), the need to include or not some investigations such as EEG studies and the prediction of adverse events, such as anxiolytic and amnesia potentials, is considered. The place, within the scope of CNS Safety Pharmacology, of studies intended to document the potential for drug abuse and dependence is mentioned and some information given about the methods generally used. PMID- 10860003 TI - Basic safety pharmacology: the cardiovascular system. AB - The cardiovascular system is one of the primary vital functions which have to be examined during safety pharmacology studies. Cardiovascular system functioning is maintained by cardiac electrical activity and by pump-muscle function which contribute to haemodynamic efficacy. The aim of cardiovascular safety pharmacology is to evaluate the effects of test substances on the most pertinent components of this system, in order to detect potentially undesirable effects, before engaging in clinical trials. In the basic programme, a detailed haemodynamic evaluation is carried out in the anaesthetized dog. It is completed by cardiac and/or cellular electrophysiology investigations in order to assess the arrhythmogenic risk. The basic programme can be preceded by rapid and simple testing procedures, during the early drug discovery stage. It should be completed, if necessary, by specific supplementary studies, depending on the data obtained during the early clinical trials. The following article describes and presents an analytic strategy aimed at problems of cardiovascular risk. PMID- 10860004 TI - [The respiratory system and safety pharmacology]. AB - Many drugs affect respiration in man. The changes in respiratory function following their administration are due to a direct effect on the respiratory system or are the consequence of a central, metabolic (alteration of the acid base balance) or vascular effect (pulmonary hypertension). Regulatory documents (CPMP, FDA and MHW drafts) are in agreement in considering the respiratory system as a vital function to explore during safety pharmacology studies. On the basis of these recommendations, the first studies to be performed should be on conscious unrestrained animals, in general the drug being administered as a single dose. The effects on the respiratory function are best studied by plethysmography in the guinea-pig or rat. The measurement of ventilatory parameters--respiratory rate, tidal volume, inspiratory time, expiratory time, peak inspiratory flow, peak exploratory flow and resistance--allow the differentiation between drugs affecting respiratory control and those altering lung mechanical properties. The pulmonary hypertension risk could be evaluated in the dog during haemodynamic studies. Finally, the study of the effects on blood gases should always be carried out in conscious animals, usually in the dog. For drugs belonging to pharmacological classes presenting a high respiratory risk, complementary studies should be considered. The extrapolation from healthy persons to ill patients (especially chronic respiratory insufficiency patients) of the incurred risk is often difficult. For this reason, it is very useful to study the effect of such drugs on the respiratory function of pathological animals. PMID- 10860005 TI - [Safety pharmacology from the regulatory veiwpoint: past, present and future]. AB - Safety pharmacology, a discipline unrecognized until the last few years, is now the subject of Japanese, American, European and ICH draft guidelines. The aim of these guidelines, either of a general nature or targeted at one particular organ, is to define the necessary pharmacology studies as well as the conditions in which these studies must be performed to ensure the safe use of new molecules when they are administered to man. This presentation attempts to give a comparative analysis of these documents and to place them in the context of future international harmonization. PMID- 10860006 TI - Renal safety pharmacology: value of sensitized experimental models. AB - The kidneys are an important target for toxic effects of drug candidates. It is mandatory to select accurate, clinically relevant parameters in order to be in a position to detect putative nephrotoxic effects during the safety pharmacology programme. The glomerular filtration rate appears to be of major interest since it is associated with the definition of acute renal failure. Measurement of the renal blood flow, proteinuria, enzymuria, fractional excretion of sodium, etc. are also highly useful to detect any possible renal impact of a new compound. Although the rat is, by far, the most widely used animal species, there are no specific (clinically relevant) reasons to choose it. Various parameters may vary according to the species, sex, strain, age, etc. Since in most cases acute renal failure occurs following administration of drugs in patients with pre-existing risk factors, it is suggested that sensitized animal models be validated and used (salt depletion, dehydration, co-administration of pharmacologic agents, etc.). PMID- 10860007 TI - [Telemetry: the importance of surgical aspects]. AB - Telemetry allows the monitoring of multiple parameters, such as blood pressure, temperature, motor activity and ECG, without any stressful handling or restraint. The invasive phase (i.e. the surgical implantation of a transmitter), which takes place before the start of the study, is of critical importance. Good surgical techniques as described below are essential. All risks of trauma or infection should be avoided, particularly if the animals are to be used in long-term studies. The utility of telemetry may be complemented with other atraumatic techniques, for instance the use of indwelling i.v. infusion catheters allowing long-term studies without any extrinsic stress resulting from the dosing procedure or routine examinations. PMID- 10860008 TI - The preclinical assessment of the risk for QT interval prolongation. AB - Some drugs have been reported to induce severe ventricular arrhythmias, including torsades de pointes, and have been responsible in some cases for sudden death of patients. Although the mechanisms of these arrhythmias are not well understood, they are often, but not always, associated with QT interval prolongation. Regulatory authorities (CPMP in Europe) have recently pointed out the necessity to assess most carefully the potential, especially of non-cardiovascular drugs, for QT interval prolongation. Different methodological approaches are presented in this paper and experimental protocols are suggested; limitations and advantages of the presently available in vitro and in vivo models are discussed. It appears that both in vitro and in vivo approaches are complementary. In particular it is pointed out that only the in vitro models using isolated cardiac tissues (Purkinje fibres or papillary muscles) enable assessment of the drug properties under low cardiac rhythm conditions. This model allows us to mimic pathological situations of long QT interval (such as acquired or congenital long QT syndrome) in which most of the major clinical problems are encountered. Finally, a strategy for the preclinical assessment of the potential of a molecule for QT interval prolongation is presented. PMID- 10860009 TI - [Pharmacoepidemiology and pharmacoeconomics]. PMID- 10860010 TI - [Pharmacoepidemiology: definitions, problems, methodology]. AB - Pharmacoepidemiology aims to complete the evaluation of drugs made before approval, by providing reliable information concerning effectiveness, safety and utilization of medicines in realistic conditions. The goal may be only descriptive or aetiological. In the latter, the conclusions from observational studies can be jeopardized by systematic errors and cannot achieve the robustness of experimental designs. According to directionality, three main types of studies can be identified: cross-sectional, prospective and retrospective. Prospective and retrospective studies can be based on a single group (descriptive studies) or include a reference group (comparative or aetiologic studies). The interest of prospective studies is reduced when (1) the incidence of the considered event becomes low or (2) one intends to assess the effects of various causal factors. Retrospective studies are approaching their limits when (1) the prevalence of the exposure is low in the source-population or (2) several events or outcomes are concerned. PMID- 10860011 TI - [Pharmacoeconomics as a tool for the synthesis of available information on drugs]. AB - Data on refundable pharmaceutical drugs are numerous and of various kinds. The clinical approach and the economic approach could be used together. This new approach allows for not only the synthesis but also the structuring of all the available information. The new information obtained is involved in decision trees. PMID- 10860012 TI - [Use of computerized data in pharmacoepidemiology]. AB - Positive and negative effects of long-term therapy are sometimes difficult to highlight, for instance in gerontology or in respiratory medicine, and the use of computerized data may be a good alternative. Data collection is made of three steps. First, a study population is identified from disease criteria. All treatments used during a given study period are then extracted from computerized files; these data may be linked, when needed, to detailed patient surveys. Finally, all outcomes of the study period are retrieved, possibly using validated proxies. This method also allows for assessment of the quality of prescribing and the burden of disease. Asthma studies are used as examples. PMID- 10860013 TI - [Epidemiology and economics of chemotherapy combinations in the treatment of advanced breast cancer]. AB - 125 oncologists, taken at random from a French professional list, followed up a cohort of 836 patients suffering from advanced breast cancer and treated by chemotherapy. The following data were collected prospectively: socio-demographics of patients and practitioners, disease history, characteristics of one chemotherapy cycle (type, utility and cost). A total of 89 chemotherapy combinations were described during the first-line treatment, independently of doses, eight combinations representing 70 per cent of all the prescriptions. No statistically significant differences were found when comparing the utility of the most often used combinations. Chemotherapy was the main cost driver of administration, rising to 90 per cent of the total cost when using taxans, followed by CSF. The cost price of taxan administration was twice to three times that of FEC or vinorelbine associations. These differences were poorly taken into account by the French budget allocation system as based on Diagnosis Related Groups. PMID- 10860014 TI - Excess costs related to non-steroidal anti-inflammatory drug utilization in general practice. AB - Data concerning the reasons for consultation and the use of NSAIDs were collected in 4643 patients, seen by 126 GPs over 2 days' consultation. In all, 11.6 per cent took NSAIDs. They were older (49 vs. 46 years, p = 0.02), took more drugs (3 vs. 2.5, p < 0.01), and more had ADRs (8 vs 2 per cent) than non-users, even after correction for age, sex and number of drugs taken. Some 33 per cent of NSAID users also took adjuvant medication for the prevention of gastric injury (including with COX-2 inhibitors meloxicam, nimesulide). Estimated excess costs associated with NSAID use were high, related to excess consultations (GP or specialist, for ADRs, approx. 5-8 million Euros per year in France) and to use of preventive medication (100 million Euros per year at least). PMID- 10860015 TI - [Cost of hospitalizations for adverse drug effects]. AB - The French network of Regional Pharmacovigilance Centres evaluated in 1997 the prevalence of adverse effects of drugs (AED). In 1998, and again with the support of the French Drug Agency, in collaboration with the company CEMKA for economic evaluations, the incidence of AED-related hospitalizations in the medical department of French public hospitals was studied. The evaluation was performed over 14 consecutive days in 62 hospital departments, which were selected randomly. The total number of 3137 patients were hospitalized for a mean duration of 9 days and they were using a mean number of six different drugs. Taking into account the number of about 4 million patients admitted per year in the hospitals represented, it was estimated that the total number of AED-related hospitalizations amounts to about 130,000 annually (CI95%: 100,916-156,620). Using established cost calculations for hospitalized days (AP-HP, results of 1996) the mean cost for an AED-related hospitalization was estimated to be about FF16,000. PMID- 10860016 TI - [National observation of prescriptions and consumption and pharmacoepidemiology]. PMID- 10860017 TI - Endogenous benzodiazepines. AB - The existence of endogenous benzodiazepines such as diazepam and nordiazepam has been provided in human blood and brains as well as in medicinal plants and foods. It must be stressed, however, that in plasma and brain tissue there are also other benzodiazepine-like compounds termed 'endozepines' which are not halogenated. A synthetic pathway for the production of benzodiazepine-like compounds and endozepines has not yet been found, hence it may be surmised that these compounds could be of exogenous source. Changes in the level of endogenous circulating benzodiazepines due to food or drug ingestion could be responsible for pathological conditions. Clinical experiments were designed in order to study the levels of the endogenous benzodiazepines in vegetables and in the blood of control subjects and of cirrhotic patients. These patients accumulate benzodiazepines because of decreased liver metabolization capacity and impaired renal secretion, reaching plasma concentrations similar to those recorded in commercial benzodiazepine consumers. PMID- 10860018 TI - Clinical methodology for testing of anxiolytic drugs. AB - Diagnostic criteria and classification are changing. It is no longer acceptable to include patients with a general diagnosis of any anxiety, or neurotic anxiety. Regardless of the reference system used, DSM IV or ICD 10, anxiety disorders are now detailed in separate entities. General anxiety disorder, GAD, which is pivotal for the evaluation of new products, can only be claimed after the elimination of all the others, and is relatively rare. The inclusion of such outpatients is further complicated, as comorbidity is frequently associated with GAD--alcoholism, major depression, dysthymia, personality disorders, somatic disease likely to interfere with patient evaluation--and leads to exclusions, and also because the requested duration for the syndrome, prior to inclusion, is six months, which means six months without psychotropic drugs, including excessive alcohol consumption. As to patient evaluation, the reference scale remains the HAM-A. It should show a score above 20 at baseline. It has been designed to assess the level of anxiety of patients presenting with the diagnosis of anxiety, but not the diagnosis of GAD, and, clearly, in relation to the expected results obtained with BZD, which are still the standard reference drugs. The same is true for the other investigator scales and self-rating scales. Moreover, the criteria defining clinical improvement are still discussed. More generally, clinical testing in comparison with placebo and reference drugs is particularly important for anxiolytic drugs. The optimal dose range should be investigated in phase I, evidence of sedative or disinhibiting effects, and in phase II, defining the minimal active dose. Longer duration of treatment should be scrutinized in phase III, in order to check on long-term efficacy, recurrences and relapses. The effects of drug withdrawal should also be studied: withdrawal syndrome, rebound, recurrence, dependence. It currently looks difficult to market new anxiolytic drugs, and clinical research mainly provides an extension of the indications for antidepressant drugs in anxiety. PMID- 10860019 TI - [Pharmacology of recombinant cytokines]. AB - The explosive growth in cytokines has been followed by many attempts to bring them into clinical use. Immediate applications are already recognized in cancer and infectious diseases. Future applications are foreseeable in inflammatory and auto-immune diseases. The use of accurate and sensitive methods for cytokine measurements in body fluids is an absolute prerequisite to define the pharmacological effect of parenterally administered recombinant cytokines. Enzyme linked immunosorbent assay (ELISA) has become the most convenient method but there is an urgent need for a real standardization of this technique. Moreover, ELISA may be susceptible to cross-reactivity due to the high percentage of amino acid homology between the various cytokines. The pharmacokinetic profile of recombinant cytokines may be influenced by endogenous production, receptor binding effect, receptor antagonists and soluble receptors. Cytokines elicit an immunogenic response and anticytokine antibodies should be monitored. Serum half life of elimination is about 4 h after subcutaneous administration. In contrast with conventional drugs, pharmacokinetic data do not provide useful information for the design of a clinical protocol, and the rational choice of the unit dose and dosing schedule should be based on biological considerations. In vitro studies on the duration of receptor occupancy required for effector augmentation provide one of the bases for the choice of therapeutic protocol. Recombinant cytokines share biological activities and synergize with or antagonize one another so that it is difficult to evaluate their effects in clinical studies. Thus, pharmacological results do not always correlate with therapeutic effect. There is no direct relationship between dose and activity. One must determine the optimum biological dose (OBD), which is the minimal dose resulting in a significant augmentation of effector cell activity correlating with the therapeutic response. Surrogate markers may help to assess the clinical response such as 2',5'-oligo(A) synthetase or neopterin following interferon administration. Cytokines' adverse effects are difficult to foresee in the human because studies in rodents and dogs cannot fully predict them because of their species specificity. New relevant animal models are needed such as transgenic animals and parallel animal models. Pro-inflammatory cytokines inhibit cytochrome P-450 and have the potential to cause drug-cytokine interactions. PMID- 10860020 TI - Glucocorticoid hormones in the regulation of cell death. AB - The immune T-cell compartment maintains the capability to respond to a wide variety of antigens (Ag). This whole process is regulated by lymphocyte apoptosis (programmed cell death, PCD) and involves the coordinated expression of a great number of genes including those coding for cytokines and their receptors, such as for example IL-2/IL-2R and the Fas/FasL systems and those coding for transcription factors, including the NF-kB complex, involved in T-cell activation and apoptosis in that they simultaneously activate cell suicide and an anti-death programme. This binary effect, PCD activation and inhibition, is due on one hand to GCH-induced activation of the caspases cascade and on the other to the induction of expression of a new gene that we have named GILZ. In fact, GILZ over expression in transfected cells inhibits the sequential increase of NF-kB/DNA binding activity, IL-2 production and IL-2R expression, and transcription of the Fas/FasL complex that follows TCR triggering and plays an important role in the control of T-lymphocyte apoptosis. These results indicate a new mechanism responsible for the GCH-mediated inhibition of T-cell death and activation that could contribute to anti-inflammatory and immunosuppressive efficacy. PMID- 10860021 TI - Interleukin-1 in the central nervous system: from physiology to pathology. AB - A classification on the basis of time-course effect is proposed to describe the pleiotropic actions of interleukin-1 (IL-1) on the central nervous system (CNS); two main time-frames, minutes-to-days and days-to-years, are distinguished. The former includes the central aspects of acute-phase response with fever, altered food and water intake, sleepiness, sickness behaviour and neuroendocrine changes. Apart from stress response triggered by immune-inflammatory stimuli, the concept that IL-1 mediates other types of stress is also reviewed, showing that the cytokine may have a role in mediating hypothalamic responses to restrain stress and nociceptive stimuli. The days-to-years time-frame includes several CNS disorders accompanied by inappropriate and/or sustainedly elevated IL-beta production: ischaemia, Alzheimer's disease, HIV-related dementia and experimental allergic encephalomyelitis-multiple sclerosis. In all cases, IL-beta is not envisioned as an aetiological factor, but it contributes significantly to the maintenance of disease state. Current and perspective therapeutic approaches involving the modulation of IL-beta production and effects are briefly discussed. PMID- 10860022 TI - Clinical pharmacology of immunosuppressive drugs: year 2000--time for alternatives. AB - Perspectives in immunosuppressive drug therapy have changed rapidly in the past few years with the appearance on the market of several new entities. Used for organ transplantation, bone-marrow transplantation and more recently in some auto immune diseases, the usual classical scheme consisting of corticoids, azathioprine (1970) and cyclosporin (1980), with or without an induction period with antilymphocyte antibodies, has varied little except for the monoclonal antibody OKT3. The considerable evolution due to the introduction of cyclosporin almost twenty years ago has reached its limits. The new perspectives offer two aspects: (1) on one hand, the specificity of each compound: tacrolimus, Prograf, Fujisawa; mycophenolate mofetyl (MMF), CellCept, Roche; cyclosporin, Neoral, Novartis; basiliximab, Simulect, Novartis; and dacliximab, Zenapax, Roche; monoclonal humanized antibodies, rapamycine or sirolimus, Rapamune, Wyeth; or its derived form RAD Novartis; (2) on the other hand, all these products represent alternatives to the present scheme in a field where coprescription is the rule. These alternatives encounter one main difficulty: the evaluation and the organization of the different possible combinations have to be done within a small series of patients. It is essential to note that the market authorizations have been given on the basis of a precise scheme of dosage regimen from the pivotal studies and that extrapolation from these conditions, in particular choice of the doses, has required thorough reflection. These recent developments need optimization between an improvement of immunosuppression and a higher risk for infection and malignancy. PMID- 10860023 TI - Cardiac K+ channels and drug-acquired long QT syndrome. AB - The hallmark of long QT syndromes (LQTS) is an abnormal ventricular repolarization characterized by a prolonged QT interval on the electrocardiogram and a propensity to the occurrence of syncopes resulting from polymorphic ventricular tachycardia, called torsades de pointes. They may degenerate to ventricular fibrillation, possibly causing sudden death. Congenital LQTS, which implicates at least six chromosomal loci, LQT1 to LQT6, three of them corresponding to mutations concerning the coding of K+ channel proteins, give useful information about the mechanism underlying the arrhythmia. One of the potassium channel genes implicated in congenital LQTS is HERG, which encodes the IKr current channel protein. This current has provided a relevant insight into the occurrence of drug-acquired LQTS, since all drugs associated with torsades, such as erythromycin, terfenadine, haloperidol, or cisapride, also block IKr. PMID- 10860024 TI - Effects of calcium channel blockers on cloned cardiac K+ channels IKr and IKs. AB - Cloned HERG and KvLQT1-IsK K+ channels have been expressed in mammalian cells and assayed as a target for calcium channel blockers. These channels generate the rapid and slow components of the cardiac delayed rectifier K+ current, and mutations can affect them that lead to long QT syndromes. HERG is blocked by bepridil (EC50 = 0.55 microM), verapamil (EC50 = 0.83 microM) and mibefradil (EC50 = 1.43 microM), whereas nitrendipine and diltiazem have negligible effects. Steady-state activation and inactivation parameters are shifted to more negative values in the presence of the blockers. Similarly, KvLQT1-IsK is inhibited by bepridil (EC50 = 10.0 microM) and mibefradil (EC50 = 11.8 microM), whilst being insensitive to nitrendipine, diltiazem or verapamil. This work may help to understand the mechanisms of action of verapamil in certain ventricular tachycardias as well as some of the deleterious adverse cardiac events associated with bepridil and mibefradil. PMID- 10860025 TI - [Use of the CD-ROM "Tox-Didact" for teaching of toxicology and pharmacology]. AB - Tox-Didact is a multimedia teaching software package for initial training, continuing training and self-learning of toxicology and pharmacology. This software covers a large part of toxicology in its acute and chronic pathology using several approaches: drugs (salicylics, paracetamol, lithium ...); toxins (lead, methanol, carbon monoxide ...); drug addiction and doping (cocaine, heroin, LSD, amphetamines ...); systemic targets (kidneys, skin, liver ...). Tox Didact is currently composed of 39 modules in validation, each tackling the diagnosis, biological surveillance, treatment, prevention and documentation of a real clinical case. Each module is organized around four types of questions, requiring a choice either (drug, symptom, formula ...) or an open response. Each validated answer is analysed by software which then comments on or corrects it. The essential points which characterize this software are: its multidisciplinarity (toxicology, pharmacology, semiology ...), its reliability (validated by experts), its simplicity of use. It is modular and offers an interactive teaching approach. The objective is to create a portable multimedia tool operational with all computer systems (IBM PC, Macintosh). This program is sustained by the Region des Pays de la Loire and by the Multimedia Resource Office of the French Ministry of National Education. PMID- 10860026 TI - [Adverse effects of locally applied drugs]. AB - Topical drugs, such as non-steroidal anti-inflammatory drugs (NSAIDs), are available without prescription and are widely used for moderate, acute and chronic painful conditions. However, the different characteristics of their adverse reactions remain largely unknown. The present work was undertaken in order to quantify ADRs reported to our centre with topical drugs during a period of 5 years. The retrospective study was performed using the spontaneous reports to the Midi-Pyrenees Pharmacovigilance Centre from 1993 to 1997. The following routes of administration were selected: cutaneous, ocular, pulmonary, vaginal, intra-articular, auricular, nasal, buccal and pharyngeal. One hundred and twenty eight notifications were found in the database. Thirty-eight cases (29.7 per cent) were between 60 and 75 years old, 25 cases (19.5 per cent) between 45 and 60 years old and 22 cases (17.2 per cent) between 30 and 45 years old. The most frequently reported ADRs were cutaneous (n = 71, 55.5 per cent) cardiovascular (n = 16, 12.5 per cent), ocular (n = 13, 10.2 per cent), neurological (n = 12, 9.4 per cent) and oro-pharyngeal (n = 7, 5.4 per cent). Systemic ADRs (45 cases, 35 per cent) occurred mainly with aerosols (n = 10) and patches (n = 6). Twenty seven reports (21 per cent) were classified as 'serious' including one death (misuse). The main pharmacotherapeutic agents involved were rheumatological [(n = 29) including 20 NSAIDs], dermatological (n = 27), ophthalmological (n = 16), pneumological (n = 15) and cardiovascular (n = 15) drugs. In 21 cases the rechallenge was positive. In eight cases (6.3 per cent), ADRs occurred after self medication. This study underlines the importance of monitoring ADRs of topical drugs. The seriousness of such ADRs and the occurrence of systemic effects with such drugs seems rather frequent (as shown by topical NSAIDs-induced photosensitization). PMID- 10860027 TI - Warning! One buflomedil may hide another one! AB - A 75-year-old woman experienced fever and convulsions. She was treated for diabetes mellitus, angina pectoris and also for arteritis with Buflomedil Merck (3 tab/d). Further investigations failed to find any aetiology. Buflomedil dosage was elevated to 6.3 mg/l (N = 4-4.5 mg/l). The drug was discontinued and there was no recurrence of symptoms. There was no evidence of error in dosage or interaction. A failure of the generic product was suspected. Only a pharmacist solved the problem. Fonzylane (buflomedil) had recently been switched to Buflomedil Merck. The patient misunderstood the change and took both drugs! Our purpose is not to report a known effect but to emphasize the importance of extending the information given to the patient and the risk of misuse of the generic product. PMID- 10860028 TI - [Lyell's syndrome and phenobarbital: two cases]. PMID- 10860029 TI - Latex allergy: a potential liability issue. PMID- 10860030 TI - Establishing a baseline: the medical health history and other patient record considerations. PMID- 10860031 TI - Clinical evaluation of the use of calcium sulfate in regenerative periodontal surgery for the treatment of Class III furcation involvement. AB - The purpose of this study was clinical evaluation of the effectiveness of a composite graft, consisting of demineralized freeze-dried bone and doxycycline hyclate (4:1 by vol.) in combination with a resorbable calcium sulfate barrier in the treatment of Class III furcation involvement. A total of 24 sites in seven patients having adult periodontitis with at least two mandibular molars with class III furcation involvement were randomly allocated into two equal groups. The experimental sites received the composite graft in the furcation area covered by the barrier buccally and lingually. The control sites were treated by surgical debridement only. Baseline preoperative data, including probing depth, clinical attachment level, gingival recession and radiographic bone level, were recorded. During surgery, direct linear and volumetric measurements of the defects were taken. At 12 months postoperatively, all sites were surgically re-entered and all measurements were again recorded. Student's t-test and analysis of variance of the data showed: (1) The experimental sites had a significant (P < .05) gain in clinical attachment while the control sites did not (1.44 + 0.33 mm vs. 0.11 + 0.35 mm); (2) the control sites showed greater (P < .05) gingival recession than the experimental sites (0.93 + 0.43 vs. 0.31 + 0.26); (3) when compared to control sites, the experimental group showed significantly (P < .05) greater vertical defect fill (2.24 + 0.79 mm vs. 0.45 + 0.82 mm) and volumetric fill (37.2% vs. 7.5%). In conclusion, the combined composite graft and resorbable barrier utilized in this study may have a favorable effect on the treatment outcome of mandibular class III furcations 12 months postoperatively when compared to surgical debridement only. PMID- 10860032 TI - Reimplantation of an avulsed tooth after prolonged storage. AB - This report demonstrates that if an intact avulsed tooth is retrieved, stored carefully and treated, it can be re-implanted successfully, even after more than 42 hours outside the alveolus. The tooth can regain its position as a stable member of the arch, and its osseous and gingival complex can be restored. PMID- 10860033 TI - On predicting prognosis for single teeth. Case reports. AB - Two cases are presented where teeth initially deemed hopeless and scheduled for extraction were, at the patient's request, treated. The so-called hopeless teeth responded well to regenerative periodontal therapy. This supports findings of previous studies that only on healthy teeth can treatment outcome be predicted with confidence. PMID- 10860034 TI - Evaluation of adjunctive tetracycline fiber therapy with scaling and root planing: short-term clinical results. AB - The purpose of this single-blind, randomized study was to evaluate the clinical efficacy of slow-release tetracycline fibers as an adjunct to initial periodontal therapy in moderate to advanced periodontitis patients who had no periodontal treatment before. A group of 17 patients with at least two sites in each quadrant with probing pocket depths > 5 mm that bled on probing were included in the study. After scaling and root planing, each selected site was randomly assigned to one of two treatment groups: tetracycline fiber therapy and control group. Plaque index, gingival index, bleeding on probing, probing pocket depth, and clinical attachment level were measured at baseline and at 1, 3, and 7 weeks. The change from the baseline of each measurement was studied using analysis of variance with subject and treatment effects and a baseline covariate. Results of this investigation demonstrated that tetracycline fiber treatment as adjunct to scaling and root planing is effective in regard to probing pocket depth reduction and bleeding on probing. On the other hand, fiber application seems to have no beneficial effect on gain of clinical attachment level within limits of this study. PMID- 10860035 TI - Oral giant pyogenic granulomas associated with facial skin hemangiomas (Sturge Weber syndrome). AB - This is a case report of two patients, aged 26 and 22, who suffered from congenital hemangioma on their faces and pronounced gingival overgrowth localized parallel to extraoral lesions. Prior to surgical intervention the hygienic conditions were improved in several sessions by means of professional preventive treatment and oral hygiene instructions. Histologic examination of both cases revealed a highly vascularized pattern of pyogenic granuloma. One of the cases was associated with a pregnancy. These patients can be classified as Sturge-Weber syndrome. Postsurgical treatment consisted of efficient plaque control and adequate oral prophylaxis sessions every 3 months. The large gingival overgrowth was not observed to recur in 2 and 4 years, respectively, of follow-up. PMID- 10860036 TI - Dental millennium series, Part 1. Dentistry in 1000 AD. PMID- 10860037 TI - Dental millennium series (continued), Part 2. Barbitonsores and dentatores. PMID- 10860038 TI - Psychological models and strategies to guide dentist-patient interaction. PMID- 10860039 TI - External bacterial contamination of local anaesthetic cartridges. AB - In this study external bacterial contamination of local anaesthetic cartridges from newly opened and open containers was examined. Colonies of mainly Gram positive cocci and Gram-positive rods were grown from cartridges from both the freshly opened and open containers. However, the total number of colonies grown from open containers was significantly higher than that from freshly opened ones. It was concluded that where local anaesthetic cartridges are bulk-packed in containers, strict infection control measures are to be instituted in clinical practice. We suggest that these include keeping containers tightly capped, removing cartridges only when needed, using forceps to handle cartridges and swabbing cartridges with alcohol prior to loading into syringes. PMID- 10860040 TI - Tissue morphogenesis and tissue engineering by bone morphogenetic proteins. PMID- 10860041 TI - The gingival prosthesis--a literature review. PMID- 10860042 TI - A comparison of service profiles in alternative dental care delivery systems. AB - The financial crisis within the health care sector has resulted in the search by medical benefit funds of worker organisations for cost-effective alternatives to the private health care sector. This study compares the profile of services rendered to members of a union medical benefit fund by an 'in house' dental clinic and private dental practitioners. A total of 2,858 union clinic claims and 10,811 private practitioner accounts were selected. Each treatment code was categorised into either preventive, restorative, extraction or denture services. For the union clinic and private dental practitioners a total of 3,021 and 15,853 treatment codes respectively were included in the analysis. Significant differences were found in the profile of services rendered by the union clinic and private dental practitioners. The union clinic provided predominantly preventive and denture-related treatment compared with private practitioners who provided mainly preventive and extraction services. Members and their dependants are far more likely to have dentures made through the dental clinic (the odds of having dentures made at the clinic were 4.67 times the odds of having dentures made through private practitioners). On the other hand, members were more likely to receive all the other services (restorative, preventive and extractions) from private practitioners. However, the nature of the study design and the findings do not allow conclusions to be drawn regarding whether these differences were due to demand or supplier influences. PMID- 10860043 TI - Industrial dental erosion: a cross-sectional, comparative study. AB - Occupational exposure to sulphuric acid mist (H2SO4) is a health hazard. The threshold limit value-time weighted average (TLV-TWA) of exposure to H2SO4 recommended by the American Conference of Governmental Industrial Hygienists (ACGIH, 1994-1996) is 1 mg/m3. This single-blind study conducted in an electro winning facility in South Africa, compared dental erosion of anterior and premolar teeth of male workers exposed daily to H2SO4 in an exposed group (H2SO4 range: 0.3 mg/m3-1 mg/m3) and an unexposed group (H2SO4 range: 0.1 mg/m3-0.3 mg/m3). The exposed group comprised all workers at the facility exposed to the aforesaid range of H2SO4 (N = 103). A total of 102 unexposed subjects similar in composition with respect to age and length of service were randomly selected from the rest of the 700 workers at the facility. A questionnaire was administered to seek information on possible worker habits associated with dental erosion and to determine perceptions of oral function. Clinical examinations assessed prevalence and severity of dental erosion. The mean age of subjects was 31.4 years and mean length of service 4.2 years. In the exposed group 48% complained of pain and sensitivity on their teeth compared with the 31% of unexposed persons (P = 0.020). Dental erosion was present in exposed (96%) and unexposed (75%) subjects. Exposed subjects were more likely to develop erosion than unexposed subjects, the odds ratio being 5.531 within the confidence limits 2.167 < OR < 14.117. There was a significant difference in the severity of tooth surface loss between exposed and unexposed groups (P = 0.001). Dental erosion was most severe in the anterior teeth and occurred mostly on the labial and incisal surfaces. PMID- 10860044 TI - Attitudes to water fluoridation in South Africa 1998. Part I. Analysis by age, sex, population and province. AB - The successful implementation of water fluoridation (WF) depends upon support from the public. A survey was conducted in 1998 by the Human Sciences Research Council to assess attitudes of the South African population towards WF. A representative sample of 2,220 persons over the age of 18 years was interviewed. Questions relating to current knowledge, sources of information, the purpose of water fluoridation and its desirability as a public health measure were put to respondents. Only 25% of the population had heard or read of WF. The major sources of information were the electronic and printed media (40% and 27% respectively), while 2.5% heard about it from dentists. Just over a third of the sample (35.5%) identified the purpose of WF as protecting teeth from decay; 28% said it was to purify water and 28.5% were uncertain of, or did not know its purpose. Nearly two-thirds (61.9%) indicated that fluoride should be added to drinking water if it can reduce tooth decay, while 9% were not in favour and 29% were uncertain. The results suggest that most people do not know what WF is or does, though most were in agreement that it should be implemented to reduce tooth decay. PMID- 10860045 TI - The child dental patient. Part II: An approach to the management of fearful children. AB - The first part of this review, published last month, dealt with understanding the nature and prevalence of dental fears among children. This article is concerned with investigating ways in which these fears could best be managed. PMID- 10860046 TI - [An examination program for prospective dental students]. AB - The aim of this study was to develop a selection programme for prospective dental students at the University of Pretoria. A study of relevant literature was undertaken, and experts at universities in South Africa, Britain and Belgium were interviewed, in order to research and analyse existing selection procedures with a view to developing a concept selection programme. By means of a job analysis questionnaire relevant job requirements were ascertained, which were utilised as criteria for a concept selection programme. PMID- 10860047 TI - Evaluating the efficacy of curing lights. AB - The use of light curing units has increased tremendously over the past few years with the introduction of photoactivated composite restorative resins. The advantages of light-cured composites are well documented especially the ease of placement of restorations as a result of extended working time and control of setting. Many of the advantages of these composites are dependent on the adequate polymerisation and therefore the source of the visible light, especially the wavelength and the intensity of the light source. The aims of this study were (i) to determine the intensity output of curing lights in state clinics, private dental practices and dental clinics run by trade unions using a radiometer and (ii) to relate their output to various factors that may affect the intensity of the light produced by each unit. The light intensity of each light curing unit was measured using the Efos Cure Rite radiometer (Cure Rite, Efos model #8000; Efos Inc, Mississauga, Canada). Mean readings ranged from a high of 448 to a low of 22 mW/cm2 for curing lights at the time of evaluation. Nineteen (54.3%) of the light curing units were functioning at optimal intensities, followed by 10 (28.6%) functioning at levels between 150 and 300 mW/cm2 and 6 (17.1%) functioning at levels far below that required to achieve adequate photocuring. No statistically significant differences were found when comparing intensity readings with age, last maintenance service and bulb replacement (P > 0.05). All of the clinicians interviewed (100%) expressed subjective satisfaction with the performance of the light curing units, even though 45.7% functioned below optimal intensities (P > 0.05). Not one of the clinicians expressed dissatisfaction with the performance of any of the light curing units. The results of this study showed that the light intensities f light curing units used within private dental practices, state and trade union dental clinics were inadequate for optimum curing. PMID- 10860048 TI - A feeding adaptation by an infant with a cleft palate. AB - One of the primary problems in the management and care of infants born with cleft palates is that of achieving an adequate seal of the cleft to allow proper swallowing to take place. This article describes an interesting case in which a 14-month-old baby, who had received no surgical or prosthodontic treatment for her cleft palate, had developed her own 'obturation' mechanism to enable her to swallow efficiently. PMID- 10860049 TI - Oral lipoid proteinosis. AB - This literature review of oral lipoid proteinosis and a case report of an infant with this condition describe an uncommon condition inherited as an autosomal recessive trait, but one which is more likely to occur in this country than elsewhere. The article discusses other lesions which could be mistaken for oral lipoid proteinosis, and offers some pertinent aspects of its diagnosis and management. PMID- 10860050 TI - Assessment of periodontal status and treatment needs of a disabled population using the CPITN. AB - The Community Periodontal Index of Treatment Needs (CPITN) was used to assess the periodontal status of 213 handicapped persons attending seven institutions in Johannesburg. Fewer than 2% had healthy mouths, 8% had bleeding only, followed by calculus (46%), shallow pockets (40%) and deep pockets (4%). The mean number of sextants with bleeding or higher score was 5.9. Oral hygiene instruction was indicated for 98% and prophylaxis for 90% of the participants. The CPITN was easily used in the disabled population but may overestimate treatment need in view of the current understanding that periodontal disease does not automatically progress from a low CPITN level to the next. A more appropriate measure of treatment need in handicapped persons is required. PMID- 10860051 TI - Gunshot injuries of the orofacial region in Nigerian civilians. AB - The dearth of information from Africa on gunshot injuries to the orofacial region in a civilian population prompted this retrospective survey. Sociodemographic data, manifestations at admission, type of firearm inflicting injuries, mechanism of injury, radiographs, treatment and complications were reviewed. All injuries occurred in the civilian population. The majority of patients (95.5%) were males, involved mainly in recreational hunting expeditions. The Dane gun was mostly used (86.4%) and accidental discharge of the weapon was recorded in 59.1% of the cases, 40.9% of these being accidentally self-inflicted. Overall, the left half of the face received more of the missiles. Complications were seen in 63.6% of the patients and the most common of these was loss of sight. The mortality rate was 22.7%. Late presentation was a frequent occurrence. It is suggested that Nigerian law enforcement agencies devise strategies to monitor the sale, acquisition and use of such firearms. PMID- 10860052 TI - The apically repositioned flap in tooth exposure. AB - Methods of exposing impacted teeth in order to bring them into the line of the arch include gingivectomy, the apically repositioned flap and closed eruption techniques. These procedures aim to facilitate the eruption of the impacted tooth with a minimum of disruption or damage to the tooth itself or adjacent structures. The aim of this paper is to discuss the various surgical methods of exposing impacted teeth and to help to identify where the use of the apically repositioned flap is indicated. Clinical examples are presented and a surgical method for carrying out this procedure recommended. PMID- 10860053 TI - Immediate cantilevered resin-bonded bridgework: a case report. AB - Resin-bonded bridgework (RBB) is now an accepted alternative to conventional bridgework in specific cases. This article reviews the immediate replacement of two anterior teeth with the resin-bonded technique and describes the use of immediate cantilevered RBB in the aesthetic treatment of a patient with class II division II malocclusion on a moderate skeletal II base. Both maxillary lateral incisors were extracted and immediately replaced with cantilevered RBB. This simple and conservative approach produced an instant solution to a difficult aesthetic problem of proclined lateral incisors. The reported RBB continues to function successfully after 15 months. PMID- 10860054 TI - The effect of elastomeric impression materials on the growth of micro-organisms. AB - The influence of elastomeric impression materials on the growth of micro organisms was examined in vitro. Bacillus subtilis was inoculated into broth containing impression materials and incubated at 37 degrees C for 72 hours. Express STD Putty, President Putty and Jet-Light Body, Low and Very High Viscosity Permagum and Provil L stimulated growth whereas Impregum-F and Express Light Body inhibited growth. The influence of Impregum-F and Express Light Body on oral micro-organisms was investigated further. Broth extracts were prepared by soaking these materials in Todd Hewitt broth for either 5 or 10 minutes. Thereafter, the extracts were inoculated with oral strains of Staphylococcus aureus, Streptococcus mutans and Candida albicans, incubated at 37 degrees C for 72 hours and plated on blood and Sabourauds agar to test for the presence of viable micro-organisms. The 10-minute broth extracts killed all the test isolates which suggests that impressions taken with Impregum-F and Express Light Body may not require disinfecting. PMID- 10860055 TI - Full dentures--back to basics. AB - This article is the first one of a series on the basic essentials in full denture construction. It deals with the introductory visit and the importance that it has in the success or otherwise of the clinical procedure. Future articles will discuss the various clinical stages in full denture construction. PMID- 10860056 TI - Functional appliances: mode of action and clinical use. AB - Functional appliances are frequently used in the treatment of class II division 1 malocclusions, either in isolation or, more commonly, before a course of fixed appliance therapy. Extensive speculation and investigation into the precise mode of action of these appliances has recently focused on their growth-restraining effect on the maxilla, their growth-enhancing effect on the mandible and dentoalveolar and soft-tissue effects. The different types of functional appliances are described in this article, with emphasis placed on practitioners adopting a component approach to design. A checklist is provided to aid identification of the more common problems occurring during treatment. PMID- 10860057 TI - The child dental patient. Part I. The nature and prevalence of children's dental fears. AB - For various reasons, fear of dentists and dentistry is not uncommon among children. These fears not only disrupt the performance of clinical procedures, they also tend to undermine the relationship between dentist and child patient. It would seem to make sense then, to address this vexing problem in which two major issues are involved. The first is concerned with the prevalence of dental fears among children and the second deals with their management. For reasons of convenience, clarity and brevity, these issues are dealt with in two separate papers. The first paper (Part I) focuses on acquiring an understanding of the nature and prevalence of dental fears among children and the second (Part II) is concerned with investigating ways in which these fears could best be managed. Parts I and II appear in successive issues of the SADJ. PMID- 10860058 TI - Assessment of patients for orthognathic surgery. AB - Rapid advances in orthognathic surgery now allow the clinician to treat severe dentofacial deformities that were once only manageable by orthodontic camouflage. These cases were often compromised with unacceptable facial esthetics and unstable occlusal results. Over the past 25 years, there have been numerous improvements in technology and the surgical management of dentofacial deformities. These progressions now allow more predictable surgical outcomes, which ensure patient satisfaction. Not all patients are candidates for surgical treatment; therefore, patient assessment and selection remains paramount in the process of diagnosing and treatment planning for this type of irreversible treatment. The inclusion of patients in the decision-making process increases their awareness and acceptance of the final result. The past three decades indicate an increased usage of orthodontic treatment by both children and adults. Patient demographic profiles for severe occlusal and facial characteristics are presented in an effort to understand the epidemiological factors of malocclusion and predict the population's need for this service. PMID- 10860059 TI - Patient-centered outcomes in surgical and orthodontic treatment. AB - Patient-centered health care has two characteristics: it is closely congruent with and responsive to the patient's wants, needs, and preferences, and it considers the psychological, social, cultural, and economic dimensions of the patient in addition to physical findings. The ultimate benefit to the patient has always been a primary concern of clinicians providing coordinated orthodontic and orthognathic surgical treatment. However, in the past 10 years, a much greater emphasis has been placed in both medicine and dentistry on the patient as a coparticipant in decision making and the process of treatment. In addition, it has been realized the success of treatment must be defined not just in terms of the objective findings of clinicians, but also in the context of the patient's perceptions of what was achieved. This article discusses the impact of the new emphasis on patient-centered care on clinical practice and research in orthodontics and orthognathic surgery and provides two examples of how patient centered outcomes can be assessed and used to improve the quality of care in these patients. PMID- 10860060 TI - Surgical versus orthodontic correction for Class II patients: age and severity in treatment planning and treatment outcome. AB - Treatment options for Class II malocclusion include orthognathic surgery. Treatment choices are particularly difficult for young patients because of the uncertainty regarding future growth. Surgical treatment has generally been considered necessary for older patients with more severe Class II problems. The treatment records of more than 500 patients with Class II malocclusion were reviewed. Patients were grouped according to their initial treatment plan (surgery or orthodontics) and treatment outcome (overjet [OJ] reduced to < 4 mm or not). Discriminant function analyses using data from the patient's pretreatment cephalogram were used to determine whether age, in combination with malocclusion severity, could predict the choice of treatment, and whether a simple set of pretreatment variables could predict the success or failure of OJ reduction. The derived equations were tested in a similar group of growing Class II children. Although the data showed clinicians use patient's age in determining treatment choice, age did not seem to be associated with treatment outcome. The majority of the variability that determined the success or failure of OJ reduction was not explained by patient's age or malocclusion severity. These findings suggest other factors, including psychosocial variables, need to be explored if we are to gain a better understanding of why treatments succeed or fail. PMID- 10860061 TI - Bilateral sagittal split osteotomies in an ambulatory care setting. AB - Orthognathic surgery has made many advances since its inception. This, combined with modern day health economics, has necessitated changes in orthognathic surgical practice. The bilateral sagittal split osteotomy is evaluated as an ambulatory surgical procedure. Studies have proven the sagittal split osteotomy can be performed as an ambulatory procedure. Patient costs, as well as other issues that impact on the patient and health care provider, are reviewed. The need for additional outcome data to determine whether patient care or perception of care is affected is discussed. PMID- 10860062 TI - Complex orthodontic problems: the orthognathic patient with temporomandibular disorders. AB - The diagnosis and treatment of temporomandibular disorders (TMD) remain controversial despite considerable research and publication in this area. The relationship of these problems to dental and skeletal malocclusion is equally debatable. Recent studies suggest that although malocclusion may have a role, it is a small one. Accordingly, treatment of TMD with occlusion-altering therapy, such as orthodontics and orthognathic surgery, should be limited to specific situations. This report discusses the management of patients with coexisting TMD and skeletal malocclusion. Current concepts in clinical and radiographic diagnosis are discussed, as well as an overview of noninvasive therapy. A case report is used to illustrate an approach to diagnosis and treatment planning in an individual with active TMD and a skeletal malocclusion requiring orthognathic surgery for correction. PMID- 10860063 TI - How to avoid surgical failures. AB - Problems and failures in orthodontic-orthognathic surgical treatment are frequently discussed among colleagues, but not often written about. This lack of documentation may be attributed to a natural inclination for us to report our successes and hide our failures. More positively, we believe the lack of written material on complications probably relates more to the overwhelming success rate in these procedures than to an inclination to hide failures. In addition, some complications are clearly a result of the orthodontic care before and/or after the surgical procedure. This article concentrates on exploring the many complications in orthognathic surgery that are more a result of the orthodontic phase of care, and how the orthodontist can react to and manage surgical complications to achieve a successful result. The more the orthodontist understands the nature of surgical problems, the more able he/she is to manage them. PMID- 10860064 TI - Nonsurgical management of the anterior open bite: a review of the options. AB - The open bite malocclusion has been described as being of 2 types: dental and skeletal. Proper differentiation is essential in determining the appropriate corrective measures. Dental open bites are generally more responsive to treatment with orthodontics alone, whereas skeletal open bites often require a combination of orthodontics and orthognathic surgery. Patient selection and treatment principles for nonsurgical open bite treatment are discussed, and a review of various methods of treatment for the skeletal open bite is presented. Posttreatment stability and retention concerns are addressed. PMID- 10860065 TI - Terminology: semantics of postorthodontic treatment changes in the dentition. AB - Correct terminology eliminates confusion in communication between clinicians, as well as between clinician and patient. Long-term stability seems to be an elusive goal, because the terminology in this respect eludes to the various changes occurring in the posttreatment dentition. However, a stable orthodontic result can be achieved when the physiologic changes that naturally occur in the dentition are considered as part of the long-term result. Standardization of terminology is therefore important. PMID- 10860066 TI - Methodologies for evaluating long-term stability of dental relationships after orthodontic treatment. AB - A comprehensive review of literature which considers methodologies for studying long-term occlusal stability is presented. This article focuses on the evaluation of plaster study models because occlusal changes are best reflected in longitudinal casts. Of particular interest is the assessment of crowding in the dentition and the various physical and mathematical procedures used to evaluate the measurement of space available. Indices used to assess the overall occlusal results of treatment are also presented. PMID- 10860067 TI - A review of changes in lower arch alignment from seven to fifty years. AB - Changes in alignment in the untreated lower arch were studied at various developmental stages: 7 to 10 years, 10 to 12 years, 12 to 15 years, 13 to 18 years, 18 to 21 years, 21 to 28 years, and 18 to 50 years. On average, crowding decreased between 7 and 12 years and increased thereafter. The maximum increase occurred in the teenage years between 13 and 18, little or no change occurred in the third decade, and small increases occurred later in life. The possible cause of these changes is discussed in relation to the deterioration in alignment reported in orthodontically treated patients after retention. PMID- 10860068 TI - A longitudinal evaluation of extraction versus nonextraction treatment with special reference to the posttreatment irregularity of the lower incisors. AB - A tendency exists in contemporary orthodontics to pursue a completely non extraction philosophy. Moreover, it has been shown that the extraction versus non extraction debate is still with us. Controversy exists as to which treatment decision will eventually lead to orthodontic stability. It is thus imperative to conduct investigations on long-term changes of the dentition in both treatment regimens. The present study serves as an example of such a longitudinal study. A random sample, inclusive of both extraction and non-extraction treatments, was examined with respect to long-term stability and an assessment was made as to whether one treatment option favors success over the other. It was concluded that the correct initial treatment choice will not only lead to correction of the malocclusion, but will also ensure clinically acceptable stability with no significant differences between extraction and non-extraction treatments. PMID- 10860069 TI - Transverse dimension and long-term stability. AB - This article emphasizes the critical importance of the skeletal differential between the width of the maxilla and the width of the mandible. Undiagnosed transverse discrepancy leads to adverse periodontal response, unstable dental camouflage, and less than optimal dentofacial esthetics. Hundreds of adult retreatment patients corrected for significant maxillary transverse deficiency using surgically assisted maxillary expansion (similar to osseous distraction) has produced excellent stability. Eliciting tooth movement for children (orthopedics, lip bumper, Cetlin plate) in all three planes of space by muscles, eruption, and growth, develops the broader arch form (without the mechanical forces of fixed or removable appliances) and has also demonstrated impressive long term stability. PMID- 10860070 TI - Postretention mandibular incisor stability after premolar serial extractions. AB - The purpose of this study was to evaluate the mandibular incisor alignment in serial extraction cases, using the longitudinal dental cast records of the Burlington Growth Center as a control sample. Various parameters were investigated and the statistical differences determined between the treated and untreated groups. The results were also compared with data from serial extraction groups that subsequently had orthodontic treatment. Untreated subjects and subjects treated only with serial extractions showed similar longitudinal changes. However, the extraction group that also received orthodontic treatment appeared to show more lower incisor crowding long-term. No predictors for stability of clinical significance could be determined. Mechanotherapy influences the craniofacial and dentoalveolar dimensions, which appear to cause more long term lower incisor crowding. PMID- 10860071 TI - Stability and relapse of mandibular anterior alignment: University of Washington studies. AB - For more than 40 years, research in the Department of Orthodontics, University of Washington (Seattle, WA) has focused on a growing collection of more than 800 sets of patient records to assess stability and relapse of orthodontic treatment. All patients had completed treatment a decade or more before the last set of data. Evaluation of treated premolar extraction patients, treated lower incisor extraction patients, treated non-extraction cases with generalized spacing, patients treated with arch enlargement strategies, and untreated normals showed similar physiologic changes: (1) Arch length decreases after orthodontic treatment. (2) Arch width measured across the mandibular canine teeth typically reduces posttreatment, whether or not the case was expanded during treatment. (3) Mandibular anterior crowding during the posttreatment phase is a continuing phenomenon well into the 20-to-40 years age bracket and likely beyond. (4) Third molar absence or presence, impacted or fully erupted, seems to have little effect on the occurrence or degree of relapse. (5) The degree of post-retention anterior crowding is both unpredictable and variable and no pretreatment variables either from clinical findings, casts, or cephalometric radiographs before or after treatment seem to be useful predictors. PMID- 10860072 TI - Stability of orthodontic treatment: myth or reality--a peek at the future. PMID- 10860073 TI - Prophylactic antibiotic therapy prior to dental treatment for patients with end stage renal disease. AB - In the United States, there is a large and growing population of patients undergoing dialysis because of end-stage renal disease (ESRD). These patients present special management considerations for dentists, including antibiotic prophylaxis for the prevention of bacterial endocarditis (BE). ESRD patients, particularly those with an arteriovenous shunt for hemodialysis access, are predisposed to valvular endocarditis. Thus, BE prevention is the primary goal of antibiotic prophylaxis prior to dental or other invasive procedures in these patients. Bacteremia may predispose to infection of synthetic vascular access grafts, although this form of endovascular infection in ESRD patients has not been as well-characterized as BE. Antibiotic prophylaxis may be of some benefit for prevention of synthetic graft infections as well as BE. Poor dentist and physician compliance with BE prophylaxis regimens, as well as errors in dosing, timing, or duration of prophylaxis, have been reported. These problems are of particular concern in the treatment of chronically ill patients. In this article, we review the rationale for prophylactic antibiotic therapy prior to dental procedures in ESRD patients with vascular access. We also elaborate on the current American Heart Association guidelines for BE prophylaxis, and address special considerations for ESRD patients. PMID- 10860074 TI - Nursing assistants' opinions of oral health care provision. AB - Understanding the attitudes and perceptions about oral health in nursing assistants (NAs) may facilitate efforts to improve daily oral care in long-term care settings. By exploring the attitudes of individuals charged with daily oral care, we may gain insight into the level of care provided for the residents. To explore motivation for oral care by NAs, we developed a 28-item survey. The survey included descriptive information and a 20-item Likert-type instrument dealing with oral care for self-care and dependent individuals. Factor analysis was used to test the validity of the constructs intended to be measured by the survey items. The results indicated favorable responses to knowledge items and items related to the importance of oral health in general. However, the responses to questions related to amount of time to perform mouth care, the risk of being bitten by a resident, resident cooperation, and myths about oral health in aging revealed significant variation by NAs descriptive variables. An understanding of the implications of NAs' perceptions, values, and knowledge may provide impetus for new strategies for improving oral health and daily care in long-term-care facilities. PMID- 10860075 TI - Perineural spread of squamous cell carcinoma of the lip: the importance of follow up and collaboration. AB - Perineural spread (PNS) of mucosal squamous cell carcinoma of the head and neck region occurs with a reported frequency of 2% to more than 27%. Patients previously diagnosed with and treated for head and neck cancer should be closely followed by both their physician and their dentist in order to facilitate the coordination of care. This case history demonstrates the results that can occur when a team approach to head and neck cancer is not followed, especially in a patient who is an infrequent and somewhat reluctant health care utilizer. Despite mandibular pain, the patient, who had a history of a carcinoma of the lower lip and had developed PNS, was not referred to a dentist. In patients with a previous history of squamous cell carcinoma, sensory and/or motor changes must be closely monitored when there is a suspicion of PNS. The outcome of this case supports improved collaborations between physicians and dentists in following head and neck cancer patients. PMID- 10860076 TI - Dental neglect: definition and prevention in the Louisiana Developmental Centers for patients with MRDD. AB - When dental services and appropriate use of restraints are used for persons who are severely and profoundly mentally retarded and developmentally disabled (MRDD), oral health is enhanced and dental neglect is controlled or even reduced. This paper defines dental neglect and examines the consequences to persons who are MRDD when appropriate restraints and preventive dental home care are not used. Oral health outcome assessment data of this MRDD population indicate the presence or absence of dental neglect. The outcome assessment measures the level of oral health within the developmental center. The Louisiana Department of Health and Hospitals (DHH) operates nine developmental centers, where approximately 2000 individuals with severe to profound mental handicaps reside. Two-thirds (66%) of this population are resistive to the delivery of clinical and preventive dental services. The responsibilities and roles of dental health professionals, long-term-care facility administrators, and direct service staff are discussed to help reduce dental neglect in these centers. For these individuals to receive quality and comprehensive dental care, appropriate use of restraint/physical hold is necessary. The Dental Health Resource Program (DHRP) developed guidelines for delivering preventive and clinical dental services to reduce confusion and misinterpretation of the definition and use of restraint in the Louisiana Developmental Centers (LADCs). PMID- 10860077 TI - Respiratory pathogen colonization of the dental plaque of institutionalized elders. AB - Although it has been established that aspiration of pharyngeal bacteria is the major route of infection in the development of nosocomial pneumonia, colonization of the pharyngeal mucosa by respiratory pathogens has been shown to be a transient phenomenon. It has been suggested that the dental plaque may constitute an additional, possibly more stable, reservoir of respiratory pathogens. The purpose of this study was to assess the prevalence of oral colonization by potential respiratory pathogens in a group of elderly (mean age = 75.9 yrs) chronic-care-facility residents (n = 28) and a group of age-, gender-, and race matched outpatient control subjects (n = 30), with specific attention to plaque present on tooth, denture, and oral mucosal surfaces. Plaque scores on teeth and dentures were significantly higher in the chronic-care-facility (CCF) subjects than in the dental outpatient control (DOC) subjects (PII 2.3 vs. 1.2 and denture plaque 1.4 vs. 0.3). While no subjects in the DOC group were found to be colonized with respiratory pathogens (> 1.0% of the cultivable aerobic flora), 14.3% (4/28) of the CCF subjects were found to be colonized. Oral colonization with respiratory pathogens in CCF subjects was associated with the presence of chronic obstructive pulmonary disease (COPD) and higher plaque scores. These results suggest that deficient dental plaque control and the presence of COPD may be related to respiratory pathogen colonization of dental plaque in chronic-care facility residents. PMID- 10860078 TI - A solitary maxillary central incisor treated orthodontically: a case report. AB - The presentation of a solitary maxillary central incisor is reported and its association with other congenital abnormalities discussed. It is an important finding for dental health professionals, since it may indicate the presence of other significant midline congenital abnormalities. It may also indicate the presence of associated disorders that profoundly affect growth and development and which, once identified, may be treated. PMID- 10860079 TI - Geriatric dentistry in long-term-care facilities: current status and future implications. AB - The status of oral health care of geriatric patients in long-term-care facilities is reviewed. Barriers to improving the dental management of institutionalized geriatric patients are discussed. The elderly population segment is the fastest growing population, which presents a great burden to dental health professionals. Institutionalized geriatric patients are currently not receiving adequate care, and the number of these patients will double by the year 2020. The future implications are of considerable importance. Commitment by dental professionals to meet the oral health needs of this underserved population is key to the improvement of and change in the current status of this population. PMID- 10860080 TI - Oral and maxillofacial pathology case of the month. Chronic desquamative gingivitis. PMID- 10860081 TI - Increased risk for dental caries in asthmatic children. PMID- 10860082 TI - Oral and maxillofacial pathology case of the month. Xerostomia. PMID- 10860083 TI - The 1998 C.T. Rowland Award orthodontic case report. PMID- 10860084 TI - Prevalence of pulpal calcifications in the primary dentition of Hispanic children. PMID- 10860085 TI - Oral and maxillofacial pathology case of the month. Fungiform papilla. PMID- 10860086 TI - The Declaration of Helsinki: revising ethical research guidelines for the 21st century. PMID- 10860087 TI - Male hormonal contraception: a safe, acceptable and reversible choice. PMID- 10860088 TI - Prostate cancer: what should be the sequel to diagnosis? PMID- 10860089 TI - Evidence for evidence-based medicine at the coalface. PMID- 10860090 TI - Death and readmission in the year after hospital admission with cardiovascular disease: the Hunter Area Heart and Stroke Register. AB - OBJECTIVES: To compare outcomes one year after hospital admission for patients initially discharged with a diagnosis of acute myocardial infarction (AMI), other ischaemic heart disease (other IHD), congestive heart failure (CHF) or stroke. DESIGN: Cohort study. SETTING: Hunter Area Heart and Stroke Register, which registers all patients admitted with heart disease or stroke to any of the 22 hospitals in the Hunter Area Health Service in New South Wales. PATIENTS: 4981 patients with AMI, other IHD, CHF or stroke admitted to hospital as an emergency between 1 July 1995 and 30 June 1997 and followed for at least one year. MAIN OUTCOME MEASURES: Death from any cause or emergency hospital readmission for cardiovascular disease. RESULTS: In-hospital mortality varied from 1% of those with other IHD to 22% of those with stroke. Almost a third of all patients discharged alive (and 38% of those aged 70 or more) had died or been readmitted within one year. This varied from 22% of those with stroke to 49% of those with CHF. The causes of death and readmission were from a spectrum of cardiovascular disease, regardless of the cause of the original hospital admission. CONCLUSIONS: Data from this population register show the poor outcome, especially with increasing age, among patients admitted to hospital with cardiovascular disease. This should alert us to determine whether optimal secondary prevention strategies are being adopted among such patients. PMID- 10860091 TI - HIV and AIDS in aboriginal and Torres Strait Islander Australians: 1992-1998. The National HIV Surveillance Committee. AB - OBJECTIVE: To describe the epidemiological pattern of newly diagnosed HIV infection and AIDS among Indigenous Australians. DESIGN AND SETTING: National surveillance for newly diagnosed HIV infection and AIDS in Australia. Information on Indigenous status was sought at HIV/AIDS notification in all State/Territory health jurisdictions, except the Australian Capital Territory, and Victoria before June 1998. MAIN OUTCOME MEASURES: Number of people with newly diagnosed HIV per year and population rate of HIV diagnosis; demographic characteristics of people with HIV and AIDS diagnoses by Indigenous status. RESULTS: From 1992 to 1998, 127 Indigenous Australians were newly diagnosed with HIV infection and 55 were diagnosed with AIDS. The population rate of HIV diagnosis among Indigenous Australians (5.23/100,000 per year) was similar to that among non-Indigenous Australians (5.51/100,000 per year). The annual number of HIV diagnoses among Indigenous people was relatively stable, but among non-Indigenous people it declined steadily over time. A higher proportion of Indigenous people diagnosed with HIV were women (26.8% v 8.9%; P < 0.001). Although male homosexual contact was the predominant source of exposure for both Indigenous (46.7%) and non Indigenous (75.0%) people with HIV infection, exposure by heterosexual contact (36.7% v 15.3%; P < 0.001) was reported more frequently among Indigenous people. CONCLUSION: Although HIV incidence was similar among Indigenous and non Indigenous Australians, the lack of a recent decline in incidence and the higher proportion of Indigenous people exposed to HIV by heterosexual contact indicate the need to intensify interventions to prevent HIV transmission among Indigenous people. PMID- 10860092 TI - Prostate cancer in Victoria in 1993: patterns of reported management. AB - OBJECTIVE: To describe the management of newly diagnosed prostate cancer in 1993 during the early prostate specific antigen (PSA) era. DESIGN: Survey of medical practitioners involved in the management of a total sample of incident prostate cancer cases selected from a population-based cancer registry. The survey was conducted in 1996, and the sample was followed up until 1998, to obtain five-year survival data on all patients. SETTING: The State of Victoria, including both public and private health sectors. PATIENTS: All men who were newly diagnosed with prostate cancer in the six months January-June 1993. MAIN OUTCOME MEASURES: Reported management by method of diagnosis; staging investigations; and treatment by observation, hormonal therapy, radical radiotherapy or radical prostatectomy. RESULTS: 1048 of 1117 (94%) cases diagnosed were surveyed. Most of the men (858 [82%]) were older than 65 years: 117 (11%) cancers were detected by screening asymptomatic men, and a further 269 (26%) were found by testing of men with symptoms ("case-found"). The 259 (25%) men treated with definitive local therapies (prostatectomy and curative radiotherapy) were younger (< 75 years), and their disease was clinically more localised (clinical stage, T1-2) and they were often found by screening or case-finding. Men given hormonal therapy (407; 39%) or managed without treatment (373; 36%) tended to be older and more likely to have been diagnosed by transurethral resection of the prostate (TURP). The overall relative survival at five years was 86% and was decreased in men with cancers of higher histological grade or more advanced clinical stage, or who had higher PSA levels. CONCLUSIONS: Although a third of patients were detected by screening or case-finding early in the PSA era, definitive local therapies were used infrequently (25% of the total sample). Most received appropriate treatment. PMID- 10860093 TI - Angle grinder injuries: a cause of serious head and neck trauma. AB - Over the past 12 months, the Victorian Trauma Centre at the Alfred Hospital, Melbourne, has dealt with serious head and neck injuries associated with angle grinder use. Three cases are presented, documenting the circumstances and severity of these injuries and subsequent management. Angle grinder injuries are a source of serious morbidity and mortality, much of which is preventable. PMID- 10860094 TI - Erectile dysfunction, sildenafil and cardiovascular risk. AB - Cardiovascular risk factors are commonly associated with erectile dysfunction and should be identified and treated. Patients with cardiovascular diseases should be assessed and counselled regarding their fitness for sexual activity. The danger of concurrent use of sildenafil and nitrates under any circumstances, regardless of age and sex, must be highlighted at all levels of the community. Sildenafil is absolutely contraindicated in patients receiving treatment with long-acting nitrates for ischaemic heart disease. Patients who need sublingual short-acting nitrates infrequently should not be precluded from taking sildenafil, provided they are aware that sildenafil is not to be taken within 24 h of taking the nitrate. PMID- 10860095 TI - Is the grass greener? The link between cannabis and psychosis. PMID- 10860096 TI - Clinical practice guidelines: reality bites. AB - In 1995, the National Health and Medical Research Council (NHMRC) announced its commitment to developing evidence-based clinical practice guidelines "to promote best practice linked to outcomes and effective cost management". To date, resources for dissemination and implementation have been identified for only two guidelines developed by the NHMRC as part of that commitment. Clinical practice guidelines for the management of men with lower urinary tract symptoms (LUTS) were launched in 1997. We were members of the working party that developed these guidelines, and here we give our personal account, offering insights into the tensions and contradictions impeding the translation of political commitment to evidence-based medicine into policy and practice. PMID- 10860097 TI - The Declaration of Helsinki and research in vulnerable populations. AB - Mooted changes to the Declaration on the agenda of the World Medical Association have sparked a vigorous debate on international research issues. The medical, research and ethics communities in Australia need to participate more broadly in this debate. PMID- 10860098 TI - Hospitals and hospitalists: an alternative view. PMID- 10860099 TI - Hospitals and hospitalists: an alternative view. PMID- 10860100 TI - Addressing dysfunctional doctors. PMID- 10860101 TI - The fate of "intramuscular" iron injections. PMID- 10860102 TI - Care of older people in acute-care hospitals: do we know how? PMID- 10860103 TI - Photosensitivity associated with herbal preparations of St John's wort (Hypericum perforatum) PMID- 10860104 TI - Pelvic fractures diagnosed by bone scintigraphy in patients with normal radiographs after a fall. PMID- 10860105 TI - Acute and recurrent skin ulceration after spider bite. PMID- 10860106 TI - Humour in medical teaching. PMID- 10860107 TI - Radiation protection recommendations as applied to the disposal of long-lived solid radioactive waste. A report of The International Commission on Radiological Protection. AB - (79) Waste, by definition, has no benefit. It should be viewed as one aspect of the beneficial practice that gave rise to it. Furthermore, radioactive waste management should be placed in the context of the management of society's waste in general. (80) A major issue in evaluating the acceptability of a disposal system for long-lived solid radioactive waste is that doses or risks may arise from exposures in the distant future. There is uncertainty surrounding any estimate of these doses or risks due to lack of knowledge about future conditions. Such exposures are treated as potential exposures as their magnitude depends on future processes and conditions that have probabilities associated with them. (81) Nevertheless, the Commission recognises a basic principle that individuals and populations in the future should be afforded at least the same level of protection from the action of disposing of radioactive waste today as is the current generation. This implies use of the current quantitative dose and risk criteria derived from considering associated health detriment. Therefore, protection of future generations should be achieved by applying these dose or risk criteria to the estimated future doses or risks in appropriately defined critical groups. These estimates should not be regarded as measures of health detriment beyond times of around several hundreds of years into the future. In the case of these longer time periods, they represent indicators of the protection afforded by the disposal system. (82 Constrained optimisation is the central approach to evaluating the radiological acceptability of a waste disposal system; dose or risk constraints are used rather than dose or risk limits. By this transition from limitation to optimisation, the needs of practical application of the radiological protection system to the disposal of long-lived solid waste disposal are met: determination of acceptability now for exposures that may occur in the distant future. Optimisation should be applied in an iterative manner during the disposal system development process and should particularly cover both site selection and repository design. (83) Two broad categories of exposure situations should be considered: natural processes and human intrusion. The latter only refers to intrusion that is inadvertent. The radiological implications of deliberate intrusion into a repository are the responsibility of the intruder. Assessed doses or risks arising from natural processes should be compared with a dose constraint of 0.3 mSv per year or its risk equivalent of around 10(-5) per year. With regard to human intrusion, the consequences from one or more plausible stylized scenarios should be considered in order to evaluate the resilience of the repository to such events. (84) The Commission considers that in circumstances where human intrusion could lead to doses to those living around the site sufficiently high that intervention on current criteria would almost always be justified, reasonable efforts should be made at the repository development stage to reduce the probability of human intrusion or to limit its consequences. In this respect, the Commission has previously advised that an existing annual dose of around 10 mSv per year may be used as a generic reference level below which intervention is not likely to be justifiable. Conversely, an existing annual dose of around 100 mSv per year may be used as a generic reference level above which intervention should be considered almost always justifiable. Similar considerations apply in situations where the thresholds for deterministic effects in relevant organs are exceeded. (85) Compliance with the constraints can be assessed by utilising either an aggregated risk-oriented approach, with a risk constraint, or a disaggregated dose/probability approach, with a dose constraint, or a combination of both. A similar level of protection can be achieved by any of these approaches; however, more information may PMID- 10860108 TI - Progression of consequences among heavy-drinking college students. AB - Among a sample of 180 male and 226 female undergraduates, 84.2% reported a heavy drinking episode (5+ drinks for men, 4+ for women) within the previous 90 days. Principal-components analysis revealed 3 alcohol-related problem factors among the heavy drinkers (Careless Behavior, Risky/Reckless Behavior, and Authority Problems). Nearly all heavy drinkers experienced a careless behavior that was due to drinking (92.7%), and many reported a risky/reckless behavior (60.2%), yet only one third (33.9%) experienced an authority problem. Guttman scaling procedures revealed a progression from Careless Behavior to Reckless/Risky Behavior to an Authority Problem. Heavy drinkers with an authority problem drank more frequently, consumed more when drinking, endorsed more alcohol expectancies, and reported earlier ages of initial and regular drinking than other groups. PMID- 10860109 TI - An event-based analysis of aggression women experience in bars. AB - The environmental and social circumstances in bars are likely to contribute to an individual's risk for becoming a victim. The present study was designed to assess differences that exist in these circumstances between times when aggression does and does not occur in bars. Participants were a subsample of 46 women who had previously participated in a larger survey of female bar drinkers. Using daily logs and biweekly interviews over 12 weeks, the author assessed the drinking patterns and aggressive experiences of these women. At times when aggression occurred women spent less time in the bar, consumed more alcohol, and reported feeling more intoxicated. Differences were found between sexual and nonsexual incidents and among the women on the basis of the type of aggression they had experienced. Women who experienced both sexual and nonsexual aggression reported using other drugs in addition to alcohol when drinking in bars. These findings and their implications for prevention are discussed. PMID- 10860110 TI - Determinants and effects of attentional bias in smokers. AB - Much research has shown that individuals exhibit an attentional bias to stimuli related to their current concerns or pathologies. Using the emotional Stroop task, we investigated attentional bias in smokers. Ninety-six smokers either abstained from smoking for 24 hr or smoked normally before color-naming smoking related and neutral words. Both a blocked format (smoking and neutral words presented in separate blocks) and an unblocked format (smoking and neutral words presented in a mixed random sequence) were used. In the blocked format, abstinence caused an attentional bias to smoking-related stimuli, and the degree of attentional bias predicted the latency to the first cigarette of the morning. However, different results were obtained from the unblocked version of the task. We conclude that the emotional Stroop task is a useful tool to measure attentional bias in smokers and could be used in cessation studies. PMID- 10860111 TI - Gender differences and similarities in the personality correlates of adolescent alcohol problems. AB - The personality traits of behavioral under-control (BU) and negative emotionality (NE) are associated with alcohol problems. The authors examined gender differences in the associations of BU and NE with alcohol problems in 710 adolescents recruited from community and treatment sources. Multiple measures were used to characterize each construct, and the specified 2-factor model provided a reasonably good fit to the data. ANCOVAs were used to examine each construct by gender across four groups: never-regular drinkers, regular drinkers, and those with DSM-IV alcohol abuse and alcohol dependence. Males had significantly higher BU and lower NE than did females. BU and NE both increased with degree of alcohol problems. However, there was not a significant Gender x Alcohol Group interaction for BU or NE. Although there are gender differences in levels of BU and NE, mechanisms of alcohol involvement related to these 2 personality traits may operate similarly in adolescent males and females. PMID- 10860112 TI - Determinants of attrition from cessation treatment in smokers with a history of major depressive disorder. AB - Attrition from smoking cessation treatment by individuals with a history of major depression was investigated. An investigation of preinclusion attrition examined differences between eligible smokers who did (n = 258) and did not (n = 100) attend an initial assessment session. Postinclusion attrition was investigated by comparing early dropouts (n = 33), late dropouts (n = 27), and treatment completers (n = 117). Those who failed to attend the assessment session were more likely to be female, to smoke cigarettes with higher nicotine content, and to have a history of psychotropic medication use. Early-treatment dropouts reported a higher smoking rate than late-treatment dropouts and endorsed more symptoms of depression than late dropouts and treatment completers. Results are compared with previous investigations of smoking cessation attrition, and implications for treatment and attrition prevention are discussed. PMID- 10860113 TI - DWI prevention: profiles of drinkers who serve as designated drivers. AB - This study was designed to profile drinkers who serve as designated drivers (DDs) and to determine if drinkers who are at risk for driving while intoxicated (DWI) serve as DDs. Bivariate and logistic regression analyses on data from 1,393 computer-assisted telephone interviews (CATIs) and 913 bar-room surveys showed that DDs, relative to non-DDs, tend to be at-risk, heavier drinkers. Logistic regression using CATI data showed that DDs were more often heavy drinkers and reported higher levels of driving after drinking and riding with intoxicated drivers (RID). Logistic regression using bar-room data showed that DDs reported more driving after drinking, in spite of drinking less often outside the home. DDs were also much more likely to have used a DD. These findings are consistent with those from several related studies that showed that drinkers who used DDs or free safe (taxi) rides tended to be heavier drinkers who reported more DWI and RID (B. D. Caudill, W. M. Harding, & B. Moore, in press-a, in press-b). Future research may benefit from examining why at-risk drinkers take steps to avoid DWI on some occasions but not others. PMID- 10860114 TI - Problem drinking behavior in two community-based samples of adults: influence of gender, coping, loneliness, and depression. AB - The authors examined the extent to which the relationships among coping, loneliness, depression, and 3 problematic drinking behaviors varied as a function of gender in 2 community-based samples of young adults (19-39 years old). Regression analyses revealed that (a) after controlling for the quantity and frequency of alcohol typically consumed, the 3 psychosocial variables were significantly related to frequency of intoxication, binge drinking, and drink tossing behaviors; (b) not all predictors were related to all problem drinking behaviors; (c) the predictors that were significant varied as a function of the 2 cohorts; and (d) with the exception of frequency of intoxication in the younger cohort, the associations between the predictors and problematic drinking behaviors tended to be similar for men and women. Future directions for research are discussed. PMID- 10860115 TI - Demographic, individual, and interpersonal predictors of adolescent alcohol and marijuana use following treatment. AB - A vulnerability model of adolescent substance abuse treatment outcome provided the basis for selection of demographic, individual, interpersonal, and treatment factors to predict the follow-up use of alcohol and marijuana in a sample of adolescents (N = 225) with psychoactive substance use disorders. Pretreatment levels of sibling substance use and aftercare participation predicted alcohol and marijuana use during the first 6 months posttreatment. Pretreatment levels of deviant behavior also predicted the use of marijuana at 6-month follow-up. Peer substance use at intake and 6-month posttreatment both predicted substance use frequency outcomes at 12-month follow-up. Alcohol and marijuana use frequencies at 6-month follow-up also predicted continued use for these substances throughout the remainder of the 1st posttreatment year. Shorter treatment length and being male were risk factors for alcohol use during the 2nd half of the 1st posttreatment year. Elevated psychological substance dependence at 6-month follow up was a unique risk factor for subsequent marijuana use. Findings support conceptual models that attempt to explain adolescent substance abuse treatment outcome in terms of relationships among demographic, individual, interpersonal, and treatment factors. PMID- 10860116 TI - Parenting practices as predictors of substance use, delinquency, and aggression among urban minority youth: moderating effects of family structure and gender. AB - This study examined how parenting factors were associated with adolescent problem behaviors among urban minority youth and to what extent these relationships were moderated by family structure and gender. Sixth-grade students (N = 228) reported how often they use alcohol, smoke cigarettes, or engage in aggressive or delinquent behaviors; a parent or guardian reported their monitoring and other parenting practices. Findings indicated that boys and those from single-parent families engaged in the highest rates of problem behavior. More parental monitoring was associated with less delinquency overall, as well as less drinking in boys only. Eating family dinners together was associated with less aggression overall, as well as less delinquency in youth from single-parent families and in girls. Unsupervised time at home alone was associated with more smoking for girls only. Implications for prevention interventions are discussed. PMID- 10860117 TI - Alcohol expectancies as a mediator of the relationship between injury and readiness to change drinking behavior. AB - There is a well-established relationship between alcohol expectancies and drinking behavior. The purpose of the present study was to extend the literature by examining the role of alcohol expectancies in determining readiness to change drinking behavior among injured emergency department patients who screened positive for hazardous drinking. Negative expectancies were found to partially mediate the relationships of alcohol-related injuries and injury aversiveness to readiness to change drinking behavior. Results suggest that negative alcohol expectancies are a potential means of increasing patients' readiness to change drinking behavior. PMID- 10860118 TI - Group self-identification as a prospective predictor of drug use and violence in high-risk youth. AB - This study provides a 1-year prospective analysis of group self-identification as a predictor of adolescent drug use and violence. Youth identified with discrete groups. In most comparisons, 1 year later, a high-risk group reported greater levels of drug use and violence-related exposure than other groups, and the statistical relation between group self-identification and drug use or violence remained after baseline assessment of the drug use or violence measure was controlled for. This is the 1st study to demonstrate that group self identification is a significant prospective predictor of drug use and other problem behaviors. PMID- 10860119 TI - Predicting substance abuse among youth with, or at high risk for, HIV. AB - This article describes data from 4,111 males and 4,085 females participating in 10 HIV/AIDS service demonstration projects. The sample was diverse in age, gender, ethnicity, HIV status, and risk for HIV transmission. Logistic regression was used to determine the attributes that best predict substance abuse. Males who were younger; HIV positive; homeless; involved in the criminal justice system; had a sexually transmitted disease (STD); engaged in survival sex; and participated in risky sex with men, women, and drug injectors were most likely to have a substance abuse history. For females, the same predictors were significant, with the exception of having an STD. Odds ratios as high as 6 to 1 were associated with the predictors. Information about sexual and other risk factors also was highly predictive of substance abuse issues among youth. PMID- 10860120 TI - Measuring adolescent alcohol outcome expectancies. AB - The psychometric properties and construct validity of the Comprehensive Effects of Alcohol (CEOA) questionnaire were compared with those of the Alcohol Expectancy Questionnaire--Adolescent version (AEQ-A) in relation to adolescent alcohol consumption. Both measures of adolescent alcohol expectancies were found to be internally reliable and temporally stable. Alcohol use was significantly associated with subjective evaluations for Cognitive and Behavioral Impairment and Self-Perception on the CEOA and with expected effects for Cognitive and Motor Impairment and Changes in Social Behavior on the AEQ-A. Compared with the AEQ-A, the CEOA explained more variance in quantity (28%) and a similar variance in frequency (15%) for adolescent alcohol use (AEQ-A quantity = 20%, frequency = 15%). Whereas the general content and psychometric properties of the 2 measures are markedly similar, the Likert response format, shorter length, and assessment of both expected effects and subjective evaluations with the CEOA may offer measurement advantages over the AEQ-A. PMID- 10860121 TI - Charles University and pigment cell research. AB - A short review on the pigment cell research at Charles University in Prague is presented. PMID- 10860122 TI - Surgical therapy of thymomas. AB - Surgical treatment of thymomas is indicated for Masaoka stage 1 to 3. We are not in favor of mini-invasive techniques. We consider a gold standard to be sternotomy followed by a tumor removal and extended thymectomy. We are convinced it is necessary to perform sternotomy, tumor removal and extended thymectomy after a thymoma resection through thoracotomy to prevent a late onset of myasthenia gravis. In stages 2 to 3 actinotherapy along with chemotherapy should follow surgery to increase the patient's chances for a prolonged survival (Tab. 10). PMID- 10860123 TI - [New methods of surgery on the arteries supplying the brain]. AB - INTRODUCTION: Brain ischaemia as a sequelae of atherosclerosis involving the brain-supplying arteries is one of the commonest causes of death in the industrialized nations. MATERIAL AND METHODS: A total of 236 procedures on the internal and common carotid arteries in 227 patients with a mean age of 67 years were performed. General anaesthesia was used in 18.5% of patients, cervical block in 81.5%. RESULTS: The hospitalization morbidity of our group of patients was 8.9%, hospitalization mortality was zero. No association between the technique of anaesthesia and morbidity was found. However, cervical block allowed a marked shortening of postoperative hospitalization time from 5.5 days to 2 days. CONCLUSION: The results obtained justify surgical procedures on the internal and common carotid arteries even in asymptomatic stenoses, and are especially useful before a scheduled cardiac surgical or vascular surgical procedure. PMID- 10860124 TI - Huntington's disease: the relationship between clinical signs, CAG repeats and the atrophy of the caudate nucleus in CT scans. AB - We compared clinical data from 45 patients with Huntington's Disease (HD) with CAG triplet repeats and the planimetric measurement of the caudate nucleus head area (CNHA) in CT scans. The mean age of patients was 50.4 yrs (SD +/- 10.2), the mean duration of HD 7.4 yrs (4.6), the mean age at the onset of HD 43.1 yrs (11.1). HD started with motor symptoms in 28 patients, with psychiatric symptoms in 14 patients, the history was unknown in 3 patients. The paternal transmission was observed in 29 patients, the maternal one in 12 patients, unknown in 4 patients. The mean number of CAG repeats was 46.6 (6.1). The mean CNHA was 0.4 cm2 (0.1). We found statistically significant reversed correlation between CAG repeats and the age at the onset of HD (p < 0.0001, r -0.6). The earlier onset of HD in patients with the paternal transmission compared to the maternal one was found statistically significant (p < 0.05). This phenomenon was not related to the larger number of CAG triplets in patients with the paternal transmission. No differences either of the age at the onset of HD or numbers of CAG repeats were found between subgroups of HD patients starting with motor or psychiatric symptoms. We also observed the significant reversed correlation between the duration of HD and CNHA measurement (p < 0.001, r -0.5). Even in the earliest stage of HD patients showed the marked atrophy of CNHA. PMID- 10860125 TI - [Sleep apnea syndrome in acromegaly. Treatment and development-- retrospective analysis]. AB - In total 56 subjects with acromegaly (37 men and 19 females) were examined in attempt to find sleep apnoea syndrome (SAS). The examination consisted of clinical examination and of all-night monitoring MESAM4 or polyMESAM. The diagnosis of SAS was established in 76.4% of subjects: 31 men and 11 females, average age 51.8 (SD +/- 9.6) years. The average ODI (oxygen desaturation index- number of oxygen saturation drops per 1 hour of sleep) of SAS patients was 29.2 (+/- 20.7). The therapy of SAS was recommended to 27 patients: sleeping position on the side (3 patients), reduction of the weight (8 patients), change of hypnotic drug (1 patient) and CPAP--continuous positive airway pressure (24 patients). CPAP titration was performed in 18 patients (in one subject the titration was repeated 4 years later). CPAP was titrated within all-night polysomnography in 10 subjects and in 9 subjects using self adjusting CPAP. Primary acceptance of CPAP was 94.4%. The average CPAP pressure was 7.8 (+/- 2.1) mbar. The theoretical duration of CPAP use was 546.6 (+/- 533.7) days. Long term compliance was considered as sufficient (weekly 25 hours or more) in 66.7% of patients. Ten patients underwent important acromegaly therapy or its change during the follow up and the improvement or the disappearance of SAS symptoms occurred in 6 of them. PMID- 10860126 TI - [Excitatory amino acid antagonists and potentiation of cortical evoked potentials]. AB - Changes of evoked potentials under the influence of NBQX (a non-NMDA receptor antagonist), MK-801 (a NMDA receptor antagonist) and GDEE (a nonselective antagonist of glutamate receptor) were studied. GDEE augmented potentiation and was marked progression of potentiation with increasing number of stimuli. There was no potentiation of responses in relation to the number of stimulus in a series in experiments with both MK-801 and NBQX. Interpretation of results with NBQX and MK-801 is difficult. PMID- 10860127 TI - [Contribution of professor R. Cihak to paleoanthropology and historical anthropology]. PMID- 10860128 TI - Incidence of intravenous site reactions in neurotrauma patients receiving valproate or phenytoin. AB - OBJECTIVE: To determine the incidence of intravenous site reactions to phenytoin and valproate in a large population of patients with neurotrauma. DESIGN: Retrospective chart review of two double-blind, randomized clinical trials evaluating the use of antiepileptic drugs to prevent posttraumatic seizures in patients with neurotrauma: phenytoin versus placebo (n = 390), and valproate versus phenytoin with placebo (n = 385). Information collected from the charts included the number, type, and location of intravenous lines and intravenous site events. SETTING: Tertiary care trauma and university teaching hospital. MAIN RESULTS: Intravenous site reactions occurred in 18% and 25% of patients receiving valproate or phenytoin, respectively, with the majority of events (70%) occurring in the first intravenous site. Patients received the neurosurgery study drug (NSSD) by either central or peripheral lines; all intravenous site reactions occurred in peripheral administration sites. When patients who received the drug by central line during the course of therapy were excluded, the estimated incidence of site reactions was 21% and 30% for valproate and phenytoin, respectively (p = 0.056). The time to the first event was shorter with phenytoin compared with valproate (2.0 +/- 1.3 vs. 3.0 +/- 1.9 d; p = 0.009). Fewer adverse events were noted with phenytoin in the phenytoin-without-valproate study than in the phenytoin-with-valproate study, with 4.3% and 8.2% of intravenous site events recorded in patients receiving placebo or phenytoin, respectively. There was no significant difference in the number of intravenous lines per patient used during NSSD drug infusion for phenytoin versus placebo or phenytoin versus valproate. CONCLUSIONS: Both intravenous phenytoin and valproate resulted in intravenous site reactions, with the loading doses responsible for the majority of the events. PMID- 10860129 TI - Long-term patient adherence to antiretroviral therapy. AB - OBJECTIVE: To measure patient adherence to antiretroviral therapy over a two-year period and to identify factors impacting adherence. METHODS: In a regional HIV treatment center, 100 consecutive patients starting any new antiretroviral agent were enrolled in this study, which consisted of a one-year retrospective data review and a one-year prospective component. The tools used for evaluating adherence were the monthly prescription refill data and a patient questionnaire. Data analyzed included overall adherence, adherence to individual antiretrovirals, and change in adherence over time, as well as factors reported as influencing adherence. RESULTS: Greater than 80% adherence in taking prescribed doses was seen in 75% of patients during the retrospective phase of the study; adherence increased to 84% in the prospective phase. Throughout the prospective phase of the study, monthly median adherence rates were 98-100%. Suboptimal adherence secondary to pill fatigue or number of daily pills did not occur. Reported nonadherence to dietary restrictions varied among drugs. The primary cause given for poor adherence was difficulty remembering followed by inconvenient dosing schedule and difficulty scheduling administration times around meals. At least one adherence tool was used by 61% of patients. A diagnosis of AIDS was associated with lower adherence in our patient population (p = 0.039); substance abuse and psychiatric history had no influence. CONCLUSIONS: Adherence to antiretroviral treatment regimens did not diminish over the two years studied. Several patients with poor adherence were identified, emphasizing the importance of addressing this issue both prior to and throughout treatment. A personalized approach by healthcare providers can optimize patient adherence to antiretroviral therapy by providing careful drug selection in addition to routine follow-up and the provision of information, feedback, and reminder systems. PMID- 10860130 TI - Pharmacists' knowledge and attitudes toward herbal medicine. AB - OBJECTIVE: The use and sales of herbal medications have increased dramatically over the past several years. Pharmacists are in an ideal position to educate patients about herbal medicines. This study was intended to determine the knowledge and attitudes of pharmacists regarding herbal medications. METHODS: A survey was distributed to pharmacists at several state and regional meetings in Virginia and North Carolina between August and October 1998. The survey evaluated demographic data, attitudinal scales, and a 15-item herbal medicine knowledge test. Pharmacists immediately returned the surveys to the distributor on completion. RESULTS: Of the 217 surveys distributed, 164 met the inclusion criteria for further evaluation. Of the pharmacists surveyed, 68.0% practiced in a community pharmacy, 45.1% had previous continuing education on herbal medications, and 73.6% sold herbal medications in their practice settings. The average score on the herbal knowledge test was 6.3 (maximum score of 15). Pharmacists with previous continuing education scored significantly higher than those without prior continuing education (p < 0.001). Of the 15 questions, the five that pharmacists were most likely to answer correctly assessed the uses of herbal medications. Additionally, pharmacists with prior continuing education or with access to herbal medication information at their practice site were more likely to agree that providing information about herbal medication is a pharmacist's professional responsibility (p = 0.02 and p = 0.01, respectively). CONCLUSIONS: The findings from this study demonstrate that pharmacists were more likely to answer correctly about the uses of herbal medications than about drug interactions, adverse drug effects, and precautions of herbal medications. Additionally, pharmacists with previous continuing education on herbal medications were more knowledgeable about these products. With the increasing use of herbal medications, there is a greater need for pharmacy training programs in this area. PMID- 10860131 TI - Selective prescribing of spasmolytics. AB - BACKGROUND: Daily clinical practice often differs largely from the clinical trial setting, so extrapolation of outcomes from trial data, such as safety, effectiveness, and economic outcomes, can be deceptive. Prescribers may intend to treat a selected group of patients with new drugs; this practice could result in significant bias in assessing outcomes of these agents during their use in daily clinical practice. OBJECTIVE: To evaluate what type of patient received tolterodine compared with the spasmolytic drugs previously marketed (oxybutynin, flavoxate, emepronium). DESIGN: An observational, follow-up study. SETTING: Eighteen collaborating community pharmacies. PATIENTS: Aged > or = 18 years, noninstitutionalized; initial therapy with tolterodine, oxybutynin, flavoxate, or emepronium. RESULTS: Tolterodine was often used as a second-line and even as a third-line treatment, and was prescribed to a "polluted" population in terms of concomitant psychotropic medication. Tolterodine users were 7.5 times more likely to have received another spasmolytic drug (RR 7.5, 95% CI 4.8 to 11.9). In addition, these patients more frequently used antiparkinsonian drugs (RR 4.1, 95% CI 1.6 to 10.4) as well as antipsychotic drugs (RR 2.9, 95% CI 1.4 to 6.2). There was a small difference in concomitant use of antidepressants and benzodiazepines between patients receiving tolterodine versus those taking other spasmolytic drugs. CONCLUSIONS: Tolterodine is prescribed for a population differing from that receiving previously marketed spasmolytic drugs. Selective prescribing should recognized when evaluating new drugs in daily clinical practice. Policy makers, such as pharmacy and therapeutics committees, should consider this aspect in their formulary decisions since selective prescribing can lead to unjustified conclusions about a drug's therapeutic effects (e.g., efficacy, safety, cost effectiveness). PMID- 10860132 TI - Helicobacter pylori eradication in clinical practice: retreatment rates and costs of competing regimens. AB - OBJECTIVE: To measure outcomes for eradication regimens for Helicobacter pylori infection in routine clinical practice. DESIGN: Retrospective analysis of an integrated medical and pharmacy claims database identified patients treated from June 1, 1995 through May 31, 1996, and followed the patients' claims until December 31, 1996. SETTING: The database represented multiple health plans throughout the US. PATIENTS: Patients were > or = 16 years old, continuously enrolled from April 1, 1995, to December 31, 1996, and met clinical (diagnostic or procedural) criteria. INTERVENTION: Patient cohorts were treated with bismuth based triple (n = 98), proton-pump inhibitor (PPI)-based triple (n = 180), or PPI based dual (n = 337) regimens. OUTCOME MEASURES: Retreatment; monthly postregimen medical expense, controlling for preregimen expense; and drug cost per successfully treated patient. Cox regression (retreatment analysis) and ANCOVA (postregimen expense analysis) adjusted for age, gender, diagnostic/procedural criteria met by patient, and specialty physician use. RESULTS: Retreatment rates were higher (p < 0.05) for PPI-based dual than bismuth-based or PPI-based triple therapy cohorts. Retreatment rates for bismuth- and PPI-based triple-therapy cohorts were not significantly different. Total and follow-up (excluding the first 2 wk of treatment) expenses were higher for retreated than nonretreated patients (p < 0.01). Total expenses were higher for the PPI-dual cohort (p < 0.05) than for triple cohorts. Drug costs per successfully treated patient were $30 for bismuth-based, $172 for PPI-based triple, and $208 for PPI-based dual therapy regimens. CONCLUSIONS: PPI-based dual-therapy regimens are not cost effective in H. pylori treatment. Further study should compare more costly (PPI based) versus less costly (bismuth-based) triple-therapy regimens. PMID- 10860133 TI - Potentiation of warfarin anticoagulation associated with topical methyl salicylate. AB - OBJECTIVE: To report a case of international normalized ratio (INR) elevation resulting from the administration of topical methyl salicylate in a patient whose INR was previously stable while she received warfarin anticoagulation. CASE SUMMARY: A 22-year-old white woman presented with an INR of 12.2 after applying a topical pain-relieving gel to her knees daily for eight days. The potentiation of the warfarin anticoagulation was attributed to the low-dose methyl salicylate contained in the product. DISCUSSION: Methyl salicylate is systemically absorbed through the skin in measurable amounts, and may increase warfarin action by affecting vitamin K metabolism or by displacing warfarin from protein-binding sites. While several investigators have reported this interaction with use of high-dose methyl salicylate, this case indicates that a significant interaction can occur with the use of lower topical doses of methyl salicylate as well. CONCLUSIONS: Healthcare providers and patients taking warfarin must be aware of the potential hazard of using topical methyl salicylate in combination with warfarin. PMID- 10860134 TI - Interaction between warfarin and trazodone. AB - BACKGROUND: It is well known that there are many drug interactions involving warfarin. However, few data have been supplied to guide clinicians concerning the interaction between trazodone and warfarin. CASE SUMMARY: Three clinically significant cases of suspected trazodone and warfarin interactions were identified in a retrospective chart review based on changes in the prothrombin time (PT) and international normalized ratio (INR) that were not explained by other factors. In each of the cases, the INR changed by > or = 1.0 after the initiation or discontinuation of trazodone. In the patients who started trazodone, a subsequent decrease in the PT and INR resulted; conversely, the PT and INR increased in the patient who stopped trazodone therapy. Although none of the patients experienced adverse effects due to the marked changes in PT and INR, the warfarin dosages had to be adjusted accordingly on initiation and discontinuation of trazodone. DISCUSSION: These cases show that there is a potentially clinically significant interaction between trazodone and warfarin. The time to onset of the interaction is variable; the mechanism behind it is not known, but it may involve substrate or protein-binding competition. CONCLUSIONS: The use of trazodone on an as-needed basis for sleep is strongly discouraged in patients who are receiving warfarin, due to the difficulty of achieving a therapeutic PT and INR. Until more is known, patients and clinicians should be educated about this potential interaction and monitor for changes in the anticoagulant effects when trazodone is initiated or stopped. PMID- 10860135 TI - Acquired factor VIII inhibitor in a patient with chronic myelogenous leukemia receiving interferon-alfa therapy. AB - OBJECTIVE: To report a case of an acquired factor VIII inhibitor associated with the use of interferon-alfa. CASE SUMMARY: A 58-year-old white man with newly diagnosed chronic myelogenous leukemia (CML) was initially treated with hydroxyurea. Interferon-alfa therapy was started six weeks later in order to enhance the response, with gradual reduction and eventual discontinuation of hydroxyurea. Interferon-alfa was continued for one year. Following bone marrow aspiration at one year, the patient developed significant bleeding and bruising at the site of extraction. His hemoglobin decreased from 11.3 to 9.3 g/dL and his activated partial thromboplastin time was elevated at 72 seconds. The factor VIII concentration was 0.02 units/mL; factor VIII inhibitor concentration was 58 Bethesda units. A diagnosis of an acquired factor VIII inhibitor was made, and the patient was treated with activated factor VII concentrates and prednisone. Interferon-alfa was discontinued, and the inhibitor subsequently disappeared over the next six weeks. The patient did not have any further bleeding problems. DISCUSSION: Acquired factor VIII inhibitors other than in patients with hemophilia are rare. To date, there are no reported cases of factor VIII inhibitors associated with CML. Moreover, the temporal association with interferon-alfa administration suggests a causal relationship. There are only two previous case reports suggesting interferon-alfa as a cause of factor VIII inhibitors. CONCLUSIONS: Induction of factor VIII inhibitors is a serious potential complication of therapy with interferon-alfa. We suggest that a diagnosis of an acquired factor VIII inhibitor be considered in patients who experience unexplained bleeding with interferon-alfa therapy. PMID- 10860136 TI - Zuclopenthixol-associated neutropenia and thrombocytopenia. AB - OBJECTIVE: To report a case of neutropenia and thrombocytopenia secondary to use of zuclopenthixol in a schizophrenic patient. CASE SUMMARY: A 66-year-old white man with chronic schizophrenia was referred to the hospital due to neutropenia and thrombocytopenia that developed shortly after initiation of zuclopenthixol therapy. Prior to zuclopenthixol administration, his white blood cell and platelet counts were 8.5 x 10(9) cells/L3 and 305 cells x 10(9)/L, respectively. Progressive reduction in leukocyte and platelet counts occurred, reaching a nadir of 2.9 x 10(9) cells/L3 (granulocytes 18.9%) and 109 cells x 10(9)/L, respectively. Zuclopenthixol was discontinued on admission, resulting in complete recovery within the next five days. DISCUSSION: Neutropenia and thrombocytopenia are well-known complications of antipsychotic drug therapy. Zuclopenthixol, a well-established antipsychotic agent, has relatively few adverse effects. The rapid decrease of white blood cell and platelet counts following the initiation of zuclopenthixol, as well as the rapid recovery, implicate zuclopenthixol as the predominant cause for neutropenia and thrombocytopenia in this patient. CONCLUSIONS: Although neutropenia and thrombocytopenia are rare complications of zuclopenthixol therapy, monitoring blood counts in patients receiving this agent seems to be justified. PMID- 10860137 TI - Treatment options for rheumatoid arthritis: celecoxib, leflunomide, etanercept, and infliximab. AB - OBJECTIVE: To review new pharmacologic agents approved for use in the management of rheumatoid arthritis (RA). DATA SOURCES: A MEDLINE search (1966-January 2000) was conducted to identify English-language literature available on the pharmacotherapy of RA, focusing on celecoxib, leflunomide, etanercept, and infliximab. These articles, relevant abstracts, and data provided by the manufacturers were used to collect pertinent data. STUDY SELECTION: All controlled and uncontrolled trials were reviewed. DATA EXTRACTION: Agents were reviewed with regard to mechanism of action, efficacy, drug interactions, pharmacokinetics, dosing, precautions/contraindications, adverse effects, and cost. DATA SYNTHESIS: Traditional pharmacologic treatments for RA have been limited by toxicity, loss of efficacy, or both. Increasing discoveries into the mechanisms of inflammation in RA have led to the development of new agents in hopes of addressing these limitations. With the development of celecoxib, a selective cyclooxygenase-2 inhibitor, the potential exists to minimize the gastrotoxicity associated with nonsteroidal antiinflammatory drugs. Leflunomide has been shown to be equal to or less efficacious than methotrexate, and may be beneficial as a second-line disease-modifying antirheumatic drug (DMARD). The biologic response modifiers, etanercept and infliximab, are alternatives that have shown benefit alone or in combination with methotrexate. However, they should be reserved for patients who fail to respond to DMARD therapy. Further studies should be conducted to evaluate the long-term safety and efficacy of these agents as well as their role in combination therapy. CONCLUSIONS: Celecoxib, leflunomide, etanercept, and infliximab are the newest agents approved for RA. Clinical trials have shown that these agents are beneficial in the treatment of RA; however, long-term safety and efficacy data are lacking. PMID- 10860138 TI - Citalopram in the treatment of depression. AB - OBJECTIVE: To review the efficacy and safety of citalopram in the treatment of depression. DATA SOURCES: MEDLINE search (1966-April 2000), Current Contents search, additional references listed in articles, and unpublished data obtained from the manufacturer were used to identify data from scientific literature. Studies evaluating citalopram (i.e., abstracts, clinical trials, data on file with the manufacturer) were considered for inclusion. STUDY SELECTION: English language literature was reviewed to evaluate the pharmacology, pharmacokinetics, therapeutic use, and adverse effects of citalopram. DATA EXTRACTION: Controlled animal and human clinical studies published in the English-language literature were reviewed and evaluated. Clinical trials selected for inclusion were limited to those in human subjects and included data from animals if human data were not available. DATA SYNTHESIS: Citalopram is an antidepressant belonging to the class of selective serotonin-reuptake inhibitors (SSRIs) available for the treatment of depression. Citalopram offers therapeutic efficacy similar to that of the other SSRIs and a more favorable adverse effect profile than that of the tricyclic antidepressants (TCAs). Citalopram does not cause anticholinergic or cardiovascular adverse effects associated with the TCAs. Citalopram is the most selective SSRI and, unlike other SSRIs, seems to be relatively free of interaction mediated by the cytochrome P450 system. Citalopram is also the least expensive antidepressant available to date. This review of citalopram includes data from clinical trials comparing safety, tolerability, efficacy, and pharmacoeconomics with TCAs and SSRIs. CONCLUSIONS: Clinical trials demonstrate that citalopram's therapeutic efficacy is significantly greater than that of placebo and is comparable with that of other antidepressants. Citalopram has a favorable adverse effect profile, and thus may be useful in treating depressed patients who cannot tolerate anticholinergic or cardiovascular adverse effects associated with TCAs. It may also be useful in patients with comorbid illnesses requiring concomitant medicines. PMID- 10860139 TI - Ambulatory care pharmacy services: has the agenda changed? AB - OBJECTIVE: To provide an extensive review of ambulatory care clinical pharmacy services and evaluate the services and research data in the field. DATA SOURCES: MEDLINE was searched from January 1992 through July 1999. Search terms included pharmacy, clinical pharmacy, and pharmaceutical care, cross-referenced with ambulatory care, primary care, family medicine, and managed care. STUDY SELECTION: Relevant peer-reviewed studies and reports since our previous article in 1992 were selected and described. Literature prior to 1992 was briefly reviewed. DATA SYNTHESIS: The relevant literature was reviewed and some examples from the authors' institutions are provided. Much research has continued to be published documenting the value of clinical pharmacy services in ambulatory care, including in community pharmacy, anticoagulation services, family medicine, primary care clinics, Veterans Affairs Medical Centers, and managed care. However, these innovative services are underrepresented in the community at large. The vast majority of the public does not have access to these types of services. CONCLUSIONS: There will be continued and dramatic expansion of ambulatory care pharmacy services in the new decade beginning in the year 2000. It will be critical that standards of practice be very high. We believe there is a critical need for visible demonstration projects and large multicenter research projects that demonstrate the value of these services. PMID- 10860140 TI - Indirect modulation of dopamine D2 receptors as potential pharmacotherapy for schizophrenia: II. Glutamate (Ant)agonists. AB - OBJECTIVE: To summarize the published preclinical and clinical data that suggest the possible use of glutamate receptor agonists or antagonists as novel antipsychotic agents. DATA SOURCES: Primary and review articles were identified by MEDLINE search (from 1966 to December 1999) and through secondary sources. STUDY SELECTION AND DATA EXTRACTION: All of the articles identified from the data sources were evaluated and all information deemed relevant was included. DATA SYNTHESIS: The standard antipsychotic drugs, whose clinical activity correlates with affinity for dopamine D2 receptors, alleviate some of the positive symptoms of schizophrenia, but have limited impact on negative symptoms. Several lines of evidence implicate glutamate-receptor system dysfunction(s) in schizophrenia, either as causative or contributory factors. In addition, several standard antipsychotic drugs modulate glutamate or glutamate receptor activity, suggesting an alternative view of their mechanism of antipsychotic action. Preliminary studies have shown that drugs which modulate glutamate brain concentrations have positive effects in animal models of schizophrenia. CONCLUSIONS: A role for glutamate in the pathogenesis or pharmacotherapy of schizophrenia is suggested from anatomic (interactions between glutamatergic and dopaminergic systems in relevant brain regions), physiologic (implication of glutamate-receptor dysfunction), and pharmacologic (modulation of glutamate or glutamate receptors) evidence. Therefore, compounds that function at glutamate receptors might represent a novel approach to the treatment of the disease or to the amelioration of symptoms, either as monotherapy or as an adjunct to dopamine D2 receptor antagonists. PMID- 10860141 TI - Zanamivir in the treatment and prevention of influenza. AB - OBJECTIVE: To evaluate the efficacy of zanamivir in the prevention and treatment of influenza. DATA SOURCES: Medical literature was accessed through MEDLINE (1966 June 1999). Key search terms included zanamivir, GG167, and influenza. Additional information was obtained from GlaxoWellcome, Inc. DATA SYNTHESIS: Zanamivir is the first in a new class of drugs to be developed for the treatment of influenza. An evaluation of clinical trials using inhaled zanamivir was conducted to determine its efficacy. CONCLUSIONS: Zanamivir appears to shorten the median duration of influenza symptoms by up to 2.5 days when compared with placebo. It was well tolerated in clinical trials, with mild adverse effects occurring in a small percentage of subjects. PMID- 10860142 TI - Use of gabapentin in the treatment of neuropathic pain. AB - OBJECTIVE: To evaluate the role of gabapentin for the treatment of neuropathic pain. DATA SOURCES: Clinical literature was identified through MEDLINE (from 1990 to October 1999). Key search terms were gabapentin and pain. DATA SYNTHESIS: Neuropathic pain can be a problematic, chronic syndrome that is frequently refractory to current drug treatments. Gabapentin is a newer generation antiepileptic drug that is commonly used in treatment of neuropathic pain. An evaluation of clinical trials using gabapentin to treat neuropathic pain was performed. CONCLUSIONS: Gabapentin appears to be effective in treating various neuropathic pain disorders. Gabapentin may have advantages over current therapies, such as a favorable safety profile and lack of drug interactions; however, cost issues and limited experience may limit the use of gabapentin as a first-line option. PMID- 10860143 TI - Chest and back pain associated with a six-hour infusion of liposomal daunorubicin. PMID- 10860144 TI - Polyethylene vials of calcium gluconate reduce aluminum contamination of TPN. PMID- 10860145 TI - Aseptic meningitis after iohexol myelography. PMID- 10860146 TI - Fluorouracil cream: don't forget to wash your hands. PMID- 10860147 TI - WA joins quest for legal ban on tail docking. PMID- 10860148 TI - Dockers' advocate has a really bad day! PMID- 10860149 TI - Charles Wright caught on wrong foot. PMID- 10860150 TI - Pulmonary cryptococcosis and Capillaria aerophila infection in an FIV-positive cat. AB - A 12-year-old, FIV-positive, domestic longhair cat was presented with a history of sneezing and coughing during the previous seven months. On thoracic radiographs, a prominent bronchial pattern and three focal, opacified nodules were seen. Cytology of bronchoalveolar lavage fluid demonstrated spherical, capsulate, narrow-necked, budding yeasts within macrophages. Culture of the fluid yielded a heavy growth of Cryptococcus neoformans var neoformans. The serum latex cryptococcal antigen agglutination test titre was 158. The cat was treated with itraconazole and the cough resolved over a 5-month period but then recurred. Repeat thoracic radiographs showed resolution of the pulmonary nodules but a persistent bronchial pattern. Adult nematodes and ova with morphology characteristic of Capillaria aerophila were seen in bronchoalveolar lavage fluid and no yeasts were cultured from the fluid. The cryptococcal titre was zero. The lungworm infection was treated successfully with abamectin and the cough resolved. Immunosuppression related to FIV infection may have predisposed this cat to sequential respiratory tract infections. PMID- 10860151 TI - Successful treatment of invasive nasal cryptococcosis in a ferret. AB - Invasive cryptococcal rhinitis due to Cryptococcus neoformans var gattii was diagnosed in a castrated, 5-year-old, albino ferret with subcutaneous swelling of the nasal bridge. The diagnosis was based on histology, needle aspirate cytology and positive culture on Sabouraud's dextrose agar and birdseed agar. The ferret was successfully treated using a long course of itraconazole (25 to 33 mg orally once daily with food) subsequent to surgical debulking of the lesion, using sequential cryptococcal antigen titre determinations to guide therapy. PMID- 10860152 TI - Dermoid cyst in the tongue of a dog. AB - A German Shepherd Dog presented with recurrent intermandibular and intralingual swelling. Cytological and microbiological findings on fluid aspirated from the mass were consistent with an infected cyst lined by epithelium. The sinus was explored and an epithelium-lined cystic structure was extirpated from the frenulum and body of the tongue. This structure was diagnosed histologically as a dermoid cyst. PMID- 10860153 TI - Juvenile nephropathy in a boxer, a rottweiler, a collie and an Irish wolfhound. AB - Juvenile nephropathy was diagnosed in a Boxer, a Rottweiler, a Collie and an Irish Wolfhound dog, each presenting with signs compatible with chronic renal failure. The diagnosis in each case was based on the presence of persistence of poorly differentiated tissue (immature glomeruli and/or tubules, persistent mesenchyme) on histopathologic examination. Although juvenile nephropathy has been reported in many breeds of dog, this is the first report of the condition in the Collie and the Irish Wolfhound and only the second description in the Boxer and the Rottweiler. The possibility of an inherited origin of the condition in these four breeds is at present unknown. PMID- 10860154 TI - Perirenal pseudocysts in 26 cats. AB - OBJECTIVE: To evaluate clinical features, anatomical location, nature of pseudocyst fluid, results of surgical treatment and links with underlying renal disease in cats with perirenal pseudocysts. DESIGN: A retrospective study of 26 affected cats, including 8 treated surgically. RESULTS: Nineteen (73%) affected cats were male. The median age was 11 years. Most presented for abdominal enlargement and had varying degrees of renal dysfunction on presentation. Thirteen cats (50%) had bilateral pseudocysts. The pseudocyst fluid was a transudate or modified transudate in all cases. All surgically treated cats had subcapsular perirenal pseudocysts. Associated renal lesions were identified in all cats that had renal biopsies or detailed ultrasonographic examinations. Surgery relieved clinical signs but did not stop progression of renal disease. Cats survived a median of 9 months after surgery and survival was correlated statistically to degree of azotaemia at presentation. Percutaneous drainage of pseudocysts was ineffective in controlling long-term fluid accumulation. CONCLUSIONS: Subcapsular perirenal pseudocysts are formed in cats by accumulation of transudate between the capsule and parenchyma of the kidney as a result of underlying parenchymal disease. Pseudocyst formation can occur at variable stages of renal dysfunction. Resection of the pseudocyst wall is usually effective in eliminating signs but does not stop progression of renal disease. The prognosis for cats with pseudocyst formation is related to the degree of renal dysfunction at time of diagnosis. PMID- 10860155 TI - Establishment, validation and use of the Kielstein-Rapp-Gabrielson serotyping scheme for Haemophilus parasuis. AB - OBJECTIVES: To produce antisera to the 15 recognised reference strains of the Kielstein-Rapp-Gabrielson (KRG) serotyping scheme for Haemophilus parasuis, validate those sera and use them to serotype 46 Australian field isolates of H parasuis. DESIGN: Antisera were produced in rabbits and validated by cross testing with the reference strains and re-testing 15 Australian field isolates of H parasuis that had been previously serotyped in the United States of America. The validated antisera were then used to determine the serovar of 46 Australian isolates. RESULTS: Monospecific antisera were produced for 14 of the 15 KRG serovars of H parasuis. Two Australian field isolates, confirmed previously as serovars 1 and 7, were used to produce monospecific antisera for serovars 1 and 7 respectively. The antiserum for serovar 4 gave a one-way cross reaction with the antigen of serovar 14. The typing antisera correctly typed all 15 H parasuis that had been previously typed by antisera produced overseas. The 46 field isolates were shown to belong to serovars 2 (two isolates), 4 (one isolate), 5 (18 isolates), 12 (two isolates) and 13 (four isolates). The remaining 19 isolates were non-typable. CONCLUSION: Serotyping of H parasuis isolates is now available in Australia. H parasuis serovars 5 and 13 remain the predominant serovars present in Australian pigs. PMID- 10860156 TI - Novel Propionibacterium infection in cattle. AB - OBJECTIVE: To describe four cases of infection in cattle, from geographically different places, with a presumptive new species of Propionibacterium, which causes granulomatous lesions in the head, thorax, abdomen, pelvic area and skin. PROCEDURE: Gross lesions, ranging from 0.5 to 15 cm and detected during routine carcase inspection at the abattoir, were submitted to the laboratory for routine testing in the National Granuloma Submission Program. The bacterium isolated was identified using morphological characteristics, biochemical reactions, cell wall components, products of fermentation and 16S rRNA gene sequencing. RESULTS: Gross lesions submitted for examination consisted of a fibrous outer capsule enclosing thick yellow pus-like material. A Gram-Glynn stain of the histological sections revealed colonies of Gram-positive, filamentous, branching bacteria. Bacteriological culture, cell wall analysis, biochemical reactions and 16S rRNA sequencing identified the organism as a Propionibacterium sp closely related to P cyclohexanicum and the P freudenreichii cluster. CONCLUSION: This is the first report of a Propionibacterium sp closely related to P cyclohexanicum and the P freudenreichii cluster associated with extensive granulomatous lesions in cattle in Queensland. Sequencing data are suggestive of a previously undescribed species of the Propionibacterium genus. PMID- 10860157 TI - Application of PCR assays to determine the genotype of Babesia bovis parasites isolated from cattle with clinical babesiosis soon after vaccination against tick fever. AB - OBJECTIVE: To demonstrate the value of PCR assays to determine the genotypes of Babesia bovis in cattle with clinical signs of babesiosis within 3 weeks after vaccination against tick fever. DESIGN: Samples from 5 cases of babesiosis in cattle soon after vaccination against tick fever were analysed in two PCR assays. PROCEDURE: Parasite DNA was purified from blood taken from cattle with signs of babesiosis within 3 weeks of vaccination against tick fever. DNA was also prepared from the tissues of animals that died of babesiosis. Two PCR assays that amplify repeat sequences of DNA within the B bovis genes, Bv80 and BvVA1, were used to differentiate the genotypes of field isolates and vaccine strains of B bovis. RESULTS: One of the five cases of babesiosis was found to be caused by a vaccine strain, but PCR analyses showed that the predominant isolate in the other four cases was not the vaccine strain. CONCLUSIONS: PCR assays on the DNA of B bovis obtained from the blood or tissues of cattle clinically affected with tick fever within 3 weeks after vaccination are useful to distinguish between vaccine strains and field isolates as the source of infection. PMID- 10860158 TI - Cryptosporidium meleagridis in an Indian ring-necked parrot (Psittacula krameri). AB - OBJECTIVE: To perform a morphological and genetic characterisation of a Cryptosporidium infection in an Indian ring-necked parrot (Psittacula krameri) and to compare this with C meleagridis from a turkey. DESIGN: Tissue and intestinal sections from an Indian ring-necked parrot were examined microscopically for Cryptosporidium. The organism was also purified from the crop and intestine, the DNA extracted and a portion of the 18S rDNA gene amplified, sequenced and compared with sequence and biological information obtained for C meleagridis from a turkey as well as sequence information for other species of Cryptosporidium. RESULTS: Morphological examination of tissue sections from an Indian ring-necked parrot revealed large numbers of Cryptosporidium oocysts attached to the apical border of enterocytes lining the intestinal tract. Purified Cryptosporidium oocysts measured about 5.1 x 4.5 microns, which conformed morphologically to C meleagridis. The sequence obtained from this isolate was identical to sequence information obtained from a C meleagridis isolate from a turkey. CONCLUSION: Cryptosporidium meleagridis was detected in an Indian ring-necked parrot using morphological and molecular methods. This is the first time that this species of Cryptosporidium has been reported in a non galliform host and extends the known host range of C meleagridis. PMID- 10860159 TI - DNA fingerprinting of Australian isolates of Mycobacterium avium subsp paratuberculosis using IS900 RFLP. AB - OBJECTIVES: To evaluate additional restriction enzymes for IS900 RFLP of Mycobacterium avium subsp paratuberculosis and examine the genetic diversity among Australian isolates for epidemiological studies of Johne's disease. DESIGN AND PROCEDURE: Seventy-one isolates of M paratuberculosis from cattle, sheep, goat, alpaca and rhinoceros in six Australian States and the Northern Territory, reference strains and reference DNA from previously characterised strains were tested for genetic variation. Bst EII, Pvu II and Pst I restriction enzymes were used, and four others (Bam HI, Alu I, Xho I and Dra I) were assessed for their ability to detect polymorphisms. Multiple isolates from some animals were tested. RESULTS: Bam HI, was the most effective enzyme for identifying polymorphisms (12 types), followed by Bst EII (11 types). Both Pvu II and Pst I were relatively ineffectual. Fifteen different types were identified, 12 in clinical isolates. Most isolates were cattle (C) strains and fell into the C1 (n = 28) and C3 (n = 32) groupings. All isolates from alpaca were type C1, and bovine isolates were commonly C1 (n = 15) or C3 (n = 28). All of the sheep were infected with sheep (S) strains; no S strains were identified in cattle. Two of six isolates from one animal had single band differences. CONCLUSION: The epidemiological features of M paratuberculosis in Australia are similar to those reported in New Zealand, where cattle and sheep are commonly infected with different strains. However, because of the lack of polymorphism identified within the major groups, it is unlikely that DNA fingerprinting will have a significant role in epidemiological studies of Johne's disease, unless an unusual strain in being studied. PMID- 10860160 TI - Langhans' giant cells in the gills of an Atlantic salmon. PMID- 10860161 TI - Pathology of melioidosis in captive marine mammals. PMID- 10860162 TI - Piglets born from vitrified early blastocysts using a simple technique. PMID- 10860163 TI - Sell kangaroo meat. PMID- 10860164 TI - Pups' tails saved. PMID- 10860165 TI - 'Strange' US idea. PMID- 10860166 TI - The Minnesota Multiphasic Personality Inventory and chronic pain: a conceptual analysis of a long-standing but complicated relationship. AB - The Minnesota Multiphasic Personality Disorder (MMPI) and its successor, the MMPI 2, have a long-standing tradition in the assessment of patients with chronic pain. With the introduction of more narrowly defined and factor-analyzed pain inventories, however, the utility of the MMPI-2 for pain assessment has been brought into question. In this review, the relevant literature is carefully scrutinized from a conceptual and historical perspective. It is concluded that many of the (recent) criticisms are largely ungrounded. Rather than the test itself being at fault or of little utility in the field of pain assessment, it has simply been applied inappropriately (i.e., for determination of pain etiology or underlying personality structure "explaining" the chronic pain). In conclusion, it is suggested that the application of the MMPI-2 in the assessment of patients with chronic pain should correspond more closely to the original aims and psychometric properties of the tool--that is, for screening and the generation of hypotheses regarding comorbid psychopathology and personality features having the potential to complicate the treatment process. Guidelines for clinical interpretation of MMPI-2 profiles with regard to chronic pain are provided. PMID- 10860167 TI - Maternal depression and parenting behavior: a meta-analytic review. AB - The results of 46 observational studies were analyzed to assess the strength of the association between depression and parenting behavior and to identify variables that moderated the effects. The association between depression and parenting was manifest most strongly for negative maternal behavior and was evident to a somewhat lesser degree in disengagement from the child. The association between depression and positive maternal behavior was relatively weak, albeit significant. Effects for negative maternal behavior were moderated by timing of the depression: Current depression was associated with the largest effects. However, residual effects of prior depression were apparent for all behaviors. Socioeconomic status, child age, and methodological variables moderated the effects for positive behavior: Effects were strongest for studies of disadvantaged women and mothers of infants. Studies using diagnostic interviews and self-report measures yielded similar effects, suggesting that deficits are not specific to depressive disorder. Research is needed to identify factors that affect the magnitude of parenting deficits among women who are experiencing depression and other psychological difficulties. PMID- 10860168 TI - A modern behavioral perspective on child conduct disorder: integrating behavioral momentum and matching theory. AB - It has been suggested that knowledge produced within the operant laboratory is of little or no use to clinicians. I argue, on the contrary, that laboratory science has provided clinicians with two general principles that may expand the focus of behavioral family therapy to incorporate a wide range of clinical interventions that have heretofore been considered nonbehavioral. These principles, matching theory and behavioral momentum, outline the relativity of reinforcement and the persistence of behavior in the absence of reinforcement, respectively. These principles make specific predictions concerning clinical interventions aimed not only at identified reinforcement contingencies, but also the context within which reinforcement contingencies are operative. This expanded behavioral formulation allows both the clinician and the researcher a framework for designing, implementing, and assessing techniques that target cognition, affect, and interpersonal relationships, as well as specific behaviors. PMID- 10860169 TI - The relationship between eating disorders and substance use: moving beyond co prevalence research. AB - The frequent comorbidity of eating disorders and substance use has been demonstrated consistently by research. Less is known about the basis of this relationship. A review of the literature indicates that the hypotheses proposed to clarify the etiological relationship between eating disorders and substance use have not been supported sufficiently or consistently by empirical evidence. General criticisms include: a lack of well developed models, a reliance on co prevalence data, and a lack of integration of knowledge from eating disorder and substance use research. It is suggested that an understanding of the etiological relationship between eating disorders and substance use will be arrived at only after fundamental inquiries into the functional relationship between eating disorders symptomatology and substance use patterns have been conducted. A behavioral assessment approach is offered as a means of evaluating the functional relationship between eating disorder symptomatology and substance use. PMID- 10860170 TI - A review of expressed emotion research in health care. AB - Much research has been carried out on the impact of family relationships on the development and course of different illness. Research on Expressed Emotion (EE) developed out of studies of the impact of family members on patients with schizophrenia, and has provided us with a robust measure of relatives' emotional attitudes towards patients, which has now been applied in the study of numerous psychiatric and medical illnesses. This review outlines the history of EE research in schizophrenia, and discusses the evidence for the association between family EE and the course of schizophrenic illness. Some of the factors which might moderate the association between EE and illness course are outlined and the issues of the meaning and development of EE are discussed in the light of recent theoretical advances. The application of the EE methodology in other psychiatric and medical conditions is then reviewed and conclusions are drawn about the extent to which EE predicts illness course in conditions other than schizophrenia. Consideration is given to the ways in which the application of the paradigm to a variety of illnesses or conditions with different features can enhance our understanding of the EE construct. PMID- 10860171 TI - Psychiatric comorbidity in white and African-American illicit substance abusers: evidence for differential etiology. AB - Research on psychiatric comorbidity among opiate and cocaine addicts has consistently found African-Americans to report fewer symptoms of anxiety and affective disorders than Whites. The current article reviews the research on these racial differences, evaluates various interpretations of these differences, and discusses the limitations of past research. It is concluded that Whites and African-American addicts differ in their underlying reasons for abusing drugs. Drug addiction among Whites appears to be related largely to psychopathology, whereas Black drug abuse is best understood in terms of social and environmental factors. Treatment implications are also discussed. PMID- 10860172 TI - Thrombolytic therapy for acute myocardial infarction in the elderly. PMID- 10860173 TI - Primary angioplasty for acute myocardial infarction in the elderly. AB - Elderly patients with acute myocardial infarction present a formidable therapeutic challenge. Although there appears to be a survival benefit from thrombolytic therapy for the eligible elderly patient, persistent concerns regarding the risk of intracranial hemorrhage impedes utilization in this age group. Primary or direct angioplasty of the infarct artery has been proven to be an effective modality for reperfusion. Randomized comparisons suggest an advantage over thrombolysis in terms of achieving superior patency and mitigating recurrent ischemic events. Primary angioplasty expands the reperfusion population by including many patients ineligible for thrombolysis and is more effective for treating patients at high risk, such as those with cardiogenic shock. Acute angiography accumulates important prognostic and decision-facilitating information. The benefits of primary angioplasty are more impressive for the aging patient. The survival gain and reduction in intracranial hemorrhage may combine to magnify the advantages of performing angioplasty on patients in this group. Emerging evidence concerning the aging population validates continued examination of this invasive reperfusion approach. PMID- 10860174 TI - Percutaneous revascularization for unstable angina in the elderly. PMID- 10860175 TI - Anti-thrombotic therapy for elderly patients with acute coronary syndromes. PMID- 10860176 TI - Beta-blockers, angiotensin-converting enzyme inhibitors, and calcium antagonists in treatment of elderly patients with acute myocardial infarction. AB - Administration of beta-blockers reduces mortality among old persons during and after acute myocardial infarction. The American College of Cardiology/American Heart Association guidelines recommend that persons without contraindications to use of beta-blockers should be administered beta-blockers within a few days of myocardial infarction (if administration is not initiated acutely) and that their administration should be continued indefinitely. These guidelines also recommend the use of angiotensin converting enzyme inhibitors in treating persons within the first 24 h of suspected onset of acute myocardial infarction with ST-segment elevation in two or more anterior precordial leads or with congestive heart failure in the absence of significant hypotension or other contraindications to use of ACE inhibitors; and persons during and after convalescence from acute myocardial infarction with congestive heart failure associated with an abnormal left ventricular ejection fraction (LVEF) or with asymptomatic left ventricular systolic dysfunction with a LVEF < 40%. These guidelines state that there are no class I indications for using calcium antagonists after myocardial infarction. If patients have persistent angina pectoris after myocardial infarction despite treatment with beta-blockers and nitrates or hypertension inadequately controlled by other drugs, administration of a nondihydropyridine calcium antagonist such as verapamil or diltiazem should be added to the therapeutic regimen if the LVEF is normal. If the LVEF is abnormal, administration of amlodipine or felodipine should be added to the therapeutic regimen. PMID- 10860177 TI - Mortality, mode of death and risk indicators for death during 5 years after coronary artery bypass grafting among patients with and without a history of diabetes mellitus. AB - OBJECTIVE: To describe mortality, mode of death, risk indicators for death and symptoms of angina pectoris among survivors during 5 years after coronary artery bypass grafting (CABG) among patients with and without a history of diabetes mellitus. METHODS: All patients in western Sweden who underwent CABG without concomitant valve surgery and who had no previous CABG between June 1988 and June 1991 were entered prospectively in this study. After 5 years, information on deaths that had occurred was obtained for the analysis. RESULTS: In all, 1998 patients were included in the analysis; 242 (12%) had a history of diabetes. Among the non-diabetic patients, 5-year mortality was 12.5%; the corresponding relative risk for diabetic patients was 2.1 (95% confidence interval 1.6 to 2.9). A history of diabetes was an independent risk indicator of death; there was no significant interaction between any other risk indicator and diabetes. Independent risk indicators for death among diabetic patients were: current smoking, renal dysfunction and left ventricular ejection fraction < 0.40. Compared with non-diabetic patients, those with diabetes more frequently died in hospital, died a cardiac death, or had death associated with the development of acute myocardial infarction and with symptoms of congestive heart failure. Among survivors, diabetic patients tended to have more angina pectoris 5 years after CABG than did those without diabetes. CONCLUSION: During a period of 5 years after CABG, diabetic patients had a mortality twice that of non-diabetic patients. The increased risk included death in hospital, cardiac death and death associated with development of acute myocardial infarction and with symptoms of congestive heart failure. PMID- 10860178 TI - Effects of valsartan and 17 beta-estradiol on the oxidation of low-density lipoprotein in vitro. AB - BACKGROUND: The 'sartans' are antagonists of the angiotensin type 1 (AT1) receptor that are mainly used for treatment of hypertension. Little is known about AT1-independent effects of these substances and interactions with other drugs used for prevention of cardiovascular diseases. Postmenopausal estradiol replacement therapy has been shown to exert beneficial antiathero-sclerotic properties by inhibiting oxidation of low-density lipoprotein (LDL). OBJECTIVE: In the present study, the effects of valsartan alone and in combination with 17 beta-estradiol on the oxidation of isolated human LDL were investigated. METHODS: Oxidation of LDL, which was triggered by copper (II) chloride, was monitored spectrometrically at 234 nm. The test substances were added in vitro. RESULTS: Valsartan alone increased the duration of resistance of LDL to oxidation by 75.3 +/- 5.7 min at 5 mumol/l and by 138.2 +/- 8.1 min at 10 mumol/l. 17 beta estradiol alone delayed the onset of oxidation of LDL by 75.7 +/- 5.1 min at 1 mumol/l. With the combination of 5 and 10 mumol/l valsartan with 1 mumol/l estradiol the time to onset of oxidation of LDL was increased by 142.8 +/- 4.9 and 215.3 +/- 6.9 min, respectively. CONCLUSIONS: There has been demonstrated an antioxidative effect of valsartan that was additive to that of 17 beta-estradiol. Thus this combination has the potential to be useful in the treatment of postmenopausal women with hypertension. PMID- 10860179 TI - Use of nitric-oxide-eluting polymer-coated coronary stents for prevention of restenosis in pigs. AB - BACKGROUND: Restenosis after angioplasty remains an unresolved problem despite an increase in use of coronary stents. It has been theorized that nitric oxide (NO) exerts several actions that can prevent restenosis. These include inhibition of proliferation of smooth muscle cells, prevention of arterial spasms, and decreasing aggregation of platelets in response to exposure to collagen. OBJECTIVE: To determine whether NO coated stents decrease restenosis in a pig balloon injury model. METHODS: We used coronary stents impregnated with a slow release precursor of NO in the porcine model of restenosis. Tantalum coil coronary stents (Cordis) were coated with a polymer impregnated with a slow release precursor of NO. Polymer-coated stents without active precursors were used as controls. Oversized stents were mounted on a delivery balloon and subsequently deployed in the right coronary and left anterior descending arteries of each animal. RESULTS: Repeated recording of angiograms demonstrated that changes in minimum lumen diameter on going from immediately after stenting to 28 day follow-up for the control and NO-eluting-stent groups were similar, namely decreases of 1.89 +/- 0.33 and 2.08 +/- 0.28 mm, respectively. The morphometric results, showing that severe luminal narrowing occurred for both groups, were similar. The percentage area stenoses were 85 +/- 5% for the control group and 84 +/- 6% for the NO-eluting group. Histology demonstrated that profuse formation of neointima and an inflammatory cell infiltrate occurred. CONCLUSIONS: Severe diameter stenosis occurred both for control and for treatment groups. The degree of angiographic stenosis was markedly worse than that previously reported for this model. Sustained release of a precursor of NO did not prevent restenosis in this model. This might have been due to a lack of efficacy of nitric oxide or to a profuse and overwhelming stimulatory effect of the polymer in the coated stents. PMID- 10860180 TI - Immediate effect of Biosense guided percutaneous direct myocardial revascularization with holmium:yttrium aluminium garnet laser on myocardial contractility assessed by transesophageal echocardiography. AB - OBJECTIVE: To evaluate immediate changes in left ventricular wall motion in patients treated using Biosense direct myocardial revascularization laser system. METHODS: Regional wall motion in 10 patients undergoing catheter-based direct myocardial revascularization using a holmium:yttrium aluminium garnet laser was assessed by transesophageal echocardiography before and immediately after the procedure. RESULTS: Mild deterioration in wall-motion score occurred rarely for only three of 160 (1.9%) segments and did not induce clinical heart failure. CONCLUSION: With the current catheter-based laser myocardial revascularization strategy, mild deterioration in wall motion of treated segments was rarely observed and did not effect overall left ventricular function or induce clinical congestive heart failure. PMID- 10860181 TI - Impact of exercise-induced coronary vasomotion on anti-ischemic therapy. AB - Coronary vasomotion has an important role in the regulation of myocardial perfusion. During dynamic exercise, normal coronary arteries dilate, whereas stenotic arteries constrict. This exercise-induced vasoconstriction has been associated with the occurrence of myocardial ischemia and has been believed to be the result of endothelial dysfunction, with a reduced release or production of EDRF, increased sympathetic stimulation, enhanced platelet aggregation with release of thromboxane A2 and serotonin, or a passive collapse of the disease free wall segment within the stenosis (the Bernoulli effect), or a combination of any of these. More recently, it has been realized that pharmacological treatment might prevent exercise-induced vasoconstriction and, thus, reduce myocardial ischemia and the occurrence of angina pectoris. Vasodilators such as nitrates, calcium antagonists or alpha-receptor blockers dilate the coronary arteries and prevent coronary stenosis narrowing during exercise. In contrast, beta-blocking agents are associated with coronary vasoconstriction at rest, but--conversely- can induce coronary vasodilatation during exercise. Pharmacological treatment in patients with stable angina pectoris may improve myocardial ischemia by reducing pre- and afterload, myocardial contractility, oxygen consumption, and vasomotor tone. However, coronary collateral perfusion can modify these effects by shunting blood from the non-ischemic to the ischemic region (collateral flow) or by shunting blood from the ischemic to the non-ischemic zone (coronary steal phenomenon). Typically, a steal phenomenon has been reported in patients receiving either dipyridamole or calcium antagonists, whereas a reversed steal has been described after beta-blockade, with an increase in contralateral tone shunting blood from the non-ischemic to the ischemic zone (reverse steal phenomenon). PMID- 10860182 TI - Bibliography. Current world literature. PMID- 10860183 TI - The changing lifestyle in the world. Body weight and what else? AB - Body weight and the prevalence of obesity are rising so rapidly in many countries that the World Health Organization has recognized that there is a "global epidemic of obesity." The prevalence of type 2 diabetes is rising in parallel. In view of its associated cardiovascular complications, we are facing a severe public health problem. Both obesity and type 2 diabetes have a combined genetic and environmental background, but the epidemic must be due to major changes in the environment. By definition, obesity is a result of a positive energy balance, which usually amounts to a tiny proportion of the total energy turnover. Energy intake, energy expenditure, and energy accumulation (as fat) may all be primarily disturbed. There is a great, and still insufficiently understood, variation in prevalence of obesity and in the rate of change of the prevalence. The prevailing contention is that the epidemic is due to the changes in the society--the so called modernization--leading to overnutrition and a sedentary life. These factors are likely contributors, but it has been difficult to provide consistent evidence for their effects. In Denmark, a steep rise has taken place in the prevalence of obesity among schoolboys and young men in two phases linked to the birth cohorts of the 1940s and of the mid-1960s and later. This rise suggests that environmental influences operating early in life are involved. In conclusion, a global obesity epidemic is developing, but the causes of the epidemic are not yet clear and more research is needed to establish the grounds for prevention. PMID- 10860184 TI - Type 2 diabetes worldwide according to the new classification and criteria. AB - Two major reports have recently revised the classification of and diagnostic criteria for diabetes. Classification was previously based on the need for insulin (insulin-dependent or non-insulin-dependent), but this has become increasingly confusing. Now, the type of diabetes is determined by the etiological process rather than the treatment modality. Type 1 diabetes is thus characterized by islet cell destruction and type 2 diabetes by a combination of defects in insulin secretion and action. An individual with either type of diabetes may be on any treatment modality. This classification should prove to be more logical and, for example, allow latent autoimmune diabetes in adults, which typically does not require insulin at presentation, to be classified as type 1 diabetes. The fasting plasma glucose diagnostic threshold for diabetes has been lowered to 7.0 mmol/l (126 mg/dl), and impaired fasting glucose (fasting plasma glucose 6.1-6.9 mmol/l [110-125 mg/dl]) has been introduced as a new category of intermediate glucose metabolism. These changes recognize that the old fasting threshold did not match the 2-h (postload) threshold well and that both micro- and macrovascular disease develop at lower fasting glucose levels than previously recognized. Although the prevalences of diabetes according to the new fasting and 2-h criteria are now similar in most populations, the actual individuals identified as having diabetes are often different. Over 30% of all those with diabetes have a nondiabetic fasting glucose but still have increased cardiovascular mortality. Thus, it is important to retain the oral glucose tolerance test for the diagnosis of diabetes. PMID- 10860185 TI - Background and recruitment data for the U.S. Diabetes Prevention Program. AB - The objective of the Diabetes Prevention Program (DPP) is to prevent or delay the development of type 2 diabetes in those high-risk individuals who have tested positive for impaired glucose tolerance on an oral glucose tolerance test. The DPP is a multicenter randomized clinical trial in the U.S. with three intervention arms--lifestyle, metformin, and placebo--with 1,000 participants in each arm who will be recruited over a 3-year recruitment period and will be followed for 3 years after the study-wide closing date for recruitment, resulting in a 3- to 6-year participant follow-up interval. The primary outcome is the development of diabetes according to the revised American Diabetes Association criteria, confirmed with a repeat test. Recruitment ended in the spring of 1999. As of 14 October 1998, the DPP had screened 133,683 individuals, of whom 26,518 had an oral glucose tolerance test, resulting in 3,048 randomized participants (585 of who were former troglitazone arm participants). Of the randomized participants, approximately 45% belong to an ethnic minority group, 67% are women, and 10% are > or = 65 years old. In conclusion, recruitment of subjects for the DPP has been highly successful, particularly with respect to recruitment of minority participants. PMID- 10860186 TI - Course of glomerular filtration rate in albuminuric type 2 diabetic patients with or without diabetic glomerulopathy. AB - OBJECTIVE: To evaluate and compare the clinical course and prognosis in type 2 diabetic patients with persistent albuminuria, with biopsy-proven diabetic glomerulosclerosis (DG), or with nondiabetic glomerulopathies (NDG). RESEARCH DESIGN AND METHODS: A kidney biopsy was performed in 34 consecutive type 2 diabetic patients with persistent albuminuria (> or = 300 mg/24 h). Glomerular filtration rate (GFR) (51Cr-EDTA) was determined at least once a year, and albuminuria, arterial blood pressure, and HbA1c were determined every 3-6 months. RESULTS: The biopsy revealed DG in 26 patients (25 men/1 woman) (DG group), age 52 +/- 2 (mean +/- SEM) years, and NDG in 8 patients (7 men/1 woman) (NDG group), age 54 +/- 3 years. The patients were followed for a median of 7.7 years (range 1.0-14.2). In the DG group, GFR decreased from 82 (24-146) to 38 (2-116) ml.min 1.1.73 m-2 (P < 0.001), with a median rate of decline in GFR of 5.6 (0.3-21.6) ml.min-1.year-1, and in the NDG group, GFR decreased from 107 (89-135) to 90 (17 119) ml.min-1.1.73 m-2 (P < 0.05), with a median rate of decline in GFR of 1.3 (0.3-7.6) ml.min-1.year-1 (P < 0.05 between groups). In the DG group, albuminuria increased from 1.4 (0.3-7.2) to 2.6 (0.1-21.6) g/24 h (P < 0.05) and in the NDG group, decreased from 2.2 (0.8-8.7) to 0.8 (0.2-2.5) g/24 h (P = 0.05). Mean arterial blood pressure (MABP) decreased from 118 +/- 3 to 104 +/- 3 mmHg (P < 0.05) in the DG group, whereas it remained unchanged in the NDG group (106 +/- 3 vs. 105 +/- 3 mmHg). In the DG group, the rate of decline in GFR correlated with systolic blood pressure (r = 0.62, P < 0.001), MABP (r = 0.52, P < 0.01), albuminuria (r = 0.55, P < 0.005), and GFR at entry (r = -0.45, P < 0.05). CONCLUSIONS: Our study demonstrated a more rapid decline in GFR and a progressive rise in albuminuria in type 2 diabetic patients with DG compared with type 2 diabetic patients with NDG. PMID- 10860187 TI - Long-term results of the Kumamoto Study on optimal diabetes control in type 2 diabetic patients. AB - OBJECTIVE: To examine whether intensive glycemic control could decrease the frequency or severity of diabetic microvascular complications, an 8-year prospective study of Japanese patients with type 2 diabetes was performed. RESEARCH DESIGN AND METHODS: A total of 110 patients with type 2 diabetes (55 with no retinopathy [the primary prevention cohort] and 55 with simple retinopathy [the secondary intervention cohort]) were randomly assigned to multiple insulin injection therapy (MIT) groups and administered three or more daily insulin injections or assigned to conventional insulin injection therapy (CIT) groups and administered one or two daily intermediate-acting insulin injections. Worsening of microvascular complications was regularly assessed during 8 years. Two or more steps up in the 19 stages of the modified Early Treatment of Diabetic Retinopathy Study classification in retinopathy and one or more stages up among three stages in nephropathy (normoalbuminuria, microalbuminuria, and albuminuria) were defined as worsening of complications. RESULTS: In both primary prevention and secondary intervention cohorts, the cumulative percentages of worsening in retinopathy and nephropathy were significantly lower (P < 0.05) in the MIT group than in the CIT group. In neurological tests after 8 years, the MIT group showed significant improvement (P < 0.05) in the median nerve conduction velocities (motor and sensory nerves), whereas the CIT group showed significant deterioration (P < 0.05) in the nerve conduction velocities and vibration threshold. From this study, the glycemic threshold to prevent the onset and progression of diabetic microvascular complications was as follows: HbA1c < 6.5%, fasting blood glucose concentration < 110 mg/dl, and 2-h postprandial blood glucose concentration < 180 mg/dl. CONCLUSIONS: Intensive glycemic control can delay the onset and progression of the early stages of diabetic microvascular complications in Japanese patients with type 2 diabetes. PMID- 10860188 TI - Optimal glycemic control in type 2 diabetic patients. Does including insulin treatment mean a better outcome? AB - Type 2 diabetes is a progressive disease with a significant risk for developing late complications. This article presents evidence related to the effect of glycemic control on the outcome of daily symptoms, microvascular complications, and macrovascular complications. Literature limited to Medline and the Cochrane Library was searched primarily for randomized clinical trials. In terms of education, present intervention studies indicate a positive effect on surrogate end points such as glycemic control, knowledge, practical skills, and psychological performance. Studies show improved glycemic control and plasma lipid profiles after moderate weight reduction. However, this positive effect is limited in time because weight is regained. With regard to oral blood glucose lowering drugs, clinical trials show a significant blood glucose-lowering effect of different available drugs. Both sulfonylurea and metformin have been shown to significantly reduce the risk of microvascular complications. In the U.K. Prospective Diabetes Study, intensive treatment with metformin in obese subjects reduced the risk for any diabetes-related event and stroke. A major problem is that many patients gradually experience increasing hyperglycemia, creating the need for combined treatment with several drugs including insulin. Insulin treatment has been shown to be effective in achieving satisfactory glycemic control over several years. There is also a positive effect on hard end points such as microvascular disease in the eye, kidney, and nerves. In conclusion, present evidence shows that optimal glycemic control can be attained in people with type 2 diabetes, resulting in fewer disease-related symptoms and a reduced risk of late complications. PMID- 10860189 TI - Prevalence and determinants of microalbuminuria in high-risk diabetic and nondiabetic patients in the Heart Outcomes Prevention Evaluation Study. The HOPE Study Investigators. AB - OBJECTIVE: To describe the characteristics of diabetic and nondiabetic participants in the Heart Outcomes Prevention Evaluation (HOPE) Study who are at high risk of developing cardiovascular (CV) disease and who have microalbuminuria (MA), and to identify the key determinants of MA in these two groups. RESEARCH DESIGN AND METHODS: Albuminuria was measured in 97% of patients enrolled in the HOPE Study as part of the MICRO-HOPE (MA, CV, and Renal Outcomes in HOPE) substudy. Baseline clinical characteristics of diabetic and nondiabetic participants with MA were recorded, and the univariate and multivariate relationship between these characteristics and the presence of MA was estimated for both groups. RESULTS: Baseline urinary albumin determinations were available in 3,574 (97.8%) diabetic participants and 5,708 (97.0%) nondiabetic participants. MA was detected in 1,151 (32.2%) diabetic participants and 837 (14.7%) nondiabetic participants. Age, waist-to-hip ratio, diabetes, smoking, hypertension, vascular disease, and left ventricular hypertrophy were independent determinants of MA in all participants. In diabetic participants, the odds of MA increased 16% for every 10.4 years of diabetes duration, and increased 8% for every 0.9% increase in glycated hemoglobin (assuming a GHb assay with an upper limit of 6% in the nondiabetic range). CONCLUSIONS: MA is independently associated with several risk factors for CV and renal disease in both diabetic and nondiabetic individuals at high risk for CV disease. PMID- 10860190 TI - Determinants of elevated urinary albumin in the 4,937 type 2 diabetic subjects recruited for the DIABHYCAR Study in Western Europe and North Africa. AB - OBJECTIVE: Whether ACE inhibition is useful for type 2 diabetic patients with micro- and macroalbuminuria remains unknown. The Non-Insulin-Dependent Diabetes, Hypertension, Microalbuminuria, Cardiovascular Events and Ramipril (DIABHYCAR) Study was set up to address this issue through a multicenter double-blind parallel placebo-controlled > or = 3-year trial in Europe and North Africa. In this article, we report the characteristics of the randomized patients. RESEARCH DESIGN AND METHODS: The main selection criteria were as follows: men or women aged > or = 50 years with type 2 diabetes treated with oral antidiabetic drugs, with or without hypertension, with a plasma creatinine level < 150 mumol/l, and with persistent micro- or macroalbuminuria, as assessed centrally by two successive urine samples containing a urinary albumin concentration > or = 20 mg/l. Patient characteristics were studied by comparing patients who were randomized to those who were not, taking their geographical origin into account. RESULTS: There were 25,455 patients screened for urinary albumin (20,296 from France, 918 from Germany, 1,019 from Northwest Europe, 969 from Central Europe, 959 from Mediterranean Europe, and 1,294 from North Africa). Of these patients, 4,937 were randomized. Compared with the nonrandomized patients, the randomized patients were older, more often men, more obese, had higher systolic/diastolic blood pressure and plasma glucose, smoked more tobacco, drank more alcohol, and had complications more frequently. Using a logistic regression analysis, all the above-mentioned items appeared as independent determinants for randomization into the study, with the exception of alcohol intake. The contribution of each item varied slightly from one geographical origin to another. CONCLUSIONS: The physical, biological, and behavioral characteristics create a poor renal and cardiovascular prognosis for the type 2 diabetic patients randomized to the DIABHYCAR Study because of micro- and macroalbuminuria. Testing the usefulness of ACE inhibition for the type 2 diabetic patients with microalbuminuria seems feasible through the DIABHYCAR Study. PMID- 10860191 TI - Lipid intervention trials in diabetes. AB - Most clinical trials of lipid intervention and coronary artery disease prevention have been conducted in study populations that exclude diabetic individuals. Three trials have conducted post hoc analyses of their diabetic subgroups. One of these was a primary intervention trial with gemfibrozil (Helsinki Heart Study). Although this trial found a reduction in coronary events, the numbers were too small to reach significance. The two other trials (the Scandinavian Simvastatin Survival Study [4S] and Cholesterol and Recurrent Events Trial [CARE]) were secondary intervention trials conducted with hydroxymethylglutaryl-CoA reductase inhibitors, simvastatin, and pravastatin. Both of these trials found a reduction in coronary events. Although these two trials present the strongest evidence in support of the clinical benefits of lipid reduction in diabetes, they must be interpreted with caution. They are post hoc subgroup analyses, they looked at mainly hypercholesterolemic populations, and they are secondary intervention studies. Four studies aimed at testing the "lipid hypothesis" specifically in diabetes are currently under way. Three of these studies (Fenofibrate Intervention and Event Lowering in Diabetes [FIELD], Collaborative Atorvastatin Diabetes Study [CARDS], and Lipids in Diabetes Study [LDS]) are primary prevention trials, with clinical events as the primary end point. FIELD uses micronized fenofibrate, CARDS uses atorvastatin, and LDS uses both micronized fenofibrate and cerivastatin alone or in combination. These trials are in the early stages of starting or recruiting. One study (Diabetes Atherosclerosis Intervention Study [DAIS]) using micronized fenofibrate is nearing completion. It is an angiographic study that combines those with and without preexisting clinical coronary disease. PMID- 10860192 TI - Effect of blood pressure control on diabetic microvascular complications in patients with hypertension and type 2 diabetes. AB - OBJECTIVE: The Appropriate Blood Pressure Control in Diabetes (ABCD) Trial is a prospective randomized blinded clinical trial that compares the effects of intensive versus moderate blood pressure control on the incidence and progression of type 2 diabetic complications. The current article discusses the results of 5.3 years of follow-up of 470 patients with hypertension and evaluates the effects of intensive and moderate blood pressure therapy using nisoldipine versus enalapril as the initial antihypertensive medication for nephropathy, retinopathy, and neuropathy. RESEARCH DESIGN AND METHODS: The 470 hypertensive subjects, defined as having a baseline diastolic blood pressure of > or = 90 mmHg, were randomized to intensive blood pressure control (diastolic blood pressure goal of 75 mmHg) versus moderate blood pressure control (diastolic blood pressure goal of 80-89 mmHg). RESULTS: The mean blood pressure achieved was 132/78 mmHg in the intensive group and 138/86 mmHg in the moderate control group. During the 5-year follow-up period, no difference was observed between intensive versus moderate blood pressure control and those randomized to nisoldipine versus enalapril with regard to the change in creatinine clearance. After the first year of antihypertensive treatment, creatinine clearance stabilized in both the intensive and moderate blood pressure control groups in those patients with baseline normo- or microalbuminuria. In contrast, patients starting with overt albuminuria demonstrated a steady decline in creatinine clearance of 5-6 ml.min 1.1.73 m-2 per year throughout the follow-up period whether they were on intensive or moderate therapy. There was also no difference between the interventions with regard to individuals progressing from normoalbuminuria to microalbuminuria (25% intensive therapy vs. 18% moderate therapy, P = 0.20) or microalbuminuria to overt albuminuria (16% intensive therapy vs. 23% moderate therapy, P = 0.28). Intensive therapy demonstrated a lower overall incidence of deaths, 5.5 vs. 10.7%, P = 0.037. Over a 5-year follow-up period, there was no difference between the intensive and moderate groups with regard to the progression of diabetic retinopathy and neuropathy. In addition, the use of nisoldipine versus enalapril had no differential effect on diabetic retinopathy and neuropathy. CONCLUSIONS: Blood pressure control of 138/86 or 132/78 mmHg with either nisoldipine or enalapril as the initial antihypertensive medication appeared to stabilize renal function in hypertensive type 2 diabetic patients without overt albuminuria over a 5-year period. The more intensive blood pressure control decreased all-cause mortality. PMID- 10860193 TI - Efficacy of diuretics and beta-blockers in diabetic hypertensive patients. Results from a meta-analysis. The INDANA Steering Committee. AB - OBJECTIVE: To review the effectiveness of diuretic or beta-blocker-based treatment of hypertension in diabetic patients. RESEARCH DESIGN AND METHODS: A meta-analysis on individual patient data was performed on four trials of the treatment of hypertension in which diabetic patients were included and treated with first-line diuretics or beta-blockers. The main outcomes were the relative risk of death, fatal or nonfatal stroke, fatal or nonfatal coronary events, and major cardiovascular events. RESULTS: There were 92 diabetic patients who received first-line beta-blockers and 1,008 who received diuretics. In the control groups, diabetic patients had nearly twice the risk of any outcome when compared with nondiabetic patients. The same blood pressure reduction was achieved under treatment in the diabetic and nondiabetic patients, except for systolic pressure, which decreased more in the nondiabetic patients at 1 year. In the 15,843 nondiabetic patients, the risk of all four outcomes was reduced significantly in the treated group. In the 2,254 diabetic patients, the risk reduction was significant only for fatal and nonfatal stroke (36%, P = 0.011) and major cardiovascular events (20%, P = 0.032), but not for death (5%, P = 0.65) and fatal or nonfatal coronary events (15%, P = 0.23). However, no heterogeneity was detected between diabetic patients and nondiabetic patients for any outcome. The numbers of outcomes avoided for 1,000 patients treated for 5 years were higher in diabetic patients (e.g., 38 major cardiovascular events) than with nondiabetic patients (e.g., 28 major cardiovascular events). CONCLUSIONS: These results show that hypertensive diabetic patients benefit from first-line treatment with diuretics. No conclusion can be drawn for beta-blockers, owing to the small sample size. PMID- 10860194 TI - Pleiotropic effects of statins in atherosclerosis and diabetes. AB - OBJECTIVE: To investigate the direct anti-atherosclerotic properties of statins. RESEARCH DESIGN AND METHODS: Using in vitro and ex vivo models, the effect of different statins on key events involved in atherogenesis has been investigated. We studied the ability of statins to modulate modified LDL-induced cholesterol esterification, metalloproteinase secretion by macrophages, and arterial myocyte migration and proliferation. The mechanisms underlying the inhibitory effect of statins have also been explored. Finally, the antiproliferative effect of sera from statin-treated patients has been confirmed in a cell culture system. RESULTS: Fluvastatin, simvastatin, lovastatin, atorvastatin, and cerivastatin, but not pravastatin, dose-dependently decrease smooth muscle cell (SMC) migration and proliferation. Moreover, statins are able to reduce cholesterol accumulation in macrophages in vitro by blocking cholesterol esterification and endocytosis of modified lipoproteins and matrix-degrading enzyme secretion. This in vitro inhibition was completely prevented by mevalonate and partially by all-trans farnesol and all-trans geranylgeraniol, confirming the specific role of isoprenoid metabolites (probably through prenylated protein[s]) in regulating these cellular events. The inhibitory effect of statins on SMC proliferation has been shown in different models of proliferating cells, such as cultured arterial myocytes and rapidly proliferating carotid and femoral intimal lesions in rabbits, independently of their ability to reduce plasma cholesterol. Finally, ex vivo studies showed that sera from fluvastatin-treated patients interfere with SMC proliferation. CONCLUSIONS: These results suggest that 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors exert a direct anti atherosclerotic effect in the arterial wall, beyond their effects on plasma lipids that could translate into a more significant prevention of cardiovascular disease. These findings provide a basis for the beneficial effect of statins in clinical trials also involving diabetic patients--a population with a higher absolute risk of recurrent cardiovascular events. PMID- 10860195 TI - Dietary trans fatty acid. AB - Trans fatty acids are unsaturated fatty acids that contain at least one double bond in the trans configuration. In the diet they occur at relatively low levels in meat and dairy products as a by-product of fermentation in ruminant animals or in hydrogenated fats as a consequence of the hydrogenation process. In general, dietary hydrogenated fat/trans fatty acids have been reported to increase LDL cholesterol levels relative to oil in the natural state or cis fatty acids. In contrast, dietary hydrogenated fat/trans fatty acids have been reported have to have little effect or decrease HDL cholesterol levels, the later observation restricted to relatively high intakes of trans fatty acids. These two effects result in higher, therefore less favorable, total or LDL cholesterol/HDL cholesterol ratios. Significant increases in Lp(a) levels have been reported after consumption of diets relatively high in trans fatty acids compared with either unsaturated or saturated fatty acids. However, the magnitude of the change is for the most part small and the physiological significance of this observation has yet to be resolved. Data related to the mechanism by which hydrogenated fat/trans fatty acids alter serum lipid levels and other risk factors for cardiovascular disease are in the nascent stages. At this time it would appear prudent that public health recommendations should be aimed at encouraging the moderate consumption of products low in saturated fat or minimally hydrogenated. Trans fatty acids intake should not be stressed at the expense of saturated fat but should augment it. PMID- 10860196 TI - The cost-effectiveness of exercise training for the primary and secondary prevention of cardiovascular disease. AB - BACKGROUND: Although exercise training improves cardiovascular disease (CVD) risk factors, few studies have evaluated its potential long-term cost-effectiveness. METHODS: Using the Cardiovascular Disease Life Expectancy Model, a validated disease simulation model, we calculated the life expectancy of average 35- to 74 year-old Canadians found in the 1992 Canadian Heart Health Survey. The impacts of exercise training on cardiovascular risk factors were estimated as a 4% decrease in low-density lipoprotein (LDL) cholesterol, a 5% increase in high-density lipoprotein (HDL) cholesterol, and a 6 mm Hg decrease in both systolic and diastolic blood pressure. Exercise adherence was estimated at 50% for the first year and 30% for all additional years. Costs for a supervised exercise program determined from Canadian sources and converted to US dollars were estimated at $605 for the first year (medical evaluation, stress test, exercise prescription, and program costs) and $367 for all additional years (program costs). For an unsupervised program, the costs were estimated at $311 for the first year and $73 for all additional years. RESULTS: The cost-effectiveness (CE) of an unsupervised exercise program (1996 U.S. dollars) was less than $12,000 per year of life saved (YOLS) for all individuals. The CE of a supervised exercise program was less than $15,000/YOLS for men with CVD, and between $12,000 and $43,000 for women with CVD and men without CVD. CONCLUSIONS: Given the relatively few risks, substantial long-term benefits, and modest costs, an unsupervised exercise training program represents good value for all. A more expensive supervised exercise program is also cost-effective for most individuals with CVD. PMID- 10860197 TI - Validity and reliability of the 6-minute walk test in a cardiac rehabilitation population. AB - PURPOSE: Although the 6-minute walk test is commonly used to assess the functional status of patients with severe cardiopulmonary disease, few studies have tested its value in a cardiac rehabilitation (CR) population with milder disease status. The purpose of this study was to examine the validity and reliability of the 6-minute walk in a Phase II/III CR program. METHODS: Ninety four patients (61 men, 33 women) aged 63 +/- 10 years completed three 6-minute walks on nonconsecutive days. Patients also completed the Duke Activity Status Index (DASI) and the Short Form 36 Health Survey (SF-36). In addition, maximum metabolic equivalents (METs) from a symptom-limited graded exercise test were obtained from files. RESULTS: The 6-minute walk was linearly related to maximum METs (r = 0.687, P < 0.001), supporting the validity of the test. Patients walked significantly farther in each 6-minute walk (F = 19.83, P < 0.001), and strong test-retest reliability was demonstrated (intraclass correlation = 0.97). Distance walked decreased with older age (F = 19.49, P < 0.001), with men walking farther than women (F = 7.19, P < 0.01). The 6-minute walk was moderately correlated with scores from the DASI (r = 0.502, P < 0.001), and the Physical Function subscale of the SF-36 (r = 0.624, P < 0.001). CONCLUSIONS: The 6-minute walk is a valid and reliable method of assessing functional ability in a Phase II/III CR population. A learning effect of 6% was observed over the three walks; however, it is unknown if this learning effect will be retained over time. This test may be particularly valuable to smaller CR centers that want to document functional improvements but do not have access to conventional treadmill tests. PMID- 10860198 TI - Comorbidities and the entry of patients with peripheral arterial disease into an exercise rehabilitation program. AB - BACKGROUND: Exercise rehabilitation is advocated to improve function in patients with peripheral arterial disease (PAD) who have intermittent claudication. Patients with PAD often have comorbid medical problems that may interfere with their ability to safely participate in exercise rehabilitation programs. There is a paucity of information regarding the medical comorbidities and the evaluation of PAD patients before their participation in exercise rehabilitation studies. The purpose of this study was to identify comorbidities that predicted exclusion of PAD patients from participation in an aerobic exercise rehabilitation clinical trial. METHODS: This was a prospective cohort study of 225 consecutive outpatient volunteers (mean age 68 +/- 8 years, SD) with a history of Fontaine Stage II PAD recruited for exercise rehabilitation. Patient eligibility was determined by a history and physical exam, blood chemistries, measurement of ankle to brachial index (ABI), and an exercise treadmill test. RESULTS: Seventy-nine volunteers (35%) were medically ineligible: 22 because of symptomatic coronary artery disease, 12 because of severe PAD, and the rest for a variety of medical problems. In stepwise logistic regression analyses, low ABI and use of insulin were predictors of exclusion, whereas peripheral revascularization was an indicator of inclusion. Age, a history of coronary artery disease, myocardial infarction, coronary bypass surgery, and hypertension were not independent determinants of eligibility. CONCLUSIONS: Insulin-requiring diabetes and a low ABI increase the likelihood that older patients with PAD will be ineligible to participate in a research exercise rehabilitation program, whereas peripheral revascularization was associated with inclusion. Whether intensive medical management and peripheral revascularization would enable the claudicants deemed ineligible for entry into the exercise rehabilitation program to safely exercise remains to be determined. PMID- 10860199 TI - Risk profile and health awareness in male offspring of parents with premature coronary heart disease. AB - BACKGROUND: The offspring of parents who suffer from premature coronary heart disease have a significantly higher risk of early cardiac death than controls. A genetic predisposition is compounded by a commonality of environmental risk factors within families. Increasing awareness, early detection and modification of risk factors are essential components of an effective public health strategy to protect this highly vulnerable population. METHODS: The sons (n = 571) of parents with premature coronary heart disease attended the Toronto Rehabilitation Centre for a risk factor evaluation that included an interview with questionnaire, measurement of body dimensions and blood lipids, and cardiopulmonary exercise testing. A follow-up questionnaire was sent out 2 years after the evaluation. RESULTS: Despite concern about family history, 23% of subjects were smokers and 75% were inactive. Objective data confirmed a substantial prevalence of cardiac risk factors: less than optimal cardiovascular fitness (48%), overweight (34%), total cholesterol > or = 200 mg/dL (46%), high density lipoprotein cholesterol < or = 35 mg/dL (26%), low-density lipoprotein cholesterol > or = 160 mg/dL (16%), triglycerides > or = 200 mg/dL (27%), and lipoprotein (a) > 30 mg/dL (24%). Although almost all had a family physician whom they had seen an average of 1.8 times in the past year, and 4.7 times in the previous 3 years, screening and risk factor intervention strategies were disappointing. Two-year follow-up data showed a heightened health awareness, with a greater proportion of subjects exercising and attempting to maintain an appropriate body mass. CONCLUSIONS: The male offspring of parents who have suffered a premature coronary event exhibit a substantial prevalence of modifiable risk factors. The family physician can play an essential role in promoting a healthy lifestyle through risk reduction counselling and screening. PMID- 10860200 TI - Modest effects of exercise training alone on coronary risk factors and body composition in coronary patients. AB - BACKGROUND: Cardiac rehabilitation programs have evolved to become secondary prevention centers. However, the independent effect of exercise alone on coronary risk factors and body composition in patients with coronary artery disease has not been well studied. OBJECTIVE: The aim of this study was to determine the effect of exercise training alone, without modification of dietary intake, on coronary risk factors and body composition in a coronary population. METHODS: The authors studied 82 coronary patients (23 females and 59 males) aged 61.2 +/- 12.2 years (mean +/- SD) before and after a 3-month exercise training program. Outcome variables included serum lipid values, glucose, insulin, body composition, body fat distribution, macronutrient intake, and peak aerobic capacity. RESULTS: Neither male nor female patients experienced a significant overall improvement in plasma cholesterol, low-density lipoprotein (LDL)-cholesterol, triglycerides, glucose, or insulin levels after the 3-month exercise training program. Dietary macronutrient intake was unaltered during the study period. Peak aerobic capacity increased by 3.4 +/- 4.7 ml/kg/min (17%, P < 0.0001) and high-density lipoprotein (HDL)-cholesterol increased from 38 +/- 10 to 41 +/- 11 mg/dL (8%, P < 0.001) after the rehabilitation program. Patients with baseline triglyceride levels over 200 mg/dL experienced a 22% decrease (from 374 +/- 205 to 293 +/- 190 mg/dL; P < 0.05) after conditioning. Patients with baseline HDL-cholesterol levels under 35 mg/dL also improved overall by 17% (from 29 +/- 3 to 34 +/- 5 mg/dL; P < 0.0001). Exercise-induced changes in plasma HDL-cholesterol were more related to changes in body composition and/or body fat distribution, rather than changes in peak aerobic capacity. CONCLUSION: Exercise conditioning alone resulted in relatively modest risk factor improvements in coronary patients after 3 months. High-density lipoprotein cholesterol measures increased by 3 +/- 8 mg/dL (8%). Patients with baseline triglyceride elevations experienced a 22% decrease. On the other hand, there were no overall effects on body weight, total cholesterol, LDL-cholesterol, triglycerides, glucose, or insulin levels. For most patients, exercise effects were minimal and nutritional and medical therapy will need to be used more aggressively to attain nationally recognized risk factor goals. PMID- 10860201 TI - Cardiac rehabilitation participation patterns in a large, tertiary care center: evidence for selection bias. AB - BACKGROUND: Clinical practice guidelines have been published for cardiac rehabilitation, directing programs to address secondary risk-reduction issues. The role of risk factor profiles in the referral of patients to cardiac rehabilitation programs has not been evaluated. METHODS: Patients from the Cardiovascular Information Registry at the Cleveland Clinic Foundation (CCF) who entered the CCF hospital-based cardiac rehabilitation program (n = 371) were compared with those who did not participate in the CCF program (n = 2960) with respect to gender, demographics, and risk factor profile for CAD. A random subset of those who did not participate in the CCF program (n = 100) was interviewed by phone to determine participation patterns in other rehabilitation programs. RESULTS: Only 11% of patients participated in CCF-based program. Standard risk factors were similar between participants and nonparticipants. Rehabilitation patients were younger (63 +/- 10 versus 66 +/- 10, P < 0.01) and as a group had better left ventricular function (moderate-severe left ventricle: 16% versus 23%, P < 0.01) than nonparticipants. Women were underrepresented in the CCF rehabilitation population (20% versus 30%, P < 0.01). Of the phone survey sample, 21% of patients entered other community-based rehabilitation programs. Similar trends with respect to risk factors, younger age, and better left ventricular function were noted for the community subset. However, women accounted for a greater percentage of the participants in the community programs than the CCF based program (42.8% versus 19.7%, P < 0.03). CONCLUSIONS: Conclusions based on institution-specific programs likely underestimate overall participation in cardiac rehabilitation. Traditional risk factors apparently are not considered when referring patients to cardiac rehabilitation programs. Younger patients with lower mortality risks preferentially participate in rehabilitation programs. Women are more likely to participate in community-based programs. Overall use of cardiac rehabilitation programs remains low. PMID- 10860202 TI - Patients with dilated cardiomyopathy and less than 20% ejection fraction increase exercise capacity and have less severe arrhythmia after controlled exercise training. AB - The results of this pilot study suggest that patients with a diminished ejection fraction as low as 16% can safely perform an exercise program. A significant improvement in peak VO2 and maximal work rate was achieved. Moreover, this study suggests that exercise training might diminish the severity of asymptomatic ventricular arrhythmia; however a larger controlled study is needed to confirm these findings. PMID- 10860203 TI - Welcome to medical humanities--and why. PMID- 10860204 TI - Tuberculosis, non-compliance and detention for the public health. PMID- 10860205 TI - Public deliberation and private choice in genetics and reproduction. AB - The development of human genetics raises a wide range of important ethical questions for us all. The interpersonal dimension of genetic information in particular means that genetics also poses important challenges to the idea of patient-centredness and autonomy in medicine. How ought practical ethical decisions about the new genetics be made given that we appear, moreover, no longer to be able to appeal to unquestioned traditions and widely shared communitarian values? This paper argues that any coherent ethical approach to these questions must be able both to uphold the moral status of the individual and at the same time recognise the communitarian, interpersonal dimensions both of the world in which we live and of personal autonomy itself. The paper then goes on to propose an approach to the resolution of the ethical questions raised by the use of the new genetics in reproductive choice through the development of a coherent and principled process of public reason and justification oriented towards the support and development of personal autonomy. PMID- 10860207 TI - Who should know about our genetic makeup and why? AB - Recent developments in biology have made it possible to acquire more and more precise information concerning our genetic makeup. Although the most far-reaching effects of these developments will probably be felt only after the Human Genome Project has been completed in a few years' time, scientists can even today identify a number of genetic disorders which may cause illness and disease in their carriers. The improved knowledge regarding the human genome will, it is predicted, in the near future make diagnoses more accurate and treatments more effective, and thereby considerably reduce and prevent unnecessary suffering. On the other hand, however, the knowledge can also be, depending on the case, futile, distressing or plainly harmful. This is why we propose to answer in this paper the dual question: who should know about our genetic makeup and why? Through an analysis of prudential, moral and legal grounds for acquiring the information, we conclude that, at least on the levels of law and social policy, practically nobody is either duty-bound to receive or entitled to have that knowledge. PMID- 10860206 TI - Human embryonic stem cells and respect for life. AB - The purpose of this essay is to stimulate academic discussion about the ethical justification of using human primordial stem cells for tissue transplantation, cell replacement, and gene therapy. There are intriguing alternatives to using embryos obtained from elective abortions and in vitro fertilisation to reconstitute damaged or dysfunctional human organs. These include the expansion and transplantation of latent adult progenitor cells. PMID- 10860208 TI - Working with mentally ill homeless persons: should we respect their quest for anonymity? AB - In recent years, the homeless population has received much attention as authorities attempt to comprehend this phenomenon and offer solutions. When striving to establish a relationship with the homeless person, many problems arise. We encounter this dilemma when respecting the right of the mentally ill to dwell neglected in the streets and simultaneously observe their inability to comprehend provisions such as housing, shelter, medical and mental care which contribute to their human dignity. The polarities of autonomy versus involuntary treatment are highlighted when treating the homeless population. PMID- 10860209 TI - Can unequal be more fair? A response to Andrew Avins. AB - In this paper, we respond to Andrew Avins's recent review of methods whose use he advocates in clinical trials, to make them more ethical. He recommends in particular, "unbalanced randomisation". However, we argue that, before such a recommendation can be made, it is important to establish why unequal randomisation might offer ethical advantages over equal randomisation, other things being equal. It is important to make a pragmatic distinction between trials of treatments that are already routinely available and trials of restricted treatments. We conclude that unequal randomisation could, indeed, be an ethical compromise between protecting the interests of participants and those of society. PMID- 10860210 TI - Examining consent within the patient-doctor relationship. AB - The notion of consent which rose to the forefront in biomedical ethics as an attempt to safeguard patients' autonomy, is relatively new. The notion itself requires qualification, for it precludes neither duress nor ignorance. More seriously, I argue here that consent is redundant except in situations where paternalism prevails. Paradoxically, these are the very situations where it may be difficult to uphold or to verify voluntary consent. I suggest that a request based relationship has the potential to overcome these difficulties. It enhances patients' participation in decision making, requires that the patients remain in command and avoids their subordination. Request is also more conducive to treatments that are representative of patients' own values and perceptions. In practice, what one wants and what one agrees to, often concur. But these are not conceptually identical issues, and they carry important differences of emphasis. PMID- 10860211 TI - Justified deception? The single blind placebo in drug research. AB - "Run-in" and "washout" periods involving the withholding of medication are widely used in drug research trials in pursuit of both patient safety and scientific reliability. Such no-medication periods can be justified ethically provided that they are apparent to patients, who can thereby properly consent to undergoing them. Less widespread, but still common, is the practice of "single blinding" no medication periods, concealing them from patients by means of placebo. Whilst all placebos involve a measure of concealment, their use is typically justified in drug research trials (i) by their preserving the uncertainty generated by the random allocation of different treatments within a drug trial; and (ii) by the researchers openly declaring both the randomisation process and the chances of receiving placebo. In the single blind placebo "run-in" or "washout", neither of these conditions is met. This paper considers three possible defences of the practice of using single blind placebo "run-ins" or "washouts" and finds them all to fail; the practice appears ethically unjustified. PMID- 10860212 TI - Like marriage, without the romance. AB - Physicians and philosophers have contributed to the field of medical ethics several different paradigms for the physician-patient relationship. Here I suggest another: marriage. Patients usually enter into relationships as we enter marriage: we allow our high hopes to obscure the possibility of deep disappointment. The argument of the essay encourages renewed focus on the contractual element of physician-patient relationships. PMID- 10860213 TI - Ethics consultation on demand: concepts, practical experiences and a case study. AB - Despite the increasing interest in clinical ethics, ethics consultation as a professional service is still rare in Europe. In this paper I refer to examples in the United States. In Germany, university hospitals and medical faculties are still hesitant about establishing yet another "committee". One of the reasons for this hesitation lies in the ignorance that exists here about how to provide medical ethics services; another reason is that medical ethics itself is not yet institutionalised at many German universities. The most important obstacle, however, may be that medical ethics has not yet demonstrated its relevance to the needs of those caring for patients. The Centre for Ethics and Law, Freiburg, has therefore taken a different approach from that offered elsewhere: clinical ethics consultation is offered on demand, the consultation being available to clinician(s) in different forms. This paper describes our experiences with this approach; practical issues are illustrated by a case study. PMID- 10860214 TI - At the coalface: medical ethics in practice. A double dose of double effect. AB - This paper presents a clinically orientated illustration of the doctrine of double effect. The case of an elderly gentleman with advanced cancer is discussed, with particular emphasis on two dilemmas encountered during the terminal phase of his illness. The author describes how the doctrine of double effect was applied to help the team make some complex management decisions. PMID- 10860215 TI - To give or sell human gametes--the interplay between pragmatics, policy and ethics. AB - The ever-growing acceptance and use of assisted human reproduction techniques has caused demand for "donated" sperm and eggs to outstrip supply. Medical professionals and others argue that monetary reward is the only way to recruit sufficient numbers of "donors". Is this a clash between pragmatics and policy/ethics? Where monetary payments are the norm, alternative recruitment strategies used successfully elsewhere may not have been considered, nor the negative consequences of commercialism on all participants thought through. Considerations leading some countries to ban the buying and selling of sperm, eggs and embryos are outlined and a case made that the collective welfare of all involved parties be the primary consideration in this, at times heated, debate. PMID- 10860216 TI - Paying for informed consent. AB - The Japanese Ministry of Health and Welfare has implemented a policy of paying physicians to explain the nature of the patient's medical condition and the treatment plan. We describe the precepts of this policy and examine ethical dimensions of this development. We question whether this policy will be sufficient to ensure patients will have the opportunity to become informed participants in medical decision making. The policy also raises a broader philosophical question as to whether informed consent is a fundamental ethical requirement of all doctor-patient encounters or an option that can be exercised by physicians for financial gain. The impact of this policy in Japan merits continued observation from abroad. PMID- 10860217 TI - Rationality and the wish to die--a response to Clarke. PMID- 10860218 TI - Hymenorrhaphy: what happens behind the gynaecologist's closed door? PMID- 10860219 TI - On an alleged problem for voluntary euthanasia. PMID- 10860220 TI - Appropriate use of DNR orders: a practical approach. AB - Too often, physicians, patients, and families make end-of-life care decisions despite poor physician-patient communication and misunderstanding by the patient and family about the effectiveness of cardiopulmonary resuscitation. We describe an approach to resolving conflict and reaching consensus on end-of-life care. This approach supports physician judgment to withhold futile treatment within the constraints of law and patient autonomy. PMID- 10860221 TI - How should patients taking the discontinued diabetes drug troglitazone be managed? PMID- 10860222 TI - When do you x-ray ankle sprains in patients with acute ankle injuries? PMID- 10860223 TI - Cancer genetics for the clinician: recommendations on screening for BRCA1 and BRCA2 mutations. AB - Mutations in the BRCA1 and BRCA2 genes confer a greatly increased risk of breast cancer, ovarian cancer, and other malignant diseases. This paper offers recommendations about who should be screened for BRCA mutations, and insights into the ramifications of BRCA testing. PMID- 10860224 TI - Transesophageal echocardiography: first-line imaging for aortic diseases. AB - Transesophageal echocardiography (TEE) is now commonly used to evaluate the thoracic aorta, because it is widely available and provides high-resolution images and flow information by Doppler. This article reviews the essential features on TEE of acute and chronic aortic diseases, such as aortic dissection, aneurysm, and atherosclerosis, and discusses its strengths, weaknesses, and indications. PMID- 10860225 TI - Alpha-blockers and congestive heart failure: early termination of an arm of the ALLHAT trial. AB - The Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) is a large, randomized double-blind study comparing four antihypertensive agents (chlorthalidone, doxazosin, amlodipine, and lisinopril) in hypertensive patients older than 55 years. The doxazosin arm was terminated early, when the trial's safety and monitoring board noted a twofold higher incidence of congestive heart failure in patients receiving doxazosin than in those receiving chlorthalidone (8.13% vs 4.45% at 4 years, P < .001). PMID- 10860226 TI - Acute respiratory distress syndrome: low-stretch ventilation improves survival. AB - In a recent major study, patients with acute respiratory distress syndrome or acute lung injury were randomly assigned to have their respirators set to deliver tidal volumes of either 6 mL/kg or a more-traditional 12 mL/kg. Mortality in the low-tidal-volume group was 31.0%, compared with 39.8% in the traditional-tidal volume group, a 22% difference (P = .007). PMID- 10860227 TI - Effective treatment of Alzheimer disease and its complications. AB - In Alzheimer disease, therapies to improve the core symptoms and perhaps even slow disease progression include cholinesterase inhibitors, receptor agonists, antiinflammatory drugs, and antioxidants. Neuroleptics, antiepileptics, and nondrug approaches are used to control and relieve complications such as delusions, hallucinations, paranoia, and agitated behavior. We outline a practical approach to the use of these therapies. PMID- 10860228 TI - Shortcomings of coronary angiography. PMID- 10860229 TI - Shortcomings of coronary angiography. PMID- 10860230 TI - The HOPE study. PMID- 10860231 TI - Concurrent factor V Leiden and prothrombin G20210A gene mutations in a patient with a history of recurrent thrombosis. AB - We report a case of a 35-year-old male with a history of recurrent thromboembolic events, who presented to the emergency room with right sided weakness and difficulty with speech. The patient's past medical history included two myocardial infarctions, two deep vein thromboses, and a pulmonary embolism. Subsequent laboratory evaluation indicated that the patient was heterozygous for both the factor V Leiden and prothrombin G20210A mutations. This case report emphasizes the importance of evaluating patients with suspected hereditary thrombophilia for both of these mutations. PMID- 10860232 TI - Eccrine sweat gland carcinoma: a case report and review of diagnosis and treatment. AB - Carcinomas of the skin appendices are rare neoplasms but for prognostic reasons it is important to differentiate them from the indolent squamous and basal cell carcinomas, as their behavior is more aggressive. We report on a case of eccrine sweat gland carcinoma that displayed all the typical features of those neoplasms. The patient sought medical attention after a lesion in his foot, already present for four years, began to enlarge and developed satellite lesions. The pathological diagnosis was made only after the lesion was initially misdiagnosed as basal cell carcinoma of the skin. Multiple chemotherapeutic regimens and radiation therapy were administered with only temporary benefit. The patient developed distant metastatic disease but survived with metastases for three years. He died nine years after the initial lesion developed in his foot and five years after the diagnosis. The diagnosis of sweat gland carcinomas can be facilitated by histochemical stains. In contrast to squamous and basal cell carcinomas of the skin, these are generally positive for the carcinoembryonic antigen (CEA). Once metastatic, these neoplasms are only infrequently, and usually briefly, responsive to either chemotherapy or radiotherapy and new treatments are urgently needed. Early recognition of the entity may allow more timely treatment. PMID- 10860233 TI - Dorsal thoracic cord compression from a spinal angiolipoma: case report and brief comment. AB - Angiolipomas are benign tumors usually found in the forearms of young adults. To the best of our knowledge only 63 cases of spinal angiolipomas have been reported in the literature up until 1999 (MEDLINE 1966-1999). We report a case of a spinal angiolipoma causing dorsal cord compression in a 44-year-old woman presenting with subacute lower limb paresthesias, in the absence of sensory or motor findings, which mimicked multiple sclerosis by history. Operative intervention was curative. PMID- 10860234 TI - Hospitals still essential but no longer the center of health-care delivery. If not hospitals, where's the center? PMID- 10860235 TI - Dr. Gurdon Wadsworth Russell's account of the 1853 railroad accident at Norwalk, Connecticut. A commentary on the person, the times, and the event. PMID- 10860236 TI - [Analysis of newborns' health status in twin pregnancies according to the duration of pregnancy and its delivery based on the materials of Reproduction and Obstetrics Clinic in 1995-1999]. AB - On the ground of the analysis of 79 twin pregnancies, the valuation of newborns' condition in Apgar's scale, according to the delivery means, has been executed. The lack of differences between the mean values of the scale has been ascertained. There was no indication of any difference between the condition of the twin I and II. Furthermore, there has been executed a detailed comparative analysis of the newborns' condition in the four periods of the pregnancy duration: 23-27, 28-32, 33-37 and 38-42 weeks of gestation. It has been ascertained that the newborns delivered through the abdominal delivery were in better condition than those born in the spontaneous delivery, in the 28-32 weeks of gestation period. The use of rather intraspinal rather than general anesthesia in the 33-37 weeks period gave better results by improving the newborns' condition. Moreover, there has been stated a similar condition of the newborns delivered through either spontaneous or abdominal delivery with intraspinal anesthesia in the periods of 33-37 and 38-42 weeks of gestation. PMID- 10860237 TI - [Family deliveries: socio-psychological analysis of participants]. AB - In the paper we undertook the qualitative analysis of the questionnaires of 60 couples who took part in the so called "family deliveries". We started that in most of this kind of deliveries took part the marriages aged between 26-30 years, of higher school education and living in towns. The cardinal motive of the couples to choose this kind of delivery was the need of "psychic support". To both of them the most important source of preparing to the familiar delivery was self-teaching and for the bearing women the most important was the concentration on the aim: "the husband-father is still present with me and I can undertake a spirit talk with him". Most of the participants appreciated very high our care to secure the best terms of safe delivery (in the special conditions) and the level of knowledge and professional preparation of our stuff to guide this kind of delivery. PMID- 10860238 TI - [Late maternity: the phenomenon of the end of the 20th century -- the estimate of clinical condition of mature newborns born at the Institute of Gynecology and Obstetrics in Lodz by older mothers]. AB - In the paper, the mature newborns of women over 40 were compared to the mature ones of young mothers. The babies were born in the Institute of Gynecology and Obstetrics in Lodz, in the years 1997-1998, and the comparison was done by the analysis of some neonatal parameters. The newborns of older women were delivered by cesarean section mainly. The birth weights average of the index babies was lower than the birth weights average of the controlled ones. Some clinical parameters in both analysed groups were similar. The most important difference concerned the congenital malformations and intrauterine hypotrophy--these disorders were clearly more frequent among the newborns of older mothers. PMID- 10860239 TI - [Active labor]. AB - In the paper we underlined the benefits of the labour in the vertical position in contrast to many disadvantages of parturition in recumbent or semi-recumbent position. These not natural positions are the consequences of the development of obstetrics manoeuvres as late as 200 years ago. In our opinion there is the necessity of the changing our minds on pros and cons of horizontal position during labour. PMID- 10860240 TI - [Clinical analysis of 1300 family deliveries]. AB - We presented the course of 1300 family deliveries and their comparison with the control group. We stated the elongation of the first stage of delivery (correspondingly 7 hours 10 minutes and 4 hours 10 minutes) and the second stage of delivery (70 minutes and 25 minutes). At the same time we observed that the obstetric procedures were much limited, especially the indication for caesarean sections (correspondingly 2.7% and 5.4%), and the general condition of new-borns estimated immediately after delivery according to Apgar scale was better too. The bad and poor general condition (estimated from 1 to 6 points) in the group of family deliveries was observed in 4.4% cases in comparison with 6.2% in the control group. PMID- 10860241 TI - [The influence of computer supervision of deliveries on the medical procedures during labor and neonatal post-delivery status]. AB - OBJECTIVES: The aim of the study was estimation of the influence of computer supervision of delivery on the way of medical management during partus. MATERIALS AND METHODS: Authors analyzed 10,983 labours which took place in years 1990-1999 in University Clinic of Reproduction and Obstetrics of Medical Academy in Wroclaw. Cases of labours without computer monitoring was the first analyzed group, and the cases of computer monitoring deliveries was the second group. In both groups, the count of cesarean sections performed because of first symptoms of fetal asphyxia, and Apgar score gained by newborns, were taken into consideration. RESULTS AND CONCLUSIONS: The frequency of cesarean sections and neonatal Apgar score gained by newborns were compared in two characterized group in order to evaluation the influence of computer monitoring deliveries on the neonatal status. No impact of computer supervision on the way of delivery procedures, quality of work and neonatals state was observed. PMID- 10860242 TI - [Analysis of twin pregnancies on the ground of women hospitalized in the Department of Reproduction and Obstetrics Medical University in Wroclaw in 1995 1999]. AB - The aim of this study was the general valuation of the course and the delivery means of the twin pregnancies. The research material composed of 83 from among 5540 pregnant women hospitalized in the Department of Reproduction and Obstetrics Medical University of Wroclaw in the years 1995-1999. The mean body mass values and the condition of the newborns have been analyzed on the ground of Apgar's scale, according to the date of delivery. In the period between 23 and 42 week of pregnancy a very high correlation between fetus' body mass and a high correlation between Apgar's scale and the pregnancy's duration has been ascertained. These values have also been estimated in particular periods: 23-27, 28-32, 33-37 and 38 42 weeks of gestation. Statistical analysis didn't indicate any difference between the mean values of Apgar's scale of the newborns from the periods of 33 37 and 38-42 weeks of gestation. There was no evidence of differences either in Apgar's scale values or in the twins' I and II body masses, as well in the whole examined group as in particular periods. PMID- 10860243 TI - [Delivery school at Municipal Hospital of Obstetrics and Gynecology, primary care: analysis of its activity]. AB - We presented the analysis of aims and effects of function the Delivery School, which is situated at Municipal Hospital Obst./Gynec. in Zory. PMID- 10860244 TI - [Rooming-in: a new standard in obstetrics and neonatology]. AB - A bibliography review concerning to mother and child care in "rooming-in" system (RI) was shown in this paper. Propagation of this form of care, main advantages of RI concerning breast-feeding promotion, organization of medical staff work and opinion of women in puerperium about the system were taken into consideration. There was also discussed "Humane Neonatal Care Initiative" which concentrates on dissemination of "rooming-in" in neonatal intensive care units. It issued from this analysis that newborn separation from mother is one of the main reason disordering lactation and rising of emotional link between mother and child. PMID- 10860245 TI - [The analysis of 135 water births]. AB - DESIGN: The authors showed the results of the study of the influence of warm tub bath during delivery on 135 women. MATERIALS AND METHODS: In a prospective study 135 women bathed in a warm tub bath during first and second stage of labor after a strictly normal pregnancy, ending with spontaneous delivery at term. A control group consisted of 135 women fulfilling the same criteria, but who did not take the bath during labor. The newborns weight, their condition, perineum injury, time of first and second stage of delivery and number of periteotomies was analyzed. CONCLUSIONS: The observed features were undistinguishing. PMID- 10860246 TI - [Delivery with husband]. AB - On the basis of the own questionnaire, the research concerning 60 married couples, experiencing family delivery, was made. It was stated that there is strong emotional bound between parents and strong need for experiencing the delivery together. PMID- 10860247 TI - [Fetal pulse oximetry in second stage of labor]. AB - OBJECTIVE: To compare the mean values of fetal oxygen saturation with fetal heart rate pattern in the second stage of labor (Melchior classification) MATERIAL AND METHODS: The study included 30 parturients in gestational age 37-41 weeks. Fetal oxygen saturation was recorded and averaged over the last 30 min of the second stage of labor Simultaneously, fetal heart rate and uterine contractions were monitoring. Fetal heart rate patterns were assessed according to Melchior's classification. At birth, the cord acid-base parameter (pH) was calculated. RESULTS: The lowest value of fetal oxygen saturation over the last 30 min of labor was 10%, the highest 60%, and the mean value 39.1 +/- 12.5%. Statistically, significant correlation between the mean value of oxygen saturation over the last 30 min of labor and pH arterial values (n = 25, p = 0.004, r = 0.42) was found. There was no statistically significant correlation between fetal oxygen saturation and pH venous values (n = 24, p. = 0.006, r = 0.49), but trend towards significance could be observed. Statistical analysis revealed that arterial pH was significantly correlated to Melchior's classification. The lowest pH and FSpO2 values coexisted with type 3 and 4 of FHR patterns according to Melchior's classification. CONCLUSIONS: The mean values of fetal oxygen saturation over the last 30 min of labor significantly correlate with arterial pH values. These values correlate with FHR abnormalities according to Melchior's classification. Fetal pulse oximetry seems to be an important, additional method assessing fetal well-being in the second stage of labor. PMID- 10860248 TI - [Spectral analysis of fetal heart rate variability]. AB - OBJECTIVE: To assess power spectral densities and power spectrum of FHR variability in two categories of fetal activity, quiescence and breathing movements, in pregnancies complicated by diabetes mellitus or intrauterine growth retardation. MATERIAL AND METHODS: The study group included 70 women, 35 in uncomplicated pregnancies and 35 with above-mentioned complications. The spectral analysis of FHR variability in fetal quiescence or breathing activity was performed. The power spectrum was analyzed at frequencies 0-1.0 Hz. Successively, the power spectral density was calculated. It has been presented the plots of spectral densities of beat-to-beat variabilities well during breathing episodes as "no breathing". Subsequently, the power spectra were calculated in both analyzed activities and groups. RESULTS: The highest values of power spectral densities (0.118 +/- 0.025) at 0.6-0.8 Hz were found in uncomplicated pregnancies. In pregnancies complicated by diabetes mellitus (0.048 +/- 0.049) or IUGR (0.042 +/- 0.041) these values were statistically significantly. At 0.6 Hz and 0.8 Hz power spectra of FHR variability were lower in quiescence comparing to breathing activity, as well in uncomplicated as complicated pregnancies. At these frequencies power spectra during breathing movements were significantly lower in complicated pregnancies comparing to uncomplicated. COMMENTS: The results of this study confirm the usefulness of spectral analysis in the assessment of FHR variability. The observed disturbances may reflect an abnormal fetal reactivity. The evaluation of breathing and cardiovascular systems interaction allows to study indirectly the central nervous system that coordinates both activities. PMID- 10860249 TI - [Back pain in pregnant women]. AB - In the paper a problem of spinal ailments in women in the third trimester of pregnancy is presented. The study concerns the changes of both body posture and pulvis position. Some disadvantageous situations pregnant women meet with, which cause feeling of pain in spine are analyzed. The alarming lack of proper knowledge about phenomenon of spinal pain in pregnant women is also pointed out. PMID- 10860250 TI - [Simplified cesarean section technique in the author's version]. AB - The paper contains the description of the modification of simplified method for caesarean section, based on the Joel-Cohen and Misgav Ladach method. Advantages of the simplified methods are described on the basis of the cited papers. Following stages of procedure are described in details. PMID- 10860251 TI - [The use of oxytocin and prostaglandin in pregnancies after cesarean delivery or uterine surgery]. AB - OBJECTIVES: Evaluation of risks involved with the use of oxytocin and prostaglandin in pregnancies after caesarean delivery or uterine surgery. MATERIALS AND METHODS: This study involved 267 pregnant women who underwent caesarean delivery or gynaecological surgery within the uterus (myomectomy or Strassmann corrective surgery). Oxytocin and prostaglandin was used to induce uterine contraction in these women. We analysed their delivery methods and the frequency of complications like uterine rupture and rupture of past caesarean scar. RESULTS: Uterine rupture occurred in one women had history of caesarean delivery (0.5%), and rupture past history of caesarean scar occurred in eight women (3.5%) who previously underwent caesarean delivery. Complications were not found in women with past history of myomectomy or Strassmann Corrective surgery. We did not observe an increase in complications among women who received prostaglandin compared to those who received oxytocin. CONCLUSIONS: 1. Cardiotocographic monitor during labour reduces the risk of uterine rupture and rupture of past caesarean scar in pregnant women with past history of caesarean delivery or other uterine surgeries. 2. The use of prostaglandin in comparison to oxytocin does not increase complications in women with past history of uterine surgery. 3. The choice of delivery method in pregnant women with past history of uterine should be individualised. PMID- 10860252 TI - Increased maternal plasma levels of soluble vascular cell adhesion molecule-1 (VCAM-1) in pregnancy induced hypertension (PIH). AB - One of the reason of PIH problems may be due to the presence of increased circulating levels of cell adhesion molecules, markers of endothelial damage and leukocyte activation. The objective was to evaluate the plasma levels of soluble vascular cell adhesion molecule in maternal peripheral blood of patients with PIH (pregnancy induced hypertension) and compared to those of normal healthy women with uncomplicated pregnancy. Maternal plasma samples were prepared from peripheral venous blood collected from 10 patients with PIH and 10 matched normotensive patients with uncomplicated pregnancies. Samples were assayed for soluble VCAM-1 by specific enzyme-linked immunosorbent assay (ELISA). Women with PIH had significantly higher plasma level of soluble VCAM-1 as compared with healthy pregnant women without PIH (653.50 vs. 456.39 ng/mL, respectively, p < 0.005). Our results on the increased plasma levels of soluble VCAM-1 in patients with PIH provide evidence for endothelial activation of PIH. It suggest that increased plasma level of soluble VCAM-1 could be an early marker of the maternal syndrome of PIH. PMID- 10860253 TI - PIH is associated with an increase of trophoblasts circulating in maternal blood. AB - One of the reason of PIH problems may be due to exposition to placental trophoblast. The objective of the work was to evaluate the number of trophoblast cells deported into maternal peripheral blood of patients with PIH (pregnancy induced hypertension) as compared to normal pregnancy. Trophoblasts have been detected, by cytofluorimetry, in peripheral maternal venous blood of hypertensive woman (15 cases) and normotensive pregnancy (16 cases). Women with PIH had statistically significant (p < 0.005) higher trophoblasts number than that found in normotensive pregnant women without PIH (16 cases). Our results indicate that the increased trophoblasts deportation into peripheral blood could be a marker of the maternal syndrome of PIH. PMID- 10860254 TI - [Comparative clinical analysis of cesarean section technique by Misgav Ladach method and Pfennenstiel method]. AB - Clinical and biochemical parameters were analysed in 55 patients who underwent a caesarean section performed using Misgav Ladach method compared to reference group of 41 patients who underwent caesarean section using Pfannenstiel method. Shortened operation time, shortened hospitalisation time and less postoperative morbidity were observed in the Misgav Ladach group. This kind of method seems to have advantages in comparison to Pfannenstiel method. PMID- 10860255 TI - [The use of forceps delivery in own analysis]. AB - OBJECTIVES: The clinical analysis of deliveries ended by forceps over the period of ten years. DESIGN: Review of perinatal outcome and indications to use outlet and low forceps or midforceps. MATERIALS AND METHODS: Author analysed 137 forceps deliveries in comparison to control group of 250 normal, vaginal labours. Obstetrical history, indications to use vaginal operation, duration of labour, hospitalisation time, newborns state in Apgar score or arterial cord pH, PaO2, and fetal or maternal injures were statistically analysed. The American College of Obstetricians and Gynecologists (ACOG) 1988 forceps classification be adopted for deliveries. Using outlet, low forceps and midforceps concerned with vaginal operation. RESULTS: The common indications to use outlet or low forceps were prolonged second stage of labour. The most frequent indication for the midforceps was a risk of fetal asphyxia and neonatal hypoxia. A major fetal injury occurred in midforceps, particularly with fetal head rotations. Furthermore, midforceps delivery increased incidence of maternal perineal trauma. The outlet or low forceps was safe for fetal outcome and trauma of the birth canal in comparison to normal vaginal delivery. CONCLUSIONS: The prophylactic use of outlet or low forceps has beneficial impact on the neonate because it shortens second stage of labour and decreased the incidence of neonatal hypoxia. The midforceps delivery increased a perinatal disorders and using cesarean section are better for child and mother. PMID- 10860256 TI - [Analysis of bacterial flora in the uterine cervix in women during preterm labor]. AB - The aim of this study was to estimate type and frequency the bacterial flora of uterine cervix of female in case of preterm labor. There were analyzed 153 uterine cervix swabs of women before 37 weeks of pregnancy. It has been shown that the main etiologic factors of infestations were Escherichia coli and Staphylococcus epidermidis. Positive results of bacterial investigation has been occurred more frequently in multiparas with precocious ammonirrhea. PMID- 10860257 TI - [Activity of N-b-D-acetylglucosaminidase molecular forms in amniotic fluid of tobacco smoking pregnant women with oligohydramnios or premature rupture of the membrane complications]. AB - OBJECTIVES: Morphological lesions in placenta and changed metal distribution through placenta, which were observed in pregnancy exposed to tobacco smoke, could cause alternations in lysosomal enzymes secretion to amniotic fluid. DESIGN: Assessment of total activity and molecular forms of N-beta-D acetylglucosaminidase in amniotic fluid in pregnancy with oligohydramnios or PROM. MATERIALS AND METHODS: The materials for studies were amniotic fluids collected from pregnancy with oligohydramnios and from pregnancy with PROM. Determination of N-beta-D-acetylglucosaminidase activity was performed with use of sodium salt of 3-crezolosulfthaleinyl-N-beta-D-acetylglucosamide as a substrate (test Boehringer, Niemcy). Creatinine was determined by kinetic method (test of Analco-GBG firm). Protein was determine by Bredford method. NAG-B was assessed after previous thermal NAG-A inactivation in 56 degrees C, for 2 hours and the difference between activity of total form and termostabile form of NAG were assessed. The microsomal and cytosol fractions were separated by ultracentrifugation. RESULTS AND CONCLUSIONS: The statistically significant growth of NAG activity in amniotic fluid in pregnant with oligohydramnios was observed. The cytosol form took over 90% of whole activity. The activity of NAG microsomal fraction was lower in amniotic fluid in women with PROM diagnosis. The positive coleration between NAG-A activity and the Pb level was observed in this group of women as well. In the second group of women with oligohydramnios, the positive coleration between NAG and Cd level, NAG-A and level, NAG-B and Pb level were demonstrated. Obtained results show on damage of placental and foetal membranes cell structure. It could be caused by metal ions cumulation and releasing of molecular N-beta-D-acetylglucosaminase form to amniotic fluid. PMID- 10860258 TI - [Evaluation of the neonatal status from pregnancies complicated by PROM or oligohydramnios in smoking women]. AB - OBJECTIVES: The aim of the study was analysis of neonatal status from pregnancies complicated oligohydramnios and from pregnancies complicated premature rupture of the membranes (PROM). MATERIALS AND METHODS: Authors analyzed 15 newborns from pregnancies with oligohydramnios diagnosed and 15- from pregnancies complicated PROM. Mothers of these newborns smoked cigarettes or were exposed to tobacco smoke. The features taken into consideration was prematurity, hypotrophy, malformations, birth weight, Apgar scale gained by newborns in first minutes after delivery. RESULTS AND CONCLUSIONS: The higher perinatal mortality, more frequent occurrence of hypotrophy and malformations were shown in newborns from pregnancies complicated oligohydramnios. The most frequent developmental anomaly was defect of nervosal system. Status of newborns from PROM pregnancies was getting better in according to lengthening duration of gestation. It wasn't stated in case of pregnancies with oligohydramnios complications. PMID- 10860259 TI - [Meaning of opening form of internal cervical opening in prognosis of cervical insufficiency]. AB - The introduction of the ultrasonographical examinations into the obstetrical diagnostics has created further possibilities in the process of recognition of the isthmico-cervical insufficiency. The qualification of the opening forms of the internal cervical os, described as Y, V and U forms, is possible with the utilization of transperineal and, especially, transvaginal sonography. The aim of this study was the valuation of the ultrasonographical examinations' usefulness in the diagnostics of the isthmico-cervical insufficiency and the estimation of the opening form of the internal cervical os as a prognostic factor in the pregnancy course. 265 cases of pregnant women hospitalized in Department of Fertility and Obstetrics Medical University of Wroclaw have been analyzed. On the ground of the executed examinations it has been ascertained that the ultrasonographical examination, considering the valuation of the opening form of the internal cervical os, is essential in the diagnostics of the isthmico cervical insufficiency. Furthermore, it has been proved that in the aspect of the obtainment of the full-term pregnancy, the worst-prognosing form of the internal cervical os is the U form and the statement of a U or V formed opening confirms the necessity of the circular suture of the cervix. PMID- 10860260 TI - [Cesarean section by the Misgav Ladach+ with the abdominal opening surgery by the Joel Cohen method]. AB - The purpose of this study was to compare 90 Misgav-Ladach cesarean section by the Joel-Cohen method with 45 others with Pfannenstiel incision. The Misgav-Ladach technique involves the Joel-Cohen method, that is a superficial transverse cut in the cutis, a small midline incision in the fascia, then blunt preparation of deeper layers, including the peritoneum, followed by manual transverse traction applied to tear the recti muscles and subcutis. The uterus is also opened using the blunt preparation after a small cut in the midline. After the delivery of the fetus and placenta the uterus is lifted through the incision onto the draped abdominal wall. Then the uterus is closed with one layer of continuous vicryl suture. The abdomen is closed by a continuous suture of fascia, and widely spaced silk stitches of the skin. We sometimes use continuous suture of the skin. We do not close visceral and parietal peritoneum, recti muscles and subcutis. In our experience Misgav-Ladach method is 50% less time consuming, it reduces blood loss by about 250 ml. and allows for a much faster delivery of the fetus than Pfannenstiel method. The post operative outcome of the two methods is similar. Using the blunt preparation in the Joel-Cohen method causes less trauma and shortens convalescence time. We therefore recommend Misgav-Ladach method for cesarean section. PMID- 10860261 TI - [Frequency cesarean section in pregnant women with uterine myoma]. AB - We were investigated in the connection of the frequency of cesarean sections at pregnant women with uterus myomatous and physiological pregnancies. 189 pregnant patients with myomas have been analyzed retrospectively 72 women with physiological pregnancies belong to the controlled group. Our results didn't confirm the higher risk of cesarean sections in analyzed group. PMID- 10860262 TI - [HELLP syndrome: problems in diagnosis and treatment illustrated on the basis of the fully symptomatic case]. AB - HELLP syndrome is a serious complication of Pregnancy-Induced Hypertension (PIH) which is very dangerous for the mother and her foetus. The prognosis depends on early diagnosis and correct treatment. The etiopathogenesis of the syndrome is still investigated but remains unclear. Taking as an example the precisely monitored, fully symptomatic case of the HELLP syndrome, we present review of modern opinions on pathogenesis, recognition and treatment. PMID- 10860263 TI - [Fibronectin level in amniotic fluid as an index of unavoidable labor]. AB - During uterus contractions detaching of amino-chorionic layer from uterus wall occurs and released fibronectin penetrates into amniotic fluid. The aim of this study was to estimate in a quantity mode the presence of fibronectin in amniotic fluid and to find the dependence between the fibronectin level in amniotic fluid and the period of time from collecting the sample to the labor. We wanted also to find the dependence between fibronectin level in amniotic fluid and duration of pregnancy, preterm rupture of amniotic membranes, patients' age, parity and number of deliveries. We analysed 86 pregnant women where we estimated the fibronectin level in specimens of amniotic fluid. During carrying out the experiment we noted that fibronectin is present in amniotic fluid and can be identified in a quantity mode. We have proved dependence between fibronectin level in amniotic fluid and the period of time from collecting the sample, up to the delivery. Fibronectin level in amniotic fluid in pregnancies uncomplicated with premature delivery was on the average 350 mg/ml. Increase of fibronectin in amniotic fluid above 700 mg/ml points at detaching of amino-chorionic layer and the occurrence of unavoidable preterm labor at the time no longer than 24 hours. Fibronectin level in amniotic fluid doesn't depend of pregnancy duration, preterm rupture of amniotic membranes. PMID- 10860264 TI - [Useful of cerebral placental ratio in pregnancies complicated by intrauterine growth retardation and it correlation with perinatal outcome]. AB - OBJECTIVE: The aim of this study was to asses usefulness of cerebro-placental ratio in the estimation of the intrauterine fetal well being in pregnancies complicated by intrauterine growth retardation and prediction of perinatal outcome. MATERIAL AND METHODS: We investigated 22 pregnant women between 28th and 40th week of pregnancy with IUGR detected by ultrasound examination. 19 pregnant women between 28th and 41st week of pregnancy was control group. We measured parameters blood flow in umbilical arteries and in middle cerebral arteries in both groups. We calculated cerebro placental ratios(CPR, CPP). We divided pregnant women with IUGR int 2 groups depending on correct (CPR, CPP > 1.08) or incorrect (CPR, CPP < 1.08). In both groups we analyzed perinatal outcome. RESULTS: In group pregnancies complicated with IUGR cerebro-placental ratios (CPR and CPP) were statistically significant lower than in control group. (for CPR p < 0.015 and for CPP p < 0.033). Sensitivity of cerebro placental ratio in screening small gestational age fetuses was 59% and specificity 89%. Sensitivity of cerebro placental ratio in predicting adverse perinatal outcome was 85% and specificity was 82%. CONCLUSIONS: Statistically significant decrease of cerebro-placental ratios is observed in pregnancies complicated wit IUGR. Cerebro placental ratio is very useful tool for prediction of adverse perinatal outcome. PMID- 10860265 TI - [Levels of Cd, Pb in blood and Zn, Cu, Cd, Pb in amniotic fluid of tobacco smoking women during pregnancy complicated oligohydramnios or premature rupture of membranes]. AB - OBJECTIVES: The ingredients of tobacco smoke have the impact on uterine blood vessels. They caused vascular insufficiency of placenta during development of gestation and changes in placental tissue and fetal membranes. It manifest changeable metal permeability and others symptoms. DESIGN: Assessment of metal contents in blood and amniotic fluid in pregnancies with oligohydramnios and PROM. MATERIALS AND METHODS: The materials for studies were blood and amniotic fluid of 30 pregnancies. All of them had made amino-punction between 20 and 38 week of pregnancy before procedure of amniotic fluid supplementation. 15 patients had diagnosed oligohydramnios and the next 15 women had diagnosed premature rupture of membranes. Metals were determined by method of electrothermal atomical absorbic spectrophotometry (ET-ASA). RESULTS AND CONCLUSIONS: Twice lower concentration of Zn and Cd and ten times lower concentration of Pb in amniotic fluids in examined women than women in normal pregnancy were observed. The women, with oligohydramnios in earlier period of gestation, smoked many more cigarettes and cotinine++ and Cd. were much higher. Both these facts had relevance with each other certainly. The stillborns were many more in this group. The different distribution of Cd, Pb, Zn in pregnant women with PROM and oligohydramnios, comparing with women in normal pregnancy was demonstrated. It is probable to be effect of placental tissue and fetal membranes disfunction. PMID- 10860266 TI - [The Doppler cerebroplacental ratios and perinatal outcome in post-term pregnancy]. AB - Our purpose was to determine whether the Doppler cerebroplacental ratios predicts perinatal outcome in postterm pregnancy. The middle cerebral to umbilical artery resistant and pulsatility indices (MCA PI/UA PI and MCA RI/UA RI) were measured in 59 postterm pregnancies. We found significant correlation between MCA PI/UA PI, MCA RI/UA RI, nonstress testing and intrapartum fetal heart rate assessment. There was also an association between MCA PI/UA PI and 1- and 5-minute Apgar score. We conclude that the Doppler cerebroplacental ratios provide useful information about perinatal outcome. PMID- 10860267 TI - [The use of Misoprostol preparation (Cytotec) in induction of labor at prolonged pregnancy]. AB - OBJECTIVES: Safety and effectiveness testing of Misoprostol use at prolonged pregnancies. DESIGN: Prospective, clinical study. MATERIAL AND METHODS: 50 pregnant women with prolonged pregnancy, monocyesis, cephalic longitudinal foetus lie, existing foetal membranes and lack of spontaneous delivery action. Women were given 50 micrograms Misoprostol (Cytotec) to posterior vaginal fornix in case to provoke delivery. Effectiveness of inducing, delivery lasting, way of its finishing and infant condition at birth were controlled. Results were matched with control group of 35 patients with physiological pregnancy, who delivered in spontaneous partus. RESULTS: Effective provocation was observed at 38 pregnant. Natural way delivery was observed at 40 patients. In 10 cases caesarean section was done. Lasting time of birth, way of finishing, infant condition at birth and number of complication do not differ statistically between examined and control group. CONCLUSION: Misoprostol can be effective and save of delivery induction in prolonged pregnancy. PMID- 10860268 TI - [Comparative analysis of modification of Misgav-Ladach and Pfannenstiel methods for cesarean section in the material of Fetal-Maternal Clinical Department PMMH RI between 1994-1999]. AB - PURPOSE: Comparative analysis of own modification of Misgav-Ladach (mML) and Pfannenstiel methods for caesarean section in the material of Fetal-Maternal Medicine Clinical Department PMMH-RI between 1994-99. MATERIAL AND METHODS: Study group consists of 242 patients. In all women from this group we performed caesarean section using Misgav-Ladach method. Among all patients from control group counting 285 women we performed caesarean section applying Pfannenstiel method. To analyse clinical postoperative course in both groups we took account several parameters. RESULTS: Statistical analysis revealed that most of clinical postoperative course parameters was significantly better values in the study group we performed caesarean section using Misgav-Ladach method. CONCLUSIONS: The benefits of Misgav-Ladach method, with less pain post-operatively and quicker recovery, are all a by-product of doing the least harm during surgery and removing every unnecessary step. This method is appealing for its simplicity, ease of execution and its time-saving advantage. PMID- 10860269 TI - [The evaluation of respiratory distress syndrome (RDS) risk factor evidence in newborns]. AB - The aim of this study was to evaluate RDS risk factors in newborns of mothers with define pregnancy complications. The study group included mothers with imminent preterm delivery, intrauterine growth retardation, gestation cholestasis, serological collision, oligo- and polyhydramnios and foetus life threatens risk factors. RDS appeared in newborns born up till 35 weeks of gestation, for the risk factors analysis only preterm delivery, which were ended until 36 weeks of gestation were taken into consideration. Hypertension appeared to be a significant risk factor increasing the risk of RDS evidence 5 times and asphyxia, which increased by 4. In the logistic regression analysis model hypertension showed to increase the risk of RDS evidence 6 times, even when the time of delivery was considered. A, when the time of gestation was prolonged even one week the risk of RDS was decreased by 30%. The was no significant increase in RDS evidence in offspring of mothers with diabetes mellitus, cholestasis gravidarum, hypotrophia foetus or oligo and polyhydramnios. PMID- 10860270 TI - [Evaluation of predictive value of lung maturity in newborns based on effectiveness of amniotic fluid study]. AB - The aim of this study was to evaluate the predictive value of respiratory distress syndrome-RDS evidence using amniotic fluid diagnostic tests. 200 amniotic fluid samples obtained by amniocentesis were study to evaluate pulmonary tissue maturity status. The PG test was 30% specific and 100% sensitive; FLM test showed 50% specificity and 67% sensitivity. The Sbarr tests OD 400 and 650 showed high specificity, relatively 88% and 71% and a low sensitive of 21% and 29%. Woyton test showed 62% specificity and 36% sensitivity. Foam test had 11% specificity and 100% sensitivity. None of these testes used alone cannot predict the newborn pulmonary tissue maturity status and therefore all these test should be used combined. In our study RDS appeared in 8.4% of study newborns, and there was no evidence of RDS in newborns born after 35 weeks of gestation. PMID- 10860271 TI - [Cesarean section for the second twin]. AB - Twin pregnancies delivered after 26 weeks gestation were reviewed. Of 167 twin pregnancies 101 (60.5%) were delivered by cesarean section. There were 8 combined vaginal-abdominal deliveries, including 4.8% of all twins delivered and 7.3% of all twins delivered by cesarean section. The commonest indication for the cesarean birth of the second twin was transverse lie. The data suggest that cesarean section of the second twin is not possible to predict by obstetrics variables including antepartum ultrasound. In order to decrease the rate of combined vaginal-abdominal deliveries the need to reassess the currently protocol of management for twin delivery is indicated. PMID- 10860272 TI - [Vaginal delivery after previous cesarean: comparison between own method and Weinstein's predictive score]. AB - 121 women after previous caesarean section were examined. The amount of successful labours were compared (according to the own diagnostic card) with the foreseeing probability of vaginal labours according to Weinstein's score. 63% pregnant woman was classified to the natural labour on the base of the own diagnostic card, probability of natural labour according to Weinstein's score were more or equal than 67%. There were no agreement between prognostic Weinstein's scale and our own method in the cases of failed attempt of labour followed by caesarean section. PMID- 10860273 TI - [Ultrasound evaluation of fetal lung maturity in imminent preterm labor cases]. AB - The development of ultrasound examination lets us perform the biophysical assessment of fetus lungs. From 1986, at the Ivth Clinic of Obstetrics and Gynecology in Tychy, examinations of fetus lung were performed with Siemens and Bruel & Kjaer real-time ultrasonographic equipped. The elasticity of fetal lung (DLS--dynamic lung score) was estimated by taking a transversal section through the fetal chest in a plane at fetal in a plane at fetal heart and presented in 3 grade scale: I-grade--lack of elasticity (immaturity of lungs), II-grade--non complete elasticity, III-grade--normal elasticity (maturity of lungs). PMID- 10860274 TI - [Management of breech delivery in term pregnancy: own diagnostic and prognostic model]. AB - Using own diagnostic and prognostic model, 1157 pregnant women with breech presentation in term were analysed. In 1008 cases (87.1%) they delivered according to prognosis established formerly. Manual support in delivery was required in 748 cases (64.6%). Elective caesarean section was performed in 260 women (22.5%). In 149 cases (12.9%) immediate caesarean section appeared, because of additional reasons, despite normal delivery prognosis. PMID- 10860275 TI - [Relationship between Doppler velocimetry at middle cerebral artery and umbilical artery and status of newborn]. AB - Aim of investigations was qualification of account between Doppler parameters in estimation of fetal state. Investigations one passed on 30 fetuses and newborn children in pregnancies brought. Doppler parameters one priced at use of sonographic device Toshiba SSH 140 A/G and searchers of type convex about working frequency 3.75 MHz. Following Doppler flow blood parameters were analyzed: maximum blood speed (V1) average blood speed (V2) and minimum blood speed (V3), systolic/diastolic ratio (S/D), resistance index (RI), pulsatile index (PI) and proper flow in umbilical cord vein (MF) in following dishes of feto-placental circulation: middle cerebral artery (MCA) and umbilical cord artery (UA). Acid base equilibrium and gasometry of blood in umbilical cord dishes one marked at use of device Ciba-Corning 278 Blood Gas System and parameters of oxygenation of blood at use of device Ciba-Corning 270 CO-OXIMETER. At new-born children one priced pH-metry (pH) and gasometry (pO2, pCO2, BAA) in blood umbilical cord arterial and venous were measured. The newborn children were estimated by Apgar score. There were following essential statistical correlations between Doppler parameters of fetal blood flow and with parameters of acid-base equilibrium of new-born child: 1/ between V2 and V3 in UA and with supply of rules (BAA) in UV (p = 0.027; p = 0.009) and UA (p = 0.035; p = 0.003) and venous pH (p = 0.022; p = 0.009); 2/ between RI in UA and BAA in UV (p = 0.006) and UA (p = 0.010); 3/ between PI in UA and BAA in UV (p < 0.0001) and UA (p < 0.0001) and pH venous (p < 0.0001). We can conclude that Doppler investigations only by measure of parameters of blood flow in middle cerebral artery and umbilical artery in expectation of state of birth new-born child priced across parameters of acid base equilibrium and Apgar score are not very useful, however they are helpful. PMID- 10860276 TI - [The analysis of estrogen and pregesterone receptors in leiomyomas cell in different phases of menstrual cycle]. AB - The aim of a study was immunocytochemical analysis of estrogen (ER) and progesterone (PgR) receptors in uterine leiomyomas cells derived from patients who underwent an operation in three different phases: in the follicular phase of the menstrual cycle (12 cases), in the luteal phase (20 cases) and in the postmenopausal age (18 cases). The examined receptors reflect the tissue sensibility to activity of sex steroid hormones and play an important role in the pathogenesis of uterine leiomyomas. Performed studies showed that expression of ER and PgR does not depend on the phase of the menstrual cycle and that it seems to be similar like in the postmenopausal stage. In all groups PgR expression was significantly higher then the expression of ER. In the group of patients in which the surgery was performed during the luteal phase of the menstrual cycle was demonstrated negatively correlation (-0.4846) between the expression of ER in relation to the expression of PgR. PMID- 10860277 TI - Cardiac function evaluated by transesophageal echocardiography during cardiopulmonary bypass. AB - OBJECTIVE: To evaluate cardiac function at cardiopulmonary bypass weaning, we applied a new technique clinically to determine the approximated Emax without using a conductance catheter. METHODS: Subjects were 5 patients. The left ventricular end-systolic pressure was obtained by overlaying the radial arterial pressure curve on the left ventricular pressure curve. Left ventricular end systolic volume was assessed by a transesophageal echographic apparatus. At cardiopulmonary bypass weaning, volume loading was applied to increase left atrial pressure by a few mmHg while fixing the pump flow rate at half flow. Changes in left ventricular end-systolic volume and approximated left ventricular end-systolic pressure for total heart beat were plotted during this period, and the gradient of the regression line was taken as approximated Emax. RESULTS: Approximated Emax ranged from 1.29 to 3.28 (mean 2.13 +/- 0.72), and its correlation coefficient was 0.80 +/- 0.06. CONCLUSION: Our new technique is useful in evaluating cardiac function during cardiopulmonary bypass. PMID- 10860278 TI - Long-term results of root reconstruction using the Carrel patch. AB - OBJECTIVE: The treatment of annuloaortic ectasia in patients, including those with Marfan syndrome, ascending aortic dissection, and other disorders of the ascending aorta and aortic valve presents a surgical challenge that has, unfortunately, shown high hospital mortality up to now. Improvements in graft materials and advanced surgical techniques have, however, begun to reduce hospital mortality. SUBJECTS AND METHODS: We retrospectively analyzed the records of 47 consecutive patients who undergoing aortic root reconstruction using the Carrel patch between January 1991 and March 1999. RESULTS: Postoperative complications included myonephrotic metabolic syndrome caused by femoral artery cannulation in 2 patients. Reexploration was done to halt bleeding in 2 patients. In 1 of 4 acute dissection patients, retrograde filling of the false lumen was demonstrated postoperatively. Overall surgical mortality in this series was 2.1% (1 of 47). The cardiac-event free rate was 98% at 5 years and 88% at 8 years. Actual survival is 97.8% at 8 years. No anastomosis complications were seen during follow-up (average: 32.7 months) (about 2.73 years). CONCLUSION: Surgery is considered feasible in any anatomic variation of aortic root disease, even in coronary ostial minimal dislocation, and the Carrel patch holds hope in preventing of anastomotic pseudaneurysm and ensuring long-term survival. Our experience suggests that modified Bentall operation, or aortic root remodeling using the Carrel patch, has few late-term complications, even in Marfan patients. PMID- 10860279 TI - Percutaneous cardiopulmonary support with heparin-coated circuits in postcardiotomy cardiogenic shock. Efficacy and comparison with left heart bypass. AB - OBJECTIVE: Percutaneous cardiopulmonary support, a simplified form of venoarterial bypass, using totally heparin-coated circuits, has recently come into clinical use. To clarify its efficacy in postcardiotomy cardiogenic shock to aid weaning from cardiopulmonary bypass, we compared results of percutaneous cardiopulmonary support with those of left heart bypass using a centrifugal pump. METHODS: We reviewed 18 patients treated between 1991 and 1998 who could not be weaned from cardiopulmonary bypass. Nine were aided by totally heparin-coated percutaneous cardiopulmonary support (PCPS group), and 9 supported by left heart bypass using a centrifugal pump (LHB group). In both groups, activated clotting time was controlled at 150-200 seconds using minimal doses of heparin as needed. RESULTS: Weaning and survival rates were higher in the PCPS group than in the LHB group (100% vs 55.6%, and 66.7% vs 22.2%). The PCPS group had a smaller amount of blood loss and needed a smaller amount of blood components in the immediate postoperative period. One percutaneous cardiopulmonary support patient required surgical re-exploration for postoperative bleeding (11.1%), but no clinical thromboembolic event occurred in the PCPS group. In the LHB group, 5 patients underwent surgical re-exploration for postoperative bleeding (55.6%), and 2 underwent thrombus extirpation in the left ventricle (22.2%). CONCLUSIONS: Although this study was retrospective and historical backgrounds could have been involved, our data suggest that totally heparin-coated percutaneous cardiopulmonary support system appears more effective as an aid to weaning from cardiopulmonary bypass and in short-term circulatory support for patients in postcardiotomy cardiogenic shock. PMID- 10860280 TI - Experimental myocardial preservation study of adding perfluorochemicals (FC43) in lidocaine cardioplegia. AB - OBJECTIVE: Lidocaine exhibits a cardioplegic action via acute inhibition of sodium influx into the myocardial cells. In terms of the cardiac function and calcium dynamics in the myocardial cells, we investigated the myocardial protective effect of addition of FC43 of Perfluorochemicals, which has an excellent oxygen transport function to meet the myocardial oxygen demand, on lidocaine-induced cardioplegia. METHODS: Isolated rat hearts were perfused with Langendorff mode and were divided to three experimental groups. During of preservation, these hearts were perfused continuously with the next three solution, potassium chloride was added to Krebs-Henseleit bicarbonate buffer to make potassium concentration of 20 mM in the first group (Group A), 2 mM lidocaine was added to Krebs-Henseleit bicarbonate buffer in the second group (Group B), and 2 mM lidocaine and 20% FC43 were added to Krebs-Henseleit bicarbonate buffer in the third group (Group C). After 60 minutes of continuous perfusion, the cardiac function and the intracellular calcium concentration in Groups A and B during cardioplegia were measured. Furthermore, after 360 minutes of continuous coronary perfusion, the cardiac function were measured in Group B and Group C. RESULTS AND CONCLUSIONS: Lidocaine cardioplegia showed a good recovery of cardiac function, because lidocaine induced prompt cardiac arrest by blocking sodium influx and inhibited the intracellular calcium overload by the following inhibition of sodium-calcium channels. Moreover, our results suggested that combining Perfluorochemicals with lidocaine produced a more effective myocardial-preservation that meets the myocardial oxygen demand during long-term cardiac arrest. PMID- 10860281 TI - Life-saving strategy for left ventricular free wall rupture after acute myocardial infarction. Infarction-covering repair on the ruptured site under the beating heart. AB - OBJECTIVE: Left ventricular free wall rupture after acute myocardial infarction is a serious complication with high mortality. For life-saving, it is important how to maintain poor hemodynamics till operation. We have consistently made it our strategy to attach percutaneous cardiopulmonary support system and intra aortic balloon pumping immediately after the diagnosis regardless of the type of left ventricular free wall rupture and of the hemodynamic conditions, and perform an infarction-covering repair under the beating heart. We have studied the short term and middle-term results after the operations, and have evaluated the efficacy and problems of this procedure. METHODS: Since September 1994, we have performed this method in six of eight patients with left ventricular free wall rupture. RESULTS: As results, five of the six patients (83%) were saved including two cases of blow-out type. Our strategy for left ventricular free wall rupture showed several advantages for preoperative and intraoperative maintenance of the hemodynamic conditions, and for preservation of some reversible myocardium by the simple technique of infarction-covering repair under the beating heart. These resulted in shortening the operation time, decreasing the incidence of low cardiac output syndrome, and obtaining a satisfactory rate of life-saving. CONCLUSION: We believe that this infarction-covering repair based on our strategy is effective for life-saving during the acute period. PMID- 10860282 TI - Evaluation of leukocyte-reducing arterial line filter (LG6) for postoperative lung function, using cardiopulmonary bypass. AB - OBJECTIVE: To prevent postoperative pulmonary dysfunction, we have investigated the effect of the Leuko-Guard 6 leukocyte-reducing arterial line filter (LG6) on postoperative lung function. METHODS: Twenty-six cases of adult valvular heart disease were included in this study. Thirteen cases were operated upon using the LG6 (Group LG), and 13 cases were operated upon using a conventional arterial line filter (Group C). Neutrophil, polymorphonuclear leukocyte elastase and lipoperoxide were measured for this study, and the lung function was evaluated using the Oxygenation Index (PaO2/FiO2). RESULTS: Statistically significant differences were observed in neutrophil counts between Group LG and Group C (LG = 2225 +/- 572/mm3, C = 3157 +/- 1413/mm3, p = 0.04) at 5 minutes after the onset of cardiopulmonary bypass. In simultaneous blood sampling from the pulmonary artery and the pulmonary vein, the sequestration of neutrophil in the lung decreased in Group LG after the discontinuation of cardiopulmonary bypass. Release of polymorphonuclear leukocyte elastase from the lungs was significantly decreased (p = 0.04) in the Group LG at 1 hour post-bypass. Significant differences were observed in the Oxygenation Index between Group LG and Group C (LG = 398 +/- 72, C = 326 +/- 71, p = 0.019) at 3 hours post-bypass. CONCLUSION: We concluded that LG6 improved the postoperative lung function, and its mechanism might be derived from the prevention of leukosequestration in the lungs that occurs during the rewarming phase due to selective absorption of activated leukocyte by the LG6. PMID- 10860283 TI - Endoluminal laser application under percutaneous cardiopulmonary support in severe tracheal stenosis. AB - Laser ablation under bronchoscopic guidance was conducted on 2 patients with severe tracheal stenosis. Case 1 was a 57-year-old man admitted to our emergency unit because asphyxia. Bronchoscopy showed the lumen occluded at the bifurcation by tracheal cancer. Case 2 was a 62-year-old woman who underwent tracheostomy elsewhere for respiratory failure caused by a brain contusion and was treated for 3 months. After transfer to our emergency unit, bronchoscopy showed severe tracheal stenosis. Tracheoplasty conducted under bronchoscopy used a noncontact Nd:YAG laser at an output of 10-40 W and irradiation time of 1 second per shot. Total irradiation energy was 1700-1900 J. Percutaneous cardiopulmonary support was used during the laser procedure due to asphyxia. All procedures were completed satisfactorily and clinical symptoms improved dramatically in both cases. Laser tracheoplasty under bronchoscopic guidance treated severe stenosis safely and completely. Percutaneous cardiopulmonary support was very useful in preventing severe respiratory failure or asphyxia during this procedure. PMID- 10860284 TI - Surgical management of ductus arteriosus aneurysm in adults. AB - Aneurysm of the ductus arteriosus is rare, especially in adults, and preoperative diagnosis is very difficult, requiring intraoperative diagnosis. A 71-year-old man was preoperatively diagnosed with ductus arteriosus aneurysm by computed tomographic scanning, magnetic resonance imaging, and aortography, which demonstrated a saccular aneurysm of the distal aortic arch. Under partial extracorporeal circulation, the aneurysm was replaced by an artificial vascular graft. The location and morphology of the aneurysm confirmed the preoperative diagnosis. The postoperative course was uneventful. Surgery for this condition is safe and is recommended the same as for common thoracic aortic aneurysm. PMID- 10860285 TI - Reoperative coronary artery bypass via left thoracotomy. AB - The patient was a 49-year-old woman. When she was 39 years old, she underwent coronary artery bypass grafting (left internal thoracic artery to left anterior descending artery, saphenous vein graft to first diagonal branch). At the age 48, she had effort angina. On coronary angiography, triple-vessel disease was found, and she was treated conservatively. Progression of the disease was confirmed with detection of the left circumflex artery associated with jeopardized collateral to the right coronary artery showing total occlusion. The patient underwent reoperation. Since the left internal thoracic artery was patent despite occlusion of the saphenous vein graft, the approach of left thoracotomy was employed. Under cardiopulmonary bypass with ventricular fibrillation and left vent through left atrial appendage, the right radial artery was anastomosed to the left circumflex artery from the descending thoracic aorta, and the right gastroepiploic artery was anastomosed to the right coronary artery (4AV branch). Patency of the bypass was confirmed postoperatively. We consider this operative technique was especially useful for reoperation in cases of a patent internal thoracic artery in which left thoracotomy can be conducted safely. PMID- 10860286 TI - Ultrasonic aortic valve decalcification. AB - We report a case of calcific aortic stenosis in a 79-year-old man who had undergone aortocoronary bypass. Since ordinary valve replacement was problematic because of severe annular calcification, a small annular diameter, and three patent vein grafts, we performed ultrasonic aortic valve decalcification. As a result, the pressure gradient across the aortic valve decreased from 100 mmHg to 25 mmHg, and the patient is doing well to date at two years after the operation. Although long-term results of ultrasonic aortic valve decalcification are still undetermined, it appears worth considering in cases in which ordinary valve replacement could be extremely difficult. PMID- 10860287 TI - Angiosarcoma of the chest wall. AB - A 50-year-old man sustaining bilateral chest wall angiosarcomas at intervals of several months underwent lesion resection. Angiosarcoma is so rare that we found no case in the literature who had undergone bilateral chest wall surgery for such tumors. His second tumor was thought to be metastatic rather than primary. Despite the 2 surgeries, irradiation, and chemotherapy, however, his prognosis was grave, as in other reports on angiosarcoma. PMID- 10860288 TI - Thoracolithiasis. AB - Thoracolithiasis without any history of chest traumas or interventions is pathologically rare, with only 9 cases including our 2, reported thus far in the literature. Case 1: A 76-year-old man admitted to our hospital had an abnormal shadow in chest radiography that gradually enlarged. Serum carcinoembrionic antigen was slightly elevated during follow-up. A milky white tumor 1.5 cm in diameter with many projections was found in the thoracic cavity and removed by thoracoscopy. Histopathological examination showed the tumor to consist of fibrous tissue with fatty necrosis at the core. Case 2: A 54-year-old woman admitted to our hospital had an abnormal shadow in chest screening radiography in 1998. Transbronchial biopsy showed this shadow to be lung adenocarcinoma. A small trigonal pyramid-shaped milky white nodule 5 mm in diameter was found in the thorax during lobectomy for lung cancer. Histopathological examination showed this nodule also to consist of fibrous tissue with fatty necrosis. PMID- 10860289 TI - Pulmonary atresia with intact ventricular septum, antegrade coronary-right ventricular sinusoidal communication, and Wolff-Parkinson-White syndrome. AB - A fenestrated Fontan operation was performed successfully in a patient with pulmonary atresia with intact ventricular septum, an antegrade sinusoidal communication, and Wolff-Parkinson-White syndrome. Unlike most cases, blood flow in the sinusoidal communication was antegrade, from the left anterior descending artery to the right ventricle. This is the first report of the combination of pulmonary atresia with intact ventricular septum and Wolff-Parkinson-White syndrome. PMID- 10860290 TI - Surgical angioplasty of ruptured left anterior descending coronary artery without cardiopulmonary bypass. AB - We report a case of successful off-pump surgical angioplasty in the left anterior descending coronary artery using a saphenous vein patch. A 49-year-old male with left main disease and severe cerebrovascular stenosis underwent off-pump bypass to the left anterior descending artery. Postoperative angiography showed anastomotic stenosis where balloon angioplasty was performed. However, it resulted in coronary perforation complicated with epicardial tamponade, and emergency off-pump surgical angioplasty was performed. This case demonstrated that coronary arterial rupture could be managed by surgical angioplasty without cardiopulmonary bypass when coronary artery perforation occurred. PMID- 10860291 TI - Aneurysm in the pulmonary trunk associated with atrial septal defect, a left coronary artery fistula to the pulmonary trunk, and valvular pulmonary stenosis. AB - A 78-year-old woman with an aneurysm in the pulmonary trunk associated with an atrial septal defect, left anterior descending coronary artery fistula to the pulmonary trunk and valvular pulmonary stenosis is reported. The aneurysm showed gradual dilatation over 16 years and was successfully treated using aneurysmorrhaphy. Although there has been some controversy regarding the optimum management for a pulmonary artery aneurysm, surgical correction is thought to be essential for aneurysms associated with congenital cardiac anomalies because of the high incidence of rupture. PMID- 10860292 TI - 61-year-old man with midsternal chest pain, mid-back pain, and lower extremity weakness: a clinicopathologic correlation conference from the University of Oklahoma College of Medicine. PMID- 10860293 TI - Drug seeking behavior--a differential approach. AB - "Drug seeker" is a common term for patients who are perceived to be seeking drugs that might be abused or used in an illegal fashion. While there are patients who seek drugs for totally illegitimate reasons, most of these patients have some type of underlying medical issue. A differential model is proposed using four categories of seeker. The first is fraudulent behavior, second is addictive disease, third is under-treated symptomatology such as pain, and last is psychiatric disorders. Each area is discussed with useful examples of behavior in each category. With a good working knowledge of these areas described, physicians can more effectively manage this difficult group of patients. PMID- 10860294 TI - Effects of smoking on health care costs. AB - Smoking is the leading preventable cause of death in the United States. The US Centers for Disease Control and Prevention (CDC) estimate that smoking kills approximately 419,000 people in the United States each year. Cigarette smoking is the nation's leading cause of premature mortality, and is responsible for one third of all deaths among working-age Americans. Smoking cigarettes is both psychologically and physiologically addictive. Smoking is an important risk factor for cardiovascular diseases, especially coronary artery disease, stroke, carcinoma of the lung, chronic bronchitis, chronic obstructive pulmonary disease, and emphysema. It also increases the risk for peripheral vascular disease and is associated with cancers of the larynx, oral cavity, esophagus, pancreas, and urinary bladder. Smoking by pregnant women can cause adverse health effects on their babies, like low birth weight and preterm delivery; increases the risk of miscarriage; and has also been found to be an important cause of sudden infant death syndrome. Careless smoking also can cause severe burn injuries and death. Many of these adverse effects of smoking occur in "second-hand" smokers. PMID- 10860295 TI - Folic acid knowledge and multivitamin use among Oklahoma women. PMID- 10860296 TI - Comparison of mortality between respondents and non-respondents in a mail survey. AB - We conducted a population-based mail survey and prospectively compared mortality between respondents and non-respondents. Age-adjusted mortality rates for all causes were higher among non-respondents than among respondents in both sexes. Age-adjusted rate ratios were 1.50 for males and 1.33 for females. Non respondents also had, in both sexes, higher mortality for three leading causes of death, namely, cancer, heart diseases and stroke than respondents. In particular, the difference between the two groups was much greater for cardiovascular disease than for cancer. Our results suggested that prospective studies using data from respondents to mail surveys in Japan would have underestimated the mortality for cardiovascular disease. PMID- 10860297 TI - Risk factors for cigarette smoking among high school students in Okinawa, Japan. AB - The purpose of this study was to examine the relationships between risk factors and smoking status among high school students in Okinawa, Japan. We also examined if there is a dose-response relation between the number of risk factors and smoking status. Self-reported questionnaires including smoking status and potential risk factors were conducted using a sample of 1,029 students of a public senior high school in Okinawa. The percentage of smokers was 40.0% for males and 10.6% for females, and it was significantly higher for males than females. As a result of multivariate analysis, we identified four significant risk factors; peer smoking, attitude of peer toward subject smoking, intention to smoke at the age 20, and alcohol drinking. The number of these risk factors was linearly associated with increased percentage of smokers, and a linear trend was significant for both gender students. Additionally, magnitude of risk for smoking among females became considerably great compared with those of males as the number of risk factors increased. In conclusion, this study was the first study in Japan to indicate a significant dose-response relationship between the number of risk factors and smoking status among high school students. We also found that females with many risk factors had extremely increased vulnerability to smoking compared to male counterpart. These findings may be useful to identify high-risk students who need more intensive smoking prevention programs and to develop the content of effective interventions. PMID- 10860298 TI - Relationship between health practices and education level in the rural Japanese population. AB - Past studies in Europe and the USA have found that people with higher education levels have better health practices. The aim of this study was to examine the association between health practices and education level among people in a rural Japanese community. Data were derived from the Ohsaki National Health Insurance Cohort Study, which has been following 52,029 NHI beneficiaries, aged 40 to 79 years, in Miyagi Prefecture, Japan. The relationship between education level and seven health indices (smoking, drinking, body mass index, sleeping, exercise, breakfast, and snacks) was analyzed. Higher education was associated with shorter sleeping hours for both men and women, and lower BMI for women. In age groups younger than 70 years, people with higher education tended to exercise more. Smoking for women, alcohol consumption, and a Health Practices Index were not related to education levels. These results are different from those from Europe and the USA. This study suggest that the relationship between health practices and education level is weaker in Japan than in Europe and the USA. PMID- 10860299 TI - Study of the smoking behavior of medical doctors in Fukui, Japan and their antismoking measures. AB - We conducted a survey on smoking among all members of the medical association in Fukui Prefecture, using a questionnaire to be filled in by the subjects. The survey was conducted from December of 1996 to February of 1997, and the return rate was 90.8%. The main results of this survey were as follows: the prevalence of current smoking among medical doctors was 26.0% (male: 27.8%, female: 5.2%), which was lower than that of adults in the general population. The prevalence of past smoking among doctors 20 to 34 years old by age cohort was highest and that among doctors 35 years old and higher declines as age cohort increased. Doctors' participation in activities for the prevention of smoking in the general society was also found to be at a low level. PMID- 10860300 TI - Apolipoprotein E, methylenetetrahydrofolate reductase (MTHFR) mutation and the risk of senile dementia--an epidemiological study using the polymerase chain reaction (PCR) method. AB - We examined apolipoprotein E (Apo E) polymorphism and methylenetetrahydrofolate reductase (MTHFR) 677 C to T mutation by using the polymerase chain reaction (PCR) method in 100 elderly Japanese aged 60 or more, and assessed whether these genetic factors are associated with an increased risk for the clinical phenotypes of senile dementia, Alzheimer's disease (AD) and vascular dementia (VD) by cross sectional survey. It was found that the Apo E epsilon 4 allele were associated with an increased prevalence of AD as previously reported. Although, it was not strongly related to the severity of senile dementia, a weak association between the ApoE genotype and the severity of dementia was suggested. The proportion of patients with senile dementia was higher in the group of carriers of MTHFR mutation than in the group of noncarriers. Furthermore, the proportion of male patients with senile dementia was higher in the group of homozygous for the mutation (+/+) than the group without the mutation (-/-). Notably in VD patients, 5 of 7 males had the +/+ genotype. The results suggest that the ApoE epsilon 4 genotype and the MTHFR mutation are associated with the clinical phenotype and the clinical onset of senile dementia. PMID- 10860301 TI - Depressive mood and suicide among middle-aged workers: findings from a prospective cohort study in Nagoya, Japan. AB - BACKGROUND: In Japan, mortality from suicide has peaked around 50 years old among men, with increasing trend after 65 years old, and this peak became more apparent in recent years. Beside this, "psychological autopsy" has revealed depression as one of the most important risk factors for suicide. There is, however, no cohort study which examined the relationship between depressive mood measured by simple method and suicide in middle-aged general population. METHODS: In 1989, baseline information was collected by a self-administered questionnaire, and 18,450 workers were followed up to March 31, 1995. All deaths observed during active service were identified, and when retired, its date was recorded. Among 5,352 male workers aged between 40 to 54, 11 committed suicide during follow-up period of 5 years. Analysis were carried out by Cox's proportional hazard model, controlling for age. RESULTS: Those who slept 9 hours or more per night demonstrated 12.14-fold risk of suicide compared with those who slept less than 9 hours. Smokers were more likely to commit suicide than non-smokers. Those who answered affirmatively to more than 7 out of 12 questions, which were derived from Zung self-rating depression scale, experienced an increased risk of suicide (RR 9.95; 95%CI: 1.89-52.44), even after adjusting for other confounding factors. CONCLUSION: We found an association between depressive mood and subsequent suicide in a middle-aged workers. Detailed observation and follow-up of those with depressive mood should be systematically organized with due attention and caution. PMID- 10860302 TI - Factors influencing the visual acuity of primary school pupils. AB - To investigate the factors influencing the visual acuity of primary school pupils, an epidemiological study of 480 pupils in the 6th grade (11-12 years of age) was conducted in 8 primary schools in Sapporo City, Japan. Questionnaires were used to inquire into their current and past visual acuity and related factors. Dividing the subjects into those whose visual acuity of both eyes was 0.7 or more and those whose visual acuity of at least one eye was less than 0.7, odds ratios of various factors were calculated. Lifestyle and dietary factors showed no significant odds ratios. Parental myopia and age of parents at the birth of subjects showed significant odds ratios. Visual acuity of the pupils whose parents were myopic or older than 30 years when they were born tended to have worse visual acuity as they got older. Till the ages 11 or 12, hereditary factors seem much more contributory to visual acuity than environmental ones. PMID- 10860303 TI - Exposure of Japanese school children to smoking-related environmental factors. AB - Japan has no legal restrictions on cigarette advertising and vending machines. This lack of smoking control measures is a possible contributor to smoking initiation by adolescents. This study was conducted to provide primary data on environmental factors related to smoking, such as cigarette advertising and candy cigarettes, that influence elementary school children in Japan. A cross-sectional survey was conducted with a self-administered questionnaire at two elementary schools in Kitakyushu City, Japan in 1995. Questionnaire sheets were anonymously filled out by 282 elementary school children at school. The effective response rate was 91.5% (128 boys and 130 girls). Over 90% of respondents had seen cigarette advertising on TV, candy cigarettes and cigarette vending machines. Over 75% had at least one smoker in their family. Fewer female children expressed an intent to smoke in the future despite the fact that there were no significant sex differences in smoking-related experiences. Children were higher exposed to cigarette advertising on TV, candy cigarettes, vending machines and family members' smoking. Control of such smoking-related factors in the environment would be crucial to keeping children from initiating smoking behavior. PMID- 10860304 TI - Do taught courses on community medicine change knowledge status regarding clinical epidemiology and biostatistics in medical students? AB - This study aimed to explore the changes in medical student's knowledge and attitudes regarding clinical epidemiology and biostatistics (CEB) after community medicine (CM) taught courses. All the 3rd (before CM-taught courses) and 4th year (after CM-taught courses) undergraduate students of Dhaka Medical College, Bangladesh, were given a questionnaire concerning some introductory level problems on CEB and attitudes towards them. Mean knowledge scores were not statistically different between these two groups: 3.70 and 3.85 (out of 9) on clinical epidemiology; 0.20 and 0.18 (out of 4) on biostatistics; and 3.91 and 4.04 as a total (out of 13) among them, respectively. Most of the 3rd and 4th year students agreed that CEB is essential for smooth understanding of clinical medicine and journals, and asserted to include it in CM-taught courses. Since the current CM-curriculum does not offer any improvement of knowledge among them, well-planned taught courses on it should be included as a component of CM. PMID- 10860305 TI - Personality and dietary habits. AB - BACKGROUND: The personality of healthy individuals has not been well studied in relation to health consciousness, dietary habits and actual food intake, simultaneously. OBJECTIVE: Our objective was to study the association between personality and dietary habits. DESIGN: Information on dietary habits, including taste preferences and the frequency of food consumption, was collected through a questionnaire from 76 male and 394 female students. The personality of students was determined by a modified NEO-FFI test. Health status, height, body weight, body fat percentage and blood pressure were measured by physical examination. Main outcome measures were personality scores as indicators of a healthy dietary pattern. RESULTS: Food intake was influenced by neuroticism (N), extraversion (E), openness (O) and agreeableness (A) of personality. Taste preferences and receptivity to dietary advice were also influenced by personality: the odds ratios (ORs) between the high and low tertiary points of the NEO-FFI scores for salty and sweet taste preferences were significantly higher in the group that scored high for neuroticism (N) (salty taste preference: OR = 2.25, NS in males and OR = 2.39, 95%CI = 1.16-4.93 in females; sweet taste preference: OR = 21.00, 95%CI = 2.40-183.99 in males and OR = 3.33, 95%CI = 1.61-6.91 in females). On the other hand, the groups with high scorer for O and A did not like salty tastes. The groups with high scores for A and C were receptive to dietary advice. High scores of each N, E, O, A, and C factor were characterized by distinguishable, dietary habits and lifestyle. For nutritional or health education, group classes are sufficient for high A and O. High C scorer displayed discrepancies between health consciousness and dietary habits, so intervention or a close follow-up by medical professionals would be necessary to improve the health of individuals in this group. High E scorer possessed a confident attitude towards their health, but they were not interested in developing healthy habits. High N scorer was adverse to receiving health information and learning healthy dietary habits. CONCLUSION: Personality determined by NEO-FFI test was related to dietary habits and health attitude. Effective health education methods must take the personality of the targeted individuals into consideration. PMID- 10860306 TI - Why nursing? PMID- 10860307 TI - Education. It's about students. PMID- 10860308 TI - Legal and ethical issues. Diversity in nursing education--Part III. PMID- 10860309 TI - International affairs. Journey around the globe. PMID- 10860310 TI - Insights and inquiry: why would one want to be president of a major association? PMID- 10860311 TI - Comparison of Internet versus lecture instructional methods for teaching nursing research. AB - Although many higher education programs are using the Internet to teach classes, there are few published reports on the effectiveness of this method on test scores or student satisfaction. The purpose of this study was to compare test and student satisfaction scores of graduate nursing students who take a nursing research course via the Internet with those of students who take the same course via traditional lecture instruction. In addition, student technical support use and Internet student lecture attendance also were examined. A total of 97 students (Internet, 44; lectures, 53) participated. There were no significant differences in test scores and overall course student satisfaction (P > .05). However, the Internet students reported significantly higher (P = .04) stimulation of learning compared with the traditional lecture students. Technical support use by the Internet students was high initially and was related to software problems. Of interest were the large proportion of Internet students (73 percent) who attended at least 3 of the 10 lectures. Use of the Internet to teach graduate-level nursing research can provide comparable learning and student satisfaction to traditional lecture instructional methods. PMID- 10860312 TI - IRB-identified ethical issues in nursing research. AB - Ethical issues in nursing research protocols submitted to a School of Nursing institutional review board (IRB) were identified by examining the letters sent to researchers whose protocols required revision or were not-approved. Themes were extracted from the ethical issues and placed into the following categories: The informed consent document, barriers to informed consent, subject benefit, subject risk, confidentiality, and problems with specific populations. The protocols were coded by the researcher's status (student/nonstudent), type of review (expedited/full committee), vulnerability of the population (vulnerable/nonvulnerable), and the type of subject (healthy volunteer/at risk/patient/family/provider). A total of 157 protocols were examined; of these, 45.9 per cent were approved without requiring revision. Revisions were required in 46.5 per cent of the protocols and 8.3 per cent were not-approved. Problems with the form used to document informed consent were found in 43.3 per cent. The next most commonly identified theme was "information needed" in 30.6 per cent of the protocols, followed up by "inadequate explanation of benefit" in 22.3 per cent of protocols. The nursing research protocols in this study were found to be more vulnerable to ethical problems arising from the relationship between the researcher and subject than from physical harm. It is suggested that nurses give extra care to issues of coercion, deception, and attention to problems uncovered in the research process. PMID- 10860313 TI - Supporting nursing students with learning disabilities: a metacognitive approach. AB - As more students with diagnosed learning disabilities (LD) enroll in college nursing programs, nursing educators must increase their understanding of ways to support these students without compromising their academic standards. This article describes a framework for understanding LD including issues in diagnosis, intervention, and accommodation and is designed to assist nursing faculty in maximizing the likelihood of success of these students. Specifically, this article examines the characteristics of students with LD from a metacognitive perspective, identifies curricular issues specific to a typical nursing curriculum, reviews legal obligations under the Americans with Disabilities Act (1990), and provides examples of classroom and clinical modifications including teaching approaches that can benefit all students. PMID- 10860314 TI - An interdisciplinary team model for substance abuse prevention in communities. AB - Recognizing the continuing threat of alcohol, tobacco, and other drug abuse and the mandate for health care reform with emphasis on community-based care and prevention, the University of Texas-Houston Health Science Center School of Nursing developed a model to link faculty to communities to provide culturally competent, scientifically based, preventive interventions. Faculty and community associates engaged in individual and group training activities such as seminars, courses, and off-site meetings. The Preventive Intervention Research Cycle was used to structure prevention activities and assure scientific rigor. In addition to the specific outcomes of five preventive interventions, the project resulted in increased faculty scholarship in the field, increased community awareness and sustained interventions related to substance abuse, enhanced curriculum for students, and expanded collaborations with other community-based organizations. Collaborative interdisciplinary partnerships between academic institutions and community organizations are critical to the development of the science of substance abuse prevention. PMID- 10860315 TI - Nursing drives interdisciplinary health care workforce planning in Wisconsin: one state's experience. AB - The Consortium for Primary Care in Wisconsin convened a forum to develop an interdisciplinary primary care workforce plan to address issues related to the supply of and demand for primary health care providers in Wisconsin. Nursing leaders played a pivotal role in making this effort successful and in ensuring that the focus would be on all primary health care professionals, not just physicians. This process used a primary care workforce planning tool (IRM) developed by the Bureau of Health Professions, U.S. Public Health Service which allowed Wisconsin to (1) examine its own workforce needs with data produced in Wisconsin, (2) compare the state's situation with national trends, and (3) include these data and projections in a cooperative process for state-level planning for interdisciplinary workforce development. The Bureau has encouraged other states and organizations to adopt a similar strategy through a series of IRM workshops in which the Wisconsin process serves as a model for training materials developed for these workshops. The Wisconsin planning process is an innovative model for other states to follow in facilitating workforce development and serves to encourage other states to share their experiences in the academic literature. PMID- 10860317 TI - Oral and maxillofacial pathology. PMID- 10860316 TI - "Enter to learn, go forth to serve": service learning in nursing education. AB - The Pew Health Professions Commission (1998) has recommended community-based service learning as an integral part of nursing education. Service learning activities in a baccalaureate nursing program with an integrated curricular model are described with a women's health elective used as one example of the integration of service learning with clinical experiences. Based on clinical journal entries, students identified benefits of service learning including (1) a sense of personal satisfaction, (2) professional growth, (3) a higher level of critical thinking skills, (4) preparation for nursing practice in a dynamic and diverse health care delivery system, and (5) an increased awareness of unmet needs in clients, families, communities, and populations. Students are truly engaged because they have the opportunity to apply theoretical concepts in giving service, creating a "capacity for connectedness," and learning social responsibility as professionals. PMID- 10860318 TI - Treating bruxism and clenching. PMID- 10860319 TI - Amalgam bonding. PMID- 10860320 TI - Antibiotic prophylaxis. PMID- 10860321 TI - Oral cancer in young adults. PMID- 10860322 TI - Smoking during pregnancy increases risk of cleft lip and palate. PMID- 10860323 TI - Obesity related to periodontal disease. PMID- 10860324 TI - Red wine may reduce oral cancer risks. PMID- 10860325 TI - Practicing dentistry in the age of telemedicine. AB - BACKGROUND: This article examines teledentistry and some of its current legal issues. Topics include licensure, malpractice, technology and ethics. General recommendations for the dental practitioner are included. The literature review includes state and federal laws, the Telemedicine Report to Congress and numerous articles (both printed and electronic) associated with the topic. Sources were selected for timeliness and relevance to legal issues and implications of telemedicine/teledentistry for the dental practitioner. CONCLUSIONS: Numerous issues require resolution before telemedicine and teledentistry will truly realize their enormous potential to increase access to health care while decreasing health care costs. These issues include interstate licensure, jurisdiction and malpractice, as well as technological, security and ethical questions. PRACTICE IMPLICATIONS: Telemedicine and teledentistry are relatively new to the dental field. Many of the legal issues reviewed have yet to be resolved by the legislature or the courts. Furthermore, technology has not yet progressed to the point where the practitioner can be certain that no technological failure will occur during a teledental consultation. In spite of these problems, the potential of telemedicine and teledentistry is tremendous. Improvement in accessibility of health care and lowered health care costs are only two of the many advantages that will emerge as telemedicine and teledentistry become integrated with, and fundamentally change, the practice of medicine and dentistry. PMID- 10860326 TI - Risk of enamel fluorosis in nonfluoridated and optimally fluoridated populations: considerations for the dental professional. AB - BACKGROUND: Few studies have evaluated the impact of specific fluoride sources on the prevalence of enamel fluorosis in the population. The author conducted research to determine attributable risk percent estimates for mild-to-moderate enamel fluorosis in two populations of middle-school-aged children. METHODS: The author recruited two groups of children 10 to 14 years of age. One group of 429 had grown up in nonfluoridated communities; the other group of 234 had grown up in optimally fluoridated communities. Trained examiners measured enamel fluorosis using the Fluorosis Risk Index and measured early childhood fluoride exposure using a questionnaire completed by the parent. The author then calculated attributable risk percent estimates, or the proportion of cases of mild-to moderate enamel fluorosis associated with exposure to specific early fluoride sources, based on logistic regression models. RESULTS: In the nonfluoridated study sample, sixty-five percent of the enamel fluorosis cases were attributed to fluoride supplementation under the pre-1994 protocol. An additional 34 percent were explained by the children having brushed more than once per day during the first two years of life. In the optimally fluoridated study sample, 68 percent of the enamel fluorosis cases were explained by the children using more than a pea sized amount of toothpaste during the first year of life, 13 percent by having been inappropriately given a fluoride supplement, and 9 percent by the use of infant formula in the form of a powdered concentrate. CONCLUSIONS: Enamel fluorosis in the nonfluoridated study sample was attributed to fluoride supplementation under the pre-1994 protocol and early toothbrushing behaviors. Enamel fluorosis in the optimally fluoridated study sample was attributed to early toothbrushing behaviors, inappropriate fluoride supplementation and the use of infant formula in the form of a powdered concentrate. CLINICAL IMPLICATIONS: By advising parents about the best early use of fluoride agents, health professionals play an important role in reducing the prevalence of clinically noticeable enamel fluorosis. PMID- 10860327 TI - Cosmetic oral and maxillofacial surgery options. AB - BACKGROUND: Dentistry and its related specialties have made exponential increases in the functional and cosmetic treatment of the maxillofacial region. Oral and maxillofacial surgeons historically have been involved in functional and cosmetic rejuvenation of the face, and newer technologies have enhanced the ability to make patients look and feel better. METHODS: Cosmetic oral and maxillofacial surgery is being taught in residency programs, is included in the oral and maxillofacial surgery board examinations and represents a part of contemporary oral and maxillofacial surgery. The author discusses common facial rejuvenation procedures with an emphasis on newer treatment technologies. RESULTS: Many oral and maxillofacial surgeons have the ability to improve the esthetics of the maxillofacial area and related structures. The large number of aging baby boomers and technological advances in cosmetic facial surgery have made these procedures easier to perform and more popular than ever. CONCLUSION: A global diagnosis and treatment plan to include facial esthetics can enhance cosmetic dentistry and serve to frame the work of the restorative dentist. The oral and maxillofacial surgeon can help the dentist and patient pursue both functional and cosmetic improvement with safe and effective procedures. CLINICAL IMPORTANCE: All dentists should be aware and abreast of advances in all areas of dentistry and have a basic understanding of available procedures that can benefit their patients. Cosmetic oral and maxillofacial surgery can enhance the work of the restorative dentist and improve facial esthetics, as well as enhance the well-being of the patient. PMID- 10860328 TI - Treating obstructive sleep apnea and snoring: assessment of an anterior mandibular positioning device. AB - BACKGROUND: Dental devices have been used to help manage snoring and obstructive sleep apnea, or OSA. This article reports on patients' compliance with and complications of long-term use of an anterior mandibular positioning, or AMP, device. METHODS: The device used was a custom-made, two-piece, full-coverage, adjustable acrylic appliance, connected with Herbst attachments. The appliance was used nightly and advanced the mandible by 75 percent of the patient's maximum protrusive distance. Patients were telephoned to determine whether they were still using the AMP device. If not, they were asked when and why they stopped using it. The study sample included 65 consecutive patients with mild-to-moderate obstructive sleep apnea and snoring. RESULTS: Long-term use (three years or more) of the AMP device in these patients was 51 percent (27 of 53 patients). Of the 53 responding patients, 40 percent reported jaw/facial muscle pain, 40 percent had occlusal changes, 38 percent reported tooth pain, 30 percent reported jaw joint pain and 30 percent experienced xerostomia. Of the 27 long-term AMP users, 22 rated themselves as being very satisfied and four as somewhat satisfied; one was neither satisfied nor dissatisfied with the appliance. CONCLUSIONS: It was determined that with use of the AMP device, 40 percent of patients will develop some minor complications of jaw, mouth and/or tooth pain, and approximately 26 percent of long-term users might experience a painless but irreversible change in their occlusion. Annual follow-up office visits with the dentist appear necessary for early detection of these changes. CLINICAL IMPLICATIONS: Patients with mild to-moderate OSA who receive a two-piece, adjustable AMP device should be informed that 50 percent of patients quit using the device in a three-year period and some will experience shifts in their occlusion. PMID- 10860329 TI - Differentiating HIV-1 parotid cysts from papillary cystadenoma lymphomatosum. AB - BACKGROUND: Patients with parotid cystic lesions may first be seen in the dental office. These conditions most often represent either papillary cystadenoma lymphomatosum, or PCL, or lymphoepithelial cysts associated with human immunodeficiency virus, or HIV, disease. The authors present a case report to illustrate the differential diagnosis. CASE DESCRIPTION: PCL represents a benign, usually unilateral, circumscribed parotid tumor with cystic elements. HIV associated lymphoepithelial cysts of the parotid gland usually are seen bilaterally, create cosmetic concerns and are hallmarked by an associated cervical lymphadenopathy. Therapy for PCL demands surgical excision, while patients with HIV-associated lymphoepithelial cysts may be treated with antiviral therapy and undergo periodic monitoring by a physician. CLINICAL IMPLICATIONS: As a member of the health care team, the dentist must be familiar with head and neck swellings. Early clinical recognition of parotid swellings leads to successful treatment. PMID- 10860330 TI - A laser-powered hydrokinetic system for caries removal and cavity preparation. AB - BACKGROUND: Laser systems have been developed for the cutting of dental hard tissues. The erbium, chromium:yttrium-scandium-gallium-garnet, or Er,Cr:YSGG, laser system used in conjunction with an air-water spray has been shown to be efficacious in vitro for cavity preparation. METHODS: The authors randomly selected subjects for cavity preparation with conventional air turbine/bur dental surgery or an Er,Cr:YSGG laser-powered system using a split-mouth design. They prepared Class I, III and V cavities, placed resin restorations and evaluated subjects on the day of the procedure and 30 days and six months postoperatively for pulp vitality, recurrent caries, pain and discomfort, and restoration retention. Sixty-seven subjects completed the study. RESULTS: There were no statistical differences between the two treatment groups for the parameters measured with one exception; there was a statistically significant decrease in discomfort levels for the laser system at the time of cavity preparation for subjects who declined to receive local anesthetic. CONCLUSIONS: The Er,Cr:YSGG laser system is effective for preparation of Class I, III and V cavities and resin restorations are retained by lased tooth surfaces. CLINICAL IMPLICATIONS: Hard-tissue cutting lasers are being introduced for use in operative dentistry. In this study, an Er,Cr:YSGG laser has been shown to be effective for cavity preparation and restoration replacement. PMID- 10860331 TI - Disinfection and communication practices: a survey of U.S. dental laboratories. AB - BACKGROUND: The need to disinfect impressions is crucial to prevent the transmission of infectious diseases. The authors report the results of a survey of U.S. dental laboratory directors. The survey was designed to determine how well dental laboratory personnel are communicating with dentists regarding the disinfection of impressions, and, in turn, what laboratory technicians are doing to protect themselves against microbial cross-contamination. METHODS: Four hundred dental laboratory directors were selected in a blinded and random manner. To create a geographically representative sample, an equal number of laboratory directors from the East, Midwest and West were interviewed. A survey consisting of 16 open-ended questions was conducted by trained interviewers via 10- to 15 minute telephone interviews. All dental laboratory directors stated that they were thoroughly familiar with their laboratory's disinfection protocol. RESULTS: The survey documented that the majority of impressions were made of polyvinyl (57 percent) or polyether (27 percent) materials. Only 44 percent of the respondents stated that they knew if the impressions they received had been disinfected. Twenty-three percent of the laboratory directors did not know the method of disinfection used, and 47 percent did not know the length of time involved. Forty five percent of the respondents reported that they receive inadequate instruction in regard to disinfection techniques. No one class of impression materials was found to be more problematic than others by the laboratory directors. CONCLUSIONS: The results indicate a significant and problematic lack of communication between these team members. The responses also suggested that laboratory-perceived problems with impressions were not linked to any particular type of material, but more to the disinfection technique used. PRACTICE IMPLICATIONS: Lack of communication between dentists, staff members and dental laboratory personnel, along with poor training of laboratory personnel in disinfection techniques, may have a direct effect on the prosthetic results achieved in dental practices. PMID- 10860332 TI - Coronary artery stents: review and patient-management recommendations. AB - BACKGROUND: Coronary artery stents are metallic scaffold devices that physically support narrowed coronary arteries to alleviate symptoms of ischemic coronary artery disease. They are placed during invasive procedures similar to that of percutaneous transluminal coronary angioplasty, and patients are maintained with antiplatelet medications to lessen the chances of stent stenosis. METHODS: The authors provide a brief overview of coronary artery stents and discuss the dental management of patients who have received stents. CONCLUSIONS: After stent placement, patients usually are maintained with antiplatelet regimens, which may necessitate choosing medications that do not potentiate their effects. Any discussion as to the possible need for antibiotic prophylaxis of patients with stents largely is missing from the literature. Recent literature, however, indicates that antibiotic prophylaxis, if required, may only be needed during the first few weeks after stent placement. CLINICAL IMPLICATIONS: Dental professionals should become knowledgeable about coronary artery stents. Although these devices have a higher success rate than other procedures in alleviating symptoms of ischemic coronary artery disease, some patients are still at risk of experiencing significant cardiac events. PMID- 10860333 TI - The inevitable maladies of the mature dentition. AB - Patients who are in their mature years should be advised about the numerous ways in which teeth and supporting structures degenerate in the later years of life. This knowledge will allow patients to make special efforts to prevent, reduce, eliminate or treat the oral problems associated with aging. PMID- 10860334 TI - Maturity and oral health: live longer and better. PMID- 10860335 TI - Alternative method for making ideal contacts when placing direct posterior composite resin. PMID- 10860336 TI - Placing the single-tooth, screw-retained implant prosthesis. PMID- 10860337 TI - Presenting the California Dental Association's case. The oral argument in CDA vs. Federal Trade Commission. PMID- 10860338 TI - Patients' expectations for oral health care in the 21st century. AB - BACKGROUND: This article examines trends in patient demographics and dental disease patterns. Data suggest the patient expectations about oral health are increasing, as is their knowledge of oral health services. CLINICAL IMPLICATIONS: Changing patient demographics and technological advances will lead to higher patient expectations and greater demands for oral health care in the 21st century than they had been during most of the 20th century. PMID- 10860339 TI - Trends in preventive care: caries risk assessment and indications for sealants. AB - BACKGROUND: In the 21st century, risk assessment models will continue to be developed. By understanding patients' susceptibility to disease, better treatment and preventive regimens can be offered. As the causative agent of dental caries is bacterial, the interaction between the susceptible host, the causative agent and the environment determine whether caries occurs--regardless of the patient's age. CLINICAL IMPLICATIONS: This article reviews risk assessment for dental caries and the implication for developing preventive strategies. It also describes the indications and uses of sealants in the prevention of dental caries. PMID- 10860340 TI - Rationale and treatment approach in minimally invasive dentistry. AB - BACKGROUND: Current methods of detecting caries, especially fissure caries, are inaccurate, causing some caries to go undetected until it has reached more advanced stages. Minimally invasive dentistry is a philosophy in which the goal of intervention to conserve healthy tooth structure. The authors review the rationale and role of air abrasion in successful practice in the 21st century that includes the philosophy of minimal intervention. CLINICAL IMPLICATIONS: This objective encompasses a range of clinical procedures that includes assessment of caries risk to reinforce patient self-help, early detection of the disease before lesion cavitation to fortify the oral environment, restoration of fissure caries with maximum retention of sound tooth structure and sealant placement in unaffected areas. This conservative approach minimizes the restoration/re restoration cycle, thus benefiting the patient over a lifetime. PMID- 10860341 TI - The science of bonding: from first to sixth generation. AB - BACKGROUND: Adhesive dentistry has revolutionized restorative dental practice during the past 30 years. Improved adhesive materials have made resin-based composite restorations more reliable and long-standing. CLINICAL IMPLICATIONS: This article reviews the evolution of bonding from the first generation to current bonding materials. It discusses the composition and effectiveness of the various iterations. Current products are highlighted to improve clinical use and performance of the materials. PMID- 10860342 TI - The spectrum of composites: new techniques and materials. AB - BACKGROUND: During the past 25 years, advances in adhesive technology and composite-based resins have provided dentists and patients with new treatment options. This technology provides patients with more tooth-conserving and highly esthetic restorations. CLINICAL IMPLICATIONS: This article reviews advances in composite-based resin materials. It discusses composition and classification of current resin-based composite. It also reviews techniques for successful placement of these materials and provides a discussion of current concepts of polymerization. PMID- 10860343 TI - Trends in surgical and nonsurgical periodontal treatment. AB - BACKGROUND: New research is demonstrating that a person's total health is indeed related to his or her oral health. Elimination of all oral infections, including gingivitis and periodontis, is important to overall health. CLINICAL IMPLICATIONS: This article reviews recent evidence on the systemic and oral connection and discusses these findings as they relate to patient care. The article examines trends in nonsurgical and surgical therapy that will successfully arrest periodontal infections. Opportunities for early diagnosis and prevention will play an increasing role in dental practice in the future as patients understand the importance of oral health to overall health. PMID- 10860344 TI - 21st-century endodontics. AB - BACKGROUND: Endodontics as a discipline has offered patients the opportunity to maintain their natural teeth. As the population expands and ages, the demand for endodontic therapy can be expected to increase as patients seek dental options to keep their teeth for a lifetime. CLINICAL IMPLICATIONS: New materials, techniques and instruments are entering the market-place to assist dentists in providing patients with more predictable and reliable endodontic treatment. In addition, these new systems make the delivery of endodontic services more efficient. This article describes these advances in endodontic treatment for dentists interested in incorporating these advances into their clinical practice. PMID- 10860345 TI - Porcelain esthetics for the 21st century. AB - BACKGROUND: Dental procedures play a vital role in the modern dental practice. Considerable research has addressed improvements in the properties of dental porcelains. CLINICAL IMPLICATIONS: This article examines the trends in the scientific advances in dental porcelains. It highlights properties of the new low fusing porcelains and describes indications for their use. New luting cements also are addressed. PMID- 10860346 TI - Current trends in removable prosthodontics. AB - BACKGROUND: This article discusses trends in the demographics and treatment of the edentulous patient. It is clear that there still is a tremendous need for removable-prosthodontic services today. While the basic process of making dentures has changed little over the past several decades, new materials and techniques can help laboratories and clinicians provide functional, esthetic restorations that offer exceptional value to patients. Implant treatment is a tremendous adjunct to removable prosthodontics in the treatment of edentulous patients, but it is not within the financial reach of all dental patients. CLINICAL IMPLICATIONS: The clinical skills required to deliver excellent complete denture care are also paramount to successful implant prosthodontics (fixed and removable) and esthetic dentistry. Even so, the opportunities to develop these skills and the interest appear to be decreasing at the same time that the need is projected to increase. In service to our patients, the profession must examine this trend closely. PMID- 10860347 TI - The clinical significance of the digital patient record. AB - BACKGROUND: Computer technology has revolutionized the way the world does business, allowing us to work faster, smarter and more efficiently than ever before. Computers first made their way into the dental office in the late 1960s as an accounts receivable device. Today, we can digitize anything and recall it in the operatory with the patient. CLINICAL IMPLICATIONS: This article discusses new trends in the digital patient record and the benefits this technology provides to the dental team in terms of improved data collection and recording. It also discusses the benefits a digital patient record provides to patients, as well as how to communicate patients' oral health needs using these electronic tools. PMID- 10860348 TI - [Feasibility study of CdTe Semiconductor detector for gamma camera--evaluation of planar images]. AB - To evaluate the performance of a semiconductor detector for use in a gammacamera system, we assembled a detector with a small field of view--1 inch x 1 inch and 1 inch x 2 inch--made from CdTe (Cadmium telluride). We then compared the planar images and energy resolution of the resulting detectors against those of a conventional gammacamera. Pixel pitch of the detector was 1.6 mm x 1.6 mm, and was manufactured by Acrorad Corporation. Average FWHM of the energy spectrum for the semiconductor detector was 5.11% (SD: 0.80%, Best: 3.26%, Worst: 6.68%). The planar images obtained were of a letter phantom made from pieces of lead and of an IMP brain phantom. Since the field of view of the semiconductor detector was small, the image of the IMP brain phantom was acquired by moving the semiconductor over the collimated detector module until the area of the entire phantom was covered. The images from the semiconductor assembly were compared with those from a conventional gammacamera using the same conditions, and it was found that visual image quality was superior to those of the conventional camera system. PMID- 10860349 TI - [The studies of hemodynamic changes and liver uptake in a combination of ATP stress test and low workload exercise test on myocardial scintigraphy]. AB - A pharmacological adenosine-tri-phosphoric acid (ATP) stress test has been used in patients who can not perform an enough exercise stress test. However, falling blood pressure during the stress test and increased liver uptake of the tracer are often found in patients undergoing the ATP test. To prevent these phenomena, a combination of ATP stress test and low workload exercise test (ATP & EX) is proposed. The usefulness of this newly developed stress test was elucidated from two viewpoints. Firstly, the changes of hemodynamic parameters were measured in 34 patients: 17 undergoing ATP alone and 17 undergoing ATP & EX. Systolic blood pressure fell from 150 +/- 20 mmHg to 126 +/- 16 mmHg (p < 0.05) for ATP alone. However, it changed from 141 +/- 19 mmHg to 149 +/- 31 mmHg (ns) for ATP & EX. There was a significant fall in systolic blood pressure (> 30 mmHg) in 58.8% for ATP alone and 5.9% for ATP & EX (p < 0.01). Secondly, the ROI count in the liver and heart on an anterior projection image were measured in 38 patients: 11 undergoing ATP alone, 13 undergoing ATP & EX, and 14 undergoing an ergometer exercise test (EX). The ROI count in the liver at 60 minutes after tracer injection were 29.0 +/- 10.7 count/pixel, 21.4 +/- 5.2 count/pixel, 18.3 +/- 4.5 count/pixel for ATP alone, ATP & EX and EX, respectively. The activities for ATP & EX and EX were lower than that for ATP alone (p < 0.05 and p < 0.01). Thus, ATP & EX decreased the rates of the fall of systolic blood pressure and decreased liver uptake of the tracer compared with ATP alone. In conclusion, ATP & EX is a useful stress method for myocardial perfusion scintigraphy in patients who can not perform the enough exercise stress test. PMID- 10860350 TI - [Transient left ventricular dysfunction is still present one hour after exercise stress test: evaluation by gated SPECT with 99mTc-labeled perfusion agent]. AB - It has been reported that quantitative gated SPECT (QGS) has revealed post-stress dysfunction of the left ventricle (LV) 30 minutes after a stress test. The purpose of this study was to determine whether post-stress dysfunction of LV is still present one hour after an exercise stress test. The subjects comprised 152 patients (124 males and 28 females, mean age 59 +/- 10 years). Exercise stress myocardial scintigraphy was performed using a one-day, stress and rest protocol. 99mTc labeled myocardial perfusion tracer, tetrofosmin, 370 MBq was injected at the end-point of a supine ergometer stress test for stress imaging. ECG gated SPECT was carried out 1 hour after injection. Three hours later, 740 MBq to 1100 MBq of 99mTc labeled myocardial perfusion tracer was injected for rest imaging. ECG gated SPECT was again performed 1 hour after injection. We divided the subjects into four groups according to the severity score of defects on the stress image and the presence or absence of fill-in; normal (NOR, n = 59), myocardial infarct (MI, n = 65), small ischemia (S-IS, n = 13) and large ischemia (L-IS, n = 15). Post-stress dysfunction is defined according to two criteria: 1) rest LVEF--post-stress LVEF > or = 5% and 2) post-stress ESV--rest ESV > or = 5 ml. The frequency of post-stress dysfunction was 3.4%, 9.1%, 23.1% and 40% in NOR, MI, S-IS and L-IS, respectively. Post-stress LV dysfunction was found to be more frequent in the large ischemia group. In conclusion, post-stress dysfunction was present 1 hour after the stress test and was more frequent in the large ischemia group. PMID- 10860351 TI - [Clinical usefulness of delayed exercise images on 99mTc-Tetrofosmin myocardial SPECT in the diagnosis of vasospastic angina pectoris]. AB - This study was designed to evaluate the clinical usefulness of delayed exercise images in 99mTc-tetrofosmin (TF) myocardial SPECT in the diagnosis of vasospastic angina pectoris. We studied 30 patients with vasospastic angina, 10 of 30 patients (group A) had both effort and rest angina, 20 of 30 patients (group B) had rest angina. A 370 MBq of TF was intravenously injected at peak exercise, and initial (EX-I) and delayed exercise (EX-D) images were obtained at 30 min and 180 min after the injection. An additional 740 MBq of TF was intravenously reinjected after EX-D image acquisition, and rest images were obtained 30 min after the reinjection. The left ventricular wall was divided into 9 segments. Regional myocardial uptakes of TF were scored by 4-point defect score (0 = normal, 1 = mildly reduced, 2 = moderately reduced, and 3 = severely reduced). Total defect score (TDS) was calculated from the sum of defect scores in 9 segments. Reverse redistribution (RR) was defined as increase of more than 2 in TDS on EX-D images. In group A, 4 of 10 cases (40%) showed decreased uptake on EX-I images, 6 of 10 cases (60%) revealed RR on EX-D images, and none of the patients showed decreased uptake on rest images. In group B, no one showed decreased uptake on EX-I and rest images, 11 of 20 cases (55%) revealed RR on EX-D images. The mean +/- SD of TDS were 2.9 +/- 3.4, 5.1 +/- 4.5, 0.5 +/- 0.5 on EX-I, EX-D, rest images in group A, and serially 0.4 +/- 0.5, 3.3 +/- 3.6, 0.4 +/- 0.5 in group B. Regional wall motion abnormality was reduced in regions with RR. RR on EX-D images may reflect ischemic damaged but viable myocardium in vasospastic angina. The clinical usefulness of exercise-rest TF imaging in detection of organic coronary artery disease has been well established. Therefore, exercise-rest TF imaging with additional delayed exercise image could evaluate not only organic coronary artery disease but also coronary artery vasospasm. PMID- 10860352 TI - [Assessment of most appropriate background subtraction method for quantification of 123I-metaiodobenzylguanidine (MIBG) myocardial uptake by comparing with plasma ANP and BNP]. AB - BACKGROUND: Quantification of 123I-metaiodobenzylguanidine (MIBG) myocardial uptake is widely accepted as a useful tool for estimating the severity of congestive heart failure. However, most reliable method has not been determined yet because of the difficulty of background (BG) subtraction. In this study, the most appropriate BG subtraction method was evaluated as compared with plasma atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), norepinephrine (NEP) and left ventricular ejection fraction (LVEF). METHODS: Twenty-one patients with chronic heart failure were examined. After bolus injection of 123I-MIBG (111 MBq), dynamic anterior chest images were collected every second for 2 minutes. Planar anterior chest images were obtained 15 minutes (early image) and 4 hours later (delayed image) respectively. The parameters for quantification of 123I MIBG myocardial uptake used in this study were heart to mediastinum ratio (H/M), myocardial washout rate and uptake ratio. Background was calculated using the region of interest (ROI) placed over a part of upper mediastinum, whole upper mediastinum, right lung and pericardial space respectively. The values of these parameters were calculated with and without BG subtraction and compared with plasma ANP, BNP and others. RESULTS: H/M did not correlate with ANP or BNP. Myocardial washout rate without BG subtraction showed significant correlation with ANP (p < 0.01) and BNP (p < 0.05). After BG subtraction employing ROI placed over the part of upper mediastinum and whole upper mediastinum, myocardial washout rate showed better correlation with BNP (p < 0.01). Myocardial uptake ratio did not showed any correlation with ANP or BNP without BG subtraction. However, myocardial uptake ratio showed significant correlation with BNP (p < 0.05) after subtraction of upper mediastinal BG. BG subtraction using ROI over right lung or pericardial area revealed poor results in both myocardial washout rate- and uptake ratio. CONCLUSION: BG subtraction using ROI over the upper mediastinum is likely to be suitable for quantitative analysis of 123I-MIBG myocardial scintigram. PMID- 10860353 TI - [Influence of the radioactive strontium (89Sr) using for nuclear medical radiation therapy upon radioactive draining-water system]. AB - PURPOSE: Strontium-89 chloride (89Sr) is a new radiopharmaceutical that provides effective pain relief for metastatic bone lesions, and is expected to be available soon in the palliative management for metastatic bone pain in Japan. Because of relatively long physical half life (50.5 days), 89Sr may affect to the radioactive draining-water system by exceeding the limits of activity concentration for radioactive drain. In this article, the influence of 89Sr use on the radioactive drainage system was simulated. METHODS: The standard tank capacity of drainage and draining frequency was determined from the results of questionnaire carried out for the nationwide medical and research institutes where radioisotope treatment are performed. On the assumption that 89Sr of 148 MBq for one therapy was used twice a week and several common radionuclides were used as the same activity as used at Chiba Cancer Center, the influence of 89Sr was estimated. The calculation was performed using the activity contamination ration into the draining-water system of each radionuclide of 0.01, which was legally determined. RESULTS: The simulation revealed that the sum of the contamination ratios of individual radionuclides exceeded a legal value of 1.0 in standard drainage with the capacity of 5 m3 and 10 m3 and draining frequency of 7 times per year. The actual contamination ratios of common radiopharmaceuticals measured at Chiba Cancer Center ranged from 1/100 to 1/1000 of the legal values. CONCLUSION: It is necessary that the legal value of activity contamination ratios into the draining-water system should be reassessed before starting 89Sr therapy. PMID- 10860354 TI - [The twenty-first report on survey of the adverse reaction to radiopharmaceuticals (the 24th survey in 1998). Subcommittee of Safety Issue for the Radiopharmaceuticals. Medical and Pharmaceutical Committee. Japan Radioisotope Association]. PMID- 10860355 TI - [Basic study on anti-bacterial urethral catheter. I. Development of a new anti bacterial coating material for silicon catheters]. AB - In order to develop a new anti-bacterial urethral catheter, we studied anti bacterial and anti-adherent coating material suitable for silicon catheters. Several aspects of various silver compounds were examined, including anti bacterial activity, chemical property and toxicity. Among silver citrate, silver phosphate and silver oxide, which were found to have excellent anti-bacterial activities, silver citrate was regarded as the material of choice for anti bacterial coating in terms of durable activity and biological safety. It was also found that several surfactants inhibited bacterial adherence to the surface of silicon catheters. Among them soybean lecithin exhibited excellent anti-adherent activity in a dose dependent manner. Finally, a mixture of silver citrate, soybean lecithin and liquid silicon at the ratio of 2:2:8 was regarded as an ideal anti-bacterial coating material for silicon catheters. PMID- 10860356 TI - [Basic study on anti-bacterial urethral catheter. II. Potency of a new anti bacterial catheter and its durability in experimental models]. AB - The potency and effectiveness of an anti-bacterial catheter coated with a mixture of silver citrate, soybean lecithin and liquid silicon at the ratio of 2:2:8 were compared with those of commercially available anti-bacterial and conventional urethral catheters. This new anti-bacterial catheter showed a strong activity and excellent durability in ordinary in vitro experimental studies. In the present series we have developed new in vitro experimental models for the evaluation of anti-bacterial catheters in inhibiting bacterial ascent via intraluminal or extraluminal route. The characteristic features of the silver citrate/lecithin catheter, namely strong activity and excellent durability, were confirmed using these new models that mimic urinary catheter-associated clinical infections. PMID- 10860357 TI - [Detection of genotypic and phenotypic drug-resistant HIV-1 in patients receiving antiretroviral therapy]. AB - We have analyzed the sequences of HIV-1 reverse transcriptase and protease genes in peripheral blood mononuclear cells obtained from patients receiving antiretroviral therapy to evaluate the drug resistance-associated mutations. Of 84 HIV-1-infected individuals treated with reverse transcriptase inhibitors, 43 (51.2%) have been found to carry amino acid substitutions predicted to acquire drug-resistances. One to 3 mutations at amino acid residues reported to be associated with protease inhibitor-resistance were detected in more than 80% of protease inhibitor-naive patients. However, these pre-existing mutations did not seem to raise a real resistance after the initiation of therapy with protease inhibitors. Phenotypic resistance assay was performed with 6 clinical isolates to compare with genotypic resistance. In most of the cases, phenotype was correlated with genotypic changes, however, two strains which were isolated from patients having no experience of chemotherapy showed a decrease in susceptibility to several drugs without any resistance-related mutations detected in their genes. Taken together, determination of phenotypic resistance is necessary, especially when a newly-infected patients starts antiviral therapy. PMID- 10860358 TI - [Retrospective studies of penicillin-resistant pneumococcal acute otitis media in infants and children--epidemiological study]. AB - Retrospective studies of the prognostic factors of infantile acute otitis media caused by Penicillin-resistant Streptococcus pneumoniae were performed. In this study, the following items were discussed the following items; age, sex, presence of nurturance, presence of acute otitis media, presence of sibling, presence of past antibiotic therapy, presence of past therapy for this type of otitis media, MIC of PCG, MIC of CDTR. 1. Statistically significant results were obtained in the following items; age, presence of acute otitis media, presence of past antibiotic therapy, presence of past therapy of this type of otitis media (especially that treated by an otolaryngologist). 2. In previous reports, persistent and recurrent otitis media was influenced by participation in kindergarten groups. However in this study, a statistically significant difference was not recognized. 3. Statistically significant results were not recognized in the MIC of PCG and the MIC of CDTR for PRSP. 4. In this study, immunities of each infants were not discussed, but it was necessary for us to examine the factors of immunity in infants, such as specific pneumococcal antigen and type-specific IgG2 antibody and comparison of such titers to the various results of treatment. PMID- 10860359 TI - [Measures against hospital infection for students in medical schools in Japan--a survey by questionnaires]. AB - We sent questionnaires to 80 medical schools in Japan to know what the problems in hospital infection for students in medical schools are. Seventy-one % were sent back to us. There have been hospital infection from patients to students in 12% of the medical schools, included 3 cases each of measles, chickenpox and mumps, 2 cases each of rubella, hepatitis B and tuberculosis. Fourteen % of the medical schools had reports about the past history of infection and vaccination from students, 70% of the schools determined their antibodies and the 28% did nothing. Ninety-three%, 25%, 23%, 18%, and 15%, of the schools determined antibodies for hepatitis B, rubella, measles, chickenpox and mumps, respectively. Some assays of the measurement were low in sensitivity. The cost for the determination was fully paid in 48% of the schools, but only partially supported in 35%. Tuberculin reaction was performed in 40% of the schools and then BCG was done in 57%. Vaccination was recommended in 40% of the schools. The cost of vaccination was all paid in 38% of them, which was only for hepatitis B, and partially supported in 15% of them. PMID- 10860360 TI - [Genetic characterization of Norwalk-like viruses (NLVs) detected in Japan from 1989 to 1998 and establishment of consensus primers]. AB - By means of reverse transcription-polymerase chain reaction using various pairs of primers, 902 strains of Norwalk-like viruses (NLVs) were detected during 10 years from 1989 to 1998 in the nationwide area of Japan including Hokkaido and Okinawa. Nucleotide sequencing was done on 177 strains, and we found that 153 strains (86%) of them belonged to geno-group II (GII). GII could be classified into five subgroups (G2A, G2B, G2C, G2D and G2E) based on the amino acid sequences showing a similarity of more than 91%. Among them G2C and G2D (named JP1) as well as G2E (named JP2), which were occasionally identified in Japan (detection rate of 17%) but rarely detected in the other countries, revealed to form new subgroups. From the above analysis, three pairs of new primers were established as consensus primers for NLVs. All the three sets of primers, P1/P2,P1/P3 and Y1/Y2, amplified both GI and GII equally and efficiently. PMID- 10860361 TI - [Amantadine usage for influenza A during an influenza outbreak in a nursing home]. AB - An outbreak of an influenza like illness was found in a nursing home in Fukuoka in January, 1999. Results of hemagglutinin inhibition tests with paired sera of patients and rapid diagnosis kit for influenza A indicated that an influenza A (H3N2) outbreak had occurred. A total of 15 patients with influenza like illness from one residential area of the nursing home were administered amantadine, 100 mg per day for five days. Clinical records of 264 residents were surveyed retrospectively from the tenth to the thirty-first of January, 1999. Influenza like illness was found in 112 residents (42.4%). The incidence of influenza like illness differed by residential area, ranging from 27.6% to 54.0%. The mean duration of fever was 3.6 days among patients administered amantadine. The mean duration was 4.4 days for patients not administered amantadine. The incidence of influenza like illness decreased rapidly after amantadine administration in the residential area where amantadine administration was done. These results suggest that amantadine is effective in mitigating influenza symptoms in the elderly. Amantadine may be useful for diminishing the influence of influenza A outbreaks in nursing homes. PMID- 10860362 TI - [Epidemiological analysis of influenza B virus belonging to B/Victoria/2/87 lineage isolated in off-season of 1998 and late epidemic season in Shizuoka Prefecture]. AB - In Shizuoka Prefecture, influenza B viruses belonging to B/Victoria/2/87 lineage caused an outbreak among school children in the off-season of 1998, and the same B viruses were mainly isolated during the following epidemic. Low titer of HI antibody among children for influenza B virus belonging to the same lineage was a recognized factor in the causation of the distinctive epidemic. The herald virus strains seemed to be antigenically similar to the late epidemic virus strains, but the former strains were not genetically close to the latter. This indicates that the herald viruses are not always the parental viruses for the following influenza season. PMID- 10860363 TI - [Emphysematous cystitis and neuropathy; a report of the case with diabetes mellitus]. AB - A study was made of a 55 years old male, who suffered from emphysematous cystitis with diabetes mellitus. He had multiple complications due to diabetic neuropathy such as foot ulceration, oculomotor nerve palsy, peroneal nerve palsy and a neurogenic bladder. Klebsiella pneumoniae and Pseudomonous aeruginosa were cultured from urine specimens. There have been only 19 reported cases of emphysematous cystitis since 1962. Fourteen of these cases had diabetes mellitus. PMID- 10860364 TI - [A rapidly fatal case of severe falciparum malaria complicated with high-level metabolic acidosis]. AB - A 67-year-old male was admitted with consciousness disturbance (JCS, III-200) after completing a 12-day tour to east Africa without malaria chemoprophylaxis. When he visited the hospital one day prior to the admission complaining of fever and a slightly sore throat, he did not mention the travel history. Soon after his travel history was revealed, blood films were prepared which showed abundant ring forms accompanied with a small number of trophozoites and schizonts of Plasmodium falciparum, with the parasitemia of 26%. Despite intravenous quinine infusion, first that of loading dose, his consciousness state (JCS, III-300), renal and hepatic functions and anemia (Hb, 5.8 g/dL) deteriorated progressively. Moreover, metabolic acidosis worsened with pH of 6.954, HCO3- of 3.4 mEq/L, BE of--27.0 mEq/L, PCO2 15.5 mmHg by arterial blood gas analysis, although he received a large volume of sodium bicarbonate solution. The patient died on the 4th day of his illness. According to the literature, it is suggested that the treatment of metabolic acidosis in severe faciparum malaria with sodium bicarbonate is sometimes harmful, since it can result in sodium overloading, which may then precipitate pulmonary edema/ARDS. However, alternative treatment regimens have not yet been established. Future investigation on the etiology and the proper treatment of metabolic acidosis associated with severe falciparum malaria is highly needed. PMID- 10860365 TI - Governor's address to MSNJ House of Delegates. PMID- 10860366 TI - Harold T. Shapiro, PhD. PMID- 10860367 TI - Investing tobacco settlement monies with care. PMID- 10860368 TI - Suggestions for the House of Delegates. PMID- 10860369 TI - Radiation risks of high-dose fluoroscopy. AB - Radiation-induced skin injury is an underdiagnosed, significant complication for patients undergoing fluoroscopy-guided interventional procedures. With proper equipment, fluoroscopic technique, and physician education, patient radiation exposure can be decreased by 75% or more and skin injuries can be minimized. PMID- 10860370 TI - Medicare's evaluation and management codes. PMID- 10860371 TI - Dr. Hugh Mercer: physician and soldier. PMID- 10860372 TI - Senior health issues. AB - Thanks to improved living conditions, better nutrition, preventive care, increased fitness, antibiotics, and other factors, more Americans will remain healthy through their 80s and beyond. As people live longer, however, so do the demands for services and products that respond to their needs. Will the medical field be prepared? PMID- 10860373 TI - A longitudinal study of orthodontic treatment need in Dunedin schoolchildren. AB - The Index of Orthodontic Treatment Need (IOTN) was used to assess unmet orthodontic treatment need in 152 13-year-old Dunedin schoolchildren, and to compare the findings with those obtained in the same children 3 years previously. The children were randomly selected from Dunedin schools as 10-year-olds, and had not received orthodontic treatment. Approximately 86 percent of the 13-year-old children had "No-little" need for orthodontic treatment when assessed by the child-assessed Aesthetic Component (AC) and the examiner-assessed AC. Slightly less than half the children had "No-little" need for orthodontic treatment when assessed with the Dental Health Component (DHC). More 10- and 13-year-old children "Needed" orthodontic treatment with the DHC than with the AC. Both the examiner-assessed AC and the DHC assessed significantly fewer 13-year-olds as needing orthodontic treatment than the same children as 10-year-olds. Complete agreement between the grades assigned at 10 and 13 years occurred in 30-43 percent of the children and, in the treatment-need categories, between 53 percent (DHC) and 84 percent (child-assessed AC) of the children. The fall in treatment need over the 3-year period may be due to selection bias, over-sensitivity of the IOTN to mixed dentition traits, or both. Although a number of 10-year-old children were assigned different grades as 13-year-olds, many remained within the same treatment category. The apparent stability of the IOTN to assess treatment need in 10- and 13-year-old children is attributed to the grouping of different occlusal traits in the same treatment-need category, and to the small number of treatment-need categories in each component. PMID- 10860374 TI - Jaw and tooth abnormalities detected on panoramic radiographs in New Zealand children aged 10-15 years. AB - Panoramic radiographs of 1,608 children and adolescents aged 10 to 15 years (797 males and 811 females) were reviewed to determine the prevalence of tooth and jaw abnormalities. Abnormalities were detected on 21 percent of the radiographs (23 percent females and 17.3 percent males); 879 teeth were diagnosed with abnormalities on 331 radiographs. The more common abnormalities were malpositioned teeth, missing teeth, misshaped teeth, and teeth with hypoplastic appearance. Bony abnormalities and growth problems were detected in a few radiographs. This study demonstrates the value of panoramic radiography in detecting or confirming dental abnormalities, and supports recommendations on the use of panoramic radiography to aid in the assessment of dental development. PMID- 10860375 TI - Dental care for children under general anaesthesia by private dental practitioners in New Zealand. AB - An overall reduction of approximately one-third in the availability of private dental care under general anaesthesia in New Zealand has occurred in the past 5 years. Private dentists providing dental care under general anaesthesia are disproportionately located in Auckland. Specialist anaesthetists or general medical practitioners are used to provide almost all the general anaesthetics; approximately half the dentists providing this service continue to use their dental surgeries for the procedure. Private dentists provide approximately one third of the dental care under general anaesthesia for children each month in New Zealand, but utilise a greater number of sessions per month than the public sector hospitals. Fees associated with dental care under general anaesthesia for children provided by private dentists are predominantly privately funded. Barriers to dental care for children provided by private dentists are primarily cost, difficulties for the dentists and anaesthetists to fit a general anaesthetic session into the practising day, and difficulties providing care for children under 3 years of age and for those with medical problems and disabilities. PMID- 10860376 TI - Implant site development using orthodontic extrusion: a case report. AB - One of the most important factors in the successful placement of endosseous implants is the presence of adequate alveolar bone at the recipient site. Alveolar bone loss associated with destructive periodontal disease frequently results in osseous defects that may complicate subsequent implant placement. Typically, such defects are treated prior to or at the time of implant surgery using the principles of guided bone regeneration. Under certain circumstances, however, such defects may be managed non-surgically by orthodontic extrusion. Orthodontic extrusion can be used to increase the vertical bone height and volume and to establish a more favourable soft-tissue profile prior to implant placement. The addition, the increase in the vertical osseous dimension at interproximal sites may assist in the preservation of the interdental papillae and can further enhance gingival aesthetics. This report illustrates the treatment sequence for site development with orthodontic extrusion prior to immediate implant placement. PMID- 10860377 TI - Clinical applications of new advances in occlusal caries diagnosis. AB - Because of the difficulties associated with the diagnosis of occlusal caries, many clinicians have adopted the philosophy of "watch and wait". The decision to treat, be it by sealing or cavity preparation, is often made after the caries process is well established, and either bonding techniques fail, or unnecessary sound tooth structure is lost. The current diagnostic model of visual, probe, and radiographic examination is qualitative, subject to operator interpretation, and consequently can produce varied diagnoses from dentists examining the same patient. Recent advances in caries diagnosis and an understanding of the caries process, fissure morphology, and bonding principles allow early intervention. Mirror and probe examination is only 25 percent accurate in detecting early occlusal caries. The use of caries detection dye and laser caries diagnosis raises diagnostic accuracy beyond 90 percent. Hidden caries is now an historic phrase. Early and accurate diagnosis of occlusal caries enables successful prevention and minimal intervention restorative techniques, ending the common evolution from occlusal restorations through to cusp restorations, crowns, and endodontics. PMID- 10860378 TI - Epidemiology in practice. PMID- 10860379 TI - [Roles of magnetic resonance imaging in management of bone tumors]. AB - The roles of magnetic resonance imaging (MRI) in the diagnosis and treatment of bone tumors are reviewed. Most bone tumors can be detected on plain radiography or bone scintigraphy. MRI is helpful in detecting tumors that do not destroy bone matrix or suppress reactive bone formation. Detailed analysis by plain radiography is still the most reliable method for differentiating between benign and malignant bone tumors. The T1 and T2 values, internal texture, and peritumoral edema depicted on MRI are not helpful for this differentiation. In characterizing the histologic types of bone tumors, MRI is of some advantage. For example, MRI can demonstrate cartilage matrix, hemoglobin metabolites, vascular components, and fat contents more clearly than conventional radiological techniques. MRI is now indispensable for the preoperative delineation of malignant bone tumors, because of its excellent soft tissue contrast and multiplanar imaging capability. In this article, the guidelines for evaluation of the surgical margin advocated by the JOA Musculo-skeletal Tumor Committee are introduced for radiologists. MRI monitoring of malignant bone tumors after chemotherapy or surgery can reveal change in the size of enhanced areas that may reflect viable tumors. Dynamic MRI is helpful to differentiate recurrent tumors from granulation tissue. PMID- 10860380 TI - [Imaging of tumors of the spine and spinal cord]. AB - Imaging of the spine and spinal cord has traditionally been accomplished with plain radiography, myelography, and CT. Recently, MR imaging has become the technique of choice in the assessment of lesions of the spine and spinal cord. MR imaging provides accurate localization of intramedullary, intradural extramedullary, and extradural tumors. Ependymomas and low-grade astrocytomas are the most common intramedullary tumors. MR imaging findings are distinguishable by the delineation and size of the lesion, and the signal intensity on T2-weighted images. Other less common tumors include malignant astrocytomas, hemangioblastomas, and intramedullary metastasis. Numerous foci of high-velocity signal loss are seen in the hemangioblastomas. Metastasis, meningiomas, and schwannomas are the most common intradural extramedullary tumors. Meningiomas are characterized by dural enhancement on postcontrast T1-weighted images. Schwannomas and neurofibromas often erode bony structures and appear to be dumbbell-shaped. Epidural metastasis accounts for the majority of extradural tumors. Primary malignant extradural tumors include lymphomas, chordomas, and so on. The most common primary benign extradural tumor is hemangioma, which often appears to be hyperintense on both T1-weighted and T2-weighted images. Intramedullary non-neoplastic lesions include demyelinating, vascular, and infectious diseases. Diffuse, peripheral, or speckled contrast enhancement, and lack of contrast enhancement may suggest non-neoplastic lesions. PMID- 10860381 TI - [Changes in MR imaging appearance of breast cancer after intra-arterial infusion chemotherapy]. AB - The purpose of this study was to evaluate whether the characteristic change in breast cancer related to chemotherapeutic response (CR) and the effect of invasion and toxicity in the skin and pectoralis muscle exist on MR imaging after intra-arterial infusion chemotherapy. A total of 11 patients with histologically proven breast cancer underwent MR study before and after chemotherapy. Changes in images and the dynamic curve-after-chemotherapy were evaluated, including time to maximum signal intensity (SI) and the early phase enhance ratio (EPER) in the tumor. In the tumor, changes in the dynamic curve, time to maximum SI, EPER and necrosis did not correlate with CR, but change in SI on T2-weighted images was suggested to do so. Changes in the dynamic curve and images in the pectoralis muscle and in images on the skin were suggested to correlate with CR. In addition, images changed for the worse in many cases of invasion and toxicity in the pectoralis muscle and in some cases of invasion in the skin. In conclusion, tumors had fewer imaging changes correlating with CR after intra-arterial infusion chemotherapy. Changes for the worse in images of the pectoralis muscle and skin may be useful for the evaluation of invasion. PMID- 10860382 TI - [Therapeutic effect of TAE in patients with hepatocellular carcinoma using color Doppler imaging with intravenous ultrasound contrast agent]. AB - We evaluated the therapeutic effect of TAE in 9 nodules with hepatocellular carcinoma (HCC) using color Doppler flow imaging with an intravenous ultrasound contrast agent. The intratumoral color signal enhancement that was detected in 7 nodules resulted in complete disappearance after TAE. The other 2 nodules without color signal enhancement showed well-differentiated HCC with fatty degeneration on histological study. The intratumoral enhancement noted in dynamic MRI in 7 of the 9 nodules resulted in complete disappearance after TAE. Color Doppler flow imaging with an intravenous contrast agent is a promising method for assessing the therapeutic effect of TAE. PMID- 10860383 TI - [Pitfalls in the assessment of radioresponse as determined by tumor regression: consideration based on the location and histologic constitution of tumors]. AB - PURPOSE: To prove the following hypotheses regarding tumor shrinkage after radiotherapy. Tumors located on an outer tissue surface, e.g. esophageal tumors, shrink faster than parenchymal tumors, e.g. lymph-node metastasis, because two clearance mechanisms, exfoliation and absorption, can operate in the former type of tumors whereas only absorption can function in the latter. Tumors which are being controlled do not necessarily respond completely, because tumors are constituted not only of tumor cells but also stromal tissues that are difficult to be absorbed. MATERIALS AND METHODS: Long-term shrinkage patterns of a parenchymal tumor were determined by using 18 curatively irradiated hepatomas. Preoperatively irradiated thymomas (10) and lymph-node metastases (37) from head and neck cancers were examined histopathologically. Twenty-one esophageal cancers were used for intra-patient response comparison between the primary disease and the lymph-node metastases. RESULTS: Shrinkage patterns were generally biphasic: rapid exponential regression followed by a plateau phase. Histologically, thymomas generally consisted of predominant fibrous tissues and few remaining tumor cells. Radioresponse did not predict the presence of remaining cancer cells in the lymph nodes. Esophageal-cancer radioresponse was always higher for the primary disease than the lymph-node metastases. CONCLUSION: The location and histologic constitution of tumors must be taken into account in predicting radiocurability using radioresponse. PMID- 10860384 TI - [Weight of feces and its daily fluctuation in young women. Part 1. A survey of the relation fecal weight and dietary habits and life-styles]. AB - This study investigates the relation of fecal production and dietary habits and life-styles in four 21 to 22 year-old healthy female students. The survey was conducted over 30 days and was repeated twice. All feces that were discharged were collected and weighed. The subjects performed very little physical exercise. The fecal weight, the number of defecations per day, gastrointestinal symptoms, feeling of incomplete defecation and of abdominal distention were recorded. The fecal weight was converted to autocorrelation, and the day-by-day variation was examined by a time series analysis (correlogram). Free access to foods was allowed. The weight of each food item was weighed for nutritional evaluation. The daily number of steps walked and sleeping hours were taken as indicators of life style. The average fecal weight ranged from 96.8 g/day to 127.8 g/day, with a grand mean for the four subjects of 94.1 g/day. The average number of times of defecation during the 60 days period was 53 to 72, or 0-3 a day. The subjects tended to have feeling of incomplete defecation when the stool was hard and fecal weight was less than 100 g per day, whereas the subjects felt incomplete defecation less frequently when the stool was well-formed or pasty. The time series analysis by correlogram indicates that the variation in fecal weight formed a 3-4 day cycle and that the cycle was irrelevant to fecal weight. This survey shows that there was no apparent correlation among the fecal weight and nutrient intake, the number of meals per day, the number of steps walked or sleeping hours per day. It also indicates that defecation factors differ from individual to individual. PMID- 10860385 TI - [A study of institutional medical care of female victims of sexual assault and violence]. AB - OBJECTIVE: The purpose of the study is to clarify the present situation of medical care for victims of sexual assault and violence. Medical facilities in two wards in Tokyo were studied in order to know what problems regarding medical care exist and how to support female victims. METHODS: In April 1998, we distributed questionnaires to 338 medical facilities covering all the clinics and hospitals, that had more than only otorhinolaryngology and ophthalmology, in Kouto-ku and Sumida-ku, Tokyo. The questionnaire included questions about individual experience of consulting with sexual assault and violence against women, the number of victims in the last year, and their understandings for victims. RESULT: 1) 76 of the respondents completed the answer sheet by themselves. The mean age of the subjects was 57.4 years old, 16.3% of them had seen sexual assault victims, and about 36.8% had cared for victims of violence. 2) 67 victims of sexual assault and violence were reported in the previous year. 36% of victims of sexual assault were reported by facilities related to obstetrics, and 85% of victims of violence were reported by general medical facilities. 3) As for understandings for victims, those who thought the victims were responsible for the sexual assault also regarded violence as caused by carelessness of victims. CONCLUSION: Medical facilities may be an important place to care for victims of sexual assault and violence against women. There are few data available as to how many women suffer from sexual violence. This study showed for the first time the reality of sexual assault and violence from the viewpoints of medical facilities in Japan, although it had some limitations. It is necessary for more discussion about roles of medical care for female victims of sexual assault and violence. PMID- 10860386 TI - [Availability of antibacterial towels and fabrics against enterohemorrhagic Escherichia coli]. AB - OBJECTS: After outbreaks of enterohemorrhagic Escherichia coli, various varieties of antibacterial kitchen and table materials are being used in Japan. The actual of availability of towels and fabrics, designated as antibacteria, against enterohemorrhagic E. coli was evaluated. METHODS: For an indicator strain a clinically isolated strain of E. coli O157:H7 C2 in Canada was utilized. Material kept in a tube was autoclaved at 121 degrees C for 15 mins, and 200 microliters of the bacterial culture was inoculated on the material, and cultured at 35 degrees C for an adequate interval. Colony forming units (CFU) of the materials were estimated. We also studied bactericidal effects of copper-fixed towels after washing. Materials without antibacterial processing or white cotton towels were used as controls. RESULTS: CFU of E. coli on silver- or copper-fixed materials at 18 hrs after inoculation were markedly decreased, compared to CFU of bacteria on the controls at 0 hr after inoculation. These materials were found to have bactericidal activity. Three materials of 23 samples were found to have bacterial inhibitory activity, because CFU of bacteria on the 3 materials at 18 hrs after inoculation were less than CFU of bacteria on the controls at 18 hrs. Thirteen materials (56.5%) were found to have no evident bactericidal nor inhibitory activity. CFU of silver- or copper-fixed materials were clearly decreased at 3 hrs and 20 mins after inoculation, respectively. It was also recognized that bactericidal activity was sustained at 18 hrs after inoculation even after the copper-fixed towel was washed 50 times. CONCLUSION: Evaluation of bactericidal effects of towels and fabrics, that claim to be antibacterial, showed that a few materials had bactericidal or inhibitory activity. Consumers must not have too much confidence in claims of a product being antibacteria. Even if antibacterial materials are available, the most important measure for us is to wash sufficiently and maintain sanitary conditions. PMID- 10860387 TI - [A study of the guideline of case management for persons with mental disorders analyzed with fidelity index. The conditions of effective implementation]. AB - OBJECTIVE: The present study examines the condition of effective case management based on the guideline of case management for persons with mental disorders. METHODS: A total of 295 clients were admitted to a case management trial for about 2 months. A fidelity index of program implementation and outcome measures of case management were developed. The relationship between implementation of critical program components measured by fidelity index, and outcome was analyzed. RESULTS: Fidelity Index was significantly correlated with outcome measures of clients, staffs and the care system. Effective but not-easily-feasible elements of program were having case-conference, application, of informal support and development of services. CONCLUSION: It is necessary to facilitate application of informal support with liaison with public health nurses, to standardize case conference and to place development of care services in the core of programs for ensuring effective implementation of case management for persons with mental disorders in Japan. PMID- 10860388 TI - [An oral health scoring system for promoting 8020 achievement in residents]. AB - The present study was undertaken to develop a self-scoring system which can be used by a resident to check lifestyle. The oral health scoring system which we used in Tobishima village, Aichi-ken, was named SAWAYAKA score. A total of 777 subjects were examined. The subjects responded to a questionnaire regarding their past individual lifestyles and dietary habits. Oral health conditions were also examined by dentists. The odds ratio and 95% confidence interval were calculated both from retained tooth numbers and the questionnaire. Questions with significant odds ratio were selected and the partial regression coefficients of quantification II method by Hayashi were calculated. The results are as follows; 1) Eleven questions showed a significant odds ratio between retained tooth numbers and past lifestyle and dietary habits. The questions involved the frequency of snack intake, tooth brushing frequency, having own tooth brush, smoking, drinking, having a hobby, having a family dentist, consulting a dentist before a problem got serious, gum bleeding, swollen gums and toothache caused by sensitivity to cold water. 2) The eleven items were analysed by using Hayashi's quantification II method. 3) The results showed that unswollen gums affected the retention of teeth by the range of 1.240. Toothache caused by sensitivity to cold water affected the retention of teeth by the range of 0.765. Having a hobby affected the retention of teeth by the range of 0.691. 4) The "SAWAYAKA" score was used to select important items, excluding drinking. 5) When results were analysed with the SAWAYAKA score, an average of 9.6 was obtained. It was concluded that the scoring list could be used for checking resident's lifestyles, and for promoting the preservation of more than 20 teeth at the age of 80. PMID- 10860389 TI - [A comparison between three methods to investigate falls among the elderly living in the community]. AB - PURPOSE: Interview survey is the often selected method to investigate falls among the elderly. However, the reliability of data obtained by that method has not be confirmed. The aim of this study is to clarify the number of intervals of recollection that should be specified to ensure accurate data when we investigate falls by using interview surveys among the elderly. METHODS: We carried out a total health survey among the elderly in Shigenobu town, and reported about the frequency of falls and its relating factors. Sex, age, walking ability over a distance of 1 km and hospitalization experience for past 1 year were related to falls significantly. In this study, we set 3 groups matching for the above factors in addition to history of falls during the preceding a year, and for one year we asked about falls every month for the first group (116 participants), every 3 months for the second group (116 participants) respectively. The third group (118 participants) was asked once at the end of this study. These three groups totaled 350 persons who were participants of physical fitness measurements that we carried out in Shigenobu town. The existence of falls was investigated by mail, and the nonrespondents were questioned by direct telephone calls. RESULTS: For reasons such as death, moving out, hospitalization and answer denial, persons whose answer was not obtained were excluded from the analysis. Finally, 87.1% in the 1st group, and 89.7% in the 2nd group and 96.6% in the 3rd group were analyzed. Statistically significant difference for the above factors related to falls did not exist among the 3 groups. Annual incidence for falls tended to be more frequent in the 1st group that in the 3rd group (20.5% in the 1st group, 15.9% in the 2nd group and 6.4% in the 3rd group) in males. In females, such a difference was not observed. (26.3% in the 1st group, 18.3% in the 2nd group and 20.9% in the 3rd group). CONCLUSIONS: There were no significant differences for the factors related to falls in 3 groups, therefore, the groups had almost similar backgrounds. In men, the difference of annual reported incidence between the 1st group and the 3rd group may be due to differences in the method of recollection. PMID- 10860390 TI - [An outbreak of enterohemorrhagic Escherichia coli O111 infections at a nursery school in Hiroshima Prefecture]. PMID- 10860391 TI - [Outpatients' knowledge about health and welfare services and the related factors. Using the questionnaire investigation of university hospital]. PMID- 10860392 TI - Impact of age on associations between weight and mortality. AB - The effect of age on the weight associated with the lowest mortality and the effect of age on the mortality risk associated with obesity are issues fraught with methodologic complexities. Current evidence supports the notion that the body mass index associated with the lowest mortality falls within the range of 18.5 to 24.9 in men and women between the ages of 30 and 74. The impact of age on the mortality risk associated with obesity changes with age, however, and the direction of the trend depends upon the measure used. PMID- 10860393 TI - Hypervitaminosis A and bone. AB - Animal, human, and in vitro data all indicate that excess vitamin A stimulates bone resorption and inhibits bone formation. This combination would be expected to produce bone loss and to contribute to osteoporosis development and may occur with relatively low vitamin A intake. It is possible that unappreciated hypervitaminosis A contributes to osteoporosis pathogenesis. PMID- 10860394 TI - Functional glycerol kinase activity and the possibility of a major role for glyceroneogenesis in mammalian skeletal muscle. AB - According to textbook descriptions of glycerol metabolism, liver and kidney are the only tissues that express significant glycerol kinase activity. Thus esterification of fatty acids to triglycerides in peripheral tissues such as skeletal muscle and adipose tissue is presumed to be dependent on the synthesis of glycerol-3-phosphate from glucose. This report describes exciting new data indicating that, although low, the glycerol kinase activity of skeletal muscle is functional. Interestingly, the results also suggest that neither glycerol nor glucose is the major substrate for the synthesis of muscle triglyceride glycerol. Rather, glyceroneogenesis, the synthesis of glycerol-3-phosphate from lactate, may play an as yet under-appreciated, but quantitatively important, role. PMID- 10860395 TI - Low-fat, high-carbohydrate diets and atherogenic risk. AB - The debate surrounding the use of low-fat, high-carbohydrate diets has raised concerns regarding the atherogenicity of triglyceride-rich lipoproteins. A recent study has demonstrated that feeding a diet high in carbohydrate delays very low density lipoprotein (VLDL) clearance. The same diet fed to mildly hyperinsulinemic subjects delays both VLDL and chylomicron clearance without affecting triglyceride production. PMID- 10860396 TI - Cellular retinoic acid-binding protein II: a coactivator of the transactivation by the retinoic acid receptor complex RAR.RXR. AB - Cellular retinoic acid-binding protein (CRABP) appears in vertebrate organisms in two isoforms, CRABPI and II, with distinct distribution and functions. CRABPII, when combined with its retinoic acid (RA) ligand, interacts specifically with the liganded RA receptor complex (RAR.RXR), which is bound to the RA response elements of particular genes in order to greatly activate their expression. Thus, liganded CRABPII acts as a coactivator for RAR.RXR. PMID- 10860397 TI - Syndrome X: medical nutrition therapy. AB - A significant number of Americans are at risk for developing a condition of insulin resistance termed Syndrome X. Dyslipidemia, resistance to insulin, obesity, and blood pressure elevation--the deadly quartet--describe Syndrome X, which increases atherogenic risk and contributes to coronary artery disease. Lifestyle factors such as overeating and physical inactivity play a pivotal role in Syndrome X. This deadly duet has been aptly coined "hyperactive fork" and "hypoactive foot," respectively. In addition, emerging evidence suggests that certain nutrients may help protect against Syndrome X. This review provides a brief discussion of diet and lifestyle factors related to Syndrome X. PMID- 10860398 TI - Morphometric analysis of the human cervical motoneurons in the aging process. AB - We examined 12 human cervical spinal cords to study the numbers and transverse cell body areas of the motoneurons in relation to the aging process. Our conclusion was that cervical motoneurons regulate their function by increasing in size to compensate the loss in numbers that occur as people get older. PMID- 10860399 TI - Myofiber length and myofiber arrangement in the antebrachial and leg muscles of sheep. AB - The myofibers of the skeletal muscle vary in length in muscle and from muscle to muscle. Information about myofiber length and myofiber arrangement is necessary to understand the architecture of muscles and differences in myofiber length in muscles. The purpose of the present study was to determine myofiber lengths, ratio of myofiber length to muscle length, and myofiber arrangement of the antebrachial and leg muscles in the sheep. The myofibers of the antebrachial muscles ranged from 7.3 mm (m. extensor carpi ulneris, bipennate muscle) to 39.6 mm (m. flexor digitorum profundus caput radiale, unipennate muscle) in length. The myofibers of the leg muscles ranged from 10.0 mm (m. flexor digitorum superficialis, multipennate muscle) to 119.2 mm (m. soleus, fusiform muscle) in length. The ratio of myofiber length to muscle length in the fusiform muscle was higher than that in the uni-, bi-, and multipennate muscles. The unipennate muscles tended to have a higher ratio of myofibers length to muscle length than did the bi- and multipennate muscles. No difference in the ratio of myofiber length to muscle length existed between the bipennate muscles and multipennate muscles. PMID- 10860400 TI - Effects of long-term treatment with caffeine on the ultrastructure of the golden hamster parathyroid gland and tibia. AB - The ultrastructure of the parathyroid gland and the SEM appearances of the tibia were studied in hamsters with and without administration of caffeine. Caffeine was treated orally each day at either 2.5 mg (low dose) or 10 mg (high dose) per 100 g body weight for a period of 17 or 32 days. Statistical analysis showed no significant differences among all groups examined regarding the serum calcium level. Transmission electron microscopy of the parathyroid gland revealed that the volume densities occupied by the mitochondria, Golgi complexes and rough endoplasmic reticulum of caffeine-treated groups were found significantly higher when compared with controls. The number of secretory granules observed close to the cell membrane per total amount of these granules revealed significant increase in all caffeine-treated animals. The bone mineral content (BMC) values were closely related to body weight. In the high dose caffeine-treated hamsters increment of the mean BMC and body weight values was significantly lower than those of the controls after 32 days. In the scanning electron microscopic studies of the tibia, no alteration in the morphometric parameters was demonstrated. It is considered that the synthesis and release of parathyroid hormone is stimulated following caffeine consumption. Our data suggest that although chronic administration of caffeine in the hamster may slightly increase bone turnover as evidenced by the BMC decrease, bone morphometry was not altered. Thus the osteoporotic changes were not proved in this study. PMID- 10860401 TI - Gender dimorphism of axons in the human lateral corticospinal tract. AB - Axons in the lateral corticospinal tract (LCST) were analyzed morphometrically on 37 cadavers (24 males and 13 females). The results revealed gender differences at the first lumbar cord level. Whereas the shape of the axons showed small differences (they were slightly more circular in males than in females), significant differences regarding the average area and diameter were found for the first time: these showed significant reduction with age in the case of males, but not in the case of females. PMID- 10860402 TI - Peroxidase activity in the submandibular gland of the house musk shrew, Suncus murinus (Soricidae, Insectivora). AB - Endogenous peroxidase activity in the submandibular gland of the house musk shrew, Suncus murinus was cytochemically investigated by light and electron microscopy using 3,3'-diaminobenzidine-tetrahydrochloride salt (DAB). The submandibular glands of male Suncus murinus at 8-month-olds were excised and diced into small pieces. In general, salivary glands are structurally divided into a terminal portion comprising a secretory portion and duct system. The submandibular gland of the Suncus murinus, the terminal portions consisted of proximal and distal acinar cells. On the other hand, a granular duct cell of the duct system contained a number of characteristic myelin-like bodies. In the present study, the peroxidase reaction products were localized in the secretory granules of the proximal acinar cells and in the endoplasmic reticulum, Golgi apparatus and myelin-like bodies of the granular duct cells. These reaction products were reduced when 5 mM 3-amino-1,2,4-triazole was added to the reaction medium. Additionally, release of peroxidase into the lumen was observed. In conclusion, the proximal acinar and granular duct cells formed peroxidase and may have performed excretory secretions. Moreover, the peroxidase positive myelin like body consisted of lamellated membrane and its outer surface membrane continued to the endoplasmic reticulum. PMID- 10860403 TI - Sexual dimorphism of the human cerebral pallium. AB - Sexual dimorphism was found regarding the cortico-medullary volume ratio in human brains throughout the age range, without the need for statistical analysis. After measurement of the areas of the whole pallium, cerebral cortex and cerebral medullary substance in 10 mm thick slices with the help of an image-analyzer, the total volume of the brain was calculated by integrating the volumes of each slice. The sexual dimorphism found in the cortico-medullary volume ratio may be of importance to understand sex differences, to solve questions related to various activities, functions, behaviours and responses, or to evaluate various pathologic conditions including non-physiological cerebral atrophy. PMID- 10860404 TI - SPR Award, 1999. For distinguished contributions to psychophysiology: Steven A. Hillyard. AB - At the Thirty-Ninth Annual Meeting of the Society for Psychophysiological Research (SPR), the award for Distinguished Contributions to Psychophysiology was presented to Steven A. Hillyard. The following is the citation given by Risto Naatanen on behalf of the Society's Awards Committee on October 9, 1999. PMID- 10860405 TI - Cognitive and emotional modulation of the cardiac defense response in humans. AB - The cognitive and emotional modulation of the cardiac defense response was investigated in this study. One hundred forty-four participants were exposed to three presentations of an intense auditory stimulus while performing one of four attentional tasks: a control task, an external perceptual tracking task, and two internal tasks presented at either easy or difficult memory loads. State anxiety was also manipulated by requiring each group to perform either with or without the threat of shock. Heart rate and vasomotor activity were recorded. Results indicated that only the externally directed tracking task led to potentiation of the cardiac response. No predicted effects for attentional demands were obtained and the anxiety manipulation did not appear to have an effect. Differences between measures were also observed, particularly with respect to response habituation. Unlike cardiac activity, vasomotor responses displayed resistance to habituation. The results are discussed in relation to contemporary accounts of defensive responding. PMID- 10860406 TI - Masked fear words produce increased SCRs: an anomaly for Ohman's theory of pre attentive processing in anxiety. AB - A. Ohman and J.J.F. Soares (1994) demonstrated that masked presentation of phobic pictures produces increased skin conductance responses (SCRs) in phobic subjects. A. Ohman (1993) explained this phenomenon in terms of a hypothetical "feature detector" that identifies physical characteristics of stimuli and activates the arousal system without involving significance evaluation or consciousness. By exposing spider phobics to spider words, general threat words, and neutral words instead of pictures, this explanation was tested. Words were presented both masked and unmasked while electrodermal activity was measured. Under unmasked conditions, SCRs were largest for spider words followed by general threat words, then neutral words. When masked, the difference between spider words and general threat words disappeared but SCRs remained significantly smaller for neutral words. It is concluded that activation of the arousal system by masked threat cues does not necessarily depend on their perceptual characteristics. PMID- 10860407 TI - Cardiovascular and electrodermal responses to support and provocation: interpersonal methods in the study of psychophysiological reactivity. AB - This study examined the joint and independent effects of experimentally manipulated social contexts and individual differences in hostility and perceived social support on physiological responses to a social stressor, while illustrating the use of the interpersonal circumplex for integrative social psychophysiological research. Undergraduate women completed a speech task in a supportive, neutral, or provoking context and completed measures of hostility and perceived social support. The provoking context evoked the largest blood pressure and heart rate (HR) responses, followed by the neutral and the supportive context. Social context also influenced HR and electrodermal reactivity during task preparation. Hostility elicited higher systolic blood pressure (SBP) reactivity during preparation, speech, and recovery. Perceived social support interacted with context to affect SBP and HR during speech and preparation. The roles of interpersonal characteristics and contexts in the physiological stress response and the utility of interpersonal methods in studying these associations are discussed. PMID- 10860408 TI - Effects of paced respiration on heart period and heart period variability. AB - The present study investigated psychophysiological responses to paced respiration of different frequencies. Twenty men and 20 women (mean age: 24.3 years) underwent five breathing conditions (paced with 0.15 Hz, 0.20 Hz, 0.25 Hz, 0.30 Hz, and unpaced), each lasting 5 min. As dependent physiological measures heart period, and different heart period variability (HPV) parameters were assessed. Psychological variables consisted of mood estimates as well as rated accuracy and effort to follow the pacing rhythm. HPV decreased with higher breathing frequencies, under paced and unpaced conditions, whereas mood ratings did not change. Subjects indicated more effort and less accuracy in following the pacing signal, the more its frequency differed from their spontaneous breathing frequency. The comparison of a spontaneous breathing condition with a frequency matched paced condition revealed that pacing per se provoked a reduction in heart period. Because this decrease was not accompanied by changes in any of the HPV frequency components, their validity as measures of autonomic control needs to be questioned. PMID- 10860409 TI - The respiratory-related evoked potential: effects of attention and occlusion duration. AB - The present study assessed the effects of occlusion duration and attention on components of the respiratory-related evoked potential (RREP). Twenty-nine channel evoked potential recordings were obtained from 12 young adults exposed to a pseudorandom sequence of 100-, 200-, 400-, and 800-ms inspiratory occlusions, under attend and ignore conditions. Results demonstrated that the duration of an inspiratory occlusion does not affect RREP components systematically, highlighting the importance of the onset of the occlusion in producing the cortical responses. Attention resulted in augmentation of the N1, P2, and P3 components but did not affect the early latency Nf and P1 components. P1, N1, and P3 occurred with shorter latencies in the attend condition. One subject with poor duration estimation ability displayed substantially delayed P3 latency. This result highlights the relationship between P3 and perception of respiratory somatosensory information. PMID- 10860410 TI - ERP differences in visual attention processing between attention-deficit hyperactivity disorder and control boys in the absence of performance differences. AB - Event-related potentials (ERPs) were recorded during a visual two-choice reaction time (RT) task in attention-deficit hyperactivity disorder (ADHD) and control boys selected using strict inclusion and exclusion criteria. No group differences were found in mean RT and correct responses. Although early occipital ERPs were not affected in the ADHD group, the peak latency of early anterior ERPs (N1, P1, N2) was significantly delayed. ADHD showed a larger effect of stimulus type on the frontal negativity (N530) and the posterior late negativity (nSW) and a smaller effect of stimulus type on anterior N2 and posterior P3b amplitude. The development of N530 and P450 amplitude across blocks of five trials was analyzed using orthogonal polynomial trend analysis of variance software. In the control group, P450 amplitude to "frequent" stimuli reduced across blocks. In the ADHD group, N530 amplitude increased for "rare" stimuli across blocks. It is suggested that the ADHD group showed a lack of automatization of the categorization process with increasing time on task for which they compensated by controlled attentional processes. PMID- 10860411 TI - Attentional capacity, a probe ERP study: differences between children with attention-deficit hyperactivity disorder and normal control children and effects of methylphenidate. AB - In the present study it was investigated whether the smaller P3s in attention deficit hyperactivity disorder (ADHD) children are caused by a shortage of capacity underlying P3 processes or whether they are due to a capacity allocation problem. Also, effects of methylphenidate on these processes were investigated. Performance and event-related potentials (ERPs) of 14 ADHD and 14 control children were measured using an irrelevant-probe technique. Three types of task irrelevant visual probes (standards, deviants, and novels) were presented against the background of two visual tasks that varied in task difficulty. The parietal P3 wave was measured in response to task stimuli and probes. ADHD subjects made significantly fewer correct detections than normal controls in both the easy and the hard tasks. Controls showed an enhanced P3 to task-relevant stimuli in the hard task, whereas ADHD children did not. Probe (novel) P3 amplitudes decreased from the easy to the hard task to the same extent in both groups. Methylphenidate enhanced the percentage of correct responses and task P3 amplitudes in both the easy and the hard task but probe P3 amplitudes were not influenced by methylphenidate. It was concluded that ADHD children do not suffer from a shortage in attentional capacity; rather, the evidence is in favor of a problem with capacity allocation. Furthermore, methylphenidate had enhancing effects on performance and ERPs, but did not improve the capacity-allocation deficit. PMID- 10860412 TI - Detecting the onset of the lateralized readiness potential: a comparison of available methods and procedures. AB - Studies that measure the onset of the lateralized readiness potential (LRP) could well provide researchers with important new data concerning the information processing locus of experimental effects of interest. However, detecting the onset of the LRP has proved difficult. The present study used computer simulations involving both human and artificial data, and both stimulus- and response-locked effects, to compare a wide variety of techniques for detecting and estimating differences in the onset latency of the LRP. Across the two sets of simulations, different techniques were found to be the most accurate and reliable for the analysis of stimulus- and response-locked data. On the basis of these results, it is recommended that regression-based methods be used to analyze most LRP data. PMID- 10860413 TI - Phasic heart period reactions to cued threat and nonthreat stimuli in generalized anxiety disorder. AB - The hallmark of generalized anxiety disorder (GAD) is chronic uncontrollable worry. A preattentive bias toward threat cues and hypervigilance may support this ongoing state of apprehension. A study was conducted to bridge the attentional and physiological underpinnings of GAD by examining phasic heart period (HP) responses to cued threat and nonthreat stimuli. Thirty-three GAD clients and 33 nonanxious control participants engaged in an S1-S2 procedure that employed cued threat and nonthreat word stimuli, during which phasic HP reactions were recorded. As compared with the control group, the GAD group showed (1) smaller cardiac orienting responses and impaired habituation of cardiac orienting to neutral words, (2) HR acceleration in response to threat words, and (3) a conditioned anticipatory HR deceleration to threat words over repeated trials. The cardiac-autonomic underpinnings of GAD appear to rigidly maintain precognitive defensive responses against threat. This portrayal is discussed in the context of an integrative model that depicts diminished global adaptive variability in GAD. PMID- 10860414 TI - P300, N400, aerobic fitness, and maximal aerobic exercise. AB - Electrophysiological effects of aerobic fitness and maximal aerobic exercise were investigated by comparing P300 and N400 before and after a maximal cycling test. Event-related potentials (ERPs) were obtained from 20 students divided into two matched groups defined by their aerobic fitness level (cyclists vs. sedentary subjects). The session of postexercise ERPs was performed immediately after body temperature and heart rate returned to preexercise values. At rest, no significant differences were observed in ERP parameters between cyclists and sedentary subjects. This finding argued against the hypothesis that ERP modifications may be directly assumed by aerobic fitness level. The postexercise session of ERPs showed a significant P300 amplitude increase and a significant P300 latency decrease in all subjects. Similarly, N400 effect increased significantly after the maximal exercise in all subjects. ERP changes were of the same magnitude in the two groups. The present study argues for a general arousing effect of maximal aerobic exercise, independently of the aerobic fitness level. PMID- 10860415 TI - Vagal changes following cancer chemotherapy: implications for the development of nausea. AB - Many physiological changes that occur contemporaneously with nausea are mediated by the autonomic nervous system, but the specific autonomic changes associated with nausea have not been characterized. Cardiac parasympathetic (vagal) activity as indicated by heart rate variability, measured as the standard deviation of successive differences (SDSD) in beat-to-beat intervals, was assessed in 24 women with ovarian cancer immediately prior to and accompanying nausea that occurred following anticancer chemotherapy. A progressive increase in SDSD followed infusion of the chemotherapy agent, indicating a rise in cardiac parasympathetic (vagal) activity, with onset of nausea consistently occurring after the peak activity had been reached, at a time when SDSD was decreasing. An increase in parasympathetic activity seems to set the stage for the expression of nausea but an additional stimulus is apparently needed to finally trigger the event. PMID- 10860416 TI - Changes in spinal excitability during choice reaction time: the H reflex as a probe of information transmission. AB - The aim of the present study was to investigate the modulations in amplitude of H reflexes elicited in a hand muscle, the flexor pollicis brevis, during the performance of a choice reaction time (RT) task in which this muscle was directly involved. Ten subjects were to choose between a left- or a right-thumb key-press according to the lateral location of a flash of light. The stimulus-response mapping was either compatible or incompatible. Hoffman reflexes were elicited at different times during the RT by stimulation of the median nerve. Twenty-five milliseconds before the voluntary response, the amplitude of the H reflex suddenly increased when the muscle was involved in the response and decreased symmetrically when the muscle was not involved in the response. Mapping compatibility exerted no detectable influence on the changes in spinal excitability. The latter result supports the assumptions that are at the core of Sternberg's additive factor method. PMID- 10860417 TI - [Treatment of sepsis]. PMID- 10860418 TI - [Coagulation disorders in sepsis]. PMID- 10860419 TI - [Mechanisms in sepsis induced by gram positive microorganisms]. PMID- 10860420 TI - [Sepsis--pathogenesis]. PMID- 10860421 TI - [Adult respiratory distress syndrome in the year of 2000]. PMID- 10860422 TI - [Empirical antibiotic treatment of sepsis]. PMID- 10860423 TI - [Dopamine in "kidney-friendly" doses]. PMID- 10860424 TI - [Continuous venovenous hemodiafiltration in critically ill patients with acute renal failure]. AB - The development of computerized machines with a simple user-friendly interface to perform continuous venovenous hemodiafiltration (CVVHDF) has resulted in a break through of CVVHDF in the treatment of acute renal failure (ARF) in Danish intensive care units. During CVVHDF the blood is submitted to a combination of dialysis and ultrafiltration. In contrast to intermittent haemodialysis (HD), CVVHDF can be used in critically ill patients with unstable circulation. Biocompatible membranes are used. During treatment with CVVHDF, cytokines are removed from the blood partly by ultrafiltration, partly by adsorption to the filter. The clinical importance of this is not yet known. Patients with ARF treated with CVVHDF seem to be more likely to show renal recovery than those treated with HD. There are few prospective investigations of the effect of CVVHDF on mortality, but all comparisons of CVVHDF with HD indicate a trend in favor of CVVHDF. PMID- 10860425 TI - [Non-antibiotic treatment of sepsis]. AB - Treatment of patients with sepsis with monoclonal antibodies against lipopolysaccharide, tumour necrosis factor or treatment with soluble TNF receptors or interleukin-1 receptor antagonist have not had any effect on mortality. Treatment with intravenous polyclonal nonspecific immunoglobulin has been shown to have beneficial efficacy on mortality. However, the number of patients included in the latter treatment studies are limited, the patients studied are heterogeneous and do not allow to conclude that sepsis patients should be routinely treated with immunoglobulins. PMID- 10860426 TI - [Incidence, severity and mortality of acute respiratory failure in Denmark]. AB - To determine the incidence, severity and 90-day mortality of acute respiratory failure (ARF), acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) we conducted an eight-week prospective cohort study in 48 Danish ICUs, during which all ICU admissions (n = 6647) > 14 years of age were assessed. ARF was defined as intubation and mechanical ventilation > 24 hrs. ALI and ARDS were defined in accordance with the American-European Consensus Conference criteria. Among the 813 patients included, 552 were diagnosed with ARF, 117 with ALI and 95 with ARDS. The incidences (patients per 100.000/yrs) for ARF were 84.8, for ALI 17.8 and for ARDS 14.6. The 90-day mortality was 46.3% for ARF patients without ALI/ARDS, 47.3% for ALI patients and 46.2% for patients with ARDS. Compared to previously reported figures, the ARDS mortality is in the lower range whereas the incidence is slightly higher. This probably reflects a broader selection of patients when using the consensus criteria to define the ARDS population as opposed to definitions previously used. PMID- 10860427 TI - [Antibiotic pre-hospital treatment and the course of meningococcal disease]. AB - The aim of the study was to assess the effect of pre-hospital antibiotic treatment given by general practitioners to patients with meningococcal disease. It was carried out as a 16-year population-based historical follow-up study based on referral letters and hospital records in the County of North Jutland, Denmark, and included 320 patients with meningococcal disease, of whom 302 were examined by a general practitioner before admission to hospital. The main outcome measure was death. We found that 44 patients (14.6%) were given antibiotic treatment by the referring general practitioner. Nine of these (20.5%) died, compared with 16 (6.2%) patients who did not receive pre-hospital antibiotic treatment. The presence of skin bleeding, petechiae and impaired consciousness were strongly associated with case fatality. Even after adjustment for these variables the odds ratio for death in patients treated with antibiotics was high (3.2; 95% CI 0.9 10.6). In the 15 patients with skin bleeding (ecchymoses, suggillations) the case fatality rate was 100% in patients treated with antibiotics, and 50% in patients who did not receive antibiotics before hospitalization. It is concluded that pre hospital treatment is mainly given to the most severe cases with expected high case fatality, and this confounding by indication was probably not fully adjusted for with the available data. The results contradict previous findings and provide reason to doubt the benefit of pre-hospital antibiotic treatment in patients with meningococcal disease. PMID- 10860428 TI - [Two thousands seven hundred and thirty nine episodes of bacteremia in the county of Northern Jutland 1996-1998. Presentation of a regional clinical database]. AB - A complete registration of bacteraemias has been undertaken in the County of Northern Jutland (population 493,000) since 1996, in the form of a bacteraemia register maintained by the Department of Clinical Microbiology, Aalborg Hospital, which serves the seven hospitals in the county. A follow-up is conducted on all patients with positive blood cultures. We registered 2,739 bacteraemias during 1996-1998. Eighty-nine percent of bacteraemias were monomicrobial and 11% polymicrobial. Among monomicrobial bacteraemias the predominant bacteria were Staphylococcus aureus (17%), Streptococcus pneumoniae (12%), Escherichia coli (31%), and other enterobacteria (14%). The source (focus) of infection was identified in 82% and the main foci were the urogenital tract (28%), the respiratory tract (14%), the digestive tract (13%), the hepato-biliary tract (8%), and intravascular devices (8%). The overall 30 day case fatality rate (CFR) was 21.5%. At first notification antibiotic therapy was appropriate in 54% of bacteraemias (CFR 14.8%), antibiotic therapy was inappropriate or missing in 38% (CFR 21.0%), active therapy had been withdrawn in 0.3%, in 5% the patient had died, and in 2% therapy had been completed or the patient had been discharged alive. The register has thus created a platform for research projects regarding risk factors, therapy and prognosis of bacteraemia. PMID- 10860429 TI - [Life-threatening puerperal sepsis caused by group A streptococci]. AB - A case of postpartum streptococcal toxic shock syndrome due to group A streptococci is described. The patient suffered multiorgan failure, was critically ill and stayed in the intensive care unit for 18 days before recovering. The patient received massive antibiotic treatment as well as intravenous immunoglobulin therapy. The case is described to draw attention to the reoccurrence of serious group A streptococcal infections. PMID- 10860430 TI - [Pseudomonas aeruginosa as the cause of meningitis in a patient with epidural catheter]. AB - A case of epidural infection following epidural catheterization is presented. The clinical signs initially were backpain and a small swelling at the site of insertion. Treatment with dicloxacillin was begun, presuming a Staphylococcus infection. The symptoms persisted. Weeks later the patient developed meningitis and Pseudomonas aeruginosa was cultivated. Antibiotic treatment was changed to ceftazidime, netilmycin and ciprofloxacin. Complete recovery followed. PMID- 10860431 TI - [Fatal outcome of Strongyloides stercoralis infection in a patient with no previously known immunosuppression]. AB - A case of fatal infection with Strongyloides stercoralis in a previously healthy young African with no known immunosuppression is reported. The patient suffered severe gastrointestinal bleeding and despite intensive treatment died of multiorgan failure. Postmortem, signs of ulcerating T-cell lymphoma were found in a jejunal specimen, and the patient was found to be infected with HTLV-1. Gastrointestinal bleeding in relation to infection with Strongyloides stercoralis is discussed as well as possible relations between the severity of infection, lymphoma and HTLV-1. PMID- 10860432 TI - [Picture of the month. Pseudomonas aerupinosa sepsis]. PMID- 10860433 TI - [Optimal fluid therapy in acute bacterial meningitis--from waterline to desert wandering]. PMID- 10860434 TI - [Women physicians are performing research--but few will become professors]. PMID- 10860436 TI - [Ethical tensions in therapeutic research]. PMID- 10860435 TI - [Equality--also in academic medicine]. PMID- 10860437 TI - [Art and music in intensive care units]. PMID- 10860438 TI - [Drug therapy of influenza--a comment?]. PMID- 10860439 TI - [Influenza--drug therapy and prevention]. PMID- 10860440 TI - [What is the patient's HIV status?]. PMID- 10860441 TI - [Molecular biology and counseling of patients and their families]. PMID- 10860442 TI - [The experience of medical support for the troops in World War II: its assessment and importance 55 years after the Great Victory]. PMID- 10860443 TI - [The organization of surgical care in a frontier hospital at the stage of troop withdrawal from Afghanistan]. PMID- 10860444 TI - [Military medical statistics: the status, problems and outlook]. AB - The reform of medical service of the RF Armed Forces has objectively required the conduction of quite definite changes and military and medical statistics as an applied science, the teaching subject and field of practical activity of medical service. The results obtained during analysis of experience of the Armed Forces personnel medical support for the last years have shown that it is necessary to widen the scope of military and medical statistics applications by military physicians, medical specialists of medical service head-quarters in the form of their informative and statistic activity which is one of the integrated part of medical service in military men. The article presents the main results of military and medical statistics reformation, as well as principal trends of its further improvement. PMID- 10860445 TI - [Diagnosis and the technology for optimizing the medical support of a troop unit]. AB - The work is devoted to investigation of the system of military unit medical support with the use of principles and states of organizational diagnosis; development of the method allowing to assess its functional activity; and determination of optimization trends. Basing on the conducted organizational diagnosis and expert inquiry the informative criteria were determined which characterize the stages of functioning of the military unit medical support system. To evaluate the success of military unit medical support the complex multi-criteria pattern was developed and algorithm of this process optimization was substantiated. Using the results obtained, particularly realization of principles and states of decision taking theory in machine program it is possible to solve more complex problem of comparison between any number of military units: to dispose them according to priority decrease; to select the programmed number of the best and worst; to determine the trends of activity optimization in corresponding medical service personnel. PMID- 10860446 TI - [The central hemodynamics and renin-angiotensin-aldosterone system in servicemen with arterial hypertension who participated in battle actions]. PMID- 10860447 TI - [The treatment of middle-aged and elderly patients with femoral neck fractures]. PMID- 10860448 TI - [A trial of the use of new tableted forms of central-action analgesics]. PMID- 10860449 TI - [A bronchial obstructive syndrome in pneumonia patients: its clinical and pharmacotherapeutic aspects]. PMID- 10860450 TI - [A comparative trial of the clinical use of new antidepressants]. PMID- 10860451 TI - [A trial of the use of Triderm preparations]. PMID- 10860452 TI - [A device for extracting pins from the long bones]. PMID- 10860453 TI - [A new homeopathic medicine kit]. PMID- 10860454 TI - [The treatment of acute intestinal diarrheal infections]. AB - The article presents a short characteristic of the general therapeutic principles in intestinal diarrheal infections, the main of which are the following: early beginning, individual approach and complex therapy. Therapeutic schemes of the most frequent diseases are given. Special attention is paid to the great importance of rational bacterio-therapy in current treatment of acute intestinal diarrheal infections. Individual approach considers determination or supposition of disease etiology and clinical form before treatment, as well as evaluation of state pathogenesis taking into account the body physiologic specificity, severity and period of disease, possible complications and concomitant disease. Complex therapy supposes simultaneous effect on disease agent, several links of its pathogenesis, struggle with intoxication, restoration of the disturbed functions of organs and systems, increase in body defense, treatment of concomitant diseases. PMID- 10860455 TI - [Viral hepatitis B: the metabolism of the B-group vitamins against a background of glucocorticosteroid treatment]. PMID- 10860456 TI - [The assessment and correction of the body functional state of aviation specialists working with video display terminals]. AB - The investigations have shown that military and aviation specialist working with video-display terminals are exposed to unfavourable effect of professional environment of anthropogenic genesis. It is accompanied with development of energo-informative stress. Manifestations of stress-effect have a cumulative character. One can notice the decrease in physiologic reserve level, homeostatic potential, worsening of the quality of operators' professional health and safety in the "man-machine" system. The proposed and appropriated methods of premorbid state diagnosis and working capacity support have confirmed its adequate efficiency both in experimental conditions and in aviation specialists against the background of their professional activity. PMID- 10860457 TI - [Efim Ivanovich Smirnov: the lessons of World War II (on the 55th anniversary of the Great Victory)]. PMID- 10860458 TI - [Veteran medics on World War II]. PMID- 10860459 TI - [Isaak Iosifovich Rogozin (on the centenary of his birth)]. PMID- 10860460 TI - [Organizer of military public health, clinician, scientist and teacher (on the 75th anniversary of the birth of V. S. Matkovskii)]. PMID- 10860461 TI - [The 60th anniversary of the Arkhangel'sk Military Hospital]. PMID- 10860462 TI - [Guarding the health of aviation command personnel]. PMID- 10860463 TI - [Military medicine in World War II (on the 55th anniversary of the Great Victory)]. PMID- 10860464 TI - [The state legal and expert identification aspects of protecting the right of the person for citizens of the Russian Federation]. AB - The article deals with the questions concerning the problem of person identification, different extreme situations. This problem is closely connected with ensuring of national security of Russia. The authors give the definition of person's right in contact with constant threat to violate this right. Two categories of person's properties are analysed (material and not material) with the help of which it is possible to establish identity of man and his personality. It is noted that protection of person's rights is carried out in six forms. For the first time the possibilities to create organisational system of person identification in the form of Federal service are discussed that must be legislatively secured. Scientific investigation of this problem allowed the authors to submit concrete proposals concerning the question of organisation of state service for person identification. PMID- 10860465 TI - [The humanitarian-legal aspects of medical support for the multinational peace keeping forces in Yugoslavia]. PMID- 10860466 TI - [The outlook for the organization of specialized ambulatory surgical care]. PMID- 10860467 TI - [The intraoperative leaving of foreign bodies as a defect in surgical treatment]. AB - The work presents literary data and our own study results obtained in the cases of intraoperative leaving of foreign bodies in different categories and types of surgical interventions. Influence of the fact of foreign body leaving on outcome is analysed. The authors describe the causes of intraoperative leaving of foreign bodies and preventative measures of such medical care defect. Recommendations on medical tactics in the case of detection of foreign body left during operation are given. PMID- 10860469 TI - [A trial of the use of ronkoleukin in the treatment and prevention institutions of the Navy]. PMID- 10860468 TI - [The clinical and economic aspects of treating community-acquired pneumonias]. PMID- 10860470 TI - [The importance of stimulation of the statokinetic system in the combined treatment of patients with the initial manifestations of vascular insufficiency in the vertebrobasilar arteries]. PMID- 10860471 TI - [Low-intensity magnetic-laser therapy in the combined treatment of alcoholics with neurotic disorders]. PMID- 10860472 TI - [Claritin in the combined therapy of neurodermatoses]. PMID- 10860473 TI - [The potentials for using xidifon in diseases of the locomotor system]. PMID- 10860474 TI - [Visual color stimulation as a nondrug method for eliminating emotional stress]. PMID- 10860475 TI - [Current problems in the theory of the epidemic process]. PMID- 10860476 TI - [The current aspects of the prevention of hepatitis B at hemodialysis centers (a review of the literature)]. PMID- 10860477 TI - [The effect of circadian rhythms on the body functional status and the work capacity in operators]. AB - Lately more and more attention is given to the indices of efficiency and safety of operators' activities because of their special complexity and mistake value. Operators' work is connected with intensive processing of encoded information and is accompanied with development of significant fatigue, stereotype cyclic alternation, changes in work and rest regimen--with necessity of working at night. Effect of circadian biorhythms considerably influences on characteristics of operators' activities. The existing organizational approach considers the operator only as being present or absent at working place. The fact that operator can be present but his working capacity is not sufficient is taken into account not enough and that's why requires consideration of operators' work and elaboration of organizational measures. PMID- 10860478 TI - [The adaptive pharmacological correction of functional disorders in young recruits with hypotrophy]. AB - Low level of physical development, physiologic reserves, signs of psychical disadaptation, frequent functional disorders in cardiovascular system and gastrointestinal tract are noted in recruits with hypotrophy. Using of rapidly acting adaptogenes during 1 month (phytopreparation elixir "Altai" and "Vitavis" in tablets) significantly improves body state: increase in the level of physiologic reserves, physical working capacity, IMT, body unspecific resistance; improvement in indices of hemodynamics, metabolism and immunity; decrease in asthenization and psychical disadaptation. The state of adaptation and increased resistance in this recruit group remains during the following months of service. PMID- 10860479 TI - [The characteristics of the delivery of medical care in frostbite in the Soviet Finnish War]. PMID- 10860480 TI - [The history of research into the problem of the evacuation of the wounded and sick by aviation transport]. PMID- 10860481 TI - [The 55th anniversary of the Forensic Medical Laboratory of the Northern Caucasus Military District]. PMID- 10860482 TI - [Directives from the commander of the Main Military Medical Administration of the Ministry of Defense of the Russian Federation]. PMID- 10860483 TI - [A statistic study of airborne pollen grains]. AB - Since 1969, authors have been doing a statistical analysis of airborne pollen grains, aiming at the clarification of their atmospheric behaviors, in connection with an increase in a number of pollinosis cases. The airborne pollens were collected three times a week starting from January 1 and ending December 31 with the help of cascade impactor, by which 6001 of air was suctioned at the rate of 51 per min for 2 h from 10:00 a.m. to 12:00 at the rooftop of one of the buildings (16 m high) in the Narashino Campus of Toho University, Funabashi, Chiba. The collected airborne pollens were numerated and classified under microscope according to the Ikuse's classification system for Japanese pollens. The number of days (n) was plotted as a function of daily number of a certain type of airborne pollens (x), not fitting to a simple normal distribution. However, the data were found to approximate a normal distribution when x was log transformed as ln (x + 1). Among the statistical methods tested, semilogarithmic plot, Weibull plot, Edwards' plot and circular graph method were found to be useful in the follow-up of seasonal variation of environmental pollen grains, especially when used in combination. PMID- 10860484 TI - [Application of radionuclides in pharmaceutical fields]. AB - Application studies of radionuclides in pharmaceutical fields are described concerning two major categories, such as the use of gamma-emitter in nuclear medicine and beta-emitter in tracer investigation. In the former radionuclide an effort has been made to develop radioactive new diagnostic agents in the nephro urology field. Of several agents examined two new radiopharmaceuticals, 99mTc malic acid and 99mTc-dimercaptopropionic acid proved to be sufficient for renal imaging with adequate concentration in experimental animals and humans. Useful information was obtained for diagnosis of renal diseases using both the labeled agents. Identification and purity tests of radiopharmaceuticals also set forth for official compendia such as the Japanese pharmacopoeia and so on. In the latter one the fates of two medicines, thalidomide and norphenylephrine, were studied and the safety evaluation of some xenobiotics including food additives, plasticizers and domestic chemical products, etc. was carried out in animals using synthesized 14C- or 3H-labeled compounds. In addition, the metabolism of n propyl-N-nitrosourea, a carcinogen, in rats, and chemical modification of high molecular substances such as DNA, RNA and proteins by 1,1'-ethylene-bis(1 nitrosourea) (EBNU), a carcinostatic agent, were studied in vitro by the use of the tracer technique. PMID- 10860485 TI - [Functional arrangement of genomic DNA and structure of nuclear matrix]. AB - The nuclear matrix (nuclear scaffold), the RNA-protein skeleton of the nucleus, has a role in the organization and function of nuclear DNA. Nuclear processes associated with the nuclear matrix include transcription, replication, repair and splicing. We have purified a nuclear matrix protein, P130, which binds to several matrix attachment regions (MARs). Since the nucleotide sequence of P130 cDNA cloned by us was closely similar to that of matrin 3 cDNA cloned, except for two incorrect nucleotides within the matrin 3 coding region, and since the functions of matrin 3 were unknown, P130, referred to as P130/Mat3, was functionally characterized. The primary structure deduced for P130/Mat3 contained two DNA binding domains with C2H2-type zinc finger motif and two RNA binding domains. In addition, there were a nuclear localization signal and several phosphorylation sites for tyrosine or serine/threonine protein kinases, suggesting its multiple functions. MAR inserted upstream from the SV40 promoter in pMAR/luc assisted luciferase gene transcription in a transient expression system in Ac2F cells. Cotransfection of a plasmid carrying P130/Mat3 cDNA downstream from the CMV promoter into Ac2F cells produced this protein a level 4 times higher than that in wild-type Ac2F, causing 20 times higher luciferase activity from pMAR/luc than that induced by pMAR/luc alone. These findings indicated that MAR functions as a cis-element to which P130/Mat3 binds as one of the possible transactivators. Nuclear matrix proteins, which are tissue- and cell-type-specific, are altered with transformation and state of differentiation. We have shown that an MAR binding protein, P230, is detectable in rat hepatoma cells but not in normal liver, and suggested that this protein is a diagnostic and prognostic marker for liver cancer. It is clear that nuclear matrix proteins hold a considerable promise as diagnostic tools for pathologists. Present evidence, including our data, suggests that nuclear matrix proteins may be useful biomarkers of neoplastic disease in the serum, body fluids, and tissues. Nuclear matrix proteins are also potential candidates for the use as tumor prognostic factors and targets of anticancer drugs through apoptosis. We will discuss screening of drugs that interact with nuclear matrix proteins and influence nuclear events. PMID- 10860486 TI - [Pigment molecules linked to polymer support: blue rayon, blue chitin, and green chitosan-synthesis and applications]. AB - The fact that hemin can inhibit the mutagenic activity of compounds bearing polycyclic structures is ascribable to the ability of the porphyrin structure to complex with the planar surface of the mutagens. The elucidation of this mechanism has led to the discovery of copper phthalocyanine trisulfonate (cpt) as an efficient ligand to trap polycyclic compounds on polymeric supports that bear cpt through covalent bond linkages. In blue cotton, the support for cpt is cotton, in blue rayon, it is amorphous rayon, and in blue chitin it is poly-N acetylglucosamine. Using these blue materials, we have successfully isolated mutagens of polycyclic structures, e.g., heterocyclic amines, from environmental complex mixtures such as food, urine, feces, and river water. Preparation and properties of these adsorbents are described. Chlorophyllin linked to Sepharose and chitosan is also described. The use of these green materials is discussed. PMID- 10860487 TI - [Application study of drug information to proper use of drugs--estimation of adverse drug reactions by the evaluation scores of subjective symptoms (complaints) and background of patients]. AB - We have built a database for CARPIS (Case Reports of Adverse Drug Reaction and Poisoning Information System) since 1987, and the case reports of adverse drug reactions accumulated in the CARPIS database to be total about 20,000. The purpose of our study is to develop, implement, and assess an estimation procedure for preventing the adverse drug reactions by subjective symptoms (complaints) of patients using this CARPIS. The case reports of the adverse reactions are not a systematical study and are elapse reports by the case, which encountered on the way of the cure. Therefore, it might be possible to use as the data if not gaining enough evaluation. Especially, the abstracts in the annual meeting of the society are fragmentary and does not often have objective evidence. Therefore, we contrived an original evaluation method and carefully selected the case reports for our study. Also, the adverse reactions are very colorful and it is difficult to handle them at once. Therefore, we study every organ and want to aim to make a finally all-round format. We reported on the Liver disorders, the Extra-pyramidal symptoms, the Drug eruption, the Leukopenia and so on. We discovered that there was an adverse reaction which shows very peculiar subjective symptoms (ex. Extra pyramidal symptoms) and unpeculiar subjective symptoms (ex. Leukopenia). We want to show our point of view and the future of our study below. PMID- 10860488 TI - [The mechanism of alteration of monoamine metabolism in brain regions in marble burying behavior-isolated housing mice and effect of oren-gedoku-to on this alteration]. AB - The mechanism of the alteration in marble burying behavior-isolated housing (MBB IH) mice was investigated. The determination of hypothalamus monoamine and serum corticosterone contents indicated that MBB-IH mice readily responded to the stress stimuli in conditioned fear stress. Six drugs, such as buspirone (10 mg/kg, p.o.), zimelidine (10 mg/kg, p.o.), clomipramin (10 mg/kg, p.o.), yohimbine (5 mg/kg, p.o.), ethyl beta-carboline-3-carboxylate (beta-CCE, 5 mg/kg, p.o.) and flumazenil (15 mg/kg, p.o.) were singly and/or three times administered to MBB-IH mice. Their inhibitory activity on the MBB-IH mice was considered by the use of activity profiles consisting of spontaneous locomotor activity, marble burying behavior and hypothalamus monoamine content. Using these profiles, we calculated the activities as vector in three-dimensional space, and compared the distance from the control point (DCP). DCPDOPAC and DCP5-HIAA were shortened by single administration of beta-CCE and flumazenil. Oren-gedoku-to (30 and 300 mg/kg, p.o.) shortened the DCPDOPAC and DCP5-HIAA similarly to beta-CCE. The blended crude drugs in Oren-gedoku-to, Coptis rhizome (636.0 mg/kg, p.o.), Scutellaria root (644.4 mg/kg, p.o.) and Gardenia fruit (894.8 mg/kg, p.o.) shortened the DCPDOPAC. Coptis rhizome and Scutellaria root also shortened the DCP5-HIAA. These results suggest that GABA neuron function intensely affects the alteration of MBB-IH and Oren-gedoku-to has the intrinsic benzodiazepine-like activity. PMID- 10860489 TI - [Estimation of adverse drug reactions by the evaluation scores of subjective symptoms (complaints) and background of patients. IV. Drug eruptions]. AB - OBJECTIVES: The purpose of this study is to develop, implement, and assess an estimation procedure for preventing adverse drug reaction by subjective symptoms (complaints) of patients. This time, we focused and studied on drug eruptions. METHODS: We have built a database for CARPIS (Case Reports of Adverse Drug Reaction and Poisoning Information System) since 1987, and the case reports of adverse drug reactions accumulated in the CARPIS database to be total about 20,000. We studied 1473 cases of drug eruptions cumulated in CARPIS database. The evaluation scores were created based on the subjective symptoms and backgrounds of the patients. We estimated 1473 cases using this evaluation scores. RESULTS: We could estimate 1455 cases (98.8%) in 1473 cases to be drug eruptions using this evaluation scores. The validity of this evaluation scores were sensitivity = 98.8%, specificity = 91.0% and predictive value of positive test = 99.4%. The positive likelihood ratio was 11.0 and negative likelihood ratio was 0.01. CONCLUSIONS: This study confirmed the validity of our evaluation scores. We reported the evaluation scores about drug-induced liver diseases, drug-induced extra-pyramidal symptoms and drug-induced leukopenia before. In order to apply these evaluation scores to the clinical practice, we prepared an evaluation form for subjective symptoms and backgrounds of the patients with adverse drug reactions. PMID- 10860490 TI - [GC-MS determination of l-ephedrine and d-pseudoephedrine in human plasma]. AB - To determine l-ephedrine (E) and d-pseudoephedrine (PE) concentrations in human plasma simultaneously, we used a selected-ion monitoring method with gas chromatography-mass spectrometry (GC-MS) using deuterium-labeled E and PE as internal standards. The E and PE in human plasma were extracted with hexane ethylacetate (9/1) under alkaline conditions and were easily converted into their heptafluorobutyryl derivatives by treating with heptafluorobutyrylimidazole. The calibration curves of E and PE showed a good linearity in the range from 0.82 to 81.9 ng/ml for E, and from 0.41 to 41.0 ng/ml for PE, respectively. The limit of detection was 0.82 ng/ml for E and 0.41 ng/ml for PE in human plasma, respectively. PMID- 10860491 TI - [The anti-Candida activity of shikon]. AB - Antifungal activity of Shikon, roots of Lithospermum erythrorhizon and Arnebia euchroma was investigated in vitro. The extracts containing the pigments of Ko shikon or Nan-shikon showed the antifungal activities against Candida albicans. Acetylshikonin, one of these pigments, inhibited the fungal growth at MIC 15.6 micrograms/ml (RPMI24 h) or 3.9 micrograms/ml (YNB24 h). PMID- 10860492 TI - Enzymatic microtiter plate-based nitrate detection in environmental and medical analysis. AB - Our microtiter plate assay is based on the enzymatic reduction of nitrate by dissimilatory nitrate reductase from Pseudomonas stutzeri [EC 1.7.99.4]. Exogenous redox mediators like methyl viologen, methylene blue, and cibachron blue were applied to reduce nitrate reductase. Concentrations of 0.02-0.9 mM nitrate can be detected with +/-6% standard deviation, by using a photometric Griess reaction for the formed nitrite. Nitrate reductase is stable in the pH range 6.5-9.0 and works in the temperature range 4-76 degrees C. The assay shows no interferences with salt content up to 1 M chloride or 11 mM chlorate, and serum albumin content up to 50 mg/ml. The time demand of our two-step procedure is 20 min/100 samples. Nitrate reductase could be conserved on site of the wells of microtiter plates for at least 6 months at room temperature. The nitrate assay was applied in environmental and consumer goods analysis, and for medical diagnostics in human plasma samples. PMID- 10860493 TI - Determination of the binding parameters of drug to protein by equilibrium dialysis/piezoelectric quartz crystal sensor. AB - A novel method, equilibrium dialysis/piezoelectric quartz crystal sensor, applied to determine the binding parameters of diethyldithiocarbamate to human plasma protein is proposed. Based on the investigation of the equilibrium reaction for the binding of drug to protein, the related theoretical equations for this binding were derived. By monitoring the frequency responses of a copper-plated piezoelectric quartz crystal sensor to drug in and out of a dialysis membrane after equilibrium, the binding parameters were determined, i.e., 0.375 micromol g(-1) for beta(p), 6.496 microM for K(dp), 141.99 L mmol(-1) for K(p), and 0.043 for N. These values were in good agreement with reference values. It was found that this method may have application for studying the characteristics of the interaction between other drugs and proteins. PMID- 10860494 TI - High-resolution magic angle spinning (1)H NMR spectroscopy of intact liver and kidney: optimization of sample preparation procedures and biochemical stability of tissue during spectral acquisition. AB - High-resolution magic angle spinning (MAS) (1)H NMR spectroscopy has been used to investigate the biochemical composition of whole rat renal cortex and liver tissue samples. The effects of a number of sample preparation procedures and experimental variables have been investigated systematically in order to optimize spectral quality and maximize information recovery. These variables include the effects of changing the sample volume in the MAS rotor, snap-freezing the samples, and the effect of organ perfusion with deuterated saline solution prior to MAS NMR analysis. Also, the overall biochemical stability of liver and kidney tissue MAS NMR spectra was investigated under different temperature conditions. We demonstrate improved resolution and line shape of MAS NMR spectra obtained from small spherical tissue volume (12 microl) rotor inserts compared to 65 microl cylindrical samples directly inserted into the MAS rotors. D(2)O saline perfusion of the in situ afferent vascular tree of the tissue immediately postmortem also improves line shape in MAS NMR spectra. Snap-freezing resulted in increased signal intensities from alpha-amino acids (e.g., valine) in tissue together with decreases in renal osmolytes, such as myo-inositol. A decrease in triglyceride levels was observed in renal cortex following stasis on ice and in the MAS rotor (303 K for 4 h). This work indicates that different tissues have differential metabolic stabilities in (1)H MAS NMR experiments and that careful attention to sample preparation is required to minimize artifacts and maintain spectral quality. PMID- 10860495 TI - A scintillation proximity assay for poly(ADP-ribose) polymerase. AB - Poly(ADP-ribose) polymerase (PARP) is an abundant nuclear protein in most of the eukaryotic tissues. When activated by DNA damage, PARP synthesizes poly(ADP ribose) from NAD. Conventional radioactive PARP enzyme assay requires the separation of the polymer product from the NAD substrate, a rate-limiting step that hampers large-scale chemical library screening to identify novel small molecule PARP inhibitors. By using biotinylated NAD, we have developed a scintillation proximity assay (SPA) for PARP. We demonstrated that PARP can incorporate the biotinylated ADP-ribose units into the radioactive poly(ADP ribose) polymer, which can directly bind and excite the streptavidin-conjugated scintillation beads. PARP-SPA can be readily adapted to a 96-well format for automatic high-throughput screening for PARP inhibitors. PMID- 10860496 TI - Influence of cell fixation on chromatin topography. AB - Using in situ hybridization techniques, a fixation step must precede denaturation to prevent disintegration of the chromosomes. The analysis of nuclei fixed by paraformaldehyde, preserving the native structure (three-dimensional or 3D fixation and analysis) has become possible with the development of confocal microscopy; however, the analysis of those fixed by methanol and acetic acid, dehydrating the nuclei (two-dimensional or 2D fixation and analysis), remains a very valuable tool for practical use in diagnostics and also in many cases for research. We compared both types of fixation and analyses using different cell lines and different DNA probes. Fixation of cells by methanol and acetic acid leads to the enlargement of contact of nuclei with the slide surface, resulting in a substantial increase of nuclear diameter, flattening of the nucleus, and consequently to a distortion of gene topology. Our results indicate that chromatin structures located in the outer parts of the nuclear volume (e.g., heterochromatin of some centromeres) are relatively shifted to the membrane of these nuclei, keeping the absolute centromere-membrane distance constant. On the other hand, euchromatin located in the inner parts of the nuclear volume is not shifted outside proportionally to the increase of molecular dimensions; consequently, the relative distances for the center of nucleus to gene are smaller after methanol-acetic acid fixation. The limitations of the analysis of dehydrated preparations for practical use and in research are discussed. PMID- 10860497 TI - Catalytic chromatography. AB - Catalytic chromatography exploits both specific biological affinity and catalytic specificity to selectively purify enzymes. Two different applications are presented. Purification of EcoRI restriction endonuclease to apparent homogeneity was accomplished in a single step with significantly greater yield and purification than was obtained with affinity chromatography. An attempt to purify the multiple DNA polymerase activities of Escherichia coli was also developed. Five well-resolved peaks of DNA polymerase activity were fractionated. In this new chromatographic mode, the enzyme binds immobilized substrate coupled to a column in the absence of some required cofactor. When the missing cofactor is added, the enzyme converts substrate to product and selectively elutes from the column. PMID- 10860498 TI - A branched DNA signal amplification assay to quantitate messenger RNA of human uncoupling proteins 1, 2, and 3. AB - Uncoupling proteins (UCP) are inner mitochondrial membrane transporters which dissipate the proton gradient, releasing stored energy as heat. Three subtypes of UCP have been identified so far. The regulation of UCP expression is mainly controlled at the transcriptional level, thus making the measurement of UCP mRNA beneficial for both diagnosis and research of weight disorders and diabetes. We have developed an assay using the branched DNA signal amplification assay (bDNA assay) to quantitatively measure the mRNA levels for human UCP1, 2, and 3. UCP subtype-specific primers were designed for the assay. RNA transcripts of each UCP generated by in vitro transcription were used to validate the specificity and sensitivity of the assay. The quantitative measurement of UCP mRNA was further demonstrated with cultured cells and human tissue. A comprehensive survey of UCP expression from 17 human tissues measured by the newly developed assay is provided. The method described here offers a rapid, sensitive, specific, and quantitative assay for measurement of human UCP mRNA. PMID- 10860499 TI - Histidine-tagged ubiquitin substitutes for wild-type ubiquitin in Saccharomyces cerevisiae and facilitates isolation and identification of in vivo substrates of the ubiquitin pathway. AB - A general method for purification of any substrate of the ubiquitin pathway, the major eukaryotic proteolytic pathway, should utilize the common characteristic of covalent linkage of ubiquitin to substrate lysyl residues. The utility of a N terminal histidine-tagged ubiquitin (HisUb) for in vivo conjugation and isolation of ubiquitinated proteins by metal chelation chromatography is conditioned by the requirement that HisUb conjugate to the same set of proteins as wild-type ubiquitin. Stringent in vivo tests with Saccharomyces cerevisiae strains expressing ubiquitins only from plasmids were performed to show that HisUb could substitute for wild-type ubiquitin. The utility of HisUb as a method for purification of proteins ubiquitinated in vivo was demonstrated by metal chelation chromatography of yeast extracts expressing HisUb and immunoblotting for Rpb1, the largest subunit of RNA polymerase II. A fraction of Rpb1 was present in the ubiquitinated form in vivo. The ability to use HisUb expression in transgenic organisms that retain expression of their endogenous ubiquitin genes was demonstrated through transgenic Arabidopsis thaliana expressing HisUb or its variant HisUbK48R. UbK48R is a version of ubiquitin capable of conjugation to proteins, but cannot serve as an attachment site for ubiquitin via the major in vivo interubiquitin linkage. Whereas transgenic plants expressing HisUb showed insignificant enrichment of ubiquitinated proteins, transgenic Arabidopsis lines expressing HisUbK48R gave a much better yield. PMID- 10860500 TI - Genetically fused protein A-luciferase for immunological blotting analyses. AB - The gene expression plasmid pMALU5 for the fusion protein of protein A (SpA) with a complete sequence of firefly luciferase (Luc) was constructed. The fused gene was expressed in Escherichia coli, and the resulting SpA-Luc fusion protein was purified by one-step affinity chromatography on IgG-Sepharose. The protein retained both activities: IgG binding capability of protein A and enzymatic activity of luciferase. Blotting analyses were performed with the fusion protein to determine a tumor marker of alpha-fetoprotein (AFP). AFP was detected at the lowest detection limit of 5 pg by dot blotting and Western blotting. The SpA-Luc fusion protein provides a highly selective, sensitive, and versatile marker for blotting analyses. PMID- 10860501 TI - Detection of DNA via an ion channel switch biosensor. AB - Detection of DNA by an ion channel switch biosensor has been demonstrated in a model system, using single-stranded oligonucleotide sequences of 52-84 bases in length. Two different biotinylated probes are bound, via streptavidin, either to the outer region of a gramicidin ion channel dimer or to an immobilized membrane component. The ion channels are switched off upon detection of DNA containing complementary epitopes to these probes, separated by a nonbinding region, at nanomolar levels. The DNA cross-links the ion channel to the immobilized species, preventing ions passing through the channel. Addition of DNase I after the target DNA has been added switches the ion channels on. The DNA response is dependent on the rate of hybridization of the individual probes to their complementary epitopes, as shown by using a single probe against DNA containing a repeat of the complementary epitope. These results were correlated with hybridization rates determined using surface plasmon resonance (BIAcore 2000), and with free energies of dimer formation for the probes. PMID- 10860502 TI - Modified telomeric repeat amplification protocol: a quantitative radioactive assay for telomerase without using electrophoresis. AB - A polymerase chain reaction (PCR)-based radioactive telomerase assay was developed in our laboratory which is quantitative and does not require electrophoretic evaluation (designated as TP-TRAP; it utilizes two reverse primers). The main steps of the assay include (1) extension of a 20-mer oligonucleotide substrate (MTS) by telomerase, (2) amplification of the telomerase products in the presence of [(3)H]dTTP using the substrate oligonucleotide and two reverse primers (RPC3, 38 mer; RP, 20 mer), (3) isolation of the amplified radioactive dsDNA by precipitation and filtration, (4) determination of the radioactivity of the acid-insoluble DNA. The length of the telomerase products does not increase on amplification. This valuable feature of the assay is achieved by utilization of the two reverse primers and a highly specific PCR protocol. The assay is linear, accurate, and suitable for cell biological studies where slight quantitative differences in telomerase activity must be detected. The assay is also suitable for screening and characterization of telomerase inhibitors, as shown with a chemically modified oligonucleotide reverse transcriptase inhibitor [(s(4)dU)(35)]. PMID- 10860503 TI - Determination of ascorbic acid and dehydroascorbic acid in biological samples by high-performance liquid chromatography using subtraction methods: reliable reduction with tris[2-carboxyethyl]phosphine hydrochloride. AB - Determination of dehydroascorbic acid in biological samples most commonly involves indirect measurement. The concentration is calculated by subtraction of the measured ascorbic acid concentration from that of total ascorbic acid analyzed after reduction of the dehydroascorbic acid present; a methodology also referred to as subtraction methods. Consequently, successful determination of dehydroascorbic acid is dependent on proper sample handling, quantitative reduction of the compound, and accurate quantification of both ascorbic acid and total ascorbic acid. In this paper, the recently introduced reductant tris[2 carboxyethyl]phosphine (TCEP) is evaluated as a reliable alternative to the commonly used reducing agent dithiothreitol (DTT). The results show that TCEP offers a more efficient reduction of dehydroascorbic acid at low pH compared to that of DTT. Moreover, while DTT maintains a reducing sample environment for less than 24 h, TCEP show complete protection from oxidation of ascorbic acid for at least 96 h following sample preparation. Removal of TCEP prior to analysis is unnecessary. A revised HPLC-EC method incorporating TCEP as reductant as well as the coanalysis of isoascorbic acid and uric acid is presented. The within- and between-day coefficients of variation for the complete assay are less than 1.5 and 3.5% for all analytes. As a whole, the method presented here is simpler and more reliable than existing methods. PMID- 10860504 TI - Cytostar-T scintillating microplate assay for measurement of sodium-dependent bile acid uptake in transfected HEK-293 cells. AB - Real-time measurements of bile acid uptake into HEK-293 cell monolayers expressing the human sodium/bile acid cotransporters have been demonstrated using Cytostar-T microplates with an integral scintillating base. In these 96-well microplates, which permits culturing and observation of adherent cell monolayers, uptake of (14)C-labeled glycocholate and taurocholate into transfected HEK-293 cells was time-dependent, sodium-stimulated, and saturable. The sodium-activated uptake of 30 microM [(14)C]glycocholate (GC) via the ileal (IBAT) and liver (LBAT) transporters was 30-40 times higher than GC uptake in a sodium-free background. In addition, ouabain inhibition of the plasma membrane Na(+), K(+) ATPase, causing the sodium gradient to collapse, resulted in total loss of glycocholate transport. Induction of gene expression by sodium butyrate showed that the amount of labeled bile acid accumulated in the cell monolayers at steady state was a function of the total amount of transporter expressed. Uptake of labeled bile acids was inhibited both by the specific IBAT inhibitor, 2164U90, and by various bile acids. No major difference was observed between IBAT and LBAT in their specificity for the bile acids tested while the dihydroxy bile acids had the highest affinity for both the transporters studied. The Cytostar-T proximity assay has been demonstrated to be an accurate and reproducible method for monitoring specific bile acid transport in transfected mammalian cells and the results are similar to those obtained by traditional methods. We conclude that the technique is an attractive approach to the cellular study of membrane transport of radiolabeled solutes in general and suggest a role in screening and characterization of novel transport inhibitors. PMID- 10860505 TI - Effects of dithiothreitol on protein activity unrelated to thiol-disulfide exchange: for consideration in the analysis of protein function with Cleland's reagent. AB - Dithiothreitol (DTT) is widely used to reduce disulfide bonds in the analysis of protein structure and function. However, thiol-disulfide exchange is not the only mechanism whereby DTT can alter protein function. We observe that DTT diminishes the carbohydrate binding activity of a cysteineless mutant of pigpen as well as it inhibits the intact molecule. Lack of inhibition by threitol, a derivative of the four-carbon sugar threose, indicates that the thiol groups of DTT are required for inhibition, and also that DTT is not acting as a simple carbohydrate competitor. Moreover, inhibition of pigpen-carbohydrate binding is not likely due to metal chelation because pigpen binding to carbohydrate is insensitive to EDTA and 1, 10-phenanthroline, which would otherwise be expected to mimic the DTT effect. Our results suggest that DTT can interact with protein domains in the absence of cysteine residues, and that the biochemical reactivity of DTT is not necessarily one and the same with its assumed biochemical specificity. PMID- 10860506 TI - Development of a scintillation proximity assay for beta-ketoacyl-acyl carrier protein synthase III. AB - Assays of beta-ketoacyl-acyl carrier protein synthases III (KASIII; FabH), a key enzyme initiating bacterial type II fatty acid biosynthesis, usually involve incubation of radiolabeled acetyl-coenzyme A and malonyl-acyl carrier protein (MACP). The radiolabeled acetoacetyl-ACP product is precipitated and separated from the substrate before quantitation. We have developed a scintillation proximity assay (SPA) where use of biotinylated MACP (BMACP) allows the generation of a biotinylated acetoacetyl-ACP. This product, when captured by the streptavidin-coated scintillant-impregnated microspheres, generates an SPA signal. A BMACP K(m) of 7.1 microM was determined using this SPA with the Streptomyces glaucescens FabH. A similar MACP K(m) (6 microM) was determined in a precipitation assay, demonstrating that BMACP is an effective substrate for FabH. IC(50) values of 15.2 microM (SPA) and 24.8 microM were obtained with iodoacetamide and the S. glaucescens FabH. Comparable IC(50) values of 160 microM (SPA) and 125 microM were also obtained with the antibiotic thiolactomycin and the Escherichia coli FabH. These observations demonstrate that FabH inhibitors can be readily detected using a SPA with BMACP and that the effectiveness of inhibitors in the SPA is comparable to that obtained using MACP and a standard TCA precipitation assay. A FabH SPA adaptable to high-throughput screening should facilitate the discovery of potential novel antibiotics. PMID- 10860507 TI - Quantitative measurement of sphingosine 1-phosphate by radioreceptor-binding assay. AB - In Chinese hamster ovary cells overexpressing Edg-1, one of the sphingosine 1 phosphate (S1P) receptor subtypes, [(3)H]S1P binding was displaced by unlabeled S1P with IC(50), a half-maximal concentration to inhibit the binding, of about 20 nM. This radioreceptor binding was used for quantitative measurement of S1P. Among the various lipids employed, only sphingosylphosphorylcholine (SPC), other than S1P, practically displaced the binding; however, the potency of SPC was about 100 to 1000 times less than that of S1P. Thus, SPC bound to the S1P receptors inefficiently. Furthermore, before the application of test samples to this assay, S1P was partially purified: the lipid was extracted first into the aqueous phase and separated from other lipids under alkaline conditions, and then reextracted into the chloroform phase under acidic conditions. With this assay, we could specifically and quantitatively measure S1P from 2 to 40 pmol per assay well in biological samples including serum samples and various tissues. This assay also allowed us to measure the change in cellular S1P content in U937 cells after treatment with exogenous sphingosine. PMID- 10860508 TI - Affinity biosensor for avidin using a double functionalized dendrimer monolayer on a gold electrode. AB - We have developed an affinity biosensor system based on avidin-biotin interaction on a gold electrode. As the building block of an affinity-sensing monolayer, a fourth-generation (G4) poly(amidoamine) dendrimer having partial ferrocenyl tethered surface groups was prepared and used. The unmodified surface amine groups from dendrimers were functionalized with biotinamidocaproate, and the biotinylated and electroactive dendritic monolayer was constructed on a gold electrode for the affinity-sensing surface interacting with avidin. An electrochemical signal from the affinity biosensor was generated by free glucose oxidase in electrolyte, depending on the degree of coverage of the sensing surface with avidin. The sensor signal decreased correlatively with increasing avidin concentration and approached a minimum level when the sensing surface was fully covered with avidin. The detection limit of avidin was about 4.5 pM, and the sensor signal was linear ranging from 1.5 pM to 10 nM under optimized conditions. From the kinetic analysis using the biotinylated glucose oxidase, an active enzyme coverage of 2.5 x 10(-12) mol/cm(2) on the avidin-pretreated surface was registered, which demonstrates the formation of a spatially ordered and compact protein layer on the derivatized electrode surface. PMID- 10860509 TI - Competitive hybridization: theory and application in isolation and quantification of differentially regulated genes. AB - Competitive hybridization is a simple yet powerful method that was developed to screen cDNA libraries for differentially regulated genes. The method is based on competition between unlabeled cDNA from the mRNA of one sample and labeled cDNA from another sample. By manipulating the amount of competing unlabeled cDNA, background signals from the nonregulated genes can be increased or reduced, enabling the signals from differentially regulated genes to be contrasted and to be identified in a quantitative manner. To demonstrate the feasibility of the method, we screened a citrus cDNA library for ethylene-induced genes and identified three genes with different levels of ethylene induction. The mathematical basis of the method and its possible application in gene chip technology are discussed. PMID- 10860510 TI - Identification of sulfonylureas in serum by electrospray mass spectrometry. AB - Identification of sulfonylureas in serum is important in the diagnosis of hypoglycemic crisis of unknown origin. Methods based on HPLC with UV or fluorescence detection may give false positive results. Mass spectrometry may successfully avoid this problem. The described method allows the simultaneous identification and quantification of tolbutamide, chlorpropamide, glibenclamide, and glipizide in human serum using one of the tested sulfonylureas as the internal standard. Serum purification is carried out by solid-phase extraction with ENVI-C18 cartridges and samples are analyzed by liquid chromatography electrospray mass spectrometry. For all drugs, the limit of detection and the limit of quantification are about 2 and 10 ng/ml, respectively. PMID- 10860511 TI - A fiber-optic microarray biosensor using aptamers as receptors. AB - A fiber-optic biosensor using an aptamer receptor has been developed for the measurement of thrombin. An antithrombin DNA aptamer was immobilized on the surface of silica microspheres, and these aptamer beads were distributed in microwells on the distal tip of an imaging fiber. A different oligonucleotide bead type prepared using the same method as the aptamer beads was also included in the microwells to measure the degree of nonspecific binding. The imaging fiber was coupled to a modified epifluorescence microscope system, and the distal end of the fiber was incubated with a fluorescein-labeled thrombin (F-thrombin) solution. Nonlabeled thrombin could be detected using a competitive binding assay with F-thrombin. The aptamer beads selectively bound to the target and could be reused without any sensitivity change. The fiber-optic microarray system has a detection limit of 1 nM for nonlabeled thrombin, and each test can be performed in ca. 15 min including the regeneration time. PMID- 10860512 TI - Electrophoresis of proteins and protein-protein complexes in a native agarose gel. PMID- 10860513 TI - Phosphohistidine analysis using reversed-phase thin-layer chromatography. PMID- 10860514 TI - An in-gel assay of a recombinant western corn rootworm (Diabrotica virgifera virgifera) cysteine proteinase expressed in yeast. PMID- 10860515 TI - Polymerase dependence of autosticky polymerase chain reaction. PMID- 10860516 TI - A noncommercial dual luciferase enzyme assay system for reporter gene analysis. PMID- 10860517 TI - Reaction of Tris(2-carboxyethyl)phosphine (TCEP) with maleimide and alpha haloacyl groups: anomalous elution of TCEP by gel filtration. PMID- 10860518 TI - Geophagy among primates: adaptive significance and ecological consequences. AB - We review geophagy, or soil ingestion, in primates. This behaviour is widespread and is presumed to be important to health and nutrition. Primates may engage in geophagy for one or a combination of reasons. Here we present, and make a preliminary assessment of, six nonexclusive hypotheses that may contribute to the prevalence of geophagy. Four hypotheses relate to geophagy in alleviating gastrointestinal disorders or upsets: (1) soils adsorb toxins such as phenolics and secondary metabolites; (2) soil ingestion has an antacid action and adjusts the gut pH; (3) soils act as an antidiarrhoeal agent; and (4) soils counteract the effects of endoparasites. Two hypotheses pertain to geophagy in supplementing minerals and/or elements: (5) soils supplement nutrient-poor diets and (6) soils provide extra iron at high altitudes. In addition to these hypotheses, geophagy may satiate olfactory senses, serve as a famine food and finally may have no function at all. We draw together a large body of information from various sources to assess these hypotheses and suggest some tests to understand the function of geophagy. Our review suggests that primates engage in geophagy for a number of reasons that are nonexclusive. We conclude that mineral supplementation, adsorption of toxins, treatment of diarrhoea and pH adjustment of the gut seem the most plausible reasons why primates engage in geophagy. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860519 TI - Referential labelling in Diana monkeys. AB - Animal semantic communication has received considerable theoretical and empirical attention because of its relevance to human language. Advances have been made by studies of alarm-call behaviour in nonhumans. In monkeys, for example, there is evidence that recipients have a fairly sophisticated understanding of a call's meaning; that is, the predator type usually associated with a certain alarm call. Little is known, however, about the mental mechanisms that drive call production in nonhuman primates. In some nonprimate species, it has been found that signallers do not respond to a predator's physical features but instead seem to respond to its relative threat or direction of attack. In these species, therefore, alarm calls do not denote different predator categories but simply reflect different types or levels of danger. Because different predator categories typically impose different types and degrees of threat it is entirely possible that nonhuman primates also respond to threat rather than a predator's category. This study examined how wild Diana monkeys, Cercopithecus diana, of the Tai forest, Ivory Coast, label predation events. By altering playback stimuli and the position of a concealed speaker, I investigated whether Diana monkeys respond with acoustically different alarm calls depending on a predator's (1) distance (close versus far), (2) elevation (above versus below), or (3) category (eagle versus leopard). Analysis of male and female alarm-call behaviour showed that Diana monkeys consistently responded to predator category regardless of immediate threat or direction of attack. Data further suggested that, in addition to predator category, monkeys' alarm calls might also convey information about the predator's distance. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860520 TI - Behaviour plasticity without learning: phenotypic and genetic variation of naive Daphnia in an ecological trade-off. AB - The swimming behaviour of adult Daphnia largely governs their depth, which has a direct effect on both individual foraging success and predation avoidance. We treated individual swimming behaviour as a threshold character and used directional changes in average clonal depth within experimental tubes as a test for character plasticity. We compared the swimming behaviours of two cohabiting, phenotypically similar Daphnia (Daphnia galeata and Daphnia galeata-Daphnia rosea hybrid) to determine (1) whether there is inherited variation (H(2)) for different traits (responses to hunger and predator cues); (2) whether changes in genetic parameters (norm of reaction and character state) across four environments could be simulated by combinations of the presence or absence of a predator cue and high or low hunger levels; and (3) whether these Daphnia would respond to directional selection, particularly in a trade-off environment (high hunger and predator cue treatments). Responses of both D. galeata and the D. galeata-rosea hybrid to hunger and a predator cue and the trade-off environment were plastic. Daphnia galeata expressed significant genetic variation within (H(2)) and between environments (heritability of plasticity). Both the character state and norm of reaction estimates of heritable variation in the hybrid were close to zero. Genetic correlations were positive and stable across six environmental pairs in Daphnia galeata. Most hybrid genetic correlations were not significant. The phenotypic distributions of both D. galeata and the hybrid were bimodal in the trade-off environment. The D. galeata distribution was partly due to between-clone variation and the hybrid distribution was almost entirely due to within-clone variation. Genetic variation expressed by D. galeata in the trade off environment appears to depend on both clone by environment interactions and stable inherited differences. These results indicate that while plastic phenotypic responses cause a similar opportunity for selection in D. galeata and the hybrid, their responses to selection would be different in the trade-off and in related environments. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860521 TI - Flower choice by naive young crab spiders and the effect of subsequent experience. AB - Initial responses of naive individuals to critical environmental stimuli provide important information about the innate contribution to behaviour, and subsequent responses to the same stimuli may show the role of experience in mediating those initial responses. To test the role of these factors, I measured initial patch choices and giving-up responses of just-emerged, naive, second-instar crab spiders, Misumena vatia, on several hunting sites they encountered after leaving their natal nests. In follow-up tests I measured the effects of these experiences on subsequent patch choice decisions. The choice of hunting sites is a vital decision at all stages of the life cycle for sit-and-wait predators such as Misumena. In their initial tests these spiderlings remained more frequently on goldenrod (Solidago spp.) flowers than on green or yellow goldenrod buds, a preference they retained through tests run on 5 consecutive days. Individuals on green and yellow buds shifted sites more quickly and frequently than those from flowers, and made most of these moves to flowers, which attracted many more prey than did buds. These differences were not affected by age, energetic condition, or loss of information over the period of the experiment. Once spiderlings moved from buds, they showed a high, increasing tendency to move from buds in subsequent runs, those from flowers showed a consistently low tendency. These results suggest that spiderlings retain their innate behavioural patterns through the second instar, but that experience also plays a modest role in patch choice at this stage. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860522 TI - The evolution of postcopulatory displays in dabbling ducks (Anatini): a phylogenetic perspective. AB - Although postcopulatory displays are widespread in animals, little is known about their function or the intended receiver. The postcopulatory displays of dabbling ducks are among the best described for any animal group. We documented the presence of initial and additional postcopulatory displays in nearly all dabbling duck species. We then reconstructed the evolution of postcopulatory displays in dabbling ducks using a phylogeny derived from mitochondrial DNA sequences. The display immediately following copulation (the initial display) is highly stereotyped in most species and shows extreme phylogenetic conservation. In contrast, the performance of additional displays is less stereotyped and less phylogenetically conservative. We review the possible functions of postcopulatory displays. Using evidence from display orientation, display form and phylogenetic reconstruction, we suggest that the most likely functions of postcopulatory displays in dabbling ducks are pair bond maintenance, individual identification, or signalling a successful copulation. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860523 TI - Contingencies in the behaviour of the crab Heterozius rotundifrons. AB - We recorded the responses of individual intertidal crabs, Heterozius rotundifrons, to stimuli presented singly and in combinations in the laboratory. Undisturbed crabs did not respond to the introduction of odour from a crushed conspecific but did respond strongly to food odour. Undisturbed crabs responded equally to food odour alone and a combination of food and odour from a crushed conspecific. When tactile stimulation was applied, as when the crab is grasped by a predator, individual H. rotunidfrons assumed a rigid, appendage-extended posture for several minutes. Tests with predatory fish showed that this catatonic posture is a very effective predator-defence mechanism. The duration of the catatonic state was decreased by the addition of food odour but increased by the addition of alarm odour (crushed conspecific) or the combination of alarm and food odours. Thus, which chemical stimulus was dominant was reversed by tactile input (i.e. dominance was contingent upon context). The effect of alarm odour on food odour responses lasted 4 h. Visual input in the form of a shadow passing over the crabs, either before or after tactile induction of the catatonic state, also increased the duration of that state. However, the duration of the catatonic state following exposure to both cues associated with danger (shadow+alarm odour) was similar to that of the control level. The crabs appeared to switch strategies when three cues associated with danger (tactile grasping, alarm odour and shadows) were detected, either simultaneously or over a 4-h period. The results illustrate the highly contingent nature of the behaviour of these crabs. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860524 TI - 'Culture' in quail: social influences on mate choices of female Coturnix japonica. AB - We have shown previously that after a focal female Japanese quail, Coturnix japonica, sees a conspecific male mating, the focal female's tendency to affiliate and to mate with that male is significantly increased. Here we describe two experiments demonstrating that a focal female quail that has seen a conspecific male mating subsequently shows an enhanced tendency to affiliate not only with that particular male, but also with other males that share his characteristics. The results have important implications for our understanding of the role of social learning in the evolution of male characteristics. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860525 TI - Female-female aggression and female mate choice on black grouse leks. AB - We studied female-female aggression in relation to female mate choice in black grouse, Tetrao tetrix, in central Finland, in 1994-1998. Aggression occurred on average every other minute when there was more than one female on a territory, and aggressive behaviour was most prominent when several females attended the lek. Interactions tended to be proportionally most frequent on the territories of the highest-ranking males, although not significantly so. Females that were chased by other females did not mate with lower-ranking males than their aggressors did. Furthermore, chased females were only rarely (6% of cases) forced to move off the territory by agonistic interactions and copulations were disrupted by other females even less often (3% of cases). The choice of a mating territory did not depend on the outcome of aggression even though the aggressors were more likely to mate on the territory where aggression occurred than elsewhere. There was a marginally significant tendency for aggressors to mate earlier in the season. Females placed themselves further away from other females on the territory when soliciting a copulation than just before aggression. Our results suggest that aggression between females does not effectively constrain female choice in black grouse. Its function may be to aid females to secure undisturbed mating opportunities for themselves rather than to prevent others from mating with a particular male. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860526 TI - Patch time allocation and patch sampling by foraging great and blue tits. AB - The rate at which parents deliver energy to their brood is an important factor in avian reproduction because poor condition caused by malnutrition may reduce the offspring's survival to breeding. Models of central place foraging predict that nesting parents should optimize their prey delivery rate by minimizing travelling distances and by selecting patches where the gain per unit cost is high. I investigated the allocation of searching time amongst food patches in the home ranges of breeding great tits, Parus major, and blue tits P. caeruleus, by radiotracking. The density of locations in individual trees was positively correlated with prey biomass within trees and negatively with the distance of the trees from the nest. These two factors explained 52% of the variance in the allocation of the birds' search time. In rich patches, food was reduced considerably within 20 m of the nests, and the birds' travelling distances increased significantly during the nestling period. In parallel to foraging selectively in rich resources near the nest, the birds continually sampled the trees in their territory. The average surplus search time due to resource exploration was 1.52 times (range 1.25-1.99) the expected search time if the birds had exclusively used the most profitable patch. Despite considerable effort in patch sampling, the overall search time per unit prey was 30% better than expected by an equal use of trees. The results suggest that foraging tit parents come close to the maximum rate of prey delivery possible in a given patch distribution. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860527 TI - Biparental care and obligate monogamy in the rock-haunting possum, Petropseudes dahli, from tropical Australia. AB - Monogamy is rare among mammals, including marsupials. I studied the social organization of the little-known rock-haunting possum in Kakadu National Park in Northern Australia. Preliminary field observations revealed that the majority of possums live in cohesive groups consisting of a female-male pair and young, suggesting a monogamous mating system. I used radiotracking to determine home range patterns, and observations to measure the degree of symmetry between the sexes in maintaining the pair bond and initiating changes in group activity. I also measured the extent of maternal and paternal indirect and direct care. Nocturnal observations and radiotelemetric data from 3 years showed that six possum groups maintained nonoverlapping home ranges with long-term consorts and young sharing dens. Males contributed more than females to maintaining the pair bond but they contributed equally to parental care. For the first time, the parental behaviours of bridge formation, embracing, marshalling of young, sentinel behaviour and tail beating are reported in a marsupial. Males participated to a high degree in maintaining relationships with one mate and their offspring. Collectively, these results suggest that the mating system of this wild population of rock-haunting possums is obligate social monogamy. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860528 TI - Helper contributions in the cooperatively breeding laughing kookaburra: feeding young is no laughing matter. AB - I studied the contributions of individuals to incubation and nestling feeding in a population of cooperatively breeding laughing kookaburras, Dacelo novaeguineae. In most cooperatively breeding birds where nest success is limited by nestling starvation, related helpers increase the overall level of provisioning to the nest, thus boosting the production of nondescendent kin. However, although partial brood loss is the largest cause of lost productivity in kookaburra nests, additional helpers failed to increase overall provisioning. Instead, all group members, but especially helpers, reduced their feeding contributions as group size increased. Breeders and helpers reduced the size of prey delivered, and helpers also reduced the number of feeding visits. An important benefit of helping in kookaburras may be to allow all group members to reduce their effort. Within groups, contributions to care depended on status, sex, group size and the brood size. Breeding males delivered the most food. Breeding females provisioned less than their partner, but their effort was comparable to that of male helpers. Female helpers contributed the least food. Incubation effort followed similar patterns. The relatedness of helpers to the brood had no impact on their provisioning. Across all group sizes, helpers generally brought larger items to the nest than breeders. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860529 TI - Differential responses of kin and nonkin salmon to patterns of water flow: does recirculation influence aggression? AB - Studies conducted in laboratory streams have shown that juvenile salmonid fish (parr) can differentiate between their kin and nonkin and may be less aggressive towards their kin. Chemicals produced by salmonids are also known to be used as cues to aid kin recognition. We tested the hypothesis that the ability of Atlantic salmon, Salmo salar, to recognize kin and hence modulate their level of aggression is influenced by the amount of water recirculation, which would be expected to affect the concentration of odour. Levels of aggression were similar between pairs of kin and pairs of nonkin when there was negligible recirculation of water. However, when water was recirculated, pairs of nonkin were on average 1.56 times more aggressive than pairs of kin, owing to an increase in attacks by the dominant fish. The data do not support the idea that odour concentration affects kin recognition and hence reduces aggression among siblings. Instead they indicate that recirculation of water instigates heightened aggression in dominant fish but only towards nonkin subordinates. The study suggests that the advantages for juvenile salmonids of associating with kin vary spatially, being influenced by water flow dynamics. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860530 TI - Olfactory information transfer in the honeybee: compared efficiency of classical conditioning and early exposure. AB - We investigated the ability of honeybees, Apis mellifera, to use olfactory information gained in a given experimental context, in other contexts. First, restrained bees were subjected to a Pavlovian associative learning procedure, based on the conditioning of the proboscis extension response (PER), where a floral odour was paired with a sugar reward. We observed the orientation behaviour of conditioned and naive bees in a four-armed olfactometer with four contiguous fields either scented with the conditioning odour or unscented. Information transfer was clearly shown, conditioned bees orienting towards the conditioning odour, whilst naive bees shunned it. Second, the effect of passive olfactory exposures during the bees' development was assessed in two behavioural contexts: either orientation in the olfactometer or a PER conditioning procedure. Two exposure periods were applied: (1) the pupal stage (9 days before emergence); (2) the early adult stage (8 days after emergence). No effect of preimaginal exposure was recorded, but exposure during the early adult stage induced a higher choice frequency of the odour field in the olfactometer, and lower learning performance in the PER conditioning assay. These observations show that olfactory information gained during development can modify bees' later behaviour in different contexts: this is another instance of olfactory information transfer in bees. These results also suggest that nonassociative learning phenomena, taking place at a critical period during development, might be involved in the maturation of the bees' olfactory system, and in the organization of odour mediated behaviours. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860531 TI - Towards the integration of social dominance and spatial structure. AB - My aim was to show how individual-oriented (or artificial life) models may provide an integrative background for the development of theories about dominance by including effects of spatial structure. Dominance interactions are thought to serve two different, contrasting functions: acquisition of high rank and reduction of aggression. The model I present consists of a homogeneous virtual world inhabited by artificial agents whose actions are restricted to grouping and dominance interactions in which the effects of winning and losing are self reinforcing. The two functions are implemented as strategies to initiate dominance interactions and the intensity of aggression and dominance perception (direct or memory based) are varied experimentally. Behaviour is studied by recording the same behavioural units as in real animals. Ranks appear to differentiate more clearly at high than at low intensity of aggression and also more in the case of direct than of memory-based rank perception. Strong differentiation of rank produces a cascade of unexpected effects that differ depending on which function is implemented: for instance, a decline in aggression, spatial centrality of dominants and a correlation between rank and aggression. Insight into the origination of these self-organized patterns leads to new hypotheses for the study of the social behaviour of real animals. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860532 TI - Sex differences in weather sensitivity can cause habitat segregation: red deer as an example. AB - Sex differences in habitat use (habitat segregation) are widespread in sexually dimorphic ungulates. A possible cause is that males are more sensitive to weather than females, leading to sex differences in sheltering behaviour (the 'weather sensitivity hypothesis'). However, this hypothesis has never been tested. We considered the allometric rates of net energy gain during times of cold weather and food shortage in a model. We argue that the higher absolute heat losses relative to intake rates of larger ungulates should indeed lead to higher weather sensitivity in males than in females. Furthermore, we tested the weather sensitivity hypothesis empirically in red deer, Cervus elaphus, on the Isle of Rum, U.K. We predicted that (1) use of relatively exposed, high-quality forage habitat should be negatively influenced by bad weather; and (2) this influence should be stronger in males. We found that bad weather (strong wind, low temperature, heavy rain) in winter and spring influenced use of high-quality forage habitat negatively in all deer; that adult males responded more strongly to low temperature and strong wind than did females; and that adult males foraged on windy days at better sheltered sites than did females. Thus, the weather sensitivity hypothesis is supported both theoretically and empirically. We suggest that the weather sensitivity hypothesis can potentially explain winter habitat segregation in a large number of ungulate species. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860534 TI - A simple technique for estimating a constant daily survival rate for reproductive attempts. PMID- 10860533 TI - How do ants assess food volume? AB - By comparing the behaviour of Lasius niger scouts at sucrose droplets of different volumes, we empirically identified the criterion used by each scout to assess the amount of food available as well as the rules governing its decision to lay a recruitment trail. When scouts discovered food volumes exceeding the capacity of their crop (3 or 6 ul), 90% immediately returned to the nest laying a recruitment trail. In contrast, when smaller food droplets (0.3, 0.7 or 1 ul) were offered, several scouts stayed on the foraging area, presumably exploring it for additional food. If unsuccessful, they returned to the nest without laying a trail. The droplet volume determined the percentage of trail-laying ants but had no influence on the intensity of marking when this was initiated. The key criterion that regulated the recruiting behaviour of scouts was their ability to ingest their own desired volume. This volume acted as a threshold triggering the trail-laying response of foragers. Collective regulation of foraging according to food size resulted from the interplay between the distribution of these desired volume thresholds among colony members and the food volume available. We relate some aspects of the foraging ecology of aphid-tending ants to this decision making process. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10860536 TI - Block of cardiac ATP-sensitive K(+) channels reduces hydroxyl radicals in the rat myocardium. AB - The present study examined whether opening of an ATP-sensitive K(+) (K(ATP)) channel can induce hydroxyl free radical ((*)OH) generation in the rat myocardium. Sodium salicylate in Ringer's solution (0.5 nmol/microl/min) was infused directly through a microdialysis probe to detect the generation of (*)OH as reflected by the nonenzymatic formation of 2,3-dihydroxybenzoic acid (DHBA). Induction of cromakalim (100 microM), a K(ATP) channel opener, through the microdialysis probe significantly increased the level of 2,3-DHBA. Another K(ATP) channel opener, nicorandil, also increased the level of 2,3-DHBA. When iron(II) was administered to cromakalim-pretreated animals, a marked elevation of DHBA was observed, compared with iron(II) only-treated animals. A positive linear correlation between iron(II) and formation of (*)OH, trapped as DHBA in the dialysate, was shown (r(2) = 0.988). When corresponding experiments were performed with nicorandil-treated animals, a positive linear correlation between iron(II) and DHBA in the dialysate was shown (r(2) = 0.988). However, the presence of glibenclamide (1-50 microM) decreased the cromakalim-induced 2,3-DHBA formation in a concentration-dependent manner (IC(50) = 9.1 microM). 5 Hydroxydecanoate (5-HD; 100 microM), another K(ATP) channel antagonist, also decreased cromakalim-induced (*)OH formation. The IC(50) value for 5-HD against cromakalim-evoked increase in 2,3-DHBA was 107.2 microM. In the presence of glibenclamide (10 microM), the heart was subjected to myocardial ischemia for 15 min by occlusion of the left anterior descending coronary artery (LAD). When the heart was reperfused, the normal elevation of 2,3-DHBA in the heart dialysate was not observed in animals pretreated with glibenclamide (10 microM). When corresponding experiments were performed with 5-HD (100 microM) pretreated animals, the same results were obtained. These results suggest that opening of cardiac K(ATP) channels may cause (*)OH generation. PMID- 10860537 TI - Structural and functional characterization of BnSP-7, a Lys49 myotoxic phospholipase A(2) homologue from Bothrops neuwiedi pauloensis venom. AB - BnSP-7, a Lys49 myotoxic phospholipase A(2) homologue from Bothrops neuwiedi pauloensis venom, was structurally and functionally characterized. Several biological activities were assayed and compared with those of the chemically modified toxin involving specific amino acid residues. The cDNA produced from the total RNA by RT-PCR contained approximately 400 bp which codified its 121 amino acid residues with a calculated pI and molecular weight of 8.9 and 13,727, respectively. Its amino acid sequence showed strong similarities with several Lys49 phospholipase A(2) homologues from other Bothrops sp. venoms. By affinity chromatography and gel diffusion, it was demonstrated that heparin formed a complex with BnSP-7, held at least in part by electrostatic interactions. BnSP-7 displayed bactericidal activity and promoted the blockage of the neuromuscular contraction of the chick biventer cervicis muscle. In addition to its in vivo myotoxic and edema-inducing activity, it disrupted artificial membranes. Both BnSP-7 and the crude venom released creatine kinase from the mouse gastrocnemius muscle and induced the development of a dose-dependent edema. His, Tyr, and Lys residues of the toxin were chemically modified by 4-bromophenacyl bromide (BPB), 2-nitrobenzenesulfonyl fluoride (NBSF), and acetic anhydride (AA), respectively. Cleavage of its N-terminal octapeptide was achieved with cyanogen bromide (CNBr). The bactericidal action of BnSP-7 on Escherichia coli was almost completely abolished by acetylation or cleavage of the N-terminal octapeptide. The neuromuscular effect induced by BnSP-7 was completely inhibited by heparin, BPB, acetylation, and CNBr treatment. The creatine kinase releasing and edema-inducing effects were partially inhibited by heparin or modification by BPB and almost completely abolished by acetylation or cleavage of the N-terminal octapeptide. The rupture of liposomes by BnSP-7 and crude venom was dose and temperature dependent. Incubation of BnSP-7 with EDTA did not change this effect, suggesting a Ca(2+)-independent membrane lytic activity. BnSP-7 cross-reacted with antibodies raised against B. moojeni (MjTX-II), B. jararacussu (BthTX-I), and B. asper (Basp-II) myotoxins as well as against the C-terminal peptide (residues 115 129) from Basp-II. PMID- 10860538 TI - Mechanistic studies of Escherichia coli adenosine-5'-phosphosulfate kinase. AB - Adenosine-5'-phosphosulfate kinase (APS kinase) catalyzes the formation of 3' phosphoadenosine 5'-phosphosulfate (PAPS), the major form of activated sulfate in biological systems. The enzyme from Escherichia coli has complex kinetic behavior, including substrate inhibition by APS and formation of a phosphorylated enzyme (E-P) as a reaction intermediate. The presence of a phosphorylated enzyme potentially enables the steady-state kinetic mechanism to change from sequential to ping-pong as the APS concentration decreases. Kinetic and equilibrium binding measurements have been used to evaluate the proposed mechanism. Equilibrium binding studies show that APS, PAPS, ADP, and the ATP analog AMPPNP each bind at a single site per subunit; thus, substrates can bind in either order. When ATPgammaS replaces ATP as substrate the V(max) is reduced 535-fold, the kinetic mechanism is sequential at each APS concentration, and substrate inhibition is not observed. The results indicate that substrate inhibition arises from a kinetic phenomenon in which product formation from ATP binding to the E. APS complex is much slower than paths in which product formation results from APS binding either to the E. ATP complex or to E-P. APS kinase requires divalent cations such as Mg(2+) or Mn(2+) for activity. APS kinase binds one Mn(2+) ion per subunit in the absence of substrates, consistent with the requirement for a divalent cation in the phosphorylation of APS by E-P. The affinity for Mn(2+) increases 23-fold when the enzyme is phosphorylated. Two Mn(2+) ions bind per subunit when both APS and the ATP analog AMPPNP are present, indicating a potential dual metal ion catalytic mechanism. PMID- 10860539 TI - Inhibition of nitric oxide synthase isoforms by tris-malonyl-C(60)-fullerene adducts. AB - C(3)-tris-malonyl-C(60)-fullerene and D(3)-tris-malonyl-C(60)-fullerene derivatives inhibit citrulline and NO formation by all three nitric oxide synthase isoforms in a manner fully reversible by dilution. The inhibition of citrulline formation by C(3)-tris-malonyl-C(60)-fullerene occurs with IC(50) values of 24, 17, and 123 microM for the neuronal, endothelial, and inducible nitric oxide synthase (NOS) isoforms, respectively. As measured at 100 microM l arginine, neuronal NOS-catalyzed nitric oxide formation was inhibited 50% at a concentration of 25 microM C(3)-tris-malonyl-C(60)-fullerene. This inhibition was a multisite, positively cooperative inhibition with a Hill coefficient of 2.0. C(3)-tris-malonyl-C(60)-fullerene inhibited the arginine-independent NADPH oxidase activity of nNOS with an IC(50) value of 22 microM but had no effects on its cytochrome c reductase activity at concentrations as high as 300 microM. The inhibition of nNOS activity by C(3)-tris-malonyl-C(60)-fullerene reduced the maximal velocity of product formation but did not alter the EC(50) value for activation by calmodulin. C(3)-tris-malonyl-C(60)-fullerene reduced the maximal velocity of citrulline formation by inducible NOS without altering the K(m) for l arginine substrate or the EC(50) value for tetrahydrobiopterin cofactor. As measured by sucrose density gradient centrifugation, fully inhibitory concentrations of C(3)-tris-malonyl-C(60)-fullerene did not produce a dissociation of nNOS dimers into monomers. These observations are consistent with the proposal that C(3)-tris-malonyl-C(60)-fullerene inhibits the inter-subunit transfer of electrons, presumably by a reversible distortion of the dimer interface. PMID- 10860540 TI - Peroxidase-catalyzed formation of quercetin quinone methide-glutathione adducts. AB - The oxidation of quercetin by horseradish peroxidase/H(2)O(2) was studied in the absence but especially also in the presence of glutathione (GSH). HPLC analysis of the reaction products formed in the absence of GSH revealed formation of at least 20 different products, a result in line with other studies reporting the peroxidase-mediated oxidation of flavonoids. In the presence of GSH, however, these products were no longer observed and formation of two major new products was detected. (1)H NMR identified these two products as 6-glutathionylquercetin and 8-glutathionylquercetin, representing glutathione adducts originating from glutathione conjugation at the A ring instead of at the B ring of quercetin. Glutathione addition at positions 6 and 8 of the A ring can best be explained by taking into consideration a further oxidation of the quercetin semiquinone, initially formed by the HRP-mediated one-electron oxidation, to give the o quinone, followed by the isomerization of the o-quinone to its p-quinone methide isomer. All together, the results of the present study provide evidence for a reaction chemistry of quercetin semiquinones with horseradish peroxidase/H(2)O(2) and GSH ultimately leading to adduct formation instead of to preferential GSH mediated chemical reduction to regenerate the parent flavonoid. PMID- 10860541 TI - The cyanide-resistant alternative oxidases from the fungi Pichia stipitis and Neurospora crassa are monomeric and lack regulatory features of the plant enzyme. AB - Both plant and fungal mitochondria have cyanide-resistant alternative oxidases that use reductant from the mitochondrial ubiquinone pool to reduce oxygen to water in a reaction that conserves no energy for ATP synthesis. The dimeric plant alternative oxidase is relatively inactive when its subunits are linked by a disulfide bond. When this bond is reduced, the enzyme can then be stimulated by its activators, alpha-keto acids. A Cys in the N-terminal section of the protein is responsible for both of these features. We examined the alternative oxidases in mitochondria isolated from two fungi Neurospora crassa and Pichia stipitis for dimeric structure, ability to form an intermolecular disulfide, and sensitivity to alpha-keto acids. Neither of the two fungal alternative oxidases could be covalently linked by diamide, which induces disulfide bond formation between nearby Cys residues, nor could they be cross-linked by a Lys-specific reagent or glutaraldehyde at concentrations which cross-link the plant alternative oxidase dimer completely. Alternative oxidase activity in fungal mitochondria was not stimulated by the alpha-keto acids pyruvate and glyoxylate. Pyruvate did stimulate activity when succinate was the respiratory substrate, but this was not a direct effect on the alternative oxidase. In contrast, added GMP was a strong activator of fungal alternative oxidase activity. Analysis of plant and fungal alternative oxidase protein sequences revealed a unique domain of about 40 amino acids surrounding the regulatory Cys in the plant sequences that is not present in the fungal sequences. This domain may be where dimerization of the plant enzymes occurs. In contrast to plant enzymes, the fungal alternative oxidases studied here are monomeric and their activities are independent of alpha-keto acids. PMID- 10860542 TI - fMLP-induced arachidonic acid release in db-cAMP-differentiated HL-60 cells is independent of phosphatidylinositol-4, 5-bisphosphate-specific phospholipase C activation and cytosolic phospholipase A(2) activation. AB - In inflammatory cells, agonist-stimulated arachidonic acid (AA) release is thought to be induced by activation of group IV Ca(2+)-dependent cytosolic phospholipase A(2) (cPLA(2)) through mitogen-activated protein kinase (MAP kinase)- and/or protein kinase C (PKC)-mediated phosphorylation and Ca(2+) dependent translocation of the enzyme to the membrane. Here we investigated the role of phospholipases in N-formylmethionyl-l-leucyl-l-phenylalanine (fMLP; 1 nM 10 microM)-induced AA release from neutrophil-like db-cAMP-differentiated HL-60 cells. U 73122 (1 microM), an inhibitor of phosphatidyl-inositol-4,5-biphosphate specific phospholipase C, or the membrane-permeant Ca(2+)-chelator 1, 2-bis?2 aminophenoxyethane-N,N,N',N'-tetraacetic acid (10 microM) abolished fMLP-mediated Ca(2+) signaling, but had no effect on fMLP-induced AA release. The protein kinase C-inhibitor Ro 318220 (5 microM) or the inhibitor of cPLA(2) arachidonyl trifluoromethyl ketone (AACOCF(3); 10-30 microM) did not inhibit fMLP-induced AA release. In contrast, AA release was stimulated by the Ca(2+) ionophore A23187 (10 microM) plus the PKC activator phorbol myristate acetate (PMA) (0.2 microM). This effect was inhibited by either Ro 318220 or AACOCF(3). Accordingly, a translocation of cPLA(2) from the cytosol to the membrane fraction was observed with A23187 + PMA, but not with fMLP. fMLP-mediated AA release therefore appeared to be independent of Ca(2+) signaling and PKC and MAP kinase activation. However, fMLP-mediated AA release was reduced by approximately 45% by Clostridium difficile toxin B (10 ng/ml) or by 1-butanol; both block phospholipase D (PLD) activity. The inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC), D609 (100 microM), decreased fMLP-mediated AA release by approximately 35%. The effect of D609 + 1-butanol on fMLP-induced AA release was additive and of a magnitude similar to that of propranolol (0.2 mM), an inhibitor of phosphatidic acid phosphohydrolase. This suggests that the bulk of AA generated by fMLP stimulation of db-cAMP-differentiated HL-60 cells is independent of the cPLA(2) pathway, but may originate from activation of PC-PLC and PLD. PMID- 10860543 TI - Mitochondrial adaptations to obesity-related oxidant stress. AB - It is not known why viable hepatocytes in fatty livers are vulnerable to necrosis, but associated mitochondrial alterations suggest that reactive oxygen species (ROS) production may be increased. Although the mechanisms for ROS mediated lethality are not well understood, increased mitochondrial ROS generation often precedes cell death, and hence, might promote hepatocyte necrosis. The aim of this study is to determine if liver mitochondria from obese mice with fatty hepatocytes actually produce increased ROS. Secondary objectives are to identify potential mechanisms for ROS increases and to evaluate whether ROS increase uncoupling protein (UCP)-2, a mitochondrial protein that promotes ATP depletion and necrosis. Compared to mitochondria from normal livers, fatty liver mitochondria have a 50% reduction in cytochrome c content and produce superoxide anion at a greater rate. They also contain 25% more GSH and demonstrate 70% greater manganese superoxide dismutase activity and a 35% reduction in glutathione peroxidase activity. Mitochondrial generation of H(2)O(2) is increased by 200% and the activities of enzymes that detoxify H(2)O(2) in other cellular compartments are abnormal. Cytosolic glutathione peroxidase and catalase activities are 42 and 153% of control values, respectively. These changes in the production and detoxification of mitochondrial ROS are associated with a 300% increase in the mitochondrial content of UCP-2, although the content of beta-1 ATP synthase, a constitutive mitochondrial membrane protein, is unaffected. Supporting the possibility that mitochondrial ROS induce UCP-2 in fatty hepatocytes, a mitochondrial redox cycling agent that increases mitochondrial ROS production upregulates UCP-2 mRNAs in primary cultures of normal rat hepatocytes by 300%. Thus, ROS production is increased in fatty liver mitochondria. This may result from chronic apoptotic stress and provoke adaptations, including increases in UCP-2, that potentiate necrosis. PMID- 10860544 TI - Metal-catalyzed oxidation of alpha-synuclein in the presence of Copper(II) and hydrogen peroxide. AB - alpha-Synuclein is a component of abnormal protein depositions of Lewy bodies and senile plaques found in Parkinson's and Alzheimer's diseases, respectively. By using chemical coupling reagents such as dicyclohexylcarbodiimide or N (ethoxycarbonyl)-2-ethoxy-1, 2-dihydroquinoline, the protein was shown to experience self-oligomerization in the presence of either copper(II) or Abeta25 35. The oligomers which appeared as a ladder on a 10-20% Tricine/SDS-PAGE have been suggested to participate in the formation of protein aggregations by possibly providing a nucleation center. Since oxidatively modified protein could increase its own tendency toward protein aggregation, metal-catalyzed oxidation of alpha-synuclein has been examined with copper(II) and hydrogen peroxide in the absence of the coupling reagent. Intriguingly, the protein was also self oligomerized into an SDS-resistant ladder on the gel. This biochemically specific copper-mediated oxidative oligomerization was shown to be dependent upon the acidic C-terminus of alpha-synuclein because the C-terminally truncated proteins such as alpha-syn114 and alpha-syn97 were not affected by the metal and hydrogen peroxide. More importantly, the oxidative oligomerization was synergistically enhanced by the presence of Abeta25-35, indicating that the peptide interaction with alpha-synuclein facilitated the copper(II) binding to the acidic C-terminus and subsequent oxidative crosslinking. It has been, therefore, suggested that abnormalities in copper and H(2)O(2) homeostasis and certain pathological factors functionally similar to the Abeta25-35 could play critical roles in the metal catalyzed oxidative oligomerization of alpha-synuclein, which may lead to possible protein aggregation and neurodegenerations. PMID- 10860545 TI - Characterization of two novel defense peptides from pea (Pisum sativum) seeds. AB - A fraction that possesses antifungal activity against Aspergillus niger has been isolated from seeds of the pea (Pisum sativum) by ammonium sulfate fractionation followed by gel filtration on Sephadex G-75. On further purification by reverse phase high performance liquid chromatography, two small cysteine-rich polypeptides were obtained (Psd1 and Psd2). They are localized primarily in vascular bundles and epidermis tissues of pea pods and exhibit high antifungal activity toward several fungi, displaying IC(50) values ranging from 0.04 to 22 microg/ml. This inhibitory activity decreases when A. niger growth medium is supplemented with cations such as Ca(2+), Mg(2+), Na(+), and K(+). Although the primary sequence of both Psd1 and Psd2 shows homology with other plant defensins, they cannot easily be assigned to any established group. PMID- 10860546 TI - Purification and characterization of mevalonate kinase from suspension-cultured cells of Catharanthus roseus (L.) G. Don. AB - Mevalonate kinase was purified to homogeneity from Catharanthus roseus (L.) G. Don suspension-cultured cells. The purified enzyme had an M(r) of 104,600 and a subunit size of about 41,500. Kinetic studies indicated an ordered sequential mechanism of action, in which mevalonate was the first substrate to bind and ADP was the last product to leave the enzyme. True values for the kinetic constants were determined for mevalonate, with K(ma) = 76 microM and K(ia) = 74 microM, and for ATP, with K(mb) = 0.13 mM and K(ib) = 0. 13 mM; the true V(max) was calculated to be 138.7 nkat/mg of protein. Product inhibition was only detectable at rather high concentrations: above 0.7 mM for 5-phosphomevalonate and above 2 mM for ADP, with an ADP/ATP ratio of at least 1. Mevalonate kinase activity was shown to be strongly inhibited by farnesyl diphosphate. Farnesyl diphosphate acted as a competitive inhibitor toward ATP, with a K(i) value of 0.1 microM. Mevalonate kinase activity was dependent on the presence of divalent ions. At a concentration of 2 mM, Mg(2+) and Mn(2+) were best and equally effective in sustaining activity; compared to Mg(2+) and Mn(2+), relative activities of 35, 30, 16, 4.8, and 3.4% were detected at equimolar concentrations of Zn(2+), Fe(2+), Co(2+), Ca(2+), and Ni(2+), respectively. The pH-dependent activity profile of mevalonate kinase showed a broad pH optimum between pH 7 and 10, with a maximum at about pH 8.9. PMID- 10860547 TI - Comparative induction of CYP1A1 expression by pyridine and its metabolites. AB - We compared pyridine and five of its metabolites in terms of (i) in vivo induction of CYP1A1 expression in the lung, kidney, and liver in the rat and (ii) in vitro binding to, and activation of, the aryl hydrocarbon receptor (AhR) in cytosol from rat liver or Hepa1c1c7 cells. Following a single 2.5 mmol/kg ip dose of either pyridine, 2-hydroxpyridine, 3-hydroxypyridine, 4-hydroxypyridine, N methylpyridinium, or pyridine N-oxide, CYP1A1 activity (ethoxyresorufin O deethylase), protein level (as determined by Western blotting), and mRNA level (as determined by Northern blotting) were induced by pyridine, N methylpyridinium, and pyridine N-oxide in the lung, kidney, and liver. The induction by N-methylpyridinium or pyridine N-oxide was comparable to or greater than that by pyridine in some tissues. 2-Hydroxypyridine and 3-hydroxypyridine caused tissue-specific induction or repression of CYP1A1, whereas 4 hydroxypyridine had no effect on the expression of the enzyme. Pyridine and its metabolites elicited weak activation of the aryl hydrocarbon receptor in a gel retardation assay in cytosol from rat liver but not Hepa 1c1c7 cells. However, the receptor activation did not parallel the in vivo CYP1A1 induction by the pyridine compounds, none of which inhibited binding of ?(3)H2,3,7, 8 tetrachlorodibenzo-p-dioxin to AhR in a competitive assay in rat liver cytosol. The findings are consistent with a role of pyridine metabolites in CYP1A1 induction by pyridine but do not clearly identify the role of aryl hydrocarbon receptor in the induction mechanism. PMID- 10860548 TI - Cell adhesive sequences in mouse laminin beta1 chain. AB - Laminin-1, a major component of the basement membrane, consists of three different chains, alpha1, beta1, and gamma1. We sought to identify cell adhesive sequences from the mouse laminin beta1 chain by testing HT-1080 fibrosarcoma and B16-F10 melanoma cells for binding to 187 overlapping synthetic peptides which covered the entire chain. Fourteen peptides showed cell adhesive activities with either peptide-conjugated Sepharose beads or peptide-coated plates or both. Additional cells, including neuronal, endothelial, and salivary gland cells, showed biological responses in a cell type-specific manner. B-7, B-133, and B-160 showed the most potent cell attachment. Cell binding on three peptides (B-34, B 133, and B-160) was inhibited by EDTA. Cell adhesion to 11 of the 12 active peptides was inhibited to varying degrees by heparin. Of the 17 active peptides identified in the laminin beta1 chain in this and other studies, 8 are clustered on the amino terminal globular domain, suggesting a possible important role in cell binding for this domain that may be multifunctional. These data demonstrate that the laminin beta1 chain has multiple active sites for cell adhesion, some of which are cell-type specific. PMID- 10860549 TI - Cress and potato soluble epoxide hydrolases: purification, biochemical characterization, and comparison to mammalian enzymes. AB - Affinity chromatographic methods were developed for the one-step purification to homogeneity of recombinant soluble epoxide hydrolases (sEHs) from cress and potato. The enzymes are monomeric, with masses of 36 and 39 kDa and pI values of 4.5 and 5.0, respectively. In spite of a large difference in sequence, the two plant enzymes have properties of inhibition and substrate selectivity which differ only slightly from mammalian sEHs. Whereas mammalian sEHs are highly selective for trans- versus cis-substituted stilbene oxide and 1,3 diphenylpropene oxide (DPPO), plant sEHs exhibit far greater selectivity for trans- versus cis-stilbene oxide, but little to no selectivity for DPPO isomers. The isolation of a covalently linked plant sEH-substrate complex indicated that the plant and mammalian sEHs have a similar mechanism of action. We hypothesize an in vivo role for plant sEH in cutin biosynthesis, based on relatively high plant sEH activity on epoxystearate to form a cutin precursor, 9,10 dihydroxystearate. Plant sEHs display a high thermal stability relative to mammalian sEHs. This stability and their high enantioselectivity for a single substrate suggest that their potential as biocatalysts for the preparation of enantiopure epoxides should be evaluated. PMID- 10860550 TI - Human fatty acid omega-hydroxylase, CYP4A11: determination of complete genomic sequence and characterization of purified recombinant protein. AB - The gene of the human fatty acid omega-hydroxylase, CYP4A11, has been isolated from a human BAC library, and its complete genomic sequence has been determined. The CYP4A11 gene spanned 12,568 bp and contained 12 exons. The known PPAR recognition elements (PPRE), which were reported to be involved in the induction of CYP4A6 by clofibric acid, were not observed within the 5'-flanking region of the CYP4A11 gene. The recombinant CYP4A11 protein expressed in Escherichia coli using the pCWOri expression vector was purified to an almost electrophoretically homogeneous state with a specific content of 6.4 nmol of P450/mg of protein. This P450 exhibited omega-hydroxylation activity toward laurate, with a turnover number of 14.7 nmol/min/nmol of P450. The apparent K(m) and V(max) values were 56.7 microM and 15.2 nmol/min/nmol of P450, respectively. It also showed omega hydroxylation activity toward palmitate, with a turnover number of 0.78 nmol/min/nmol of P450. Although several reports from other groups described that CYP4A11 preparations catalyzed omega-hydroxylation of arachidonic acid, our purified recombinant protein exhibited no activity toward arachidonic acid nor prostaglandin A(1). PMID- 10860551 TI - Allantoate amidinohydrolase (Allantoicase) from Chlamydomonas reinhardtii: its purification and catalytic and molecular characterization. AB - An allantoate-degrading enzyme has been purified to electrophoretic homogeneity for the first time from a photosynthetic organism, the unicellular green algae Chlamydomonas reinhardtii. The purification procedure included a differential protein extraction followed by conventional steps such as ammonium sulfate fractionation, gel filtration, anion exchange chromatography, and preparative electrophoresis. Under the routine assay conditions (7 mM allantoate), specific activity for the purified enzyme was 185 U/mg, which rose to 225 U/mg under kinetic considerations (saturating substrate). Therefore, a turnover number of 4.5 x 10(4) min(-1) can be deduced for the 200-kDa protein. The enzyme is a true allantoicase (EC 3.5.3.4) that catalyzes the degradation of allantoate to ( )ureidoglycolate and (+)ureidoglycolate to glyoxylate. The enzyme exhibited hyperbolic kinetic for allantoate and ureidoglycolate with K(m) values of 2 and 0.7 mM, respectively. V(max) of the reaction with allantoate as substrate was nine times higher than that with ureidoglycolate. The native enzyme has a molecular weight of 200 kDa and consists of six identical or similar-sized subunits of 34 kDa each, organized in two trimers of 100 kDa. Each subunit has five cysteine residues, four of which are involved in disulfide bonds, with a total of 12 disulfide bonds in the 200-kDa protein. Allantoate inhibits competitively the reaction with ureidoglycolate as substrate. In addition, buffers and group-specific reagents affect the activity in the same manner irrespective of the substrate used. Those results suggest that both substrates use the same active site. The effect of group-specific reagents suggest that the amino acids histidine, tyrosine, and cysteine are essentials for the allantoicase activity with both substrates. PMID- 10860552 TI - Localization of C-terminal domains required for the maximal activity or for determination of substrate preference of maize branching enzymes. AB - Previous analysis of a chimeric enzyme mBEII-IBspHI, in which the C-terminal 229 amino acids of maize endosperm branching enzyme isoform II (mBEII) are replaced by the corresponding 284 amino acids of isoform I (mBEI), suggested that the carboxyl terminus of maize branching enzymes may be involved in catalytic efficiency and substrate preference. In the present study, additional hybrids of mBEI and mBEII were generated and expressed in Escherichia coli BL21 (DE3) to dissect the structure/function relationships of the C-terminal regions of maize branching enzymes. A truncated form of purified mBEII-IBspHI, which lacks the C terminal 58 amino acids, retained similar levels of V(max) in branching activity, K(m) for reduced amylose AS 320, and substrate preference for amylose than amylopectin when compared to mBEII-IBspHI. This indicates that the C-terminal extension derived from mBEI is not required for either catalysis or substrate preference. However, deletion of an additional 87 amino acids from the carboxyl terminus resulted in complete loss of activity. Replacement of the deleted C terminal additional 87 amino acids with the corresponding 79 amino acids from mBEII restored 25% of the mBEII-IBspHI branching activity without altering substrate preference. It thus appears that a C-terminal region encompassing Leu649-Asp735 of mBEII-IBspHI is required for maximum catalytic efficiency. Another C-terminal region, residues Gln510-Asp648, of mBEII-IBspHI (Gln476-Asp614 of mBEI) may be involved in substrate-preference determination. PMID- 10860553 TI - Inhibition selectivity of grapefruit juice components on human cytochromes P450. AB - Five compounds including furanocoumarin monomers (bergamottin, 6', 7' dihydroxybergamottin (DHB)), furanocoumarin dimers (4-??6-hydroxy-71-?(1-hydroxy 1-methyl)ethyl-4-methyl-6-(7-oxo-7H- furo?3,2-g1benzopyran-4-yl)-4-hexenyl]oxy] 3,7-dimethyl- 2-octenyl]oxy]-7H-furo[3,2-g]?1benzopyran-7-one (GF-I-1) and 4-??6 hydroxy-7??4-methyl-1-(1-methylethenyl)-6-(7-oxo-7H-furo?3, 2-g1benzopyran-4-yl) 4-hexenyloxy-3, 7-dimethyl-2-octenyloxy-7H-furo?3,2-g1benzopyran-7-one (GF-I-4)), and a sesquiterpene nootkatone have been isolated from grapefruit juice and screened for their inhibitory effects toward human cytochrome P450 (P450) forms using selective substrate probes. Addition of ethyl acetate extract of grapefruit juice into an incubation mixture resulted in decreased activities of CYP3A4, CYP1A2, CYP2C9, and CYP2D6. All four furanocoumarins clearly inhibited CYP3A4 catalyzed nifedipine oxidation in concentration- and time-dependent manners, suggesting that these compounds are mechanism-based inhibitors of CYP3A4. Of the furanocoumarins investigated, furanocoumarin dimers, GF-I-1 and GF-I-4, were the most potent inhibitors of CYP3A4. Inhibitor concentration required for half maximal rate of inactivation (K(I)) values for bergamottin, DHB, GF-I-1, and GF-I 4 were calculated, respectively, as 40.00, 5. 56, 0.31, and 0.13 microM, whereas similar values were observed on their inactivation rate constant at infinite concentration of inhibitor (k(inact), 0.05-0.08 min(-1)). Apparent selectivity toward CYP3A4 does occur with the furanocoumarin dimers. In contrast, bergamottin showed rather stronger inhibitory effect on CYP1A2, CYP2C9, CYP2C19, and CYP2D6 than on CYP3A4. DHB inhibited CYP3A4 and CYP1A2 activities at nearly equivalent potencies. Among P450 forms investigated, CYP2E1 was the least sensitive to the inhibitory effect of furanocoumarin components. A sesquiterpene nootkatone has no significant effect on P450 activities investigated except for CYP2A6 and CYP2C19 (K(i) = 0.8 and 0.5 microM, respectively). PMID- 10860554 TI - Promoter activity and regulation of the CYP4F2 leukotriene B(4) omega-hydroxylase gene by peroxisomal proliferators and retinoic acid in HepG2 cells. AB - The human liver CYP4F2 gene (Accession No. AF221943) encodes a leukotriene B(4) omega-hydroxylase that metabolizes leukotriene B(4) (LTB(4)) to a less potent proinflammatory eicosanoid, 20-OH-LTB(4). We sequenced a 6.7-kb genomic fragment of the human CYP4F2 gene that has the first five exons and 500 bp of the 5' flanking region. The major transcription start site was found to be 49 bp upstream of the 3' end of exon 1 and the ATG translation initiation codon was located in exon 2. Besides the TATA box at -39 bp and basal transcription factor binding sites, the promoter region and 412-bp intron 1 have several putative binding sites for nuclear factors that may mediate the inflammatory response and lipid homeostasis. We found two DR1 elements in the 5' promoter, a DR2 element in intron 1, and RXR/RAR binding sites in both intron 1 and the 5' promoter. DNase I footprinting revealed three protected sequences, with the region containing two CAATT boxes at -71 and -111 bp important in CYP4F2 gene expression. Luciferase reporter assays showed that the 500-bp upstream sequence has strong promoter activity. Transient transfection experiments identified two sites in the 5' promoter and intron 1 that cooperate in gene transcription while exon 1 and a GC rich region flanking exon 1 inhibit transcription. trans-Retinoic acid and 9-cis retinoic acid stimulate promoter activity 3- and 6-fold, respectively, while cotransfection with RXRalpha or RAR/RXRalpha further enhanced activity. Peroxisome proliferators inhibit CYP4F2 gene promoter activity and cotransfection with PPARalpha or PPARalpha/RXRalpha can slightly attenuate this inhibition. Both saturated fatty acids and 12-hydroxydodecanoic acid (12-OH-C(12)) can stimulate CYP4F2 gene promoter activity. Therefore, the CYP4F2 gene is repressed by peroxisomal proliferators and induced by retinoic acid, with RAR/RXRalpha mediating the induction while PPARalpha/RXR functions neither in the repression nor in the induction by peroxisomal proliferators or retinoic acid. PMID- 10860555 TI - Alternative splice variant of gamma-calmodulin-dependent protein kinase II alters activation by calmodulin. AB - Calmodulin-dependent protein kinase II (CaMKII) is a ubiquitous, multifunctional enzyme family involved in the regulation of a variety of Ca(2+)-signaling pathways. These family members are expressed from four highly homologous genes (alpha, beta, gamma, and delta) with similar catalytic properties. Additional isoforms of each gene, created by alternative splicing of variable regions I-XI, are differentially expressed in various cell types. gammaB, gammaC, gammaD, gammaE, gammaF, gammaGs, and gammaH CaMKII isoforms are expressed in the biliary epithelium; however, little is known about their roles in these cells. We began our studies into the function of these variable regions by examining the effects of variable region I on kinase activation and calmodulin binding. Activities and calmodulin binding properties of gammaB and gammaGs, which differ only by the exclusion or inclusion of this region, were compared. The K(0.5) for calmodulin was 2.5-fold lower for gammaGs than gammaB. In contrast, gammaB bound calmodulin more tightly in a calmodulin overlay assay. Mutation of variable regions I's charged residue, gammaGs-R318E, resulted in an enzyme with intermediate activation properties but a calmodulin affinity similar to gammaB. Thus, variable region I appears to modulate calmodulin sensitivity, in part, through charge charge interactions. This altered threshold of activation may modulate cellular responses to gradients of Ca(2+)/calmodulin in the biliary tract. PMID- 10860556 TI - A 66-base-pair enhancer module activates the expression of a distinct isoform of UDP-glucuronosyltransferase family 1 (UGT1A2) in primary hepatocytes. AB - UGT1A2, an isoform of the UDP-glucuronosyltransferase family 1 (UGT1), is not expressed in the rat liver, but its expression was highly induced in primary cultures of rat hepatocytes. In primary hepatocytes that had been cultured for 70 h, the amount of UGT1A2 mRNA was 100 times higher than that in the rat liver. Deletion analysis of a 4.8-kb promoter region of the UGT1A2 gene revealed that a 66-nucleotide region between -307 and -242 upstream of the transcription start site was required for induction of UGT1A2 expression. The 66-nucleotide region acted on a heterologous promoter in a manner independent of its position and orientation in reporter constructs. Gel mobility shift assay showed that a specific binding protein to this region appeared in the nuclei of cultured hepatocytes, but was not present in the rat liver. DNase I protection analysis revealed the existence of a CTGGCAC core sequence between -274 and -268 of the UGT1A2 promoter. Methylation interference assay showed that the guanine residues at -294 and -287 on the upper strand and the guanine residue at -267 on the lower strand as well as the core sequence were required for the DNA-protein interaction. These results suggest that the 66-nucleotide region, which was designated culture-associated expression responsive enhancer module (CEREM), interacts with a specific nuclear protein and enhances the expression of UGT1A2 in cultured hepatocytes. PMID- 10860557 TI - Insect melanogenesis. III. Metabolon formation in the melanogenic pathway regulation of phenoloxidase activityy by endogenous dopachrome isomerase (decarboxylating) from Manduca sexta. AB - Tyrosinase initiates melanogenesis in a variety of organisms. The nature of melanin formed is modified subsequently by dopachrome isomerase and other melanogenic proteins. Earlier, we reported the partial purification of dopachrome isomerase (decarboxylating) from the hemolymph of Manduca sexta and demonstrated the generation of a new quinone methide intermediate during melanogenesis (Sugumaran, M., and Semensi, V. (1991) J. Biol. Chem. 266, 6073-6078). In this paper, we report the purification of this enzyme to homogeneity and a novel inhibition mechanism for regulation of phenoloxidase activity. The activity of phenoloxidase isolated from M. sexta was markedly inhibited by purified dopachrome isomerase. In turn, phenoloxidase also reciprocated by inhibiting the isomerase activity. Preformed dopaminechrome did not serve as the substrate for the isomerase; but dopaminechrome that generated in situ by phenoloxidase was readily converted to melanin pigment by the phenoloxidase/isomerase mixture. Furthermore, the isomerase, which has a molecular weight of about 40,000 in native state, exhibited retardation during affinity electrophoresis on sodium dodeyl sulfate (SDS)-polyacrylamide gel electrophoresis gel copolymerized with tyrosinase and migrated with a molecular weight of 50,000, indicating complex formation with phenoloxidase. Electrophoresis of pupal cuticular extract on polyacrylamide gel, followed by activity staining revealed the presence of a protein band carrying both phenoloxidase and isomerase activity. Accordingly, a high-molecular-weight melanogenic complex was isolated from the pharate cuticle of M. sexta. The complex catalyzed the generation of melanochrome from dopa, while the free phenoloxidase produced only dopachrome from the same substrate. When the complex was treated with trace amounts of SDS, which inhibited the activity of dopachrome isomerase present in the complex, then only the conversion of dopa to dopachrome was observed. These studies confirm the formation of a melanogenic complex between phenoloxidase and dopachrome isomerase. By forming a complex and regulating each other's activity, these two enzymes seem to control the levels of endogenous quinones. PMID- 10860558 TI - Oxidation chemistry of 1,2-dehydro-N-acetyldopamines: direct evidence for the formation of 1,2-dehydro-N-acetyldopamine quinone. AB - Two-electron oxidation of catecholamines either by phenol oxidase or by chemical oxidants such as sodium periodate produces their corresponding o-quinones as observable products. But, in the case of 1,2-dehydro-N-acetyldopamine, an important insect cuticular sclerotizing precursor, phenol oxidase catalyzed oxidation has been reported to generate a quinone methide analog as a transient, but first observable product. ?Sugumaran, M., Semensi, V., Kalyanaraman, B., Bruce, J. M., and Land, E. J. (1992) J. Biol. Chem. 267, 10355-10361. The corresponding quinone has escaped detection until now. However, in this paper, for the first time, we present direct evidence for the formation of dehydro-N acetyldopamine quinone and show that it can readily be produced from the tautomeric quinone methide imine amide during the chemical oxidation of dehydro-N acetyldopamine under acidic conditions. This situation is in sharp contrast to other known alkyl-substituted catechol oxidations, where quinone is the first observable product and quinone methide is the subsequently generated product. Dehydro-N-acetyldopamine quinone thus formed is also highly unstable. Semiempirical molecular orbital calculation also indicates that quinone methide imine amide is more stable than the quinone. Chemical considerations indicate that the quinone methide tautomer, and not the dehydro-N-acetyldopamine quinone, is responsible for crosslinking the structural proteins and chitin polymer in the insect cuticle. Therefore, the quinone methide tautomer, and not the quinone, is the key reactive intermediate aiding the hardening of insect cuticle. PMID- 10860559 TI - The methionine chain elongation pathway in the biosynthesis of glucosinolates in Eruca sativa (Brassicaceae). AB - Glucosinolates are nitrogen- and sulfur-containing plant natural products that have become increasingly important in human affairs as flavor precursors, cancer prevention agents, and crop protectants. While many glucosinolates are biosynthesized from common amino acids, the major glucosinolates in economically important species of the Brassicaceae, such as Brassica napus (oilseed rape), are thought to be formed from chain-elongated derivatives of methionine or phenylalanine. We investigated the chain elongation pathway for methionine that is involved in glucosinolate biosynthesis in Eruca sativa. Isotopically labeled methionine and acetate were administered to cut leaves and the major product, 4 methylthiobutylglucosinolate (isolated as its desulfated derivative), was analyzed by MS and NMR. Administration of ?U-(13)Cmethionine showed that the entire carbon skeleton of this amino acid, with the exception of the COOH carbon, is incorporated as a unit into 4MTB. Administration of ?(13)C- and ?(14)Cacetate gave a labeling pattern consistent with the operation of a three-step chain elongation cycle which begins with the condensation of acetyl-CoA with a 2-oxo acid derived from methionine and ends with an oxidative decarboxylation forming a new 2-oxo acid with one additional methylene group. Administration of ?(15)Nmethionine provided evidence for the transfer of an amino group to the chain-elongated 2-oxo acid, forming an extended amino acid which serves as a substrate for the remaining steps of glucosinolate biosynthesis. The retention of a high level of (15)N in the products suggests that the amino transfer reactions and the chain elongation cycle are localized in the same subcellular compartment. PMID- 10860560 TI - Effects of morphological complexity on phonological output deficits in fluent and nonfluent aphasia. AB - This study examined the effects of morphological complexity on aphasic speakers with lexical-phonological output deficits. Subjects were two fluent and two nonfluent aphasic speakers who repeated morphologically simple words at the same level of accuracy and whose errors were virtually all phonological in nature. They were asked to repeat a variety of morphologically complex (i.e., affixed) words. Results were interpreted within our two-stage model of lexical phonological production (Kohn & Smith, 1994, 1995), which we expand to include a distinction between derivational and inflectional morphology. When comparing overall performance levels between morphologically simple and complex words, only three subjects exhibited more difficulty repeating the morphologically complex targets. Nevertheless, when comparing repetition accuracy between different types of morphologically complex words (e.g., derived vs. inflected), all four subjects displayed similar patterns. These findings suggested that while morphological complexity has different effects on the two stages within the lexical phonological output system, the relative effects of different morphological structures are constant. At the level of error analysis, patterns of affix errors distinguished the nonfluent from the fluent subjects in ways that are reminiscent of the affix errors associated with agrammatic and paragrammatic speech. This finding raised questions concerning the relationship between morphosyntactic and morphophonological deficits. PMID- 10860561 TI - Verb retrieval in brain-damaged subjects: 1. Analysis of stimulus, lexical, and conceptual factors. AB - Verb retrieval for action naming was assessed in 53 brain-damaged subjects by administering a standardized test with 100 items. The goal of the study was to gain further insight into the nature of verb processing impairments by investigating the influence of several kinds of stimulus, lexical, and conceptual factors on the subjects' performance at the level of group tendencies and also at the level of individual differences. (1) Stimulus factors: visual complexity, familiarity, image agreement, and one vs. two pictures (which corresponds to ongoing vs. completed actions); (2) lexical factors: name agreement, verb frequency, and whether the root of the target verb has a homophonous noun; (3) conceptual factors: whether the action is done with the hand or the body, whether the action involves one or two core participants, whether the undergoer of the action has a change of internal state, whether the undergoer has a change of spatial location, and whether the actor makes use of an instrument in carrying out the action. The subjects were divided into an impaired group (n = 19) and an unimpaired group (n = 34) on the basis of their overall performance on the test. For both groups of subjects, verb retrieval was significantly affected by the following factors: familiarity, image agreement, name agreement, homophonous noun, and undergoer change of location. These results indicate that, at the level of group analysis, some factors have a stronger influence on verb retrieval for action naming than others. Moreover, the finding that the two groups exhibited the same general pattern of factor sensitivity suggests that although the processing efficiency of the mechanisms that subserve verb retrieval is degraded in the impaired group, the basic functional properties of these mechanisms may not be qualitatively very different from those of the unimpaired group. Further analyses were conducted at the level of individual subjects and revealed a considerable amount of variation with regard to factor sensitivity. Many patterns of associations and dissociations of factors were found across the subjects, which suggests that the task of retrieving verbs for naming actions is quite complex and that different subjects can be influenced by different properties of both the stimuli and the target verbs. PMID- 10860562 TI - Verb retrieval in brain-damaged subjects: 2. Analysis of errors. AB - Verb retrieval for action naming was assessed in 53 brain-damaged subjects by administering a standardized test with 100 items. In a companion paper (Kemmerer & Tranel, 2000), it was shown that impaired and unimpaired subjects did not differ as groups in their sensitivity to a variety of stimulus, lexical, and conceptual factors relevant to the test. For this reason, the main goal of the present study was to determine whether the two groups of subjects manifested theoretically interesting differences in the kinds of errors that they made. All of the subjects' errors were classified according to an error coding system that contains 27 distinct types of errors belonging to five broad categories-verbs, phrases, nouns, adpositional words, and "other" responses. Errors involving the production of verbs that are semantically related to the target were especially prevalent for the unimpaired group, which is similar to the performance of normal control subjects. By contrast, the impaired group had a significantly smaller proportion of errors in the verb category and a significantly larger proportion of errors in each of the nonverb categories. This relationship between error rate and error type is consistent with previous research on both object and action naming errors, and it suggests that subjects with only mild damage to putative lexical systems retain an appreciation of most of the semantic, phonological, and grammatical category features of words, whereas subjects with more severe damage retain a much smaller set of features. At the level of individual subjects, a wide range of "predominant error types" were found, especially among the impaired subjects, which may reflect either different action naming strategies or perhaps different patterns of preservation and impairment of various lexical components. Overall, this study provides a novel addition to the existing literature on the analysis of naming errors made by brain-damaged subjects. Not only does the study advance our knowledge of the relatively under investigated topic of action naming errors, but it also approaches the analysis from the point of view of a detailed, theoretically motivated, and reliable error coding system. PMID- 10860563 TI - In search of the language switch: An fMRI study of picture naming in Spanish English bilinguals. AB - For many years, researchers investigating the brain bases of bilingualism have concentrated on two basic questions. The first concerns the nature of language representation. That is, are a bilinguals' two languages represented in distinct or overlapping areas of the brain. The second basic question in the neuropsychology of bilingualism concerns the neural correlates of language switching, that is, the areas that are active when bilinguals switch from one language to the other. Performance between single-language and dual-language picture naming was compared in a group of six Spanish-English bilinguals using behavioral measures and functional magnetic resonance imaging. Participants showed slower reaction times and increased activation in the dorsolateral prefrontal cortex in the mixed language condition relative to single language condition. There was no evidence that each language was represented in different areas of the brain. Results are consistent with the view that language switching is a part of a general executive attentional system and that languages are represented in overlapping areas of the brain in early bilinguals. PMID- 10860564 TI - Mirror writing and handedness. AB - It has been reported that left-handed subjects are better able to write in mirror reversed script than right-handers (Tankle & Heilman, 1983). Vaid and Stiles Davis (1989) conducted studies which led them to contradict the supposed superiority of left-handers in this area. In these studies, left as well as right handed subjects were examined under normal- and mirror-writing conditions. Both examinations included the analysis of writing time and the accuracy of mirror writing (error rates). Using a digitizing tablet, we examined normal- and mirror writing performance of left-handers, right-handers, and left-handed subjects who habitually write with their right hand. Our results support the finding of Tankle and Heilman (1983) that left-handers perform better in mirror-writing tasks. PMID- 10860565 TI - Perception of dynamic acoustic patterns by an individual with unilateral verbal auditory agnosia. AB - Previous studies have found that subjects diagnosed with verbal auditory agnosia (VAA) from bilateral brain lesions may experience difficulties at the prephonemic level of acoustic processing. In this case study, we administered a series of speech and nonspeech discrimination tests to an individual with unilateral VAA as a result of left-temporal-lobe damage. The results indicated that the subject's ability to perceive steady-state acoustic stimuli was relatively intact but his ability to perceive dynamic stimuli was drastically reduced. We conclude that this particular aspect of acoustic processing may be a major contributing factor that disables speech perception in subjects with unilateral VAA. PMID- 10860566 TI - Right hemisphere semantic processing of visual words in an aphasic patient: an fMRI study. AB - This study was designed to identify the neural network supporting the semantic processing of visual words in a patient with large-scale damage to left hemisphere (LH) language structures. Patient GP, and a control subject, RT, performed semantic and orthographic tasks while brain-activation patterns were recorded using functional magnetic resonance imaging. In RT, the semantic orthographic comparison activated LH perisylvian and extrasylvian temporal regions comparable to the network of areas activated by non-brain-damaged subjects in other neuroimaging studies of semantic discrimination. In GP, the same comparison activated homologous right-hemisphere regions, demonstrating the ability of the right hemisphere to subserve visual lexicosemantic processes. The results are discussed within the context of the normal right hemisphere's capacity for semantic processing of visual words. Examining results from functional neuroimaging studies on recovery in the context of innate hemispheric abilities may enable reconciliation of disparate claims about mechanisms supporting recovery from aphasia. PMID- 10860567 TI - Packing of myosin molecules in muscle thick filaments. AB - The backbone of the myosin filament is an aggregate of alpha-helical coiled coil myosin rods. Its surface forms a three-stranded helix composed of myosin heads. Currently there is no adequate model to describe the organization of the myosin filament. It is proposed here that, in cross-section the light meromyosin (LMM) of 18 myosin molecules form an outer tube, with nine S2 forming the interior core. At the surface of the thick filament, myosin heads are arranged in three rows, giving the filament a periodicity of 14.3 nm per three myosin molecules. Two of these molecules are organized at an angle of 120 degrees to each other on the same level, while the third is shifted 7.2 nm along the filament axis. This packing gives a striation pattern of 7.2 nm by electron microscopy. An alternative model is also possible, in which the heads of the myosin molecules are uniformly spaced at an interval of 14.3 nm along the filament axis. The packing of individual molecules within the myosin filament is based on a regular pattern of charge on the 28 amino-acid repeat in the rod domain. PMID- 10860568 TI - Speeding up kinesin-driven microtubule gliding in vitro by variation of cofactor composition and physicochemical parameters. AB - So far, there has been a discrepancy between the velocities of kinesin-dependent microtubule motility measured in vitro and within cells. By changing ATP, Mg(2+), and kinesin concentrations, pH and ionic strength, we tried to find conditions that favour microtubule gliding across kinesin-covered glass surfaces. For porcine brain kinesin, we found that raising the molar Mg(2+)/ATP ratio can substantially elevate gliding velocity. Gliding became also faster after temperature elevation or lowering the number of kinesin molecules bound to the glass surface. The highest mean gliding velocity (1.8 microm/s+/-0.09 microm/s), approaching velocities measured for anterograde transport in vivo, was achieved by combination of favourable factors (2.5 m m ATP, 12.5 m m Mg(2+), 37 degrees C, 450 kinesin molecules/microm(2)). PMID- 10860569 TI - From hormone signal, via the cytoskeleton, to cell growth in single cells of tobacco. AB - Cultured mesophyll protoplasts of Nicotiana tabacum L. can be hormonally induced into different developmental pathways. In a medium containing auxins (NAA) and cytokinins (BAP) cells divide and eventually give rise to calli. When only auxins are present cells elongate and finally differentiate into very long tubular cells. We focused on the sequence of events leading to elongation. When cultured in a high (1 mg/l) auxin concentration elongating cells seem to pass a certain threshold and increase their nuclear DNA up to about 16C. Cells cultured in a low (0.065 mg/l) auxin concentration only have C-values up to 4C, are unable to pass this threshold and finally fail to elongate. Besides the concentration dependence of the auxin signal, the efflux of auxin seems to be necessary for elongation since addition of TIBA drastically reduces the amount of elongating cells. Concomitant with the changes in nuclear physiology, auxin-induced axiality is seen as sequential rearrangements of microtubules and actin-filaments and of cell wall cellulose microfibrils from 'randomly' arranged in spherical cells to an orientation perpendicular to the long axis of elongating cells. PMID- 10860570 TI - Interaction of plant polysomes with the actin cytoskeleton. AB - Protein composition and functional activity of various polysome subpopulations isolated from Vicia faba L. leaves and Triticum aestivum L. and Hordeum vulgare L. seedlings were studied. Membrane- and cytoskeleton-bound polysomes were more active in the wheat germ cell-free translational system than free polysomes. Several non-ribosomal proteins were detected in the polysome preparations by gel electrophoresis and Western blot analysis: (1) a canonical actin of mol wt 42 kDa; (2) a 40 kDa protein, demonstrating affinity for ribosomes, sharing some determinants with actin, and present predominantly in the subpopulations of bound polysomes; and (3) an acidic ribosome-associated p40 evenly distributed between free and bound polysomes. The possibility of involvement of these proteins in interactions between polysomes and the actin cytoskeleton is discussed. PMID- 10860571 TI - Change in the activity of lectins in cell structure of winter wheat crown under the action of low-temperature hardening and fusicoccin. AB - Haemagglutinating activity was determined in cell walls and total cell organelles of crown cells of Winter wheat (Triticum aestivum L. ) plants. The effect of fusicoccin (FC) was investigated using fractions obtained from plants hardened for 7 days at 2 degrees C and from untreated plants. FC concentration (5x10(-7) m) increased the frost resistance of the plants. The temporal pattern of lectin activity during hardening could be described by a single-peak curve. In the cell wall fraction, the highest activity manifested itself after one-day hardening, and in the fraction of organelles it peaked after five-days hardening. The carbohydrate specificity of lectins also changed during hardening; cell wall lectins completely lost their capacity for interaction with uridine diphosphoglucose, glucose 6-phosphate, D-galactosamine, and N-acetylglucosamine and the lectins of organelles retained some affinity only for amino sugars. After hardening the test plants, the activity of the lectins increased substantially in the cell walls and plastids, decreased in the nuclei, and was practically flat in mitochondria and microsomes. Consequently, low temperature and FC with their antistress effect improved frost resistance and stimulated the activity of the lectins of some cell structures of the tillering node of winter wheat. A similar action of low temperature and FC in increasing the activity of lectins of plastids was found. Further information was obtained on the subcellular localization of lectins providing additional information on their possible participation in the development of frost resistance of winter wheat. PMID- 10860572 TI - Effect of abscisic acid and cold acclimation on the cytoskeletal and phosphorylated proteins in different cultivars of Triticum aestivum L. AB - In winter wheat, the tubulin and 60 kDa-phosphorylated proteins/actin ratio is considerably higher in the roots than in the leaves. Differences in the content of the main cytoskeletal proteins were also found in the leaves of the different cultivars. It is suggested that the lower amount of the tubulin and 60 kDa phosphorylated proteins and higher content of actin determine the greater tubulin cytoskeletal stability in the leaves and their higher frost resistance, as compared with the roots. Also, it is possible that the higher content of the tubulin and 60 kDa-phosphorylated proteins defines the lower microtubule (MT) stability in the leaves of the low frost resistant cultivar than in the leaves of the more frost resistant ones. In the roots and leaves of the low frost resistant cultivar, the low stability of the numerous tubulin structures is apparently one reason for the abscisic acid (ABA)-induced reduction of the cytoskeletal and 60 kDa-phosphorylated proteins in the cells. The cold acclimation compensated the ABA effect in the roots of the very frost resistant cultivar in the most extent. This suggests the existence of the different pathways in the increased plant cell frost resistance through the action of ABA and low temperature. PMID- 10860573 TI - Cytoskeleton-induced alterations of the lectin activity in winter wheat under cold hardening and abscisic acid (ABA). AB - The roots and leaves of 7-day seedlings of three winter wheat cultivars differing in frost resistant were used to study changes in lectin activity under cytoskeleton modifiers (DMSO-7%; colchicine-1 m m; oryzalin-15 microm; cytochalasin B-15 microm) of non-hardened (23 degrees C) and hardened (2-3 degrees C, 3-7 day) plants. Plants were grown with ABA (30 microm) or without ABA. Pretreatment with colchicine, oryzalin [inhibitors of microtubules (MT) polymerization], cytochalasin B [inhibitor of microfilament (MF) polymerization] increased the activity of cell wall lectins, although pretreatment with DMSO (stabilizer of microtubules) decreased the activity. Both hardening and ABA decreased the effect of the cytoskeletal modifiers. These results could be explained by the appearance of tolerant MTs with less affinity. It is probable that increase in the activity of cell wall lectins may be the compensatory mechanism which stabilizes the cytoskeleton structure in conditions tending to disrupt it. The genotype with low resistance had higher sensitivity of lectin activity to cytoskeleton modifiers than the frost resistant genotype. The results suggest that leaves have more stable MTs and MFs and stronger MT-MF binding than roots. PMID- 10860574 TI - Influence of inhibitors of cytoskeleton proteins on water exchange of wheat roots under the after-action of water stress. AB - The dynamics of water molecular state and transport in winter wheat (Triticum aestivum L.) of roots different resistance cultivars was studied by a biophysical method, Nuclear Magnetic Resonance (NMR), and a physiological method, Water Holding Capacity (WHC). The effective coefficient of water self-diffusion (D(eff)), spin-spin relaxation times (T(2)) and WHC were measured after structural modification of cytoskeleton by colchicine and cytochalasin B after the action of water stress. New information about molecular mechanisms of water state and water transport regulation determined by the influence of dynamic cytoskeleton structure has been obtained. This is very important for the development of a fundamental theory of water exchange in plants, and for the ways of its optimization under conditions of environmental stress. PMID- 10860575 TI - Influence of cytoskeletal agents on the respiratory electron transport pathways in the cells of winter wheat leaves. AB - The effects of actin and tubulin polymerization inhibitors on the respiratory electron transport pathway activities were investigated using abscisic acid (ABA) and cold-treated winter wheat seedling leaves. In unstressed control plants, cytochalasin B (15 microm) decreased the capacity of the cytochrome pathway, but stimulated the cyanide-resistant pathway, whereas oryzalin (15 microm) produced the opposite effects. Cold hardening (3 degrees C for 7 days) and ABA treatment 30 microm changed the respiratory pattern in a similar manner to cytochalasin B but to lesser effects. This points to cold- and ABA-induced reduction in microfilament sensitivity to these drugs and hence stabilization of actin dependent processes. In contrast, oryzalin had only weak effects on control samples and its effects were strengthened in the presence of the cytoskeleton modifying factors. The data suggest that the potential targets for the agent either increase and/or the degree of involvement of microtubules in the respiratory chain regulation, and therefore that the cytoskeleton can modify the functioning of the respiratory electron transport pathways in winter wheat cells. PMID- 10860576 TI - Ultrastructure of Chenopodium rubrum L. apices during the effect of fusicoccin and photoperiodic induction. AB - Plant transition from vegetative to reproductive development is associated with ultrastructural changes in stem apices. Those seen in Chenopodium rubrum L. under the influence of fusicoccin in many ways resemble those induced by a short-day treatment favourable to flowering. This suggests that fusicoccin can play a definite (physiological) role in plant development. PMID- 10860578 TI - Cannabinoid CB(1) receptor expression in rat spinal cord. AB - While evidence implicates the endogenous cannabinoid system as a novel analgesic target at a spinal level, detailed analysis of the distribution of the cannabinoid receptor CB(1) in spinal cord has not been reported. Here, immunocytochemical studies were used to characterize the CB(1) receptor expression in rat spinal cord. Staining was found in the dorsolateral funiculus, the superficial dorsal horn (a double band of CB(1) immunoreactivity (ir) in laminae I and II inner/III transition), and lamina X. Although CB(1)-ir was present in the same laminae as primary afferent nociceptor markers, there was limited colocalization at an axonal level. Interruption of both primary afferent input by dorsal root rhizotomy and descending input by rostral spinal cord hemisection produced minor changes in CB(1)-ir. This and colocalization of CB(1) ir with interneurons expressing protein kinase C subunit gamma-ir suggest that the majority of CB(1) expression is on spinal interneurons. These data provide a framework and implicate novel analgesic mechanisms for spinal actions of cannabinoids at the CB(1) receptor. PMID- 10860577 TI - HLA-DQ polymorphism influences progression of demyelination and neurologic deficits in a viral model of multiple sclerosis. AB - The importance of genetic susceptibility in determining the progression of demyelination and neurologic deficits is a major focus in neuroscience. We studied the influence of human leukocyte antigen (HLA)-DQ polymorphisms on disease course and neurologic impairment in virus-induced demyelination. HLA-DQ6 or DQ8 was inserted as a transgene into mice lacking endogenous expression of MHC class I (beta(2)m) and class II (H2-A(beta)) molecules. Following Theiler's murine encephalomyelitis virus (TMEV) infection, we assessed survival, virus persistence, demyelination, and clinical disease. Mice lacking expression of endogenous class I and class II molecules (beta(2)m(o) Abeta(o) mice) died 3 to 4 weeks postinfection (p.i.) due to overwhelming virus replication in neurons. beta(2)m(o) Abeta(o) DQ6 and beta(2)m(o) Abeta(o) DQ8 mice had increased survival and decreased gray matter disease and virus replication compared to nontransgenic littermate controls. Both beta(2)m(o) Abeta(o) DQ6 and beta(2)m(o) Abeta(o) DQ8 mice developed chronic virus persistence in glial cells of the white matter of the spinal cord, with greater numbers of virus antigen-positive cells in beta(2)m(o) Abeta(o) DQ8 than in beta(2)m(o) Abeta(o) DQ6 mice. At day 45 p.i., the demyelinating lesions in the spinal cord of beta(2)m(o) Abeta(o) DQ8 were larger than those in the beta(2)m(o) Abeta(o) DQ6 mice. Earlier and more profound neurologic deficits were observed in beta(2)m(o) Abeta (o) DQ8 mice compared to beta(2)m(o) Abeta(o) DQ6 mice, although by 120 days p.i. both strains of mice showed similar extent of demyelination and neurologic deficits. Delayed-type hypersensitivity and antibody responses to TMEV demonstrated that the mice mounted class II-mediated cellular and humoral immune responses. The results are consistent with the hypothesis that rates of progression of demyelination and neurologic deficits are related to the differential ability of DQ6 and DQ8 transgenes to modulate the immune response and control virus. PMID- 10860579 TI - Expression and alternative splicing of mouse Gfra4 suggest roles in endocrine cell development. AB - Members of the GDNF protein family signal through receptors consisting of a GPI linked GFRalpha subunit and the transmembrane tyrosine kinase Ret. Here we characterize the mouse Gfra4 and show that it undergoes developmentally regulated alternative splicing in several tissues. The mammalian GFRalpha4 receptor lacks the first Cys-rich domain characteristic of other GFRalpha receptors. Gfra4 is expressed in many tissues, including nervous system, in which intron retention leads to a putative intracellular or secreted GFRalpha4 protein. Efficient splicing occurs only in thyroid, parathyroid, and pituitary and less in adrenal glands. A splice form that leads to a GPI-linked GFRalpha4 receptor is expressed in juvenile thyroid and parathyroid glands. In newborn and mature thyroid as well as in parathyroid and pituitary glands major transcripts encode for a putative transmembrane isoform of GFRalpha4. Significant loss of thyroid C cells in Ret deficient mice suggests that C cells and cells in adrenal medulla, which also express Ret, may require signaling via the GFRalpha4-Ret receptor. Finally, in human, GFRalpha4 expression may restrict the inherited cancer syndrome multiple endocrine neoplasia type 2, associated with mutations in RET, to these cells. PMID- 10860580 TI - Loss of cadherin-11 adhesion receptor enhances plastic changes in hippocampal synapses and modifies behavioral responses. AB - Cadherins organize symmetrical junctions between the pre- and postsynaptic membranes in central synapses. One of them, cadherin-11 (cad11), is expressed in the limbic system of the brain, most strongly in the hippocampus. Immunohistochemical studies of the hippocampus showed that cad11 proteins were densely distributed in its synaptic neuropil zones; in cultured hippocampal neurons, their distribution often overlapped with that of synaptophysin, and also occasionally with that of GluR1 at spines. To assess the role of cad11 in synaptic formation and/or function, we analyzed brains of cad11-deficient mice. In these mice, long-term potentiation (LTP) in the CA1 region of the hippocampus was, unexpectedly, enhanced; and the level of LTP saturation was increased. In behavioral tests, the mutant mice showed reduced fear- or anxiety-related responses. These results suggest that the cad11-mediated junctions may modulate synaptic efficacy, confining its dynamic changes to a limited range, or these junctions are required for normal development of synaptic organization in the hippocampus. PMID- 10860581 TI - Metalloprotease-disintegrin (ADAM) genes are widely and differentially expressed in the adult CNS. AB - ADAM family of metalloprotease-disintegrins, including enzymes that process TNF alpha and beta-amyloid precursor protein, has been indicated in neuronal development, but the role of these protease/adhesion/signaling proteins in adult nervous system remains poorly understood. Present study provides a systematic examination of ADAM gene expression in rodent CNS, showing the first quantitative characterization of ADAM mRNA distribution therein. At least 17 ADAM mRNAs were expressed. Individual ADAM mRNAs and their isoforms showed strikingly different expression patterns. Expression of mRNAs for ADAM10, the putative alpha secretase, and ADAM17 (TACE), also indicated in APP processing, was further characterized using in situ hybridization. Expression of ADAM10 mRNA was widespread, while ADAM17 showed a more restricted pattern. Altogether, the wide and differential expression of ADAM mRNAs suggests versatile roles for ADAMs in the adult CNS. PMID- 10860582 TI - Expression of neurexin Ialpha splice variants in sympathetic neurons: selective changes during differentiation and in response to neurotrophins. AB - Neurexins are a surprisingly diverse group of alternatively spliced proteins possibly involved in neural cell recognition processes. We find neurexin Ialpha and its splice variants highly conserved between mammals and birds. In vivo, neurexin Ialpha is expressed in sympathetic neurons during target innervation and relative expression levels of splice variants change with development. In vitro, no such changes are observed in the absence of growth factors, indicating that interactions with the environment are required to modify the splicing pattern. Specific alterations in splice variant expression are induced in vitro by neurotrophins. Expression patterns of splice variants in vivo and neurotrophin induced regulation without changes in cell composition in vitro demonstrate that neurexin splice variant expression varies during differentiation of individual neurons. Our data suggest that changes in neurexin splice variants contribute to alterations of neuronal cell surface properties during target innervation. PMID- 10860583 TI - Fractal texture analysis of perfusion lung scans. AB - The purpose of this study is to investigate if fractal texture analysis can assist in the diagnostic interpretation of perfusion lung scans. Forty-five perfusion scans were acquired from patients with clinical suspicion of acute pulmonary embolism (PE) who underwent pulmonary angiography for final diagnosis. Fractal texture analysis was performed on 270 regions of interest (ROIs) extracted from the posterior view of the lung scans. Specifically, there were 94 normally perfused ROIs and 176 abnormal ROIs representing various lung diseases including PE and obstructive pulmonary disease (OPD). The average fractal dimension (FD) of normal ROIs was statistically significantly higher than that of abnormal ROIs. Furthermore, the FDs of abnormal ROIs with PE were significantly lower than the FDs of ROIs with OPD present. PMID- 10860584 TI - Accuracy of two dipolar inverse algorithms applying reciprocity for forward calculation. AB - Two inverse algorithms were applied for solving the EEG inverse problem assuming a single dipole as a source model. For increasing the efficiency of the forward computations the lead field approach based on the reciprocity theorem was applied. This method provides a procedure to calculate the computationally heavy forward problem by a single solution for each EEG lead. A realistically shaped volume conductor model with five major tissue compartments was employed to obtain the lead fields of the standard 10-20 EEG electrode system and the scalp potentials generated by simulated dipole sources. A least-squares method and a probability-based method were compared in their performance to reproduce the dipole source based on the reciprocal forward solution. The dipole localization errors were 0 to 9 mm and 2 to 22 mm without and with added noise in the simulated data, respectively. The two different inverse algorithms operated mainly very similarly. The lead field method appeared applicable for the solution of the inverse problem and especially useful when a number of sources, e.g., multiple EEG time instances, must be solved. PMID- 10860585 TI - 3-D reconstruction from tomographic data using 2-D active contours. AB - Reconstructing three-dimensional (3-D) shapes of structures like internal organs from tomographic data is an important problem in medical imaging. Various forms of the deformable surface model have been proposed to tackle it, but they are either computationally expensive or limited to tubular shapes. In this paper a 3 D reconstruction mechanism that requires only 2-D deformations is proposed. Advantages of the proposed model include that it is conformable to any 3-D shape, efficient, and highly parallelizable. Most importantly, it requires from the user an initial 2-D contour on only one of the tomograph slices to start with. Experimental results are shown to illustrate the performance of the model. PMID- 10860586 TI - Optimizing drug regimens in cancer chemotherapy by an efficacy-toxicity mathematical model. AB - In cancer chemotherapy, it is important to design treatment strategies that ensure a desired rate of tumor cell kill without unacceptable toxicity. To optimize treatment, we used a mathematical model describing the pharmacokinetics of anticancer drugs, antitumor efficacy, and drug toxicity. This model was associated with constraints on the allowed plasma concentrations, drug exposure, and leukopenia. Given a schedule of drug administrations, the mathematical model optimized the drug doses that can minimize the tumor burden while limiting toxicity at the level of the white blood cells. The main result is that the optimal drug administration is an initial high-dose chemotherapy up to saturation of constraints associated with normal cell toxicity and a maintenance continuous infusion at a moderate rate. Data related to etoposide investigations were used in a feasibility study. Simulations with the optimized protocol showed better performances than usual clinical protocols. Model-based optimal drug doses provide for greater cytoreduction, while limiting the risk of unacceptable toxicity. PMID- 10860587 TI - Surface Pressure, Hysteresis, Interfacial Tension, and CMC of Four Sorbitan Monoesters at Water-Air, Water-Hexane, and Hexane-Air Interfaces. AB - The purpose of this study was to investigate the interfacial properties of sorbitan monoesters (Span 20, 40, 60, and 80). The surface pressure was investigated at the water-air interface using a Langmuir-Blodgett apparatus. Interfacial tensions at n-hexane-air and water-n-hexane interfaces were measured by a du Nouy tensiometer. The effects of different surface-active agents and their concentrations on the interfacial properties of surfactant films were determined. With saturated sorbitan monoesters the lengthening of the hydrocarbon chain increases the collapse pressure and molecular area at the water-air interface. Unsaturated Span 80 had a lower collapse pressure and a larger molecular area than its saturated counterpart Span 60. Under compression expansion cycles, all sorbitan monoesters showed hysteresis effects. At the n hexane-air interface there were no differences in the interfacial tension between different sorbitan monoesters. At the water-n-hexane interface, differences in CMCs were small, but the surface excess of Span 80 was markedly smaller and the molecular area larger than the corresponding values of other sorbitan monoesters. Copyright 2000 Academic Press. PMID- 10860588 TI - BET Measurements: Outgassing of Minerals. AB - Outgassing minerals at elevated temperatures prior to BET measurements can lead to phase changes, especially in the case of amorphous and poorly crystalline materials. In order to evaluate the applicability of the BET method when low outgassing temperatures are required, selected aquifer minerals were outgassed at different temperatures and for different times. The studied minerals are 2-line ferrihydrite, goethite, lepidocrocite, quartz, calcite, alpha-alumina, and kaolinite. The results demonstrate that measured specific surface areas of iron oxides are strongly dependent on outgassing conditions because the surface area increased by 170% with increasing temperature. In the poorly crystalline minerals, phase changes caused by heating were observed at temperatures lower than 100 degrees C. Therefore low outgassing temperatures are preferable for minimizing phase changes. As demonstrated in this study, stable BET values can be obtained by increasing the outgassing time without heating iron oxides. For quartz, calcite, alpha-alumina, and kaolinite, stable BET values were obtained after outgassing the minerals at 100 to 250 degrees C for 2 h. However, outgassing these minerals at room temperature (20 degrees C) only resulted in minor errors, implying that aquifer sediments containing poorly crystalline materials can be outgassed at low temperatures if the outgassing time is increased. Scanning electron microscopy of the studied minerals demonstrated that the particle size as calculated from BET data compares well with particle size observed by scanning electron microscopy images. Copyright 2000 Academic Press. PMID- 10860589 TI - Analytical Solutions for Bikinetic Coagulation: Incorporation of a Maximum Size Class. AB - Analytical solutions are derived for both orthokinetic and perikinetic coagulation rate mechanisms by considering bikinetic collisions of the agglomerating particles. The bikinetic behavior assumes that only equal-sized particles collide to form larger aggregates. In addition to the bikinetic assumption used to simplify the rate equations, the coagulating particles are assumed to approach a maximum size class with increasing time. Mechanisms of perikinetic, orthokinetic, differential sedimentation, and flow-induced breakup were examined in the analyses. A solution incorporating flow-induced breakup forced the equilibrium population to a maximum size class that was less than an initially assumed maximum size class, consistent with coagulation modeling. This breakup equation is shown to be beneficial to researchers seeking to develop predictive models for orthokinetic coagulation by establishing estimates for breakup constants. The new solutions are contrasted with the existing simplified models and found to be more characteristic of actual coagulation behavior. The new solutions approach a maximum particle size class and conserve mass at increasing time, rather than an asymptotic approach to zero particles or zero mass as predicted by the existing models. Applications of the new coagulation rate solutions are useful in estimating particle collision efficiency and breakup rate constants and in determining the accuracy of numerical simulations of coagulation processes. Copyright 2000 Academic Press. PMID- 10860590 TI - Scintigraphic Measurement of Liquid Holdup in Foam Fractionation Columns. AB - The profile of liquid holdup in foam fractionation columns was studied as a function of time by the use of a nuclear scintigraphic imaging technique. Sodium [(99m)Tc]pertechnetate was employed as the imaging agent and served as a marker for the amount of water present in the foam during the foam fractionation of bovine serum albumin. A gamma scintigraphic camera fitted with a pinhole collimator was used to acquire the data in a dynamic manner. The greatest changes in the liquid holdup of the foam were observed in the region just above the foam retentate interface, particularly when a gas dispersion frit with large nominal frit porosity was employed to generate the foam. Beyond these first few centimeters, the volume fraction of water in the foam did not change appreciably. This suggests that under the conditions employed, increases in foam column height would not substantially improve the performance of the foam fractionation of bovine serum albumin. In the case of foam generated with a frit of small porosity, the liquid holdup profile indicated a higher fraction of water than observed with the larger frit. The use of scintigraphic imaging can provide valuable data concerning the liquid fraction that is not easily obtained with methods previously used in the study of drainage in foam fractionation columns. Copyright 2000 Academic Press. PMID- 10860591 TI - Monitoring Protein Fouling of Metal Surfaces via a Quartz Crystal Microbalance. AB - A quartz crystal microbalance (QCM) has been used to study the fouling of chromium and hydrophobically modified gold surfaces when heated with milk proteins. Solutions of pure beta-lactoglobulin and a commercial skimmed milk powder were studied at 3 wt%, neutral pH, and before and after heating the solutions to 80 degrees C. The ease of removal of the adsorbed protein by rinsing with buffer and 1 wt% Tween 20 was also studied. The beta-lactoglobulin behaved rather similarly on the hydrophobic gold and chromium surfaces: Tween 20 was not particularly effective in removing this protein after heating. On the other hand, Tween 20 seemed more efficient at removing the heated skimmed milk protein from the hydrophobic gold surface, but less efficient at removing the skimmed milk from the chromium surface (which also exhibited the highest adsorbed amounts of either protein). On chromium, trypsin followed by buffer removed almost all the beta-lactoglobulin but had little effect on the adsorbed layers from skimmed milk. These changes are interpreted in terms of the hydrodynamic thickness of the adsorbed films and lead to the conclusion that the QCM is a highly sensitive way of monitoring adsorbed film properties during heating, cooling, and detergent action. Copyright 2000 Academic Press. PMID- 10860592 TI - Instability of the Contact Line between Three Fluid Phases during Mass Transfer Processes. AB - The movable contact line between two liquids and a gas phase sensitively reacts to small disturbances in the force equilibrium. The shape of the contact line and the adjoining interfaces is determined by the interface and surface tensions, the contact angles, the density differences (hydrostatic pressure), and the Laplace capillary pressure. When these change, the three-phase contact line can deform and even become unstable. Interface and surface tension depend on the concentration and temperature. During mass transport processes (concentration changes) various forms of the instability of the contact line can be observed: Oscillations of a circular contact line (regular expansion and reduction); Single deformations (bulges) which quickly disappear again; -Deformations (bulges) which run along the boundary line; -Periodically generated and damped deformations with different modes. The behavior of the three-phase contact line is of practical importance for coalescence processes and for spontaneous emulsification on liquid surfaces. Copyright 2000 Academic Press. PMID- 10860593 TI - An in situ Infrared Spectroscopic Study of Glutamic Acid and of Aspartic Acid Adsorbed on TiO(2): Implications for the Biocompatibility of Titanium. AB - In situ ATR-IR spectroscopy has been applied to the study of glutamic (Glu) and aspartic (Asp) acid adsorbed on amorphous TiO(2) particle films. Unlike Asp, which gives evidence of one major adsorbed species, Glu yields several spectroscopically distinct structures upon adsorption to TiO(2). The pH dependence of Glu and Asp adsorption is also different, with Glu adsorbing markedly to TiO(2) at pH where electrostatic interactions between the surface and adsorbate are unfavorable. Application of the Langmuir model to adsorption isotherms yields a single binding constant for Asp and two binding constants for Glu, further supporting the evidence for different adsorbed Glu species. This is the first investigation of the molecular structure of Glu and Asp species adsorbed on amorphous TiO(2) using in situ ATR-IR spectroscopy. Copyright 2000 Academic Press. PMID- 10860594 TI - Force Calibration in Lateral Force Microscopy. AB - An analysis of the contact mechanics and the forces of interaction in lateral force microscopy measurements is presented. This analysis allows for a new method of interpretation of the frictional forces, the lateral contact stiffness, and the contact shear strength. The technique was developed for the interpretation of frictional data obtained with colloidal probes, although results are presented which illustrate its ability to interpret measurements recorded with both colloidal probes and standard atomic force microscopy tips. The technique is found to compensate for the variations in the contact geometry, giving repeatable results for probes of different sizes. A critical review of other techniques which have been employed to interpret the frictional force in lateral force microscopy is also presented. Copyright 2000 Academic Press. PMID- 10860595 TI - Some New Data for Metal Desorption on Inorganic-Organic Hybrid Materials. AB - A 2(4-1) fractional factorial design is employed to study Cu(II) desorption on organofunctionalized silicas. Two types of silicas, at amounts of 100 and 200 mg and two acidic levels, and contact times were employed. Our study indicates that the effects of HCl and contact time on the percentage number of moles of copper are 13.23 and 18.53, respectively. However, the binary interaction of these factors is very small. Three important antagonistic and synergistic binary effects involving the silica type, acidic quantity, and contact time factors are found. The silica mass showed an insignificant principal effect. The desorption data are compared with those previously published concerning adsorption processes on both functionalized silicas. Copyright 2000 Academic Press. PMID- 10860596 TI - Thermodynamic Approach to the Wetting of Solids by Surface-Active Agents: A Supplementary Approach to the Gibbs Formalism. AB - The adsorption of a solute on a solid can be followed by contact angle measurements of a drop of the solution on the solid. The Gibbs isotherm model can be used for quantitative interpretation of wettability variations. Its use in linking the wettability to the adsorption isotherm involves assimilating the Gibbs' planes to the surfaces themselves. Within this framework, these interpretations lead to the conclusion that adsorption of surface-active agents is greater on solid-vapor interfaces than on solid-liquid interfaces, for hydrophilic solids. This is not the only approach. Thermodynamics allows other formalisms, the conclusions of which can be completely different. We present a thermodynamic approach which explicitly reveals relationships between surface tensions and contents of surfaces, without referring to the Gibbs' plane. This permits us to explain the behavior of a drop of surfactant solution put on hydrophilic or hydrophobic solids with conclusions different from those reached using the Gibbs approach. We show that all these thermodynamic approaches are linked; they do not dismiss one another but give different views of the same phenomenon. Copyright 2000 Academic Press. PMID- 10860597 TI - Cationic Mixed Micelles in the Presence of beta-Cyclodextrin: A Host-Guest Study. AB - The conductances of trimethyltetradecylammonium bromide (TTAB)+triphenyltetradecylphosphonium bromide (TTPB) and TTAB+trimethylhexadecylammonium bromide (HTAB) over the entire mole fraction range of TTAB (alpha(TTAB)) were measured in water and in beta-cyclodextrin+water (CD+W) mixtures at fixed 4 and 8 mM of CD at 30 degrees C. The conductivity plots for both binary mixtures show a single break from which the mixed critical micelle concentration (cmc) and degree of micelle ionization (chi) were computed. From the slopes of the conductivity curves, the equivalent ionic conductivities of the monomeric (Lambda(m)), associated (Lambda(ass)), and the micelle (Lambda(mic)) states were calculated and discussed with respect to the surfactant CD complexation in the whole mole fraction range of both surfactant binary mixtures. The association constant (K) between the respective monomeric surfactant and CD cavity of fixed 4 mM CD was computed by considering 1:1 association from the surface tension measurements. A comparison among the K values for HTAB-CD, TTAB-CD, and TTPB-CD shows that the former complexation is significantly stronger in comparison to the other ones due to the longer hydrophobic tail. The nonideality in mixed micelle formation in pure water was evaluated by using the regular solution theory, and it was observed that both binary mixtures exhibit close to ideal behavior. Copyright 2000 Academic Press. PMID- 10860598 TI - Brownian Dynamics Simulation of Film Formation of Mixed Polymer Latex in the Water Evaporation Stage. AB - Brownian dynamics simulations of the filming process of a mixed polymer latex in the water evaporation stage were performed in order to explore the effect of surface potential on latex particle packing and distribution at a temperature far below the glass transitions of polymers in bulk. Polymer latex particles are modeled as spheres that interact via DLVO potential with various surface charge densities for emulsifier-free emulsion polymerized particles and dispersion polymerized particles. It is found that the distribution of modeled poly(methyl methacrylate) and polystyrene latex particles in the finally formed film exhibits a noticeable dependence of surface potentials of latex particles. When the difference of the surface potentials between binary mixed latex particles is small, the particles distribute randomly. In contrast, when the difference of the surface potentials between binary mixed latex particles is large, heterocoagulation occurs and the polymer latex in which the repulsive electrostatic potential is weak will form clusters in the film. The results are in agreement with laser confocal fluorescence microscopy observations of fluorescent dye labeled poly(methyl methacrylate) and polystyrene mixed latex films. The correlation between latex particles increases with increasing repulsive electrostatic potential, and the spatial order can be obtained at the end of the water evaporation stage. Copyright 2000 Academic Press. PMID- 10860599 TI - Preparation of Monodispersed Solid Lipid Microspheres Using a Microchannel Emulsification Technique. AB - Monodispersed solid lipid microspheres consisting of high melting point edible oil were prepared as a novel sophisticated material. To prepare the monodispersed solid lipid microspheres a temperature-controlled microchannel (MC) emulsification process was devised. The prepared microspheres had diameters of approximately 20 um or more and the standard deviation of the diameters was less than 1 um. The effects of different surfactants, the pressure and the shape of the MC on the MC emulsification, and the diameter of the prepared droplet were studied. The experimental results are discussed using the new mechanism of droplets formation which is caused by the interfacial tension. Copyright 2000 Academic Press. PMID- 10860600 TI - Reanalysis and New Measurements of N(2) and CH(4) Adsorption on Ice and Snow. AB - Numerous literature data indicate that the mean heat of adsorption of a monolayer of N(2) (DeltaQ(N(2))) on ice or snow at 77.15 K, determined by volumetric methods, is highly variable, suggesting that ice surface properties strongly depend on its mode of formation and its thermal history. Less numerous data on CH(4) adsorption show smaller variations of DeltaQ(CH(4)). If such variations are real, the extrapolation to atmospheric chemistry models of adsorption parameters measured on laboratory-made ice may be unwarranted. We have measured CH(4) adsorption on variable amounts of a crushed ice sample, to show that when the total surface area of the sample is below a threshold value, DeltaQ(CH(4)) decreases. We identify the cause of this artifact as an error in the molar budget, because the temperature gradient in the tube connecting the introduction and expansion volumes is not taken into account. Performing an adequate molar budget suppresses this artifact, except for ice samples with very small total surface areas, where the resolution of the manometer becomes a limiting factor and a further decrease in DeltaQ(CH(4)) is observed. Error in DeltaQ(gas) results in large errors in surface area, and we suggest that the value of DeltaQ(gas) obtained can be used to test the reliability of the surface area measurement. Copyright 2000 Academic Press. PMID- 10860601 TI - Adhesion between Silica Particle and Mica Surfaces in Water and Electrolyte Solutions. AB - An atomic force microscope (AFM) is used to study the adhesion between a silica sphere and a mica plate in pure water and solutions of monovalent cations (LiCl, NaCl, KCl, and CsCl). It is found that the adhesive force depends not only on the electrolyte concentration but also on the hydration enthalpy of cations and the contact time of the particle on the surface. Possible mechanisms by which the observed phenomena can be explained consistently are discussed extensively. It is suggested that the adhesive force is closely related to the structure of the layer of cations and water molecules adsorbed on the surfaces: the strong adhesive force is obtained when highly hydrated cations (Li(+), Na(+)) are adsorbed to form a thick but weakly adsorbed layer, while the weak adhesive force is observed when poorly hydrated cations (Cs(+), K(+)) are adsorbed to form a thin but strongly adsorbed layer. Copyright 2000 Academic Press. PMID- 10860602 TI - Measurement and Network Modeling of Liquid Permeation into Compacted Mineral Blocks. AB - A microbalance has been used to measure the rate of uptake of a wetting fluid, 1,3-propandiol, into a cube of compacted calcium carbonate. The cube had sides 12 mm long, with a wax band applied to the outer perpendicular edges of one basal plane to prevent external surface uptake, and the liquid was applied in a highly controlled manner at this single face only. The percolation characteristics of an identical sample were measured by mercury porosimetry. A three-dimensional void structure was generated with the same percolation characteristics using a software package called "Pore-Cor." The wetting of 1,3-propandiol into this model structure was then calculated using an extended Lucas-Washburn equation, developed by Bosanquet, which includes viscous, inertial, and capillary force effects. Neither the experimental nor the simulated wetting can be explained in terms of an "hydraulic stream tube" or "effective hydraulic radius" model. A mathematical function is presented which compensates for the differences in the boundary conditions between the simulation and the experiment. The wetting is found to be initially slowed by inertial flow, then speeded up to a t(0.8) dependence by the connectivity of the three-dimensional void network. The effect of the inertial flow is most pronounced for larger pores. Copyright 2000 Academic Press. PMID- 10860603 TI - Benzenecarboxylate Surface Complexation at the Goethite (alpha-FeOOH)/Water Interface. AB - A surface complexation model describing the adsorption of three benzenecarboxylates (phthalate, trimellitate, and pyromellitate) on goethite (alpha-FeOOH) was calibrated on data using goethite particles of 37 and 43 m(2)/g surface area. The models predict potentiometric titration and batch adsorption data with the multisite complexation model coupled with the three-plane model to account for surface electrostatics. The modeling parameters were found to be similar to those calibrated on benzenecarboxylate adsorption data on goethite particles of 90 m(2)/g (Boily et al. Geochim. Cosmochim. Acta, in press). The significance of the benzenecarboxylate-dependent values of the modeling parameters is also discussed. The values of the capacitances of the inner- and outer-Helmholtz planes were shown to be important modeling parameters to model the benzenecarboxylate-dependent slopes of the adsorption edges. It was shown that the larger the charge of the ligand, the larger the capacitance of the outer Helmholtz plane. Copyright 2000 Academic Press. PMID- 10860604 TI - Measurement of the Low-Frequency Dielectric Properties of Colloidal Suspensions: Comparison between Different Methods. AB - A careful analysis of the main factors that affect the low-frequency dielectric measurements of conducting liquid samples is carried out. The influence of the type of the measurement cell, the calibration method, and the type of the instrument used, on the spectra obtained using the variable electrode spacing technique, is investigated. Permittivity and conductivity measurements in the 10 Hz to 10 MHz range are reported for low (sigma approximately 0.01 S/m) and high (sigma approximately 0.7 S/m) conductivity samples, both electrolyte solutions and polystyrene particle suspensions. Two measurement cells are evaluated: one made of glass currently used at Granada and the other made of acrylic currently used at Tucuman. Two calibration methods, the classical Short/Open correction and the quadrupolar technique (similar to the Short/Open/Load correction), are contrasted, and two impedance analyzers, the HP 4284 A and the HP 4192 A, are compared. Copyright 2000 Academic Press. PMID- 10860605 TI - Effects of Adsorbed Polyaniline on Redox Processes on As(2)O(3) Surfaces. AB - Polyaniline deposited on As(2)O(3) surface resulted in a new material, which was characterized by infrared spectoscopy, thermogravimetry, differential scanning calorimetry, scanning electron microscopy, X-ray diffraction, and cyclic voltammetry. The mass percentage of polymer deposited on oxide surface is approximately 13%. The scanning electron microscopy images as well as the X-ray diffraction patterns provided conclusive evidence that the oxide surface is coated by the polymer. The cyclic voltammograms of the polyaniline adsorbed on As(2)O(3) surface showed that the adsorbate exerts remarkable effects on redox processes on this oxide. The pure oxide exhibited two oxidation/reduction peaks at 0.25/-0.06 and 0.47/-0.25 V attributed tentatively to the processes As(2)O(3)(s)+6H(+)+6e(-)=2As(s)+3H(2)O and As(s)+3H(+)+3e(-)=AsH(3)(g), respectively. The polyaniline-coated sample exhibited a better-defined voltammogram in which the first oxidation peak of the oxide had its intensity increased about four times. Copyright 2000 Academic Press. PMID- 10860606 TI - Surface Charge of Silver Iodide and Several Metal Oxides. Are All Surfaces Nernstian? AB - Charge titration data for silver iodide, titanium dioxide, aluminum oxide, silica dioxide and ferric hydroxide are analyzed using a generic site binding model that has the Nernst equation as a limiting form. Both the hypernetted chain and the nonlinear Poisson-Boltzmann approximations are used to relate the diffuse double layer potential to the surface charge, and a zeroth-order Stern layer is used to give the surface potential. In all cases it is shown that in the vicinity of the point of zero charge the Nernst equation accurately gives the surface charge. Copyright 2000 Academic Press. PMID- 10860607 TI - Adsorption of PBTCA on Alumina Surfaces and Its Influence on the Fractal Characteristics of Sediments. AB - The adsorption of 2-phosphonobutane-1,2,4-tricarboxylic acid (PBTCA) on Al(2)O(3) powder has been studied as a function of pH and concentration. The adsorption density of PBTCA is found to decrease with an increase of pH. Zeta potential measurements show that the addition of PBTCA results in a dramatic increase in the absolute zeta potential, as well as a shift of the isoelectric point to the more acidic region. PBTCA considerably enhances the stability of the alumina suspension via an electrostatic mechanism. The surface properties of alumina suspensions are examined by using Auger electron microscopy and FTIR analysis. Chemical interactions take place at the solid/water interface by forming complexes between Al(3+) ions and PBTCA. The dispersing ability of PBTCA is believed to be related to its high adsorption ability and the high number of dissociable protons. The morphology of the sediments is observed with scanning electron microscopy. It is found that the sediment surfaces exhibit fractal characteristics. The fractal dimension values of sediments are correlated with PBTCA concentration in the experimental range. Copyright 2000 Academic Press. PMID- 10860608 TI - Effects of Cetylpyridinium Chloride on Phase and Rheological Behavior of the Diluted C(12)E(4)/Benzyl Alcohol/Water System. AB - The influence of a small amount of cetylpyridinium chloride on the phase diagram of the diluted C(12)E(4)/benzyl alcohol/water system is studied. The presence of the ionic surfactant results in the stabilization of the vesicles in relation to the open bilayers. The bilayer structures are less sensitive to shear effects than for the system without cationic surfactant. Moreover, the formation of the mixed micelle phase is also favored. The transition from spherical to rod micelles has been detected from viscosity measurements. The micelles located close to the two-phase region (at the lower amount of cationic surfactant) are the largest and most flexible micelles. Copyright 2000 Academic Press. PMID- 10860609 TI - Steric Stabilization of Colloids by Poly(dimethylsiloxane) in Carbon Dioxide: Effect of Cosolvents. AB - Steric stabilization and flocculation of colloids with surface-grafted poly(dimethylsiloxane) (PDMS) chains are examined in liquid and supercritical carbon dioxide with and without hexane as a cosolvent. Neither poly(methyl methacrylate) (PMMA) nor silica particles with grafted 10,000 g/mol PDMS could be stabilized in pure CO(2) at pressures up to 345 bar at 25 degrees C and 517 bar at 65 degrees C without stirring. The addition of 15 wt% hexane to CO(2) led to stable dispersions with sedimentation velocities of 0.2 mm/min for 1-2 um PMMA particles. The critical flocculation pressure of the colloids in the hexane/CO(2) mixture, determined from turbidity versus time measurements, was found to be the same for silica and PMMA particles and was well above the upper critical solution pressure for the PDMS-CO(2) system. The addition of a nonreactive cosolvent, hexane, eliminates flocculation of PMMA particles synthesized through dispersion polymerization in CO(2) with PDMS-based surfactants. Copyright 2000 Academic Press. PMID- 10860610 TI - Spreading of Surfactant Solutions over Hydrophobic Substrates. AB - The spreading of surfactant solutions over hydrophobic surfaces is considered from both theoretical and experimental points of view. Water droplets do not wet a virgin solid hydrophobic substrate. It is shown that the transfer of surfactant molecules from the water droplet onto the hydrophobic surface changes the wetting characteristics in front of the drop on the three-phase contact line. The surfactant molecules increase the solid-vapor interfacial tension and hydrophilize the initially hydrophobic solid substrate just in front of the spreading drop. This process causes water drops to spread over time. The time of evolution of the spreading of a water droplet is predicted and compared with experimental observations. The assumption that surfactant transfer from the drop surface onto the solid hydrophobic substrate controls the rate of spreading is confirmed by our experimental observations. Copyright 2000 Academic Press. PMID- 10860611 TI - A Mathematical Model for Crater Defect Formation in a Drying Paint Layer. AB - Certain deep indentations observed in dry coatings are referred to as "craters". They are believed to arise from gradients in the coating surface tension. A mathematical model of surface-tension-gradient-driven flow, using the lubrication approximation for thin layers, is developed to study the formation of craters. The paint is modeled as consisting of an evaporating "solvent" part and a nonvolatile "resin" part. Surface tension gradients on the coating surface arise due to a nonuniform distribution of surfactant. Axisymmetric numerical simulations using the model are performed to explore two candidate crater production mechanisms: an initial release of concentrated surfactant and a steady surfactant source. The effects of changes in various properties, such as the paint drying rate, the surfactant diffusivity, and the viscosity increase during drying, are examined. The model produces craters with large diameters, pronounced rims, and central peaks, similar to those seen in practice. Drying rate has a large influence on crater diameter and depth, by limiting flow due to surface tension gradients within a given time. Reduction of the paint viscosity increase during drying causes increased flow rates, leading to larger craters. A preexisting layer of surfactant on the paint surface sharply reduces the extent of cratering. Surfactant diffusion also tends to reduce the severity of cratering by alleviating surface tension gradients. In some cases, a simplified form of the drying model may be used to quickly approximate the results of the full model. The model provides useful insights into the craters seen in industrial coating applications. Copyright 2000 Academic Press. PMID- 10860612 TI - In Situ Study of the Specific Adsorption of HSO(4)(-)/SO(4)(2-) Ions on Hematite by Radiotracer Technique. AB - The specific adsorption of radiolabeled sulfate ions from perchlorate supporting electrolyte on hematite powder has been investigated. It was demonstrated that a measurable adsorption occurs only at pH values under 5. The adsorption capacity of a given amount of hematite powder tends to a limiting value with decreasing pH. A Langmuir-like adsorption isotherm describes the concentration dependence of the adsorption of sulfate species. It was found that the experimental results are in good agreement with the model suggested recently (Hug, S. J. J. Colloid Interface Sci. 188, 415 (1997)). Copyright 2000 Academic Press. PMID- 10860613 TI - Effect of a Dynamic Stern Layer on the Sedimentation Velocity and Potential in a Dilute Suspension of Colloidal Particles. AB - In this paper the theory of the sedimentation velocity and potential (gradient) in a dilute suspension of charged spherical colloidal particles developed by Ohshima et al. (H. Ohshima, T. W. Healy, L. R. White, and R. W. O'Brien, J. Chem. Soc., Faraday Trans. 2, 80, 1299 (1984)) has been modified to include the presence of a dynamic Stern layer on the particle surfaces. The starting point has been the theory that Mangelsdorf and White (C. S. Mangelsdorf, and L. R. White, J. Chem. Soc., Faraday Trans. 86, 2859 (1990)) developed to calculate the electrophoretic mobility of a colloidal particle allowing for the lateral motion of ions in the inner region of the double layer (dynamic Stern layer). The effects of varying the different Stern layer parameters on the sedimentation velocity and potential are discussed and compared to the case when a Stern layer is absent. The influence of electrolyte concentration and zeta potential of the particles is also analyzed. The results show that regardless of the chosen set of Stern layer and solution parameters, the presence of a dynamic Stern layer causes the sedimentation velocity to increase and the sedimentation potential to decrease, in comparison with the standard case (no Stern layer present). These changes are almost negligible when sedimentation velocity is concerned, but they are very important when it comes to the sedimentation potential. A justification for this fact can be given in terms of an Onsager reciprocal relation, connecting the magnitudes of the sedimentation potential and the electrophoretic mobility. As previously reported, the presence of a dynamic Stern layer exerts a great influence on the electrophoretic mobility of a colloidal particle, and by means of the Onsager relation, the same is confirmed to occur when the sedimentation potential is concerned. Copyright 2000 Academic Press. PMID- 10860614 TI - Density Functional Theory and the Capillary Evaporation of a Liquid in a Slit. AB - Density functional theory is applied to a Lennard-Jones fluid near a single hard wall and in a slit formed by two walls. We use some simplified versions of the Weeks-Chandler-Andersen (WCA) and the Barker-Henderson (BH) theories. Only the most crude mean field version of the WCA theory, in which the hard-sphere correlation function is set equal to unity for all distances, seems useful. Use of the full WCA approximation is impractical because the effective hard-sphere diameter is density dependent. Generally, the best results are obtained using the BH macroscopic compressibility approximation. Our earlier study of "evaporation" of Lennard-Jones molecules in a slit is extended to other densities using the mean field theory. Copyright 2000 Academic Press. PMID- 10860615 TI - Temperature Influence of Benzene Adsorption by a Microporous Silica. AB - The theory for volume filling of micropores is used to describe benzene adsorption isotherms measured over a 25 K temperature range. The adsorption potential or molar work of adsorption for the isotherm at 298 K is derived and compared with Weibull, Gaussian, and gamma potential distribution functions. The Weibull function is fitted via a two-term Dubinin-Radushkevich (D-R) equation. The closest data fit occurs for the gamma distribution. The two-term D-R potentials are interpreted as indicating adsorption by primary micropores followed sequentially by secondary micropores. Analysis of the distribution of adsorption enthalpy for the porous solid compared with a nonporous standard suggests that the predominant pore width is 1.2 nm. The interpretation of the differential molar adsorption entropy at 298 K suggests that strongly localized adsorption occurs in the primary micropores and two-dimensional translational motion with rotation in the plane of the ring occurs in the secondary micropores. Copyright 2000 Academic Press. PMID- 10860616 TI - An Infrared Study of the Amine-Catalyzed Reaction of Methoxymethylsilanes with Silica. AB - A thin film infrared technique was used to investigate the reaction of methoxysilanes and amines with the silica surface. The low-frequency region contains bands due to Si-O-Si modes that are used to distinguish between hydrogen bonded and chemisorbed species. It is shown that the competitive adsorption of amines and CH(3)OSi(CH(3))(3) differs from the results obtained using (CH(3)O)(2)Si(CH(3))(2) or (CH(3)O)(3)SiCH(3). The monomethoxysilane does not displace preadsorbed triethylamine whereas the triethylamine is displaced from the surface by both (CH(3)O)(2)Si(CH(3))(2) and (CH(3)O)(3)SiCH(3). In the reverse sequence, the triethylamine displaces all three methoxysilanes on the surface. When 1:1 mixtures of methoxysilanes and triethylamine (or propylamine) are co-added to silica, the amine preferentially adsorbs and is only displaced by subsequent chemisorption of the silane. The implication of these results for using a two-step amine-catalyzed reaction of methoxysilanes on silica is discussed. Copyright 2000 Academic Press. PMID- 10860617 TI - Phenomena Affecting the Equilibrium of Al(III) and Zn(II) Extraction with Winsor II Microemulsions. AB - The extraction of Al(III) and Zn(II) from an aqueous solution with two water-in oil microemulsions, one containing di(2-ethylhexyl)phosphoric acid (DEHPA), was investigated to aid the understanding of the role of the extractant and the metal specific characteristics in the mechanism of microemulsion extraction. The extraction of Al with the DEHPA microemulsion increased by a factor of about 10 with respect to that in the conventional DEHPA system, whereas the extraction of Zn was lower than that in the single DEHPA system. Extraction with the DEHPA-free microemulsion was very low, showing that metal ion solubilization was not important in the mechanism of microemulsion extraction. It is proposed that the effect of the mixed microemulsion on the metal distribution coefficient is the result of the balance between a decrease in the complexation reaction yield due to the interaction between butanol and DEHPA, and the adsorption of the metal complex at the macro- and microinterfaces. The former leads to a decrease in Zn(II) extraction and the latter to Al(III) extraction synergism. Copyright 2000 Academic Press. PMID- 10860618 TI - A Hyperbranched Poly(ethyleneimine) Grown on Surfaces. AB - Aminosilylated substrate was treated with aziridines in order to prepare hyperbranched polymers on solid supports such as silicon wafer and fused silica. It is observed that the primary amine on the substrate is good enough to initiate the ring-opening polymerization of aziridine. Measuring the thickness of the film and the absolute density of the primary amine functionality shows that a very highly branched poly(ethyleneimine) is formed upon the reaction. The surface density of the primary amine functional group (-NH(2)) on the very top surface increased dramatically (from 3.5 amines/nm(2) to 66 amines/nm(2)). A protected aziridine, benzyl 1-aziridinecarboxylate, was employed for the stepwise growth of the film. Two step-processes, chain growth and deprotection, were successful in growing the molecular layer, and a linear chain was formed without branching. Copyright 2000 Academic Press. PMID- 10860619 TI - The enhancement of the ability of mouse sperm to survive freezing and thawing by the use of high concentrations of glycerol and the presence of an Escherichia coli membrane preparation (Oxyrase) to lower the oxygen concentration. AB - The cryobiological preservation of mouse spermatozoa has presented difficulties in the form of poor motilities or irreproducibility. We have hypothesized several underlying problems. One is that published studies have used concentrations of the cryoprotectant glycerol that are substantially lower (<0.3 M) than the approximately 1 M concentrations that are optimal for most mammalian cells. Another may arise from the known high susceptibility of mouse sperm to free radical damage. We have been able to obtain high motilities in 0.8 M glycerol provided that the exposure time is held to approximately 5 min to minimize toxicity and provided that the glycerol is added and removed stepwise to minimize osmotic shock. Since free radical damage in mouse sperm is proportional to the oxygen concentrations, we have determined the consequences of reducing the oxygen to <3% of atmospheric by maintaining the sperm in contact with an Escherichia coli membrane preparation, Oxyrase, from the moment of collection throughout the assessment of motility. Prior studies have shown that the procedure significantly reduces damage from centrifugation and osmotic shock. In the experiments reported here we obtained approximately 50% motility relative to untreated controls when suspensions containing 3.8% Oxyrase were exposed approximately 5 min to a solution of 0.8 M glycerol and 0.17 M (10%) raffinose in a supplemented PBS and then frozen at approximately 25 degrees C/min to -75 degrees C. In the absence of Oxyrase, the normalized motility dropped to 31%. The protection by Oxyrase was in part a consequence of minimizing centrifugation damage, but in part it reflected a reduction in freeze-thaw damage. Preliminary experiments indicate that the number of motile sperm after cryopreservation in Oxyrase is higher when the sperm are collected without swim-up than when they are collected by swim-up. This is in part due to the fact that more cells are collected in the absence of swim-up and in part due to a greater protective effect of Oxyrase on those cells. The minimum temperature in these initial experiments was limited to -75 degrees C to avoid the potential contribution of other injurious factors between -75 and -196 degrees C. PMID- 10860620 TI - Cryohemostasis of uncontrolled hemorrhage from liver injury. AB - Uncontrolled hemorrhage is the primary cause of death in both blunt and penetrating liver trauma. Cryohemostasis was attempted in the past for elective liver surgery but did not gain popularity. During past decades, cryoequipment was refined and successfully used for tumor ablation. The purpose of the present study was to assess the efficacy of cryosurgery as a potential adjuvant hemostatic technique in the treatment of grades III-IV liver injuries. A standard liver crush-evulsion injury was created in pigs. In the control group, the liver was left to bleed freely. In the experimental group, the severed liver surface was immediately frozen to -160 degrees C for 10 min, spontaneously thawed, and left to bleed thereafter. Blood pressure, pulse rate, urine output, and serum lactate were monitored. The total blood loss was measured 180 min after liver injury was inflicted. The volume of frozen liver parenchyma was measured. For further laboratory evaluation, three additional experimental animals were not sacrificed and recovered. Cryohemostasis significantly reduced blood loss and substantially attenuated hemorrhagic shock. The frozen liver parenchyma underwent necrosis but did not jeopardize survival. Cryosurgery may be an efficient adjuvant technique in the early control of hemorrhage in grades III-IV liver injury. PMID- 10860621 TI - Distribution and characterization of recrystallization inhibitor activity in plant and lichen species from the UK and maritime Antarctic. AB - Extracts from a range of evolutionarily diverse plant and lichen species from the UK and maritime Antarctic have been assayed for inhibition of ice recrystallization. Approximately 25% of overwintering UK species and all Antarctic species exhibited antifreeze activity when exposed to low temperature. Preliminary characterization of the active extracts has demonstrated that the molecules co-opted to antifreeze activity by different species are biochemically diverse. PMID- 10860622 TI - Vitrification enhancement by synthetic ice blocking agents. AB - Small concentrations of the synthetic polymer polyvinyl alcohol (PVA) were found to inhibit formation of ice in water/cryoprotectant solutions. Ice inhibition improved with decreasing molecular weight. A PVA copolymer of molecular weight 2 kDa consisting of 20% vinyl acetate was found to be particularly effective. PVA copolymer concentrations of 0.001, 0.01, 0.1, and 1% w/w decreased the concentration of glycerol required to vitrify in a 10-ml volume by 1, 3, 4, and 5% w/w, respectively. Dimethyl sulfoxide concentrations required for vitrification were also reduced by 1, 2, 2, and 3% w/w, respectively. Crystallization of ice on borosilicate glass in contact with cryoprotectant solutions was inhibited by only 1 ppm of PVA copolymer. Devitrification of ethylene glycol solutions was also strongly inhibited by PVA copolymer. Visual observation and differential scanning calorimeter data suggest that PVA blocks ice primarily by inhibition of heterogeneous nucleation. PVA thus appears to preferentially bind and inactivate heterogeneous nucleators and/or nascent ice crystals in a manner similar to that of natural antifreeze proteins found in cold hardy fish and insects. Synthetic PVA-derived ice blocking agents can be produced much less expensively than antifreeze proteins, offering new opportunities for improving cryopreservation by vitrification. PMID- 10860623 TI - Factors affecting the survival of frozen-thawed mouse spermatozoa. AB - Mouse epididymal spermatozoa were frozen in solutions containing various compounds with different molecular weights, and the factors affecting the postthawing survival were examined. Monosaccharides (glucose, galactose) had almost no protective effect regardless of the concentration and the temperature of exposure. On the other hand, disaccharides (sucrose, trehalose) and trisaccharides (raffinose, melezitose) resulted in higher survival rates, especially at a concentration of around 0.35 mol/kg H(2)O (0.381-0.412 Osm/kg). Macromolecules, such as PVP10, Ficoll 70, bovine serum albumin, and skim milk had almost no effect, but compounds with a molecular weight of about 800, such as metrizamide and Nycodenz, had some protective effect. When a raffinose solution was supplemented with 10% metrizamide, resulting in an osmolality of approximately 0.400 Osm/kg, a high survival rate was obtained. Solutions at about 0.400 Osm/kg containing trehalose alone, trehalose + metrizamide, raffinose alone, and raffinose + metrizamide, were all effective for sperm freezing; frozen thawed sperm could fertilize oocytes, and the resultant embryos could develop to live young after transfer. For freezing mouse spermatozoa, aqueous solutions at approximately 0.400 Osm/kg containing a disaccharide or a trisaccharide seem to be effective. PMID- 10860624 TI - Assessment of some critical factors in the freezing technique for the cryopreservation of precision-cut rat liver slices. AB - A number of studies on the cryopreservation of precision-cut liver slices using various techniques have been reported. However, the identification of important factors that determine cell viability following cryopreservation is difficult because of large differences between the various methods published. The aim of this study was to evaluate some important factors in the freezing process in an effort to find an optimized approach to the cryopreservation of precision-cut liver slices. A comparative study of a slow and a fast freezing technique was carried out to establish any differences in tissue viability for a number of endpoints. Both freezing techniques aim at the prevention of intracellular ice formation, which is thought to be the main cause of cell death after cryopreservation. Subsequently, critical variables in the freezing process were studied more closely in order to explain the differences in viability found in the two methods in the first study. For this purpose, a full factorial experimental design was used with 16 experimental groups, allowing a number of variables to be studied at different levels in one single experiment. It is demonstrated that ATP and K(+) content and histomorphology are sensitive parameters for evaluating slice viability after cryopreservation. Subsequently, it is shown that freezing rate and the cryopreservation medium largely determine the residual viability of liver slices after cryopreservation and subsequent culturing. It is concluded that a cryopreservation protocol with a fast freezing step and using William's Medium E as cryopreservation medium was the most promising approach to successful freezing of rat liver slices of those tested in this study. PMID- 10860625 TI - Cryoprotectants lead to phenotypic adaptation to freeze-thaw stress in Lactobacillus delbrueckii ssp. bulgaricus CIP 101027T. AB - This study was conducted to investigate the ability of cryoprotective chemicals to induce phenotypic cryoadaptation in Lactobacillus delbrueckii ssp. bulgaricus CIP 101027T. Tolerance to negative temperature stress (freezing at -20 degrees C and thawing at 37 degrees C) was induced by pretreatment with Me(2)SO, glycerol, lactose, sucrose, and trehalose. Interestingly, Me(2)SO has a significantly greater cryoprotective effect than glycerol. Furthermore, lactose, sucrose, and trehalose, often referred to as osmotica, were shown to have greater cryoadaptive than cryoprotective properties. These results suggest that bacteria such as L. delbrueckii ssp. bulgaricus could be phenotypically adapted to freezing and thawing by an osmotic stress applied prior to freeze-thaw stress. PMID- 10860626 TI - Glutathione content during the rinsing and rewarming process of rat hepatocytes preserved in University of Wisconsin solution. AB - The addition of glutathione (GSH) to University of Wisconsin (UW) solution increases the intracellular content of GSH and decreases the release of lactate dehydrogenase used here as a measure of cell viability. However, we found a depletion of GSH when the cells were transferred from UW solution to the rewarming solution. This could sensitize the cells to various forms of oxidative injury. In this study we examined how different compositions of rinsing and rewarming solutions affected the GSH content and the viability of hepatocytes after 72 h of cold storage. For both the rinsing and the rewarming steps we used a Krebs-Henseleit solution with the addition of GSH, methionine, or both GSH and methionine. We found no loss of GSH when the hepatocytes were rinsed in the presence of 3 mM GSH. During the rewarming step we observed a loss of GSH in all of the study groups, but the cells that were incubated with 1 mM methionine showed a lesser depletion of GSH and improved viability. This finding may have valuable applications in hepatocellular transplantation and in the development of bioartificial liver support devices. PMID- 10860627 TI - Literature review: supplemented phase diagram of the trehalose-water binary mixture. AB - Trehalose is of great interest in many fields, including freeze-drying, cryoprotection, and anhydrobiosis. Although data for the trehalose-water supplemented phase diagram have previously appeared in the literature, the data have been widely scattered and reported in several units. In this study, literature data for the binary trehalose-water system were collected and analyzed. The literature data were found to be reasonably consistent, with substantial agreement on the melting points for water, trehalose, and trehalose dihydrate and the glass transition temperature of water. There was also good agreement for the solubility, freezing, and glass transition curves. However, there was no general agreement on the glass transition temperature of pure trehalose. Additionally, the trehalose-water glass transition curve was modeled using the Gordon-Taylor equation, with a value for k of 5.2. The collected data in this report will be of much use in further studies of the protective abilities of trehalose. PMID- 10860628 TI - Isolated cardiac cells for electropharmacological studies. AB - Single cardiac myocytes provide a model widely used to characterize the electrophysiological properties of drugs and to identify new therapeutic targets. This review focuses on isolation procedures to obtain single cardiac myocytes from several mammal species, including humans, and on patch-clamp technique as a useful method to investigate the molecular mechanism of druy actions. PMID- 10860629 TI - Endothelial cells in culture: a model for studying vascular functions. AB - Vascular endothelium - lining the inner side of blood bessels - is one of the largest secretory tissues of the body. Therefore, understanding the cellular and molecular biology of the endothelial cells is essential for the development of new approaches for both the prevention and therapy of cardiovascular diseases. To this aim, in vitro cultures of endothelial cells provide a valuable technical resource. This review focuses on some of the critical phases of the endothelial cells culturing methodology such as: i) isolation and growing of endothelial cells; ii) identification of endothelial cells by morphological, biochemical and cellular markers; iii) studying endothelial cells in function of vascular pharmacology, vasomotor tone, vessel remodelling (angiogenesis/apoptosis), blood haemostasis, inflammatory reactions, and molecular engineering. Practical suggestions for culturing endothelial cells are presented while pros and cons of each method are discussed. PMID- 10860630 TI - Thiamin status in furosemide-treated rats. AB - Rats were treated with furosemide at intraperitoneal doses of 10, 20, and 30 mg kg(-1)day(-1)for 21 days, and some indices of thiamin status (liver, blood and urine thiamin concentrations, erythrocyte transketolase (TK) activity, and thiamin pyrophosphate (TPP) effect, which refers to the increase in TK activity in a deficient haemolysate due to the addition of TPP before incubation) were measured. The results indicated that, at doses of 20 and 30 mg kg(-1)(but not 10 mg kg(-1)), furosemide produced significant effects on these indices suggestive of thiamin deficiency. In another experiment, the effect of thiamin treatment (20 and 100 microg kg(-1)) in rats concomitantly given furosemide (30 mg kg(-1)) was investigated. The high dose of the vitamin was sufficient to overcome the biochemical signs of thiamin deficiency induced by furosemide. PMID- 10860631 TI - The situation of OTC drugs in italy compared to the other EU states. AB - Even if a specific directive has been approved many years ago, the situation of self-medication products (OTC) in EU countries is still far from being harmonized. In Italy the market is lower than that of most other countries; in order to solve some of the major problems that led to this situation a guideline, concerning the criteria for the definition of an OTC product, and the characteristics of the label and the package leaflet, was recently published. In this document the characteristics of OTC, such as composition, indications and duration of the treatment are assessed. The European Commission has recently published a guideline on the readability of labels and package leaflets of medicinal products for human use. The two documents stated the same principles and the Italian document is in agreement with the European guidelines. In this paper the Italian situation of OTC products (definition and presentation) is presented and discussed. PMID- 10860632 TI - A proposal to improve the supply of orphan drugs. AB - Industrial medicinal products are not always available for the treatment of some specific patients or diseases. Appropriate research on orphan drugs, neglected doses or administration routes and patients' sensitivity to excipients, as an example, might lead to interesting results in this respect. In the United States, a specific legislation on orphan drugs dates back to 1983; in the European Union, regulations are currently being studied. The industrial production of orphan drugs might be improved following the adoption of adequate incentives. New regulations should also take care of neglected doses, and stimulate the preparation of galenic products, as an example by including a number of monographs on orphan drugs in Pharmacopoeias and National Formularies. As a matter of fact, magistral and officinal preparations made in community and hospital pharmacies in accordance with a medical prescription or with the prescription of a Pharmacopoeia or an Official Formulary, can represent an effective alternative. In this paper, the regulations enforced in the USA and Japan, the European situation, possible incentives to the industry and the role of galenic products will be discussed. PMID- 10860633 TI - Cost-effectiveness analysis of oral N-acetylcysteine as a preventive treatment in chronic bronchitis. AB - Chronic bronchitis has a prevalence of approximately 11% in the population aged over 35 years and its frequent acute exacerbations (AECBs) are an important cause of morbidity and costs in health-care resources. Oral N -acetylcysteine (NAC) is administered during the winter months as a way of reducing AECBs. This cost effectiveness analysis was done from the payers' point of view in the Swiss health-care system, based on a retrospective analysis of published placebo controlled studies. The pooled data show that continuous administration of 400 mg day(-1)per os of NAC leads to a significant reduction in the number of AECBs (NAC: 16.2 vs 25.2% AECBs per month); a significantly smaller percentage of days of sick leave (NAC: 3.6 vs 5.3%) and a lower rate of hospitalizations (NAC: 1.5 vs 3.5% over a period of 6 months). Taking into account the poor compliance of these patients, calculations assumed a compliance of 80%. Direct costs were those of an NAC treatment, the management of an AECB (biological tests in 59%, X-rays in 65% and pulmonary function tests in 45%; antibiotics 70%, bronchodilators in 89%, corticosteroids in 24% and 'others' in 25% of the patients), and of hospitalizations (estimated at 10 days per case). Based on these figures, the mean direct costs of an untreated patient were CHF 869 vs CHF 700 in the NAC treated patient. Univariate sensitivity analysis indicated that cost neutrality is reached with 0.6 (<0.25-1. 94, 95% CI) AECBs per 6 months. Indirect costs (based on sick leave) were also significantly different; the mean in untreated patients was CHF 1324 vs CHF 779 in the NAC-treated patients. CONCLUSION: Treating chronic bronchitis patients with NAC during the winter months is cost effective both from the payer's and a social point of view. PMID- 10860634 TI - Action of ozonized water in preclinical inflammatory models. AB - Ozonized water increases oxygen local tension. The ozone by itself produces inflammatory reactions in lung epithelium. The action of ozonized water during the inflammatory process was investigated. In rat paw edema induced by carrageenin, the inflammatory process was stimulated by ozonized water (1.2 microg, p.o.) when applied 30 min prior to the induction. In the inflammatory process induced by Freund's Complete Adjuvant, the ozonized water did not interfere in the inflammatory reaction. Furthermore, in gastric ulcers models induced by stress, there was a significant reduction in the incidence of ulcers types I, II and III (P<0.05, Student's t -test). PMID- 10860635 TI - Pharmacological activity of natural lipids on a skin barrier disruption model. AB - The lipids of the stratum corneum are considered responsible for the most important functions of the skin, such as the transepidermal water loss, as well as the transdermal penetration of the chemical substances. Topical application of lipids similar to the physiological stratum corneum (SC) on barrier disrupted skin, could enhance the recovery rate of the skin barrier. A mixture of natural lipids or liposomes with the same lipid composition, were applied and their pharmacological action was investigated. The tests were done in vivo, on the back of hairless mice. Comparative results were obtained and showed that the liposomes had a higher turnover of the skin barrier in contrast to that of the mechanical mixture of lipids. PMID- 10860636 TI - Glucocorticoid-induced alterations in the rate of diaphragmatic fatigue. AB - These experiments tested the hypothesis that in vitro diaphragmatic fatigue resistance is enhanced in animals treated with glucocorticoids. Female Sprague Dawley rats (4 months old) were randomly assigned to a control (N =12) or glucocorticoid treatment group (N =12). Treatment animals were injected daily for 8 days with prednisolone (5 mg kg(-1)); control animals were injected with the same volume of the vehicle. Twenty-four hours after the last injection, the following in vitro diaphragmatic contractile properties were examined in costal diaphragm strips: maximal twitch (P(t)) half time to peak tension (1/2 TPT), half relaxation time (1/2 RT), force-frequency relationship, and the rate of fatigue development. Diaphragmatic fatigue was assessed by monitoring the decrease in force production (measured as percent of initial force) over a 60-min contractile period. The in vitro fatigue protocol incorporated a supramaximal stimulus delivered at 30 Hz every 2 s with a train duration of 250 ms (duty cycle 12.5%). Citrate synthase (CS), superoxide dismutase (SOD), and water content of the costal diaphragm were also determined. Glucocorticoid administration induced an 18.9% (P<0.05) decrease in animal body weight when compared to the control. Similar weight losses also occurred in the diaphragm with a decrease (P<0.05) in mass of 16.5% compared to the control. Furthermore, prednisolone treatment resulted in a significant reduction in the cross-sectional area (CSA) of type IIb fibres with no change in the CSA area of type I and IIa fibres. 1/2 TPT and 1/2 RT were significantly prolonged (P<0.05) in the glucocorticoid treated rats whereas the force-frequency curve was unaltered (P>0.05). Fatigue resistance was greater in the glucocorticoid group (P<0.05); the relative force production differed between groups at the end of 1 min of contractions and remained different throughout the 60-min fatigue protocol. Citrate synthase, SOD, and water content were not different between groups. These experiments support the hypothesis that costal diaphragm strips of glucocorticoid-treated rats possess a greater resistance to fatigue. We postulate that this fatigue resistance is due to glucocorticoid-induced changes muscle fibre type composition. PMID- 10860637 TI - Nitric oxide and prostanoid-dependent relaxation induced by angiotensin II in the isolated precontracted mouse tracheal muscle and the role of potassium channels. AB - In this study we examined the mechanism of the concentration-dependent relaxant effect of angiotensin II (A II) on mouse isolated tracheal rings precontracted by carbachol. The contractile effect of carbachol is increased while the relaxant effect of A II decreased by pretreatment with N(G)-nitro-L-arginine methyl ester (L-NAME). This finding suggests that the L-arginine nitric oxide (NO) pathway is activated by the peptide. The relaxing effect of A II was also mediated through the release of cyclo-oxygenase products of arachidonic acid, as evidenced by the inhibition of the response by lysine acetylsalicylic acid (ASA) pretreatment. A II also caused a relaxation in the isolated tracheal rings precontracted by K(+)(>60 m m). K(+)-channel blockers partially inhibited the relaxant effect of A II in carbachol-precontracted mouse tracheal rings. A non-selective blocker of K(+)channels, tetraethylammonium, completely abolished the relaxation induced by A II. Among the other channel blockers, the inhibitory effect of apamine and glibenclamide was higher than that of iberiotoxin, indicating the involvement of K(ATP)and small conductance K(Ca)channels in the relaxing response to the octapeptide. These results suggest that the mechanisms, involved in the relaxation induced by A II in the isolated mouse tracheal rings precontracted by carbachol, were firstly l -arginine NO and cyclo-oxygenase products of arachidonic acid, secondly K(+)channels. PMID- 10860638 TI - Mechanisms for the hepatoprotective action of kolaviron: studies on hepatic enzymes, microsomal lipids and lipid peroxidation in carbontetrachloride-treated rats. AB - The present work examines the protective mechanisms of a biflavonoid fraction of an extract from Garcinia kola seeds, kolaviron, in rats treated with carbontetrachloride (CCl(4)). CCl(4)administered at a dose of 1.2 g kg(-1), three times a week for 2 weeks, significantly depressed the activities of microsomal aniline hydroxylase, aminopyrine N -demethylase, ethoxyresorufin O -demethylase and p -nitroanisole O -demethylase. Kolaviron (200 mg kg(-1)), administered for 14 days consecutively, inhibited (P<0.001) the CCl(4)mediated decrease in the activities of these enzymes by 60, 65, 55, and 63%, respectively. Kolaviron reduced the CCl(4)increase in the cholesterol/phospholipid ratio. Similarly, kolaviron attenuated the toxic onslaught imposed by CCl(4)on 5'nucleotidase, glucose 6-phosphatase (microsomal marker enzymes) and malondialdehyde formation by 41, 54 and 77%, respectively. Kolaviron elicited 168% and 234% increases in the activity of UDP-glucuronosyl transferase and glutathione S -transferase. Simultaneous administration of kolaviron with CCl(4)modulated the effect of CCl(4)on the activities of these enzymes. On the basis of the above data, it can be postulated that kolaviron exerts its protective action against carcinogen induced liver damage, first, by acting as an in vivo natural antioxidant and, second, by enhancement of drug-detoxifying enzymes. PMID- 10860639 TI - The effect of rebamipide on cisplatin-induced nephrotoxicity in rats. AB - This study aimed to evaluate the protective effect of rebamipide (free radical scavenger) against the nephrotoxic effect induced by cisplatin in normal rats. Twenty-four male Wister albino rats were divided equally into four groups: control, rebamipide, cisplatin and cisplatin plus rebamipide-treated groups. Nephrotoxicity was induced with single intravenous (i.v.) cisplatin dose of 6 mg kg(-1)and measured through the estimation of kidney weight, serum albumin (Alb), serum creatinine (Cr), blood urea nitrogen (BUN), kidney glutathione (GSH) and malondialdehyde (MDA) production. In the cisplatin-treated group the kidney weight as a percent of the total body weight, serum Alb, serum Cr, BUN, GSH content and MDA amount were: 0.61+/-0.054%, 2.84+/-0.24 g dl(-1), 2.99+/-0.10 mg dl(-1), 147.08+/-7.46 mg dl(-1), 3.11+/-0.238 micromol g(-1)and 1449. 09+/-127.36 nmol g(-1), respectively. All the previous changes were significantly (P<0.01) different from the corresponding values in the control group. In addition, histopathological examination of the kidney tissue revealed degenerative cellular material and apoptotic tubular cells were seen in the renal tubules. Rebamipide treatment (140 mg kg(-1), i.p.) for 1 week ameliorated all the previous changes and the results recorded for the cisplatin plus rebamipide-treated group were: 0.45+/-0.035%, 4.17+/-0.091 g dl(-1), 1.37+/-0.209 mg dl(-1), 72.25+/-5.14 mg dl( 1), 5.063+/-0.269 micromol g(-1)and 560.23+/-21.98 nmol g(-1)for the previous tests, respectively. Furthermore, significant improvement in the kidney histopathology was observed. The results of this study clearly revealed that rebamipide protected the kidney against the nephrotoxic effect of cisplatin. These results suggest that lipid peroxidation is not the only mechanism by which cisplatin induced nephrotoxicity. More investigations are needed to confirm the effect of rebamipide and at the same time to elucidate the exact mechanism by which cisplatin induces nephrotoxicity. PMID- 10860640 TI - Stereoselective biotransformation of cicletanine in cultured rat and human hepatocytes. AB - Cicletanine [(+/-)-C] is a racemic furopyridine derivative used as an antihypertensive agent. Pharmacokinetic and metabolic studies have shown that cicletanine is rapidly and almost fully metabolized into sulfo- and glucuro conjugated metabolites. However, the stereoselective metabolism of cicletanine is not well-known in humans. In the present study, the stereoselective aspect of cicletanine metabolism was investigated in cultured hepatocytes from humans and rats. The two enantiomers of cicletanine were both strongly metabolized in rat hepatocytes. So, after 24 h of incubation, very low amounts of free cicletanine were found [1.2% for (+)-C and 2.7% for (-)-C], respectively. In addition (+/-)-C was mainly biotransformed into conjugated metabolites: (-)-C mainly transformed into (-)-C-glucuronide (78.3+/-6.4%) and (+)-C mainly into (+)-C-sulfate (87.4+/ 3.3%). In human hepatocytes, inter-individual variations were more marked than in rat hepatocytes. In addition (+/-)-C biotransformation in human was lower than the one observed in rat. (-)-C enantiomer was more metabolized than (+)-C. After a 24-h incubation the percentages of free (+)-C and (-)-C were 32.3+/-16.4 and 8.2+/-10.3, respectively. On the contrary to rat hepatocytes, both cicletanine enantiomers in humans were mainly metabolized into glucuroconjugated metabolites. These results clearly demonstrated that cicletanine underwent stereospecific metabolism in both rat and human hepatocytes. PMID- 10860641 TI - Isoteoline, a putative serotonin antagonist, inhibits meta chlorophenylpiperazine, but not 1-(2, -dimethoxy-4-iodphenyl)-2-aminopropane and 8-hydroxy-2-(di-n-propylamino)-tetraline-induced increase of serum prolactin levels. AB - Serotonin, in addition to dopamine and other factors, is known to participate in the control of prolactin (PRL) and gonadotropins secretion. Isoteoline (IST), a putative serotonin antagonist and dopamine agonist, was studied for its neuroendocrine effects on PRL, follicle-stimulating hormone (FSH) and luteinizing hormone (LH). IST was given intraperitoneally to adult male rats at doses of 0.25, 1 and 4 mg kg(-1)alone and 30 min prior to the injection of three 5-HT agonists with preferential affinity for various receptor subtypes: meta chlorophenylpiperazine (m CPP) for 5-HT2C; 1-(2, 5-dimethoxy-4-iodophenyl)-2 aminopropane (DOI) for 5-HT2A and 8-hydroxy-2-(di- n -propylamino)tetralin (8-OH DPAT) for 5-HT1A. m CPP (2.5 mg kg(-1)), DOI (2.5 mg kg(-1)) and 8-OH-DPAT (1 mg kg(-1)) increased the serum PRL levels to a similar value, without affecting FSH and LH concentrations. IST by itself modified neither PRL nor gonadotropins serum levels. IST antagonized the m CPP-induced elevation in serum PRL, the lowest dose being the most effective. It had no effect on DOI and 8-OH-DPAT-induced increases of PRL levels and produced no significant changes in the gonadotropins levels when used as an antagonist. The results are discussed in terms of the likely involvement of serotonin vs dopamine mechanism in the effect of IST. It is concluded that the inhibition of the m CPP-induced rise of PRL levels by IST confirmed the serotonin antagonistic activity, previously demonstrated for this compound in other studies. The present results are also suggestive of possible selectivity of this antagonism of IST for the 5-HT2C vs 5-HT2A and 5-HT1A receptors, all of which are involved in the control of PRL secretion. PMID- 10860642 TI - Effect of calcium channel blockers on baroreceptor reflex in anaesthetized cats. AB - The baroreflex-induced changes in heart rate in chloralose anaesthetized and artificially ventilated cats (2.5-4.0 kg) before and after pretreatment with calcium channel blockers (CCBs) were compared. Baroreflex mediated changes in heart rate (HR) were elicited by raising and lowering the systemic blood pressure with intravenous injections of phenylephrine and sodium nitroprusside, respectively. The effects of three CCBs, verapamil, diltiazem and nifedipine administered either intravenously (i.v.) or intracisternally (i.c.) were studied. Verapamil administration markedly inhibited the reflex bradycardia as well as the tachycardia following either i.v. or i.c. administration. Intracisternally, a relatively smaller dose of verapamil produced an effect comparable in magnitude and duration, to a higher i.v. dose. The reflex bradycardia was inhibited following i.v., but not i.c. administration of nifedipine while the reflex tachycardia was not affected significantly by either i.v. or i.c. nifedipine. Intravenous diltiazem did not appear to affect the reflex bradycardia or tachycardia significantly. It is suggested that verapamil administration interacts with central cardiovascular integrating mechanisms to reduce the gain of the baroreflex function. Nifedipine and diltiazem are relatively free from this effect. PMID- 10860643 TI - Mitochondrial COI sequences of brachiopods: genetic code shared with protostomes and limits of utility for phylogenetic reconstruction. AB - A 1230-bp region of the cytochrome c oxidase subunit I (COI) gene of mitochondrial DNA of each of 16 brachiopod species, representing all five living orders, was amplified by polymerase chain reaction and sequenced. Pairwise comparisons of sequence differences plotted against divergence times estimated from the brachiopod fossil record revealed that, although there are considerable variations in the expected substitution rate among different lineages, amino acid substitutions of the COI sequences may largely become saturated in 100 Ma, due mostly to multiple substitutions at the same site. Coinciding with this result, phylogenetic analysis indicated low bootstrap values for nodes corresponding to divergence events that occurred before 100 Ma, suggesting that COI sequences are suitable only for inference of phylogenetic events subsequent to the Mesozoic. Examination of brachiopod codons corresponding to invariant amino acids in the COI of various other animals suggest the nonuniversal codon relationships UGA = Trp, AUA = Met, AAA/G = Lys, and AGA/G = Ser. These are identical to those in mollusks, annelids, and arthropods, consistent with the conclusion that brachiopods are protostomes, as indicated by previous molecular analyses. PMID- 10860644 TI - Molecular phylogenetic analysis of the Pneumoroidea (Orthoptera, Caelifera): molecular data resolve morphological character conflicts in the basal acridomorpha. AB - A key transition in the evolution of the insect suborder Caelifera (Orthoptera; Insecta) was from predominantly non-angiosperm-feeding basal lineages to the modern acridomorph fauna (grasshoppers and related insects). However, because of conflicts in the distribution of several complex morphological characters, the relationships of the presumed intermediates, and in particular of the superfamily Pneumoroidea, are presently unclear. We undertook a phylogenetic study of representatives of all of the transitional acridomorph families using mitochondrial and nuclear DNA sequences. No support for pneumoroid monophyly was obtained from nonparametric bootstrap analysis. Furthermore, adopting a maximum likelihood approach, specific hypotheses of relationships within the Pneumoroidea were firmly rejected using parametric bootstrapping and Kishino-Hasegawa tests. The results indicate that the Pneumoroidea are at best a grade. This distinction implies that the evolution of the proposed pneumoroid synapomorphies, femoro abdominal stridulation and simple male genital structure, might previously have been misinterpreted as cases of single character gains or losses within lineages. Reconstructions of character states for the femoro-abdominal stridulation indicate that, in fact, multiple losses or gains are equally likely. An important implication of our findings is that, in grasshoppers, auditory tympana may have evolved before stridulation, supporting the argument that the original function of tympana may have been related not to conspecific communication but to predator detection. Overall, the results of this study emphasize the high information content of these minor groups (in this case, the four intermediate families under consideration contain only 0.2% of extant orthopteran species diversity). Our analyses also demonstrate the advantages of model-based methods in analyzing systematic problems and, in particular, of the importance of testing specific phylogenetic hypotheses when a priori support for groupings (e.g., from nonparametric bootstrapping) is marginal. PMID- 10860645 TI - Rapidly evolving lineages impede the resolution of phylogenetic relationships among Clitellata (Annelida). AB - The phylogenetic relationships of the Clitellata were investigated using a data set with published and new complete or partial 18S rRNA and mtCOI gene sequences of 13 and 49 taxa representing 8 and 14 families, respectively. Three different alignments were considered for 18S, and the possible influence of departures from rate constancy among sites was evaluated by analyses using a Gamma model of rate heterogeneity. Maximum-likelihood estimates of the shape parameter alpha of the Gamma distribution were very low, whatever the alignment or the gene considered, suggesting that phylogenetic reconstructions taking into account the rate heterogeneity among sites are likely to be the most reliable. Analyzed separately, the two genes did not resolve the relationships among the Clitellata, but the consensus tree was congruent with the morphology-based relationships. Our data suggest the inclusion of the Euhirudinea, Acanthobdellida, and Branchiobdellida in the Oligochaeta and suggest the Lumbriculidae as the link between both assemblages. Although separate analyses of both genes, as well as different alignments for the 18S rRNA sequences, yielded conflicting results concerning the phylogenetic position of leeches and leech-like worms vis-a-vis the Oligochaeta, subsequent analyses using the Gamma model greatly reduced the observed inconsistencies. Our analyses show that among the Clitellata, the leeches and the leech-like and gutless worms represent significantly faster evolving lineages. It is suggested that the observed higher mutation rates may be explained by the fact that these lineages contain almost exclusively commensal and/or parasitic taxa. PMID- 10860646 TI - Evolution in the high Andes: the phylogenetics of Muscisaxicola ground-tyrants. AB - Phylogenetic relationships within the genus Muscisaxicola, a primarily Andean group of tyrant-flycatchers, were studied using complete sequences of the mitochondrial genes COII and ND3. Relationships among Muscisaxicola species were found to differ substantially from those of previous views, suggesting convergence in traditional avian taxonomic characters within the genus. The 11 species of large, gray, "typical" Muscisaxicola flycatchers (including M. grisea, newly restored to species status) formed a distinct clade, consisting of two major groups: a clade of 6 species breeding primarily in the central Andes and a clade of 5 species breeding primarily in the southern Andes. The other 2 species traditionally placed in this genus, M. fluviatilis, an Amazonian species, and M. maculirostris, were both rather divergent genetically from the typical species, although M. maculirostris may be the sister taxon to the typical clade. The patterns of sympatry exhibited by Muscisaxicola species in the high Andes appear to be the consequence of speciation and secondary contact within regions of the Andes, rather than a result of dispersal between regions. Diversification of the typical Muscisaxicola species appears to have occurred during the middle and late Pleistocene, suggesting generally that taxa of the high Andes and Patagonia may have been greatly influenced by mid-to-late Pleistocene events. There were likely several independent developments of migration within this genus, but migration is probably ancestral in the southern clade, with subsequent loss of migration in two taxa. PMID- 10860647 TI - Phylogenetic analysis of Tubificidae (Annelida, Clitellata) based on 18S rDNA sequences. AB - Tubificids are aquatic clitellate worms, but recent analyses of morphological characters suggested that this family, as currently recognized, is paraphyletic. Sequences of the 18S rDNA gene of 40 protostome worm species (including 13 representatives of the Tubificidae) and 2 mollusc species were cladistically analyzed to test the monophyly of the Tubificidae and that of some of its constituent subfamilies. Under all alignments tested, the same general phylogenetic pattern emerged. The data support the idea that the Naididae, another clitellate taxon, is associated with some "rhyacodriline" groups within the Tubificidae. The data also corroborate the idea that the Tubificinae and the Limnodriloidinae are monophyletic but indicate that the Rhyacodrilinae and the Phallodrilinae are not. Bathydrilus does not appear to be closely related to other "phallodriline" genera. PMID- 10860648 TI - Resolving phylogeny at the family level by mitochondrial cytochrome oxidase sequences: phylogeny of carrion beetles (Coleoptera, Silphidae). AB - We investigated the phylogenetic relationships of carrion beetles (Coleoptera, Silphidae) using 2094 bp of their mitochondrial cytochrome oxidase subunit I and II and tRNA leucine gene sequences. Shorter fragments of this gene region previously have been used to establish generic relationships in insects. In this study, they provided more than sufficient resolution, although the third positions of the protein-coding sequences reached saturation for the deeper divergences. This first published phylogeny for the Silphidae comprises 23 species from 13 genera sampled across the geographic range of the family. In addition, we included species from three related families as outgroups. One of these families, the Agyrtidae, was, until recently, included in the Silphidae, but its resolution here justifies its current position as a separate family. The silphid subfamilies Nicrophorinae and Silphinae are monophyletic in all analyses. All genera for which several species were sampled are supported as monophyletic groups, with the exception of the genus Silpha. European and North American representatives of two Nicrophorus species described from both continents are supported as each others' closest relatives. The lineage that colonized Gondwanaland and that most likely originated in the Palearctic is the most basal within the Silphinae. PMID- 10860649 TI - Phylogeny of the lady fern group, tribe Physematieae (Dryopteridaceae), based on chloroplast rbcL gene sequences. AB - Nucleotide sequences of the chloroplast gene rbcL from 42 species of the fern tribe Physematieae (Dryopteridaceae) were analyzed to gain insights into the inter- and intrageneric relationships and the generic circumscriptions in the tribe. The phylogenetic relationships were inferred using the neighbor-joining and maximum-parsimony methods, and both methods produced largely congruent trees. These trees reveal that: (1) Athyrium, Cornopteris, Pseudocystopteris, and Anisocampium form a clade and Athyrium is polyphyletic; (2) Deparia sensu lato is monophyletic and Dictyodroma formosana is included in the Deparia clade; (3) Diplaziopsis forms a clade with Homalosorus, which is isolated from the other genera of the Physematieae; (4) Monomelangium is included in the monophyletic Diplazium clade; and (5) Rhachidosorus is not closely related to either Athyrium or Diplazium. PMID- 10860650 TI - One species or several? Discordant patterns of geographic variation between allozymes and mtDNA sequences among spiders in the genus Metepeira (Araneae: Araneidae). AB - Paradoxically, an allozyme study of Metepeira "spinipes" (sensu lato) demonstrated extensive gene flow among four populations whose members are nevertheless morphologically and behaviorally distinct. Initially, the authors tentatively concluded that the populations exhibited panmixis and suggested that local environmental effects accounted for the apparent morphological and behavioral differences. However, they later concluded that such differences were too great to be accounted for by the environment alone and that the four populations actually represented three different species. To confirm that the allozyme results were, in fact, artifactual, we reexamined the relationships among these populations by sequencing a portion of the 12S mtDNA ribosomal subunit. In contrast to the allozyme result, our results demonstrate good agreement between patterns of genetic and morphological/behavioral variation. We suggest (1) that the allozyme allele frequencies are homogenized by balancing selection, not gene flow as was previously concluded, and therefore (2) that this study provides another instance in which inferences about population structures from allozyme data are misleading. PMID- 10860651 TI - Ribosomal DNA evidence and disjunctions of western American Portulacaceae. AB - Phylogenetic analysis of ribosomal DNA internal transcribed spacer sequences from 35 members of western American Portulacaceae plus seven Portulacaceae outgroups generally supports morphologically based interpretations of multiple intercontinental disjunctions. The data neither support nor refute monophyly of the western American group but strongly support a group comprising the western American taxa plus Phemeranthus, the only strictly American genus of the morphology-based eastern American/African group of Portulacaceae, along with the Australian genus Parakeelya. Support is strong for the monophyly of Calandrinia, Montiopsis, Lewisia, Claytonia, and Montia, along with a sister relationship of the last two. The data neither strongly support nor refute the morphologically based diagnosis of Cistanthe, but they strongly support a clade including the North American Cistanthe section Calyptridium and the South American Cistanthe sections Amarantoideae and Philippiamra. The internal transcribed spacer data fail to resolve the phylogenetic relationships among most of the western American lineages, suggesting either rapid radiation or, alternatively, erratic evolution of the internal transcribed spacer. The internal transcribed spacer and morphological evidence together suggest that in this group there have been 8-13 dispersal and colonization events across >2000 km (1 for every 15-26 extant species in this group). The internal transcribed spacer data document complex molecular evolutionary patterns, including strong substitution biases, among-site rate heterogeneity, positional bias for deamination-type substitutions, nonstationarity, and variable rates of insertion/deletion. Our phylogenetic conclusions, however, do not appear to be sensitive to these patterns. PMID- 10860652 TI - Higher-level phylogeny of the Therevidae (Diptera: insecta) based on 28S ribosomal and elongation factor-1 alpha gene sequences. AB - Therevidae (stilleto flies) are a little-known family of asiloid brachyceran Diptera (Insecta). Separate and combined phylogenetic analyses of 1200 bases of the 28S ribosomal DNA and 1100 bases of elongation factor-1alpha were used to infer phylogenetic relationships within the family. The position of the enigmatic taxon Apsilocephala Krober is evaluated in light of the molecular evidence. In all analyses, molecular data strongly support the monophyly of Therevidae, excluding Apsilocephala, and the division of Therevidae into two main clades corresponding to a previous classification of the family into the subfamilies Phycinae and Therevinae. Despite strong support for some relationships within these groups, relationships at the base of the two main clades are weakly supported. Short branch lengths for Australasian clades at the base of the Therevinae may represent a rapid radiation of therevids in Australia. PMID- 10860653 TI - Phylogenetic relationships, character evolution, and biogeography of the subfamily Lygosominae (Reptilia: Scincidae) inferred from mitochondrial DNA sequences. AB - Phylogenetic relationships among the lygosomine skinks were inferred from 1249 base positions of mitochondrial DNA sequences of 12S and 16S rRNA genes. The monophyly of this subfamily was confirmed and the presence of five distinct infrasubfamilial lineages detected. Of these, the Sphenomorphus group appears to have diverged first, followed by the Lygosoma and Egernia groups in order, leaving the Eugongylus and Mabuya groups as sister groups. Our results did not support monophyly of the Mabuya group sensu lato (i.e., an assemblage of the Lygosoma, Egernia, and Mabuya groups), for which a number of morphological and karyological studies demonstrated a considerable similarity. Our results also contradict the previous hypothesis, formulated on the basis of morphological and immunological data, which argued for the sister relationship between the Egernia and the Eugongylus groups. Morphological and karyological characters used to define the Mabuya group (sensu lato) may actually represent plesiomorphic states. The phylogenetic diversity of lygosomine skinks in the Australian region appears to have increased through multiple colonizations from Southeast Asia. PMID- 10860654 TI - Timing the eastern Asian-eastern North American floristic disjunction: molecular clock corroborates paleontological estimates. AB - Sequence data of the chloroplast gene rbcL were used to estimate the time of the well-known eastern Asian-eastern North American floristic disjunction. Sequence divergence of rbcL was examined for 22 species of 11 genera (Campsis, Caulophyllum, Cornus, Decumaria, Liriodendron, Menispermum, Mitchella, Pachysandra, Penthorum, Podophyllum, and Phryma) representing a diverse array of flowering plants occurring disjunctly in eastern Asia and eastern North America. Divergence times of putative disjunct species pairs were estimated from synonymous substitutions, using rbcL molecular clocks calibrated for Cornus. Relative rate tests were performed to assess rate constancy of rbcL evolution among lineages. Corrections of estimates of divergence times for each species pair were made based on rate differences of rbcL between Cornus and other species pairs. Results of these analyses indicate that the time of divergence of species pairs examined ranges from 12.56 +/- 4.30 million years to recent (<0.31 million years), with most within the last 10 million years (in the late Miocene and Pliocene). These results suggest that the isolation of most morphologically similar disjunct species in eastern Asia and eastern North America occurred during the global climatic cooling period that took place throughout the late Tertiary and Quaternary. This estimate is closely correlated with paleontological evidence and in agreement with the hypothesis that considers the eastern Asian eastern North American floristic disjunction to be the result of the range restriction of a once more or less continuously distributed mixed mesophytic forest of the Northern Hemisphere that occurred during the late Tertiary and Quaternary. This implies that in most taxa the disjunction may have resulted from vicariance events. However, long-distance dispersal may explain the disjunct distribution of taxa with low divergence, such as Menispermum. PMID- 10860655 TI - Phylogenetic utility of the nuclear gene dopa decarboxylase in noctuoid moths (Insecta: Lepidoptera: noctuoidea). AB - Phylogenetic utility for the nuclear gene encoding dopa decarboxylase (DDC), little used in systematics, was recently demonstrated within the noctuid moth subfamily Heliothinae. Here we extend the test of the utility of a 709-bp DDC fragment to deeper levels, analyzing 49 species representing major groups across the superfamily Noctuoidea. Parsimony, distance, and maximum-likelihood analyses recover all or nearly all of a set of "test clades" supported by clear morphological synapomorphies, spanning a wide range of taxonomic levels. DDC also upholds a recent proposal that the Noctuidae are paraphyletic. Nt3 contributes a majority of the signal and recovers the basal split between Notodontidae and all other noctuoids, despite a plateau of nt3 divergence at this level. However, nonsynonymous changes also support groups at all levels, and in contrast to nt3, amino acid divergence shows no plateau. The utility of DDC promises to extend back to the early Tertiary and Cretaceous, a time span for which few suitable genes have been identified. PMID- 10860656 TI - Mitochondrial DNA variation among worldwide populations of gypsy moths, Lymantria dispar. AB - Gypsy moth populations from Japan, mainland Asia, Europe, Tunisia, and North America were analyzed for variation in mitochondrial DNA (mtDNA) sequences from three gene regions. These samples resolve into four groups, representing gypsy moths from (1) Okinawa, Japan, (2) Hokkaido, Japan, (3) Honshu and Kyushu, Japan and mainland Asia, and (4) Europe, Tunisia, and North America. Some patterns of geographic variation observed for mtDNA (for example, the distinctiveness of gypsy moths from Hokkaido, Japan) coincide with those observed by Goldschmidt from analyses of morphology, life history, and intersexuality. Other patterns (relative sequence homogeneity across Asia, Honshu, and Kyushu and reduced levels of variation in mainland Japan) do not. PMID- 10860657 TI - First comprehensive low-density horse linkage map based on two 3-generation, full sibling, cross-bred horse reference families. AB - Two 3-generation full-sibling reference families have been produced and form a unique resource for genetic linkage mapping studies in the horse. The F(2) generations, now comprising 61 individuals, consist of 28- to 32-day-old embryos removed nonsurgically from two pairs of identical twin mares. The same stallion sired all F(2)s such that the two full-sibling families are half-sibling with respect to each other. The families are crossbred to maximize levels of heterozygosity and include Arabian, Thoroughbred, Welsh Cob, and Icelandic Horse breeds. Milligram quantities of DNA have been isolated from each embryo and from blood samples of the parents and grandparents. The families have been genotyped with 353 equine microsatellites and 6 biallelic markers, and 42 linkage groups were formed. In addition, the physical location of 85 of the markers is known, and this has allowed 37 linkage groups to be anchored to the physical map. The inclusion of dams in the genotyping analysis has allowed the generation of a genetic map of the X chromosome. Markers have been assigned to all 31 autosomes and the X chromosome. The average interval between markers on the map is 10.5 cM, and the linkage groups collectively span 1780 cM. The results demonstrate the benefits for horse linkage mapping studies of genotyping on these unique full sibling families, which comprise relatively few individuals, by the generation of a comprehensive low-density map of the horse genome. PMID- 10860658 TI - Flow cytometry-based minisequencing: a new platform for high-throughput single nucleotide polymorphism scoring. AB - Single-nucleotide polymorphisms (SNPs) are the most abundant type of human genetic variation. These variable sites are present at high density in the genome, making them powerful tools for mapping and diagnosing disease-related alleles. We have developed a sensitive and rapid flow cytometry-based assay for the multiplexed analysis of SNPs based on polymerase-mediated primer extension, or minisequencing, using microspheres as solid supports. The new method involves subnanomolar concentrations of sample in small volumes ( approximately 10 microl) which can be analyzed at rates of one sample per minute or faster, without a wash step. Further, genomic analysis using multiplexing microsphere arrays (GAMMArrays), enables the simultaneous analysis of dozens, and potentially hundreds of SNPs per sample. We have tested the new method by genotyping the Glu69 variant from the HLA DPB1 locus, a SNP associated with chronic beryllium disease, as well as HLA DPA1 alleles using the multiplexed method. The results demonstrate the sensitivity and accuracy of flow cytometry-based minisequencing, a powerful new tool for genome- and global-scale SNP analysis. PMID- 10860659 TI - An expressed sequence tag database of T-cell-enriched activated chicken splenocytes: sequence analysis of 5251 clones. AB - The cDNA and gene sequences of many mammalian cytokines and their receptors are known. However, corresponding information on avian cytokines is limited due to the lack of cross-species activity at the functional level or strong homology at the molecular level. To improve the efficiency of identifying cytokines and novel chicken genes, a directionally cloned cDNA library from T-cell-enriched activated chicken splenocytes was constructed, and the partial sequence of 5251 clones was obtained. Sequence clustering indicates that 2357 (42%) of the clones are present as a single copy, and 2961 are distinct clones, demonstrating the high level of complexity of this library. Comparisons of the sequence data with known DNA sequences in GenBank indicate that approximately 25% of the clones match known chicken genes, 39% have similarity to known genes in other species, and 11% had no match to any sequence in the database. Several previously uncharacterized chicken cytokines and their receptors were present in our library. This collection provides a useful database for cataloging genes expressed in T cells and a valuable resource for future investigations of gene expression in avian immunology. A chicken EST Web site (http://udgenome. ags.udel. edu/chickest/chick.htm) has been created to provide access to the data, and a set of unique sequences has been deposited with GenBank (Accession Nos. AI979741 AI982511). Our new Web site (http://www. chickest.udel.edu) will be active as of March 3, 2000, and will also provide keyword-searching capabilities for BLASTX and BLASTN hits of all our clones. PMID- 10860660 TI - Significant evidence for linkage of mite-sensitive childhood asthma to chromosome 5q31-q33 near the interleukin 12 B locus by a genome-wide search in Japanese families. AB - Childhood-onset asthma is frequently found in association with atopy. Although asthmatic children may develop IgE antibodies against variety of allergens, asthma is associated primarily with allergy to house-dust mites, molds, or other allergens. In this study, we conducted a genome-wide linkage search in 47 Japanese families (197 members) with more than two mite-sensitive atopic asthmatics (65 affected sib-pairs) using 398 markers. Multipoint linkage analysis was carried out for atopic asthma as a qualitative trait using the MAPMAKER/SIB program. We observed significant evidence for linkage with maximum lod scores (MLS) of 4.8 near the interleukin 12 B gene locus on chromosome 5q31-q33. In addition, suggestive evidence on 4q35 with MLS = 2.7 and on 13q11 with MLS = 2.4 was obtained. The other possible linkage regions included 6p22-p21.3 (MLS = 2.1), 12q21-q23 (MLS = 1.9), and 13q14.1-q14.3 (MLS = 2.0). Many of the linkage loci suggested in this study were at or close to those suggested by genome-wide studies for asthma in Caucasian populations. The present study suggests the contribution of the interleukin 12 B gene or nearby gene(s) to mite-sensitive atopic asthma and a considerable number of genetic variants common across Caucasians and Japanese populations contributing to asthma, although the relative importance of various variants may differ between the groups. PMID- 10860661 TI - Sequence-ready BAC contig, physical, and transcriptional map of a 2-Mb region overlapping the mouse chromosome 6 host-resistance locus Cmv1. AB - The host-resistance locus Cmv1 controls viral replication of mouse cytomegalovirus (MCMV) in the spleen of infected mice. Cmv1 maps on distal chromosome 6, very tightly linked to the Ly49 gene family within a 0.35-cM interval defined proximally by Cd94/Nkg2d and distally by D6Mit13/D6Mit111/D6Mit219/Prp/Kap. To facilitate the cloning of the gene, we have created a high-resolution physical map of the Cmv1 genetic interval that is based on long-range restriction mapping by pulsed-field gel electrophoresis, fluorescence in situ hybridization analysis of interphase nuclei, and the assembly of a cloned contig. A contig of BAC and YAC clones was assembled using probes derived from the minimal genetic interval. Individual clones from the region were validated by (1) restriction digest fingerprinting, (2) STS content mapping, (3) Southern hybridizations, and (4) sequencing and mapping of clone ends. This contig contains 25 YACs anchored by 71 STSs and 73 BACs anchored by 40 STSs. We also report the cloning of 31 new STSs and 18 new polymorphic markers. A minimum tiling path was defined that consists of either 4 YACs or 13 BACs covering 1.82 Mb between D6Ott8, the closest proximal marker, and D6Ott115, the closest distal marker. Gene distribution in the region includes 14 Ly49 genes as well as 3 new additional transcripts. This high-resolution, sequence-ready BAC contig provides a backbone for the identification of Cmv1 and its relationship with genes involved in innate immunity. PMID- 10860662 TI - cDNA cloning, genomic structure, and chromosomal localization of three members of the human fatty acid desaturase family. AB - The insertion of double bonds into specific positions of fatty acids is achieved by the action of distinct desaturase enzymes. Here we report the cloning and characterization of three members of the fatty acid desaturase (FADS) gene family in humans. Initially identified as cDNA fragments by direct cDNA selection within a defined 1.4-Mb region in 11q12-q13.1, full-length fatty acid desaturase-1 (FADS1) and fatty acid desaturase-2 (FADS2) transcripts were obtained by EST sequence assembly. A third member, fatty acid desaturase-3 (FADS3), was identified in silico revealing 62 and 70% nucleotide sequence identity with FADS1 and FADS2, respectively. The three genes are clustered within 92 kb of genomic DNA located 2 kb telomeric to FEN1 and 50 kb centromeric to VMD2 and are likely to have arisen evolutionarily from gene duplication as they share a remarkably similar exon/intron organization. Protein database searches identified FADS1, FADS2, and FADS3 as fusion products composed of an N-terminal cytochrome b5-like domain and a C-terminal multiple membrane-spanning desaturase portion. PMID- 10860663 TI - Mouse RNA helicase II/Gu: cDNA and genomic sequences, chromosomal localization, and regulation of expression. AB - RNA helicase II/Gu (RH II/Gu) is a mammalian nucleolar RNA helicase previously identified using an autoimmune serum from a patient with watermelon stomach disease. RH II/Gu can unwind double-stranded RNA and can fold or introduce a secondary structure to a single-stranded RNA. These two enzymatic activities reside in two separate domains of the RH II/Gu molecule. The present study reports the molecular analysis of the cDNA and genomic sequences of the mouse RH II/Gu, its chromosomal localization, and the regulation of expression. The cDNA derived amino acid sequence shows three tandem repeats at the NH(2)-terminal end of the protein, which are not conserved in the human homologue. Each repeat has 37 amino acids that are rich in basic residues. The helicase and foldase domains are highly conserved between the mouse and the human RH II/Gu. The basic promoter region of the mouse RH II/Gu gene is within 300 nucleotides upstream of a putative ATG initiation codon. Upstream of this promoter region is a silencer that represses transcription of the mouse RH II/Gu gene. This inhibitory region contains three 38-nucleotide repeats in tandem. The mouse RH II/Gu consists of 14 exons and 13 introns. The 3' flanking sequence of the gene contains three putative polyadenylation sites but only two sites are probably functional as shown by Northern blot analysis and 3' end sequences of mouse RH II/Gu cDNA in the EST database. These two alternative polyadenylation sites are approximately 240 and 2100 nucleotides from the TGA stop codon. Both mouse and human RH II/Gu genes are localized on chromosome 10. The availability of the mouse RH II/Gu gene will facilitate its functional analysis including creation of a mouse deficient in RH II/Gu protein. PMID- 10860664 TI - Characterization of the human and mouse HEY1, HEY2, and HEYL genes: cloning, mapping, and mutation screening of a new bHLH gene family. AB - Many basic helix-loop-helix (bHLH) transcription factors are known as key regulators of embryonic development or differentiation in various species. We have isolated and characterized three new hairy-related bHLH transcription factor genes from mouse and human (hairy and Enhancer-of-split related with YRPW motif; HEY1, HEY2, and HEYL). All three HEY genes have a similar genomic structure with five exons. Together with a highly related Drosophila homologue, they form a new bHLH gene subfamily that is different from both hairy and the known vertebrate Hes and Her genes. While the overall structure with the bHLH domain, Orange domain, and WRPW motif is similar, the last motif is changed to KPYRPWG in Hey1/2 and absent in HeyL. This and other sequence features suggest Hey proteins to have unique functional properties. The genes were mapped by fluorescence in situ hybridization and RH mapping to the following human chromosomes: (HEY1) 8q21, (HEY2) 6q21, and (HEYL) 1p34.3. Based on expression patterns and map location, HEY genes are candidates for several human or mouse disease loci. However, initial screening of DNA from affected individuals for two human disorders and four mouse mutants did not reveal any diagnostic alterations in the coding regions. PMID- 10860665 TI - cDNA cloning and mapping of mouse pleckstrin (Plek), a gene upregulated in transformation-resistant cells. AB - Changes that occur during tumor promotion, the rate-limiting phase of multistep carcinogenesis, may offer the best targets for prevention of cancer or reversal of early disease. The murine epidermal JB6 promotion-sensitive (P+) and resistant (P-) cell lines provide a cell culture model for tumor promoter-induced neoplastic transformation ideally suited to the identification of molecular events that mediate or inhibit transformation. A differential display comparison of P+ and P- cell mRNAs yielded seven differentially expressed sequences. One of the sequences preferentially expressed in P- cells identified an approximately 3. 6-kb message that was induced to higher levels in P- cells following exposure to the tumor promoter 12-O-tetradecanoylphorbol acetate than in P+ cells. The message was detected in mRNA from heart, lung, and spleen. cDNA cloning of the P- preferential sequence revealed a high degree of identity to human pleckstrin (PLEK), the major PKC substrate in platelets (Tyers et al., 1988, Nature 333: 470). We report the complete mouse cDNA sequence of pleckstrin and the localization of the gene to chromosome 11, its expression in a nonhematopoetic cell line, and its potential role in blocking neoplastic transformation. PMID- 10860666 TI - Identification and gene organization of three novel members of the IL-1 family on human chromosome 2. AB - Members of the IL-1 family of cytokines are important in mediating inflammatory responses. The genes encoding IL-1alpha, IL-beta, and the IL-1 receptor antagonist (IL-1Ra) are clustered within 450 kb on human chromosome 2q. By searching the EST databases and sequencing this region of chromosome 2, we have identified three novel genes that show homology to the IL-1 family, which we have named IL-1-related protein 1, 2, and 3 (IL-1RP1, IL-1RP2, and IL-1RP3). All three genes contain a signature motif common to the IL-1 family and appear to be more closely related to IL-1Ra. Similar to the intracellular form of IL-1Ra, these genes lack conventional hydrophobic signal sequences. The expression of these genes appears to be highly restricted to various epithelial cell populations. Our results demonstrate the existence of additional IL-1 gene family members within the previously defined IL-1 cluster and point to this region of chromosome 2 as an evolutionary hotspot for IL-1 gene duplication. These genes may prove to have an important role in inflammatory responses. PMID- 10860667 TI - Cloning and expression analysis of a novel member of the facilitative glucose transporter family, SLC2A9 (GLUT9). AB - Several lines of evidence suggest the existence of additional members of the mammalian facilitative glucose transporter family. A human cDNA sequence corresponding to a novel member of the glucose transporter family (GLUT1-5) was identified (GLUT9; HGMW-approved symbol SLC2A9), and it encodes a putative transporter of 540 amino acids. The predicted protein has sequence identity of 44 and 38% to Glut5 and Glut1, respectively. Based on hydropathic analysis, the novel transporter's predicted topology consists of 12 transmembrane domains, similar to the other family members. Northern analysis reveals three mRNA species: a major transcript of 1.9 kb and two other transcripts of 3.1 and 5.0 kb, found primarily in kidney and liver, but present at low levels in several other tissues. GLUT9 was localized to chromosome 4 using a monochromosomal human/rodent somatic cell hybrid mapping panel. A portion of the GLUT9 cDNA is represented in a National Center for Biotechnology Information UniGene cluster, which maps to chromosome 4p15.3-p16. PMID- 10860668 TI - Construction of a physical and transcript map flanking the imprinted MEST/PEG1 region at 7q32. AB - MEST/PEG1, a gene expressed paternally in mesodermal derivatives in early embryonic stages, is the first imprinted gene mapped to chromosome 7q32. Since imprinted genes are clustered in general at a chromosomal region, we speculated that a similar imprinted-gene cluster may exist at chromosome region 7q32 and that the functions of some such genes may contribute to the phenotype of Silver Russell syndrome including maternal uniparental disomy for chromosome 7 (maternal UPD7). As an initial step toward the isolation of imprinted genes at 7q32, we adopted an integrated approach involving the construction of a PAC contig and ESTs mapping in the vicinity of MEST. Here, we have constructed a complete contig of PAC and BAC clones and a transcript map spanning the entire approximately 1-Mb region between D7S530 and D7S649. We developed 60 novel STSs and precisely mapped 47 genes/ESTs. This map contains a putative autistic disorder locus that has been suggested to be localized near markers D7S530 and D7S684. This integrated physical and transcript map provides a valuable resource for identification of an imprinted gene(s) in this region as well as a candidate gene(s) for autistic disorder. PMID- 10860669 TI - Chromosomal mapping and genomic organization of an evolutionarily conserved zinc finger gene ZNF277. AB - A novel C2H2 zinc finger gene, ZNF277, has been localized to human chromosome 7q31.1. The gene is encoded by 12 exons in a genomic fragment of >100 kb between the microsatellite markers D7S523 and D7S471, deleted in a number of malignancies. The predicted open reading frame (ORF) of 438 amino acids shows an overall homology of 50% to the putative ORF F46B6.7 of Caenorhabditis elegans. The presence of a 30-amino-acid coiled-coil domain in both the C. elegans ORF F46B6.7 and ZNF277 is suggestive of functional similarities. ESTs for the murine orthologue ZFP277 are found in early embryonic stem cells, 16-cell stage embryo, and blastocysts. The evolutionary conservation and the expression profile suggest ZNF277 to be a critical regulator of development and differentiation. PMID- 10860671 TI - Leptin and related peptides PMID- 10860670 TI - Editor's introduction PMID- 10860672 TI - Understanding the neural control of ingestive behaviors: helping to separate cause from effect with dehydration-associated anorexia. AB - Eating and drinking are motivated behaviors that are made up of coordinated sets of neuroendocrine, autonomic, and behavioral motor events. Although the spinal cord, hindbrain, and hypothalamus contain the motor neurons and circuitry sufficient to maintain the reflex parts of these motor events, inputs from the telencephalon are required to furnish the behavioral components with a motivated (goal-directed) character. Each of these motor events derives from the complex interaction of a variety of sensory inputs with groups of neural networks whose components are distributed throughout the brain and collectively support motor expression and coordination. At a first approximation based on a variety of data, these networks can be divided into three groups: networks that stimulate, those that inhibit, and those that disinhibit motor functions. A fourth contributor is the circadian timing signal that originates in the hypothalamic suprachiasmatic nucleus and provides the temporal anchor for the expression of all behaviors. This article discusses the nature of these networks using neuroanatomical (tract tracing and neuropeptide in situ hybridization), endocrine, and behavioral evidence from a variety of experimental models. A persistent problem when studying the control of food intake from a neural systems perspective has been the difficulty in separating those neuronal changes that result in hunger from those that are as a consequence of eating. To address this problem, dehydration associated anorexia is presented as a particularly useful experimental model because it can be used to distinguish between neural mechanisms underlying anorexia and those changes that occur as a consequence of anorexia. The article concludes by highlighting the potential role of neuropeptidergic action in the operation of these networks, using forebrain neuropeptidergic innervation of the parabrachial nucleus as an example. PMID- 10860673 TI - How tight are your genes? Transcriptional and posttranscriptional regulation of the leptin receptor, NPY, and POMC genes. AB - In the past few years, there has been exponential growth in our knowledge of genes that control food intake and metabolism. Most of this research has demonstrated either an increased or decreased expression of these "obesity genes" in response to changes in nutritional status. Ultimately, these changes reflect modifications in the rate of gene transcription, mRNA stability, translation initiation, or posttranslational processing. Few laboratories have examined specifically which of these molecular mechanisms are responsible for obesity gene regulation, and thus, the field is wide open for exploration. In addition, it is possible that some forms of human obesity may be caused by inherited mutations in transcription factors or other regulatory molecules rather than base pair mutations in the obesity genes themselves. This article focuses on the regulation of the leptin receptor, NPY, and POMC genes, and explores what is known about the regulation of these obesity genes in response to food intake or changes in body fat stores. Connections between regulation of these genes and some inherited forms of human obesity are made. PMID- 10860674 TI - CNS melanocortin system involvement in the regulation of food intake. AB - Accumulating evidence indicates that the central melanocortin (MC) system plays a key role in the regulation of food intake and energy balance. This evidence includes findings that either spontaneous genetic mutations or targeted gene deletions that impair melanocortin signaling cause disrupted food intake and body weight control. In addition, expression of the mRNA that encodes the endogenous agonists and antagonists for CNS melanocortin receptors is regulated by changes in energy balance and body-adiposity signals. Finally, administration of both natural and synthetic ligands to MC receptors produces changes in food intake. The data collectively suggest a critical role for melanocortin signaling in the control of energy balance. PMID- 10860675 TI - Leptin and metabolic control of reproduction. AB - Leptin treatment prevents the effects of fasting on reproductive processes in a variety of species. The mechanisms that underlie these effects have not been elucidated. Progress in this area of research might be facilitated by viewing reproductive processes in relation to mechanisms that maintain fuel homeostasis. Reproduction, food intake, and fuel partitioning can be viewed as homeostatic responses controlled by a sensory system that monitors metabolic signals. These signals are generated by changes in intracellular metabolic fuel availability and oxidation rather than by changes in the amount of body fat or by changes in any aspect of body composition. Leptin might be viewed as either a mediator or as a modulator of the intracellular metabolic signal. Consistent with its purported action as a mediator of the metabolic signal, leptin synthesis and secretion are influenced acutely by changes in metabolic fuel availability, and these changes might lead to changes in reproductive function. The effects of leptin treatment on reproduction are blocked by treatments that inhibit intracellular fuel oxidation. Metabolic signals that inhibit reproduction in leptin-treated animals might act via neural pathways that are independent of leptin's action. Alternatively, both leptin and metabolic inhibitors might interact at the level of intracellular fuel oxidation. In keeping with the possibility that leptin modulates the metabolic signal, leptin treatment increases fuel availability, uptake, and oxidation in particular tissues. Leptin might affect reproduction indirectly by altering fuel oxidation or other peripheral processes such as gastric emptying. Reproductive processes are among the most energetically expensive in the female repertoire. Because leptin increases energy expenditure while simultaneously inhibiting energy intake, it may have limited use as a long term treatment for infertility. PMID- 10860676 TI - Ultrastructural localization of the receptor for leptin in the rat hypothalamus. AB - Ultrastructural localization of the leptin receptor in the rat hypothalamus was studied by immunocytochemistry. The antiserum against the leptin receptor which was used specifically recognized the carboxy terminal of the cytoplasmic domain. Intense leptin receptor immunoreactivity was detected in the arcuate, paraventricular, and ventromedial nuclei of the hypothalamus and in the lateral hypothalamic area. At the ultrastructural level, leptin receptor-like immunoreactivity appeared to be concentrated predominantly in perikarya and dendrites of these areas and strong immunolabeling for the leptin receptor was detected in the plasma membrane, rough endoplasmic reticulum, Golgi apparatus, and cytoplasmic matrix. This study provides the first detailed fine structure of leptin receptor-immunoreactive neurons in the rat hypothalamus. It may help to provide better understanding of the functions of leptin in the rat hypothalamus. PMID- 10860677 TI - Differential distribution and regulation of OX1 and OX2 orexin/hypocretin receptor messenger RNA in the brain upon fasting. AB - To further understand the functions of the orexin/hypocretin system, we examined the expression and regulation of the orexin/hypocretin receptor (OX1R and OX2R) mRNA in the brain by using quantitative in situ hybridization. Expression of OX1R and OX2R mRNA exhibited distinct distribution patterns. Within the hypothalamus, expression for the OX1R mRNA was largely restricted in the ventromedial (VMH) and dorsomedial hypothalamic nuclei, while high levels of OX2R mRNA were contained in the paraventricular nucleus, VMH, and arcuate nucleus as well as in mammilary nuclei. In the amygdala, OX1R mRNA was expressed throughout the amygdaloid complex with robust labeling in the medial nucleus, while OX2R mRNA was only present in the posterior cortical nucleus of amygdala. High levels of OX2R mRNA were also observed in the ventral tegmental area. Moreover, both OX1R and OX2R mRNA were observed in the hippocampus, some thalamic nuclei, and subthalamic nuclei. Furthermore, we analyzed the effect of fasting on levels of OX1R and OX2R mRNA in the hypothalamic and amygdaloid subregions. After 20 h of fasting, levels of OX1R mRNA were significantly increased in the VMH and the medial division of amygdala. An initial decrease (14 h) and a subsequent increase (20 h) in OX1R mRNA levels after fasting were observed in the dorsomedial hypothalamic nucleus and lateral division of amygdala. Levels of OX2R mRNA were augmented in the arcuate nucleus, but remained unchanged in the dorsomedial hypothalamic nucleus, paraventricular hypothalamic nucleus, and amygdala following fasting. The time dependent and region-specific regulatory patterns of OX1R and OX2R suggest that they may participate in distinct neural circuits under the condition of food deprivation. PMID- 10860678 TI - Appearance of a nocturnal peak of leptin secretion in the pubertal rat. AB - Whether leptin is involved in the timing of puberty remains highly controversial in the rat. Daytime leptin secretion shows little change during the transition into adulthood. Because leptin exhibits a diurnal variation in the adult, it is possible that the ontogeny of such a rhythm provides important information for the timing of puberty. To begin to evaluate this hypothesis, we determined the development of the diurnal leptin secretion in the rat. The young females were raised in a light-controlled environment (12L, 0700 h light on). A cannula was placed in the right atrium on the previous day, and blood samples were collected every 4 h on Days 21, 24, 28, 32, 36 (1 day after vaginal opening), and 48 (adult, diestrus of estrous cycle). In addition to vaginal opening, plasma prolactin levels were determined as an endocrine index of puberty. Changes in food intake were monitored because nocturnal food intake has been considered to be a synchronizer for the leptin rhythm. This pattern of food intake was clearly evident throughout the ages studied. By contrast, there was no leptin rhythm at 21 and 24 days of age. Beginning at 28 days, leptin secretion exhibited a significant nocturnal peak (2300 h); this nocturnal peak increased in amplitude at 32 and 36 days and was still apparent in the cycling adult at Day 48. Plasma prolactin did not exhibit a diurnal rhythm but it increased from Days 32 to 48. The present findings indicate that in the rat, both the appearance of the nocturnal leptin rhythm and the nocturnal increase in circulating leptin levels during development carry information for timing the onset of puberty. PMID- 10860679 TI - Changes in leptin levels during lactation: implications for lactational hyperphagia and anovulation. AB - In these studies we investigated the time course of changes in circulating leptin levels in lactating rats and the dependence of these changes on the energetic cost of lactation and evaluated the contribution of changes in leptin levels to lactational hyperphagia and infertility. In the first experiment, plasma leptin levels were measured on Days 5, 10, 15, 20, and 25 postpartum in freefeeding lactating rats and age-matched virgin females. Retroperitoneal and parametrial fat pads weights were obtained from the same females. In the second experiment the same measures, together with plasma insulin and prolactin levels, were taken on Days 15 and 20 postpartum from galactophore-cut and sham-operated females. In Experiments 3 and 4, the effects of exogenous leptin administration, either subcutaneously (sc) or intracerebroventricularly (icv), on lactational anovulation, maternal food intake, and dam and litter weights were examined. Circulating leptin levels decreased in lactating rats. Leptin levels were highly positively correlated with fat pad weight. Eliminating the energetic costs of lactation by preventing milk delivery induced dramatic increases in plasma leptin and insulin levels and also increased adiposity. Exogenous leptin administration did not affect length of lactational anovulation but reduced food intake, maternal body weight, and litter weight gain when given centrally and maternal body weight when given systemically. Together, these data show that the energetic costs of lactation are associated with a fall in circulating leptin levels but that these do not make a major contribution to the suppression of reproduction in lactating rats; however, they may be permissive to the hyperphagia of lactation. PMID- 10860680 TI - Chronic leptin administration in developing rats reduces stress responsiveness partly through changes in maternal behavior. AB - In adult rodents, leptin has been shown to significantly alter the activity of several neuroendocrine functions, including the activity of the hypothalamic pituitary-adrenal (HPA) axis. Leptin is generally believed to be inhibitory to HPA activity in adults. Developing rat pups have high circulating levels of leptin, which begs the question of leptin's physiological role in controlling basal and stress-induced adrenocortical activity in neonatal rats. In this study, we treated rat pups daily from days 2-9 (or 6-10) of life with either vehicle or leptin (1 or 3 mg/kg body wt, ip) and determined the effects on body weight gain, fat pad deposits, and HPA activity in 10-day-old pups. We measured hypothalamic CRF mRNA levels in vehicle- and leptin-treated pups by in situ hybridization and determined plasma ACTH, corticosterone, and leptin concentrations under basal conditions or following exposure to a 3-min ether stress. Because leptin activates sympathetic activity and energy expenditure in adults and possibly also in rat pups, and because litter temperature is an important determinant of maternal behavior, we also investigated whether chronic leptin administration would modify aspects of maternal care that are important for the maintenance of HPA function. Chronic leptin treatment increased circulating levels of leptin and had significant dose-related metabolic effects, including reduced body weight gain and fat pad weight in 10-day-old pups. Basal expression of CRF mRNA in the PVN or secretion of ACTH and corticosterone was not modified by leptin treatment. In contrast, chronically elevated leptin concentrations during the neonatal period significantly lowered CRF expression in the PVN 60 min after stress and reduced the duration of the ACTH response to stress in pups, suggesting that glucocorticoid feedback on the HPA axis might be altered by this treatment. In addition, mothers caring for pups injected with leptin displayed longer bouts of anogenital licking of pups than mothers of vehicle-treated rats. Given that this particular type of pup stimulation has been shown to influence stress responsiveness, it is possible that the maternal response modulates the effects of exogenous leptin treatment. In conclusion, our results demonstrate that the leptin signal is functional during the early developmental period and that leptin can modulate the hormonal response to stress in young rats either by a direct effect on the HPA axis or indirectly through changing some aspects of maternal behavior. PMID- 10860681 TI - Effects of intracerebroventricular leptin administration on feeding and sexual behaviors in lean and obese female Zucker rats. AB - Obese Zucker rats (fa/fa) are characterized by inadequate leptin signaling caused by a mutation in the leptin receptor gene. Obese Zucker females are infertile and hyporesponsive to the inductive effects of ovarian hormones on sexual behaviors. Leptin treatment reverses aspects of reproductive dysfunction due to perturbations in energy balance in other animal models. Our first experiment tested the hypothesis that intracerebroventricular (icv) leptin administration would enhance the display of sexual behaviors in obese Zucker females. A second experiment compared lean and obese Zucker females' responses to leptin, during fed and fasted conditions. Ovariectomized (OVX) Zucker rats were implanted with lateral ventricular cannulae. In Experiment 1, fasted, obese females received estradiol benzoate, progesterone, and icv injections of 3, 18, or 36 microg murine leptin or vehicle. Leptin administration reduced food intake, but did not enhance sexual behaviors. In Experiment 2, steroid-replaced, OVX lean and obese females (from a different source than those in Experiment 1) received icv injections of vehicle or 3 or 36 microg leptin under fed and fasted conditions. Leptin treatment reduced food intake and weight gain in the fed, but not the fasted, condition in both genotypes. Sexual receptivity and locomotion were not affected, but icv leptin injections reduced proceptive behaviors in ad libitum fed rats. These data confirm previous reports that centrally administered leptin decreases food intake and weight gain in obese Zucker rats; results from Experiment 2 suggest that lean and obese females are similarly responsive to these actions of leptin. Contrary to our hypothesis, leptin treatment did not stimulate sexual behaviors; rather, the hormone appears to inhibit the display of sexual proceptivity in ad libitum-fed lean and obese Zucker female rats. PMID- 10860682 TI - Twenty-four-hour profiles of serum leptin in siberian and golden hamsters: photoperiodic and diurnal variations. AB - Serum leptin concentrations were obtained from male Siberian hamsters (Phodopus sungorus) and golden hamsters (a.k.a. Syrian, Mesocricetus auratus) housed on long [light:dark (LD) 16:8] and short (LD 6:18) photoperiods for 10-11 weeks. Blood samples were collected at 45-min intervals for 24 h from individual animals using an in-dwelling atrial catheter. In Siberian hamsters, exposure to short photoperiods as compared to long photoperiods reduced body weight (32.5 +/- 1.5 vs 47.7 +/- 1.1 g) and leptin (24-h mean: 5.3 +/- 0.4 ng/ml vs 18.6 +/- 2.1 ng/ml). Although photoperiod influenced the temporal distribution of leptin in golden hamsters, the main effect of photoperiod on leptin levels in golden hamsters did not reach significance (24-h mean: 7.1 +/- 1.0 ng/ml vs 5.1 +/- 0.8 ng/ml.). Body weights of golden hamsters did not vary significantly following exposure to short photoperiod for 11 weeks (178.3 +/- 3.6 g in LD 6:18 vs 177.8 +/- 7.3 g in LD 16:8). There was no nocturnal increase in serum leptin in either species. Marked interindividual differences were apparent in individual leptin profiles. Periodogram analysis revealed that only a few animals exhibited 24-h periodicities; the presence of a significant 24-h periodicity was more common in hamsters exposed to short days. Photoperiod-associated differences in the 24-hour profile of leptin secretion may be the result of photoperiod-associated changes in feeding behavior or metabolism. A full understanding of the regulation of leptin secretion in multiple time domains may enhance our understanding of the function of leptin. PMID- 10860683 TI - Fathers, fat, and maternal energetics in a biparental hamster: paternal presence determines the outcome of a current reproductive effort and adipose tissue limits subsequent reproductive effort. AB - Djungarian hamster females, Phodopus campbelli, are severely constrained in their ability to reproduce successfully and lose 20% of their body weight by the time pups are weaned. In the wild and in the laboratory, biparental care improves maternal reproductive success. Two experiments quantified the effects of paternal presence and partial lipectomy [surgical depletion of parametrial white adipose tissue (PWAT) on day 8 of the 18-day gestation] on maternal energy balance, reproductive success, and investment in a subsequent reproductive attempt. Paired females reproduced successfully, maintained body weight, and invested in a second litter. Removal of the male decreased pup survival, growth, and readiness for dispersal by 18 days of age. Solitary females lost 10% of their body weight by the birth and a further 10% by day 18 after the birth. Thus, paternal presence balanced maternal energy budgets during reproduction and prevented a 20% loss in body weight. Equivalent weight loss occurs in response to other maternal stressors, therefore 20% may be the maximum tolerable weight loss in this species. Fresh weight of interscapular brown adipose tissue was predicted by the extent of maternal hyperthermia but not by maternal energy balance or lipectomy. Partial lipectomy did not adversely affect the female or the first litter but decreased the probability of investment in a second reproductive attempt and halved the size of the second litter. This effect may have been due to the 0.1% of body weight amount of lipid removed or may reflect a specialized role for PWAT in adjusting maternal investment. PMID- 10860684 TI - The cost of hospital-acquired infection and the value of infection control. PMID- 10860685 TI - Do antimicrobials have a role in preventing septicaemia following instrumentation of the urinary tract? AB - Urinary tract instrumentation is a significant cause of septicaemia. Review of the literature suggests that selective use of antimicrobials would reduce the risk of septicaemia as this varies between patients and with procedures. Antimicrobial prophylaxis is indicated for patients at high risk of endocarditis, or who are neutropenic. For patients without these risk factors, it is indicated for open, transurethral, or certain forms of laser prostatectomy or trans-rectal prostate biopsy. For cystoscopy, antimicrobials are indicated for patients with preoperative bacteriuria or a preoperative indwelling catheter. Single dose aminoglycosides or oral fluoroquinolones are the agents of choice with the exception of the prevention of endocarditis, where combinations active against streptococci are recommended. For other instrumentations, the risk of antimicrobial toxicity probably outweighs the benefits and a risk-reduction strategy is recommended. Further studies are required to provide definitive answers in many of these areas. PMID- 10860686 TI - Nosocomial pulmonary infection by antimicrobial-resistant bacteria of patients hospitalized in intensive care units: risk factors and survival. AB - The objectives of this study were to identify the risk factors of nosocomial pulmonary infection (NPI) in intensive care units (ICUs) associated with antimicrobial-resistant bacteria (NPI-ARB) and to compare survival after NPI-ARB with NPI due to antimicrobial-sensitive bacteria (NPI-ASB). We analysed data from a surveillance network monitoring nosocomial infections in 27 mixed ICUs in the south-east of France. NPI surveillance data were recorded for 628 patients with documented NPI. The patients were stratified into 2 groups by type of pneumonia: NPI-ASB (445 patients) vs. NPI-ARB (183 patients). Variables associated with NPI ARB were identified++ by multivariate logistic regression. Survival was calculated using the Kaplan-Meier method. A medical condition for ICU admission [odds ratio (OR) 1.98, 95% confidence interval (95% CI) 1.35-2.91], transfer from another hospital ward [OR 1.66, 95% CI (1.14-2.42)], a colonized central venous catheter [OR 3.47, 95% CI (1.46-8.21)], a stay of [eight days [OR 1.02, 95% CI (1.01-1. 05)] and mechanical ventilation [OR 2.10, 95% CI (1.31-3.36)] were independent risk factors of NPI-ARB. Median survival was 35 days after NPI-ARB and 32 days after NPI-ASB (P=0.92). Survival after bacterial NPI was not associated with antimicrobial susceptibility. PMID- 10860687 TI - Role of infection control measures in limiting morbidity associated with multi resistant organisms in critically ill patients. AB - A retrospective comparative study was performed to determine the impact of infection control measures (ICMs) on colonization and infections due to methicillin-resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae (producing transferable extended-spectrum beta-lactamase, KPESBL), and multi resistant Enterobacter aerogenes (MREA) in intensive care unit patients. Infection Control Measures included surveillance cultures, isolation procedures and mupirocin for MRSA nasal carriage. The numbers of patients infected and/or colonized by MRSA, KPESBL or MREA were compared during two consecutive one-year periods (Period 1 before ICMs, and Period 2 after ICMs). The antibiotic consumption during the two periods was analysed. In Period 1 and Period 2, respectively, the rate of patients infected or colonized by at least one of the three organisms was 15% and 6.8% (P=0.001); by MRSA 7.7% and 2.6% (P=0. 004); by KPESBL 1.7% and 0% (P=0.25); and by MREA 5.6% and 4.3% (P=0. 47). During Period 2, there was a clear-cut decrease in the percentage of patients infected by MRSA (P=0.018), a non-significant decrease in those infected by KPESBL (P=0.06), and no decrease in patients infected by MREA (P=0.22). When calculated per 1000 patient-days, for Period 1 and Period 2, respectively, the rate of patients infected or colonized by at least one of the three organisms was 11.9 and 8.8; for MRSA it was 4 and 2.2; for KPESBL it was 1 and 0; and for MREA it was 4 and 4. Antibiotic cost was pound98.7 in Period 1 and pound62.7 in Period 2. ICMs contributed to the control of infections and colonizations due to MRSA and KPESBL but not those due to MREA. PMID- 10860688 TI - Randomized multi-centre trial of the effects of a catheter coated with hydrogel and silver salts on the incidence of hospital-acquired urinary tract infections. AB - Catheters coated with hydrogel and silver salts have been proposed to prevent hospital-acquired urinary tract infections (UTI). We carried out a randomized, prospective, double-blind multi-centre trial to compare those catheters with classical urinary tract catheters. We included in the study 199 patients requiring urethral catheterization for more than three days: 109 in group 1 (classical catheter) and 90 in group 2 (catheter coated with hydrogel and silver salts). Urine from the patients was tested for 10 days after the insertion of the catheter (reactive dipsticks each day and diagnostic urinalysis every two days). The UTI associated with catheterization was defined on the basis of bacterial and cytological criteria (>10(5)cfu bacteria per mL and >10 leucocytes per mm(3)). Twenty-two UTIs were recorded: 13 in group 1 and nine in group 2. The cumulative incidence of UTI associated with catheterization was 11.1% overall, 11.9% for group 1 and 10% for group 2; the odds ratio was 0.82 (95% confidence interval: 0.30 to 2. 20); the cumulative incidence for UTI, calculated by the Kaplan-Meier method was 36.3 overall, 35.2 in group 1 and 36.0 in group 2; the overall incidence density was 19 per thousand days of catheterization, 21 in group 1 and 18 in group 2. The differences between the two groups were not significant. Overall, we feel that there is not enough evidence to conclude that catheters coated with silver salts and hydrogel give greater protection than classical catheters and to recommend widespread use. PMID- 10860689 TI - Clinical and molecular epidemiology of hospital Enterococcus faecium isolates in eastern France. Members of Reseau Franc-Comtois de Lutte contr les Infections Nosocomiales. AB - We carried out a surveillance study of Enterococcus faecium isolates in the Franche-Comteregion of France over three years. Clinical and epidemiological strains were characterized by antibiotype and genotype (pulsed field gel electrophoresis, PFGE). Three case-control studies were performed to identify risk factors for colonization/infection with three defined resistant phenotypes (amoxycillin, high-level gentamicin and high-level kanamycin). The crude incidence of colonization/infection was 0.156%, and 68.8% of cases were classified as hospital-acquired. Incidence did not differ according to the type of hospitalization (middle term or acute care). The urinary tract was the major site of infection. Resistance rates were: 45.8% (amoxycillin), 18.7% (high-level gentamicin), 61.4% (high-level kanamycin) and 3.1% (vancomycin). No isolate produced b-lactamase and one isolate carried the vanA gene. PFGE revealed two major epidemic patterns each including resistant strains isolated in different hospitals and during different periods in the study. Previous antimicrobial treatment was not identified as a risk factor for colonization/infection with any resistant phenotype. Despite the low frequency of vancomycin-resistant isolates in this study, resistant strains were widely disseminated and had characteristics enabling them to persist and spread. If these strains acquired the vanA gene, the risk of an outbreak would be large. So, the prevalence of vancomycin-resistant E. faecium in hospitals should be carefully monitored in the future. PMID- 10860690 TI - Nosocomial outbreak of ampicillin resistant Enterococcus faecium: risk factors for infection and fatal outcome. AB - A nosocomial outbreak caused by ampicillin resistant Enterococcus faecium (ARE) was detected at a Norwegian university hospital in January 1995. Prior to this outbreak, ARE were not common in this hospital or other hospitals in Norway. During 1995 and 1996, a total of 149 cases with clinical ARE infection were detected prospectively. A case control study was performed by allocating controls matched for gender, age and ward of admission. Altogether, 123 case control pairs with mean age 70.1 years were included. Isolates from 89 (72. 4%) of the cases were identical or related to the defined outbreak strain as determined by pulsed field gel electrophoresis (PFGE). In 75 of the patients (60.9%), ARE caused urinary tract infection, five (4.1%) had bacteraemia, 33 (26.8%) had wound infection and 10 (8.1%) had other infections. In a logistic regression model for 1:1 matched samples, the following factors were identified as significant risk factors for ARE infection: underlying neurological disease (OR=33.5), prescription of antimicrobial agents for more than 10 days (OR=8. 99), prescription of cephalosporins (OR=4.69), underlying gastrointestinal disease (OR=3.36) and length of hospital stay per day (OR=1.04). The intrahospital death rate for the cases was 18.7% compared with 8.9% for the controls, corresponding to an excess mortality attributable to ARE infection of 9.8%. A history of carbapenem prescription was the only independent factor contributing to death (OR=5.64) when comparing ARE patients dying in hospital to those surviving. PMID- 10860691 TI - Predominant staphylococci in the intensive care unit of a paediatric hospital. AB - Coagulase-negative staphylococci cause a significant number of infections, especially in immunocompromised patients, including premature neonates. Nosocomial strains present in the environment create a special risk.We studied staphylococci isolated from the intensive care unit of a paediatric teaching hospital over the period of six months in 1997. Biotyping and species identification were performed; resistance to methicillin and other beta-lactam antibiotics and patterns of resistance to antimicrobial agents were determined. Staphylococcus cohnii was the predominant species of 147 isolates of staphylococci recovered from the ward environment. Strains were resistant to several antibiotics and 97% were resistant to methicillin. In isolates from infants (72) methicillin-resistant strains of Staphylococcus epidermidis were predominant. Susceptibility to beta-lactams (penicillin, amoxycillin, amoxycillin clavulanic acid and cephalosporins: cephalothin, cefuroxime and cefotaxime) showed differences between the two species. Some S. cohnii were susceptible to penicillin and amoxycillin despite methicillin-resistance. S. epidermidis were relatively susceptible to amoxycillin-clavulanic acid and cephalosporins. All strains investigated were susceptible to vancomycin, but nearly 30% demonstrated high-level resistance to mupirocin. The search for strains of the same origin showed clones belonging to S. epidermidis, S. hominis and S. saprophyticus but not S. cohnii.A large number of multiresistant, phenotypically different S. cohnii strains surviving in the ward environment may provide a reservoir of antimicrobial resistance genes. PMID- 10860692 TI - Can household dishwashers be used to disinfect medical equipment? AB - In hospitals, medical instruments are usually cleaned and disinfected in a washer/disinfector. However, it is not efficient for small hospitals or general practitioners to purchase such machines as they would not be working to capacity. We investigated the possibility of cleaning and disinfecting medical equipment in a conventional household dishwasher modified to achieve a temperature of 71 degrees C. For this purpose we contaminated screws with different test soils containing either bacterial (100 screws) or viral (106 screws) suspensions. Test organisms were re-isolated from 2% of the screws after bacterial contamination and 4.7% of those with viral contamination. In both cases we found dishwasher processing to be a suitable means of disinfecting medical instruments. PMID- 10860693 TI - Outbreak of scabies in Norwegian nursing homes and home care patients: control and prevention. AB - Over a period of five months (October 1998-February 1999), an outbreak of scabies affected 19 persons associated with a nursing home in Oslo, Norway. Scabies was diagnosed in 13 patients; six long-term patients, five short-term patients also cared for at home, and two home care patients associated with the same institution. Six healthcare workers who had assisted with infected patients in their own homes were also diagnosed with scabies. Two separate index cases were found. Both had had pruritus for several months, diagnosed as eczema, and were repeatedly treated at dermatology outpatient clinics before the diagnosis was made. Both index cases were cared for at home and in the nursing home (short term). Repeated treatments with permethrin were tried before effective treatment with benzyl benzoate. Altogether 370 persons (patients, staff, relatives) were treated. In June-July 1999, scabies was diagnosed in two other nursing homes; six patients or staff, and 156 persons were treated. Patients with scabies were contact isolated and disinfection and cleaning was performed. Simultaneous treatment and washing or disinfection of clothing, bedding and environment of all potentially affected individuals is imperative to control an outbreak of scabies. PMID- 10860694 TI - A prospective clinical study to investigate the microbial contamination ofa needleless connector. AB - Needleless connectors, which allow direct access to intravascular catheters, are widely used in clinical practice. The benefits of these devices to healthcare workers are well documented; however, the potential risk of microbial contamination and associated infection is unclear. This clinical study evaluated microbial contamination rates for a needleless connector, Connecta Clave(R) (CC(R)), as compared to a conventional three-way tap, which was connected to the hubs of central venous catheters (CVC) immediately following insertion. Patients in the study group had CC(R) attached to the three-way taps, whereas the control group had standard entry port caps. On removal (up to 72 h) the connectors were studied for microbial contamination. There was no significant difference between the number of three-way taps contaminated on the internal surface with micro organisms in the control group with entry port caps (19/132, 14%) compared to the group with CC(R) (18/105, 17%). Sixteen percent (27/173) of the CC(R) were contaminated with micro-organisms on the internal surfaces. The external surface of 33% (27/82) of the CC(R) silicone seals were contaminated after clinical use. Micro-organisms were also isolated from 9% (8/91) of the silicone seals after disinfection. The use of this needleless connector, compared to standard caps therefore does not appear to increase the risk of infection via the internal lumen of three-way taps. PMID- 10860695 TI - Does MRSA affect patient outcomes in the elderly? A retrospective pilot study. AB - We retrospectively studied admissions to our geriatric acute assessment and rehabilitation ward over a one-year period, to identify those with Methicillin resistant Staphylococcus aureus (MRSA) and determine whether this affected outcomes. Two hundred and thirty eight admissions of 204 patients were analysed and 9.8% of patients were MRSA positive. Demographics did not differ between MRSA positive and negative patients. Respiratory co-morbidity was more common in MRSA positive patients. Rates of functional decline did not differ between the two groups. Those colonized or infected by MRSA had a significantly longer stay (51.4 vs. 32.2 days, P=0.03), perhaps due to isolation and limited rehabilitation. The virulence of MRSA may be less in these patients, therefore isolation may be inappropriate and counter-productive. PMID- 10860696 TI - Cumulative benefit games: achieving cooperation when players discount the future. AB - Experimental studies with captive animals show strong preferences for immediate reward. Several authors have argued that these tendencies to discount delayed reward may severely limit the Iterated Prisoner's Dilemma game as a model of animal cooperation. This paper explores a simple mechanism, dubbed cumulative games, that can, in principle, promote cooperative action even when there is strong temporal discounting. In the simplest type of cumulative game a pair of players does not receive benefits at the end of each play, as in a conventional repeated game, but must complete a sequence of games before collecting the accumulated benefits. In a preliminary analysis pitting tit-for-tat against all D, I show that accumulation can promote a conditionally cooperative strategy even when there is strong temporal discounting. However, the delays created by accumulation de-value the pairwise interaction, so although the relative value of cooperation increases, the total value of the interaction decreases. I investigate accumulation further by simulating the evolution of a broader class of strategies. These simulation studies show that accumulation, and small discounting rates (high future value) can both promote cooperative action. The limitations of these results are discussed. PMID- 10860697 TI - Mechanisms of cortical electrical activity and emergence of gamma rhythm. AB - A continuum model of the electrical activity of the cerebral cortex is described which predicts the occurrence of a resonance in the gamma range near 40 Hz. The emergence of this resonance is due to two refinements to a previous model, namely the inclusion of a modulation of synaptic strength due to finite reversal potentials, and use of parameters that better match physiological measurements. Analytical expressions for the fixed points of the system and for its linear dynamics are found in terms of average neuronal properties, and together explain the occurrence and modulation of the gamma-like resonance. The analytical results are confirmed by a numerical simulation. PMID- 10860698 TI - Exploratory recruitment plasticity in a social spider (Anelosimus eximius). AB - Exploratory recruitment was investigated in an artificial experimental set-up on location in French Guyana. Groups of 200 freshly collected spiders of the neotropical social theridiid Anelosimus eximius were released on an open circular surface and offered a choice between two accessible shelters. Results indicated that a clear-cut collective decision was not always reached, unlike what we found using a different set-up in another set of experiments. Simulations were conducted using available information in order to explore the potential causes for this difference. Theoretical projections fit experimental data and strongly suggest that variability in the collective response results from a combination between modifications of the environment's properties and alteration of the recruitment procedure. Multiple variants of the theoretical set-up (including external bias) are investigated and emphasize plasticity in the collective response. New experimental studies are suggested and adaptative value of exploratory recruitment in social spiders is briefly discussed. PMID- 10860699 TI - Head-on collisions of waves in an excitable FitzHugh-Nagumo system: a transition from wave annihilation to classical wave behavior. AB - For the particular case of an excitable FitzHugh-Nagumo system with diffusion, we investigate the transition from annihilation to crossing of the waves in the head on collision. The analysis exploits the similarity between the local and the global phase portraits of the system. We find that the transition has features typical of the nucleation theory of first-order phase transitions, and may be understood through purely geometrical arguments. In the case of periodic boundary conditions, the transition is an infinite-dimensional analog of the creation and the vanishing of limit cycles via a homoclinic Andronov bifurcation. Both before and after the transition, the behavior of a single cell continues to be typical for excitable systems: a stable equilibrium state, and a threshold above which an excitation pulse can be induced. The generality and qualitative character of our argument shows that the phenomenon described can be observed in excitable systems well beyond the particular case presented here. PMID- 10860700 TI - Predictive modeling of mixed microbial populations in food products: evaluation of two-species models. AB - Predictive microbiology is an emerging research domain in which biological and mathematical knowledge is combined to develop models for the prediction of microbial proliferation in foods. To provide accurate predictions, models must incorporate essential factors controlling microbial growth. Current models often take into account environmental conditions such as temperature, pH and water activity. One factor which has not been included in many models is the influence of a background microflora, which brings along microbial interactions. The present research explores the potential of autonomous continuous-time/two-species models to describe mixed population growth in foods. A set of four basic requirements, which a model should satisfy to be of use for this particular application, is specified. Further, a number of models originating from research fields outside predictive microbiology, but all dealing with interacting species, are evaluated with respect to the formulated model requirements by means of both graphical and analytical techniques. The analysis reveals that of the investigated models, the classical Lotka-Volterra model for two species in competition and several extensions of this model fulfill three of the four requirements. However, none of the models is in agreement with all requirements. Moreover, from the analytical approach, it is clear that the development of a model satisfying all requirements, within a framework of two autonomous differential equations, is not straightforward. Therefore, a novel prototype model structure, extending the Lotka-Volterra model with two differential equations describing two additional state variables, is proposed to describe mixed microbial populations in foods. PMID- 10860701 TI - Translocation induced outgrowth of fungi in nutrient-free environments. AB - The study of alternatives to chemical methods of nematode control in agriculture has received significant recent interest. One such method is biological control using nematode trapping fungi such as Arthrobotrys superba. To understand how these fungi can be implemented as effective nematicides, it is essential to study their outgrowth into soil from localized nutrient resources. In this paper, we use a mathematical model to investigate the outgrowth of fungi into an environment essentially without available nutrients capable of supporting net growth. By comparing model solutions with experimental results, we show that in such circumstances, continual mycelial expansion can only be obtained if internal metabolites are actively translocated through the mycelium. Predictions are made concerning the maximal extension possible from a given quantity of nutrients and a testable functional relationship between the two is derived. Using this modelling technique we are able to map not only biomass extent but also biomass distribution at each stage. The type of environmental heterogeneity investigated here is encountered by many species of fungi in nature and is of relevance for the introduction of specific fungi into soil for biological control or bioremediation purposes. PMID- 10860702 TI - Removing allometric effects of body size in morphological analysis. AB - In the present paper, a normalization technique to scale data that exhibit an allometric growth is presented and the way it has to be used is described. It is shown how the method has been derived from the theoretical equations of allometric growth. Consequently, the method completely removes all the information related to size, not only scaling all individuals to the same size, but also adjusting their shape to that they would have in the new size according to allometry. In the particular case of isometry when the measures are of identical dimension, this normalization coincides with ratios (one of the most popular methods but only valid in this particular case). This procedure is a theoretical generalization of the technique used by Thorpe (1975, Biol. J. Linn. Soc.7, 27-43; 1976, Biol. Rev.51, 407-452) which was recorded as one of the most efficient methods in the empirical evaluation done by Reist (1985, Can. J. Zool.63, 1429-1439). PMID- 10860703 TI - Optimal stopover decisions under wind influence: the effects of correlated winds. AB - Wind speed and direction have a significant effect on a flying bird's ground speed. Migrants are therefore expected to be sensitive to wind conditions and this should have consequences for optimal strategies of stopover and refuelling. Based on an earlier model of time-minimizing migration which includes wind condition, we investigate the consequences of the temporal correlation of wind conditions. Day-to-day changes in wind conditions are modelled with a two-state Markov process and an expression for the expected speed of migration is derived. The policy of the migrants is described by two parameters: a day t(g) when the birds start to leave whenever favourable conditions occur and a later day t(b)when they leave even in unfavourable winds. The model predicts that in most cases departures should be close to the date which is predicted by a wind-free deterministic model and that the birds should never leave without wind assistance. Only if the probability that the condition remains the same on the following day is close to 1 should the birds leave even in unfavourable conditions shortly after the deterministic optimal date. If the transition matrix is highly asymmetrical, i.e. if it is very probable that unfavourable conditions remain and that favourable conditions will change into unfavourable, then the birds are predicted to start using good winds several days before the deterministic optimal date. An analysis of six years of wind data from two sites in Sweden shows that wind directions on successive days are in fact correlated in all years. PMID- 10860704 TI - Population genetics of rhizobia: construction and analysis of an "Infection and Release" model. AB - A mathematical model is created to assess the inputs of sym gene transfer of in planta multiplication and of interstrain competition into dynamics of the rhizobia populations. Their microevolution is presented as a series of the "infection and release" cycles; each cycle includes transfer of sym genes from virulent initial symbionts to avirulent local bacteria yielding the virulent novel symbionts; competition between initial symbionts and novel symbionts for the host nodulation; multiplication of initial symbionts and novel symbionts in planta and their release into soil; competition between the released novel symbionts and resident local bacteria for ex planta survival. A recurrent equation is created to determine the number of novel symbionts at each cycle of evolution of the closed bacteria-plant system. Its analysis demonstrates that under certain, really allowable values of the introduced parameters two major effects may occur: (a) rapid multiplication of novel symbionts arisen from sym gene transfer; and (b) increase of frequency of rare local bacteria genotypes after acquisition of virulence. Multiplication of very rare strains (p<10(-19)) in the plant-associated bacteria population is possible at certain parameters of the system. Variation of the sizes of bacteria populations and of the parameters for interstrain competition may influence the evolutionary rate of the bacteria population. The "infection and release" model represents a selective mechanism which may be responsible for a high taxonomic diversity of rhizobia and for a panmictic structure of their populations. PMID- 10860705 TI - Adaptation to spatially heterogeneous modifying and adaptive environments. AB - The development of an individual's phenotype is influenced by environmental factors (the modifying environment) which may differ from those factors (the adaptive environment) that decide on the adaptational value of the developed phenotype. The shapes of adaptationally optimal norms of reaction are therefore essentially determined by associations between these two environmental components together with the degree of adaptational sensitivity of the developed phenotypes. Two complementary aspects of optimality are accounted for: (a) environments can be optimal for a given norm of reaction and (b) norms of reaction can be optimal for a given environment. The results are obtained for random distribution of genotypes over environmental conditions and under the physiologically reasonable premise that fitness is a function of the costs of modification and adaptation. It turned out that the associations of adaptive and modifying environments are the primary sources of adaptational optimization. More specifically, it is shown that (i) independence between the two environmental components constitutes an adaptationally optimal environment only for norms of reaction in which all phenotypes are adaptively insensitive; (ii) if costs of modification do not depend on the environment, and if the two environmental components are not associated, adaptationally optimal norms of reaction can always be realized through phenogenetic invariance; (iii) as a rule, adaptively sensitive phenotypes developed under strong environmental associations necessitate phenogenetic plasticity for the optimal norm of reaction; (iv) a norm of reaction which is adaptationally optimal in its adaptationally optimal environment can always be realized through phenogenetic invariance, if costs of modification do not vary with the environment. These results reveal an important role of patterns of adaptive sensitivity of phenotypes, which may even surpass that of shapes of norms of reaction in adaptational processes. PMID- 10860706 TI - Simulated navigation based on observed gradients of atmospheric trace gases (Models on pigeon homing, part 3). AB - An earlier developed model, simulating pigeon homing based on fictitious gradients of atmospheric odours, was applied to actually observed spatial distributions of volatile hydrocarbons. The model calculations demonstrate that sufficient information on a bird's current position with respect to home can be derived from the ratios among three or more chemical compounds which gradually vary over distances of several hundreds of kilometres, differently in different directions. Flight directions computed by model birds from such observed ratios are roughly but not perfectly homeward-oriented from most positions within the investigated radius of 200 km around home. Performances of model birds are at least as good as those of real pigeons in the field. According to calculations using atmospheric data collected under different wind directions, the birds might, but possibly need not, take the current weather conditions into account when evaluating olfactory signals. It is necessary, however, that the birds acquire, during their long-term stay at the home site, some knowledge of the directions of relevant gradients. Homing experiments with pigeons as well as measurements of atmospheric trace substances are consistent with the hypothesis that this knowledge is gained by correlating wind directions with specific changes of ratios among a number of compounds. This assumed process requires further elucidation. PMID- 10860707 TI - Language evolution and information theory. AB - This paper places models of language evolution within the framework of information theory. We study how signals become associated with meaning. If there is a probability of mistaking signals for each other, then evolution leads to an error limit: increasing the number of signals does not increase the fitness of a language beyond a certain limit. This error limit can be overcome by word formation: a linear increase of the word length leads to an exponential increase of the maximum fitness. We develop a general model of word formation and demonstrate the connection between the error limit and Shannon's noisy coding theorem. PMID- 10860708 TI - A comment on sex ratio with intrasex sibling cooperation. PMID- 10860709 TI - Cancer, somatic mutation and stem cells. PMID- 10860710 TI - Functional role of mycobacteriophage transfer RNAs. PMID- 10860711 TI - A solid phase plate assay for HIV-1 genotyping. AB - A prototype solid phase plate assay (SPPA) for the detection and genotyping of HIV-1 subtypes was developed using a PCR-based capture hybridization format. Well characterized HIV-1 reference controls of known subtypes and subtype specific capture oligonucleotide probes targeting several regions of the envelope (env) gene of HIV-1 were selected to develop the assay. The subtype specific oligonucleotide probes were covalently bound to microtubes in an ordered pattern and biotin labelled primers were used to amplify the target sequences. The PCR products were hybridized to the corresponding oligonucleotide probes, and colorimetrically detected by a chromogenic reaction using a standard microplate reader. All the HIV-1 subtype reference controls specifically hybridized to the corresponding capture probes and negligible cross-hybridization between subtypes was observed. To demonstrate the performance and reproducibility of the SPPA system and its validation with clinical samples, several human plasma samples of unknown and known HIV-1 subtypes were tested. The SPPA is highly specific and unambiguously identify the major subtypes of the HIV-1 M and O groups. This assay could be a useful method for subtyping samples of HIV-1 infected individuals and for disease management. PMID- 10860712 TI - A simple and reliable method for the detection of the 30delG mutation of the CX26 gene. AB - Mutations in the CX26 gene (GJB2), encoding the gap-junction protein Connexin-26, have been shown to be the major cause of non-syndromic recessive deafness. Among these mutations, the deletion of a guanine within the stretch of six G between nucleotide positions +30 and +35 of the CX26 cDNA (30delG) accounts for the majority of this kind of deafness. Molecular detection of the 30delG mutation is usually performed by direct sequencing analysis of PCR products or by SSCP. To detect this mutation we developed an easy and reliable method, based on PCR, followed by a non-radioactive sandwich hybridization on microtiter plates. We tested 188 individuals recruited from the genetic counseling service for deaf people at the Pasteur Hospital and at the Armand-Trousseau Children's Hospital, Paris, France between April 1997 and September 1998. Our screening method is simple, uses stable and safe reagents, and employs inexpensive equipment. As such, it is suitable for widespread use in genetic diagnosis. PMID- 10860713 TI - Evaluation of the accuracy and reproducibility of a practical PCR panel assay for rapid detection and differentiation of Mycobacterium avium subspecies. AB - The Mycobacterium avium subspecies (MAs) include the closely related MAs avium and MAs paratuberculosis. This study was conducted to evaluate the performance of a PCR panel assay as a diagnostic tool to detect and differentiate MAs avium and MAs paratuberculosis infection. Specific oligonucleotides primers derived from the 16 S rRNA (MAs) sequence, insertion elements IS 901 (MAs avium), IS 1245 Mycobacterium avium complex (MAC), IS 900 (MAs paratuberculosis), and the hspX (MAs paratuberculosis) gene sequences were synthesized and used in preassembled PCR reaction mixtures. These five primer sets made up the PCR panel assay. To determine the accuracy of the PCR panel assay for MAs avium and MAs paratuberculosis strain detection and differentiation, lysates of mycobacterial DNA from 120 (n=120) strains were tested with the PCR panel assay by one laboratory (#1). The PCR panel assay specifically detected and differentiated 91/91 (100%) of MAs avium and MAs paratuberculosis strains tested in this study. The PCR panel assay also specifically differentiated all MAs avium and MAs paratuberculosis strains from all but one (M. intracellulare, serovar 23) of the other mycobacterial strains tested. To confirm the accuracy and evaluate the reproducibility of the PCR panel assay, samples were numbered and given to a different laboratory (#2) as 'unknowns' for identification by the PCR panel assay. In this study, the overall accuracy for strain identification using the PCR panel assay was 99.2% (119/120). The reproducibility of the PCR panel assay when comparing data from laboratory #1 with laboratory #2 was found to be 100% (120/120). These results indicate that this 'easy-to-use', rapid PCR method can accurately and reliably detect and differentiate closely related MAs avium and MAs paratuberculosis from each other and from other mycobacterial species. The PCR panel assay can also differentiate mixed cultures of MAs. The simplicity of this PCR method could be beneficial to laboratories that test for members of MA. PMID- 10860714 TI - Identification of bacteria from a non-healing diabetic foot wound by 16 S rDNA sequencing. AB - Approximately 10-20% of diabetic foot wounds fail initial antibiotic treatment. It is generally believed that several bacterial species may be present in these types of wounds. Because some of these organisms cannot be easily cultured, proper identification is problematic and thus, appropriate treatment modalities cannot be applied. This report examined the bacterial flora present in a chronic diabetic foot wound that failed antibiotic treatment. A tissue sample was collected from the base of the wound and used for standard microbiological culturing. DNA from the sample was used to amplify bacterial 16 S rDNA gene sequences and a library of these sequences was made. The clones were placed into two major groups on the basis of their melting temperatures. Representatives of these groups were sequenced, and information was used to identify the bacteria present in the wound. The culture-based method identified a single anaerobic species, Bacteroides fragilis. The method employing rDNA sequencing identified B. fragilis as a dominant organism and Pseudomonas (Janthinobacterium) mephitica as a minor component. The results indicate that rDNA sequencing approach can be an important tool in the identification of bacteria from wounds. PMID- 10860715 TI - PCR-RFLP and sequence analysis of a non-ribosomal fragment for genetic characterization of European stone fruit yellows phytoplasmas infecting various Prunus species. AB - A 927 bp non-ribosomal fragment was used to assess the genetic variability of the European stone fruit yellows (ESFY) phytoplasma infecting 14 different Prunus species. For this, 175 isolates originating from four different Mediterranean countries were tested by PCR-RFLP analysis with seven restriction enzymes. No polymorphism among the ESFY phytoplasma could be observed but 12 out of 18 restriction sites differed between the homologous fragments of ESFY and apple proliferation (AP) phytoplasmas. An 846 bp fragment of a French ESFY isolate was sequenced, it included the 3'-end of a putative nitroreductase gene, an intergenic region and a truncated open reading frame. This ESFY phytoplasma sequence showed 89.7% identity with the equivalent AP phytoplasma nucleotide sequence (83. 9% identity at the amino acid level). The G+C content of the entire sequence was extremely low (15.4%) and A+T-rich codons were highly preferred in codon usage. In this paper, we report the presence of the ESFY phytoplasma for the first time in Turkey and in five Prunus hosts never reported previously. Our results also indicate that the ESFY phytoplasma isolates affecting various Prunus species are genetically homogenous but can be distinguished from the AP phytoplasma. Therefore, they are likely to represent different taxons. PMID- 10860716 TI - PCR detection of Giardia lamblia in stool: targeting intergenic spacer region of multicopy rRNA gene. AB - A PCR based detection that amplifies the 552-bp intergenic spacer (IGS) region of multicopy rRNA gene of Giardia lamblia and 320-bp internal sequences to first PCR product has been used in diagnosis of giardiasis in stool sample. The primers were found highly specific to Giardia spp. only, because no amplification was observed with DNAs from other enteric pathogens like Escherichia coli, Shigella dysenteriae and Entamoeba histolytica. The test could detect even less than 2 pg of genomic DNA from Giardia trophozoites. In direct diagnosis of Giardia lamblia in stool samples, it was observed that PCR amplification of IGS followed by nested PCR could enhance the sensitivity and specificity of the tests manifold and the system was able to detect as low as 10 parasites in 100 microl of stool. The comparative evaluation of the present system with conventional microscopy, CIEP and ELISA in the diagnosis of giardiasis from diarrhoeic stool samples and control subjects demonstrated a 100% correlation among nested PCR, microscopic examination and ELISA in patients suggestive of giardiasis (Group I) and control subjects (Group II). In Group I cases (patients suffering from other than giardiasis), CIEP, ELISA and nested PCR showed better results than microscopic examination. However, among them, PCR was found most sensitive and specific because 20% positivity was noticed by PCR whereas CIEP and ELISA showed only 7.14% and 12.85%, respectively. Break-up results showed that all the samples which were positive by CIEP or ELISA, also found positive by PCR. The present observation clearly suggests the use of PCR that amplifies the intergenic spacer region of multicopy rRNA gene of Giardia lamblia followed by nested PCR for routine, quick and reliable detection of Giardia lamblia in stool samples. PMID- 10860717 TI - Deletion polymorphism in the coding region of the human NESP55 alternative transcript of GNAS1. AB - NESP55, a novel member of the chromogranins, was originally implicated as a precursor of a peptide LSAL with 5-HT1B receptor antagonist activity. In humans, NESP55 (MIM 139320) is encoded by an alternative transcript of GNAS1, the gene encoding the guanine nucleotide-binding alpha subunit of G(S). As a result of the potential relevance of NESP55 to serotoninergic neurotransmission, we screened its sequence using genomic DNA pools from autistic disorder, obsessive-compulsive disorder (OCD) probands and control subjects. Six single nucleotide polymorphisms (SNPs) were identified and the allele frequencies of those SNPs were determined. In addition, a 24-bp in-frame deletion in the coding region was found in one of the OCD probands. To further investigate its pattern of inheritance and the relevance to studied phenotypes, we genotyped 123 total subjects from autism, OCD and attention deficit hyperactivity disorder (ADHD) families. The deletion was detected only in one OCD family and followed Mendelian inheritance. All subjects with the deletion were heterozygous. However, there are no specific behavioural or physical alterations in the subjects with this deletion variant. The physiological role of NESP55 in serotoninergic neurotransmission as well as the effect of the deletion on its function should be evaluated in future studies. PMID- 10860718 TI - A common T/C polymorphism in the promoter region of the beta T-cell receptor gene. AB - Polymorphisms in the T-cell receptor genes can provide important information for the study of the immune response system, particularly for autoimmune diseases. This report characterizes a common T to C polymorphism in the promoter of the beta 2 constant chain of the T-cell receptor, which abolishes a recognition site for BglII restriction endonuclease. PMID- 10860719 TI - Interaction of translation initiation factor IF1 with the E. coli ribosomal A site. AB - Initiation Factor 1 (IF1) is required for the initiation of translation in Escherichia coli. However, the precise function of IF1 remains unknown. Current evidence suggests that IF1 is an RNA-binding protein that sits in the A site of the decoding region of 16 S rRNA. IF1 binding to 30 S subunits changes the reactivity of nucleotides in the A site to chemical probes. The N1 position of A1408 is enhanced, while the N1 positions of A1492 and A1493 are protected from reactivity with dimethyl sulfate (DMS). The N1-N2 positions of G530 are also protected from reactivity with kethoxal. Quantitative footprinting experiments show that the dissociation constant for IF1 binding to the 30 S subunit is 0.9 microM and that IF1 also alters the reactivity of a subset of Class III sites that are protected by tRNA, 50 S subunits, or aminoglycoside antibiotics. IF1 enhances the reactivity of the N1 position of A1413, A908, and A909 to DMS and the N1-N2 positions of G1487 to kethoxal. To characterize this RNA-protein interaction, several ribosomal mutants in the decoding region RNA were created, and IF1 binding to wild-type and mutant 30 S subunits was monitored by chemical modification and primer extension with allele-specific primers. The mutations C1407U, A1408G, A1492G, or A1493G disrupt IF1 binding to 30 S subunits, whereas the mutations G530A, U1406A, U1406G, G1491U, U1495A, U1495C, or U1495G had little effect on IF1 binding. Disruption of IF1 binding correlates with the deleterious phenotypic effects of certain mutations. IF1 binding to the A site of the 30 S subunit may modulate subunit association and the fidelity of tRNA selection in the P site through conformational changes in the 16 S rRNA. PMID- 10860720 TI - Sequences within Pr160gag-pol affecting the selective packaging of primer tRNA(Lys3) into HIV-1. AB - The selective packaging of the primer tRNA(Lys3) into HIV-1 particles is dependent upon the viral incorporation of the Pr160gag-pol precursor protein. In order to map a tRNA(Lys3) binding site within this precursor, we have studied the effects of mutations in Pr160gag-pol upon the selective incorporation of tRNA(Lys3). Many of these mutations were placed in a protease-negative HIV-1 proviral DNA to prevent viral protease degradation of the mutant Gag-Pol protein. C-terminal deletions of protease-negative Gag-Pol that removed the entire integrase sequence and the RNase H and connection subdomains of reverse transcriptase did not inhibit the incorporation of either the truncated Gag-Pol or the tRNA(Lys3), indicating that these regions are not required for tRNA(Lys3) binding. On the other hand, larger C-terminal deletions, which also remove the thumb subdomain sequence, did prevent tRNA(Lys3) packaging, without inhibiting viral incorporation of the truncated Gag-Pol, indicating a possible interaction between thumb subdomain sequences and tRNA(Lys3). While point mutations K249E, K249Q, and R307E in the primer grip region of the thumb subdomain have been reported to inhibit the in vitro interaction of mature reverse transcriptase with the anticodon loop of tRNA(Lys3), we find that these mutations do not inhibit tRNA(Lys3) packaging into the virus, which supports other work indicating that the anticodon loop of tRNA(Lys3) is not involved in interactions with Pr160gag pol during tRNA(Lys3) packaging. PMID- 10860721 TI - Genomic sequences of bacteriophages HK97 and HK022: pervasive genetic mosaicism in the lambdoid bacteriophages. AB - We report the complete genome DNA sequences of HK97 (39,732 bp) and HK022 (40,751 bp), double-stranded DNA bacteriophages of Escherichia coli and members of the lambdoid or lambda-like group of phages. We provide a comparative analysis of these sequences with each other and with two previously determined lambdoid family genome sequences, those of E. coli phage lambda and Salmonella typhimurium phage P22. The comparisons confirm that these phages are genetic mosaics, with mosaic segments separated by sharp transitions in the sequence. The mosaicism provides clear evidence that horizontal exchange of genetic material is a major component of evolution for these viruses. The data suggest a model for evolution in which diversity is generated by a combination of illegitimate and homologous recombination and mutational drift, and selection for function produces a population in which most of the surviving mosaic boundaries are located at gene boundaries or, in some cases, at protein domain boundaries within genes. Comparisons of these genomes highlight a number of differences that allow plausible inferences of specific evolutionary scenarios for some parts of the genome. The comparative analysis also allows some inferences about function of genes or other genetic elements. We give examples for the generalized recombination genes of HK97, HK022 and P22, and for a putative headtail adaptor protein of HK97 and HK022. We also use the comparative approach to identify a new class of genetic elements, the morons, which consist of a protein-coding region flanked by a putative delta 70 promoter and a putative factor-independent transcription terminator, all located between two genes that may be adjacent in a different phage. We argue that morons are autonomous genetic modules that are expressed from the repressed prophage. Sequence composition of the morons implies that they have entered the phages' genomes by horizontal transfer in relatively recent evolutionary time. PMID- 10860722 TI - Genomic sequence and analysis of the atypical temperate bacteriophage N15. AB - N15 is a temperate bacteriophage that forms stable lysogens in Escherichia coli. While its virion is morphologically very similar to phage lambda and its close relatives, it is unusual in that the prophage form replicates autonomously as a linear DNA molecule with closed hairpin telomeres. Here, we describe the genomic architecture of N15, and its global pattern of gene expression, which reveal that N15 contains several plasmid-derived genes that are expressed in N15 lysogens. The tel site, at which processing occurs to form the prophage ends is close to the center of the genome in a similar location to that occupied by the attachment site, attP, in lambda and its relatives and defines the boundary between the left and right arms. The left arm contains a long cluster of structural genes that are closely related to those of the lambda-like phages, but also includes homologs of umuD', which encodes a DNA polymerase accessory protein, and the plasmid partition genes, sopA and sopB. The right arm likewise contains a mixture of apparently phage- and plasmid-derived genes including genes encoding plasmid replication functions, a phage repressor, a transcription antitermination system, as well as phage host cell lysis genes and two putative DNA methylases. The unique structure of the N15 genome suggests that the large global population of bacteriophages may exhibit a much greater diversity of genomic architectures than was previously recognized. PMID- 10860723 TI - DNA melting and promoter clearance by eukaryotic RNA polymerase I. AB - Ribosomal RNA transcription initiation requires the melting of DNA to form an open complex, formation of the first few phosphodiester bonds, commencement of RNA polymerase I movement along the DNA, clearance of the promoter, and the formation of a steady-state ternary elongation complex. We examined DNA melting and promoter clearance by using potassium permanganate, diethylpyrocarbonate and methidiumpropylEDTA.Fe(II) footprinting. In combination, these methods demonstrated: (1) TIF-IB and RNA polymerase I are the only proteins required for formation of an initial approximately 9 base-pair open promoter region. This finding contradicts earlier results using diethylpyrocarbonate alone, which suggested an RNA synthesis requirement for stable melting. (2) DNA melting is temperature-dependent, with a tm between 15 and 20 degrees C. (3) Temperature dependency of melting, as well as stalling the polymerase at sites close to the transcription start site revealed that the melted DNA region initially opens upstream of the transcription initiation site, and enlarges in a downstream direction coordinate with initiation, eventually attaining a steady-state transcription bubble of approximately 19 base-pairs. (4) The RNA-DNA hybrid protects the template DNA from single-strand footprinting reagents. The hybrid is 9 bp in length, consistent with the longer hybrid estimated by some for the Escherichia coli polymerase and with the hybrids estimated for eukaryotic polymerases II and III. PMID- 10860724 TI - Mutations in the N-terminal region of RecA that disrupt the stability of free protein oligomers but not RecA-DNA complexes. AB - We have introduced targeted mutations in two areas that make up part of the RecA subunit interface. In the RecA crystal structure, cross-subunit interactions are observed between the Lys6 and Asp139 side-chains, and between the Arg28 and Asn113 side-chains. Unexpectedly, we find that mutations at Lys6 and Arg28 impose sever defects on the oligomeric stability of free RecA protein, whereas mutations at Asn113 or Asp139 do not. However, Lys6 and Arg28 mutant proteins showed an apparent normal formation of RecA-DNA complexes. These results suggest that cross subunit contacts in this region of the protein are different for free RecA protein filaments versus RecA-DNA nucleoprotein filaments. Mutant proteins with substitutions at either Lys6 or Arg28 show partial inhibition of DNA strand exchange activity, yet the mechanistic reasons for this inhibition appear to be distinct. Although Lys6 and Arg28 appear to be more important to the stability of free RecA protein, as opposed to the stability of the catalytically active nucleoprotein filament, our results support the idea that the cross-subunit interactions made by each residue play an important role in optimizing the catalytic organization of the active RecA oligomer. PMID- 10860725 TI - Crystal structure of an adenine bulge in the RNA chain of a DNA.RNA hybrid, d(CTCCTCTTC).r(gaagagagag). AB - Crystal structure of a DNA.RNA hybrid, d(CTCCTCTTC).r(gaagagagag), with an adenine bulge in the polypurine RNA strand was determined at 2.3 A resolution. The structure was solved by the molecular replacement method and refined to a final R-factor of 19.9% (Rfree 22.2%). The hybrid duplex crystallized in the space group I222 with unit cell dimensions, a = 46.66 A, b = 47.61 A and c = 54.05 A, and adopts the A-form conformation. All RNA and DNA sugars are in the C3'-endo conformation, the glycosyl angles in anti conformation and the majority of the C4'-C5' torsion angles in g+ except two trans angles, in conformity with the C3'-endo rigid nucleotide hypothesis. The adenine bulge is looped out and it is also in the anti C3'-endo conformation. The bulge is involved in a base-triple (C.g)*a interaction with the end base-pair (C9.g10) in the minor groove of a symmetry-related molecule. The 2' hydroxyl group of g15 is hydrogen bonded to O2P and O5' of g17, skipping the bulged adenine a16 and stabilizing the sugar phosphate backbone of the hybrid. The hydrogen bonding and the backbone conformation at the bulged adenine site is very similar to that found in the crystal structure of a protein-RNA complex. PMID- 10860726 TI - The crystal structure of the octamer [r(guauaca)dC]2 with six Watson-Crick base pairs and two 3' overhang residues. AB - The crystal structure of an alternating RNA octamer, r(guauaca)dC (RNA bases are in lower case while the only DNA base is in upper case), with two 3' overhang residues one of them a terminal deoxycytosine and the other a ribose adenine, has been determined at 2.2 A resolution. The refined structure has an Rwork 18.6% and Rfree 26.8%. There are two independent duplexes (molecules I and II) in the asymmetric unit cell, a = 24.95, b = 45.25 and c = 73.67 A, with space group P2(1)2(1)2(1). Instead of forming a blunt end duplex with two a+.c mispairs and six Watson-Crick base-pairs, the strands in the duplex slide towards the 3' direction forming a two-base overhang (radC) and a six Watson-Crick base-paired duplex. The duplexes are bent (molecule I, 20 degrees; molecule II, 25 degrees) and stack head-to-head to form a right-handed superhelix. The overhang residues are looped out and the penultimate adenines of the two residues at the top end (A15) are anti and at the bottom (A7) end are syn. The syn adenine bases form minor groove A*(G.C) base triples with C8-H...N2 hydrogen bonds. The anti adenine in molecule II also forms a triple and a different C2-H...N3 hydrogen bond, while the other anti adenine in molecule I does not, it stacks on the looped out overhang base dC. The 3' terminal deoxycytosines form two stacked hemiprotonated trans d(C.C)+ base-pairs and the pseudo dyad related molecules form four consecutive deoxyribose and ribose zipper hydrogen bonds in the minor groove. PMID- 10860727 TI - The solution structure and internal motions of a fragment of the cytidine-rich strand of the human telomere. AB - We present the solution structure of d(CCCTA2CCCTA2CCCTA2CCCT), a fragment of the vertebrate telomere which folds intramolecularly. The four cytidine stretches form an i-motif which includes six intercalated C.C+ pairs and terminates with the cytidines at the 5' extremity of each stretch. Above, the second TA2 linker loops across one of the narrow grooves, while at the bottom, the first and third linkers loop across the wide grooves. At 30 degrees C, the spectra of the first and third linkers are quasi-degenerate. Severe broadening at lower temperature indicates that this results from motional averaging between at least two structures of each bottom loop, and makes it impossible to solve the configuration of the bottom loops directly, in contrast to the rest of the structure. We therefore turned to the modified sequence d(CCCTA(2)5MCCCTA2CCCUA2CCCT) in which the two base substitutions (underlined) break the quasi-symmetry between linkers 1 and 3. The three loops follow approximately the hairpin "second pattern" of Hilbers. In the first loop, T4 is in the syn orientation, whereas its analog in the third loop, U16, oriented anti, is in a central location, where it interacts with bases of both loops, thus contributing to their tight association. The only motion is a syn/anti flip of A18 in the third loop. Returning to the telomere fragment, we show that each of the bottom loops switches between the structures identified in the first and third loops of the modified structure. The motions are concerted, and the resulting configurations of the bottom loop cluster present a bulge to either right (T4 syn) or left (T16 syn). PMID- 10860728 TI - NMR structure of stem-loop SL2 of the HIV-1 psi RNA packaging signal reveals a novel A-U-A base-triple platform. AB - The genome of the human immunodeficiency virus type-1 (HIV-1) contains a stretch of approximately 120 nucleotides known as the psi-site that is essential for RNA packaging during virus assembly. These nucleotides have been proposed to form four stem-loops (SL1-SL4) that have both independent and overlapping functions. Stem-loop SL2 is important for efficient recognition and packaging of the full length, unspliced viral genome, and also contains the major splice-donor site (SD) for mRNA splicing. We have determined the structure of the 19-residue SL2 oligoribonucleotide by heteronuclear NMR methods. The structure is generally consistent with the most recent of two earlier secondary structure predictions, with residues G1-G2-C3-G4 and C6-U7 forming standard Watson Crick base-pairs with self-complementary residues C16-G17-C18-C19 and A12-G13, respectively. However, residue A15, which is located near the center of the stem, does not form a predicted bulge, and residues A5 and U14 do not form an expected Watson-Crick base-pair. Instead, these residues form a novel A5-U14-A15 base-triple that appears to be stabilized by hydrogen bonds from A15-H61 and -H62 to A5-N1 and U14 O2, respectively; from A5-H61 to U14-O2, and from C16-H42 to U14-O2'. A kink in the backbone allows the aromatic rings of the sequential U14-A15 residues to be approximately co-planar, adopting a stable "platform motif" that is structurally similar to the A-A (adenosine) platforms observed in the P4-P6 ribozyme domain of the Tetrahymena group I intron. Platform motifs generally function in RNA by mediating long-range interactions, and it is therefore possible that the A-U-A base-triple platform mediates long-range interactions that either stabilize the psi-RNA or facilitate splicing and/or packaging. Residue G8 of the G8-G9-U10-G11 tetraloop is stacked above the U7-A12 base-pair, and the remaining tetraloop residues are disordered and available for potential interactions with either other RNA or protein components. PMID- 10860729 TI - Conservation and variation in superantigen structure and activity highlighted by the three-dimensional structures of two new superantigens from Streptococcus pyogenes. AB - Bacterial superantigens (SAgs) are a structurally related group of protein toxins secreted by Staphylococcus aureus and Streptococcus pyogenes. They are implicated in a range of human pathologies associated with bacterial infection whose symptoms result from SAg-mediated stimulation of a large number (2-20%) of T cells. At the molecular level, bacterial SAgs bind to major histocompatability class II (MHC-II) molecules and disrupt the normal interaction between MHC-II and T-cell receptors (TCRs). We have determined high-resolution crystal structures of two newly identified streptococcal superantigens, SPE-H and SMEZ-2. Both structures conform to the generic bacterial superantigen folding pattern, comprising an OB-fold N-terminal domain and a beta-grasp C-terminal domain. SPE-H and SMEZ-2 also display very similar zinc-binding sites on the outer concave surfaces of their C-terminal domains. Structural comparisons with other SAgs identify two structural sub-families. Sub-families are related by conserved core residues and demarcated by variable binding surfaces for MHC-II and TCR. SMEZ-2 is most closely related to the streptococcal SAg SPE-C, and together they constitute one structural sub-family. In contrast, SPE-H appears to be a hybrid whose N-terminal domain is most closely related to the SEB sub-family and whose C terminal domain is most closely related to the SPE-C/SMEZ-2 sub-family. MHC-II binding for both SPE-H and SMEZ-2 is mediated by the zinc ion at their C-terminal face, whereas the generic N-terminal domain MHC-II binding site found on many SAgs appears not to be present. Structural comparisons provide evidence for variations in TCR binding between SPE-H, SMEZ-2 and other members of the SAg family; the extreme potency of SMEZ-2 (active at 10(-15) g ml-1 levels) is likely to be related to its TCR binding properties. The smez gene shows allelic variation that maps onto a considerable proportion of the protein surface. This allelic variation, coupled with the varied binding modes of SAgs to MHC-II and TCR, highlights the pressure on SAgs to avoid host immune defences. PMID- 10860730 TI - Design, characterization, and structure of a biologically active single-chain mutant of human IFN-gamma. AB - A mutant form of human interferon-gamma (IFN-gamma SC1) that binds one IFN-gamma receptor alpha chain (IFN-gamma R alpha) has been designed and characterized. IFN gamma SC1 was derived by linking the two peptide chains of the IFN-gamma dimer by a seven-residue linker and changing His111 in the first chain to an aspartic acid residue. Isothermal titration calorimetry shows that IFN-gamma SC1 forms a 1:1 complex with its high-affinity receptor (IFN-gamma R alpha) with an affinity of 27(+/- 9) nM. The crystal structure of IFN-gamma SC1 has been determined at 2.9 A resolution from crystals grown in 1.4 M citrate solutions at pH 7.6. Comparison of the wild-type receptor-binding domain and the Asp111-containing domain of IFN gamma SC1 show that they are structurally equivalent but have very different electrostatic surface potentials. As a result, surface charge rather than structural changes is likely responsible for the inability of the His111-->Asp domain of to bind IFN-gamma R alpha. The AB loops of IFN-gamma SC1 adopt conformations similar to the ordered loops of IFN-gamma observed in the crystal structure of the IFN-gamma/IFN-gamma R alpha complex. Thus, IFN-gamma R alpha binding does not result in a large conformational change in the AB loop as previously suggested. The structure also reveals the final six C-terminal amino acid residues of IFN-gamma SC1 (residues 253-258) that have not been observed in any other reported IFN-gamma structures. Despite binding to only one IFN-gamma R alpha, IFN-gamma SC1 is biologically active in cell proliferation, MHC class I induction, and anti-viral assays. This suggests that one domain of IFN-gamma is sufficient to recruit IFN-gamma R alpha and IFN-gamma R beta into a complex competent for eliciting biological activity. The current data are consistent with the main role of the IFN-gamma dimer being to decrease the dissociation constant of IFN-gamma for its cellular receptors. PMID- 10860731 TI - The atomic structure of pentameric lumazine synthase from Saccharomyces cerevisiae at 1.85 A resolution reveals the binding mode of a phosphonate intermediate analogue. AB - Lumazine synthase of Saccharomyces cerevisiae is a homopentamer with a molecular weight of 90 kDa. Crystals of the recombinant enzyme with a size of up to 1.6 mm were obtained. The space group is P4(1)2(1)2 with lattice dimensions 82.9 A x 82.9 A x 300.2 A. X-ray diffraction data collected under cryogenic conditions were complete to 1.85 A resolution. The structure of the enzyme in complex with the intermediate analogue, 5-(6-D-ribitylamino-2,4-dihydroxypyrimidine-5-yl)-1 pentyl-p hosphonic acid was solved via molecular replacement using the structure of the Bacillus subtilis enzyme as search model and was refined to a final R factor of 19.8% (Rfree: 22.5%). The conformation of the active site ligand of the enzyme mimicks that of the Schiff base intermediate of the enzyme-catalyzed reaction. The data enable the reconstruction of the reactant topology during the early steps of the catalytic reaction. Structural determinants, which are likely to be responsible for the inability of the S. cerevisiae enzyme to form icosahedral capsids, will be discussed. PMID- 10860732 TI - Four crystal structures of the 60 kDa flavoprotein monomer of the sulfite reductase indicate a disordered flavodoxin-like module. AB - Escherichia coli NADPH-sulfite reductase (SiR) is a 780 kDa multimeric hemoflavoprotein composed of eight alpha-subunits (SiR-FP) and four beta-subunits (SiR-HP) that catalyses the six electron reduction of sulfite to sulfide. Each beta-subunit contains a Fe4S4 cluster and a siroheme, and each alpha-subunit binds one FAD and one FMN as prosthetic groups. The FAD gets electrons from NADPH, and the FMN transfers the electrons to the metal centers of the beta subunit for sulfite reduction. We report here the 1.94 A X-ray structure of SiR FP60, a recombinant monomeric fragment of SiR-FP that binds both FAD and FMN and retains the catalytic properties of the native protein. The structure can be divided into three domains. The carboxy-terminal part of the enzyme is composed of an antiparallel beta-barrel which binds the FAD, and a variant of the classical pyridine dinucleotide binding fold which binds NADPH. These two domains form the canonic FNR-like module, typical of the ferredoxin NADP+ reductase family. By analogy with the structure of the cytochrome P450 reductase, the third domain, composed of seven alpha-helices, is supposed to connect the FNR-like module to the N-terminal flavodoxine-like module. In four different crystal forms, the FMN-binding module is absent from electron density maps, although mass spectroscopy, amino acid sequencing and activity experiments carried out on dissolved crystals indicate that a functional module is present in the protein. Our results clearly indicate that the interaction between the FNR-like and the FMN-like modules displays lower affinity than in the case of cytochrome P450 reductase. The flexibility of the FMN-binding domain may be related, as observed in the case of cytochrome bc1, to a domain reorganisation in the course of electron transfer. Thus, a movement of the FMN-binding domain relative to the rest of the enzyme may be a requirement for its optimal positioning relative to both the FNR-like module and the beta-subunit. PMID- 10860733 TI - 1.9 A resolution crystal structure of the Saccharomyces cerevisiae Ran-binding protein Mog1p. AB - The 1.9 A resolution X-ray crystal structure of Ran-binding protein Mog1p shows that it has a unique fold based on a six-stranded antiparallel beta-sheet backed on both sides by an extensive alpha-helix. The topology of some elements of Mog1p secondary structure resemble a portion of nuclear transport factor 2 (NTF2), but the hydrophobic cavity and surrounding negatively charged residues that are important in the NTF2-RanGDP interaction are not conserved in Mog1p. In addition to binding RanGTP, Mog1p forms a 1:1 complex with RanGDP and so binds Ran independent of its nucleotide state. Mog1p and NTF2 compete for binding to RanGDP indicating that their binding sites on RanGDP are sufficiently close to prevent both proteins binding simultaneously. Although there may be some overlap between the Mog1p and NTF2 binding sites on RanGDP, these sites are not identical. Sequence analysis of Mog1p homologues from Schizosaccharomyces pombe, human, and Caenorhabditis elegans in the context of the Mog1p crystal structure indicates the presence of a cluster of highly conserved surface residues consistent with an interaction site for Ran. PMID- 10860734 TI - Direct visualisation of the beta-sheet structure of synthetic Alzheimer's amyloid. AB - Amyloid fibrils are a major pathological feature of Alzheimer's disease as well as other amyloidoses including the prion diseases. They are an unusual phenomenon, being made up of different, normally soluble proteins which undergo a profound conformational change and assemble to form very stable, insoluble fibrils which accumulate in the extracellular spaces. In Alzheimer's disease the amyloid fibrils are composed of the A beta protein. Knowledge of the structure of amyloid is essential for understanding the abnormal assembly and deposition of these fibrils and could lead to the rational design of therapeutic agents for their prevention or disaggregation. Here we reveal the core structure of an Alzheimer's amyloid fibril by direct visualisation using cryo-electron microscopy. Synthetic amyloid fibrils composed of A beta residues 11 to 25 and 1 to 42 were examined. The A beta (11-25) fibrils are clearly composed of beta sheet structure that is observable as striations across the fibres. The beta strands run perpendicular to the fibre axis and the projections show that the fibres are composed of beta-sheets with the strands in direct register. This observation has implications not only for the further understanding of amyloid, but also for the development of cryo-electron microscopy for direct visualisation of secondary structure. PMID- 10860735 TI - The final stages of folding of the membrane protein bacteriorhodopsin occur by kinetically indistinguishable parallel folding paths that are mediated by pH. AB - The folding of the transmembrane protein bacteriorhodopsin that occurs during the binding of its retinal cofactor is investigated in a membrane-like environment. Changes in the retinal absorption band reveal two transient retinal-protein intermediate states, with apparent absorption maxima at 380 nm and 440 nm, respectively. Studies on a bacteriorhodopsin mutant of Lys216, which cannot bind retinal covalently, add to evidence that retinal is non-covalently bound in these intermediate states. The two retinal-protein intermediates are genuine intermediate states that form in parallel, each with an observed rate constant of 1.1 s-1. Meanwhile no formation of the folded state is detected. Folded bacteriorhodopsin, with all trans retinal covalently bound, forms from both retinal-bound intermediates with the same apparent rate constant of 0.0070 s-1 that is independent of retinal concentration. Retinal isomerisation then occurs with a rate constant of 0.00033 s-1 to give bacteriorhodopsin containing all trans and 13 cis-retinal. These results provide experimental evidence for multiple folding routes for a membrane protein that are pH dependent, with pH conditions determining the apparent folding route. These observed parallel folding paths are kinetically indistinguishable, which contrasts with most other observations of parallel folding pathways where only pathways with different kinetics have been reported. Furthermore, together with previous work, this study shows that bacteriorhodopsin has to populate at least two folding intermediates, during folding in the mixed lipid micelles investigated here, before the final fold is attained. PMID- 10860736 TI - A small synthetic peptide, which inhibits the p53-hdm2 interaction, stimulates the p53 pathway in tumour cell lines. AB - The hdm2 protein negatively regulates p53 tumour suppressor activity. Upon binding to p53, hdm2 stimulates p53 degradation and inhibits its transcriptional activity. Moreover, the hdm2 protein is overexpressed in various tumours inactivating p53. We report here that an octamer synthetic peptide derived from p53 inhibits the p53-hdm2 interaction in vitro. In cellular assays, this untagged peptide penetrates tumour cells and induces the accumulation of p53. The accumulation of p53 leads to its activation. Two gene products transcriptionally regulated by p53, p21Waf1/Cip1 and hdm2, are induced in the presence of the peptide. When used with tumour cells that overexpress hdm2, the peptide induces the death of these tumour cells by apoptosis. The mode of action of this peptide differs from that of DNA-damaging agents (e.g. cisplatin) in that it does not induce p53 phosphorylation on serine 15. This work validates with a low molecular mass molecule our current knowledge on the regulation of the p53 pathway by the hdm2 protein. It also shows that inhibitors of the p53-hdm2 interaction are very attractive candidates for the activation of the p53 pathway in tumours expressing wild-type p53. PMID- 10860737 TI - Hydrogen bonding and catalysis: a novel explanation for how a single amino acid substitution can change the pH optimum of a glycosidase. AB - The pH optima of family 11 xylanases are well correlated with the nature of the residue adjacent to the acid/base catalyst. In xylanases that function optimally under acidic conditions, this residue is aspartic acid, whereas it is asparagine in those that function under more alkaline conditions. Previous studies of wild type (WT) Bacillus circulans xylanase (BCX), with an asparagine residue at position 35, demonstrated that its pH-dependent activity follows the ionization states of the nucleophile Glu78 (pKa 4.6) and the acid/base catalyst Glu172 (pKa 6.7). As predicted from sequence comparisons, substitution of this asparagine residue with an aspartic acid residue (N35D BCX) shifts its pH optimum from 5.7 to 4.6, with an approximately 20% increase in activity. The bell-shaped pH activity profile of this mutant enzyme follows apparent pKa values of 3.5 and 5.8. Based on 13C-NMR titrations, the predominant pKa values of its active-site carboxyl groups are 3.7 (Asp35), 5.7 (Glu78) and 8.4 (Glu172). Thus, in contrast to the WT enzyme, the pH-activity profile of N35D BCX appears to be set by Asp35 and Glu78. Mutational, kinetic, and structural studies of N35D BCX, both in its native and covalently modified 2-fluoro-xylobiosyl glycosyl-enzyme intermediate states, reveal that the xylanase still follows a double-displacement mechanism with Glu78 serving as the nucleophile. We therefore propose that Asp35 and Glu172 function together as the general acid/base catalyst, and that N35D BCX exhibits a "reverse protonation" mechanism in which it is catalytically active when Asp35, with the lower pKa, is protonated, while Glu78, with the higher pKa, is deprotonated. This implies that the mutant enzyme must have an inherent catalytic efficiency at least 100-fold higher than that of the parental WT, because only approximately 1% of its population is in the correct ionization state for catalysis at its pH optimum. The increased efficiency of N35D BCX, and by inference all "acidic" family 11 xylanases, is attributed to the formation of a short (2.7 A) hydrogen bond between Asp35 and Glu172, observed in the crystal structure of the glycosyl-enzyme intermediate of this enzyme, that will substantially stabilize the transition state for glycosyl transfer. Such a mechanism may be much more commonly employed than is generally realized, necessitating careful analysis of the pH-dependence of enzymatic catalysis. PMID- 10860738 TI - Co-evolution of proteins with their interaction partners. AB - The divergent evolution of proteins in cellular signaling pathways requires ligands and their receptors to co-evolve, creating new pathways when a new receptor is activated by a new ligand. However, information about the evolution of binding specificity in ligand-receptor systems is difficult to glean from sequences alone. We have used phosphoglycerate kinase (PGK), an enzyme that forms its active site between its two domains, to develop a standard for measuring the co-evolution of interacting proteins. The N-terminal and C-terminal domains of PGK form the active site at their interface and are covalently linked. Therefore, they must have co-evolved to preserve enzyme function. By building two phylogenetic trees from multiple sequence alignments of each of the two domains of PGK, we have calculated a correlation coefficient for the two trees that quantifies the co-evolution of the two domains. The correlation coefficient for the trees of the two domains of PGK is 0. 79, which establishes an upper bound for the co-evolution of a protein domain with its binding partner. The analysis is extended to ligands and their receptors, using the chemokines as a model. We show that the correlation between the chemokine ligand and receptor trees' distances is 0.57. The chemokine family of protein ligands and their G-protein coupled receptors have co-evolved so that each subgroup of chemokine ligands has a matching subgroup of chemokine receptors. The matching subfamilies of ligands and their receptors create a framework within which the ligands of orphan chemokine receptors can be more easily determined. This approach can be applied to a variety of ligand and receptor systems. PMID- 10860739 TI - Transcription activation by the Escherichia coli cyclic AMP receptor protein: determinants within activating region 3. AB - At Class II CRP-dependent promoters, the Escherichia coli cyclic AMP receptor protein (CRP) activates transcription by making multiple interactions with RNA polymerase (RNAP). Two discrete surfaces of CRP, known as Activating Region 1 (AR1) and Activating Region 2 (AR2), interact with the C-terminal and N-terminal domains, respectively, of the alpha subunit of RNAP. Activating Region 3 (AR3) is a third separate surface of CRP, which is thought to interact with a target in the C-terminal region of the RNAP sigma(70) subunit. We have used a CRP mutant that functions primarily via AR3, CRP HL159 KE101 KN52, as a tool to identify residues within AR3 that are important for activation. This was achieved by screening a random mutant library of the gene encoding CRP HL159 KE101 KN52 for positive control mutants at Class II CRP-dependent promoters, and also by performing alanine scanning mutagenesis. Using both in vivo reporter assays and in vitro transcription assays, we measured the effects of key substitutions within AR3 on transcription activation in both CRP HL159 KE101 KN52 and wild-type CRP. We show that a cluster of negatively charged surface-exposed residues at positions 53, 54, 55 and 58 is required for optimal activation at a Class II, but not at a Class I, CRP-dependent promoter. We conclude that these residues in AR3 of CRP form an activatory determinant for Class II transcription activation. Abortive initiation assays were used to show that this activatory determinant accelerates the rate of isomerisation from the closed to open complex at a Class II CRP-dependent promoter. AR3 of CRP also contains an inhibitory determinant: the lysine residue at position 52 of CRP is inhibitory to maximal levels of transcription activation from Class II promoters. We show that the negative effects of K52 are not simply due to "masking" of the negatively charged residues at positions 53, 54, 55 and 58. Our results suggest that, during activation by wild-type CRP, the activatory and inhibitory determinants of AR3 balance each other. Thus, activation is predominantly determined by AR1 and AR2. PMID- 10860740 TI - Interactions between activating region 3 of the Escherichia coli cyclic AMP receptor protein and region 4 of the RNA polymerase sigma(70) subunit: application of suppression genetics. AB - The Escherichia coli cyclic AMP receptor protein, CRP, induces transcription at Class II CRP-dependent promoters by making three different activatory contacts with different surfaces of holo RNA polymerase. One contact surface of CRP, known as Activating Region 3 (AR3), is functional in the downstream subunit of the CRP dimer and is predicted to interact with region 4 of the RNAP sigma(70) subunit. We have previously shown that a mutant CRP derivative that activates transcription primarily via AR3, CRP HL159 KE101 KN52, requires the positively charged residues K593, K597 and R599 in sigma(70) for activation. Here, we have used the positive control substitution, EK58, to disrupt AR3-dependent activation by CRP HL159 KE101 KN52. We then screened random mutant libraries and an alanine scan library of sigma(70) for candidates that restore activation by CRP HL159 KE101 KN52 EK58. We found that changes at R596 and R599 in sigma(70) can restore activation by CRP HL159 KE101 KN52 EK58. This suggests that the side-chains of both R596 and R599 in sigma(70) clash with K58 in CRP. Maximal activation by CRP HL159 KE101 KN52 EK58 is achieved with the substitutions RE596 or RD596 in sigma(70). We propose that there are specific charge-charge interactions between E596 or D596 in sigma(70) and K58 in AR3. Thus, no increase in activation is observed in the presence of another positive control substitution, EG58 (CRP HL159 KE101 KN52 EG58). Similarly, both sigma(70) RE596 and sigma(70) RD596 can restore activation by CRP EK58 but not CRP EG58, and they both decrease activation by wild-type CRP. We suggest that E596 and D596 in sigma(70) can positively interact with K58 in AR3, thereby enhancing activation, but negatively interact with E58, thereby decreasing activation. The substitution, KA52 in AR3 increases Class II CRP-dependent activation by removing an inhibitory lysine residue. However, this increase is not observed in the presence of either sigma(70) RE596 or sigma(70) RD596. We conclude that the inhibitory side-chain, K52 in AR3, clashes with R596 in sigma(70). Finally, we show that the sigma(70) RE596 and RD596 substitutions affect CRP-dependent activation from Class II, but not Class I, promoters. PMID- 10860741 TI - Overextended RNA:DNA hybrid as a negative regulator of RNA polymerase II processivity. AB - An eight nucleotide RNA:DNA hybrid at the 3' end of the transcript is required for the stability of the elongation complex (EC) of RNA polymerase II. A non template DNA strand is not needed for the stability of the EC, which contains this minimal hybrid. Here, we apply a recently developed method for promoter independent assembly of functional EC of RNA polymerase II from synthetic RNA and DNA oligonucleotides to study the minimal composition of the nucleic acid array required for stability of the complex with RNA longer than eight nucleotides. We found that upon RNA extension beyond 14-16 nt in the course of transcription, non template DNA becomes essential for maintaining a stable EC. Our data suggest that the overextended RNA:DNA hybrid formed in the absence the non-template DNA acts as a negative regulator of EC stability. The dissociation of the EC correlates with the backsliding of the polymerase along the overextended hybrid. The dual role of the hybrid provides a mechanism for the control of a correct nucleic acid architecture in the EC and of RNA polymerase II processivity. PMID- 10860742 TI - Changes in the 17 bp spacer in the P(R) promoter of bacteriophage lambda affect steps in open complex formation that precede DNA strand separation. AB - Tau plots and temperature-shift experiments were used to determine which step in the formation of transcriptionally-competent open complexes is affected by changing the length of the 17 bp spacer separating the -10 and -35 consensus regions of the P(R) promoter of bacteriophage lambda. Abortive initiation assays at 37 degrees C indicate that the primary effect of insertion of a base-pair, thereby increasing spacer length to 18 bp, is a decrease in k(f), the rate constant for conversion from closed (RP(c)) to open (RP(o)) complexes, by approximately a factor of 4. The mutation did not significantly affect K(B), the equilibrium constant for formation of closed complexes, and decreased K(B)k(f) by a factor of 3. Deletion of a bp to create a 16 bp spacer had a much greater effect, decreasing the measured value of k(f) by a factor of about 25 to 30, and K(B)k(f) by a factor of 7 to 8. When the values of the parameters for the deletion mutant were corrected for incomplete occupancy of RP(o) at equilibrium, the effects of the deletion were even greater. In particular, the corrected value of K(B)k(f) was about 15 times lower than the corresponding value for two promoters with wild-type spacing. Based on temperature shift experiments, the changes in spacer length did not affect the equilibrium at 20 degrees C between RP(i), a stable intermediate in which DNA strands are not separated, and RP(o). Although differential sensitivity of single-stranded bases to KMnO(4) indicated that in about 20% of the open complexes at 20 degrees C the DNA strands are not fully separated (RP(o1)), the distribution between these complexes and RP(o2) (DNA strands fully separated) was also not affected significantly by changes in spacer length. Thus, changes in spacer length primarily affect k(2), the rate constant for conversion of RP(c) to RP(i), which corresponds to a nucleation of DNA strand-separation. Application of published data and/or algorithms for determining effects of nucleotide sequence on twist angle or rise at individual bp steps does not provide a simple explanation of the difference in promoter strength between P(R) derivatives with 16 bp spacing and those with 18 bp spacing. PMID- 10860743 TI - Identification and characterisation of the selenocysteine-specific translation factor SelB from the archaeon Methanococcus jannaschii. AB - Selenocysteine insertion into archaeal selenopolypeptides is directed through an mRNA structure (the SECIS element) situated in the 3' non-translated region like in eukaryotes. To elucidate the mechanism how this element affects decoding of an in-frame UGA with selenocysteine the open reading frames of the genome of Methanococcus jannaschii were searched for the existence of a homolog to the bacterial specialized translation factor SelB. The product of the open reading frame MJ0495 was identified as the archaeal SelB homolog on the basis of the following characteristics: (1) MJ0495 possesses sequence features characteristic of bacterial SelB; (2) purified MJ0495 displays guanine nucleotide binding properties like SelB; and (3) it preferentially binds selenocysteyl-tRNA(Sec). In contrast to bacterial SelB, however, no binding of MJ0495 protein to the SECIS element of the mRNA was found under the experimental conditions employed which correlates with the fact that MJ0495 lacks the C-terminal domain of the bacterial SelB protein known to bind the SECIS element. It is speculated that in Archaea the functions of bacterial SelB are distributed over at least two proteins, one, serving as the specific translation factor, like MJ0495, and another one, binding to the SECIS which interacts with the ribosome and primes it to decode UGA. PMID- 10860744 TI - Two nuclear localization signals in the HIV-1 matrix protein regulate nuclear import of the HIV-1 pre-integration complex. AB - Replication of HIV-1 in non-dividing and slowly proliferating cell populations depends on active import of the viral pre-integration complex (PIC) into the cell nucleus. While it is commonly accepted that this process is mediated by an interaction between the HIV-1 PIC and the cellular nuclear import machinery, controversial results have been reported concerning the mechanisms of this interaction. Here, we demonstrate that a recently identified nuclear localization signal within the HIV-1 matrix protein (MA), MA NLS-2, together with previously described MA NLS-1, mediates nuclear import of the HIV-1 PIC. Inactivation of both MA NLSs precluded nuclear translocation of MA and rendered the virus defective in nuclear import and replication in non-dividing macrophage cultures, even when functional Vpr and integrase (IN), two more viral proteins implicated in HIV-1 nuclear import, were present. Taken together, these results indicate that Vpr does not function as an independent nuclear import factor and demonstrate that HIV-1 MA, by virtue of its two nuclear localization signals, regulates HIV-1 nuclear import. PMID- 10860745 TI - Biochemical analyses of the AF10 protein: the extended LAP/PHD-finger mediates oligomerisation. AB - Leukaemogenesis correlates with alterations in chromatin structure brought about by the gain or loss of interactive domains from regulatory factors that are disrupted by chromosomal translocations. The gene MLL, a target of such translocation events, forms chimaeric fusion products with a variety of partner genes. While MLL appears to be involved in chromatin-mediated gene regulation, the functions of its partner genes are largely speculative. We report the biochemical analysis of the MLL partner gene AF10 and its possible role in leukaemogenesis. AF10 has been reported to be re-arranged with genes other than MLL leading to the same phenotype, a myeloid leukaemia. We have identified a novel protein-protein interaction motif in the AF10 protein comprising the extended LAP/PHD-finger. This domain mediates homo-oligomerisation of recombinant AF10 and is conserved in several proteins, including MLL itself. AF10 binds cruciform DNA via a specific interaction with an AT-hook motif and is localised to the nucleus by a defined bipartite nuclear localisation signal in the N terminal region. PMID- 10860746 TI - Mutations at position A960 of E. coli 23 S ribosomal RNA influence the structure of 5 S ribosomal RNA and the peptidyltransferase region of 23 S ribosomal RNA. AB - The proximity of loop D of 5 S rRNA to two regions of 23 S rRNA, domain II involved in translocation and domain V involved in peptide bond formation, is known from previous cross-linking experiments. Here, we have used site-directed mutagenesis and chemical probing to further define these contacts and possible sites of communication between 5 S and 23 S rRNA. Three different mutants were constructed at position A960, a highly conserved nucleotide in domain II previously crosslinked to 5 S rRNA, and the mutant rRNAs were expressed from plasmids as homogeneous populations of ribosomes in Escherichia coli deficient in all seven chromosomal copies of the rRNA operon. Mutations A960U, A960G and, particularly, A960C caused structural rearrangements in the loop D of 5 S rRNA and in the peptidyltransferase region of domain V, as well as in the 960 loop itself. These observations support the proposal that loop D of 5 S rRNA participates in signal transmission between the ribosome centers responsible for peptide bond formation and translocation. PMID- 10860747 TI - Coordinated control of XerC and XerD catalytic activities during Holliday junction resolution. AB - Site-specific recombinases XerC and XerD function in the segregation of circular bacterial replicons. In a recombining nucleoprotein complex containing two molecules each of XerC and XerD, coordinated reciprocal switches in recombinase activity ensure that only XerC or XerD is active at any one time. Mutated recombinases that carry sub?stitutions of a catalytic arginine residue stimulate cleavage and strand exchange mediated by the partner recombinase on DNA substrates that are normally recombined poorly by the partner. This is associated with a reciprocal impairment of the recombinase's own ability to initiate catalysis. The extent of this switch in catalysis is modulated by changes in recombination site sequence and is not a direct consequence of any catalytic defect. We propose that altered interactions between the mutated proteins and their wild-type partners lead to an increased level of an alternative Holliday junction intermediate that has a conformation appropriate for resolution by the partner recombinase. The results indicate how subtle changes in protein-DNA architecture at a Holliday junction can redirect recombination outcome. PMID- 10860748 TI - Evidence from mutational specificity studies that yeast DNA polymerases delta and epsilon replicate different DNA strands at an intracellular replication fork. AB - Although polymerases delta and epsilon are required for DNA replication in eukaryotic cells, whether each polymerase functions on a separate template strand remains an open question. To begin examining the relative intracellular roles of the two polymerases, we used a plasmid-borne yeast tRNA gene and yeast strains that are mutators due to the elimination of proofreading by DNA polymerases delta or epsilon. Inversion of the tRNA gene to change the sequence of the leading and lagging strand templates altered the specificities of both mutator polymerases, but in opposite directions. That is, the specificity of the polymerase delta mutator with the tRNA gene in one orientation bore similarities to the specificity of the polymerase epsilon mutator with the tRNA gene in the other orientation, and vice versa. We also obtained results consistent with gene orientation having a minor influence on mismatch correction of replication errors occurring in a wild-type strain. However, the data suggest that neither this effect nor differential replication fidelity was responsible for the mutational specificity changes observed in the proofreading-deficient mutants upon gene inversion. Collectively, the data argue that polymerases delta and epsilon each encounter a different template sequence upon inversion of the tRNA gene, and so replicate opposite strands at the plasmid DNA replication fork. PMID- 10860749 TI - Cross-linking constraints on F-actin structure. AB - The DNase I binding loop (residues 38-52), the hydrophobic plug (residues 262 274), and the C terminus region are among the structural elements of monomeric (G ) actin proposed to form the intermonomer interface in F-actin. To test the proximity and interactions of these elements and to provide constraints on models of F-actin structure, cysteine residues were introduced into yeast actin either at residue 41 or 265. These mutations allowed for specific cross-linking of F actin between C41 and C265, C265 and C374, and C41 and C265 using dibromobimane and disulfide bond formation. The cross-linked products were visualized on SDS PAGE and by electron microscopy. Model calculations carried out for the cross linked F-actins revealed that considerable flexibility or displacement of actin residues is required in the disulfide cross-linked segments to fit these filaments into model F-actin structures. The calculated, cross-linked structures showed a better fit to the Holmes rather than the refined Lorenz model of F actin. It is predicted on the basis of such calculations that image reconstruction of electron micrographs of disulfide cross-linked C41-C374 F-actin should provide a conclusive test of these two similar models of F-actin structure. PMID- 10860750 TI - Influence of transfer RNA tertiary structure on aminoacylation efficiency by glutaminyl and cysteinyl-tRNA synthetases. AB - The position of the tertiary Levitt pair between nucleotides 15 and 48 in the transfer RNA core region suggests a key role in stabilizing the joining of the two helical domains, and in maintaining the relative orientations of the D and variable loops. E. coli tRNA(Gln) possesses the canonical Pu15-Py48 trans pairing at this position (G15-C48), while the tRNA(Cys) species from this organism instead features an unusual G15-G48 pair. To explore the structural context dependence of a G15-G48 Levitt pair, a number of tRNA(Gln) species containing G15 G48 were constructed and evaluated as substrates for glutaminyl and cysteinyl tRNA synthetases. The glutaminylation efficiencies of these mutant tRNAs are reduced by two to tenfold compared with native tRNA(Gln), consistent with previous findings that the tertiary core of this tRNA plays a role in GlnRS recognition. Introduction of tRNA(Cys) identity nucleotides at the acceptor and anticodon ends of tRNA(Gln) produced a tRNA substrate which was efficiently aminoacylated by CysRS, even though the tertiary core region of this species contains the tRNA(Gln) G15-C48 pair. Surprisingly, introduction of G15-G48 into the non-cognate tRNA(Gln) tertiary core then significantly impairs CysRS recognition. By contrast, previous work has shown that CysRS aminoacylates tRNA(Cys) core regions containing G15-G48 with much better efficiency than those with G15-C48. Therefore, tertiary nucleotides surrounding the Levitt pair must significantly modulate the efficiency of aminoacylation by CysRS. To explore the detailed nature of the structural interdependence, crystal structures of two tRNA(Gln) mutants containing G15-G48 were determined bound to GlnRS. These structures show that the larger purine ring of G48 is accommodated by rotation into the syn position, with the N7 nitrogen serving as hydrogen bond acceptor from several groups of G15. The G15-G48 conformations differ significantly compared to that observed in the native tRNA(Cys) structure bound to EF-Tu, further implicating an important role for surrounding nucleotides in maintaining the integrity of the tertiary core and its consequent ability to present crucial recognition determinants to aminoacyl-tRNA synthetases. PMID- 10860751 TI - The 1.5 A resolution crystal structure of the carbamate kinase-like carbamoyl phosphate synthetase from the hyperthermophilic Archaeon pyrococcus furiosus, bound to ADP, confirms that this thermostable enzyme is a carbamate kinase, and provides insight into substrate binding and stability in carbamate kinases. AB - Carbamoyl phosphate (CP), an essential precursor of arginine and the pyrimidine bases, is synthesized by CP synthetase (CPS) in three steps. The last step, the phosphorylation of carbamate, is also catalyzed by carbamate kinase (CK), an enzyme used by microorganisms to produce ATP from ADP and CP. Although the recently determined structures of CPS and CK show no obvious mutual similarities, a CK-like CPS reported in hyperthermophilic archaea was postulated to be a missing link in the evolution of CP biosynthesis. The 1.5 A resolution structure of this enzyme from Pyrococcus furiosus shows both a subunit topology and a homodimeric molecular organization, with a 16-stranded open beta-sheet core surrounded by alpha-helices, similar to those in CK. However, the pyrococcal enzyme exhibits many solvent-accessible ion-pairs, an extensive, strongly hydrophobic, intersubunit surface, and presents a bound ADP molecule, which does not dissociate at 22 degrees C from the enzyme. The ADP nucleotide is sequestered in a ridge formed over the C-edge of the core sheet, at the bottom of a large cavity, with the purine ring enclosed in a pocket specific for adenine. Overall, the enzyme structure is ill-suited for catalyzing the characteristic three-step reaction of CPS and supports the view that the CK-like CPS is in fact a highly thermostable and very slow (at 37 degrees C) CK that, in the extreme environment of P. furiosus, may have the new function of making, rather than using, CP. The thermostability of the enzyme may result from the extension of the hydrophobic intersubunit contacts and from the large number of exposed ion-pairs, some of which form ion-pair networks across several secondary structure elements in each enzyme subunit. The structure provides the first information on substrate binding and catalysis in CKs, and suggests that the slow rate at 37 degrees C is possibly a consequence of slow product dissociation. PMID- 10860752 TI - Crystal structure of a pol alpha family DNA polymerase from the hyperthermophilic archaeon Thermococcus sp. 9 degrees N-7. AB - The 2.25 A resolution crystal structure of a pol alpha family (family B) DNA polymerase from the hyperthermophilic marine archaeon Thermococcus sp. 9 degrees N-7 (9 degrees N-7 pol) provides new insight into the mechanism of pol alpha family polymerases that include essentially all of the eukaryotic replicative and viral DNA polymerases. The structure is folded into NH(2)- terminal, editing 3' 5' exonuclease, and polymerase domains that are topologically similar to the two other known pol alpha family structures (bacteriophage RB69 and the recently determined Thermococcus gorgonarius), but differ in their relative orientation and conformation. The 9 degrees N-7 polymerase domain structure is reminiscent of the "closed" conformation characteristic of ternary complexes of the pol I polymerase family obtained in the presence of their dNTP and DNA substrates. In the apo-9 degrees N-7 structure, this conformation appears to be stabilized by an ion pair. Thus far, the other apo-pol alpha structures that have been determined adopt open conformations. These results therefore suggest that the pol alpha polymerases undergo a series of conformational transitions during the catalytic cycle similar to those proposed for the pol I family. Furthermore, comparison of the orientations of the fingers and exonuclease (sub)domains relative to the palm subdomain that contains the pol active site suggests that the exonuclease domain and the fingers subdomain of the polymerase can move as a unit and may do so as part of the catalytic cycle. This provides a possible structural explanation for the interdependence of polymerization and editing exonuclease activities unique to pol alpha family polymerases. We suggest that the NH(2)-terminal domain of 9 degrees N-7 pol may be structurally related to an RNA-binding motif, which appears to be conserved among archaeal polymerases. The presence of such a putative RNA- binding domain suggests a mechanism for the observed autoregulation of bacteriophage T4 DNA polymerase synthesis by binding to its own mRNA. Furthermore, conservation of this domain could indicate that such regulation of pol expression may be a characteristic of archaea. Comparion of the 9 degrees N-7 pol structure to its mesostable homolog from bacteriophage RB69 suggests that thermostability is achieved by shortening loops, forming two disulfide bridges, and increasing electrostatic interactions at subdomain interfaces. PMID- 10860753 TI - The crystal structure of the penicillin-binding protein 2x from Streptococcus pneumoniae and its acyl-enzyme form: implication in drug resistance. AB - Penicillin-binding proteins (PBPs), the primary targets for beta-lactam antibiotics, are periplasmic membrane-attached proteins responsible for the construction and maintenance of the bacterial cell wall. Bacteria have developed several mechanisms of resistance, one of which is the mutation of the target enzymes to reduce their affinity for beta-lactam antibiotics. Here, we describe the structure of PBP2x from Streptococcus pneumoniae determined to 2.4 A. In addition, we also describe the PBP2x structure in complex with cefuroxime, a therapeutically relevant antibiotic, at 2.8 A. Surprisingly, two antibiotic molecules are observed: one as a covalent complex with the active-site serine residue, and a second one between the C-terminal and the transpeptidase domains. The structure of PBP2x reveals an active site similar to those of the class A beta-lactamases, albeit with an absence of unambiguous deacylation machinery. The structure highlights a few amino acid residues, namely Thr338, Thr550 and Gln552, which are directly related to the resistance phenomenon. PMID- 10860754 TI - Protein packing: dependence on protein size, secondary structure and amino acid composition. AB - We have used the occluded surface algorithm to estimate the packing of both buried and exposed amino acid residues in protein structures. This method works equally well for buried residues and solvent-exposed residues in contrast to the commonly used Voronoi method that works directly only on buried residues. The atomic packing of individual globular proteins may vary significantly from the average packing of a large data set of globular proteins. Here, we demonstrate that these variations in protein packing are due to a complex combination of protein size, secondary structure composition and amino acid composition. Differences in protein packing are conserved in protein families of similar structure despite significant sequence differences. This conclusion indicates that quality assessments of packing in protein structures should include a consideration of various parameters including the packing of known homologous proteins. Also, modeling of protein structures based on homologous templates should take into account the packing of the template protein structure. PMID- 10860755 TI - Enhanced genome annotation using structural profiles in the program 3D-PSSM. AB - A method (three-dimensional position-specific scoring matrix, 3D-PSSM) to recognise remote protein sequence homologues is described. The method combines the power of multiple sequence profiles with knowledge of protein structure to provide enhanced recognition and thus functional assignment of newly sequenced genomes. The method uses structural alignments of homologous proteins of similar three-dimensional structure in the structural classification of proteins (SCOP) database to obtain a structural equivalence of residues. These equivalences are used to extend multiply aligned sequences obtained by standard sequence searches. The resulting large superfamily-based multiple alignment is converted into a PSSM. Combined with secondary structure matching and solvation potentials, 3D PSSM can recognise structural and functional relationships beyond state-of-the art sequence methods. In a cross-validated benchmark on 136 homologous relationships unambiguously undetectable by position-specific iterated basic local alignment search tool (PSI-Blast), 3D-PSSM can confidently assign 18 %. The method was applied to the remaining unassigned regions of the Mycoplasma genitalium genome and an additional 13 regions were assigned with 95 % confidence. 3D-PSSM is available to the community as a web server: http://www.bmm.icnet.uk/servers/3dpssm PMID- 10860756 TI - Mutational investigation of the specificity determining region of the Src SH2 domain. AB - SH2 domains are protein modules which bind tyrosine phosphorylated sequences in many signaling pathways. These domains contain two regions with specialized functions: residues in one region form a deep pocket into which the phosphotyrosine of the target inserts, while the other region contains the so called "specificity determining residues" which interact with the three residues C-terminal to the phosphotyrosine in the target. Here, titration calorimetry and site-directed mutagenesis have been used to probe the importance of eight specificity determining residues of the SH2 domain of the Src kinase involved in contacts with its tyrosine phosphorylated consensus peptide target (sequence pYEEI where pY indicates a phosphotyrosine). Mutating six of these eight residues to Ala individually, resulted in a threefold or less loss in binding affinity; hence the majority of the residues in the specificity determining region are by themselves of minimal importance for binding. Two residues were found to have significant effects on binding: Tyr betaD5 and Lys betaD3. Tyr betaD5 was the most crucial residue as evidenced by the 30-fold loss in affinity when Tyr betaD5 is mutated to Ile. However, while this mutation eliminated the specificity of the Src SH2 domain for the pYEEI peptide sequence, it was not sufficient to switch the specificity of the Src SH2 domain to that of a related SH2 domain which has an Ile at the betaD5 position. Mutation of Lys betaD3 to an Ala residue resulted in a modest reduction in binding affinity (sevenfold). It is interesting that this mutation resulted in a change of specificity affecting the selection of the +1 position residue C-terminal to the phosphotyrosine. Except for the Lys betaD3 +1 Glu interaction which is significantly coupled, only weak energetic coupling was observed across the binding interface, as assessed using double mutant cycles. The results of this study suggest that interactions involving the specificity determining region of SH2 domains may be insufficient by themselves to target single SH2 domains to particular phosphorylated sites. PMID- 10860757 TI - Conformational analysis of long spacers in PROSITE patterns. AB - To determine if variable sequences (spacers) between conserved positions in a sequence motif or pattern share a consensus structure, three-dimensional structures containing PROSITE patterns with spacers of fixed length greater than three residues were analyzed. Structural similarities of a given pattern were evaluated by computing the backbone phi, psi and side-chain chi1 dihedral order parameters. The exact bias information in analyzing the conformational variability of the patterns was taken into account by introducing a new parameter, the bias coefficient, which describes the number and distribution of residue types found at each position of a pattern in the structures. The results of the analyses show that backbone conformational heterogeneity at a given position in a sequence motif does not necessarily correlate with the residue-type variability at that position, and the long spacer region can adopt a well-defined backbone conformation, in addition to the conserved residues. Furthermore, a PROSITE pattern may be redefined to yield two or more "refined" regular expressions, each corresponding to a distinct backbone conformation. A way in which the observed structural consensus in a pattern may be employed to improve the accuracy of function prediction from sequence is suggested. PMID- 10860758 TI - Canine coronary microvessel NO production regulates oxygen consumption in ecNOS knockout mouse heart. AB - We have previously shown that NO production by tissues following stimulation with bradykinin or other agonists can regulate oxygen consumption in skeletal muscle, heart and kidney. From those studies and from those using agonists, which classically release NO from blood vessels and which are unable to regulate tissue oxygen consumption in heart from ecNOS knockout mice, we concluded that vascular NO production is capable of regulating tissue oxygen consumption. The goal of these studies was to directly address the concept that NO production by blood vessels can regulate tissue oxygen consumption using a classical transfer paradigm. Microvessels, capable of producing NO, were prepared from canine hearts using a sieving technique, cardiac tissue was taken from mice lacking the ability to produce NO from ecNOS (ecNOS -/- mice) and tissue oxygen consumption measured in vitro using a Clark type electrode in a sealed chamber. Bradykinin (10(-7)to 10(-4)M) had no effect on tissue oxygen consumption when administered to heart from ecNOS -/mice as expected and no effect on oxygen consumption by isolated canine coronary microvessels (0+/-5% at 10(-5)M). However when coronary microvessels were co-incubated with heart from ecNOS -/- mice, bradykinin caused a dose dependent reduction in tissue oxygen consumption reaching a maximum of 44+/-10% at 10(-4)M. The effects of bradykinin were entirely abolished by L NAME. The calculated concentration range for NO in these studies was 2.9 to 293 n M, within estimated physiologic range for the activity of NO on cytochrome oxidase. These data indicate that coronary microvessels can regulate cardiac oxygen consumption through a NO dependent mechanism. PMID- 10860759 TI - Cardioprotective effects of 17 beta-estradiol produced by activation ofmitochondrial ATP-sensitive K(+)Channels in canine hearts. AB - We have previously demonstrated the effects of estrogen on modulation of myocardial ATP-sensitive K(+)(K(ATP)) channel. Previous studies have demonstrated that activation of mitochondrial K(ATP)channel is a major contributor of ischemic cardioprotection. The purpose of the present study was to investigate the role of K(ATP)channel in estrogen-induced myocardial protection after ischemia/reperfusion in dogs. Anaesthetized dogs were subjected to 60 min of left anterior descending coronary artery occlusion followed by 2 h of reperfusion. In a first study to characterize effects of sex and the dose-response profile of estrogen on infarct size, the drug was intravenously administered at 10 or 20 microg/kg. In a second study to investigate the cardioprotective mechanisms of estrogen, vehicle, preconditioning or 17 beta -estradiol (10 microg/kg) was given, beginning 15 min prior to the 60 min occlusion period in the presence or absence of 5-hydroxydecanoate (5-HD). In the first study, administration of 17 beta -estradiol resulted in a significant, dose-dependent limitation of infarct size. Estrogen administration provided myocardial protection of similar magnitude in both males and females. In the second study, infarct size in control animals averaged 39+/-5% of the risk region, compared with 14+/-5% of the risk region in estrogen-treated dogs and 6+/-5% of the risk region in preconditioning dogs (both P<0.0001 v controls). Pretreatment with 5-HD completely abolished preconditioning and estrogen-induced cardioprotection. Estrogen limits myocardial infarction size resulting from coronary artery occlusion and reperfusion in a dose-dependent fashion, irrespective of gender difference. The infarct size-limiting effect of estrogen++ was abolished by 5-HD, suggesting that the cardioprotective effect of estrogen may result from activation of myocardial mitochondrial K(ATP)channels. PMID- 10860760 TI - Exogenous nitric oxide can trigger a preconditioned state through a free radical mechanism, but endogenous nitric oxide is not a trigger of classical ischemic preconditioning. AB - Nitric oxide (NO) has been reported to play an important role in the late phase of ischemic preconditioning (PC) in the rabbit heart. However, the role of NO in the early phase of ischemic PC ("classical PC") is controversial. Accordingly, the present study was designed to determine whether NO contributes to the cardioprotective effect of classical PC in rabbits. Isolated hearts experienced 30 min of regional ischemia followed by 120 min of reperfusion. Infarct size was measured with triphenyltetrazolium chloride. In control hearts infarction was 30.2+/-3.3% of the risk zone. PC with 5 min of global ischemia and 10 min of reperfusion reduced infarct size to 10.2+/-2.4% (P<0.05). Perfusion with 2 microm S-nitroso-N-acetylpenicillamine (SNAP), a NO donor, in lieu of ischemia mimicked PC (4.4+/-1.9% infarction, P<0.01 v control). To determine whether this protection was dependent on either protein kinase C (PKC) as has previously been demonstrated for classical PC or free radicals known to be produced during exogenous NO administration, chelerythrine (5 microm), a PKC inhibitor, or N-(2 mercaptopropionyl)-glycine (300 microm), a free radical scavenger, was administered with or shortly after SNAP. Neither drug had any independent effect on infarct size, and each blocked SNAP's cardioprotection (31.0+/-5.1 and 25.7+/ 5.2% infarction, resp.). N(omega)-nitro- L -arginine methyl ester (L -NAME, 100 microm), a NO synthase inhibitor, failed to block the cardioprotection from the above ischemic PC protocol (9.5+/-2.8% infarction, P<0.05 v control). L -NAME alone had no effect on infarct size (30.6+/-2.7%). These results suggest that the beneficial effect of exogenous NO production during SNAP pretreatment is mediated by a protein kinase C-dependent pathway via MPG-sensitive oxidants. However, we were unable to show any contribution of endogenous NO to classical PC's protection in isolated rabbit hearts. PMID- 10860761 TI - Late post-myocardial infarction induces a tetrodotoxin-resistant Na(+)Current in rat cardiomyocytes. AB - Left ventricular remodeling after myocardial infarction is accompanied by electrical abnormalities that might predispose to rhythm disturbances. To get insight into the ionic mechanisms involved, we studied myocytes isolated from four different regions of the rat ventricles, 4-6 months after ligation of the left coronary artery. Using the whole-cell patch-clamp technique, we never observed T-type Ca(2+)current in both diseased and control hearts. In contrast, in 41 out of 78 cells isolated from 16 post-myocardial infarcted rats, analysed in the presence of 30 m m Na(+)ions, we found a tetrodotoxin (TTX)-resistant Na(+)current with quite variable amplitude in every investigated region. Albeit being resistant to 100 microM TTX, this Na(+)-dependent current was highly sensitive to lidocaine since 3 microM lidocaine induced about 65% tonic block. It was also inhibited by 5 microM nifedipine and 2 m m Co(2+), but was insensitive to 100 microM Ni(2+). The TTX-resistant Na(+)channel availability was shifted rightward by 25-30 mV with respect to TTX-sensitive Na(+)current; therefore, a large "window current" might flow in the voltage range from -70 to -20 mV. In conclusion, in late post-myocardial infarction, a Na(+)current with specific kinetics and pharmacology may provide inward charges in a critical range of membrane voltages that are able to alter action potential time course and trigger ventricular arrhythmia. These apparent new characteristics of the Na(+)channel might result in part from environmental changes during heart remodeling. PMID- 10860762 TI - Slowly inactivating component of sodium current in ventricular myocytes is decreased by diabetes and partially inhibited by known Na(+)-H(+)Exchange blockers. AB - Recent evidence has suggested a major role for a slowly inactivating component of Na(+)current (I(NaL)) as a contributor to ischemic Na(+)loading. The purposes of this study were to investigate veratrine and lysophosphatidylcholine (LPC) induced I(NaL)in single ventricular myocytes of normal and diabetic rats and to analyse the effects on this current of three pharmacological agents, known as Na(+)/H(+)exchange inhibitors, whose selectivity has been questioned in several studies. A decrease in Na(+)/H(+)exchange activity has been previously shown to be associated with diabetes, and this has been found to confer some protection to the diabetic heart after an episode of ischemia/reperfusion. Recordings were made using the whole-cell patch-clamp technique. I(NaL)was stimulated either by veratrine (100 mg/ml) or by LPC (10 micromol/l) applied extracellularly. Veratrine as well as LPC-induced I(NaL)was found to be significantly decreased in ventricular myocytes isolated from diabetic rat hearts. Veratrine- and LPC induced I(NaL)in ventricular myocytes of normal rats was significantly (in the range 10(-7)to 10(-4)mol/l) inhibited by the Na(+)/H(+)exchange blockers HOE 694, EIPA and HOE 642. HOE 694 was the most potent inhibitor, followed by the amiloride derivative EIPA and HOE 642. The sensitivity of veratrine-induced I(NaL)to inhibition by HOE 694 and EIPA was markedly reduced in diabetic ventricular myocytes, with no observed inhibition by HOE 642. These data may have important implications as to the protection that may be afforded against ischemic and reperfusion injury, especially during ischemia and when ischemia occurs in a diabetic situation. PMID- 10860763 TI - The alpha(1)-adrenoceptor subtype- and protein kinase C isoform-dependence of Norepinephrine's actions in cardiomyocytes. AB - Catecholamines modulate cardiac function at least in part through alpha(1) adrenergic receptors linked to the activation of protein kinase C (PKC). This study examines the molecular forms of the alpha(1)-receptor and PKC that mediate norepinephrine's actions in cardiomyocytes; distinct approaches (activation dependent down-regulation of PKC isoforms) and novel reagents (A61603, an alpha(1A/c)-receptor agonist) are used to resolve this issue which has been the focus of dispute in previous studies. Norepinephrine (NE) induces a rise in diacylglycerol levels which is sustained for 24 h and is associated with the translocation (at 5 min) and down-regulation (at 24 h) of PKC delta and PKC xi (but not PKC alpha). The selective targeting of the alpha(1)-adrenergic receptor to activate novel PKC isoforms is remarkable, given an 8-fold greater abundance of PKC alpha relative to PKC xi in this preparation. NE activates the extracellular signal-regulated protein kinase (ERK) subfamily of mitogen activated protein kinases through a PKC delta/PKC xi -dependent pathway. WB-4101 (alpha(1A/c)- and alpha(1D)-receptor antagonist) and 5-methylurapidil (alpha(1A/c)-receptor antagonist) inhibit norepinephrine-dependent accumulation of inositol phosphate and diacylglycerol, down-regulation of PKC delta and PKC xi, and activation of ERK. Each of these responses is stimulated by A61603, but not attenuated by high concentrations of chloroethylclonidine (which irreversibly inactivates the alpha(1B)-, and to a lesser extent, the alpha(1D)-receptor) or BMY 7378 (selective alpha(1D)-receptor antagonist). A61603 also activates p38 MAPK and induces hypertrophy. These studies establish that NE's actions in cardiomyocytes can be attributed to the alpha(1A/c)-adrenergic receptor subtype and nPKC isoforms, thereby identifying specific targets for the development of pharmaceuticals to influence cardiac contractile function and/or growth responses. PMID- 10860764 TI - Characterization of G-protein signaling in ventricular myocytes from the adult mouse heart: differences from the rat. AB - We have developed a high yield technique for isolating ventricular myocytes from adult mouse hearts. This collagenase-trypsin procedure yields 3 6x10(6)cells/heart. The cells are rod-shaped, roughly 20 microM x 100 microM and Ca(++)tolerant, with viability of 65-80%. Binding studies with [(125)I]ICYP demonstrate the presence of beta -adrenergic receptors at a density of 83 fmol/mg membrane protein. Assessment of the effects of the beta(1)-specific antagonist CGP 20712A on [(125)I]ICYP binding and on isoproterenol (ISO)-sensitive adenylyl cyclase activity indicates that 67% of the receptors are beta(1)and 33% are beta(2), compared to 16-20%beta(2)in rat myocytes. Mouse myocytes respond to isoproterenol to produce cyclic AMP with an EC(50) approximately 110+/-20 n M. A functional G(i)pathway is demonstrated by inhibition of ISO-stimulated cyclic AMP accumulation by endothelin, carbachol and ATP and by sensitivity of this inhibition to pertussis toxin. As assessed by inositol phosphate production, endothelin and ATP stimulate the activity of the G(q)-phospholipase C pathway, whereas carbachol, PGF(2 alpha)and alpha(1)-adrenergic receptor agonists show no significant effect. The inability of alpha(1)-adrenergic receptor agonists to induce phosphoinositide hydrolysis in mouse myocytes differs from a several fold alpha(1)-adrenergic activation that occurs in rat. Biochemical and pharmacological profiles, as well as the need for modifications in experimental design, indicate that mouse myocytes differ substantially from rat cardiac myocytes. PMID- 10860765 TI - Intracellular free calcium and mitochondrial membrane potential in ischemia/reperfusion and preconditioning. AB - Moderation of calcium perturbations has been implicated in ischemic preconditioning. As mitochondria possess an effective Ca(2+)transporting system driven by the mitochondrial membrane potential, experiments were performed to study time-averaged intracellular free calcium and the mitochondrial membrane potential during preconditioning and ischemia-reperfusion. Isolated rat hearts were subjected to 5 min of preconditioning, a 9-min intervening reperfusion and 21 min of ischemia with subsequent reperfusion. The hearts were preloaded with the Ca(2+)indicator Fura-2 or the mitochondrial membrane potential probe safranine. A method was devised for correction for NADH autofluorescence in time averaged Ca(2+)probing with Fura-2. The pH dependence of the apparent dissociation constant of the Ca(2+)complex of Fura-2 was determined. Intracellular free Ca(2+)increased during the 5-min ischemia, and this was reversed upon reperfusion. During protracted ischemia a continual Ca(2+)rise was observed when the fluorescence data were corrected for changes in pH. An initial sharp Fura-2 fluorescence spike upon final reperfusion was caused by a pH dependent change in the dissociation constant of the Ca(2+)complex of Fura-2. In preconditioned hearts the free Ca(2+)was somewhat lower during reperfusion, but a major effect of preconditioning was observed during the prolonged ischemia. The decrease in mitochondrial membrane potential during prolonged ischemia was faster in the preconditioned heart with no difference during the final reperfusion. The effect of preconditioning on cell survival was reflected in a decrease in the post-ischemic washout of creatine kinase. The moderation of the ischemic and post ischemic intracellular Ca(2+)increase, and the acceleration of the ischemic mitochondrial membrane potential decrease by ischemic preconditioning is in accord with the notion of the involvement of mitochondrial ATP sensitive K(+)channels in preconditioning. In studies on ischemia it is absolutely necessary to correct for the pH-sensitivity of the apparent dissociation constant of the calcium complex of Fura-2 to obtain reliable data for intracellular free calcium. PMID- 10860766 TI - Increased susceptibility to development of triggered activity in myocytes from mice with targeted disruption of endothelial nitric oxide synthase. AB - Nitric oxide generated by cardiac myocytes or delivered by drugs has been shown to regulate cardiac contractile function and has been implicated in suppressing some cardiac arrhythmias, although this remains controversial. We examined the ability of the soluble cardiac glycoside, ouabain, to trigger arrhythmic contractions in ventricular myocytes isolated from mice lacking a functional endothelial nitric oxide synthase gene (eNOS(null)). Arrhythmic activity, defined as aftercontractions, was induced with ouabain (50 micromol/L) and recorded using a video-motion detector in isolated, electrically driven single ventricular myocytes from adult eNOS(null)or from their wild-type (WT) littermates. The rate of ouabain-induced arrhythmic contractions was significantly higher in eNOS(null)myocytes than in WT myocytes. Application of the NO donor S-nitroso acetylcysteine (SNAC) significantly diminished the frequency of arrhythmic contractions in eNOS(null)myocytes. The antiarrhythmic effect of NO, whether generated by eNOS in WT cells or by SNAC, could be partially reversed by 1H [1,2,4]oxadiazolo-[4, 3-a]- quinoxalin-1-one (ODQ), a specific soluble guanylyl cyclase inhibitor. Ouabain significantly increased intracellular cGMP in WT but not eNOS(null)hearts, and this cGMP response was blocked by ODQ. Since cardiac glycoside- induced aftercontractions are activated by the transient inward current (I(ti)), the role of NO in ouabain (100 micromol/L)- induced I(ti)was examined using the nystatin-perforated patch-clamp technique. The frequency of ouabain-induced I(ti)was significantly higher in eNOS(null)myocytes than in WT myocytes, and this could be suppressed by SNAC. These data demonstrate that NO derived from myocyte eNOS activation suppresses ouabain-induced arrhythmic contractions by a mechanism that might involve activation of guanylyl cyclase and elevation of cGMP. PMID- 10860767 TI - Different regulation of cardiac and renal corticosteroid receptors in aldosterone salt treated rats: effect of hypertension and glucocorticoids. AB - This study analysed the regulation of cardiac mineraloreceptor (MR) and glucoreceptor (GR) in aldosterone-salt treatment (AST). AST causes hypertension, left ventricle (LV) hypertrophy and decreases plasma corticosterone level. Ribonuclease protection assay and Western blot analysis showed a rise of MR mRNA (1.5- and 1.4-fold at day 15 and 30, respectively) and protein levels (1.8- and 4.1-fold at day 30 and 60, respectively) in the LV, but not in either the right ventricle (RV) or in kidney of treated rats. Addition of MR antagonist spironolactone (20 mg/kg/day) for 30 days failed to prevent these changes but was able to reduce AST-induced cardiac fibrosis. Similar hypertension-induced MR upregulations were observed in the LV of AngII-hypertensive rats and of 12-week old SHR when compared to 4-week-old prehypertensive SHR. AST also enhanced left ventricular GR mRNA (2.0- and 3.0-fold at day 7 and 15, respectively) and protein contents (2.0- and 1.7-fold at day 30 and 60, respectively). In contrast to MR, GR levels were also upregulated in both RV and kidney. Such an upregulation was equally observed at mRNA and protein levels in LV, RV and kidney after adrenalectomy (15 days) and was prevented in both tissues after glucocorticoid replacement (adrenalectomy + dexamethasone at 100 micro g/kg/day for 15 days). Therefore, MR level may be controlled by hemodynamical factors whereas that of GR depends upon glucocorticoids level. PMID- 10860768 TI - Exogenous neopterin causes cardiac contractile dysfunction in the isolated perfused rat heart. AB - Neopterin is known in humans as a sensitive marker for diseases associated with increased activity of the cellular immune system. Recent studies report neopterin also to exhibit distinct effects: neopterin induces inducible nitric oxide synthase expression in rat vascular smooth muscle cells and activates translocation of nuclear factor- kappa B. Neopterin may also induce oxidative stress causing apoptotic cell death, or superinduce tumor necrosis factor- alpha mediated apoptosis. Observing these effects in cell cultures, we were interested in possible consequences of neopterin on cardiac function in the isolated perfused rat heart. The influence of neopterin in three different concentrations (10 micromol/l, 50 micromol/l, 100 micromol/l) on cardiac contractility parameters and coronary vascular resistance were studied in 67 male Sprague Dawley rats using the temperature-controlled and pressure-constant Langendorff apparatus with retrograde perfusion of the aorta with a Krebs-Henseleit buffer. Treatment with 100 micromol/l neopterin resulted in a significant decrease in coronary flow and cardiac contractility. Coronary flow decreased from 15.2 to 9.5 ml/min (P=0.002), left ventricular pressure from 80 to 52 mmHg (P=0. 002), rate of pressure fall from 1605 to 923 mmHg/s (P=0.001) and rate of pressure rise from 2862 to 1709 mmHg/s (P=0.001). Concentrations lower than 100 micromol/l neopterin had no significant effect on cardiac function. Our study demonstrates a considerable influence of exogenous neopterin on cardiac performance in the Langendorff model of isolated perfused rat hearts. This has to be considered a potential pathogenic factor of cardiac disturbances in diseases in which high concentrations of neopterin are released due to immune activation. At present the exact mechanism remains unclear. PMID- 10860769 TI - Isolation and characterization of the murine cardiotrophin-1 gene: expression and norepinephrine-induced transcriptional activation. AB - Cardiotrophin-1 (CT-1) is a novel cytokine capable of inducing hypertrophy in cardiac myocytes and belongs to the interleukin-6 family that exert their biological effects through gp130. To clarify the involvement and pathophysiological role of CT-1 in myocardial diseases, it is important to characterize the regulation of CT-1 gene expression. In this study, we isolated and characterized the mouse CT-1 gene and studied the expression of CT-1 mRNA under norepinephrine (NE) stimulation. The mouse CT-1 gene constitutes 5.4 kilobases (kb) in length and consists of three exons and two introns. When nucleotide sequences of the coding regions of exons were compared with those of human, exon 1, 2 and 3 share 96%, 84% and 81% homology, respectively. The 2.2 kb of 5; flanking lesion of the mouse CT-1 gene contains a variety of transcription factor binding motif (e.g. CREB, MyoD, NF-IL6, Nkx2.5, GATA). Fluorescent in situ hybridization (FISH) analysis demonstrated that the mouse CT-1 gene was located on chromosome 7F3. The expression of CT-1 mRNA in cardiac myocytes was markedly augmented by NE stimulation, both in vivo and in vitro. Promoter analysis using deletion constructs of the CT-1 gene indicated that the NE responsive element located between -2174/-1540 and this region contained the cAMP responsive element (CRE). Electrophoretic gel mobility shift assays showed enhanced binding activity to the CRE motif in the nuclear extracts from NE-stimulated cardiac myocytes. These studies indicate that CT-1 is abundantly expressed in the heart and that the CRE is a possible cis -acting element of the CT-1 gene under NE-stimulation. These data suggest that the CT-1 gene expression is regulated, at least partially, by transcriptional machinery. PMID- 10860770 TI - Loading of calcium and strontium into the sarcoplasmic reticulum in rat ventricular muscle. AB - Previous work suggests that strontium ions (Sr(2+)) are less effective than calcium ions (Ca(2+)) at supporting excitation-contraction (EC) coupling in cardiac muscle. We therefore tested whether this was due to differences in the uptake and release of Ca(2+)and Sr(2+)by the sarcoplasmic reticulum (SR) of rat ventricular trabeculae and myocytes at 22-24 degrees C. In permeabilized trabeculae, isometric contractions activated by exposure to Ca(2+)- and Sr(2+) containing solutions produced similar maximal force, but were four times more sensitive to Ca(2+)than to Sr(2+). The rate of loading and maximal SR capacity for caffeine-releasable Ca(2+)and Sr(2+)were similar. In isolated, voltage clamped ventricular myocytes, the SR content was measured as Na(+)-Ca(2+)exchange current during caffeine-induced SR cation releases. The SR Ca(2+)load reached a steady maximum during a train of voltage clamp depolarizations. A similar maximal Sr(2+)load was not observed, suggesting that the SR capacity for Sr(2+)exceeds that for Ca(2+). Therefore, the relative inability of Sr(2+)to support cardiac EC coupling appears not to be due to failure of the SR to sequester Sr(2+). Instead, increases in cytosolic [Sr(2+)] seem to poorly activate Sr(2+)release from the SR. PMID- 10860771 TI - Differential translocation or phosphorylation of alpha B crystallin cannot be detected in ischemically preconditioned rabbit cardiomyocytes. AB - Alpha B Crystallin (alpha BC) is a putative effector protein of ischemic preconditioning (IPC), that is phosphorylated on Ser 45 by ERK1/2 and Ser 59 by the p38 MAPK substrate, MAPKAPK-2. Translocation and phosphorylation of alpha BC was determined in cytosolic and cytoskeletal fractions by 1D SDS-PAGE and IEF, or using Ser 45 and Ser 59 phospho-specific antibodies in: (1) control rabbit cardiomyocytes; (2) cells preconditioned by 10 min in vitro ischemia; or after pre-treatment with specific inhibitors of (3) Ser/Thr protein phosphatase 1/2A (calyculin A); (4) p38 MAPK (SB203580); or (5) ERK 1/2 (PD98059); all prior to 180 min ischemia. Ischemia induced a cytosolic to cytoskeletal translocation of alpha BC, which was similar in all the groups. Highly phosphorylated isoforms (D1/2) of alpha BC were present in cytosolic but not cytoskeletal fractions at 0 min ischemia. By 60-90 min ischemia, D1/2 isoforms had translocated to the cytoskeletal fraction. Calyculin A maintained D1/2 levels throughout prolonged ischemia. SB203580 decreased alpha BC phosphorylation. Neither PD98059 nor IPC altered alpha BC phosphorylation during prolonged ischemia. It is concluded that alpha BC phosphorylation during ischemia is regulated by p38 MAPK but not by ERK 1/2. The inability to detect a correlation between IPC protection and either alpha BC translocation or phosphorylation suggests that the proteins in the highly phosphorylated isoform bands of alpha BC quantitated in this study are not protective end effectors of classical IPC. PMID- 10860772 TI - 6-Hydroxydopamine-induced developmental cardiac alterations in morphology, calmodulin content, and K(2+)-mediated [Ca(2+)](i)Transient of chicken cardiomyocytes. AB - Calcium and the calcium-calmodulin-mediated processes have been implicated in cardiac hypertrophy. The purpose of this study was to investigate whether there is a potential role played by the calcium-calmodulin processes in cardiac morphogenesis and malformations, especially in the development of cardiac hypertrophy. Recently, the authors reported that 6-hydroxydopamine, an adrenergic neurotoxin can produce malformations in various organs including ventricular septal lesions and cardiac hypertrophy in the developing chicken embryo. Morphological studies revealed areas of coagulative necrosis, with broken nuclear membranes, swollen mitochondria and dilations of the ventricles, as well as thickening of ventricular walls reminiscent of cardiac hypertrophy. The observation that 6-hydroxydopamine treatment on day 3 of incubation produced a dose-dependent increase in both heart and brain calmodulin levels on day 11 of incubation and an increase in the sensitivity to external potassium induction of intracellular free calcium transient in incubation day 14 chicken cardiomyocytes in culture, leading to an increase in intracellular free calcium is reported here. However, sodium/potassium adenosinetriphosphatase activity showed no significant change on days 12 and 16 of incubation. The effect appears to be relatively specific since 5-hydroxydopamine, a chemical isomer of 6 hydroxydopamine, failed to produce a similar sensitivity change of potassium induced intracellular calcium transient. PMID- 10860773 TI - The novel glycolipid RC-552 attenuates myocardial stunning and reduces infarct size in dogs. AB - The novel glycolipid RC-552 shares common structural features with the natural products lipid A and the previously described cardioprotectant monophosphoryl lipid A. RC-552 administered to dogs as a bolus intravenous dose (35-70 microg/kg) either 24 h or 10 min prior to 60 min of regional myocardial ischemia and 3 h of reperfusion significantly (P<0.05 v control) reduced infarct size (IS) as assessed by triphenyltetrazolium staining from 27.0+/-2.3% of the area-at-risk (AAR) to 13.3+/-2.2% and 15.0+/-3.0%, respectively. Administration of the non specific inducible nitric oxide synthase (iNOS) inhibitor aminoguanidine (30 mg/kg, subcutaneously) 1 h prior to ischemia blocked the ability of RC-552 (35 microg/kg, 24 h pretreatment) to reduce infarct size. Intravenous pretreatment with RC-552 (35 microg/kg) either 24 h or 10 min prior to five 5 min repetitive cycles of ischemia and reperfusion significantly improved regional myocardial segment shortening (percentage of control) at all time points during 2 h of reperfusion in dogs. These effects of RC-552 in either cardiac injury model occurred independent of differences in AAR, transmural blood flow during ischemia or hemodynamics throughout the experiment. In contrast with monophosphoryl lipid A (MLA), which has also been reported to be cardioprotective at similar doses in dogs, RC-552 was approximately 100 times less prone to cause fever in the USP rabbit pyrogen test. Likewise, RC-552 did not induce secretion of the proinflammatory cytokines TNF, IL-6 or IL-8 from THP-1 cells or alter the expression of adhesion molecules on human neutrophils at concentrations up to 10 microg/ml. MLA was active in these systems at concentrations in the range 0.1-1.0 microg/ml. In conclusion, RC-552 reduces myocardial infarct size and stunning in dogs in the absence of residual immunomodulatory activity. PMID- 10860774 TI - IL-1 beta increases abundance and activity of the negative transcriptional regulator yin yang-1 (YY1) in neonatal rat cardiac myocytes. AB - Current research from both clinical and basic science perspectives indicates that cytokines play an important role in the genesis of cardiovascular pathology. Specifically, levels of cytokines such as interleukin-1 (IL-1), tumor necrosis factor- alpha (TNF- alpha), and interleukin-6 (IL-6) have been found to be elevated in both acute myocardial injury as well as situations of chronic dysfunction. Further, therapies directed primarily at interfering with cytokine action have suggested that such an immunomodulatory approach may be beneficial in some of these circumstances of myocardial injury. We recently reported that IL-1 beta induces a hypertrophic state in cultured neonatal rat cardiac myocytes that differs from other well described hypertrophic phenotypes in terms of myocardial gene expression (such as skeletal alpha -actin, sACT), an effect that appeared to co-localize with that of the negative regulator yin yang-1 (YY1).(1)In the present study, we further localize the area in the sACT promoter responsible for the IL-1 effect. These investigations indicate that sequences in and around the third upstream serum response element (SRE3) bind YY1 and are required for IL-1 beta mediated repression. This element is also capable of transferring both IL-1 beta and YY1-mediated transcriptional repression to a heterologous promoter. In support of an IL-1 beta induced post-translational modification of YY1 that results in an increase in DNA-binding activity,(32)P-labeling experiments reveal an increase in phosphorylated YY1 in IL-1 beta treated cells and phosphatase treated myocyte nuclear proteins lose their ability to bind to the YY1 site. In summary, these results provide evidence that sequences within the SRE3 of the skeletal actin promoter represent an IL-1 beta response element and suggest that IL-1 beta activates the negative transcription factor YY1 by both transcriptional and post-transcriptional mechanisms. PMID- 10860775 TI - Functional evidence for a cyclic-AMP related mechanism of action of the beta(2) adrenoceptor in human ventricular myocytes. AB - The human ventricle contains both beta(1)- and beta(1)-adrenoceptors (AR) and both have been shown to be present on a single myocyte. In animal ventricular myocardium there is evidence that beta(1)ARs increase cardiac contraction by non cAMP-dependent mechanisms. We have used the anti-adrenergic effects of carbachol and the cAMP antagonist Rp -cAMPS to investigate the functional contribution of cAMP to beta(2)AR responses in human ventricular myocytes isolated from cardiac biopsies or explants. Concentration-response curves to isoproterenol (Iso) were constructed in the absence and presence of a beta(1)AR antagonist, CGP 207 12A (300 nmol/l) to determine the contribution of the beta(2)AR to contraction. The cells were rechallenged with sub-maximal dose of Iso under beta(2)AR-specific conditions and Rp -cAMPS (100-200 micromol/l) or carbachol (1-3 microm/l) added. Rp -cAMPS significantly decreased contraction amplitude (% shortening; Iso 7.1+/ 0.7, Iso+Rp -cAMPS 3.5+/-0.5, n=7, P<0.001) though not completely to the baseline (2.2+/-0.6, n=7). Rechallenge with Iso alone reversed the effects of Rp -cAMPS, and subsequent addition of the beta(1)AR antagonist ICI 118,551 reduced the response to baseline (1.6+/-0.3, n=4) confirming beta(2)AR involvement. Similarly, carbachol decreased Iso-stimulated contraction from 7.5+/-1.2% to 3.2+/-0.9% (P<0.05, n=4), but not completely to basal levels (1.6+/-0.3%). These results provide functional evidence for a predominantly cAMP-mediated mechanism of contractile stimulation by beta(1)ARs in human ventricular myocardium, although a small contribution from a non-cAMP dependent pathway may occur. PMID- 10860776 TI - Voltage-gated K(+)Channel, Kv4.2, localizes predominantly to the transverse-axial tubular system of the rat myocyte. AB - Kv4.2 subunit, a member of K(+)channel gene family, is considered to play a major role in the formation of depolarization-activated transient outward K(+)current channels in the mammalian heart. We investigated the subcellular localization of Kv4.2 subunit in the rat heart by immunofluorescence and immunoelectron microscopy. In atrial cells, Kv4.2 immunofluorescent staining was intensely observed in the peripheral sarcolemma and the intercalated disks, but seldom found in transverse tubules, which are rare or absent in atrial cells. In ventricular cells, the labeling of Kv4.2 immunofluorescent staining was found throughout the entire cell membrane, and the staining was stronger in the transverse-axial tubular system than in the peripheral sarcolemma. Correlative immunoconfocal and immunoelectron microscopy using FluoroNanogold confirmed that Kv4.2 distributed in the transverse-axial tubular system including the longitudinally oriented axial tubules. Immunogold electron microscopy of ultrathin cryosections revealed that Kv4.2 was distributed on the plasma membranes of the T-tubules. The extensive distribution of Kv4.2 on the entire cell membrane of myocytes would provide rat myocardial cells with a large capability for the transport of K(+)ions through the channels in the repolarization phase. PMID- 10860777 TI - Aging reduces the cardioprotective effect of ischemic preconditioning in the rat heart. AB - Multiple brief periods of ischemia in the mammalian heart elicits protection against morphologic and functional damage caused by longer-duration ischemia. Preconditioning-induced protection against post-ischemic contractile dysfunction has been reported to be depressed with aging of the adult heart. This study was undertaken to determine whether aging of the adult myocardium reduces the preconditioning-induced attenuation of necrosis observed with ischemia. Isolated, perfused hearts obtained from Fischer 344 rats of either 3 (young) or 22 (aged) months of age were paced and instrumented for determination of developed left ventricular pressure. Necrosis was determined with triphenyltetrazolium. In the absence of preconditioning, young and aged adult hearts made globally ischemic for 45 min developed necrosis involving 53+/-6% and 49+/-6% of the myocardium, respectively. Contractile function (+dP/dt(max)) at 90 min of reperfusion was depressed by 80% in young and 52% in aged hearts, compared to values obtained prior to preconditioning. Preconditioning with two 5 min ischemia/5 min reperfusion cycles significantly reduced necrosis development and enhanced reperfusion contractile function in young hearts. However, in aged adult hearts, the preconditioning did not significantly reduce the development of necrosis or enhance reperfusion contractile function. These data suggest that aging reduces the effectiveness of preconditioning in providing cardioprotection against ischemic-induced myocardial necrosis. PMID- 10860778 TI - Response to letter to the editor by eisner et al PMID- 10860779 TI - The Rotational Spectrum of OCCCS Revisited. AB - The rotational spectrum of the linear chain molecule tricarbon oxide sulfide, OCCCS, was recorded at room temperature between 118 and 179 GHz using a new commercial millimeter-wave spectrometer. The observed transitions were analyzed together with hitherto unassigned transitions from measurements between 78 and 118 GHz. About 1900 rotational transitions of OCCCS in the vibrational ground state and in 13 excited bending states could be assigned, the latter including the eighth, ninth, and tenth excited state of the low-lying bending mode nu(7) as well as the second excited state of the bending mode nu(6). Besides the determination of effective constants for these newly identified vibrational states, our data also served to considerably improve the effective constants for some of the bending states already characterized in previous publications. Copyright 2000 Academic Press. PMID- 10860780 TI - The ins and outs of a polyglutamine neurodegenerative disease: spinocerebellar ataxia type 1 (SCA1). AB - Polyglutamine neurodegenerative disorders are characterized by the expansion of a glutamine tract within the mutant disease-causing protein. Expression of the mutant protein induces a progressive loss of neuronal function and the subsequent neurodegeneration of a set of neurons characteristic to each disease. Spinocerebellar ataxia type 1 (SCA1) is one polyglutamine disease where various experimental model systems, in particular transgenic mice, have been utilized to dissect the molecular and cellular events important for disease. This review summarizes these findings and places them in a context of potential future research directions. PMID- 10860781 TI - Binding partners of Alzheimer's disease proteins: are they physiologically relevant? AB - Protein-protein interactions are a molecular basis for the structural and functional organization within cells. They are mediated by a growing number of protein modules that bind peptide targets. Alterations in binding affinities can have serious consequences for some essential cellular processes. The three proteins identified to have mutations in their corresponding genes leading to presenile Alzheimer dementia (AD)-the amyloid precursor protein (APP) and presenilin 1 and 2-all interact with other proteins. The nature and function of these interacting proteins may contribute to elucidating the proper physiological functions of the AD proteins. APP-interacting proteins are pointing toward a function of APP in cell adhesion and neurite outgrowth and signaling. Proteins interacting with the presenilins however are more diverse in nature linking presenilin function to regulation in different signaling pathways including Wnt and Notch but also in apoptosis and Ca(2+) homeostasis. Further research however is still needed to delineate the exact functional relevance of these interactions with respect to the physiological functions of the AD proteins in particular and the contribution of these proteins to AD pathogenesis in general. PMID- 10860782 TI - Acetylcholinesterase-positive fiber deafferentation and cell shrinkage in the septohippocampal pathway of aged amyloid precursor protein london mutant transgenic mice. AB - Several lines of evidence implicate a cholinergic deficit in Alzheimer's disease (AD). Transgenic mice that overexpress clinical mutants of the human amyloid precursor protein (APP) have been generated that recapitulate many aspects of AD. We now analyzed the cholinergic system in aged APP/London transgenic mice. The major finding was the reorganization of acetylcholinesterase-positive fibers within the hippocampus and the reduced size of cholinergic cells in the medial septum. The reduction of acetylcholinesterase-positive fibers in the subiculum together with increased fiber density in the CA1 and in the dentate gyrus suggests a synaptic sprouting compensatory mechanism within the hippocampus. In the cortex, amyloid plaques were associated with intense acetylcholinesterase activity and surrounded by dystrophic acetylcholinesterase-positive fibers. Nevertheless, the overall pattern of cholinergic innervation was unchanged. These results demonstrate that overexpression of APP/London caused, besides amyloid plaques in aged mouse brain, also cholinergic deafferentation and cholinergic cell shrinkage. PMID- 10860783 TI - Neuronal death in newborn striatum after hypoxia-ischemia is necrosis and evolves with oxidative stress. AB - The mechanisms for neurodegeneration after hypoxia-ischemia (HI) in newborns are not understood. We tested the hypothesis that striatal neuron death is necrosis and evolves with oxidative stress and selective organelle damage. Piglets ( approximately 1 week old) were used in a model of hypoxia-asphyxia and survived for 3, 6, 12, or 24 h. Neuronal death was progressive over 3-24 h recovery, with approximately 80% of putaminal neurons dead at 24 h. Striatal DNA was digested randomly at 6-12 h. Ultrastructurally, dying neurons were necrotic. Damage to the Golgi apparatus and rough endoplasmic reticulum occurred at 3-12 h, while most mitochondria appeared intact until 12 h. Mitochondria showed early suppression of activity, then a transient burst of activity at 6 h, followed by mitochondrial failure (determined by cytochrome c oxidase assay). Cytochrome c was depleted at 6 h after HI and thereafter. Damage to lysosomes occurred within 3-6 h. By 3 h recovery, glutathione levels were reduced, and peroxynitrite-mediated oxidative damage to membrane proteins, determined by immunoblots for nitrotyrosine, occurred at 3-12 h. The Golgi apparatus and cytoskeleton were early targets for extensive tyrosine nitration. Striatal neurons also sustained hydroxyl radical damage to DNA and RNA within 6 h after HI. We conclude that early glutathione depletion and oxidative stress between 3 and 6 h reperfusion promote damage to membrane and cytoskeletal proteins, DNA and RNA, as well as damage to most organelles, thereby causing neuronal necrosis in the striatum of newborns after HI. PMID- 10860784 TI - Accumulation of insoluble alpha-synuclein in dementia with Lewy bodies. AB - The alpha-synuclein (alpha SN) protein is thought to play a central role in the pathogenesis of neurodegenerative diseases where it aggregates to form intracellular inclusions. We have used Western blotting to examine the expression levels and solubility of alpha SN in brain homogenates from dementia with Lewy bodies (DLB), Parkinson's disease (PD), Alzheimer's disease (AD), and normal controls using samples from the parahippocampus/transentorhinal cortex. Compared to controls, DLB brains accumulate significantly greater amounts of sodium dodecyl sulfate (SDS)-soluble and SDS-insoluble alpha SN but levels of TBS soluble alpha SN did not change. Levels of synaptophysin, a marker of synaptic integrity, were significantly lower in DLB cases than in normal aged controls regardless of whether concurrent changes of AD were present. This limbic synaptic dysfunction may contribute to cognitive impairment in DLB. Whether aggregated alpha SN is a cause or effect of the disease process in DLB and PD remains to be determined, but the presence of aggregated alpha SN is consistent with a pathogenesis similar to that associated with aggregates of Abeta amyloid in AD. PMID- 10860785 TI - Increased beta-carboline 9N-methyltransferase activity in the frontal cortex in Parkinson's disease. AB - Enzymatic beta-carboline N-methyltransferase activities generate N-methylated beta-carbolinium cations that are analogs of the parkinsonian-producing neurotoxin MPP+. We measured beta-carboline-2N-methyltransferase and beta carboline-9N-methyltransferase activities in the supernatant and particulate fractions from postmortem human brains. These N-methyltransferase activities were assessed in the substantia nigra, putamen, and frontal cortex from control and Parkinson's disease cases. No significant differences were measured in any brain region in particulate and supernatant fraction beta-carboline 2N methyltransferase activity or particulate fraction beta-carboline 9N methyltransferase activity. Likewise, supernatant fraction beta-carboline 9N methyltransferase activity was similar in the putamen and substantia nigra from Parkinson's disease and control cases. Unexpectedly, supernatant fraction beta carboline 9N-methyltransferase activity was increased fourfold in Parkinson's disease frontal cortex (P < 0.05), suggesting that beta-carboline N-methylation may play a role in Parkinson's disease. PMID- 10860786 TI - Adenovirus-mediated expression of ciliary neurotrophic factor (CNTF) rescues axotomized rat retinal ganglion cells but does not support axonal regeneration in vivo. AB - Rat optic nerve (ON) transection leads to mainly apoptotic cell death of about 85% of the retinal ganglion cell (RGC) population within 14 days after lesion. In the present study, we tested the effect of adenovirally delivered CNTF (Ad-CNTF) on survival and regeneration of axotomized adult RGCs in vivo. Single intravitreal Ad-CNTF injection led to stable CNTF mRNA and protein expression for at least 18 days and significantly enhanced RGC survival by 155% when compared to control animals 14 days after ON transection. ON stump application of Ad-CNTF also resulted in an increased number of surviving RGCs. Ad-CNTF injection led to better preservation of intraretinal RGC axons but did not support regeneration of axotomized RGCs into a peripheral nerve graft. Thus, adenovirus-mediated neurotrophic factor supply is a suitable approach for reducing axotomy-induced RGC death in vivo and may constitute a relevant strategy for clinical treatment of traumatic brain injury. PMID- 10860787 TI - Regionally specific sensitivity differences in fMRI and PET: where do they come from? AB - In this paper we report three neuroimaging studies of language that investigate potential sources of inconsistency in measured hemodynamic responses: (1) between sessions for fMRI, including differences in hormonal status, (2) between sessions for PET, and (3) between scanning modalities (PET and fMRI). Differences in evoked responses between sessions of the same modality were small. In particular we did not find any effect of hormone levels when testing during the first and third weeks of the menstrual cycle (although we cannot exclude the possibility that activation in the temporoparietal regions is sensitive to hormonal status). Comparing the two modalities showed that prefrontal regions were more activated in fMRI than in PET. This may relate to task switching between blocks in fMRI that is not induced by PET paradigms or increased error variance in these regions for PET. In contrast, temporal activations were found in PET more than in fMRI. We attribute the lack of temporal activations, in fMRI, to a combination of factors, including susceptibility artifacts, anticipatory activity during the control condition, discontinuous sampling of peristimulus time, and differences in the source, acquisition, and analysis of the measured signals. It is concluded that although there is sufficient reproducibility of results for these paradigms within each modality, the regionally specific differences in sensitivity found between modalities warrant further investigation. These regionally specific differences are important for a properly qualified interpretation of activation profiles in fMRI. PMID- 10860788 TI - Susceptibility-induced loss of signal: comparing PET and fMRI on a semantic task. AB - Functional magnetic resonance imaging (fMRI) has become a popular tool for investigations into the neural correlates of cognitive activity. One limitation of fMRI, however, is that it has difficulty imaging regions near tissue interfaces due to distortions from macroscopic susceptibility effects which become more severe at higher magnetic field strengths. This difficulty can be particularly problematic for language tasks that engage regions of the temporal lobes near the air-filled sinuses. This paper investigates susceptibility-induced signal loss in the temporal lobes and proposes that by defining a priori regions of interest and using the small-volume statistical correction of K. J. Worsley, S. Marrett, P. Neelin, A. C. Vandal, K. J. Friston, and A. C. Evans (1996, Hum. Brain Mapp. 4: 58-83), activations in these areas can sometimes be detected by increasing the statistical power of the analysis. We conducted two experiments, one with PET and the other with fMRI, using almost identical semantic categorization paradigms and comparable methods of analysis. There were areas of overlap as well as differences between the PET and fMRI results. One anticipated difference was a lack of activation in two regions in the temporal lobe on initial analyses in the fMRI data set. With a specific region of interest, however, activation in one of the regions was detected. These experiments demonstrate three points: first, even for almost identical cognitive tasks such as those in this study, PET and fMRI may not produce identical results; second, differences between the two methods due to macroscopic susceptibility artifacts in fMRI can be overcome with appropriate statistical corrections, but only partially; and third, new data acquisition paradigms are necessary to fully deal with susceptibility-induced signal loss if the sensitivity of the fMRI experiment to temporal lobe activations is to be enhanced. PMID- 10860789 TI - Magnetic resonance microscopy of the C57BL mouse brain. AB - With the rapid progression in gene technologies, transgenic, targeted, and chemically induced mutations in mice are continually created. The major goal of these studies is to understand and characterize the effects of genotype on anatomy, physiology, and behavior and ultimately the role of genotype in development of disease. The demand for imaging techniques with high spatial resolution potential is rising because such imaging tools would expedite anatomical phenotyping in the genetically altered mice. Magnetic resonance microscopy (MRM) is a noninvasive, inherently three-dimensional (3D) imaging technique capable of visualizing several anatomical structures in the small mouse. The 3D nature of MRM also allows for interpretation of complex spatial relationships between substructures, which is important when phenotyping anatomically. The goal of this paper is to systematically describe three major brain regions in the C57BL/6J mouse at microanatomical spatial resolution ranges using in vitro MRM. We explore different MR contrast parameters, voxel sizes, and signal-to-noise ratios to best characterize C57BL/6J mouse brain microstructure by MRM. Further, we compare all MRM images with Nissl-stained brain sections. Major findings were as follows: T2* MR images visualized several gross anatomical regions in the mouse brain but not, for example, subregions within the hippocampus. Diffusion proton stains on the other hand were superior to T2* MR images and delineated many subregions within the hippocampus proper. Finally, contrast enhancement facilitated visualization of hippocampal anatomy on the T2* MR images. The results of this study are part of an ongoing initiative at our Center focused on creating a complete C57BL/6J mouse anatomical 3D image database by MRM. PMID- 10860790 TI - Attention to speed of motion, speed discrimination, and task difficulty: an fMRI study. AB - We studied the functional neuroanatomy of attention to speed of motion using functional magnetic resonance imaging in eight healthy subjects, who performed a speed discrimination (SID) task using a random textured pattern moving at a reference speed of 6 deg/s. During the control condition (DIM), with retinal stimulation identical to that during SID, subjects detected the dimming of the central fixation point. Attention to speed (SID compared to DIM) activated mainly ventral V3 and V4, dorsal V3 and V3A. Compared to a fixation control condition, speed discrimination recruited a large visuomotor network, including hMT/V5+. However, hMT/V5+ was only marginally more active during speed discrimination than during dimming detection. Thus hMT/V5+ is involved in speed discrimination, in line with the speed discrimination impairments following hMT/V5+ lesions, but our results suggest that this activity simply reflects the processing of motion rather than attention to speed. Manipulating the difficulty of the speed discrimination task over a large range of the psychometric curve revealed that increasing difficulty linearly increases activity in right frontal regions, as well as in lateral occipital and dorsal parietal regions. A weak effect of difficulty was also observed in dorsal V3. PMID- 10860791 TI - Hemispheric activation of anterior and inferior prefrontal cortex during verbal encoding and recognition: a PET study of healthy volunteers. AB - Evidence of bilateral prefrontal activation during memory encoding and retrieval has increased attention given to anatomical subdivisions within the prefrontal cortex. The current study examined anterior and inferior aspects of the prefrontal cortex to determine their degree of functional and hemispheric overlap during encoding and recognition. Cerebral blood flow of 25 healthy volunteers was measured using PET (15)O-water methods during four conditions: resting baseline, sequential finger movement, word encoding, and word recognition. Resting and motor images were averaged to provide a single reference that was subtracted from encoding and recognition using statistical parametric mapping (SPM96). Memory conditions were also subtracted from each other to identify differences in regional activity. Subjects performed well (86% correct) and had a slightly conservative response bias. Baseline subtraction from encoding revealed focal activation of left inferior prefrontal cortex (area 45) without significant contralateral activation. Recognition minus baseline subtraction produced a focal right anterior prefrontal activation (areas 9 and 10) that was not present in the left hemisphere. Bilateral effects were seen in area 45 during recognition. Subtraction of memory tasks from each other did not reveal any areas of greater activity during encoding. However, the recognition task produced greater activation in right area 9 extending into the anterior cingulate. Greater activity during recognition was also observed in left insula and bilateral visual integration areas. These results are discussed in relation to the prevailing model of prefrontal hemispheric asymmetry during episodic memory. PMID- 10860792 TI - Brain areas involved in rapid categorization of natural images: an event-related fMRI study. AB - Event-related fMRI was used to investigate brain activation during a visual go/no go categorization task based on colored photographs of natural scenes, similar to a previous ERP study by Thorpe et al. (1996, Nature 381: 520-522). Subjects had to press a key when an animal was present in the display. Stimuli were flashed for 33 ms using an intertrial interval of 5 s and a design that carefully balanced targets and distractors in a pseudo-random sequence. Activation produced by targets and distractors was compared with two different techniques, one based on correlations with the stimulation pattern, the other using simple t score statistics to compare selected scans. The contralateral primary motor cortex and the ipsilateral cerebellum were both more active following target trials than following distractors, thus confirming the sensitivity of the method. Differential activity was also seen in the posterior cingulate cortex, the fusiform, and the parahippocampic gyri. Activity in such structures could underlie the differential evoked-potentials reported previously in the same task. Surprisingly, in these visual structures, the signal was stronger following distractor trials than target ones. This result could be due to more prolonged processing on distractor trials. Alternatively, it could be that target detection induces strong activation of a small proportion of neurons, which, because of competitive inhibitory mechanisms, could result in a decrease in activity for the population as a whole. We suggest that this kind of mechanism could also account for the decreases in signal observed in perceptual priming experiments. PMID- 10860793 TI - The role of lateral occipitotemporal junction and area MT/V5 in the visual analysis of upper-limb postures. AB - Humans, like numerous other species, strongly rely on the observation of gestures of other individuals in their everyday life. It is hypothesized that the visual processing of human gestures is sustained by a specific functional architecture, even at an early prelexical cognitive stage, different from that required for the processing of other visual entities. In the present PET study, the neural basis of visual gesture analysis was investigated with functional neuroimaging of brain activity during naming and orientation tasks performed on pictures of either static gestures (upper-limb postures) or tridimensional objects. To prevent automatic object-related cerebral activation during the visual processing of postures, only intransitive postures were selected, i. e., symbolic or meaningless postures which do not imply the handling of objects. Conversely, only intransitive objects which cannot be handled were selected to prevent gesture related activation during their visual processing. Results clearly demonstrate a significant functional segregation between the processing of static intransitive postures and the processing of intransitive tridimensional objects. Visual processing of objects elicited mainly occipital and fusiform gyrus activity, while visual processing of postures strongly activated the lateral occipitotemporal junction, encroaching upon area MT/V5, involved in motion analysis. These findings suggest that the lateral occipitotemporal junction, working in association with area MT/V5, plays a prominent role in the high-level perceptual analysis of gesture, namely the construction of its visual representation, available for subsequent recognition or imitation. PMID- 10860794 TI - Anatomically informed basis functions. AB - This paper introduces the general framework, concepts, and procedures of anatomically informed basis functions (AIBF), a new method for the analysis of functional magnetic resonance imaging (fMRI) data. In contradistinction to existing voxel-based univariate or multivariate methods the approach described here can incorporate various forms of prior anatomical knowledge to specify sophisticated spatiotemporal models for fMRI time-series. In particular, we focus on anatomical prior knowledge, based on reconstructed gray matter surfaces and assumptions about the location and spatial smoothness of the blood oxygenation level dependent (BOLD) effect. After reconstruction of the grey matter surface from an individual's high-resolution T1-weighted MRI, we specify a set of anatomically informed basis functions, fit the model parameters for a single time point, using a regularized solution, and finally make inferences about the estimated parameters over time. Significant effects, induced by the experimental paradigm, can then be visualized in the native voxel-space or on the reconstructed folded, inflated, or flattened cortical surface. As an example, we apply the approach to a fMRI study (finger opposition task) and compare the results to those of a voxel-based analysis as implemented in the Statistical Parametric Mapping package (SPM99). Additionally, we show, using simulated data, that the approach offers several desirable features particularly in terms of superresolution and localization. PMID- 10860795 TI - Application of cortical unfolding techniques to functional MRI of the human hippocampal region. AB - We describe a new application of cortical unfolding to high-resolution functional magnetic resonance imaging (fMRI) of the human hippocampal region. This procedure includes techniques to segment and unfold the hippocampus, allowing the fusiform, parahippocampal, perirhinal, entorhinal, subicular, and CA fields to be viewed and compared across subjects. Transformation parameters derived from unfolding high-resolution structural images are applied to coplanar, functional images, yielding two-dimensional "unfolded" activation maps of hippocampi. The application of these unfolding techniques greatly enhances the ability of fMRI to localize and characterize signal changes within the medial temporal lobe. Use of this method on a novelty-encoding paradigm reveals a temporal dissociation between activation along the collateral sulcus and activation in the hippocampus proper. PMID- 10860796 TI - Areas 3a, 3b, and 1 of human primary somatosensory cortex. Part 2. Spatial normalization to standard anatomical space. AB - Interindividual topographical variability of cytoarchitectonically defined somatosensory areas 3a, 3b, and 1 was analyzed in the standard anatomical format of a computerized brain atlas. T1-weighted magnetic resonance images were obtained from 10 postmortem brains. The brains were serially sectioned at 20 mcm, sections were stained for cell bodies, and areas 3a, 3b, and 1 were defined with an observer-independent cytoarchitectonic technique. After correction of the sections for deformations due to histological processing, the 3-D reconstructed histological volumes of the individual brains and the volume representations of the cytoarchitectonic areas were adapted to the reference brain of a computerized atlas. Corresponding areas were superimposed in the 3-D space of the reference brain. These population maps describe, for each voxel, how many brains have a representation of one particular cytoarchitectonic area. Each area's extent is very variable across different brains, but representations of areas 3a, 3b, and 1 in >/=50% of the brains were found in the fundus of the central sulcus, its caudal bank, and on the crown of the postcentral gyrus, respectively. Volumes of interest (VOIs) were defined for each area in which >/=50% of the brains have a representation of that area. Despite close spatial relationship of areas 3a, 3b, and 1 in the postcentral gyrus, the three VOIs overlap by <1% of their volumes. Functional imaging data can now be brought into the same standard anatomical format, and changes in regional cerebral blood flow can be calculated in VOIs of areas 3a, 3b, and 1, which are derived from genuine cytoarchitectonic data. PMID- 10860797 TI - Supplementary motor area activation preceding voluntary movement is detectable with a whole-scalp magnetoencephalography system. AB - Despite the fact that the knowledge about the structure and the function of the supplementary motor area (SMA) is steadily increasing, the role of the SMA in the human brain, e.g., the contribution of the SMA to the Bereitschaftspotential, still remains unclear and controversial. The goal of this study was to contribute further to this discussion by taking advantage of the increased spatial information of a whole-scalp magnetoencephalography (MEG) system enabling us to record the magnetic equivalent of the Bereitschaftspotential 1, the Bereitschaftsfeld 1 (BF 1) or readiness field 1. Five subjects performed a complex, and one subject a simple, finger-tapping task. It was possible to record the BF 1 for all subjects. The first appearance of the BF 1 was in the range of 1.9 to -1.7 s prior to movement onset, except for the subject performing the simple task (-1 s). Analysis of the development of the magnetic field distribution and the channel waveforms showed the beginning of the Bereitschaftsfeld 2 (BF 2) or readiness field 2 at about -0.5 s prior to movement onset. In the time range of BF 1, dipole source analysis localized the source in the SMA only, whereas dipole source analysis containing also the time range of BF 2 resulted in dipole models, including dipoles in the primary motor area. In summary, with a whole-head MEG system, it was possible for the first time to detect SMA activity in healthy subjects with MEG. PMID- 10860798 TI - Variability in fMRI: an examination of intersession differences. AB - The results from a single functional magnetic resonance imaging session are typically reported as indicative of the subject's functional neuroanatomy. Underlying this interpretation is the implicit assumption that there are no responses specific to that particular session, i.e., that the potential variability of response between sessions is negligible. The present study sought to examine this assumption empirically. A total of 99 sessions, comprising 33 repeats of simple motor, visual, and cognitive paradigms, were collected over a period of 2 months on a single male subject. For each paradigm, the inclusion of session-by-condition interactions explained a significant amount of error variance (P < 0.05 corrected for multiple comparisons) over a model assuming a common activation magnitude across all sessions. However, many of those voxels displaying significant session-by-condition interactions were not seen in a multisession fixed-effects analysis of the same data set; i.e., they were not activated on average across all sessions. Most voxels that were both significantly variable and activated on average across all sessions did not survive a random-effects analysis (modeling between-session variance). We interpret our results as demonstrating that correct inference about subject responses to activation tasks can be derived through the use of a statistical model which accounts for both within- and between-session variance, combined with an appropriately large session sample size. If researchers have access to only a single session from a single subject, erroneous conclusions are a possibility, in that responses specific to this single session may be claimed to be typical responses for this subject. PMID- 10860799 TI - Characterizing the hemodynamic response: effects of presentation rate, sampling procedure, and the possibility of ordering brain activity based on relative timing. AB - Rapid-presentation event-related functional MRI (ER-fMRI) allows neuroimaging methods based on hemodynamics to employ behavioral task paradigms typical of cognitive settings. However, the sluggishness of the hemodynamic response and its variance provide constraints on how ER-fMRI can be applied. In a series of two studies, estimates of the hemodynamic response in or near the primary visual and motor cortices were compared across various paradigms and sampling procedures to determine the limits of ER-fMRI procedures and, more generally, to describe the behavior of the hemodynamic response. The temporal profile of the hemodynamic response was estimated across overlapping events by solving a set of linear equations within the general linear model. No assumptions about the shape were made in solving the equations. Following estimation of the temporal profile, the amplitude and timing were modeled using a gamma function. Results indicated that (1) within a region, for a given subject, estimation of the hemodynamic response is extremely stable for both amplitude (r(2) = 0.98) and time to peak (r(2) = 0.95), from one series of measurements to the next, and slightly less stable for estimation of time to onset (r(2) = 0.60). (2) As the trial presentation rate changed (from those spaced 20 s apart to temporally overlapping trials), the hemodynamic response amplitude showed a small, but significant, decrease. Trial onsets spaced (on average) 5 s apart showed a 17-25% reduction in amplitude compared to those spaced 20 s apart. Power analysis indicated that the increased number of trials at fast rates outweighs this decrease in amplitude if statistically reliable response detection is the goal. (3) Knowledge of the amplitude and timing of the hemodynamic response in one region failed to predict those properties in another region, even for within-subject comparisons. (4) Across subjects, the amplitude of the response showed no significant correlation with timing of the response, for either time-to-onset or time-to-peak estimates. (5) The within-region stability of the response was sufficient to allow offsets in the timing of the response to be detected that were under a second, placing event-related fMRI methods in a position to answer questions about the change in relative timing between regions. PMID- 10860800 TI - Extrastriatal mean regional uptake of fluorine-18-FDOPA in the normal aged brain- an approach using MRI-aided spatial normalization. AB - The aim of this study was to define the mean regional 6-[(18)F]fluoro-l-dopa (FDOPA) uptake rate constant (K(i)) values in the striatal and extrastriatal regions of the brain of normal subjects using magnetic resonance imaging (MRI) aided spatial normalization of the FDOPA K(i) image and using automatic region of interest (ROI) analysis. Dynamic three-dimensional FDOPA positron emission tomography (PET) and three-dimensional magnetic resonance (MR) images were acquired in 13 aged normal subjects. The FDOPA add image and the K(i) image of each subject were transformed into standard stereotactic space with the aid of individual coregistered MR image. The mean regional K(i) values of the striatal and extrastriatal regions before normalization were compared with the respective values after normalization. Then automatic ROI analysis was performed on the MRI aided spatially normalized K(i) images of the 13 normal subjects. The K(i) values on original images and those on spatially normalized images were in good agreement, indicating that the spatial normalization technique did not change the regional K(i) values appreciably. Automatic ROI analysis of the spatially normalized FDOPA K(i) images of the normal subjects, showed high K(i) values in ventral and dorsal regions of the midbrain, amygdala, hippocampus, and medial prefrontal cortex, in addition to caudate nucleus and putamen, which correspond to the dopaminergic projections in the brain. Spatial normalization technique helped to establish a database of FDOPA K(i) images of normal subjects and high K(i) values were observed widely besides striatal regions corresponding to the dopaminergic projections in the brain. PMID- 10860801 TI - Structural group analysis of functional activation maps. AB - We present here a new method for cerebral activation detection over a group of subjects. This method is performed using individual activation maps of any sort. It aims at processing a group analysis while preserving individual information and at overcoming as far as possible limitations of the spatial normalization used to compare different subjects. We designed it such that it provides the individual occurrence of the activations detected at a group level. The localization can then be performed on the individual anatomy of each subject. The analysis starts with a hierarchical multiscale object-based description of each individual map. These descriptions are then compared, rather than comparing the images directly. The analysis is thus performed at an object level instead of voxel by voxel. It is made using a comparison graph, on which a labeling process is performed. The label field on the graph is modeled by a Markov random field, which allows us to introduce high-level rules of interrogation of the data. The process has been evaluated on simulated data and real data from a PET protocol. PMID- 10860802 TI - Testing for neural responses during temporal components of trials with BOLD fMRI. AB - Some cognitive neuroscientific hypotheses might concern neural responses occurring during particular periods of time in a behavioral trial. Here, these particular periods of time are referred to as temporal components of the trial. A difficulty in using BOLD fMRI to test hypotheses about neural responses during temporal components is that some information is irretrievably lost when neural responses are hemodynamically transformed. As a result, one cannot in general use the fMRI signal to unambiguously specify if there was a neural response during a given temporal component. However, adoption of a linear-time invariant model for the transform from neural signal to fMRI signal and constraint of the space of underlying neural waveforms might allow one to ask such questions. Here, the basic theory relevant to this issue and a corresponding method are discussed. The application of this method to fMRI time series data collected during the performance of a delayed-response trial is provided as an illustrative example. PMID- 10860803 TI - Temporal filtering of event-related fMRI data using cross-validation. AB - To circumvent the problem of low signal-to-noise ratio (SNR) in event-related fMRI data, the fMRI experiment is typically designed to consist of repeated presentations of the stimulus and measurements of the response, allowing for subsequent averaging of the resulting data. Due to factors such as time limitation, subject motion, habituation, and fatigue, practical constraints on the number of repetitions exist. Thus, filtering is commonly applied to further improve the SNR of the averaged data. Here, a time-varying filter based on theoretical work by Nowak is employed. This filter operates under the stationary wavelet transform framework and is demonstrated to lead to good estimates of the true signals in simulated data. The utility of the filter is also shown using experimental data obtained with a visual-motor paradigm. PMID- 10860804 TI - Voxel-based morphometry--the methods. AB - At its simplest, voxel-based morphometry (VBM) involves a voxel-wise comparison of the local concentration of gray matter between two groups of subjects. The procedure is relatively straightforward and involves spatially normalizing high resolution images from all the subjects in the study into the same stereotactic space. This is followed by segmenting the gray matter from the spatially normalized images and smoothing the gray-matter segments. Voxel-wise parametric statistical tests which compare the smoothed gray-matter images from the two groups are performed. Corrections for multiple comparisons are made using the theory of Gaussian random fields. This paper describes the steps involved in VBM, with particular emphasis on segmenting gray matter from MR images with nonuniformity artifact. We provide evaluations of the assumptions that underpin the method, including the accuracy of the segmentation and the assumptions made about the statistical distribution of the data. PMID- 10860805 TI - Kinetic modeling of ATP synthesis by ATP synthase and its mechanistic implications. AB - Based on the torsional mechanism of ATP synthesis by ATP synthase, a kinetic scheme has been developed in this work. The scheme considers adenine nucleotide transport, binding of substrates ADP and P(i), unbinding of product ATP, and ATP synthesis. This kinetic scheme has been analyzed mathematically, and a kinetic model has been obtained to explain the experimentally observed hyperbolic Michaellian dependence of the rate of ATP synthesis on the ADP concentration by ATP synthase under physiological steady-state operating conditions. The principal results of the kinetic model have been compared with the experimental data and an estimate of the enzymological kinetic parameters V(max), K(M), and K(I) has been determined. Mechanistic implications arising from further analysis of the kinetic model have been discussed. These biological implications provide deep insight into the sequence of events leading to ATP synthesis. PMID- 10860806 TI - A mechanism for action of oscillating electric fields on cells. AB - The biological effects of electromagnetic fields have seriously concerned the scientific community and the public as well in the past decades as more and more evidence has accumulated about the hazardous consequences of so-called "electromagnetic pollution." This theoretical model is based on the simple hypothesis that an oscillating external electric field will exert an oscillating force to each of the free ions that exist on both sides of all plasma membranes and that can move across the membranes through transmembrane proteins. This external oscillating force will cause a forced vibration of each free ion. When the amplitude of the ions' forced vibration transcends some critical value, the oscillating ions can give a false signal for opening or closing channels that are voltage gated (or even mechanically gated), in this way disordering the electrochemical balance of the plasma membrane and consequently the whole cell function. PMID- 10860807 TI - Phosphorylation of RTP, an ER stress-responsive cytoplasmic protein. AB - RTP, also called Drg1/Cap43/rit42/TDD5/Ndr1, was originally identified as a homocysteine-responsive gene product, and is now considered to be involved in stress responses, atherosclerosis, carcinogenesis, differentiation, androgen responses, hypoxia, and N-myc pathways. We raised an antiserum against a recombinant human RTP. Western blot analysis showed that RTP expression was induced in human umbilical vein endothelial cells under conditions causing endoplasmic reticulum stress. RTP was partially phosphorylated at seven or more sites. The phosphorylation was reversible, and was enhanced by an increased level of intracellular cAMP and inhibited by both a protein kinase A inhibitor and a calmodulin kinase inhibitor. Protein kinase A directly phosphorylated recombinant RTP in vitro. The phosphorylated forms were abundant in cells at the early log phase, and then decreased with increasing cell density. These data demonstrated that RTP is a phosphorylated stress-responsive protein, and its phosphorylation may be related to cell growth. PMID- 10860808 TI - Human fetal skin fibroblast migration stimulated by the autocrine growth factor bFGF is mediated by phospholipase A(2) via arachidonic acid without the involvement of pertussis toxin-sensitive G-protein. AB - We reported previously that human fetal skin fibroblast migration into a denuded area was stimulated by an autocrine factor, basic fibroblast growth factor (bFGF). Since the signal transduction pathway of this migration is unknown, we attempted to clarify it by comparing this fibroblast migration with a previously reported bovine endothelial cell migration into a wounded area stimulated by an addition of bFGF, in which the bFGF signal was mediated by phospholipase A(2) coupled G-protein and phospholipase A(2) (PLA(2)) via arachidonic acid. Our study demonstrated that pertussis toxin, a specific inhibitor of PLA(2)-coupled G protein, did not suppress human fetal skin fibroblast migration, but 2-(p amylcinnamyl)amino-4-chlorobensoic acid (ONO-RS-082), a PLA(2) inhibitor, did. Since ONO-RS-082 is a non-specific PLA(2) inhibitor, a cytoplasmic, Ca-dependent PLA(2) (cPLA(2)) inhibitor, AACOCF3, was examined. AACOCF3 suppressed cell migration in certain concentrations. The PLA(2) inhibitor-suppressed cell migration was restored by adding arachidonic acid, and cell migration suppressed by anti-bFGF antibodies was restored by adding arachidonic acid. In addition, pertussis toxin did not suppress arachidonic acid release, which shows an action of PLA(2), but AACOCF3 did. These results indicate that human fetal skin fibroblast migration stimulated by an autocrine factor, bFGF, was mediated by PLA(2) via arachidonic acid without the involvement of PLA(2)-coupled G-protein. PMID- 10860809 TI - Molecular cloning of the gene encoding flavoredoxin, a flavoprotein from Desulfovibrio gigas. AB - Sulfate-reducing bacteria are rich in unique redox proteins and electron carriers that participate in a variety of essential pathways. Several studies have been carried out to characterize these proteins, but the structure and function of many are poorly understood. Many Desulfovibrio species can grow using hydrogen as the sole energy source, indicating that the oxidation of hydrogen with sulfite as the terminal electron acceptor is an energy-conserving mechanism. Flavoredoxin is an FMN-binding protein isolated from the sulfate-reducing bacteria Desulfovibrio gigas that participates in the reduction of bisulfite from hydrogen. Here we report the cloning and sequencing of the flavoredoxin gene. The derived amino acid sequence exhibits similarity to several flavoproteins which are members of a new family of flavin reductases suggested to bind FMN in a novel mode. PMID- 10860810 TI - Identification of age-dependent changes in expression of senescence-accelerated mouse (SAMP8) hippocampal proteins by expression array analysis. AB - Aging is associated with extensive cognitive impairments, although the biochemical and physiological basis of these deficits are unknown. As the hippocampus plays a vital role in cognitive functions, we have selected this tissue to analyze changes in gene expression at two different ages. Array technology is utilized to explore how gene expression in hippocampus is affected by accelerated cognitive impairment in Senescence-Accelerated Mouse (SAM P8) strain. We show that the expression of genes associated with stress response and xenobiotic metabolism are strongly affected at a time when cognitive impairment occurs. Affected genes include those involved both in signaling and chaperone function. The effector and regulator family of chaperones, which play an important role in protein folding, and also the xenobiotic metabolizing enzymes that play crucial role in antioxidant systems, show significant changes in gene expression between 4 and 12 months. PMID- 10860811 TI - Sexually dimorphic expression of two types of DM (Doublesex/Mab-3)-domain genes in a teleost fish, the Tilapia (Oreochromis niloticus). AB - Sex determination consists of somatic and germ-line sex differentiation hierarchies whose interaction is poorly understood. A single gene known to control somatic sex determination, the DM-domain containing (Doublesex/Mab-3 DNA binding motif) gene, is highly conserved across species. Vertebrate DMRT1 (DM related transcription factor 1) expression occurs predominantly in testis. We, however, isolated two distinct DM-domain cDNAs from tilapia testis and ovary, named tDMRT1, and tDMO (DM-domain gene in Ovary), respectively. Despite high homology in the DM-domain, there is little similarity outside the DM-domain. A male specific motif is absent in tDMO indicating a similarity with the female type of doublesex in Drosophila. In contrast to the alternatively spliced male and female types of doublesex, tDMRT1 and tDMO cDNAs are encoded by two different genes. The mutually exclusive nature of tDMRT1 and tDMO expression in the testis or ovary suggests that they both play important roles in gonadal development and/or function. PMID- 10860812 TI - Transduction of Apaf-1 or caspase-9 induces apoptosis in A-172 cells that are resistant to p53-mediated apoptosis. AB - p53 replacement gene therapy has been carried out clinically for cancers with p53 mutations; however, some cancers are resistant to p53 gene therapy. In this study, we transduced A-172 and U251 cells harboring p53 mutations with wild-type p53 using adenovirus vectors to induce wild-type p53 protein at similar expression levels. A-172 cells did not undergo apoptosis after p53 transduction, whereas U251 cells were markedly sensitive to p53-mediated apoptosis. A-172 cells showed higher endogenous expression of Bcl-X(L) than U251, and transduction of Bcl-X(L) repressed p53-mediated apoptosis in U251 cells, suggesting that high endogenous expression of Bcl-X(L) renders A-172 cells, at least in part, resistant to p53-mediated apoptosis. We transduced A-172 cells and U251 cells with the Apaf-1 or caspase-9 genes; both are downstream components of p53 mediated apoptosis. We found that A-172 cells were highly sensitive to Apaf-1- and caspase-9-mediated apoptosis. The results indicate that A-172 cells harboring mutant p53 were not susceptible to p53-mediated apoptosis, possibly due to high endogenous expression of Bcl-X(L). Transduction of Apaf-1 or caspase-9 would override the resistance mechanism of apoptosis in A-172 cells. These findings provide potentially a novel approach in killing cancers that are resistant to p53 replacement gene therapy. PMID- 10860813 TI - Synergistic induction of apoptosis in murine hepatoma Hepa1-6 cells by IFN-gamma and TNF-alpha. AB - In this study, we examined the susceptibility of murine hepatoma Hepa1-6 cells to undergo IFN-gamma- and/or TNF-alpha-induced apoptosis. IFN-gamma or TNF-alpha alone had no demonstrable cytotoxic effects, whereas IFN-gamma and TNF-alpha in combination induced apoptosis drastically in Hepa1-6 cells. During this apoptosis, an increase in caspase-3- and -8-like protease activities and activation of caspase-3, identified by the appearance of its p17 fragment, were observed. Moreover, the cytotoxic induction and caspase-3 activation were effectively inhibited by Z-Asp-CH(2)-DCB (Z-Asp), a caspase inhibitor. Further, an elevation of cytochrome c in the cytosol, in a parallel to activation of caspase-3, was observed in a time-dependent manner. Concurrently, up-regulation of caspase-11 gene expression and processing of procaspase-11 were detected during this apoptosis. These results suggest that the caspase-3 activation, the release of cytochrome c from mitochondria, and increased caspase-11 gene expression involve in synergistic induction of apoptosis in Hepa1-6 cells by IFN gamma and TNF-alpha. PMID- 10860814 TI - Regulation and significance of hepatocyte-derived matrix metalloproteinases in liver remodeling. AB - Regulation in expression and activation of proteinases is one of the most important mechanisms in organ morphogenesis. In this study, we investigated the expression of MMPs in primary hepatocytes and their roles in liver remodeling. A hepatocyte proliferation initiating cytokine, TNFalpha, induced MMP-9 expression in these cells while the expression of MMP-2 did not change by zymography analysis. Interestingly, both the induced MMP-9 expression and hepatocyte proliferation by TNFalpha were synergistically enhanced by HGF in vitro. The increased proliferation was suppressed by MMP inhibitor TIMP-1, suggesting that cytokine-induced MMP regulates proliferation. The increased expression of MMP-9 by the cytokines was inhibited by cytochalasin D or colchicine but not by PI3 kinase inhibitor wortmannin. In addition, co-stimulation by TNFalpha and HGF of spheroidal hepatocytes cultured in 3-dimensional collagen gel drastically induced morphological changes by cell extension and migration in the gel, which was in parallel with the induced expression of MMP-9 and was inhibited by TIMP-1 and -2. The MMP activity was also detected in vivo in the regenerating liver after partial hepatectomy by in situ zymography. These results suggest the roles of MMPs produced by parenchymal cells in liver remodeling. PMID- 10860815 TI - Cell-type specificity of l-leucyl l-leucine methyl ester. AB - l-Leucyl l-leucine methyl ester (LeuLeuOMe) is a lysosomotropic agent which is converted to a membranolytic compound by dipeptidyl peptidase I and kills human leukocytes such as CD8+ T cells and monocytes but not B cells. The reagent has also been used in mice on the assumption that the cell-type specificity to murine leukocytes is the same as that to human leukocytes. During study on the effect of LeuLeuOMe on antigen-driven IL-2 production using murine splenocytes as antigen presenting cells, however, we noticed that murine B cells were sensitive to LeuLeuOMe. We therefore examined the cell-type specificity using murine splenocytes and peritoneal macrophages. Flow cytometric analysis revealed that the most sensitive cells to LeuLeuOMe were CD8+ cells and that CD19+ cells (B cells) were as sensitive as CD3+ cells (T cells). Murine splenic B cells, which were either positively or negatively sorted with a cell sorter, were also sensitive to LeuLeuOMe, whereas human peripheral blood B cells, which were positively sorted, were not. Peritoneal macrophages were the most insensitive to LeuLeuOMe. Thus, this study demonstrated that the cell-type specificity to murine leukocytes is different from that to human leukocytes. PMID- 10860816 TI - False fertilization in sea urchin eggs induced by diabolin, a 120K kelp protein. AB - A 120K lectin-like protein was isolated from the kelp Laminaria diabolica (Oni kombu), with a unique activity to induce false fertilization specifically in the eggs of the sea urchin Hemicentrotus pulcherrimus. The protein designated as "diabolin" rendered the unfertilized egg forms and elevated the fertilization envelope without insemination at 18 nM half-maximally. Those eggs with elevated fertilization envelopes, however, could not enter into normal cleavage or further development, and hence the proliferation of the sea urchin was hindered. Diabolin, thus, by its unique defense mechanism protects the kelp from the predator sea urchin. It was partially sequenced and found to have the highest homology with phytoene dehydrogenase from the plant virus Erwinia uredovora. A question was left to be solved as to how the kelp on the southeast coast of Hokkaido Island could develop the defense mechanism against the sea urchin on Honshu Island separated by Tsugaru Straits. PMID- 10860817 TI - Molecular alignment of proteins in bicellar solutions: quantitative evaluation of effects induced in 2D COSY spectra. AB - Partial molecular alignment leads to an incomplete averaging of anisotropic magnetic interactions such as magnetic dipole interaction or chemical shift anisotropy. In the present contribution we quantitatively describe and evaluate the effects induced by the addition of magnetically oriented lipid bicelles in homonuclear two-dimensional (2D) NMR correlation (COSY) spectra of proteins. It is shown that 2D COSY experiments allow the measurement of H(N)-H(alpha) residual dipole couplings of positive sign which can be used for structure refinement. In contrast to the double- and triple-resonance experiments previously proposed, these measurements can be carried out even on nonisotope-enriched samples. PMID- 10860818 TI - The HIV-1 cell entry inhibitor T-20 potently chemoattracts neutrophils by specifically activating the N-formylpeptide receptor. AB - T-20, a synthetic peptide corresponding to the heptad repeat sequence of HIV-1 gp41, blocks HIV-1 entry by targeting gp41, and is currently in clinical trials as an anti-retroviral agent. We recently reported that in vitro T-20 also functions as a phagocyte chemoattractant and a chemotactic agonist at the phagocyte N-formylpeptide receptor (FPR). Here we show that T-20 is also a potent chemotactic agonist in vitro at a related human phagocyte receptor FPRL1R. To test the relative importance of FPR and FPRL1R in primary cells, we identified the corresponding mouse T-20 receptors, mFPR and FPR2, which are both expressed in neutrophils, and compared T-20 action on neutrophils from wild type and mFPR knockout mice. Surprisingly, although T-20 activates mFPR and FPR2 in transfected cells with equal potency and efficacy in both calcium flux and chemotaxis assays, neutrophils from mFPR knockout mice did not respond to T-20. These results provide genetic evidence that FPR is the major phagocyte T-20 receptor in vivo and point to the potential feasibility of studying T-20 effects on immunity in a mouse model. This may help define the cause of local inflammation after T-20 injection that has recently been reported in Phase I clinical trials. PMID- 10860819 TI - Bone morphogenetic protein-2 blocks MDA MB 231 human breast cancer cell proliferation by inhibiting cyclin-dependent kinase-mediated retinoblastoma protein phosphorylation. AB - Bone morphogenetic protein-2 (BMP-2) has been shown to act as an antiproliferative agent for a number of different cell types. We show that BMP-2 dose-dependently inhibits growth of MDA MB 231 human breast cancer cells. Epidermal growth factor (EGF) stimulates DNA synthesis and entry of these cells into the S-phase. BMP-2 inhibits EGF-induced DNA synthesis by arresting them in G1 phase of the cell cycle. BMP-2 increases the level of cyclin kinase inhibitor p21. Furthermore, we show that exposure of MDA MB 231 cells to BMP-2 stimulates association of p21 with cyclin D1 and with cyclin E resulting in the inhibition of their associated kinase activities. Finally, BMP-2 treatment is found to cause hypophosphorylation of the retinoblastoma protein (pRb), a key regulator of cell cycle progression. Our data provide a mechanism for the antiproliferative effect of BMP-2 in the breast cancer cells. PMID- 10860820 TI - p21-activated kinase PAK phosphorylates desmin at sites different from those for Rho-associated kinase. AB - p21-activated kinase (PAK) and Rho-associated kinase (Rho-kinase) have been shown to induce Ca(2+)-independent contraction of smooth muscle. PAK-induced contraction of Triton-skinned smooth muscle correlates with increased phosphorylation of caldesmon and desmin, although the role of desmin phosphorylation has remained obscure. Here we report that desmin serves as an excellent substrate for PAK in vitro. PAK phosphorylated desmin in a GTP. Cdc42/Rac-dependent manner. Phosphorylation of desmin by PAK dramatically inhibited its filament-forming ability. PAK phosphorylated mainly serine residues of the head domain of desmin, and the major phosphorylation sites differed from those for Rho-kinase. These results suggest that different site-specific phosphorylation of desmin via two divergent protein kinases downstream of Rho family GTPases would seem to increase the regulatory potential for organization of desmin filaments. PMID- 10860821 TI - Tensin can induce JNK and p38 activation. AB - Cells organize diverse types of specialized adhesion sites upon attachment to extracellular matrix (ECM) components. One of the physiological roles of such cell-ECM interactions is to initiate and regulate adhesion-mediated signal transduction responses. The association of cells with fibronectin fibrils has been shown to regulate the JNK and p38 signaling pathways. We tested whether tensin, a cytoskeletal component localized to both focal contacts and fibronectin associated fibrillar adhesions, can induce these signaling pathways. We found that tensin overexpression resulted in activation of both the c-Jun amino terminal kinase (JNK) and p38 pathways. Tensin-mediated JNK activation was independent of the activities of the small GTP binding proteins Rac and Cdc42, but did depend on SEK, a kinase involved in the JNK pathway. We suggest that tensin may directly activate the JNK and p38 pathways, acting downstream or independent of the activities of the small GTP binding proteins Rac and Cdc42. PMID- 10860822 TI - Molecular cloning of neuronally expressed mouse betaPix isoforms. AB - Pix, a Pak-interacting exchange factor, is known to be involved in the regulation of Cdc42/Rac GTPases and Pak kinase activity. In this study, we cloned the cDNAs encoding two betaPix isoforms from mouse brain cDNA library. Both of the cloned genes, designated betaPix-b and betaPix-c (GenBank Accession Nos. AF247654 and AF247655, respectively), have a novel insert region consisting of 59 amino acid residues. In betaPix-c, 75 amino acid residues are deleted in the proline-rich region at the carboxyl-terminus of betaPix. In situ hybridization studies with insert region-specific probe in rat embryo show that insert region-containing isoforms are expressed mainly in the central nervous system. Moreover, temporal expression pattern of isoforms is correlated with the active neurogenesis period in the cerebral cortex and cerebellum. These results strongly suggest that betaPix isoforms may play important roles in the cellular events required for brain development such as neuronal migration. PMID- 10860823 TI - Functional roles of histidine and tyrosine residues in the H(+)-peptide transporter PepT1. AB - Histidyl residues in peptide transporters PepT1 and PepT2 are believed to participate in proton and substrate binding and to be crucial to the transporters' functional activities. In the present study, we performed mutagenesis of rabbit PepT1. We mutated three histidine residues (H57, H111, and H121) predicted to reside in transmembrane segments, as well as tyrosine residues adjacent to H57. Functional analysis of wild-type and mutant PepT1 expressed in Xenopus oocytes, using both the radiotracer methods and two-microelectrode voltage-clamping, revealed that not only the H57 but also the aromatic residues near H57 were essential for the normal function of PepT1, in agreement with the concept that aromatic residues stabilize the charge on H(+) when interacting with H57. While mutagenesis at H111 did not significantly affect the activity of PepT1, mutagenesis at H121 had profound implications. The substrate affinities for H121 mutants were decreased depending both on the charge of the substrate and the charge on the substituted residues at position 121. We propose that H57 and H121 are intimately involved in the binding of the coupling ion H(+) and the recognition of transportable peptide substrates, respectively. PMID- 10860824 TI - Plasma membrane receptors for the cancer-regulating progesterone metabolites, 5alpha-pregnane-3,20-dione and 3alpha-hydroxy-4-pregnen-20-one in MCF-7 breast cancer cells. AB - Recent observations indicate that the progesterone metabolite, 5alpha-pregnane 3,20-dione (5alphaP), which is produced at higher levels in tumorous breast tissue, promotes cell proliferation and detachment, whereas 3alpha-hydroxy-4 pregnen-20-one (3alphaHP), which is produced at higher levels in nontumorous breast tissue, suppresses proliferation and detachment of MCF-7 breast cancer cells. The objective of the current study was to determine the presence and characteristics of binding sites for these endogenous putative cancer-regulating steroid hormones. Radiolabeled 5alphaP and 3alphaHP were used in radioligand binding assays on MCF-7 cell (membrane, cytosolic, and nuclear) fractions. Binding of [(3)H]5alphaP and [(3)H]3alphaHP was observed only in the plasma membrane fraction, whereas estradiol binding sites were confirmed in the cytosolic and nuclear fractions. The respective membrane binding sites exhibited specificity for the 5alphaP and 3alphaHP ligands with no appreciable displacement at 200- to 500-fold excess by other steroids. The association rate constants were calculated as 0. 107/min and 0.0089/min and the dissociation rate constants were 0. 049 9 and 0.011 for 5alphaP and 3alphaHP, respectively. Saturation analyses indicated single classes of molecules with dissociation constants of 4.5 and 4.87 nM and receptor densities of 486 and 629 fmol/mg protein, respectively, for 5alphaP and 3alphaHP. Exposure of MCF-7 cells to estradiol for 1, 24, 48, and 72 h resulted in 2.3, 4. 2-, 2.99-, and 1.7-fold increases, respectively, in 5alphaP receptor density. 3alphaHP resulted in partial suppression of the estradiol mediated increase in 5alphaP receptor density. This is the first report of receptors for the progesterone metabolites, 5alphaP and 3alphaHP, of their occurrence in breast cancer cell membranes, and of the induction of 5alphaP receptors by estradiol. The results provide further support for the potential importance of progesterone metabolites in breast cancer. PMID- 10860825 TI - Multiple promoters regulate human calcitonin receptor gene expression. AB - To initiate studies on the transcriptional regulation of the human calcitonin receptor (hCTR) gene, a 4.9-kb hCTR promoter fragment was cloned and hCTR transcriptional initiation sites were mapped in human osteoclasts, kidney, and breast cancer cell line T47D. RT-PCR detected additional hCTR transcripts initiating at least 1 kb 5' to the transcripts mapped above, demonstrating that the hCTR gene is regulated by at least two separate promoters (hCTRP1 and hCTRP2). Transcripts initiating from the upstream promoter (hCTRP2) have a novel 5' untranslated region (5' UTR). Transfection of T47D breast cancer cells with hCTR promoter/luciferase deletion constructs demonstrated that the cloned 4.9-kb hCTR promoter fragment contains both hCTRP1 and hCTRP2 promoters. Fine deletion mapping of the more transcriptionally active hCTRP1 promoter demonstrated that only 97 bp of the hCTRP1 5' flanking region is required for the majority of its transcriptional activity. PMID- 10860826 TI - Prostaglandin E(2) upregulates cyclooxygenase-2 expression in lipopolysaccharide stimulated RAW 264.7 macrophages. AB - Prostaglandin E(2) (PGE(2)) has been implicated in the regulation of inflammatory and immunological events. Using RAW 264.7 macrophages, the present study investigates the influence of PGE(2) on the expression of cyclooxygenase-2 (COX 2). Incubation of cells with PGE(2) increased lipopolysaccharide (LPS)-induced COX-2 mRNA levels in a concentration-dependent manner. Upregulation of COX-2 expression by PGE(2) was completely abolished by the specific adenylyl cyclase inhibitor 2',5'-dideoxyadenosine and mimicked by butaprost, a selective agonist of the adenylyl cyclase-coupled PGE(2) receptor subtype 2 (EP(2)), or 11-deoxy PGE(1), an EP(2)/EP(4) receptor agonist. By contrast, the EP(3)/EP(1) receptor agonists 17-phenyl-omega-trinor PGE(2) and sulprostone left LPS-induced COX-2 expression virtually unaltered. Upregulation of LPS-induced COX-2 expression and subsequent PGE(2) synthesis was also observed in the presence of the cell permeable cAMP analogue dibutyryl cAMP and the adenylyl cyclase activator cholera toxin. Together, our data demonstrate that PGE(2) potentiates COX-2 mRNA expression via an adenylyl cyclase/cAMP-dependent pathway. In conclusion, upregulation of COX-2 expression via an autocrine feed-forward loop may in part contribute to the well-known capacity of PGE(2)/cAMP to modulate inflammatory processes. PMID- 10860827 TI - Transcriptional regulation of human cardiac homeobox gene CSX1. AB - Cardiac homeobox gene Csx/Nkx-2.5 is essential for normal heart development and morphogenesis and is the earliest marker for cardiogenesis. To elucidate the regulatory mechanisms of Csx/Nkx-2.5 expression, we have isolated and characterized the upstream regulatory region of human Csx/Nkx-2.5 (CSX1). Transfection of the reporter gene containing a 965-bp CSX1 5' flanking region indicated that this region confers cardiomyocyte-predominant expression of CSX1. Deletion and mutational analyses revealed two positive cis-regulatory elements in this region that are essential for CSX1 expression in cardiomyocytes. Electrophoretic mobility shift assay revealed that nuclear proteins prepared from cardiac myocytes bound to these elements in a sequence-specific manner. The identification of cis-regulatory sequences of the Csx/Nkx-2.5 gene will facilitate further analysis for the upstream regulatory factors that control the expression of Csx/Nkx-2.5 and the process of vertebrate heart development. PMID- 10860828 TI - Acetaminophen inhibits iNOS gene expression in RAW 264.7 macrophages: differential regulation of NF-kappaB by acetaminophen and salicylates. AB - Acetaminophen is a widely used analgesic and anti-inflammatory drug that is considered a good alternative to salicylates for individuals who cannot tolerate salicylates. Nitric oxide (NO) synthesized by inducible nitric oxide synthase (iNOS) has been implicated as a mediator of inflammation. Recent evidence suggests that anti-inflammatory effect of salicylates lies in the inhibition of iNOS, but nothing has been reported about the direct effect of iNOS expression by acetaminophen. The present study was designed to elucidate sequentially the action mechanisms of acetaminophen and salicylates (aspirin and sodium salicylate) on lipopolysaccharide (LPS) plus interferon-gamma (IFN-gamma)-induced iNOS expression in RAW 264.7 macrophages. Both acetaminophen and salicylates inhibited NO production and iNOS protein expression in a dose dependent manner. Acetaminophen inhibited iNOS mRNA expression, promoter activity of iNOS gene and nuclear factor-kappa B (NF-kappaB) binding activity induced by LPS plus IFN gamma, whereas salicylates did not show any effect on them. In addition, salicylates did not affect on iNOS mRNA stability induced by LPS plus IFN-gamma. Furthermore, the inhibition of iNOS protein expression and NO production by salicylates was disappeared when salicylates were added for only 5 h to inhibit the early event of iNOS expression. Aspirin also dose dependently inhibited iNOS enzyme activity in cell-free extracts, whereas no significant differences were observed in extracts treated with sodium salicylate or acetaminophen. These findings suggest that acetaminophen may exert analgesic or anti-inflammatory effect by inhibiting iNOS expression induced by LPS plus IFN-gamma at transcriptional level by suppression of NF-kappaB binding activity, whereas salicylates exert its effect by inhibiting iNOS expression at the translational or posttranslational level. PMID- 10860829 TI - NFATz: a novel rel similarity domain containing protein. AB - Nuclear Factor of Activated T cell (NFAT) is a family of transcription factors that are important for the coordinate expression of various cytokines and immunoregulatory cell surface molecules in T cells and other types of cells in the immune system. In addition, analysis of gene disrupted mice revealed that some members of NFAT family are important for the development of myocardium, myocardial hypertrophy, and mesenchymal stem cells. NFAT family proteins have two conserved domains, the NFAT Homology Domain (NHD) and the Rel Similarity Domain (RSD). The RSD is DNA binding and AP-1 interacting domain which has structural similarity to the Rel Homology Region, the DNA binding domain of Rel family proteins. The NHD is a regulatory domain required for the Ca regulated translocation of NFAT. We report here the isolation and initial characterization of a novel RSD containing protein designated NFATz. NFATz has a RSD but no NHD. NFATz protein is localized in the nucleus without Ca signal. There is no detectable binding to a typical NFAT site even in the presence of AP-1, and it is not capable of activating transcription through the NFAT site. The chromosomal location determined by FISH revealed that NFATz and NFATx genes are in the same region. PMID- 10860830 TI - Developmental expression of maf-1 messenger ribonucleic acids in rat kidney by in situ hybridization histochemistry. AB - maf is a family of oncogenes, originally identified from the avian oncogenic retrovirus AS42, which encodes a nuclear bZip transcription factor protein. It has been reported that maf family genes have critical roles in embryological development and cellular differentiation. In this study, the distribution of maf 1 genes, the rat homologues of mafB, was examined in rat kidneys at the embryonic stages from 13 days, gestation (E13) through E21 and then 1, 2, 4, and 8 weeks after birth by in situ hybridization with (35)S-labeled antisense riboprobes. The cellular localization was determined using double in situ hybridization. Expression of maf-1 mRNA appeared weakly on E15 and was restricted to glomerular visceral epithelial cells during the pre- and postnatal stages until 2 weeks after birth and then gradually diminished. Double in situ hybridization demonstrated that maf-1 mRNA-positive cells in glomerulus also expressed Pod-1 gene, suggesting that maf-1 mRNA was expressed in the podocyte. These findings suggest that the expression of maf-1 gene may be involved in embryological development and/or differentiation of the kidney. PMID- 10860831 TI - Plant rac proteins induce superoxide production in mammalian cells. AB - The small GTP-binding protein family including Rac proteins represents a paradigm for signaling molecules shared by animal and plants. In mammalian cells, Rac induces the activation of NADPH oxidase leading to superoxide production. In plants, evidence suggests that resistance to pathogens depends on superoxide that is generated via NADPH oxidase-like enzymes. We have identified four closely related Rho/Rac genes from Zea mays that exhibit a high degree of homology to the human Rac. We hypothesized that these plant Rac proteins could function as their mammalian counterpart and activate an enzymatic complex that leads to superoxide production. Here, we show that like human Rac1, activated Zea mays Rac genes can induce superoxide production, when expressed in a mammalian system: NIH 3T3 cells. Our results suggest that in plants, Rac proteins can function as activators of oxidative burst and indicate the remarkable functional and structural conservation of Rho/Rac proteins between plant and animal kingdoms during evolution. PMID- 10860832 TI - Dendritic translocation of the rat ferritin H chain mRNA. AB - To elucidate the mechanism regulating the selective transport of mRNAs to synaptic sites, we compared the synaptosomal mRNAs with those from the forebrain using the differential display method. The ferritin H chain mRNA was found to be highly enriched in the synaptosomes. In situ hybridization for the ferritin H chain mRNA in the cultured dissociated neurons and in the hippocampal brain slices demonstrated its existence in the dendritic region. These data clearly indicate the selective translocation of the ferritin H chain mRNA into the dendrites and suggested the local expression of ferritin at the synapse. PMID- 10860833 TI - Novel Cdk inhibitors restore TGF-beta sensitivity in cdk4 overexpressing epithelial cells. AB - Transforming growth factor-beta (TGF-beta) is a potent mitogen that effects a wide variety of cells by blocking cell growth. TGF-beta acts by interacting with components of cell cycle machinery to cause G1 arrest and in mink lung epithelial cells (Mv1Lu) it does so by inhibiting Cdk4 synthesis. Overexpression of Cdk4 in these cells (B7) renders them resistant to the effects of TGF-beta. Here we report that two novel Cdk inhibitors (pyridopyrimidines) that not only inhibit Cdk4 and Cdk2 in an in vitro kinase assay but also, in the absence of TGF-beta, block growth of Mv1Lu cells in G1 more efficiently than their B7 (overexpressing Cdk4) counterparts. Interestingly, these inhibitors restored sensitivity of B7 cells towards TGF-beta. This may have implications for the treatment of tumors that have lost TGF-beta responsiveness due to deregulated cellular growth in vivo. These Cdk inhibitors could therefore be used in conjunction with TGF-beta to understand the mechanism of growth arrest in normal versus tumour cells. PMID- 10860834 TI - Computational and experimental analysis identifies many novel human genes. AB - Because of advances in automation, human genomic sequences are being deposited in public databases at a dramatic rate. However, the process of detecting genes in these sequences is still something of an art. Here we describe the implementation and testing of a relatively straightforward computational approach, the Virtual Transcribed Sequence project, which analyzes their gene content using the gene prediction program GENSCAN (GENSCAN 1.0 1,2) in combination with similarity-based methods. This approach identifies many novel human genes not found even in EST databases. PMID- 10860835 TI - Activation of human CAII gene promoter by v-Src: existence of Ras-dependent and independent pathways. AB - Carbonic anhydrase II (CAII) catalyzes the reversible hydration of carbon dioxide and plays key roles in acid base homeostasis in mammals. We found that human CAII gene promoter could be activated in human cells such as HeLa and T47D cells when the CAII promoter-luciferase gene was transfected with v-Src and assayed as a reporter of the promoter activity. Kinase negative mutants of Src, in contrast, showed little activation. The activation was completely suppressed with the introduction of a dominant-negative Ras in T47D cells, while no suppression was observed in HeLa cells. Introduction of various kinds of deletions into the CAII promoter revealed two essential regions responsible for this activation. No activation, however, was observed in activated Fyn-transfected human cells or in v-Src-transfected rodent cells. These findings suggest that Src can modulate the human CAII promoter by exerting its tyrosine kinase activity in certain human cells, and that two types of Src signaling pathways, Ras-dependent and independent, exist in a cell type dependent manner. PMID- 10860836 TI - Polyglutamine-mediated aggregation and cell death. AB - The expansion of CAG repeats is the genetic defect underlying eight neurodegenerative diseases. A common feature of these disorders is the presence of intracellular aggregates in neuronal cells. It is still unclear the significance of these cellular inclusions in the neurodegenerative process, since cell death without aggregate formation has been reported. We have constructed a synthetic fusion protein containing 17 or 43 CAG repeats and the green fluorescent protein that recapitulates the features of CAG-expanded alleles. Expression of 43, but not 17 CAG repeats results in formation of nuclear aggregates in human neuroblastoma cells. Moreover, the normal allele (17 CAG) is sequestered in the inclusion bodies. The presence of nuclear inclusions tightly correlates with apoptosis in cells expressing the protein encoding 43 CAG repeats. Cells harboring nuclear aggregates stop proliferation and undergo apoptosis. Moreover, the inhibition of protein degradation pathway increases intracellular aggregates and cell death. These data indicate that intranuclear aggregates induce apoptosis and suggest that the degradation of unfolded proteins improves cell survival. PMID- 10860837 TI - Constitutive cellular expression of PI 3-kinase is distinct from transient expression. AB - The discovery that the PTEN tumor suppressor encodes a phosphoinositide 3 phosphatase has raised interest in the effects of constitutive activation of PI 3 kinase. To gain insight into PI 3-kinase function, we have stably expressed a myristoylated form of the catalytic subunit p110alpha (myr-p110) in cells. The myr-p110 associated with the endogenous p85 regulatory subunit and retained lipid and protein kinase activity. Stable lines expressing myr-p110 had 2- to 4-fold more PI 3-kinase activity than controls. Expression of myr-p110 altered cellular morphology and increased the saturation density in culture. These clones were morphologically transformed but Akt and pp70(s6k) were not constitutively activated in contrast to transient assays and from tumor cell lines deficient in PTEN. In addition, the ability of PDGF to induce activation of Akt and pp70(s6k) was diminished. Therefore, expression of a myristoylated PI 3-kinase in murine fibroblasts induces a morphological transformation of the cells. PMID- 10860838 TI - Ligand binding properties of binary complexes of heparin and immunoglobulin-like modules of FGF receptor 2. AB - Epithelial cells, which express FGFR2IIIb, bind and respond to FGF-1, FGF-7 and FGF-10, but not FGF-2. Stromal cells, which bind and respond to FGF-1 and FGF-2, but not FGF-7 and FGF-10, express FGFR2IIIc or FGFR1IIIc. Here we show that when both isolated FGFR2betaIIIb and FGFR2betaIIIc or their common Ig module II are allowed to affinity select heparin from a mixture, the resultant binary complexes bound FGF-1, FGF-2, and FGF-7 with nearly equal affinity. In addition, FGF-2 and FGF-7 bound to both heparin-Ig module IIIb and IIIc complexes, but FGF-1 bound to neither Ig module III. The results show that in isolation both Ig modules II and III of FGFR2 can interact with heparin and that each exhibits a binding site for FGF. We suggest that the specificity of FGFR2IIIb and FGFR2IIIc is dependent on the cell membrane environment and heparin/heparan sulfate. Ig modules II and III cooperate both within monomers and across dimers with cellular heparan sulfates to confer cell type-dependent specificity of the FGFR complex for FGF. PMID- 10860839 TI - Structure and developmental expression of the mouse CCK-B receptor gene. AB - Cholecystokinin (CCK) and gastrin exert their effects through two receptors, the CCK-A and CCK-B receptors. We have cloned the mouse CCK-B receptor gene (Cckbr) and determined its complete genomic structure, nucleotide sequence, and tissue specific expression pattern. Cckbr is divided into five exons spanning 11 kb. A primer extension assay was used to map the transcription initiation site to 199 bp upstream of the translational start site. Rapid amplification of cDNA ends was used to localize the 3' end downstream of an atypical polyadenylation site (GATAAA). Mouse Cckbr transcripts were most abundant in brain and stomach, but were also detected in colon, kidney, ovary, and pancreas. Prenatal expression of both CCK-A and CCK-B receptors in various tissues was analyzed by RT-PCR. The expression pattern was similar to the adult pattern, suggesting that receptor transcription is an early event in gastrointestinal development. PMID- 10860840 TI - Structural basis of functional mimicry between carbohydrate and peptide ligands of con A. AB - Crystallographic studies have shown independent binding sites for sugar and peptide ligands of concanavalin A, although they were considered functional mimics based on biochemical experiments. The topological correlation of 12 residue peptide with different carbohydrate ligands of concanavalin A showed similarity between trimannose and the YPY region of the peptide establishing structural mimicry. Molecular docking of trimannose and the YPY motif on the reciprocal binding sites revealed equivalent interactions and energetics implying that the peptide-binding sites may constitute additional sugar-binding subsites of concanavalin A. The binding of a mannose-rich neoglycoprotein with significantly higher affinity compared with that of the methyl alpha-d mannopyranoside is consistent with this interpretation. PMID- 10860841 TI - Modulation of the chaperone activities of Hsc70/Hsp40 by Hsp105alpha and Hsp105beta. AB - Hsp105alpha and Hsp105beta are stress proteins found in various mammals including human, mouse, and rat, which belong to the Hsp105/Hsp110 protein family. To elucidate their physiological functions, we examined here the chaperone activity of these stress proteins. Hsp105alpha and Hsp105beta prevented the aggregation of firefly luciferase during thermal denaturation, whereas the thermally denatured luciferase was not reactivated by itself or by rabbit reticulocyte lysate (RRL). On the other hand, Hsp105alpha and Hsp105beta suppressed the reactivation of thermally denatured luciferase by RRL and of chemically denatured luciferase by Hsc70/Hsp40 or RRL. Furthermore, although Hsp105alpha and Hsp105beta did not show ATPase activity, the addition of Hsp105alpha or Hsp105beta to Hsc70/Hsp40 enhanced the amount of hydrolysis of ATP greater than that of the Hsp40 stimulated Hsc70 ATPase activity. These findings suggest that Hsp105alpha and Hsp105beta are not only chaperones that prevent thermal aggregation of proteins, but also regulators of the Hsc70 chaperone system in mammalian cells. PMID- 10860842 TI - Both amino- and carboxyl-terminal domains of TRAF3 negatively regulate NF-kappaB activation induced by OX40 signaling. AB - OX40 is a member of the tumor necrosis factor receptor (TNF-R) superfamily. We observed that overexpression of OX40 activated NF-kappaB, which was inhibited by dominant negative forms of TRAF2, NF-kappaB-inducing kinase (NIK), and IkappaB kinase (IKK) alpha. This indicates that OX40 signaling leads to NF-kappaB activation through the same cascade as TNF-R2. We then investigated the negative regulatory function of TRAF3 on OX40-induced NF-kappaB activation. TRAF3 blocked OX40-, TRAF2-induced NF-kappaB activation, but not NIK- and IKKalpha-induced NF kappaB activation, indicating that TRAF3 blocks the pathway between TRAF2 and NIK. C-terminal deletion mutants as well as the N-terminal deletion mutant of TRAF3 inhibited NF-kappaB activation induced by OX40 or TRAF2. Since TRAF3 bound to OX40 through the C-terminal TRAF domain, the C-terminal domain is likely to work as a dominant negative mutant to compete the recruitment of TRAF2 to the receptor, which transmits the signal from OX40 to the downstream, NIK kinase. On the other hand, the N-terminal domain of TRAF3 seems to affect the downstream of TRAF2 binding. Thus, it is suggested that TRAF3 actively inhibits NF-kappaB activation induced by OX40. PMID- 10860843 TI - Identification of a GPI-anchored type HDL-binding protein on human macrophages. AB - To identify the HDL3-binding proteins on human macrophages, we examined the involvement of GPI-anchored protein in the binding of HDL3, and tried to purify HDL3-binding protein. From membrane fractions of macrophages, we obtained 80- and 130-kDa HDL3-binding proteins by ligand blotting. Treatment of macrophages with phosphatidylinositol-specific phospholipase C (PI-PLC) significantly decreased the specific HDL3-binding in a dose-dependent manner. Furthermore, treatment with mannosamine, which blocks GPI-anchor formation, decreased specific HDL3-binding in a dose-dependent manner. PI-PLC treatment released from the cells the proteins with an M(r) of 80 kDa, which could also bind HDL3. PI-PLC as well as mannosamine treatment markedly reduced cholesterol efflux from macrophages in association with the decreased HDL-binding. Using HDL3-affinity chromatography, we purified 80-kDa GPI-anchored type HDL3-binding protein. In summary, we demonstrate the implication of 80-kDa GPI-anchored protein in the binding of HDL3 to human macrophages, which might have some role in reverse cholesterol transport. PMID- 10860844 TI - High-density lipoproteins protect endothelial cells from tumor necrosis factor alpha-induced apoptosis. AB - High-density lipoproteins (HDL) levels have been shown to be inversely correlated with coronary heart disease, but the mechanisms of the direct protective effect of HDL on endothelial cells are not fully understood. The apoptosis of endothelial cells induced by cytokines and/or oxidized low-density lipoproteins, etc. may provide a mechanistic clue to the "response-to-injury" hypothesis of atherogenesis. Here we report that HDL prevent the apoptosis of human umbilical venous endothelial cells (HUVECs) induced by tumor necrosis factor-alpha (TNF alpha) via an inhibition of CPP32-like protease activity. The incubation of HUVECs with TNF-alpha significantly increased the CPP32-like protease activity, and induced apoptosis. Preincubation of HUVECs with HDL before incubation with TNF-alpha significantly suppressed the increase in the CPP32-like protease activity, preventing apoptosis in a concentration-dependent manner. These results suggest that HDL prevent the suicide pathway leading to apoptosis of endothelial cells by decreasing the CPP32-like protease activity and that HDL thus play a protective role against the "response-to-injury" hypothesis of atherogenesis. PMID- 10860845 TI - Dominant expression of a novel splice variant of caspase-8 in human peripheral blood lymphocytes. AB - Caspase-8 is an apical and critical proteolytic enzyme in the cascade of apoptosis. As a result of alternative splicing, the generation of at least 7 isoforms of caspase-8 has been reported. The existence of multiple isoforms that lack the essential domains for apoptosis suggests the possible role of these isoforms on the regulation of apoptosis. Here we report a novel longer isoform of caspase-8 (caspase-8L) that was generated by alternative splicing of intron 8, thereby carrying a 136-bp insertion and frame shift of the transcript. The transcript encoded N-terminal two repeats of death effector domain (DED) of caspase-8, but lacking the C-terminal half of the proteolytic domain. Reverse transcriptase (RT)-polymerase chain reaction (PCR) analysis revealed the dominant expression of caspase-8L transcript compared to the intact form of caspase-8 in human peripheral blood lymphocyte (PBL) and T cells. In patients with systemic lupus erythematosus (SLE), imbalanced expression of caspase-8L transcript was identified. These results suggest the important role of caspase-8L in the modulation of apoptosis. PMID- 10860846 TI - The repressive function of AP2 transcription factor on the hepatocyte growth factor gene promoter. AB - Hepatocyte growth factor is an important multifunctional growth factor whose gene expression is tightly regulated at the transcriptional level. Previous studies from our laboratory have shown that several cis-acting elements are present in the promoter and proximal promoter region of the HGF gene. In this study, we have uncovered that AP2 transcription factor specifically binds to a regulatory site located at -230 to -260 in the upstream region of the HGF gene promoter. Gelshift and supershift assays confirmed that AP2 has high binding affinity to this region. Functional studies which introduced a mutation in the AP2 core binding region as well as cotransfection experiments using an AP2 expression vector revealed that AP2 exerts a repressive role on the HGF gene promoter activity. The AP2 binding site overlaps with those of NF1 and USF/E-box binding sites which we have recently shown to constitute a composite multifunctional docking site for the members of the NF1 and USF transcription factor families. An inverse correlation was noted between AP2 binding activity to this composite site and HGF gene expression in different cell lines. Therefore, AP2-mediated repression of the HGF gene promoter may be part of the molecular mechanism responsible for regulating HGF expression. PMID- 10860847 TI - Morphological alteration of X-ray induced partially transformed human cells by transfection with a small c-myc DNA sequence. AB - During attempts to transform a normal human fibroblast strain (GM730) by X irradiation, we obtained a partially transformed cell strain (GM730pt) which demonstrates several aspects of the transformed phenotype including morphological changes, increased saturation density, growth in soft agar, and focus formation in long-term cultures. When GM730pt cells were transfected with the feline c-myc gene, morphology of the cells changed dramatically following seven days of expression. Transfection of other plasmid DNAs or oncogenes such as pUC8, pSV2neo, src, sis, and H-ras had little or no effects on the phenotype of GM730pt cells. On the other hand, a gel purified, small fragment of c-myc DNA had a complete cell alteration activity. Furthermore, Bal 31 deletion and M13 sequencing experiments showed that the alteration seen in GM730pt cells is delimited to a 24 nucleotide stretch (active myc element) from the second intron of the feline c-myc gene that contains a T-rich sequence. PMID- 10860848 TI - Temperature dependence of force, velocity, and processivity of single kinesin molecules. AB - Using the bead assay in optical microscopy equipped with optical tweezers, we have examined the effect of temperature on the gliding velocity, force, and processivity of single kinesin molecules interacting with a microtubule between 15 and 35 degrees C. The gliding velocity increased with the Arrhenius activation energy of 50 kJ/mol, consistent with the temperature dependence of the microtubule-dependent ATPase activity. Also, the average run length, i.e., a measure of processivity of kinesin, increased on increasing temperature. On the other hand, the generated force was independent of temperature, 7.34 +/- 0.33 pN (average +/- S.D., n = 70). The gliding velocities decreased almost linearly with an increase in force irrespective of temperature, implying that the efficiency of mechano-chemical energy conversion is maintained constant in this temperature range. Thus, we suggest that the force generation is attributable to the temperature-insensitive nucleotide-binding state(s) and/or conformational change(s) of kinesin-microtubule complex, whereas the gliding velocity is determined by the ATPase rate. PMID- 10860849 TI - Hepatocyte growth factor enhances MMP activity in human endothelial cells. AB - Scatter factor (SF) or hepatocyte growth factor (HGF) has been identified as an angiogenic factor. Angiogenesis requires not only tube formation but also invasion of pericytes and extracellular matrix (ECM) remodeling to promote new vessel stabilization. In the current study, the effect of SF/HGF on endothelial cell (EC) production of matrix metalloproteinases (MMPs) was explored. We showed that SF/HGF enhanced MT1-MMP synthesis and induced MMP-2 activation in two human EC lines: dermal microvessel EC and coronary arterial EC. Furthermore, SF/HGF accelerated EC invasion into matrix, an activity that could be inhibited by a MMP inhibitor. We also demonstrated that the MAP kinase cascade is critical in signal transduction pathway from SF/HGF stimulation to MT1-MMP up-regulation. The current study indicates that MMP activation is a novel effect of SF/HGF on ECs. PMID- 10860850 TI - alpha(1)-adrenoceptor subtypes differentially couple to growth promotion and inhibition in Chinese hamster ovary cells. AB - We have compared the coupling of human alpha(1A)-, alpha(1B)-, and alpha(1D) adrenoceptors (expressed at approximately 2000 fmol/mg protein in Chinese hamster ovary cells) to cellular growth promotion (as assessed by [(3)H]thymidine incorporation) and related signaling mechanisms. Maximum elevation of intracellular Ca(2+) by the three subtypes occurred with the rank order alpha(1A) (1691 nM) > alpha(1D) (1215 nM) > alpha(1B) (360 nM). In contrast, activation of the ERK, JNK, and p38 forms of mitogen-activated protein kinases occurred with the rank order alpha(1D) > alpha(1A) > alpha(1B). alpha(1A)-Adrenoceptor stimulation inhibited basal and growth factor-stimulated [(3)H]thymidine incorporation by 74%, and this was mitigated by p38 inhibition. In contrast, alpha(1D)-adrenoceptor stimulation enhanced cellular growth by 136%, and this was blocked by two distinct inhibitors of ERK activation. We conclude that within a given cell type alpha(1)-adrenoceptor subtypes can have opposite effects on cellular growth, although their proximal signal transduction displays only quantitative differences. PMID- 10860851 TI - Structural and functional characterizations of the 5'-flanking region of the mouse glucagon receptor gene: comparison with the rat gene. AB - A putative proximal promoter was defined previously for the mouse glucagon receptor (GR) gene. In the present study, a distal promoter was characterized upstream from a novel non-coding exon revealed by the 5'-rapid amplification of cDNA ends from mouse liver tissue. The 5'-flanking region of the mouse GR gene was cloned up to 6 kb and the structural organization was compared to the 5' untranslated region of the rat gene cloned up to 7 kb. The novel exon, separated by an intron of 3.8 kb from the first coding exon, displayed a high homology (80%) with the most distal of the two untranslated exons found in the 5' region of the rat GR gene. The mouse distal promoter region, extending up to -1 kb from the novel exon, displayed 85% identity with the rat promoter. Both contain a highly GC-rich sequence with five putative binding sites for Sp1, but no consensus TATA or CAAT elements. To evaluate basal promoter activities, 5' flanking sequences of mouse or rat GR genes were fused to a luciferase reporter gene and transiently expressed in a mouse and in a rat cell line, respectively or in rat hepatocytes. Both mouse and rat distal promoter regions directed a high level of reporter gene activity. Deletion of the Sp1 binding sites region or mutation of the second proximal Sp1 sequence markedly reduced the distal promoter activity of the reporter gene. The mouse proximal promoter activity was 2- to 3 fold less than the distal promoter, for which no functional counterpart was observed in the similar region of the rat gene. PMID- 10860852 TI - VIP activates G(s) and G(i3) in rat alveolar macrophages and G(s) in HEK293 cells transfected with the human VPAC(1) receptor. AB - We have characterized vasoactive intestinal peptide (VIP) receptor/G-protein coupling in rat alveolar macrophage (AM) membranes and find that pertussis toxin treatment and antisera against G(alphai3) and G(alphas) reduce high-affinity (125)I-VIP binding, indicating that both G(alphas) and G(alphai3) couple to the VIP-receptor. The predominant VIP-receptor subtype in AM is VPAC(1) and we examined the G-protein interactions of the human VPAC(1) that had been transfected into HEK293 cells. VPAC(1) has a molecular mass of 56 kDa; GTP analogs reduced (125)I-VIP binding to this protein demonstrating that high affinity binding of VIP to the receptor requires coupling to G-protein. Functional VIP/VPAC(1)/G-protein complexes were captured by covalent cross linking and analyzed by Western blotting. The transfected human VPAC(1) receptor in HEK293 was found to be coupled to G(alphas) but not G(alphai) or G(alphaq). Furthermore, pertussis toxin treatment had no effect on VPAC(1)/G-protein coupling in these cells. These observations suggest that the G-proteins activated by VPAC(1) may be dependent upon species and cell type. PMID- 10860853 TI - Cloning and characterization of alphaP integrin in embryos of the sea urchin Strongylocentrotus purpuratus. AB - Differentially expressed integrins have been shown to be involved in the intricate cell movements that occur during early development. Because the migration and movement of cells have been well characterized in sea urchin embryos, we searched for alpha-integrin subunits in this organism. An alpha integrin subunit, alphaP, was cloned from Strongylocentrotus purpuratus mesenchyme blastula stage mRNA by RT-PCR and RACE and found to exhibit 74-77% sequence similarity to mammalian alpha(5), alpha(8), alpha(IIb), and alpha(v) integrin. The 8-kb transcript was most abundant at the prism stage, although low levels could be detected at all stages by Northern blot analysis and RT-PCR. A polyclonal antibody to this novel integrin was generated against a 100-amino-acid alphaP fragment fused to glutathione S-transferase and shown to recognize a 180 kDa alpha-integrin in the egg and in all stages of embryogenesis studied. PMID- 10860854 TI - Translocation of TRAF proteins regulates apoptotic threshold of cells. AB - Tumor necrosis factor (TNF) receptor-associated factors (TRAFs) are involved in signaling pathways triggered by members of the TNF receptor (TNFR) family and other cell surface proteins. After recruitment to a receptor, TRAFs initiate formation of multiprotein complexes that induce downstream events, such as translocation of transcription factor nuclear factor kappaB (NF-kappaB) and activation of c-Jun N-terminal kinase (JNK). Several proteins in these complexes play important roles in regulation of apoptosis. However, the fate of TRAF containing complexes once assembled in response to receptor multimerization is not understood. In this report, we demonstrate that crosslinking of TNFR family members or interaction of TRAF2 with the cytoplasmic protein A20 leads to intracellular translocation of TRAF2. This redistribution leads to depletion of the cytoplasmic pool of TRAF2. The ratio between soluble and insoluble TRAF2 determines the sensitivity of cells to TNF-alpha-induced apoptosis and may play an important role in limiting further TRAF-dependent signal transduction. PMID- 10860855 TI - Porphyrin biosynthesis intermediates are not regulating delta-aminolevulinic acid transport in Saccharomyces cerevisiae. AB - In Saccharomyces cerevisiae, as in all eukaryotic organisms, delta-aminolevulinic acid (ALA) is a precursor of porphyrin biosynthesis, a very finely regulated pathway. ALA enters yeast cells through the gamma-aminobutyric acid (GABA) permease Uga4. The incorporation of a metabolite into the cells may be a limiting step for its intracellular metabolization. To determine the relationship between ALA transport and ALA metabolization, ALA incorporation was measured in yeast mutant strains deficient in the delta-aminolevulinic acid-synthase, uroporphyrinogen III decarboxylase, and ferrochelatase, three enzymes involved in porphyrin biosynthesis. Results presented here showed that neither intracellular ALA nor uroporphyrin or protoporphyrin regulates ALA incorporation, indicating that ALA uptake and its subsequent metabolization are not related to each other. Thus a key metabolite as it is, ALA does not have a transport system regulated according to its role. PMID- 10860856 TI - Refinement of the expression pattern of a mouse homologue of the porcine 135-kDa alpha-d-mannosidase (MAN2B2). AB - In the article by Hiramoto et al. (1997) (Stage-specific expression of a mouse homologue of the porcine 135 kDa alpha-d-mannosidase (MAN2B2) in type A spermatogonia, Biochem. Biophys. Res. Commun. 241, 439-445) a new homologue to the porcine epididymis-specific 135 kDa alpha-d-mannosidase (MAN2B2) was cloned from a mouse testis cDNA library. Northern blot analysis with RNA of different tissues showed a testis-specific expression by using a mouse MAN2B2 oligonucleotide. In situ hybridization revealed that the mouse MAN2B2 transcript is localized exclusively in spermatogonia. In consequence to this it was proposed by Hiramoto et al. that the mouse MAN2B2 homologue could serve as a marker for spermatogonia. By repeating the published experiments we observed a different expression pattern of the mouse MAN2B2 gene. Northern blot analysis with either testicular RNA from prepubertal males or with testicular RNA from the W/W(v) mutant mouse which lacks germ cells showed an expression of the MAN2B2 gene. Furthermore, Northern blot analysis with RNA from different somatic tissues revealed that the gene is ubiquitously expressed. Therefore, our refinement clearly demonstrates that the MAN2B2 mouse homologue is not specifically expressed in spermatogonia. PMID- 10860857 TI - Characterization of insulin receptor substrate 3 in rat liver derived cells. AB - In rat liver derived HTC cells transfected with and expressing human insulin receptors, there are multiple p60-70 proteins that are tyrosine phosphorylated following insulin treatment of cells. Employing antibodies to insulin receptor substrate 3 (alpha-IRS-3), we found that IRS-3 is a major p60 phosphoprotein that is tyrosine phosphorylated following insulin treatment of cells and interacts with phosphatidylinositol-3-kinase (PI3K). Majority of IRS-3 when phosphorylated appears to interact with PI3K. Tyrosine phosphorylation of IRS-3 is robust at 2 min and steadily increases up to 30-90 min of insulin treatment. Following insulin treatment of cells, some high molecular weight phosphoproteins are coimmunoprecipitated with alpha-IRS-3. In summary, IRS-3 is the major p60 protein that is tyrosine phosphorylated and interacts with PI3K in HTC rat liver derived cells following insulin treatment of cells. Unlike related IRS-1/2 that is transiently phosphorylated, IRS-3 shows robust and prolonged tyrosine phosphorylation upon insulin treatment of cells and may play a role in delayed and/or prolonged insulin actions. PMID- 10860858 TI - Sensitivity of orexin-A binding to phospholipase C inhibitors, neuropeptide Y, and secretin. AB - The binding of [(125)I] orexin-A (Ox-A) to particulates from Chinese hamster ovary (CHO) cells expressing the cloned orexin-A receptor, or from rat forebrain areas, was sensitive to blockers of phosphatidylinositol-specific phospholipase C (PtdIns-PLC) U-73122 and ET-18-OCH(3), little affected by phospholipase A(2) inhibitor quinacrine, and not sensitive to D609, a xanthate inhibitor of phosphatidylcholine-selective PLC. Interaction of the receptor with a PtdIns-PLC was further indicated by a large sensitivity of the binding to Ca(2+). Up to 50% of the binding was sensitive to the G-protein nucleotide site agonist GTP-gamma S. Ligand attachment to the orexin-A receptor thus depends on an association with both PtdIns-PLC and G-protein alpha-subunits. In all paradigms examined, the binding of [(125)I]orexin-A was competed by human/rat neuropeptide Y (hNPY) and porcine secretin with a potency similar to orexin-A (IC(50) range 30-100 nM). The rank order of potency for NPY-related peptides was hNPY > porcine peptide YY (pPYY) > (Leu(31), Pro(34)) human PYY > human PYY(3-36) > hNPY free acid > human pancreatic polypeptide. Among secretin-related peptides, the rank order of potency was porcine secretin > or = orexin-A > human pituitary adenylate cyclase activating peptide > orexin-B > porcine vasoactive intestinal peptide. Among opioid peptides, rat beta-endorphin and camel delta-endorphin were much less active than NPY and secretin, and two enkephalins were inactive at 1 microM. In view of high abundance of NPY in forebrain, the above cross-reactivity could indicate a significant contribution of NPY to signaling via orexin-A receptors. PMID- 10860859 TI - Telomerase: not just black and white, but shades of gray. AB - Telomerase, the telomeric DNA reverse transcriptase, plays a key role in the maintenance of telomeres in mammals and is required for immortalization of primary cells. Inexplicably, telomerase activation is sometimes associated with telomere shortening and inhibition leads not only to apoptosis but also increased tumorigenicity in rapidly renewing tissues of mouse and man. This article reviews the current evidence, both in vitro and in vivo, for telomerase function and the potential mechanisms, downstream of telomerase, in telomere signaling involving both the tumor-suppressor p53-dependent and independent pathways. PMID- 10860860 TI - The LIM-domain protein FHL1 (SLIM 1) exhibits functional regulation in skeletal muscle. AB - The LIM domain protein FHL1 (SLIM 1) transcript is preferentially expressed in postnatal skeletal muscle but almost nothing is known about its function in this tissue. In this study we have examined the expression of the FHL1 transcript at the cellular level by in situ hybridisation. Muscle fibers exist as a number of discrete subpopulations or "types" which are differentiated by their contractile and metabolic properties. It was observed that the FHL1 transcript was not fiber type specific but was however more abundant in oxidative fibers. Muscle atrophy induced by disuse caused a significant decline in the expression of the transcript but atrophy induced by short-term denervation did not. Hypertrophy of skeletal muscle induced by passive stretch was associated with an up-regulation of the FHL1 transcript. These data are consistent that FHL1 is involved in synthetic processes within the muscle fibre. PMID- 10860861 TI - Polymorphism of the 5' untranslated region of NHE1 gene associated with type-I diabetes. AB - The ubiquitous form of the sodium-hydrogen exchanger, NHE1, is devoted to the regulation of intracellular pH and cell volume. In addition, NHE1 activity is stimulated by growth factors and increased NHE rates are found in both circulating and immortalized cells during diabetes or diabetic nephropathy. In this context, we searched for polymorphisms of the 5'-flanking regulatory region of NHE1 gene in subjects with type-I diabetes. We identified a C/T transition 696 bases upstream the translation initiation start site which disrupts a repeated palindromic GC sequence. The TT genotype was significantly more frequent in type 1 diabetics and may have functional importance. Genetic linkage between NHE1 and diabetes has been previously described in NOD mice strains with consequences on NHE rates. Hence, the polymorphism described hereby may act as a predisposition factor to type-I diabetes or to diabetic complications, and may be useful to investigate the genetic involvement of NHE1 in human pathophysiology. PMID- 10860862 TI - Dental cells express factors that regulate bone resorption. AB - Odontoblasts and osteoblasts produce similar highly mineralized extracellular matrices. In bone, osteoblasts/stromal cells regulate osteoclast (ocl) formation and bone resorption by producing factors like osteoprotegerin (OPG), osteoclast differentiating factor (ODF/RANKL), and macrophage colony-stimulating factor (M CSF) that interact with hematopoietic ocl precursor cells. Using odontoblast and pulp cell lines, we detected a constitutive expression of OPG, RANKL, and M-CSF mRNA in both cell types. OPG and RANKL proteins were also detectable. In vivo, RANKL and OPG were localized to odontoblasts, ameloblasts, and pulp cells in developing mouse teeth by immunohistochemistry. In a coculture system, we found the dental cells to be inhibitory to ocl formation from spleen and bone marrow precursors, despite their production of osteoclast stimulatory factors. Our data indicate for the first time that dental cells express factors important in regulation of osteoclastogenesis and bone resorption. Since both stimulatory (RANKL, M-CSF) and inhibitory (OPG) factors are expressed, a balance between positive and negative factors may contribute to regulation of bone resorption. PMID- 10860863 TI - Association of the inositol (1,4,5)-trisphosphate receptor ligand binding site with phosphatidylinositol (4,5)-bisphosphate and adenophostin A. AB - The inositol 1,4,5-trisphosphate receptor (InsP(3)R) is activated by InsP(3) binding to amino-terminal ligand binding domain (InsP(3)R-N). Recently we reported functional coupling of phosphatidylinositol (4, 5)-bisphosphate (PIP(2)) to the InsP(3)R. Specific binding of PIP(2) to InsP(3)R-N domain was postulated as a part of the InsP(3)R-PIP(2) functional coupling model. Here we utilized bacterially expressed and purified InsP(3)R-N domain to characterize its binding specificity for InsP(3), Adenophostin A (AdA) and the water-soluble PIP(2) analog dioctanoyl-(4,5)PIP(2) (ShPIP(2)). Obtained data led us to conclude that specific InsP(3), AdA, and ShPIP(2) binding sites are located within the InsP(3)R-N domain, that the extra receptor binding element responsible for enhanced binding of AdA is an integral part of the InsP(3)R-N domain, that ShPIP(2) is able to displace InsP(3) from the InsP(3)R-N, but InsP(3) or AdA is unable to completely displace ShPIP(2). These results support the InsP(3)R-PIP(2) functional coupling model and provide novel insights into InsP(3)R ligand specificity. PMID- 10860864 TI - Ligands for the peroxisome proliferator-activated receptor-gamma and the retinoid X receptor-alpha exert synergistic antiproliferative effects on human coronary artery smooth muscle cells. AB - In human coronary artery vascular smooth muscle (hcaVSM) cells, the mechanisms that mediate the antiproliferative effects of ligands for the peroxisome proliferator-activated receptor-gamma (PPAR gamma) and the retinoid X receptor alpha (RXR alpha) are unclear. Dimerization of PPAR gamma with RXR alpha and occupancy by both ligands is required for maximal activation. Accordingly, we determined whether the antiproliferative activity of the PPAR gamma ligands, troglitazone or 15-deoxy-Delta-12,14-prostaglandin J2 (15d-PGJ2), was enhanced with the RXR alpha ligand, 9-cis-retinoic acid (9-cis-RA). Incubation of actively proliferating hcaVSM cells with either troglitazone or 15d-PGJ2 resulted in a dose-dependent inhibition of proliferation with half-maximal inhibitory concentrations (IC(50)s) of 13 and 2 microM, respectively. Quiescent cells incubated with troglitazone or 15d-PGJ2 and subsequently stimulated with PDGF-BB showed a concentration-dependent decrease in the active form of MAP kinase, suggesting that inhibition of cell growth by troglitazone may involve the MAP kinase pathway, an important growth activation pathway in VSM cells. Incubation of cells with either 0.1 or 1.0 microM 9-cis-RA inhibited cell growth to a similar degree. Addition of troglitazone or 15d-PGJ2 to cells in combination with either concentration of 9-cis-RA resulted in a striking increase in growth inhibition, and was accompanied by an approximately 4-fold reduction in the IC(50)s for both PPAR gamma ligands. These findings imply that RXR alpha activation by 9-cis-RA synergistically enhanced inhibition of hcaVSM cell growth. The precise nature of this cooperative interaction between PPAR gamma and RXR alpha remains to be determined. PMID- 10860865 TI - Differential gene expression in synovium of rheumatoid arthritis and osteoarthritis. AB - Rheumatoid arthritis (RA) and osteoarthritis (OA) are the major types of arthritis. Although both diseases are characterized by joint destruction, their etiologies are different. To get insights into pathophysiological pathways, we used the suppression subtractive hybridization (SSH) method to identify differentially expressed genes in RA. DNA sequencing identified 12 gene products including cytoskeletal gamma-actin and extracellular matrix components such as fibronectin, collagen III alpha(1), and superficial zone protein. Interferon gamma-inducible genes such as a novel thiol reductase, two genes of unknown function (HSIFNIN4, RING3), and annexin II were also found. Two genes encoded proteins involved in proliferation such as elongation factor 1 alpha and the granulin precursor. Furthermore, the protease cathepsin B and synovial phospholipase A2 group IIA were detected by SSH. To confirm the differential expression of the genes, we performed RT-PCR analyses of RA and OA synovial tissues. Compared to OA patients, 9 of the 12 genes were overexpressed in RA, suggesting that SSH is a powerful tool for the detection of differential gene expression in synovial tissues. Further characterization of the gene products may help to identify pathophysiological mechanisms in arthritic diseases. PMID- 10860866 TI - Contribution of the plasmin/matrix metalloproteinase cascade to the retraction of human fibroblast populated collagen lattices. AB - To assess the contribution of the plasmin/matrix metalloproteinase cascade in lattices retraction, human gingival fibroblast-populated collagen lattices were supplemented with plasminogen. The rate of lattice retraction was enhanced by addition of plasminogen. This effect was concomitant to plasmin generation, prostromelysin-1 and procollagenase activation. Plasminogen-mediated initiation of that proteolytic cascade was accompanied by conspicuous changes in cell morphology and collagen fibers organization. At day 1 of culture fibroblasts shifted from a rounded (control) to an elongated (in presence of plgn) shape. At the latest stage of retraction, intense vacuolization around fibroblasts was noticed in plgn-supplemented lattices which paralleled the increased collagen degradation. Plgn-enhancing influence on the initial phase of lattice retraction could be totally annihilated by either aprotinin or Batimastat. Those data emphasize the crucial importance of the plasmin-MMP proteolytic cascade in granulation tissue retraction in a healing wound. PMID- 10860867 TI - Rat beta 1-adrenergic receptor regulatory region containing consensus AP-2 elements recognizes novel transactivator proteins. AB - beta 1-Adrenergic receptors (beta1-ARs) serve as important regulators of central nervous system (CNS)-mediated behavior and several neural functions, including mood, memory, neuroendocrine control, and stimulation of autonomic function. Using beta 1-AR-luciferase reporter recombinants, we have previously determined that important beta 1-AR genetic elements controlling expression within the C6 glioma cell line are contained within the region -396 to -299, relative to the translational start site. By conducting progressive internal deletions of the rat beta 1-AR 5' flanking region and with the use of beta 1-AR-luciferase recombinants, we have verified that this region contains the primary beta 1-AR promoter and/or major regulatory elements. To begin the identification of protein factors involved in beta 1-AR transcriptional activity conferred by this beta 1 AR region and flanking sequences, we conducted electrophoretic mobility shift assays using defined beta 1-AR DNA subregion probes. One probe (GS-1), encompassing the region -396 to -367, was found to produce two major and two minor mobility shift complexes when bound to nuclear extracts from the beta 1-AR expresser C6 cell line. UV-crosslinking of DNA-protein complexes, coupled with DNase I digestion, indicated that this beta 1-AR region interacts with one major protein of approximately 117 kDa molecular weight and additional minor proteins. GS-1 DNA-protein complexes were observed using beta 1-AR expresser tissues in the CNS, including cortex, hippocampus, and olfactory bulb. No DNA-protein complexes were observed when using nuclear extracts from beta 1-AR nonexpresser tissues; in some cases, using L6 cells, previously characterized to express little or no beta1-ARs, a reduction in intensities of the DNA-protein complexes was observed. Competition experiments indicate that nuclear protein binds to one of two subregions within the GS-1 sequence that contain AP-2-like consensus elements. Recombinant AP-2 protein will bind to both the beta 1-AR GS-1 promoter fragment and commercially available AP-2 consensus element control probes. Interestingly, using antibody supershift and immunoblotting experiments, no supershifts were observed and the major 117-kDa protein was not immunoreactive to antibodies recognizing either AP-2 alpha or AP-2 beta. These results support our contention that this beta 1-AR regulatory region contains AP-2 consensus elements that recognize novel transactivator proteins. PMID- 10860868 TI - Relative potency values derived from hepatic vitamin A reduction in male and female Sprague-Dawley rats following subchronic dietary exposure to individual polychlorinated dibenzo-p-dioxin and dibenzofuran congeners and a mixture thereof. AB - This study investigated the potency of individual polychlorinated dibenzo-p dioxins (PCDDs) and dibenzofurans (PCDFs) to reduce hepatic vitamin A in the rat. Dose-response relationships were determined following long-term dietary exposure to 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,4,7, 8 pentachlorodibenzofuran, 1,2,3,4,8-pentachlorodibenzofuran, 1,2,3, 7,8 pentachlorodibenzofuran, 1,2,3,6,7,8-hexachlorodibenzofuran, 1,2, 3,7,8 pentachlorodibenzo-p-dioxin, octachlorodibenzo-p-dioxin, octachlorodibenzofuran, or mixtures of some of these congeners. The aim was to estimate vitamin A-related relative potency (REP) values for each congener in relation to that of TCDD and to investigate if these values were in accordance with REP values estimated for the subchronic toxicity observed in the same study. An additional aim was to investigate if the effect on hepatic vitamin A levels was additive compared to the effect of the individual congeners. The obtained results demonstrate that hepatic vitamin A reduction occurs as a consequence of long-term low-level exposure to 2,3,7, 8-substituted but not to non-2,3,7,8-substituted congeners. Female rats were slightly more responsive to this effect as judged from the lower EC50 values for all the congeners in this sex. The vitamin A-related REP values were similar for female and male rats and were in good agreement with the estimated REP values for subchronic toxicity in the same animals. The vitamin A effect of the individual congeners in the mixture tended to be somewhat less than pure additive for male rats and very close to pure additive for female rats. In conclusion, the presented data show that reduction of hepatic vitamin A is a sensitive marker of an altered retinoid homeostasis following long-term low-dose exposure to dioxin-like compounds, which essentially conforms to their assumed additive mechanism of action. PMID- 10860869 TI - Differential expression of two CYP1A genes in rainbow trout (Oncorhynchys mykiss). AB - Differential expression of two rainbow trout CYP1A genes was measured in vivo and in vitro in response to treatment with the model CYP1A inducers beta naphthoflavone (BNF), 3-methylcholanthrene (3-MC), isosafrole (ISF), and 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD, only in vitro). Originally described by Berndtson and Chen (Arch. Biochem. Biophys. 310, 187-195, 1994) as CYP1A1 and CYP1A2, these genes were renamed CYP1A3 and CYP1A1, respectively, by the P450 nomenclature committee. A significant, differential, inducer-dependent induction of the two CYP1A mRNAs, as measured by RNase protection assay, was observed in vivo. CYP1A3 and CYP1A1 mRNA levels in liver were significantly induced 50- and 18-fold, respectively, following ip injection with BNF. Conversely, CYP1A3 and CYP1A1 mRNA levels were significantly induced 5- and 66-fold, respectively, following ip injection with 3-MC. Isosafrole had no significant effect on in vivo induction of CYP1A mRNA levels. In primary cultures of hepatocytes, BNF, 3-MC, ISF, as well as TCDD all significantly induced CYP1A3 and CYP1A1 mRNA levels compared to controls. The differential induction of the two CYP1A genes was not as evident in vitro as in vivo. In addition, reanalysis and sequence comparison of the these two trout CYP1A genes with the first trout CYP1A cDNA described by Heilmann et al. (DNA 7, 379-387, 1988) indicate that the Heilmann cDNA is a hybrid of the two trout genes. The 5' portion of the cDNA sequence (212 bp) was determined by sequencing of a genomic clone and is 100% identical to the trout CYP1A3 gene. The majority of the cDNA sequence (2377 bp), however, was sequenced from a partial cDNA clone and is 99.2% identical to trout CYP1A1. Although the nomenclature of these two trout CYP1A genes is undergoing revision, these results demonstrate a differential, inducer-dependent response to model mammalian CYP1A inducers. PMID- 10860870 TI - Irreversible effects of neonatal exposure to p-tert-octylphenol on the reproductive tract in female rats. AB - It has been known for many years that administration of androgens or estrogens at critical periods of development in mammals causes severe long-term effects on the endocrine/genital systems. The environmental pollutant p-tert-octylphenol (OP) possesses a weak but clear estrogen agonist activity in in vitro and in vivo studies. In the present study, effects of neonatal exposure to OP on the reproductive tract of female rats were investigated. Newborn female pups were injected with 100 mg/kg OP subcutaneously within 24 h after birth. Administration was repeated every other day until postnatal day 15 (total of eight doses). Before weaning, serum follicle-stimulating hormone (FSH) and luteinizing hormone (LH) remained at low levels during OP exposure, although the serum FSH peak and the high LH level were obvious in the controls. Histologically, inhibition of uterine gland genesis was apparent. The day of vaginal opening was about 4 days earlier in OP-treated animals than in controls. Persistent estrus was consistently observed in OP-treated animals. Atrophic and polycystic ovaries without corpora lutea showed anovulation. In the endometrium, cell-proliferative activity and cell-death were increased and decreased, respectively, and expression of estrogen receptor alpha mRNA was apparent by in situ hybridization. Unexpectedly, endometrial hyperplasias appeared at 8 weeks of age. After ovariectomy, vaginal smears immediately became of castration type and the uterus was atrophied. These results suggested that neonatal exposure to a high dose of OP alters developmental hormonal secretion presumably due to a hypothalamo pituitary-ovarian disorder, with accelerated vaginal opening, subsequent persistent estrus, and uterine endometrial hyperplasia. The changes in the uterus and vagina are ovary-dependent. PMID- 10860871 TI - The aryl hydrocarbon receptor has a role in the in vivo maturation of murine bone marrow B lymphocytes and their response to 2,3,7,8-tetrachlorodibenzo-p-dioxin. AB - The ligand-activated aryl hydrocarbon receptor (AHR) is a cytosolic DNA binding protein. Although no biologic role for AHR has been elucidated, it mediates the immunotoxicity of xenobiotics such as 2, 3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and its targeted inactivation produces abnormal immune system development. While investigators have demonstrated AHR's involvement in TCDD induced B lymphocyte functional alterations, little is known about the receptor's possible role in early B cell maturation and whether exogenous ligands change this process. The purpose of this study was to determine, (1) whether bone marrow B lymphocyte maturation is affected by AHR presence, (2) if so, its relative importance in hematopoietic and/or nonhematopoietic elements and, (3) whether TCDD alters this process. Radiation chimeras were produced that were AHR positive (Ahr+/+) or negative (Ahr-/-) in either their nonhematopoietic or hematopoietic elements, or both. Marrow cells were analyzed for alterations in B lymphocyte maturation stage cell numbers in both vehicle- and TCDD-treated animals. Our results showed that (1) Ahr-/- animals had significantly higher numbers of pro/pre-B cells than Ahr+/+ animals, (2) TCDD treatment of Ahr+/+ animals produced a decrease in pro/pre-B cell numbers, whereas no effect was observed on Ahr-/- animals, and (3) AHR is required in both hematopoietic and stromal elements for maintenance of B cell subset maturation profiles. PMID- 10860872 TI - Chemical index for volume 165 PMID- 10860873 TI - Human xenograft models for virus infection. PMID- 10860874 TI - The application of transgenic and knockout mouse technology for the study of viral pathogenesis. PMID- 10860875 TI - Neural stem cells as tools for understanding retroviral neuropathogenesis. AB - The discovery within the past decade that neural stem cells (NSCs) from the developing and adult mammalian brain can be propagated, cloned, and genetically manipulated ex vivo for ultimate transfer back into the CNS has opened the door to a novel means for modifying the CNS environment for experimental and therapeutic purposes. While a great deal of interest has been focused on the properties and promise of this new technology, especially in regard to cellular replacement and gene therapy, this minireview will focus on the recent use of NSCs to study the neuropathogenesis of the murine oncornaviruses. In brief, the use of this NSC-based approach has provided a means for selective reconstitution within the brain, of specific retroviral life cycle events, in order to consider their contribution to the induction of neurodegeneration. Furthermore, by virtue of their ability to disseminate virus within the brain, NSCs have provided a reliable means for assessing the true neurovirulence potential of murine oncornaviruses by directly circumventing a restriction to virus entry into the CNS. Importantly, these experiments have demonstrated that the neurovirulence of oncornaviruses requires late virus life cycle events occurring specifically within microglia, the resident macrophages of the brain. This initial application of NSC biology to the analysis of oncornavirus-CNS interactions may serve as an example for how other questions in viral neuropathogenesis might be addressed in the future. PMID- 10860876 TI - Detection and characterization of functional T-cell epitopes on the structural proteins VP2, VP3, and VP4 of foot and mouth disease virus O1 campos. AB - Foot and mouth disease virus (FMDV) is the cause of a widespread infectious disease affecting cloven-hoofed animals. It is controlled by vaccination with immune-inactivated virus grown in tissue culture. However, peptide vaccines represent a safer alternative to the current virus-inactivated immunogens. Their design requires the identification and evaluation of the sequences recognized by T- and B-lymphocytes. Four structural proteins, VP1, VP2, VP3, and VP4, comprise the viral capsid of the FMDV, but only VP1 has been extensively studied regarding the existence of relevant T-cell epitopes. Here, we utilize a murine model to present a functional T-cell epitope mapping on the complete sequences of VP2, VP3, and VP4 of FMDV O1 Campos. We used two in vitro assays to describe 13 amino acid sequences, each one of them including at least one T-cell epitope. The in vivo T-cell helper function of these sequences was studied in an adoptive cell transfer assay in mice. Immunization experiments with a fusion peptide containing one of the sequences characterized were also done comparing the helper activity of this sequence with other T-cell epitopes included in the major immunogenic region of VP1. PMID- 10860877 TI - Entry of R5X4 and X4 human immunodeficiency virus type 1 strains is mediated by negatively charged and tyrosine residues in the amino-terminal domain and the second extracellular loop of CXCR4. AB - CXCR4 mediates the fusion and entry of X4 and R5X4 strains of human immunodeficiency virus type 1 (HIV-1). The residues involved in CXCR4 coreceptor function have not all yet been identified, but tyrosine and negatively charged residues in the amino-terminal domain of CCR5 were shown to be indispensable for gp120 binding and entry of R5 and R5X4 strains. We therefore evaluated the role of such residues in CXCR4 coreceptor function by replacing tyrosines (Y), aspartic acids (D), and glutamic acids (E) with alanines (A) and testing the ability of these mutants to mediate the entry of X4 and R5X4 HIV-1 isolates. Our results show that viral entry depends on YDE-rich clusters in both the amino terminus and the second extracellular loop of CXCR4. Different viral isolates vary in their dependence on residues in one or the other domain. The determinants of CXCR4 coreceptor function are, therefore, more diffuse and isolate-dependent than those of CCR5. PMID- 10860878 TI - Membrane fusion between retroviral particles: host-range extension and vaccine prospects. AB - We have analyzed if different populations of retroviral particles carrying the viral and cellular receptors of membrane viruses, respectively, are able to specifically fuse with each other. Using the glycoprotein of human immunodeficiency virus type 1 and its cellular receptor complex, we demonstrate that interviral membrane fusion can, indeed, occur and that the resultant fused viral structures are able to infect cells and transduce a marker gene. On the one hand, these results have relevance for the development of vaccine strategies based on fusion-induced conformational epitopes on the viral glycoprotein. However, in addition to this potential practical application, the results obtained (which were extended to include analyses with the vesicular stomatitis virus G protein and its cellular receptor) have far-reaching implications for in vivo situations in which simultaneous infections with different membrane viruses can occur. PMID- 10860879 TI - Absence of coreceptor switch with disease progression in human immunodeficiency virus infections in India. AB - The envelope glycoprotein of the human immunodeficiency virus (HIV) utilizes CD4 as a receptor and CCR5 and/or CXCR4 as coreceptor to gain entry into the cell. The CCR5-tropic viruses, observed early in infection, could be important in transmission and the CXCR4-tropic viruses, observed late, may play an important role in disease progression. Viruses from 40 HIV-positive, asymptomatic or symptomatic individuals in India were isolated. Of 40 isolates 39 used CCR5. Thirty-three isolates were subtype C, 3 isolates were subtype A, and 4 isolates were HIV-2. Only 1 HIV-2 isolate, from a symptomatic individual, was dualtropic. Therefore, a majority of isolates from India belonged to subtype C and all the isolates utilized CCR5 exclusively irrespective of HIV disease status. PMID- 10860880 TI - The movement protein-triggered in situ conversion of potato virus X virion RNA from a nontranslatable into a translatable form. AB - Plant virus-encoded movement protein(s) (MP), and for many viruses the coat protein (CP), is required to mediate viral spread between plant cells via plasmodesmata (PD). Most probably, the genomic RNA of potexviruses moves through PD as assembled virions and/or as ribonucleoprotein complexes containing the CP and 25-kDa MP. Here we report that encapsidated potato virus X (PVX) virion RNA, which is nontranslatable in a cell-free protein synthesizing system, can be converted into a fully translatable form after interaction of intact PVX particles with the PVX 25-kDa MP. The 25-kDa MP molecules bind selectively to only one extremity of the viral particle (that presumably contains the 5' end of the genomic RNA). The process of complex formation is ATP-independent; i.e., the ATPase activity of the 25-kDa MP is not involved in the binding of the MP to PVX virion. PMID- 10860881 TI - Iduronic acid-containing glycosaminoglycans on target cells are required for efficient respiratory syncytial virus infection. AB - Respiratory syncytial virus (RSV) is an important human respiratory pathogen, particularly in infants. Glycosaminoglycans (GAGs) have been implicated in the initiation of RSV infection of cultured cells, but it is not clear what type of GAGs and GAG components are involved, whether the important GAGs are on the virus or the cell, or what the magnitude is of their contribution to infection. We constructed and rescued a recombinant green fluorescent protein (GFP)-expressing RSV (rgRSV) and used this virus to develop a sensitive system to assess and quantify infection by flow cytometry. Evaluation of a panel of mutant Chinese hamster ovary cell lines that are genetically deficient in various aspects of GAG synthesis showed that infection was reduced up to 80% depending on the type of GAG deficiency. Enzymatic removal of heparan sulfate and/or chondroitin sulfate from the surface of HEp-2 cells also reduced infection, and the removal of both reduced infection even further. Blocking experiments in which RSV was preincubated with various soluble GAGs revealed the relative blocking order of: heparin > heparan sulfate > chondroitin sulfate B. Iduronic acid is a component common to these GAGs. GAGs that do not contain iduronic acid, namely, chondroitin sulfate A and C and hyaluronic acid, did not inhibit infection. A role for iduronic acid-containing GAGs in RSV infection was confirmed by the ability of basic fibroblast growth factor to block infection, because basic fibroblast growth factor binds to GAGs containing iduronic acid. Pretreatment of cells with protamine sulfate, which binds and blocks GAGs, also reduced infection. In these examples, infection was reduced by pretreatment of the virus with soluble GAGs, pretreatment of the cells with GAG-binding molecules, pretreatment of the cells with GAG-destroying enzymes or in cells genetically deficient in GAGs. These results establish that the GAGs involved in RSV infection are present on the cell rather than on the virus particle. Thus, the presence of cell surface GAGs containing iduronic acid, like heparan sulfate and chondroitin sulfate B, is required for efficient RSV infection in cell culture. PMID- 10860882 TI - Coxsackievirus infection of the pancreas: evaluation of receptor expression, pathogenesis, and immunopathology. AB - Coxsackievirus type B (CVB) infection of the pancreas induces a massive cellular infiltrate composed of natural killer cells, T cells, and macrophages and leads to the destruction of exocrine tissue. The physiological manifestations of pancreatic CVB infection are correlated with viral tropism; the virus infects acinar cells but spares the islets of Langerhans. Here we evaluate the mechanisms underlying pancreatic inflammation and destruction and identify the determinants of viral tropism. T-cell-mediated immunopathology has been invoked, along with direct virus-mediated cytopathicity, to explain certain aspects of CVB-induced pancreatic disease. However, we show here that in the pancreas, the extent of inflammation and tissue destruction appears unaltered in the absence of the cytolytic protein perforin; these findings exclude any requirement for perforin mediated lysis by natural killer cells or cytotoxic T cells in CVB3-induced pancreatic damage. Furthermore, perforin-mediated cytotoxic T-cell activity does not contribute to the control of CVB infection in this organ. In addition, we demonstrate that the recently identified coxsackie-adenovirus receptor is expressed at high levels in acinar cells but is barely detectable in islets, which is consistent with its being a major determinant of virus tropism and, therefore, of disease. However, further studies using various cell lines of pancreatic origin reveal secondary determinants of virus tropism. PMID- 10860883 TI - Sequence analysis and genome organisation of poinsettia mosaic virus (PnMV) reveal closer relationship to marafiviruses than to tymoviruses. AB - Sequence comparison and genome organisation of poinsettia mosaic virus (PnMV), a putative member of the tymoviruses, revealed a closer relationship to marafiviruses. The complete nucleotide sequence of PnMV was determined. The 6099 nt RNA genome encodes a putative 221-kDa polyprotein that lacks a stop codon between the replicase and the coat protein genes, as in most tymovirus RNAs. The genomic RNA has a poly(A) tail at its 3'-terminus in contrast to the tRNA-like structure found in the RNA of most tymoviruses, and no homology was observed to the conserved noncoding region of the tymoviral 3'-termini. The tymobox of PnMV, a 16-nt region of the subgenomic RNA (sgRNA) promoter shared by most tymoviruses, differs in 3 nt from the RNA sequence of tymoviruses but is identical to the sequence of marafiviruses. At least three sgRNAs were found in PnMV-infected Euphorbia pulcherrima and in isolated PnMV particles; one that is 650 nt long encodes the 21.4-kDa coat protein, and the others are about 3.5 and 1.7 kb and contain the 5'- and the 3'-terminal parts of genomic RNA, respectively. Like tymoviruses, PnMV particles sediment as top and bottom components. The particles of the top component contain the sgRNA (650 nt) encoding the coat protein, and those of bottom component contain both genomic and sgRNAs. PMID- 10860884 TI - Forced retroevolution of an RNA bacteriophage. AB - The operator hairpin ahead of the replicase gene in RNA bacteriophage MS2 contains overlapping signals for binding the coat protein and ribosomes. Coat protein binding inhibits further translation of the gene and forms the first step in capsid formation. The hairpin sequence was partially randomized to assess the importance of this structure element for the bacteriophage and to monitor alternative solutions that would evolve on the passaging of mutant phages. The evolutionary reconstruction of the operator failed in the majority of mutants. Instead, a poor imitation developed containing only some of the recognition signals for the coat protein. Three mutants were of particular interest in that they contained double nonsense codons in the lysis reading frame that runs through the operator hairpin. The simultaneous reversion of two stop codons into sense codons has a very low probability of occurring. Therefore the phage solved the problem by deleting the nonsense signals and, in fact, the complete operator, except for the initiation codon of the replicase gene. Several revertants were isolated with activities ranging from 1% to 20% of wild type. The operator, long thought to be a critical regulator, now appears to be a dispensable element. In addition, the results indicate how RNA viruses can be forced to step back to an attenuated form. PMID- 10860885 TI - Structure of transcripts and proteins encoded by U79-80 of human herpesvirus 6 and its subcellular localization in infected cells. AB - We analyzed the U79-80 gene of human herpesvirus 6 (HHV-6), which is predicted to be a positional homolog of the UL112-113 gene of human cytomegalovirus (HCMV). The U79-80 gene encoded a family of nuclear proteins of 36, 41, 44, and 59 kDa. These proteins had common amino termini and were generated by complex alternative splicing. Transcripts from U79-80 appeared as early as 3 h postinfection and could be detected in the presence of phosphonoformate. U79-80 proteins were seen as early as 8 h postinfection and could be detected in the presence of phosphonoformate but not in the presence of cycloheximide combined with actinomycin D treatment. The U79-80 proteins were localized to the nucleus of infected cells, where they were detected as a speckled or punctuate pattern. Moreover, the U79-80 proteins colocalized with the components of the viral DNA replication machinery and appeared to distribute adjacent to or touching nuclear domain 10, where viral DNA replication occurs. From the sequence analysis of genomic DNA, the predicted amino acid similarity between U79-80 and UL112-113 was lower than between other genes, but the characteristics of the transcripts and proteins encoded by U79-80 were similar to those of UL112-113. These results suggest that the U79-80 proteins have a role in viral DNA replication and are functional homologues of the UL112-113 proteins. PMID- 10860886 TI - Molecular architecture of bacteriophage T4 capsid: vertex structure and bimodal binding of the stabilizing accessory protein, Soc. AB - T4 encodes two dispensable proteins that bind to the outer surface of the mature capsid. Soc (9 kDa) stabilizes the capsid against extremes of alkaline pH and temperature, but Hoc (40 kDa) has no perceptible effect. Both proteins have been developed as display platforms. Their positions on the hexagonal surface lattice of gp23*, the major capsid protein, were previously defined by two-dimensional image averaging of negatively stained electron micrographs of elongated variant capsids. We have extended these observations by reconstructing cryo-electron micrographs of isometric capsids produced by a point mutant in gene 23, for both Hoc+.Soc+ and Hoc+.Soc- phages. The expected T = 13 lattice was observed, with a single Hoc molecule at the center of each gp23* hexamer. The vertices are occupied by pentamers of gp24*: despite limited sequence similarity with gp23*, the respective monomers are similar in size and shape, suggesting they may have the same fold. However, gp24* binds neither Hoc nor Soc; in situ, Soc is visualized as trimers at the trigonal points of the gp23* lattice and as monomers at the sites closest to the vertices. In solution, Soc is a folded protein ( approximately 10% alpha-helix and 50-60% beta sheet) that is monomeric as determined by analytic ultracentrifugation. Thus its trimerization on the capsid surface is imposed by a template of three symmetry-related binding sites. The observed mode of Soc binding suggests that it stabilizes the capsid by a clamping mechanism and offers a possible explanation for the phenotype of osmotic shock resistance. PMID- 10860887 TI - Molecular characterization of Nipah virus, a newly emergent paramyxovirus. AB - Recently, a new paramyxovirus, now known as Nipah virus (NV), emerged in Malaysia and Singapore, causing fatal encephalitis in humans and a respiratory syndrome in pigs. Initial studies had indicated that NV is antigenically and genetically related to Hendra virus (HV). We generated the sequences of the N, P/C/V, M, F, and G genes of NV and compared these sequences with those of HV and other members of the family Paramyxoviridae. The intergenic regions of NV were identical to those of HV, and the gene start and stop sequences of NV were nearly identical to those of HV. The open reading frames (ORFs) for the V and C proteins within the P gene were found in NV, but the ORF encoding a potential short basic protein found in the P gene of HV was not conserved in NV. The N, P, C, V, M, F, and G ORFs in NV have nucleotide homologies ranging from 88% to 70% and predicted amino acid homologies ranging from 92% to 67% in comparison with HV. The predicted fusion cleavage sequence of the F protein of NV had a single amino acid substitution (K to R) in comparison with HV. Phylogenetic analysis demonstrated that although HV and NV are closely related, they are clearly distinct from any of the established genera within the Paramyxoviridae and should be considered a new genus. PMID- 10860888 TI - Antigenic properties of recombinant envelope glycoproteins derived from T-cell line-adapted isolates or primary human immunodeficiency virus isolates and their relationship to immunogenicity. AB - The native envelope glycoproteins of primary HIV-1 virions have weaker antigenicity than do T-cell laboratory-adapted (TCLA) viruses. These antigenic properties require further evaluation if recombinant envelope glycoproteins are produced as part of a vaccine strategy. In this study, we compared the antigenicity of recombinant envelope glycoproteins derived from three primary isolates (PI) (HIV-1(BX08), HIV-1(CHA), and HIV-1(133)) and two TCLA viruses (HIV 1(HXB2) and HIV-1(MN)) produced using the Semliki Forest virus (SFV) system. This analysis was performed by radioimmunoprecipitation assays and flow cytometry. The results suggest that the SFV produces envelope glycoproteins with features in common with the envelopes found in naturally occurring virions. In particular, the PI envelopes had weak heterogeneous antigenic properties. However, the cytometric analysis also showed that there was less envelope glycoprotein on the cell surface for the PI envelopes than for those of TCLA viruses, suggesting differences in their intracellular trafficking. The immunogenic properties of the various envelope glycoproteins were evaluated in mice using recombinant SFV particles as vaccine vectors. The PI envelopes were less immunogenic than the TCLA envelopes, probably due to both their low antigenicity and cell surface expression level. Thus, it may be difficult to design an effective vaccine based on native recombinant PI envelopes. PMID- 10860889 TI - Internal and 3' RNA initiation by Qbeta replicase directed by CCA boxes. AB - RNA initiation by Qbeta replicase directed by the short-sequence CCA at the 3' end of all RNAs amplified by this enzyme has been studied. Most CCA repeats in an RNA consisting of 12 CCAs serve as independent sites of de novo RNA initiation, with initiation occurring opposite the 3'-C residue of each CCA. Qbeta replicase is thus capable of internal initiation remote from the 3'-end, although predominant initiation occurs close to the 3'-end. The precise 3'-terminal sequence in (CCA)(n)-containing RNAs influences the number and position of active initiation sites near the 3'-terminus. C residues are required at the initiation site, whereas the position of purines (especially A residues) influences the selection of initiation sites. The template activity of (CCA)(n) RNAs is positively correlated with the number of CCA repeats. Three CCA repeats added to the 3'-end of a nontemplate 83-nt RNA are sufficient to activate transcription by Qbeta replicase. These experiments show that CCA boxes can act as strong initiation sites in the absence of specific cis-acting signals derived from Qbeta RNA, although the efficiency of initiation is modulated by surrounding sequence. PMID- 10860890 TI - Genotype-specific synthesis and secretion of spliced hepatitis B virus genomes in hepatoma cells. AB - Hepatitis B virus-infected patients frequently have viral particles with DNA derived from differently spliced RNA. Which factors influence the synthesis of these splice variants is unclear. We analysed the type of splice variants produced from different genotypes and determined whether they are secreted as efficiently as wild-type virus. We demonstrate production of a single splice variant from genotypes D, C, and E as dominant species in two hepatoma cell lines. The type of minor splice variants synthesised varied between genotypes but was identical in both hepatoma cell lines. A novel splice variant with a deletion in the core gene was identified for genotype D. Viral DNA from intracellular compared with extracellular viral particles was spliced approximately five times more often than wild-type-sized genomes. A variable amount of the major splice variant was also identified in sera from patients infected with genotypes A, D, and C. These data indicate genotype A-, C-, D-, and E- as well as hepatoma cell line-independent synthesis of a dominant single splice variant and argue for a biological function of the corresponding splice sites. This study clearly demonstrates the intracellular accumulation of viral particles containing spliced genomes and offers a tool for the investigation of underlying mechanisms. PMID- 10860891 TI - Mixed viral infection identified using heteroduplex mobility analysis (HMA). AB - It is now recognised that mixed viral infection, or infection of an individual with two or more distinct strains of a single viral species, often occurs particularly with RNA viruses. Current methods for detection of mixed infection normally involve genotyping or cloning and DNA sequencing. These methods are not always accurate or sensitive at detecting mixed infection and cannot be used for large numbers of samples. Furthermore subsequent sequence determination of the coinfecting viruses is labour intensive. This paper describes a simple, generic method based upon PCR and heteroduplex mobility analysis (HMA) that can be used to rapidly determine mixed infection with two strains of the same virus. The utility of this method is illustrated with hepatitis C virus (HCV) and TT virus (TTV) as examples. PCR-HMA detected mixed infection in 3 (8%) of 38 sera from intravenous drug users (IVDU) and 28 (30%) of 70 TTV-positive sera from Australia, China, and Vietnam. HMA can also be used to screen recombinant colonies to identify the sequences of the coinfecting viruses. The methods described here could be applied to analyse any PCR product containing two or more divergent sequences, whether derived from viruses, bacteria, or eukaryotic organisms. PMID- 10860892 TI - Role of type I IFNs in the in vitro attenuation of live, temperature-sensitive vaccine strains of human respiratory syncytial virus. AB - The contributions of type I interferons (IFNs) to the in vitro attenuation of three temperature-sensitive (Ts) subgroup A and one subgroup B deletion mutant RSV strains were evaluated. The ability of these vaccine viruses to induce IFNs at their permissive and restrictive temperatures and their sensitivity to the antiviral effects of exogenous I IFNs were tested in human lung epithelial A549 cells. Our results show that the highly attenuated and immunogenic subgroup A vaccine strain Ts1C produced higher levels of IFN-beta than its parent RSS-2 or two related strains, Ts1A and Ts1B, at their permissive temperature. Growth of RSV-infected A549 cultures at restrictive temperatures or prior UV inactivation of the virus abolished the observed induction of IFN-beta, suggesting a strict requirement of viral replication for cellular IFN induction. The enhanced induction of IFN-beta by the highly immunogenic Ts1C at permissive temperature may be an advantageous characteristic of a live intranasal vaccine candidate. The subgroup B strain RSV B1 and its mutant cp-52 (with SH and G gene deletions) both induced similar but low levels of IFN-beta. Hence the observed overattenuation of cp-52 in human infants is probably not due to enhanced IFN induction during its replication in the host. The ability of cp-52, which does not express the SH and G proteins, to induce IFN-beta levels similar to those of its parent strain suggests that these viral proteins may not have a role in the induction of IFN beta in the host. In addition, both subgroup A and B mutants and their respective parent strains were similarly resistant to the antiviral effects of exogenous IFN alpha or -beta. Therefore, increased sensitivity of the mutants to IFNs does not seem to contribute to their attenuation. PMID- 10860893 TI - Introduction PMID- 10860894 TI - Management of the adverse effects of antiretroviral therapy and medication adherence. AB - A commonly cited cause of poor adherence to highly active antiretroviral therapy (HAART) is adverse drug reactions. Short-term adverse effects are potential threats to successful introduction and maintenance of HAART. The long-term toxicities of HAART are still emerging and being defined, as evidenced by the recently described metabolic disorders (i.e., the syndrome of maldistribution, hyperlipemia, glucose intolerance and insulin resistance). With 14 licensed agents in 2000, other agents in common use, and numerous combinations of >/=3 drugs, awareness and recognition of adverse effects are increasingly important for clinicians and patients. The common adverse drug reactions encountered with HAART, including new agents and their impact on patient adherence, are reviewed. Current strategies to anticipate and mitigate adverse effects are summarized. PMID- 10860895 TI - Clinical utility of testing human immunodeficiency virus for drug resistance. AB - Human immunodeficiency virus (HIV) type 1 drug-resistance testing is quickly moving from the research laboratory to the clinic as data defining its utility as a prognostic indicator of response to therapy become available. In July 1998, a panel of the International AIDS Society-USA did not recommend the widespread application of resistance testing, but by May 2000 this panel endorsed and recommended the incorporation of resistance testing in patient-care management. Considerable data supporting the use of drug-resistance testing have now been published or presented at international conferences. These data strongly suggest that drug-resistance testing is of considerable value in many clinical settings. Prospective trials of resistance testing as a clinical management tool are still ongoing, and the long-term benefits still need to be evaluated. Nevertheless, early results from several studies showed a significantly better virological response when treatment regimens were based on resistance-testing data, rather than on the standard of care. HIV drug-resistance testing is also useful as a tool for new antiretroviral drug design and development, as well as for monitoring the spread of primary HIV drug resistance. PMID- 10860896 TI - Using pharmacokinetics to optimize antiretroviral drug-drug interactions in the treatment of human immunodeficiency virus infection. AB - Better understanding of the pharmacokinetics of antiretroviral drugs has resulted in the design of combination therapies for the treatment of human immunodeficiency virus (HIV) infection. This has improved the bioavailability and prolonged the plasma half-life of some of the drugs, resulting in enhanced antiviral activity. However, antiviral combination therapy can also result in adverse drug-drug interactions and diminished antiretroviral activity. In this review, we examine drug interactions involving combinations of protease inhibitors, combinations of protease inhibitors with nonnucleoside reverse transcriptase inhibitors, and combinations of nucleoside analogues for the treatment of patients with HIV infection. We discuss examples and mechanisms of pharmacokinetic interactions that improve or decrease antiviral efficacy. PMID- 10860897 TI - Initial therapy with protease inhibitor-sparing regimens: evaluation of nevirapine and delavirdine. AB - We have compared the results (on-treatment analyses) of 2 randomized clinical trials of protease inhibitor-sparing regimens in drug-naive patients. In the INCAS (Italy, Netherlands, Canada, Australia) study, the mean decrease in plasma viral load over 52 weeks was 2.2 log(10) copies/mL in 40 patients who were receiving zidovudine/didanosine/nevirapine (18 [45%] had maximal suppression), with a mean increase in CD4 T cell counts of 139 cells/microL. In protocol 0021 Part II, the mean decrease in plasma viral load over 52 weeks was 2.1 log(10) copies/mL in 34 patients who were receiving zidovudine/lamivudine/delavirdine (20 [59%] had maximal suppression), with a mean increase in CD4 T cell counts of 88 cells/microL. The virologic and immunologic efficacy of the 2 triple-drug regimens are similar. Until results of long-term studies are available to establish whether a preferred approach to initial therapy exists, nonnucleoside reverse transcriptase inhibitors may be a valuable alternative to protease inhibitors in the initial therapy of antiretroviral-naive, moderately immunosuppressed patients. PMID- 10860898 TI - HIV protease inhibitor-related lipodystrophy syndrome. AB - Human immunodeficiency virus (HIV) protease inhibitor (PI) therapy is frequently associated with a syndrome increasingly referred to as lipodystrophy syndrome, which is characterized by peripheral lipoatrophy, fat accumulation within the abdomen, in the breasts of women, and over the cervical vertebrae ("buffalo hump"), hyperlipidemia, and insulin resistance. In the largest study to date, peripheral lipoatrophy (an estimated 0.35-kg fat loss per month overall from the face, limbs, and upper trunk) was observed in association with all licensed PIs after a median 10 months of PI therapy. Diabetes mellitus type II appears to be a related, but less common, adverse effect. The lipodystrophy syndrome may be a result of the inhibition of 2 proteins involved in lipid metabolism that have significant homology to the catalytic site of HIV protease-namely, cytoplasmic retinoic acid binding protein type 1 and low density lipoprotein-receptor-related protein. PMID- 10860899 TI - Role of hydroxyurea in treatment of disease due to human immunodeficiency virus infection. AB - The potential role of hydroxyurea in the treatment of human immunodeficiency virus (HIV) infection was first supported by in vitro experiments that demonstrated control of viral production in activated and resting T cells. More recently, controlled clinical trials demonstrated that the addition of hydroxyurea to nucleoside-including regimens (chiefly of didanosine but also of stavudine and lamivudine) enhances their antiviral potency. It is believed that the cytostatic effect of hydroxyurea is at least partially responsible for its antiviral effect, through the down-modulation of cellular proliferation. Such an effect has also been credited for the blunted CD4 T cell responses that are characteristically observed when hydroxyurea is added to nucleoside-including regimens. The adjunctive antiviral effect of hydroxyurea-as well as its favorable dosing schedule, safety profile, and cost-makes it a very attractive addition to our therapeutic armamentarium. Further research is urgently needed to delineate the most appropriate use of this compound in the setting of HIV antiretroviral therapy. PMID- 10860900 TI - Pharmacodynamics of human immunodeficiency virus type 1 protease inhibitors. AB - Many factors are involved in the success or failure of antiretroviral therapy. Recent data suggest that there are significant differences in drug absorption and disposition for the protease inhibitor class of antiretroviral drugs, and relationships between plasma concentrations and their antiviral effect have been described. Consequently, the issue of whether therapeutic drug monitoring should be employed for patients receiving treatment with these drugs has arisen. Several criteria must be met before a drug is considered a candidate for therapeutic drug monitoring. These criteria include pharmacological, clinical, and analytic components. Although not all the necessary criteria have yet been met, some of these components have been defined, and additional data are being generated. However, prospectively designed clinical trials must be completed to determine if monitoring protease inhibitor plasma concentrations provides additional clinical benefit to the patient. PMID- 10860901 TI - HIV-1 protease inhibitors. AB - Treatment of human immunodeficiency virus type 1 (HIV-1) infection with regimens that include protease inhibitors (PIs) has contributed to marked improvements in HIV-related disease progression and mortality. Five PIs are approved by the US Food and Drug Administration and have potent activity in vitro. PIs with 2 nucleoside analogue reverse transcriptase inhibitors have demonstrated prolonged suppression of HIV-1 replication in treated patients and improvements in disease progression and mortality. PIs combined with nonnucleoside reverse transcriptase inhibitors or other PIs produce marked antiretroviral effects. Although not all patients have prolonged responses to PIs, and salvage treatment has had mixed results for patients who have not responded to initial PI therapy or whose HIV RNA levels have relapsed during such therapy, newer PIs currently being developed hold promise. Most patients can successfully tolerate PI-including regimens; however, long-term side effects, such as body fat redistribution, insulin resistance, and increased serum lipids, are now being observed in some patients receiving PI-including therapy. PMID- 10860902 TI - Factors affecting adherence to antiretroviral therapy. AB - In both clinical trials and clinical practice, nonadherence to medications is widespread among patients with chronic diseases. The shift to combination therapies for treating human immunodeficiency virus (HIV)-infected individuals has increased adherence challenges for both patients and health-care providers. Estimates of average rates of nonadherence to antiretroviral therapy range from 50% to 70%. Adherence rates of <80% are associated with detectable viremia in a majority of patients. The principal factors associated with nonadherence appear to be patient-related, including substance and alcohol abuse. However, other factors may also contribute, such as inconvenient dosing frequency, dietary restrictions, pill burden, and side effects; patient-health-care provider relationships; and the system of care. We discuss the major reasons reported by HIV-infected individuals for not taking their medications. Improving adherence probably requires clarifying the treatment regimen and tailoring it to patient lifestyles. PMID- 10860903 TI - Determinants of virological response to antiretroviral therapy: implications for long-term strategies. AB - A variety of factors can contribute to the failure of combination antiretroviral therapy to durably suppress viral replication in patients infected with human immunodeficiency virus (HIV). Patients who have a low CD4(+) T cell count or high plasma viral load before therapy is initiated are at high risk for subsequent virological failure. Previous therapy is also a strong determinant of subsequent virological response, presumably because of pre-existing viral resistance. Drug exposure, as determined by adherence, drug absorption, and drug metabolism, has a significant impact on future long-term virological responses. Although definitive proof is lacking, some tissues may have limited drug penetration, thus allowing for ongoing viral replication. Understanding why combination therapy fails for HIV-infected patients may allow clinicians to individualize treatment strategies. Unfortunately, almost any factor (drug, host, or viral) that leads to virological failure of an initial combination regimen is likely to persist-and perhaps become more challenging-once a salvage regimen is initiated. PMID- 10860904 TI - Resistance against reverse transcriptase inhibitors. AB - The response to antiretroviral therapy in human immunodeficiency virus (HIV) infected patients is limited by the emergence of drug resistance. This resistance is a consequence of the high rate of HIV mutation, the high rate of viral replication (especially when potent multidrug therapies are not used or taken reliably), and the selective effect of these drugs, which favors emergence of mutations that can establish clinical drug resistance. The introduction of highly active antiretroviral therapy (HAART), which typically includes at least 2 nucleoside reverse transcriptase inhibitors (RTIs) and a protease inhibitor or a nonnucleoside RTI, for most treatment-naive patients results in a reduction of viral load below the limit of detection determined by currently available HIV RNA assays. It is this marked reduction that results in durable viral suppression, usually only possible by the simultaneous use of 3 or 4 drugs. The RTI components of HAART are crucial for these benefits of combination therapy. Specific amino acid changes are associated with resistance to several RTIs, but new mutation complexes have been observed that can confer broad cross-resistance within this class. Genotypic and phenotypic resistance assays to measure drug resistance are being developed, but refinements in both methodology and our ability to interpret results of these assays are necessary before they are introduced into widespread clinical use. PMID- 10860905 TI - Rationale for the use of hydroxyurea as an anti-human immunodeficiency virus drug. AB - Hydroxyurea has been extensively used in medical practice, mainly for treating chronic myelogenous leukemia, sickle cell anemia, and other diseases. In light of its ability to inhibit DNA synthesis and to induce cell cycle arrest through inhibition of ribonucleotide reductase, the effects of hydroxyurea on replication of human immunodeficiency virus type 1 (HIV-1) have been investigated. In vitro hydroxyurea has been shown to block HIV-1 reverse transcription and/or replication in quiescent peripheral blood mononuclear cells (PBMC) and macrophages. Hydroxyurea was also found to be synergistic with the nucleoside reverse transcriptase inhibitor didanosine and to inhibit HIV-1 replication in activated PBMC; this inhibition may be due to a reduction in deoxynucleoside triphosphate pool sizes. Finally, hydroxyurea has been shown to sensitize didanosine-resistant mutants. Hydroxyurea may therefore be useful for limiting the spread of didanosine-resistant HIV-1 variants. The favorable toxicity profile of hydroxyurea and the lack of significant overlapping toxicities with some of the nucleoside reverse transcriptase inhibitors, as well as their distinct mechanisms of action, have provided further rationale for use of these agents in combination therapies. PMID- 10860906 TI - New perspectives on the Mesozoic seed fern order Corystospermales based on attached organs from the Triassic of Antarctica. AB - A new Triassic corystosperm is described from the Shackleton Glacier region of Antarctica. The compression fossils include cupulate organs (Umkomasia uniramia) and leaves (Dicroidium odontopteroides) attached to short shoot-bearing branches. The cupulate organs occur in groups near the apices of the short shoots, and each consists of a single axis with a pair of bracts and a subapical whorl of five to eight ovoid cupules. This unique architecture indicates that the cupules are individual megasporophylls rather than leaflets of a compound megasporophyll. A branch bearing an attached D. odontopteroides leaf provides the first unequivocal evidence that Umkomasia cupulate organs and Dicroidium leaves were produced by the same plants. Although this had previously been assumed based on organ associations, the new specimens are important in demonstrating that a single species of corystosperm produced the unique cupulate organs described here and the geographically and stratigraphically widespread and common D. odontopteroides leaf. Therefore, biostratigraphic, paleoecological, and phylogenetic studies that treat Dicroidium leaf morphospecies as proxies for biological species of entire plants should be reconsidered. Phylogenetic analysis suggests that the corystosperm cupule is an unlikely homologue for the angiosperm carpel or outer integument. PMID- 10860907 TI - Effects of atmospheric CO2 enrichment on the growth and development of Hymenocallis littoralis (Amaryllidaceae) and the concentrations of several antineoplastic and antiviral constituents of its bulbs. AB - Two 2-yr crops of tropical spider lily (Hymenocallis littoralis) plants were grown in field soil in clear-plastic-wall open-top enclosures in the Sonoran Desert environment of central Arizona. Half of the plants were exposed to ambient air of 400 ppm atmospheric CO(2) concentration and half of them were exposed to air of 700 ppm CO(2). This 75% increase in the air's CO(2) content resulted in a 48% increase in aboveground plant biomass and a 56% increase in belowground (bulb) biomass. It also increased the concentrations of five bulb constituents that have been demonstrated to possess anticancer and antiviral activities. Mean percentage increases in these concentrations were 6% for a two-constituent (1:1) mixture of 7-deoxynarciclasine and 7-deoxy-trans-dihydronarciclasine, 8% for pancratistatin, 8% for trans-dihydronarciclasine, and 28% for narciclasine, for a mean active ingredient percentage concentration increase of 12%. Combined with the 56% increase in bulb biomass, these percentage concentration increases resulted in a mean active ingredient increase of 75% for the 75% increase in the air's CO(2) concentration used in our experiments. PMID- 10860908 TI - Genetic discontinuity revealed by chloroplast microsatellites in eastern North American Abies (Pinaceae). AB - Development of conservation strategies for Fraser fir (Abies fraseri) in the southern Appalachian Mountains depends in part on recognition of the extent to which Fraser fir is genetically distinct from the closely related balsam (A. balsamea) and intermediate (A. balsamea var. phanerolepis) fir. These sibling species have exhibited intergrading, clinal variation in morphological, chemical, and genetic characteristics in prior research. Chloroplast microsatellite markers were polymerase chain reaction amplified from genomic DNA samples of 78 individuals representing the geographic ranges of Fraser, balsam, and intermediate fir. Gene diversity levels at two loci ranged among taxa from 0.65 to 0.84. Allele frequencies demonstrated significant differentiation among taxa, with R(ST) values of 0.36 and 0.10. Haplotype diversity and D(SH) were highest for balsam fir and lowest for intermediate fir. A haplotype network analysis based on allele size distribution for the two loci revealed two distinct clusters of haplotypes and population-specific haplotypes. Ninety-two percent of the haplotypes in one cluster were from balsam fir and intermediate fir, and 84% of the haplotypes in the other cluster were from Fraser fir and intermediate fir. The genetic differentiation of chloroplast DNA markers provides justification for the recognition of Fraser fir as a distinct Management Unit (MU) for conservation purposes, regardless of its taxonomic classification. PMID- 10860909 TI - Patterns of genetic variation in rare and widespread plant congeners. AB - Rare species are typically considered to maintain low levels of genetic variation, and this view has been supported by several reviews of large numbers of isozyme studies. Although these reviews have provided valuable data on levels of variability in plant species in general, and rare species in particular, these broad overviews involve comparisons that may confound the effects of rarity with a multitude of other factors that affect genetic variability. Additionally, the statistical analyses employed assume the data to be independent, which is not the case for organisms that share a common phylogenetic history. As the role of evolutionary history and historical constraints has become better understood, more researchers have studied widespread congeners when investigating the genetic diversity of rare species in an effort to control for these effects. We summarize the available data from such studies, comparing for rare and widespread congeners (1) the levels of genetic variability at the population and species levels and (2) measures of population substructuring. At the population level, we summarized data for percentage polymorphic loci (%P(pop)), mean number of alleles per locus (A(pop)), and observed heterozygosity (H(o)). Species-level measures used were percentage polymorphic loci (%P(spp)), mean number of alleles per locus (A(spp)), and total genetic diversity (H(T)). Indices of population subdivision (either F(ST) or G(ST)) were also examined. Using Wilcoxon signed rank tests, we found significant, but small, differences between rare and widespread species for all diversity measures except H(T). However, there does not appear to be a difference between rare and widespread congeners in terms of how genetic variation is partitioned within and among populations. Levels of diversity, for all measures examined, between rare and widespread congeners are highly correlated. PMID- 10860910 TI - Intersectional gene flow between insular endemics of Ilex (Aquifoliaceae) on the Bonin Islands and the Ryukyu Islands. AB - Hybridization and introgression play important roles in plant evolution, and their occurrence on the oceanic islands provides good examples of plant speciation and diversification. Restriction fragment length polymorphisms (RFLPs) and trnL (UAA) 3'exon-trnF (GAA) intergenic spacer (IGS) sequences of chloroplast DNA (cpDNA), and the sequences of internal transcribed spacer (ITS) of nuclear ribosomal DNA were examined to investigate the occurrence of gene transfer in Ilex species on the Bonin Islands and the Ryukyu Islands in Japan. A gene phylogeny for the plastid genome is in agreement with the morphologically based taxonomy, whereas the nuclear genome phylogeny clusters putatively unrelated endemics both on the Bonin and the Ryukyu Islands. Intersectional hybridization and nuclear gene flow were independently observed in insular endemics of Ilex on both sets of islands without evidence of plastid introgression. Gene flow observed in these island systems can be explained by ecological features of insular endemics, i.e., limits of distribution range or sympatric distribution in a small land area. PMID- 10860911 TI - Production and survivorship of the functional stolons of giant cutgrass, Zizaniopsis miliacea (Poaceae). AB - Giant cutgrass [Zizaniopsis miliacea], a tall emergent grass native to the southeastern United States, was studied in two Florida lakes. In Lake Seminole (15 176 ha) giant cutgrass forms large expanding stands, but in Lake Alice (9 ha) it is confined to a stable narrow fringe. By monitoring individual plants in Lake Seminole, it was found that an average decumbent flowering stem produced three flowers and ten nodes, 80% of which became rooted in the substrate. Such flowering stem development could potentially result in stand expansion of 2.2-2.7 m/yr, depending upon water levels and rates of node rooting. Once flowering stems became decumbent in Lake Alice, they typically broke, producing no more than two flowers with four nodes in a growing season. While still attached to the parent plant, few of these nodes were able to become rooted in the substrate, limiting the rate of stand expansion in Lake Alice. Sections of flowering stems bearing axillary shoots that were detached from the parent plant and free-floating could become rooted on reaching shallow water and produce robust, new, flowering plants. This interesting mode of population dispersal and spread has important implications for the distribution and management of giant cutgrass. PMID- 10860912 TI - Absence of conspecific pollen advantage in the dynamics of an Ipomopsis (Polemoniaceae) hybrid zone. AB - The frequency of hybrid formation in angiosperms depends on how often heterospecific pollen is transferred to the stigma and on the success of that heterospecific pollen at fertilizing ovules. Even if heterospecific pollen is capable of effecting fertilization it may perform poorly when conspecific pollen is also available on the stigma. We applied pollen mixtures to stigmas to determine how pollen interactions affect siring success and the frequency of hybrid formation between two species of Ipomopsis (Polemoniaceae) in Colorado. Plants of both parental species and natural hybrids were pollinated with I. aggregata and I. tenuituba pollen in ratios of 100:0, 80:20, 50:50, 20:80, and 0:100 by mass. Plants were homozygous for different alleles at an isozyme marker, allowing us to distinguish the type of pollen parent for 2166 viable seeds from 273 fruits. In contrast to studies of many other hybridizing taxa, there was no evidence of an advantage to conspecific pollen, nor did composition of the stigmatic pollen load affect seed set. Instead, the frequency of seeds sired by a given species was proportional to its representation in the pollen load. In this hybrid zone, both the frequency of first-generation hybrid formation and the relative male fitness of the two parental species should be predictable from the rates of pollen transfer to stigmas. PMID- 10860913 TI - Pollination biology of two chiropterophilous agaves in Arizona. AB - I studied the pollination biology of two closely related species of agave, Agave palmeri and A. chrysantha (Agavaceae), which exhibit several chiropterophilous (bat-pollinated) traits. Floral studies, floral visitor observations, and pollination studies were conducted over four summers at six different sites to examine floral traits and determine the relative importance of diurnal vs. nocturnal pollinators. Agave chrysantha appears to have developed minor shifts in several floral characters that enhance diurnal pollination. Although floral shifts towards diurnal pollination were fewer in A. palmeri, stigmas were diurnally receptive and copious floral rewards were available in the morning, indicating that some adaptations exist to allow for multiple pollinators. Differences in fruit and seed set between naturally day- and night-pollinated umbels for both species were either not significant or significantly higher in day-pollinated plants. Bats were not important pollinators of A. chrysantha, and the mutualistic relationship between A. palmeri and the lesser long-nosed bat was found to be asymmetric. "Bat-adapted" floral traits appear to be flexible enough to respond to the climatic and pollinator unpredictability experienced by agaves at the northern edge of their distribution. This variability may be a more important factor affecting evolution of floral characters than a particular pollinator. PMID- 10860914 TI - Correlation between male and female reproduction in the subdioecious herb Astilbe biternata (Saxifragaceae). AB - Genotypic trade-offs between male and female reproduction are commonly assumed in theoretical studies of the evolution of gender specialization. Although these trade-offs are supported by higher seed production of females than hermaphrodites in natural populations of gynodioecious species, comparisons between male and female reproductive allocation among hermaphrodite individuals under controlled conditions are rare. We assessed phenotypic and genotypic correlations between stamen and fruit production in fruiting males of the near-dioecious herb Astilbe biternata. In the field, we found a significant negative phenotypic correlation between stamen production and fruit production within individuals that produced both stamens and fruit as well as higher fruit set in females than fruiting males. The negative correlation between fruit and stamen production that was observed in the field was also apparent across clonally propagated genotypes. These results suggest that negative genetic correlations between male and female reproduction may limit the independent evolution of fruit and stamen production in A. biternata. PMID- 10860915 TI - Pollen quality limits seed set in Burchardia umbellata (Colchicaceae). AB - In self-incompatible plants, interference by self pollen or genetically related pollen can potentially exacerbate pollen limitation, although this has rarely been demonstrated. We examined the breeding system, pollen limitation, and pollen interference using self- and cross- pollinations and pollen supplementations in Burchardia umbellata, an insect-pollinated lilioid monocot. Ovule fertilization and seed set were less following selfing than crossing (22 vs. 78% and 4 vs. 73%, respectively), indicating partial self-incompatibility. Flowers were partially protandrous, and flowers opened concurrently on plants potentially allowing self pollen interference. Natural seed set was pollen limited and varied within and among years, probably due to variation in flowering plant density. Interference by self or genetically related pollen caused pollen limitation as evidenced by increased seed set of bagged cross-pollinated plants compared to unbagged pollen supplemented plants in two years. In 1996, both fertilization and seed set increased in response to cross-pollination, indicating that interference occurred in the style and ovary. In 1997, only seed set increased after cross-pollination indicating that interference occurred in the ovary. Inappropriate pollen deposition may contribute to pollen limitation more often than previously recognized and should select for floral traits that decrease deposition of self or related pollen. PMID- 10860916 TI - Allelochemical autotoxicity in the emergent wetland macrophyte Juncus effusus (Juncaceae). AB - Bioassays for allelochemical toxicity of aboveground Juncus effusus tissues were conducted with seeds and seedlings of Eleocharis obtusa and Scirpus cyperinus, two emergent sedge species (Cyperaceae) found sympatric with J. effusus, and with seeds and seedlings of J. effusus itself to evaluate potential autotoxicity. Bioassays were performed under controlled, axenic conditions with aqueous shoot extract treatments simulating in situ dissolved organic carbon concentrations. With respect to the two sedge species, neither shoot development nor seedling biomass accrual was significantly suppressed by lyophilized whole extracts from J. effusus. Although the extracts induced no significant reduction in growth of E. obtusa or S. cyperinus, biomass-specific chlorophyll a concentration was significantly reduced in E. obtusa seedlings. In contrast, seedlings of J. effusus exhibited significant reductions of biomass and chlorophyll a concentrations, and seedling shoot development was retarded in response to leachate exposure. Results of the present study suggest that J. effusus seedlings possess autotoxic sensitivity to extracts of dead, aboveground tissues of adult plants. PMID- 10860917 TI - Factors affecting establishment of a gypsophyte: the case of Lepidium subulatum (Brassicaceae). AB - The restriction of vascular plants to gypsum-rich soils under arid or semiarid climates has been reported by many authors in different parts of the world. However, factors controlling the presence of gypsophytes on these soils are far from understood. We investigated the establishment of Lepidium subulatum, a gypsophyte, in a nondisturbed semiarid gypsum-soil landscape in central Spain, both from spatial and temporal perspectives. Over 1400 seedlings were tagged, and their growth and survival were monitored for a 2-yr period. Several biotic and abiotic variables were measured to determine the factors controlling the emergence and early survival. These variables included the cover of annual plants, bryophytes, lichens, litter, gypsum crystals, bare fraction and cover of each perennial plant, and several soil properties (gravel, fine gravel, and fine earth fraction, conductivity, pH, gypsum content, organic matter and penetrometer soil resistance). Our results support the linkage of gypsophily with some physical properties of the surface crust. Seedlings tended to establish on the gypsum surface crust, and their survival was size dependent, probably as a consequence of the necessity of rooting below the surface crust before summer drought arrives. However, once seedlings emerged, a higher survival rate occurred on the alluvial soils of the piedmont-slope boundary where soil crusts are absent or thinner. We conclude that Lepidium subulatum may be considered a refuge model endemic with a distribution range that occupies a reduced fraction of a wider habitat from which it is probably excluded by competition. PMID- 10860918 TI - Phylogeny and biogeography of Juglans (Juglandaceae) based on matK and ITS sequence data. AB - We investigated phylogenetic and biogeographic relationships within Juglans (walnuts), a Tertiary disjunct genus, using 15 species of Juglans and related (Juglandaceae) outgroups. The relationships were analyzed using nucleotide sequences of the chloroplast gene matK and its flanking spacers and of the internal transcribed spacers (ITS) and 5.8S gene of the nuclear ribosomal DNA. The DNA sequences provided 246 informative characters for parsimony analysis. ITS data supported as monophyletic groups the four generic sections, Cardiocaryon, Dioscaryon, Rhysocaryon, and Trachycaryon. Within Rhysocaryon, the temperate black walnuts and the tropical black walnuts were supported as monophyletic groups. When the two data sets were combined, J. cinerea was nested within Cardiocaryon. Combined analysis with published nuclear DNA restriction site data placed J. cinerea in a monophyletic group with Cardiocaryon. These analyses consistently supported Juglans as a monophyletic group and as the sister group to the genus Pterocarya. The results of this work are consistent with the known geological history of Juglans. The fossil record suggests that the butternuts had evolved by the early Oligocene in North America. The presence of butternuts in Eurasia could be the result of migration from North America to Eurasia during the warming trend of the mid Oligocene. PMID- 10860919 TI - Phylogenetic resolution within the tribe Episcieae (Gesneriaceae): congruence of ITS and NDHF sequences from parsimony and maximum-likelihood analyses. AB - Generic relationships within Episcieae were assessed using ITS and ndhF sequences. Previous analyses of this tribe have focussed only on ndhF data and have excluded two genera, Rhoogeton and Oerstedina, which are included in this analysis. Data were analyzed using both parsimony and maximum-likelihood methods. Results from partition homogeneity tests imply that the two data sets are significantly incongruent, but when Rhoogeton is removed from the analysis, the data sets are not significantly different. The combined data sets reveal greater strength of relationships within the tribe with the exception of the position of Rhoogeton. Poorly or unresolved relationships based exclusively on ndhF data are more fully resolved with ITS data. These resolved clades include the monophyly of the genera Columnea and Paradrymonia and the sister-group relationship of Nematanthus and Codonanthe. A closer affinity between Neomortonia nummularia and N. rosea than has previously been seen is apparent from these data, although these two species are not monophyletic in any tree. Lastly, Capanea appears to be a member of Gloxinieae, although C. grandiflora remains within Episcieae. Evolution of fruit type, epiphytic habit, and presence of tubers is re-examined with the new data presented here. PMID- 10860920 TI - Pollination by flies, bees, and beetles of Nuphar ozarkana and N. advena (Nymphaeaceae). AB - Nuphar comprises 13 species of aquatic perennials distributed in the temperate Northern Hemisphere. The European species N. lutea and N. pumila in Norway, the Netherlands, and Germany are pollinated by bees and flies, including apparent Nuphar specialists. This contrasts with reports of predominant beetle pollination in American N. advena and N. polysepala. We studied pollination in N. ozarkana in Missouri and N. advena in Texas to assess whether (1) there is evidence of pollinator shifts associated with floral-morphological differences between Old World and New World species as hypothesized by Padgett, Les, and Crow (American Journal of Botany 86: 1316-1324. 1999) and (2) whether beetle pollination characterizes American species of Nuphar. Ninety-seven and 67% of flower visits in the two species were by sweat bees, especially Lasioglossum (Evylaeus) nelumbonis. Syrphid fly species visiting both species were Paragus sp., Chalcosyrphus metallicus, and Toxomerus geminatus. The long-horned leaf beetle Donacia piscatrix was common on leaves and stems of N. ozarkana but rarely visited flowers. Fifteen percent of visits to N. advena flowers were by D. piscatrix and D. texana. The beetles' role as pollinators was investigated experimentally by placing floating mesh cages that excluded flies and bees over N. advena buds about to open and adding beetles. Beetles visited 40% of the flowers in cages, and flowers that received visits had 69% seed set, likely due to beetle-mediated geitonogamy of 1st-d flowers. Experimentally outcrossed 1st-d flowers had 62% seed set, and open-pollinated flowers 76%; 2nd-d selfed or outcrossed flowers had low seed sets (9 and 12%, respectively). Flowers are strongly protogynous and do not self spontaneously. Flowers shielded from pollinators set no seeds. A comparison of pollinator spectra in the two Old World and three New World Nuphar species studied so far suggests that the relative contribution of flies, bees, and beetles to pollen transfer in any one population depends more on these insects' relative abundances (and in the case of Donacia, presence) and alternative food sources than on stamen length differences between Old World and New World pond-lilies. PMID- 10860921 TI - Regulation of the MDR1 gene by transcriptional repressors selected using peptide combinatorial libraries. AB - The ability to selectively regulate the expression of genes implicated in cancer or other diseases could have important ramifications for both basic research and for therapy. Using peptide combinatorial libraries expressed in yeast, we have screened for novel zinc finger proteins that selectively bind to an overlapping EGR1/SP1/WT1 regulatory site in the promoter of the MDR1 multidrug resistance gene. The novel proteins were only moderately effective in blocking transcription by simple masking of the target site. However, when coupled to mammalian transactivator or repressor domains, they could selectively modulate the expression of reporter genes having promoters containing the MDR1 target site. Moreover, they could also regulate transcription of the chromosomal MDR1 gene. Thus, in K562 cells, 12-O-tetradecanoylphorbol-13-acetate-inducible expression of P-glycoprotein, the product of MDR1 gene, was strongly and selectively inhibited by the presence of a repressor protein targeted to the MDR1 promoter. These studies potentially provide a novel alternative approach to the control of multidrug resistance. They also provide important insights into strategies for developing selective regulators of gene expression. PMID- 10860922 TI - Subunit-specific action of an anticonvulsant thiobutyrolactone on recombinant glycine receptors involves a residue in the M2 membrane-spanning region. AB - The anticonvulsant alpha-ethyl, alpha-methyl-gamma-thiobutyrolactone (alphaEMTBL) potentiates the response to a submaximal concentration of glycine produced by receptors composed of human glycine alpha1-subunits but reduces the response of receptors composed of rat glycine alpha3-subunits. Both the potentiating and blocking actions of alphaEMTBL are reduced by higher concentrations of glycine. The subunit specificity of alphaEMTBL block is conferred by a residue in the second membrane-spanning region (M2), which is alanine in the alpha3-subunit (A254) and glycine in the alpha1-subunit. The mutation A254G in the alpha3 subunit removes blocking by alphaEMTBL and reveals potentiation. Picrotin, a picrotoxinin analog, blocks responses of receptors composed of either alpha1 or alpha3-subunits. Blocking of alpha3 receptors by picrotin is reduced in the presence of alphaEMTBL, indicating that the mechanisms interact at some point, but the mutation alpha3 A254G does not remove block by picrotin. However, mutation of a nearby residue alpha3 T258F does remove block by picrotin, picrotoxinin and alphaEMTBL. These observations suggest that alphaEMTBL and picrotin act on glycine alpha3 receptors to produce block by an allosteric mechanism that involves overlapping sets of residues in the M2 region. Coexpression of the alpha3-subunit with the beta-subunit of the glycine receptor also removes block by alphaEMTBL and reveals potentiation, suggesting that receptors containing either alpha3 or alpha1 glycine receptor subunits are potentiated in the adult brain. PMID- 10860923 TI - Sarco-endoplasmic ATPase blocker 2,5-Di(tert-butyl)-1, 4-benzohydroquinone inhibits N-, P-, and Q- but not T-, L-, or R-type calcium currents in central and peripheral neurons. AB - The effects of 2,5-di(tert-butyl)-1,4-benzohydroquinone (tBHQ), a synthetic phenolic antioxidant and a blocker of the sarco-endoplasmic ATPase, were evaluated on low and high voltage-activated Ca(2+) currents (ICas) with rodent dorsal root ganglion, hippocampal, and motor neurons. In all cell types tested, tBHQ (IC(50) = 35 microM) blocked ICa at concentrations used to inhibit sarco endoplasmic ATPase. This effect was specific to tBHQ because the other sarco endoplasmic reticulum calcium ATPase pump inhibitors (thapsigargin and cyclopiazonic acid) had no effect. Selective blockade of the N-type current with omega-conotoxin GVIA and of P- (motoneuron) or Q-type currents (hippocampal neuron) with omega-agatoxin IVA indicated that tBHQ inhibited N, P, and Q types of ICa. tBHQ had no effect on nitrendipine-sensitive (L-type) and residual drug resistant (R-type) ICa, nor on the low voltage-activated T-type ICa. Contrary to neuronal cells, the L-type ICa was inhibited by tBHQ in a differentiated mouse neuroblastoma and rat glioma hybrid cell line. Injection of cDNAs encoding the alpha1A, alpha1B, alpha1C, and alpha1E subunits into oocytes showed that tBHQ blocked ICas at the level of the pore-forming protein. This effect of tBHQ on ICa should be considered when interpreting results obtained with tBHQ used on neuronal preparations. It also may be useful for developing new strategies for the generation of more potent intracellular calcium transient inhibitors. PMID- 10860924 TI - Anticancer derivative of butyric acid (Pivalyloxymethyl butyrate) specifically potentiates the cytotoxicity of doxorubicin and daunorubicin through the suppression of microsomal glycosidic activity. AB - Pivalyloxymethyl butyrate (AN9) is an anticancer derivative of butyric acid. In this study, doxorubicin (DXR) and AN9 synergistically inhibited the growth of lymphoma and lung carcinoma cells, whereas there was no synergy between AN9 and antimetabolites. AN9 did not affect the intracellular uptake of DXR. Among anthracyclines and their derivatives, the synergistic effect was prominent in compounds with a daunosamine moiety, suggesting that AN9 may affect the catabolism of these compounds. The degradation of DXR in the extract from AN9 treated cells was much less than that in extract from untreated cells. AN9 did not directly inhibit the enzyme activity but rather suppressed expression of the enzyme. With respect to the expression of drug resistance-related genes, there was no significant difference between untreated and AN9-treated cells. However, AN9 significantly down-regulated the levels NADPH-cytochrome P450 reductase and DT-diaphorase mRNA in the presence of DXR but not the level of xanthine oxidase mRNA. The enhancement of the sensitivity to anthracyclines was closely associated with the suppression of the mRNA expression. PMID- 10860925 TI - Functional analysis of a tryptophan-less P-glycoprotein: a tool for tryptophan insertion and fluorescence spectroscopy. AB - P-glycoprotein (Pgp) functions as an ATP-dependent drug efflux pump to confer multidrug resistance to tumor cells. In the absence of a high-resolution structure for this protein, several important and intriguing aspects of Pgp structure and function remain poorly understood. Fluorescence spectroscopy of endogenous or genetically engineered tryptophan residues represents a potentially powerful method to probe static and dynamic aspects of Pgp at high resolution. We have used site-directed mutagenesis to modify the wild-type (WT) mouse mdr3 Pgp for tryptophan fluorescence spectroscopy by replacement of all 11 tryptophan residues individually with phenylalanine. None of the 11 tryptophans were found to be absolutely essential for Pgp activity, because Chinese hamster ovary cells transfected and overexpressing this mutant Trp-less mdr3 cDNA (mdr3F(1-11)) become multidrug-resistant and can carry out active transport of vinblastine, colchicine, and Calcein-AM. The mdr3F(1-11) mutant has reduced activity compared with WT Mdr3, and shows a unique pattern of drug resistance clearly distinct from WT and, as opposed to the latter, can neither confer FK-506 resistance nor functionally complement ste6 in yeast. Studies with Pgp mutants containing either single or double tryptophan residues or with chimeric molecules constructed between wild-type Pgp and mdr3F(1-11) indicated that no single tryptophan residue was responsible for the reduced activity of the mdr3F(1-11) mutant. Likewise, all but one chimeric Pgp preserved the unique drug resistance profile of the mdr3F(1 11) mutant. Altogether, we show that a Trp-less Pgp is functionally active and can be used as a molecular backbone for insertion of tryptophans in strategic locations to probe various aspects of Pgp function. PMID- 10860926 TI - Two-stage glucocorticoid induction of CYP3A23 through both the glucocorticoid and pregnane X receptors. AB - Glucocorticoid inducibility of the CYP3A23 gene is conferred by a multisite unit comprising binding sites for several members of the nuclear receptor superfamily of transcription factors, including the chicken ovalbumin upstream promoter transcription factor COUP-TF, pregnane X receptor (PXR), and hepatocyte nuclear factor 4 (HNF-4). The presence of three binding sites, each of which interacts with more than one factor, contributes to the complexity of the CYP3A23 glucocorticoid-responsive region. Despite the glucocorticoid sensitivity of this gene, direct binding of ligand-activated glucocorticoid receptor (GR) to the CYP3A23 dexamethasone-responsive region (DexRE) is not required for induction. This study demonstrates that DexRE-2 is the key element within the CYP3A23 proximal promoter, conferring ligand sensitivity via its interaction with the PXR/RXRalpha heterodimer. The DexRE-1 and HNF-4 sites are not ligand-responsive, but are essential accessory elements required for full promoter inducibility. In addition to ligand-mediated activation of PXR, the overall induction response involves a GR-mediated stimulation of PXR and RXRalpha expression. Hence, the induction pathway can be divided into two stages. In stage one, maximal induction requires a GR-dependent increase in PXR and RXRalpha expression, and stage two is characterized by direct transcriptional activation of CYP3A23, which is dependent on ligand-activated PXR as well as accessory factors bound at the DexRE-1 and HNF 4 sites. Because multiple proteins bind at each element within the glucocorticoid responsive region, factors not contributing to ligand responsiveness, such as chicken ovalbumin upstream promoter-transcription factor, may modulate the response through competitive interactions. PMID- 10860927 TI - Mechanism of action of the potent sodium-retaining steroid 11, 19 oxidoprogesterone. AB - We have demonstrated previously that a planar conformation of the molecular frame is required for steroids to acquire optimal sodium-retaining activity and binding properties to the mineralocorticoid receptor (MR). One of the most active sodium retaining compounds tested in those studies was 11, 19-oxidoprogesterone. Despite its biological potency, the relative affinity of 11,19-oxidoprogesterone for the MR is 5-fold lower than that of 21-deoxycorticosterone and 10-fold lower than aldosterone. Such a discrepancy may be assigned to uncommon biopharmacological properties of this synthetic steroid or an unusual molecular mechanism of action. In this work, we studied the biopharmacological and mechanistic features of 11,19 oxidoprogesterone. We show that both the pharmacokinetic properties of 11,19 oxidoprogesterone and its ability to transform and translocate the MR into the nucleus are undistinguishable from aldosterone. However, the capability of the serine/threonine phosphatase inhibitor tautomycin to impair nuclear translocation of the aldosterone-MR complex is not observed for the 11,19-oxidoprogesterone-MR complex. In addition, the binding properties of both steroids are differentially affected by modification of crucial lysyl residues of the MR. Kinetic studies performed on the aldosterone-MR complex in the presence of low concentrations of oxidopregnane suggest that 11,19-oxidoprogesterone may bind to the MR in a different binding site from the aldosterone binding pocket. Consistent with this postulate, a biologically inactive dose of 0.6 ng of oxidopregnane is able to potentiate the mineralocorticoid effect of a suboptimal dose of aldosterone. PMID- 10860928 TI - Acetyl-boswellic acids are novel catalytic inhibitors of human topoisomerases I and IIalpha. AB - Acetyl-boswellic acids (acetyl-BA) are pentacyclic triterpenes derived from the gum resin of frankincense. We have previously shown that these compounds are effective cytotoxic agents, acting through a mechanism that appears to involve the inhibition of topoisomerase activity. We have now investigated the mechanism of action of acetyl-BA and show that these compounds are more potent inhibitors of human topoisomerases I and IIalpha than camptothecin, and amsacrine or etoposide, respectively. Our data demonstrate that acetyl-BA and, to a lesser extent, some other pentacyclic triterpenes, such as betulinic acid, ursolic acid, and oleanolic acid, inhibit topoisomerases I and IIalpha through a mechanism that does not involve stabilization of the cleavable complex or the intercalation of DNA. Surface plasmon resonance analysis revealed that topoisomerases I and IIalpha bind directly to an immobilized derivative of acetyl-BA. This acetyl-BA derivative interacts with human topoisomerases through high-affinity binding sites yielding K(D) values of 70.6 nM for topoisomerase I and 7.6 nM for topoisomerase IIalpha. Based on our data, we propose that acetyl-BA inhibit topoisomerases I and IIalpha through competition with DNA for binding to the enzyme. Thus, acetyl-BA are a unique class of dual catalytic inhibitors of human topoisomerases I and IIalpha. PMID- 10860929 TI - Brilliant blue G selectively blocks ATP-gated rat P2X(7) receptors. AB - There are few antagonists selective for subtypes of the several P2X receptors, but these are needed to identify the receptors expressed on native cells and tissues. In particular, P2X(4) and P2X(7) receptor subunits are colocalized on immune, epithelial, and exocrine gland cells, but both are relatively insensitive to suramin and pyridoxal-5-phosphate-6-azo-2',4'-disulfonic acid derivative. In this article, we show that Coomassie Brilliant Blue G selectively inhibits P2X(7) receptors with nanomolar affinity. We measured currents in response to P2X receptor activation in HEK293 cells heterologously expressing human or rat P2X(1), P2X(2), P2X(3), P2X(2/3), P2X(4), P2X(1/5), and P2X(7) receptors. Brilliant Blue G produced a noncompetitive inhibition of rat and human P2X(7) receptors with IC(50) values of 10 and 200 nM, respectively. IC(50) values for inhibition of the other receptors ranged from 2 to >30 microM; the rat and human P2X(4) receptors showed IC(50) values of >10 and 3.2 microM. Coomassie Blue G also blocked YO-PRO1 uptake and membrane blebbing, which are uniquely associated with activation of P2X(7) receptors. Thus, Brilliant Blue G is at least 1000-fold more potent at rat P2X(7) receptors than at rat P2X(4) receptors. PMID- 10860930 TI - Characterization of the UDP-glucuronosyltransferase 1A locus in lagomorphs: evidence for duplication of the UGT1A6 gene. AB - The UGT1 locus is felt to be highly conserved between species, as is evident from the characterization of the locus in rodents and humans. In rabbits, cDNAs encoding proteins homologous to human UGT1A4, UGT1A6, and UGT1A7 have previously been identified. Here we demonstrate by Southern blot analysis, using exon 1 divergent 5' segments from rabbit UGT1A4 and UGT1A6 cDNAs, the existence of a cluster of highly related genes that are homologous to each of these exon 1 sequences. In comparing rabbit and human, it is evident that the UGT1A4 and UGT1A6 gene clusters in rabbit have undergone gene duplication. This is particularly evident with rabbit UGT1A6. The human UGT1A6 cDNA anneals to only a single gene fragment, as displayed by Southern blot analysis, indicating that the UGT1A6 exon 1 sequence is highly conserved. However, up to six rabbit UGT1A6 genes could be predicted from Southern blot analysis. To examine the potential linkage of the rabbit UGT1A6 genes, multiple UGT1A6 exons were identified from genomic DNA by extended polymerase chain reaction techniques and cloning of the UGT1A6 exon 1 sequences. Five unique UGT1A6 exon 1 gene sequences were characterized that could be predicted to encode proteins that are 98% similar in amino acid structure. Using a conserved region of the rabbit UGT1A6 cDNA as a probe to screen cDNA libraries, we identified a second UGT1A6 cDNA, termed UGT1A6alpha. In addition, a cDNA that encodes a protein similar to human UGT1A3 was also cloned. Characterization of UGT1A6alpha demonstrated the protein to be 98.9% identical to UGT1A6. The expression of rabbit UGT1A3, UGT1A4, and UGT1A6 displayed catalytic activities similar to their human orthologs. However, UGT1A6alpha was catalytically divergent from UGT1A6, indicating that UGT1A6 and UGT1A6alpha do not arise from allelic polymorphism. These results demonstrate that lagomorphs have evolved at least five additional UGT1A6 genes, an event that is not duplicated in rodents or humans. PMID- 10860931 TI - Anticonvulsants but not general anesthetics have differential blocking effects on different T-type current variants. AB - The sensitivity to anticonvulsants and anesthetics of Ca(2+) currents arising from alpha1G and alpha1H subunits was examined in stably transfected HEK293 cells. For comparison, in some cases blocking effects on dorsal root ganglion (DRG) T currents were also examined under identical ionic conditions. The anticonvulsant, phenytoin, which partially blocks DRG T current, blocked alpha1G current completely but with weaker affinity ( approximately 140 microM). Among different cells, alpha1H current exhibited either of two responses to phenytoin. In one subpopulation of cells, phenytoin produced a partial, higher affinity block (IC(50) approximately 7.2 microM, maximum block approximately 43%) similar to that in DRG neurons. In other cells, phenytoin produced complete, but lower affinity, blockade (IC(50) approximately 138 microM, maximum block approximately 89%). Another anticonvulsant, alpha-methyl-alpha-phenylsuccinimide (MPS), blocked DRG current partially, but blocked both alpha1G and alpha1H currents completely with weaker affinity ( approximately 1.7 mM). These data suggest that higher affinity blockade of T-type currents by phenytoin and MPS may require additional regulatory factors that can contribute to native T-type channels. In contrast, anesthetics blocked all T current variants similarly and completely. Block of alpha1G current by anesthetics had the following order of potency: propofol (IC(50) approximately 20.5 microM) > etomidate ( approximately 161 microM) = octanol ( approximately 160 microM) > isoflurane ( approximately 277 microM) > ketamine ( approximately 1.2 mM), comparable with results on DRG T currents. Barbiturates completly blocked alpha1G currents with potency [thiopental ( approximately 280 microM), pentobarbital ( approximately 310 microM), phenobarbital ( approximately 1.54 mM)] similar to that in DRG cells. The effects of propofol, octanol, and pentobarbital on alpha1H currents were indistinguishable from effects on alpha1G currents. PMID- 10860932 TI - Subtype-selective inhibition of neuronal nicotinic acetylcholine receptors by cocaine is determined by the alpha4 and beta4 subunits. AB - Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels of the central and peripheral nervous system that regulate synaptic activity from both pre- and postsynaptic sites. Nicotine binding to brain nAChRs is thought to underlie the induction of behavioral addiction to nicotine, probably as a result of desensitizing/inhibitory effects. Here, another commonly abused drug, cocaine, is shown to selectively inhibit particular nAChR subtypes with a potency in the low micromolar range by interacting with separate sites associated with the alpha4 and beta4 nAChR subunits. Chimeric receptor subunits and site-directed mutants were used to localize sequence determinants of cocaine affinity to: 1) a region of alpha4 located between residues 128 and 267 and 2) a site within the pore-lining M2 domain of beta4 that includes the 13' phenylalanine residue. The voltage dependence for inhibition associated with each site is consistent with these results. Analysis of the effects of incorporation of mutant and chimeric subunits also permitted identification of sequence elements important in receptor activation. For alpha3-containing receptors, a region or regions contained within the N-terminal extracellular domain of neuronal beta subunits influence the time course of responses to acetylcholine. Conversely, the 13' residue of the beta4 subunit M2 region is important in determining acetylcholine potency, indicating a role for this residue in agonist binding/gating processes. In summary, the present work describes sequence elements critical in both cocaine inhibition and acetylcholine activation of nAChRs and indicates that nAChRs may provide a site of interaction for the effects of nicotine and cocaine in the nervous system. PMID- 10860933 TI - Inhibition of cell surface expression by mutant receptors demonstrates that D2 dopamine receptors exist as oligomers in the cell. AB - Numerous mutant G protein-coupled receptors with diminished or no function have been described that are naturally occurring or that are the product of gene manipulation. It has largely been assumed that receptor mutants do not affect the function of the wild-type receptor; however, the occurrence of G protein-coupled receptor dimerization suggests the possibility that an intermolecular interaction between mutant and wild-type receptors can occur. We have shown previously that the D2 dopamine receptor (D2DR) exists as dimers in cell lines and brain tissue. In this study, we demonstrated that mutant D2DR can modulate the function of the wild-type D2DR. While attempting to elucidate the structure of the D2DR dimer, we demonstrated that nonfunctional D2DR substitution and truncation mutants antagonized wild-type D2DR function. Furthermore, from analyses of this interaction between the receptor mutants and the D2DR, using photoaffinity labeling, we provide evidence that the D2DR is oligomeric in the cell. PMID- 10860934 TI - Molecular basis for proton regulation of glycine transport by glycine transporter subtype 1b. AB - In the central nervous system, glycine is a coagonist with glutamate at the N methyl-D-aspartate subtype of ionotropic glutamate receptors. The GLYT1b subtype of glycine transporters is expressed in similar regions of the brain as the excitatory N-methyl-D-aspartate receptors and has been postulated to regulate glycine concentrations within excitatory synapses. We have expressed GLYT1b in Xenopus laevis oocytes and used electrophysiological techniques to investigate the pH regulation of glycine transporter function. We found that H(+) inhibits glycine transport by a noncompetitive mechanism, with half-maximal inhibition occurring at concentrations found in both physiological and pathological conditions. Charge-to-flux experiments revealed that the decreased current measured corresponds to a decreased influx of [(3)H]glycine and that the proton inhibition of GLYT1b does not alter the coupling ratio of transport. The membrane potential does not affect proton inhibition of transport, suggesting that the site of action on GLYT1b is not within the electric field of the membrane. Mutation of histidine 421 to an alanine residue, in the fourth extracellular loop of GLYT1b, renders the transporter insensitive to regulation by pH, but does not seem to alter the kinetics of glycine transport. These results suggests that histidine 421 is responsible for mediating the inhibitory actions of protons. Proton modulation of GLYT1b may be an important factor in determining the dynamics of excitatory neurotransmission. PMID- 10860935 TI - Pharmacological properties of 5-Hydroxytryptamine(4) receptor antagonists on constitutively active wild-type and mutated receptors. AB - We studied the pharmacological properties of twenty-four 5-hydroxytryptamine (5 HT)(4) receptor ligands known to act as antagonists on 5-HT(4) receptors positively coupled to adenylyl cyclase endogenously expressed in mouse colliculi neurons. In COS-7 cells expressing human or mouse 5-HT(4(a)) receptors (100-8000 fmol/mg of protein), we found neutral antagonists, partial agonists, and inverse agonists. The majority of neutral antagonists belong to the benzodioxanyl ketone class, whereas partial agonists belong to different chemical classes. We found only two inverse agonists, GR 125487 and SB 207266, which are both indoles. Analysis of pharmacological characteristics of the constitutively active wild type and constitutively active mutated receptors revealed that 1) the ratio between the efficiencies of the full agonist 5-HT and the partial agonist RS 23597 was invariable when the receptor density increased, but was dependent on receptor structure; 2) similarly, the efficacy of the inverse agonist SB 207266 was not dependent on receptor density but was dependent on receptor structure; 3) when the receptor concentration increased, the EC(50) values of the full agonist 5-HT were not modified and the increase in basal constitutive activity, as well as its stimulation by 5-HT, followed a parallel evolution; and 4) the stimulation of basal constitutive activity by 5-HT was not modified by the overexpression of Galphas. All these results indicate that in COS-7 cells, the coupling of the 5 HT(4) receptor to adenylyl cyclase was linear with no indication of spare receptors even at high receptor density (8 pmol/mg). These results are also in accordance with a precoupling between the activated receptor (f(R*)) and adenylyl cyclase. Such observations allowed us to use the two-state model to calculate the constant J, i.e., the equilibrium allosteric constant denoting the ratio of the receptor in the inactive versus active state (J = [R]/[R*]). We found that J was a receptor structural characteristic, independent of receptor density. PMID- 10860936 TI - Elevation of intracellular cAMP inhibits growth factor-mediated matrix metalloproteinase-9 induction and keratinocyte migration. AB - Receptor tyrosine kinases are regulators of diverse cellular functions including cell growth, cell survival, differentiation, locomotion, and morphogenesis. Activation of the cAMP-dependent protein kinase A inhibits receptor tyrosine kinase-stimulated growth responses in a number of cell types. In this study, we investigated the consequences of elevated cAMP on growth factor-mediated keratinocyte migration and matrix metalloproteinase (MMP)-9 induction in a human keratinocyte cell line. We found that elevation of intracellular cAMP by forskolin abolishes epidermal growth factor (EGF)- or scatter factor/hepatocyte growth factor-dependent colony dispersion. Concentrations of forskolin that inhibit growth factor-induced motility also eliminate EGF- or scatter factor/hepatocyte growth factor-dependent induction of the 92-kDa gelatinase/MMP 9. In contrast to findings obtained in fibroblasts, elevated intracellular cAMP did not interfere with growth factor-dependent activation of the p42/44 extracellular signal-regulated kinases, indicating that cAMP-dependent inhibition of migration and MMP-9 induction does not occur through perturbation of the extracellular signal-regulated kinases/mitogen-activated protein kinase pathway. However, forskolin effectively inhibited EGF-dependent activation of c-Jun N terminal kinase and p38, demonstrating that cAMP selectively interferes with a different subset of growth factor-induced mitogen-activated protein kinase signaling cascades than reported previously in fibroblasts. These findings illustrate that EGF concurrently activates multiple mitogen-activated protein kinase signaling cascades in keratinocytes and suggests that each pathway contributes to maximal EGF-dependent migration and proteinase induction. PMID- 10860937 TI - Activation of c-Ha-ras by benzo(a)pyrene in vascular smooth muscle cells involves redox stress and aryl hydrocarbon receptor. AB - Repeated cycles of vascular injury by benzo(a)pyrene (BaP) increase the onset and progression of atherosclerotic lesions in laboratory animals. This atherogenic response is partly mediated by activation of cis-acting antioxidant/electrophile response elements that enhance c-Ha-ras transcription in vascular smooth muscle cells (vSMCs). Activation of antioxidant/electrophile responsive cis-acting elements may depend on metabolism of BaP by cytochrome P450s to intermediates that induce oxidative stress and modulate gene expression. To test this hypothesis, we evaluated mitogen-activated c-Ha-ras expression in vSMCs treated with BaP or its metabolic intermediates alone, and in combination with agents that modulate cellular redox status. BaP (0.3 and 3 microM), BaP-3, 6-quinone (0.3 microM), or hydrogen peroxide (50 microM) enhanced serum-activated c-Ha-ras. Ellipticine (0.01 nM), a known inhibitor of cytochrome P450 metabolism and aryl hydrocarbon receptor (AhR) antagonist, inhibited c-Ha-ras induction by BaP (3 microM). Serum challenge of G(0) synchronized cultures of vSMCs with DL buthionine-(S,R)-sulfoximine (0.1 mM), a depletor of cellular glutathione, increased c-Ha-ras mRNA levels during the early phase of the mitogenic response. Combined BaP/DL-buthionine-(S, R)-sulfoximine challenge was cytotoxic to the cells and inhibited c-Ha-ras expression, whereas up-regulation of antioxidant capacity by N-acetylcysteine (0.5 mM) precluded BaP-induced ras expression. BaP increased formation of reactive oxygen species and depleted cellular glutathione, but these changes did not correlate with the kinetics of c-Ha-ras induction. BaP did not enhance c-Ha-ras expression in vSMCs from AhR knockout mice, although aryl hydrocarbon hydroxylase activity was constitutively expressed in these cells. These results suggest that c-Ha-ras activation in vSMCs by BaP involves a redox-sensitive mechanism that is coupled to AhR receptor-dependent functions. PMID- 10860938 TI - Overexpression of dynamin is induced by chronic stimulation of mu- but not delta opioid receptors: relationships with mu-related morphine dependence. AB - Several studies using selective opioid agonists or mice with a deletion of the mu opioid receptor, have shown that morphine dependence is essentially due to chronic stimulation of mu- but not delta-opioid receptors. Because dependence is assumed to be related to persistent intracellular modifications, we have investigated modifications putatively induced by chronic activation of mu receptors with morphine or selective agonists in vitro in SH-SY5Y cells and in vivo in different strains of mice, including mice lacking the mu-opioid receptor gene. The results show a similar down-regulation and desensitization of mu and delta binding sites, whereas an overexpression of dynamin occurred only with mu agonists, strongly suggesting the relevance of this up-regulation with the opiate dependence. Moreover, translocation of overexpressed dynamin from intracellular pools to plasma membranes was observed in chronic morphine-treated rats. This recruitment could be critically involved in long-lasting changes such as alterations of axonal transport observed in opioid dependence. PMID- 10860939 TI - Cellular response to a glutathione S-transferase P1-1 activated prodrug. AB - TER286 [gamma-glutamyl-alpha-amino-beta(2-ethyl-N,N,N', N'-tetrakis(2 chloroethyl)phosphorodiamidate)-sulfonyl-propionyl-( R)- (-) phenylglycine] is a novel nitrogen mustard prodrug that is preferentially activated by glutathione S transferase P1-1 (GSTP1-1). A human promyelocytic leukemia /TER286-resistant cell line was selected by chronic, long-term exposure to the prodrug. Although resistance was not readily achieved, eventually a 5-fold resistant clone was isolated. Cross-resistance to melphalan occurred, but not to doxorubicin (Adriamycin), taxol, and gamma-glutamyl-S-(benzyl)cysteinyl-R(-)-phenyl glycine diethyl ester, a GSTP1-1 inhibitor. The protein and transcript levels and enzymatic activity of GSTP1-1 were reduced significantly in the selected resistant line. GSTalpha levels were unchanged, and GSTmu was undetectable. Although glutathione levels were elevated in human promyelocytic leukemia/TER286 cells, no changes in the expression of thiol-related genes including gamma glutamylcysteine synthetase, gamma-glutamyl transpeptidase, or multidrug resistance protein were found. A 7-fold increase in catalase expression in the resistant cell line indicated an adaptive response to oxidative and electrophilic stress, and this was also reflected in the lower prevalence of drug-induced DNA single-strand breaks in the resistant cells. Mouse embryo fibroblast GSTP1-1(-/-) cells exhibited 2-fold resistance to TER286 compared with GSTP1-1(+/+) cells. NIH3T3 cells transfected with combinations of gamma-GCS and multidrug resistance protein exhibited enhanced resistance to TER286, although the degree of resistance was impaired by cotransfection of GSTP1-1. These results are consistent with responses in the TER286-resistant cells indicative of GSTP1-1 mediated mechanism of activation. In consequence, these data support the rationale that tumors expressing high levels of GSTP1-1 will be more sensitive to the cytotoxic effects of the drug. PMID- 10860940 TI - Scanning mutagenesis identifies amino acid side chains in transmembrane domain 5 of the M(1) muscarinic receptor that participate in binding the acetyl methyl group of acetylcholine. AB - The exofacial part of transmembrane domain 5 (TMD 5) of the cationic amine binding subclass of 7-transmembrane receptors is thought to be important in binding the side chain of the agonist. Residues Ile-188 through Ala-196 in TMD 5 of the M(1) muscarinic acetylcholine receptor (mAChR) have been studied by Cys- and Ala-scanning mutagenesis. The results are consistent with a helical conformation for this sequence. The positively charged sulfhydryl reagent N trimethyl-2-aminoethyl methanethiosulfonate reacted selectively with Phe-190 --> Cys, Thr-192 --> Cys, and Ala-193 --> Cys, indicating that the face of TMD 5 accessible from the binding site crevice is consistent with a recent model by Baldwin and colleagues of the transmembrane domain of the 7-transmembrane receptors. In contrast, the acetylcholine derivative bromoacetylcholine reacted selectively with Thr-192 --> Cys, which forms the focus of a group of amino acids (Ile-188, Thr-189, Thr-192, Ala-196) whose mutation decreased the binding affinity of the transmitter ACh itself. The center of this patch of residues is offset to one side of the binding pocket, suggesting that a rotation of TMD 5, relative to that implied by the Baldwin model, may be necessary to optimize the anchoring of acetylcholine within the binding site of the M(1) mAChR. An induced rotation of TMD 5 could contribute to the formation of the activated state of the receptor. PMID- 10860941 TI - Differences in the formation of PPARalpha-RXR/acoPPRE complexes between responsive and nonresponsive species upon fibrate administration. AB - Peroxisome proliferator-activated receptor-alpha (PPARalpha) is responsible for the hypolipidemic, peroxisome proliferation and carcinogenic effects of fibrates. Rats and mice are responsive, but guinea pigs and primates are resistant to the proliferative and carcinogenic effects of these drugs, but the hypolipidemic effect is still manifest. It is not yet clear whether humans should be considered unresponsive, and there is concern about the long-term safety of fibrates. We present molecular evidence for the reported resistance of human cells to peroxisome proliferation by describing a deficient interaction of nuclear extracts from human cells with an acyl-CoA oxidase (ACO)-peroxisome proliferator response element probe upon fibrate addition. Electrophoretic mobility shift assay analysis showed that ciprofibrate elicited a concentration-dependent increase in the binding of nuclear extracts from cells of rat (Morris) and human (HepG2) origin to an ACO-peroxisome proliferator response element probe, although in HepG2 cells the increase was of marginal statistical significance. In Morris cells, the increase was more marked than in HepG2 cells (4-fold versus 1.5-fold at 0.2 mM ciprofibrate), and maximal binding was achieved earlier in Morris (30 min) than in HepG2 cells (3 h). Morris cells responded to the addition of ciprofibrate by increasing the levels of ACO mRNA, whereas HepG2 did not. The ratio between PPARbeta/PPARalpha mRNAs was higher in HepG2 cells than in Morris cells (3.2 versus 1.9), pointing to an antagonizing effect of PPARbeta on PPARalpha activity. These results were obtained in untransfected cells expressing their own basal set of receptors. We also provide evidence of the translocation of PPARalpha from the cytosol to the nucleus upon activation by ciprofibrate. PMID- 10860942 TI - Allosteric interactions of staurosporine and other indolocarbazoles with N [methyl-(3)H]scopolamine and acetylcholine at muscarinic receptor subtypes: identification of a second allosteric site. AB - We have studied the interactions of five indolocarbazoles with N-[methyl (3)H]scopolamine (NMS) and unlabeled acetylcholine at M(1)-M(4) muscarinic receptors, using equilibrium and nonequilibrium radioligand binding studies. The results are consistent with an allosteric model in which the primary and allosteric ligands bind simultaneously to the receptor and modify each other's affinities. The compounds were generally most active at M(1) receptors. [(3)H]NMS binding was enhanced by staurosporine, KT5720, and KT5823 at M(1) and M(2) receptors, and by K-252a at M(1) receptors. Go 7874 reduced [(3)H]NMS affinity by up to threefold for all subtypes. A range of cooperative effects with acetylcholine was seen, and, at the M(1) receptor, KT5720 had a log affinity of 6.4 and enhanced acetylcholine affinity by 40%. The compounds inhibited the dissociation of [(3)H]NMS to different extents across the receptor subtypes, with the largest effects at M(1) receptors. In equilibrium binding studies the inhibitory potency of gallamine at M(1) receptors was not affected by KT5720, indicating that these agents bind to two distinct allosteric sites and have neutral cooperativity with each other. In contrast, gallamine and staurosporine had a negatively cooperative or competitive interaction at M(1) receptors. Similarly, the potency and relative effectiveness of KT5720 for inhibiting [(3)H]NMS dissociation from M(1) receptors were not affected by gallamine or brucine, but were affected in a complex manner by staurosporine. These results demonstrate that there are at least two distinct allosteric sites on the M(1) receptor, both of which can support positive cooperativity with acetylcholine. PMID- 10860943 TI - Induction of metabolism-dependent and -independent neutrophil apoptosis by clozapine. AB - Clozapine, an atypical antipsychotic used in the treatment of refractory schizophrenia, causes neutropenia and agranulocytosis in 3 and 0.8% of patients, respectively. Clozapine undergoes bioactivation to a chemically reactive nitrenium ion, which has been shown to cause neutrophil cytotoxicity. To define further the mechanism of cell death, we have investigated the toxicity of clozapine, its stable metabolites, and its chemically reactive nitrenium ion to neutrophils and lymphocytes. Clozapine was able to induce neutrophil apoptosis at therapeutic concentrations (1-3 microM) only when it was bioactivated to the nitrenium ion. The parent drug caused apoptosis at supratherapeutic concentrations (100-300 microM) only. Neutrophil apoptosis induced by the nitrenium ion, but not by the parent drug itself, was inhibited by antioxidants and genistein and was accompanied by cell surface haptenation (assessed by flow cytometry) and glutathione depletion. Dual-color flow cytometry showed that neutrophils that were haptenated were the same cells that underwent apoptosis. No apoptosis of lymphocytes was evident with the nitrenium ion or the parent drug, despite the fact that the former caused cell surface haptenation, glutathione depletion, and loss of membrane integrity. Demethylclozapine, the major stable metabolite in vivo, showed a profile that was similar to, although less marked than that observed with clozapine. N-oxidation of clozapine or replacement of the nitrogen (at position 5) by sulfur produced compounds that were entirely nontoxic to neutrophils. In conclusion, the findings of the study expand on potential mechanisms of clozapine-induced cytotoxicity, which may be of relevance to the major forms of toxicity encountered in patients taking this drug. PMID- 10860944 TI - Molecular characterization of the melanin-concentrating hormone/receptor complex: identification of critical residues involved in binding and activation. AB - A molecular model of the human melanin-concentrating hormone (MCH) peptide was constructed and docked into a helical, bacteriorhodopsin-based model of the recently identified human MCH receptor. From this hormone-receptor complex, potential sites of agonist-receptor interaction were identified, and site directed mutagenesis was used to substitute residues predicted to reside within the receptor binding pocket. Substitution of Asp(123)(3.32) in the third transmembrane domain of the receptor resulted in a loss of detectable (125)I-MCH binding and of MCH-stimulated Ca(2+) flux; cell surface expression of the mutant receptor was not affected. Arg(11) and Arg(14) of the MCH ligand were identified as potential sites of interaction with Asp(123)(3.32). [Ala(14)]-MCH was equipotent to native MCH in its ability to bind to and activate the wild-type MCH receptor, whereas [Ala(11)]-MCH displayed a 3000-fold reduction in binding affinity and a complete loss of measurable functional activity. Furthermore, [Lys(11)]-MCH and [D-Arg(11)]-MCH displayed reduced affinity for the receptor. [Lys(11)]-MCH was observed to be a partial agonist, eliciting approximately 67% of the native peptide's activity in a Ca(2+) flux assay, and [D-Arg(11)]-MCH was determined to be a functional antagonist with a K(b) valve of 15.8 microM. These data provide evidence that a basic moiety with specific stereochemical requirements at this site is needed for receptor activation. We conclude that both Asp(123)(3.32) in the MCH receptor and Arg(11) in the MCH peptide are required for the formation of the MCH peptide/receptor complex and propose that they form a direct interaction that is critical for receptor function. PMID- 10860945 TI - A galpha(s) carboxyl-terminal peptide prevents G(s) activation by the A(2A) adenosine receptor. AB - The molecular mechanisms of interaction between G(s) and the A(2A) adenosine receptor were investigated using synthetic peptides corresponding to various segments of the Galpha(s) carboxyl terminus. Synthetic peptides were tested for their ability to modulate binding of a selective radiolabeled agonist, [(3)H]2-[4 (2-carboxyethyl)phenylethylamino]-5'-N-ethylcarboxam idoade nosine ([(3)H]CGS21680), to A(2A) adenosine receptors in rat striatal membranes. The Galpha(s) peptides stimulated specific binding both in the presence and absence of 100 microM guanosine-5'-O-(3-thiotriphosphate) (GTPgammaS). Three peptides, Galpha(s)(378-394)C(379)A, Galpha(s)(376-394)C(379)A, and Galpha(s)(374 394)C(379)A, were the most effective. In the presence of GTPgammaS, peptide Galpha(s)(374-394)C(379)A increased specific binding in a dose-dependent fashion. However, the peptide did not stabilize the high-affinity state of the A(2A) adenosine receptor for [(3)H]CGS21680. Binding assays with a radiolabeled selective antagonist, [(3)H]5-amino-7-(2-phenylethyl)-2-(2-furyl)pyrazolo[4, 3-e] 1,2,4-triazolo[1,5-c]pyrimidine ([(3)H]SCH58261), showed that the addition of the Galpha(s) peptide modified the slope of the 5'-N-ethylcarboxamidoadenosine (NECA) competition curve, suggesting modulation of receptor affinity states. In the presence of GTPgammaS, the displacement curve was right-shifted, whereas the addition of Galpha(s)(374-394)C(379)A caused a partial left-shift. Both curves were fitted by one-site models. This same Galpha(s) peptide was also able to disrupt G(s)-coupled signal transduction as indicated by inhibition of the A(2A) receptor-stimulated adenylyl cyclase activity without affecting either basal or forskolin-stimulated enzymatic activity in the same membrane preparations. Shorter peptides from Galpha(s) and Galpha(i1/2) carboxyl termini were not effective. NMR spectroscopy showed the strong propensity of peptide Galpha(s)(374 394)C(379)A to assume a compact carboxyl-terminal alpha-helical conformation in solution. Overall, our results point out the conformation requirement of Galpha(s) carboxyl-terminal peptides to modulate agonist binding to rat A(2A) adenosine receptors and disrupt signal transduction. PMID- 10860946 TI - Activation of p55 tumor necrosis factor-alpha receptor-1 coupled to tumor necrosis factor receptor-associated factor 2 stimulates intercellular adhesion molecule-1 expression by modulating a thapsigargin-sensitive pathway in human tracheal smooth muscle cells. AB - Tumor necrosis factor-alpha (TNFalpha) stimulates the expression of intercellular adhesion molecule-1 (ICAM-1) by activating the transcription factor nuclear factor-kappaB (NF-kappaB) in human airway smooth muscle (ASM) cells. This study characterizes the receptor involved as well as critical downstream signaling events mediating cytokine-induced NF-kappaB activation and ICAM-1 expression. TNFalpha stimulation for 1 to 4 h induced ICAM-1 expression in human ASM cells. This rapid TNFalpha-induced ICAM-1 expression enhanced T-lymphocyte adhesion to ASM cells, which was inhibited by anti-ICAM-1 antibodies. Using immunostaining, we demonstrated that TNFalpha receptors TNFR1 and TNFR2 are expressed on native human tracheal smooth muscle. Treatment of cells with htr-9, an antibody that specifically activates TNFR1, also stimulated expression of ICAM-1 mRNA and protein. Utr-1, a blocking antibody to TNFR2, did not affect TNFalpha-mediated ICAM-1 expression. Both TNFalpha and htr-9 increased luciferase activity in ASM cells transfected with a NF-kappaB reporter plasmid. Overexpression of a dominant negative TNF receptor-associated factor 2 construct, lacking the NH(2)-terminal RING finger, completely abrogated both TNFalpha- and htr-9-mediated increases in NF-kappaB reporter activity. Thapsigargin, an agent that depletes intracellular calcium stores, abrogated both cytokine-mediated NF-kappaB-dependent ICAM-1 mRNA transcription and protein expression but had no effect on IkappaB degradation. In addition, chelating cytosolic calcium with 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid acetoxymethyl ester also inhibited cytokine TNFalpha-induced ICAM-1 expression. These data suggest that TNFR1, through a TNF receptor associated factor 2-NF-kappaB signaling pathway, mediates TNFalpha-induced expression of ICAM-1 on ASM cells by involving a thapsigargin-sensitive signaling pathway. PMID- 10860947 TI - 50 million years of chordate evolution: seeking the origins of adaptive immunity. PMID- 10860948 TI - Genome, diversity, and origins: the Y chromosome as a storyteller. PMID- 10860949 TI - Atherosclerosis: the emerging role of inflammation and the CD40-CD40 ligand system. PMID- 10860950 TI - Natural T cells: cranking up the immune system by prompt cytokine secretion. PMID- 10860951 TI - Gelsolin and ADF/cofilin enhance the actin dynamics of motile cells. PMID- 10860952 TI - Protection of auditory receptors and neurons: evidence for interactive damage. PMID- 10860953 TI - Variation and evolution in plants and microorganisms: toward a new synthesis 50 years after Stebbins. PMID- 10860954 TI - G. Ledyard Stebbins (1906-2000): an appreciation. PMID- 10860955 TI - Solution to Darwin's dilemma: discovery of the missing Precambrian record of life. AB - In 1859, in On the Origin of Species, Darwin broached what he regarded to be the most vexing problem facing his theory of evolution-the lack of a rich fossil record predating the rise of shelly invertebrates that marks the beginning of the Cambrian Period of geologic time ( approximately 550 million years ago), an "inexplicable" absence that could be "truly urged as a valid argument" against his all embracing synthesis. For more than 100 years, the "missing Precambrian history of life" stood out as one of the greatest unsolved mysteries in natural science. But in recent decades, understanding of life's history has changed markedly as the documented fossil record has been extended seven-fold to some 3,500 million years ago, an age more than three-quarters that of the planet itself. This long-sought solution to Darwin's dilemma was set in motion by a small vanguard of workers who blazed the trail in the 1950s and 1960s, just as their course was charted by a few pioneering pathfinders of the previous century, a history of bold pronouncements, dashed dreams, search, and final discovery. PMID- 10860956 TI - The chimeric eukaryote: origin of the nucleus from the karyomastigont in amitochondriate protists. AB - We present a testable model for the origin of the nucleus, the membrane-bounded organelle that defines eukaryotes. A chimeric cell evolved via symbiogenesis by syntrophic merger between an archaebacterium and a eubacterium. The archaebacterium, a thermoacidophil resembling extant Thermoplasma, generated hydrogen sulfide to protect the eubacterium, a heterotrophic swimmer comparable to Spirochaeta or Hollandina that oxidized sulfide to sulfur. Selection pressure for speed swimming and oxygen avoidance led to an ancient analogue of the extant cosmopolitan bacterial consortium "Thiodendron latens." By eubacterial archaebacterial genetic integration, the chimera, an amitochondriate heterotroph, evolved. This "earliest branching protist" that formed by permanent DNA recombination generated the nucleus as a component of the karyomastigont, an intracellular complex that assured genetic continuity of the former symbionts. The karyomastigont organellar system, common in extant amitochondriate protists as well as in presumed mitochondriate ancestors, minimally consists of a single nucleus, a single kinetosome and their protein connector. As predecessor of standard mitosis, the karyomastigont preceded free (unattached) nuclei. The nucleus evolved in karyomastigont ancestors by detachment at least five times (archamoebae, calonymphids, chlorophyte green algae, ciliates, foraminifera). This specific model of syntrophic chimeric fusion can be proved by sequence comparison of functional domains of motility proteins isolated from candidate taxa. PMID- 10860957 TI - Dynamic evolution of plant mitochondrial genomes: mobile genes and introns and highly variable mutation rates. AB - We summarize our recent studies showing that angiosperm mitochondrial (mt) genomes have experienced remarkably high rates of gene loss and concomitant transfer to the nucleus and of intron acquisition by horizontal transfer. Moreover, we find substantial lineage-specific variation in rates of these structural mutations and also point mutations. These findings mostly arise from a Southern blot survey of gene and intron distribution in 281 diverse angiosperms. These blots reveal numerous losses of mt ribosomal protein genes but, with one exception, only rare loss of respiratory genes. Some lineages of angiosperms have kept all of their mt ribosomal protein genes whereas others have lost most of them. These many losses appear to reflect remarkably high (and variable) rates of functional transfer of mt ribosomal protein genes to the nucleus in angiosperms. The recent transfer of cox2 to the nucleus in legumes provides both an example of interorganellar gene transfer in action and a starting point for discussion of the roles of mechanistic and selective forces in determining the distribution of genetic labor between organellar and nuclear genomes. Plant mt genomes also acquire sequences by horizontal transfer. A striking example of this is a homing group I intron in the mt cox1 gene. This extraordinarily invasive mobile element has probably been acquired over 1,000 times separately during angiosperm evolution via a recent wave of cross-species horizontal transfers. Finally, whereas all previously examined angiosperm mtDNAs have low rates of synonymous substitutions, mtDNAs of two distantly related angiosperms have highly accelerated substitution rates. PMID- 10860958 TI - The evolution of RNA viruses: A population genetics view. AB - RNA viruses are excellent experimental models for studying evolution under the theoretical framework of population genetics. For a proper justification of this thesis we have introduced some properties of RNA viruses that are relevant for studying evolution. On the other hand, population genetics is a reductionistic theory of evolution. It does not consider or make simplistic assumptions on the transformation laws within and between genotypic and phenotypic spaces. However, such laws are minimized in the case of RNA viruses because the phenotypic space maps onto the genotypic space in a much more linear way than on higher DNA-based organisms. Under experimental conditions, we have tested the role of deleterious and beneficial mutations in the degree of adaptation of vesicular stomatitis virus (VSV), a nonsegmented virus of negative strand. We also have studied how effective population size, initial genetic variability in populations, and environmental heterogeneity shapes the impact of mutations in the evolution of vesicular stomatitis virus. Finally, in an integrative attempt, we discuss pros and cons of the quasispecies theory compared with classic population genetics models for haploid organisms to explain the evolution of RNA viruses. PMID- 10860959 TI - Effects of passage history and sampling bias on phylogenetic reconstruction of human influenza A evolution. AB - In this paper we determine the extent to which host-mediated mutations and a known sampling bias affect evolutionary studies of human influenza A. Previous phylogenetic reconstruction of influenza A (H3N2) evolution using the hemagglutinin gene revealed an excess of nonsilent substitutions assigned to the terminal branches of the tree. We investigate two hypotheses to explain this observation. The first hypothesis is that the excess reflects mutations that were either not present or were at low frequency in the viral sample isolated from its human host, and that these mutations increased in frequency during passage of the virus in embryonated eggs. A set of 22 codons known to undergo such "host mediated" mutations showed a significant excess of mutations assigned to branches attaching sequences from egg-cultured (as opposed to cell-cultured) isolates to the tree. Our second hypothesis is that the remaining excess results from sampling bias. Influenza surveillance is purposefully biased toward sequencing antigenically dissimilar strains in an effort to identify new variants that may signal the need to update the vaccine. This bias produces an excess of mutations assigned to terminal branches simply because an isolate with no close relatives is by definition attached to the tree by a relatively long branch. Simulations show that the magnitude of excess mutations we observed in the hemagglutinin tree is consistent with expectations based on our sampling protocol. Sampling bias does not affect inferences about evolution drawn from phylogenetic analyses. However, if possible, the excess caused by host-mediated mutations should be removed from studies of the evolution of influenza viruses as they replicate in their human hosts. PMID- 10860960 TI - Bacteria are different: observations, interpretations, speculations, and opinions about the mechanisms of adaptive evolution in prokaryotes. AB - To some extent, the genetic theory of adaptive evolution in bacteria is a simple extension of that developed for sexually reproducing eukaryotes. In other, fundamental ways, the process of adaptive evolution in bacteria is quantitatively and qualitatively different from that of organisms for which recombination is an integral part of the reproduction process. In this speculative and opinionated discussion, we explore these differences. In particular, we consider (i) how, as a consequence of the low rates of recombination, "ordinary" chromosomal gene evolution in bacteria is different from that in organisms where recombination is frequent and (ii) the fundamental role of the horizontal transmission of genes and accessory genetic elements as sources of variation in bacteria. We conclude with speculations about the evolution of accessory elements and their role in the adaptive evolution of bacteria. PMID- 10860961 TI - Evolution of RNA editing in trypanosome mitochondria. AB - Two different RNA editing systems have been described in the kinetoplast mitochondrion of trypanosomatid protists. The first involves the precise insertion and deletion of U residues mostly within the coding regions of maxicircle-encoded mRNAs to produce open reading frames. This editing is mediated by short overlapping complementary guide RNAs encoded in both the maxicircle and the minicircle molecules and involves a series of enzymatic cleavage-ligation steps. The second editing system is a C(34) to U(34) modification in the anticodon of the imported tRNA(Trp), thereby permitting the decoding of the UGA stop codon as tryptophan. U-insertion editing probably originated in an ancestor of the kinetoplastid lineage and appears to have evolved in some cases by the replacement of the original pan-edited cryptogene with a partially edited cDNA. The driving force for the evolutionary fixation of these retroposition events was postulated to be the stochastic loss of entire minicircle sequence classes and their encoded guide RNAs upon segregation of the single kinetoplast DNA network into daughter cells at cell division. A large plasticity in the relative abundance of minicircle sequence classes has been observed during cell culture in the laboratory. Computer simulations provide theoretical evidence for this plasticity if a random distribution and segregation model of minicircles is assumed. The possible evolutionary relationship of the C to U and U-insertion editing systems is discussed. PMID- 10860962 TI - Population structure and recent evolution of Plasmodium falciparum. AB - Plasmodium falciparum is the agent of malignant malaria, one of mankind's most severe maladies. The parasite exhibits antigenic polymorphisms that have been postulated to be ancient. We have proposed that the extant world populations of P. falciparum have derived from one single parasite, a cenancestor, within the last 5, 000-50,000 years. This inference derives from the virtual or complete absence of synonymous nucleotide polymorphisms at genes not involved in immune or drug responses. Seeking to conciliate this claim with extensive antigenic polymorphism, we first note that allele substitutions or polymorphisms can arise very rapidly, even in a single generation, in large populations subject to strong natural selection. Second, new alleles can arise not only by single-nucleotide mutations, but also by duplication/deletion of short simple-repeat DNA sequences, a process several orders of magnitude faster than single-nucleotide mutation. We analyze three antigenic genes known to be extremely polymorphic: Csp, Msp-1, and Msp-2. We identify regions consisting of tandem or proximally repetitive short DNA sequences, including some previously unnoticed. We conclude that the antigenic polymorphisms are consistent with the recent origin of the world populations of P. falciparum inferred from the analysis of nonantigenic genes. PMID- 10860963 TI - Transposons and genome evolution in plants. AB - Although it is known today that transposons comprise a significant fraction of the genomes of many organisms, they eluded discovery through the first half century of genetic analysis and even once discovered, their ubiquity and abundance were not recognized for some time. This genetic invisibility of transposons focuses attention on the mechanisms that control not only transposition, but illegitimate recombination. The thesis is developed that the mechanisms that control transposition are a reflection of the more general capacity of eukaryotic organisms to detect, mark, and retain duplicated DNA through repressive chromatin structures. PMID- 10860964 TI - Maize as a model for the evolution of plant nuclear genomes. AB - The maize genome is replete with chromosomal duplications and repetitive DNA. The duplications resulted from an ancient polyploid event that occurred over 11 million years ago. Based on DNA sequence data, the polyploid event occurred after the divergence between sorghum and maize, and hence the polyploid event explains some of the difference in DNA content between these two species. Genomic rearrangement and diploidization followed the polyploid event. Most of the repetitive DNA in the maize genome is retrotransposable elements, and they comprise 50% of the genome. Retrotransposon multiplication has been relatively recent-within the last 5-6 million years-suggesting that the proliferation of retrotransposons has also contributed to differences in DNA content between sorghum and maize. There are still unanswered questions about repetitive DNA, including the distribution of repetitive DNA throughout the genome, the relative impacts of retrotransposons and chromosomal duplication in plant genome evolution, and the hypothesized correlation of duplication events with transposition. Population genetic processes also affect the evolution of genomes. We discuss how centromeric genes should, in theory, contain less genetic diversity than noncentromeric genes. In addition, studies of diversity in the wild relatives of maize indicate that different genes have different histories and also show that domestication and intensive breeding have had heterogeneous effects on genetic diversity across genes. PMID- 10860965 TI - Flower color variation: a model for the experimental study of evolution. AB - We review the study of flower color polymorphisms in the morning glory as a model for the analysis of adaptation. The pathway involved in the determination of flower color phenotype is traced from the molecular and genetic levels to the phenotypic level. Many of the genes that determine the enzymatic components of flavonoid biosynthesis are redundant, but, despite this complexity, it is possible to associate discrete floral phenotypes with individual genes. An important finding is that almost all of the mutations that determine phenotypic differences are the result of transposon insertions. Thus, the flower color diversity seized on by early human domesticators of this plant is a consequence of the rich variety of mobile elements that reside in the morning glory genome. We then consider a long history of research aimed at uncovering the ecological fate of these various flower phenotypes in the southeastern U.S. A large body of work has shown that insect pollinators discriminate against white phenotypes when white flowers are rare in populations. Because the plant is self-compatible, pollinator bias causes an increase in self-fertilization in white maternal plants, which should lead to an increase in the frequency of white genes, according to modifier gene theory. Studies of geographical distributions indicate other, as yet undiscovered, disadvantages associated with the white phenotype. The ultimate goal of connecting ecology to molecular genetics through the medium of phenotype is yet to be attained, but this approach may represent a model for analyzing the translation between these two levels of biological organization. PMID- 10860966 TI - Gene genealogies and population variation in plants. AB - Early in the development of plant evolutionary biology, genetic drift, fluctuations in population size, and isolation were identified as critical processes that affect the course of evolution in plant species. Attempts to assess these processes in natural populations became possible only with the development of neutral genetic markers in the 1960s. More recently, the application of historically ordered neutral molecular variation (within the conceptual framework of coalescent theory) has allowed a reevaluation of these microevolutionary processes. Gene genealogies trace the evolutionary relationships among haplotypes (alleles) with populations. Processes such as selection, fluctuation in population size, and population substructuring affect the geographical and genealogical relationships among these alleles. Therefore, examination of these genealogical data can provide insights into the evolutionary history of a species. For example, studies of Arabidopsis thaliana have suggested that this species underwent rapid expansion, with populations showing little genetic differentiation. The new discipline of phylogeography examines the distribution of allele genealogies in an explicit geographical context. Phylogeographic studies of plants have documented the recolonization of European tree species from refugia subsequent to Pleistocene glaciation, and such studies have been instructive in understanding the origin and domestication of the crop cassava. Currently, several technical limitations hinder the widespread application of a genealogical approach to plant evolutionary studies. However, as these technical issues are solved, a genealogical approach holds great promise for understanding these previously elusive processes in plant evolution. PMID- 10860967 TI - Toward a new synthesis: major evolutionary trends in the angiosperm fossil record. AB - Angiosperm paleobotany has widened its horizons, incorporated new techniques, developed new databases, and accepted new questions that can now focus on the evolution of the group. The fossil record of early flowering plants is now playing an active role in addressing questions of angiosperm phylogeny, angiosperm origins, and angiosperm radiations. Three basic nodes of angiosperm radiations are identified: (i) the closed carpel and showy radially symmetrical flower, (ii) the bilateral flower, and (iii) fleshy fruits and nutritious nuts and seeds. These are all coevolutionary events and spread out through time during angiosperm evolution. The proposal is made that the genetics of the angiosperms pressured the evolution of the group toward reproductive systems that favored outcrossing. This resulted in the strongest selection in the angiosperms being directed toward the flower, fruits, and seeds. That is why these organs often provide the best systematic characters for the group. PMID- 10860968 TI - Reproductive systems and evolution in vascular plants. AB - Differences in the frequency with which offspring are produced asexually, through self-fertilization and through sexual outcrossing, are a predominant influence on the genetic structure of plant populations. Selfers and asexuals have fewer genotypes within populations than outcrossers with similar allele frequencies, and more genetic diversity in selfers and asexuals is a result of differences among populations than in sexual outcrossers. As a result of reduced levels of diversity, selfers and asexuals may be less able to respond adaptively to changing environments, and because genotypes are not mixed across family lineages, their populations may accumulate deleterious mutations more rapidly. Such differences suggest that selfing and asexual lineages may be evolutionarily short-lived and could explain why they often seem to be of recent origin. Nonetheless, the origin and maintenance of different reproductive modes must be linked to individual-level properties of survival and reproduction. Sexual outcrossers suffer from a cost of outcrossing that arises because they do not contribute to selfed or asexual progeny, whereas selfers and asexuals may contribute to outcrossed progeny. Selfing and asexual reproduction also may allow reproduction when circumstances reduce opportunities for a union of gametes produced by different individuals, a phenomenon known as reproductive assurance. Both the cost of outcrossing and reproductive assurance lead to an over representation of selfers and asexuals in newly formed progeny, and unless sexual outcrossers are more likely to survive and reproduce, they eventually will be displaced from populations in which a selfing or asexual variant arises. PMID- 10860969 TI - Hybridization as a stimulus for the evolution of invasiveness in plants? AB - Invasive species are of great interest to evolutionary biologists and ecologists because they represent historical examples of dramatic evolutionary and ecological change. Likewise, they are increasingly important economically and environmentally as pests. Obtaining generalizations about the tiny fraction of immigrant taxa that become successful invaders has been frustrated by two enigmatic phenomena. Many of those species that become successful only do so (i) after an unusually long lag time after initial arrival, and/or (ii) after multiple introductions. We propose an evolutionary mechanism that may account for these observations. Hybridization between species or between disparate source populations may serve as a stimulus for the evolution of invasiveness. We present and review a remarkable number of cases in which hybridization preceded the emergence of successful invasive populations. Progeny with a history of hybridization may enjoy one or more potential genetic benefits relative to their progenitors. The observed lag times and multiple introductions that seem a prerequisite for certain species to evolve invasiveness may be a correlate of the time necessary for previously isolated populations to come into contact and for hybridization to occur. Our examples demonstrate that invasiveness can evolve. Our model does not represent the only evolutionary pathway to invasiveness, but is clearly an underappreciated mechanism worthy of more consideration in explaining the evolution of invasiveness in plants. PMID- 10860970 TI - The role of genetic and genomic attributes in the success of polyploids. AB - In 1950, G. Ledyard Stebbins devoted two chapters of his book Variation and Evolution in Plants (Columbia Univ. Press, New York) to polyploidy, one on occurrence and nature and one on distribution and significance. Fifty years later, many of the questions Stebbins posed have not been answered, and many new questions have arisen. In this paper, we review some of the genetic attributes of polyploids that have been suggested to account for the tremendous success of polyploid plants. Based on a limited number of studies, we conclude: (i) Polyploids, both individuals and populations, generally maintain higher levels of heterozygosity than do their diploid progenitors. (ii) Polyploids exhibit less inbreeding depression than do their diploid parents and can therefore tolerate higher levels of selfing; polyploid ferns indeed have higher levels of selfing than do their diploid parents, but polyploid angiosperms do not differ in outcrossing rates from their diploid parents. (iii) Most polyploid species are polyphyletic, having formed recurrently from genetically different diploid parents. This mode of formation incorporates genetic diversity from multiple progenitor populations into the polyploid "species"; thus, genetic diversity in polyploid species is much higher than expected by models of polyploid formation involving a single origin. (iv) Genome rearrangement may be a common attribute of polyploids, based on evidence from genome in situ hybridization (GISH), restriction fragment length polymorphism (RFLP) analysis, and chromosome mapping. (v) Several groups of plants may be ancient polyploids, with large regions of homologous DNA. These duplicated genes and genomes can undergo divergent evolution and evolve new functions. These genetic and genomic attributes of polyploids may have both biochemical and ecological benefits that contribute to the success of polyploids in nature. PMID- 10860971 TI - Toward antibody-catalyzed hydrolysis of organophosphorus poisons. AB - We report here our preliminary results on the use of catalytic antibodies as an approach to neutralizing organophosphorus chemical weapons. A first-generation hapten, methyl-alpha-hydroxyphosphinate Ha, was designed to mimic the approach of an incoming water molecule for the hydrolysis of exceedingly toxic methylphosphonothioate VX (1a). A moderate protective activity was first observed on polyclonal antibodies raised against Ha. The results were further confirmed by using a mAb PAR 15 raised against phenyl-alpha-hydroxyphosphinate Hb, which catalyzes the hydrolysis of PhX (1b), a less toxic phenylphosphonothioate analog of VX with a rate constant of 0.36 M(-1) x min(-1) at pH 7.4 and 25 degrees C, which corresponds to a catalytic proficiency of 14,400 M(-1) toward the rate constant for the uncatalyzed hydrolysis of 1b. This is a demonstration on the organophosphorus poisons themselves that mAbs can catalytically hydrolyze nerve agents, and a significant step toward the production of therapeutically active abzymes to treat poisoning by warfare agents. PMID- 10860972 TI - The asymptotic distribution of canonical correlations and variates in cointegrated models. AB - The cointegrated model considered here is a nonstationary vector autoregressive process in which some linear functions are stationary and others are random walks. The first difference of the process (the "error-correction form") is stationary. Statistical inference, such as reduced rank regression estimation of the coefficients of the process and tests of hypotheses of dimensionality of the stationary part, involves the canonical correlations between the difference vector and the relevant vector of the past of the process. The asymptotic distributions of the canonical correlations and the canonical vectors under the assumption that the process is Gaussian are found. PMID- 10860973 TI - Museum specimen data predict crop damage by tropical rodents. AB - Museum collections constitute a massive store of information on biological diversity. We used museum specimen data to generate ecological niche models that provide predictions of geographic distributions of native rodent pest species and agricultural census data that summarize the geographic distribution of nine crops in the state of Veracruz, Mexico, as well as crop losses between planting and harvest. Herein, we show that crop damage is related significantly to the predicted presence of rodent species for seven of nine crops. Museum collections may thus provide important baseline information for designing land-use and agricultural pest-management programs. PMID- 10860974 TI - Suppressed T helper 2 immunity and prolonged survival of a nematode parasite in protein-malnourished mice. AB - Protein malnutrition may increase susceptibility to gastrointestinal parasitic infections, possibly as a result of impaired intestinal and/or systemic T helper 2 (Th2) effector responses induced by down-regulation of Th2 cytokines and/or up regulation of Th1 cytokines. To test this hypothesis, female BALB/c mice (n = 18/diet) were fed a control (24%), marginal (7%), or deficient (3%) protein diet and given a challenge infection with Heligmosomoides polygyrus. The 3% mice had higher worm burdens at 1, 2, and 4 weeks postchallenge infection (pci), lower increases in serum IgE, reduced intestinal eosinophilia, and depressed mucosal mast cell proliferation and activation at 1-2 weeks pci. To determine whether these suppressed effector responses resulted from altered spleen and mesenteric lymph node (MLN) cytokine production, cells were restimulated in vitro with parasite antigen and cytokine concentrations were measured. Deficient MLN cells secreted significantly less IL-4 and more IFN-gamma at 1-2 weeks pci than did control MLN cells. Deficient spleen cells also secreted more IFN-gamma at 2 weeks pci compared with control spleen cells. From reverse transcription-PCR analyses, the 3% mice also had lower IL-4 mRNA level in spleen and MLN at 1-2 weeks pci. Our study supports the hypothesis that protein malnutrition increases the survival of a nematode parasite by decreasing gut-associated IL-4 (Th2) and increasing IFN-gamma (Th1) within 2 weeks pci, leading to reduced intestinal and systemic Th2 effector responses. PMID- 10860975 TI - Long-range order in the src SH3 folding transition state. AB - One of the outstanding questions in protein folding concerns the degree of heterogeneity in the folding transition state ensemble: does a protein fold via a large multitude of diverse "pathways," or are the elements of native structure assembled in a well defined order? Herein, we build on previous point mutagenesis studies of the src SH3 by directly investigating the association of structural elements and the loss of backbone conformational entropy during folding. Double mutant analysis of polar residues in the distal beta-hairpin and the diverging turn indicates that the hydrogen bond network between these elements is largely formed in the folding transition state. A 10-glycine insertion in the n-src loop (which connects the distal hairpin and the diverging turn) and a disulfide crosslink at the base of the distal beta-hairpin exclusively affect the folding rate, showing that these structural elements are nearly as ordered in the folding transition state as in the native state. In contrast, crosslinking the base of the RT loop or the N and C termini dramatically slows down the unfolding rate, suggesting that dissociation of the termini and opening of the RT loop precede the rate-limiting step in unfolding. Taken together, these results suggest that essentially all conformations in the folding transition state ensemble have the central three-stranded beta-sheet formed, indicating that, for the src homology 3 domain, there is a discrete order to structure assembly during folding. PMID- 10860976 TI - Pausing by bacterial RNA polymerase is mediated by mechanistically distinct classes of signals. AB - Transcript elongation by RNA polymerase is discontinuous and interrupted by pauses that play key regulatory roles. We show here that two different classes of pause signals punctuate elongation. Class I pauses, discovered in enteric bacteria, depend on interaction of a nascent RNA structure with RNA polymerase to displace the 3' OH away from the catalytic center. Class II pauses, which may predominate in eukaryotes, cause RNA polymerase to slide backwards along DNA and RNA and to occlude the active site with nascent RNA. These pauses differ in their responses to antisense oligonucleotides, pyrophosphate, GreA, and general elongation factors NusA and NusG. In contrast, substitutions in RNA polymerase that increase or decrease the rate of RNA synthesis affect both pause classes similarly. We propose that both pause classes, as well as arrest and termination, arise from a common intermediate that itself binds NTP substrate weakly. PMID- 10860977 TI - Osmoregulated ABC-transport system of Lactococcus lactis senses water stress via changes in the physical state of the membrane. AB - An osmoregulated ABC transporter (OpuA) with novel structural features has been identified that responds to water stress. This glycine betaine transport system consists of an ATP-binding/hydrolyzing subunit (OpuAA) and a protein (OpuABC) that contains both the translocator and the substrate-binding domain. The components of OpuA have been overexpressed, purified, and functionally incorporated into liposomes with an ATP-regenerating system in the vesicle lumen. A transmembrane osmotic gradient (outside hyperosmotic relative to the inside) of both ionic and nonionic compounds was able to osmotically activate OpuA in the proteoliposomal system. Hypoosmotic medium conditions inhibited the basal activity of the system. The data show that OpuAA and OpuABC are sufficient for osmoregulated transport, indicating that OpuA can act both as osmosensor and osmoregulator. Strikingly, OpuA could also be activated by low concentrations of cationic and anionic amphipaths, which interact with the membrane. This result indicates that activation by a transmembrane osmotic gradient is mediated by changes in membrane properties/protein-lipid interactions. PMID- 10860978 TI - Structural hierarchy in erythrocruorin, the giant respiratory assemblage of annelids. AB - Many annelids, including the earthworm Lumbricus terrestris, have giant cooperative respiratory proteins (molecular masses greater than 3.5 million Da) freely dissolved in the blood, rather than packaged in cells. These complexes, termed either erythrocruorins or hemoglobins, are assembled from many copies of both hemoglobin subunits and nonhemoglobin or "linker" subunits. In this paper, we present the crystal structure of Lumbricus erythrocruorin at 5.5-A resolution, which reveals a remarkable hierarchical organization of 144 oxygen-binding hemoglobin subunits and 36 nonhemoglobin linker subunits. The hemoglobin chains arrange in novel dodecameric substructures. Twelve trimeric linker complexes project triple-stranded helical coiled-coil "spokes" toward the center of the complex; interdigitation of these spokes appears crucial for stabilization. The resulting complex of linker chains forms a scaffold on which twelve hemoglobin dodecamers assemble. This structure specifies the unique, self-limited assemblage of a highly cooperative single molecule. PMID- 10860979 TI - Structure-based discovery of an organic compound that binds Bcl-2 protein and induces apoptosis of tumor cells. AB - Bcl-2 and related proteins are key regulators of apoptosis or programmed cell death implicated in human disease including cancer. We recently showed that cell permeable Bcl-2 binding peptides could induce apoptosis of human myeloid leukemia in vitro and suppress its growth in severe combined immunodeficient mice. Here we report the discovery of HA14-1, a small molecule (molecular weight = 409) and nonpeptidic ligand of a Bcl-2 surface pocket, by using a computer screening strategy based on the predicted structure of Bcl-2 protein. In vitro binding studies demonstrated the interaction of HA14-1 with this Bcl-2 surface pocket that is essential for Bcl-2 biological function. HA14-1 effectively induced apoptosis of human acute myeloid leukemia (HL-60) cells overexpressing Bcl-2 protein that was associated with the decrease in mitochondrial membrane potential and activation of caspase-9 followed by caspase-3. Cytokine response modifier A, a potent inhibitor of Fas-mediated apoptosis, did not block apoptosis induced by HA14-1. Whereas HA14-1 strongly induced the death of NIH 3T3 (Apaf-1(+/+)) cells, it had little apoptotic effect on Apaf-1-deficient (Apaf-1(-/-)) mouse embryonic fibroblast cells. These data are consistent with a mechanism by which HA14-1 induces the activation of Apaf-1 and caspases, possibly by binding to Bcl-2 protein and inhibiting its function. The discovery of this cell-permeable molecule provides a chemical probe to study Bcl-2-regulated apoptotic pathways in vivo and could lead to the development of new therapeutic agents. PMID- 10860980 TI - A second calcineurin binding site on the NFAT regulatory domain. AB - NFATc (a member of the family of nuclear factors of activated T cells) is a transcriptional factor responsible for the Ca(2+)-inducible activation of cytokine genes during the immune response. In resting T cells, NFATc is retained in the cytoplasm by a mechanism that depends on multiple phosphorylations in an N terminal regulatory domain. Physical interaction with and dephosphorylation by Ca(2+)-activated calcineurin (Cn) allows the protein to enter the nucleus, where it binds to specific sites in cytokine gene promoters. Previous studies had identified a peptide segment in NFATc that binds Cn stably. Here we report the identification of a second Cn-binding element in NFATc, which synergizes with the previously identified element. Although these sequences are conserved in all isoforms of NFAT, we find that the two sites contribute differentially to the overall affinity for Cn in an isoform-dependent manner. The regulatory domain of NFAT also was found to be entirely devoid of structure, both in the phosphorylated and unphosphorylated state. This finding suggests that the NFAT regulatory domain does not undergo phosphorylation-induced conformational switching, but instead requires partner proteins to control accessibility of the NFAT nuclear localization and nuclear export signals. PMID- 10860981 TI - A modular chitin-binding protease associated with hemocytes and hemolymph in the mosquito Anopheles gambiae. AB - Sp22D, a modular serine protease encompassing chitin binding, low density lipoprotein receptor, and scavenger receptor cysteine-rich domains, was identified by molecular cloning in the malaria vector, Anopheles gambiae. It is expressed in multiple body parts and during much of development, most intensely in hemocytes. The protein appears to be posttranslationally modified. Its integral, putatively glycosylated form is secreted in the hemolymph, whereas a smaller form potentially generated by proteolytic processing is associated with the tissues. Bacterial challenge or wounding result in low-level RNA induction, but the protein does not bind to bacteria, nor is its processing affected by infection. However, Sp22D binds to chitin with high affinity and undergoes transient changes in processing during pupal to adult metamorphosis; it may respond to exposure to naked chitin during tissue remodeling or damage. PMID- 10860982 TI - Differential ligand-dependent protein-protein interactions between nuclear receptors and a neuronal-specific cofactor. AB - Nuclear receptors are transcription factors that require multiple protein-protein interactions to regulate target gene expression. We have cloned a 27-kDa protein, termed NIX1 (neuronal interacting factor X 1), that directly binds nuclear receptors in vitro and in vivo. Protein-protein interaction between NIX1 and ligand-activated or constitutive active nuclear receptors, including retinoid related orphan receptor beta (RORbeta) (NR1F2), strictly depends on the conserved receptor C-terminal activation function 2 (AF2-D). NIX1 selectively binds retinoic acid receptor (RAR) (NR1A) and thyroid hormone receptor (TR) (NR1B) in a ligand-dependent manner, but does not interact with retinoid X receptor (RXR) (NR2B) or steroid hormone receptors. Interestingly, NIX1 down-regulates transcriptional activation by binding to ligand-bound nuclear receptors. A 39-aa domain within NIX1 was found to be necessary and sufficient for protein-protein interactions with nuclear receptors. Northern blot analysis demonstrates low abundance RNA messages only in brain and neuronal cells. In situ hybridization and immunohistochemistry revealed that NIX1 expression is restricted to the central nervous system and could be confined to neurons in the dentate gyrus of the hippocampus, the amygdala, thalamic, and hypothalamic regions. In summary, protein-protein interactions between the neuronal protein NIX1 and ligand activated nuclear receptors are both specific and selective. By suppressing receptor-mediated transcription, NIX1 implements coregulation of nuclear receptor functions in brain. PMID- 10860983 TI - Characterization of bacteriophage T4-coordinated leading- and lagging-strand synthesis on a minicircle substrate. AB - The DNA replication complex of bacteriophage T4 has been assembled as a single unit on a minicircle substrate with a replication fork that permits an independent measurement of the amount of DNA synthesis on both the leading and lagging strands. The assembled replisome consists of the T4 polymerase [gene product 43 (gp43)], clamp protein (gp45), clamp loader (gp44/62), helicase (gp41), helicase accessory factor (gp59), primase (gp61), and single-stranded DNA binding protein (gp32). We demonstrate that on the minicircle the synthesis of the leading and lagging strands are coordinated and that the C-terminal domain of the gp32 protein regulates this coordination. We show that the reconstituted replisome encompasses two coupled holoenzyme complexes and present evidence that this coupling might include a gp43 homodimer interaction. PMID- 10860984 TI - The histone deacetylase-3 complex contains nuclear receptor corepressors. AB - Acetylation and deacetylation of nucleosomal histones have profound effects on gene transcription in all eukaryotes. In humans, three highly homologous class I and four class II histone deacetylase (HDAC) enzymes have been identified to date. The class I deacetylases HDAC1 and HDAC2 are components of multisubunit complexes, one of which could associate with the nuclear hormone receptor corepressor, N-CoR. N-CoR also interacts with class II deacetylases HDAC4, HDAC5, and HDAC7. In comparison with HDAC1 and HDAC2, HDAC3 remains relatively uncharacterized, and very few proteins have been shown to interact with HDAC3. Using an affinity purification approach, we isolated an enzymatically active HDAC3 complex that contained members of the nuclear receptor corepressor family. Deletion analysis of N-CoR revealed that HDAC3 binds multiple N-CoR regions in vitro and that all of these regions are required for maximal binding in vivo. The N-CoR domains that interact with HDAC3 are distinct from those that bind other HDACs. Transient overexpression of HDAC3 and microinjection of Abs against HDAC3 showed that a component of transcriptional repression mediated by N-CoR depends on HDAC3. Interestingly, data suggest that interaction with a region of N-CoR augments the deacetylase activity of HDAC3. These results provide a possible molecular mechanism for HDAC3 regulation and argue that N-CoR is a platform in which distinct domains can interact with most of the known HDACs. PMID- 10860985 TI - Selenium-dependent metabolism of purines: A selenium-dependent purine hydroxylase and xanthine dehydrogenase were purified from Clostridium purinolyticum and characterized. AB - During purification of the selenium-dependent xanthine dehydrogenase (XDH) from Clostridium purinolyticum, another hydroxylase was uncovered that also contained selenium and exhibited similar spectral properties. This enzyme was purified to homogeneity. It uses purine, 2OH-purine, and hypoxanthine as substrates, and based on its substrate specificity, this selenoenzyme is termed purine hydroxylase (PH). The product of hydroxylation of purine by PH is xanthine. A concomitant release of selenium from the enzyme and loss of catalytic activity on treatment with cyanide indicates that selenium is essential for PH activity. Selenium-dependent XDH, also purified from C. purinolyticum, was found to be insensitive to oxygen during purification and to use both potassium ferricyanide and 2,6-dichloroindophenol as electron acceptors. Selenium is required for the xanthine-dependent reduction of 2, 6-dichloroindophenol by XDH. Kinetic analyses of both enzymes revealed that xanthine is the preferred substrate for XDH and purine and hypoxanthine are preferred by PH. This characterization of these selenium-requiring hydroxylases involved in the interconversion of purines describes an extension of the pathway for purine fermentation in the purinolytic clostridia. PMID- 10860986 TI - Homologs of the yeast Sec complex subunits Sec62p and Sec63p are abundant proteins in dog pancreas microsomes. AB - Cotranslational protein transport into dog pancreas microsomes involves the Sec61p complex plus a luminal heat shock protein 70. Posttranslational protein transport into the yeast endoplasmic reticulum (ER) involves the so-called Sec complex in the membrane, comprising a similar Sec61p subcomplex, the putative signal peptide receptor subcomplex, and the heat shock protein 40-type subunit, Sec63p, plus a luminal heat shock protein 70. Recently, human homologs of yeast proteins Sec62p and Sec63p were discovered. Here we determined the concentrations of these two membrane proteins in dog pancreas microsomes and observed that the canine homologs of yeast proteins Sec62p and Sec63p are abundant proteins, present in almost equimolar concentrations as compared with Sec61alphap monomers. Furthermore, we detected fractions of these two proteins in association with each other as well as with the Sec61p complex. The J domain of the human Sec63p was shown to interact with immunoglobulin heavy chain binding protein. Thus, the membrane of the mammalian ER contains components, known from the posttranslationally operating protein translocase in yeast. We suggest that these components are required for efficient cotranslational protein transport into the mammalian ER as well as for other transport processes. PMID- 10860987 TI - Measuring the rate of intramolecular contact formation in polypeptides. AB - Formation of a specific contact between two residues of a polypeptide chain is an important elementary process in protein folding. Here we describe a method for studying contact formation between tryptophan and cysteine based on measurements of the lifetime of the tryptophan triplet state. With tryptophan at one end of a flexible peptide and cysteine at the other, the triplet decay rate is identical to the rate of quenching by cysteine. We show that this rate is also close to the diffusion-limited rate of contact formation. The length dependence of this end-to end contact rate was studied in a series of Cys-(Ala-Gly-Gln)(k)-Trp peptides, with k varying from 1 to 6. The rate decreases from approximately 1/(40 ns) for k = 1 to approximately 1/(140 ns) for k = 6, approaching the length dependence expected for a random coil (n(-3/2)) for the longest peptides. PMID- 10860988 TI - Interference fine structure and sarcomere length dependence of the axial x-ray pattern from active single muscle fibers. AB - Axial x-ray diffraction patterns from single intact fibers of frog skeletal muscle were recorded by using a highly collimated x-ray beam at the European Synchrotron Radiation Facility. During isometric contraction at sarcomere lengths 2.2-3.2 microm, the M3 x-ray reflection, associated with the repeat of myosin heads along the filaments, was resolved into two peaks. The total M3 intensity decreased linearly with increasing sarcomere length and was directly proportional to the degree of overlap between myosin and actin filaments, showing that it comes from myosin heads in the overlap region. The separation between the M3 peaks was smaller at longer sarcomere length and was quantitatively explained by x-ray interference between myosin heads in the two overlap regions of each sarcomere. The relative intensity of the M3 peaks was independent of sarcomere length, showing that the axial periodicities of the nonoverlap and overlap regions of the myosin filament have the same value, 14.57 nm, during active contraction. In resting fibers the periodicity is 14.34 nm, so muscle activation produces a change in myosin filament structure in the nonoverlap as well as the overlap part of the filament. The results establish x-ray interferometry as a new tool for studying the motions of myosin heads during muscle contraction with unprecedented spatial resolution. PMID- 10860989 TI - Real-time light-driven dynamics of the fluorescence emission in single green fluorescent protein molecules. AB - Real-time single-molecule fluorescence detection using confocal and near-field scanning optical microscopy has been applied to elucidate the nature of the "on off" blinking observed in the Ser-65 --> Thr (S65T) mutant of the green fluorescent protein (GFP). Fluorescence time traces as a function of the excitation intensity, with a time resolution of 100 micros and observation times up to 65 s, reveal the existence of a nonemissive state responsible for the long dark intervals in the GFP. We find that excitation intensity has a dramatic effect on the blinking. Whereas the time during which the fluorescence is on becomes shorter as the intensity is increased, the off-times are independent of excitation intensity. Statistical analysis of the on- and off-times renders a characteristic off-time of 1.6 +/- 0.2 s and allows us to calculate a transition yield of approximately 0.5 x 10(-5) from the emissive to the nonemissive state. The saturation excitation intensity at which on- and off-times are equal is approximately 1.5 kW/cm(2). On the basis of the single-molecule data we calculate an absorption cross section of 6.5 x 10(-17) cm(2) for the S65T mutant. These results have important implications for the use of the GFP to follow dynamic processes in time at the single-molecular level. PMID- 10860990 TI - Native topology determines force-induced unfolding pathways in globular proteins. AB - Single-molecule manipulation techniques reveal that stretching unravels individually folded domains in the muscle protein titin and the extracellular matrix protein tenascin. These elastic proteins contain tandem repeats of folded domains with beta-sandwich architecture. Herein, we propose by stretching two model sequences (S1 and S2) with four-stranded beta-barrel topology that unfolding forces and pathways in folded domains can be predicted by using only the structure of the native state. Thermal refolding of S1 and S2 in the absence of force proceeds in an all-or-none fashion. In contrast, phase diagrams in the force-temperature (f,T) plane and steered Langevin dynamics studies of these sequences, which differ in the native registry of the strands, show that S1 unfolds in an allor-none fashion, whereas unfolding of S2 occurs via an obligatory intermediate. Force-induced unfolding is determined by the native topology. After proving that the simulation results for S1 and S2 can be calculated by using native topology alone, we predict the order of unfolding events in Ig domain (Ig27) and two fibronectin III type domains ((9)FnIII and (10)FnIII). The calculated unfolding pathways for these proteins, the location of the transition states, and the pulling speed dependence of the unfolding forces reflect the differences in the way the strands are arranged in the native states. We also predict the mechanisms of force-induced unfolding of the coiled-coil spectrin (a three-helix bundle protein) for all 20 structures deposited in the Protein Data Bank. Our approach suggests a natural way to measure the phase diagram in the (f,C) plane, where C is the concentration of denaturants. PMID- 10860991 TI - Chimeric green fluorescent protein-aequorin as bioluminescent Ca2+ reporters at the single-cell level. AB - Monitoring calcium fluxes in real time could help to understand the development, the plasticity, and the functioning of the central nervous system. In jellyfish, the chemiluminescent calcium binding aequorin protein is associated with the green fluorescent protein and a green bioluminescent signal is emitted upon Ca(2+) stimulation. We decided to use this chemiluminescence resonance energy transfer between the two molecules. Calcium-sensitive bioluminescent reporter genes have been constructed by fusing green fluorescent protein and aequorin, resulting in much more light being emitted. Chemiluminescent and fluorescent activities of these fusion proteins have been assessed in mammalian cells. Cytosolic Ca(2+) increases were imaged at the single-cell level with a cooled intensified charge-coupled device camera. This bifunctional reporter gene should allow the investigation of calcium activities in neuronal networks and in specific subcellular compartments in transgenic animals. PMID- 10860992 TI - Interferon regulatory factor subcellular localization is determined by a bipartite nuclear localization signal in the DNA-binding domain and interaction with cytoplasmic retention factors. AB - The transduction of type I interferon signals to the nucleus relies on activation of a protein complex, ISGF3, involving two signal transducers and activators of transcription (STAT) proteins, STAT1 and STAT2, and the interferon (IFN) regulatory factor (IRF) protein, p48/ISGF3gamma. The STAT subunits are cytoplasmically localized in unstimulated cells and rapidly translocate to the nucleus of IFN-stimulated cells, but the p48/ISGF3gamma protein is found in both the nucleus and the cytoplasm, regardless of IFN stimulation. Here, we demonstrate that p48 is efficiently and constitutively targeted to the nucleus. Analysis of the subcellular distribution of green fluorescent protein-p48 fragments indicates that p48 contains a bipartite nuclear retention signal within its amino-terminal DNA-binding domain. This signal is preserved in two other IRF proteins involved in immune responses, ICSBP and IRF4. Mutations to clustered basic residues within amino acids 50-100 of p48 or IRF4 disrupt their nuclear accumulation, and DNA-binding ability is not required for nuclear targeting. This is the only example of a nuclear localization signal for any ISGF3 component and assigns a second function to the IRF DNA-binding domain. We also demonstrate that the nuclear distribution of p48 is dramatically altered by coexpression of the STAT2 protein, indicating that STAT2 forms a cytoplasmic complex with p48, overriding the intrinsic p48 nuclear targeting. Retention by STAT2 may serve to regulate the activity of free p48 and/or guarantee that cytoplasmic pools of preassociated STAT2:p48 are available for rapid activation of the IFN response. These findings suggest that analogous mechanisms may exist for regulating the distribution of other IRF proteins. PMID- 10860993 TI - huASH1 protein, a putative transcription factor encoded by a human homologue of the Drosophila ash1 gene, localizes to both nuclei and cell-cell tight junctions. AB - During animal development, regions of the embryo become committed to position specific identities, which are determined by spatially restricted expression of Hox/homeotic genes. This expression pattern is initially established by the activity of the segmentation genes and is subsequently maintained during the proliferative stage through the action of transcription factors encoded by the trithorax (trx) and Polycomb (Pc) groups of genes. trithorax (trx)and ash1 (absent, small, or homeotic 1) are members of the Drosophila trx group. Their products are associated with chromosomes and are believed to activate transcription of target genes through chromatin remodeling. Recently, we reported molecular studies indicating that TRX and ASH1 proteins act in concert to bind simultaneously to response elements located at close proximity within the same set of target genes. Extension of these and other studies to mammalian systems required identification and cloning of the mammalian homologue of ash1 (the mammalian homologue of trx, ALL-1, was previously cloned). We have identified a human expressed sequence tag (EST) clone with similarity to the SET domain of Drosophila ASH1, and used it to clone the human gene. huASH1 resides at chromosomal band 1q21. The gene is expressed in multiple tissues as an approximately 10.5-kb transcript and encodes a protein of 2962 residues. The protein contains a SET domain, a PHD finger, four AT hooks, and a region with homology to the bromodomain. The last region is not present in Drosophila ASH1, and as such might confer to the human protein a unique additional function. Using several anti-huASH1 Ab for immunostaining of cultured cells, we found that the protein is distributed in intranuclear speckles, and unexpectedly also in intercellular junctions. Double-immunofluorescence labeling of huASH1 and several junctional proteins localized the huASH1 protein into tight junctions. The significance of huASH1 dual location is discussed. In particular, we consider the possibility that translocation of the protein between the junctional membrane and the nucleus may be involved in adhesion-mediated signaling. PMID- 10860994 TI - Identification and characterization of a p53 homologue in Drosophila melanogaster. AB - The tumor suppressor gene p53 in mammalian cells plays a critical role in safeguarding the integrity of genome. It functions as a sequence-specific transcription factor. Upon activation by a variety of cellular stresses, p53 transactivates downstream target genes, through which it regulates cell cycle and apoptosis. However, little is known about p53 in invertebrates. Here we report the identification and characterization of a Drosophila p53 homologue gene, dp53. dp53 encodes a 385-amino acid protein with significant homology to human p53 (hp53) in the region of the DNA-binding domain, and to a lesser extent the tetramerization domain. Purified dp53 DNA-binding domain protein was shown to bind to the consensus hp53-binding site by gel mobility analysis. In transient transfection assays, expression of dp53 in Schneider cells transcriptionally activated promoters that contained consensus hp53-responsive elements. Moreover, a mutant dp53 (Arg-155 to His-155), like its hp53 counterpart mutant, exerted a dominant-negative effect on transactivation. Ectopic expression of dp53 in Drosophila eye disk caused cell death and led to a rough eye phenotype. dp53 is expressed throughout the development of Drosophila with highest expression levels in early embryogenesis, which has a maternal component. Consistent with this, dp53 RNA levels were high in the nurse cells of the ovary. It appears that p53 is structurally and functionally conserved from flies to mammals. Drosophila will provide a useful genetic system to the further study of the p53 network. PMID- 10860995 TI - Inhibition of sonic hedgehog autoprocessing in cultured mammalian cells by sterol deprivation. AB - Sonic hedgehog (Shh) is a signaling molecule that is important for defining patterning in the developing vertebrate central nervous system. After translation, Shh autoproteolyzes and covalently attaches cholesterol to the newly formed carboxyl terminus, a modification crucial for normal Shh signaling. Presented here is evidence that acute severe sterol deprivation in cultured Chinese hamster ovary cells expressing mouse Shh (mShh) inhibits autoprocessing of the protein. These conditions allowed the first detailed kinetic analysis of mShh autoprocessing and turnover rates revealing that cells rapidly degrade both precursor and mature mShh regardless of sterol content and sterol deprivation increases the rate of precursor degradation. Inhibition of mShh autoprocessing also allowed the determination of the subcellular localization of mShh precursor which accumulates in a pre-medial Golgi intracellular compartment. Finally, the precursor form of mShh that results from autoprocessing inhibition appears to accumulate as an amide rather than a stable thioester. PMID- 10860996 TI - GLUT8 is a glucose transporter responsible for insulin-stimulated glucose uptake in the blastocyst. AB - Mammalian preimplantation blastocysts exhibit insulin-stimulated glucose uptake despite the absence of the only known insulin-regulated transporter, GLUT4. We describe a previously unidentified member of the mammalian facilitative GLUT superfamily that exhibits approximately 20-25% identity with other murine facilitative GLUTs. Insulin induces a change in the intracellular localization of this protein, which translates into increased glucose uptake into the blastocyst, a process that is inhibited by antisense oligoprobes. Presence of this transporter may be necessary for successful blastocyst development, fuel metabolism, and subsequent implantation. Moreover, the existence of an alternative transporter may explain examples in other tissues of insulin regulated glucose transport in the absence of GLUT4. PMID- 10860997 TI - Crosstalk pathway for inhibition of glucocorticoid-induced apoptosis by T cell receptor signaling. AB - Activation of the glucocorticoid receptor (GR) triggers apoptosis in T cells. However, activation of the T cell antigen receptor (TCR) blocks glucocorticoid induced apoptosis, implying functional crosstalk between these two distinct signaling systems. By reconstructing or selectively blocking TCR-stimulated signaling pathways, we show here that TCR activation of the mitogen-activated protein kinase kinase/extracellular signal regulated kinase (MEK/ERK) cascade via Ras is necessary and sufficient to inhibit GR-mediated death in immortalized T and thymocyte cell lines and in primary T cells. Moreover, we found that activation of various pathway components (TCR, Ras, MEK1) altered the transcriptional regulatory activity of GR. In contrast, phosphatidylinositol 3 kinase and Akt, which down-regulate other lymphocyte apoptosis pathways, did not inhibit glucocorticoid-induced apoptosis. Our findings, which link signaling from the TCR cell surface receptor to that from the GR intracellular receptor, demonstrate the importance of the integration of signal transduction pathways in defining regulatory circuits. Because the TCR/Ras/MEK pathway has been shown previously to be essential for positive selection of thymocytes, the TCR/Ras/MEK signaling to GR crosstalk described herein may affect T cell development and homeostasis. PMID- 10860998 TI - A complex of iron and nucleic acid catabolites is a signal that triggers differentiation in a freshwater protozoan. AB - The polymorphic ciliated protozoan Tetrahymena vorax can undergo differentiation from the microstomal form, which normally feeds on bacteria and other particulate matter, into the macrostomal cell type, which is capable of ingesting prey ciliates. The process is triggered by exposure of the microstome to an inducer contained in stomatin, an exudate of the prey. To establish the identity of the signal, stomatin was fractionated by combinations of cation exchange, HPLC, and TLC, and the fractions were assayed for biological activity. Although no single active fraction of purified inducer was obtained, all fractions with activity contained ferrous iron and the nucleic acid catabolites hypoxanthine (6 oxypurine) and uracil (2, 4-dioxopyrimidine), probably in a chelated form. The activity of synthetic complexes containing these three components is equivalent to stomatin. These results indicate a role for ferrous iron and its potential in chelated form to signal differentiation in certain protozoa and, perhaps, in other organisms as well. PMID- 10860999 TI - DER signaling restricts the boundaries of the wing field during Drosophila development. AB - Arthropod and vertebrate limbs develop from secondary embryonic fields. In insects, the wing imaginal disk is subdivided early in development into the wing and notum subfields. The activity of the Wingless protein is fundamental for this subdivision and seems to be the first element of the hierarchy of regulatory genes promoting wing formation. Drosophila epidermal growth factor receptor (DER) signaling has many functions in fly development. Here we show that antagonizing DER signaling during the second larval instar leads to notum to wing transformations and wing mirror-image duplications. DER signaling is necessary for confining the wing subregion in the developing wing disk and for the specification of posterior identity. To do so, DER signaling acts by restricting the expression of Wingless to the dorsal-posterior quadrant of wing discs, suppressing wing-organizing activities, and by cooperating in the maintenance of Engrailed expression in posterior compartment cells. PMID- 10861000 TI - Circadian rise in maternal glucocorticoid prevents pulmonary dysplasia in fetal mice with adrenal insufficiency. AB - The hypothalamic-pituitary-adrenal (HPA) axis, including hypothalamic corticotropin-releasing hormone (CRH) and pituitary corticotropin, is one of the first endocrine systems to develop during fetal life, probably because glucocorticoid secretion is necessary for the maturation of many essential fetal organs. Consistent with this, pregnant mice with an inactivating mutation in the Crh gene deliver CRH-deficient offspring that die at birth with dysplastic lungs, which can be prevented by prenatal maternal glucocorticoid treatment. But children lacking the ability to synthesize cortisol (because of various genetic defects in adrenal gland development or steroidogenesis) are not born with respiratory insufficiency or abnormal lung development, suggesting that the transfer of maternal glucocorticoid across the placenta might promote fetal organ maturation in the absence of fetal glucocorticoid production. We used pregnant mice with a normal HPA axis carrying fetuses with CRH deficiency to characterize the relative contributions of the fetal and maternal adrenal to the activity of the fetal HPA axis, and related these findings to fetal lung development. We found that in the presence of fetal adrenal insufficiency, normal fetal lung development is maintained by the transfer of maternal glucocorticoid to the fetus, specifically during the circadian peak in maternal glucocorticoid secretion. PMID- 10861001 TI - Isolation and characterization of BEN, a member of the TFII-I family of DNA binding proteins containing distinct helix-loop-helix domains. AB - The transcriptional regulation of the Hoxc8 gene is controlled during early mouse embryogenesis by an enhanceosome-like control region, termed the early enhancer (EE), located 3 kb upstream from the Hoxc8 translation start site. The EE is involved in establishing the posterior expression pattern of Hoxc8 at embryonic day (E) 8.5-9. 0. Genetic and biochemical data have shown that nuclear factors interact with this region in a sequence-specific manner. We have used a yeast one hybrid screen in a search for transcription factors that bind to EE motifs and have isolated a novel murine DNA-binding protein, termed BEN (binding factor for early enhancer). The ORF of BEN encodes a protein of 1072 amino acids and contains six helix-loop-helix domains, a hydrophobic leucine zipper-like motif, and a serine-rich repeat. The murine BEN gene is structurally similar to the human gene TFII-I in that both genes encode unique 95-amino acid long helix loop/span-helix domains. The BEN gene produces several major transcripts (3.6, 4.4, and 5.9 kb) present in most adult tissues and shows discrete spatial and temporal domains of expression in areas of epithelial-mesenchymal interaction during mouse embryogenesis from E9.5 to E12.5. Several BEN-encoded polypeptides of different sizes ranging from 165 to 40 kDa were identified by Western blot analysis using BEN-specific polyclonal Abs. We propose, on the bases of sequence homology, that BEN is the mouse ortholog of the recently described human gene, WBSCR11, known also as GTF2IRD1, GTF3, Cream1, and MusTRD1. This gene is deleted hemizygously in individuals with Williams Syndrome, an autosomal dominant genetic condition characterized by complex physical, cognitive, and behavioral traits resulting from a perturbed developmental process. PMID- 10861002 TI - The multitype zinc-finger protein U-shaped functions in heart cell specification in the Drosophila embryo. AB - Multitype zinc-finger proteins of the Friend of GATA/U-shaped (Ush) class function as transcriptional regulators of gene expression through their modulation of GATA factor activity. To better understand intrinsic properties of these proteins, we investigated the expression and function of the ush gene during Drosophila embryogenesis. ush is dynamically expressed in the embryo, including several cell types present within the mesoderm. The gene is active in the cardiogenic mesoderm, and a loss of function results in an overproduction of both cardial and pericardial cells, indicating a requirement for the gene in the formation of these distinct cardiac cell types. Conversely, ectopic expression of ush results in a decrease in the number of cardioblasts in the heart and the inhibition of a cardial cell enhancer normally regulated by the synergistic activity of the Pannier and Tinman cardiogenic factors. These findings suggest that, similar to its known function in thoracic bristle patterning, Ush functions in the control of heart cell specification through its modulation of Pannier transcriptional activity. ush is also required for mesodermal cell migration early in embryogenesis, where it shows a genetic interaction with the Heartless fibroblast growth factor receptor gene. Taken together, these results demonstrate a critical role for the Ush transcriptional regulator in several diverse processes of mesoderm differentiation and heart formation. PMID- 10861003 TI - Population genetic implications from sequence variation in four Y chromosome genes. AB - Some insight into human evolution has been gained from the sequencing of four Y chromosome genes. Primary genomic sequencing determined gene SMCY to be composed of 27 exons that comprise 4,620 bp of coding sequence. The unfinished sequencing of the 5' portion of gene UTY1 was completed by primer walking, and a total of 20 exons were found. By using denaturing HPLC, these two genes, as well as DBY and DFFRY, were screened for polymorphic sites in 53-72 representatives of the five continents. A total of 98 variants were found, yielding nucleotide diversity estimates of 2.45 x 10(-5), 5. 07 x 10(-5), and 8.54 x 10(-5) for the coding regions of SMCY, DFFRY, and UTY1, respectively, with no variant having been observed in DBY. In agreement with most autosomal genes, diversity estimates for the noncoding regions were about 2- to 3-fold higher and ranged from 9. 16 x 10( 5) to 14.2 x 10(-5) for the four genes. Analysis of the frequencies of derived alleles for all four genes showed that they more closely fit the expectation of a Luria-Delbruck distribution than a distribution expected under a constant population size model, providing evidence for exponential population growth. Pairwise nucleotide mismatch distributions date the occurrence of population expansion to approximately 28,000 years ago. This estimate is in accord with the spread of Aurignacian technology and the disappearance of the Neanderthals. PMID- 10861004 TI - Recent common ancestry of human Y chromosomes: evidence from DNA sequence data. AB - We consider a data set of DNA sequence variation at three Y chromosome genes (SMCY, DBY, and DFFRY) in a worldwide sample of human Y chromosomes. Between 53 and 70 chromosomes were fully screened for sequence variation at each locus by using the method of denaturing high-performance liquid chromatography. The sum of the lengths of the three genes is 64,120 bp. We have used these data to study the ancestral genealogy of human Y chromosomes. In particular, we focused on estimating the expected time to the most recent common ancestor and the expected ages of certain mutations with interesting geographic distributions. Although the geographic structure of the inferred haplotype tree is reminiscent of that obtained for other loci (the root is in Africa, and most of the oldest non African lineages are Asian), the expected time to the most recent common ancestor is remarkably short, on the order of 50,000 years. Thus, although previous studies have noted that Y chromosome variation shows extreme geographic structure, we estimate that the spread of Y chromosomes out of Africa is much more recent than previously was thought. We also show that our data indicate substantial population growth in the effective number of human Y chromosomes. PMID- 10861005 TI - Ultraviolet pigments in birds evolved from violet pigments by a single amino acid change. AB - UV vision has profound effects on the evolution of organisms by affecting such behaviors as mating preference and foraging strategies. Despite its importance, the molecular basis of UV vision is not known. Here, we have transformed the zebra finch UV pigment into a violet pigment by incorporating one amino acid change, C84S. By incorporating the reverse mutations, we have also constructed UV pigments from the orthologous violet pigments of the pigeon and chicken. These results and comparative amino acid sequence analyses of the pigments in vertebrates demonstrate that many avian species have achieved their UV vision by S84C. PMID- 10861006 TI - Derivation of the relationship between neutral mutation and fixation solely from the definition of selective neutrality. AB - A mutation whose fixation is independent of natural selection is termed a neutral mutation. Therefore selective neutrality of a mutation can be defined by independence of its fixation from natural selection. By the population genetic approach, Kimura [Kimura, M. (1962) Genetics 47, 713-719] predicted that the probability of fixation of a neutral mutation (u) is equal to the frequency of the new allele at the start (p). The approach traced the temporal sequence of the fixation process, and the prediction was obtained by assuming the selective equality of neutral mutant and wild-type alleles during the fixation process. The prediction, however, has not been verified by observation. In the present study, I search for the mathematical equation that represents the definition of selective neutrality. Because the definition concerns only mutation and fixation, an ideal approach should deal only with these and not with the intervening process of fixation. The approach begins by analysis of the state of fixation of a neutral mutation, and its relation with the initial state is deduced logically from the definition. The approach shows that the equality of the alleles during the fixation process is equivalent to the equality of probability of their ultimate fixation in a steady state. Both are manifestations of the definition of selective neutrality. Then, solely from this dual nature of the definition, the equality between u and p is derived directly. Therefore, the definition of selective neutrality can be represented by the equation u = p. PMID- 10861007 TI - Transposon diversity in Arabidopsis thaliana. AB - Recent availability of extensive genome sequence information offers new opportunities to analyze genome organization, including transposon diversity and accumulation, at a level of resolution that was previously unattainable. In this report, we used sequence similarity search and analysis protocols to perform a fine-scale analysis of a large sample ( approximately 17.2 Mb) of the Arabidopsis thaliana (Columbia) genome for transposons. Consistent with previous studies, we report that the A. thaliana genome harbors diverse representatives of most known superfamilies of transposons. However, our survey reveals a higher density of transposons of which over one-fourth could be classified into a single novel transposon family designated as Basho, which appears unrelated to any previously known superfamily. We have also identified putative transposase-coding ORFs for miniature inverted-repeat transposable elements (MITEs), providing clues into the mechanism of mobility and origins of the most abundant transposons associated with plant genes. In addition, we provide evidence that most mined transposons have a clear distribution preference for A + T-rich sequences and show that structural variation for many mined transposons is partly due to interelement recombination. Taken together, these findings further underscore the complexity of transposons within the compact genome of A. thaliana. PMID- 10861008 TI - Chromosome instability contributes to loss of heterozygosity in mice lacking p53. AB - The p53 tumor suppressor protein participates in multiple cellular processes including cell cycle checkpoints and programmed cell death. In cell lines, loss of p53 function is associated with increased genetic instability including aneuploidy, gene amplification, and point mutation. Although similar genetic instability often accompanies the progression of malignancy in tumors, its role in tumor initiation in normal cells is not clear. To study whether or not loss of p53 leads to genetic instability in normal cells in vivo, we have examined mechanisms of loss of heterozygosity (LOH) at the Aprt (adenine phosphoribsyltransferase) and flanking loci in normal fibroblasts and T lymphocytes of p53-deficient mice. Somatic cell variants that arose in vivo as a consequence of genetic or epigenetic alterations abolishing Aprt function were selected and expanded in vitro by virtue of their resistance to 2,6-diaminopurine (DAP). We observed that p53 null mice produced about three times as many DAP resistant fibroblast colonies than wild-type mice, but the frequency of DAP resistant T lymphocyte colonies was not significantly changed. Mitotic recombination, but not point mutation, partly accounted for the increase in the frequency of DAP-resistant fibroblasts. Most significantly, chromosome loss/duplication and interstitial deletion, which were extremely rare events in the wild-type mice, represented a significant proportion of LOH events in both fibroblasts and T lymphocytes of p53 null mice. Also, increased interstitial deletion was observed in fibroblasts of p53 heterozygous mice. These data suggest that increased genetic variation, including chromosome instability, starts at the initiation stage of tumorigenesis when functional p53 is absent or reduced. PMID- 10861009 TI - G protein-coupled receptor kinase-5 regulates thrombin-activated signaling in endothelial cells. AB - We studied the function of G protein-coupled receptor kinases (GRKs) in the regulation of thrombin-activated signaling in endothelial cells. GRK2, GRK5, and GRK6 isoforms were expressed predominantly in endothelial cells. The function of these isoforms was studied by expressing wild-type and dominant negative (dn) mutants in endothelial cells. We determined the responses to thrombin, which activates intracellular signaling in endothelial cells by cleaving the NH(2) terminus of the G protein-coupled proteinase-activated receptor-1 (PAR-1). We measured changes in phosphoinositide hydrolysis and intracellular Ca(2+) concentration ([Ca(2+)](i)) in response to thrombin as well as the state of endothelial activation. In the latter studies, the transendothelial monolayer electrical resistance, a measure of the loss of endothelial barrier function, was measured in real time. Of the three isoforms, GRK5 overexpression was selective in markedly reducing the thrombin-activated phosphoinositide hydrolysis and increased [Ca(2+)](i). GRK5 overexpression also inhibited the thrombin-induced decrease in endothelial monolayer resistance by 75%. These effects of GRK5 overexpression occurred in association with the specific increase in the thrombin induced phosphorylation of PAR-1. In contrast to the effects of GRK5 overexpression, the expression of the dn-GRK5 mutant produced a long-lived increase in [Ca(2+)](i) in response to thrombin, whereas dn-GRK2 had no effect. These results indicate the crucial role of the GRK5 isoform in the mechanism of thrombin-induced desensitization of PAR-1 in endothelial cells. PMID- 10861010 TI - T cell immunity to copolymer 1 confers neuroprotection on the damaged optic nerve: possible therapy for optic neuropathies. AB - We recently reported that the posttraumatic spread of degeneration in the damaged optic nerve can be attenuated by the adoptive transfer of autoimmune T cells specific to myelin basic protein. However, it would be desirable to obtain immune neuroprotection free of any possible autoimmune disease. In an attempt to obtain disease-free immune neuroprotection, we used the synthetic four-amino acid polymer copolymer 1 (Cop-1), which is known not to be encephalitogenic despite its cross-reactivity with myelin basic protein. We show here that active immunization with Cop-1 administered in adjuvant, as well as adoptive transfer of T cells reactive to Cop-1, can inhibit the progression of secondary degeneration after crush injury of the rat optic nerve. These results have implications for the treatment of optic neuropathies. PMID- 10861011 TI - Multiple sclerosis: comparison of copolymer-1- reactive T cell lines from treated and untreated subjects reveals cytokine shift from T helper 1 to T helper 2 cells. AB - Copolymer 1 (COP), a standardized mixture of synthetic polypeptides consisting of l-glutamic acid, l-lysine, l-alanine, and l-tyrosine, has beneficial effects in multiple sclerosis and experimental autoimmune encephalomyelitis. We selected a panel of 721 COP-reactive T cell lines (TCL) from the blood of COP-treated and untreated multiple sclerosis patients and from healthy donors by using the split well cloning technique. All TCL selected with COP proliferated in response to COP but not to myelin basic protein (MBP). Conversely, 31 control TCL selected with MBP proliferated in response to MBP but not to COP. We used intracellular double immunofluorescence flow cytometry for quantitative analysis of cytokine production (IL-4, IFN-gamma) by the TCL. The majority of the COP-reactive TCL from untreated multiple sclerosis patients and normal donors predominantly produced IFN-gamma and, accordingly, were classified as T helper 1 cells (TH1). In contrast, the majority of the COP-reactive TCL from COP-treated patients predominantly (but not exclusively) produced IL-4-i.e., were TH2 (P < 0.05 as assessed by using a suitable preference intensity index). Longitudinal analyses revealed that the cytokine profile of COP-reactive TCL tends to shift from TH1 to TH2 during treatment. Interestingly, although there was no proliferative cross reaction, about 10% of the COP-reactive TCL responded to MBP by secretion of small amounts of IL-4 or IFN-gamma, depending on the cytokine profile of the TCL. These results are consistent with a protective effect of COP-reactive TH2 cells. It is hypothesized that these cells are activated by COP in the periphery, migrate into the central nervous system, and produce immunomodulatory cytokines after local recognition of MBP. PMID- 10861012 TI - Inhibition of CD40 signaling limits evolution of established atherosclerosis in mice. AB - Interruption of inflammatory pathways may provide a novel approach to the therapy of atherosclerosis. Recently, we and others have implicated the immune mediator dyad CD40/CD40L (CD40 ligand), which is expressed on endothelial and smooth muscle cells, macrophages, and T lymphocytes within human atherosclerotic lesions, in aspects of atherogenesis and the acute coronary syndromes, including regulation of matrix metalloproteinases, procoagulant activity, cytokines, etc. In vivo, interruption of CD40 signaling reduced the initiation and early phases of atheroma formation in hypercholesterolemic mice. However, whether interruption of CD40 signaling can retard the progression or even regress established lesions remains unknown. We report here that anti-CD40L antibody treatment of randomly assigned low-density lipoprotein receptor-deficient mice during the second half of a 26-week regimen of high-cholesterol diet did not regress, but did significantly reduce further evolution of established atherosclerotic lesions within the aortic arch and particularly the thoracic and abdominal aorta, as compared with control treatment (application of rat-IgG or saline; 13 weeks, continued high-cholesterol diet). In addition to limiting lesion progression, anti-CD40L treatment changed the composition of atheroma in manners thought to favor plaque stability, e.g., reduced relative content of macrophages and lipid, as well as increased relative content of smooth muscle cells and collagen. These data implicate CD40/CD40L as crucial mediators not only in the initial events of atherogenesis but also during the evolution of established atheroma. This study lends further support to the importance of this specific inflammatory signaling pathway in atherosclerosis and its complications. PMID- 10861013 TI - Both early and delayed anti-CD40L antibody treatment induces a stable plaque phenotype. AB - In the present study, we investigated the role of the CD40L-CD40 pathway in a model of progressive atherosclerosis. ApoE-/- mice were treated with an anti CD40L antibody or a control antibody for 12 wk. Antibody treatment started early (age 5 wk) or was delayed until after the establishment of atherosclerosis (age 17 wk). In both the early and delayed treatment groups, anti-CD40L antibody did not decrease plaque area or inhibit lesion initiation or age-related increase in lesion area. The morphology of initial lesions was not affected, except for a decrease in T-lymphocyte content. Effects of anti-CD40L antibody treatment on the morphology of advanced lesions were pronounced. In both the early and delayed treatment groups, T-lymphocyte content was significantly decreased. Furthermore, a pronounced increase in collagen content, vascular smooth muscle cell/myofibroblast content, and fibrous cap thickness was observed. In the delayed treatment group, a decrease in lipid core and macrophage content occurred. Interestingly, advanced lesions of anti-CD40L antibody-treated mice exhibited an increased transforming growth factor beta1 immunoreactivity, especially in macrophages. In conclusion, both early and delayed treatment with an anti-CD40L antibody do not affect atherosclerotic lesion initiation but do result in the development of a lipid-poor collagen-rich stable plaque phenotype. Furthermore, delayed treatment with anti-CD40L antibody can transform the lesion profile from a lipid-rich to a lipid-poor collagen-rich phenotype. Postulated mechanisms of this effect on plaque phenotype are the down-regulation of proinflammatory pathways and up-regulation of collagen-promoting factors like transforming growth factor beta. PMID- 10861014 TI - Tumor selective G2/M cell cycle arrest and apoptosis of epithelial and hematological malignancies by BBL22, a benzazepine. AB - Two distinct benzodiazepine binding sites have been identified, (i) a central site restricted to brain and (ii) a ubiquitously expressed mitochondrial binding site, the so-called peripheral-type benzodiazepine receptor (PBR). In this paper, we show that a benzazepine referred to as BBL22 (2-amino 9-chloro-7-(2 fluorophenyl)-5H-pyrimidol[5,4-d][2]benzazepine), which is classified as a PBR ligand based on structure, induces arrest in G(2)/M phase of the cell cycle in human tumor cell lines of both epithelial and hematopoietic cellular origin. After G(2)/M arrest, several tumor types, notably prostate and certain breast cancer lines exhibited significant apoptosis. Ideally, cancer therapies should selectively target tumor cells while sparing normal cell counterparts. BBL22 exhibited such selectivity, as it did not affect the growth and survival of nonmalignant breast and prostate epithelial lines. Moreover, BBL22 demonstrated structural requirements for this selective antitumor activity as 11 structurally related PBR ligands, including high-affinity ligands Ro5-4864 and PK11195, failed to induce tumor cell growth arrest or apoptosis. The in vivo antitumor activity of BBL22 was examined in a human xenograft model of androgen-independent prostate cancer where BBL22 significantly reduced the growth of PC3 prostate tumors without eliciting overt toxicity. Identification of BBL22 represents a tumor selective therapeutic strategy for a variety of human tumors. PMID- 10861015 TI - Targeted disruption of an erythrocyte binding antigen in Plasmodium falciparum is associated with a switch toward a sialic acid-independent pathway of invasion. AB - Erythrocyte invasion by Plasmodium requires molecules present both on the merozoite surface and within the specialized organelles of the apical complex. The Plasmodium erythrocyte binding protein family includes the Plasmodium falciparum sialic acid-binding protein, EBA-175 (erythrocyte binding antigen 175), which binds sialic acid present on glycophorin A of human erythrocytes. We address the role of the conserved 3'-cysteine rich region, the transmembrane, and cytoplasmic domains through targeted gene disruption. Truncation of EBA-175 had no measurable effect on either the level of EBA-175 protein expression or its subcellular localization. Similarly, there appears to be no impairment in the ability of soluble EBA-175 to be released into the culture supernatant after schizont rupture. Additionally, the 3'-cys rich region, transmembrane, and cytoplasmic domains of EBA-175 are apparently non-essential for merozoite invasion. In contrast, erythrocyte invasion via the EBA-175/glycophorin A route appears to have been disrupted to such a degree that the mutant lines have undergone a stable switch in invasion phenotype. As such, EBA-175 appears to have been functionally inactivated within the truncation mutants. The sialic acid independent invasion pathway within the mutant parasites accounts for approximately 85% of invasion into normal erythrocytes. These data demonstrate the ability of P. falciparum to utilize alternate pathways for invasion of red blood cells, a property that most likely provides a substantial survival advantage in terms of overcoming host receptor heterogeneity and/or immune pressure. PMID- 10861016 TI - AML1/ETO-expressing nonleukemic stem cells in acute myelogenous leukemia with 8;21 chromosomal translocation. AB - Leukemia-specific AML1/ETO transcripts are detectable in most patients with t(8;21) acute myelogenous leukemia (AML) in long-term remission. To understand the inconsistency between the clinical cure and the presence of "residual disease" at a molecular level, we separated and identified the cells expressing AML1/ETO by phenotype and function. Here we demonstrate that AML1/ETO transcripts are present in a fraction of stem cells, monocytes, and B cells in remission marrow, and in a fraction of B cells in leukemic marrow, but not in T cells. AML1/ETO transcripts also were demonstrated in a fraction of colony-forming cells of erythroid, granulocyte-macrophage, and/or megakaryocyte lineages in both leukemic and remission marrow. These data strongly suggest that the acquisition of the t(8;21) occurs at the level of stem cells capable of differentiating into B cells as well as all myeloid lineages, and that a fraction of the AML1/ETO expressing stem cells undergo additional oncogenic event(s) that ultimately leads to transformation into AML. PMID- 10861017 TI - A conserved amino acid sequence directing intracellular type III secretion by Salmonella typhimurium. AB - Type III secretion systems (TTSS) are important virulence factors that Gram negative bacteria use to translocate proteins into the cytoplasm of eukaryotic host cells. Salmonellae encode two virulence-associated TTSS. The Salmonella pathogenicity island 1 (SPI1)-encoded TTSS is active on contact with host cells, whereas the Salmonella pathogenicity island 2 (SPI2)-encoded TTSS is expressed after phagocytosis of bacteria by host cells. Previously, no consensus signal sequence for translocation has been identified among TTSS effector proteins. In this work, seven proteins, termed Salmonella-translocated effectors (STE), are described that contain conserved amino acid sequences that direct translocation by TTSS in Salmonella typhimurium. STE that are coordinately regulated with SPI2 gene expression contain translocation signals that are recognized by the SPI2 but not by the SPI1 TTSS. STE that are constitutively expressed contain signals that direct translocation through both SPI1 and SPI2 TTSS. Of the seven STE examined, SspH1 and SspH2 have been previously shown to be translocated and involved in virulence; SlrP and SifA were identified as virulence factors, but were not previously known to be associated with TTSS; and SseI, SseJ, and SifB were previously unidentified. Three STE genes (sspH1, sspH2, and sseI) are located within temperate bacteriophages, suggesting a common mechanism for the dissemination of more recently evolved STE. PMID- 10861018 TI - Steroid receptor coactivator-1 (SRC-1) mediates the development of sex-specific brain morphology and behavior. AB - Steroid hormone action during brain development exerts profound effects on reproductive physiology and behavior that last into adulthood. A variety of in vitro studies indicate that steroid receptors require nuclear receptor coactivators for efficient transcriptional activity. To determine the functional significance of the nuclear receptor coactivator SRC-1 in developing brain, we investigated the consequence of reducing SRC-1 protein during sexual differentiation of the brain. We report that reducing SRC-1 protein interferes with the defeminizing actions of estrogen in neonatal rat brain. Our data indicate that SRC-1 protein expression is critically involved in the hormone dependent development of normal male reproductive behavior and brain morphology. PMID- 10861019 TI - Immunogold evidence that neuronal gap junctions in adult rat brain and spinal cord contain connexin-36 but not connexin-32 or connexin-43. AB - Physiological and ultrastructural evidence indicates that gap junctions link many classes of neurons in mammalian central nervous system (CNS), allowing direct electrical and metabolic communication. Among at least six gap junction-forming connexin proteins in adult rat brain, connexin- (Cx) 32, Cx36, and Cx43 have been reported to occur in neurons. However, no connexin has been documented at ultrastructurally defined neuronal gap junctions. To address this question directly, freeze-fracture replica immunogold labeling (FRIL) and immunofluorescence (IF) were used to visualize the subcellular and regional localization of Cx36 in rat brain and spinal cord. Three antibodies were generated against different sequences in Cx36. By Western blotting, these antibodies detected protein at 36 and 66 kDa, corresponding to Cx36 monomer and dimer forms, respectively. After double-labeling for Cx36 and Cx43 by FRIL, neuronal gap junctions in inferior olive, spinal cord, and retina were consistently immunogold-labeled for Cx36, but none were labeled for Cx43. Conversely, Cx43 but not Cx36 was detected in astrocyte and ependymocyte gap junctions. In >250 Cx32/Cx43 single- and double-labeled replicas from 10 CNS regions, no neuronal gap junctions were labeled for either Cx32 or Cx43. Instead, Cx32 and Cx43 were restricted to glial gap junctions. By IF, Cx36 labeling was widely distributed in neuropil, including along dendritic processes and within neuronal somata. On the basis of FRIL identification of Cx36 in neuronal gap junctions and IF imaging of Cx36 throughout rat brain and spinal cord, neuronal gap junctions containing Cx36 appear to occur in sufficient density to provide widespread electrical and metabolic coupling in adult CNS. PMID- 10861020 TI - Attentional modulation of effective connectivity from V2 to V5/MT in humans. AB - The nonlinear nature of integration among cortical brain areas renders the effective connectivity between them inherently dynamic and context-sensitive. One emerging architectural principle of functional brain organization, which rests explicitly on these nonlinear interactions, is that neuronal responses expressed at any level in a sensory hierarchy reflect an interaction between (i) bottom up "driving" afferents from lower cortical areas and (ii) backwards "modulatory" inputs from higher areas that mediate top-down contextual effects. A compelling example is attentional modulation of responses in functionally specialized sensory areas. The aim of this work was to demonstrate that parietal regions may mediate selective attention to motion by modulating the effective connectivity from early visual cortex to the motion-sensitive area V5/MT. Using functional magnetic resonance imaging, and an analysis of effective connectivity based on nonlinear system identification, we found that backwards modulatory influences from the posterior parietal cortex are sufficient to account for a significant component of attentional modulation of V5/MT responses to "driving" inputs from V2. By explicitly modeling interactions among inputs to V5/MT, we were able to make inferences about the influences of V2 inputs and their concomitant activity dependent modulation by parietal afferents. The latter effects embody dynamic changes in effective connectivity that may underlie attentional mechanisms. These results speak to the context-sensitive nature of functional integration in the brain and provide empirical evidence that attentional effects may be mediated by backwards connections, of a modulatory sort, in humans. PMID- 10861021 TI - Complementary roles of neurotrophin 3 and a N-methyl-D-aspartate antagonist in the protection of noise and aminoglycoside-induced ototoxicity. AB - Recent progress has been made regarding the prevention of hearing loss. However, the complete protection of both hair cells and spiral ganglion neurons, with restored function, has not yet been achieved. It has been shown that spiral ganglion neuronal loss can be prevented by neurotrophin 3 (NT3) and hair cell damage by N-methyl-D-aspartate (NMDA) receptor antagonists. Here we demonstrate that the combined treatment with MK801, a NMDA antagonist, and NT3 protect both cochlear morphology and physiology from injury. Pretreatment with MK801 prevented hearing loss and the dendrites of the spiral ganglion neurons from swelling after noise-induced damage. The acute phase of insult with the aminoglycoside antibiotic amikacin resulted in swollen afferent dendrites beneath the inner hair cells. The chronic phase resulted in complete hair cell loss and near-complete loss of spiral ganglion neurons. This damage caused a near-complete loss of hearing sensitivity as displayed by elevated (>90-dB sound pressure levels) auditory brainstem response thresholds. The treatment of amikacin-exposed animals with MK801 gave only a partial protection of hearing. However, the combined treatment with NT3 and MK801 in the amikacin-comprised ear resulted in improved mean hearing within 20 dB of normal. Furthermore, hair cell loss was prevented in these animals and spiral ganglion neurons were completely protected. These results suggest that the NMDA antagonist MK801 protects against noise-induced excitotoxicity in the cochlea whereas the combined treatment of NT3 and MK801 has a potent effect on preserving both auditory physiology and morphology against aminoglycoside toxicity. PMID- 10861022 TI - Coupling between changes in human brain temperature and oxidative metabolism during prolonged visual stimulation. AB - A fundamental discovery of modern human brain imaging with positron-emission tomography that the blood flow to activated regions of the normal human brain increases substantially more than the oxygen consumption has led to a broad discussion in the literature concerning possible mechanisms responsible for this phenomenon. Presently no consensus exists. It is well known that oxygen delivery is not the only function of systemic circulation. Additional roles include delivery of nutrients and other required substances to the tissue, waste removal, and temperature regulation. Among these other functions, the role of regional cerebral blood flow in local brain temperature regulation has received scant attention. Here we present a theoretical analysis supported by empirical data obtained with functional magnetic resonance suggesting that increase in regional cerebral blood flow during functional stimulation can cause local changes in the brain temperature and subsequent local changes in the oxygen metabolism. On average, temperature decreases by 0.2 degrees C, but individual variations up to +/-1 degrees C were also observed. Major factors contributing to temperature regulation during functional stimulation are changes in the oxygen consumption, changes in the temperature of incoming arterial blood, and extensive heat exchange between activated and surrounding brain tissue. PMID- 10861023 TI - In vivo detection of amyloid plaques in a mouse model of Alzheimer's disease. AB - Strategies for treating Alzheimer's disease (AD) include therapies designed to decrease senile plaque (SP) formation and/or promote clearance of SPs, but clinical trials of these treatments are limited by the lack of effective methods to monitor changes in plaque burden in the brains of living AD patients. However, because SPs are extracellular deposits of amyloid-beta peptides (Abeta), it may be possible to eventually develop radioligands that cross the blood-brain barrier (BBB) and label SPs so they can be visualized by current imaging methods. As a first step toward the generation of such a radioligand, we developed a probe, [(trans,trans)-1-bromo-2, 5-bis-(3-hydroxycarbonyl-4-hydroxy)styrylbenzene (BSB)], and we report here that BSB has the following properties essential for a probe that can detect SPs in vivo. First, BSB sensitively labels SPs in AD brain sections. Second, BSB permeates living cells in culture and binds specifically to intracellular Abeta aggregates. Third, after intracerebral injection in living transgenic mouse models of AD amyloidosis, BSB labels SPs composed of human Abeta with high sensitivity and specificity. Fourth, BSB crosses the BBB and labels numerous AD-like SPs throughout the brain of the transgenic mice after i.v. injection. Thus, we conclude that BSB is an appropriate starting point for future efforts to generate an antemortem diagnostic for AD. PMID- 10861024 TI - T-type alpha 1H Ca2+ channels are involved in Ca2+ signaling during terminal differentiation (fusion) of human myoblasts. AB - Mechanisms underlying Ca(2+) signaling during human myoblast terminal differentiation were studied using cell cultures. We found that T-type Ca(2+) channels (T-channels) are expressed in myoblasts just before fusion. Their inhibition by amiloride or Ni(2+) suppresses fusion and prevents an intracellular Ca(2+) concentration increase normally observed at the onset of fusion. The use of antisense oligonucleotides indicates that the functional T-channels are formed by alpha1H subunits. At hyperpolarized potentials, these channels allow a window current sufficient to increase [Ca(2+)](i). As hyperpolarization is a prerequisite to myoblast fusion, we conclude that the Ca(2+) signal required for fusion is produced when the resting potential enters the T-channel window. A similar mechanism could operate in other cell types of which differentiation implicates membrane hyperpolarization. PMID- 10861025 TI - Enhanced learning after genetic overexpression of a brain growth protein. AB - Ramon y Cajal proposed 100 years ago that memory formation requires the growth of nerve cell processes. One-half century later, Hebb suggested that growth of presynaptic axons and postsynaptic dendrites consequent to coactivity in these synaptic elements was essential for such information storage. In the past 25 years, candidate growth genes have been implicated in learning processes, but it has not been demonstrated that they in fact enhance them. Here, we show that genetic overexpression of the growth-associated protein GAP-43, the axonal protein kinase C substrate, dramatically enhanced learning and long-term potentiation in transgenic mice. If the overexpressed GAP-43 was mutated by a Ser --> Ala substitution to preclude its phosphorylation by protein kinase C, then no learning enhancement was found. These findings provide evidence that a growth related gene regulates learning and memory and suggest an unheralded target, the GAP-43 phosphorylation site, for enhancing cognitive ability. PMID- 10861026 TI - Cutting edge: administration of anti-CD40 ligand and donor bone marrow leads to hemopoietic chimerism and donor-specific tolerance without cytoreductive conditioning. AB - Transplantation tolerance, defined as allograft acceptance by an immunocompetent recipient in the absence of long-term immunosuppression, has remained an elusive goal in clinical transplantation. Robust experimental tolerance induction strategies have in common methods to induce mixed hemopoietic chimerism. To date, however, chimerism induction across allogeneic barriers has required recipient conditioning with irradiation or cytoablative agents. In this paper we show that B6 recipients of fully allogeneic BALB/c skin grafts treated with repeated doses of donor bone marrow and anti-CD40 ligand (CD40L) develop durable (>300 days), readily detectable (6-12%) multilineage hemopoietic chimerism, indefinite allograft acceptance (>300 days), and donor-specific tolerance to secondary skin grafts. Analysis of the TCR repertoire of treated mice indicates that the underlying mechanisms of tolerance are in part mediated by deletion of donor reactive T cells. These data demonstrate that durable hemopoietic chimerism and robust transplantation tolerance can be achieved without cytotoxic conditioning using a potentially clinically applicable regimen. PMID- 10861027 TI - Cutting edge: antilisterial activity of CD8+ T cells derived from TNF-deficient and TNF/perforin double-deficient mice. AB - The mechanisms by which CD8+ T cells mediate immunity against bacterial pathogens remain largely unknown. Perforin-dependent cytolysis plays a role, but is not required for CD8+ T cell-mediated immunity against Listeria monocytogenes. TNF is essential for CD8+ T cell immunity to L. monocytogenes, but the cellular source of TNF is undefined. TNF-deficient and TNF/perforin double-deficient mice were used to generate CD8+ T cells specific for an L. monocytogenes-derived Ag. Wild type and TNF-deficient CD8+ T cells mediated antilisterial immunity in wild-type but not TNF-deficient host mice, revealing that CD8+ T cell-derived TNF is not required for CD8+ T cell-mediated antilisterial immunity, but demonstrating a role for TNF derived from other cell types. TNF/perforin double-deficient CD8+ T cells mediated antilisterial immunity in the liver, but not in the spleen, of wild-type recipient mice, suggesting that perforin-independent immunity in the spleen requires CD8+ T cell-derived TNF. PMID- 10861028 TI - Cutting edge: differential expression of chemokines in Th1 and Th2 cells is dependent on Stat6 but not Stat4. AB - The in vivo function of Th cell subsets is largely dependent on the ability of differentiated CD4+ T cells to be recruited to specific sites and secrete restricted sets of cytokines. In this paper we demonstrate that Th1 and Th2 cells secrete discrete patterns of chemokines, small m.w. cytokines that function as chemoattractants in inflammatory reactions. Th2 cells secrete macrophage-derived chemokine and T cell activation gene 3, and acquisition of this pattern of expression is dependent on Stat6. In contrast, Th1 cells secrete lymphotactin and RANTES, though unlike IFN-gamma, expression of these chemokines is independent of Stat4. We further show that supernatants from activated Th2 cells preferentially induce the chemotaxis of Th2 over Th1 cells, corresponding with Stat6-dependent expression of CCR4 and CCR8 in Th2 cells. These data provide the basis for restricted and direct T cell-mediated cellular recruitment to sites of inflammation. PMID- 10861029 TI - Cutting edge: hierarchy of chemokine receptor and TCR signals regulating T cell migration and proliferation. AB - Chemokines play an important role in establishing the distribution of lymphocyte subpopulations in primary and secondary lymphoid tissues and in the recruitment of leukocytes to sites of inflammation. However, the potential of chemokines to down-regulate immune responses has not been demonstrated. We now show that certain chemokine gradients have the potential to suppress T cell activation by preventing formation of the immunological synapse, the specialized cell-cell junction that forms before a T cell can be fully activated. Our data reveals an immunosuppressive potential of chemokines engaging the CXCR3 and CCR7 receptors, but not the CXCR4, CCR2, CCR4, or CCR5 receptors. These results suggest a novel mechanism for T cell ignorance of agonist MHC-peptide complexes based on dominant chemokine gradients. PMID- 10861030 TI - Cutting edge: introduction of an endopeptidase cleavage motif into a determinant flanking region of hen egg lysozyme results in enhanced T cell determinant display. AB - The choice of which determinants of a whole Ag will be presented on cell surface MHC class II molecules after uptake and processing by APC is the result of the interplay between structural characteristics of the Ag and the processing machinery of the APC. In this study, we demonstrate that introduction of a dibasic motif adjacent to a subdominant determinant enhances the presentation of this determinant from the whole molecule. This is the first report showing that a single amino acid substitution in a whole Ag, designed to introduce an endopeptidase recognition site, enhances display of class II-restricted determinants, most likely by creating a peptide chain cleavage in the antigenic molecule. Our findings have important implications for the understanding of immunodominance and for vaccine design. PMID- 10861031 TI - T cell tolerance based on avidity thresholds rather than complete deletion allows maintenance of maximal repertoire diversity. AB - Given the flexible nature of TCR specificity, deletion or permanent disabling of all T cells with the capacity to recognize self peptides would severely limit the diversity of the repertoire and the capacity to recognize foreign Ags. To address this, we have investigated the patterns of CD8+ CTL reactivity to a naturally H 2Kb-presented self peptide derived from the elongation factor 1alpha (EF1alpha). EF1alpha occurs as two differentially expressed isoforms differing at one position of the relevant peptide. Low avidity CTLs could be raised against both variants of the EF1alpha peptide. These CTLs required 100-fold more peptide-H-2Kb complexes on the target cell compared with CTLs against a viral peptide, and did not recognize the naturally expressed levels of EF1alpha peptides. Thus, low avidity T cells specific for these self peptides escape tolerance by deletion, despite expression of both EF1alpha isoforms in dendritic cells known to mediate negative selection in the thymus. The low avidity in CTL recognition of these peptides correlated with low TCR affinity. However, self peptide-specific CTLs expressed elevated levels of CD8. Furthermore, CTLs generated against altered self peptide variants displayed intermediate avidity, indicating cross-reactivity in induction of tolerance. We interpret these data, together with results previously published by others, in an avidity pit model based on avidity thresholds for maintenance of both maximal diversity and optimal self tolerance in the CD8+ T cell repertoire. PMID- 10861032 TI - Induction of permanent mixed chimerism and skin allograft tolerance across fully MHC-mismatched barriers by the additional myelosuppressive treatments in mice primed with allogeneic spleen cells followed by cyclophosphamide. AB - A pure method of drug (cyclophosphamide plus busulfan)-induced skin allograft tolerance in mice that can regularly overcome fully H-2-mismatched barriers in mice has been established. The components of the method are i.v. administration of 1 x 108 allogeneic spleen cells on day 0, i.p. injection of 200 mg/kg CP and 25 mg/kg busulfan on day 2, and i.v. injection of T cell-depleted 1 x 107 bone marrow cells from the same donor on day 3. Recipient B10 (H-2b; IE-) mice prepared with this conditioning developed donor-specific tolerance, and long lasting survival of skin allografts was shown in almost of the recipient mice. In the tolerant B10 mice prepared with new conditioning, stable multilineage mixed chimerism was observed permanently, and IE-reactive Vbeta11+ T cells were reduced in periphery as seen in untreated B10.D2 (H-2d; IE+) mice. The specific tolerant state was confirmed by the specific abrogation against donor Ag in the assays of CTL activity and MLR and donor-specific acceptance in the second skin grafting. These results demonstrated that the limitation of standard protocol of cyclophosphamide-induced tolerance, which have been reported by us since 1984, can be overcome by the additional treatments with the myelosuppressive drug busulfan, followed by 1 x 107 T cell-depleted bone marrow cells. To our knowledge, this is the first report to induce allograft tolerance with a short course of the Ag plus immunosuppressive drug treatment without any kind of mAbs (pure drug-induced tolerance). PMID- 10861033 TI - Genetic mapping of two murine loci that influence the development of IL-4 producing Thy-1dull gamma delta thymocytes. AB - IL-4-producing gamma delta cells belong to a novel subset of gamma delta lymphocytes that expresses a very restricted repertoire of TCRs. To gain a deeper insight into the development and in vivo functions of these cells, we have analyzed the genetic control of their representation in the thymus. Using an intercross between C57BL/6 and DBA/2 mice we found two loci on chromosomes 13 and 17-named LadT1 and LadT2, respectively-with marked influence in their development. The LadT2 locus does not appear to be the MHC locus. The region identified on mouse chromosome 13 contains the structural genes for TCR gamma as well as the IL-9 gene, which has been suggested as a candidate gene influencing the complex pathogenesis of asthma. PMID- 10861034 TI - Polyethylene glycol-modified GM-CSF expands CD11b(high)CD11c(high) but notCD11b(low)CD11c(high) murine dendritic cells in vivo: a comparative analysis with Flt3 ligand. AB - Dendritic cells (DC) are potent APCs that can be characterized in the murine spleen as CD11b(high)CD11c(high) or CD11b(low)CD11c(high). Daily injection of mice of Flt3 ligand (FL) into mice transiently expands both subsets of DC in vivo, but the effect of administration of GM-CSF on the expansion of DC in vivo is not well defined. To gain further insight into the role of GM-CSF in DC development and function in vivo, we treated mice with polyethylene glycol modified GM-CSF (pGM-CSF) which has an increased half-life in vivo. Administration of pGM-CSF to mice for 5 days led to a 5- to 10-fold expansion of CD11b(high)CD11c(high) but not CD11b(low)CD11c(high) DC. DC from pGM-CSF-treated mice captured and processed Ag more efficiently than DC from FL-treated mice. Although both FL- and pGM-CSF-generated CD11b(high)CD11c(high) DC were CD8alpha-, a greater proportion of these DC from pGM-CSF-treated mice were 33D1+ than from FL-treated mice. CD11b(low)CD11c(high) DC from FL-treated mice expressed high levels of intracellular MHC class II. DC from both pGM-CSF- and FL-treated mice expressed high levels of surface class II, low levels of the costimulatory molecules CD40, CD80, and CD86 and were equally efficient at stimulating allogeneic and Ag-specific T cell proliferation in vitro. The data demonstrate that treatment with pGM-CSF in vivo preferentially expands CD11b(high)CD11c(high) DC that share phenotypic and functional characteristics with FL-generated CD11b(high)CD11c(high) DC but can be distinguished from FL-generated DC on the basis of Ag capture and surface expression of 33D1. PMID- 10861035 TI - IL-12 responsiveness and expression of IL-12 receptor in human peripheral blood monocyte-derived dendritic cells. AB - We analyzed the expression of IL-12Rbeta1 and IL-12Rbeta2 and the role of IL-12 in the activation of monocyte-derived dendritic cells (DCs) via IL-12Rbeta1 mediated signaling events. Flow cytometric analysis revealed that IL-12Rbeta1 was expressed in T cells, Con A blasts, and monocyte-derived DCs, but not in monocytes, while its transcript was detected in all of these cell types. Transcriptional expression of IL-12Rbeta2 was observed in T cells, Con A blasts, and monocyte-derived DCs, but not monocytes. The ligation of DCs as well as Con A blasts by IL-12 induced the production of GM-CSF, IL-1beta, IL-6, TNF-alpha, and IFN-gamma at the transcription levels. Furthermore, stimulation of DCs with IL-12 induced IL-12p40 transcript, but not IL-12p35 transcript, whereas this stimulation caused the expressions of both transcripts in Con A blasts. Stimulation of DCs with IL-12 caused a tyrosine phosphorylation of several intracellular proteins, and the pattern of these events were distinct from those of IL-12-stimulated Con A blasts. IL-12 also induced tyrosine phosphorylation of IL-12Rbeta1 as well as recruitment of several tyrosine-phosphorylated proteins to IL-12Rbeta1 in DCs and Con A blasts. Receptor engagement of DCs as well as Con A blasts by IL-12 resulted in activation of Janus kinase 2 and Tyk2 kinases and Stat3 and Stat4 transcription factors and the association of these proteins to IL 12Rbeta1. Stimulation with IL-12 caused a tyrosine phosphorylation and enzymatic activity of a family of mitogen-activated protein kinases, p38mapk. These results suggest that IL-12 acts directly on DCs to induce their functional activation via IL-12Rbeta1-mediated signaling events. PMID- 10861036 TI - Rapid, B lymphoid-restricted engraftment mediated by a primitive bone marrow subpopulation. AB - Utilizing multiparameter flow cytometry, we have defined a subset of bone marrow cells containing lymphoid-restricted differentiation potential after i.v. transplantation. Bone marrow cells characterized by expression of the Sca-1 and c kit Ags and lacking Ags of differentiating lineages were segregated into subsets based on allele-specific Thy-1.1 Ag expression. Although hematopoietic stem cells were recovered in the Thy-1.1low subset as previously described, the Thy-1.1neg subset consisted of progenitor cells that preferentially reconstituted the B lymphocyte lineage after i.v. transplantation. Recipients of Thy-1.1neg cells did not survive beyond 30 days, presumably due to the failure of erythroid and platelet lineages to recover after transplants. Thy-1.1neg cells predominantly reconstituted the bone marrow and peripheral blood of lethally irradiated recipients with B lineage cells within 2 weeks, although a low frequency of myeloid lineage cells was also detected. In contrast, myeloid progenitors outnumbered lymphoid progenitors when the Thy-1.1neg population was assayed in culture. When Thy-1. 1low stem cells were rigorously excluded from the Thy-1.1neg subset, reconstitution of T lymphocytes was rarely observed in peripheral blood after i.v. transplantation. Competitive repopulation studies showed that the B lymphoid reconstitution derived from Thy-1.1neg cells was not sustained over a 20 wk period. Therefore, the Thy-1. 1neg population defined in these studies includes transplantable, non-self-renewing B lymphocyte progenitor cells. PMID- 10861037 TI - Subtle effects on myelin basic protein-specific T cell responses can lead to a major reduction in disease susceptibility in experimental allergic encephalomyelitis. AB - The presence of potentially autoreactive T cells is a necessary, but not sufficient, condition for the development of autoimmune disease. However, the relationship between T cell response and susceptibility to disease is not straightforward. In this report, we use experimental allergic encephalomyelitis as a model to demonstrate that subtle alterations of the T cell response to an encephalitogenic epitope are sufficient to cause a dramatic decrease in disease susceptibility. Transgenic expression of a fusion protein of hen egg lysozyme and an encephalitogenic peptide of myelin basic protein (MBP) residues 84-105, coexpressed with MHC class II, causes profound tolerance to hen egg lysozyme, while maintaining a near normal response to MBP. Detailed analysis of the T cell repertoire of transgenic animals using a panel of T cell hybridomas revealed a highly selective loss of one minor component of the response to the MBP84-104 region. Despite this, transgenic animals were highly resistant to experimental allergic encephalomyelitis induction with the MBP peptide, indicating that minor changes to the T cell repertoire may result in major alterations in disease susceptibility. Possible reasons for this are discussed. PMID- 10861038 TI - An amiloride-sensitive and voltage-dependent Na+ channel in an HLA-DR-restricted human T cell clone. AB - We investigated changes in voltage-gated Na+ currents and effects of extracellular Na+ on proliferation in HLA-DR-restricted human CD4+ alphabeta T cells after stimulation with a non-self antigenic peptide, M12p54-68. In the absence of antigenic peptide, neither single (n = 80) nor APC-contacted (n = 71) T cells showed voltage-gated inward currents recording with whole-cell patch clamp techniques, even with Ca2+ and Na+ ions present in the perfusion solution. However, with the same recording conditions, 31% (26 of 84) of APC-contacted T cells stimulated with the antigenic peptide showed voltage-dependent inward currents that were elicited from -60 mV. The inward currents were not inhibited in extracellular Ca2+-free conditions or in the presence of 1 mM NiCl2. However, they were completely inhibited in extracellular Na+-free conditions, which were made by replacing Na+ with iso-osmotic N-methyl-d -glucamine or choline. The Na+ currents were insensitive to tetrodotoxin, a classical blocker of Na+ channels, but were dose-dependently inhibited by amiloride, a potassium-sparing pyrazine diuretic. Furthermore, the Ag-specific proliferative response of T cells was completely inhibited in Na+-free Tyrode's solution and was suppressed by amiloride in a dose-dependent manner. Our findings suggest that activation of amiloride-sensitive and voltage-gated Na+ channels would be an important step to allow an adequate influx of Na+ and maintain a sustained high Ca2+ level during T cell activation. PMID- 10861039 TI - Positive impact of inhibitory Ly49 receptor-MHC class I interaction on NK cell development. AB - NK cells can kill MHC-different or MHC-deficient but not syngeneic MHC-expressing target cells. This MHC class I-specific tolerance is acquired during NK cell development. MHC recognition by murine NK cells largely depends on clonally distributed Ly49 family receptors, which inhibit NK cell function upon ligand engagement. We investigated whether these receptors play a role for the development of NK cells and provide evidence that the expression of a Ly49 receptor transgene on developing NK cells endowed these cells with a significant developmental advantage over NK cells lacking such a receptor, but only if the relevant MHC ligand was present in the environment. The data suggest that the transgenic Ly49 receptor accelerates and/or rescues the development of NK cells which would otherwise fail to acquire sufficient numbers of self-MHC-specific receptors. Interestingly, the positive effect on NK cell development is most prominent when the MHC ligand is simultaneously present on both hemopoietic and nonhemopoietic cells. These findings correlate with functional data showing that MHC class I ligand on all cells is required to generate functionally mature NK cells capable of reacting to cells lacking the respective MHC ligand. We conclude that the engagement of inhibitory MHC receptors during NK cell development provides signals that are important for further NK cell differentiation and/or maturation. PMID- 10861040 TI - Anti-TCR-specific DNA vaccination demonstrates a role for a CD8+ T cell clone in the induction of allograft tolerance by donor-specific blood transfusion. AB - Donor-specific allograft tolerance can be induced in the adult rat by pregraft donor-specific blood transfusion (DST). This tolerance appeared to be mediated by regulatory cells and to the production of the suppressive cytokine TGF-beta1. A potential immunoregulatory CD8+ clone bearing a Vbeta18-Dbeta1-Jbeta2.7 TCR gene rearrangement was previously identified in DST-treated recipients. To assess the functional role of this T cell clone in the induction of tolerance by DST, we have vaccinated DST-treated recipients with a plasmid construct encoding for the Vbeta18-Dbeta1-Jbeta2.7 TCR beta-chain. DST-induced allograft tolerance was abolished by anti-TCR Vbeta18-Dbeta1-Jbeta2.7 DNA vaccination in six of seven recipients, whereas vaccination with the vector alone, or with the construct encoding a TCR Vbeta13 beta-chain, had no effect. However, the transcript number of the Vbeta18-Dbeta1-Jbeta2.7 chain was unchanged in allografts from vaccinated DST-treated rats, suggesting that this clone was not depleted by vaccination, but rather was altered in its function. Moreover, TCR Vbeta18-Dbeta1-Jbeta2.7 DNA vaccination restored the anti-donor alloantibody production, partially restore the capacity of spleen cells from tolerized recipients to proliferate in vitro against donor cells, and decreased the inhibitory effect of TGF-beta1, seen in DST-treated recipients, in spleen cells from vaccinated DST-treated ones. This study strongly suggests that this CD8+ TCR Vbeta18-Dbeta1-Jbeta2.7 T cell clone has an effective immunoregulatory function in allograft tolerance induced by DST. PMID- 10861041 TI - Rebound from nitric oxide inhibition triggers enhanced monocyte activation and chemotaxis. AB - Exposure of human peripheral blood monocytes to the NO donor S-nitroso-N-acetyl DL-penicillamine (SNAP) resulted in a rapid shift in cellular conformation of spontaneously activated cells from ameboid to round. The population of activated cells, approximately 7. 1 +/- 1.2%, was reduced 7-fold to 1.1 +/- 0.4% following 0.5 h exposure to SNAP. Observation of monocytes for 6 h demonstrated a gradual release from NO inhibition initiating at 2.5 h following SNAP treatment and a period of hyperactivity that was maximal at approximately 5 h following SNAP exposure. During the rebound from the NO inhibition phase, there was a significant increase in the population of activated monocytes and an increased responsiveness to chemotactic agents such as IL-1, IL-8, and fMLP relative to that of cells treated with the chemotactic agents alone. Conformational changes induced by SNAP were associated with a reduction in F-actin and loss of filopodial extension. The loss and recovery of F-actin staining paralleled changes in cell activity, suggesting that NO may alter cellular activity by modulation of cytoskeletal actin. These data taken together suggest that inhibition of monocyte activity by NO results in an excitatory phase observed subsequent to release from NO inhibition and increased sensitivity to chemotactic agents. We propose that this rebound from NO inhibition may provide increased immunosurveillance to rectify immunological problems that have been encountered during the period of inhibition. PMID- 10861042 TI - Integrated signals between IL-13, IL-4, and IL-5 regulate airways hyperreactivity. AB - In this investigation, we have examined the integrated relationship between IL 13, IL-4, and IL-5 for the development of airways hyperreactivity (AHR) in a model of asthma in BALB/c mice. Sensitization and aeroallergen challenge of both wild-type (WT) and IL-13 gene-targeted (IL-13-/-) mice induced allergic disease that was characterized by pulmonary eosinophilia and AHR to beta-methacholine. Although these responses in IL-13-/- mice were heightened compared with WT, they could be reduced to the level in nonallergic mice by the concomitant neutralization of IL-4. Mice in which both IL-4 and IL-13 were depleted displayed a marked reduction in tissue eosinophils, despite the development of a blood eosinophilia. Similar neutralization of IL-4 in WT mice only partially reduced AHR with no effect on tissue eosinophilia. In addition, neutralization of IL-5 in IL-13-/- mice, but not in WT mice, inhibited AHR, suggesting that tissue eosinophilia is linked to the mechanism underlying AHR only in the absence of IL 13. Additionally, mucus hypersecretion was attenuated in IL-13-/- mice, despite the persistence of AHR. Taken together, our data suggest both a modulatory role for IL-13 during sensitization and a proinflammatory role during aeroallergen challenge. The latter process appears redundant with respect to IL-4. PMID- 10861043 TI - Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide inhibit T cell-mediated cytotoxicity by inhibiting Fas ligand expression. AB - We reported recently that the neuropeptides vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) protect CD4+ T cells against Ag-induced apoptosis by down-regulating the expression of Fas ligand (FasL). Because the cytotoxic activity of CD8+ CTLs is mediated through two mechanisms, which involve the perforin/granzyme and the FasL/Fas pathways, in this study we investigated the effects of VIP/PACAP on the generation and activity of allogeneic CTLs, of CD8+ T1 and T2 effector cells and of alloreactive peritoneal exudate cytotoxic T cells (PEL) generated in vivo. VIP/PACAP did not affect perforin/granzyme-mediated cytotoxicity, perforin gene expression, or granzyme B enzymatic activity, but drastically inhibited FasL/Fas-mediated cytotoxicity against allogeneic or syngeneic Fas-bearing targets. VIP/PACAP inhibit CTL generation, but not the activity of competent CTLs. The inhibition is associated with a profound down-regulation of FasL expression, and these effects are mediated through both VPAC1 and VPAC2 receptors. VIP/PACAP inhibit the FasL/Fas-mediated cytotoxicity of T1 effectors and do not affect T2 cytotoxicity, which is entirely perforin/granzyme mediated. Similar effects were observed in vivo. Both the FasL/Fas-mediated cytotoxicity and FasL expression of cytotoxic allogeneic PELs generated in vivo in the presence of VIP or PACAP were significantly reduced. We conclude that, similar to their effect on CD4+ T cells, the two structurally related neuropeptides inhibit FasL expression in CD8+ cytotoxic T cells and the subsequent lysis of Fas-bearing target cells. PMID- 10861044 TI - Thapsigargin-induced degranulation of mast cells is dependent on transient activation of phosphatidylinositol-3 kinase. AB - Thapsigargin, which elevates cytosolic calcium levels by inhibiting the sarcoplasmic/endoplasmic reticulum calcium-dependent ATPase, was tested for its ability to degranulate bone marrow-derived mast cells (BMMCs) from src homology 2 containing inositol phosphatase +/+ (SHIP+/+) and SHIP-/- mice. As was found previously with steel factor, thapsigargin stimulated far more degranulation in SHIP-/- than in SHIP+/+ BMMCs, and this was blocked with the phosphatidylinositol 3 (PI-3) kinase inhibitors, LY294002 and wortmannin. In contrast to steel factor, however, this heightened degranulation of SHIP-/- BMMCs was not due to a greater calcium influx into these cells, nor was the thapsigargin-induced calcium influx inhibited by LY294002, suggesting that the heightened thapsigargin-induced degranulation of SHIP-/- BMMCs was due to a PI-3 kinase-regulated step distinct from that regulating calcium entry. An investigation of thapsigargin-stimulated pathways in both cell types revealed that MAPK was heavily but equally phosphorylated. Interestingly, the protein kinase C inhibitor, bisindolylmaleimide (compound 3), totally blocked thapsigargin-induced degranulation in both SHIP+/+ and SHIP-/- BMMCs. As well, thapsigargin stimulated a PI-3 kinase-dependent, transient activation of protein kinase B, and this activation was far greater in SHIP-/- than in SHIP+/+ BMMCs. Consistent with this, thapsigargin was found to be a potent survival factor, following cytokine withdrawal, for both cell types and was more potent with SHIP-/- cells. These studies have both identified an additional PI-3 kinase-dependent step within the mast cell degranulation process, possibly involving 3-phosphoinositide-dependent protein kinase-1 and a diacylglycerol-independent protein kinase C isoform, and shown that the tumor-promoting activity of thapsigargin may be due to its activation of protein kinase B and subsequent promotion of cell survival. PMID- 10861045 TI - Duplicated DQ haplotypes increase the complexity of restriction element usage in cattle. AB - The MHC of cattle encodes two distinct isotypes of class II molecules, DR and DQ. Unlike humans, cattle lack the DP locus and about half the common haplotypes express duplicated DQ genes. The number and frequency of DQA and DQB alleles means that most cattle are heterozygous. If inter- and/or intrahaplotype pairing of DQA and DQB molecules occurs, cattle carrying DQ-duplicated haplotypes may express more restriction elements than would be predicted by the number of expressed alleles. We are investigating whether duplicated haplotypes cause differences in immune response, particularly in terms of generating protective immunity. We have analyzed the Ag-presenting function of DQ molecules in two heterozygous animals, one of which carries a duplicated haplotype. We compared the class II isotype specificity of T cell clones recognizing a putative vaccinal peptide from foot-and-mouth disease virus (FMDV15). We show for the first time that bovine T cells can recognize Ag in the context of DQ molecules. We also present evidence that interhaplotype pairings of DQA and DQB molecules form functional restriction elements. Both animals showed distinct biases to usage of particular restriction elements. Mainly DQ-restricted clones were derived from the animal with duplicated DQ genes, whereas the majority of clones from the animal with a single DQ gene pair were DR restricted. Furthermore, haplotype bias was observed with both animals. These experiments show that understanding of class II chain pairing in addition to knowledge of the genotype may be important in vaccine design where effective epitope selection is essential. PMID- 10861046 TI - Lipopolysaccharide stimulates the proliferation of human CD56+CD3- NK cells: a regulatory role of monocytes and IL-10. AB - NK cells recognize and kill tumor cells and normal cells, and these play an important role in immune defense in cancer, infectious disease, and autoimmunity. NK killing is regulated by positive or negative signals derived from the interaction of surface receptors with ligands on the target cells. However, the mechanisms controlling the proliferation and maintenance of NK cells in normal human individuals are less clearly defined. In this study, using an entirely autologous system, we demonstrate that human peripheral blood CD3-CD56+, killer cell-inhibitory receptor (KIR)-expressing cells proliferate and expand in response to LPS. These responses are enhanced in the presence of anti-IL-10 receptor-blocking Abs or on the removal of CD14+ cells from the cultures. This enhancement is also reflected in substantial increases in cytolytic activity and IFN-gamma production. The negative effect of CD14+ cells may also be IL-10 mediated, IL-10 being lost from the culture supernatants of CD14-depleted PBMC and rIL-10 reversing the effect of this depletion. On the other hand, mRNA for the p35 and p40 subunits of IL-12 is still induced in CD14-depleted cultures. The expansion of CD3-CD56+ cells was also inhibited by CTLA4-Ig, indicating a role for CD80/86. B lymphocytes were not required for the expansion of CD3-CD56+ cells, whereas removal of MHC class II+ cells from CD14-depleted cultures resulted in a complete abrogation of these responses. Expansion of CD3-CD56+ cells was reconstituted in MHC class II-depleted cell cultures by adding back monocyte-derived dendritic cells. These results indicate that the responses of CD3-CD56+ NK cells to LPS may be driven by a MHC class II+ B7+ CD14- peripheral population, most likely blood dendritic cells. PMID- 10861047 TI - Linkage of the CCR5 Delta 32 mutation with a functional polymorphism of CD45RA. AB - A 32-bp deletion in CCR5 (CCR5 Delta 32) confers to PBMC resistance to HIV-1 isolates that use CCR5 as a coreceptor. To study this mutation in T cell development, we have screened 571 human thymus tissues for the mutation. We identified 72 thymuses (12.6%) that were heterozygous and 2 (0.35%) that were homozygous for the CCR5 Delta 32 mutation. We found that thymocyte development was normal in both CCR5 Delta 32 heterozygous and homozygous thymuses. In 3% of thymuses we identified a functional polymorphism of CD45RA, in which cortical and medullary thymocytes failed to down-regulate the 200- and 220-kDa CD45RA isoforms during T cell development. Moreover, we found an association of this CD45 functional polymorphism in thymuses with the CCR5 Delta 32 mutation (p = 0.00258). In vitro HIV-1 infection assays with CCR5-using primary isolates demonstrated that thymocytes with the heterozygous CCR5 Delta 32 mutation produced less p24 than did CCR5 wild-type thymocytes. However, the functional CD45RA polymorphism did not alter the susceptibility of thymocytes to HIV-1 infection. Taken together, these data demonstrate association of the CCR5 Delta 32 mutation with a polymorphism in an as yet unknown gene that is responsible for the ability to down-regulate the expression of high m.w. CD45RA isoforms. Although the presence of the CCR5 Delta 32 mutation down-regulates HIV-1 infection of thymocytes, the functional CD45RA polymorphism does not alter the susceptibility of thymocytes to HIV-1 infection in vitro. PMID- 10861048 TI - Dendritic cells in the induction of protective and nonprotective anticryptococcal cell-mediated immune responses. AB - Dendritic cells (DC) can be divided into three subsets, Langerhans cells, myeloid DC (MDC), and lymphoid DC (LDC), based upon phenotypic and functional differences. We hypothesized that different DC subsets are associated with the development of protective vs nonprotective cell-mediated immune (CMI) responses against the fungal pathogen, Cryptococcus neoformans. To test this, mice were immunized with protective and/or nonprotective immunogens, and DC subsets in draining lymph nodes were assessed by flow cytometry. The protective immunogen (cryptococcal culture filtrate Ag-CFA), in contrast to the nonprotective immunogen (heat-killed cryptococci-CFA), the nonprotective immunogen mixed with the protective immunogen (cryptococcal culture filtrate Ag + heat-killed cryptococci-CFA), or controls, stimulated significant increases in total leukocytes, Langerhans cells, MDC, LDC, and activated CD4+ T cells in draining lymph nodes. The protective immune response resulted in significantly increased levels of anticryptococcal delayed-type hypersensitivity reactivity and activated CD4+ T cells at the delayed-type hypersensitivity reaction site. Draining lymph node LDC:MDC ratios induced by the protective immunogen were significantly lower than the ratios induced by either immunization in which the nonprotective immunogen was present. In contrast, mice given the nonprotective immunogen had LDC:MDC ratios similar to those of naive mice. Our data indicate that lymph node Langerhans cells and MDC are APC needed for induction of the protective response. The predominance of LDC in mice undergoing nonprotective responses suggests that lymph node LDC, like splenic LDC, are negative regulators of CMI responses. In addition to showing DC subsets associated with functional differences, our data suggest that the LDC:MDC balance, which can be modulated by the Ag, determines whether protective or nonprotective anticryptococcal CMI responses develop. PMID- 10861049 TI - CD1d-specific NK1.1+ T cells with a transgenic variant TCR. AB - The majority of T lymphocytes carrying the NK cell marker NK1.1 (NKT cells) depend on the CD1d molecule for their development and are distinguished by their potent capacity to rapidly secrete cytokines upon activation. A substantial fraction of NKT cells express a restricted TCR repertiore using an invariant TCR Valpha14-Jalpha281 rearrangement and a limited set of TCR Vbeta segments, implying recognition of a limited set of CD1d-associated ligands. A second group of CD1d-reactive T cells use diverse TCR potentially recognizing a larger diversity of ligands presented on CD1d. In TCR-transgenic mice carrying rearranged TCR genes from a CD1d-reactive T cell with the diverse type receptor (using Valpha3. 2/Vbeta9 rearrangements), the majority of T cells expressing the transgenic TCR had the typical phenotype of NKT cells. They expressed NK1.1, CD122, intermediate TCR levels, and markers indicating previous activation and were CD4/CD8 double negative or CD4+. Upon activation in vitro, the cells secreted large amounts of IL-4 and IFN-gamma, a characteristic of NKT cells. In mice lacking CD1d, TCR-transgenic cells with the NKT phenotype were absent. This demonstrates that a CD1d-reactive TCR of the "non-Valpha 14" diverse type can, in a ligand-dependent way, direct development of NK1.1+ T cells expressing expected functional and cell-surface phenotype characteristics. PMID- 10861050 TI - cGMP-mediated inhibition of TNF-alpha production by the atrial natriuretic peptide in murine macrophages. AB - The atrial natriuretic peptide (ANP) is suggested to regulate inflammatory response by alteration of macrophage functions. The aim of this study was to investigate whether ANP influences production of TNF-alpha. TNF-alpha production in murine bone marrow-derived macrophages was induced by LPS, and TNF-alpha secretion (+/-ANP) was determined by L929 bioassay. ANP dose dependently (10-8-10 6 M) inhibited TNF-alpha release by up to 95%. The effect was mediated via the guanylate cyclase-coupled A receptor, as was shown by employing dibutyryl-cGMP, the cGMP-inhibitory compound Ly-83583, and the A receptor antagonist HS-142-1. A specific ligand of the natriuretic peptide "clearance" receptor inhibited TNF alpha production only at 10-7 and 10-8 M, but not at 10-6 M. The B receptor ligand C-type natriuretic peptide showed no TNF-alpha-inhibitory effect. To investigate the underlying mechanism of ANP-mediated TNF-alpha inhibition, Northern blot was performed. ANP-treated macrophages displayed decreased TNF alpha-mRNA levels. Besides the known inhibition of NF-kappaB activation, in this study we demonstrated that ANP also attenuates the activation of the proinflammatory transcription factor AP-1 (gel shift assay). ANP did not alter subunit composition of AP-1 complexes, as was shown by supershift assays applying anti-c-jun and anti-c-fos Abs. To get information on the ANP effect for human inflammatory processes, we investigated cytokine production in human LPS activated blood. ANP significantly attenuated production of TNF-alpha and IL 1beta without affecting production of IL-10 and IL-1ra. In summary, ANP was shown to attenuate TNF-alpha production of LPS-activated macrophages via cGMP. The inhibition is suggested to involve transcriptional processes that are the result of reduced activation of responsible transcription factors. PMID- 10861051 TI - IL-4 and IFN-gamma up-regulate substance P receptor expression in murine peritoneal macrophages. AB - While the ability of macrophages to express authentic substance P receptors (i.e., NK-1 receptors) has been inferred from radioreceptor binding assays and functional assays and, most recently, by identification of NK-1 receptor mRNA expression, we know little about NK-1 expression at the protein level or what host factors might up-regulate expression of this receptor. In the present study we demonstrate that the cytokines IL-4 and IFN-gamma can increase the expression of NK-1 receptors on murine peritoneal macrophages. Specifically, we show that IL 4 and IFN-gamma can elicit increases in the level of mRNA encoding the NK-1 receptor by up to 12- and 13-fold, respectively. Furthermore, these cytokines can significantly increase the expression of the NK-1 receptor protein as measured by Western blot and FACS analysis using specific Abs developed in our laboratory. In addition, we have demonstrated the ability of both IL-4 and IFN-gamma to enhance the ability of macrophages to bind substance P as measured by radiolabeled binding assay. The observation that the level of expression of this receptor protein can be enhanced by cytokines that promote either cell-mediated (Th1) or humoral (Th2) immune responses supports the idea that this receptor can be induced during either type of immune response. As such, these results may point to a more ubiquitous role for substance P in the generation of optimal immune responses than previously appreciated. PMID- 10861052 TI - Epithelial cell-specific laminin 5 is required for survival of early thymocytes. AB - The gene LamC2 encoding the gamma2 chain of laminin 5, an epithelial cell specific extracellular matrix protein, was identified in a PCR-based subtracted cDNA library from mouse thymic stromal cells. The mRNA existed in two alternative forms (5.1 and 2.4 kb). The full-length message was highly expressed in SCID thymus and in a nurse cell line, but not in other thymic epithelial cell lines, while the short form was more widely expressed. In situ hybridization and immunohistochemical staining revealed laminin 5 expression mostly in the subcapsular region of the adult thymus. Addition to fetal thymic organ cultures of a cell adhesion-blocking mAb to the alpha3 chain of laminin 5 interrupted T cell development. There was a 40% reduction in the total yield of thymocytes, and the most profound decrease (75-90%) was seen in the CD25+CD44+ and CD25+CD44 subsets of the CD4-CD8- double negative fraction. Most of the surviving double negative thymocytes expressed Sca-1, and there were significant increases in the number of cells with CD69 expression and in the fraction of annexin V-stained cells. None of these changes were observed with a nonblocking anti-laminin alpha3 chain mAb. These results suggest that the interaction between double negative thymoctyes and laminin 5 made by subcapsular epithelial cells is required for the survival and differentiation of mouse thymocytes. PMID- 10861053 TI - Systemic administration of agonist peptide blocks the progression of spontaneous CD8-mediated autoimmune diabetes in transgenic mice without bystander damage. AB - Insulin-dependent diabetes is an autoimmune disease targeting pancreatic beta islet cells. Recent data suggest that autoreactive CD8+ T cells are involved in both the early events leading to insulitis and the late destructive phase resulting in diabetes. Although therapeutic injection of protein and synthetic peptides corresponding to CD4+ T cell epitopes has been shown to prevent or block autoimmune disease in several models, down-regulation of an ongoing CD8+ T cell mediated autoimmune response using this approach has not yet been reported. Using CL4-TCR single transgenic mice, in which most CD8+ T cells express a TCR specific for the influenza virus hemagglutinin HA512-520 peptide:Kd complex, we first show that i.v. injection of soluble HA512-520 peptide induces transient activation followed by apoptosis of Tc1-like CD8+ T cells. We next tested a similar tolerance induction strategy in (CL4-TCR x Ins-HA)F1 double transgenic mice that also express HA in the beta-islet cells and, as a result, spontaneously develop a juvenile onset and lethal diabetes. Soluble HA512-520 peptide treatment, at a time when pathogenic CD8+ T cells have already infiltrated the pancreas, very significantly prolongs survival of the double transgenic pups. In addition, we found that Ag administration eliminates CD8+ T cell infiltrates from the pancreas without histological evidence of bystander damage. Our data indicate that agonist peptide can down-regulate an autoimmune reaction mediated by CD8+ T cells in vivo and block disease progression. Thus, in addition to autoreactive CD4+ T cells, CD8+ T cells may constitute targets for Ag-specific therapy in autoimmune diseases. PMID- 10861054 TI - Human mast cells transmigrate through human umbilical vein endothelial monolayers and selectively produce IL-8 in response to stromal cell-derived factor-1 alpha. AB - Mature mast cells are generally considered to be less mobile cells residing within tissue sites. However, mast cell numbers are known to increase in the context of inflammation, and mast cells are recognized to be important in regulating local neutrophil infiltration. CXC chemokines may play a critical role in this process. In this study two human mast cell-like lines, HMC-1 and KU812, and human cord blood-derived primary cultured mast cells were employed to examine role of stromal cell-derived factor-1 (SDF-1) in regulating mast cell migration and mediator production. It was demonstrated that human mast cells constitutively express mRNA and protein for CXCR4. Stimulation of human mast cells with SDF-1, the only known ligand for CXCR4, induced a significant increase in intracellular calcium levels. In vitro, SDF-1 alpha mediated dose-dependent migration of human cord blood-derived mast cells and HMC-1 cells across HUVEC monolayers. Although SDF-1 alpha did not induce mast cell degranulation, it selectively stimulated production of the neutrophil chemoattractant IL-8 without affecting TNF-alpha, IL 1beta, IL-6, GM-CSF, IFN-gamma, or RANTES production, providing further evidence of the selective modulation of mast cell function by this chemokine. These findings provide a novel, SDF-1-dependent mechanism for mast cell transendothelial migration and functional regulation, which may have important implications for the local regulation of mast cells in disease. PMID- 10861055 TI - The polarization defect of Wiskott-Aldrich syndrome macrophages is linked to dislocalization of the Arp2/3 complex. AB - Wiskott-Aldrich syndrome (WAS) is an X-linked recessive disorder originally characterized by the clinical triad eczema, thrombocytopenia, and severe immunodeficieny, with recurrent bacterial and viral infections, indicating a profound immune cell defect. Such altered immune cells include monocytes, macrophages, and dendritic cells, which were reported to display disturbed cell polarization or chemotaxis. WAS is caused by mutations in the WAS protein (WASp), which is thought to organize the actin cytoskeleton through the Arp2/3 complex. Here we show that the Arp2/3 complex is an integral part of podosomes, actin-rich adhesion structures of macrophages, and that WAS macrophages fail to organize the Arp2/3 complex into podosomes. We also demonstrate that microinjection of a C terminal acidic stretch of WASp into normal macrophages displaces Arp2/3 from podosomes and, in combination with chemoattractant stimulation of cells, induces a phenotype resembling the polarization-defective phenotype of stimulated WAS macrophages. These findings point to an important role of the Arp2/3 complex in polarization and migration of immune cells. PMID- 10861056 TI - Microchimerism, donor dendritic cells, and alloimmune reactivity in recipients of Flt3 ligand-mobilized hemopoietic cells: modulation by tacrolimus. AB - Flt3 ligand (FL) is a potent hemopoietic growth factor that strikingly enhances stem cells and dendritic cells (DC) in vivo. We examined the impact of infusing FL-mobilized bone marrow (BM) cells on microchimerism and anti-donor reactivity in normal and tacrolimus-immunosuppressed, noncytoablated allogeneic recipients. BM from B10 (H2b) mice given FL (10 microg/day; days 0-8; FL-BM) contained a 7 fold higher incidence of potentially tolerogenic immature CD11c+ DC (CD40low, CD80low, CD86low, MHC IIlow) that induced alloantigen-specific T cell hyporesponsiveness in vitro. C3H (H2k) mice received 50 x 106 normal or FL-BM cells (day 0) and tacrolimus (2 mg/kg/day; days 0-12). On day 15, enhanced numbers of donor (IAb+) cells were detected in the thymi and spleens of FL-BM recipients. Tacrolimus markedly enhanced microchimerism, which declined as a function of time. Ex vivo splenocyte proliferative and CTL responses and Th1 cytokine (IFN-gamma) production in response to donor alloantigens were augmented by FL-BM infusion, but reduced by tacrolimus. Systemic infusion of purified FL-BM immature DC, equivalent in number to that in corresponding whole BM, confirmed their capacity to sensitize, rather than tolerize, recipient T cells in vivo. In vitro, tacrolimus suppressed GM-CSF-stimulated growth of myeloid DC from normal BM much more effectively than from FL-BM without affecting MHC class II or costimulatory molecule expression. Infusion of normal B10 BM cells at the time of transplant prolonged C3H heart allograft survival, whereas FL-BM cells did not. A therapeutic effect of tacrolimus on graft survival was observed in combination with normal, but not FL-BM cells. These findings suggest the need for alternative immunosuppressive strategies to calcineurin inhibition to enable the engraftment, survival, and immunomodulatory function of FL-enhanced, immature donor DC. PMID- 10861057 TI - Macrophage-derived chemokine and EBI1-ligand chemokine attract human thymocytes in different stage of development and are produced by distinct subsets of medullary epithelial cells: possible implications for negative selection. AB - The chemoattractant activity of macrophage-derived chemokine (MDC), EBI1-ligand chemokine (ELC), and secondary lymphoid tissue chemokine (SLC) on human thymocytes was analyzed. Both ELC and SLC caused the accumulation of CD4+CD8- or CD4-CD8+ CD45RA+ thymocytes showing high CD3 expression. By contrast, a remarkable proportion of MDC-responsive thymocytes were CD4+CD8+ cells exhibiting reduced levels of CD8 or CD4+CD8- cells showing CD3 and CD45R0, but not CD45RA. MDC-responsive thymocyte suspensions were enriched in cells expressing the MDC receptor, CCR4, selectively localized to the medulla, and in CD30+ cells, whereas ELC-responsive thymocytes never expressed CD30. Reactivity to both MDC and ELC was localized to cells of the medullary areas, but never in the cortex. Double immunostaining showed no reactivity for either MDC or ELC by T cells, macrophages, or mature dendritic cells, whereas many medullary epithelial cells were reactive to MDC or ELC. However, MDC reactivity was consistently localized to the outer wall of Hassal's corpuscles, whereas ELC reactivity was often found in cells surrounding medullary vessels, but not in Hassal's corpuscles. Moreover, while most MDC-producing cells also stained positive for CD30L, this molecule was never found on ELC-producing cells. We suggest therefore that CD30L-expressing MDC-producing medullary epithelial cells attract CCR4-expressing thymocytes, thus favoring the CD30/CD30L interaction, and therefore the apoptosis, of cells that are induced to express CD30 by autoantigen activation. By contrast, ELC production by CD30L-lacking medullary epithelial cells may induce the migration into periphery of mature thymocytes that have survived the process of negative selection. PMID- 10861058 TI - An essential contribution by IFN-gamma to CD8+ T cell-mediated rejection of pancreatic islet allografts. AB - CD8+ T cells have long been considered to be the prototypical cytotoxic lymphocyte subpopulation. However, whether alloreactive CD8+ T cells require traditional cytolytic pathways such as perforin and Fas ligand (FasL) to mediate graft rejection has been a controversial issue. In the present studies, we examined the role of varied effector pathways in CD8+ T cell-mediated rejection of pancreatic islet allografts. Our goal was to systematically determine the relative requirements, if any, of perforin and FasL as well as the proinflammatory cytokine IFN-gamma in triggering graft destruction. To study CD8+ T cell effector pathways independently of other lymphocyte populations, purified alloreactive CD8+ T cells were adoptively transferred into severe combined immune deficient (SCID) recipients bearing established islet allografts. Results indicate that to reject established islet allografts, primed CD8+ T cells do not require the individual action of the conventional cytotoxic effectors perforin and Fas ligand. In contrast, the ability to produce IFN-gamma is critical for efficient CD8+ T cell-mediated rejection of established islet allografts. Furthermore, alloreactive CD8+ TCR transgenic T cells (2C) also show IFN-gamma dependence for mediating islet allograft rejection in vivo. We speculate from these results that the production of IFN-gamma by alloreactive CD8+ T cells is a rate-limiting step in the process of islet allograft rejection. PMID- 10861059 TI - TCR/CD3-Induced activation and binding of Emt/Itk to linker of activated T cell complexes: requirement for the Src homology 2 domain. AB - Expressed in mast and T cells/inducible T cell tyrosine kinase (Emt/Itk), a Tec family protein tyrosine kinase, is critical for the development and activation of T lymphocytes. The mechanism through which Emt/Itk mediates its effector functions is poorly understood. In this study, we show that the Emt/Itk Src homology 2 (SH2) domain is critical for the transphosphorylation and activation of Emt/Itk catalytic activity that is mediated by TCR/CD3 engagement. Furthermore, we find that the Emt/Itk SH2 domain is essential for the formation of TCR/CD3-inducible Emt/Itk-LAT complexes, whereas the SH3 domain and catalytic activity are not required. The Emt/Itk-linker of activated T cells (LAT) complexes are biologically important because Jurkat T cells with deficient LAT expression (JCaM2) fail to increase Emt/Itk tyrosine phosphorylation upon TCR/CD3 stimulation. Confocal microscopy reveals that in activated cells, LAT complexes colocalize with TCR/CD3. The present data suggest that upon TCR/CD3 engagement, the Emt/Itk SH2 domain mediates the formation of a molecular complex containing Emt/Itk, LAT, and TCR/CD3; this complex is essential for Emt/Itk activation and function. PMID- 10861060 TI - The HTLV-I tax protein transcriptionally modulates OX40 antigen expression. AB - OX40 is a member of the TNF receptor family, expressed on activated T cells. It is the only costimulatory T cell molecule known to be specifically up-regulated in human T cell leukemia virus type-I (HTLV-I)-producing cells. In a T cell line, OX40 surface expression was shown to be induced by HTLV-I Tax alone. To understand molecular mechanisms of OX40 gene regulation and modulation by HTLV-I Tax, we have cloned the human OX40 gene and analyzed its 5'-flanking region. By reporter gene analysis with progressive 5' deletions from nucleotides -1259 to 64, we have defined a 157-bp DNA fragment as a minimal promoter for constitutive expression. In addition, we show that in the OX40+ cell line, Co, Tax is able to further increase OX40 surface expression. Up-regulation of OX40 promoter activity by Tax requires two upstream NF-kappaB sites, which are not active in the constitutive OX40 expression. Their deletion abrogates Tax responsiveness in reporter gene analysis. The site-directed mutagenesis of each NF-kappaB site demonstrates that cooperative NF-kappaB binding is a prerequisite for Tax directed activity as neither site alone is sufficient for a full Tax responsiveness of the OX40 promoter. Upon Tax expression, both sites bind p65 and c-Rel. These data provide new insight into the direct regulation of OX40 by Tax and add to our understanding of the possible role of the OX40/OX40 ligand system in the proliferation of HTLV-I+ T cells. PMID- 10861061 TI - An IFN-gamma-inducible transcription factor, IFN consensus sequence binding protein (ICSBP), stimulates IL-12 p40 expression in macrophages. AB - IL-12 is a cytokine that links innate and adaptive immunity. Its subunit p40 is induced in macrophages following IFN-gamma/LPS stimulation. Here we studied the role for IFN consensus sequence binding protein (ICSBP), an IFN-gamma/LPS inducible transcription factor of the IFN regulatory factor (IRF) family in IL-12 p40 transcription. Macrophage-like cells established from ICSBP-/- mice did not induce IL-12 p40 transcripts, nor stimulated IL-12 p40 promoter activity after IFN-gamma/LPS stimulation, although induction of other inducible genes was normal in these cells. Transfection of ICSBP led to a marked induction of both human and mouse IL-12 p40 promoter activities in ICSBP+/+ and ICSBP-/- cells, even in the absence of IFN-gamma/LPS stimulation. Whereas IRF-1 alone was without effect, synergistic enhancement of promoter activity was observed following cotransfection of ICSBP and IRF-1. Deletion analysis of the human promoter indicated that the Ets site, known to be important for activation by IFN gamma/LPS, also plays a role in the ICSBP activation of IL-12 p40. A DNA affinity binding assay revealed that endogenous ICSBP is recruited to the Ets site through protein-protein interaction. Last, transfection of ISCBP alone led to induction of the endogenous IL-12 p40 mRNA in the absence of IFN-gamma and LPS. Taken together, our results show that ICSBP induced by IFN-gamma/LPS, acts as a principal activator of IL-12p40 transcription in macrophages. PMID- 10861062 TI - Altered peptide ligand-mediated TCR antagonism can be modulated by a change in a single amino acid residue within the CDR3 beta of an MHC class I-restricted TCR. AB - The Ag receptor of cytotoxic CD8+ T lymphocytes recognizes peptides of 8-10 aa bound to MHC class I molecules. This Ag recognition event leads to the activation of the CD8+ lymphocyte and subsequent lysis of the target cell. Altered peptide ligands are analogues derived from the original antigenic peptide that commonly carry amino acid substitutions at TCR contact residues. TCR engagement by these altered peptide ligands usually impairs normal T cell function. Some of these altered peptide ligands (antagonists) are able to specifically antagonize and inhibit T cell activation induced by the wild-type antigenic peptide. Despite significant advances made in understanding TCR antagonism, the molecular interactions between the TCR and the MHC/peptide complex responsible for the inhibitory activity of antagonist peptides remain elusive. To approach this question, we have identified altered peptide ligands derived from the vesicular stomatitis virus peptide (RGYVYQGL) that specifically antagonize an H 2Kb/vesicular stomatitis virus-specific TCR. Furthermore, by site-directed mutagenesis, we altered single amino acid residues of the complementarity determining region 3 of the beta-chain of this TCR and tested the effect of these point mutations on Ag recognition and TCR antagonism. Here we show that a single amino acid change on the TCR CDR3 beta loop can modulate the TCR-antagonistic properties of an altered peptide ligand. Our results highlight the role of the TCR complementarity-determining region 3 loops for controlling the nature of the T cell response to TCR/altered peptide ligand interactions, including those leading to TCR antagonism. PMID- 10861063 TI - IL-10 gene expression is controlled by the transcription factors Sp1 and Sp3. AB - IL-10 is an 18-kDa cytokine with a key role in homeostatic control of inflammatory and immune responses. We have investigated how transcription of the IL-10 gene is regulated, so as to be able to understand the circumstances of IL 10 expression in both health and disease. In the mouse, IL-10 gene expression is regulated by a TATA-type promoter with a critical cis-acting element containing GGA repeats located at -89 to -77. Its complementary sequence is similar to the cis-acting elements (TCC repeats) in the promoters of genes encoding epidermal growth factor receptor and CD58. All these elements comprise a common CCTCCT sequence with less conserved C + T-rich sequences. Eliminating this CCTCCT sequence results in a marked reduction in promoter activity, suggesting a necessary role in IL-10 gene expression. Despite its dissimilarity to the G + C rich Sp1 consensus sequence (GC box), Sp1 and Sp3 transcription factors could be shown to bind to this motif. The requirement for Sp1 and Sp3 in transcription of IL-10 was confirmed using Drosophila SL2 cells, which lack endogenous Sp factors. These results suggest that the transcription of IL-10 is positively regulated by both Sp1 and Sp3. PMID- 10861064 TI - Posttranscriptional regulation of IL-10 gene expression through sequences in the 3'-untranslated region. AB - IL-10 is an 18-kDa immunoregulatory cytokine the transcription of which is controlled by the ubiquitously expressed transcription factors Sp1 and Sp3. Although many cell types express IL-10 mRNA, not all make detectable amounts of protein, and levels of protein expression vary enormously. We show here that much of this variation can be accounted for by posttranscriptional mechanisms. Multiple copies of potential mRNA destabilizing motifs AUUUA and related sequences can be found to the 3'-untranslated region (UTR) of IL-10 mRNA distributed through three potential regulatory regions. Evidence of RNA destabilizing activities in all three regions was deduced from luciferase reporter assays. The half-life of RNA containing the 3'-UTR of IL-10 mRNA was quite short in both nonstimulated (t1/2 = 1 h), and PMA-stimulated EL-4 cell (t1/2 = 3 h). In contrast, the half-life of RNA lacking the 3'-UTR was much longer (t1/2 = >12 h) whether cells were stimulated or not. This suggests that many cells are poised to secrete IL-10 and will do so if they receive appropriate posttranscriptional signals. PMID- 10861065 TI - The duration of nuclear residence of NFAT determines the pattern of cytokine expression in human SCID T cells. AB - The expression of cytokine genes and other inducible genes is crucially dependent on the pattern and duration of signal transduction events that activate transcription factor binding to DNA. Two infant patients with SCID and a severe defect in T cell activation displayed an aberrant regulation of the transcription factor NFAT. Whereas the expression levels of the NFAT family members NFAT1, -2, and -4 were normal in the patients' T cells, dephosphorylation and nuclear translocation of these NFAT proteins occurred very transiently and incompletely upon stimulation. Only after inhibition of nuclear export with leptomycin B were we able to demonstrate a modest degree of nuclear translocation in the patients' T cells. This transient activation of NFAT was not sufficient to induce the expression of several cytokines, including IL-2, IL-3, IL-4, and IFN-gamma, whereas mRNA levels for macrophage inflammatory protein-1alpha, GM-CSF, and IL-13 were only moderately reduced. By limiting the time of NFAT activation in normal control cells using the calcineurin inhibitor cyclosporin A, we were able to mimic the cytokine expression pattern in SCID T cells, suggesting that the expression of different cytokine genes is differentially regulated by the duration of NFAT residence in the nucleus. PMID- 10861066 TI - Members of the Ikaros gene family are present in early representative vertebrates. AB - Members of the Ikaros multigene family of zinc finger proteins are expressed in a tissue-specific manner and most are critical determinants in the development of both the B and T lymphocytes as well as NK and dendritic APC lineages. A PCR amplification strategy that is based on regions of shared sequence identity in Ikaros multigene family members found in mammals and several other vertebrates has led to the recovery of cDNAs that represent the orthologues of Ikaros, Aiolos, Helios, and Eos in Raja eglanteria (clearnose skate), a cartilaginous fish that is representative of an early divergence event in the phylogenetic diversification of the vertebrates. The tissue-specific patterns of expression for at least two of the four Ikaros family members in skate resemble the patterns observed in mammals, i.e., in hematopoietic tissues. Prominent expression of Ikaros in skate also is found in the lymphoid Leydig organ and epigonal tissues, which are unique to cartilaginous fish. An Ikaros-related gene has been identified in Petromyzon marinus (sea lamprey), a jawless vertebrate species, in which neither Ig nor TCRs have been identified. In addition to establishing a high degree of evolutionary conservation of the Ikaros multigene family from cartilaginous fish through mammals, these studies define a possible link between factors that regulate the differentiation of immune-type cells in the jawed vertebrates and related factors of unknown function in jawless vertebrates. PMID- 10861067 TI - Ly-6I, a new member of the murine Ly-6 superfamily with a distinct pattern of expression. AB - A new member of the mouse Ly-6SF, designated Ly-6I, has been isolated as a gene homologous to a segment of the Ly-6C gene. A single allelic difference in the mature protein sequence was identified, which is similar to other Ly-6SF members. Ly-6I mRNA has been detected in a wide range of tissues and cell lines, and a rabbit polyclonal Ab has been used to determine that Ly-6I protein is present at a low constitutive level on cell lines from several different lineages. In contrast to Ly-6C and Ly-6A/E, the Ly-6I gene is only weakly responsive to IFNs. Expression in vivo is most abundant on bone marrow populations and is coexpressed with Ly-6C on granulocytes and macrophages. However, Ly-6I is also expressed on immature B cell populations that do not express Ly-6C. Expression on mature B cells in spleen is uniformly low. Similarly, Ly-6I is expressed on TCRlow/int, but not TCRhigh, thymocytes. Ly-6I is re-expressed on Ly-6Chigh T cells in the periphery. Thus, Ly-6I may be a useful marker to define maturation stages of both T and B lymphocytes as well as subsets of monocytes and granulocytes. PMID- 10861068 TI - Tapasin-mediated retention and optimization of peptide ligands during the assembly of class I molecules. AB - The murine class I H-2Kb molecule achieves high level surface expression in tapasin-deficient 721.220 human cells. Compared with their behavior in wild-type cells, Kb molecules expressed on 721.220 cells are more receptive to exogenous peptide, undergo more rapid surface decay, and fail to form macromolecular peptide loading complexes. As a result, they are rapidly transported to the cell surface, reflecting a failure of endoplasmic reticulum retention mechanisms in the absence of loading complex formation. Despite the failure of Kb molecules to colocalize to the TAP and their rapid egress to the cell surface, Kb is still capable of presenting TAP-dependent peptides in the absence of tapasin. Furthermore, pool sequencing of peptides eluted from these molecules revealed strict conservation of their canonical H-2Kb-binding motif. There was a reduction in the total recovery of peptides associated with Kb molecules purified from the surface of tapasin-deficient cells. Comparison of the peptides bound to Kb in the presence and absence of tapasin revealed considerable overlap in peptide repertoire. These results indicate that in the absence of an interaction with tapasin, Kb molecules fail to assemble with calreticulin and TAP, yet they are still capable of acquiring a diverse array of peptides. However, a significant proportion of these peptides appear to be suboptimal, resulting in reduced cell surface stability of Kb complexes. Taken together, the findings indicate that tapasin plays an essential role in the formation of the class I loading complex, which retains class I heterodimers in the endoplasmic reticulum until optimal ligand selection is completed. PMID- 10861069 TI - Dominant epitopes and allergic cross-reactivity: complex formation between a Fab fragment of a monoclonal murine IgG antibody and the major allergen from birch pollen Bet v 1. AB - The symptoms characteristic of allergic hypersensitivity are caused by the release of mediators, i.e., histamine, from effector cells such as basophils and mast cells. Allergens with more than one B cell epitope cross-link IgE Abs bound to high affinity FcepsilonRI receptors on mast cell surfaces leading to aggregation and subsequent mediator release. Thus, allergen-Ab complexes play a crucial role in the cascade leading to the allergic response. We here report the structure of a 1:1 complex between the major birch pollen allergen Bet v 1 and the Fab fragment from a murine monoclonal IgG1 Ab, BV16, that has been solved to 2.9 A resolution by x-ray diffraction. The mAb is shown to inhibit the binding of allergic patients' IgE to Bet v 1, and the allergen-IgG complex may therefore serve as a model for the study of allergen-IgE interactions relevant in allergy. The size of the BV16 epitope is 931 A2 as defined by the Bet v 1 Ab interaction surface. Molecular interactions predicted to occur in the interface are likewise in agreement with earlier observations on Ag-Ab complexes. The epitope is formed by amino acids that are conserved among major allergens from related species within the Fagales order. In combination with a surprisingly high inhibitory capacity of BV16 with respect to allergic patients' serum IgE binding to Bet v 1, these observations provide experimental support for the proposal of dominant IgE epitopes located in the conserved surface areas. This model will facilitate the development of new and safer vaccines for allergen immunotherapy in the form of mutated allergens. PMID- 10861070 TI - Ascaris suum, an intestinal parasite, produces morphine. AB - The parasitic worm Ascaris suum contains the opiate alkaloid morphine as determined by HPLC coupled to electrochemical detection and by gas chromatography/mass spectrometry. The level of this material is 1168 +/- 278 ng/g worm wet weight. Furthermore, Ascaris maintained for 5 days contained a significant amount of morphine, as did their medium, demonstrating their ability to synthesize the opiate alkaloid. To determine whether the morphine was active, we exposed human monocytes to the material, and they immediately released nitric oxide in a naloxone-reversible manner. The anatomic distribution of morphine immunoreactivity reveals that the material is in the subcuticle layers and in the animals' nerve chords. Furthermore, as determined by RT-PCR, Ascaris does not express the transcript of the neuronal mu receptor. Failure to demonstrate the expression of this opioid receptor, as well as the morphine-like tissue localization in Ascaris, suggests that the endogenous morphine is intended for secretion into the microenvironment. PMID- 10861071 TI - Senescent jejunal mast cells and eosinophils in the mouse preferentially translocate to the spleen and draining lymph node, respectively, during the recovery phase of helminth infection. AB - Because mice infected with Trichinella spiralis experience a pronounced, but transient, mastocytosis and eosinophilia in their intestine, this disease model was used to follow the fate of senescent T cell-dependent mast cells (MCs) and eosinophils. Very few MCs or eosinophils undergoing apoptosis were found in the jejunum during the resolution phase of the infection, even though apoptotic MCs were common in the large intestine. Although the mesenteric draining lymph nodes contained large numbers of apoptotic eosinophils, MCs were rarely found at this location. During the recovery phase, large numbers of MCs were present in the spleen, and many of these cells possessed segmented nuclei. These splenic MCs were not proliferating. Although MCs from the jejunum and spleen of noninfected mice failed to express mouse MC protease (mMCP) 9, essentially all of the MCs in the jejunal submucosa and spleen of T. spiralis-infected mice expressed this serine protease during the recovery phase. The MCs in the jejunum expressed mMCP 9 before any mMCP-9-containing cells could be detected in the spleen. The fact that mMCP-9-containing MCs were detected in splenic blood vessels as these cells began to disappear from the jejunum supports the view that many jejunal MCs translocate to the spleen during the recovery phase of the infection. During this translocation process, some senescent jejunal MCs undergo nuclear segmentation. These studies reveal for the first time different exit and disposal pathways for T cell-dependent eosinophils and MCs after their expansion in the jejunum during a helminth infection. PMID- 10861072 TI - CD8+ CTL from lungs of Mycobacterium tuberculosis-infected mice express perforin in vivo and lyse infected macrophages. AB - CD8+ T lymphocytes have been implicated in the protective immune response against human and murine tuberculosis. However, the functional role that this cell subset plays during the resolution of infection remains controversial. In this study, we demonstrate the presence of Mycobacterium tuberculosis-specific CD8+ CTL in the lungs and lung-draining lymph nodes of mice infected with M. tuberculosis via the aerosol or i.v. route. These cells expressed perforin in vivo and specifically recognized and lysed M. tuberculosis-infected macrophages in a perforin-dependent manner after a short period of in vitro restimulation. The efficiency of lysis of infected macrophages was dependent upon the time allowed for interaction between macrophage and M. tuberculosis bacilli. Recognition of infected targets by CD8+ CTL was beta 2-microglobulin and MHC class I dependent and was not CD1d restricted. The presented data indicate that CD8+ T cells contribute to the protective immune response during M. tuberculosis infection by exerting cytotoxic function and lysing infected macrophages. PMID- 10861073 TI - IL-4-independent inhibition of IL-12 responsiveness during Leishmania amazonensis infection. AB - Leishmania amazonensis induces a nonhealing infection in C3H mice, whereas infection with Leishmania major is self-healing. We found that C3H mice infected with L. amazonensis exhibited decreased IL-12 production, which could account for the susceptibility to this organism. However, exogenous IL-12 administration failed to induce a healing immune response. The failure of L. amazonensis infected C3H mice to respond to IL-12 was associated with a specific defect in IL 12 receptor beta2 (IL-12Rbeta2) mRNA expression by CD4+ T cells. Furthermore, decreased IL-12Rbeta2 mRNA expression correlated with a decrease in the IL-12 signaling capacity of the lymph node (LN) cells. IL-4 did not contribute to susceptibility or down-regulation of the IL-12Rbeta2 subunit, because IL-4-/- mice remained susceptible to L. amazonensis infection, even after IL-12 administration, and CD4+ cells from infected IL-4-/- mice also had reduced expression of IL-12Rbeta2 mRNA. These results demonstrate that regulation of the IL-12 receptor, independent of IL-4, is a point of control for the immune response to leishmaniasis. In contrast to experimental L. major infections, where host genetics control susceptibility, these studies demonstrate that the lack of IL-12 responsiveness may be dictated by the pathogen, rather than the host. PMID- 10861074 TI - Delta-9-tetrahydrocannabinol inhibits antitumor immunity by a CB2 receptor mediated, cytokine-dependent pathway. AB - In this study, we show that Delta-9-tetrahydrocannabinol (THC), the major psychoactive component of marijuana, suppresses host immune reactivity against lung cancer. In two different weakly immunogenic murine lung cancer models, intermittent administration of THC (5 mg/kg, four times/wk i.p. for 4 wk) led to accelerated growth of tumor implants compared with treatment with diluent alone. In contrast to our findings in immunocompetent mice, THC did not affect tumor growth in tumor-bearing SCID mice. The immune inhibitory cytokines, IL-10 and TGF beta, were augmented, while IFN-gamma was down-regulated at both the tumor site and in the spleens of THC-treated mice. Administration of either anti-IL-10- or anti-TGF-beta-neutralizing Abs prevented the THC-induced enhancement in tumor growth. Both APC and T cells from THC-treated mice showed limited capacities to generate alloreactivity. Furthermore, lymphocytes from THC-treated mice transferred the effect to normal mice, resulting in accelerated tumor growth similar to that seen in the THC-treated mice. THC decreased tumor immunogenicity, as indicated by the limited capacity for tumor-immunized, THC-treated mice to withstand tumor rechallenge. In vivo administration of a specific antagonist of the CB2 cannabinoid receptor also blocked the effects of THC. Our findings suggest the THC promotes tumor growth by inhibiting antitumor immunity by a CB2 receptor-mediated, cytokine-dependent pathway. PMID- 10861075 TI - T cell vaccination in mice with invasive pulmonary aspergillosis. AB - Aspergillus fumigatus, an opportunistic fungal pathogen, is responsible for multiple airway diseases of an allergic and a nonallergic nature. In a murine model of invasive pulmonary aspergillosis, resistance is associated with a decreased lung inflammatory pathology and the occurrence of an IL-12-dependent Th1-type reactivity that are both impaired by IL-4. In the present study we assess the ability of Aspergillus crude culture filtrate Ags and the recombinant allergen Asp f 2 to induce protective antifungal responses in mice with invasive pulmonary aspergillosis. Similar to what occurred upon nasal exposure to viable A. fumigatus conidia, treatment of immunocompetent mice with Aspergillus crude culture filtrate Ags resulted in the development of local and peripheral protective Th1 memory responses, mediated by Ag-specific CD4+ T cells producing IFN-gamma and IL-2 capable of conferring protection upon adoptive transfer to naive recipients. Protective Th1 responses could not be observed in mice deficient of IFN-gamma or IL-12 and did not occur in response to Asp f 2, which, on the contrary, elicited high level production of inhibitory IL-4. The results show that Ags of Aspergillus exist with the ability to induce both Th1- and Th2 type reactivity during infection, a finding that suggests a possible mechanism through which potentially protective immune responses are inhibited in mice with the infection. However, the occurrence of Th1-mediated resistance upon vaccination with Aspergillus crude culture filtrate Ags, suggests the existence of fungal Ags useful as a candidate vaccine against invasive pulmonary aspergillosis. PMID- 10861076 TI - CD4+ T cells acting independently of antibody contribute to protective immunity to Plasmodium chabaudi infection after apical membrane antigen 1 immunization. AB - Apical membrane Ag 1 (AMA1) is a leading malaria vaccine candidate. Homologues of AMA1 can induce protection in mice and monkeys, but the mechanism of immunity is not understood. Mice immunized with a refolded, recombinant, Plasmodium chabaudi AMA1 fragment (AMA1B) can withstand subsequent challenge with P. chabaudi adami. Here we show that CD4+ T cell depletion, but not gammadelta T cell depletion, can cause a significant drop in antiparasite immunity in either immunized normal or immunized B cell KO mice. In normal mice, this loss of immunity is not accompanied by a decline in Ab levels. These observations indicate a role for AMA1-specific Ab-independent T cell-mediated immunity. However, the loss of immunity in normal CD4+ T cell-depleted mice is temporary. Furthermore, immunized B cell KO mice cannot survive infection, demonstrating the absolute importance of B cells, and presumably Ab, in AMA1-induced immunity. CD4+ T cells specific for a cryptic conserved epitope on AMA1 can adoptively transfer protection to athymic (nu/nu) mice, the level of which is enhanced by cotransfer of rabbit anti-AMA1 specific antisera. Recipients of rabbit antisera alone do not survive. Some protected recipients of T cells plus antisera do not develop their own AMA 1 specific Ab response, suggesting that AMA 1-specific CMI alone can protect mice. These data are the first to demonstrate the specificity of any protective CMI response in malaria and have important implications for developing a malaria vaccine. PMID- 10861077 TI - The chemokine fractalkine inhibits Fas-mediated cell death of brain microglia. AB - Fractalkine is a CX3C-family chemokine, highly and constitutively expressed on the neuronal cell surface, for which a clear CNS physiological function has yet to be determined. Its cognate receptor, CX3CR-1, is constitutively expressed on microglia, the brain-resident macrophages; however, these cells do not express fractalkine. We now show that treatment of microglia with fractalkine maintains cell survival and inhibits Fas ligand-induced cell death in vitro. Biochemical characterization indicates that this occurs via mechanisms that may include 1) activation of the phosphatidylinositol-3 kinase/protein kinase B pathway, resulting in phosphorylation and blockade of the proapoptotic functions of BAD; 2) up-regulation of the antiapoptotic protein Bcl-xL; and 3) inhibition of the cleavage of BH3-interacting domain death agonist (BID). The observation that fractalkine serves as a survival factor for primary microglia in part by modulating the protein levels and the phosphorylation status of Bcl-2 family proteins reveals a novel physiological role for chemokines. These results, therefore, suggest that the interaction between fractalkine and CX3CR-1 may play an important role in promoting and preserving microglial cell survival in the CNS. PMID- 10861078 TI - Expression of P-selectin at low site density promotes selective attachment of eosinophils over neutrophils. AB - The selective interaction of neutrophils with E-selectin and eosinophils with P selectin has been previously reported, but the relevance of selectin site density and fluid shear has not been studied in detail. We have developed a new approach to examine these interactions in cell suspensions that integrates an on-line cone plate viscometer with a flow cytometer. We find that eosinophils and neutrophils both use P-selectin glycoprotein ligand-1 to form stable conjugates with P selectin Chinese hamster ovary cell transfectants, with a preferential adhesion of eosinophils. Further, the difference in cell adhesion between neutrophils and eosinophils is magnified at P-selectin expression levels below approximately 20 sites/microm2, a range likely to be relevant to endothelial cell expression levels in conditions associated with eosinophilia. The unique behavior is retained over shear rates ranging from 100 to 1500/s but is magnified at low shear. Results from parallel-plate flow chamber assays suggest that preferential eosinophil adhesion reflects an enhanced efficiency of initial PSGL-1 bond formation with P-selectin rather than a unique ability of eosinophils to mediate rolling interactions of longer duration on low-density P-selectin substrates. These differences may account in part for the increase in eosinophil accumulation in allergic diseases. PMID- 10861079 TI - Proteolysis of human monocyte CD14 by cysteine proteinases (gingipains) from Porphyromonas gingivalis leading to lipopolysaccharide hyporesponsiveness. AB - Cysteine proteinases (gingipains) elaborated from Porphyromonas gingivalis exhibit enzymatic activities against a broad range of host proteins and are considered key virulence factors in the onset and development of adult periodontitis and host defense evasion. In this study, we examined the ability of arginine-specific gingipains (high molecular mass Arg-specific gingipain (HRGP) and Arg-specific gingipain 2) and lysine-specific gingipain (KGP) to cleave monocyte CD14, the main receptor for bacterial cell surface components such as LPS. Binding of anti-CD14 mAb MY4 to human monocytes was almost completely abolished by 0.3 microM HRGP and KGP treatments for 15 min, and 1 microM RGP2 for 30 min. In contrast, the expressions of Toll-like receptor 4, and CD18, CD54, CD59, and HLA-A, -B, -C on monocytes were slightly increased and decreased, respectively, by 0. 3 microM HRGP and KGP. This down-regulation resulted from direct proteolysis, because 1) gingipains eliminated MY4 binding even to fixed monocytes, and 2) CD14 fragments were detected in the extracellular medium by immunoblot analysis. Human rCD14 was degraded by all three gingipains, which confirmed that CD14 was a substrate for gingipains. TNF-alpha production by monocytes after HRGP and KGP treatments was decreased at 1 ng/ml, but not at 20 microg/ml LPS, indicating that gingipains inhibited a CD14-dependent cell activation. These results suggest that gingipains preferentially cleave monocyte CD14, resulting in attenuation of the cellular recognition of bacteria, and as a consequence sustain chronic inflammation. PMID- 10861080 TI - Group B Streptococcus induces TNF-alpha gene expression and activation of the transcription factors NF-kappa B and activator protein-1 in human cord blood monocytes. AB - It has been postulated that production of TNF-alpha is central to the pathogenesis of septic shock induced by group B Streptococcus (GBS). In vitro studies using human cord blood monocytes have demonstrated that GBS induces TNF alpha secretion, but little is known about the intracellular signaling pathways of TNF-alpha induction. In this report we show that heat-killed serotype III GBS induces host cell signal transduction pathways that lead to activation of the transcription factors NF-kappaB and AP-1. Using adenoviral transfer of IkappaBalpha (IkappaBalpha overexpression), the production of TNF-alpha induced by whole GBS was inhibited by only 20%. We also show that the p38 mitogen activated protein kinase (MAPK) pathway is involved in GBS-induced TNF-alpha secretion, because TNF-alpha protein and mRNA levels in the presence of a specific inhibitor of p38 MAPK, SB 202190, were dramatically diminished. EMSAs showed that SB 202190 inhibited GBS-induced AP-1 activation, but had no effect on NF-kappaB-DNA binding activity. These results indicate that both NF-kappaB and AP 1 (via p38 MAPK) are involved in the regulation of TNF-alpha production in GBS stimulated neonatal monocytes. Therefore, disrupting the signal transduction pathways induced by GBS has the potential to attenuate the production of immune response mediators, thereby halting or possibly reversing the course of this potentially fatal disease. PMID- 10861081 TI - Divergent effects of platelet-endothelial cell adhesion molecule-1 and beta 3 integrin blockade on leukocyte transmigration in vivo. AB - The final stage in the migration of leukocytes to sites of inflammation involves movement of leukocytes through the endothelial cell layer and the perivascular basement membrane. Both platelet-endothelial cell adhesion molecule-1 (PECAM 1/CD31) and the integrin alphavbeta3 have been implicated in this process, and in vitro studies have identified alphavbeta3 as a heterotypic ligand for PECAM-1. In the present study we have addressed the roles of these molecules by investigating and comparing the effects of PECAM-1 and alphavbeta3 blockade on leukocyte migration in vivo. For this purpose we have examined the effects of neutralizing Abs directed against PECAM-1 (domain 1-specific, mAb 37) and beta3 integrins (mAbs 7E3 and F11) on leukocyte responses in the mesenteric microcirculation of anesthetized rats using intravital microscopy. The anti-PECAM-1 mAb suppressed leukocyte extravasation, but not leukocyte rolling or firm adhesion, elicited by IL-1beta in a dose-dependent manner (e.g., 67% inhibition at 10 mg/kg 37 Fab), but had no effect on FMLP-induced leukocyte responses. Analysis by electron microscopy suggested that this suppression was due to an inhibition of neutrophil migration through the endothelial cell barrier. By contrast, both anti-beta3 integrin mAbs, 7E3 F(ab')2 (5 mg/kg) and F11 F(ab')2 (5 mg/kg), selectively reduced leukocyte extravasation induced by FMLP (38 and 46%, respectively), but neither mAb had an effect on IL-1beta-induced leukocyte responses. These findings indicate roles for both PECAM-1 and beta3 integrins in leukocyte extravasation, but do not support the concept that these molecules act as counter-receptors in mediating leukocyte transmigration. PMID- 10861082 TI - Macrophage-induced neutrophil apoptosis. AB - Macrophages (Mphi) contribute to the resolution of early inflammation by recognizing and ingesting apoptotic polymorphonuclear neutrophils (PMN). In addition, experiments reported here demonstrated that Mphi can actively induce PMN apoptosis. Coculture of cells from 2- or 5-day-old wounds in rats, or of Mphi purified from such preparations, with PMN-rich wound cell populations obtained 1 day after wounding increased PMN apoptosis by >3-fold. Neither resident- nor Proprionibacterium acnes-elicited peritoneal Mphi-induced PMN apoptosis. Apoptosis was not mediated by a soluble factor and required E:T contact. Fixed wound-Mphi and membrane isolates from viable Mphi were as effective as intact cells in inducing PMN apoptosis. Mphi-induced apoptosis was inhibited by peptide Arg-Gly-Asp-Ser, anti-beta3 (CD61) Ab, CD36 peptide, or anti-TNF-alpha Ab. Soluble TNF-alpha did not induce PMN apoptosis. In additional studies, K562 cells (negative for beta3, TNF-alpha, and Fas ligand) transfected to express either alphavbeta3 integrin, an uncleavable membrane form of TNF-alpha, or both were used in cocultures with wound PMN. Only the double transfectants were able to induce PMN apoptosis, an effect inhibited by anti-beta3 (CD61) or anti-TNF-alpha Abs. These results demonstrate that wound Mphi induce PMN apoptosis through a constitutive effector mechanism requiring both intercellular binding through integrin-ligand interactions and membrane-bound TNF-alpha. PMID- 10861083 TI - The LFA-1 integrin supports rolling adhesions on ICAM-1 under physiological shear flow in a permissive cellular environment. AB - The LFA-1 integrin is crucial for the firm adhesion of circulating leukocytes to ICAM-1-expressing endothelial cells. In the present study, we demonstrate that LFA-1 can arrest unstimulated PBL subsets and lymphoblastoid Jurkat cells on immobilized ICAM-1 under subphysiological shear flow and mediate firm adhesion to ICAM-1 after short static contact. However, LFA-1 expressed in K562 cells failed to support firm adhesion to ICAM-1 but instead mediated K562 cell rolling on the endothelial ligand under physiological shear stress. LFA-1-mediated rolling required an intact LFA-1 I-domain, was enhanced by Mg2+, and was sharply dependent on ICAM-1 density. This is the first indication that LFA-1 can engage in rolling adhesions with ICAM-1 under physiological shear flow. The ability of LFA-1 to support rolling correlates with decreased avidity and impaired time dependent adhesion strengthening. A beta2 cytoplasmic domain-deletion mutant of LFA-1, with high avidity to immobilized ICAM-1, mediated firm arrests of K562 cells interacting with ICAM-1 under shear flow. Our results suggest that restrictions in LFA-1 clustering mediated by cytoskeletal attachments may lock the integrin into low-avidity states in particular cellular environments. Although low-avidity LFA-1 states fail to undergo adhesion strengthening upon contact with ICAM-1 at stasis, these states are permissive for leukocyte rolling on ICAM-1 under physiological shear flow. Rolling mediated by low-avidity LFA-1 interactions with ICAM-1 may stabilize rolling initiated by specialized vascular rolling receptors and allow the leukocyte to arrest on vascular endothelium upon exposure to stimulatory endothelial signals. PMID- 10861084 TI - Cyclic tensile strain acts as an antagonist of IL-1 beta actions in chondrocytes. AB - Inflammatory cytokines play a major role in cartilage destruction in diseases such as osteoarthritis and rheumatoid arthritis. Because physical therapies such as continuous passive motion yield beneficial effects on inflamed joints, we examined the intracellular mechanisms of mechanical strain-mediated actions in chondrocytes. By simulating the effects of continuous passive motion with cyclic tensile strain (CTS) on chondrocytes in vitro, we show that CTS is a potent antagonist of IL-1 beta actions and acts as both an anti-inflammatory and a reparative signal. Low magnitude CTS suppresses IL-1 beta-induced mRNA expression of multiple proteins involved in catabolic responses, such as inducible NO synthase, cyclo-oxygenase II, and collagenase. CTS also counteracts cartilage degradation by augmenting mRNA expression for tissue inhibitor of metalloproteases and collagen type II that are inhibited by IL-1 beta. Additionally, CTS augments the reparative process via hyperinduction of aggrecan mRNA expression and abrogation of IL-1 beta-induced suppression of proteoglycan synthesis. Nonetheless, the presence of an inflammatory signal is a prerequisite for the observed CTS actions, as exposure of chondrocytes to CTS alone has little effect on these parameters. Functional analysis suggests that CTS-mediated anti inflammatory actions are not mediated by IL-1R down-regulation. Moreover, as an effective antagonist of IL-1 beta, the actions of CTS may involve disruption/regulation of signal transduction cascade of IL-1 beta upstream of mRNA transcription. These observations are the first to show that CTS directly acts as an anti-inflammatory signal on chondrocytes and provide a molecular basis for its actions. PMID- 10861085 TI - The role of the CC chemokine, RANTES, in acute lung allograft rejection. AB - Lung transplantation is a therapeutic option for patients with end-stage lung disease. Acute allograft rejection is a major complication of lung transplantation and is characterized by the infiltration of activated mononuclear cells. The specific mechanisms that recruit these leukocytes have not been fully elucidated. The CC chemokine, RANTES, is a potent mononuclear cell chemoattractant. In this study we investigated RANTES involvement during acute lung allograft rejection in humans and in a rat model system. Patients with allograft rejection had a 2.3-fold increase in RANTES in their bronchoalveolar lavages compared with healthy allograft recipients. Rat lung allografts demonstrated a marked time-dependent increase in levels of RANTES compared with syngeneic control lungs. RANTES levels correlated with the temporal recruitment of mononuclear cells and the expression of RANTES receptors CCR1 and CCR5. To determine RANTES involvement in lung allograft rejection, lung allograft recipients were passively immunized with either anti-RANTES or control Abs. In vivo neutralization of RANTES attenuated acute lung allograft rejection and reduced allospecific responsiveness by markedly decreasing mononuclear cell recruitment. These experiments support the idea that RANTES, and the expression of its receptors have an important role in the pathogenesis of acute lung allograft rejection. PMID- 10861086 TI - Differential involvement of Src family kinases in Fc gamma receptor-mediated phagocytosis. AB - The tyrosine phosphorylation cascade originated from Fc gamma receptors (Fc gamma Rs) is essential for macrophage functions including phagocytosis. Although the initial step is ascribed to Src family tyrosine kinases, the role of individual kinases in phagocytosis signaling is still to be determined. In reconstitution experiments, we first showed that expression in the RAW 264.7 cell line of C terminal Src kinase (Csk) inhibited and that of a membrane-anchored, gain-of function Csk abolished the Fc gamma R-mediated signaling that leads to phagocytosis in a kinase-dependent manner. We next tested reconstruction of the signaling in the membrane-anchored, gain-of-function Csk-expressing cells by introducing Src family kinases the C-terminal negative regulatory sequence of which was replaced with a c-myc epitope. Those constructs derived from Lyn and Hck (a-Lyn and a-Hck) that associated with detergent-resistant membranes successfully reconstructed Fc gamma R-mediated Syk activation, filamentous actin rearrangement, and phagocytosis. In contrast, c-Src-derived construct (a-Src), that was excluded from detergent-resistant membranes, could not restore the series of phagocytosis signaling. Tyrosine phosphorylation of Vav and c-Cbl was restored in common by a-Lyn, a-Hck, and a-Src, but Fc gamma RIIB tyrosine phosphorylation, which is implicated in negative signaling, was reconstituted solely by a-Lyn and a-Hck. These findings suggest that Src family kinases are differentially involved in Fc gamma R-signaling and that selective kinases including Lyn and Hck are able to fully transduce phagocytotic signaling. PMID- 10861087 TI - Lethal granuloma disintegration in mycobacteria-infected TNFRp55-/- mice is dependent on T cells and IL-12. AB - Genetically susceptible, TNFRp55 gene-deficient (TNFRp55-/-) mice succumb to infection with Mycobacterium avium. Before their death, M. avium-infected TNFRp55 /- mice develop granulomatous lesions that, in contrast to granulomas in wild type syngeneic mice, undergo acute disintegration. To determine the factors involved in these events, we depleted T cell subsets or neutralized the inflammatory cytokines IFN-gamma, IL-12, or TNF in TNFRp55-/- mice infected i.v. with M. avium. Infected TNFRp55-/- mice treated with a control mAb became moribund between days 26 and 34 postinfection, showing widespread inflammatory cell apoptosis within disintegrating granulomas. In contrast, TNFRp55-/- mice depleted of either CD4+ or CD8+ cells after granuloma initiation stayed healthy until at least day 38 postinfection and showed no signs of granuloma destruction. Neutralization of IL-12, but not of IFN-gamma or TNF, also protected M. avium infected TNFRp55-/- mice from granuloma decomposition and from premature death. Treatment with dexamethasone or with a specific inhibitor of inducible NO synthase did not prevent granuloma dissolution or death of TNFRp55-/- mice. In conclusion, granuloma disintegration in TNFRp55-/- mice is a lethal event that is dependent on IL-12 and that is mediated by an excess of T cells. PMID- 10861088 TI - Ribozymes as tools for therapeutic target validation in arthritis. AB - In this paper we describe a method for validating therapeutic gene targets in arthritic disease. Ribozymes are catalytic oligonucleotides capable of highly sequence-specific cleavage of RNA. We designed ribozymes that cleave the mRNA encoding stromelysin, a matrix metalloproteinase implicated in cartilage catabolism. Ribozymes were initially screened in cultured fibroblasts to identify sites in the mRNA that were accessible for binding and cleavage. Accessible sites for ribozyme binding were found in various regions of the mRNA, including the 5' untranslated region, the coding region, and the 3' untranslated region. Several ribozymes that mediated sequence-specific and dose-dependent inhibition of stromelysin expression were characterized. Site selection in cell culture was predictive of in vivo bioactivity. An assay for measuring cartilage catabolism in rabbit articular cartilage explants was developed. Ribozymes inhibited IL-1 stimulated stromelysin mRNA expression in articular cartilage explants, yet failed to inhibit proteoglycan degradation. This indicated that up-regulation of stromelysin was not essential for IL-1-induced cartilage catabolism. Broad applications of this approach in therapeutic target validation are discussed. PMID- 10861089 TI - Critical involvement of the chemotactic axis CXCR4/stromal cell-derived factor-1 alpha in the inflammatory component of allergic airway disease. AB - Stromal cell-derived factor-1alpha/beta (SDF-1alpha/beta) is phylogenetically a primitive chemokine widely expressed in a variety of tissues and cell types. This expression is detectable in the absence of stimuli provided by bacterial or viral infections and allergic or autoimmune disorders. Based on these and other findings, SDF-1alpha has not been considered an inflammatory chemokine, but, rather, has been believed to be involved in certain homeostatic processes, such as leukocyte recirculation. SDF-1alpha is a potent chemoattractant for lymphocytes and monocytes that mediates its activity via the chemokine receptor CXCR4. Study of the role of SDF-1alpha/CXCR4 in vivo during inflammation has been limited by the fact that transgenic mice that have been made deficient in either molecule die early in life due to developmental defects. The present study was aimed at evaluating the functional relevance of the SDF-1alpha/CXCR4 axis during an inflammatory process. Neutralizing Abs to CXCR4 reduced lung eosinophilia (bronchoalveolar lavage fluid and interstitium) by half, indicating that CXCR4 mediated signals contribute to lung inflammation in a mouse model of allergic airway disease (AAD). This reduction in inflammation was accompanied by a significant decrease in airway hyper-responsiveness. SDF-1alpha neutralization resulted in similar reduction in both lung allergic inflammation and airway hyper responsiveness. Retroviral delivery of a CXCR4 cDNA to leukocytes resulted in greater inflammation when transduced mice were subjected to a mouse model of AAD. These results highlight that, although considered a noninflammatory axis, the involvement of CXCR4 and SDF-1alpha is critical during AAD, and this receptor and its ligand are potentially relevant in other inflammatory processes. PMID- 10861090 TI - Mechanism of paclitaxel activity in Kaposi's sarcoma. AB - Kaposi's sarcoma (KS) is an angioproliferative disease characterized by proliferation of spindle-shaped cells predominantly of endothelial cell origin, neoangiogenesis, inflammatory cell infiltration, and edema. At least in early stage, KS behaves as a reactive lesion sustained by the action of inflammatory cytokines and growth factors, has a polyclonal nature, and can regress. However, in time it can become monoclonal, especially in the nodular stage, evolving into a true sarcoma, likely in association with the increased expression of antiapoptotic oncogenes. We have recently demonstrated by immunohistochemical analysis that Bcl-2, a proto-oncogene known to prolong cellular viability and to antagonize apoptosis, is highly expressed in spindle cells and vessels of both AIDS-KS and classical KS lesions and that its expression increases with lesion stage. Paclitaxel, a microtubule-stabilizing drug known to inhibit Bcl-2 antiapoptotic activity and to be highly effective in the treatment of certain neoplasms, has recently been found to be active also in patients with advanced HIV-associated KS. In this report we investigated the mechanism(s) of paclitaxel activity in KS. By using a model of experimental KS induced by the inoculation of KS-derived spindle cells in nude mice and primary cultures of KS spindle cells, we found that paclitaxel promotes regression of KS lesions in vivo and that it blocks the growth, migration, and invasion of KS cells in vitro. Furthermore, paclitaxel treatment promoted apoptosis and down-regulated Bcl-2 protein expression in KS cells in vitro and in KS-like lesions in mice. Our results suggest that paclitaxel interferes with KS by down-regulating Bcl-2 antiapoptotic effect. PMID- 10861091 TI - Regulation of human cell engraftment and development of EBV-related lymphoproliferative disorders in Hu-PBL-scid mice. AB - Human PBMC engraft in mice homozygous for the severe combined immunodeficiency (Prkdcscid) mutation (Hu-PBL-scid mice). Hu-PBL-NOD-scid mice generate 5- to 10 fold higher levels of human cells than do Hu-PBL-C.B-17-scid mice, and Hu-PBL-NOD scid beta2-microglobulin-null (NOD-scid-B2mnull) mice support even higher levels of engraftment, particularly CD4+ T cells. The basis for increased engraftment of human PBMC and the functional capabilities of these cells in NOD-scid and NOD scid-B2mnull mice are unknown. We now report that human cell proliferation in NOD scid mice increased after in vivo depletion of NK cells. Human cell engraftment depended on CD4+ cells and required CD40-CD154 interaction, but engrafted CD4+ cells rapidly became nonresponsive to anti-CD3 Ab stimulation. Depletion of human CD8+ cells led to increased human CD4+ and CD20+ cell engraftment and increased levels of human Ig. We further document that Hu-PBL-NOD-scid mice are resistant to development of human EBV-related lymphoproliferative disorders. These disorders, however, develop rapidly following depletion of human CD8+ cells and are prevented by re-engraftment of CD8+ T cells. These data demonstrate that 1) murine NK cells regulate human cell engraftment in scid recipients; 2) human CD4+ cells are required for human CD8+ cell engraftment; and 3) once engrafted, human CD8+ cells regulate human CD4+ and CD20+ cell expansion, Ig levels, and outgrowth of EBV-related lymphoproliferative disorders. We propose that the Hu-PBL-NOD-scid model is suitable for the in vivo analysis of immunoregulatory interactions between human CD4+ and CD8+ cells. PMID- 10861092 TI - Functional reconstitution of class II MHC-restricted T cell immunity mediated by retroviral transfer of the alpha beta TCR complex. AB - Transfer of the alphabeta TCR genes into T lymphocytes will provide a means to enhance Ag-specific immunity by increasing the frequency of tumor- or pathogen specific T lymphocytes. We generated an efficient alphabeta TCR gene transfer system using two independent monocistronic retrovirus vectors harboring either of the class II MHC-restricted alpha or beta TCR genes specific for chicken OVA. The system enabled us to express the clonotypic TCR in 44% of the CD4+ T cells. The transduced cells showed a remarkable response to OVA323-339 peptide in the in vitro culture system, and the response to the Ag was comparable with those of the T lymphocytes derived from transgenic mice harboring OVA-specific TCR. Adoptive transfer of the TCR-transduced cells in mice induced the Ag-specific delayed-type hypersensitivity in response to OVA323-339 challenge. These results indicate that alphabeta TCR gene transfer into peripheral T lymphocytes can reconstitute Ag specific immunity. We here propose that this method provides a basis for a new approach to manipulation of immune reactions and immunotherapy. PMID- 10861093 TI - Tetramer-guided analysis of TCR beta-chain usage reveals a large repertoire of melan-A-specific CD8+ T cells in melanoma patients. AB - The assessment of the TCR repertoire expressed by tumor-specific CD8+ T lymphocytes has been hampered to date by the difficulty of targeting the analysis to lymphocytes directed against a single epitope. In the present study we have used fluorescent A2/Melan-A tetramers in conjunction with anti-CD8 and anti-TCR beta-chain variable (BV) mAbs to analyze by flow cytometry the BV segment usage by Melan-A-specific CD8+ T cells in tumor-infiltrated lymph nodes (TILN) and tumor-infiltrating lymphocytes (TIL) from A2 melanoma patients. Analysis of TILN populations revealed small proportions of A2/Melan-A tetramer+ cells expressing many different BV together with over-representation of A2/Melan-A tetramer+ cells expressing certain BVs. The BV usage by A2/Melan-A tetramer+ lymphocytes in TIL was more restricted than that in TILN. Moreover, the predominant BV segments were quite distinct in populations derived from different patients. A2/Melan-A tetramer+ cells expressing the dominant BVs found in TILN could also be found in the corresponding peptide-stimulated autologous PBMC, although A2/Melan-A tetramer+ lymphocytes expressing additional BVs were also identified. Together, these results suggest that a large and diverse repertoire of Melan-A-specific T cells using different BV TCR segments is available in A2 melanoma patients. PMID- 10861094 TI - Repeated administration of cytosine-phosphorothiolated guanine-containing oligonucleotides together with peptide/protein immunization results in enhanced CTL responses with anti-tumor activity. AB - The development of therapeutic anti-cancer vaccines designed to elicit CTL responses with anti-tumor activity has become a reality thanks to the identification of several tumor-associated Ags and their corresponding peptide T cell epitopes. However, peptide-based vaccines, in general, fail to elicit sufficiently strong CTL responses capable of producing therapeutic anti-tumor effects (i.e., prolongation of survival, tumor reduction). Here we report that repeated administration of synthetic oligonucleotides containing foreign cytosine phosphorothiolated guanine (CpG) motifs increased 10- to 100-fold the CTL response to immunization with various synthetic peptides corresponding to well known T cell epitopes. Moreover, repeated CpG administration allowed the induction of CTL to soluble protein even in the absence of additional adjuvant. Our results indicate that the potentiating effect of CpG in CTL responses required the participation of Th lymphocytes. Repeated CpG administration resulted in overt splenomegaly and lymphadenopathy with a significant increase in the numbers of CTL precursors and dendritic cells. Protein vaccination in combination with repeated CpG therapy was effective in delaying tumor cell growth and extending survival in mice bearing melanoma tumors. These findings support the contention that repeated administration of CpG-oligonucleotides enhances the effect of peptide and protein vaccines leading to potent anti-tumor responses, presumably through the induction of Th1 and dendritic cells, which are essential for optimal CTL responses. The immunostimulatory properties of CpG motifs may be key in inducing a consistent long term immunity to tumor-associated Ags when using peptides or proteins as T cell-inducing vaccines. PMID- 10861095 TI - IFN-beta-1b inhibits IL-12 production in peripheral blood mononuclear cells in an IL-10-dependent mechanism: relevance to IFN-beta-1b therapeutic effects in multiple sclerosis. AB - IL-12 is a proinflammatory cytokine secreted by dendritic cells in response to microbial Ags and mitogens. IL-12 is thought to contribute to the pathogenesis of autoimmune diseases such as multiple sclerosis (MS). This is based on studies in experimental allergic encephalomyelitis and the demonstration that PBMC IL-12 production correlates with disease progression in MS. IFN-beta-1b is an effective treatment for MS, which is thought to involve in part inhibition of proinflammatory cytokines. In this study we examined the effect of in vitro treatment with IFN-beta-1b, on mitogen-induced IL-12 production in human PBMC and myelin basic protein-specific T cell lines obtained from healthy donors and MS patients. We demonstrate that IFN-beta-1b significantly inhibits inducible IL-12 p40 up to 80% and biologically active IL-12 p70 up to 70% beginning at a dose of 10 IU/ml. This inhibition is IL-10 dependent, as it could be blocked by anti-IL 10 but not anti-IL-4 or control Abs. Thus, endogenously produced IL-10 is a required cofactor for the IFN-beta-1b inhibitory effect on IL-12 to occur. We conclude that IFN-beta-1b has a profound inhibitory effect on PBMC IL-12 production in vitro, and that this effect is IL-10 dependent. These findings are potentially relevant to the therapeutic mechanism of IFN-beta-1b in MS. PMID- 10861096 TI - Heparan sulfate-like proteoglycans mediate adhesion of human malignant melanoma A375 cells to P-selectin under flow. AB - Selectins, a family of cell adhesion molecules, bind to sialylated and fucosylated carbohydrates, such as sialyl Lewisx (SLex) and its derivatives, as their minimal recognition motif. Here we report that P-selectin bound to human malignant melanoma A375 cells and mediated their adhesion under flow. However, probing with a specific Ab failed to detect any apparent expression of SLex. This finding was bolstered by reduced expression of alpha-1,3-fucosyltransferase VII mRNA and by absence of the cell surface expression of P-selectin glycoprotein ligand-1. Instead, they expressed heparan sulfate-like proteoglycans on their cell surfaces. Treatment with beta-d -xyloside (a proteoglycan biosynthesis inhibitor) or heparinases could reduce the binding of these cells to P-selectin. In the competition assays, heparin, but not other proteoglycans, could abolish the P-selectin recognition. Further, we found that P-selectin could bind specifically to human tongue squamous cancer Tca-8113 cells, which had negative staining of SLex but positive staining of heparan sulfates. Both beta-d -xyloside and heparinases could reduce the binding of P-selectin to Tca-8113 cells. Our results thus indicate that heparan sulfate-like proteoglycans can mediate adhesion of certain types of non-blood borne, "epithelial-like" human cancer cells to P-selectin. PMID- 10861097 TI - Flt3-ligand and granulocyte colony-stimulating factor mobilize distinct human dendritic cell subsets in vivo. AB - Dendritic cells (DCs) have a unique ability to stimulate naive T cells. Recent evidence suggests that distinct DC subsets direct different classes of immune responses in vitro and in vivo. In humans, the monocyte-derived CD11c+ DCs induce T cells to produce Th1 cytokines in vitro, whereas the CD11c- plasmacytoid T cell derived DCs elicit the production of Th2 cytokines. In this paper we report that administration of either Flt3-ligand (FL) or G-CSF to healthy human volunteers dramatically increases distinct DC subsets, or DC precursors, in the blood. FL increases both the CD11c+ DC subset (48-fold) and the CD11c- IL-3R+ DC precursors (13-fold). In contrast, G-CSF only increases the CD11c- precursors (>7-fold). Freshly sorted CD11c+ but not CD11c- cells stimulate CD4+ T cells in an allogeneic MLR, whereas only the CD11c- cells can be induced to secrete high levels of IFN-alpha, in response to influenza virus. CD11c+ and CD11c- cells can mature in vitro with GM-CSF + TNF-alpha or with IL-3 + CD40 ligand, respectively. These two subsets up-regulate MHC class II costimulatory molecules as well as the DC maturation marker DC-lysosome-associated membrane protein, and they stimulate naive, allogeneic CD4+ T cells efficiently. These two DC subsets elicit distinct cytokine profiles in CD4+ T cells, with the CD11c- subset inducing higher levels of the Th2 cytokine IL-10. The differential mobilization of distinct DC subsets or DC precursors by in vivo administration of FL and G-CSF offers a novel strategy to manipulate immune responses in humans. PMID- 10861098 TI - CTL control of EBV in nasopharyngeal carcinoma (NPC): EBV-specific CTL responses in the blood and tumors of NPC patients and the antigen-processing function of the tumor cells. AB - Undifferentiated nasopharyngeal carcinoma (NPC) is latently infected with EBV and expresses a restricted number of viral proteins. Studies in healthy virus carriers have demonstrated that at least some of these proteins can act as targets for HLA class I-restricted CTLs. Therefore we have explored the possibility of a CTL-based therapy for NPC by characterizing EBV-specific CTL responses in 10 newly diagnosed NPC cases and 21 healthy virus carriers from Southeast Asia. Using the autologous EBV-transformed lymphoblastoid cell line, virus-specific CTL were reactivated in vitro from PBMC, cloned, and screened for cytotoxicity against target cells expressing individual EBV proteins from recombinant vaccinia vectors. EBV-specific CTLs were identified in 6 of 10 patients and 14 of 21 controls and mainly targeted the EBV nuclear Ag 3 (EBNA3) family of viral latent proteins. However, in 3 of 10 patients and 11 of 21 controls, CTLs specific for the NPC-associated protein LMP2 were also detected, albeit at low frequency. EBV-specific CTLs were detected in tumor biopsy material obtained from 3 of 6 of the patients, indicating that functional CTL are present at the tumor site, but none was specific for tumor-associated viral proteins. To assess the Ag-presenting function in NPC we studied two NPC-derived cell lines (C15 and c666.1) and demonstrated that both were capable of processing and presenting endogenously synthesized protein to HLA class I-restricted CTL clones. Overall, our data provide a sound theoretical basis for therapeutic strategies that aim to boost or elicit LMP2-specific CTL responses in NPC patients. PMID- 10861099 TI - Clonal expansion of infiltrating T cells in the spinal cords of SJL/J mice infected with Theiler's virus. AB - Intracerebral infection of susceptible mice with Theiler's murine encephalomyelitis virus results in immune-mediated inflammatory demyelination in the white matter and consequent clinical symptoms. This system has been utilized as an important virus model for human multiple sclerosis. Although the potential involvement of virus-specific Th cells has been studied extensively, very little is known about the nature of T cells infiltrating the CNS during viral infection and their role in the development of demyelinating disease. In this study, the clonal nature of T cells in the spinal cord during the disease course was analyzed using size spectratyping and sequencing of the TCR beta-chain CDR3 region. These studies clearly indicate that T cells are clonally expanded in the CNS after viral infection, although the overall TCR repertoire appears to be diverse. The clonal expansion appears to be Ag-driven in that it includes Th cells specific for known viral epitopes. Interestingly, such restricted accumulation of T cells was not detectable in the infiltrates of mice with proteolipid protein peptide-induced experimental autoimmune encephalomyelitis. The initial T cell repertoire (7-9 days postinfection) seems to be more diverse than that observed in the later stage (65 days) of virally induced demyelination, despite the more restricted utilization of Vbeta subfamilies. These results strongly suggest continuous stimulation and clonal expansion of virus-specific T cells in the CNS of Theiler's murine encephalomyelitis virus-infected mice during the entire course of demyelinating disease. PMID- 10861100 TI - Imiquimod: an immune response modifier. PMID- 10861101 TI - Immunomodulatory and pharmacologic properties of imiquimod. AB - Imiquimod, an imidazoquinoline amine, is an immune response modifier recently approved for the topical treatment of external genital and perianal warts. Although the majority of immunomodulatory agents available or in development inhibit pathways involved in immune activation, imiquimod is unique in that it activates immune function. The exact mechanism of imiquimod's antiviral activity is unknown; however, its effects are likely to be related to its immunomodulating properties. Although in vitro studies have shown that imiquimod has no direct antiviral effects, the drug does exhibit antiviral and antitumor effects in vivo through induction of cytokines and enhancement of cell-mediated cytolytic antiviral activity. Imiquimod stimulates the innate immune response through induction of cytokines, and the cellular arm of acquired immunity through induction of interferon-alpha (IFN-alpha), IFN-gamma, and interleukin-12. Results from animal studies have indicated a possible use for imiquimod in the prevention and treatment of herpes simplex virus infection. In addition, recent studies demonstrated that imiquimod activates Langerhans' cells and enhances allergic contact hypersensitivity. PMID- 10861102 TI - Imiquimod in clinical practice. AB - Numerous clinical trials have shown topical imiquimod (Aldara) to be effective and safe for the treatment of anogenital warts. In addition, the recurrence rate appears to be lower than those reported for many standard therapies of genital warts. Clinical experience with this medication since its availability 2 years ago has shown that it is important in the therapy of this disease. However, it is not the ideal treatment in all cases, and the advantages and disadvantages in individual patients must be considered. In addition, the antiviral and antiproliferative effects of imiquimod, as well as early reports, indicate that this medication may prove useful in the treatment of other skin diseases, including some nongenital warts and molluscum contagiosum. PMID- 10861103 TI - Human papillomavirus infections: epidemiology, pathogenesis, and host immune response. AB - Human papillomaviruses (HPVs) are ubiquitous and often cause lesions on the skin that come to the attention of the dermatologist. Skin lesions, or warts, often occur on the hands or soles of the feet and can cause embarrassment or discomfort. Genital HPV infections are transmitted by sexual contact. Infections associated with some HPV types have a high risk of progressing to carcinoma. This review discusses the molecular biology and genetics of human papillomaviruses and provides an overview of the virology, pathology, clinical manifestations, and host immune response to infection. PMID- 10861104 TI - Immunomodulatory therapy in the management of viral infections in patients with HIV infection. AB - Human immunodeficiency virus (HIV) causes disease by infecting lymphocytes and progressively destroying critical regulatory and effector cells of the immune system, leaving patients vulnerable to a number of bacterial, fungal, and viral infections. Facial herpes (herpes simplex virus-1 [HSV-1]), genital herpes (HSV 2), herpes zoster (varicella zoster virus), oral hairy leukoplakia (Epstein-Barr virus), Kaposi's sarcoma (HHV-8), molluscum contagiosum, condyloma acuminata (human papillomavirus [HPV-6, HPV-11]), plantar warts (HPV-1), and facial warts and flat warts (HPV-5) are some of the cutaneous viral diseases most commonly seen in HIV-infected patients. Two immunomodulatory agents, imiquimod (Aldara), shown to be safe and effective in the management of genital warts, and alitretinoin gel, shown to be safe and effective in the treatment of Kaposi's sarcoma, may offer a new therapeutic approach to treatment of cutaneous viral diseases. There is a strong scientific rationale to suggest that imiquimod and alitretinoin gel may be useful in the treatment of a variety of cutaneous viral diseases that have been shown to respond to immunomodulatory drugs. This represents a new approach in the therapeutic treatment paradigm for treatment of cutaneous viral diseases at their site of infection. PMID- 10861105 TI - [Effects of pravastatin on hepatic plasminogen activator inhibitor 1 mRNA expression in rabbits with fatty liver]. AB - OBJECTIVE: To explore the effects of Pravastatin on the changes of plasminogen activator inhibitor 1 (PAI-1) gene expression of hepatic tissue in rabbit with fatty liver. METHODS: We observed the influences of Pravastatin on the rabbit model of hyperlipidemia and fatty liver induced by high fat diet for 12 weeks, and the plasma PAI-1 activity and the hepatic PAI-1 mRNA expression in therapeutic group (n=10); and meanwhile, designed the model group (n=10) and normal group (n=7) as control. RESULTS: Severe hepatocellular steatosis was found in model group compared with normal group. Plasma lipids and PAI-1 activity increased in model group in 6 and 12 weeks, and was significantly higher than normal group in 12 weeks. PAI-1 mRNA relative values in hepatic tissue in model group were higher than those in normal group. Compared with model group, hepatic pathological manifestation with the respect of steatosis had no obvious changes in therapeutic group, but the lipid content and PAI-1 activity in the plasma and the expression of PAI-1 mRNA in the liver were significantly reduced. The activity of PAI was (14.0A2. 5)X10(-3)U/L in model group, and (8.6A2.0)X10(-3)U/L in therapeutic group in 12 weeks. CONCLUSION: Pravastatin could inhibit over expression of PAI-1 mRNA in the rabbit liver with hyperlipidemia and fatty liver, and reduce its plasma PAI-1 activity. PMID- 10861106 TI - [Effect of lipids on I and III procollagen mRNA expression of hepatic stellate cells]. AB - OBJECTIVE: To study the effect of lipids on I and III procollagen mRNA expression of hepatic stellate cells (HSC). METHODS: HSC were isolated and cultured from the liver of Wistar rats by in situ perfusion with pronase and collagenase and the density gradient centrifugation with Nycodenz. HSC were incubated with triglyceride (25mg/L) and very low-density lipid (VLDL, 25 mg/L) for 10d, respectively. I and III procollagen mRNA expression of HSC was detected by Northern blot hybridization. RESULTS: After HSC was stimulated with triglyceride and VLDL, the expression of I and III procollagen mRNA increased obviously. CONCLUSION: Lipids may promote the expression of I and III procollagen mRNA, and may be associated with fatty liver and hepatic fibrogenesis. PMID- 10861107 TI - [The relationship between nonalcoholic fatty liver and insulin resistance with abnormal glucose metabolism]. AB - OBJECTIVE: To explore the correlation of nonalcoholic fatty liver (called "fatty liver" for short) and insulin resistance with abnormal glucose metabolism. METHODS: The subjects were 48 patients with fatty liver, while the controls were 15 normal persons. Oral glucose tolerance test (OGTT) and insulin release test were performed; and the area under curve of serum glucose (AUCSG), area under curve of insulin (AUCIns) and insulin sensitivity index (AUCSG/AUCIns) were calculated. RESULTS: The insulin level in fatty liver group was significantly higher than that in normal control group at 60 min, 120 min and 180 min after glucose intake (P<0.01) with the postpond peak. The blood glucose level in the fatty liver group was significantly higher than that in normal control group at all time intervals except at 180 min after glucose intake (P<0.05). The AUCIns and AUCSG in the patient group were significantly higher than in the control group 209.56 vs 124.22, 24.72 vs 19.96, respectively (P<0.01). The insulin sensitivity index in the patient group was significantly lower than that in the control group (P<0. 05). There is no significant difference in the insulin sensitivity index between different degree of fatty liver (P>0.05). CONCLUSION: It is evident that patients with fatty liver have insulin resistance with abnormal glucose metabolism, which is not related with the extent of fat infiltration in the liver, suggesting that insulin resistance with abnormal glucose metabolism is a very important contributor to the pathogenesis of fatty liver. PMID- 10861109 TI - [Dynamic changes of hepatitis B virus during treatment with lamivudine versus lamivudine plus famciclovir]. AB - OBJECTIVE: To study whether combination therapy with lamivudine (LAM) and famciclovir (FCV) is effective for treatment of Chinese chronic HBV infection. METHODS: Chronic hepatitis B-infected patients treated with either LAM (n=9, 150 mg daily for 12 weeks) or LAM plus FCV (n=12, 150 mg LAM daily plus 500 mg FCV daily for 12 weeks). Serial serum HBV DNA were determined. A mathematical model was applied to analyze the dynamics of viral clearance. RESULTS: HBV clearance from patients treated with LAM was biphasic attenuation with mean antiviral efficacy of 0.94 while a combination therapy with LAM and FCV displayed an increased mean antiviral efficacy of 0. 988. There was an increased magnitude of the first phase of clearance from 1.1 log(10) to 1.9 log(10). Mean log(10) HBV viral decline were 1.8A0.2 for LAM treated group and 2.5A0.8 for combination group. CONCLUSION: Combination therapy of LAM and FCV has higher efficacy than LAM alone in suppressing HBV replication in Chinese chronic HBeAg-positive carriers. PMID- 10861108 TI - [Proliferation and apoptosis of hepatic stellate cells and effects of compound 861 on liver fibrosis]. AB - OBJECTIVE: To investigate the proliferation and apoptosis of the hepatic stellate cell (HSC) in vitro and in vivo and the effects of Chinese herb, compound 861. METHODS: The in vitro study was carried out on the culture of hepatic stellate cell line. Various concentrations of compound 861 were added and incubated. Cell proliferation was detected with MTT colorimetric assay. Cell apoptosis was detected by electron microscopy, flow cytometry and TUNEL. The subjects of in vivo study were patients with chronic hepatitis B. RESULTS: Compound 861 could significantly inhibit HSC proliferation and increase the apoptosis rate of HSC dose-dependently and time-dependently compared with the control group. After compound 861 incubation for 48h, the apoptosis rate of HSC was 25.9% compared with 9.2% in control group (P<0.05). The clinical study elucidated that the HSC was activated and proliferated in patients with chronic hepatitis B. After compound 861 treatment for 6 months, the number of activated HSC decreased and the apoptosis of HSC could be seen in the liver biopsy. CONCLUSION: Apoptosis of activated HSC exists in vitro and in vivo, stimulating its apoptosis may play an important role in the treatment of liver fibrosis. PMID- 10861110 TI - [Significance and expression of insulin-like growth factor II and its receptor in hepatocellular carcinogenesis]. AB - OBJECTIVE: To provide information on the potential role of insulin-like growth factor II (IGF-II) and insulin-like growth factor II receptor (IGF-IIR) during different liver diseases and hepatocarcinogenesis. METHODS: Southern hybridization was used to detect HBV DNA integration in various kinds of liver tissues, and DNA-RNA in situ hybridization to observe and analyze the mRNA of IGF II and IGF-II receptor in chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC). RESULTS: The expression of IGF-II and IGF-II receptor mRNA was observed not only in HCC, but also in chronic hepatitis and cirrhosis. An increasing gradient of IGF-II and IGF-IIR Mrna expression was in turn from chronic hepatitis (33. 3%), HCC (66.7%) to cirrhosis (72.0%). Strongly positive expression was found in liver cell dysplasia, regenerative nodules and poorly differentiated HCC cells. CONCLUSION: IGF-II and its receptor might play an important role in the development of HCC. PMID- 10861111 TI - [Examination and evaluation of sIL-6R, sgp130 in serum in patients with chronic hepatopathy]. AB - OBJECTIVE: To observe the variation of soluble interleukin-6 receptor (sIL-6R) and soluble interleukin-6 receptor beta strands (sgp130) in patients with chronic hepatopath. METHODS: We examined sIL-6R and sgp130 level in 40 patients with chronic hepatitis (CH), 15 with cirrhosis following hepatitis, and 35 normal controls (NC) in serum by enzyme linked immunosorbent assay (ELISA) method. RESULTS: The content of sIL-6R and sgp130 in serum was higher in CH group (224.27 mug/L and 489.35 mug/L, respectively) than NC group (174.81 mug/L and 273.64 mug/L respectively). The two parameters above in patients with hepatocirrhosis were higher than those in patients with chronic hepatitis, and the content in order of quantity was severe>moderate>slight. There was significant difference among the three groups (P<0.05, P<0.01), and positive correlation between sIL-6R and sgp130 level (r=0.481, P<0.05), between sIL-6R, sgp130 level and total bilirubin level (r=0.417, r=0. 428, P<0.01). While sIL-6R, sgp130 was no significant correlation to ALT (r=0.173, r=0.182, P>0.05). CONCLUSION: sIL-6R and sgp130 in serum are associated with the development of chronic hepatopath, and therefore can guide the assessment of prognosis. PMID- 10861112 TI - [Ultrastructure study of hepatocellular carcinoma treated by diamminedichloroplatinum]. AB - OBJECTIVE: To study the changes of the ultrastructure of hepatocellular carcinoma (HCC) after being treated by diamminedichloroplatinum. METHODS: The changes of morphological feature of HCC treated by diamminedichloroplatinum were observed with transmission electron microscope. RESULTS: The cells lost its specialized cell surface elements such as microvilli and cell-cell junctions. The nucleus chromatin condensed and segregated. The cytoplasm was hollow. The ribosomes on the endoplasmic reticulum reduced or lacked. The glycogen granules reduced. The mitochondria vacuolalized or produced flocculi, and the apoptotic bodies could be found. CONCLUSION: The changes of the ultrastructure of HCC treated by diamminedichloroplatinum correspond with the procedure of programmed cell death. Apoptosis of HCC cells can be induced by diamminedichloroplatinum. PMID- 10861113 TI - [Pathological study of the therapeutic effect of compound qinggan granula on chronic hepatitis C]. AB - OBJECTIVE: To assess histopathologically the therapeutic effect of Compound Qinggan Granula (QTG) on chronic hepatitis C. METHODS: Among 48 cases of chronic hepatitis C, 36 were treated by QTG (30g for one time and three times a day) for 6 months, and 12 were not treated with any drugs as control. Liver biopsy was done in all patients before and after the observation except 3 patients who refused to take biopsy again. Specimen were stained with HE and James method, and their grading of inflammation (G), staging of fibrosis (S) and scoring for both were then evaluated. RESULTS: In the treatment group, 14 and 26 cases showed a decreasing grading (42. 5%) and inflammation scoring (78.8%), respectively. Score for G was reduced from 8.42 to 5.58 (P<0.05). As for the fibrosis, the stage didn't change significantly (P>0.05), but the score was reduced in 19 cases (57.6%) from 3.83 to 3.13 (P<0.05). In the control group, there were not evident changes for grading and staging. The scoring of G increased from 7 to 8.6 and S from 4.15 to 6.6 (P<0.05). CONCLUSION: QTG is effective in treating chronic hepatitis C by attenuating inflammation and stopping or reversing the fibrosis. PMID- 10861114 TI - [Effect of anti-endotoxin therapy on vaso-active substances in decompensated liver cirrhosis]. AB - OBJECTIVE: To study the anti-endotoxin therapy on the plasma levels of nitric oxide, prostacyclin, tumor necrosis factor-alpha in patients with liver cirrhosis after hepatitis. METHODS: Thirty patients with decompensated liver cirrhosis accepted anti-endotoxin therapy with oral Amoxycillin and Smedta for 2 weeks. Plasma levels of endotoxin, nitric oxide, prostacyclin and tumor necrosis factor alpha were detected before and after therapy in study group and control group, respectively. The relationship between endotoxin and vasoactive substances and between the four substances and the status of patients were analyzed. RESULTS: The four substances were all increased significantly (P<0.01) in patients with liver cirrhosis compared with control group and decreased obviously after treatment for 2 weeks (endotoxin from 0.546X10(21) U/L to 0. 347X10(21) U/L, no from 56.498 mumol/L to 31.256 mumol/L, 6-keto-PGF1 alpha from 716.964 ng/L to 539.867ng/L, and TNF-alpha from 3.090 mug/L to 1.750 mug/L (P<0.01). CONCLUSION: The plasma levels of nitric oxide, prostacyclin and tumor necrosis factor-alpha increased because of endotoxemia. Amoxycillin and Smecta can clear endotoxin out effectively. PMID- 10861115 TI - [The percentage of hepatitis B virus precore A83 mutant and its dynamicchange in fulminant hepatitis B]. AB - OBJECTIVE: In order to clarify whether hepatitis B virus precore A83 mutant is related to the pathogenesis of fulminant hepatitis B or not. METHODS: The percentage of hepatitis B virus precore A83 mutant and the quantity of HBV DNA in sera from 9 patients with fulminant hepatitis B were assayed by the method of densitometry after mispairing PCR-restriction fragment length polymorphism (mpPCR RFLP) and quantitative PCR, respectively. RESULTS: The HBV precore A83 mutant was found in sera of 6 out of 9 (66.7%) fulminant hepatitis patients while none of them were infected with the mutant strain alone, and all were mixed with wild strain. The percentage of the mutant over fifty percent is only in 2 cases in whom one was 59.3% (survival) and another 64.8% (died). The percentage below 31.0% was in 4 cases and all of them died. It was observed that appearance or disappearance of the A83 mutant and its dynamic change of percentage were consistent with the fluctuation of HBV DNA level but there was no correlation statistically (r=0.602, 0.583; P>0.05). CONCLUSION: The mutant strain alone has little correlation to the occurrence of fulminant hepatitis. It might be merely a accompaniment to follow the replication of HBV under the highly immune pressure. PMID- 10861116 TI - [A randomized controlled study of ligustrazine in combination with propranolol for prevention of recurrent esophageal varices bleeding]. AB - OBJECTIVE: To assess the efficacy of Ligustrazine in combination with propranolol in the prevention of recurrent esophageal varices bleeding following liver cirrhosis, and its act mechanism. METHODS: A prospective controlled study was conducted on 74 patients, in whom 38 belonged to treatment group, and 36 to control group. By detecting the portal system hemodynamics in patients with portal hypertension and esophageal varices using color Doppler ultrasound technique, the therapeutic efficacy and safety were investigated. Meanwhile the blood pressure, heart rate, hepatic and renal function were dynamically observed. RESULTS: After four weeks administration of the drugs, the flow of portal vein and splenic vein, the diameter of portal vein and splenic vein in the treatment group were significantly decreased with the values of 1152.36A387.46 ml/min, 529.35A326.31 ml/min, 1.36A0.28 cm, and 0.94A0.19 cm, respectively. No adverse effect was observed on the system circulation and the liver function. In the follow-up period of two years, the rebleeding rate and mortality rate in the placebo group were higher than that in the treatment group, but patients with liver cirrhosis Grade C in the two groups were not significant different. CONCLUSION: Low dose ligustrazine plus propranolol is a safe and effective therapy in preventing recurrent esophageal varices bleeding, and worth further trial. PMID- 10861117 TI - [Cloning of partial cDNA of rat connective tissue growth factor and expression of its mRNA in experimental liver fibrosis]. AB - OBJECTIVE: To clone the partial cDNA sequence of rat connective tissue growth factor (CTGF) and investigate the mRNA expression of CTGF and transforming growth factor-beta 1 (TGF-beta 1) in rat experimental liver fibrosis. METHODS: A 430 base pair sequence of rat CTGF Cdna was cloned by reverse transcription polymerase chain reaction. Sixteen female Wistar rats were allocated into bile duct occlusion group (n=8) and sham-operated group (n=8). The liver tissue mRNA levels of CTGF and TGF-beta 1 were measured by multiprobe ribonuclease protection assay (RPA). RESULTS: The cloned partial cDNA sequence of rat CTGF was 95% homology (at nucleic acid level) with the mouse counterpart. In the liver of rats with biliary fibrosis mRNA levels of TGF-beta 1 and CTGF were 4 and 7 times as high as in those of sham-operated rats, respectively. CONCLUSION: The mRNA expression of CTGF is remarkably upregulated in rat liver fibrogenesis. PMID- 10861118 TI - [The expression and localization of wild-type p53-GFP fused gene on human high metastasis hepatocellular carcinoma cell line]. AB - OBJECTIVE: To study the normal expression and location of wt-p53 protein on human high-metastasis hepatocellular carcinoma (HCC) cell line. METHODS: According to the sequence of human wild-type p53, the mp53 cDNA in which stopped codon TGA was mutated to TGG, was amplified by PCR, ligated to EGFP gene, of pEGFP-N1. The recombinant plasmid was transfected into MHCC97, in which p53 gene was mutated. The expression of human p53 protein was observed by fluorescence microscopy. RESULTS: There was a p53 mutation on 249 codon in human high-metastasis hepatocellular carcinoma cell line MHCC97, but no mutation in low-metastasis HCC cells LD20. The two genes of wt-p53 and GFP were fused in codon frame by DNA sequencing. The fluorescence microscopy showed that fluorescence light was spread in all of MHCC97 cells transfected with pEGFP-N1, but fluorescence light was focused on the nucleus of MHCC97 cells transfected with pEGFP-53. CONCLUSION: The p53 mutation on 249 codon may be related to the character of metastasis of HCC cells. The human wild-type p53-GFP fusion protein is expressed in MHCC97 cell line and located in cell nucleus, which is similar to the expression of wt-p53 protein. PMID- 10861119 TI - [Changes of collagenase activity in immune hepatic fibrosis following pig's serum injection and therapeutic effect of HanDanGanLe]. AB - OBJECTIVE: To investigate the changes of collagenase activity in immune hepatic fibrosis following pig's serum injection and the therapeutic effect of HanDanGanLe. METHODS: Pig's serum was injected intraperitoneally to Wistar rats to duplicate the hepatic fibrosis model due to immunologic injury. HanDanGanLe was used as therapeutic medicine and colchicine as control. The animals were killed at 12th week to detect hepatic collagenase activity, the staining semiquantity of hepatic collagenous fibre, and the content of hepatic collagen. RESULTS: HanDanGanLe can increase collagenase activity, decrease the content of collagenous fibre and collagen in the liver. The beneficial effect achieved in HanDanGanLe-treatment group especially in high-dose group in comparison to colchicine-treatment group. CONCLUSION: HanDanGanLe can effectively increase collagenase activity, improve collagen degradation, and abate hepatic fibrosis. PMID- 10861120 TI - [Radiation-induced apoptosis and p53, bcl-2 gene expression products in QGY-7703 cell line in vitro]. AB - OBJECTIVE: To investigate the dose response and time course of radiation-induced apoptosis and gene expression products of p53, bcl-2 genes in human hepatocarcinoma cell line QGY-7703 in vitro. METHODS: Apoptosis was induced by radiation and quantitated by Flow Cytometry. The oncoproteins of p53 and bcl-2 genes after radiation were detected immunocytochemically on cell cytospinned slides. The data were analyzed with Sony Mias-300 Spato Immage Analyzer. RESULTS: The percentage of apoptosis in unradiation QGY-7703 was 4. 79%. After radiation, the apoptotic fraction increased with incubation time and radiation dose. Both gene expression products increased with incubation time and reached plateau at 6h post-radiation. CONCLUSION: After radiation, the response and time course of radiation-induced apoptosis in QGY-7703 cell line are similar to those of p53, bcl-2 oncoproteins. The later proceeds by the apoptosis peak about 6 h. The results suggest that p53 and bcl-2 genes are the regulating factors partially in radiation-induced apoptosis of QGY-7703 cell line. PMID- 10861121 TI - [Influence of HBV/C mutation on HLA expression of host cells]. AB - OBJECTIVE: To study the influence of HBV/C mutation on host cellular HLA expression. METHODS: HBV expression vectors carrying wild or variant HBV/C gene were constructed and transferred into HepG(2) cells. HBV/C gene expression on the host cells was identified. The expression of HLA-I and HLA-DR on the host cell membrane was detected. RESULTS: DNA segments similar with HBV/C gene size and HBcAg were detected by PCR and Western blot analysis, respectively. Almost all HepG(2) cells expressed HLA-I but not HLA-DR. The fluorescence intensity of HLA-I expression on host cells was different: HepG(2) was 57.8 and wild vector 54.3. The wild HBV/C gene declined significantly to 31.2. While V60, G87 and L97 increased to 43.0, 54.0 and 69.4, respectively. L97 was greatly increased with 11.6 higher than HepG(2). HLA-DR had no significant expression. CONCLUSION: Replication and expression of HBV/C gene can influence HLA-I expression on the host cells directly. Influence of variant gene on HLA-I expression is different from the wild gene. This might relate with the exacerbation of diseases resulting from HBV mutation. PMID- 10861137 TI - The ryanodine receptor: cause or consequence of diabetic heart failure? PMID- 10861138 TI - Bringing light to the dark side. PMID- 10861139 TI - The three components of hyperoxia. PMID- 10861140 TI - Anesthesia-related cardiac arrest in children: initial findings of the Pediatric Perioperative Cardiac Arrest (POCA) Registry. AB - BACKGROUND: The Pediatric Perioperative Cardiac Arrest (POCA) Registry was formed in 1994 in an attempt to determine the clinical factors and outcomes associated with cardiac arrest in anesthetized children. METHODS: Institutions that provide anesthesia for children are voluntarily enrolled in the POCA Registry. A representative from each institution provides annual institutional demographic information and submits anonymously a standardized data form for each cardiac arrest (defined as the need for chest compressions or as death) in anesthetized children 18 yr of age or younger. Causes and factors associated with cardiac arrest are analyzed. RESULTS: In the first 4 yr of the POCA Registry, 63 institutions enrolled and submitted 289 cases of cardiac arrest. Of these, 150 arrests were judged to be related to anesthesia. Cardiac arrest related to anesthesia had an incidence of 1.4 +/- 0.45 (mean +/- SD) per 10,000 instances of anesthesia and a mortality rate of 26%. Medication-related (37%) and cardiovascular (32%) causes of cardiac arrest were most common, together accounting for 69% of all arrests. Cardiovascular depression from halothane, alone or in combination with other drugs, was responsible for two thirds of all medication-related arrests. Thirty-three percent of the patients were American Society of Anesthesiologists physical status 1-2; in this group, 64% of arrests were medication-related, compared with 23% in American Society of Anesthesiologists physical status 3-5 patients (P < 0.01). Infants younger than 1 yr of age accounted for 55% of all anesthesia-related arrests. Multivariate analysis demonstrated two predictors of mortality: American Society of Anesthesiologists physical status 3-5 (odds ratio, 12.99; 95% confidence interval, 2.9-57.7), and emergency status (odds ratio, 3. 88; 95% confidence interval, 1.6-9.6). CONCLUSIONS: Anesthesia-related cardiac arrest occurred most often in patients younger than 1 yr of age and in patients with severe underlying disease. Patients in the latter group, as well as patients having emergency surgery, were most likely to have a fatal outcome. The identification of medication-related problems as the most frequent cause of anesthesia-related cardiac arrest has important implications for preventive strategies. PMID- 10861141 TI - Supplemental intraoperative oxygen augments antimicrobial and proinflammatory responses of alveolar macrophages. AB - BACKGROUND: The first goal was to test the hypothesis that 100% inspired oxygen maintained for approximately 8 h intraoperatively is not associated with impaired pulmonary oxygenation. The authors also tested the hypothesis that intraoperative inhalation of 100% oxygen augments proinflammatory and antimicrobial responses of alveolar macrophages during anesthesia and surgery. METHODS: The authors studied patients administered 100% oxygen (n = 30) and 30% oxygen (n = 30) during propofol-fentanyl general anesthesia. Alveolar macrophages were harvested by bronchoalveolar lavage immediately, 2, 4, and 6 h after induction of anesthesia, and at the end of surgery. The authors measured "opsonized" and "unopsonized" phagocytosis and microbicidal activity. RNA was extracted from harvested cells and cDNA was synthesized. The expression of interleukin(IL)-1beta, IL-6, IL-8, interferon-gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) was measured by semiquantitative polymerase chain reaction. RESULTS: Gene expression of all proinflammatory cytokines except IL-6 increased fourfold to 20-fold over time in both groups. However, expression of TNF-alpha and IL-8, IFN-gamma, and IL 6 and IL-1beta was 2-20 times greater in patients administered 100% than in those administered 30% oxygen. Unopsonized and opsonized phagocytosis and microbicidal activity decreased progressively, with the decreases being nearly twice as great during inhalation of 30% oxygen versus 100% oxygen. CONCLUSION: Inhalation of 100% oxygen improved intraoperative decreases in phagocytic and microbicidal activity possibly because expression of proinflammatory cytokines was augmented. These data therefore suggest that intraoperative inhalation of 100% oxygen augments antimicrobial and proinflammatory responses in alveolar macrophages during anesthesia and surgery. PMID- 10861142 TI - Pharyngolaryngeal, neck, and jaw discomfort after anesthesia with the face mask and laryngeal mask airway at high and low cuff volumes in males and females. AB - BACKGROUND: There is controversy over (1) the relative incidence of sore throat between the face mask (FM) and laryngeal mask airway (LMA), (2) the efficacy of LMA intracuff pressure reduction as a mechanism for minimizing sore throat, and (3) the relative incidence of sore throat with the LMA between males and females. In a randomized double-blind study, the authors compared laryngopharyngeal, neck, and jaw discomfort with the FM and LMA at high and low cuff volumes in males and females. METHODS: Three hundred adult patients were randomly assigned to three equal-sized groups for airway management: (1) the FM, (2) the LMA with a fully inflated cuff (LMA-High), or (3) the LMA with a semi-inflated cuff (LMA-Low). Anesthesia was administered with propofol, nitrous oxide, oxygen, and isoflurane. In the FM group, a Guedel-type oropharyngeal airway and jaw thrust were used only if necessary. In the LMA groups, cuff inflation was achieved with either 15 or 30 ml for the size 4 (females) and 20 or 40 ml for the size 5 (males). The LMA was removed when the patient was awake. Patients were questioned 18-24 h postoperatively about surgical pain, sore throat, sore neck, sore jaw, dysphonia, and dysphagia, and about whether they were satisfied with their anesthetic. RESULTS: The incidence of sore throat was lower in the FM (8%) than the LMA-High (42%) and LMA-Low (20%) groups (both: P < or = 0.02). The incidence of sore neck was higher for the FM (14%) than the LMA-High group (6%; P = 0.05) but similar to the LMA-Low group (8%). The incidence of sore jaw was higher in the FM (11%) than the LMA-High (3%) and LMA-Low (3%) groups (both: P = 0. 02). There were no differences among groups for surgical pain or dysphonia. The incidence of dysphagia was lower in the FM (1%) than the LMA-High group (11%; P = 0.003), but similar to the LMA-Low group (1%). The incidence of sore throat and dysphagia was lower in the LMA-Low group than the LMA-High group for both males and females (all: P < or = 0.04). There were no differences in discomfort levels between males and females in any group. Two patients from the FM group and one from the LMA-High group were not satisfied with their anesthetic. These complaints were unrelated to postoperative morbidity. CONCLUSION: The LMA causes more sore throat and dysphagia but less jaw pain than the FM. Sore throat and dysphagia are more common with the LMA if the initial cuff volume is high. There are no differences in discomfort levels between males and females. However, these discomforts do not influence patient satisfaction after LMA or FM anesthesia. PMID- 10861143 TI - Comparison of isoflurane effects on motor evoked potential and F wave. AB - BACKGROUND: Volatile anesthetics produce surgical immobility by suppressing the motor system. The anesthetic action site in the motor pathway is unclear. Anesthetic effects on the whole and the lower portion of motor pathway can be studied by measuring the motor evoked potentials (MEP) and the F wave. This study measured the effect of isoflurane on the MEP and the F wave. METHODS: With institutional review board approval, we studied 12 adult patients with American Society of Anesthesiologists physical status I or II. After intubation, anesthesia was maintained with nitrous oxide/oxygen and propofol infusion. MEPs were elicited by transcranial electrical stimuli (train-of-five pulse; stimuli intensity 40-160 mA) through electrodes placed in the scalp at C3/C4 positions and recorded at the anterior tibialis muscle with an Axon Sentinel-4EP monitor. F waves were elicited by an electrode fixed over the posterior tibial nerve at the medial malleolus and recorded at the abductor hallucis muscle. After end-tidal concentration of isoflurane was maintained at 0.5% for 20 min, the MEP and F wave were measured again. MEP and F-wave changes before and after isoflurane were analyzed using paired Wilcoxon test with Bonferroni correction. The difference between the changes in MEP and F wave was analyzed using Friedman's test. RESULTS: Motor evoked potential amplitudes (median, 205 microV; 25th-75th percentiles, 120-338 microV), F-wave amplitude (median, 100 microV; 25th-75th percentiles, 64.2-137.5 microV), and F-wave persistence (59 +/- 29%) were decreased to 0 microV (0-15 microV), 49 microV (12.4-99.6 microV), and 30 +/- 31%, respectively, by 0.5% isoflurane. MEP amplitude suppression was different from the changes in F-wave amplitude and persistence (P < 0.02). CONCLUSIONS: Isoflurane 0.5% suppresses the motor pathway by decreasing both MEP and F wave. The MEP is suppressed more than the F wave. PMID- 10861144 TI - In vivo whole-body resting energy expenditure and insulin action in human malignant hyperthermia. AB - BACKGROUND: Malignant hyperthermia (MH) is a pharmacogenetic disease triggered by volatile anesthetics or succinylcholine. The disorder is heterogenetic and caused by abnormal calcium regulation within skeletal muscle cells. No clear metabolic differences have been found in MH-susceptible (MHS) persons in vivo while not having MH episodes, but some reported signs suggest that insulin action and energy turnover might be altered in muscle of MHS persons. METHODS: In fasting and insulin-stimulated conditions, using the glucose clamp technique and indirect calorimetry, we assessed in vivo resting energy expenditure (REE) and nutrient utilization rates in 10 MHS, 5 MH-equivocal (MHE) and 10 MH-negative (MHN) persons from 14 families. With a model using the persons' fat-free mass, fat mass, age, and gender, we calculated their predicted REE and compared it with measured REE in 10 MHS and 10 MHN persons (measured - predicted = residual REE). RESULTS: In vivo measured REE and glucose disposal rates were similar in 10 MHS and 10 MHN persons. Only during insulin stimulation was residual REE greater in MHS persons (6.4%; P = 0. 013). CONCLUSIONS: In vivo insulin action is unimpaired in MHS persons. Although the absolute values of whole-body REE are the same in MHS and MHN persons, the part of REE independent of the determinants fat-free mass, fat mass, age, and gender is moderately greater in MHS than in MHN persons during insulin exposure. This suggests that MH susceptibility might influence insulin-stimulated energy turnover in muscle. PMID- 10861145 TI - Relation between perioperative hypertension and intracranial hemorrhage after craniotomy. AB - BACKGROUND: Previous data suggest that systemic hypertension (HTN) is a risk factor for postcraniotomy intracranial hemorrhage (ICH). The authors examined the relation between perioperative blood pressure elevation and postoperative ICH using a retrospective case control design. METHODS: The hospital's database of all patients undergoing craniotomy from 1976 to 1992 was screened. Coagulopathic and unmatchable patients were excluded. There were 69 evaluable patients who developed ICH postoperatively (n = 69). A 2-to-1 matched (by age, date of surgery, pathologic diagnosis, surgical procedure, and surgeon) control group without postoperative ICH was assembled (n = 138). Preoperative, intraoperative, and postoperative blood pressure records (up to 12 h) were examined. Incidence of perioperative HTN (blood pressure > or = 160/90 mmHg) and odds ratios for ICH were determined. RESULTS: Of the 11,214 craniotomy patients, 86 (0.77%) suffered ICH, and 69 fulfilled inclusion criteria. The incidence of preoperative HTN was similar in the ICH (34%) and the control (24%) groups. ICH occurred 21 h (median) postoperatively, with an interquartile range of 4-52 h. Sixty-two percent of ICH patients had intraoperative HTN, compared with only 34% of controls (P < 0.001). Sixty-two percent of the ICH patients had prehemorrhage HTN in the initial 12 postoperative hours versus 25% of controls (P < 0.001), with an odds ratio of 4.6 (P < 0.001) for postoperative ICH. Hospital stay (median, 24.5 vs. 11.0 days), and mortality (18.2 vs. 1.6%) were significantly greater in the ICH than in the control groups. CONCLUSIONS: ICH after craniotomy is associated with severely prolonged hospital stay and mortality. Acute blood pressure elevations occur frequently prior to postcraniotomy ICH. Patients who develop postcraniotomy ICH are more likely to be hypertensive in the intraoperative and early postoperative periods. PMID- 10861146 TI - Regulation of proprioceptive memory by subarachnoid regional anesthesia. AB - BACKGROUND: Patient perception of limb position during regional anesthesia is frequently incorrect. The existing model ascribes this misperception, or phantom sensation, as a reversion to a fixed, slightly flexed, body schema. A model was developed to evaluate the influence of limb position changes on the incidence of incorrect or phantom sensations during regional anesthesia. METHODS: Forty American Society of Anesthesiologists physical status I-III adult patients undergoing genitourinary procedures under subarachnoid anesthesia were assigned to a lidocaine or bupivacaine treatment group and randomly assigned to one of four time groups (1, 4, 7, and 10 min). After blockade, patients were placed supine and blinded to limb positioning manipulations. One leg was flexed and the contralateral leg extended, with leg positions subsequently reversed at the assigned time point. At 10 min, patients were asked to identify the position of each leg. Percentage of incorrect response was analyzed using a logistic regression model with two independent variables: treatment and time. A supplemental study was undertaken to evaluate the observed difference in incorrect perceptions relative to flexed first versus extended limb first sequencing. RESULTS: The inability to perceive a change in limb position under regional anesthesia is dependent on the time after the block that the position change is initiated in relation to the onset characteristics of the local anesthetic. A phantom sensation of an extended leg position clearly exists. The flexed-first limb has a significantly higher incidence of incorrect or phantom perceptions. CONCLUSION: Proprioceptive memory involves a dynamic neuroplastic imprinting process that is influenced by limb or joint position prior to onset of regional anesthesia. This contrasts with previously held beliefs of a purely fixed body schema. PMID- 10861147 TI - Usefulness of continuous oxygen insufflation into trachea for management of upper airway obstruction during anesthesia. AB - BACKGROUND: Severe complications associated with upper airway obstruction often occur during the perioperative period. Development of a simple and reliable technique for reversing the impaired airway patency may improve airway management. The purpose of the current study is to evaluate the usefulness of transtracheal oxygen insufflation (TTI) for management of upper airway obstruction during anesthesia and to explore the mechanisms of TTI in detail. METHODS: During propofol anesthesia in eight spontaneously breathing patients, the upper airway cross-sectional area and pressure-flow measurements during neck flexion with TTI were compared with those during triple airway maneuvers (TAM) without TTI. Blood gas analyses assessed efficacy of CO2 elimination during TTI in an additional nine patients. RESULTS: TTI achieved adequate PaCO2 and PaO2 levels equivalent to those during TAM. In addition to a significantly smaller cross-sectional area during TTI, the location and slope of the pressure-flow relation during TTI completely differed from those during TAM, indicating that upper airway resistance was much higher during TTI. Notably, minute ventilation during TTI was significantly smaller than that during TAM, suggesting reduced dead space or other mechanisms for CO2 elimination. CONCLUSIONS: TTI is capable of maintaining adequate blood gases through mechanisms different from those of conventional airway support in anesthetized subjects with upper airway obstruction. PMID- 10861148 TI - Correlation properties and complexity of perioperative RR-interval dynamics in coronary artery bypass surgery patients. AB - BACKGROUND: Dynamic measures of heart rate variability (HRV) may uncover abnormalities that are not easily detectable with traditional time and frequency domain measures. The purpose of this study was to characterize changes in RR interval dynamics in the immediate postoperative phase of coronary artery bypass graft (CABG) surgery using traditional and selected newer dynamic measures of HRV. METHODS: Continuous 24-h electrocardiograph recordings were performed in 40 elective CABG surgery patients up to 72 h postoperatively. In one half of the patients, Holter recordings were initiated 12-40 h before the surgery. Time and frequency domain measures of HRV were assessed. The dynamic measures included a quantitative and visual analysis of Poincare plots, measurement of short- and intermediate-term fractal-like scaling exponents (alpha1 and alpha2), the slope (beta) of the power-law regression line of RR-interval dynamics, and approximate entropy. RESULTS: The SD of RR intervals (P < 0.001) and the ultra-low-, very-low , low-, and high-frequency power (P < 0.01, P < 0.001, P < 0.001, P < 0.01, respectively) measures in the first postoperative 24 h decreased from the preoperative values. Analysis of Poincare plots revealed increased randomness in beat-to-beat heart rate behavior demonstrated by an increase in the ratio between short-term and long-term HRV (P < 0.001) after CABG. Average scaling exponent alpha1 of the 3 postoperative days decreased significantly after CABG (from 1.22 +/- 0.15 to 0.85 +/- 0.20, P < 0.001), indicating increased randomness of short term heart rate dynamics (i.e., loss of fractal-like heart rate dynamics). Reduced scaling exponent alpha1 of the first postoperative 24 h was the best HRV measure in differentiating between the patients that had normal ( 48 h, n = 7) intensive care unit stay (0.85 +/- 0.17 vs. 0.68 +/- 0.18; P < 0.05). In stepwise multivariate logistic regression analysis including typical clinical predictors, alpha1 was the most significant independent predictor (P < 0.05) of long intensive care unit stay. None of the preoperative HRV measures were able to predict prolonged intensive care unit stays. CONCLUSIONS: In the selected group of patients studied, a decrease in overall HRV was associated with altered nonlinear heart rate dynamics after CABG surgery. Current results suggest that a more random short-term heart rate behavior may be associated with a complicated clinical course. Analysis of fractal-like dynamics of heart rate may provide new perspectives in detecting abnormal cardiovascular function after CABG. PMID- 10861149 TI - Airway occlusion pressure to titrate positive end-expiratory pressure in patients with dynamic hyperinflation. AB - BACKGROUND: Although the use of external positive end-expiratory pressure (PEEP) is recommended for patients with intrinsic PEEP, no simple method exists for bedside titration. We hypothesized that the occlusion pressure, measured from airway pressure during the phase of ventilator triggering (P0.1t), could help to indicate the effects of PEEP on the work of breathing (WOB). METHODS: Twenty patients under assisted ventilation with chronic obstructive pulmonary disease were studied with 0, 5, and 10 cm H2O of PEEP while ventilated with a fixed level of pressure support. RESULTS: PEEP 5 significantly reduced intrinsic PEEP (mean +/- SD, 5.2 +/- 2.4 cm H2O at PEEP 0 to 3.6 +/- 1.9 at PEEP 5; P < 0.001), WOB per min (12. 6 +/- 6.7 J/min to 9.1 +/- 5.9 J/min; P = 0.003), WOB per liter (1.2 +/- 0.4 J/l to 0.8 +/- 0.4 J/l; P < 0.001), pressure time product of the diaphragm (216 +/- 86 cm H2O. s-1. min-1 to 155 +/- 179 cm H2O. s-1. min-1; P = 0.001) and P0.1t (3.3 +/- 1.5 cm H2O to 2.3 +/- 1.4 cm H2O; P = 0.002). At PEEP 10, no further significant reduction in muscle effort nor in P0.1t (2.5 +/- 2.1 cm H2O) occurred, and transpulmonary pressure indicated an increase in end expiratory lung volume. Significant correlations were found between WOB per min and P0.1t at the three levels of PEEP (P < 0.001), and between the changes in P0.1t versus the changes in WOB per min (P < 0.005), indicating that P0.1t and WOB changed in the same direction. A decrease in P0.1 with PEEP indicated a decrease in intrinsic PEEP with a specificity of 71% and a sensitivity of 88% and a decrease in WOB with a specificity of 86% and a sensitivity of 91%. CONCLUSION: These results show that P0.1t may help to assess the effects of PEEP in patients with intrinsic PEEP. PMID- 10861150 TI - Pulsus alternans during general anesthesia with halothane: effects of permissive hypercapnia. AB - BACKGROUND: Pulsus alternans is a classic type of abnormal pulse. It can be defined as a regular alternation of pulse amplitude in which runs of weak and strong beats follow each other alternatively without any change in cycle length. It may be a sign of severe decompensated congestive heart failure. The authors infrequently encountered some cases of pulsus alternans during halothane anesthesia with spontaneous respiration in otherwise normal subjects in association with high levels of end-tidal carbon dioxide. This study was conducted to determine if there is any relation between this phenomena and hypercapnia. METHODS: One hundred twenty patients undergoing elective lower extremity surgery were selected. Halothane was used for maintenance of anesthesia, and the patients were allowed to breath spontaneously. The occurrence of pulsus alternans was determined by plethysmographic display of pulse wave and then confirmed by palpation of the radial artery. RESULTS: Ten patients (8.3%) developed pulsus alternans together with elevated levels of end-tidal carbon dioxide (57 +/- 4 mmHg vs. 41 +/- 4 mmHg in patients without pulsus alternans [mean +/- SD]). The pulsus alternans disappeared after switching to controlled ventilation and 15-20% reduction in end-tidal carbon dioxide. During the period of pulsus alternans, vital signs and electrocardiography remained within normal limits. CONCLUSIONS: There may be some relation between occurrence of pulsus alternans and hypercapnia during halothane anesthesia. Pulsus alternans occurs in a small fraction of spontaneously breathing, halothane-anesthetized patients. Although hypercapnia is clearly a factor, the mechanism of this phenomenon is unknown. PMID- 10861151 TI - Differences in cardiovascular response to airway stimulation at different sites and blockade of the responses by lidocaine. AB - BACKGROUND: Mechanical stimulation of the airways elicits abrupt cardiovascular responses (CVR) in anesthetized humans. We examined a potential difference in such responses by comparing changes in heart rate (HR) and arterial blood pressure (AP) responses to mechanical stimulation of three different parts of the airways, as well as the effects of localized airway anesthesia with lidocaine on these responses. METHODS: After induction of general anesthesia, the larynx under laryngeal mask insertion (L, n = 20), the trachea-carina under tracheal intubation (T, n = 20), or the bronchus under bronchial intubation (B, N = 20) of each patient was mechanically stimulated in a similar manner. The same stimulation was repeated in 15 patients in each group after 5 ml of 4% lidocaine had been sprayed onto the part of the airway being stimulated. To test the systemic effect, intravenous lidocaine 1 mg/kg was given to five patients in each group, followed by the same airway stimulation. Consequent changes in HR and AP were continuously recorded and analyzed. RESULTS: Significant increases in HR and AP in response to airway tactile stimulation differed in magnitude according to the stimulated sites (L > T > or = B). These responses were completely blocked by topical application of lidocaine and partially blocked by intravenous lidocaine. CONCLUSIONS: We found that CVRs to tactile stimulation differ in their magnitude at three different sites within the airways, and localized anesthesia with lidocaine can abolish these responses in humans. The inhibition of lidocaine could be mainly due to direct blockade of the mechanoreceptors of the airways and partly to its systemic effect. PMID- 10861152 TI - The ProSeal laryngeal mask airway: A randomized, crossover study with the standard laryngeal mask airway in paralyzed, anesthetized patients. AB - BACKGROUND: The ProSeal laryngeal mask airway (PLMA) is a new laryngeal mask device with a modified cuff to improve seal and a drainage tube to provide a channel for regurgitated fluid and gastric tube placement. In the present randomized, crossover study, the authors tested the hypothesis that ease of insertion, airway sealing pressure, and fiberoptic position differ between the PLMA and the standard laryngeal mask airway (LMA). For the PLMA, we also assess ease of gastric tube placement and the efficacy of an introducer tool. METHODS: Sixty paralyzed, anesthetized adult patients were studied. Both devices (only size 4) were inserted into each patient in random order. Airway sealing pressure and fiberoptic position were determined during cuff inflation from 0 to 40 ml in 10-ml increments. Gastric tube insertion was attempted with the PLMA if there was no gas leak from the drainage tube. In 60 additional patients, ease of insertion for the PLMA was compared with and without an introducer. RESULTS: First-time success rates were higher (60 of 60 vs. 52 of 60; P = 0.003) and the effective airway time shorter (9 +/- 3 s vs20 +/- 18 s; P < 0.0001) for the LMA. There were no failed uses of either device within three attempts. Airway sealing pressure was 8-11 cm H2O higher for the PLMA at all cuff volumes (P < 0.00001) and was higher in females for both devices. Fiberoptic position was better with the LMA at all cuff volumes (P < 0.00001), but vocal cord visibility was similar (LMA, 59 of 60; PLMA, 56 of 60). For the PLMA, gastric tube placement was successful in 58 of 58 patients and took 9 +/- 5 s. First-time success rates were higher (59 of 60 vs53/60; P = 0.03) and the effective airway time shorter (15 +/- 13 s vs 23 +/- 18 s; P = 0.008) with the introducer. CONCLUSION: The PLMA is capable of achieving a more effective seal than the LMA and facilitates gastric tube placement, but it is more difficult to insert unless an introducer tool is used. When correctly positioned, the PLMA isolates the glottis from the upper esophagus with possible implications for airway protection. PMID- 10861153 TI - Difficult intubation in acromegalic patients: incidence and predictability. AB - BACKGROUND: Previous studies have suggested that the incidence of difficult intubation in acromegalic patients is higher than in normal patients. However, these studies were retrospective and did not include preoperative assessment of the airways. The aims of this study were to determine the incidence of difficult intubation and to assess the usefulness of preoperative tests in predicting difficult laryngoscopy. METHODS: One hundred twenty-eight consenting acromegalic patients requiring general anesthesia and tracheal intubation were studied. Preoperatively, Mallampati classification, thyromental distance, and head and neck movement were determined in each patient. After induction of anesthesia and muscle paralysis, laryngoscopic grade was assessed during direct laryngoscopy; Cormack and Lehane grade III or IV were classified as difficult. The association of individual airway assessment with laryngeal view was evaluated using the Fisher exact test. Predictors of difficult laryngoscopy were evaluated by calculating their sensitivity and specificity. RESULTS: Laryngoscopy was difficult (grade III) in 33 of 128 patients (26%). Application of external laryngeal pressure improved laryngeal visualization to grade II in 20 of these 33 patients. In the remaining 13 patients (10%), intubation was difficult (more than two attempts, blade change, use of gum-elastic bougie). Mallampati classes 3 and 4 were significantly related to laryngoscopy grade III (Fisher exact test, P = 0.001). CONCLUSIONS: The incidence of difficult laryngoscopy and intubation in acromegalic patients is higher than in normal patients. Preoperative Mallampati scores of 3 and 4 were of value in predicting difficult laryngoscopy. Nevertheless, even this test will miss a significant number of patients with a difficult airway. PMID- 10861154 TI - Lumbar plexus block reduces pain and blood loss associated with total hip arthroplasty. AB - BACKGROUND: The usefulness of peripheral nerve blockade in the anesthetic management of hip surgery has not been clearly established. Because sensory afferents from the hip include several branches of the lumbar plexus, the authors hypothesized that a lumbar plexus block could reduce pain from a major hip procedure. METHODS: In a double-blind prospective trial, 60 patients undergoing total hip arthroplasty were randomized to receive general anesthesia with (plexus group, n = 30) or without (control group, n = 30) a posterior lumbar plexus block. The block was performed after induction using a nerve stimulator, and 0.4 ml/kg bupivacaine, 0.5%, with epinephrine was injected. General anesthesia was standardized, and supplemental fentanyl was administered per hemodynamic guidelines. Postoperative pain and patient-controlled intravenous morphine use were serially assessed for 48 h. RESULTS: The proportion of patients receiving supplemental fentanyl intraoperatively was more than 3 times greater in the control group (20 of 30 vs. 6 of 29, P = 0.001). In the postanesthesia care unit, a greater than fourfold reduction in pain scores was observed in the plexus group (visual analogue scale [VAS] pain score at arrival 1.3 +/- 2 vs. 5.6 +/- 3, P < 0.001), and "rescue" morphine boluses (administered if VAS > 3) were administered 10 times less frequently (in 2 of 28 vs. in 22 of 29 patients, P < 0.0001). Pain scores and morphine consumption remained significantly lower in the plexus group until 6 h after randomization (VAS at 6 h, 1.4 +/- 1.3 vs. 2.4 +/- 1.4, P = 0.007; cumulative morphine at 6 h, 5.6 +/- 4.7 vs. 12.6 +/- 7.5 mg, P < 0.0001). Operative and postoperative (48 h) blood loss was modestly decreased in the treated group. Epidural-like distribution of anesthesia occurred in 3 of 28 plexus group patients, but no other side-effects were noted. CONCLUSIONS: Posterior lumbar plexus block provides effective analgesia for total hip arthroplasty, reducing intra- and postoperative opioid requirements. Moreover, blood loss during and after the procedure is diminished. Epidural anesthetic distribution should be anticipated in a minority of cases. PMID- 10861155 TI - Effect of intravenous versus epidural fentanyl on the minimum local analgesic concentration of epidural bupivacaine in labor. AB - BACKGROUND: The minimum local analgesic concentration (MLAC) has been defined as the median effective local analgesic concentration (EC50) in a 20-ml volume for epidural analgesia in the first stage of labor. The aim of this study was to determine the relative local anesthetic sparing efficacies of intravenous and epidural fentanyl by comparison of their effects on the MLAC of bupivacaine. METHODS: In this double-blind, randomized, prospective study, 84 parturients at < or = 7-cm cervical dilation who requested epidural analgesia were allocated to one of two groups. After lumbar epidural catheter placement, 20 ml bupivacaine (n = 44) or bupivacaine with 3 microg/ml (60 microg) fentanyl (n = 40) was administered. The plain bupivacaine group then received 60 microg intravenous fentanyl. The bupivacaine-fentanyl group received intravenous saline. The concentration of bupivacaine was determined by the response of the previous patient in that group to a higher or lower concentration using up-down sequential allocation. Analgesic efficacy was assessed using 100-mm visual analog pain scores, with < or = 10 mm within 30 min define as effective. RESULTS: The MLAC of bupivacaine-intravenous fentanyl was 0.064% wt/vol (95% confidence interval, 0.049-0.080), and the MLAC of bupivacaine-epidural fentanyl was 0.034% wt/vol (95% confidence interval, 0.017-0.050). Epidural fentanyl significantly increased the analgesic potency of bupivacaine by a factor of 1.88 (95% confidence interval, 1.09-3.67) compared with intravenous fentanyl. The epidural fentanyl group demonstrated significantly higher dermatomal spread (P = 0.0064) and increased pruritus (P = 0. 01). CONCLUSIONS: Epidural fentanyl significantly reduced the MLAC of bupivacaine when compared with intravenous fentanyl for the parturients in this study. The significantly enhanced local anesthetic sparing, dermatomal level, and pruritus with epidural fentanyl suggest a primarily spinal site of action. PMID- 10861156 TI - Analysis of risk factors for myocardial infarction and cardiac mortality after major vascular surgery. AB - BACKGROUND: Patients undergoing vascular surgical procedures are at high risk for perioperative myocardial infarction (PMI). This study was undertaken to identify predictors of PMI and in-hospital death in major vascular surgical patients. METHODS: From the Vascular Surgery Registry (6,948 operations from January 1989 through June 1997) the authors identified 107 patients in whom PMI developed during the same hospital stay. Case-control patients (patients without PMI) were matched at a 1x:x1 ratio with index cases according to the type of surgery, gender, patient age, and year of surgery. The authors analyzed data regarding preoperative cardiac disease and surgical and anesthetic factors to study association with PMI and cardiac death. RESULTS: By using univariable analysis the authors identified the following predictors of PMI: valvular disease (P = 0.007), previous congestive heart failure (P = 0.04), emergency surgery (P = 0.02), general anesthesia (P = 0.03), preoperative history of coronary artery disease (P = 0.001), preoperative treatment with beta-blockers (P = 0.003), lower preoperative (P = 0.03) and postoperative (P = 0.002) hemoglobin concentrations, increased bleeding rate (as assessed from increased cell salvage; P = 0.025), and lower ejection fraction (P = 0.02). Of the 107 patients with PMI, 20.6% died of cardiac cause during the same hospital stay. The following factors increased the odds ratios for cardiac death: age (P = 0.001), recent congestive heart failure (P = 0.01), type of surgery (P = 0.04), emergency surgery (P = 0.02), lower intraoperative diastolic blood pressure (P = 0.001), new intraoperative ST-T changes (P = 0.01), and increased intraoperative use of blood (P = 0.005). Patients who underwent coronary artery bypass grafting, even more than 12 months before index surgery, had a 79% reduction in risk of death if they had PMI (P = 0.01). Multivariable analysis revealed preoperative definitive diagnosis of coronary artery disease (P = 0.001) and significant valvular disease (P = 0.03) were associated with increased risk of PMI. Congestive heart failure less than 1 yr before index vascular surgery (P = 0. 0002) and increased intraoperative use of blood (P = 0.007) were associated with cardiac death. The history of coronary artery bypass grafting reduced the risk of cardiac death (P = 0.04) in patients with PMI. CONCLUSIONS: The in-hospital cardiac mortality rate is high for patients who undergo vascular surgery and experience clinically significant PMI. Stress of surgery (increased intraoperative bleeding and aortic, peripheral vascular, and emergency surgery), poor preoperative cardiac functional status (congestive heart failure, lower ejection fraction, diagnosis of coronary artery disease), and preoperative history of coronary artery bypass grafting are the factors that determine perioperative cardiac morbidity and mortality rates. PMID- 10861157 TI - Attenuation of the preoperative stress response with midazolam: effects on postoperative outcomes. AB - BACKGROUND: Previously, effects of preoperative sedatives were assessed mainly with respect to preoperative outcomes such as anxiety and compliance. The purpose of this investigation was to evaluate the effects of preoperative sedatives on postoperative psychological and clinical recovery. METHODS: Patients undergoing general anesthesia and outpatient surgery were enrolled in a double-blind, randomized, placebo-controlled trial. Subjects (n = 55) were randomly assigned to receive either 5 mg intramuscular midazolam (n = 26) or a placebo injection (n = 29) at least 30 min before surgery. The anesthetic technique was controlled. Postoperative anxiety, pain, analgesic consumption, clinical recovery parameters, and global health (SF-36) were evaluated up to 1 month after surgery. RESULTS: Surgery length did not differ significantly between the treatment and placebo groups (118 +/- 45 min vs 129 +/- 53 min; P = NS). Throughout the first postoperative week, subjects in the treatment group reported a greater reduction in postoperative pain compared with subjects in the placebo group (F1,50= 3.5; P = 0.035). Moreover, at 1 week, ibuprofen use was reported by less subjects in the treatment group than in the placebo group (0% vs 17.2%; P = 0.026). Subjects in the treatment group also reported a greater reduction in postoperative anxiety throughout the follow-up period (F1,53 = 9.2; P = 0.04). However, global health indexes (SF-36) did not detect any significant differences between the two experimental groups (multivariate F1,45 = 0.44; P = 0.51). CONCLUSION: Subjects treated with midazolam preoperatively self-report improved postoperative psychological and pain recovery. However, the clinical significance of these findings is unclear at the present time. PMID- 10861158 TI - Alternative medicine use in presurgical patients. AB - BACKGROUND: A dramatic increase in the use of complementary and alternative medicines has been observed. The use of such remedies in the presurgical population has implications for the anesthesiologist because of the potential for drug interactions, side effects, and medical liability. This study was undertaken to quantify the use of herbal remedies and vitamins in the presurgical population of a large tertiary care center. METHODS: A one-page questionnaire was distributed to all patients presenting for evaluation in the preoperative clinic over an 11-week period. Patients answered questions regarding use of prescription and nonprescription medications, herbal remedies, and vitamins. RESULTS: Twenty two percent of presurgical patients reported the use of herbal remedies, and 51% used vitamins. Women and patients aged 40-60 yr were more likely to use herbal medicines. Over-the-counter medication use was strongly associated with herbal preparation use. The most commonly used compounds, from highest to lowest, included echinacea, gingko biloba, St. John's wort, garlic, and ginseng. CONCLUSIONS: Alternative medicine use is common in the preoperative period. PMID- 10861159 TI - Anesthesiologist direction and patient outcomes. AB - BACKGROUND: Anesthesia services for surgical procedures may or may not be personally performed or medically directed by anesthesiologists. This study compares the outcomes of surgical patients whose anesthesia care was personally performed or medically directed by an anesthesiologist with the outcomes of patients whose anesthesia care was not personally performed or medically directed by an anesthesiologist. METHODS: Cases were defined as being either "directed" or "undirected," depending on the type of involvement of the anesthesiologist, as determined by Health Care Financing Administration billing records. Outcome rates were adjusted to account for severity of disease and other provider characteristics using logistic regression models that included 64 patient and 42 procedure covariates, plus an additional 11 hospital characteristics often associated with quality of care. Medicare claims records were analyzed for all elderly patients in Pennsylvania who underwent general surgical or orthopedic procedures between 1991-1994. The study involved 194,430 directed and 23,010 undirected patients among 245 hospitals. Outcomes studied included death rate within 30 days of admission, in-hospital complication rate, and the failure-to rescue rate (defined as the rate of death after complications). RESULTS: Adjusted odds ratios for death and failure-to-rescue were greater when care was not directed by anesthesiologists (odds ratio for death = 1.08, P < 0.04; odds ratio for failure-to-rescue = 1.10, P < 0.01), whereas complications were not increased (odds ratio for complication = 1.00, P < 0.79). This corresponds to 2.5 excess deaths/1,000 patients and 6.9 excess failures-to-rescue (deaths) per 1,000 patients with complications. CONCLUSIONS: Both 30-day mortality rate and mortality rate after complications (failure-to-rescue) were lower when anesthesiologists directed anesthesia care. These results suggest that surgical outcomes in Medicare patients are associated with anesthesiologist direction, and may provide insight regarding potential approaches for improving surgical outcomes. (Key words: Anesthesiologists; anesthesia care team; quality of care; mortality; failure-to-rescue; complication; Medicare; general surgery; orthopedics.) PMID- 10861160 TI - Norepinephrine release from spinal synaptosomes: auto-alpha2 -adrenergic receptor modulation. AB - BACKGROUND: Clonidine produces analgesia after spinal injection by activating alpha2-adrenergic receptors. Recently, clonidine has been demonstrated to increase spinal release of norepinephrine (NE) in vivo, in contrast to that anticipated by classic presynaptic autoinhibition. The purpose of the current study was to determine if clonidine could inhibit release of NE in a preparation of spinal cord tissue lacking synaptic circuits. METHODS: Crude synaptosomes were prepared from male Sprague-Dawley rat spinal cord, loaded with [3H]NE, and stimulated by potassium chloride to release [3H]NE. Samples were incubated with clonidine in the absence or presence of various inhibitors. To study the effect of alpha2a-adrenergic receptor subtypes, some animals were pretreated with an oligodeoxynucleotide (ODN) composed of a sense or antisense sequence to a portion of this receptor. RESULTS: Potassium chloride produced a concentration-dependent increase in [3H]NE release, and this release was inhibited by clonidine with a concentration producing 50% maximal inhibition (IC50) of 1.3 microm. The effect of clonidine was inhibited by the alpha2-adrenergic antagonists, yohimbine and idazoxan, but not by alpha1-adrenergic, muscarinic, or opioid antagonists. Intrathecal pretreatment with antisense ODN to alpha2A-adrenergic receptors reduced alpha2A-adrenergic receptor protein expression compared with sense ODN control and also reduced clonidine-induced inhibition of [3H]NE release. CONCLUSIONS: These data demonstrate the existence of classic autoinhibitory alpha2-adrenergic receptors in the spinal cord, probably of the alpha2Asubtype. They further suggest that clonidine-induced stimulation of spinal NE release must occur from indirect actions, presumably due to activation of a spinal circuit. PMID- 10861161 TI - Activation of peripheral NMDA-nitric oxide cascade in formalin test. AB - BACKGROUND: It has been suggested that peripheral glutamate and nitric oxide (NO) released by tissue-damaging stimuli play an important role in peripheral nociceptive transmission. This study was conducted to determine whether NO was released in the periphery after subcutaneous injection of formalin and whether the peripheral N-methyl-d-aspartate (NMDA)-NO cascade was activated. METHODS: During pentobarbital anesthesia, a microdialysis probe was implanted subcutaneously into the glabrous skin of both hind paws of Sprague-Dawley rats. After sample collection to obtain the basal level of NO metabolites (total amount of nitrite [NO2-] and nitrate [NO3-] [NO2--NO3-]), 5% formalin was injected into the plantar surface of the right hind paw during perfusion of the dialysis catheters with artificial cerebrospinal fluid (ACSF), the NO synthase inhibitor NG-monomethyl-L-arginine acetate, or the NMDA antagonist D,l-2-amino-5 phosphonovaleric acid through a microdialysis probe. Formalin also was injected in the animals that underwent sciatic nerve sectioning. In another series of experiments, NMDA was perfused through one probe. Samples for measurement of NO2- NO3- were collected and immediately analyzed using high-performance liquid chromatography. RESULTS: Subcutaneous formalin significantly increased the dialysate concentrations of NO2--NO3- (maximum increase 144 +/- 12% of baseline value 30 min after formalin administration; P < 0.05) on the side ipsilateral to the injection. Both NG-monomethyl-l-arginine acetate and D, l-2-amino-5 phosphonovaleric acid significantly (P < 0.05) suppressed the formalin-induced increases in NO2--NO3- concentration. In the rats with denervation of the sensory nerves, formalin did not change the NO2--NO3- concentration. In addition, NMDA perfusion significantly (P < 0.05) increased the concentrations of NO2--NO3-. CONCLUSION: The results of the current study show that subcutaneous formalin injection induces peripheral release of NO, the production of which is mediated by activation of NMDA receptors in the peripheral nervous system. PMID- 10861162 TI - Low-temperature modification of the inhibitory effects of volatile anesthetics on airway smooth muscle contraction in dogs. AB - BACKGROUND: Because exposure to low temperature can modify the effect of volatile anesthetics on airway smooth muscle contraction, this study was conducted to investigate low-temperature modifications of the inhibitory effects of isoflurane and sevoflurane on canine tracheal smooth muscle tone by simultaneously measuring the muscle tension and intracellular concentration of Ca2+ ([Ca2+]i) and by measuring voltage-dependent Ca2+ channel activity. METHODS: [Ca2+]i was monitored by the 500-nm light emission ratio of preloaded fura-2, a Ca2+ indicator. Isometric tension was measured simultaneously. Whole cell patch clamp recording techniques were used to observe voltage-dependent Ca2+ channel activity in dispersed muscle cells. Isoflurane (0-3.0%) or sevoflurane (0-3%) was introduced to a bath solution at various temperatures (37, 34, or 31 degrees C). RESULTS: Low temperature (34 or 31 degrees C) reduced high-K+-induced (72.7 mm) muscle contraction and increased [Ca2+]i, but it enhanced carbachol-induced (1 microm) muscle contraction with a decrease in [Ca2+]i. The volatile anesthetics tested showed significant inhibition of both high-K+-induced and carbachol-induced airway smooth muscle contraction, with a concomitant decrease in [Ca2+]i. The inhibition of the carbachol-induced muscle contraction by volatile anesthetics was abolished partially by exposure to low temperature. Volatile anesthetics and low-temperature exposure significantly inhibited voltage-dependent Ca2+ channel activity of the smooth muscle. CONCLUSIONS: Exposure of airway smooth muscle to low temperature leads to an increase in agonist-induced muscle contractility, with a decrease in [Ca2+]i. The inhibition of voltage-dependent Ca2+ channel activity by exposure to low temperature and by volatile anesthetics cam be attributed, at least in part, to the decrease in [Ca2+]i. PMID- 10861163 TI - Effects of halothane and isoflurane on the intracellular Ca2+ transient in ferret cardiac muscle. AB - BACKGROUND: Halothane and isoflurane depress myocardial contractility by decreasing transsarcolemmal Ca2+ influx and Ca2+ release from the sarcoplasmic reticulum. Decreases in Ca2+ sensitivity of the contractile proteins have been shown in skinned cardiac fibers, but the relative importance of this effect in intact living myocardium is unknown. The aims of this study were to assess whether halothane and isoflurane decrease myofibrillar Ca2+ sensitivity in intact, living cardiac fibers and to quantify the relative importance of changes in myofibrillar Ca2+ sensitivity versus changes in myoplasmic Ca2+ availability caused by these anesthetics. METHODS: The effects of halothane and isoflurane (0 1.5 times the minimum alveolar concentration (MAC) in three equal increments) on isometric and isotonic variables of contractility and on the intracellular calcium transient were assessed in isolated ferret right ventricular papillary muscle microinjected with the Ca2+-regulated photoprotein aequorin. The intracellular calcium transient was analyzed in the context of a multicompartment model of intracellular Ca2+ buffers in mammalian ventricular myocardium. RESULTS: Halothane and isoflurane decreased contractility, time-to-peak force, time to half-isometric relaxation, and intracellular Ca2+ transient in a reversible, concentration-dependent manner. Halothane, but not isoflurane, slowed the increase and the decrease of the intracellular Ca2+ transient. Increasing extracellular Ca2+ in the presence of anesthetic to produce peak force equal to control values increased intracellular Ca2+ to values higher than control values. CONCLUSIONS: Halothane decreases myoplasmic Ca2+ availability more than isoflurane; halothane and isoflurane decrease myofibrillar Ca2+ sensitivity to the same extent; in halothane at 0.5 MAC and isoflurane at 1.0 MAC, the decrease in Ca2+ sensitivity is already fully apparent; halothane decreases intracellular Ca2+ availability more than myofibrillar Ca2+ sensitivity; and isoflurane decreases myoplasmic Ca2+ availability and Ca2+ sensitivity to the same extent, except at 1.5 times the MAC, which decreases Ca2+ availability more. PMID- 10861164 TI - Neuroprotective, anesthetic, and cardiovascular effects of the NMDA antagonist, CNS 5161A, in isoflurane-anesthetized lambs. AB - BACKGROUND: N-methyl-d-aspartate (NMDA) receptor antagonists are neuroprotective in animal models of cerebral ischemia, but adverse cardiovascular and neurobehavioral effects have precluded their clinical use. The authors present the neuroprotective, anesthetic, and cardiovascular effects of a novel NMDA antagonist, CNS 5161A. METHODS: Lambs, 4.0-6.5 kg, were anesthetized with isoflurane, intubated, and ventilated and had thermodilution catheters placed in the pulmonary artery and 20-g catheters placed in the femoral artery. The minimum alveolar concentration (MAC) of isoflurane was determined using the "bracketing technique." CNS 5161A was given as a bolus and then as an infusion at three doses. Cardiovascular measurements were determined every 15 min. Other lambs (n = 25) were subjected to cardiopulmonary bypass (CPB) with hypothermic circulatory arrest (HCA) for 120 min. Eighteen received CNS 5161A, and seven received saline vehicle. One hour after CPB, brains were perfusion-fixed and removed for in situ hybridization and immunohistochemistry analysis in half of the animals. The other half survived 48 h before their brains were examined for neuronal degeneration. RESULTS: Isoflurane at MAC significantly decreased blood pressure, heart rate, cardiac output, and systemic vascular resistance by 30-48% (n = 16; P < 0.05). CNS 5161A (n = 12) had no significant cardiovascular effects. All concentrations of CNS 5161A caused a significant reduction (21-29%) of the MAC of isoflurane (n = 12; P < 0.05). CNS 5161A, at serum concentrations greater than 25 ng/ml, completely inhibited c-fosmRNA and c-FOS protein expression in hippocampal neurons after 120 min of HCA, attenuated neuronal degeneration, and improved functional outcome by 47% (P < 0.05). CONCLUSIONS: CNS 5161A at neuroprotective concentrations before CPB-HCA significantly reduces the MAC of isoflurane without cardiovascular effects. PMID- 10861165 TI - Norepinephrine-induced apoptosis is inhibited in adult rat ventricular myocytes exposed to volatile anesthetics. AB - BACKGROUND: Volatile anesthetics are used to provide anesthesia to patients with heart disease under heightened adrenergic drive. The purpose of this study was to test whether volatile anesthetics can inhibit norepinephrine (NE)-induced apoptosis in cardiomyocytes. METHODS: Rat ventricular cardiomyocytes were exposed to NE (10 microm) alone or in the presence of increasing concentrations of isoflurane and halothane. RESULTS: Isoflurane at 1.6 minimum alveolar concentration (MAC) (4 +/- 2% [SD]) and halothane at 1.2 MAC (3 +/- 2%) abolished the percentage of cardiomyocytes undergoing NE-induced apoptosis (34 +/- 8%), as assessed by terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) (P < 0.0001). Lower concentrations of isoflurane and halothane markedly decreased the number of TUNEL-positive cells. Similarly, isoflurane at 1.6 MAC (5 +/- 3%) and halothane at 1.2 MAC (6 +/- 3%) prevented the increase in annexinV staining cardiomyocytes (38 +/- 7%; P < 0. 0001). These findings were corroborated with a decreased quantity of NE-induced DNA laddering by volatile anesthetics. Halothane at 1.2 MAC abolished the increase in TUNEL-positive cardiomyocytes exposed to the dihydropyridine Ca2+-channel agonist BAY K-8644 (1 microm) (BAY K-8644 + halothane: 3 +/- 2% vsBAY K-8644: 34 +/- 6%; P < 0. 0001) and the Ca2+-ionophore 4-bromo-A23187 (1 microm) (4-bromo-A23187 + halothane: 2 +/- 2% vs4-bromo-A23187: 13 +/- 4%; P = 0.03). NE treatment increased caspase-9 activity to 197 +/- 62% over control myocytes (P < 0.0001), whereas no caspase-8 activation was detectable. This increase in caspase-9 activity was blocked by isoflurane at 1.6 MAC and halothane at 1.2 MAC. CONCLUSIONS: Volatile anesthetics offer significant protection against beta-adrenergic apoptotic death signaling in ventricular cardiomyocytes. The authors present evidence that this protection is mainly mediated through modulation of cellular Ca2+ homeostasis and inhibition of the apoptosis initiator caspase-9. PMID- 10861166 TI - Intestinal inflammation and morphine tolerance alter the interaction between morphine and clonidine on gastrointestinal transit in mice. AB - BACKGROUND: Morphine and clonidine show synergy or antagonism inhibiting gastrointestinal transit depending on their proportion and level of effect. Their interaction during morphine tolerance and intestinal inflammation were assessed. METHODS: Gastrointestinal transit in mice was evaluated with charcoal and antitransit effects expressed as percent mean values +/- SEM. Tolerance was induced with a morphine pellet (75 mg) implanted for 72 h, and inflammation with intragastric croton oil. Dose-response curves for morphine and clonidine alone and combined at a 1:1 potency ratio were obtained, and doses producing a 50% and 60% inhibition were calculated (ED50, ED60). Interaction was established by isobolograms, interaction indexes, and analysis of variance. RESULTS: In naive and tolerant mice, the combination induced linear dose-response curves up to the ED60 and then reached a plateau. In naive mice, ED50 values were as follows: morphine 1.52 +/- 0.15 mg/kg, clonidine 0.09 +/- 0.008 mg/kg, and combined 0.506 +/- 0.084 mg/kg (0.478 +/- 0.08 mg/kg morphine plus 0.028 +/- 0.004 mg/kg clonidine). During tolerance, ED50 values were as follows: morphine 9.73 +/- 0.8 mg/kg, clonidine 0.09 +/- 0.007 mg/kg, combination 0.131 +/- 0.09 mg/kg (morphine 0. 13 +/- 0.09 mg/kg plus clonidine 0.0013 +/- 0.0005 mg/kg). In both groups, the interaction was synergistic up to the ED60 and antagonistic thereafter; synergy was enhanced during tolerance. During inflammation, ED50 values were as follows: morphine 0.17 +/- 0.04 mg/kg, clonidine 0.015 +/- 0.006 mg/kg, combined 0.62 +/- 0.04 mg/kg (morphine 0.568 +/- 0.04 mg/kg plus clonidine 0.052 +/- 0.004 mg/kg); thus, potencies of morphine and clonidine increased 9.3 and 7.1 times, while the combination remained unaltered. Moreover, inflammation transformed synergy into antagonism. CONCLUSIONS: The interaction between morphine and clonidine was significantly altered during tolerance and inflammation. During tolerance, synergy was present up to 60% effect and then became antagonistic. Inflammation converted synergy to antagonism. A common pathway in signal transduction could partially explain the results. PMID- 10861167 TI - The effects of ketamine and its enantiomers on the morphine- or dexmedetomidine induced antinociception after intrathecal administration in rats. AB - BACKGROUND: The spinal administration of some N-methyl-d-aspartate receptor antagonists results in antinociception and potentiates the effects of opioids and alpha2-adrenoceptor agonists, but ketamine and its enantiomers have not been examined. The present study investigated the interactions of racemic ketamine, R( )-ketamine and S(+)-ketamine with morphine and with dexmedetomidine. METHODS: Intrathecal catheters were implanted into male Wistar rats. Three days later, the acute nociceptive sensitivity was assessed using the tail-flick test. Analgesic latencies were converted to the percentage maximum possible effect. The dose that yielded 50% of the maximum possible effect (ED50) and dose-response and time course curves were determined for the ketamines (30-300 microg), morphine (0.1 3.0 microg), dexmedetomidine (0.3-10.0 microg), and mixtures of two doses of ketamines (30 or 100 microg) with different doses of morphine or dexmedetomidine for fixed-dose analysis. RESULTS: Neither racemic ketamine nor its enantiomers alone had a significant effect on the tail-flick test, with the exception of the highest dose of racemic ketamine, which caused motor impairment. Morphine and dexmedetomidine each produced dose-dependent antinociception, with ED50 of 1.7 microg (95% confidence interval: 1.04-2.32) and 4. 85 microg (3.96-5.79), respectively. A low dose (30 microg) of racemic ketamine or its enantiomers did not influence the ED50 of morphine significantly. Coadministration of 100 microg racemic ketamine or S(+)-ketamine, but not R(-)-ketamine, significantly enhanced and prolonged the antinociceptive effect of morphine. Both doses of racemic ketamine or its isomers significantly decreased the ED50 value for dexmedetomidine, although the higher dose of racemic or S(+)-ketamine had the highest potency. One-hundred micrograms of racemic ketamine or S(+)-ketamine also prolonged the effects of dexmedetomidine. CONCLUSIONS: These data indicate that racemic ketamine and S(+)-ketamine, but not R(-)-ketamine, exhibit similar effectiveness in potentiating the antinociceptive effects of both morphine and dexmedetomidine. PMID- 10861168 TI - Eliminating blood transfusions: new aspects and perspectives. PMID- 10861169 TI - Pioneer Chinese anesthesiologists: American influences. PMID- 10861170 TI - The ancestors of inhalational anesthesia: the Soporific Sponges (XIth-XVIIth centuries): how a universally recommended medical technique was abruptly discarded. PMID- 10861171 TI - Carbon dioxide for obstetric pneumoperitoneum. PMID- 10861172 TI - Preoperative cardiac assessment has to take into account the type of surgery. PMID- 10861173 TI - Assessment of the patient with cardiac disease. PMID- 10861174 TI - Compliance and capnography monitoring during independent lung ventilation: report of two cases. PMID- 10861175 TI - The rapid infusion system: user error in tubing connection mimicking severe hemorrhage. PMID- 10861176 TI - Anaphylaxis to penicillin on reperfusion during liver transplantation. PMID- 10861177 TI - Negative-pressure pulmonary edema in a child with hiccups during induction. PMID- 10861178 TI - A comparison of two ventilator systems using an infant lung model. PMID- 10861179 TI - Motor blockade by brachial plexus block in the sheep. PMID- 10861180 TI - Revisiting the ASA guidelines for management of a difficult airway. PMID- 10861181 TI - Another use for nasopharyngeal airway. PMID- 10861182 TI - Comparing combined spinal-epidural analgesia with conventional epidural analgesia: were the two groups identical? PMID- 10861183 TI - Another explanation. PMID- 10861184 TI - Measurement of sympathetic blockade: effect of epidural and spinal anesthesia. PMID- 10861185 TI - OR scheduling algorithms. PMID- 10861186 TI - Poor Hippocrates. PMID- 10861187 TI - Intraoperative pacemaker malfunction during a shoulder arthroscopy. PMID- 10861188 TI - Anesthetic considerations of a patient with a tongue piercing, and a safe solution. PMID- 10861189 TI - Double-access-port endotracheal tube for selective lung ventilation in pediatric patients. PMID- 10861190 TI - A versatile alternative to standard laryngoscopy. PMID- 10861191 TI - Loss of propofol during in vitro experiments. PMID- 10861192 TI - Development of an ELISA system for detection of anti-Anaplasma marginale antibodies in cattle in Brazil. AB - An ELISA test was developed for detecting antibodies against Anaplasma marginale in bovine sera. Four antigenic preparations were produced from infected red blood cells. Some aliquots of this preparation were stored at -70 degrees C with 30% DMSO in phosphate-buffered saline (PBS) and others were lysed with 0.9% NH4Cl and stored at -20 degrees C. Typical anaplasmal structures were seen by electron microscopy in the antigenic preparations containing the erythrocytes that had been stored with DMSO. The performance of the ELISA test was evaluated by testing 298 positive serum samples collected from immunized cattle, 39 negative serum samples collected from cattle imported from areas free of A. marginale and 50 samples collected from cattle naturally infected in the field. The test gave a specificity of 94.87% and a sensitivity of 100%. PMID- 10861193 TI - Scanning electron microscopy of adult Gongylonema pulchrum (Nematoda: Spirurida). AB - Scanning electron microscopy (SEM) was used to study the surface ultrastructure of adult worms of Gongylonema pulchrum. The anterior end in both sexes was covered by numerous cuticular platelets. There was a pair of lateral cervical papillac. The buccal opening was small and extended in the dorsoventral direction. Around the mouth a cuticular elevation enclosed the labia, and eight papillae were located laterodorsally and lateroventrally. Two large lateral amphids were seen. On the lateral sides of the female's tail, phasmidal apertures were observed. The caudal end of the male was asymmetrically alate and bore 10 pairs of papillae and two phasmidal apertures. PMID- 10861194 TI - Study on risk factors and their association with subclinical mastitis in lactating dairy cows in Trinidad. AB - A cross-sectional study was conducted on dairy farms in eight milking centres in Trinidad to determine the prevalence of risk factors for mastitis and to assess their relationship to occurrence of subclinical mastitis. The California mastitis test (CMT) was used to determine the prevalence of subclinical mastitis by estimating the somatic cell counts in bulk and composite milk. Of a total of 177 dairy farms studied, 121 (68.4%), 39 (22.0%) and 17 (9.6%) practised semi intensive, extensive and intensive management systems, respectively. A total of 129 (72.9%), 37 (20.9%) and 11 (6.2%) farms milked cows in parlours, stanchions and pasture/out-on-field, respectively. Based on sanitary practices, 40 (22.6%), 123 (69.5%), and 14 (7.9%) farms were classified as good, fair, and poor, respectively, while 76 (42.9%) and 60 (33.9%) farms reported to rarely experience and frequently experience water shortages, respectively. Amongst the 177 farms, only seven (4.0%) used machine-milking primarily, 152 (85.9%) screened for mastitis as a routine, 18 (10.2%) teat dipped, and 49 (27.7%) practised dry cow therapy. To detect mastitis, of 152 farms involved, 20 (13.2%) used the strip cup while only two (1.3%) employed the CMT. Pipe-borne water delivered directly from the hose was the only source of water to 91 (51.4%) farms while seven (4.0%) and eight (4.5%) farms used only well and surface water (ponds and rivers), respectively. Based on bulk milk samples, the farm prevalence of subclinical mastitis was 60.5% (107 of 177) with a range from 33.3% (centre 5H) to 100.0% (centre 2B). The difference was statistically significant (P < 0.01; chi 2). However, using composite milk, the farm prevalence of subclinical mastitis was 52.5% (93 of 177) with a range from 21.2% (centre 5H) to 92.9% (centre 2B) and again, the difference in prevalence was statistically significant (P < 0.001; chi 2). Subclinical mastitis was detected in 150 (45.0%) of 333 lactating cows screened and the range of prevalence was from 17.9% (centre 5H) to 56.3% (centre 1C). The difference was statistically significant (P < 0.001; chi 2). Of a total of 14 risk factors for mastitis studied which were related to animal husbandry, personnel, mastitis control and water, only two, the herd size and practice of dry cow therapy were significantly (P < 0.05; chi 2) associated with subclinical mastitis. It was concluded that the high prevalence of subclinical mastitis in Trinidad dairy herds could significantly reduce milk production with associated economic loss. Although a majority of the risk factors studied were not found to be significantly associated with the occurrence of subclinical mastitis possibly due to confounding factors, the need to eliminate or reduce these risk factors cannot be over-emphasized. PMID- 10861195 TI - Immunization of pups with maternally derived antibodies to canine parvovirus (CPV) using a modified-live variant (CPV-2b). AB - The results of vaccination trials carried out on pups with maternally derived antibodies (MDA) to canine parvovirus (CPV), using a modified-live CPV-2b variant vaccine (29-97/40 strain), are reported. The vaccine was able to overcome the obstacle of MDA, and to elicit protective immunity in 100% of the pups whose antibody titres were 1:10-1:40, 83% of the pups with titres of 1:80, 57% of the pups with titres of 1:160, and even in 60% of the pups with antibody titres of 1:320. PMID- 10861196 TI - Experiment to determine limits of tolerance for fumonisin B1 in weaned piglets. AB - In Hungary almost 70% of mould-affected maize inspected since 1993 was found to be contaminated with fumonisin B1 (FB1) (mean 2.6-8.65 mg/kg; maximum 9.8-75.1 mg/kg), the degree of this contamination was found to increase from year to year (Fazekas et al., 1997b). In this experiment, in order to define tolerance limit values, the effect of exposing weaned piglets to FB1 in low doses over a 4-week period was examined. The experiment was performed with 20 weaned barrows of Danish Landrace breed. After a 5-day adaptation period cultures of the fungus Fusarium moniliforme were mixed into the animals' feed in concentrations that resulted in a daily intake of fumonisin B1 of 0, 10, 20 and 40 mg/kg feed. Feeding with the toxin was observed to exert no significant effect on body weight gain or feed consumption in the animals, no clinical signs were observed and no mortality traceable to toxic effects occurred. In computer tomography examinations performed in the second and fourth weeks mild and more severe pulmonary oedema was diagnosed in the experimental animals. The processes developing in the pulmonary parenchyma were corroborated by the mathematical and statistical evaluation procedures applied. The haematological parameters examined revealed no change attributable to toxic effects, while with respect to the biochemical parameters, an increase in aspartate aminotransferase (AST) activity dependent on dosage, indicating a pathological change in the liver, was ascertained in all three experimental groups. The free sphinganine to sphingosine ratio (SA/SO), which is regarded as the most sensitive bioindicator of fumonisin toxicosis, showed an increase proportionate to toxin concentration for all three dosages. Dissection revealed mild cases of pulmonary oedema in three of the animals given doses of 10 p.p.m. (n = 4), two mild and two severe cases in those exposed to 20 p.p.m. (n = 5), and severe cases in all five animals given 40 p.p.m. The oedema of non-inflammatory origin was confirmed by histopathological examinations. The findings of this experiment which indicate that in this study FB1 administered in substantially lower concentrations than those reported in the literature resulted in severe pathological changes, point to the importance of studies involving even lower doses. PMID- 10861197 TI - Effect of shearing on the incidence of caseous lymphadenitis in Awassi sheep in Jordan. AB - A total of 876 sheep from five flocks in north Jordan were selected to study the effect of shearing on the incidence of caseous lymphadenitis (CLA). The animals were divided into two age groups, sheep aged 1-2 years and those aged > or = 3 years. Blood samples were collected from the animals at the time of shearing and again 6 months later. A toxin enzyme-linked immunosorbent assay was used to identify sheep that had been infected with Corynebacterium pseudotuberculosis. The point prevalences of CLA were 6.59% and 21.06% in the 1-2-year and > or = 3 year age groups, respectively, and were significantly higher (P < 0.01) in the > or = 3-year age group. The overall prevalence among all ages was 15.3%. In the shorn sheep, the incidence of CLA was 22.46% and 9.47% in the 1-2-year and > or = 3-year age groups, respectively, and was significantly higher (P < 0.05) in the 1 2-year age group. In the control animals, the incidence was 8% and 5.26% in the 1 2-year and > or = 3-year age groups, respectively, and was different (P < 0.01) between the shorn (22.46%) and control (8%) animals of the 1-2-year age group. An epidemiological survey of 35 sheep farms revealed the prevalence of CLA, shearing wounds and unhygienic conditions during shearing in all farms. In conclusion, the prevalence of CLA increases with age and the incidence increases only in young sheep after shearing. Sheep are sheared under unhygienic conditions, which may be a contributing factor in increasing both the prevalence and the incidence of CLA. PMID- 10861198 TI - Type- and subtype-specific RT-PCR assays for avian influenza A viruses (AIV). AB - Reverse transcriptase (RT) PCR assays have been developed to improve the diagnosis of avian influenza A. RT-PCR using primers complementary to a conserved region of the matrix protein was assessed as being suitable for the detection of influenza A virus RNA from poultry as well as from pigs, horses and humans, regardless of the haemagglutinin (HA) and neuraminidase (NA) subtype. Therefore, this RT-PCR is a valuable tool to confirm the initial diagnosis of any influenza A infection. As a second approach, experiments were performed to identify the HA gene encoding the post-translational cleavage site of potentially highly pathogenic AIV isolates by RT-PCR. The principal aim was to design one universal primer pair for each virus subtype, H5 and H7, respectively, which allows the detection of all strain variants using only one consistent method. To realize this objective, it was necessary to develop 'wobble' primers. AIV RNAs from seven H5 and 11 H7 subtype viruses included in the investigations were specifically recognized by RT-PCR using these primers. This method therefore provides a rapid, subtype-specific diagnosis and subsequent sequencing of H5 and H7 avian influenza viruses. PMID- 10861199 TI - A survey of more than 11 years of neurologic diseases of ruminants with special reference to transmissible spongiform encephalopathies (TSEs) in Greece. AB - The first cases of scrapie were detected in Greece in a flock of sheep in October 1986. All the animals of the affected flock and all sheep in two flocks that were in contact were killed and buried. A systematic investigation of all available cases with signs indicating a neurological disease started in sheep and goats in late 1986, as well as in cattle in 1989. The investigation was based on clinical examination, necropsy or macroscopical examination of the brain and viscera, and histological examination of the brain in all animals except those with coenurosis. Histological examinations of specimens from the spinal cord and other tissues, and if considered necessary bacteriological, toxicological and serological examinations were also carried out. In October 1997, scrapie was diagnosed in sheep of a second flock (a mixed flock of sheep and goats), grazing in a pasture close to the place where scrapie was initially detected. All animals of the second flock were also killed and buried. Diagnosis in the first flock was based on clinical signs and histological lesions, and in the second immunoblotting was also used. Distinctive lesions of scrapie were found in the brain and/or the spinal cord of eight sheep with clinical signs from the two flocks. The lesions were revealed in the brain stem and/or in the cervical spinal cord, and tended to be symmetrical. In one sheep, severe lesions in the cortex of cerebral hemispheres and of the cerebellum were also found. In the brain of two sheep from the second flock the pathological isoform of PrP protein was detected. Despite the eradication scheme applied, scrapie in sheep reappeared after 11 years in a place close to where it occurred initially. This may indicate that the effectiveness of the eradication scheme implemented was not adequate and additional approaches may be needed. PMID- 10861200 TI - Taxonomic evidence that serovar 7 of Erysipelothrix strains isolated from dogs with endocarditis are Erysipelothrix tonsillarum. AB - The levels of relatedness among strains of Erysipelothrix serovar 7 isolated from dogs with endocarditis were estimated by performing DNA-DNA hybridization experiments with the type strains of Erysipelothrix rhusiopathiae and Erysipelothrix tonsillarum. All the canine strains exhibited more than 81% hybridization with the type strain of E. tonsillarum but less than 13% hybridization with the type strain of E. rhusiopathiae. Based on DNA-DNA hybridization results we confirmed that serovar 7 of the isolates from dogs with endocarditis were conclusively identified as E. tonsillarum. These results strongly indicate that some strains of genomic E. tonsillarum are a canine pathogen. PMID- 10861201 TI - Congenital thymic aplasia in a Holstein-Israeli female calf. AB - An unusual case of congenital thymic aplasia in a Holstein-Israeli female calf is described. The most prominent clinical findings were diarrhoea and weakness. At necropsy, the only significant pathological finding was the marked decrease in thymus size. Histologically, this organ was composed of loose connective tissue in which a few lymphocytes were scattered randomly. PMID- 10861202 TI - Index of suspicion. Case 2. Chronic granulomatous disease. PMID- 10861203 TI - Index of suspicion. Case 3. Otitic hydrocephalus. PMID- 10861204 TI - DNA replication and damage checkpoints and meiotic cell cycle controls in the fission and budding yeasts. AB - The cell cycle checkpoint mechanisms ensure the order of cell cycle events to preserve genomic integrity. Among these, the DNA-replication and DNA-damage checkpoints prevent chromosome segregation when DNA replication is inhibited or DNA is damaged. Recent studies have identified an outline of the regulatory networks for both of these controls, which apparently operate in all eukaryotes. In addition, it appears that these checkpoints have two arrest points, one is just before entry into mitosis and the other is prior to chromosome separation. The former point requires the central cell-cycle regulator Cdc2 kinase, whereas the latter involves several key regulators and substrates of the ubiquitin ligase called the anaphase promoting complex. Linkages between these cell-cycle regulators and several key checkpoint proteins are beginning to emerge. Recent findings on post-translational modifications and protein-protein interactions of the checkpoint proteins provide new insights into the checkpoint responses, although the functional significance of these biochemical properties often remains unclear. We have reviewed the molecular mechanisms acting at the DNA replication and DNA-damage checkpoints in the fission yeast Schizosaccharomyces pombe, and the modifications of these controls during the meiotic cell cycle. We have made comparisons with the controls in fission yeast and other organisms, mainly the distantly related budding yeast. PMID- 10861205 TI - Regulation of sterol regulatory-element binding protein 1 gene expression in liver: role of insulin and protein kinase B/cAkt. AB - Insulin stimulates the transcription of the sterol regulatory- element binding protein (SREBP) 1/ADD1 gene in liver. Hepatocytes in primary culture were used to delineate the insulin signalling pathway for induction of SREBP1 gene expression. The inhibitors of phosphoinositide 3-kinase (PI 3-kinase), wortmannin and LY 294002, abolished the insulin-dependent increase in SREBP1 mRNA, whereas the inhibitor of the mitogen- activated protein kinase cascade, PD 98059, was without effect. To investigate the role of protein kinase B (PKB)/cAkt downstream of PI 3 kinase, hepatocytes were transduced with an adenovirus encoding a PKB--oestrogen receptor fusion protein. The PKB activity of this recombinant protein was rapidly activated in hepatocytes challenged with 4-hydroxytamoxifen (OHT), as was endogenous PKB in hepatocytes challenged with insulin. The addition of OHT to transduced hepatocytes resulted in accumulation of SREBP1 mRNA, with a time course and magnitude similar to the effect of insulin in non-transduced cells. The level of SREBP1 mRNA was not increased by OHT in hepatocytes expressing a mutant form of the recombinant protein whose PKB activity was not activated by OHT. Thus acute activation of PKB is sufficient to induce SREBP1 mRNA accumulation in primary hepatocytes, and might be the major signalling event by which insulin induces SREBP1 gene expression in the liver. PMID- 10861206 TI - Activation of extracellular signal-regulated protein kinase1,2 results in down regulation of decorin expression in fibroblasts. AB - Decorin is a small leucine-rich extracellular matrix proteoglycan, the expression of which is down-regulated in proliferating and malignantly transformed cells. In the present study we show that the expression of decorin in fibroblasts is suppressed by epidermal growth factor (EGF) and PMA, and that the effect of both is potently inhibited by blocking the extracellular signal-regulated protein kinase (ERK)1,2 signalling pathway (Raf/MEK1,2/ERK1,2) with the specific MAPK/ERK kinase (MEK)1,2 inhibitor, PD98059. In addition, specific activation of ERK1,2 by adenovirus-mediated expression of constitutively active MEK1 in dermal fibroblasts results in marked reduction in decorin mRNA abundance and production. Co-transfection of NIH-3T3 fibroblasts with human decorin promoter/chloramphenicol acetyltransferase (CAT) construct (pDEC--879/CAT) in combination with the expression vectors for constitutively active Raf-1 and MEK1 markedly suppressed decorin promoter activity. Co-transfections of human decorin promoter 5'-deletion constructs with constitutively active MEK1 expression vector identified the region -278 to -188 as essential for ERK1,2 mediated down regulation of decorin promoter activity. These results show that activation of the ERK1,2 signalling pathway by a mitogenic growth factor, a tumour promoter or transformation suppresses decorin gene expression in fibroblasts, which in turn may promote proliferation and migration of normal and malignant cells. PMID- 10861207 TI - Characterization and regulation of Leishmania major 3-hydroxy-3-methylglutaryl CoA reductase. AB - In eukaryotes the enzyme 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase catalyses the synthesis of mevalonic acid, a common precursor to all isoprenoid compounds. Here we report the isolation and overexpression of the gene coding for HMG-CoA reductase from Leishmania major. The protein from Leishmania lacks the membrane domain characteristic of eukaryotic cells but exhibits sequence similarity with eukaryotic reductases. Highly purified protein was achieved by ammonium sulphate precipitation followed by chromatography on hydroxyapatite. Kinetic parameters were determined for the protozoan reductase, obtaining K(m) values for the overall reaction of 40.3+/-5.8 microM for (R,S)-HMG-CoA and 81.4+/ 5.3 microM for NADPH; V(max) was 33.55+/-1.8 units x mg(-1). Gel-filtration experiments suggested an apparent molecular mass of 184 kDa with subunits of 46 kDa. Finally, in order to achieve a better understanding of the role of this enzyme in trypanosomatids, the effect of possible regulators of isoprenoid biosynthesis in cultured promastigote cells was studied. Neither mevalonic acid nor serum sterols appear to modulate enzyme activity whereas incubation with lovastatin results in significant increases in the amount of reductase protein. Western- and Northern-blot analyses indicate that this activation is apparently performed via post-transcriptional control. PMID- 10861208 TI - Oxidation of ubiquinol by peroxynitrite: implications for protection of mitochondria against nitrosative damage. AB - A major pathway of nitric oxide utilization in mitochondria is its conversion to peroxynitrite, a species involved in biomolecule damage via oxidation, hydroxylation and nitration reactions. In the present study the potential role of mitochondrial ubiquinol in protecting against peroxynitrite-mediated damage is examined and the requirements of the mitochondrial redox status that support this function of ubiquinol are established. (1) Absorption and EPR spectroscopy studies revealed that the reactions involved in the ubiquinol/peroxynitrite interaction were first-order in peroxynitrite and zero-order in ubiquinol, in agreement with the rate-limiting formation of a reactive intermediate formed during the isomerization of peroxynitrite to nitrate. Ubiquinol oxidation occurred in one-electron transfer steps as indicated by the formation of ubisemiquinone. (2) Peroxynitrite promoted, in a concentration-dependent manner, the formation of superoxide anion by mitochondrial membranes. (3) Ubiquinol protected against peroxynitrite-mediated nitration of tyrosine residues in albumin and mitochondrial membranes, as suggested by experimental models, entailing either addition of ubiquinol or expansion of the mitochondrial ubiquinol pool caused by selective inhibitors of complexes III and IV. (4) Increase in membrane-bound ubiquinol partially prevented the loss of mitochondrial respiratory function induced by peroxynitrite. These findings are analysed in terms of the redox transitions of ubiquinone linked to both nitrogen centred radical scavenging and oxygen-centred radical production. It may be concluded that the reaction of mitochondrial ubiquinol with peroxynitrite is part of a complex regulatory mechanism with implications for mitochondrial function and integrity. PMID- 10861209 TI - Acylphosphatase possesses nucleoside triphosphatase and nucleoside diphosphatase activities. AB - We have demonstrated that acylphosphatase possesses ATP-diphosphohydrolase (apyrase-like) activity. In fact, acylphosphatase first catalyses the hydrolysis of the gamma-phosphate group of nucleoside triphosphates, and then attacks the beta-phosphate group of the initially produced nucleoside diphosphates, generating nucleoside monophosphates. In contrast, it binds nucleoside monophosphates but does not catalyse their hydrolyses. The calculated k(cat) values for the nucleoside triphosphatase activity of acylphosphatase are of the same order of magnitude as those displayed by certain G-proteins. An acidic environment enhances the apyrase-like activity of acylphosphatase. The true nucleotide substrates of acylphosphatase are free nucleoside di- and triphosphates, as indicated by the Mg(2+) ion inhibition of the activity. We have also demonstrated that, although nucleoside triphosphates are still hydrolysed at pH 7.2 and 37 degrees C, in the presence of millimolar Mg(2+) concentrations this occurs at a lower rate. Taken together with the previously observed strong increase of acylphosphatase levels during induced cell differentiation, our findings suggest that acylphosphatase plays an active role in the differentiation process (as well as in other processes, such as apoptosis) by modulating the ratio between the cellular levels of nucleoside diphosphates and nucleoside triphosphates. PMID- 10861210 TI - Processing of N-linked oligosaccharide depends on its location in the anion exchanger, AE1, membrane glycoprotein. AB - The human erythrocyte anion exchanger (AE)1 (Band 3) contains a single complex N linked oligosaccharide that is attached to Asn(642) in the fourth extracellular loop of this polytopic membrane protein, while other isoforms (AE2, AE3 and trout AE1) are N-glycosylated on the preceding extracellular loop. Human AE1 expressed in transfected human embryonic kidney (HEK)-293 or COS-7 cells contained a high mannose oligosaccharide. The lack of oligosaccharide processing was not due to retention of AE1 in the endoplasmic reticulum since biotinylation assays showed that approx. 30% of the protein was expressed at the cell surface. Moving the N glycosylation site to the preceding extracellular loop in an AE1 glycosylation mutant (N555) resulted in processing of the oligosaccharide and production of a complex form of AE1. A double N-glycosylation mutant (N555/N642) contained both a high-mannose and a complex oligosaccharide chain. The complex form of the N555 mutant could be biotinylated showing that this form of the glycoprotein was at the cell surface. Pulse-chase experiments showed that the N555 mutant was efficiently converted from a high-mannose to a complex oligosaccharide with a half-time of approx. 4 h, which reflected the time course of trafficking of AE1 from the endoplasmic reticulum to the plasma membrane. The turnover of the complex form of the N555 mutant occurred with a half-life of approx. 15 h. The results show that the oligosaccharide attached to the endogenous site in extracellular loop 4 in human AE1 is not processed in HEK-293 or COS-7 cells, while the oligosaccharide attached to the preceding loop is converted into the complex form. PMID- 10861211 TI - Activation of phosphatidylinositol 3-kinase is required for transcriptional activity of F-type 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: assessment of the role of protein kinase B and p70 S6 kinase. AB - Previous studies have demonstrated that the F isoform of6 phosphofructo-2-kinase/fructose-2,6-bisphosphatase(6PF2K/Fru-2,6-BPase) is transcriptionally regulated by growth factors. The aim of this study was to investigate the importance of the phosphatidylinositol 3-kinase (PI 3-kinase) pathway in the regulation of 6PF2K/Fru-2,6-BPase gene expression. We have completed studies using chemical inhibitors and expression vectors for the proteins involved in this signalling cascade. Treatment of cells with LY 294002, an inhibitor of PI 3-kinase, blocked the epidermal growth factor (EGF)-dependent stimulation of 6PF2K/Fru-2,6-BPase gene transcription. Transient transfection of a constitutively active PI 3-kinase was sufficient to activate transcription from the F-type 6PF2K/Fru-2,6-BPase promoter. In contrast, co-transfection with a dominant-negative form of PI 3-kinase completely abrogated the stimulation by EGF, and down-regulated the basal promoter activity. In an attempt to determine downstream proteins that lie between PI 3-kinase and 6PF2K/Fru-2,6-BPase gene expression, the overexpression of a constitutively active form of protein kinase B (PKB) was sufficient to activate 6PF2K/Fru-2,6-BPase gene expression, even in the presence of either a dominant-negative form of PI 3-kinase or LY 294002. The over-expression of p70/p85 ribosomal S6 kinase or the treatment with its inhibitor rapamycin did not affect 6PF2K/Fru-2,6-BPase transcription. We conclude that PI 3-kinase is necessary for the transcriptional activity of F-type 6PF2K/Fru-2,6-BPase, and that PKB is a downstream effector of PI 3-kinase directly involved in the regulation of 6PF2K/Fru-2,6-BPase gene expression. PMID- 10861212 TI - Identification of a nucleoside/nucleobase transporter from Plasmodium falciparum, a novel target for anti-malarial chemotherapy. AB - Plasmodium, the aetiologic agent of malaria, cannot synthesize purines de novo, and hence depends upon salvage from the host. Here we describe the molecular cloning and functional expression in Xenopus oocytes of the first purine transporter to be identified in this parasite. This 422-residue protein, which we designate PfENT1, is predicted to contain 11 membrane-spanning segments and is a distantly related member of the widely distributed eukaryotic protein family the equilibrative nucleoside transporters (ENTs). However, it differs profoundly at the sequence and functional levels from its homologous counterparts in the human host. The parasite protein exhibits a broad substrate specificity for natural nucleosides, but transports the purine nucleoside adenosine with a considerably higher apparent affinity (K(m) 0.32+/-0.05 mM) than the pyrimidine nucleoside uridine (K(m) 3.5+/-1.1 mM). It also efficiently transports nucleobases such as adenine (K(m) 0.32+/-0.10 mM) and hypoxanthine (K(m) 0.41+/-0.1 mM), and anti viral 3'-deoxynucleoside analogues. Moreover, it is not sensitive to classical inhibitors of mammalian ENTs, including NBMPR [6-[(4-nitrobenzyl)thio]-9-beta-D ribofuranosylpurine, or nitrobenzylthioinosine] and the coronary vasoactive drugs, dipyridamole, dilazep and draflazine. These unique properties suggest that PfENT1 might be a viable target for the development of novel anti-malarial drugs. PMID- 10861213 TI - Apolipoprotein E includes a binding site which is recognized by several amyloidogenic polypeptides. AB - Inheritance of the apolipoprotein E (apoE) epsilon 4 allele is a risk factor for late-onset Alzheimer's disease (AD). Biochemically apoE is present in AD plaques and neurofibrillary tangles of the AD brain. There is a high avidity and specific binding of apoE and the amyloid beta-peptide (A beta). In addition to AD apoE is also present in many other cerebral and systemic amyloidoses, Down's syndrome and prion diseases but the pathophysiological basis for its presence is still unknown. In the present study we have compared the interaction of apoE with A beta, the gelsolin-derived amyloid fragment AGel(183-210) and the amyloidogenic prion fragments PrP(109-122) and PrP(109-141). We show that, similar to A beta, also AGel and PrP fragments can form a complex with apoE, and that the interaction between apoE and the amyloidogenic protein fragments is mediated through the same binding site on apoE. We also show that apoE increases the thioflavin-T fluorescence of PrP and AGel and that apoE influences the content of beta-sheet conformation of these amyloidogenic fragments. Our results indicate that amyloids and amyloidogenic prion fragments share a similar structural motif, which is recognized by apoE, possibly through a single binding site, and that this motif is also responsible for the amyloidogenicity of these fragments. PMID- 10861214 TI - Redox equilibria of manganese peroxidase from Phanerochaetes chrysosporium: functional role of residues on the proximal side of the haem pocket. AB - Redox potentials of recombinant manganese peroxidase from Phanerochaetes chrysosporium have been measured by cyclic voltammetry as a function of pH, between pH 4.5 and pH 10.5. They display a bimodal behaviour (characterized by an 'alkaline' and an 'acid' transition), which indicates that (at least) two protonating groups change their pK(b) values upon reduction (and/or oxidation) of the iron atom in haem. Analogous measurements have been carried out on four site directed mutants involving residues in close proximity to the proximal ligand, His(173), in order to investigate the role played by residues of the proximal haem pocket on the redox properties of this enzyme. Results obtained suggest that the protonation state of N(delta) of the proximal imidazole group is redox-linked and that it is crucial in regulating the 'alkaline' transition. On the other hand, none of the proximal mutants alters the 'acid' transition, suggesting that it is modulated by groups located in a different portion of the protein. PMID- 10861215 TI - Glucocorticoids induce a near-total suppression of hyaluronan synthase mRNA in dermal fibroblasts and in osteoblasts: a molecular mechanism contributing to organ atrophy. AB - Glucocorticoid (GC) administration induces atrophy of skin, bone, and other organs, partly by reducing tissue content of glycosaminoglycans, particularly hyaluronic acid (HA). We took advantage of the recent cloning of the three human hyaluronan synthase (HAS) enzymes (HAS1, HAS2 and HAS3), to explore the molecular mechanisms of this side effect. Northern and slot blots performed on RNA extracted from cultured dermal fibroblasts and the MG-63 osteoblast-like osteosarcoma cell line indicated that HAS2 is the predominant HAS mRNA in these cells. Incubation of both cell types for 24 h in the presence of 10(-6) M dexamethasone (DEX) resulted in a striking 97--98% suppression of HAS2 mRNA levels. Time-course studies in fibroblasts demonstrated suppression of HAS2 mRNA to 28% of control by 1 h, and to 1.2% of control by 2 h, after addition of DEX. Dose-response studies in fibroblasts indicated that the majority of the suppressive effect required concentrations characteristic of cell-surface GC receptors, a point confirmed by persistent DEX-induced suppression in the presence of RU486, an antagonist of classic cytosolic steroid hormone receptors. Nuclear run-off experiments showed a 70% suppression of HAS2 gene transcription in nuclei from DEX-treated fibroblasts, which is unlikely to fully explain the rapid 50--80-fold reduction in message levels. Experiments with actinomycin D (AMD) demonstrated that the message half-life was 25 min in cells without DEX, whereas the combination of AMD with DEX dramatically increased the half-life of HAS2 mRNA, suggesting that DEX acts by inducing a short-lived destabilizer of the HAS2 message. Direct assessment of HAS2 mRNA stability by wash-out of incorporated uridine label established a half-life of 31 min in cells without DEX, which substantially shortened in the presence of DEX. In conclusion, GCs induce a rapid and sustained, near-total suppression of HAS2 message levels, mediated through substantial decreases in both gene transcription and message stability. These effects may contribute to the loss of HA in GC-treated organs. PMID- 10861216 TI - Indirect induction of suppressor of cytokine signalling-1 in macrophages stimulated with bacterial lipopolysaccharide: partial role of autocrine/paracrine interferon-alpha/beta. AB - It has previously been reported by us that a brief prior exposure of mouse bone marrow culture-derived macrophages to bacterial lipopolysaccharide (LPS) resulted in a dramatic reduction in their ability to produce NO in response to a subsequent stimulus with either interferon-gamma (IFN-gamma) or IFN-gamma plus LPS. We show here that this brief exposure to LPS results in an impaired response to subsequently added IFN-gamma. A 2--4 h pretreatment with LPS leads to a dramatic reduction in the IFN-gamma-induced DNA-binding of the transcription factor, signal transducer and activator of transcription 1 alpha (STAT1 alpha). This loss in ability to activate STAT1 alpha temporally correlates with the LPS induced accumulation of mRNA encoding the suppressor of cytokine signalling-1 (SOCS-1). However, LPS does not directly induce the synthesis of SOCS-1. Rather, LPS induces the synthesis of autocrine/paracrine factors that are the true mediators of SOCS-1 induction. IFN-alpha/beta is one of these mediators, but plays only a partial role in the induction of SOCS-1 because neutralization of LPS-induced IFN-alpha/beta production incompletely inhibits the induction of SOCS 1. We show that mouse IFN-beta directly induces the synthesis of SOCS-1, without the need for prior protein synthesis, and does so with faster kinetics than does LPS. Our results are consistent with the non-specific nature of LPS-induced tolerance and provide a mechanistic insight into nonspecificity; LPS indirectly induces the synthesis of a protein mediator, SOCS-1, which inhibits the signalling that is induced by IFN-gamma. PMID- 10861217 TI - Characterization of the actin filament capping state in human erythrocyte ghost and cytoskeletal preparations. AB - The narrow Gaussian-length-distribution of actin filaments forming the cytoskeleton of the human erythrocyte indicates the existence of strict mechanisms for length determination and maintenance. A similar regulation is achieved in striated muscle by the capping of both the ends of the thin filaments, which consequently prevents monomer exchange. However, the ability of erythroid cytoskeletal preparations to nucleate actin polymerization has led to the proliferation of the idea that at least the barbed ends of the actin filaments are uncapped. The mechanism by which the length of the filaments is thus maintained has been left open to debate. In an effort to resolve any doubt regarding length-maintenance in human erythrocytes we have characterized the capping state of the actin filaments in a number of different ghost and cytoskeletal preparations. Under conditions of sufficiently high bivalent-cation concentration the actin filaments retain functional caps at both the barbed and pointed ends. Hence filament capping at both ends prevents redistribution of the actin monomer in a similar manner to that proposed for the thin filaments of striated muscle. Actin filament uncapping is apparently caused by the centrifugal shearing stress imposed during ghost preparation. The uncapping is more pronounced when the bivalent-cation concentration is reduced or when the membrane is removed by detergents. The effects of bivalent cations seem to be mediated through the erythroid protein spectrin, consistent with the hypothesis of Wallis et al. [Wallis, Babitch and Wenegieme (1993) Biochemistry 32, 5045--5050] that the ability of spectrin to resist shearing stress is dependent on the degree of bound bivalent cations. PMID- 10861218 TI - p40(phox) Participates in the activation of NADPH oxidase by increasing the affinity of p47(phox) for flavocytochrome b(558). AB - NADPH oxidase is one of the major components of the innate immune system and is used by phagocytes to generate microbicidal reactive oxygen species. Activation of the enzyme requires the participation of a minimum of five proteins, p22(phox), gp91(phox) (together forming flavocytochrome b(558)), p47(phox), p67(phox) and the GTP-binding protein, Rac2. A sixth protein, p40(phox), has been implicated in the control of the activity of NADPH oxidase principally based on its sequence homology to, and physical association with, other phox components, and also the observation that it is phosphorylated during neutrophil activation. However, to date its role in regulating the activity of the enzyme has remained obscure, with evidence for both positive and negative influences on oxidase activity having being reported. Data are presented here using the cell-free system for NADPH oxidase activation that shows that p40(phox) can function to promote oxidase activation by increasing the affinity of p47(phox) for the enzyme approx. 3-fold. PMID- 10861219 TI - Mitochondrial ATP production is necessary for activation of the extracellular signal-regulated kinases during ischaemia/reperfusion in rat myocyte-derived H9c2 cells. AB - To search for the stimuli involved in activating the mitogen-activated protein kinases (MAPKs) during ischaemia and reperfusion, we simulated the event in a system in vitro conducive to continuous and non-invasive measurements of several major perturbations that occur at the time: O(2) tension, mitochondrial respiration and energy status. Using H9c2 cells (a clonal line derived from rat heart), we found that activation of the extracellular signal-regulated MAPKs (ERKs) on reoxygenation was abolished if the mitochondria were inhibited prior to and during reoxygenation. Re-introduction of O(2) per se is therefore not sufficient to activate the ERKs. Recovery and maintenance of cellular ATP levels by mitochondrial respiration is necessary, although ATP recovery alone is not sufficient. ERK activation by H(2)O(2), but not phorbol esters, was also sensitive to mitochondrial inhibition. Thus, reoxygenation and H(2)O(2)-mediated oxidative stress share a mechanism of ERK activation that is ATP- or mitochondrion-dependent, and this common feature suggests that the reoxygenation response is mediated by reactive oxygen species. A correlation between ERK activity and ATP levels was also found during the anoxic phase of ischaemia, an effect that was not due to substrate limitation for the kinases. Our results reveal the importance of cellular metabolism in ERK activation, and introduce ATP as a novel participant in the mechanisms underlying the ERK cascade. PMID- 10861220 TI - The role of phosphatidylcholine in fatty acid exchange and desaturation in Brassica napus L. leaves. AB - The role of phosphatidylcholine (PC) in fatty acid exchange and desaturation was examined and compared with that of monogalactosyldiacylglycerol (MGDG) in Brassica napus leaves using (14)C-labelling in vivo. Data are presented which indicate that in the chloroplast newly formed saturated (palmitic acid, 16:0) and monounsaturated (oleic acid, 18:1) fatty acid is incorporated into MGDG and desaturated in situ. In the non-plastidic compartments, however, newly formed fatty acid is exchanged with polyunsaturated fatty acid in PC, the probable major site of subsequent desaturation. The unsaturated fatty acid is released to the acyl-CoA pool, which is then used to synthesize diacylglycerol (DAG) containing a high level of unsaturated fatty acid. This highly unsaturated DAG may be the source for the biosynthesis of other cellular glycerolipids. The generally accepted pathway in which PC is synthesized from molecular species of DAG containing 16:0 and 18:1 followed by desaturation of the 18:1 to linoleic (18:2) and linolenic (18:3) acids is questioned. PMID- 10861221 TI - The short prodomain influences caspase-3 activation in HeLa cells. AB - Proteolytic activation of caspases is a key step in the process of apoptosis. According to their primary structure, caspases can be divided into a group with a long prodomain and a group with a short prodomain. Whereas long prodomains play a role in autocatalytic processing, little is known about the function of the short prodomain, for example the prodomain of caspase-3. We constructed caspase-3 variants lacking the prodomain and overexpressed these in HeLa and yeast cells. We found that removal of the caspase-3 prodomain resulted in spontaneous proteolytic activation of the protein when expressed in HeLa cells. This processing was only partially autocatalytic, as demonstrated by a catalytically inactive caspase-3 mutant. Co-expression of the anti-apoptotic protein XIAP (X chromosome-linked inhibitor of apoptosis protein) completely blocked the observed spontaneous activation, which excluded a direct involvement of caspase-8. Our findings indicate that the short prodomain of caspase-3 serves as a silencing component in mammalian cells by retaining this executioner caspase in an inactive state. PMID- 10861222 TI - Expression, purification and characterization of recombinant human choline acetyltransferase: phosphorylation of the enzyme regulates catalytic activity. AB - Choline acetyltransferase synthesizes acetylcholine in cholinergic neurons and, in humans, may be produced in 82- and 69-kDa forms. In this study, recombinant choline acetyltransferase from baculovirus and bacterial expression systems was used to identify protein isoforms by two-dimensional SDS/PAGE and as substrate for protein kinases. Whereas hexa-histidine-tagged 82- and 69-kDa enzymes did not resolve as individual isoforms on two-dimensional gels, separation of wild-type choline acetyltransferase expressed in insect cells revealed at least nine isoforms for the 69-kDa enzyme and at least six isoforms for the 82-kDa enzyme. Non-phosphorylated wild-type choline acetyltransferase expressed in Escherichia coli yielded six (69 kDa) and four isoforms (82 kDa) respectively. Immunofluorescent labelling of insect cells expressing enzyme showed differential subcellular localization with the 69-kDa enzyme localized adjacent to plasma membrane and the 82-kDa enzyme being cytoplasmic at 24 h. By 64 h, the 69-kDa form was in cytoplasm and the 82-kDa form was only present in nucleus. Studies in vitro showed that recombinant 69-kDa enzyme was a substrate for protein kinase C (PKC), casein kinase II (CK2) and alpha-calcium/calmodulin-dependent protein kinase II (alpha-CaM kinase), but not for cAMP-dependent protein kinase (PKA); phosphorylation by PKC and CK2 enhanced enzyme activity. The 82-kDa enzyme was a substrate for PKC and CK2 but not for PKA or alpha-CaM kinase, with only PKC yielding increased enzyme activity. Dephosphorylation of both forms of enzyme by alkaline phosphatase decreased enzymic activity. These studies are of functional significance as they report for the first time that phosphorylation enhances choline acetyltransferase catalytic activity. PMID- 10861223 TI - Identification and characterization of a novel cytochrome c(3) from Shewanella frigidimarina that is involved in Fe(III) respiration. AB - Shewanella frigidimarina NCIMB400 is a non-fermenting, facultative anaerobe from the gamma group of proteobacteria. When grown anaerobically this organism produces a wide variety of periplasmic c-type cytochromes, mostly of unknown function. We have purified a small, acidic, low-potential tetrahaem cytochrome with similarities to the cytochromes c(3) from sulphate-reducing bacteria. The N terminal sequence was used to design PCR primers and the cctA gene encoding cytochrome c(3) was isolated and sequenced. The EPR spectrum of purified cytochrome c(3) indicates that all four haem irons are ligated by two histidine residues, a conclusion supported by the presence of eight histidine residues in the polypeptide sequence, each of which is conserved in a related cytochrome c(3) and in the cytochrome domains of flavocytochromes c(3). All four haems exhibit low midpoint redox potentials that range from -207 to -58 mV at pH 7; these values are not significantly influenced by pH changes. Shewanella cytochrome c(3) consists of a mere 86 amino acid residues with a predicted molecular mass of 11780 Da, including the four attached haem groups. This corresponds closely to the value of 11778 Da estimated by electrospray MS. To examine the function of this novel cytochrome c(3) we constructed a null mutant by gene disruption. S. frigidimarina lacking cytochrome c(3) grows well aerobically and its growth rate under anaerobiosis with a variety of electron acceptors is indistinguishable from that of the wild-type parent strain, except that respiration with Fe(III) as sole acceptor is severely, although not completely, impaired. PMID- 10861224 TI - Identification of three human type-II classic cadherins and frequent heterophilic interactions between different subclasses of type-II classic cadherins. AB - We identified three novel human type-II classic cadherins, cadherin-7, -9 and 10, by cDNA cloning and sequencing, and confirmed that they interact with catenins and function in cell-cell adhesion as do other classic cadherins. Cell cell binding activities of the eight human type-II classic cadherins, including the three new molecules, were evaluated by long-term cell-aggregation experiments using mouse L fibroblast clones transfected with the individual cadherins. The experiments indicated that all the type-II cadherins appeared to possess similar binding strength, which was virtually equivalent to that of E-cadherin. We next examined the binding specificities of the type-II cadherins using the mixed cell aggregation assay. Although all of the type-II cadherins exhibited binding specificities distinct from that of E-cadherin, heterophilic interactions ranging from incomplete to complete were frequently observed among them. The combinations of cadherin-6 and -9, cadherin-7 and -14, cadherin-8 and -11, and cadherin-9 and 10 interacted in a complete manner, and in particular cadherin-7 and -14, and cadherin-8 and -11 showed an indistinguishable binding specificity against other cadherin subclasses, at least in this assay system. Although these data were obtained from an in vitro study, they should be useful for understanding cadherin mediated mechanisms of development, morphogenesis and cell-cell interactions in vivo. PMID- 10861225 TI - The pepsin residue glycine-76 contributes to active-site loop flexibility and participates in catalysis. AB - Glycine residues are known to contribute to conformational flexibility of polypeptide chains, and have been found to contribute to flexibility of some loops associated with enzymic catalysis. A comparison of porcine pepsin in zymogen, mature and inhibited forms revealed that a loop (a flap), consisting of residues 71--80, located near the active site changed its position upon substrate binding. The loop residue, glycine-76, has been implicated in the catalytic process and thought to participate in a hydrogen-bond network aligning the substrate. This study investigated the role of glycine-76 using site-directed mutagenesis. Three mutants, G76A, G76V and G76S, were constructed to increase conformational restriction of a polypeptide chain. In addition, the serine mutant introduced a hydrogen-bonding potential at position 76 similar to that observed in human renin. All the mutants, regardless of amino acid size and polarity, had lower catalytic efficiency and activated more slowly than the wild-type enzyme. The slower activation process was associated directly with altered proteolytic activity. Consequently, it was proposed that a proteolytic cleavage represents a limiting step of the activation process. Lower catalytic efficiency of the mutants was explained as a decrease in the flap flexibility and, therefore, a different pattern of hydrogen bonds responsible for substrate alignment and flap conformation. The results demonstrated that flap flexibility is essential for efficient catalytic and activation processes. PMID- 10861226 TI - Reconstitution of purified, active and malonyl-CoA-sensitive rat liver carnitine palmitoyltransferase I: relationship between membrane environment and malonyl-CoA sensitivity. AB - Carnitine palmitoyltransferase I (CPT I) catalyses the initial step of fatty acid import into the mitochondrial matrix, the site of beta-oxidation, and its inhibition by malonyl-CoA is a primary control point for this process. The enzyme exists in at least two isoforms, denoted L-CPT I (liver type) and M-CPT I (skeletal-muscle type), which differ in their kinetic characteristics and tissue distributions. A property apparently unique to L-CPT I is that its sensitivity to malonyl-CoA decreases in vivo with fasting or experimentally induced diabetes. The mechanism of this important regulatory effect is unknown and has aroused much interest. CPT I is an integral outer-membrane protein and displays little activity after removal from the membrane by detergents, precluding direct purification of active protein by conventional means. Here we describe the expression of a 6 x His-tagged rat L-CPT I in Pichia pastoris and purification of the detergent-solubilized enzyme in milligram quantities. Reconstitution of the purified product into a liposomal environment yielded a 200--400-fold increase in enzymic activity and restored malonyl-CoA sensitivity. This is the first time that a CPT I protein has been available for study in a form that is both pure and active. Comparison of the kinetic properties of the reconstituted material with those of L-CPT I as it exists in mitochondria prepared from yeast over-expressing the enzyme and in livers from fed or fasted rats permitted novel insight into several aspects of the enzyme's behaviour. The malonyl-CoA response of the liposomal enzyme was found to be greater when the reconstitution procedure was carried out at 22 degrees C compared with 4 degrees C (IC(50) approximately 11 microM versus 30 microM, respectively). When the sensitivities of L-CPT I in each of the different environments were compared, they were found to decrease in the following order: fed liver>fasted liver approximately liposomes prepared at 22 degrees C approximately P. pastoris mitochondria>liposomes prepared at 4 degrees C. In addition, pre-treatment of L-CPT I liposomes with the membrane-fluidizing reagent benzyl alcohol caused densensitization to the inhibitor. In contrast with the variable response to malonyl-CoA, the liposomal L-CPT I displayed a pH profile and kinetics with regard to the carnitine and acyl-CoA substrates similar to those of the enzyme in fed or fasted liver mitochondria. However, despite a normal sensitivity to malonyl-CoA, L-CPT I in P. pastoris mitochondria displayed aberrant behaviour with regard to each of these other parameters. The kinetic data establish several novel points. First, even after stringent purification procedures in the presence of detergent, recombinant L-CPT I could be reconstituted in active, malonyl-CoA sensitive form. Second, the kinetics of the reconstituted, 6 x His-tagged L-CPT I with regard to substrate and pH responses were similar to what is observed with rat liver mitochondria (whereas in P. pastoris mitochondria the enzyme behaved anomalously), confirming that the purified preparation is a suitable model for studying the functional properties of the enzyme. Third, wide variation in the response to the inhibitor, malonyl CoA, was observed depending only on the enzyme's membrane environment and independent of interaction with other proteins. In particular, the fluidity of the membrane had a direct influence on this parameter. These observations may help to explain the mechanism of the physiological changes in the properties of L CPT I that occur in vivo and are consistent with the current topographical model of the enzyme. PMID- 10861227 TI - Lithocholic acid and sulphated lithocholic acid differ in the ability to promote matrix metalloproteinase secretion in the human colon cancer cell line CaCo-2. AB - The human colon carcinoma cell line CaCo-2 has the ability to sulphate the secondary bile acid lithocholic acid (LA), whereas other primary or secondary bile acids were not sulphated [Halvorsen, Kase, Prydz, Gharagozlian, Andresen and Kolset (1999) Biochem. J. 343, 533--539]. To study the biological implications of this modification, CaCo-2 cells were incubated with either LA or sulphated lithocholic acid (3-sulpholithocholic acid, SLA), and in some experiments with taurine-conjugated lithocholic acid. Increased secretion of matrix metalloproteinases (MMPs) correlates with transformation of colon epithelial cells. When CaCo-2 cells were incubated with LA, the secretion of MMP-2 was found to increase approx. 60% when analysed by gelatin zymography, and 80% when analysed by Western blotting. SLA, in contrast, did not affect the level of MMP-2 secretion, and after zymography the level of enzyme activity was 78% of control values after 18 h incubation. The secretion of MMPs is linked to increased cellular invasion and, in tumours, to increased capacity for metastasis. The ability of CaCo-2 cells to invade in a chamber assay was stimulated after exposure to LA, whereas SLA-treated cells did not differ from control cells. LA therefore seems to induce a more invasive CaCo-2 cell phenotype, as judged by the two parameters tested, whereas the sulphated counterpart, SLA, did not have these effects. Sulphation of LA in the colon may be an important mechanism to decrease the potential LA has to promote a malignant epithelial phenotype. PMID- 10861228 TI - Pyruvate dehydrogenase kinase from Arabidopsis thaliana: a protein histidine kinase that phosphorylates serine residues. AB - Pyruvate dehydrogenase kinase (PDK) is the primary regulator of flux through the mitochondrial pyruvate dehydrogenase complex (PDC). Although PDKs inactivate mitochondrial PDC by phosphorylating specific Ser residues, the primary amino acid sequence indicates that they are more closely related to prokaryotic His kinases than to eukaryotic Ser/Thr kinases. Unlike Ser/Thr kinases, His kinases use a conserved His residue for phosphotransfer to Asp residues. To understand these unique kinases better, a presumptive PDK from Arabidopsis thaliana was heterologously expressed and purified for this investigation. Purified, recombinant A. thaliana PDK could inactivate kinase-depleted maize mitochondrial PDC by phosphorylating Ser residues. Additionally, A. thaliana PDK was capable of autophosphorylating Ser residues near its N-terminus, although this reaction is not part of the phosphotransfer pathway. To elucidate the mechanism involved, we performed site-directed mutagenesis of the canonical His residue likely to be involved in phosphotransfer. When His-121 was mutated to Ala or Gln, Ser autophosphorylation was decreased by 50% and transphosphorylation of PDC was decreased concomitantly. We postulate that either (1) His-121 is not the sole phosphotransfer His residue or (2) mutagenesis of His-121 exposes an additional otherwise cryptic phosphotransfer His residue. Thus His-121 is one residue involved in kinase function. PMID- 10861229 TI - Unifying mechanism for Aplysia ADP-ribosyl cyclase and CD38/NAD(+) glycohydrolases. AB - Highly purified Aplysia californica ADP-ribosyl cyclase was found to be a multifunctional enzyme. In addition to the known transformation of NAD(+) into cADP-ribose this enzyme is able to catalyse the solvolysis (hydrolysis and methanolysis) of cADP-ribose. This cADP-ribose hydrolase activity, which becomes detectable only at high concentrations of the enzyme, is amplified with analogues such as pyridine adenine dinucleotide, in which the cleavage rate of the pyridinium-ribose bond is much reduced compared with NAD(+). Although the specificity ratio V(max)/K(m) is in favour of NAD(+) by 4 orders of magnitude, this multifunctionality allowed us to propose a 'partitioning' reaction scheme for the Aplysia enzyme, similar to that established previously for mammalian CD38/NAD(+) glycohydrolases. This mechanism involves the formation of a single oxocarbenium-type intermediate that partitions to cADP-ribose and solvolytic products via competing pathways. In favour of this mechanism was the finding that the enzyme also catalysed the hydrolysis of NMN(+), a substrate that cannot undergo cyclization. The major difference between the mammalian and the invertebrate enzymes resides in their relative cyclization/hydrolysis rate constant ratios, which dictate their respective yields of cADP-ribose (ADP ribosyl cyclase activity) and ADP-ribose (NAD(+) glycohydrolase activity). For the Aplysia enzyme's catalysed transformation of NAD(+) we favour a mechanism where the formation of cADP-ribose precedes that of ADP-ribose; i.e. macroscopically the invertebrate ADP-ribosyl cyclase conforms to a sequential reaction pathway as a limiting form of the partitioning mechanism. PMID- 10861230 TI - Investigation of the slow inhibition of almond beta-glucosidase and yeast isomaltase by 1-azasugar inhibitors: evidence for the 'direct binding' model. AB - (-)-1-Azafagomine [(3R,4R,5R)-4,5-dihydroxy-3-hydroxymethylhexahydropyridazine; inhibitor 1] is a potent glycosidase inhibitor designed to mimic the transition state of a substrate undergoing glycoside cleavage. The inhibition of glycosidases by inhbitor 1 and analogues has been found to be a relatively slow process. This 'slow inhibition' process was investigated in the inhibition of almond beta-glucosidase and yeast isomaltase by inhibitor 1 and analogues. Progress-curve experiments established that the time-dependent inhibition of both enzymes by inhibitor 1 was a consequence of relatively slow dissociation and association of the inhibitor from and to the enzyme, and not a result of slow interchanges between protein conformations. A number of hydrazine-containing analogues of inhibitor 1 also inhibited beta-glucosidase and isomaltase slowly, while the amine isofagomine [(3R,4R,5R)-3,4-dihydroxy-5-hydroxymethylpiperidine; inhibitor 5] only inhibited beta-glucosidase slowly. Inhibitor 1 and related inhibitors were found to leave almond beta-glucosidase with almost identical rate constants, so that the difference in K(i) values depended almost entirely on changes in the binding rate constant, k(on). The same trend was observed for the inhibition of yeast isomaltase by inhibitor 1 and a related inhibitor. The values of the rate constants were obtained at 25 degrees C and at pH 6.8. PMID- 10861231 TI - Interaction of C3b(2)--IgG complexes with complement proteins properdin, factor B and factor H: implications for amplification. AB - Nascent C3b can form ester bonds with various target molecules on the cell surface and in the fluid phase. Previously, we showed that C3b(2)--IgG complexes represent the major covalent product of C3 activation in serum [Lutz, Stammler, Jelezarova, Nater and Spath (1996) Blood 88, 184--193]. In the present report, binding of alternative pathway proteins to purified C3b(2)--IgG complexes was studied in the fluid phase by using biotinylated IgG for C3b(2)--IgG generation and avidin-coated plates to capture complexes. Up to seven moles of properdin 'monomer' bound per mole of C3b(2)--IgG at physiological conditions in the absence of any other complement protein. At low properdin/C3b(2)--IgG ratios bivalent binding was preferred. Neither factor H nor factor B affected properdin binding. On the other hand, properdin strongly stimulated factor B binding. Interactions of all three proteins with C3b(2)--IgG exhibited pH optima. An ionic strength optimum was most pronounced for properdin, while factor B binding was largely independent of the salt concentration. C3b(2)--IgG complexes were powerful precursors of the alternative pathway C3 convertase. In the presence of properdin, C3 convertase generated from C3b(2)--IgG cleaved about sevenfold more C3 than the enzyme generated on C3b. C3b(2)--IgG complexes could therefore maintain the amplification loop of complement longer than free C3b. PMID- 10861232 TI - Epidermal growth factor regulation of glutathione S-transferase gene expression in the rat is mediated by class Pi glutathione S-transferase enhancer I. AB - Using chloramphenicol acetyltransferase assays we showed that epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha), and 3,3',4,4',5 pentachlorobiphenyl (PenCB) induce class Pi glutathione S-transferase (GSTP1) in primary cultured rat liver parenchymal cells. GSTP1 enhancer I (GPEI), which is required for the stimulation of GSTP1 expression by PenCB, also mediates EGF and TGF alpha stimulation of GSTP1 gene expression. However, hepatocyte growth factor and insulin did not stimulate GPEI-mediated gene expression. On the other hand, the antioxidant reagents butylhydroxyanisole and t-butylhydroquinone, stimulated GPEI-mediated gene expression, but the level of GSTP1 mRNA was not elevated. Our observations suggest that EGF and TGF alpha induce GSTP1 by the same signal transduction pathway as PenCB. Since the sequence of GPEI is similar to that of the antioxidant responsive element (ARE), some factors which bind to ARE might play a role in GPEI-mediated gene expression. PMID- 10861233 TI - Low-density lipoprotein activates the small GTPases Rap1 and Ral in human platelets. AB - Physiological concentrations of low-density lipoprotein (LDL) sensitize blood platelets to alpha-thrombin- and collagen-induced secretion, and after prolonged contact trigger secretion independent of other agonists. Here we report that LDL activates the small GTPases Rap1 and Ral but not Ras, as assessed by specific precipitation of the GTP-bound enzymes. In unstirred suspensions, the inhibitor SB203580 blocks Rap1 activation by 60-70%, suggesting activation via p38 mitogen activated protein kinase and a second, unidentified route. Inhibitors of cyclooxygenase (indomethacin) and the thromboxane A(2) (TxA(2)) receptor (SQ30741) induce complete inhibition, indicating that Rap1 activation is the result of TxA(2) formation. Stirring reveals a second, TxA(2)-independent Rap1 activation, which correlates quantitatively with a slow induction of dense granule secretion. Both pathways are unaffected by inhibitors of ligand binding to integrin alpha(IIb)beta(3). The results suggest that Rap1 and Ral, but not Ras, may take part in signalling routes initiated by LDL that initially enhance the sensitivity of platelets to other agonists and later trigger LDL-dependent secretion. PMID- 10861234 TI - Regulation of ecdysteroid signalling: molecular cloning, characterization and expression of 3-dehydroecdysone 3 alpha-reductase, a novel eukaryotic member of the short-chain dehydrogenases/reductases superfamily from the cotton leafworm, Spodoptera littoralis. AB - One route of inactivation of ecdysteroids in insects involves ecdysone oxidase catalysed conversion into 3-dehydroecdysone (3DE), followed by irreversible reduction by 3DE 3 alpha-reductase to 3-epiecdysone. The 3DE 3 alpha-reductase has been purified and subjected to limited amino acid sequencing. It occurs as two distinct forms, including a probable trimer of subunit molecular mass of approx. 26 kDa. A reverse-transcriptase PCR-based approach has been used to clone the cDNA (1.2 kb) encoding the 26 kDa protein. Northern blotting showed that the mRNA transcript was expressed in Malpighian tubules during the early stage of the last larval instar. Conceptual translation of the 3DE 3 alpha-reductase cDNA and database searching revealed that the enzyme belongs to the short-chain dehydrogenases/reductases superfamily. Furthermore, the enzyme is a novel eukaryotic 3-dehydrosteroid 3 alpha-reductase member of that family, whereas vertebrate 3-dehydrosteroid 3 alpha-reductases belong to the aldo-keto reductase (AKR) superfamily. Enzymically active recombinant 3DE 3 alpha-reductase has been produced using a baculovirus expression system. Surprisingly, we observed no similarity between this 3DE 3 alpha-reductase and a previously reported 3DE 3 beta-reductase, which acts on the same substrate and belongs to the AKR family. PMID- 10861235 TI - Definition of Munc13-homology-domains and characterization of a novel ubiquitously expressed Munc13 isoform. AB - Munc13 proteins constitute a family of three highly homologous molecules (Munc13 1, Munc13-2 and Munc13-3). With the exception of a ubiquitously expressed Munc13 2 splice variant, Munc13 proteins are brain-specific. Munc13-1 has a central priming function in synaptic vesicle exocytosis from glutamatergic synapses. In order to identify Munc13-like proteins that may regulate secretory processes in non-glutamatergic neurons or non-neuronal cells, we developed protein profiles for two Munc13-homology-domains (MHDs). MHDs are present in a wide variety of proteins, some of which have previously been implicated in membrane trafficking reactions. Taking advantage of partial sequences in the human expressed sequence tag (EST) database, we characterized a novel, ubiquitously expressed, rat protein (Munc13-4) that belongs to a subfamily of Munc13-like molecules, in which the typical Munc13-like domain structure is conserved. Munc13-4 is predominantly expressed in lung where it is localized to goblet cells of the bronchial epithelium and to alveolar type II cells, both of which are cell types with secretory function. In the present study we identify a group of novel proteins, some of which may function in a Munc13-like manner to regulate membrane trafficking. The MHD profiles described in the present study are useful tools for the identification of Munc13-like proteins, that would otherwise have remained undetected. PMID- 10861236 TI - FM1-43 reports plasma membrane phospholipid scrambling in T-lymphocytes. AB - We have found using imaging techniques that stimulating Jurkat human leukaemic T cells with ionomycin in the presence of FM1-43, a dye used to monitor exocytosis and endocytosis, causes large (6--10-fold) increases in FM1-43 fluorescence. These responses are too large to be caused by exocytosis. Instead, three lines of evidence suggest that FM1-43 is responding to phospholipid scrambling. First, ionomycin also stimulates increases in the fluorescence of annexin V, a phosphatidylserine-specific probe, while thapsigargin does not stimulate fluorescence increases of either probe. Secondly, cells that exhibit FM1-43 fluorescence increases after ionomycin stimulation stain with annexin V once FM1 43 is washed out. Thirdly, ionomycin stimulates uptake of 7-nitrobenz-2-oxa-1,3 diazole-labelled phosphatidylcholine, a specific assay for scramblase activity, whereas thapsigargin does not. We find that FM1-43 reports phospholipid scrambling with 'better' kinetics than annexin V, and does require extracellular Ca(2+) to report phospholipid scrambling. We suggest that FM1-43 may be a useful probe to study the dynamics of phospholipid scrambling. The results are the first demonstration that FM1-43 can respond significantly to a biological process other than vesicular trafficking. PMID- 10861237 TI - A single amino acid substitution (Trp(666)-->Ala) in the interbox1/2 region of the interleukin-6 signal transducer gp130 abrogates binding of JAK1, and dominantly impairs signal transduction. AB - gp130 is the common signal-transducing receptor chain of interleukin (IL)-6-type cytokines. Here we describe, for the first time, a single amino acid substitution (Trp(666)-->Ala) in the membrane-proximal interbox1/2 region that abrogates activation of STAT (signal transducer and activator of transcription) transcription factors and the proliferative response of pro-B-cell transfectants. Moreover, association of the Janus kinase JAK1 is prevented. No signalling of heterodimeric IL-5 receptor (IL-5R)/gp130 chimaeras occurs in COS-7 cells, even when only a single cytoplasmic chain of a gp130 dimer contains the Trp(666)Ala mutation, indicating that it acts dominantly. PMID- 10861238 TI - Non-linear antigenic regions in epidermal growth factor (EGF) and transforming growth factor alpha (TGF alpha) studied by EGF-TGF alpha chimaeras. AB - With the help of 16 chimaeras between human epidermal growth factor (hEGF) and human transforming growth factor alpha (hTGF alpha), a detailed analysis was performed on the epitope recognized by two polyclonal antibodies raised against hEGF, and one polyclonal antibody raised against hTGF alpha. All three antibodies recognized essentially the same antigenic site, a non-linear and conformation dependent sequence that is located near the second and fourth disulphide-bonded cysteines and that includes the start of the B-loop beta-sheet. The epitope recognized by the anti-hEGF antibodies was further characterized using 8 chimaeras between hEGF and an EGF-repeat from Drosophila Notch and was found to include Met(21), Ala(30) and Asn(32). All three polyclonal antibodies were able to neutralize the biological activity of the respective growth factor when tested on 32D murine haematopoietic progenitor cells transfected with ErbB-1, indicating that the receptor binding domain is shielded upon binding of the antibody. PMID- 10861239 TI - Kinetic properties of missense mutations in patients with glutathione synthetase deficiency. AB - Patients with hereditary glutathione synthetase (GS) (EC 6.3.2.3) deficiency present with variable clinical pictures, presumably related to the nature of the mutations involved. In order to elucidate the relationship between genotype, enzyme function and clinical phenotype, we have characterized enzyme kinetic parameters of missense mutations R125C, R267W, R330C and G464V from patients with GS deficiency. One of the mutations predominantly affected the K(m) value, with decreased affinity for glycine, two mutations influenced both K(m) and V(max) values, and one mutation reduced the stability of the enzyme. This characterization agrees well with predictions based on the recently reported crystal structure of human GS. Thus our data indicate that different mutations can affect the catalytic capacity of GS by decreasing substrate affinity, maximal velocity or enzyme stability. PMID- 10861240 TI - Targeting motifs and functional parameters governing the assembly of connexins into gap junctions. AB - To study the assembly of gap junctions, connexin--green-fluorescent-protein (Cx- GFP) chimeras were expressed in COS-7 and HeLa cells. Cx26-- and Cx32--GFP were targeted to gap junctions where they formed functional channels that transferred Lucifer Yellow. A series of Cx32--GFP chimeras, truncated from the C-terminal cytoplasmic tail, were studied to identify amino acid sequences governing targeting from intracellular assembly sites to the gap junction. Extensive truncation of Cx32 resulted in failure to integrate into membranes. Truncation of Cx32 to residue 207, corresponding to removal of most of the 78 amino acids on the cytoplasmic C-terminal tail, led to arrest in the endoplasmic reticulum and incomplete oligomerization. However, truncation to amino acid 219 did not impair Cx oligomerization and connexon hemichannels were targeted to the plasma membrane. It was concluded that a crucial gap-junction targeting sequence resides between amino acid residues 207 and 219 on the cytoplasmic C-terminal tail of Cx32. Studies of a Cx32E208K mutation identified this as one of the key amino acids dictating targeting to the gap junction, although oligomerization of this site-specific mutation into hexameric hemichannels was relatively unimpaired. The studies show that expression of these Cx--GFP constructs in mammalian cells allowed an analysis of amino acid residues involved in gap-junction assembly. PMID- 10861241 TI - Heterologous expression, functional characterization and localization of two isoforms of the monkey iron transporter Nramp2. AB - Natural resistance-associated macrophage protein 2 (Nramp2) has been suggested to be involved in transferrin-independent iron uptake. Two isoforms of the Nramp2 gene generated by alternative splicing of the 3' exons were identified in mouse, rat and human, but it is unclear if they perform distinct functions. To rationalize our previous work, which indicated an increase in iron deposition in a Parkinsonian monkey brain, two monkey Nramp2 isoforms were isolated for a comparative study to assess their relative iron-uptake abilities, tissue distribution and subcellular localization. The monkey Nramp2 isoforms, 2a and 2b, exhibit approx. 98% identity at the amino acid level when compared with the human homologues. The Nramp2a transcript contains a canonical iron-responsive element (IRE), whereas that of Nramp2b lacks the IRE motif in the 3' untranslated region. By reverse transcriptase (RT)-PCR, the mRNAs of both isoforms were detected in all tissues examined. The amino acid differences at the C-terminus neither affected the protein expression levels in HEK-293T and COS-7 cells nor altered the subcellular localization and tissue distribution of the isoforms. Similar levels of iron uptake were detected in the HEK-293T cells transfected with either the Nramp2a or 2b gene, and a reduction of iron from the ferric (Fe(3+)) to the ferrous (Fe(2+)) state is necessary before transport can take place. However, this transferrin-independent uptake of iron into the cells is not a Ca(2+) dependent process. PMID- 10861242 TI - Oligomerization of beta-amyloid of the Alzheimer's and the Dutch-cerebral haemorrhage types. AB - A novel ELISA has been developed which detects oligomerization of beta-amyloid (A beta). Oligomerization, fibrillization and neurotoxicity of native A beta associated with Alzheimer's disease (AD) type has been compared with E22Q A beta (amyloid beta-protein containing residues 1--40 with the native Glu at residue 22 changed to Gln) implicated in Dutch cerebral haemorrhage disease. Solutions of A beta rapidly yield soluble oligomers in a concentration-dependent manner, which are detected by the ELISA, and by size-exclusion gel chromatography. Conformational changes from disordered to beta-sheet occur more slowly than oligomerization, and fibrils are produced after prolonged incubation. The E22Q A beta oligomerizes, changes conformation and fibrillizes more rapidly than the native form and produces shorter stubbier fibrils. Aged fibrillar preparations of E22Q A beta are more potent than aged fibrils of native A beta in inducing apoptotic changes and toxic responses in human neuroblastoma cell lines, whereas low-molecular-mass oligomers in briefly incubated solutions are much less potent. The differences in the rates of oligomerization of the two A beta forms, their conformational behaviour over a range of pH values, and NMR data reported elsewhere, are consistent with a molecular model of oligomerization in which strands of A beta monomers initially overcome charge repulsion to form dimers in parallel beta-sheet arrangement, stabilized by intramolecular hydrophobic interactions, with amino acids of adjacent chains in register. PMID- 10861243 TI - Oxalate oxidases and differentiating surface structure in wheat: germins. AB - Oxalate oxidases (OXOs) have been found to be concentrated in the surface tissues of wheat embryos and grains: germin is concentrated in root and leaf sheaths that surround germinated embryos; pseudogermin (OXO-psi) is concentrated in the epidermis and bracts that 'encircle' mature grains. Most strikingly, the epidermal accumulation of OXO-psi was found to presage the transition of a delicate 'skin', similar to the fragile epidermis of human skin, into the tough shell (the miller's 'beeswing') that is typical of mature wheat grains. A narrow range of oxalate concentration (1--2 mM) in the hydrated tissues of major crop cereals (barley, maize, oat, rice, rye and wheat) contrasted with wide variations in their OXO expression, e.g. cold-tolerant and cold-sensitive varieties of maize have similar oxalate contents but the former was found to contain approx. 20-fold more germin than did the latter. Well-known OXOs in sorghum, a minor cereal, and beet, a dicotyledon, were found to have little antigenic relatedness to the germins, but the beet enzyme did share some of the unique stability properties that are peculiar to the germin-like OXOs that are found only in the major crop cereals. Their concentration in surface structures of domesticated wheat suggests a biochemical role for germin-like OXOs: programmed cell death in surface tissues might be a constitutive as well as an adaptive form of differentiation that helps to produce refractory barriers against tissue invasion by predators. Incidental to the principal investigation, and using an OXO assay (oxalate-dependent release of CO(2)) that did not rely on detecting H(2)O(2), which is often fully degraded in cell extracts, it was found that OXO activity in soluble extracts of wheat was manifested only in standard solution assays if the extract was pretreated in a variety of ways, which included preincubation with pepsin or highly substituted glucuronogalactoarabinoxylans (cell-wall polysaccharides). PMID- 10861244 TI - Ca(2+) signals mediated by Ins(1,4,5)P(3)-gated channels in rat ureteric myocytes. AB - Localized Ca(2+)-release signals (puffs) and propagated Ca(2+) waves were characterized in rat ureteric myocytes by confocal microscopy. Ca(2+) puffs were evoked by photorelease of low concentrations of Ins(1,4,5)P(3) from a caged precursor and by low concentrations of acetylcholine; they were also observed spontaneously in Ca(2+)-overloaded myocytes. Ca(2+) puffs showed some variability in amplitude, time course and spatial spread, suggesting that Ins(1,4,5)P(3) gated channels exist in clusters containing variable numbers of channels and that within these clusters a variable number of channels can be recruited. Immunodetection of Ins(1,4,5)P(3) receptors revealed the existence of several spots of fluorescence in the confocal cell sections, supporting the existence of clusters of Ins(1,4,5)P(3) receptors. Strong Ins(1,4,5)P(3) photorelease and high concentrations of acetylcholine induced Ca(2+) waves that originated from an initiation site and propagated in the whole cell by spatial recruitment of neighbouring Ca(2+)-release sites. Both Ca(2+) puffs and Ca(2+) waves were blocked selectively by intracellular applications of heparin and an anti Ins(1,4,5)P(3)-receptor antibody, but were unaffected by ryanodine and intracellular application of an anti-ryanodine receptor antibody. mRNAs encoding for the three subtypes of Ins(1,4,5)P(3) receptor and subtype 3 of ryanodine receptor were detected in these myocytes, and the maximal binding capacity of [(3)H]Ins(1,4,5)P(3) was 10- to 12-fold higher than that of [(3)H]ryanodine. These results suggest that Ins(1,4,5)P(3)-gated channels mediate a continuum of Ca(2+) signalling in smooth-muscle cells expressing a high level of Ins(1,4,5)P(3) receptors and no subtypes 1 and 2 of ryanodine receptors. PMID- 10861245 TI - Molecular modelling and site-directed mutagenesis of the inositol 1,3,4,5 tetrakisphosphate-binding pleckstrin homology domain from the Ras GTPase activating protein GAP1IP4BP. AB - GAP1(IP4BP) is a Ras GTPase-activating protein (GAP) that in vitro is regulated by the cytosolic second messenger inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P(4)]. We have studied Ins(1,3,4,5)P(4) binding to GAP1(IP4BP), and shown that the inositol phosphate specificity and binding affinity are similar to Ins(1,3,4,5)P(4) binding to Bruton's tyrosine kinase (Btk), evidence which suggests a similar mechanism for Ins(1,3,4,5)P(4) binding. The crystal structure of the Btk pleckstrin homology (PH) domain in complex with Ins(1,3,4,5)P(4) has shown that the binding site is located in a partially buried pocket between the beta 1/beta 2- and beta 3/beta 4-loops. Many of the residues involved in the binding are conserved in GAP1(IP4BP). Therefore we generated a model of the PH domain of GAP1(IP4BP) in complex with Ins(1,3,4,5)P(4) based on the Btk Ins(1,3,4,5)P(4) complex crystal structure. This model had the typical PH domain fold, with the proposed binding site modelling well on the Btk structure. The model has been verified by site-directed mutagenesis of various residues in and around the proposed binding site. These mutations have markedly reduced affinity for Ins(1,3,4,5)P(4), indicating a specific and tight fit for the substrate. The model can also be used to explain the specificity of inositol phosphate binding. PMID- 10861246 TI - Identification and expression of NEU3, a novel human sialidase associated to the plasma membrane. AB - Several mammalian sialidases have been described so far, suggesting the existence of numerous polypeptides with different tissue distributions, subcellular localizations and substrate specificities. Among these enzymes, plasma-membrane associated sialidase(s) have a pivotal role in modulating the ganglioside content of the lipid bilayer, suggesting their involvement in the complex mechanisms governing cell-surface biological functions. Here we describe the identification and expression of a human plasma-membrane-associated sialidase, NEU3, isolated starting from an expressed sequence tag (EST) clone. The cDNA for this sialidase encodes a 428-residue protein containing a putative transmembrane helix, a YRIP (single-letter amino acid codes) motif and three Asp boxes characteristic of sialidases. The polypeptide shows high sequence identity (78%) with the membrane associated sialidase recently purified and cloned from Bos taurus. Northern blot analysis showed a wide pattern of expression of the gene, in both adult and fetal human tissues. Transient expression in COS7 cells permitted the detection of a sialidase activity with high activity towards ganglioside substrates at a pH optimum of 3.8. Immunofluorescence staining of the transfected COS7 cells demonstrated the protein's localization in the plasma membrane. PMID- 10861247 TI - Impaired nitric oxide production in coronary endothelial cells of the spontaneously diabetic BB rat is due to tetrahydrobiopterin deficiency. AB - Endothelial cells (EC) from diabetic BioBreeding (BB) rats have an impaired ability to produce NO. This deficiency is not due to a defect in the constitutive isoform of NO synthase in EC (ecNOS) or alterations in intracellular calcium, calmodulin, NADPH or arginine levels. Instead, ecNOS cannot produce sufficient NO because of a deficiency in tetrahydrobiopterin (BH(4)), a cofactor necessary for enzyme activity. EC from diabetic rats exhibited only 12% of the BH(4) levels found in EC from normal animals or diabetes-prone animals which did not develop disease. As a result, NO synthesis by EC of diabetic rats was only 18% of that for normal animals. Increasing BH(4) levels with sepiapterin increased NO production, suggesting that BH(4) deficiency is a metabolic basis for impaired endothelial NO synthesis in diabetic BB rats. This deficiency is due to decreased activity of GTP-cyclohydrolase I, the first and rate-limiting enzyme in the de novo biosynthesis of BH(4). GTP-cyclohydrolase activity was low because of a decreased expression of the protein in the diabetic cells. PMID- 10861248 TI - Inhibition of Ca(2+) signalling by p130, a phospholipase-C-related catalytically inactive protein: critical role of the p130 pleckstrin homology domain. AB - p130 was originally identified as an Ins(1,4,5)P(3)-binding protein similar to phospholipase C-delta but lacking any phospholipase activity. In the present study we have further analysed the interactions of p130 with inositol compounds in vitro. To determine which of the potential ligands interacts with p130 in cells, we performed an analysis of the cellular localization of this protein, the isolation of a protein-ligand complex from cell lysates and studied the effects of p130 on Ins(1,4,5)P(3)-mediated Ca(2+) signalling by using permeabilized and transiently or stably transfected COS-1 cells (COS-1(p130)). In vitro, p130 bound Ins(1,4,5)P(3) with a higher affinity than that for phosphoinositides. When the protein was isolated from COS-1(p130) cells by immunoprecipitation, it was found to be associated with Ins(1,4,5)P(3). Localization studies demonstrated the presence of the full-length p130 in the cytoplasm of living cells, not at the plasma membrane. In cell-based assays, p130 had an inhibitory effect on Ca(2+) signalling. When fura-2-loaded COS-1(p130) cells were stimulated with bradykinin, epidermal growth factor or ATP, it was found that the agonist-induced increase in free Ca(2+) concentration, observed in control cells, was inhibited in COS 1(p130). This inhibition was not accompanied by the decreased production of Ins(1,4,5)P(3); the intact p130 pleckstrin homology domain, known to be the ligand-binding site in vitro, was required for this effect in cells. These results suggest that Ins(1,4,5)P(3) could be the main p130 ligand in cells and that this binding has the potential to inhibit Ins(1,4,5)P(3)-mediated Ca(2+) signalling. PMID- 10861249 TI - Deactivation of neutrophil NADPH oxidase by actin-depolymerizing agents in a cell free system. AB - The cell-free activation of human neutrophil NADPH oxidase (O(2)(-)-generating enzyme) is enhanced by actin [Morimatsu, Kawagoshi, Yoshida and Tamura (1997) Biochem. Biophys. Res. Commun. 230, 206--210]. In an attempt to elucidate the mechanism, we examined the effect of actin-depolymerizing agents on the duration of NADPH oxidase in a cell-free system. The addition of DNase I, an F-actin depolymerizing protein, caused an accelerated deactivation of the oxidase. The deactivation was also facilitated by latrunculin A, a sponge toxin that depolymerizes F-actin. Exogenously added actin prevented the deactivation by DNase I or latrunculin A, whereas EDTA accelerated a dilution-induced deactivation of the oxidase and Mg(2+) ions retarded it. The stability in dilution was found to correlate well with free Mg(2+) concentration. Estimation of F-actin in the system showed that F-actin increased during the oxidase activation and that DNase I or EDTA decreased F-actin content in parallel with the activity. Treatment of the cell-free mixture with a chemical cross-linker prevented the deactivation and F-actin decrease by EDTA. Taken together, these results suggest that actin filaments which grow during the activation of NADPH oxidase prolong the lifetime of the oxidase. PMID- 10861250 TI - Sj-FABPc fatty-acid-binding protein of the human blood fluke Schistosoma japonicum: structural and functional characterization and unusual solvent exposure of a portal-proximal tryptophan residue. AB - Sj-FABPc of the blood fluke of humans, Schistosoma japonicum, is a member of the FABP/P2/CRBP/CRABP family of beta-barrel cytosolic fatty-acid-binding and retinoid-binding proteins. Sj-FABPc has at least eight different variants encoded by a single-copy polymorphic gene. In fluorescence-based assays, recombinant Sj FABPc was found to bind 11-(dansylamino)undecanoic acid (DAUDA), inducing a shift in peak fluorescence emission from 543 to 493 nm. A similar spectral change was observed in dansyl-amino-octanoic acid (in which the dansyl fluorophore is attached at the alpha-carbon rather than the omega-carbon of DAUDA), indicating that the ligand enters entirely into the binding site. Sj-FABPc also bound the naturally fluorescent cis-parinaric acid, as well as oleic acid and arachidonic acid, by competition, but not all-trans-retinol. Dissociation constants were, for cis-parinaric acid, K(d)=2.5+/-0.1 microM (mean+/-S.E.M.) and an apparent stoichiometry consistent with one binding site per molecule of Sj-FABPc and, for oleic acid, K(i) approximately 80 nM. A deletion mutant from which alpha-II was absent failed to bind ligand. Sj-FABPc modelled well to known structures of the protein family; an unusually solvent-exposed Trp side chain was evident adjacent to the presumptive portal through which ligand is thought to enter and leave. Intrinsic fluorescence analyses of Sj-FABPc and of the deletion mutant (from which Trp-27 is absent) confirmed the unusual disposition of this side chain. Virtually all members of the FABP/P2/CRBP/CRABP protein family have prominent hydrophobic side chains in this position, with the exception of liver FABP and ileal FABP, which instead have charged side chains. Liver FABP is known to be distinct from other members of the protein family in that it does not seem to contact membranes to collect and deposit its ligand. It is therefore postulated that the unusually positioned apolar side chains in Sj-FABPc and others in the family are important in interactions with membranes or other cellular components. PMID- 10861251 TI - The molecular genetics of familial intrahepatic cholestasis. PMID- 10861252 TI - Inducible nitric oxide synthase: a little bit of good in all of us. AB - The established dogma regarding the different isoforms of nitric oxide has been that constitutively expressed nitric oxide synthase is an extremely important homeostatic regulator of numerous important physiological processes whereas the inducible form of nitric oxide synthase underlies injury associated with intestinal inflammation. In this brief overview, I review some of the literature that clearly supports this contention, particularly the dramatically beneficial effects of oral L-NAME administration to animals with colitis induced by trinitrobenzene sulphonic acid (TNBS). However, I also highlight some of the gastrointestinal data that does not fit this simple tidy paradigm, particularly with respect to the inducible form of nitric oxide synthase (iNOS). For example, iNOS induced healing of skin and the intestinal mucosa, killing of certain bacteria, regulation of T cell proliferation and differentiation (Th1 v Th2), and control of leucocyte recruitment may mask or counter the toxic metabolites that are produced by iNOS. Perhaps it is not surprising that one does not always obtain benefit from inhibiting all iNOS either by gene deletion or by systemic NOS inhibition. I raise some potential flaws in our approaches to studying iNOS. For example, to date no attempts have been made to selectively inhibit iNOS in single cell types. Global inhibition of all iNOS assumes that the large variety of cell types that can produce iNOS have identical functions. Finally, I attempt to highlight areas that require additional investigation and issues that have not been explored. PMID- 10861253 TI - H pylori and Lewis antigens. PMID- 10861254 TI - A little rest and relaxation. PMID- 10861255 TI - MMPs in the gut: inflammation hits the matrix. PMID- 10861256 TI - Heparin and inflammation: a new use for an old GAG? PMID- 10861257 TI - Antimicrobial peptides in innate intestinal host defence. PMID- 10861258 TI - Association of peptic ulcer with increased expression of Lewis antigens but not cagA, iceA, and vacA in Helicobacter pylori isolates in an Asian population. AB - BACKGROUND: Studies in Western populations suggest that cagA, iceA, and vacA gene status in Helicobacter pylori isolates is associated with increased virulence and peptic ulcer disease. AIM: To investigate the relationship between peptic ulcer and expression of Lewis (Le) antigens as well as cagA, iceA, and vacA in H pylori isolates in Singapore. METHODS: Expression of Le antigens in H pylori isolates obtained from patients with dyspepsia was measured by enzyme linked immunosorbent assay. The cagA, iceA, and vacA status was determined by polymerase chain reaction. RESULTS: Of 108 H pylori isolates, 103 (95.4%) expressed Le(x) and/or Le(y), while Le(a) and Le(b) were expressed in 23 (21.3%) and 47 (43.5%) isolates, respectively. Expression of two or more Le antigens (Le(x), Le(y), Le(a), or Le(b)) was significantly higher in H pylori isolated from ulcer patients than in non-ulcer patients (89.6% v 73.2%, p=0.035). There were no significant differences in the prevalence of cagA or iceA1 in H pylori isolates from peptic ulcer and non-ulcer patients (86.6% v 90.2% for cagA; 70.1% v 68.3% for iceA1), and no association of peptic ulcer with any specific vacA genotype. CONCLUSIONS: The present study indicates that peptic ulcer disease is associated with increased expression of Lewis antigens but not cagA, iceA, or vacA genotype in H pylori isolates in our population. This suggests that cagA, iceA, and vacA are not universal virulence markers, and that host-pathogen interactions are important in determining clinical outcome. PMID- 10861259 TI - High prevalence of potentially virulent strains of Helicobacter pylori in the general male British population. AB - BACKGROUND: Strains of Helicobacter pylori that express the cytotoxin associated gene product A (CagA) may be more strongly associated with serious gastric diseases, such as gastric cancer and peptic ulceration, than other strains. Data, however, are sparse on the prevalence, risk factors, and other correlates of these strains in the general population. AIM: To characterise aspects of the seroepidemiology of CagA(+) strains of H pylori in the general British population. METHODS: We measured serum IgG antibodies to mixed H pylori antigens and separately to CagA in 1025 men aged 40-59 years who were randomly selected from a larger group of participants in a community based survey conducted in 18 different British towns. RESULTS: Overall, 44% (95% confidence interval 41-47%) of the men were seropositive to CagA antibodies, representing about 61% (57-65%) of the men seropositive to mixed antigen H pylori. The risk factors for seropositivity to CagA antibodies were similar to those for seropositivity to mixed antigen H pylori, apart from an increased prevalence of reported bedroom sharing in childhood (p<0.01). CONCLUSION: In a nationwide study of potentially virulent H pylori strains, there was a high prevalence of the infection, with some evidence that acquisition of such strains might occur earlier in life than other strains. PMID- 10861260 TI - No relation between body mass and gastro-oesophageal reflux symptoms in a Swedish population based study. AB - BACKGROUND: There is a widespread notion that obesity leads to gastro-oesophageal reflux but scientific evidence of an association is limited and inconsistent. AIMS: To estimate the strength of the association between body mass and reflux symptoms, we performed a population based cross sectional interview study. SUBJECTS: Population based, randomly selected, middle aged or elderly persons in Sweden in 1995-1997. METHODS: At face-to-face interviews we asked a stratified sample of Swedes about body measures and occurrence of reflux symptoms. Odds ratios (OR) with 95% confidence intervals (CI), calculated by logistic regression with multivariate adjustments for covariates, were the measures of association. RESULTS: Reflux symptoms occurring at least once a week more than five years before the interview were reported by 135 (16%) of the 820 interviewees. Among those who had ever been overweight during adulthood (body mass index (BMI) > or =25 kg/m(2)), the OR of having recurrent reflux symptoms was 0.99 (95% CI 0.66 1.47) compared with those who were never overweight. There was no association between BMI at age 20, BMI 20 years before the interview, or maximum adult BMI and occurrence of reflux symptoms: ORs per unit increase in BMI were 1. 00 (95% CI 0.93-1.09), 1.03 (95% CI 0.96-1.10), and 1.01 (95% CI=0.95-1.07), respectively. There was no association between BMI and severity or duration of reflux symptoms. CONCLUSIONS: Gastro-oesophageal reflux symptoms occur independently of body mass index. Weight reduction may not be justifiable as an antireflux therapy. PMID- 10861261 TI - Effect of atropine on proximal gastric motor and sensory function in normal subjects. AB - BACKGROUND AND AIMS: Distension of the proximal stomach is a major stimulus for triggering transient lower oesophageal sphincter (LOS) relaxations. We have shown recently that atropine inhibits triggering of transient LOS relaxations in both normal subjects and patients with gastro-oesophageal reflux disease. Atropine could potentially act centrally by inhibiting the central integrating mechanism in the brain stem, or act peripherally by altering the response of the stomach to distension. The aim of this study was to investigate the effect of atropine on fasting gastric compliance and postprandial gastric tone using an electronic barostat. METHODS: Fasting and postprandial proximal gastric motor and sensory functions were assessed in 10 normal healthy volunteers. Oesophageal manometry and pH were simultaneously measured. On separate days, atropine (15 microg/kg bolus, 4 microg/kg/h intravenous infusion) or saline was given and maintained for the duration of the recording period. RESULTS: In the fasting period, atropine significantly reduced minimum distending pressure (5.5 (0.4) v 4.4 (0.4) mm Hg; p<0.005) and increased proximal gastric compliance (81.3 (5.3) v 102. 1 (8.7) ml/ mm Hg; p<0.05). In response to a meal, maximal gastric relaxation was similar on both study days. However, during atropine infusion, there was no recovery of proximal gastric tone in the two hour postprandial observation period. Postprandial fullness scores were higher during atropine infusion and correlated with changes in intrabag volume. Atropine significantly reduced the rate of postprandial transient LOS relaxations: first hour, 7.0 (5.3-10.0) v 3.0 (1.0 4.0) (p<0.02); second hour, 5.0 (3.3-5.8) per hour v 1.0 (0-3.0) per hour (p<0.05). CONCLUSIONS: In humans, fasting and postprandial proximal gastric motor function is under cholinergic control. Atropine induced inhibition of transient LOS relaxations is unlikely to be caused by its effect on the proximal stomach, but rather by a central action on the integrating mechanisms in the brain stem. PMID- 10861262 TI - DNA microsatellite instability and mismatch repair protein loss in adenomas presenting in hereditary non-polyposis colorectal cancer. AB - BACKGROUND AND AIM: Hereditary non-polyposis colorectal cancer (HNPCC), as its name implies, is associated with few adenomas, and the early evolution of colorectal neoplasia is poorly understood. In this study our aim was to clarify the genetic profiles of benign polyps in subjects with HNPCC using a combined molecular and immunohistochemical approach. METHODS: Thirty adenomas and 17 hyperplastic polyps were obtained from 24 affected HNPCC subjects. DNA was extracted from paraffin embedded tissue by microdissection and analysed for the presence of microsatellite instability (MSI) and mutations in five genes known to be targets in mismatch repair deficiency (TGFbetaRII, IGF2R, BAX, hMSH3, and hMSH6). Serial sections were stained by immunohistochemistry for hMLH1 and hMSH2. RESULTS: Twenty four (80%) of 30 adenomas showed MSI. Of MSI positive adenomas, 66.7% showed MSI at more than 40% of markers (high level of MSI (MSI-H)). Two of 17 hyperplastic polyps revealed MSI at one marker (low level of MSI (MSI-L)). A significant association was found between MSI-H and high grade dysplasia in adenomas (p=0.004). Eight of nine adenomas with mutations of coding sequences revealed high grade dysplasia and all nine were MSI-H. Four of the nine ranged in size from 2 to 5 mm. The presence of the hMSH6 mutation was significantly correlated with high levels of MSI (80% of markers) (p<0.02). Twenty four adenomas gave evaluable results with immunohistochemistry. One of six (17%) microsatellite stable, six of seven (86%) MSI-L, and 11 of 11 (100%) MSI-H adenomas showed loss of either hMLH1 or hMSH2. CONCLUSIONS: Most adenomas in subjects with a definite diagnosis of HNPCC show MSI (80%). The finding of MSI-L is usually associated with loss of expression of hMLH1 or hMSH2, unlike the situation in MSI-L sporadic colorectal cancer. The transition from MSI-L to MSI-H correlated with the finding of high grade dysplasia and mutation of coding sequences and may be driven by mutation of secondary mutators such as hMSH3 and hMSH6. Advanced genetic changes may be present in adenomas of minute size. PMID- 10861263 TI - Neoplastic progression occurs through mutator pathways in hyperplastic polyposis of the colorectum. AB - AIM: Colorectal cancer has been described in association with hyperplastic polyposis but the mechanism underlying this observation is unknown. The aim of this study was to characterise foci of dysplasia developing in the polyps of subjects with hyperplastic polyposis on the basis of DNA microsatellite status and expression of the DNA mismatch repair proteins hMLH1, hMSH2, and hMSH6. MATERIALS AND METHODS: The material was derived from four patients with hyperplastic polyposis and between one and six synchronous colorectal cancers. Normal (four), hyperplastic (13), dysplastic (13), and malignant (11) samples were microdissected and a PCR based approach was used to identify mutations at 10 microsatellite loci, TGFbetaIIR, IGF2R, BAX, MSH3, and MSH6. Microsatellite instability-high (MSI-H) was diagnosed when 40% or more of the microsatellite loci showed mutational bandshifts. Serial sections were stained for hMLH1, hMSH2, and hMSH6. RESULTS: DNA microsatellite instability was found in 1/13 (8%) hyperplastic samples, in 7/13 (54%) dysplastic foci, and in 8/11 (73%) cancers. None of the MSI-low (MSI-L) samples (one hyperplastic, three dysplastic, two cancers) showed loss of hMLH1 expression. All four MSI-H dysplastic foci and six MSI-H cancers showed loss of hMLH1 expression. Loss of hMLH1 in MSI-H but not in MSI-L lesions showing dysplasia or cancer was significant (p<0.001, Fisher's exact test). Loss of hMSH6 occurred in one MSI-H cancer and one MSS focus of dysplasia which also showed loss of hMLH1 staining. CONCLUSION: Neoplastic changes in hyperplastic polyposis may occur within a hyperplastic polyp. Neoplasia may be driven by DNA instability that is present to a low (MSI-L) or high (MSI-H) degree. MSI-H but not MSI-L dysplastic foci are associated with loss of hMLH1 expression. At least two mutator pathways drive neoplasia in hyperplastic polyposis. The role of the hyperplastic polyp in the histogenesis of sporadic DNA microsatellite unstable colorectal cancer should be examined. PMID- 10861264 TI - Increased expression of collagenase-3 (MMP-13) and MT1-MMP in oesophageal cancer is related to cancer aggressiveness. AB - BACKGROUND: Collagenase-3 (matrix metalloproteinase-13, MMP-13) is a recently identified human MMP with broad substrate specificity which can be activated by membrane type 1 (MT1) matrix metalloproteinase in vitro. These may play a critical role in cancer aggressiveness. AIMS: To examine the clinical significance of collagenase-3 expression and the cooperative role of MT1-MMP in human oesophageal carcinomas. PATIENTS: Forty five individuals with oesophageal carcinoma who underwent surgery without preoperative treatment. METHODS: The tumour/normal (T/N) ratios of collagenase-3 and MT1-MMP mRNA expression in 45 human oesophageal carcinomas were determined by northern blot analysis. The production and localisation of collagenase-3 and MT1-MMP proteins were investigated by immunohistochemistry, western blot analysis, and zymography. RESULTS: The mean T/N ratio of collagenase-3 mRNA was 3.5 and that of MT1-MMP 2.1. There was a significant correlation between collagenase-3 and MT1-MMP mRNA expression (p<0.001). Twenty two cases with a collagenase-3 T/N ratio >3.5 showed a significantly higher frequency of vascular involvement and lymph node metastasis, and tended to be at a more advanced stage than 23 cases with a T/N ratio < or =3.5 (p<0.05). Western blot analysis and zymography demonstrated production of collagenase-3 protein in tumour tissues but not in normal tissues. Immunohistochemical studies revealed that collagenase-3 was localised predominantly in tumour cells and MT1-MMP was detected in the same collagenase-3 positive cells; there was a significant association between collagenase-3 and MT1 MMP protein expression (p<0.05). With regard to prognosis, the survival time for subjects in the high collagenase-3 group (T/N ratio >3.5) was significantly worse (p<0.05). CONCLUSIONS: These data suggest that production of collagenase-3 together with MT1-MMP is implicated in tumour aggressiveness and prognosis in human oesophageal carcinomas. PMID- 10861265 TI - Imbalance of stromelysin-1 and TIMP-1 in the mucosal lesions of children with inflammatory bowel disease. AB - BACKGROUND: Degradation of the extracellular matrix and ulceration of the mucosa are major features of inflammatory bowel disease (IBD). One of the most important enzymes in degrading the matrix and produced in excess by cytokine activated stromal cells, is stromelysin-1. The activity of stromelysin-1 is controlled by tissue inhibitor of metalloproteinase (TIMP-1), its natural inhibitor. In model systems excess stromelysin-1 produces mucosal degradation. METHODS: Quantitative competitive RT-PCR was used to analyse stromelysin-1 and TIMP-1 transcripts; western blotting was used to measure the amount of stromelysin-1 and TIMP-1 protein in biopsy samples from children with IBD. RESULTS: In biopsies from patients with active Crohn's disease (n=24), ulcerative colitis (n=23), and controls (n=16), TIMP-1 transcripts and protein were abundant and unchanged. Stromelysin-1 transcripts and protein were markedly elevated in mucosal biopsies obtained from inflamed sites of patients with active IBD but were not elevated in adjacent endoscopically normal mucosa (n=10). Elevated levels of stromelysin-1 transcripts in active Crohn's disease (n=5) returned to normal levels following treatment with enteral nutrition. CONCLUSIONS: Stromelysin-1 is markedly overexpressed at inflamed sites in patients with IBD whereas TIMP-1 remains unaltered. Excess stromelysin-1 is likely to be responsible for loss of mucosal integrity in IBD. PMID- 10861266 TI - Differential expression of matrix metalloproteinases and their tissue inhibitors in colon mucosa of patients with inflammatory bowel disease. AB - BACKGROUND/AIMS: Alterations in synthesis and breakdown of extracellular matrix components are known to play a crucial role in tissue remodelling during inflammation and wound healing. Degradation of collagens is highly regulated by a cascade of matrix metalloproteinases (MMPs). The current study was therefore designed to determine gene expression patterns of MMPs and their tissue inhibitors (TIMPs) in single endoscopic biopsies of patients with inflammatory bowel disease (IBD). PATIENTS/METHODS: mRNA expression was measured by quantitative competitive polymerase chain reaction (PCR) in biopsies from patients with ulcerative colitis (n=21) and Crohn's disease (n=21). Protein expression was analysed by western blotting and immunohistochemistry. RESULTS: MMP-2, MMP-14, and TIMP-1 mRNAs were marginally increased in inflamed, but 9-12 fold increased in ulcerated colonic mucosa in IBD whereas TIMP-2 mRNA expression remained unchanged. MMP-1 and MMP-3 mRNA expression correlated well with the histological degree of acute inflammation, resulting in more than 15-fold increased MMP-1 and MMP-3 mRNA levels in inflamed versus normal colon samples from patients with ulcerative colitis and Crohn's disease. CONCLUSION: Profound overexpression of MMP-1 and MMP-3 mRNA transcripts suggests an important role for these enzymes in the process of tissue remodelling and destruction in inflammatory bowel disease. PMID- 10861267 TI - Antineutrophil antibodies associated with ulcerative colitis interact with the antigen(s) during the process of apoptosis. AB - BACKGROUND: Cell death by apoptosis seems to be an important mechanism for translocation to the cell surface of a variety of intracellular components capable of inducing autoantibody production. AIMS: To identify the cellular location of antigen (Ag)-antineutrophil cytoplasmic antibodies (ANCA) in non apoptotic human neutrophils, and to assess if ANCA associated with ulcerative colitis reacts with neutrophil antigen(s) during neutrophil apoptosis. The cellular distribution of Ag-ANCA in apoptotic neutrophils was also investigated. METHODS: Sera from 18 ulcerative colitis patients known to be positive for perinuclear IgG-ANCA (titre > or =1/320), as assessed by indirect immunofluorescence (IIF), were analysed by immunofluorescent confocal laser scanning microscopy. ANCA were identified with fluorescein isothiocyanate (FITC) and tetramethylrhodamine isothiocyanate (TRITC) in non-apoptotic and apoptotic neutrophils, respectively. Apoptotic and non-apoptotic DNA was labelled with FITC and propidium iodide, respectively. Cycloheximide was added to polymorphonuclear leucocyte culture to induce apoptosis. RESULTS: Three patterns of scanning laser immunofluorescence microscopy in non-apoptotic neutrophils were observed with respect to cellular ulcerative colitis associated ANCA distribution: (1) diffuse nuclear localisation (16.7%); (2) nuclear localisation in the nuclear periphery (50%); and (3) mixed nuclear and cytoplasmic localisation (33.4%). In all sera ANCA fluorescence colocalised almost completely with apoptotic DNA, with persistence of a diffuse and intense fluorescence. No significant changes in ANCA titres were found in non-apoptotic neutrophils. CONCLUSIONS: The antigen(s) of ANCA associated with ulcerative colitis seems to be localised in most cases in the neutrophil nucleus. The almost identical colocalisation of ANCA and apoptotic cleaved DNA suggests that intracellular DNA redistribution during neutrophil apoptosis may play a role in antigen exposure to the immune system and ANCA production in ulcerative colitis. PMID- 10861268 TI - Non-pathogenic bacteria elicit a differential cytokine response by intestinal epithelial cell/leucocyte co-cultures. AB - BACKGROUND AND AIM: Intestinal epithelial cells (IEC) are thought to participate in the mucosal defence against bacteria and in the regulation of mucosal tissue homeostasis. Reactivity of IEC to bacterial signals may depend on interactions with immunocompetent cells. To address the question of whether non-pathogenic bacteria modify the immune response of the intestinal epithelium, we co cultivated enterocyte-like CaCO-2 cells with human blood leucocytes in separate compartments of transwell cultures. METHODS: CaCO-2/PBMC co-cultures were stimulated with non-pathogenic bacteria and enteropathogenic Escherichia coli. Expression of tumour necrosis factor alpha (TNF-alpha), interleukin (IL)-1beta, IL-8, monocyte chemoattracting protein 1 (MCP-1), and IL-10 was studied by enzyme linked immunosorbent assays (cytokine secretion) and by semiquantitative reverse transcription-polymerase chain reaction. RESULTS: Challenge of CaCO-2 cells with non-pathogenic E coli and Lactobacillus sakei induced expression of IL-8, MCP-1, IL-1beta, and TNF-alpha mRNA in the presence of underlying leucocytes. Leucocyte sensitised CaCO-2 cells produced TNF-alpha and IL-1beta whereas IL-10 was exclusively secreted by human peripheral blood mononuclear cells. CaCO-2 cells alone remained hyporesponsive to the bacterial challenge. Lactobacillus johnsonii, an intestinal isolate, showed reduced potential to induce proinflammatory cytokines but increased transforming growth factor beta mRNA in leucocyte sensitised CaCO-2 cells. TNF-alpha was identified as one of the early mediators involved in cellular cross talk. In the presence of leucocytes, discriminative activation of CaCO-2 cells was observed between enteropathogenic E coli and non-pathogenic bacteria. CONCLUSION: The differential recognition of non pathogenic bacteria by CaCO-2 cells required the presence of underlying leucocytes. These results strengthen the hypothesis that bacterial signalling at the mucosal surface is dependent on a network of cellular interactions. PMID- 10861269 TI - Heparin attenuates TNF-alpha induced inflammatory response through a CD11b dependent mechanism. AB - BACKGROUND: In addition to its anticoagulant properties, heparin has anti inflammatory effects, the molecular and mechanistic bases of which are incompletely defined. AIMS: The current studies were designed to test the hypothesis that heparin abrogates the expression or function of leucocyte endothelial adherence molecules which are fundamental to the acute inflammatory response. METHODS: The effects of heparin on tumour necrosis factor alpha (TNF alpha) induced leucocyte rolling, adhesion, and migration as well as vascular permeability were assessed in rat mesenteric venules using intravital microscopy. Expression of adhesion molecules was quantitated using a double radiolabelled monoclonal antibody (mAb) binding technique in vivo (P-selectin, intercellular cell adhesion molecule type 1 (ICAM-1), and vascular cell adhesion molecule 1 (VCAM-1)) or flow cytometry (CD11a, CD11b, and L-selectin). Ex vivo binding of heparin to neutrophils was assessed by flow cytometry. RESULTS: TNF-alpha induced a significant increase in leucocyte rolling, adhesion, and migration, and vascular permeability, coincident with a significant increase in expression of P selectin, ICAM-1, and VCAM-1. Ex vivo assessment of blood neutrophils showed significant upregulation of CD11a and CD11b and significant downregulation of L selectin within five hours of TNF-alpha administration. Heparin pretreatment significantly attenuated leucocyte rolling, adhesion, and migration but did not affect expression of cell adhesion molecules or vascular permeability elicited by TNF-alpha administration. Binding of heparin was significantly increased on blood neutrophils obtained five hours after TNF-alpha administration. Preincubation with an anti-CD11b mAb but not with an anti-CD11a or anti-L-selectin antibody significantly diminished heparin binding ex vivo. CONCLUSIONS: Our results support the concept that the anti-inflammatory effects of heparin involve attenuation of a CD11b dependent adherent mechanism. PMID- 10861270 TI - Blockade of the integrin alphaLbeta2 but not of integrins alpha4 and/or beta7 significantly prolongs intestinal allograft survival in mice. AB - BACKGROUND: Small bowel transplantation remains a difficult therapeutic option endangered by a high rate of rejection and infectious complications. To improve these clinical results, it is mandatory to set up animal models to test alternative immunosuppressive regimens which may lead to immunotolerance. AIMS: To determine the value of blockade of alphaLbeta2 (LFA-1) and alpha4 and beta7 integrins (alpha4beta1, alpha4beta7, and alphaEbeta7) in the prevention of rejection of fetal small bowel grafts in mice and the effect of the association of calcineurin dependent drugs in anti-LFA-1 treated mice. METHODS: Adult recipient mice engrafted with allogeneic fetal small bowel received a short course of anti-alpha4 and/or anti-LFA-1 monoclonal antibodies (mAb) with or without FK506 or cyclosporin A. In addition, in a set of experiment, beta7-/- mice were used as recipients. Graft biopsies were performed and processed for standard histology. RESULTS: Blockade of the pathways of the integrins alpha4 and beta7 had a modest or no effect on intestinal graft survival. In contrast, transitory, short administration of anti-LFA-1 monoclonal antibody alone, when started before engraftment (day -1), allowed long term survival of intestinal grafts, even when associated with calcineurin dependent drugs. However, early withdrawal of FK506 reversed the immunosuppressive effect of anti-LFA-1 treatment. CONCLUSION: These results suggest that firstly, anti-LFA-1, but not anti-alpha4 mAb treatment, may be useful in improving the results of intestinal transplantation, and secondly, that this treatment is not incompatible with long term administration of tacrolimus currently used in the prevention of small bowel graft rejection in humans. PMID- 10861271 TI - Role of urokinase and its receptor in basal and stimulated colonic epithelial cell migration in vitro. AB - BACKGROUND: Migration of colonic epithelial cells is important for mucosal repair following injury. The urokinase (u-PA) system regulates migration in other cell types. AIM: To examine the role of u-PA and its receptor (u-PAR) in colonic epithelial cell migration. METHODS: Migration was assessed over 24 hours in circular wounds made in confluent monolayers of LIM1215 and Caco-2 human colon cancer cells. The function of u-PA and u-PAR was ablated with antisense oligonucleotides to block expression, with synthetic u-PA peptides to block interaction, and with aprotinin to block u-PA mediated proteolysis. RESULTS: Migration was stimulated two to threefold by exogenous u-PA, an effect dependent on u-PAR binding but independent of u-PA mediated mitogenesis and proteolysis. Expression of u-PA and u-PAR was inhibited by 80% by the appropriate antisense oligonucleotide. Basal migration and the motogenic effects of butyrate, epidermal growth factor, and phorbol-12-myristate-13-acetate were suppressed by the u-PAR antisense oligonucleotide (40-60%) but were at best minimally affected following inhibition of u-PA expression and binding. CONCLUSIONS: In an in vitro model of wounded colonic epithelium, u-PAR promotes cell migration through mechanisms that are not exclusively dependent on u-PA binding. Therefore, u-PA and u-PAR may contribute to colonic mucosal repair in vivo. PMID- 10861272 TI - Glucagon-like peptide-2 enhances intestinal epithelial barrier function of both transcellular and paracellular pathways in the mouse. AB - BACKGROUND AND AIMS: Glucagon-like peptide-2 (GLP-2) is a recently identified potent intestinotrophic factor. We have evaluated the effect of GLP-2 treatment on intestinal epithelial barrier function in mice. METHODS: CD-1 mice were injected subcutaneously with GLP-2 or a protease resistant analogue, h[Gly(2)]GLP 2, twice daily for up to 10 days. Saline injected mice served as controls. Jejunal segments were mounted in Ussing chambers. Tissue conductance was measured and unidirectional fluxes were determined for (i) Na(+) and the small inert probe Cr-EDTA (both transported via the paracellular pathway) and (ii) the macromolecule horseradish peroxidase (HRP, transported via the transcellular pathway). RESULTS: Mice treated with GLP-2 or h[Gly(2)]GLP-2 for 10 days demonstrated significantly reduced intestinal conductance and fluxes of Na(+), Cr EDTA, and HRP. Electron microscopy confirmed that GLP-2 reduced endocytic uptake of HRP into enterocytes. Functional changes (evident by four hours) preceded morphological changes (evident by 48 hours). CONCLUSIONS: GLP-2 enhances intestinal epithelial barrier function by affecting both paracellular and transcellular pathways and thus may be of therapeutic value in a number of gastrointestinal conditions. PMID- 10861273 TI - Evaluation of prognosis of squamous cell carcinoma of the oesophagus by endoscopic ultrasonography. AB - BACKGROUND/AIMS: For pretherapeutic staging of squamous cell carcinoma of the oesophagus, endoscopic ultrasonography (EUS) is considered the most profitable modality because it can provide cross sectional imaging of the tumour. The aim of this study was to evaluate the relation between prognosis and EUS findings, especially tumour area, in squamous cell carcinoma of the oesophagus. PATIENTS/METHODS: A total of 113 patients with squamous cell carcinoma of the oesophagus underwent EUS for pretherapeutic examination at Nagoya University Hospital. We compared EUS findings, histological results, and outcome. In addition, we measured the area of the tumour on EUS images (n=113) and evaluated if EUS area correlated with volume of the tumour on histological findings (n=50). RESULTS: The overall accuracy rate of EUS was 83.2% (94/113) for depth of tumour invasion and 67.6% (69/102) for perioesophageal lymph node metastasis. The EUS area increased in proportion to the development of tumour infiltration, and patients with lymph node metastasis had a larger EUS area than patients without lymph node metastasis. There was a close correlation between EUS area and volume of the tumour on histological findings. If EUS area of the tumour was less than 50 mm(2), the five year survival rate was 100%. As EUS area increased, the survival rate decreased. CONCLUSIONS: Measurement of EUS area of the tumour is reliable for quantification of the tumour and prediction of prognosis in patients with squamous cell carcinoma of the oesophagus. PMID- 10861274 TI - Diagnostic accuracy and interobserver agreement of CT colonography (virtual colonoscopy). AB - BACKGROUND AND AIMS: Computed tomographic (CT) colonography or virtual colonoscopy (VC) is a non-invasive imaging method proposed for screening patients with colorectal neoplasias. Our aims were to study the diagnostic accuracy and interobserver agreement of VC for correct patient identification compared with conventional colonoscopy (CC). METHODS: This was a prospective study of 50 patients successively undergoing VC and CC. Multiplanar two dimensional CT images and three dimensional VC were constructed using surface rendering software and interpreted by two independent investigator teams. VC findings were compared with those of CC. Interobserver agreement was determined using kappa statistics. RESULTS: CC found 65 polyps in 24 patients. For identification of patients with polyps > or =10 mm, the sensitivity of VC was 38% and 63%, and specificity was 74% and 74% for teams 1 and team 2. Interobserver agreement was good (kappa 0.72). For patients with polyps of any size, the sensitivity of VC was 75% and 71%, and specificity was 62% and 69% for teams 1 and 2. Interobserver agreement was fair (kappa 0.56). Accuracy improved when comparing the results of the first 24 with the last 26 patients. CONCLUSIONS: In our experience, VC had a low diagnostic value for identification of patients with colorectal neoplasias. Interobserver agreement for VC interpretation was fair. These results may be explained by software imperfections and a learning curve effect. PMID- 10861275 TI - Hepatitis C virus related cirrhosis: time to occurrence of hepatocellular carcinoma and death. AB - BACKGROUND: In patients with hepatitis C virus (HCV) infection and cirrhosis, long term outcome and the incidence of hepatocellular carcinoma (HCC) are still debated. DESIGN: From January 1987 to January 1997, 416 patients (240 male, median age 57 years) with uncomplicated Child-Pugh A HCV related cirrhosis were followed in two Paris area centres from diagnosis of cirrhosis until death or reference date (1 June 1998). The analysis used a three state disability model generalising the Cox model. RESULTS: Of the 416 patients, 60 developed HCC with a five year rate of 13.4% (95% confidence interval (CI) 9.0-17.8%) and 83 died (including 34 with HCC), with a five year death rate of 15.3% (95% CI 12.6 18.0%). By multivariable analysis, time to HCC relied on age (hazard ratio (HR) 1.05 per year; p=0.0005), male sex (HR 2.13; p=0.01), oesophageal varices (HR 2.36; p= 0.008), decreased platelet count (HR 0.99; p=0. 03), and bilirubin level (HR 1.01; p=0.003), while death after HCC was mainly related to tobacco consumption (HR 1.04; p=0.0006). In contrast, death free of HCC was dependent on age (HR 1.04; p=0.01), oesophageal varices (HR 2.75; p=0.001), low platelet count (HR 0.99; p=0.006), and albumin level (HR 0.90; p=0.0001). CONCLUSION: The incidence of HCC and mortality should be higher in these patients than previously stated, and prognostic factors of HCC and death are closely related age and symptoms of portal hypertension. PMID- 10861276 TI - Cytotoxic T lymphocyte responses and CTL epitope escape mutation in HBsAg, anti HBe positive individuals. AB - BACKGROUND/AIMS: Clearance of hepatitis B virus (HBV) is characterised by a strong cytotoxic T cell response. Persistence of HBV in chronic hepatitis B carriers may be related to failure of this response. The aim of this study was to determine whether HLA class I restricted cytotoxic T lymphocyte (CTL) responses persist in anti-hepatitis B e (HBe) positive / HBV DNA negative individuals, and to correlate the presence of viral CTL epitope mutation with clinical outcome. METHODS: An HLA/HBV dual transfectant model was used to demonstrate these CTL responses in individuals chronically infected with HBV. Subsequently, a known hepatitis B core (HBc) CTL epitope was sequenced in a family of five chronically infected individuals all sharing a HLA allele (HLA-A68.1). RESULTS: Low level HLA class I restricted cytotoxic T cell responses were detected in the peripheral blood of five of eight anti-HBe positive individuals. In the family of HLA-A68.1 positive chronically infected individuals, mutation of the HLA-A68.1 restricted hepatitis B core antigen (HBcAg) CTL epitope STLPETTVVRR was found in all four anti-HBe positive individuals but not in the sole hepatitis B e antigen (HBeAg) positive patient. CONCLUSION: These data are consistent with a continued immune selection pressure on HBV in anti-HBe positive chronically infected individuals with low replicating HBV infection and suggest that mutation of a CTL epitope may be a consequence of the immune response, as opposed to the cause of viral persistence. PMID- 10861277 TI - Insulin and gall stones: a population case control study in southern Italy. AB - BACKGROUND: Hyperinsulinaemia has been associated with many common diseases in developed countries, such as ischaemic heart disease and colon cancer. Gall stones are also very prevalent in these countries but little is known about the association between insulin and gall stones. AIMS: To study the relationships between insulin and the incidence of gall stones in a sample of the general population. SUBJECTS AND METHODS: Between May 1985 and June 1986, systematic sampling from the electoral register of Castellana, a small town in southern Italy, yielded 2472 subjects who had their gall bladder checked for gall stones by ultrasonography. Between May 1992 and June 1993, 1962 of the 2235 subjects without gall stones at the first examination agreed to a further ultrasound examination. A total of 101 subjects with newly diagnosed gall stones and 303 randomly chosen controls entered the study. Serum insulin was determined by radioimmunoassay, and concentrations of cholesterol, cholesterol high density lipoprotein (HDL), glucose, and triglycerides by standard enzymatic colorimetric methods. Unconditional multiple logistic regression was used to study the association between insulin and gall stones, controlling for the most common confounding factors. RESULTS: In individuals with no clinical diagnosis of diabetes and serum glucose <7 mmol/l, insulin was associated with gall stones. This association persisted even after controlling for sex, age, body mass index, and serum glucose. The risk of gall stones in the highest quintile of serum insulin was 2.66 (95% confidence interval 1.04-6.72; chi(2) test for trend, p=0.03). The association of insulin with gall stones persisted when total and HDL cholesterol were entered in the logistic regression models, and only slightly decreased when serum triglycerides were included in the model. CONCLUSIONS: The results of the study indicate that hyperinsulinaemia may play an important role in the aetiology of gall stones even in individuals without diabetes and with normal serum glucose levels. PMID- 10861282 TI - Polymorphisms of the CYP2D6 gene increase susceptibility to ankylosing spondylitis. AB - Ankylosing spondylitis (AS) is a common and highly familial rheumatic disorder. The sibling recurrence risk ratio for the disease is 63 and heritability assessed in twins >90%. Although MHC genes, including HLA-B27, contribute only 20-50% of the genetic risk for the disease, no non-MHC gene has yet been convincingly demonstrated to influence either susceptibility to the disease or its phenotypic expression. Previous linkage and association studies have suggested the presence of a susceptibility gene for AS close to, or within, the cytochrome P450 2D6 gene (CYP2D6, debrisoquine hydroxylase) located at chromosome 22q13.1. We performed a linkage study of chromosome 22 in 200 families with AS affected sibling-pairs. Association of alleles of the CYP2D6 gene was examined by both case-control and within-family means. For case-control studies, 617 unrelated individuals with AS (361 probands from sibling-pair and parent-case trio families and 256 unrelated non-familial sporadic cases) and 402 healthy ethnically matched controls were employed. For within-family association studies, 361 families including 161 parent-case trios and 200 affected sibling-pair families were employed. Homozygosity for poor metabolizer alleles was found to be associated with AS. Heterozygosity for the most frequent poor metabolizer allele (CYP2D6*4) was not associated with increased susceptibility to AS. Significant within-family association of CYP2D6*4 alleles and AS was demonstrated. Weak linkage was also demonstrated between CYP2D6 and AS. We postulate that altered metabolism of a natural toxin or antigen by the CYP2D6 gene may increase susceptibility to AS. PMID- 10861278 TI - DNA mismatch repair genes and colorectal cancer. PMID- 10861283 TI - The neurofibromatosis 2 tumor suppressor protein interacts with hepatocyte growth factor-regulated tyrosine kinase substrate. AB - The neurofibromatosis 2 tumor suppressor protein schwannomin/merlin is commonly mutated in schwannomas and meningiomas. Schwannomin, a member of the 4.1 family of proteins, which are known to link the cytoskeleton to the plasma membrane, has little known function other than its ability to suppress tumor growth. Using yeast two-hybrid interaction cloning, we identified the HGF-regulated tyrosine kinase substrate (HRS) as a schwannomin interactor. We verified the interaction by both immunoprecipitation of endogenous HRS with endogenous schwannomin in vivo as well as by using bacterially purified HRS and schwannomin in vitro. We narrowed the regions of interaction to include schwannomin residues 256-579 and HRS residues from 480 to the end of either of two HRS isoforms. Schwannomin molecules with a L46R, L360P, L535P or Q538P missense mutation demonstrated reduced affinity for HRS binding. As HRS is associated with early endosomes and may mediate receptor translocation to the lysosome, we demonstrated that schwannomin and HRS co-localize at endosomes using the early endosome antigen 1 in STS26T Schwann cells by indirect immunofluorescence. The identification of schwannomin as a HRS interactor implicates schwannomin in HRS-mediated cell signaling. PMID- 10861284 TI - Deletion in the promoter region and altered expression of Pitx3 homeobox gene in aphakia mice. AB - Mouse aphakia (ak) is a recessive phenotype that spontaneously occurs in the 129/Sv-SlJ strain and is characterized by small eyes that lack a lens. We have recently identified a homeobox-containing gene, Pitx3, and have shown that it is expressed in the developing lens and maps to chromosome 19 close to ak in mouse. Human PITX3 gene was found to underlie anterior segment dysgenesis and cataracts. We have now obtained the entire sequence of the mouse Pitx3 gene including 10 kb of the 5' region and 5 kb of the 3' region. Of several microsatellite repeat regions identified within the Pitx3 sequence, one was informative for linkage analysis. No recombination was observed between ak and the Pitx3 marker, indicating that these two loci are closely linked (0.2 +/- 0.2 cM). Additionally, Pitx3 transcripts were not detected in the ak/ak mice either in the lens placode or at later developmental stages of the lens by in situ hybridization. Since no differences were previously found between ak/ak and wild-type sequences in the Pitx3 coding region, we hypothesized that an etiologic mutation is located in the promoter or other regulatory regions. To test this hypothesis we studied the 5' flanking region of the Pitx3 gene. This analysis revealed a deletion of 652 bp located 2.5 kb upstream from the start point of the Pitx3 5' UTR sequence in ak/ak mice. The deletion co-segregated with the ak mutation and was not detected in 16 samples from 10 different mouse strains including the founder strains. Analysis of the 652 bp region identified sequences similar to consensus binding sites for transcription factors AP-2 and Maf that were shown to play a critical role in lens determination. These lines of evidence suggest that the abnormal ocular development in the aphakia mouse is due to the deletion upstream of the Pitx3 gene. PMID- 10861285 TI - Human GRB10 is imprinted and expressed from the paternal and maternal allele in a highly tissue- and isoform-specific fashion. AB - As part of a systematic screen for novel imprinted genes of human chromosome 7 we have investigated GRB10, which belongs to a small family of adapter proteins, known to interact with a number of receptor tyrosine kinases and signalling molecules. Upon allele-specific transcription analysis involving multiple distinct splice variants in various fetal tissues, we found that human GRB10 is imprinted in a highly isoform- and tissue-specific manner. In fetal brains, most variants are transcribed exclusively from the paternal allele. Imprinted expression in this tissue is not accompanied by allele-specific methylation of the most 5' CpG island. In skeletal muscle, one GRB10 isoform, gamma1, is expressed from the maternal allele alone, whereas in numerous other fetal tissues, all GRB10 splice variants are transcribed from both parental alleles. A remarkable finding is paternal-specific expression of GRB10 in the human fetal brain, since, in the mouse, this gene is transcribed exclusively from the maternal allele. To our knowledge, this is the first example of a gene that is oppositely imprinted in mouse and human. PMID- 10861286 TI - Therapeutic liver repopulation in a mouse model of hypercholesterolemia. AB - Liver repopulation constitutes an attractive approach for the treatment of liver disorders or of diseases requiring abundant secretion of an active protein. We have described previously a model of selective repopulation of a normal liver by Fas/CD95-resistant hepatocytes, in which we achieved up to 16% hepatocyte repopulation. In the present study, we investigated the therapeutic efficacy of this strategy. With this aim, apolipoprotein E (ApoE) knockout mice were transplanted with Fas/CD95-resistant hepatocytes which constitutively express ApoE. Transplanted mice were submitted to weekly injections of non-lethal doses of the Fas agonist antibody Jo2. After 8 weeks of treatment, we obtained up to 30% of the normal level of plasma ApoE. ApoE secretion was accompanied by a drastic and significant decrease in total plasma cholesterol, which even fell to normal levels. Moreover, this secretion was sufficient to markedly reduce the progression of atherosclerosis. These results demonstrate the efficacy of this repopulation approach for correcting a deficiency in a protein secreted by the liver. PMID- 10861287 TI - A neonatal lethal mutation in FGFR3 uncouples proliferation and differentiation of growth plate chondrocytes in embryos. AB - We have generated the first mouse model of fibro-blast growth factor receptor 3 (Fgfr3) with the K644E mutation, which accurately reflects the embryonic onset of a neonatal lethal dwarfism, thanatophoric dysplasia type II (TDII). Long-bone abnormalities were identified as early as embryonic day 14, during initiation of endochondral ossification. Increased expression of PATCHED: (PTC:) was observed, independent of unaltered expression of parathyroid hormone-related peptide (PTHrP) receptor and Indian Hedgehog (IHH:), suggesting a new regulatory role for Fgfr3 in embryos. We demonstrate that the mutation enhances chondrocyte proliferation during the early embryonic skeletal development, in contrast to previous reports that showed decreased proliferation in postnatal-onset dwarf mice with activating Fgfr3 mutations. This suggests that signaling through Fgfr3 both promotes and inhibits chondrocyte proliferation, depending on the time during development. In contrast, suppressed chondrocyte differentiation was observed throughout the embryonic stages, defining decreased differentiation as the primary cause of retarded longitudinal bone growth in TDII. This model was successfully crossed with a cartilage-specific CRE: transgenic strain, excluding the lung as the primary cause of lethality. PMID- 10861288 TI - A deletion encompassing Zic3 in bent tail, a mouse model for X-linked neural tube defects. AB - Bent tail is a mouse model for human neural tube defects. Bent tail mice are characterized by a shortened, kinked tail. We have observed numerous aberrations in Bent tail embryos including exencephaly, rotation defects and occasionally omphalocele, orofacial schisis and situs abnormalities. Exencephaly was seen in >10% of all embryos and resulted from a closure defect of the hindbrain. Bent tail maps to the proximal part of the X chromosome. By haplotype analysis we have appointed the Bent tail locus to a 1.1 cM interval between markers DXMit159 and DXMit143. Subsequent analysis has revealed the presence of a deletion in all affected animals. The deletion is approximately 1 Mb in size and encompasses the gene for ZIC:3, a zinc finger transcription factor expressed in murine neuroectoderm and dorsal axial mesoderm during neurulation. ZIC:3 is a homolog of the Drosophila segmentation gene odd-paired. Although the Bent tail phenotype probably is the result of the deletion of several genes, combining data on ZIC:3 expression and function of ZIC: genes in the mouse shows that deletion of Zic3 alone is compatible with a major role of this gene in the congenital malformations of the Bent tail mouse. In man, mutations in ZIC3 are associated with situs abnormalities. These patients occasionally also show spina bifida, indicating that genetic variation in human ZIC3 may contribute to other congenital malformations, including neural tube defects. PMID- 10861289 TI - Mammalian artificial chromosome formation from circular alphoid input DNA does not require telomere repeats. AB - Mammalian artificial chromosomes (MACs) form in HT1080 cells after transfecting linear yeast artificial chromosome constructs minimally containing competent alphoid arrays, a selectable marker and terminal human telomere repeats. Restrictions on the structure of input DNA in MAC formation were investigated by transfecting circular or linear alphoid constructs with or without human telomere arrays and by varying the position and orientation of the telomere arrays on input linear constructs. Circular input DNA efficiently produced MACs. Absence of telomere arrays from circular input molecules did not significantly alter MAC formation rates. Linear constructs capped with telomere arrays generated MACs effectively, but a severe reduction in MAC formation was observed from linear constructs lacking telomere arrays. Human telomere arrays positioned 1-5 kb from linear construct ends and in either orientation were able to promote MAC formation with similar efficiencies. Both circular and linear input constructs generated artificial chromosomes that efficiently segregated in the absence of selection. Telomeres were not detected on the MACs, regardless of the inclusion of telomere arrays on input DNA, suggesting that circular chromosomes were formed. We found no evidence for acquisition of host cell DNA, which is consistent with de novo chromosome assembly. PMID- 10861290 TI - Expression of the PTEN tumour suppressor protein during human development. AB - The tumour suppressor gene PTEN, localized to 10q23.3, is the susceptibility gene for Cowden syndrome (CS) and Bannayan-Riley-Ruvalcaba (BRR) syndrome, two hamartoma syndromes with an increased risk of breast and thyroid tumours. Somatic mutations have been found in a variety of human tumours. Functional studies have revealed that PTEN plays a fundamental role in cellular growth, death, adhesion and migration. RNA in situ hybridization using the pten coding region in mouse embryos showed ubiquitous transcription, providing evidence that pten could play a versatile role throughout murine development. Nothing is known regarding the pattern of PTEN expression during human development. Here, we present the pattern of PTEN expression during human development using a specific monoclonal antibody and examine the relationship of the temporal and spatial expression pattern to the clinical manifestations of CS and BRR, the somatic genetic data in sporadic cancers, the murine knockout models and the RNA expression data in mouse embryos. We observed mainly high-level PTEN expression in tissues (e.g. skin, thyroid and central nervous system) known to be involved in CS and BRR. In addition, we identified tissues (e.g. peripheral nervous system, autonomomic nervous system and upper gastrointestinal tract) with high PTEN expression not commonly known to play a role in these syndromes nor in sporadic tumorigenesis in those organs. This knowledge may help in identifying roles for PTEN which, as of today, are unknown or even unsuspected. PMID- 10861291 TI - Strong homophilic interactions of the Ig-like domains of polycystin-1, the protein product of an autosomal dominant polycystic kidney disease gene, PKD1. AB - The 14 kb mRNA of the polycystic kidney disease gene PKD1 encodes a novel large (approximately 460 kDa) protein, polycystin-1, of unknown function that is responsible for autosomal dominant polycystic kidney disease (ADPKD). The unique organization of multiple adhesive domains of polycystin-1, including 16 Ig-like domains (or PKD domains) suggests that it may play an important role in cell cell/cell-matrix interactions. Here we demonstrate the localization of polycystin 1 to epithelial cell-cell contacts in culture. These results along with structural predictions prompted us to propose that polycystin-1 is involved in cell-cell adhesion through its cluster of Ig-like repeats. We show that Ig-like domains II-XVI are involved in strong calcium-independent homophilic interactions in vitro. Domains XI-XVI form interactions with high affinity (K(d) = 60 nM) and domains II-V exhibit the lowest binding affinity (K(d) = 730 nM) in these studies. Most importantly, we show that antibodies raised against Ig-like domains of polycystin-1 disrupt cell-cell interactions in MDCK cell monolayers, thus indicating that polycystin-1 is directly involved in the cell-cell adhesion process. Collectively, these data suggest that interactions of the Ig-like repeats of polycystin-1 play an important role in mediating intercellular adhesion. We suggest that the loss of these interactions due to mutations in polycystin-1 may be an important step in cystogenesis. PMID- 10861292 TI - Common chromosomal fragile site FRA16D sequence: identification of the FOR gene spanning FRA16D and homozygous deletions and translocation breakpoints in cancer cells. AB - Fluorescence in situ hybridization of a tile path of DNA subclones has previously enabled the cyto-genetic definition of the minimal DNA sequence which spans the FRA16D common chromosomal fragile site, located at 16q23.2. Homozygous deletion of the FRA16D locus has been reported in adenocarcinomas of stomach, colon, lung and ovary. We have sequenced the 270 kb containing the FRA16D fragile site and the minimal homozygously deleted region in tumour cells. This sequence enabled localization of some of the tumour cell breakpoints to regions which contain AT rich secondary structures similar to those associated with the FRA10B and FRA16B rare fragile sites. The FRA16D DNA sequence also led to the identification of an alternatively spliced gene, named FOR (fragile site FRA16D oxidoreductase), exons of which span both the fragile site and the minimal region of homozygous deletion. In addition, the complete DNA sequence of the FRA16D-containing FOR intron reveals no evidence of additional authentic transcripts. Alternatively spliced FOR transcripts (FOR I, FOR II and FOR III) encode proteins which share N terminal WW domains and differ at their C-terminus, with FOR III having a truncated oxidoreductase domain. FRA16D-associated deletions selectively affect the FOR gene transcripts. Three out of five previously mapped translocation breakpoints in multiple myeloma are also located within the FOR gene. FOR is therefore the principle genetic target for DNA instability at 16q23.2 and perturbation of FOR function is likely to contribute to the biological consequences of DNA instability at FRA16D in cancer cells. PMID- 10861293 TI - Regions of genomic instability on 22q11 and 11q23 as the etiology for the recurrent constitutional t(11;22). AB - The constitutional t(11;22)(q23;q11) is the only known recurrent, non Robertsonian translocation. To analyze the genomic structure of the breakpoint, we have cloned the junction fragments from the der(11) and der(22) of a t(11;22) balanced carrier. On chromosome 11 the translocation occurs within a short, palindromic AT-rich region (ATRR). Likewise, the breakpoint on chromosome 22 has been localized within an ATRR that is part of a larger palindrome. Interestingly, the 22q11 breakpoint falls within one of the 'unclonable' gaps in the genomic sequence. Further, a sequenced chromosome 11 BAC clone, spanning the t(11;22) breakpoint in 11q23, is deleted within the palindromic ATRR, suggesting instability of this region in bacterial clones. Several unrelated t(11;22) families demonstrate similar breakpoints on both chromosomes, indicating that their translocations are within the same palindrome. It is likely that the palindromic ATRRs produce unstable DNA structures in 22q11 and 11q23 that are responsible for the recurrent t(11;22) translocation. PMID- 10861294 TI - DNA structural properties of AF9 are similar to MLL and could act as recombination hot spots resulting in MLL/AF9 translocations and leukemogenesis. AB - The human AF9 gene at 9p22 is one of the most common fusion partner genes with the MLL gene at 11q23, resulting in the t(9;11)(p22;q23). The MLL-AF9 fusion gene is associated with de novo acute myelo-genous leukemia (AML), rarely with acute lymphocytic leukemia (ALL) and with therapy related leukemia (t-AML). The AF9 gene is >100 kb and two patient breakpoint cluster regions (BCRs) have been identified; BCR1 is within intron 4, previously called site A, whereas BCR2 or site B spans introns 7 and 8. Patient breakpoint locations were determined previously by RT-PCR and by genomic DNA cloning. In this study, we defined the exon-intron boundaries and identified several different structural elements in AF9 including a co-localizing in vivo DNA topo II cleavage site and an in vitro DNase I hypersensitive (DNase 1 HS) site in intron 7 in BCR2. Reversibility experiments demonstrated a religation of the topo II cleavage sites. The location of the in vivo topo II cleavage site was confirmed in vitro using a topo II cleavage assay. In addition, two scaffold associated regions (SARs) are located centromeric to the topo II and DNase I HS cleavage sites and border both patient breakpoint regions: SAR1 is located in intron 4, whereas SAR2 encompasses parts of exons 5-7. This study demonstrates that the patient breakpoint regions of AF9 share the same structural elements as the MLL BCR. We describe a DNA breakage and repair model for non-homologous recombination between MLL and its partner genes, particularly AF9. PMID- 10861295 TI - DSCR1, overexpressed in Down syndrome, is an inhibitor of calcineurin-mediated signaling pathways. AB - Down syndrome is one of the major causes of mental retardation and congenital heart malformations. Other common clinical features of Down syndrome include gastrointestinal anomalies, immune system defects and Alzheimer's disease pathological and neurochemical changes. The most likely consequence of the presence of three copies of chromosome 21 is the overexpression of its resident genes, a fact which must underlie the pathogenesis of the abnormalities that occur in Down syndrome. Here we show that DSCR1, the product of a chromosome 21 gene highly expressed in brain, heart and skeletal muscle, is overexpressed in the brain of Down syndrome fetuses, and interacts physically and functionally with calcineurin A, the catalytic subunit of the Ca(2+)/calmodulin-dependent protein phosphatase PP2B. The DSCR1 binding region in calcineurin A is located in the linker region between the calcineurin A catalytic domain and the calcineurin B binding domain, outside of other functional domains previously defined in calcineurin A. DSCR1 belongs to a family of evolutionarily conserved proteins with three members in humans: DSCR1, ZAKI-4 and DSCR1L2. We further demonstrate that overexpression of DSCR1 and ZAKI-4 inhibits calcineurin-dependent gene transcription through the inhibition of NF-AT translocation to the nucleus. Together, these results suggest that members of this newly described family of human proteins are endogenous regulators of calcineurin-mediated signaling pathways and as such, they may be involved in many physiological processes. PMID- 10861296 TI - The neuronal ceroid lipofuscinosis CLN8 membrane protein is a resident of the endoplasmic reticulum. AB - Progressive epilepsy with mental retardation (EPMR) is a new member of the neuronal ceroid lipofuscinoses (NCLs). The CLN8 gene underlying EPMR was recently identified. It encodes a novel 286 amino acid transmembrane protein that contains an endoplasmic reticulum (ER)-retrieval signal (KKRP) in its C-terminus. A homozygous mutation in the orthologous mouse gene (Cln8) underlies the phenotype of a naturally occurring NCL model, the motor neuron degeneration mouse (mnd). To characterize the product of the CLN8 gene and to determine its intracellular localization, we expressed CLN8 cDNA in BHK, HeLa and CHO cell lines. In western blotting and pulse-chase analyses an approximately 33 kDa protein that does not undergo proteolytic processing steps was detected. Using CLN8 and cell organelle specific antibodies with confocal immunofluorescence microscopy the CLN8 protein was shown to localize in the ER. Partial localization to the ER-Golgi intermediate compartment (ERGIC) was also observed. The ER-ERGIC localization was not altered in the CLN8 protein representing the EPMR mutation. However, mnd mutant protein was only found in the ER. Mutations in the ER retrieval signal KKRP resulted in localization of CLN8 to the Golgi apparatus. Taken together, these data strongly suggest that CLN8 is an ER resident protein that recycles between ER and ERGIC. PMID- 10861297 TI - Paradoxical influence of acid beta-galactosidase gene dosage on phenotype of the twitcher mouse (genetic galactosylceramidase deficiency). AB - We have cross-bred twitcher mice (galactosylceramidase deficiency) and acid beta galactosidase knockout mice (G(M1) gangliosidosis) and found that the acid beta galactosidase gene dosage exerts an unexpected and paradoxical influence on the twitcher phenotype. Twitcher mice with an additional complete deficiency of acid beta-galactosidase have the mildest phenotype with the longest lifespan and nearly rescued CNS pathology. In contrast, twitcher mice with a single functional acid beta-galactosidase gene have the most severe disease with the shortest lifespan, despite the fact that G(M1) gangliosidosis carrier mice with an otherwise normal genetic background are phenotypically normal. A significant proportion of these galc(-/-), bgal(+/-) mice clinically develop additional extreme hyper-reactivity and generalized seizures not seen in any other genotypes. Consistent with the clinical seizures, widespread neuronal degeneration is present in the galc(-/-), bgal(+/-) mice, most prominently in the CA3 region of the hippocampus. The double knockout mice show a massive accumulation of lactosylceramide in all tissues. The brain inexplicably contains only a half-normal amount of galactosylceramide, which may account for the mild clinical and pathological phenotype. On the other hand, brain psychosine level is increased in all twitcher mice, but galc(-/-), bgal(+/-) mice show a significantly higher level than other genotypes. The reduced galactosylceramide in the brain of the double knockout mice and the significantly higher psychosine in the brain of the galc(-/-), bgal(+/-) mice cannot readily be explained from the genotypes of these mice. These observations are contrary to the expected outcome of Mendelian autosomal recessive single gene disorders and may also be interpreted as that the acid beta-galactosidase gene functions as a modifier gene for the phenotypic expression of genetic galactosylceramidase deficiency. PMID- 10861298 TI - Functional differences of the PDS gene product are associated with phenotypic variation in patients with Pendred syndrome and non-syndromic hearing loss (DFNB4). AB - The PDS gene encodes a transmembrane protein, known as pendrin, which functions as a transporter of iodide and chloride. Mutations in this gene are responsible for Pendred syndrome and autosomal recessive non-syndromic hearing loss at the DFNB4 locus on chromosome 7q31. A screen of 20 individuals from the midwestern USA with non-syndromic hearing loss and dilated vestibular aqueducts identified three people (15%) with PDS mutations. To determine whether PDS mutations in individuals with Pendred syndrome differ functionally from PDS mutations in individuals with non-syndromic hearing loss, we compared three common Pendred syndrome allele variants (L236P, T416P and E384G), with three PDS mutations reported only in individuals with non-syndromic hearing loss (V480D, V653A and I490L/G497S). The mutations associated with Pendred syndrome have complete loss of pendrin-induced chloride and iodide transport, while alleles unique to people with DFNB4 are able to transport both iodide and chloride, albeit at a much lower level than wild-type pendrin. We hypothesize that this residual level of anion transport is sufficient to eliminate or postpone the onset of goiter in individuals with DFNB4. We propose a model for pendrin function in the thyroid in which pendrin transports iodide across the apical membrane of the thyrocyte into the colloid space. PMID- 10861299 TI - Screening for breast cancer: how useful are clinical breast examinations? PMID- 10861300 TI - Chemoprevention of lung cancer is proving difficult and frustrating, requiring new approaches. PMID- 10861301 TI - High-dose chemo for breast cancer: does it still have a chance? PMID- 10861302 TI - Embargoes and economics: the birth of biotechnology in Cuba. PMID- 10861303 TI - Does the U.S. Embargo affect cuban health care? PMID- 10861304 TI - Avalanche of direct-to-consumer drug marketing brings new questions. PMID- 10861305 TI - Cancer therapies touted in physician, hospital, web advertising. PMID- 10861306 TI - Tyrosine kinase inhibitor research presses on despite halted clinical trial. PMID- 10861307 TI - Stat bite: Use of sigmoidoscopy or proctoscopy in the U.S. by age, 1997. PMID- 10861308 TI - Findings from 752,081 clinical breast examinations reported to a national screening program from 1995 through 1998. AB - BACKGROUND AND METHODS: Mammography programs have received extensive study, but little is known about the outcome of clinical breast examinations (CBEs) performed in community settings. Consequently, we analyzed data from the National Breast and Cervical Cancer Early Detection Program on CBEs provided to low-income women from 1995 through 1998 and determined the percentage of CBEs considered to be abnormal, suspicious for cancer; the rates of cancer detection; and the sensitivity, specificity, and positive predictive value of CBEs. RESULTS: We analyzed data from 752081 CBEs and found that 6.9% of all CBEs were coded abnormal, suspicious for cancer, and that 5.0 cancers were detected per 1000 examinations (95% confidence interval [CI] = 4.9-5.2). The values observed for sensitivity (58.8%) and specificity (93.4%) were comparable to those reported for the CBE component of clinical trials. The observed positive predictive value was 4.3%. About 74% of all records also reported mammography results. The cancer detection rate among records reporting an abnormal CBE and normal mammography was 7.4 cancers per 1000 records (95% CI = 6. 3-8.4). When the CBE was normal but the mammography was abnormal, the rate was 42.0 cancers per 1000 records (95% CI = 39.9-44.1). When both CBE and mammography results were abnormal, the rate was 170.3 cancers per 1000 records (95% CI = 162.7-177.9). Cancer detection could not be attributed entirely to CBE or mammography on 38% of the records in the latter subset because the tests were performed on the same day. CONCLUSION: CBEs performed in community-based screening programs can detect breast cancers as effectively as CBEs performed in clinical trials and may modestly improve early detection campaigns. PMID- 10861309 TI - EUROSCAN, a randomized trial of vitamin A and N-acetylcysteine in patients with head and neck cancer or lung cancer. For the EUropean Organization for Research and Treatment of Cancer Head and Neck and Lung Cancer Cooperative Groups. AB - BACKGROUND: Preclinical evidence suggests that retinoids and antioxidants may prevent or delay the occurrence of cancer in the upper or lower airways, but such effects have not been reliably established in clinical studies. To assess the chemopreventive effects of vitamin A (retinyl palmitate) and N-acetylcysteine, we conducted a large randomized intervention study in patients with head and neck cancer or with lung cancer, most of whom had a history of smoking. METHODS: From June 1988 through July 1994, a total of 2592 patients (60% with head and neck cancer and 40% with lung cancer) were randomly assigned to receive 1) retinyl palmitate (300000 IU daily for 1 year followed by 150000 IU for a 2(nd) year), 2) N-acetylcysteine (600 mg daily for 2 years), 3) both compounds, or 4) no intervention. All statistical tests were two-sided. RESULTS: Of the patients, 93.5% had smoked tobacco at sometime in their lives (and 25% continued to smoke after cancer diagnosis). After a median follow-up of 49 months, 916 patients were reported with an event (recurrence, second primary tumor, or death). No statistically significant difference was observed in overall survival or event free survival between patients who received retinyl palmitate and patients who did not. Similarly, no difference was seen in overall survival or event-free survival between patients who received N-acetylcysteine and patients who did not. There was a lower incidence of second primary tumors in the no intervention arm, but the difference was not statistically significant. CONCLUSION: A 2-year supplementation of retinyl palmitate and/or N-acetylcysteine resulted in no benefit-in terms of survival, event-free survival, or second primary tumors-for patients with head and neck cancer or with lung cancer, most of whom were previous or current smokers. PMID- 10861310 TI - Effects of c-erbB2 overexpression on the drug sensitivities of normal human mammary epithelial cells. AB - BACKGROUND: Overexpression of the gene c-erbB2, which encodes a receptor tyrosine kinase, in breast tumors has been linked with either increased or decreased response of breast cancer patients to various therapies. In breast cancer cell lines, overexpression of exogenous c-erbB2 sometimes alters drug sensitivities but sometimes has no effect. To avoid the genetic complexities associated with established cancer cell lines, normal human mammary epithelial cells (HMECs) were studied to determine whether c-erbB2 overexpression by itself would alter chemosensitivity. METHODS: HMECs were designed to overexpress c-erbB2, and these cells were then evaluated for alterations in chemosensitivity. RESULTS: HMECs overexpressing c-erbB2 failed to show any alterations in chemosensitivity to a panel of chemotherapeutic agents, as indicated by 95% confidence intervals on growth curves of cells treated with or without the agent of interest. With the use of fluorescence-activated cell sorting to enrich for HMECs overexpressing c erbB2 on their surface, an 85% pure population of cells was isolated and their chemosensitivity was evaluated. Again, the cells failed to display any alterations in chemosensitivity. CONCLUSIONS: These results suggest that overexpression of c-erbB2 is not sufficient by itself to induce changes in chemosensitivity. Cellular studies using normal human cells in which the complexity of the system can be carefully controlled by the addition of one, two, or even more genes associated with cancer development may provide valuable information about how the products of the genes interact with each other and which combinations are critical in regulating chemosensitivity. PMID- 10861311 TI - Diurnal cortisol rhythm as a predictor of breast cancer survival. AB - BACKGROUND: : Abnormal circadian rhythms have been observed in patients with cancer, but the prognostic value of such alterations has not been confirmed. We examined the association between diurnal variation of salivary cortisol in patients with metastatic breast cancer and subsequent survival. We explored relationships between cortisol rhythms, circulating natural killer (NK) cell counts and activity, prognostic indicators, medical treatment, and psychosocial variables. METHODS: Salivary cortisol levels of 104 patients with metastatic breast cancer were assessed at study entry at 0800, 1200, 1700, and 2100 hours on each of 3 consecutive days, and the slope of diurnal cortisol variation was calculated using a regression of log-transformed cortisol concentrations on sample collection time. NK cell numbers were measured by flow cytometry, and NK cell activity was measured by the chromium release assay. The survival analysis was conducted by the Cox proportional hazards regression model with two-sided statistical testing. RESULTS: Cortisol slope predicted subsequent survival up to 7 years later. Earlier mortality occurred among patients with relatively "flat" rhythms, indicating a lack of normal diurnal variation (Cox proportional hazards, P =. 0036). Patients with chest metastases, as opposed to those with visceral or bone metastases, had more rhythmic cortisol profiles. Flattened profiles were linked with low counts and suppressed activity of NK cells. After adjustment for each of these and other factors, the cortisol slope remained a statistically significant, independent predictor of survival time. NK cell count emerged as a secondary predictor of survival. CONCLUSIONS: Patients with metastatic breast cancer whose diurnal cortisol rhythms were flattened or abnormal had earlier mortality. Suppression of NK cell count and NK function may be a mediator or a marker of more rapid disease progression. PMID- 10861312 TI - alpha-fetoprotein levels in maternal serum during pregnancy and maternal breast cancer incidence. AB - BACKGROUND: A full-term pregnancy is associated with a reduced risk of breast cancer, but the underlying biologic mechanism has not been elucidated. During pregnancy, maternal serum levels of alpha-fetoprotein, an estradiol-binding protein, rise sharply. In culture, alpha-fetoprotein inhibits the growth of estrogen-sensitive cells, including estrogen-sensitive breast cancer cells. Thus, we investigated whether a high level of alpha-fetoprotein in maternal serum during pregnancy is associated with a reduced risk of breast cancer. METHODS: From a population-based cohort of 42057 pregnant women in Denmark, enrolled in an alpha-fetoprotein-screening program from 1978 through 1996, we obtained a complete reproductive history, vital status, and a possible diagnosis of breast cancer (in 117 women) to the end of follow-up on September 1, 1998. RESULTS: During pregnancy, women with an alpha-fetoprotein level greater than or equal to the median value had a 41% lower risk of breast cancer than women with an alpha fetoprotein level below the median value (relative risk [RR] = 0.59; 95% confidence interval [CI] = 0.41-0. 85). RRs for breast cancer by mother's age at childbirth were as follows: 29 years or younger, RR = 0.21 (95% CI = 0.08-0.56); 30-34 years, RR = 0.61 (95% CI = 0.32-1.14); 35-37 years, RR = 0.96 (95% CI = 0.49-1.89); and 38 years or older, RR = 0.71 (95% CI = 0.29-1. 75) (P for trend =.02). Further analyses suggested that high levels of alpha-fetoprotein were associated with a reduced incidence of aggressive disease. The most striking finding was that women with high levels of serum alpha-fetoprotein, compared with women with low levels of serum alpha-fetoprotein, showed a particularly reduced incidence of large tumors (>2 cm; RR = 0.24 [95% CI = 0.11-0.50]). CONCLUSION: A high level of alpha-fetoprotein in maternal serum during any pregnancy is associated with a low overall incidence of breast cancer and, in particular, with a low incidence of advanced breast cancer at diagnosis. This association appears particularly strong for a pregnancy occurring at a young age. PMID- 10861313 TI - Genotype-phenotype relationships in U.S. melanoma-prone families with CDKN2A and CDK4 mutations. AB - BACKGROUND: Two genes have been implicated in the development of cutaneous malignant melanoma (CMM). CDK4 (the gene encoding cyclin-dependent kinase 4, an oncogene) has exhibited germline mutations found in only three melanoma-prone families to date. CDKN2A is a tumor suppressor gene that encodes p16 (which inhibits activity of the cyclin D1-CDK4 complex) with germline mutations detected in 10%-25% of melanoma-prone families, some of whom are also prone to pancreatic cancer. METHODS: We compared 104 CMM patients from 17 CDKN2A families and 12 CMM case subjects from two CDK4 families. We used nonparametric statistics to test for differences in median age at first CMM diagnosis, numbers of CMMs, and numbers of nevi. The three recurrent mutations were haplotyped. All P values were two-sided. RESULTS: The median age at CMM diagnosis (P =.70) and the median numbers of CMMs (P =.73) did not differ between CMM case subjects from CDKN2A versus CDK4 families. Assessment of CMM case subjects from CDKN2A families with and without pancreatic cancer revealed no statistically significant differences in median age at diagnosis (P =.80) or in tumor number (P =.24). There was, however, a statistically significant difference in age-adjusted median numbers of nevi (P =.004), and CMM case subjects from CDKN2A families without pancreatic cancer had greater numbers of nevi. Recurrent CDKN2A mutations were a change from valine to aspartic acid at codon 126 (n = 3) and from glycine to tryptophan at codon 101 (n = 3). Six CDKN2A families had pancreatic cancer. Both CDK4 families carried a mutation resulting in an arginine-to-cysteine substitution at codon 24. Analyses of recurrent CDKN2A and CDK4 mutations suggested common haplotypes. CONCLUSIONS: The recurrent CDKN2A mutations were observed in families with and without pancreatic cancer, which suggests that other factors may be involved in the development of pancreatic cancer. Despite hypothetical differences in the mechanisms of action between CDKN2A and CDK4, clinical factors were indistinguishable between CMM case subjects from CDKN2A versus CDK4 families. PMID- 10861314 TI - Rapid reporting and review of an increased incidence of a known adverse event. PMID- 10861315 TI - Re: Emerging technologies and cervical cancer. PMID- 10861316 TI - Re: Beta-carotene: a miss for epidemiology. PMID- 10861318 TI - RESPONSE: re: beta-carotene: a miss for epidemiology PMID- 10861317 TI - Re: Beta-carotene: a miss for epidemiology. PMID- 10861319 TI - Graft-versus-host disease in children who have received a cord-blood or bone marrow transplant from an HLA-identical sibling. Eurocord and International Bone Marrow Transplant Registry Working Committee on Alternative Donor and Stem Cell Sources. AB - BACKGROUND: Umbilical-cord blood as an alternative to bone marrow for hematopoietic stem-cell transplantation may lower the risk of graft-versus-host disease (GVHD). METHODS: We studied the records of 113 recipients of cord blood from HLA-identical siblings from the period from 1990 through 1997 and compared them with the records of 2052 recipients of bone marrow from HLA-identical siblings during the same period. The study population consisted of children 15 years of age or younger. We compared the rates of GVHD, hematopoietic recovery, and survival using Cox proportional-hazards regression to adjust for potentially confounding factors. RESULTS: Recipients of cord blood were younger than recipients of bone marrow (median age, 5 years vs. 8 years; P<0.001), weighed less (median weight, 17 kg vs. 26 kg; P<0.001), and were less likely to have received methotrexate for prophylaxis against GVHD (28 percent vs. 65 percent, P<0.001). Multivariate analysis demonstrated a lower risk of acute GVHD (relative risk, 0.41; P=0.001) and chronic GVHD (relative risk, 0.35; P=0.02) among recipients of cord-blood transplants. As compared with recovery after bone marrow transplantation, the likelihood of recovery of the neutrophil count and the platelet count was significantly lower in the first month after cord-blood transplantation (relative risk, 0.40 [P<0.001], and relative risk, 0.20 [P<0.001]), respectively. Mortality was similar in the two groups (relative risk of death in the recipients of cord blood, 1.15; P=0.43). CONCLUSIONS: Recipients of cord-blood transplants from HLA-identical siblings have a lower incidence of acute and chronic GVHD than recipients of bone marrow transplants from HLA identical siblings. PMID- 10861320 TI - Causes and outcomes of the acute chest syndrome in sickle cell disease. National Acute Chest Syndrome Study Group. AB - BACKGROUND: The acute chest syndrome is the leading cause of death among patients with sickle cell disease. Since its cause is largely unknown, therapy is supportive. Pilot studies with improved diagnostic techniques suggest that infection and fat embolism are underdiagnosed in patients with the syndrome. METHODS: In a 30-center study, we analyzed 671 episodes of the acute chest syndrome in 538 patients with sickle cell disease to determine the cause, outcome, and response to therapy. We evaluated a treatment protocol that included matched transfusions, bronchodilators, and bronchoscopy. Samples of blood and respiratory tract secretions were sent to central laboratories for antibody testing, culture, DNA testing, and histopathological analyses. RESULTS: Nearly half the patients were initially admitted for another reason, mainly pain. When the acute chest syndrome was diagnosed, patients had hypoxia, decreasing hemoglobin values, and progressive multilobar pneumonia. The mean length of hospitalization was 10.5 days. Thirteen percent of patients required mechanical ventilation, and 3 percent died. Patients who were 20 or more years of age had a more severe course than those who were younger. Neurologic events occurred in 11 percent of patients, among whom 46 percent had respiratory failure. Treatment with phenotypically matched transfusions improved oxygenation, with a 1 percent rate of alloimmunization. One fifth of the patients who were treated with bronchodilators had clinical improvement. Eighty-one percent of patients who required mechanical ventilation recovered. A specific cause of the acute chest syndrome was identified in 38 percent of all episodes and 70 percent of episodes with complete data. Among the specific causes were pulmonary fat embolism and 27 different infectious pathogens. Eighteen patients died, and the most common causes of death were pulmonary emboli and infectious bronchopneumonia. Infection was a contributing factor in 56 percent of the deaths. CONCLUSIONS: Among patients with sickle cell disease, the acute chest syndrome is commonly precipitated by fat embolism and infection, especially community-acquired pneumonia. Among older patients and those with neurologic symptoms, the syndrome often progresses to respiratory failure. Treatment with transfusions and bronchodilators improves oxygenation, and with aggressive treatment, most patients who have respiratory failure recover. PMID- 10861321 TI - Long-term treatment of primary pulmonary hypertension with aerosolized iloprost, a prostacyclin analogue. AB - BACKGROUND: Continuous intravenous infusion of epoprostenol (prostacyclin) is an effective treatment for primary pulmonary hypertension. This approach requires the insertion of a permanent central venous catheter, with the associated risk of serious complications. Recently, aerosolized iloprost, a stable prostacyclin analogue, has been introduced as an alternative therapy for severe pulmonary hypertension. METHODS: We evaluated the effects of aerosolized iloprost on exercise capacity and hemodynamic variables over a one-year period in patients with primary pulmonary hypertension. RESULTS: Twenty-four patients with primary pulmonary hypertension received aerosolized iloprost at a daily dose of 100 or 150 microg for at least one year. The mean (+/-SD) distance covered in the six minute walk test increased from 278+/-96 m at base line to 363+/-135 m after 12 months (P<0.001). During the same period, the mean pulmonary arterial pressure before the inhalation of iloprost declined from 59+/-10 mm Hg to 52+/-15 mm Hg (P=0.006), cardiac output increased from 3.8+/-1.4 liters per minute to 4.4+/-1.3 liters per minute (P=0.02), and pulmonary vascular resistance declined from 1205+/-467 dyn x sec x cm(-5) to 925+/-469 dyn x sec x cm(-5) (P<0.001). The treatment was generally well tolerated, except for mild coughing, minor headache, and jaw pain in some patients. CONCLUSIONS: Long-term treatment with aerosolized iloprost is safe and has sustained effects on exercise capacity and pulmonary hemodynamics in patients with primary pulmonary hypertension. PMID- 10861322 TI - Acromegaly caused by secretion of growth hormone by a non-Hodgkin's lymphoma. PMID- 10861323 TI - Images in clinical medicine. Bacterial perianal dermatitis. PMID- 10861324 TI - A comparison of observational studies and randomized, controlled trials. AB - BACKGROUND: For many years it has been claimed that observational studies find stronger treatment effects than randomized, controlled trials. We compared the results of observational studies with those of randomized, controlled trials. METHODS: We searched the Abridged Index Medicus and Cochrane data bases to identify observational studies reported between 1985 and 1998 that compared two or more treatments or interventions for the same condition. We then searched the Medline and Cochrane data bases to identify all the randomized, controlled trials and observational studies comparing the same treatments for these conditions. For each treatment, the magnitudes of the effects in the various observational studies were combined by the Mantel-Haenszel or weighted analysis-of-variance procedure and then compared with the combined magnitude of the effects in the randomized, controlled trials that evaluated the same treatment. RESULTS: There were 136 reports about 19 diverse treatments, such as calcium-channel-blocker therapy for coronary artery disease, appendectomy, and interventions for subfertility. In most cases, the estimates of the treatment effects from observational studies and randomized, controlled trials were similar. In only 2 of the 19 analyses of treatment effects did the combined magnitude of the effect in observational studies lie outside the 95 percent confidence interval for the combined magnitude in the randomized, controlled trials. CONCLUSIONS: We found little evidence that estimates of treatment effects in observational studies reported after 1984 are either consistently larger than or qualitatively different from those obtained in randomized, controlled trials. PMID- 10861325 TI - Randomized, controlled trials, observational studies, and the hierarchy of research designs. AB - BACKGROUND: In the hierarchy of research designs, the results of randomized, controlled trials are considered to be evidence of the highest grade, whereas observational studies are viewed as having less validity because they reportedly overestimate treatment effects. We used published meta-analyses to identify randomized clinical trials and observational studies that examined the same clinical topics. We then compared the results of the original reports according to the type of research design. METHODS: A search of the Medline data base for articles published in five major medical journals from 1991 to 1995 identified meta-analyses of randomized, controlled trials and meta-analyses of either cohort or case-control studies that assessed the same intervention. For each of five topics, summary estimates and 95 percent confidence intervals were calculated on the basis of data from the individual randomized, controlled trials and the individual observational studies. RESULTS: For the five clinical topics and 99 reports evaluated, the average results of the observational studies were remarkably similar to those of the randomized, controlled trials. For example, analysis of 13 randomized, controlled trials of the effectiveness of bacille Calmette-Guerin vaccine in preventing active tuberculosis yielded a relative risk of 0.49 (95 percent confidence interval, 0.34 to 0.70) among vaccinated patients, as compared with an odds ratio of 0.50 (95 percent confidence interval, 0.39 to 0.65) from 10 case-control studies. In addition, the range of the point estimates for the effect of vaccination was wider for the randomized, controlled trials (0.20 to 1.56) than for the observational studies (0.17 to 0.84). CONCLUSIONS: The results of well-designed observational studies (with either a cohort or a case-control design) do not systematically overestimate the magnitude of the effects of treatment as compared with those in randomized, controlled trials on the same topic. PMID- 10861326 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 19-2000. A 17-year-old boy with obstructive jaundice. PMID- 10861327 TI - The pharmaceutical industry--to whom is it accountable? PMID- 10861328 TI - The acute chest syndrome of sickle cell disease. PMID- 10861329 TI - Randomized trials or observational tribulations? PMID- 10861330 TI - Blockade of adhesion of sickle cells to endothelium by monoclonal antibodies. PMID- 10861331 TI - Should physicians prescribe religious activities? PMID- 10861332 TI - Towards optimizing the timing of the pre-exercise meal. AB - The purpose of this study was to compare the effect a 6-hr versus 3-hr prefeeding regimen on exercise performance. The subjects were 8 active women (21.4 +/- 0.9 years, 60.4 +/- 2.4 kg, 19.9 +/- 1.3% body fat, and 165.6 +/- 2.1 cm). All women completed 2 exercise trials (separated by 3-6 d) on a treadmill where they ran at moderate intensity for 30 min with 30-s sprints at 5-min intervals, followed directly by increasing incrementally the grade until volitional fatigue was achieved. The exercise trials were performed 3 hr and 6 hr after consuming 40 +/- 3 kJ/kg meal. Time to exhaustion was 0.75 min shorter (p =.0001) for the 6-H trials compared to the 3-H trials. There were no significant differences in submaximal or peak oxygen uptake, heart rate, or rating of perceived exertion (p >.05). The 6-H trials compared to the 3-H trials resulted in.05 lower RERs (p =.0002), and a 2 mmol lower blood lactate at exhaustion (p =.012). Blood glucose levels and cortisol responses to exercise were similar between trials (p >.05). However, both resting and post exercise insulin levels were lower during 6-H trials. It was concluded that performance of moderate- to high-intensity exercise lasting 35-40 min is improved by consuming a moderately-high carbohydrate, low fat, low protein meal 3-hr before exercise compared to a similar meal consumed 6 hr prior to exercise. Thus, athletes should not skip meals before competition or training sessions. PMID- 10861333 TI - Diet and short term plasma lipoprotein-lipid changes after exercise in trained Men. AB - To test the effect of diet on the short-term lipid response to exercise, fourteen moderately trained (VO2max: 50.2 +/- 6.7 ml/kg/min), healthy men (mean age: 28 +/ 4 years) were alternately fed a high fat (60 +/- 6.7% fat) and a high carbohydrate (63 +/- 3.2% carbohydrate) isoenergetic diet for 2 weeks in a randomized crossover design. During the last 4 days of the treatments, fasting total cholesterol, triglyceride, HDL-cholesterol, and HDL3-cholesterol were measured the day before, and again immediately, 24 hr, and 48 hr after exercise (4190 kJ, 70% VO2max). LDL-cholesterol and HDL2-cholesterol were calculated. Lipid concentrations were adjusted for plasma volume changes after exercise. A 2 (diet) x 4 (time) ANOVA with repeated measures revealed no significant interaction between the diet and exercise treatments. Furthermore, diet alone did not influence lipid concentrations in these trained men. Exercise resulted in an increase in HDL-C (10.7%) and HDL3-C (8.5%) concentrations and a concomitant fall in triglyceride (-25%) and total cholesterol (-3.5%). Thus, we conclude that diet composition does not affect the short-term changes in blood lipids and lipoproteins that accompany a single session of aerobic exercise in moderately trained men. PMID- 10861334 TI - Behavioral, psychological, and physical characteristics of female athletes with subclinical eating disorders. AB - The purpose of this study was to delineate and further define the behavioral, psychological, and physical characteristics of female athletes with subclinical eating disorders. Subjects consisted of 24 athletes with subclinical eating disorders (SCED) and 24 control athletes. Group classification was determined by scores on the Eating Disorder Inventory (EDI), the Body Shape Questionnaire (BSQ), and a symptom checklist for eating disorders (EDI-SC). Characteristics representative of the female athletes with subclinical eating disorders were derived from an extensive health and dieting history questionnaire and an in depth interview (the Eating Disorder Examination). Energy intake and expenditure (kcal/d) were estimated using 7-day weighed food records and activity logs. The characteristics most common in the female athletes with subclinical eating disorders included: (a) preoccupation with food, energy intake, and body weight; (b) distorted body image and body weight dissatisfaction; (c) undue influence of body weight on self-evaluation; (d) intense fear of gaining weight even though at or slightly below ( approximately 5%) normal weight; (e) attempts to lose weight using one or more pathogenic weight control methods; (g) food intake governed by self-hatred upon breaking a rule; (h) absence of medical disorder to explain energy restriction, weight loss, or maintenance of low body weight; and (i) menstrual dysfunction. Awareness of these characteristics may aid in more timely identification and treatment of female athletes with disordered eating patterns and, perhaps, prevent the development of more serious, clinical eating disorders. PMID- 10861335 TI - No effect of heavy resistance training and creatine supplementation on blood lipids. AB - In order to examine the effects of heavy resistance training and the influence of creatine supplementation on nonperformance measures of health status, 19 healthy resistance-trained men were matched and then randomly assigned in a double-blind fashion to either a creatine (n = 10) or placebo (n = 9) group. Periodized heavy resistance training was performed 3-4 times per week for 12 weeks. During the first week of training, creatine subjects consumed 25 g creatine monohydrate per day, while the placebo group ingested an equal number of placebo capsules. Five grams of supplement per day was consumed for the remainder of the study. Body composition, fasting serum creatinine, lipoproteins and triglycerides, and reported changes in body function were determined prior to and after 12 weeks of training and supplementation. After training, significant increases in body mass and fat-free mass were greater in creatine (5.2 and 4.3 kg, respectively) than placebo (3.0 and 2.1 kg, respectively) subjects. There was no change in percent body fat. Dietary energy and macronutrient distribution was not significantly different during Weeks 1 and 12. Serum creatinine was significantly elevated in creatine subjects after 1 (11.6%) and 12 weeks (13.8%); however, values were within normal limits for healthy men. There were no effects of training or supplementation on serum total cholesterol, HDL-cholesterol, LDL-cholesterol, or triglycerides. In healthy men, a 12-week heavy resistance training program, with or without creatine supplementation, did not significantly influence serum lipid profiles, subjective reports of body functioning, or serum creatinine concentrations. PMID- 10861336 TI - Sub-regional tissue morphometry in male athletes and controls using dual X-Ray absorptiometry (DXA). AB - Athletes have traditionally been evaluated for body composition by percent fat, percent muscle, and somatotype. Since the late 1980s, dual X-ray absorptiometry (DXA) has offered total and regional body composition of bone mineral content (BMC), lean tissue and fat, but studies involving athletes are rare (11) and have not included regional tissue distribution. In the present study, DXA was used to compare a total of 121 male subjects belonging to 9 different athletic groups and controls. ANOVA showed total tissue percent BMC, lean tissue, and fat were significantly different between the various athletic groups (p <.001). Regional differences in tissue distribution between different athletic groups affect BMC and lean tissue (p <.001), but not fat (p >.05). However, athletes of the leanest groups had different fat distribution to that of nonexercising controls (p <.01). It appears that fat distribution is nonspecific in its response to exercise, while lean and BMC distributions show highly specific adaptations to specific sports. PMID- 10861337 TI - The effects of choline supplementation on physical performance. AB - It has been reported that plasma choline levels decrease following certain types of strenuous exercise. Preliminary findings also suggest that a drop in plasma choline may limit physical performance, while choline supplementation may delay fatigue during prolonged efforts. A double-blind crossover design was used to determine the relationship between plasma choline and performance during and after 4 hr of strenuous exercise. Volunteers (N = 14) received either a placebo or treatment beverage (8.425 g choline citrate) prior to and midway through a 4 hr load carriage treadmill exercise (3% grade at 5.6 km/h 3 20 km) carrying a total load of 34.1 kg. Following the treadmill test, run time-to-exhaustion and squat tests were performed, and perceived exertion, plasma choline, glycerophosphocholine, and phosphatidylcholine were measured. Plasma choline levels increased 128% after the run-to-exhaustion with the choline supplemented beverage but remained unchanged with the placebo beverage. No significant effects were seen with choline supplementation on any outcome performance measure. Consequently, soldiers conditioned to carry heavy loads over long distances do not deplete plasma choline as a result of a prolonged exhaustive exercise under a placebo beverage, nor do they benefit from choline supplementation to delay fatigue under the same conditions. PMID- 10861338 TI - L-Carnitine supplementation combined with aerobic training does not promote weight loss in moderately obese women. AB - L-Carnitine (L-C) transports fatty acids into mitochondria for oxidation and is marketed as a weight loss supplement. In a double-blind investigation to test the weight loss efficacy of L-C, 36 moderately overweight premenopausal women were pair matched on Body Mass Index (BMI) and randomly assigned to two groups (N = 18). For 8 weeks the L-C group ingested 2 g twice daily of L-C, while the placebo (P) group ingested the same amount of lactose. All subjects walked for 30 min (60 70% maximum heart rate) 4 days/week. Body composition, resting energy expenditure (REE) and substrate utilization were estimated before and after treatment. For the subjects who completed the study (15 P, 13 L-C), no significant changes in mean total body mass (TBM), fat mass FM, and resting lipid utilization occurred over time, nor were there any significant differences between groups for any variable. Conversely REE increased significantly for all subjects, but no between group differences existed. Five of the L-C group experienced nausea or diarrhea and consequently did not complete the study. Eight weeks of L-C ingestion and walking did not significantly alter the TBM or FM of overweight women, thereby casting doubt on the efficacy of L-C supplementation for weight loss. PMID- 10861339 TI - The effects of Tribulus terrestris on body composition and exercise performance in resistance-trained males. AB - The purpose of this study was to determine the effects of the herbal preparation Tribulus terrestris (tribulus) on body composition and exercise performance in resistance-trained males. Fifteen subjects were randomly assigned to a placebo or tribulus (3.21 mg per kg body weight daily) group. Body weight, body composition, maximal strength, dietary intake, and mood states were determined before and after an 8-week exercise (periodized resistance training) and supplementation period. There were no changes in body weight, percentage fat, total body water, dietary intake, or mood states in either group. Muscle endurance (determined by the maximal number of repetitions at 100-200% of body weight) increased for the bench and leg press exercises in the placebo group (p <.05; bench press +/-28.4%, leg press +/-28.6%), while the tribulus group experienced an increase in leg press strength only (bench press +/-3.1%, not significant; leg press +/-28.6%, p <.05). Supplementation with tribulus does not enhance body composition or exercise performance in resistance-trained males. PMID- 10861340 TI - Evaluation of ischemic injury during liver procurement from non-heart-beating donors. AB - The aim of this study was to assess liver viability after different periods of cardiac arrest and the predictive value of two markers of ischemia-reperfusion injury. METHODS: A pig liver transplantation model of non-heart-beating donors was studied. Four donor groups were designed; three groups were submitted to different periods of cardiac arrest (20, 30 and 40 min), and the fourth group served as the control group (without cardiac arrest). In the non-heart-beating donor groups, normothermic recirculation was established 30 min prior to total body cooling. Aminotransferase, alpha-glutathione-S-transferase, and hyaluronic acid determinations as well as liver biopsies, were serially performed. RESULTS: Although hepatocellular function could be preserved after 40 min of cardiac arrest, histological lesions at 5 days were considered irreversible due to the presence of a necrotic biliary tract. An overall significant relationship was found between the time period of cardiac arrest (20, 30 or 40 min) and the levels of hyaluronic acid (p = 0.004) or alpha-glutathione-S-transferase (p = 0.01) obtained during liver procurement and transplantation. CONCLUSIONS: The period of cardiac arrest is the determinant factor of liver viability after liver transplantation from non-heart-beating donors. As early markers of endothelial or hepatocellular damage, hyaluronic acid or alpha-glutathione-S-transferase levels may help to evaluate the ischemic injury of a potential donor. PMID- 10861341 TI - Intestinal ischaemia-reperfusion increases plasma amylin concentration in rats. AB - Intestinal ischaemia-reperfusion and hyperamylinaemia are both associated with severe acute pancreatitis. The aim of this study was to examine the relationship between intestinal-ischaemia reperfusion and plasma amylin in an experimental model. Wistar rats (n = 24, 400-450 g) were divided into three groups: (1) a sham (S)-operated group (n = 7) that underwent laparotomy and isolation (without clamping) of the superior mesenteric artery, (2) an ischaemia-reperfusion (IR) group (n = 7) that had clamping of the superior mesenteric artery for 60 min followed by 15 min reperfusion, and (3) a control (C) group (n = 10) that underwent no surgery. Amylin was significantly elevated in the IR group (median 39 pM, range 30-44) compared with the S group (19 pM, range 15-45; Mann-Whitney U, p < 0.05) and the C group (24 pM, range 15-55; p < 0. 01). Insulin was significantly elevated in the IR group (2,060 pM, range 1,000-4,650) compared with the S group (558 pM, range 424-2, 020; p < 0.01). There was a significant positive correlation between amylin and insulin (R = 0.82, F = 46.6, p < 0.0001), but not between amylin and glucose or insulin and glucose. Intestinal histology was consistent with an ischaemia-reperfusion injury, whereas pancreatic histology was normal. The unique finding that plasma amylin concentration is increased with intestinal ischaemia-reperfusion injury warrants further investigation. PMID- 10861342 TI - Effect of nitric oxide on postoperative adhesion formation. AB - Peritoneal adhesions continue to be a significant cause of postoperative complications. The purpose of the present study was to investigate the effect of nitric oxide in preventing postoperative adhesion formation in rats. Three randomized groups of Sprague-Dawley rats were subjected to a standardized lesion by cecal abrasion and parietal peritoneal defect. 0.9% NaCl (control, group 1), L arginine (300 mg/kg, group 2) and Nomega-nitro arginine methyl ester (L-NAME; 25 mg/kg, group 3) were administered intraperitoneally before abdominal closure and during 3 consecutive days after surgery. Two weeks after surgery, a relaparotomy was performed and the extent of adhesion formation was determined. In groups 1 and 3 heavy adhesions were detected. In the L-arginine group, adhesion formation was significantly less than in the other groups (p < 0.05). This study showed that L-arginine reduced adhesion formation. PMID- 10861343 TI - Laparoscopic modified taylor procedure in the treatment of duodenal ulcer: technique and outcome after 5-year follow-up. AB - BACKGROUND: Recently, gastric stapling with posterior truncal vagotomy has been performed by laparoscopic surgery, as an alternative to highly selective vagotomy (HSV) and the Taylor procedure for the treatment of chronic duodenal ulcer. AIM: To investigate, after a mean 5-year follow-up, the effect of the stapling modified laparoscopic Taylor procedure, on gastric secretion, emptying and reflux as well as clinical parameters and recurrence rates in patients treated for duodenal ulcer. METHODS: 16 patients, aged 38-66 years, were treated from January 1993 to January 1996 (median 60.5 months), by the laparoscopic stapling-modified Taylor procedure, using the Endo-GIA stapler device. Assessment of the results of gastric acid secretion, solid and liquid gastric emptying, enterogastric reflux, endoscopic findings and clinical parameters, using the Visick grading, was performed. RESULTS: Endoscopy found healing ulcer in 15 patients. One patient showed signs of chronic ulcerative disease without gastritis or pyloric stenosis indicative of progressive ulcerative diathesis and was classified as Visick III. 14 patients were classified as Visick I and 1 as II. The enterogastric reflux index ranged from 0 to 26%, basal and peak acid output were 1.4 +/- 0.6 and 11.7 +/- 6.1 mmol H(+)/h, respectively. The half-emptying times of the solid and liquid meal were 82 +/- 7 and 16 +/- 6 min, respectively. These results are likely to be similar to those obtained from series of patients who underwent HSV or Taylor procedure and are closed to those from healthy controls. CONCLUSIONS: The laparoscopic modified Taylor procedure, using the Endo-GIA stapler device allows a more rapid, technically easier and radical performance of the operation with excellent long- term results and should be included in the armamentarium of the treatment of chronic duodenal ulcer. PMID- 10861344 TI - Expression of the extracellular matrix proteins collagen I, collagen III and fibronectin and matrix metalloproteinase-1 and -13 in the skin of patients with inguinal hernia. AB - Although abnormal collagen metabolism has been ascribed an important role in the high recurrence rates after surgical hernia repair, knowledge on tissue sampled in the region affected by inguinal hernias is poor. In the present study, we determined collagen type I and type III in the skin of adult patients with indirect and direct inguinal hernias by both immunohistochemistry and Western blot analysis. In addition, we quantified the immunohistochemical expression of fibronectin and matrix metalloproteinase (MMP)-1 and -13. The results indicated that the ratio of collagen type I/III was significantly decreased in the skin of patients with either indirect (n = 9) or direct hernia (n = 7), with a concomitant increase in collagen type III (p < 0.001 vs. controls, n = 7, without affection of the inguinal region). There was no significant difference between patients with indirect and direct hernia (p > 0.05). MMP-13 was not expressed in any of the skin samples investigated, whereas MMP-1 was found in the epidermis. Fibronectin was predominantly detected at the epidermal-dermal junction. MMP-1, MMP-13 and fibronectin levels were significantly different between patients and controls (p > 0. 05). We conclude that in contrast to the unchanged expression of fibronectin and MMP-1 and MMP-13, the decreased ratios of collagen tpye I/III with the basically increased amount of collagen type III could be of significant importance for the pathophysiology of hernias. The specific ratio collagen I/III probably reflects the altered structural integrity and mechanical stability of the connective tissue in both indirect and direct hernias. Moreover, our findings stress that hernias should be regarded as the manifestation of a systemic disease in the inguinal region with a genetic background, explaining the high recurrence rates after repeated suture repair, as well as the usefulness of surgical meshes in this clinical setting. PMID- 10861345 TI - Development of a model for measurement of adhesion strength of fibrin sealant to human tissue. AB - It is crucial for the surgeon to know the physical properties of a surgical sealant. Current test methods of fibrin sealant involving animal testing or in vitro testing of sealant using artificial substrates have little clinical relevance. Most of these test methods also lack accuracy and reproducibility. A new model was developed for testing strength and in vitro adhesion of fibrin sealant to vital human tissue using fresh vein leftover from coronary artery bypass grafting. The vein leftover was cut into samples and fastened in a tensiometer linked to a computer. Patient-derived fibrin sealant (0.1 ml) was applied to the tissue, and the surfaces of the tissue samples were held together for 5 min, and then automatically pulled apart by the tensiometer. Data were generated in a load cell and recorded and analysed by the computer. The reproducibility for the adhesion strength was 6.6%, adhesion energy 9.8%, and elongation at break 8.4%. The method has been considered ethical and has good reproducibility. The method can be used for standardised measurements and comparison of different types of fibrin sealant without the sacrifice of animals. PMID- 10861346 TI - Gastrectomy in the rat using two modifications of esophagojejunal anastomosis. general status, local histological changes and relationships to bone density. AB - Gastrectomy (GX) was carried out in the male rat according to the Longmire and the Roux-en-Y procedure. The focus of the postoperative investigations was to evaluate the influence of post-GX morphological changes occurring at the site of two types of end-to-end esophagojejunal anastomosis (with and without invagination), in particular food intake, body weight gain, food efficiency, hematocrit and bone density. GX failed to alter food intake, fasting blood glucose, alpha-amino nitrogen, or free fatty acids, but led to uniformly decreased body weight, food efficiency and serum gastrin, and increased serum osteocalcin, indicating high turnover osteopenia. However, irrespective of the type of (digestive tract) reconstruction (Longmire or Roux-en-Y), the invagination anastomosis was associated with lower mortality, fewer complications, less early postoperative weight loss, less intensive tissue changes at the anastomotic site, and improvement of bone density and hematocrit. Bivariate and multivariate regression analysis revealed that bone density was negatively influenced by epithelial hyperplasia of the anastomotic tissue, while hematocrit was positively influenced by bone density. In contrast, food intake appeared to have no influence. It was concluded that (1) the histological status of the esophagointestinal anastomosis varies depending on the surgical technique applied and (2) the type of anatomical reconstruction of the digestive tract (Longmire vs. Roux-en-Y) and food intake may be of minor importance for the bone and hematological status of GX rats. Future investigations are justified to clarify whether esophagojejunal proinflammatory tissue factors may contribute to the GX-mediated damage of bone mineral and bone marrow, thereby leading to low body weight. PMID- 10861347 TI - A view on the science: physical anthropology at the millennium editor. PMID- 10861348 TI - Sequence analysis of bacterial DNA in the colon and stomach of the Tyrolean Iceman. AB - The male human body found in an Alpine glacier on September 19, 1991 ("Tyrolean Iceman") has, for the first time in history, given scientists a chance to perform detailed anatomical, histological, and molecular investigations on the organs of a person from the Neolithic Age (5350-5100 B.P.). In the present study, tissue samples aseptically taken from the stomach and the colon of the mummy were utilized for DNA extraction, and the DNA was PCR-amplified, using primer pairs designed to bind to fragments of the 16s ribosomal RNA gene (16s rDNA) of a broad range of bacteria. The PCR products were cloned in plasmid vectors, and the recombinant clones (amplicons) were sequenced. The sequence data were finally used for scanning data libraries containing the corresponding sequences of present-day bacteria, to infer the putative ecophysiology of the ancient ones. The same procedure was repeated on some fragments of grass from the clothing found near the corpse. These fragments were taken as a control of the microbiological situation of the glacier. The results show that the flora of the Iceman's stomach is entirely composed of Burkholderia pickettii, an organism commonly found in aquatic habitats. The colon, on the other hand, contains several members of the fecal flora of humans, such as Clostridium perfringens, C. ghonii, C. sordellii, Eubacterium tenue, and Bacteroides sp. The Iceman's colon, however, was found to contain, rather unexpectedly, also some members of the genus Vibrio. The results are discussed in light of what is known about the preservation of microbial DNA at the Iceman's site and of previous parasitological studies performed on the Iceman himself and on human coprolites. PMID- 10861349 TI - Beta-globin gene cluster haplotypes in two North American indigenous populations. AB - Haplotypes derived from five polymorphic restriction sites were determined in 50 Carrier-Sekani and 70 Mvskoke chromosomes, and the results were integrated with those previously obtained for 11 South American Indian populations. Eleven haplotypes were identified in the Mvskokes, while five were observed in the Carrier-Sekani. As in South American natives, haplotype 2 (+----) and 6 (-++ -+) were the most prevalent among the Mvskoke (46% and 30%, respectively). In the Carrier-Sekani, haplotype 2 was also the most common, but haplotype 5 (-+ -++) was somewhat more frequent (18%) than 6 (12%). High heterozygosities, as well as genetic differentiation, were observed among these two North American and two other South American groups (Mapuche and Xavante). They could be due to non Indian admixture in the Mvskoke and Mapuche, but the findings in the other two populations require some other type of explanation. PMID- 10861350 TI - Microsatellite variation in Central Africa: an analysis of intrapopulational and interpopulational genetic diversity. AB - As a part of a research project on molecular variation in Central Africa, we have analyzed 10 microsatellites (CD4, CSFO, D3S1358, D18S51, D21S11, F13A1, FES, TH01, TPOX, and VWA) in the Bamileke and Ewondo from Cameroon and the Sanga and Mbenzele Pygmies from the Central African Republic (a total of 390 chromosomes). A statistically significant trend towards heterozygote deficiency was detected in the Mbenzele Pygmies. This was established through the use of powerful exact tests for the Hardy-Weinberg equilibrium. A certain degree of isolation and a small effective size may explain this finding. However, the lack of any substantial reduction in allelic diversity in the Mbenzele does not support the possibility that this group has a smaller effective size in evolutionary terms. A possible explanation based on ethnographic studies suggests that the gene flow from non-Pygmies to Pygmies could have been interrupted only in relatively recent times. The analysis of association between genotypes at pairs of independent loci indicates that the level of subheterogeneity is markedly lower in the Bamileke than in other sampled populations. This may be explained by the combined effect of larger population size, more rigid respect of clanic exogamy, and higher matrimonial mobility of the Bamileke. Finally, we have analyzed interpopulational relationships among our sampled populations and other Central African populations. The results are consistent with a previous study of protein loci (Spedini et al. 1999), which suggests the recent history of the Bamileke and Ewondo has led them to aquire a substantial genetic similarity. Furthermore, the Mbenzele Pygmies diverge from Biaka Pygmies, despite their common origin and geographical proximity. This is probably due to the differentiating effect of genetic drift, which is enhanced by the small effective size of Pygmy populations. PMID- 10861351 TI - Brachycephalization in Japan has ceased. AB - Somatometric data are presented which show that the rapid brachycephalization in Japan has recently ceased. The causes of brachycephalization are investigated in relation to the secular change in height. Increases in head breadth have been the main cause of brachycephalization, and its pattern of secular change is very similar to that in height. Associations between head breadth, height, and year of birth were examined by partial correlation coefficients and through a comparison of students and the general population. Brachycephalization is thought to result from increases in the growth rate for head breadth caused by improvements in nutritional levels, as seen in increases in height. Increases in height over the last 100 years have been accompanied by brachycephalization in Japanese and Koreans, but by debrachycephalization in many European populations. Increases in lateral growth in Asian heads may be related to the facial flatness which is characteristic to northern Mongoloid populations. PMID- 10861352 TI - Age-dependent cortical bone loss in women from 18th and early 19th century London. AB - Age-dependent cortical bone loss was investigated in an earlier British population. The study sample comprised female skeletons from the 18th/19th century crypt at Christ Church, Spitalfields, London. Bone loss was monitored using metacarpal radiogrammetry. Age at death was known exactly from coffin plates. Results indicated that peak cortical thickness was less than in modern subjects. Continuing periosteal apposition was evident throughout adulthood, and the rate of increase in metacarpal diameter resembled that in modern subjects. Bone loss from the endosteal surface was evident from the fifth decade onwards, and this outstripped the rate of subperiosteal gain so that there was a net loss of cortical bone with age. Cortical bone loss occurred at a similar rate to that in modern subjects. In contrast to modern populations, there was no evidence that loss of cortical bone was associated with increased propensity to fracture. The present results, together with those previously published for a British medieval skeletal assemblage, suggest that patterns of cortical bone loss in women have remained unchanged over at least the last millennium in Britain. Given the great changes in lifestyle which have occurred during this period, this suggests that lifestyle factors may be rather less important than is sometimes asserted in influencing the severity of osteoporosis, at least as far as loss of cortical bone is concerned. PMID- 10861353 TI - Comparison of population structure in Ohio's late archaic and late prehistoric periods. AB - Previous studies of population structure among prehistoric groups in the Ohio valley region have shown that hunting-gathering populations exhibited a different structure than horticultural populations. Among both Late Archaic hunter gatherers and Late Prehistoric horticulturists, covariance structures for cranial metrics were found to be homogenous within the populations, but the Late Archaic subpopulations showed little differentiation while the Late Prehistoric subpopulations exhibited a marked differentiation. Biodistance based on cranial discrete trait frequency showed similar patterns, but in the Late Archaic discrete trait distance was associated significantly with the geographical distance separating populations. The present investigation is an extension of the previous studies increasing the Late Prehistoric sample (n = 8 samples and n = 341 individuals) and using the Harpending-Ward model, modified for use with multivariate quantitative data, to estimate the effects of differential gene flow and the amount of differentiation within populations. Results of the present analyses indicate that differentiation among subpopulations, measured by minimum F(ST), was greater in the Late Prehistoric compared to the Late Archaic period. However, for both periods the minimum F(ST) is comparable to values found for historic native populations of the northeast woodlands. Analysis of differential gene flow in the Late Archaic period indicates that geographically peripheral populations were affected more by external gene flow than more central populations. Late Prehistoric populations exhibit a very complex pattern of differential gene flow. We discuss the latter pattern in terms of proposed culture change in the Late Prehistoric period of Ohio. PMID- 10861354 TI - Do big females have big pelves? AB - Previous studies have shown that maternal stature is a correlate of both pelvic size and reproductive efficiency. This study addresses the issue of body size and obstetric advantage. The relationship between pelvic size and three nonpelvic measures of body size is determined for females and males. The skeletal sample consists of blacks, whites, and Native Americans. The variables include 28 measures of the pelvis, length and head diameter of the femur, and clavicular length. The coefficient of multiple determination (CMD) is computed for each pelvic measure using multiple regression, with the three nonpelvic measures serving as the independent variables. Partial correlation coefficients are also calculated between each pelvic and nonpelvic variable, while controlling for the other two nonpelvic variables. The results show that all CMDs in females and all but one CMD in males are "low," i.e., below 33%. The sexes are nonsignificantly different in their CMDs for 22 of the 28 pelvic variables; of the six variables that are significantly different, five are of the midplane. The sexes are also broadly comparable in their partial correlations. The results are explained as follows. First, the concordance between the sexes in the relationship between pelvic size and nonpelvic measures of body size is due to their genetic similarity for homologous structures. Second, as pelvic size is at the minimum at the midplane, the sexual differences in CMDs are the result of selection with respect to obstetrics. Third, four explanations for the low CMDs are discussed: 1) lack of populationally or racially specific analysis; 2) nonlinear relationship between pelvic size and nonpelvic measures of body size; 3) combination of negative allometric selection between newborn body weight and maternal stature and weight with positive selection for maternal pelvic size; and 4) hormonally induced increase in pelvic capacity during parturition. PMID- 10861355 TI - Linear enamel hypoplasia in gibbons (Hylobates lar carpenteri). AB - This study describes the expression of linear enamel hypoplasia (LEH), a sensitive dental indicator of physiological stress, in Thailand gibbons (Hylobates lar carpenteri). Previous studies of enamel hypoplasia in hominoids have focused on great apes, with little attention given to the expression of this stress indicator in gibbons. In that gibbons differ from both monkeys and great apes in numerous life history features, LEH expression in gibbons might be expected to show significant differences from both. In this study, 92 gibbon specimens from two sites in Thailand were compared with several samples of monkeys and great apes in their expression of LEH. The intertooth distribution of LEH in gibbons was compared to that of chimpanzees and rhesus monkeys. Gibbon populations from both sites exhibit LEH frequencies intermediate between those of the monkey samples, in which LEH prevalence is usually low, and those of the great ape samples, in which LEH prevalence is high. Gibbons differ significantly from monkeys, but not great apes, in the number of individuals whose teeth record multiple stress events. Multiple episodes of stress are rarely recorded in the teeth of monkeys, while multiple stress events occur with higher frequency in gibbons and great apes. Taxonomic variation in the duration of crown formation, the prominence and spacing of perikymata on dental crowns, life history features, and/or experience of physiological stress may explain these patterns. The intertooth distribution of LEH in gibbons is, for different reasons, unlike that of either chimpanzees or rhesus monkeys. The mandibular canines of gibbons have significantly more LEH than any of their other teeth. Aspects of crown morphology, perikymata prominence/spacing, enamel thickness, and crown formation spans are potential causes of taxonomic variation in the intertooth distribution of LEH. PMID- 10861356 TI - Niche separation in Varecia variegata rubra and Eulemur fulvus albifrons: I. Interspecific patterns. AB - Niche separation was documented in a year-long study of Varecia variegata rubra and Eulemur fulvus albifrons on the Masoala Peninsula, Madagascar. Feeding trees were measured, and diet, forest height, and forest site were recorded at 5-min time points on focal animals. For time point data, multivariate and bivariate analysis of frequencies was employed to examine how niche dimensions vary between species according to sex, season, and reproductive stage. V. v. rubra feeds in larger trees than E. f. albifrons. V. v. rubra has a diet consisting mainly of fruit, whereas E. f. lbifrons has a more varied diet. V. v. ubra ranges mainly above 15 m in tree crowns, whereas E. f. albifrons ranges mainly below 15 m in a wide array of forest sites. Both species are largely frugivorous, but they harvest fruit in different-sized trees, in different quantities, and in different forest strata. Niche partitioning varies in tandem with seasonal shifts in climate and food availability and with reproductive stages. Seasonal shifts in forest site and forest height use are largely attributed to species-specific tactics for behavioral thermoregulation and predator avoidance. The diet of E. f. albifrons is diverse whether examined by season or reproductive stage. However, females of both species diversify their diets with more low-fiber protein than males during gestation, lactation, and the hot seasons. This pattern is most pronounced for V. v. rubra females and may be directly attributed to high energetic investment in reproduction. These results suggest that niche partitioning may be driven more by the energetic requirements of reproductive females than males. PMID- 10861358 TI - Calbindin D28k-immunoreactive afferent nerve endings in the laryngeal mucosa. AB - The distribution of the calbindin D28k in the laryngeal sensory structures was studied by immunohistochemistry, immunoelectronmicroscopy, and double immunofluorescence with calretinin-immunoreactivity. Moreover, origin of the nerve endings were observed using retrograde tracer, fast blue. Immunoreactivity for calbindin D28k was found in the various types of nerve endings in the larynx, namely, laminar nerve endings, nerve endings associated with the taste buds, intraepithelial nerve endings, and endocrine cells. The laminar endings with calbindin D28k-immunoreactivity were observed in the subepithelial connective tissue. In some endings, terminals were expanded. The laminar endings were also observed in the perichondrium of the epiglottic cartilage. In the epiglottic and arytenoid epithelia, thick nerve fibers with calbindin D28k-immunoreactivity ascending to taste buds and intragemmal nerve fibers were also observed. Within the epithelial layer, intraepithelial free nerve endings with calbindin D28k immunoreactivity were observed. Furthermore, diffuse endocrine cells were observed within the laryngeal epithelium. By immunoelectron microscopy, immunoreaction products in the endings mentioned above were localized in the cytoplasm of the axon terminals and nerve fibers which contained with numerous mitochondria. Out of the 100 laminar endings, 18 endings were immunopositive for both calbindin D28k and calretinin, 33 were positive for calbindin D28k but negative for calretinin, and 49 were positive for only calretinin in the double immunofluorescence microscopy. The nerve fibers associated with the taste buds and the free nerve endings, which immunostained for calbindin D28k, were not stained with antibody against calretinin. After injection of the fast blue in the laryngeal mucosa, fast blue-labeled cells were mainly observed in the nodose ganglia. Of the total number of labeled cell in the nodose and dorsal root ganglia at the level C1 to Th2, 65.1% occurred in nodose ganglia (572/879, n = 6). In the nodose ganglia, 79.7% of labeled cells (456/572) were immunoreacted for calbindin D28k. The distribution of calbindin D28k-immunoreactivity may be differnt from that of calretinin. It is suggested that calbindin D28k have regulatory role on intracellular calcium concentration in the laryngeal sensory corpuscles. PMID- 10861359 TI - Cardiac looping in the chick embryo: a morphological review with special reference to terminological and biomechanical aspects of the looping process. AB - Understanding early cardiac morphogenesis, especially the process of cardiac looping, is of fundamental interest for diverse biomedical disciplines. During the past few years, remarkable progress has been made in identifying molecular signaling cascades involved in the control of cardiac looping. Given the rapid accumulation of new data on genetic, molecular, and cellular aspects of early cardiac morphogenesis, and given the widespread interest in cardiac looping, it seems worth reviewing those aspects of the looping process that have received less attention during the past few years. These are terminological problems, the "gross" morphological aspects, and the biomechanical concepts of cardiac looping. With respect to terminology, emphasis is given to the unperceived fact that different viewpoints exist as to which part of the normal sequence of morphogenetic events should be called cardiac looping. In a short-term version, which is preferred by developmental biologists, cardiac looping is also called dextral- or rightward-looping. Dextral-looping comprises only those morphogenetic events leading to the transformation of the originally straight heart tube into a c-shaped loop, whose convexity is normally directed toward the right of the body. Cardioembryologists, however, regard cardiac looping merely as a long-term process that may continue until the subdivisions of the heart tube and vessel primordia have approximately reached their definitive topographical relationship to each other. Among cardioembryologists, therefore, three other definitions are used. Taking into account the existence of four different definitions of the term cardiac looping will prevent some confusion in communications on early cardiac morphogenesis. With respect to the gross morphological aspects, emphasis is given to the following points. First, the straight heart tube does not consist of all future regions of the mature heart but only of the primordia of the apical trabeculated regions of the future right and left ventricles, and possibly a part of the primitive conus (outflow tract). The remaining part of the primitive conus and the primordia of the great arteries (truncus arteriosus), the inflow of both ventricles, the primitive atria, and the sinus venosus only appear during looping at the arterial (truncus arteriosus) and venous pole (other primordia). Second, dextral-looping is not simply a bending of the straight heart tube toward the right of the body, as it has frequently been misinterpreted. It results from three different morphogenetic events: (a) bending of the primitive ventricular region of the straight heart tube toward its original ventral side; (b) rotation or torsion of the bending ventricular region around a craniocaudal axis to the right of the body, so that the original ventral side of the heart tube finally forms the right convex curvature and the original dorsal side forms the left concave curvature of the c-shaped heart loop; (c) displacement of the primitive conus to the right of the body by kinking with respect to the arterial pole. Third, dextral-looping does not bring the subdivisions of the heart tube and vessel primordia approximately into their definitive topographical relationship to each other. This is achieved by the morphogenetic events following dextral looping. This review seeks to bring together data from the diverse disciplines working on the developing heart. PMID- 10861360 TI - The peripapillary glia of the optic nerve head in the chicken retina. AB - Abstract The eye of reptiles and birds is characterized by an avascular retina and a vascular convolute called conus papillaris in reptiles and pecten oculi in birds which arises from the papilla nervi optici (PNO) or optic nerve head into the vitreous. At least in birds, this central part of the retina is the site of a heterogeneous population of glial cells. Muller cells reside in the retina, astrocytes in the optic nerve, and pecteneal glial cells in the pecten. The latter are developmentally related to the pigment epithelial cells. In addition to these established types of cells, there is a population of glial cells lining the base of the pecten oculi. In the present study, we investigated both the morphology and the development of these glial cells of the PNO in a series of chicken embryos. These cells were called peripapillary glial cells. They were characterized by their morphology and by their spatiotemporal expression of antigens typical of glial cells (intermediate filaments and glutamine synthetase). They reside at the border between the retina and the optic nerve and at the innermost border of the ventricular cleft representing transitional forms among Muller cells, astrocytes, and pigment epithelial cells. The developmental data suggest a migration of the perikarya of the peripapillary glia in vitread direction, which may coincide with that of the pecteneal glia. Whereas the pecteneal glial cells differentiate morphologically from E16 on, the peripapillary glia retain characteristics of radial glia by spanning the distance from the vitreous to the ventricular cleft. Blood vessels only occurred in the optic nerve head and the pecten oculi. No capillaries were found in the retinal tissue, beyond the peripapillary glia, leading us to suggest that these cells may play a role in demarcating the outer limit of vascularization. The functional properties of these cells are unknown but were discussed to include prevention of vessel growth into the avascular retina and/or axonal guidance during development. PMID- 10861361 TI - Transmission electron microscope study of bacterial morphotypes on the anterior dorsal surface of human tongues. AB - The human tongue has been the subject of many cytological and histological studies. When a literature search disclosed no reports of the ultrastructure of the morphotypes of bacteria residing on the tongue's surface, a transmission electron microscope study of ultrathin sections of bacteria obtained by scraping eight human tongues was undertaken. The scrapings from the anterior dorsal tongue surfaces, processed conventionally for electron microscope study, revealed 33-35 different bacterial morphotypes. Several of the morphotypes were unique to a tongue. Morphotype differences were also related to donor characteristics such as smoking, tongue site, location in centrifuge pellet, diet, and medications. The predominant morphotypes were Gram-positive cocci. These preliminary findings suggest that the microbiota of the human tongue and variations in that microbiota, related to physical condition, lifestyle, medications, and dietary preferences, merit more attention from anatomists. PMID- 10861362 TI - Atrial development in the human heart: an immunohistochemical study with emphasis on the role of mesenchymal tissues. AB - The development of the atrial chambers in the human heart was investigated immunohistochemically using a set of previously described antibodies. This set included the monoclonal antibody 249-9G9, which enabled us to discriminate the endocardial cushion-derived mesenchymal tissues from those derived from extracardiac splanchnic mesoderm, and a monoclonal antibody recognizing the B isoform of creatine kinase, which allowed us to distinguish the right atrial myocardium from the left. The expression patterns obtained with these antibodies, combined with additional histological information derived from the serial sections, permitted us to describe in detail the morphogenetic events involved in the development of the primary atrial septum (septum primum) and the pulmonary vein in human embryos from Carnegie stage 14 onward. The level of expression of creatine kinase B (CK-B) was found to be consistently higher in the left atrial myocardium than in the right, with a sharp boundary between high and low expression located between the primary septum and the left venous valve indicating that the primary septum is part of the left atrial gene-expression domain. This expression pattern of CK-B is reminiscent of that of the homeobox gene Pitx2, which has recently been shown to be important for atrial septation in the mouse. This study also demonstrates a poorly appreciated role of the dorsal mesocardium in cardiac development. From the earliest stage investigated onward, the mesenchyme of the dorsal mesocardium protrudes into the dorsal wall of the primary atrial segment. This dorsal mesenchymal protrusion is continuous with a mesenchymal cap on the leading edge of the primary atrial septum. Neither the mesenchymal tissues of the dorsal protrusion nor the mesenchymal cap on the edge of the primary septum expressed the endocardial tissue antigen recognized by 249 9G9 at any of the stages investigated. The developing pulmonary vein uses the dorsal mesocardium as a conduit to reach the primary atrial segment. Initially, the pulmonary pit, which will becomes the portal of entry for the pulmonary vein, is located along the midline, flanked by two myocardial ridges. As development progresses, tissue remodeling results in the incorporation of the portal of entry of the pulmonary vein in left atrial myocardium, which is recognized because of its high level of creatine. Closure of the primary atrial foramen by the primary atrial septum occurs as a consequence of the fusion of these mesenchymal structures. PMID- 10861363 TI - Chronology of the appearance of beta, A, and alpha mitochondria-rich cells in the gill epithelium during ontogenesis of the brown trout (Salmo trutta). AB - Three types of mitochondria-rich (MR) cells, the alpha, beta, and accessory cells, are observed in the gill epithelium of juvenile and adult freshwater teleosts. In addition to numerous mitochondria, their cytoplasm contains a network of membranous tubules, the tubular system, connected to the laterobasal plasma membrane. Because they are believed to play a role in ionic regulation, it is of interest to examine the order of appearance and the ultrastructural characteristics of such cells during the embryogenesis and larval life of the brown trout. Gills of embryos and fry maintained in freshwater were thus removed at different stages and prepared for transmission and scanning electron microscopic examination. One week before hatching, cells resembling the beta cells of juvenile and adult teleosts appeared first among the epithelial cells located at the base of the filaments in the gills of the brown trout larva. In addition to their tubular system, they contained numerous and large apical structures seemingly originating from the Golgi apparatus. At approximately hatching time, small pear-shaped cells were seen to be closely apposed to the lateral side of the beta cells; they were usually devoid of apical structures and were considered to be accessory cells. After yolk sac resorption, additional cells, the alpha cells, were present along the lamellae. In contrast to the beta cells, they only exhibited poorly developed apical structures. The possible role of these three types of MR cells in osmoregulation during fish development is discussed. PMID- 10861364 TI - Histo-physiology of the scent-marking glands of the penile pad, anal pouch, and the forefoot in the aardwolf (Proteles cristatus). AB - The scentmarking glands of the anal pouch, penile pad, and the forefoot of the aardwolf (Proteles cristatus) were studied by histological, histochemical, immunohistochemical methods, and by electron microscopy. The morphological observations are correlated with eco-ethological aspects of this nocturnal animal. In all studied regions there was a superficial layer of holocrine sebaceous glands and a deeper layer of apocrine scent glands; these two types of glands apparently function in concert. Only in the forefoot were additional tubular glands, resembling eccrine sweat glands found, which may improve the frictional capacities of the paw, while apocrine and holocrine glands serve scent marking functions of the forefoot. Penile pad and anal pouch are exclusively scent marking organs. The secretion modus of the apocrine glands is both via exocytosis and apocrine mechanism. Homogeneous apical, secretory granules, which contain glycoproteinaceous material, represent evidence for exocytosis. In the anal pouch, additional variably sized granules contain endogenous pigments which are probably responsible for the brownish coloration of the secretory product of the male animals. Variable heights of the glandular cells, frequent apical tall protrusions as well as pinched-off pieces of cytoplasm in the glandular tubules support the concept of an apocrine secretion in the scent glands. The immunohistochemical staining pattern of actin points to the involvement of actin filaments in the pinching-off process of the apical cell protrusion, which does not contain any cell organelles. The variable actin staining patterns suggest a dynamic process during which actin filaments form a ring or sheet at the basis of the pinching-off bleb. Proliferative and apoptotic phenomena show no preference for active and inactive glandular cells suggesting that replacement of cells occurs independently of the functional status of the glands. PMID- 10861365 TI - Differential scanning calorimetry, biochemical, and biomechanical analysis of human skin from individuals with diabetes mellitus. AB - The aim of this work was to compare biochemical, two-dimensional biomechanical and calorimetric parameters of diabetic skin vs. control skin. Skin specimens taken from the palms and backs of the hands of aged persons with non-insulin dependent diabetes mellitus (NIDDM) and of controls (CO) were compared (age range 68-85 years). Only skin specimens from individuals with diabetes mellitus (DM) showed an increased fluorescence specific for the formation of advanced glycation end-products (AGEs) and the presence of tissue AGEs, such as N(e) (Carboxymethyl)lysine (CML). Differential scanning calorimetry (DSC) revealed an elevation of the heat flow per unit mass during collagen denaturation in diabetic skin samples. However, the temperatures of the heat flow maximum and the onset of the phase transformation were not uniformly altered. Young's moduli were found to be increased in diabetic skin and correlated with AGE-fluorescence and tissue AGEs. The ratio between the Young's moduli, which defines a measure for the degree of anisotropy, was higher for dorsal skins from hands. In dorsal skin specimens from diabetic subjects the degree of anisotropy was more pronounced than in healthy controls. In general, neither of the measured parameters showed any correlation with age. However, E(1) moduli were clearly associated with the duration of diabetes. PMID- 10861366 TI - Calbindin D28k and parvalbumin immunoreactivity in the rabbit superior colliculus: an anatomical study. AB - The expression pattern of two calcium binding proteins (CaBP), calbindin D28k (CB) and parvalbumin (PV), in the superior colliculus (SC) of the adult rabbit, as well as the morphology of the immunoreactive cells were examined. The study was performed on 12 rabbits. Coronal sections from postmortem SC were analyzed by light microscopy, and drawings of CaBP-labeled cells were obtained using a drawing tube. No previous information is available on either the CB/PV expression or the morphology of CB/PV positive cells in the SC of the adult rabbit. Therefore, in this study we show that CB neurons and neuropil form three main tiers: the first located within the stratum zonale (SZ) and the upper part of the stratum griseum superficiale (SGS), the second located within the stratum griseum intermedium (SGI), and the third, located within the medial and central areas of the stratum griseum profundum (SGP). In contrast to this layer labeling, almost no CB-positivity is found within the other collicular layers. On the other hand, the densest concentration of PV labeled cells and terminals is found within a single dense tier that spanned the ventral part of the startum griseum superficiale (SGS) and the dorsal part of the stratum opticum (SO). Anti-PV neurons are also scattered through the deeper layers below the dense tier. In contrast, almost no anti-PV labeled neurons or neuropil are found within the stratum zonale (SZ) and upper SGS. This distribution represents a new pattern of sublamination in the SC of this species. All the previously described cell types in other mammals are observed in the rabbit SC: marginal cells, horizontal cells, pyriform cells, narrow-field vertical cells, wide-field vertical cells, and stellate/multipolar cells. Detailed drawings of all these cellular types are represented to show their complete morphology. The results of this study indicate that both CB and PV are present in a variety of neurons, which present a number of homologies between mammals, but have a different location and/or distribution, according to the different species. These findings are thus relevant to better understand the organisation of the SC in mammals. PMID- 10861367 TI - Neurocalcin-immunoreactive neurons in the mammalian dorsal root ganglia, including humans. AB - Neurocalcin (NC) is a recently characterized EF-hand calcium-binding protein present in a discrete population of sensory neurons and their peripheral mechanoreceptors, but its presence in peripheral nervous system neurons other than in the rat is still unknown. The present study was designed to investigate the occurrence of NC in the dorsal root ganglia (DRG) of several mammalian species (horse, buffalo, cow, sheep, pig, dog, and rat), including humans. DRG were fixed, embedded in paraffin, and processed for immunohistochemistry using a polyclonal antibody against NC. The size of the immunoreactive neurons was measured. In all species examined, NC immunoreactivity (IR) was restricted to neurons but the percentage, as well as the size of the immunoreactive neurons, varied among different species. As a rule, small neurons (diameter <20 microm) lack NC IR. In some species (pig, dog, buffalo, cow), only the largest neurons showed IR, whereas in others (sheep, horse, rat, and humans) they covered the entire range of neuron sizes. The pattern of immunostaining was cytoplasmic, although in some species (cow and buffalo), it formed a peripheral "ring." The present results demonstrate that mammalian DRG contain a subpopulation of NC positive neurons, which varies from one species to another. Based on the neuron size, the possible function of the NC-containing neurons is discussed. PMID- 10861368 TI - Anatomic genomics: systems of genes supporting the biology of systems. AB - This essay lays the groundwork for the concept that "anatomy" in the new millennium will be a subject that is increasingly based on understanding the parallel relationships between systems of genes on chromosomes and the structures defined by these genes. The concept of Anatomic Genomics is introduced in terms of systems of genes on chromosomes, which actually mirror the biology of anatomically defined systems. A case is made for the possibility that genomes may be structured in ways that make local but not necessarily global sense. In the new millennium, systems biologists have the opportunity to be the creators and purveyors of this new anatomy. PMID- 10861369 TI - A study of conformational stability of polyglycine and poly(L-alanine), and polyglycine/poly(L-alanine) blends in the solid state by (13)C cross polarization/magic angle spinning NMR. AB - 13C Cross-Polarization/Magic Angle Spinning nmr and T(1rhoH) experiments of polyglycine (PG), poly(L-alanine) (PLA), and PG/PLA blends prepared from dichloroacetic acid solution have been carried out, in order to elucidate the conformational stability of these polypeptides in the solid state. From these experimental results, it was clarified that the conformations of PG and PLA in their blends are strongly influenced by intermolecular hydrogen-bonding interactions that cause their miscibility at the molecular level. PMID- 10861370 TI - Smoluchowski dynamics of the vnd/NK-2 homeodomain from Drosophila melanogaster: second-order maximum correlation approximation. AB - The mode coupling diffusion theory is applied to the derivation of local dynamics in proteins in solution. The rotational dynamics of the bonds along the protein sequence are calculated and compared to the experimentally measured nmr (15)N spin-lattice relaxation time T(1), at 36.5, 60.8, and 81.1 MHz of the vnd/NK-2 homeodomain from Drosophila melanogaster. The starting point for the calculations is the experimental three-dimensional solution structure of the homeodomain determined by multidimensional nmr spectroscopy. The higher order mode-coupling computations are compared also with the recently published first-order approximation calculations. The more accurate calculations improve substantially the first-order ORZLD calculations and show that the role of the strength of the hydrodynamic interactions becomes crucial to fix the order of magnitude of the rotational dynanics for these very compact molecules characterized by partial screening of the internal atoms to water. However, the relative mobility of the bonds along the sequence and the differential fluctuations depend only weakly on the hydrodynamic strength but strongly on the geometry of the three-dimensional structure and on the statistics incorporated into the theory. Both rigid and fluctuating dynamic models are examined, with fluctuations evaluated using molecular dynamics simulations. The comparison with nmr data shows that mode coupling diffusion accounts for the T(1) relaxation pattern at low frequency where the rotational tumbling dominates. An important contribution of internal motions in the nanosecond time scale is seen at high frequencies and is discussed in terms of diffusive concepts. PMID- 10861371 TI - The three-dimensional structure of the 4:1 mithramycin:d(ACCCGGGT)(2) complex: evidence for an interaction between the E saccharides. AB - Mithramycin and chromomycin, two antitumor drugs, each having an identical aglycone and nearly identical disaccharide and trisaccharide side chains, have differing binding properties to a small oligonucleotide, d(ACCCGGGT)(2) (M. A. Keniry et al., Journal of Molecular Biology, 1993, Vol. 231, pp. 753-767). In order to understand the forces that induce four mithramycin molecules to bind to d(ACCCGGGT)(2) instead of two drug molecules in the case of chromomycin, the structure of the 4:2:1 mithramycin: Mg(2+):d(ACCCGGGT)(2) complex was investigated by (1)H-nmr and restrained molecular dynamics. The resulting three dimensional model showed that in order to accommodate the close approach of one neighboring mithramycin dimer, the inwardly directed CDE saccharide chain of the neighboring mithramycin dimer undergoes a conformational change such that the E saccharide no longer spans the minor groove but reorients so that the hydrophilic face of the E saccharides from the two dimers oppose each other. Two hydrogen bonds are formed between the hydroxyl groups of the two opposing E saccharide groups. The results are interpreted in terms of the differences in stereochemistry and functional group substitutions between mithramycin and chromomycin. A mithramycin dimer is able to self-associate on an oligonucleotide template because it has two hydroxyl groups on the same face of its terminal E saccharide. A chromomycin dimer is unable to self-associate because one of these hydroxyl groups is acetylated and the neighboring hydroxyl group has a stereochemistry that cannot permit close contact of the hydroxyl group with a neighbouring chromomycin dimer. PMID- 10861372 TI - The structure of gellan in dilute aqueous solution. AB - A full assignment of high-field nmr spectra of gellan was obtained in dilute aqueous solution by performing a series of selective one-dimensional nmr experiments. The observed nuclear Overhauser effects (NOEs) cannot be interpreted assuming that each sugar residue is intrinsically rigid and in a chair conformation. In fact, the rhamnose residue gives strong NOE contacts coherent only with an equilibrium involving both a chair as well as a boat (or a hemiboat) conformation. Molecular dynamic calculations performed on a heptamer with a central rhamnose support the above finding, and show a structure based on a very stiff single chain in which it is present a flipping of the rhamnose residue. At low temperatures (5-20 degrees C) in very dilute solutions (0.018 mg/mL) nmr spectra show a splitting of the resonance due to the methyl group of rhamnose residue, thus confirming the presence of a slow equilibrium among different conformers. PMID- 10861373 TI - Peptide-sandwiched protoporphyrin compounds mimicking hemoprotein structures in solution. AB - A series of covalently bound peptide-protoporphyrin-peptide compounds, also carrying naphthalene (N) to allow a photophysical investigation, were synthesized. Their general formula is P(nN)(2), where P refers to protoporphyrin IX, and n to the number of amino acids in the sequence Boc-Leu-Leu-Lys-(Ala)(x) Leu-Leu-Lys-OtBu of each backbone chain (x = 0-3; n = x + 6). Their structural features in methanol solution were investigated by ir and CD spectra, and by steady-state and time resolved fluorescence experiments as well. The ir spectra indicate that intramolecularly H-bonded conformations form, and CD data in both methanol and water-methanol mixture suggest the presence of alpha-helix structure. Quenching of excited naphthalene takes place by electronic energy transfer from singlet N* to P ground state. Fluorescence decays coupled with molecular mechanics calculations indicate that two conformers for each dimeric peptide are the major contributors to the observed phenomena. These conformers are characterized by a globular, protein-like structure, where the protoporphyrin resides in a central pocket, while the two N groups are externally situated. Of the four N linkages in the two conformers, three of them attain a very similar steric arrangement around the central P molecule, in terms of both center-to center distance and mutual orientation, while the fourth experiences a different steric disposition as compared to the others. Experimental photophysical parameters satisfactorily compare with those obtained by theoretical calculations, within the Forster mechanism for long-range energy transfer, only when the mutual orientation of the chromophores was also taken into account. This implies that interconversion among conformational substates of probes linkages is slow on the time scale of the energy transfer process. PMID- 10861374 TI - DNA and buffers: the hidden danger of complex formation. AB - The free solution electrophoretic mobility of DNA differs significantly in different buffers, suggesting that DNA-buffer interactions are present in certain buffer systems. Here, capillary and gel electrophoresis data are combined to show that the Tris ions in Tris-acetate-EDTA (TAE) buffers are associated with the DNA helix to approximately the same extent as sodium ions. The borate ions in Tris borate-EDTA (TBE) buffers interact with DNA to form highly charged DNA-borate complexes, which are stable both in free solution and in polyacrylamide gels. DNA borate complexes are not observed in agarose gels, because of the competition of the agarose gel fibers for the borate residues. The resulting agarose-borate complexes increase the negative charge of the agarose gel fibers, leading to an increased electroendosmotic flow of the solvent in agarose-TBE gels. The combined results indicate that the buffers in which DNA is studied cannot automatically be assumed to be innocuous. PMID- 10861375 TI - Solution properties of viilian, the exopolysaccharide from Lactococcus lactis subsp. cremoris SBT 0495. AB - The exopolysaccharide (EPS) "viilian" was isolated from a large-batch fermentation of Lactococcus lactis subsp. cremoris SBT 0495. After applying a newly developed purification procedure, pure viilian with a weight-averaged molar mass of 2.64 x 10(3) kg/mol was obtained in a yield of 0.6 g/L culture broth. The native EPS, as well as lower molar mass fractions obtained by sonication of the native polymer, were studied by capillary viscometry and size-exclusion chromatography (SEC) coupled to multiangle laser light scattering detection (MALLS). From the viscosity data at various ionic strengths, we extracted a Mark Houwink-Kuhn-Sakurada exponent a = 0.79, and a Smidsrod B value of 0.03. By application of the Hearst, Bohdanecky, and Odijk models for stiff polymer coils, in connection to the experimental viscosity data, we established the characteristic ratio to be C(infinity) = 44 and the intrinsic persistence length q(0) = 11.5 nm. The rms radii of gyration predicted from each of the models were in good agreement with the experimental radii (e.g., (1/2)(w) = 162 nm for native viilian in 0.2M NaNO(3)), as determined by SEC-MALLS. In addition, the Odijk model predicts correct ionic strength-linear charge density dependence of the rms radius of gyration. From the combined viscosity and SEC-MALLS experiments we concluded that, in dilute aqueous solutions, viilian behaves as an intermediately stiff, random coil polyelectrolyte system. PMID- 10861376 TI - Conformational choice at alpha,alpha-di-n-propylglycine residues: helical or fully extended structures? AB - The conformational analysis of peptides containing a single alpha, alpha-di-n propylglycine (Dpg) residue incorporated into valine-rich sequences has been undertaken in order to delineate the possible role of sequence effects in stabilizing fully extended (C(5)) or local helical conformations at this residue. The three peptides Boc-Val-Dpg-Val-OMe (3), Boc-Val-Val-Dpg-Val-OMe (4), Boc-Val Val-Dpg-Val-Val-OMe (5), have been studied by (1)H-nmr methods in chloroform (CDCl(3)) and dimethylsulfoxide (DMSO) solutions. Even in a relatively poorly solvating medium like CDCl(3), all the valine NH groups appear to be solvent exposed, suggesting an absence of folded beta-turn conformations. However, in both CDCl(3) and DMSO the Dpg NH groups in all the three peptides appear to behave like apparently solvent-inaccessible groups. In fully extended C(5) conformations, the proximity of the NH and CO groups of Dpg may preclude effective solvation due to a combination of stereoelectronic factors. Nuclear Overhauser effects provide support for the largely extended backbones. The crystal structure of peptide 3 reveals an extended conformation at Dpg (2) with straight phi = -176 degrees, psi = 180 degrees. A correlation between the crystallographically observed backbone conformation and solution nmr parameters in DMSO has been attempted using available data. Dpg residues placed in poor helix stabilizing environments may be expected to favor a local C(5) conformation. PMID- 10861377 TI - Static and dynamic light scattering from aggregating particles. AB - We use standard hydrodynamic and light scattering theories to calculate the total intensity and dynamic light scattering properties of random aggregates of spherical particles containing up to ten spheres. When the aggregates have dimensions comparable to the wavelength of light, intraaggregate interference effects can dramatically reduce the apparent size of the aggregates. These results could be significant in interpreting DNA condensation, protein polymerization, and other biomolecular aggregation reactions. PMID- 10861378 TI - The cyclic contryphan motif CPxXPXC, a robust scaffold potentially useful as an omega-conotoxin mimic. AB - Contryphan-R, from venom of the cone-shell Conus radiatus, represents a novel cyclic peptide scaffold onto which residues may be grafted to mimic unrelated protein surfaces. Three substitutions were made at the x and X positions of the disulfide-bridged motif CPxXPXC, where X and x represent any L- and D-handed residues, respectively, P represents proline or hydroxyproline, and C a half cystine. These substitutions were designed to mimic part of the pharmacophore of the unrelated globular polypeptide omega-conotoxin GVIA, which blocks N-type calcium channels. The structure of this engineered contryphan, YNK-contryphan-R ([D-Tyr4, Asn5, Lys7]contryphan-R), is shown to be similar to that of native contryphan-R (Pallaghy et al., Biochemistry, 1999, Vol. 38, pp. 13553-13559), confirming that the scaffold is robust with respect to the multiple substitutions. In particular, the alpha-beta bond vectors characterising the orientation of the side chains relative to the backbone are similar in contryphan R, YNK-contryphan-R, and omega-conotoxin GVIA, which is the required result for a scaffold-based approach to molecular design. The solution structure of YNK contryphan-R has an N-terminal, nonhydrogen-bonded, chain reversal centered on Hyp3-D-Trp4, and a C-terminal type I beta-turn. A minor form due to cis-trans isomerism of the Hyp2-Cys3 peptide bond is present in YNK-contryphan-R in a larger proportion than in contryphan-R. It is evident, particularly from the (3)J(HalphaHN) coupling constants, that YNK-contryphan-R is more flexible than contryphan-R, probably due to the absence in YNK-contryphan-R of the Pro-Trp packing present in the native molecule. Nevertheless, the structure confirms that cyclic peptide molecular designs can achieve the intended conformations. PMID- 10861379 TI - IR spectroscopy of isotope-labeled helical peptides: probing the effect of N acetylation on helix stability. AB - The effect of N-acetylation on the conformation of alanine-rich helical peptides is examined using isotope-edited Fourier transform infrared (FTIR) spectroscopy. A series of peptides with sequence AA(AAKAA)(3)AAY has been prepared; each peptide incorporates four (13)C-labeled alanines. These peptides have two amide I' bands in their FTIR spectra: one corresponding to the (12)C amino acids, and one assigned to the (13)C amino acids. The intensity and frequency of the (13)C amide I' band varies systematically with the position of the labels in the sequence and the presence or absence of an N-acetyl capping group. The intensity of the (13)C amide I' band correlates with helix stability at the labeled residues as predicted by thermodynamic models of the helix-coil transition. These results suggest that FTIR spectroscopy combined with specific isotope labeling can be used to dissect the conformation of helical peptides at the residue level. PMID- 10861380 TI - Conformational studies on a synthetic C-terminal fragment of the alpha subunit of G(S) proteins. AB - It has recently been reported that synthetic peptides corresponding to the C terminal sequence of G alpha, can be used to study the molecular mechanisms of interaction between this protein and G protein coupled receptors (Hamm et al., Science, 1988, Vol. 241, pp. 832-835). A conformational analysis on a 11 amino acids peptide from the G alpha(S) C-terminus, G alpha(S)(384-394) (H-QRMHLRQYELL OH), was performed by nmr spectroscopy and molecular modeling methods. Two dimensional nmr spectra, recorded in hexafluoroacetone/water, a mixture with structure stabilizing properties, showed an unusually high number of nuclear Overhauser effects, forming significative pattern to the drawing of a secondary structure. Conformations consistent with experimental NOE distances were obtained through molecular dynamics and energy minimization methods. These calculations yielded two stable conformers corresponding to an alpha-turn and a type III beta turn involving the last five C-terminal residues. Interestingly, the alpha-turn conformation was found to overlap with good agreement the crystallographic structure of the same fragment in the G alpha(S) protein. PMID- 10861381 TI - Direct visualization of changes in deacylated Na(+) gellan polymer morphology during the sol-gel transition. AB - Changes in gellan polymer morphology during the sol-gel transition were directly visualized by transmission electron microscopy and a model incorporating these changes and existing physical data is proposed. Our observations suggest that the most thermodynamically stable conformations of gellan polymers in solution, in the absence of added cations, are the double helix and double-helical duplexes. We have demonstrated two forms of lateral aggregation of gellan helices in the presence of Ca(2+) and K(+) ions. One type forms junction zones that lead to network formation and gelation, while the second type leads to the formation of isolated fibers of aggregated helices and inhibition of gelation. The proposed model of gellan gelation is based on these observations where thermoreversibility, gel strength, and endothermic transitions of gellan gels can be explained. PMID- 10861382 TI - How bradykinin alters the lipid membrane structure: a spin label comparative study with bradykinin fragments and other cations. AB - Electron spin resonance spectroscopy of several different spin labels was used to comparatively study the interaction of the cationic peptide hormone bradykinin (BK; Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg), and some BK fragments (des-Arg(9)-BK, des-Arg(1)-BK, and Arg-Pro-Pro-Gly-Phe or BK(1-5)), with anionic vesicles of dimyristoyl phosphatidylglycerol (DMPG). For temperatures above the lipid gel liquid crystal thermal transition (T(m) approximately 20 degrees C), membrane incorporated spin labels indicated that all peptides (total concentration of 10 mol % relative to lipid) interact with the bilayer, turning the membrane less fluid, both at its surface and center, suggesting a partial penetration of the peptides into the membrane core. However, in the lipid gel phase (t < T(m)), BK was found to display a much stronger interaction with the membrane, decreasing the bilayer fluidity. At temperatures around 15 degrees C the BK-DMPG system was found to present a hysteresis, evinced by the different electron spin resonance spectra yielded upon cooling and heating the sample. System reversibility was found at all other temperatures (0-45 degrees C). That effect could not be assigned to the BK higher concentration at the membrane surface, due to its higher net charge (2(+)) compared to the fragments (1(+)), because ten times more des-Arg(9)-BK (100 mol %) yielded opposite result. Further, that was found to be a result rather different from those elicited by the other cations tested: the monovalent Na(+), the divalent Zn(2+), and the peptide pentalysine. The data presented here are discussed in the light of the different BK and BK fragments biological activities. PMID- 10861383 TI - Time-lapse confocal reflection microscopy of collagen fibrillogenesis and extracellular matrix assembly in vitro. AB - The development of the next generation of biomaterials for restoration of tissues and organs (i.e., tissue engineering) requires a better understanding of the extracellular matrix (ECM) and its interaction with cells. Extracellular matrix is a macromolecular assembly of natural biopolymers including collagens, glycosaminoglycans (GAGs), proteoglycans (PGs), and glycoproteins. Interestingly, several ECM components have the ability to form three-dimensional (3D), supramolecular matrices (scaffolds) in vitro by a process of self-directed polymerization, "self-assembly". It has been shown previously that 3D matrices with distinct architectural and biological properties can be formed from either purified type I collagen or a complex mixture of interstitial ECM components derived from intestinal submucosa. Unfortunately, many of the imaging and analysis techniques available to study these matrices either are unable to provide insight into 3D preparations or demand efforts that are often prohibitory to observations of living, dynamic systems. This is the first report on the use of reflection imaging at rapid time intervals combined with laser-scanning confocal microscopy for analysis of structural properties and kinetics of collagen and ECM assembly in 3D. We compared time-lapse confocal reflection microscopy (TL-CRM) with a well-established spectrophotometric method for determining the self-assembly properties of both purified type I collagen and soluble interstitial ECM. While both TL-CRM and spectrophotometric techniques provided insight into the kinetics of the polymerization process, only TL-CRM allowed qualitative and quantitative evaluation of the structural parameters (e.g., fibril diameter) and 3D organization (e.g., fibril density) of component fibrils over time. Matrices formed from the complex mixture of soluble interstitial ECM components showed an increased rate of assembly, decreased opacity, decreased fibril diameter, and increased fibril density compared to that of purified type I collagen. These results suggested that the PG/GAG components of soluble interstitial ECM were affecting the polymerization of the component collagens. Therefore, the effects of purified and complex mixtures of PG/GAG components on the assembly properties of type I collagen and interstitial ECM were evaluated. The data confirmed that the presence of PG/GAG components altered the kinetics and the 3D fibril morphology of assembled matrices. In summary, TL CRM was demonstrated to be a new and useful technique for analysis of the 3D assembly properties of collagen and other natural biopolymers which requires no specimen fixation and/or staining. PMID- 10861384 TI - Resonance raman studies of heme structural differences in subunits of deoxy hemoglobin. AB - Low frequency resonance Raman (RR) spectra are reported for deoxy hemoglobin (Hb), its isolated subunits, its analogue bearing methine-deuterated hemes in all four subunits (Hb-d(4)), and the hybrids bearing the deuterated heme in only one type of subunit, which are [alpha(d4)beta(h4)](2) and [alpha(h4)beta(d4)](2). Analyzed collectively, the spectra reveal subunit-specific modes that conclusively document subtle differences in structure for the heme prosthetic groups in the two types of subunits within the intact tetramer. Not surprisingly, the most significant spectral differences are observed in the gamma(7) mode that has a major contribution from out of plane bending of the methine carbons, a distortion that is believed to relieve strain in the high-spin heme prosthetic groups. The results provide convincing evidence for the utility of selectively labeled hemoglobin hybrids in unraveling the separate subunit contributions to the RR spectra of Hb and its various derivatives and for thereby detecting slight structural differences in the subunits. PMID- 10861385 TI - Interactions of phosphatidylinositol 3-kinase Src homology 3 domain with its ligand peptide studied by absorption, circular dichroism, and UV resonance raman spectroscopies. AB - Absorption, circular dichroism (CD), and UV resonance Raman (UVRR) spectroscopies were applied to selectively examine the environmental and structural changes of Trp and Tyr residues in the phosphatidylinositol 3-kinase (PI3K) SH3 domain induced by ligand association. Comparison of the spectra of PI3K SH3 in the presence or absence of its ligand peptide RLP1 (RKLPPRPSK) indicated that RLP1 binding changed the environment of Trp55 of the SH3 to be more hydrophilic and its H bonding weaker and that of Tyr residues to be more hydrophobic. The D21N mutant (Asp21 --> Asn) of the SH3 yielded a UV CD distinct from that of the wild type, and its spectral changes induced by RLP1 binding were smaller and different from those of the wild type in absorption, CD, and UVRR spectra, suggesting that the mutation of conserved Asp21 affected the conformation of the ligand binding cleft and thus might lead to the decrease in the ligand affinity. These data provide direct evidence for the occurrence of environmental and structural changes of PI3K SH3 by the association of a ligand and the D21N mutation. PMID- 10861386 TI - Two-photon autofluorescence microscopy and spectroscopy of Antarctic fungus: new approach for studying effects of UV-B irradiation. AB - We combined two-photon fluorescence microscopy and spectroscopy to provide functional images of UV-B (280-315 nm) induced stress on an Antarctic fungus. Two photon excitation microscopy was used to characterize the distribution of autofluorescence inside the spore and the hyphae of the fungus. The imaging analysis clearly shows that the autofluorescence response of spores is higher than that of hyphae. The imaging analysis at different depths shows that, strikingly enough, the spore autofluorescence originates from the cell wall and membrane fluorophores. The spectroscopic results show moreover that the fluorescence spectra of spores are redshifted upon UV-B irradiation. Tentative identification of the chromophores involved in the autofluorescence response and their biological relevance are also discussed on the basis of a previous steady state fluorescence spectroscopic study performed on both whole spore suspension and organic-soluble extracts. PMID- 10861387 TI - Calcium-induced conformational change in fragment 1-86 of factor X. AB - Time-resolved fluorescence of the single tryptophan residue Trp41 in fragment 1 86 of factor X (FX F1-86) is studied using a time-correlated single photon counting technique with synchrotron radiation as the excitation source. Calcium ions are believed to induce a conformational change in the N-termini of the activated factor X and other vitamin K dependent proteins, which is accompanied by a decrease in fluorescence intensity. The titration with calcium yields a sigmoidal fluorescence titration curve with a transition midpoint concentration of 0.44 mM. The wavelength-dependent tryptophan fluorescence decays of the apo-FX F1-86 (in the absence of calcium) and Ca-FX F1-86 are characterized by conventional multiexponential analysis and fluorescence lifetime distribution analysis. In the absence of calcium there are three significant classes of fluorescence lifetimes (ns) that are nearly wavelength independent: 0.55 +/- 0.08 (component A), 2.6 +/- 0.1 (component B), and 5.3 +/- 0.3 (component C). However, their preexponential amplitudes vary with wavelength. The decay associated emission spectra of the individual components show that components B and C contribute over 85% to the total fluorescence for all examined wavelengths. However, in the presence of calcium, the analysis of the time-resolved fluorescence data of Ca-FX F1-86 yields four wavelength-independent lifetimes (ns) of 0.30 +/- 0.09 (component D), 0.65 +/- 0.10 (component A), 2.7 +/- 0.2 (component B), and 5.4 +/- 0.3 (component C). Calcium addition to the apo-FX F1 86 leads to a decrease in the fluorescence intensities of components B and C while their decay times remain unaffected. In Ca-FX F1-86 an additional component D arises that has a decay time of 0.30 ns and that contributes up to 35% to the total fluorescence intensity. A comparison with a previous investigation of prothrombin fragment 1 demonstrates the extensive structural and functional homology between the N termini of prothrombin and factor X(a). PMID- 10861388 TI - Raman optical activity characterization of native and molten globule states of equine lysozyme: comparison with hen lysozyme and bovine alpha-lactalbumin. AB - Vibrational Raman optical activity (ROA) spectra of the calcium-binding lysozyme from equine milk in native and nonnative states are measured and compared with those of the homologous proteins hen egg white lysozyme and bovine alpha lactalbumin. The ROA spectrum of holo equine lysozyme at pH 4.6 and 22 degrees C closely resembles that of hen lysozyme in regions sensitive to backbone and side chain conformations, indicating similarity of the overall secondary and tertiary structures. However, the intensity of a strong positive ROA band at approximately 1340 cm(-1), which is assigned to a hydrated form of alpha helix, is more similar to that in the ROA spectrum of bovine alpha-lactalbumin than hen lysozyme and may be associated with the greater flexibility and calcium-binding ability of equine lysozyme and bovine alpha-lactalbumin compared with hen lysozyme. In place of a strong sharp positive ROA band at approximately 1300 cm(-1) in hen lysozyme that is assigned to an alpha helix in a more hydrophobic environment, equine lysozyme shows a broader band centered at approximately 1305 cm(-1), which may reflect greater heterogeneity in some alpha-helical sequences. The ROA spectrum of apo equine lysozyme at pH 4.6 and 22 degrees C is almost identical to that of the holo protein, which indicates that loss of calcium has little influence on the backbone and side chain conformations, including the calcium-binding loop. From the similarity of their ROA spectra, the A state at pH 1.9 and both 2 and 22 degrees C and the apo form at pH 4.5 and 48 degrees C, which are partially folded denatured (molten globule or state A) forms of equine lysozyme, have similar structures that the ROA suggests contain much hydrated alpha helix. The A state of equine lysozyme is shown by these results to be more highly ordered than that of bovine alpha-lactalbumin, the ROA spectrum of which has more features characteristic of disordered states. A positive tryptophan ROA band at approximately 1551 cm(-1) in the native holo protein disappears in the A state, which is probably due to the presence of nonnative conformations of the tryptophans associated with a previously identified cluster of hydrophobic residues. PMID- 10861389 TI - Localization of carotenoids in plasma low-density lipoproteins studied by surface enhanced resonance Raman spectroscopy. AB - Surface-enhanced resonance Raman scattering (SERRS) spectra were measured for the beta-carotene and lycopene carotenoids present in low-density lipoproteins (LDLs), which were isolated from human plasma and adsorbed on roughened silver surfaces. The silver surface was modified by formation of a self-assembled monolayer (SAM) of carboxylate-terminated linear alkanethiols in order to simulate the LDL binding region of the cellular LDL receptor. Thiols of different chain length were used to produce SAMs of varying thicknesses. It was shown that carotenoids are not released from the LDL particle upon adsorption onto the bare and thiol modified silver surfaces. The SERRS studies indicated that beta carotene and lycopene were present in the shell of the LDL particle. The dependence of SERRS on the distance from the silver surface was different for beta-carotene and lycopene in LDL. This observation suggests that the two carotenoids are located in different places of the LDL particle. PMID- 10861390 TI - Investigation of stretching vibrations of glycosidic linkages in disaccharides and polysaccharides with use of IR spectra deconvolution. AB - The results are presented for the deconvolution of IR spectra of disaccharides and polysaccharides with alpha and beta configurations of the 1 --> 4 glycosidic linkage (maltose, cellobiose, amylose, and cellulose), as well as of their corresponding monosaccharides (alpha- and beta-D-glucose) in the 1200-920 cm(-1) frequency range. It is established that a characteristic of di- and polysaccharides with 1 --> 4 glycosidic linkage is the appearance of new absorption bands in the 1175-1140 cm(-1) spectral range, as opposed to the IR spectra of monosaccharides. This can be a spectroscopic manifestation of the glycosidic linkage formation. In the 1000-970 cm(-1) frequency range, absorption bands, which are not observed in the monomer spectrum, are separated as a result of the deconvolution of the IR spectra of cellobiose and cellulose. The number of bands in this range remains unchanged for maltose and amylose, as compared to the monomer spectra. It is shown that the application of the method of deconvolution leads to a considerable enhancement in the resolution of the absorption bands in the IR spectra of mono-, di-, and polysaccharides. PMID- 10861391 TI - Influence of scale-up on the quality of recombinant human growth hormone. AB - The aerobic fed-batch production of recombinant human growth hormone (rhGH) by Escherichia coli was studied. The goal was to determine the production and protein degradation pattern of this product during fed-batch cultivation and to what extent scale differences depend on the presence of a fed-batch glucose feed zone. Results of laboratory bench-scale, scale-down (SDR), and industrial pilot scale (3-m(3)) reactor production were compared. In addition to the parameters of product yield and quality, also cell yield, respiration, overflow, mixed acid fermentation, glucose concentration, and cell lysis were studied and compared. The results show that oxygen limitation following glucose overflow was the critical parameter and not the glucose overflow itself. This was verified by the pattern of byproduct formation where formate was the dominating factor and not acetic acid. A correlation between the accumulation of formate, the degree of heterogeneity, and cell lysis was also visualized when recombinant protein was expressed. The production pattern could be mimicked in the SDR reactor for all parameters, except for product quantity and quality, where 30% fewer rhGH degraded forms were present and where about 80% higher total yield was achieved, resulting in 10% greater accumulation of properly formed rhGH monomer. PMID- 10861392 TI - Metabolic shifts do not influence the glycosylation patterns of a recombinant fusion protein expressed in BHK cells. AB - BHK-21 cells expressing a human IgG-IL2 fusion protein, with potential application in tumor-targeted therapy, were grown under different nutrient conditions in a continuous system for a time period of 80 days. At very low glucose (< 0.5 mM) or glutamine (< 0. 2 mM) concentrations, a shift toward an energetically more efficient metabolism was observed. Cell-specific productivity was maintained under metabolically shifted growth conditions and at the same time an almost identical intracellular ATP content, obtained by in vivo (31)P NMR experiments, was observed. No significant differences in the oligosaccharide structures were detected from the IgG-IL2 fusion protein preparations obtained by growing cells under the different metabolic states. By using oligosaccharide mapping and MALDI/TOF-MS, only neutral diantennary oligosaccharides with or without core alpha1-6-linked fucose were detected that carried no, one or two beta1-4-linked galactose. Although the O-linked oligosaccharide structures that are present in the IL2 moiety of the protein were studied with less detail, the data obtained from the hydrazinolysis procedure point to the presence of the classical NeuAcalpha2-3Galbeta1-3GalNAc structure. Here, it is shown that under different defined cellular metabolic states, the quality of a recombinant product in terms of O- and N-linked oligosaccharides is stable, even after a prolonged cultivation period. Moreover, unaffected intracellular ATP levels under the different metabolic states were observed. PMID- 10861393 TI - Development of an ultra-high-temperature process for the enzymatic hydrolysis of lactose: II. Oligosaccharide formation by two thermostable beta-glycosidases. AB - During lactose conversion at 70 degrees C, when catalyzed by beta-glycosidases from the archea Sulfolobus solfataricus (SsbetaGly) and Pyrococcus furiosus (CelB), galactosyl transfer to acceptors other than water competes efficiently with complete hydrolysis of substrate. This process leads to transient formation of a range of new products, mainly disaccharides and trisaccharides, and shows a marked dependence on initial substrate concentration and lactose conversion. Oligosaccharides have been analyzed quantitatively by using capillary electrophoresis and high performance anion-exchange chromatography. At 270 g/L initial lactose, they accumulate at a maximum concentration of 86 g/L at 80% lactose conversion. With both enzymes, the molar ratio of trisaccharides to disaccharides is maximal at an early stage of reaction and decreases directly proportional to increasing substrate conversion. Overall, CelB produces about 6% more hydrolysis byproducts than SsbetaGly. However, the product spectrum of SsbetaGly is richer in trisaccharides, and this agrees with results obtained from the steady-state kinetics analyses of galactosyl transfer catalyzed by SsbetaGly and CelB. The major transgalactosylation products of SsbetaGly and CelB have been identified. They are beta-D-Galp-(1-->3)-Glc and beta-D-Galp-(1-->6)-Glc, and beta-D-Galp-(1-->3)-lactose and beta-D-Galp-(1-->6)-lactose, and their formation and degradation have been shown to be dependent upon lactose conversion. Both enzymes accumulate beta(1-->6)-linked glycosides, particularly allolactose, at a late stage of reaction. Because a high oligosaccharide concentration prevails until about 80% lactose conversion, thermostable beta-glycosidases are efficient for oligosaccharide production from lactose. Therefore, they prove to be stable and versatile catalysts for lactose utilization. PMID- 10861394 TI - Effect of variation of Klebsiella pneumoniae acetolactate synthase expression on metabolic flux redistribution in Escherichia coli. AB - Escherichia coli strains carrying the Bacillus subtilis acetolactate synthase (ALS) gene were previously shown to produce less acetate with higher ATP yields. Metabolic flux analysis was used to show that excess pyruvate was channeled into the less inhibitory product, acetoin. To further understand the role of intrinsic enzymatic properties and the effect of variations in enzyme levels in the alternation of metabolic fluxes, we constructed a chromosomal integrant of the Klebsiella pneumoniae ALS gene. The reported in vitro Michaelis-Menten constants (K(m)) for the Bacillus and the Klebsiella ALS are 13.0 mM and 8.0 mM, respectively. Furthermore, expression of the Klebsiella ALS is under the control of an inducible trp promoter system. Shake-flask experiments showed a linear induction response (the ALS activity changes from about 9 to 223 U/mg of protein when the inducer concentration [IAA] varied from 0 to 40 mg/L). Chemostat experiments showed a similar induction response. Interactions between the branched reactions catalyzed by the PFL, LDH, and the ALS enzymes at the pyruvate node were examined. The results indicate the importance of in vivo enzyme activities in the redistribution of metabolic fluxes. PMID- 10861395 TI - Statistical analysis of nonlinear parameter estimation for Monod biodegradation kinetics using bivariate data. AB - A nonlinear regression technique for estimating the Monod parameters describing biodegradation kinetics is presented and analyzed. Two model data sets were taken from a study of aerobic biodegradation of the polycyclic aromatic hydrocarbons (PAHs), naphthalene and 2-methylnaphthalene, as the growth-limiting substrates, where substrate and biomass concentrations were measured with time. For each PAH, the parameters estimated were: q(max), the maximum substrate utilization rate per unit biomass; K(S), the half-saturation coefficient; and Y, the stoichiometric yield coefficient. Estimating parameters when measurements have been made for two variables with different error structures requires a technique more rigorous than least squares regression. An optimization function is derived from the maximumlikelihood equation assuming an unknown, nondiagonal covariance matrix for the measured variables. Because the derivation is based on an assumption of normally distributed errors in the observations, the error structures of the regression variables were examined. Through residual analysis, the errors in the substrate concentration data were found to be distributed log-normally, demonstrating a need for log transformation of this variable. The covariance between ln C and X was found to be small but significantly nonzero at the 67% confidence level for NPH and at the 94% confidence level for 2MN. The nonlinear parameter estimation yielded unique values for q(max), K(S), and Y for naphthalene. Thus, despite the low concentrations of this sparingly soluble compound, the data contained sufficient information for parameter estimation. For 2-methylnaphthalene, the values of q(max) and K(S) could not be estimated uniquely; however, q(max)/K(S) was estimated. To assess the value of including the relatively imprecise biomass concentration data, the results from the bivariate method were compared with a univariate method using only the substrate concentration data. The results demonstrated that the bivariate data yielded a better confidence in the estimates and provided additional information about the model fit and model adequacy. The combination of the value of the bivariate data set and their nonzero covariance justifies the need for maximum likelihood estimation over the simpler nonlinear least squares regression. PMID- 10861396 TI - Cell damage of microcarrier cultures as a function of local energy dissipation created by a rapid extensional flow. AB - Microcarrier cultures of Chinese hamster ovary cells were subjected to a range of energy dissipations created by an abrupt contraction. These flow conditions can be characterized as a rapidly transient, extensional, and shear flow. Cell damage was measured using a lactate dehydrogenase assay. The laminar flow in the device was modeled using two commercial, computation fluid-dynamic codes: POLYFLOW and FLUENT. Cell damage was correlated to numerical values of energy dissipation. The magnitude of energy dissipation at which cell damage began to be detected, 10(4) ergs cm(-3) s(-1) (10(4) cm(2) s(-3)), is consistent with values of energy dissipation estimated in bioreactors operated under conditions which result in cell damage. This magnitude of energy dissipation is orders of magnitude lower than those values reported to cause damage to suspended animals cells which is also consistent with generally accepted experimental observations. Finally, an analysis and discussion of the presence and relative importance with re- spect to cell damage of shear vs. extensional flow is included. PMID- 10861397 TI - Effect of pH on the production of lipolyzed butter oil by a calf pregastric esterase immobilized in a hollow-fiber reactor: I. uniresponse kinetics. AB - A calf pregastric esterase immobilized in a hollow-fiber reactor was employed to hydrolyze milkfat, thereby producing a lipolyzed butteroil. The reaction kinetics can be modeled by a two-parameter model of the general Michaelis-Menten form based on a ping-pong bi-bi mechanism; the rate of enzyme deactivation can be modeled as a first-order reaction. The initial concentration of accessible glyceride bonds, [G](O), was estimated by complete saponification of the substrate butteroil as 2400 mM. An extra sum of squares test indicated that not only the parameters of the kinetic generalized Michaelis-Menten model, but also the deactivation-rate constant varied significantly with pH. The optimum pH, for lypolysis is near 6.0 at a temperature of 40 degrees C because at this pH the rate of deactivation of the esterase is minimized. PMID- 10861398 TI - Effect of anions on selective solubilization of zinc and copper in bacterial leaching of sulfide ores. AB - Bacterial leaching of sulfide ores using Thiobacillus ferrooxidans, Thiobacillus thiooxidans, or a combination of the two was studied at various concentrations of specific anions. Selective zinc and copper solubilization was obtained by inhibiting iron oxidation without affecting sulfur/sulfide oxidation. Phosphate reduced iron solubilization from a pyrite (FeS(2))-sphalerite (ZnS) mixture without significantly affecting zinc solubilization. Copper leaching from a chalcopyrite (CuFeS(2))-sphalerite mixture was stimulated by phosphate, whereas chloride accelerated zinc extraction. In a complex sulfide ore containing pyrite, chalcopyrite, and sphalerite, both phosphate and chloride reduced iron solubilization and increased copper extraction, whereas only chloride stimulated zinc extraction. Maximum leaching obtained was 100% zinc and 50% copper. Time course studies of copper and zinc solubilization suggest the possibility of selective metal recovery following treatment with specific anions. PMID- 10861399 TI - Limitations to the amplification and stability of human tissue-type plasminogen activator expression by Chinese hamster ovary cells. AB - Chinese hamster ovary cell production of recombinant tissue-type plasminogen activator (t-PA) was increased by amplification of cotransfected dihydrofolate reductase cDNA using stepwise adaptation to increasing methotrexate (MTX) concentrations. The highest producing clones were isolated at 5 microM MTX and yielded 26,000 U/10(6) cells/day t-PA (43 microgram/10(6) cells/day). Above 25 microM MTX, cell specific t-PA production rates became increasingly variable and the cDNA copynumbers decreased. No apparent correlation between the cell specific t-PA production rate and the growth rate was observed upon subcloning of the amplified cells. When MTX selection was removed, the t-PA production rate decreased up to tenfold within 40 days; this was accompanied by an up to 60% drop in cDNA copynumber. Subclones isolated after 108 days of culture in the absence of MTX were, on average, sixfold more stable than their parental cells. In culture without MTX, the maximum stable t-PA production rate obtained (over 250 days) was 7000 +/- 750 U/10(6) cells/day (approximately 12 microgram/10(6) cells/day), approximately threefold lower than the maximum unstable levels of production reached under selective pressure. Taken together, these results define a wide range of the highest t-PA expression rates obtained under MTX selection, for which stable expression without selection has not been reported. PMID- 10861400 TI - Study of drop and bubble sizes in a simulated mycelial fermentation broth of up to four phases. AB - The mean sizes and size distributions of air bubbles and viscous castor oil drops were studied in a salt-rich aqueous solution (medium), first separately, and then simultaneously as a three-phase system. The dispersion was created in a 150-mm diameter stirred tank equipped with a Rushton turbine, and the sizes were measured using an advanced video technique. Trichoderma harzianum biomass was added in some experiments to study the effect of a solid phase under unaerated and aerated conditions to give either three-or four-phase systems. In all cases, the different dispersed phases could be clearly seen. Such photoimages have never been obtained previously. For the three phases, air-oil-medium, aeration caused a drastic increase in Sauter mean drop diameter, which was greater than could be accounted for by the reduction in energy dissipation on aeration. Also, as in the unaerated case, larger drops were observed as the oil content increased. On the other hand, mean bubble sizes were significantly reduced with increasing oil phase up to 15% with bubbles inside many of the viscous drops. With the introduction of fungal biomass of increasing concentration (0.5 to 5 g L(-1)) under unaerated conditions, the Sauter mean drop diameter decreased. Finally, in the four-phase system (oil [10%]-medium-air-biomass) as found in many fermentations, all the phases (plus bubbles in drops) could clearly be seen and, as the biomass increased, a decrease in both the bubble and the drop mean diameters was found. The reduction in size of bubbles (and therefore increase in interfacial area) as the oil and bio- mass concentration increased provides a possible explanation as to why the addition of an oil phase has been reported to enhance oxygen transfer during many fermentations. PMID- 10861401 TI - Selectivity of Rhizomucor miehei lipase as affected by choice of cosubstrate system in ester modification reactions in organic media. AB - Fatty acid (FA) selectivity of immobilized Rhizomucor miehei lipase was determined for various cosubstrate systems for ester modification involving competing n-acyl-donor substrates of even-chain length (C4-C16; FA or their methyl esters, FAME) and either n-propanol or propyl acetate in hexane. Acyl chain-length optima were observed for C8 and C14/16 in all cases. Upon changing between cosubstrate systems of [FA + propanol] to [FAME + propanol] to [FAME + propyl acetate], there was a general shift in selectivity toward shorter-chain length FA (C4-C8). The greatest degree of reaction selectivity (based on ratios of selectivity constants) among the FA substrates was 3.1 for the [FA + propanol], 2.5 for the [FAME + propanol], and 1.4 for the [FAME + propyl acetate] cosubstrate systems. For esterification reactions between C6 FA and reactive members of a series of aliphatic and aromatic alcohols, the greatest degree of selectivity observed was 3.6. PMID- 10861402 TI - Room-temperature ionic liquids as replacements for organic solvents in multiphase bioprocess operations. AB - Organic solvents are widely used in a range of multiphase bioprocess operations including the liquid-liquid extraction of antibiotics and two-phase biotransformation reactions. There are, however, considerable problems associated with the safe handling of these solvents which relate to their toxic and flammable nature. In this work we have shown for the first time that room temperature ionic liquids, such as 1-butyl-3-methylimi- dazolium hexafluorophosphate, [bmim][PF(6)], can be successfully used in place of conventional solvents for the liquid-liquid extraction of erythromycin-A and for the Rhodococcus R312 catalyzed biotransformation of 1, 3-dicyanobenzene (1,3-DCB) in a liquid-liquid, two-phase system. Extraction of erythromycin with either butyl acetate or [bmim][PF(6)] showed that values of the equilibrium partition coefficient, K, up to 20-25 could be obtained for both extractants. The variation of K with the extraction pH was also similar in the pH range 5-9 though differed significantly at higher pH values. Biotransformation of 1,3-DCB in both water toluene and water-[bmim][PF(6)] systems showed similar profiles for the conversion of 1,3-DCB initially to 3-cyanobenzamide and then 3-cyanobenzoic acid. The initial rate of 3-cyanobenzamide production in the water-[bmim][PF(6)] system was somewhat lower, however, due to the reduced rate of 1,3-DCB mass transfer from the more viscous [bmim] [PF(6)] phase. It was also shown that the specific activity of the biocatalyst in the water-[bmim] [PF(6)] system was almost an order of magnitude greater than in the water-toluene system which suggests that the rate of 3-cyanobenzamide production was limited by substrate mass transfer rather than the activity of the biocatalyst. PMID- 10861403 TI - Enzymatic dehalogenation of gas phase substrates with haloalkane dehalogenase. AB - Haloalkane dehalogenase is an enzyme capable of catalyzing the conversion of short-chained (C(2)-C(8)) aliphatic halogenated hydrocarbons to a corresponding primary alcohol. Because of its broad substrate specificity for mono-, di-, and trisubstituted halogenated hydrocarbons and cofactor independence, haloalkane dehalogenases are attractive biocatalysts for gas-phase bioremediation of pollutant halogenated vapor emissions. A solid preparation of haloalkane dehalogenase from Rhodococcus rhodochrous was used to catalyze the dehalogenation reaction of 1-chlorobutane or 1,3-dichloropropane delivered in the gas phase. For optimal gas-phase dehalogenase activity, a relative humidity of 100%, a(w) = 1, was desired. With a 50% reduction in the vapor-phase hydration level, an 80% decrease in enzymatic activity was observed. The enzyme kinetics for the gas phase substrates obeyed an Arrhenius-"like" behavior and the solid haloalkane dehalogenase preparation was more thermally stable than its water-soluble equivalent. Triethylamine was added to the gaseous reaction environment in efforts to increase the rate of reaction. A tenfold increase in the dehalogenase activity for the vapor-phase substrates was observed with the addition of triethylamine. Triethylamine altered the electrostatic environment of haloalkane dehalogenase via a basic shift in local pH, thereby minimizing the effect of the pH-reducing reaction product on enzyme activity. Both organic phase and solid state buffers were used to confirm the activating role of the altered ionization state. PMID- 10861404 TI - A systematic approach to the validation of process control parameters for monoclonal antibody production in fed-batch culture of a murine myeloma. AB - A systematic approach to the validation of control ranges of control parameters for a cell culture process producing a monoclonal antibody is described. Specifically, the structure and functional activity of a monoclonal IgG1 antibody produced at the outer limits of numerical ranges of fed-batch culture control parameters such as pH and temperature were examined, with the aim of providing assurance that antibody produced under varying culture conditions was of consistent quality based on a carefully defined set of specifications. An experimental design was created using a half-fractional factorial design for fed batch culture incorporating half of the thirty two possible combinations of five selected control parameters at high and low levels. Statistical analysis of all data gathered from the study allowed an assessment of the effects of the process control parameters at either high or low outer limits on fed-batch culture response variables such as growth rate and specific antibody productivity. Measured values for the responses of growth rate and specific antibody productivity throughout this study ranged from 0.22-0.44 d(-1) and 6.4-32 microg monoclonal antibody/10(6) cells/d respectively. Analytical characterisation of monoclonal antibody purified from each fed-batch culture considered the purity, structure and biological activity of the glycoprotein. All antibody preparations were identical to each other and to the current antibody reference standard or control. Glycosylation analysis of certain samples from the study demonstrated that the distribution of glycoforms of the antibody was not affected by the varying process control conditions of the fed-batch cultures. PMID- 10861405 TI - Kinetic analysis of a tod-lux bacterial reporter for toluene degradation and trichloroethylene cometabolism. AB - Kinetics of toluene and trichloroethylene (TCE) degradation and bioluminescence from the bioreporter Pseudomonas putida B2 and TVA8 were investigated utilizing batch and continuous culture, respectively. Degradation was modeled using a Michaelis-Menten expression for the competition of two substrates for a single enzyme system, and bioluminescence was modeled assuming a luciferase enzyme saturational dependence on toluene as the inducer and growth substrate. During the batch experiments, bioluminescence increased at approximately 90 namp/min for initial toluene concentrations of 10 to 50 mg/L, but more slowly at higher toluene concentrations, suggesting maximum promoter induction at below 10 mg/L and toxic effects above 50 mg/L toluene. TCE degradation did not occur until toluene depletion, presumably due to competition between toluene and TCE for the toluene dioxygenase enzyme. During continuous culture, bioluminescence transiently increased, then gradually decreased in response to increasing step changes in toluene feed concentration. Bioluminescence in the CSTR appeared to be limited by growth substrate and/or inducer. PMID- 10861406 TI - Enhanced productivity during controlled proliferation of BHK cells in continuously perfused bioreactors. AB - A perfused cell-culture process was developed to investigate the stability of IRF 1-mediated proliferation control in BHK cells and to evaluate the efficacy of a novel promoter in these cells. The cell density of proliferation-controlled producer cells was effectively regulated for over 7 weeks in a microcarrier-based continuously perfused bioreactor. An IRF-1-inducible promoter was employed to express a heterodimeric IgG antibody as a relevant model protein. Basal expression levels were equivalent to that of a highly active viral promoter, while productivity increased up to sixfold during growth arrest. However, no stably expressing clone was isolated in this study. Protein expression decreased gradually with time and could not be induced further in subsequent growth repression cycles. The results demonstrate that the regulatory system is sufficiently stable to allow controlled growth in a continuous scalable reactor system and that productivity increases can be achieved in a proliferation controlled microcarrier culture. PMID- 10861407 TI - Framework for online optimization of recombinant protein expression in high-cell density Escherichia coli cultures using GFP-fusion monitoring. AB - A framework for the online optimization of protein induction using green fluorescent protein (GFP)-monitoring technology was developed for high-cell density cultivation of Escherichia coli. A simple and unstructured mathematical model was developed that described well the dynamics of cloned chloramphenicol acetyltransferase (CAT) production in E. coli JM105 was developed. A sequential quadratic programming (SQP) optimization algorithm was used to estimate model parameter values and to solve optimal open-loop control problems for piecewise control of inducer feed rates that maximize productivity. The optimal inducer feeding profile for an arabinose induction system was different from that of an isopropyl-beta-D-thiogalactopyranoside (IPTG) induction system. Also, model-based online parameter estimation and online optimization algorithms were developed to determine optimal inducer feeding rates for eventual use of a feedback signal from a GFP fluorescence probe (direct product monitoring with 95-minute time delay). Because the numerical algorithms required minimal processing time, the potential for product-based and model-based online optimal control methodology can be realized. PMID- 10861408 TI - Improved operational stability of peroxidases by coimmobilization with glucose oxidase. AB - The operational stability of peroxidases was considerably enhanced by generating hydrogen peroxide in situ from glucose and oxygen. For example, the total turnover number of microperoxidase-11 in the oxidation of thioanisole was increased sevenfold compared with that obtained with continuous addition of H(2)O(2). Coimmobilization of peroxidases with glucose oxidase into polyurethane foams afforded heterogeneous biocatalysts in which the hydrogen peroxide is formed inside the polymeric matrix from glucose and oxygen. The total turnover number of chloroperoxidase in the oxidation of thioanisole and cis-2-heptene was increased to new maxima of 250. 10(3) and 10. 10(3), respectively, upon coimmobilization with glucose oxidase. Soybean peroxidase, which normally shows only classical peroxidase activity, was transformed into an oxygen-transfer catalyst when coimmobilized with glucose oxidase. The combination catalyst mediated the enantioselective oxidation of thioanisole [50% ee (S)] with 210 catalyst turnovers. PMID- 10861409 TI - Growth and sporulation stoichiometry and kinetics of Coniothyrium minitans on agar media. AB - Coniothyrium minitans was cultivated on agar media with different concentrations of starch, urea, and trace elements. By means of elemental balances, the stoichiometry of growth and sporulation was established. C. minitans produced byproducts on all media, especially in the medium with high urea concentrations, where 30% of the starch was converted into byproducts. Simple empirical models were used to describe the kinetics of growth, sporulation, CO(2) production, and substrate consumption on all media. Total biomass and mycelium could be described reasonably well with the logistic law. Starch, urea, and oxygen consumption and CO(2) production could be described as a function of total biomass by the linear growth model of Pirt. There were almost no differences between media for the estimates of yield coefficients and maintenance coefficients. Only at high urea concentrations were maintenance coefficients much higher. Similar to substrate consumption and CO(2) production, the kinetics of sporulation could be described as a function of mycelium production with the linear-growth model. It is shown that sporulation of C. minitans is growth-associated. Based on kinetics, the process costs for producing spores are roughly calculated. In addition, it is shown that fermentor costs represent the majority of production costs. PMID- 10861410 TI - Oxygen-transfer strategy and its regulation effects in serine alkaline protease production by Bacillus licheniformis. AB - The effects of oxygen transfer on the production and product distribution in serine alkaline protease (SAP) fermentation by Bacillus licheniformis and oxygen transfer strategy in relation to the physiology of the bacilli were investigated on a defined medium with citric acid as sole carbon source in 3.5-dm(3) batch bioreactor systems. By forming a 3 x 3 matrix with the parameters air-inlet rates of Q(O)/V(R) = 0.2, 0.5, 1.0 vvm, and agitation rates of N = 150, 500, 750 min( 1), the effects of oxygen transfer were investigated at nine different conditions. The concentrations of the product SAP and by-products, i.e., neutral protease, alpha-amylase, amino acids, and organic acids, and SAP activities were determined throughout the bioprocess. Among the constant air-flow and agitation rate fermentations, Q(O)/V(R) = 0.5 vvm, N = 750 min(-1) oxygen-transfer conditions produced maximum SAP activity that was 500 U cm(-3), at t = 37 h. With the increase in Q(O)/V(R) and/or N, Damkohler number that is the oxygen-transfer limitation decreases; and the process passes from oxygen-transfer limited conditions to biochemical-reaction limited conditions. Further increase in SAP activity, A = 680 U cm(-3) was achieved by applying an oxygen-transfer strategy based on the analysis of the data obtained with the constant oxygen-transfer condition experiments, with a step increase in air-inlet rate, from Q(O)/V(R) = 0.2 to Q(O)/V(R) = 0.5 vvm at N = 750 min(-1) constant agitation rate at t = 24 h. Organic acids and amino acids that were excreted to the fermentation medium varied depending on the oxygen-transfer conditions. With the increase in oxygen transfer rate acetic acid concentration increased; contrarily, with the decrease in the oxygen-transfer rate the TCA-cycle organic acids alpha-ketoglutaric and succinic acids, and gluconic acid were excreted to the fermentation broth; nevertheless, the application of the oxygen-transfer strategy prevented the increase in acetic acid concentration between t = 35-38 h. Under all the oxygen transfer conditions, the amino acid having the highest concentration and the amino acid that was not excreted to the fermentation broth were lysine and asparagine, respectively; both of which belong to the aspartic acid-group amino acids. Further, this result indicates the requirement of the genetic regulation directed to the aspartic acid-group enzymes for the progress in SAP production in B. licheniformis. PMID- 10861411 TI - Determination of yeast glycogen content by individual cell spectroscopy using image analysis. AB - A rapid technique has been developed to determine the glycogen content of yeast on an individual cell basis using a combination of image analysis technology and staining of yeast cells with an I(2):KI solution. Changes in mean cellular glycogen content during alcoholic fermentation have been reported using this technique. The glycogen content of stored brewer's yeast is heterogeneous compared to freshly propagated yeast which have a more uniform distribution of glycogen. Analysis of the distribution of yeast glycogen during fermentation indicates that a fraction of yeast cells do not dissimilate glycogen. Therefore, conventional analysis of the mean glycogen content of yeast used to inoculate fermentations is of limited use, unless information regarding the proportion of cells which utilize glycogen is known. Analysis of the distribution of glycogen within a yeast population can serve as a useful indicator of yeast quality. PMID- 10861412 TI - Application of polymer-embedded proteins to fabrication of DNA array. AB - A plasma-polymerized film (PPF) of hexamethyldisiloxane [HMDS; (CH(3))(3)SiOSi(CH(3))(3)] was used to immobilize streptavidin on a glass substrate. Another layer of HMDS-PPF was also applied to the protein, which was first adsorbed to an underlayer of the same kind of film. As the result, the streptavidin was "embedded" between the two layers of HMDS, whereby biotinylated molecules could be efficiently captured. The second layer of approximately 30 to 45 A PPF was sufficient to allow the binding of biotinylated molecules, whereas thicknesses of >90 A significantly hindered the streptavidin-biotin interactions. Fluorescence analysis revealed that the absence of an HMDS plasma-polymer (HMDS PP) layer on either side of the streptavidin film resulted in a decrease in biotin binding. This immobilization technique was used to bind biotinylated oligonucleotides in sequence-specific DNA-DNA interactions. The hydrophobic properties of the plasma-polymerized HMDS thin film acted to minimize nonspecific DNA binding to the glass substrate. A DNA array was fabricated using this procedure and showed greatly decreased nonspecific DNA binding compared with a poly-L-lysine coated substrate. PMID- 10861413 TI - On-line control of fed-batch fermentation of dilute-acid hydrolyzates. AB - Dilute-acid hydrolyzates from lignocellulose are, to a varying degree, inhibitory to yeast. In the present work, dilute-acid hydrolyzates from spruce, birch, and forest residue, as well as synthetic model media, were fermented by Saccharomyces cerevisiae in fed-batch cultures. A control strategy based on on-line measurement of carbon dioxide evolution (CER) was used to control the substrate feed rate in a lab scale bioreactor. The control strategy was based solely on the ratio between the relative increase in CER and the relative increase in feed rate. Severely inhibiting hydrolyzates could be fermented without detoxification and the time required for fermentation of moderately inhibiting hydrolyzates was also reduced. The feed rate approached a limiting value for inhibiting media, with a corresponding pseudo steady-state value for CER. However, a slow decrease of CER with time was found for media containing high amounts of 5-hydroxymethyl furfural (HMF). The success of the control strategy is explained by the conversion of furfural and HMF by the yeast during fed-batch operation. The hydrolyzates contained between 1.4 and 5 g/l of furfural and between 2.4 and 6.5 g/l of HMF. A high conversion of furfural was obtained (between 65-95%) at the end of the feeding phase, but the conversion of HMF was considerably lower (between 12-40%). PMID- 10861414 TI - Optimized synthesis of L-sorbose by C(5)-dehydrogenation of D-sorbitol with Gluconobacter oxydans. AB - The optimization of L-sorbose synthesis by regiospecific dehydrogenation of D sorbitol using Gluconobacter oxydans is reported. The current L-sorbose production processes that are based on G. oxydans and other bacterial strains are suboptimal as to yield and rate of L-sorbose synthesis. One reason for these problems is the toxicity that is induced by the substrate D-sorbitol when used in concentrations of >10% (w/v). This phenomenon significantly limits the potentials of L-sorbose production from an industrial point of view. The goal of this study was to develop a fast production process that yields L-sorbose in stoichiometric amounts starting from D-sorbitol concentrations that exceed 10% (w/v). A gradual improvement of the inoculum build-up procedure, culture medium composition, and process parameters ultimately led to a theoretically maximal L-sorbose productivity (200 g L(-1) of L-sorbose from 200 g L(-1) of D-sorbitol in 28 h of fermentation) using a Gluconobacter oxydans mutant strain that was selected under conditions of substrate inhibition. Because the D-sorbitol/L&HYPHEN;sorbose bioconversion is used to mass-produce vitamin C, the procedure reported here will contribute to a more efficient and more economic synthesis of vitamin C. PMID- 10861415 TI - Immobilization of alliinase on porous aluminum oxide. AB - Membrane filters prepared from porous aluminum oxide (Anopore) were investigated for their potential use as a durable support for enzymes. Alliinase (EC 4.4.1.4) was chosen as a model enzyme for immobilization experiments. To allow for smooth fixation, the enzyme was immobilized indirectly by sugar-lectin binding. Monomolecular layers of the lectin concanavalin A and alliinase were applied by self-assembling processes. As an anchor for these layers, the sugar, mannan, was covalently coupled to the membrane surface. This procedure exhibits several advantages: (i) enzyme immobilization can be carried out under smooth conditions; (ii) immobilization needs little time; and (iii) protein layers may be renewed. PMID- 10861416 TI - Pathology of early invasive adenocarcinoma of the esophagus or esophagogastric junction: implications for therapeutic decision making. AB - BACKGROUND: As an alternative to surgical resection, endoscopic treatment modalities are being explored for the treatment of patients with early esophageal carcinoma. This study aimed to evaluate patterns of local growth and regional dissemination of early adenocarcinoma of the esophagus or esophagogastric junction, as these pathologic features may contribute to rational therapeutic decision making. METHODS: Among 173 patients who underwent esophageal resection for invasive adenocarcinoma (1993-1998), 32 (19%) had early stage cancer (pT1). Clinical records, pathology reports, and original slides of the surgically resected esophagus were reviewed in each case. RESULTS: In 12 patients tumor invasion was limited to the mucosa, whereas in 20 patients the tumor showed infiltration of the submucosa. All cancers were associated with intestinal metaplasia. Areas of high grade dysplasia accompanied 27 of the 32 cancers (84%). Intramucosal cancer had no lymph node metastasis but presented as multifocal disease in 42% of cases and extended under preexisting squamous mucosa in 17% of cases. In submucosal cancer, lymph node metastases were present in 30% of cases. Disease specific 3-year survival for patients with intramucosal cancer was 100% and for those with submucosal cancer 82% (P = not significant). CONCLUSIONS: Based on the local growth pattern of intramucosal adenocarcinoma of the esophagus or esophagogastric junction, endoscopic treatment of patients with this disease should be applied with caution. For submucosal carcinoma, surgery is the mainstay of treatment, as lymph node metastasis is frequently present. Both subclassifications of early cancer show a favorable outcome after esophagectomy. PMID- 10861417 TI - Morphometric analysis of regional lymph nodes with and without metastasis from early gastric carcinoma. AB - BACKGROUND: To the authors' knowledge detailed morphometric changes in lymph nodes with and without metastasis in patients with early gastric carcinoma remain undocumented. METHODS: Histologic slides of 1847 lymph nodes dissected from 115 consecutive patients who underwent gastrectomy for early gastric carcinoma were examined histologically and measured using computer morphometry with the public domain National Institutes of Health Image program. Quantitative data were analyzed in relation to preoperative and intraoperative clinical assessments and postoperative pathologic diagnosis. RESULTS: Metastasis was found in 11 lymph nodes (0.6%) from 8 patients (7.0%). Metastatic lymph nodes showed a mean maximum dimension of 4.8 mm, a mean area of 14.4 mm(2), and a mean ratio of maximum/minimum dimension of 1.36; the corresponding values for nonmetastatic lymph nodes were 4.7 mm (P = 0.45), 13.2 mm(2) (P = 0. 13), and 1.66 (P = 0.10), respectively. The lymph node with a metastasis was not necessarily the largest of the dissected lymph nodes from each patient, and histologically each lymph node with a metastasis showed pericancerous fibrosis in > 10% of its area. The sensitivities of preoperative computed tomography, abdominal ultrasonography (US), endoscopic US, and intraoperative assessments to diagnose metastasis were 0%, 13%, 0%, and 13%, respectively, and the sensitivities of these modalities to detect lymph nodes > 10 mm in dimension were 18%, 10%, 3%, and 10%, respectively. CONCLUSIONS: Digital quantitative analysis is useful and widely applicable to clinicopathologic evaluation. The diagnostic sensitivity of lymph node metastasis in patients with early gastric carcinoma in the current study was very low with preoperative and intraoperative assessments because lymph node metastases were small and showed subtle histologic changes of pericancerous fibrosis. PMID- 10861418 TI - Influence of gastrectomy on the distribution of intravenously administered 5 fluorouracil to regional lymph nodes in an animal model. AB - BACKGROUND: Neoadjuvant and postoperative chemotherapies have been widely employed for the treatment of patients with gastric carcinoma. However, to the authors' knowledge there have been no studies that focused on pharmacokinetic differences between antineoplastic agents administered before versus after gastrectomy. The influence of gastrectomy on the distribution of antineoplastic agents, especially to the regional lymph nodes, were examined. METHODS: Twelve pigs were divided into 2 groups of 6 each (i.e., those with and without total gastrectomy). Under general anesthesia, 10 mg/kg of 5-fluorouracil (5-FU) was injected rapidly. Total gastrectomies were performed before these injections in the group that underwent gastrectomy. In both groups a lymph node at the root of the left gastric artery (1), a preaortic lymph node (2), and venous blood were sampled in 3 animals at 15 minutes and in another 3 animals at 60 minutes after the 5-FU injection. The 5-FU concentrations in these samples were determined by high performance liquid chromatography. RESULTS: The 5-FU concentrations of 1, 2, and plasma at 15 minutes/60 minutes after injection in the nongastrectomized group were 7.75 x 10(3)/227 ng/g, 7.58 x 10(3)/439 ng/g, and 7.24 x 10(3)/417 ng/mL, respectively. The corresponding concentrations in the gastrectomized group were 6.04 x 10(3)/211 ng/g, 5.80 x 10(3)/206 ng/g, and 5.76 x 10(3)/229 ng/mL, respectively. There were no differences in 5-FU concentrations among any of the lymph nodes or the plasma, regardless of whether gastrectomy was performed. CONCLUSIONS: No influence of gastrectomy on the distribution of 5-FU to the regional lymph nodes was observed in animal models in the current study. PMID- 10861419 TI - Intra-abdominal fibrosis after systemic and intraperitoneal therapy containing fluoropyrimidines. AB - BACKGROUND: Intra-abdominal and retroperitoneal fibrosis has been described as secondary to intraperitoneal (IP) administration of several chemotherapeutic agents, including carboplatin, mitoxantrone, and the combination of 5 fluorouracil and cisplatin. The IP administration of floxuridine (FUDR) is an effective and minimally toxic treatment for patients with metastases to the peritoneum. An increasing number of patients with colorectal, gastric, or ovarian carcinoma are treated with IP chemotherapy. METHODS: The authors report two patients with metastatic colon carcinoma who experienced severe intra-abdominal fibrosis presenting as an intra-abdominal mass mimicking recurrence in one patient and diffuse encasement of the bowel in the other, after the administration of IP FUDR and leucovorin. RESULTS: Two patients with Stage III colon adenocarcinoma received postoperative adjuvant 5-fluorouracil and levamisole. They subsequently presented with a rise in carcinoembryonic antigen level and isolated liver metastasis. They underwent hepatic lobectomy with postoperative intra-arterial hepatic FUDR and systemic 5-fluorouracil and leucovorin. They each had an intra-abdominal recurrence, which was resected and treated with postoperative IP FUDR and leucovorin. They then presented with a diffuse pattern of IP fibrosis with no tumor identified. CONCLUSIONS: IP FUDR and leucovorin therapy can be associated with diffuse IP fibrosis, which in this study caused an intra-abdominal mass that was indistinguishable from recurrent malignancy in one patient and encasement of the bowel in the other. PMID- 10861420 TI - Treatment of intrahepatic malignancy with radiofrequency ablation: radiologic pathologic correlation. AB - BACKGROUND: Radiofrequency (RF)-induced tissue coagulation represents a new approach for the thermal destruction of tumors within the liver. The purpose of the current study was to 1) assess technique safety; 2) determine the extent and evolution of induced cellular damage; and 3) correlate the observed pathologic effects with radiologic studies. METHODS: Twenty-three tumors measuring 3 days after ablation showed definite, contiguous coagulative necrosis without intervening areas of viable tumor. CT and MRI scans demonstrated circumscribed hypodense, nonenhancing regions surrounding the electrode tract as early as 15 minutes after ablation. These corresponded within 2 mm to measurements of coagulation at pathology. CONCLUSIONS: RF ablation is a minimally invasive and safe approach to the treatment of tumors in the liver. Tumors treated with RF energy do not immediately demonstrate coagulative necrosis, but do show evidence of irreversible cellular damage. The extent of tumor necrosis correlates closely with findings at contrast-enhanced imaging. PMID- 10861421 TI - Prognostic significance of heat shock protein-27 expression in hepatocellular carcinoma and its relation to histologic grading and survival. AB - BACKGROUND: The expression of heat shock protein-27 (HSP-27) has been detected in some human tumors. In this study the authors investigated HSP-27 expression in patients with hepatocellular carcinoma (HCC) and examined its prognostic significance. METHODS: Expression of HSP-27 was studied in 58 HCC and adjacent noncancerous liver tissues by immunohistochemical stain. The relation between its expression and eight known prognostic factors was evaluated. RESULTS: Of the 58 HCC tissues studied, the presence of HSP-27 was demonstrated in 45 tissues (77.6%); low expression ( 25%) was demonstrated in 28 tissues. A significantly higher distribution of HSP-27 expression in HCC tissues compared with adjacent noncancerous liver tissues was obtained (P < 0.0001). Patients with high HSP-27 expression had a significantly higher histologic tumor grade than those with low HSP-27 expression (P = 0.001). The 5-year disease free survival rate of patients with high HSP-27 expression was 21.4% versus 59.3% for patients with low HSP-27 expression (P < 0.001). A similar relation was observed with overall survival (33.3% vs. 64. 8%; P = 0.009). HSP-27 expression was also identified to be a significant and powerful prognostic indicator for disease free survival (odds ratio = 2.25; P = 0.034) and for overall survival (odds ratio = 2.72; P = 0.015). CONCLUSIONS: The current study data suggest that HSP-27 expression is a powerful prognostic indicator and is related to histologic grade and survival of patients with HCC. PMID- 10861422 TI - Incidence of primary cholangiocellular carcinoma of the liver in japanese patients with hepatitis C virus-related cirrhosis. AB - BACKGROUND: Hepatitis C virus (HCV) infection is a major risk factor for the development of hepatocellular carcinoma. However, the risk factors for primary cholangiocellular carcinoma of the liver (PCC-L) have not been fully investigated. The authors determined the incidence of PCC-L in patients with HCV related cirrhosis. METHODS: Between 1980 and 1997, the authors prospectively studied 600 consecutive patients for the appearance of PCC-L; these patients were positive for HCV and later developed cirrhosis. The follow-up period ranged from 0 to 18.5 years (median, 7.2 years). RESULTS: During the observation period, PCC L developed in 14 patients (2.3%). Among these, 11 (1.8%) had cholangiocellular carcinomas and the other 3 (0.5%) had a combined type of hepatocellular and cholangiocellular carcinoma. Within the same period, hepatocellular carcinoma (HCC) developed in 206 patients (34.3%). The cumulative rates of newly diagnosed PCC-L were 1.6% at 5 years and 3.5% at 10 years, which was about 1000 times higher than the estimated incidence of PCC-L in the general population of Japan. PCC-L was treated by surgical resection in 3 patients who survived for > 3 years. However, the other 11 patients received palliative therapy or chemotherapy. The survival rates among PCC-L patients were 39.3%, 23. 6%, and 16.5% at the end of 1, 3, and 5 years, respectively, and were significantly lower than those of HCC (P = 0.0001). CONCLUSIONS: The results of this study show a relatively high incidence of PCC-L in patients with HCV-related cirrhosis, and also show that this type of liver cancer is associated with a relatively poor prognosis. These results indicate that HCV-related cirrhosis is a major risk factor for PCC-L in Japanese patients. PMID- 10861423 TI - The significance of hepatitis B virus DNA detected in hepatocellular carcinoma of patients with hepatitis C. AB - BACKGROUND: Recently, it has been reported that hepatitis B virus (HBV) DNA is detected in cancerous liver tissues in some hepatitis B surface antigen negative chronic hepatitis C patients with hepatocellular carcinoma (HCC). However, the significance of HBV DNA detected in such cases remains unclear. METHODS: The authors determined the presence or absence and the titers of HBV DNA in the liver tissue of anti-hepatitis C virus (HCV) positive/HBs antigen negative patients with HCC by polymerase chain reaction (PCR) and Southern hybridization, and correlated them with clinicopathologic parameters. RESULTS: HBV DNA was found in cancerous liver tissues from 12 (50.0%) of the 24 patients studied. However, Southern hybridization of genomic DNA in the cancerous liver tissues led to detection of HBV DNA in only 3 (12.5%) of the 24 patients, suggesting that the copy number of HBV DNA may be very low in most cases. Indeed, the titration of HBV DNA in cancerous liver tissues performed by the end point dilution method revealed that most contained less than one copy of HBV DNA per cell, although cells of monoclonal origin carrying integrated HBV DNA should have at least one copy of it. There was no significant difference in HBV DNA positivity between the HCC patients with and without underlying cirrhosis. CONCLUSIONS: The results of this study corroborate previous reports of frequent detection of HBV DNA in the liver tissue of anti-HCV positive/HBs antigen negative patients with HCC, but do not support an essential role of HBV in hepatocarcinogenesis in patients with chronic hepatitis C and occult HBV infection. PMID- 10861424 TI - Closing the gastrin loop in pancreatic carcinoma: coexpression of gastrin and its receptor in solid human pancreatic adenocarcinoma. AB - BACKGROUND: Alpha-amidated gastrin promotes the growth of nontransfected pancreatic cell lines expressing the gastrin/cholecystokinin (CCK)-B receptor. Gastrin/CCK-B and CCK-A receptors recently were demonstrated in human pancreatic adenocarcinomas, but to the authors' knowledge expression of their ligands to date have not been adequately investigated. As a prerequisite for making suggestions regarding local growth stimulation, the authors examined whether gastrin and the homologous CCK peptides as well as their specific receptors were expressed in consecutively collected solid human pancreatic adenocarcinomas. METHODS: Using a library of radioimmunoassays specific for different epitopes on proCCK, progastrin, their processing intermediates, and bioactive end products, CCK and gastrin gene expression was measured in extracts of solid human pancreatic adenocarcinomas (n = 19), resection margins (n = 15), and normal pancreatic tissue (n = 8). Moreover, CCK, CCK-A receptor, and gastrin/CCK-B receptor mRNA were measured by reverse transcriptase-polymerase chain reaction. RESULTS: Amidated gastrins were synthetized in 14 of 19 carcinomas (median, 0.4 pmol/g; range, < 0.1-84.0 pmol/g) and in 12 of 15 resection margin samples (median, 0.3 pmol/g; range, < 0.1-6.1 pmol/g). In contrast, normal human pancreatic tissue expressed only traces of poorly processed progastrin. Gastrin/CCK-B receptor mRNA was present in all carcinomas, resection margins, and normal pancreatic tissue. CCK-A receptor mRNA was detected in most tumors, but neither the mature ligands (alpha-amidated and O-sulfated CCK peptides) nor their precursors were expressed in carcinoma and normal pancreatic tissue. CONCLUSIONS: The results of the current study demonstrate that alpha-amidated gastrin peptides and their receptor invariably are coexpressed in pancreatic adenocarcinoma. Therefore these findings support the contention of a role for local gastrin regulatory mechanisms, but no CCK mechanisms, in pancreatic carcinoma. PMID- 10861425 TI - Ductal lesions in patients with chronic pancreatitis show K-ras mutations in a frequency similar to that in the normal pancreas and lack nuclear immunoreactivity for p53. AB - BACKGROUND: Chronic pancreatitis (CP) is considered to be a risk factor for the development of pancreatic carcinoma. The detection of K-ras mutations in the duodenal or pancreatic juice has been held to be a reliable tool for its early diagnosis. However, K-ras mutations also occur in hyperplastic ductal epithelium, making it difficult to interpret their role in pancreatic carcinogenesis. METHODS: The study included 30 resection specimens, 15 from patients with alcoholic CP, and 15 from patients with idiopathic CP. The mean duration of disease was 6.8 years. A total of 429 ductal lesions were classified according to the World Health Organization classification (1996) and microdissected. K-ras analysis was performed by means of polymerase chain reaction (45 cycles), constant denaturing gel electrophoresis, and sequencing. Immunostaining was performed with antibodies against p53, Ki-S5, carcinoembryonic antigen, and two types of mucins. RESULTS: The 30 specimens demonstrated all types of ductal lesions. Severe cellular atypia was not observed. A total of 429 ductal lesions were analyzed. Approximately 4.4% of the lesions (19 of 429) from 27% of the patients (8 of 30) showed K-ras mutations, but they were unrelated to the duration or type of CP. Immunostaining for mutated p53 protein always was negative. Increased proliferative activity was noted only in patients with papillary hyperplasia. No patient developed pancreatic carcinoma within a follow up period of at least 3 years. CONCLUSIONS: Ductal lesions in patients with CP exhibit K-ras mutations without additional indications of neoplastic transformation such as severe dysplasia or mutated p53 protein. Therefore, for diagnostic and therapeutic purposes, the detection of K-ras mutations should be supplemented by the demonstration of additional genetic alterations or clinical signs of malignancy. PMID- 10861426 TI - A phase II trial of biweekly high dose gemcitabine for patients with metastatic pancreatic adenocarcinoma. AB - BACKGROUND: Although the novel cytidine analog gemcitabine has shown superior antitumor activity compared with weekly bolus 5-fluorouracil in patients with advanced pancreatic carcinoma, further improvements of therapeutic results are warranted. The current Phase II study was initiated to investigate whether this might be achieved by dose intensification. METHODS: Between August 1997 and September 1998, 43 consecutive patients with metastatic pancreatic adenocarcinoma were enrolled in this multicenter Phase II trial. Patients received 4 weekly courses of gemcitabine 2200 mg/m((2)) given as intravenous infusion during 30 minutes on Days 1 and 15 for a duration of 6 months unless there was prior evidence of progressive disease. The efficacy of treatment was assessed according to standard criteria, i.e., objective response, progression free survival, and overall survival, as well as by analysis of clinical benefit response (defined as >/= 50% reduction in pain intensity, >/= 50% reduction in daily analgesic consumption, and/or >/= 20 point improvement in Karnofsky performance status that was sustained for >/= 4 consecutive weeks). RESULTS: Of 43 patients evaluable for objective response, 1 achieved complete and 8 partial remissions, for an overall response rate of 21% (95% confidence interval, 10-36%); 18 additional patients (42%) had stable and 16 (37%) progressive disease. The median time to progression was 5.3 months. Median survival was 8.8 months, and the probability of surviving beyond 12 months was 26.3%. Of 36 patients with tumor-related symptoms who were considered evaluable for clinical benefit response, 16 (44%) experienced significant palliation. The median time to achieve a clinical benefit response was 6 weeks, and its median duration was 27 weeks. Chemotherapy was well tolerated, with leukopenia/granulocytopenia representing the most common side effect. Gastrointestinal and other subjective toxicities were infrequent and generally mild. CONCLUSIONS: Biweekly high dose gemcitabine seems to represent a safe, tolerable, and effective regimen for the palliative treatment of patients with advanced pancreatic carcinoma. PMID- 10861427 TI - Intraperitoneal chemohyperthermia with mitomycin C for digestive tract cancer patients with peritoneal carcinomatosis. AB - BACKGROUND: Most patients with peritoneal carcinomatosis of digestive tract origin die within 6 months. Intraperitoneal chemohyperthermia (IPCH) associated with surgery has been reported as a possible new therapeutic approach. METHODS: A prospective Phase II trial was carried out with 83 patients who had digestive tract cancer and peritoneal carcinomatosis to evaluate the tolerance and efficacy of IPCH with mitomycin C (MMC) associated with surgery. Eighty-six IPCH treatments with MMC were given as complementary therapy after surgery (peritoneal perfusate with a 10 mg/L dose of MMC; inflow temperature, 46-49 degrees C; use of a closed circuit; duration, 90 minutes). Primary tumors were mainly gastric (in 42 cases) or colorectal (in 27 cases). RESULTS: Mortality and morbidity occurred in 3 of 83 cases and 8 of 83 cases, respectively. For patients with resectable tumors, the median survival time was 16 months when carcinomatosis was Stage I and II (malignant granulations less than 5 mm in greatest dimension), whereas it was 6 months when carcinomatosis was Stage III and IV (malignant granulations more than 5 mm in greatest dimension). For patients with resectable gastric cancer and Stage I and II carcinomatosis, 1-, 2-, and 3-year actuarial survival rates were 80%, 61%, and 41%, respectively, whereas the rate was 10% at 1 year for patients with bulky disease (Stage III and IV). CONCLUSIONS: IPCH appears to be a promising new approach to treating patients with digestive tract cancers and peritoneal carcinomatosis with small, malignant granulations (Stage I and II). PMID- 10861428 TI - Intestinal metaplasia is the probable common precursor of adenocarcinoma in barrett esophagus and adenocarcinoma of the gastric cardia. AB - BACKGROUND: Intestinal metaplasia in the tubular esophagus is the recognized precancerous lesion of adenocarcinoma in Barrett esophagus. However, it is not yet clear whether adenocarcinoma of the gastric cardia arises from the same premalignant lesion, i.e., intestinal metaplasia of the gastric cardia. The purpose of this study was to compare adenocarcinomas in Barrett esophagus and adenocarcinomas of the gastric cardia at an early stage, when it was more likely that intestinal metaplasia had not been completely overgrown by the tumor. METHODS: The authors compared the epidemiologic, clinical, and pathologic features of early stage adenocarcinoma in Barrett esophagus and adenocarcinoma of the gastric cardia from 42 patients who underwent resection surgery. The presence of intestinal metaplasia was assessed in the resected specimens by using Alcian blue (pH 2.5) staining. RESULTS: Intestinal metaplasia was detected in the mucosa adjacent to neoplasia in 25 of 26 patients with adenocarcinoma in Barrett esophagus and in 11 of 16 (69%) patients with adenocarcinoma of the gastric cardia. Patient and tumor characteristics and survival were comparable in both groups. CONCLUSIONS: Intestinal metaplasia is a very common finding in the mucosa adjacent to early stage adenocarcinoma of the gastric cardia. Adenocarcinoma in Barrett esophagus and adenocarcinoma of the gastric cardia may represent the same disease; the former arises from longer segments of intestinal metaplasia and the latter from intestinal metaplasia of the cardia. PMID- 10861429 TI - The clinical significance of CD34 expression in response to therapy of patients with acute myeloid leukemia: an overview of 2483 patients from 22 studies. AB - BACKGROUND: Although many studies have been performed to evaluate the prognostic significance of CD34 expression in acute myeloid leukemia (AML), the findings have been inconsistent. In this study, the authors reviewed such previous studies to establish a definite conclusion. METHODS: Using MEDLINE, the authors identified studies that evaluated the prognostic significance of CD34 expression in AML. The outcome measure was the complete remission rate. They used the random effect method to combine the results. Results were presented as odds ratios (ORs) with 95% confidence intervals (CIs). The ORs were less than 1 if the complete remission occurred more frequently in the CD34 negative group. RESULTS: Twenty two studies including 2483 patients were reviewed. The combined OR was 0.38 (95% CI, 0. 26-0.57), which suggested that CD34 expression was associated with a poor remission rate. However, the authors found statistical evidence of marked heterogeneity among trials (P < 0.001), especially according to time of publication. The combined OR in studies published in or after 1994 was 0.70 (95% CI, 0.47-1.09). The authors divided the studies into several subgroups, but they could not determine the reason for the heterogeneity. CONCLUSIONS: At present, CD34 expression should not be considered a marker of poor prognosis because it is not supported by the combined data from recent studies. Further studies should be conducted to investigate the intensity of CD34 expression in specific populations of patients, such as those with t(8;21) or t(15;17) translocations or the AML-M0 subtype. PMID- 10861430 TI - Ultrasound examination of regional lymph nodes significantly improves early detection of locoregional metastases during the follow-up of patients with cutaneous melanoma: results of a prospective study of 1288 patients. AB - BACKGROUND: In regional lymph node metastasis of cutaneous melanoma, the number and volume of involved lymph nodes are the most important prognostic factors. Several studies have revealed that palpation of the lymphatic drainage area(s) and regional lymph nodes has a high rate of false-negative results during follow up. The aim of the current study was to assess the sensitivity and specificity of ultrasound versus clinical diagnosis in the detection of subcutaneous and regional metastases. METHODS: During a period of 42 months, a total of 6328 lymphatic drainage areas were examined clinically and by ultrasound (7.5-10 MHz) in 1288 melanoma patients at 4435 follow-up consultations. When an ultrasound finding was suggestive of metastasis, surgery and histopathologic evaluation were performed. The results of clinical examination, ultrasound examination, and histopathologic findings were compared. RESULTS: In 504 ultrasound examinations performed on 235 patients, metastatic disease was diagnosed in 263 examinations following surgery (179 patients). Due to advanced disease or rejection, an additional 56 patients did not undergo surgery. In 239 of the 263 positive findings (90.9%), metastases from melanoma were histopathologically confirmed. In 8 cases (3%) a second malignancy and in 16 cases (6. 1%) benign lymphadenopathy was histopathologically diagnosed. Palpation of subcutaneous lymph nodes and lymph nodes gave false-negative results in 68 of the 238 cases of histopathologically proven metastases (28.6%). Clinical examination was least sensitive in the supraclavicular, axillary, and infraclavicular regions. The sensitivity and specificity for ultrasound examination were 89.2% and 99.7%, respectively, and 71.4% and 99.7% for clinical examination, respectively. CONCLUSIONS: For early diagnosis of in-transit and regional lymph node metastases in cutaneous melanoma, ultrasound scanning is distinctly superior to clinical examination. Controlled follow-up studies are proposed to examine the possible beneficial effects on survival time resulting from the ultrasound examinations of the lymphatic drainage area(s) and regional lymph nodes. PMID- 10861431 TI - Accuracy of sentinel lymph node biopsy in patients with large primary breast tumors. AB - BACKGROUND: Patients with large breast tumors are increasingly undergoing neoadjuvant treatment to downstage local disease; however, accurate staging of the axilla before the initiation of chemotherapy remains problematic. In the current study, the authors report on the accuracy of sentinel lymph node (SLN) biopsy in such patients to determine the feasibility of applying this technique before induction chemotherapy. METHODS: One hundred three patients with 104 tumors classified as American Joint Committee on Cancer (AJCC) T2 (tumor >/= 2 cm but /= 3 cm, 1 false-negative result (2% [95% exact CI, < 1-15%]) was identified, and the rate of lymph node metastasis was 62.5% (95% exact CI, 48. 5-75%) (35 of 56 tumors). Within 30 SLN positive patients with tumors >/= 3 cm and complete axillary lymph node dissection, 3 of 8 patients (37.5% [95% exact CI, 8.5-75.5%]) with micrometastasis ( 2 mm) to the SLN (P = 0.002). CONCLUSIONS: SLN biopsy for patients with large breast tumors is technically feasible and highly accurate. SLN biopsy should be considered for the staging of clinically negative axilla in patients scheduled to receive neoadjuvant chemotherapy. PMID- 10861432 TI - Lymphatic mapping with intralesional tracer administration in breast carcinoma patients. AB - BACKGROUND: The objectives of the study were to determine how often a sentinel lymph node is visualized by lymphoscintigraphy in breast carcinoma patients, how often the sentinel lymph node is identified during surgery, and the sensitivity of these procedures to identify the presence of axillary lymph node metastasis. METHODS: A total of 136 patients were enrolled in 2 hospitals. Preoperative dynamic and static lymphoscintigraphy were performed; in addition, both a vital dye and a gamma detection probe were used intraoperatively. The tracers were injected into the primary lesion. Sentinel lymph node biopsy was followed by completion axillary lymph node dissection. The sentinel lymph nodes and other axillary lymph nodes were examined routinely and by immunohistochemical staining. RESULTS: A sentinel lymph node was visualized by lymphoscintigraphy in 118 patients (87%). During the operation a sentinel lymph node was localized in 126 patients (93%). A total of 224 sentinel lymph nodes were harvested (average of 1.7 and range of 1-4 sentinel lymph nodes per patient). Of all the sentinel lymph nodes, 37 were blue (17%), 68 were radioactive (30%), and 119 were both blue and radioactive (53%). The sentinel lymph nodes contained metastatic disease in 56 patients (44%). Three sentinel lymph node biopsies were false-negative (sensitivity 95%). CONCLUSIONS: Sentinel lymph node biopsy with preoperative lymphoscintigraphy after intralesional tracer administration and intraoperative use of both a gamma detection probe and a vital dye is a reliable technique for staging the axilla of breast carcinoma patients. PMID- 10861433 TI - Differences in the pathologic features of ductal carcinoma in situ of the breast based on patient age. AB - BACKGROUND: Young patient age at diagnosis has been reported as a risk factor for recurrence in patients with ductal carcinoma in situ (DCIS) of the breast treated with breast-conserving therapy (BCT). The authors examined pathologic features of DCIS in three different age groups of patients to identify differences that might explain why young patient age at the time of diagnosis is a risk factor for recurrence. METHODS: Excised specimens from 177 breasts of 172 patients with DCIS treated with BCT were studied. All slides from all specimens were reviewed. Patients were divided into 3 age groups: those age < 45 years, those ages 45-59 years, and those age >/= 60 years. The histologic features that were quantified included most common and highest nuclear grades, DCIS architectural pattern, amount of central necrosis (quartiles), calcifications, amount of DCIS, and number of terminal duct lobular units (TDLUs) with cancerization of lobules (COL) within 0.42 cm of the margin, margin status, and size and volume of excision specimens. RESULTS: Patients age < 45 years at the time of diagnosis more frequently had higher nuclear grade DCIS (highest nuclear Grade 3: 69%, 60%, and 39%; P = 0.003), respectively and central necrosis (72%, 62%, and 44%; P = 0. 01), respectively. Although not statistically significant, younger patients tended to have comedo subtype DCIS more often (31%, 23%, and 19%; P = 0.35), respectively. Younger patients also more often had smaller initial biopsy specimen maximum dimensions (4.3 cm, 5.2 cm, and 5.7 cm; P = 0.004), respectively, with close or positive margins (89%, 61%, and 64%; P = 0.03), and more TDLUs with COL in the 0.42-cm rim of tissue adjacent to the margin (5.2, 3.6, and 1.9; P = 0.23), respectively. No other features including the amount of DCIS when classified as > 50% or > 75% of ducts, calcifications within DCIS ducts, pattern of DCIS involvement, number of slides examined, number of slides with DCIS, and mean number of DCIS ducts near the margin were found to occur more frequently in younger patients. CONCLUSIONS: Younger patients with DCIS may have an increased risk of local recurrence when treated with BCT due to smaller initial excision volumes, a greater proportion of high nuclear grade DCIS, and central necrosis. PMID- 10861434 TI - Changing incidence rate of invasive lobular breast carcinoma among older women. AB - BACKGROUND: In 1998, an unusually large number of invasive lobular breast carcinoma cases were seen at the University of Washington. The purpose of this study was to assess whether the incidence rate of invasive lobular carcinoma has been increasing disproportionately compared with the incidence rate of invasive ductal carcinoma. METHODS: Age specific and age-adjusted breast carcinoma incidence rates from 1977-1995 were obtained from the nine population-based cancer registries that participate in the Surveillance, Epidemiology, and End Results (SEER) program. Three histologic groupings were used: lobular, ductal, and all invasive breast carcinomas. Overall incidence rates for each grouping, as well as for each stage (local, regional, and distant), were obtained. RESULTS: The rate of incidence of lobular carcinoma increased steadily from 1977-1995 in women age >/= 50 years whereas it remained stable in women age < 50 years. Alternatively, the rate of incidence of ductal carcinoma increased steadily from 1977-1987, but from 1987-1995 it remained relatively constant across all age groups. CONCLUSIONS: The incidence rates of invasive lobular breast carcinomas increased steadily since 1977 whereas the incidence rates of invasive ductal carcinoma have plateaued since 1987. This rise occurred specifically among women age >/= 50 years and may be related to postmenopausal status. Further epidemiologic, clinical, and laboratory research is required to assess what factors are contributing to this trend. PMID- 10861435 TI - Hormone replacement therapy in relation to risk of lobular and ductal breast carcinoma in middle-aged women. AB - BACKGROUND: In the majority of studies, long term, recent use of hormone replacement therapy has been associated with an increased risk of breast carcinoma. However, little attention has been paid to the possibility that the magnitude of this association may vary according to the histologic type of breast carcinoma. METHODS: In this population-based case-control study, interviews were conducted with 537 female residents of King County, Washington who were ages 50 64 years and who had been diagnosed with primary breast carcinoma between January 1, 1988 and June 30, 1990. Interviews with 492 randomly selected King County women without a history of breast carcinoma served as a basis for comparison. Analyses were performed separately for women with lobular and for those with ductal tumors. RESULTS: Compared with nonusers of menopausal hormones, those who currently were using combined estrogen and progestin hormone replacement therapy (CHRT) and had done so for at least 6 months had an elevated risk of lobular breast carcinoma (odds ratio [OR] = 2.6; 95% confidence interval [95% CI], 1.1 5.8), but no change in their risk of ductal breast carcinoma was noted (OR = 0.7; 95% CI, 0.5-1. 1). The OR associated with current use of unopposed estrogen for at least 6 months was 1.5 (95% CI, 0.5-3.9) for lobular tumors and 0.7 (95% CI, 0.4-1.1) for ductal tumors. Similar results were found when cases of invasive tumor were analyzed separately. CONCLUSIONS: The results of this study suggest that CHRT use increases the risk of lobular, but not ductal, breast carcinoma in middle-aged women. PMID- 10861436 TI - A multivariate analysis of blood vessel and lymph vessel invasion as predictors of ovarian and lymph node metastases in patients with cervical carcinoma. AB - BACKGROUND: To the authors' knowledge there are few available data regarding the influence of lymphovascular space invasion, which has been examined separately as two components (lymphatic vessel invasion [LVI] and blood vessel invasion [BVI]), in the metastasis of cervical carcinoma. METHODS: LVI and BVI, which include capillary vessel invasion, were reviewed retrospectively based on the histopathologic slides of 239 women with cervical carcinoma who were treated with radical hysterectomy. The correlation between lymph node and/or ovarian metastases and LVI, BVI, and other histopathologic factors was investigated by multiple logistic regression analysis. The influence of LVI and BVI on survival was examined by Cox regression analysis. RESULTS: The rate of incidence of LVI was higher than that of BVI in all stages of cervical carcinoma (P < 0.0001 for International Federation of Gynecology and Obstetrics Stage IB and Stage II disease and P < 0.05 for Stage III disease). The incidence rate of BVI increased as LVI became more prominent and there was a significant correlation between the two findings (P < 0.0001). BVI was more frequent in adenocarcinoma/adenosquamous carcinoma than in squamous cell carcinoma (P < 0.05). LVI (P < 0.0001) and parametrial invasion (P < 0.0001) were significantly related to lymph node metastasis on multivariate analysis. Conversely, BVI (P < 0.05) and parametrial invasion (P < 0.0025), as well as adenocarcinoma or adenosquamous carcinoma (P < 0.0005), were significantly related to ovarian metastasis on multivariate analysis. With regard to the prognostic significance of these components, it was found that BVI (hazards ratio [HR] = 2.0), ovarian metastasis (HR = 6.5), and lymph node metastasis (HR = 5.5) were significantly related to a poor prognosis in women with cervical carcinoma. CONCLUSIONS: Ovarian metastasis may occur via hematogenous spread of cervical carcinoma. The results of the current study suggest that BVI, including capillary vessels, that is diagnosed separately from LVI using hematoxylin and eosin stained sections may be an important prognostic factor for patients with cervical carcinoma. PMID- 10861437 TI - Clinical characteristics of clear cell carcinoma of the ovary: a distinct histologic type with poor prognosis and resistance to platinum-based chemotherapy. AB - BACKGROUND: A retrospective review of treatment results comparing women with clear cell carcinoma of the ovary (CCC) with a group with serous adenocarcinoma of the ovary (SAC) was conducted. METHODS: Between 1988-1998, 662 patients with epithelial ovarian carcinoma were identified through the medical records department and the tumor registry at 4 institutions. After the central pathologic review, 101 patients with pure or dominant (>/= 90%) CCC (15.3%) were entered into the current study. Two hundred thirty five patients with pure SAC were selected as a group for comparison. All patients underwent staging laparotomy followed by platinum-based chemotherapy. Distribution of the International Federation of Gynecology and Obstetrics (FIGO) disease stage, response to chemotherapy, and prognosis for patients with CCC were compared with the same values in patients with SAC. RESULTS: Patients with CCC were significantly more likely to have FIGO Stage I disease than were patients with SAC (48.5% vs. 16.6%). A high recurrence rate was noted in those patients with Stage IC CCC (37%). In those patients with Stage IC disease, the survival rates for patients with CCC were lower than those for patients with SAC. The 3-year and 5-year survival rates for Stage III CCC patients were significantly lower compared with Stage III SAC patients. The response rate to platinum-based chemotherapy in patients with CCC was significantly lower than that in patients with SAC. CONCLUSIONS: CCC is an intriguing histologic type of epithelial ovarian cancer that demonstrates a clinical behavior distinctly different from that of SAC. PMID- 10861438 TI - Circulating neuroendocrine markers in patients with prostate carcinoma. AB - BACKGROUND: Circulating neuroendocrine markers were measured in patients with prostate carcinoma (PC), prostatic intraepithelial neoplasia (PIN), and benign prostatic hypertrophy (BPH) with the goal to: 1) evaluate the differences in the expression of these markers in patients with benign, premalignant, and primary or metastatic PC; 2) evaluate their prognostic significance; 3) compare values in patients with hormone-naive and hormone-refractory disease; and 4) assess changes after androgen deprivation or chemotherapy. METHODS: Serum neuron specific enolase (NSE) (immunoradiometric assay) and plasma chromogranin A (CgA) (enzyme linked immunoadsorbent assay) were evaluated in 141 patients with BPH, 54 patients with PIN, and 159 patients with PC; 119 patients were bearing hormone naive disease and 40 were bearing hormone-refractory disease. CgA was monitored in 31 patients submitted to androgen deprivation and in 24 patients receiving chemotherapy. RESULTS: Supranormal CgA was observed more frequently in patients with American Urologic Association (AUA) Stage D2 disease (45.5%) compared with those with Stage D1 disease (33.3%), Stage C disease (16.7%), Stage A/B disease (18.8%), PIN (25.9%), and BPH (17.0%) (P < 0.02). Supranormal NSE did not change in any of the patient subgroups. Elevated CgA was observed in 36.0% of patients with metastases who had hormone-naive disease and in 45.0% of patients with hormone-refractory disease (P value not significant). Supranormal NSE and CgA values were predictors for poor prognosis in patients with hormone-refractory disease. Elevated baseline CgA values decreased > 50% in 1 of 12 patients who received luteinizing hormone-releasing hormone analogs and in 2 of 12 patients who underwent chemotherapy. CONCLUSIONS: CgA appears to reflect the neuroendocrine activity of PC better than NSE. Elevated CgA values correlate with poor prognosis and are scarcely influenced by either endocrine therapy or chemotherapy. PMID- 10861439 TI - MIB-1 as a proliferative marker in transitional cell carcinoma of the bladder: clinical significance and comparison with other prognostic factors. AB - BACKGROUND: Staging and grading of transitional cell carcinoma of the bladder are generally viewed as indicators of prognosis and form the basis of therapy, but they do not predict outcome accurately. This study was designed to evaluate the value for predicting recurrence, progression, and survival of proliferation fraction in transitional cell carcinoma of the bladder determined by immunostaining of histopathologic specimens with the monoclonal antigen MIB-1. METHODS: In a prospectively followed group of 301 patients with transitional cell carcinoma of the bladder, formalin fixed tumor specimens were immunostained and the MIB-1 labeling index was determined. Crude survival, progression free survival, and recurrence free survival (for patients with Ta and T1 tumors) were assessed in univariate and multivariate analysis according to stage, grade, mitotic index of the tumor, and patient age. The median value of continuous variables was used as a cutoff point in statistical analysis. RESULTS: In univariate analysis there was a strong association between all included factors and crude survival, progression free survival, and recurrence free survival with a median follow-up period of 60 months. In multivariate analysis, crude survival and progression free survival were determined by stage (P = 0.0001) and age (P = 0.0001). Recurrence free survival for patients with Ta and T1 tumors was determined by MIB-1 labeling index (P = 0.0317), mitotic index (P = 0.0229), and age (P = 0.0001). CONCLUSIONS: MIB-1 immunostaining in transitional cell carcinoma of the bladder correlated well with grade, stage, and clinical outcome. In multivariate analysis, proliferation fraction had prognostic value in predicting recurrence free survival for patients with Ta and T1 tumors, whereas stage and age appeared to be predictors of progression free survival. PMID- 10861440 TI - Expression of hypoxia-inducible factor 1alpha in brain tumors: association with angiogenesis, invasion, and progression. AB - BACKGROUND: Hypoxia inducible factor-1 (HIF-1) plays a critical role in angiogenesis during vascular development. The authors tested the hypothesis that HIF-1 expression correlates with progression and angiogenesis in brain tumors. METHODS: The authors investigated the expression of the HIF-1alpha and HIF-1beta subunits in human glioma cell lines and brain tumor tissues using Western blot analysis and immunohistochemistry. RESULTS: In glioblastomas multiforme (GBMs), HIF-1alpha primarily was localized in pseudopalisading cells around areas of necrosis and in tumor cells infiltrating the brain at the tumor margin. In contrast, HIF-1alpha was expressed in stromal cells throughout hemangioblastomas (HBs). Like HIF-1alpha, HIF-1beta was most highly expressed in high grade tumors but was expressed more widely than HIF-1alpha, including cells away from necrotic zones. In the brains of mice injected with Glioma 261 cells, a pattern of HIF 1alpha expression identical to that observed in human GBMs was noted. CONCLUSIONS: In GBMs, the heterogeneous pattern of HIF-1alpha expression appears to be determined at least in part by tissue oxygenation, whereas in HBs the homogeneous expression of HIF-1alpha may be driven by an oncogenic rather than a physiologic stimulus. PMID- 10861441 TI - Paclitaxel-induced cell death: where the cell cycle and apoptosis come together. AB - BACKGROUND: Compelling evidence indicates that paclitaxel kills cancer cells through the induction of apoptosis. Paclitaxel binds microtubules and causes kinetic suppression (stabilization) of microtubule dynamics. The consequent arrest of the cell cycle at mitotic phase has been considered to be the cause of paclitaxel-induced cytotoxicity. However, the biochemical events, downstream from paclitaxel's binding to microtubules, that lead to apoptosis are not well understood. METHODS: The authors examined recent scientific literature about the mechanisms by which paclitaxel exerts cytotoxicity. RESULTS: In addition to an arrest of the cell cycle at the mitotic phase in paclitaxel-treated cells, recent discoveries of activation of signaling molecules by paclitaxel and paclitaxel induced transcriptional activation of various genes indicate that paclitaxel initiates apoptosis through multiple mechanisms. The checkpoint of mitotic spindle assembly, aberrant activation of cyclin-dependent kinases, and the c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK) are shown to be involved in paclitaxel-induced apoptosis. Consistent with observations that microtubules of different status (e.g., cytoskeletal microtubules vs. mitotic spindles) have different sensitivity to paclitaxel, the concentration of paclitaxel appears to be the major determinant of its apoptogenic mechanisms. CONCLUSIONS: Advances in research of the cell cycle and apoptosis have extended our understanding of the mechanisms of paclitaxel-induced cell death. Further elucidation of resistance and enhancement of paclitaxel-induced apoptosis should expedite the development of better paclitaxel-based regimens for cancer therapy. PMID- 10861442 TI - The relative risk of second nongerminal malignancies in patients with extragonadal germ cell tumors. AB - BACKGROUND: Apart from a recognized association between extragonadal mediastinal germ cell tumors (GCT) and the occurrence of hematologic malignancies, the risk of developing second nongerminal solid tumors after the diagnosis or treatment of extragonadal GCT is unknown. METHODS: Six hundred thirty-five consecutive patients with extragonadal GCT treated at 11 centers in the U.S. and Europe during the era of cisplatin-based chemotherapy (1975-1996) were included into a large database. These patients were evaluated for the occurrence of second malignancies. RESULTS: No treatment-related leukemia was observed in 611 patients treated with chemotherapy. In 7 patients, second solid tumors were observed, resulting in a frequency of 1.86% (95% confidence interval [95% CI], 1.79-1.93%) after a median follow-up of 55 months (95% CI, 50-60 months) (annual incidence, 0.30% [95% CI, 0.14-0.59]). Four solid tumors (57%) developed in patients with primary mediastinal and 3 tumors (43%) developed in patients with retroperitoneal GCT. Three patients (43%) had a nonseminomatous and 4 patients (57%) had a seminomatous histology. Six patients had been treated with chemotherapy and one patient with radiotherapy. Six of 7 solid tumors (86%) had developed within 5 years and 7 of 7 solid tumors within 10 years of diagnosis. The median time period to the occurrence of neoplasia was 47 months (range, 9-145 months). Four cutaneous tumors were observed (melanoma, two patients; basal cell carcinoma, one patient; and squamous cell carcinoma, one patient); the other three tumors were angiosarcoma, nonsmall cell lung carcinoma, and colorectal carcinoma. The overall risk for developing a second tumor was not increased compared with an age-matched general population with a standard incidence ratio (SIR) of 1.49 (95% CI, 0.60 3.06). An elevated risk for skin tumors was observed in all extragonadal GCT patients (SIR, 4.00 [95% CI, 1. 09-10.24]), as well as in the subgroup of patients treated with chemotherapy (SIR, 5.33 [95% CI, 1.45-13.65]). CONCLUSIONS: This analysis excludes an increased biologic risk of developing second solid malignancies in patients with extragonadal GCT except for the previously reported association between primary mediastinal nonseminoma and hematologic disorders. The overall risk of developing second malignancies in extragonadal GCT patients appears to be comparable to that in patients with primary testicular carcinoma. The incremental occurrence of skin malignancies in patients treated with chemotherapy should be investigated further. PMID- 10861443 TI - Testicular and paratesticular involvement by metastatic neuroblastoma. AB - BACKGROUND: Testicular and paratesticular involvement is a less familiar feature in neuroblastoma and its prognostic impact is unclear. METHODS: The records of 1076 male patients in the German cooperative neuroblastoma treatment trials were searched for patients with testicular or paratesticular involvement. RESULTS: The authors found 11 children with paratesticular or testicular involvement at the time of manifestation of the disease, 3 children with testicular involvement at the time of relapse, and 1 infant with testicular involvement at the time of progression of neuroblastoma from International Neuroblastoma Staging System (INSS) Stage 4S to Stage 4. In de novo disease, the age of manifestation did not exceed 12 months. Two children had paratesticular involvement per continuitatem by growth of a primary tumor through the inguinal channel. All others had distant primary tumor. Prognosis appeared more favorable for infants (3 of 9 died) than for older children (5 of 6 died), for children with involvement of testes and < 10% involvement of bone marrow (2 of 8 died) than for children with other distant metastasis (6 of 7 died), and for children with intrascrotal involvement at first diagnosis (4 of 11 died) than for children with intrascrotal involvement during relapse of the disease (all 4 children died). CONCLUSIONS: Paratesticular or testicular metastasis does not per se indicate unfavorable outcome and is compatible with INSS Stage 4S in infants. The age at diagnosis and the time of manifestation during disease contributed to the prognosis in those patients. PMID- 10861444 TI - Annual cancer incidence rates for Hispanics in the United States: surveillance, epidemiology, and end results, 1992-1996. AB - BACKGROUND: The expansion of the Surveillance, Epidemiology, and End Results (SEER) program and the determination of annual population estimates by county level for different racial/ethnic groups since 1990 allow the calculation of annual cancer incidence rates for Hispanics. METHODS: Incidence rates were calculated for 11 SEER areas representing 25% of the Hispanic population. Standard regression analyses of log-transformed rates were used to determine the trends of the rates. RESULTS: An important measure of the cancer burden among Hispanics is the rank order of their cancers. For Hispanic males, the five major cancers (in declining order) are prostate, lung and bronchus, colon/rectum, non Hodgkin lymphoma, and stomach cancers. For Hispanic females, the top five cancers are breast, colon/rectum, lung and bronchus, cervix, and endometrial cancers. Another measure of cancer burden is their rates relative to white non-Hispanics. Hispanic males have rates greater than white non-Hispanic males for stomach (1.6 times greater) and liver and IBD cancers (2.2), whereas Hispanic females have greater rates for cervix (2.2 times greater), liver and IBD (2.0), stomach (2.1), and gallbladder cancers (3.3). Other measures of cancer burden include the trends in Hispanic rates. Hispanic males have significant declining trends for all sites, prostate cancer, and urinary bladder cancer, and an increasing trend for liver and IBD cancers. Hispanic females have significant declining trends for cervix and urinary bladder cancers. CONCLUSIONS: The SEER cancer incidence rates and trends provide a general overview of the cancer burden among Hispanics residing in the SEER sites. This type of information is critical for determining interventions to reduce the cancer burden among Hispanics in the United States. PMID- 10861445 TI - Increased levels of promutagenic etheno-DNA adducts in colonic polyps of FAP patients. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) can regress adenomas in patients with familial adenomatous polyposis (FAP), and the mechanism involves inhibition of cyclooxygenases (COX). Reactive intermediates formed during the arachidonic acid cascade, notably by COX-2, which is upregulated in polyps of FAP patients, may promote various stages of the polyp --> adenoma --> carcinoma sequence. Etheno-DNA adducts can be derived from reactive intermediates generated during arachidonic acid metabolism and lipid peroxidation. We tested this hypothesis in colonic polyps from FAP patients and colorectal tissue from cancer patients to see whether increased formation of etheno-DNA adducts occurs. Using an ultra sensitive and specific immunoaffinity/(32)P-postlabelling method, 1, N(6) ethenodeoxyadenosine (straightepsilondA) and 3, N(4)-ethenodeoxycytidine (straightepsilondC) were quantitated in epithelial cell DNA from asymptomatic colon, FAP polyps and colon tumor tissues. Mean adduct levels in FAP polyps were 65 straightepsilondA/10(9) and 59 straightepsilondC/10(9) parent nucleotides, being 2 to 3 times higher than in unaffected colon tissue (p < 0.02 for straightepsilondA; p < 0.05 for straightepsilondC). Adduct levels in colonic epithelia decreased in the order: FAP polyps > tumor-adjacent tissue > tumor, normal and tumor-distal tissue. Based on this study, requiring confirmation in a larger number of patients and in experimental models, we have demonstrated the formation of promutagenic etheno-DNA adducts in adenomatous polyps of FAP patients that may contribute to genetic instability and cancer progression. PMID- 10861446 TI - Down-regulation of drs mRNA in human colon adenocarcinomas. AB - We have previously reported that the drs gene, whose mRNA expression is down regulated by retroviral oncogenes such as v-src and v-K-ras, has the ability to suppress transformation by v-src and v-K-ras in the rat cell line F2408. We have also isolated a human homolog of this gene (h-drs) and shown that expression of h drs mRNA is markedly reduced in a variety of human cancer cell lines, including those of carcinomas of the colon, bladder, and ovary, suggesting that down regulation of drs mRNA is correlated with the development of human cancers. To clarify the correlation between down-regulation of the drs gene and malignant tumor formation in human cancer tissues, we examined expression of drs mRNA in human normal tissues, colon adenoma, and adenocarcinoma tissues by in situ hybridization. Expression of drs mRNA was detected in most normal tissues tested, including those of the colon, bladder, and ovary. However, drs mRNA was hardly expressed in any of the colon adenocarcinoma tissues examined. Northern blot analyses confirmed these results. Neither gross deletion nor re-arrangement of the drs genome was detected by Southern blot hybridization in these adenocarcinoma tissues. drs mRNA was significantly expressed in colon adenoma with mild atypia but down-regulated in adenomas with moderate atypia and focal carcinoma. Our results indicate that down-regulation of drs mRNA is closely correlated with development of colon adenocarcinoma, suggesting a tumor suppressor function of the drs gene in human cancers. PMID- 10861447 TI - Expression of integrin alpha(v)beta(3) correlates with activation of membrane type matrix metalloproteinase-1 (MT1-MMP) and matrix metalloproteinase-2 (MMP-2) in human melanoma cells in vitro and in vivo. AB - Activation of matrix metalloproteinase-2 (MMP-2) is mediated by binding to the complex of membrane-type matrix metalloproteinase-1 (MT1-MMP) with tissue inhibitor of MMP-2 (TIMP-2) on the cell surface. Binding of MMP-2 to integrin alpha(v)beta(3) has been implicated in presenting activated MMP-2 on the cell surface of invasive cells, but interactions with the MT1-MMP-TIMP-2 system have not been considered. Therefore, we studied the expression and interaction of MT1 MMP, MMP-2 and TIMP-2 in the alpha(v)beta(3)-negative melanoma cell line BLM and in its beta(3)-transfected, alpha(v)beta(3)-expressing counterpart BLM-beta(3), both on cell lines and in xenografts. Total expression levels of MMP-2, MT1-MMP and TIMP-2 did not differ markedly between the alpha(v)beta(3)-negative and alpha(v)beta(3)-positive cells. Remarkable differences, however, exist in the presence of active MMP-2 and MT1-MMP. Zymography on cell lysates revealed that active MMP-2 was restricted to alpha(v)beta(3)-positive cell line and clearly accumulated in xenografts derived from the BLM-beta(3) cells, confirming the relevance of this integrin for MMP-2 function. Western blotting of cell lysates showed that processing of proMT1-MMP to the activated form was enhanced in BLM beta(3). The ratio of active and inactive MT1-MMP was 3-fold higher in the beta(3)-transfectants. Immunofluorescence double-labeling followed by confocal laser microscopy showed co-localization of MT1-MMP and alpha(v)beta(3) on BLM beta(3) cells. In xenografts from BLM-beta(3) cells, active MT1-MMP was markedly increased. Our results demonstrate that expression of alpha(v)beta(3) in cell lines and xenografts was accompanied by an accumulation of active MT1-MMP and MMP 2. Furthermore, MT1-MMP and alpha(v)beta(3) are co-localized on the cell membrane of tumor cells. These findings suggest that activated MT1-MMP co-localized with alpha(v)beta(3) may be involved in activation of alpha(v)beta(3)-bound MMP-2. PMID- 10861448 TI - Transforming growth factor-alpha-mediated growth pathways in human gastro intestinal cell lines in relation to the gastrin autocrine pathway. AB - Epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) increase transcription of the gastrin gene, and the gastrin peptide may be phosphorylated by EGF-stimulated tyrosine kinase. Our aims were to compare EGF/TGF-alpha interactions in 2 human gastro-intestinal cell lines: MGLVA1, with a strong gastrin autocrine pathway, and C170HM2, with a weak pathway. Both cell lines expressed the TGF-alpha gene. MGLVA1 expressed TGF-alpha protein as determined by immuno-cytochemistry, which was absent in C170HM2. Both cell lines expressed the same level of EGF receptors, as assessed by flow cytometry; however, MGLVA1 did not have enhanced in vitro proliferation in response to EGF or TGF-alpha, unlike C170HM2. The basal growth of MGLVA1 was inhibited by anti sera against TGF-alpha, the EGF receptor and G17. C170HM2 was not inhibited by any of the anti-sera. Neutralisation of TGF-alpha resulted in undetectable cell associated progastrin levels in MGLVA1 (untreated had 391.7 fmol/5 x 10(6) cells). The progastrin level of C170HM2 remained unaffected. Tyrosine kinase activity, as assessed by phosphopeptide concentration, of unstimulated MGLVA1 was 2.6 times higher than that of C170HM2 in the cell membrane fraction (0.097 compared to 0.037 microg/mg protein, p < 0.001) and 4.8 times higher in the cytosolic fraction (0.269 compared to 0.056 microg/mg protein, p < 0.05). Following treatment with EGF, the phosphopeptide concentration increased in both the membrane and cytosolic fractions of both cell lines. Tyrphostin B42, which inhibits autophosphorylation of the EGF receptor, inhibited the basal growth of MGLVA1 (IC(50) 1.3 microM) and C170HM2 (9.5 microM, p < 0.05 from MGLVA1). Herbimycin, which inhibits pp60(c-src) kinase, reduced the basal growth of MGLVA1 (0.67 microM) but not C170HM2. Immunofluorescence studies confirmed the presence of tyrosine-phosphorylated proteins and pp60(c-src) within the cytoplasm of unstimulated MGLVA1 cells. There was no specific immunofluorescence for either parameter in C170HM2 cells until after treatment with EGF. PMID- 10861449 TI - Expression of HER/erbB family of receptor tyrosine kinases and induction of differentiation by glial growth factor 2 in human rhabdomyosarcoma cells. AB - Five human rhabdomyosarcoma cell lines were used to investigate the surface expression of HER/erbB receptors, particularly of HER-2, HER-3 and HER-4, by flow cytometry. HER-2 was expressed in 3/5 rhabdomyosarcoma cell lines. HER-3 was expressed in all rhabdomyosarcoma cell lines. None of the rhabdomyosarcoma cell lines investigated showed HER-4 surface expression. To study the biological activity of HER/erbB receptors in rhabdomyosarcomas, cells were cultured in the presence of glial growth factor 2 (GGF-2), a specific ligand of HER-3 that is able to stimulate myogenesis in normal myoblasts. In 3 of 3 human rhabdomyosarcoma cell lines positive for both HER-2 and HER-3 receptors, the treatment with GGF-2 induced a significant increase in myogenic differentiation. PMID- 10861450 TI - Expression, subcellular localization and putative function of platelet-type 12 lipoxygenase in human prostate cancer cell lines of different metastatic potential. AB - The involvement of 12-lipoxygenase (12-LOX) expression and function in tumor metastasis has been demonstrated in several murine tumor cell lines. In addition, 12-LOX expression was detected in human prostatic tumors and correlated to the clinical stage of disease. Here we provide data that human prostate cancer cell lines express the platelet-type isoform of 12-LOX at both the mRNA and protein levels, and immunohistochemistry revealed 12-LOX expression in human prostate tumors. The enzyme was localized to the plasma membrane, cytoplasmic organelles and nucleus in non-metastatic cells (PC-3 nm) and to the cytoskeleton and nucleus in metastatic cells (DU-145). After orthotopic/intraprostatic injection of tumor cells into SCID mice, the metastatic prostate carcinoma cells (DU-145) expressed 12-LOX at a significantly higher level compared with the non-metastatic counterparts, PC-3nm. The functional involvement of 12-LOX in the metastatic process was demonstrated when DU-145 cells were pretreated in vitro with the 12 LOX inhibitors N-benzyl-N-hydroxy-5-phenylpentamide (BHPP) or baicalein, the use of which significantly inhibited lung colonization. These data suggest a potential involvement of 12-LOX in the progression of human prostate cancer. PMID- 10861452 TI - MAGE-B5, MAGE-B6, MAGE-C2, and MAGE-C3: four new members of the MAGE family with tumor-specific expression. AB - A number of genes of the MAGE-A, B, and C families have been shown to code for antigens that are recognized on many human tumors by autologous cytolytic T lymphocytes. These antigens ought to be strictly tumor specific because the encoding MAGE genes are not expressed in normal adult cells except for male germline cells, which lack HLA expression. To identify new genes of this type, we performed representational difference analysis on a melanoma cell line by subtraction with a normal skin sample. This led to the identification of MAGE-C2, a new member of the MAGE-C family. A search for nucleotide sequences encoding MAGE-like proteins in public databases led to the identification of three additional MAGE genes, which were named MAGE-B5, MAGE-B6, and MAGE-C3. The four new MAGE genes are not expressed in normal tissues, except for testis, and are expressed in tumors of different histological origins. Therefore, like other MAGE genes expressed specifically in tumors, MAGE-B5, MAGE-B6, MAGE-C2, and MAGE-C3 ought to encode antigens that could be targets for cancer immunotherapy. PMID- 10861451 TI - Identification and characterization of a Kunitz-type protease inhibitor in ascites fluid from patients with ovarian carcinoma. AB - Urinary trypsin inhibitor (UTI; Mr 40 kDa) is a Kunitz-type protease inhibitor that efficiently inhibits cell-associated trypsin and plasmin activities. The aim of this study is to examine the expression pattern of UTI in the human ovarian carcinoma ascites fluid by Western blotting, zymography, immunoprecipitation, immunohistochemistry, biochemical and gene analyses and animal experiments. We have identified and characterized the 40 kDa immunoreactive UTI (UTI(40)) and 8 kDa degradation fragment (UTI(8)) in ascites fluid. The levels of UTI(40) and UTI(8) are elevated in ascites fluid taken from patients with ovarian carcinoma relative to paired plasma samples. The UTI(40) and UTI(8) were identified immunologically by the reactivity with 2 different anti-UTI antibodies recognizing different epitopes of the UTI molecule, functionally by its ability to bind trypsin and structurally by its apparent molecular mass with and without deglycosylation treatment. The purified polypeptides have been sequenced and were identical with sequences obtained from UTI and the carboxyl-terminal domain of UTI, respectively. However, UTI mRNA was not detected in the ovarian carcinoma tissue and ovarian carcinoma cell lines examined. Based on extravasation experiments using intravenously injected biotinylated inter-alpha-trypsin inhibitor (IalphaI; a precursor of UTI), we conclude that UTI(40) and UTI(8) found in the ascites fluid may result from (i) the extravasation of plasma proteins such as IalphaI into the peritoneal cavity via hyperpermeable vessels and (ii) the subsequent degradation of IalphaI and UTI(40) by tumor cell associated trypsin-like enzymes. PMID- 10861453 TI - SSA/RO52gene and expressed sequence tags in an 85 kb region of chromosome segment 11p15.5. AB - Frequent allelic loss in lung cancer has been described in a region on chromosome segment 11p15.5 (LOH11A). The region is approximately 650 kb in size and flanked by the markers D11S988 centromeric and D11S860 telomeric. Clinical and cell biological studies suggest that it contains a gene associated with metastatic tumor spread. One of the genes identified within this region is SSA/Ro52, which has a RING finger domain and may be involved in gene regulation. We studied this gene for mutations using SSCP analysis and for expression using RT-PCR and Western blotting on lung cancer cell lines and tumor-normal tissue pairs. No mutations and no differences in mRNA or protein expression between tumor tissue and normal tissue pairs were identified. We discovered a novel polymorphic site (SSA44C/T) within exon 1 of this gene. Among 141 primary lung cancers, allelic loss was observed in 16% of informative cases. Our analyses excluded SSA/Ro52 as a tumor-suppressor gene in lung cancer and newly defined the centromeric border of the LOH11A region from D11S988 previously to SSA44C/T. This reduced the region of the putative suppressor gene to 460 to 485 kb. A significant difference (p = 0.01) in the frequency of alleles for this polymorphism between Caucasians and African-Americans was observed. The "T" allele frequency was 0.12 in Caucasians and 0.23 in African-Americans. A genomic EcoRI map over 85 kb surrounding the SSA/Ro52 gene was constructed, and 4 expressed sequence tags were identified by sequencing and studied. PMID- 10861454 TI - A tumor suppressor locus in familial and sporadic chordoma maps to 1p36. AB - Previous cytogenetic/FISH data have demonstrated 1p36 deletions in a relapsing familial clivus chordoma developed by a patient who has 2 daughters, respectively affected with childhood astrocytoma and clivus chordoma. Using an approach that combined the LOH (loss of heterozygosity) study of the father chordoma and the daughter astrocytoma and a segregation analysis from parents to sibs using 17 CA repeats spanning 1p36.32-1p36.11, we mapped the cancer susceptibility locus in this family to the 1p36 region. The LOH and haplotype information was elaborated using a pairwise linkage analysis that gave a maximum lod score of 1.2. Additional LOH data relating to 6 sporadic chordomas allowed us to define an SRO (the smallest region of overlapping loss) of about 25 cM from D1S2845 (1p36.31) to D1S2728 (1p36.13). Our overall findings converge on mapping to 1p36 a tumor suppressor gene involved in familial and sporadic chordoma. PMID- 10861455 TI - p53 protein accumulation and mutations in normal and benign breast tissue. AB - Mutations in the p53 gene are amongst the most common molecular changes detected in breast cancer, and there are several reports suggesting that changes in p53 may contribute to the pathogenesis of this disease. In a previous case-control study, we demonstrated that p53 protein accumulation detected by immunohistochemistry in normal or benign breast tissue was associated with a 2.5 fold increase in the risk of subsequent breast cancer. In this study, we investigated whether p53 gene mutations were present in the 29 p53 immunopositive normal or benign breast tissue samples and in 15 p53 immunonegative normal or benign breast tissue samples selected randomly from the original study. DNA was extracted from paraffin sections and underwent PCR-SSCP analysis for exons 4 to 10. PCR products that showed abnormal mobility were excised and sequenced. Sixteen (59.2%) of the 27 immunopositive breast tissue samples and 4 (26.7%) of the 15 immunonegative samples had p53 sequence changes. There was no obvious association between the occurrence of these alterations and any specific histopathologic features. Ten cases showed p53 mutations, and they were all missense base substitutions of the transition type. Thirteen other gene changes occurred in 11 breast tissue samples and consisted of 8 silent (no amino acid change), 4 intronic alterations, and 1 indeterminate alteration. One individual had both a mutation and a silent change. In summary, p53 gene alterations can occur in normal or benign breast tissue, but resolution of their role in the pathogenesis of breast cancer will require long-term follow-up studies involving comparisons of breast cancer occurrence in patients with and without p53 mutations as well as functional assays to determine their significance. PMID- 10861456 TI - Immunization of ovarian cancer patients with a synthetic Lewis(y)-protein conjugate vaccine: a phase 1 trial. AB - As the initial step in developing carbohydrate-based vaccines for the treatment of ovarian cancer patients in an adjuvant setting, 25 patients were immunized with a Lewis(y) pentasaccharide (Le(y))-keyhole limpet hemocyanin (KLH)-conjugate vaccine together with the immunological adjuvant QS-21. Four different doses of the vaccine, containing 3, 10, 30, and 60 microg of carbohydrate were administered s.c. at 0, 1, 2, 3, 7, and 19 weeks to groups of 6 patients. Sera taken from the patients at regular intervals were assayed by ELISA for reactivity with naturally occurring forms of Le(y) (Le(y)-ceramide and Le(y) mucin) and by flow cytometry and a complement-dependent cytoxicity assay for reactivity with Le(y)-expressing tumor cells. The majority of the patients (16/24) produced anti Le(y) antibodies as assessed by ELISA, and a proportion of these had strong anti tumor cell reactivity as assessed by flow cytometry and complement-dependent cytotoxicity. One serum, analyzed in detail, was shown to react with glycolipids but not with glycoproteins or mucins expressed by ovarian cancer cell line OVCAR 3. The vaccine was well tolerated and no gastrointestinal, hematologic, renal, or hepatic toxicity related to the vaccine was observed. On the basis of this study, Le(y)-KLH should be a suitable component for a polyvalent vaccine under consideration for the therapy of epithelial cancers. PMID- 10861457 TI - Apoptosis induced by immunotoxins used in the treatment of hematologic malignancies. AB - The recombinant immunotoxins anti-Tac(Fv)-PE38 (LMB-2), targeting the interleukin 2 receptor alpha subunit (IL-2Ralpha, Tac or CD25), and RFB4(dsFv)-PE38 (BL22), targeting CD22, are being evaluated in clinical trials as treatment for hematologic malignancies. The toxin moiety Pseudomonas exotoxin A (PE) of these recombinant molecules leads to the arrest of protein synthesis due to inactivation of elongation factor 2. Here, we provide evidence that cell lines derived from patients with hematologic malignancies react to immunotoxins not only with inhibition of protein synthesis but also with characteristic hallmarks of apoptosis such as caspase activation, cleavage of the "death substrate poly(ADP)-ribose polymerase and DNA laddering. Anti-Tac(Fv)-PE38 leads to a 10 fold increase in the cleavage of the fluorescent substrate DEVD-AFC, suggesting that a caspase-3-like enzyme is involved. This was verified by cleavage of caspase-3 (CPP32). MT1 cells exhibited DNA laddering after treatment with immunotoxin, which was reversed by pre-treatment with the protease inhibitor zVAD fmk. This caspase inhibitor led to an at least 5-fold improvement in cell viability without altering inhibition of protein synthesis. Interestingly, HUT 102 cells did not undergo programmed cell death after exposure to immunotoxins that kill these cells. We conclude that immunotoxins may be valuable in the treatment of cancers that are resistant toward apoptosis because their targeted killing is often facilitated by, but not completely dependent on, programmed cell death. Int. J. Cancer 87:86-94, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861458 TI - Effect of the specific cyclooxygenase-2 inhibitor meloxicam on tumour growth and cachexia in a murine model. AB - The effects of the cyclooxygenase-2 (COX-2) inhibitor, meloxicam, on tumour growth and cachexia have been determined in 2 established murine adenocarcinomas (MAC). At a dose level of 2.5 and 5.0 mgkg(-1), meloxicam produced pronounced inhibition of the growth of the MAC13 tumour, increasing the tumour volume doubling time from 2 to 5 days. Meloxicam also suppressed growth of the MAC16 tumour, which is generally refractory to standard cytotoxic agents, increasing the tumour volume doubling time from 1.5 to 2.5 days at dose levels of 0.5 and 1.0 mgkg(-1). Cachexia was also effectively attenuated at these dose levels. To investigate whether meloxicam exerted a direct effect on the cachectic process, studies on protein degradation were carried out using C(2)C(12) mouse myoblasts in response to a proteolysis-inducing factor (PIF). PIF produced maximum protein degradation at a concentration of 4.2 nM, and this was effectively attenuated by meloxicam at concentrations greater than 1 microM. This suggests that meloxicam may be capable of directly antagonizing the process of muscle catabolism in cachexia. PMID- 10861459 TI - Protective specific immunity induced by doxorubicin plus TNF-alpha combination treatment of EL4 lymphoma-bearing C57BL/6 mice. AB - The therapeutic efficacy of a single (day 8), moderate dose (4 mg/kg, i.v.) of doxorubicin (DOX, Adriamycin) combined with recombinant human TNF-alpha (3 different doses and 5 different schedules, i.v.) was evaluated in C57BL/6 mice bearing an implant (s.c.) of the DOX-sensitive, TNF-alpha-resistant EL4 lymphoma. In parallel to monitoring survival, the levels of several host anti-tumor cytolytic effector functions of splenocytes and thymocytes were evaluated throughout the treatment period and in long-term survivors (LTS). DOX treatment alone resulted in a moderate (approx. 20%) increase in life span but no cures. TNF-alpha alone, at any tested dose or schedule, had little or no positive effect on survival. The combinations of DOX and TNF-alpha were only slightly better than DOX alone with respect to the time to death of mice that died (approx. 29% increase); however, each of the combinations involving 1,000 U TNF alpha/injection produced a fraction (20% to 80%) of LTS. The host defense activities examined included those of splenic and thymic cytolytic T lymphocytes (CTL) and lymphokine-activated killer cells as well as splenic tumoricidal macrophages. Although most activities were modulated by tumor growth and/or treatment, only CTL responsiveness appeared to correlate with survival. CTL activity in the treated groups with LTS was significantly higher than in control groups late in the treatment period. Finally, ex vivo analyses of splenocytes and thymocytes together with the rejection of implanted tumor at 17 months established that LTS displayed specific long-term immune memory. PMID- 10861460 TI - Ovarian and breast cancer risks to women in families with two or more cases of ovarian cancer. AB - There are few published estimates of the risk of developing breast or ovarian cancer in women with a strong family history of ovarian cancer. As these women commonly present to family cancer clinics, accurate cancer risk estimates are needed. We have estimated these risks in women from families with 2 or more confirmed ovarian cancers in first-degree relatives using data from the UKCCCR Familial Ovarian Cancer Register. The number of cancers observed in more than 10,000 person years of follow-up was compared with the number expected based on national-, age-, sex- and period-specific incidence rates. The relative risk of ovarian cancer was found to be 7.18 (95% CI 3.82-12.3), declining from 16.0 (6.40 32.9) in women under 50 to 4.38 (1.60-9.52) in women 50 years of age and older. For breast cancer, the relative risk for women under 50 was 3.74 (2.04-6. 28) and 1.79 (1.02-2.90) for women 50 years of age and older (average RR 2.36, 1.59 3.37). These correspond to absolute risks by age 70 of 11% for ovarian cancer and 15% for breast cancer. When the analyses were restricted to families that had been negative for mutations in the breast/ovarian cancer susceptibility genes, BRCA1 and BRCA2, the ovarian cancer risk was 11.59 (3.12-29.7) and that of breast cancer 3.32 (1.52-6.31). As well as having clinical relevance, our finding may suggest that other breast/ovarian cancer genes are segregating in these families, though the possibility of undetected BRCA1/2 mutations must also be considered. PMID- 10861461 TI - Insulin-like growth factor 1 in hepatocellular carcinoma and metastatic liver cancer in men. AB - The insulin-like growth factor (IGF) axis has important autocrine, paracrine, and endocrine roles in the promotion of growth. Alterations of the IGF system have recently been implicated in the pathogenesis of several malignancies, but the relation to hepatocellular carcinoma (HCC) risk is unclear. To address this issue, we used an immunoradiometric assay to quantify IGF-1 levels in serum samples in a hospital-based, case-control study in Greece. The study subjects were all men and included 53 patients with HCC positive for hepatitis B and/or hepatitis C virus infections, 20 virus-negative HCC patients, 25 virus-negative patients with metastatic liver cancer (MLC), and 111 virus-negative control subjects. Data were analyzed by multiple linear regression, using IGF-1 as the dependent variable. The mean value of IGF-1 was 65.9 ng/ml among virus-positive HCC patients, 79.5 ng/ml among virus-negative HCC patients, 110.8 ng/ml among patients with MLC, and 174.7 ng/ml among hospital controls. After controlling for the degree of liver damage, as assessed by prothrombin time and serum albumin level, the reduction in IGF-1 level among HCC patients was found to be more than could be attributed to liver damage alone. This finding may have both diagnostic and pathophysiological implications. PMID- 10861462 TI - Projections of alcohol- and tobacco-related cancer mortality in Central Europe. AB - Central European mortality rates for cancer sites related to tobacco and alcohol have increased rapidly in recent decades. From a public health point of view, it is of considerable interest to know whether these past increases in cancer mortality will continue into the future. Cancer mortality rates for the period 1965-1994 in Bulgaria, Czech Republic and Slovakia (analysed together), Hungary, Poland, and Romania were analysed for cancers of the larynx, oral cavity and pharynx, oesophagus, bladder, kidney, and pancreas. Using a Bayesian age-period cohort approach, we have calculated smoothed observed rates. The effects of period and cohort were extrapolated to estimate mortality projections for 1995 99, 2004-09, and 2005-09. Mortality rates for all sites are projected to increase in most countries. Hungary has the highest projected rates for most sites, and particularly rapid increases are expected for cancers of the oral cavity and pharynx and of the larynx in Hungarian men. The smoothed 1990-94 male mortality rates for these two sites of 16. 32/100,000 and 8.70/100,000, respectively, are projected to reach 35. 17/100,000 for cancer of the oral cavity and pharynx and 14.12/100, 000 for cancer of the larynx by the period 2000-04. For kidney cancer, former Czechoslovakia has the highest observed and projected mortality rates. The smoothed 1990-94 rate of 8.37/100,000 is expected to increase 24% to 10.38/100,000 by 2000-04. Our results indicate that further increases may be expected on top of the already high cancer mortality levels in Central Europe. Policies to reduce alcohol consumption and prevent smoking in younger generations are necessary to reduce mortality as these cohorts age. PMID- 10861463 TI - Plant foods and risk of laryngeal cancer: A case-control study in Uruguay. AB - In order to examine the relationships between plant foods, defined as the grouping of vegetables, fruits, tubers and legumes, with the risk of developing laryngeal cancer, a case-control study was conducted in Uruguay between 1998 1999. The study included 148 cases with histologically verified squamous cell carcinoma of the larynx, which were frequency matched on age, residence and urban/rural status with 444 hospitalized controls, afflicted by non-neoplastic conditions. Both series of patients were face-to-face interviewed in the hospitals shortly after admittance using a detailed questionnaire. This questionnaire included 62 queries on food items, representative of the usual diet of the Uruguayan population. Food items and food groups were adjusted for tobacco smoking, alcohol drinking and total energy intake. High consumption of plant foods was associated with an OR of 0.42 (95% CI 0.21-0.84). Among subgroups of plant foods, fruits and raw vegetables were associated with a strong reduction in risk (OR for the highest quartile of raw vegetables 0.29, 95% CI 0.15-0.56). Also, legumes were associated with a protective effect (OR 0.62, 95% CI 0.33 1.19). Among individual food items, tomatoes and oranges were associated with the stronger protective effects (OR for tomato intake 0.32, 95% CI 0. 17-0.58). The joint effect of heavy smoking and the low intake of vegetables and fruits displayed an increased risk of 19.2 (95% CI 5. 7-64.9). PMID- 10861464 TI - Dietary antioxidant intake and the risk of cardia cancer and noncardia cancer of the intestinal and diffuse types: a population-based case-control study in Sweden. AB - In spite of diverging incidence trends, subsite, and subtype-specific gastric cancer data on the association with dietary antioxidants are sparse. We aimed to test whether the apparent protective effect of antioxidants is mainly confined to noncardia (distal) cancer of the intestinal subtype, to which most of the incidence decline in gastric cancer has been ascribed. In a Swedish study base (total population 1.3 million), we interviewed 567 cases uniformly classified to subsite (cardia vs. noncardia) and subtype (intestinal vs. diffuse), and 1165 population-based controls, frequency matched for age and sex. Serologic data on H. pylori status was available for a subset of 542 individuals. Ascorbic acid (vitamin C) was inversely associated with all subsites and subtypes of gastric cancer in a significant dose-response manner (all p<0.05), with risk reductions between 40% and 60%. beta-carotene was also strongly and negatively associated with risk, particularly with the intestinal type. The associations with alpha tocopherol (vitamin E) were less clear. The highest parallel intake of all three antioxidants (quartiles 4), compared to those with the lowest parallel intakes (quartiles 1), was associated with a 70% lower risk of developing noncardia cancer (OR 0.3, 95% CI 0.1-0.9). Our results suggest that antioxidants might be especially beneficial among subjects at increased risk for gastric cancer such as smokers and those infected by H. pylori. We conclude that a high intake of antioxidants, as a consequence of high consumption of fruit and vegetables, may lower the risk not only for gastric cancer of the intestinal type, but also for diffuse type adenocarcinoma and cardia cancer. PMID- 10861465 TI - Cigarette smoking and risk of prostate cancer in the physicians' health study (United States). AB - To assess whether cigarette smoking is associated with prostate cancer incidence or mortality, we analyzed a large cohort of 22,071 men, aged 40-84 at baseline, in the Physicians' Health Study. During an average of 12.5 years of follow-up, we documented 996 cases of prostate cancer, including 113 fatal cases. Men were categorized according to smoking status, total pack-years smoked, and duration of smoking. We used Cox proportional hazard models to estimate the relative risks associated with smoking. Compared to never smokers, the age-adjusted relative risks (RR) of total prostate cancer were 1. 14 (95% confidence interval [CI] = 1.00-1.30) for past smokers, 1.10 (95% CI = 0.78-1.55) for current smokers of less than 20 cigarettes per day, and 1.10 (95% CI = 0.84-1.44) for current smokers of 20 or more cigarettes per day. Adjustment for body mass index, height, alcohol intake, and physical activity did not materially alter these findings. No significant association was observed in analyses of total pack-years smoked or duration of smoking. The results were similar for non-fatal and fatal prostate cancer. These data indicate no material association between cigarette smoking and prostate cancer incidence or mortality. PMID- 10861466 TI - Sunscreen use and malignant melanoma. AB - In a new population-based, matched, case-control study from southern Sweden of 571 patients with a first diagnosis of cutaneous malignant melanoma, between 1995 and 1997, and 913 healthy controls aged 16 to 80 years, the association between sunscreen use and malignant melanoma was evaluated. The median sun protection factor (SPF) used by both cases and controls was 6, range 2 to 25. Sunscreen users reported greater sun exposure than non-users. Persons who used sunscreens did not have a decreased risk of malignant melanoma. Instead, a significantly elevated odds ratio (OR) for developing malignant melanoma after regular sunscreen use was found, adjusted for history of sunburns, hair color, frequency of sunbathing during the summer, and duration of each sunbathing occasion ?OR = 1.8, 95% confidence interval (CI) 1.1-2.9]. The OR was higher in subjects who reported that sunscreen use enabled them to spend more time sunbathing (adjusted OR = 8.7, 95% CI 1.0-75.8 for always vs. never use). The association appeared to hold for subjects who did not suffer from sunburns while using sunscreens, for subjects who used SPF of 10 or lower, and among men. The pattern of a significantly increased melanoma risk was seen only for lesions of the trunk. Our results are probably related mainly to earlier sunscreens of low SPF. They substantiate the hypothesis that sunscreen use, by permitting more time sunbathing, is associated with melanoma occurrence. PMID- 10861467 TI - Mutation analysis of the PTEN gene in uveal melanoma cell lines. PMID- 10861468 TI - Activated K-ras is involved in regulation of integrin expression in human colon carcinoma cells. AB - Integrins participate in controlling proliferation and migration. Therefore, changes in integrin expression might be responsible for unrestrained proliferation and invasiveness of tumor cells. Alterations of integrin subunit expression have been observed in human colon carcinoma, especially loss or reduction of the alpha5 subunit, which was observed consistently. The mechanisms responsible for reduction of alpha5 expression and alteration of expression of other integrins are not fully understood. Circumstantial evidence from previous investigations points to an involvement of activated ras oncogenes in repression of integrin expression. The K-ras protooncogene is activated by point mutation in 50% of human colon carcinomas. Thus, we choose an antisense approach for specific inactivation of activated K-ras in the human colon carcinoma cell line SW 480 in order to test whether activated K-ras contributes to changes in integrin expression on colon carcinoma cells. Cell surface expression of the alpha1 and the alpha5 subunit was increased in K-ras antisense transfected clones, cell surface expression of the alpha3 subunit and the alphav subunit was decreased. This shows, in a human system, that activated K-ras is involved in diminishing cell surface expression of the alpha1beta1 collagen/laminin receptor and the alpha5beta1 fibronectin receptor, both of which are implicated in maintenance of a non-transformed phenotype. Moreover, activated K-ras contributes to increased cell surface expression of the alpha3beta1 laminin/collagen/fibronectin receptor and the alphavbeta5 vitronectin receptor, which might play a role in metastatic behavior of tumor cells. PMID- 10861469 TI - Activation of fibroblast-derived matrix metalloproteinase-2 by colon-cancer cells in non-contact Co-cultures. AB - Stromal fibroblasts interact with invading cancer cells by secreting and activating matrix metalloproteinases (MMPs). To elucidate the mechanisms involved in the expression and activation patterns of MMPs, human colon-cancer cell lines Caco-2 and LoVo and colon-fibroblast cell line CCD18-Co were co-cultivated in non contact and contact conditions which mimic in vivo interaction between cancer cells and fibroblasts before and after cancer invasion respectively. Gelatin zymography disclosed that MMP-2 was secreted from the fibroblasts but not from the cancer cells. The quantity of fibroblast-derived MMP-2 in conditioned medium was not significantly changed in either the contact or the non-contact co cultures when compared with that of individual cultures of CCD18-Co fibroblasts. Cancer cells in non-contact co-cultures, however, enhanced the activation of fibroblast-derived MMP-2. Transcripts of membrane-type matrix metalloproteinase-1 (MT1-MMP), which is thought to be present on the cell surface and to work as a candidate activator of MMP-2, were detected in both cancer cell lines. Plasma membrane extracts of cancer cells also activated MMP-2 in conditioned media in cell-free conditions. This activation of MMP-2 may be caused by MT1-MMP of the cancer cells, since it was inhibited by a series of MMP inhibitors, including ethylenediaminetetraacetic acid (EDTA), the tissue inhibitor of metalloproteinase 2 (TIMP-2), and the MMP inhibitor CGS 27023A, but not by TIMP-1. Our data demonstrate that in non-contact co-cultures colon-cancer cells activate fibroblast-derived MMP-2 on their plasma membranes. These findings should help to elucidate the mechanism involved in the initial destruction of basement membrane by cancer cells. PMID- 10861470 TI - Differential expression of sphingolipids in MRP1 overexpressing HT29 cells. AB - We have obtained a novel multidrug resistant cell line, derived from HT29 G(+) human colon carcinoma cells, by selection with gradually increasing concentrations of the anti-mitotic, microtubule-disrupting agent colchicine. This HT29(col) cell line displayed a 25-fold increase in colchicine resistance and exhibited cross-resistance to doxorubicin, VP16, vincristine and taxol. Immunoblotting, combined with RT-PCR showed that the multidrug resistance phenotype was conferred by specific overexpression of the multidrug resistance protein 1. Confocal scanning laser microscopy revealed that multidrug resistance protein 1 specifically localized in the plasma membrane of HT29(col) cells. In a functional assay, using the fluorescent multidrug resistance protein 1 substrate 5-carboxyfluorescein, an increased efflux activity of HT29(col) cells was measured, as compared to the wild-type HT29 G(+) cells. MK571, a specific inhibitor of multidrug resistance protein 1, blocked the 5-carboxyfluorescein efflux, but only partially reversed resistance to colchicine, indicating that additional multidrug resistance mechanisms operate in HT29(col) cells. In conclusion, these results show for the first time overexpression of a functional multidrug resistance protein 1 under colchicine pressure, indicating that colchicine is not a P-glycoprotein-specific substrate. Colchicine-induced overexpression of multidrug resistance protein 1 is accompanied by a changed sphingolipid composition, i.e., enhanced levels of glucosylceramide and galactosylceramide. In addition, ceramide, a lipid messenger molecule involved in apoptosis-related signal transduction processes, was much more abundant in HT29(col) cells, which is indicative of a stress response. PMID- 10861471 TI - PAX6 methylation and ectopic expression in human tumor cells. AB - The mechanisms underlying the de novo methylation of CpG islands in human cancer remain almost completely unknown. We used a methylation-sensitive arbitrarily primed polymerase chain reaction (Ms AP-PCR) technique to scan genomic DNA for differential methylation patterns and identified a 550 bp band that was hypermethylated in a majority of colon and bladder cancer cells. This band corresponded to a CpG-rich region in exon 5 of PAX6, which is a highly conserved transcription regulatory factor involved in embryogenesis. Interestingly, exon 5 was very frequently methylated in solid tumors compared with adjacent normal tissues, 17 out of 27 (63%) in bladder, and colon, but the promoter remained unmethylated in all these cases. This methylation was not effective in blocking transcription since ectopic expression of PAX6 was seen in several tumors and cell lines with extensive exon 5 methylation. De novo methylation of the promoter was only seen in tumor cell lines and was associated with gene silencing since treatment with 5-aza-2'-deoxycytidine restored expression to the cells and resulted in a less methylated promoter. Thus, ectopic expression and hypermethylation of exon 5 of PAX6 demonstrate that methylation within a transcribed region, as opposed to promoter methylation, does not block gene expression. PMID- 10861472 TI - p185(neu) protein is required for tumor and anchorage-independent growth, not for cell proliferation of transgenic mammary carcinoma. AB - Transgenic FVB-NeuN mice (N202) bearing the rat neu protooncogene driven by the mouse mammary tumor virus promoter/enhancer develop focal mammary carcinomas overexpressing the neu-encoded p185(neu) protein. In vitro expression of p185(neu) among mammary carcinoma cultures was heterogeneous, and we could establish some cell lines and clones displaying a complete loss of p185(neu) expression, along with others with very high p185(neu) protein level. Upon in vivo injection, p185(neu)-positive cells gave rise to fast-growing tumors with a short latency, while p185(neu)-negative cells required a very long latency time, and the resulting tumors were invariably p185(neu)-positive. The lower growth ability of p185(neu)-negative cells in vivo was also confirmed in athymic nude mice. In vitro, analysis of anchorage-independent growth in soft agar revealed colony formation from p185(neu)-positive but not p185(neu)-negative cells. The direct involvement of p185(neu) in clonogenicity was demonstrated by the inhibition of p185(neu)-positive colony growth in soft agar in the presence of an anti-p185(neu) monoclonal antibody. By contrast, a higher level of anchorage dependent clonogenic growth and proliferation was observed in p185(neu)-negative cells as compared to p185(neu)-positive cells, thus explaining the relative ease with which p185(neu)-negative cell lines and clones were established in vitro. Together, the results indicate that p185(neu) expression can lead to tumor formation and metastasis through the modification of intrinsic properties of cells related to anchorage-independent growth ability rather than to proliferation or host-dependent mechanisms. PMID- 10861473 TI - Low frequency of HLA-A*0201 allele in patients with Epstein-Barr virus-positive nasal lymphomas with polymorphic reticulosis morphology. AB - Lymphoproliferative diseases of the nasal cavity and paranasal sinuses occur frequently in Asian countries and are histologically categorized as monomorphic ordinary lymphoma and polymorphic reticulosis (PR) with apparent inflammatory cell infiltration. The large atypical cells in PR show natural-killer cell nature and frequently contain Epstein-Barr virus (EBV) DNA. Among the EBV genes involved in latent infection, those encoding EBV latent membrane proteins are frequently expressed in PR. Several cytotoxic T-lymphocyte (CTL) defined epitopes have been mapped to latent membrane proteins restricted with HLA-A2, -A11 or -A24 antigens. Thus, the HLA-A allele may affect the development of PR. To examine this possibility, HLA-A alleles of 25 patients with EBV(+) PR were determined with low resolution polymerase chain reaction-based typing using HLA-A locus sequence specific primer combinations. The frequency of HLA-A alleles including HLA-A2 and -A24 antigens in PR patients was lower than that in the normal Japanese population, but the difference was not significant. Since HLA-A2-restricted CTL responses are well delineated at the A2-subtype level, the A2-subtype of PR cases with HLA-A2 antigen was further determined by high-resolution genetic typing. The frequency of HLA-A*0201 in PR was significantly lower than in the normal population (p=0.0314). The HLA-A*0201-restricted CTL responses may thus function in vivo to suppress the development of overt lymphoma. PMID- 10861474 TI - Methylation of the hMLH1 promoter but no hMLH1 mutations in sporadic gastric carcinomas with high-level microsatellite instability. AB - Microsatellite instability (MSI) in tumors from patients with hereditary non polyposis colorectal cancer (HNPCC) is caused by germline mutations in mismatch repair (MMR) genes, principally hMSH2 and hMLH1. In contrast, somatic mutations in MMR genes are relatively rare in sporadic MSI(+) colon cancers. Rather, the majority of mutation-negative, MSI(+) cases involve hypermethylation of the hMLH1 promoter and subsequent lack of expression of hMLH1. The details of the mechanisms of this epigenetic gene silencing remain to be elucidated. In some colon cancer cell lines, hMLH1 promoter methylation is accompanied by mutation of 1 of the 2 alleles, whereas in other cell lines and tumors, such combinations have not been reported. To contribute to the characterization of MSI in gastric cancer and to directly investigate whether hMLH1 promoter methylation is accompanied by gene mutation in these cancers, we have analyzed 42 gastric tumors and corresponding normal tissue for MSI, hypermethylation of the hMLH1 promoter, and mutations in hMLH1 as well as hMSH2. We found that 10 (23.8%) of 42 cases of sporadic gastric cancer were MSI(+) and that 8 had at least 2 of 12 altered microsatellite loci. All samples with at least 2 altered loci exhibited methylation of the hMLH1 promoter region, but none had detectable mutations in hMLH1 or hMSH2. Our results confirm the importance of methylation of the hMLH1 promoter region in MSI(+) gastric tumors and suggest that methylation takes place in the absence of hMLH1 mutations in these tumors. PMID- 10861475 TI - A tetranucleotide repeat polymorphism in CYP19 and breast cancer risk. AB - The CYP19 gene codes for aromatase, a key steroidogenic enzyme involved in the conversion of androgens to estrogens. A tetranucleotide (TTTA) repeat polymorphism is present in intron 4 of CYP19; 2 out of 4 breast cancer case control studies have reported a greater frequency of 2 specific alleles among affected women. We evaluated associations between CYP19 repeat alleles and breast cancer risk in a case-control study nested within the Nurses' Health Study cohort (incident cases: n=462; controls: n=618). We observed seven different CYP19 alleles (TTTA(7-13)). Compared to controls, cases had a statistically significant greater frequency of the 10 (TTTA)(10) repeat allele (10 allele: 2.3% vs. 0.7%, p = 0.005) and a nonsignificant increase in the frequency of the 12 (TTTA)(12) allele (12 allele: 3.1% vs. 2.1%, p = 0.11). A higher frequency of the 10 allele was observed in more advanced cancer cases defined as four or more involved nodes or distant metastasis [4+ nodes: 5/36 (13.9%) vs. 0-3 nodes: 13/330 (3.9%), p = 0.02]. Among controls, we found women with the 7 repeat allele to have decreased levels of estrone sulfate (-16.4%, p = 0.02), estrone (-6.1%, p = 0.22) and estradiol (-9.9%, p = 0.10), and a lower estrone/androstenedione ratio (E1/A) ( 10.5%, p = 0.08) compared to non-carriers. A higher E1/A ratio and elevated estrogen levels were observed among carriers of the 8 repeat allele; E1/A ratio (+21.0%, p = 0.003), estrone (+7.5%, p = 0.16) and estradiol (+10.8%, p = 0.08). However, we observed no evidence of an association between these alleles and breast cancer risk. We were unable to make inferences regarding the effect of the 10 allele on hormone levels due to the small number of allele carriers in the subgroup with hormone levels. As this repeat polymorphism is not close to the splice sites in intron 4, linkage disequilibrium with other functional polymorphisms in CYP19 may explain the findings of an increased association between breast cancer and the 10 allele variant of CYP19. We did not detect any sequence variants in the regulatory region or in the adipose-specific exon I.4. The lack of an established effect on CYP19 function associated with the 10 allele means that these findings should be interpreted with caution. PMID- 10861476 TI - Pre-malignant and malignant lymphoproliferations in an HCV-infected type II mixed cryoglobulinemic patient are sequential phases of an antigen-driven pathological process. AB - Type II mixed cryoglobulinemia (MC) is a systemic vasculitis characterized by the presence in the serum of a monoclonal cryoprecipitable IgM with rheumatoid factor (RF) activity. Hepatitis C virus (HCV) has been recognized as its major etiologic factor. Because MC frequently evolves into overt B-cell non-Hodgkin's lymphoma (NHL), chronic HCV infection is hypothesized to lead to both benign and malignant lymphoproliferative disease. In this study, we investigated mutations in the V(H) and V(K) genes of the B-cell clone originating the overt B-cell lymphoma in a subject with MC. Mutational patterns were analyzed longitudinally in two bone marrow biopsies obtained at the stage of MC, as well as in multiple involved tissues (bone marrow, liver, and peripheral blood cells) at the stage of overt NHL. Hybridization of variable-diversity-joining (VDJ) PCR products with a probe specific for the neoplastic clone indicated that the lymphoma originated from one of the clones over-stimulated during MC. This clone producing an IgM highly homologous to a protein with RF specificity may explain the MC syndrome in the patient. Moreover, the presence of an IgH ongoing mutation process and the expression of an Ig antigen receptor significantly homologous to an anti-HCV protein support the hypothesis that the MC syndrome and the subsequent evolution to NHL are antigen-driven lymphoproliferative processes possibly sustained by HCV. Furthermore, the marked reduction in intra-clonal diversity in the last bone marrow biopsy obtained at the stage of overt NHL points out a minor dependence of the cells on the antigen-driven mechanism, although an intrinsic propensity of the neoplastic cell to undergo replacement mutations cannot be excluded. PMID- 10861477 TI - Quantitative measurement of telomerase reverse transcriptase (hTERT) mRNA in urothelial cell carcinomas. AB - Telomerase reverse transcriptase (hTERT) messenger RNA has been detected in 95% of bladder tumors using RT-PCR. In this study, we quantified the expression of hTERT in 35 bladder urothelial cell carcinomas and in 6 normal bladder epithelia using a real-time quantitative PCR assay. hTERT expression was detected in all 35 urothelial cell carcinomas of varying grade and stage, but not in normal tissue samples. An increase in both pathological grade and clinical stage as prognostic parameters correlated with increased hTERT expression. Using different cutoff values for grades and stages, normalized hTERT expression values could discriminate among low, medium, and high grade tumors and between superficial and muscle-invasive tumors. We conclude that standardized real-time measurement of hTERT expression can be used for early tumor detection and may be used for determination of prognosis in urothelial cell carcinomas of the bladder. PMID- 10861478 TI - Distribution of 37 mucosotropic HPV types in women with cytologically normal cervical smears: the age-related patterns for high-risk and low-risk types. AB - Before guidelines can be set for the use of high-risk human papillomavirus (HR HPV) testing in cervical cancer screening and vaccine preparation, age-related prevalence of HR HPV types in cytologically normal smears has to be known. Therefore, in a cross-sectional study the prevalence of 37 different HPV genotypes and putatively unidentified HPV types was determined in 3,305 cytologically normal cervical smears from the general female population (15-69 years of age) using an HPV general primer GP5+/bioGP6+ mediated PCR assay. Subsequently, HPV-positive cervical smears were typed for 19 HR and 18 low-risk (LR) HPVs with an enzyme immunoassay using HPV type-specific oligoprobes in cocktails and individually, respectively. Overall, -HR and -LR HPV prevalences appeared to be of 4.6%, 3.3%, and 1.0%, respectively. Twenty-six different HPV types were detected in the 152 HPV-positive samples, the most prevalent types being HPV 16, 31, and 18. With regard to age, a peak prevalence of 19.6% for all HPVs was found in women 25-29 years of age, which declined to a mean of 4.3% in women over 30 years. With regard cytologically normal cervical smears (n = 3, 011) of women participating in the population-based screening program in the Netherlands (30 to 60 years), all HR HPVs showed decreased occurrence with increasing age, whereas the prevalence of LR HPV types remained constant. We suggest that screening for abnormal cytology implies screening for HR HPV infections and the subsequent treatment results in a decline of HR HPV prevalence in contrast to LR HPV prevalence during the years of screening. PMID- 10861479 TI - Second malignant tumors in patients with nasopharyngeal carcinoma and their association with Epstein-Barr virus. AB - Since previous published studies about second malignant tumors (SMTs) in nasopharyngeal carcinoma (NPC) patients usually included a limited sample size and did not attain consistent results, we conducted a large retrospective study in a cohort of 1,549 patients to assess the risk of SMT in NPC patients following radiotherapy (RT) in Taiwan. The follow-up period ranged from 2 to 16 years, with a median of 7 years. Thirty-nine patients developed SMTs during the 7,145 person year follow-up [standardized incidence ratio (SIR): 2. 8; 95% confidence interval (CI): 2.0 to 3.9]. Increased risks of developing SMTs were observed for head and neck (H/N) cancer (SIR: 16.5; 95% CI: 10.0 to 26.8), gastric cancer (SIR: 5.5; 95% CI: 2.2 to 11.4) and leukemia (SIR: 9; 95% CI: 1.9 to 26.3). Paraffin embedded specimens of secondary H/N cancer (11), secondary gastric cancer (6) and their corresponding NPC specimens were examined by EBER in situ hybridization to assess the association between Epstein-Barr virus (EBV) and these SMTs. Twenty six primary H/N and 5 gastric cancer specimens were chosen as the control groups. In H/N cancer, EBV was detected in 3.8% of the primary cancers and 9.1% of the secondary cancers. All the positive specimens resulted from hypopharyngeal cancer. Of the secondary gastric cancers, only 1 case (16.6%) was associated with EBV. None of the primary gastric cancers was associated with EBV. Our results indicate an increased risk of developing SMTs, with a preference for head and neck cancer, gastric cancer and leukemia, in NPC patients after RT in Taiwan. Only a small proportion of the secondary H/N and gastric cancers was associated with EBV. PMID- 10861480 TI - Seropositivity to human herpesvirus 8 in relation to sexual history and risk of sexually transmitted infections among women. AB - The mode of transmission of human herpesvirus 8 (HHV8) was investigated in two seroepidemiological studies of Swedish women who completed a questionnaire about sexual behavior. Seropositivity for HHV8 antibodies, measured using an indirect immunofluorescence assay, was linked to a high number (>10) of sexual partners (P < 0.004). It also correlated strongly with a history of other sexually transmitted diseases (STD; P < 0.0001), in particular with a history of Chlamydia trachomatis infection and condyloma acuminata. There was appreciable HHV8 seropositivity already among virginal or monogamous women (9%). In summary, HHV8 transmission to women in Sweden may occur nonsexually. When sexual transmission occurs, it appears to be associated with high risk-taking sexual behavior. PMID- 10861481 TI - Methylation of specific CpG sites in the promoter region could significantly down regulate p16(INK4a) expression in gastric adenocarcinoma. AB - Silencing of p16(INK4a) by methylation of the CpG islands in the promoter region has been found to be an alternative mechanism of inactivation in several tumors. However, in gastric carcinoma, the relationship between methylation status and the transcriptional silencing of the p16 gene remains to be clarified. In this study, we investigated whether methylation of a few specific CpG sites in the promoter region could significantly down-regulate p16 activity in the tumorigenesis of gastric carcinoma. By Southern analysis and bisulfite-modified genomic sequencing of 9 gastric-carcinoma cell lines, we found that the 5 cell lines (55.5%) not expressing p16 mRNA had methylated CpG sites at the promoter region of p16. In addition, we analyzed the p16-protein expression of 28 primary gastric carcinomas and their normal counterparts by immunohistochemical staining (IHC) on paraffin sections. Loss of p16 expression was detected in 6 cases (22%). In 5 out of these 6 (83%), the actual p16 gene was inactivated by de novo methylation of the promoter sites. Taken together, these results suggest a strong correlation between de novo methylation of a few specific CpG sites and transcriptional silencing of the p16 gene in gastric carcinoma. PMID- 10861482 TI - Naturally processed and concealed HLA-A2.1-restricted epitopes from tumor associated antigen tyrosinase-related protein-2. AB - In this study, a computer-assisted reverse immunology approach was utilized in order to identify potentially antigenic peptides derived from the differentiation antigen TRP-2, a melanosomal protein frequently expressed in melanoma. Among the seven peptides complying with HLA-A2.1-binding motifs, two induced specific CD8(+) cytotoxic T lymphocytes. HLA-A2.1(+) melanoma cells expressing TRP-2 were lysed by clones specific for TRP-2(360-368) (TLDSQVMSL) peptide, thus identifying it as a naturally processed epitope. Other T-cell clones directed against TRP 2(476-484) (VMGTLVALV) were unable to lyse HLA-matched TRP-2(+) cell lines. The role of intracellular proteolytic processing in the generation of this epitope was investigated by transfecting mini-genes encoding the TRP-2(476-484) peptide alone or carrying N- or C-terminal extensions. Specific T-cell clones recognized target cells expressing the cytotoxic T-lymphocyte (CTL)-defined epitope or its C terminally extended precursor, but failed to recognize cells expressing the N terminally extended TRP-2(476-484) peptide, suggesting the presence of a negative processing signal (NPS). Regarding C-terminus-flanking regions, mutational analysis indicates that the GLY485 residue plays a key role in the processing of the TRP-2(476-484) epitope. Interestingly, proteasome inhibitors preventing the generation of the MART-1/Melan-A(27-35) immunodominant melanoma tumor-associated antigen (TAA) promoted detectable presentation of TRP-2(476-484) epitope in HLA A2.1(+) and TRP-2(+) tumor lines, as witnessed by cytokine release by specific T cell clones. PMID- 10861483 TI - A novel approach for inducing enhanced and selective transgene expression in hepatocellular-carcinoma cells. AB - Utility of the alpha-fetoprotein (afp) promoter for gene therapy against hepatocellular carcinoma (HCC) is limited because of the weak promoter activity. To circumvent this, the 5.1-kb 5;-flanking sequence of the human afp gene including the entire enhancer and silencer regions as well as the promoter region was employed for achieving strong, HCC-selective transgene expression. To thoroughly inhibit the promoter activity of the 5;-flanking sequence of the human afp gene, the afp 5;-flanking region was inserted downstream of the human interleukin-2 (il-2) gene controlled by the simian-virus-40 (SV40) early promoter. il-2-production ability of HCC cells transduced with the construct was significantly enhanced compared with that transduced with the same construct lacking the afp 5;-flanking region. Importantly, il-2 production of non-HCC cells was substantially inhibited by the addition of the afp 5;-flanking region to the construct. When the afp 5;-flanking region was inserted downstream of the human tumor-necrosis-factor-alpha (tnf-alpha) gene controlled by the retrovirus long terminal-repeat (LTR) enhancer/promoter, tnf-alpha production ability of HCC cells was significantly enhanced and that of non-HCC cells was significantly suppressed compared with that transduced with the same construct lacking the afp 5;-flanking region. Our results indicated that the afp 5;-flanking region gave the enhanced HCC-selective activity to the non-tissue specific SV40 early promoter and LTR enhancer/promoter. It is essential for successful gene therapy to induce strong, target-cell-selective transgene expression. This novel strategy, therefore, may contribute to the establishment of HCC-selective cancer gene therapy. PMID- 10861484 TI - Cyclophosphamide enhances anti-tumor effect of wild-type p53-specific CTL. AB - The tumor suppressor protein p53 is overexpressed in up to 50% of all human malignancies, both in solid tumors as well as hematological malignancies, and is therefore an attractive target for immunotherapy. We have recently shown that cytotoxic T lymphocytes (CTL), raised in p53 gene deficient (p53 -/-) mice and recognizing a murine wild-type (wt) p53 peptide, were able to eradicate a mutant p53-induced and overexpressing tumor in p53 +/+ nude mice. These CTL also prevented the outgrowth of a more aggressive p53-overexpressing tumor in immunocompetent C57BL/6 mice. Importantly, this occurred in the absence of demonstrable damage to normal tissue. Possibly due to the aggressive nature of the latter tumor, adoptive transfer of wtp53-specific CTL did not result in the eradication of established tumors, either in nude or immunocompetent mice. Therefore, we explored whether the cytotoxic drug cyclophosphamide (CY) could potentiate the therapeutic activity of wtp53-specific CTL. We show here that CY acts synergistically with adoptively transferred wtp53-specific CTL in controlling the growth of an aggressive mutant p53-induced and overexpressing tumor. Previously described mechanisms underlying the synergism between CY and immune T cells were evaluated, but were not found to be operational in this model. PMID- 10861485 TI - Targeting angiogenesis inhibits tumor infiltration and expression of the pro invasive protein SPARC. AB - The solid growth of high-grade glioma appears to be critically dependent on tumor angiogenesis. It remains unknown, however, whether the diffuse infiltration of glioma cells into healthy adjacent tissue is also dependent on the formation of new tumor vessels. Here, we analyze the relationship between tumor angiogenesis and tumor cell infiltration in an experimental glioma model. C6 cells were implanted into the dorsal skinfold chamber of nude mice, and tumor angiogenesis was monitored by intravital fluorescence videomicroscopy. Glioma infiltration was assessed by the extent of tumor cell invasion into the adjacent chamber tissue and by expression of SPARC, a cellular marker of glioma invasiveness. To test the hypothesis that glioma angiogenesis and glioma infiltration are codependent, we assessed tumor infiltration in both the presence and the absence of the angiogenesis inhibitor SU5416. SU5416 is a selective inhibitor of the VEGF/Flk-1 signal-transduction pathway, a critical pathway implicated in angiogenesis. Control tumors demonstrated both high angiogenic activity and tumor cell invasion accompanied by strong expression of SPARC in invading tumor cells at the tumor host tissue border. SU5416-treated tumors demonstrated reduced vascular density and vascular surface in the tumor periphery accompanied by marked inhibition of glioma invasion and decreased SPARC expression. A direct effect of SU5416 on glioma cell motility and invasiveness was excluded by in vitro migration and invasion assays. These results suggest a crucial role for glioma-induced angiogenesis as a prerequisite for diffuse tumor invasion and a possible therapeutic role for anti-angiogenic compounds as inhibitors of both solid and diffuse infiltrative tumor growth. PMID- 10861486 TI - Anti-angiogenic activity of torilin, a sesquiterpene compound isolated from Torilis japonica. AB - Torilin is a sesquiterpene compound purified from fruits of Torilis japonica (Umbelliferae). In this study, we demonstrated the anti-angiogenic activity of torilin using in vivo and in vitro assay systems. Torilin decreased both neovascularization of chick embryos in the chorioallantoic membrane assay and basic fibroblast growth factor-induced vessel formation in the mouse Matrigel plug assay. Torilin also reduced the proliferation and tube formation of human umbilical vein endothelial cells. In addition, the concentrated conditioned media obtained from torilin-treated HepG2 human hepatoblastoma cells blocked the angiogenic activation of torilin-untreated concentrated conditioned media, indicating that torilin may have an inhibitory effect on tumor-induced angiogenesis. To determine what molecules were involved in the anti-angiogenic activity, we examined the expression of hypoxia-inducible angiogenic factors in torilin-treated HepG2 cells. Torilin significantly down-regulated the expression of hypoxia-inducible vascular endothelial growth factor and insulin-like growth factor-II. Taken together, our data suggest that torilin may be a strong angiogenic inhibitor with the ability to decrease tube formation of vascular endothelial cells and to reduce expression of angiogenic factors of tumor cells. PMID- 10861487 TI - Synthetic inhibitor of matrix metalloproteases decreases tumor growth and metastases in a syngeneic model of rat prostate cancer in vivo. AB - Members of the matrix metalloprotease (MMP) family are implicated in the progression of several malignancies including prostate cancer due to their ability to break down extracellular matrix (ECM) components. In this study, we have evaluated the ability of a synthetic MMP inhibitor (A-177430) to block tumor growth and metastases in a syngeneic model of rat prostate cancer. In an in vitro substrate assay, A-177430 exhibited nanomolar potency (IC(50) 2-6 nM) against the enzymatic activity of several MMPs. For in vivo studies, male Copenhagen rats were injected s.c. with Mat Ly Lu rat prostate cancer cells (1 x 10(6) cells ) into the right flank and animals were administered i.p.with different doses (10 100 mg/kg per day) of A-177430 for 16 days. Administration of A-177430 resulted in a dose-dependent decrease in tumor volume as compared to a control group of animals receiving vehicle alone. The maximum dose (100 mg/kg per day) of A-177430 exhibited complete arrest in tumor growth and prevented the development of macroscopic tumor metastases to lungs without exhibiting any noticeable side effects. Histologic examination of primary tumors from experimental animals showed extensive tumor necrosis and decreased tumor angiogenesis as determined by factor VIII staining of primary tumors following A-177430 treatment. These primary tumors from experimental animals also exhibited a significant increase in tumor cell DNA fragmentation as determined by TUNEL assay. Collectively, these results demonstrate the ability of MMP inhibitors to block tumor growth and metastases by blocking ECM degradation and by inhibiting tumor angiogenesis and promotion of prostate cancer cell apoptosis in vivo. PMID- 10861488 TI - Subsequent primary cancers after basal-cell carcinoma: A nationwide study in Finland from 1953 to 1995. AB - The aim of this study was to investigate whether patients with basal-cell carcinoma (BCC) of the skin have an increased risk of developing other cancers. A total of 71,924 patients diagnosed with BCC between 1953 and 1995 were identified from the Finnish Cancer Registry. They were followed up for subsequent primary cancers from the date of the first BCC diagnosis to the end of 1995. Standardized incidence ratios (SIR) with 95% confidence intervals (CI) were calculated based on national rates. Altogether 11,042 subsequent primary cancers occurred among the study cohort during 625,000 person-years of follow-up. Risk increases were observed for non-melanoma skin cancer (SIR 3.79, 95% CI 3.59-4.00) and skin melanoma (SIR 2.34, 95% CI 2.08-2.61). The five other primary sites presenting the highest SIRs were salivary glands (SIR 3.30), lip (2. 19), small intestine (1.85), nose (1.73) and pharynx (1.71). Patients who were less than 40 years of age at the time of BCC diagnosis had a significantly higher relative risk for a subsequent new cancer than the older patients (ratio of the SIRs 1.29, 95% CI 1. 10-1.51). Time since BCC diagnosis did not materially influence the overall relative risk of subsequent cancers. Part of the increase in the risk of skin cancer is likely to be due to enhanced diagnostic activity after an initial diagnosis of BCC. However, the increases in the risk of several non-cutaneous cancers suggest a generalized carcinogenic role of some factors in the BCC pathogenic pathways. PMID- 10861489 TI - Food groups and risk of squamous cell esophageal cancer in northern Italy. AB - To better understand the nutritional etiology of squamous cell esophageal cancer, we conducted a case-control study in 3 areas of northern Italy. A total of 304 incident, histologically confirmed cases of squamous cell carcinoma of the esophagus (275 men, 29 women) and 743 hospital controls (593 men, 150 women) with acute, non-neoplastic conditions, not related to smoking, alcohol consumption or long-term diet modification, were interviewed during 1992 to 1997. The validated food-frequency questionnaire included 78 questions on food items or recipes, which were then categorized into 19 main food groups, and 10 questions on fat intake pattern. After allowance for age, sex, education, area of residence, tobacco smoking, alcohol drinking and non-alcohol energy, a significant increased risk emerged for high consumption of soups (OR=2.1 for the highest vs. lowest quintile), whereas inverse associations with esophageal cancer risk were observed for pasta and rice (OR=0.7), poultry (OR=0.4), raw vegetables (OR=0.3), citrus fruit (OR=0.4) and other fruit (OR=0.5). The associations with dietary habits were consistent in different strata of tobacco smoking and alcohol drinking. Among added lipids, olive oil intake showed a significant reduction of esophageal cancer risk, even after allowance for total vegetable consumption (OR=0.4), while butter consumption was directly associated with this risk (OR=2.2). Our results thus provide further support to the evidence that raw vegetables and citrus fruit are inversely related to the risk of squamous cell esophageal cancer and suggest that olive oil may also reduce this risk. PMID- 10861490 TI - Association of menstrual and reproductive factors with breast cancer risk: results from the Shanghai Breast Cancer Study. AB - The incidence of breast cancer among women in Shanghai, a traditionally low-risk population, has increased substantially over the past 20 years. To evaluate the association of menstrual and reproductive factors with breast cancer risk and the influence of these factors on the temporal trend of breast cancer incidence, we analyzed data from the Shanghai Breast Cancer Study, a population-based case control study of breast cancer recently completed among Chinese women in urban Shanghai. In-person interviews were completed for 1,459 women newly diagnosed with breast cancer between ages 25 and 64 and for 1,556 controls frequency matched to cases by age. Unconditional logistic regression was employed to estimate adjusted odds ratios (ORs) and 95% confidence intervals (CIs) related to menstrual and reproductive factors. Earlier menarcheal age, nulliparity, and later age at first live birth were associated with increased risk of breast cancer among both pre- and post-menopausal women, while never having breast-fed and later age at menopause were associated with elevated risk only among post menopausal women. Among controls, 32% of younger women (40 years) reported starting menarche at age of 13 or younger, and this factor contributed to 44% of cases diagnosed among younger women and 26% to 28% of cases in older women. Older age at first live birth or at menopause explained a considerable portion of cases diagnosed in older, but not younger, women. Our study suggests that the changes in menstrual and reproductive patterns among women in Shanghai have contributed to the recent increase in breast cancer incidence, particularly among younger women. PMID- 10861491 TI - National cancer prevalence estimation in France. AB - In France, as in several other European countries, prevalence has to be estimated from the modelling of 2 of the 3 basic epidemiological measures of incidence, mortality, and survival. Since, in these countries, follow-up of cancer patients is only made in a few registries, we explored the feasibility of estimating prevalence in the absence of follow-up data. The method, which used only incidence and mortality, was validated on Danish data and applied to France. For this latter country, the estimation procedure is based on the recorded mortality data and an estimate of incidence for the entire country. It is applied to selected sites of cancer, which account for 80% of the estimated incidence. In 1992, the prevalence of patients who had such a diagnosis amounts to 538,000 women and 424, 000 men. The most frequent cancer sites are head and neck, breast, and large bowel. Most of the cancer sites present an increase in prevalence proportion between 1987 and 1992. The larger increases concern breast and prostate cancer. PMID- 10861492 TI - An intronic splicing mutation of the MEN1 gene. PMID- 10861493 TI - A 5178-9g--> A splice donor site mutation in intron 4 of the MEN1 gene causing multiple endocrine neoplasia type 1 PMID- 10861494 TI - Lifestyle and cancer: protection from a cancer-free spouse. PMID- 10861495 TI - Down-regulation of p27 is a significant predictor of poor overall survival and may facilitate metastasis in colorectal carcinomas. AB - The p27 gene product has been shown to have prognostic significance in a range of tumors. Down-regulation of p27 has also been implicated in loss of cell adhesion in tumor cells. Our study aimed to investigate whether p27 expression was significantly correlated with overall survival of colorectal carcinoma patients in the Singapore population, which is predominantly Chinese. Staining was performed on 136 paraffin-embedded specimens collected between 1991 and 1992 using an anti-p27 monoclonal antibody. Follow-up of patients was until time of death or for 5 years. There was a significant association between overall survival and p27 expression for all specimens. However, there was no significant correlation between p27 expression and other clinical features such as gender, age, tumor stage, differentiation, and site. When stratified by tumor stage, patients whose tumors exhibited higher metastatic potential (stage III/IV) but had strong p27 expression had a median survival that was 23 months longer than stage III/IV patients whose tumors had no or weak p27 expression. Our results thus suggest that one potential mechanism of action of p27 is to suppress metastasis possibly through its involvement in cell adhesion. PMID- 10861496 TI - Amplification and over-expression of the AIB1 nuclear receptor co-activator gene in primary gastric cancers. AB - Our analysis of chromosomal aberrations in primary gastric cancers using comparative genomic hybridization has revealed novel, high and frequent copy number increases in the long arm of chromosome 20, indicating that this region contains novel amplified genes involved in gastric cancer progression. AIB1, a member of the steroid receptor co-activator-1 family, has been cloned on 20q12 as a candidate target gene for this amplification in human breast cancers. In this study, we examined the numbers of AIB1 copies as well as their expression and relation to clinico-pathological features in 72 primary gastric cancers. AIB1 amplification was observed in 7% and over-expression in 40% of the specimens. AIB1 amplification always coincided with its over-expression, but several cases showed AIB1 over-expression without amplification, suggesting that expression of AIB1 is regulated not only by gene amplification but also by other mechanisms, such as transcriptional activation, in human gastric cancer. Gastric cancers with AIB1 amplification showed extensive lymph node metastases, liver metastases and poorer prognosis compared to those without amplification. Our results suggest that amplification and over-expression of AIB1 are likely to increase the number of malignant phenotypes of gastric cancers and that it can be expected to be useful as a marker of poor prognosis. PMID- 10861497 TI - LEA.135 expression: an independent and favorable prognostic biomarker for patients with primary invasive breast cancer. AB - The prognostic significance of LEA.135 expression, detected by immunohistochemistry in formalin-fixed and paraffin-embedded tissue sections, was evaluated and compared with the widely utilized clinicopathological parameters for patients with primary invasive breast carcinomas. Pathological parameters such as tumor size, histological tumor type, histological grade, nuclear grade, lymph node (LN) status, bone marrow (BM) status, as well as age of patient at initial diagnosis together with follow-up in years were available for this group of patients (n = 178). Among these parameters, tumor size, histological tumor type, histological grade, LN status, and BM status were individually and significantly associated with increased probability of recurrence by univariate analysis. By multivariate analysis, however, only tumor size, LN status, and BM status remained statistically significant. LEA.135-positive patients showed a statistically significant probability of not recurring (77 +/- 5% at 5 years after surgery) compared with patients who were LEA. 135-negative (49 +/- 6% at 5 years after surgery) (log-rank p < 0. 001). Furthermore, the association remained statistically significant by multivariate analysis (log-rank p = 0.019), demonstrating that LEA.135 expression independently and significantly identified breast cancer patients with favorable clinical outcome. In addition, there was a statistically significant association between loss of LEA.135 expression and poor prognosis when patients were stratified by pathological parameters. Furthermore, a subgroup of patients who were LEA. 135-positive/LN-negative experienced a decreased rate of recurrence compared with those who were LEA.135-negative/LN negative (16% vs. 27%, respectively). A similar result was also obtained when BM negative patients were stratified on the basis of LEA. 135-positive or LEA.135 negative subgroups for recurrence (18% vs. 43%, respectively). Most interestingly, the patients whose cancer cells were LEA.135-positive/LN-positive experienced a much lower rate of recurrence than those whose cells were LEA. 135 negative/LN-positive (29% vs. 57%, respectively). The results clearly demonstrate that LEA.135 expression was a significantly independent and favorable prognostic marker for patients with primary invasive breast carcinoma by both univariate and multivariate analyses. PMID- 10861498 TI - Genetic progression in microsatellite instability high (MSI-H) colon cancers correlates with clinico-pathological parameters: A study of the TGRbetaRII, BAX, hMSH3, hMSH6, IGFIIR and BLM genes. AB - Colon carcinomas with microsatellite mutator phenotype exhibit specific genetic and clinico-pathological features. This report describes the analysis of 63 "microsatellite instability-high" (MSI-H) tumors for the presence of mutations in microsatellites located in the coding regions (CDRs) of 6 genes: TGFbetaRII, BAX, hMSH3, hMSH6, IGFIIR, and BLM. The following frequencies of mutations were detected: TGFbetaRII (70%), BAX (54%), hMSH3 (36.5%), IGFIIR (22%), hMSH6 (17.5%), and BLM (16%). The overall picture revealed combinations of mutations suggestive of a progressive order of accumulation, with mutations of TGFbetaRII and BAX first, followed by frameshifts in hMSH3, hMSH6, IGFIIR, and BLM. Correlations with 12 clinico-pathological parameters revealed that tumors with frameshifts in 1 or 2 CDRs were significantly better differentiated than tumors with frameshifts in more than 2 CDRs. We also found that mutations in the hMSH3 gene were significantly associated with decreased wall invasiveness and aneuploidy, and frameshifts in the BLM gene were significantly associated with the mucinous histotype. A trend toward an association between hMSH3 and IGFIIR with the medullary and conventional adenocarcinoma histotypes, respectively, was seen. Our results strengthen the concept that mutations in target genes have a role in the tumorigenic process of MSI-H tumors, and indicate that frameshifts in microsatellites located in CDRs occur in a limited number of combinations that could determine distinct clinico-pathological traits. PMID- 10861499 TI - p27(kip1) expression in breast carcinomas: an immunohistochemical study on 512 patients with long-term follow-up. AB - p27(Kip1) (p27), a cyclin-dependent kinase inhibitor, has an important role in the progression of cells from G(1) into S phase of the cell cycle. p27 may act as a tumor suppressor, and several reports suggest that loss of its expression in breast carcinoma is related to tumor progression and poor prognosis. We evaluated p27 immunohistochemical expression in 512 consecutive cases of breast carcinoma with 9 years of median-term follow-up. p27 expression was heterogeneous and frequently less intense than in normal cells. Low p27 expression (<50% of reacting cells) was associated with grade III tumors, N0 status, estrogen receptor-negative status, and low cyclin D1 expression. In the whole series of cases, p27 expression did not predict outcome. In node-negative cases (249 patients), high p27 expression indicated poor prognosis. p27 was not prognostically relevant in the group of 223 patients with pT1 disease or in the group of 154 patients <50 years of age. We also investigated the prognostic value of the combined expression of p27 and cyclin D1, but no differences in survival were seen in this bivariate analysis. PMID- 10861500 TI - Lack of expression of c-KIT in ovarian cancers is associated with poor prognosis. AB - The c-KIT protooncogene encodes a tyrosine kinase receptor, KIT, that is expressed in many normal and cancerous tissues. In this study, we have examined the expression of c-KIT and its ligand, stem cell factor (SCF), in human epithelial ovarian tumors, in normal ovaries and in cultured ovarian surface epithelium (OSE). Cultured cells, normal tissues and tumors were analyzed by Northern and Western blot analyses, reverse transcription-polymerase chain reaction and immunohistochemistry. Normal OSE expressed SCF, but not c-KIT; however, epithelial invaginations and inclusion cysts often expressed KIT protein. Of 15 benign ovarian tumors and tumors of low malignant potential, 87% expressed c-KIT, and 92% of these co-expressed SCF, suggesting the possibility of autocrine growth regulation. Of 35 malignant ovarian cancers, 71% expressed c-KIT (92% co-expressed SCF), with a trend for decreased c-KIT expression in advanced stage disease. Of 34 patients with malignant tumors for whom follow-up information was available (median follow-up time of 24 months), 9 had tumors that did not express c-KIT, 8 (89%) of whom have died and the remaining 1 has recurrent disease. Of the 25 patients with tumors expressing c-KIT, 56% are still alive. Eight of the patients have no evidence of disease and all had KIT expressing tumors. Statistical analysis indicated that patients whose tumors did not express c-KIT had a significantly shorter (p < 0.05) disease-free survival time than patients who had KIT-expressing tumors. Our results suggest that c-KIT may play a role in early ovarian tumorigenesis, and that loss of c-KIT expression is associated with poor prognosis. PMID- 10861501 TI - Expression of TGF-beta isoforms, TGF-beta receptors, and SMAD molecules at different stages of human glioma. AB - Human gliomas express TGF-beta but, so far the expression of downstream mediators has been investigated in only a few cell lines. We have examined tissue specimens of 23 gliomas: 3 astrocytomas grade II (AST), 8 anaplastic astrocytomas grade III (AAST), and 12 glioblastoma multiforme grade IV (GBM). We analyzed the mRNA expression of TGF-beta1, TGF-beta2, TGF-beta3, the TGF-beta receptors type I (TbetaR-I) and type II (TbetaR-II), Smad2, Smad3, and Smad4. mRNA expression of IL-10 and CD95 (FAS/APO-1) were also studied. We detected increased mRNA levels of the 3 TGF-beta isoforms, correlating with the degree of malignancy. TGF-beta3 mRNA was increased, particularly in AST and AAST, while TGF-beta1 and TGF-beta2 mRNAs were strongly expressed in GBM. TGF-beta normally up-regulates the TGF-beta receptors, and TbetaR-I and TbetaR-II showed stronger expression in all gliomas when compared to normal tissues. However, the mRNA expression of Smad2, Smad3, and Smad4 was decreased in GBM. IL-10 mRNA expression was detected in glioma tissues but not in glioma cell lines. No marked increase in the expression of soluble CD95 splicing variants was found in the gliomas compared with normal tissue. However, total CD95 mRNA was elevated among GBM tissues. PMID- 10861502 TI - Epidemiologic determinants of clinically relevant prostate cancer. AB - While tumor volume and Gleason scores are the best available prognostic indicators for prostate cancer, contemporary predictive methods are unable to identify which men with Gleason scores of 7 have clinically insignificant tumors that will not progress and which men will develop highly aggressive prostate cancer. Our objective was to evaluate potential environmental determinants of significant prostate cancer. Subjects were patients identified from a university based hospital and tertiary cancer center who had undergone radical prostatectomy for prostate cancer. Cases were 103 patients whose tumor volumes were 80%). Treatments of 1 week duration revealed that the most sensitive time to AI lies in the first week (between 7 and 14 dph). Progeny testing of males from AI-treated groups gave results indicating that these were XX males, as expected. These experiments strongly implicate aromatase activity as a key factor in sexual differentiation in the Nile tilapia. PMID- 10861550 TI - Seasonal changes in circulating testosterone and androstenedione concentration and their correlation with the anomalous reproductive pattern in the male indian sheath-tailed bat, Taphozous longimanus. AB - The relationship between reproductive organs, circulating testosterone, and androstenedione concentration in the male sheath-tailed bat, Taphozous longimanus, was studied. The masses of testis, accessory sex gland (prostate, ampullary), and epididymides showed three peaks, one each in October, January, and April. Monthly changes in testosterone also peaked during October, January, and April and closely coincided with the peak spermatogenesis. Serum androstenedione concentration peaked during November and January. Testosterone showed a strong correlation with masses of testis and accessory sex glands, while androstenedione showed strong correlation with the body mass. Different threshold levels of testosterone may be required to trigger spermatogenesis, secretory activity of accessory sex glands and mating in Taphozous longimanus and may be responsible for reproductive asynchrony in this species. Higher circulating concentrations of testosterone and androstenedione throughout the year in this species, as compared with other mammalian species, may be responsible for prolonged retention of sperm in the epididymides. PMID- 10861551 TI - Implication of somatolactin in the regulation of sexual maturation and spawning of Mugil cephalus. AB - Specific antibody for chum salmon somatolactin (SL) was used for immunocytochemical investigation of SL cell activity of Mugil cephalus during the gonadal cycle in both natural habitat and captivity. The SL-immunoreactive cells showed strong and specific immunoreactivity to antichum salmon SL. The number of SL-immunoreactive cells increased, as did the secretory and synthetic activity during sexual maturation and spawning in the natural habitat. The SL cells were rather small and moderately immunoreactive in immature fish; they were enlarged, their numbers increased, and they frequently showed more SL immunoreactivity during gonadal development. In addition, during late stages of maturation, small cell size with more or less SL immunoreactive cells were noted, indicating an active release of SL granules. Prespawning females tended to have more enlarged SL cells with stronger immunoreactivity than equivalent males. The SL cells showed an increase in the secretory activity during spawning as indicated by small size and weak immunoreactivity. The SL cells of M. cephalus reared in captivity showed high activity. This may be due to the low concentration of calcium in fresh water. The gradual stimulation of SL synthesis and release during sexual maturation and spawning of M. cephalus suggest that SL may be involved in the control of some steps of reproductive processes, such as steroidogenesis, calcium metabolism, and energy mobilization. PMID- 10861552 TI - Lack of gonadotrophin-releasing hormone (GnRH) neuron response to decreasing photoperiod in thyroidectomized male starlings (Sturnus vulgaris). AB - Exposure of starlings to long days initially causes reproductive maturation, but eventually leads to photorefractoriness. During photorefractoriness, gonadotrophin-releasing hormone (GnRH) decreases in the GnRH cell bodies and fibers emanating from these to the median eminence, circulating gonadotrophin concentrations decrease to a minimum, and the gonads regress. Thyroidectomy profoundly affects these photoperiodic responses. In chronically thyroidectomized starlings, gonadal responses to changes in day length are attenuated. This investigation was conducted to determine whether, in the absence of gonadal responsiveness, the GnRH system of chronically thyroidectomized starlings responds to changes in day length. Two groups of thyroidectomized male starlings were transferred from short days (8L:16D) to long days (18L:6D) for four weeks, and testicular volume increased. One group was kept on long days (TxLD) and the other was returned to short days (TxSD). Testicular volume did not decrease in the TxSD group. The GnRH neurons of the two thyroidectomized groups were compared to those of two groups of intact starlings, one of them on long days and photorefractory (ILD), the other on short days and photosensitive (ISD). Group ILD had lower numbers of GnRH-stained cells than groups TxLD, TxSD and ISD, which did not differ in this respect. Similar differences were observed for GnRH cell size in the pre-optic area (POA) and for density of staining of GnRH fibers in the median eminence. The results confirm that thyroidectomy attenuates gonadal responses to change in day length and suggest that this results from an effect upon the central nervous system rather than a peripheral effect. PMID- 10861553 TI - Direct and correlated responses to individual selection for large adult weight in the edible snail Helix aspersa Muller. AB - A selection experiment for large adult weight based on individual performance was conducted for three generations in Helix aspersa aspersa. A second line was kept as an unselected control line. Direct response measured as deviation from the control line was 3.55 g after three generations of artificial selection, which averaged 13%. Realized heritability was 0.38 +/- 0.04. Correlated responses to selection showed a significant increase in weight after hibernation, mean egg weight and mean weight of newly-hatched snails with selection. For adult age, egg number, and hatching rate, no significant change correlated to selection was found, but this is to be confirmed. PMID- 10861554 TI - Prostaglandins and reproduction of the scallop argopecten purpuratus: II. relationship with gamete release. AB - Levels of prostaglandins E(2) and F(2alpha) were measured in male and female gonadal portions of the functional hermaphroditic scallop Argopecten purpuratus, at the time of spawning. For these measurements, mature scallops were stimulated to initiate their chain of spawning events, and prostaglandins (PGs) were analyzed in samples before stimulation, when animals appeared stimulated to release sperm, during the release of sperm, when animals appeared stimulated to release oocytes, when most of the oocytes had been released, and 24 hr after the end of spawning. The experiment was run twice, once in winter and once in spring. An additional in vitro experiment was carried out to analyze activity of the monoamines dopamine and serotonin on the levels of these prostaglandins. For this, minced tissue of the female portion of the gonad was incubated for different time periods with each of the amines at 10(-5) M. PGs were determined in the tissue samples by radioimmunoanalysis. The results showed that the amounts of the PGs increased significantly in both parts of the gonad during the global process of spawning and decreased when it had been completed. Incubation of minced gonad with the amines affected the time course of PGs variations, i.e., the effect of these compounds depended on the incubation time used. The results support a model for the regulation of spawning which assumes the occurrence of a stimulus which is detected by receptors, processed by nerve cells, and sent to the gonad where intermediation by amines (changing the time course of the process) induces liberation of gametes, in some way modulated by prostaglandins. PMID- 10861555 TI - Induction of ovulation by clomiphene citrate in the Indian vespertilionid bat, Scotophilus heathi. AB - The ovulation induction property of clomiphene citrate (CC) and human chorionic gonadotropin (hCG) was studied in Scotophilius heathi, an Indian tropical vespertilionid bat, during the period of delayed ovulation between December to early January. The results of the study showed that 10 microg of CC alone was ineffective to induce ovulation, whereas 100 microg CC and 10 IU hCG alone induced ovulation. A significant (P < 0.01) increase in the ovulation rate was observed when 10 microg CC followed by 10 IU hCG, compared to 10 IU hCG and 100 microg CC alone groups. Finally, CC at a 100 microg dose, followed by 10 IU hCG, produced superovulation (14.00 +/- 0. 70), which is significantly different in comparison to all other groups. This is the first report of ovulation induced by CC in the Indian tropical bat as well as in any animal model that exhibits temporary anovulation similar to polycystic ovary syndrome (PCOD) during the normal physiology of reproduction. PMID- 10861556 TI - Induction of gynogenesis in muskellunge with irradiated sperm of yellow perch proves diploid muskellunge male homogamety. AB - Diploid gynogenesis was induced in muskellunge Esox masquinongy using UV irradiated muskellunge sperm as the first step in producing monosex females. In this approach, we have to rely on negative controls as an indirect reference for sperm genetic material destruction. In the first experiment, equal proportions of gynogenetic females and males were produced. Negative controls, UV-irradiated sperm without heat shock, yielded some normal hatching larvae, described as spontaneous diploids. In the second experiment, muskellunge eggs were activated using sperm from yellow perch. Because hybrids between these species are not viable, we produced unambiguous gynogens. When UV-irradiated yellow perch sperm was used to inseminate muskellunge eggs, haploids resulted (22.5% +/- 2.8% survival to the eyed stage). To produce diploid gynogens, a heat shock of 31 degrees C was applied to inseminated eggs 20 min after activation for a duration of 6 min. This process yielded several hundreds of gynogens for rearing. Several treatments of masculinizing hormone, 17alpha-methyltestosterone (MT), were carried out. Fish were dissected and gonads examined histologically for sex determination. Gynogens produced using yellow-perch sperm confirmed the presence of males in the control group, whereas the MT bath treatment (400 microg/liter) resulted in the production of fish with ovotestis. These results provide evidence for male homogamety in muskellunge and imply that a change of strategy is needed to produce monosex populations. PMID- 10861557 TI - Developmental characterization and chromosomal mapping of a LacZ transgene expressed in the mouse apical ectodermal ridge. AB - The apical ectodermal ridge (AER) has an essential role in limb morphogenesis involving the specification of the proximal-distal axis of the limb. During the analysis of transgenic mice that harbor a LacZ transgene, we detected strong expression of beta-galactosidase within the AER of developing embryos. In this mouse line, called Z16, the bacterial LacZ gene is linked to a Herpes simplex virus immediate early promoter that is normally silent in mice. Embryos from other independent mouse lines harboring the same DNA construct exhibited no AER specific staining. Thus, it appears that the LacZ transgene in the Z16 line is expressed in the AER in response to regulatory influences from genomic DNA flanking the integration site. By fluorescent in situ hybridization, the transgene insertion site was mapped to chromosome 12. Hemizygous and homozygous transgenic mice appear normal and are fertile. AER specific beta-galactosidase staining was detected by 9.5 days post coitum in the forelimb and hindlimb bud. beta-galactosidase staining could be seen throughout the development of the limbs up to 14.5 days post coitum when expression was restricted to the distal-most regions of the digits of the hindlimbs. The loss of beta-galactosidase staining between digits correlated with the onset of programmed cell death, or apoptosis, in the digit interzones. LacZ expression in this transgenic line represents a useful marker for studying AER function in limb specification during mouse embryogenesis. PMID- 10861558 TI - Actin filament disruption blocks cerebellar granule neurons at the unipolar stage of differentiation in vitro. AB - Cerebellar granule neurons developing in vitro initially extend a single axon, with the Golgi apparatus and centrosome positioned at the base of this axon and then begin the transition to a bipolar morphology by rotating the Golgi centrosome to the opposite pole of the cell and extending a secondary axon; granule cells reach a mature, complex morphology by extending multiple, short dendrites by 5-6 days in vitro. (Zmuda and Rivas, 1998. Cell Motil Cytoskel 41:18 38). To test the effects of actin depolymerization on this characteristic pattern of granule cell axonogenesis, cultured granule cells were treated with either cytochalasin D (CD) or latrunculin A (Lat A) to depolymerize filamentous actin. Although actin depolymerization did not inhibit initial axon extension, it prevented the cells from proceeding on to the transitional, bipolar, or complex stages of differentiation, effectively blocking the cells at the unipolar stage of differentiation. Although the Golgi apparatus resided at the base of the axon in nontreated unipolar cells, or at the opposite pole of the cell body in nontreated transitional cells, the Golgi was randomly localized within the cytoplasm of cells that had been treated with either CD or Lat A. These results show that the transition from the unipolar to the bipolar stage and on to more mature stages of granule cell differentiation is dependent on an intact actin cytoskeleton and suggest that the characteristic pattern of granule cell differentiation may be dependent on the repositioning of the Golgi-centrosome during morphological development. PMID- 10861559 TI - Development of utricular otoliths, but not saccular otoliths, is necessary for vestibular function and survival in zebrafish. AB - We have been studying the consequences of embryonic vestibular dysfunction caused by the monolith (mnl) mutation in zebrafish. mnl is a dominant mutation that specifically inhibits formation of utricular otoliths. However, briefly immobilizing mnl/mnl embryos in agarose with the otic vesicle orientated at certain angles selectively induces or prevents formation of utricular and/or saccular otoliths. With this noninvasive technique, we generated six phenotypic classes of mnl/mnl mutants, designated S-S, U-U, U-S, S-US, U-US, and US-US, depending on which otoliths are present on each side (U, utricular otolith; S, saccular otolith). All mnl/mnl larvae survived through day 10 of development. Thereafter, S-S larvae showed a rapid decline, probably because of starvation, and none survived to adulthood. Survival rates in all other classes of mnl/mnl larvae (those having at least one utricular otolith) were close to normal. The presence or absence of utricular otoliths also correlated with vestibular function during early larval development, as measured by three criteria: First, unlike wild-type larvae, S-S mutant larvae showed almost no detectable counter rotation of the eyes when tilted tail up or tail down. Second, 95% of S-S mutant larvae never acquired the ability to maintain a balanced dorsal-up posture. Third, although most wild-type larvae responded to gentle prodding by swimming in a straight line, S-S larvae responded by swimming in rapid circles, showing sudden and frequent changes in direction ("zigzagging"), and/or rolling and spiraling. All other phenotypic classes of mnl/mnl larvae behaved normally in these assays. These data demonstrate that bilateral loss of utricular otoliths disrupts the ability to sense gravity, severely impairs balance and motor coordination, and is invariably lethal. The presence of a utricular otolith in at least one inner ear is necessary and sufficient for vestibular function and survival. In contrast, saccular otoliths are dispensable for these functions. PMID- 10861560 TI - Synaptic localization and axonal targeting of agrin secreted by ventral spinal cord neurons in culture. AB - Agrin secreted by motor neurons is a critical signal for postsynaptic differentiation at the developing neuromuscular junction. We used cultures of chick ventral spinal cord neurons with rat myotubes and immunofluorescence with species-specific antibodies to determine the distribution of agrin secreted by neurons and compare it to the distribution of agrin secreted by myotubes. In addition, we determined the distribution of agrin secreted by isolated chick ventral spinal cord neurons and rat motor neurons grown on a substrate that binds agrin. In cocultures, neuronal agrin was concentrated along axons at sites of axon-induced acetylcholine receptor (AChR) aggregation and was found at every such synaptic site, consistent with its role in synaptogenesis. Smaller amounts of agrin were found on dendrites and cell bodies and rarely were associated with AChR aggregation. Muscle agrin, recognized by an antibody against rat agrin, was found at nonsynaptic sites of AChR aggregation but was not detected at synaptic sites, in contrast to neuronal agrin. In cultures of isolated chick neurons or rat motor neurons, agrin was deposited relatively uniformly around axons and dendrites during the first 2-3 days in culture. In older cultures, agrin immunoreactivity was markedly more intense around axons than dendrites, indicating that motor neurons possess an intrinsic, developmentally regulated program to target agrin secretion to axons. PMID- 10861561 TI - Cdc42 stimulates neurite outgrowth and formation of growth cone filopodia and lamellipodia. AB - To assess the role of cdc42 during neurite development, cmyc-tagged constitutively active (CA) and dominant negative (DN) cdc42 were expressed in dissociated primary chick spinal cord neurons using adenoviral-mediated gene transfer. Three days after infection, >85% of the neurons in infected cultures expressed cdc42 proteins, as detected by indirect immunofluorescence against cmyc. Growth cones of infected neurons displayed 1.83- (CAcdc42) and 1.93-fold (DNcdc42) higher cmyc immunofluorescence per square micrometer than uninfected controls. CAcdc42 expression stimulated growth cones, almost doubling growth cone size and number of filopodia, and increased neurite growth rates by 65-89%. In neurons plated onto fibronectin, the percent of growth cones with both filopodia and lamellipodia increased from 71 to 92%. Total Texas Red-phalloidin staining in these growth cones doubled, and the percent of growth cones with F-actin localized to peripheral regions increased from 52% in controls to 78% after CAcdc42 expression. Expression of DNcdc42 did not significantly alter growth cone morphology or neurite growth rates. Addition of soluble laminin to spinal cord neurons resulted in the identical phenotype as CAcdc42 expression, including changes in growth cone morphology, F-actin localization, and neurite growth rates. Significantly, expression of DNcdc42 blocked the effects of laminin on growth cones. These results show that cdc42 promotes neurite outgrowth and filopodial and lamellipodial formation in growth cones and suggests that cdc42 and laminin share a common signaling pathway during neurite development. Addition of laminin to CAcdc42-expressing neurons is inhibitory to growth cones, indicating that laminin also may activate some other pathways. PMID- 10861562 TI - Disruption of peripheral target contact influences the development of identified central dendritic branches in a leech motor neuron in vivo. AB - Retrograde signaling from target tissues has been shown to influence many aspects of neuronal development in a number of developmental systems. In these experiments using embryonic leeches (Hirudo medicinalis), we examined how depriving a neuron of contact with its peripheral target affects the development of the cell's central arborization. We focused our attention on the motor neuron cell 3, which normally stimulates dorsal longitudinal muscle fibers to contract. At different locations in the periphery and in embryos of several different stages, we cut the nerve containing the growing axon of cell 3. This surgery led to dramatic overgrowth of cell 3's central dendritic branches, which normally accept synaptic contacts from other neurons, including the inhibitory motor neuron cell 1. When cell 3's peripheral axon was cut relatively early in development, its overgrown central branches eventually retracted. However, cells that were disrupted later in development retained their overextended branches into adulthood. In addition, if the axon was cut close to the ganglion early in development, depriving the cell of contact with any dorsal tissues, the central branches failed to retract and were instead retained into adulthood. Unlike cell 3, the central branches of cell 1, which has the same peripheral target muscles as cell 3, remained unchanged following all axotomy protocols. These results suggest that in at least some neurons contact with peripheral targets can influence development of the central processes that normally mediate synaptic contacts. PMID- 10861563 TI - Rhythmic coupling among cells in the suprachiasmatic nucleus. AB - In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a pair of structures in the hypothalamus known as the suprachiasmatic nucleus (SCN). Previous studies suggest that the basic mechanism responsible for the generation of these rhythms is intrinsic to individual cells. There is also evidence that the cells within the SCN are coupled to one another and that this coupling is important for the normal functioning of the circadian system. One mechanism that mediates coordinated electrical activity is direct electrical connections between cells formed by gap junctions. In the present study, we used a brain slice preparation to show that developing SCN cells are dye coupled. Dye coupling was observed in both the ventrolateral and dorsomedial subdivisions of the SCN and was blocked by application of a gap junction inhibitor, halothane. Dye coupling in the SCN appears to be regulated by activity-dependent mechanisms as both tetrodotoxin and the GABA(A) agonist muscimol inhibited the extent of coupling. Furthermore, acute hyperpolarization of the membrane potential of the original biocytin-filled neuron decreased the extent of coupling. SCN cells were extensively dye coupled during the day when the cells exhibit synchronous neural activity but were minimally dye coupled during the night when the cells are electrically silent. Immunocytochemical analysis provides evidence that a gap-junction-forming protein, connexin32, is expressed in the SCN of postnatal animals. Together the results are consistent with a model in which gap junctions provide a means to couple SCN neurons on a circadian basis. PMID- 10861564 TI - Involvement of L1.1 in memory consolidation after active avoidance conditioning in zebrafish. AB - To investigate the involvement of the cell adhesion molecules L1.1, L1.2, NCAM, and tenascin-C in memory formation, zebrafish (Brachydanio rerio) were trained in an active avoidance paradigm to cross a hurdle to avoid mild electric shocks after a light signal. Application of [(14)C]deoxyglucose prior to the training session revealed an increased energy demand in the optic tectum during acquisition of the active avoidance response compared with untrained fish and with fish not learning the task (nonlearners). In situ hybridization with digoxigenin-labeled cRNA probes directed against zebrafish L1.1, L1.2, NCAM, and tenascin-C revealed an enhanced expression of L1.1 and NCAM mRNA in the optic tectum of learners 3 h after acquisition of the task compared with untrained fish, nonlearners, overtrained fish, and learners decapitated 1 or 6 h after acquisition. Levels of L1.2 mRNA were not significantly increased in the tectum 3 h after learning. Tenascin-C was neither expressed in the optic tectum of untrained fish nor in the tectum of learners. To test for a possible involvement of L1.1 in memory consolidation, antibodies were injected intracerebroventricularly 1 h after the last training trial. Two days later, injected zebrafish were tested for recall and evaluated by a retention score (RS), ranging from 1.0 for immediate recall to 0.0 indicating no savings. The average retention score of L1.1 antibody-injected fish (RS = 0. 29) was different from that of tenascin-C antibody-injected (RS = 0. 71) or uninjected fish (RS = 0.78), indicating a pivotal function of L1.1 in long-term memory formation in zebrafish. PMID- 10861565 TI - Spatial, temporal, and sexually dimorphic expression patterns of the fruitless gene in the Drosophila central nervous system. AB - The fruitless (fru) gene of Drosophila produces both sex-specifically and non-sex specifically spliced transcripts. Male-specific fru products are believed to regulate male courtship. To further an understanding of this gene's behavioral role, we examined the central nervous system (CNS) for temporal, spatial, and sexually dimorphic expression patterns of sex-specific fru products by in situ hybridization and immunohistochemistry. For the latter, antibodies were designed to detect only male-specific forms of the protein (FRU(M)) or amino acid sequences that are in common among all translated products (FRU(COM)). Sex specific mRNAs and male-specific proteins were first observed in mature larvae and peaked in their apparent abundances during the first half of the pupal period. At later stages and in adults, faint mRNA signals were seen in only a few neural clusters; in contrast, relatively strong FRU(M) signals persisted into adulthood. Twenty neuronal groups composed of 1700 fru-expressing neurons were identified in the midpupal CNS. These groups overlap most of the neural sites known to be involved in male courtship. Anti-FRU(COM) led to widespread labeling of neural and nonneural tissues in both sexes, but in the female CNS, only in developing ganglia in a pattern different from that of the male's FRU(M) cells. Expression of sex-specific fru mRNAs in the CNS of males analyzed from the earliest pupal stages indicated that sex-specific alternative splicing is not the exclusive mechanism regulating expression of fruitless transcripts. PMID- 10861566 TI - The journal of pathology publishes faster on EarlyView AB - Original papers accepted by The Journal of Pathology are now published online via EarlyView once proofs have been approved by authors. Readers can therefore view papers before they are compiled into a print issue. Authors can cite the EarlyView version of the paper as published online and by using the paper's DOI (digital object identifier). Papers can therefore be read and cited in a shorter time frame, enabling faster publication for authors and a faster response to published work. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10861567 TI - Cell cycle and melanoma--two different tumours from the same cell type. AB - Melanoma of the uvea of the eye and melanoma of the skin share a common cell of origin, but differ substantially in their behaviour and response to chemotherapy. There is increasing evidence that this is related to differences in their molecular phenotype, particularly in relation to the expression of cell cycle associated proteins. Since many cytotoxic agents act by damaging DNA, resistance is often associated with intact mechanisms which allow the neoplastic cells to arrest their growth while DNA is repaired, or to resist apoptosis in response to detection of DNA damage. p53 is important to these processes, but mutation appears to be a less common event in uveal melanoma than in skin melanoma, probably due to the lack of UV exposure in the uvea. There are also differences in proliferation-associated proteins such as c-myc and cyclin D1. Overexpression of the former molecule is associated with a poor prognosis in skin melanoma, but is associated with a good prognosis in uveal melanoma, although there is considerable genetic heterogeneity within each type. While prognostic studies may therefore be of little relevance to individual patients, they continue to inform our understanding of tumour biology. PMID- 10861568 TI - Methods for imaging the structure and function of living tissues and cells: 1. Optical coherence tomography. AB - This is the first in a series of review articles which aim to present a concise and systematic overview of the principles, limitations, advantages, and uses of some of the more important recently developed techniques capable of imaging living histology. Optical coherence tomography (OCT) is now an established optical biopsy method, imaging 2-3 mm into opaque tissue. It is analogous to optical 'ultrasound' but has an outstanding resolution, being capable of imaging single cells in the intact animal via a surface, intravascular or endoscopic approach. Both two-dimensional (2D) and three-dimensional (3D) image datasets can be acquired and studied over time (4D imaging) in the live animal or human subject without the need to remove tissue or perform any tissue processing or staining. It has been used in ophthalmology, gastrointestinal tract (GI) studies, gynaecological tract investigation, and endovascular imaging, to name but a few areas. A degree of differential tissue contrast information can also be gleaned, since different tissue components give different OCT reflectivity signals such that adipose, muscle, collagen, and elastic components may all be resolved without staining. Continuing developments include faster data acquisition for real-time recording and Doppler OCT for more functional imaging. PMID- 10861569 TI - The prognostic value of cyclin D1, p53, and MDM2 protein expression in uveal melanoma. AB - Malignant uveal melanoma is the commonest primary intraocular tumour in adults. It metastasizes frequently and 50% of patients die within 10 years of diagnosis. The expression of cyclin D1, p53, and MDM2 in uveal melanoma and their relationship to metastasis-free 5-year survival was determined, in order to investigate whether these proteins help to distinguish those patients with a favourable prognosis from those with a poorer one. Ninety-six eyes enucleated for uveal melanomas were immunohistochemically analysed for the protein expression of cyclin D1 and related cell-cycle markers, p53 and MDM2. The evaluation of the specimens was undertaken by two independent pathologists without knowledge of the outcome. Statistical analysis of clinical, morphological, and immunohistological features was performed. A 'favourable outcome' was defined as survival of at least 5 years after diagnosis, without metastases (n=57). An 'unfavourable outcome' was defined as death from metastases within the first 5 years after diagnosis of uveal melanoma (n=39). Cyclin D1 positivity (>15% positive tumour cells) as well as p53 positivity (>15% positive tumour cells) was associated with an unfavourable outcome (for cyclin D1: odds ratio=4. 2, 95% confidence interval 1.5-11.8, p=0.006; for p53: odds ratio=3. 2, 95% confidence interval 1.1-9.3, p=0.03). In addition, cyclin D1 positivity was associated with the presence of extraocular extension of the tumour (p=0.01), with the mixed or epithelioid cell type (p=0. 02), and with the tumour cell MIB-1 positivity (p=0.0001). MDM2 immunoreactivity of the tumour cells showed a potential correlation with clinical outcome (odds ratio=2.1, 95% confidence interval 0.8-5. 8, p=0.13). Multiple logistic regression models showed that cyclin D1 positivity is an independent prognostic factor after control for other prognostic markers. The expression of cyclin D1 in uveal melanoma is associated with a more aggressive course and histologically unfavourable disease. This could serve as a further independent prognostic factor in uveal melanoma. PMID- 10861570 TI - EBNA-1 sequence variation in Danish and Chinese EBV-associated tumours: evidence for geographical polymorphism but not for tumour-specific subtype restriction. AB - The Epstein-Barr virus (EBV) nuclear antigen (EBNA)-1 is consistently expressed in EBV-associated tumours. Recently, EBNA-1 carboxy (C)-terminal sequence variants have been described based on the amino acid signature at codon 487, and designated prototype (P)-ala (identical to prototype B95.8 strain), P-thr, variant (V)-val, V-leu, and V-pro. These studies suggest that certain EBNA-1 variants show selective cell tropism and may be preferentially associated with different EBV-positive malignancies; for example, in contrast to P-ala subtypes, V-val appeared to be restricted to the oral compartment and to be associated with undifferentiated nasopharyngeal carcinoma (NPC). To test the hypothesis that V val subtypes are restricted in distribution, EBNA-1 variants were investigated in NPC and throat washings (TWs) from a low (Denmark) and a high (China) NPC risk area. For comparison, cases of Hodgkin's disease (HD) were also studied. V-val was found to be the dominant EBNA-1 subtype, not only in Chinese TWs and NPC biopsies, but also in Chinese HD. Furthermore, V-val was not detected in any of the Danish NPC biopsies or TW samples. These findings show that V-val is not associated with NPC, nor is it restricted to the oral compartment, but rather that it represents a dominant Asian EBNA-1 subtype, both in EBV-associated malignancies and in the general population. PMID- 10861571 TI - p53 immunoexpression in non-malignant oral mucosa adjacent to oral squamous cell carcinoma: potential consequences for clinical management. AB - p53 is a tumour suppressor gene encoding a protein whose function is impaired in a very large proportion of human cancers. The objectives of this study were to determine the natural history of p53 alterations during stages of oral carcinogenesis, by comparing p53 immunoexpression in oral squamous cell carcinomas (OSCCs), their non-malignant adjacent mucosa, and respective metastases; and to define the potential practical consequences for clinical management of p53 staining in the non-malignant adjacent mucosa. Forty-two samples of non-malignant mucosa adjacent to OSCCs, the respective carcinomas, and six lymph node metastases derived from six of the OSCCs were investigated for p53 protein expression by immunohistochemistry. Seven out of 42 (17%) non-malignant mucosal samples immediately adjacent to OSCC showed suprabasal p53 staining and this was significantly associated with moderate/severe dysplasia (p=0.02). In six of these cases (86%), the respective carcinoma showed p53 immunoexpression in more than 50% of the neoplastic cells and in the remaining case, p53 immunoexpression was found in more than 25% of the neoplastic cells. In all p53 negative carcinomas that showed p53 immunoexpression in the non-malignant adjacent mucosa, p53 staining was never detected above the basal cell layer. Lymph node metastases showed the same patterns of p53 immunoexpression as the carcinomas from which they were derived. When suprabasal p53 staining is present in non-malignant mucosa immediately adjacent to OSCCs, this suggests stable p53 alterations which are maintained upon progression to overt malignancy. The immunostaining in non-malignant mucosa of the resection margins of OSCCs might be a valuable predictor for local recurrences and may therefore have implications for the management of patients who have received surgical treatment for OSCC. PMID- 10861572 TI - c-erbB2 oncoprotein expression, gene amplification, and chromosome 17 aneusomy in apocrine adenosis of the breast. AB - Amplification of the c-erbB2 oncogene and numerical aberrations of chromosome 17 occur in human breast carcinomas. Apocrine adenosis (AA) of the breast has been shown occasionally to have c-erbB2 overexpression and a possible premalignant potential, but little is known about cellular level genetic alterations in AA of the breast. Fluorescence in situ hybridization (FISH) is a new approach to detect these. In this study, a series of AA was studied by immunohistochemistry for c erbB2 protein expression and by FISH using dual colour DNA probes for the c-erbB2 gene and the centromeric region of chromosome 17. Cell membrane immunostaining was seen in 10/18 (55.6%) AA cases, but unequivocal c-erbB2 gene amplification or chromosome 17 aneusomy was not seen. The results of this study suggest that c erbB2 overexpression without amplification may occur early in breast oncogenesis. Amplification and numerical chromosome aberrations may occur later in the pathogenesis of apocrine-derived breast carcinomas. PMID- 10861573 TI - Computer-assisted 3D mapping and morphometry of dysplastic zones in endoscopically resected colonic adenomas. AB - Three-dimensional (3D) reconstruction and morphometry of resected colonic adenomas were undertaken to extend current knowledge of clinically significant features such as the frequency of occurrence of cancer, and the size and spatial distribution of dysplastic zones in these tumours. Fifty endoscopically resected colonic adenomas were serially sectioned at intervals of 0.2 mm and the sectional images were loaded into a computer system in order to visualize the spatial distribution of dysplastic zones. These were graded into five groups according to the criteria of Morson and Dawson: normal mucosa, mild dysplasia, moderate dysplasia, severe dysplasia, and cancer. The way in which zones of different grades are distributed in a polyp was visualized in a computer display and the volume of each dysplastic zone was estimated by the Cavalieri principle. In five polyps, adenocarcinoma was found growing in an adenoma. In all of these, the cancer was surrounded by less dysplastic zones, in the form of 'cancer in adenoma'. In pedunculated polyps, submucosal invasion could occur even if the volume percent of severe dysplasia was less than 10%. In such a case, multiple biopsy specimens are advisable. Semipedunculated polyps smaller than 200 mm(3) can also harbour submucosal invasion. In this study it was found that if the adenomas had been examined by only a single section, as many as one in five of the cases in which submucosal invasion had already developed would have escaped microscopic confirmation. To prevent such diagnostic failure, it is advisable to add a few deeper sections. Thus, 3D reconstruction and morphometry have been helpful in establishing a better standard for the diagnostic histopathology of colonic tumours. PMID- 10861574 TI - Fas and Fas ligand: strong co-expression in human hepatocytes surrounding hepatocellular carcinoma; can cancer induce suicide in peritumoural cells? AB - Fas (Apo-1, CD95), a member of the nerve growth factor/tumour necrosis factor receptor superfamily, mediates apoptosis in response to agonistic antibodies or Fas ligand (Fas-L) binding. Fas has been shown to be present on hepatocyte membranes in normal liver and in chronic hepatitis C. At the present time, very limited data are available on the expression of Fas-L. This paper describes a study of 20 cases of active chronic hepatitis of different aetiologies, 20 hepatocellular carcinomas (HCCs) and the adjacent non-tumoural liver parenchyma, and five normal livers. The immunohistochemical expression of Fas and Fas-L was determined using specific monoclonal antibodies. In normal liver, Fas was faintly expressed on membranes of hepatocytes and bile duct cells, while Fas-L was negative. In active chronic hepatitis, Fas expression in hepatocytes was enhanced, resulting in a diffuse honeycomb pattern. Fas-L showed cytoplasmic positivity in hepatocytes in areas of interface hepatitis. Strong expression of Fas as well as Fas-L in the hepatocytes immediately adjacent to HCC was a constant finding. Within the HCCs, Fas-L expression was variable, but present only in a minority of cells. Fas varied from a diffuse honeycomb pattern to focal positivity in occasional cells. There was no correlation between Fas and Fas-L expression in the tumours. In conclusion, hepatocytes can co-express Fas and Fas L in areas of interface hepatitis and adjacent to HCC, suggesting that they have the ability to induce apoptosis in an autocrine or paracrine way. Within the tumour, the Fas-Fas-L apoptosis pathway seems to be little involved. PMID- 10861575 TI - The grade of pancreatic ductal carcinoma is an independent prognostic factor and is superior to the immunohistochemical assessment of proliferation. AB - Tumour grade is one of the prognostic factors in pancreatic ductal adenocarcinoma, but its value is controversial. In this study, the predictive value and the reproducibility of the WHO grading system were reconsidered and the possibility of supplementing it with the immunohistochemically assessed proliferative activity was investigated. Seventy resected ductal adenocarcinomas of the head of the pancreas were evaluated. A total of 60 HPF fields on two to four sections per tumour were screened for glandular differentiation, mucin production, mitosis, and nuclear atypia by two observers with different degrees of experience. Each criterion was scored and the grade was calculated from the mean value of all single scores. Corresponding slides were immunohistochemically stained with the proliferation marker Ki-S5. The percentage of positive nuclei was assessed and a proliferation index (PI) assigned (<10%=1; 10-50%=2; >50%=3). Multivariate analysis (Cox regression) identified grade and R stage as the most significant factors for predicting survival. The PI determined on the basis of Ki S5 staining did not prove to be an independent prognostic factor. In 30 of 70 carcinomas, it correlated with the tumour grade. Within a given tumour grade, the cases with the least favourable prognosis could be distinguished on the basis of their PI. The inter-observer variability was considerable, with the main differences occurring in the group of G1 tumours. According to the refined WHO criteria, the histopathological grade of pancreatic ductal carcinoma is an important independent prognostic factor, but reproducibility depends on the expertise of the observer. Criteria that relate to cellular and structural differentiation seem to be more predictive than those related to proliferation. PMID- 10861576 TI - Molecular analysis of E-cadherin and cadherin-11 in Wilms' tumours. AB - Different studies of Wilms' tumours have demonstrated a loss of heterozygosity (LOH) of chromosome 16q ranging from 17 to 25%. In order to search for a potential tumour suppressor gene on 16q, we chose the calcium-dependent cell adhesion molecules E-cadherin and cadherin-11 as candidate genes, which are both located on the long arm of chromosome 16. E-cadherin is known to be expressed in epithelial structures, whereas cadherin-11 is supposed to be expressed in mesenchymal structures and developing epithelium, including renal tubules. For the present study, fresh frozen tissue from 30 Wilms' tumours and corresponding non-tumour tissues were analysed. Single nucleotide polymorphisms of the E cadherin and cadherin-11 genes were chosen and analysed for allelic inactivation by polymerase chain reaction (PCR) amplification and sequence analysis. Loss of expression of one E-cadherin allele was seen in 10% (2/20) of the informative cases. Two out of 11 informative cases (18%) showed loss of expression of one cadherin-11 allele. No length alterations of either the E-cadherin or the cadherin-11 messenger RNAs were identified using reverse transcription PCR and agarose gel electrophoresis in tumour tissue. Sequencing of the entire E-cadherin coding region in seven cases showed the wild-type sequence. These data imply that E-cadherin and cadherin-11 are not likely to play typical tumour suppressor roles in Wilms' tumour. Interestingly, the E-cadherin immunohistochemistry showed a deviation from the normal reaction pattern in 50% of the cases, with 27% (8/30) showing an apical or cytoplasmic reaction and 23% (7/30) being completely negative. Northern blot analysis revealed that the overall expression of cadherin 11 is much stronger than that of E-cadherin. In several cases, the expression levels of the two genes were inversely correlated, suggesting the existence of a regulatory mechanism. Analysis of differential expression of the various cadherins and their subsequent signal transduction pathways might contribute to a better understanding of the complexity of Wilms' tumour formation. PMID- 10861577 TI - Expression of parathyroid hormone-related protein and its receptor in bone metastases from prostate cancer. AB - Studies of breast cancer suggest that parathyroid hormone-related protein (PTHrP) is important in the development of bone metastases. To determine whether PTHrP expression is important in prostate cancer metastasis, immunohistochemistry and in situ hybridization were used to assess the expression of PTHrP and its receptor in primary prostate cancer and bone metastases from both prostate and non-prostate cancers. PTHrP was expressed in more prostate primary tumours than bone metastases (p=0.003, Fisher's exact test). All bone metastases from non prostate cancers expressed PTHrP. In contrast, PTHrP receptor was expressed in all bone metastases, but in only 19% of primary prostate tumours (p=0.001). The receptor to PTHrP was found to be highly expressed in bone metastases from prostate and other primaries, whereas PTHrP protein was found to have lower expression in the bone metastases than in the primary tumours. In conclusion, the expression of the receptor to PTHrP is increased in bone metastases from prostate cancer and may play an important role in their formation. PMID- 10861578 TI - Proliferative index in phaeochromocytomas: does it predict the occurrence of metastases? AB - Evaluation of the malignant potential of phaeochromocytomas in the absence of metastases presents a formidable challenge to both clinicians and pathologists. Until now, no widely accepted clinical, histological, immunohistochemical or molecular method has become available to discriminate malignant from benign phaeochromocytomas. In other endocrine tumours, estimation of proliferative activity by MIB-1 immunostaining has emerged as a promising approach for the determination of metastatic potential. In this study, the utility of MIB-1 immunostaining as a predictive marker for the occurrence of metastases in phaeochromocytomas was evaluated. In addition, the density of S100-positive sustentacular cells was studied, since their depletion has been identified as a negative predictive marker in smaller series. Furthermore, several clinicopathological parameters were evaluated. One hundred and ten patients operated on for a total of 99 benign and 37 malignant phaeochromocytomas were studied. All malignant tumours had documented metastases. The histopathological diagnosis of primary tumours and metastases was reviewed and graded for angioinvasion, capsular extension, and intra-tumoural necrosis. The proliferative index (percentage of MIB-1-positive cells) and the density of S100-positive cells were assessed. In addition, age at resection, associated familial tumour syndromes, tumour size, and tumour location were recorded. Univariate analysis revealed statistically significant correlations between malignancy and proliferative index (p<0.0005) and depletion of S100-positive sustentacular cells (p<0.0005). Fifty per cent of the malignant, but none of the benign phaeochromocytomas had a proliferative index greater than 2.5%. Higher age at resection (p=0. 03), sporadic occurrence (p<0.0005), extra-adrenal location (p<0. 0005), tumour size (p<0.0005), and necrosis (p=0.03) were also significantly associated with malignancy. Logistic regression showed that proliferative index (p=0.0072), size (p=0.0022), and extra-adrenal location (p=0.0012) of the primary tumour were independently predictive for malignancy. In conclusion, this study indicates that assessing the proliferative activity of phaeochromocytomas by MIB 1 immunohistochemistry can predict the occurrence of metastases. The predictive value of S100 immunostaining, tumour size, and extra-adrenal location of the tumour was also confirmed. PMID- 10861579 TI - Nucleolar size indicates the rapidity of cell proliferation in cancer tissues. AB - In order to define the importance of the nucleolus in tumour pathology, the relationship between nucleolar size and function and tumour mass growth rate was studied in vivo. Ten established human cancer cell lines from colon carcinomas and neuroblastomas were inoculated subcutaneously in athymic mice and the doubling time (DT) of the xenograft tumour mass was calculated. The tumour DTs ranged from 3.2 to 15.7 days. Nucleolar size was evaluated in sections from formalin-fixed and paraffin-embedded tumour samples after silver staining for AgNOR proteins, using a specific image analysis system. The nucleolar area values were inversely related to the xenograft tumour mass DTs (r=-0.90; p<0.001). Nucleolar functional activity was also evaluated using rapid, intermediate, and slow growing tumours (one each). The values of RNA polymerase I activity measured in vitro were strongly related to the corresponding tumour DTs (r=-0. 99; p=0.03). The labelling indices (LIs) of three proliferation markers, MIB1, PCNA, and bromodeoxyuridine (BrdU), were also evaluated. As revealed by the MIB1 and PCNA LIs, almost all the cells of the xenograft tumours were cycling (86.6+/-5.6 SD and 95. 5+/-2.0 SD, respectively). Neither the MIB1, PCNA or BrdU LIs were related to the xenograft tumour mass DT, showing that the different growth rates of tumour xenografts were not due to different growth fractions, but were mainly related to different cell proliferation rates. The present data demonstrate that the size and function of the nucleolus are related to the cell proliferation rate of cancer tissue. Evaluation of nucleolar size after silver staining of AgNOR proteins represents a unique parameter for the histological assessment of rapidity of cell proliferation in tumour lesions. PMID- 10861580 TI - Comparative genomic hybridization of microdissected samples from different stages in the development of a seminoma and a non-seminoma. AB - Human testicular germ cell tumours (TGCTs) of adolescents and adults, both seminomas and non-seminomas, originate from intratubular germ cell neoplasia (IGCN). Comparative genomic hybridization (CGH) was applied to microdissected samples from different stages of the development of a seminoma and a mixed non seminoma, including IGCN of both. The different stages of the seminoma development, namely IGCN, intratubular and invasive seminoma, showed a very similar pattern of chromosomal imbalances, including gains of parts of 7, 8, 12,14, and X, and losses of parts of 3, 4, 5, 10, 11, 12q, 16, 18, 22, and Y. A more heterogeneous pattern was found for the non-seminoma. Some aberrations were present only in IGCN, or in IGCN and in all invasive components (gains of parts of 1q, 17, 19p, 20q, and 22, and losses of parts of 4, 5, 9p, 13, and 18q), while others were present in a less consistent pattern. These are the first reported CGH data from different stages in the development of TGCTs. Although only two cases were studied, the results suggest that particular numerical changes of (parts of) chromosomes are involved in the early development and progression of this cancer. PMID- 10861581 TI - Dissimilar anti-tumour reactions induced by tumour cells engineered with the interleukin-2 or interleukin-15 gene in nude mice. AB - Interleukin (IL)-15 shares immuno-stimulatory properties with IL-2 and is a potent inducer of natural killer (NK) cell function. The major histocompatibility complex (MHC) class I-negative human small cell lung cancer (SCLC) cell line N592, engineered to express a modified IL-15 cDNA (N592/IL-15), secreted biologically active IL-15 (300-500 pg/ml), capable of boosting T-cell proliferation and NK activity 'in vitro'. The effect of IL-15 gene transfer on natural immunity 'in vivo' was assessed by xenotransplants in nude mice and compared with that of the IL-2 gene. N592 cells engineered with IL-2 (N592/IL-2) were promptly rejected, while N592/IL-15 displayed a significant delay in tumour growth and a slightly reduced take rate. However, in NK-depleted nude mice, N592/IL-15 displayed the same growth kinetics as unmodified N592 cells, and N592/IL-2 grew with slightly reduced kinetics. An impressive reactive cell infiltration, consisting mainly of macrophages and granulocytes, was associated with N592/IL-2 tumour rejection, while a more evident recruitment of NK cells was found in N592/IL-15 tumours. In both N592 transfected tumours, we found expression of chemoattractant molecules, such as granulocyte macrophage-colony stimulating factor (GM-CSF) and monocyte chemoattractant protein (MCP)-1, while macrophage inflammatory protein (MIP)-2 was produced by endothelial cells only in N592/IL-2 tumours. In this tumour, very few and severely damaged microvessels were found, while microvessels were numerous in N592/IL-15 tumours. The potent recruitment of NK cells mediated by IL-15 gene transfer suggests its possible therapeutic use in tumours lacking MHC class I. PMID- 10861582 TI - Enhanced expression of vascular endothelial growth factor in pulmonary plexogenic arteriopathy due to congenital heart disease. AB - Congenital heart disease (CHD) leading to increased pulmonary blood pressure and flow is an important cause of pulmonary plexogenic arteriopathy (PPA). This type of arteriopathy tends to progress to an irreversible stage, hallmarked histologically by the emergence of a number of characteristic lesions, which include concentric laminar intimal proliferation and fibrosis, and plexiform lesions. The pathogenesis of these lesions, which connote a very poor prognosis, is not well understood. Since endothelial cell proliferation has been demonstrated in these lesions, it was hypothesized that vascular endothelial growth factor (VEGF), a key mediator of angiogenesis, might play a role in their pathogenesis. Thirty-nine patients with various types of CHD, who underwent cardiac catheterization and subsequent cardiac surgery, were studied prospectively. On the basis of a detailed assessment of the type of cardiac defect, the haemodynamic abnormalities, and the histopathological features evident from open lung biopsies, taken in all instances, patients were histologically grouped into cases with moderate PPA (n=18), advanced PPA (n=7), pulmonary congestive vasculopathy (PCV, n=5), and controls lacking pulmonary hypertension or increased pulmonary blood flow (n=4). Five patients were excluded from analysis because of inadequate sample size or quality. The presence of VEGF was assessed immunohistochemically using standard procedures and was correlated with haemodynamic and histological data. Immunoreactive VEGF was detected in pulmonary arterial smooth muscle cells and endothelial cells in 13 out of 34 cases and was more frequent and more pronounced in patients with the histological lesions of advanced PPA than in those with moderate PPA (p<0.01). VEGF positivity was particularly prominent in the lesions characteristic of advanced PPA. No difference in VEGF expression was observed between controls, PVC, and moderate PPA cases. Measured haemodynamic parameters did not differ significantly between VEGF-positive and VEGF-negative cases. We conclude that VEGF may play a role in the angioproliferative changes of advanced PPA. PMID- 10861583 TI - Altered expression of high-molecular-weight calmodulin-binding protein in human ischaemic myocardium. AB - A high-molecular-weight calmodulin-binding protein (HMWCaMBP) was previously identified and purified from the cytosolic fraction of bovine heart. Based on the sequence homology, amino acid analysis, antibody reactivity, and calpain inhibition, HMWCaMBP has been identified as a homologue of the calpain inhibitor calpastatin. In the present study the expression of HMWCaMBP was investigated in normal and ischaemic human myocardium. Western blot analysis of normal human cardiac muscle extract with the polyclonal antibody raised against bovine HMWCaMBP indicated a prominent immunoreactive band with a molecular mass of 140 kD. HMWCaMBP was localized in the cytoplasm and myofilaments of cardiac myocytes. Furthermore, Western blot analysis of normal and ischaemic cardiac tissues indicated a decrease in the expression of HMWCaMBP in ischaemic tissues. These studies were further substantiated by immunohistochemical studies, indicating strong to moderate HMWCaMBP immunoreactivity in normal cardiac muscle and poor to negative immunoreactivity in ischaemic muscle. The results obtained from the rat ischaemic model suggested that the expression of cardiac HMWCaMBP was significantly decreased during ischaemia/reperfusion. In addition, micro-calpain and m-calpain expression was higher in ischaemic cardiac tissue samples than in normal controls. The calpain inhibitory activity of ischaemic cardiac tissues was significantly lower than normal cardiac tissue samples. In some cases of cardiac ischaemia, HMWCaMBP highlighted the contraction band necrosis seen at the margins of a myocardial infarct. In vitro, HMWCaMBP was proteolysed by micro-calpain and m-calpain. These results indicate that HMWCaMBP could be susceptible to proteolysis by calpains during ischaemia or reperfusion and may play a contributory role in myocardial injury. PMID- 10861584 TI - Follicular dendritic cells share a membrane-bound protein with fibroblasts. AB - The expression of a fibroblast antigen (AS02) on a proportion of CD21+ follicular dendritic cells (FDCs) provides evidence in support of their fibroblastic reticular origin. This antigen is expressed on the membrane of tissue fibroblasts but is absent from lymphocytes, macrophages or granulocytes. The distribution of AS02 in conjunction with other FDC markers (DRC-1, RFD3, CD23, IgM, and vitronectin) showed six types of FDCs. AS02 is present in the outer layers of primary and secondary follicles, but gradually decreases and disappears in the centre of germinal centres. In contrast, there is a progressive up-regulation of the other FDC markers. AS02 is re-expressed in involuting FDCs. Intermediate forms from fibroblastic to dendritic appearance are also apparent and occasionally FDC processes contain collagen type I and IV fibres, a characteristic feature of fibroblasts. In pathological follicles the normal differentiation pattern is disrupted, with persistence of the fibroblast marker, possibly due to altered interactions between FDCs and disrupted lymphocytic patterns. These findings provide new evidence for a local differentiation pathway of fibroblasts to mature FDCs. PMID- 10861585 TI - The physical properties of biogels and their permeability for macromolecular drugs and colloidal drug carriers. AB - Macromolecular drugs, either free or complexed with colloidal drug carriers, have created a great deal of interest during the last decade. If one wants to administer these new therapeutics via the oral, nasal, and cervical routes or through the conductive airways, one of the first barriers to overcome is the mucus layer that adheres to the related epithelia. In this review, the physicochemical properties of biogels, macromolecular drugs, and colloidal drug carriers that play a major role in transport through biogels are reviewed. Also, methods of studying the mobility of macromolecular drugs and colloidal drug carriers in and through biogels are addressed. PMID- 10861586 TI - Bioadhesive-based dosage forms: the next generation. AB - Prolonged contact time of a drug with a body tissue, through the use of a bioadhesive polymer, can significantly improve the performance of many drugs. These improvements range from better treatment of local pathologies to improved drug bioavailability and controlled release to enhanced patient compliance. There are abundant examples in the literature over the past 15 years of these improvements using first generation or "off-the-shelf" bioadhesive polymers. The present mini-review will remind us of the success achieved with these first generation polymers and focus on proposals for the next-generation polymers and attendant benefits likely to occur with these improved polymeric systems. PMID- 10861587 TI - A self-complementary, self-assembling microsphere system: application for intravenous delivery of the antiepileptic and neuroprotectant compound felbamate. AB - Felbamate (FBM) is a novel antiepileptic drug (AED) and neuroprotectant (NP) compound that interacts with strychnine-insensitive (SI) glycine receptors in brain (IC(50) = 374 microM). FBM concentrations required to interact with SI glycine receptors are consistent with brain levels following oral and intraperitoneal administration of AED and NP doses. Because of the solubility limits of FBM, an intravenous (iv) form has not been developed. Nevertheless, an iv form could be important for the treatment of disorders such as status epilepticus and neuronal damage due to hypoxic/ischemic events. Substituted diketopiperazines precipitate in acid to form microspherical particles of uniform size ( approximately 2 microm). The microsphere system entraps drugs on precipitation and dissolves near physiological pH to release the drug cargo. Therefore, microspheres were used to produce an iv formulation of FBM. Mice were administered the FBM/microsphere (20-60 mg/kg FBM) and tested for protection against tonic extension seizures using maximal electroshock. The FBM/microsphere was effective in a time- and dose-dependent manner following iv administration. The median effective dose (ED(50)) for protection against MES seizures at 30 min was 27.2 mg/kg [95% confidence interval (CI) = 20.8-33.4, slope = 6.5]. The ED(50) for minimal motor impairment at 30 min was 167 mg/kg (95% CI = 155-177, slope = 28.1). Thus, the feasibility of encapsulating FBM or similar aqueous insoluble compounds in a microsphere system with delivery by the iv route for treatment of epilepsy and various central nervous system disorders has been clearly demonstrated. Studies were performed in accordance with the Guide for the Care and Use of Laboratory Animals. PMID- 10861588 TI - Flow-injection analysis of dopamine in injections with a periodate-selective electrode. AB - Dopamine determination in pharmaceutical preparations based on its oxidation with periodate (IO(4)(-)) using a new IO(4)(-)-selective electrode under flow conditions is presented. An electrode with a tubular configuration, no internal reference solution, and a PVC (31. 2%) membrane, with metaperiodate bis(triphenylphosphoranylidene)ammonium (1.3%) as ion exchanger and 2 nitrophenyloctylether (67.5%) as mediator solvent, was used. Optimization procedures were directed at potentials versus dopamine readings instead of potential versus the remaining IO(4)(-). This approach was achieved by selecting a 50-cm reactor and an overall flow of 7 mL/min, and injecting 70 microL of dopamine standards in a 3.0 x 10(-4) M IO(4)(-) solution. Under these conditions, a linearity range of 8.0 x 10(-3) to 2.7 x 10(-1) g/L, with a slope of 310.1 +/- 7.4 mV L g(-1) and a reproducibility of +/-0.4 mV, were recorded (n = 8). Interference from common excipients was negligible. Under these conditions, analysis of dopamine injections (n = 12) presenting 200 mg/injection gave average and standard deviation values of 201.0 and 3.3 mg/injection, respectively. A simple and inexpensive flow-injection analysis (FIA) manifold, with a good potentiometric detector, enabled the analysis of 200 samples/h without requiring pretreatment procedures. Comparison with the dopamine injection analysis in the United States Pharmacopoeia monograph showed good accuracy, with a relative deviation of -0.2%. PMID- 10861589 TI - The degradation of the antitumor agent gemcitabine hydrochloride in an acidic aqueous solution at pH 3.2 and identification of degradation products. AB - A study of the degradation kinetics of gemcitabine hydrochloride (2'-deoxy-2',2' difluorocytidine) in aqueous solution at pH 3.2 was conducted. The degradation of gemcitabine followed pseudo first-order kinetics, and rate constants were determined at four different temperatures. These rates were used to construct an Arrhenius plot from which degradation rates at lower temperatures were extrapolated and activation energy calculated. Four major degradation products were identified. Only one of these degradation products, the uridine analogue of gemcitabine, was a known degradation product of gemcitabine and was identified by comparison with synthesized material. The other three degradation products were isolated and characterized by spectroscopic techniques. Two of these products were determined to be the diastereomeric 6-hydroxy-5, 6-dihydro-2'-deoxy-2',2' difluorouridines, and the other product was determined to be O(6),5'-cyclo-5,6 dihydro-2'-deoxy-2', 2'-difluorouridine. The mechanisms of formation of these degradation products are discussed. PMID- 10861590 TI - Synergistic effect of low-frequency ultrasound and sodium lauryl sulfate on transdermal transport. AB - Application of low-frequency ultrasound has been shown to enhance transdermal transport of drugs (low-frequency sonophoresis). In this paper, we show that the efficacy of low-frequency ultrasound in enhancing transdermal transport can be further increased by its combination with sodium lauryl sulfate (SLS), a well known surfactant. The dependence of the ultrasound-SLS-mediated transport on ultrasound parameters, including intensity, net exposure time, and duty cycle, is discussed. The transdermal transport enhancement is proportional to ultrasound intensity as well as to exposure time, and is independent of duty cycle as long as the net exposure time is the same. The synergistic effect of SLS and ultrasound on transdermal transport increases linearly with SLS concentration. The enhancement is also proportional to the ultrasound energy density beyond a threshold value, which suggests that a certain minimum amount of energy density is required before noticeable changes in skin permeability occur. A similar dependence of the transdermal transport enhancement on energy density is observed in the case of the enhancement induced by ultrasound alone. Although the threshold energy density value in the presence of SLS is about 10 times lower than that in the case of ultrasound alone, the relationship between enhancement and energy density in the presence and in the absence of SLS is otherwise similar. Possible mechanisms for the synergistic effect of ultrasound and SLS are also discussed. PMID- 10861591 TI - Polycarbophil-cysteine conjugates as platforms for oral polypeptide delivery systems. AB - The purpose of the present study was to evaluate the potential of polycarbophil cysteine conjugates as carrier systems for orally administered peptide and protein drugs. Mediated by a carbodiimide, cysteine was covalently attached to polycarbophil. The properties of resulting conjugates, displaying 35-50 microM thiol groups per gram of polymer, to bind polypeptides and to inhibit pancreatic proteases was evaluated in vitro. Results demonstrated that only some polypeptides are immobilized to the polycarbophil-cysteine conjugate. Due to the covalent attachment of cysteine to polycarbophil, the inhibitory effect of the polymer toward carboxypeptidase A (EC 3.4. 17.1) and carboxypeptidase B (EC 3.4.17.2) could be significantly (p < 0.05) improved. As the zinc binding affinity of polycarbophil could be improved by the covalent attachment of cysteine, the raised inhibitory effect seems to be based on the complexation of this divalent cation from the enzyme structure. Whereas the covalent attachment of cysteine on polycarbophil had no influence on the enzymatic activity of trypsin (EC 3.4.21.4) and elastase (EC 3.4.21. 36), the inhibitory effect of the polymer-cysteine conjugate toward chymotrypsin (EC 3.4.21.1) was significantly (p < 0.05) higher than that of the unmodified polymer. Because of these inhibitory features, polycarbophil-cysteine conjugates seem to be a promising tool in protecting orally administered therapeutic polypeptides, which are not bound to the polymer, from presystemic metabolism in the intestine. PMID- 10861592 TI - Sustained and controlled release of daunomycin from cross-linked poly(aldehyde guluronate) hydrogels. AB - We have incorporated daunomycin, an antineoplastic agent, into a biodegradable hydrogel through a labile covalent bond. In brief, sodium alginate was chemically broken down to low molecular weight and followed by oxidation to prepare poly(aldehyde guluronate). Adipic dihydrazide was used to incorporate the drug into the polymer backbone and cross-link the polymer to form hydrogels. Daunomycin can be released from the hydrogel after the hydrolysis of the covalent linkage between the drug and the polymer. A wide range of release profiles of daunomycin (e.g., from 2 days to 6 weeks) has been achieved using these materials, and the biological activity of the released daunomycin was maintained. PMID- 10861593 TI - Liquid chromatography-mass spectrometry and proton nuclear magnetic resonance characterization of trace level condensation products formed between lactose and the amine-containing diuretic hydrochlorothiazide. AB - Trace levels of condensation products between lactose and the amine-containing diuretic hydrochlorothiazide are formed when a mixture of the two solids containing 30% weight water is heated at 60 degrees C for 2 weeks. The two most abundant condensation products were characterized by liquid chromatography-mass spectrometry (LC-MS) and proton nuclear magnetic resonance ((1)H NMR) spectroscopy. Under these relatively mild conditions of formation, the amine lactose reaction products are limited to those involving the elimination of only a single molecule of water, rather than the multiple-water eliminations associated with later stages of the Maillard reaction. The spectroscopic data clearly show that the primary condensation products are cyclic N-substituted glycosylamines rather than Schiff base, 1,2-enolic forms, or Amadori rearrangement products of identical mass. In solution, the two most abundant N substituted glycosylamines are shown to be in a kinetically slow equilibrium with each other, most likely through a mutarotation involving the intermediate formation of the acyclic Schiff base. PMID- 10861594 TI - Stable reversed vesicles in oil: characterization studies and encapsulation of model compounds. AB - The formation of reversed sucrose ester vesicles in silicon oil and mixtures of silicon oil and isopropyl palmitate was studied. The vesicles were characterized by polarized light microscopy, freeze-fracture electron microscopy, and differential scanning calorimetry. Furthermore the ability to encapsulate p aminobenzoic acid and cholesterol in such vesicles was studied. The vesicles were multilamellar and had sizes up to several micrometers. The vesicles agglomerated but did not show fusion for at least 2 years when stored at room temperature in glass vials. The encapsulation efficiency of both p-aminobenzoic acid and cholesterol strongly depended on the oil phase in which the vesicles were prepared. Reversed sucrose ester vesicles in silicon oil encapsulated nearly 100% of the amount of p-aminobenzoic acid or cholesterol present in the dispersion. These compounds were encapsulated in different compartments of the vesicles. Reversed sucrose ester vesicles offer new perspectives regarding the development of novel pharmaceutical dosage forms. PMID- 10861595 TI - The ascent of pulmonary drug delivery. AB - The origins of inhalation therapy can be traced back to the early civilizations but this route of administration was relatively uncommon until recently. Direct delivery of drugs to the lung by inhalation for the treatment of respiratory disease grew rapidly in the second half of the 20th century as a result of the availability of effective asthma drugs in convenient, portable delivery systems. In the search for non-invasive delivery of biologics, it was discovered that the large highly absorptive surface area of the lung could be used for systemic delivery of proteins such as insulin. New delivery systems with efficiency and reproducibility to match the high cost and therapeutic constraints of biologics are currently in late stage clinical trials. Even small molecular weight drugs previously administered by injection are tested via the inhalation route either to provide non-invasively rapid onset of action, or to improve the therapeutic ratio for drugs acting in the lung. Gene therapy of pulmonary disease is still in its infancy but could provide valuable solutions to currently unmet medical needs. The beginning of the new millennium is therefore likely to witness development of many valuable therapeutic products delivered by inhalation. PMID- 10861596 TI - Study of phase behavior of poly(ethylene glycol)-polysorbate 80 and poly(ethylene glycol)-polysorbate 80-water mixtures. AB - Mixtures of poly(ethylene glycols) (PEGs) with polysorbate 80 are often used to dissolve poorly water-soluble drugs in dosage forms, where polysorbate 80 helps either in enhancing dispersion or in inhibiting precipitation of drugs once the solution is mixed with water. Binary phase diagrams of polysorbate 80 with several low molecular weight PEGs and a ternary phase diagram of polysorbate 80 with PEG 400 and water are presented. Two phases were observed in the binary mixtures when the concentration of PEG 200, PEG 300, PEG 400, or PEG 600 was >55%(w/w). The miscibility of the binary mixtures increases with an increase in temperature; the upper consolute temperatures of PEG 200-polysorbate 80, PEG 300 polysorbate 80, PEG 400-polysorbate 80, and PEG 600-polysorbate 80 mixtures were 100, 85, 75, and 40 degrees C, respectively. The upper consolute temperature of PEG 1000-polysorbate 80 could not be determined because the melting temperature of the mixtures is approximately 40 degrees C and the consolute temperature appeared to be less than this temperature. The decrease in upper consolute temperature with an increase in PEG molecular weight indicated a greater miscibility of the two components. In the ternary system, phase separation of polysorbate 80 was observed when the concentration of PEG 400 was >50-60 % (w/w), possibly because of the high exclusion volume of PEG 400. PMID- 10861597 TI - Intestinal absorption of octreotide: N-trimethyl chitosan chloride (TMC) ameliorates the permeability and absorption properties of the somatostatin analogue in vitro and in vivo. AB - Octreotide acetate is a somatostatin analogue used for the control of endocrine tumors of the gastrointestinal (GI) tract and the treatment of acromegaly. The oral absorption of octreotide is limited because of the limited permeation across the intestinal epithelium. Both chitosan hydrochloride and N-trimethyl chitosan chloride (TMC), a quaternized chitosan derivative, are nonabsorbable and nontoxic polymers that have been proven to effectively increase the permeation of hydrophilic macromolecules across mucosal epithelia by opening the tight junctions. This study investigates the intestinal absorption of octreotide when it is coadministered with the polycationic absorption enhancer TMC. Caco-2 cell monolayers were used as an in vitro intestinal epithelium model, and male Wistar rats were used for in vivo studies. Octreotide with or without polymers (TMC; chitosan hydrochloride) was administered intrajejunally in rats, and serum peptide levels were measured by radioimmunoassay. All applications and administrations were performed at neutral pH values (i.e., pH = 7.4). In vitro transport studies with Caco-2 cells revealed an increased permeation of octreotide in the presence of TMC. Enhancement ratios ranged from 34 to 121 with increasing concentrations of the polymer (0.25-1.5%, w/v). In rats, 1.0% (w/v) TMC solution significantly increased the absorption of the peptide analogue, resulting in a 5-fold increase of octreotide bioavailability compared with the controls (octreotide alone). Coadministration of 1.0% (w/v) chitosan hydrochloride did not enhance octreotide bioavailability. These results in combination with the nontoxic character of TMC suggest that this polymer is a promising excipient in the development of solid dosage forms for the peroral delivery and intestinal absorption of octreotide. PMID- 10861598 TI - Structure determination from conventional powder diffraction data: application to hydrates, hydrochloride salts, and metastable polymorphs. AB - Recent advances in crystallographic computing have made it possible to solve by powder diffraction methods structures that have not been possible to solve by single-crystal methods. Although there is vast improvement in the quality of data obtained from high-intensity synchrotron radiation, we found that surprisingly reliable results can be obtained from conventional laboratory sources. In this article we examine the application of Monte Carlo/simulated annealing methods for the determination of structures ranging in complexity from 9 to 15 degrees of freedom. We re-determine the structures of papaverine hydrochloride and erythromycin A dihydrate by the powder diffraction method and compare the structures to those determined by single-crystal diffraction methods. The structure of a metastable polymorphic form of acetohexamide, form B, is solved and examined spectroscopically. Its structure has not previously been solved by single-crystal techniques because of the small size of its crystals. PMID- 10861599 TI - The italian society of surgical oncology (SICO) PMID- 10861600 TI - Locally advanced rectal cancer: a multivariate analysis of outcome risk factors. AB - BACKGROUND AND OBJECTIVES: Stages II and III rectal tumors are known as locally advanced rectal cancer (LARC) because they are characterized by a high incidence of local and distant relapses and a low probability of long-term survival. Adjuvant treatments have been advocated to ameliorate overall survival (OS), local recurrence-free survival (LRFS), and metastasis-free survival (MFS) without a univocal beneficial trend. The aim of this study was to identify the independent predictive factors of OS, LRFS, and MFS which could best select patients for adjuvant treatment of LARC. METHODS: Of 153 rectal cancer cases seen consecutively from 1991 to 1998, we studied the main clinical and pathological parameters of 73 LARCs. Clinical and pathological variables were studied by univariate analysis, and independent predictive factors were identified by multivariate analysis. RESULTS: Stages II and III rectal cancer have shown not statistically different rates of OS, LRFS, and MFS. Factors independently associated with increasing OS and MFS were low preoperative carcinoembryonic antigen level (CEA), low number of metastatic lymph nodes, low percentage of metastatic lymph nodes out of the total number of lymph nodes excised, and adjuvant treatment. Increased staging and distal resection margins < or =1 cm were shown to be independent detrimental risk factors regarding OS and MFS, respectively. Independent prognostic factors associated with a reduction in LRFS were advanced age, Hartman's resection, distal resection margins < or =1 cm, and fewer than 14 resected nodes. CONCLUSIONS: Whereas stage I rectal cancer can be treated with a good probability of cure by surgery alone, avoiding adverse effects of adjuvant regimens, the outcome of LARC appears to be positively influenced by adjuvant therapies. In LARC, an accurate study of risk factors would be useful to identify which subset of patients could be favorably influenced by postoperative radiochemotherapy. PMID- 10861601 TI - Evaluation of anal function after surgery for rectal cancer. AB - BACKGROUND AND OBJECTIVES: Sphincter-saving surgical procedures for rectal cancer have been legitimized if executed respecting the criteria of oncological radicalness. Our objective was to evaluate anal sphincteric continence after rectal cancer surgery. METHODS: A detailed questionnaire regarding continence was administered to 3 groups of patients. Group 1 was composed of 9 patients treated with a higher (>4 cm), stapled colorectal anastomosis; the 9 group 2 patients were treated with a low (< or =4 cm), stapled colorectal anastomosis; the 9 group 3 patients underwent coloanal anastomosis. RESULTS: The results were evaluated about 3 years after surgery. Continence was excellent in group 1 and very good in group 2. However, in group 3, we observed diminished gas/feces discrimination, reduced ability to postpone evacuation, and increased soiling and perianal rash. CONCLUSIONS: Anal sphincteric continence was better after surgery with a high or low colorectal anastomosis than after coloanal anastomosis. PMID- 10861602 TI - Long-term treatment with sulindac in familial adenomatous polyposis: is there an actual efficacy in prevention of rectal cancer? AB - BACKGROUND AND OBJECTIVES: Ileorectal anastomosis (IRA) is still used in the treatment of familial adenomatous polyposis (FAP). Sulindac appears to induce regression of colorectal adenomas; however, its effects in long-term therapy and in preventing carcinoma remain unclear. METHODS: Fifteen FAP patients treated by IRA received sulindac (200 mg/day) for a mean period of 48.6 +/- 28.7 (range 12 124) months. Number, size, and type of rectal polyps were assessed by endoscopic and histological evaluation every 6 months. RESULTS: Significant regression of polyps was observed in all patients after 6 months (P < 0.02). However, after a mean of 48.6 +/- 28.7 months, both number and size of polyps increased again, showing no statistical difference with baseline values. Minute polyps appeared reddish, while the largest lesions were flat or slightly elevated. Endoscopic polypectomy was necessary in 9 patients and transanal surgical excision in 3. Two patients were submitted to restorative proctectomy because of a large polyp with severe dysplasia and a rectal cancer, respectively. CONCLUSIONS: Sulindac appears to influence the morphological appearance of polyps in FAP patients, inducing apparent regression. However, at a dose of 200 mg, it does not influence the progression of polyps toward a malignant pattern. PMID- 10861603 TI - Total mesorectal excision for surgical treatment of rectal cancer. AB - BACKGROUND AND OBJECTIVES: The aim of our study was to retrospectively evaluate the results of 2 groups of patients admitted and treated for rectal cancer. METHODS: One hundred and fifty-one patients were available for evaluation. Eighty (group A) were radically operated with the standard technique; 71 (group B) underwent total mesorectal excision (TME). Groups were similar according to demographics, staging, and pathological data. Mean follow-up was 73.5 months. RESULTS: No operative mortality was observed. Complications were 15% in group A and 32% in group B. Local recurrence rates were 41.2% in group A and 12.6% in group B. Distant metastases occurred in 21.2% and 7.6%, respectively, in groups A and B. Cancer-related mortality was 62.5% in the non-TME group and 19.5% in the TME group. Overall 5-year survival rates were 32.4% in group A and 70.5% in group B. Disease-free survival rates were 25% in group A and 62.3% in group B. CONCLUSIONS: TME appears to lower the incidence of cancer-related mortality, with a higher incidence of postoperative complications. Further studies need to be done to assess the real benefits of TME in the surgical treatment of rectal cancer. PMID- 10861604 TI - Result of liver resection as treatment for metastases from noncolorectal cancer. AB - BACKGROUND AND OBJECTIVES: While liver resection for metastatic disease from colorectal cancer extends survival in selected patients, the efficacy of hepatectomy for metastases from other malignancies has not been defined. METHODS: Between 1988 and 1998, 20 hepatic resections were performed in 18 patients (2 underwent a double resection due to recurrence) as treatment of noncolorectal metastases. One-, 2-, and 5-year overall and disease-related actuarial survival rates were calculated. RESULTS: No intraoperative or early postoperative deaths were reported. Seven minor (30%) and 1 major (5%) postoperative complications occurred; mean blood loss was 401 +/- 324 ml. In 25% of patients, intra- or postoperative blood transfusion was needed. The mean postoperative hospital stay was 13. 2 days (range 9-23). The overall actuarial survival rate was 54% at 1 year, 42% at 2 years, and 21% at 5 years (mean 38 +/- 11 months). Survival is related to the primary tumor nature. CONCLUSIONS: Hepatic resection for metastases from noncolorectal carcinoma is safe and feasible, with a relatively low incidence of intra- or postoperative complications and a short hospital stay. Although it achieves good results in terms of survival in patients suffering from neuroendocrine metastases, it could also have a cytoreductive effect for other tumors. PMID- 10861605 TI - Clinicopathologic characteristics and outcome indicators in node-negative gastric cancer. AB - BACKGROUND AND OBJECTIVES: The relationship between the number of lymph nodes examined and the outcome in patients with node-negative (N(-)) gastric cancer was studied. We compared N(-) patients to those with nodal involvement (N(+)) to identify clinicopathologic characteristics of N(-) gastric cancer. Finally, we evaluated outcome indicators in this group of patients. METHODS: Of 367 patients, 130 (35.4%) were N(-). These patients were stratified according to the main prognostic variables, to assess differences with N(+) cases. A statistical analysis using the Cox model was performed to estimate outcome indicators. RESULTS: N(-) gastric carcinomas were significantly different from N(+) cases in terms of tumor depth and site, TNM stage, grading, residual disease, and vessel involvement. The overall 5-year survival rate was 72%. It was 82% in those patients with more than 15 nodes retrieved and 59% in the others. Serosal involvement, residual disease, and poor differentiation were independent prognostic factors. CONCLUSIONS: The clinicopathologic factors and outcome of N( ) cases were similar to those of early gastric cancer. At least 15 examined nodes appears to be necessary to define a case as N(-). The prognostic value of D2 lymphadenectomy in N(-) patients suggests a biologic role of micrometastases. PMID- 10861606 TI - Clinicopathologic findings and results of surgical treatment in cardiac adenocarcinoma. AB - BACKGROUND AND OBJECTIVES: There is a great deal of controversy regarding the definition, classification, and staging of cardiac adenocarcinoma (CA). Recently, a shift from distal to proximal lesions has been documented in gastric cancer. We have stratified our cases of gastric cancer as CA, distal gastric cancer (DGC), and stump cancer (SC). METHODS: Between 1986 and 1998, 450 patients with gastric cancer were operated on at our institute. The resectability rate was 81.6%. Of 367 patients, 48 were CA, 298 DGC, and 21 SC. These 3 groups were compared in terms of clinicopathologic factors and survival rates. RESULTS: CA was significantly higher in male patients and showed a prevalence of the Lauren intestinal type. Regarding staging parameters, CA showed a higher rate of T3 tumors and of resection line involvement. Five-year survival rates were 23. 2% for CA, 45.0% for DGC, and 17.4% for SC. CONCLUSIONS: A possible cause of the poor outcome of CA is presentation at a more advanced stage. CA was similar to SC as far as epidemiology, pathologic factors, and survival rates. PMID- 10861607 TI - Prognostic factors after surgical resection for pancreatic carcinoma. AB - BACKGROUND AND OBJECTIVES: Surgical resection offers the only potential cure for pancreatic carcinoma. Several recent series have reported an encouraging increase in 5-year survival rate exceeding 20% and have emphasized the importance of patient selection based on reproducible prognostic factors. The impact on survival of demographic, intraoperative, and histopatologic factors are investigated in this study. METHODS: Seventy-three patients with adenocarcinoma of the pancreas, treated at the Department of Surgery of the Catholic University of Rome during 1988-1998, were retrospectively analyzed. Survival data were reviewed, and potential prognostic factors were compared statistically by univariate and multivariate analyses. RESULTS: There was no operative mortality, and the morbidity rate was 37%. Actuarial overall and disease-specific survival rates for all 73 patients were, respectively, 27% and 31% at 3 years and 13% and 21% at 5 years, with a median survival time of 16 months. T stage and nodal status significantly affected survival according to univariate analysis (P = 0.0017 and 0.04). An impact on survival, even if not of statistical significance, was shown for other pathologic or intraoperative factors. CONCLUSIONS: T and nodal stage are the strongest independent predictors of survival. Limited intraoperative transfusion, reduced operative time, and clear margins also may play a role, which requires further confirmation in a larger series. PMID- 10861608 TI - Treatment of peritoneal carcinomatosis with intent to cure. AB - BACKGROUND AND OBJECTIVES: Low-grade malignant tumors arise in the abdomen, do not infiltrate, and "redistribute" on the peritoneum with no extraregional spreading. In these cases, aggressive surgery combined with localized chemotherapy may provide cure. METHODS: After removing the tumor with the regional peritoneum en bloc, intraabdominal hyperthermic chemoperfusion was performed throughout the abdominopelvic cavity. Alternatively, early intraabdominal chemotherapy, starting on the first postoperative day, was administered for 5 days. RESULTS: Forty patients affected with extensive peritoneal carcinomatosis underwent peritonectomy, with no residual macroscopic disease except in four cases. Seventy-five percent of the patients underwent locoregional chemotherapy. Major complications were observed in 40% of the patients and led to death in five; there was a direct correlation to the duration of surgery (P = 0.03). At a mean follow-up of 20 months, the overall 2-year survival was 61.4%, with a median survival of 30 months. CONCLUSIONS: After a learning curve of 18 months, the feasibility of the integrated treatment increased to greater than 90%, and mortality dramatically decreased. The combined treatment resulted in a high survival rate in patients with extensive carcinomatosis who were no longer responsive to traditional therapies. PMID- 10861609 TI - Thrombectomy of the inferior vena cava from recurrent low-grade endometrial stromal sarcoma: case report and review of the literature. AB - We report a rare case in which a patient successfully underwent surgical removal from the inferior vena cava of a neoplastic thrombus induced by a recurrent low grade endometrial stromal sarcoma. The patient was admitted with severe acute renal failure and a large edema on the right lower extremity. One year previously she had undergone hysterectomy and adnexectomy due to an endometrial stromal sarcoma with involvement of the tuba. Because of complete thrombosis of the right internal and common iliac veins and the inferior vena cava, she underwent thrombectomy of the inferior vena cava. The postoperative course was complicated by hydruric renal failure induced by a acute tubular necrosis. At 6-month follow up, the patient was asymptomatic with normal renal function. The ileocaval axis was patent on magnetic resonance imaging. Only 5 cases of intracaval extension of endometrial stromal sarcoma have been reported. Surgical treatment is viable due to excellent prognosis of the low-grade endometrial stromal sarcoma (80-100% 5 year survival) and likely fatal heart failure in untreated cases. PMID- 10861610 TI - Detubularized rectosigmoid neobladder in women after cystectomy for bladder cancer. AB - BACKGROUND AND OBJECTIVES: We present the long-term functional results of a new technique for bladder substitution after cystectomy for bladder cancer in women. METHODS: Between 1991 and 1995, 10 women underwent radical cystectomy for bladder cancer with a new technique. We created a detubularized rectosigmoid neobladder associated with either a terminal colostomy or intrasphincteric perineal colostomy section (Heitz-Boyer-Hovelacque). We evaluated neobladder functioning over almost 5 years by means of urodynamic studies, ultrasound scans, urograms and pouchgrams, and renal function tests. RESULTS: Neobladder function was excellent in all patients, with good diurnal and nocturnal urinary continence, voiding patterns, and preservation of the upper urinary tract. CONCLUSIONS: This new technique, which is a modification of the standard rectal or rectosigmoid neobladder technique, is a valid alternative to the ortothopic neobladder in women, with good functional results. PMID- 10861611 TI - Biological prognostic factors for early stage completely resected non-small cell lung cancer. AB - BACKGROUND AND OBJECTIVES: The different and unpredictable outcomes in early stage non-small cell lung cancer patients requires urgent research concerning the biological pathway of this neoplasm. Our study investigated the frequency of expression and the clinicopathologic and prognostic significance of a series of biological markers in stage I and II resected non-small cell lung cancer. METHODS: A total of 99 cases of pathologic stage I and II were analyzed. The mean follow-up of surviving patients was 41 months. The expressions of the following biological markers were tested: bcl-2, p53, Ki-67, angiogenesis, and tumor vessel invasion. Kaplan-Meier estimates of survival and time to recurrence were calculated for clinical variables and biological markers using Cox's model for multivariate analysis. RESULTS: Tumoral vessel invasion was present in 22 (22%) pathologic samples, the angiogenesis mean value was 37 +/- 13, and median was 35; 13 (13%) patients showed positive immunostaining for bcl-2 oncoprotein. P53 oncoprotein expression was present in 48 patients (48.5%). All samples presented Ki-67 expression (mean value = 25.3 +/- 19.3, median = 20). The pathologic staging of the tumor was the most important independent prognostic factor for survival (P = 0.037) and for recurrence of disease (P = 0.040). Tumoral vessel invasion was the only marker with an independent predictive factor for survival and recurrence of disease in the group of patients without lymph node involvement (P = 0.02). CONCLUSION: Our data do not support a relevant prognostic role for p53, bcl-2, or Ki-67 immunohistochemical markers in non-small cell cancer. Tumor vessel invasion was an independent predictive factor of poor outcome in the group of patients without lymph node involvement. Pathological stage was confirmed as the most important independent prognostic factor. PMID- 10861612 TI - Sentinel lymph node mapping in early-stage breast cancer: technical issues and results with vital blue dye mapping and radioguided surgery. AB - BACKGROUND AND OBJECTIVES: Axillary lymph node status is the most important prognostic factor in patients with operable breast cancer. Recent studies have demonstrated the possibility of identifying the sentinel lymph node (sN) as a reliable predictor of axillary lymph node status in both cutaneous melanoma and breast cancer. Sentinel lymph node identification proved feasible by either peritumoral dye injection (Patent Blue-V) or radiodetection, with identification rates of 65-97% and 92-98%, respectively. However, some important issues need further definition, namely (a) optimization of the technique for intraoperative detection of the sN, (b) predictive value of the sN with regard to axillary lymph node status, and (c) reliability of intraoperative histology of the sN. We reviewed our experience in sN detection in patients with stage I-II breast cancer to assess the feasibility and accuracy of lymphatic mapping, by vital blue dye or radioguided surgery, and sN histology as a predictor of axillary lymph node status. METHODS: Two groups of patients (55 and 48) were recruited between May 1996 and May 1997 and between October 1997 and February 1998; the patients of the first series underwent vital blue dye lymphatic mapping only, whereas those of the second series had a combined approach with both vital blue dye mapping and radioguided detection of the sN. RESULTS: In the first set of patients, the sN was identified in 36/55 patients (65.4%); sN histology predicted axillary lymph node status with a 77% sensitivity (10/13), a 100% specificity (23/23), an 88.5% negative predictive value (23/26), and an overall 91.5% accuracy (33/36). The sN was the quasi-elective site of lymph node metastases because in clinically N0 patients nodal involvement was 20-fold more likely at histology in sN than in non sN (30% and 1.5%, respectively). In the second set of patients, 49 lymphadenectomies were performed because 1 patient had bilateral breast cancer; the sN was identified in 45/49 lymphadenectomies (92%). The sN was intraoperatively negative at frozen-section examination in 33 cases, and final histology confirmed the absence of metastases in 31/33 cases (94%), whereas in 2 cases (6%) micrometastases only were detected. Final histology of the sN predicted axillary lymph node status with an 87.5% sensitivity (14/16), a 100% specificity (29/29), a 93.5% negative predictive value (29/31), and an overall 95.5% accuracy (43/45). CONCLUSIONS: Sentinel lymphadenectomy can be better accomplished when both mapping techniques (vital blue dye and radioguided surgery) are used. In this group of patients, agreement of intraoperative histology of the sN with the final diagnosis was 94%, and sN histology accurately predicted axillary lymph node status in 43/45 lymphadenectomy specimens (95.5%) in which an sN was identified. PMID- 10861613 TI - Pattern of lymphatic drainage to the sentinel lymph node in breast cancer patients. AB - BACKGROUND AND OBJECTIVE: We performed a pilot study on 30 consecutive patients undergoing sentinel node (sN) biopsy by radioguided surgery and vital blue dye mapping to determine whether there is a single sN for each breast independent of tumor site or an sN specifically related to the site of the breast neoplasm. METHODS: There were 6 groups of 5 patients; each patient had a subdermal injection of radiotracer on the tumor site plus a second injection of radiotracer that was changed in every subset of patients to test whether modifying the site or the route of injection could have impaired the proper detection of the sN. RESULTS: "False" sN were detected only in patients who had a second injection of radiotracer away from the tumor site; this occurred in 2 of 5 patients (40%) in group I, in 3 of 5 patients (60%) in group II, in all patients in group III, and in 3 of 5 patients (60%) in group IV. The different route of injection (peritumoral or subdermal) always on the tumor site that was tested in groups V and VI did not impair the proper detection of the sN. CONCLUSIONS: Our findings support the hypothesis of a precise topographic correspondence between the primary tumor and its specific sN more than the existence of a first sN in the axillary basin, which indiscriminately drains all quadrants of the breast, like "a neck of a bottle." This should be considered for the proper selection of the injection site of either vital blue dye or radiopharmaceuticals. PMID- 10861614 TI - Integrated treatment in locally advanced carcinoma of the oropharynx. AB - BACKGROUND AND OBJECTIVES: Oropharyngeal carcinoma tends to be aggressive and deeply infiltrative of nearby sites, with an high incidence of lymph node metastases. The last treatment decision generally depends on the stage of the lesion and the patient's general status. Oropharyngeal tumor is generally treated by integrated treatments. METHODS: We retrospectively studied 115 patients with locally advanced oropharyngeal tumors treated in our institution with combined therapies compare the results in two different groups of patients (surgery plus radiotherapy and chemotherapy plus radiotherapy). RESULTS: The 3-year overall survival rate in patients who underwent surgery plus radiotherapy was 82% and in those who underwent chemotherapy plus radiotherapy was 49%. CONCLUSION: The results suggest that surgery followed by radiotherapy seems to be the best treatment in the case of locally advanced oropharyngeal tumor. PMID- 10861615 TI - Current role of radiotherapy in the treatment of locally advanced laryngeal carcinomas. AB - BACKGROUND AND OBJECTIVES: During the past few years, radiotherapy (RT) has been increasingly used in combination with surgery in the treatment of locally advanced laryngeal carcinomas to improve survival rates in patients with more extensive tumors. METHODS: This is a retrospective study of a large series of stage III and IV laryngeal carcinomas, and postoperative RT was indicated for some of these cases. We retrospectively reviewed the medical records of 380 patients with stage III and IV tumors, of which 163 (43%) underwent surgery only and 217 (57%) received surgery and postoperative RT. RESULTS: The survival rates of patients who underwent surgery and RT were comparable to those of patients who underwent surgery only, but the former group was composed of subjects suffering from negative prognostic factors. CONCLUSIONS: The indications for combined treatment should be correlated with the prognostic factors to increase the survival rate of patients with stage III and IV laryngeal carcinoma. PMID- 10861624 TI - Dr. Sidney Q. Cohlan (1915-1999). PMID- 10861616 TI - Multiple primary malignant neoplasm in patients with laryngeal carcinoma. AB - BACKGROUND AND OBJECTIVES: The aim of this study was to study the incidence, localization, and nature of second primary tumors arising in patients with primary laryngeal carcinoma and their correlation with the site, staging, and grade of differentiation. METHODS: Between 1979 and 1996, 877 patients underwent laryngeal surgery (850 male and 27 female; age range = 30.5-76 years). The second primary tumor was multicentric in 7 cases (14%), systemic in 16 cases (33. 3%), and "coincidental" in 25 cases (52%). In 3 cases the tumors were simultaneous, in 7 cases they were synchronous, and in 38 cases they were metachronous. In the 48 patients with multiple primary malignant neoplasm, the index tumor was glottic in 38 cases and supraglottic in the remaining 10, but the incidence of second tumors in these two groups was virtually matching, respectively: 38/709 (5. 3%) and 10/168 (5.9%). RESULTS: This study did not identify any correlation between the second primary tumor and the extension of the first neoplasm (T) and the presence of laterocervical lymph node metastases (N). Our data showed a higher incidence of second tumor in patients having G1 carcinomas compared with the other series with G2 and G3 carcinomas. As far as survival rates are concerned, a significant reduction was observed in patients developing a second primary tumor, regardless of whether the index tumor was glottic or supraglottic. CONCLUSIONS: The incidence of new tumors in patients with laryngeal carcinoma is not linked to the site, size, staging, or grade of differentiation of the index tumor. PMID- 10861625 TI - Histogenesis control genes: embryology, wound-healing, and NF1. PMID- 10861626 TI - Case-control studies using only malformed infants who were prenatally exposed to drugs. What do the results mean? PMID- 10861627 TI - Response to "case-control study using only malformed infants who were prenatally exposed to drugs. What do the results mean?" by Prieto L and Martinez-Frias ML. PMID- 10861628 TI - Pathways to transcription. PMID- 10861629 TI - Teratogen-induced activation of ERK, JNK, and p38 MAP kinases in early postimplantation murine embryos. AB - BACKGROUND: Although many teratogens are known to activate apoptotic pathways culminating in abnormal development, little is known about how the embryo transduces a teratogenic exposure into specific responses. Signal reception and transduction are regulated by a number of signal transduction pathways, including the extracellular signal-regulated protein kinases (ERKs), c-Jun N-terminal kinases (JNKs) and the stress-activated protein kinase, p38. METHODS: To analyze the effects of teratogens on MAP kinases, we used whole embryo culture, Western blot analyses, and antibodies recognizing inactive or active MAP kinases, or both. RESULTS: We show that heat shock (HS) induces a rapid, strong, but transient activation of ERK, JNK, and p38 with maximal activation occurring within 30 min of the heat shock. By contrast, cyclophosphamide (CP) and staurosporine (ST) failed to activate ERK or JNK during the time period studied (7. 5 hr). ST and CP did induce a low but reproducible activation of p38 beginning at around 3 hr and 5 hr, respectively, after the initiation of exposure. Previous work has shown that heat shock induces elevated cell death in the embryo, primarily in the developing neuroepithelium, but not in the embryonic heart. Thus, we also compared the activation of these three MAP kinase pathways in heads, hearts, and trunks isolated from day 9 embryos exposed to 43 degrees C for 15 min. The results show that ERK, JNK, and p38 are activated in heads, hearts, and trunks. CONCLUSIONS: Our results show that day 9 embryos do activate MAP kinase signaling pathways in response to teratogenic exposures; however, activation of a particular pathway does not appear to be required for teratogen induced apoptosis. PMID- 10861630 TI - Alpha-tocopherol and gamma-tocopherol attenuate ethanol-induced changes in membrane fatty acid composition in embryonic chick brains. AB - BACKGROUND: This project investigated whether or not EtOH-induced reductions in the levels of long-chain polyunsaturated membrane fatty acids could be attenuated by exogenous exposure to either alpha-tocopherol, gamma-tocopherol, or diallyl sulfide (DAS). METHODS: At 0 days of development, fertile chicken eggs were injected with a single dose of either saline supplemented with various concentrations of EtOH, alpha- or gamma-tocopherol and EtOH, or DAS and EtOH. At 18 days of development, brains were isolated and subjected to membrane analyses. RESULTS: When exposed to EtOH, concentrations ranging from 0-60.50 microm/Kg egg, dose-dependent decreases in the levels of brain 18:0, 18:1 (n-9), 18:2 (n-6), 18:3 (n-3), and 20:4 (n-6) were observed. These ethanol-induced changes in membrane fatty acid composition correlated with ethanol-induced reductions in brain mass, brain protein levels, acetylcholine esterase (AChE) activities and correlated with increased lipid hydroperoxide levels. Exposure to either 2.5 microm alpha-tocopherol/Kg egg and 6.050 mm EtOH/Kg egg, or 2.5 microm alpha tocopherol/ Kg egg and 6.050 mm EtOH/Kg egg attenuated EtOH-induced changes in membrane fatty acid composition, brain mass, brain protein levels, AChE activities, and lipid hydroperoxide levels. Embryonic exposure to the cytochrome p450-2E1 inhibitor, diallyl sulfide (DAS), also attenuated EtOH-induced decreases in long-chain, unsaturated membrane fatty acids. However, embryonic exposure to DAS promoted abnormally low brain mass. CONCLUSION: EtOH-induced reductions in the levels of brain long-chain polyunsaturated fatty acid are caused by lipid peroxidation. PMID- 10861631 TI - Risk factors for congenital hypothyroidism: an investigation of infant's birth weight, ethnicity, and gender in California, 1990-1998. AB - BACKGROUND: Approximately 85% of primary congenital hypothyroidism (CH) is sporadic and due to malformations of the thyroid gland. Past studies have reported an increased birth weight among infants with CH. We have attempted to replicate and expand these observations, examining the association between different birth weight categories and CH stratified by infant's sex. We have also examined the prevalence of CH by mother's age and infant's ethnicity, gender, and year of birth. METHODS: A cross-sectional study was conducted on 5, 049,185 infants screened by the statewide California Newborn Screening Program between 1990 and 1998, an estimated 98.6% of all newborns in the state. Dried blood spots from a heel stick were assayed for thyroxine (T4), and presumptive positives had follow-up assays of thyroid-stimulating hormone (TSH) to determine definite positives. RESULTS: A total of 1,806 cases of CH were identified. The following findings are unlikely to be due to chance. Compared with infants with birth weights of 3,000-3,499 g, infants weighing <2,000 g and those weighing >/=4,500 g had a twofold or greater increase in the prevalence of CH. This was not explained as a result of confounding by the infant's ethnicity or gender. Compared with whites, elevated prevalence rates were found in most ethnic groups, which include the following: Hispanics, Chinese, Vietnamese, Asian Indians, Filipinos, Middle Easterners, and Hawaiians. As reported previously, black infants had about one third the prevalence rate of whites. We also observed the frequently described female preponderance of CH. The female excess was maintained at all birth weights, however it varied by infant's ethnicity. Trends in the prevalence of CH were not associated with mother's age or with the time interval between 1990 and 1998. CONCLUSIONS: We observed an increased risk of CH in both low-birth-weight (<2,000-g) and macrosomic (>/=4,500-g) infants. This U-shaped association has not been described in past studies. We have also expanded the previously described ethnic differences in CH risk to include ethnic groups not previously studied. The unique pattern of CH occurrence suggests that further studies to define modifiable risk factors may be useful. PMID- 10861632 TI - Hypothesis: folate-responsive neural tube defects and neurocristopathies. AB - BACKGROUND: What accounts for the wide spectrum of folate-responsive dysmorphogeneses? Both embryonic and fetal cells are entirely dependent on maternal folate to support their requirement for precisely timed proliferative bursts during gestation. Folate receptors (FRs) mediate transport into cells and are central to transplacental maternal-to-fetal folate transport. FRs are also critical for neural tube and neural crest development because recent murine "knock-out" and "knock-down" of FRs results in a high percentage of folate responsive neural tube defects (NTDs) and neurocristopathies. HYPOTHESIS: Central to our hypothesis is the fact that folate deficiency is accompanied by a reduction in the proliferative capacity of highly mitotic neural tube or neural crest cells. Therefore, depending on when in pregnancy various cohorts of highly proliferative cells are deprived of folate, and the origin of the affected cells will determine the type of developmental dysmorphogenesis. Thus, selective folate deficiency in early pregnancy of only highly proliferative neural tube or neural crest cells predisposes to NTDs or gross dysmorphogenesis, respectively. Folate deficiency that compromises placental development will predispose to small-for date babies due to an overall nutrient deficiency, and the development of folate insufficiency later in pregnancy could predispose to more subtle midline birth defects involving atresia of neural crest cell-derived structures. Finally, a congenital folate transport defect would only be corrected by suprapharmacological doses of folate, which ensures passive diffusion. CONCLUSION: This hypothesis can explain the results of several earlier and more recent clinical trials on folate supplementation in pregnancy, but it also raises the possibility that there may be several as yet undiscovered neurocristopathies that are folate responsive. Teratology 62:42-50, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861633 TI - Appropriate use of animal models in the assessment of risk during prenatal development: an illustration using inorganic arsenic. AB - BACKGROUND: Assessing risks to human development from chemical exposure typically requires integrating findings from laboratory animal and human studies. METHODS: Using a case study approach, we present a program designed to assess the risk of the occurrence of malformations from inorganic arsenic exposure. We discuss how epidemiological data should be evaluated for quality and criteria for determining whether an association is causal. In this case study, adequate epidemiological data were not available for evaluating the potential effect of arsenic on development. Consequently, results from appropriately designed, conducted, and interpreted developmental toxicity studies, which have been shown to be predictive of human risk under numerous scenarios, were used. In our case study, the existing animal data were not designed appropriately to assess risk from environmental exposures, although such studies may be useful for hazard identification. Because the human and animal databases were deficient, a research program comprising modern guideline toxicological studies was designed and conducted. RESULTS: The results of those studies in rats, mice, and rabbits indicate that oral and inhalational exposures to inorganic arsenic do not cause structural malformations, and inhalational exposures produced no developmental effects at all. The new study results are discussed in conjunction with considerations of metabolism, toxicokinetics, and maternal toxicity. CONCLUSIONS: Based on the available experimental data, and absent contrary findings from adequately conducted epidemiological studies, we conclude that exposure to inorganic arsenic by environmentally relevant routes poses no risk of the occurrence of malformations and little risk of other prenatal developmental toxicity in developing humans without concomitant and near-lethal toxicological effects in mothers. PMID- 10861634 TI - HPV-DNA is not detectable in outgrowing cells from explant cultures of skin lesions established at the air-liquid-interface. AB - Keratinocyte cultures established from HPV containing skin cancers were described earlier to lose their HPV DNA after passaging in vitro. A different approach was therefore used in this study. Explant cultures were generated by depositing small pieces of various benign and (pre)malignant skin specimens of renal transplant recipients and non-immunosuppressed patients on fibroblast-populated collagen lattices or on de-epidermized dermis. Subsequently, the cultures were maintained at the air-liquid interface. At various time points, samples were collected for both HPV analysis, using a nested PCR approach, and morphology. The outgrowing keratinocytes developed into multilayered epithelial structures showing terminal differentiation. No histological differences were observed between cultures established from HPV positive and negative lesions. Eighteen biopsy specimens were tested for their HPV content before and after culture. Before culture 11 out of these skin specimens contained DNA of the Epidermodysplasia Verruciformis related HPV types (EV-HPV). Comparison of the HPV types detected in two different parts of the same skin specimen before culture was strongly suggestive for a non homogeneous distribution of EV-HPV in the lesions. From the explant cultures derived from the 11 HPV-positive biopsies, 31 samples from the originally explanted pieces of tissue and 38 samples from the outgrowing multilayered epithelial sections were collected. HPV DNA was detected in 10 of the 31 and in 3 of the 38 samples (Chi-square test, P = 0.01), respectively. These results indicate that EV-HPV positive keratinocytes do not efficiently proliferate or lose their HPV DNA in this culture system or EV-HPV DNA is present in only a few basal cells, making it improbable that these cells are located at the outgrowing margins. PMID- 10861635 TI - Human papillomavirus infection and non-melanoma skin cancer in immunosuppressed and immunocompetent individuals. AB - The role of human papillomavirus (HPV) in anogenital carcinogenesis is established firmly, but a similar role in non-melanoma skin cancer remains speculative. Certain immunosuppressed individuals have an increased incidence of both viral warts and non-melanoma skin cancer, that has prompted the suggestion that HPV may play a pathogenic role. Differences in the techniques used to detect HPV DNA in skin, however, have led to discrepancies in the prevalence and spectrum of HPV types reported in these malignancies. This study describes the use of a comprehensive degenerate PCR technique to compare the HPV status of 148 Non-melanoma skin cancers from immunosuppressed and immunocompetent individuals. HPV DNA was detected in 37/44 (84.1%) squamous cell carcinomas, 18/24 (75%) basal cell carcinomas and 15/17 (88.2%) premalignant skin lesions from the immunosuppressed group compared with 6/22 (27.2%) squamous cell carcinomas, 11/30 (36.7%) basal cell carcinomas and 6/11 (54. 4%) premalignancies in the immunocompetent group. Epidermodysplasia verruciformis HPV types prevailed in all lesion types from both groups of patients. In immunosuppressed individuals, cutaneous HPV types were also identified at high frequency, and co-detection of multiple HPV types within single tumours was commonly observed. This study represents the largest and most comprehensive analysis of the HPV status of non melanoma skin cancers yet undertaken; whereas there are clearly significant differences in non-melanoma skin cancers from immunosuppressed and immunocompetent populations, we provide evidence that the prevalence and spectrum of HPV types does not differ in squamous cell carcinomas, basal cell carcinomas or premalignancies within the two populations. These data have important implications for future investigation of the role of HPV in cutaneous carcinogenesis at a functional level. PMID- 10861636 TI - Analysis of relative binding affinity of E7-pRB of human papillomavirus 16 clinical variants using the yeast two-hybrid system. AB - A number of genotypes of the human papillomaviruses (HPV) are associated with malignancies of the uterine cervix. Sequencing work has revealed the existence of intratype HPV variants with minor differences in the nucleotide sequence. More recent data suggest the possibility that some of the variants may have different modes of clinical manifestation. In this study, sequences of the E6 and E7 oncogenes of 17 HPV16 isolates derived from PAP smear samples of Taiwanese patients were analyzed. A number of E6 and E7 novel variants were found. Particularly, a prevalent (64.7%) E6 polymorphic site A442C with an E113D amino acid substitution seems specific to Taiwanese patients. In E7, two novel but silent polymorphic sites G663A (41.2%) and T846C (88.2%) were also prevalent in the samples analyzed. The yeast two-hybrid system was adopted for rapid assessment of relative E7-pRb binding affinity in the variants. The relative binding affinities of the E7 proteins of different HPV types to pRB were in close agreement with previous biochemical data. A T663G/C24W polymorphic change in E7 correlated with a decrease in E7-pRb relative binding affinity the significance of which remains to be clarified. This semi-quantitative biochemical and genetic approach may be useful as a first step in the development of clinical protocols for the screening and identification of important HPV variants for clinical interpretation and for further functional analysis by transfection or other bioassays. PMID- 10861637 TI - Genetic changes in the interferon sensitivity determining region of hepatitis C virus during the natural course of chronic hepatitis C. AB - Amino acid mutations in the interferon sensitivity determining region (ISDR) are closely associated with the response to interferon in patients with hepatitis C virus genotype 1b (HCV-1b) infection. In this study, 36 patients chronically infected with HCV-1b, with no history of interferon therapy with respect to ISDR changes in HCV were studied. Two serum samples were obtained from each patient, with an average interval of 3.5 years, and predominant nucleotide and amino acid sequences of the ISDR at initial and subsequent time points were compared for each patient. Three of 12 patients with the wild type ISDR (no amino acid mutation in the ISDR compared to the consensus sequence) changed to the intermediate type (1 to 3 mutations) at later time points, whereas the other 9 still had the wild type. Similarly, 2 of 18 patients with the intermediate type changed to the wild type, whereas the other 16 patients continued to have the intermediate type. One of 6 patients with the mutant type (4 or more mutations) changed to the intermediate type, and the other 5 continued to have the mutant type. Although ISDR nucleotide changes/site/year were not significantly different among the 3 groups of patients, percentages of non-synonymous nucleotide changes were greater in the mutant type (63%) than the wild (9%) or the intermediate type (20%) (P < 0.05). These results show that mutations in the ISDR do not occur frequently, suggesting that interferon sensitivity does not change greatly during the natural course of the disease in each patient. PMID- 10861638 TI - 93G, a novel sporadic strain of hepatitis E virus in South China isolated by cell culture. AB - The nucleotide sequence of Hepatitis E virus (HEV) serous isolates (G-9 and G-20) from Guangzhou, South China, which has been reported previously, are divergent significantly from those of other reported HEV isolates. In order to investigate more extensively the Guangzhou isolate, the 93G strain was isolated from the faecal sample of the same individual as G-9 by A549 cell culture and identified immunologically and by molecular biological techniques. The results showed that strain 93G could be propagated in an A549 cell line causing cytopathic effects. The viral particles were aggregated by a specific antibody to HEV Chinese Xinjiang strain (87A) observed using immunoelectron microscopy and were similar morphologically to HEV from other sources. In this study, an indirect fluorescent antibody assay was first developed to examine HEV antigen in the infected cells, by immunofluorescence in the cytoplasm and on the surface membrane of the cells. The 58-kDa and 82-kDa native structural proteins of HEV were also identified in this study by Western blotting. The 93G genome showed high homology (93%) with G 9 previously reported but was also as divergent from the Burmese, Mexican, Chinese Xinjiang isolates and the recently reported US-1 isolate, as was G-9. The data presented indicate that 93G propagated in A549 cells, together with its related serum isolate G-9, represents another HEV strain circulating in China and is responsible for some sporadic hepatitis E infections. PMID- 10861639 TI - Molecular epidemiology of GB virus C/hepatitis G virus in Athens, Greece. AB - The relevance of GB virus C/hepatitis G virus (GBV-C/HGV) infections in liver pathology remains unclear. To investigate the epidemiology of GBV-C/HGV in Athens, Greece, sera from 512 subjects were screened for present and past markers of GBV-C/HGV infection using a reverse transcription-polymerase chain reaction (RT-PCR) and a serological assay, respectively. GBV-C/HGV RNA was detected in 18/56 (32.1%), 12/42 (28.6%), and 16/55 (29.1%) patients with acute hepatitis B, C, or non-A-E, and in 5/58 (8.6%) and 18/68 (26.5%) patients with chronic hepatitis B or C, respectively, as well as in 50/133 (37.6%) hemodialysis patients and 10/100 (10%) healthy individuals. The data indicated that GBV-C/HGV seroprevalence is age-dependent; thus, GBV-C/HGV RNA and anti-E2 positivity were shown to be associated with younger age [odds ratio 0.98, 95% confidence interval (CI) 0. 97-1.00, P = 0.017] and older age (odds ratio 1.03, 95% CI 1.01-1.05, P = 0.002), respectively. No significant associations were identified between GBV C/HGV RNA status and alanine aminotransferase (ALT) levels in either hepatitis or hemodialysis patients. Nevertheless, GBV-C/HGV RNA-positive acute non-A-E hepatitis patients were more likely to manifest a more severe clinical form of acute hepatitis (P = 0.024). Phylogenetic analysis of partial 5'-untranslated region sequences isolated from 18 viremic individuals showed that most GBV-C/HGV strains circulating in the greater metropolitan area of Athens belong to the 2a subgroup. A genetically diverse type 2 sequence that may represent a novel subtype within group 2 was also characterized. PMID- 10861640 TI - Molecular epidemiology of astroviruses in Japan from 1995 to 1998 by reverse transcription-polymerase chain reaction with serotype-specific primers (1 to 8). AB - In addition to the serotype-specific primers described previously (1 to 7), a new serotype 8-specific primer has been designed, allowing detection of all astrovirus serotypes. A total of 1,382 diarrheal stool samples in 5 regions in Japan were examined by reverse transcription-polymerase chain reaction (RT-PCR). The incidence of astrovirus infection in all 5 regions was 5.9% (82 of 1,382 samples) and infection occurred mainly from November to April. Serotypes 1, 3, and 4 were detected in 66, 14, and 2 of the 82 positive samples, respectively. None of the other serotypes was detected. The highest detection rate was from 0 to 1 year old, 39.0%, and the next highest was from 1 to 2 years old, 34.1%. The primers provide a useful approach for study of the epidemiology of astroviruses. PMID- 10861641 TI - Human herpesvirus 8-encoded interleukin-6 homologue (viral IL-6) induces endogenous human IL-6 secretion. AB - We found that human herpesvirus 8-encoded IL-6 (vIL-6) induced endogenous human IL-6 (huIL-6) secretion from various cell lines (MT-4, THP-1, U937, Raji, and CESS) including patients with multicentric Castleman's disease. Especially, in MT 4 cells, huIL-6 was enhanced with vIL-6 by 30-fold compared with that of control. In addition, reverse transcriptase-polymerase chain reaction confirmed that vIL-6 induced huIL-6 expression in MT-4 cells. Our novel finding of the huIL-6 induction by vIL-6 indicates that vIL-6 may play a significant role in the pathogenesis of HHV-8 associated diseases. PMID- 10861642 TI - No association of borna disease virus with psychiatric disorders among patients in northern Kyushu, Japan. AB - There is controversy over the prevalence of Borna disease virus (BDV) antibodies and its RNA in the peripheral blood mononuclear cells (PBMCs) of psychiatric patients, and the contribution of BDV to human psychiatric disorders. We examined 299 plasma and 229 PBMC samples. No plasma samples were positive for BDV-p40, p24 or gp18 antibodies by western blot analysis. The prevalence of BDV RNA in the psychiatric (schizophrenic) patients (1.8%) was not significantly different from that in the healthy volunteers (0.6%). The nucleotide sequences of BDV p40 and p24 were highly conserved with those of BDV He/80. Our results suggested that there is a lack of association between BDV infection and psychiatric disorders among the patients in Northern Kyushu, Japan. PMID- 10861643 TI - Differential detection of rhinoviruses and enteroviruses RNA sequences associated with classical immunofluorescence assay detection of respiratory virus antigens in nasopharyngeal swabs from infants with bronchiolitis. AB - To define the role of enteroviruses and human rhinoviruses as etiological agents in childhood bronchiolitis, clinical aspirates from 84 infants admitted to hospital with symptoms of obstructive bronchiolitis were tested by picornavirus RT-PCR assay, adenovirus PCR assay and classical immunofluorescence antigen detection of common respiratory viral agents. Respiratory syncytial viruses (A&B) were detectable in 45 of 84 (53.6%) nasopharyngeal aspirates from infants with bronchiolitis, whereas coronaviruses, influenza viruses, and parainfluenza viruses were not detectable in the same samples. Adenoviruses were detectable by PCR in 11 of 84 (13.1%) nasopharyngeal swabs. By using a picornavirus RT-PCR assay followed by a differential molecular hybridisation, rhinovirus and enterovirus RNA sequences were detected in 16 of 84 (19%) and in 10 of 84 (11.9%) of the nasopharyngeal swabs tested. Positive human rhinovirus or enterovirus RT PCR assay, however, was the only evidence of respiratory infection in 8 of 84 (9.5%) and in 7 of 84 (8.33%) of the studied patients. Respiratory syncytial viruses, human rhinoviruses, adenoviruses, and enteroviruses occur in dual infections detected in 18 of 84 (21.4%) respiratory samples tested. The median duration of stay in hospital was not significantly different between the patients demonstrating a single viral infection and those with a dual viral infection (6.22 +/- 2.07 vs. 5. 04 +/- 0.95 days; P > 0.05). In summary, combination of molecular and classical detection assays of common viruses can be used to demonstrate enterovirus and human rhinovirus respiratory infection in childhood bronchiolitis, and provides an improved approach to obtain new insights into concomitant viral respiratory tract infection in infants. PMID- 10861644 TI - TT virus infection in human immunodeficiency virus type 1 infected mothers and their infants. AB - Serum TT virus (TTV) DNA was determined in 83 human immunodeficiency virus type 1 (HIV 1) infected mothers [46 intravenous drug user and 37 non-intravenous drug user women] and their infants. Twenty-nine (34.9%) mothers were TTV infected. Infection was more frequent among intravenous drug user than non-intravenous drug user mothers [21/46 (45.6%) vs. 8/37 (21.6%); relative risk (RR): 2.1; 95% confidence limits (95% CL): 1.1-4.2; P = 0.023] and among intravenous drug users who carried on injecting than in those who had given it up [10/14 (71.4%) vs. 11/32 (34.3%); RR: 2.1 (95%CL: 1.2-3.7); P = 0. 021]. Infection was not related to age, CD4-positive T-lymphocyte counts, HIV 1 load, hepatitis B (HBV), G/GB-C (GBV-C/HGV), C (HCV) virus exposure. Eight (27.5%) infants born to TTV infected (but none of those born to TTV uninfected) mothers were TTV infected at a median age of 1.5 (range: 0.6-2.8) months. Infants born by vaginal/emergency caesarean delivery were more frequently infected than those born by elective caesarean delivery [7/16 (43.7%) vs. 1/13 (7.6%); RR: 2.1; 95%CL: 1.2-3.5; P = 0.033]. Infection in infants was not related to maternal CD4-positive T-lymphocyte counts, HIV 1 load, and HIV 1, HBV, GBV-C/HGV, or HCV transmission. No infant became TTV infected thereafter. No TTV infected child [follow-up: 31 (median; range: 6-60) months] showed signs of liver disease; five infants cleared TTV DNA after 22 (median; range: 6-60) months. TTV infection in HIV 1 infected women is prevalently related to intravenous drug user. The findings suggest that infants may acquire TTV at birth. Infection may persist without evident liver disease. PMID- 10861646 TI - Proceedings of the international symposium on treatment of chronic hepatitis B infection with lamivudine, an oral antiviral drug PMID- 10861645 TI - Prevalence of primary resistance to zidovudine and lamivudine in drug-naive human immunodeficiency virus type-1 infected patients: high proportion of reverse transcriptase codon 215 mutant in circulating lymphocytes and free virus. AB - The presence of primary zidovudine (AZT)-resistance (mutation T215Y/F) or lamivudine (3TC)-resistance (mutation M184V) was evaluated in 90 drug-naive patients infected with human immunodeficiency virus type-1 (HIV-1) between 1987 and 1997. The proportion of mutant strains in proviral samples or plasma viral samples was determined using a differential hybridization assay. Mutation T215Y/F was found in five (5.6%) patients infected between 1994 and 1997, whereas none of these patients harbored the mutation M184V. The T215Y/F mutation was present in the virus and/or provirus and persisted for at least two years. In one patient, the mutant provirus was associated with only wild-type free virus. Four of these patients were followed, and two were treated subsequently to a regimen containing AZT but with low response. The persistence of primary resistance mutations might depend on the proportion of these mutations at the time of infection, although mutant provirus might not give rise to replicating variants. PMID- 10861647 TI - Hepatitis B: an important public health issue. AB - Hepatitis B is one of the most common infectious diseases in the world. It has been estimated that 350 million people world-wide are chronic hepatitis B virus (HBV) carriers. The global prevalence of chronic HBV infection varies widely, from high (>/=8%, e.g., Africa, Asia and the Western Pacific) to intermediate (2 7% e.g., Southern and Eastern Europe) and low (<2%, e.g., Western Europe, North America and Australia). The predominant routes of transmission vary according to the endemicity of the HBV infection. In areas of high endemicity, perinatal transmission is the main route of transmission, whereas in areas of low endemicity, sexual contact amongst high-risk adults is the predominant route. Between one-third and one-quarter of people infected chronically with HBV are expected to develop progressive liver disease (including cirrhosis and primary liver cancer). Although mass vaccination programmes have begun to control the spread of HBV infection, therapeutic intervention is the only option for those with established chronic HBV-associated liver disease. Until recently, the only treatment for chronic hepatitis B was the immune modulator, interferon (IFN) alpha. IFN alpha treatment has several disadvantages; it is expensive, it must be administered by injection, there are side effects, and IFN alpha is poorly tolerated. Lamivudine, a nucleoside analogue, is the first effective, and well tolerated, oral treatment for chronic hepatitis B. In conclusion, although we are still some way from eradicating or curing chronic hepatitis B, the advent of lamivudine allows new populations to benefit from therapy and helps to address the global public health problem of hepatitis B. PMID- 10861648 TI - Profound suppression of hepatitis B virus replication with lamivudine. AB - The therapeutic goals in chronic hepatitis B are to prevent or decrease cirrhosis and hepatocellular carcinoma in patients with pre-cirrhotic or early cirrhotic disease and to stabilise patients with end-stage cirrhosis. Lamivudine is an oral nucleoside analogue that suppresses hepatitis B virus (HBV) replication, and so may achieve both these treatment objectives. The active 5'-triphosphate metabolite of lamivudine has two modes of viral suppression. First, it mimics deoxycytidine triphosphate and is incorporated into newly synthesised HBV DNA to cause chain termination. Second, it demonstrates competitive inhibition of viral DNA-dependent and RNA-dependent DNA polymerase activity (i.e., reverse transcriptase activity). Lamivudine may, therefore, act at four possible stages during HBV replication: reverse transcription of pre-genomic mRNA into nascent minus-strand DNA; formation of plus strand DNA from nascent minus-strand DNA; completion of double-stranded DNA; and formation of covalently closed circular DNA. In clinical studies, lamivudine therapy reduced serum HBV DNA and this was associated with significant improvements in liver histology and significant and sustained enhancement of the proliferative CD4-mediated response to HBeAg and hepatitis B core antigen (HBcAg), and an increased frequency of HBeAg-specific T cells. HBV DNA concentrations often returned to pre-treatment values when therapy ended prior to the loss of hepatitis B e antigen (HBeAg). Although the emergence of HBV variants with a mutation in the YMDD (tyrosine-methionine-aspartate aspartate) motif has been observed, such variants show reduced susceptibility to lamivudine due to limited replication competence, and their emergence is not a signal to cease lamivudine therapy. In conclusion, viral suppression with lamivudine offers a means of disease improvement and immunological control in chronic hepatitis B. PMID- 10861649 TI - Hepatitis B e antigen seroconversion: effects of lamivudine alone or in combination with interferon alpha. AB - Seroconversion of hepatitis B e antigen (HBeAg) is an important marker for resolution of active hepatitis B virus (HBV) infection and for a long-term positive response to treatment. Lamivudine, a nucleoside analogue, is the first effective oral treatment for chronic hepatitis B in patients with evidence of viral replication and liver disease. When appropriate patient groups are compared, treatment with lamivudine for 1 year leads to HBeAg seroconversion in a similar proportion of patients as a standard course of interferon (IFN) alpha therapy. Seroconversion increases during prolonged therapy (up to 3 years), and is sustained post-treatment in more than three-quarters of patients. Response rates are related to the pretreatment level of serum alanine aminotransferase (ALT) and reach 65% in those patients with serum ALT > 5 x upper limit of normal (ULN) after one year. For patients with pretreatment ALT > 2 x ULN, response was seen in 38% after one year, rising to 65% after 3 years. To date, combination with IFN and lamivudine has not been shown to confer additional benefit compared with lamivudine monotherapy. Lamivudine is effective and appropriate for use in a greater proportion of HBV infected patients than IFN alpha, particularly those infected at birth or in early childhood. Furthermore, because seroconversion after lamivudine is not normally associated with a severe flare of liver disease, as seen with IFN, it is more suitable for use in patients with active liver disease and cirrhosis. In conclusion, lamivudine is more suitable than IFN for a broad range of patients, including those with severe liver disease, recurrent flares, pre-core mutant HBV and those who have failed previously IFN treatment or are immunosuppressed. Lamivudine is also better tolerated than IFN. PMID- 10861650 TI - Liver disease-significant improvement with lamivudine. AB - The natural history of chronic hepatitis B virus (HBV) infection is highly variable, ranging from a benign course to one of shortened life expectancy. Liver histology represents an accurate tool for assessing progressive liver disease, and has been used in five recent Phase III clinical trials of the oral nucleoside analogue, lamivudine, 100 mg/day, in patients with chronic hepatitis B. Significant improvements in the Knodell histological activity index (HAI) score were reported with lamivudine, with greater decreases noted after 2 years of therapy, consistent with continued alanine transaminase (ALT) normalisation. Histological data showed that lamivudine therapy can resolve or lessen the progression of fibrosis, and reduce the progression to cirrhosis in patients with chronic hepatitis B. These trials also showed that lamivudine provoked significant enhancement of hepatitis B e antigen (HBeAg) seroconversion compared with placebo, and had a profound effect on serum HBV DNA, resulting in rapid suppression of viraemia. The emergence of variants with a mutation in the YMDD (tyrosine-methionine-aspartate-as-partate) motif did not cause significant worsening of the Knodell HAI score. In conclusion, lamivudine is the first oral antiviral therapy for the treatment of chronic hepatitis B. It reduces significantly the severity of liver disease and reduces progression to cirrhosis. In addition, because lamivudine is well tolerated it represents a viable therapeutic option that may improve the prognosis of patients with chronic hepatitis B. PMID- 10861651 TI - Lamivudine for hepatitis B in clinical practice. AB - Lamivudine is a potent, once-daily, oral antiviral therapy that is effective and well tolerated in most patient groups with chronic hepatitis B virus infection, including those with pre-core mutant infection. Studies to date show that lamivudine suppresses serum viral replication, causing reductions in serum hepatitis B virus (HBV) DNA and enhancing hepatitis B e antigen (HBeAg) seroconversion (loss of HBeAg plus presence of antibodies to HBeAg [anti-HBe]). Lamivudine also improves liver disease, as shown by normalisation of alanine transaminase (ALT) levels and reduced progression to cirrhosis. Lamivudine is effective in patients who are interferon (IFN) alpha naive and in those who have failed to respond to IFN alpha, and it suppresses HBV in decompensated liver disease and in liver transplantation. Variants with mutations in the YMDD (tyrosine-methionine-aspartate-aspartate) motif may emerge with prolonged lamivudine therapy, but most patients maintain clinical control. Lamivudine has a safety profile similar to that of placebo and it is better tolerated than IFN alpha. In conclusion, lamivudine represents a major advance in the therapeutic options available for the management of patients with chronic hepatitis B and should now be considered the drug of choice for most patients who require treatment. PMID- 10861652 TI - Management of hepatitis B in China. AB - A randomised, multicentre, double-blind, placebo controlled trial was conducted in Chinese patients with chronic hepatitis B to compare the efficacy of once daily lamivudine and placebo on serum HBV DNA, and to assess the long-term efficacy and safety of lamivudine. Patients received lamivudine 100 mg (n = 322) or placebo (n = 107) once daily for 12 weeks, and were then offered open-label lamivudine treatment for 2 years. Lamivudine therapy resulted in increased hepatitis B e antigen (HBeAg) loss and seroconversion (loss of HBeAg plus the development of antibodies to HBeAg) in patients with high baseline serum alanine aminotransferase (ALT) concentrations. At 2 years, loss of HBeAg was achieved by 27% (38/140), 38% (25/66) and 60% (9/15), and seroconversion was achieved by 17% (24/140), 24% (16/66) and 33% (5/15) of patients with baseline serum alanine aminotransferase (ALT) concentrations of >1 x upper limit of normal (ULN), >2 x ULN and >5 x ULN, respectively. With lamivudine treatment, serum HBV DNA decreased rapidly to very low concentrations and remained low throughout the 2 years of the study. At 1 year, 15% (43/295) of patients in the lamivudine group had developed YMDD (tyrosine-methionine-aspartate-aspartate amino acid motif of HBV polymerase) variant HBV. These patients derived clinical benefit with continued lamivudine therapy, demonstrated by serum HBV DNA and ALT concentrations below baseline, or normal serum ALT concentrations. Lamivudine was well tolerated and an effective once-daily oral therapy for Chinese patients with chronic hepatitis B with viral replication and liver disease. PMID- 10861653 TI - Response of pre-core mutant chronic hepatitis B infection to lamivudine. AB - The proportion of chronic liver disease associated with the pre-core mutant of hepatitis B virus (HBV) infection is increasing, particularly in Mediterranean Europe and in Asia. The pre-core mutant HBV is unable to produce hepatitis B e antigen (HBeAg), so that patients with this variant do not present with HBV characterised by HBeAg in the serum. Pre-core mutant chronic hepatitis B infection usually proceeds to serious liver disease. Wild-type HBV infection may be mild and respond relatively well to interferon (IFN) alpha therapy, but IFN alpha is not an effective therapeutic option in pre-core mutant hepatitis B infection and new therapeutic options are needed. Clinical data show that lamivudine is an effective treatment for patients with pre-core mutant hepatitis B. There is profound suppression of HBV replication and improvement in indicators of liver disease in most patients. In conclusion, lamivudine is suitable for treatment of a wide range of patients with chronic hepatitis B, including those with pre-core mutant HBV infection. PMID- 10861654 TI - End-stage liver disease and liver transplantation: role of lamivudine therapy in patients with chronic hepatitis B. AB - Chronic infection with hepatitis B virus (HBV) is a common cause of advanced liver disease, including end-stage liver disease. Liver transplantation is generally regarded as the treatment of choice for decompensated cirrhosis. Although long-term prophylaxis with hepatitis B immune globulin (HBIg) has improved significantly the outcome after transplantation, about 20-36% of transplant recipients still develop recurrent hepatitis B and have reduced survival. Moreover, HBIg prophylaxis has a number of disadvantages, including high cost, difficulty in administration and tolerability problems. Lamivudine, an oral nucleoside analogue, is a potent inhibitor of HBV replication and has been investigated in end-stage liver disease and liver transplantation. Treatment with lamivudine results in suppression of viral replication, and clinical improvement and stabilisation of some patients with end-stage liver disease, leading to increased pre-transplant survival as well as a reduced need for transplantation. Prophylaxis with lamivudine is also effective in preventing recurrent HBV infection and graft reinfection after transplantation, although a combination of lamivudine plus HBIg is preferable to prevent the emergence of YMDD variant HBV (tyrosine-methionine-aspartate-aspartate amino acid motif of HBV polymerase). Lamivudine is also effective for the treatment of recurrent hepatitis B after transplantation, based on improvement in virological parameters of infection as well as clinical and histological manifestations of disease. In all of these clinical settings, lamivudine is well tolerated and dose reduction is not required. In conclusion, lamivudine has a potential role for the treatment of patients with hepatitis B-related end-stage liver disease, for prophylaxis in patients undergoing liver transplantation, and in the treatment of recurrent or de novo HBV infection after transplantation. PMID- 10861656 TI - Folic acid: abortifacient or pseudoabortifacient? PMID- 10861657 TI - Heterogeneity in Paget disease of the bone. AB - Paget disease of the bone is a common skeletal disorder. Recently, a gene for Paget disease was localized to 18q with subsequent evidence for linkage heterogeneity. We report the identification and clinical characterization of a large pedigree of Paget disease and demonstrate that the Paget disease gene in this pedigree is not linked to the region on 18q, thus confirming linkage heterogeneity. PMID- 10861658 TI - De novo trisomy 16p11.2-qter: report of an infant. AB - We report on a four-month-old girl with a de novo trisomy 16q [47,XX, +del(16)(p11.2).ish del(16)(p11.2)(wcp16+,D16Z2+,tel16q+, tel16p-)]. She had minor facial anomalies, limb anomalies, urogenital abnormalities, and severe cardiovascular defects. Autopsy confirmed left hypoplastic lung, total anomalous pulmonary venous drainage via coronary sinus, persistent left superior vena cava, patent ductus arteriosus, secundum atrial septal defect, bilateral hydronephrosis and hydroureters, uterus bicornis, and ovarian hypoplasia. Short tandem repeat polymorphism analysis indicated that the additional, structurally abnormal chromosome 16 was maternal in origin. PMID- 10861659 TI - Tertiary trisomy due to a reciprocal translocation of chromosomes 5 and 21 in a four-generation family. AB - Tertiary trisomy, or double trisomy, is a rare occurrence. We present two individuals with a previously unreported tertiary trisomy for chromosomes 5p and 21q in an eight-generation pedigree. Their phenotypes are compared with other partial trisomies of either 5p or 21q from the literature. The propositus was diagnosed with trisomy 21 at 2 years of age after a karyotype study for short stature and developmental delay. His phenotype was described as atypical for Down syndrome. He presented at 9 years of age because of pervasive behavioral problems and obesity. He was brachycephalic with a flattened nasal bridge, but he lacked other characteristics of trisomy 21. Because of lack of phenotypic evidence of Down syndrome, a repeat karyotype was obtained and showed 47,XY, +der(21)t(5;21)(p15.1; q22.1), incorporating partial trisomies of both chromosomes 5 and 21. Mother had a balanced translocation, 46, XX,t(5;21)(p15.1; q22.1); 8 other relatives were examined. The translocation originated from the maternal great-grandmother, but only the propositus and his mentally retarded aunt had a similar phenotye and the derivative chromosome. Fluorescence in situ hybridization showed absence of band 21q22.2 in the derivative chromosome of the propositus and his aunt, indicating that neither had trisomy for the Down syndrome critical region. These cases represent a unique double partial trisomy of chromosome arms 5p and 21q that occurred because of 3:1 malsegregation of a reciprocal translocation. These cases further demonstrate that phenotypic discordance with cytogenetic results dictate further investigation using advanced cytogenetic hybridization. PMID- 10861660 TI - Mosaic telomeric (2;14) association in a child with motor delay. AB - In a 6-year-old girl referred because of mild motor delay and hyperextensible joints, chromosome analysis disclosed a derivative chromosome consisting of end to-end fusion of chromosomes 2 and 14. Two cell lines existed in which this telomere association was present, one with a 45,XX,tas(2;14)(q37;p11) karyotype and one with a 45,XX,tas(2;14) (q37;q32) karyotype. The cell line with the telomeric fusion of 2q and 14p was present in 90% of the cells; a telomeric fusion of 2q and 14q was seen in the remaining 10% of the cells. In both association complexes, only the centromere of chromosome 14 was active. Fluorescence in situ hybridization with telomere and subtelomere probes disclosed no deletion of chromosomal material. Microsatellite analysis showed that the patient had a normal biparental contribution of chromosomes 14. PMID- 10861661 TI - Paternal UPD15: further genetic and clinical studies in four Angelman syndrome patients. AB - Among 25 patients diagnosed with Angelman syndrome, we detected 21 with deletion and 4 with paternal uniparental disomy (UPD), 2 isodisomies originating by postzygotic error, and 1 MII nondisjunction event. The diagnosis was obtained by molecular techniques, including methylation pattern analysis of exon 1 of SNRPN and microsatellite analysis of loci within and outside the 15q11-q13 region. Most manifestations present in deletion patients are those previously reported. Comparing the clinical data from our and published UPD patients with those with deletions we observed the following: the age of diagnosis is higher in UPD group (average 7 3/12 years), microcephaly is more frequent among deletion patients, UPD children start walking earlier (average age 2 9/12 years), whereas in deletion patients the average is 4 (1/2) years, epilepsy started later in UPD patients (average 5 10/12 years) than in deletion patients (average 1 11/12 years), weight above the 75th centile is reported mainly in UPD patients, complete absence of speech is more common in the deleted (88.9%) than in the UPD patients because half of the children are able to say few words. Thus, besides the abnormalities already described, the UPD patients have somewhat better verbal development, a weight above the 75th centile, and OFC in the upper normal range. PMID- 10861662 TI - Delayed membranous ossification of the cranium associated with familial translocation (2;3)(p15;q12). AB - The relationship of delayed membranous cranial ossification to cranium bifidum and parietal foramina syndromes is unclear. We report on a family with delayed cranial membranous ossification (OMIM 155980) that segregates with an apparently balanced reciprocal translocation between chromosomes 2 and 3. The propositus had apparently low-set ears, proptosis, and a soft skull at birth. A radiographic survey of the skeleton showed markedly decreased ossification of the cranial bones and no other skeletal abnormalities. The mother and maternal grandmother of the propositus have brachycephaly, hypertelorism, and a history of a soft skull at birth. Chromosome analysis of peripheral blood from the propositus showed 46,XY,t(2;3)(p15;q12). The propositus, mother, and grandmother carry the same reciprocal translocation, whereas the mother's two phenotypically normal sibs have a normal karyotype. We used an STS-linked BAC resource to define the translocation breakpoint by identifying flanking BAC clones from both chromosomes 2, 1006D24 (D2S2279) and 1060A5 (D2S2231), and chromosome 3, 3D17 (WI8558) and 3D18 [CITB Human BAC Library, J.R.K.]. This represents the second report of a family with delayed membranous ossification of the cranium and the first report of the phenotype segregating with a chromosome rearrangement. PMID- 10861663 TI - Carrier testing in fragile X syndrome: effect on self-concept. AB - The purpose of the study was to explore self-concept in women at risk for inheriting the fragile X mutation. Time 1 measures were obtained prior to carrier testing and Time 2 measures were collected approximately 5 months after learning carrier status. The sample consisted of 42 women from 17 families. Measures included the Tennessee Self-Concept Scale (TSCS), the fragile X Visual Analog Scale (VAS), and a structured interview. The TSCS provided a global measure of self-concept and the fragile X VAS and structured interview provided a contextual measure of self related to carrier status. Results indicated that there were no differences initially between carriers and noncarriers and no change from Time 1 to Time 2 on the TSCS. Analysis of the Time 1 fragile X VAS means for the total sample found a reduction in positive feelings about self. Analysis of the Time 2 fragile X VAS found that noncarriers reported improvement in feelings about self, with no change in feelings about self found in the carriers. Responses from the structured interview indicated that the feelings regarding self in the context of genetic testing are not related to global self-concept, but result from concerns regarding the implications of a positive carrier test for themselves and their families. This information highlights areas related to carrier testing that warrant further investigation and may ultimately result in modifications to the genetic counseling. PMID- 10861664 TI - Genetic disorders among Palestinian Arabs: 3. Autosomal recessive disorders in a single village. AB - Autosomal recessive diseases are common in the Arab population of Israel, mostly as a result of the high rate of consanguinity. They represent a major factor in the mortality and morbidity of the population. Since the distribution of genetic disorders in this population is not uniform, the present study was performed to determine the frequency and impact of recessive disorders within a single village. We demonstrate the existence of at least 19 autosomal recessive disorders in a village of about 8,600 inhabitants chosen at random. Since most of the disorders were chronic, the prevalence of recessive conditions in the village at the time of the study was at least 1/70, leading to a very high burden to the population and the health services. PMID- 10861665 TI - Absence of mutations in the homeodomain of the MSX1 gene in patients with hypodontia. AB - Hypodontia, the congenital absence of one or a few permanent teeth, is one of the most frequent alterations of the human dentition. Although hypodontia does not represent a public health problem, it may cause both speech and masticatory dysfunction and esthetic problems. A missense mutation in the homeodomain of MSX1 gene has been associated with hypodontia of second premolars and third molars in humans. However, another study excluded this gene as causative locus for hypodontia of incisors and premolars. To further investigate the role of the MSX1 gene in human hypodontia, we analyzed the homeobox region of the MSX1 gene in 20 individuals with different patterns of familial or isolated hypodontia. The direct sequencing of PCR products did not show any polymorphisms or mutations in the human MSX1 gene. Our results indicate that inactivation of MSX1 gene in humans must have a highly selective effect on dentition, and other genes must be involved in the cause of hypodontia in humans. PMID- 10861666 TI - Ocular manifestations in Proteus syndrome. AB - We report on the ocular manifestations of a Proteus syndrome patient. Several of the manifestations are due to severe maldevelopment and malfunction of the neuroretina including strabismus, nystagmus, high myopia, and retinal pigmentary abnormalities. In reviewing the literature, strabismus and epibulbar tumors were recorded most commonly. Some articles about presumed Proteus syndrome are spurious; these have not been included here. Also, because of anecdotal and nonsystematic study of the eye and because of the ascertainment bias inherent in literature reports, numbers of cases of each ocular manifestation have not been tabulated. PMID- 10861667 TI - Congenital insensitivity to pain with anhidrosis (CIPA) in Israeli-Bedouins: genetic heterogeneity, novel mutations in the TRKA/NGF receptor gene, clinical findings, and results of nerve conduction studies. AB - Congenital insensitivity to pain with anhidrosis (CIPA), a rare and severe disorder, comprises absence of sensation to noxious stimuli, inability to sweat, and recurrent episodes of hyperthermia. It has a relatively high prevalence in the consanguineous Israeli-Bedouins. Clinical studies of 28 patients are reported here. Using the linkage analysis approach, we linked the disease in 9 of 10 unrelated Israeli-Bedouin families with CIPA to the TrkA gene, which encodes the receptor for nerve growth factor. In one family, linkage was excluded, implying that another gene, yet unidentified, is involved. Two new mutations in the tyrosine kinase domain of the TrkA gene were identified in our CIPA patients: a 1926-ins-T in most of the southern Israeli-Negev CIPA patients, and a Pro- 689 Leu mutation in a different isolate of Bedouins in northern Israel. Eight prenatal diagnoses were made in the southern Israeli-Negev Bedouins, two by linkage analysis and six by checking directly for the 1926-ins-T mutation. Three polymorphisms in the TrkA protein kinase encoding domain were also observed. PMID- 10861668 TI - Opitz trigonocephaly (C)-like syndrome, or Bohring-Opitz syndrome: another example. PMID- 10861669 TI - Severe end of Opitz trigonocephaly C syndrome. PMID- 10861670 TI - Bohring syndrome. PMID- 10861671 TI - Growth curves for height and head circumference. PMID- 10861672 TI - No evidence supporting MTHFR as a risk factor in the development of familial NSCLP. PMID- 10861673 TI - Heterogeneity and minor anomalies. PMID- 10861674 TI - Yellow nail syndrome presenting as non-immune hydrops: second case report. AB - The yellow nail syndrome is characterized by slowly growing yellow discolored nails and lymphoedema, with onset generally after puberty. We report on a newborn infant who, at 23 weeks, was found to have hydrops on antenatal ultrasonography and bilateral chylothorax at delivery. His mother has the yellow nail syndrome, with typical nail changes, and bronchiectasis. There seemed to be no other etiology for the non-immune hydrops, and this is the second documented case of the prenatal manifestation of this condition. PMID- 10861675 TI - New form of idiopathic osteolysis: nodulosis, arthropathy and osteolysis (NAO) syndrome. AB - We describe 10 patients (6 females and 4 males) from 6 unrelated families with an autosomal recessive disease characterized by simultaneous presentation of nodulosis, arthropathy and osteolysis. They were followed up regularly at King Faisal Specialist Hospital and Research Center in Saudi Arabia for clinical evaluation, serial blood work-up, and evaluating radiological changes. Nodulosis and arthropathy were the clinical criteria for inclusion in this study, and the ten patients fulfilled these criteria. All patients had nodulosis and distal arthropathy. Eight patients (80%) presented with deformed hands and four (40%) with painful hands. All patients had parents who were first cousins and three families had more than one affected child, the finding suggesting autosomal recessive inheritance. Osteopenia and undertubulation of bones distally more than proximally, and upper limbs affected more often than lower limbs, were found in all patients. Osteolysis was seen in carpal and tarsal bones. Other common findings were sclerotic cranial sutures, brachycephaly, and broad medial clavicles. This novel phenotype should be considered in the differential diagnosis of chronic arthritis. Familial arthropathies are more often seen in communities where interfamilial marriage is common. Such a collection of patients is ideal for homozygosity mapping of the disease locus. PMID- 10861676 TI - Inherited multicentric osteolysis with arthritis: a variant resembling Torg syndrome in a Saudi family. AB - The autosomal recessive multicentric osteolytic disorders of childhood-Torg, Winchester, and Francois syndromes-predominantly affect the carpal, tarsal, and interphalangeal joints, and their progressive bone loss and crippling arthritic deformities mimic severe juvenile rheumatoid arthritis. In a consanguineous Saudi Arabian family two affected sibs with facial anomalies and short stature displayed a distal arthropathy of the metacarpal, metatarsal, and interphalangeal joints starting in the first few months of life that eventually progressed to the proximal joints and resulted in crippling ankylosis and severe generalized osteopenia. Facial changes included proptosis, a narrow nasal bridge, bulbous nose, and micrognathia. In addition, they had large, painful fibrocollagenous palmar and plantar pads and mild body hirsutism. Affected individuals were of normal intelligence and had normal renal function. Routine hematologic, chemistry, and rheumatoid studies were within normal limits. Histologic examination of bone marrow and an interphalangeal joint biopsy were not informative. The autosomal recessive inheritance, clinical, and radiologic characteristics of the affected sibs suggested that they had a form of multicentric osteolysis most closely resembling the Torg syndrome, but with a unique facial appearance, fibrocollagenous pads, and body hirsutism not noted in the original description of the syndrome. PMID- 10861677 TI - Distal limb deficiencies, oral involvement, and renal defect: report of a third patient and confirmation of a distinct entity. AB - In 1987 Buttiens and Fryns [1987: Am J Med Genet 27:651-660] reported on two sibs, brother and sister, with severe distal limb defects, micrognathia, and mild to moderate mental retardation. The male also showed severe myopia and oligomeganephronia. To the best of our knowledge, no other similar patients have been described since. We report on a boy with a similar phenotype. . PMID- 10861678 TI - Clinical findings in a patient with FGFR1 P252R mutation and comparison with the literature. AB - We report on a patient with the skeletal findings of Jackson-Weiss syndrome, who manifests only mild craniofacial anomalies. Molecular analysis of her fibroblast growth factor receptor 1 gene (FGFR1) identified a heterozygous P252R missense mutation, previously only reported with FGFR1-Pfeiffer syndrome like manifestations. Mutations in the immunoglobulin-like, II-III (IgII-III) linker region of FGFR1 and FGFR3 molecules may present as a skeletal dysplasia affecting the appendicular skeleton including, brachydactyly, short broad middle phalanges, phalangeal epiphyseal coning and broad halluces. This communication is a further example of the phenomenon of an activated FGFR molecule resulting in overlapping manifestations in FGFR syndromes. PMID- 10861679 TI - Genetic testing, adverse selection, and the demand for life insurance. AB - The dramatic increase in genetic testing for adult-onset diseases has created a debate regarding whether or not insurance companies should be able to use genetic test results in underwriting. We use data from women who have been tested for the BRCA1 gene mutation along with data from otherwise comparable untested women to assess the potential for adverse selection in the life insurance market when tested individuals know their genetic test results but insurers do not. Our analyses show that women who test positive for the BRCA1 gene mutation do not capitalize on their informational advantage by purchasing more life insurance than those women who have not undergone genetic testing. PMID- 10861680 TI - Multicolor fluorescence in situ hybridization analysis of meiotic chromosome segregation in a 47,XYY male and a review of the literature. AB - The frequencies of aneuploid and diploid sperm were determined in a 47,XYY male using multi-color fluorescence in situ hybridization (FISH) analysis, and compared with those from 10 control donors. A total of 30,078 sperm from the patient was scored, 15,044 by two-color FISH for chromosomes 13 and 21, and 15,034 by three-color FISH for the sex chromosomes using chromosome 1 as an internal autosomal control for diploidy and lack of hybridization. The frequencies of X-bearing (49.73%) and Y-bearing sperm (49.46%) in control males were not significantly different from the expected 50% (chi(2)-test for goodness of fit). The ratio of 24,X (50.60%) to 24, Y sperm (48.35%) in the patient, however, was significantly different from the controls (P = 0.0144, chi(2)-test for independence) and from the expected 1:1 ratio (P = 0.0055, chi(2)-test for goodness of fit). There was no significant increase in the frequency of diploid sperm when compared with the controls (chi(2)-test for independence). Significantly increased frequencies were found for 24,YY (0.07% vs. 0.02%, P = 0.0009) and 24,XY (0.44% vs. 0.29%, P = 0.0025), but not for 24,XX (0.05% vs. 0.05%, P > 0. 05), 24,+13 (0.07% vs. 0.07%, P > 0.05) or 24,+21 sperm (0.21% vs. 0. 18%, P > 0.05) in the 47,XYY male when compared with control donors (chi(2) test for independence). Our results support the theory that loss of the extra Y chromosome occurs during spermatogenesis in most cells. In this XYY patient there was a significant increase in the frequency of sperm with sex chromosomal abnormalities but no suggestion of an inter-chromosomal effect on autosomes. All 3-color FISH studies in the literature demonstrate a significantly increased risk of gonosomal aneuploidy in XYY males, with the risk being on the order of 1%. PMID- 10861681 TI - Syndrome of short stature, mental deficiency, microcephaly, ectodermal dysplasia, and multiple skeletal anomalies. AB - We report on two brothers with mental deficiency, short stature of prenatal onset, microcephaly, alopecia/sparse hair, follicular ichthyosis, multiple skeletal anomalies, and recurrent respiratory infections. The younger brother has celiac disease, cryptorchidism, inguinal herniae, and hypohidrosis, while the older brother has hidrotic ectodermal dysplasia, juvenile autoimmune thyroiditis, hypolacrimation, photophobia, and optic atrophy. Striking resemblance exists between our patients and those previously reported by Schinzel ?1980: Helv Paediatr Acta 35:243-251 and van Gelderen ?1982: Am J Med Genet 13:383-387. The fact that boys are born to young and healthy nonconsanguineous parents and there are no other affected relatives suggests autosomal or X-linked recessive inheritance or parental germinal mosaicism for a dominant mutation. PMID- 10861682 TI - Severe phenotypes associated with inactive ring X chromosomes. AB - Mental retardation and congenital malformations in individuals with small ring X chromosomes are often due to the functional disomy that results from failure of these chromosomes to undergo X inactivation. Such chromosomes either lack the XIST locus or do not express it. We have carried out genetic analysis of the ring X chromosomes from two girls with a 45,X/46,X,r(X) karyotype, mental retardation, and a constellation of abnormalities characteristic of the severe phenotype due to X disomy. In each case the ring X chromosome included an intact XIST locus which was expressed; the breakpoints were distal to DXS128, and therefore outside the XIC region; transcription analysis of alleles at the androgen receptor locus confirmed that these were inactive chromosomes. The characteristics of the XIST RNA were similar to the wild-type. Additional studies in cultured fibroblasts showed a second ring in a small percentage of the cells. The association of severe phenotype with an inactive X chromosome most likely reflects the presence of a second ring X chromosome which was active at least in some tissues during embryogenesis, but is no longer prominent in the tissues we analyzed. PMID- 10861683 TI - Are hereditary hemochromatosis mutations involved in Alzheimer disease? AB - Mutations in the class I-like major histocompatibility complex gene called HFE are associated with hereditary hemochromatosis (HHC), a disorder of excessive iron uptake. We screened DNA samples from patients with familial Alzheimer disease (FAD) (n = 26), adults with Down syndrome (DS) (n = 50), and older (n = 41) and younger (n = 52) healthy normal individuals, for two HHC point mutations C282Y and H63D. Because the apolipoprotein E (ApoE) E4 allele is a risk factor for AD and possibly also for dementia of the AD type in DS, DNA samples were also ApoE genotyped. Chi-squared analyses were interpreted at the 0.05 level of significance without Bonferroni corrections. In the pooled healthy normal individuals, C282Y was negatively associated with ApoE E4, an effect also apparent in individuals with DS but not with FAD. Relative to older normals, ApoE E4 was overrepresented in both males and females with FAD, consistent with ApoE E4 being a risk factor for AD; HFE mutations were overrepresented in males and underrepresented in females with FAD. Strong gender effects on the distribution of HFE mutations were apparent in comparisons among ApoE E4 negative individuals in the FAD and healthy normal groups (P < 0.002). Our findings are consistent with the proposition that among ApoE E4 negative individuals HFE mutations are predisposing to FAD in males but are somewhat protective in females. Further, ApoE E4 effects in our FAD group are strongest in females lacking HFE mutations. Relative to younger normals there was a tendency for ApoE E4 and H63D to be overrepresented in males and underrepresented in females with DS. The possibility that HFE mutations are important new genetic risk factors for AD should be pursued further. PMID- 10861684 TI - Severe acro-renal-uterine-mandibular syndrome. AB - Although limb and renal defects occur together in a variety of patterns of multiple malformations, familial cases of acro-renal disorders are rare. In 1980, Halal et al. ?Am J Med Genet 5:277-284 described two sisters with unusual limb deficiencies, renal anomalies, and mandibular hypoplasia and termed this condition acro-renal-mandibular syndrome. A girl reported earlier by Fitch and Lachance ?1972; Can Med Assoc J 107:653-656 had similarly limb and renal findings, but an apparently normal jaw. We document three sibs with unusual limb deficiencies, renal agenesis, uterine anomalies in the two females, and orofacial defects, who clearly have a similar but more severe type of acrorenal disorder, apparently inherited as an autosomal recessive condition. The sibs with limb deficiencies and renal agenesis reported by Hennekam et al. ?1994; Am J Med Genet 53:102-107 appear to be additional cases of this very rare disorder, the pathogenesis of which may be related to abnormal epithelial-mesenchymal interactions. PMID- 10861685 TI - Tetracuspid aortic valve in a patient with 22q11.2 microdeletion. PMID- 10861686 TI - Trisomy of 5p and marker chromosomes. PMID- 10861687 TI - Hemochromatosis gene nomenclature. PMID- 10861688 TI - Flow cytometric measurement of telomere length. AB - The regulation of telomere length may be involved in the cellular physiology of senescence, reproduction, cancer, immune response to infection, and possibly immune deficiency. The measurement of telomere length, critical to research in this area, has traditionally been performed by Southern blot analysis, which is cumbersome and time consuming. Several alternative methods have been described in recent years. Some, such as pulsed-field electrophoresis, slot blots, and centromere-to-telomere ratio measurements are essentially improvements to the Southern blot technique. However, other methods such as fluorescent in situ hybridization on metaphase chromosome spreads and flow cytometry-based fluorescent in situ hybridization represent a completely new technical approach to the problem. In this review, we compare methods, with particular emphasis placed on flow cytometric techniques for measuring telomere length in situ and identifying potential areas where improvements may still be made. PMID- 10861689 TI - Use of specimen mammography-guided FNA (fine-needle aspirates) for flow cytometric multiple marker analysis and immunophenotyping in breast cancer. AB - A pilot study of a novel translational research method to simultaneously assay multiple molecular markers and DNA in fine-needle aspirates (FNA) of mammographically detected breast lesions is described. Specimen mammography guided 20-gauge FNAs obtained from 86 lesions and 22 areas of normal tissue were analyzed by multiparameter flow cytometry for DNA content, her2/neu, transforming growth factor alpha (TGF alpha), and the epithelial marker cytokeratin (CK) simultaneously. Epithelial cell her2/neu positivity was detected in 12 of 44 (27%) of invasive ductal carcinomas and 3 of 9 (33%) ductal carcinoma in situ (DCIS), 10 of 30 (33%) benign lesions, and 4 of 22 (18%) normal tissue aspirates. All lesions and normal tissue showed a similar positive rate for TGFalpha ranging from 61 to 76%. The CK(+)TGF alpha(-)her2/neu(+) immunophenotype was more frequently positive in aneuploid tumors (22%) than all other lesions (7%) (P < 0.05). Specimen mammography-guided FNAs provide fresh cells for flow cytometric multiple marker analysis and immunophenotyping of clinically occult breast lesions and normal tissue. PMID- 10861690 TI - Multisite comparison of methods for the quantitation of the surface expression of CD38 on CD8(+) T lymphocytes. The ACTG Advanced Flow Cytometry Focus Group. AB - We evaluated the effect of specimen processing variations and quantitation methods on quantitative determination of CD38 expression on CD8 T lymphocytes. Neither lysing reagent (ammonium chloride versus BD FACSlyse), fixation (paraformaldehyde versus no final fixation step), nor acquisition delay (acquisition within 6 h after fixation versus 24 h after fixation) had a significant effect on CD38 relative fluorescent intensity or CD38 quantitative estimates (RFI or antibodies bound per cell). The only significant difference in fluorescent intensity and CD38 antibodies bound per cell (ABC) was encountered when whole blood was held for 24 h prior to staining and fixation and then acquired after another 24-h hold. However, for all sample processing methods above, the CD4 biologic calibrator and QuantiBRITE bead methods gave significantly different estimates of CD38 intensity. In many cases, however, these differences are relatively small and were more pronounced in certain laboratories. We conclude that there is some flexibility in sample processing methods for quantitative CD38 determination; however, it is preferable for a laboratory to employ one method of fluorescence quantitation calculation consistently because small differences are detected between different methods. Cytometry (Comm. Clin. Cytometry) 42:174-179, 2000. PMID- 10861692 TI - Expansions of clonal and oligoclonal T cells in B-cell chronic lymphocytic leukemia are primarily restricted to the CD3(+)CD8(+) T-cell population. AB - B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of mature-appearing clonal B cells exhibiting coexpression of CD5 and CD23. In addition to the accumulation of neoplastic B cells, numerous T-cell abnormalities also occur in B-CLL patients. In this study, the presence, and distribution within the T-cell subsets, of clonal/oligoclonal T cells was studied. Multicolor flow cytometric techniques were employed using combinations of anti-CD3, anti CD4, and anti-CD8 antibodies coupled with antibodies specific for V(alpha) and V(beta) T-cell receptor (TCR) epitopes. Molecular studies of TCR gene sequences were done to confirm the presence of clonal/oligoclonal T-cell populations. In the flow cytometric studies, examination of V(alpha)/V(beta)expression found evidence of clonal/oligoclonal expansion in 9 of 19 patients studied. In eight of the nine patients, the expansions were restricted to the CD3(+)CD8(+) cell population. Molecular analyses were performed in 16 patients, 12 of whom showed a clonal or oligoclonal pattern. Of the four patients who were negative in the molecular analyses, all demonstrated flow cytometric evidence of clonal/oligoclonal expansions. Thus, when the flow cytometric and molecular analyses were considered together, all 16 patients for whom parallel analyses were done showed evidence of clonal/oligoclonal expansions. These results confirm previous work demonstrating that the majority of B-CLL patients harbor clonal/oligoclonal expansions within the T-cell population. Additionally, based on the relative numbers of cells expressing specific V(alpha) or V(beta)epitopes, these results show that these expansions occur primarily within the CD3(+)CD8(+) T-cell population. PMID- 10861691 TI - Circulating CD4+ T lymphocytes with intracellular but no surface CD3 antigen in five of seven patients consecutively diagnosed with angioimmunoblastic T-cell lymphoma. AB - Angioimmunoblastic T-cell lymphoma (AITL) accounts for less than 1% of all lymphatic malignancies. Oligoclonality or monoclonality for any of the T-cell receptor (TCR) chain genes can be demonstrated in the majority of the cases. During systematic screening for the presence of circulating lymphocytes with atypical coexpression of differentiation antigens in patients with T-cell lymphomas, we have discovered a minor population (accounting for 0.2% to 10.% of all lymphocytes) of atypical lymphocytes in the blood of five of seven patients consecutively diagnosed in 1997/1998 by lymph node histology to have AITL. The major distinguishing feature of these cells consists of the lack of the surface expression of the CD3 antigen, but not of the intracellular expression. These cells express the T-cell antigens CD2 and CD5 on their surface, but not CD7, and they express CD4 and CD45 at numbers of molecules per cell typical for T lymphocytes. Gene scan analyses for the TCR gamma chain revealed oligoclonality of these flow-sorted cells in one patient and monoclonality in two patients, the same patterns of TCR gamma chain gene as determined processing the respective diagnostic lymph nodes. Circulating CD4-expressing T lymphocytes with exclusively cytoplasmic expression of CD3 appear to represent the malignant population in patients with histologically diagnosed AITL. PMID- 10861693 TI - Reliability of DNA cytometric S-phase analysis in surgical biopsies: assessment of systematic and sampling errors and comparison between results obtained by image and flow cytometry. AB - Three major parameters in DNA histograms that contribute to the reliability of S phase analysis were evaluated. These parameters are (1) the extent of background in relation to the amount of S-phase cells (and the validity of its subtraction), (2) the size of the "free" S-phase range (S(free)), and (3) the sampling error of cell counting. Tests in histograms obtained from surgical biopsies by flow cytometry (FCM) showed that the background subtraction is reliable if the found S phase fraction is higher than the fraction of background events in the histogram range of the cell population. The size of S(free) was determined in computer generated test histograms as a function of variables such as the coefficient of variation (CV) and the DNA index (DI). To calculate the sampling error of cell counting above background and in S(free), a model was developed that was validated by experimental data. This error can serve as an indicator of the uncertainty in S-phase analysis. The poor correlation found between %S values measured by image cytometry (ICM) and FCM in surgical biopsies was assigned to high uncertainty by low cell numbers in ICM histograms. A method is proposed to estimate quantitatively the reliability of S-phase analysis that can facilitate the interpretation of results. PMID- 10861694 TI - Contribution of automated hematology analysis to the detection of apoptosis in peripheral blood lymphocytes. AB - Automated hematology analyzers (analyzers) can provide complete blood counts and white blood cell (WBC) differentials in clinical laboratories and alert users to the presence of quantitative and qualitative cell abnormalities through cautionary flags. In this study, we applied analyzers to the screening of apoptotic cells in peripheral blood and examined the triggering capacity of cautionary flags to detect apoptotic cell populations. EDTA-anticoagulated fresh peripheral blood from patients with acute infectious mononucleosis containing atypical lymphocytes comprising 12.3 +/- 4. 0% of WBC was applied to a Beckman Coulter MAXM A/L Retic (MAXM) analyzer. The lymphocyte cluster spread upward in VOLUME/DF1 scattergrams and the threshold lines between lymphocyte and monocyte clusters shifted upward. Flags for the number and percentage of lymphocytes, variant lymphocytes, and blast cells were generally present for samples containing atypical lymphocytes. After the blood from acute infectious mononucleosis patients was incubated for 4 h at 37 degrees C, peripheral blood smears revealed the presence of morphologically apoptotic cells comprising 9.0 +/ 4.2% of WBC and a comparable reduction of lymphocytes. On the MAXM analyzer, the apoptotic lymphocyte cluster appeared under the lymphocyte cluster in VOLUME/DF1 scattergrams. However, no specific flag was present to alert users to the presence of the apoptotic lymphocyte cluster. We conclude that visual inspection of scattergrams generated by the MAXM analyzer can be useful for the detection of apoptotic lymphocytes in peripheral blood. Cytometry (Comm. Clin. Cytometry) 42:209-214, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861695 TI - A rapid measurement of apoptosis-associated light scatter changes using a hematology analyzer. AB - The alternative application of an automated hematology analyzer, H*3 system, has been described for the detection of apoptosis. Apoptosis induction by the topoisomerase I inhibitor, camptothecin (CAM) on several cell lines is followed by typical morphological alterations. On the H*3 cytogram, measurement of CAM treated cells revealed an increased population of cells with reduced size suggesting cell contraction during apoptosis. The decreased LUC/Lymph ratio also indicated the enhanced degree of apoptosis directly correlated with increasing CAM concentration and/or incubation period. Quantitative analysis shows a good correlation between the H*3 measurement and flow cytometry measurements of Annexin V-fluorescein isothiocyanate-labeled method. Thus, the H*3 measurement, under an appropriate adjustment, can be used as a rapid monitor for evaluating the degree of apoptotic changes in drug susceptibility testing of homogeneous cell samples. PMID- 10861696 TI - Influence of season on the incidence of DNA hypodiploidy in urinary cytology. AB - OBJECTIVE: This study was conducted to determine if an incidence of hypodiploidy in urinary specimens is related to seasonal temperature changes. MATERIALS AND METHODS: DNA ploidy was evaluated on 10,846 urinary specimens fixed in buffered alcohol (MOPSO/NaCl + ETOH) and received over a one year period from numerous sites throughout the United States. The percentage of hypodiploid (DNA index < 0.8) cases was evaluated in each month. As a control, DNA ploidy results from 3, 755 prostate biopsies, fixed in 10% neutral buffered formalin, received during the winter and summer months of the same year, were evaluated. RESULTS: The average percentage of hypodiploidy in cytologic specimens during the summer months was 19.6% compared to 5. 4% in the winter and early spring months (range: 20.6-4.8%). The average percentage of hypodiploid cells in histologic specimens was 0.8% for both the summer and winter months (range: 1.73-0.36%). CONCLUSIONS: The rate of hypodiploidy in urinary cytology seems to be temperature related. The hypodiploidy rate of histologic specimens fixed in formalin shows no fluctuation with the seasons. PMID- 10861697 TI - Coronary revascularization in heart transplant recipients by excimer laser angioplasty. AB - BACKGROUND AND OBJECTIVE: Aggressive development of allograft coronary artery disease is a major cause of death in heart transplant recipients. Percutaneous balloon angioplasty is considered suboptimal for complex lesions in native coronary vessels and heart transplant recipients, alike. Excimer laser energy (308-nm wavelength) can successfully remove and vaporize atherosclerotic plaques in native coronary vessels; however, its application in heart transplant recipients has not been studied clinically yet. STUDY DESIGN/MATERIALS AND METHODS: Six heart transplant recipients underwent percutaneous excimer laser (CVX-300, Spectranetics, Colorado Springs, CO) coronary angioplasty for treatment of a total of 10 discrete, obstructive coronary artery lesions. By using concentric or eccentric multifiber laser catheters, energy parameters were set at a fluence of 45 mJ/mm(2) or 60 mJ/mm(2) with a frequency of 25 Hz and 40 Hz, respectively, with a pulse duration of 135 ns and output of 200 mJ/pulse. The "saline flush" and "pulse and retreat" lasing techniques were used. In each case, adjunct balloon angioplasty was performed; in five lesions, an intracoronary stent was implanted. Angiographic evaluation was performed by visual assessment. RESULTS: Each procedure was successful as defined by laser recanalization of the target lesion (reduction of target lesion stenosis in more than 20%) and subsequent adequate final luminal patency (reduction of target lesion stenosis to less than 50%) and absence of any major in-cardiac catheterization complication (such as perforation, acute closure, dissection, emergency coronary artery bypass surgery), or in-hospital complications (such as death, myocardial infarction, cardiac enzyme elevation, major bleeding), or need for surgical revascularization. A 92 +/- 5% preprocedural percent diameter stenosis was reduced by laser to 35 +/- 16% and by adjunct balloon angioplasty in all lesions and stenting in five lesions, to final residual stenosis of 2 +/- 6%. Angiographic follow-up between 2 and 6 months after the procedure demonstrated a target lesion restenosis rate of 22%. CONCLUSION: Percutaneous excimer laser is safe and efficacious in the treatment of focal obstructive lesions caused by allograft coronary artery disease. These data represent an early clinical experience; thus, the long-term outcome of this revascularization method in recipients of heart transplantation will have to be determined by a large scale prospective, randomized, multicenter clinical study. PMID- 10861698 TI - Preliminary study of in vivo autofluorescence of nasopharyngeal carcinoma and normal tissue. AB - BACKGROUND AND OBJECTIVE: In nasopharyngeal cancer, conventional white light endoscopy does not provide adequate information to detect the flat/small lesion and identify the margin of observable tumor. In the present study, we evaluate the potential of light-induced fluorescence spectroscopic imaging for the localization of cancerous nasopharyngeal tissue. STUDY DESIGN/MATERIALS AND METHODS: We built a multiple channel spectrometer specifically for the investigation of fluorescence collected by a conventional endoscopic system. Nasopharyngeal fluorescence were measured in vivo from 27 subjects during the routine endoscopy. The biopsy specimens for histologic analysis were taken from the tissue sites where the fluorescence were measured. RESULTS: Two algorithms to discriminate the nasopharyngeal carcinoma from normal tissue were created based on the good correlation between the tissue autofluorescence and histologic diagnosis. For the two-wavelength algorithm, carcinoma can be differentiated from normal tissue with a sensitivity and specificity of 93% and 92%, respectively. For the three-wavelength algorithm with compensation of variation of blood content in tissue, a sensitivity of 98% and specificity of 95% were achieved. CONCLUSION: Fluorescence endoscopic imaging used with the algorithms developed in this report is an efficient method for detecting the nasopharyngeal carcinoma. PMID- 10861699 TI - MS-2 fibrosarcoma characterization by laser induced autofluorescence. AB - BACKGROUND AND OBJECTIVE: MS-2 fibrosarcoma implanted in BALB-CDF1 mice was investigated by frequency and time domain measurements of the autofluorescence (AF) radiation emitted upon excitation by a N(2) laser beam (337.1 nm). STUDY DESIGN/MATERIALS AND METHODS: AF spectra were obtained by using a spectrograph, a multichannel plate and an optical multichannel analyzer for the steady state detection. Time-resolved spectra were performed by means of a monochromator, a photomultiplier, and a digital signal analyzer. RESULTS: Spectral measurements show that the autofluorescence intensity of pathologic tissue is lower than that of healthy one in the 400- to 500- spectral region. In the same spectral range, we found the fluorescence decay to be the sum of a fast and a slow component. The lifetime of the fast component of tumoral tissue is significantly lower than that of healthy samples. CONCLUSION: Frequency and time domain measurements used in combination show that MS2-fibrosarcoma is characterized by the probable presence of the free form of NADH. PMID- 10861700 TI - Enhanced cell death in NR-S1 tumor by photodynamic therapy: possible involvement of Fas and Fas ligand system. AB - BACKGROUND AND OBJECTIVES: To investigate the effect of photodynamic therapy (PDT) on cell death in malignant tumor tissue, the frequency of terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL)-positive cells and the possible involvement of Fas and Fas ligand system were evaluated. STUDY DESIGN/MATERIALS AND METHODS: NR-S1 tumor-bearing C3H/HeNCrj mice were treated by PDT with Photofrin(R) (12 mg/kg body weight) and Nd:YAG dye laser (630 nm, 10 Hz, 150 J/cm(2)). Paraffin-embedded tissue sections from the excised tumor tissues at 6, 12, 24, 48 hours after PDT were analyzed by TUNEL for the occurrence of apoptosis and by immunohistochemistry for Fas and Fas ligand (FasL) expression. TUNEL-positive cells as well as Fas- or FasL-positive cells were counted and expressed as a percentage of positive cells per total cells. RESULTS: Based on the percent area of tumor necrotic foci, the most effective conditions for PDT were first determined. Under these conditions, PDT increased the number of TUNEL-positive tumor cells at 12 hours after irradiation. In parallel with the increase in TUNEL-positive cells, Fas-positive tumor cells were also found in the same area where many TUNEL-positive tumor cells were found. The expression of Fas ligand was found in the tumor cells surrounding TUNEL-positive cells on serial sections. A significant increase in FasL-positive lymphocytes was observed at 12 hours, whereas the infiltration of such lymphocytes into the area where TUNEL positive tumor cells were observed was rare. CONCLUSION: The possible role of Fas/FasL system in the cell death induced by PDT with Photofrin(R) and Nd:YAG dye laser was suggested. Moreover, the role of infiltrated lymphocytes seemed not to be so much in this model. PMID- 10861701 TI - Effect of photodynamic therapy in a multimodal approach for advanced carcinoma of the gastro-esophageal junction. AB - Background and Objective We wanted to determine the role of additional photodynamic therapy in a multimodal approach for the treatment of patients with advanced cancer of the gastro-esophageal junction. Study Design/Materials and Methods We reviewed 53 patients, after endoluminal palliation, with advanced cancer of the gastro-esophageal junction. Combined dilatation and retrograde tumor disobliteration with Nd-YAG laser before photodynamic therapy (PDT), brachyradiotherapy, or both, became necessary in 12 patients. Brachyradiotherapy was carried out in all patients. PDT before brachyradiotherapy was performed in 25 patients. The endoluminal treatment was completed by external beam irradiation in 30 patients (15 cases with PDT and 15 without PDT) with an at least fair performance status. RESULTS: Photodynamic therapy showed a significant difference regarding the mean opening of the tumor stenosis (mean, 6.4 mm; P = 0.0002), the mean decrease in tumor length (3.1 cm; P = 0.00001) and the increase in median survival (13. 8 months; P = 0.001). The combined multimodal approach by using PDT, brachyradiotherapy and external beam irradiation showed a median survival of 16.8 months. However, additional external beam irradiation showed no significant difference (P = 0.11). The rate of severe complications was 5.7%. The mortality rate was 1.9%. CONCLUSION: Photodynamic therapy has been shown to be an effective treatment for palliation of advanced cancer at the gastro-esophageal junction. The use of PDT combined with irradiation was associated with an acceptable survival rate, low rates of complications and reasonable quality of life. PMID- 10861702 TI - Temperature mapping of magnetic resonance-guided laser interstitial thermal therapy (LITT) in lymphangiomas of the head and neck. AB - BACKGROUND AND OBJECTIVE: Lymphangiomas of the tongue and neck are uncommon benign congenital lymphatic tumors. These vascular lesions are difficult to treat, frequently recur, and can cause patients significant morbidity. Treatment may also be complicated by adjacent vital anatomic structures. Magnetic resonance (MR)-controlled laser-induced interstitial thermotherapy (LITT) has been proven to be a noninvasive safe treatment. Real-time monitoring of tissue temperature with thermosensitive sequences allows controlled coagulation necrosis. STUDY DESIGN/MATERIALS AND METHODS: LITT was performed in a lymphangioma specimen ex vivo. In four patients (eight procedures) with lymphangiomas of the tongue and neck, MR-guided LITT was performed with a percutaneous approach in a multiapplicator technique. The laser system consisted of a titanium catheter and a protective catheter. The dome of the fiber end had a diameter of 1.4 mm with an active length of 20 mm. Temperature sensitive sequences were used in a 0.5 T open configured MR scanner with the proton frequency shift technique to map the spatial and temporal distribution of Nd:YAG laser effects (7 Watts, 30 pulses per second, 10 minutes/location). Postoperative MR follow-up was performed at 1 week and at 3 months. In three patients, partial resection of the tumor was performed 6 months after LITT. RESULTS: In three patients, MR clearly showed a diminished tumor volume. All four patients reported subjective amelioration and in three patients former functional problems, such as speech and swallowing were improved. MR thermometry allowed accurate demarcation of changes by heat and distinction of affected tumor volume (3.0 cm +/- 0.3 cm). The histology of the patients 6 months after LITT showed laser-induced fibrosis of former lymphatic tissue. CONCLUSION: The results suggest that LITT can be performed safely with tissue preserving of vital structures and can be effective in the treatment of deep tumors, such as lymphangiomas. However, given the nature of the lesion, the potential for recurrence exists no matter what modality is chosen. PMID- 10861703 TI - Pulsed Nd:YAG laser irradiation of the tooth pulp in the cat: II. Effect of scanning lasing. AB - BACKGROUND AND OBJECTIVE: The purpose of the present study was to assess whether "scan irradiation" with a pulsed Nd:YAG laser could produce changes in intrapulpal nerve activities and pulpal blood flow and to investigate whether it would cause tissue damage in the pulp. STUDY DESIGN/MATERIALS AND METHODS: The pulsed Nd:YAG laser was used to irradiate, in a scanning manner, the canine tooth pulp in sodium pentobarbitone anesthetized cats. The compound action potentials and spike response in the functional single afferent nerve fibers were recorded while responding to various external stimuli applied to the exposed dentin. Histologic observation was performed to detect lasing-induced tissue changes. RESULTS: Pulpal compound action potentials evoked by various external stimuli were significantly reduced (P < 0.05) and unit firings were observed in both functional single A delta- and C-fibers during irradiation. Unit responses to external mechanical stimulation of the dentin completely disappeared after "scan irradiation" with the pulsed Nd:YAG laser. Histologic observation revealed that irradiation with the laser produced tissue damage in the pulp. CONCLUSION: "Scan irradiation" with the pulsed Nd:YAG laser of cat's teeth produced alterations in the intrapulpal nerve activities, as well as caused tissue damage in the pulp. PMID- 10861704 TI - Lack of effect of pulsed low-intensity infrared (820 nm) laser irradiation on nerve conduction in the human superficial radial nerve. AB - BACKGROUND AND OBJECTIVE: To investigate the effect of pulsed low-intensity laser irradiation on nerve conduction in the human superficial radial nerve and on temperature in the skin overlying the nerve. STUDY DESIGN/MATERIALS AND METHODS: Thirty-two healthy human volunteers were recruited and randomly assigned to either placebo, laser 1 (9.12 Hz), laser 2 (73 Hz), or control groups (n = 8 all groups). A GaAlAs laser diode (820 nm, 50 mW peak) was used to irradiate the skin overlying the right superficial radial nerve at three points (1.2 J per point; energy density, 9.55 J/cm(2)). Antidromic action potentials were recorded from the superficial radial nerve preirradiation and at 5, 10, and 15 minutes after irradiation. Skin temperature was monitored concomitantly by using two surface thermistor probes attached to the skin overlying the nerve. RESULTS: Repeated measures analysis of variance showed no significant differences between groups for negative peak latency nor skin temperature data after laser irradiation. CONCLUSION: This study has demonstrated that laser irradiation at the radiant exposure and pulsing parameters indicated did not produce any specific neurophysiologic effects in this model of nerve function. PMID- 10861705 TI - Laser-tissue interaction with a continuous wave 3-mcm fibre laser: preliminary studies with soft tissue. AB - BACKGROUND AND OBJECTIVE: Lasers operating at wavelengths in the mid-infrared region have become increasingly popular for applications in areas of surgery and medicine. Advances in fibre laser technology have introduced a highly efficient, compact, diode-pumped source operating at around the 3-mcm wavelength. This study examines the effects of this recently developed laser on soft biological tissue. STUDY DESIGN/MATERIALS AND METHODS: Chicken breast and liver tissue samples were exposed to 800 mW continuous wave laser power, at a wavelength of 2.71 mcm, with incident spot sizes of around 150 mcm. Samples were inspected grossly immediately after laser irradiation and also prepared for histologic processing. RESULTS: After irradiation, visual assessment of changes at sample surfaces indicated a region of thermally affected tissue surrounding the ablation crater. This region was observed to grow in size to around 1.0 mm in diameter after 3 seconds of laser exposure at 800 mW. An ablation velocity of 0.80 mm.s(-1) was determined in chicken breast for the same incident laser parameters. Analysis of samples irradiated at 800 mW and processed for histology revealed minimal damage at hole boundaries and no signs of char formation, providing incident exposure times were restricted to below around 0. 5 seconds. CONCLUSION: This fibre laser source has demonstrated its potential to fulfil medical applications, enabling accurate, precise tissue removal to proceed at a rapid ablation rate. The efficiency and small size of the laser are attractive features. PMID- 10861707 TI - Early prenatal management of a fetal ventricular tachycardia treated in utero by amiodarone with long term follow-up. AB - Fetal cardiac arrhythmias are one of the causes of intra-uterine congestive heart failure and non-immune hydrops fetalis leading to fetal death. As ventricular tachycardia (VT) is rarely diagnosed in utero, it leads to emergency deliveries. We report a prenatal diagnosis of fetal tachycardia at 20 weeks of gestation associated with non-immune hydrops fetalis. The tachycardia seemed to be supraventricular and was initially treated by digoxin and sotalol. The hydrops increased and sotalol was stopped in order to give the mother a high dose of amiodarone by mouth over a long period. Although the tachycardia, which the ECG recorded at birth revealed to be of ventricular origin, persisted but at a lower rate, the new treatment proved successful. The child is three years old now and health, though with persistent VT. In conclusion, fetal tachycardia with similar ventricular and atrial rates can be a VT and the drug of choice in this case seems to be amiodarone. PMID- 10861708 TI - The applicability of different PCR-based methods for fetal RHD and K1 genotyping: a prospective study. AB - The applicability of different PCR-based assays for fetal RHD and K1 genotyping using DNA isolated from uncultured amniotic fluid cells has been tested prospectively: cord blood serotyping served as a control. For RHD genotyping, DNA was amplified with PCRs specific for RHD exon 7, the 3'-non-coding region and intron 4, using standard conditions. The results of these three separate assays were compared to those of a newly-developed multiplex PCR, simultaneously amplifying six regions of RHD. The PCRs analysing the 3'-non-coding region or intron 4 often yielded false-negative results or no results at all. Results of the exon 7 PCR and of the multiplex PCR always corresponded with postnatal serotyping, the multiplex PCR having the advantage of analysing six RHD-specific exons simultaneously. For K1 genotyping, two different PCR-based assays, both analysing the presence of T578C in the KEL gene, were applied. With the first method, a consensus 740-bp product of the KEL gene was amplified and subsequently specifically digested. As we were not able to obtain any PCR product from amniotic fluid DNA, we developed a new K1-specific PCR, amplifying a fragment of 91 bp only in cases of K1-positivity. With this PCR, all K1 genotyping results (n=30) correctly predicted the phenotypes. We conclude that fetal RHD and K1 genotyping can be performed reliably with DNA from uncultured amniotic fluid cells. PMID- 10861709 TI - Prognostic factors associated with congenital cystic adenomatoid malformation of the lung. AB - This study presents 18 cases of prenatally diagnosed congenital cystic adenomatoid malformation (CCAM) to identify potential factors that could predict prognosis. Comparisons of prenatal parameters were made between fetuses that survived and those that died perinatally. It was found that microcystic lesion, bilateral lung involvement and hydrops were each highly correlated with poor prognosis, while neither polyhydramnios nor mediastinal shift was significantly associated with had outcome. Fetal interventions were indicated only in two of the surviving cases: a thoracocentesis and a cysto-amniotic shunt. A therapeutic amniocentesis was performed in one case of polyhydramnios. The diagnosis of CCAM was histologically confirmed in all cases by necropsy or by postnatal lobectomy. PMID- 10861710 TI - Prenatal UPD testing survey in Robertsonian translocations. AB - A systematic search was made for uniparental disomy (UPD) in familial or de novo balanced Robertsonian translocations, identified by prenatal cytogenetic investigations. Parent-of-origin studies were performed using molecular markers for both chromosomes involved in the translocation. No UPD cases were identified out of 23 analysed cases. The results presented here, combined with other available data, provide preliminary elements for genetic counselling in these common chromosomal rearrangements. PMID- 10861711 TI - Long-term effect of prospective detection of high genetic risk on couples' reproductive life: data for thalassaemia. AB - Prospective risk detection with availability of prenatal diagnosis is the best service currently available for couples at high genetic risk Here we describe the long term effect of this service on the reproductive life of 102 couples at risk of thalassaemia, whose risk was detected prospectively by carrier screening, who made use of prenatal diagnosis, and where the woman is now over 40. Overall outcome for couples is described in terms of number of favourable versus unfavourable pregnancy outcomes. (A favourable pregnancy outcome = unaffected livebirth, or affected livebirth resulting from informed parental choice.) The 102 couples had a total of 356 pregnancies, including 302 viable pregnancies, and 88% achieved a family unburdened by thalassaemia. 68% of viable pregnancies had a favourable outcome, but only 43% of couples had only favourable outcomes, and 26% lost two or more viable wanted pregnancies. When early losses are included 58% of pregnancies had a favourable outcome, but only 30% of couples had only favourable outcomes, and 41% lost two or more pregnancies. Even with the best available service, at risk couples remain victims of chance, and a significant minority experience great difficulty in obtaining even one healthy child. Research is needed on approaches that may allow couples better control of reproductive outcomes. PMID- 10861712 TI - Prenatal diagnosis of hypoparathyroidism retardation and dysmorphism (HRD) syndrome. AB - We used linkage analysis for prenatal diagnosis of the recently reported hypoparathyroidism, retardation, and dysmorphism (HRD) syndrome. Five cases from four families were evaluated. Three fetuses were carriers and were born healthy. Two fetuses were affected but the parents decided not to terminate the pregnancies. The diagnosis of HRD syndrome was confirmed in these newborns. This is the first report about prenatal diagnosis of HRD syndrome. PMID- 10861713 TI - Quantitative FISH analysis and in vitro suspension cultures of erythroid cells from maternal peripheral blood for the isolation of fetal cells. AB - Several techniques for the enrichment of nucleated fetal red blood cells present in maternal blood have been reported. Here we describe the use of a quantitative fluorescence in situ hybridization (FISH) method and in vitro suspension cultures of erythroid cells from newborn cord blood and maternal peripheral blood. Together with a rapid high performance liquid chromatography (HPLC) method, that allows us to determine as few as 100 cells containing haemoglobin F (HbF), we have scrutinized the reported enrichment methods for fetal nucleated cells in peripheral maternal blood. One hundred FISH analyses on maternal peripheral blood were performed. The method comprises a cell lysis method for depletion of red cells with minimal losses of nucleated cells, uniform numbers of cells (750 000 cells each) on microscopic slides, and inclusion of internal controls to monitor the efficacy of hybridization. Twenty-six cultures of pure erythroid progenitor cells from maternal peripheral blood were analysed for the expansion of fetal cells. To generate these in vitro cultures, nucleated cells from 10-20 ml of peripheral blood from 26 pregnant women were grown in media containing growth factors and hormones to yield over 10(7) of immature erythroid cells within two weeks. Of those, 13 cultures were from pregnancies with confirmed male fetuses. A total of approximately 8x10(8) maternal cells were added into tissue culture medium for these 13 cultures, resulting in about 2x10(8) nearly pure erythroid cells after two weeks. Whereas fetal cells, alone or added into cultures of peripheral blood, grow rapidly and can be detected quantitatively, we could not find any fetal cells in cultures from maternal blood. Likewise, in 7.5x10(7) peripheral blood cells probed by FISH analysis (half of which were from pregnancies with male fetuses) no single Y chromosome was detected. In summary, suspension cultures of erythroid cells can be established routinely and easily. With the quantitative FISH technique used, 750 000 cells per slide can be screened reliably for cells with Y chromosomes. However, the stringent quality criteria and most elaborate methods indicate that fetal cells in maternal peripheral blood can not be found using the current technology. PMID- 10861714 TI - Acceptability of serum screening as an alternative to cytogenetic diagnosis of down syndrome among women 35 years or older in Hong Kong. AB - The addition of second trimester serum markers to maternal age increases the efficacy of screening for Down syndrome by maternal age alone. Among women aged 35 years or older, serum screening makes a large proportion of amniocentesis unnecessary. However, there are ethical and medicolegal concerns about serum screening in 'old' women, largely because some of the pregnancies affected by Down syndrome and other chromosomal abnormalities may not be detected. We investigated the acceptability of serum screening in women aged 35 years or older when it was offered as an alternative to prenatal cytogenetic diagnosis after detailed counselling. Women referred for prenatal diagnosis of Down syndrome because of advanced maternal age were given the options of cytogenetic diagnosis by chorionic villus sampling (CVS) or amniocentesis. As an alternative, they could choose to undergo second trimester serum human chorionic gonadotrophin (hCG) and alpha-fetoprotein (AFP) screening first before deciding on whether to undergo amniocentesis. Between January 1997 and October 1999, 3419 subjects were recruited. 1807 women (52. 9%) chose to undergo serum screening, 1516 women (44.3%) chose to have amniocentesis and 96 women chose to have CVS (2.8%). The proportion of women who chose serum screening rose steadily from 38. 8% in the year of 1997 to 63.4% in 1999. Significantly fewer Chinese women chose serum screening than non-Chinese. The decision as to whether to undergo an invasive diagnostic procedure or to be content with the relatively safer but less accurate screening test varies, being affected by the women's background and culture. PMID- 10861715 TI - The influence of ethnic origin on first trimester biochemical markers of chromosomal abnormalities. AB - In a first trimester study of 5422 Caucasian women, 752 Afro-Caribbean women and 170 Asian women we have shown that the median maternal serum marker MoMs for free beta-hCG and PAPP-A were 19% and 48% higher in Afro-Caribbean women and 19% higher and 35% higher in Asian women, compared to Caucasian women. Correcting for maternal weight made very little difference to the effect in Afro-Caribbeans (21% and 57% higher after weight correction) but reduced the effect in Asians (4% and 17% higher after weight correction ). It is estimated that correcting for maternal weight and ethnicity overall would increase the detection rate by a modest 1.4%. However, the effect on an individual's risk could result in as much as a two-fold increase in the patient specific risk for trisomy 21. The impact of ethnic origin seems to be greater than that observed with second trimester biochemical markers and larger studies are required in order to develop robust algorithms for correcting for ethnic origin in the first trimester. PMID- 10861716 TI - Screening for triploidy by fetal nuchal translucency and maternal serum free beta hCG and PAPP-A at 10-14 weeks of gestation. AB - In 25 cases of triploidy at 10-14 weeks of gestation, compared with 947 controls, the median multiple of the median (MoM) fetal nuchal translucency (NT) thickness was significantly increased (1.89 MoM), and maternal serum total and free beta human chorionic gonadotrophin (hCG) were increased (3.13 MoM and 4.59 MoM respectively), alpha fetoprotein (AFP) was increased (2.14 MoM), and pregnancy associated plasma protein A (PAPP-A) was decreased (0.12 MoM). There are two types of triploidy. In type I, where the additional chromosome set is of paternal origin, the placenta is partially molar and the fetus is relatively well-grown. Type II, where the extra chromosome set is of maternal origin, is characterized by a small normal looking placenta and severe asymmetrical fetal growth restriction. In type I triploidy there was increased fetal NT (2.76 MoM), maternal serum total hCG (4.91 MoM), free beta-hCG (8.04 MoM), and AFP (3.22 MoM), and mildly decreased PAPP-A (0.75 MoM). In type II triploidy fetal NT was not increased (0.88 MoM), and there was a decrease in maternal serum total hCG (0.16 MoM), free beta-hCG (0.18 MoM), PAPP-A (0.06 MoM) and AFP (0.77 MoM). We conclude that a large proportion of triploidy cases of both phenotypes could be identified in the first trimester using NT, maternal serum free beta-hCG and PAPP A with a combination of trisomy 21 risk and an atypicality approach. PMID- 10861717 TI - Prenatal ultrasonographic diagnosis of the popliteal pterygium syndrome. AB - The prenatal ultrasound identification of a cleft lip and palate, equinovarus feet with severe lower limb malposition and genital abnormalities led to the prenatal diagnosis of popliteal pterygium syndrome in a pregnant mother suspected to have a mild expression of this autosomal dominant condition. However, in sporadic cases with lack of a family history for this rare syndrome, prenatal diagnosis may be difficult to ascertain. PMID- 10861718 TI - Norman-Roberts syndrome: prenatal diagnosis and autopsy findings. AB - We report on the autopsy findings of a male fetus in the 27th week of gestation with Norman-Roberts syndrome. The unaffected parents are first cousins and have a five-year-old child with a low, sloping forehead, broad and prominent nasal bridge, widely set eyes, severe psychomotor retardation, and an agyric cortex. Prenatal diagnosis showed a small head at the 25th week of gestation. At this time, a slowing-down of the growth of the sonographic measurements of the biparietal diameter and head circumference was found. Both the biparietal diameter (57 mm, <5th percentile) and the head circumference (207 mm, <5th percentile) showed a delay of at least two weeks in comparison with other non cephalic somatometric parameters, that were normal for the gestational age (femur length: 46 mm=median value). After termination of pregnancy, post-mortem examination showed a normotrophic fetus with microcrania and marked microcephaly (brain weight: 50 g), low, sloping forehead, broad and prominent nasal bridge, and widely set eyes. The cerebral hemispheres displayed an almost completely smooth surface with poorly defined sylvian fissures and failure of operculization of the insula. Microscopic examination showed a predominantly four-layered cortex (lissencephaly type I). Karyotype was normal and in situ hybridization did not show any deletion in the Miller-Dieker/isolated lissencephaly critical region on 17p13.3. The syndromes with lissencephaly are reviewed. PMID- 10861719 TI - Subtle familial unbalanced translocation t(8;11)(p23.2;p15.5) in two fetuses with Beckwith-Wiedemann features. AB - We describe a subtle translocation t(8;11)(p23.2;p15.5) ascertained after two induced abortions in the same sibship because of the discovery of fetal hydrops on ultrasound examination. Initial cytogenetic studies performed on cultured amniotic fluid cells were considered as normal in both fetuses. High resolution banding analysis and FISH studies performed on the parents' chromosomes revealed a paternal translocation t(8;11)(p23.2;p15.5). Retrospective FISH analysis of both fetuses showed that they carried the same chromosomal imbalance including a distal monosomy 8pter and a distal trisomy 11pter. The phenotypes of the fetuses were re-examined and found to be compatible with Beckwith-Wiedemann syndromes (BWS). FISH analysis using an IGF2 probe demonstrated the presence of three copies of the IGF2 gene. This study highlights the value of searching for subtle chromosome rearrangements in families with recurrent unexplained multiple malformation syndromes discovered prenatally. Also, it contributes to a better delineation of the prenatal phenotype of BWS. Finally, it sheds new light on the aetiology of non-immune hydrops fetalis. PMID- 10861720 TI - Delay of chorioamniotic fusion: relation to chromosomal anomalies. PMID- 10861721 TI - Obstetrical three-dimensional ultrasound in the visualization of the intracranial midline and corpus callosum of fetuses with cephalic position. PMID- 10861722 TI - First trimester nuchal anomalies as a prenatal sign of Zellweger syndrome. PMID- 10861723 TI - Update on prenatal diagnosis of true mosaic trisomy 17 in amniocyte cultures. PMID- 10861724 TI - Three-dimensional sonographic visualization of a fetal omphalocele at 14 weeks of gestation. PMID- 10861725 TI - Transient fetal blood flow redistribution induced by maternal diabetic ketoacidosis diagnosed by Doppler ultrasonography. PMID- 10861726 TI - Current awareness in prenatal diagnosis. AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of prenatal diagnosis. Each bibliography is divided into 17 sections: 1 Books, Reviews & Symposia; 2 General Interest; 3 Normal Fetal Development; 4 Gametogenesis and Pre-implantation Diagnosis; 5 First Trimester Diagnosis; 6 Second Trimester Diagnosis; 7 Fetal Diagnosis by Ultrasound and Other Imaging; 8 Maternal Screening; 9 Screening for Carriers of Genetic Abnormality; 10 Technological Developments; 11 Confined Placental Mosaicism and Uniparental Disomy; 12 Molecular Cytogenetics; 13 Fetal Cells in Maternal Circulation; 14 Fetal Therapy; 15 Psychosocial Aspects; 16 Epidemiology and Environmental Factors; 17 Developmental Pathology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted PMID- 10861727 TI - The 2-pyridone antibacterial agents: bacterial topoisomerase inhibitors. AB - Many attempts have been made to prepare analogs of 4-quinolone antibacterial agents bearing novel ring systems, which might retain the favorable properties of these widely used antibacterial agents and at the same time increase activity against multidrug-resistant bacteria, streptococci, and anaerobic microorganisms. One such attempt involved bioisosteric exchange of the 1-N atom and 4a-C atom of naphthyridones, quinolones, and benzoxazines to produce a family of highly active pyridopyrimidines, quinolizines, and ofloxacin bioisosteres. These new antibacterial agents have been named collectively as the 2-pyridones. Many hundreds of 2-pyridones have been synthesized and evaluated in vitro and in vivo, and selected members are advancing toward human clinical trials. Preparation of these bioisosteres required the development of enabling chemistry, as previous methods were unsuccessful in producing the needed core structures. This review compares the structure-activity relationships of these agents with known trends among 4-quinolones, from which it is seen that there are many parallels, but also some significant departures as well. Generally, 2-pyridones are more highly active in vitro and in vivo and more water soluble than comparable 4-quinolones. These properties are posited to arise from electronic and conformational alternations in these new substances. Selected members show excellent pharmacodynamic properties, justifying the view that this is a very promising new class of totally synthetic antibacterial agents. PMID- 10861728 TI - New antiarrhythmic agents: a conceptually novel approach. AB - Synthesis and biological evaluation of novel phenoxyalkyl amines exhibiting both class Ib and class III type electrophysiological properties are described. Two compounds showed excellent antiarrhythmic effects against reperfusion induced arrhythmias. PMID- 10861729 TI - Multicomponent domino reactions for the synthesis of biologically active natural products and drugs. AB - A main issue in modern synthetic organic chemistry, which deals with the preparation of natural products, pharmaceuticals, diagnostics, agrochemicals, and other important materials, is the improvement of efficiency, the avoidance of toxic reagents, the reduction of waste, and the responsible treatment of our resources. One of the ways to fulfill these goals is the development and use of domino processes, which consist of several bond-forming reactions and which allow the highly efficient synthesis of complex molecules starting from simple substrates. Herein, the combination of several catalytic bond-forming transformations is clearly most appropriate. The synthesis of the enantiopure alkaloid (-)-hirsutine 22, which has a strong inhibitorial effect on influenza A viruses, was accomplished using a biomimetic domino Knoevenagel-hetero-Diels Alder-solvolysis-hydrogenation process. In a similar way the alkaloids (+) dihydrocorynantheine 23 and (-)-dihydroantirhine 24 as well as heterosteroids 62, D-homosteroids 65 and 68, and azasteroids 25 are prepared. In addition, novel steroid alkaloids 26 are accessible by a combination of the formation of an iminium salt, a hydride shift, and an alkylation. The anti-leukemic pentacyclic ( )-cephalotaxine 27 is obtained by a combination of two Pd-catalyzed reactions. PMID- 10861731 TI - The impact of chirality of the fluorinated volatile inhalation anaesthetics on their clinical applications. AB - This review discusses the various chromatographic enantioseparation methods on an analytical and preparative scale for fluorinated inhalation anaesthetics used clinically, namely halothane, enflurane, desflurane and isoflurane. The differences in the pharmacodynamics and pharmacokinetics between the enantiomers of those anaesthetics are presented. It can be concluded that using a single enantiomer for these fluorinated anaesthetics is advantageous over using the racemic mixture. The racemic switch to a single enantiomer for these fluorinated volatile anaesthetics offers a more effective and safe general anaesthetic. PMID- 10861732 TI - Determination of Ibuprofen in human plasma by high-performance liquid chromatography: validation and application in pharmacokinetic study. AB - A specific method for the simultaneous determination of S-(+)Ibuprofen and R-( )Ibuprofen enantiomers in human plasma is described. Adopting a high-performance liquid chromatographic (HPLC) system with spectrofluorometer detector, the compounds were extracted from plasma in alcohol medium and were separated on C18 column, using a solution of acetonitrile-water-acetic acid-triethylamine as mobile phase. The limit of quantitation was 0.1 microg/mL for both compounds. The method was validated by intra-day assays at three concentration levels and was used in a kinetic study in healthy volunteers. During the study we carried out inter-day assays to confirm the feasibility of the method. PMID- 10861733 TI - Enantioselective HPLC analysis of propafenone and of its main metabolites using polysaccharide and protein-based chiral stationary phases. AB - HPLC on chiral stationary phases has been used for the enantioselective assay of propafenone (PPF), 5-hydroxypropafenone (PPF-50H) and N-despropylpropafenone (PPF NOR) enantiomers. The results obtained on Chiralpak AD column showed that it is useful for the resolution of PPF and of its main metabolites, although the peaks obtained for PPF-NOR were not symmetrical under the conditions investigated. This column and circular dichroism-based detection system were used to determine the absolute configuration of the eluates. Furthermore, the influence of the mobile phase composition on the resolution of PPF and of its main metabolites was investigated on cellulose derivatives (Chiralcel OD-H and Chiralcel OD-R) and protein (Chiral AGP and Ultron ES-OVM)-based chiral stationary phases. The enantiomers of PPF were resolved on all the columns, except for the Ultron ES OVM. This column, the Chiralpak AD and the Chiralcel OD-H columns were suitable for the resolution of the PPF-50H enantiomers. The PPF-NOR enantiomers were resolved on the Chiralpak AD, Chiral AGP and Chiralcel OD-R columns. PMID- 10861734 TI - Visible diode laser induced fluorescence detection of doxorubicin in plasma using pressurized capillary electrochromatography. AB - Pressurized capillary electrochromatography is a variant of capillary electrochromatography (CEC) in which the driving force is both electroosmotic and hydraulic. The inlet of the CEC capillary is pressurized using an HPLC pump, and an electric field is simultaneously applied. This work describes a method for the analysis of doxorubicin. Doxorubicin was reacted with Cy5.29.OSu in acetonitrile. The derivative was confirmed by RP-TLC. A CEC system equipped with a VDLIF detector was constructed and used to analyze the derivative. The reaction mixture was injected onto a capillary packed in-house with 3 microm C-18 Luna particles and separation was carried out at 25 kV using 70% acetonitrile/ 30% phosphate (10 mM, pH = 4.8) as the mobile phase. The derivatization reaction was optimized by the investigation of parameters such as reaction time, temperature and concentration of label in order to increase the yield of the derivative. The optimal conditions were determined to be 30 min, 80 degrees C and 50 nmol/mL, respectively. Doxorubicin was extracted from plasma using solid-phase extraction under alkaline conditions, derivatized and injected onto the CEC-VDLIF system. The selectivity of the assay was demonstrated by a lack of interfering peaks due to plasma constituents across the elution window of the derivative peak in blank plasma extracts (n = 6 sources). The limit of detection (LOD) of the assay in plasma calculated as 3 s(b)/m was determined to be 1.7 ng/mL. The precision of the assay determined at a concentration of 167.7 ng/mL (n = 5) was found to be within 7.04 %RSD. PMID- 10861735 TI - Enantioselective determination of bromoisovalerylurea by liquid chromatography on chiral stationary phase in reversed- or normal-phase partition mode. AB - Bromoisovalerylurea (bromvalerylurea) is a sedative-hypnotic given orally as a racemate. Enantiomers of this drug could be separated by high-performance liquid chromatography on the three chiral stationary phases (a vancomycin-bonded, beta cyclodextrin derivative-bonded, or urea derivative-bonded phase). Biological fluids of human subjects who had ingested toxic or therapeutic doses of the racemate were chromatographed after liquid-liquid extraction. The (+)-enantiomer concentration was almost equal to the (-)-enantiomer concentration in the serum of one overdosed patient. In all the other subjects, the (+)-enantiomer was less than the (-)-enantiomer in their sera and saliva. The data suggest that the drug is absorbed non-stereoselectively from the gastrointestinal tract and eliminated from the blood stereoselectively. PMID- 10861736 TI - Determining the influence of storage time on the level of propofol in blood samples by means of chromatography. AB - Due to unsatisfactory equipment efficiency and the time consuming manual procedures of sample preparation, drug analyses in physiological fluids and tissues frequently have to be carried out a few days after the sample collection. This is especially the case with investigations which require the examination of materials for which a large number of samples is necessary. The paper deals with the influence of storing blood samples on the level of propofol in blood and plasma. Propofol (2,6-diisopropylphenol, Diprivan) is a very popular intravenous agent used both for the induction and the maintenance of anaesthesia in human and veterinary patients as well as in laboratory animals. The results obtained show that, due to distinct losses of propofol in samples during their storage, the comparison of data estimated for subsequent days after sampling can lead to misleading or even wrong conclusions. The speed of drug diminution depends both on the type of blood and the anticoagulant used. The established interdependencies between the change in the level of propofol in blood and plasma samples and their storage time show that analogous investigations of other pharmaceutical agents are necessary. PMID- 10861737 TI - Retention of barbituric acid derivatives on immobilized artificial membrane stationary phase and its correlation with biological activity. AB - A series of 30 barbituric acid derivatives were subjected to high-performance liquid chromatography (HPLC) on the 'immobilized artificial membrane' (IAM) column with acetonitrile buffer mobile phase. The retention parameter log k(IAM) was related to the logarithms of partition coefficients determined in octanol water partition system, log P, to a thin-layer chromatographic (TLC) parameter from partition TLC, R(m0), to an adsorption HPLC retention parameter, log k(0), and to a solubility parameter, delta. It was demonstrated that log k(IAM) correlated significantly to the other parameters of barbiturates determined in partition systems but not to delta. However, log k(IAM) appeared to be a distinctive descriptor of hydrophobicity of barbiturates as compared to the standard log P parameters. The parameter log k(IAM) was shown to correlate with bioactivity data of the agents studied. PMID- 10861738 TI - Determination of terbinafine in tissues. AB - Terbinafine and N-demethyl terbinafine concentrations were determined simultaneously in rat tissues by a high-performance liquid chromatography method. This method involved the homogenization of tissues (except for skin) followed by a liquid-liquid extraction. Skin samples were dissolved in sodium hydroxide prior to extraction. Terbinafine and its N-demethylated metabolite were assayed using a C(18) reversed-phase column with a mobile phase of acetonitrile and water (40:60) containing ortho phosphoric acid (0.02 M) and triethylamine (0.01 M), and UV detection (at 224 nm). The standard curve for the assay (constructed using clotrimazole as internal standard) was linear over the concentration range 100 3000 ng/g in skin and 10-600 ng/g in all other tissues. The inter- and intra-day precision for both terbinafine and metabolite was between 0.2% and 16%. The limit of quantification was 10 ng/g in all tissues and 100 ng/g in skin. This assay was found to be reliable and reproducible for the determination of terbinafine and N demethyl terbinafine concentration in all rat tissues and has been used for tissue distribution studies. PMID- 10861739 TI - Determination of YM992, a novel selective serotonin reuptake inhibitor, in rat and dog plasma by high-performance liquid chromatography with fluorescence detection. AB - A high-performance liquid chromatographic method with fluorescence detection was developed for the determination of (S)-2-[[(7-fluoro-4 indanyl)oxy]methyl]morpholine monohydrochloride (YM992) in plasma. Plasma samples were extracted with n-hexane under alkali condition. After the organic solvent was evaporated to dryness, the residue was treated with 4-fluoro-7 nitrobezofurazan (NBD-F) in borate buffer (pH 7.5) at room temperature for 20 min. The reaction was terminated with hydrochloric acid and the resultant solution was injected onto HPLC without further purification. No interfering peak was observed at the retention time of YM992 or the internal standard. The calibration curve was linear with the concentration of YM992 up to 200 ng/ml. The limit of quantitation was 1 ng/ml. The intra- and inter-day relative standard deviation was less than 5.6% and 4.1%, respectively, and the intra- and inter-day relative error ranged from -3.0% to 17.2% and 2.8% to 7.5%, respectively. Using the assay, the plasma concentration of YM992 could be determined up to 8 and 10 h after the oral administration of YM992 to rats and dogs, respectively. PMID- 10861740 TI - Simultaneous determination of cefazolin in rat blood and brain by microdialysis and microbore liquid chromatography. AB - A sensitive microbore liquid chromatographic method combined with the minimally invasive technique of microdialysis was devised for simultaneously and continuously monitoring the levels of unbound blood and brain cefazolin in rats. Microdialysis probes were inserted into the jugular vein and brain striatum for blood and brain sampling, respectively. Chromatographic conditions consisted of a mobile phase of methanol-acetonitrile-100 mM monosodium phosphoric acid (20:10:70, v/v, pH 4.5) pumped through a microbore reversed-phase column at a flow rate of 0.05 mL/min. The ultraviolet detection wavelength was set at 270 nm. An on-line design allowed direct and continuous analysis of protein-free samples in the dialysate. Microdialysis probes, being home-made, were screened for acceptable in vivo recovery. Chromatographic resolution and detection were validated for response linearity as well as intra-day and inter-day variabilities. This method was then applied to pharmacokinetic profiling of protein unbound cefazolin in both the blood and brain following intravenous administration (10 mg/kg, i.v., n = 6). Rapid appearance of cefazolin in the rat brain striatal dialysate following drug injection suggested good blood-brain barrier penetration. According to a non-compartmental pharmacokinetics model, the area under the concentration (AUC) vs time ratio of cefazolin in rat brain and blood was 6%. PMID- 10861742 TI - Introduction PMID- 10861741 TI - Morris Pollard: a pioneer in developing animal models for prostate cancer. PMID- 10861743 TI - Mouse prostate reconstitution model system: A series of in vivo and in vitro models for benign and malignant prostatic disease. AB - BACKGROUND: An elucidation of the complex, morphological and molecular changes that underlie benign and malignant prostatic disease will likely lead to improved methods of diagnosis and therapy for those disorders. To identify and understand the interrelation of the phenotypic and genetic changes inherent in these important diseases requires the development and use of in vivo and in vitro models that closely mimic specific aspects of the disease process. Once the suspected molecular underpinnings of prostatic disease are uncovered, in vivo and in vitro models will be required for further testing of the functional significance of specific genetic alterations as they are identified. In addition models of prostatic disease are necessary to evaluate novel therapeutic approaches. METHODS: The mouse prostate reconstitution (MPR) model system was developed more than a decade ago with these specific needs in mind. Over the years, specific modifications of the MPR model have demonstrated its versatility and applicability for the study of benign and malignant prostatic disease, including metastatic progression. RESULTS: We discuss various modifications of the MPR model system made for its application to specific aspects of prostatic disease; the clinically relevant information that has been gleaned thus far from the use of this model system; and advances on the horizon for the expansion of its role in prostate research. CONCLUSIONS: The MPR model system has contributed substantially to the understanding and treatment of benign and malignant prostatic diseases. Additional modifications in this series of in vivo and in vitro models will likely lead to further advances. PMID- 10861744 TI - Isolation and characterization of mouse probasin: An androgen-regulated protein specifically expressed in the differentiated prostate. AB - BACKGROUND: The development and growth of the prostate gland is regulated, in part, by a variety of steroid and polypeptide growth-factor hormones. As a consequence of hormone action, the prostate gland will produce a number of tissue restricted gene products. Characterization of the regulation, expression, and function of genes encoding prostate-specific proteins is critical to our understanding of prostate biology. Probasin is a prostate-specific gene originally isolated from the rat and has been exploited as a marker of prostate differentiation and to elucidate androgen action. Furthermore, a number of transgenic mouse models of prostate cancer have been established based on the regulatory elements derived from the rat probasin gene. In this report, we describe the isolation and characterization of the mouse probasin ortholog to further facilitate studies related to hormone action in the prostate and the generation and characterization of novel autochthonous models of prostate cancer. METHODS: Mouse probasin cDNA was isolated from a phage library, and the DNA sequence was determined. The predicted protein sequence was used to generate specific oligonucleotide primers and antibodies. Probasin protein and RNA expression were examined by immunobloting, immunohistochemistry, and RT-PCR, in normal mouse prostate tissue and tumor tissues derived from the autochthonous "transgenic adenocarcinoma of the mouse prostate" (TRAMP) model. Regulation of probasin expression in response to surgical castration and hormone supplementation was also characterized. RESULTS: Several points of evolutionary sequence conservation were identified between mouse and rat probasin, especially in the 3' untranslated region. Specific polyclonal antibodies were generated to peptide fragments, and the temporal and spatial pattern of probasin expression was examined. The expression of probasin was primarily localized to the apical membrane of differentiated secretory epithelium. Probasin mRNA and protein were absent from the poorly differentiated tissue of TRAMP tumors. Probasin was found to be androgen-regulated. In contrast to data from studies on rat probasin, no postcastration rebound of mouse probasin mRNA was observed. CONCLUSIONS: Probasin is a marker of differentiation and androgen action in the mouse prostate, and strong sequence conservation between mouse and rat probasin supports an essential role for this gene in the biology of the prostate gland. Isolation and characterization of mouse probasin will facilitate further development and analysis of autochthonous mouse models of prostate cancer. PMID- 10861745 TI - Use of nude mouse xenograft models in prostate cancer research. AB - BACKGROUND: Our understanding of the mechanisms of (progressive) growth of prostatic cancer has been largely obtained through the study of experimental animal models. To be able to validate new concepts, representative model systems of human origin that mimic the clinical process of the disease in patients are essential. Unfortunately, the limited number of human prostate tumor models has considerably hampered research. METHODS: Various research groups have put much effort in the development of human prostate tumor xenograft models, and large numbers of clinical prostate tumors were heterotransplanted in immune-deficient host animals. This huge effort has resulted in a number of tumor lines which are reviewed here. RESULTS: Up to now, approximately 25 xenograft models of human prostate cancer have been established and reported in the literature. The available xenografts seem to represent the various stages of clinical prostate cancer, such as early progression and transition from androgen-dependent to androgen-independent growth. In addition, recent efforts are concentrating on the establishment of in vitro cell lines from these xenografts as well as on the development of (bone) metastatic variants. CONCLUSIONS: Xenograft models are important for elucidating regulatory pathways of tumor growth and progression and are indispensible for testing of new treatment modalities. PMID- 10861746 TI - Life expectancy, antagonistic pleiotropy, and the testis of dogs and men. AB - BACKGROUND: Prostate cancer and benign prostatic hyperplasia are important age related prostatic diseases that are under the influence of testicular hormones. However, the disparity between male and female life expectancy within the human population cannot be explained solely by the prevalence of prostatic disease related mortality. The purpose of this paper is to explore the possibility that the testis exerts a detrimental effect on life span. METHODS: First, we review previously published and unpublished data on the influence of the testis on the life span of dogs and men. Aging in pet dogs and men is then discussed in terms of evolutionary theory, emphasizing the significance of a prolonged postreproductive life span and possible consequences of late-acting deleterious genes in these two species. Finally, we present preliminary data that orchiectomy can reduce DNA damage within the brain of elderly male dogs. RESULTS AND CONCLUSIONS Taken together, these observations raise the intriguing possibility that interventions to antagonize the testis might have much broader therapeutic applications that will extend well beyond the treatment of prostate cancer. PMID- 10861747 TI - Prostatic neoplasia in transgenic mice with prostate-directed overexpression of the c-myc oncoprotein. AB - BACKGROUND: Promoter elements within the 5' DNA region of the rat C(3)1 gene have been shown to direct prostate-specific expression of gene products when they are fused through recombinant DNA procedures and used to produce transgenic mice. In order to test the in vivo effects of chronic overexpression of the mouse c-myc protooncogene on the prostate glands of transgenic mice, we created several lines of C(3)1-c-myc transgenic mice and then examined the phenotype of males with this genetic alteration. METHODS: The modified promoter and 5' region of the rat C(3)1 gene was fused to the coding region of the mouse c-myc gene using recombinant DNA techniques. This DNA was used to create three different founder lines of transgenic mice. Tissues from males and females heterozygous for the transgene were examined for expression of the recombinant mouse c-myc mRNA by an RNase protection assay. Prostates from males were examined for expression of recombinant c-myc mRNA by in situ hybridization. Thin sections of fixed ventral prostates from males were analyzed by microscopy for histological abnormalities. RESULTS: Three different lines of transgenic mice were obtained from these procedures. These mice demonstrated expression of recombinant mouse c-myc mRNA in the testis and ventral prostates of males and in the uterus of females. In situ hybridization demonstrated that the epithelial cells were the source of recombinant c-myc expression in the ventral prostates of the transgenic lines. Microscopic analysis of the ventral prostates from these mice demonstrated abnormalities in epithelial cell morphology seemingly typical of an intraepithelial neoplasia-like phenotype. However, none of the males of any of the lines developed overt prostatic adenocarcinoma over their lifetimes. CONCLUSIONS: Chronic overexpression of c-myc in the ventral prostate epithelial cells of C3(1)-c-myc transgenic mice leads to the development of epithelial cell abnormalities similar to those seen in low-grade prostatic intraepithelial neoplasia in humans. These abnormalities were not found to progress to adenocarcinoma over the lifetimes of the transgenic mice, suggesting the need for additional oncogenic changes in the pathway to prostatic adenocarcinomas. Furthermore, our cumulative experience with the use of the C3(1) gene promoter in the generation of transgenic mice suggests that the probasin promoter element provides a much more specific and effective means to target transgenes to the prostate glands of mice. PMID- 10861748 TI - Prostatic intraepithelial neoplasia: animal models 2000. AB - Numerous animal models of preinvasive prostate cancer have been described in the past decade. Differences among models account for their variable applicability for answering specific research questions. The dog is the only known nonhuman spontaneous animal model of prostate cancer, but numerous transgenic and therapy induced models have been generated in mice and rats. This report summarizes existing models of prostatic intraepithelial neoplasia and their relevance for the study of human prostate cancer. PMID- 10861749 TI - The Dunning model. PMID- 10861750 TI - Thapsigargin induces a calmodulin/calcineurin-dependent apoptotic cascade responsible for the death of prostatic cancer cells. AB - BACKGROUND: New agents are required for the treatment of androgen-independent prostate cancer. Due to the low rate of proliferation of these malignant cells, agents which can activate the apoptotic death of these cells without requiring the cells being in the proliferative cell cycle are critically required. Thapsigargin (TG), via its ability to perturb intracellular free calcium [Ca(2+)](i), is such a cell proliferation-independent cytotoxic agent. The present study focuses on more completely describing the biochemical cascade during the apoptotic death of androgen-independent prostate cancer cells induced by TG and on the mechanistic requirements for this death. METHODS: A variety of cell and molecular biology techniques (e.g., time-lapse video, fluorescence image analysis, Northern and Western blotting) were used to examine the temporal relationship between changes in [Ca(2+)](i), GADD 153 transcription, translocation of the NFATc transcription factor to the nucleus, translocation of BAD from the cytosol to the mitochondria, caspase 9 activation, DNA fragmentation, and the loss of clonogenic survival induced by TG treatment of both human TSU-prl and rat AT3.1 prostate cancer cells in vitro. Additional studies using both microinjection of inhibitors of calmodulin and DNA transfections to induce expression of Ca(2+) binding proteins, e.g., calbindin, were performed to evaluate the causal relationship between [Ca(2+)](i) elevation, calmodulin/calcineurin activation, and apoptosis of prostate cancer cells. RESULTS: Using simultaneous fluorescence ratiometric and phase contrast image analysis in individual cells followed longitudinally for several days, it was documented that TG induced early (1-12 hr) moderate (i.e., <500 nM) elevation in [Ca(2+)](i). During this early rise in [Ca(2+)](i), genes like GADD 153 are induced at the transcriptional level. This early rise is followed by a return of [Ca(2+)](i) to baseline (i.e., approximately 50 nM) before the induction of a delayed (i.e., >12 hr) secondary rise ( approximately 10 microM) in [Ca(2+)](i). During the secondary rise in [Ca(2+)](i), Ca(2+) binds to calcineurin and calmodulin, allowing these proteins to form a complex which activates calcineurin's latent phosphatase activity. Once activated, calcineurin dephosphorylates NFATc and BAD, allowing translocation of these proteins to the nucleus and mitochondria, respectively. BAD translocation induces the release of cytochrome C from the mitochondria into the cytoplasm, which results in activation of caspase 9 and DNA fragmentation. If the TG-induced rise in [Ca(2+)](i) is blocked by overexpressing calbindin, or if calmodulin function is inhibited, these apoptotic events are prevented. CONCLUSIONS: TG induces the apoptotic death of prostate cancer cells via the activation of a reversible signaling phase induced by a transient nanomolar rise in [Ca(2+)](i), which involves new gene transcription and translation. This reversible signaling phase is followed by an irreversible commitment to undergo the execution phase which is induced by a secondary micromolar rise in [Ca(2+)](i). This secondary [Ca(2+)](i) rise irreversibly commits the cell to a calmodulin/calcineurin-dependent cascade, which results in DNA and cellular fragmentation into apoptotic bodies. PMID- 10861751 TI - Role of procathepsin D activation peptide in prostate cancer growth. AB - BACKGROUND: Enzymatically inactive procathepsin D secreted from cancer cells has been confirmed to play a role in breast cancer development. We focused on prostate cancer and the role of activation peptide in mitogenic activity. METHODS: Synthetic peptides and monoclonal antibodies raised against individual fragments of activation peptide were employed. Cell proliferation was measured by MTT (3-[4,5-dimethylthiatol-2-yl]-2,5-diphenyl tetrazolium bromide) assay or by in vivo growth in nude mice. RESULTS: We demonstrated that the growth factor activity of activation peptide is localized in amino-acid region 27-44. In addition, both anti-activation peptide and anti-27-44 peptide antibodies administered in vivo inhibited the growth of human prostate tumors in mice. CONCLUSIONS: Based on these data, we hypothesize that the interaction of procathepsin D activation peptide with an unknown receptor is mediated by amino acid sequence 27-44. This interaction leads in certain types of tumor to a proliferation and higher motility. Blocking of this interaction by antibodies or antagonists might be a valuable tool in prostate cancer inhibition. PMID- 10861752 TI - Estradiol causes a dose-dependent stimulation of prostate growth in castrated beagle dogs. AB - BACKGROUND: Previous studies have shown that chronic treatment of castrate dogs with androgen and estrogen results in significant prostate growth. Estrogen treatment of castrate dogs in the absence of androgen has resulted in conflicting data as reported by several authors. The purpose of this experiment was to evaluate the effect of a physiological dose of estradiol on prostate growth in dogs, using ultrasound to study size changes over time. METHODS: Dogs (n = 25) were randomly divided into groups (n = 5) and treated as follows: castration alone (CC), castration plus low dose estradiol (E(2) low), castration plus high estradiol (E(2) high), castration plus estradiol and androstanediol (E(2)A), or no treatment (normal controls, NC). Silastic implants containing 5alpha-androstan 3alpha-17beta-diol (3alphadiol), and/or 17beta-estradiol were used for continous delivery of steroids. Prostate volume was measured by transrectal ultrasonography, and blood was drawn for hormone and sex hormone binding globulin (SHBG) determinations. RESULTS: Results show that serum estradiol and SHBG levels were fairly constant over 12 weeks in all groups. Estradiol-treated groups had mean serum estradiol values of approximately 40 and 60 pg/ml, respectively. Initially, all groups had similar prostate volumes. Over 12 weeks the castrate dogs had a decline in prostate volume, whereas the intact dogs and those treated with E(2) and 3alpha-diol maintained a constant prostate volume. Estradiol treatment caused a large, late onset (week 7), dose-dependent increase in prostate volume relative to the intact group (P < 0.01). At 12 weeks, animals were euthanized and prostates weighed. The mean prostate weights in each group were: NC 14.8 +/- 2. 9, CC 2.4 +/- 0.5, E(2)A 9.7 +/- 2.0, E(2) low 21.7 +/- 4.3, and E(2) high 63.6 +/- 12.6 g (geometric mean +/- SEM). Histologically, prostates of estrogen-treated dogs showed metaplastic squamous epithelium. CONCLUSIONS: These results demonstrate that estradiol causes marked dose-dependent stimulation of prostate growth in the castrate dog. PMID- 10861753 TI - Expression of proinflammatory genes during estrogen-induced inflammation of the rat prostate. AB - BACKGROUND: Exposure of male Wistar rats to estradiol-17beta (E(2)) in the presence or absence of dihydrotestosterone propionate (DHT) was previously shown to result in prostate inflammation. The present study examines, for the first time, changes in the expression level of several proinflammatory genes during the course of this experimentally induced prostatitis. METHODS: Adult male Wistar rats were given chronic exposure to E(2) + DHT by capsule implantation or were injected with E(2) for short-term exposure. Semiquantitative RT-PCR was employed to measure changes in proinflammatory transcript levels in the separated lobes of the prostate after various times of exposure to estrogen with or without DHT. RESULTS: We observed an upregulation of IL-1beta, IL-6, MIP-2, and inducible nitric oxide synthase (iNOS) after only 4 days treatment with E(2). After 4 weeks of treatment with E(2) + DHT, a significant increase in transcript levels of IL 4, IL-5, IL-6, MIP-2, eotaxin, and iNOS was detected, while IL-1beta and TNF alpha transcript levels only increased slightly. No increase in transcript levels for cyclooxygenase-2 (cox-2), IFN-gamma, IL-2, or IL-12 was observed. CONCLUSIONS: Upregulation of proinflammatory transcripts occurred shortly after exposure to E(2) and well before any inflammatory cells were observed in the prostate. The pattern of gene expression resembled a T(H)2-type helper-cell response. PMID- 10861754 TI - Pharmacodynamic model of testosterone suppression after intramuscular depot estrogen therapy in prostate cancer. AB - BACKGROUND: A long-acting parenteral depot estrogen, polyestradiol phosphate (PEP), which has been in clinical use for several years in combination therapy, has been reevaluated pharmacokinetically and clinically as a single treatment. The present report describes a model predicting the effect on testosterone flux achieved with this estrogen drug. METHODS: Data on serum levels of estradiol and testosterone from a single-dose study, in prostate cancer patients as well as data from injections of 240 or 320 mg PEP each fourth week, were used for pharmacokinetic/dynamic modeling. RESULTS: Serum concentrations of estradiol were governed by a flip-flop mechanism when administered as PEP. An indirect-response model fitted to individual data showed a value of about 500 pmol estradiol/l serum to get a 50% suppression of serum testosterone concentrations. CONCLUSIONS: This model could successfully predict the serum levels of estradiol and testosterone after repeated injections at different doses and was also used to simulate the testosterone suppressing effect of a new dose regimen. PMID- 10861755 TI - Correlation of increased apoptosis and proliferation with development of prostatic intraepithelial neoplasia (PIN) in ventral prostate of the Noble rat. AB - BACKGROUND: Imbalance between cell proliferation and cell apoptosis has been considered a key factor in carcinogenesis. Prostatic intraepithelial neoplasia (PIN) is the most likely precancereous lesion and represents the major target for chemoprevention of prostate cancer. The proliferative and apoptotic activities involved in the development of PIN remain to be elucidated. METHODS: Ventral prostates were removed from Noble rats that were treated with a combination of testosterone (T) and estradiol (E(2)) for certain periods of time, and processed for histopathological grading. To evaluate the relationship between cell proliferation and apoptosis, immunohistochemistry for proliferating cell nuclear antigen (PCNA), Ki-67, and in situ DNA nick labeling (TUNEL) for identifying apoptotic cells, were performed on paraffin sections from prostate samples with PIN lesions. The results were correlated with expression patterns of Bcl-2 and Bax, two proteins related to cell survival and cell apoptosis. RESULTS: Pathologically, low-grade PIN (LGPIN) and high-grade PIN (HGPIN) were observed in ducts or alveoli after 3 and 5 months of T + E(2) treatment, respectively. Quantitative evaluation of Ki-67 showed an increased proliferative activity in HGPIN. In contrast to normal prostatic ducts and alveoli, which showed no positive staining for Ki-67 in the nuclei of luminal cells, 25% Ki-67-positive cells were detected in luminal cells of HGPIN. Only 7.5% Ki-67-positive cells were found belonging to the basal cell type. The Ki-67 index showed a higher growth rate from normal to HGPIN. The PCNA results showed a similar expression pattern to that of Ki-67 in normal prostate, LGPIN, and HGPIN. Apoptotic index (number of apoptotic cells/total number of cell counted) was significantly higher (P = 0.028) in HGPIN (3.23%) than in control prostate (1.19%). In contrast to control prostate, which showed no definite expression of Bcl-2, an intense positive expression of Bcl-2 in HGPIN was observed. Positive expression of Bax protein was observed in glandular epithelial cells of normal control prostate and HGPIN as well. CONCLUSIONS: Overexpression of Bcl-2 and higher Ki-67 or PCNA indices in HGPIN suggest that abnormal growth of premalignant lesions might result from an increase in cell proliferation. An increased apoptotic rate in HGPIN further implicates that active apoptosis may accelerate cell turnover in the development of premalignant lesions of the prostate. PMID- 10861756 TI - Cell kinetics and differentiation after hormonal-induced prostatic hyperplasia in the dog. AB - BACKGROUND: Our aim was to characterize the immunophenotypical changes in canine prostate epithelium after hormonal-induced benign prostatic hyperplasia (BPH). METHODS: Castrated dogs (aged 1-2 and 9-12 years) were treated with vehicle (group C), androstanediol (group A), or androstanediol plus estradiol (group AE). Surgical prostate biopsies were obtained before and after castration and after hormonal treatment. Tissue sections were stained using antibodies specific for basal cells (34betaE12), transiently proliferating (TP)/amplifying cells (RCK103), and luminal exocrine cells (RGE53). RESULTS: Castration resulted in a marked reduction in specific immunoreactivity associated with luminal secretory cells and basal cells in young dogs. In older dogs the number of basal cells remained constant. Hormonal treatment (AE) resulted in an increased number of cells with an immunophenotype that was associated with the TP/amplifying cell compartment and hyperplastic luminal epithelium. CONCLUSIONS: The relative increase in TP/amplifying cells in hormonally induced BPH in the dog is in line with a stem-cell-derived proliferation. Moreover, the finding of androgen independent basal cells in the prostate of older dogs may contribute to the enhanced risk of development of BPH with increasing age. PMID- 10861757 TI - PSA is a candidate self-antigen in autoimmune chronic prostatitis/chronic pelvic pain syndrome. AB - BACKGROUND: Previous studies demonstrated that recognition of seminal plasma antigens can occur in patients with chronic prostatitis/chronic pelvic pain syndrome. This suggests that an autoimmune component may contribute to symptoms in some men. To determine if any of the principal secretory proteins of the prostate could be candidate antigens in autoimmune prostatitis, we examined the recall proliferative response of purified CD4 T cells in patients with chronic prostatitis and in normal volunteers using purified seminal plasma antigens and autologous dendritic cells. METHODS: Peripheral blood mononuclear cells were harvested from 14 patients with chronic prostatitis and 12 normal volunteers by density gradient centrifugation. The stimulating cells were irradiated autologous dendritic cells produced by culture of monocyte-enriched fractions with IL-4 and Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF). Purified CD4 T cells were the responding population. Recall proliferation assays were performed, using purified seminal plasma proteins as antigens. RESULTS: In 14 patients with chronic prostatitis, we detected a greater than 2-fold increase in proliferative response to PSA compared to control in 5 patients (36%). No response to Prostatic Acid Phosphatase (PAP) or beta-microseminoprotein was observed in these 14 patients. In 12 normal volunteer donors with no history of genitourinary disease or symptoms, no proliferative response above background was observed for any prostatic antigen. CONCLUSIONS: The data suggest that some men with symptoms of chronic prostatitis have evidence of a proliferative CD4 T-cell response to PSA. PSA is a candidate antigen in chronic prostatitis/chronic pelvic pain syndrome and may be an appropriate target for immunotherapy for prostatic cancer. PMID- 10861758 TI - Suramin potently inhibits the enzymatic activity of PSM. AB - BACKGROUND: The polysulfonated napthlyurea suramin has shown significant antitumor activity in patients with hormone-refractory metastatic prostate cancer. The mechanism by which suramin exerts this effect is unknown. In 1993, prostate-specific membrane antigen (PSM) was identified as a prostate biomarker that is elevated in hormone-refractory and metastatic prostate cancer. PSM is a glutamate exocarboxypeptidase capable of cleaving the terminal alpha-linked glutamate from the dipeptide N-acetyl-aspartyl-glutamate (NAAG) and the gamma linked glutamates from folate polyglutamate. METHODS: Using a NAAG hydrolytic radioenzymatic assay, we tested whether suramin had any effect on the enzymatic activity of PSM. RESULTS: We demonstrate that suramin potently inhibits the enzymatic activity of PSM with a K(i) = 15 nM and 68 nM for the membrane associated and soluble forms of PSM, respectively. In addition, we show that suramin inhibition of PSM enzyme activity displays the kinetics of a classic competitive inhibitor. CONCLUSIONS: This is one of the most potent activities described for suramin to date and may represent a portion of its pharmacologic and/or toxicological mechanism of action. PMID- 10861759 TI - Effects of voltage-gated ion channel modulators on rat prostatic cancer cell proliferation: comparison of strongly and weakly metastatic cell lines. AB - BACKGROUND: The strongly metastatic MAT-LyLu and the weakly metastatic AT-2 rat prostatic cancer cell lines have been shown to express voltage-gated ion channels differentially. In the present study, the possible contribution of voltage-gated ion channel activity to the proliferation of these cell lines was investigated, in a comparative approach. METHODS: Several voltage-gated ion channel modulators were tested for their effects on proliferation over 54 hr, using an in vitro assay. The modes of action of the chemicals were monitored by electrophysiological (patch-clamp) recording. RESULTS: The voltage-gated K(+) channel blockers 4-aminopyridine (4-AP; 2 mM), margatoxin (5 nM), charybdotoxin (4.5 nM), and verapamil (50 microM) inhibited the K(+) channels of both cell lines by between 38-65% and reduced the proliferation of the AT-2 cell line, in a dose-dependent manner, by 8-51%. However, only 4-AP reduced proliferation of the MAT-LyLu cell line. Tetrodotoxin (6 microM) blocked completely the voltage-gated Na(+) channel expressed selectively in the MAT-LyLu cell line, but had no effect on the proliferation of either cell line. On the other hand, the presumed Na(+) channel "opener" veratridine (10-50 microM) reduced significantly, in a dose dependent manner, the proliferation of both cell lines by up to approximately 30%. CONCLUSIONS: We conclude that the mechanism(s) controlling the proliferation of the weakly metastatic AT-2 cells involves voltage-gated K(+) channels. In contrast, the proliferation of strongly metastatic MAT-LyLu cells is much less dependent upon voltage-gated K(+) channel activity. PMID- 10861760 TI - Endothelium-derived factors as paracrine mediators of prostate cancer progression. AB - BACKGROUND: Vascular endothelium represents a complex network of cells producing a large number of active substrates affecting physiologic, metabolic, and immunologic properties of the whole organism, as well as particular organs or tissues. The potential influence of endothelium-derived paracrine factors on prostate cancer progression has only begun to be examined. METHODS: This review summarizes recent literature on endothelium-derived factors, including vasoactive agents, peptide growth factors, cytokines, and colony-stimulating factors, involved in the development and progression of prostate cancer. RESULTS: Endothelial cells produce an array of active substrates, many of which have been shown to influence prostate cancer growth. Available data demonstrate the positive impact of such molecules as endothelin-1, basic FGF, TGF-beta, IL-6, and IL-8 on prostate cancer progression. Many other endothelium-derived factors NO, IGF, PDGF, IL-1, G-CSF, and GM-CSF (Nitric Oxide, Insulin-Like Growth Factor, Platelet-Derived Growth Factor, Interleukin-1, Granulocyte Colony Stimulating Factor, and Granulocyte-Macrophage Colony Stimulating Factor) are, at best, implicated in prostate cancer growth, and in most cases support cancer progression. CONCLUSIONS: A better understanding of endothelium-derived factors, as paracrine mediators of prostate carcinogenesis and progression, should aid in the development of novel therapeutic strategies. PMID- 10861761 TI - Leukemia after exposure to benzene: temporal trends and implications for standards. AB - BACKGROUND: Benzene is a human leukemogen. Risk assessment, and the setting of occupational and environmental standards, has assumed that risk is constant in time after a unit of exposure. Leukemia risk is known to vary with time after exposure to ionizing radiation. METHODS: A matched case-control study of leukemia risk in relation to the temporal pattern of benzene exposures was performed using data from the National Institute of Occupational Safety and Health. RESULTS: Leukemia risk following exposure to benzene varied with time in a manner similar to that following exposure to ionizing radiation. More recent exposures were more strongly associated with risk than were more distant ones. There was no significant relation between leukemia death and benzene exposures incurred more than 20 years previously. CONCLUSIONS: Recent analyses of specific occupational and environmental carcinogens, including benzene and radon, have indicated that cancer risk tends to decline as the time from exposure increases. This suggests that standards for the control of occupational or public risk must be selected to control exposures over a narrower time frame than the usual lifetime one. In the case of benzene, it would appear that risk is attributable primarily to exposures incurred during the previous 10 to 20 years, with exposures in the most recent 10 years being the most potent. To limit risk, exposures must be controlled during that interval. It is important that epidemiologists explore the temporal pattern of risk in their studies to facilitate the risk assessment of other carcinogens. PMID- 10861762 TI - Silica, silicosis, and lung cancer: a risk assessment. AB - BACKGROUND: To investigate exposure-response relationships for silica, silicosis, and lung cancer. METHODS: Quantitative review of the literature identified in a computerized literature search. RESULTS: The risk of silicosis (ILO category 1/1 or more) following a lifetime of exposure at the current OSHA standard of 0.1 mg/m(3) is likely to be at least 5-10% and lung cancer risk is likely to be increased by 30% or more. The exposure-response relation for silicosis is nonlinear and reduction of dust exposures would have a greater than linear benefit in terms of risk reduction. Available data suggests that 30 years exposure at 0.1 mg/m(3) might lead to a lifetime silicosis risk of about 25%, whereas reduction of the exposure to 0.05 mg/m(3) might reduce the risk to under 5%. CONCLUSIONS: The lifetime risk of silicosis and lung cancer at an exposure level of 0.1 mg/m(3) is high. Lowering exposures to the NIOSH recommended limit if 0.05 mg/m(3) may have substantial benefit. PMID- 10861763 TI - Case-control assessment of the association between non-Hodgkin's lymphoma and occupational radiation with doses assessed using a job exposure matrix. AB - BACKGROUND: Epidemiologic data for an association between radiation exposure and non-Hodgkin's lymphoma (NHL) have been inconclusive though the strongest evidence has been provided by studies of patients treated with radiotherapy. METHODS: We evaluated the association between occupational radiation exposure and non Hodgkin's lymphoma in men using a population-based case-control study with 1,056 case and 1,860 control subjects sampled from eight geographic areas in the United States. Because dosimetry data were not available, doses were estimated for individuals who reported occupational radiation exposure using a radiation job exposure matrix developed for this purpose. Conditional logistic regression was used to model the association between reported occupational radiation exposure and NHL incidence. RESULTS: We found that most men (> 90%) did not report exposure to occupational sources of radiation. Among those who reported exposure, estimated cumulative doses were low, with an estimated mean of less than 0.02 Gray and a maximum of 0.12 Gray. The risk for NHL was not associated with ever having reported an occupational radiation exposure (OR = 0.90, 95% CI = 0.74 1.10) nor was there evidence of a dose-response relationship between risk and either the estimated cumulative doses or duration of exposure. CONCLUSIONS: The findings in this study are consistent with results from most current research on occupational radiation and NHL risk that have found no increased risk of NHL at low levels of occupational radiation exposure. While it should be noted that exposure misclassification likely biased our results toward the null, this large population-based case-control study adds to existing evidence which suggests that there is little to no increased risk for NHL associated with exposure to low levels of radiation such as that commonly found in many occupational settings. PMID- 10861764 TI - A retrospective job exposure matrix for estimating exposure to 2,3,7, 8 tetrachlorodibenzo-p-dioxin. AB - BACKGROUND: A job exposure matrix was developed to estimate the 2,3, 7,8 tetrachlorodibenzo-p-dioxin exposure of 3,538 workers who produced 2,4,5 trichlorophenol and its derivatives. METHODS: Daily TCDD exposure scores that were plant, process, and period specific were estimated for each job title as the product of 1) the concentration of TCDD (microg/g); 2) a qualitative factor to account for the extent of worker contact and 3) time exposed to TCDD contamination. Daily scores were summed to compute individual cumulative TCDD exposure scores. RESULTS: Daily TCDD exposure scores ranged from 0.001 to 1,250. Cumulative TCDD scores ranged from 0.002 to 1,559,430. The 393 workers with records of chloracne in the TCDD exposure cohort (11%) had markedly higher cumulative scores than those with no record of chloracne (a median score of 11,546 vs. 77). CONCLUSIONS: The cumulative TCDD exposure scores incorporate both duration and level of exposure, and permit the relative ranking of worker exposures for the evaluation of exposure-response relationships between TCDD exposure and mortality in an updated cohort study analysis. PMID- 10861765 TI - Slip and fall-related injuries in relation to environmental cold and work location in above-ground coal mining operations. AB - BACKGROUND: The association between slip and fall-related injuries and environmental temperature was examined for mostly enclosed (inside vehicles, machinery, or buildings), outdoor (outside, not enclosed), and enclosed/outdoor jobs in the coal mining industry to see if differences existed among the three work locations that had varying exposure to cold temperatures. METHODS: Temperature data from the National Climatic Data Center and injury data from the Mine Safety and Health Administration were evaluated from 1985-1990 for seven states. Proportionate methods were used to examine the relationship between slips and falls and temperature. RESULTS: Proportionate injury ratios of slips and fall related injuries increased as temperature declined for all three work locations. Proportion of slips and fall-related injuries that occurred while running/walking increased with declining temperature, with the ground outside as the most common source of these injuries. CONCLUSIONS: Outside movement becomes a greater hazard at freezing temperatures for workers in all locations, not just outdoor workers. Any intervention methods geared toward reducing injury incidents facilitated by cold weather must also be directed toward workers who spend time in more enclosed locations. PMID- 10861766 TI - Occupational injuries in the mining industry and their association with statewide cold ambient temperatures in the USA. AB - BACKGROUND: Relatively few occupational epidemiological studies have been conducted concerning the association between cold ambient temperatures and cold exposure injuries, and fewer still of traumatic occupational injuries and cold ambient temperatures. METHODS: The association of ambient temperature and wind data from the National Climatic Data Center with injury data from mines reported to the Mine Safety and Health Administration (MSHA) was evaluated over a 6 year period from 1985-1990; 72,716 injuries from the seven states with the most numerous injuries were included. Temperature and wind data from each state's metropolitan weather stations were averaged for each day of the 6 year period. A weighted linear regression tested the relationship of ungrouped daily temperature and injury rate for all injury classes. For cold exposure injuries and fall injuries, relative incidence rates for grouped temperature data were fit with Poisson regression. RESULTS: As temperatures decreased, injury rates increased for both cold exposure injuries and slip and fall injuries. The association of slip and fall injuries with temperature was inverse but not strictly linear. The strongest association appeared with temperatures 29 degrees F and below. The injury rates for other accident categories increased with increasing ambient temperatures. CONCLUSIONS: This study suggests that statewide average ambient temperature reflects the expected association between the thermal environment and cold exposure injuries for workers, but more importantly, documents an association between ambient temperatures and occupational slip and fall injuries. PMID- 10861767 TI - Motor vehicle manufacturing and prostate cancer. AB - BACKGROUND: The purpose of this investigation was to evaluate the relation between employment in motor vehicle manufacturing (MVM) and fatal prostate cancer. METHODS: The study included 322 prostate cancer deaths occurring in 1973 through 1987 and 1,285 controls, selected from a cohort of 126,100 male MVM workers. RESULTS: Men employed in casting operations had an odds ratio of 1.5 (95% CI = 1. 1-2.0). The association was consistent across casting facilities and was attributable primarily to work in core and mold making (OR = 1.5, 95% CI = 1.1-2.2) and metal melting and pouring jobs (OR = 1.9, 95% CI = 1.0-3.6). Other results included ORs of 1.9 (95% CI = 1.0-3.7) for warehousing and distribution operations and 2.1 (95% CI = 1.2-3. 7) for electric and electronic equipment manufacturing. The latter two associations exhibited little internal consistency. CONCLUSIONS: The relationships seen in this study were weak and may have been due to chance. Core and mold making and metal melting and pouring foundry operations entail potential exposure to metal dusts and fumes, to polycyclic aromatic hydrocarbons (PAHs), and to other chemicals. However, associations between these exposures and prostate cancer have not been reported consistently, nor have other studies of foundry workers consistently noted an excess of prostate cancer. PMID- 10861768 TI - Safety awareness among New York farmers. AB - BACKGROUND: This study was conducted to assess the health status and safety practices among year-round adult farm workers and residents and included a telephone interview survey of 1,727 persons from 552 farms. METHODS: Logistic regression was used to analyze four safety questions. RESULTS: Among 541 farm owner/operators significant predictors of making substitutions in the use of chemicals and major changes to equipment include younger age, more persons assisting on the farm, and higher gross sales. Having training is associated with having more than a high school education. Among all participants the perception that personal protective equipment are useful is associated with being younger, male, an owner/operator or worker, and having at least a high school education. CONCLUSIONS: These findings suggest that older and less educated farmers should be targeted for health and safety programs. PMID- 10861769 TI - Incidence of unintentional injuries in farming based on one year of weekly registration in Danish farms. AB - BACKGROUND: In Denmark, farming ranks as the industry with the highest incidence rate of fatal injuries. For nonfatal injuries, insufficient registration practices prevent valid comparisons between occupations. This study examines the occurrence of farm accidents and injuries, as well as work-specific factors, via weekly registration in a representative sample of 393 farms in one county during 1 year. METHODS: From a random sample of 794 farms, (10% of farms in the county of Ringkoebing, Denmark) 393 farms with 1,597 residents and employees participated in a 1-year self-registration of work-related unintentional incidents. The procedure included a detailed registration of hours spent on all main working tasks. Weekly recording of incident occurrence or nonoccurrence resulted in the completion of 19,782 registration forms. Three months after incident occurrence, a telephone interview was conducted about the related work situation and resulting injuries. RESULTS: During the 12-month period, 479 occupational accidents were reported, of which 389 resulted in an injury. The absolute number of injuries increased with number of work hours, but there was no relative increase of incidence by work hours. Persons below the age of 50 had slightly less than a doubled risk compared with those over 50 years of age. No other marked, reliable age effect was found. There was, however, a seasonal variation, with summer and autumn having a double relative incidence compared with winter and spring. Among farm owners, 35% experienced at least one injury per year, while this was the case for 17% of farm laborers. When adjusting for work hours, the increased frequency of injuries among farm owners was reduced to a factor of 1.5. Animal-related work was the most common injury mechanism. Repair and maintenance work was found to be the most dangerous task relative to the number of task-specific work hours. Subgroups of tasks with a markedly increased injury rate were moving animals within the farm, veterinary procedures, and repair of field machinery and stable equipment. CONCLUSIONS: Farm injuries occur among 32% of full-time farmers and farm laborers each year. A quarter of these require professional treatment. This area calls for preventive action. PMID- 10861770 TI - Occupational bladder cancer mortality among racial and ethnic minorities in 21 states. AB - BACKGROUND: Occupational bladder cancer mortality among minority racial/ethnic groups is not well described compared to occupational bladder cancer mortality among non-minority males in the United States. METHODS: Race/ethnicity- and sex specific bladder cancer mortality (1985-1992) of workers employed in 21 states was examined using a proportionate mortality study design. Mortality of specific racial/ethnic/occupational groups was compared separately with workers in the specific occupation and with members of the specific racial/ethnic group. RESULTS: This study identified elevated bladder cancer mortality among African American males and females and Latino males in several occupational groups with exposure to suspected bladder carcinogens as well as among Asian males in sales (PMR = 2. 13) and Asian females in the personal services industry (PMR = 5.25; CI: 1.64-16.75). CONCLUSIONS: Surveillance of occupational cancer risks among racial/ethnic minorities using regularly available death certificate data is facilitated when states code both usual occupation/industry and race/ethnicity. PMID- 10861771 TI - The prevalence of sensorineural symptoms attributable to hand-transmitted vibration in Great Britain: a national postal survey. AB - BACKGROUND: Exposure to hand-transmitted vibration (HTV) can cause sensorineural symptoms in the upper limb, but its impact has not previously been assessed in the general population. METHODS: To investigate, we mailed a questionnaire about exposures to HTV, finger blanching and sensory symptoms (numbness or tingling) in the upper limbs to a population sample comprising 21,201 working-aged men and women selected at random from the age-sex registers of 34 British general practices, and a further 993 randomly selected from the pay records of the armed services. Associations were explored using multiple logistic regression models to adjust for confounding, with the resultant odds ratios converted into prevalence rate ratios (PRs). RESULTS: Of 12,907 respondents, 2,607 (20.2%) reported sensory symptoms in the upper limb during the past week. Sensory symptoms were more prevalent in those with blanching, and were commonly associated with exposure to HTV, especially in men. In comparison with men who had never been exposed to HTV, the PR in men exposed both at work and in leisure was 2.2 (95% CI 1.9-2.4). Associations were found even in those who had never blanched. CONCLUSIONS: Sensorineural symptoms in the upper limbs are common. HTV is an important risk factor for such complaints in the general population. PMID- 10861772 TI - A study of post-traumatic shingles as a work related injury. AB - BACKGROUND: After chicken pox, the herpes varicella-zoster (HVZ) virus may remain dormant in the dorsal root ganglion until later reactivation causes shingles, characterized by painful dysesthesias and cutaneous vesicular eruptions along a unilateral dermatome. Shingles as a work-related injury has not been previously addressed in the medical literature. Case History We present a 50-year old female hospital employee who, while working, sustained an acute, traumatic hyperextension injury to her right wrist, hand, and fingers. Although she initially responded to treatment for flexor tendinitis, she suddenly developed shingles in the right C5-C6 dermatomes. She was treated with famcyclovir and her skin lesions resolved, but post-herpetic neuralgia persisted. CONCLUSIONS: It was felt that her shingles was causally related to her occupational injury since trauma (previously reported to precipitate shingles) was her only risk factor and the timing and location of the lesions corresponded closely to the occupational injury. In addition to appropriately diagnosing and treating their patients, workers' compensation physicians often must determine if a particular condition was caused by the original work-related incident. Clinicians who treat trauma patients and injured workers should be aware of post-traumatic shingles and understand the causal relationship of this uncommon but clinically important phenomenon. PMID- 10861773 TI - Heterogeneity and statistical significance in meta-analysis: an empirical study of 125 meta-analyses. AB - For meta-analysis, substantial uncertainty remains about the most appropriate statistical methods for combining the results of separate trials. An important issue for meta-analysis is how to incorporate heterogeneity, defined as variation among the results of individual trials beyond that expected from chance, into summary estimates of treatment effect. Another consideration is which 'metric' to use to measure treatment effect; for trials with binary outcomes, there are several possible metrics, including the odds ratio (a relative measure) and risk difference (an absolute measure). To examine empirically how assessment of treatment effect and heterogeneity may differ when different methods are utilized, we studied 125 meta-analyses representative of those performed by clinical investigators. There was no meta-analysis in which the summary risk difference and odds ratio were discrepant to the extent that one indicated significant benefit while the other indicated significant harm. Further, for most meta-analyses, summary odds ratios and risk differences agreed in statistical significance, leading to similar conclusions about whether treatments affected outcome. Heterogeneity was common regardless of whether treatment effects were measured by odds ratios or risk differences. However, risk differences usually displayed more heterogeneity than odds ratios. Random effects estimates, which incorporate heterogeneity, tended to be less precisely estimated than fixed effects estimates. We present two exceptions to these observations, which derive from the weights assigned to individual trial estimates. We discuss the implications of these findings for selection of a metric for meta-analysis and incorporation of heterogeneity into summary estimates. Published in 2000 by John Wiley & Sons, Ltd. PMID- 10861774 TI - Cumulative cause-specific mortality for cancer patients in the presence of other causes: a crude analogue of relative survival. AB - A common population-based cancer progress measure for net survival (survival in the absence of other causes) of cancer patients is relative survival. Relative survival is defined as the ratio of a population of observed survivors in a cohort of cancer patients to the proportion of expected survivors in a comparable set of cancer-free individuals in the general public, thus giving a measure of excess mortality due to cancer. Relative survival was originally designed to address the question of whether or not there is evidence that patients have been cured. It has proven to be a useful survival measure in several areas, including the evaluation of cancer control efforts and the application of cure models. However, it is not representative of the actual survival patterns observed in a cohort of cancer patients. This paper suggests a measure for cumulative crude (in the presence of other causes) cause-specific probability of death for a population diagnosed with cancer. The measure does not use cause of death information which can be unreliable for population cancer registries. Point estimates and variances are derived for crude cause-specific probability of death using relative survival instead of cause of death information. Examples are given for men diagnosed with localized prostate cancer over the age of 70 and women diagnosed with regional breast cancer using Surveillance, Epidemiology and End Results (SEER) Program data. The examples emphasize the differences in crude and net mortality measures and suggest areas where a crude measure is more informative. Estimates of this type are especially important for older patients as new screening modalities detect cancers earlier and choice of treatment or even 'watchful waiting' become viable options. Published in 2000 by John Wiley & Sons, Ltd. PMID- 10861775 TI - Comparison of different approaches to incidence prediction based on simple interpolation techniques. AB - The paper compares three different methods for performing disease incidence prediction based on simple interpolation techniques. The first method assumes that the age-period specific numbers of observed cases follow a Poisson distribution and the other two methods assume a normal distribution for the incidence rates. The main emphasis of the paper is on assessing the reliability of the three methods. For this purpose, ex post predictions produced by each method are checked for different cancer sites using data from the Cancer Control Region of Turku in Finland. In addition, the behaviour of the estimators of predicted expected values and prediction intervals, crucial for investigation of the reliability of prediction, are assessed using a simulation study. The prediction method making use of the Poisson assumption appeared to be the most reliable of the three approaches. The simulation study found that the estimator of the length of the prediction interval produced by this method has the smallest coverage error and is the most precise. PMID- 10861776 TI - On the inclusion of prevalent cases in HIV/AIDS natural history studies through a marker-based estimate of time since seroconversion. AB - In most cohort studies of HIV infection and AIDS, seroprevalent cases provide a substantial amount of information. Inclusion of these people in natural history studies requires a fairly unbiased method to estimate their seroconversion distribution. When a cohort-based estimate is not feasible, an alternative is to estimate individual seroconversion distributions, based on marker values at entry. In this paper, a non-parametric marker-based estimation method is developed. The method is applied to data from the Amsterdam cohort study on homosexual men. For seroprevalent cases who entered the study between October 1984 and April 1985, individual seroconversion distributions are estimated based on their first measured CD4 count. In subsequent survival analyses, dates of seroconversion are estimated via conditional mean imputation. Inclusion of these seroprevalent cases greatly improves the quality of the data. Age at seroconversion is a significant cofactor for disease progression, a result not found when analysis is restricted to those who seroconvert. To incorporate the uncertainty in the imputed date of seroconversion, a bootstrap procedure is developed for the computation of p-values and confidence intervals. In our analyses, standard procedures, which ignore the uncertainty in the imputed date of seroconversion, perform almost as well. PMID- 10861777 TI - Summarizing the predictive power of a generalized linear model. AB - This paper studies summary measures of the predictive power of a generalized linear model, paying special attention to a generalization of the multiple correlation coefficient from ordinary linear regression. The population value is the correlation between the response and its conditional expectation given the predictors, and the sample value is the correlation between the observed response and the model predicted value. We compare four estimators of the measure in terms of bias, mean squared error and behaviour in the presence of overparameterization. The sample estimator and a jack-knife estimator usually behave adequately, but a cross-validation estimator has a large negative bias with large mean squared error. One can use bootstrap methods to construct confidence intervals for the population value of the correlation measure and to estimate the degree to which a model selection procedure may provide an overly optimistic measure of the actual predictive power. PMID- 10861778 TI - The (in)validity of sensitivity and specificity. AB - This paper is a legacy of the first author, and after her untimely death reconstructed by the second author as a tribute to Irene Guggenmoos's contribution to biostatistics. It discusses two different views on diagnostic testing: the classical view in which sensitivity and specificity of a diagnostic test are considered universal constants, and the more statistical point of view that focuses on predictive values. The differences between the two paradigms are outlined and practical examples are discussed to show that the familiar concepts of sensitivity and specificity must be handled with care and not used indiscriminately. PMID- 10861779 TI - Modelling covariance structure in the analysis of repeated measures data. AB - The term 'repeated measures' refers to data with multiple observations on the same sampling unit. In most cases, the multiple observations are taken over time, but they could be over space. It is usually plausible to assume that observations on the same unit are correlated. Hence, statistical analysis of repeated measures data must address the issue of covariation between measures on the same unit. Until recently, analysis techniques available in computer software only offered the user limited and inadequate choices. One choice was to ignore covariance structure and make invalid assumptions. Another was to avoid the covariance structure issue by analysing transformed data or making adjustments to otherwise inadequate analyses. Ignoring covariance structure may result in erroneous inference, and avoiding it may result in inefficient inference. Recently available mixed model methodology permits the covariance structure to be incorporated into the statistical model. The MIXED procedure of the SAS((R)) System provides a rich selection of covariance structures through the RANDOM and REPEATED statements. Modelling the covariance structure is a major hurdle in the use of PROC MIXED. However, once the covariance structure is modelled, inference about fixed effects proceeds essentially as when using PROC GLM. An example from the pharmaceutical industry is used to illustrate how to choose a covariance structure. The example also illustrates the effects of choice of covariance structure on tests and estimates of fixed effects. In many situations, estimates of linear combinations are invariant with respect to covariance structure, yet standard errors of the estimates may still depend on the covariance structure. PMID- 10861780 TI - Tutorial in biostatistics-longitudinal data analysis (repeated measures) in clinical trials. PMID- 10861781 TI - Fluorescent myelin proteins provide new tools to study the myelination process. AB - We present here a new approach which permits us to follow myelin proteins within living, actively myelinating cells. We have developed probes to study the spatial and temporal incorporation of proteins into the myelin sheath by expressing myelin proteins fused to the green fluorescent protein (GFP). GFP from the jellyfish Aequorea victoria and its derivatives, e.g., blue fluorescent protein (BFP) were used as molecular reporters to monitor the intracellular distribution of myelin proteins. Fusion proteins (14 kD myelin basic protein [MBP]-GFP, 21 kD MBP-GFP) were expressed in primary Schwann cells (SCs) and their distribution was monitored by confocal microscopy. The autofluorescent chimeric proteins were readily visualized and their subcellular localization was unaffected by the GFP reporter. However, because of the length of culturing time necessary to establish permanent cell lines, we found that it was not possible to obtain MBP-GFP stable SCs that also were capable of myelinating neuronal axons. We therefore devised a way of introducing vectors under conditions where cells are dividing in response to endogenous stimuli, and therefore are still capable of myelinating. We designed a protocol in which SCs cocultured with dorsal root ganglion (DRG) neurons are transfected while they are actively dividing. SCs transfected in this way exhibit a good level of protein expression and retain their myelinating phenotype. The fusion protein expression lasts long enough to observe "green myelin. " These fluorescently tagged myelin proteins will allow high-resolution examination of the protein and membrane traffic in normal myelinating cells. PMID- 10861782 TI - Acylation of myelin Po protein is required for adhesion. AB - The extracellular domains of myelin Po protein interact homophilically and hence hold myelin compact at the intraperiod line. The cytoplasmic domain of Po, however, can also affect the interactions of its extracellular sequences. Po is acylated, mostly with palmitic acid, at Cys 153, just at the transmembrane:cytoplasmic domain interface. Here we show that Po mutated at Cys 153 to alanine (C153A), is not acylated and is not adhesive. Like wild-type Po, C153A Po clusters within the membrane and seems to interact with the cytoskeleton. On the other hand, the rate of turnover of C153A Po in transfected Chinese hamster ovary cells is almost 4 times faster than wild-type Po. The increased instability of C153A Po compared to wild-type Po may account for its loss of adhesion. PMID- 10861783 TI - Absence of P0 leads to the dysregulation of myelin gene expression and myelin morphogenesis. AB - P0, the major peripheral nervous system (PNS) myelin protein, is a member of the immunoglobulin supergene family of membrane proteins and can mediate homotypic adhesion. P0 is an essential structural component of PNS myelin; mice in which P0 expression has been eliminated by homologous recombination (P0-/-) develop a severe dysmyelinating neuropathy with predominantly uncompacted myelin. Although P0 is thought to play a role in myelin compaction by promoting adhesion between adjacent extracellular myelin wraps, as an adhesion molecule it could also have a regulatory function. Consistent with this hypothesis, Schwann cells in adult P0-/ mice display a novel molecular phenotype: PMP22 expression is down-regulated, MAG and PLP expression are up-regulated, and MBP expression is unchanged. As in quaking viable mutant mice (qk(v)), which have uncompacted myelin morphologically similar to that found in P0-/- mice, neither the qKI-6 or qKI-7 proteins are expressed in P0-/- peripheral nerve. In addition to these changes in gene expression in the P0 knockout, PLP/DM-20 accumulates in the endoplasmic reticulum of P0-/- Schwann cells, whereas MAG accumulates in redundant loops of uncompacted myelin, not at nodes of Ranvier or Schmidt-Lantermann incisures. Taken together, these results demonstrate that P0 is involved, either directly or indirectly, in the regulation of both myelin gene expression and myelin morphogenesis. PMID- 10861784 TI - Neurotrophin-3 (NT-3) diminishes susceptibility of the oligodendroglial lineage to AMPA glutamate receptor-mediated excitotoxicity. AB - Prior reports demonstrated that cells of the oligodendroglial lineage are susceptible to excitotoxic necrosis mediated by alpha-amino-3-hydroxy-5 methylisoxazole-4-propionic acid glutamate receptors (AMPA-GluR), and also showed that these cells express the high affinity neurotrophin receptors, TrkC and TrkA. We now report that: a) oligodendroglial progenitors (OP) and immature oligodendroglia are more vulnerable to AMPA-GluR-mediated excitotoxicity than are mature oligodendroglia; b) TrkC expression falls substantially during differentiation of cultured OP to mature oligodendroglia, whereas TrkA expression increases markedly; and c) neurotrophin-3, and to a lesser extent, nerve growth factor, protect the oligodendroglial lineage against AMPA-GluR-mediated excitotoxicity. PMID- 10861785 TI - sICAM-1 and TNF-alpha induce MIP-2 with distinct kinetics in astrocytes and brain microvascular endothelial cells. AB - The dysfunction of the blood-brain barrier (BBB) occurring after traumatic brain injury (TBI) is mediated by intracerebral neutrophil accumulation, chemokine release (e.g., interleukin (IL)-8) and upregulation of adhesion molecules (e.g., intercellular adhesion molecule (ICAM)-1). In patients with severe TBI, we previously found that elevated cerebrospinal fluid (CSF) IL-8 and soluble (s)ICAM 1 correlate with BBB dysfunction, and this prompted us to concomitantly monitor IL-8, sICAM-1 and their stimulator tumor necrosis factor (TNF)-alpha in CSF. Potential mechanisms for upregulation of the IL-8 analogue, murine macrophage inflammatory protein (MIP)-2, and sICAM-1 at the BBB were studied using cultured mouse astrocytes and brain microvascular endothelial cells (MVEC). In CSF of seven patients, IL-8 and sICAM-1 were elevated for 19 days after severe TBI, whereas TNF-alpha exceeded normal values on 9 days. Stimulation of MVEC and astrocytes with TNF-alpha simultaneously induced the release of MIP-2 reaching saturation by 4-8 hr and of sICAM-1 increasing continuously from 2-4 hr to 12 hr. Augmented sICAM-1 production correlated with enhanced membrane-bound (m)ICAM-1 expression in both cell types (r(s) = 0.96 and 0.90, P < 0.0001), but was markedly higher in astrocytes. The release of sICAM-1 was not influenced by IL-8 or MIP-2, although astrocytes and MVEC expressed the IL-8/MIP-2 receptor (CXCR-2) as determined by FACS analysis. Instead, we found that sICAM-1 strongly induced MIP-2 secretion by both cell types with kinetics differing from those evoked by TNF-alpha. If added together, sICAM-1 and TNF-alpha synergistically induced MIP-2 production suggesting the involvement of two different pathways for MIP-2 regulation. PMID- 10861786 TI - The mechanism of the neurotransmitter release in growth cones. AB - The growth cone is considered the precursor of the presynaptic terminal. To elucidate the minimal molecular machinery required for exocytosis, we examined the characteristics of alpha-latrotoxin-induced exocytosis in growth cones. In isolated growth cones (IGC), neurotransmitters were released in a SNARE-dependent manner, but rab3A cycling was blocked. By supplying rabphilin, a rab3A acceptor found in low levels in IGC, the IGC obtained as high an exocytotic efficiency as adult synaptosomes, and the complete GDP-GTP conversion of rab3A occurred on growth cone vesicles (GCV). GCVs bound SNAREs but not NSF or alpha-SNAP; whereas in the rabphilin-supplied IGC, GCVs recruited both NSF and alpha-SNAP, to form the SNARE-NSF-SNAP complex. These results suggest that rab3A cycling is dependent upon the accumulation of rabphilin and is completed later than the SNARE mechanism, and that rabphilin is involved in determining the efficiency of exocytosis by modifying the SNARE mechanism. PMID- 10861787 TI - Efficient expression of tetracycline-responsive gene after transfection of dentate gyrus neurons in vitro. AB - Gene transfer into neurons both in vivo and in vitro may aid in understanding of gene regulation and function in nerve cells. Especially desirable is ability to control the gene expression. In this study we developed conditions for transfection of hippocampal dentate gyrus neurons in dissociated cultures in vitro by calcium-phosphate method. Furthermore, we describe an effective use of tetracycline responsive gene promoter (Tet-On) system for the controlled and very efficient expression of transfected genes. Under optimal conditions as established in this study, efficiency of transfection of neurons with green fluorescent protein (GFP) driven by constitutive cytomegalovirus (CMV) early promoter reached 2.7%. With tetracycline responsive promoter percentage of GFP positive neurons raised in the presence of tetracycline analog, doxycycline up to 20%. Application of the Tet-On system resulted in almost 10-fold induction of GFP expression. PMID- 10861788 TI - Vasopressin increases [Ca(2+)](i) in differentiated astrocytes by activation of V1b/V3 receptors but has no effect in mature cortical neurons. AB - Vasopressin (AVP) plays an important role in regulation of astrocytic, but not neuronal, water content and cell volume during hydro-osmotic challenge. To investigate the intracellular mechanism(s) signaling this response, [Ca(2+)](i) was measured fluorometrically in cultured cerebrocortical astrocytes and neurons, obtained from neonatal and fetal mouse brains, and matured during the culturing period. In astrocytes, [Ca(2+)](i) increased with an EC(50) of between 10(-10) and 10(-9) M AVP, the maximum increase was approximately 100 nM, and the response was independent of extracellular Ca(2+), identifying the receptor as being of the V1b/V3 subtype. In contrast, AVP had no effect on [Ca(2+)](i) in cortical neurons. This cellular difference is consistent with the ability of AVP to increase water permeability in astrocytes but not in neurons. PMID- 10861789 TI - Retinoic acid potentiated the protective effect of NGF against staurosporine induced apoptosis in cultured chick neurons by increasing the trkA protein expression. AB - Nerve growth factor (NGF) has already been shown to protect neurons and PC12 cells from cell death induced by different stimuli. When chick embryonic neurons were exposed to staurosporine (200 nM, 24 hr), the percentage of apoptotic neurons increased from 15% in controls to 80%, but the treatment with NGF alone did not show any neuroprotection. In the presence of retinoic acid (RA, 5 microM), however, NGF (20 pg/ml) reduced staurosporine-induced damage to 42% apoptotic neurons compared to 58% in the presence of RA (5 icroM) alone. TrkA protein expression in chick neurons was markedly reduced by staurosporine, but was found to be increased in the presence of RA and NGF compared with the treatment with staurosporine alone. The antiapoptotic effect caused by RA and NGF was abolished by the tyrosine kinase inhibitor K-252a, as well as by anti-trkA antibodies and anti-NGF antibodies suggesting that the increase in trkA protein expression contributed to its mechanism of action. In addition, RA-enhanced 2.6 fold the NGF secretion from cultured rat cortical astrocytes and conditioned medium of RA-treated astrocytes reduced the percentage of apoptotic chick neurons after a 24 hr-incubation with staurosporine in the same manner as the external addition of RA and NGF. Increasing the endogenous synthesis of growth factors as well as the expression of their receptors by small, blood-brain barrier-permeable drugs was suggested as a promising concept for neuroprotection. PMID- 10861790 TI - Proton magnetic resonance spectroscopy of cerebrospinal fluid in neurodegenerative disease: indication of glial energy impairment in Huntington chorea, but not Parkinson disease. AB - Metabolite levels in cerebrospinal fluid from patients with Parkinson disease or Huntington chorea were compared with the levels in healthy controls using proton magnetic resonance spectroscopy. No significant differences were found for any metabolite measured in cerebrospinal fluid from patients with Parkinson disease compared to controls. Slight but significantly reduced levels of both lactate and citrate, however, were found in cerebrospinal fluid from patients with Huntington chorea compared to controls. This suggests possible impairment of both glycolysis and tricarboxylic acid cycle function. The reduction in lactate found in the present study may reflect neuronal loss. The decrease in citrate supports the theory of mitochondrial dysfunction in the brain of patients with Huntington chorea, but also suggests that there may be an important astrocytic component in this disease. If so, it would certainly have implications for neuronal function. PMID- 10861791 TI - Brain-derived neurotrophic factor and tyrosine kinase receptor TrkB in rat brain are significantly altered after haloperidol and risperidone administration. AB - The antipsychotics haloperidol and risperidone are widely used in the therapy of schizophrenia. The former drug mainly acts on the dopamine (DA) D(2) receptor whereas risperidone binds to both DA and serotonin (5HT) receptors, particularly in the neurons of striatal and limbic structures. Recent evidence suggests that neurotrophins might also be involved in antipsychotic action in the central nervous system (CNS). We have previously reported that haloperidol and risperidone significantly affect brain nerve growth factor (NGF) level suggesting that these drugs influence the turnover of endogenous growth factors. Brain derived neurotrophic factor (BDNF) supports survival and differentiation of developing and mature brain DA neurons. We hypothesized that treatments with haloperidol or risperidone will affect synthesis/release of brain BDNF and tested this hypothesis by measuring BDNF and TrkB in rat brain regions after a 29-day treatment with haloperidol or risperidone added to chow. Drug treatments had no effects on weight of brain regions. Chronic administration of these drugs, however, altered BDNF synthesis or release and expression of TrkB immunoreactivity within the brain. Both haloperidol and risperidone significantly decreased BDNF concentrations in frontal cortex, occipital cortex and hippocampus and decreased or increased TrkB receptors in selected brain structures. Because BDNF can act on a variety of CNS neurons, it is reasonable to hypothesize that alteration of brain level of this neurotrophin could constitute one of the mechanisms of action of antipsychotic drugs. These observations also support the possibility that neurotrophic factors play a role in altered brain function in schizophrenic disorders. PMID- 10861792 TI - Electrochemical monitoring of superoxide anion production and cerebral blood flow: effect of interleukin-1 beta pretreatment in a model of focal ischemia and reperfusion. AB - Conditions associated with systemic infection, such as endotoxinemia, are known to increase the levels of pro-inflammatory cytokines such as interleukin (IL)-1 in the central nervous system. Systemic infection has been shown to be a common preexisting condition in patients with stroke. To examine a possible consequence of systemic infection, we used a novel electrochemical technique, which combines measurement of cerebral blood flow with measurement of superoxide anion concentrations, to examine the effect of pretreatment of pial vasculature with a proinflammatory cytokine, IL-1 beta, on cerebral blood flow and superoxide anion concentration in a rat model of middle cerebral artery occlusion and reperfusion. In addition, neutrophil recruitment was measured using an immunohistochemical technique. Our results indicate that exposure of pial and cerebral vasculature to IL-1 beta significantly accelerates recruitment of neutrophils, reduces cerebral blood flow, and increases superoxide anion concentration at the pial surface during reperfusion. These results support the idea that prior exposure of brain vasculature to IL-1 beta results in acceleration of cerebrovascular injury by accelerating recruitment of neutrophils, which secrete superoxide anion, during reperfusion. This finding has possible implications for the treatment of stroke with reperfusion agents in patients with preexisting infections. PMID- 10861793 TI - Toxicity of various amyloid beta peptide species in cultured human blood-brain barrier endothelial cells: increased toxicity of dutch-type mutant. AB - The amyloid beta peptide (A beta) is the major component of the neuritic and cerebrovascular amyloid plaques that are one of the characteristic features of Alzheimer's disease (AD). This peptide has been shown to be toxic to several relevant cell types, including neurons, cerebrovascular smooth muscle cells, and endothelial cells. We have studied the toxic effects of both soluble and aggregated species of A beta(1-40) and the mutation A beta(1-40)Glu-->Gln(22), which is the major species deposited in the cerebrovascular blood vessels of victims of hereditary cerebral hemorrhage with amyloidosis, Dutch type. We find that aggregates of both peptides, as well as of A beta(1-42) and A beta(25-35), are toxic to cultured human cerebrovascular endothelial cells (hBEC) obtained from the brain of a victim of AD (at doses lower than those that are toxic to CNS neurons or leptomeningeal smooth muscle cells). Soluble A beta(1-40) Gln(22) is equally toxic to hBEC, whereas wild-type A beta(1-40) is toxic only at higher doses. This toxicity is seen at the lowest dose of A beta(1-40) Gln (22) used, 20 nM. The soluble A beta(1-40)Gln(22) aggregates on the surface of the cells, in contrast to A beta(1-40), and its toxicity can be blocked both by an inhibitor of free radical formation and by Congo red, which inhibits amyloid fibril formation. We discuss the possibility that the enhanced toxicity of A beta(1-40)Gln(22) is mediated by a A beta receptor on the endothelial cells. PMID- 10861794 TI - Broad specificity of GDNF family receptors GFRalpha1 and GFRalpha2 for GDNF and NTN in neurons and transfected cells. AB - The glial cell line-derived neurotrophic factor (GDNF) family of ligands binds to lipid anchored proteins termed GDNF family receptor (GFR)alphas, and then activates the RET receptor tyrosine kinase, by ligand GFRalpha. The binding of soluble GFRalphas to transfected cells suggested that different GFRalphas were dedicated to particular ligands, with GDNF acting primarily or entirely through GFRalpha1, and neurturin (NTN), through GFRalpha2. More recent evidence has suggested the possibility of cross-talk between these ligands and the two receptors. We examined here whether crosstalk between the GDNF ligands and the GFRalphas is biologically relevant, using midbrain dopaminergic, and parasympathetic, submandibular gland neurons. By biochemical and genetic addition and/or deletion of GFRalpha1 and 2, we show that in both neuronal cell types, robust biological activities of GDNF or NTN can be mediated by either GFRalpha1 or GFRalpha2, although GDNF is slightly more potent in dopaminergic (DA) neurons which normally express GFRalpha1, and NTN in submandibular neurons which normally express GFRalpha2. Throughout the body, GDNF and NTN are likely to have important biological actions on both GFRalpha1- and GFRalpha2-expressing cells. PMID- 10861795 TI - Comparative evaluation of cytokine profiles and reactive gliosis supports a critical role for interleukin-6 in neuron-glia signaling during regeneration. AB - Using reverse transcription polymerase chain reaction (RT-PCR), we have studied the temporal expression of interleukin-1beta (IL-1beta), interleukin-6 (IL-6), transforming growth factor-beta 1 (TGF-beta 1), and tumor necrosis factor-alpha (TNF-alpha) mRNAs in three axotomy paradigms with distinct functional outcomes. Axotomy of adult rat facial motoneurons results in neuronal regeneration, axotomy of neonatal facial motoneurons results in neuronal apoptosis, and axotomy of rubrospinal neurons results in neuronal atrophy. Our RT-PCR findings show that a significant and sustained upregulation of IL-6 mRNA is associated uniquely with the regeneration of adult facial motoneurons. Histochemical studies using IL-6 immunohistochemistry show intense IL-6 immunoreactivity in axotomized adult facial motoneurons. Assessment of reactive glial changes with astroglial and microglial markers reveals that the reactive gliosis following adult facial nerve axotomy is more intense than that observed in either of the other two paradigms. Exposure of cultured microglial cells to IL-6 stimulates microglial proliferation in a dose-dependent manner. Cultured microglia also show expression of IL-6 receptor mRNA, as determined by RT-PCR. Our findings support the idea that reactive gliosis is required for neuron regeneration to occur, and more specifically, they suggest that neuron-derived IL-6 serves as a signalling molecule that induces microglial proliferation during motoneuron regeneration. PMID- 10861796 TI - Wnt-1 dependent activation of the survival factor NF-kappaB in PC12 cells. AB - Expression of the Wnt-1 oncogene in PC12 cells induces morphological and biochemical changes, including up-regulation of cell adhesion and lack of differentiation in response to growth factors. The survival of PC12 cells is known to be mediated in part by phosphatidylinositol-3 kinase (PI-3 kinase) dependent activation of the transcription factor nuclear factor-kappaB (NF kappaB). We investigated the effect of Wnt-1 expression on cell survival and NF kappaB activation using PC12 cells expressing Wnt-1 (PC12/Wnt1) and a reporter vector in which firefly luciferase expression is under the control of NF-kappaB consensus sequences. Serum deprivation caused apoptosis and decreased NF-kappaB activity in wild type PC12 cells. PC12/Wnt-1 cells showed less apoptosis in the absence of serum, and the levels of NF-kappaB activity were higher than in wild type PC12 cells. NF-kappaB activity was also increased by the transient expression of Wnt-1 in PC12 cells and it was completely inhibited in both PC12 and PC12/Wnt-1 cells by a dominant negative mutant IkappaB-alpha that has been shown to prevent NF-kappaB activation. Agents known to inhibit NF-kappaB-induced apoptosis in PC12 as well as in PC12/Wnt-1 cells, indicating a role of NF-kappaB activation in the anti-apoptotic effect of Wnt-1. Inhibition of PI-3 kinase with wortmannin, or with a dominant negative p85 regulatory subunit of the PI-3 kinase, blocked NF-kappaB activity in PC12 cells but caused only partial inhibition in PC12/Wnt-1 cells. The effect of Wnt-1 in activating NF-kappaB can be mimicked by inhibition of glycogen synthase kinase-3beta (GSK-3beta) with lithium or with a dominant negative GSK-3beta. Our results show that expression of Wnt-1 increases survival of PC12 cells in the absence of serum by activating the anti-apoptotic factor NF-kappaB. Wnt-1-induced activation of NF-kappaB is partially independent of PI-3 kinase and can be mimicked by inhibition of GSK 3beta. PMID- 10861797 TI - The third fibronectin type III repeat is required for L1 to serve as an optimal substratum for neurite extension. AB - As a means of defining functionally important regions of the L1 neuronal cell adhesion molecule, neurite outgrowth from cerebellar neurons was compared on monolayers of L1-negative B28 glioma cells, B28 cells transfected with wild-type human L1, and B28 cells transfected with variant forms of L1. Neurite outgrowth on L1-positive B28 cells is greatly enhanced over that seen on parental B28 cells. Neurite outgrowth on B28 cells expressing L1 variants that lack either the first or the fifth fibronectin type III repeat is comparable to that seen on monolayers expressing wild-type L1. In contrast, B28 cells expressing L1 without the third fibronectin type III repeat do not support neurite outgrowth above the background level seen on parental B28 cells. This suggests that the third fibronectin type III repeat plays a key role in the ability of L1 to promote neurite extension. This is consistent with reports that the third fibronectin type III repeat mediates L1 homomultimerization and integrin binding and that plasmin cleavage within this domain interferes with L1 function by abolishing these molecular interactions. PMID- 10861798 TI - M2 mutations of the nicotinic acetylcholine receptor increase the potency of the non-competitive inhibitor phencyclidine. AB - Phencyclidine (PCP) is a non-competitive inhibitor of the nicotinic acetylcholine receptor (nAChR) with biphasic characteristics. At low and high micromolar concentrations, PCP inhibits nAChR from fetal mouse muscle, whereas at intermediate concentrations PCP does not inhibit the receptor. The present study was performed to determine whether the high and low concentration effects of PCP on mouse nAChR were due to interactions of this blocker with channel lining amino acids. In order to test this hypothesis, we examined the ability of PCP to inhibit acetylcholine-induced currents from wild-type nAChR and nAChR in which amino acid substitutions were made in the 6', 8' and 10' positions of the M2 transmembrane segments of the receptor. Fetal mouse nAChR from BC(3)H-1 cells were expressed in Xenopus laevis oocytes and studied using the two-electrode voltage clamp technique. The results of this study reveal that in native fetal muscle receptor, PCP potency is not affected by membrane potential between -80 mV and -30 mV. The potency of PCP is increased by mutations in M2 6', 8', and 10' positions. This increase in potency cannot be explained merely by either changes in hydrophobicity/hydrophilicity of amino acids at these positions or by side chain size. A model proposing extra-luminal inhibitory and regulatory sites for PCP explains the lack of voltage-dependency, the biphasic effect of PCP, and the fact that all M2 mutations increased PCP potency (by disrupting the link with the regulatory sites). PMID- 10861799 TI - Oligodendrocyte progenitor cells internalize ferritin via clathrin-dependent receptor mediated endocytosis. AB - We previously demonstrated ferritin binding is specific to white matter in mouse and human brain tissue and is not found within Multiple Sclerotic plaques. These results suggest that ferritin receptors are selectively expressed on oligodendrocytes. The present studies were designed to test the hypothesis that oligodendrocyte progenitor cells selectively bind ferritin and internalize it by methods consistent with receptor-mediated endocytosis. Using a cell culture system enriched for oligodendrocyte progenitor cells, we determined, that oligodendrocyte progenitor cells bind ferritin in a saturable and competitive manner with a K(d) of 5 nM and a receptor density of 0.06 fmol bound/20,000 cells. FITC tagged ferritin is internalized by A2B5, O4 or CNPase expressing cells in the culture, but not by GFAP+ cells. The uptake of ferritin into the oligodendrocyte progenitors was inhibited by treating the cells with inhibitors of receptor mediated endocytosis (hypertonic medium, potassium deficient medium, ATP depletion, sulfhydryl reagents). In addition exogenous ferritin decreased iron responsive element/iron regulatory protein binding indicating that the iron within the internalized ferritin is released and contributes to the intracellular iron pool. Given the relatively high amount of iron that can be delivered via ferritin, and the selective distribution of ferritin receptors in the white matter tracts in vivo, we propose that ferritin is a major source of iron for oligodendrocytes. PMID- 10861800 TI - Novel method for the labeling of distant neuromuscular junctions. AB - Essential to understanding the roles proteins and structural elements play at the synapse is to understand the development, remodeling and reinnervation of peripheral neuromuscular junctions. It has, however, been a challenging task to label and visualize neuromuscular junctions. In this paper we demonstrate how adenovirus technology can be combined with intraspinal microinjection techniques to follow both the development and the reinnervation of a distant peripheral neuromuscular junction in the rat. A recombinant adenovirus containing VAMP-2 (synaptobrevin-2) was fused to the green fluorescent protein (GFP) and microinjected into the region of the lumbar motor neurons. We were able to follow the neuronal incorporation, axonal transport and synaptic localization of the GFP VAMP-2 using fluorescence microscopy. GFP-VAMP-2 was found in neuronal cell bodies, selected sciatic nerve axons and was concentrated in the presynaptic nerve terminal. During reinnervation of the neuromuscular junction, GFP-VAMP-2 allows us to follow the time course of junctional reinnervation. Thus, the microinjection of microliter amounts of labeled recombinant virus into locations far distant from target regions can be used to efficiently study the formation of neuromuscular junctions with a minimum of trauma to the animal. PMID- 10861801 TI - Expression of peroxisome proliferator-activated receptors (PPARS) in human astrocytic cells: PPARgamma agonists as inducers of apoptosis. AB - We report the isolation by RT-PCR of partial cDNAs encoding the human peroxisome proliferator-activated receptor (PPAR) isoforms PPARbeta and -gamma in human primary astrocytes (HPA) as well as in the human malignant astrocytoma cell line T98G. In contrast, we failed to detect PPARalpha mRNA in either of these two cell types. Because PPARbeta is ubiquitously expressed but has, as yet, no known function, we pursued our functional studies of these cells with regard to PPARgamma. To that end, we showed that PPARgamma protein is abundantly expressed in both cell types, having a molecular weight of approximately 50 kDa. Immunocytochemistry revealed a predominantly nuclear localization of this receptor. Moreover, incubation of the two cell types with 1-12 mcM 15-deoxy PGJ(2) or 1-12 mcM ciglitazone, both of which are agonists of PPARgamma, induced loss of cellular viability as assessed by the MTT assay after a 4 hr incubation. Reduced cellular viability as a consequence of exposure to PGJ(2) or ciglitazone resulted from induction of apoptosis, as assessed by DNA fragmentation and Hoechst staining, and involves activation of the CPP32 (caspase-3) protease. These data show that modulation of the process of apoptosis is one function of PPARgamma in cells derived from the human astrocytic lineage. PMID- 10861802 TI - Pertussis toxin treatment prevents 5-HT(5a) receptor-mediated inhibition of cyclic AMP accumulation in rat C6 glioma cells. AB - We have investigated the functional coupling of the rat 5HT(5a) receptor subtype to adenylate cyclase in a rat C6 glioma cell line. In 5HT(5a) receptor transfected cells, 5HT caused a concentration-dependent inhibition of forskolin stimulated cAMP accumulation, with an EC(50) value of 41 nM and a maximal effect of 57% inhibition. This effect was dependent on the concentration of forskolin used to elevate cAMP levels. Methiothepin (1 mcM), which has high affinity for the 5HT(5a) receptor, antagonized the 5HT(5a) receptor-mediated inhibition, and unmasked a stimulation of cAMP formation similar to that observed in untransfected cells, whereas ketanserin (0.1 mcM) enhanced the inhibitory effect of 5HT. Pertussis toxin treatment (0.5 mcg/ml) completely blocked the inhibitory effect of 5HT on cAMP formation, also revealing increase in cAMP accumulation. Pretreatment of the transfected membranes with pertussis toxin abolished subsequent ADP-ribosylation of a 41 kDa protein, correlating the cAMP effect with a functional uncoupling of an inhibitory G protein from its receptor. These results demonstrate an efficient functional coupling of the rat 5HT(5a) receptor to the inhibition of adenylate cyclase via a pertussis toxin-sensitive G[alpha(i)], inhibitory G-protein. PMID- 10861803 TI - Antidepressant-induced regulation of 5-HT(1b) mRNA in rat dorsal raphe nucleus reverses rapidly after drug discontinuation. AB - Serotonin release from dorsal raphe projections in the forebrain is regulated by terminal 5-HT(1B) autoreceptors; dysregulation of these receptors may be involved in the pathophysiology of clinical depression. Using in situ hybridization, we have previously reported that fluoxetine reduces 5-HT(1B) mRNA in rat dorsal raphe nucleus (DRN) in a time-dependent and reversible manner. In this study we examined longer term treatment (8 weeks) with several different serotonin selective reuptake inhibitors (SSRIs) or a tricyclic antidepressant on 5-HT(1B) mRNA regulation in DRN and hippocampus, and evaluated the stability of these drugs' effects after drug discontinuation. Fluoxetine (5 mg/kg/d), paroxetine (5 mg/kg/d), sertraline (10 mg/kg/d) or nortriptyline (10 mg/kg/d) was administered to rats via subcutaneous osmotic minipumps. Paroxetine and fluoxetine reduced DRN 5-HT(1B) mRNA by 36% and 27%, respectively whereas sertraline had a no significant effect. After 3-14 days of drug washout, DRN 5-HT(1B) mRNA levels in SSRI treated rats were no longer different from control. 5-HT(1B) mRNA levels in hippocampus were not affected by SSRI drugs at any timepoint. Nortriptyline had no significant effect on 5-HT(1B) mRNA in either DRN or hippocampus. These results confirm that SSRI antidepressants reduce presynaptic 5-HT(1B) mRNA selectively, and that this effect is maintained for at least 8 weeks of antidepressant treatment but reverses rapidly after discontinuation. Furthermore, it is possible that washout after chronic antidepressant treatment, that is routinely used in functional assays of autoreceptor action in animal models, may lead to more rapid reversal of biological effects than has previously been thought. PMID- 10861804 TI - Single-unit and polygraphic recordings associated with systemic or local pharmacology: a multi-purpose stereotaxic approach for the awake, anaesthetic free, and head-restrained rat. AB - In order to avoid any artifactual pharmacological interferences with anaesthetic agents, a procedure has been developed for working on the awake, anaesthetic-free rat in a head-restrained condition. It allows, on the same animal and over several consecutive days, single-unit recordings in combination with systemic or local pharmacology (microiontophoresis or micropressure ejections), as well as monitoring vigilance states via the electroencephalogram and the electromyogram. After the cementing of a special "U"-shaped device on its skull under general anaesthesia, the animal is progressively habituated to stay daily, for several hours, under a painless corresponding stereotaxic restraint. This system can be easily adapted to different stereotaxic frames and, because of its spatial flexibility for targetting the desired rostrocaudal or lateral positions, allows access to a large number of cerebral structures. Experiments performed on Globus Pallidus, Substantia Nigra, and Locus Coeruleus neurons, combining the different possibilities of this system, are reported. They demonstrate, on the awake anaesthetic-free head-restrained rat, and under suitable ethical conditions, the feasibility of single-unit recordings of identified neurons associated with the study of their pharmacological reactivity after systemic or local drug administrations without any other drug interferences, and in physiologically relevant conditions such as the spontaneous alternance of vigilance states. PMID- 10861805 TI - Increased septal 5-HIAA efflux in rats that do not develop learned helplessness after inescapable stress. AB - Learned helplessness is a behavioral deficit that can be induced by exposure to inescapable stress. Previous studies have implicated the lateral septum in mediating this phenomenon, and in this brain region, serotonin plays an important role in the development, maintenance, prevention, and reversal of learned helplessness behavior. Using the technique of in vivo microdialysis, we measured the efflux of serotonin (5-HT), dopamine (DA), and their respective metabolites, 5-hydroxyindoleacetic acid (5-HIAA) and 3, 4-dihydroxyphenylacetic acid (DOPAC), from the lateral septum of rats that either developed or did not develop learned helplessness. During the microdialysis session all rats were subjected to restraint stress. Control groups included naive, home cage rats as well as tested control rats that were subjected to the identical handling, restraint, and shuttlebox testing as the rats that received inescapable shock. Overall, levels of 5-HIAA were significantly higher in non-helpless rats. There were no significant effects of restraint or differences in levels of 5-HT, DA, or DOPAC. We propose that this increase in 5-HIAA is indicative of an overall increase in serotonin metabolism in the lateral septum of rats that do not become helpless after inescapable stress. This increased serotonin metabolism in the lateral septum may protect the animal from adverse behavioral consequences of inescapable stress. J. Neurosci. Res. 61:101-106, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861806 TI - Postnatal development of unipolar brush cells in the cerebellar cortex of cat. AB - The postnatal developmental distribution pattern of metabotropic glutamate receptor (mGluR1a) immunoreactive unipolar brush cells (UBCs) was studied in the cerebellar cortex of kittens. On the day of birth (P0) UBCs are already present in the white matter in lobule X of the vermis, but only a few of these cell seemed to migrate to the deeper region of the internal granular layer. By the end of the first week (P8) UBCs were seen to invade the white matter + internal granular layer of lobules IX, VIII, I, and II of the vermis, and they spread further in the transitory area medio-laterally from the vermis toward the cerebellar hemispheres. By P15, UBCs appeared in lobules III and VII of the vermis, as well as in corresponding lobules of the neocerebellum, with especially high numbers in lobule VII. By P22, UBCs migrated further after their medio lateral course in the neocerebellum, and began to invade lobules V and VI. At P62 the amount of UBCs in midsagittal planes of early developing vermal lobules (I, II, VII-X) resembled the P132 or adult pattern. The medio-lateral migration and incorporation of UBCs into the late-developing cerebellar lobules V and VI was completed only by P132, when the spatial distribution of UBCs in both the vermal and neocerebellar lobules was comparable to that seen in the 1 year old young adult cat. Although by P132 the postnatal migration of the vast majority of UBCs seemed to be completed, in the cerebellum of adult cats a few migrating UBCs could still be observed in the white matter of the cerebellar lobules, and beneath the ependyma of the fourth ventricle. It is concluded that during ontogenesis the migration course of UBCs follows essentially the developmental sequence of cerebellar lobules, although the incorporation of UBCs into the internal granular layer continues until 4 months postnatally, i.e., much beyond the apparent completion (about two months postnatally) of cytoarchitectonic built up of the cerebellar cortex of kittens. PMID- 10861807 TI - Decreased tissue factor and tissue-plasminogen activator antigen in relapsed acute promyelocytic leukemia. AB - This study evaluated hemostatic data in 28 patients with newly diagnosed acute promyelocytic leukemia (APL) and 15 patients with relapsed APL. Activated partial thromboplastin time and prothrombin time were prolonged at initial onset of APL. Plasma level of fibrinogen was significantly decreased in patients with initial disease of APL, but it was not decreased significantly during the relapse of APL. Plasma fibrin and fibrinogen degradation products levels were significantly increased and platelet counts significantly decreased in both groups. Plasma levels of antiplasmin significantly decreased at initial onset but not during relapse. Plasma levels of antithrombin were within normal range in patients with initial disease but significantly decreased in those with relapse. Plasma levels of D-dimer, soluble fibrin monomer (sFM), plasmin-plasmin inhibitor complex (PPIC), and thrombin antithrombin complex (TAT) levels were significantly high in both groups. Plasma levels of PPIC, sFM, and D-dimer were significantly higher at initial onset of APL than during relapse. However, there was no significant difference in DIC score between patients with initial onset and those with relapse; plasma levels of tissue factor (TF) significantly increased in both groups, but they were significantly higher at initial onset of APL than during relapse. TF and tissue type plasminogen activator (t-PA) antigen levels in leukemic cell lysate were significantly increased in both groups, and they were significantly lower during relapse than at initial onset. Hemostatic abnormalities occurring in patients with relapsed APL might be the result of the decrease of TF and t-PA in leukemic cells. These findings suggest that DIC in APL patients with relapse might not be caused only by TF and t-PA and thus should be treated with different therapy from patients with initial onset of APL. PMID- 10861808 TI - HbF production in beta thalassaemia heterozygotes for the IVS-II-1 G-->A beta(0) globin mutation. Implication of the haplotype and the (G)gamma-158 C-->T mutation on the HbF level. AB - We studied the presence of the XmnI site and the beta-globin haplotype in 24 individuals, carriers of the IVSII-1 G-->A beta(0)-globin mutation, of whom fourteen had no detectable levels of HbF, while ten coming from 5 families, presented HbF levels ranging from 1.7 to 9% of the total Hb. Of these beta thalassaemia heterozygotes with fetal hemoglobin, 6 were females and 4 were males with median HbF levels of 4.85% and 4% respectively, and an excess of (G)gamma chains (range (G)gamma/(A)gamma: 55/45-70/30). Of the group of carriers of beta thalassaemia with HbF < 0.1, in all cases except one, IVSII-1 mutation was found associated with XmnI polymorphic site. Haplotype analysis in these individuals revealed that in 10 cases IVSII-1 was linked with haplotype IIIb, in 1 case with haplotype IIIa, and in 3 cases with haplotype IX. On the other hand, in the group of carriers with measurable levels of HbF, IVSII-1 was always associated with haplotype IIIa and the XmnI site was either in-homozygous or the heterozygous state in-cis or in-trans with the mutated beta-globin gene. In conclusion the results of the study of these families seem that XmnI site in-cis with the IVSII 1 does not induce HbF production when this beta(0)-thalassaemia mutation is associated with IIIb or IX haplotype. On the other hand the (G)gamma -158 C-->T mutation is associated with small amounts of HbF in IVSII-1 heterozygotes, when the beta-globin mutation is linked to haplotype IIIa. PMID- 10861809 TI - Hematopoietic recovery after IEV chemotherapy for malignant lymphoma followed by different cytokines can be monitored by analysis of Galpha 16 and CD34. AB - The hematopoiesis-specific G protein alpha subunit Galpha16 is a specific element in the signal transduction of the early hematopoietic cytokine network. As Galpha16 mRNA can be detected in early hematopoietic progenitor cells, RT-PCR for Galpha16 can be used as a sensitive marker of hematopoietic activity. The aim of this study was to test the possible use of Galpha16 determinations for monitoring cytokine effects on hematopoietic recovery after chemotherapy in patients. We correlated presence of Galpha16 mRNA and CD34 surface antigen with hematopoietic recovery in six lymphoma patients undergoing salvage therapy with different cytokine support (IEV followed by G-CSF, IL-3, or placebo). Regardless of different cytokine schedules with different time courses, hematopoietic recovery was always preceded by transcription of Galpha16. Monitoring the expression of Galpha16 mRNA by RT-PCR is a highly sensitive diagnostic tool for analyzing hematopoietic recovery after chemotherapy and for characterizing the effects of cytokines on hematopoiesis. PMID- 10861810 TI - Many unbalanced translocations show duplication of a translocation participant. Clinical and cytogenetic implications in myeloid hematologic malignancies. AB - If a translocation is followed by loss of one of the two derivative chromosomes, the result is an unbalanced translocation, showing monosomy for the segments making up the lost derivative. We have found that in most unbalanced translocations, a third event takes place: a morphologically normal copy of one of the two translocation participants is added to the karyotype. This creates a complex abnormal karyotype with monosomy, disomy, and trisomy for different segments of the translocation participants. We have examined 82 unbalanced translocations from 77 patients, 73 of whom had a myeloid hemopoietic malignancy. Acquisition of a normal copy of a translocation participant was found in 49 translocations. Twenty-five of these showed trisomy for 1q. In 16 of the 25 1q trisomic cases the translocation was t(1;7)(q10;p10) (trisomy for 1q and monosomy for 7q). Patients with trisomy for 1q were younger than the remaining patients. Whereas those with t(1;7))(q10;p10) showed brief survivals, those with trisomy 1q but monosomy for regions other than 7q survived longer than the remaining patients. We conclude that most unbalanced translocations involve a partial trisomy, that 1q is trisomic far more frequently than any other segment, and that partial trisomy is associated with patient age and survival. PMID- 10861811 TI - Routine chest radiography for the evaluation of febrile neutropenic patients after autologous stem cell transplantation. AB - Chest radiographs are routinely obtained for diagnostic evaluation of neutropenic febrile patients. We investigated the frequency of chest radiographic abnormalities during febrile episodes after autologous PBSC transplants and assessed the relationship of these abnormalities to past history of pulmonary disease, pre-transplant chest radiographic abnormalities, and pulmonary signs or symptoms at time of fever. We also studied the impact of chest radiographic findings on patient management. Sixty-one consecutive adult autologous PBSC transplant recipients were studied. Fifty-two (85%) developed fever, and 20 (38%) of these showed new chest radiographic abnormalities suggestive of pulmonary infection. Patients with pre-transplant chest radiographic abnormalities were more likely to develop additional abnormalities with fever post-transplant. Pulmonary symptoms or signs had low sensitivity or specificity for predicting radiographic abnormalities. Only 40% of patients with pulmonary symptoms or signs had an abnormal chest radiograph. Twenty-six percent of patients with abnormal chest radiographs had no clinical findings suggestive of pulmonary infection. The identification of chest radiographic abnormality did not change empiric antibiotic treatment in any patient. The role of routine chest radiography for diagnostic evaluation of febrile autologous PBSC transplant patients should be re evaluated. PMID- 10861812 TI - Chemotherapy alone versus surgery followed by chemotherapy for stage I/IIE large cell lymphoma of the stomach. AB - The optimal treatment of localized large-cell lymphoma of the stomach remains controversial. In particular, the role of surgical resection of the primary tumor needs to be clearly defined. We have reviewed all patients with a diagnosis of gastric lymphoma and treated in our institutions between 1988 and 1998. Patients fulfilling the following criteria were included in this study: (1) histologically proven large-cell lymphoma of the stomach; (2) adequate pathological materials and complete clinical information for analysis; (3) clinical stage I/II disease according to the Musshoff modification of Ann Arbor system; and (4) received primary chemotherapy alone with anthracycline- or anthracenedione-containing regimens (group A) or curative surgery followed by adjuvant chemotherapy (group B). There were 38 and 21 patients in group A and group B, respectively. All pertinent clinicopathologic features were similar between the two groups of patients, except that patients of group A had significantly more stage II-2 disease (P = 0.004). Of group A, among 36 patients who could be evaluated for response to chemotherapy, there were 29 complete and 1 partial responses, with an overall response rate of 83.3% (95% CI, 71.1-95.5%). The projected 5-year relapse free survival (RFS) and overall survival (OS) were 86.0% (95% CI, 73.3-98.7%) and 72.6% (95% CI, 57.0-88.2%), respectively. On the other hand, the projected 5-year RFS and OS of group B were 77.9% (95% CI, 58.0-97.8%) and 77.8% (95% CI, 57.9-97. 7%), respectively, not significantly different from that of group A. Our data suggest that systemic chemotherapy alone may be a reasonable alternative treatment for stage I/II large-cell lymphoma of the stomach. Resection of the primary tumor before systemic chemotherapy does not appear to improve the cure rate of this group of patients. PMID- 10861813 TI - Diagnosis of the hemolytic state using serum levels of erythrocyte adenylate kinase. AB - Red cell hemolysis is classically diagnosed by a combination of nonspecific laboratory tests, including serum bilirubin, LDH, and the reticulocyte count. None of these tests alone or in combination has the specificity to reliably ascertain the presence of hemolysis. We have previously demonstrated that erythrocyte adenylate kinase (EAK) is a red cell specific enzyme released from damaged red cells. Its activity can be measured in serum by rapid electrophoresis or immunological methods and correlates linearly with the degree of hemolysis in vitro. We now report on a clinical study comparing EAK levels in patients with and without hemolysis. The clinical diagnosis of hemolysis was established in hospitalized patients with anemia by the combined elevation of the bilirubin, LDH, and reticulocyte count in the absence of liver disease and demonstrable blood loss. The normal range of serum EAK was determined in 30 healthy nonanemic voluntary blood donors and was 0-3.5 Units (mean = 0.5). In 25 patients with hemolytic anemia due to sickle cell disease, hemolytic transfusion reactions, or TTP, the mean EAK level was 62.4 with a range 0-298 Units (P < 0.001 compared to normals). Levels of EAK exceeded the normal range in 24 of 25 patients (96%). In a control group of 44 hospitalized patients with liver disease or myocardial infarction and no clinical evidence of hemolysis, the mean EAK level was 0.12 with a range of 0-3.2 (P = 0.1, NS compared to normals and P < 0.001 compared to patients with hemolysis). None of the control patients had EAK levels that exceeded the normal range. The diagnostic sensitivity of the EAK assay for hemolysis, as calculated according to Baye's algorithm, was 96%, with a specificity and accuracy of 97%. Measurement of serum EAK represents a highly sensitive and specific test for the diagnosis of hemolytic anemia. PMID- 10861814 TI - The Gardos channel is responsible for CDNB-induced dense sickle cell formation. AB - The red blood cells (RBCs) derived from blood taken from homozygous sickle cell (SS) patients demonstrate densities that are inversely proportional to the intracellular reduced glutathione (GSH) content. Addition of 1 mM 1-chloro-2,4 dinitrobenzene (CDNB) to low-density sickle cells (LDSS), at 4 degrees C, results in a shift of LDSS erythrocytes to high-density sickle cells (HDSS), with corresponding decreases in GSH. We have previously demonstrated that this CDNB effect was due to increased K(+) leakage and that dense cell formation could be inhibited by clotrimazole (specific for the Gardos channel) but not DIOA (specific for the K(+)-Cl(-) co-transport system) at pH 7.4 (Shartava et al. Am. J. Hematol. 1999;62:19-24). Here we demonstrate that clotrimazole (10 microM) inhibits dense cell formation at pH 7.1 and 6.8, while DIOA (1 mM) has no effect. As pH 6.8 is the optimal pH for the K(+)-Cl(-) co-transport system, we can now reasonably conclude that damage to the Gardos channel is responsible for CDNB induced dense cell formation. PMID- 10861815 TI - Mantle cell lymphoma. A clinicopathologic study of 68 cases from the Nebraska Lymphoma Study Group. AB - Although mantle cell lymphoma (MCL) is considered a distinctive disease entity within non-Hodgkin's lymphoma (NHL), the cytology and growth pattern of MCL can be quite variable and the clinical significance of these features is unclear. Also, the role of anthracyclines in the management of MCL is unclear. Therefore, we examined our experience with MCL in an effort to clarify these important issues. We identified 68 patients with MCL who were evaluated clinically and treated by the Nebraska Lymphoma Study Group. Treatment consisted of combination chemotherapy containing an anthracycline in 76% of the patients. The cases were grouped by blastic or lymphocytic cytology, and the latter were divided by growth pattern into nodular (or mantle-zone) and diffuse types. The clinical and pathological variables were then evaluated for their prognostic value. The median overall survival (OS) and failure-free survival (FFS) for the entire group were 38 months and 12 months, respectively, and there was no survival advantage for those who received an anthracycline. The cases were grouped as follows: blastic type, 26%; nodular lymphocytic type, 44%; and diffuse lymphocytic type, 30%. Both the cytology and pattern of growth were predictive of OS and FFS. The median OS was as follows: blastic type, 55 months; nodular lymphocytic type, 50 months; and diffuse lymphocytic type, 16 months (P = 0.0038). The clinical features that predicted for a shorter survival included bone marrow involvement, advanced stage disease, B symptoms, a poor performance score, and the International Prognostic Index. We conclude that new therapeutic approaches, with the patients stratified by histologic type and clinical prognostic factors, are clearly needed for MCL. PMID- 10861816 TI - Co-existence of cutaneous T-cell lymphoma and B hairy cell leukemia. AB - A primary cutaneous form of peripheral T-cell lymphoma (PTCL) and a low grade B cell non-Hodgkin's lymphoma that was classified as a variant of hairy cell leukemia (HCL) were simultaneously diagnosed in a 79-year-old woman by both phenotypic and genotypic analyses. The coexistence of a T- and B-cell lymphoma in the same patient is rare, and, to our knowledge, this particular association has not been previously described. The patient was referred to our Department for evaluation of multiple cutaneous itchy, reddish plaques; laboratory analyses disclosed a lymphocytosis, that presented 6 years earlier. A bone marrow aspirate showed a 50% B-cell interstitial infiltrate, while a skin biopsy surprisingly revealed a PTCL. Clonality of both neoplastic processes was assessed by Southern blot analysis. The indolent clinical course of the cutaneous disease, and the low and stable number of circulating neoplastic T cells supported the diagnosis of a mycosis fungoides (MF)-like PTCL. Possible oncogenic events and/or putative underlying viral infections which could have played a role in the occurrence of B and T-cell non-Hodgkin's lymphomas in the same patient are discussed. PMID- 10861817 TI - Serial analysis of MLL-AF4 chimeric message through successful bone marrow transplantation in a patient with t(4;11)-positive infant-ALL. AB - A 28-month-old girl with acute lymphoblastic leukemia (ALL) showing a t(4;11)(q21;q23) karyotype successfully underwent allogeneic bone marrow transplantation (BMT) at relapse. The chimeric MLL-AF4 message on her bone marrow (BM) specimens, examined by reverse transcriptase-polymerase chain reaction, was detectable at diagnosis, relapse, and just before BMT but became undetectable following BMT. She has since maintained complete remission. This observation suggests that detection of the chimeric message in BM may be useful to predict clinical outcome in patients with t(4;11) ALL. PMID- 10861818 TI - beta-thalassemia intermedia caused by compound heterozygosity for Hb Malay (beta codon 19 AAC-->AGC; asn-->Ser) and codons 41/42 (-CTTT) beta(0)-thalassemia mutation. AB - We report a case of beta-thalassemia intermedia caused by compound heterozygosity for hemoglobin (Hb) Malay and codon 41/42 (-CTTT) beta(0)-thalassemia mutation in a 38-year-old Chinese woman. This patient has long-standing anemia with a baseline Hb level of around 70 g/L. She worked as a full-time cashier and had not required regular blood transfusions. Nevertheless, she had splenomegaly necessitating splenectomy, cholelithiasis, and iron overload. This case illustrates the varied phenotypic expression associated with compound heterozygosity for Hb Malay and other beta-thalassemia mutations. Since Hb Malay migrates as Hb A on electrophoresis and chromatography, this variant Hb mutation ought to be included in the differential diagnosis for beta-thalassemia major or intermedia patients of Southeast Asian descent who are reported to have Hb A on the basis of Hb analysis. The possible presence of this mutation should also be considered in appropriate cases for genetic counseling in couples at risk of conceiving fetuses with beta-thalassemia major or intermedia. PMID- 10861819 TI - Hyperviscosity syndrome secondary to a myeloma-associated IgG(1)kappa paraprotein strongly reactive against the HIV-1 p24 gag antigen. AB - Hyperviscosity syndrome secondary to hypergammaglobulinemia is a rare and potentially fatal complication in patients with human immunodeficiency virus type 1 (HIV-1) infection. We studied an HIV-1-positive patient with symptomatic hyperviscosity attributable to IgG(1)kappa multiple myeloma. The patient initially responded to plasmapheresis and was subsequently treated with cytotoxic immunosuppressive chemotherapy. The patient remained asymptomatic during a 3-year follow-up period. The monoclonal IgG(1)kappa gammopathy evolved to a biclonal variant of the same subtype with an expansion of marrow plasma cell population. Western blot analysis demonstrated that this myeloma-associated paraprotein was strongly reactive against the HIV-1 p24 gag antigen. PMID- 10861820 TI - Report of a factor VIII inhibitor in a patient with autoimmune lymphoproliferative syndrome. AB - The occurrence of factor VIII inhibitors in non-hemophilic patients is a rare event with a potentially lethal outcome. Despite its infrequent occurrence, the association of this inhibitor with multiple autoimmune diseases is well recognized. We report the case of a patient with the recently described autoimmune lymphoproliferative syndrome (ALPS) who developed an inhibitor to factor VIII. ALPS is a disease characterized by defective lymphocyte apoptosis due to inherited mutations in genes that regulate apoptosis, with the resulting enlargement of lymphoid organs and a variety of autoimmune manifestations. Published 2000 Wiley-Liss, Inc. PMID- 10861821 TI - Ruptured Klebsiella pneumoniae liver abscess after high-dose cyclophosphamide for severe aplastic anemia. AB - A 19-year-old woman with severe aplastic anemia who had previously failed antithymocyte globulin/cyclosporine A received high-dose cyclophosphamide without bone marrow rescue. On day +14, she complained of right upper quadrant abdominal pain and fever. A CT scan of the abdomen showed multiple liver abscesses with rupture and Klebsiella pneumoniae was isolated from blood. In spite of aggressive antibiotic therapy, she rapidly deteriorated and died of overwhelming sepsis. To our knowledge, our patient is the first case of fatal ruptured liver abscess after high-dose cyclophosphamide in a patient with severe aplastic anemia. PMID- 10861822 TI - Subcutaneous panniculitis by Epstein-Barr virus-infected natural killer (NK) cell proliferation terminating in aggressive subcutaneous NK cell lymphoma. AB - We describe here a case involving a patient presenting initially with subcutaneous panniculitis, which developed after 12 years into aggressive subcutaneous natural killer (NK) cell lymphoma with peripheral blood involvement and hemophagocytosis. The surface marker of lymphoid cells in peripheral blood was CD2+3-7+8-16+56+. Skin biopsies were taken in May 1986 and June 1998. The initial biopsy revealed a diffuse proliferation of atypical lymphoid cells in the subcutaneous tissue with panniculitis, while the second biopsy revealed the presence of large lymphoid cells in the subcutaneous tissue with necrotic changes, consistent with a diagnosis of malignant lymphoma (diffuse pleomorphic type). The lymphoid cells from these two specimens were positive for CD56 and such cytotoxic molecules as T-cell intracellular antigen-1 (TIA-1), granzyme B, and, interestingly, also positive for Epstein-Barr (EB) virus by in situ hybridization. This suggests that chronic EB virus infections play an important role in the early stages of tumorigenesis and in the progression of NK cell lymphoproliferative disorders. PMID- 10861823 TI - Combined heterozygosity of factor V leiden and the G20210A prothrombin gene mutation in a patient with cerebral cortical vein thrombosis. AB - Cerebral venous thrombosis (CVT) is a rare type of stroke with a variety of causes. Several reports have suggested that either factor V Leiden or G20210A prothrombin gene mutation is associated with an increased risk of CVT. The genetic thrombophilias are typically associated with other predisposing factors. We report a unique case of CVT in a patient with both the factor V Leiden and the G20210A prothrombin gene mutations without other identifiable precipitating factors in a 28-year-old white male in good health. MRI and cerebral arterial angiography showed cerebral cortical venous thrombosis. This case suggests that combined heterozygous individuals may be particularly prone to spontaneous thrombosis, like CVT. PMID- 10861824 TI - Chimeric del20q in a case of chronic neutrophilic leukemia. AB - Chronic neutrophilic leukemia (CNL) is a rare myeloproliferative disorder in which recurrent abnormalities of chromosome 20 have been reported. We report the case of a 76-year-old woman with CNL with partial deletion of the long arm of chromosome 20 in a subset of bone marrow metaphases, suggesting coexistence of a clonal stem cell disorder and normal hematopoiesis. Review of the literature suggests that such mosaicism is common in CNL, possibly accounting for the favorable prognosis observed in many patients with this disorder. PMID- 10861825 TI - B-cell bradycardia: carotid sinus massage by a high-grade lymphoma. PMID- 10861826 TI - Relapse after autologous hematopoietic stem cell transplantation in acute myeloid leukemia, a single center experience over 13 years. PMID- 10861827 TI - Fludarabine and risk of hepatitis B virus reactivation in chronic lymphocytic leukemia. PMID- 10861828 TI - Fine structure of the spermatophore and spermatozoa in inseminated females of Pergamasus mites (Acari: Gamasida: Pergamasidae). AB - This study describes the spermatophore of Pergamasus mites after transfer into the female endogynial sac and modifications of the structure of the female penetrating spermatozoa. The spermatophore is saccular and contains largely unmodified spermatozoa and variously structured secretions. The spermatozoa that leave the spermatophore reach the inner (proximal) end of the vaginal duct where they presumably escape from the genital tract to continue their route through the hemocoel into the ovarian tissue where fertilization occurs. During this transit, the structure of such spermatozoa changes considerably, in particular when in contact with the ovary. These alterations include modifications of the cell periphery and of certain inclusion bodies. Furthermore, the spermatozoa form protrusions that fit into corresponding invaginations of the somatic tissue of the ovary. PMID- 10861829 TI - Serial homology of the mandible and maxilla in the jumping bristletail Pedetontus unimaculatus Machida, based on external embryology (Hexapoda: Archaeognatha, Machilidae). AB - The development of the mandible and maxilla is examined with the scanning electron microscope in the Archaeognatha. Serial homology is discussed to elucidate the general construction of the hexapod mandible. The part comparable to the maxillary palp does not develop in the mandible. Thus, the mandible is coxopodal in origin, and not telognathic but coxognathic. The mandible proper is subdivided into two in late embryonic development, and the smaller proximal and larger distal parts are homologized with the maxillary cardo and stipes, respectively, being subcoxal and coxal in nature. The partition into the "mandibular cardo" in which the mandibular monocondyle is formed and the "mandibular stipes" is recognized as a cuticular ridge or the "mandibular basal ridge" in the postembryonic stages including the imaginal. The molar and incisor are comparable in position and homologized with the maxillary lacinia and galea, respectively. The lacinia and galea could be morphologically interpreted as being the endites of the maxillary coxae I and II, respectively, and the molar and incisor might represent the mandibular coxae I and II as their constituents or endites. PMID- 10861830 TI - Septomaxilla of nonmammalian synapsids: soft-tissue correlates and a new functional interpretation. DELETE. AB - The function of the septomaxilla of nonmammalian synapsids has long been problematic. Distinctive features of this bone, including a prominent intranarial process and a septomaxillary canal and foramen, are characteristic of pelycosaurs and nonmammalian therapsids, but are lost in their mammalian descendants. Numerous contradictory reconstructions have been proposed for the soft anatomy associated with the septomaxilla of nonmammalian synapsids. This review supports the following conclusions: 1) No particular correlation exists between the septomaxilla and the vomeronasal organ (VNO), and the most likely location for the VNO is on the dorsal surface of the palatal process of the vomer; 2) The most likely occupant of the septomaxillary canal is the nasolacrimal duct, which opened either anterior or medial to the intranarial process, near the opening of the VNO duct; and 3) The occupant of the septomaxillary foramen remains uncertain. These conclusions suggest that the functional significance of the septomaxilla in the nonmammalian synapsids is tied to that of the nasolacrimal duct. The association of this duct and the VNO in these animals resembles the condition in Recent amphibians and lepidosaurs, in which the nasolacrimal duct supplies orbital fluids to the VNO, apparently to enhance vomeronasal function. The peculiar shape of the synapsid septomaxilla may have served to collect vomeronasal odor molecules. The changes of the septomaxilla in early mammals, and its nearly complete loss in extant mammals, are probably correlated with a dissociation of the nasolacrimal duct and VNO, and functional changes in both structures. PMID- 10861831 TI - Strain in the galago facial skull. AB - Little experimental work has been directed at understanding the distribution of stresses along the facial skull during routine masticatory behaviors. Such information is important for understanding the functional significance of the mammalian circumorbital region. In this study, bone strain was recorded along the dorsal interorbit, postorbital bar, and mandibular corpus in Otolemur garnettii and O. crassicaudatus (greater galagos) during molar chewing and biting. We determined principal-strain magnitudes and directions, compared peak shear-strain magnitudes between various regions of the face, and compared galago strain patterns with similar experimental data for anthropoids. This suite of analyses were used to test the facial torsion model (Greaves [1985] J Zool (Lond) 207:125 136; [1991] Zool J Linn Soc 101:121-129; [1995] Functional morphology in vertebrate paleontology. Cambridge: Cambridge University Press, p 99-115). A comparison of galago circumorbital and mandibular peak strains during powerful mastication indicates that circumorbital strains are very low in magnitude. This demonstrates that, as in anthropoids, the strepsirhine circumorbital region is highly overbuilt for countering routine masticatory loads. The fact that circumorbital peak-strain magnitudes are uniformly low in both primate suborders undermines any model that emphasizes the importance of masticatory stresses as a determinant of circumorbital form, function, and evolution. Preliminary data also suggest that the difference between mandibular and circumorbital strains is greater in larger-bodied primates. This pattern is interpreted to mean that sufficient cortical bone must exist in the circumorbital region to prevent structural failure due to nonmasticatory traumatic forces. During unilateral mastication, the direction of epsilon(1) at the galago dorsal interorbit indicates the presence of facial torsion combined with bending in the frontal plane. Postorbital bar principal-strain directions during mastication are oriented, on average, very close to 45 degrees relative to the skull's long axis, much as predicted by the facial torsion model. When chewing shifts from one side of the face to the other, there is a characteristic reversal or flip-flop in principal-strain directions for both the interorbit and postorbital bar. Although anthropoids also exhibit an interorbital reversal pattern, peak-strain directions for this clade are opposite those for galagos. The presence of such variation may be due to suborder differences in relative balancing-side jaw-muscle force recruitment. Most importantly, although the strain-direction data for the galago circumorbital region offer support for the occurrence of facial torsion, the low magnitude of these strains suggests that this loading pattern may not be an important determinant of circumorbital morphology. PMID- 10861832 TI - Nervous and sensory system correlates of an epibenthic evolutionary radiation in antarctic notothenioid fishes, genus Trematomus (Perciformes; Nototheniidae). AB - The perciform suborder Notothenioidei consists of 120 species, with 94 confined to the Antarctic Region of the Southern Ocean. On the Antarctic shelf, this phyletic radiation has been accompanied by a substantial morphological and ecological diversification towards a pelagic existence. For example, the primarily benthic genus Trematomus contains an epibenthic radiation that includes T. loennbergii, T. lepidorhinus, and T. eulepidotus. By comparing these epibenthic species with three congeneric benthic species (T. scotti, T. pennellii, and T. bernacchii) we tested three null hypotheses regarding brain variation in Antarctic trematomids: 1) that there is no difference in brain morphology among the six species; 2) that phylogenetic and ecological factors do not influence brain morphology; and 3) that peripheral sensory structures do not influence brain morphology. We rejected each of these hypotheses, leading us to conclude that Trematomus brains vary interspecifically, between benthic and epibenthic species, and with a species' depth distribution. Further, we conclude that brain variation is correlated with differences in peripheral sensory systems and motor activity. Specifically, epibenthic Trematomus have larger percentages of their brain volume devoted to lateral line mechanoreceptive and motor (cerebellar) structures. Species living at greater depths have low ratios of cones:rods in the retina and larger olfactory structures. PMID- 10861833 TI - Ultrastructure of the skin mechanoreceptors of the Chinese giant salamander, Andrias davidianus. AB - The ultrastructure of two kinds of mechanoreceptive organs, pit organs and neuromasts, in the skin of adult giant salamanders (Andrias davidianus) was studied by transmission electron microscopy. Neuromasts and pit organs differ in their types of synapses, the spatial distribution of kinocilia on sensory cells, and in the degree to which sensory cells are separated by processes of the supporting cells; the two organs probably serve complementary functions. The neuromasts in A. davidianus differ from those of other salamanders in the orientation of kinocilia, in the extent of intrusion of supporting cells into the sensory layer, in the degree of thickening of the synaptic membranes, in the distribution of synaptic spheres, and by the absence of a cupula. PMID- 10861834 TI - Regulation of inducible bradykinin B1 receptor gene expression through absence of internalization and resensitization. AB - Rapid induction and down-regulation of bradykinin B1 receptor (BKB1R) gene expression is tightly regulated at the transcriptional and mRNA levels (Zhou et al. [1998] Biochem. J. 330:361-366; Zhou et al. [1999] Mol. Cell Biol. Res. Commun. 1:29-35). Here we explore regulation of BKB1R expression at the protein level. To make this inducible gene express constitutively, we utilized a bicistronic mammalian expression vector (pCMin) for stable transfection of the BKB1R gene into human lung fibroblasts, IMR90SV40. The BKB1R displayed a high affinity and specificity (K(d) = 0.5 nM) for desArg(10)-kallidin. The receptor mediated such signaling events as arachidonic acid (ARA) release, phosphoinositide (PI) turnover and Ca(2+)-flux. The receptor function proved differentially desensitized. For example, after initial exposure to desArg(10) kallidin, a second stimulation with desArg(10)-kallidin did not induce further Ca(2+)-flux or ARA-release while PI-turnover continued unabated. Unlike most of the G-protein coupled receptors, the BKB1R did not internalize within 60 min of exposure to 10 nM desArg(10)-kallidin. It also did not resensitize. Thus, the duration and signal capacity of the BKB1R at the protein level is regulated through lack of internalization, an absence of resensitization and a lack of desensitization for certain events such as PI turnover. In fact, the absence of BKB1R resensitization is likely a very important contributor to the rapid disappearance of this inducible receptor. PMID- 10861835 TI - Role of cyclic AMP in idiopathic nephrotic syndrome: a pathway involving a decrease in glomerular cell heparan sulfates? AB - The physiopathological mechanisms of idiopathic nephrotic syndrome involve a circulating plasma factor and a decrease in HS in the glomerular basement membrane. Previous studies have demonstrated that plasma from patients with INS decreases glomerular cell HS in vitro. We examined the involvement of cyclic adenosine monophosphate (cAMP) in this interaction. We studied the effect of plasma from patients with INS on mesangial cell cAMP. We also determined mesangial cell HS when cAMP levels were modified using a cationic membrane after metabolic labeling. Cellular cAMP levels increased significantly when mesangial cells were incubated with plasma from patients with INS in comparison with control plasma (+77%, P = 0.01). Forskolin and IBMX, which increased cellular cAMP, decreased HS levels (-21 +/- 9% and -15 +/- 6% respectively, P < 0.05 for both), whereas dideoxyadenosine, which decreased cellular cAMP, increased HS levels (+24 +/- 7%, P < 0.05). Plasma from patients with INS decreased glomerular cell HS in comparison with control plasma (-34 +/- 8%, P < 0,05). This effect was abolished when cells were preincubated with ddAdo to prevent an increase in cAMP levels. We conclude that in mesangial cells, plasma from patients with INS increases cAMP levels, and that cAMP mediates a decrease in HS levels. Moreover, the action of plasma from patients on HS was inhibited when an increase in cAMP was prevented. cAMP may therefore be instrumental in the negative effect of the plasma factor on mesangial cell HS. PMID- 10861836 TI - ERK1/2 phosphorylation, induced by electromagnetic fields, diminishes during neoplastic transformation. AB - It has been suggested that electromagnetic (EM) fields can act as co-promoters during neoplastic transformation. To examine this possibility, we studied the effects of 0.8-, 8-, 80-, and 300-microT 60-Hz electromagnetic (EM) fields in INITC3H/10T1/2 mouse fibroblast cells. These cells are transformed carcinogenically by methylcholanthrene, but the neoplastic phenotype can be suppressed indefinitely by the presence of retinyl acetate (RAC) in the culture medium. The effects of EM field exposures were examined at three stages: (1) before initiation of transformation (i.e., RAC in the culture media); (2) early in the transformation process (4 days after withdrawal of RAC); and (3) at full of neoplastic transformation (10 days after withdrawal of RAC). EM field exposures induced significant increases in protein levels for hsp70 and c-Fos and in AP-1 binding activity. EM fields induced phosphorylation of MAPK/ERK1/2 before the onset of transformation, but these increases diminished during the transformation process. No phosphorylation in the other major extracellular stress pathway, SAPK/JNK, was detected in cells exposed to EM fields at any time before, during, or after neoplastic transformation. Human cells HL60, MCF7, and HTB124, exposed to EM fields, also showed MAPK/ERK1/2 phosphorylation. Cells treated with the phorbol ester, TPA, served as positive controls for AP-1 activation, c-Fos protein synthesis, and ERK1/2 phosphorylation. There was no indication that EM fields affected the rate of cell transformation or acted as a co-promoter, under the conditions of this study. PMID- 10861837 TI - Cytokine-specific induction of the TGF-beta inducible early gene (TIEG): regulation by specific members of the TGF-beta family. AB - Select members of the TGF-beta family of cytokines play key regulatory roles in skeletal development, structure, and turnover. This laboratory has previously reported that TGF-beta treatment of immortalized normal human fetal osteoblast (hFOB) cells results in the rapid induction of the mRNA levels of a TGF-beta inducible early gene (TIEG) followed by changes in cell proliferation and bone matrix protein production. Previous studies have also shown that nonmembers of the TGF-beta superfamily showed little or no induction of TIEG mRNA. This article further addresses the cytokine specificity of this TIEG induction by examining whether activin and select bone morphogenetic proteins, (BMP-2, BMP-4, and BMP 6), which are representative of different subfamilies of this superfamily, also induce the expression of TIEG in hFOB cells. However, TGF-beta remained the most potent of these cytokines, inducing TIEG mRNA steady-state levels at 0.1 ng/ml, with a maximum induction of 24-fold at 2.0 ng/ml. The BMP-2 (16-fold), BMP-4 (4 fold), and activin (1-3-fold) also induced TIEG mRNA levels, but at reduced degrees compared to TGF-beta (24-fold), and only at much higher cytokine concentrations, e.g., 50-100 ng/ml, compared to 2 ng/ml for TGF-beta. BMP-6 showed no effect on TIEG mRNA levels. The TIEG protein levels generally correlated with the mRNA steady-state levels. As with TGF-beta, BMP-2 treatment of hFOB cells was shown by confocal microscopy to induce a rapid translocation of the TIEG protein to the nucleus. In summary, the relative potencies of these TGF beta family members to induce TIEG expression generally follows the general osteoinductive capacity of these cytokines, with TGF-beta >>> BMP-2 > BMP-4 > activin >> BMP-6. PMID- 10861838 TI - Osteogenic imprinting upstream of marrow stromal cell differentiation. AB - Five spontaneously transformed cell lines were established from a population of murine bone marrow stromal cells (BMSCs) and the expression profiles of phenotype characteristic genes, patterns of in vitro differentiation, and osteogenic capacity after in vivo transplantation were determined for each. All the clones expressed stable levels of cbfa1, the osteogenic "master" gene, whereas the levels of individual phenotypic mRNAs were variable within each, suggestive of both maturational and phenotypic plasticity in vitro. Varying levels of collagen type I and alkaline phosphatase (AP) were expressed in all the clonal lines. The clonal lines with proven in vivo osteogenic potential (3 out of 5) had a high proliferation rate and expressed bone sialoprotein (BSP), whereas the two nonosteogenic clones proliferated more slowly and never expressed BSP. Bone nodules were only observed in 2 out of 3 of the osteogenic lines, and only 1 out of three formed cartilage-like matrix in vitro. There was no evidence of chondrogenesis in the nonosteogenic lines. By contrast, LPL was expressed in two osteogenic and in two nonosteogenic lines. These results demonstrate the presence of multipotential and restricted progenitors in the murine stromal system. cbfa1, collagen type I, and AP expression were common to all, and therefore presumably early, basic traits of stromal cell lines that otherwise significantly differ with respect to growth and differentiation potential. This finding suggests that an osteogenic imprinting lies upstream of diversification, modulation, and restriction of stromal cell differentiation potential. PMID- 10861839 TI - Thapsigargin-induced grp78 expression is mediated by the increase of cytosolic free calcium in 9L rat brain tumor cells. AB - Exposure of 9L rat brain tumor cells to 300 nM thapsigargin (TG), a sarcoendoplasmic Ca(2+)-ATPases inhibitor, leads to an immediate suppression of general protein synthesis followed by an enhanced synthesis of the 78-kDa glucose regulated protein, GRP78. Synthesis of GRP78 increases significantly and continues to rise after 4 h of treatment, and this process coincides with the accumulation of grp78 mRNA. TG-induced grp78 expression can be suppressed by the cytosolic free calcium ([Ca(2+)](c)) chelator dibromo-1, 2 bis(aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA) in a concentration dependent manner. Induction of grp78 is completely abolished in the presence of 20 microM BAPTA under which the TG-induced increase of [Ca(2+)](c) is also completely prevented. By adding ethyleneglycol bis(beta-aminoethyl)ether N,N,N',N' tetraacetic acid in the foregoing experiments, in a condition such that endoplasmic reticulum calcium ([Ca(2+)](ER)) is depleted and calcium influx from outside is prevented, TG-induced grp78 expression is also abolished. These data lead us to conclude that increase in [Ca(2+)](c), together with the depletion of [Ca(2+)](ER), are the major causes of TG-induced grp78 expression in 9L rat brain tumor cells. By using electrophoretic mobility shift assays (EMSA), we found that the nuclear extracts prepared from TG-treated cells exhibit an increase in binding activity toward the extended grp78 promoter as well as the individual cis acting regulatory elements, CRE and CORE. Moreover, this increase in binding activity is also reduced by BAPTA. By competitory assays using the cis-acting regulatory elements as the competitors as well as the EMSA probes, we further show that all of the tested cis elements-CRE, CORE, and C1-are involved in the basal as well as in the TG-induced expression of grp78 and that the protein factor(s) that binds to the C1 region plays an important role in the formation and maintenance of the transcription complex. PMID- 10861840 TI - Shark cartilage extract interferes with cell adhesion and induces reorganization of focal adhesions in cultured endothelial cells. AB - In this study, we examined the effects of shark cartilage extract on the attachment and spreading properties and the focal adhesion structure of cultured bovine pulmonary artery endothelial cells. Treatment with cartilage extract resulted in cell detachment from the substratum. Immunofluorescence staining of those treated cells that remained attached showed that, instead of being present in both central and peripheral focal adhesions as in control cells, both integrin alpha(v)beta(3) and vinculin were found only in peripheral focal adhesion and thinner actin filament bundles were seen. In addition to causing cell detachment, cartilage extract partially inhibited the initial adherence of the cells to the substratum in a dose-dependent manner. Integrin alpha(v)beta(3) and vinculin staining of these cells also showed a peripheral focal adhesion distribution pattern. Vitronectin induced cell spreading in the absence of serum, but was blocked by simultaneous incubation with cartilage extract, which was shown to inhibit both integrin alpha(v)beta(3) and vinculin recruitment to focal adhesion and the formation of stress fibers. Dot binding assays showed that these inhibitory effects on cell attachment and spreading were not due to direct binding of cartilage extract components to integrin alpha(v)beta(3) or vitronectin. Shark cartilage chondroitin sulfate had no inhibitory effect on either cell attachment or spreading of endothelial cells. These results show that the inhibitory effects of cartilage extract on cell attachment and spreading are mediated by modification of the organization of focal adhesion proteins. PMID- 10861841 TI - Potent inhibitory action of red wine polyphenols on human breast cancer cells. AB - Breast cancer (one of the most common malignancy in Western societies), as well as esophagus, stomach, lung, bladder, and prostate cancer, depend on environmental factors and diet for growth and evolution. Dietary micronutriments have been proposed as effective inhibitory agents for cancer initiation, progression, and incidence. Among them, polyphenols, present in different foods and beverages, have retained attention in recent years. Red wine is a rich source of polyphenols, and their antioxidant and tumor arresting effects have been demonstrated in different in vitro and in vivo systems. In the present study, we have measured the antiproliferative effect of red wine concentrate, its total polyphenolic pool, and purified catechin, epicatechin, quercetin, and resveratrol, which account for more than 70% of the total polyphenols in red wine, on the proliferation of hormone sensitive (MCF7, T47D) and resistant (MDA MB-231) breast cancer cell lines. Our results indicate that polyphenols, at the picomolar or the nanomolar range, decrease cell proliferation in a dose- and a time-dependant manner. In hormone sensitive cell lines, a specific interaction of each polyphenol with steroid receptors was observed, with IC(50)s lower than previously described. Interaction of polyphenols with steroid receptors cannot fully explain their inhibitory effect on cell proliferation. In addition, discrete antioxidant action on each cell line was detected under the same concentrations, both by modifying the toxic effect of H(2)O(2), and the production of reactive oxygen species (ROS), after phorbol ester stimulation. Our results suggest that low concentrations of polyphenols, and consecutively, consumption of wine, or other polyphenol-rich foods and beverages, could have a beneficial antiproliferative effect on breast cancer cell growth. PMID- 10861842 TI - Major DNA replication initiation sites in the c-myc locus in human cells. AB - DNA replication initiation sites and initiation frequencies over 12. 5 kb of the human c-myc locus, including 4.6 kb of new 5' sequence, were determined based on short nascent DNA abundance measured by competitive polymerase chain reaction using 21 primer sets. In previous measurements, no comparative quantitation of nascent strand abundance was performed, and distinction of major from minor initiation sites was not feasible. Two major initiation sites were identified in this study. One predominant site has been located at approximately 0.5 kb upstream of exon 1 of the c-myc gene, and a second new major site is located in exon 2. The site in exon 2 has not been previously identified. In addition, there are other sites that may act as less frequently used initiation sites, some of which may correspond to sites in previous reports. Furthermore, a comparison of the abundance of DNA replication intermediates over this same region of the c-myc locus between HeLa and normal skin fibroblast (NSF) cells indicated that the relative distribution was very similar, but that nascent strand abundance in HeLa cells was approximately twice that in NSF relative to the abundance at the lamin B2 origin. This increased activity at initiation sites in the c-myc locus may mainly be influenced by regulators at higher levels in transformed cells like HeLa. PMID- 10861843 TI - Human umbilical vein endothelial cells (HUVECs) show Ca(2+) mobilization as well as Ca(2+) influx upon hypoxia. AB - Bleb formation is an early event of cellular damage observed in a variety of cell types upon hypoxia. Although we previously found that the [Ca(2+)](i) rise before bleb formation only at the same loci of HUVECs upon hypoxia (localized [Ca(2+)](i) rise), the mode of the [Ca(2+)](i) rise remains ill-defined. In order to clarify the mechanisms causing the localized [Ca(2+)](i) rise in hypoxia challenged HUVECs, we studied the effects of several Ca(2+) channel blockers or a Ca(2+) chelator, EGTA, which reduces extracellular Ca(2+) concentration on the hypoxia-induced localized [Ca(2+)](i) rise and bleb formation by employing a confocal laser scanning microscopy (CLSM). After the initiation of hypoxia, [Ca(2+)](i) rose gradually in a localized fashion up to 15 min, which was associated with bleb formation at the same loci. The maximal [Ca(2+)](i) rise was 435 +/- 84 nM at the loci of bleb formation. Ca(2+) channel blockers including Ni(2+) (non-specific, 1 mM), nifedipine (L type, 10 microM), nicardipine (L + T type, 10 microM), and cilnidipine (L + N type, 10 microM) did not inhibit either the localized [Ca(2+)](i) rise or bleb formation. Although both the localized [Ca(2+)](i) rise and bleb formation were inhibited by lowering extracellular Ca(2+) concentration below 100 nM, a diffuse [Ca(2+)](i) rise through the cytoplasm remained without bleb formation, which was inhibited by a phospholipase C (PLC) inhibitor, U73122. In conclusion, hypoxia causes both the Ca(2+) mobilization and the Ca(2+) influx in HUVECs and the Ca(2+) influx through unknown Ca(2+) channels is responsible for the localized [Ca(2+)](i) rise integral to bleb formation. PMID- 10861844 TI - Osteopontin-induced, integrin-dependent migration of human mammary epithelial cells involves activation of the hepatocyte growth factor receptor (Met). AB - Osteopontin (OPN) is a secreted glycophosphoprotein which induces migration of mammary carcinoma cells, and has been implicated in the malignancy of breast carcinoma. Hepatocyte growth factor (HGF) induces cell migration of several mammary epithelial cell (MEC) lines, via activation of its cognate receptor (Met). This study examines the mechanism of OPN-induced MEC migration, in terms of the cell surface integrins involved and induction of the HGF/Met pathway. Three different MEC cell lines were used, representing different stages of tumor progression: 21PT, non-tumorigenic; 21NT, tumorigenic; non-metastatic; and MDA-MB 435, tumorigenic, highly metastatic. Human recombinant OPN was found to induce the migration of all three lines. OPN-induced migration of 21PT and 21NT cells was alphavbeta5 and beta1-integrin dependent, and alphavbeta3-independent, while that of MDA-MB-435 cells was alphavbeta3-dependent. HGF also induced migration of all three cell lines, and a synergistic response was seen to HGF and OPN together. The increased migration response to OPN was found to be associated with an initial increase in Met kinase activity (within 30 min), followed by an increase in Met mRNA and protein expression. OPN-induced cell migration is thus mediated by different cell surface integrins in MEC lines representing different stages of progression, and involves activation of the HGF receptor, Met. PMID- 10861845 TI - Gene therapy for bone formation: in vitro and in vivo osteogenic activity of an adenovirus expressing BMP7. AB - Bone morphogenetic proteins (BMPs) are well-established agents for inducing orthotopic and ectopic bone formation. However, their clinical usefulness as regenerative agents may be limited by a short in vivo half-life and low specific activity. BMP gene therapy is an alternative route for exploiting the bone inductive activity of this class of molecules. To test the feasibility of this approach, we examined the osteogenic activity of AdCMV-BMP7, an adenovirus containing BMP7 cDNA under control of the CMV promoter that was constructed using Cre/lox recombination (Hardy et al. [1997] J. Virol. 71:1842-1849). Adenovirus vectors were shown to readily infect a wide variety of cell types in vitro including osteoblasts, fibroblasts, and myoblasts. COS7 cells transduced with AdCMV-BMP7 produced high levels of BMP-7 (approximately 0.5 microg/10(6) cells). Furthermore, transduction of C2C12 murine myoblast cells with AdCMVBMP-7 suppressed the muscle phenotype and induced in vitro osteoblast differentiation. To test its in vivo biological activity, AdCMV-BMP7 was mixed with a bovine bone derived collagen carrier (10(8) plaque-forming units virus/site) and was implanted into mouse muscle and dermal pouches. In both cases, an ossicle containing cortical and trabecular bone and a clearly defined marrow cavity formed at the site of virus implantation within 4 weeks. These data demonstrate that AdCMV-BMP7 transduced cells produce biologically active BMP-7 both in vitro and in vivo and show that gene therapy by direct viral transduction using a virus/matrix implant may be a viable route for stimulating bone regeneration. PMID- 10861846 TI - Overexpression of a secretory form of FGF-1 promotes MMP-1-mediated endothelial cell migration. AB - A coordinated interaction between fibroblast growth factors (FGFs) and matrix metalloproteinases (MMPs) is implicated in migration of microvascular endothelial cells (ECs), an early stage of angiogenesis. Specifically, we investigated microvascular ECs migration in vitro, which can be initiated by the overexpression of a secretory form of the angiogenic fibroblast growth factor-1 (FGF-1) and mediated through the enzymatic activity of matrix metalloproteinase-1 (MMP-1). MMP-1 is a member of the MMP family with a propensity for degradation of interstitial type I collagen. We stably overexpressed a chimeric FGF-1 construct composed of the FGF-4 signal-peptide gene, linked in-frame to the FGF-1 coding frame gene (sp-FGF-1), in cultured postcapillary venular ECs. The presence of the biologically active form of FGF-1 was readily detected in the conditioned medium of ECs transfected with sp-FGF-1 construct as demonstrated by DNA synthesis assay. The sp-FGF-1-, but not the plasmid vector alone-transfected ECs, exhibited an altered morphology as demonstrated by their conversion from a classic cobblestone form to a fibroblastlike shape that featured prominent neuritelike extensions. Addition of the anti-FGF receptor 1 antibody (FGFR1 Ab) reverted the transformed phenotype of sp-FGF-1 transfectants. This suggests that the resulting phenotypic transformation in sp-FGF-1 transfectants requires an uninterrupted interaction between the FGF-1 ligand and its receptor. We studied migration of cells through matrices of either highly pure collagen I or reconstituted basement membrane (matrigel) and found that sp-FGF-1-transfected cells migrated two times and six times faster than the vector control transfectants in the respective matrices. We further demonstrated that the enhanced migration rate of sp-FGF-1 transfected EC coincided with the induction of their MMP-1 mRNA level and increased enzymatic activity. The enhanced migratory activity of sp-FGF-1 could be blocked with a selective inhibitor of MMP-1. These results suggest that the multipotent FGF-1 plays a key role in the early stages of angiogenesis, by mediating MMP-1 proteolytic activity. PMID- 10861847 TI - Mercuric chloride stimulates distinct signal transduction pathway for DNA synthesis in a T-cell line, CTLL-2. AB - Exposure of an interleukin-2 (IL-2)-dependent murine T-cell line (CTLL-2) to mercuric chloride in in vitro culture induced a low but definite level of DNA synthesis in the absence of exogenous IL-2, and further enhanced the IL-2-induced DNA synthesis. Addition of anti-IL-2 or anti-IL-4 antibody to the culture, which neutralized all of the IL-2 or IL-4 activity, respectively, never inhibited the mercuric chloride-mediated DNA synthesis. Correspondingly, no detectable level of IL-2, IL-4, and IL-15 mRNA was found in mercuric chloride-treated CTLL-2 cells in our test condition. Stimulation of CTLL-2 cells with IL-2 induced phosphorylation on extracellular signal-regulated kinases more intensively than on c-Jun NH2 terminal kinases (JNKs), and provoked tyrosine phosphorylation of Janus kinases (JAKs) and signal transducers and activators of transcription (STATs). In contrast, by mercuric chloride stimulation, JNKs and c-Jun were preferentially phosphorylated, but no detectable level of phosphorylation was induced on JAKs and STATs. These findings provided a possibility that mercuric chloride promoted lymphocyte proliferation through a JNK-linked signal cascade in CTLL-2 cells, which differs from that triggered by IL-2. PMID- 10861848 TI - Construction of a cosmid library of DNA replicated early in the S phase of normal human fibroblasts. AB - We constructed a subgenomic cosmid library of DNA replicated early in the S phase of normal human diploid fibroblasts. Cells were synchronized by release from confluence arrest and incubation in the presence of aphidicolin. Bromodeoxyuridine (BrdUrd) was added to aphidicolin-containing medium to label DNA replicated as cells entered S phase. Nuclear DNA was partially digested with Sau 3AI, and hybrid density DNA was separated in CsCl gradients. The purified early-replicating DNA was cloned into sCos1 cosmid vector. Clones were transferred individually into the wells of 96 microtiter plates (9,216 potential clones). Vigorous bacterial growth was detected in 8,742 of those wells. High density colony hybridization filters (1, 536 clones/filter) were prepared from a set of replicas of the original plates. Bacteria remaining in the wells of replica plates were combined, mixed with freezing medium, and stored at -80 degrees C. These pooled stocks were analyzed by polymerase chain reaction to determine the presence of specific sequences in the library. Hybridization of high-density filters was used to identify the clones of interest, which were retrieved from the frozen cultures in the 96-well plates. In testing the library for the presence of 14 known early-replicating genes, we found sequences at or near 5 of them: APRT, beta-actin, beta-tubulin, c-myc, and HPRT. This library is a valuable resource for the isolation and analysis of certain DNA sequences replicated at the beginning of S phase, including potential origins of bidirectional replication. PMID- 10861849 TI - Human monocyte/neutrophil elastase inhibitor (MNEI) is regulated by PU.1/Spi-1, Sp1, and NF-kappaB. AB - Human monocyte/neutrophil elastase inhibitor (MNEI) is a specific inhibitor of the neutrophil azurophil granule proteases including elastase. To understand the physiological mechanisms that regulate expression of MNEI, we dissected a 1.0 kb region upstream of exon 1. On transient transfection, promoter activity of MNEI luciferase constructs was highest in U937 myeloid cells, followed by K562 hematopoietic cells, followed by HeLa cervical carcinoma cells, indicating that the MNEI promoter is most active in myeloid cells and is also active in non myeloid cells. Three transcription factor binding elements, which confer the majority of activity, are located within the first 180 base pairs of the promoter, one of which, located at -128, was active in U937 and K562 cells but inactive in non-myeloid HeLa cells. The three proximal elements were identified by transient transfection, mutation, gel shift and competition assays as Sp1 at 170, PU.1/Spi-1 at -128, and Sp1 at -66. The trans-acting factors that bind and control these elements were detected, and their identity confirmed by antibody supershift assays. Further upstream at -821, an additional regulatory element was identified controlled by NF-kappaB, which supports the highest levels of MNEI transcriptional activity. In U937 cells, reporter gene expression by the MNEI luciferase construct that included the NF-kappaB element was two- to three-fold greater than the construct without the element. In addition, treatment of myeloid cells with lipopolysaccharide, a complex glycolipid of gram-negative bacteria, activated NF-kappaB to bind the -821 element, together suggesting that enhancement of expression of the anti-inflammatory MNEI gene is linked to innate immune responses to bacterial infection. PMID- 10861850 TI - Expression of replication factor C 40-kDa subunit is down-regulated during neonatal development in rat ventricular myocardium. AB - During neonatal development, cardiac myocytes undergo a transition from hyperplastic to hypertrophic growth. Whether these cells are terminally differentiated and permanently withdrawn from the cell cycle shortly after birth is controversial. Nevertheless, the clinical observation that functionally significant myocardial regeneration has not been documented in cardiovascular disease or injury during adulthood seems to support the notion that the vast majority of cardiac myocytes do not proliferate once they differentiate. Regardless of the controversy, the elucidation on how mitosis is blocked in cardiac myocytes may facilitate development of new cardiovascular therapies, based on the regeneration of the adult myocardium. To better understand postnatal myocardial development, we performed suppression subtractive hybridization to isolate genes that are differentially expressed in day one or day seven postnatal rat ventricular myocardium. Here we report the down-regulated mRNA expression of the 40-kDa subunit of replication factor C (RFC p40 or RFC2), which is an essential processive factor for proliferating cellular nuclear antigen-dependent DNA replication during neonatal myocardial development. PMID- 10861851 TI - Role of endogenous regucalcin in the regulation of Ca(2+)-ATPase activity in rat liver nuclei. AB - The role of endogenous regucalcin in the regulation of Ca(2+)-ATPase, a Ca(2+) sequestrating enzyme, in rat liver nuclei was investigated. Nuclear Ca(2+)-ATPase activity was significantly reduced by the addition of regucalcin (0.1-0.5 microM) into the enzyme reaction mixture. The presence of anti-regucalcin monoclonal antibody (25 or 50 ng/ml) caused a significant elevation of Ca(2+)-ATPase activity; this effect was completely abolished by the addition of regucalcin (0.1 microM). The effect of anti-regucalcin antibody (50 ng/ml) in increasing Ca(2+) ATPase activity was completely prevented by the presence of thapsigargin (10(-6) M), an inhibitor of Ca(2+) sequestrating enzyme, N-ethylmaleimide (1 mM), a modifying reagent of thiol groups, or vanadate (10(-5) M), an inhibitor of phosphorylation of the enzyme by ATP, which revealed an inhibitory effect on nuclear Ca(2+)-ATPase activity. Meanwhile, the effect of anti-regucalcin antibody (50 ng/ml) was significantly enhanced by the addition of calmodulin (5 microg/ml), which could increase nuclear Ca(2+)-ATPase activity. In addition, the effect of antibody (50 ng/ml) was significantly reduced by the presence of trifluoperazine (20 microM), an antagonist of calmodulin. These results suggest that the endogenous regucalcin in liver nuclei has a suppressive effect on nuclear Ca(2+)-ATPase activity, and that regucalcin can inhibit an activating effect of calmodulin on the enzyme. PMID- 10861852 TI - Hydrogen peroxide-mediated, lysyl oxidase-dependent chemotaxis of vascular smooth muscle cells. AB - Lysyl oxidase (LO), an enzyme secreted by vascular smooth muscle cells (VSMC), initiates the covalent crosslinking of polypeptide chains within collagen and elastin. The present study reveals that purified LO strongly induces directional migration of VSMC in an in vitro assay system. LO-dependent chemotaxis, but not chemokinesis, was abolished by beta-aminopropionitrile, an active site inhibitor of LO, or by catalase, as well as by prior heat denaturation. This indicates that the H(2)O(2) product of amine oxidation by LO is critical to the expression of its chemotactic activity. The results indicate that the chemotactic response requires direct access between LO and a substrate molecule (or molecules) tightly associated with the VSMC. The addition of LO to VSMC elevated the levels of intracellular H(2)O(2), enhanced stress fiber formation, and focal adhesion assembly, is consistent with the induction of the chemotactic response. PMID- 10861853 TI - Interaction of hCLIM1, an enigma family protein, with alpha-actinin 2. AB - Enigma proteins are proteins that possess a PDZ domain at the amino terminal and one to three LIM domains at the carboxyl terminal. They are cytoplasmic proteins that are involved with the cytoskeleton and signal transduction pathway. By virtue of the two protein interacting domains, they are capable of protein protein interactions. Here we report a study on a human Enigma protein hCLIM1, in particular. Our study describes the interaction of the human 36 kDa carboxyl terminal LIM domain protein (hCLIM1), the human homologue of CLP36 in rat, with alpha-actinin 2, the skeletal muscle isoform of alpha-actinin. hCLIM1 protein was shown to interact with alpha-actinin 2 by yeast two-hybrid screening and immunochemical analyses. Yeast two-hybrid analyses also demonstrated that the LIM domain of hCLIM1 binds to the EF-hand region of alpha-actinin 2, defining a new mode of LIM domain interactions. Immunofluorescent study demonstrates that hCLIM1 colocalizes with alpha-actinin at the Z-disks in human myocardium. Taken together, our experimental results suggest that hCLIM1is a novel cytoskeletal protein and may act as an adapter that brings other proteins to the cytoskeleton. PMID- 10861854 TI - Relative abundance of different cadherins defines differentiation of mesenchymal precursors into osteogenic, myogenic, or adipogenic pathways. AB - Cadherins, a family of cell-cell adhesion molecules, provide recognition signals that are important for cell sorting and aggregation during tissue development. This study was performed to determine whether distinct cadherin repertoires define tissue-specific lineages during differentiation of immature C3H10T1/2 and C2C12 mesenchymal cells. Both cell lines expressed mRNA for N-cadherin (N-cad), cadherin-11 (C11), and R-cadherin (R-cad). After induction of osteogenesis by recombinant human BMP-2 (rhBMP-2) treatment, steady state N-cad mRNA slightly increased in C3H10T1/2 cells. Likewise, the abundance of C11 mRNA increased in both cell lines, although the changes were more remarkable in C2C12 cells. By contrast, R-cad expression was almost shut off by rhBMP-2. The immature but committed osteoblastic MC3T3-E1 cells exhibited only minor changes in N-cad and C11 mRNA abundance after rhBMP-2 treatment. Whereas adipogenic differentiation was associated with a net decrease of N-cad and C11 expression in C3H10T1/2 cells, induction of myogenesis in C2C12 cells resulted in up-regulation of N-cad, while R-cad mRNA became undetectable in either case. Similarly, the adipocytic 3T3-L1 cells expressed very low levels of all cadherins when fully differentiated. Therefore, the repertoire of cadherins present in undifferentiated mesenchymal cells undergoes distinct changes during transition to mature cell phenotypes. Although neither N-cad nor C11 represent strict tissue specific markers, the relative abundance of these mesenchymal cadherins defines lineage-specific signatures, perhaps providing recognition signals for aggregation and differentiation of committed precursors. PMID- 10861855 TI - Glutathione is a factor of resistance of Jurkat leukemia cells to nitric oxide mediated apoptosis. AB - We have previously reported that nitric oxide (NO) stimulates apoptosis in different human neoplastic lymphoid cell lines through mitochondrial damage (including degradation of cardiolipin, a major mitochondrial lipid) followed by activation of caspases. Here we demonstrate that Jurkat human leukemia cells which survive after 24 h treatment with NO form subpopulations with higher and lower cardiolipin content (designated as NAO(high) and NAO(low), respectively). Sorted NAO(high) cells were found to survive in culture whereas sorted NAO(low) cells died. Moreover, NAO(high) cells acquired an increased resistance to the exposure to NO donors which remained unchanged during long-term culture. These cells showed a similar cardiolipin content and expressed the same level of anti apoptotic proteins Bcl-2 and Bcl-x(L) as APO-S unsorted cells but contained significantly higher concentration of the antioxidant glutathione. Depletion of glutathione in these cells with buthionine-sulfoximine (BSO) correlated with a significant stimulation of NO-mediated apoptosis whereas the exposure of NO sensitive APO-S cells to the glutathione precursor N-acetylcysteine (NAC) resulted in a substantial suppression of this effect. Our data suggest a complex mechanism of the resistence to NO-induced apoptosis in Jurkat human leukemia cells in which glutathione plays an important role. PMID- 10861856 TI - TGF-beta signaling in A549 lung carcinoma cells: lipid second messengers. AB - Transforming growth factor-beta (TGF-beta) is a potent inducer of numerous extracellular matrix components, largely through a transcriptional mechanism. To define the postreceptor signaling pathways used by TGF-beta in the induction of extracellular matrix gene expression, we have utilized the human lung carcinoma cell line, A549, in transfection experiments with the TGF-beta inducible reporter construct, p3TP-Lux. Previous work from this laboratory using pharmacologic agents suggested that a phosphatidylcholine-specific phospholipase C and protein kinase C may be involved in early aspects of TGF-beta signaling. Here we provide evidence that TGF-beta induces a rapid and transient increase in diacylglycerol (DAG) production. When cells transfected with the p3TP-Lux reporter plasmid are simultaneously treated with TGF-beta and a DAG kinase inhibitor, we observed a higher level of luciferase than with TGF-beta alone. We also find elevated levels of phosphocholine in cells following TGF-beta treatment. Further, exogenously added bacterial phosphatidylcholine phospholipase C (PC-PLC) is capable of inducing expression of the p3TP-Lux reporter to the same extent as TGF-beta indicating that the bacterial PC-PLC can mimic the TGF-beta effect. In contrast, neither hexanoyl sphingosine (a ceramide analogue) nor arachadonic acid induce expression of the p3TP-Lux reporter. Measurements with the fluorescent, calcium sensitive dye, FURA2, indicated that there was no change in intracellular calcium in response to TGF-beta. Furthermore, buffering intracellular calcium with the calcium chelating agent BAPTA/AM failed to block TGF-beta induction of the p3TP Lux reporter. Thus the TGF-beta signaling pathway appears to involve the production of diacylglycerol but is independent of calcium. PMID- 10861857 TI - Selenoorganic compound, ebselen, inhibits nitric oxide and tumor necrosis factor alpha production by the modulation of jun-N-terminal kinase and the NF-kappab signaling pathway in rat Kupffer cells. AB - In response to the bacterial endotoxin, LPS, Kupffer cells are induced to express NO and TNF-alpha. These compounds are involved in hepatic inflammation/injury, especially that associated with endotoxic shock. In this study, we demonstrate that ebselen (2-phenyl-1,2-benzisoselenazol-3[2H]one), a selenoorganic compound, blocks LPS-induced NO and TNF-alpha production by cultured rat liver Kupffer cells. LPS can activate both the NF-kappaB signaling pathway and MAPK signal transduction pathways such as JNK and p38 MAPK. We find that ebselen inhibits LPS induced NF-kappaB nuclear translocalization, and also suppresses the LPS-induced phosphorylation of JNK, but not the phosphorylation of p38 MAPK. This inhibition of signal transduction leads to a decrease in the transcription of TNF-alpha and the inducible isoform of NO. Furthermore, ebselen inhibits LPS-induced COX-2 expression, which is responsible for proinflammatory prostaglandin production, without affecting constitutive COX-1 expression. These data suggest the mechanism by which ebselen acts as an antiinflammatory agent, and also suggest that ebselen may be potent in preventing hepatic injury such as endotoxic shock, in which Kupffer cell activation has been implicated. PMID- 10861858 TI - Secretory products from PC-3 and MCF-7 tumor cell lines upregulate osteopontin in MC3T3-E1 cells. AB - Tumor cells frequently have pronounced effects on the skeleton including bone destruction, bone pain, hypercalcemia, and depletion of bone marrow cells. Despite the serious sequelae associated with skeletal metastasis, the mechanisms by which tumor cells alter bone homeostasis remain largely unknown. In this study, we tested the hypothesis that the disruption of bone homeostasis by tumor cells is due in part to the ability of tumor cells to upregulate osteopontin (OPN) mRNA in osteoblasts. Conditioned media were collected from tumor cells that elicit either osteolytic (MCF-7, PC-3) or osteoblastic responses (LNCaP) in animal models and their effects on OPN gene expression were compared using an osteoblast precursor cell line, MC3T3-E1 cells. Secretory products from osteolytic but not osteoblastic tumor cell lines were demonstrated to upregulate OPN in osteoblasts while inhibiting osteoblast proliferation and differentiation. Signal transduction studies revealed that regulation of OPN was dependent on both protein kinase C (PKC) and the mitogen-activated protein (MAP) kinase cascade. These results suggest that the upregulation of OPN may play a key role in the development of osteolytic lesions. Furthermore, these results suggest that drugs that prevent activation of the MAP kinase pathway may be efficacious in the treatment of osteolytic metastases. PMID- 10861859 TI - Opposing early inhibitory and late stimulatory effects of insulin-like growth factor-I on myogenin gene transcription. AB - Insulinlike growth factors (IGFs) stimulate skeletal muscle cell differentiation in association with an increase in the mRNA of myogenin, a member of the MyoD family of skeletal muscle-specific transcription factors that plays an essential role in the differentiation process. However, this is a relatively late effect, requiring treatment periods of >24 h. In contrast, IGFs initially inhibit skeletal muscle cell differentiation, associated with a marked reduction in myogenin mRNA. The mechanisms by which IGF-I initially inhibits and subsequently stimulates myogenin expression are unknown. In the first 24 h, we find that IGF-I inhibits myogenin gene transcription by >80% but has no effect on myogenin mRNA stability. Similarly, in the first 24 h, IGF-I markedly inhibits myogenin promoter activity; the sequence -145 to -9 of the myogenin gene is sufficient to confer this inhibitory effect of IGF-I. In contrast, 48 h of treatment with IGF-I results in an increase in myogenin promoter activity that parallels the increase in myogenin steady-state mRNA. This increase in promoter activity is completely prevented in constructs lacking the sequence -1,565 to -375 of the myogenin gene. These data indicate that the early inhibitory and late stimulatory effects of IGF I on myogenin expression are mediated at the level of transcription, and that these time-dependent, opposing effects of IGF-I on myogenin transcription are mediated by distinct regions of the myogenin gene. To our knowledge, this is the first demonstration of a gene whose promoter activity is initially inhibited and subsequently stimulated by IGF-I. PMID- 10861860 TI - Proteinase expression during differentiation of human osteoclasts in vitro. AB - Osteoclasts are macrophage-derived polykaryons that degrade bone in an acidic extracellular space. This differentiation includes expression of proteinases and acid transport proteins, cell fusion, and bone attachment, but the sequence of events is unclear. We studied two proteins expressed at high levels only in the osteoclast, cathepsin K, a thiol proteinase, and tartrate-resistant acid phosphatase (TRAP), and compared this expression with acid transport and bone degradation. Osteoclastic differentiation was studied using human apheresis macrophages cocultured with MG63 osteosarcoma cells, which produce cytokines including RANKL and CSF-1 that mediate efficient osteoclast formation. Immunoreactive cathepsin K appeared at 3-5 days. Cathepsin K activity was seen on bone substrate but not within cells, and cathepsin K increased severalfold during further differentiation and multinucleation from 7 to 14 days. TRAP also appeared at 3-5 d, independently of cell fusion or bone attachment, and TRAP activity reached much higher levels in osteoclasts attached to bone fragments. Two proteinases that occur in the precursor macrophages, cathepsin B, a thiol proteinase related to cathepsin K, and an unrelated lysosomal aspartate proteinase, cathepsin D, were also studied to determine the specificity of the differentiation events. Cathepsin B occurred at all times, but increased two- to threefold in parallel with cathepsin K. Cathepsin D activity did not change with differentiation, and secreted activity was not significant. In situ acid transport measurements showed increased acid accumulation after 7 days either in cells on osteosarcoma matrix or attached to bone, but bone pit activity and maximal acid uptake required 10-14 days. We conclude that TRAP and thiol proteinase expression begin at essentially the same time, and precede cell fusion and bone attachment. However, major increases in acid secretion and proteinases expression continue during cell fusion and bone attachment from 7 to 14 days. PMID- 10861861 TI - Glycosylated nuclear lectin CBP70 also associated with endoplasmic reticulum and the Golgi apparatus: does the "classic pathway" of glycosylation also apply to nuclear glycoproteins? AB - The subcellular plurilocalization of some lectins (galectin-1, galectin-3, galectin-10, calreticulin, etc.) is an intriguing problem, implying different partners according to their localization, and involvement in a variety of cellular activities. For example, the well-known lectin, galectin-3, a lactose binding protein, can act inside the nucleus in splicing events, and at the plasma membrane in adhesion, and it was demonstrated that galectin-3 interacts in the cytoplasm with Bcl-2, an antiapoptotic protein. Some years ago, our group isolated a nuclear lectin CBP70, capable of recognizing N-acetylglucosamine residues. This lectin, first isolated from the nucleus of HL60 cells, was also localized in the cytoplasm. It has been demonstrated that CBP70 is a glycosylated lectin, with different types of glycosylation, comparing cytoplasmic and nuclear forms. In this article, we have studied the localization of CBP70 in undifferentiated HL60 cells by electron microscopy, immunofluorescence analysis, and subcellular fractionation. The results obtained clearly demonstrated that CBP70 is a plurilocalized lectin that is found in the nucleus, at the endoplasmic reticulum, the Golgi apparatus, and mitochondria, but not at the plasma membrane. Because CBP70, a nuclear glycoprotein, was found to be associated also with the endoplasmic reticulum and the Golgi apparatus where the glycosylation take place, it raised the question: where does the glycosylation of nuclear proteins occur? PMID- 10861862 TI - Protein-protein interactions that precede the nuclear entry of goat uterine estrogen receptor under cell-free conditions. AB - Mechanisms associated with a regulated nuclear entry of the goat uterine estrogen receptor (gER), under the influence of estradiol, have been examined using a cell free system. The gER transport into the nucleus is mediated by a 55-kDa cytosolic protein, p55. Experimental evidence has been provided to demonstrate that p55 binds to the nuclear localization signals (NLS) on the gER. Under hormone-free conditions, a 28-kDa protein, p28, binds to the NLS and prevents the p55 interaction with the NLS. This inhibition is reversed by a 73-kDa protein, p73. It appears that p73 associates with the hormone binding domain (HBD) of the gER under hormone-free conditions. Estradiol binding to the HBD facilitates p73 interaction with p28. This leads to the dissociation of p28 from the NLS, which, in turn, facilitates the binding of p55 to the NLS on the gER. Both p28 and p55 cross-react with monoclonal antibodies against hsp-25 and hsp-70, indicating a possibility that p28 and p55 belong to a superfamily of molecular chaperones. J. Cell. Biochem. 78:650-665, 2000. PMID- 10861863 TI - Somatostatin analog SMS 201995 inhibits proliferation in human leukemia T-cell line: relevance of the adenylyl cyclase stimulation. AB - Octreotide SMS 201995 is a stable somatostatin (SRIF) analog with potent antiproliferative actions in numerous cell types including normal T lymphocytes. It is currently used in the clinical treatment of different malignancies. However, the possible beneficial actions of octreotide in T-cell leukemia have not been addressed before, although these cells express SRIF receptors. For instance, human leukemia Jurkat T cells have been shown to express a single SRIF receptor isotype: sst3 that can be pharmacologically targeted by octreotide. In this study, we therefore studied SMS 201995 effects on in vitro [(3)H CH3]thymidine incorporation in Jurkat T cells. Our data show that octreotide inhibits the proliferation of Jurkat cells both in the absence and in the presence of mitogens. By contrast, SRIF28, an endogenous SRIF analog sharing with SMS 201995 an almost identical affinity for somatostatin sst3 receptors, increases [(3)H-CH3]thymidine uptake in both mitogen-activated and nonactivated cells. To assess the mechanisms of the opposite actions of these two analogs on leukemia T-cell proliferation, we next studied their effects on adenylyl cyclase activity in whole Jurkat cells. At least in the presence of mitogens, SMS 201995 significantly enhances the adenylyl cyclase activity whereas SRIF28 inhibits it. Taken together these data are in accordance with the current hypothesis according to which increase and decrease in cAMP production are required to allow the inhibition and stimulation of T-cell proliferation, respectively. They also point to a potential therapeutic benefit of SMS 201995 in the management of human T cell leukemia. PMID- 10861864 TI - Hypoxia induces differential expression of the integrin receptors alpha(vbeta3) and alpha(vbeta5) in cultured human endothelial cells. AB - The integrins alpha(vbeta3) and alpha(vbeta5) have been implicated in playing a key role in the process of angiogenesis. In this study, we examined the effects of hypoxia, an important stimulus of angiogenesis, on the differential expression of the integrin subunits beta(3) and beta(5). beta(3) and beta(5) messenger RNA (mRNA), protein levels, and alpha(v)beta(3) function were measured in human umbilical vein endothelial cells (HUVECs) cultured under normoxic and hypoxic (1% O(2)) conditions. Cells exposed to hypoxic conditions for up to 72 h showed gradually increased mRNA levels of alpha(V) and beta(3), peaking at 24 h, in comparison with cells cultured under normoxic conditions. However, beta(5) mRNA levels, under the same hypoxic conditions, remained at a constant level. Results from Western blot analysis of HUVECs, cultured under hypoxic conditions, paralleled those of the Northern analysis with an increased expression in alpha(v)beta(3) protein levels, measured by blotting with LM609, evident by 24 h. alpha(v)beta(5) protein levels, measured by blotting with P1F6, did not change for up to 72 h. HUVECs cultured under hypoxic conditions for 72 h showed increased attachment to fibrinogen, an alpha(v)beta(3) mediated process. These results indicate that hypoxia can increase expression of alpha(v)beta(3) in HUVECs, and that hypoxic regulation of alpha(v)beta(3) may be an important regulator of angiogenesis. PMID- 10861865 TI - Differential susceptibility of normal and PARP knock-out mouse fibroblasts to proteasome inhibitors. AB - Recently we found a clearly reduced basal level of wt p53 protein in PARP deficient cells. Interestingly, PARP deficiency affected only regularly spliced (RS) wt p53. No significant difference of the p53 transcription rate was observed between wt and PARP-lacking cells. To clarify whether the reduction of RS p53 protein is due to a lower translation rate or rather to its instability in the absence of functional PARP, we investigated the effect of the inhibition of proteasome activity and nuclear export on the p53 level. The p53 half-life was approximately eight-fold decreased in PARP-lacking cells. Surprisingly, treatment with three proteasome inhibitors increased RS p53 in normal but not in PARP deficient cells. However, the inhibition of nuclear export resulted in a considerable accumulation of RS p53 in the latter. Therefore, we decided to increase concentrations of the inhibitors. Their higher concentrations strongly affected viability of normal, but not of PARP-deficient cells, about 70% of MEFs died. Interestingly, higher concentrations of proteasome inhibitors resulted in the appearance of RS p53 in PARP-lacking fibroblasts. Reconstitution of PARP deficient cells with PARP restored the normal susceptibility to proteasome inhibitors thereby unequivocally demonstrating that the enhanced cytotoxicity of proteasome inhibitors and their action on p53 level depends on the presence of functional PARP. PMID- 10861866 TI - Fungi as elicitors of insect immune responses. PMID- 10861867 TI - Adult Colorado potato beetles, Leptinotarsa decemlineata compensate for nutritional stress on oryzacystatin I-transgenic potato plants by hypertrophic behavior and over-production of insensitive proteases. AB - Protease inhibitors have been proposed as potential control molecules that could be engineered into potato plants for developing crops resistant to the Colorado potato beetle, Leptinotarsa decemlineata, a major pest of potato and other Solanaceae. In this study, we examined the effects of feeding young female beetles with foliage from a cultivar of the "Kennebec" potato line (K52) transformed with a gene encoding oryzacystatin I (OCI), a specific cysteine proteinase inhibitor with proven activity against cathepsin H-like enzymes of larvae and adults of the potato beetle. To evaluate the insect's performance, we collected data over a 16-d postemergence period on survival, diapause incidence, foliage consumption, weight gain, and oviposition of females. Tested individuals were fed untransformed (control) and OCI-transformed foliage at two stages of potato leaf differentiation, corresponding to "low" and "high" levels of OCI expression in leaves of K52. The OCI-expressing foliage did not affect female survival (close to 100%), incidence of diapause (15-30%), relative growth rate (RGR) during postemergence growth (5-9% d(-1)) or maximum weight reached (140-160 mg). Neither did it affect female reproductive fitness as measured by preoviposition time (8-9 d), 16-d fecundity (220-290 eggs), or egg eclosion incidence (86-91%). However, nutritional stress to females feeding on OCI foliage was evident, as reflected in their lower efficiency of conversion of ingested foliage (ECI) during postemergence growth, increased foliage consumed per egg laid (up to 119% more), and adaptation of their digestive proteolytic system to the inhibitory effect of OCI. Interestingly, beetles fed foliage expressing the highest level of OCI reacted rapidly to the presence of OCI by producing OCI insensitive proteases, and exhibiting strong hypertrophic behavior by ingestion of 2.4-2.5 times more OCI rich foliage apparently as a compensatory response for nutritional stress due to the protease inhibitor in their diet. PMID- 10861868 TI - Effect of whitefly parasitoids on the cuticular lipid composition of Bemisia argentifolii (Homoptera: aleyrodidae) nymphs. AB - Experiments were conducted to determine the effects of whitefly parasitoids on the cuticular lipid composition of the silverleaf whitefly, Bemisia argentifolii Bellows and Perring [=sweetpotato whitefly, Bemisia tabaci (Gennadius), Biotype B] nymphs. The cuticular lipids of B. argentifolii nymphs that had been attacked by parasitic wasps, either Eretmocerus mundus Mercet or Encarsia pergandiella Howard, were characterized by capillary gas chromatography and CGC-mass spectrometry and the results compared with the cuticular lipids of unparasitized nymphs. Previous studies with B. argentifolii nymphs had shown that wax esters were the major components of the cuticular lipids with lesser amounts of hydrocarbons, long-chain aldehydes, and long-chain alcohols. No appreciable changes in lipid composition were observed for the cuticular lipids of E. pergandiella-parasitized nymphs as compared to unparasitized controls. However, the cuticular lipids from nymphs parasitized by E. mundus contained measurable quantities of two additional components in their hydrocarbon fraction. Analyses and comparisons with an authentic standard indicated that the two hydrocarbons were the even-numbered chain length methyl-branched alkanes, 2-methyltriacontane and 2-methyldotriacontane. The occurrences and possible functions of 2 methylalkanes as cuticular lipid components of insects are discussed and specifically, in regard to host recognition, acceptance, and discrimination by parasitoids. Published 2000 Wiley-Liss, Inc. PMID- 10861869 TI - Carbohydrate metabolism during starvation in the silkworm Bombyx mori. AB - The effect of starvation on carbohydrate metabolism in the last instar larvae of the silkworm Bombyx mori was examined. Trehalose concentration in the hemolymph increased slightly during the first 6 h of starvation and decreased thereafter, whereas glucose concentration decreased rapidly immediately after diet deprivation. Starvation-induced hypertrehalosemia was completely inhibited by neck ligation, suggesting that starvation stimulates the release of a hypertrehalosemic factor(s) from the head. The percentage of active glycogen phosphorylase in the fat body increased within 3 h of starvation and its glycogen content decreased gradually. These observations suggest that production of trehalose from glycogen is enhanced in starved larvae. However, hypertrehalosemia during starvation cannot be explained by the increased supply of trehalose into hemolymph alone, as similar changes in phosphorylase activity and glycogen content in the fat body were observed in neck-ligated larvae, in which hemolymph trehalose concentration did not increase but decreased gradually. When injected into larvae, trehalose disappeared from hemolymph at a rate about 40% lower in starved larvae than neck-ligated larvae. The hemolymph lipid concentration increased during starvation, suggesting that an increased supply of lipids to tissues suppresses the consumption of hemolymph trehalose and this is an important factor in hypertrehalosemia. PMID- 10861870 TI - PRESTO-SENSE: an ultrafast whole-brain fMRI technique. AB - A new ultrafast MR imaging method is proposed and tested, which enables whole brain fMRI with a true temporal resolution of 1 sec. The method combines a 3D PRESTO pulse sequence with the concept of sensitivity-encoding with multiple receiver coils (SENSE). The so-called PRESTO-SENSE technique is demonstrated on a set of functional block-type motor and visual experiments and compared with conventional functional imaging techniques, such as PRESTO and EPI. Comparable image quality and activation areas are found with all sequences. The noise characteristics of the proposed method are analyzed in detail and their implications for ultrafast fMRI studies are discussed. Magn Reson Med 43:779-786, 2000. PMID- 10861871 TI - Interleaved spiral cine coronary artery velocity mapping. AB - Navigator-echo controlled interleaved spiral cine coronary-artery velocity maps were acquired in eight free-breathing volunteers. Good quality data were achieved in all subjects with phasic flow being clearly demonstrated. The improved efficiency of spiral k-space coverage compared to that of more conventional spin warp techniques resulted in a mean scan time for 256 x 256 pixel-matrix studies of 94 sec (range 69 - 123 sec, SD = 20 sec), the navigator acceptance rate being typically 40-45%. Repeat scans in five subjects showed a high degree of reproducibility, with no significant differences occurring in the peak velocities (14.6+/-1.4 cm/sec vs. 14.5+/-1.5 cm/sec, P = ns) nor in the mean velocities measured over all cine phases (8.3+/-2.1 cm/sec vs. 8.7+/-1.6 cm/sec, P = ns). The potential benefits of using this sequence for assessing coronary blood flow and flow reserve are discussed. Magn Reson Med 43:787-792, 2000. PMID- 10861872 TI - Imaging pulmonary blood flow and perfusion using phase-sensitive selective inversion recovery. AB - A technique is described for imaging pulmonary blood flow using a phase-sensitive selective inversion recovery (PS-SIR) sequence. PS-SIR image reconstruction provides excellent contrast, differentiating fully relaxed inflowing blood from inverted blood and lung tissue. The magnetization of the inverted tissues remains negative at any inversion delay less than that at which the magnetization of the lung tissue is nulled, whereas that of the fully relaxed inflowing blood is always positive. Pulmonary blood flow can be observed by tracking the propagation of the pixels with positive values. Five healthy volunteers were imaged. The normal pattern of blood flow advancing from the central arteries toward the peripheries and into the lung parenchyma with return toward the center via draining veins was depicted. The method offers promise for evaluating pulmonary blood flow without the need for image subtraction or contrast administration. Magn Reson Med 43:793-795, 2000. PMID- 10861873 TI - Temperature monitoring of internal body heating induced by decoupling pulses in animal (13)C-MRS experiments. AB - A temperature monitoring method to promote safety with regard to tissue heating induced by RF irradiation during MRI procedures, especially carbon-13 magnetic resonance spectroscopy ((13)C-MRS), is proposed. The method is based on the temperature dependence of the water proton chemical shift (-0.01 ppm/ degrees C) combined with phase mapping. Using this method, temperature changes were measured in rats (n = 4) employing practical (1)H-decoupled (13)C-MRS pulse sequences for 1D projections (TR = 1000 ms, acquisition time = 15 ms, matrix = 256, spatial resolution = 0.2 mm) and 2D images (TR = 1500 ms, acquisition time = 840 ms, matrix = 128x32, spatial resolution = 0.8x1.5 mm). Measurement error was 0.18 degrees C (SD) for 1D acquisition and 0.39 degrees C (SD) for 2D acquisition, demonstrating the feasibility of this temperature mapping method. Further studies should be conducted in human subjects to monitor patient safety and to optimize the pulse sequences employed. Magn Reson Med 43:796-803, 2000. PMID- 10861874 TI - Electron paramagnetic resonance oxygen mapping (EPROM): direct visualization of oxygen concentration in tissue. AB - Tissue oxygen content is a central parameter in physiology but is difficult to measure. We report a novel procedure for spatial mapping of oxygen by electron paramagnetic resonance (EPR) utilizing a spectral-spatial imaging data set, in which an EPR spectrum is obtained from each image volume element. From this data set, spatial maps corresponding to local spin density and maximum EPR spectral line amplitude are generated. A map of local EPR spectral linewidth is then computed. Because linewidth directly correlates with oxygen concentration, the linewidth image provides a map of oxygenation. This method avoids a difficulty inherent in other oxygen content mapping techniques using EPR, that is, the unwanted influence of local spin probe density on the image. We provide simulation results and data from phantom studies demonstrating the validity of this method. We then apply the method to map oxygen content in rat tail tissue and vasculature. This method provides a new, widely applicable, approach to direct visualization of oxygen concentration in living tissue. Magn Reson Med 43:804-809, 2000. PMID- 10861875 TI - Corrections for off-resonance effects and incomplete saturation in conventional (two-site) saturation-transfer kinetic measurements. AB - The effects of off-resonance irradiation and incomplete saturation on saturation transfer measurements of chemical-exchange rates are discussed. With off resonance effects there is no exact formula for the exchange rate constant from spin D to spin E, k(DE), in terms of observable signal intensities and relaxation rates. However, k(DE) can be estimated by measuring the effective spin-lattice relaxation rate constant of spin D when spin E is saturated, *R(1), plus signal intensities with no RF irradiation, M(0D) and M(0E); with irradiation of a control position, M'(D) and M'(E); and with saturation of spin E, *M(D) and *M(E). Several formulas are compared and the best formula for calculating k(DE) appears to be either k(DE) = *R(1) [(M'(D)- *M(D))/M(0D)]/[(M'(E) - *M(E))/M(0E)], or the same formula with M(0D) and M(0E) replaced by M'(D) and M'(E). These formulas are exact with incomplete saturation and no off-resonance effects, and are better than previously published formulas when off-resonance effects are present. More accurate formulas are available if signal intensities and relaxation rates can be measured while the exchange process is stopped. Magn Reson Med 43:810-819, 2000. PMID- 10861876 TI - Simultaneous quantitative cerebral perfusion and Gd-DTPA extravasation measurement with dual-echo dynamic susceptibility contrast MRI. AB - Quantification of cerebral perfusion using dynamic susceptibility contrast MRI generally relies on the assumption of an intact blood-brain barrier. The present study proposes a method to correct the tissue response function that does not require this assumption, thus, allowing perfusion studies in, for example, high grade brain tumors. The correction for contrast extravasation in the tissue during the bolus passage is based on a two-compartment kinetic model. The method separates the intravascular hemodynamic response and the extravascular component and returns the corrected tissue response function for perfusion quantification as well as the extravasation rate constant of the vasculature. Results of simulation experiments with different degrees of contrast extravasation are presented. The clinical potential is illustrated by determination of the perfusion and extravasation of a glioblastoma multiforme. The correction scheme proves to be fast and reliable even in cases of low signal-to-noise ratio. It is applicable whether extravasation occurs or not. When extravasation is present, application of the proposed method is mandatory for accurate cerebral blood volume measurements. Magn Reson Med 43:820-827, 2000. PMID- 10861877 TI - Measurement of cell density and necrotic fraction in human melanoma xenografts by diffusion weighted magnetic resonance imaging. AB - The aim of this study was to investigate whether apparent diffusion coefficients (ADCs) could be used as measures of cell density and necrotic fraction of tumors. Tumors of four human melanoma xenograft lines were subjected to diffusion weighted magnetic resonance imaging (DWI). ADCs were calculated from the images and related to cell density and necrotic fraction, as determined from histological sections. A significant correlation was found between the ADC of the viable tissue and cell density, regardless of whether tumors of different lines or different regions within individual tumors were considered. Necrosis was found in two of the lines. A single region of massive necrosis that could be differentiated from the viable tissue in ADC maps was found in one line, whereas a number of smaller necrotic regions that could not be identified in ADC maps were found in the other line. Tumor ADC was significantly correlated with the necrotic fraction of the former, but not of the latter line. Our results suggest that ADCs can be used as measures of cell density and necrotic fraction of some but not of all tumors, depending on whether the individual necrotic regions are large enough to be differentiated from the viable tissue with the obtained spatial resolution of the DW images. Magn Reson Med 43:828-836, 2000. PMID- 10861878 TI - Compartmental study of diffusion and relaxation measured in vivo in normal and ischemic rat brain and trigeminal nerve. AB - The correlation between the apparent diffusion coefficient (ADC) and T(2) of water in rat brain and trigeminal nerve was investigated using a hybrid diffusion weighted-CPMG imaging sequence. Little dependence of ADC on T(2) was found in brain regions of interest, which is postulated to be due to rapid exchange between intra- and extracellular water. Conversely, the ADC of water in trigeminal nerve was found to change significantly with echo time (TE). Parallel to the nerve and with a constant diffusion time (t(diff) = 10.8 ms), the ADC increased by approximately 30% between TEs of 25 ms and 185 ms; perpendicular to the nerve, the ADC decreased by a similar amount over the same range of TE. Measurements made following the onset of global ischemia yielded lower ADCs, with similar dependence on TE. Observations that transverse relaxation of water in nerves is multiexponential have previously been interpreted in terms of microanatomical compartments in slow exchange. In the context of this interpretation, our data suggest that diffusional anisotropy is greater outside than within the myelinated axons. Further, data following the onset of global ischemia suggest that the mechanism(s) by which ADC is reduced affect most or all microanatomical environments of nerve, at least insofar as they are represented over the TE domain investigated. Magn Reson Med 43:837-844, 2000. PMID- 10861879 TI - Utility of simultaneously acquired gradient-echo and spin-echo cerebral blood volume and morphology maps in brain tumor patients. AB - An interleaved gradient-echo (GE) / spin-echo (SE) EPI sequence was used to acquire images during the first pass of a susceptibility contrast agent, in patients with brain tumors. Maps of 1) GE (total) rCBV (relative cerebral blood volume), 2) SE (microvascular) rCBV, both corrected for T(1) leakage effects, and 3) (DeltaR(2)*/DeltaR(2)), a potential marker of averaged vessel diameter, were determined. Both GE rCBV and DeltaR(2)*/DeltaR(2) correlated strongly with tumor grade (P = 0.01, P = 0.01, n = 15), while SE rCBV did not (P = 0.24, n = 15). When the GE rCBV data were not corrected for leakage effects, the correlation with tumor grade was no longer significant (P = 0.09, n = 15). These findings suggest that MRI measurements of total blood volume fraction (corrected for agent extravasation) and DeltaR(2)*/DeltaR(2), as opposed to maps of microvascular volume, may prove to be the most appropriate markers for the evaluation of tumor angiogenesis (the induction of new blood vessels) and antiangiogenic therapies. Magn Reson Med 43:845-853, 2000. PMID- 10861880 TI - In vivo gallbladder bile diffusion coefficient measurement by diffusion-weighted echo planar imaging in hamster fed normal and lithogenic diets. AB - It is shown that in vivo measurement of bile water apparent diffusion coefficient (ADC) by diffusion-weighted echo-planar imaging (EPI) in hamster gallbladder is possible providing motion artifact-free ADC values. These ADC values are used to estimate bile viscosity variation induced by normal diets, cholesterol gallstone inducing diets, and an antilithiasic drug, and to determine if a link exists between bile viscosity and cholesterol gallstone formation. Measurements were performed at 4.7 T with respiratory triggering in five groups of hamsters fed a commercial (RC) or a semisynthetic (SSD) diet, a SSD containing 0.2% hyodeoxycholic acid (SSD+HDC) and two lithogenic diets (LD5, LD10). ADC decreased significantly in LD10 (2.15+/-0.07x 10(-3) mm(2)s(-1)) and SSD+HDC (2.03+/-0.04) compared to RC (2.40+/-0.05) but not in the most lithogenic LD5 diet (2.33+/ 0.06). No direct relationship was found between bile viscosity and gallstone incidence; however, viscosity seems to be related to lipid contents of diets. Magn Reson Med 43:854-859, 2000. PMID- 10861881 TI - Optimization and evaluation of the signal intensity change in multisection oxygen enhanced MR lung imaging. AB - The behavior of the signal intensity in MRI of human lungs was investigated during inhalation of pure oxygen. Nine volunteers were examined, five using a breath-hold and four using a non-breath-hold technique. Four coronal slices were acquired in each volunteer using an inversion recovery turbo spin-echo sequence. The inversion time of the sequence was optimized for maximum contrast. Breathing of pure oxygen and room air was alternated in the volunteers. Breath-hold and non breath-hold cases were compared. Breathing pure oxygen lead to a statistically significant signal intensity increase (up to 18%) compared to breathing room air. In addition, T(1) maps were acquired during breathing 100% oxygen and room air. Inhalation of pure oxygen reduced the mean T(1) time of the lungs from 1280 (+/ 85) msec to 1224 (+/-139) msec without breath-hold and from 1219 (+/-176) to 1074 (+/-92) msec with breath-hold. Therefore, an optimized sequence and measurement protocol provided significant signal intensity changes utilizing 100% oxygen. Magn Reson Med 43:860-866, 2000. PMID- 10861882 TI - Reconstructing MR images from undersampled data: data-weighting considerations. AB - Data which are sampled more densely than the Nyquist limit in k-space are weighted prior to reconstruction by the inverse of the local sampling density. This work considers the effects of weighting data that are sampled less densely than the Nyquist limit. It specifically analyzes azimuthally undersampled projection reconstruction, variable density spirals, and variable density phase encoding. Effects on resolution, aliasing, and SNR are given. Higher resolution is obtained by weighting undersampled data according to the inverse of sampling density, while better SNR and less aliasing artifact are obtained by weighting undersampled data uniformly. Magn Reson Med 43:867-875, 2000. PMID- 10861883 TI - MRI of focal cerebral ischemia using (17)O-labeled water. AB - This work presents a novel approach for quantifying low concentrations of H(2)(17)O in vivo and explores its utility for assessing cerebral ischemia. Oxygen-17 enriched water acts as a T(2) shortening contrast agent whose effect can be suppressed by decoupling at the (17)O frequency during TE interval in a spin-echo MR image. Serial T(2)-weighted echo planar images were acquired in phantoms and rat brain with decoupler power alternated every eight images. The resulting periodic signal change (proportional to H(2)(17)O concentration) was detected by cross-correlating the square-wave decoupler power timecourse with the signal intensity in each voxel. Natural abundance (0.037 atom%) images of H(2)(17)O in rat brain were generated. The transverse relaxivity of H(2)(17)O in brain was estimated, R(2) = 2.4+/-0.5 s(-1)(atom%)(-1). After bolus injection of 1 ml of 10 atom% H(2)(17)O, brain H(2)(17)O concentration was estimated at 0.06+/ 0.01 atom%. In the rat focal ischemia model, (17)O cross-correlation maps compared well with diffusion and Gd-DTPA perfusion images to indicate infarct location. Magn Reson Med 43:876-883, 2000. PMID- 10861884 TI - Visualization of pressure distribution within loaded joint cartilage by application of angle-sensitive NMR microscopy. AB - High-resolution MRI measurements on knee joints show a multilaminar appearance of the cartilage. This intracartilaginar structure, visualized as hypointense zones in T(2)-weighted MR images is based on the dipolar interaction of water molecules within regions of anisotropic arrangement of collagen network. Using the different angle dependence of the MR signal, zones of radially and tangentially oriented network structures can be distinguished. Information equivalent to that from polarization light microscopy can be derived noninvasively. This is demonstrated by polarization light microscopic reference investigations. It is shown that this multilaminar MRI appearance is sensitively influenced by mechanical stress. A model explaining the contrary behavior of loaded tangential and radial network structures is given. Based on this pressure dependence, a noninvasive determination of mechanical properties is possible. Using the variation of size and intensity of the hypointense zones under pressure, dynamic high resolution MRI yields noninvasive information about the intracartilaginar pressure distribution similar to photoelastic measurements. Magn Reson Med 43:884 891, 2000. PMID- 10861885 TI - Combined connectivity and a gray-level morphological filter in magnetic resonance coronary angiography. AB - A connectivity algorithm combined with a new gray-level morphological filter dramatically improves the segmentation of tortuous coronary arteries from 3D MRI. Small coronary arteries are segmented from the larger ventricles with a new filter. These blood vessels are segmented from the noise background with connectivity. Coronary angiograms were computed in nine datasets acquired on volunteers with 3D stack of spirals and contrast-enhanced navigator sequences by both a maximum intensity projection and surface rendering. Surface images provided depth information needed to distinguish branching arteries from crossing veins. Magn Reson Med 43:892-895, 2000. PMID- 10861886 TI - A protocol for assessing subtraction errors of arterial spin-tagging perfusion techniques in human brain. AB - A protocol for assessing signal contributions from static tissue (subtraction errors) in perfusion images acquired with arterial spin-labeling (ASL) techniques in human brain is proposed. The method exploits the reduction of blood T(1) caused by the clinically available paramagnetic contrast agent, gadopentetate dimeglumine (Gd-DTPA). The protocol is demonstrated clinically with multislice FAIR images acquired before, during, and after Gd-DTPA administration using a range of selective inversion widths. Perfusion images acquired postcontrast for selective inversion widths large enough (threshold) to avoid interaction with the imaging slice had signal intensities reduced to noise level, as opposed to subtraction errors manifested on images acquired using inversion widths below the threshold. The need for these experiments to be performed in vivo is further illustrated by comparison with phantom results. The protocol allows a one-time calibration of relevant ASL parameters (e.g., selective inversion widths) in vivo, which may otherwise cause subtraction errors. Magn Reson Med 43:896-900, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861887 TI - Temperature monitoring in fat with MRI. AB - The aim of the study was to test the hypothesis that fast spin echo T(1)-weighted images can be used to quantify the temperature in fat during thermal therapy in vivo. An MR compatible positioning device was used to manipulate focused ultrasound transducers in an MRI scanner. This system was used to sonicate fat tissue around the kidneys of 12 rabbits at various power levels for 10 to 20 sec. The scan parameters of T(1)-weighted fast spin echo (FSE) sequence were varied to optimize signal intensity characteristics while maintaining short scan times. An invasive optical probe was used to calibrate the temperature related signal intensity changes. For the T(1)-weighted FSE sequence, the signal intensity decreased with the temperature elevation at the rate of 0.97+/-0.02%/ degrees C. The single focused transducer produced a contrast-to-noise ratio more than 10 at power levels below the tissue damage threshold. The signal intensity was linearly dependent on the power, despite the measured temperatures being well above the coagulation threshold. This study demonstrates that T(1)-weighted FSE MRI sequences can be used to quantify the temperature elevation in fat in vivo during short focused ultrasound exposures. This can be very important for breast tumor surgery, fat ablation, and for treating deep seated tumors through superficial fat layers. Magn Reson Med 43:901-904, 2000. PMID- 10861888 TI - High-speed interlaced spin-echo magnetic resonance imaging. AB - A new method is introduced for increasing the efficiency in multislice single spin-echo MRI. The method interlaces the excitation and measurement of different slices, resulting in an effective use of the echo delay time between RF excitation and reception. Under certain conditions, the method allows for scan time reduction compared to standard single spin-echo MRI, in particular for long echo times. The technique is demonstrated in examples of brain scans, indicating that a substantial increase is scan speed can be achieved without loss in image signal-to-noise ratio or contrast. Potential applications include perfusion imaging using T(2)-contrast agents, as well as BOLD-based functional imaging. Magn Reson Med 43:905-908, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10861889 TI - Monitoring of high-intensity focused ultrasound-induced temperature changes in vitro using an interleaved spiral acquisition. AB - An interleaved, spoiled gradient-echo spiral acquisition technique was implemented to monitor high-intensity focused ultrasound heating of porcine kidney ex vivo by measuring temperature induced phase shifts in the detected MR signal. Echo time, flip angle, repetition time, number of interleaves, and readout time were varied to observes effects on temperature sensitivity and phase difference noise. The temperature response of the interleaved spiral acquisition was found to be comparable to a spoiled fast gradient-echo sequence of comparable in-plane spatial resolution. However, when imaging with an optimal echo time, spiral acquisition offers dramatically increased temporal resolution for comparable spatial resolution. Magn Reson Med 43:909-912, 2000. PMID- 10861890 TI - Multiple inversion recovery MR subtraction imaging of human ventilation from inhalation of room air and pure oxygen. AB - The feasibility of MR subtraction imaging of lung ventilation using air against oxygen using a multiple inversion recovery half-Fourier single-shot turbo spin echo (MIR-HASTE) sequence was investigated. Eight healthy, nonsmoking volunteers (3 males, 5 females; from 27 to 48 years of age) were studied on a 1.5 T MR unit. The ventilation image was obtained from the subtraction of the images acquired with the subject inhaling room air and 100% oxygen. By suppressing the signal from subcutaneous fat and thoracic muscle, MIR-HASTE improved the subtraction of signal arising from background tissues. Lung parenchyma, pulmonary veins, descending aorta, spleen, and kidney showed high signal difference, but pulmonary arteries exhibited minimal signal difference. Because of minimal signal change in the pulmonary arteries after inhalation of 100% oxygen, the average signal decreases in the left and right lungs including hilus and periphery amounted to only 19.4+/-4.5 and 20.2+/-3.4%, respectively, compared with regional averages of 23.6+/-5.4 and 24.1+/-3.1% for both lung peripheries alone. Magn Reson Med 43:913 916, 2000. PMID- 10861892 TI - Color schemes to represent the orientation of anisotropic tissues from diffusion tensor data: application to white matter fiber tract mapping in the human brain PMID- 10861891 TI - Probe with chest shielding for improved breast MRI. AB - The design and construction of an RF coil system for use in MR breast imaging is described. The two-ring, tuned Helmholtz coil, with its axis perpendicular to the chest, surrounds a single pendant breast and is coupled both internally and to the MRI transmitter/receiver by mutual induction. The addition of two symmetrical RF shields minimizes losses in the chest and significantly improves performance. Images obtained from eight healthy volunteers showed that the coil permitted imaging of breasts of diverse size with an in-plane resolution of 0.27 x 0.27 mm and a slice thickness of less than 2 mm at a field strength of 3 T as well as 1.5 T. The use of shields with surface coils in general is advocated as a method for improving signal-to-noise ratio. Magn Reson Med 43:917-920, 2000. PMID- 10861893 TI - Haseman and Elston revisited. AB - Haseman and Elston (H-E) [1972] proposed a method to detect quantitative trait loci by linkage to a marker. The squared sib-pair trait difference is regressed on the proportion of marker alleles the pair is estimated to share identical by descent: a significantly negative regression coefficient suggests linkage. It has been shown that a maximum likelihood method that directly models the sib-pair covariance has more power. This increase in power can also be obtained using the H-E regression procedure by changing the dependent variable from the squared difference to the mean-corrected product of the sibs' trait values. Multiple sibs in a sibship can be accommodated by allowing for the correlations between pairs of products in a generalized least squares procedure. Multiple trait loci, including epistatic interactions, involve only multiple linear regression. Multivariate traits can use the method of Amos et al. [1990] to find the linear function of the traits that maximizes the evidence for linkage, which now leads more simply to a test of significance. Multiple markers can be the basis of a multipoint analysis. Results of simulation studies for a continuous trait are presented that investigate Type I error and power. A similar general scheme can be used to study affected sib pairs, testing whether their identity by descent sharing probabilities are greater than would be expected in the absence of linkage, and to study other types of relative pairs. PMID- 10861894 TI - Circumventing multiple testing: a multilocus Monte Carlo approach to testing for association. AB - Advances in marker technology have made a dense marker map a reality. If each marker is considered separately, and separate tests for association with a disease gene are performed, then multiple testing becomes an issue. A common solution uses a Bonferroni correction to account for multiple tests performed. However, with dense marker maps, neighboring markers are tightly linked and may have associated alleles; thus tests at nearby marker loci may not be independent. When alleles at different marker loci are associated, the Bonferroni correction may lead to a conservative test, and hence a power loss. As an alternative, for tests of association that use family data, we propose a Monte Carlo procedure that provides a global assessment of significance. We examine the case of tightly linked markers with varying amounts of association between them. Using computer simulations, we study a family-based test for association (the transmission/disequilibrium test), and compare its power when either the Bonferroni or Monte Carlo procedure is used to determine significance. Our results show that when the alleles at different marker loci are not associated, using either procedure results in tests with similar power. However, when alleles at linked markers are associated, the test using the Monte Carlo procedure is more powerful than the test using the Bonferroni procedure. This proposed Monte Carlo procedure can be applied whenever it is suspected that markers examined have high amounts of association, or as a general approach to ensure appropriate significance levels and optimal power. PMID- 10861895 TI - Model-free sib-pair linkage analysis: combining full-sib and half-sib pairs. AB - When sampling full-sibs for linkage studies, half-sibs are often available. Not only are half-sibs convenient to sample, but they can sometimes offer greater power than full-sibs. We propose a method to combine the information from full sibs and half-sibs into a single test for linkage. This method is based on the Haseman and Elston [1972] method of regressing the squared trait-difference for a pair of sibs (either full- or half-sibs) on the estimated proportion of alleles shared identical by descent. To approximate the distribution of the test statistic, we propose a correction factor that considers the correlation among sibs, and demonstrate by simulations that this approximation works well in many situations, although there are some conditions for which the statistic can have an inflated Type-I error rate. The main appeal of our proposed method is the speed at which it can be computed, offering a rapid way to perform genome-wide linkage screens. PMID- 10861896 TI - Apo E genotype, diabetes, and peripheral arterial disease in older men: the Honolulu Asia-aging study. AB - The epsilon4 allele of the gene coding for apolipoprotein (apo) E is associated with an atherogenic lipid profile that has been linked to increased risk of coronary artery disease (CAD). Apo E genotype may also be associated with peripheral arterial disease (PAD). If present, this association may be modified by diabetes, which is also associated with dyslipidemia that predisposes to macrovascular disease. Observable associations between both ApoE genotype and diabetes with PAD may be confounded by smoking, a potent PAD risk factor that is unrelated to lipids. From 1991 to 1993, apo E genotypes (2/3, 3/3, 3/4), PAD (defined as ankle-brachial index [ABI] <0.9), diabetes (prevalent and newly diagnosed), and smoking history (ever/never) were determined for 3,161 Japanese American men aged 71-93. Data on hypertension and other potential confounders were also collected. Logistic regression was used to determine odds ratios (OR) between groups cross-categorized by apo E genotype and diabetes with prevalence of PAD, within strata of smoking. In each smoking stratum, non-diabetic apo epsilon3/3 carriers were considered the reference. Among ever-smokers, there was no association between apo E and PAD, regardless of diabetes status. Among never smokers there appeared to be both apo E- and diabetes effects on PAD prevalence. Compared to the non-diabetic epsilon3/3 group, the ORS of PAD were 2.3 (1.2-4.4) and 2.0 (1.1-3.4) for epsilon3/3 newly-diagnosed and epsilon3/3 prevalent diabetic subjects, respectively. Associations were stronger among diabetic individuals in the epsilon3/4 group: the ORS were 3.0 (1.1-8.8) and 4.1 (1.9-8.7) for epsilon3/4 newly-diagnosed and epsilon3/4prevalent diabetic subjects, respectively. Despite associations whose pattern and magnitude suggested interaction between apo E genotype and diabetes on PAD prevalence among never smokers, formal testing of this interaction did not reach statistical significance. Our finding of an apo E-PAD association among never-smokers may result from the effects of an apo epsilon4-related atherogenic lipid profile on peripheral arteries. Further studies are needed to clarify the potential mediating role of diabetes on the apo E-PAD association. PMID- 10861897 TI - Extensive association analysis between the CETP gene and coronary heart disease phenotypes reveals several putative functional polymorphisms and gene-environment interaction. AB - An extensive association analysis of a candidate gene for coronary heart disease, Cholesteryl Ester Transfer Protein (CETP) gene, was performed. Ten polymorphisms, out of which three were newly identified in regulatory regions, were investigated for association with myocardial infarction (MI) and 2 MI endophenotypes (CETP mass and HDL-cholesterol level) in 568 MI patients and 668 controls. The polymorphisms affecting codon 405 (Ile(405)Val) and the nucleotide 524 downstream from the stop codon (G(+524)T) were almost completely concordant and associated with plasma CETP mass (P < 0.001). The polymorphisms -629 (located in promoter), intron1 (Taq1B) and intron7 were almost completely concordant and associated with plasma CETP mass (P < 0.0001) and HDL-cholesterol levels (P < 0.0001). This latter association was not found in teetotalers and increased with the quantity of alcohol consumed. Heavy drinkers (>75g/day) homozygous for the (-628)A allele had a reduced risk of MI (OR = 0. 33, P < 0.02). Subjects both homozygous for (451)Arg and heterozygous for (373)Pro had decreased plasma HDL-cholesterol levels and this effect increased with alcohol consumption. The results illustrate the complexity of polymorphism-phenotype associations. They suggest that the CETP gene may carry several functional polymorphisms. Observed interactions between alcohol consumption and polymorphisms associated with HDL-cholesterol level constitute concrete examples of gene-environment interactions. Furthermore, the pattern of association between HDL-cholesterol levels and the polymorphisms at codons 373 and 451 illustrated how two polymorphisms may be confounders (in the usual epidemiological sense) one for the other: their marginal effects are neutralized because of linkage disequilibrium and thus are not detectable by standard univariate association analysis. PMID- 10861898 TI - Evidence for major genes influencing pulmonary function in the NHLBI family heart study. AB - Segregation analysis was performed on the pulmonary measures forced expiratory volume in one second (FEV1), forced vital capacity (FVC), and the ratio of FEV1/FVC in 455 randomly ascertained families from the NHLBI Family Heart Study (FHS). Gender specific standardized residuals were used as the phenotypic variable in both familial correlation and segregation analyses. These residuals represented adjustments for the effects of age, age(2), age(3), Body Mass Index (BMI, kg/m(2)), height, the ratio of waist to hip measurements (WHR), the presence of coronary heart disease, smoking history, and pack years for current smokers. Sibling correlations were not different from parent-offspring correlations for all three traits, and heritability estimates for FEV1, FVC, and the FEV1/FVC ratio were 0. 515, 0.540, and 0.449, respectively. Segregation analysis of FEV1, a trait that measures airflow, indicated that a dominant major gene best fits the data, although a residual familial correlation supports the presence of an additional polygenic or common environmental component. For FVC, a trait that measures lung volume, alternative models could not be statistically differentiated, but the transmission probabilities do not support a Mendelian major gene. The best model for FEV1/FVC ratio is a non-Mendelian codominant model, perhaps due to the mixing of the individual underlying distributions influencing airflow and lung volume. These results support the hypothesis that complex relationships exist for lung function traits and that multiple genes and environmental factors influence lung function. PMID- 10861899 TI - Tailoring wine yeast for the new millennium: novel approaches to the ancient art of winemaking. AB - Yeasts are predominant in the ancient and complex process of winemaking. In spontaneous fermentations, there is a progressive growth pattern of indigenous yeasts, with the final stages invariably being dominated by the alcohol-tolerant strains of Saccharomyces cerevisiae. This species is universally known as the 'wine yeast' and is widely preferred for initiating wine fermentations. The primary role of wine yeast is to catalyze the rapid, complete and efficient conversion of grape sugars to ethanol, carbon dioxide and other minor, but important, metabolites without the development of off-flavours. However, due to the demanding nature of modern winemaking practices and sophisticated wine markets, there is an ever-growing quest for specialized wine yeast strains possessing a wide range of optimized, improved or novel oenological properties. This review highlights the wealth of untapped indigenous yeasts with oenological potential, the complexity of wine yeasts' genetic features and the genetic techniques often used in strain development. The current status of genetically improved wine yeasts and potential targets for further strain development are outlined. In light of the limited knowledge of industrial wine yeasts' complex genomes and the daunting challenges to comply with strict statutory regulations and consumer demands regarding the future use of genetically modified strains, this review cautions against unrealistic expectations over the short term. However, the staggering potential advantages of improved wine yeasts to both the winemaker and consumer in the third millennium are pointed out. PMID- 10861900 TI - Mitotic recombination in yeast: elements controlling its incidence. AB - Mitotic recombination is an important mechanism of DNA repair in eukaryotic cells. Given the redundancy of the eukaryotic genomes and the presence of repeated DNA sequences, recombination may also be an important source of genomic instability. Here we review the data, mainly from the budding yeast S. cerevisiae, that may help to understand the spontaneous origin of mitotic recombination and the different elements that may control its occurrence. We cover those observations suggesting a putative role of replication defects and DNA damage, including double-strand breaks, as sources of mitotic homologous recombination. An important part of the review is devoted to the experimental evidence suggesting that transcription and chromatin structure are important factors modulating the incidence of mitotic recombination. This is of great relevance in order to identify the causes and risk factors of genomic instability in eukaryotes. PMID- 10861901 TI - A history of research on yeasts 2: Louis Pasteur and his contemporaries, 1850 1880. PMID- 10861902 TI - Budding yeast as a model organism for population genetics. AB - Population genetics is a highly theoretical field in which many models and theories of broad significance have received little experimental testing. Microbes are well-suited for empirical population genetics since populations of almost any size may be studied genetically, and because many have easily controlled life cycles. Saccharomyces cerevisiae is almost ideal for such studies as the growing body of knowledge and techniques that have made it the best characterized eukaryote genome also allow the experimental manipulation and analysis of its population genetics. In experiments to date, the evolution of laboratory yeast populations has been observed for up to 1000 generations. In several cases, adaptation has occurred by gene duplications. The interaction between mutation, selection and genetic drift at varying population sizes is a major area of theoretical study in which yeast experiments can provide particularly valuable data. Conflicts between gene-level and among-cell selection, and co-evolution between genes within a genome, are additional topics in which a population genetics perspective may be particularly helpful. The growing field of genomics is increasingly complementary with that of population genetics. The characterization of the yeast genome presents unprecedented opportunities for the detailed study of evolutionary and population genetics. Conversely, the redundancy of the yeast genome means that, for many open reading frames, deletion has only a quantitative effect that is most readily observed in competitions with a wild-type strain. PMID- 10861903 TI - The yeast Ty virus-like particles. AB - Virus-like particle (VLP) assembly is a crucial step of the life cycle of retrotransposons. The S. cerevisiae Ty elements represent an interesting model for the analysis of these particles and thus have been studied extensively. Our current knowledge of the organisation and assembly of Ty1 and Ty3 VLPs is reviewed here. This includes the mechanism of assembly, the role of the Tya core protein during VLP formation and the RNA packaging process. The physical properties of Ty1 VLPs are also described and the latest three-dimensional Ty1 VLP reconstructions are shown. In addition, the relevance of these studies is discussed in the context of retro-element biology. PMID- 10861904 TI - The importance of ATP as a regulator of glycolytic flux in Saccharomyces cerevisiae. AB - The control of glycolytic flux in the yeast Saccharomyces cerevisiae was studied by using permeabilized cells. Cells were harvested from chemostat cultures and, after removal of the cell wall, nystatin was used to permeabilize the spheroplasts. By this method it is possible to study the performance and regulation of a complete and functional metabolic pathway and not only a single enzymatic step. The results showed that ATP has a strong negative effect on glycolytic activity affecting several of the glycolytic enzymes. However, the main targets for ATP inhibition was phosphofructokinase and pyruvate kinase. Phospofructokinase was inhibited by ATP concentrations starting at about 1-2 mM, while pyruvate kinase required ATP levels above 2.5 mM before any inhibition was visible. These ATP concentrations were in the same range as measured for nitrogen and glucose-limited cells cultivated in chemostat cultures. Other potential candidates as enzymes susceptible to ATP inhibition included hexokinase and enolase. The ATP:ADP ratio, as well as trehalose-6-phosphate levels, did not seem to influence the glycolytic activity. PMID- 10861905 TI - Mutational analysis of the karmellae-inducing signal in Hmg1p, a yeast HMG-CoA reductase isozyme. AB - In response to elevated levels of HMG-CoA reductase, an integral endoplasmic reticulum (ER) membrane protein, cells assemble novel ER arrays. These membranes provide useful models for exploration of ER structure and function, as well as general features of membrane biogenesis and turnover. Yeast express two functional HMG-CoA reductase isozymes, Hmg1p and Hmg2p, each of which induces morphologically different ER arrays. Hmg1p induces stacks of paired nuclear associated membranes called karmellae. In contrast, Hmg2p induces peripheral ER membrane arrays and short nuclear-associated membrane stacks. In spite of their ability to induce different cellular responses, both Hmg1p and Hmg2p have similar structures, including a polytopic membrane domain containing eight predicted transmembrane helices. By examining a series of recombinant HMG-CoA reductase proteins, our laboratory previously demonstrated that the last ER-lumenal loop (Loop G) of the Hmg1p membrane domain contains a signal needed for proper karmellae assembly. Our goal was to examine the primary sequence requirements within Loop G that were critical for proper function of this signal. To this end, we randomly mutagenized the Loop G sequence, expressed the mutagenized Hmg1p in yeast, and screened for inability to generate karmellae at wild-type levels. Out of approximately 4000 strains with Loop G mutations, we isolated 57 that were unable to induce wild-type levels of karmellae assembly. Twenty-nine of these mutants contained one or more point mutations in the Loop G sequence, including nine single point mutants, four of which had severe defects in karmellae assembly. Comparison of these mutations to single point mutations that did not affect karmellae assembly did not reveal obvious patterns of sequence requirements. For example, both conservative and non-conservative changes were present in both groups and changes that altered the total charge of the Loop G region were observed in both groups. Our hypothesis is that Loop G serves as a karmellae-inducing signal by mediating protein-protein or protein-lipid interactions and that amino acids revealed by this analysis may be important for maintaining the proper secondary structure needed for these interactions. PMID- 10861906 TI - A transcriptional autoregulatory loop for KIN28-CCL1 and SRB10-SRB11, each encoding RNA polymerase II CTD kinase-cyclin pair, stimulates the meiotic development of S. cerevisiae. AB - Alkalization of the medium is associated with and required for the cellular development to meiosis and sporulation in the yeast Saccharomyces cerevisiae. To elucidate the molecular mechanisms for the significance of external alkalization, we isolated mutants defective in division arrest at G1 phase under an alkaline condition. The mutants obtained had recessive alleles of SRB10 encoding the cyclin (SRB11)-dependent protein kinase that phosphorylates the CTD domain of the largest subunit of RNA polymerase II and negatively regulates the transcriptional initiation of certain genes. A delta srb11 deletion mutant showed the same cell cycle defect. When shifted to alkali, wild-type cells decreased transcript levels of G1-cyclin genes (CLN1 to CLN3) and KIN28-CCL1 (encoding another CTD kinase cyclin pair which, in contrast, stimulates the promoter clearance and transcriptional elongation in most genes), resulting in the accumulation of G1 cells and the hypophosphorylated form of RNA polymerase II and in an increase in cell size. However, under the same conditions, a delta srb10 mutant was defective in these events, except the downregulation of CLN1 and CLN2. The delta srb10 mutation also influenced on the transcript levels of meiosis-inducing genes called IME1 and IME2: the mutation elevated the transcript level of IME1 but reduced that of IME2, resulting in partial defects in premeiotic DNA synthesis and meiosis. Overexpression of KIN28 and CCL1 in wild-type cells impaired the alkali-induced G1 arrest and the rate of meiosis and elevated the transcript levels of SRB11 and IME1. These results indicate that a transcriptional autoregulatory loop for KIN28-CCL1 and SRB10-SRB11 is important for G1 arrest and meiosis. We also found that environmental conditions for meiosis finely regulate the transcript levels of KIN28 and CCL1, such that nitrogen starvation first elevates them but subsequent alkalization of medium decreases them. PMID- 10861907 TI - The ALS6 and ALS7 genes of Candida albicans. AB - ALS genes of Candida albicans encode a family of cell-surface glycoproteins that are composed of an N-terminal domain, a central domain of a tandemly repeated motif, and a relatively variable C-terminal domain. Although several ALS genes have been characterized, more ALS-like sequences are present in the C. albicans genome. Two short DNA sequences with similarity to the 5' domains of known ALS genes were detected among data from the C. albicans genome sequencing project. Probes developed from unique regions of these sequences were used to screen a genomic library from which two full-length genes, designated ALS6 and ALS7, were cloned. ALS6 and ALS7 encode features similar to other genes in the ALS family and map to chromosome 3, a chromosome previously not known to encode ALS sequences. ALS6 and ALS7 are present in all C. albicans strains examined. Additional analysis suggested that some C. albicans strains have another ALS gene with a 5' domain similar to that of ALS6. Characterization of ALS7 revealed a novel tandemly repeated sequence within the C-terminal domain. Unlike other ALS family tandem repeats, the newly characterized ALS7 repeat does not appear to define additional genes in the ALS family. However, our data and information from the C. albicans genome sequencing project suggest that there are additional ALS genes remaining to be characterized. PMID- 10861908 TI - Rapid and reliable protein extraction from yeast. AB - The methods currently used for protein extraction from yeast are either laborious or insufficiently reliable. Here I report a method for protein extraction for electrophoretic analysis that is both easy and reliable. In this method, yeast cells are subjected to mild alkali treatment and then boiled in a standard electrophoresis loading buffer. The method was tested for different strains of Saccharomyces cerevisiae and for yeast Hansenula polymorpha DL-1. It yields virtually complete extraction independently of the strain, growth conditions and protein molecular weight and allows working with small amounts of yeast cells grown on agar plates. PMID- 10861909 TI - A family of multifunctional thiamine-repressible expression vectors for fission yeast. AB - A series of thiamine-repressible shuttle vectors has been constructed to allow a more efficient DNA manipulation in Schizosaccharomyces pombe. These high-copy number vectors with regulatable expression (pJR) are based on the backbone of the pREP-3X, pREP-41X and pREP-81X plasmids. The pJR vectors are all uniform in structure, containing: (a) sequences for replication (ori) and selection (AmpR) in Escherichia coli; (b) the f1 ori sequence of the phage f1 for packaging of ssDNA, making them suitable for site-directed mutagenesis; and (c) the ars1 sequence for replication in S. pombe. The pJR vectors differ among them in: (a) the selectable marker (Saccharomyces cerevisiae LEU 2 gene, which complements S. pombe leu1- gene and S. pombe ura4+ and his3+ genes); (b) the thiamine repressible nmt1 promoter (3X, 41X and 81X with extremely high, moderate or low transcription efficiency, respectively); and (c) the multiple cloning site (two multiple cloning sites, with 12 restriction sites each). The expression level of the pJR vectors has been analysed using the beta-galactosidase gene as reporter. Three levels of expression for each nmt1 promoter version, with any selectable marker and for either repressed or induced conditions, have been found. The expression is dependent on the distance to the initiation codon, varying from 0.001 to 15 times the activity characterized for the pREP plasmids. Also, the gene expression has been found to be extremely sensitive to the nucleotide sequence prior to the initiation codon, being up to 50-fold higher with an A/T sequence than with a G/C sequence. Finally, the beta-galactosidase mRNA levels were found to be similar in each nmt1 series, suggesting a translational effect on gene expression. As a result, any of these 18 new vectors allow performing gene expression in fission yeast, as well as a more versatile cloning, sequencing and mutagenesis, directly in the plasmid without the need for subcloning into intermediary vectors. PMID- 10861910 TI - Current awareness on yeast. PMID- 10861911 TI - 'Pressing issues in the dementias and dementia servicesrising single quote, left (low) symposium 28-29 january 1999 royal college of physicians, london: foreword PMID- 10861912 TI - Predictors of positive and negative appraisal among Cuban American caregivers of Alzheimer's disease patients. AB - OBJECTIVE: This study investigated predictors of positive (satisfaction) and negative (burden) appraisal among Cuban American (CA) caregivers of Alzheimer's disease (AD) patients. DESIGN: Cross-sectional study of AD patients and their family caregivers. SETTING: A university-affiliated outpatient memory disorders clinic. SUBJECTS: A convenience sample of 40 CA family caregivers of patients diagnosed with probable or possible AD according to NINCDS-ADRDA diagnostic criteria. MEASURES: AD patients: Mini-Mental State Examination (MMSE), Blessed Dementia Scale (BDS) and the Behavioral Pathology in Alzheimer's Disease Rating Scale (BEHAVE-AD). Caregivers: Caregiving Burden Scale (CBS), Caregiving Satisfaction Scale (CSS), Perceived Emotional Support scale (PES) and the Short Form-36 Health Survey-General Health Index (GH). RESULTS: Appraised burden was predicted by increased patient behavioral pathology, female caregiver gender and lower levels of perceived emotional support. The association between older caregiver age and increased burden approached significance. Older caregiver age and higher levels of perceived social support were shown to predict appraised satisfaction. Post-hoc analyses also indicated that length of residence in the United States, a measure of acculturation, was not associated with positive or negative appraisal. CONCLUSION: Appraised burden and satisfaction represent important outcomes of dementia care that show relations with distinct factors among CA caregivers. It is clear that further research is warranted in order to ascertain the relationship of ethnicity or culture to the process and psychological consequences of dementia caregiving. Continued investigations into predictors of caregiving satisfaction are also recommended. PMID- 10861913 TI - The implication of selection bias in clinical studies of late life depression: an empirical approach. AB - OBJECTIVES: It is supposed that selection bias precludes the extrapolation of results of studies carried out in a clinical setting to the general population. There is little empirical evidence demonstrating the degree to which those depressed in the community are different from those treated in clinical settings. This study compared elderly patients with major depression admitted to a psychiatric hospital with those living in the community. METHODS: All elderly (55 years and older) patients admitted between 1990 and 1992 to a psychiatric hospital with DSM major depression as the primary diagnosis (n=104), were compared with all elderly patients with the same diagnosis (n=59) who were participating in a large community study (Longitudinal Aging Study, Amsterdam). Data were gathered from the clinical sample using chart-reviews while the community-based sample was interviewed. The two groups were compared with respect to differences in demographic variables, presenting symptoms, risk factors and treatment. RESULTS: The following characteristics were significantly more prevalent in the clinical sample: late onset of the depression, threat of suicide, conflicts with significant others and use of antidepressant medication. Chronic physical illness was the only characteristic that was more prevalent in the community sample. CONCLUSION: The results confirm that elderly patients treated in clinical psychiatry represent a group with more threatening and more disruptive depressive illness. Major depression in the community was more often associated with chronic physical illness, which may hamper the recognition and treatment of depression. As the two samples were similar in all other respects, selection bias, hampering comparison of results of studies carried out across treatment settings, appears to have a very limited effect. PMID- 10861914 TI - Some predictors of mortality in acutely medically ill elderly inpatients. AB - BACKGROUND: The prevalence of depression and suicidal ideation in acutely medically ill elderly inpatients is high. Depression and suicidal ideation are associated with increased mortality. METHOD: The following were examined among acutely medically ill elderly inpatients: the association between mortality at 6 8 month follow-up period and Brief Assessment Scale (BAS-DEP) depression caseness and scores, using BAS-DEP items of 'a wish to die', 'pessimism' and 'life not worth living', functional disability measured by the London Handicap Scale (LHS) and the Barthel Index (BI), suicidal ideation measured by the Beck Suicidal Ideation Scale (BSSI), severity of physical illness, previous deliberate self harm and demographic variables. RESULTS: On univariate analysis, significant associations between mortality and being married, previous deliberate self-harm, higher scores on the BAS-DEP item of pessimism, lower scores on the LHS and the BI and higher scores on the BSSI were observed. On multivariate analysis only LHS scores and BSSI scores independently predicted mortality. CONCLUSIONS: It is hypothesized that suicidal ideation and functional disability may have a causal effect on mortality. This hypothesis could be tested by early identification of suicidal ideation and/or functional disability and subsequent interventions specifically designed to improve these two facets using a randomized controlled design. PMID- 10861915 TI - Therapy with the elderly: introducing psychodynamic psychotherapy to the multi disciplinary team. AB - Psychotherapy with the elderly has tended to be restricted to family or behavioural paradigms with some contribution from cognitive approaches, but at least until recently, very little input from psychodynamic therapy. The reasons for this are discussed and include both patient and therapist factors. Freud himself was pessimistic regarding the prospects of working with the elderly, a viewpoint he changed as he aged. As the discipline of old age psychiatry also matures there may be increasing scope for collaboration between the mental health of the elderly teams and psychodynamically orientated therapists. We describe here one model of such collaboration, a weekly case discussion seminar, conducted in an acute admission old age psychiatry in-patient unit. The role of the seminar in discussing difficult cases will be described and a case will be discussed to illustrate the work. Finally, recommendations for broadening the scope of the work are advanced. PMID- 10861916 TI - A long-term, multicenter, open-label study of risperidone in elderly patients with psychosis. On behalf of the Risperidone Working Group. AB - RATIONALE: Studies have shown that risperidone is safe and efficacious in young and middle-aged adults with chronic schizophrenia, but considerably fewer data are available on the treatment of elderly patients with schizophrenia or other psychotic disorders, particularly long-term outcomes. OBJECTIVE: A 12-month, open label study was conducted to assess the effects of risperidone in elderly, chronically ill, psychotic patients. METHODS: This study enrolled 180 elderly, chronically ill, psychotic patients (median age, 72 years [range 54-89]), 97 of whom completed the 12-month study. At endpoint, the mean dose of risperidone was 3.7 mg/day. RESULTS: Clinical improvement (> or =20% reduction in Positive and Negative Syndrome Score [PANSS] total score) was achieved by 54% of patients at endpoint. There were significant reductions in PANSS total, subscale (positive, negative, and general psychopathology), and cognition cluster scores at endpoint (p<0.001). Clinical Global Impressions severity of illness scores showed continued improvement through month 12 (p<0.001). In contrast, PANSS data from a historical comparable control group of patients receiving conventional antipsychotic agents showed no symptom improvement over a 12-month treatment period. The severity of preexisting extrapyramidal symptoms (EPS) in patients treated with risperidone decreased significantly from baseline to endpoint (p<0.001), and the use of antiparkinsonian medication decreased from 41.1% of patients before the trial to 25.6% during the trial. There were no spontaneous reports of tardive dyskinesia (TD) and the incidence of assessed TD was 4.3% in contrast to the expected 26% reported in middle-aged and elderly patients receiving conventional antipsychotic agents for 1 year. CONCLUSIONS: Long-term treatment with risperidone was associated with continued symptom improvement, a decrease in the severity of preexising EPS, and a low incidence of TD in elderly psychotic patients. PMID- 10861917 TI - Social factors and the outcome of dementia. AB - Previous studies have linked reduced survival in dementia with male sex, older age, longer duration of illness and increased severity of cognitive impairment. However, little is known about the potential influence of social factors (such as life events and social support) on the outcome of dementia. Sixty recently admitted patients with dementia (27 in-patients and 33 day patients) were given detailed psychiatric and social assessments. These included information on life events and social supports. At follow-up, 3 years later, data were collected on outcome. The hypotheses were that adverse life events and lack of social support would be associated with reduced survival. Nearly half the patients (48%) died during the follow-up period. The experience of life events before the first assessment was not associated with outcome. However, receiving meals on wheels (p=0.01) was associated with reduced survival, while attending a day centre (p=0.06) and having support from relatives (p=0.06) were associated with increased survival. Higher dependency (p=0.004) and poorer physical health (p=0.07) were associated with reduced survival. These results suggest that factors related to social support are associated with the outcome of dementia. In particular, receiving meals on wheels or home help may be a marker of a lack of social support, which influences outcome. Further studies are required to examine these associations in more detail. PMID- 10861918 TI - Community screening interview for dementia (CSI 'D'); performance in five disparate study sites. AB - The Community Screening Interview for Dementia (CSI 'D') was developed as a screening instrument for dementia for use in cross-cultural studies. It consists of two components, a cognitive test for non-literate and literate populations and an informant interview regarding performance in everyday living. The development of the CSI 'D', involving harmonization, translation, back translation and pilot testing, for use in five sites is described. The results demonstrate the adaptability and utility of the CSI 'D' in populations from very different socioeconomic backgrounds. The inclusion of informant data adds significantly to the performance of the CSI 'D' as a dementia screen. The combination of informant and cognitive scores in a discriminant score produces better sensitivity and specificity for dementia than cognitive scores alone. The informant score has a significant independent effect in predicting dementia. PMID- 10861919 TI - The effects of group work therapy in patients with Alzheimer's disease. PMID- 10861920 TI - Acute confusional states during treatment with risperidone. PMID- 10861921 TI - Ethics and the anti-dementia drugs. PMID- 10861922 TI - Diagnostic disclosure in dementia: an opportunity for intervention? AB - OBJECTIVE: To find out from people with dementia what they were worried about in relation to their diagnosis, and how they changed their behaviour in relation to these worries.Design. Consecutive case series of people with early dementia presenting for neuropsychological assessment. METHODS: People's ability to engage in talking about dementia was assessed by asking three standardised questions. Two measures of cognitive function, the MMSE and MEAMS were given to all. Those who could engage were asked two open-ended questions in relation to what they worried about and how learning they had dementia had affected them. RESULTS: The commonest worries related to fear of others finding out, fears of social embarrassment, long term dependency needs and not being listened to. Commonest effects were social withdrawal and hypervigilance for evidence of cognitive failures. CONCLUSIONS: People with dementia who know their diagnosis have worries which effect their behaviour in a way likely to result in low self-esteem, self stigmatisation and impaired quality of life. PMID- 10861923 TI - Clock-drawing: is it the ideal cognitive screening test? AB - OBJECTIVE: The clock-drawing test has achieved widespread clinical use in recent years as a cognitive screening instrument and a significant amount of literature relates to its psychometric properties and clinical utility. This review aims to synthesize the available evidence and assess the value of this screening test according to well-defined criteria. DESIGN: A Medline and Psycho-info literature search of all languages was done from 1983 to 1998 including manual cross referencing of bibliographies. A brief summary of all original scoring systems is provided as well as a review of replication studies. Psychometric data including correlations with other cognitive tests were recorded. Qualitative aspects of the test are also described. RESULTS: Among published studies, the mean sensitivity (85%) and specificity (85%) of the clock-drawing test are impressive. Correlations with the Mini-Mental State Examination and other cognitive tests was high, generally greater than r = 0.5. High levels of inter-rater and test-re-test reliability and positive predictive value are recorded and despite significant variability in the scoring systems, all report similar psychometric properties. The clock test also shows a sensitivity to cognitive change with good predictive validity. CONCLUSIONS: The clock-drawing test meets defined criteria for a cognitive screening instrument. It taps into a wide range of cognitive abilities including executive functions, is quick and easy to administer and score with excellent acceptability by subjects. Together with informant reports, the clock drawing test is complementary to the widely used and validated Mini-Mental State Examination and should provide a significant advance in the early detection of dementia and in monitoring cognitive change. A simple scoring system with emphasis on the qualitative aspects of clock-drawing should maximize its utility. PMID- 10861924 TI - Treatment for Alzheimer's disease. PMID- 10861925 TI - Current awareness. Bibliography. PMID- 10861926 TI - Protein threading using PROSPECT: design and evaluation. AB - The computer system PROSPECT for the protein fold recognition using the threading method is described and evaluated in this article. For a given target protein sequence and a template structure, PROSPECT guarantees to find a globally optimal threading alignment between the two. The scoring function for a threading alignment employed in PROSPECT consists of four additive terms: i) a mutation term, ii) a singleton fitness term, iii) a pairwise-contact potential term, and iv) alignment gap penalties. The current version of PROSPECT considers pair contacts only between core (alpha-helix or beta-strand) residues and alignment gaps only in loop regions. PROSPECT finds a globally optimal threading efficiently when pairwise contacts are considered only between residues that are spatially close (7 A or less between the C(beta) atoms in the current implementation). On a test set consisting of 137 pairs of target-template proteins, each pair being from the same superfamily and having sequence identity /=40 or van der Waals overlaps >/=0.4 A) greatly improves the clustering of rotamer populations. Asn, Gln, or His side chains additionally benefit from flipping of their planar terminal groups when required by atomic overlaps or H-bonding. Sensitivity to skew and to the boundaries of chi angle bins is avoided by using modes rather than traditional mean values. Rotamer definitions are listed both as the modal values and in a preferred version that maximizes common atoms between related rotamers. The resulting library shows significant differences from previous ones, differences validated by considering the likelihood of systematic misfitting of models to electron density maps and by plotting changes in rotamer frequency with B-factor. Few rotamers now show atomic overlaps in ideal geometry; those overlaps are relatively small and can be understood in terms of bond angle distortions compensated by favorable interactions. The new library covers 94.5% of examples in the highest quality protein data with 153 rotamers and can make a significant contribution to improving the accuracy of new structures. Proteins 2000;40:389 408. PMID- 10861931 TI - A comprehensive analysis of the Greek key motifs in protein beta-barrels and beta sandwiches. AB - The Greek key motifs are the topological signature of many beta-barrels and a majority of beta-sandwich structures. An updated survey of these structures integrates many early observations and newly emerging patterns and provides a better understanding of the unique role of Greek keys in protein structures. A stereotypical Greek key beta-barrel accommodates five or six strands and can have 12 possible topologies. All except one six-stranded topologies have been observed, and only one five-stranded topologies have been seen in actual structures. Of the representative beta-barrel structures analyzed here, half have left-handed Greek keys. This result challenges the empirical claim of the handedness regularity of Greek keys in beta-barrels. One of the five-stranded topologies that has not been observed in beta-barrels comprises two overlapping Greek keys. The two three-dimensional forms of this topology constitute a structural unit that is present in a vast majority of known beta-sandwich structures. Using this unit as the root, we have built a new taxonomy tree for the beta-sandwich folds and deduced a set of rules that appear to constrain how other beta-strands adjoin the unit to form a larger double-layered structure. These rules, though derived from a larger data set, are essentially the same as those drawn from earlier studies, suggesting that they may reflect the true topological constraints in the design of beta-sandwich structures. Finally, a novel variant of the Greek key motif (defined here as the twisted Greek key) has emerged which introduces loop crossings into the folded structures. Proteins 2000;40:409-419. PMID- 10861932 TI - Cell surface receptors and their ligands: in vitro analysis of CD6-CD166 interactions. AB - CD6 is a cell surface receptor belonging to the scavenger receptor cysteine-rich (SRCR) protein superfamily (SRCRSF). It specifically binds activated leukocyte cell adhesion molecule (ALCAM, CD166), a member of the immunoglobulin (Ig) superfamily (IgSF). CD166 was among the first molecules identified as a ligand for an SRCRSF receptor, and the CD6-CD166 interaction was the first interaction characterized involving SRCRSF and IgSF proteins. We focus here on what has been learned about the specifics of the CD6-CD166 interaction from in vitro analysis. The studies are thought to provide an instructive example for the analysis of interactions between single-path transmembrane cell surface proteins. Using soluble recombinant forms, the extracellular binding domains of receptor and ligand have been identified and characterized in a variety of assay systems. Both CD6 and CD166 have been subjected to intense mutagenesis and monoclonal antibody (mAb) binding studies and residues critical for their interaction have been identified. The availability of structural prototypes of both superfamilies has made it possible to map the binding site in CD166 and, more recently, in CD6 and compare these regions to epitopes of mAbs that block, or do not block, the interaction. In addition, the molecular basis of observed cross-species receptor ligand interactions could be rationalized. These studies illustrate the value of structural templates for the interpretation of sequence and mutagenesis analyses. Proteins 2000;40:420-428. PMID- 10861933 TI - Side chains in transmembrane helices are shorter at helix-helix interfaces. AB - Transmembrane helices from crystallographically determined structures were analyzed to determine the distribution of side chains inside and outside helix helix interfaces. Two structural characteristics were explored: (1) the number of atoms outside the interfaces that belong to the side chains with the C(alpha) atoms inside the interfaces, as well as the opposite, inside/outside number (conformation-dependent values) and (2) the side-chain length (depends only on the residue type and does not depend on the side-chain conformation). The results showed that the interface side chains tend to be bent away from the interacting helix. The most important finding, however, is that the side chains in the interface areas, on average, are shorter than in the noninterface areas. Proteins 2000;40:429-435. PMID- 10861934 TI - A new fold in the scorpion toxin family, associated with an activity on a ryanodine-sensitive calcium channel. AB - We determined the structure in solution by (1)H two-dimensional NMR of Maurocalcine from the venom of Scorpio maurus. This toxin has been demonstrated to be a potent effector of ryanodyne-sensitive calcium channel from skeletal muscles. This is the first description of a scorpion toxin which folds following the Inhibitor Cystine Knot fold (ICK) already described for numerous toxic and inhibitory peptides, as well as for various protease inhibitors. Its three dimensional structure consists of a compact disulfide-bonded core from which emerge loops and the N-terminus. A double-stranded antiparallel beta-sheet comprises residues 20-23 and 30-33. A third extended strand (residues 9-11) is perpendicular to the beta-sheet. Maurocalcine structure mimics the activating segment of the dihydropyridine receptor II-III loop and is therefore potentially useful for dihydropyridine receptor/ryanodine receptor interaction studies. Proteins 2000;40:436-442. PMID- 10861935 TI - Side-chain conformational entropy in protein unfolded states. AB - The largest force disfavoring the folding of a protein is the loss of conformational entropy. A large contribution to this entropy loss is due to the side-chains, which are restricted, although not immobilized, in the folded protein. In order to accurately estimate the loss of side-chain conformational entropy that occurs upon folding it is necessary to have accurate estimates of the amount of entropy possessed by side-chains in the ensemble of unfolded states. A new scale of side-chain conformational entropies is presented here. This scale was derived from Monte Carlo computer simulations of small peptide models. It is demonstrated that the entropies are independent of host peptide length. This new scale has the advantage over previous scales of being more precise with low standard errors. Better estimates are obtained for long (e.g., Arg and Lys) and rare (e.g., Trp and Met) side-chains. Excellent agreement with previous side-chain entropy scales is achieved, indicating that further advancements in accuracy are likely to be small at best. Strikingly, longer side chains are found to possess a smaller fraction of the theoretical maximum entropy available than short side-chains. This indicates that rotations about torsions after chi(2) are significantly affected by side-chain interactions with the polypeptide backbone. This finding invalidates previous assumptions about side chain-backbone interactions. Proteins 2000;40:443-450. PMID- 10861936 TI - Protein fold recognition by total alignment probability. AB - We present a protein fold-recognition method that uses a comprehensive statistical interpretation of structural Hidden Markov Models (HMMs). The structure/fold recognition is done by summing the probabilities of all sequence to-structure alignments. The optimal alignment can be defined as the most probable, but suboptimal alignments may have comparable probabilities. These suboptimal alignments can be interpreted as optimal alignments to the "other" structures from the ensemble or optimal alignments under minor fluctuations in the scoring function. Summing probabilities for all alignments gives a complete estimate of sequence-model compatibility. In the case of HMMs that produce a sequence, this reflects the fact that due to our indifference to exactly how the HMM produced the sequence, we should sum over all possibilities. We have built a set of structural HMMs for 188 protein structures and have compared two methods for identifying the structure compatible with a sequence: by the optimal alignment probability and by the total probability. Fold recognition by total probability was 40% more accurate than fold recognition by the optimal alignment probability. Proteins 2000;40:451-462. PMID- 10861937 TI - Turning an opinion inside-out: Rees and Eisenberg's commentary (Proteins 2000;38:121-122) on "Are membrane proteins 'inside-out' proteins?" (Proteins 1999;36:135-143). PMID- 10861938 TI - The duplication of an eight-residue helical stretch in Staphylococcal nuclease is not helical: a model for evolutionary change. AB - A common method of evolutionary change is gene duplication, followed by other events that lead to new function, decoration of folds, oligomerization, or other changes. As part of a study on the potential for evolutionary change created by duplicated sequences, we have carried out a crystallographic study on a mutant of Staphylococcal nuclease in which residues 55-62 have been duplicated in a wild type variant termed PHS. In the parental protein (PHS) these residues form the first two turns of a helix running from residue 54 to 68 (hereafter designated as helix I). The crystal structure of the mutant is very similar to that of the parental, with helix I being unaltered. The duplicated residues are accommodated by expanding an existing loop N-terminal to helix I. In addition, circular dichroism (CD) studies have been carried out on a parental peptide containing helix I with six flanking residues at each terminus (residues 48-74) and on the same peptide expanded by the duplication, as a function of 2,2,2-trifluoroethanol (TFE) concentration. Each peptide possesses only modest helical propensity in solution. Our data, which is different from what was observed in T4 lysozyme, show that the conformation of the duplicated sequence is determined by a balance of sequential and longer-range effects. Thus duplicating sequence need not mean duplicating structure. Proteins 2000;40:465-472. PMID- 10861939 TI - The esterase from the thermophilic eubacterium Bacillus acidocaldarius: structural-functional relationship and comparison with the esterase from the hyperthermophilic archaeon Archaeoglobus fulgidus. AB - The esterase from the thermophilic eubacterium Bacillus acidocaldarius is a thermophilic and thermostable monomeric protein with a molecular mass of 34 KDa. The enzyme, characterized as a "B-type" carboxylesterase, displays the maximal activity at 65 degrees C. Interestingly, it is also quite active at room temperature, an unusual feature for an enzyme isolated from a thermophilic microorganism. We investigated the effect of temperature on the structural properties of the enzyme, and compared its structural features with those of the esterase from the hyperthermophilic archaeon Archaeoglobus fulgidus. In particular, the secondary structure and the thermal stability of the esterase were studied by FT-IR spectroscopy, while information on the conformational dynamics of the enzyme were obtained by frequency-domain fluorometry and anisotropy decays. Our data pointed out that the Bacillus acidocaldarius enzyme possesses a secondary structure rich in alpha-helices as described for the esterase isolated from Archaeoglobus fulgidus. Moreover, infrared spectra indicated a higher accessibility of the solvent ((2)H(2)O) to Bacillus acidocaldarius esterase than to Archaeoglobus fulgidus enzyme suggesting, in turn, a less compact structure of the former enzyme. The fluorescence studies showed that the intrinsic tryptophanyl fluorescence of the Bacillus acidocaldarius protein was well represented by the three-exponential model, and that the temperature affected the protein conformational dynamics. The data suggested an increase in the protein flexibility on increasing the temperature. Moreover, comparison of Bacillus acidocaldarius esterase with the Archaeoglobus fugidus enzyme fluorescence data indicated a higher flexibility of the former enzyme at all temperatures tested, supporting the infrared data and giving a possible explanation of its unusual relative high activity at low temperatures. Proteins 2000;40:473-481. PMID- 10861940 TI - Structures of scrambled disulfide forms of the potato carboxypeptidase inhibitor predicted by molecular dynamics simulations with constraints. AB - The structures of two species of potato carboxypeptidase inhibitor with nonnative disulfide bonds were determined by molecular dynamics simulations in explicit solvent using disulfide bond constraints that have been shown to work for the native species. Ten structures were determined; five for scrambled A (disulfide bonds between Cys8-Cys27, Cys12-Cys18, and Cys24-Cys34) and five for the scrambled C (disulfide bonds Cys8-Cys24, Cys12-Cys18, and Cys27-Cys34). The two scrambled species were both more solvent exposed than the native structure; the scrambled C species was more solvent exposed and less compact than the scrambled A species. Analysis of the loop regions indicates that certain loops in scrambled C are more nativelike than in scrambled A. These factors, combined with the fact that scrambled C has one native disulfide bond, may contribute to the observed faster conversion to the native structure from scrambled C than from scrambled A. Results from the PROCHECK program using the standard parameter database and a database specially constructed for small, disulfide-rich proteins indicate that the 10 scrambled structures have correct stereochemistry. Further, the results show that a characteristic feature of small, disulfide-rich proteins is that they score poorly using the standard PROCHECK parameter database. Proteins 2000;40:482 493. PMID- 10861941 TI - Search for the most stable folds of protein chains: III. Improvement in fold recognition by averaging over homologous sequences and 3D structures. AB - Three-dimensional (3D) protein fold recognition by query sequence can be improved using information of fold recognition yielded by the sequences homologous to the query one. This idea is now used more and more widely. Our paper presents its consequent development. We suggest incorporating information both on the sequences homologous to the query protein sequence and the 3D structures homologous to the target (already deciphered) protein folds. We show that both these tricks, and especially their combination reduces errors in fold recognition by the threading method. Proteins 2000;40:494-501. PMID- 10861942 TI - Application of multiple sequence alignment profiles to improve protein secondary structure prediction. AB - The effect of training a neural network secondary structure prediction algorithm with different types of multiple sequence alignment profiles derived from the same sequences, is shown to provide a range of accuracy from 70.5% to 76.4%. The best accuracy of 76.4% (standard deviation 8.4%), is 3.1% (Q(3)) and 4.4% (SOV2) better than the PHD algorithm run on the same set of 406 sequence non-redundant proteins that were not used to train either method. Residues predicted by the new method with a confidence value of 5 or greater, have an average Q(3) accuracy of 84%, and cover 68% of the residues. Relative solvent accessibility based on a two state model, for 25, 5, and 0% accessibility are predicted at 76.2, 79.8, and 86. 6% accuracy respectively. The source of the improvements obtained from training with different representations of the same alignment data are described in detail. The new Jnet prediction method resulting from this study is available in the Jpred secondary structure prediction server, and as a stand-alone computer program from: http://barton.ebi.ac.uk/. Proteins 2000;40:502-511. PMID- 10861943 TI - Dynamics of proteins predicted by molecular dynamics simulations and analytical approaches: application to alpha-amylase inhibitor. AB - The dynamics of alpha-amylase inhibitors has been investigated using molecular dynamics (MD) simulations and two analytical approaches, the Gaussian network model (GNM) and anisotropic network model (ANM). MD simulations use a full atomic approach with empirical force fields, while the analytical approaches are based on a coarse-grained single-site-per-residue model with a single-parameter harmonic potential between sufficiently close (r Butia). The Comet assay was more sensitive and also showed a direct relationship between age and damage, and an inverse relationship between temperature and damage index. PMID- 10861947 TI - Quantitative structure-activity relationship of flavonoids for inhibition of heterocyclic amine mutagenicity. AB - The mutagenic/carcinogenic heterocyclic amines formed during the cooking of protein foods have been determined to be a potential risk to human health. Therefore, mitigation measures are beginning to be studied. A recent finding is that the induction of mutation in Salmonella by these amines can be inhibited by the addition of flavonoids to the assay. This study combines data on the inhibitory process with structural, ab initio quantum chemical, hydropathic, and antioxidant factors to develop a quantitative structure-activity relationship (QSAR) database and statistical analysis. For 39 diverse flavonoids the inhibitory potency varied approximately 100-fold. Three predictive variables, in order of decreasing contribution to variance, are: (1) a large dipole moment; (2) after geometric minimization of energy, a small departure from planarity (i.e., small dihedral angle between the benzopyran nucleus and the attached phenyl ring), and a low rotational energy barrier to achieving planarity; and (3) fewer hydroxyl groups on the phenyl ring. However, these variables account for less than half of the variance in inhibitory potency of the flavonoids. Frontier orbital energies and antioxidant or radical scavenging properties showed little or no relationship to potency. We conclude that interference by the flavonoids with cytochrome P450 activation of the promutagens is the probable mechanism for inhibition of mutagenesis, and suggest avenues for further research. Environ. Mol. Mutagen. 35:279-299, 2000 Published 2000 Wiley-Liss, Inc. PMID- 10861948 TI - In vitro determination of carcinogenicity of sixty-four compounds using a bovine papillomavirus DNA-carrying C3H/10T(1/2) cell line. AB - A new in vitro test for predicting rodent carcinogenicity is evaluated against a testing database of 64 chemicals including both genotoxic and nongenotoxic carcinogens and carcinogens that normally require addition of an S-9 microsomal fraction for detection in the bacterial mutagenicity assay. The assay uses focus formation in a stable, bovine papillomavirus type 1 (BPV-1) DNA carrying C3H/10T(1/2) mouse embryo fibroblast cell line (T1) that does not require transfection, infection with virus, isolation of primary cells from animals, or addition of a microsomal fraction. Of a total database of 64 compounds, 92% of the carcinogens, promoters, or noncarcinogens were correctly predicted. Based on previously reported results, the test of bacterial mutagenicity would have correctly predicted 58% of carcinogens, promoters or noncarcinogens and the Syrian hamster embryo test would have correctly predicted 87% of carcinogens, promoters, or noncarcinogens of this database. Of carcinogens that normally require addition of an S-9 fraction, T1 cells correctly predicted rodent carcinogenicity of polyaromatic hydrocarbons, aflatoxins, azo-compounds, nitrosamines, and hydrazine without the addition of an S-9 fraction. Of nongenotoxic carcinogens, T1 cells correctly predicted diethylstilbestroel, diethylhexylphthalate, acetamides, alkyl halides, ethyl carbamate, and phorbol ester tumour promoters. PMID- 10861949 TI - Cell transformation and genotoxicity induced by bis(2, 3-dichloro-1-propyl) ether. AB - Bis(dichloropropyl) ether isomers have been identified in a petrochemical plant effluent through a toxicity identification evaluation study in the United States. They have also been observed in the microgram per liter range along one of the largest rivers in Europe, the Elbe River. In the present investigation, the genotoxic and transforming activity of a bis(dichloropropyl) ether isomer, bis(2,3-dichloro-1-propyl) ether, was assayed in vitro. The results demonstrate that bis(2,3-dichloro-1-propyl) ether is a potent mutagen in Salmonella typhimurium strains TA 100, TA 1535, and to a lesser extent in strain TA 98, but only when tested in the presence of a metabolic activation system (S9 mix). We have also investigated the induction of micronuclei by bis(2,3-dichloro-1-propyl) ether in the metabolically competent cell line, MCL-5. A linear, dose-dependent increase in micronuclei was observed following exposure to bis(2,3-dichloro-1 propyl) ether. The DNA strand-breaking capacity of this chemical was assessed in the alkaline single-cell gel electrophoresis ("comet") assay with MCL-5 cells. Bis(2,3-dichloro-1-propyl) ether clearly induced DNA strand breaks in the 4.5 45.5 microg/ml dose range. The ether also induced malignant transformation in C3H/M2 mouse fibroblasts after metabolic activation (S9 mix). Thus, it must be suspected that bis(2, 3-dichloro-1-propyl) ether may possess a carcinogenic potential. Since the compound along with its isomers is present in considerable concentrations in surface water, their elimination is a matter of significant public concern. PMID- 10861950 TI - The tumor promoter TPA enhances benzo[a]pyrene and benzo[a]pyrene diolepoxide mutagenesis in Big Blue mouse skin. AB - The Big Blue mouse was used to investigate the role of cell proliferation in mutation fixation in the mouse back skin model of carcinogenesis. Phorbol 12 myristate 13 acetate (TPA) was applied to the dorsum of Big Blue mice to manipulate cell proliferation, and benzo[a]pyrene (BaP) or BaP-diolepoxide (BPDE) was applied to produce premutagenic DNA damage. Mutations in the lacI transgene of skin DNA were measured. BaP and BPDE elevated mutant frequency, DNA adducts, and cell damage over untreated and acetone-treated mice. BPDE-DNA adducts peaked within 30 min of exposure and DNA adducts, formed after application of both BaP and BPDE, declined rapidly with time. As the dose of BaP increased (4 to 64 microg), DNA adducts, mutant frequency, and cell damage increased in a dose dependent manner. TPA applied after BaP and BPDE further increased mutant frequency, DNA adducts, and cell damage, while variably affecting mitotic index and other measures of cell proliferation. TPA became less effective at increasing mitotic index as the dose of BaP increased, although all measures of cell proliferation, taken together, increased. The most effective production of DNA adducts and mutations occurred when the carcinogen was applied simultaneously with or within 1 hr of TPA. Mutations induced by BPDE were predominantly base substitutions: of these base substitutions, 35% were G:C --> A:T transitions, and 36% were G:C --> T:A and 29% G:C --> C:G transversions. Approximately 88% of all mutations and 100% of base substitutions were at G:C sites; 60% of all mutations and 70% of the base substitution mutations occurred at CpG sites. A:T --> G:C transitions were not found. All of the single-base deletions were at G:C base pairs. PMID- 10861951 TI - Activation of MeIQ (2-amino-3,4-dimethylimidazo- [4,5-f]quinoline) by sequence variants of recombinant human cytochrome P450 1A2. AB - Understanding the relationships between the sequences and catalytic activities of P450 enzymes that catalyze the bioactivation of mutagens and carcinogens is an important goal in mutation research. Escherichia coli strain DJ4309 expresses recombinant human P450 1A2 and activates promutagens such as MeIQ (2-amino-3, 4 dimethylimidazo[4,5-f]quinoline), as measured by induction of reverse mutations detected as lacZ(+) colonies on minimal lactose (ML) plates. Pools of P450 1A2 mutants were constructed by polymerase chain reaction (PCR) mutagenesis of putative substrate recognition sites (SRSs). Cultures of individual clones were patched onto MeIQ/ML plates and the growth of revertant microcolonies within each patch was inspected after 2 days of incubation. Beginning with a pool of several thousand clones, we identified 25 distinct P450 1A2 SRS variants with altered activities. In this study, the MeIQ dose-responses of all the variants are reported. The implications of the results are considered with reference to published models of the protein's structure. PMID- 10861952 TI - Differences in the response to oxidative stress and mutant frequency in CD (Sprague-Dawley) and Fisher 344 rats due to an induced inflammatory response. AB - In this study, the rodent air pouch model was used to examine the production and processing of oxidative DNA damage in two strains of rats commonly used in toxicity testing. An inflammatory response was induced by injecting zymosan A (50 mg) into an air pouch on male CD (Sprague-Dawley [S-D]) and Fisher 344 (F-344) rats, and the animals were then sacrificed at 1, 3, 7, 14, and 28 days (n = 6 per time point per strain). Tissues from the lining of the air pouch were collected for 8-hydroxy-2'-deoxyguanosine (8-OH-dG) analysis and for paraffin embedding. Significant (P < 0.01) increases in 8-OH-dG were observed after 1 day in the DNA from cells lining the air pouch of zymosan A-treated versus control S-D (101.5 +/ 27.1 vs. 23.1 +/- 2. 7 8-OH-dG/dG x 10(5)) and F-344 (51.4 +/- 5.3 vs. 14.4 +/- 0.6 8-OH-dG/dG x 10(5)) rats. By 28 days, 8-OH-dG levels had returned to background in S-D rats, but remained elevated in F-344 rats. The frequency of apoptosis was evaluated using the in situ end-labeling (TUNEL) assay, which revealed that zymosan A-treated S-D rats had a significantly (P < 0.05) higher frequency of apoptosis compared to zymosan A-treated F-344 rats. To examine the potential consequences of these differences in endogenously produced DNA damage and apoptosis, we measured mutations at the hprt locus in fibroblasts of the pouch lining and observed a significant (P < 0.05) increase in the mutant frequency at day 28 in F-344 rats (54.2 +/- 13.6 mutants per 10(6) cells) compared to controls (4.5 +/- 2.0 mutants per 10(6) cells). The mutant frequency was not increased in S-D rats. These data demonstrate that strain differences in the production and processing of oxidative DNA damage due to an inflammatory response may impact the long-term pathologic consequences of chronic inflammation. Environ. Mol. Mutagen. 35:336-342, 2000 Published 2000 Wiley-Liss, Inc. PMID- 10861953 TI - Tributyltin induces cytogenetic damage in the early life stages of the marine mussel, Mytilus edulis. AB - Using an integrated approach, the genotoxic potential of bis(tri-n-butyltin), an antifouling agent known to disrupt endocrine system in marine invertebrates, has been evaluated in the embryo-larval stages of the edible mussel, Mytilus edulis. While evaluating the genotoxic potential, the toxicity of tributyltin was also taken into account by determining the maximum tolerated dose. The study suggested that tributyltin is capable of inducing cytogenetic damage (sister chromatid exchanges and chromosomal aberrations) in this target species. The study emphasises the need for further investigations of the potential genotoxic effects of this and other endocrine disrupters on ecologically relevant aquatic invertebrates, which contribute to the maintenance of ecosystems and that could potentially be harmful to human health via the food chain. PMID- 10861954 TI - Asymmetric synthesis of 2,3-methanoleucine stereoisomers from common intermediates. AB - 2,3-Methanoamino acids are useful probes for studying the bioactive conformation of peptides and for investigating the effect of local conformational constraints on the activity of peptidomimetics. We synthesized all four stereoisomers of Cbz protected 2, 3-methanoleucine for incorporation into peptidomimetic inhibitors of calpain. While the synthesis of 2,3-methanoamino acids has been previously reported, our procedure offers a versatile route in which the pair of diastereomers of each geometric isomer was synthesized from a common intermediate. PMID- 10861955 TI - Apparent and true enantioselectivity in enantioseparations. AB - The separation factor of two compounds in chromatography is the ratio of their equilibrium constants or retention factors. This parameter is universally employed to investigate their resolution and to optimize the experimental conditions of their analysis. In enantioseparations, the situation is more complex because there is a mixed retention mechanism. The retention factor is the sum of two contributions, one enantioselective, the other nonselective. Although both contribute to retention, the latter being identical for the two enantiomers and does not contribute to their separation. We show how these two contributions can be measured and how it becomes necessary to distinguish between the apparent, alpha(app), and the true, alpha(true), separation factors. The existence of nonselective sites is responsible for alpha(app) being less than alpha(true). Depending on the difference between these two factors, the more effective approach to improve a separation is either to increase the enantioselectivity or to reduce the nonselective interactions. Practical applications to separations of different beta-blockers on cellobiohydrolase are discussed. The apparent enantioselectivity of alprenolol is larger and increases faster with increasing pH than that of the more hydrophobic propranolol, in spite of the importance of hydrophobic interactions in the enantioselective mechanism. These two unexpected properties are discussed and explained. PMID- 10861956 TI - Chiral discrimination of N-carbazole-carbonyl derivatives of alpha-amino acids with a short linear alkyl side chain by bovine serum albumin. AB - Chiral discrimination of racemic carbazole carbonyl (CC)-amino acids with linear alkyl sidechain (C(1)-C(4)) by bovine serum albumin (BSA) was investigated by competitive replacement experiments using dansyl-L-proline and dansyl-D-norvaline as fluorescent probes. It was found that the CC derivatives of the D-forms of alanine (C(1)), amino butyric acid (C(2)), norvaline (C(3)), and norleucine (C(4)) bound to the dansyl-L-proline site much more strongly than their L-forms, whereas the interactions between both enantiomers of these amino acids with dansyl-D-norvaline site were slight. PMID- 10861957 TI - Enzymatic resolution to (-)-ormeloxifene intermediates from their racemates using immobilized Candida rugosa lipase. AB - In the synthesis of (-)-ormeloxifene, a drug candidate recently under development, enzymatic resolution of potential intermediates can be carried out using a simple, practical method. Five commercially available lipases, Candida rugosa lipase, Candida antarctica lipase B, Mucor miehei lipase, Pseudomonas cepacia lipase, and Humicola lanuginosa lipase, all immobilized on Accurel(R), were initially screened in combination with four different substrates belonging to the class of phenyl esters. Excellent stereoselectivity was observed using C. rugosa lipase with an acetate as substrate, but low reaction rates were observed in scale-up experiments. However, by changing the acyl part of the ester into a hexanoyl moiety and subjecting this substrate to enzymatic hydrolysis in aqueous acetonitrile at room temperature by C. rugosa lipase, it became possible to run the reaction to a 50% conversion on a 10 g scale within a period of 4 h, obtaining a phenolic product of more than 95% ee that could be converted to the target molecule, (-)-ormeloxifene, in two synthetic steps. Simple recovery of the immobilized enzyme by filtration allowed multiple recycling of the catalyst without significant loss of enzymatic activity. Capillary electrophoresis with sulfobutyl ether beta-cyclodextrin as a chiral buffer additive and acetonitrile as an organic modifier was demonstrated to provide an excellent chiral analytical tool for monitoring the enzymatic reactions. PMID- 10861958 TI - Trifluoromethanesulfonamide, diphenylphosphoramide and diphenylthiophosphoramide of (R)-(+)-1,1'-binaphthyl-2,2'-diamine as chiral catalyst ligands for the titanium(IV) alkoxide-promoted addition of diethylzinc to aldehydes AB - Chiral trifluoromethanesulfonamide 4, diphenylphosphoramides 5 and 6, and phenylthiophosphoramide 7 were prepared from the reaction of trifluoromethanesulfonic anhydride, diphenylphosphinic chloride, and diphenylthiophosphinic chloride with (R)-(+)-1,1'-binaphthyl-2, 2'-diamine, respectively. They were used as catalytic chiral ligands in the asymmetric addition reaction of diethylzinc to aldehydes in the presence of titanium(IV) isopropoxide to give the corresponding sec-alcohols with 43-54%, 18-22%, 30-34%, and 52-64% enantiomeric excess, respectively. Copyright 2000 Wiley-Liss, Inc. PMID- 10861959 TI - Total synthesis of gigantetrocin A. AB - A highly efficient synthetic method for the trans-tetrahydrofuran (THF) ring building block was established and the title compound was synthesized in 19 steps from trans-1,4-dichloro-2-butene via a convergent route with a Wittig reaction as the key step. PMID- 10861962 TI - Publisher's note PMID- 10861960 TI - Chiral high-performance liquid chromatographic analysis of antifungal SCH 56592 and evaluation of its chiral inversion in animals and humans. AB - SCH 56592 is a novel triazole antifungal agent that is active both orally and intravenously in animal models of infection. This compound is in Phase II-III clinical trials for the treatment of systemic fungal infections. SCH 56592 is a single enantiomer with four stereogenic centers; therefore, it was necessary to evaluate the possible chiral inversion of this drug candidate in animals and humans. Thus, chiral high-performance liquid chromatographic (HPLC) methods have been developed to separate SCH 56592 from its diastereomers and to evaluate its chiral inversion in rats, dogs, cynomolgus monkeys, and humans. Chiral HPLC analysis involved the use of a Chiralcel OD column set at 39 degrees C with a mobile phase of hexane-ethanol-diethylamine and a fluorescence detector set at an excitation wavelength of 270 nm and an emission wavelength of 390 nm. Plasma or serum samples were subjected to solid phase extraction on a C(2) cartridge followed by HPLC analysis. The method was sensitive with a limit of quantitation of 0.1 microg/ml in dog serum. The linearity was satisfactory, as shown by correlations of >0.997 and by visual examination of the calibration curves. The precision and accuracy were satisfactory, as indicated by coefficients of variation (CV) ranging from 1.1 to 12.1% and bias values ranging from -11.0 to 9.0%. Chiral HPLC analysis indicated that SCH 56592 was not subjected to chiral inversion in rats, dogs, cynomolgus monkeys, and humans. PMID- 10861963 TI - Phytochemistry, pharmacology and toxicity of Rhazya stricta decne: a review. AB - Phytochemical, pharmacological and toxicological properties of the medicinal plant Rhazya stricta Decne. are reviewed. Several types of alkaloids and a few flavonoids have been isolated and their structures and stereochemistry characterized. However, in most cases the biological activity of these compounds has not been studied. Most of the pharmacological activity of the plant resides in its alkaloidal fractions which cause depression of the central nervous system and hypotension. Extracts of R. stricta appear to have low toxicity, although its use in pregnant women may be inadvisable. PMID- 10861964 TI - Blood pressure depression by the fruit juice of Carica papaya (L.) in renal and DOCA-induced hypertension in the rat. AB - A crude ethanol extract was prepared from the unripened fruit of Carica papaya. Lethality studies showed a dose-mortality relationship with an LD(50) of 325.2 mg/kg in mice administered i.p. Male albino Wistar rats were randomly divided into three batches (15 rats per batch)-renal, DOCA-salt hypertensives and normotensives. Each batch was further divided into three groups-the untreated, hydrallazine and extract treated groups. The mean arterial blood pressure (MAP) and the heart rate were measured in all groups. From the results, the basal (control) MAP were 93.8 +/- 4.5, 175.2 +/- 5. 1 and 181.3 +/- 6.2 mmHg in the normotensive, renal and DOCA-salt hypertensives, respectively. Both hydrallazine (200 microg/100 g i. v) and extract (20 mg/kg.i.v) produced a significant depression of MAP in all groups (p < 0.01 vs controls), but the extract produced about 28% more depression of MAP than hydrallazine in the hypertensive groups. In another group of rats, the extract failed to depress the MAP in rats pretreated with propranolol, but atropine and noradrenaline pretreatment did not prevent the action of the extract on blood pressure. In vitro studies using isolated rabbit arterial (aorta, renal and vertebral) strips showed that the extract (10 microg/mL) produced relaxation of vascular muscle tone which was, however, attenuated by phentolamine (0.5-1.5 microg/mL). It is concluded that the fruit juice of C. papaya probably contains antihypertensive agent(s) which exhibits mainly alpha-adrenoceptor activity. PMID- 10861965 TI - Antiinflammatory and antinociceptive effects of 1,8-cineole a terpenoid oxide present in many plant essential oils. AB - 1,8-Cineole (cineole), a terpenoid oxide present in many plant essential oils displays an inhibitory effect on some types of experimental inflammation in rats, i.e. paw oedema induced by carrageenan and cotton pellet-induced granuloma. Cineole also inhibits in mice, the acetic acid-induced increase in peritoneal capillary permeability and the chemical nociception induced by intraplantar formalin and intraperitoneal acetic acid. Activity was present in these tests, at an oral dose range of 100-400 mg/kg. In the formalin test, the antinociceptive effect of cineole was not reversed by pretreatment of mice with naloxone (1 mg/kg, s.c.), a mu-opioid receptor antagonist, suggesting the involvement of a non-opioid mechanism. Cineole demonstrated a significant inhibitory effect on locomotion and also potentiated the pentobarbital sleeping time in mice, indicating a plausible depressant effect on the central nervous system. The present results, when taken together with the recent reports that describe the inhibitory effects of cineole on the formation of prostaglandins and cytokines by stimulated monocytes in vitro, may provide additional evidence for its potential beneficial use in therapy as an antiinflammatory and analgesic agent. PMID- 10861966 TI - Protection of epithelial cells against influenza A virus by a plant derived biological response modifier Ledretan-96. AB - A multicomponent herbal formula Ledretan-96 was tested on an epithelial tissue culture cell line (MDCK) for its protective activity against cytopathic effects caused by influenza A virus. The whole formula and each of its 23 individual components were tested in the same system. The results indicated that the formula, when prepared according to established procedure, in the form of decoction, is active in protecting epithelial cells against damage caused by influenza A virus used at different dosages. Of the 23 components tested, only one, Terminalia chebula, showed a significant protective effect when applied to the epithelial cells individually. Controls for these studies included cell cultures exposed to individual components and the complete formula without virus; the cultures were monitored for any toxic effects by morphology and protein synthesis. The protective effects of Ledretan-96 and Terminalia chebula could be distinguished from toxicity against the epithelial cells. In addition, the results indicated that the complete formula maintained antiviral activity at a higher therapeutic index than the Terminalia chebula extract alone. PMID- 10861967 TI - Application of flow injection--chemiluminescence to the study of radical scavenging activity in plants. AB - Chemiluminescence (CL) was observed during the oxidation of luminol (2 mg/L). mediated by 0.06% hydrogen peroxide (H(2)O(2)) and cytochrome c (10 mg/L). CL intensity was decreased by the presence of radical scavengers and the reduction was linearly proportional to the concentration and ability of scavengers; butylated hydroxytoluene (BHT), caffeic acid and gallic acid. The order of effectiveness as radical scavengers was gallic acid > caffeic acid > BHT, which shows that the number of hydroxyl groups (OH) in the B-ring of flavonoids plays a key role in a good radical scavenging activity. Of eight catechins obtained from green tea extracts, (-)-catechin was the least effective and (-)-epigallocatechin gallate (EGCg) showed the strongest activity. This result indicates that the stereoscopic structure between the C-3 group and the B ring of flavonoids as well as substituents at the C-3 position make a contribution to radical scavenging activity. Of the tested Chinese herbal ingredients, five species of ingredients represented more than 90% of the radical scavenging activity. PMID- 10861968 TI - Biological disposition of boldine: in vitro and in vivo studies. AB - Boldine is a natural compound with well-established free radical scavenger and hepatoprotective properties. The further exploration of its actual therapeutic potential as an antioxidant is, however, partially limited by the absence of knowledge on its pharmacokinetics. In the present studies, we provide information on the in vitro and in vivo biological disposition of boldine. The addition of 200 microM boldine to an isolated rat hepatocyte suspension was followed by a time-dependent (0-60 min) disappearance of boldine from the extracellular medium. This decline was associated with an early (first 2 min) and swift accumulation (1600 microM) of boldine within the cells. Although the intracellular concentration of boldine diminished, boldine was always found to occur within the cells at concentrations substantially higher than those initially added to the preparation. Boldine was also concentration-dependently removed from the extracellular medium by isolated rat livers portally perfused with the antioxidant. In vivo studies, conducted in rats, revealed that following either its oral or its intravenous administration, plasma boldine concentrations declined rapidly and according to an apparently first order type of kinetics. After its oral administration (50 or 75 mg/kg), boldine was rapidly (within 30 min) absorbed and preferentially concentrated in the liver, with substantially lower concentrations being found in the brain and heart. Maximal hepatic concentrations of boldine were found to be equal to or greater than those needed to afford antioxidant and hepatoprotective effects in vitro. PMID- 10861969 TI - Aloe polymannose enhances anti-coxsackievirus antibody titres in mice. AB - Aloe polymannose (AP), a high mannose biological response modifier (BRM) purified from the Aloe barbadensis Miller plant, was tested for activity in enhancing antibody titres against coxsackievirus B3 (CVB3) and CVB3-induced myocarditis in murine models of the disease. Inoculation of mice with AP over a range of three nontoxic doses and in varying schedules did not reduce virus titres in heart tissues or ameliorate virus-induced cardiopathological alterations during acute disease. However, this BRM was found to significantly enhance titres of anti-CVB3 antibodies produced during acute infection of three strains of mice with CVB3. Simultaneous intraperitoneal inoculation of AP at a dose of 0.5 mg/kg body weight per mouse with purified CVB3 significantly increased ELISA titres of anti-CVB3 antibodies and the proportion of mice with these titres, compared with similar parameters in mice inoculated only with CVB3. The data conclusively show that AP can immunopotentiate antibody production against capsid protein epitopes of a nonenveloped picornavirus and suggest this BRM (AP) might be of benefit in enhancing antibody titres against other enteroviruses during a natural infection and poliovirus vaccine strains. PMID- 10861970 TI - Antimicrobial activity of essential oils and ethanol extract of Phlomis fruticosa L. (Lamiaceae). AB - The essential oils and an ethanol extract of Phlomis fruticosa L. were evaluated for antibacterial and antifungal activities. Seven bacterial and seven fungal species were used. Among them were human, animal and plant pathogens, food poisoning bacteria and fungi which are known as potential mycotoxin producers. The essential oils showed antibacterial activity against Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae and Micrococcus luteus. The essential oils extracted from the plants collected from two different localities showed similar antibacterial activities. The antifungal activity of the essential oils was positive against Aspergillus niger, A. ochraceus, Cladosporium cladosporioides, Fusarium tricinctum and Phomopsis helianthi. The ethanol extract showed antibacterial activity against Staphylococcus aureus and Bacillus subtilis and antifungal activity against Aspergillus niger, A. ochraceus, Cladosporium cladosporioides, Fusarium tricinctum and Phomopsis helianthi. PMID- 10861971 TI - Evaluation of antipyretic potential of Nelumbo nucifera stalk extract. AB - The ethanol extract of stalks of Nelumbo nucifera (NNSE) was evaluated for its antipyretic potential on normal body temperature and yeast induced pyrexia in rats. NNSE showed significant activity in both the models at oral doses of 200 and 400 mg/kg. NNSE at a dose of 200 mg/kg was found to produce significant lowering of normal body temperature up to 3 h and at 400 mg/kg it caused significant lowering of body temperature up to 6 h after its administration. In the model of yeast provoked elevation of body temperature NNSE showed dose dependent lowering of body temperature up to 4 h at both the doses and the results were comparable to that of paracetamol, a standard antipyretic agent. PMID- 10861972 TI - Antinociceptive effect of some Argentine medicinal species of Eupatorium. AB - Eupatorium laevigatum, E. arnottianum and E. subhastatum, plants used in Argentine folk medicine for the treatment of inflammation and pain related problems, were evaluated for analgesic activity. The infusions of these species (500 mg/kg, p.o.) produced a reduction in the number of stretches of 46.6%, 41.5% and 35.6% respectively, in the acetic acid induced writhing test. This antinociceptive effect of the infusions was not reversed by pretreatment with naloxone. The infusions studied did not produce antinociceptive effects when assayed in the hot plate test. These results suggest that the analgesic activity is exerted by a mechanism unrelated to interaction with opioid systems. PMID- 10861973 TI - Studies on antibacterial activity of Ficus racemosa Linn. leaf extract. AB - Extracts of Ficus racemosa Linn. leaves were tested for antibacterial potential against Escherichia coli ATCC 10536, Basillus pumilis ATCC 14884, Bacillus subtilis ATCC 6633, Pseudomonas aeruginosa ATCC 25619 and Staphylococcus aureus ATCC 29737. The effects produced by the extracts were significant and were compared with chloramphenicol. The petroleum ether extract was the most effective against the tested organisms. PMID- 10861974 TI - The repellant and antifeedant properties of Cyperus articulatus against Tribolium casteneum Hbst. AB - Cyperus articulatus is an insect repellant plant commonly found in Northern Nigeria and used traditionally in pest control. The light petroleum and methanol extracts of the plant's rhizome were evaluated against Tribolium casteneum Hbst (the red flour beetle) using standard techniques. The methanol extract showed more antifeedant property than the light petroleum extract, while both the extracts were observed to have similar repellant actions. PMID- 10861975 TI - Isothiocyanates in myrosinase treated herb extract of Cleome chrysantha decne. and their antimicrobial activities. AB - GC/MS analysis of the volatiles produced by the action of endogenous myrosinase in Cleome chrysantha Decne. herb, showed three major components, 1-isocyano-4 methyl benzene, gamma-muurolene and (cis) nerolidole (21.72%, 12.15% and 10.39%, respectively). 1-isocyano-4-methyl benzene is not produced from myrosinase treated seeds of Cleome chrysantha Decne., while muurolene and nerolidole were found at higher concentrations 16.1% and 13.67%, respectively. Some other isothiocyanates identified in the seeds and leaves, but in low percentages, were 2-methylbutyl isothiocyanate, 4-methylthiobutyl isothiocyanate and isothiocyanatomethyl benzene. Enzymatic hydrolysis of the ethanol extract of Cleome chrysantha Decne., produced two non-volatile isothiocyanates, 4, 6 dimethyltetrahydro-1,3-oxazine-2-thione and N-(-4-methyl-sulphinyl-3-butenyl) isothiocyanate which was established by aniline and methanolic ammonia to give the stable form of phenyl thiourea. They were identified by their spectroscopic data. These two isothiocyanates and the volatiles of the herb showed good antimicrobial activity against E. coli, Pseudomonas putida and Rhizobium meloloti (gram-negative bacteria) and moderate activity against Bacillus subtilis and Streptococcus lactis (gram-positive bacteria). PMID- 10861976 TI - The effect of extracts of Cynomorium coccineum and Withania somnifera on gonadotrophins and ovarian follicles of immature Wistar rats. AB - The effects of water extracts of Cynomorium coccineum and Withania somnifera on ovarian follicular development and serum levels of FSH and LH were studied in immature 17-day-old and 25-day-old-Wistar rats. Water extracts of the plants were given to the animals per os in a dose of 47 mg/100 g body weight for 6 days. Serum levels of FSH and LH were measured by ELISA. Folliculogenesis was studied with a light microscope. In 25-day-old rats, extracts of both plants elicited significant changes in gonadotrophin levels coupled with a significant increase in ovarian weight and profound folliculogenesis. Numerous primary, secondary, tertiary and antral follicles were present. A distinct zona pellucida was not seen and the oocyte was often detached. In 17-day-old animals there was a significant increase-in body weight but without significant changes in the ovarian weight and folliculogenesis. PMID- 10861977 TI - Garlic and neem leaf extracts enhance hepatic glutathione and glutathione dependent enzymes during N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced gastric carcinogenesis in rats. AB - The protective effect of garlic (Allium sativum L.) and neem leaf (Azadirachta indica A. Juss.) was investigated on hepatic lipid peroxidation and antioxidant status during N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced gastric carcinogenesis in male Wistar rats. Enhanced lipid peroxidation in the liver of tumour-bearing animals was accompanied by significant decreases in the activities of glutathione peroxidase (GPx), glutathione-S-transferase (GST), gamma-glutamyl transpeptidase (GGT) and reduced glutathione (GSH) levels. Administration of garlic and neem leaf extracts significantly lowered lipid peroxidation and enhanced the hepatic levels of glutathione and glutathione dependent enzymes. We speculate that garlic and neem leaf significantly alter cancer development at extrahepatic sites by influencing hepatic biotransformation enzymes and antioxidants. PMID- 10861978 TI - Inhibitory effects of astragali radix, crude drug in Oriental medicines on lipid peroxidation and protein oxidative modification of mouse brain homogenate by copper. AB - Astragali Radix, the root of Astragalus membranaceus Bunge, is a crude drug used widely in Oriental medicines. It is a major component of Ougi-Keishi-gomotsu-to, a traditional herbal medicine, used for neurop patients with abnormal sensations and neuropathic pain of the legs. It was shown to have inhibitory effects on lipid peroxidation and protein oxidative modification by copper. The effects were similar to and stronger than those of mannitol and superoxide dismutase as free radical scavengers. These results demonstrated that Astragali Radix has inhibitory effects on oxidative stress induced by metal. PMID- 10861979 TI - Patents alert PMID- 10861980 TI - Selected bibliography PMID- 10861981 TI - Rapid determination of chlorogenic acid and related compounds in sunflower seeds by high-performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry. AB - High-performance liquid chromatography (HPLC) in combination with atmospheric pressure chemical ionization mass spectrometry (APcI-MS) was applied to the determination of the phenolic fraction found in methanolic extracts of sunflower seeds (mainly chlorogenic acid and derived compounds). These extracts were directly separated by HPLC and detected by both negative and positive APcI-MS. Abundant structural information about these compounds can be obtained even at low extraction cone voltages. This method has been shown to be a rapid and effective method for the analysis of crude extracts from sunflower seeds. PMID- 10861982 TI - Liquid chromatography/electrospray tandem mass spectrometry method for the quantitation of fosinoprilat in human serum using automated 96-well solid-phase extraction for sample preparation. AB - A sensitive, specific, accurate and reproducible liquid chromatography/electrospray tandem mass spectrometry method was developed and validated for the quantitation of fosinoprilat in 0.2 mL of human serum. The method employed acidification (with pH 4.0 sodium acetate buffer) of the serum samples to minimize the hydrolysis of the prodrug fosinopril to fosinoprilat prior to purification by automated 96-well solid-phase extraction. The required chromatographic separation of fosinoprilat and fosinopril was achieved isocratically on a Luna C8 analytical column (2 x 50 mm, 3 microm). The total run time was 2 min. The mobile phase contained methanol and water with 10 mM ammonium acetate. Detection was by positive ion electrospray tandem mass spectrometry. The standard curve, which ranged from 2.00 to 500 ng/mL, was fitted to a 1/x(2) weighted linear regression model. Fosinoprilat quality control (QC) samples used to determine the accuracy and precision of the method were prepared in human serum at concentrations of 5.00, 200, 400 and 1000 ng/mL. The assay accuracy was within 8% (dev). The intra- and inter-assay precisions were within 6 and 3% (RSD), respectively. Fosinopril QC samples used to gauge the rate of hydrolysis of fosinopril to fosinoprilat during the assay procedure were prepared in human serum at 500 ng/mL. The hydrolysis of fosinopril to fosinoprilat was 90%) is possible from the CID spectra of singly charged peptides using the SCIEX Predict Sequence routine. Ion production at pressures near 1 Torr (rather than in vacuum) is found to give reduced metastable fragmentation, particularly for higher mass molecular ions. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10861987 TI - Consequence of boar edible tissue consumption on urinary profiles of nandrolone metabolites. I. Mass spectrometric detection and quantification of 19 norandrosterone and 19-noretiocholanolone in human urine. AB - For the first time in the field of steroid residues in humans, demonstration of 19-norandrosterone (19-NA: 3alpha-hydroxy-5alpha-estran-17-one) and 19 noretiocholanolone (19-NE: 3alpha-hydroxy-5beta-estran-17-one) excretion in urine subsequent to boar consumption is reported. Three male volunteers agreed to consume 310 g of tissues from the edible parts (meat, liver, heart and kidney) of a boar. The three individuals delivered urine samples before and during 24 h after meal intake. After deconjugation of phase II metabolites, purification and specific derivatisation of target metabolites, the urinary extracts were analysed by mass spectrometry. Identification was carried out using measurements obtained by gas chromatography/high resolution mass spectrometry (GC/HRMS) (R = 7000) and liquid chromatography/tandem mass spectrometry (LC/MS/MS) (positive electrospray ionisation (ESI+)). Quantification was realised using a quadrupole mass filter. 19-NA and 19-NE concentrations in urine reached 3.1 to 7.5 microg/L nearby 10 hours after boar tissue consumption. Levels returned to endogenous values 24 hours after. These two steroids are usually exploited to confirm the exogenous administration of 19-nortestosterone (19-NT: 17beta-hydroxyestr-4-en-3-one), especially in the antidoping field. We have thus proved that eating tissues of non-castrated male pork (in which 17beta-nandrolone is present) might induce some false accusations of the abuse of nandrolone in antidoping. PMID- 10861988 TI - Sodium ion attachment reactions in an ion trap mass spectrometer AB - The capabilities of ion traps to perform attachment reactions with alkali cations using classical scanning sequences have been exploited here with an ion trap mass spectrometer equipped with an external ion source to generate the reagent Na(+) ions. Kinetic studies have shown that, as expected, the attachment efficiency is very high, near-collision efficiency, and illustrate how the present method is particularly well suited for ion trap mass spectrometers. The large adaptability of the experimental conditions suggests that a wide range of organic molecules, characterized by a large alkali ion affinity, could be readily detected even at low levels. Applications of sodium ion attachment reactions are illustrated by the detection and characterization of explosives and some of their correlated pyrolytic degradation products. Detection -limits for phthalate compounds are shown to reach the low ng range of injected samples, without any noticeable difficulties in the full scan mode of acquiring mass spectra. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10861989 TI - Electron impact induced mass spectral study of 2- and 4-ethoxycarbonylalkylthio-5 bromo-6-methyluracils AB - Electron impact induced mass spectral fragmentation of ten new 2- and 4 ethoxycarbonylalkylthio-5-bromo-6-methyluracils have been investigated. Fragmentation pathways are proposed on the basis of accurate mass and B/E and B(2)/E linked scan spectra measurements. The data obtained create the basis for the distinction of isomers. Copyright 2000 John Wiley & Sons Ltd. PMID- 10861990 TI - Fragmentation mechanisms in electron ionization mass spectrometry of some new thiazoles PMID- 10861991 TI - Notes on current safety issues in MRI. AB - The present unsatisfactory state of the worldwide regulatory system for whole body magnetic resonance is explored. The priorities of a number of the most important regulators are outlined, and the differences between factors affecting the safety of patients on the one hand, and of those operating equipment on the other, are discussed. At the end, a strategy is outlined for obtaining the data needed to establish new, more firmly based, limits of operation. A prime factor in getting this data will be collaboration between the regulators, users of equipment, and manufacturers. A measure of urgency is desirable as the longer the present situation is allowed to persist the greater the risk that the stresses already present in it will become intolerable. PMID- 10861992 TI - High-field quantitative transverse relaxation time, magnetization transfer and apparent water diffusion in experimental rat brain tumour. AB - The potential of quantitative parameter images of transverse relaxation time T(2), apparent diffusion coefficient (ADC) and magnetization transfer ratio (MTR) to characterize experimental brain tumours was studied. Necrosis or haemorrhage can be detected using either MTR, ADC or T(2) (necrosis-MTR reduced by 35%, ADC and T(2) increased respectively by 170% and 100% compared with normal brain tissue; haemorrhage-MTR increased by 60%, ADC and T(2) decreased by 40% and 20%, respectively). Normal brain tissue can only be distinguished from tumour on T(2) and MTR parameter images. However, for small tumours (10 microl), the best contrast is observed with MTR, ca. 30%, whereas for T(2) the contrast is ca. 10%. PMID- 10861993 TI - Double-resonance J-edited (1)H-NMR detection of 6-(13)C-D-2-deoxyglucose uptake in glioma cells. AB - The C6 methylene protons were selectively detected in (1)H-NMR spectra of intact glioma cells incubated with 6-(13)C-D-2-deoxyglucose (6-(13)C-2dG), a (13)C enriched glucose analog that is suitable for monitoring glucose utilization in brain tumors. Spectral editing via (1)H-(13)C scalar coupling was performed with twin spin-echo double resonance (T-SEDOR), a pulse sequence which combines chemical specificity and high sensitivity, requires no solvent pre-saturation, and can easily be adapted to imaging protocols. This work demonstrates the suitability of the pulse sequence for monitoring 6-(13)C-2dG uptake in living cells in vitro, in spite of line-broadening and the occurrence of other strong signals in the spectral region of interest (3.5-4.4 ppm). PMID- 10861994 TI - Proton NMR chemical shifts and coupling constants for brain metabolites. AB - Proton NMR chemical shift and J-coupling values are presented for 35 metabolites that can be detected by in vivo or in vitro NMR studies of mammalian brain. Measurements were obtained using high-field NMR spectra of metabolites in solution, under conditions typical for normal physiological temperature and pH. This information is presented with an accuracy that is suitable for computer simulation of metabolite spectra to be used as basis functions of a parametric spectral analysis procedure. This procedure is verified by the analysis of a rat brain extract spectrum, using the measured spectral parameters. In addition, the metabolite structures and example spectra are presented, and clinical applications and MR spectroscopic measurements of these metabolites are reviewed. PMID- 10861995 TI - Diffusion weighted imaging and magnetic resonance spectroscopy in a low flow ischaemia model due to endothelin induced vasospasm. AB - The aim of this MR study was to determine if vasospasm induced by application of endothelin-1 (ET-1) in the rat brain would model the abnormalities attributed to vasospasm described in patients with subarachnoid haemorrhage (SAH) with reversible neurological deficits. Following application of ET-1 in concentrations of 10(-4) M or 10(-6) M to the middle cerebral artery, there was an immediate drop in pH, an increase in the inorganic phosphate (P(i)) to phosphocreatine (PCr) ratio and elevated lactate. There was gradual recovery to control in the 10(-6) M group, but in the 10(-4) M group there was a loss of approximately 10% in the absolute signal intensities of PCr and adenosine triphosphate (ATP). In a second similarly treated group of animals, the area of the hemisphere with a low apparent diffusion coefficient (ADC) was 27 +/- 6% at 30 min and remained at about 20-21% at 90 min and beyond. Together these data suggest that the regions with persistently low ADC were metabolically compromised, with incomplete recovery of PCr and ATP, and represent irreversibly damaged tissue. This raises the possibility that MR spectroscopy and imaging could be a sensitive indicator of tissue viability. This is a potentially useful model of low flow as seen in clinical vasospasm following SAH. PMID- 10861996 TI - Quantification of short echo time (1)H signals: a response letterp. PMID- 10861997 TI - Current awareness in NMR in biomedicine AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of NMR in biomedicine. Each bibliography is divided into 9 sections: 1 Books, Reviews ' Symposia; 2 General; 3 Technology; 4 Brain and Nerves; 5 Neuropathology; 6 Cancer; 7 Cardiac, Vascular and Respiratory Systems; 8 Liver, Kidney and Other Organs; 9 Muscle and Orthopaedic. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. PMID- 10861998 TI - Identification of genes expressed in the epithelium of porcine oviduct containing early embryos at various stages of development. AB - As a first step toward elucidation of the action of factors secreted by the epithelium of oviduct, differential display reverse transcription-polymerase chain reaction (DDRT-PCR) was used in this study to identify transcripts of such oviductal factors in gilts carrying various stages of early embryo development post hormone-induced ovulation. A total of 13 differentially expressed transcripts were identified between 50 and 120 hr post-hCG injection (between 1- and 8-cell embryonic stages). Twelve of these transcripts were found to be initially expressed at 96 hr post-hCG injection (at 4-cell embryonic stage) and beyond. Three of such genes were shown by sequence analysis to be the porcine transforming growth factor-alpha, the porcine transforming growth factor-beta binding protein II and a porcine astral natriuretic factor receptor-like transcript. Only one differentially expressed gene was detected between 50-60 and 85 hr post-hCG injection, and this gene turned out to be the porcine follicle stimulating hormone receptor. The remaining eight transcripts detected by DDRT PCR were novel. Moreover, most of these newly expressed genes were found to be turned on at a time coincidental with that of the 4-cell block of porcine embryos cultured in vitro. Our results demonstrate that DDRT-PCR is a feasible approach for rapid identification of genes that are differentially expressed in oviductal epithelium. Some of the genes thus identified may be important for unhindered development of embryos in the oviduct. PMID- 10861999 TI - Xenopus laevis TRK-fused gene (TFG) is an SH3 domain binding protein highly expressed in the cement gland. AB - TRK-fused gene (TFG) was originally identified in humans as the N-terminus of an oncogenic fusion protein TRK-T3, associated with papillary thyroid carcinoma. An amino-terminal coiled coil domain of TFG is responsible for mediating oligomerization of the TRK-T3 oncoprotein, resulting in constitutive activation of the TRK protein tyrosine kinase and oncogenesis. We have cloned the Xenopus laevis homologue of TFG and demonstrated that xTFG was highly expressed in the cement gland of tailbud embryos. Overexpression of xTFG2-136 (including the coiled coil domain) in early embryos, via mRNA microinjection as well as transgenic expression using the recently described restriction enzyme mediated integration (REMI) transgenesis, did not alter embryonic development or development of a functional cement gland, despite clear evidence that xTFG2-136 strongly interacted with endogenous xTFG. Finally, we have identified a potential SH3 binding motif in xTFG (and in TFG) and have demonstrated that xTFG selectively interacted with SH3 domains of Src, PLCgamma, and the p85 phosphatidylinositol 3-kinase subunit. PMID- 10862000 TI - Monoalleleic transcription of the insulin-like growth factor-II gene (Igf2) in chick embryos. AB - A polymorphism in the igf2 gene of chickens was identified using NlaIII (GenBank accession number AF218827). In some embryos, the igf2 alleles were expressed monoallelically from either maternal or paternal alleles. These data demonstrate that genomic imprinting is not confined to mammalian vertebrates and suggest that genomic imprinting evolved at an early stage of vertebrate evolution. The observations that the igf2 gene is imprinted in a minority of embryos suggest that the imprinting in birds is unrelated to embryonic growth. Genome imprinting may provide opportunities for evolution of genes in a nonexpressed state. In poultry breeding, the presence of imprinted genes may make a major contribution to unequal performance in reciprocal matings between commercial lines. PMID- 10862001 TI - Apoptosis regulation in the testis: involvement of Bcl-2 family members. AB - Using immunohistochemical techniques and Western blot analysis, the possible role of Bcl-2 family members Bax, Bcl-2, Bcl-x(s), and Bcl-x(l) in male germ cell density-related apoptosis and DNA damage induced apoptosis was studied. The apoptosis inducer Bax was localized in all mouse and human testicular cell types, but despite the fact that irradiation induces its transcriptional activator, p53 in the human, Bax expression did not change after irradiation. The apoptosis inhibitor Bcl-2 appeared to be present in late spermatocytes and spermatids and was up-regulated in these cells after a dose of 4 Gy of X-rays. Finally, Bcl-x was expressed in both the mouse and human testis. The apoptosis inhibiting long transcripts of Bcl-x, Bcl-x(l), were expressed in spermatogonia and spermatocytes and were up-regulated after X-irradiation. The apoptosis inducing shorter form of Bcl-x, Bcl-x(s), was found to be expressed only in somatic cells, like peritubular and Leydig cells. While Bax is important in germ cell density regulation, Bax expression did not change after DNA damage inflicted by X radiation. Hence, spermatogonial apoptosis after X-irradiation may not be induced via the apoptosis inducer Bax. Furthermore, as Bcl-x(l), but not Bcl-2, is present in spermatogonia and spermatocytes, Bcl-x(l) may regulate germ cell density, possibly in cooperation with Bax. As Bcl-x(l) expression is enhanced after irradiation, this protein may also have a role in the response of spermatogonia and spermatocytes to irradiation. PMID- 10862002 TI - Transfection of mouse eggs and embryos using DNA combined to cationic liposomes. AB - We have used plasmid DNA in combination with cationic liposomes to transfect mouse eggs and embryos. The plasmid was rhodamine labeled, which allowed a direct visualization of the DNA uptake by the cells. Immature eggs, collected from the ovaries, were easily transfected, but once the egg was ovulated the zona pellucida (ZP) acted as a barrier and prevented transfection. Permeabilization or removal of the ZP was therefore a requirement to allow transfection. Transfected eggs were capable of being fertilized in vitro giving raise to embryos that expressed the recombinant protein. Morulae and blastocysts were also transfected when the ZP was permeabilized, but the efficiency of transfection decreased and in some cases not all the blastomeres incorporated the plasmid. Pronuclear embryos were cultured and showed expression of the transgene from the 2-cell stage. This indicates that liposome-transfection of oocytes or pronuclear embryos could be a simple and suitable method to introduce foreign genes in embryos and perhaps could be also useful to generate transgenic animals. PMID- 10862003 TI - Effect of short interspersed element sequences on the integration and expression of a reporter gene in the preimplantation-stage mouse embryos. AB - Based on the assumption that foreign DNA sequences may have increased chance of integration into the host genome if they are flanked by high copy-numbered genomic sequences such as SINEs (short interspersed elements), we investigated the integration frequency of Lac Z reporter gene flanked by a fused B1/B2 in an in vivo system using pronuclear microinjection technique in the mouse. The SINE flanked DNA showed a 4-fold increased integration frequency of the reporter gene than the control DNA (63% vs. 16%). Moreover, the level of beta-galactosidase expression, estimated from the X-Gal staining intensity in transgenic embryos, was greatly higher in SINE-carrying DNA. These results suggest that the SINE sequences can serve a very useful tool in improving the efficiency of current transgenic animal technology. PMID- 10862004 TI - Electrofusion of two-cell bovine embryos for the production of tetraploid blastocysts in vitro. AB - Tetraploid bovine blastocysts were produced experimentally by electrofusion of in vitro matured and fertilized, zona-enclosed two-cell embryos (33-35 hr after initiation of sperm-egg incubation) using three fusion protocols. Field strengths of 1.0, 1.4, and 2.4 kV/cm were tested and the rate of fusion, subsequent cleavage, and blastocyst development were measured for each. High rates of fusion (76.5% +/- 2.8%), cleavage (72.5% +/- 7.4%) and blastocyst development (56.1% +/- 6.4%) were achieved with the application of 1. 4 kV/cm as a single 100 microseconds pulse. Embryos were scored 30 and 60 min after stimulation for fusion. No time effect for fusion, cleavage, or blastocyst development was observed. Chromosome preparations of day 7 blastocysts revealed 12.5% of fused embryos were tetraploid. This is a significant increase from that found in nonfused embryos where spontaneous tetraploidy did not occur. An electrical stimulus of 1.0 kV/cm applied as two 50-microseconds pulses produced significantly less one-cell embryos (64.2% +/- 3.0%) compared to 1.4 kV/cm while cleavage (79.9% +/- 3.4) and blastocyst development (44.6% +/- 4.0%) were not different from that for unexposed control embryos (89.5% +/- 2.3% and 57.2% +/- 3.2%, respectively). Embryos fused at 2.4 kV/cm applied as a single 30 microseconds pulse (69.7% +/- 5.7%) showed significantly lower cleavage (72.1% +/ 3.7%) and blastocyst rates (40.2% +/- 4.6%) compared to the unexposed control. PMID- 10862005 TI - Oviductal plasminogen activator inhibitor-1 (PAI-1): mRNA, protein, and hormonal regulation during the estrous cycle and early pregnancy in the pig. AB - Recent identification of plasminogen activator inhibitor-1 (PAI-1) in the pig oviduct has prompted an evaluation of its mRNA, protein synthesis, and hormonal regulation during the estrous cycle and early pregnancy, defined as time prior to and after maternal recognition of pregnancy. To examine PAI-1 protein synthesis, oviductal tissue was collected from European Large White and Chinese Meishan gilts on days 0, 2, and 5 of early pregnancy, divided into three functional segments, and cultured. Culture media was collected and de novo synthesized PAI-1 analyzed by 2D-SDS-PAGE, fluorography, and densitometry. To determine hormonal regulation of PAI-1 synthesis and secretion, four groups of ovariectomized (OVX) cross-bred gilts were each treated with one of four steroid regimens (corn oil, estrogen, progesterone, or estrogen + progesterone) and tissue collected for RNA or cultured. Steady-state mRNA levels of PAI-1 were evaluated throughout the estrous cycle in cross-bred gilts. To compare steady-state PAI-1 mRNA levels between cyclic and pregnant cross-bred gilts, tissue was collected on days 0, 2, and 12. Quantitative analysis of steady-state levels of PAI-1 mRNA were analyzed by dot-blot hybridization and densitometry. A greater (P < 0.01) synthesis and secretion of PAI-1 protein was found in the isthmus portion of the oviduct relative to either the ampulla or infundibulum regardless of day of pregnancy or breed. No difference could be detected for PAI-1 protein between breeds. The Large White had a greater (P < 0.05) secretion of PAI-1 on day 2 of early pregnancy relative to other days examined. Whole oviductal tissue from cross-bred gilts was found to have a significantly greater amount of PAI-1 mRNA on days 1 and 2 compared to other days examined, while the isthmus had significantly greater levels of mRNA on days 2 and 12. A significant effect of day and segment was detected for levels of PAI-1 mRNA from cyclic and early pregnant cross-bred gilts. PAI-1 mRNA was found to be significantly greater in the isthmus than other segments, regardless of day of the estrous cycle or pregnancy. An interaction was detected for estrogen and progesterone on PAI-1 mRNA (P < 0.05) and protein (P = 0.09). Estrogen was found to inhibit PAI-1 protein synthesis and also inhibited progesterone-mediated stimulation of PAI-1 mRNA. Our results demonstrate expression of PAI-1 mRNA and protein are highest on day 2 of early pregnancy, which is consistent with its proposed function of protecting the oocyte/embryo from enzymatic degradation and/or extracellular matrix remodeling of both oviduct and early cleavage-stage embryo. PMID- 10862006 TI - Tumor necrosis factor-alpha stimulates prostaglandin F2alpha secretion by bovine luteal cells via activation of mitogen-activated protein kinase and phospholipase A2 pathways. AB - It has been well demonstrated that tumor necrosis factor-alpha (TNFalpha) stimulates prostaglandin (PG) F2alpha secretion by bovine corpus luteum (CL) in vitro. The objective of the present study was to clarify the intracellular signaling pathway of TNFalpha to stimulate PGF2alpha production in cultured bovine luteal cells. Bovine luteal cells that were obtained from mid- (days 8-12 after ovulation) CL were incubated with TNFalpha (0.6 nM) and/or various compounds as follows: U-73122 (an inhibitor of phospholipase [PL] C), ACA (an inhibitor of PL-A2), H-89 (an inhibitor of protein kinase [PK] A), calphostin C (an inhibitor of PK-C), L-NAME/L-NORG (inhibitors of nitric oxide synthase), and PD98059 (an inhibitor of mitogen-activated protein kinase [MAPK] kinase). Although U-73122 (0. 1-10 microM), H-89 (0.1-10 microM), calphostin C (0.01-1 microM) and L-NAME/L-NORG (1-100 microM) did not affect TNFalpha-induced PGF2alpha secretion by the cultured cells, ACA (1-100 microM) and PD98059 (0.1 100 microM) inhibited TNFalpha-stimulated PGF2alpha secretion by the cells in a dose-dependent fashion (P < 0.05 or lower). These findings suggest that TNFalpha activates the MAPK and PL-A2 pathways in bovine luteal cells to stimulate PGF2alpha secretion. PMID- 10862007 TI - Diffusible highly glycosylated protein from Bufo arenarum egg-jelly coat: biological activity. AB - L-HGP is a highly glycosylated protein from Bufo arenarum egg-jelly coat that diffuses into the surrounding medium when the strings of oocytes are incubated in saline solutions. L-HGP was purified from egg water and the estimated percentage of L-HGP/total protein in egg water was estimated in 30%. In the present study we examine, by indirect immunofluorescence, the effect of L-HGP on acrosome status of homologous spermatozoa. A high percentage (77%) of sperm lost the acrosome when incubated in 10% Ringer solution buffered with 10 mM Tris-HCl, pH 7.6, during 60 min, a condition that resembles egg-jelly osmolarity. The addition of purified L-HGP to the incubation medium prevents acrosome breakdown. The acrosome integrity is maintained for at least 1 hr. This effect is specific for L-HGP at concentration ranging from 0.01 to 0.1 mg/ml since neither BSA nor fetuin seems to have similar activity at similar concentrations. The same effect was observed when spermatozoa were incubated in egg water. Preliminary results suggest that L HGP binds to B. arenarum spermatozoan membranes. PMID- 10862008 TI - Molecular cloning of rat sperm galactosyl receptor, a C-type lectin with in vitro egg binding activity. AB - Rat sperm galactosyl receptor is a member of the C-type animal lectin family showing preferential binding to N-acetylgalactosamine compared to galactose. Binding is mediated by a Ca(2+)-dependent carbohydrate-recognition domain (CRD) identical to that of the minor variant of rat hepatic lectin receptor 2/3 (RHL 2/3). The molecular organization of the genomic DNA, cDNA, and derived amino acid sequence of rat testis galactosyl receptor have been determined and in vitro fertilization studies were conducted to ascertain its role. We have determined that the rat testis galactosyl receptor gene generates two mRNA species: one species, designated liver-type, is identical to RHL-2/3; the other, designated testis-type, contains one unspliced intron (86 nt) which alters the reading frame and changes the amino acid sequence of the carboxyl terminus. As a result, the CRD (glutamine-proline-aspartic acid/QPD) and flanked Ca(2+)-binding amino acid sequences were not present in the testis-type protein. Northern and Southern blots demonstrated presence of transcripts with unspliced intron in rat sperm but not liver. Similarly, antibody, raised against a synthetic 12-amino acid peptide (p12) encoded by the unspliced intron, recognized in immunoblots a 54 kDa receptor protein in protein extracts from testis but not from liver. Immunofluorescence and immunogold electron microscopy studies demonstrated that both protein species localized on the plasma membrane surface of the head and tail of rat sperm. Furthermore, capacitated rat sperm preincubated with polyclonal antisera to RHL-2/3 or to the CRD of the liver-type galactosyl receptor showed a statistically significant decrease in the in vitro fertilization rate. We conclude that rat sperm galactosyl receptor may play a role in egg binding and that an undetermined molecular mechanism operates to generate two proteins with identical intracellular amino terminal domain but only one of them displays a CRD and associated Ca(2+)-binding sites at the carboxyl terminal extracellular domain. PMID- 10862009 TI - Analysis of the role of egg integrins in sperm-egg binding and fusion. AB - Sperm-egg fusion is believed to be mediated via specific molecular interactions. Integrin alpha6beta1 is a strong candidate for a sperm receptor on the egg plasma membrane. However, the ability of the egg integrin alpha6beta1 to interact with molecules on intact sperm has not yet been proven. In this report, possible involvement of integrin alpha6beta1 in sperm-egg interactions was examined by biochemical and immunocytochemical analyses. To identify egg molecules that specifically interact with sperm, we first incubated sperm with biotin-labeled egg surface proteins. Under this condition, solubilized proteins from eggs inhibited sperm-egg fusion. Western blot analysis under reducing conditions indicated that a major-labeled band of 135 kDa bound to sperm. An immunodepletion experiment using the anti-integrin alpha6 antibody GoH3 indicated that the 135 kDa egg surface molecule that bound to sperm was the integrin alpha6 subunit. To investigate the potential involvement of integrin alpha6beta1 in sperm-egg fusion, we next examined the localization of integrin alpha6 and beta1 subunits before and after fertilization by confocal laser microscopy. At an early stage of sperm-egg fusion, the integrin alpha6 and beta1 subunits were accumulated at the sperm binding site. The frequency of cluster formation was closely related to that of sperm-egg fusion, indicating that integrin receptors are accumulated by sperm destined for fusion. Taken together, these results strongly suggest that the integrin alpha6beta1 is involved in sperm-egg binding leading to fusion via direct association of the integrin alpha6 with sperm. PMID- 10862010 TI - Cloning, sequencing, and expression analysis of mouse glucosamine-6-phosphate deaminase (GNPDA/oscillin). AB - It was reported that a hamster protein, called "oscillin," with a sequence related to that of an Escherichia coli GNPDA triggered Ca(2+) oscillations in mammalian oocytes when introduced into their cytoplasm upon fertilization. Recently, it was shown that GNPDA/oscillin is ubiquitously expressed in rat tissues and that a recombinant hamster GNPDA/oscillin protein does not exhibit oscillin activity when injected into oocytes. In the mouse, the nature and role of such a GNPDA/oscillin is not known, but another candidate protein, tr-kit, has been proposed as a sperm factor causing oocyte activation. In order to clarify this issue, we have characterized the mouse homolog of hamster and human GNPDA/oscillin, and examined its expression along with that of tr-kit, in parallel. We report here the molecular cloning and sequencing of mouse GNPDA/oscillin, which shows over 96% identity with the hamster and human homologs. Using specific primers, we performed an RT-PCR analysis to determine the tissue distribution of mouse GNPDA/oscillin mRNA. Unlike tr-kit mRNA which is expressed solely in mouse testis, GNPDA/oscillin mRNA is detected in unfertilized oocytes and in all tissues examined including testis, heart, thymus, liver, ovary, uterus, kidney, spleen, and lung. The protein itself is also detected in all tissues examined by Western blots. Indirect immunofluorescence studies, using an antibody raised against hamster GNPDA, demonstrate that GNPDA is lost with the acrosome reaction of mouse spermatozoa, is localized in the equatorial and neck regions of the human spermatozoa and the post-acrosomal region of the hamster spermatozoa. Our results thus indicate that mouse GNPDA/oscillin, the homolog of hamster oscillin, unlike tr-kit, does not exhibit some of the required characteristics expected from a putative sperm-derived oocyte-activating factor. PMID- 10862011 TI - Effects of platelet activating factor on capacitation and acrosome reaction in mouse spermatozoa. AB - Platelet activating factor (PAF) plays an important role in mammalian reproduction. The aim of this study was to investigate the effects of PAF on capacitation and acrosome reaction of mouse spermatozoa by chlortetracycline (CTC) fluorescence assay and coomassie blue staining. The percentage of capacitated mouse spermatozoa was increased (P < 0.05) by incubation with 50 ng/ml PAF for 20-120 min. The peak response occurred between 80 to 100 min of exposure to PAF. In contrast, the effects of PAF on acrosome reaction may be not receptor-mediated since lyso-PAF had the same effects. Ionophore A23187 stimulated an increase in acrosome-reacted spermatozoa of PAF-treated spermatozoa, but not of lyso-PAF-treated ones. These results suggest that PAF mainly acts on sperm capacitation. PMID- 10862012 TI - The impact of comorbidity on the survival of patients with squamous cell carcinoma of the head and neck. AB - BACKGROUND: In North America, cigarette smoking and/or alcohol consumption not only cause head and neck cancer, they also cause many of the other diseases, illnesses, and conditions, also known as comorbidities, frequently found in our patients. Comorbidities can influence treatment decision making and treatment outcome. The aim of this study is to quantify the increased risk of comorbidity in our patients. METHOD: The survival of 655 consecutive patients with squamous cell carcinoma from a regional cancer center is analyzed. We compare the survival curves for all-cause death, death from cancer, and death from noncancer causes to the expected survival of age/sex-matched populations of Ontario residents, Canadian smokers, and Canadian nonsmokers. RESULTS: Of those patients who had not survived 5 years, 59% died of their index tumor, 23% would have been expected to die if they did not have head and neck cancer, and 18% died of the increased comorbidity associated with being a patient with head and neck cancer. DISCUSSION: Comorbidity, and specifically the increased comorbidity found in patients with head and neck cancer, is an important factor in overall survival. PMID- 10862013 TI - Anterior mandibular osteotomy for tumor extirpation: a critical evaluation. AB - BACKGROUND: Anterior mandibular osteotomy gives excellent access to the oropharynx for oncologic resections. Although rigid internal fixation has improved results in stability and bone healing, side effects and complications of the surgical procedure must be considered. METHODS: Between 1991 and 1997, 64 patients underwent a transmandibular approach for benign or malignant tumors of the head and neck. Sixty-one of them were included in a 6-month follow-up study. Final results and major and minor complications were recorded. RESULTS: Three (5%) cases of non-union and infection of the osteotomy site required plate removal, curettage, and reosteosynthesis. Occlusal disturbances and local periodontal disease occurred in 2 (3%) patients. Thirty-two (52%) patients complained of sensitivity disturbances, 18 (30%) of temporomandibular joint pain, and 44 (72%) of a limitation in opening the mouth. Six patients (10%) had a cosmetic complaint. CONCLUSION: Dental condition and periodontal care in the vicinity of the osteotomy are important factors in reducing the risk of major complications. Trismus, limitation of joint motion, and pain cannot be directly related to the technique; they depend on the extent of tumor resection and postoperative radiotherapy. PMID- 10862014 TI - Overexpression of p53 in squamous cell carcinoma of the tongue in young patients with no known risk factors is not associated with mutations in exons 5-9. AB - BACKGROUND: This study investigated the status of the p53 tumor suppressor gene in patients less than 40 years of age who had squamous cell carcinoma of the tongue develop with no known risk factors. METHODS: Histologic sections from 21 patients were prepared from formalin-fixed, paraffin-embedded tissue and were processed for standard immunohistochemistry for detection of the p53 protein. In addition, tumors were evaluated by single-strand conformation polymorphism and by DNA sequencing to identify potential mutations in the conserved exons (5-9) of the p53 gene. RESULTS: Eighty-one percent (17 of 21) of the patients overexpressed p53 by immunohistochemical analysis. However, none of these patients demonstrated mutations in exons 5-9 of the gene. CONCLUSIONS: These data suggest that the molecular mechanisms by which the young individuals with no risk factors had altered p53 function in oral squamous cell carcinoma may differ from those of the more typical population of individuals who have this malignancy develop. PMID- 10862015 TI - Epstein-Barr virus detection in neck metastases by in-situ hybridization in fine needle aspiration cytologic studies: an aid for differentiating the primary site. AB - BACKGROUND: Nasopharyngeal carcinoma (NPC) is strongly associated with Epstein Barr virus (EBV). The metastasis to cervical lymph nodes represents a frequent initial manifestation of NPC. The usefulness of EBV detection by polymerase chain reaction (PCR) in the diagnosis of occult NPC with cervical metastasis has been reported. Our previous study showed that EBER1 in-situ hybridization was somewhat more sensitive and specific than PCR in detecting EBV in the evaluation of specimens from a population at high risk for NPC. METHODS: Fine-needle aspiration cytologic specimens of neck masses from 30 patients were investigated, including 10 NPC primary tumors, 19 squamous cell carcinomas from other sites of the head and neck (9 oral cavity, 2 paranasal sinuses, 2 oropharynx, 3 larynx, and 3 hypopharynx), and 1 diffuse large-cell lymphoma. EBER1 in-situ hybridization was performed on direct smears made from aspirates. RESULTS: EBER1 signals were detected in all neck metastases from the nasopharynx but none of the specimens from other primary sites. CONCLUSIONS: This study suggests that EBER1 in-situ hybridization can be used as a supplemental tool for differential diagnosis whenever fine-needle aspiration cytologic examination is presented with a neck metastasis without knowing the primary site. PMID- 10862016 TI - Antisense oligonucleotides against protein kinase CK2-alpha inhibit growth of squamous cell carcinoma of the head and neck in vitro. AB - BACKGROUND: Human squamous cell carcinomas of the head and neck (SCCHN) overexpress the protein kinase CK2, and elevated CK2 activity correlates with aggressive tumor behavior and poor clinical outcome. We therefore investigated whether interference with CK2 expression would inhibit SCCHN cell growth in vitro. METHODS: We targeted the catalytic (alpha) subunit of CK2 using an antisense oligodeoxynucleotide (ODN) strategy. Human Ca9-22 cells derived from SCCHN were transfected with CK2-alpha sense, nonsense, or antisense ODN; CK2 activity was measured; and the effect on CK2 activity and on cell growth was determined. RESULTS: Transfection of Ca9-22 cells with antisense CK2-alpha ODN resulted in significantly decreased CK2 kinase activity associated with nuclear chromatin and in dose-dependent growth inhibition of Ca9-22 cells in vitro. CONCLUSIONS: Interference with the protein kinase CK2 signal in SCCHN cells may offer a novel anticancer strategy for this malignancy. PMID- 10862017 TI - Telomerase activity in head and neck tumors after introduction of wild-type p53, p21, p16, and E2F-1 genes by means of recombinant adenovirus. AB - BACKGROUND: Telomerase (reverse transcriptase) has been shown to play a role in the process of cellular immortalization. METHODS: Telomerase activity was determined in 11 head and neck squamous cell carcinoma (SCCHN) cell lines. The effects of wild-type p16, p21, E2F-1, and p53 genes on telomerase activity were examined by introducing the wild-type genes into two SCCHN cell lines by means of a recombinant adenovirus. RESULTS: We found elevated telomerase activity in 10 of the 11 SCCHN cell lines tested. When we infected Tu-138 and Tu-167 cell lines with wild-type p16, p21, E2F-1, and p53 genes, we found that p16 had little effect on telomerase activity. Both E2F-1 and p53 were known to induce apoptosis in SCCHN cell lines. Significantly reduced telomerase activity by p53 in both cell lines and E2F-1 in Tu-167 cells was in agreement with suppression of cell growth. Overexpression of p21 also exhibited reduction in telomerase activity. CONCLUSIONS: We conclude from this study that overexpression of E2F-1 and p53 can reverse telomerase activity in SCCHN cell lines and that telomerase activity may be involved in cancer cell immortalization. PMID- 10862018 TI - MRI-guided needle localization in the head and neck using contemporaneous imaging in an open configuration system. AB - BACKGROUND: MRI-guided procedures have previously been limited by technical difficulties, including the need for MRI-compatible instruments, slow image acquisition time, and the closed nature of conventional MRI scanners. The development of open configuration MRI systems with in-room, contemporaneous imaging has greatly increased the potential for MRI-guided interventional procedures. We evaluate our clinical experience applying this technology to the head and neck. METHODS: An open design 0.2T magnet combined with an in-room monitor was used for 24 MRI-guided needle localization procedures in the head and neck. Success of the procedures was based on the ability to accurately position the instrument in the target region to allow biopsy or treatment. RESULTS: In all 24 cases placement of the instrument within the target tissue was successful. CONCLUSION: MRI-guided needle-localization procedures in an open design magnet with in-room, contemporaneous image monitoring offer advantages over previous conventional interventional MRI systems by allowing interactive guidance with near real-time imaging feedback. As a result, procedure time is reduced and accuracy of instrument positioning is increased. PMID- 10862019 TI - Papillary squamous cell carcinomas of the upper aerodigestive tract: a clinicopathologic and molecular study. AB - BACKGROUND: The limited studies and the small number of published cases of papillary squamous cell carcinoma have precluded accurate assessment of the biologic characteristics of this lesion. METHODS: Thirty-eight of the carcinomas were studied. In-situ hybridization and polymerase chain reaction were performed to detect human papilloma virus (HPV) and p53 expression. RESULTS: HPV was found in 4 of 14 assessable carcinomas by in-situ hybridization and in 5 of 14 by polymerase chain reaction. The most frequently identified HPVs were HPVs in 6/11 and 16/18 patients. In general, a reciprocal relationship was found between p53 and HPV prevalence. The most lethal site for this tumor was the sinonasal tract, whereas patients with papillary squamous cell carcinomas of the larynx had the best outlook. Eleven of 25 (44%) assessable patients died of disease (mean time interval, 2 year). CONCLUSIONS: Papillary squamous cell carcinoma of the upper aerodigestive tract is a distinct variant of squamous cell carcinoma. As such and because of its putative association with HPV, papillary squamous cell carcinoma could be an informative model for defining how viral oncogenes cooperate with other factors in genomic instability, carcinogenesis, and tumor development. PMID- 10862020 TI - Staging computed tomography of the thorax for nasopharyngeal carcinoma. AB - BACKGROUND: To assess the role of staging CT of the thorax in advanced nodal stage nasopharyngeal carcinoma and to examine the hypothesis that contiguous spread of nodal metastases from the supraclavicular region to the upper mediastinal region occurs in this cancer. METHODS: Forty-four patients with newly diagnosed nasopharyngeal carcinoma with neck node metastases to the supraclavicular region (ie, AJCC N3b stage) underwent contrast-enhanced CT (CECT) thorax for staging. CT findings and clinical outcome were analyzed. RESULTS: No patient was found to have intrathoracic metastasis, although 1 had hepatic metastases on CECT of the thorax, resulting in upstaging in 1 of 44 (2%) of patients. With a median follow-up time of 21 months, 3 patients had lung metastases and 2 had axillary nodal metastases develop without evidence of upper mediastinal nodal metastases. CONCLUSION: Staging CECT of the thorax has a very low yield in nasopharyngeal carcinoma, even in advanced nodal disease. The hypothesis that contiguous spread of nodal metastases from the supraclavicular region to the upper mediastinum is not substantiated, and no evidence suggests that radiation therapy for N3b-stage disease needs to encompass the upper mediastinum. PMID- 10862021 TI - Metalloproteinase-9 immunoexpression and angiogenesis in thyroid follicular neoplasms: relation to clinical and histopathologic features. AB - BACKGROUND: Thyroid follicular neoplasms (adenoma and carcinoma) may pose considerable difficulties to the differential diagnosis. Because such a distinction is not possible at fine-needle aspiration, surgery is often necessary. Clinical information such as age, sex, and node size is important in case of suspected carcinoma. Follicular carcinoma is characterized by capsular invasion, vascular invasion, and metastatic dissemination mainly by the hematogenic pathway. This invasion depends on collagen degradation in capsule and in subendothelial basement membrane. Collagen degradation has been widely researched in the angiogenesis process and in the hematogenic dissemination mechanism. In this study, we performed clinical and histopathologic assessment of 74 follicular neoplasms, as well as immunohistochemical reactions for CD-34 protein to estimate angiogenesis and for metalloproteinase-9, an enzyme that degrades type IV collagen. METHODS: The research was carried out retrospectively in 74 patients who had surgery and were followed up at HC-FMUSP and IBCC. Clinical, histologic, and immunohistochemical variables were compared among the groups of follicular neoplasms and a control group of 36 patients with colloid goiter. RESULTS: No significant statistical difference was found between patients with follicular adenoma and thyroid follicular carcinoma concerning sex (p =.092), age (p =.098), thyroid node size (p =.426), vascularization (p =.388), and immunostaining intensity for metalloproteinase-9 (p =.055). The proportion of immunoreactive cells for metalloproteinase-9 in follicular carcinoma cases was higher than that observed in follicular adenoma cases (p <.001). Patients in more advanced stages of carcinoma were more than 45 years old (p =.006), presented extensive invasion (p <.001), had less vascularization (p =.046), and a had higher proportion of immunoreactive cells for metalloproteinase-9 (p <.001). CONCLUSIONS: The proportion of immunoreactive cells for metalloproteinase-9 in follicular carcinoma was higher than that observed in follicular adenoma, with a significant statistical difference (p <.001). This method must be developed to apply in material obtained by fine-needle aspiration to differentiate follicular adenoma from carcinoma. PMID- 10862022 TI - Cervical node metastases in laryngeal and hypopharyngeal cancer: a prospective analysis of prevalence and distribution. AB - BACKGROUND: We have prospectively analyzed the prevalence and distribution of histologic cervical node metastases in laryngeal and hypopharyngeal squamous carcinoma to determine the most appropriate form of neck dissection. METHODS: We have examined specimens from 100 consecutive patients in whom neck dissection was part of the primary treatment of laryngeal and hypopharyngeal carcinoma. Fifty eight patients were treated by unilateral or bilateral selective dissection of levels I to IV +/- VI for N0 disease and 42 by comprehensive dissection for N+ disease. Assessment was by separation of the specimens into node levels at the time of surgery and embedding all the resected material for histologic analysis. RESULTS: Nodal metastases were found in 36% of ipsilateral and 27% of contralateral dissections in the N0 cases. The corresponding prevalences in N+ cases were 90% and 37%, respectively. All metastases in N0 and N1 disease were confined to levels II, III, IV, and VI. Metastases to levels I and V were infrequent even in N+ disease. CONCLUSIONS: Our results support the use of elective dissection of node levels II to IV for N0 laryngeal and hypopharyngeal carcinoma. We suggest the inclusion of level VI nodes for tumors invading the subglottis, pyriform fossa apex, and postcricoid region. The prevalence of bilateral metastases is great enough in midline or bilateral tumors to justify bilateral selective dissection. It is possible that selective neck dissection is also adequate for small palpable metastases, but greater numbers are required to confirm this. PMID- 10862023 TI - Treatment of the contralateral negative neck in supraglottic cancer patients with unilateral node metastases (N1-3). AB - BACKGROUND: Elective treatment of the contralateral N0 neck in supraglottic cancer patients with unilateral metastases is controversial. METHODS: We reviewed 127 N1-3 cases with contralateral negative necks to compare elective contralateral dissection (ED: 24 cases) with a contralateral wait-and-see policy (WS: 103 cases) and subsequent delayed therapy (SDT: 40 cases) when contralateral disease became evident. Prognostic factors were studied to identify the risk of contralateral disease. RESULTS: Nine of 24 (37.5%) ED patients had occult contralateral metastases, and 40 of 103 (38.8%) WS patients had a delayed contralateral failure. Supraglottic cancers involving or extending up to the midline had a higher risk of contralateral metastases compared with well lateralized tumors (p =.049). The risk of contralateral neck disease was more influenced by tumor site and stage than by histopathologic characteristics of ipsilateral metastases. WS patients with contralateral neck relapse showed a higher risk of distant metastases and of level I and V neck involvement than ED cases with no difference in terms of survival. CONCLUSIONS: The risk of contralateral occult neck involvement in supraglottic laryngeal cancers with unilateral metastases is high (about 40%), particularly for more advanced lesions extending to or involving the midline larynx; thus, a bilateral neck treatment in such cases is recommended. PMID- 10862024 TI - Dysphagia in treated nasopharyngeal cancer. AB - OBJECTIVES: To investigate the prevalence of long-term dysphagia in patients treated for nasopharyngeal carcinoma (NPC) by radiotherapy. Study Design Questionnaire-based assessment, clinical examination, and videofluoroscopic assessment of 50 patients, ages 26 to 75 years (average, 49 years), treated for NPC 12 to 119 months (average, 56 months) previously with no evidence of disease recurrence. METHODS: Administered questionnaire assessment of patients eating and swallowing. Clinical examination by a single experienced clinician. Videofluoroscopy was used to record swallowing of solid, paste, and liquid bolus. Pharyngeal transit time (PTT) was recorded, and the video recordings were assessed by two experienced observers for abnormalities. RESULTS: Fifty patients completed the questionnaire and were examined. Seventy-six percent reported dysphagia, 97% had xerostomia, and 78% had no gag reflex. Forty-nine patients underwent videofluoroscopy. Abnormal pharyngeal contraction was observed on videofluoroscopy in 93% of the subjects. Silent aspiration was observed in 22% of the patients. PTT was prolonged from a normal of 1 second to 1.9 seconds for solid, 1.7 seconds for paste, and 1.3 seconds for liquid consistencies. CONCLUSIONS: Subjective and objective swallowing abnormalities are common after radiotherapy for NPC. The implications of this finding and possible causes are discussed. PMID- 10862025 TI - Pretreatment factors predicting quality of life after treatment for head and neck cancer. AB - BACKGROUND: Quality of life (QOL) has become an important issue in head and neck cancer. Explanation of factors predicting QOL after treatment has important implications for patient management. METHODS: In this prospective study we analyzed which pretreatment factors predicted QOL after surgery and/or radiotherapy with curative intent in a cohort of 153 patients with cancer of the oral cavity, oropharynx, hypopharynx, or larynx. The patients completed the EORTC Core Questionnaire, the EORTC Head and Neck Cancer module, and the Center for Epidemiologic Studies Depression scale before treatment and 6 and 12 months later. The influence of gender, age, performance status, and depressive symptoms at baseline, site, stage, and treatment on QOL (and its dimensions) and depressive symptoms after 6 and 12 months was studied, using linear regression analysis. RESULTS: A high level of depressive symptoms and a low performance status at baseline and combination treatment were significant predictors of increased severity of symptoms and poor functioning after treatment. Treatment was a predictor of head and neck symptoms, whereas performance status and depressive symptoms were predictors of general symptoms and functioning. Gender and age had little predictive value. CONCLUSIONS: Patients with depressive symptoms or a low performance status who receive combination treatment for cancer of the head and neck are at risk for physical and psychologic morbidity after treatment. Special attention should be given to these patients in rehabilitation programs. PMID- 10862026 TI - National cancer database report on chondrosarcoma of the head and neck. AB - BACKGROUND: Management of chondrosarcoma of the head and neck is largely based on single-institution reports with small numbers accrued over several decades. METHODS: The American College of Surgeons' National Cancer Data Base included 400 cases of chondrosarcoma of the head and neck diagnosed between 1985 and 1995. Chi square analyses of selected contingency tables and Wilcoxon regression analyses of selected survival stratifications were performed. RESULTS: Histologic types included conventional (80.8%), myxoid (10.5%), and mesenchymal (8.8%). The mesenchymal and myxoid subtypes were rare among white patients (17.1%) and more common among African-American (31.8%) and Hispanic patients (44.9%). Treatment was most commonly surgery alone (59.5%) and surgery with irradiation (21.0%). Disease-specific survival was 87.2% at 5 years and 70.6% at 10 years. Worse 5 year survival was associated with higher grade (67.3%), regional or distant spread (71.0%), and the myxoid (45.0%) or mesenchymal (53.2%) subtypes. CONCLUSIONS: Chondrosarcoma of the head and neck encompasses a variety of lesions that differ substantially by demographic and tumor characteristics. Individual tumors can be classified further according to site of origin, histologic subtype, and tumor grade, which can be used to predict biologic behavior and prognosis. PMID- 10862027 TI - Supraglottic cancer. PMID- 10862028 TI - Subglottic stenosis secondary to extramedullary hematopoiesis in a patient with postpolycythemia myeloid metaplasia. AB - BACKGROUND: Extramedullary hematopoiesis (EMH) is known to occur in myeloproliferative disorders and hemoglobinopathies and is usually seen in the spleen and liver. METHODS: We report the first case of EMH causing subglottic stenosis in a woman with postpolycythemia myeloid metaplasia (PPMM). A tracheotomy was performed to maintain the airway and local radiotherapy was given. RESULTS: Two months after the radiotherapy was completed laryngoscopy showed an unobstructed airway with no evidence of disease, and the patient was successfully decanulated. Magnetic resonance imaging 8 months after radiotherapy confirmed the absence of local disease. CONCLUSION: Consideration should be given to EMH as a possible cause of airway obstruction in the differential diagnosis of a patient with a history of PPMM. PMID- 10862029 TI - Genetics of uterine leiomyomata. AB - Leiomyomata represent the most common gynecologic tumors in women of reproductive age and are the primary indication for hysterectomy in the United States. Cytogenetic and genetic studies have in recent years advanced our understanding of the etiology of these tumors. Cytogenetic aberrations involving chromosomes 6, 7, 12, and 14 constitute the major chromosomal abnormalities seen in leiomyomata and have led to the discovery that disruptions or dysregulations of HMGIC and HMGIY contribute to the development of these tumors. Based on the finding of a variety of other consistent chromosomal aberrations detected in these tumors, other genes with fundamental roles in the pathobiology of uterine leiomyomata await identification. Genes Chromosomes Cancer 28:235-245, 2000. PMID- 10862030 TI - Involvement of the X chromosome in non-Hodgkin lymphoma. AB - Gain of an X chromosome is observed as a secondary, acquired karyotypic alteration in a significant proportion of malignant lymphomas. To determine the potential involvement of X-linked genes in neoplastic development, we have analyzed the inactivation status of the supernumerary X chromosome in lymphomas in both male and female patients. In males, neither methylation of FMR1 nor expression of XIST was detected, demonstrating that the duplicated chromosome was not subject to inactivation. In females, both expressed polymorphisms and polymorphisms associated with methylation differences between the active and inactive X chromosome were analyzed to determine whether the duplicated chromosome was active or inactive. To facilitate this analysis, allele-specific PCR primers were designed for detection of previously described polymorphisms in the IDSX and G6PD genes. The female lymphomas were shown to be clonal in origin, and duplication of either the active (5 cases) or inactive (4 cases) X chromosome was observed. Correlations between clinical status and the inactivation status of the X chromosome involved in the duplication were not observed in our relatively small sample, although 4/4 informative cases with a t(14;18) showed duplication of the active X chromosome. In the course of these studies, we detected hypermethylation of the androgen receptor (AR) locus in an extremely high proportion of both male (7/9) and female (9/10) samples. These results are discussed with respect to whether sex chromosome aneuploidies in tumors are involved in, or simply the result of, the neoplastic process. Genes Chromosomes Cancer 28:246-257, 2000. PMID- 10862032 TI - Mapping of a minimal deleted region in human hepatocellular carcinoma to 1p36.13 p36.23 and mutational analysis of the RIZ (PRDM2) gene localized to the region. AB - Human chromosome band 1p36 commonly undergoes loss of heterozygosity (LOH) in hepatocellular carcinoma (HCC) but the minimal deleted region remains to be mapped. This chromosomal region contains a candidate HCC suppressor gene, RIZ (PRDM2), that is a member of the PR (PRDI-BF1-RIZ homology)-domain-containing zinc finger gene family. One characteristic of this family is the unusual yin yang involvement in human cancers. The PR-domain-containing RIZ1 product of the RIZ locus, in contrast to the PR-domain-minus product RIZ2, is commonly lost or underexpressed in HCC. Furthermore, RIZ1 can induce cell cycle arrest, apoptosis, or both and suppress HCC tumorigenicity in nude mice. To help identify the putative HCC locus on 1p36 and to evaluate a genetic role of RIZ in HCC, we studied 97 HCC cases and mapped a minimal deleted region in HCC to 1p36.13-p36. 23 between markers D1S434 and D1S436. Notably, RIZ mapped within this region and was found to undergo LOH in 37% (25/67) of HCC cases. Single-strand conformation polymorphism (SSCP) analysis, however, did not show mutations in the PR-domain region of RIZ1 in 49 cases of HCC examined. Our data suggest that the RIZ locus is a target of frequent deletion in HCC, but that the more common way of RIZ inactivation in HCC may not involve mutations that alter peptide sequences. Genes Chromosomes Cancer 28:269-275, 2000. PMID- 10862031 TI - Genetic analysis of the APAF1 gene in male germ cell tumors. AB - Cytogenetic and molecular analyses have shown that the chromosome band 12q22 is recurrently deleted in male germ cell tumors (GCTs), indicating the presence of a candidate tumor suppressor gene (TSG) in this region. To identify the TSG, we mapped the APAF1 gene, a proapoptotic mammalian homologue of ced-4, to chromosomal band 12q22, that suggested that this might be the candidate deleted gene in GCTs. We further localized the gene between the polymorphic markers D12S1671 and D12S1082 at 12q22 to determine the role of APAF1 in the pathogenesis of GCT, and we characterized its normal genomic structure and analyzed its alterations in GCTs. The APAF1 gene comprises 27 exons, with the coding region spanning 26. The region containing APAF1 was found to be deleted in GCT by fluorescence in situ hybridization analysis, but without evidence of coding sequence alterations. RT-PCR and Western blot analysis showed APAF1 gene expression at detectable levels in all GCT cell lines analyzed. An aberrant-sized APAF1 protein was seen in one cell line. This and 2 other cell lines carrying APAF1 deletions also exhibited defects in dATP-mediated caspase-3 activation. Caspase-3 activity was effectively restored by addition of recombinant caspase-9 and APAF1 proteins, and to a lesser extent by caspase-9 alone, but not by APAF1 alone. These data do not support a TSG role for APAF1, but defects in other components of the apoptotic pathway that may be related to 12q22 deletion cannot be ruled out. Genes Chromosomes Cancer 28:258-268, 2000. PMID- 10862033 TI - Molecular analysis of chromosome arm 17q gain in neuroblastoma. AB - Complete or partial gain of the long arm of chromosome 17 (17q) has been shown recently by molecular cytogenetic techniques to be the most frequent chromosomal change in neuroblastoma and to be associated with adverse prognosis. Few reports, however, have focused on the precise mapping of the commonly overrepresented region. We have investigated 17q gain by the analysis of allelic imbalances at microsatellite loci dispersed along chromosome 17 in a series of 69 neuroblastomas. Allelic imbalances for at least two consecutive loci were observed in 39/59 informative cases, that is in agreement with previously reported frequencies of 17q gain. In a subset of the cases, comparative genomic hybridization analysis established the relationship between these allelic imbalances and the gain of 17q material. A partial 17q gain was observed in 9 cases, delineating a common region of 17q gain between the marker D17S787 (75 cM, 360 cR) and the telomere. In most cases, molecular results were suggestive of partial tri- or tetrasomy, whereas in 4 cases a higher copy number was documented. Our results also confirm that the presence of additional 17q material is closely associated with 1p36 deletion, MYCN amplification, and diploid or tetraploid chromosomal content. Genes Chromosomes Cancer 28:276-284, 2000. PMID- 10862034 TI - Localization of a novel tumor metastasis suppressor region on the short arm of human chromosome 2. AB - Much of the lethality of malignant neoplasms is attributable directly to their ability to develop secondary growths in organs at a distance from the primary tumor mass, whereas few patients die from their primary neoplasm. Little is known about the molecular mechanism of tumor metastasis, however, which is controlled by a variety of positive and negative factors. In the search for metastasis suppressor genes, we have used the microcell-mediated chromosome transfer method and a rat prostate tumor model in SCID mice. When human chromosome 2 was introduced into the highly metastatic rat prostatic tumor cell, AT6.1, the metastatic ability of this cell was significantly (>99%) decreased in animals. An STS-based PCR analysis for 8 hybrid clones indicates that the suppressor activity is located in the p25-22 region of the chromosome. Furthermore, the AT6.1 cell with human chromosome 2 showed a reduced ability to invade Matrigel, suggesting that the suppressor activity is involved in the step of tumor invasion during the progression of prostate cancer. We have also examined the status of the suppressor region on chromosome 2 in human prostate cancer specimens and found that this region was often lost in high-grade tumors. These results suggest that the putative suppressor gene on chromosome 2 is functionally involved in the progression of human prostate cancer. Genes Chromosomes Cancer 28:285-293, 2000. PMID- 10862035 TI - Pancreatic acinar carcinoma shows a distinct pattern of chromosomal imbalances by comparative genomic hybridization. AB - Pancreatic acinar cell carcinoma (PAC) is a rare pancreatic tumor for which no information about chromosomal anomalies is available. We examined six primary PACs by comparative genomic hybridization (CGH). All cases showed chromosomal changes. A total of 106 gains and 48 losses was detected. Consensus regions of gain were identified on chromosomes 1, 12, and X: 1q21 in four cases, 1q42 in three cases, 12p11.2 in four cases, and Xq12-21 in three cases. Recurrent losses were found at 16p13.2-p13.1 in three cases and at 16q23 in three cases. To verify these chromosomal imbalances, microsatellite analysis of matched normal and tumor DNA was performed using PCR-amplified markers for chromosomes 1, 12, and 16 in the regions showing nonrandom gains or losses. This analysis showed allelic imbalances in tumor DNA consistent with the CGH profiles. Our CGH study suggests that PAC shows a characteristic pattern of chromosomal alterations, involving gain at 1q, 12p, and Xq and loss of sequences at 16p and 16q. This pattern appears unique among solid tumors and is markedly different from that detected in pancreatic ductal carcinomas by the same technique. This suggests that PAC tumorigenesis involves different molecular pathways than those involved in the more common pancreatic ductal tumors. Genes Chromosomes Cancer 28:294-299, 2000. PMID- 10862036 TI - An Alu-mediated 7.1 kb deletion of BRCA1 exons 8 and 9 in breast and ovarian cancer families that results in alternative splicing of exon 10. AB - Constitutive large deletions and duplications of BRCA1 resulting from Alu mediated recombination account for a significant proportion of disease-causing mutations in breast and/or ovarian cancer families. Using Southern blot analysis and a protein truncation test (PTT), we have identified a 7.1 kb germline deletion in two families with breast and ovarian cancer. This deletion, which includes exons 8 and 9 and leads to a frameshift at the mRNA level, appears to result from homologous recombination between closely related Alu repeats, one in intron 7 and one in intron 9. In addition to the transcript without exons 8 and 9, analysis of RNA by protein truncation test from individuals with the deletion also identified the presence of alternative splicing of exon 10 from the mutant allele, which results in a transcript that lacks exons 8, 9, and 10. Of interest is that the two American families who carry this deletion are of northern European ancestry and share a common haplotype, suggesting that this deletion may represent a founder mutation. Genes Chromosomes Cancer 28:300-307, 2000. PMID- 10862037 TI - Chromosomal alterations in 15 breast cancer cell lines by comparative genomic hybridization and spectral karyotyping. AB - Breast cancer cell lines have been widely used as models in functional and therapeutical studies, but their chromosomal alterations are not well known. We characterized the chromosomal aberrations in 15 commonly used human breast carcinoma cell lines (BT-474, BT-549, CAMA-1, DU4475, MCF7, MDA-MB-134, MDA-MB 157, MDA-MB-361, MDA-MB-436, MPE600, SK-BR-3, T-47D, UACC-812, UACC-893, and ZR 75-1) by comparative genomic hybridization (CGH) and spectral karyotyping (SKY). By CGH the most frequent gains were detected at 1q, 8q, 20q, 7, 11q13, 17q, 9q, and 16p, whereas losses were most common at 8p, 11q14-qter, 18q, and Xq. SKY revealed a multitude of structural and numerical chromosomal aberrations. Simple translocations, typically consisting of entire translocated chromosome arms, were the most common structural aberrations. Complex marker chromosomes included material from up to seven different chromosomes. Evidence for a cytogenetic aberration not previously described in breast cancer, the isoderivative chromosome, was found in two cell lines. Translocations t(8;11), t(12;16), t(1;16), and t(15;17) were frequently found, although the resulting derivative chromosomes and their breakpoints were strikingly dissimilar. The chromosomes most frequently involved in translocations were 8, 1, 17, 16, and 20. An excellent correlation was found between the number of translocation events found by SKY in the individual cell lines, and the copy number gains and losses detected by CGH, indicating that the majority of translocations are unbalanced. Genes Chromosomes Cancer 28:308-317, 2000. PMID- 10862038 TI - Twenty-four-color spectral karyotyping reveals chromosome aberrations in cytogenetically normal acute myeloid leukemia. AB - Multicolor spectral karyotyping allows simultaneous visualization of all human chromosomes and screening for chromosomal rearrangements without a priori knowledge of any abnormalities involved. Based on this potentially increased sensitivity, we investigated, in a preliminary manner, whether spectral karyotyping could detect cytogenetic aberrations in karyotypically normal leukemia. The test population was comprised of 28 cryopreserved, cytogenetically normal acute myeloid leukemia (AML) samples from patients registered to a randomized trial for previously untreated AML (SWOG 9031). Two normal and 12 samples with known cytogenetic aberrations were used to validate and establish the diagnostic accuracy of the spectral karyotyping assay and instrumentation in a clinical setting. Enumeration and region-specific DNA fluorescence in situ hybridization (FISH) probes verified discrepant results. In the validation data set, spectral karyotyping refined complex karyotypic rearrangements in six cases and defined the chromosomal origin of a "jumping" homogeneously staining region; however, the technology was less sensitive in the detection of subtelomeric rearrangements and double minute chromosomes. In the test population, spectral karyotyping identified previously undetected cytogenetic aberrations in two cases (7%) of karyotypically normal AML: a cryptic 11q23 translocation in 20/20 cells and a minor monosomy 7 clone in 3/21 cells (FISH, 10.5%). Both of these abnormalities are considered to confer a poor prognosis when based on classical cytogenetic prognostic criteria. As an adjunct to classical cytogenetics and standard FISH analyses, the additive resolution of spectral karyotyping, in particular, with chromosome paints spiked with subtelomeric and/or locus-specific probes, may allow significant gains to be made in diagnostic accuracy and recognition of genotype/phenotype prognostic relationships, and in defining underlying biologic mechanisms in cancer. Genes Chromosomes Cancer 28:318-328, 2000. PMID- 10862039 TI - Chromosomal regions involved in the pathogenesis of osteosarcomas. AB - The comparative genomic hybridization technique (CGH) was used to identify common chromosomal imbalances in osteosarcomas (OS), which frequently display complex karyotypic changes. We analyzed 13 high-grade primary tumors, 5 corresponding cell lines, 2 primary tumors grade 2, and 1 recurrent tumor from a total of 16 patients. Some of the CGH results have been verified by fluorescence in situ hybridization (FISH) studies. Gains of chromosomal material were more frequent than losses. Most common gains were observed at 8q (11 cases), 4q (9 cases), 7q (8 cases), 5p (7 cases), and 1p (8 cases). The smallest regions of overlap have been narrowed down to 8q23 (10 cases), 4q12-13 (8 cases), 5p13-14 (7 cases), 7q31 32 (7 cases), 8q21 (7 cases), and 4q28-31 (5 cases). These data demonstrate that a number of chromosomal regions and even two distinct loci on 4q and 8q are involved in the pathogenesis of OS, with gain of 4q12-13 chromosomal material representing a newly identified locus. Seven of 16 cases displayed, besides gain of 8q23 sequences, gain of MYC copies in CGH and FISH. Previous CGH reports confined gain of 8q material to 8cen-q13, 8q21.3-8q22, and 8q23-qter, whereas our data suggest that the loci 8q21 and 8q23-24 are affected in the development of OS. In contrast to recent reports, copy number increases at 8q and 1q21 did not have an unfavorable impact on prognosis in the present series. Genes Chromosomes Cancer 28:329-336, 2000. PMID- 10862040 TI - Loss of heterozygosity analysis of 13q and 14q in human malignant mesothelioma. AB - Cytogenetic investigations of malignant mesothelioma (MM) have revealed frequent losses in chromosomes 13 and 14, suggesting that inactivation of tumor suppressor genes (TSGs) residing in these chromosomes may contribute to mesothelial cell tumorigenesis. To define the shortest region of overlap (SRO) of deletions from these chromosomes, we performed loss of heterozygosity (LOH) analyses on 30 MMs using 25 microsatellite markers in 13q and 21 markers in 14q. Twenty of the 30 MMs (67%) showed allelic loss of at least one marker in 13q. The SRO of deletions was delineated as an approximately 7 centiMorgan region, flanked by markers D13S1253 and D13S291, located at 13q13.3-14.2. Thirteen of the 30 MMs (43%) displayed allelic losses from 14q, with at least three distinct regions of LOH located at segments q11.2-13.2, q22.3-24.3, and q32. 12. These data highlight a single region of chromosomal loss in 13q in many MMs, implicating the involvement of a TSG that is critical to the pathogenesis of this malignancy. In contrast, the lower incidence and diffuse pattern of allelic losses in 14q suggest that several TSGs in this chromosome arm may contribute to tumorigenic progression in a subset of MMs. Genes Chromosomes Cancer 28:337-341, 2000. PMID- 10862041 TI - Frequent allelic imbalance suggests involvement of a tumor suppressor gene at 1p36 in the pathogenesis of human lung cancers. AB - The short arm of chromosome 1 is among the most frequently affected regions in various types of common adult cancers as well as in neuroblastoma. In a previous study of ours, frequent allelic imbalance at the TP73 locus at 1p36 was noted in lung cancer despite the absence of TP73 mutations. This suggested the possible existence of an as yet unidentified tumor suppressor gene on 1p. Our initial attempt using the candidate gene approach did not yield any somatic mutations in the 14-3-3sigma gene (official gene symbol, SFN), a mediator of G2 arrest by TP53. Detailed deletion mapping of the telomeric region of 1p was thus carried out as an initial step toward positional cloning. We used seven polymorphic markers in addition to TP73 to examine 61 primary lung cancers. Allelic imbalance at one or more loci of 1p36 was observed in 30 of the 61 cases, whereas D1S508 at 1p36.2 exhibited the highest frequency (45%) of allelic imbalance among the 1p36 markers examined. In contrast, two proximal markers at 1p32-34 showed significantly less frequent (11-14%) allelic imbalance. Consequently, the present study identified the shortest region of overlap between D1S507 and TP73, which included the most frequently affected marker, D1S508. In addition, several cases exhibited allelic imbalance confined to a subtelomeric region distal to D1S2845 at 1p36.3. The present findings warrant future studies to identify the putative tumor suppressor gene(s) at 1p36 to gain a better understanding of the molecular pathogenesis of lung cancer. Genes Chromosomes Cancer 28:342-346, 2000. PMID- 10862042 TI - Locus-specific multifluor FISH analysis allows physical characterization of complex chromosome abnormalities in neoplasia. AB - Novel techniques in molecular cytogenetics have radically improved the ability to characterize genetic changes in neoplastic cells. In parallel, a rapid development in high-throughput genomics has resulted in detailed physical maps of the human genome. Combining these two fields, we have developed a method for the simultaneous visualization of several physically defined segments along a chromosome. Seven YAC clones and one subtelomeric cosmid clone from chromosome 12 were labeled with unique combinations of four fluors and hybridized to metaphase chromosomes from neoplastic cells. In a uterine leiomyoma and a myxoid liposarcoma with translocations 12;14 and 12;16, the breakpoints in chromosome 12 could be localized to the HMGIC and CHOP regions, respectively. In the other tumors, more complex aberrations were visualized, including two inversions in 12q with a common breakpoint between MDM2 and D12S332 in a pleomorphic adenoma, amplification of MDM2 and CDK4 in ring chromosomes from a malignant fibrous histiocytoma, and amplification of KRAS2 together with other unbalanced rearrangements in two pancreatic adenocarcinomas. Combinatorially labeled single copy probes may thus simultaneously provide physical localization of breakpoints and an overview of complex structural rearrangements. Genes Chromosomes Cancer 28:347-352, 2000. PMID- 10862043 TI - Risk of false positive results in comparative genomic hybridization. AB - Comparative genomic hybridization, a widely used method for screening for genomic imbalances, suffers from a lack of standardized evaluation. In order to compare a recently proposed data-driven procedure with the commonly used fixed cutoff values, we tested 257 events by both procedures as well as by fluorescence in situ hybridization using selected probes. With the data-driven procedure, a much higher fraction (42/218 vs. 8/218) of false positive results was obtained, whereas a higher sensitivity with respect to the detection of imbalances (30/39 vs. 19/39) was reached. Based on the significantly higher positive likelihood ratios and the positive predictive value, we strongly recommend the use of fixed diagnostic thresholds, because the alternative procedure generates an unacceptably high portion of incorrectly scored chromosomal imbalances. Genes Chromosomes Cancer 28:353-357, 2000. PMID- 10862044 TI - Two identical triplet sisters carrying a germline BRCA1 gene mutation acquire very similar breast cancer somatic mutations at multiple other sites throughout the genome. AB - Monozygotic twins, each of whom has breast cancer, offer a natural study population for gene-environmental interactions as causation of cancer, because they are genetically identical. If heritable factors play a large role in the origin of a neoplasm, disease concordance should be significant in monozygotic twins. Two monozygotic triplet sisters carrying a germline BRCA1 gene mutation (5382insC) who both developed breast cancer at early ages were studied for loss of heterozygosity (LOH) in their microdissected, paraffin-embedded tumors along with control blood and stromal breast tissue at 19 chromosomal arms using 161 microsatellite markers. Microdissected areas of normal lobular and ductal epithelium and ductal in situ carcinoma were also studied for LOH using a subset of microsatellite markers. The mother's DNA (extracted from peripheral blood lymphocytes) was analyzed to determine the parental allele under LOH in each case. Both tumors demonstrated similar histologic features suggestive of a secretory variant of ductal carcinoma. The tumors from both sisters had similar overall LOH frequency expressed by the fractional allelic loss (FAL) indices (0.56 vs. 0.60) and demonstrated concordance for loss or retention at 82 of 97 informative markers (85% correlation). In addition, detailed mapping analysis of several chromosomal arms revealed that identical breakpoints were detected in both tumors at several chromosome regions. Finally, in both sisters' tumors, when a chromosome exhibited allelic loss, all of the markers exhibited LOH of the same parental allele even when there were intervening regions of retention of heterozygosity. In contrast, 17 archival sporadic breast carcinomas demonstrated a wide range of FAL indexes and highly individual patterns of LOH. Our findings support the hypothesis that inherited factors play a role in the development of the multiple somatic deletions occurring in breast carcinomas. Whether one of these factors is the mutant BRCA1 allele or some other gene(s) remains to be determined. Genes Chromosomes Cancer 28:359-369, 2000. PMID- 10862045 TI - Detection of chromosome imbalances in retinoblastoma by parallel karyotype and CGH analyses. AB - We have studied a series of 20 primary retinoblastomas by karyotypic analysis and comparative genomic hybridization (CGH), to perform an exhaustive evaluation of chromosome imbalances in this tumor. In addition, 4 tumors were studied by CGH only. On the whole, CGH results were largely in agreement with those of karyotypic analysis and with known cytogenetic data. The most frequent imbalances were +6p (13/24 cases), +1q (12/24), -16/-16q (11/24), and +2p (9/24). Recurrent high-level amplifications were observed in 2p23-25 and 1q21. Amplification of 2p23-25, present in 4 cases among which 3 showed double-minute chromosomes, was related to MYCN amplification, as demonstrated by FISH and PCR. No evident correlation was found in this small series between any of the imbalances identified and either the differentiation or the histoprognostic risk. PMID- 10862046 TI - Marginal zone B-cell lymphomas (MZBL) arising at different sites represent different biological entities. AB - Most entities of B-cell malignant non-Hodgkin's lymphomas (NHL) are characterized by typical primary chromosomal changes such as the t(14;18) in follicular lymphoma or the t(11;14) in mantle cell lymphoma. In contrast, marginal zone B cell lymphomas (MZBL), arising at different nodal and extranodal sites, are poorly characterized on the genetic level. We performed cytogenetic investigations in 20 splenic and in 10 nodal MZBL and analyzed 52 MZBL (including 12 MALT-type lymphomas) for deletions of TP53, D13S25, and RB1 loci by fluorescence in situ hybridization. A new nonrandom chromosomal aberration, del(10)(q22q24), was found as a clonal anomaly in 3 out of 20 cases of splenic MZBL. Further recurring abnormalities such as del(7q) or trisomy 3 were found to be characteristic chromosomal changes in a subset of splenic MZBL. TP53 was deleted in 5/25 cases of splenic MZBL. Deletions involving band 13q14 were only rarely encountered, challenging a previous report that stated a dissociated D13S25-RB1 status as characteristic in splenic MZBL. There are fundamental differences between the different subtypes of marginal zone lymphomas as defined with current classification schemes. Splenic MZBL, in contrast to most other entities of B-cell NHL, seems to constitute a heterogeneous disease especially with regard to genetic alterations. del(10)(q22q24) could be of importance at least in a subset of this lymphoma entity. PMID- 10862047 TI - Loss of heterozygosity in spontaneous and X-ray-induced intestinal tumors arising in F1 hybrid min mice: evidence for sequential loss of APC(+) and Dpc4 in tumor development. AB - Min (multiple intestinal neoplasia) mice carry a mutant allele of the murine Apc (adenomatous polyposis coli) locus and are predisposed to intestinal adenoma formation in the intestinal tract. Early studies have shown complete loss of function of Apc by whole chromosome loss on the tumor-sensitive C57BL/6J genetic background and in AKR x B6 F1 hybrids. Gamma-radiation-induced chromosomal losses focus the critical region on wt Apc, but because of the limited number of polymorphic markers used, no other critical regions of loss on chromosome 18 were identified. Using intestinal tumors arising spontaneously and induced by X-rays in CBA/H x C57BL/6J F1 hybrid mice and high-resolution microsatellite loss of heterozygosity (LOH) techniques, we provide mapping data for wt Apc loss, which confirms and extends earlier observations. In addition, high-frequency loss events at the Dpc4 locus were found in both spontaneous and radiation-induced tumors. These data identified LOH of Dpc4 as a critical secondary event following complete functional loss of Apc. LOH across the Trp53 genomic region of chromosome 11 was not observed. No LOH was recorded for the Mom1 candidate gene Pla2g2a or for 9 out of 10 polymorphic markers from the Mom1 genomic region on murine chromosome 4. One marker mapping distal to Pla2g2a showed LOH in a small minority of spontaneous tumors. These data support the contention that Mom1 does not act as a classical tumor suppressor. Overall, our data indicates a significant role for Dpc4 mutation in intestinal tumor progression in the mouse and provides further evidence for the importance of interstitial chromosome losses in radiation tumorigenesis. PMID- 10862048 TI - Improving degenerate oligonucleotide primed PCR-comparative genomic hybridization for analysis of DNA copy number changes in tumors. AB - Combining degenerate oligonucleotide-primed PCR (DOP-PCR) with comparative genomic hybridization (CGH) has made it possible to analyze genomic changes in single cells. Although DOP-PCR-CGH methodology has been reported, the reproducibility of the method has been uncertain. We have developed a reproducible DOP-PCR-CGH protocol by systematically evaluating different labeling methods (including nick translation, PCR incorporation, and random-primed labeling) and different hybridization mixtures (including amplified test DNA vs. amplified reference DNA, termed homo-hybridization; and amplified test DNA vs. unamplified reference DNA or vice versa, termed hetero-hybridization). We have analyzed DNA samples obtained from 16 tissue sources including fresh/frozen normal and tumor samples, formalin fixed and paraffin embedded tumor tissue, and tumor cell lines by using differently labeled probes and hybridization combinations, and we calculated the corresponding rate (CR) of DOP-PCR-CGH with standard CGH. We found that homo-hybridization produced reproducible results with high CRs as compared to standard CGH (91-100% CR, mean 97%); In contrast, hetero hybridization failed to generate reproducible hybridization with low CRs (57-97% CR, mean 80%; chi(2) = 1245.8, P<0.0001), high background, uneven hybridization, and false deletions or amplifications. In addition, our improved DOP-PCR protocol raised the amplification efficiency at least five times as compared to previously reported protocols, allowing for the detection of genomic imbalances in as little as 12.5 pg of starting DNA. In conclusion, the DOP-PCR-CHG homo-hybridization method, especially when combined with labeling by nick translation, is reliable and reproducible. The method can be used in screening for genomic imbalances using minute amounts of tumor DNA, thereby facilitating CGH application. Genes Chromosomes Cancer 28:395-403, 2000. PMID- 10862049 TI - Analysis of G(1)/S checkpoint regulators in metastatic melanoma. AB - We have analyzed the expression of the CDKN1A (p21(CIP1)), CDKN1B (p27(Kip1)), TP53, RB1 and MDM2 proteins and tumor cell proliferation by immunohistochemical staining in 59 cases of metastatic melanoma. The genomic status of the CDKN2A (INK4-ARF, p16/p14(ARF)), CDKN2B (p15) and CDKN2C (p18) genes was determined by PCR-SSCP (single-strand conformation polymorphism) in 46 of these cases. These results were correlated with various clinico-pathological parameters, including the outcome of combined chemoimmunotherapy. We found positive correlations between the expression of CDKN1A and MDM2 (r = 0.5063, P = 0.001), between the expression of CDKN1B and RB1 (r = 0.5026, P = 0.001), and between RB1 expression and tumor cell proliferation (0.5564, P<0.001). Two mutations in the CDKN2A (p16) gene were detected, including a novel base change AAC-->ATC (Asn to Ile) at codon 71, that also changes the codon 85 of the alternative reading frame gene p14(ARF) from CAA to CAT (Gln to His). Homozygous deletion at exon 2 of the CDKN2A (INK4 ARF) gene was detected in six cases. In seven cases, the 540C-->G polymorphism in the 3'UTR of the CDKN2A (p16) gene was found in linkage disequilibrium with the 74C-->A polymorphism in intron 1 of the CDKN2B gene (P < 0.0001). These cases had significantly lower expression of the TP53 protein (P = 0.0032). Both 540C-->G and 580C-->T polymorphisms in the 3'UTR of the CDKN2A (p16) gene were associated with significantly shorter progression time from primary to metastatic disease (P = 0.0071). We conclude, that although none of the analyzed cell cycle regulators could be singled out as a major prognostic factor, G(1)/S checkpoint abnormalities remain one of the most significant factors in the development of malignant melanoma. PMID- 10862050 TI - RT-PCR analysis of the MOZ-CBP and CBP-MOZ chimeric transcripts in acute myeloid leukemias with t(8;16)(p11;p13). AB - The translocation t(8;16)(p11;p13) is associated with a subtype of acute monocytic leukemia (AML M5) characterized morphologically by erythrophagocytosis and clinically by a poor prognosis. The t(8;16) fuses the MOZ gene from 8p11 with the CBP (also named CREBBP) gene from 16p13. Previously published studies of MOZ and CBP rearrangements in t(8;16)-positive AML have used fluorescence in situ hybridization and Southern blot methodologies, whereas attempts to amplify and to analyze further the chimeric MOZ-CBP and CBP-MOZ transcripts by means of reverse transcriptase-polymerase chain reaction (RT-PCR) have largely been unsuccessful. In the only t(8;16) that has been described at the sequence level using RT-PCR, the CBP-MOZ fusion was found to be out-of-frame, suggesting that the reciprocal MOZ-CBP transcript is the essential one for leukemogenesis. We have developed an RT-PCR strategy that enables us to detect the MOZ-CBP as well as the CBP-MOZ fusions in the two AML M5 with t(8;16)(p11;p13) analyzed. In both leukemias, the combination of a MOZ forward and a CBP reverse primer amplified a strongly expressed 1,128 bp fragment (type I transcript) and a weakly expressed 415 bp fragment (type II transcript). In the type I transcript, nucleotide (nt) 3,745 of MOZ was fused in-frame with nt 284 of CBP, whereas in the type II transcript, nt 3,745 of MOZ was fused out-of-frame with nt 997 of CBP. Nested PCR with a combination of two forward CBP and two reverse MOZ primers amplified CBP-MOZ chimeric transcripts in both cases. Direct sequence analysis showed that nt 283 of CBP was fused in-frame with nt 3,746 of MOZ, that the initiation ATG codon of the CBP gene remained intact, and that there was no mutation or deletion in the part of the CBP gene included in the CBP-MOZ transcript. Thus, the data we present are not informative with regard to the question whether it is the MOZ-CBP or the CBP-MOZ transcript that is leukemogenic. The present RT-PCR method may be of value for rapid identification of the t(8;16) and also for further molecular genetic studies of the two fusion transcripts and their roles in leukemogenesis. PMID- 10862051 TI - Chromosome 17 loss-of-heterozygosity studies in benign and malignant tumors in neurofibromatosis type 1. AB - Neurofibromatosis type 1 (NF1) is a common autosomal dominant condition characterized by benign tumor (neurofibroma) growth and increased risk of malignancy. Dermal neurofibromas, arising from superficial nerves, are primarily of cosmetic significance, whereas plexiform neurofibromas, typically larger and associated with deeply placed nerves, extend into contiguous tissues and may cause serious functional impairment. Malignant peripheral nerve sheath tumors (MPNSTs) seem to arise from plexiform neurofibromas. The NF1 gene, on chromosome segment 17q11.2, encodes a protein that has tumor suppressor function. Loss of heterozygosity (LOH) for NF1 has been reported in some neurofibromas and NF1 malignancies, but plexiform tumors have been poorly represented. Also, the studies did not always employ the same markers, preventing simple comparison of the frequency and extent of LOH among different tumor types. Our chromosome 17 LOH analysis in a cohort of three tumor types was positive for NF1 allele loss in 2/15 (13%) dermal neurofibromas, 4/10 (40%) plexiform neurofibromas, and 3/5 (60%) MPNSTs. Although the region of loss varied, the p arm (including TP53) was lost only in malignant tumors. The losses in the plexiform tumors all included sequences distal to NF1. No subtle TP53 mutations were found in any tumors. This study also reports the identification of both NF1 "hits" in plexiform tumors, further supporting the tumor suppressor role of the NF1 gene in this tumor type. PMID- 10862052 TI - Multiple copies of mutant BRCA1 and BRCA2 alleles in breast tumors from germ-line mutation carriers. AB - Inactivation of the BRCA1 and BRCA2 breast cancer susceptibility genes has been reported to occur via a germ-line mutation of one allele and a somatic loss of the remaining wild-type allele. We investigated the genetic mechanisms behind the second event in breast tumors from 17 BRCA1 and eight BRCA2 germ-line mutation carriers, as compared with 21 sporadic breast tumors. Microsatellite markers intragenic or in close proximity to both genes were used to analyze imbalances between the mutant and wild-type alleles. The actual and relative gene copy numbers were scored by fluorescence in situ hybridization (FISH) analysis of tumor cells using locus and centromere specific probes. All but one of the informative BRCA1 and BRCA2 tumors exhibited allelic imbalance and loss of the corresponding wild type allele. In contrast to sporadic tumors, however, where allelic imbalance at the BRCA1 and BRCA2 loci correlated well with relative copy number losses by FISH, a simple reduction to a single copy (average copy number ratio 2:1) was found in only two BRCA1 (12%) and four BRCA2 (50%) tumors. The majority of BRCA1 and BRCA2 tumors showed a copy number reduction (relative to reference probe with ratios 4:2, 3:2, 4:3) at corresponding loci, suggesting that a specific physical deletion of the wild-type BRCA gene allele has been followed by a duplication of the remaining mutant allele via polyploidization. Several tumors contained multiple copies of BRCA1 and BRCA2 genes without relative copy number changes, implying that loss of wild-type alleles is executed by alternative mechanisms such as mitotic recombination, non-disjunctional chromosomal loss with or without reduplication, or by gene conversion. A paradoxical relative copy number gain of the mutant allele was evident in three BRCA1 tumors (18%), which could be of biological relevance if a dominant negative or gain-of-function model was ascribed for certain BRCA1 mutants. Our results indicate that complex genetic alterations are operational at the BRCA1 and BRCA2 loci in tumors from genetically predisposed individuals. PMID- 10862053 TI - Sequence variants of the axin gene in breast, colon, and other cancers: an analysis of mutations that interfere with GSK3 binding. AB - Axin is a recently discovered component of a multiprotein complex containing APC, beta-catenin, GSK3, and PP2A, which functions in the degradation of the beta catenin protein. As part of WNT signal transduction, the function of the Axin complex is inhibited, leading to the accumulation of beta-catenin. The inappropriate stabilization of beta-catenin has been implicated in a range of human tumors. Two oncogenic mechanisms leading to beta-catenin stabilization are the loss of the APC tumor suppressor protein and the mutational activation of beta-catenin, such that the Axin/APC complex can no longer regulate it. Studies in Drosophila and mammalian tissue culture showed loss of Axin function interfered with beta-catenin turnover and activated beta-catenin/TCF-dependent transcription. Based on these observations, Axin was screened for mutations in a range of human tumor cell lines and primary breast tumor samples. We identified two sequence variants causing amino acid substitutions in four colon cancer cell lines, a Ser-to-Leu at residue 215 in LS513 and a Leu-to-Met at residue 396 in HCT-8, HCT-15, and DLD-1. The Axin L396M mutation was selected for further study since it lay within a region that was shown to interact with glycogen synthase kinase-3. Biochemical and functional studies showed that the L396M change interfered with Axin's ability to bind GSK3. Interestingly, this mutation and a neighboring L392M change differentially altered Axin's ability to interfere with two upstream activators of TCF-dependent transcription, Frat1 and Disheveled. PMID- 10862054 TI - BOLD-f MRI response to single-pulse transcranial magnetic stimulation (TMS). AB - Five healthy volunteers were studied using interleaved transcranial magnetic stimulation/functional magnetic resonance imaging (TMS/fMRI) and an averaged single trial (AST) protocol. Blood oxygenation level-dependent (BOLD)-fMRI response to single TMS pulses over the motor cortex was detectable in both the ipsilateral motor cortex under the TMS coil and the contralateral motor cortex, as well as bilaterally in the auditory cortex. The associated BOLD signal increase showed the typical fMRI hemodynamic response time course. The brain's response to a single TMS pulse over the motor cortex at 120% of the level required to induce thumb movement (1.0%-1.5% signal increase) was comparable in both level and duration to the auditory cortex response to the sound accompanying the TMS pulse (1.5% -2.0% signal increase). PMID- 10862055 TI - Quantification of endothelial permeability, leakage space, and blood volume in brain tumors using combined T1 and T2* contrast-enhanced dynamic MR imaging. AB - This study describes a method for imaging brain tumors that combines T1-weighted (T1W) and T2*-weighted (T2*W) dynamic contrast-enhanced acquisitions. Several technical improvements have been made to produce high-quality three-dimensional mapping of endothelial permeability surface area product (k) and leakage space (vl), based on T1W data. Tumor blood volume maps are obtained from T2*W images with a complete removal of residual relaxivity effects. The method was employed in 15 patients with brain tumors (5 gliomas, 5 meningioma, and 5 acoustic schwannoma). Mean values of vl were significantly greater in acoustic schwannomas (53% +/- 9%) than in meningiomas (34% +/- 7%) or gliomas (22% +/- 4%). Mean values of vl in meningioma were significantly greater than those of gliomas. Mean values of rCBV correlated closely with k. There was also a positive correlation between k and vl for pixels with low k values. This relationship was weaker in areas of high k. The highest mean ratios of k to vl (k(ep)) were seen in two patients with glioblastoma, one patient with transitional cell meningioma, and one patient with angioblastic meningioma. Pixel-by-pixel comparison showed a strong correlation between rCBV and k in 11 of 15 patients. However, decoupling between pixel-wise rCBV and k was found in four patients who had lesions with moderate k and vl elevation but no increase of rCBV. Results from this study suggest that in assessing the angiogenic activities in brain tumors it is advisable to monitor simultaneously changes in tumor blood volume, vessel permeability, and leakage space of tumor neovasculature. PMID- 10862056 TI - Magnetization transfer and multicomponent T2 relaxation measurements with histopathologic correlation in an experimental model of MS. AB - Magnetization transfer and multicomponent T2 imaging techniques were implemented to study guinea pig in vivo. A chronic-progressive model of experimental allergic encephalomyelitis (EAE) was produced, and the inflammatory component of the disease was manipulated using antibodies against integrin. The magnetization transfer ratio (MTR) and T2 relaxation properties were measured in normal appearing white matter (NAWM) with histological comparisons. Significant reductions in both the mean MTR and the myelin water percentage were measured in NAWM of EAE guinea pig brain. However, the MTR and myelin water percentage appear to measure different aspects of pathology in NAWM in EAE. Reductions in the MTR were prevented or reversed with suppression of inflammation. However, modulation of inflammatory activity was not reflected in the measurement of the myelin water percentage. Since the amount of myelin is not expected to vary with inflammatory related changes, these observations support our hypothesis that the MTR is sensitive to physiological changes to myelin induced by inflammation, while the short T2 component is a more specific indicator of myelin content in tissue. Pathologic features other than demyelination may be important in the determination of the MTR. PMID- 10862057 TI - Magnetic resonance spectroscopy (MRS) in five patients with treated propionic acidemia. AB - Propionic acidemia is an inherited disorder caused by a defect of propionyl CoA carboxylase. Untreated, propionic acidemia leads to metabolic decompensation and toxic encephalopathy. We report on the magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) findings in five children who were properly treated by protein restriction and carnitine supplementation, during a phase of clinically and metabolically stable conditions. The examinations were performed on a whole-body 1.5 T scanner. During the observation period, from 1992 to 1996 we employed long echo time single-voxel spectroscopy and chemical shift imaging in addition to a conventional MRI protocol. The two children with the longest delay before onset of therapy showed cerebral atrophy. MRS yielded elevated lactate peaks in four of the children. These results indicate that MRS can detect metabolic alterations in the brains of children with propionic acidemia during metabolically stable conditions. The presence of lactate could be caused by hampered aerobic oxidation within the citrate cycle due to intracellular elevated propionic metabolites. PMID- 10862058 TI - Correlation of high-resolution breast MR imaging with histopathology; validation of a technique. AB - A high-resolution three-dimensional surface gradient coil set was used to obtain magnetic resonance (MR) images of breast specimens, using a gradient-echo pulse sequence (TR/TE 1000/8 msec, flip angle 75 degrees), with 117 micrometer in-plane resolution and 1 mm slice thickness. Breast tissues were obtained from one autopsy and three surgical specimens. High-resolution breast MR images and histopathology sections (7 micrometer thickness) were acquired in the same anatomical plane. Radiographs were acquired of the sliced specimens (approximately 5 mm thick) so that images from all three methods could be correlated. It was found that in vitro high-resolution breast MRI correlated well with low-resolution microscopic histology, demonstrating normal anatomy (lobules, ducts, connective tissue strands, blood vessels) and pathology (tumor content, margins, and presence of microcalcifications) of the breast more clearly than conventional pre-gadolinium breast MRI. High-resolution breast MRI may improve specificity, when added to a conventional breast MRI protocol. PMID- 10862059 TI - Magnetic resonance imaging of regional myocardial perfusion in patients with single-vessel coronary artery disease: quantitative comparison with (201)Thallium SPECT and coronary angiography. AB - The clinical value of magnetic resonance perfusion imaging (MRI) was investigated by quantitative comparison with (201)thallium-single-photon emission computed tomography ((201)TI-SPECT) and quantitative coronary angiography (QCA). Short axis imaging was performed during dipyridamole administration in 13 patients with single-vessel coronary artery disease. Using inner and outer contours, the myocardium was divided into 30 contiguous, radial regions. Defining a perfusion defect as a region with less than 90% of maximum (201)TI intensity, nine patients had a matching perfusion defect, two had no defect on both (201)TI-SPECT or MRI, and one had a defect on (201)TI-SPECT but not on MRI. One patient had a defect on both modalities but with inaccurate localization. Three perfusion parameters were investigated: a) maximum contrast enhancement (MCE); b) slope of the signal intensity versus time curve; and c) inverse mean transit time (1/MTT). The sensitivity and specificity of MCE in the detection of perfusion abnormalities with TI-SPECT as the reference method were 71% and 71%, respectively (slope 77% and 61%, 1/MTT 44% and 70%). Furthermore, correlations were calculated per patient for the entire circumference of the short-axis myocardium. Median correlations were as follows: MCE 0.92, slope 0.91, and 1/MTT 0.40. Mismatches between (201)TI defects and defects on MRI resulted in low mean correlations (MCE 0.45, slope 0.46, and 1/MTT 0.26). There was a trend between severity of perfusion defects on MRI (using MCE) and QCA stenosis area (r = -0.56, P = 0.06). Thus, MRI and (201)TI-SPECT demonstrate fair agreement in the assessment of perfusion defects but show moderate correlation when the entire short-axis myocardium is correlated. PMID- 10862060 TI - 1H magnetic resonance imaging of human lung using inversion recovery turbo spin echo. AB - Evaluation of lung pathologies using magnetic resonance imaging remains limited, primarily due to the lung's low proton density and high density of magnetic field susceptibility gradients. It is hypothesized that visualization of the lung is possible if signal intensity from muscle and/or fat is suppressed or reduced. Using the inversion recovery and frequency selective saturation pulse with a half Fourier single-shot turbo spin-echo (HASTE) or a segmented, centric reordered turbo spin-echo (TSE) readout, signal intensity and contrast of tissues can be manipulated to enhance the visibility of the lung. Multislice images of the lung from 10 healthy volunteers were acquired with negligible motion artifacts. Peripheral pulmonary vessels appear well delineated. T(1) maps of the lung are also presented; the overall average was 1335 +/- 85 msec and 1245 +/- 93 msec with the volunteers performing breath-holding on end-expiration and end inspiration, respectively. This difference is statistically significant, at P < 0.01. PMID- 10862061 TI - Three-dimensional MR imaging of pulmonary vessels and parenchyma with NC100150 injection (Clariscan). AB - The influence of increasing doses of NC100150 Injection (Clariscantrade mark) and echo times on visualization of pulmonary vessels and parenchyma was evaluated. The effects of 0.5, 1, 2, 4, and 8 mg Fe/kg NC100150 Injection and echo times (TE) of 1.1, 1.8, 2. 2, and 4.3 msec were determined in six dogs using breath hold three-dimensional (3D) spoiled gradient-echo magnetic resonance (MR) sequence. At 2 mg Fe/kg and TE of 1.1 msec, the signal-to-noise ratio of the central pulmonary arteries and parenchyma was significantly increased (5.3 +/- 2.2 to 50.3 +/- 2.4) and (2.2 +/- 0. 9 to 6.4 +/- 1.1), respectively. Using the TE of 1.1 msec, signal intensity in the main arteries continued to increase with increasing dose. Moreover, the enhancement of pulmonary parenchyma and microvasculature had a positive dose response. 3D MR imaging with ultrashort echo time and 2 mg Fe/kg NC100150 Injection produces angiograms with strong vascular contrast and allows qualitative assessment of pulmonary parenchyma and microvasculature. PMID- 10862062 TI - MR imaging of solitary pulmonary lesion: emphasis on tuberculomas and comparison with tumors. AB - The aim of this study was to determine whether solitary pulmonary tuberculoma and malignant tumor can be differentiated on the basis of magnetic resonance (MR) signal intensity. Twenty-eight patients with solitary pulmonary lesions were prospectively studied with MR imaging: T1-weighted, enhanced T1-weighted, proton density-weighted, and T2-weighted spin echo images were obtained. The confirmation methods used were computed tomography (CT)-guided biopsy in seven patients with lung cancer and four patients with tuberculosis; surgery in ten patients with lung cancer and five patients with tuberculosis; and laboratory data in two patients with tuberculosis. Morphologic features and MR signal intensity were examined in detail. As the test for detection of tuberculoma, signal difference on T2-weighted images was carefully analyzed. The signal intensity ratio of the nodule to thoracic muscle signal intensity was measured. The signal intensities obtained from the lung cancers and tuberculomas were variable on pre-and post-enhanced T1-weighted images and proton density-weighted images. Masses were hypointense in 2 of 17 patients with lung cancer and in 9 of 11 patients with tuberculoma on T2-weighted images (sensitivity 82%, specificity 89%, accuracy 87%). The mean signal intensity ratios of the tuberculomas to muscle were significantly lower than those of malignant tumors on T1-weighted, enhanced T1-weighted, proton density-weighted, and T2-weighted images (P < 0.0001). After gadolinium-DTPA enhancement, 2 malignant tumors and 7 tuberculomas showed a marginal rim enhancement pattern, whereas 15 malignant tumors and 2 tuberculomas revealed a diffuse enhancement. The results of MR imaging were consistent with those of CT in 84% of the patients. MR imaging is a helpful adjunctive method in terms of differentiating a tuberculoma from a malignant tumor. PMID- 10862063 TI - Time-of-flight MR angiography with Gd-DTPA hexamethylene diamine co-polymer blood pool contrast agent: comparison of enhanced MRA and conventional angiography for arterial stenosis induced in rabbits. AB - Vascular stenoses were induced in the external iliac arteries of New Zealand white rabbits by a combination of hypercholesterolemic diet and repeat balloon injury. Two-dimensional (2D) and three-dimensional (3D) time-of-flight (TOF) magnetic resonance angiography (MRA) was performed with a specifically designed phased array coil in a 1.5 T system. Enhancement with gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA) hexamethylene diamine co-polymer (Nycomed: NC 22181), a blood pool MR contrast agent, was measured after contrast administration and compared with pre-contrast images at the same levels. Vessel diameter measurements were obtained at multiple levels and compared with comparable levels on conventional angiograms of the same animals. Stable enhancement, averaging 227% above baseline, was observed with the 3D TOF MRA over the 40 minutes of this study. Enhancement was not observed with the 2D TOF technique. Measurement of the smallest vessels in this study with 3D TOF MRA was slightly improved following contrast enhancement, although both pre- and post contrast diameter measurements tended to underestimate the assumed true vessel diameter. Thus, Gd-DTPA hexamethylene diamine co-polymer (Nycomed: NC 22181), a blood pool MR contrast agent, produces significant, stable enhancement with the 3D TOF technique and may improve MRA measurement of small vessels. PMID- 10862064 TI - Improved tissue characterization of focal liver lesions with ferumoxide-enhanced T1 and T2-weighted MR imaging. AB - The purpose of our study was to evaluate the potential value of ferumoxide enhanced T1-weighted magnetic resonance (MR) imaging for tissue characterization of focal liver lesions when combined with T2-weighted sequences. Images were acquired within 30 minutes after the end of ferumoxide administration, when ferrite particles were not totally cleared from the intravascular compartment. Thirty-eight patients with 47 focal liver lesions underwent T1-weighted gradient echo (TR/TE 150/4.1 msec) and T2-weighted fast spin-echo (3180-8638/90 msec) MR imaging at 1.5 T before and after intravenous administration of ferumoxides (10 micromol/kg body weight). A qualitative and quantitative analysis was performed. During the early phase after infusion of ferumoxide, blood vessels showed hypersignal intensity on T1-weighted fast low-angle shot (FLASH) images, while liver signal decreased. Hemangiomas showed both homogeneous and inhomogeneous enhancement patterns, and liver metastasis most typically showed ring enhancement. Hypervascular tumors (hepatocellular carcinomas and focal nodular hyperplasias) showed a slight degree of homogeneous enhancement. Quantitatively, the degree of enhancement and lesion-to-liver contrast on ferumoxide-enhanced images were significantly different among these tumors. Our results demonstrate that distinct enhancement patterns obtained on ferumoxide-enhanced T1-weighted MR imaging improve tissue characterization of focal liver lesions when combined with T2-weighted images. PMID- 10862065 TI - Phase II double-blind, dose-ranging clinical evaluation of gadobenate dimeglumine in focal liver lesions: with analysis of liver and kidney signal change on early and delayed imaging. AB - To evaluate the effect of contrast dose using gadobenate dimeglumine, 30 patients with focal liver lesions documented by computed tomography or ultrasound were studied by magnetic resonance imaging at 1.5 T. Patients received one of four doses of gadobenate dimeglumine (0.025, 0.05, 0.1, or 0.2 mmol/kg) or saline. The order of dosage was randomized, with both the physician and patient blinded to the administered dose. Scans were obtained before, immediately following injection, and after 80 minutes of delay. Enhancement effects were quantified by region of interest measurements. Films were also reviewed in a randomized prospective fashion by an abdominal radiologist blinded to contrast dose and diagnosis. Higher doses led to a statistically significant improvement in enhancement of normal liver, both on immediate (P = 0.01 for the comparison of 0.1 and 0.2 mmol/kg immediately post-contrast) and delayed scans (P = 0.003 for the same comparison). Liver-lesion contrast-to-noise ratio also increased with dose, although results for most comparisons by dose were not statistically significant. Scans following gadobenate dimeglumine injection were judged to provide additional diagnostic confidence sufficient to affect patient management in 10 of 24 cases. In seven cases this information was provided by dynamic scans, in one case by delayed scans, and in two cases by both dynamic and delayed scans. In 2 of the 10 cases the dose was 0.025 mmol/kg, in 2 cases 0.05 mmol/kg, in 3 cases 0.1 mmol/kg, and in 3 cases 0.2 mmol/kg. Gadobenate dimeglumine is effective for imaging of focal liver lesions at a range of doses, with trends toward improved diagnostic information at higher doses. PMID- 10862066 TI - Detection of focal hepatic lesions: comparison of unenhanced and SHU 555 A enhanced MR imaging versus biphasic helical CTAP. AB - The purpose of this study was to compare the diagnostic sensitivity of unenhanced magnetic resonance (MR) imaging, and MR imaging with a new superparamagnetic iron oxide (SPIO)-enhanced contrast agent (SHU 555 A) with biphasic helical computed tomography during arterial portography (CTAP) in patients with focal liver lesions. Eighteen patients with a total of 91 (78 malignant, 13 benign) proven liver lesions underwent unenhanced short tau inversion recovery (STIR), T2 weighted (T2-w) TSE, and SHU 555 A-enhanced T2-w turbo spin-echo (TSE) MR imaging and biphasic helical CTAP. The standard of reference was histopathologic analysis of resected specimens in 59 lesions, intraoperative ultrasound with biopsy in 20 lesions, and CT-guided biopsy and follow-up in 12 lesions. Diagnostic performance of the imaging modalities was compared quantitatively and qualitatively by assessing lesion involvement in liver segments. There were 68 lesions detected on unenhanced T2-w TSE, which resulted in a sensitivity of 75%. With the STIR sequence, 76 lesions were detected, for a sensitivity of 84%, and with SHU 555 A enhanced MRI, 84 lesions were detected, for a sensitivity of 92%. CTAP detected 88 lesions, for a sensitivity of 97%. The accuracy for unenhanced T2-w TSE was 98%, for STIR 99%, for enhanced-MRI 100%, and for CTAP 95%. The specificity was 100% for SHU 555 A-enhanced MRI and 95% for CTAP. SHU 555 A-enhanced MRI was superior to nonenhanced MRI (P < 0.05) and equivalent to CTAP in terms of sensitivity. Due to the absence of false-positive results on SHU 555 A-enhanced MRI, the specificity and accuracy of enhanced MRI were higher than those of CTAP, but the difference was not statistically significant (P = 0.134). PMID- 10862067 TI - Single-shot fluid attenuated inversion recovery (FLAIR) magnetic resonance imaging of the bladder. AB - The purpose of this study was to reduce artifacts and increase imaging speed in fluid-attenuated inversion recovery (FLAIR) imaging of the urinary bladder. An existing half-Fourier, single-shot fast spin-echo imaging sequence was modified to allow presaturation with a non-slice-selective inversion recovery pulse (NSI SSFLAIR). Four independent, blinded readers rated severity of bladder artifacts and image quality in six normal male volunteers. NSI SSFLAIR effectively suppressed bladder urine signal in all six cases using a TI of 2900-3100 msec. Although NSI SSFLAIR images were noisier than standard fast spin-echo images, imaging time was only 10 seconds per slice location. Furthermore, perceived image sharpness was only minimally reduced, and conspicuity of the seminal vesicles and peripheral zone of the prostate were nearly equivalent. NSI SSFLAIR provides rapid T2-weighted imaging of the bladder wall and perivesicular tissues with nearly complete negation of signal from urine in the bladder. PMID- 10862068 TI - Detection of articular cartilage lesions: experimental evaluation of low- and high-field-strength MR imaging at 0.18 and 1.0 T. AB - The objective of this study was to compare the diagnostic performance of a dedicated orthopedic magnetic resonance (MR) imaging system (0.18 T) and a conventional MR imaging system (1.0 T) in the detection of articular cartilage lesions. Fifty knee joint specimens of pigs with artificially created articular cartilage lesions of different diameters, grades (2-3), and localizations, as well as 50 joints with intact articular cartilage, were imaged at 0. 18 and 1.0 T. Diagnostic performance was determined by means of receiver operating characteristics (ROC) analysis with three independent observers. For none of the pulse sequences used at 0.18 T or 1.0 T areas under ROC curves (A(z)) showed significant differences between the three observers. A(z) values from averaged data were as follows: a) 0.18 T: T1-weighted spin echo (SE): 0.70, proton-density weighted SE: 0.59, T2-weighted SE: 0.61, two-dimensional (2D) gradient-echo (GRE): 0.73, 3D GRE: 0.75; and b) 1.0 T: T1-weighted SE: 0.73, fat-suppressed T2 weighted turbo-SE: 0. 79, 2D fast low-angle shot (FLASH): 0.79, fat-suppressed 3D FLASH: 0. 96, and water-excited 3D double-echo steady state (DESS): 0.96. With the use of 3D pulse sequences, the high-field system demonstrated a significantly better diagnostic performance than the low-field system in the detection of grades 2 and 3 articular cartilage lesions (P < 0.001). PMID- 10862069 TI - Heterogeneity of cartilage laminae in MR imaging. AB - The purpose of this study was to investigate the discrepancy in the number of laminae observed in magnetic resonance (MR) images of articular cartilage (the magic angle effect in MRI of cartilage). Microscopic MR imaging (muMRI) experiments were carried out at 14-micrometer pixel resolution on full-depth cartilage-bone plugs from several locations (central, intermediate, and peripheral) on the humeral heads of two young healthy beagles. When the articular surface of the plug was perpendicular to the direction of the magnetic field, the cartilage appeared to have two layers in the plugs from the central locations of the humeral head, three layers in the plugs from the greater tubercle side of the humeral head, and three or five layers in the plugs from the lesser tubercle side. This heterogeneity of cartilage laminae was observed within a single humeral head and was symmetrical about the median plane of the animal. This result suggests that some structural variations related to cartilage structure in various regions of load bearing may cause some unique laminar patterns seen in MRI of cartilage. This novel and new observation may resolve the controversy about whether cartilage appears as two or three layers in MR images. A comprehensive model for the collagen structure over a curved two-dimensional surface of a joint is suggested as a replacement of the classic three-zone model of fiber orientation in collagen. This heterogeneity of cartilage laminae is speculated to be related to the load-bearing status of the tissue in the joint. The ability to visualize such structural heterogeneity is important because of the direct connection between collagen structure and the mechanical characteristics of cartilage. PMID- 10862070 TI - A new polysaccharide macromolecular contrast agent for MR imaging: biodistribution and imaging characteristics. AB - The aims of this study were to characterize certain physicochemical, pharmacokinetic, and enhancement properties of a new macromolecular contrast agent, carboxymethyl hydroxyethyl starch-(Gd-DO3A)(35) [CMHES-(Gd-DO3A)(35)], consisting of a polysaccharide backbone covalently derivatized with multiple macrocyclic chelating groups for gadolinium. CMHES-(Gd-DO3A)(35) has an average molecular weight of 72 kD and a plasma half-time of 8.4 hours. T1 and T2 relaxivities are 14.1 +/- 0.1 L mmol(-1) * sec(-1) and 17.8 +/- 0.9 L mmol(-1) * sec(-1), respectively, for each gadolinium ion measured at 39 degrees C and 20 Mhz; this T1 relaxivity is more than 4 times that of gadopentetate. Seven days after intravenous administration only relatively small amounts of gadolinium could be detected in blood or other tissues of rats. The compound was well tolerated in diagnostic dosages by all experimental animals. Magnetic resonance angiography performed within 1 hour of CMHES-(Gd-DO3A)(35) administration showed a near-constant and strong enhancement of blood in arteries and veins. Analysis of dynamic enhancement patterns of experimental tumors (MAT-LyLu prostate cancer implanted in rats) following intravenous CMHES-(Gd-DO3A)(35) administration yielded quantitative estimates of tumor plasma volume and microvessel permeability; the demonstrated hyperpermeability of tumor microvessels was easily distinguished from the absence of measurable microvascular permeability in non neoplastic soft tissues. PMID- 10862071 TI - Nonlinear smoothing for reduction of systematic and random errors in diffusion tensor imaging. AB - Calculation and sorting of the eigenvectors of diffusion using diffusion tensor imaging has previously been shown to be sensitive to noise levels in the acquired data. This sensitivity manifests as random and systematic errors in the diffusion eigenvalues and derived parameters such as indices of anisotropy. An optimized application of nonlinear smoothing techniques to diffusion data prior to calculation of the diffusion tensor is shown to reduce both random and systematic errors, while causing little blurring of anatomical structures. Conversely, filtering applied to calculated images of fractional anisotropy is shown to fail in reducing systematic errors and in recovering anatomical detail. Using both real and simulated brain data sets, it is demonstrated that this approach has the potential to allow acquisition of data that would otherwise be too noisy to be of use. PMID- 10862072 TI - Hybrid cardiac imaging with MR-CAT scan: a feasibility study. AB - We demonstrate the feasibility of a new versatile hybrid imaging concept, the combined acquisition technique (CAT), for cardiac imaging. The cardiac CAT approach, which combines new methodology with existing technology, essentially integrates fast low-angle shot (FLASH) and echoplanar imaging (EPI) modules in a sequential fashion, whereby each acquisition module is employed with independently optimized imaging parameters. One important CAT sequence optimization feature is the ability to use different bandwidths for different acquisition modules. Twelve healthy subjects were imaged using three cardiac CAT acquisition strategies: a) CAT was used to reduce breath-hold duration times while maintaining constant spatial resolution; b) CAT was used to increase spatial resolution in a given breath-hold time; and c) single-heart beat CAT imaging was performed. The results obtained demonstrate the feasibility of cardiac imaging using the CAT approach and the potential of this technique to accelerate the imaging process with almost conserved image quality. PMID- 10862073 TI - Hospital response to DRG refinements: the impact of multiple reimbursement incentives on inpatient length of stay. AB - Recent research has warned that the introduction of Diagnosis Related Groups (DRGs) based on hospital treatment decisions will lead to an increase in the rate of marginal procedures and to a resumption of high medical expenditure growth rates. This paper explores the often contradictory effects of the multiple reimbursement incentives created by refinements to the Prospective Payment System (PPS) (principally, the introduction of procedure-based DRGs) on hospital resource allocation. Three effects are examined in the paper: (i) the change in primary or payment-related procedures owing to marginal reimbursement incentives; (ii) the change in secondary or non-payment-related services owing to average price incentives; and (iii) the change in average severity of both medical and surgical admissions. The model suggests that the anticipated positive effect of marginal reimbursement incentives on overall hospital resource use may be offset by several factors, most notably the lower average payment incentives of non procedural DRGs. PMID- 10862074 TI - Willingness to pay for improved respiratory and cardiovascular health: a multiple format, stated-preference approach. AB - This study uses stated-preference (SP) analysis to measure willingness to pay (WTP) to reduce acute episodes of respiratory and cardiovascular ill health. The SP survey employs a modified version of the health state descriptions used in the Quality of Well Being (QWB) Index. The four health state attributes are symptom, episode duration, activity restrictions and cost. Preferences are elicited using two different SP formats: graded-pair and discrete-choice. The different formats cause subjects to focus on different evaluation strategies. Combining two elicitation formats yields more valid and robust estimates than using only one approach. Estimates of indirect utility function parameters are obtained using advanced panel econometrics for each format separately and jointly. Socio economic differences in health preferences are modelled by allowing the marginal utility of money relative to health attributes to vary across respondents. Because the joint model captures the combined preference information provided by both elicitation formats, these model estimates are used to calculate WTP. The results demonstrate the feasibility of estimating meaningful WTP values for policy-relevant respiratory and cardiac symptoms, even from subjects who never have personally experienced these conditions. Furthermore, because WTP estimates are for individual components of health improvements, estimates can be aggregated in various ways depending upon policy needs. Thus, using generic health attributes facilitates transferring WTP estimates for benefit-cost analysis of a variety of potential health interventions. PMID- 10862075 TI - Mother's willingness to pay for her own and her child's health: a contingent valuation study in Taiwan. AB - We use the contingent valuation (CV) method to estimate mothers' willingness to pay (WTP) to protect themselves and their children from suffering a minor illness a cold-in Taiwan. WTP is specified as a hedonic function of the duration and severity of the cold (measured alternatively by symptoms experienced and the Quality of Well-Being (QWB) index) and of respondents' socioeconomic characteristics. The average mother is willing to pay more to protect her child than herself from suffering a cold. Median WTP to avoid the average mother's and child's colds are US$37 and US$57, respectively. Adjusting for the greater duration and severity of the average mother's cold suggests that WTP to prevent comparable illnesses is approximately twice as large for the child as for the mother. We also find that mother's WTP is about 20% greater to prevent a son's than a daughter's illness. PMID- 10862076 TI - Fieller's method and net health benefits. AB - Statistical and conceptual difficulties complicate the estimation of the incremental cost-effectiveness ratio (ICER). An alternative approach is to measure cost-effectiveness by the incremental net health benefit (INHB), defined as the difference in mean effectiveness of a new treatment compared with a standard, adjusted for cost difference by subtracting the health foregone if purchasing care at the rate of a marginally cost-effective therapy. Because net health benefit (NHB) is dependent on this threshold rate, one can construct confidence intervals for the INHB at various values of the rate. It turns out that the set of rates where new and standard are not significantly different is equal to the Fieller's method confidence set for the ICER. We review the derivation of the Fieller's method confidence set, present numerical examples, and discuss the implications of our result for the calculation and interpretation of NHB analyses. PMID- 10862077 TI - Evidence-based medicine and health economics: a case study of end stage renal disease. AB - This paper explores the potential for use of an economic evaluation framework alongside systematic reviews. Clinical issues in dialysis therapy for end stage renal disease are used as case studies. The effectiveness data required were obtained from a systematic review of randomized controlled trials. Resource use and cost data were obtained from three sources; the identified randomized controlled trials, a separate review of observational studies and primary data collection. The results of the case studies show that, although simple economic evaluations were possible, issues arose, such as how transferable results are between settings and how appropriate it is to focus on the average patient. The interface between economic evaluation and systematic reviews needs to be further developed in order to ensure that the best available evidence can be used to inform future policy and research. PMID- 10862078 TI - Discrete time representation of the formula for calculating DALYs. AB - The global burden of disease (GBD) was measured using a new indicator called disability-adjusted life years (DALYs). The formula to calculate DALYs is based on the idea of time being a continuous variable, which is not consistent with the way economic and health data are collected and reported, and is also different from the concept of time used in the vast majority of policy analyses. Use of this formula gives rise to a time-aggregation bias in the estimates of GBD. Based on discrete time representation and the key principles outlined in the GBD study, a new formula for estimating DALYs is derived in this paper. The properties of the two formulae are compared and contrasted and the implications of using the new formula are discussed. The results show that there is an appreciable difference in percentage terms (14.06%) between the burden of cataract in Sub Saharan Africa in 1990 calculated using the new and the old formulae. The global burden of diseases and injuries as previously reported in the GDB study may, therefore, be underestimated and the relative positions of some diseases and injuries, and hence the relative priorities of related interventions, may shift if the more appropriate, discrete-time formula is used. The difference is greatest for diseases of short duration (e.g. infectious diseases). PMID- 10862079 TI - Twenty two novel mutations of the factor VII gene in factor VII deficiency. AB - Factor VII is a vitamin K-dependent coagulation protease essential for the initiation phase of normal hemostasis. The human factor VII gene (FVII, also known as F7) spans 13 kb and is located on chromosome 13, 2.8 kb upstream of the factor X gene. In the Greifswald FVII deficiency study the molecular basis for inherited factor VII was investigated. All exons, exon-intron boundaries and the promotor of the FVII gene were amplified by PCR and directly sequenced. 87 unrelated probands with reduced or low FVII activities were investigated. Thirty four different FVII gene lesions were analyzed in 101 FVII alleles of 77 unrelated probands. Twenty-two of these FVII gene lesions are novel FVII variations. The 34 different lesions comprise 31 point mutations and three small deletions. A transition in the CpG doublet accounted for 12 of the 34 different mutants. Sixteen mutations were noted only once. The missense mutation A294V and the double mutation A294V; 11128delC in exon 8 were by far the most common mutations found in this study. The haplotype of the different mutant FVII alleles were analyzed using six polymorphisms of the FVII gene. The haplotypes were identified in 29 mutant FVII alleles. Five different haplotypes are linked to the mutant FVII alleles. Except for one, the same haplotype was detected in FVII genes with an identical FVII gene mutation. Different haplotypes were identified in two patients with the mutant allele A206T. It is likely that identical mutant FVII alleles with the same haplotype share the same origin. PMID- 10862080 TI - Analysis of CDKN1C in Beckwith Wiedemann syndrome. AB - In this study we have examined 32 patients with Beckwith Wiedemann Syndrome (BWS) for mutations affecting the CDKN1C gene, including seven cases of familial BWS. Mutations were not detected in the coding region of the CDKN1C gene in any individual with BWS. However in two patients, two G/A base substitutions at adjacent positions in the 5'UTR were detected. These substitutions were also found in normal controls. Expression of CDKN1C in somatic tissues was examined in 18 of the 32 cases using semi-quantitative RT-PCR. CDKN1C expression was significantly reduced in the peripheral blood of three cases compared with controls. These results suggest that, although coding region mutations in the CDKN1C gene are rare in BWS, mutations disrupting CDKN1C expression may be found. Three of five informative patients exhibited biallelic CDKN1C expression in lymphocytes, cord blood, and kidney tissue, respectively. Biallelic expression was not associated with overall CDKN1C levels significantly different to those in controls. Patients who expressed CDKN1C biallelically, or who were low CDKN1C expressors, maintained monoallelic methylation in the Differentially Methylated Region 2 (DMR2) of the IGF2 locus. One patient expressing CDKN1C biallelically, maintained imprinted gene expression at the IGF2 locus. These results suggest that biallelic CDKN1C expression does not significantly perturb the overall levels of CDKN1C expression in somatic tissue. They also confirm other studies showing that the mechanisms associated with regulating CDKN1C expression and imprinting are separate from those regulating IGF2 imprinting. PMID- 10862081 TI - Phenotype-genotype relationships in PEX10-deficient peroxisome biogenesis disorder patients. AB - The peroxisome biogenesis disorders (PBD) are characterized by neural, hepatic, and renal deficiencies, severe mental retardation, and are often lethal. These disorders are genetically and phenotypically heterogeneous and are caused by defective peroxisomal protein import and decreased peroxisomal metabolic function. Mutations in PEX10 have been identified in patients from complementation group 7 (CG7) of the PBDs and we report here an analysis of the genotypes and phenotypes of PEX10-deficient patients. All four PEX10-deficient Zellweger Syndrome (ZS) patients were found to have nonsense, frameshift, or splice site mutations that remove large portions of the PEX10 coding region. In contrast, a more mildly affected PEX10-deficient neonatal adrenoleukodystrophy patient expressed a PEX10 allele with a missense mutation, H290Q, affecting the C terminal zinc-binding domain of the PEX10 product. These results support the hypothesis that severe, loss-of-function mutations in PEX genes cause more severe clinical phenotypes, whereas mildly affected PBD patients have PEX gene mutations that retain residual function. To quantitate the effects of the PEX10 mutations identified here and elsewhere we employed a functional complementation assay. Surprisingly, we observed that nonsense and frameshift mutations predicted to delete the C-terminal 2/3 (R125X) or 1/3 (c.704insA) of the protein displayed nearly normal PEX10 activity. Even more surprising, we found that the unexpectedly high PEX10 activity displayed by these cDNAs could be eliminated by removing or mutating segments of the PEX10 cDNA downstream of the mutations. Although these results demonstrate serious flaws in the PEX10 functional complementation assay, they do suggest that the C-terminal zinc-binding domain is critical for PEX10 function. PMID- 10862082 TI - Clinical differences in patients with mitochondriocytopathies due to nuclear versus mitochondrial DNA mutations. AB - Defects in oxidative phosphorylation (OXPHOS) are genetically unique because the different components involved in this process, respiratory chain enzyme complexes (I, III, and IV) and complex V, are encoded by nuclear and mitochondrial genome. The objective of the study was to assess whether there are clinical differences in patients suffering from OXPHOS defects caused by nuclear or mitochondrial DNA (mtDNA) mutations. We studied 16 families with > or = two siblings with a genetically established OXPHOS deficiency, four due to a nuclear gene mutation and 12 due to a mtDNA mutation. Siblings with a nuclear gene mutation showed very similar clinical pictures that became manifest in the first years (ranging from first months to early childhood). There was a severe progressive course. Seven of the eight children died in their first decade. Conversely, siblings with a mtDNA mutation had clinical pictures that varied from almost alike to very distinct. They became symptomatic at an older age (ranging from childhood to adulthood), with the exception of defects associated with Leigh or Leigh-like phenotype. The clinical course was more gradual and relatively less severe; four of the 26 patients died, one in his second year, another in her second decade and two in their sixth decade. There are differences in age at onset, severity of clinical course, outcome, and intrafamilial variability in patients affected of an OXPHOS defect due to nuclear or mtDNA mutations. Patients with nuclear mutations become symptomatic at a young age, and have a severe clinical course. Patients with mtDNA mutations show a wider clinical spectrum of age at onset and severity. These differences may be of importance regarding the choice of which genome to study in affected patients as well as with respect to genetic counseling. PMID- 10862083 TI - A novel St(a) glycophorin produced via gene conversion of pseudoexon III from glycophorin E to glycophorin A gene. AB - Stone (St(a)) is a variant antigen carried on human erythrocyte MNSs glycophorins (GPSt(a)) that are genetically associated with splicing mutations in GPA genes or with hybrid formation between GPA and GPB genes. Here we identify the first and rare gene conversion event in which GPE, the third member of the family, recombined with GPA, giving rise to a GPA-E-A hybrid gene encoding the St(a) antigen. Western blot detected expression in the proband of both GPA and GPSt(a) on the plasma membrane. Southern blot showed a new restriction fragment from the GPSt(a) gene, indicating an altered exon III-intron 3 junction. Sequencing of RT PCR products identified one full-length and two shortened glycophorin cDNAs. The shortened forms were derived from GPSt(a) lacking one (exon III) and two exons (exon III and IV), respectively. To define the molecular basis for exon skipping, the genomic region spanning exon III of the GPSt(a) gene was amplified and sequenced. This revealed transfer from GPE to GPA of a DNA segment containing the pseudoexon III and its silent donor splice site. Thus, the inactivation of GPA exon III by conversion of a silent GPE donor splice site portrays a new molecular mechanism for St(a) antigen expression in human erythrocytes. PMID- 10862084 TI - Exhaustive mutation analysis of the NF1 gene allows identification of 95% of mutations and reveals a high frequency of unusual splicing defects. AB - Neurofibromatosis type 1 (NF1) is one of the most common autosomal dominant disorders and is caused by mutations in the NF1 gene. Mutation detection is complex due to the large size of the NF1 gene, the presence of pseudogenes and the great variety of possible lesions. Although there is no evidence for locus heterogeneity in NF1, mutation detection rates rarely exceed 50%. We studied 67 unrelated NF1 patients fulfilling the NIH diagnostic criteria, 29 familial and 38 sporadic cases, using a cascade of complementary techniques. We performed a protein truncation test starting from puromycin-treated EBV cell lines and, if no mutation was found, continued with heteroduplex, FISH, Southern blot and cytogenetic analysis. We identified the germline mutation in 64 of 67 patients and 32 of the mutations are novel. This is the highest mutation detection rate reported in a study of typical NF1 patients. All mutations were studied at the genomic and RNA level. The mutational spectrum consisted of 25 nonsense, 12 frameshift, 19 splice mutations, six missense and/or small in-frame deletions, one deletion of the entire NF1 gene, and a translocation t(14;17)(q32;q11.2). Our data suggest that exons 10a-10c and 37 are mutation-rich regions and that together with some recurrent mutations they may account for almost 30% of the mutations in classical NF1 patients. We found a high frequency of unusual splice mutations outside of the AG/GT 5 cent and 3 cent splice sites. As some of these mutations form stable transcripts, it remains possible that a truncated neurofibromin is formed. PMID- 10862085 TI - A comparison of fluorescent SSCP and denaturing HPLC for high throughput mutation scanning. AB - We examined 67 different mutations in 16 different amplicons in a comparison of mutation detection by fluorescent single strand conformation polymorphism (F SSCP) and by denaturing HPLC (DHPLC). F-SSCP was used to analyze fluorescent amplicons with internal size standards and automated fragment analysis (GeneScan, PE Applied Biosystems, Foster City, CA). In DHPLC, unlabelled amplicons were analyzed by reverse phase HPLC with fragment detection by absorbance at 260nm. Both methods had high sensitivity (95-100%) and specificity (100%). Overall, F SSCP with external temperature control was the more sensitive method, but DHPLC was particularly useful for the rapid analysis of novel fragments. PMID- 10862086 TI - Higher resolution microplate array diagonal gel electrophoresis: application to a multiallelic minisatellite. AB - The 5' polymorphic region of the insulin (INS, MIM# 176730) gene contains a variable tandem repetition of 14-15 bp (a variable number of tandem repeats (VNTR) locus). After PCR amplification, we achieved precise sizing of class I alleles (range 641 to 843 bp) on 96-well open-face polyacrylamide microplate array diagonal gel electrophoresis (MADGE) gels, obtaining resolution of the 2% mobility difference which represents one tandem repeat. PCR products were run double-stranded, but no additional bands were generated except in the case of differences of three, two, and one repeat between alleles; none compromised allele identification, and in the latter case the heteroduplex was a useful confirmation signal. No end labelling of primers was required, as the sensitive Vistra Green intercalating dye for double strands was used for visualization of bands from diluted samples. Duracryl, a high mechanical-strength polyacrylamide derivative, proved to have good resolution properties for electrophoresis. A co run ladder ensured precise binning without inter-lane variability. Simultaneous electrophoresis of gels in a thermostatically controlled tank allowed up to 1,000 samples to be run in 90 min. Gels were analyzed using a FluorImager 595 fluorescent scanning system, and alleles identified using a combination of Phoretix software for band migration measurement and Microsoft Excel to compute allele sizes. Unlike other systems for minisatellite allele sizing, throughput was not limited (in time or cost) by electrophoresis. PMID- 10862087 TI - Identification of 5 novel mutations in the AGXT gene. AB - In order to identify additional genotypes in primary hyperoxaluria type 1, we sequenced the AGXT genes of 9 patients. We report 5 new mutations. Three are splice-site mutations situated at the end of intron 4 and 8 (647-1G>A, 969-1G>C, 969-3C>G), one is a missense mutation in exon 5 (D183N), and one is a short duplication in exon 2 (349ins7). Their consequence is always a lack of enzymatic activity of the Alanine-Glyoxylate Aminotransferase (AGT); for 4 of them, we were able to deduce that they were associated to the absence of AGT protein. These mutations are rare, as they have been found on one allele in our study (except 969-3C>G present in 2 unrelated families), and have not been previously reported. PMID- 10862088 TI - A novel nonsense mutation (Q509X) in three Italian late-infantile neuronal ceroid lipofuscinosis children. AB - We identified a novel nonsense CLN2 mutation (Q509X) in three Italian children with classical late-infantile neuronal ceroid lipofuscinosis (LINCL) from two unrelated families. The mutation introduced a premature stop codon in exon 12 of the CLN2 gene, resulting in a protein lacking the last 54 residues. Haplotype analysis suggested independent origin of the mutation in our families. The protein truncation test was employed to verify the functional consequences of the novel Q509X mutation. In Patient 1, the mutant alleles were transcribed but translated in a shorted peptide suggesting that the Q509X mutation is likely to interfere with CLN2p function. While expanding the list of genetic variants in LINCL, our findings represent the first molecular characterization of LINCL patients in South Europe. PMID- 10862089 TI - Identification of four single nucleotide polymorphisms in DNA repair genes: XPA and XPB (ERCC3) in Polish population. AB - A deficiency in DNA repair is associated with increased cancer risk. Inter individual variations in DNA repair capacity observed in humans may result from genetic polymorphisms in DNA repair genes. In order to provide a basis for future functional and molecular epidemiology studies on cancer susceptibility, we screened 35 individuals for polymorphisms in coding regions of XPA and XPB genes involved in nucleotide excision repair (NER). Relevant cDNA sequences were amplified by PCR, sequenced with fluorescently labeled terminators and analyzed with automated sequencer. Two polymorphisms in XPB were found: AAA-->AGA (445A>G; GenBank M31899) causing K117R substitution and GGC-->TGC (1299G>T; GenBank M31899) causing G402C exchange. Also, two polymorphisms in XPB were detected: CGA ->CAA (709G>A; GenBank D14533) causing R228Q exchange, and A-->G (23A>G; GenBank D14533) substitution in the 5' non-coding region of the gene. The three aforementioned amino acid substitutions were uncommon in this population (1.4%). In contrast, the substitution located 4 nucleotides upstream of the ATG start codon of XPB was frequent (57%). To our best knowledge this is the first report of these sequence variants. The location of these polymorphisms in evolutionary conserved regions suggest that they may be of functional significance. PMID- 10862090 TI - Fanconi anemia A due to a novel frameshift mutation in hotspot motifs: lack of FANCA protein. AB - Homozygosity for a frameshift mutation at codon 1213 of FANCA gene was identified in a Turkish patient. Immunoprecipitation-western blot analysis showed the complete absence of the FANCA protein band. This novel mutation, a deletion of T at position 3639 in exon 37 (3639delT), is responsible for the disease and causes premature termination of translation 32 aa downstream. The deletion is (i) the T residue of 2 overlapping TGAGGC and CCTG hot spot motifs, (ii) flanked by several direct repeats, (iii) surrounded by the highly GC rich region that have frequently been identified at the site of human DNA deletions. The patient is the third living child of a first degree cousin marriage. The major abnormalities of the patient at the age of 6 months were growth retardation, microcephaly, hypoplastic right thumb, distal displacements of both thumbs and pelvic displacement of left kidney. Hematological presentation of the disease started before the age of 4 years. PMID- 10862091 TI - Spectrum of COL4A5 mutations in Finnish Alport syndrome patients. AB - Alport syndrome (AS) is a hereditary kidney disorder, mainly caused by mutations in the X-chromosomal gene (COL4A5) encoding the type IV collagen a5 chain. In this study, detection of COL4A5 mutations was performed in 17 Finnish Alport syndrome families. Regions around the 51 previously known exons, as well as the two recently characterized exons 41A and 41B in COL4A5, were PCR-amplified from the patient DNA. Direct sequencing of the amplified products was performed and mutations were found in 12 families. None of the mutations involved exons 41A or 41B. Three of the mutations were potential splicing mutations, two of which were studied at the mRNA level. Seven of the mutations were single base substitutions, and two were deletions. In five families, no mutations were found. PMID- 10862092 TI - Identification of novel mutations in Spanish patients with muscle carnitine palmitoyltransferase II deficiency. AB - Carnitine palmitoyltransferase II (CPT II) deficiency is the most common recessively inherited disorder of lipid metabolism affecting skeletal muscle and the most frequent cause of hereditary myoglobinuria. We studied 5 Spanish patients with CPT II deficiency from four unrelated families. Four patients had the typical clinical phenotype of muscle CPT II deficiency with recurrent episodes of myoglobinuria, triggered by prolonged exercise, fasting, or fever, and marked elevation of creatine kinase values during metabolic crisis. One patient had exercise-related myalgia, cramps and moderate elevation of serum CK values, but had never had myoglobinuria. Molecular analysis showed that three patients were heterozygous for the S113L mutation and one patient heterozygous for the P50H substitution. To identify the mutations in the other alleles of our patients we amplified and sequenced genomic DNA fragments encompassing the entire coding region and intron/exon boundaries of the CPT2 gene. We found the recently reported 178 insT/del 25 bp in one patient. Three novel mutations were identified: a Y120C substitution that leads to a nonconservative amino acid replacement; a 36-38 insGC mutation that results in premature termination of the translation; and an I502T substitution that affects a conserved amino acid residue in the CPT II protein. Our data confirm the molecular heterogeneity of patients with CPT II deficiency, and suggest that the ethnic origin has to be taken into account before performing mutation analysis in these patients. PMID- 10862093 TI - Novel frameshift mutations in the RP2 gene and polymorphic variants. AB - Mutations in the RP2 gene located on Xp11.23 are associated with X-linked retinitis pigmentosa (XLRP), a severe form of progressive retinal degeneration which leads to complete loss of vision in affected males. To date, 14 different mutations in the RP2 gene have been reported to cause XLRP, the majority of which lead to a coding frameshift within the gene and predicted truncation of the protein product. We here report two novel frameshift mutations in RP2 identified in XLRP families by PCR-SSCP and direct sequencing, namely 723delT and 796 799del. Four single nucleotide polymorphisms (SNPs) within the coding region of RP2 are also described (105A>T, 597T>C, 844C>T, 1012G>T), the first polymorphisms to be reported within this gene of unknown function, two of which alter the amino acid sequence. The current study extends the XLRP mutation profile of RP2 and highlights non-pathogenic coding sequence variations which may facilitate both functional studies of the gene and analysis of intragenic allelic contribution to the phenotype. PMID- 10862094 TI - Survey of the coding region of the HERG gene in long QT syndrome reveals six novel mutations and an amino acid polymorphism with possible phenotypic effects. AB - Analysis of the entire coding region of the HERG gene of 39 Finnish LQTS patients revealed eight mutations, six of which are hitherto unreported. All these mutations are located in the evolutionarily conserved regions of HERG, including the transmembrane domains (P451L, Y569H, 1631delAG, G584S, G601S, T613M) and the cytoplasmic N-terminus (453delC, R176W) of the channel. Our present and earlier results suggest that the LQT2 subtype accounts for approximately 20-30% of LQTS cases in Finland. We also report the first common amino acid polymorphism (K897T) of the HERG channel, with allele frequencies of 0.84 and 0.16. Investigation of 170 genetically homogenous LQT1 patients suggests that this polymorphism may influence QT interval in female individuals. PMID- 10862095 TI - Identification of a novel mutation (c279delC) and a polymorphism (c291C>G) in the von Hippel-Lindau gene in Italian patients. PMID- 10862096 TI - A novel deletion (IVS11+3del4) identified in the human PAX6 gene in a patient with aniridia. PMID- 10862097 TI - Identification of 3 novel mutations (Y4236X, Q3820X, 11745+2 ins3) in autosomal dominant polycystic kidney disease 1 gene (PKD1). PMID- 10862098 TI - Human beta defensin 1 gene: six new variants. PMID- 10862099 TI - A novel complex mutation in exon 8 of RB1 in a case of isolated bilateral retinoblastoma. PMID- 10862100 TI - Identification of a polymorphism (D366H) in the endoglin gene in Chinese. PMID- 10862101 TI - W179R: a novel missense mutation in the peripherin/RDS gene in a family with autosomal dominant retinitis pigmentosa. PMID- 10862102 TI - Novel cardiac beta-myosin heavy chain gene missense mutations (R869C and R870C) that cause familial hypertrophic cardiomyopathy. PMID- 10862103 TI - Identification of two novel mutations (E332X and c1536delC) in the RPGR gene in two Chinese families with X-linked retinitis pigmentosa. PMID- 10862104 TI - New missense mutation (G626V) in the predicted selectivity filter of the HERG channel associated with familial long QT syndrome. PMID- 10862105 TI - Identification of a novel BLM missense mutation (2706T>C) in a Moroccan patient with Bloom's syndrome. PMID- 10862106 TI - Novel 67-bp insertional mutation in the ASS gene in a patient with citrullinemia. PMID- 10862107 TI - Snail nervous system: from classical histology to chemical and molecular neuroanatomy. PMID- 10862108 TI - Structure and function in the cerebral ganglion. AB - Evidence is reviewed to evaluate whether the term "brain is justified in referring to the snail's cerebral ganglion. The focus of the review is terrestrial species, with particular attention given to the genus Helix. In accordance with a standard definition of "brain, the cerebral ganglion is found to be differentiated both structurally and functionally. It receives convergent sensory inputs from a variety of anterior sensory organs plus the posterior body wall. Its outputs comprise motor commands directed towards anterior muscle systems, e.g., the tentacles and the penis, as well as premotor commands directed towards executory centers in other ganglia, e.g., the buccal, visceral, and pedal ganglia. Of the three major divisions in the ganglion, the procerebrum and the mesocerebrum are the most differentiated, whereas the metacerebrum is the least differentiated. The specializations of the procerebrum for olfactory functions, and the mesocerebrum for reproductive functions, reflect the importance of adaptations for feeding and mating in the evolution of the Gastropoda. PMID- 10862109 TI - Topographic organization of efferent neurons with different neurochemical characters in the cerebral ganglia of the snail Helix pomatia. AB - This study provides a description of the organization of neurons efferent to different head areas in the cerebral ganglia of Helix pomatia, revealed by simultaneous Ni-lysine and Co-lysine back-filling of different pairs of cerebral nerves. The backfills show that labeled cerebral neurons that innervate the head areas are concentrated in seven representation foci distributed in different parts of the cerebral ganglia. Almost each head area is represented in each focus. At a gross level, the representation of the different head areas in the representation foci shows a topographic arrangement. Each focus is constituted by neurochemically different groups of neurons. All head areas are innervated by serotonin-containing fibers from a single focus (Focus 2) and by dopamine containing fibers from Foci 1, 2, and 4. However, they are innervated by CARP and FMRFamide-containing fibers from all of the foci. The combination of retrograde labeling with 5, 6-dihydroxytriptamine induced pigment labeling of serotonin containing neurons or with fluorescence tyrosinehydroxylase immunocytochemistry to detect dopamine-containing neurons showed that the different head areas are topographycally represented in the clusters of both the serotonin- and dopamine containing cells. The combination of Ni-lysine backfillings from different cerebral nerves with fluorescence CARP and FMRFamide immunocytochemistry revealed that the head areas are represented also in both the CARP and FMRFamide immunoreactive groups of neurons in the different foci. PMID- 10862110 TI - Ultrastructural aspects of peptidergic modulation in the peripheral nervous system of Helix pomatia. AB - The ultrastructural characteristics of peptidergic peripheral contacts in the snail, Helix pomatia, were investigated, with special attention to the innervation of the heart, buccal mass, and salivary gland by Mytilus inhibitory peptide-immunoreactive neurons. Following the application of correlative light- and electron-microscopic pre-embedding immunocytochemistry, the peripheral tissues reveal a rich innervation by Mytilus inhibitory peptide-immunoreactive elements. These neurons establish three types of neuromuscular contacts in the heart and buccal mass: (1) close (16-20 nm) unspecialized membrane contacts; (2) contacts with a relative wide (40-100 nm) intersynaptic cleft; and (3) labeled varicosties located freely in the extracellular space, far (0. 5-several microm) from the muscle cells. In the salivary gland, the immunoractive profiles contact both the muscular and glandular elements with close (type 1) and wider (type 2) membrane attachments. The great majority of Mytilus inhibitory peptide immunoreactive profiles contain an ultrastructurally uniform population of large (120-150 nm) electron dense granules. The ultrastructural features of the innervation by Mytilus inhibitory peptide-immunoreactive elements are compared with those established by immunogold labelled FMRFamide-containing profiles in the heart and salivary gland. These latter display similarities in forming the different kinds of intercellular contacts, and differences in the morphological variability of the content of granules in the immunolabeled profiles. The results suggest diverse, non-synaptic modulatory roles of neuropeptides in the peripheral nervous system of Helix pomatia, including localized membrane effects and neurohormonal-like remote global controls, that may also be of significance in orchestrating the effects of neuropeptides released at the same time on different targets. PMID- 10862111 TI - Gene expression and function of FMRFamide-related neuropeptides in the snail Lymnaea. AB - FMRFamide and a large family of related peptides (FaRPs) have been identified in every major metazoan phylum examined, including chordates. In the pulmonate snail Lymnaea this family of neuropeptides is encoded by a five-exon locus that is subject to alternative splicing. The two alternative mRNA transcripts are expressed in the CNS in a mutually exclusive manner at the single cell level, resulting in the differential distribution of the distinct sets of FaRPs that they encode in defined neuronal networks. Biochemical peptide purification, single-cell analysis by mass spectroscopy, and immunocytochemistry have led to an understanding of the post-translational processing patterns of the two alternative precursor proteins and identified at least 12 known and novel peptides contained in neuronal networks involved in cardiorespiration, penial control and withdrawal response. The pharmacological actions of single or co expressed peptides are beginning to emerge for the cardiorespiratory network and its central and peripheral targets. Peptides derived from protein precursor 1 and contained in the heart excitatory central motoneurons E(he) have distinct functions and also act in concert in cardiac regulation, based on their unique effects on heartbeat and their differential stimulatory effects on second messenger pathways. Precursor-2 derived peptides, contained in the Visceral White Interneuron, a key neuron of the cardiorespiratory network, have mostly inhibitory effects on the VWI's central postsynaptic target neurons but with some of the peptides also exhibiting excitatory effects on the same cells. PMID- 10862112 TI - Giant identified NO-releasing neurons and comparative histochemistry of putative nitrergic systems in gastropod molluscs. AB - Gastropod molluscs provide attractive model systems for behavioral and cellular analyses of the action of nitric oxide (NO), specifically due to the presence of many relatively giant identified nitrergic neurons in their CNS. This paper reviews the data relating to the presence and distribution of NO as well as its synthetic enzyme NO synthase (NOS) in the CNS and peripheral tissues in ecologically and systematically different genera representing main groups of gastropod molluscs. Several species (Lymnaea, Pleurobranchaea, and Aplysia) have been analyzed in greater detail with respect to immunohistochemical, biochemical, biophysical, and physiological studies to further clarify the functional role of NO in these animals. PMID- 10862113 TI - Insights into early molluscan neuronal development through studies of transmitter phenotypes in embryonic pond snails. AB - Pond snails have long been the subject of intense scrutiny by researchers interested in general principles of development and also cellular and molecular neurobiology. Recent work has exploited both these fields of study by examining the ontogeny of the nervous system in these animals. Much of this work has focussed upon the development of specific transmitter phenotypes to provide vignettes of neuronal subpopulations that can be traced from early embryonic life through to adulthood. While such studies have generally confirmed previous explanations of gangliogenesis in gastropods, they have also indicated the presence of several neurons that appear earlier and in positions inconsistent with classical views of gastropods neurogenesis. The earliest of these cells contain FMRFamide-related peptides and have anteriorly projections that mark the future locations of ganglia and interconnecting pathways that will comprise the postembryonic central nervous system. These posterior, peptidergic cells, as well as certain, apical, monoaminergic neurons, disappear and apparently die near the end of embryonic life. Finally, populations of what appear to be peripheral sensory neurons begin to express catecholamines by around midway through embryonic life. Like several of the neurons expressing a variety of transmitters in the developing central ganglia, the catecholaminergic peripheral cells persist into postembryonic life. Transmitter phenotypes, cell shapes and locations, and neuritic morphologies all suggest that many of the neurons observed in early embryonic pond snails have recognizable homologues across the molluscs. Such observations have profoundly altered our views of neurogenesis in gastropods over the last few years. They also suggest the promise for pond snails as fruitful models for studying the roles and mechanisms for pioneering fibres, cues triggering apoptosis, and contrasting origins and mechanisms employed for generating central vs. peripheral neurons within a single organism. PMID- 10862114 TI - Regeneration as an application of gastropod neural plasticity. AB - Gastropod research is providing many insights into mechanisms of neural regeneration. These observations were made possible by the pioneering work of individuals who described the nervous systems of gastropods, mapped prominent neurons and determined their roles and connections, and developed the techniques for culturing them. This information has allowed questions about injury responses, target selection, and pathway cues to be explored at the level of individually identified neurons. Because of gastropod studies, more is known about axon sealing, growth cone formation and behavior, signals that travel from the site of axotomy to the soma, and the second messengers that are activated there. The responses in neurons and non-neuronal cells during neural development and injury are coordinated by chemical messenger systems that are highly conserved, including neurotransmitters, cytokines, and neurotrophins. The nervous system is modified in learning paradigms by some of the same messenger systems activated by injury, because learning and injury both challenge neurons to change. The conservation of basic mechanisms that coordinate neuronal plasticity allows us to approach basic questions in relatively simple nervous systems with reasonable confidence that the findings will be relevant for other nervous systems, including possible applications to the mammalian nervous system. PMID- 10862115 TI - Immunohistochemical myofiber typing and high-resolution myofibrillar lesion detection in LR white embedded muscle. AB - We have developed a method of fixing, embedding, sectioning, and staining that allows high-resolution detection of myofibrillar structure and myosin immunocytochemical muscle fiber typing in serial semithin sections of LR White plastic embedded muscle at the light microscopic level. Traditional approaches, such as cryostat sections, permit fiber typing, but small myofibrillar lesions (1 3 sarcomeres) are difficult to detect because of section thickness. Semithin sections of hydrophobic resins do not stain well either histochemically or immunocytochemically. Electron microscopy can resolve lesions and discriminate fiber types based on morphology, but the sampling area is small. Our goal was to develop a rapid method for defining both fiber type and high-resolution primary myofibrillar lesion damage. Mild fixation (1-4% paraformaldehyde, 0. 05-0.1% glutaraldehyde) and embedment in a hydrophilic resin (LR White) were used. Myofibrillar structure was extremely well preserved at the light microscopic (LM) level, and lesions could be readily resolved in Toluidine blue stained 500-nm sections. Fiber type was defined by LM immunomyosin staining of serial plastic semithin sections, which demonstrated reciprocal staining patterns for "fast (Sigma M4276) and "total" (skeletal muscle) myosins (Sigma M7523). PMID- 10862116 TI - Improvement of light collection efficiency of lens-coupled YAG screen TV system for a high-voltage electron microscope. AB - A new lens coupling television (TV) system using a YAG (Yttrium Aluminum Garnet: Y(3)Al(5)O(12) : Ce(3+)) single crystal screen has been developed for a high voltage electron microscope (HVEM), and its performance is examined. The system, using a combination of YAG and lenses, is less damaged by high-energy electron irradiation and reduces the influence of X-rays on the image. YAG screens have not been used for lens-coupling systems, because the high refractive index (n = 1.84) of YAG results in a low light collection efficiency for emitted light. This disadvantage is overcome by combining a thin YAG disk screen (thickness; 100 microm) with a glass hemisphere whose refractive index is 1.81. We found that the light intensity is almost the same as that obtained with a conventional P22 powder screen and lenses system. The resolution is about 55 microm on the YAG screen, and this value is 1.3 times higher than that measured by the conventional system. Shading and distortion do not affect TV observation. Detection quantum efficiency, obtained after correction of the channel mixing effect, is about 0.1. PMID- 10862117 TI - Miniature mass analyzers AB - Increased efforts are being made to develop miniature mass spectrometers, including those which are hand-portable, and to retain the performance characteristics of traditional laboratory instruments as much as possible in the miniature instruments. This review of miniature mass analyzers emphasizes analytical performance and compares the relative merits of each type of miniature mass analyzer. Miniature instruments discussed include sector, Wien filter, time of-flight, linear quadrupole, quadrupole ion trap and Fourier transform ion cyclotron resonance mass spectrometers, as well as combinations of and variations on these major types. Special considerations that apply to small mass analyzers are noted and suggestions are made regarding the possible future development of this field. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10862119 TI - Reaction competition in the fragmentation of protonated dipeptides AB - A major low-energy fragmentation reaction of many protonated dipeptides involves cleavage of the amide bond resulting in formation of either the y(1)" ion or the a(1) ion. For a series of protonated dipeptides H-Val-Xxx-OH it is observed that log(y(1)"/a(1)) is a linear function of the proton affinity of the variable C terminal amino acid. For the series of protonated dipeptides H-Xxx-Phe-OH log(a(1)/y(1)") gives a poor correlation with the proton affinity or gas-phase basicity of H-Xxx-OH. However, a good limited correlation of log(a(1)/y(1)") with the Taft-Topsom sigma(alpha) for the alkyl group is observed when Xxx is an aliphatic amino acid. It is proposed that fragmentation occurs by initial formation of a proton-bound complex of an aziridinone and an amino acid which may fragment to form either a protonated amino acid (y(1)") or an N-protonated aziridinone with the corresponding neutrals being an aziridinone and an amino acid. Ab initio calculations show that the N-protonated aziridinone is unstable and fragments by loss of CO to form the a(1) immonium ion. However, the proton bound complex of an aziridinone and an amine base is a stable species which exists in a potential well. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10862118 TI - Novel procedure for the identification of proteins by mass fingerprinting combining two-dimensional electrophoresis with fluorescent SYPRO red staining. AB - The fluorescent sensitive SYPRO Red dye was successfully employed to stain proteins in two-dimensional gels for protein identification by peptide mass fingerprinting. Proteins which are not chemically modified during the SYPRO Red staining process are well digested enzymatically in the gel and hence the resulting peptides can be efficiently eluted and analysed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A SYPRO Red two-dimensional gel of a complex protein extract from Candida albicans was analysed by MALDI-TOF MS. The validity of SYPRO Red staining was demonstrated by identifying, via peptide mass fingerprinting, 10 different C. albicans proteins from a total of 31 selected protein spots. The peptide mass signal intensity, the number of matched peptides and the percentage of coverage of protein sequences from SYPRO Red-stained proteins were similar to or greater than those obtained in parallel with the modified silver protein gel staining. This work demonstrates that fluorescent SYPRO Red staining is compatible with the identification of proteins separated on polyacrylamide gel and that it can be used as an alternative to silver staining. As far as we know, this is the first report in which C. albicans proteins separated using 2-D gels have been identified by peptide mass fingerprinting. The improved technique described here should be very useful for carrying out proteomic studies. PMID- 10862120 TI - Mass spectrometric analysis of hydroxyproline glycans. AB - Mass spectrometric techniques are presented which allow one to analyze the sugar part bound to hydroxyproline in hydroxyproline-rich glycoproteins. The hydroxyproline (Hyp) glycans obtained by alkaline hydrolysis give abundant [M + Na](+) ions by electrospray ionization which after collision-induced dissociation (CID) yield inter alia [Hyp - H + Na](+). In mixtures a parent ion scan of this species will indicate the various molecular species which can then be analyzed by MS(n) after CID in an ion trap, where successive losses of the sugar units are observed. Methylation techniques allow one to distinguish between linear and branched isomeric structures. PMID- 10862121 TI - Gas chromatography/negative ion chemical ionization mass spectrometry and liquid chromatography/electrospray ionization tandem mass spectrometry quantitative profiling of N-acetylcysteine conjugates of valproic acid in urine: application in drug metabolism studies in humans. AB - We report a GC/NICI-MS assay and a LC/ESI-MS/MS assay for the analysis of N acetylcysteine (NAC) conjugates of (E)-2,4-diene VPA (NAC I and NAC II) identified in humans. The assay also includes the analysis of the NAC conjugate of 4,5-epoxy VPA (NAC III), an identified metabolite in rats treated with 4-ene VPA for its use in metabolic studies in animals. The highly sensitive GC/MS assay was designed to monitor selectively the diagnostic and most abundant [M - 181](-) fragment anion of the di-PFB derivatives of NAC I, NAC II, and NAC IV, the internal standard (IS) and the PFB derivative of NAC III. The higher selectivity of LC/MS/MS methodology was the basis for an assay which could identify and quantitate the underivatized conjugates simultaneously using MRM of the diagnostic ions m/z 130 and 123 arising from the CID of their protonated molecular ions [MH](+). The GC/MS assay employed liquid-liquid extraction whereas the LC/MS/MS assay used a solid-phase extraction procedure. Linearity ranges of the calibration curves were 0.10-5.0microg ml(-1) by GC/MS and 0.10-1.0microg ml( 1) by LC/MS/MS for NAC I, NAC II and NAC III (r(2) = 0.999 or better). Both assays were validated for NAC I and NAC II and provided good inter- and intra assay precision and accuracy for NAC I and NAC II. The LOQ by LC/MS/MS was 0.1microg ml(-1), representing 1 ng of NAC I and NAC II. The same LOQ (0.1microg ml(-1)) was observed by GC/MS and was equivalent to 100 pg of each metabolite. NAC III was detected at concentrations as low as 0.01 microg ml(-1) by both methods. The total urinary excretion of the NAC conjugates in four patients on VPA therapy was determined to be 0.004-0.088% of a VPA dose by GC/MS and 0.004-0. 109% of a VPA dose by LC/MS/MS. PMID- 10862122 TI - Regioisomeric differentiation of 2,3- and 3,4-methylenedioxy ring-substituted phenylalkylamines by gas chromatography/tandem mass spectrometry AB - Numerous abused drugs of the 3,4-methylenedioxymetamphetamine (MDMA; Ecstasy; N methyl-1-(3,4-methylenedioxyphenyl)-2-propaneamine) type and various alkyl chain- and aromatic ring-substituted isomers give very similar electron ionization (EI) mass spectra. This seriously affects the analysis of especially ring regioisomeric drug variants. Using collision-induced dissociation (CID) (argon) under EI and chemical ionization, the mass spectra of 18 2,3- and 3, 4 methylenedioxy ring-substituted phenylethylamines were recorded. These techniques permitted an unequivocal differentiation of all studied ring regioisomeric methylenedioxyphenylethylamines. CID mass spectrometry therefore appear to be a reliable tool to establish the kind of ring substitution pattern in regioisomeric methylenedioxyphenalkylamines. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10862123 TI - New results on the mass spectra of cinchona alkaloids AB - The electrospray ionization (ESI) mass spectra of 16 cinchona alkaloid compounds were studied for the first time. The electron ionization (EI) spectra of 22 cinchona alkaloids were also recorded, 14 of which had not been examined previously. In the case of EI the characteristic direction of the fragmentation is the scission of the C8-C9 bond. Under EI the cleavage of the C4'-C9 bond occurs only in the case of hydrogenated cinchona alkaloids, whereas the C9-O bond cleavage can be observed in the case of ester and ether derivatives. At a low capillary exit voltage (CapEx) in the ESI measurements there is no fragmentation, and only the [M + H](+) and in some cases the double protonated [M + 2H](2+) ions can be detected. On increasing the CapEx the characteristic primary direction is the cleavage of the C9-O bond, which was observed in the case of epialkaloids and esterified or etherified cinchona derivatives, respectively. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10862124 TI - Simultaneous determination of risperidone and 9-hydroxyrisperidone in plasma by liquid chromatography/electrospray tandem mass spectrometry. AB - A simple and highly sensitive liquid chromatographic/electrospray tandem mass spectrometric (LC/MS/MS) assay was developed for the simultaneous determination of risperidone (RSP) and its major circulating metabolite 9-hydroxyrisperidone (9 OH-RSP) in the plasma of humans and rats. A simple one-step solvent extraction with 15% methylene chloride in pentane was used to isolate the compounds from plasma. The compounds were eluted from a phenyl-hexyl column and detected with a Perkin-Elmer SCIEX API2000 triple-quadrupole mass spectrometer using positive ion atmospheric pressure electrospray ionization and multiple reaction monitoring. The assay was linear over the range 0.1-100 ng ml(-1) when 0.5 ml of plasma was used in the extraction. The overall intra- (within-day) and inter- (between days) assay variations were < 11%. The variations in the concentrations of two long term quality control samples from pooled patient plasma samples analyzed over a period of 6 months were approximately 10%. The analysis time for each sample was 4 min and more than 100 samples could be analyzed in one day by running the system overnight. The assay is simple, highly sensitive, selective, precise and fast. This method is being used for the therapeutic drug monitoring of schizophrenic patients treated with RSP and to study the pharmacokinetics and tissue distribution of RSP and 9-OH-RSP in rats. PMID- 10862125 TI - Matrix-assisted laser desorption/ionization mass spectrometric quantification of the mu opioid receptor agonist DAMGO in ovine plasma. AB - The synthetic opioid peptide analog Tyr-D-Ala-Gly-N-methyl-Phe-Gly-ol (DAMGO), which is a mu opioid receptor-selective agonist, was quantified in ovine plasma samples with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), using delayed extraction and a reflectron. The internal standard was pentadeuterated DAMGO. Timed-ion selection was used to select the precursor ion. The analysis of the post-source decay fragments improved the detection sensitivity, and the use of the precursor-product ion relationship optimized the specificity. For plasma samples, the inter-assay variability of this method was 6.4% (n = 79) and the intra-assay variability was 6.0% (n = 10). The variability for controls was 3.4% (n = 43). The profile of DAMGO amount versus time was determined in sheep plasma, and the corresponding pharmacokinetic data were calculated. PMID- 10862127 TI - Structural analysis of synthetic homo- and copolyesters by electrospray ionization on a fourier transform ion cyclotron resonance mass spectrometer AB - The molecular structure of a series of homo- and copolyesters was studied using sustained off-resonance irradiation collisionally activation dissociation on a Fourier transform ion cyclotron resonance mass spectrometer. Electrospray ionization was used as an ionization technique. The most important fragmentation pathways of the homopolyesters poly(dipropoxylated bisphenol-A/adipic acid) and poly(dipropoxylated bisphenol-A/isophthalic acid) were studied. Six different dissociation mechanisms were observed which are very similar to the mechanisms found to occur during pyrolysis of these compounds. Four of these mechanisms are a result of cleavages of the ester bond and the others are due to cleavages of the ether bond or bisphenol-A unit. Some of the fragments expected are not present in the spectrum, indicating that each fragment has a specific sodium affinity. Sequence-specific fragments of two of the three copolyester sequences that theoretically can exist were experimentally observed. Fragments that originate from the third sequence are not unique and can also be formed from other sequences. Therefore, it was not possible to determine the presence of the third sequence. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10862126 TI - Use of stable isotope labeling technique and mass isotopomer distribution analysis of [(13)C]palmitate isolated from surfactant disaturated phospholipids to study surfactant in vivo kinetics in a premature infant. AB - Pulmonary surfactant is a complex mixture of phospholipids and proteins which lowers surface tension and maintains alveolar expansion at end expiration. Developmental and genetic disruption of pulmonary surfactant metabolism leads to respiratory distress in newborns. Stable isotope labeling of metabolic precursors of disaturated phospholipids, the most abundant and specific component of pulmonary surfactant, permits the measurement of the kinetics of surfactant metabolism in vivo. We measured [U-(13)C(6)]glucose incorporation into palmitic acid derived from disaturated surfactant phospholipids. A 24 h infusion of [U (13)C(6)]glucose (140 mg kg(-1)) was administered to a premature infant who required mechanical ventilation for respiratory distress syndrome; tracheal aspirate samples were obtained at the start of the infusion and at regular intervals for the next 70 h. Each tracheal aspirate sample was incubated with osmium tetroxide to isolate disaturated surfactant phospholipids. Methyl esters of the fatty acids in the disaturated phospholipids were prepared and the enrichment of [(13)C]methyl palmitate was measured by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combination/isotope ratio mass spectrometry (GC/C/IRMS). Mass isotopomer distribution analysis (MIDA) was used to calculate the fractional synthetic rate (FSR) of palmitate synthesized from acetate. With both GC/MS and GC/C/IRMS, palmitate (13)C enrichment was first detected 12.3 h after the start of the tracer infusion. The enrichment increased in a linear fashion, reached a peak at 47 h and remained constant in the remainder of the samples. The FSR of palmitate from acetate was 5.2% per day. Stable isotope techniques and MIDA will provide insights into the kinetics of surfactant metabolism in newborns with respiratory dysfunction. PMID- 10862128 TI - Current awareness AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of mass spectrometry. Each bibliography is divided into 11 sections: 1 Books, Reviews & Symposia; 2 Instrumental Techniques & Methods; 3 Gas Phase Ion Chemistry; 4 Biology/Biochemistry: Amino Acids, Peptides & Proteins; Carbohydrates; Lipids; Nucleic Acids; 5 Pharmacology/Toxicology; 6 Natural Products; 7 Analysis of Organic Compounds; 8 Analysis of Inorganics/Organometallics; 9 Surface Analysis; 10 Environmental Analysis; 11 Elemental Analysis. Within each section, articles are listed in alphabetical order with respect to author (5 Weeks journals - Search completed at 22nd. Mar 2000) PMID- 10862129 TI - Introduction PMID- 10862130 TI - Simulation-based medical planning for cardiovascular disease: visualization system foundations. AB - A technique for visualizing computational models along with volumetric imaging data in a real-time, interactive, simulation-based medical planning system for cardiovascular disease treatment is described. This technique involves an ordered rendering of faceted geometry and volumetric image data. We have developed a software system based on this image-fusion technique that is capable of capturing and representing the inherent anatomic constraints of an individual patient. Such constraints must be represented accurately in a medical planning system to ensure the validity of a potential procedure. A hypothetical clinical scenario is described for which vascular treatment plans were constructed pre-operatively without reference to the physical anatomic structure. These models were later embedded into patient-specific diagnostic MRA scans to establish the anatomic context for physiologic observations. PMID- 10862131 TI - Development and application of a virtual environment for reconstructive surgery. AB - OBJECTIVE: This paper details the development and application of a Virtual Environment for Reconstructive Surgery (VERS). It addresses the technical and user-interface challenges in developing such a system, and the lessons learned during application of the system in the case of a 17-year-old boy with a severe facial defect arising from the removal of a soft-tissue sarcoma. MATERIALS AND METHODS: Computed tomography (CT) scans were segmented into bone and soft-tissue classifications using traditional and novel algorithms, a surface mesh was generated, and imaging artifacts were removed, yielding a mesh suitable for visualization. This patient-specific mesh was then used in a virtual environment by the surgeons for preoperative visualization of the defect, planning of the surgery, and production of a custom surgical template to aid in repairing the defect. RESULTS: This system was successfully used to plan the surgery of the patient and to produce a custom, patient-specific template that was used to harvest bone from a donor site in order to reconstruct the defect. CONCLUSION: Despite technical challenges, virtual-environment surgical planning is useful as a clinical tool for preoperative visualization, cephalometric analysis, and surgical intervention. It can provide a more precise surgical result than would otherwise be realized using traditional methods. PMID- 10862132 TI - Comparative tracking error analysis of five different optical tracking systems. AB - OBJECTIVE: Effective utilization of an optical tracking system for image-based surgical guidance requires optimal placement of the dynamic reference frame (DRF) with respect to the tracking camera. Unlike other studies that measure the overall accuracy of a particular navigation system, this study investigates the precision of one component of the navigation system: the optical tracking system (OTS). The precision of OTS measurements is quantified as jitter. By measuring jitter, one can better understand how system inaccuracies depend on the position of the DRF with respect to the camera. MATERIALS AND METHODS: Both FlashPointtrade mark (Image Guided Technologies, Inc., Boulder, Colorado) and Polaristrade mark (Northern Digital Inc., Ontario, Canada) optical tracking systems were tested in five different camera and DRF configurations. A linear testing apparatus with a software interface was designed to facilitate data collection. Jitter measurements were collected over a single quadrant within the camera viewing volume, as symmetry was assumed about the horizontal and vertical axes. RESULTS: Excluding the highest 5% of jitter, the FlashPoint cameras had an RMS jitter range of 0.028 +/- 0.012 mm for the 300 mm model, 0.051 +/- 0.038 mm for the 580 mm model, and 0.059 +/- 0.047 mm for the 1 m model. The Polaris camera had an RMS jitter range of 0.058 +/- 0.037 mm with an active DRF and 0.115 +/- 0.075 mm with a passive DRF. CONCLUSION: Both FlashPoint and Polaris have jitter less than 0.11 mm, although the error distributions differ significantly. Total jitter for all systems is dominated by the component measured in the axis directed away from the camera. PMID- 10862133 TI - Accuracy of muscle moment arms estimated from MRI-based musculoskeletal models of the lower extremity. AB - OBJECTIVE: Biomechanical models that compute the lengths and moment arms of soft tissues are broadly applicable to the treatment of movement abnormalities and the planning of orthopaedic surgical procedures. The goals of this study were to: (i) develop methods to construct subject-specific biomechanical models from magnetic resonance (MR) images, (ii) create models of three lower-extremity cadaveric specimens, and (iii) quantify the accuracy of muscle-tendon lengths and moment arms estimated using these models. MATERIALS AND METHODS: Models describing the paths of the medial hamstrings and psoas muscles for a wide range of body positions were developed from MR images in one joint configuration by defining kinematic models of the hip and knee, and by specifying "wrapping surfaces" that simulate interactions between the muscles and underlying structures. Our methods for constructing these models were evaluated by comparing hip and knee flexion moment arms estimated from models of three specimens to the moment arms determined experimentally on the same specimens. Because a muscle's moment arm determines its change in length with joint rotation, these comparisons also tested the accuracy with which the models could estimate muscle-tendon lengths over a range of hip and knee motions. RESULTS: Errors in the moment arms calculated with the models, averaged over functional ranges of hip and knee flexion, were less than 4 mm (within 10% of experimental values). CONCLUSION: The combination of MR imaging and graphics-based musculoskeletal modeling provides an accurate and efficient means of estimating muscle-tendon lengths and moment arms in vivo. PMID- 10862134 TI - Computer-assisted training and learning in surgery. AB - The teaching and learning of surgery is a time-honored tradition based upon the "see one, do one, teach one" apprenticeship model. Recent improvement of this model has centered upon incremental change in skills teaching and testing and curricular development. Economic pressures have strained the resources of academic health centers and faculty responsible for teaching surgery, even as information technology has opened new avenues for obtaining and benefitting from relevant information. Combining the tools of simulation theory, virtual reality, and the principles of adult education offers new opportunities to optimize surgical education as we enter a more highly connected and interdependent era, where the boundaries between teacher and student blur as the modern surgeon truly becomes a lifelong learner. PMID- 10862135 TI - Abstract: molecular imaging of vascular receptors: implications in the design of anti-angiogenic therapeutics PMID- 10862136 TI - Abstract: MRI: basic principles and future potential PMID- 10862137 TI - Abstract: real-time interactive MRI for cardiac applications PMID- 10862138 TI - Abstract: cost-effectiveness analysis of new image-based screening technologies PMID- 10862139 TI - Abstract: alternative visualization and analysis of volumetric data PMID- 10862140 TI - A flow injection chemiluminescence method using Cr(III) as a catalyst for determining hydrogen peroxide. Application to H(2)O(2) determination in cultures of microalgae. AB - Flow injection analysis has been applied to the determination of hydrogen peroxide produced by some different species of microalgae. The method is based on the luminol-H(2)O(2) chemiluminescence reaction using Cr(III) as a catalyst. Optimum experimental conditions for the method have been studied and trace amounts of hydrogen peroxide determined with detection limits of 4 10(-8) mol/L. The method using Cr(III) was compared with that using horseradish peroxidase as the catalyst. PMID- 10862141 TI - Determination of hydrogen peroxide by micro-flow injection-chemiluminescence using a coupled flow cell reactor chemiluminometer. AB - A novel flow cell reactor was developed for micro-flow injection determination of hydrogen peroxide (H(2)O(2)) using horseradish peroxide (HRP)-catalysed luminol chemiluminescence. The newly developed flow cell reactor for a chemiluminometer allowed mixing of the chemiluminescent reagents in front of a photomultiplier for maximum detection of the emitted light. The rapid mixing allowed a decrease in the flow rate of the pump to 0.1-0.01 mL/min, resulting in increased sensitivity of detection of light. The flow cell reactor was made by packing HRP-immobilized gels into a flow cell (Teflon tube; 6 cm x 0.98 mm i.d.) located in the cell holder of a chemiluminometer (flow-through type). The HRP-immobilized gels were made by immobilizing HRP onto the Chitopearl gel by the periodate method. H(2)O(2) specimens (50 microL) were injected into a stream of water delivered at a flow rate of 0.1 mL/min and mixed with a luminol solution (0.56 mmol/L in Tricine buffer, pH 9.2) delivered at 0.1 mL/min in the flow cell reactor. Within run reproducibility of the assay of H(2)O(2) was 2.4% (4.85 micromol/L; flow rate 0.1 mL/min, injection interval 10 min). The reproducibility of the H(2)O(2) assay was influenced by the flow rates and the injection intervals of the H(2)O(2) specimens. As the flow rates decreased, both the light intensity and the light duration increased. Optimal light intensity was obtained at a luminol concentration of 3-8 mmol/L, but 0.56 mmol/L was sufficient for assay of H(2)O(2) in clinical specimens. At a luminol concentration of 0.56 mmol/L, the regression equation of the standard curve for H(2)O(2) (0-9.7 micromol/L) was Y = 27.5 X(2) + 394 X + 58.9 (Y = light intensity; X = concentration of H(2)O(2)) and the detection limit of H(2)O(2) was 0.2 micromol/L. This method was used to assay glucose (2.7-16.7 mmol/L) based on a glucose oxidase (20 U/mL, pH 7.4) reaction. The standard curve for glucose was Y = 167 X(2) - 351 X + 1484 (Y = light intensity; X = glucose). The within-run reproducibility for an aqueous glucose standard (2.7 mmol/L) and a control serum (glucose, 5 mmol/L) was 4.48% (n = 5) and 5.70% (n = 9), respectively. PMID- 10862142 TI - Functional states of polymorphonuclear leukocytes determined by chemiluminescent kinetic analysis. AB - During the respiratory burst, upon stimulation with both soluble and particulate matter, polymorphonuclear leukocytes (PMN) generate reactive oxygen species (ROS) and emit chemiluminescence (CL) as a result of metabolic activation. The measurement of CL has been demonstrated to be a useful tool for in vitro assessment of the opsonophagocytic function of PMN. Using component analysis of CL kinetics, we characterized the functional state of PMN by three parameters of the respiratory burst: capacity, effectiveness and velocity (CEV space). The possibility of delimiting eight different functional states of PMN is discussed. The CL kinetics shown by blood PMN in different functional states was analysed, and revealed six out of eight functional states. We conclude that CEV-estimated functional states of PMN are relative, depending on both PMN readiness to generate ROS and conditions of the CL test. PMID- 10862143 TI - Activation of peripheral phagocytes in BCG-vaccinated subjects. AB - The role of polymorphonuclear leukocytes (PMNs) in the immune defence against intracellular bacteria has long been neglected. Only recently have studies begun to address this issue. In this study the behavior of peripheral PMNs in Bacillus Calmette-Guerin (BCG) vaccinated subjects was investigated. Twenty healthy and purified protein derivative-negative adults were studied before, and two and four months after, BCG administration. Luminol-amplified chemiluminescence (CL) emission was evaluated in whole blood phagocytes using a soluble stimulus, such as phorbol mirystate acetate, or particulates such as zymosan opsonized with homologous (OZH) or autologous (OZA) serum. Specific IgG, IgA and IgM against antigen -60 by ELISA, total immunoglobulin, C3 and C4 components of complement, were assessed by immunochemical tests. The results revealed a late heightened production of reactive oxygen intermediates in vaccinated subjects in presence of OZA and OZH. Our findings confirm that the role of PMNs and their mediators in immunoregulation of intracellular diseases needs to be re-evaluated. PMID- 10862144 TI - Involvement of cyclic nucleotides and IP(3) in the regulation of luminescence in the brittlestar Amphipholis squamata (Echinodermata). AB - We investigated the effects of cyclic nucleotides (cGMP, cAMP) and the phosphoinositide IP(3) on the luminescence of the ophiuroid Amphipholis squamata. The cGMP analogue, dibutyryl-cGMP, and the guanylate cyclase activator, sodium nitroprusside, had no effect on the luminescence. The cAMP analogue, dibutyryl cAMP, and the adenylate cyclase activator, forskolin, triggered luminescence. Moreover, the adenylate cyclase inhibitor, MDL-12330A, significantly reduced ACh induced luminescence. The phospholipase C inhibitor, U-73122, also significantly reduced ACh-induced luminescence. The results suggest that ACh-induced luminescence is mediated by both cAMP and IP(3) pathways but not by cGMP. The effects of calcium-free ASW confirmed this hypothesis. A hypothetical scheme of the transduction mechanisms involved in the intracellular control of luminescence is presented. PMID- 10862145 TI - Effect of aqueous extract of cigarette smoke on peripheral blood polymorphonuclear leukocytes chemiluminescence. AB - Cigarette smoke induces a vast cohort of deleterious effects on biological structures. In the present paper, the effect of aqueous extract of cigarette smoke on the activity of polymorphonuclear leukocytes was studied. Although the aqueous extract of cigarette smoke inhibits the luminol oxidation catalysed by horseradish peroxidase, it strongly interacts with polymorphonuclear leukocytes and inhibits their phorbol-induced chemiluminescence in the presence of either luminol or lucigenin. The results indicate that at least some of the components of the aqueous extract of cigarette smoke may strongly interfere with polymorphonuclear cells, contributing to the deleterious effects of smoke products. PMID- 10862146 TI - Chemiluminescent determination of hydrogen peroxide with 9 acridinecarbonylimidazole and use in measurement of glucose oxidase and alkaline phosphatase activity. AB - Several activated derivatives of 9-acridinecarboxylic acid were prepared in order to investigate their utility for detection of hydrogen peroxide. One of these derivatives, 9-acridinecarbonylimidazole (I), is especially stable and is a useful reagent for measuring, by chemiluminescence, the activity of a number of enzymes that directly produce peroxide, including glucose oxidase. Other enzymes can also be assayed if an appropriate intermediate substrate exists that ultimately produces hydrogen peroxide after being acted upon by the enzyme. 5 Bromo-4-chloro-3-indolyl phosphate (BCIP) and 3-indolyl phosphate (3-IP) are such substrates for alkaline phosphatase. The detection limits for both of these enzymes are in the 1-10 amol range. Other enzymes that can potentially be assayed using I include oxidases, hydrolases and dehydrogenases. Negative assays for compounds that consume or bind peroxide such as reducing agents, antioxidants, catalases and peroxidases are also feasible. PMID- 10862147 TI - Chemiluminescence and reactive oxygen scavenging activities of the hydrogen peroxide/gallic acid/lactoperoxidase system. AB - Photon emission in the hydrogen peroxide/gallic acid/lactoperoxidase system was studied using visible region chemiluminescence. The photon emission intensity showed a pH-dependence curve with a maximum at pH 4.5-5.0. The spectral analysis showed to be at 510 nm and its energy was 61.0 kcal/mol at either pH 4.5 or pH 7.0. In addition, near-infrared spectral analysis at 1270 nm suggested that this photon emission from the hydrogen peroxide/gallic acid/lactoperoxidase system was produced without generation of (1)O(2). The gallic acid/lactoperoxidase system, based on the chemiluminescence system, has superoxide-, hydroxyl radical scavenging activities and antioxidative activity. These results are strong evidence that the gallic acid/lactoperoxidase system is one of the reactive oxygen scavenging systems. PMID- 10862149 TI - Characterization of an amphioxus wnt gene, AmphiWnt11, with possible roles in myogenesis and tail outgrowth. AB - The full-length sequence and developmental expression of an amphioxus Wnt gene (AmphiWnt11) are described. A phylogenetic analysis of all known full-length Wnt11 sequences indicates that a gene duplication occurred at the base of the vertebrate Wnt11 clade. The developmental expression domains of AmphiWnt11 resemble those of Wnt11 homologs in vertebrates. The earliest detectable expression is transiently associated with the dorsal lip of the blastopore. At the neurula stage, AmphiWnt11 is expressed in myotomal muscle cells; however, AmphiWnt11 transcription is not associated with metameric pre-patterning prior to morphological segmentation. Finally, in amphioxus and the vertebrates, Wnt11 homologs are expressed in anteroventral ectoderm and in association with the tailbud and the tail fin. Thus, in amphioxus and lower vertebrates, the posterior expression of Wnt11 may be involved in tail fin outgrowth, and this ancient genetic program might have been co-opted at least in part for lateral appendage development during vertebrate evolution. genesis 27:1-5, 2000. PMID- 10862148 TI - Comparative studies of the chemiluminescent horseradish peroxidase-catalysed peroxidation of acridan (GZ-11) and luminol reactions: effect of pH and scavengers of reactive oxygen species on the light intensity of these systems. AB - In this study, the chemiluminescent horseradish peroxidase/H(2)O(2)-catalysed oxidation of acridan (GZ-11) substrate was compared with the well-characterized light-producing luminol reaction. p-Iodophenol and p-phenylphenol were used as enhancers, respectively, for the luminol and acridan reactions. These two light producing systems showed significant differences in relation to the effect of pH, as well as the effect of scavengers of reactive oxygen species, on the light intensity. Light production measured could be as low as pH 2.6 in the acridan reaction, whereas light emission was not detected in the luminol system below pH 5.6. In contrast with the luminol system, it was found that superoxide dismutase does not inhibit the light intensity of the acridan system. This suggests that superoxide anion does not participate in the mechanism of the light-emitting steps of the acridan reaction. Addition of hydroxyl radical scavengers, mannitol and benzoate, to the acridan reaction medium had no appreciable effect on the chemiluminescent intensity, indicating that hydroxyl radicals do not interfere in light-emitting steps. In addition, the peroxidation of the acridan substrate was found to be very slow at pH 5.6 in the absence of the enhancer, p-phenylphenol, whereas in its presence a rapid degradation of the acridan substrate was observed. Therefore, it is suggested that the enhancer might be initially oxidized by the HRP/H(2)O(2) system, resulting in the formation of the enhancer radical, which could be the actual oxidizing agent of the acridan substrate. Together, the data presented in this paper indicate that the chemiluminescent horseradish peroxidase-catalysed peroxidation of acridan (GZ-11) is more specific than the luminol reaction for the reactive oxygen species involved in the light emitting steps, i. e, H(2)O(2). PMID- 10862150 TI - Evidence for the differential regulation of Nkx-6.1 expression in the ventral spinal cord and foregut by Shh-dependent and -independent mechanisms. AB - The Nkx-6.1 homeodomain transcription factor was previously shown to be expressed in ventral neural progenitor cells and subsequently subsets of unidentified motor neurons during early neural development. In this study, we identify a specific subpopulation of motor neurons, the median half of the lateral motor neuron column (LMCm), that retain a strong expression of Nkx-6.1. In addition, we report novel patterns of Nkx-6.1 expression in several mesenchymal tissues surrounding Sonic hedgehog (Shh)-expressing cells, including ventral spinal meninges, esophageal mesenchyme, and dorsal tracheal mesenchyme. Whereas Shh signaling is required for Nkx-6.1 expression in the ventral neural tube and spinal meninges, an Shh-independent pathway appears to operate in regulating Nkx-6.1 expression in the foregut. The persistent and robust expression of Nkx-6.1 in motor neurons and mesenchymal cells suggests an important role for Nkx-6.1 in controlling cell fate specification and differentiation. genesis 27:6-11, 2000. PMID- 10862151 TI - Requirement of LIM domains for LIM1 function in mouse head development. AB - Lim1, also known as Lhx1, encodes a LIM homeodomain transcription factor that is essential for head development in the mouse. As with other LIM homeodomain proteins, LIM1 has two LIM domains located N-terminal to the homeodomain, with each LIM domain containing two zinc finger motifs. LIM domains can physically interact with other proteins to form protein complexes that regulate transcription. Previous studies have suggested that LIM domains negatively regulate the transcriptional activity of their associated homeodomains. To investigate the requirement of LIM domains for LIM1 activity, we have mutated the Lim1 gene to alter the conserved amino acid residues that are required for zinc finger structure within both of the LIM domains. Although mice homozygous for this Lim1 allele express the mutant mRNA and protein appropriately, they are a phenocopy for Lim1-null mice. These results suggest that the integrity of the LIM domains is essential for LIM1 activity in mouse head development. genesis 27:12 21, 2000. PMID- 10862153 TI - Retinal axon misrouting at the optic chiasm in mice with neural tube closure defects. AB - In a new mouse mutant, circletail (Crc), failure of neural tube closure (embryonic day [E] 8-9) is associated with errors in retinal axon projection at the optic chiasm (E12-18), such that many axons normally projecting contralaterally instead grow to ipsilateral targets. Although the architecture of the chiasmatic region is altered, neurons and glia containing putative cues for axon guidance are present. The aberrant ipsilateral-projecting cells originate from a nonrandom expansion of the wild-type uncrossed retinal region. These axon pathway defects are found in two other mutants with cephalic neural tube defects (NTD), loop-tail (Lp) and Pax3 (splotch; Sp(2H)). Crc is phenotypically similar to Lp, exhibiting an open neural tube from midbrain to tail (craniorachischisis), while splotch has spina bifida with or without a cranial NTD. The retinal axon abnormalities occur only in the presence of NTD and not in homozygous mutants lacking cranial NTD. Thus, failure of neural tube closure is associated with failure of many retinal axons to cross the ventral midline. This study therefore reveals an unexpected connection between closure of the neural tube at the dorsal midline and development of ventral axon tracts. genesis 27:32-47, 2000. PMID- 10862152 TI - Mesenchymal-epithelial transition in the developing metanephric kidney: gene expression study by differential display. AB - The developing metanephric kidney is a convenient model to study molecular events associated with epithelial cell differentiation. To determine the genes involved in the defining event of this process, namely, the conversion of metanephric mesenchyme to the epithelium of the nephron, we applied differential display (DD) techniques. Explants of rat metanephric mesenchymes were induced to condense ex vivo with fibroblast growth factor 2 (FGF2) or to form tubules with FGF2 and conditioned medium (CM) from a cell line (RUB1) of ureteric bud, the renal inductive tissue. Three time points (6, 24, and 72 h) were chosen to track the dynamics of gene expression during morphogenesis. Seventy-two up- or down regulated mRNAs were identified, including 36 novel sequences and those of cell cycle regulatory proteins (TGF-beta2, Cyclin D1, p57Kip2), transcription factors (beta-catenin, Sox11, DP1), signaling proteins (SH3-domain binding protein, G protein-coupled receptor, Ser-Thr protein kinase), cell adhesion molecules (syndecan-4, integrin-beta1), and also gene33, H19, SM20, IGFBP5, MAMA receptor, lectin, keratin, beta-tubulin, calreticulin, GRP78, ERp72, MnSoD, thioredoxin, and others. Some have previously been associated with kidney development and serve as good controls for expected changes, while most have not been linked with kidney epithelial cell differentiation. Using thin sections of embryonic kidney and labeled antisense RNA probes, we applied RNA hybridization to confirm the results of DD and related the expression of these genes to specific cell lineages of the developing kidney. These results provide a window into the events that mediate this critical differentiation process and suggest that a limited number of interrelated events direct the epithelial conversion of metanephric mesenchyme. genesis 27:22-31, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10862155 TI - Classical respiratory physiology-gone the way of the dinosaurs? Do we need a jurassic park? PMID- 10862156 TI - Overall and peripheral inhomogeneity of ventilation in patients with stable cystic fibrosis. AB - We studied distribution of ventilation in patients with cystic fibrosis (CF) who had not had an exacerbation for some time. Patients performed either the vital capacity nitrogen (N(2)) single-breath washout test (VC test) or a modified single-breath washout consisting of 1 L inspired from functional residual capacity (FRC test) of 90% oxygen (O(2)), 5% helium (He), and 5% sulfur hexafluoride (SF(6)). We computed the slopes of phase III of N(2) concentration from the VC test (S(N2) (VC)) and the phase III slopes of the He (S(He)): The SF(6) (S(SF6)), and curves from the FRC test. S(N2) (VC) may be regarded as an index of overall ventilation and the difference (S(SF6) - S(He)) as an index of peripheral ventilation. Three groups were studied: CF, 28 CF patients (8-36 years of age); normal controls (NC), 33 normal nonsmokers (9-55 years of age); and a smoking group (SG), 42 non-CF smoking patients (39-79 years of age). Compared to the NC group, S(N2) (VC) is increased in the CF group, reflecting an overall ventilation impairment. There is no difference in S(N2) (VC) between the CF group and the SG group, suggesting that S(N2), though sensitive, is nonspecific. Compared to both NC and SG groups, (S(SF6) - S(He)) is decreased in the CF group, being on the average negative. This may imply that there is a peripheral impairment in the distribution of ventilation that originates in terminal and respiratory bronchioles. Negative (S(SF6) - S(He)) is statistically associated with the youngest CF patients, suggesting that terminal and respiratory bronchiolar involvement is linked to early stages of the disease. In older CF patients, (S(SF6) - S(He)) is more often positive, suggesting that even more distal airways, such as alveolar ducts, become involved in peripheral inhomogeneity of ventilation. PMID- 10862157 TI - Risk factors for emergence of Stenotrophomonas maltophilia in cystic fibrosis. AB - The number of patients with cystic fibrosis (CF) whose sputum culture has yielded Stenotrophomonas maltophilia has increased in the last 5 years at St. Christopher's Hospital for Children. We conducted a case-control study to determine risk factors for recovery of S. maltophilia in respiratory secretions from patients with CF. We reviewed the outpatient and inpatient records of patients colonized with S. maltophilia between 1993 and 1997, and of age-matched (at time of initial recovery of S. maltophilia) control patients with CF who had never had a positive sputum culture for S. maltophilia. Variables included age at time of CF diagnosis, gender, severity of CF (based on Shwachman-Kulczycki (S-K) scores and spirometry), frequency of hospitalizations, use of oral, intravenous, or inhaled antibiotics, and use of oral or inhaled corticosteroids in the 2 years prior to the first isolation of S. maltophilia from respiratory secretions. Statistical methods included stepwise logistic regression to determine risk factors for acquisition of S. maltophilia. During the study period, 58 patients with CF had a positive sputum or deep throat culture for S. maltophilia. The distribution of S. maltophilia acquisition by year increased from 7 patients in 1993 (incidence, 2.8%) to 16 in 1997 (incidence, 6.2%). Patients positive for S. maltophilia were found to have significantly worse growth parameters, S-K score, and spirometric values than S. maltophilia-negative CF controls (P < 0.05). Stepwise logistic regression demonstrated that treatment with long-term antibiotics (P = 0.0016) and number of days of intravenous antibiotic therapy (P = 0.035) were significant risk factors for S. maltophilia colonization in our group of CF patients. We conclude that patients with CF whose respiratory secretions yield S. maltophilia have an overall worse clinical status at the time of initial S. maltophilia isolation than noncolonized patients, and that preceding treatment with antibiotics may have predisposed them to the acquisition of this bacterium in their respiratory secretions. PMID- 10862158 TI - Effect of a short course of rhDNase on cough and mucociliary clearance in patients with cystic fibrosis. AB - The aim of the study was to measure the effect of a short course of recombinant human deoxyribonuclease I (rhDNase) on ciliary and cough clearance in a group of cystic fibrosis patients, using a radioaerosol and gamma camera technique. Patients were initially randomized to receive either rhDNase (2.5 mg qd) or placebo. Following the measurement of baseline clearance, patients were given a 7 day course of either rhDNase or placebo. The patient then returned on the seventh day for follow-up clearance measurements. This was followed by a 2-week washout period before the whole process was repeated with the alternative inhalation solution. On each of the study days, mucociliary clearance was initially measured for a period of 60 min (IC). This was followed by cough clearance (CC) measurements for 30 min, during which patients were requested to cough a total of 120 times. Post-cough clearance (PCC) was then measured for a further 60 min. Thirteen patients completed the study. Patients' age ranged between 18-38 years, and they had baseline values of FEV(1) of 27-103% of predicted values. Following completion of the course of rhDNase, there was a mean percent increase from baseline of 7.5% for FEV(1) and 5.4% for FVC% (P = 0. 03). There was a small, nonsignificant increase in IC (6.2 +/- 3.6%) on the rhDNase arm compared with the placebo arm (-2.3 +/- 2.9%), P = 0.1. No changes were seen in either CC (1.0 +/- 3.2% [rhDNase] vs. 1.9 +/- 2.4% [placebo], P = 0.9) or PCC (-0.7 +/- 1.5% [rhDNase] vs. 0.9 +/- 1.7% [placebo], P = 0.3). Patients who achieved a 10% or greater improvement in FEV(1) (n = 5) in response to rhDNase did not show any greater change in clearance than nonresponders. In conclusion, we were unable to demonstrate any improvements in either ciliary or cough clearance in response to a short course of rhDNase. The mechanism of action of this drug in vivo remains uncertain. PMID- 10862159 TI - French multicenter randomized double-blind placebo-controlled trial on nebulized amiloride in cystic fibrosis patients. The Amiloride-AFLM Collaborative Study Group. AB - The effect of amiloride, a sodium channel blocker, has been evaluated in a multicenter randomized double-blind placebo-controlled trial in cystic fibrosis patients more than 5-years-old (n = 137) whose forced vital capacity (FVC), forced expiratory volume in 1 sec (FEV(1)), and forced mid-expiratory flow (FEF(25-75)) were not below 50%, 50%, and 30% of reference values, respectively. Patients were randomly allocated to two parallel groups. Sixty-four patients were chronically colonized with Pseudomonas aeruginosa; they received either amiloride or placebo as a nebulized solution three times daily for 6 months. Routine treatments were continued. Patients chronically colonized with Pseudomonas received nebulized colimycine twice a day for a month during the third and sixth months of treatment. Bronchopulmonary exacerbations were treated in the usual way. The effects of the amiloride treatment were assessed at the end of the 6 month treatment period. The effects on FVC and secondarily on FEV(1), FEF(25-75), the number of days on antibiotic therapy, the Shwachman score, a nutritional index (weight/height(2)), the change in sputum bacterial flora, and nocturnal cough were assessed. For the patients not chronically colonized with Pseudomonas, the effect of the treatment was also evaluated by counting chronic colonizations with pathogens appearing during the trial period. The present study failed to demonstrate any significant benefit of amiloride over placebo on FVC, FEV(1), and the other secondary endpoints in the studied population. Neither the chronically colonized, nor the noncolonized patients benefited. The confidence intervals of the differences between treatment groups indicated small differences that were most likely of no clinical significance. Complementary analyses taking into account the gender, the type of mutation, the subpopulations whose FVC and FEV(1) were below 80% of normal values at the beginning of the study, and also patients less than 10 years old, did not show any statistically or clinically significant improvements following amiloride therapy. PMID- 10862160 TI - Effects of birthweight and oxygen supplementation on lung function in late childhood in children of very low birth weight. AB - Impaired respiratory function has been found frequently in ex-premature children, but it is unclear which specific factors influence this impairment the most. The aim of this study was to determine the importance of the contributions of birth weight, gestational age, neonatal respiratory disease, and its treatment on subsequent childhood lung function at age 11 years in a cohort of children of very low birth weight (VLBW; 2,000 g) of similar age. VLBW children were shorter and lighter than controls (P < 0.0001) at 11 years of age, and had reduced expiratory flows (P < 0.00001) and forced vital capacities (P < 0.001). The residual volume to total lung capacity ratio (RV/TLC ratio) was increased (P < 0.00001), while total lung capacity (TLC) remained unchanged. Those with bronchopulmonary dysplasia (BPD) had the lowest mean expiratory flows. Males had lower expiratory flows than females. On univariate analysis, gestational age by itself accounted for 8.8% of the explained variance in FEV(1) at 11 years of age, but birth weight accounted for 16% on its own; both together accounted for a further 0.2% (16.2%), suggesting that the latter was the dominant factor. On multivariate analysis, the contribution of birth weight and gestational age was small, and the best predictors at 11 years of age, which together explained 43.4% of the total variance in FEV(1), were log days of supplemental oxygen (9.6%) and a reported history of asthma (10.8%). For FEF(25-75), these predictors explained 7.2% and 13.4%, respectively, of the total explained variance of 40.6%. The relation between neonatal oxygen supplementation and childhood FEV(1) was such that up to 20 days of supplemental oxygen had little effect on subsequent FEV(1) at 11 years of age, but each additional week of supplemental oxygen after that time was associated with a progressive reduction in FEV(1) of 3%. These data confirm the significant role of supplemental oxygen in the neonatal period and a history of asthma on the subsequent reduction of expiratory flows in VLBW children. Birth weight was a more important prenatal factor than gestational age, but both were of lesser predictive significance than either supplemental oxygen or a reported history of asthma. PMID- 10862162 TI - Input respiratory impedance measured by head generator in preschool children. AB - The upper airway wall motion represents a serious problem when measuring the input impedance of the respiratory system (Zrs) by the forced oscillation technique, particularly in young children. To minimize this error, it has been proposed to vary transrespiratory pressure around the head rather than directly at the mouth, using the head generator technique (HGT). The aim of this study was to collect normative data in preschool children in whom the technique may prove most useful. Zrs was measured using HGT and 4-32-Hz pseudorandom noise input in 127 healthy children. Age ranged from 2. 8 to 7.4 years and height (H) from 0.89 to 1.29 m. The fast Fourier transforms of pressure and flow allowed us to calculate respiratory system resistance (Rrs(f)) and reactance (Xrs(f)) at each frequency (f). Resonant frequency (fn), respiratory system inertance (Irs), and compliance (Crs) were derived from the Xrs(f) data. The technique was accepted by more than 95% of the children. A coherence function or = 10% was associated with poorer metastasis-free survival but p53 was not. GENETICS: Type of fusion transcripts (SYT-SSX1 or SYT-SSX2) and Ki-67 were assessed in fresh frozen tissue from 33 patients. The 5-year metastasis-free survival for patients with SYT-SSX1 was 42% versus 89% for those with SYT-SSX2. The hazard ratio for metastasis associated with the SYT-SSX1 fusion transcripts was 7 (95% CI 1.5-36, log-rank p = 0.004). There was a significant association between SYT-SSX1 and high tumor proliferation rate. Comparative Genomic Hybridization revealed DNA sequence copy number changes in 35 of 69 tumor specimens. The frequency of aberrations/tumor were higher in monophasic tumors than in biphasic. Gains of chromosome 8 were associated with large tumors (> 5 cm). There was no obvious association between secondary aberrations and clinical outcome. CONCLUSIONS: Large tumor size, local recurrence, histologic Grade IV, MIB1 index > or = 10 and possibly SYT-SSX1 fusion transcript were associated with impaired clinical outcome. PMID- 10862211 TI - "Solutions" doesn't provide any new answers. PMID- 10862212 TI - Awareness, perception, and knowledge of heart disease risk and prevention among women in the United States. American Heart Association Women's Heart Disease and Stroke Campaign Task Force. AB - CONTEXT: One of 2 women in the United States dies of heart disease or stroke, yet women are underdiagnosed and undertreated for these diseases and their risk factors. Informed decisions to prevent heart disease and stroke depend on awareness of risk factors and knowledge of behaviors to prevent or detect these diseases. OBJECTIVE: Assess (1) knowledge of risks of heart disease and stroke and (2) perceptions of heart disease and its prevention among women in the United States. DESIGN AND SETTING: Telephone survey conducted in 1997 of US households, including an oversample of African American and Hispanic women. PARTICIPANTS: One thousand respondents 25 years or older; 65.8% white, 13.0% African American, and 12.6% Hispanic. MAIN OUTCOME MEASURES: Knowledge of heart disease and stroke risks, perceptions of heart disease, and knowledge of symptoms and preventive measures. RESULTS: Only 8% of the respondents identified heart disease and stroke as their greatest health concerns; less than 33% identified heart disease as the leading cause of death. More women aged 25 to 44 years identified breast cancer as the leading cause of death than women 65 years or older. Women aged 25 to 44 years indicated they were not well informed about heart disease and stroke. Although 90% of the women reported that they would like to discuss heart disease or risk reduction with their physicians, more than 70% reported that they had not. CONCLUSIONS: Most women do not perceive that heart disease is a substantial health concern and report that they are not well informed about their risk. Age influenced knowledge to a greater extent than ethnicity. Programs directed at young women that address the effects of lifestyle behaviors on long-term health are needed. Better communication between physicians and patients is also warranted. PMID- 10862213 TI - Heart disease prevention in US women. There is more work to be done. PMID- 10862214 TI - Preventive attitudes and beliefs of deaf and hard-of-hearing individuals. AB - OBJECTIVE: To investigate the unique health care issues of deaf and hard-of hearing (D&HH) persons by studying their attitudes, beliefs, and behaviors toward preventive medicine. DESIGN: A self-administered, cross-sectional survey, written in a format comprehensible to persons whose primary language is American Sign Language. POPULATION: One hundred forty D&HH persons recruited from southeastern Michigan, Chicago, Ill, and Rochester, NY, and 76 hearing subjects from southeastern Michigan and Rochester. RESULTS: No significant differences existed between D&HH or hearing persons from different states. However, numerous differences existed between D&HH and hearing persons. Deaf and hard-of-hearing persons were less likely to report receiving preventive information from physicians or the media, and more likely to report receiving it from a Deaf club. They rated the following physician-initiated procedures as less important than hearing persons: discussion of alcohol consumption, smoking, depression, and diet, plus screening for hypertension, hearing loss, and cancer. Deaf and hard-of hearing persons often considered a preventive procedure important if it was reported performed at their last health maintenance examination. They were less likely to report being asked about alcohol consumption and smoking, or to having been examined for hypertension, cancer, height, and weight. They were more likely to report receiving a hearing examination, mammogram, and Papanicolaou smear. Deaf and hard-of-hearing persons were less likely to report believing that smoking less, exercising regularly, maintaining ideal weight, and regular physical examinations improve health. Differences existed within the D&HH cohort depending on the respondent's preferred language (oral English vs American Sign Language); our sample size was too small for a complete assessment of these differences. CONCLUSIONS: Deaf and hard-of-hearing persons appear to have unique knowledge, attitudes, and behaviors regarding preventive medicine, and their attitudes are influenced by their personal experiences with physicians. Preventive practices addressed during health visits may differ between D&HH and hearing patients. Further research is needed to clarify the reasons for these differences, including within D&HH subgroups, and to develop effective mechanisms to improve the health care of all D&HH persons. PMID- 10862215 TI - Prediction of probable Alzheimer disease in patients with symptoms suggestive of memory impairment. Value of the Mini-Mental State Examination. AB - BACKGROUND: The Mini-Mental State Examination (MMSE) is a widely used diagnostic tool for dementia. Its use as a predictive indicator of probable Alzheimer disease (AD) has not been established. OBJECTIVES: To determine the accuracy of the MMSE in predicting emergent AD in a sample of patients who were referred because of symptoms suggestive of memory problems and to determine whether an abbreviated version of the MMSE could be developed that would be as accurate as the full MMSE in predicting emergent AD. DESIGN: Inception cohort of participants with symptoms suggestive of memory impairment by their family physicians were given baseline assessments, including MMSE. After 2 years, the participants' conditions were diagnosed following the standard criterion for AD. Diagnosticians were blind to baseline scores. SETTING AND PARTICIPANTS: One hundred eighty-three community-residing participants were referred by their family physicians to a university teaching hospital research investigation. After baseline screening, 165 participants were included in the study who did not have dementia and had no identifiable cause for memory impairment. After 2 years, 29 participants met criteria for AD, 98 did not develop dementia, 18 developed vascular lesions or non-AD dementia, and 20 did not return. MAIN OUTCOME MEASURE: Diagnostic classification of AD or no evidence of dementia. RESULTS: Logistic regression model was significant. At a cutoff score of 24 or less, sensitivity was 31%; specificity, 96%; with a likelihood ratio of 7.75. A reduced model of 2 subtests was identified with a sensitivity of 41%; specificity, 98%; with a likelihood ratio of 20.70. CONCLUSIONS: Results suggest that the full or abbreviated MMSE is useful in predicting emergent AD in patients with positive test results. However, it is not recommended for use as a screening or diagnostic instrument since a negative test result did not rule out emergent AD. It is recommended as a tool to identify those needing closer monitoring. PMID- 10862216 TI - How often do office blood pressure measurements fail to identify true hypertension? An exploration of white-coat normotension. AB - BACKGROUND: The often-observed differences between ambulatory (ABP) and office blood pressure (OBP) measurements have brought attention to the problem of misdiagnoses. Although there has been much focus on white-coat hypertension (elevated OBP with normal ABP means), few studies have examined "white-coat normotension" (WCN; normal OBP with elevated ABP means). OBJECTIVES: To describe patients with WCN in terms of prevalence and quantitative differences between ABP and OBP; to identify psychological and demographic features that discriminate them from true normotensive patients; and to offer possible corrections for diagnostic limitations of OBP measurements in clinical practice. DESIGN AND METHODS: Five OBP measurements and 10- to 12-hour daytime ABP monitoring in 319 presumed healthy participants. RESULTS: Prevalence rates of WCN were 23% for systolic BP and 24% for diastolic BP. Participants with WCN were more often male, past smokers, and older and consumed more alcohol. Increasing the number of office readings and discarding the first office reading did not improve the accuracy of OBP measurements. Participants with BP of 10 mm Hg above or below the 140/90 office reading cutoff showed the lowest accuracy, with more than 50% of normotensive diagnoses being incorrect. CONCLUSIONS: Office measures of BP lack sensitivity, missing a sizable portion of individuals who have hypertensive mean ABP measurements. Subjects with WCN differ from true normotensive subjects on several demographic and lifestyle variables. Only those office readings averaging 20 points above or below the 140/90 cutoff represent safe diagnostic information. PMID- 10862217 TI - Does the structure of clinical questions affect the outcome of curbside consultations with specialty colleagues? AB - BACKGROUND: Clinical questions frequently arise during the practice of medicine, and primary care physicians frequently use curbside consultations with specialty physicians to answer these questions. It is hypothesized that well-formulated clinical questions are more likely to be answered and less likely to receive a recommendation for formal consultation. OBJECTIVE: To assess the relationship between the structure of clinical questions asked by family physicians and the response of specialty physicians engaged in curbside consultations. DESIGN AND PARTICIPANTS: A case series of clinical questions asked during informal consultations between 60 primary care and 33 specialty physicians using an e-mail service. Curbside consultation questions were sent, using e-mail, to academic specialty physicians by primary care physicians (faculty, residents, and community practitioners) in eastern Iowa. MAIN OUTCOME MEASURES: Questions were analyzed to determine the clinical task and to identify 3 components: an intervention, a comparison, and an outcome. Consultants' responses were analyzed to identify whether questions were answered and whether consultants recommended formal consultation. RESULTS: There were 708 questions in this analysis: 278 (39.3%) were diagnosis questions, 334 (47.2%) were management questions, 57 (8.0%) were prognosis questions, and 39 (5.5%) were requests for direction. Clinical questions were less likely to go unanswered or receive a recommendation for formal consultation when the question identified the proposed intervention (odds ratio, 0.54; 95% confidence interval, 0.34-0.86; P = .006) and desired outcome (odds ratio, 0.46; 95% confidence interval, 0.29-0.69; P < .001). Only 271 (38.3%) of 708 curbside consult questions identified both of these components. CONCLUSION: Medical specialists' responses to curbside consultation questions seem to be affected by the structure of these clinical questions. PMID- 10862218 TI - Infantile Henoch-Schonlein purpura. AB - Henoch-Schonlein purpura is a common cause of vasculitis in children. This condition is unusual in infants and children younger than 2 years. We describe a 4-month-old infant with infantile Henoch-Schonlein purpura and review the clinical spectrum, differential diagnoses, and the histopathologic features of the disease. Its relations to Henoch-Schonlein purpura in older children are discussed. PMID- 10862219 TI - Maximum antihypertensive effect from different classes of antihypertensives. PMID- 10862220 TI - Methicillin-resistant Staphylococcus aureus infections in 2 pediatric outpatients. AB - Methicillin-resistant Staphylococcus aureus (MRSA) infections are an emerging problem in children. The following are 2 case reports of unsuspected MRSA infections: the first is an infant with cervical adenitis and the second is a child with a deep infection of the toe. Both patients failed outpatient therapy with oral cephalosporins and required hospitalization for surgical drainage. Both patients had cultures positive for MRSA at surgery. Neither patient had any risk factors for acquiring MRSA. Thus, outpatients with presumed staphylococcal infections who fail oral therapy with cephalosporins may be infected with MRSA. PMID- 10862221 TI - Failure of treatment with cephalexin for Lyme disease. AB - CONTEXT: Lyme disease typically presents with a skin lesion called erythema migrans (EM), which though often distinctive in appearance may be confused with cellulitis. The first-generation cephalosporin, cephalexin monohydrate, is effective for treating bacterial cellulitis but has not been recommended or studied for treating Lyme disease because of poor in vitro activity. OBJECTIVE: To describe the outcome of patients with EM who were treated with cephalexin. PATIENTS AND METHODS: Patients presenting with EM to the Lyme Disease Diagnostic Center in Westchester, NY (May 1992-September 1997). A 2-mm punch biopsy specimen of the leading edge of the EM lesion and/or blood was cultured for Borrelia burgdorferi. RESULTS: Eleven (2.8%) of 393 study patients had been initially treated with cephalexin prior to our evaluation; 9 (82%) were originally diagnosed with cellulitis. Cephalexin was administered for 8.6 days (range, 2-21 days) prior to presentation. All 11 patients had clinical evidence of disease progression, including 8 whose rash enlarged, 2 who developed seventh-nerve palsy (1 with new EM lesions), and 1 who developed new EM lesions. Borrelia burgdorferi grew in cultures from 5 patients despite a mean of 9.8 days of treatment with cephalexin (range, 5-21 days). CONCLUSION: Cephalexin should not be used to treat early Lyme disease and should be prescribed with caution during the summer months for patients believed to have cellulitis in locations where Lyme disease is endemic. PMID- 10862222 TI - Home HIV testing: why not in Canada? PMID- 10862223 TI - Patient compliance with drug therapy for diabetic nephropathy. PMID- 10862224 TI - Access to the morning-after pill in BC. PMID- 10862225 TI - Access to the morning-after pill in BC. PMID- 10862226 TI - Managing hypertension in patients with renal disease and diabetes. PMID- 10862227 TI - Managing hypertension in patients with renal disease and diabetes. PMID- 10862228 TI - Folate and vitamin B12 status of women in Newfoundland at their first prenatal visit. AB - BACKGROUND: Newfoundland has one of the highest rates of neural tube defects in North America. Given the association between low maternal folic acid levels and neural tube defects, a cross-sectional study was conducted to obtain base-line data on the folate and vitamin B12 status of a sample of women in Newfoundland who were pregnant. METHODS: Blood samples were collected between August 1996 and July 1997 from 1424 pregnant women in Newfoundland during the first prenatal visit (at approximately 16 weeks' gestation); this represented approximately 25% of the women in Newfoundland who were pregnant during this period. The samples were analysed for serum folate, vitamin B12, red blood cell folate and homocysteine. RESULTS: Median values for serum folate, red blood cell folate and serum vitamin B12 were 25 nmol/L, 650 nmol/L and 180 pmol/L, respectively. On the basis of the interpretive criteria used for red blood cell folate status, 157 (11.0%) of the 1424 women were deficient (< 340 nmol/L) and a further 180 (12.6%) were classified as indeterminate (340-420 nmol/L). Serum homocysteine levels, measured in subsets of the red blood cell folate status groups, supported the inadequate folate status. Serum vitamin B12 levels of 621 (43.6%) women were classified as deficient or marginal; however, the validity of the interpretive criteria for pregnant women is questionable. INTERPRETATION: A large proportion of pregnant women surveyed in Newfoundland in 1997 had low red blood cell folate levels. PMID- 10862229 TI - Chest pain with nondiagnostic electrocardiogram in the emergency department: a randomized controlled trial of two cardiac marker regimens. AB - BACKGROUND: Early detection of acute myocardial infarction (AMI) may save lives. In the emergency setting, it is unclear whether the early use of certain cardiac markers (myoglobin and cardiac troponin I [cTnI]) assists in making appropriate decisions whether to admit or discharge patients with chest pain of possible ischemic cause who have nondiagnostic electrocardiograms (ECGs). We performed a study to determine whether the addition of new cardiac markers in the emergency department results in improved clinical decisions. METHODS: A single-blind randomized controlled trial was conducted between June 1997 and June 1998 in a tertiary care emergency department in Kingston, Ont. Of 296 patients aged 30 years or more who presented to the emergency department with chest pain and nondiagnostic ECGs, 146 were randomly assigned to the intervention group (determination of baseline creatine kinase [CK] level, CK MB fraction and cTnI level, and myoglobin level at baseline and at 2 hours) and 150 to the control group (determination of baseline CK level and CK MB fraction). Outcome measures included the rate of admission to the inpatient cardiology service and length of stay in the emergency department. RESULTS: Of the 296 patients, 34 (11.5%) received a diagnosis of AMI in the emergency department, and 92 (31.1%) had chest pain of noncardiac cause. Patients in the intervention group were less likely than those in the control group to be admitted to the cardiology service (67 [45.9%] v. 81 [54.0%]). The absolute difference in the proportion (8.1% [95% confidence interval -3.3 to 19.5]), although potentially important clinically, was not statistically significant. The length of stay in the emergency department was essentially the same in the 2 study groups. At 30 days, the proportions of patients with a diagnosis of recurrent angina (58.2% in the intervention group and 58.0% in the control group) and AMI (12.3% and 14.7%) were also similar. INTERPRETATION: The optimal cardiac marker panel to be used in the emergency department remains unknown. The addition of serial testing of myoglobin with cTnI confirmation to the standard panel did not substantially change the clinical management or outcomes of patients presenting with chest pain and nondiagnostic ECGs. PMID- 10862230 TI - Presence of spirochete causing Lyme disease, Borrelia burgdorferi, in the blacklegged tick, Ixodes scapularis, in southern Ontario. PMID- 10862231 TI - Folic acid: the opportunity that still exists; [comment]. PMID- 10862232 TI - Lyme borreliosis in Ontario: determining the risks. PMID- 10862233 TI - Promoting parental leave for female and male physicians. Gender Issues Committee of the Council of Ontario Faculties of Medicine. PMID- 10862234 TI - Rheumatology: 4. Acute monoarthritis. PMID- 10862235 TI - Withholding life-sustaining treatment: are adolescents competent to make these decisions? PMID- 10862236 TI - Building a "brain attack" team to administer thrombolytic therapy for acute ischemic stroke. AB - Before tissue plasminogen activator (tPA) was licensed for use in Canada, in February 1999, the Calgary Regional Stroke Program spearheaded the development and organization of local resources to use thrombolytic therapy in patients who had experienced acute ischemic stroke. In 1996 special permission was obtained from the Calgary Regional Health Authority to use intravenously administered tPA for acute ischemic stroke, and ethical and scientific review boards approved the protocols. After 3 years our efforts have resulted in improved patient outcomes, shorter times from symptom onset to treatment and acceptable adverse event rates. Areas for continued improvement include the door-to-needle time and broader education of the public about the symptoms of acute ischemic stroke. PMID- 10862237 TI - MDs' bid to raise drinking age meets opposition. PMID- 10862238 TI - New rapid HIV test opens Pandora's box of ethical concerns. PMID- 10862239 TI - Boys and girls and healthy behaviour. PMID- 10862240 TI - Pinpointing the genes at work in osteoarthritis and cardiovascular disease. PMID- 10862241 TI - Rapid HIV testing. PMID- 10862242 TI - Alberta ignores vocal opposition, presses ahead with law to expand role of private clinics. PMID- 10862243 TI - Will litigation become part of public health arsenal in Canada's war against smoking? PMID- 10862244 TI - Dr. Martin pushes private medicine, but not even reformers appear to be listening. PMID- 10862245 TI - [The Hemoccult II test: results of 16 years of screening tests at the Tumor Prevention Service of the City of Paris]. AB - OBJECTIVE: Colorectal cancer is a major health problem causing high mortality and short survival after diagnosis. Both the cancer itself and its precancerous lesion, the adenomatous polyp, can provoke occult bleeding. Screening programs for cancer of the colon use the Hemocult II test to detect such occult bleeding. We report the results obtained after 16 years of screening with the Hemocult II at the Paris tumor prevention center. METHODS: Since 1980, an annual Hemocult II test has been proposed to all consulting patients over the age of 40 years. Tests were self-administrated and returned to the center by mail. After rehydration, the tests were read by center nurses. RESULTS: The test was well accepted by the patients (78-90%). Over 16 years, the test led to the diagnosis of 18 cases of cancer, two-third of which were in an early stage (Dukes A) as well as 84 adenomatous polyps including 25% with signs of moderate to severe dysplasia and one case of diffuse adenomatous polyposis. Test sensitivity was 60%, with a 97% specificity and an overall (cancer and precancerous lesions) positive predictive value of 28.3%. In light of published data on the adenoma-cancer relationship, it can be estimated that 6 cancers were avoided. CONCLUSION: Hemoccult II is an easy to perform low-cost test well accepted by patients. Its use in a screening program can detect early stage precancerous and cancerous lesions of the colon. PMID- 10862246 TI - [Treatment of nocturnal enuresis in France]. AB - OBJECTIVE: To evaluate the current incidence of nocturnal enuresis in France and the ways this condition is managed. PATIENTS AND METHODS: A survey was conducted amongst 3,803 school children (5 to 10 years old). In addition, management of nocturnal enuresis proposed by various specialists were collected through an analysis of the literature. RESULTS: The incidence of nocturnal enuresis was found to be 9.2% (11.2% in the subgroup of patients 5 to 7 years of age). 42% of the children (the most severe cases) stated they were bothered by their problem and that it affected their social life, although the mothers had a tendency to minimize the impact of enuresis. 66% of the mothers of children suffering from moderate to severe enuresis did address the problem to a medical doctor, mainly a general practitioner. Amongst these latter, 20% did not propose any solution. The remaining mainly proposed advise for lifestyle and dietary habits as well as a "wait and see" attitude. As a specific treatment, drugs (oxybutynin 48%, desmopressin 22%, imipramine 12%) were prescribed more often than alarms as drugs proved to be more effective than alarms. 48% of the mothers of enuretic children considered that doctors do not pay enough attention to this problem. Options for specific treatments of nocturnal enuresis vary from one specialist to another. CONCLUSION: A consensus about the most appropriate management approach to nocturnal enuresis is needed. Management of voiding dysfunction should be part of the medical curriculum. PMID- 10862247 TI - [Extra-pyramidal syndrome induced by donepezil]. AB - BACKGROUND: The cholinergic hypothesis of Alzheimer's disease is the basis of a new class of drugs: acetylcholinesterase inhibitors. These drugs have few side effects, mainly digestive disorders. CASE REPORTS: Extra-pyramidal side effects with severe gait disorders were observed in 3 patients with Alzheimer's dementia treated with donepezil. This drug was associated with paroxetine or a neuroleptic. In 2 of the 3 cases, the extra-pyramidal effects disappeared when donepezil was discontinued. DISCUSSION: Extra-pyramidal syndromes in elderly subjects with cognitive impairment are difficult to interpret. The possible causes include interactions between acetylcholinesterase inhibitors, neuroleptics and serotonine reuptake inhibitors and Lewy body dementia. PMID- 10862248 TI - [Abdominal tuberculosis: deceptive and still encountered]. AB - BACKGROUND: Extrapulmonary manifestations of tuberculosis are increasing in incidence. Abdominal tuberculosis may mimic a variety of gastrointestinal disorders. The diagnosis of abdominal tuberculosis is still difficult to establish before surgery. CASE REPORTS: We report 3 cases of abdominal tuberculosis in immunocompetent individuals. One patient presented with an ileocecal mass mimicking cancer. The second one presented with fever, ileocecal mass and ascites leading to the diagnosis of appendiceal peritonitis. The last patient was admitted for ascites, ovarian mass and high CA 125 serum level simulating ovarian cancer with peritoneal carcinomatosis. COMMENTS: In cases of abdominal tuberculosis when standard investigations are unhelpful, a PCR should be performed. Estimation of adenosine deaminase in ascitic fluid is an easy and reliable method for diagnosing tuberculous ascites. With these non invasive diagnostic procedures, surgery should be reserved only to patients with complications. PMID- 10862249 TI - [Collaboration between the geriatrician and the surgeon: a major contribution for the aged patient in rehabilitation]. AB - BACKGROUND: Subtrochanteric fractures are complex fractures requiring prudent reeducation. In elderly patients who also have other illnesses, reeducation of walking may be quite difficult. CASE REPORT: An 89-year-old woman was referred for physical therapy after osteosynthesis of a comminutive subtrochanteric fracture. Weight bearing was contraindicated for 3 months due to her "incapacity to perform reeducation exercises". The patient also had several serious co morbidities which had to be integrated into the reeducation program. DISCUSSION: In cases of complex fractures, reeducation must take into account other comorbid conditions. Outcome can be favorable, but requires a well organized interdisciplinary approach associating the geriatrics and the orthopedics teams. PMID- 10862250 TI - [Digital clubbing associated with primary alveolar proteinosis: possible implication of growth factors]. PMID- 10862251 TI - [Myopericarditis disclosing acute toxoplasmosis]. PMID- 10862252 TI - [Postpartum myocardiopathy]. PMID- 10862253 TI - [Thank you to M. Souques and K. Zummer]. PMID- 10862254 TI - [Screening for colorectal cancer]. PMID- 10862255 TI - [Hemorrhoids, classical surgery or mechanical endoanal excision?]. PMID- 10862256 TI - [8th consensus conference of the French Society for Medical Emergencies. Emergency management of adult nephrotic colic. April 23 1999-Marseille]. PMID- 10862257 TI - [What aspirin dose after carotid endarteriectomy? Commentary on an explanatory study]. PMID- 10862258 TI - [Vascular risk and menopause: role of hormone replacement therapy]. AB - RATIONALE: Menopause dramatically increases cardiovascular risk in women who lack the protecting effects of estrogens. This emphasizes the importance of hormone replacement therapy (HRT) which, until recently, was considered beneficial in spite of the heterogeneous nature of clinical studies. Such a benefit was supposed to result from the favorable effects of estrogens, including lipid lowering and a complex vasodilating action including the improvement of vascular endothelial function. RATIONALE REVISITED: The results of the HERS study have however questioned these affirmations because of the lack of clinical benefit due to HRT in secondary prevention of coronary artery disease in postmenopausal women, with an increased vascular risk during the first year of treatment. In clinical practice, it is not recommended to start HRT after a cardiac attack. PRACTICAL ATTITUDE: In fact, the HERS study used a conjugated equine estrogen and progestin (with possible deleterious effects) and cannot be compared to French prescription habits which are based on the use of natural hormones. However, it remains to be demonstrated that some HRT regimens, in particular those using progesterone, may provide a real benefit in terms of lower cardiovascular morbidity-mortality. PMID- 10862259 TI - [Leishmaniasis: update]. AB - PARASITE IDENTIFICATION: Leishmaniases are a group of parasitic diseases affecting 88 countries in four continents. Isoenzymatic electrophoresis, presently the gold standard technique for Leishmania identification, has led to the development of the phenetic and phylogenetic classifications commonly in use. Study of the genome of these parasites is particularly important because of the features it presents. PARASITE CYCLE: Parasite transmission to mammals is facilitated by sandfly saliva. The parasite invades the human dermis by complex mechanisms involving molecular interactions allowing the parasite to survive after intracellular penetration. CLINICAL EXPRESSION: Clinical outcome of leishmaniases depends on both the Leishmania species tropism and the host's immune response. Immunosuppressive status, whatever its origin, facilitates Leishmania infection and worsens the clinical picture. TREATMENT: No new developments have occurred for a long time in the treatment of leishmaniases. Individual prophylactic measures are limited to insecticide impregnated bednets for human use and insecticide collars for prevention of canine leishmaniases. PMID- 10862260 TI - [Beta-blocker intoxication]. AB - SEVERITY: Beta-blocker intoxication is not frequent but can produce particularly severe or fatal conditions which must not be underestimated. Severity of beta blocker intoxication varies from one compound to another. The more toxic drugs are propranolol, sotalol, oxprenolol, metoprolol, atenolol, acebutolol, labetalol, and carvedilol. Besides the drug type, history taking can provide a precise assessment of risk, particularly important in when elderly patients with a cardiovascular history have taken more than 20 tablets, when emergency care is provided late, and when other cardiotoxic or psychotoxic drugs have been coingested. IMPLICATIONS: The diagnosis of beta-blocker intoxication must be suspected in any case associating hypotension and bradycardia. The main cardiovascular complications are cardiogenic shock, atrio-ventricular conduction disorders, and obviously life-threatening ventricular arrhythmia with cardiac arrest. Centrally induced respiratory arrest is a rare but dreadful consequence which can occur suddenly even without hemodynamic failure. Neurologic toxicity is mainly expressed by consciousness disorders and more sporadically by seizures. Laboratory tests show variable serum potassium, lactic acidosis, hypoxia hypercapnia resulting from hypoventilation, and rarely hypoglycemia. The ECG should be recorded early because electrocardiographic signs usually appear before clinical signs and QRS enlargement is a factor predictive of severe ventricular arrhythmia. INTENSIVE CARE: The patient must be placed in an intensive care unit for continuous multiparametric monitoring. Besides gastric evacuation and symptomatic measures, treatment essentially requires glucagon for its positive inotropic effect after high intravenous doses. If glucagon infusion is ineffective or unavailable, an alternative would be to use high doses of vasoactive agents, choosing isoproterenol or epinephrine as the first intention drugs. PMID- 10862261 TI - [Left retro-aortic renal vein]. PMID- 10862262 TI - The SEEM: selective estrogen enzyme modulators in breast cancer. AB - Human breast cancer tissue contains all the enzymes (estrone sulfatase, 17 beta hydroxysteroid dehydrogenase, aromatase) involved in the last steps of estradiol biosynthesis. This tissue also contains sulfotransferase for the formation of the biologically inactive estrogen sulfates. In the past years, it has been demonstrated that various progestins (promegestone, nomegestrol acetate, medrogestone) as well as tibolone and its metabolites are potent inhibitors of sulfatase and 17 beta-hydroxysteroid dehydrogenase activities. It was also shown that medrogestone, nomegestrol acetate, promegestone or tibolone can stimulate the sulfotransferase activity for the local production of estrogen sulfates. All these data, in addition to numerous agents which can block the aromatase action, lead to the new concept of Selective Estrogen Enzyme Modulators (SEEM) which can largely apply to breast cancer tissue. The exploration of various progestins and other active agents in trials with breast cancer patients, showing an inhibitory effect on sulfatase and 17 beta-hydroxysteroid dehydrogenase, or a stimulatory effect on sulfotransferase, will provide a new option in the treatment of this disease. PMID- 10862263 TI - Conjugated estrogens--the natural SERMs. AB - Tissue selective and metabolite replacement therapy may become a new aspect in hormone replacement therapy (HRT). In addition to the naturally secreted hormones, there are also the later formed metabolites that exert a characteristic pharmacological profile. This mechanism is well known in thyroid replacement therapy, when triiodothyronine, the metabolite of thyroxine, is added to substitution therapy. The same is true for testosterone replacement therapy, when dihydrotestosterone is used for replacement. Also in menopausal HRT these aspects will gain tremendous importance. Progesterone metabolites have a strong clinical potency as neurosteroids, and estradiol metabolites are important factors in angiogenesis and angiostasis. Conjugated estrogens consist of different metabolites such as 16-hydroxy-equilin, which has no angiogenetic effect compared with 16-hydroxy-estrone. Estrone sulfate, the main component in conjugated estrogens, can be activated into estrone and 17 beta-estradiol in a tissue specific manner. This aspect will become of interest in clinical practice with HRT. PMID- 10862264 TI - Conjugated estrogens and breast cancer risk. AB - Available epidemiologic data suggest the possibility that the use of oral conjugated equine estrogens (CEE) 0.625 mg/day as a first-choice dose could be associated with a very limited (if any) breast cancer risk increase. Some biological peculiarities of oral CEE back the possibility of a limited detrimental effect on breast tissue, due to either direct or indirect actions. Direct actions. Some experimental findings suggest that the 17 alpha dihydroderivatives of equilenin and equilin (15% of the CEE components) have a non-estrogenic or even an anti-estrogenic effect on breast tissue. This could partially counterbalance the stimulatory action of the other CEE components. Indirect actions. Oral estrogens, through their metabolic and hepatocellular effects (emphasized by the first liver passage) cause a sharp increase in sex hormone binding globulin (SHBG) level which is followed by a lower quantity of both estrogen and androgen in the free, bioavailable, form. More importantly, they cause a decrease in circulating insulin-like growth factor I (IGF-I) activity, due to both a reduction in IGF-I synthesis by the liver and an increase in IGF-binding protein-1 level. A strong relationship between breast cancer risk and the concentration of circulating IGF-I in premenopausal women has been recently found. Actually, estrogens and IGF-I have a synergistic effect on cell proliferation, and IGF-I is necessary for maximum estrogen-receptor activation in breast cancer cell lines. The possibility does exist that the SHBG level increase and the IGF-I bioavailability decrease, caused by oral CEE, balance the increased estrogen stimulation on breast tissue. PMID- 10862265 TI - Modification of serum lipids and cardiovascular risk by estrogenic active compounds. AB - Cardiovascular disease is an important cause of death among women. After the menopause there is a steep elevation of low-density lipoprotein (LDL) cholesterol and lipoprotein(a) concomitantly with other risk factors, followed by a striking increase in arteriosclerotic vascular disease, compatible with a lack of estrogens as a causative factor. The benefit of hormone replacement therapy has been documented in several large studies, although rigorous randomized trials are still under way. However, cardiovascular events have been reduced by approximately 50%. In order to optimize beneficial effects and minimize side effects, the characteristics of selective estrogenic active compounds are a major target of research. The anti-estrogens are the prototype of partial and selective estrogen functions. The components of conjugated estrogens are under investigation for their specific effects. For example, both 17 alpha dihydroequilin sulfate and 17 alpha-dihydroequilenin sulfate seem to exert antioxidant activity without significant proliferative effects on uterine or breast tissues. This suggests a selective estrogenic activity dependent on the predominant type of estrogen receptor in the particular tissue. Equilin sulfate exerts a significantly stronger antioxidant activity than 17 beta-estradiol in vitro, and delta 8-estrone sulfate increases the lag time for low-density lipoprotein oxidation in vivo. These actions on lipoproteins may add to the quantitative changes of lipoproteins, while other effects are independent of lipoproteins. The 17 alpha-dihydroequilin sulfate and equilin sulfate improved the action of insulin in vivo. Data from rhesus monkeys treated with 17 alpha dihydroequilenin sulfate indicate that additional mechanisms are probably responsible for the observed cardiovascular protection. Further studies need to be conducted in order to identify selective estrogen receptor modulators and assess their potential, especially in lowering cardiovascular risk. PMID- 10862266 TI - Quantitative assessment of bone mineral density during estrogen replacement therapy. PMID- 10862267 TI - Role of progestins in the premenopausal climacteric. AB - Both the prevention of diseases and therapy of symptoms in premenopausal women is a demanding task for the gynecologist. Present day knowledge allows a differentiated approach for the use of the various progestins in the premenopausal period. A women-specific medicine needs individual selection of the progestins, both for substitution and treatment. The various partial effects of the progestins, such as androgenic, antiandrogenic, estrogenic, antiestrogenic, glucocorticoid etc., have to be taken into account and should be applied according to the individuals' needs. The benefits of progestins can be obtained in various clinical conditions, including bleeding disorders; benign proliferative diseases (such as benign breast disease, endometrial hyperplasia and endometriosis); contraception; androgenization; and gynecological oncology. PMID- 10862268 TI - Lipid profile in chronic renal failure at a moderate altitude of 2250 m. AB - There is no statistically significant difference between the two groups (CRF and control) as regard the serum lipid values regardless of the severity, age and etiology. The dyslipidemia in present study not correlate with the risk of developing cardiovascular disease reinforcing the earlier finding of increased levels of HDL-c at higher altitudes. Still in practice the lipid profile should be estimated in all patients and those found to be inappropriately high with increased cardiovascular risk should be treated as recommended in the general population. PMID- 10862269 TI - Role of ketamine in convulsions. AB - It can be concluded from the present study that ketamine showed dose-dependent anticonvulsant effect on MES in mice. It is presumed that this anticonvulsant effect of ketamine could be due to blockade of excitatory amino acid NMDA receptors. It was potentiated by anticonvulsants like diazepam and DPH but was found to be insensitive to naloxone. These findings suggest the involvement of NMDA receptors and their antagonists in epilepsy. Ketamine thus can be given as add-on therapy in refractory cases, and may prove to be useful as an anticonvulsant in future. PMID- 10862270 TI - Mycoplasmal antibodies as determined with an enzyme-linked immunosorbent assay, in tubal factor infertility. AB - A total of 81 infertile women, who had been referred for diagnostic loparoscopy, were tested for the presence of antibodies to Mycoplasma hominis and T mycoplasma. Out of 81, 30 had tubal adhesions and 51 had unilateral/bilateral tubal blockage. Antibodies to M. hominis were found in 21/30 (70%) and 14/51 (27.45%) women, antibodies to T-mycoplasma in 12/20 (40% and 39/51 (76.47%) women with tubal disorder. In a control group of 40 pregnant women, antibodies to the same two organisms occurred in 10% and 32.5%. Antibodies to M. hominis and T mycoplasma were significantly (P < 0.001) more common in women with tubal disorder. Our results confirm the important role of M. hominis and T-mycoplasma in the aetiology of tubal infertility. PMID- 10862271 TI - Drug resistance pattern and phage types of Salmonella typhi isolates in Manipal, South Karnataka. AB - Out of the 226 strains of S. typhi isolated over a period of three years 57.9% of them were multidrug resistant. 8.8% of the isolates were sensitive to all the drugs tested. A significant decline in hte number of multidrug resistant strains was observed in this region. Majority of the isolates belonged to phage type E (75.7%) and Biotype 1 (93.8%). All strains isolated were sensitive to ciprofloxacin, norfloxacin and ceftriaxone. PMID- 10862272 TI - Chronic lymphatic leukemia presenting as hemolytic anaemia. AB - In CLL autoimmune phenomena manifest most commonly as autoimmune hemolytic anemia followed by thrombocytopenic purpura and pure red cell aplasia. Here an elderly lady presenting with AIHA as the presenting manifestation of CLL is described. PMID- 10862273 TI - Donor selection for allogeneic bone marrow transplantation. PMID- 10862274 TI - AIDS cure from donkey's milk? 'Immuno-stimulants in milk could provide cure for cancer, TB also'. PMID- 10862275 TI - Asthma drug from African soil. PMID- 10862276 TI - New therapy replaces major uterine surgery. PMID- 10862277 TI - Noteworthy discoveries in molecular biology show promise for longer lives in the future. PMID- 10862278 TI - Aetiopathogenesis of keloids. PMID- 10862279 TI - A study of changes in the status of autonomic nervous system in primary open angle glaucoma cases. AB - Control of Intra Ocular Pressure (IOP) is influenced by both divisions (sympathetic and para-sympathetic) of Autonomic Nervous System (ANS). The present study was conducted to confirm existence of any association of ANS with primary open angle glaucoma (POAG), if present. Systemic autonomic function tests were assessed in fifty patients of POAG with fifty normal subjects matched for sex and age as control using several well established tests based on cardiovascular reflex responses to standardised stimuli viz. Resting heart rate (RHR), Corrected QT Interval (QTc), T-wave amplitude, Standing to Lying Ratio (SLR); Valsalva Ratio; Galvanic Skin Resistance (GSR) and Cold Pressor Test (CPR). Tests were conducted in the Department of Physiology, Maulana Azad Medical College and associated Glaucoma Clinic of Guru Nanak Eye Centre from june '94-May '95. Subjects were randomly chosen from known POAG cases with IOP--25 +/- 5.08 mm Hg. The results showed decrease in both sympathetic and para-sympathetic activity of autonomic function tests in POAG suggesting associated autonomic dysfunction. Sympathetic under-activity is seen in 36 patients out of 50 (73%), POAG subjects while para-sympathetic activity is decreased in 43 of 50 (86%) of the POAG subjects when compared with normal control group. The results help concluding that POAG is associated with autonomic dysfunction with decreased activity of both sympathetic and para-sympathetic divisions of ANS. PMID- 10862280 TI - Unexplained fever-analysis of 233 cases in a referral hospital. AB - A study was conducted to analyse the causes of fever of unknown origin (FUO) in a teaching hospital in central India. Study subjects consisted of 233 patients having FUO admitted in the medical ward. Specific causes of FUO were identified in 73.4% cases. The commonest causes (46.4%) were of infectious diseases origin foremost being enteric fever (29.6%) followed by malaria (9.0%) and tuberculous fever (5.2%). Chloroquine responsive fever accounted for 26% cases of FUO. Enteric fever were seen more commonly in younger adults less than 50 years, tuberculous fever presented usually after four weeks of onset of symptoms and more in elderly patients aged 50 years or more. Intermittent type of fever was more commonly recorded in infectious diseases. Approach to causes of FUO should be focused primarily on infectious diseases followed by other specific investigations. Empirical treatment of cases having intermittent fever with chloroquine seems justifiable even in absence of malarial parasite in peripheral blood smear. PMID- 10862281 TI - Epistaxis: study of aetiology, site and side of bleeding. AB - The present study comprises 300 cases of epistaxis. The analysis of these cases revealed a higher incidence in young males. Unilateral bleeding was seen in almost 60% each of indoor and outdoor cases. Litte's area was the most common site responsible for epistaxis in 28.8% of the indoor and 26.2% of the outdoor patients. Hypertension was the most common systemic cause among indoor patients (62.2%) and sickle cell disorder among the outdoor patients (37.5%). Atrophic rhinitis with myiasis was the local cause of epistaxis in maximum (27%) of the indoor patients and traumatic epistaxis was the commonest cause (33%) among outdoor patients-fingernail trauma in 75.9% of them. Idiopathic epistaxis contributed for 16.5% indoor and 26.1% of outdoor cases. Intractable epistaxis was seen in one case following accidental facial trauma. PMID- 10862282 TI - The top medical advances of 1998. PMID- 10862283 TI - Diagnosis of Chlamydia trachomatis: laboratory methods. PMID- 10862284 TI - Neurogenic appendicopathy--role of enterochromaffin cells in its pathogenesis. AB - One hundred cases of neurogenic appendicopathy were histochemically studied for schwann cells and enterochromaffin cells. The early phase, labelled as neuro appendicopathy (29 cases) showed minimum to moderate number of extraepithelial enterochromaffin cells without neurogenous hyperplasia. In 53 cases, there was intra and submucosal neural hyperplasia with increase in the extraepithelial enterochromaffin cells, representing the active phase. The late phase known as obliterative neurogenic appendicopathy, showed extraepithelial enterochromaffin cells and schwann cell proliferation of variable grades (18 cases). The origin of extraepithelial enterochromaffin cells is related to proliferating nerve plexus, rather than epithelial enterochromaffin cells. PMID- 10862285 TI - Neutrophil functions in rheumatoid arthritis. AB - Neutrophils play an important role in the pathogenesis of rheumatoid arthritis by accumulation and liberation of active proteolytic enzymes. Despite the active participation of the neutrophils, the patients afflicted with rheumatoid arthritis are prone to multiple infections. We studied neutrophil functions in 20 rheumatoid arthritis patients in active disease and equal number in remission and 20 healthy normal controls. No change in neutrophil function was seen in patients in remission. Phagocytic capacity of the neutrophils in active disease was found to be significantly reduced (p < 0.05). This inversly correlated with the rheumatoid factor (r = -0.128, p = 1). Random migration and chemotaxis was statistically reduced when compared with either healthy controls (p < 0.01) or when compared with patients in remission (p < 0.01). The chemotaxis inhibition was further enhanced by autologus serum (p < 0.05). The serum from patients with active disease also reduced chemotaxis of neutrophils from normal individuals (p < 0.01), indicating reduced cellular response as well as inhibitors in serum. The positive correlation (r = 0.466, p < 0.01) with rheumatoid factor, suggests the inhibitory activity may be due to the circulating rheumatoid factor in the active disease. The postulate that prior saturation of neutrophil receptors with immune complexes lower phagocytosis as well as chemotaxis is sustained. Destruction of chemotaxis receptors by release of various strong oxidative enzymes by neutrophils may also be a factor. Normal leucocytes are seen to take up immunoglobulins from diseases serum but not from normal serum. This uptake of diseased serum may be responsible for reducing the chemotactic and phagocytic function of neutrophils and hence increased incidence of infection in these patients. PMID- 10862286 TI - Immunohistopathological reactions for liver-specific membrane lipo-protein in experimental autoimmune hepatitis. AB - Antibody to the hepatocyte membrane protein, was induced in inbred strain C57BL/6 and C3H mice by immunisation with 100,000 g supernatant of syngeneic liver homogenate in CFA. Three weekly intraperitoneal injection of 200 ul of liver homogenate with CFA for continuous 4 weeks gave the best possible result. Histopathological changes were characterised mainly by perivascular inflammatory infiltrates and hepatocyte necrosis which mimicked human autoimmune hepatitis. In one of the immunological parameters, antibody to hepatocyte membrane protein (LSP) has been demonstrate by ouchterlony method in the test serum of those animals, who had received weekly doses of liver antigen. Thus in experimental autoimmune liver disease, semi-purified syngeneic liver fluid (S-100) leads to hepatic destruction and to an inflammatory process with several features in common with human chronic aggressive hepatitis. The presence of antibody against syngeneic liver antigen (S-100) in the test sera emphasizes that hepatocyte membrane protein does have an important role in liver tissue pathogenesis and disease process in experimental model. In this study we tried to prove that hepatocyte membrane protein may act as a target antigen in developing experimental autoimmune hepatitis. PMID- 10862287 TI - Red cell alloimmunization in multi-transfused chronic renal failure patients undergoing hemodialysis. AB - Red blood cell (RBC) transfusions are frequently used in the management of patients with chronic renal failure (CRF) undergoing hemodialysis for dialysis related anaemia. Consequently, they are subject to all hazards associated with repeated transfusions, such as red cell alloimmunization. A retrospective study was performed to estimate the frequency of alloimmunization against red cell antigens in multitransfused CRF patients. A total of 81 patients (67 males & 14 females) with CRF were studied who received a mean of 8.5 units of RBC matched for ABO & Rh(D) antigens only. Using standard techniques (indirect antiglobulin test, enzyme, polyethylene glycol, and low ionic strength solution), we observed a RBC alloimmunization rate of 9.8% (8/81). Nine alloantibodies were detected in 8 patients, and most (88%) involved antigens in the Rhesus & Kell systems. No correlation was observed with the alloantibody formation & number of units transfused. The calculated risk of 1.3% observed in the present study, suggests that renal failure patients are not at a higher risk of red cell alloimmunization than the general population. PMID- 10862288 TI - Cytomorphological spectrum of cysticercosis--a review of 132 cases. AB - A retrospective analysis of fine needle aspirates of 132 cases of cysticercosis presenting as palpable nodule is presented. In 98 cases, larval parts, detached hooklets and scolex established the diagnosis; in another 24 cases, the background inflammatory pattern was helpful in suggesting the diagnosis of a parasitic lesion. PMID- 10862289 TI - Role of enzyme linked immunosorbent assay in the diagnosis of suspected cases of genito urinary tuberculosis. AB - The aim of study was evaluation of the utility of ELISA test using A60 Antigen for rapid diagnosis of Genitourinary Tuberculosis in various age groups. ELISA test based on mycobacterial antigen A60 (Anda biological, France) was used to estimate specific IgG antibodies in the sera of fifty four suspected cases of Genito urinary tuberculosis. (GUT)Sera of 30 montoux negative healthy adults (age/sex matched) were taken as control by detecting IgG anti bodies to A60 antigen. It was concluded from this study that IgM was positive in 87.0% of cases. PMID- 10862290 TI - Study of diagnostic modalities and pathology of Helicobacter pylori infection in children. AB - To evaluate various diagnostic tests for Helicobacter pylori (Hp) in children, and to study the spectrum of endoscopic and histological changes in the stomach and duodenum of children with gastroduodenal disorders, associated with Hp infection Children below 12 years of age with various gastroduodenal disorders requiring upper gastrointestinal endoscopy were studied. Endoscopic biopsy specimens were collected from duodenum and antrum. Apart from histopathological examination of biopsy material, rapid urease test (RUT) of the antral biopsy specimen and blood examination to estimate specific IgG antibodies to Hp by Indirect Solid Phase Enzyme Immunoassay was performed. Forty seven children were included. Nine (19.1%) of them were positive both by serology and RUT. Seven (14.9%) were positive by histology. A significant correlation of Hp was noticed with chronic antral gastritis (p = 0.002) and chronic duodenitis (p = 0.006). Age equal to or more than 10 years was found to be significant risk factor for acquiring Hp infection. Prevalence of Hp in children with gastroduodenal complaints was found to be 19%. Both RUT and serology were found to be reliable diagnostic tests for Hp as compared with histology. Antral gastritis and chronic duodenitis had a significant correlation with Hp colonization. PMID- 10862291 TI - Bacteraemia in a tertiary care urban hospital in south India. AB - A total of 1727 blood samples were cultured aerobically over a one year period, of which 201(11.8%) were positive. The ratio of Gram positive to Gram negative bacteraemia was 1:1. The three antimicrobials having the highest activities against the Gram positive isolates were amikacin, cefotaxime, and ciprofloxacin to which 88.5, 81.7 and 80.7 percent of the strains were susceptible: and the same agents were equally effective against Gram negative organisms with 84.5, 75.3 and 70.1 percent efficacy respectively. Coagulase negative Staphylococcus spp. was the most frequent organism isolated(60; 29.8%), followed by Pseudomonas aeruginosa (40; 19.9%), and Staphylococcus aureus (34; 16.9%). PMID- 10862292 TI - Virulence factors in uropathogenic E. coli. AB - Escherichia coli isolates from urinary tract infections often exhibit characters different from those isolated from normal faecal samples. Adherence to uroepithelial cells, nature of lipopolysaccharide O antigen and mannose resistant haemagglutination of human erythrocytes are some of the important virulence factors proposed in the pathogenesis of urinary tract infections caused by E. coli. In the present study a total of 100 strains of E. coli isolated from symptomatic cases of urinary tract infections (with significant bacteriuria) were studied for these properties. Faecal isolates of E. coli from adult healthy individuals were also studied as controls. As many as 58 uropathogenic strains showed high affinity for attachment to uroepithelial cells while 28 strains showed adherence at moderate degree. Agglutination of human erythrocytes was induced by as many as 70 uropathogenic strains while in 32 strains haemagglutination was not affected by D-mannose. In control group, adherence was observed in eight strains while 28 strains were haemagglutinating. Of these 28 strains, D-mannose resistant haemagglutination was observed in only one faecal strain. In uropathogenic group O4 was isolated with maximum frequency (12%) followed by O101, O135 and O6. PMID- 10862293 TI - Connective tissue stromal changes in tumours and tumour-like lesions of the breast. AB - Between June 1994 and December 1995, one hundred and fifteen tumours and tumour like lesions of the breast were studied in the Department of Pathology, Kasturba Medical College, Mangalore. Neoplasms constituted 80.9% and tumour like lesions accounted for 19.13%. Among the tumours, 51.6% were benign and 48.4% were malignant. Tumour-like lesions included the entire spectrum of fibrocystic disease. Tumours were common between 2nd and 8th decades while tumour-like lesions were uncommon above 6th decade. Whereas benign tumours exhibited ground substance, collagen and reticulin fibres in varying amounts, tumour-like lesions and carcinoma showed more of collagen fibres and less of ground substance. Elastosis and lymphoplasmacytic infiltrate were prominent features in carcinomas rather than in benign lesions. PMID- 10862294 TI - Role of serology in the diagnosis of herpes simplex encephalitis. AB - Twenty one cerebrospinal fluid (CSF)/brain tissue (16 CSF and 5 brain tissue) from patients clinically suspected of herpes simplex encephalitis (HSE) were collected during one year period and was subjected for the detection of HSV type I and type II antigen by direct Immunofluorescence (DIF). Paired serum and CSF samples obtained from 12 patients were tested for herpes simplex virus antibodies by indirect immunofluorescence test. Of the 21 cases, two were positive for HSV-1 antigen in CSF and one in brain tissue by DIF. Virus specific IgG antibody in paired CSF and serum samples was positive in one case only. In none, virus could be grown in verocell line. In 2 Patients antemortem diagnosis was possible and parenteral acyclovir could be administered in one case that recovered following treatment. Thus, besides antibody detection, direct immunofluorescence is an useful and rapid method for the early diagnosis of HSE. PMID- 10862295 TI - Association of tuberculosis with malignancy at KIMIO--an oncology centre. AB - The association of tuberculosis and malignancy was studied at an oncology centre in Bangalore. The study period was from January 1981 to December 1995. A total of 8779 clinical material obtained from patients were screened for Mycobacterium tuberculosis infection. Out of which 675 were positive for acidfast bacilli, 385 from non malignant conditions and 290 from malignant conditions. Highest incidence is seen in Head and Neck cancer (42%) followed by Gastrointestinal cancer (14.1%), Lung cancer (13.8%), Haematological cancer (10.7%), Reproductive cancer (10.3%) and miscellaneous group (9%), Antibiogram of Mycobacterial cultures was done in 282 subjects. Resistance patterns to antitubercular drugs showed highest with Isonicotinic acid hydrochloride (INH) (17.7%), followed by para amino salicylic acid (PAS) (8.5%), Streptomycin (SM) (6.7%), Rifampicin (RIF) (4.6%) and Ethambutol (EM) (0.35%). PMID- 10862296 TI - Clinicopathological correlation of diaphragmatic contraction band necrosis in a neonatal and infantile population--an autopsy study. AB - Contraction band necrosis' is a lesion widely studied in myocardial fibres (Heart) and to some extent in skeletal and smooth muscles. The phenomenon of Diaphragmatic Contraction Band necrosis (D-CBN) occurring in the diaphragms of 53 neonates and infants removed at autopsy were studied microscopically. Of these D CBN was present in 19 (35.85%) cases. D-CBN presented in two morphological patterns (I) solid ('Block-like') segmental necrosis in 31.58% cases or (ii) Ribbon-like transfiber (shredded appearance) bands in 15.79% cases. A combination of both the above lesions were observed in a vast majority of cases, i.e. 52.63% cases. A clinicopathological correlation was attempted as regards cause and mode of death with occurrence and severity of D-CBN. It was found that severe D-CBN was present in the group of birth asphyxia (26.92%) and infant infections (20.0%). The presence and frequency of D-CBN in autopsied subjects proved useful in interpreting the cause and mode of death. PMID- 10862297 TI - Cytologic diagnosis of Enterobius vermicularis eggs in an enterocutaneous fistula. AB - A middle-aged female underwent a laparotomy for suspected ovarian cancer and developed a discharging sinus in the right iliac fossa. Smears of the discharge showed helminthic eggs which were characterised as those of Enterobius Vermicularis. The possibility of an enterocutaneous fistula was suggested which was subsequently confirmed during a relook laparotomy. PMID- 10862298 TI - Fasciolopsis buski (giant intestinal fluke)--a case report. AB - A girl, aged 20 years presented with diarrhoea, vomiting, pain abdomen and loss of weight, the routine Stool examination revealed Fasciolopsis buski (giant intestinal fluke) in large numbers. Despite treatment with Praziquantel, she died after three days. PMID- 10862299 TI - Clear cell hidradenoma of the eyelid: a case report. AB - Sweat gland tumours are extremely rare in the eyelids. We report a case of a clear cell hidradenoma (nodular hidradenoma) in an elderly female, who had presented with a nodular swelling in a eyelid. Clear cell hidradenomas arise as intradermal nodules from eccrine sweat glands. Ultrastructural and enzyme histochemical studies have shown nodular hidradenomas to be intermediate between eccrine poroma and eccrine spiradenoma. No apocrine differentiation has ever been observed in these tumours. Malignant forms are distinctly unusual. This case is being documented for the extremely uncommon presentation of this tumour as an eyelid mass. Complete primary excision is advocated and local steroid preparations should bot be used. PMID- 10862300 TI - Senile systemic amyloidosis--a case report. AB - An 85 years old female presented with acute pain and weakness in left lower extremity and doppler evidence of femoropopliteal block was made which subsequently proved fatal. Necropsy revealed extensive amyloid deposition in the heart and amyloid angiopathy in rest of the organs. PMID- 10862301 TI - FNAC of papillary and solid epithelial neoplasm of pancreas--a case report. AB - A case of solid papillary epithelial neoplasm (PSEN) of pancreas in a young woman is reported in which the nature of tumour was recognised pre-operatively by ultrasound guided Fine needle aspiration. The pre-operative cytologic diagnosis enabled prompt and appropriate surgical treatment. FNAC revealed large cell clumps in the aspirate showing branching papillary appearance in which multiple layers of tumour cells surrounded central vascular stalks. The above was confirmed on histopathological examination of the excised tumour tissue. PMID- 10862302 TI - Coat's disease: an uncommon lesion of eye--a case report. AB - Coat's Disease, first reported in 1908, is a rare disease which is usually seen in young males presenting with complaints of unilateral vision loss. Microscopically, retinal telangiectasis and exudative retinal detachment is seen. Attempts should be made for differentiating and early detection of this disease to avoid enucleation of eye ball. Here we discuss a case report of a child manifesting as coat's disease in which a clinical diagnosis of Retinoblastoma was given and eye was enucleated. PMID- 10862303 TI - Acute pneumonitis with pulmonary hemorrhage an uncommon and potentially fatal complication of systemic lupus erythematosus: a case report. AB - Acute pneumonitis with diffuse alveolar haemorrhage is potentially fatal. When it occurs in a patient of systemic lupus erythematosus, the primary disease itself may be responsible for it; rather than any complicating infection or metabolic/physiological derangement. Diagnosis of primary pulmonary involvement by systemic lupus erythematosus can only be made on open lung biopsy coupled with immunofluorescent and/or ultrastructural studies. Early diagnosis of acute pulmonary complications in systemic lupus erythematosus patients is essential as specific management is reported to improve the chances of recovery. PMID- 10862304 TI - Pneumocystis carnii pneumonia in AIDS--an autopsy report. AB - Pneumocystis carinii pneumonia (PCP), the most common presenting manifestation in patients with AIDS from western countries, holds the distinction for being the first opportunistic infection that was associated with AIDS. There is marked paucity of clinically diagnosed and pathologically confirmed cases of PCP in India. This case represents the first complete autopsy report of pneumocystis carinii pneumonia inpatient with AIDS from our country. A high index of clinical suspicion and microscopic confirmation is needed to avert mortality due to PCP in patients with AIDS. PMID- 10862305 TI - Pure lipoma of the uterus: an extremely rare entity. PMID- 10862306 TI - Acanthameba keratitis. PMID- 10862307 TI - Hepatitis C--a transfusion associated hepatitis. PMID- 10862308 TI - Supravital staining and bright field microscopy: a simple technique for urine sediment. PMID- 10862309 TI - Scorpion--stings the limb and stuns the heart? PMID- 10862310 TI - Scorpion envenomation as a risk factor for development of dilated cardiomyopathy. AB - OBJECTIVES: Though scorpion envenomation is known to lead to acute myocarditis and a reversible decrease in left ventricular function, it has not been implicated as an etiological factor in idiopathic dilated cardiomyopathy. We studied the association of idiopathic dilated cardiomyopathy with a history of scorpion sting as well as with socio-economic status, history of smoking and alcoholism, rural habitation, and history of snake bites. METHODS: Consecutive cases of idiopathic dilated cardiomyopathy were recruited for this study: The association with putative risk factors was studied using a case-control study design with two sets of controls. One set of controls were age and sex matched inpatients selected at random, the other set of controls were spouse, or if not available, a close relative, ordinarily resident with the patient. RESULTS: On analysis, none of the factors except scorpion envenomation had a significant association. A past history of scorpion envenomation had an adjusted odds ratio of 8.01 (3.55-18.06) when compared to one set of controls and an odds ratio of 8.33 (6.55-10.59) when compared to the second group of controls. CONCLUSIONS: Our study indicates that a history of scorpion envenomation acts as a risk factor for the subsequent development of idiopathic dilated cardiomyopathy. Despite an apparently complete recovery from a scorpion sting, many patients probably retain sub-clinical deficits that predispose to the development of cardiomyopathy later in life, when other factors get added on. The known association of cardiomyopathy with catecholamine excess in experimental situations in animal studies, and in other disease states in humans, supports this hypothesis. PMID- 10862311 TI - Prevalence of coronary risk factors in non-insulin dependent (type 2) diabetics. AB - OBJECTIVES: A cross sectional study was conducted to find the prevalence of coronary risk factors in non-insulin dependent diabetic (NIDDM) patients and to compare and co-relate these risk factors in type II diabetics with and without electrocardiographic and/or symptomatic evidence of coronary heart disease (CHD). METHODS: One hundred sixty-seven consecutive NIDDM patients (77 males, and 90 females) attending the diabetic clinic at Dr. RML Hospital, New Delhi were studied. Only known NIDDM cases, already on treatment and without any history of ketosis or congestive heart failure were included. Coronary risk factors comprising of age, gender, duration and treatment for diabetes, smoking, physical activity, hypertension, truncal obesity, lipids, microalbuminuria (semiquantitative) and glycemic control have been particularly ascertained in all the cases. The data was analysed using 'Epi Info version 6.0'. RESULTS: The mean age of patients was 53.12 year and 8.86 year was the mean duration of diabetes. 28.6% of the diabetic men were found to be currently smoking and/or consuming alcohol, 82% were involved in sedentary physical activity and 20.4% had family history of CHD. Central obesity was observed in 46.7% of the cases; more so in females. 31.74% of cases were hypertensive; more females than males had hypertension (33.8% vs 30%). Poor glycemic control (HbA1c > = 9.5%) was seen in 16.8% of the cases. In about 52.5% of the total group hypertriglyceridemia was noted. Microalbuminuria could be found in 35.93%. CHD was diagnosed in 15.57% of cases in this study. CONCLUSIONS: The present study revealed that high levels of serum cholesterol (p = 0.000004), LDL (p = 0.00003), HbA1c (p = 0.002), microalbuminuria (p = 0.000006) and hypertension (p = 0.00006) are significant associates of CHD in NIDDM (both the sexes). Among the female NIDDM cases, in addition BMI (p = 0.01), Waist-hip ratio (WHR) (p = 0.003) and low HDL level (p = 0.008) are important correlates of CHD. Multiple logistic regression analysis was used to allow for confounding between variables. Microalbuminuria alone entered the 'best' model for CHD prediction. Other risk factors, though significant, provided inadequate models for CHD prediction. PMID- 10862312 TI - Effect of fiberoptic bronchoscopy on arterial blood gases and cardiac rhythm at a moderate altitude of 2250 meters. AB - OBJECTIVES: To study the effects of fiberoptic bronchoscopy (FOB) at an altitude of 2250 m on arterial blood gases (ABG) and cardiac rhythm abnormality. METHODS: Fifty consecutive patients undergoing fiberoptic bronchoscopy were evaluated for the arterial blood gases and cardiac rhythm changes at Shimla (a moderate altitude of 2250 m), where there is a state of ambient hypoxia. RESULTS: The changes were noted in five stages ranging from the levels before the procedure till 15 minutes after the completion of the procedure. The mean fall in PaO2 levels in this study was 8 +/- 2.45 mm Hg and the fall was maximum at the end of procedure. Both smokers and nonsmokers showed a significant fall but the fall was more severe in smokers. The mean fall in SaO2 in this study was 3%. The increase in heart rate and blood pressure during FOB was significant as compared to baseline levels. There was no significant change in PH, PaCO2, HCO3. The commonest rhythm abnormality noted was sinus tachycardia which was well tolerated. No major cardiac arrhythmia was noted. It was further seen that the duration of the procedure and type of special procedure undertaken did not effect the levels significantly. Cyanosis was the commonest complication encountered (36%) and was seen more frequently in smokers and those with age more than 40 years. It was observed during the induction of bronchoscope and also during the further negotiation of the bronchoscope into the smaller branches of bronchial tree. CONCLUSION: The changes in ABG and cardiac rhythm are comparable to the studies at sea level except the increased incidence of cyanosis. PMID- 10862313 TI - Prevalence and incidence of type-2 diabetes and impaired glucose tolerance in a selected Indian urban population. AB - AIM: To study the impact of diabetes mellitus on a selected Indian urban population. METHODS: The staff of Indian Institute of Technology in Chennai and their relatives were screened for diabetes by oral glucose tolerance test (OGTT) in 1992. But those on treatment for diabetes were not screened by OGTT. All those found to have diabetes during initial screening were excluded from further follow up. Those without diabetes were followed with repeat OGTT one year later in 1993. RESULTS: A total of 1198 persons, 455 (38.0%) females and 743 (62.0%) males, participated in this study. While 116 (9.7%), 80 (69.0%) males and 36 (31.0%) females, suffering from diabetes were exempted from OGTT, the remaining 1082 (90.3%), 663 (61.3%) males and 419 (38.7%) females, were screened by OGTT. Among the 663 males, 450 (67.9%) were normal, 155 (23.4%) had impaired glucose tolerance (IGT) and 58 (8.7%) had diabetes. Among the 419 females, 275 (65.6%) were normal, 120 (28.6%) had IGT and 24 (5.7%) had diabetes. Out of 1000 persons without diabetes, 696 (69.6%), 444 (63.8%) with normal glucose tolerance and 252 (36.2%) with IGT had participated in the repeat screening by OGTT after one year. One (0.7%) normal person and 14 (5.5%) with IGT, progressed to diabetes in one year. All had type-2 diabetes and non type-1 diabetes. Of 444 normal persons 34 (7.7%) developed IGT during one year. CONCLUSIONS: The prevalence of 9.7% and 7.7% for known and newly detected type-2 diabetes respectively and the annual incidence of 2.2% indicate the magnitude of impact of diabetes mellitus on this population. Though IGT was found to be a risk factor, factors leading to its progression to diabetic state could not be identified in this study. PMID- 10862314 TI - Tumour necrosis factor-alpha and interleukin-2 in pus aspirate and blood in patients with amoebic liver abscess. AB - OBJECTIVES: Cell mediated immunity (CMI), cytokines and humoral immunity have been implicated in the pathogenesis of invasive amoebiasis. METHODS: The role of cytokines--tumour necrosis factor-alpha (TNF-alpha) and interleukin 2 (IL-2) in blood and pus aspirate was studied in 20 patients of amoebic liver abscess (ALA), before and after treatment and 10 controls. RESULTS: The mean TNF-alpha levels (pg/ml) in the controls and before treatment in the patients in serum and pus were 24.3 +/- 11.6, 28 +/- 14.5 and 161.2 +/- 81.3 (p < 0.002) respectively. The mean IL-2 levels (pg/ml) in the controls, serum and pus aspirate in the patients prior to treatment were 10.3 +/- 8.5, 39.2 +/- 26.1 and 117.0 +/- 65.9 respectively. The levels in the patients after therapy, increased to 47 +/- 25.7 (p < 0.001) and 134 +/- 59.4 (p < 0.01). CONCLUSIONS: The higher levels of TNF alpha and IL-2 in the pus aspirate compared to blood pre treatment, supports the role of locally released cytokines in the target organ i.e. liver in amoebiasis. The rise in values observed after therapy are indicative of increased macrophage activity due to CMI occurring late in the course of the disease which may contribute to disease limitation and localisation in amoebiasis. The study suggests that locally released cytokines play an important role in the pathogenesis of ALA. PMID- 10862315 TI - Neurological complications of eclampsia. AB - OBJECTIVE: A prospective study was conducted to evaluate the various neurological (clinical, radiological and EEG) complications in patients of eclampsia. METHODS: Thirty nine patients of eclampsia were studied regarding neurological findings at presentation and electroencephalographic (EEG) tracings were recorded in each patient. Patients with an abnormal neurologic examination and/or focal or lateralizing findings on EEG, underwent a CT scan (n = 18). Foetal and maternal outcome were recorded. RESULTS: The age of the patients ranged from 19-30 (mean +/- SD, 24.2 +/- 3.5) years thirty six patients (92%) had seizures in the antenatal period, 2 (5.4%) patients developed post partum eclampsia and 1 (2.6%) patient had seizures before and after delivery. A diffuse encephalopathy was seen in 9 patients (23.1%), 4 patients (10.2%) had hemiparesis and 1 patient (2.6%) had papilledema. EEG abnormalities were seen in 29 cases (74%) and included generalized slowing (n = 19), generalized sharp waves (n = 9), focal slowing (n = 4), focal sharp waves (n = 2) and spikes (generalized and focal) were seen in 1 patient each. Abnormal CT scan was seen in 10 cases (n = 18). Five patients had generalized infarct was seen in 1 patient each. There were 8 (20.5%) still births and 31 (19.5%) live births and no maternal mortality. CONCLUSIONS: Antenatal seizures occur in > 90% cases of eclampsia and less than 10% cases have seizures after delivery. A diffuse encephalopathy is the commonest clinical abnormality along with generalized slowing on EEG. Although cerebral oedema is common focal infarcts may be seen on CT scan. PMID- 10862316 TI - Clinical profile of headache and cranial neuralgias. AB - OBJECTIVES: The present study, first of its kind from Kashmir Valley, was conducted on 2982 patients to depict the pattern of various headache types and cranial neuralgias. Besides demographic parameters, various factors influencing the frequency of headaches and cranial neuralgias were also analysed. METHODS: Patients presenting with the chief complaint of headache and facial pain were included in this study. The diagnosis of different headache types and cranial neuralgias were established after following the criteria as devised by the International Headache Society (1988). RESULTS: Mean age at presentation for all headache types/cranial neuralgias was 24.5 years (range 7-74 years) with a male:female ratio of 2:1. Tension headache and migraine was found in 1988 (66.6%) and 407 (13.6%) cases respectively. Cranial neuralgias were observed in 20 (1%) cases. CONCLUSIONS: Tension headache and migraine were the commonest forms of headache disorders. Militancy related stress and handicrafts profession were the main predisposing factors for tension and cervicogenic headaches respectively. Ramadan fasting was the prime precipitating factor for migraine. PMID- 10862317 TI - Rapid diagnosis of falciparum malaria by detection of Plasmodium falciparum HRP-2 antigen. AB - OBJECTIVE: Malaria is a resurging problem all over the country and rapid diagnosis is mandatory to decrease the morbidity and mortality and for control of malaria. In the current study the aim was to evaluate the usefulness of rapid Plasmodium falciparum antigen detection and to compare its utility over conventional peripheral thick and thin smear examination. METHODS: Three hundred fifty seven randomly selected patients with pyrexia and or atypical presentations of malaria, found initially negative for malaria were subjected to thick and thin smear examination and Plasmodium falciparum antigen detection test by using commercially available Parasight F. kit. RESULTS: 54.6% of cases presented with pyrexia, while other presentations of falciparum malaria were less frequently encountered (162/357). Eighty five patients (23.8%) were diagnosed as having falciparum malaria based on smear/Parasight F. Test. Eighty- four of these patients were positive for Parasight F. test and only 34.51% of these cases were also positive on smear examination. CONCLUSION: The antigen detection test for Plasmodium falciparum is useful for rapid diagnosis of Plasmodium falciparum malaria. It could detect 65.5% cases of falciparum malaria which were initially negative by peripheral smear examination. Hence, this technique is superior to peripheral smear staining and helps early diagnosis. PMID- 10862318 TI - Need for education on footcare in diabetic patients in India. AB - OBJECTIVES: The patient himself plays the crucial role in the prevention of diabetic foot disease and therefore education on foot care is important. In this study, we have evaluated the knowledge of the diabetic subjects regarding the foot problems and the care of feet in order to identify areas that require stress in the education programme. PATIENTS AND METHODS: Two hundred and fifty, consecutive cases of Type 2 diabetes (M:F, 176:74, age 57.2 +/- 9.7 yrs, duration 12.9 +/- 7.9 yrs) were selected for this study from the out-patient department of our hospital. A questionnaire was filled up for each patient by personal interview. The total score was 100 and a score of < 50 was considered as a low score for foot care knowledge. RESULTS: A score of < 50 was obtained in 67.2%. Low score was more common in women (78.5%) than in men (62.5%) (chi 2 = 5.26, P = 0.022). Low scores (< 50) were more common among those with lower level of formal education (chi 2 = 70.0, P < 0.0001), there were more women with low educational status. Significant foot problems like gangrene, foot ulcers were present in 27.2% and low scores were more common among those with these complications (82% vs 62%) (chi 2 = 8.3, P = 0.004). In general the scores on awareness of general foot care principles and basic facts about the foot complications were poor. Most of them (72%) had good knowledge about the right usage of foot wear. There was a trend to have lower scores with poor formal education (chi 2 = 51.1, P < 0.0001) and also with increasing age. There was no correlation between the scores and the number of hospital visits. Multiple linear regression analyses showed that 31.2% of the variations in the scores were explained by the level of education. CONCLUSIONS: This study underscores the importance of patient education on foot care principles, especially so, considering the magnitude of the problem of diabetes and the lower levels of literacy and poor socio economic status of many patients in this country. PMID- 10862319 TI - Respiratory complications in postoperative patients. AB - OBJECTIVES: This study was conducted to see the extent of respiratory morbidity in the general surgical unit of a teritiary care teaching hospital and to look for probable factors that were responsible for them. METHODS: A prospective study was conducted over a six month period, of patients who underwent both elective and emergency surgeries. Patients were assessed pre-operatively, on the fifth post operative day and at the time of discharge for respiratory complications. RESULTS: Five hundred eighty four consecutive patients who underwent surgeries were studied. Eighty one of them (13.9%) had respiratory complications. Pneumonia was the most common complication (68%). The others included pleural effusion, empyema, pneumothorax and exacerbations of asthma and chronic obstructive pulmonary disease (COPD). One patient developed ARDS (Adult respiratory distress syndrome) and died. Patients who underwent upper abdominal surgery (both elective and emergency), those who had a stay in the surgical ICU for more than 24 hours and those who were on the ventilator for more than 24 hours had a higher incidence of respiratory complications (p < 0.001). CONCLUSION: Respiratory complications increase the morbidity in post operative patients. Pre-operative respiratory illnesses, upper abdominal surgery, ICU stay and mechanical ventilation in the post-operative period predispose patients to respiratory complications. Pre-operative respiratory assessment and treatment of any underlying respiratory disorder is necessary and may decrease the morbidity in surgical patients. PMID- 10862320 TI - Medicine in the next millennium--manpower needs. PMID- 10862321 TI - Artificial liver support systems. PMID- 10862322 TI - Ischemic nephropathy. AB - Atherosclerotic narrowing of the renal arteries may result in severe consequences including chronic renal ischemia, renal artery atheroembolism and renal vascular hypertension. Ischemic renal disease is increasingly recognised as a potentially treatable cause of chronic renal failure. Its precise prevalence is still poorly determined as there is no population based studies. The patients with ARD, particularly those with high grade stenosis and systolic hypertension are at very high risk for renal atrophy and renal failure. Angiogram is usually required to confirm the diagnosis. However, the diagnosis is likely in the elderly patient with systemic atherosclerosis and hypertension in whom a rapid rise in serum creatinine concentration is associated with decreased renal length. Disease is associated with high mortality when treated medically. In contrast, clinical improvement is reported after renal revascularisation. Therefore, consider the diagnosis in the patients at risk, because revascularisation (surgical or endovascular) can successfully preserve renal function in selected patients. PMID- 10862323 TI - Primary hypertrophic osteoarthropathy. PMID- 10862324 TI - Hemiparesis: an uncommon complication of falciparum malaria. PMID- 10862325 TI - Wallenberg's lateral medullary syndrome: a new non-vascular cause. PMID- 10862326 TI - Rupture of sinus of Valsalva aneurysm and coronary arterio-venous fistula. PMID- 10862327 TI - A story of "Ayurvedic" tablets and misled epileptic patients. PMID- 10862328 TI - Alprazolam poisoning. PMID- 10862329 TI - Tropical diabetic hand syndrome--Indian experience. PMID- 10862330 TI - Reversible complete heart block in Grave's disease. PMID- 10862331 TI - Complicated tropical pyomyositis caused by multiple organisms. PMID- 10862332 TI - Diuretics in the management of hypertension. PMID- 10862333 TI - Drug abuse with a difference. PMID- 10862334 TI - Behaviour of healed duodenal ulcer on maintenance therapy: age related. PMID- 10862335 TI - Gas gangrene following intramuscular injection. PMID- 10862336 TI - Trigeminal neuropathy in NIDDM. PMID- 10862337 TI - Leptin--the fat controller. PMID- 10862338 TI - Hepatic artery aneurysm. PMID- 10862339 TI - Exercise induced proteinuria is an early indicator of diabetic nephropathy. PMID- 10862341 TI - Feelings of knowing in the Ranschburg effect. AB - In two experiments, we assessed feelings of knowing (FOKs) for the Ranschburg effect to examine the types of retrieval ease that affect FOKs. In the Ranschburg effect, retrieval performance for repeated items differs from nonrepeated items in supramemory span tasks. We found that FOKs are affected by memory manipulations that affect recall processes, but not by manipulations that affect recognition. This suggests that processes that affect recognition, such as target familiarity, do not affect FOKs, whereas processes that affect recall, such as response suppression and guessing factors, affect FOKs. We propose that an integrated theory of FOKs must include mechanisms responsive to both encoding and retrieval factors (such as retrieval accessibility and cue familiarity), which are highly susceptible to output interference. PMID- 10862340 TI - Automatic and voluntary control of attention in young and older adults. AB - Young and older adults searched for a target character in a 3-item display. On each trial, both a symbolic cue (arrow at fixation) and a spatial cue (abrupt onset of one item) could indicate the target's position. Participants were told to use the central arrow cue on all trials because it had 75% validity. The onset cue also had 75% validity for half the participants and 25% validity for the other half. Both age groups showed about the same cost and benefit effects for the central arrow cues, but the abrupt onsets had much larger cuing effects for older adults. Young adults were able to suppress at least partially an automatic attentional response to an abrupt onset item when the arrow cue preceded the onset and had a higher validity than the onset cue. Older adults appeared to be less able to inhibit their responses to abrupt onsets and to disengage their attention from invalid onset cues than were the young adults. PMID- 10862342 TI - Face-name mediated learning and long-term retention: the role of images and imagery processes. AB - Subjects were presented with videotapes wherein 24 students introduced themselves for 20 s while the subjects were given image mediators as learning aids. After initial recall and before recall after 1 week (Experiment 1) or 3 weeks (Experiment 2), subjects rated whether they thought they did (session 1) or would (session 2) remember each name and rated the clarity of the mental images of the faces, with the names used as cues. The image clarity of faces was significantly related to long-term retention of names, the clarity of images decreased with time, and the process of making ratings and reconstructing mental images of faces and face-name connections greatly facilitated long-term retention of names. The results are discussed in the context of a multiphase processing model. PMID- 10862343 TI - Total time and efficient time management: in search of optimal learning and retention via study-test-rest presentation programs. AB - Bugelski's total time hypothesis (TTH) assumes that a constant time X is needed to learn Y regardless of how time X is programmed, with no possibility of optimization. Izawa's study-test-rest (STR) hypothesis assumes that optimization in learning and retention is possible. Examined were the effects of varied study (S) and test (T) presentation programs on learning and retention, with total time held constant. Participants learned a list of 12 paired associates under three presentation program types. In S + T list repetitions, each pair was presented once per S cycle and tested once per T cycle; both S and T were repeated in list form. In S + T item repetitions, each pair was repeated 1-6 times before the next pair in the list was presented on both S and T cycles. For S/T alternations, the entire list of pairs was studied and then each pair was tested, with S and T cycles alternating. Under each program type, there were four presentation rates. Item repetition led to more learning than list repetition when the presentation rate was very fast. Learning and retention were greatest with the intermediate S/T alternation program. These findings supported Izawa's STR hypothesis but strongly contradicted TTH. PMID- 10862344 TI - Retention of problem solutions: the re-solution effect. AB - Four experiments compared the re-solution performance of prior solvers with that of prior nonsolvers given the correct solutions. Experiments 1 and 2 challenged Weisberg and Alba's (1981) contention that solving a problem and being shown the solution yield equivalent problem knowledge. In both experiments, students who initially solved problems showed near-perfect recall of the solutions after a 1 week delay, far superior to recall by students who had been shown the correct answers. In Experiment 3, solves showed poor solution retention when the connection between the problem and the solution was not meaningful. Experiment 4 showed that with meaningful problems, solvers and those merely provided with solutions have qualitatively different problem representations. The findings can be explained in terms of differential understanding of problems and their solutions. PMID- 10862345 TI - How the challenge of explaining learning influenced the origins and development of John B. Watson's behaviorism. AB - Before he invented behaviorism, John B. Watson considered learning one of the most important topics in psychology. Watson conducted excellent empirical research on animal learning. He developed behaviorism in part to promote research and elevate the status of learning in psychology. Watson was much less successful in the adequacy and originality of the mechanisms he proposed to explain learning. By assimilating the method of classical conditioning and adopting Pavlov's theory of stimulus substitution, Watson linked behaviorism with a new method that could compete with both Titchener's method of introspection and Freud's methods of psychoanalysis. Watson's interest in explaining psychopathology led to the discovery of conditioned emotional responses and a behavioristic explanation for the learning of phobic behavior. Watson established learning as a central topic for basic research and application in American psychology. PMID- 10862346 TI - [The problems in the chemotherapy of tuberculosis taking into account new data on its causative organism]. PMID- 10862347 TI - [The action of its own antibiotic on the producer of imbricin growing on an agarized medium]. AB - The action of imbricin on its own producer Streptomyces imbricatus grown on an agarized medium was studied. Comparatively low concentrations of the antibiotic were shown to have a high lethal action on the streptomycete. The morphological and cultural features of S. imbricatus did not change under the action of imbricin while the variation with respect to the antibiotic production property markedly increased. After the strain exposure to 200 micrograms/ml of imbricin, a stable variant with the antibiotic potency 20 per cent higher than that of the initial organism was isolated. PMID- 10862348 TI - [The effect of fluoroquinolones on the adhesive properties of different plasmidovars of Yersinia pseudotuberculosis]. AB - The effect of fluoroquinolones such as ciprofloxacin, pefloxacin and norfloxacin on adhesion of Yersinia pseudotuberculosis was studied. It was shown that the effect of the fluoroquinolones was different: decreasing or increasing. The same effect was also observed in the closely related strains of Y. pseudotuberculosis. In the strains not dominating in the polyclonal population the adhesion decreased under the effect of the fluoroquinolones. In the strains of the dominant clones the effect on the adhesion was not single valued. PMID- 10862349 TI - [The participation of the transport-barrier functions of the plasma membrane in the development of fluoroquinolone (ciprofloxacin) resistance in Acholeplasma laidlawii]. AB - The role of transport activity of Acholeplasma laidlawii plasmatic membrane in the development of resistance to ciprofloxacin was investigated. It was shown that ethidium bromide used as fluoroquinolone analogue in plasmatic membrane efflux pump was accumulated in ciprofloxacin-resistant cells in much less amount. It was estimated that ethidium bromide efflux depended on temperature, glucose and transmembrane electro-chemical proton potential. Inhibitors of efflux systems -reserpine and verapamil enhanced the ethidium bromide accumulation much more intensively in ciprofloxacin resistant cells. The results of investigation allowed to consider the existence of active efflux system for toxic agents in acholeplasma; in the case of ciprofloxacin-resistant strain these systems are inducible. PMID- 10862350 TI - [The role of antibacterial prophylaxis and therapy in pancreonecrosis]. AB - The research of efficiency of different antibacterial prophylactic and therapy procedures among 89 patients suffering destructive pancreatitis is presented in the article. Determined that optimized tactics of antibacterial prophylactic and therapy with using of such medicines like carbapenems have the important value in cardinal improvement of the results of multi-stage surgical and intensive treatment of the patients suffering pancreonecrosis. PMID- 10862351 TI - [The efficacy of meropenem in treating abdominal sepsis in surgical patients]. PMID- 10862352 TI - [The antibacterial therapy of pneumonias in children. The Commission on Antibiotic Policy, Ministry of Public Health of the Russian Federation and the Russian Academy of Medical Sciences]. PMID- 10862353 TI - [The problems of antibacterial therapy in pediatric practice]. PMID- 10862354 TI - [Khuan Khuanovich Planel'es (Juan J. Planeles) (on the centenary of his birth)]. PMID- 10862355 TI - Molecular cloning and characterization of mouse testis poly(A) binding protein II encoded by the Pabp3 gene, which transcomplements meiotic mutant sme2 of S. pombe. AB - A cDNA clone from a mouse testis cDNA library was isolated by the transcomplementation method using a Schizosaccharomyces pombe meiotic mutant (sme2) that is defective in meiosis I. The cDNA clone isolated has an open reading frame encoding 302 amino acids constituting a protein with a strong similarity to mouse poly(A) binding protein II (mPABII) and bovine poly(A) binding protein II (PABII). PABII is known to bind to the growing poly(A) tail and stimulates poly(A) polymerase, which catalyzes the polymerization of the mRNA poly(A) tail. Northern blot analysis of the cDNA clone identified as mPABII revealed a single transcript of 1.2 kb. This was detectable exclusively in adult testis. Immunohistochemical analysis using a polyclonal antibody demonstrated that mPABII protein was expressed in the nucleus at specific stages from late pachytene spermatocytes to round spermatids. Genetic mapping showed the Pabp3 gene encoding mPABII to be located near position 19.5 on mouse chromosome 14. These results suggest that mPABII might be involved in specific spermatogenetic cell differentiation. PMID- 10862356 TI - Genetic analysis of Bemisia (Hemiptera: Aleyrodidae) populations by isoelectric focusing electrophoresis. AB - Twenty-one whitefly populations in the genus Bemisia were evaluated for genetic variation at 3 allozyme loci. Nine of the 22 populations that exhibited polymorphic loci were subjected to allozyme analysis using a minimum of 10 enzymes, representing 10 to 14 distinct loci. Among those nine variants examined, calculated genetic distances ranged between 0.03 and 0.52, with three main groups emerging from the analysis. One group comprised two closely related Western Hemisphere variants of B. tabaci: type A from California, United States and a geographically proximal population from Culiacan, Mexico. A second cluster contained five collections previously identified as B. tabaci type B and Bemisia argentifolii, while a third group contained a single population from Benin, Africa. The latter two groups were grouped separately from New World populations and are thought to have a recent origin in the Eastern Hemisphere. PMID- 10862357 TI - A phylogenetic view on species radiation in Apodemus inferred from variation of nuclear and mitochondrial genes. AB - Species of field mice (genus Apodemus) are the most common rodents inhabiting woodlands and forests of the Palaearctic region. We examined the cytochrome b (cyt b) gene in mitochondrial DNA (1140 bp) and the interphotoreceptor retinoid binding protein (IRBP) gene in nuclear DNA (1152 bp) in nine species of Apodemus. Based on the genetic variation, the nine species were grouped into four lineages: (1) Agrarius group (A. agrarius, A. peninsulae, A. semotus, and A. speciosus), (2) Argenteus group (A. argenteus), (3) Gurkha group (A. gurkha), and (4) Sylvaticus group (A. alpicola, A. flavicollis, and A. sylvaticus). It was shown that these four lineages diverged within a short period of evolutionary time, suggestive of a radiation event. Soon after the radiation, the Agrarius group was likely to have differentiated again into the species lineages simultaneously. In contrast, the European clade, the Sylvaticus group, radiated rather recently. The relative ratio of the extent of sequence divergence among the four main lineages to that among the members of the subfamily Murinae (including Mus and Rattus) was calculated to be 72.4% in the cyt b gene with transversional substitutions, and 58.5% in the IRBP gene with all substitutions, using the Kimura two-parameter method. The value for the three European lineages was 27.6% in the cyt b gene and 12.3% in the IRBP gene. These results may have a correlation with the notion that deciduous broadleaf forests remained in Central East Asia through the late Tertiary to the present, while those in Europe to a large extent had disappeared by the Pliocene. PMID- 10862358 TI - Insulin-like growth factor-I cDNA gene transfer in vitro and in vivo. AB - Our hypothesis is that gene transfer of an IGF-I CMV-cDNA with cholesterol containing cationic liposomes is an efficient tool for transient transfection of growth factors in vitro and in vivo. In vitro, we transiently cotransfected IGF-I cDNA with a CMV construct and a Lac Z beta-galactosidase cDNA/CMV construct using cholesterol containing cationic liposomes and measured beta-galactosidase and IGF I mRNA and protein. In vivo, we subcutaneously injected 3-month-old male Sprague Dawley rats with IGF-I cDNA and beta-galactosidase cDNA into rat skin. After IGF I and beta-galactosidase were cotransfected into PC12 cells, Northern blot analysis showed that the peak time of IGF-I expression was 2 days for mRNA and 5 days for protein. In vivo, a cDNA/liposome ratio of 1:2 was most effective. IGF-I protein expression in IGF-I-transfected skin resulted in significant transfection from day 5 to day 7. In situ determination of beta-galactosidase activity confirmed that transfections resulted in a restricted expression area. PMID- 10862359 TI - PCR cloning of cerambycidae Parechthistatus gibber (Pg) homeobox genes. PMID- 10862360 TI - Autonomy in physician-assisted suicide. PMID- 10862361 TI - Autonomy in physician-assisted suicide. PMID- 10862362 TI - Health care on Main Street. PMID- 10862363 TI - Separate execution and organ donation. PMID- 10862364 TI - No painless death yet for European euthanasia debate. PMID- 10862365 TI - Justice and managed care. Four principles for the just allocation of health care resources. AB - The debate about justice and health care has occurred largely at a remove from the institutions it concerns; it has been about our most general moral principles, and about what things we value. This debate has foundered. But if the debate is turned in another direction, toward some moral principles that are widely accepted within those institutions, and toward principles that have to do with control over allocation decisions rather than with actually how to make those decisions, agreement may be nearer at hand. PMID- 10862366 TI - The improbable future of employment-based insurance. AB - Voluntary, employment-based insurance is afflicted by a variety of internal imbalances: inequities in the way health insurance is paid for, a conflict of interest in the selection of health insurance, the concentration of the healthy and the sick into separate plans, and free-ridership. But while all these imbalances generate severe problems, those seeking to reform health insurance in the United States should concentrate on the last two. PMID- 10862367 TI - Making room for alternatives. PMID- 10862368 TI - Accountability in science and government: is access the answer? PMID- 10862369 TI - Nondirective counseling or advice? Psychotherapy as value laden. PMID- 10862370 TI - Heroes in bioethics. AB - Throughout his remarkable and too-brief career, Benjamin Freedman was concerned with the ethical standards of ethicists themselves. He worried that ethicists had been bought out, as he knew of none whose opposition to an employer had ever led to being fired. If we look today for what he sought--a bioethical hero--Benjy is himself still the preeminent example. PMID- 10862371 TI - Prayer as therapy. A challenge to both religious belief and professional ethics. The Anglican Working Group in Bioethics. AB - Scientists seeking hard evidence of prayer's curative powers misunderstand the nature of prayer in the Western theistic traditions. Yet theistically consonant ways in which religious belief may influence health do not figure as they should in current professional practice. PMID- 10862372 TI - The Canadian Biotechnology Advisory Committee. PMID- 10862373 TI - Organ wars: another battle. PMID- 10862374 TI - Are pro-welfare state and full-employment policies possible in the era of globalization? AB - There is a widely held belief in U.S. and European economic, political, and academic circles that economic globalization has considerably diminished states' power to follow public policies identified with the social democratic tradition, such as full-employment policies, comprehensive and universal provision of welfare state services, and state regulatory interventions in labor markets and economic policies. And large sectors of the European center-left and left parties believe that European monetary integration made expansionist and full-employment policies practically impossible, except when realized at the European continental level. This article presents empirical information that questions these positions. It documents how specific governments in Europe have been able to carry out such public policies during these years of economic globalization and monetary integration. Some countries (such as Sweden and Finland) that had carried out these policies then later weakened their implementation did so in response to political changes mostly unrelated to globalization of the economy or monetary integration. The article also analyzes and documents how countries that had followed expansionist and full-employment policies have responded to the globalization of financial markets. PMID- 10862375 TI - Excerpts from Joseph Stiglitz's speech to the World Bank, April 1999. AB - The Journal has published several articles critical of the World Bank and the International Monetary Fund. These articles have shown the damage caused by the neoliberal policies advocated by these agencies to the health and quality of life of the people in countries where such policies are carried out. Published here are excerpts of a speech given by Joseph Stiglitz, senior economist of the World Bank, in which he finally recognizes the damage these policies have caused in Russia, where life expectancy has fallen quite dramatically during the years of neoliberal reform. The question triggered by his speech is why the World Bank continues its neoliberal policies. PMID- 10862376 TI - Competition and containment in health care. AB - In the United Kingdom and elsewhere, the preconditions for well-functioning internal markets (in relation to market structure, transaction costs, and information) may not exist in health care. Similar doubts exist about the impact of internal markets on cost-effectiveness. While the quantity of health care has increased, the effects on quality are ambiguous and costs have not been successfully restrained. With respect to equity of health care, fears have been raised that sections of the population may be discriminated against. In the United Kingdom, resources have been shifted away from deprived areas and toward the more affluent. Health care services are once again being reformed, by New Labour in the United Kingdom and similar administrations elsewhere. The rhetoric of competition has given way to talk of partnership. The imposition of new forms of rationing has been reshaped, not abandoned. Additional funding is required, along with an effective commitment to the pursuit of equity and quality in health care. PMID- 10862377 TI - The state, the market, and general practice: the Australian case. AB - This article examines the development of general practice in the latter half of the 20th century, documenting the issues of concern to both the profession and the state. General practice developed hand in hand with the welfare state in Australia. As the structural changes associated with restructuring of the welfare state have advanced, so have the fortunes of general practice declined, despite significant attempts in the 1970s and 1980s to "save" general practice by both the profession and the state. These structural changes have operated on two fronts, the economic and the cultural. On the economic, changes to the employment of general practitioners clearly indicate ongoing proletarianization, particularly in a changing environment of labor-capital relations. At the cultural level, development of the self-help and the women's movements and the elective affinity of these groups with the individualism of the new right are leading to deprofessionalization. The author advances this argument in a review of general practice over the last 40 years and in a case study of community health services. Theoretically he argues for a combination of the proletarianization and the deprofessionalization theses. PMID- 10862378 TI - New labour and Britain's National Health Service: an overview of current reforms. AB - Britain's National Health Service (NHS) has been the subject of unprecedented market reforms, which have failed to solve its problems. The New Labour government elected in 1997 has halted the drive toward the marketization of health care and replaced cost with quality as the central concern of NHS administration and policy. Major changes are occurring in the regulation of professional activity, with profound implications for the medical profession and the health service. The authors discuss these changes and possible future problems for the NHS. PMID- 10862379 TI - Does social policy matter? Poverty cycles in OECD countries. AB - Traditionally, poverty was linked to an individual's family phase. This article examines to what extent poverty cycles are still apparent in OECD countries. By combining data on social policy programs and data on income distribution, the authors compare trends between nations. The main question is, how successful have various sociopolitical solutions been in eliminating poverty? Here the focus is on family policy and pensions. Improvements in social policies have impacts on poverty cycles in all countries. In most countries poverty among the elderly has declined, and the young have replaced the old as the lowest income group. In many countries the poverty cycles have flattened out, and life phase is no longer as important as it used to be. Some differences between nations remain, however. High poverty rates among families continue to be an Anglo-American problem, and improvements in this area have been only marginal. Social policy provisions are important for explaining both cross-national variation in poverty and changes over time. The impact is clearest among pensioners. Family-related poverty is lowest in countries that have combined cash benefits with public child-care services that facilitate parents' participation in the labor market. PMID- 10862380 TI - Legislative proposals for reversing the cancer epidemic and controlling run-away industrial technologies. AB - An interlocking legislative complex is proposed for the control of carcinogenic and other adverse impacts of established run-away petrochemical and radionuclear technologies, with particular reference to winning the losing war against cancer. These proposals are also applicable to the poorly recognized, potentially adverse public health and environmental hazards of emerging technologies, particularly genetically engineered food production. The proposals embody fundamental democratic rights--the right to know and balanced and transparent decision making -the "Precautionary Principle," reduction in the use of toxics, incentives for the development of safe industrial technologies, and criminal sanctions for suppression or manipulation of information. PMID- 10862381 TI - Critical assessment of opposing views on trends in childhood cancer. AB - Two articles reaching opposite conclusions on the current trends in childhood cancer have recently appeared in the literature. One concluded that pediatric cancers have increased dramatically, suggesting an effect from environmental hazards; the other concluded that rates for the major pediatric cancers have remained fairly stable, except for modest increases due to improvements in diagnosis or reporting. This review discusses the reasons for this discrepancy, including differences in the populations, age groups, and time periods analyzed. The arguments in favor of an increase are examined and shown to provide no convincing evidence that environmental pollutants have increased pediatric cancer rates over the past 20 to 30 years. Any suggested increase appears to be the result of non-causal factors, such as selective analysis and reporting, residual confounding by age, random variation, and stepwise improvements in diagnosis and classification. PMID- 10862382 TI - The construction of gender and mental health in Nordic psychotropic-drug advertising. AB - The authors examine the advertisements for psychotropic drugs in the major medical journals of Denmark, Finland, Norway, and Sweden in 1975, 1985, and 1995, with the object of illuminating the gender construction of the portrayed user. Using both a longitudinal and a cross-sectional approach, the study looked for a common Nordic gender display and whether it varied over time. The Nordic journals clearly conveyed a message that psychotropics are a gendered product, but without any uniform pattern. In 1975, men dominated the gender portrayals in Finland and Denmark, and women in Norway and Sweden. In 1985, the pattern was reversed: women dominated in Finland and Denmark, and men in Sweden and Norway. By 1995, the advertisements were mainly for antidepressants, and women were portrayed as the predominant users in Denmark, Finland, and Norway; the Swedish journal displayed couples only. In advertisements with dual-gender positions, however, the focus was on the female; they showed that the drug would assist her in fulfilling the expected supportive female gender behavior. PMID- 10862383 TI - Genetic technologies and achieving health for populations. AB - The remarkable progress in genetics over the last 50 years has led to the development of genetic technologies to identify or alter genes in living organisms, and these technologies can be applied to people. This article presents background information on the role of genetics in human disease, outlines the technologies, and discusses the sources of the strong push for a genetic approach to ill-health and some implications and harmful consequences of using these genetic technologies. The determinants of most diseases are complex and are embedded in a social context. To focus on only one strand of this web--the genetic strand--because it is one that may be amenable to biological/pharmaceutical treatment, although profitable for industry, does not address other important determinants of health and may lead to a harmful overemphasis on genetic approaches. The author outlines some limitations to the potential contribution of genetic technologies to population health across the globe and the need for policy development if these technologies are to have an appropriate place in health care. PMID- 10862384 TI - Popular opinion on tax cuts and Social Security. AB - A review of recent opinion polls reveals the U.S. public's views on budget priorities and Social Security. The public wants more spending on Social Security, Medicare, and other domestic programs, chiefly education and health care, and prefers these spending priorities--by up to a 70 percent majority--to paying down the national debt and cutting taxes. The public supports the Social Security system but doubts it can continue to deliver the goods. To remedy this problem, it is willing not only to use part of the surplus but to raise the cap on payroll taxes. The public does not support benefit cuts or an increase in the retirement age. And the public remains unsure to hostile about the role of the stock market, whether in individual accounts or in the Social Security trust fund. PMID- 10862385 TI - Arthroscopic monopolar radiofrequency thermal stabilization for chronic lateral ankle instability: a preliminary report on 10 cases. AB - This study represents a preliminary review of 10 patients having undergone arthroscopic monopolar thermal stabilization for ankle instability from October 1996 to June 1998. All patients in this study expressed mild to moderate chronic ankle instability complaints and were dissatisfied with their attempts at conservative care. Subjective clinical results were evaluated in all patients having undergone this procedure utilizing a modified version of the American Orthopedic Foot and Ankle Society (AOFAS) Ankle-Hindfoot Scale. In addition, eight of these patients underwent pre- and postoperative stress radiographs. The average age of the patient population in this study was 34.5 +/- 9.26 years. The preoperative AOFAS scores averaged 58.3 +/- 8.96 and the postoperative were 88.1 +/- 11.09 points. Patients returned to full activities on the average of 3 months. Postoperative ankle varus stress test reduced on the average of 2.8 degrees +/- 2.77 degrees, while the anterior drawer measurements reduced 4.8 +/- 1.83 mm. The reduction in anterior drawer test amounted to an approximate 60% decrease in talar excursion postoperatively. All patients who underwent this procedure achieved ankle stability and commented that they would undergo the procedure again. PMID- 10862386 TI - Distal oblique osteotomy of the first metatarsal for the correction of hallux limitus and rigidus deformity. AB - The purpose of this study was to evaluate the effectiveness of the distal oblique osteotomy of the first metatarsal (a triplanar shortening decompression osteotomy) to correct stage I and II hallux rigidus. The osteotomy cut is performed from dorsal-distal to plantar-proximal with an angle ranging from 35 degrees to 45 degrees in the sagittal plane. The capital fragment is then displaced plantarly and proximally and fixed with two screws and the metatarsal head is remodeled. From January 1993 through December 1995, a total of 26 patients (21 females and 5 males) underwent 30 distal oblique osteotomies of the first metatarsal (22 unilateral and 4 bilateral). The mean age of the patients was 54 years and the mean follow-up was 21 months. Patient satisfaction and objective clinical and radiographic measurements were evaluated. Patients' satisfaction was measured postoperatively with a modification of the University of Maryland 100-Point Painful Foot Center Scoring System. The results were: 84% good to excellent; 7% fair; and 9% poor. Radiographic measurements included: intermetatarsal angle mean: preop = 12.2 degrees, postop = 8.6 degrees; proximal articular set angle mean: preop = 11.8 degrees; postop = 10.3 degrees. There was no evidence of avascular necrosis in any of the cases. Clinical findings were: dorsiflexion of the first metatarsophalangeal joint: preop = 22 degrees, postop = 45 degrees; plantarflexion of the first metatarsophalangeal joint: preop = 15 degrees, postop = 18 degrees; hallux purchase power: preop = 2.5, postop = 2.3; pain on the second and third metatarsophalangeal joints, associated with excessive pressure on the central metatarsal heads: preop--present in 10 patients, postop--present in 12 patients; forefoot supination angle: preop = 13 degrees, postop = 7 degrees. PMID- 10862387 TI - Sliding oblique osteotomy for the treatment of hallux abducto valgus associated with functional hallux limitus. AB - This is a retrospective study of 27 patients (35 feet) with hallux abducto valgus associated with hallux limitus who underwent a sliding oblique osteotomy for surgical treatment between August 1997 and June 1998. Radiographic analysis and range-of-motion measurements were evaluated with an average follow-up of 65 days (range, 26-100). Preoperative criteria included < 45 degrees of dorsiflexion of the first metatarsophalangeal joint with weightbearing, no evidence of degenerative joint disease at the first metatarsocuneiform joint, and no previous surgical procedures on the first ray. The average preoperative intermetatarsal angle was 9 degrees, hallux abductus angle 17 degrees, and first metatarsal declination angle 15 degrees. The average postoperative intermetatarsal angle was 6.6 degrees, hallux abductus angle 10.3 degrees, and first metatarsal declination angle 21.7 degrees. Eighteen patients (22 feet) had a follow-up of over 6 weeks, and the first metatarsophalangeal joint was evaluated. The average gain in postoperative range of motion with weightbearing was 22.3 degrees. PMID- 10862388 TI - A double-blind study of the effect on hemostasis of nabumetone (Relafen) compared to placebo. AB - A double-blind, placebo-controlled clinical trial comparing the effect on hemostasis of nabumetone (Relafen) to placebo in patients who were about to undergo forefoot surgery was performed. Aspirin and nonsteroidal anti inflammatory drugs (NSAIDs) are reported to inhibit platelet cyclooxygenase activity, resulting in altered platelet function and thus potentially enhanced bleeding. Nabumetone has been reported to have no effect on platelet aggregation and bleeding time in normal volunteers and in patients who have undergone knee arthroscopy. After fulfilling the inclusion criteria and after a 1-week washout period (acetaminophen controlled), 15 patients were enrolled in the nabumetone group and 15 patients were randomized in the placebo group. Hemostatic parameters [prothrombin time (PT), partial thromboplastin time (PTT), and Ivy bleeding time (IBT)] were assessed at baseline, visit 2, visit 3, and final visit. No meaningful differences were observed between treatment groups in any of the measured hemostatic parameters. No significant adverse events were reported. There was no significant change from baseline for PT, PTT, and IBT in the nabumetone group (PT, p < .06; PTT, p < .64; IBT, p < .17) versus the placebo group (PT, p < .61; PTT, p < .63; IBT, p < .25). The lack of bleeding diathesis and significant prolongation of PT, PTT, or IBT in this study suggests that nabumetone in dosages up to 1000 mg/day can be administered safely in the immediate preoperative period to patients undergoing forefoot surgery. PMID- 10862389 TI - Metatarsal insufficiency fractures in previously undiagnosed osteoporosis patients. AB - This paper, along with a review of osteoporosis, examines 21 patients (15 women and 6 men) who presented to the senior author between May 1997 and January 1999 with unexplained metatarsal fractures. All 21 patients agreed to bone density testing. Twenty of the 21 had bone densities significantly below the mean for corresponding age, gender, and race. The average bone density for the 21 patients was 2.1 standard deviations below the expected mean for the corresponding 30-year old reference population and 1.7 standard deviations below the mean for an age, gender, weight, and ethnicity matched population. Distribution and location of the fractures were also investigated. The Body Mass Indices were calculated for all 21 patients, but did not seem to play a role in the fractures. Pertinent medical histories and possible factors for comorbidity are also presented. In all cases successful treatment consisted of guarded weightbearing in a padded boot. The authors conclude that there is a previously unreported correlation between metatarsal insufficiency fractures and low bone mass in both genders, confirmed by the abnormal bone mineral density testing. They also point out that men should be made aware that they can suffer from this disease. PMID- 10862390 TI - Pedal macrodactyly: coverage of a large defect with a rectus abdominus free flap. AB - The authors report a case of a unique reconstructive approach for an isolated macrodactyly of the lower extremity in an otherwise healthy African male. Surgical treatment included excision and local resection of the affected hypertrophied skin, soft tissue, and bone. A rectus abdominis free-tissue transfer and split-thickness skin graft were used for coverage of the defect. The foot healed without complication, and at 2-year follow-up, the patient had an aesthetically pleasing and fully functional result. PMID- 10862391 TI - Remitting seronegative symmetrical synovitis with pitting edema (RS3PE) syndrome: a review of the literature and a report of three cases. AB - Remitting seronegative symmetrical synovitis with pitting edema (RS3PE) syndrome is a disease that commonly produces symmetrical synovitis and swelling of both the upper and lower extremities. It generally involves the wrists, hands, feet, and ankles of the affected individual. This syndrome most often resembles that of polymyalgia rheumatica and rheumatoid arthritis and usually affects elderly Caucasian males. Serological testing is typically negative except for a mild to moderate elevation of the erythrocyte sedimentation rate. The HLA-B7 phenotype is present in approximately 50% of patients with this syndrome. Treatment of RS3PE syndrome is heralded by the predictable response to low-dose corticosteroid or hydroxychloroquine therapy. There has been no previous mention of this condition in the podiatric literature. Presented below is a review of this syndrome and three case studies. PMID- 10862392 TI - The accessory soleus and recurrent tarsal tunnel syndrome: case report of a new surgical approach. AB - The accessory soleus muscle is a rare anatomic variant, which presents as a mass in the posteromedial aspect of the ankle. This anomaly has been linked with compression neuropathy of the posterior tibial nerve. The authors present a case of tarsal tunnel syndrome in which the presence of an accessory soleus was unrecognized at the time of the original procedure, but was utilized during the revisional operation to provide safe coverage of the posterior tibial nerve. PMID- 10862393 TI - Fracture of the entire posterior process of the talus: a case report. AB - A fracture of the entire posterior process of talus is rare. In this case report, the authors present a case of isolated displaced fracture of the entire posterior process, which was treated by early open reduction and internal fixation through a posterolateral approach. The fracture healed without evidence of avascular necrosis of either the posterior fragment or the talar body. The aim of this article is to highlight the importance of early internal fixation of this uncommon type of fracture. PMID- 10862394 TI - [The intraspecific taxonomic structure and identification of closely related species of ixodid ticks (Ixodidae)]. AB - Two variants of intraspecific morphological differentiation of 7 palearctic species of Ixodidae and respectively two variants of their intraspecific taxonomic pattern have been revealed on the basis of the study of geographic variation of each species. Three species--Ixodes pavlovskyi Pom., 1946, Haemaphysalis erinacei Pavesi, 1884, Hyalomma asiaticum Sch. et Schl., 1929, are subdivided into subspecies, and four ones--I. ricinus (L., 1758), I. persulcatus Sch., 1930, I. crenulatus Koch, 1844, Dermacentor marginatus (Sulzer, 1776),- into morphotypes by the degree of distinction of the whole complex of active stages in ontogenesis. Acknowledging the conventional character of the categories of subspecies and morphotype their criteria have been formulated for ixodid ticks. Paleogeographic and ecological grounds are given. Some data of geographic variation of immature stages favour the identification of closely related species in concrete localities. PMID- 10862395 TI - [Resting nymphal stages and the nature of individual development in trombiculid mites (Acariformes: Trombiculidae)]. AB - A morphological expression of quiescent nymphal instars (calyptostases) in trombiculid mites (Trombiculidae) is analysed. The analysis takes into consideration ontogenetic functions of different instars during the individual development of these mites, which includes an alternation of active and regressive quiescent instars. Cyclic start and blocking of different integumental differentiation programs in Trombiculidae is caused by the cyclic reduction and renewal of particular functions of respective instars, whereas internal organs of these mites do not transform significantly during the life cycle. The reduction of some functions, particularly the locomotion and feeding, in the quiescent nymphal instars with changed integumental function has an inevitable result in a complication and prolongation of the moulting process. The moulting of these instars begins at once after finishing the previous moulting cycle, i.e. it is realized in "automatic pattern" without the intermoult phase. In general, the organization of both quiescent proto- and tritonymph is the same, because it expresses concordant morphological correlations and synchronous reductions of certain functions and structures in these instars during the evolutionary process. At the same time, the trombiculid mites and, perhaps, other representatives of Parasitengona reveal the most generalized type of ontogenesis in Acariformes within the "alternating calyptostasy" phenomenon. PMID- 10862396 TI - [The development of the genital system in Ichthyocotylurus variegatus (Trematoda: Strigeidae)]. AB - The development of Ichthyocotylurus variegatus from metacercariae to ovigerous adults in the natural definitive host (Larus ridibundus) was examined at 24 h intervals. The metacercariae develop to the adults through a somatic growth of hindbody; the gametogeny and vitellogenesis take 4-6 days. During the first day of development the pair of testes are separated from the genital anlage. The ovary differentiates one day later. Genital ducts are completely differentiated on 4th day after the beginning of spermatogenesis and cogenesis. The vitelline cells developed from 2nd to 5th day. They take an origin from the subtegumental cells of hindbody. The hind part of uterus in strigeoid adults is identical to the metraterm of other trematodes. The spermaduct along all its length works as a spermatic bladder. PMID- 10862397 TI - [The occurrence of monogeneans of the subfamily Capsalinae (Capsalidae)- parasites of marine fishes]. AB - The data on an occurrence of capsalines in marine fishes are analyzed. 33 capsaline species among 55 recently known species are monoxienous, that points to the high specialization of these monogeneans. The majority of species are recorded from three subfamilies of perciform fishes: Scombridae, Istiophoridae, Xiphiidae. Possible factors causing the high specificity in monogeneans are discussed. PMID- 10862398 TI - [The role of fish parasites in fresh-water ecosystems exemplified by a parasite of the smelt (Osmerus eperlanus)]. AB - The European smelt Osmerus eperlanus had been accidentally introduced into the ecosystem of the Syamozero Lake (Karelia). The population of this species has achieved a high density and caused serious changes in the structure and trophic relationships of fish community of the Syamozero ecosystem. The microsporidia Glugea hertwigi Weisenberg, 1921 has become a new and super-dominant parasite of the european smelt in this ecosystem. The invasion of microsporidia has caused a mass death of fishes, that has led to changes in population structure of the smelt and lowered a fish catch. The present study suggests to show a role of parasites in the ichthyocenosis structure regulation in freshwater ecosystem. PMID- 10862399 TI - [The ultrafine morphology of the integument of thorny-headed worms (Acanthocephala)]. AB - Different cited and author's data on the ultrastructural morphology of tegument in Acanthocephala are analyzed and discussed. PMID- 10862400 TI - [The mosquito fauna of Smolensk and Kaluga provinces]. AB - The mosquitos collected in Smolensk and Kaluga provinces in 1985 by the expedition team of the D. I. Ivanovskii Institute of Virology were identified by V. N. Danilov in 1986. Among the females of mosquitos collected in Smolensk Province four genera and 16 species were recorded; two genera (Culiseta, Mansonia) and 9 species were new ones for this region. Taking in account the reference data (excluding Anopheles hircanus and Aedes behningi, the presence of which here is doubtful) there 22 species of mosquitos of five genera (Anopheles- 3, Aedes--16, Mansonia, Culiseta, Culex--one of each) are recently known in Smolensk Province. Among the females of mosquitos collected in Kaluga Province, two genera and 8 species were found including one genus (Mansonia) and three species being a new ones for this region. Together with the reference data, there 17 species of mosquitos of four genera (Anopheles--2, Aedes--13, Mansonia, Culex- one of each) are known now for Kaluga Province. PMID- 10862401 TI - [The manifestation of valiporiasis in cultivated fishes]. AB - Lethal cases of valiporiasis in the silver carps of the age 3 and 4 years are described. The resistance of hybrids between different forms of Cyprinus carpio to the infection with Valipora campylancristrota is discussed. PMID- 10862402 TI - [The fertility of thorny-headed worms in the genus Echinorhynchus (Acanthocephala: Echinorhynchidae) from Lake Baikal]. AB - The total number of eggs (absolute fecundity) and the number of eggs per 1 gr of body weight (relative fecundity) of Echinorhynchus borealis Linstow, 1901, E. salmonis salmonis Muller, 1784, E. s. baicalensis Bogolepova, 1957 and E. truttae Schrank, 1788 from the Baikal lake was examinide. The absolute fecundity decreases in the row or species: E. borealis (26,200 eggs), E. s. salmonis (110,113), E. truttae (8123), E. s. baicalensis (6087). The fecundity of acanthocephalans depends on the host species. The dependence of the whole number of larvae upon the body length of females (r = 0.804) is shown. PMID- 10862403 TI - [The developmental cycle of the trematode Glypthelmins rugocaudata (Plagiorchidae) under the conditions in the Maritime Territory]. AB - It is found out, that the life cycle of the trematode Glyphthelmins rugocaudata (Yoshida, 1916) in the Primorye region includes two intermediate hosts (mollusc Lymnaea pacifampla and dragonfly larvae of the genera Cordulia and Lestes) and a final host, frog species Rana migrimaculata and R. semiplicata. PMID- 10862404 TI - [Gyrodactylus onegensis sp. n. (Monogenea)--a parasite of the bullhead (Cottus gobio L.)]. AB - A new species, Gyrodactylus onegensis sp. n., is described from gills of the freshwater sculpin Cottus gobio. PMID- 10862405 TI - [The results of a study of the species composition of fish parasites in the river basins of northeastern European Russia. Leeches (Hirudinea), mollusks (Mollusca), crayfish (Crustacea) and arachnids (Arachnida)]. AB - The report contains the results of faunistic study of parasitic Hirudinea, Mollusca, Crustacea and Arachnida occurring on fishes in the region of the S. Dvina, Mezen' and Pechora river basins. The parasite fauna in the first basin is represented by 3 species of leeches, 4 species of molluscs, 13 species of crustaceans and 1 species of water mites, in the second basin--4, 1, 11, 0; in the third basin--2, 7, 7, 1 species respectively. Total number of parasites in the region investigated: 4 leech species, 5 molluscs species, 16 crustacean species and 1 water mite species. PMID- 10862406 TI - Small axons relative to number of myelin lamellae in Charcot-Marie-Tooth disease 1A with peripheral myelin protein 22 gene duplication. AB - In myelinated fibers of the sural nerves in Charcot-Marie-Tooth disease 1A (CMT 1A), with peripheral myelin protein (PMP) 22 gene duplication, it has been controversial whether axonal attenuation occurs or the myelin sheath thickens. Therefore, the relationship between the transverse axonal area and the number of myelin lamellae was morphometrically studied in myelinated fibers of the sural nerves in CMT 1A with PMP22 gene duplication to re-evaluate such relationships as are revealed in hereditary motor sensory neuropathy, type 1 without genetic diagnosis. In electron microscopic studies both the axonal circumference and natural log (ln) axonal area were significantly (P < 0.01) smaller in the CMT 1A group (n = 8) than in the control group (n = 9). Myelin lamellae numbers, however, were similar in both groups. In a regression analysis relating ln axonal areas to the numbers of myelin lamellae, ln axonal area in the CMT 1A group was significantly (P < 0.05) smaller than in the control group in myelinated fibers with the numbers of myelin lamellae equal to or over 40. Therefore, it was concluded that the axonal area was smaller in large-diameter myelinated fibers in the CMT 1A group than in the control group. We speculated on the existence of axonal attenuation due to the impaired effect of Schwann cells on myelinated axons in CMT 1A with PMP22 gene duplication. PMID- 10862407 TI - Investigation of possible involvement of several genes related to development of hepatocarcinogenesis in rats. AB - A comparative study on the possible involvement of several genes in the susceptibility of chemical carcinogenesis was carried out using carcinogen resistant DRH rat and -sensitive Donryu and F344 rats. Previously, we observed that the induction of glutathione S-transferase placental form (GST-P) in the liver of Donryu rats by 3'-methyl-4-dimethylaminoazobenzene (3'-Me-DAB) was significantly greater than that of DRH rats. In the present study, we tentatively determined base sequences of the enhancer region including GPE-I and GPE-II (GST P enhancers I and II) of GST-P genes of DRH, Donryu and F344 rats, but we did not observe any nucleotide polymorphism around these regions. Furthermore, the mRNA levels of silencer binding protein (NFA-1) for the GST-P promoter of rat liver were also similar in the DRH and Donryu rats. Since clonal expansion of putative preneoplastic GST-P-positive foci in the DRH rat liver was significantly suppressed during 3'-Me-DAB administration, we examined whether two opposite growth controlling factors, TGF-alpha and TGF-beta, may participate in such suppression of growth. It was supposed that mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R), at least in part, activates TGF-beta preproprotein. However, we observed that the levels of M6P/IGF2R mRNA in the livers of DRH were not higher than those of Donryu rats after being fed 3'-Me-DAB for 8 weeks. Another important factor in the carcinogenesis is insulin-like growth factor II itself. Although liver tumors induced by 3'-Me-DAB in F344 had high levels of IGF-II mRNA, little IGF-II gene expression existed in normal adult livers of Donryu, F344 and DRH rats. High levels of IGF-II mRNA were detected similarly in the livers of neonates from all these three strains of rats. Finally, we detected a significant increase of AFP (alpha-fetoprotein) mRNA in the livers of Donryu rats around 6 to 8 weeks from the start of 3'-Me-DAB feeding, which is in parallel with detrimental effects of this carcinogen on these rats. A reduced induction of AFP mRNA was observed in DRH rats under the same conditions. Further study will be needed to explain the lower tumor susceptibility in the DRH rat. PMID- 10862408 TI - Concentrations and consequent radiation doses of uranium-238, thorium-232 and potassium-40 contained in the front glass of CRTs. AB - Concentrations of 238U, 232Th, and 40K contained in the front glass of CRTs (cathode ray tube) of three television sets are measured by gamma-ray spectrometry on the assumption of radioactive equilibrium in uranium and thorium series. The concentrations of 238U and 232Th are also measured by neutron activation analysis. The obtained results by the two different methods agree well within experimental uncertainty, showing that the radioactive equilibrium is found to be established in the present samples, and that the concentrations of 238U and 232Th in the glass can be measured by the gamma-ray spectrometry as well as by the neutron activation analysis. We also found that although the obtained concentrations of 238U (6.5-13.5 ppm in weight) and 232Th (8.4-15.1 ppm) vary considerably between these three kinds of CRTs, those of 40K (average: 7.45 +/- 0.05 ppm) are almost equal for all samples. Moreover, the radiation dose rates are estimated for gamma-rays from the CRT. At distances larger than 30 cm from the surface of the front glass, even the dose rate (5.65 x 10(-3) microSv/h) for the CRT containing the highest concentration of 238U and 232Th is smaller than one-tenth of that (7 x 10(-2) microSv/h) for background radiation, and that is practically negligible. PMID- 10862409 TI - Cardiac and peripheral vascular responses to head-up tilt during whole body thermal stress. AB - During acute orthostatic stress, neurally mediated control of cardiac output (CO) and total peripheral vascular resistance (TPR) play an important role in the maintenance of systemic blood pressure. To examine the influence of thermal stress on the CO and TPR responses to orthostatic stress, 10 healthy male volunteers were exposed to normothermic control conditions followed by whole-body thermal stress produced by a cold or hot water-perfused suit during 5 min-70 degrees head-up tilt (HUT). HUT increased mean arterial pressure (MAP) by 3% of the pre-tilt value during normothermic control and cooling, whereas it decreased MAP by 4% of the pre-tilt value during heating. HUT decreased CO by 16-17% of the pre-tilt value under each thermal condition. The increase of TPR during HUT was exaggerated during cooling and inhibited during heating compared to normothermic control. Tilt-induced decrease of skin blood flow was greater during heating than cooling. These results suggest that the smaller increase of TPR rather than the CO change is responsible for the decreased MAP during acute orthostatic stress in hyperthermic humans. The contribution of skin vascular constriction to TPR changes during HUT is increased during heating and decreased during cooling. PMID- 10862410 TI - [Contamination of a bronchial fiberscope by mycobacteria linked to an automated bronchoscope disinfection machine]. AB - Mycobacteria are being isolated with increasing frequency from automated bronchoscope disinfection machines. This has led to misdiagnosis and nosocomial infections. In this study, we isolated Mycobacterium chelonae from a bronchoscope disinfection machine and found one strain to be resistant to 2% glutaraldehyde and sensitive to 70% ethanol. Since we began cleaning the sink of the machine with 70% ethanol, no mycobacteria has been seen throughout the machine. PMID- 10862411 TI - [Occupationally induced hydrofluoric acid burns: an analysis of 9 patients from the aspect of occupational health]. AB - We report here 9 patients suffering from hydrofluoric acid burn who visited our clinic from July, 1979 to February, 2000. These 9 cases occupied 25% of all chemical burn cases experienced in our clinic. All the patients were men ranging in age from 20 to 53 (mean age 35 years; average 36.8 years). At the time of accidental exposures, 6 patients had been engaged in washing or cleaning work, and 2 had been changing the parts of instruments containing hydrofluoric acid. Eight patients received burns on the hands and/or fingers. During the work, 2 patients had used vinyl chloride or rubber gloves, but three patients employed no protection for the hands. After the symptoms began to develop, it was found that the glove of one patient had a pin hole. Coupled with the occasions described in previous reports, the causal factors of hydrofluoric acid burn could be divided as follows: 1) negligence or carelessness of workers, in particular skilled persons, in handling hydrofluoric acid, 2) ignorance of the dangerousness of hydrofluoric acid, 3) the presence of pin hole (s) in protection gloves, and 4) unexpected accident. Hydrofluoric acid is one of the most corrosive inorganic acids, and can produce progressive and serious tissue necrosis with severe pain. To prevent burns due to this chemical, enlightenment and reeducation of the workers regarding the hazard of hydrofluoric acid are necessary. PMID- 10862412 TI - [A case of occupational contact dermatitis due to hydroxylamine]. AB - A 36-year-old man, working in a chemical industry, had a generalized pruritic eruption. A forty-eight hour patch test revealed positivity for 1% hydroxylamine. Prevention of exposure to this chemical resulted in a dramatic improvement of the symptoms. Based on these findings, we diagnosed this case as occupational contact dermatitis due to hydroxylamine. There has been few case reports of contact dermatitis due to hydroxylamine. Histopathological examination revealed a marked spongiosis and a spongiotic bulla formation in the epidermis and follicular infundibulum, suggesting an allergic reaction. PMID- 10862413 TI - [A case of squamous cell carcinoma that developed on chronic tar dermatosis]. AB - A 79-year-old Japanese man visited our clinic for evaluation of a large tumor on the left wrist and multiple keratotic tumors. He had handled creosote oil, which is a purified product of coal tar production, for 50 years. Physical examinations revealed poikiloderma with multiple hyperkeratotic tumors on the back of his hand and forearm bilaterally, and a cauliflower-like tumor, 80 x 60 x 15 mm in size, on the left wrist. Histopathologically, the large tumor showed a proliferation of atypical squamoid cells with many keratinization foci, indicating well differentiated squamous cell carcinoma. PMID- 10862414 TI - [Hyperferritinemia and diseases]. AB - Ferritin is the principal iron storage protein participating in iron metabolism. As serum ferritin levels often reflect the amount of storage iron in the body, physicians have measured serum ferritin in order to evaluate iron deficiency or overload. Although a rise in serum ferritin concentration occurs in iron overload, hyperferritinemia without it has been reported in some inflammatory diseases and malignancies. Some cytokines have been reported to be responsible for the elevation of ferritin production. Studies on serum isoferritin in adult Still's disease and other diseases, especially measurements of the proportion of glycosylated ferritin, have been widely accepted. Pathophysiological properties of the increased serum ferritin are not clear. However, we should be aware that the hyperferritinemia is not a result, but is profoundly participating in the disease process. PMID- 10862415 TI - Toxins on the firing range. PMID- 10862416 TI - Biohazards--emerging diseases. A Plum of an Island. PMID- 10862417 TI - Soothing the inflamed brain. PMID- 10862418 TI - SNPs of disease. PMID- 10862419 TI - Asthma worldwide. PMID- 10862421 TI - Tantalizing tubes PMID- 10862420 TI - Paleontology's Indiana Jones. PMID- 10862422 TI - Waging a new kind of war. A scourge of small arms. PMID- 10862423 TI - Waging a new kind of war. Invisible wounds. PMID- 10862424 TI - Waging a new kind of war. Children of the gun. PMID- 10862425 TI - Dwarf galaxies & starbursts PMID- 10862426 TI - Cell communication: the inside story. PMID- 10862427 TI - Reading the bones of La Florida. PMID- 10862428 TI - Computing with molecules. PMID- 10862430 TI - Home is where the ECG is. PMID- 10862429 TI - Looking for life below the bottom. PMID- 10862431 TI - Mathematical recreations. Paradox lost PMID- 10862432 TI - The Internet as hardware PMID- 10862433 TI - Are cirrhosis and its complications inevitable in hepatitis C virus infection? PMID- 10862434 TI - Modern treatment of ovarian cancer. PMID- 10862435 TI - Management of deep vein thrombosis and pulmonary embolism. PMID- 10862437 TI - Setting up a pain management programme. The Ayrshire experience. AB - A controlled trial of an outpatient cognitive behavioural pain management programme for sufferers of non-cancer chronic pain is described. A multidisciplinary team set up a programme of ten half day sessions for groups of ten to fourteen patients aiming to improve activity levels and control over pain; to reduce maladaptive pain behaviours and drug intake; to mitigate negative mood; to modify unhelpful beliefs and to maintain treatment gains by operant and cognitive methods. Self report questionnaires were employed before and six weeks, six months and one year after the programme. Fifty-eight patients entered the study group and 39 patients completed the programme and initial follow up with further attrition in long term follow up. There were no changes in the waiting list control group of twelve subjects but the study group made significant short and long term improvements in pain severity, activity levels, mood, coping and experienced fewer catastrophizing thoughts. PMID- 10862436 TI - Optimising direct access ECHO referral in suspected heart failure. AB - The objective was to prospectively validate a method of increasing the sensitivity, specificity and negative predictive value of a normal ECG in the exclusion of left ventricular systolic dysfunction by the addition of clinical history. We performed a prospective three year study of all referrals to our direct access ECHO service for assessment of LV function. The ECG was reported blind of the result of the ECHO, history of MI or not was noted, and result of the ECHO predicted. Over three years 416 patients were assessed for the presence or absence of left ventricular systolic dysfunction and consequent changes in clinical management. A total of 320(77%) of patients referred with suspected left ventricular dysfunction were found to have normal left ventricular function. Of the 250(60%) patients treated prior to referral for assessment, 183(73%) were treated inappropriately. The combination of a normal ECG and a negative history of myocardial infarction had a sensitivity of 98% and a negative predictive value of 99% in the assessment of LV function. This was an improvement over a normal ECG alone. Our study shows that diagnosis and treatment of heart failure in the community remains sub-optimal. The combination of a normal ECG and no previous history of myocardial infarction is shown to be a sensitive and accurate predictor of normal left ventricular function. If adopted by general practitioners this would be a valuable method of optimising the use of echocardiography in patients with suspected left ventricular dysfunction. PMID- 10862438 TI - Response to open access endoscopy findings by general practitioners guidelines need education for implementation. AB - General practitioners may gain valuable information from the use of open access endoscopy. The benefit to the individual patient depends on the interpretation of the endoscopy findings and the subsequent action. The aim of the study was to determine GPs response to open access endoscopy findings of three conditions with possible malignant complications: Barrett's oesophagus, gastric ulcer and colonic adenomatous polyps. The study took place at Ninewells Hospital, Dundee. Using the endoscopy unit's records for the year, 1 January 1995 to 31 December 1995, all patients having had an open access upper gastro-intestinal endoscopy or sigmoidoscopy were identified. Case-notes were reviewed of patients who had Barrett's oesophagus, gastric ulcer or colonic polyps diagnosed. During the year, 1158 upper gastro-intestinal endoscopies and 293 sigmoidoscopies were performed by the open access service. The referral rates for the conditions were as follows: Barrett's oesophagus 56%; Gastric ulcers 56%; Adenomatous polyps 88%; Non adenomatous polyps 12.5%. The provision of guidelines does not ensure a high referral rate, education is a vital partner. PMID- 10862439 TI - A reappraisal of the role of chlorambucil in patients with end stage ovarian cancer who have previously been treated with platinum regimens. AB - The role of chlorambucil in end stage platinum resistant epithelial ovarian cancer was evaluated in women with end stage ovarian cancer. They had received platinum based chemotherapy and all other intravenous chemotherapeutic options had been exhausted. Over a 15 year period, 30 patients were identified. The median age was 64.5 years (range 45-81). The median number of chlorambucil pulses was 4 (range 1-16). The median survival following the introduction of chlorambucil was 5.5 months (range 0.72-38.8). The 22 patients who survived for longer than three months were significantly younger than those who did not (p = 0.03). Apart from two patients who developed transient myelosupression there were no toxic side effects. Chlorambucil should be considered as a therapeutic option in end stage ovarian cancer. It is has minimal toxicity, and can be prescribed safely for long term use. In younger women, an increase in benefit may be anticipated. PMID- 10862440 TI - Subacute gastric volvulus--a rare cause of vomiting in the elderly. AB - We report a case of an important and uncommon cause of vomiting in an elderly female patient who had no previous apparent gastrointestinal problems. A diaphragmatic hernia with gastric volvulus, which presented non-specifically but was an important diagnosis to make. PMID- 10862441 TI - Neuroendocrine pancreatic cancer: an unusual case of pancreatitis. AB - Acute pancreatitis is a common clinical problem. This case of acute pancreatitis was due to a most uncommon underlying aetiology, a non-functioning neuroendocrine tumour of the pancreas. PMID- 10862442 TI - Excision of a remarkable tumour of the upper jaw in 1834 by Robert Liston. AB - A series of pre-operative casts of the head, one of plaster of Paris and the other of wax, have recently been discovered in the Department of Anatomy, Edinburgh, of a patient with an immense tumour of the left maxillary antrum which produced an enormous degree of facial distortion. These casts complement a series of engravings published in the contemporary literature. This lady's tumour was successfully excised by Robert Liston in 1834 in the Royal Infirmary of Edinburgh, only a month before he left Edinburgh for London. The tumour was believed to be benign, and was removed without the benefit of anaesthesia. The patient returned the following summer to have a gold palate fitted, and while her voice was initially indistinct, it subsequently recovered. PMID- 10862443 TI - Inverted papilloma. AB - BACKGROUND: Inverted papilloma, a benign sinonasal lesion, constitutes 0.5% to 4% of all nasal tumors. Its local aggressiveness, high recurrence rate, association with malignancy or malignant transformation, and tendency toward multicentricity lead most physicians to advocate radical surgery. Nonetheless, conservative surgery is effective for selected patients with limited disease. This study was done to reassess the efficacy of both radical and conservative surgery. METHODS: The records of all patients with the diagnosis of inverted papilloma treated at the Taipei Veterans General Hospital between January 1986 and October 1998 were collected. A minimal follow-up period of 12 months was required for inclusion in the study. Clinical manifestations, radiologic findings, methods of treatment, pathology reports and recurrence rates were all retrospectively reviewed. RESULTS: Sixty patients with an average age at diagnosis of 58 years were studied. The follow-up period ranged from 12 to 195 months, with a mean of 43 months. The most common presenting symptom was unilateral nasal obstruction. The duration of symptoms ranged from one month to 30 years, with a mean of 36 months. The lateral wall and middle meatus were the most commonly involved sites. Bone erosion and intracranial or intraorbital extension were observed in some patients. Medial maxillectomy was the most common surgical treatment, followed by functional endoscopic sinus surgery and the Caldwell-Luc operation. The overall recurrence rate was 23%, with an average interval from initial treatment to recurrence of 42 months (range, 2-93 months). The recurrence rates for the two groups undergoing medial maxillectomy and conservative surgery were 16% and 27%, respectively. CONCLUSIONS: Early diagnosis relies on high suspicion and biopsy. The treatment modality is related to the location and extent of the disease, which is assessed by preoperative imaging studies. Any tumor with extensive involvement of the sinuses or inaccessibility to the endoscope should be treated with traditional medial maxillectomy for best control. The endoscopic technique is good for preoperative biopsy and follow-up. PMID- 10862444 TI - Changes in superoxide dismutase activity and mRNA in vivo after short-term supplementation with trilinolein in rats. AB - BACKGROUND: Oxidative damage plays a central role in atherogenesis and antioxidation defense mechanisms may prevent atherosclerosis. This study evaluated the effect of short-term supplementation of the natural lipophilic antioxidant trilinolein on superoxide dismutase (SOD) activity and SOD-mRNA gene expression in vivo in rat vital organs. METHODS: Male Wistar rats (n = 8) were injected intraperitoneally with trilinolein (1 mM/ml/kg/day in 0.5% ethanol) daily for three consecutive days. Two control groups (n = 8) were administered saline or 0.5% ethanol in saline, respectively, for three days. RESULTS: Assay of SOD activity and SOD-mRNA by Northern blotting in rat liver, spleen and brain showed significant increases in SOD activity and increased SOD-mRNA gene expression. CONCLUSIONS: The natural lipophilic antioxidant trilinolein potentiates the SOD antioxidation defense mechanism and increases gene expression of SOD-mRNA after short-term supplementation in rats. PMID- 10862445 TI - Meropenem versus imipenem/cilastatin in the treatment of sepsis in Chinese patients. AB - BACKGROUND: Meropenem and imipenem are beta-lactam antibiotics of the carbapenem group. Carbapenems have bactericidal activity against a broad spectrum of bacteria, including most gram-positive cocci, gram-negative bacilli and anaerobes. Experience in using meropenem in Chinese patients has not been previously reported. METHODS: Meropenem (2 g daily) and imipenem/cilastatin (2 g daily) were compared in an open, randomized, prospective study on the treatment of hospitalized Chinese septic patients. All participants (male or female) were hospitalized with a diagnosis of sepsis. All patients were randomly allocated to one of the two treatment groups: the meropenem group or the imipenem/cilastatin group. Clinical status was evaluated daily during treatment and at the end of therapy or when treatment was withdrawn. Patients were checked every day for potential side-effects, according to subjective and objective symptoms. RESULTS: Fifty-three patients were enrolled in the study; 50 were evaluated for clinical efficacy and 27 patients were evaluated for bacteriologic efficacy. The most frequent clinical diagnoses were pneumonia and urinary tract infection. The predominant pathogens were Pseudomonas aeruginosa, Escherichia coli and Enterobacter cloacae. There were 31 pathogens isolated from 27 patients. A single pathogen was identified in 23 patients, and two pathogens were isolated from four patients. Satisfactory clinical outcome (excellent and good) was 84% in the meropenem group and 76% in the imipenem/cilastatin group. Satisfactory bacteriologic response was 80% in the meropenem group and 75% in the imipenem/cilastatin group. Transiently elevated liver enzymes were the most common side-effect. One patient treated with imipenem/cilastatin experienced a seizure during the study, while another patient treated with meropenem withdrew due to urticaria. CONCLUSIONS: The efficacy and safety data presented in this report indicate that meropenem was well tolerated and appeared to be as effective as standard monotherapy with imipenem in bacteremic patients. Meropenem and imipenem/cilastatin were highly effective for the treatment of bacteremia in Chinese patients and only mild or negligible side-effects were noted. PMID- 10862446 TI - Randomized, double-blind comparison of irbesartan and enalapril for treatment of mild to moderate hypertension. AB - BACKGROUND: Irbesartan is a newly developed angiotensin II receptor antagonist. Its antihypertensive efficacy and safety in Taiwanese patients with mild to moderate hypertension remains to be determined. METHODS: This was a multicenter, double-blind, randomized, parallel group study. One hundred and sixteen patients from three centers were enrolled. After a placebo lead-in period of 14 days, 55 patients (24-75 years-of-age) who had a mean seated diastolic blood pressure of 95 to 110 mmHg were randomized to once-daily treatment with irbesartan 150 mg or enalapril 10 mg. Doses were doubled at week 4 if trough seated diastolic blood pressure was 90 mmHg or more. Trough blood pressure was measured at zero, two, four and eight weeks of treatment. RESULTS: Both treatments lowered blood pressure with no significant difference in efficacy between treatment groups. Irbesartan 150 mg to 300 mg provided reductions in trough seated systolic and diastolic blood pressures at week 8 of -16.5 mmHg and -7.2 mmHg, respectively, with 36% of patients having a favorable response. Similarly, enalapril 10 mg to 20 mg reduced systolic and diastolic blood pressure by -10.6 mmHg and -5.0 mmHg, respectively, with a response rate of 43%. Headache, malaise and dizziness were the major adverse reactions observed in both groups. The incidence of drug related cough was significantly higher with enalapril (18%) than with irbesartan (0%). CONCLUSIONS: Irbesartan 150 mg to 300 mg once daily was as effective in lowering blood pressure as enalapril 10 mg to 20 mg once daily. Both irbesartan and enalapril were well tolerated, while there was a significantly lower incidence of cough with irbesartan compared with enalapril. PMID- 10862447 TI - Computerized rotational vestibular testing in normal subjects. AB - BACKGROUND: As an integral part of the contemporary vestibular testing battery, computerized vestibular rotational testing provides physiologic stimuli and quantitative evaluation of the vestibulo-ocular reflex function of the horizontal semicircular canals. Clinically, it is most commonly used in the sinusoidal harmonic acceleration test (SHAT) and velocity step test. Because the results for a given subject may vary when tested using different facilities, the purpose of this study is to establish the normative data for our laboratory. METHODS: Fifty six normal subjects underwent the SHAT and velocity step test. Three parameters of SHAT--gain, phase and symmetry--were measured and recorded at 0.02, 0.05, 0.09 and 0.10 Hz. In the velocity step test, the three parameters of postrotatory nystagmus-time constant, maximum slow component eye velocity and directional preponderance were measured and recorded. RESULTS: The means +/- standard deviations of gain in the SHAT were 0.47 +/- 0.16 at 0.02 Hz, 0.50 +/- 0.16 at 0.05 Hz, 0.50 +/- 0.16 at 0.09 Hz and 0.53 +/- 0.17 at 0.10 Hz. The coefficient of variation for SHAT gain was 0.32. The time constant means and standard deviations in the velocity step test were 13.44 +/- 3.53 and 13.52 +/- 3.69 for clockwise and counterclockwise rotations, respectively. CONCLUSIONS: We conclude that computerized rotational vestibular testing is precise. PMID- 10862448 TI - Treatment of acetabular fractures: 10-year experience. AB - BACKGROUND: Acetabular fracture is a controversial and difficult fracture to manage. We retrospectively evaluated the outcome of the traditional management of acetabular fractures. METHODS: From 1987 to 1996, 112 cases of acetabular fracture presented at the Taipei Veterans General Hospital and were managed surgically in 73 cases and nonsurgically in 39 cases. The follow-up period was 90 (36-140) months. RESULTS: In the nonsurgically managed group, congruent reduction was achieved in 29 cases (74.3%) and good to excellent functional results were achieved in 25 cases (64.1%). In the surgically managed group, congruent reduction was achieved in 60 cases (82.2%) and good to excellent functional results were achieved in 57 cases (74.3%). In 51 (45.5%) cases, early or late complications developed after management, including one femoral artery perforation, one screw penetration, three wound infections, one iatrogenic sciatic nerve injury, one deep vein thrombosis, 21 heterotopic ossifications, two chondrolyses, three avascular necroses of the femoral head and 18 cases of symptomatic traumatic arthritis. CONCLUSIONS: The functional results correlated well with the final congruity of the joints and the severity of the complications. PMID- 10862449 TI - Far lateral lumbar disc herniation. AB - BACKGROUND: Far lateral lumbar disc herniation is an uncommon condition that may compress the nerve root outside the vertebral canal and in its extraforaminal course. The traditional midline interlaminar approach for the exploration of far lateral lumbar disc herniation is often difficult because the intervertebral articulation obviates a direct view of the course of the extraspinal nerve. In this report, we present two surgical approaches for the treatment of far lateral lumbar disc herniation: the paramedian muscle-splitting microtechnique and the enlarged midline approach. METHODS: Eight patients with far lateral lumbar disc herniation were found among 160 lumbar disc operations in 160 patients. According to computed tomography results, we divided patients with far lateral lumbar disc herniations into two groups; the extraforaminal and foraminal groups. Clinical presentation, imaging studies and surgical approach were thoroughly reviewed. RESULTS: Three patients in the extraforaminal group underwent the paramedian muscle-splitting microtechnique. Two patients in the foraminal group underwent the enlarged midline approach. The other three were operated on before the introduction of the paramedian muscle-splitting microtechnique and the enlarged midline approach. One of these patients who underwent the traditional interlaminar approach with resection of the lateral portion of facet joint, received additional instrumentation and fusion for the prevention of further instability. All had good results and no further surgical treatment was necessary. CONCLUSIONS: The incidence of far lateral lumbar disc herniation was 5% of all surgically treated disc herniations at our institution. For the extraforaminal group, the paramedian muscle-splitting microtechnique is the surgical route of choice. For the foraminal group, the enlarged midline approach is better than the traditional, interlaminar approach in saving the facet joint and preventing postoperative instability. PMID- 10862450 TI - Review of trophoblastic disease at Taipei Veterans General Hospital. AB - BACKGROUND: Trophoblastic diseases are well known and encountered frequently within most oriental populations except the Japanese. In recent decades, fewer cases have been reported in Taiwan. The purpose of this study was to review and discuss all patients diagnosed with trophoblastic disease at one particular Taiwanese medical center. METHODS: Sixty-four patients with malignant gestational trophoblastic disease (GTD) were treated at the Taipei Veterans General Hospital from 1977 to 1995. All cases, except those of placental-site trophoblastic disease, were included in this study. RESULTS: Of the 64 cases of GTD identified, 36 were nonmetastatic and 28 were metastatic. The common metastatic sites were the lungs, followed by the brain and/or liver. Six patients died of the disease. The majority of these patients (5/6) suffered from liver and/or brain metastases. CONCLUSIONS: GTD was found to be a highly chemosensitive and curable disease. However, a significant proportion of patients die of the disease. More effective therapeutic protocols may be required in such patients to improve the survival rate. PMID- 10862451 TI - Fluidity and fatty acid components of erythrocyte membranes in diabetic retinopathy. AB - BACKGROUND: Recently, numerous anomalies of erythrocyte rheology and function have been reported in diabetic patients. These changes are thought to be related to the alteration of erythrocyte membrane fluidity and may play an important role in diabetic retinopathy (DR). Nonetheless, the principal factors contributing to fluidity change remain undetermined. The influence of erythrocyte membrane fatty acid components on membrane fluidity and retinopathy was evaluated in patients with type 2 diabetes. METHODS: The fatty acid components and fluidity of erythrocyte membranes in 63 patients with type 2 diabetes with and without retinopathy were examined using high-performance liquid chromatography and fluorescence polarization. RESULTS: The content and composition of erythrocyte membrane arachidonate (C20:4n-6) was significantly lower in diabetics than in controls, and diabetics had significantly increased erythrocyte membrane microviscosity compared with controls. Furthermore, membrane microviscosity was higher in diabetics with DR than those without DR. In the diabetic patients, arachidonic acid contents of erythrocyte membranes were significantly, inversely correlated with erythrocyte membrane microviscosity, fasting blood glucose, glycosylated hemoglobin, triglycerides and cholesterol, and positively correlated with the insulin sensitivity index. CONCLUSIONS: The results suggest that erythrocyte membrane fatty acid components may contribute to alterations in membrane fluidity in patients with type 2 diabetes. Alterations in membrane fluidity may play an important role in the development of diabetic microangiopathy. PMID- 10862452 TI - Spuriously high CK-MB isoenzyme activity mimicking acute myocardial infarction in a patient with adenocarcinoma of the rectum. AB - A 58-year-old Chinese male presented with precordial distress and was found to have elevated creatine kinase (CK)-myocardial bound (MB) (determined by the Kodak Ektachem Clinical Chemistry Slide method) activity and an abnormal electrocardiogram. Eventually, the patient was diagnosed with an annular ulcerative tumor 10 cm above the anal orifice. The tumor was an adenocarcinoma of the rectum and was immunoreactive for CK-BB (brain type). CK isoenzyme electrophoresis disclosed both BB and MM (muscle type) bands. We concluded that the level of CK-MB determined using the Kodak Ektachem Clinical Chemistry Slide method was spuriously elevated in our patient with adenocarcinoma of the rectum, without evidence of myocardial injury. Institutions using the Kodak Ektachem Clinical Chemistry Slide system for CK should be aware of this possibility. PMID- 10862453 TI - Postictal psychosis with forced normalization. AB - A 22-year-old female patient with epilepsy for more than six years who had prominent psychiatric manifestations, including paranoia, delusions and hallucinations, after a series of major seizure attacks caused by sudden withdrawal of anticonvulsant medication was monitored. This reported episode of psychosis occurred eight hours after the last seizure attack and lasted for two weeks. The psychosis gradually disappeared after administration of lorazepam. The electroencephalogram (EEG) performed during the period of psychosis showed intermittent slow activity in the bilateral frontal regions of the brain. There were active bisynchronous epileptiform discharges bisynchronously in both frontotemporal regions after the resolution of the psychotic episode. The clinical picture and course were consistent with the diagnosis of "postictal psychosis", and the transient near normal EEG during psychosis was most likely a phenomenon known as "forced normalization". The patient has not had a similar psychotic attack since the one reported here. PMID- 10862454 TI - Squamous cell carcinoma of the right upper lung congenital tracheal bronchus. AB - Tracheal bronchus, directly arising from the abnormal tracheal wall, is a rare congenital anomaly of the airway. Lung cancer developing in a tracheal bronchus is even more rare. A 68-year-old man had a mass shadow over the right upper lung on chest radiography. Bronchoscopy revealed an abnormally displaced bronchus stemming from the right side of the tracheal wall, 2 cm above the carina. Arising from this tracheal bronchus was a tumor, a biopsy of which showed squamous cell carcinoma. During surgery, a bronchus arising from the right lateral wall of the trachea and supplying the apical segment of the right upper lobe was found. There was a tumor occluding the lumen of the tracheal bronchus, causing distal obstructive pneumonitis. The patient underwent right upper lobectomy with closure of the tracheal bronchus and radical lymph node dissection. The subsequent pathologic examination of the specimen revealed squamous cell carcinoma with surrounding pneumonitis. Bronchogenic cancers developing in a tracheal bronchus are rarely reported in the English language literature. Surgical resection appears to be the treatment of choice. PMID- 10862456 TI - [Pleural mesothelioma: current status]. PMID- 10862455 TI - Cytomegalovirus pericarditis with cardiac tamponade in a young infant. AB - The principal viruses implicated in pericarditis are enteroviruses. Cytomegalovirus pericarditis is quite rare and has been reported in immunocompromised patients with acquired immunodeficiency syndrome, malignant neoplasm or organ transplantation. We report a three-month-old male infant who suffered from cough and rhinorrhea for two weeks. He developed shortness of breath for three days, and fever for one day, prior to admission. Physical examination revealed tachycardia, tachypnea, pale conjunctiva, hepatomegaly, and a muffled heart sound without significant murmur. Chest radiography showed marked enlargement of the cardiac silhouette. Echocardiography demonstrated a large amount of pericardial effusion with impaired diastolic ventricular function. After pericardial drainage and supportive treatment, the fluid gradually disappeared. Viral culture of the pericardial fluid and serologic data confirmed a cytomegalovirus infection. Cytomegalovirus pericarditis should be included in the differential diagnosis of pericardial effusion in a young infant. PMID- 10862457 TI - [Changes in the hemostatic system after laparoscopic cholecystectomy]. AB - A 30 degrees reverse Trendelenburg position and the pneumoperitoneum performed in patients undergoing at Laparoscopic Cholecystectomy (LC) cause haemodynamic modifications in inferior cava vein and in femoral veins producing a venous stasis in lower limbs. So that activation of coagulation with hypercoagulability occurs. The aim of this study is to value the modification of the haemocoagulative and fibrinolytic factors during LC and OC. 18 patients with symptomatic and non complicated lithiasis were examined. They were divided in two groups, nine patients for each group. LC was performed in first group and OC in the other group. Prothrombin time (PT), plasma-activated partial thromboplastin time (PTT), Antithrombin III (AT III) fibrinogen degradation products (FDP) were valued before and at 6, 12, 24, 48 hours after operation. For the statistical analysis Student "t" for average comparison was used. There were no relevant alterations of PT, PTT and ATIII after operation in the two groups. FP was increased after surgery, especially at 24th-48th hours in the group in which OC (p < 0.05) was performed. FDP were increased in both groups but only at 24 and 48 hour after OC (p < 0.05) the result was significant. Earlier mobilization and reduced invasivity of laparoscopy could contrast thrombotic effects of the blood stasis in the lower limbs when 30 degrees-45 degrees reverse Trendelenburg and pneumoperitoneum are performed. PMID- 10862458 TI - [Clinical considerations on malignant carcinoids: bronchial, gastric, ileocolic involvement]. AB - Six cases of malignant neuroendocrine tumours (3 bronchial, 1 ileocolic) are presented; diagnostic and clinical features, functional behaviour and the results of the treatment are discussed. Early functional manifestation (diarrhoea) was observed only in the first case of ileocolic tumour: in this patient the diarrhoea disappeared after the operation but further relapse occurred at the recurrence of the pathology. In the second ileocolic tumour diarrhoea with increased levels of 5 HT appeared later on, after the operation, expressing repeated disease. In one malignant bronchial carcinoid with early spinal secondary, the typical flushes appeared only after the explosion of metastatic spread. No functional disorders were observed in one patient with metastasis localised only in the bones. On the basis of actual knowledge the authors discuss the pathogenetic hypothesis of these events. Even though these experience limited, it may remark the importance of the quantity and quality of produced hormones, of the localization of metastasis in the liver and portal district and the unconstant production of clinical symptoms. The importance of A-chromogranin and of NSE as specific markers is discussed; the studies on the chromogranin have furthermore demonstrated the possible effect of the inhibitors of protonic pump on structural, hyper-dysplastic modification of gastric mucosa and this requires further investigation. PMID- 10862459 TI - [Benign schwannoma of the pelvic retroperitoneum. Report of a case and review of the literature]. AB - Benign schwannoma is a tumor arising from Schwann cells (forming the neural sheath of peripheral nerves). The retroperitoneal location is unusual (0.5-5% of cases). Most common locations are cranial nerves (especially the 8th pair) and, in peripheral nerve system the neck, mediastinum and extremities. To this date the known cases of benign retroperitoneal schwannoma are about 60, of which less than 20 in the pelvis. The low frequency of this tumor and the lack of specific instrumental signs and objective symptoms (since it develops in a deep and broad region as retroperitoneum) make presurgical diagnosis very difficult. It can be confirmed only during surgery and definitive histological examination. The information provided by ultrasonography, CT and MR help to limit diagnostic hypothesis, but they don't show any pathognomonic images. The resection of this tumor is the appropriate treatment, even though it is really a complex one. Prognosis is quite good since post-surgical recurrences are unusual. If they appear is probably because excision wasn't radical. Complete resection is the best treatment for retroperitoneal pelvic schwannoma and today it can be performed also by laparoscopy. Partial resection can be used when the mass is strongly connected to essential organs in order to prevent iatrogenic harms (neural deficit, vessel lesions); this may occur in 10% of cases. This paper describes a benign schwannoma with pelvic retroperitoneal location, incidentally discovered during a routine gynecological check up. The purpose of this study is to review current therapeutic and diagnostic techniques in retroperitoneal pelvic schwannoma (including a review of current literature) and to identify th problems that can be encountered in the differential diagnosis of this unusual disease from other neoplasms occurring in the same place. PMID- 10862460 TI - [Recurrent giant retroperitoneal leiomyosarcoma. Report of a clinical case]. AB - The Authors report the case of a male patient, 52 year old, suffering from retroperitoneal leiomyosarcoma, submitted to multiple operations in the space of about seven years, for the presence of liver metastases and local relapses; the good general health state, the moderate grade of the neoplasm and the disease's fair interval free, have justified the therapeutic attitude adopted; the patient at present enjoys good health. PMID- 10862461 TI - [Annular pancreas in adults: diagnostic considerations on a case]. AB - Annular Pancreas (AP) is a rare congenital anomaly that usually presents in childhood with symptoms referable to duodenal obstruction; nonetheless, this condition can manifest in adulthood with abdominal pain, pancreatitis, duodenal ulcer, pancreatic head mass. The Authors hereby discuss a case of AP observed in a 63 year-old patient in which EUS played a decisive role in achieving a certain diagnosis. PMID- 10862462 TI - [Surgical treatment of total rectal prolapse: Delorme's technique]. AB - Rectal total prolapsus is a cause of a permanent disablement more founded in old women. Many pathogenetic hypotheses are made up to date time, many surgical treatments were proposed for the treatments of this disease: through perineal, abdominal or combined way. The results are different, of course, in a short and a long time, whether in the matter of relapse of the prolapse, or in morbidity in connection to surgical treatment and his sequences. After a review of the most frequent techniques now in use, the Authors report their experience on 6 patients operated from 6 January 1988 to June 1993, giving reasons for their preference to Delorme's operation through perineal way. PMID- 10862463 TI - [Rubber band ligation of hemorrhoids. 5-year follow-up]. AB - The Authors quickly reviewing the various surgical ambulatorial methods of hemorrhoids, considered the rubber band ligation, technique. The purpose of this study is that to define the real advantages of the before said technique through the analysis of the cases operated. From December 1992 through 1997 100 patient with hemorrhoids were observed. Of these, 37 have been submitted to rubber band ligation, for a total of 135 bindings. In 26 cases (86%) the Authors observed disappearance of the bleeding and the local troubles. In eight patients complete reduction of the mucous prolapse was obtained. Two patient have continued to complain even if with small intensity, the subjective symptoms, that induced the Authors recommend surgical therapy. The last two patients have been submitted with success to hemorrhoidectomy and reduction of the concomitant prolapse. The advantages of the rubber band ligation are indisputable: it offers the possibility of a definitive, ambulatorial with least risk solution to without necessity of hospitalisation and anesthesia, repeatable in the time and with an overall percentage of complications (0.4%) inferior to that of the surgical hemorrhoidectomy. PMID- 10862464 TI - [Elective and emergency minimally invasive surgery in pleural diseases]. AB - In the last years the video-assisted thoracic surgery (V.A.T.S.) assumed an important order for the diagnosis and treatment of the pleural disease. In this particular field, the procedure allows obtaining almost the same outcomes of traditional surgery and is very safety. V.A.T.S. reduces hospital time and trauma with a fast return to the working life. The Authors describe their experience and outline the diagnostic and therapeutic indications for the treatment of choice and emergency. PMID- 10862465 TI - [The mechanisms of the curative action of systemic enzyme therapy]. AB - Employment of the authors' highly sensitive biochemical method allowed the fact of resorption of proteolytic enzymes after their peroral ingestion to be confirmed, with a question having been raised as to further fate of proteinases and mechanisms of curative action of combined enzyme preparations. The analysis of relevant data from published literature and the results of the author's own studies permitted identifying the most important moments of realization of the systemic enzyme therapy effects, which are as follows: 1) activation of receptors on the surface of enterocytes; 2) formation of activated alpha 2-macroglobulin (alpha 2-M) under the action of proteinases from the intestines, and realization of catalytic activity of those enzymes involved in this restrictor of proteinases; 3) modulation of cell activity by complexes alpha 2-M-proteinase; 4) activation of the system of mononuclear phagocytes; 5) influence on the balance of bodily endogenous proteolytic systems. PMID- 10862466 TI - [The clinico-pathogenetic characteristics of neurosis-like states in the participants in the cleanup of the aftermath of the accident at the Chernobyl Atomic Electric Power Station]. AB - A clinical and paraclinical examination was done in 120 individuals with neurosis like disorders, who had taken part in the elimination of the aftermath of the Chernobyl Nuclear Power Plant breakdown. A complex of etiological factors in the disorder has been identified. The most important syndrome was cerebral asthenia associated with depressive disorders and apparent vegetovascular dysfunction. Cerebral influences were implicated in the origination and development of neurosis-like disorders, with functional inadequacies having been disclosed in the patients' bodily catecholamine and immune systems. PMID- 10862467 TI - [The toxic and hygienic characteristics of the new synthetic organic flocculants AES-5, AES-7 and AES-10]. AB - A toxicological and hygienic characterization is submitted of novel synthetic organic flocculant AEC-5, AEC-7, AEC-10 which are low-toxicity substances and are classified under the fourth class of hazards. They have no skin-resorptive, locally irritative action and are endowed with a weak cumulative activity of functional character. The AEC-5 flocculant exerts a moderately manifest sensitizing effect in the dermal route of entry. PMID- 10862468 TI - [Angiotensin-converting enzyme inhibitors in the treatment of arterial hypertension]. AB - Angiotensin-converting enzyme (ACE) inhibitors have come into widespread use as a treatment option for patients with arterial hypertension, which fact can be explained by their apparent hypotensive effect, a small percent of side effects, protective effects on target organs, and a possibility of reversibility of essential hypertension- and symptomatic hypertensions-related changes in the cardiovascular system. There are a great many of drug preparations belonging in the group of ACE-inhibitors (capoten, enalapril, renitec (enalapril maleate), benazepril, quinapril, cilazopril, lizinopril, ramipril--to name but few of them. Specificities of different representatives of ACE-inhibitors, as to their clinical efficacy were found to be related to different pharmacokinetic properties of drugs. PMID- 10862469 TI - [Agents for the metabolic correction of myocardial energy metabolism in treating patients with ischemic heart disease]. AB - Correction of the metabolic link of the natural course of ischemic heart disease (IHD) was found out to be a real reserve of enhancement of therapy efficiency in treating this condition. A total of 137 IHD patients were examined, presenting with functional class II-III exertional angina, their age ranging between 67 to 86 years. Kinds of metabolic complexes to be used in IHD treatment included amino acids, antioxidants, cofactors, energy-providing compounds that enhance efficiency of basic methods of IHD therapy. A positive therapeutic effect of metabolic correction treatments was evidenced by earlier dispelling of electrocardiographic signs of myocardial ischemia, higher tolerance to physical load, improvement in parameters characterizing cardio-hemodynamics. PMID- 10862470 TI - [The ultrastructural changes in the air-blood barrier in the acute period of a myocardial infarct with the development of pneumonia (data from autopsy materials)]. AB - In this paper, the author highlights the ultrastructural changes in the lung respiratory zone during the acute period of myocardial infarction in origination and development of pneumonia. The development of inflammatory process was found out to be preceded by microcirculation disorders, focal damage to the gas-blood barrier, and lung tissue bacterial contamination at day 2 to 4 of the course of myocardial infarction. The exudate was seen to contain much fibrin, alveolar macrophages, and erythrocytes. The lung tissue develops albuminous and fatty degeneration with the alveolar septa and lumina storing fibrin and fatty drops. PMID- 10862471 TI - [Lipid peroxidation, the antioxidant system and the trace element level in acute pancreatitis]. AB - Overall, thirty-two patients with acute pancreatitis (edematous, destructive, purulent forms) were studied for the state of lipid peroxidation (LPO), antioxidant defence (AOD), and trace element status. The development of destructive and purulent forms of acute pancreatitis was found out to be accompanied by activation of LPO processes, AOD breakdown, with profound disturbances having been disclosed in the metabolism of zinc, selenium, cuprum, manganum, that play an important part in LPO, AOD processes, and in those of immune defence. The secured results suggest that there is a need for us to include into a complex therapy of acute pancreatitis antioxidants and some trace elements (zinc, celenium) and that it is expedient to use indices for the LPO, AOD systems, and for trace element status in the differential diagnosis of clinical forms of acute pancreatitis, prognostication of development of purulent complications. PMID- 10862472 TI - [The neutral lipid and total phospholipid content of the saliva in gingivitis and periodontitis]. AB - With the aid of a high-performance thin-layer and gas-liquid chromatography, an increase has been found out in the mixed saliva content of long chain fatty acids, triacylglycerin, diacylglycerin, cholesterin, cholesterol esters, and total phospholipids in parodontium tissues inflammation (gingivitis, parodontitis). In the gingiva fluid in parodontitis, the content of volatile fatty acids (C3-C6) and their isomeric forms was found to have gotten increased. Based on comparison of the content of certain lipid fractions in mixed and parotid saliva it has been shown that the main source of augmentation of fatty acid is the gingiva fluid, neutrophiles, and anaerobic microorganisms. PMID- 10862473 TI - [The effect of wheat germ oil on the antioxidant system of animals]. AB - An experimental study was made on wheat-germ oil as a source of easily assimilable vitamin E. In rats, the above oil intake results in a rapid increase in the content of vitamin E in the brain, liver, heart, lungs, kidneys, and spleen. There has been studied a change in the intensity of lipid peroxidation processes. It is shown that in oral administration wheat germ oil efficiently saturates the body with vitamin E. In all tissues, the work of the [symbol: see text]-tocopherol redox-system is noted to be stimulated, LPO inhibited, with eicosanoid synthesis proceeding under immediate control of the system of fat soluble vitamins. The total state of dynamic equilibrium in the LPO-AOA system has effectively shifted to the side of predominance of the AD system components. The total tissue AOA remained stable in most organs. PMID- 10862474 TI - [The action in vitro of a permanent and an alternating magnetic field and of cytosar on the colony- and cluster-forming properties of bone marrow cells from hematologic patients]. AB - The bone marrow of hematological patients was studied as were effects of cytozar at different dilutions on the bone marrow cells. The analysis of the secured results showed the magnetic field to be a powerful biologically active factor capable of changing clonogenic properties, proliferative and differentiative potential of hemopoietic cells precursors. A long exposure brings about a decline in the colony-forming activity of bone-marrow cells, that is to say, with prolongation of time of the exposition the magnetic field becomes an unfavourable factor for the leukotic cell culture development. Similar results were obtained with cytozar in the culture while studying the colony-forming capability of bone marrow cells. Hence it appears that action of magnetic field under long exposition to it, combined exposure to magnetic fields, are, in some cases, comparable to effects of cytozar. It is possible that concurrent prescription of cytozar together with magnetization of blood in patients with hemoblastoses should permit enhancing the efficacy of treatment. Employment of weak magnetic fields is a satisfactory and worth-while therapeutic attempt to be made in a multimodality treatment of patients with hemoblastoses. PMID- 10862475 TI - [The prognosis for the occurrence of inflammatory genital diseases in girls and young women]. AB - With the purpose of predicting probability of origination of inflammatory diseases of the genital organs among young female patients and their diagnosis during the early stages of development thereof, an algorithm has been worked out together with a mathematical model of probability of origination of genitalia inflammatory diseases at young age, based on prevailing social and economic and medical and biological risk factors. Correspondence between the secured indices across all samples among girls came up to 95.9%, that for the young women group was 99.3%. High significance and relative simplicity of the above algorithm for prediction of risk of development of inflammatory diseases of genitalia in girls and young women suggest to us that it can surely come into widespread use. PMID- 10862476 TI - [Anatomical variants of the lymphatic bed of the human epididymis]. AB - In a series of 59 male subjects aged between 1 to 86 years, the human epididymis intraorgan lymphatic bed was studied using subserosal injection of Herot's mass. It is shown that the epididymal lymphatic bed consists of the surface- and intraorgan plexuses of lymphatic vessels. The surface plexus is represented by subserosal, surface and deep intracapsular networks of lymphatic vessels. The development of the surface plexus is at its greatest on the dorsal surface of the proximal segment of the organ. The intraorgan plexus of lymphatic vessels resembling baskets braids the segments of the duct and cones of the small seminal ducts. One is supposed to know and take into account the topography of lymphatic vessels in performing surgical interventions on the epididymis in order that it might be possible to prevent their injury and reduce the incidence of complications related to disturbances in the lymphatic drainage. PMID- 10862477 TI - [The characteristics of the attitude to disease of people with venereal diseases]. AB - The article focuses on psychological aspects of spread of venereal diseases, such as accentuation and types of the patients' attitude towards illness. Patterns have been studied of types of accentuation and types of attitude towards illness in men and women. Psychological features as risk factors for venereal diseases are characterized together with results of treatment. The conducted investigation permits elaborating a system of psychocorrection and psychorehabilitation of patients with venereal diseases. PMID- 10862478 TI - [The prostaglandin level in patients with an alcohol dependence syndrome]. AB - A change in the state of the cardiovascular system in patients presenting with gamma alcohol syndrome is mediated by changes in the neurohumoral system and in regulation of the vascular tone and changes in prostaglandins (PG). To study the above-named changes, male subjects aged between 24 to 60 years were examined as were 20 essentially healthy men in the same age bracket. The examined patients demonstrated a marked activation of all kinds of PG during early stages of the presence of gamma alcoholism syndrome. With prolongation of duration of the underlying pathology there comes to be formed a dysbalance with predominance of pressor fractions of PG. PMID- 10862479 TI - [The treatment of peptic ulcer patients with a disordered motor-evacuatory function of the digestive canal]. AB - Submitted in the paper are basic mechanisms of development of disorders of gastrointestinal motility and evacuatory function in ulcer of the stomach and duodenum. Pharmacological correction of the above disorders is considered in some detail and discussed as thoroughly as possible. Ways for enhancement of motility and normalization of the evacuatory function of the stomach and duodenum are outlined, indications and contraindications are determined, specificities of each of the drugs employed are described. In the author's own investigations, coordinax has been shown to be superior to other available prokinetics. PMID- 10862480 TI - [The dynamics of the humoral immunity indices and of the beta 2-microglobulin level in children with chronic hepatitis]. AB - Studied in children with chronic hepatitis B (n = 38) were indices for humoral immunity, such as blood serum content of immunoglobulins A, G, M, and beta 2 microglobulin. The revealed increase in the level of the indices under study suggest to us an apparent tension of the humoral link of immunity. The polyenzymic drug preparation wobenzym in the complex of therapeutic measures instituted in the above children was found out to have a positive effect on humoral homeostasis. PMID- 10862481 TI - [The prevention and treatment of enterocolitis in children with Hirschsprung's disease]. AB - Enterocolitis remains the most grave and very dangerous complication of Hirschsprung's disease. Its etiology is not fully understood. It can occur in children both before and after surgery. The article focuses on the course of enterocolitis in 35 pediatric patients with Hirschsprung's disease. The clinical course varied from symptom-free course to an aggregate of manifest symptoms of enterocolitis and rapid progression of pathological changes. In all above pediatric patients, a therapeutic and prophylactic complex of measures was inaugurated involving urgent decompression of colon with irrigations and symptomatic therapy as well. Early diagnosis of Hirschsprung's disease is regarded as a mainstay in enterocolitis prophylaxis. The most effective method of prophylaxis and treatment of enterocolitis is decompression of a stretched colon with the aid of the irrigation technique. PMID- 10862482 TI - [Insulin monotherapy in patients with diabetes mellitus type 2 developing secondary resistance to sulfanilamide preparations]. AB - Patients with type-II diabetes mellitus differ widely in degrees of beta-cell functional activity, with some of them requiring insulinization as a corrective measure for hyperglycemia. It is a well-chosen and a propitious treatment option in those patients presenting with secondary resistance to sulfa drugs. PMID- 10862483 TI - [The importance of crystallography in the diagnosis of pyelo- and glomerulonephritis in children]. AB - The article provides characterization of a new crystallography technique with making use of Petri dishes and a 2% solution of CuCl2 in pyelonephritis and glomerulonephritis. As many as 116 children living in the city of Kiev were enrolled in the study. Different patterns of crystals in the above illnesses have come to be seen in macro- and micro-examinations. PMID- 10862484 TI - [The diagnosis of autonomic disorders in premature newborns]. AB - A diagnostic algorithm has been worked out for identification of sympathicotonia and parasympathicotonia in newborn infants. The algorithm is calculated on the basis of results of observations carried out in 286 premature infants who had been examined three times during the first fourteen days of their lives. Included in the algorithm are 11 indices of informative value. Score total +13 suggest to us sympathicotonia while -13 indicates parasympathicotonia, with probability threshold being 95%. The above method permits the determination of direction of autonomic disorders in newborn infants, in which case there is no necessity whatever to conduct elaborate and expensive investigations (spectral analysis of the heart's rhythm, measurement of hormones and mediators. PMID- 10862485 TI - [The development of the skin-optical perception of color and images in blind schoolchildren on an "internal visual screen"]. AB - In profound impairement of vision the function of colour and seen objects perception is absent, with the person being unable to orient himself in space. The uncovered sensory sensations of colour allowed their use in training the blind in recognizing the colour of paper, fabric, etc. Further study in those having become blind will, we believe, help in finding eligible people and relevant approaches toward educating the blind, which will make for development of the trainee's ability to recognize images on the "inner visual screen". PMID- 10862486 TI - [The clinical use of blood ultrafiltration in leptospirosis patients with acute kidney and liver failure]. AB - The use of ultrafiltration of blood in clinical settings in the treatment of patients with leptospirosis presenting with acute renal failure permits enhancing a detoxicating effect due to elimination with the ultrafiltrate of up to 30% of products of nitrogenous metabolism. This allows the water balance to be corrected, homeostasis, the functional activity of the cardiovascular, respiratory systems to be stabilized. Therapeutic effectiveness is enhanced that is evidenced by a 20% decline in the incidence of complications and in case fatality rate in grave forms of leptospirosis. PMID- 10862487 TI - [The prenatal diagnosis of placental insufficiency in pregnant women with a herpetic infection]. AB - Our objective in this study was to find out exactly the diagnostic criteria for dysfunction of the fetoplacental complex in pregnant women with herpes infection. A total of 62 pregnant women who ran a high risk for intrauterine infection were examined. As many as 87 percent of female patients were found to be PHV-infected displaying manifestations of the immunodeficiency syndrome such as increased pathological antibody formation and activation of mechanisms of cell immunosuppression, which fact results in disturbance in the fetomaternal immune relations. Routine methods of obstetric examination lack informative value, which fact necessitates conducting a combined echographic and immunological investigation in order that we might be able to establish a perinatal prognosis. PMID- 10862488 TI - [Surgical methods for delivery in modern obstetrics and their influence on maternal and infant health]. AB - The article addresses issues of comparative characterization of deliveries involving surgery and impact thereof on the health of the mother and her child. Risk factors are identified that the mother and her child run in sectio cesarea, in application of obstetrical forceps, and in vacuum-extraction of the fetus. Cesarean section was found out to be the most acceptable mode of delivery in origination of organic and functional nervous system involvement in children but the most ill-chosen and unpropitious one in the mother, especially so in those groups at risk for bleeding, septic complications, and genital endometriosis. Among those surgical methods of delivery being the least traumatic to the mother are obstetrical forceps and vacuum-extraction of the fetus. PMID- 10862489 TI - [The correction of the immune status at the postoperative rehabilitative stage in lung cancer patients by using millimeter-wave resonance therapy]. AB - Results are submitted of study into effects of microwave resonance therapy (MRT) on parameters characterizing cell immunity in those lung cancer patients having undergone a radical operation. This study comprised 58 patients as the main group, with 31 patients being controls. Indices for cell immunity were also studied in 20 donors. There was a rise in T-lymphocytes after MRT in the postoperative period. MRT contributed to prolongation of patients' lives, reduction of the incidence of chemotherapy-related dyspeptic complications and that of leukopenia cases by 8 and 9 percent respectively. PMID- 10862490 TI - [The treatment of the initial stage of cervical cancer]. AB - The analysis of the author's own experience and of relevant published literature suggest that it is expedient to further elaborate methods for treatment of incipient forms of uterine cervix cancer aimed to preserve the genital organs in women, especially in those at childbearing age. Worthy of mention in this respect is the progress made by the Kiev City Oncological Centre. In choosing a treatment option, it is necessary that etiopathogenic risk factors for development of cancer of the uterine cervix by analyzed and taken account of. PMID- 10862491 TI - [The characteristics of the ultrastructural organization of the cervix uteri mucosa after the combined treatment of a papillomavirus infection]. AB - The paper presents findings obtained from the electron microscopy investigation designed to study the mucosa of the vaginal portion of the uterine cervix in patients with atypical condyloma 45 days after CO2 laser destruction had been performed and system interferon therapy conducted. The proposed combined method was found out to encourage normalization of the uterine cervix structure, with the above therapy being capable of activating the local immune response that prevents the disease recurrences. PMID- 10862492 TI - [A case of spontaneous rupture of the inferior epigastric artery]. PMID- 10862493 TI - [The effect of manufacturing factors in asphalt-bitumen plants on the health of the workers]. AB - Comprehensive investigations designate to study the working conditions and health status of workers of asphalt-bitumen plants were conducted. Measurable in the breathing zone of workers of basic professions were toxic substances that have a toxic effect on the body (benzene, oil hydrocarbons, toluene, nitric, carbonic oxides, and benz(a)pyrene). There was noted a high incidence of abnormalities of the internal organs, with the cardiovascular and respiratory pathology ranking first in frequency to other disorders. Studies on the allergologic and atopic status showed that as far as the development of the respiratory organ pathology and the external breathing functional shifts are concerned, the immunity status related specificities of the body are to be considered along with in-plant factors. Measures have been designed aimed to prevent the development of disorders. PMID- 10862494 TI - [Changes in the interleukin-6 and interleukin-10 concentrations in the blood plasma of miners working in deep coal mines]. AB - Blood plasma levels of interleukin-6 (IL-6) and interleukin-10 (IL-10) were measured in 45 miners working in a deep coal mine immediately after work shift using an immunoenzyme technique. The highest IL-6 level was recorded in those miners engaged in hard work under most adverse conditions of underground workings -it was found to exceed the control values. The same group of workers demonstrated the lowest level of IL-10 that differed from the control value. Miners aged between 41 to 50 years working in a coal mine, their underground service duration 16 to 20 years, displayed a decline in the level of IL-6. The coal mine miners with the 11- to 15-year service duration revealed an increase in the level of IL-10. PMID- 10862495 TI - [The combination of reflexotherapy and homeopathy in treating patients with facial nerve neuropathy]. AB - Used in a multiple-modality treatment of 103 patients presenting with facial nerve neuropathy were infrared laser puncture and homeopathy. The former treatments were administered with the aid of the apparatus [symbol: see text] 001 at wave-length 890 nm, average power 15 mW/cm2. Homeopathy therapy was with those drug preparations meant to deal with constitutional and symptomatic problems. As many as 103 patients with facial nerve neuropathy derived apparent benefit from treatment, which fact was confirmed by electrophysiological findings as was by those from acupuncture diagnosis techniques. PMID- 10862496 TI - [The social hygiene assessment of mortality among adolescents and young people in Ukraine]. AB - An analysis is submitted of age- and sex-specific mortality among adolescents and young people (young men and women) aged 15-19, 20-24, 25-29) of Ukraine. The main causes of death in the above groups of population related to sex and age and place of residence (town, village) are stated. Measures to be designed and implemented to reduce death rates from the most important causes in juveniles and young people are outlined. PMID- 10862497 TI - [The characteristics of the delivery of hospital care to elderly patients under the current conditions]. AB - Based on the epidemiological survey and materials on hospitalization of pensionable population in different regions of Ukraine real medical services requirements were determined for this category of population. 43.3% of urban population older than those capable of working need to be kept under continuous medical surveillance, the same being true of 37.2% of rural population. 10.6% and 8.3% of population respectively are in want of intensive short-term care in the in-patient departments of hospitals followed by supplementary treatment with making use of out-patient facilities. 4.3% and 9.5% of population need to be admitted into social care units and into nurse-tending units as well as into those units designed to provide long-term treatment for chronically sick patients. Models have been elaborated of the required duration, staging, and forms of in-patient treatment of pensionable patients as exemplified by cardiological patients. It is shown that treatment involving intensive care in a specialized in-patient facility and subsequent completion of the patient care in the day hospital, home hospital, unit for a prolonged treatment of chronically sick patients is more expedient from the economical standpoint than staying in hospital over the whole period of time required for the patient to resume his/her health. PMID- 10862499 TI - [In Process Citation] PMID- 10862498 TI - [The sociomedical problems of disability in elderly persons]. AB - Indices were analyzed for invalidity in pensionable persons residing in Chernivtsi Province, presenting with disorders of respiratory organs and circulatory system. By comparing intensive indices for primary disablement in pensioners and those able to work and data on distribution of primary disablement according to forms of diseases it is shown that in persons of greater age complications of chronic unspecific lung diseases are "hidden" in the cerebrovascular pathology. Ways for realization of the program intended to rehabilitate disabled persons of greater age are outlined. PMID- 10862500 TI - Lesioning of locus coeruleus projections by DSP-4 neurotoxin treatment: effect on amphetamine-induced hyperlocomotion and dopamine D2 receptor binding in rats. AB - DSP-4 is a neurotoxin highly selective for the noradrenergic nerve terminals of the locus coeruleus projections. Data on the effect of DSP-4 treatment on amphetamine-induced hyperlocomotion are contradictory. In this study, DSP-4 (50 mg/kg) caused reduction of noradrenaline levels by 70% in the cerebral cortex and by 79% in the cerebellum. This treatment resulted in upregulation of dopamine D2 receptors in the striatum as evidenced by [3H]-raclopride binding. In an open field test, DSP-4 reduced locomotor activity. D-Amphetamine (1.5 mg/kg) caused a similar increase in locomotor activity in control and DSP-4-pretreated animals not familiar to the apparatus. However, when the rats were habituated to the test apparatus, the effect of amphetamine on horizontal activity was significantly larger in the DSP-4-pretreated animals. These data suggest that supersensitivity of D2 receptors develops after locus coeruleus denervation, but that the enhanced efficacy of amphetamine in DSP-4-treated rats is masked by neophobia. PMID- 10862501 TI - Antioxidative and prooxidative action of stilbene derivatives. AB - The effects of stilbene derivatives, including resveratrol, diethylstilboestrol and stilbene, as antioxidants or prooxidants were examined. Resveratrol and diethylstilboestrol, but not stilbene, strongly inhibited NADPH- and adenosine 5' diphosphate (ADP)-Fe3+-dependent lipid peroxidation at the initial and propagation stages. In addition, phenolic stilbenes also inhibited ultraviolet light-induced lipid peroxidation. Resveratrol and diethylstilboestrol efficiently scavenged 2,2'-azobis-(2-amidinopropane)-dihydrochloride peroxyl radicals. However, 2,2'-diphenyl-p-picrylhydrazyl radicals were trapped only by resveratrol, but not by diethylstilboestrol. These results suggest that the inhibitory effect of phenolic stilbenes on lipid peroxidation was due to their scavenging ability of lipid peroxyl and/or carbon-cantered radicals. Resveratrol efficiently reduced ADP-Fe3+, but not EDTA-Fe3+. Stilbenes and diethylstilboestrol did not reduce either ADP-Fe3+ or EDTA-Fe3+. The strand breaks of DNA were stimulated during the interaction of resveratrol with ADP-Fe3+ in the presence of H2O2. These results suggest that phenolic stilbenes act as antioxidants of membrane lipids and that resveratrol has a prooxidative effect DNA damage during interaction with ADP-Fe3+ in the presence of H2O2. PMID- 10862502 TI - Myrica nagi attenuates cumene hydroperoxide-induced cutaneous oxidative stress and toxicity in Swiss albino mice. AB - In recent years, considerable efforts have been made to identify new chemopreventive agents which could be useful for man. Myrica nagi, a subtropical shrub, has been shown to possess significant activity against hepatotoxicity and other pharmacological and physiological disorders. We have shown a chemopreventive effect of Myrica nagi on cumene hydroperoxide-induced cutaneous oxidative stress and toxicity in mice. Cumene hydroperoxide treatment at a dose level of 30 mg/animal/0.2 ml acetone enhances susceptibility of cutaneous microsomal membrane for iron-ascorbate-induced lipid peroxidation and induction of xanthine oxidase activity which are accompanied by decrease in the activities of cutaneous antioxidant enzymes such as catalase, glutathione peroxidase, glutathione reductase, glucose-6-phosphate dehydrogenase and depletion in the level of cutaneous glutathione. Parallel to these changes a sharp decrease in the activities of phase II metabolizing enzymes such as glutathione S-transferase and quinone reductase has been observed. Application of Myrica nagi at doses of 2.0 mg and 4.0 mg/kg body weight in acetone prior to that of cumene hydroperoxide (30 mg/animal/0.2 ml acetone) treatment resulted in significant inhibition of cumene hydroperoxide-induced cutaneous oxidative stress and toxicity in a dose-dependent manner. Enhanced susceptibility of cutaneous microsomal membrane for lipid peroxidation induced by iron ascorbate and xanthine oxidase activities were significantly reduced (P<0.05). In addition the depleted level of glutathione, the inhibited activities of antioxidants, and phase II metabolizing enzymes were recovered to a significant level (P<0.05). The protective effect of Myrica nagi was dose-dependent. In summary our data suggest that Myrica nagi is an effective chemopreventive agent in skin and capable of ameliorating cumene hydroperoxide induced cutaneous oxidative stress and toxicity. PMID- 10862503 TI - Selegiline metabolism and cytochrome P450 enzymes: in vitro study in human liver microsomes. AB - Although being a drug therapeutically used for a long time, the enzymatic metabolism of selegiline has not been adequately studied. In the current work we have studied the cytochrome P450 (CYP)-catalyzed oxidative metabolism of selegiline to desmethylselegiline and 1-methamphetamine and the effects of selegiline, desmethylselegiline and 1-methamphetamine on hepatic CYP enzymes in human liver microsomes in vitro. The apparent Km values for desmethylselegiline and 1-methamphetamine formation were on an average 149 microM and 293 microM, and the apparent Vmax values, 243 pmol/min./mg and 1351 pmol/min./mg, respectively. Furafylline and ketoconazole, the known reference inhibitors for CYP1A2 and CYP3A4, respectively, inhibited the formation of desmethylselegiline with Ki value of 1.7 microM and 15 microM. Ketoconazole inhibited also the formation of 1 methamphetamine with Ki of 18 microM. Fluvoxamine, an inhibitor of CYP1A2, CYP2C19 and CYP3A4, inhibited the formation of desmethylselegiline and 1 methamphetamine with Ki values of 9 and 25 microM, respectively. On the basis of these results we suggest that CYP1A2 and CYP3A4 contribute to the formation of desmethylselegiline and that CYP3A4 participates in the formation of 1 methamphetamine. In studies with CYP-specific model activities, both selegiline and desmethylselegiline inhibited the CYP2C19-mediated S-mephenytoin 4' hydroxylation with average IC50 values of 21 microM and 26 microM, respectively. The Ki for selegiline was determined to be around 7 microM. Selegiline inhibited CYP1A2-mediated ethoxyresorufin O-deethylation with a Ki value of 76 microM. Inhibitory potencies of selegiline, desmethylselegiline and 1-methamphetamine towards other CYP-model activities were much lower. On this basis, selegiline and desmethylselegiline were shown to have a relatively high affinity for CYP2C19, but no evidence about selegiline metabolism by CYP2C19 was obtained. PMID- 10862504 TI - Ultrastructural evidence of the protective effect of Na+/H+ exchange inhibition on the in vitro damage induced by ischaemia reperfusion in the interventricular septum of the rabbit heart. AB - We investigated the effects of the Na+/H+ antiporter inhibitor, dimethylamiloride, on myocardial injury after 1 h global ischaemia and 30 min. reperfusion in the isolated arterially perfused interventricular septum of the rabbit heart. After ischaemia and reperfusion challenge, dimethylamiloride significantly increased the recovery of developed tension in a dose-dependent manner, and significantly decreased the maximal increase in resting tension. Ultrastructural analysis of myocytes submitted to the experimental in vitro model supported functional maintenance of physiologically-like conditions. Where myocardial portions were submitted to ischaemic conditions and reperfusion, myocyte cell damage reached usual characteristics of infarct-like induced lesions. Intracellular oedema, severe disruption of myofibrils with loss of muscle striation and both swelling and fragmentation of mitochondria were the main characteristics observed. Dimethylamiloride treatment clearly modifies ultrastructural findings towards the normalization of cell shape and structure, only a slight-middle intracellular oedema and contraction bands were found. On the basis of the present results, we suggest that the protective effects exhibited by dimethylamiloride on the ischaemic myocardium are compatible with its Na+/H+ antiporter inhibition properties, they diminish Na+ accumulation and then either Ca2+ overload or non-exocytotic noradrenaline release during the ischaemia and reperfusion challenge. PMID- 10862505 TI - Hydrolytic and metabolic characteristics of the esters of 1-(3'-hydroxypropyl)-2 methyl-3-hydroxypyridin-4-one (CP41), potentially useful iron chelators. AB - 1-(3'-Hydroxypropyl)-2-methyl-3-hydroxypyridin-4-one (CP41) has been extensively investigated as an orally effective iron chelator. In order to improve the pharmacokinetic and metabolic properties of CP41, eleven aromatic esters have been synthesised and tested as potential prodrugs. In the present study, the hydrolytic rates of these CP41 esters in phosphate buffer (pH2.0 and pH 7.4), rat blood and rat liver homogenate have been determined and found to cover a wide range. Generally, they possessed relatively slow hydrolytic rates in phosphate buffer (0-50 nmol/ml/hr at pH 2.0 and 0-140 nmol/ml/hr at pH 7.4). The hydrolytic rates in rat blood fell in the range of 9-5766 nmol/ml blood/hr and in rat liver homogenate 1-800 micromol/g liver tissue/hr. All esters possess a higher lipophilicity than that of the parent compound CP41. Although no apparent relationship was observed between the lipophilicities and hydrolytic rates, the esters with relatively higher hydrolytic rates in liver homogenate tend to possess higher iron scavenging efficacies. Further investigation of the metabolism of selected CP41 esters indicates that metabolism is a key factor influencing the efficacy of CP41 esters, as some esters can be metabolically inactivated in the liver in preference to undergoing ester hydrolysis. Ester design, combined with a knowledge of the prodrug metabolism, is a useful strategy for the production of 3-hydroxypyridin-4-ones with enhanced iron scavenging efficacy. PMID- 10862506 TI - Dose-dependent protection by lipoic acid against cisplatin-induced nephrotoxicity in rats: antioxidant defense system. AB - This study was designed to investigate the role of graded doses of lipoic acid pretreatment against cisplatin-induced nephrotoxicity. Male Wistar rats were divided into six groups and treated as follows: 1) vehicle (saline) control; 2) cisplatin (16 mg/kg, intraperitoneally); 3) lipoic acid (100 mg/kg, intraperitoneally); 4) cisplatin plus lipoic acid (25 mg/kg); 5) cisplatin plus lipoic acid (50 mg/kg) and 6) cisplatin plus lipoic acid (100 mg/kg). Rats were sacrificed three days after treatment, and plasma as well as kidneys were isolated and analyzed. Plasma creatinine increased (677% of control) following cisplatin administration alone which was decreased by lipoic acid in a dose dependent manner. Cisplatin-treated rats showed a depletion of renal glutathione (GSH), increased oxidized GSH and decreased GSH/GSH oxidized ratio (62%, 166% and 62% of control), respectively which were restored with lipoic acid pretreatment. Renal superoxide dismutase, catalase, glutathione peroxidase (GSH peroxidase) and glutathione reductase activities decreased (62%, 75%, 62% and 80% of control), respectively, and malondialdehyde content increased (204% of control) following cisplatin administration, which were restored with increasing doses of lipoic acid. The renal platinum concentration increased following cisplatin administration, which was possibly decreased by chelation with lipoic acid. The data suggest that the graded doses of lipoic acid effectively prevented a decrease in renal antioxidant defense system and prevented an increase in lipid peroxidation, platinum content and plasma creatinine concentrations in a dose dependent manner. PMID- 10862507 TI - CYP1A1 but not CYP1A2 proteins are expressed in human lymphocytes. PMID- 10862508 TI - K-Ras mutations and benign pancreatic disease. AB - This review addresses the history of the ras oncogene, the techniques used to detect molecular alterations in the ras oncogene, and the application of polymerase chain reaction (PCR)-based methods to determine point mutations in clinical samples of patients with pancreatic diseases, namely pancreatic carcinoma and chronic pancreatitis. The frequency of ras mutations in pancreatic carcinoma is high, ranging from 70 to almost 100%. The frequence of ras mutations in chronic pancreatitis, either in pancreatic tissue or pancreatic secretions, vary between 0 and 100%. This wide range in part may be owing to differences in sampling, DNA extraction, or PCR method. The meaning of a k-ras mutation is under debate. Taking into account the positivity of ductal hyperplasias in normal pancreas and ras mutations in normal appearing duct cells, this molecular finding may not mean anything. In contrast, ras mutations are associated with smoking, one acknowledged risk factor for pancreatic carcinoma. The need for large prospective cohort studies is emphasized. PMID- 10862509 TI - Metabolism of the hamster pancreatic carcinogen methyl-2-oxopropylnitrosamine by hamster liver and pancreas. AB - BACKGROUND: The mechanism whereby methyl-2-oxopropylnitrosamine (MOP) is activated remains unknown. To begin investigating this mechanism, we followed MOP disappearance during its incubation with liver and pancreatic slices and homogenates from Syrian hamsters and rats. METHODS: After the incubations, disappearance of 100 microM MOP and appearance of a metabolite was followed by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection. RESULTS: Disappearance rates were 1.2 nmol/mg protein/h for hamster liver slices; zero for hamster pancreatic slices, ducts and acini; zero for rat liver and pancreatic slices; and 11.8, 12.8, 1.3, and 2.3 nmol MOP/mg/h for hamster liver homogenate and cytosol, and hamster pancreas homogenate and microsomes, respectively. The principal MOP metabolite was identified as methyl-2 hydroxypropylnitrosamine (MHP) by its HPLC behavior and its 1H-NMR and mass spectra. MHP yields were generally similar to MOP consumption, but were zero for hamster pancreatic homogenate despite its ability to metabolize MOP. CONCLUSION: MOP is a pancreatic carcinogen in hamsters but not in rats. In metabolic studies, hamster liver slices and homogenate (especially the cytosol) produced MHP from MOP. This is probably an inactivation reaction. Hamster pancreas homogenate (especially the microsome fraction), but not rat pancreas homogenate, metabolized MOP without forming MHP, indicating another route of metabolism, perhaps activation to give the proximal carcinogen. PMID- 10862510 TI - Immunohistochemical analysis of apoptosis-related proteins in human embryonic and fetal pancreatic tissues. AB - BACKGROUND: The growth of both cancer cells and fetal tissue is rapid; however, cancer cells de-differentiate and proliferate in a disorderly manner, whereas fetal tissues differentiate and proliferate in an orderly manner. Thus, there may be both common and different factors that are involved in the process of the uncontrolled cell growth of pancreatic cancers and the development of the fetal pancreas. The common part of the mechanisms should be in the regulation of the cell cycle, resulting in rapid proliferation via such mechanisms as growth stimulation and avoidance of apoptosis. Therefore, in the current study we investigated the expression of apoptosis-related proteins in fetal pancreatic tissues. METHODS: Sixteen human embryonic and fetal pancreatic tissues obtained between 6 and 32 wk of gestation were used. We immunohistochemically examined the protein expression of Bcl-2, Bcl-XL, Mcl-1, and Bax. Further, the expression of insulin, glucagon, and proliferting cell nuclear antigen (PCNA), and TdT-mediated dUTP-biotin nick-end labeling (TUNEL) staining were examined. RESULTS: In embryonic and fetal pancreatic tissues, Bcl-2 was not detected in any type of pancreatic cell (acinar, ductal, or islet). Bcl-XL was expressed in all types of pancreatic cells throughout the gestation. Mcl-1 was expressed in all types of pancreatic components, and strongly expressed in the margin of the islets. Bax, a pro-apoptotic protein, was expressed in all components. PCNA was strongly expressed in the embryonic and fetal pancreas, especially in early stages of gestation; however, TUNEL staining was negative in all samples. At least one antiapoptotic protein was expressed in all types of pancreatic cells. CONCLUSION: The results of the current study indicate that active proliferation and avoidance of apoptosis take place in embryonic and fetal pancreatic tissues, which may be controlled by particular combinations of apoptosis-related proteins. Among these proteins, Bcl-XL and Mcl-1 may play an important role in the proliferation and differentiation of the embryonic and fetal pancreas. PMID- 10862511 TI - Effect of herbal medicine Saiko-keishi-to (TJ-10) on rat spontaneous chronic pancreatitis: comparison with other herbal medicines. AB - BACKGROUND: In an attempt to obtain evidence of the beneficial effects of TJ-10, we investigated the gene expression of PAP, an acute phase protein specific for pancreatitis in rat spontaneous chronic pancreatitis. METHODS: Four-wk-old male WBN/Kob rats were fed with MB-3 pellet diet containing herbal medicine. There were two administration groups for each drug: the prophylactic group administered from 4-12 wk, and the therapeutic group administered from 12-20 wk. Untreated control rats were fed with MB-3 alone. Histopathologic changes and PAP gene expressions were analyzed at 12 and 20 wk. RESULTS: In the prophylactic group, TJ 10-treated WBN/Kob rats showed no evidence of pancreatitis, and there was the amelioration of pancreatitis in the pancreata of the rats treated with other herbal medicines except TJ-24 at 12 wk. PAP mRNA was not expressed in the TJ-10 treated rats, and PAP gene expression was suppressed in rats treated with other drugs except TJ-107. In the therapeutic group, the amelioration of pancreatitis was seen only in TJ-10-treated rats, but PAP gene expression was significantly suppressed in the rats treated with all herbal medicines tested, compared with that in untreated control rats. CONCLUSION: An herbal medicine Saiko-keishi-to (TJ-10) delayed the onset of chronic pancreatitis in the WBN/Kob rat, and suppressed the pancreatitis-associated protein (PAP) gene expression more significantly than other herbal medicines. PMID- 10862512 TI - Long-term results after surgery for chronic pancreatitis. AB - AIM: To determine the early and late morbidity and mortality after surgical treatment of chronic pancreatitis. METHODS: We determined long-term outcome and early and late morbidity and mortality, respectively, in 484 consecutive patients undergoing surgery for chronic pancreatitis from 1976 through 1997. Sixty-five percent of the patients had small duct disease (main pancreatic duct <7 mm), whereas 35% had large duct disease. Indications for operation were pain (95%), suspicion of malignancy (28%), and complications involving adjacent organs (35%). Pseudocysts were present in 27% of patients. Hospital morbidity (8 vs 23%, p = 0.0002) and mortality (0 vs 1.9%, p = 0.12) were less after drainage procedures (n = 162) than after pancreatic resections (n = 286). Among resectional procedures, total pancreatectomy had the highest 30-d operative mortality (5%) and morbidity rates (47%), followed by pancreatoduodenectomy (3 and 32%, respectively). The best results with pain relief occurred after proximal pancreatic resection (89% after mean follow-up of 6.5 yr). The number of patients able to function normally after surgical treatment increased from 39 to 79% (p < 0.001). Long-term survival of our patients was lower than expected rates based on Minnesota life tables analysis (p < 0.0001) especially in alcoholics. Patients undergoing a ductal drainage procedure had the longest survival, whereas those after total pancreatectomy had the shortest survival (p = 0.06). Pancreatic insufficiency, peptic ulcer, and/or anastomotic ulcers caused significant morbidity after total pancreatectomy and pancreatoduodenectomy. A small percentage (3%) developed pancreatic cancer. CONCLUSIONS: Operative treatment of chronic pancreatitis, when indicated, can be performed safely with good results in terms of pain relief and quality of life. Resectional procedures (especially total pancreatectomy) are associated with higher early and late morbidity, greater perioperative mortality, and lower survival rates compared with drainage procedures. Abstinence from alcohol is associated with longer survival rates, which, however, still remain lower than expected rates. PMID- 10862513 TI - Endoscopic treatment of the main pancreatic duct: correlations among morphology, manometry, and clinical follow-up. AB - BACKGROUND AND AIM: During the course of chronic pancreatitis, the gradual increase in the main pancreatic duct pressure is the main pathophysiological factor responsible for pain, but up to now, the intra ductal pressure has never been measured during and after endoscopic stenting and correlated with clinical results. Pressure measurements of this kind could thus provide objective information about the useful duration of stenting period. METHODS: Main pancreatic duct pressure was measured by performing endoscopic manometry on 13 chronic pancreatitis symptomatic patients (10 men, 3 women, mean age: 45.1+/-7.9 yr); clinical follow-up was carried out for a period of 29.0+/-16.1 mo. Before treatment, the main anatomical alteration present was a localized stenosis of the main pancreatic duct, i.e., one with a diameter of less than 2 mm (chronic pancreatitis alone), 10 cases; chronic pancreatitis associated with pancreas divisum, 3 cases). Stenosis was treated by endoscopic stenting: 7 F stent (7 cases) and 12 F stent (6 cases). The pressure was measured simultaneously in the duodenum (zero level) and within the main pancreatic duct, using an electronic device, The pancreatico-duodenal gradient was taken to be the difference between the pressure in the main pancreatic duct and the duodenum. RESULTS: The endoscopic stenting induced a nonsignificant decrease in the intraductal pressure (p = 0.16). Among the 9 patients with a normal pressure at the end of the stenting and a successful anatomical outcome, 6 were painless during the follow up period whereas 3 presented with recurrent pancreatic-type pain. The remaining 4 patients were symptom-free during the entire follow-up period, although the main pancreatic duct pressure was high at the end of the stenting and the stenosis was not completely cured. CONCLUSION: The intraductal pressure at the end of the stenting period was perfectly correlated with the anatomical result, whether or not it was successful, but was not an accurate predictor of a favorable clinical outcome in patients with a poor anatomical result. PMID- 10862514 TI - The effect of chronic exercise on the rat pancreas. AB - BACKGROUND: We recently demonstrated that chronic physical exercise increases pancreatic protein content and basal amylase secretion. It is unknown whether chronic exercise causes hypertrophy or proliferation of pancreatic acinar cells. METHODS: Female F344 rats (age, 6 wk) were divided into control (n = 7) and exercise (n = 6) groups. Food consumption was matched between the 2 groups. Rats in the control group were kept sedentary. Rats in the exercise group were exercised for 60 min, 5 d/wk during the experiment. After 8 wk, the pancreas and hindlimb muscles were rapidly excised and weighed. Protein and DNA content and enzyme activity in pancreatic tissue were measured. Pancreatic tissues from control and exercised rats were also prepared for transmission electron microscopy. RESULTS: Inhibition of growth and hypertrophy of hindlimb muscles were exhibited by the exercise group. In the exercise group, pancreatic wet weight, protein content, and amylase and lipase activities, but not DNA content, were significantly higher than those in the control group. Electron micrographs clearly revealed that acinar cells were hypertrophied and zymogen granules were increased in number in exercised rats. CONCLUSION: Chronic endurance exercise increases pancreatic weight, protein content and enzyme activity through hypertrophy of acinar cells. PMID- 10862515 TI - The effect of interleukin-6 on bacterial translocation in acute canine pancreatitis. AB - BACKGROUND: Bacterial translocation from the gut to mesenteric lymph nodes and other extraintestinal sites is an important source of infection in acute pancreatitis. Impaired host immunity is known to promote bacterial translocation. Interleukin-6 (IL-6) is a multifunctional cytokine that regulates the immune response, acute phase reaction, and hematopoiesis. METHODS: Twenty-four mongrel dogs (18-29 kg) were studied in four equal groups. In Groups I and II, acute pancreatitis was induced by direct pressure injection of 4% taurocholate and trypsin into the pancreatic duct at laparotomy. Groups III and IV had only laparotomy. Group I and III dogs were given IL-6 (50 microg/kg/d, sq) daily starting 24 h after operation and Group II and IV dogs received an equal volume of saline administered at similar time. All animals had blood drawn for culture, complete blood count (CBC), platelets, erythrocyte sedimentation rate (ESR), C reactive protein (CRP), and amylase on d 0, 1, 4, and 7. On d 7, mesenteric lymph nodes (MLN), spleen, liver, pancreas, and cecum were harvested for pathology study and for cultures of aerobic and anaerobic bacteria. Quantitative cecal cultures of aerobic and anaerobic bacteria were obtained. RESULTS: All Group I and Group II dogs had severe pancreatitis. The increase of plasma CRP in Group I was sustained throughout treatment (1.3+/-0.3 on d 0 vs 3.1+/-0.3*, 3.0+/-0.3*, and 2.9+/-0.3* on d 1,4, and 7, respectively). Plasma CRP was increased in Group II on d 1 and d 4 (1.3+/-0.3 mg/dL on d 0 vs 3.6+/-0.3* mg/dL on d 1, and 3.1+/ 0.3* on d 4, *p < 0.05). There were no differences in white blood cell (WBC) count, differential, platelets, and ESR between Groups I and II. Bacterial translocation to MLN was lower in Group I (1/6) than in Group II (6/6) (p < 0.05). All 6 dogs in Group II had bacterial spread to distant sites compared to 2 of 6 dogs in Group I (p = 0.066). Both MLN and other distant organ cultures were negative in Group III and only 1 of 6 MLN cultures was positive in Group IV. CONCLUSIONS: IL-6 treatment decreases bacterial translocation to MLN and may be beneficial in reducing septic complications in acute pancreatitis. PMID- 10862516 TI - Ectopic pancreas: a rare cause of pyloric stenosis. PMID- 10862517 TI - Locoregional complications of pancreatic necrosis. PMID- 10862518 TI - Suggestions for the nomenclature of human alleles: relevance to ecogenetics, pharmacogenetics and molecular epidemiology. AB - The current number of 9422 symbols for human gene names (http://www.gene.ucl.ac.uk/nomenclature/) is expected to increase 7- to 15-fold over the next 2 years. In and around each gene, a tremendous degree of single nucleotide polymorphism (SNP) heterogeneity is now realized to exist. This review is intended to be visionary, to point out some of the enormously complex nomenclature issues that we face, and to offer some reasonable solutions to these issues. For example, I believe that a 'gene' should be defined as that region from the furthest 5'-ward enhancer to at least 150 bases downstream of the last exon. Just as established rules are critically important for the systematic naming of all new genes, standardized nomenclature rules for the naming of allelic variants are also desperately needed. The evolving consensus for naming the alleles of all human genes (ideally based on evolutionarily diverging haplotype patterns) is described herein. Because of the anticipated explosion in finding new genes and allelic variants due to high-throughput resequencing and DNA-chip technologies, this excess of new knowledge will undoubtedly overwhelm their publication by scientific journals alone. I suggest that the best approach to this staggering 'information overload' is to place the data on appropriate web sites--with numerous links between sites, and frequent updates of all information -so that colleagues in all fields of medical and genetic research can remain knowledgeable. Examples of successful web sites to date include those for the cytochrome P450 (CYP) genes and human CYP alleles, UDP glycosyltransferase (UGT) genes and human alleles, human N-acetylaminotransferase (NAT2, NAT1) alleles, and aldehyde dehydrogenase (ALDH) genes and human alleles. Many more web sites will be necessary. For each site, the webmaster will need to be responsible, accurate, energetic, highly organized, and keen to keep the site current. I believe that interactive discussions on these sites should be encouraged, and advisory committees must be willing to check frequently to ensure that all new information is accurate. Lastly, for the field of molecular epidemiology, the importance of correlating an informative genotype with an unequivocal phenotype is emphasized, and the emerging realization that racial and ethnic groups are highly admixed is summarized and updated. PMID- 10862519 TI - Update on consensus arylamine N-acetyltransferase gene nomenclature. PMID- 10862520 TI - Molecular analysis of the N-acetyltransferase 1 gene (NAT1*) using polymerase chain reaction-restriction fragment-single strand conformation polymorphism assay. AB - One major interest to analyse the extent of N-acetyltransferase 1 (NAT1*) allelic variation in the human population stems to a great extent from the possible association of interindividual differences in the metabolism of aromatic amines with certain chemically induced diseases, including cancer. Considering the increasing number of mutations in the NAT1 gene that are detected, NAT1* genotyping using conventional polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) or allele-specific amplification assays has become complicated. We developed a rapid and powerful strategy allowing the full characterization of NAT1* alleles. This method, based on single-strand conformation polymorphism analysis of a unique PCR product encompassing the entire intronless NAT1*-coding region along with additional flanking segments in the 5' and 3' untranslated regions, was then applied to DNA samples from 270 individuals. Nine NAT1* allelic variants, including two novel (NAT1*28 and NAT1*29), and 15 different genotypes were identified. This approach could be advantageously used in epidemiological studies to provide more definite data on suspected associations between NAT1* genotype and certain pathological processes. PMID- 10862521 TI - Alcohol consumption, glutathione S-transferase M1 and T1 genetic polymorphisms and breast cancer risk. AB - To evaluate the potential association between GSTM1 and GSTT1 genotypes and development of breast cancer, a hospital based case-control study was conducted in a South Korean study population consisting of 189 histologically confirmed incident breast cancer cases and their 189 age-matched control subjects with no present or previous history of cancer. A multiplex polymerase chain reaction method was used for the genotyping analyses and statistical evaluations were performed by unconditional logistic regression model. The GSTM1 null genotype was significantly associated with breast cancer risk in premenopausal women [odds ratio (OR) = 2.0, 95% confidence interval (CI) = 1-3.7], but not in the postmenopausal women (OR = 0.9, 95% CI = 0.5-1.9), nor in all women grouped together (OR = 1.3, 95% CI = 0.8-1.1). The GSTT1 null genotype posed a similar risk of breast cancer with an OR of 1.6 (95% CI = 1.0-2.5) for the total breast cancer group, OR of 1.7 (95% CI = 0.9-3.2) for pre-menopausal women, and OR of 1.3 (95% CI = 0.6-2.8) for post-menopausal women. The breast cancer risk associated with concurrent lack of both GSTM1 and GSTT1 genes was 2.2 (95% CI = 1.1-4.5), and the risk increased as the number of null genotype increased (P for trend = 0.03). When the data were stratified by the known risk factors of breast cancer, a significant interaction was observed between the GSTM1 genotypes and alcohol consumption (P for interaction = 0.03). An especially remarkable risk of breast cancer was observed for alcohol-consuming premenopausal women lacking both the GSTM1 and GSTT1 genes (OR = 5.3, 95% CI = 1.0-27.8) compared to those with both of the genes. Our findings thus suggest a novel gene-environment interaction which plays an important role in the individual susceptibility to breast cancer. p6 PMID- 10862522 TI - The relationship between genotype and chromosome aberration frequencies in a normal adult population. AB - Cancer susceptibility differences may be attributed in part to genetic variation in genes involved in metabolism of environmental procarcinogens. Increased risks for some cancers have been linked to polymorphisms in certain phase I and II genes, and have been associated with genomic instability and chromosomal aberrations. Aberration frequencies in general, and stable aberration frequencies (translocations and insertions) in particular, are used as biomarkers for disease. Thus, knowledge of the genetic factors that influence the frequency of stable aberrations in a normal population is important for cancer risk determination. In this work, genotypes for a number of xenobiotic enzymes (CYPIA1, CYP2D6, GSTM1, GSTT1, GSTP1, NAT1, NAT2 and epoxide hydrolase) and stable aberration frequencies were determined for 65 normal individuals aged 19 77 years. The population was divided at age 60 years for analysis because there was a significant difference in stable aberration frequencies between these groups. Subjects with low levels (0-66th percentile) of stable aberrations were compared to those with high levels (67th percentile and above). Of all the genotypes studied, only NAT2 showed a notable difference between the high and the low stable aberration groups in the percentage of polymorphisms observed, and this was seen only in the older subjects group. All individuals in the older-high stable aberration group were NAT2 rapid acetylator smokers. NAT2 slow acetylator smokers had significantly lower stable aberration frequencies compared to the NAT2 rapid acetylator smokers. Following previous work showing an increased risk of cancer associated with high levels of aberrations (above the 66th percentile), we hypothesize that smokers with the NAT2 rapid acetylator genotype may be at an increased risk for cancer. PMID- 10862523 TI - The human peroxisome proliferator-activated receptor alpha gene: identification and functional characterization of two natural allelic variants. AB - Peroxisome proliferator-activated receptor (PPAR)alpha-null mice have a defect in fatty acid metabolism but reproduce normally. The lack of a detrimental effect of the null phenotype in development and reproduction opens up the possibility for null or variant PPARalpha gene (PPARA) alleles in humans. To search the coding region and splice junctions for mutant and variant PPARalpha alleles, the human PPARalpha gene was cloned and characterized, and sequencing by polymerase chain reaction was carried out. Two point mutations in the human gene were found in the DNA binding domain at codons for amino acids 131 and 162. The allele containing the mutation in codon 162 (CTT to GTT, L162V) designated PPARA*3, was found at a high frequency in a Northern Indian population. Transfection assays of this mutant showed that the non-ligand dependent transactivation activity was less than one-half as active as the wild-type receptor. PPARA*3 was also unresponsive to low concentrations of ligand as compared to the wild-type PPARA*1 receptor. However, the difference is ligand concentration-dependent; at concentrations of the peroxisome proliferator Wy-14 643 > 25 microM, induction activity was restored in this variant's transactivation activity to a level five-fold greater as compared with wild-type PPARA*1 with no ligand. The mutation in codon 131 (CGA to CAA, R131Q), designated PPARA*2 is less frequent than PPARA*3, and the constitutive ligand independent activity was slightly higher than PPARA*1. Increasing concentrations of Wy-14 643 activated PPARA*2 similar to that observed with PPARA*1. The biological significance of these novel PPARalpha alleles remains to be established. It will be of great interest to determine whether these alleles are associated with differential response to fibrate therapy. PMID- 10862524 TI - The relationship between dopamine D2 receptor polymorphism at the Taq1 A locus and therapeutic response to nemonapride, a selective dopamine antagonist, in schizophrenic patients. AB - Previous studies have demonstrated that subjects with one or two A1 alleles of dopamine D2 receptor (DRD2) polymorphism at the Taq1 A locus have lower DRD2 density than those with no A1 allele. The present study aimed to examine whether the Taq1 A DRD2 genotypes are related to therapeutic response to nemonapride, a selective dopamine antagonist, in schizophrenic patients. The subjects were 25 acutely exacerbated schizophrenic inpatients who had received no medication for at least 1 month before the study. The fixed dose (18 mg/day) of nemonapride was administered to each patient for 3 weeks. The clinical status was prospectively monitored by the Brief Psychiatric Rating Scale (BPRS) before, and 3 weeks after, the treatment. The Taq1 A genotypes (A1 and A2 alleles) were determined by the polymerase chain reaction method. Three patients were homozygous for the A1 allele, 11 were heterozygous for the A1 and A2 alleles, and 11 were homozygous for the A2 allele. The patients with one or two A1 alleles (n = 14) showed significantly higher percentage improvement in total BPRS and positive symptoms than those with no A1 allele (n = 11) after 3-week treatment while the percentage improvement in other subgrouped symptoms (negative, anxiety-depression, excitement and cognitive symptoms) was similar between the two genotype groups. The present results suggest that the Taq1 A DRD2 polymorphism is related to early therapeutic response to nemonapride in schizophrenic patients, possibly by modifying the efficiency of DRD2 antagonism of the drug in the central nervous system. PMID- 10862525 TI - Polymorphisms in P450 CYP1B1 affect the conversion of estradiol to the potentially carcinogenic metabolite 4-hydroxyestradiol. AB - Most drug metabolizing cytochrome P450s (P450) are predominantly expressed in the liver. In contrast, human CYP1B1 is an extrahepatic P450 which is overexpressed in many tumours and has been strongly implicated in the activation of carcinogens. Rare allelic variants of the CYP1B1 gene which encode an inactive protein have been identified. However, four polymorphisms which most likely do not abolish functionality have been described. In this report, we have characterized the functional consequences of these. A CYP1B1 cDNA, identical to a cDNA published previously, served as a template to introduce allelic changes either separately or in combination. The resulting effects on CYP1B1 activity were determined in membranes isolated from Escherichia coli which coexpressed CYP1B1 together with P450 reductase. None of the allelic changes affected the CYP1B1 expression level. The allelic changes Arg48 to Gly, Ala19 to Ser and Asn453 to Ser had little influence on the Vmax and the Km of the CYP1B1 mediated 2- and 4-hydroxylation of estradiol. In contrast, the Km of these metabolic pathways was increased at least three-fold by the allelic change Va432 to Leu or by simultaneously changing Val432 to Leu and Asn453 to Ser. However, these alterations had little effect on the kinetic parameters of other CYP1B1 mediated reactions such as the epoxidation of (-)-trans-(7R,8R)-benzo[a]pyrene 7,8 dihydrodiol as determined by (r-7,t-8,t-9,c-10)-benzo[a]pyrene tetraol formation, or such as the O-dealkylation of ethoxyresorufin and the 1'-hydroxylation of bufuralol. Molecular modelling suggests that amino acid residue 432 of CYP1B1 may be involved in the interaction between CYP1B1 and P450 reductase. Since 4 hydroxyestradiol has been implicated in hormonal carcinogenesis and CYP1B1 is expressed in target tissues, the data presented demonstrate that polymorphisms in CYP1B1 have the potential to affect disease susceptibility. PMID- 10862526 TI - Molecular genetic basis for deficient acetaminophen glucuronidation by cats: UGT1A6 is a pseudogene, and evidence for reduced diversity of expressed hepatic UGT1A isoforms. AB - The domestic cat has a significantly lower capacity to glucuronidate planar phenolic xenobiotics compared with most other mammalian species. The aim of this study was to determine the mechanistic basis for this anomaly. Current knowledge of the substrate specificity of UDP-glucuronosyltransferase (UGT) isoforms indicates that the cat may either lack or poorly express UGT1A6. Initially, a novel cloning technique was used to identify UGT1A genes expressed in cat liver. Only two unique UGT1A isoforms could be discriminated. The first (28%, of clones) was most homologous to UGT1A1 (the bilirubin-UGT), while the second (72% of clones) showed homology to several isoforms, but could not be unambiguously identified, and was designated cat UGT1A02. Southern blot analysis confirmed the presence of a single UGT1A6-homologous region in the cat genome. Subsequent cloning and sequencing of the entire UGT1A6 exon 1 coding region revealed five deleterious genetic mutations. Identical mutations were found by sequencing of UGT1A6 exon 1 from five other unrelated cats. Four of these five genetic lesions were also identified in the UGT1A6 exon 1 region of a margay (Leopardus wiedii). Finally, RT-PCR of liver mRNA from four different cats confirmed the presence of UGT1A1 and UGT1A02, but not UGT1A6. In conclusion, UGT1A6 is a pseudogene in the domestic cat and in at least one other phylogenetically related species. Furthermore, cats appear to have a less diverse pattern of UGT1A isoform expression compared with other species. Such differences most likely reflect the highly carnivorous diet of Feliform species and resultant minimal exposure to phytoalexins. PMID- 10862527 TI - Retraining of a competitive master athlete following traumatic injury: a case study. AB - PURPOSE: The purpose of this study was to examine the physiological effects of detraining and retraining in a female master cyclist (age, 49.5 yr; wt, 54 kg) following a surgically-treated clavicular fracture complicated by brachial plexus impingement. METHODS: Variables associated with cycling performance, including VO2max, lactate threshold (LT), power output at a blood lactate concentration of 4 mM (LT(4 mM)), peak power output (PPO), muscular resistance to fatigue measured by a timed ride to exhaustion at 110% of peak power output (PPO110), and body composition (hydrostatic weight) were assessed 2 d before the injury when the subject was at the peak of her competitive season, and at days 0, 14, 28, 42, and 77 of the retraining period. Retraining gradually increased from 3 h x wk(-1) to 9-10 h x wk(-1) with an increase in intensity from approximately 70 to 95+% of HRmax. RESULTS: Detraining resulted in a 25.7% decrease in VO2max and a 16.7% and 18.9% decrease in LT and LT(4 mM), respectively, while peak power output and PPO110 declined 18.2% and 16.6%, respectively. Body fat percent increased 2.1 percentage points, while fat-free mass decreased nearly 2 kg. After 2 wk of retraining, all variables except the LT and LT(4 mM) had improved considerably; however, VO2max was still 14.8% lower and PPO and PPO110 were 12.7% and 5.7% lower than preinjury values. By the 11th week of retraining, all variables had essentially returned to their preinjury values. CONCLUSION: These data demonstrate a pattern of retraining in which aerobic power steadily improved over 6 wk, while measures of lactate threshold did not change until the fourth week of retraining when the intensity of training was markedly increased. Additional data are needed to determine whether this pattern of retraining would be consistent in other master athletes. PMID- 10862528 TI - Effect of exercise intensity on bone density, strength, and calcium turnover in older women. AB - PURPOSE: This study examined the effects of 24 wk of high intensity strength training or low intensity walking on lumbar bone mineral density (BMD), muscular strength, and calcium turnover in Australian women either taking hormone replacement therapy (HRT) or not taking HRT. METHODS: A subject pool of 64 women between 45-65 yr and randomly allocated into weights (N = 21), walking (N = 20), weightsHRT (N = 14), and walkingHRT (N = 9) groups completed this study. All subjects trained twice weekly in either a 50-min walking or weight-training program (60-90% IRM). Measurements included maximal isometric knee strength, IRM bench press, IRM squat, isokinetic back strength, lumbar (L2-L4) BMD, serum osteocalcin, and urinary deoxypyridinoline crosslinks (Dpd). RESULTS: No significant group differences in BMD were evident at the completion of training. However, a significant (P < 0.05) within group change was apparent for the walking group since BMD decreased 1.3% below baseline testing. Osteocalcin levels increased significantly (P < 0.05) in the walking (22%) group. Maximal bench press and squat strength improved significantly (P < 0.05) in the weights (25.8% and 37.7%) and weightsHRT (25.4% and 35.7%) groups. The weights group also increased significantly (P < 0.05) in isokinetic back strength (22.2%). CONCLUSION: It was concluded that short-term high intensity resistance training provides an effective means for increasing muscular strength in women between 45 and 65 yr. The training effects on lumbar BMD were not apparent in the present study. PMID- 10862529 TI - Effects of plyometric jump training on bone mass in adolescent girls. AB - PURPOSE: The purpose of this study was to investigate the effects of 9 months of plyometric jump training on bone mineral content (BMC), lower extremity performance, and static balance in adolescent girls (aged 14.6 +/- 0.5 yr; 22.7 +/- 14.0 months past menarche). METHODS: Exercisers (N = 25) trained 30-45 min, three times per week, performing various exercises using weighted vests (squats, lunges, calf raises) and plyometrics (hopping, jumping, bounding, and box depth jumps). The program was designed to load the lower extremities. Controls (N = 28), matched to exercisers for age and months past menarche, maintained their usual activities. The following were assessed at baseline and 9 months: BMC, strength by isokinetic dynamometry, power (Wingate), and static balance. RESULTS: Repeated measures ANOVA revealed no significant differences between groups for BMC, nor were the changes in anthropometric or performance variables, analyzed by MANOVA, significant. In follow-up analyses, t-tests for independent samples revealed that both groups experienced a significant (P < 0.01) increase in percent change in bone mass compared to zero, for the whole body (mean: 3.7% exercisers, 3.6% controls), femoral neck (4.5% vs 2.4%), lumbar spine (L2-4) (6.6% vs 5.3%), and femoral shaft (3.4% vs 2.3%), but only the exercisers improved BMC of the greater trochanter (3.1% vs 1.9%). Furthermore, the exercise group significantly improved knee extensor strength (14.7% vs 7.3%) and medial/lateral balance (38.1% vs 9.5%), whereas the control group demonstrated no changes. The variety of lateral movement activities performed by the exercise group may have contributed to the differences observed between groups for greater trochanter bone mineral density (BMD), leg strength, and medial/lateral balance. CONCLUSION: The trends observed in bone mass between groups suggest that plyometric jump training continued over a longer period of time during adolescent growth may increase peak bone mass. PMID- 10862531 TI - Cardiac fatigue following prolonged endurance exercise of differing distances. AB - PURPOSE: Recent echocardiographic studies have reported cardiac dysfunction following ultra-endurance exercise in trained individuals. The duration of exercise required to elicit cardiac dysfunction and the mechanisms underlying this phenomenon have not been fully elucidated. The aim of the present study was to examine the presence of cardiac dysfunction following a half-Ironman and Ironman triathlon in trained individuals. METHODS: 14 male triathletes (age: 32 +/- 5 yr; height: 180 +/- 8 cm; body mass: 75 +/- 9 kg) completed a half-Ironman triathlon. Following a 4-wk period, 10 of the original 14 triathletes completed an Ironman triathlon. All triathletes were assessed using ECG, echocardiography, and blood analysis pre-, immediately post-, and 48 h postrace for both distances. RESULTS: Echocardiographic results indicated diastolic and systolic left ventricular dysfunction, for both race distances, which were associated with altered relaxation characteristics and a reduced inotropic contractility, respectively. Following 48-h recovery, all echocardiographic measures were similar to resting values. Creatine kinase MB (CKMB) was significantly elevated immediately postrace for both distances; however, it accounted for less than 5% of the total CK value and in the presence of an elevated total CK and CKMM implied that the elevated CKMB was noncardiac in origin. Troponin-T, however, was significantly elevated immediately postrace for both distances and returned to normal following 48-h recovery indicating myocardial damage. CONCLUSIONS: Ironman and half-Ironman competition resulted in reversible abnormalities in resting left ventricular diastolic and systolic function. Results suggest that myocardial damage may be, in part, responsible for cardiac dysfunction, although the mechanisms responsible for this cardiac damage remain to be fully elucidated. PMID- 10862530 TI - Training in hypoxia: modulation of metabolic and cardiovascular risk factors in men. AB - PURPOSE: This study was designed to determine changes in metabolic and cardiovascular risk factors following normobaric hypoxic exercise training in healthy men. METHODS: Following a randomized baseline maximal exercise test in hypoxia and/or normoxia, 34 physically active subjects were randomly assigned to either a normoxic (N = 14) or a hypoxic (N = 18) training group. Training involved 4 wk of cycling exercise inspiring either a normobaric normoxic (F(IO2) = approximately 20.9%) or a normobaric hypoxic (F(IO2) = approximately 16.0%) gas, respectively, in a double-blind manner. Cycling exercise was performed three times per week for 20-30 min at 70-85% of maximum heart rate determined either in normoxia or hypoxia. Resting plasma concentrations of blood lipids, lipoproteins, total homocysteine, and auscultatory arterial blood pressure responses at rest and in response to submaximal and maximal exercise were measured before and 4 d after physical training. RESULTS: Total power output during the training period was identical in both normoxic and hypoxic groups. Lean body mass increased by 1.4 +/- 1.5 kg following hypoxic training only (P < 0.001). While dietary composition and nutrient intake did not change during the study, both normoxic and hypoxic training decreased resting plasma concentrations of nonesterified fatty acids, total cholesterol, high density lipoprotein (HDL), and low density lipoprotein (LDL) (P < 0.05 - < 0.001). Apolipoproteins AI and B decreased following normoxic training only (P < or = 0.001). Plasma concentrations of resting total homocysteine decreased by 11% following hypoxic training (P < or = 0.05) and increased by 10% (P < 0.05) following normoxic training. These changes were independent of changes in serum vitamin B12 and red cell folate which remained stable throughout. A decreased lactate concentration during submaximal exercise was observed in response to both normoxic and hypoxic training. Hypoxic training decreased maximal systolic blood pressure by 10 +/- 9 mm Hg (P < 0.001) and the rate pressure product by 14 +/- 23 mm Hg x beats x min(-1)/100 (P < or = 0.001) and increased maximal oxygen uptake by 0.47 +/- 0.77 L x min(-1) (P < 0.05). CONCLUSION: Normoxic and hypoxic training was associated with significant improvements in selected risk factors and exercise capacity. The stimulus of intermittent normobaric hypoxia invoked an additive cardioprotective effect which may have important clinical implications. PMID- 10862532 TI - Enhanced cardiovascular hemodynamics in endurance-trained postmenopausal women athletes. AB - PURPOSE: We sought to determine whether older women athletes who had habitually performed vigorous endurance exercise training had higher stroke volumes and cardiac outputs than sedentary postmenopausal women during maximal exercise. METHODS: Seventeen endurance-trained, postmenopausal women athletes (age 65 +/- 4 yr; VO2max 2.11 +/- 0.31 L x min(-1), 38.3 mL x kg(-1) x min(-1)) and 14 sedentary, postmenopausal women (age 63 +/- 5 yr; VO2max 1.41 +/- 0.22 L x min( 1), 23.7 +/- 3.5 mL x kg(-1) x min(-1)) performed maximal treadmill exercise while cardiac output (via acetylene rebreathing) and other cardiovascular hemodynamics were measured. Approximately half of the subjects in each group were on hormone replacement therapy (HRT). RESULTS: The greater VO2max of the athletes was the result of a greater cardiac output (12.8 +/- 1.6 vs. 9.3 +/- 1.4 L x min( 1)) resulting from their significantly larger stroke volume (80 +/- 10 vs 57 +/- 10 mL) at maximal exercise. There were no significant differences in maximal cardiac output or maximal stroke volume related to HRT status in the sedentary women or athletes. CONCLUSIONS: These data indicate that endurance-trained, competitive, postmenopausal women have higher stroke volumes and cardiac outputs during maximal exercise, than their sedentary peers. However, these data suggest that HRT may not affect maximal CV function in sedentary or endurance-trained postmenopausal women. PMID- 10862533 TI - Cardiovascular responses during prolonged exercise at ventilatory threshold in boys and men. AB - PURPOSE: The purpose of this study was to examine the cardiovascular responses during prolonged exercise in boys and men at an intensity set relative to ventilatory threshold (VT). METHODS: Eight boys (10-13 yr) and 10 men (18-25 yr) completed an orientation trial, a maximal exercise test, and a 40-min submaximal exercise bout at an intensity equal to the VO2 at VT (approximately 64.5% VO2max). RESULTS: Heart rate (HR) was higher and stroke volume (SV) was lower in the boys compared with the men (P < or = 0.05). From 10 to 40 min, HR significantly increased 9.5% and 13.6% and SV significantly decreased 8.8% and 11.6% in the boys and men, respectively. Despite the tendency for the changes in HR and SV to be greater in the men, the group-by-time interaction was not significant. Cardiac output was greater in the men (P < or = 0.05) but remained constant over time (P > 0.05). In men, mean arterial blood pressure was higher (P < or = 0.05) and decreased 4.2% over time. In boys, mean arterial blood pressure remained constant, which resulted in a significant group-by-time interaction. Total peripheral resistance (TPR) was significantly higher in the boys and remained constant over time (P > 0.05). From 0 to 40 min, the decrease in plasma volume was significantly greater in the men (-10.2%) than the boys (-5.7%) but was unrelated to the changes in SV in either group (P > 0.05). CONCLUSION: In conclusion, the cardiovascular responses during prolonged exercise are similar in boys and men, although there is a tendency for the magnitude of cardiovascular drift to be greater in the men. PMID- 10862534 TI - Relationship between plasma lactate parameters and muscle characteristics in female cyclists. AB - PURPOSE: In a previous study, we showed that when six different plasma lactate parameters (LPs) were compared, the LP determined by the Dmax method was the best predictor of 1-h cycling performance in women. The present study extended these findings to determine whether or not the relationship between the following six LPs and endurance performance could be explained by their relationship with muscle fiber characteristics: 1) lactate threshold (LT; the power output at which plasma lactate concentration begins to increase above the resting level during an incremental exercise test), 2) LT1 (the power output at which plasma lactate increases by 1 mmol x L(-1) or more), 3) LT(D) (the lactate threshold calculated by the Dmax method), 4) LT(MOD) (the lactate threshold calculated by a modified Dmax method), 5) L4 (the power output at which plasma lactate reaches a concentration of 4 mmol x L(-1)), and 6) LT(LOG) (the power output at which plasma lactate concentration begins to increase when the log([La-]) is plotted against the log(power output)). METHODS: Twelve trained female cyclists (27.3 +/- 5.4 yr) first completed an incremental cycle test to determine both their LPs and peak VO2. One week later, endurance performance was assessed as the average absolute power output maintained during a 1-h endurance test (OHT). Resting muscle was sampled by needle biopsy from m. vastus lateralis and analyzed for fiber type diameter, fiber type percentage, 2-oxoglutarate dehydrogenase (OGDH) activity, and phosphofructokinase (PFK) activity. RESULTS: OHT performance was more strongly correlated with all LPs (r = 0.71-0.89, P < 0.05) than with peak VO2 (L x min(-1), r = 0.65, P < 0.05). OGDH activity, PFK activity, and the percentage of Type I fibers were not related to peak VO2, any of the LPs, or OHT performance. The diameter of the Type II fibers, however, was negatively related to OHT performance (r = -0.77, P < 0.01) and to four of the LPs (r = -59 to 0.86, P < 0.001). CONCLUSIONS: These correlations, which indicate that large Type II fibers may impair endurance performance, may be the result of greater production and/or reduced removal of lactate from the larger, glycolytic Type II fibers. LPs most strongly correlated with Type II fiber diameter were also most strongly correlated with OHT performance. PMID- 10862535 TI - Interleukins 1-beta, -8, and histamine increases in highly trained, exercising athletes. AB - PURPOSE: Exercise-induced hypoxemia (EIH) in highly trained athletes is associated with an increase in histamine release (%H) during exercise. Certain cytokines, known as histamine-releasing factors, are capable of interacting with basophils and/or mast cells to cause the release of histamine. The aim of this study was to determine whether the increased histamine release in highly trained athletes is related to a high plasma level in interleukin-1 beta (IL-1beta), IL 3, or IL-8 in arterial blood. METHODS: These parameters were measured in 11 endurance athletes (23.2 +/- 1.2 yr (mean +/- SEM)) known to develop exercise induced hypoxemia and 11 control subjects (25.0 +/- 1.1 yr) at rest, during an incremental exhaustive exercise test, and at the fifth minute of recovery. RESULTS: Histamine release increased between rest and maximal exercise in the athletes (P < 0.01), showing a strong correlation with EIH (r = 0.76, P < 0.01) and was unchanged in the controls. IL-3 plasma concentration was not altered with training and/or with exercise. Circulating IL-8 levels were not different between trained and untrained subjects at any testing level and increased at maximal exercise in both groups (P < 0.01). IL-1beta plasma levels were higher in athletes than in controls (P < 0.05) at each testing level and increased during exercise only in the athletes (P < 0.05). CONCLUSION: An elevated concentration of IL-1beta in plasma and its association with increased IL-8 levels during exercise may partly explain the increase in %H associated with EIH in highly trained athletes. Histamine, IL-8, and IL-1beta releases during exercise reflect an inflammatory reaction, which is probably involved in EIH. PMID- 10862536 TI - Effect of exercise-induced arterial O2 desaturation on VO2max in women. AB - PURPOSE: We have recently reported that many healthy habitually active women experience exercise induced arterial hypoxemia (EIAH). We questioned whether EIAH affected VO2max in this population and whether the effect was similar to that reported in men. METHODS: Twenty-five healthy young women with widely varying fitness levels (VO2max, 56.7 +/- 1.5 mL x kg(-1) x min(-1); range: 41-70 mL x kg( 1) x min(-1)) and normal resting lung function performed two randomized incremental treadmill tests to VO2max (FIO2: 0.21 or 0.26) during the follicular phase of their menstrual cycle. Arterial blood samples were taken at rest and near the end of each workload during the normoxic test. RESULTS: During room air breathing at VO2max, SaO2 decreased to 91.8 +/- 0.4% (range 87-95%). With 0.26 FIO2, SaO2, at VO2max remained near resting levels and averaged 96.8 +/- 0.1% (range 96-98%). When arterial O2 desaturation was prevented via increased FIO2, VO2max increased in 22 of the 25 subjects and in proportion to the degree of arterial O2 desaturation experienced in normoxia (r = 0.88). The improvement in VO2max when systemic normoxia was maintained averaged 6.3 +/- 0.3% (range 0 to +15%) and the slope of the relationship was approximately 2% increase in VO2max for every 1% decrement in the arterial oxygen saturation below resting values. About 75% of the increase in VO2max resulted from an increase in VO2 at a fixed maximal work rate and exercise duration, and the remainder resulted from an increase in maximal work rate. CONCLUSIONS: These data demonstrate that even small amounts of EIAH (i.e., >3% delta SaO2 below rest) have a significant detrimental effect on VO2max in habitually active women with a wide range of VO2max. In combination with our previous findings documenting EIAH in females, we propose that inadequate pulmonary structure/function in many habitually active women serves as a primary limiting factor in maximal O2 transport and utilization during maximal exercise. PMID- 10862537 TI - Comparison of sympathetic nerve responses to neck and forearm isometric exercise. AB - PURPOSE: Although the autonomic and cardiovascular responses to arm and leg exercise have been studied, the sympathetic adjustments to exercise of the neck have not. The purpose of the present study was twofold: 1) to determine sympathetic and cardiovascular responses to isometric contractions of the neck extensors and 2) to compare sympathetic and cardiovascular responses to isometric exercise of the neck and forearm. METHODS: Muscle sympathetic nerve activity (MSNA), mean arterial pressure (MAP), and heart rate were measured in nine healthy subjects while performing isometric neck extension (INE) and isometric handgrip (IHG) in the prone position. After a 3-min baseline period, subjects performed three intensities of INE for 2.5 min each: 1) unloaded (supporting head alone), 2) 10% maximal voluntary contraction (MVC), and 3) 30% MVC, then subjects performed two intensities (10% and 30% MVC) of IHG for 2.5 min. RESULTS: Supporting the head by itself did not significantly change any of the variables. During [NE, MAP significantly increased by 10 +/- 2 and 31 +/- 4 mm Hg and MSNA increased by 67 +/- 46 and 168 +/- 36 units/30 s for 10% and 30% MVC, respectively. IHG and INE evoked similar responses at 10% MVC, but IHG elicited higher peak MAP and MSNA at 30% MVC (37 +/- 7 mm Hg (P < 0.05) and 300 +/- 48 units/30 s (P < 0.01) for IHG, respectively). CONCLUSIONS: The data indicate that INE can elicit marked increases in MSNA and cardiovascular responses but that it evokes lower peak responses as compared to IHG. We speculate that possible differences in muscle fiber type composition, muscle mass, and/or muscle architecture of the neck and forearm are responsible for these differences in peak responses. PMID- 10862538 TI - Left ventricular volumes and hemodynamic responses to postexercise ischemia in healthy humans. AB - PURPOSE: The purpose of this study was to determine the cardiac mechanisms involved in cardiovascular adjustments during postexercise circulatory occlusion (OCCL). METHOD: Heart rate (HR), mean arterial pressure (MAP), left ventricular end-diastolic (EDV) and end-systolic volumes (ESV), stroke volume (SV), cardiac output (CO), and total peripheral vascular resistance (total peripheral resistance (TPR)) were assessed in nine healthy volunteers during rest and static exercise at 30% of maximum voluntary contraction followed by either OCCL for 3 min or non-OCCL in a randomized crossover protocol. RESULTS: During handgrip, HR (+20%; P < 0.001), CO (+11%; P = 0.003), MAP (+18%; P = 0.001), and TPR (+6%; P = 0.004) increased, SV (-8%; P = 0.001) and EDV (-5%; P < 0.001) decreased, while ESV did not change (P > 0.05). These responses were similar between conditions (P > 0.05). During OCCL, HR, SV, and CO returned to baseline, whereas MAP (+19%; P < 0.001) and TPR (+9%; P = 0.004) remained elevated. EDV (+12%; P < 0.001) and ESV (+23%; P < 0.001) increased in parallel above resting values. CONCLUSION: Activation of muscle metaboreceptors during OCCL increased MAP by elevating TPR. Despite the higher afterload and increased ESV, CO and SV were kept similar to resting values because EDV also increased, implying the involvement of the Frank Starling mechanism. PMID- 10862539 TI - Self-administered questionnaire compared with interview to assess past-year physical activity. AB - PURPOSE: The Modifiable Activity Questionnaire (MAQ) is a physical activity questionnaire shown to be both valid and reliable and was initially designed to be interviewer-administered. After translation and adaptation into French, the objective of the study was to compare past-year physical activity data obtained by self-administration of this questionnaire and by interviewer-administration. METHODS: 84 subjects (22 men, 62 women, age 36-63 yr) enrolled in an ongoing prospective study in France (the SUpplementation en VItamines et Mineraux AntioXydants or SU.VI.MAX study) completed both versions of the questionnaire in a randomized order with a mean (SD) delay of 7.9 (5.8) d between the two modes. Past-year leisure and occupational physical activity were expressed as both h x wk(-1) and MET-h x wk(-1) of activity, and television watching was expressed as h x d(-1). Analysis of variance on ranks was used to compare activity variables obtained by self-administration and interview. Agreement was assessed by nonparametric intraclass correlation coefficients. RESULTS: A significant effect of the mode of administration was found only for past-year leisure physical activity with lower values reported in self-administered conditions compared with interview. However, a high level of concordance between the two modes of administration was observed for all variables as shown by the intraclass correlation coefficients: 0.90 for leisure physical activity (h x wk(-1)), 0.82 for occupational activity (h x wk(-1)), 0.83 for total (leisure and occupational combined) physical activity (h x wk(-1)), and 0.97 for television viewing (h x d( 1)). CONCLUSIONS: The agreement between the two modes of administration of the questionnaire suggests that the self-administered version of the MAQ is a valuable tool to assess past-year physical activity and inactivity in self administered conditions. This instrument could be used in large-scale population studies investigating the relationships between physical activity and health outcomes. PMID- 10862541 TI - Running economy of African and Caucasian distance runners. AB - PURPOSE: Anecdotal evidence suggests an advantageous physiological endowment of the African endurance athlete. Higher fractional utilization of VO2max has been suggested but not measured directly, and investigations of running economy have been inconclusive. The aim of the current study was to measure a) running economy and b) fractional utilization of VO2max, in African and Caucasian 10-km runners of similar body mass. METHODS: Eight African and eight Caucasian runners had no significant difference in mean race time (32.8 +/- 2.8, 32.0 +/- 2.5 min, respectively), body mass (61.4 +/- 7.0, 64.9 +/- 3.0 kg), age, body fat, or lean thigh volume. Caucasian runners were 6 cm taller (P < 0.05). Subjects completed a progressive treadmill VO2peak test. On a separate day, subjects completed two 6 min workloads (16.1 km x h(-1) and 10-km race pace) separated by 5 min. RESULTS: Mean VO2peak was 13% lower in the Africans (61.9 +/- 6.9, 69.9 +/- 5.4 mL x kg( 1) x min(-1), P = 0.01). At 16.1 km x h(-1), the Africans were 5% more economical (47.3 +/- 3.2, 49.9 +/- 2.4 mL x kg(-1) x min(-1), P < 0.05). This difference increased to 8% (P < 0.01) when standardized per kg(0.66). At race pace, the Africans utilized a higher %VO2peak (92.2 +/- 3.7, 86.0 +/- 4.8%, P < 0.01) and had higher HR (185 +/- 9, 174 +/- 11 b x min(-1), P < 0.05) and plasma [ammonia] (113.2 +/- 51, 60.3 +/- 16.9 micromol x L(-1), P < 0.05). Despite the higher relative workload, the plasma [lactate] was not different (5.2 +/- 2.0, 4.2 +/- 1.7 mmol x L(-1), NS). CONCLUSIONS: This study indicates greater running economy and higher fractional utilization of VO2peak in African distance runners. Although not elucidating the origin of these differences, the findings may partially explain the success of African runners at the elite level. PMID- 10862540 TI - Fascicle length of leg muscles is greater in sprinters than distance runners. AB - PURPOSE: The purpose of this study was to compare architectural characteristics of leg muscles of sprinters and distance runners. METHODS: Skeletal muscle architectural characteristics were studied in 23 elite male 100-m sprinters (SPR, 10.0-10.9 s for 100 m), 24 elite male distance runners (DR, 13.5-14.5 min for 5000 m), and 24 untrained male controls. Fascicle pennation angle and isolated muscle thickness of the vastus lateralis and gastrocnemius medialis and lateralis muscles were measured in vivo by ultrasound, and fascicle length was estimated. RESULTS: Standing height and upper and lower limb lengths were similar among the groups. Body weight was significantly greater in SPR than in either DR or controls, which were similar. Muscle thickness of the vastus lateralis and gastrocnemius medialis and lateralis muscles was significantly greater in SPR than in either DR or controls, which were similar. In all muscles, pennation angle was similar between SPR and controls, but less than DR. Fascicle length of the vastus lateralis muscle (absolute and relative to limb length) was greatest in SPR and least in DR with control values being between the athlete groups. Fascicle length of the gastrocnemius medialis muscle (absolute and relative to limb length) was greater in SPR than in either DR or controls, which were similar. Fascicle length of the gastrocnemius lateralis muscle (absolute and relative to limb length) was significantly greater in SPR than DR. Absolute fascicle length in gastrocnemius lateralis muscle was similar between DR and controls; however, relative to limb length DR was significantly less. CONCLUSION: Greater fascicle length and lesser pennation angle observed in leg muscles of SPR, compared with DR, would appear to favor shortening velocity as required for greater running speed. PMID- 10862542 TI - Maximal lactate-steady-state independent of performance. AB - PURPOSE: The maximal lactate steady state (MLSS) corresponds to the highest workload that can be maintained over time without a continual blood lactate accumulation. MLSS and MLSS intensity have been speculated to depend on performance. Experimental proof of this hypothesis is missing. METHODS: 33 male subjects (age: 23.7 +/- 5.5 yr, height: 181.2 +/- 5.3 cm, body mass: 73.4 +/- 6.4 kg) performed an exhausting incremental load test to measure peak workload and three to six 30-min constant load tests on a cycle ergometer to determine MLSS. RESULTS: MLSS (4.9 +/- 1.4 mmol x L(-1)) was independent of MLSS workload (3.4 +/ 0.6 W x kg(-1)) and peak workload (4.8 +/- 0.6 W x kg(-1)). MLSS intensity (71.1 +/- 6.7%) did not correlate with peak workload or MLSS (P > 0.05). A positive correlation was found between peak workload and MLSS workload (r = 0.82, P < 0.001). CONCLUSIONS: MLSS and MLSS intensity are independent of performance but subjects with higher maximum performance have higher MLSS workloads. The combination of various fitness related effects on both, the production and the disappearance of lactate during exercise, may explain that different MLSS workloads coincide with similar levels of MLSS and MLSS intensity. PMID- 10862543 TI - Longitudinal changes in young people's short-term power output. AB - PURPOSE: The influences of age, body size, skin-fold thickness, gender, and maturation on the short-term power output of young people were examined using multilevel modelling. METHODS: Subjects were 97 boys and 100 girls, aged 12.2 +/- 0.4 yr at the onset of the study. Sexual maturity was classified according to Tanner's indices of pubic hair. Peak power (PP) and mean power (MP) were determined on two occasions 1 yr apart using the Wingate Anaerobic Test (WAnT). The data were analyzed using multilevel regression modelling. RESULTS: Initial models identified body mass and stature as significant explanatory variables with an additional positive effect of age, which was smaller for girls' MP. A significant gender difference was apparent for both power indices with girls achieving lower values than boys. A significant incremental effect of later maturity (stages 4 and 5 for pubic hair development) was identified for MP only. Subsequent incorporation of sum of two skin-fold thicknesses into the model yielded significant negative parameter estimates for PP and MP and negated both the stature effects and the maturation influence upon MP. CONCLUSION: There are gender differences in the longitudinal growth of performance on the WanT. Regardless of gender differences, body mass and skin-fold thicknesses appear to be the best anthropometric predictors of WAnT determined PP and MP in young people. PMID- 10862544 TI - An integrated biomechanical analysis of high speed incline and level treadmill running. AB - PURPOSE: Recent sprint training regimens have used high-speed incline treadmill running to provide enhanced loading of muscles responsible for increasing forward running speed. The goal of this study was to document the joint kinematics, EMG, and swing-phase kinetics of incline treadmill running at 4.5 m x s(-1) with a 30% grade, and compare these data to that of level running under similar conditions. METHODS: Sagittal plane video (200 Hz) and EMG from eight lower extremity muscles were recorded during each of three locomotion conditions: incline running at 4.5 m x s(-1) and 30% grade (INC), level running at 4.5 m x s(-1) (LSS), and level running at the same stride frequency as INC (LSSF). A rigid body model was used to estimate net muscle power and work values at the hip, knee, and ankle during swing. Timing and amplitude of EMG signals for each muscle relative to footstrike were compared between conditions. RESULTS: Stride frequency and percentage of stride spent in stance were significantly higher during INC (1.78 Hz; 32.8%) than in the LSS (1.39 Hz; 28.8%) condition. Stride frequency played an important role, as most measures were more similar between INC and LSSF. Extensor range of motion of all joints during push-off was higher for INC. During INC, average EMG amplitude of the gastrocnemius, soleus, rectus femoris, vastus lateralis, and gluteus maximus were higher during stance, whereas the hamstrings activity amplitudes were lower. Average power and energy generated during hip flexion and extension in the swing phase were greatest during INC. CONCLUSIONS: These data suggest that compared with LSSF and LSS, INC provides enhanced muscular loading of key mono- and bi-articular muscles during both swing and stance phases. PMID- 10862545 TI - The influence of walking speed on dynamic loading on the human musculoskeletal system. AB - PURPOSE: A study was conducted to investigate the effects of walking speed on the magnitude of the heel strike initiated shock waves that propagate throughout the human musculoskeletal system. METHODS: Subjects walking on a treadmill at various speeds were used to acquire the experimental data regarding the heel strike initiated shock waves. Fifteen young healthy men participated in this study. Each one walked on the treadmill at five various speeds, and their associated cadences, while the heel strike induced shock waves were recorded at the tibial tuberosity for each speed during a 16-s period at a sampling rate of 1 kHz. RESULTS: The obtained data reveal a significant increase in the dynamic loading experienced by the human musculoskeletal system with an increase in the walking speed. CONCLUSION: The analysis of the recorded acceleration suggests that an increase in walking speed contributes to an increase in dynamic loading on the human musculoskeletal system. Further, the evidence indicates that dynamic loading increases with the increase in speed at five times the rate of the ground reaction force increase. PMID- 10862546 TI - Passive energy return after repeated stretches of the hamstring muscle-tendon unit. AB - It has been shown that five repetitive static stretches of human hamstring muscle, each lasting 90 s and separated by 30 s, altered the passive properties on a short-term basis. However, a total of 7.5 min (5 x 90 s) of stretching for a single muscle group may be an unrealistic stretching program. PURPOSE: The present investigation examined whether three repeated 45-s static stretches had a measurable effect on the passive properties of the hamstring muscle-tendon unit, in vivo. METHODS: Resistance to stretch was defined as the passive moment (Nm) offered by the hamstring muscle group during passive knee extension using a KinCom dynamometer as previously described (Kinetic Communicator, Chattecx Corp., Chattanooga, TN). The static stretch exercise was administered to the left lower extremity of all subjects and consisted of a dynamic phase of passive knee extension to a predetermined final joint angle followed by a 45-s static phase. The procedure was repeated for a total of three 45-s static stretches with 30-s rest period between stretches. RESULTS: There was a significant decline in resistance over the 45-s the static phase in stretch 1 (20 +/- 3%) and stretch 3 (18 +/- 3%), P < 0.001. Further, the absolute or relative decline in resistance over time stretch 1 and 3 were equivalent. The mean resistance in stretch 1 and 3, expressed as the log(e) of time, yielded an equal and highly linear relationship (r2 = 0.96 +/- 0.01); the slope and intercept did not differ. In the dynamic phase of the stretch, the energy of stretch 1 and 3 were similar. CONCLUSIONS: These data suggest that the static stretching protocol used in the present study had no short-term effect on the viscoelastic properties of human hamstring muscle group. PMID- 10862547 TI - Maximal aerobic power, lactate threshold, and running performance in master athletes. AB - PURPOSE: This study sought to determine how lactate threshold (LT) is related to running performance in older male and female runners, if LT changes significantly with age, and if gender alters the relationship between LT and performance in older runners. METHODS: Subjects were 168 master runners (111 men, 57 women) selected from a longitudinal study, who ran at least 10 miles x wk(-1) for 5 yr or more. VO2max was measured on a treadmill and body composition by hydrostatic weighing. Blood samples taken each minute of exercise were analyzed for lactate concentration and LT determined as the breakpoint in lactate accumulation. Performance times and training histories were self-reported by questionnaire. RESULTS: Men had significantly greater body mass, fat-free mass (FFM), and VO2max (L x min(-1); mL x kg(-1) x min(-1)) than women. FFM and VO2max (L x min(-1); mL x kg(-1) x min(-1)) declined with age in both men and women. Running performance was significantly different between men and women and declined with age in both. LT (L x min(-1); mL x kg(-1) x min(-1)) was significantly different between men and women, and declined significantly with age in men, whereas LT (%VO2max) did not differ between men and women and increased significantly with age in both. VO2max (mL x kg(-1) x min(-1)) was the most significant predictor of performance in both men and women, whereas LT (L x min(-1)) added to the prediction of 5-km and 10-km performance in women. CONCLUSION: The results of this study demonstrate that VO2max (mL x kg(-1) x min(-1)) is a better predictor of performance than LT in older male and female runners. Additionally, LT as a percentage of VO2max increases significantly with age. PMID- 10862548 TI - Physiological loads in the team technical and free routines of synchronized swimmers. AB - PURPOSE: The purpose of the present study was to examine the physiological loads on synchronized swimmers during team technical and free routines by measuring their blood lactate concentrations and percentage of peak blood lactate concentration. METHODS: Four trained college female synchronized swimmers participated as the subjects. The blood lactate concentration was measured in the first and middle periods and after the team technical and free routines. Peak blood lactate concentration was measured after maximum exertion in 100-m freestyle swimming. RESULTS: Average values and SD of blood lactate concentration and the percentage of peak blood lactate concentration after the team technical and free routines were 4.7 +/- 1.1 mmol x L(-1), 46.2 +/- 11.0% and 4.3 +/- 1.1 mmol x L(-1), 42.8 +/- 11.5%, respectively. The blood lactate concentration and percentage of peak blood lactate concentration after the team technical routine were significantly higher than those in the first period, and the blood lactate concentration after the team free routine was significantly higher than in the middle period. CONCLUSIONS: The blood lactate concentration of synchronized swimmers during the team technical and free routines in the present study tended to increase with the performance time. Thus, the predominant sources of energy may be phosphocreatine stores and aerobic metabolism during these routines, although glycolysis may also play an important role in relation to the energy requirements in the final period. PMID- 10862549 TI - The influence of direct supervision of resistance training on strength performance. AB - PURPOSE: The purpose of this study was to compare changes in maximal strength, power, and muscular endurance after 12 wk of periodized heavy-resistance training directly supervised by a personal trainer (SUP) versus unsupervised training (UNSUP). METHODS: Twenty moderately trained men aged 24.6 +/- 1.0 yr (mean +/- SE) were randomly assigned to either the SUP group (N = 10) or the UNSUP group (N = 8). Both groups performed identical linear periodized resistance training programs consisting of preparatory (10-12 repetitions maximum (RM)), hypertrophy (8 to 10-RM), strength (5 to 8-RM), and peaking phases (3 to 6-RM) using free weight and variable-resistance machine exercises. Subjects were tested for maximal squat and bench press strength (1-RM), squat jump power output, bench press muscular endurance, and body composition at week 0 and after 12 wk of training. RESULTS: Mean training loads (kg per set) per week were significantly (P < 0.05) greater in the SUP group than the UNSUP group at weeks 7 through 11 for the squat, and weeks 3 and 7 through 12 for the bench press exercises. The rates of increase (slope) of squat and bench press kg per set were significantly greater in the SUP group. Maximal squat and bench press strength were significantly greater at week 12 in the SUP group. Squat and bench press 1-RM, and mean and peak power output increased significantly after training in both groups. Relative local muscular endurance (80% of 1-RM) was not compromised in either group despite significantly greater loads utilized in bench press muscular endurance testing after training. Body mass, fat mass, and fat-free mass increased significantly after training in the SUP group. CONCLUSION: Directly supervised, heavy-resistance training in moderately trained men resulted in a greater rate of training load increase and magnitude which resulted in greater maximal strength gains compared with unsupervised training. PMID- 10862550 TI - Altered ankle joint proprioception in subjects suffering recurrent ankle sprains. PMID- 10862551 TI - Altered ankle joint proprioception in subjects suffering recurrent ankle sprains. PMID- 10862552 TI - A synopsis of the Third International Federation of Societies of Toxicologic Pathologists conference. PMID- 10862553 TI - DNA repair: kinetics and thresholds. AB - DNA damage is a critical factor in the initiation of chemically induced toxicities (including cancer), and the repair of this damage represents the cell's first line of defense against the deleterious effects of these agents. The various mechanisms of DNA repair are reviewed briefly and the actions of the DNA repair protein O6-alkylguanine DNA alkyltransferase (ATase) are used to illustrate how DNA repair can protect cells against alkylating agent-induced toxicities, mutagenesis, clastogenesis, and carcinogenesis. The effectiveness of this repair protein can be measured based on its ability to deplete levels of its promutagenic substrate O6-methylguanine (O6-meG) in the DNA of cells. These studies reveal that the repair of O6-meG from DNA occurs heterogeneously, both intra- and intercellularly. Even in cells that repair O6-meG hyperefficiently, certain regions of chromatin DNA are repaired with difficulty, and in other regions they are not repaired at all; most likely this lack of repair is a result of the location of the lesion in the DNA sequence. When individual cells are compared within a tissue, some cells are clearly repair deficient, because the O6 meG can persist in DNA for many weeks, whereas in other cells, it is removed within a matter of hours. The role of these repair-deficient cells as targets for alkylating agent induced carcinogenesis is considered. The mechanisms of the homeostatic control of DNA repair function in mammalian cells are not yet well understood. Because there are now indications of the mechanisms by which the level of DNA damage may be sensed (and so influence the activity of the ATase repair protein), this is an important area for future study. PMID- 10862554 TI - Metabolic detoxification: implications for thresholds. AB - The fact that chemical carcinogenesis involves single, isolated, essentially irreversible molecular events as discrete steps, several of which must occur in a row to finally culminate in the development of a malignancy, rather suggests that an absolute threshold for chemical carcinogens may not exist. However, practical thresholds may exist due to saturable pathways involved in the metabolic processing, especially in the metabolic inactivation, of such compounds. An important example for such a pathway is the enzymatic hydrolysis of epoxides via epoxide hydrolases, a group of enzymes for which the catalytic mechanism has recently been established. These enzymes convert their substrates via the intermediate formation of a covalent enzyme-substrate complex. Interestingly, the formation of the intermediate proceeds faster by orders of magnitude than the subsequent hydrolysis, ie, the formation of the terminal product. Under normal circumstances, this does not pose a problem, since the microsomal epoxide hydrolase (mEH), the epoxide hydrolases with the best documented importance in the metabolism of carcinogens, is highly abundant in the liver, the organ with the highest capacity to metabolically generate epoxides. Computer simulation provides evidence that the high amount of mEH enzyme is favorable for the control of the steady-state level of a substrate epoxide and can keep it extremely low. However, once the mEH is titrated out under conditions of extraordinarily high epoxide concentration, the epoxide steady-state level steeply rises, leading to a sudden burst of the genotoxic effect of the noxious agent. This prediction of the computer simulation is nicely supported by experimental work. V79 Chinese hamster cells that we have genetically engineered to express human mEH at about the same level as that observed in human liver are completely protected from any measurable genotoxic effect of the model compound styrene oxide (STO) up to a dose of 100 microM in the cell culture medium (toxicokinetic threshold). In V79 cells that do not express mEH, STO leads to the formation of DNA strand breaks in a dose-dependent manner with no toxicokinetic threshold observable. Above 100 microM, the genotoxic effect of STO in the mEH-expressing cell line parallels the one in the parental cell line. Thus, the saturable protection from STO-induced strand breaks by mEH represents a typical example of a practical threshold. However, it must be pointed out that even in the presence of protective amounts of mEH, a minute but definite level of STO is present that does not contribute sufficiently to the strand break formation to overcome the background noise of the detection procedure. As pointed out above, absolute thresholds probably do not exist in chemical carcinogenesis. PMID- 10862555 TI - Mechanistic basis for nonlinearities and thresholds in rat liver carcinogenesis by the DNA-reactive carcinogens 2-acetylaminofluorene and diethylnitrosamine. AB - To explore differences in mechanisms of carcinogenicity at low and high exposures, we have conducted a series of exposure-response studies of hepatocarcinogenesis in rats using 2 well-studied DNA-reactive carcinogens, 2 acetylaminofluorene and diethylnitrosamine. In these studies, we have used intraperitoneal injection or intragastric instillation to deliver exact doses during an initiation segment followed by phenobarbital as a liver tumor promoter to enhance manifestation of initiation. This protocol results in carcinogenicity comparable to that produced by lifetime exposure to the carcinogens. Our findings in these experiments provide evidence for the following: (a) formation of DNA adducts can be nonlinear, with a plateau at higher exposures; (b) cytotoxicity shows no-effect levels and is related to exposure; (c) compensatory hepatocyte proliferation shows no-effect levels and can be supralinear at high exposures; (d) formation of preneoplastic hepatocellular altered foci can show no-effect levels and appears supralinear at high exposures; (e) no-effect levels can exist for tumor development, and the exposure response can be supralinear. We interpret these findings to reflect thresholds for hepatocellular initiating effects of these carcinogens and exaggerated responses at high exposures attributable to cytotoxicity and compensatory hepatocyte proliferation. Such enhanced proliferation of hepatocytes harboring DNA damage likely results in an exaggerated yield of mutations in critical genes, leading to supralinear initiation of carcinogenesis. Thus, mechanisms differ between low and high exposures. Based on these observations, we suggest that linear extrapolation from high toxic exposures to postulated low-exposure effects of DNA-reactive carcinogens can yield overestimates. Such extrapolation must be supported by mechanistic information. The finding of no-effect levels provides a basis for understanding why low-level environmental exposures of humans to even DNA reactive carcinogens may convey no cancer risk. PMID- 10862556 TI - Industry viewpoint on thresholds for genotoxic carcinogens. AB - Modern chemical control of pests has contributed to a dramatic improvement in public welfare since its introduction 50 years ago. Millions of lives have been saved through the control of disease vectors, and millions more have been improved by the use of chemicals to produce an inexpensive and abundant food supply. Hundreds of pesticidally active ingredients are in commercial use today, and among these are found genotoxic and nongenotoxic carcinogens. In the United States, the Environmental Protection Agency regulates carcinogens using threshold and nonthreshold approaches depending upon the outcome of a weight-of-evidence determination. More than one-half of all pesticides with some evidence of carcinogenic potential are regulated by the nonthreshold approach. The limitations on product use imposed by this approach have restricted the number of products available to growers and to the public. This restriction has had a direct impact on industry with respect to commercial success and financial returns on investment as well as an indirect impact on the industry's ability to fund the discovery and development of new compounds. This paper explores the question of how well regulation by the nonthreshold approach has achieved the goal of protecting public health, whether it does this better than the alternative use of the threshold approach, and whether the incremental protection it affords is a meaningful public benefit that justifies the aforementioned impacts on industry. PMID- 10862557 TI - DNA adducts: endogenous and induced. AB - Human exposure to DNA damaging agents can arise from exogenous sources or endogenous processes that occur normally or in pathological states. DNA isolated from human tissues, obtained from the very young to the old, contains detectable amounts of a number of different types of DNA adducts that reflect exposure to both known carcinogens and as yet unidentified genotoxic agents. The levels of DNA damage observed in human studies as a result of exogenous exposures (noniatrogenic) is of the order of 1 adduct per 10(7)-10(9) normal DNA bases, whereas that arising from endogenous exposures may potentially be several orders of magnitude higher. Large interindividual variations in DNA adduct levels have been reported, and these are probably the result of host and environmental factors, although variation in analytical and sampling procedures may also play a role. It is important to recognize that the presence of DNA adducts in a tissue does not necessarily indicate a specific tumorigenic risk for that tissue, as other factors downstream of DNA adduct formation (including DNA repair and cell proliferation) play an important role in determining overall risk. PMID- 10862559 TI - Endocrine disruptors: overview and a pathologist's perspective. PMID- 10862558 TI - An introduction to endocrine modulators. PMID- 10862560 TI - Endocrine modulators in the food chain and environment. AB - Recently, considerable attention has been focused on certain environmental contaminants--"endocrine disruptors"--of industrial origin that may mimic the action of sex hormones. Natural compounds and their effects on other types of hormonal activity (eg, on adrenal or thyroid function) have for some reason not provoked similar attention. As exemplified by tributyltin and certain bioaccumulating chlorinated compounds, available evidence indicates that "endocrine disruption" caused by xenobiotics is primarily an ecotoxicologic problem. In mammals, certain phenylmethyl-substituted siloxanes have been found to be by far the most potent endocrine disrupters among various synthetic xenobiotics. On the other hand, it has not been possible to scientifically substantiate either certain alarming reports of powerful synergistic effects between chlorinated pesticides or the alleged adverse effects on the male reproductive tract in rodents (induced by alkylphenols and plasticizers at extremely low exposures). Whereas there is compelling evidence that estrogens in certain foods and herbal medicines can induce hormonal changes in women as well as overt toxicity in men, existing data are insufficient to support a causal relationship between exposure of the general human population to nonpharmaceutical industrial chemicals and adverse effects operating via the endocrine system. Moreover, in terms of magnitude and extent, all such exposures to so-called endocrine disruptors are dwarfed by the extensive use of oral contraceptives and estrogens for treatment of menopausal and postmenopausal disorders. Also, the exposure to hormonally active xenobiotics is virtually insignificant when compared with the intake of the phytoestrogens that are present in food and beverages, and it is even more insignificant when compared with certain herbal potions used in "alternative medicine." Furthermore, while there has been much concern about negligible exposures to xenobiotics with weak hormonelike activities, the potent endocrine disruptor licorice is freely given to children. Long-term exposure to this substance induces severe toxic symptoms of mineral corticoid hormone imbalance. Although exposures to xenobiotics and many natural compounds occur by identical routes of administration and may contribute to the same toxicological end point, they are, regrettably, judged by completely different standards. As is the case with all other chemicals, rational risk assessment and risk management of man-made and natural endocrine modulators must be based on the mode of action and dose-response relationships. Such end points as the induction of reproductive developmental effects, cancer, etc, relating to actual exposures must also be taken into consideration. PMID- 10862561 TI - Validation of in vitro and in vivo methods for assessing endocrine disrupting chemicals. AB - The concepts that require validation in terms of the subject of endocrine disruption are listed and discussed. The main mechanisms by which endocrine disruption can occur are identified, and the assays required for the detection of adverse endocrine disruption toxicities associated with these mechanisms are discussed. The process of assay validation is considered. The validation of structure-activity relationships, the need for reference chemicals, and the problems recently encountered when attempting to reproduce endocrine disruption data are also explored. The most important conclusions derived from this analysis are that given the immature state of research into endocrine disruption toxicity, testing strategies and the types of assay employed should be kept under constant review; inevitably researchers need to accept the fact that future revision of each assay will be required. Second, given the current absence of any chemical that is universally accepted to be devoid of endocrine toxicity, assay specificity will be difficult to assess, and that imposes the need for alternative objective criteria for assessing the value of individual assays. PMID- 10862562 TI - Endocrine modulators: risk characterization and assessment. AB - Over the last several years, information has been accumulating that suggests that adverse effects are being induced in certain wildlife species, and perhaps also in humans, as a consequence of exposure to man-made chemicals that have been released into the environment. Many of these effects have been attributed to interactions with various hormone systems in endocrine tissues. Most often the effects observed have been effects on reproduction and development, although there are also (often conflicting) data regarding an association between exposure and certain kinds of cancer, particularly cancers of reproductive tissues such as breast and testis; effects on the immune system have also been noted. The substances to which these attributes have been ascribed have come to be known as "endocrine modulators" or "endocrine disruptors." The full nature and scope of the "problem" of endocrine modulators/disruptors is currently a matter of great debate, both within and outside of the scientific community. Regulatory authorities around the world are being asked what their position is on this issue and what, if any, regulatory strategies they are developing to address the problem. In many cases, because of the nature of the legislation under which governments manage chemicals, regulatory decisions must be informed by risk assessment. This presentation will describe the general approach to the risk assessment of endocrine modulators/disruptors as practiced by the US government, with particular focus on the current practices/policies of the US Environmental Protection Agency. PMID- 10862563 TI - Cell death and cell proliferation in the control of normal and neoplastic tissue growth. AB - The development of reliable methodology for the assessment of rates of cell replication and cell death has enabled the study of how these 2 fundamentally opposed processes work to form and maintain tissue and to remodel tissue following diseases resulting in cell loss. The balance between these 2 processes and the consequences of an imbalance are fundamental to a clearer understanding of how hyperplasia and neoplasia develop in tissues under the influence of chemicals and drugs. An understanding of the changes that occur in target organs and tissues following chemical or drug exposure has enabled a better understanding of the mechanism by which these chemicals are able to induce cancer after prolonged exposure. Studies of the control of cell replication and the changes that occur following drug exposure have defined 2 types of response, 1 in which the cell replicative response is sustained and the other in which the cell replicative response is transient and occurs during the first few days of exposure. Although regulatory and scientific opinion appears ready to accept sustained cell replicative processes as an increased risk factor in the development of cancer, the role played by transient increases in cell replication remains unclear. Concurrent events in target organs following treatment with chemicals that induce transient increases in cell replication have revealed that the rates of apoptosis are suppressed at the same time as the cell replication levels are induced. Additional evidence suggests that growth and antigrowth factors are central in controlling these responses. Escape from the regulatory action of these factors is postulated to be one of the ways in which nongenotoxic carcinogenic chemicals, such as the peroxisome proliferators and sodium phenobarbitone, may induce cancer, with apoptosis playing a key role in the process. PMID- 10862564 TI - The use of genetically altered animals in toxicology. PMID- 10862565 TI - The use of genetically modified animals in carcinogenicity bioassays. PMID- 10862566 TI - Histopathologic approaches to detect changes indicative of immunotoxicity. AB - Toxicologic pathology is crucial in the identification and characterization of health effects following exposure to xenobiotics, mainly in toxicity experiments in rodents. Regarding regulatory toxicology, histopathology of lymphoid organs and tissues is a cornerstone in the identification of immunotoxic compounds. A 2 tier testing system is usually employed in which the first tier is a general screen for (immuno)toxicity and the second tier consists of specific immune function studies, including host resistance tests or mechanistic studies. The role attributed to histopathology of lymphoid organs in the updated Organisation for Economic Cooperation and Development and Food and Drug Administration guidelines requires improvement and standardization of the histopathology procedures. Optimalization and standardization was started in an international collaborative immunotoxicity study (ICICIS). However, several problems were left unaddressed, mostly because of the few compounds tested in this study. Based on the results of the ICICIS study and the morphologic changes induced by immunotoxic/immunomodulatory compounds observed in other investigations, suggestions are given to further improve the identification and (semi)quantification of histopathologic changes in lymphoid organs and tissues. PMID- 10862567 TI - US FDA "Redbook II" immunotoxicity testing guidelines and research in immunotoxicity evaluations of food chemicals and new food proteins. AB - The rapid advances in the field of immunology and an understanding of the potential adverse effects of xenobiotics on the immune system have resulted in the development of a discipline in toxicology now referred to as immunotoxicology. This discipline has evolved steadily over the last 2 decades as a result of research in the national and international communities. Various US, European, and Japanese regulatory agencies have recognized a need to promulgate testing guidelines for immunotoxicity in support of the approval process involving toxicological testing. The US Food and Drug Administration "Redbook II" guidelines and some of the research conducted in support of the concepts and testing strategies are presented here. Concerns raised with regard to these guidelines are included, as are on-going initiatives in development of experimental approaches for assessing allergic potential and/or hypersensitivity responses to new foods and food constituents. PMID- 10862568 TI - Responses of the immune system to injury. AB - Three categories of immunotoxic effects are identified: direct immunotoxicity, hypersensitivity, and autoimmunity. Direct immunotoxicity consists of immunosuppression and immunostimulation. Total abrogation of the immune response (immunosuppression) results in more frequent, severe, and often atypical and relapsing infections and lymphomas. Immunostimulation is associated with febrile reactions, the induction/facilitation of autoimmune diseases and allergic reactions to unrelated allergens, and impaired hepatic drug biotransformation. Hypersensitivity is manifested by a variety of symptoms involving either antigen specific or non-antigen-specific humoral and cellular adverse responses. Autoimmune reactions are divided into organ-specific and systemic reactions. Because of the involvement of many redundant mechanisms, it is difficult to predict responses of the immune system to a given immunotoxic injury. In laboratory animals, histologic but also functional changes are necessary to show evidence of and to predict such adverse responses. PMID- 10862569 TI - Use of transgenic animals for carcinogenicity testing: considerations and implications for risk assessment. AB - Advances in genetic engineering have created opportunities for improved understanding of the molecular basis of carcinogenesis. Through selective introduction, activation, and inactivation of specific genes, investigators can produce mice of unique genotypes and phenotypes that afford insights into the events and mechanisms responsible for tumor formation. It has been suggested that such animals might be used for routine testing of chemicals to determine their carcinogenic potential because the animals may be mechanistically relevant for understanding and predicting the human response to exposure to the chemical being tested. Before transgenic and knockout mice can be used as an adjunct or alternative to the conventional 2-year rodent bioassay, information related to the animal line to be used, study design, and data analysis and interpretation must be carefully considered. Here, we identify and review such information relative to Tg.AC and rasH2 transgenic mice and p53+/- and XPA-/- knockout mice, all of which have been proposed for use in chemical carcinogenicity testing. In addition, the implications of findings of tumors in transgenic and knockout animals when exposed to chemicals is discussed in the context of human health risk assessment. PMID- 10862570 TI - Brugada syndrome: an electrocardiographic diagnosis not to be missed. PMID- 10862571 TI - Stamps in Cardiology. Hypertension. PMID- 10862572 TI - Role of transoesophageal echocardiography in infective endocarditis. PMID- 10862573 TI - Fair comparison of mortality data following cardiac surgery. PMID- 10862574 TI - Cardiac surgical mortality: the tip of the quality assurance iceberg. PMID- 10862575 TI - Interleukin 6: a message from the heart. PMID- 10862576 TI - Images in cardiology. Saddle pulmonary embolism: diagnosis by spiral computed tomography. PMID- 10862577 TI - Treatment options for coarctation of the aorta. PMID- 10862578 TI - Images in cardiology. Mitral regurgitation secondary to ruptured papillary muscle. PMID- 10862580 TI - Images in cardiology: Echohistological correlation in Candida albicans endocarditis. PMID- 10862579 TI - Antiarrhythmics--from cell to clinic: past, present, and future. PMID- 10862581 TI - Infective endocarditis: clinical spectrum, presentation and outcome. An analysis of 212 cases 1980-1995. AB - OBJECTIVE: To evaluate recent changes in the spectrum and clinical presentation of infective endocarditis and to determine predictors of outcome. DESIGN: A retrospective case study. METHODS: Demographic, clinical, and echocardiographic characteristics were examined in 212 patients who fulfilled the Duke criteria for infective endocarditis between January 1980 and December 1995 to assess changes in clinical presentation and survival. RESULTS: Clinical presentation and course did not change significantly during the study period despite the concurrent introduction of new diagnostic tools (for example, transoesophageal echocardiography). In-hospital mortality was 15% and remained unchanged. Neurological symptoms on admission, arthralgia, and weight loss were all independent risk factors for adverse outcome (odds ratios 26.1, 6.2, and 4.2, respectively). Age, prosthetic valve disease, previous antibiotic treatment, renal insufficiency, surgical treatment, and the type of valve involved were not predictive of mortality. In contrast to all other major reports, Streptococcus viridans was the most common causative organism in intravenous drug users (52%). CONCLUSIONS: Despite the introduction of new diagnostic tools, the course of infective endocarditis has remained unchanged over a period of 16 years. Evidence of early dissemination of the disease to other sites was associated with adverse outcome. Even in elderly patients, early aggressive treatment seems to be effective. PMID- 10862582 TI - Images in cardiology. Coronary artery calcification. PMID- 10862583 TI - Prevalence of the Brugada sign in idiopathic ventricular fibrillation and healthy controls. AB - OBJECTIVE: To determine the prevalence of the Brugada sign (right bundle branch block with ST elevation in V1-V3) in idiopathic ventricular fibrillation and in an age matched healthy population. DESIGN: ECGs from 39 consecutive patients with idiopathic ventricular fibrillation and 592 healthy controls were reviewed. They were classified as definite, questionable, and no Brugada sign (according to predetermined criteria) by four investigators blinded to the subjects' status. RESULTS: Eight patients (21%) with idiopathic ventricular fibrillation but none of the 592 controls had a definite Brugada sign (p < 0.005). Thus the estimated 95% confidence limits for the prevalence of a definite Brugada sign among healthy controls was less than 0.5%. A questionable Brugada sign was seen in two patients with idiopathic ventricular fibrillation (5%) but also in five controls (1%) (p < 0.05). Normal ECGs were found following resuscitation and during long term follow up in 31 patients with idiopathic ventricular fibrillation (79%). Patients with idiopathic ventricular fibrillation and a normal ECG and those with the Brugada syndrome were of similar age and had similar spontaneous and inducible arrhythmias. However, the two groups differed in terms of sex, family history, and the incidence of sleep related ventricular fibrillation. CONCLUSIONS: A definite Brugada sign is a specific marker of arrhythmic risk. However, less than obvious ECG abnormalities have little diagnostic value, as a "questionable" Brugada sign was observed in 1% of healthy controls. In this series of consecutive patients with idiopathic ventricular fibrillation, most had normal ECGs. PMID- 10862584 TI - Images in cardiolgy: Apparent induction of ventricular tachycardia after "appropriate pacing" by an implantable dual chamber defibrillator: confusing ICD electrograms. PMID- 10862585 TI - Randomised double blind trial of oral versus intravenous flecainide for the cardioversion of acute atrial fibrillation. AB - OBJECTIVE: To investigate whether an oral loading dose of flecainide is as safe and effective as intravenous flecainide for the cardioversion of acute atrial fibrillation. DESIGN: Prospective, randomised, double blind, double placebo study. SETTING: Cardiac care unit of a large district general hospital in the UK. PATIENTS AND METHODS: 79 patients presenting with symptomatic acute atrial fibrillation: patients were given either intravenous flecainide (n = 39) or a solution of oral flecainide (n = 40), with appropriate placebos. All patients were heparinised during the study. PRIMARY OUTCOME MEASURES: Safety; mean time to cardioversion; proportion of patients restored to sinus rhythm at two hours and eight hours after treatment. Analysis was by intention to treat. RESULTS: There were no differences in baseline characteristics between the oral and intravenous groups. Both forms of flecainide were well tolerated, with no adverse clinical events during the study. The mean time to cardioversion was 110 minutes in the oral group and 52 minutes in the intravenous group (p = 0.002). Two hours after treatment, 27 of the 40 patients in the oral group (68%) and 25 of the 39 in the intravenous group (64%) had reverted to sinus rhythm (p = 0.74). Eight hours after treatment, 30 patients in the oral group (75%) and 28 in the intravenous group (72%) had reverted to sinus rhythm (p = 0.76). CONCLUSIONS: Intravenous flecainide restored sinus rhythm more rapidly than oral flecainide, but at two hours and eight hours after treatment there was no difference in the proportion of patients cardioverted by the two approaches. These results suggest a role for oral loading doses of flecainide in the treatment of acute or symptomatic paroxysmal atrial fibrillation. PMID- 10862586 TI - Influence of previous aspirin treatment and smoking on the electrocardiographic manifestations of injury in acute myocardial infarction. AB - OBJECTIVE: To examine demographic and clinical characteristics of patients with acute myocardial infarction in order to identify factors affecting the electrocardiographic evolution of injury. METHODS: Prospective cohort study of 1399 consecutive patients with a first myocardial infarction. Baseline clinical data associated with ST elevation and Q wave development were identified and 12 month survival was estimated. RESULTS: Smoking had complex effects on the evolution of injury, increasing the odds of ST elevation (odds ratio (OR) 1.61; 95% confidence interval (CI) 1.08 to 2.36), but reducing the odds of Q wave development (OR 0.69, 95% CI 0.49 to 0.96). The effects of previous aspirin treatment were more consistent with reductions in the odds of ST elevation (OR 0.57, 95% CI 0.35 to 0.94) and Q wave development (OR 0.53, 95% CI 0.34 to 0. 84). ST elevation and Q wave development were both associated with an adverse prognosis, with estimated 12 month survival rates of 80. 6% (95% CI 78.2% to 83.1%) and 80.0% (95% CI 77.5% to 82.5%), respectively, compared with 86.5% (95% CI 81.2% to 91.9%) and 89.9% (95% CI 86.2% to 93.7%) for patients without these ECG changes. CONCLUSIONS: The thrombogenicity of the blood may be a major determinant of infarct severity. Smoking increases thrombogenicity and the likelihood of ST elevation, but because coronary occlusion is relatively more thrombotic in smokers, responses to both endogenous and exogenous thrombolysis are better, reducing the risk of Q wave development. Previous aspirin treatment reduces thrombogenicity, protecting against ST elevation and Q wave development. PMID- 10862588 TI - Images in cardiology. Visualising fatty deposits in familial arrhythmogenic right ventricular cardiomyopathy by magnetic resonance imaging. PMID- 10862587 TI - Oxygen uptake versus exercise intensity: a new concept in assessing cardiovascular exercise function in patients with congenital heart disease. AB - OBJECTIVE: To assess the relation between exercise intensity and oxygen uptake during graded exercise in paediatric patients who underwent surgical repair of congenital heart disease, and to compare it with conventional measures of aerobic exercise function. DESIGN: Cross sectional study. Exercise testing was performed on a treadmill and gas exchange was measured on a breath by breath basis. PATIENTS: 29 patients who underwent an atrial switch operation for transposition of the great arteries (TGA) (mean (SD) age at testing 10.3 (2.5) years) and 30 patients who underwent total repair of tetralogy of Fallot (TF) (age 12.1 (3.3) years) performed graded exercise testing. Exercise responses were compared with data obtained in 24 normal controls (age 11.4 (2.6) years). RESULTS: The slope of oxygen uptake versus exercise intensity averaged 1.50 (0. 64) ml O(2)/min(2)/kg in the patients with TGA and 1.68 (0.75) ml O(2)/min(2)/kg after TF repair, both lower (p < 0.005) than in normal controls (2.42 (0.68) ml O(2)/min(2)/kg). The lower slope of oxygen uptake was correlated with a subnormal value for ventilatory anaerobic threshold, which averaged 78.0 (13.3)% of normal in TGA and 85.1 (10.6)% in TF. This was associated with a steeper slope (p = 0.001) of carbon dioxide output versus oxygen uptake above the ventilatory anaerobic threshold in TGA (1.26 (0.20)) and TF (1.20 (0. 18)) compared with the normal controls (1.05 (0.13)), and also a steeper slope of ventilation versus carbon dioxide in TGA (47.0 (15. 4)) and TF (41.5 (13.7)) than in the controls (30.3 (8.5)). CONCLUSIONS: Calculation of the steepness of the slope of oxygen uptake versus exercise intensity is a valid measurement of oxygen flow to the exercising tissues, which may be limited in congenital heart disease. PMID- 10862589 TI - Haemodynamic correlates and prognostic significance of serum uric acid in adult patients with Eisenmenger syndrome. AB - OBJECTIVE: To assess haemodynamic correlates and prognostic significance of serum uric acid in adult patients with Eisenmenger syndrome. DESIGN: Retrospective observational study. SETTING: Tertiary referral centre. PATIENTS: 94 adult patients with Eisenmenger syndrome who were diagnosed between September 1982 and July 1998. MAIN OUTCOME MEASURES: Serum uric acid was measured in all patients, together with clinical and haemodynamic variables related to mortality. RESULTS: Serum uric acid was raised in patients with Eisenmenger syndrome compared with age and sex matched control subjects (7.0 v 4.7 mg/dl, p < 0.0001) and increased in proportion to the severity of New York Heart Association functional class. Serum uric acid was positively correlated with mean pulmonary arterial pressure (r = 0.30, p = 0.0052) and total pulmonary resistance index (r = 0.55, p < 0.0001), and negatively correlated with cardiac index (r = -0.50, p < 0.0001). During a mean follow up period of 97 months, 38 patients died of cardiopulmonary causes. Among various clinical, echocardiographic, and laboratory variables, serum uric acid remained predictive in multivariate analysis. Kaplan-Meier survival curves based on median serum uric acid showed that patients with high values had a significantly worse survival rate than those with low values (log lank test: p = 0.0014 in male patients, p = 0.0034 in female patients). CONCLUSIONS: Serum uric acid increases in proportion to haemodynamic severity in adult patients with Eisenmenger syndrome and is independently associated with long term mortality. PMID- 10862590 TI - Intracardiac pressures in the human fetus. AB - OBJECTIVE: To obtain normal values for intracardiac pressures in the human fetus. DESIGN: Intracardiac pressures were measured directly in the four chambers of the human fetal heart during clinically indicated invasive obstetric procedures. SETTING: Department of fetal medicine in a tertiary referral centre. PATIENTS: 39 fetuses between 16 and 29 weeks of gestation. RESULTS: The ventricular waveforms obtained were similar to those found in postnatal life. There was an increase in ventricular systolic and end diastolic pressures with advancing gestation. There was no difference between left and right ventricular pressures. Atrial pressures were equal and remained constant in the gestational age range studied. CONCLUSIONS: Fetal cardiovascular pressure measurements in the normal fetus assist in understanding the fetal circulation, and provide a basis for the assessment of cases of congenital heart disease that may be amenable to intrauterine treatment. PMID- 10862591 TI - Images in cardiology. Defective limb growth after retrograde balloon valvuloplasty. PMID- 10862592 TI - Formula and nomogram for the sphygmomanometric calculation of the mean arterial pressure. PMID- 10862593 TI - Stent treatment for coarctation of the aorta: intermediate term follow up and technical considerations. AB - OBJECTIVE: To report the initial and intermediate term results of stent implantation in children with coarctation of the aorta. PATIENTS AND DESIGN: 17 patients with coarctation of the aorta underwent stent implantation (median age 11 years, range 0.4-15 years); six were treated for isolated coarctation, nine for recurrent coarctation (five after surgical repair and four after balloon dilatation), and two for complex long segment coarctation. INTERVENTIONS: The procedure was guided by a second catheter placed transseptally in the left ventricle or the aorta proximal to the coarctation site, for angiographic and haemodynamic monitoring during the procedure. Twenty two stents were implanted in 17 patients. One of the patients with long segment coarctation received four stents and the other three. Palmaz 4014 stents were placed in 11 patients, Palmaz 308 in five, and Palmaz 154 in one. RESULTS: Immediately after stent implantation the peak systolic gradient (mean (SD)) fell from 50.0 (24.5) to 2.1 (2.4) mm Hg (p < 0.05). The diameter of the stenotic lesion increased from 5.1 (1.5) mm to 13.9 (2.4) mm (p < 0.05). There were no deaths or procedure related complications. At a median follow up of 33 months, no cases of recoarctation were identified, either clinically (0/17; 0%, 95% confidence interval (CI) 0% to 19%) or angiographically (0/13; 0%, 95% CI 0% to 25%). CONCLUSIONS: Stent implantation for the treatment of coarctation of the aorta appears to have very low morbidity and mortality, and reasonable intermediate term results. Long term freedom from recoarctation using this method remains to be determined in comparison with simple balloon dilatation. PMID- 10862594 TI - Images in cardiology. Adenosine testing for accessory pathways: not as simple as it seems. PMID- 10862595 TI - Limitations of the Parsonnet score for measuring risk stratified mortality in the north west of England. The North West Regional Cardiac Surgery Audit Steering Group. AB - OBJECTIVE: To study the use of the Parsonnet score to predict mortality following adult cardiac surgery. DESIGN: Prospective study. SETTING: All centres performing adult cardiac surgery in the north west of England. SUBJECTS: 8210 patients undergoing surgery between April 1997 and March 1999. MAIN OUTCOME MEASURES: Risk factors and in-hospital mortality were recorded according to agreed definitions. Ten per cent of cases from each centre were selected at random for validation. A Parsonnet score was derived for each patient and its predictive ability was studied. RESULTS: Data collection was complete. The operative mortality was 3.5% (95% confidence interval 3.1% to 3.9%), ranging from 2.7% to 3.8% across the centres. On validation, the incidence of discrepancies ranged from 0% to 13% for the different risk factors. The predictive ability of the Parsonnet score measured by area under the receiver operating characteristic curve was 0.74. The mean Parsonnet score for the region was 7.0, giving an observed to expected mortality ratio of 0.51 (range 0.4 to 0.64 across the centres). A new predictive model was derived from the data by multivariate analysis which includes nine objective risk factors, all with a significant association with mortality, which highlights some of the deficits of the Parsonnet score. CONCLUSIONS: Risk stratified mortality data were collected on 100% of patients undergoing adult cardiac surgery in two years within a defined geographical region and were used to set an audit standard. Problems with the Parsonnet score of subjectivity, inclusion of many items not associated with mortality, and the overprediction of mortality have been highlighted. PMID- 10862596 TI - Likely variations in perioperative mortality associated with cardiac surgery: when does high mortality reflect bad practice? AB - OBJECTIVE: Several methods exist for estimating the risk of perioperative mortality based on preoperative risk factors; graphical methods such as the variable life adjusted display (VLAD) can be used to examine how an individual surgeon's performance for a series of operations fares against what would be expected, given the case mix. This study aimed to devise a method for assessing the natural variation in outcome in order to assist with making judgements about individual performance, in particular whether seemingly poor performance could have occurred by chance. METHOD: The risk scoring system has been derived and validated locally for cardiac surgery. A method is described for calculating the probability that an observed number of deaths occurs within a sequence of operations if perioperative mortality is regarded as a chance event with an expected value derived from the risk score. To illustrate this method, nested prediction intervals are superimposed onto VLAD plots for a series of 393 isolated coronary artery bypass and isolated valve operations performed by a single surgeon. RESULTS: Using the locally derived risk score, the VLAD plot for the individual surgeon shows a net life gain of about 6 over the predicted number of survivors, which is observed to be within the 90% prediction interval. If the Parsonnet scoring system is used instead of the locally derived risk score, the net life gain is considerably overestimated. CONCLUSIONS: The nested prediction intervals are straightforward to generate and can be integrated into a visually informative display. As an indication of the inherent variability in outcome, they have a valuable role in the monitoring of surgical performance. PMID- 10862597 TI - Interleukin 6 expression in coronary circulation after coronary angioplasty as a risk factor for restenosis. AB - OBJECTIVE: To investigate changes in cytokine expression in the coronary circulation induced by percutaneous transluminal coronary angioplasty (PTCA). METHODS: The study involved 32 patients with ischaemic heart disease who underwent elective PTCA for isolated stenotic lesions of the left coronary artery. Ten patients had plain old balloon angioplasty, 10 had percutaneous transluminal rotational atherectomy, and 12 had stent implantation. Blood samples were drawn from the coronary sinus before and immediately after PTCA. Plasma concentrations of interleukin 6 (IL-6), platelet derived growth factor (PDGF), monocyte chemoattractant protein 1 (MCP-1), and macrophage coronary stimulating factor (M-CSF) were measured. The patients were scheduled for follow up angiography six months after PTCA. Late loss index was calculated using quantitative coronary angiography. RESULTS: IL-6 concentrations in coronary sinus blood increased immediately after PTCA (p < 0.001), but there was no change in PDGF, MCP-1, or M-CSF. There was a positive correlation between changes in IL-6 concentrations immediately after PTCA and late loss index six months after PTCA (r = 0.73, p < 0.001). IL-6 concentrations in coronary sinus blood were higher in patients with late restenosis than in those without restenosis (p < 0.001). CONCLUSIONS: PTCA induces IL-6 production in the coronary circulation. This may induce subsequent inflammatory responses in injured vessels and play an important role in late restenosis after PTCA. PMID- 10862598 TI - Decreased adrenomedullin production in the coronary circulation of patients with coronary artery disease. PMID- 10862599 TI - Coronary disease. Acute coronary syndromes: presentation--clinical spectrum and management. PMID- 10862600 TI - Coronary disease. Management of the post-myocardial infarction patient: rehabilitation and cardiac neurosis. PMID- 10862601 TI - Cardiomyopathy. Diagnosis and management of dilated cardiomyopathy. PMID- 10862602 TI - Capture and fusion beats during atrial fibrillation and ventricular tachycardia. AB - Two patients were presented, and two previously unreported observations were made. Patient 1, a 50 year old man with episodic palpitations and dizziness for 10 years, exhibited initiation of idiopathic ventricular tachycardia (VT) by atrial fibrillation (AF). Patient 2, a 43 year old woman with a structurally normal heart but recurrent palpitations for one year, demonstrated fusion and capture beats during simultaneous VT and AF. An explanation is given as to why the latter phenomenon is rarely observed. PMID- 10862603 TI - Coronary artery spasm induced by carotid sinus massage. AB - A 60 year old man with a history of frequent episodes of chest pain and dizziness was referred for evaluation of coronary artery disease. He had no significant coronary artery stenosis at baseline coronary angiography. A carotid sinus massage was performed for evaluation of carotid sinus hypersensitivity in the patient. Both heart rate and blood pressure decreased a little, and returned to baseline level immediately after carotid sinus massage. However, 2.5 minutes after carotid sinus massage, ECG showed ST segment elevation in leads II, III, and aVF. Four minutes after carotid sinus massage, he had chest pain with a progressive elevation in the ST segment in the same leads, when he had 99% focal spasm in the right coronary artery. The vasospasm induced by carotid sinus massage was reproducible over several minutes and resolved spontaneously. Coronary artery spasm may be provoked by the enhanced vagal activation due to carotid sinus massage. PMID- 10862605 TI - Guidelines for the management of iron deficiency anaemia. British Society of Gastroenterology. PMID- 10862606 TI - The inhibitor of apoptosis, cIAP2, functions as a ubiquitin-protein ligase and promotes in vitro monoubiquitination of caspases 3 and 7. AB - The inhibitor of apoptosis, cIAP2, contains a putative Ring finger motif at the C terminus. Using in vitro ubiquitination assays, we found that the Ring finger of cIAP2 alone possesses intrinsic ubiquitin ligase activity and promotes substrate independent ubiquitination. It also promotes ubiquitination of caspases 3 and 7 but not caspase-1. The Ring fingers of c-Cbl and Apc11 failed to promote caspase 7 ubiquitination, suggesting that the Ring finger of cIAP2 itself is involved in substrate recognition. PMID- 10862604 TI - UK guidelines on the management of variceal haemorrhage in cirrhotic patients. British Society of Gastroenterology. PMID- 10862607 TI - The conformation-dependent interaction of alpha 2-macroglobulin with vascular endothelial growth factor. A novel mechanism of alpha 2-macroglobulin/growth factor binding. AB - alpha(2)-Macroglobulin (alpha(2)M) is a highly conserved proteinase inhibitor present in human plasma at high concentration (2-4 mg/ml). alpha(2)M exists in two conformations, a native form and an activated, receptor-recognized form. While alpha(2)M binds to numerous cytokines and growth factors, in most cases, the nature of the alpha(2)M interaction with these factors is poorly understood. We examined in detail the interaction between alpha(2)M and vascular endothelial growth factor (VEGF) and found a novel and unexpected mechanism of interaction as demonstrated by the following observations: 1) the binding of VEGF to alpha(2)M occurs at a site distinct from the recently characterized growth factor binding site; 2) VEGF binds different forms of alpha(2)M with distinct spatial arrangement, namely to the interior of methylamine or ammonia-treated alpha(2)M and to the exterior of native and proteinase-converted alpha(2)M; and 3) VEGF (molecular mass approximately 40 kDa) can access the interior of receptor recognized alpha(2)M in the absence of a proteinase trapped within the molecule. VEGF bound to receptor-recognized forms of alpha(2)M is internalized and degraded by macrophages via the alpha(2)M receptor, the low density lipoprotein receptor related protein. Oxidation of both native and receptor-recognized alpha(2)M results in significant inhibition of VEGF binding. We also examined the biological significance of this interaction by studying the effect of alpha(2)M on VEGF-induced cell proliferation and VEGF-induced up-regulation of intracellular Ca(2+) levels. We demonstrate that under physiological conditions, alpha(2)M does not impact the ability of VEGF to induce cell proliferation or up regulate Ca(2+). PMID- 10862608 TI - Production of ceramides causes apoptosis during early neural differentiation in vitro. AB - To investigate signal transduction pathways leading to apoptosis during the early phase of neurogenesis, we employed PCC7-Mz1 cells, which cease to proliferate and begin to differentiate into a stable pattern of neurons, astroglial cells, and fibroblasts upon incubation with retinoic acid (RA). As part of lineage determination, a sizable fraction of RA-treated cultures die by apoptosis. Applying natural long-chain C(16)-ceramides as well as membrane-permeable C(2)/C(6)-ceramide analogs caused apoptosis, whereas the biologically nonactive C(2)-dihydroceramide did not. Treating PCC7-Mz1 stem cells with a neutral sphingomyelinase or with the ceramidase inhibitor N-oleoylethanolamine elevated the endogenous ceramide levels and concomitantly induced apoptosis. Addition of RA caused an increase in ceramide levels within 3-5 h, which reached a maximum (up to 3.5-fold of control) between days 1 and 3 of differentiation. Differentiated PCC7-Mz1 cells did not respond with ceramide formation and apoptosis to RA treatment. The acidic sphingomyelinase contributed only weakly and the neutral Mg(2+)-dependent and Mg(2+)-independent sphingomyelinases not at all to the RA-mediated production of ceramides. However, ceramide increase was sensitive to the ceramide synthase inhibitor fumonisin B(1), suggesting a crucial role for the de novo synthesis pathway. Enzymatic assays revealed that ceramide synthase activity remained unaltered, whereas serine palmitoyltransferase (SPT), a key enzyme in ceramide synthesis, was activated approximately 2.5-fold by RA treatment. Activation of SPT seemed to be mediated via a post-translational mechanism because levels of the mRNAs coding for the two SPT subunits were unaffected. Expression of marker proteins shows that ceramide regulates apoptosis, rather than differentiation, during early neural differentiation. PMID- 10862609 TI - Dehydroascorbic acid transport by GLUT4 in Xenopus oocytes and isolated rat adipocytes. AB - Dehydroascorbic acid (DHA), the first stable oxidation product of vitamin C, was transported by GLUT1 and GLUT3 in Xenopus laevis oocytes with transport rates similar to that of 2-deoxyglucose (2-DG), but due to inherent difficulties with GLUT4 expression in oocytes it was uncertain whether GLUT4 transported DHA (Rumsey, S. C. , Kwon, O., Xu, G. W., Burant, C. F., Simpson, I., and Levine, M. (1997) J. Biol. Chem. 272, 18982-18989). We therefore studied DHA and 2-DG transport in rat adipocytes, which express GLUT4. Without insulin, rat adipocytes transported 2-DG 2-3-fold faster than DHA. Preincubation with insulin (0.67 micrometer) increased transport of each substrate similarly: 7-10-fold for 2-DG and 6-8-fold for DHA. Because intracellular reduction of DHA in adipocytes was complete before and after insulin stimulation, increased transport of DHA was not explained by increased internal reduction of DHA to ascorbate. To determine apparent transport kinetics of GLUT4 for DHA, GLUT4 expression in Xenopus oocytes was reexamined. Preincubation of oocytes for >4 h with insulin (1 micrometer) augmented GLUT4 transport of 2-DG and DHA by up to 5-fold. Transport of both substrates was inhibited by cytochalasin B and displayed saturable kinetics. GLUT4 had a higher apparent transport affinity (K(m) of 0.98 versus 5.2 mm) and lower maximal transport rate (V(max) of 66 versus 880 pmol/oocyte/10 min) for DHA compared with 2-DG. The lower transport rate for DHA could not be explained by binding differences at the outer membrane face, as shown by inhibition with ethylidene glucose, or by transporter trans-activation and therefore was probably due to substrate-specific differences in transporter/substrate translocation or release. These novel data indicate that the insulin-sensitive transporter GLUT4 transports DHA in both rat adipocytes and Xenopus oocytes. Alterations of this mechanism in diabetes could have clinical implications for ascorbate utilization. PMID- 10862611 TI - Import of a cytosolic protein into lysosomes by chaperone-mediated autophagy depends on its folding state. AB - We have analyzed the folding state of cytosolic proteins imported in vitro into lysosomes, using an approach originally developed by Eilers and Schatz, (Eilers, M., and Schatz, G. (1986) Nature 322, 228-232) to investigate protein import into mitochondria. The susceptibility toward proteases of mouse dihydrofolate reductase (DHFR), synthesized in a coupled transcription-translation system with rabbit reticulocytes, decreased in the presence of its substrate analogue, methotrexate. This analogue complexes with high affinity with the in vitro synthesized DHFR and locks it into a protease-resistant folded conformation. DHFR was taken up by freshly isolated rat liver lysosomes and methotrexate reduced this uptake by about 80%. A chimeric DHFR protein, which carries the N-terminal presequence of subunit 9 of ATP synthase preprotein from Neurospora crassa fused to its N terminus, was taken up by lysosomes more efficiently. Again, methotrexate abolished the lysosomal uptake of the fusion protein, which was partially restored by washing of methotrexate from DHFR or by adding together methotrexate and dihydrofolate, the natural substrate of DHFR. Immunoblot analysis with anti-DHFR of liver lysosomes and of other fractions, isolated from rats starved for 88 h and treated with lysosomal inhibitors, suggests that DHFR is degraded by chaperone-mediated autophagy. Competition with ribonuclease A and stimulation by ATP/Mg(2+) and the heat shock cognate protein of 73 kDa show that the lysosomal uptake of the fusion protein also occurs by this pathway. It is concluded that the lysosomal uptake of cytosolic proteins by chaperone-mediated autophagy mainly occurs by passage of the unfolded proteins through the lysosomal membrane. Therefore, this mechanism is different from protein transport into peroxisomes, but similar to the import of proteins into the endoplasmic reticulum and mitochondria. PMID- 10862610 TI - Identification and functional characterization of human soluble epoxide hydrolase genetic polymorphisms. AB - Human soluble epoxide hydrolase (sEH), an enzyme directing the functional disposition of a variety of endogenous and xenobiotic-derived chemical epoxides, was characterized at the genomic level for interindividual variation capable of impacting function. RNA was isolated from 25 human liver samples and used to generate full-length copies of soluble epoxide hydrolase cDNA. The resulting cDNAs were polymerase chain reaction amplified, sequenced, and eight variant loci were identified. The coding region contained five silent single nucleotide polymorphisms (SNPs) and two variant loci resulting in altered protein sequence. An amino acid substitution was identified at residue 287 in exon 8, where the more common arginine was replaced by glutamine. A second variant locus was identified in exon 13 where an arginine residue was inserted following serine 402 resulting in the sequence, arginine 403-404, instead of the more common, arginine 403. This amino acid insertion was confirmed by analyzing genomic DNA from individuals harboring the polymorphic allele. Slot blot hybridization analyses of the liver samples indicated that sEH mRNA steady-state expression varied approximately 10-fold. Transient transfection experiments with CHO and COS-7 cells were used to demonstrate that the two new alleles possess catalytic activity using trans-stilbene oxide as a model substrate. Although the activity of the glutamine 287 variant was similar to the sEH wild type allele, proteins containing the arginine insertion exhibited strikingly lower activity. Allelic forms of human sEH, with markedly different enzymatic profiles, may have important physiological implications with respect to the disposition of epoxides formed from the oxidation of fatty acids, such as arachidonic acid-derived intermediates, as well in the regulation of toxicity due to xenobiotic epoxide exposures. PMID- 10862612 TI - KATP channels regulate mitogenically induced proliferation in primary rat hepatocytes and human liver cell lines. Implications for liver growth control and potential therapeutic targeting. AB - To determine whether K(ATP) channels control liver growth, we used primary rat hepatocytes and several human cancer cell lines for assays. K(ATP) channel openers (minoxidil, cromakalim, and pinacidil) increased cellular DNA synthesis, whereas K(ATP) channel blockers (quinidine and glibenclamide) attenuated DNA synthesis. The channel inhibitor glibenclamide decreased the clonogenicity of HepG2 cells without inducing cytotoxicity or apoptosis. To demonstrate the specificity of drugs for K(+) channels, whole-cell patch-clamp recordings were made. Hepatocytes revealed K(+) currents with K(ATP) channel properties. These K(+) currents were augmented by minoxidil and pinacidil and attenuated by glibenclamide as well as tetraethylammonium, in agreement with established responses of K(ATP) channels. Reverse transcription of total cellular RNA followed by polymerase chain reaction showed expression of K(ATP) channel specific subunits in rat hepatocytes and human liver cell lines. Calcium fluxes were unperturbed in glibenclamide-treated HepG2 cells and primary rat hepatocytes following induction with ATP and hepatocyte growth factor, respectively, suggesting that the effect of K(ATP) channel activity upon hepatocyte proliferation was not simply due to indirect modulation of intracellular calcium. The regulation of mitogen-related hepatocyte proliferation by K(ATP) channels advances our insights into liver growth control. The findings have implications in mechanisms concerning liver development, regeneration, and oncogenesis. PMID- 10862613 TI - SUMO-1 conjugation to human DNA topoisomerase II isozymes. AB - Topoisomerase I-mediated DNA damage induced by camptothecin has been shown to induce rapid small ubiquitin-related modifier (SUMO)-1 conjugation to topoisomerase I. In the current study, we show that topoisomerase II-mediated DNA damage induced by teniposide (VM-26) results in the formation of high molecular weight conjugates of both topoisomerase IIalpha and IIbeta isozymes in HeLa cells. Immunological characterization of these conjugates suggests that both topoisomerase IIalpha and IIbeta isozymes are conjugated to SUMO-1. The involvement of SUMO-1/UBC9 in the modification of topoisomerase II isozymes is also supported by the demonstration of physical interaction between topoisomerase II and SUMO-1/UBC9. Surprisingly, ICRF-193, which does not induce topoisomerase II-mediated DNA damage but traps topoisomerase II into a circular clamp conformation, is also shown to induce similar SUMO-1 conjugation to topoisomerase II isozymes. In addition, we show that both oxidative and heat shock stresses, which can cause protein damage, rapidly increase nuclear SUMO-1 conjugates. These studies raise the question on whether SUMO-1 conjugation to topoisomerases is an indirect result of a DNA damage response or a direct result because of protein conformational changes. PMID- 10862614 TI - Hypermodification of tRNA in Thermophilic archaea. Cloning, overexpression, and characterization of tRNA-guanine transglycosylase from Methanococcus jannaschii. AB - tRNA is structurally unique among nucleic acids in harboring an astonishing diversity of modified nucleosides. Two structural variants of the hypermodified nucleoside 7-deazaguanosine have been identified in tRNA: queuosine, which is found at the wobble position of the anticodon in bacterial and eukaryotic tRNA, and archaeosine, which is found at position 15 of the D-loop in archaeal tRNA. From homology searching of the Methanococcus jannaschii genome, a gene coding for an enzyme in the biosynthesis of archaeosine (tgt) was identified and cloned. The tgt gene was overexpressed in an Escherichia coli expression system, and the recombinant tRNA-guanine transglycosylase enzyme was purified and characterized. The enzyme catalyzes a transglycosylation reaction in which guanine is eliminated from position 15 of the tRNA and an archaeosine precursor (preQ(0)) is inserted. The enzyme is able to utilize both guanine and the 7-deazaguanine base preQ(0) as substrates, but not other 7-deazaguanine bases, and is able to modify tRNA from all three phylogenetic domains. The enzyme shows optimal activity at high temperature and acidic pH, consistent with the optimal growth conditions of M. jannaschii. The nature of the temperature dependence is consistent with a requirement for some degree of tRNA tertiary structure in order for recognition by the enzyme to occur. PMID- 10862615 TI - Differential effects of low density lipoproteins on insulin-like growth factor-1 (IGF-1) and IGF-1 receptor expression in vascular smooth muscle cells. AB - Low density lipoproteins (LDLs) play an important role in the pathogenesis of atherosclerosis. LDL has been shown to be mitogenic and proapoptotic for vascular smooth muscle cells. However, the mechanisms are poorly understood and may result from an alteration in intracellular mitogenic signaling either directly by LDL or indirectly through an autocrine effect involving growth factor secretion and/or growth factor receptor expression. Insulin-like growth factor-1 (IGF-1) is an autocrine/paracrine factor for vascular smooth muscle cells and has potent anti apoptotic effects. Thus, we hypothesized that part of the proliferative responses to LDLs may be explained by its modulation of IGF-1 or IGF-1 receptor (IGF-1R) expression. Treatment of rat vascular smooth muscle cells with increasing doses of native LDL dose-dependently increased IGF-1 mRNA by up to 2.6-fold; however, native LDL had no effect on IGF-1R mRNA expression. In contrast, the same doses of oxidized LDL significantly reduced IGF-1 and IGF-1R mRNA by 80 and 61%, respectively, and reduced IGF-1 and IGF-1R protein expression by 63 and 46%. In addition, native and oxidized LDL significantly increased IGF-1-binding protein-2 and IGF-1-binding protein-4 expression as measured by Western ligand blot. Most interestingly, anti-IGF-1 antiserum completely inhibited LDL-induced but not serum-induced increase in (3)H-thymidine incorporation, indicating a requirement for IGF-1 in the LDL-stimulated mitogenic signaling pathway. In summary, these results suggest that native and oxidized LDLs have differential effects on IGF-1 and IGF-1R expression. Because IGF-1 is a potent survival factor for vascular smooth muscle cells, our findings suggest that moderately oxidized LDL may favor proliferation of smooth muscle cells, whereas oxidized LDL may contribute to plaque apoptosis by local depletion of IGF-1 and IGF-1R. PMID- 10862616 TI - Conformational requirements of collagenous peptides for recognition by the chaperone protein HSP47. AB - The collagen binding chaperone HSP47 interacts with procollagen in the endoplasmic reticulum and plays a crucial role in the biosynthesis of collagen. We recently demonstrated that typical collagen model peptides, (Pro-Pro-Gly)(n), possess sufficient structural information for interaction with HSP47 (Koide, T., Asada, S., and Nagata, K. (1999) J. Biol. Chem. 274, 34523-34526). Here we show that binding of (Gly-Pro-Pro)(n) peptides to HSP47 can be detected using the two hybrid system in yeast if a trimerizing domain is fused to the C termini of the peptides. Some peptides interacted with HSP47 at a lowered assay temperature at 24 degrees C but not at 30 degrees C, indicating the importance of conformational change of the substrate peptides. To analyze the spectrum of HSP47 substrate sequences, we performed two-hybrid screening of collagen-like peptides in designed random peptide libraries using HSP47 as a bait. In selected peptides, the enrichment ratio calculated for each amino acid residue correlated strongly with the contribution of the residue to triple-helix stability independently determined using synthetic collagen model peptides. Taken together, our results suggest that HSP47 preferentially recognizes collagenous Gly-X-Y repeats in triple-helical conformation. We also demonstrated that screening of combinatorial peptide libraries is a powerful strategy to determine conformational requirements as well as the elucidation of binding motifs in primary structure. PMID- 10862617 TI - The proliferative and migratory activities of breast cancer cells can be differentially regulated by heparan sulfates. AB - To explore how heparan sulfate (HS) controls the responsiveness of the breast cancer cell lines MCF-7 and MDA-MB-231 to fibroblast growth factors (FGFs), we have exposed them to HS preparations known to have specificity for FGF-1 (HS glycosaminoglycan (HSGAG A)) or FGF-2 (HSGAGB). Proliferation assays confirmed that MCF-7 cells were highly responsive to FGF-2 complexed with GAGB, whereas migration assays indicated that FGF-1/HSGAGA combinations were stimulatory for the highly invasive MDA-MB-231 cells. Quantitative polymerase chain reaction for the levels of FGF receptor (FGFR) isoforms revealed that MCF-7 cells have greater levels of FGFR1 and that MDA-MB-231 cells have greater relative levels of FGFR2. Cross-linking demonstrated that FGF-2/HSGAGB primarily activated FGFR1, which in turn up-regulated the activity of mitogen-activated protein kinase; in contrast, FGF-1/HSGAGA led to the phosphorylation of equal proportions of both FGFR1 and FGFR2, which in turn led to the up-regulation of Src and p125(FAK). MDA-MB-231 cells were particularly responsive to vitronectin substrates in the presence of FGF-1/HSGAGA, and blocking antibodies established that they used the alpha(v)beta(3) integrin to bind to it. These results suggest that the clustering of particular FGFR configurations on breast cancer cells induced by different HS chains leads to distinct phenotypic behaviors. PMID- 10862618 TI - Geldanamycin induces ErbB-2 degradation by proteolytic fragmentation. AB - Exposure of carcinoma cell lines to the antibiotic geldanamycin induces the degradation of ErbB-2, a co-receptor tyrosine kinase that is frequently overexpressed in certain tumors. Using ErbB-2 mutants expressed as chimeric receptors or green fluorescent protein fusion proteins, we report that the kinase domain of ErbB-2 is essential for geldanamycin-induced degradation. The kinase domain of the related epidermal growth factor receptor was not sensitive to this drug. The data further indicate mechanistic aspects of ErbB-2 degradation by geldanamycin. The data show that exposure to the drug induces at least one cleavage within the cytoplasmic domain of ErbB-2 producing a 135-kDa fragment and a 23-kDa fragment. The latter represents the carboxyl-terminal domain of ErbB-2, whereas the former represents the ectodomain and part of the cytoplasmic domain. Degradation of the carboxyl-terminal fragment is prevented by proteasome inhibitors, whereas degradation of the membrane-anchored 135-kDa ErbB-2 fragment is blocked by inhibitors of the endocytosis-dependent degradation pathway. Confocal microscopy studies confirm a geldanamycin-induced localization of ErbB-2 on intracellular vesicles. PMID- 10862619 TI - Interaction between yeast sgs1 helicase and DNA topoisomerase III. AB - The Saccharomyces cerevisiae Sgs1 protein is a member of the RecQ family of DNA helicases that includes the human Bloom's syndrome and Werner's syndrome proteins. In this work, we report studies on the interaction between Sgs1 and DNA topoisomerase III in vitro and in vivo. Affinity chromatography experiments with various fragments of Sgs1, a 1447-amino acid polypeptide, suggested that its N terminal one-fifth was sufficient for interaction with DNA topoisomerase III. Gel electrophoretic mobility shift assays also indicated that a fragment Sgs1(1-283), containing residues 1-283, inhibited the binding of DNA topoisomerase III to single-stranded DNA. A shorter protein fragment containing residues 1-107 also showed partial inhibition in these assays. Studies of a sgs1 top1 double mutant lacking both Sgs1 and DNA topoisomerase I showed that the slow growth phenotype of this double mutant is suppressed by expressing full-length Sgs1, but not Sgs1 without the N-terminal 107 amino acid residues. In sgs1 top3 cells devoid of DNA topoisomerase III, however, expression of full-length Sgs1 or Sgs1 lacking the N terminal 107 amino acid residues has the same effect of reducing the growth rate of the double mutant. These in vitro and in vivo data indicate that Sgs1 and DNA topoisomerase III physically interact and that this interaction is physiologically significant. PMID- 10862620 TI - Cloning and characterization of erythroid-specific DNase I-hypersensitive site in human rhesus-associated glycoprotein gene. AB - Rhesus-associated glycoprotein is a critical co-factor in the expression of rhesus blood group antigens. We identified and cloned an erythroid-specific major DNase I-hypersensitive site located about 10 kilobases upstream from the translation start site of the RHAG gene. A short core enhancer sequence of 195 base pairs that corresponded with the major hypersensitive site and possessed position- and orientation-independent enhancer activity in K562 cells. In vitro DNase I footprint analysis revealed four protected regions in the core enhancer; two GATA motifs, an Ets-like motif and an unknown motif. The GATA motifs bound GATA-1 and mutagenesis analysis revealed that the proximal one is critical for the enhancing activity. Homology plot analysis using the 5' sequence of the mouse RHAG gene revealed four homologous stretches and multiple insertions of repetitive sequences among them; four LINE/L1 and four Alu in the human and as well as one LINE/L1 and one LTR/MaLR in the mouse gene. The highly conservative enhancer region was flanked by SINE and LINE/L1 in both species. These results suggest that the 5'-flanking sequence of RHAG gene is a preferable target sequence for retroviral transposition and that the enhancer was inserted in the same manner, resulting in the acquisition of erythroid dominant expression. PMID- 10862621 TI - Modulation of the murine peroxisome proliferator-activated receptor gamma 2 promoter activity by CCAAT/enhancer-binding proteins. AB - Peroxisome proliferator-activated receptor gamma (PPARgamma) and CCAAT/enhancer binding proteins (C/EBPs) are transcriptional regulators essential for adipocyte differentiation and function. Previous findings indicate that PPARgamma2 transcription is regulated by members of the C/EBP family. We demonstrate here that C/EBPalpha and C/EBPdelta, but not C/EBPbeta, induce the activity of the PPARgamma2 promoter in transiently transfected 3T3-L1 preadipocytes and bind to two juxtaposed low affinity C/EBP binding sites. Results obtained with chimeras containing interchanged C/EBPalpha-C/EBPbeta N-terminal transactivation domain and C-terminal DNA binding dimerization domain indicate that the N-terminal part of C/EBPbeta prevents it from binding to the PPARgamma2 promoter. Indeed, deletion mutants of C/EBPbeta lacking the N-terminal part of the molecule are able to bind to the PPARgamma2 promoter. We further demonstrate that deletion of a region located between amino acids 184-212, upstream of the DNA binding domain, permits C/EBPbeta binding to the PPARgamma2 promoter, implicating an inhibitory region in C/EBPbeta for modulating DNA binding specificity to the PPARgamma2 promoter. In summary, this study indicates that C/EBPbeta but not C/EBPalpha or C/EBPdelta is unable to bind to C/EBP binding sites in the mouse PPARgamma2 promoter. The lack of binding is due to a region N-terminal of the C/EBPbeta DNA binding domain. Our findings illustrate a mechanism by which C/EBP isoforms differentially modulate the transactivation of the PPARgamma2 promoter. PMID- 10862622 TI - Cloning, overexpression, and characterization of peroxiredoxin and NADH peroxiredoxin reductase from Thermus aquaticus. AB - The genes for peroxiredoxin (Prx) and NADH:peroxiredoxin oxidoreductase (PrxR) have been cloned from the thermophilic bacterium Thermus aquaticus. prx is located upstream from prxR, the two genes being separated by 13 bases. The amino acid sequences show that Prx is related to two-cysteine peroxiredoxins from a range of organisms and that PrxR resembles NADH-dependent flavoenzymes that catalyze the reduction of peroxiredoxins in mesophilic bacteria. The sequence of PrxR also resembles those of thioredoxin reductases (TrxR) from thermophiles but with an N-terminal extension of about 200 residues. PrxR has motifs for two redox active disulfides, one in the FAD-binding site, as occurs in TrxR, and the other in the N-terminal extension. The molecular masses of the monomers of Prx and PrxR are 21.0 and 54.9 kDa, respectively; both enzymes exist as multimers. The recombinant flavoenzyme requires 3 mol equivalents of dithionite for full reduction, as is consistent with 1 FAD and 2 disulfides per monomer. PrxR and Prx together catalyze the anaerobic reduction of hydrogen peroxide. The activity of Prx is much less than has been observed with homologous proteins. Prx appears to be inactivated by cumene hydroperoxide. PrxR itself has low peroxidase activity. PMID- 10862623 TI - Heat and chemical shock potentiation of glucocorticoid receptor transactivation requires heat shock factor (HSF) activity. Modulation of HSF by vanadate and wortmannin. AB - Heat shock and other forms of stress increase glucocorticoid receptor (GR) activity in cells, suggesting cross-talk between the heat shock and GR signal pathways. An unresolved question concerning this cross-talk is whether heat shock factor (HSF1) activity is required for this response. We addressed this issue by modulating HSF1 activity with compounds acting by distinct mechanisms: sodium vanadate (SV), an inhibitor of protein phosphatases; and wortmannin, an inhibitor of DNA-dependent protein kinase. Using HSF1- and GR-responsive CAT reporters, we demonstrate that SV inhibits both HSF1 activity and the stress potentiation of GR, while having no effect on the hormone-free GR or HSF1. Paradoxically, SV increased hormone-induced GR activity in the absence of stress. In contrast, wortmannin increased HSF1 activity in stressed cells and had no effect on HSF1 in the absence of stress. Using the pMMTV-CAT reporter containing the negative regulatory element 1 site for DNA-dependent protein kinase, wortmannin was found to increase the GR response. However, in cells expressing a minimal promoter lacking negative regulatory element 1 sites, wortmannin had no effect on the GR in the absence of stress but increased the stress potentiation of GR. Our results show that the mechanism by which GR activity is increased in stressed cells requires intrinsic HSF1 activity. PMID- 10862624 TI - Transgenic animal models for the analysis of the renal endothelin system. PMID- 10862625 TI - Renal effects of smoking: potential mechanisms and perspectives. PMID- 10862626 TI - ACTH revisited--potential implications for patients with renal disease. PMID- 10862627 TI - Encrusted pyelitis: an underdiagnosed condition? PMID- 10862628 TI - The role of cytokines in skeletal remodelling: possible consequences for renal osteodystrophy. PMID- 10862629 TI - Borderline kidney graft donors--what are the problems? PMID- 10862630 TI - Are there two types of malnutrition in chronic renal failure? Evidence for relationships between malnutrition, inflammation and atherosclerosis (MIA syndrome). PMID- 10862631 TI - Nephrology, dialysis and transplantation in Shanghai, 1999. PMID- 10862632 TI - Capillary/myocyte mismatch in the heart in renal failure--a role for erythropoietin? AB - BACKGROUND: Chronic renal failure is characterized by remodeling of the heart with left ventricular hypertrophy (increasing oxygen demand) and capillary deficit leading to capillary/myocyte mismatch (decreasing oxygen supply). Erythropoietin (Epo) has known angiogenic properties causing endothelial cell activation, migration and sprouting, mediated at least in part via the JAK/STAT (Janus kinase/signal transducers and activators of transcription) pathway. In uraemic cardiac hypertrophy the presence of diminished capillary supply implies that capillary growth does not keep pace with development of hypertrophy. To investigate whether this was due to a deficit of the angiogenic hormone Epo we examined whether Epo levels are altered and whether an increase in haematocrit by administration of rhEpo influences capillary supply, i.e. capillary/myocyte mismatch in experimental renal failure. METHOD: Male Spraque-Dawley rats were either subjected to partial renal ablation or sham operation. Only modest amounts of renal tissue were removed so that the rats were not anemic. Subgroups of rats received either human (rh)Epo alone or in combination with unspecific antihypertensive treatment (dihydralazine plus furosemide) in order to control the Epo induced rise in blood pressure. Capillary supply was measured stereologically as capillary length per volume myocardium using the orientator method. RESULTS: Capillary length density was reduced by approximately 25% after partial renal ablation (3237+/-601 vs 4293+/-501 mm/mm(3) in controls). It was not statistically different in animals with partial renal ablation+rhEpo+antihypertensive treatment (3620+/-828 mm/mm(3)) compared to partial ablation alone. CONCLUSION: The study shows that lack of Epo does not cause, or contribute to, the deficit of capillary growth in the hypertrophied left ventricle of rats with renal failure. In addition, a rise in haematocrit is not accompanied by beneficial effects on alterations of cardiovascular structure in experimental renal failure. PMID- 10862633 TI - Antenatal Bartter syndrome with sensorineural deafness: refinement of the locus on chromosome 1p31. AB - BACKGROUND: Recently a locus for antenatal Bartter syndrome associated with sensorineural deafness was mapped to human chromosome 1p31 in a single consanguineous Bedouin family (Brennan et al. Am J Hum Genet 1998; 62: 355-361). METHODS: By haplotype analysis we demonstrate linkage to this locus in nine consanguineous families with antenatal Bartter syndrome associated with sensorineural deafness. RESULTS: The critical interval compatible with linkage was refined to 4.0 cM by two novel recombinational events with markers D1S2661 and D1S475. CONCLUSION: We thereby confirmed this gene locus and distinguished this clinical subtype from other variants of Bartter syndrome as a new disease entity. PMID- 10862634 TI - Prevalence and characterization of renal tubular acidosis in patients with osteopenia and osteoporosis and in non-porotic controls. AB - BACKGROUND: Chronic metabolic acidosis may increase alkali mobilization from the bone and thus promote the development of osteoporosis. The objective of the current study was to compare urinary acidification in patients with reduced bone mineral content with that in control subjects with normal bone density. METHODS: Forty-six subjects (41 females, 5 males) with osteopenia or osteoporosis were studied. In none of the subjects were overt metabolic acidosis, derangement of potassium homeostasis, or renal insufficiency present. Distal tubular acidification was studied by means of oral ammonium chloride loading test (0.1 g/kg body weight) and the oral frusemide test (40 mg). In addition the frusemide test was performed in 20 healthy age- and sex-matched controls (17 females, 3 males). RESULTS: In all control subjects a urinary pH <5. 5 was observed following the ingestion of 40 mg frusemide. In contrast, in patients with reduced bone mineral density incomplete renal tubular acidosis type I (RTA I) was diagnosed in 10 of 46 subjects (22%) by oral ammonium chloride loading test. Disorders possibly related to RTA I were detected in eight of these 10 patients. Thirty-six patients had a normal urinary pH response following oral ammonium chloride loading. Oral frusemide, 40 mg, failed to lower urinary pH <5.5 in sixteen patients (35%), these included 10 subjects with incomplete RTA I, and six subjects with a normal oral ammonium chloride loading test. An abnormal frusemide test was found in 35% of patients with reduced bone mass and in none of the normal controls (chi(2)=7.39; P<0.01). With the ammonium chloride test as the gold standard for diagnosis of distal RTA, the frusemide test showed a sensitivity of 1.0 (95% CI, 0.69-1.0) and a specificity of 0.89 (95% CI, 0.78 0.96) for the diagnosis of distal RTA. Patients with incomplete RTA I were younger than those without incomplete RTA I (42+/-16 vs 54+/-14 years; P=0.025; mean+/-SD). Basal serum bicarbonate concentrations and capillary pH did not differ between the groups. CONCLUSION: Incomplete RTA I may be prevalent in a significant proportion of patients suffering from osteopenia or osteoporosis. The outcome of the frusemide test suggests either a defect of the H(+)ATPase in the cortical collecting tubule (CCT) or a defective Na(+) reabsorption in the CCT. Prospective studies are needed to further elucidate the impact of incomplete RTA I on the development of reduced bone mineral content. PMID- 10862635 TI - Adult-onset idiopathic nephrotic syndrome associated with pure diffuse mesangial hypercellularity. AB - BACKGROUND: Pure diffuse mesangial hypercellularity (DMH), in its primary form, is a relatively rare histological finding and few data exist in the literature regarding its clinical course and prognosis in nephrotic adults with this diagnosis. METHODS: We retrospectively analysed the clinical and histological data of 28 adult nephrotic patients (13 male) with this diagnosis with regard to response to the treatment, outcome and prognostic indicators. RESULTS: Of 25 patients treated with prednisolone (Pred), nine (36%) showed complete remission (CR) of proteinuria, eight (32%) partial remission (PR) and eight (32%) did not respond at all (NR). The combination of cyclosporin treatment with prednisolone of those with PR or NR produced one further complete and two partial remissions. At the end of follow-up (mean 64 months), 10 patients (40%) were in CR, nine (36%) in PR and six (24%) were NR and remained nephrotic. Renal function remained unchanged in patients with CR or PR. In contrast, the six non-responders progressed to end-stage renal disease (ESRD). Compared with non-responders, patients who responded to Pred were older and had normal renal function at presentation. This group also had less mesangial sclerosis and severe tubulointerstitial fibrosis and none showed synechiae with Bowman's capsule. IgM mesangial deposits were observed in 22% of patients with CR in response to Pred, in 37% of those with PR and in 100% of non-responders, who finally progressed to ESRD. A multivariate analysis of clinical and histological features at biopsy showed persistent nephrotic syndrome (P<0.001), the severity of DMH (P<0.03) and the presence of mesangial IgM (P<0. 01) to have independent predictive value for ESRD. This analysis also demonstrated that only mesangial sclerosis (P<0.03) and the presence of mesangial IgM (P<0.002) independently predicted the response to therapy. CONCLUSIONS: DMH associated with idiopathic nephrotic syndrome is a heterogeneous entity. Patients who respond to therapy (completely or partially) have a benign course similar to that of minimal change nephrotic syndrome. They are usually older and have normal renal function at presentation, whereas 'sclerotic' lesions are less frequent findings in initial biopsies. Non responders tend to be younger and progress to ESRD. Most of them have impaired renal function at first assessment and more prominent 'sclerotic' lesions on initial biopsies. Mesangial IgM is an independent marker of poorer response to treatment and progression to ESRD but it lacks specificity. PMID- 10862636 TI - Effects of angiotensin II blockade on nitric oxide blood levels in IgA nephropathy. AB - BACKGROUND: The effects of renin-angiotensin system blockade on nitric oxide (NO), especially in pathological conditions, are far from being established. The influence of kinins and angiotensin type 2 receptor are largely speculative and based mainly on animal studies. This study was aimed to address these aspects in humans. METHODS: Eight IgA nephropathy patients with documented clinical and histological indicators of poor prognosis were given 50 mg of losartan, 10 mg of enalapril, and 40 mg of the NO donor isosorbide 5 mononitrate (as a control of NO generation) in randomized order for 7 days each. Treatment periods were separated by washout periods of 7 days each. Laboratory investigations were performed before and after each study period. Seven healthy controls received losartan and enalapril according to the same study design. RESULTS: Glomerular filtration rate remained stable while effective renal plasma flow increased with each treatment (P<0.05). Under losartan and enalapril, filtration fraction fell (P=0.02), plasma renin activity increased (P<0.05) and urinary aldosterone concentration decreased (P=0.02). Angiotensin-converting enzyme activity was reduced to the limit of detection under enalapril (P<0.001). Blood NO, detected as nitrosylhaemoglobin by a recently developed technique of spin-trap electron paramagnetic resonance, increased significantly, as expected, during treatment with isosorbide 5 mononitrate (P=0.01), with enalapril (P<0.05), and also with losartan (P<0.05). Unlike losartan, enalapril significantly reduced albuminuria (P=0.01) in this short-term period. In the seven healthy controls, neither enalapril nor losartan were able to increase blood NO levels significantly. CONCLUSIONS: Blood levels of nitrosylhaemoglobin, a surrogate marker of NO, increased under blockade of the renin-angiotensin system in patients with IgA nephropathy, but not in healthy volunteers. This increase could contribute to changes of effective renal plasma flow in renal disease states. PMID- 10862637 TI - Increased blood levels of platelet-activating factor in insulin-dependent diabetic patients with microalbuminuria. AB - BACKGROUND: Platelet-activating factor (PAF), a phospholipid mediator of inflammation, may induce an enhanced size- and charge-dependent glomerular permeability in experimental animals. Studies on the role of PAF in enhanced glomerular permeability in the early phase of diabetic nephropathy are still lacking. METHODS: We evaluated the intravascular levels of PAF and its main catabolic enzyme, the PAF-specific plasma acetyl-hydrolase (PAF-AH), in basal conditions and after exercise, in normo- or micro-albuminuric insulin-dependent diabetic (IDD) patients and in normal subjects. RESULTS: The results obtained indicate that the concentration of PAF in whole blood was significantly enhanced in basal conditions, during and after exercise in all microalbuminuric IDD patients, but not in normoalbuminuric IDD or in control subjects. The increased concentration of PAF did not correlate with changes in the activity of PAF-AH, suggesting an enhanced production rather than a decreased catabolism of PAF. CONCLUSIONS: These results indicate an association between increased production of PAF and enhanced glomerular permeability in microalbuminuric IDD patients. PMID- 10862638 TI - Genetic polymorphisms of the renin-angiotensin system and complications of insulin-dependent diabetes mellitus. AB - OBJECTIVE: Patients with insulin-dependent diabetes mellitus (IDDM) have a high risk of developing diabetic nephropathy, retinopathy and cardiovascular diseases. The contribution of gene polymorphisms of the renin angiotensin system to these complications is controversial and may differ among populations. METHODS: In 257 Dutch IDDM patients (188 with urinary albumin excretion (UAE) <30 mg/24 h), logistic regression analysis was used to study the relationships among, on the one hand, the insertion/deletion gene polymorphism of the angiotensin-converting enzyme gene (ACE-ID), the M235T gene polymorphism of the angiotensinogen gene (AGT-M235T), and the A1166C gene polymorphism of the angiotensin type 1 receptor gene (AT1-A1166C), and, on the other hand, UAE, retinopathy, hypertension, and coronary heart disease. RESULTS: The T-allele of the AGT-M235T polymorphism was associated with an increased risk of an elevated UAE (odds ratio (OR) 3.03; 95% confidence interval (CI) 1.06-8.61), but only when interaction with the D-allele of the ACE-ID polymorphism was considered. A previously described positive interaction between the T-allele of the AGT-M235T polymorphism and the D-allele of the ACE-ID polymorphism could not be confirmed. The T-allele was also associated with an increased risk of retinopathy (OR 3.89, 95% CI 1.79-8.47). The CC-genotype of the AT1-A1166C polymorphism was associated with hypertension (OR 3.58; 95% CI 1. 23-10.37). CONCLUSIONS: In a Dutch IDDM population, including 69 patients with (incipient) diabetic nephropathy, the T-allele of the AGT-M235T polymorphism is associated with an elevated UAE and diabetic retinopathy and the CC-genotype of the AT1-A1166C polymorphism is associated with hypertension. A previously described interaction between the AGT-M235T and the ACE-ID polymorphisms could not be confirmed. Since the number of nephropathic patients in this study is small, these conclusions must be interpreted with caution. PMID- 10862639 TI - Increased frequency of angiotensin-converting enzyme DD genotype in patients with type 2 diabetes in Taiwan. AB - BACKGROUND: Diabetes is one of the major causes of end-stage renal failure in the Taiwanese population. Previous studies have shown that angiotensin-converting enzyme (ACE) inhibitor can improve glucose utilization and suppress hepatic glucose production and the renin-angiotensin system may play an important role in the initiation and progression of diabetic nephropathy. Thus, ACE gene polymorphism may be associated with type 2 diabetes and diabetic nephropathy. METHODS: To investigate the distribution of ACE-I/D genotype in type 2 diabetes and diabetic nephropathy, we examined 336 patients with type 2 diabetes (157 without nephropathy and 179 with nephropathy) and 263 age-matched normal controls. The diagnosis of nephropathy was made when daily protein loss exceeded 500 mg. ACE gene polymorphism was analysed by use of polymerase chain reaction. RESULTS: Our study revealed that the frequency of the D allele of the ACE gene was 29.3% in normal controls. The frequency of ACE DD genotype was significantly higher in type 2 diabetics compared with normal controls (18.2 vs 9.1%, P<0.01). The frequency of ACE DD genotype in patients with diabetic nephropathy was significantly higher than in patients without nephropathy (22.3 vs 13.4%, P<0.05). To determine whether ACE gene polymorphism was associated with the severity of diabetic nephropathy, we divided patients with diabetic nephropathy into dialysis and non-dialysis groups. The frequency of ACE DD genotype in the dialysis group was significantly higher than in non-dialysis group (28.7 vs 15.3%, P<0.05). CONCLUSION: Our results indicate that the frequency of ACE DD genotype is markedly higher in patients with type 2 diabetes, and the ACE DD genotype is significantly associated with diabetic nephropathy. PMID- 10862640 TI - Arterial stiffening and vascular calcifications in end-stage renal disease. AB - BACKGROUND: Epidemiological studies have identified aortic stiffness as an independent predictor of cardiovascular mortality in end-stage renal disease (ESRD) patients. In these patients, aortic pulse wave velocity (PWV) was associated with mediacalcosis, but the influence of arterial calcifications on the viscoelastic properties of large arteries was not well characterized. The purpose of the present study was to analyse the influence of arterial calcifications on arterial stiffness in stable haemodialysed patients. METHODS: We studied 120 stable ESRD patients on haemodialysis. All patients underwent B mode ultrasonography of common carotid artery (CCA), aorta, and femoral arteries to determine CCA distensibility, the elastic incremental modulus (Einc), and the presence of vascular calcifications. All patients underwent measurement of aortic PWV and echocardiogram. The presence of calcifications was analysed semiquantitatively as a score (0 to 4) according to the number of arterial sites with calcifications. RESULTS: Our observations indicate that arterial and aortic stiffness is significantly influenced by the presence and extent of arterial calcifications. The extent of arterial calcifications is in part responsible for increased left ventricular afterload, and is inversely correlated with stroke volume. The influence of calcifications is independent of the role of ageing and blood pressure. Arterial calcifications density increases with age, duration of haemodialysis, the fibrinogen level, and the prescribed dose of calcium-based phosphate binders. CONCLUSIONS: The results of this study showed that the presence of vascular calcifications in ESRD patients was associated with increased stiffness of large capacity, elastic-type arteries, like the aorta and CCA. The extent of arterial calcifications increased with the use of calcium based phosphate-binders. PMID- 10862641 TI - Management of anaemia in United Kingdom renal units: a report from the UK Renal Registry. AB - BACKGROUND: Morbidity and mortality of patients undergoing renal replacement therapy is influenced by the adequacy of correction of renal anaemia. The Renal Association has set standards for attainment of a target haemoglobin of 10 g/dl. This study compared the management of anaemia in dialysis patients in nine renal units in the UK. METHODS: A cross-sectional analysis was carried out on data submitted electronically to the UK Renal Registry. There were 1449 haemodialysis patients and 741 peritoneal dialysis patients in the nine renal centres analysed. Individual patient data were collected on haemoglobin, ferritin, erythropoietin prescription, and pre- and post-dialysis urea concentrations. RESULTS: None of the centres achieved the standard of more than 80% of haemodialysis patients with a haemoglobin of greater than 10 g/dl. Three centres achieved this standard for peritoneal dialysis. There was wide variation between centres in the percentage reaching the target. Differences in ferritin, erythropoietin prescription, and dialysis doses between centres could not entirely explain the variations between centres. Females had lower haemoglobin than males despite a greater proportion being treated with erythropoietin. There was a trend of increasing haemoglobin concentration during the study period in haemodialysis but not in peritoneal dialysis patients. CONCLUSIONS: The Renal Association standards for management of anaemia are not being met. The data allow renal centres to compare their practices with others to identify areas that might be improved. Further analysis may allow a benchmark to be determined of what it is possible to achieve by best practice. PMID- 10862643 TI - Insufficient penetration of systemic vancomycin into the PermCath lumen. AB - BACKGROUND: Catheter infection is a major cause of morbidity and catheter loss in chronic haemodialysis patients. There has been a large discrepancy in the catheter salvage rate, after an episode of documented bacteraemia, whether the patients receive systemic antibiotic alone or systemic antibiotics concomitant with 'antibiotic-lock technique' (20-30% vs 100%, respectively). To test the hypothesis that vancomycin may not adequately penetrate into the lumen of the catheter, despite therapeutic plasma levels, a series of in-vivo, ex-vivo, and in vitro experiments were performed. METHODS: We compared serum and intralumenal (0.3-0.5 ml aspirate from venous port of the catheter) vancomycin concentrations in 24 chronic haemodialysis patients, with documented bacteraemia, who had received prior systemic vancomycin therapy with 14 similar patients who had additionally received 'vancomycin-lock technique' (100 microg/ml of vancomycin in heparin solution) after each haemodialysis session. RESULTS: Despite serum vancomycin concentration of approximately 17 microg/ml in each group, the vancomycin concentration in the venous hub of the catheter was only 0.2+/-0.6 microg/ml in the former group, in sharp contrast to 125. 6+/-13 microg/ml in the latter group. In the ex-vivo experiment, four uninfected PermCaths which had been removed were immediately fixed and studied with scanning electron microscopy. No cellular or fibrin barrier could be found at the terminal pore of the catheter interfering with the diffusion of vancomycin from plasma into the catheter lumen. In the in-vitro experiments, three PermCaths filled with standard heparin solution were incubated for 48 h in 100 ml of plasma containing 20 microg/ml of vancomycin. Vancomycin concentration was measured in 0.3-0.5 ml solution aspirated from each port of the catheters. Vancomycin concentration was 0.2+/-0.1 microg/ml in the aspirated samples. Finally, two PermCaths filled with the standard heparin solution were incubated for 48 h in 100 ml of plasma containing 20 microg/ml of vancomycin, after which the catheters were sectioned at 4-cm intervals. Only the distal 4 cm of the catheters had vancomycin concentrations of 2 and 5 microg/ml, the remaining segments had levels 0.05). Geometric mean (interquartile range) homocysteine at baseline was 20.0 (16.8-24.5) and 19.5 (15.3-22.0) micromol/l for the high-and low-flux groups respectively (P=0.80), and levels did not change significantly during the study. We did demonstrate a more pronounced intradialytic effect of high-flux dialysis on homocysteine levels, which fell during dialysis by 42%, compared to 32% with low-flux dialysis (P<0. 001). CONCLUSIONS: In this randomized controlled trial, the effects of high-flux and low-flux haemodialysis on homocysteine and lipid profiles were comparable. The greater intradialytic effect of high-flux dialysis on homocysteine did not translate into a significant difference in pre-dialysis levels after 3 months of study. PMID- 10862644 TI - Ambulatory blood pressure monitoring in potential renal transplant donors. AB - BACKGROUND: Hypertension is considered to be a contraindication in potential renal transplant donors. Ambulatory blood pressure monitoring (ABPM) was developed as an alternative to in-office blood pressure measurement (OBPM). The aim of this study was to determine the sensitivity of ABPM in revealing hypertension in potential renal transplant donors, and to measure the correlation between ABPM results and target organ damage. METHODS: The study included 126 potential living-related renal transplant donors. The potential donors's blood pressures were measured during three separate clinic visits and then evaluated using 24-h ABPM. Cardiac and ophthalmological examinations were also performed to investigate target organ damage in all of the donors. RESULTS: According to the OBPM, 89 potential donors were normotensive and 37 had borderline or mild hypertension. Of the normotensive group, six were diagnosed as hypertensive after 24-h ABPM, and these subjects had target organ involvement. The status of the other 83 donors remained unchanged after ABPM and investigation for target organ damage. Thirteen of the 37 subjects who had borderline or mildly elevated pressures on OBPM were classified as normotensive after ABPM. These 13 individuals exhibited no hypertension-related target organ damage. The other 24 patients who had been classified as borderline or mildly hypertensive on OBPM fulfilled the criteria for hypertension after ABPM, and hypertensive changes were found at target organ evaluation. Before donor nephrectomy, 94 subjects who were classified as normotensive prior to transplantation underwent renal angiography for routine pretransplant evaluation, and none showed hypertensive or atherosclerotic changes. CONCLUSION: In our study, ABPM was found to be more sensitive than OBPM in terms of identifying hypertension in potential renal transplant donors. PMID- 10862645 TI - Withdrawal of steroids from triple-drug therapy in kidney transplant patients. AB - BACKGROUND: In renal transplant patients with stable graft function, triple-drug immunosuppression may not be necessary, while withdrawal of steroids may eliminate side effects. The primary aim of this study was to assess the risk of rejection after steroid withdrawal. METHODS: A total of 88 patients with stable graft function and serum creatinine <160 micromol/l, treated with cyclosporin A, azathioprine and prednisone were randomized into group A (n=46) with a gradual prednisone reduction to zero in the course of 6 months, and group B (n=42) on triple-drug therapy without change. At the time of randomization, fine-needle aspiration biopsy (FNAB) was carried out in all of the patients. After stopping steroids, the patients were followed up for a period of 12 months. RESULTS: Four patients failed to complete steroid withdrawal, three due to rejection, and one due to leukopenia. The proportion of rejection in three patients in group A (6.6%) was not significantly different from rejection in two patients in group B (4.8%). The mean value of serum creatinine was not significantly different in both groups in the course of follow-up. A finding of some degree of immunological activity in FNAB was made in four patients in each group, but none of these patients developed rejection. Compared with group B, significant decreases in serum cholesterol and blood leukocytes were observed in group A. Prednisone withdrawal did not have any influence on hypertension and serum triglycerides. CONCLUSIONS: Gradual withdrawal of steroids is not associated with a higher risk for rejection and has a beneficial effect on serum total cholesterol levels. FNAB was not a useful tool for predicting rejection. PMID- 10862646 TI - Expression of chemokines and their receptors in nephrotoxic serum nephritis. AB - BACKGROUND: Chemokines play a major role in leukocyte infiltration in inflammatory kidney diseases. The specificity of the chemokine action is determined by the restricted expression of the corresponding receptors on leukocytes. We therefore simultaneously studied the expression of CC-chemokine and CC-chemokine receptor 1-5 (CCR 1-5) mRNA in an accelerated model of nephrotoxic nephritis in CD-1 mice. METHODS: Kidneys were harvested at day 0, 2 and 7. Induction of nephritis was confirmed by assessment of albuminuria by ELISA and by histological evaluation. RNA was prepared from cortex and isolated glomeruli. RNase protection assays were performed to study the expression of chemokines, RNase protection assays as well as quantitative RT-PCR assays to study the expression of chemokine receptors. RESULTS: In the cortex of nephritic kidneys mRNA for MCP-1 was increased 5-fold on day 2 and increased 4-fold on day 7 as compared to controls. mRNA for RANTES was increased 5-fold on day 7 and mRNA for IP-10 6-fold on day 7. The increase of mRNA for the chemokine receptors CCR1 and 5 was between 2-fold and 3-fold determined by RNase protection assay and for CCR1, 2 and 5 between 2- and 4-fold as determined by RT-PCR. In isolated glomeruli we found by RT-PCR an increase of CCR1, CCR2 and CCR5 of between 3 and 12-fold. CONCLUSION: These results show that chemokines and their specific chemokine receptors are increased in parallel in this model of glomerulonephritis, consistent with the potential role of the chemokine system in leukocyte recruitment to the immune injured kidney. PMID- 10862647 TI - A severe systemic inflammatory reaction following therapy with montelukast (Singulair). PMID- 10862648 TI - Minimal change nephrotic syndrome associated with rifampicin treatment. PMID- 10862649 TI - Post-infectious glomerulonephritis in a patient with vesicorenal malacoplakia- coincidence or causal relationship? PMID- 10862650 TI - Reversibility of hepatorenal syndrome in an anuric patient with child C cirrhosis requiring haemodialysis for 7 weeks. PMID- 10862651 TI - Interaction between theophylline and tacrolimus in a renal transplant patient. PMID- 10862652 TI - Low-grade lymphoma in a cadaveric renal transplant donor following organ transplantation: recipient management and outcome. PMID- 10862653 TI - Premature closure or do not get lost in your diagnostic work-up and blame it on the patient. PMID- 10862654 TI - Acute renal failure and thrombocytopenia after ticlopidine--not necessarily thrombotic thrombocytopenic purpura. PMID- 10862655 TI - Images in Nephrology. Renal cortical nephrocalcinosis. PMID- 10862656 TI - Metabolic acidosis, hypokalaemia and acute renal failure with a normal urine output. PMID- 10862658 TI - Herve favre PMID- 10862657 TI - Practical haemodialysis began with cellophane and heparin: the crucial role of William Thalhimer (1884-1961). PMID- 10862659 TI - The influence of automated peritoneal dialysis on the decrease in residual renal function. PMID- 10862660 TI - Horseshoe kidney and Turner syndrome. PMID- 10862661 TI - Elution of glomerular bound C3 by glucosamine in a case of acute glomerulonephritis. PMID- 10862662 TI - Membranous glomerulonephritis with nephrotic syndrome in a HIV positive patient- remarkable remission with triple therapy. PMID- 10862663 TI - Colchicine in the treatment of renal amyloidosis secondary to familial Mediterranean fever. PMID- 10862664 TI - Missed double-J stent by ultrasonography. PMID- 10862665 TI - The short evaluation of the injured patients with acute renal failure who required dialysis and transferred to our centre during the Marmara earthquake. PMID- 10862666 TI - Retroperitoneal fibrosis of unknown origin. PMID- 10862667 TI - Haemodialysis for hyperammonaemic encephalopathy. PMID- 10862668 TI - Spontaneous remission of recurrent secondary hyperparathyroidism PMID- 10862669 TI - Indinavir pharmacokinetics in haemodialysis. PMID- 10862670 TI - Regression of post-transplant Kaposi sarcoma after discontinuing cyclosporin and giving mycophenolate mofetil instead. PMID- 10862672 TI - Announcements PMID- 10862671 TI - Subcapsular haematoma: a cause of post biopsy oliguria in renal allografts. PMID- 10862673 TI - Recombinant protein production driven by the tryptophan promoter is tightly controlled in ICONE 200, a new genetically engineered E. coli mutant. AB - Batch processes for recombinant gene expression in prokaryotic systems should optimally comprise a growth phase with minimal promoter activity followed by a short phase favoring expression. The strong promoter of the tryptophan operon (Ptrp) gives high-level expression of recombinant proteins in E. coli. The inefficiency to control basal expression before induction is however a major obstacle towards the use of Ptrp, especially in the case of toxic proteins. To circumvent this problem, a novel E. coli strain has been generated. This mutant, named ICONE 200 (Improved Cell for Over and Non-leaky Expression), underwent replacement of tnaA, the tryptophanase encoding gene, with the trpR gene encoding the aporepressor of Ptrp. Detailed analysis of ICONE 200 showed that tryptophan, in addition to its natural role of Ptrp co-repressor, was able to induce trpR through the tryptophan-inducible tryptophanase promoter/operator. Consequently, Ptrp-dependent expression was efficiently repressed in the presence of tryptophan and was turned on, as in wild-type E. coli, as soon as tryptophan was exhausted from the medium. ICONE 200 has the capacity to express a wide range of proteins including toxic proteins such as HIV-1 protease and poliovirus 2B protein. ICONE 200 is a new host carrying stable, targeted, and marker-free genetic modifications and a candidate for large-scale applications. PMID- 10862674 TI - Mechanochemical manipulation of hepatocyte aggregation can selectively induce or repress liver-specific function. AB - Controlled activation of hepatocyte aggregation is critical to three-dimensional (3D) multicellular morphogenesis during native regeneration of liver as well as tissue reconstruction therapies. In this work, we quantify the stimulatory effects of two model hepatotrophic activators, epidermal growth factor (EGF) and hepatocyte growth factor (HGF), on the aggregation kinetics and liver-specific function of hepatocytes cultured on organotypic substrates with differing mechanical resistivity. Substrate-specific morphogenesis of cultured hepatocytes is induced on a tissue basement membrane extract, Matrigel, formulated at two distinct levels of mechanical compliance (storage modulus G', at oscillatory shear rate 1 rad/s, was 34 Pa for basal Matrigel and 118 Pa for crosslinked Matrigel). Overall, we report that growth factor stimulation selectively promotes the kinetics of aggregation in the form of two-dimensional corded aggregates on basal Matrigel and three-dimensional spheroidal aggregates on crosslinked Matrigel. Our analysis also indicates that costimulation with EGF and HGF (20 ng/mL each) cooperatively maximizes the kinetics of aggregation in a substrate specific manner. In addition, we show that the role of growth factor stimulation on hepatocyte function is sensitively governed by the mechanical compliance of the substrate. In particular, on matrices with high compliance, costimulatory aggregation is shown to elicit a marked increase in albumin secretion rate, whereas on matrices with low compliance aggregation results in effective functional repression to basal, unstimulated levels. Thus, our studies highlight a novel interplay of physicochemical parameters of the culture microenvironment, leading to selective enhancement or repression of differentiated functions of hepatocytes, in concert with the activation of cellular morphogenesis. PMID- 10862675 TI - Hydrolytic activity in baker's yeast limits the yield of asymmetric 3-oxo ester reduction. AB - Microbial reductions of ketones hold great potential for the production of enantiopure alcohols, as long as highly selective redox enzymes are not interfered with by competing activities. During reduction of ethyl 3-oxobutanoate by baker's yeast (Saccharomyces cerevisiae) to ethyl (S)-3-hydroxybutanoate, a high enantiomeric excess (> 99%) can be obtained. However, reported yields do not exceed 50-70%. In this article, three main causes are shown to be responsible for these low to moderate yields. These are evaporation of the substrate and product esters, absorption or adsorption of the two esters by the yeast cells and hydrolysis of the two esters by yeast enzymes. The hydrolysis products are further metabolized by the yeast. By reducing the evaporation and absorption losses, the reduction yield can easily be improved to about 85%. Improvement of the efficiency of the reduction and hence the reduction/hydrolysis ratio should lead to a further increase in yield. PMID- 10862676 TI - Data model for the elimination of matrix effects in enzyme-based flow-injection systems. AB - This contribution presents a new conceptional enzyme-based flow injection analysis (FIA) system for the process and quality control of food processing and biotechnological systems. It provides the determination of different analytes in distinct process media on the base of a common experimental set-up. In contrast to known comparable systems, analysis is performed without the commonly used sample preparation and dilution steps. Instead, the adaptation to the necessary measurement range is realized by optimization of intrinsic system parameters. The central principle of the work presented is the elimination of occurring interferences by the heterogeneous matrix of the process sample. Based on a particular injection mode, the application of dehydrogenases as indicator enzymes and a specially developed data model using cognitive methods, cross sensitivities of the detector as well as disturbed reaction rates of the enzymes could be almost completely compensated. Two applications are presented, the analysis of ethanol in non-alcoholic beer and the online determination of D-/L-lactate during a lactic acid fermentation, which reveal the advantage of the developed system. PMID- 10862677 TI - Biodegradation kinetics of benzene, toluene, and phenol as single and mixed substrates for Pseudomonas putida F1. AB - Although microbial growth on substrate mixtures is commonly encountered in bioremediation, wastewater treatment, and fermentation, mathematical modeling of mixed substrate kinetics has been limited. We report the kinetics of Pseudomonas putida F1 growing on benzene, toluene, phenol, and their mixtures, and compare mathematical models to describe these results. The three aromatics are each able to act as carbon and energy sources for this strain. Biodegradation rates were measured in batch cultivations following a protocol that eliminated mass transfer limitations for the volatile substrates and considered the culture history of the inoculum and the initial substrate to inoculum mass ratio. Toluene and benzene were better growth substrates than phenol, resulting in faster growth and higher yield coefficients. In the concentration ranges tested, toluene and benzene biodegradation kinetics were well described by the Monod model. The Monod model was also used to characterize phenol biodegradation by P. putida F1, although a small degree of substrate inhibition was noted. In mixture experiments, the rate of consumption of one substrate was found to be affected by the presence of the others, although the degree of influence varied widely. The substrates are catabolized by the same enzymatic pathway, but purely competitive enzyme kinetics did not capture the substrate interactions well. Toluene significantly inhibited the biodegradation rate of both of the other substrates, and benzene slowed the consumption of phenol (but not of toluene). Phenol had little effect on the biodegradation of either toluene or benzene. Of the models tested, a sum kinetics with interaction parameters (SKIP) model provided the best description of the paired substrate results. This model, with parameters determined from one- and two-substrate experiments, provided an excellent prediction of the biodegradation kinetics for the three-component mixture. PMID- 10862678 TI - A two-reservoir, hollow-fiber bioreactor for the study of mixed-population dynamics: design aspects and validation of the approach. AB - A two-reservoir, membrane bioreactor for carrying out studies of mixed-population dynamics in batch fermentations is presented. Mixing requirements and design aspects for the validity of the approach are given and discussed. Equations describing mixing times between the reservoirs are presented and compared to the experimental results. The validity of the approach is demonstrated by the study of an amensalistic-type interaction, the protein-mediated killer phenomenon between two Saccharomyces cerevisiae strains. The validation consisted in the comparison between the results obtained in actual mixed culture and the results obtained by keeping the strains separated. A good agreement was found which demonstrates the viability of the designed bioreactor. PMID- 10862679 TI - Microbial dynamics in a continuous stirred tank bioreactor exposed to an alternating sequence of organic compounds. AB - Microbial dynamics during aerobic biodegradation of an alternating mixture of organic compounds was investigated experimentally in a continuous stirred tank bioreactor (CSTB). A mathematical model describing this system was developed and tested using the experimental results. A model microbial culture consisting of Pseudomonas sp. JS150, a monochlorobenzene (MCB) degrader, and Xanthobacter autotrophicus GJ10, a 1,2-dichloroethane (DCE) degrader, each with exclusive degradation capabilities, was used. The CSTB was inoculated with both microbial strains and exposed to an alternating sequence of the two compounds at noninhibitory concentrations. Concentrations of each microbial strain, of each organic compound, and of degradation product evolved, as well as specific microbial activities via oxygen uptake tests, were monitored. Reduction of the residual DCE discharged from the bioreactor after an MCB to DCE transition was successfully achieved by continuously feeding a low flow of a concentrated solution of both compounds. PMID- 10862680 TI - Glucagon-induced self-association of recombinant proteins in Escherichia coli and affinity purification using a fragment of glucagon receptor. AB - The specific molecular interactions of alpha-helical peptide, human glucagon (i.e., intermolecular self-association and specific receptor-binding affinity) provided a rationale for using the glucagon as the fusion expression partner to achieve high productivity of foreign proteins both in vivo (in bacterial fusion expression system) and in vitro (in affinity column chromatography). The fusion of glucagon peptide(s) effectively promoted homogeneous aggregate formation of recombinant proteins while avoiding intermolecular crosslinking by disulfide bridges. High sensitivity of the self-aggregation to sequence effects resulted from two distinct nonpolar domains of glucagon, determining specificity of molecular interaction and aggregate size of recombinant proteins. An N-terminal domain of glucagon molecule (Phe6-Tyr10-Tyr13) could be a certain hydrophobic moiety involved in intermolecular self-association (probably, via helix-helix docking), while a C-terminal domain (Phe22-Trp25-Leu26) seems to critically affect the oligomer size in the off-pathway aggregation of synthesized fusion proteins. An N-terminal extracellular domain of human glucagon receptor was recombinantly expressed in Escherichia coli, immobilized to a chromatography column, and efficiently renatured to a conformation that attains high specificity in interaction with N-terminus glucagon molecules of recombinant fusion proteins. Through column chromatography employing the receptor fragment as affinity ligand, the recombinant proteins were efficiently purified from total intracellular proteins, and the long-term ligand stability was evidently proven through multiple cyclic-purification experiments. Major scaffolds for using protein ligands are large-scale production in a low-cost expression system and long-term stable operation with selective-binding affinity. From this point of view, the extracellular fragment of human glucagon receptor used in this study seems to be a new potent ligand for fusion protein-based affinity chromatography. PMID- 10862681 TI - Studies on the interaction of fermentation and microfiltration operations: erythromycin recovery from Saccharopolyspora erythraea fermentation broths. AB - Changes in fermentation media not only affect the performance of the fermentation itself (with regard to the kinetics of biomass and product formation and the yields obtained) but also the initial product-recovery operations downstream of the fermentor. In this work, microfiltration experiments to remove Saccharopolyspora erythraea biomass from fermentation broth and to recover erythromycin were carried out using two fundamentally different media; a soluble complex medium (SCM) and an oil-based process medium (OBM). Small-scale batch fermentations of 14-L working volume were carried out in triplicate using both media. Broth samples were taken from each fermentation at regular intervals from the end of the exponential-growth phase onwards. These were then processed using a Minitan II (acrylic), tangential crossflow-filtration module, fitted with a single 60 cm(2) Durapore hydrophilic 0.2 microm membrane, operated in concentration mode. The OBM fermentations produced higher titers of erythromycin but required longer fermentation times due to increased lag phases and slower maximum-growth rates. The OBM also increased the loading on the membrane; at maximum product titers residual oil concentrations of 3 g. L(-1), antifoam concentrations of 2 g. L(-1) and flour concentrations estimated at approximately 10 g/L(-1) were typical. It was found that both the permeate flux and erythromycin transmission were affected by the choice of medium. The OBM had significantly lower values for both parameters (12.8 Lm(-2) h(-1) and 89.6% respectively) than the SCM (35.9 Lm(-2) h(-1) and 96.7% respectively) when the fermentations were harvested at maximum erythromycin titers. Transmission of erythromycin stayed approximately constant as a function of fermentation time for both media, however, for the OBM the permeate flux decreased with time which correlated with an increase in broth viscosity. The relatively poor microfiltration performance of the OBM medium was, however, offset by the higher titers of erythromycin that were achieved during the fermentation. The filtration characteristics of the SCM broth did not show any correlation with either broth viscosity or fermentation time. Image-analysis data suggested that there was a correlation between hyphal morphology (main hyphal length) and permeate flux (no such correlation was found for the OBM broth). Moreover, it has been shown for the OBM broth that the residual flour had a profound effect on the microfiltration characteristics. The influence of the residual flour was greater than that imposed by the morphology and concentration of the biomass. The understanding of the factors governing the interaction of the fermentation and microfiltration operations obtained in this work provides a first step towards optimization of the overall process sequence. PMID- 10862682 TI - Performance of small-scale CHO perfusion cultures using an acoustic cell filtration device for cell retention: characterization of separation efficiency and impact of perfusion on product quality. AB - Several small-scale Chinese hamster ovary (CHO) suspension cultures were grown in perfusion mode using a new acoustic filtration system. The separation performance was evaluated at different cell concentrations and perfusion rates for two different CHO cell lines. It was found that the separation performance depends inversely on the cell concentration and perfusion rate. High media flow rates as well as high cell concentrations resulted in a significant drop in the separation performance, which limited the maximal cell concentration achievable. However, packed cell volumes of 10% to 16% (corresponding to 3 to 6. 10(7) cells/mL) could be reached and were maintained without additional bleeding after shifting the temperature to 33 degrees C. Perfusion, up to 50 days, did not harm the cells and did not result in a loss of performance of the acoustic filter as often seen with other perfusion systems. Volumetric productivities in perfusion mode were 2- to 12-fold higher for two cell lines producing two different glycoproteins when compared to fed-batch or batch processes using the same cell lines. Product concentrations were in the range of 20% to 80% of batch or fed-batch culture, respectively. In addition, using the protease-sensitive product rhesus thrombopoietin, we could show that cultivation in perfusion mode drastically reduced proteolysis when compared to a batch culture without addition of protease inhibitors such as leupeptin. PMID- 10862683 TI - Biotechnology for the production of nutraceuticals enriched in conjugated linoleic acid: II. Multiresponse kinetics of the hydrolysis of corn oil by a Pseudomonas sp. lipase immobilized in a hollow-fiber reactor. AB - The hydrolysis of corn oil in the presence of a lipase from Pseudomonas sp. immobilized within the walls of a hollow-fiber reactor was studied at 30 degrees C. To assess the selectivity of this immobilized enzyme, the effluent concentrations of five different free fatty acids were measured using high performance liquid chromatography (HPLC). Several rate expressions associated with a generic ping-pong bi-bi mechanism were used to fit the experimental data for this lipase-catalyzed reaction. A multiresponse nonlinear regression method was employed to determine the kinetic parameters associated with these rate expressions. Quasi-optimum operating conditions corresponded to 30 degrees C and a buffer pH value of 7.0. Under these conditions, the concentration of free linoleic acid (C18:2) (the fatty acid of primary interest) in the effluent oil stream for a fluid residence time of 6 h was approximately 0.5 M. PMID- 10862684 TI - Chemoenzymatic construction of a four-component Ugi combinatorial library. AB - The chemoenzymatic preparation of a nine-member Ugi condensation library is described. The carboxylic acid and amine precursors are based on 3 hydroxybutyrate and 4-amino-1-butanol, respectively, and have been acylated selectively using a variety of acyl donors catalyzed by porcine pancreatic lipase. The enzyme is selective for the hydroxyl functionalities on both precursors, thereby yielding 3-acyl-butyric acid and 4-amino-1-acyl compounds. These enzymatically generated derivatives were then subjected to a four-component Ugi condensation reaction in the presence of acetaldehyde and methyl isocyanoacetate. Isolated yields of the alpha-(acylamino)amide Ugi products ranged from 72-95%. The inherent chemoselectivity of enzymatic catalysis may play an increasingly important role in expanding the structural diversity that can be achieved by chemical multicomponent condensation reactions. PMID- 10862685 TI - A novel approach to the recovery of biologically active oligosaccharides from milk using a combination of enzymatic treatment and nanofiltration. AB - A new easily scalable approach to the recovery of biologically active oligosaccharides from milk has been developed which relies on the combination of enzymatic treatment of defatted milk using beta-galactosidase and nanofiltration. It was shown that enzymatic hydrolysis of lactose significantly improves the efficiency and selectivity of membrane-based separations. With the best membrane, as much as 6.7 g of oligosaccharides (containing very little contaminating lactose) could be obtained from one liter of defatted human milk in just four nanofiltration cycles. The human milk oligosaccharides recovered by this method were shown to inhibit binding of intimin, an adhesion molecule of enteropathogenic Escherichia coli, to epithelial cells in vitro. No significant difference in the oligosaccharide profile between samples prepared by this method and conventional gel-permeation chromatography was found. The developed approach is also suitable for the recovery of substantial quantities of tri- and tetra saccharides from caprine milk. PMID- 10862686 TI - Making the most of peer review. PMID- 10862687 TI - Spikes versus BOLD: what does neuroimaging tell us about neuronal activity? PMID- 10862688 TI - Growth of the NMDA receptor industrial complex. PMID- 10862689 TI - What goes out must come in. PMID- 10862690 TI - Regulating olfactory receptor expression: controlling globally, acting locally. PMID- 10862692 TI - A new high for alternative splicing. PMID- 10862691 TI - Proto-mapping the areas of cerebral cortex: transcription factors make the grade. PMID- 10862693 TI - What is it like to be an animal? PMID- 10862695 TI - Why you should look where you are going. PMID- 10862694 TI - Alternative role for prolactin-releasing peptide in the regulation of food intake. AB - Prolactin-releasing peptide (PrRP) is a peptide ligand for the human orphan G protein-coupled receptor hGR3/GPR10 and causes the secretion of prolactin from anterior pituitary cells. However, the lack of immunoreactive staining for PrRP in the external layer of the median eminence seems to rule out this peptide as a classical hypophysiotropic hormone and, furthermore, PrRP is less effective than another inducer of prolactin secretion, thyrotropin-releasing hormone, both in vitro and in vivo. Here we show a reduction in the expression of PrRP mRNA during lactation and fasting and an acute effect of PrRP on food intake and body weight, supporting the hypothesis of an alternative role for the peptide. PMID- 10862696 TI - The potential effectiveness of simulations versus phenomenological models. PMID- 10862697 TI - From form to function: calcium compartmentalization in dendritic spines. AB - Dendritic spines compartmentalize calcium, and this could be their main function. We review experimental work on spine calcium dynamics. Calcium influx into spines is mediated by calcium channels and by NMDA and AMPA receptors and is followed by fast diffusional equilibration within the spine head. Calcium decay kinetics are controlled by slower diffusion through the spine neck and by spine calcium pumps. Calcium release occurs in spines, although its role is controversial. Finally, the endogenous calcium buffers in spines remain unknown. Thus, spines are calcium compartments because of their morphologies and local influx and extrusion mechanisms. These studies highlight the richness and heterogeneity of pathways that regulate calcium accumulations in spines and the close relationship between the morphology and function of the spine. PMID- 10862698 TI - Proteomic analysis of NMDA receptor-adhesion protein signaling complexes. AB - N-methyl-d-aspartate receptors (NMDAR) mediate long-lasting changes in synapse strength via downstream signaling pathways. We report proteomic characterization with mass spectrometry and immunoblotting of NMDAR multiprotein complexes (NRC) isolated from mouse brain. The NRC comprised 77 proteins organized into receptor, adaptor, signaling, cytoskeletal and novel proteins, of which 30 are implicated from binding studies and another 19 participate in NMDAR signaling. NMDAR and metabotropic glutamate receptor subtypes were linked to cadherins and L1 cell adhesion molecules in complexes lacking AMPA receptors. These neurotransmitter adhesion receptor complexes were bound to kinases, phosphatases, GTPase activating proteins and Ras with effectors including MAPK pathway components. Several proteins were encoded by activity-dependent genes. Genetic or pharmacological interference with 15 NRC proteins impairs learning and with 22 proteins alters synaptic plasticity in rodents. Mutations in three human genes (NF1, Rsk-2, L1) are associated with learning impairments, indicating the NRC also participates in human cognition. PMID- 10862699 TI - Calcium channel gating and modulation by transmitters depend on cellular compartmentalization. AB - Voltage-gated Ca2+ channels participate in dendritic integration, yet functional properties of Ca2+ channels and mechanisms of their modulation by neurotransmitters in dendrites are unknown. Here we report how pharmacologically identified Ca2+ channels behave in different neural compartments. Whole-cell and cell-attached patch-clamp recordings were made on both cell bodies and electrically isolated dendrites of sympathetic neurons. We found not only that Ca2+ channel populations differentially contribute to somatic and dendritic currents but also that families of Ca2+ channels display gating properties and neurotransmitter modulation that depend on channel compartmentalization. By comparison with their somatic counterparts, dendritic N-type Ca2+ currents were hypersensitive to neurotransmitters and G proteins. Single-channel analysis showed that dendrites express a unique N-type channel that has enhanced interaction with Gbetagamma. Thus Ca2+ channels in dendrites seem to be specialized elements with unique regulatory mechanisms. PMID- 10862700 TI - Area identity shifts in the early cerebral cortex of Emx2-/- mutant mice. AB - The specification of area identities in the cerebral cortex is a complex process, primed by intrinsic cortical cues and refined after the arrival of afferent fibers from the thalamus. Little is known about the genetic control of the early steps of this process, but the distinctive expression pattern of the homeogene Emx2 in the developing cortex has prompted suggestions that it is critical in this context. We tested this hypothesis using Emx2 -/- mice. We found that the normal spectrum of cortical areal identities was encoded in these mutants, but areas with caudal-medial identities were reduced and those with anterior-lateral identities were relatively expanded in the cortex. PMID- 10862701 TI - Mutually exclusive expression of odorant receptor transgenes. AB - To study the mutually exclusive expression of odorant receptor (OR) genes, we generated transgenic mice that carried the murine OR gene MOR28. Expression of the transgene and the endogenous MOR28 was distinguished by using two different markers, beta-galactosidase and green fluorescent protein (GFP), respectively. Double staining of the olfactory epithelium revealed that the two genes were rarely expressed simultaneously in individual olfactory neurons. A similar exclusion was also observed between differently tagged but identical transgenes integrated into the same locus of one particular chromosome. Although allelic inactivation has been reported for the choice between the maternal and paternal alleles, this is the first demonstration of mutually exclusive activation among non-allelic OR gene members with identical coding and regulatory sequences. Such an unusual mode of gene expression, monoallelic and mutually exclusive, has previously been shown only for the antigen-receptor genes of the immune system. PMID- 10862702 TI - Competition at silent synapses in reinnervated skeletal muscle. AB - Synaptic connections are made and broken in an activity-dependent manner in diverse regions of the nervous system. However, whether activity is strictly necessary for synapse elimination has not been resolved directly. Here we report that synaptic terminals occupying motor endplates made electrically silent by tetrodotoxin and alpha-bungarotoxin block were frequently displaced by regenerating axons that were also both inactive and synaptically ineffective. Thus, neither evoked nor spontaneous activation of acetylcholine receptors is required for competitive reoccupation of neuromuscular synaptic sites by regenerating motor axons. PMID- 10862703 TI - Laminar sources of synaptic input to cortical inhibitory interneurons and pyramidal neurons. AB - The functional role of an individual neuron within a cortical circuit is largely determined by that neuron's synaptic input. We examined the laminar sources of local input to subtypes of cortical neurons in layer 2/3 of rat visual cortex using laser scanning photostimulation. We identified three distinct laminar patterns of excitatory input that correspond to physiological and morphological subtypes of neurons. Fast-spiking inhibitory basket cells and excitatory pyramidal neurons received strong excitatory input from middle cortical layers. In contrast, adapting inhibitory interneurons received their strongest excitatory input either from deep layers or laterally from within layer 2/3. Thus, differential laminar sources of excitatory inputs contribute to the functional diversity of cortical inhibitory interneurons. PMID- 10862704 TI - Visual input induces long-term potentiation of developing retinotectal synapses. AB - Early visual experience is essential in the refinement of developing neural connections. In vivo whole-cell recording from the tectum of Xenopus tadpoles showed that repetitive dimming-light stimulation applied to the contralateral eye resulted in persistent enhancement of glutamatergic inputs, but not GABAergic or glycinergic inputs, on tectal neurons. This enhancement can be attributed to potentiation of retinotectal synapses. It required spiking of postsynaptic tectal cells as well as activation of NMDA receptors, and effectively occluded long-term potentiation (LTP) of retinotectal synapses induced by direct electrical stimulation of retinal ganglion cells. Thus, LTP-like synaptic modification can be induced by natural visual inputs and may be part of the underlying mechanism for the activity-dependent refinement of developing connections. PMID- 10862705 TI - A direct quantitative relationship between the functional properties of human and macaque V5. AB - The nature of the quantitative relationship between single-neuron recordings in monkeys and functional magnetic resonance imaging (fMRI) measurements in humans is crucial to understanding how experiments in these different species are related, yet it remains undetermined. We measured brain activity in humans attending to moving visual stimuli, using blood oxygenation level-dependent (BOLD) fMRI. Responses in V5 showed a strong and highly linear dependence on increasing strength of motion signal (coherence). These population responses in human V5 had a remarkably simple mathematical relationship to previously observed single-cell responses in macaque V5. We provided an explicit quantitative estimate for the interspecies comparison of single-neuron activity and BOLD population responses. Our data show previously unknown dissociations between the functional properties of human V5 and other human motion-sensitive areas, thus predicting similar dissociations for the properties of single neurons in homologous areas of macaque cortex. PMID- 10862706 TI - Surround modulation in human vision unmasked by masking experiments. AB - The responses of neurons in cat and monkey primary visual cortex are modulated by stimuli outside the classical receptive field. Here we report psychophysical evidence from masking experiments for two distinct types of surround modulation, one narrowly tuned to iso-orientation (stimuli with center and surround at the same orientation) and the other broadly tuned to cross-orientation (center and surround at perpendicular orientations). Surround modulation at iso- and cross orientations showed distinct contrast dependencies, and high-contrast cross oriented surrounds were able to completely eliminate masking. Surround modulation was modeled by subtracting divisive inhibition that raised the gain of spatial filters. PMID- 10862707 TI - An 'automatic pilot' for the hand in human posterior parietal cortex: toward reinterpreting optic ataxia. AB - We designed a protocol distinguishing between automatic and intentional motor reactions to changes in target location triggered at movement onset. In response to target jumps, but not to a similar change cued by a color switch, normal subjects often could not avoid automatically correcting fast aiming movements. This suggests that an 'automatic pilot' relying on spatial vision drives fast corrective arm movements that can escape intentional control. In a patient with a bilateral posterior parietal cortex (PPC) lesion, motor corrections could only be slow and deliberate. We propose that 'on-line' control is the most specific function of the PPC and that optic ataxia could result from a disruption of automatic hand guidance. PMID- 10862708 TI - Independence of perceptual and sensorimotor predictions in the size-weight illusion. AB - The smaller of two equally weighted objects is judged to be heavier when lifted. Here we disproved a leading hypothesis that this size-weight illusion is caused by a mismatch between predicted and actual sensory feedback. We showed that when subjects repeatedly lifted equally heavy large and small objects in alternation, they learned to scale their fingertip forces precisely for the true object weights and thus exhibited accurate sensorimotor prediction. The size-weight illusion nevertheless persisted, suggesting that the illusion can be caused by high-level cognitive and perceptual factors and indicating that the sensorimotor system can operate independently of the cognitive/perceptual system. PMID- 10862709 TI - AAA domains and organization of the dynein motor unit. AB - Dyneins contain one-three microtubule motor units that are each derived from the C-terminal globular head of a heavy chain. The N-terminal regions of the heavy chains form stems that are required for intra-dynein associations. The microtubule-binding sites are located at the terminus of a short stalk that emanates from each globular head. Recent electron microscopic analysis indicates that the dynein head has a heptameric toroidal organization. This finding is echoed by the identification of six AAA (ATPases associated with cellular activities) domains and a seventh unrelated unit within this heavy chain region. At least two of these AAA domains can bind nucleotide, although only one appears able to hydrolyze ATP. Several other AAA domain proteins exhibit a similar annular organization of six AAA units. Detailed structural information is available for several AAA proteins, including N-ethylmaleimide-sensitive vesicle fusion protein and the RuvB motor involved in DNA migration and resolution of Holliday junctions. The resulting structural parallels allow intriguing predictions to be made concerning dynein organization and motor function. PMID- 10862710 TI - Intranuclear trafficking of transcription factors: implications for biological control. AB - The subnuclear organization of nucleic acids and cognate regulatory factors suggests that there are functional interrelationships between nuclear structure and gene expression. Nuclear proteins that are localized in discrete domains within the nucleus include the leukemia-associated acute myelogenous leukemia (AML) and promyelocytic leukemia (PML) factors, the SC-35 RNA-processing factors, nucleolar proteins and components of both transcriptional and DNA replication complexes. Mechanisms that control the spatial distribution of transcription factors within the three-dimensional context of the nucleus may involve the sorting of regulatory information, as well as contribute to the assembly and activity of sites that support gene expression. Molecular, cellular, genetic and biochemical approaches have identified distinct protein segments, termed intranuclear-targeting signals, that are responsible for directing regulatory factors to specific subnuclear sites. Gene rearrangements that remove or alter intranuclear-targeting signals are prevalent in leukemias and have been linked to altered localization of regulatory factors within the nucleus. These modifications in the intranuclear targeting of transcription factors might abrogate fidelity of gene expression in tumor cells by influencing the spatial organization and/or assembly of machineries involved in the synthesis and processing of gene transcripts. PMID- 10862711 TI - Differential requirement for individual sarcoglycans and dystrophin in the assembly and function of the dystrophin-glycoprotein complex. AB - Sarcoglycan is a multimeric, integral membrane glycoprotein complex that associates with dystrophin. Mutations in individual sarcoglycan subunits have been identified in inherited forms of muscular dystrophy. To evaluate the contributions of sarcoglycan and dystrophin to muscle membrane stability and muscular dystrophy, we compared muscle lacking specific sarcoglycans or dystrophin. Here we report that mice lacking (delta)-sarcoglycan developed muscular dystrophy and cardiomyopathy similar to mice lacking (gamma) sarcoglycan. However, unlike muscle lacking (gamma)-sarcoglycan, (delta) sarcoglycan-deficient muscle was sensitive to eccentric contraction-induced disruption of the plasma membrane. In the absence of (delta)-sarcoglycan, (alpha) , (beta)- and (gamma)-sarcoglycan were undetectable, while dystrophin was expressed at normal levels. In contrast, without (gamma)-sarcoglycan, reduced levels of (alpha)-, (beta)- and (delta)-sarcoglycan were expressed, glycosylated and formed a complex with each other. Thus, the elimination of (gamma)- and (delta)-sarcoglycan had different molecular consequences for the assembly and function of the dystrophin-glycoprotein complex. Furthermore, these molecular differences were associated with different mechanical consequences for the muscle plasma membrane. Through this in vivo analysis, a model for sarcoglycan assembly is proposed. PMID- 10862712 TI - Role of the ribosome in sequence-specific regulation of membrane targeting and translocation of P-glycoprotein signal-anchor transmembrane segments. AB - It is thought that the topology of a polytopic protein is generated by sequential translocation and membrane integration of independent signal-anchor and stop transfer sequences. Two well-characterized cell-free systems (rabbit reticulocyte lysate and wheat germ extract) have been widely used to study the biogenesis of secretory and membrane proteins, but different results have been observed with proteins expressed in these two different systems. For example, different topologies of P-glycoprotein (Pgp) were observed in the two systems and the cause was thought to be the source of ribosomes. To understand how the ribosome is involved in dictating membrane translocation and orientation of polytopic proteins, individual signal-anchor sequences of Pgp were dissected and examined for their membrane targeting and translocation in a combined system of wheat germ ribosomes (WGR) and rabbit reticulocyte lysate (RRL). Addition of wheat germ ribosomes to the rabbit reticulocyte lysate translation system can enhance, reduce, or have no effect on the membrane targeting and translocation of individual Pgp signal-anchor sequences, and these effects appear to be determined by the amino acid residues flanking each signal-anchor. Ribosomes regulate the membrane targeting and translocation of Pgp signal-anchors in a polytopic form differently from the same signal-anchors in isolation. Furthermore, we demonstrated that ribosomes regulate the membrane targeting and translocation of each signal-anchor cotranslationally and that this activity of ribosomes is associated with the 60S subunit. Based on this and previous studies, we propose a mechanism by which ribosomes dynamically dictate the membrane targeting and translocation of nascent polytopic membrane proteins. PMID- 10862713 TI - Syk-dependent phosphorylation of microtubules in activated B-lymphocytes. AB - Syk is a protein-tyrosine kinase that is essential for B-lymphocyte development and B-cell signaling. Syk phosphorylates tubulin on tyrosine both in vitro and in intact lymphocytes. Here we show that (alpha)-tubulin present within the cytoskeletal microtubule network was phosphorylated in a Syk-dependent manner following the activation of B-cells by engagement of the B-cell antigen receptor or by treatment with the phosphotyrosine phosphatase inhibitor, pervanadate. Immunofluorescence staining of microtubule cytoskeletons and western blotting studies with antibodies to phosphotyrosine confirmed the phosphorylation of polymerized tubulin in Syk-expressing, but not Syk-deficient, cells. At low concentrations of pervanadate, centrosomes appeared to be preferentially tyrosine phosphorylated. Tubulin phosphorylated to a high stoichiometry on tyrosine assembled into microtubules in vitro, and preassembled microtubules were also phosphorylated by Syk kinase in vitro. Thus, Syk has the capacity to interact with microtubule networks within the B-lymphocyte and catalyzes the phosphorylation of the (alpha)-tubulin subunit. Syk-dependent phosphorylation of microtubules may affect the ability of the microtubule cytoskeleton to serve as a platform upon which signaling complexes are assembled. PMID- 10862714 TI - The role of retinoic acid receptors in neurite outgrowth from different populations of embryonic mouse dorsal root ganglia. AB - Dorsal root ganglion (DRG) neurons can be categorised into at least three types, based upon their neurotrophin requirement for survival. We have analysed the expression of the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs) in NGF, NT-3 and BDNF dependent neurons isolated from embryonic day (E)13.5 mouse DRG. We show that each population of neurons expressed each of the three RXRs, (alpha), (beta) and (gamma); however, whilst the NGF and NT-3 dependent neurons expressed each of the RARs (alpha), (beta) and (gamma), the BDNF dependent neurons only expressed RAR(alpha) and (beta). When retinoic acid was added to each of the neuronal classes only the NGF and NT-3 dependent neurons responded by extending neurites, and this response involved the upregulation of RAR(beta)(2). This specificity was confirmed by the use of receptor-selective agonists as only a RAR(beta)-selective compound stimulated neurite outgrowth. These results suggest a role for RA acting via RAR(beta)(2) in the outgrowth of neurites. PMID- 10862715 TI - Protein kinase C(alpha) actively downregulates through caveolae-dependent traffic to an endosomal compartment. AB - Receptor desensitization occurs through receptor internalization and targeting to endosomes, a prerequisite for sorting and degradation. Such trafficking processes may not be restricted to membrane associated receptors but may also play an important role in the downregulation of cytoplasmic transducers such as protein kinase C (PKC). It is demonstrated here that acute TPA exposure induces the transport of activated PKC(alpha) from the plasma membrane to endosomes. This process requires PKC activity and catalytically competent PKC can even promote a similar process for a truncated regulatory domain PKC(&agr;) protein. It is established that PKC(&agr;) is targeted to the endosome compartment as an active kinase, where it colocalizes with annexin I, a substrate of PKC. Thus, PKC(alpha) downregulation shares features with plasma membrane associated receptor sorting and degradation. However, it is shown that PKC(&agr;) delivery to the endosome compartment is not a Rab5 mediated process in contrast to the well characterised internalisation of the transferrin receptor. An alternative route for PKC(alpha) is evidenced by the finding that the cholesterol binding drugs nystatin and filipin, known to inhibit caveolae mediated trafficking, are able to block PKC(alpha) traffic and down regulation. Consistent with this, the endosomes where PKC(alpha) is found also contain caveolin. It is concluded that the initial step in desensitisation of PKC(alpha) involves active delivery to endosomes via a caveolae mediated process. PMID- 10862716 TI - High resolution analysis of interphase chromosome domains. AB - Chromosome territories need to be well defined at high resolution before functional aspects of chromosome organization in interphase can be explored. To visualize chromosomes by electron microscopy (EM), the DNA of Chinese hamster fibroblasts was labeled in vivo with thymidine analogue BrdU. Labeled chromosomes were then segregated during several cell cycles to obtain nuclei containing only 2 to 3 labeled chromosomes. Subsequent immunocytochemical detection of BrdU allowed analysis by EM of chromosome territories and subchromosomal domains in well preserved nuclei. Our results provide the first high resolution visualization of chromosomes in interphase nuclei. We show that chromosome domains are either separated from one another by interchromatin space or are in close contact with no or little intermingling of their DNA. This demonstrates that, while chromosomes form discrete territories, chromatin of adjacent chromosomes may be in contact in limited regions, thus implying chromosome chromosome interactions. Chromosomes are organized as condensed chromatin with dispersed chromatin extending into the interchromatin space that is largely devoid of DNA. The interchromatin space, which is known to be involved in various nuclear functions, forms interconnecting channels running through and around chromosome territories. Functional implications of this organization are discussed. PMID- 10862717 TI - Identification and molecular-genetic characterization of a LAMP/CD68-like protein from Caenorhabditis elegans. AB - Lysosome associated membrane proteins (LAMPs) constitute a family of vertebrate proteins located predominantly in lysosomes, with lesser amounts present in endosomes and at the cell surface. Macrosialin/CD68s are similar to LAMPs in their subcellular distribution and amino acid sequence and presumed structure across the carboxyl terminal two thirds of their length. The functions of LAMPs and CD68s are not known. In the present study, a bioinformatics approach was used to identify a Caenorhabditis elegans protein (LMP-1) with sequence and presumed structural similarity to LAMPs and CD68s. LMP-1 appears to be the only membrane protein in C. elegans that carries a GYXX(phi) vertebrate lysosomal targeting sequence at its C terminus (where (phi) is a large, hydrophobic residue). LMP-1 was found to be present from early embryonic stages through adulthood and to be predominantly localized at the periphery of a population of large, membrane-bound organelles, called granules, that are seen throughout the early embryo but in later stages are restricted to the cells of the intestine. Analysis of an LMP-1 deficient C. elegans mutant revealed that LMP-1 is not required for viability under laboratory conditions, but the absence of LMP-1 leads to an alteration in intestinal granule populations, with apparent loss of one type of granule. PMID- 10862718 TI - Different properties of two isoforms of annexin XIII in MDCK cells. AB - Annexins form a family of proteins that are widely expressed and known to bind membranes in the presence of calcium. Two isoforms of the annexin XIII subfamily are expressed in epithelia. We previously reported that annexin XIIIb is apically localized in MDCK cells and that it is involved in raft-mediated delivery of apical proteins. We have now analyzed the properties of annexin XIIIa, which differs from annexin XIIIb by a deletion of 41 amino acids in the amino-terminal domain, and is distributed both apically and basolaterally. Annexin XIIIa binding to membranes is independent of calcium but requires its myristoyl amino-terminal modification, as observed with annexin XIIIb. Our biochemical and functional data show that annexin XIIIa behaves differently in the apical and in the basolateral compartments. Whereas annexin XIIIa apically can associate with rafts independently of calcium, the basolateral pool requires calcium for this. Annexin XIIIa, like annexin XIIIb, stimulates apical transport of influenza virus hemagglutinin but, in contrast, only annexin XIIIa inhibits basolateral transport of vesicular stomatitis virus G protein. Our results suggest that annexin XIIIa and XIIIb have specific roles in epithelial cells, and because of their structural similarities, these isoforms offer interesting tools for unravelling the functions of annexins. PMID- 10862719 TI - CD44 expression and regulation during mammary gland development and function. AB - The CD44v6 epitope has been widely reported to be expressed in human mammary carcinomas, yet its prognostic significance is controversial and its function in mammary tumors and mammary glands is unknown. To begin to resolve these issues, we analysed in detail the normal postnatal expression patterns and regulation of the CD44v6 epitope in murine mammary glands. We demonstrate that significant CD44v6 epitope expression is first seen during puberty, and that after puberty CD44v6 epitope expression follows the estrous cycle. CD44v6 epitope expression is observed in the myoepithelium and also less widely in luminal epithelial cells. During lactation, CD44v6 epitope expression is turned off and reappears during involution. The CD44 variant isoform bearing the v6 epitope is CD44v1-v10. Using HC11, a mammary epithelial cell line with stem cell characteristics, and facilitated by the cloning of the murine CD44 promoter, we show that growth factors and hormones which regulate ductal growth and differentiation modulate CD44 transcription. Together our data suggest that the CD44v6 epitope is expressed in mammary epithelial stems cells and in lineages derived from these cells, and that CD44v6 expression is regulated in part by hormones and growth factors such as IGF-1 and EGF which regulate the growth and differentiation of the mammary epithelium. The function of these same growth factors and hormones is often perturbed in mammary carcinomas, and we suggest that CD44v6 expression in tumors reflects this perturbation. We conclude that the expression of the CD44v6 epitope observed in some mammary tumors reflects the stem cell origin of breast tumors, and that whether or not the CD44v6 epitope is expressed in a mammary tumor is determined by the differentiation status of the tumor cells. PMID- 10862720 TI - Sustained elevation in inositol 1,4,5-trisphosphate results in inhibition of phosphatidylinositol transfer protein activity and chronic depletion of the agonist-sensitive phosphoinositide pool. AB - The 43 kDa inositol polyphosphate 5-phosphatase (5-phosphatase) hydrolyses the signalling molecules inositol 1,4,5-trisphosphate (Ins(1,4,5)P(3)) and inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4, 5)P(4)) in a signal-terminating reaction. We have utilised cell lines that stably underexpress the 43 kDa 5-phosphatase, as a model system to investigate whether Ins(1,4,5)P(3) can control the rate of its own formation by regulating the resupply of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)). A sustained 2.6-fold elevation in the basal concentration of Ins(1,4,5)P(3), in cell lines underexpressing the 43 kDa 5-phosphatase, correlated with a 32% reduction in the total cellular mass of PtdIns(4,5)P(2). The depletion in cellular PtdIns(4,5)P(2) was confined to a Triton-insoluble cell compartment, enriched in caveolin. In resting cells with elevated Ins(1,4,5)P(3) concentrations resulting from underexpression of the 43 kDa 5-phosphatase, phosphatidylinositol (PtdIns) and phosphatidylinositol 4-phosphate (PtdIns(4)P) were depleted by 50% and PtdIns(4,5)P(2) by 61% in the caveolin-enriched Triton insoluble compartment. Agonist stimulation resulted in the rapid turnover of phosphoinositides in the caveolin-enriched Triton-insoluble fraction of vector transfected cells, but not in cells with high basal Ins(1,4,5)P(3) concentrations. Depletion of phosphoinositides from the caveolin-enriched Triton insoluble pool in cells underexpressing the 43 kDa 5-phosphatase did not result from activation of phospholipase C isoenzymes, or inhibition of PtdIns 4-kinase or PtdIns(4)P 5-kinase activities. Significant inhibition of phosphatidylinositol transfer protein (PITP) activity (up to 70%) was observed in cells with elevated basal Ins(1,4,5)P(3) concentrations; however, no reduction in PITP(&agr;) protein expression was detected. These studies indicate that chronic elevation in cellular Ins(1,4,5)P(3) concentrations decreases the PITP-mediated resupply of phosphoinositides in the caveolin-enriched agonist-sensitive pool. PMID- 10862721 TI - GABA and responses to GABA in the stomatogastric ganglion of the crab Cancer borealis. AB - The multifunctional neural circuits in the crustacean stomatogastric ganglion (STG) are influenced by many small-molecule transmitters and neuropeptides that are co-localized in identified projection neurons to the STG. We describe the pattern of gamma-aminobutyric acid (GABA) immunoreactivity in the stomatogastric nervous system of the crab Cancer borealis and demonstrate biochemically the presence of authentic GABA in C. borealis. No STG somata show GABA immunoreactivity but, within the stomatogastric nervous system, GABA immunoreactivity co-localizes with several neuropeptides in two identified projection neurons, the modulatory proctolin neuron (MPN) and modulatory commissural neuron 1 (MCN1). To determine which actions of these neurons are evoked by GABA, it is necessary to determine the physiological actions of GABA on STG neurons. We therefore characterized the response of each type of STG neuron to focally applied GABA. All STG neurons responded to GABA. In some neurons, GABA evoked a picrotoxin-sensitive depolarizing, excitatory response with a reversal potential of approximately -40 mV. This response was also activated by muscimol. In many STG neurons, GABA evoked inhibitory responses with both K(+)- and Cl(-) dependent components. Muscimol and beta-guanidinopropionic acid weakly activated the inhibitory responses, but many other drugs, including bicuculline and phaclofen, that act on vertebrate GABA receptors were not effective. In summary, GABA is found in projection neurons to the crab STG and can evoke both excitatory and inhibitory actions on STG neurons. PMID- 10862722 TI - Three-dimensional loading and growth of the zygomatic arch. AB - Despite a number of previous biomechanical studies on the zygomatic arch, unanswered questions remain about its three-dimensional loading and growth. Using young miniature swine, we have for the first time recorded strains from both the medial and lateral aspects of the squamosal bone during mastication and masseter muscle stimulation. Strains from the zygomatic bone flange and zygomatic arch growth data were also obtained from the same animals. A second study on a younger group of animals examined the growth of the zygomatic flange following partial removal of the masseter. Strain data indicated that the squamosal bone is bent out-of-plane and that this pattern of loading is quite different from that of the adjacent zygomatic bone, which experiences much lower strains with little evidence of out-of-plane bending. Surprisingly, strains were higher in the zygomatic flange during contralateral chews and contralateral masseter stimulations than during ipsilateral chews/stimulations. These strains proved to arise from movement of the condyle, explaining why partial removal of the masseter had little effect on the growth of the flange. Other growth results indicated an approximately threefold greater rate of subperiosteal deposition on the lateral surface of the squamosal bone than on the zygomatic bone. This difference in growth rate is attributed to the presence of sutures that contribute to the lateral displacement of the zygomatic bone but not the squamosal bone. This explanation does not exclude the possibility that the rapid apposition on the lateral squamosal surface is regulated by the high surface strains that result from out-of-plane bending. PMID- 10862723 TI - Lipid compositional correlates of temperature-adaptive interspecific differences in membrane physical structure. AB - Teleost species from cold environments possess more disordered brain synaptic membranes than species from warm habitats, thereby providing equivalent physical structures at their respective habitat temperatures. We have related this adaptive interspecific biophysical response to the fatty acid composition of brain membranes from 17 teleost species obtained from Antarctic, temperate and semi-tropical waters, as well as from rat and turkey as representative homeotherms. Cold-adaptive increases in membrane disorder (determined by fluorescence anisotropy with diphenylhexatriene as probe) were correlated with large and linear increases in the proportion of unsaturated fatty acids, from 35 to 60 % in phosphatidylcholine (PtdCho) and from 55 to 85 % in phosphatidylethanolamine (PtdEth). For PtdCho, the cold-adaptive increase in unsaturation was associated almost entirely with increased proportions (from 7 to 40 %) of polyunsaturated fatty acids (PUFAs), with mono-unsaturates (MUFAs) providing an approximately constant proportion in all species. Exactly opposite effects were evident for phosphatidylethanolamine (PtdEth). Thus, the compositional adaptation for PtdCho occurred largely by exchange of polyunsaturated and mono-unsaturated fatty acid in the sn-2 position, whilst for PtdEth it involved exchanges between saturates and mono-unsaturates at the sn-1 position. This difference may be related to the different molecular shapes of the two phosphoglycerides and the need to maintain the balance between bilayer stabilising and -destabilising tendencies. This comparative study provides a more comprehensive view of the compositional adjustments that accompany and perhaps account for temperature-adaptive interspecific differences in membrane physical structure. PMID- 10862724 TI - The visually controlled prey-capture behaviour of the European mantispid Mantispa styriaca. AB - Mantispids (Mantispa styriaca) are predatory insects; on bright sunny days, they wait in ambush for insect prey. The prey is captured as soon as it is within reach by means of lightning-speed strikes with the powerful forelegs. The strikes can take less than 60 ms. The mantispid accomplishes this almost as effectively as the larger praying mantis, which occupies a similar habitat, even though the praying mantis has apposition eyes with a high-resolution fovea, whereas the mantispid has unspecialized optical superposition eyes. Mantispa styriaca reacts to an item of prey when the latter covers a critical visual angle. The detection of prey immediately triggers adjustment reactions in the mantispid, which attempts to position the prey item in the visual field of both eyes and in the capture zone. Irrespective of the size of the prey, the capture reaction of the mantispid is always triggered if the distance to the prey falls below a certain critical value. As indicated by the analysis of individual video frames, immediately before an aimed strike, the item of prey is always positioned exactly in the centre of the binocular field of vision in the extended midsagittal plane of the mantispid's head. The strike may be triggered by the ommatidia of the left and right eyes, the lines of sight of which converge precisely on this region. The principal conclusion to be drawn is that the prey-capture behaviour of the mantispid appears to be based on a triangulation mechanism. PMID- 10862725 TI - Selective heating of vibrissal follicles in seals (Phoca vitulina) and dolphins (Sotalia fluviatilis guianensis). AB - The thermal characteristics of the mystacial vibrissae of harbour seals (Phoca vitulina) and of the follicle crypts on the rostrum of the dolphin Sotalia fluviatilis guianensis were measured using an infrared imaging system. Thermograms demonstrate that, in both species, single vibrissal follicles are clearly defined units of high thermal radiation, indicating a separate blood supply to these cutaneous structures. It is suggested that the high surface temperatures measured in the area of the mouth of the follicles is a function of the sinus system. In seals and dolphins, surface temperature gradually decreased with increasing distance from the centre of a follicle, indicating heat conduction from the sinus system via the follicle capsule to adjacent tissues. It is suggested that the follicular sinus system is a thermoregulatory structure responsible for the maintenance of high tactile sensitivity at the extremely low ambient temperatures demonstrated for the vibrissal system of seals. The vibrissal follicles of odontocetes have been described as vestigial structures, but the thermograms obtained in the present study provide the first evidence that, in Sotalia fluviatilis, the follicles possess a well-developed sinus system, suggesting that they are part of a functional mechanosensory system. PMID- 10862726 TI - Size matters: ontogenetic variation in the three-dimensional kinematics of steady speed locomotion in the lizard Dipsosaurus dorsalis. AB - Although many studies have investigated how locomotor capacities change with size, few studies have examined whether different-sized individuals within a species have similar kinematics during locomotion. We quantified the skeletal limb morphology and the three-dimensional kinematics of the hindlimb of four sizes (4-66 g) of the lizard Dipsosaurus dorsalis moving steadily at both the walk-run transition (50 % duty factor) and at a moderately fast speed of 250 cm s(-)(1). We used analyses of variance to test whether limb movements changed with size and to determine whether size and speed had interactive effects on kinematics. The disproportionately long hindlimbs of smaller lizards partly contributed to their relatively greater (i.e. adjusted by snout-vent length) values of linear kinematic variables. Both relative linear and angular kinematics changed significantly with both size and speed, both of which had widespread interactive effects. By having more extension of the knee and ankle joints, and thus a relatively higher hip height during stance, the slow-speed movements of small lizards displayed some of the characteristics of the fast-speed movements in larger lizards. Further, approximately one-fifth and two-fifths of the strides of the two smallest size classes were digitigrade at the lower and higher speeds, respectively, whereas the two largest size classes always had a plantigrade foot posture. Some of the most striking effects of size on kinematics were most evident at the lower of the two speeds. Unlike interspecific studies, which show that the limbs often become more crouched with decreased size, the more extended limbs of smaller lizards in this study suggest that variation in size alone cannot be the causal reason for differences in limb posture. PMID- 10862727 TI - Influence of tail shape on tadpole swimming performance. AB - Many tadpoles respond to insect predators by developing deeper, and sometimes longer, tails. It has been assumed that the larger tail enhances aspects of swimming performance, because deep-tailed tadpoles survive well when confronted with hunting predators. We tested this hypothesis using both naturally occurring and surgically created variation in tail morphology of Hyla versicolor tadpoles. We measured swimming performance (maximum speed, time to reach a 2.5 cm radius, and angle of escape) and morphology (size and shape of the body and tail) in 288 tadpoles, of which half possessed the predator-induced morphology and the other half were from predator-free ponds. Large tadpoles swam faster than small ones, and shape was significantly correlated with size-corrected swimming performance. The fastest tadpoles had relatively shallow bodies and tail fins, and short tails; there was no difference in swimming performance between predator-induced and no-predator tadpoles. We performed an experiment to create independent variation in tail depth and length by surgically manipulating tail shape in 270 tadpoles. Three tail-length treatments reduced the length of the tail fin by 21 %, 34 % and 55 %; three tail-depth treatments reduced the maximum depth of the tail fin by 11 %, 34 % and 59 %; two additional treatments controlled for the effects of anaesthesia and surgery. The angle of escape was unaffected by surgery. Maximum speed and minimum escape time were both significantly impaired by the high-removal treatments, but showed no evidence of decline until 30 % of the tail (length or depth) was removed. These results suggest that the relatively deep tails in predator-induced tadpoles (approximately 10 % deeper than in no predator tadpoles) do not improve performance in burst swimming. Thus, predator induced tadpoles are less vulnerable to predation for reasons other than enhanced swimming performance. PMID- 10862728 TI - Terrestrial locomotion in the black-billed magpie: kinematic analysis of walking, running and out-of-phase hopping. AB - The inter-limb kinematic patterns of walking, running and out-of-phase hopping in black-billed magpies (Pica pica) were studied using high-speed video recordings. The flexion/extension patterns of the joints were similar between the gait types, suggesting that the within-leg control of the angular excursions is similar. This result is further supported by the fact that running and hopping are alternative gaits at speeds higher than walking; however, magpies show a preference for hopping. Moreover, only small differences occur between the kinematic patterns of the two limbs during out-of-phase hopping, during which the legs are believed to have different functions. The hindlimb kinematic patterns of magpies are like those of other flying and more terrestrial bird species; however, striking differences are found in comparison with humans at the level of the internal angles. This is probably due to the differences in the morphology and configuration of their legs. PMID- 10862729 TI - Drinking in snakes: kinematic cycling and water transport. AB - Snakes are purported to drink by sucking water into their mouths and then compressing the oral cavity to force water into the oesophagus. Video recordings of drinking behaviour in 23 snakes representing 14 species from three families, combined with simultaneous recordings of water volumes consumed, show that all the snakes vary widely in the amount of water taken in when drinking. This variation is not correlated with kinematic events. Kinematic recordings and indirect measurements of water flow suggest that moving water into the mouth can be decoupled from the processes that move water into the oesophagus and that, infrequently, water may continue flowing into the mouth during both opening (suction) and closing (presumed compression) of the mouth. Drinking in snakes is not a simple, stereotyped behaviour. Different snake species differ in both drinking kinematics and water inflow patterns. Vertical excursions of the mandible are smallest in booids and larger, but highly variable, in different viperids and colubrids. Cyclic movements of the tongue seen in booids are not evident in viperids or colubrids. All the snakes usually take in water at rates far below their potential maximum rate. Although drinking is apparently achieved by suction, a single model cannot explain all water movement patterns in snakes. At a practical level, functional morphological studies of drinking in snakes (and possibly many other animals) must demonstrate that fluid flow actually correlates with kinematic events. Without such an empirical demonstration, interpretation of other measurements (pressure, movement, etc.) is unlikely to produce meaningful models. PMID- 10862730 TI - The development of fatigue quality in high- and low-stressed tendons of sheep (Ovis aries). AB - The time taken to rupture in cyclic fatigue tests, to a stress of 45 MPa, was used to compare the fatigue quality of tendons from sheep of varying ages. Muscle and tendon cross-sectional areas were used to calculate the stress-in-life of each tendon. For any given age, high-stressed plantaris tendons were of a higher fatigue quality than low-stressed extensor tendons. Both fatigue quality and stress-in-life increased with age for each tendon type. High-stressed tendons are subjected to large increases in stress-in-life during growth, and fatigue quality increased significantly with this stress. This relationship was not seen, however, in low-stressed tendons, which are not subjected to a comparable range of stresses over time. It is possible that cells modify tendon fatigue quality in response to tendon loading history. Whilst Young's modulus was seen to increase with age, no difference was detected between high- and low-stressed tendons. PMID- 10862731 TI - Preferred speed and cost of transport: the effect of incline. AB - Preferred speed is the behavioral tendency of animals to utilize a relatively narrow set of speeds near the middle of a much broader range that they are capable of using within a particular gait. Possible explanations for this behavior include minimizing musculoskeletal stresses and maximizing energetic economy. If preferred speed is determined by energetic economy (cost of transport, C(T)), then shifts in preferred speed should produce shifts in C(T). To test this hypothesis, preferred speeds were measured in trotting horses on the level and on an incline. The preferred trotting speed decreased from 3.29+/-0.24 m s(-)(1) on the level to 3.05+/-0.30 m s(-)(1) (means +/- s.d., N=6) on an 11.8 % incline. The rate of oxygen consumption was measured as a function of trotting speed on a treadmill and was a curvilinear function of speed in all horses under both conditions (level and 10 % incline). This curvilinear relationship resulted in a C(T) that was a U-shaped function of speed. The speed at which C(T) was minimal (i.e. at which trotting was most energetically economical) was very near the preferred speed on the level and decreased on the incline, again to a speed near the preferred speed on the incline. PMID- 10862732 TI - Evidence for the role of a Na(+)/HCO(3)(-) cotransporter in trout hepatocyte pHi regulation. AB - The mechanisms of intracellular pH (pHi) regulation were examined in hepatocytes of the rainbow trout Oncorhynchus mykiss. pHi was monitored using the pH sensitive fluorescent dye BCECF, and the effects of various media and pharmacological agents were examined for their influence on baseline pHi and recovery rates from acid and base loading. Rates of Na(+) uptake were measured using (22)Na, and changes in membrane potential were examined using the potentiometric fluorescent dye Oxonol VI. The rate of proton extrusion following acid loading was diminished by the blockade of either Na(+)/H(+) exchange (using amiloride) or anion transport (using DIDS). The removal of external HCO(3)(-) and the abolition of outward K(+) diffusion by the channel blocker Ba(2+) also decreased the rate of proton extrusion following acid load. Depolarization of the cell membrane with 50 mmol l(-)(1) K(+), however, did not affect pHi. The rate of recovery from base loading was significantly diminished by the blockade of anion transport, removal of external HCO(3)(-) and, to a lesser extent, by blocking Na(+)/H(+) exchange. The blockade of K(+) conductance had no effect. The decrease in Na(+) uptake rate observed in the presence of the anion transport blocker DIDS and the DIDS-sensitive hyperpolarization of membrane potential during recovery from acid loading suggest that a Na(+)-dependent electrogenic transport system is involved in the restoration of pHi after intracellular acidification. The effects on baseline pHi indicate that the different membrane exchangers are tonically active in the maintenance of steady-state pHi. This study confirms the roles of a Na(+)/H(+) exchanger and a Cl(-)/HCO(3)(-) exchanger in the regulation of trout hepatocyte pHi and provides new evidence that a Na(+)/HCO(3)(-) cotransporter contributes to pHi regulation. PMID- 10862733 TI - ATP production from the oxidation of sulfide in gill mitochondria of the ribbed mussel Geukensia demissa. AB - The ribbed mussel Geukensia demissa inhabits intertidal Spartina grass marshes characterized by sulfide-rich sediments. Sulfide poisons aerobic respiration, and G. demissa may cope in this seemingly inhospitable environment by oxidizing sulfide in gill mitochondria. Well-coupled mitochondria isolated from G. demissa gills were used to investigate sulfide oxidation and ATP synthesis. State 3 respiration, maximally stimulated by 5 micromol l(-)(1) sulfide with a P/O ratio of 0.89 and a respiratory control ratio (RCR) of 1.40, remained refractory to sulfide at higher concentrations except in the presence of salicylhydroxamic acid (SHAM), an inhibitor of alternative oxidases. Sulfide-stimulated ATP production was 3-5 times greater than that stimulated by malate and succinate, respectively, giving an ATP/sulfide ratio of 0.63. The inhibition of sulfide-stimulated respiration and ATP production by the complex III inhibitors myxothiazol and antimycin A, respectively, suggests that electrons enter the electron transport chain before complex III. Combined with in vivo evidence for electron entry at cytochrome c, these data suggest that more than one type of sulfide-oxidizing enzyme may function in G. demissa gills. The SHAM-sensitive pathway of electron flux may be a critical component of a physiological strategy to tolerate sulfide. We conclude that G. demissa exploits the energy available from its reduced environment by using sulfide as a respiratory substrate for cellular ATP production. PMID- 10862734 TI - The effects of cell ageing on protein synthesis in rainbow trout (Oncorhynchus mykiss) red blood cells. AB - The effects of cell age on protein synthesis were examined in the nucleated red blood cells of rainbow trout (Oncorhynchus mykiss). Total DNA content was unaffected by cell age, whereas total RNA content in young red blood cells was roughly ten times as high as that in old red blood cells. The mRNA levels for haemoglobin, carbonic anhydrase and the chloride/bicarbonate (Cl(-)/HCO(3)(-)) exchanger were also approximately tenfold higher in young red blood cells. Although young red blood cells synthesized roughly five times more protein under steady-state conditions, total protein concentration was not affected by cell age. Despite large reductions in mRNA levels with red blood cell ageing, the concentrations and/or activities of the respiratory proteins were largely preserved. In contrast, the ability to mount a heat shock response was greatly reduced in older red blood cells. Young red blood cells produced 13 times more heat shock protein 70 mRNA following heat shock and four times more 70 kDa protein after recovery. They also transcribed much more heat shock cognate 71 and heat shock factor mRNA than did older red blood cells under steady-state conditions. PMID- 10862735 TI - The resting membrane potential of white muscle from brown trout (Salmo trutta) exposed to copper in soft, acidic water. AB - Previously, the distribution of ammonia between the intracellular and extracellular compartments has been used to predict a significant depolarisation of the resting membrane potential (E(M)) of white muscle from brown trout (Salmo trutta) exposed to a sub-lethal combination of copper and low pH. However, this prediction is based upon two assumptions (i) a relatively high membrane permeability for the ammonium ion with respect to that for ammonia gas and (ii) that this is unaltered by exposure to copper and low pH. Since there is conflicting evidence in the literature of the validity of these assumptions, in the present study E(M) was directly measured in white muscle fibres of trout exposed to copper and low pH (E(M)=-52.2+/-4.9 mV) and compared with that of unexposed, control animals (E(M)=-86.5+/-2.9 mV) (means +/- s.e.m., N=6). In confirming the predicted depolarisation, these data support the hypothesis of electrophysiological impairment as a factor in the reduction in the swimming performance of trout exposed to these pollutants. In addition, the results of this study support the role of a significant permeability of the muscle membrane to NH(4)(+) in determining the distribution of ammonia in fish. PMID- 10862736 TI - Measuring dimensions: the regulation of size and shape. AB - Over many years evidence has accumulated that plants and animals can regulate growth with reference to overall size rather than cell number. Thus, organs and organisms grow until they reach their characteristic size and shape and then they stop - they can even compensate for experimental manipulations that change, over several fold, cell number or average cell size. If the cell size is altered, the organism responds with a change in cell number and vice versa. We look at the Drosophila wing in more detail: here, both extracellular and intracellular regulators have been identified that link cell growth, division and cell survival to final organ size. We discuss a hypothesis that the local steepness of a morphogen gradient is a measure of length in one axis, a measure that is used to determine whether there will be net growth or not. PMID- 10862737 TI - Basic helix-loop-helix proteins and the timing of oligodendrocyte differentiation. AB - An intracellular timer in oligodendrocyte precursor cells is thought to help control the timing of their differentiation. We show here that the expression of the Hes5 and Mash1 genes, which encode neural-specific bHLH proteins, decrease and increase, respectively, in these cells with a time course expected if the proteins are part of the timer. We show that enforced expression of Hes5 in purified precursor cells strongly inhibits the normal increase in the thyroid hormone receptor protein TR(&bgr;)1, which is thought to be part of the timing mechanism; it also strongly inhibits the differentiation induced by either mitogen withdrawal or thyroid hormone treatment. Enforced expression of Mash1, by contrast, somewhat accelerates the increase in TR(beta)1 protein. These findings suggest that Hes5 and Mash1 may be part of the cell-intrinsic timer in the precursor cells. PMID- 10862738 TI - The repressor and activator forms of Cubitus interruptus control Hedgehog target genes through common generic gli-binding sites. AB - The Drosophila Gli homolog Cubitus interruptus (Ci) controls the transcription of Hedgehog (Hh) target genes. A repressor form of Ci arises in the absence of Hh signalling by proteolytic cleavage of intact Ci, whereas an activator form of Ci is generated in response to the Hh signal. These different activities of Ci regulate overlapping but distinct subsets of Hh target genes. To investigate the mechanisms by which the two activities of Ci exert their opposite transcriptional effect, we dissect here the imaginal disc enhancer of the dpp gene, which responds to both activities of Ci. Within a minimal disc enhancer, we identify the DNA sequences that are necessary and sufficient for the control by Ci, show that the same sequences respond to the activator and repressor forms of Ci, and demonstrate that their activities can be replaced by a single synthetic Gli binding site. We further show that the enhancer sequences of patched, a gene responding only to the activator form of Ci, effectively integrate also the repressor activity of Ci if placed into a dpp context. These results provide in vivo evidence against the employment of distinct binding sites for the different forms of Ci and suggest that target genes responding to only one form must have acquired distant cis-regulatory elements for their selective behavior. PMID- 10862739 TI - (beta)-catenin mediates the specification of endoderm cells in ascidian embryos. AB - In the present study, we addressed the role of (beta)-catenin in the specification of embryonic cells of the ascidians Ciona intestinalis and C. savignyi and obtained the following results: (1) During cleavages, (beta)-catenin accumulated in the nuclei of vegetal blastomeres, suggesting that it plays a role in the specification of endoderm. (2) Mis- and/or overexpression of (beta) catenin induced the development of an endoderm-specific alkaline phosphatase (AP) in presumptive notochord cells and epidermis cells without affecting differentiation of primary lineage muscle cells. (3) Downregulation of (beta) catenin induced by the overexpression of cadherin resulted in the suppression of endoderm cell differentiation. This suppression was compensated for by the differentiation of extra epidermis cells. (4) Specification of notochord cells did not take place in the absence of endoderm differentiation. Both the overexpression of (beta)-catenin in presumptive notochord cells and the downregulation of (beta)-catenin in presumptive endoderm cells led to the suppression of Brachyury gene expression, resulting in the failure of notochord specification. These results suggest that the accumulation of (beta)-catenin in the nuclei of endoderm progenitor cells is the first step in the process of ascidian endoderm specification. PMID- 10862740 TI - Misexpression of basic helix-loop-helix genes in the murine cerebral cortex affects cell fate choices and neuronal survival. AB - To investigate the role(s) of basic helix-loop-helix genes (bHLH) genes in the developing murine cerebral cortex, Mash1, Math2, Math3, Neurogenin1 (Ngn1), Ngn2, NeuroD, NeuroD2 and Id1 were transduced in vivo into the embryonic and postnatal cerebral cortex using retrovirus vectors. The morphology and location of infected cells were analyzed at postnatal stages. The data indicate that a subset of bHLH genes are capable of regulating the choice of neuronal versus glial fate and that, when misexpressed, they can be deleterious to the survival of differentiating neurons, but not glia. PMID- 10862741 TI - An orthologue of the kit-related gene fms is required for development of neural crest-derived xanthophores and a subpopulation of adult melanocytes in the zebrafish, Danio rerio. AB - Developmental mechanisms underlying traits expressed in larval and adult vertebrates remain largely unknown. Pigment patterns of fishes provide an opportunity to identify genes and cell behaviors required for postembryonic morphogenesis and differentiation. In the zebrafish, Danio rerio, pigment patterns reflect the spatial arrangements of three classes of neural crest derived pigment cells: black melanocytes, yellow xanthophores and silver iridophores. We show that the D. rerio pigment pattern mutant panther ablates xanthophores in embryos and adults and has defects in the development of the adult pattern of melanocyte stripes. We find that panther corresponds to an orthologue of the c-fms gene, which encodes a type III receptor tyrosine kinase and is the closest known homologue of the previously identified pigment pattern gene, kit. In mouse, fms is essential for the development of macrophage and osteoclast lineages and has not been implicated in neural crest or pigment cell development. In contrast, our analyses demonstrate that fms is expressed and required by D. rerio xanthophore precursors and that fms promotes the normal patterning of melanocyte death and migration during adult stripe formation. Finally, we show that fms is required for the appearance of a late developing, kit-independent subpopulation of adult melanocytes. These findings reveal an unexpected role for fms in pigment pattern development and demonstrate that parallel neural crest-derived pigment cell populations depend on the activities of two essentially paralogous genes, kit and fms. PMID- 10862742 TI - Establishing neuronal identity in vertebrate neurogenic placodes. AB - The trigeminal and epibranchial placodes of vertebrate embryos form different types of sensory neurons. The trigeminal placodes form cutaneous sensory neurons that innervate the face and jaws, while the epibranchial placodes (geniculate, petrosal and nodose) form visceral sensory neurons that innervate taste buds and visceral organs. In the chick embryo, the ophthalmic trigeminal (opV) placode expresses the paired homeodomain transcription factor Pax3 from very early stages, while the epibranchial placodes express Pax2. Here, we show that Pax3 expression in explanted opV placode ectoderm correlates at the single cell level with neuronal specification and with commitment to an opV fate. When opV (trigeminal) ectoderm is grafted in place of the nodose (epibranchial) placode, Pax3-expressing cells form Pax3-positive neurons on the same schedule as in the opV placode. In contrast, Pax3-negative cells in the grafted ectoderm are induced to express the epibranchial placode marker Pax2 and form neurons in the nodose ganglion that express the epibranchial neuron marker Phox2a on the same schedule as host nodose neurons. They also project neurites along central and peripheral nodose neurite pathways and survive until well after the main period of cell death in the nodose ganglion. The older the opV ectoderm is at the time of grafting, the more Pax3-positive cells it contains and the more committed it is to an opV fate. Our results suggest that, within the neurogenic placodes, there does not appear to be a two-step induction of 'generic' neurons followed by specification of the neuron to a particular fate. Instead, there seems to be a one-step induction in which neuronal subtype identity is coupled to neuronal differentiation. PMID- 10862743 TI - Regulation of retinoic acid signaling during lung morphogenesis. AB - Little is known about how retinoic acid (RA) synthesis, utilization and metabolism are regulated in the embryonic lung and how these activities relate to lung pattern formation. Here we report that early lung bud formation and subsequent branching morphogenesis are characterized by distinct stages of RA signaling. At the onset of lung development RA signaling is ubiquitously activated in primary buds, as shown by expression of the major RA-synthesizing enzyme, RALDH-2 and activation of a RARE-lacZ transgene. Nevertheless, further airway branching appears to require downregulation of RA pathways by decreased synthesis, increased RA degradation in the epithelium via P450RAI-mediated metabolism, and inhibition of RA signaling in the mesenchyme by COUPTF-II expression. These mechanisms controlling local RA signaling may be critical for normal branching, since we show that manipulating RA levels in vitro to maintain RA signaling activated as in the initial stage, leads to an immature lung phenotype characterized by failure to form typical distal buds. We show that this phenotype likely results from RA interfering with the establishment of a distal signaling center, altering levels and distribution of Fgf10 and Bmp4, genes that are essential for distal lung formation. Furthermore, RA upregulates P450RAI expression, suggesting the presence of feedback mechanisms controlling RA availability. Our study illustrates the importance of regional mechanisms that control RA availability and utilization for correct expression of pattern regulators and normal morphogenesis during lung development. PMID- 10862744 TI - Temporal regulation of apterous activity during development of the Drosophila wing. AB - Dorsoventral axis formation in the Drosophila wing depends on the activity of the selector gene apterous. Although selector genes are usually thought of as binary developmental switches, we find that Apterous activity is negatively regulated during wing development by its target gene dLMO. Apterous-dependent expression of Serrate and fringe in dorsal cells leads to the restricted activation of Notch along the dorsoventral compartment boundary. We present evidence that the ability of cells to participate in this Apterous-dependent cell-interaction is under spatial and temporal control. Apterous-dependent expression of dLMO causes downregulation of Serrate and fringe and allows expression of delta in dorsal cells. This limits the time window during which dorsoventral cell interactions can lead to localized activation of Notch and induction of the dorsoventral organizer. Overactivation of Apterous in the absence of dLMO leads to overexpression of Serrate, reduced expression of delta and concomitant defects in differentiation and cell survival in the wing primordium. Thus, downregulation of Apterous activity is needed to allow normal wing development. PMID- 10862745 TI - Formation of the definitive endoderm in mouse is a Smad2-dependent process. AB - TGFbeta growth factors specify cell fate and establish the body plan during early vertebrate development. Diverse cellular responses are elicited via interactions with specific cell surface receptor kinases that in turn activate Smad effector proteins. Smad2-dependent signals arising in the extraembryonic tissues of early mouse embryos serve to restrict the site of primitive streak formation and establish anteroposterior identity in the epiblast. Here we have generated chimeric embryos using lacZ-marked Smad2-deficient ES cells. Smad2 mutant cells extensively colonize ectodermal and mesodermal populations without disturbing normal development, but are not recruited into the definitive endoderm lineage during gastrulation. These experiments provide the first evidence that TGFbeta signaling pathways are required for specification of the definitive endoderm lineage in mammals and identify Smad2 as a key mediator that directs epiblast derivatives towards an endodermal as opposed to a mesodermal fate. In largely Smad2-deficient chimeras, asymmetric nodal gene expression is maintained and expression of pitx2, a nodal target, is also unaffected. These results strongly suggest that other Smad(s) act downstream of Nodal signals in mesodermal populations. We found Smad2 and Smad3 transcripts both broadly expressed in derivatives of the epiblast. However, Smad2 and not Smad3 mRNA is expressed in the visceral endoderm, potentially explaining why the primary defect in Smad2 mutant embryos originates in this cell population. PMID- 10862746 TI - Role of frizzled 7 in the regulation of convergent extension movements during gastrulation in Xenopus laevis. AB - Wnt signalling plays a crucial role in the control of morphogenetic movements. We describe the expression and functional analyses of frizzled 7 (Xfz7) during gastrulation in Xenopus. Low levels of Xfz7 transcripts are expressed maternally during cleavage stages; its zygotic expression strongly increases at the beginning of gastrulation and is predominantly localized to the presumptive neuroectoderm and deep cells of the involuting mesoderm. Overexpression of Xfz7 in the dorsal equatorial region affects the movements of convergent extension and delays mesodermal involution. It alters the correct localization, but not the expression, of mesodermal and neural markers. These effects can be rescued by extra-Xfz7, which is a secreted form of the receptor that also weakly inhibits convergent extension when overexpressed. This suggests that the wild-type and truncated receptors have opposing effects when coexpressed and that overexpression of Xfz7 causes an increased signalling activity. Consistent with this, Xfz7 biochemically and functionally interacts with Xwnt11. In addition, Dishevelled, but not (&bgr;)-catenin, synergizes with Xfz7 to affect convergent extension. Furthermore, overexpression of Xfz7 and Xwnt11 also affects convergent extension in activin-treated animal caps, and this can be efficiently reversed by coexpression of Cdc42(T17N), a dominant negative mutant of the small GTPase Cdc42 known as a key regulator of actin cytoskeleton. Conversely, Cdc42(G12V), a constitutively active mutant, rescues the effects of extra-Xfz7 on convergent extension in a dose-dependent manner. That both gain-of-function and loss-of function of both frizzled and dishevelled produce the same phenotype has been well described in Drosophila tissue polarity. Therefore, our results suggest an endogenous role of Xfz7 in the regulation of convergent extension during gastrulation. PMID- 10862747 TI - Imprint switching for non-random X-chromosome inactivation during mouse oocyte growth. AB - In mammals, X-chromosome inactivation occurs in all female cells, leaving only a single active X chromosome. This serves to equalise the dosage of X-linked genes in male and female cells. In the mouse, the paternally derived X chromosome (X(P)) is imprinted and preferentially inactivated in the extraembryonic tissues whereas in the embryonic tissues inactivation is random. To investigate how X(P) is chosen as an inactivated X chromosome in the extraembryonic cells, we have produced experimental embryos by serial nuclear transplantation from non-growing (ng) oocytes and fully grown (fg) oocytes, in which the X chromosomes are marked with (1) an X-linked lacZ reporter gene to assay X-chromosome activity, or (2) the Rb(X.9)6H translocation as a cytogenetic marker for studying replication timing. In the extraembryonic tissues of these ng/fg embryos, the maternal X chromosome (X(M)) derived from the ng oocyte was preferentially inactivated whereas that from the fg oocyte remained active. However, in the embryonic tissues, X inactivation was random. This suggests that (1) a maternal imprint is set on the X(M) during oocyte growth, (2) the maternal imprint serves to render the X(M) resistant to inactivation in the extraembryonic tissues and (3) the X(M) derived from an ng oocyte resembles a normal X(P). PMID- 10862748 TI - Transforming growth factor beta3 induces cell death during the first stage of mammary gland involution. AB - Involution of the mammary gland following weaning is divided into two distinct phases. Initially, milk stasis results in the induction of local factors that cause apoptosis in the alveolar epithelium. Secondly after a prolonged absence of suckling, the consequent decline in circulating lactogenic hormone concentrations initiates remodeling of the mammary gland to the virgin-like state. We have shown that immediately following weaning TGFbeta3 mRNA and protein is rapidly induced in the mammary epithelium and that this precedes the onset of apoptosis. Unilateral inhibition of suckling and hormonal reconstitution experiments showed that TGFbeta3 induction is regulated by milk stasis and not by the circulating hormonal concentration. Directed expression of TGFbeta3 in the alveolar epithelium of lactating mice using a beta-lactoglobulin promoter mobilized SMAD4 translocation to the nucleus and caused apoptosis of these cells, but not tissue remodeling. Transplantation of neonatal mammary tissue derived from TGFbeta3 null mutant mice into syngenic hosts resulted in a significant inhibition of cell death compared to wild-type mice upon milk stasis. These results provide direct evidence that TGFbeta3 is a local mammary factor induced by milk stasis that causes apoptosis in the mammary gland epithelium during involution. PMID- 10862749 TI - TRA-1A regulates transcription of fog-3, which controls germ cell fate in C. elegans. AB - In C. elegans, the zinc-finger protein TRA-1A is thought to be the final arbiter of somatic sexual identity. We show that fog-3, which is required for germ cells to become sperm rather than oocytes, is a target of TRA-1A. First, northern analyses and RT-PCR experiments indicate that expression of fog-3 is controlled by tra-1. Second, studies of double mutants show that this control could be direct. Third, the fog-3 promoter contains multiple sites that bind TRA-1A in gel shift assays, and mutations in these sites alter activity of fog-3 in vivo. These results establish fog-3 as one of the first known targets of transcriptional regulation by TRA-1A. Furthermore, they show that tra-1 controls a terminal regulator of sexual fate in germ cells, just as it is thought to do in the soma. PMID- 10862750 TI - Nuclear import of cubitus interruptus is regulated by hedgehog via a mechanism distinct from Ci stabilization and Ci activation. AB - The Hedgehog (Hh) signal is transduced via Cubitus interruptus (Ci) to specify cell fates in the Drosophila wing. In the absence of Hh, the 155 kDa full-length form of Ci is cleaved into a 75 kDa repressor. Hh inhibits the proteolysis of full-length Ci and facilitates its conversion into an activator. Recently, it has been suggested that Hh promotes Ci nuclear import in tissue culture cells. We have studied the mechanism of Ci nuclear import in vivo and the relationship between nuclear import, stabilization and activation. We found that Ci rapidly translocates to the nucleus in cells close to the anteroposterior (AP) boundary and this rapid nuclear import requires Hh signaling. The nuclear import of Ci is regulated by Hh even under conditions in which Ci is fully stabilized. Furthermore, cells that exhibit Ci stabilization and rapid nuclear import do not necessarily exhibit maximal Ci activity. It has been previously shown that stabilization does not suffice for activation. Consistent with this finding, our results suggest that the mechanisms regulating nuclear import, stabilization and activation are distinct from each other. Finally, we show that cos2 and pka, two molecules that have been characterized primarily as negative regulators of Ci activity, also have positive roles in the activation of Ci in response to Hh. PMID- 10862751 TI - Dual roles of Wnt signaling during chondrogenesis in the chicken limb. AB - Long bones of the appendicular skeleton are formed from a cartilage template in a process known as endochondral bone development. Chondrocytes within this template undergo a progressive program of differentiation from proliferating to postmitotic prehypertrophic to hypertrophic chondrocytes, while mesenchymal cells immediately surrounding the early cartilage template form the perichondrium. Recently, members of the Wnt family of secreted signaling molecules have been implicated in regulating chondrocyte differentiation. We find that Wnt-5a, Wnt-5b and Wnt-4 genes are expressed in chondrogenic regions of the chicken limb: Wnt-5a is expressed in the perichondrium, Wnt-5b is expressed in a subpopulation of prehypertrophic chondrocytes and in the outermost cell layer of the perichondrium, and Wnt-4 is expressed in cells of the joint region. Misexpression experiments demonstrate that two of these Wnt molecules, Wnt-5a and Wnt-4, have opposing effects on the differentiation of chondrocytes and that these effects are mediated through divergent signaling pathways. Specifically, Wnt-5a misexpression delays the maturation of chondrocytes and the onset of bone collar formation, while Wnt-4 misexpression accelerates these two processes. Misexpression of a stabilized form of beta-catenin also results in accelerated chondrogenesis, suggesting that a beta-catenin/TCF-LEF complex is involved in mediating the positive regulatory effect of Wnt-4. A number of the genes involved in Wnt signal tranduction, including two members of the Frizzled gene family, which are believed to encode Wnt-receptors, show very dynamic and distinct expression patterns in cartilaginous elements of developing chicken limbs. Misexpression of putative dominant-negative forms of the two Frizzled proteins results in severe shortening of the infected cartilage elements due to a delay in chondrocyte maturation, indicating that an endogenous Wnt signal does indeed function to promote chondrogenic differentiation. PMID- 10862752 TI - Shoot meristem size is dependent on inbred background and presence of the maize homeobox gene, knotted1. AB - The knotted1 (kn1) gene of maize is expressed in meristems and is absent from leaves, including the site of leaf initiation within the meristem. Recessive mutations of kn1 have been described that limit the capacity to make branches and result in extra carpels. Dominant mutations suggest that kn1 function plays a role in maintaining cells in an undifferentiated state. We took advantage of a Ds induced dominant allele in order to screen for additional recessive alleles resulting from mobilization of the Ds element. Analysis of one such allele revealed a novel embryonic shoot phenotype in which the shoot initiated zero to few organs after the cotyledon was made, resulting in plants that arrested as seedlings. We refer to this phenotype as a limited shoot. The limited shoot phenotype reflected loss of kn1 function, but its penetrance was background dependent. We examined meristem size and found that plants lacking kn1 function had shorter meristems than non-mutant siblings. Furthermore, meristems of restrictive inbreds were significantly shorter than meristems of permissive inbreds, implying a correlation between meristem height and kn1 gene function in the embryo. Analysis of limited shoot plants during embryogenesis indicated a role for kn1 in shoot meristem maintenance. We discuss a model for kn1 in maintenance of the morphogenetic zone of the shoot apical meristem. PMID- 10862753 TI - Restricted patterning of vestigial expression in Drosophila wing imaginal discs requires synergistic activation by both Mad and the drifter POU domain transcription factor. AB - The Drosophila Vestigial protein has been shown to play an essential role in the regulation of cell proliferation and differentiation within the developing wing imaginal disc. Cell-specific expression of vg is controlled by two separate transcriptional enhancers. The boundary enhancer controls expression in cells near the dorsoventral (DV) boundary and is regulated by the Notch signal transduction pathway, while the quadrant enhancer responds to the Decapentaplegic and Wingless morphogen gradients emanating from cells near the anteroposterior (AP) and DV boundaries, respectively. MAD-dependent activation of the vestigial quadrant enhancer results in broad expression throughout the wing pouch but is excluded from cells near the DV boundary. This has previously been thought to be due to direct repression by a signal from the DV boundary; however, we show that this exclusion of quadrant enhancer-dependent expression from the DV boundary is due to the absence of an additional essential activator in those cells. The Drosophila POU domain transcriptional regulator, Drifter, is expressed in all cells within the wing pouch expressing a vgQ-lacZ transgene and is also excluded from the DV boundary. Viable drifter hypomorphic mutations cause defects in cell proliferation and wing vein patterning correlated with decreased quadrant enhancer-dependent expression. Drifter misexpression at the DV boundary using the GAL4/UAS system causes ectopic outgrowths at the distal wing tip due to induction of aberrant Vestigial expression, while a dominant-negative Drifter isoform represses expression of vgQ-lacZ and causes severe notching of the adult wing. In addition, we have identified an essential evolutionarily conserved sequence element bound by the Drifter protein with high affinity and located adjacent to the MAD binding site within the quadrant enhancer. Our results demonstrate that Drifter functions along with MAD as a direct activator of Vestigial expression in the wing pouch. PMID- 10862754 TI - Structural requirements for notch signalling with delta and serrate during the development and patterning of the wing disc of Drosophila. AB - The delta and Serrate proteins interact with the extracellular domain of the Notch receptor and initiate signalling through the receptor. The two ligands are very similar in structure and have been shown to be interchangeable experimentally; however, loss of function analysis indicates that they have different functions during development and analysis of their signalling during wing development indicates that the Fringe protein can discriminate between the two ligands. This raises the possibility that the signalling of delta and Serrate through Notch requires different domains of the Notch protein. Here we have tested this possibility by examining the ability of delta and Serrate to interact and signal with Notch molecules in which different domains had been deleted. This analysis has shown that EGF-like repeats 11 and 12, the RAM-23 and cdc10/ankyrin repeats and the region C-terminal to the cdc10/ankyrin repeats of Notch are necessary for both delta and Serrate to signal via Notch. They also indicate, however, that delta and Serrate utilise EGF-like repeats 24-26 of Notch for signalling, but there are significant differences in the way they utilise these repeats. PMID- 10862755 TI - Synergistic activation of NF-kappa B by functional cooperation between vav and PKCtheta in T lymphocytes. AB - Here we identified PKCtheta as an activator of transcription factor NF-kappaB in T cells. PKCtheta-induced NF-kappaB activation was synergistically augmented by Vav. Several experimental approaches revealed that PKCtheta is located downstream from Vav in the control of the pathway leading to synergistic NF-kappaB activation. In addition to the synergistic activation cascade, Vav also triggered NF-kappaB activity on a separate route. CD3/CD28-induced activation of NF-kappaB was inhibited by dominant negative forms of Vav or PKCtheta, revealing their essential role in this activation pathway. The Vav/PKCtheta-mediated signals preferentially activated IkappaB kinase beta. Vav and PKCtheta were found to be constitutively associated in unstimulated T cells. Only the ligation of the costimulatory CD28 receptor, but not of the T cell receptor, resulted in the transient dissociation of the Vav-PKCtheta complex. In contrast, T cell receptor/CD28 costimulation resulted in faster dissociation and slower reassociation kinetics. PMID- 10862756 TI - FADD is required for DR4- and DR5-mediated apoptosis: lack of trail-induced apoptosis in FADD-deficient mouse embryonic fibroblasts. AB - TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) is a member of the tumor necrosis factor family that can kill a wide variety of tumor cells but not normal cells. TRAIL-induced apoptosis in humans is mediated by its receptors DR4 (TRAIL-R1) and DR5 (TRAIL-R2). What constitutes the signaling molecules downstream of these receptors, however, remains highly controversial. Using the FADD dominant negative molecule, several groups have reached different conclusions with respect to the role of FADD in TRAIL-induced apoptosis. More recently, using FADD-deficient (-/-) mouse embryonic fibroblasts, Yeh et al. (Yeh, W.-C., Pompa, J. L., McCurrach, M. E., Shu, H.-B., Elia, A. J., Shahinian, A., Ng, M., Wakeham, A., Khoo, W., Mitchell, K., El-Deiry, W. S., Lowe, S. W., Goeddel, D. V., and Mak, T. W. (1998) Science 279, 1954-1958) concluded that DR4 utilizes a FADD-independent apoptotic pathway. The latter experiment, however, involved transient overexpression, which often leads to nonspecific aggregation of death domain-containing receptors. To address this issue in a more physiological setting, we stably transfected mouse DR4/5, human DR4, or human DR5 into FADD(-/-) mouse embryonic fibroblast cells. We showed that FADD(-/-) MEF cells stably transfected with TRAIL receptors are resistant to TRAIL-mediated cell death. In contrast, TRAIL receptors stably transfected into heterozygous FADD(+/-) cells or FADD(-/-) cells reconstituted with a FADD retroviral construct are sensitive to the TRAIL cytotoxic effect. We conclude that FADD is required for DR4- and DR5-mediated apoptosis. PMID- 10862757 TI - GATA zinc finger interactions modulate DNA binding and transactivation. AB - GATA-1 and other vertebrate GATA factors contain a DNA binding domain composed of two adjacent homologous zinc fingers. Whereas only the C-terminal finger of GATA 1 is capable of independent binding to the GATA recognition sequence, double GATA sites that require both fingers for high affinity interaction are found in several genes. We propose a mechanism whereby adjacent zinc fingers interact to influence the binding and transactivation properties of GATA-1 at a subset of DNA binding sites. By using two such double GATA sites we demonstrate that the N terminal finger and adjacent linker region can alter the binding specificity of the C-terminal finger sufficiently to prevent it from recognizing some consensus GATA sequences. Therefore, the two zinc fingers form a composite binding domain having a different DNA binding specificity from that shown by the constituent single C-terminal finger. Furthermore, we compare two of these double sites and show that high affinity binding of GATA-1 to a reporter gene does not necessarily induce transactivation, namely the sequence of the DNA-binding site can alter the ability of GATA-1 to stimulate transcription. PMID- 10862758 TI - Aggregation chaperones enhance aggregation and storage of secretory proteins in endocrine cells. AB - Calcium-induced aggregation has been proposed to play a role in the sorting and storage of secretory proteins in secretory granules of endocrine cells. The regulation of this process is not known. Hexahistidine epitope tags were used to create aggregation chaperones that enhance the calcium-induced aggregation of secretory granule proteins in vitro. Indeed, 100% recovery of the aggregating target protein was achieved without any modification of the target protein. The aggregation chaperone is not trapped in the aggregates. Co-expression of His(6) tagged secreted alkaline phosphatase and the regulated secretory protein chromogranin A resulted in an increased chromogranin storage in secretory granules, and stimulated secretion of chromogranin A increased 50%. However, secretion of secreted alkaline phosphatase was not affected by the hexahistidine epitope tag. Thus, calcium-induced aggregation is not a passive process; rather, aggregation and sorting of secretory proteins can be regulated by aggregation chaperones in the secretory pathway of endocrine cells. PMID- 10862759 TI - Transforming growth factor-beta 1-induced activation of the ERK pathway/activator protein-1 in human lung fibroblasts requires the autocrine induction of basic fibroblast growth factor. AB - Transforming growth factor-beta (TGF-beta) is involved in multiple processes including cell growth and differentiation. In particular, TGF-beta has been implicated in the pathogenesis of fibrotic lung diseases. In this study, we examined regulation of the mitogen-activated protein kinase pathway by TGF-beta1 in primary human lung fibroblasts. TGF-beta1 treatment resulted in extracellular signal-regulated kinase (ERK) pathway activation in a delayed manner, with maximal activity at 16 h. ERK activation occurred concomitantly with the induction of activator protein-1 (AP-1) binding, a nuclear factor required for activation of multiple genes involved in fibrosis. AP-1 binding was dependent on ERK activation, since the MEK-1 (mitogen-activated protein kinase kinase) inhibitor PD98059 inhibited TGF-beta1-induced binding. Induction of the receptor tyrosine kinase-linked growth factor, basic fibroblast growth factor (bFGF) protein expression temporally paralleled the activation of ERK/AP-1. Induction of AP-1 by TGF-beta1-conditioned medium was observed at 2 h, similar to AP-1 induction in response to exogenous bFGF. Dependence of ERK/AP-1 activation on bFGF induction was demonstrated by inhibition of TGF-beta1-induced ERK/AP-1 activation when conditioned medium from TGF-beta1-treated cells was incubated with bFGF-neutralizing antibody. Together, these results demonstrate that TGF beta1 regulates the autocrine induction of bFGF, resulting in activation of the ERK mitogen-activated protein kinase pathway and induction of AP-1 binding. PMID- 10862760 TI - Insulin responsiveness of the glucagon gene conferred by interactions between proximal promoter and more distal enhancer-like elements involving the paired domain transcription factor Pax6. AB - Regulation of gene transcription is an important aspect of insulin's action. However, the mechanisms involved are poorly understood. Insulin inhibits glucagon gene transcription, and insulin deficiency is associated with hyperglucagonemia that contributes to hyperglycemia in diabetes mellitus. Transfecting glucagon reporter fusion genes into a glucagon-producing pancreatic islet cell line, a 5' , 3'-, and internal deletion analysis, and oligonucleotide cassette insertions failed in the present study to identify a single insulin-responsive element in the glucagon gene. They rather indicate that insulin responsiveness depends on the presence of both proximal promoter elements and more distal enhancer-like elements. When the paired domain transcription factor Pax6 binding sites within the proximal promoter element G1 and the enhancer-like element G3 were mutated into GAL4 binding sites, the expression of GAL4-Pax6 and GAL4-VP16 restored basal activity, whereas only GAL4-Pax6 restored also insulin responsiveness. Likewise, GAL4-CBP activity was inhibited by insulin within the glucagon promoter context. The results suggest that insulin responsiveness is conferred to the glucagon gene by the synergistic interaction of proximal promoter and more distal enhancer-like elements, with Pax6 and its potential coactivator the CREB-binding protein being critical components. These data thereby support concepts of insulin-responsive element-independent mechanisms of insulin action to inhibit gene transcription. PMID- 10862761 TI - GATA-1 bends DNA in a site-independent fashion. AB - The DNA binding domain of GATA-1 consists of two adjacent homologous zinc fingers, of which only the C-terminal finger binds DNA independently. Solution structure studies have shown that the DNA is bent by about 15 degrees in the complex formed with the single C-terminal finger of GATA-1. The N-terminal finger stabilizes DNA binding at some sites. To determine whether it contributes to DNA bending, we have performed circular permutation DNA bending experiments with a variety of DNA-binding sites recognized by GATA-1. By using a series of full length GATA-1, double zinc finger, and single C-terminal finger constructs, we show that GATA-1 bends DNA by about 24 degrees, irrespective of the DNA-binding site. We propose that the N- and C-terminal fingers of GATA-1 adopt different orientations when bound to different cognate DNA sites. Furthermore, we characterize circular permutation bending artifacts arising from the reduced gel mobility of the protein-DNA complexes. PMID- 10862762 TI - Heterologous desensitization mediated by G protein-specific binding to caveolin. AB - We examined the notion that sequestration of G protein subunits by binding to caveolin impedes G protein reassociation and leads to transient, G protein specific desensitization of response in dispersed smooth muscle cells. Cholecystokinin octapeptide (CCK-8) and substance P (SP) were used to activate G(q/11), cyclopentyl adenosine (CPA) was used to activate G(i3), and acetylcholine (ACh) was used to activate both G(q/11) and G(i3) via m3 and m2 receptors, respectively. CCK-8 and SP increased only Galpha(q/11), and CPA increased only Galpha(i3) in caveolin immunoprecipitates; caveolin and other G proteins were not increased. ACh increased both Galpha(q/11) and Galpha(i3) in a time- and concentration-dependent fashion: only Galpha(q/11) was increased in the presence of an m2 antagonist, and only Galpha(i3) was increased in the presence of an m3 antagonist. To determine whether transient G protein binding to caveolin affected subsequent responses mediated by the same G protein, PLC-beta activity was measured in cells stimulated sequentially with two different agonists that activate either the same or a different G protein. After treatment of the cells with ACh and an m2 antagonist, the phospholipase C-beta (PLC-beta) response to CCK-8 and SP, but not CPA, was decreased; conversely, after treatment of the cells with ACh and an m3 antagonist, the PLC-beta response to CPA, but not CCK-8 or SP, was decreased. Similarly, after treatment with CCK-8 or SP, the PLC-beta response mediated by G(q/11) only was decreased, whereas after treatment with CPA, the PLC-beta response mediated by G(i3) only was decreased. A caveolin binding Galpha(q/11) fragment blocked the binding of activated Galpha(q/11) but not Galpha(i3) to caveolin-3 and prevented desensitization of the PLC-beta response mediated only by other G(q/11)-coupled receptors. A caveolin-binding Galpha(i3) fragment had the reverse effect. Thus, transient binding of receptor activated G protein subunits to caveolin impedes reassociation of the heterotrimeric species and leads to desensitization of response mediated by other receptors coupled to the same G protein. PMID- 10862763 TI - Prohibitins, stomatins, and plant disease response genes compose a protein superfamily that controls cell proliferation, ion channel regulation, and death. AB - Prohibitins, stomatins, and a group of plant defense response genes are demonstrated to belong to a novel protein superfamily. This superfamily is bound by similar primary and secondary predicted protein structures and hydropathy profiles. A PROSITE-formatted regular expression was generated that is highly predictive for identifying members of this superfamily using PHI-BLAST. The superfamily is named PID (proliferation, ion, and death) because prohibitins are involved in proliferation and cell cycle control, stomatins are involved in ion channel regulation, and the plant defense-related genes are involved in cell death. The plant defense gene family is named HIR (hypersensitive induced reaction) because its members are associated with hypersensitive reactions involving cell death and pathogen resistance. For this study, eight novel maize genes were introduced: four closely related to prohibitins (Zm-phb1, Zm-phb2, Zm phb3, and Zm-phb4), one to stomatins (Zm-stm1), and three to a gene implicated in plant disease responses (Zm-hir1, Zm-hir2, and Zm-hir3). The maize Zm-hir3 gene transcript is up-regulated in a disease lesion mimic mutation (Les9), supporting a role in maize defense responses. Members of this gene superfamily are involved in diverse functions, but their structural similarity suggests a conserved molecular mechanism, which we postulate to be ion channel regulation. PMID- 10862765 TI - The NH2-terminal region of focal adhesion kinase reconstitutes high affinity IgE receptor-induced secretion in mast cells. AB - Focal adhesion kinase (FAK) is tyrosine-phosphorylated by adherence of cells and also by FcepsilonRI aggregation in RBL-2H3 mast cells. Using phosphorylation site specific antibodies, we observed that FcepsilonRI activation in these cells led to an increase in FAK phosphorylation at the same tyrosine residues that are phosphorylated by integrin-induced activation. Previous studies in the 3B6 line, a FAK-deficient variant of the RBL-2H3 cells, suggest that FAK plays a role in FcepsilonRI-induced secretion. Stable cell lines expressing either full-length or truncated forms of FAK were isolated after transfection of the FAK-deficient 3B6 variant cells. The NH(2) domain of FAK, which lacks the enzymatic and the COOH terminal regions, was sufficient to reconstitute secretion. The different truncated forms of FAK were still tyrosine-phosphorylated after FcepsilonRI aggregation. Therefore, the kinase domain and the COOH-terminal region are not essential for FcepsilonRI-induced tyrosine phosphorylation of FAK or for secretion. Taken together, our data demonstrate that the reconstitution of secretion is dissociated from FAK activation and that the NH(2)-terminal region of FAK is the only critical element that may play a role in FcepsilonRI-induced secretion by acting as an adaptor or linker molecule. PMID- 10862764 TI - Distinctive regulatory and metabolic properties of glycogen-targeting subunits of protein phosphatase-1 (PTG, GL, GM/RGl) expressed in hepatocytes. AB - Glycogen-targeting subunits of protein phosphatase-1 facilitate interaction of the phosphatase with enzymes of glycogen metabolism. We have shown that overexpression of one member of the family, protein targeting to glycogen (PTG), causes large increases in glycogen storage in isolated hepatocytes or intact rat liver. In the current study, we have compared the metabolic and regulatory properties of PTG (expressed in many tissues), with two other members of the gene family, G(L) (expressed primarily in liver) and G(M)/R(Gl) (expressed primarily in striated muscle). Adenovirus-mediated expression of these proteins in hepatocytes led to the following key observations. 1) G(L) has the highest glycogenic potency among the three forms studied. 2) Glycogen synthase activity ratio is much higher in G(L)-overexpressing cells than in PTG or G(M)/R(Gl) overexpressing cells. Thus, at moderate levels of G(L) overexpression, glycogen synthase activity is increased by insulin treatment, but at higher levels of G(L) expression, insulin is no longer required to achieve maximal synthase activity. In contrast, cells with high levels of PTG overexpression retain dose-dependent regulation of glycogen synthesis and glycogen synthase enzyme activity by insulin. 3) G(L)- and G(M)/R(Gl)-overexpressing cells exhibit a strong glycogenolytic response to forskolin, whereas PTG-overexpressing cells are less responsive. This difference may be explained in part by a lesser forskolin induced increase in glycogen phosphorylase activity in PTG-overexpressing cells. Based on these results, we suggest that expression of either G(L) or G(M)/R(Gl) in liver of diabetic animals may represent a strategy for lowering of blood glucose levels in diabetes. PMID- 10862766 TI - Zipper-mediated oligomerization of the mixed lineage kinase SPRK/MLK-3 is not required for its activation by the GTPase cdc 42 but Is necessary for its activation of the JNK pathway. Monomeric SPRK L410P does not catalyze the activating phosphorylation of Thr258 of murine MITOGEN-ACTIVATED protein kinase kinase 4. AB - Src homology 3 domain-containing proline-rich kinase (SPRK)/mixed lineage kinase 3 is a serine/threonine kinase that has been identified as an upstream activator of the c-Jun NH(2)-terminal kinase (JNK) pathway. SPRK is capable of activating MKK4 by phosphorylation of serine and threonine residues, and mutant forms of MKK4 that lack the phosphorylation sites Ser(254) and Thr(258) block SPRK-induced JNK activation. A region of 63 amino acids following the kinase domain of SPRK is predicted to form a leucine zipper. The leucine zipper domain of SPRK has been shown to be necessary and sufficient for SPRK oligomerization, but its role in regulating activation of SPRK and downstream signaling remains unclear. In this study, we substituted a proposed stabilizing leucine residue in the zipper domain with a helix-disrupting proline to abrogate zipper-mediated SPRK oligomerization. We demonstrate that constitutively activated Cdc42 fully activates this monomeric SPRK mutant in terms of both autophosphorylation and histone phosphorylation activity and induces the same in vivo phosphorylation pattern as wild type SPRK. However, this catalytically active SPRK zipper mutant is unable to activate JNK. Our data show that the monomeric SPRK mutant fails to phosphorylate one of the two activating phosphorylation sites, Thr(258), of MKK4. These studies suggest that zipper-mediated SPRK oligomerization is not required for SPRK activation by Cdc42 but instead is critical for proper interaction and phosphorylation of a downstream target, MKK4. PMID- 10862767 TI - 14-3-3 interacts with regulator of G protein signaling proteins and modulates their activity. AB - Regulator of G protein signaling (RGS) proteins function as GTPase-activating proteins (GAPs) that stimulate the inactivation of heterotrimeric G proteins. We have recently shown that RGS proteins may be regulated on a post-translational level (Benzing, T., Brandes, R., Sellin, L., Schermer, B., Lecker, S., Walz, G., and Kim, E. (1999) Nat. Med. 5, 913-918). However, mechanisms controlling the GAP activity of RGS proteins are poorly understood. Here we show that 14-3-3 proteins associate with RGS7 and RGS3. Binding of 14-3-3 is mediated by a conserved phosphoserine located in the Galpha-interacting portion of the RGS domain; interaction with 14-3-3 inhibits the GAP activity of RGS7, depends upon phosphorylation of a conserved residue within the RGS domain, and results in inhibition of GAP function. Collectively, these data indicate that phosphorylation-dependent binding of 14-3-3 may act as molecular switch that controls the GAP activity keeping a substantial fraction of RGS proteins in a dormant state. PMID- 10862769 TI - 31P and 1H NMR studies of the effect of the counteracting osmolyte trimethylamine N-oxide on interactions of urea with ribonuclease A. AB - 31P NMR spectroscopy has been used to show that the activity of RNase A, which is lowered in the presence of urea, can be recovered with trimethylamine-N-oxide (TMAO). A 1:1 ratio of TMAO:urea was sufficient to recover the enzyme activity. (1)H nuclear Overhauser effect spectroscopy NMR studies with RNase A have shown that even at relatively low effective concentrations of TMAO, some modification of the three-dimensional structure of the biomolecule is apparent. PMID- 10862768 TI - Structural basis for the feedback regulation of Escherichia coli pantothenate kinase by coenzyme A. AB - Pantothenate kinase (PanK) is a key regulatory enzyme in the coenzyme A (CoA) biosynthetic pathway and catalyzes the phosphorylation of pantothenic acid to form phosphopantothenate. CoA is a feedback inhibitor of PanK activity by competitive binding to the ATP site. The structures of the Escherichia coli enzyme, in complex with a nonhydrolyzable analogue of ATP, 5'-adenylimido diphosphate (AMPPNP), or with CoA, were determined at 2.6 and 2.5 A, respectively. Both structures show that two dimers occupy an asymmetric unit; each subunit has a alpha/beta mononucleotide-binding fold with an extensive antiparallel coiled coil formed by two long helices along the dimerization interface. The two ligands, AMPPNP and CoA, associate with PanK in very different ways, but their phosphate binding sites overlap, explaining the kinetic competition between CoA and ATP. Residues Asp(127), His(177), and Arg(243) are proposed to be involved in catalysis, based on modeling of the pentacoordinate transition state. The more potent inhibition by CoA, compared with the CoA thioesters, is explained by a tight interaction of the CoA thiol group with the side chains of aromatic residues, which is predicted to discriminate against the CoA thioesters. The PanK structure provides the framework for a more detailed understanding of the mechanism of catalysis and feedback regulation of PanK. PMID- 10862770 TI - Ca2+ regulation of gelsolin by its C-terminal tail. AB - Gelsolin is activated by Ca(2+) to sever actin filaments. Ca(2+) regulation is conferred on the N-terminal half by the C-terminal half. This paper seeks to understand how Ca(2+) regulates gelsolin by testing the "tail helix latch hypothesis," which is based on the structural data showing that gelsolin has a C terminal tail helix that contacts the N-terminal half in the absence of Ca(2+). Ca(2+) activation of gelsolin at 37 degrees C occurs in three steps, with apparent K(d) for Ca(2+) of 0.1, 0.3, and 6.4 x 10(-6) m. Tail helix truncation decreases the apparent Ca(2+) requirement for severing to 10(-7) m and eliminates the conformational change observed at 10(-6) m Ca(2+). The large decrease in Ca(2+) requirement for severing is not due to a change in Ca(2+) binding nor to Ca(2+)-independent activation of the C-terminal half per se. Thus, the tail helix latch is primarily responsible for transmitting micromolar Ca(2+) information from the gelsolin C-terminal half to the N-terminal half. Occupation of submicromolar Ca(2+)-binding sites primes gelsolin for severing, but gelsolin cannot sever because the tail latch is still engaged. Unlatching the tail helix by 10(-6) m Ca(2+) releases the final constraint to initiate the severing cascade. PMID- 10862771 TI - Prostaglandin H synthase. Effects of peroxidase cosubstrates on cyclooxygenase velocity. AB - Many cosubstrates for the peroxidase activity of prostaglandin H synthase-1 (PGHS 1) have been reported to produce a large (2-7-fold) increase in the cyclooxygenase velocity in addition to a substantial increase in the number of cyclooxygenase catalytic turnovers. The large stimulation of cyclooxygenase velocity has become an important criterion for evaluation of putative PGHS reaction mechanisms. This criterion has been a major weakness of branched-chain tyrosyl radical mechanisms, which correctly predict many other cyclooxygenase characteristics. Our computer simulations based on a branched-chain mechanism indicated that the uncorrected oxygen electrode signals commonly used to monitor activity can seriously overestimate the effects of cosubstrate on cyclooxygenase velocity. The simulation results prompted re-examination of the effect of several cosubstrates (phenol, acetaminophen, N,N,N',N'-tetramethylphenylenediamine, and Trolox) on PGHS-1 cyclooxygenase velocity. Cyclooxygenase kinetics were examined at reduced temperature or elevated pH, where the oxygen electrode signal can be corrected to provide reliable oxygen consumption trajectories. The cosubstrates produced only a slight (10-60%) stimulation of the cyclooxygenase velocity. Peroxidase cosubstrates thus have a much smaller stimulatory effect on cyclooxygenase velocity than previously reported. This corrects a longstanding misperception of cosubstrate effects, provides more realistic kinetic constraints on PGHS mechanisms, and removes what was a major deficiency of branched-chain tyrosyl radical mechanisms. PMID- 10862772 TI - Characterization of the fasting-induced adipose factor FIAF, a novel peroxisome proliferator-activated receptor target gene. AB - Fasting is associated with significant changes in nutrient metabolism, many of which are governed by transcription factors that regulate the expression of rate limiting enzymes. One factor that plays an important role in the metabolic response to fasting is the peroxisome proliferator-activated receptor alpha (PPARalpha). To gain more insight into the role of PPARalpha during fasting, and into the regulation of metabolism during fasting in general, a search for unknown PPARalpha target genes was performed. Using subtractive hybridization (SABRE) comparing liver mRNA from wild-type and PPARalpha null mice, we isolated a novel PPARalpha target gene, encoding the secreted protein FIAF (for fasting induced adipose factor), that belongs to the family of fibrinogen/angiopoietin-like proteins. FIAF is predominantly expressed in adipose tissue and is strongly up regulated by fasting in white adipose tissue and liver. Moreover, FIAF mRNA is decreased in white adipose tissue of PPARgamma +/- mice. FIAF protein can be detected in various tissues and in blood plasma, suggesting that FIAF has an endocrine function. Its plasma abundance is increased by fasting and decreased by chronic high fat feeding. The data suggest that FIAF represents a novel endocrine signal involved in the regulation of metabolism, especially under fasting conditions. PMID- 10862773 TI - Characterization of a novel 8-oxoguanine-DNA glycosylase activity in Escherichia coli and identification of the enzyme as endonuclease VIII. AB - 8-Oxoguanine (G*), induced by reactive oxygen species, is mutagenic because it mispairs with A. The major G*-DNA glycosylase (OGG), namely, OGG1 in eukaryotes, or MutM in Escherichia coli, excises G* when paired in DNA with C, G, and T, but not A, presumably because removal of G* from a G*.A pair would be mutagenic. However, repair of G* will prevent mutation when it is incorporated in the nascent strand opposite A. This could be carried out by a second OGG, OGG2, identified in yeast and human cells. We have characterized a new OGG activity in E. coli and then identified it to be endonuclease VIII (Nei), discovered as a damaged pyrimidine-specific DNA glycosylase. Nei shares sequence homology and reaction mechanism with MutM and is similar to human OGG2 in being able to excise G* when paired with A (or G). Kinetic analysis of wild type Nei showed that it has significant activity for excising G* relative to dihydrouracil. The presence of OGG2 type enzyme in both E. coli and eukaryotes, which is at least as efficient in excising G* from a G*.A (or G) pair as from a G*.C pair, supports the possibility of G* repair in the nascent DNA strand. PMID- 10862774 TI - Domain 26 of tropoelastin plays a dominant role in association by coacervation. AB - The temperature-dependent association of tropoelastin molecules through coacervation is an essential step in their assembly leading to elastogenesis. The relative contributions of C-terminal hydrophobic domains in coacervation were assessed. Truncated tropoelastins were constructed with N termini positioned variably downstream of domain 25. The purified proteins were assessed for their ability to coacervate. Disruption to domain 26 had a substantial effect and abolished coacervation. Circular dichroism spectroscopy of an isolated peptide comprising domain 26 showed that it undergoes a structural transition to a state of increased order with increasing temperature. Protease mapping demonstrated that domain 26 is flanked by surface sites and is likely to be in an exposed position on the surface of the tropoelastin molecule. These results suggest that the hydrophobic domain 26 is positioned to play a dominant role in the intermolecular interactions that occur during coacervation. PMID- 10862775 TI - Unraveling the amino acid sequence crucial for heparin binding to collagen V. AB - We have previously shown that a recombinant 12-kDa fragment of the collagen alpha1(V) chain (Ile(824)-Pro(950)), referred to as HepV, binds to heparin and heparan sulfate (Delacoux, F., Fichard, A., Geourjon, C., Garrone, R., and Ruggiero, F. (1998) J. Biol. Chem. 273, 15069-15076). No consensus sequence was found in the alpha1(V) primary sequence, but a cluster of 7 basic amino acids (in the Arg(900)-Arg(924) region) was postulated to contain the heparin-binding site. The contribution of individual basic amino acids within this sequence was examined by site-directed mutagenesis. Further evidence for the precise localization of the heparin-binding site was provided by experiments based on the fact that heparin can protect the alpha1(V) chain heparin-binding site from trypsin digestion. The results parallel the alanine scanning mutagenesis data, i.e. heparin binding to the alpha1(V) chain involved Arg(912), Arg(918), and Arg(921) and two additional neighboring basic residues, Lys(905) and Arg(909). Our data suggest that this extended sequence functions as a heparin-binding site in both collagens V and XI, indicating that these collagens use a novel sequence motif to interact with heparin. PMID- 10862776 TI - Rho GTPases mediate the regulation of cochlear outer hair cell motility by acetylcholine. AB - Outer hair cells are the mechanical effectors of the cochlear amplifier, an active process that improves the sensitivity and frequency discrimination of the mammalian ear. In vivo, the gain of the cochlear amplifier is regulated by the efferent neurotransmitter acetylcholine through the modulation of outer hair cell motility. Little is known, however, regarding the molecular mechanisms activated by acetylcholine. In this study, intracellular signaling pathways involving the small GTPases RhoA, Rac1, and Cdc42 have been identified as regulators of outer hair cell motility. Changes in cell length (slow motility) and in the amplitude of electrically induced movement (fast motility) were measured in isolated outer hair cells patch clamped in whole-cell mode, internally perfused through the patch pipette with different inhibitors and activators of these small GTPases while being externally stimulated with acetylcholine. We found that acetylcholine induces outer hair cell shortening and a simultaneous increase in the amplitude of fast motility through Rac1 and Cdc42 activation. In contrast, a RhoA- and Rac1 mediated signaling pathway induces outer hair cell elongation and decreases fast motility amplitude. These two opposing processes provide the basis for a regulatory mechanism of outer hair cell motility. PMID- 10862777 TI - Phosphorylation of serine 43 is not required for inhibition of c-Raf kinase by the cAMP-dependent protein kinase. AB - The activity of the serine/threonine kinase c-Raf (Raf) is inhibited by increased intracellular cAMP. This is believed to require phosphorylation with the cAMP dependent protein kinase (PKA), although the mechanism by which PKA inhibits Raf is controversial. We investigated the requirement for PKA phosphorylation using Raf mutants expressed in HEK293 or NIH 3T3 cells. Phosphopeptide mapping of (32)P labeled Raf (WT) or a mutant lacking a putative PKA phosphorylation site (serine to alanine, S43A) confirmed that serine 43 (Ser(43)) was the major cAMP (forskolin)-stimulated phosphorylation site in vivo. Interestingly, the EGF stimulated Raf kinase activity of the S43A mutant was inhibited by forskolin equivalently to that of the WT Raf. Forskolin also inhibited the activation of an N-terminal deletion mutant Delta5-50 Raf completely lacking this phosphorylation site. Although WT Raf was phosphorylated by PKA, phosphorylation did not inhibit Raf catalytic activity in vitro, nor did forskolin treatment inhibit the activity of an N-terminally truncated Raf protein (Raf 22W) or a full-length Raf protein (Raf-CAAX) expressed in NIH 3T3 cells. In contrast, forskolin inhibited the EGF dependent activation of a Raf isoform (B-Raf), lacking an analogous phosphorylation site to Ser(43). Thus, these results demonstrate that PKA exerts its inhibitory effects independently of direct Raf phosphorylation and suggests instead that PKA prevents an event required for the EGF-dependent activation of Raf. PMID- 10862778 TI - Interaction with beta-arrestin determines the difference in internalization behavor between beta1- and beta2-adrenergic receptors. AB - The beta(1)-adrenergic receptor (beta(1)AR) shows the resistance to agonist induced internalization. As beta-arrestin is important for internalization, we examine the interaction of beta-arrestin with beta(1)AR with three different methods: intracellular trafficking of beta-arrestin, binding of in vitro translated beta-arrestin to intracellular domains of beta(1)- and beta(2)ARs, and inhibition of betaAR-stimulated adenylyl cyclase activities by beta-arrestin. The green fluorescent protein-tagged beta-arrestin 2 translocates to and stays at the plasma membrane by beta(2)AR stimulation. Although green fluorescent protein tagged beta-arrestin 2 also translocates to the plasma membrane, it returns to the cytoplasm 10-30 min after beta(1)AR stimulation. The binding of in vitro translated beta-arrestin 1 and beta-arrestin 2 to the third intracellular loop and the carboxyl tail of beta(1)AR is lower than that of beta(2)AR. The fusion protein of beta-arrestin 1 with glutathione S-transferase inhibits the beta(1)- and beta(2)AR-stimulated adenylyl cyclase activities, although inhibition of the beta(1)AR-stimulated activity requires a higher concentration of the fusion protein than that of the beta(2)AR-stimulated activity. These results suggest that weak interaction of beta(1)AR with beta-arrestins explains the resistance to agonist-induced internalization. This is further supported by the finding that beta-arrestin can induce internalization of beta(1)AR when beta-arrestin 1 does not dissociate from beta(1)AR by fusing to the carboxyl tail of beta(1)AR. PMID- 10862779 TI - Marrow stromal stem cells. PMID- 10862780 TI - Myogenic stem cells for the therapy of primary myopathies: wishful thinking or therapeutic perspective? PMID- 10862781 TI - The cells that knew too much. PMID- 10862782 TI - The future of allogeneic hematopoietic stem cell transplantation: minimizing pain, maximizing gain. PMID- 10862783 TI - Immune recognition of OxLDL in atherosclerosis. PMID- 10862784 TI - Suppressed smooth muscle proliferation and inflammatory cell invasion after arterial injury in elafin-overexpressing mice. AB - Elastases degrade the extracellular matrix, releasing growth factors and chemotactic peptides, inducing glycoproteins such as tenascin, and thereby promoting vascular cell proliferation and migration. Administration of serine elastase inhibitors reduces experimentally induced vascular disease. The ability to mount an intrinsic anti-elastase response may, therefore, protect against intimal/medial thickening after vascular injury. To investigate this, we showed that wire-induced endothelial denudation of the carotid artery is associated with transient elevation in elastase activity and confirmed that this is abolished in transgenic mice overexpressing the serine elastase inhibitor, elafin, targeted to the cardiovascular system. Ten days after injury, nontransgenic littermates show vessel enlargement, intimal thickening, increased medial area and cellularity, and 2-fold increase in tenascin. Injured vessels in transgenic mice become enlarged but are otherwise similar to sham-operated controls. Injury-induced vessel wall thickening, which is observed only in nontransgenic mice, is related to foci of neutrophils and macrophages, in addition to smooth muscle cells that fail to stain for alpha-actin and are likely dedifferentiated. Our study therefore suggests that a major determinant of the vascular response to injury is the early transient induction of serine elastase activity, which leads to cellular proliferation and inflammatory cell migration. PMID- 10862785 TI - IL-4 gene therapy for collagen arthritis suppresses synovial IL-17 and osteoprotegerin ligand and prevents bone erosion. AB - Bone destruction is the most difficult target in the treatment of rheumatoid arthritis (RA). Here, we report that local overexpression of IL-4, introduced by a recombinant human type 5 adenovirus vector (Ad5E1mIL-4) prevents joint damage and bone erosion in the knees of mice with collagen arthritis (CIA). No difference was noted in the course of CIA in the injected knee joints between Ad5E1mIL-4 and the control vector, but radiographic analysis revealed impressive reduction of joint erosion and more compact bone structure in the Ad5E1mIL-4 group. Although severe inflammation persisted in treated mice, Ad5E1mIL-4 prevented bone erosion and diminished tartrate-resistant acid phosphatase (TRAP) activity, indicating that local IL-4 inhibits the formation of osteoclast-like cells. Messenger RNA levels of IL-17, IL-12, and cathepsin K in the synovial tissue were suppressed, as were IL-6 and IL-12 protein production. Osteoprotegerin ligand (OPGL) expression was markedly suppressed by local IL-4, but no loss of OPG expression was noted with Ad5E1mIL-4 treatment. Finally, in in vitro studies, bone samples of patients with arthritis revealed consistent suppression by IL-4 of type I collagen breakdown. IL-4 also enhanced synthesis of type I procollagen, suggesting that it promoted tissue repair. These findings may have significant implications for the prevention of bone erosion in arthritis. PMID- 10862786 TI - Inhibition of cystic fibrosis transmembrane conductance regulator by novel interaction with the metabolic sensor AMP-activated protein kinase. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) is an ATP-gated Cl(-) channel that regulates other epithelial transport proteins by uncharacterized mechanisms. We employed a yeast two-hybrid screen using the COOH terminal 70 residues of CFTR to identify proteins that might be involved in such interactions. The alpha1 (catalytic) subunit of AMP-activated protein kinase (AMPK) was identified as a dominant and novel interacting protein. The interaction is mediated by residues 1420-1457 in CFTR and by the COOH-terminal regulatory domain of alpha1-AMPK. Mutations of two protein trafficking motifs within the 38-amino acid region in CFTR each disrupted the interaction. GST fusion protein pull-down assays in vitro and in transfected cells confirmed the CFTR-alpha1-AMPK interaction and also identified alpha2-AMPK as an interactor with CFTR. AMPK is coexpressed in CFTR-expressing cell lines and shares an apical distribution with CFTR in rat nasal epithelium. AMPK phosphorylated full-length CFTR in vitro, and AMPK coexpression with CFTR in Xenopus oocytes inhibited cAMP activated CFTR whole-cell Cl(-) conductance by approximately 35-50%. Because AMPK is a metabolic sensor in cells and responds to changes in cellular ATP, regulation of CFTR by AMPK may be important in inhibiting CFTR under conditions of metabolic stress, thereby linking transepithelial transport to cell metabolic state. PMID- 10862787 TI - Deactivation of peroxisome proliferator-activated receptor-alpha during cardiac hypertrophic growth. AB - We sought to delineate the molecular regulatory events involved in the energy substrate preference switch from fatty acids to glucose during cardiac hypertrophic growth. alpha(1)-adrenergic agonist-induced hypertrophy of cardiac myocytes in culture resulted in a significant decrease in palmitate oxidation rates and a reduction in the expression of the gene encoding muscle carnitine palmitoyltransferase I (M-CPT I), an enzyme involved in mitochondrial fatty acid uptake. Cardiac myocyte transfection studies demonstrated that M-CPT I promoter activity is repressed during cardiac myocyte hypertrophic growth, an effect that mapped to a peroxisome proliferator-activated receptor-alpha (PPARalpha) response element. Ventricular pressure overload studies in mice, together with PPARalpha overexpression studies in cardiac myocytes, demonstrated that, during hypertrophic growth, cardiac PPARalpha gene expression falls and its activity is altered at the posttranscriptional level via the extracellular signal-regulated kinase mitogen-activated protein kinase pathway. Hypertrophied myocytes exhibited reduced capacity for cellular lipid homeostasis, as evidenced by intracellular fat accumulation in response to oleate loading. These results indicate that during cardiac hypertrophic growth, PPARalpha is deactivated at several levels, leading to diminished capacity for myocardial lipid and energy homeostasis. PMID- 10862788 TI - Natural antibodies with the T15 idiotype may act in atherosclerosis, apoptotic clearance, and protective immunity. AB - The immune response to oxidized LDL (OxLDL) may play an important role in atherogenesis. Working with apoE-deficient mice, we isolated a panel of OxLDL specific B-cell lines that secrete IgM Abs that specifically bind to oxidized phospholipids such as 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3 phosphorylcholine (POVPC). These Abs block uptake of OxLDL by macrophages, recognize similar oxidation-specific epitopes on apoptotic cells, and are deposited in atherosclerotic lesions. The Abs were found to be structurally and functionally identical to classic "natural" T15 anti-PC Abs that are of B-1 cell origin and are reported to provide optimal protection from virulent pneumococcal infection. These findings suggest that there has been natural selection for B-1 cells secreting oxidation-specific/T15 antibodies, both for their role in natural immune defense and for housekeeping roles against oxidation-dependent neodeterminants in health and disease. PMID- 10862789 TI - Abnormal functional and morphological regulation of the gastric mucosa in histamine H2 receptor-deficient mice. AB - To clarify the physiological roles of histamine H2 receptor (H2R), we have generated histamine H2R-deficient mice by gene targeting. Homozygous mutant mice were viable and fertile without apparent abnormalities and, unexpectedly, showed normal basal gastric pH. However, the H2R-deficient mice exhibited a marked hypertrophy with enlarged folds in gastric mucosa and an elevated serum gastrin level. Immunohistochemical analysis revealed increased numbers of parietal and enterochromaffin-like (ECL) cells. Despite this hypertrophy, parietal cells in mutant mice were significantly smaller than in wild-type mice and contained enlarged secretory canaliculi with a lower density of microvilli and few typical tubulovesicles in the narrow cytoplasm. Induction of gastric acid secretion by histamine or gastrin was completely abolished in the mutant mice, but carbachol still induced acid secretion. The present study clearly demonstrates that H2R mediated signal(s) are required for cellular homeostasis of the gastric mucosa and normally formed secretory membranes in parietal cells. Moreover, impaired acid secretion due to the absence of H2R could be overcome by the signals from cholinergic receptors. PMID- 10862790 TI - A role for NF-kappaB-dependent gene transactivation in sunburn. AB - Exposure of skin to ultraviolet (UV) radiation is known to induce NF-kappaB activation, but the functional role for this pathway in UV-induced cutaneous inflammation remains uncertain. In this study, we examined whether experimentally induced sunburn reactions in mice could be prevented by blocking UV-induced, NF kappaB-dependent gene transactivation with oligodeoxynucleotides (ODNs) containing the NF-kappaB cis element (NF-kappaB decoy ODNs). UV-induced secretion of IL-1, IL-6, TNF-alpha, and VEGF by skin-derived cell lines was inhibited by the decoy ODNs, but not by the scrambled control ODNs. Systemic or local injection of NF-kappaB decoy ODNs also inhibited cutaneous swelling responses to UV irradiation. Moreover, local UV-induced inflammatory changes (swelling, leukocyte infiltration, epidermal hyperplasia, and accumulation of proinflammatory cytokines) were all inhibited specifically by topically applied decoy ODNs. Importantly, these ODNs had no effect on alternative types of cutaneous inflammation caused by irritant or allergic chemicals. These results indicate that sunburn reactions culminate from inflammatory events that are triggered by UV-activated transcription of NF-kappaB target genes, rather than from nonspecific changes associated with tissue damage. PMID- 10862791 TI - CD4(+) Valpha14 natural killer T cells are essential for acceptance of rat islet xenografts in mice. AB - Pancreatic islet transplantation represents a potential treatment for insulin dependent diabetes mellitus. However, the precise cellular and molecular mechanisms of the immune reactions against allogeneic and xenogeneic transplanted islets remain unclear. Here, we demonstrate that CD4(+) Valpha14 natural killer T (NKT) cells, a recently identified lymphoid cell lineage, are required for the acceptance of intrahepatic rat islet xenografts. An anti-CD4 mAb, administrated after transplantation, allowed islet xenografts to be accepted by C57BL/6 mice, with no need for immunosuppressive drugs. The dose of anti-CD4 mAb was critical, and the beneficial effect appeared to be associated with the reappearance of CD4(+) NKT cells at around 14 days after transplantation. Interestingly, rat islet xenografts were rejected, despite the anti-CD4 mAb treatment, in Valpha14 NKT cell-deficient mice, which exhibit the normal complement of conventional lymphoid cells; adoptive transfer of Valpha14 NKT cells into Valpha14 NKT cell deficient mice restored the acceptance of rat islet xenografts. In addition, rat islet xenografts were accepted by Valpha14 NKT mice having only Valpha14 NKT cells and no other lymphoid cells. These results indicate that Valpha14 NKT cells play a crucial role in the acceptance of rat islet xenografts in mice treated with anti-CD4 antibody, probably by serving as immunosuppressive regulatory cells. PMID- 10862792 TI - Enteroaggregative Escherichia coli expresses a novel flagellin that causes IL-8 release from intestinal epithelial cells. AB - Enteroaggregative Escherichia coli (EAEC) is an emerging cause of acute and persistent diarrhea worldwide. EAEC infections are associated with intestinal inflammation and growth impairment in infected children, even in the absence of diarrhea. We previously reported that prototype EAEC strains rapidly induce IL-8 production by Caco-2 intestinal epithelial cells, and that this effect is mediated by a soluble, heat-stable factor released by these bacteria in culture. We herein report the cloning, sequencing, and expression of this biologically active IL-8-releasing factor from EAEC, and its identification as a flagellin that is unique among known expressed proteins. Flagella purified from EAEC 042 and several other EAEC isolates potently release IL-8 from Caco-2 cells; an engineered aflagellar mutant of 042 does not release IL-8. Finally, cloned EAEC flagellin expressed in nonpathogenic E. coli as a polyhistidine-tagged fusion protein maintains its proinflammatory activity. These findings demonstrate a major new means by which EAEC may cause intestinal inflammation, persistent diarrhea, and growth impairment that characterize human infection with these organisms. Furthermore, they open new approaches for diagnosis and vaccine development. This novel pathogenic mechanism of EAEC extends an emerging paradigm of bacterial flagella as inflammatory stimuli. PMID- 10862793 TI - Mixed chimerism and tolerance without whole body irradiation in a large animal model. AB - Mixed hematopoietic chimerism may provide a treatment for patients with nonmalignant hematologic diseases, and may tolerize patients to organ allografts without requiring chronic immunosuppression. However, the toxicity of the usual conditioning regimens has limited the clinical applicability of this approach. These regimens generally include some level of whole body irradiation (WBI), which is thought to facilitate engraftment either by making room for donor hematopoietic stem cells or by providing sufficient host immunosuppression to enable donor cells to engraft. Here, we have established mixed chimerism across both minor and major histocompatibility barriers in swine, by using high doses of peripheral blood stem cells in the absence of WBI. After mixed chimerism was established, swine leukocyte antigen-matched (SLA-matched) donor skin grafts were tolerated and maintained for a prolonged period, whereas third-party SLA-matched skin was rejected promptly. Donor-matched kidney allografts were also accepted without additional immunosuppression. Because of its low toxicity, this approach has potential for a wide range of clinical applications. Our data may indicate that niches for engrafting stem cells are filled by mass action and that WBI, which serves to empty some of these niches, can be omitted if the donor inoculum is sufficiently large and if adequate host T-cell depletion is achieved before transplant. PMID- 10862794 TI - Redistribution of substrates to adipose tissue promotes obesity in mice with selective insulin resistance in muscle. AB - Obesity and insulin resistance in skeletal muscle are two major factors in the pathogenesis of type 2 diabetes. Mice with muscle-specific inactivation of the insulin receptor gene (MIRKO) are normoglycemic but have increased fat mass. To identify the potential mechanism for this important association, we examined insulin action in specific tissues of MIRKO and control mice under hyperinsulinemic-euglycemic conditions. We found that insulin-stimulated muscle glucose transport and glycogen synthesis were decreased by about 80% in MIRKO mice, whereas insulin-stimulated fat glucose transport was increased threefold in MIRKO mice. These data demonstrate that selective insulin resistance in muscle promotes redistribution of substrates to adipose tissue thereby contributing to increased adiposity and development of the prediabetic syndrome. PMID- 10862795 TI - Distinct roles for the NF-kappaB1 (p50) and c-Rel transcription factors in inflammatory arthritis. AB - Rheumatoid arthritis (RA) is a complex disease, with contributions from systemic autoimmunity and local inflammation. Persistent synovial joint inflammation and invasive synovial pannus tissue lead to joint destruction. RA is characterized by the production of inflammatory mediators, many of which are regulated by the Rel/NF-kappaB transcription factors. Although an attractive target for therapeutic intervention in inflammatory diseases, Rel/NF-kappaB is involved in normal physiology, thus global inhibition could be harmful. An alternate approach is to identify and target the Rel/NF-kappaB subunits critical for components of disease. To assess this, mice with null mutations in c-rel or nfkb1 were used to examine directly the roles of c-Rel and p50 in models of acute and chronic inflammatory arthritis. We found c-Rel-deficient mice were resistant to collagen induced arthritis but had a normal response in an acute, destructive arthritis model (methylated BSA/IL-1 induced arthritis) suggesting c-Rel is required for systemic but not local joint disease. In contrast, p50-deficient mice were refractory to induction of both the chronic and acute arthritis models, showing this subunit is essential for local joint inflammation and destruction. Our data suggest Rel/NF-kappaB subunits play distinct roles in the pathogenesis of inflammatory arthritis and may provide a rationale for more specific therapeutic blockade of Rel/NF-kappaB in RA. PMID- 10862796 TI - Delayed catabolism of apoB-48 lipoproteins due to decreased heparan sulfate proteoglycan production in diabetic mice. AB - We used wild-type (WT) mice and mice engineered to express either apoB-100 only (B100 mice) or apoB-48 only (B48 mice) to examine the effects of streptozotocin induced diabetes (DM) on apoB-100- and apoB-48-containing lipoproteins. Plasma lipids increased with DM in WT mice, and fat tolerance was markedly impaired. Lipoprotein profiles showed increased levels and cholesterol enrichment of VLDL in diabetic B48 mice but not in B100 mice. C apolipoproteins, in particular apoC I in VLDL, were increased. To investigate the basis of the increase in apoB-48 lipoproteins in streptozotocin-treated animals, we characterized several parameters of lipoprotein metabolism. Triglyceride and apoB production rates were normal, as were plasma lipase activity, VLDL glycosaminoglycan binding, and VLDL lipolysis. However, beta-VLDL clearance decreased due to decreased trapping by the liver. Whereas LRP activity was normal, livers from treated mice incorporated significantly less sulfate into heparan sulfate proteoglycans (HSPG) than did controls. Hepatoma (HepG2) cells and endothelial cells cultured in high glucose also showed decreased sulfate and glucosamine incorporation into HSPG. Western blots of livers from diabetic mice showed a decrease in the HSPG core protein, perlecan. Delayed clearance of postprandial apoB-48-containing lipoproteins in DM appears to be due to decreased hepatic perlecan HSPG. PMID- 10862797 TI - Altered expression of fatty acid-metabolizing enzymes in aromatase-deficient mice. AB - Hepatic steatosis is a frequent complication in nonobese patients with breast cancer treated with tamoxifen, a potent antagonist of estrogen. In addition, hepatic steatosis became evident spontaneously in the aromatase-deficient (ArKO) mouse, which lacks intrinsic estrogen production. These clinical and laboratory observations suggest that estrogen helps to maintain constitutive lipid metabolism. To clarify this hypothesis, we characterized the expression and activity in ArKO mouse liver of enzymes involved in peroxisomal and mitochondrial fatty acid beta-oxidation. Northern analysis showed reduced expression of mRNAs for very long fatty acyl-CoA synthetase, peroxisomal fatty acyl-CoA oxidase, and medium-chain acyl-CoA dehydrogenase, enzymes required in fatty acid beta oxidation. In vitro assays of fatty acid beta-oxidation activity using very long (C24:0), long (C16:0), or medium (C12:0) chain fatty acids as the substrates confirmed that the corresponding activities are also diminished. Impaired gene expression and enzyme activities of fatty acid beta-oxidation were restored to the wild-type levels, and hepatic steatosis was substantially diminished in animals treated with 17beta-estradiol. Wild-type and ArKO mice showed no difference in the binding activities of the hepatic nuclear extracts to a peroxisome proliferator response element. These findings demonstrate the pivotal role of estrogen in supporting constitutive hepatic expression of genes involved in lipid beta-oxidation and in maintaining hepatic lipid homeostasis. PMID- 10862798 TI - Two defects contribute to hypothalamic leptin resistance in mice with diet induced obesity. AB - Obesity in humans and in rodents is usually associated with high circulating leptin levels and leptin resistance. To examine the molecular basis for leptin resistance, we determined the ability of leptin to induce hypothalamic STAT3 (signal transducer and activator of transcription) signaling in C57BL/6J mice fed either low-fat or high-fat diets. In mice fed the low-fat diet, leptin activated STAT3 signaling when administered via the intraperitoneal (ip) or the intracerebroventricular (icv) route, with the half-maximal dose being 30-fold less when given by the icv route. The high-fat diet increased body-weight gain and plasma leptin levels. After 4 weeks on the diet, hypothalamic STAT3 signaling after ip leptin administration was equivalent in both diet groups. In contrast, peripherally administered leptin was completely unable to activate hypothalamic STAT3 signaling, as measured by gel shift assay after 15 weeks of high-fat diet. Despite the absence of detectable signaling after peripheral leptin at 15 weeks, the mice fed the high-fat diet retained the capacity to respond to icv leptin, although the magnitude of STAT3 activation was substantially reduced. These results suggest that leptin resistance induced by a high-fat diet evolves during the course of the diet and has at least two independent causes: an apparent defect in access to sites of action in the hypothalamus that markedly limits the ability of peripheral leptin to activate hypothalamic STAT signaling, and an intracellular signaling defect in leptin-responsive hypothalamic neurons that lies upstream of STAT3 activation. PMID- 10862799 TI - Enhanced RANK ligand expression and responsivity of bone marrow cells in Paget's disease of bone. AB - Paget's disease is characterized by highly localized areas of increased osteoclast (OCL) activity. This suggests that the microenvironment in pagetic lesions is highly osteoclastogenic, or that OCL precursors in these lesions are hyperresponsive to osteoclastogenic factors (or both). To examine these possibilities, we compared RANK ligand (RANKL) mRNA expression in a marrow stromal cell line developed from a pagetic lesion (PSV10) with that in a normal stromal cell line (Saka), and expression in marrow samples from affected bones of Paget's patients with that in normal marrow. RANKL mRNA was increased in PSV10 cells and pagetic marrow compared with Saka cells and normal marrow, and was also increased in marrow from affected bones compared with uninvolved bones from Paget's patients. Furthermore, pagetic marrow cells formed OCLs at much lower RANKL concentrations than did normal marrow. Anti-IL-6 decreased the RANKL responsivity of pagetic marrow to normal levels, whereas addition of IL-6 to normal marrow enhanced RANKL responsivity. Thus, RANKL expression and responsivity is increased in pagetic lesions, in part mediated by IL-6. These data suggest that the combination of enhanced expression of RANKL in affected bones and increased RANKL sensitivity of pagetic OCL precursors may contribute to the elevated numbers of OCLs in Paget's disease. PMID- 10862800 TI - The serotonin transporter in the midbrain of suicide victims with major depression. AB - BACKGROUND: The involvement of serotonin in depression and suicide has been proposed, because major depression is successfully treated by medications that specifically block the serotonin transporter, and there is evidence for a decrease in serotonin transporters in major depression and suicide. The midbrain dorsal raphe nucleus (DR) has been implicated as a site for diminished serotonergic activity in that suicide victims with major depression have a significant increase in serotonin-1A autoreceptors in the DR. METHODS: [(3)H]Paroxetine was used to label the serotonin transporter in the subnuclei of the DR at several rostral-to-caudal levels of the midbrain in ten pairs of suicide victims with major depression and age-matched psychiatrically normal control subjects. RESULTS: There was a significant increase in serotonin transporters in the entire DR progressing from rostral-to-caudal levels in both normal control subjects and suicide victims with major depression. At comparable rostral-to-caudal levels, there were no significant differences in [(3)H]paroxetine binding between depressed suicide victims and normal control subjects in either the entire DR or its constituent subnuclei. CONCLUSIONS: The pathophysiology of serotonin mechanisms in suicide victims with major depression does not appear to involve alterations in the binding of [(3)H]paroxetine to the serotonin transporter in the midbrain DR. PMID- 10862801 TI - Open-label adjunctive topiramate in the treatment of bipolar disorders. AB - BACKGROUND: To preliminarily explore the spectrum of effectiveness and tolerability of the new antiepileptic drug topiramate in bipolar disorder, we evaluated the response of 56 bipolar outpatients in the Stanley Foundation Bipolar Outcome Network (SFBN) who had been treated with adjunctive topiramate in an open-label, naturalistic fashion. METHODS: In this case series, response to topiramate was assessed every 2 weeks for the first 3 months according to standard ratings in the SFBN, and monthly thereafter while patients remained on topiramate. Patients' weights, body mass indices (BMIs), and side effects were also assessed. RESULTS: Of the 54 patients who completed at least 2 weeks of open label, add-on topiramate treatment, 30 had manic, mixed, or cycling symptoms, 11 had depressed symptoms, and 13 were relatively euthymic at the time topiramate was begun. Patients who had been initially treated for manic symptoms displayed significant reductions in standard ratings scores after 4 weeks, after 10 weeks, and at the last evaluation. Those patients who were initially depressed or treated while euthymic showed no significant changes. Patients as a group displayed significant decreases in weight and BMI from topiramate initiation to week 4, to week 10, and to the last evaluation. The most common adverse side effects were neurologic and gastrointestinal. CONCLUSIONS: These preliminary open observations of adjunctive topiramate treatment suggest that it may have antimanic or anticycling effects in some patients with bipolar disorder, and may be associated with appetite suppression and weight loss that is often viewed as beneficial by the patient and clinician. Controlled studies of topiramate's acute and long-term efficacy and side effects in bipolar disorder appear warranted. PMID- 10862802 TI - Family history and symptom levels during treatment for bipolar I affective disorder. AB - BACKGROUND: Studies of family history and lithium response in patients with bipolar affective disorder have produced mixed results, but the majority have shown relationships between the presence of affective disorder among relatives and positive responses to lithium in probands. The analysis presented here sought to confirm and to further characterize such relationships. METHODS: Subjects described here participated in a long-term, prospective follow-up; had a history of Research Diagnostic Criteria manic disorder or schizoaffective disorder, manic type; and took lithium for periods of 26 weeks or longer. The majority participated in a family study in which first-degree relatives were directly interviewed. Morbidity during lithium and during anticonvulsant trials was quantified in alternative ways, as were the risks among first-degree relatives for bipolar I and nonbipolar affective disorders. RESULTS: Familial loading for bipolar affective disorder was not associated with better outcomes during lithium treatment. Rather, the presence of major depressive disorder (MDD) among relatives was associated with slower improvement during acute treatment and with higher symptom levels during continuing treatment. Findings for morbidity during anticonvulsant treatment were similar. The patients who experienced symptom persistence with lithium did so as well during periods of anticonvulsant treatment and during periods without thymoleptics. CONCLUSIONS: A family history of MDD may have an enduring and negative prognostic significance that manifests across treatment conditions. Though difficult to reconcile with several earlier studies, these findings invite replication and further exploration. PMID- 10862803 TI - Increased neurogenesis in a model of electroconvulsive therapy. AB - BACKGROUND: Electroconvulsive therapy (ECT) is a widely used and efficient treatment modality in psychiatry, although the basis for its therapeutic effect is still unknown. Past research has shown seizure activity to be a regulator of neurogenesis in the adult brain. This study examines the effect of a single and multiple electroconvulsive seizures on neurogenesis in the rat dentate gyrus. METHODS: Rats were given either a single or a series of 10 electroconvulsive seizures. At different times after the seizures, a marker of proliferating cells, Bromodeoxyuridine (BrdU), was administered to the animals. Subsequently, newborn cells positive for BrdU were counted in the dentate gyrus. Double staining with a neuron-specific marker indicated that the newborn cells displayed a neuronal phenotype. RESULTS: A single electroconvulsive seizure significantly increased the number of new born cells in the dentate gyrus. These cells survived for at least 3 months. A series of seizures further increased neurogenesis, indicating a dose-dependent mechanism. CONCLUSIONS: We propose that generation of new neurons in the hippocampus may be an important neurobiologic element underlying the clinical effects of electroconvulsive seizures. PMID- 10862804 TI - 5-HT(1A) agonist potential of pindolol: electrophysiologic studies in the dorsal raphe nucleus and hippocampus. AB - BACKGROUND: The ability of pindolol to block 5-HT(1A) autoreceptors on serotonin containing neurons in the raphe nuclei is thought to underlie the clinical reports of enhanced efficacy and rate of improvement in depressed patients treated with pindolol/selective serotonin reuptake inhibitor (SSRI) combinations. Selectivity for somatodendritic 5-HT(1A) autoreceptors is a crucial requirement, as blockade of postsynaptic 5-HT(1A) sites may jeopardize the therapeutic response. Previous investigators have probed the effects of pindolol on serotonergic dorsal raphe cell firing in animal species; here we confirm their findings and extend them to include observations on postsynaptic 5-HT(1A) receptors in the hippocampus. METHODS: Extracellular single-unit recordings were made in rats using standard electrophysiologic techniques. Firing rates of serotonin-containing neurons in the dorsal raphe nucleus and CA3 hippocampal pyramidal neurons were monitored and the effects of pindolol given alone or in combination with an SSRI (fluoxetine) or a 5-HT(1A) antagonist (WAY-100,635) were determined. RESULTS: Pindolol inhibited the firing rates of serotonergic dorsal raphe neurons in a dose-dependent manner. Recovery to baseline firing rates was gradual, but this inhibition could be acutely reversed by WAY-100,635. A range of pindolol doses failed to block the inhibitory effects of fluoxetine on dorsal raphe cell firing. In the hippocampus, pindolol also inhibited cell firing as a function of dose, although these effects were insensitive to WAY-100,635 treatment. CONCLUSIONS: The ability of pindolol to inhibit serotonergic dorsal raphe cell firing is indicative of its agonist potential and is consistent with previous studies. The lack of observable antagonism of the SSRI-induced slowing of raphe unit activity casts doubt on the suitability of this mechanism of action to account for the positive findings in clinical studies utilizing pindolol/SSRI combinations. The 5-HT(1A)-independent inhibition of hippocampal CA3 cell firing by pindolol suggests that this compound invokes multiple pharmacologic actions, all of which need to be assimilated into any proposed mechanism of action. PMID- 10862805 TI - Hippocampus and entorhinal cortex in frontotemporal dementia and Alzheimer's disease: a morphometric MRI study. AB - BACKGROUND: Magnetic resonance imaging (MRI) of hippocampal atrophy is a sensitive but not specific method to support the clinical diagnosis of early Alzheimer's disease (AD). We recently described our findings that atrophy of the entorhinal cortex (ERC) in frontotemporal dementia (FTD) is equal to that found in AD but that hippocampal atrophy in FTD is less than that found in AD. The MRI volumes of these structures provide a topographic representation of the region of interest. We hypothesized that two different dementias with distinct histopathologic and clinical features might, in addition to quantitative patterns, display topographically different patterns of atrophy. METHODS: We adopted a morphometric approach to monitor the pattern of atrophy of the hippocampus and the ERC by computing two-dimensional profiles from MRI volumes of the structures in control subjects and patients with FTD and AD. RESULTS: Compared with control subjects, atrophy of the hippocampus in patients with AD was diffuse. In patients with FTD, atrophy of the hippocampus was localized predominantly in the anterior hippocampus, suggesting a different pattern of hippocampal atrophy in FTD compared with AD. The amount and pattern of atrophy of the entorhinal cortex was virtually equal in both demented groups. CONCLUSIONS: This study provides novel data on the nature of medial temporal lobe atrophy in FTD. Morphometric MRI may be a useful technique for characterizing different patterns of atrophy in primary degenerative dementias in vivo. PMID- 10862806 TI - Frontal P300 decrements, alcohol dependence, and antisocial personality disorder. AB - BACKGROUND: The purpose of this study was to examine the independent and interactive effects of alcohol dependence, antisocial personality disorder (ASPD), and age on brain function. METHODS: P300 event-related potentials (ERPs) were recorded from 393 alcohol-dependent and 170 non-alcohol-dependent adults while they performed a visual oddball task. The two subject groups were further subdivided based upon age and the presence/absence of ASPD. RESULTS: Alcohol dependence was associated with a significant P300 amplitude decrement at anterior electrode sites only. Antisocial personality disorder was also associated with reduced P300 amplitudes at anterior electrode sites; however, the effects were only significant among subjects 30 years of age or younger. To validate this association between ASPD and P300 amplitude a correlational analysis was performed; the correlation between anterior P300 amplitude and the total number of childhood conduct disorder and adult ASPD symptoms was significant. CONCLUSIONS: The P300 amplitude decrement found at anterior electrode sites among subjects with ASPD is consistent with the results of numerous ERP, neuroimaging, or neuropsychologic studies of anterior brain function. Our study is unique in suggesting that the effects of ASPD on anterior brain function are best detected during early adulthood. The study also suggests that the detrimental neurophysiologic effects of alcohol dependence predominantly involve the anterior brain. PMID- 10862807 TI - Thrice-weekly versus daily buprenorphine maintenance. AB - BACKGROUND: Buprenorphine is a promising alternative to methadone or levo-acetyl alpha methadol for opioid agonist maintenance treatment, and thrice-weekly dosing would facilitate its use for this purpose. METHODS: After a 3-day induction, opioid-dependent patients (n = 92) were randomly assigned to daily clinic attendance and 12-weeks maintenance treatment with sublingual buprenorphine administered double blind either daily (n = 45; 16 mg/70 kg) or thrice weekly (n = 47; 34 mg/70 kg on Fridays and Sundays and 44 mg/70 kg on Tuesdays). Outcome measures include retention, results of 3x/week urine toxicology tests, and weekly self-reported illicit drug use. RESULTS: There were no significant differences at baseline in important social, demographic, and drug-use features. Retention was 71% in the daily and 77% in the 3x/week conditions. The proportion of opioid positive urine tests decreased significantly from baseline in both groups and averaged 57% (daily) and 58% in 3x/week. There were no significant differences between groups in self-reported number of bags of heroin used for any day of the week, including Thursdays (48-72 hours following the last buprenorphine dose for subjects in the 3x/week condition), or in medication compliance (92%, 91%) and counseling attendance (82%, 82%). CONCLUSIONS: At an equivalent weekly dose of 112 mg/70 kg, thrice-weekly and daily sublingual buprenorphine appear comparable in efficacy with regard to retention and reductions in illicit opioid and other drug use. These findings support the potential for utilizing thrice-weekly buprenorphine dosing in novel settings. PMID- 10862808 TI - Disulfiram versus placebo for cocaine dependence in buprenorphine-maintained subjects: a preliminary trial. AB - BACKGROUND: We examined the effects of disulfiram versus placebo on cocaine dependence in buprenorphine-maintained subjects. METHODS: Opioid and cocaine dependent subjects (n = 20) were induced onto buprenorphine maintenance, then randomized to disulfiram (250 mg q.d. ; n = 11) or placebo (n = 9) treatment for 12 weeks. RESULTS: Groups were comparable at baseline on demographic measures and on baseline measures of drug-use severity. Fifteen subjects completed the study, including 8 subjects randomized to disulfiram (72.7%) and 7 subjects randomized to placebo (77.8%). The total number of weeks abstinent from cocaine was significantly greater on disulfiram versus placebo (mean +/- SD: 7.8 +/- 2.6 vs. 3.3 +/- 0.5, p <.05) and the number of days to achieving 3 weeks (24.6 +/- 15.1 vs. 57.8 +/- 7.7, p <.01) of continuous cocaine abstinence was significantly lower in disulfiram compared with placebo. The number of cocaine-negative urine tests during the trial were also higher on disulfiram (14.7) than on placebo (8.6); furthermore, subjects in the disulfiram group achieved consistently higher rates of cocaine-negative urine tests in each 3-week interval and the increase over time was faster in the disulfiram compared with placebo. CONCLUSIONS: This preliminary study suggests the potential efficacy of disulfiram versus placebo for treatment of cocaine dependence in buprenorphine-maintained patients. PMID- 10862809 TI - Hippocampal volume in primary unipolar major depression: a magnetic resonance imaging study. AB - BACKGROUND: Previous studies have shown that major depression is frequently accompanied by hypercortisolemia. There is some evidence suggesting that an increase in the glucocorticoid levels may make hippocampal cells more vulnerable to insults caused by hypoxia, hypoglycemia, or excitatory neurotransmitters. Using magnetic resonance imaging (MRI), the hippocampi of patients with major depression were measured and compared with values observed in control subjects. METHODS: Thirty-eight patients with primary unipolar major depression were recruited. Twenty control subjects were matched for age, gender, and years of education. The hippocampal volume was measured from coronal MRI scans in all of the subjects. Patients were also grouped and compared as responders and nonresponders to treatment with fluoxetine of 20 mg/day, for 8 weeks. Hamilton Depression Rating Scale (HDRS) was used to determine the severity of depression. RESULTS: No significant differences were observed between the hippocampal volumes of patients with major depression and control subjects; however, a significant correlation was observed between the left hippocampal volume of men and their HDRS baseline values. In addition, female responders had a statistically significant higher mean right hippocampal volume than nonresponders. CONCLUSIONS: The results of our study indicate no reduction in the volume of the hippocampus in patients with major depression. Nonetheless, the results do suggest that the effects of disease severity, gender, and treatment response may influence hippocampal volume. PMID- 10862811 TI - Membrane effects of trifluoperazine, dibucaine and praziquantel on human erythrocytes. AB - Trifluoperazine (TFP) is a potent antipsychotic agent, dibucaine (DBC) is a local anaesthetic and praziquantel (PZQ) is a highly effective agent against schistosomiasis. The present work was conducted to (i) investigate the cytotoxic effects of TFP, DBC and PZQ on human erythrocyte membranes; and (ii) compare the alterations induced by the cationic drugs (TFP and DBC) with those induced by the uncharged compound (PZQ), in an attempt to have a better insight on the pathways of each drug-membrane interaction. The erythrocyte morphological alterations induced by sublytic concentrations of TFP, DBC and PZQ were evaluated by scanning electron microscopy and expressed quantitatively by the morphological index. Haemolysis and release of membrane lipids (phospholipids and cholesterol) produced by selected concentrations of TFP, DBC and PZQ, were compared with those resulting from the corresponding triple concentrations of each drug. Our results showed that the uncharged molecule of PZQ induces the same morphological alterations (stomatocytosis) as the cationic drugs TFP and DBC. Haemolysis was shown to vary with the drug used and to be concentration-dependent, with values approximately 10-fold more elevated for TFP and DBC than for PZQ, which revealed a maximum of 6% haemolysis for the highest concentration tested. Different concentration-response curves were obtained for lipid elution, although the profiles of cholesterol and phospholipids released were similar for all drugs. Nevertheless, at a fixed rate of 50% haemolysis, TFP induced a approximately 2 fold increment in the elution of cholesterol when compared with that produced by DBC (P<0. 05). The different effects induced by TFP, DBC and PZQ on erythrocyte morphology, haemolysis and lipid exfoliation are related to the physical and chemical characteristics of each compound. These results suggest that distinct cell membrane interaction pathways lead to drug-specific mechanisms of cytotoxicity. PMID- 10862812 TI - Sensitivity of cytokinesis to hydrophobic interactions. Chemical induction of bi and multinucleated cells. AB - We have tested whether cytokinesis is as sensitive to hydrophobic interactions as karyokinesis, and evaluated the usefulness of the frequency of binucleated cells as end-point. Treating cultured cells for 2 or 24 h, with different lipophilic alcohols and chlorinated hydrocarbons made this possible. Colcemid and cytochalasin B were applied as positive controls for inhibition of karyokinesis and cytokinesis, respectively. Several-fold increases of binucleated cells could be seen with cytochalasin B after 2 h of treatment, while there was no increase with colcemid, which instead blocked cells in prometaphase/metaphase. The solvent acted primarily through hydrophobic interactions. For each solvent, the blocking of cells in prometaphase/metaphase and a minor increase in binucleated cells, were seen at approximately the same concentration; the binucleated cells probably emanated from cells in anaphase/telophase at the start of treatment. We conclude that the spindle function and cleavage show similar sensitivity to hydrophobic interactions. After prolonged treatment, allowing escape from the metaphase block, the solvents induced binucleated and multinucleated cells. By forming the quotient between multinucleated (MULTI) and binucleated (BIN) cells one could distinguish between effects primarily on the spindle or cytokinesis, respectively. All solvents, and a combination of colcemid and cytochalasin B, showed quotients intermediate between those observed with colcemid (high MULTI/BIN) and cytochalasin B (low MULTI/BIN), respectively. Both protocols revealed the same relationship between lowest active concentration and lipophilicity for the solvents, implying that concentration, not dose were of prime importance. The specific inhibitors acted at low concentrations in relation to lipophilicity, clearly demonstrating their chemical mechanisms. This approach can be used for rapid screening of potential aneugens, distinguishing between routes, and when lipophilicity is known, also reveal the principal mechanism of action, i.e. physico-chemical or chemical. PMID- 10862813 TI - DNA damage induced by catechol derivatives. AB - We investigated the effect of catechol derivatives, including dopa, dopamine, adrenaline and noradrenaline, on DNA damage and the mechanisms of DNA strand breakage and formation of 8-hydroxyguanine (8HOG). The catechol derivatives caused strand breakage of plasmid DNA in the presence of ADP-Fe(3+). The DNA damage was prevented by catalase, mannitol and dimethylsulfoxide, suggesting hydroxyl radical (HO..)-like species are involved in the strand breakage of DNA. Iron chelators, such as desferrioxamine and bathophenanthroline, and reduced glutathione also inhibited the DNA damage. Deoxyribose, a molecule that is used to detect HO,, was not degraded by dopa in the presence of ADP-Fe(3+). By adding EDTA, however, dopa induced the marked deoxyribose degradation in the presence of ADP-Fe(3+), indicating that EDTA may extract iron from ADP-Fe(3+) to catalyze HO. formation by dopa. Thus, EDTA was a good catalyst for HO.-generation, whereas it did not promote the strand breakage of DNA. However, calf thymus DNA base damage, which was detected as 8-HOG formation, was caused by dopa in the presence of EDTA Fe(3+), but not in the presence of ADP-Fe(3+). The 8HOG formation was also inhibited by catalase and HO. scavengers, indicating that HO&z.rad; was involved in the base damage. These results suggest that DNA strand breakage is due to ferryl species rather than HO., and that 8HOG formation is due to HO. rather than ferryl species. PMID- 10862814 TI - Identification of NF-kappaB in the marine fish Stenotomus chrysops and examination of its activation by aryl hydrocarbon receptor agonists. AB - Members of the Rel family of proteins have been identified in Drosophila, an echinoderm, Xenopus, birds and mammals. Dimers of Rel proteins form the transcription factor nuclear factor kappaB (NF-kappaB) that rapidly activates genes encoding cytokines, cell surface receptors, cell adhesion molecules and acute phase proteins. Evidence suggests that xenobiotic compounds also may alter the activation of NF-kappaB. This study had a dual objective of identifying members of the Rel family and examining their activation by xenobiotic compounds in a marine fish model, scup (Stenotomus chrysops). A DNA-protein crosslinking technique demonstrated that liver, kidney and heart each had at least three nuclear proteins that showed specific binding to an NF-kappaB consensus sequence, with molecular weights suggesting that the proteins potentially corresponded to mouse p50, p65 (RelA) and c-rel. In addition, an approximately 35kD NF-kappaB binding protein was evident in liver and kidney. The 50 kD protein was immunoprecipitated by mammalian p50-specific antibodies. The presence of Rel members in fish implied by those results was confirmed by RT-PCR cloning of a Rel homology domain (an apparent c-rel) from scup liver. NF-kappaB activation occurred in vehicle-treated fish, but this appeared to decrease over time. In fish treated with 0.01 or 1 mg 3,3',4,4', 5-pentachlorobiphenyl per kg, NF-kappaB activation in liver did not decrease, and there was a 6-8-fold increase in activation 16-18 days following treatment. Treatment with 10 mg benzo[a]pyrene/kg had no effect on NF-kappaB-DNA binding, either at 3 or 6 days following treatment. The data show that the Rel family of proteins is present in fish, represented at least by a p50/105 homologue, and support a hypothesis that some aryl hydrocarbon receptor agonists can activate NF-kappaB in vivo. PMID- 10862815 TI - Contrasting role of Na(+) ions in modulating Cu(+2) or Cd(+2) induced hepatocyte toxicity. AB - Previously we showed that hepatocyte lysis induced by Cu(+2)/Cd(+2) could be partly attributed to membrane lipid peroxidation induced by Cu(+2) or mitochondrial toxicity induced by Cd(+2) [5]. Changes in Na(+) and Ca(+2) homeostasis induced when Cu(+2) was incubated with hepatocytes markedly differed from that induced by Cd(+2). Na(+) omission from the media or addition of the Na(+)/H(+) exchange inhibitor 5-(N,N-dimethyl)-amiloride markedly increased Cu(+2) cytotoxicity even though Cu(+2) did not increase hepatocyte Na(+) when the media contained Na(+). Intracellular Ca(+2) levels however were markedly increased when the hepatocytes were incubated with Cu(+2) in a Na(+) free media and removing media Ca(+2) with EGTA also prevented Cu(+2) induced hepatocyte cytotoxicity. This suggests that intracellular Ca(+2) accumulation contributes to Cu(+2) induced cytotoxicity and a Na(+)-dependent Ca(+2) transporter is involved in controlling excessive Ca(+2) accumulation caused by Cu(+2). The omission of Cl(-) from the media or addition of glycine, a Cl(-) channel blocker also enhanced Cu induced cytotoxicity. By contrast Cd(+2) induced cytotoxicity was prevented by Na(+) omission from the media or by the addition of 5-(N,N-dimethyl) amiloride. Furthermore the omission of Cl(-) from the media or addition of glycine also prevented Cd(+2) induced hepatocyte toxicity. A hypotonic media also increased Cd(+2) but not Cu(+2) induced hepatocyte cytotoxicity. This suggests that Cd(+2) but not Cu(+2) cytotoxicity could be partly attributed to disruption of cell volume regulation mechanisms. The increased osmotic load caused by the uncontrolled accumulation of intracellular Na(+) in Cd(+2) treated hepatocytes likely resulted from the activation of Na(+)/H(+) exchanger and the Na(+)/HCO(3)( ) cotransporter by the acidosis and ATP depletion caused by mitochondrial toxicity. PMID- 10862816 TI - Quantitation of benzo[a]pyrene-DNA adducts by postlabeling with 14C-acetic anhydride and accelerator mass spectrometry. AB - Quantitation of carcinogen-DNA adducts provides an estimate of the biologically effective dose of a chemical carcinogen reaching the target tissue. In order to improve exposure-assessment and cancer risk estimates, we are developing an ultrasensitive procedure for the detection of carcinogen-DNA adducts. The method is based upon postlabeling of carcinogen-DNA adducts by acetylation with 14C acetic anhydride combined with quantitation of 14C by accelerator mass spectrometry (AMS). For this purpose, adducts of benzo[a]pyrene-r-7,t-8 dihydrodiol-t-9,10-epoxide (BPDE) with DNA and deoxyguanosine (dG) were synthesized. The most promutagenic adduct of BPDE, 7R,8S,9R-trihydroxy-10S-(N(2) deoxyguanosyl)-7,8,9, 10-tetrahydrobenzo[a]pyrene (BPdG), was HPLC purified and structurally characterized. Postlabeling of the BPdG adduct with acetic anhydride yielded a major product with a greater than 60% yield. The postlabeled adduct was identified by liquid chromatography-mass spectrometry as pentakis(acetyl) BPdG (AcBPdG). Postlabeling of the BPdG adduct with 14C-acetic anhydride yielded a major product coeluting with an AcBPdG standard. Quantitation of the 14C postlabeled adduct by AMS promises to allow detection of attomolar amounts of adducts. The method is now being optimized and validated for use in human samples. PMID- 10862817 TI - Effect of substituents on microsomal reduction of benzo(c)fluorene N-oxides. AB - The potential benzo(c)fluorene antineoplastic agent benfluron (B) displays high activity against a broad spectrum of experimental tumours in vitro and in vivo. In order to suppress some of its undesirable properties, its structure has been modified. Benfluron N-oxide (B N-oxide) is one of benfluron derivatives tested. The main metabolic pathway of B N-oxide is its reduction to tertiary amine B. A key role of cytochrome P4502B and P4502E1 in B N-oxide reduction has been proposed in the rat. Surprisingly, B N-oxide is reduced also in the presence of oxygen although all other N-oxides undergo reduction only under anaerobic conditions. With the aim to determine the influence of the N-oxide chemical structure and its redox potential on reductase affinity, activity and oxygen sensitivity five relative benzo(c)fluorene N-oxides were prepared. A correlation between the redox potential measured and the non-enzymatic reduction ability of the substrate was found, but no effect of the redox potential on reductase activity was observed. Microsomal reductases display a high affinity to B N-oxide (apparent K(m) congruent with0. 2 mM). A modification of the side-chain or nitrogen substituents has led to only a little change in apparent K(m) values, but a methoxy group substitution on the benzo(c)fluorene moiety induced a significant K(m) increase (ten-fold). Based on kinetic study results, the scheme of mechanism of cytochrome P450 mediated benzo(c)fluorene N-oxides reduction have been proposed. All benzo(c)fluorene N-oxides under study were able to be reduced in the presence of oxygen. Changes in the B N-oxide structure caused an extent of anaerobic conditions preference. The relationship between the benzo(c)fluorene N oxide structure and the profile of metabolites in microsomal incubation was studied and important differences in the formation of individual N-oxide metabolites were found. PMID- 10862818 TI - Styrene oxide-induced 2'-deoxycytidine adducts: implications for the mutagenicity of styrene oxide. AB - The reaction between 2'-deoxycytidine and styrene 7,8-oxide (SO) resulted in alkylation at the 3-position and at the O(2)-position through the alpha- and beta carbons of the epoxide but at the N(4)-position only through the alpha-carbon. The 3-alkylated adducts were found to deaminate to the corresponding 2' deoxyuridine adducts (37 degrees C, pH 7.4) with half-lives of 6 min and 2.4 h for the alpha- and beta-isomers, respectively. The N(4)-alkylated products were stable at neutral pH. The O(2)-alkylated products were unstable being prone to depyrimidation and to isomerisation between alpha- and beta-isomers. In SO treated double-stranded DNA, enzymatic hydrolysis allowed the identification of the beta3-deoxyuridine and alphaN(4)-deoxycytidine adducts (1.9 and 0.5% of total alkylation, respectively), in addition to the previously identified DNA-adducts. The 3-substituted uracil may have implications for the mutagenicity of SO. PMID- 10862819 TI - Preventive aspirin treatment of streptozotocin induced diabetes: blockage of oxidative status and revertion of heme enzymes inhibition. AB - Some late complications of diabetes are associated with alterations in the structure and function of proteins due to glycation and free radicals generation. Aspirin inhibits protein glycation by acetylation of free amino groups. In the diabetic status, it was demonstrated that several enzymes of heme pathway were diminished. The aim of this work has been to investigate the in vivo effect of short and long term treatment with acetylsalicylic acid in streptozotocin induced diabetic mice. In both treatments, the acetylsalicylic acid prevented delta aminolevulinic dehydratase and porphobilinogen deaminase inactivation in diabetic mice and blocked the accumulation of lipoperoxidative aldehydes. Catalase activity was significantly augmented in diabetic mice and the long term treatment with aspirin partially reverted it. We propose that oxidative stress might play an important role in streptozotocin induced diabetes. Our results suggest that aspirin can prevent some of the late complications of diabetes, lowering glucose concentration and probably inhibiting glycation by acetylation of protein amino groups. PMID- 10862820 TI - 2,3,7,8-Tetrachlorodibenzo-p-dioxin alters melatonin metabolism in fish hepatocytes. AB - Pineal hormone melatonin is an important regulator of endocrine and circadian rhythms in vertebrates. Since liver is assumed to be the major organ in the metabolism of this indole hormone, we investigated the effect of the known Ah receptor agonist, 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) on melatonin metabolism in fish hepatocytes as well as the in vitro effect of melatonin on trout hepatic microsomal cytochrome P4501A (CYP1A) catalyst. Primary cell cultures of rainbow trout hepatocytes were exposed to [3H]melatonin (1 nM to 1 microM) alone and in combination with TCDD (50 pM) at 15 degrees C for 24 or 48 h. Analysis of melatonin and its metabolites in the culture medium and hepatocytes by HPLC revealed that about 96% of the added [3H]melatonin was metabolised after 24 h in both control and TCDD treated cultures. 3H radioactivity was found mainly in the culture medium and less than 5% of the total 3H-radioactivity retained inside hepatocytes. Of the HPLC separated metabolites, one coeluted with 6-hydroxymelatonin and one unknown metabolite eluted after 6-hydroxymelatonin. In addition, two other metabolites were more water-soluble than 6-hydroxymelatonin and were considered to be conjugated products. Treatment of the hepatocytes with TCDD increased the amount of the major oxidated product, 6-hydroxymelatonin, about 2.5-fold after 24 h and 1.2 fold after 48 h exposure, respectively when compared with the control cultures. Whereas the amount of the unknown metabolite eluting after 6-hydroxymelatonin decreased about 1.3-fold after 24 h and 1.2-fold after 48 h exposure, respectively. Melatonin alone did not affect P4501A associated EROD-activity or CYP1AmRNA levels in the primary hepatocyte cultures. TCDD-treatment increased EROD-activity 3 to 5-fold and respective CYP1AmRNA content 6 to 14-fold, when compared with the control or melatonin-treated cultures. Furthermore, melatonin competitively inhibited EROD-activity in liver microsomes with a Ki value of 62.06+/-3.78 microM. The results show that TCDD alters metabolic degradation of melatonin in hepatocytes and suggest that P4501A may be an important P450 isoenzyme involved in oxidative metabolism of melatonin in fish liver. PMID- 10862821 TI - Ethanol alters angiotensin II stimulated mitogen activated protein kinase in hepatocytes: agonist selectivity and ethanol metabolic independence. AB - Angiotensin II activated mitogen-activated protein kinase (MAPK) (p42 and p44) in rat hepatocytes exposed to ethanol and the relevance of ethanol metabolism on this activation was investigated. Hepatocytes, isolated from rat liver, were treated with or without ethanol for 24 h. Angiotensin II, vasopressin, insulin, serum and epinephrine significantly increased hepatocyte MAPK activity. Platelet activating factor (PAF), tumor necrosis factor-alpha (TNF-alpha), and insulin like growth factor-1 (IGF-1) had little effect on MAPK activation. Interestingly, among the above agonists, which activated hepatocyte MAPK, ethanol exposure potentiated only angiotensin II and epinephrine-stimulated MAPK. Thus, potentiation of MAPK by ethanol exhibited agonist selectivity. In contrast to several other cells, there was prevalence of p42 over p44 MAPK band in hepatocytes. Angiotensin II treatment caused a rapid activation (peak 5 min) of MAPK followed by a decrease to basal levels in 30 min. Exposure with 100 mM ethanol potentiated the angiotensin II stimulated MAPK activity. This potentiation was partially blocked by pertussis toxin suggesting it to be a G protein-dependent event. Treatment of the hepatocytes with pyrazole (an inhibitor of ethanol metabolism) or acetaldehyde (an ethanol metabolite) had no effect on potentiation. Thus, ethanol potentiation of hepatocyte MAPK is agonist-selective and independent of ethanol metabolism. PMID- 10862822 TI - Differential effects of acute administration of haloperidol and clozapine on ethanol-induced ascorbic acid release in rat striatum. AB - Antipsychotic drugs were initially considered to act predominantly through their antagonism at dopamine D(2)-like receptors. However, reports have demonstrated that the typical neuroleptic drug haloperidol and the atypical neuroleptic drug clozapine showed differential actions in clinical, behavioral and biochemical studies. Since ascorbic acid has a potential usefulness in psychological therapeutics, the present study investigates the actions of these two drugs on ethanol-induced ascorbic acid release in the striatum in order to help explain the different mechanisms of these drugs. The results showed that clozapine, at the doses of 15 and 30 mg/kg, i.p., had no effect on basal ascorbic acid release. However, a synergistic tendency at a dose of 15 mg/kg and a significant synergism at a dose of 30 mg/kg were observed on ascorbic acid release when clozapine was used with ethanol. In contrast, haloperidol, at the doses of 0.5, 1.0 and 2.0 mg/kg, i.p., administered alone did not affect the basal release of striatal ascorbic acid, and when used together with ethanol had neither a potentiating nor an antagonizing effect on ethanol-induced ascorbic acid release. Chlorpromazine, a nonselective dopamine receptor antagonist, at the dose of 5 mg/kg, i.p., affected neither the basal nor the ethanol-induced ascorbic acid release. Ritanserin, a 5-HT(2) receptor antagonist, at the dose of 1 mg/kg, s.c., significantly antagonized ethanol-induced ascorbic acid release. These results demonstrate that clozapine dose-dependently potentiates the stimulatory effect of ethanol on striatal ascorbic acid release and this effect of clozapine may not be related to its dopamine D(2) receptor antagonism. PMID- 10862823 TI - Brain phospholipase A(2)-arachidonic acid cascade is involved in the activation of central sympatho-adrenomedullary outflow in rats. AB - The present experiments were designed to explore the role of the brain phospholipase A(2)-arachidonic acid cascade in the activation of central sympatho adrenomedullary outflow in rats, using melittin (an activator of phospholipase A(2)) and arachidonic acid. Intracerebro-ventricularly administered melittin (2.5, 10, and 25 microg/animal) or arachidonic acid (75, 150, 300 microg/animal) effectively and dose dependently elevated plasma levels of adrenaline and noradrenaline. The elevation of both catecholamines induced by melittin (10 microg/animal) was abolished by centrally administered mepacrine (an inhibitor of phospholipase A(2)), but not by neomycin (an inhibitor of phospholipase C). However, mepacrine had no effect on the increase induced by arachidonic acid (150 microg/animal). Indomethacin (an inhibitor of cyclooxygenase) abolished all responses induced by melittin and arachidonic acid. Furegrelate (an inhibitor of thromboxane A(2) synthase) abolished the elevation of adrenaline induced by melittin and arachidonic acid, but had no effect on the elevation of noradrenaline induced by these compounds. These results suggest that activation of the brain phospholipase A(2)-arachidonic acid cascade facilitates the central sympatho-adrenomedullary outflow in rats. Brain thromboxane A(2) is involved in the activation of central adrenomedullary outflow and an active metabolite of arachidonic acid other than thromboxane A(2) may be involved in activation of the central sympathetic outflow. PMID- 10862824 TI - Interactions of glutamate receptor agonists with long-term potentiation in the rat hippocampal slice. AB - Previous work has described the apparent desensitisation of neuronal networks in the rat neocortex to amino acid agonists, following prior exposure several minutes earlier. Since long-term potentiation is believed to involve activation of amino acid receptors, we have now sought to determine whether long-term potentiation can modify the sensitivity of neurones to glutamate receptor agonists in rat hippocampal slices. Responses were measured as the change in population spike or postsynaptic potential (e.p.s.p.) size. Two applications of N methyl-D-aspartate (NMDA), quinolinic acid, alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) or kainate, 45 min apart, did not exhibit any apparent desensitisation. However, the induction of long-term potentiation produced a marked loss of sensitivity to quinolinic acid, with smaller effects on NMDA, AMPA and kainate responses. No marked changes were obtained of e.p. s.p. size. In order to localise the cellular sites of these changes, agonists were also applied by microiontophoresis to the cell bodies or dendritic regions of CA1 neurones. Responses to quinolinic acid showed apparent desensitisation at both sites, whereas no decrease was observed in responses to NMDA or AMPA application. The induction of long-term potentiation again produced a decrease in the size of responses to NMDA and AMPA. Inhibition of nitric oxide (NO) synthase prevented the long-term potentiation-induced loss of responsiveness to NMDA, but not AMPA, implying a role for NO in the loss of NMDA sensitivity. Recordings of single cell activity during the iontophoretic application of agonists and induction of long term potentiation showed that responses to NMDA were often suppressed to a greater extent than to quinolinic acid. The results indicate that long-term potentiation can modify the sensitivity of hippocampal neurones to glutamate receptor agonists, and that differences exist in the pharmacology of NMDA and quinolinic acid. PMID- 10862825 TI - Central administration of alpha-melanocyte stimulating hormone inhibits fasting- and neuropeptide Y-induced feeding in neonatal chicks. AB - In the present study, the effect of intracerebroventricular (i.c.v.) administration of alpha-melanocyte stimulating hormone (alpha-MSH) on food intake of neonatal chicks was examined. In experiment 1, i.c. v. injection of alpha-MSH (0.04, 0.2 and 1 microg) significantly inhibited food intake of 3-h fasted chicks in a dose-dependent manner. In experiment 2, alpha-MSH strongly inhibited neuropeptide Y-induced feeding when neuropeptide Y (2.5 microg) and several doses of alpha-MSH were given simultaneously i.c.v. These results suggest that alpha MSH plays an important role in the regulation of food intake of neonatal chicks. PMID- 10862826 TI - Ischaemia selectivity confers efficacy for suppression of ischaemia-induced arrhythmias in rats. AB - Eight novel and three reference antiarrhythmics were investigated in anaesthetised rats for antiarrhythmic actions, as well as for effects on the electrocardiogram (ECG) under normal and "simulated ischaemic" conditions. In rats subjected to coronary artery occlusion lidocaine, (+/-)-trans-[2-(4 morpholinyl)-cyclohexyl]naphthyl-1-acetate, RSD1000 and (+/-)-trans-[2-(4 morpholinyl)-cyclohexyl]-2-(1-naphthyl)propionate, RSD1030, (Group A) produced dose-related and complete antiarrhythmic protection. Group B compounds, such as (+/-)-trans-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]-3, 4-dichlorocinnamamide, RSD995, produced complete antiarrhythmic protection but had aberrant dose response curves. Group C compounds, such as quinidine and flecainide, failed to give full antiarrhythmic protection and had shallow dose-response curves. The potency of Group A compounds, but not Group B or C compounds, for ECG actions indicative of Na(+) channel blockade (prolongation of PR and QRS intervals) were significantly increased under "simulated ischaemic" conditions ([K(+)] 10 mM and pH 6.4) in isolated rat hearts. Thus, compounds with ischaemia-selective actions provided superior protection against ischaemia-induced arrhythmias in rats. PMID- 10862827 TI - Changes in local cerebral blood flow in photochemically induced thrombotic occlusion model in rats. AB - We demonstrated earlier that in a photochemically induced thrombotic occlusion model, a reperfusion-like phenomenon may be involved in the progress of brain damage. Therefore, we now investigated changes in local cerebral blood flow in a photochemical model compared with changes in a thermocoagulated occlusion model, using autoradiography. At 5 min, and 3, 6 and 24 h after middle cerebral artery occlusion, local cerebral blood flow was measured by intravenous injection of 4 iodo[N-methyl-14C]antipyrine (20 mu Ci). In the ischemic core zone, the reduction in blood flow was similar in the two models. However, blood flow in the ischemic border zone in the photochemical model decreased transiently in the third hour after ischemia and then increased again, while the blood flow in a thermocoagulated model continued to decrease. Time courses of brain damage formation in both models were no different up to 24 h. These findings suggest that the transient reduction in cerebral blood flow in the third hour following ischemia may contribute to a reperfusion-like phenomenon in a photochemical model. PMID- 10862828 TI - Effects of imidapril and captopril on streptozotocin-induced diabetic nephropathy in mice. AB - We investigated whether the prevention of the development of diabetic nephropathy by angiotensin-converting enzyme inhibitors is associated with decreases in renal angiotensin-converting enzyme activity and/or blood pressure in diabetic mice. C57Bl/6 mice were injected with streptozotocin (200 mg/kg, i.v.) and randomized to receive either imidapril (1 and 5 mg/kg) or captopril (10 and 50 mg/kg) or vehicle by gavage for 28 days. Each assay was performed on 8-10 mice from each treatment. At 28 days after the start of drug treatment, imidapril and captopril significantly reduced blood pressure of the diabetic mice, and this effect of captopril was stronger than that of imidapril. On the other hand, inhibition of renal angiotensin-converting enzyme activity by imidapril was stronger than that by captopril. Imidapril and captopril dose-dependently inhibited urinary albumin excretion to similar extents, but they failed to inhibit the renal hypertrophy and elevation of creatinine clearance. Total renal angiotensin-converting enzyme activity was significantly reduced in diabetic mice, but immunohistochemical localization of angiotensin-converting enzyme was intensive in the vasculature and glomeruli of the diabetic kidney. In conclusion, both effects on blood pressure and angiotensin-converting enzyme activity may be involved in the prevention of development of diabetic nephropathy by imidapril and captopril in streptozotocin-induced diabetic mice. The data suggest that the degrees of contribution of their effects on blood pressure and renal angiotensin-converting enzyme activity to the inhibition of urinary albumin excretion may be different between the two angiotensin-converting enzyme inhibitors. PMID- 10862829 TI - Effect of nepadutant at tachykinin NK(2) receptors in human intestine and urinary bladder. AB - We have characterized the action of the tachykinin NK(2) receptor antagonist nepadutant (c?[(beta-D-GlcNAc)Asn-Asp-Trp-Phe-Dpr-Leu]c(2 beta-5 beta)?) in the human isolated ileum, colon and urinary bladder. Nepadutant (30-1000 nM) competitively antagonized neurokinin A- or [beta Ala(8)]neurokinin A-(4-10) induced contractions in all tissues, with pK(B)=8.3 (ileum and colon) and pK(B)=8.5 (bladder). In contrast, the nonpeptide tachykinin NK(2) receptor antagonist SR 48968 (or (S)-N-methyl-N [4-acetylamino-4-phenylpiperidino)-2-(3, 4 dichlorophenyl) butyl] benzamide) (30-1000 nM) produced insurmountable antagonism in all preparations. The tachykinin NK(2) receptor blockade produced by nepadutant in the colon was fully reversed by washout, whereas that produced by SR 48968 was not. Nepadutant (1 microM) greatly reduced (by 70-80%) the nonadrenergic noncholinergic (NANC) contractile off-response evoked by electrical field stimulation in the human ileum, and almost abolished it in the presence of the tachykinin NK(1) receptor antagonist GR 82334 (or: [[(S,S) Pro-Leu (spiro gamma-lactam)](9,10),Trp(11)]Physalaemin (1-11)) (1 microM). The present results show that nepadutant is a potent, competitive and reversible antagonist at human tachykinin NK(2) receptors and provide further evidence that tachykinins act as excitatory NANC neurotransmitters in the human small intestine. PMID- 10862830 TI - In vitro anti-inflammatory effects of cynaropicrin, a sesquiterpene lactone, from Saussurea lappa. AB - We investigated in vitro anti-inflammatory effects of cynaropicrin, a sesquiterpene lactone from Saussurea lappa, on tumor necrosis factor (TNF)-alpha and nitric oxide (NO) release, and lymphocyte proliferation. Cynaropicrin strongly inhibited TNF-alpha release from lipopolysaccharide-stimulated murine macrophage, RAW264.7 cells, and differentiated human macrophage, U937 cells, proved to produce notable amount of TNF-alpha. It also potently attenuated the accumulation of NO released from lipopolysaccharide- and interferon-gamma stimulated RAW264.7 cells in a dose-dependent manner. In addition, the immunosuppressive effects of the compound on lymphocyte proliferation in response to mitogenic stimuli were examined. Cynaropicrin also dose-dependently suppressed the proliferation of lymphocytes from splenocytes and interleukin-2-sensitive cytotoxic T lymphocyte, CTLL-2 cells, stimulated by lipopolysaccharide, concanavalin A, phytohemagglutinin and interleukin-2. However, treatment with sulphydryl compound, L-cysteine, abrogated all these inhibitory effects. These results suggest that cynaropicrin may participate in the inflammatory response by inhibiting the production of inflammatory mediators and the proliferation of lymphocytes and its inhibitory effect is mediated through conjugation with sulphydryl groups of target protein(s). PMID- 10862831 TI - Oral administration of branched chain amino acids improves virus-induced glucose intolerance in mice. AB - We investigated the therapeutic effect of branched chain amino acids (BCAA) on mice with glucose intolerance induced by encephalomyocarditis virus (EMCV). Male DBA/2 mice were divided into three groups: treated with BCAA, (such as valine, leucine, and isoleucine), untreated, and control. BCAA-treated and -untreated groups were inoculated intraperitoneally with the NDK25 variant of EMCV at 200 plaque-forming units per mouse. The BCAA-treated group was administered orally 0.9 g/kg/day of each BCAA from the day after viral inoculation. The control group neither received virus inoculation nor was treated with BCAA. One week after inoculation, oral glucose tolerance tests (OGTT) were performed. After the glucose loading at 1.5 g/kg of body weight, blood glucose levels in the untreated group were 92.0+/-10.0 mg/dl at baseline, 224.6+/-10.9 mg/dl at 30 min, and 169.4+/-21.4 mg/dl at 60 min, which were significantly (P<0.05) higher than those in the control group (62. 7+/-3.6 mg/dl, 167.2+/-16.4, and 83.8+/-6.0 mg/dl, respectively). Blood glucose levels in the BCAA-treated group were 54.5+/-3.7 mg/dl at baseline, 145.2+/-8.7 mg/dl at 30 min, and 128.7+/-18.3 mg/dl at 60 min after the glucose loading, which were not significantly higher than those in the control group. Immunoreactive insulin levels at 30 min after the glucose loading were lower in the untreated group than in the control group at 1 week after virus inoculation. Histological investigations showed that the grade of insulitis in the pancreas of mice of the BCAA-treated group was lower than that of the mice of the untreated group. These results suggest that oral administration of BCAA is able to improve glucose intolerance induced by EMCV. PMID- 10862832 TI - Emergency contraception: advance provision in a young, high-risk clinic population. AB - OBJECTIVE: To assess whether advance provision of emergency contraception increases its use and whether it has secondary effects on regular contraceptive use. METHODS: We conducted a controlled trial of female clients, aged 16-24 years, who attended a publicly funded family planning clinic. Women were systematically assigned to receive an advance provision of emergency contraception and education (treatment) or education only (control). Among 263 participants enrolled (133 treatment, 130 control), follow-up was completed in 213 (111 treatment, 102 control). The main outcome measures were emergency contraception knowledge and use, frequency of unprotected sex, and pattern of contraceptive use in the past 4 months. RESULTS: Participants were aware of emergency contraception at follow-up, but the treatment group was three times as likely to use it (P =.006). Although the treatment group did not report higher frequencies of unprotected sex than the control group, women in the treatment group (28%) were more likely than those in the control group (17%) to report using less effective contraception at follow-up compared with enrollment (P =.05). The proportion of women in both groups who reported consistent pill use increased from enrollment to follow-up (34% versus 45%); however, the control group (58%) was more likely than the treatment group (32%) to report consistent pill use at follow-up (P =.03). CONCLUSION: Use of emergency contraception was increased by providing it in advance, but not by education alone. Changes to less effective contraceptive methods and patterns of pill use were potentially negative effects that need to be explored in relation to observed benefits. PMID- 10862833 TI - Gonadotropin and body mass index: high FSH levels in lean, normally cycling women. AB - OBJECTIVE: To characterize the relationships between body mass index (BMI) and LH or FSH levels over the cycle in normally cycling women. METHODS: We compared baseline characteristics, cycle characteristics, follicle sizes, and daily hormone levels among women with low (n = 22), normal (n = 63), or high (n = 22) BMIs over 326 cycles. RESULTS: There were no significant differences in age or other lifestyle characteristics between groups. High BMI was significantly associated with younger age at menarche and less sleeping time. No differences were observed between high- and low-BMI groups in cycle length or diameter of the dominant follicle. Luteinizing hormone levels were significantly higher only in the beginning of the cycle in women with low BMIs than in those with high BMIs. Follicle-stimulating hormone levels were also significantly higher but were high during all three phases of the cycle (early follicular, periovulatory, and luteal phases). Mean levels were approximately 1. 9, 1.8, and 1.2 times higher, respectively, in the low-BMI group than the high-BMI group. CONCLUSION: Luteinizing hormone levels and BMI were inversely associated in normally cycling women during the early follicular phase. Follicule-stimulating hormone levels and BMI were inversely associated during the whole cycle, independent of age. PMID- 10862834 TI - Effect of non-weight-bearing body fat on bone mineral density before and after menopause. AB - OBJECTIVE: To investigate the difference in the effect of non-weight-bearing body fat mass on bone mineral density between premenopausal and postmenopausal women. METHODS: We studied 252 regularly menstruating premenopausal women and 213 postmenopausal women with right side dominance. Age, years since menopause (in postmenopausal women), height, weight, and body mass index were recorded. Bone mineral density of non-weight-bearing sites (ie, arms), weight-bearing sites (ie, lumbar spine including L2-4 and legs), and body fat mass were measured by whole body scanning with dual-energy x-ray absorptiometry. Body fat mass was also measured by dual energy x-ray absorptiometry. RESULTS: Body fat mass did not differ between groups. In postmenopausal women, body fat mass correlated positively with bone mineral density of the left leg (r =. 41, P <.001), right leg (r =.36, P <.001), left arm (r =.31, P <. 001), and lumbar spine (r =.27, P <.001). The correlation between body fat mass and bone mineral density of the left arm remained significant after adjusting for age, years since menopause, and height. In premenopausal women, body fat mass correlated positively with bone mineral density of left leg (r =.37, P <.001) and right leg (r = 0.31, P <.001), but correlated weakly with bilateral arms (r < or =.19) and lumbar spine bone mineral density (r = 0.13, P <.05). CONCLUSION: The effect of non-weight-bearing body fat on bone mineral density was greater in postmenopausal than premenopausal women. PMID- 10862835 TI - Effect of patient position on clinical evaluation of pelvic organ prolapse. AB - OBJECTIVE: To compare the severity of pelvic organ prolapse between examinations performed in dorsal lithotomy position and examinations performed upright in a birthing chair using the Pelvic Organ Prolapse Quantification System (POPQ). METHODS: One hundred eighty-nine consecutive women were evaluated between April 1997 and September 1998. All women were examined in the dorsal lithotomy position and in a birthing chair at a 45 degrees angle. Degree of pelvic organ prolapse was assessed using the POPQ. RESULTS: When examined upright, 133 patients (70%) had the same stage of prolapse, whereas 49 (26%) had a higher stage and seven (4%) had a lower stage. Of patients who were stage 0 or I when examined in lithotomy position, 23 (36%) were stage II or greater when examined upright. Similarly, of patients who were stage II in lithotomy, 17 (23%) were stage III or higher when examined upright. There was a statistically significant increase in the degree of prolapse at all the POPQ measurements (P <.05 for each point), except for measurement of total vaginal length. Forty-eight percent of patients had at least one measurement increase by 2 cm or more when examined upright. Logistic regression identified no patient characteristics that were independently associated with a significant increase in stage or POPQ values with change in examination position. CONCLUSION: The degree of pelvic organ prolapse assessed with the patient in the lithotomy position correlates well with assessment performed upright; however, overall there is a higher degree of prolapse with upright examination. PMID- 10862836 TI - Bacteriology and treatment of malodorous lower reproductive tract in gynecologic cancer patients. AB - OBJECTIVE: To determine the bacteriology of lower genital tract cancers to direct potential treatment modalities and to determine the impact of treatment on quality of life. METHODS: Gram stain, saline preparations, tumor pH determinations, and anaerobic and aerobic tumor cultures were obtained from 13 consecutive patients with malodorous gynecologic cancers and 13 patients (controls) with nonmalodorous tumors. All patients with odor were treated with topical metronidazole for 7 days. Odor assessment questionnaires were administered daily in the treatment group. Quality-of-life evaluation was assessed using the Functional Assessment of Cancer Therapy questionnaire before and after treatment. RESULTS: Cancer of the cervix (n = 21) was the most common primary site and accounted for 81% (95% confidence interval 61%, 93%) of malodorous gynecologic cancers. Eight of 13 (62%) patients with malodorous tumors had bacterial vaginosis compared with four of 13 (31%) of those without odor (P =.11). Aerobic and anaerobic bacteria were isolated with equal frequency from malodorous gynecologic cancers. Results of odor assessment questionnaires showed a graded improvement with topical antibiotic therapy (P <.001). The Functional Assessment of Cancer Therapy questionnaire indicated improved quality of life after therapy (P =.02). CONCLUSION: Most patients with odor had bacterial vaginosis and had an improvement in odor with topical metronidazole. Therefore, this treatment might be useful for patients with malodorous pelvic tumors. PMID- 10862837 TI - BRCA1 germline mutations in women with uterine serous papillary carcinoma. AB - OBJECTIVE: To determine the possible effects and incidence of BRCA1 and BRCA2 germline mutations in uterine serous papillary carcinoma. METHODS: We screened DNA from 12 women with uterine serous papillary carcinoma for BRCA1 and BRCA2 germline mutations common in the Jewish population (BRCA1-185delAG and 5382insC, BRCA2-6174delT). In women with germline mutations, tumor DNA was screened for loss of heterozygosity at the appropriate loci. RESULTS: Nine women were of Jewish Ashkenazi origin and three were non-Ashkenazi. Two of nine Ashkenazi women were carriers of germline mutations: one 185delAG mutation and one 5382insC mutation. Five women had histories of breast carcinoma before diagnosis of uterine serous papillary carcinoma. Family histories of seven women had at least one first-degree relative with malignant disease. Of those, four had at least one first-degree relative with breast, ovarian, or colon carcinoma. Both carriers had strong family histories of breast-ovarian carcinoma. Loss of heterozygosity analysis found loss of the wild-type BRCA1 allele in the primary uterine tumors. CONCLUSION: BRCA1 germline mutations were observed in two of nine of the women in this series. The loss of heterozygosity in the tumor tissue of the carriers, coupled with the high frequency of family and patient histories of breast or ovarian malignancies, suggest that uterine serous papillary carcinoma might be a manifestation of familial breast-ovarian cancer. PMID- 10862838 TI - Anthropometric estimation of maternal body composition in late gestation. AB - OBJECTIVE: To construct a model to estimate maternal body composition in late gestation using anthropometric measurements. METHODS: Twenty healthy pregnant women at 30 weeks' gestation had estimates of body composition using hydrodensitometry, with corrections for residual lung volume, and total body water using H(2)(18)O (development group). Total body water was estimated from (18)O abundances measured by gas-isotope-ratio mass spectrometry. Maternal age, height, weight, and seven skinfold sites were correlated with fat mass using stepwise regression analysis. The anthropometric model to estimate fat mass was then tested prospectively in a second group of 20 subjects and correlated with underwater weighing and total body water measurements (validation group). Statistical analysis used chi(2), paired t and Wilcoxon sign-rank tests. RESULTS: There were no statistically significant differences in maternal demographics between groups. The fat mass of development group subjects using underwater weighing and total body water was 22.7 +/- 7.6 kg. Using the development group, a model was derived that explained 91% of the variance in fat mass by underwater weighing and total body water using maternal weight and triceps, subscapular, and suprailiac skinfolds (r(2) = 0.91, P <.001). When tested prospectively in the validation group, the correlation remained statistically significant (r(2) = 0.89, P <.001). There was no statistically significant (P =.88) difference between the anthropometric estimates of fat mass and underwater weighing and total body water measurements (95% confidence interval -2.476, 2.748 kg of fat mass). CONCLUSION: This anthropometric model can be used to predict maternal fat mass in late gestation. PMID- 10862839 TI - Riboflavin deficiency and preeclampsia. AB - OBJECTIVE: To examine in a prospective study riboflavin deficiency as a predisposing factor for preeclampsia in a high-risk collective of pregnant women in Zimbabwe. METHODS: At an antenatal clinic in Bulawayo, Zimbabwe, 154 women at increased risk for preeclampsia were observed prospectively until delivery. Riboflavin status was determined using the erythrocyte glutathione reductase activation coefficient test on the day of antenatal booking. Riboflavin deficiency was expressed by erythrocyte glutathione reductase activation coefficient of 1.4 or greater. RESULTS: Riboflavin deficiency was frequently found among the study population (33.8%). Incidence rose toward the end of pregnancy (27.3% at 29-36 weeks' gestation compared with 53.3% at over 36 weeks). In the riboflavin-deficient group, mothers were more likely to develop preeclampsia (28.8%) than in the riboflavin-adequate group (7.8%; P <.001, odds ratio [OR] 4.7, 95% confidence interval [CI] 1.8-12.2). The calculated concentrations of intracellular free flavin adenine dinucleotide were significantly lower in patients who developed preeclampsia than in normal pregnancies (P <.05). CONCLUSION: Riboflavin deficiency should be considered a possible risk factor for preeclampsia. Insufficient concentrations of the riboflavin-derived cofactors flavin adenine dinucleotide and flavin adenine mononucleotide could contribute to the established pathophysiologic changes including mitochondrial dysfunction, enhanced oxidative stress, and disturbances in nitric oxide release. PMID- 10862840 TI - Severe preeclampsia and high frequency of genetic thrombophilic mutations. AB - OBJECTIVE: To determine whether severe preeclampsia is associated with genetic thrombophilic mutations or other types of thrombophilia. METHODS: A case-control study compared 63 consecutive women with severe preeclampsia evaluated at our institution between November 1997 and April 1999 with 126 control women matched for age and ethnicity. All of these women were tested several months after delivery for mutations of factor V Leiden, methylenetetrahydrofolate reductase, and prothrombin gene; for deficiencies of protein C, protein S, and antithrombin III; and for the presence of anticardiolipin antibodies. RESULTS: Thirty-five study women (56%) had a thrombophilic mutation compared with 24 control women (19%), P <.001. Seven other study women (11%) had other thrombophilias, compared with one control woman (0.8%), P <.01. Within the study group, women with thrombophilia delivered at an earlier gestational age, and their neonates' birth weights were lower compared with those of women without thrombophilia. CONCLUSION: Because thrombophilia was found in 67% of women with severe preeclampsia, we suggest that women who have severe preeclampsia should be tested for thrombophilia. PMID- 10862841 TI - Polymorphism in the glutathione S-transferase P1 gene and risk for preeclampsia. AB - OBJECTIVE: To determine whether genetic variability in biotransformation enzymes contributes to individual differences in susceptibility to preeclampsia or the syndrome of hemolysis, elevated liver enzymes, and low platelets (HELLP). METHODS: Polymorphisms in the genes of glutathione S-transferases and cytochrome P-450 1A1 were assessed by polymerase chain reaction in 170 nonpregnant women with a history of preeclampsia, 90 of whom had HELLP syndrome, and 109 healthy control women with an uncomplicated obstetric history. chi(2) analysis was used for statistical evaluation of differences in polymorphism rates. RESULTS: A higher frequency of the glutathione S-transferase P1b-1b genotype was observed in preeclamptic women than in controls (14% in preeclampsia and 5% in controls; odds ratio 3.4, 95% confidence interval 1.2, 10. 6, P =.02). Genetic polymorphisms in other glutathione S-transferases and cytochrome P-450 1A1 genes occurred equally frequently in cases and controls. In women with a history of preeclampsia, there were no differences in the occurrence of the genetic polymorphisms investigated in women who either did or did not develop the HELLP syndrome. CONCLUSION: Women with the glutathione S-transferase P1b-1b genotype, which could result in lower glutathione S-transferase detoxification capacity, might have higher susceptibility to preeclampsia. However, polymorphisms in glutathione S transferase genes do not seem to be a risk factor for development of the HELLP syndrome. PMID- 10862842 TI - Lipoprotein particles in preeclampsia: susceptibility to oxidative modification. AB - OBJECTIVE: To investigate the susceptibility to oxidation of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) in women with preeclampsia. METHODS: Plasma levels of total cholesterol, total triglyceride, and concentrations of cholesterol, triglyceride, and protein in LDL and HDL were measured in 12 preeclamptic women and 12 normal pregnant women. Oxidation of LDL or HDL was assessed by incubation with copper ions and evaluated by monitoring the kinetics of conjugated diene formation. RESULTS: The plasma levels of total triglyceride and concentration of LDL protein were significantly higher in preeclamptic women than in normals. Levels of HDL lipid did not differ significantly. Analysis of kinetics of conjugated diene production showed a significantly shorter lag time for LDL (83.1 +/- 5.5 minutes versus 67.4 +/- 10.2 minutes, P <.001) and HDL (76.9 +/- 7.3 minutes versus 59.5 +/- 9.2 minutes, P <.001) and a significantly higher oxidation rate for LDL (3.6 +/- 0.4 nmol/minutes/mg LDL versus 4.4 +/- 1.0 nmol/minutes/mg LDL, P <.05) in preeclamptic women. CONCLUSION: Low-density lipoprotein and HDL particles were more susceptible to oxidative modification, and plasma concentration of LDL particles, but not of HDL particles, was increased in preeclampsia. PMID- 10862843 TI - Expanded-spectrum antibiotics with preterm premature rupture of membranes. AB - OBJECTIVE: To compare maternal infection rates, neonatal sepsis rates, and bacterial resistance patterns in cases of neonatal sepsis for three antibiotic protocols for women with preterm premature rupture of membranes (PROM). METHODS: From January 1, 1988 to February 28, 1998, women with preterm PROM not requiring immediate delivery were treated according to one of three antibiotic protocols. During three distinct periods, patients received no antibiotics, intravenous ampicillin for 48 hours followed by oral amoxicillin, or intravenous ticarcillin clavulanic acid for 48 hours followed by oral amoxicillin-clavulanic acid. Rates of chorioamnionitis, endometritis, and neonatal sepsis were compared, as were antimicrobial resistance patterns. Statistical analysis was done using chi(2) analysis, Fisher exact test, and the log-likelihood ratio test. The Bonferroni correction was used for multiple comparisons. RESULTS: During the three periods, preterm PROM was diagnosed in 1695 women. The incidence of endometritis was lower during the third (5.3%) compared with the first (15.1%, P <.001) and second (11.6%, P <.001) protocols. Chorioamnionitis rates were 13.6%, 12.7%, and 15.6% (P =.34) for the first, second, and third periods, respectively, and neonatal sepsis rates were 2.2%, 0.6%, and 1.1% (P =.08), respectively. Neonatal sepsis with gram-negative (P =.02) and ampicillin-resistant (P =.04) organisms was more likely when mothers received antepartum ampicillin or ticarcillin-clavulanic acid. CONCLUSION: Antibiotic therapy for patients with preterm PROM was associated with a decrease in the rate of endometritis and a trend toward less neonatal sepsis but an increase in the proportion of gram-negative and ampicillin resistant organisms causing neonatal sepsis. PMID- 10862844 TI - Serum soluble Fas levels in ovarian cancer. AB - OBJECTIVE: To determine the value of serum soluble Fas levels as a prognostic marker for survival of women with ovarian cancer and as a discriminator between benign and malignant adnexal masses. METHODS: Serum soluble Fas levels were measured with an enzyme-linked immunosorbent assay in 52 women with ovarian cancer, 30 women with benign ovarian cysts, and 35 healthy women. RESULTS: Median serum soluble Fas levels in women with ovarian cancer, women with benign ovarian cysts, and healthy women were 3.7 (range 1.6-14.5), 2.3 (range 1.3-4.1), and 1.5 ng/mL (range 0.1-5.6), respectively (P <. 001). A univariate logistic regression model showed a significant influence of serum soluble Fas and CA 125 levels on the odds of presenting with ovarian cancer versus benign cysts (P <.001 and P =. 001, respectively). In a multivariable logistic regression model for soluble Fas and CA 125, both markers showed a statistically significant influence on the odds of presenting with ovarian cancer versus benign cysts (P =.01 and P =.01, respectively). Increased pretreatment serum soluble Fas levels were associated with shortened disease-free and overall survival (P =.002 and P =.001, respectively). A multivariable Cox regression model identified serum soluble Fas levels as a significant prognostic factor for disease-free and overall survival, independent of tumor stage (P =. 04 and P =.03, respectively). CONCLUSION: Soluble Fas levels might be useful as a discriminator between benign ovarian cysts and ovarian cancer, adding to the information obtained with the use of the established tumor marker CA 125. Pretreatment serum soluble Fas levels also might be an independent prognostic factor for disease-free and overall survival. PMID- 10862846 TI - Prospective evaluation of logistic regression models for the diagnosis of ovarian cancer. AB - OBJECTIVE: To test the accuracy of three logistic regression models in diagnosing malignancy in women with adnexal masses. METHODS: This was a prospective collaborative study. Women were recruited from three hospitals and all assessments were performed at the Gynaecology Ultrasound Unit, King's College Hospital. One hundred women with known adnexal masses were examined preoperatively. The demographic, biochemical, and sonographic data recorded for each patient included age, menopausal status, CA 125 levels, ultrasound morphology, and Doppler blood flow analysis. The diagnosis of malignancy was made for each woman using three logistic regression models previously described by Alcazar et al, Tailor et al, and Timmerman et al. Variables used in these models were then combined to form a new model. The results were compared with the final histopathologic diagnosis. RESULTS: Sixty-seven women had benign tumors and 33 had ovarian cancer. Women with malignant tumors were older than those with benign masses. There were significant differences in CA 125 levels, presence of papillary proliferations, and ascites between the two groups. The sensitivities and specificities achieved respectively by the models were as follows: 45% and 93% with Tailor et al's model, 9% and 99% with Alcazar et al's model, and 73% and 91% with Timmerman et al's model. There was no significant improvement over the performance of Timmerman et al's model and the new combined model. CONCLUSION: All models performed less well than originally reported. Combining the models did not lead to a significant improvement in performance. Larger sample sizes that incorporate all types of ovarian tumors are necessary to design more accurate diagnostic models. PMID- 10862845 TI - Reported ovarian cancer screening among a population-based sample in Washington state. AB - OBJECTIVE: To assess the prevalence of reported ovarian cancer screening among a population-based sample of women from Washington state and identify factors that influence the decision to be screened. METHODS: A population-based sample of 6749 women aged 54-84 years, living in 40 predominately rural communities in Washington state, was surveyed about their utilization of ultrasonography and CA 125 for ovarian cancer screening. We also assessed relevant demographic, family history, psychosocial, and health behavior variables. RESULTS: After exclusions, data from 4938 respondents were available. Two percent (n = 96) reported having been screened. Multiple logistic regression identified ovarian cancer worry, contact with an obstetrician-gynecologist, and family history of ovarian cancer as independently associated with screening. Based on self-reported family histories, 27 women had pedigrees consistent with high risk of ovarian cancer, but none of those women reported having been screened. CONCLUSION: Ovarian cancer screening is rare. Women at high risk of it might not be getting recommended screening. PMID- 10862847 TI - The obstetrician-gynecologist's role in vaccine-preventable diseases and immunization. AB - OBJECTIVE: To assess by survey the immunization role currently played obstetrician-gynecologists in the state of Michigan. METHODS: Masked questionnaires requesting demographic, knowledge-based, practice, and attitudinal data were sent to 850 ACOG-registered fellows. RESULTS: Three hundred sixty-five physicians responded, 313 of whom were in active practice. Most were male (70%) and graduated from medical school between 1970 and 1989 (68%). The majority provided both obstetric and gynecologic services. The minority (47%) specifically identified themselves as primary care providers. Only 15% of respondents considered screening for vaccine-preventable diseases to be outside the realm of routine obstetric-gynecologic care. In practice, however, 19% did not screen their obstetric patients for any vaccine-preventable diseases, and only 10% assessed their patients for all nine vaccine-preventable diseases listed in the questionnaire. In gynecologic patients, almost 40% of physicians did not assess for any vaccine-preventable disease. A wide range in knowledge level was identified concerning vaccine-preventable diseases, immunization recommendations, and vaccine safety. CONCLUSION: These data show a discrepancy between perceived responsibilities and actual practice patterns of obstetrician-gynecologists regarding vaccine-preventable diseases and the immunization of women. Limitations in current knowledge and practical concerns specific to vaccine administration contribute to this disparity. PMID- 10862848 TI - Dietary caffeine intake and the risk for detrusor instability: a case-control study. AB - OBJECTIVE: To determine whether there is an association in women between caffeine intake and risk for detrusor instability. METHODS: Women were included if they had symptoms of urinary incontinence, completed a 48-hour voiding diary detailing fluid and caffeine intake, and had undergone standardized multichannel urodynamics. The study group had 131 women with detrusor instability on provocative cystometry and maximum urethral closure pressure greater than 20 cm of water. The control group had 128 women without detrusor instability on provocative cystometry and maximum urethral closure pressure greater than 20 cm of water. For statistical comparison, women were divided into the following three groups on the basis of caffeine intake: minimal (< 100 mg/day), moderate (100-400 mg/day), and high (> 400 mg/day). RESULTS: The mean caffeine intake of women with detrusor instability (484 +/- 123 mg/day) was significantly higher than that of controls (194 +/- 84 mg/day, P =.002). On univariate analysis, significant risk factors for detrusor instability were age, smoking status, and caffeine intake. On multivariate analysis, the statistically significant association between high caffeine intake and detrusor instability persisted after controlling for age and smoking (odds ratio [OR] 2.4, 95% confidence interval [CI] 1.1, 6.5, P =.018). When women with moderate caffeine intake were compared with those with minimal caffeine intake, the risk for detrusor instability was lower and did not reach significant levels (OR 1.5, 95% CI 0.1, 7.2, P =.093). CONCLUSION: An association between high caffeine intake and detrusor instability was seen in this population. Larger studies are required to determine whether the association is causal. PMID- 10862849 TI - Comparison of endocervical curettage and endocervical brushing. AB - OBJECTIVE: To compare endocervical brushing with endocervical curettage with respect to diagnostic yield by histology and patient discomfort. METHODS: Nonpregnant women referred for colposcopy because of abnormal Papanicolaou test results were randomized to endocervical sampling with either a metal curette (endocervical curettage [ECC]) or an endocervical brush. Extensive endocervical canal brushing was performed. All samples were submitted for histologic study. Results were evaluated against the histologic findings in electroconization specimens in a masked fashion. Pain scores were recorded using Melzack's Present Pain Intensity Scale. RESULTS: During the study period, 315 patients were randomized to the techniques: 157 to ECC and 158 to endocervical brushing. Of the 315 patients, 147 also underwent electroconization. Overall false-positive rates were 28.6% for endocervical brushing and 30.8% for ECC. False positives were due to contamination of the endocervical sample by lesional epithelium near the external os. The proportion of scanty specimens obtained by endocervical brushing (7. 6%) was higher than that obtained by ECC (2.5%) (P =.041). One sample obtained by brushing was insufficient for diagnosis; none obtained by ECC were insufficient. There were no statistically significant differences in the median pain scores between the two groups. CONCLUSION: The techniques were similar in terms of diagnostic yield and patient discomfort. Endocervical brushing had lower false-positive rates than those reported in the literature for cytologic analysis. Although ECC remains the method of choice for evaluation of the endocervical canal, brushing is an acceptable alternative. PMID- 10862850 TI - Economic burden of hospitalizations for preterm labor in the United States. AB - OBJECTIVE: To determine patient factors associated with hospital care costs for preterm labor and to develop a clinically applicable cost model for evaluating economic consequences of interventions to reduce preterm-labor hospitalizations. METHODS: Maryland state hospital discharge data from 1993-1996 were used to identify hospitalizations for preterm labor without delivery and preterm labor with early delivery. Median regression was used to determine the association between patient factors and hospital care costs in Maryland and to develop a model to estimate hospital care costs nationally. National estimates of hospitalizations for preterm labor were from the 1994 National Hospital Discharge Survey. RESULTS: During the 4-year study period, there were 25,104 hospitalizations for preterm labor, undelivered, and preterm labor with early delivery in Maryland. Maternal comorbidity, antenatal procedures, types of insurance, and lengths of stay associated significantly with hospital costs for preterm labor. National costs for preterm labor, undelivered, were more than $360 million. Incremental costs for preterm labor with early delivery, compared with term delivery, ranged from $21 million to $191 million. Total expenditures for preterm-labor hospitalization for the United States were estimated in excess of $820 million. CONCLUSION: Hospitalizations for preterm labor comprise a substantial portion of maternal cost of perinatal care in the United States. Maternal comorbidity and procedures account for major differences in costs per admission. Strategies to reduce hospitalizations for preterm labor should focus on economic and clinical outcomes in evaluating their overall values. PMID- 10862851 TI - Amniotic fluid indices of fetal pulmonary maturity with preterm premature rupture of membranes. AB - OBJECTIVE: To evaluate the performance of the TDx/TDxFLx fetal lung maturity II assay (Abbott Laboratories; Abbott Park, IL) on amniotic fluid (AF) specimens collected vaginally from women with preterm premature rupture of membranes (PROM). METHODS: We reviewed charts of patients with preterm PROM treated at Shands Hospital at the University of Florida from January 1, 1995, to June 30, 1999. Negative predictive values (prediction of the absence of neonatal respiratory distress) of mature (at or above 55 mg/g) and borderline (40-54 mg/g) test results were calculated for 153 women. RESULTS: Respiratory distress syndrome (RDS) occurred with frequencies of one in 42 and three in 29 cases with mature and borderline test results, respectively. All cases of RDS were mild, defined as sustained tachypnea with or without need for supplemental oxygen. With an immature (less than 40 mg/g) test result, 20 of 82 infants developed RDS, and half of those cases were severe, defined as needing mechanical ventilation. Negative predictive values of mature and borderline tests were 97.6% (95% confidence interval [CI] 92.9, 100) and 89.7% (95% CI, 78.3, 100), respectively. CONCLUSION: Mature results from fetal lung maturity tests of vaginally collected AF predict the absence of RDS with a high degree of accuracy. PMID- 10862853 TI - Urinary tract infections during pregnancy and mental retardation and developmental delay. AB - OBJECTIVE: To investigate the association between urinary tract infections during pregnancy and mental retardation or developmental delay in infants. METHODS: An inception cohort design was used to analyze Medicaid maternal and infant-linked records and vital records for 41,090 pregnancies from 1995-1998. RESULTS: The relative risk (RR) for mental retardation or developmental delay among infants of mothers with diagnosed urinary tract infections but no antibiotic claims was 1.31 with a 95% confidence interval (CI) of 1. 12, 1.54 compared with the group without urinary tract infections. The RR for infants of mothers with urinary tract infections without antibiotic claims was 1.22 (95% CI 1.02, 1.46) compared with infants of mothers with urinary tract infections and antibiotic claims. The RR was significant in the first trimester (1.46, 95% CI 1.07, 1.99) and third trimester (1.41, 95% CI 1.11, 1.79) after controlling for race and gestational age at birth. CONCLUSION: There was a statistically significant association between maternal urinary tract infections without evidence of antibiotics and mental retardation or developmental delay in infants. The relationship persisted when we assumed that over 30% of women who had antibiotic claims filled but did not take the medicine, and 40% of the women who did not have antibiotic claims did take the medication. PMID- 10862852 TI - Extra-amniotic saline, laminaria, or prostaglandin E(2) gel for labor induction with unfavorable cervix: a randomized controlled trial. AB - OBJECTIVE: To determine which of three methods of cervical ripening resulted in the lowest cesarean rate in women with unfavorable cervices and indications for labor induction. METHODS: Consenting women with singleton gestations, vertex presentations, and unfavorable cervices (dilatation under 2 cm and effacement under 75%) were randomly assigned to laminaria and standard intravenous oxytocin, serial doses of intracervical prostaglandin (PG) E(2) gel (Prepidil, Pharmacia & Upjohn, Inc., Kalamazoo, MI) 0.5 microg every 6 hours for two doses followed by oxytocin if indicated, or extra-amniotic saline infusion and oxytocin. RESULTS: An interim analysis after recruitment of 321 subjects, 67% of the planned sample, found similar cesarean rates for the three groups (laminaria 36%; PGE(2) gel 33%; saline infusion 29%; P =.59); however, the mean randomization-to-delivery interval was significantly longer in the PGE(2) group. Stochastic curtailment, as part of the interim analysis, indicated a low likelihood of achieving a statistically significant difference in cesarean rates between PGE(2) gel and the other two groups. Therefore, we completed the study with saline infusion and laminaria. The saline infusion and laminaria groups had similar preinduction characteristics. The cesarean rates were similar (saline infusion 25.4% versus laminaria 30.3%; P =.32), but the mean interval from randomization to delivery was shorter in the saline infusion group (18.0 versus 21.5 hours, P =.002). There were no significant differences in selected maternal and neonatal morbidities. CONCLUSION: Cervical ripening with extra-amniotic saline infusion, PGE(2), or laminaria resulted in comparable cesarean rates in women with an unfavorable cervix and indications for labor induction. Extra-amniotic saline infusion had the shortest randomization-to-delivery interval without increasing maternal or neonatal morbidity. PMID- 10862854 TI - Indomethacin tocolysis and risk of necrotizing enterocolitis. AB - OBJECTIVE: To investigate the possible association of indomethacin tocolysis with neonatal necrotizing enterocolitis. METHODS: A case-control study was performed for the period November 1, 1997, through May 1, 1999. All cases of proven necrotizing enterocolitis were ascertained, and four controls for each case were randomly identified from all Special Care Nursery admissions before 37 weeks' gestation without necrotizing enterocolitis during that same period. RESULTS: During the 18-month period there were 24 cases of necrotizing enterocolitis out of 10,200 deliveries. Infants with necrotizing enterocolitis were more preterm (29.7 +/- 3.9 compared with 32.7 +/- 6.0 weeks; P =.03) and had lower birth weights (1453 +/- 777 compared with 1820 +/- 678 g; P =.02) compared with controls (n = 96). Respiratory distress syndrome (RDS) and sepsis were both significantly associated with an increased risk of necrotizing enterocolitis: 16 of 24 cases compared with 40 of 96 controls had RDS (odds ratio [OR] 2.8, 95% confidence interval [CI] 1.0, 8.3) and 14 of 24 cases compared with 11 of 96 controls were septic (OR 10.8, 3.4, 95% CI 34.2). Indomethacin as a single agent was not associated with necrotizing enterocolitis (OR 1.0, 95% CI 0.2, 4.8). Using a logistic regression model, necrotizing enterocolitis was strongly associated with sepsis (adjusted OR 8.5, 95% CI 2.2, 32.5). When sepsis was removed from the model, double-agent tocolytic therapy was significantly associated with necrotizing enterocolitis (adjusted OR 6.9, 95% CI 1.1, 43.6). CONCLUSION: Tocolysis with indomethacin as a single agent was not associated with necrotizing enterocolitis in this case-control study. Combination tocolytic therapy may be a marker for subclinical infection and not causally related to necrotizing enterocolitis. PMID- 10862855 TI - Magnesium sulfate protection of fetal rat brain from severe maternal hypoxia. AB - OBJECTIVE: To determine whether severe maternal hypoxia affects fetal rat physical characteristics and causes neuronal damage, and whether magnesium sulfate can decrease these effects. METHODS: At 17 days gestation, rats were randomly assigned to one of four groups that received saline injections and room air (n = 6), magnesium sulfate and room air (n = 5), saline and hypoxia (n = 5) or magnesium sulfate and hypoxia (n =5). Maternal magnesium sulfate or saline injections were given for 4 hours. In groups 3 and 4 this was followed by a hypoxia chamber protocol that included a gas mixture of 9% oxygen and 3% carbon dioxide for 2 hours. After 72 hours of recovery, fetuses were delivered abdominally, perfused transcardially, and brains removed intact. Fetal body and brain weight and size were measured. Brains were embedded in paraffin, sectioned, and stained. A neuropathologist masked to the protocol performed histologic grading of brain regions. RESULTS: Exposure to the hypoxia chamber resulted in decreased maternal oxygen tension and pH (from 82.8 +/- 20.0 to 49.2 +/- 14.4 mmHg, and from 7.37 +/- 0.05 to 7.20 +/- 0.04, respectively; P <.005). Magnesium sulfate administration resulted in higher magnesium levels in blood (from 1. 52 +/- 0.2 to 3.77 +/- 0.7 mg/dL; P <.001). The hypoxia protocol resulted in a significant decrease in fetal body and brain size, but not weight. Hypoxia also caused an increase in the proportion of fetal rats that had brain injury, including shrinkage of cells and karyorrhexis in the hippocampus and thalamus (from 0% to 38.1% and 38.9%, respectively; P <.05). Magnesium sulfate reduced these effects on fetal brain histopathology and size. CONCLUSION: Severe maternal rat hypoxia resulted in significant fetal neuronal damage and decreased fetal body and brain size. Maternal magnesium sulfate administration reduced the effect of hypoxia on fetal brain histopathology and size without affecting body size. PMID- 10862856 TI - Prevention of perinatal group B streptococcal infection: current controversies. AB - Group B streptococcus (GBS) is the most frequent cause of neonatal sepsis in the United States. The Centers for Disease Control and Prevention (CDC) issued guidelines for its prevention in 1996. This article details areas of controversy with those guidelines and offers recommendations for resolution. We recommend that a prevention policy be adopted by all hospitals. If a screening-based policy is chosen, compliance is essential. Penicillin is the antibiotic of choice for GBS prevention. Increasing resistance to clindamycin and erythromycin might eliminate them as alternative choices in patients allergic to penicillin. Group B streptococcal prophylaxis might not be necessary in women who have repeat elective cesarean delivery. In asymptomatic women, a positive urine culture for GBS should be considered clinically equivalent to a positive vaginal or rectal sample for screening. Neonatal sepsis caused by organisms other than GBS must be monitored carefully by all hospitals providing obstetrics services. PMID- 10862857 TI - Development of an objective structured assessment of technical skills for obstetric and gynecology residents. AB - OBJECTIVE: To develop an objective structured assessment for evaluating surgical skills of obstetrics and gynecology residents and to evaluate the reliability and validity of the assessment. METHODS: A seven-station, objective, structured assessment of technical skills was administered to 24 residents. The test included laparoscopic procedures (port placement, salpingostomy, suturing, vessel ligation) and open abdominal procedures (hypogastric ligation, repair of enterotomy, salpingo-oophorectomy.) All surgical tasks were done on pigs. Residents were timed and assessed at each station using three methods of scoring, a task-specific checklist, global rating scale, and pass-fail grade. RESULTS: Assessment of construct validity (the ability of the test to discriminate among residency levels) found significant differences on the checklist and the global rating scale by residency level. Reliability indices calculated with Cronbach's alpha were 0.89 for the global rating scale and 0.89-0.95 for the individual skills checklists. Interrater reliability was 0.87 for the global rating scale and 0.78-0.98 for the checklists. CONCLUSION: Objective, structured assessment of technical skills can assess residents' surgical skills with high reliability and validity. These assessments have possible application for identifying residents who need additional training and might provide a mechanism to ensure competence of surgical skills. PMID- 10862858 TI - Before Kubler-Ross: lessons about grief from the book of Job. AB - Medicine is as old as the human species, and medical literature is among the earliest writing. Current research is of great help in identifying new interventions, but a great deal of the art of medicine is showcased in ancient works. The 20th century saw a plethora of books, articles, and monographs on the subjects of grief, death and dying, and suffering, but none of these has provided greater insight than the biblical book of Job. Excerpts from Job illustrate both the nature of grief and appropriate intervention when it is confronted. PMID- 10862859 TI - A new plant assemblage (microfossil and megafossil) from the Lower Old Red Sandstone of the Anglo-Welsh Basin: its implications for the palaeoecology of early terrestrial ecosystems. AB - Lower Old Red Sandstone deposits penetrated by a series of cored boreholes near Newport (South Wales) have been sedimentologically logged, and recovered plant assemblages (microfossil and megafossil) investigated. Sedimentological logging indicates that the deposits are typical of the extensive terrestrial-fluviatile floodplain deposits of the Anglo-Welsh Basin. Palynomorph assemblages have been recovered from a number of horizons and comprise entirely terrestrial forms (spores and phytodebris). They essentially represent a single assemblage, belonging to the middle subzone of the micrornatus-newportensis sporomorph assemblage biozone, and indicate an Early Devonian (mid-Lochkovian) age. The new biostratigraphical data enables correlation with other Lower Old Red Sandstone deposits of the Anglo-Welsh Basin, and the deposits are assigned to the lower part of the St. Maughan's Group. A plant megafossil/mesofossil assemblage recovered from one of the spore-bearing horizons includes a zosterophyll assigned to Zosterophyllum cf. fertile. This is the earliest reported zosterophyll from the Anglo-Welsh Basin. The new palynological/palaeobotanical data provide important information on the palaeoecology and palaeobiogeography of the vegetation of the southeastern margin of the Old Red Sandstone continent during Lochkovian times. Palaeogeographical variation in the distribution of plant microfossils and megafossils is interpreted as reflecting differences between the flora of the lowland floodplain and inland intermontaine basins, although this is to a certain extent overprinted by variation due to localized differences in environmental conditions. PMID- 10862860 TI - A new calymmate mimosoid polyad from the Miocene of Argentina. AB - A palynomorph with an unequivocal relationship to the eight-grain polyads of the mimosoid genus Calliandra Benth., is described from the Miocene sediments of San Juan Province, Argentina. Comparison of the fossil palynomorph with polyads of the extant Calliandra species shows a resemblance to those which have one, highly specialized, appendiculate monad. The new palynomorph has a rudimentary appendix, apparently transitional in the path leading to the more highly developed appendiculate forms in the extant Calliandra group. This specialized polyad type is considered to be one of the most highly evolved forms in subfamily Mimosoideae. The closest affinity of the new fossil polyad is with the eight grain calymmate polyads of Calliandra chilensis Benth., a species which has developed in the extra-tropical, xerophilous shrub, habitat of north-central Chile. The disappearance of Calliandra species in San Juan Province is thought to be related to the culmination of the Andean rising, and the consequent interruption to the Pacific Ocean climatic influence. This new discovery is the first fossil record of Calliandra for Argentina, as well as being the most southerly and the oldest. It reinforces the hypothesis of an early origin and diversification for the Leguminosae in Tropical America. PMID- 10862861 TI - Pollen diagrams and prehistoric fields: the case of Bronze Age Haarlem, the Netherlands. AB - The excavation of a Bronze Age field surrounded by peat in the vicinity of the Dutch town of Haarlem afforded a good opportunity to study the pollen rain released by such prehistoric fields. Pollen analysis of a core obtained from a peat deposit at a distance of 10m from the field's border revealed only a weak signal of a possible field. The conclusion is that the presence of prehistoric fields is difficult to detect by means of pollen analysis alone. PMID- 10862862 TI - Evaluation of Holocene pollen records from the Romanian Plain. AB - This study is a critical review of pollen analyses carried out on Holocene sequences from 15 sites in and near the Romanian Plain. Three sites come from natural sediments, 10 sites are from anthropogenic deposits and two are from both anthropogenic and natural settings. The general reconstruction is of a steppe forest-steppe vegetation through the Holocene. The nature of the deposits, however, casts doubts on this reconstruction. Deposits of archaeological sites generally yield pollen spectra that are influenced by human activities and thus unsuitable for vegetation reconstructions. Loess deposits are also unfavorable for pollen preservation because of high pH and porosity. Consequently, pollen spectra from loess deposits are strongly biased by selective pollen destruction. Research and experiments carried out by several authors suggest that spectra dominated by Asteraceae, Poaceae, Chenopodiaceae or Pinus pollen in soils and loess are a result of selective pollen destruction, especially if low pollen concentrations, progressive pollen deterioration or high frequencies of deteriorated or unidentifiable pollen are evidenced. The fact that pollen records from the Romanian Plain come from loess, alkaline peat or archaeological sites reduces their reliability for reconstructions of vegetation. The vegetation history of similar regions in Hungary, Bulgaria and Turkey suggests that early Holocene steppe vegetation was gradually replaced by forest or forest-steppe vegetation in the late Holocene. Records from lake sediments are required to find out whether the Holocene vegetation history of the Romanian Plain was similar. PMID- 10862863 TI - Hypothetical way of pollen aperture patterning. 2. Formation of polycolpate patterns and pseudoaperture geometry. AB - Deviant forms of polycolpate pollen, differing from the typical pattern in the number and arrangement of apertures, are found to be similar in distantly related dicotyledon taxa. The range of variation of common and deviant aperture patterns may be arranged as a continuous series, which may be described as a gradual and geometrically regular transformation of the deviant form with a meridional circular colpus to one of the common polycolpate conditions. Similar series have been observed in the taxa with colporate and pseudocolpate pollen. All possible spatial isomers and their mirror symmetrical variants of the deviant polycolpate and polypseudocolpate pollen have been predicted in terms of the suggested regularities of aperture multiplication. Some of them have been identified in the samples studied. PMID- 10862864 TI - Tritaenia Maegdefrau et Rudolf, Mesozoic 'Sciadopitys-like' leaves in mass accumulations. AB - The Late Jurassic to Early Cretaceous genus Tritaenia Maegdefrau et Rudolf 1969 is problematic because of: (1) missing authentic material of its type species, T. linkii (Roemer 1839) Maegdefrau et Rudolf; and (2) Watson and Harrison's (1998) synonymization of T. linkii with Pseudotorellia heterophylla Watson. This paper: (1) rectifies the status of T. linkii on the basis of newly recovered specimens carrying the original author's authentication; and (2) gives the basis for rejecting Watson and Harrison's claim that T. linkii and T. crassa (Seward) Bose et Manum 1991 represent linear leaves of the heterophyllous taxon Pseudotorellia heterophylla. The three species of Tritaenia known to date (T. crassa, T. linkii, T. scotica) are reviewed, and the genus is compared with other Mesozoic so-called 'Sciadopitys-like' hypostomatic leaves with a median stomatal zone, many of which occur in mass accumulations such as T. linkii. Deciduousness is indicated for T. linkii and T. crassa by their occurrence in mass accumulations and the possession of well-developed abscission scars. Known mass accumulations of fossil foliage are reviewed and their implications for palaeoenvironmental interpretations discussed. PMID- 10862866 TI - Yeast as a Cell Factory. PMID- 10862865 TI - Radiizonates arcuatus, a distinctive new miospore species from the Lower Carboniferous of Western Gondwana. AB - A new species of trilete zonate miospores, Radiizonates arcuatus, is established for Lower Carboniferous Western Gondwanan forms hitherto ascribed misguidedly to Radiizonates genuinus (Jushko) Loboziak and Alpern (1978), a Russian Lower Carboniferous species. The latter binomen is, moreover, not a valid combination and is more correctly designated as Vallatisporites genuinus (Jushko) Byvsheva, 1980. R. arcuatus is, from records to date, confined to westerly parts of Gondwana (Brazil, North Africa and Middle East), in which it is characteristic of Early Carboniferous strata, albeit with some slightly older and slightly younger occurrences. PMID- 10862867 TI - Expression and secretion of human alpha1(I) procollagen fragment by Hansenula polymorpha as compared to Pichia pastoris. AB - Secretion of a human collagen alpha1(I) chain fragment was achieved in Hansenula polymorpha using the native alpha1(I) procollagen secretory signal sequence. The N-terminal propeptide in the fragment was cleaved off during secretion, yielding the N-terminus of mature alpha1(I) collagen. In Pichia pastoris transformants, the expression of the fragment could be detected on RNA-level, but the product could not be determined extracellularly. After fusion of the fragment with a myc HIS6 epitope, the intact product was found intracellularly. The difference in the extracellular level of the protein between the two expression hosts is most likely caused by difference in secretion efficiency. PMID- 10862868 TI - Comparative expression and purification of human glutamic acid decarboxylase from Saccharomyces cerevisiae and Pichia pastoris. AB - The yeast cell factory is a potentially useful source of proteins in general. They include glutamic acid decarboxylase (GAD), which is one of the major autoantigens for Type 1 diabetes. We have created a hybrid form of GAD consisting of amino acids 1-101 of the human GAD67 protein fused to amino acids 96-585 of the human GAD65 protein, and have modified this to include a C-terminal hexa Histidine (H6) tag sequence. This hybrid GAD67/65-H6 was expressed in two yeast hosts: constitutively under the control of the plasmid phosphoglycerate kinase promoter (PGK1) in Saccharomyces cerevisiae, and inducibly under the control of the chromosomal alcohol oxidase promoter (AOX1) in Pichia pastoris. Enzymatically active hybrid GAD was prepared from yeast lysates by purification either on an affinity column based on the GAD-1 monoclonal antibody, or by metal-affinity chromatography. The purified GAD67/65-H6 was radiolabelled with iodine-125 and tested with Type 1 diabetes sera in a radioimmunoprecipitation assay, and results were compared with those using untagged GAD67/65 and those using porcine brain GAD. The results of enzymatic and immunological assays show hybrid GAD67/65 is isolated at high specific activity and moderate yield, and the addition of the H6 tag sequences or the choice of yeast strain did not appreciably affect enzyme activity, percentage recovery of GAD, protein purification, or the utility in diagnosis of diabetes in terms of specificity and sensitivity to the various sera. PMID- 10862869 TI - Heterologous expression of the Kluyveromyces marxianus endopolygalacturonase gene (EPG1) using versatile autonomously replicating vector for a wide range of host. AB - A versatile plasmid shuttle vector system pKDU7 was constructed, which is useful for the heterologous gene expression in a wide range of Kluyveromyces and Saccharomyces strains. This cloning vector was constructed using the 1.6-um circular plasmid pKD1 of Kluyveromyces drosophilarum, the URA3 gene of K. marxianus as well as the pUC19 sequences. The stability of vector in transformants strongly depends on the integrity of the functionally important elements of pKD1. It was shown by comparison of three recombinant vectors, which possessed the pKD1 sequence inserted in different ways. The efficient transformation and stability maintenance of the vector constructed in various strains of Kluyveromyces and Saccharomyces was shown by the expression of the EPG1 gene of the Kluyveromyces marxianus encoding pectin-degrading endopolygalacturonase. PMID- 10862870 TI - Effect of different carbon sources on lipase production by Candida rugosa. AB - Different carbon sources affecting growth and lipase production in Candida rugosa were studied by using batch cultures on defined medium. Carbohydrates and acids non-related to fats did not induce lipase production. The highest yields of enzyme were obtained with lipids or fatty acids as carbon sources. Tween 80 stimulated lipase biosynthesis and secretion outside the cell. Combinations of two types of substrates, carbohydrates and fatty acids, did not improve lipase production, and in some cases, their consumption was produced in a sequential pattern. Glucose presented a repressing effect on lipase production. Moreover, glucose was found to be effective in stimulating lipase secretion by cells with a high level of cell-bound lipase activity because of their previous growth in oleic acid. PMID- 10862871 TI - Use of gene shuffles to study the cytoplasmic transcription system operating on Kluyveromyces lactis linear DNA plasmids. AB - A modified double selection approach for manipulation of the cytoplasmic plasmids k1 and k2 from dairy yeast Kluyveromyces lactis has been exploited to investigate promoter and gene function. Using TRP1-mediated integration of a LEU2 gene fusion, we have shown that expression of the selection marker is strictly dependent on the k2 promoter UCS5. Also, k2ORF6, the gene encoding the RNA polymerase specific for UCS recognition, is functional when shuffled between the plasmids. Once transplaced onto k1 by means of gene shuffling, the hybrid ORF6 complemented an orf6 deletion created on plasmid k2 eventually yielding yeast strains that contained only two recombinant plasmids: a k2 derivative (rk2/6) with a k2orf6::TRP1 gene deletion, and a k1 derivative (rk1/6) carrying the transplaced ORF6 allele along with the LEU2 marker. This interchangeability of both UCS promoter activity and gene function between k2 and k1 supports the concept of an autonomous transcription system that operates on these nonconventional yeast plasmids. PMID- 10862872 TI - Yield improvement of heterologous peptides expressed in yps1-disrupted Saccharomyces cerevisiae strains. AB - Heterologous protein expression levels in Saccharomyces cerevisiae fermentations are highly dependent on the susceptibility to endogenous yeast proteases. Small peptides, such as glucagon and glucagon-like-peptides (GLP-1 and GLP-2), featuring an open structure are particularly accessible for proteolytic degradation during fermentation. Therefore, homogeneous products cannot be obtained. The most sensitive residues are found at basic amino acid residues in the peptide sequence. These heterologous peptides are degraded mainly by the YPS1 encoded aspartic protease, yapsin1, when produced in the yeast. In this article, distinct degradation products were analyzed by HPLC and mass spectrometry, and high yield of the heterologous peptide production has been achieved by the disruption of the YPS1 gene (previously called YAP3). By this technique, high yield continuous fermentation of glucagon in S. cerevisiae is now possible. PMID- 10862873 TI - Influence of magnesium ions on heat shock and ethanol stress responses of Saccharomyces cerevisiae. AB - This study has highlighted the role of magnesium ions in the amelioration of the detrimental effects of ethanol toxicity and temperature shock in a winemaking strain of Saccharomyces cerevisiae. Specifically, results based on measurements of cellular viability and heat shock protein synthesis together with scanning electron microscopy have shown that, by increasing the bioavailability of magnesium ions, physiological protection is conferred on yeast cells. Elevating magnesium levels in the growth medium from 2 to 20 mM results in repression of certain heat shock proteins following a typical heat shock regime (30-42 degrees C shift). Seed inocula cultures prepropagated in elevated levels of magnesium (i.e. 'preconditioned') also conferred thermotolerance on cells and repressed the biosynthesis of heat shock proteins. Similar results were observed in response to ethanol stress. Extra- and intracellular magnesium may both act in the physiological stress protection of yeast cells and this approach offers potential benefits in alcoholic fermentation processes. The working hypothesis based on our findings is that magnesium protects yeast cells by preventing increases in cell membrane permeability elicited by ethanol and temperature-induced stress. PMID- 10862874 TI - Simultaneous genomic overexpression of seven glycolytic enzymes in the yeast Saccharomyces cerevisiae. AB - Fusions of the glycolytic genes TPI1, PGK1, ENO1, PYK1, PDC1, and ADH1 with the lacZ reporter gene of Escherichia coli and a lacZ fusion construct of a 390-bp fragment from the promoter of the HXT7 gene were assayed for beta-galactosidase activity. The glycolytic promoters were induced after addition of glucose to ethanol-grown cells, whereas the HXT7 promoter fragment showed a constitutive beta-galactosidase expression on both carbon sources. The genes coding for the seven enzymes of lower glycolysis Tdh, Pgk, Gpm, Eno, Pyk, Pdc, and Adh were simultaneously put under the control of the same strong promoter, a truncated HXT7 promoter that is constitutively active on ethanol as well as on glucose medium. Genomic expression of the glycolytic genes under the control of this promoter, resulted in an at least 2-fold overexpression. The gene MSG5 was isolated, coding for a protein phosphatase normally involved in cell cycle regulation, as a factor that possibly influences the expression of the HXT7 gene. However, overexpression of MSG5 had no effect on the expression of the HXT7/lacZ fusion, whereas a deletion of this gene resulted in a decreased expression of beta-galactosidase. PMID- 10862875 TI - Respirofermentative metabolism in Kluyveromyces lactis: Insights and perspectives. AB - Yeasts do not form a homogeneous group as far as energy-yielding metabolism is concerned and the fate of pyruvate, a glycolytic intermediate, determines the type of energy metabolism. Kluyveromyces lactis has become an alternative to the traditional yeast Saccharomyces cerevisiae owing to its industrial applications as well as to studies on mitochondrial respiration. In this review we summarize the current knowdeledge about the K. lactis respirofermentative metabolism, taking into account the respiratory capacity of this yeast and the molecular mechanisms controlling its regulation, giving an up-to-date picture. PMID- 10862876 TI - An interlaboratory comparison of physiological and genetic properties of four Saccharomyces cerevisiae strains. AB - To select a Saccharomyces cerevisiae reference strain amenable to experimental techniques used in (molecular) genetic, physiological and biochemical engineering research, a variety of properties were studied in four diploid, prototrophic laboratory strains. The following parameters were investigated: 1) maximum specific growth rate in shake-flask cultures; 2) biomass yields on glucose during growth on defined media in batch cultures and steady-state chemostat cultures under controlled conditions with respect to pH and dissolved oxygen concentration; 3) the critical specific growth rate above which aerobic fermentation becomes apparent in glucose-limited accelerostat cultures; 4) sporulation and mating efficiency; and 5) transformation efficiency via the lithium-acetate, bicine, and electroporation methods. On the basis of physiological as well as genetic properties, strains from the CEN.PK family were selected as a platform for cell-factory research on the stoichiometry and kinetics of growth and product formation. PMID- 10862877 TI - Improved protocols for quantitative determination of metabolites from biological samples using high performance ionic-exchange chromatography with conductimetric and pulsed amperometric detection. AB - Simple and reliable protocols are described for an extensive analysis of metabolites in extracts from different biological sources. The separation was performed by high performance ionic-exchange chromatography (HPIC) at alkaline pH using two types of chromatography columns and two detection methods. Organic acids and inorganic anions were separated on an ionPac AS11 column using a 0.5 to 35 mM Na0H gradient. Detection limits in the range of milligrams per liter were achieved by use of a conductivity detector equipped with an anion self regenerating suppressor. Twelve phosphorylated compounds belonging to the glycolytic and the pentose phosphate pathways could be resolved on a CarboPac PA1 column using a Na0H/Na-acetate gradient. Quantification was achieved by pulsed amperometry with detection limits in the micromolar range. Cell extracts obtained by extraction in boiling buffered ethanol described previously could be directly injected onto HPIC columns for the separation of metabolites because the extraction procedure affected neither the retention time nor the stability of most of the metabolites, and yielded very clean chromatograms. These improved protocols were applied for a dynamic analysis of intracellular metabolites in Saccharomyces cerevisiae in response to a glucose pulse. PMID- 10862878 TI - Regulation of fermentative capacity and levels of glycolytic enzymes in chemostat cultures of Saccharomyces cerevisiae. AB - Regulation of fermentative capacity was studied in chemostat cultures of two Saccharomyces cerevisiae strains: the laboratory strain CEN.PK113-7D and the industrial bakers' yeast strain DS28911. The two strains were cultivated at a fixed dilution rate of 0.10 h(-1) under various nutrient limitation regimes: aerobic and anaerobic glucose limitation, aerobic and anaerobic nitrogen limitation on glucose, and aerobic ethanol limitation. Also the effect of specific growth rate on fermentative capacity was compared in glucose-limited, aerobic cultures grown at dilution rates between 0.05 h(-1) and 0.40 h(-1). Biomass yields and metabolite formation patterns were identical for the two strains under all cultivation conditions tested. However, the way in which environmental conditions affected fermentative capacity (assayed off-line as ethanol production rate under anaerobic conditions) differed for the two strains. A different regulation of fermentative capacity in the two strains was also evident from the levels of the glycolytic enzymes, as determined by in vitro enzyme assays. With the exception of phosphofructokinase and pyruvate decarboxylase in the industrial strain, no clear-cut correlation between the activities of glycolytic enzymes and the fermentative capacity was found. These results emphasise the need for controlled cultivation conditions in studies on metabolic regulation in S. cerevisiae and demonstrate that conclusions from physiological studies cannot necessarily be extrapolated from one S. cerevisiae strain to the other. PMID- 10862879 TI - A short review on the role of glutathione in the response of yeasts to nutritional, environmental, and oxidative stresses. AB - Glutathione (L-gamma-Glutamyl-L-Cysteinylglycine) appears as the major nonprotein thiol compound in yeasts. Recent advances have shown that glutathione (GSH) seems to be involved in the response of yeasts to different nutritional and oxidative stresses. When the yeast Saccharomyces cerevisiae is starved for sulfur or nitrogen nutrients, GSH may be mobilized to ensure cellular maintenance. Glutathione S-transferases may be involved in the detoxification of electrophilic xenobiotics. Vacuolar transport of metal derivatives of GSH ensure resistance to metal stress. Growth of methylotrophic yeasts on methanol results in the formation of an excess formaldehyde that is detoxified by a GSH-dependent formaldehyde dehydrogenase. Growth of yeasts on glycerol results in the accumulation of methylglyoxal detoxified by the glyoxalase pathway. Glutathione per se can react with oxidative agents or is involved in the oxidative stress response through glutathione peroxidase. PMID- 10862880 TI - The influence of hexoses addition on the fermentation of d-xylose in Debaryomyces hansenii under continuous cultivation. AB - The effect of hexoses (glucose and galactose) addition to the feed xylose mineral medium of Debaryomyces hansenii chemostat cultures grown at a constant dilution rate of 0.055 h(-1) was studied. Xylitol was the major product detected amongst all tested conditions. The maximal values for xylitol yield and volumetric productivity (0.56 gg(-1) xylose and 0.21 gl(-1)h(-1), respectively) were obtained for a glucose/xylose feeding ratio of 10%, showing that the addition of small amounts of glucose, but not galactose, enhanced the xylitol production. A xylitol yield increase of 30%, compared with the sole xylose-containing feed medium, was observed. It was found that the oxygen requirement for D. hansenii growth is lower under glucose compared with xylose. Ethanol and glycerol were only produced for glucose/xylose feeding ratio above 30%. The byproducts accumulation was correlated with glucose metabolism, because a direct relationship between the increase of ethanol (and glycerol) concentration and the increase of glucose in the feed medium was found. PMID- 10862881 TI - Mutations in GAL2 or GAL4 alleviate catabolite repression produced by galactose in Saccharomyces cerevisiae. AB - Galactose does not allow growth of pyruvate carboxylase mutants in media with ammonium as a nitrogen source, and inhibits growth of strains defective in phosphoglyceromutase in ethanol-glycerol mixtures. Starting with pyc1, pyc2, and gpm1 strains, we isolated mutants that eliminated those galactose effects. The mutations were recessive and were named dgr1-1 and dgr2-1. Strains bearing those mutations in an otherwise wild-type background grew slower than the wild type in rich galactose media, and their growth was dependent on respiration. Galactose repression of several enzymes was relieved in the mutants. Biochemical and genetic evidence showed that dgr1-1 was allelic with GAL2 and dgr2-1 with GAL4. The results indicate that the rate of galactose consumption is critical to cause catabolite repression. PMID- 10862882 TI - Air pressure effects on biomass yield of two different Kluyveromyces strains. AB - The use of air pressure as a way of improving oxygen transfer in aerobic bioreactors was investigated. To compare the air pressure effects with traditional air bubbled cultures, experiments using a pressure reactor and a stirred flask, with the same oxygen transfer rate, were made. Kluyveromyces marxianus is an important industrial yeast and some of it show a "Kluyver effect" for lactose: even under oxygen limited growth conditions, certain disaccharides that support aerobic, respiratory growth, are not fermented. This study deals with the effect of increased pressure on the physiological behavior of two Kluyveromyces strains: K. marxianus ATCC10022 is a lactose-fermenting strain, whereas K. marxianus CBS 7894 has a Kluyver-effect for lactose. For K. marxianus ATCC10022 an air pressure increase of 2 bar led to a 3-fold increase in biomass yield. When air pressure increased an enhancement of ethanol oxidation of cell yeasts was also observed. Batch cultures of K. marxianus CBS 7894 exhibited different growth behaviour. Its metabolism was always oxidative and ethanol was never produced. With the increase in air pressure, it was possible to increase the productivity in biomass of K. marxianus CBS 7894. As a response to high oxygen concentrations, due to the increase in oxygen partial pressure, oxidative stress in the cells was also studied. Antioxidant defences, such as superoxide dismutase, catalase, and glutathione reductase, were at high activity levels, suggesting that these yeast strains could tolerate the increased pressures applied. PMID- 10862883 TI - Enrichment of threonine content in Saccharomyces cerevisiae by pathway engineering. AB - In a previous work, we have investigated the effect of amplifying individually the genes of the threonine biosynthetic pathway on threonine accumulation by yeast. Here, we present the results of the simultaneous amplification of these genes in strains with different genetic backgrounds. These strains carry a mutant HOM3-R2 allele (coding for a feedback-insensitive aspartate kinase), and/or a mutant cha1 allele that makes it defective in threonine degradation by the catabolic L-serine (L-threonine) deaminase. The results show that the amplification of the clustered genes affects threonine and homoserine accumulation only when it includes the HOM3 gene, or when combined with a HOM3-R2 mutation. Similarly, the cha1 mutation is only effective when a certain amount of threonine is reached. Threonine overproduction affects other cellular functions such as the accumulation of other amino acids, the cell growth and metabolite excretion, probably reflecting a redirection of the carbon flux in the central metabolism. PMID- 10862884 TI - Regulation of primary carbon metabolism in Kluyveromyces lactis. AB - In the recent past, through advances in development of genetic tools, the budding yeast Kluyveromyces lactis has become a model system for studies on molecular physiology of so-called "Nonconventional Yeasts." The regulation of primary carbon metabolism in K. lactis differs markedly from Saccharomyces cerevisiae and reflects the dominance of respiration over fermentation typical for the majority of yeasts. The absence of aerobic ethanol formation in this class of yeasts represents a major advantage for the "cell factory" concept and large-scale production of heterologous proteins in K. lactis cells is being applied successfully. First insight into the molecular basis for the different regulatory strategies is beginning to emerge from comparative studies on S. cerevisiae and K. lactis. The absence of glucose repression of respiration, a high capacity of respiratory enzymes and a tight regulation of glucose uptake in K. lactis are key factors determining physiological differences to S. cerevisiae. A striking discrepancy exists between the conservation of regulatory factors and the lack of evidence for their functional significance in K. lactis. On the other hand, structurally conserved factors were identified in K. lactis in a new regulatory context. It seems that different physiological responses result from modified interactions of similar molecular modules. PMID- 10862885 TI - Production of a heterologous endo-1,4-beta-xylanase in the yeast Pichia stipitis with an O(2)-regulated promoter. AB - The Cryptococcus albidus XLN-gene (encoding endo-1,4-beta-xylanase) was expressed in the yeast Pichia stipitis under the control of the PsADH2-promoter, which is activated under O(2) limitation. The resulting transformant produced endo-1,4 beta-xylanase after a shift to anoxic conditions. Endo-1,4-beta-xylanase production was enhanced by limited aeration after the shift. PMID- 10862886 TI - The role of metabolic engineering in the improvement of Saccharomyces cerevisiae: utilization of industrial media. AB - Metabolic engineering has become a very important approach to strain improvement in parallel with classical strain development. Although Saccharomyces cerevisiae has been domesticated for ethanol and bread production, there are still some fundamental problems associated with its industrial use. The industrially used carbon sources often consist of a sugar mixture, and due to glucose repression these sugars are utilized sequentially, resulting in prolonged production time. In this article we discuss the application of metabolic engineering for construction of glucose-derepressed strains and specify advantages as well as difficulties associated with this approach. PMID- 10862887 TI - The methylotrophic yeast Hansenula polymorpha: a versatile cell factory. AB - The development of heterologous overexpression systems for soluble proteins has greatly advanced the study of the structure/function relationships of these proteins and their biotechnological and pharmaceutical applications. In this paper we present an overview on several aspects of the use of the methylotrophic yeast Hansenula polymorpha as a host for heterologous gene expression. H. polymorpha has been successfully exploited as a cell factory for the large-scale production of such components. Stable, engineered strains can be obtained by site directed integration of expression cassettes into the genome, for which various constitutive and inducible promoters are available to control the expression of the foreign genes. New developments have now opened the way to additional applications of H. polymorpha, which are unprecedented for other organisms. Most importantly, it may be the organism of choice for reliable, large-scale production of heterologous membrane proteins, using inducible intracellular membranes and targeting sequences to specifically insert these proteins stably into these membranes. Furthermore, the use of H. polymorpha offers the possibility to accumulate the produced components into specific compartments, namely peroxisomes. These organelles are massively induced during growth of the organism on methanol and may occupy up to 80% of the cell volume. Accumulation inside peroxisomes prevents undesired modifications (e.g. proteolytic processing or glycosylation) and is also in particular advantageous when proteins are produced which are toxic or harmful for the host. PMID- 10862888 TI - Accelerated prediction of recombinant protein production in Saccharomyces cerevisiae by using rapid monitoring techniques. AB - The use of a stopped-flow analyser for the monitoring of the production of a secreted recombinant protein, a wild-type cutinase, from S. cerevisiae CEN.PK111 32D pUR7320 is described. Induction is through use of a galactose promoter, and the monitoring facility is used to record the formation of the cutinase and cell density with optical density measurements. A range of induction conditions was studied with a view to using the monitoring to predict the likely level of cutinase formation. Results achieved within 4 to 5 h of induction were of sufficient quality to allow the use of simple modelling relating cutinase formation and cell production to predict likely final specific activities of the product. The utility of such monitoring and prediction is discussed with regards to improved process confidence and definition during fermentation production. PMID- 10862889 TI - Yeast cell attachment: a tool modulating wall composition and resistance to 5 bromo-6-azauracil. AB - The attachment of Candida utilis, Kluyveromyces lactis, and Saccharomyces cerevisiae cells stimulates an increase in the content of cell wall polysaccharides and mannoproteins, accompanied by increased resistance to the inhibitory effect of 5-bromo-6-azauracil. The covalent attachment of viable yeasts was accomplished (via dialdehyde-amino spacers) by reaction of aldehyde groups of the carrier with reactive amino groups in accessible cell surface proteins. The employed technique enables the optimization of yeast sources of beta-1,3-, beta-1,6- glucans, mannan, and mannoprotein. The modulatory effect of the cell attachment is discussed. PMID- 10862890 TI - From gene to product in yeast: production of fungal cutinase. AB - In the mid-1970s, information technology and recombinant DNA technology were considered as the breakthrough technologies of the final quarter of the 20th century. Now, about 25 years later, information technology has penetrated deeply into our society and nearly everyone uses this technology. Compared to the formidable success of information technology, the progress in the commercialization of recombinant DNA technology is moderate, even when taking into account that all that is related to the technological application of biological sciences needs extensive safety testing. However, there are signs that the speed of this commercialization will increase in the first decade of the 21st century. Moreover, new breakthroughs in our understanding of the complete genetic make up of eukaryotes will contribute to this increase in speed. An important aspect of the commercialization of this technology is the development of cells as factories for the production of valuable and/or useful molecules. Lower eukaryotes, such as yeasts and molds, are the most promising candidates to become the factories of the future, but at present these factories still contains a lot of process lines that may be superfluous under the well controlled conditions in fermentors. On the other hand, the speed and yield of these cellular production lines can be increased by eliminating the rate-determining steps of these process lines. In this contribution to the European Union symposium from Cell to Factory, some steps in the improvement of S. cerevisiae as cell factories for (heterologous) hydrophobic molecules are presented. PMID- 10862891 TI - Nutrient-induced signal transduction through the protein kinase A pathway and its role in the control of metabolism, stress resistance, and growth in yeast. AB - Yeast cells growing in the presence of glucose or a related rapidly-fermented sugar differ strongly in a variety of physiological properties compared to cells growing in the absence of glucose. Part of these differences appear to be caused by the protein kinase A (PKA) and related signal transduction pathways. Addition of glucose to cells previously deprived of glucose triggers cAMP accumulation, which is apparently mediated by the Gpr1-Gpa2 G-protein coupled receptor system. However, the resulting effect on PKA-controlled properties is only transient when there is no complete growth medium present. When an essential nutrient is lacking, the cells arrest in the stationary phase G0. At the same time they acquire all characteristics of cells with low PKA activity, even if there is ample glucose present. When the essential nutrient is added again, a similar PKA dependent protein phosphorylation cascade is triggered as observed after addition of glucose to glucose-deprived cells, but which is not cAMP-mediated. Because the pathway involved requires a fermentable carbon source and a complete growth medium, at least for its sustained activation, it has been called "fermentable growth medium (FGM)-induced pathway." PMID- 10862892 TI - Mathematical modeling of citric acid production by repeated batch culture. AB - A mathematical model has been created for the process of citric acid biosynthesis by yeast (mutant strain Yarrowia lipolytica) cultivated by the repeated batch (RB) method on ethanol under conditions of nitrogen limitation. The model accounts for cell growth as a function of nitrogen concentration in the culture liquid; nitrogen uptake by growing cells; citric acid production; pH control in the fermentor by means of NaOH addition; and changes in system volume. The model represents a system of five nonlinear differential equations. Experimental measurements of cell concentration, citric acid concentration, and cultivation broth volume were used with the least squares method to determine the values of eight model parameters. The parameter values obtained were consistent with literature data and general concepts of cell growth and citric acid biosynthesis. The model has been used to predict optimum RB culture conditions. PMID- 10862893 TI - Continuous cider fermentation with co-immobilized yeast and Leuconostoc oenos cells. AB - Ca-alginate matrix was used to co-immobilize Saccharomyces bayanus and Leuconostoc oenos in one integrated biocatalytic system in order to perform simultaneously alcoholic and malo-lactic fermentation of apple juice to produce cider, in a continuous packed bed bioreactor. The continuous process permitted much faster fermentation compared with the traditional batch process. The flavor formation was also better controlled. By adjusting the flow rate of feeding substrate through the bioreactor, i.e. its residence time, it was possible to obtain either "soft" or "dry" cider. However, the profile of volatile compounds in the final product was modified comparatively to the batch process, especially for higher alcohols, isoamylacetate, and diacetyl. This modification is due to different physiology states of yeast in two processes. Nevertheless, the taste of cider was quite acceptable. PMID- 10862894 TI - Lactic acid bacteria as a cell factory: rerouting of carbon metabolism in Lactococcus lactis by metabolic engineering. AB - Lactic acid bacteria display a relatively simple metabolism wherein the sugar is converted mainly to lactic acid. The extensive knowledge of metabolic pathways and the increasing information of the genes involved allows for the rerouting of natural metabolic pathways by genetic and physiological engineering. We discuss several examples of metabolic engineering of Lactococcus lactis for the production of important compounds, including diacetyl, alanine and exopolysaccharides. PMID- 10862895 TI - Protease produced by Pseudomonas aeruginosa K-187 and its application in the deproteinization of shrimp and crab shell wastes. AB - In addition to chitinase/lysozyme, Pseudomonas aeruginosa K-187 also produced a protease useful for the deproteinization of shrimp and crab shell wastes. The optimal culture conditions for P. aeruginosa K-187 to attain the highest protease activity were investigated and discussed. The highest protease activity was as high as 21.2 U/ml, 10-fold that (2.2 U/ml) obtained prior to optimization. The protease of P. aeruginosa K-187, produced under the optimal culture conditions, was tested for crustacean waste deproteinization. The percent of protein removal for shrimp and crab shell powder (SCSP) after 7-day incubation was 72%, while that of natural shrimp shell (NSS) and acid-treated SCSP was 78% and 45%, respectively. In contrast, with the protease produced under pre-optimization conditions, the percent of protein removal for SCSP, NSS, and acid-treated SCSP was 48%, 55%, and 40%, respectively. For comparison, three other protease producing microbes were tested for crustacean waste deproteinization. However, they were shown to be less efficient in deproteinization than P. aeruginosa K 187. The crude protease produced by P. aeruginosa K-187 can be covalently immobilized on a reversibly soluble polymeric support (hydroxypropyl methycellulose acetate succinate). The immobilized enzyme was soluble above pH 5.5 but insoluble below pH 4.5. Immobilization efficiency was 82%. The immobilized enzyme was stable between pH 6 and 9 and at temperatures below 60 degrees C. The optimum pH and temperature for the immobilized enzyme was pH 8 and 50 degrees C. The half-life of the immobilized enzyme was 12 days, longer than that of free protease (8 days). The utilization of the immobilized enzyme for the deproteinization of SCSP has resulted in a 67% protein removal. By contrast, SCSP protein removal by using free enzymes was 72%. The protease was further purified and characterized. The purification steps included ammonium sulfate precipitation, DEAE-Sepharose CL-6B ion-exchange chromatography, and Sephacryl S 200 gel-permeation chromatography. The enzyme had a molecular weight estimated to be 58.8 kDa by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified enzyme was active from pH 7 to 9 and its optimal pH was 8. PMID- 10862896 TI - Effect of thermal and chemical denaturants on Thermoanaerobacter ethanolicus secondary-alcohol dehydrogenase stability and activity. AB - Thermoanaerobacter ethanolicus 39E secondary-alcohol dehydrogenase (2 degrees ADH) was optimally active near 90 degrees C displaying thermostability half-lives of 1.2 days, 1.7 h, 19 min, 9.0 min, and 1.3 min at 80 degrees C, 90 degrees C, 92 degrees C, 95 degrees C, and 99 degrees C, respectively. Enzyme activity loss upon heating (90-100 degrees C) was accompanied by precipitation, but the soluble enzyme remaining after partial inactivation retained complete activity. Enzyme thermoinactivation was modeled by a pseudo-first order rate equation suggesting that the rate determining step was unimolecular with respect to protein and thermoinactivation preceded aggregation. The apparent 2 degrees ADH melting temperature (T(m)) occurred at approximately 115 degrees C, 20 degrees C higher than the temperature for maximal activity, suggesting that it is completely folded in its active temperature range. Thermodynamic calculations indicated that the active folded structure of the 2 degrees ADH is stabilized by a relatively small Gibbs energy (triangle upG(stab.)(double dagger) = 110 kJ mol(-1)). 2 degrees ADH catalytic activities at 37 degrees C to 75 degrees C, were 2-fold enhanced by guanidine hydrochloride (GuHCl) concentrations between 120 mM and 190 mM. These results demonstrate the extreme resistance of this thermophilic 2 degrees ADH to thermal or chemical denaturation; and suggest increased temperature or GuHCl levels seem to enhance protein fixability and activity. PMID- 10862897 TI - Selective production of L-aspartic acid and L-phenylalanine by coupling reactions of aspartase and aminotransferase in Escherichia coli. AB - With L-aspartate (L-Asp) as the amino donor, L-phenylalanine (L-Phe) can be prepared from phenylpyruvate (PPA) via an amination reaction mediated by aminotransferase (encoded by aspC). On the other hand, L-Asp can be produced by an aspartase (encoded by aspA) -catalyzed reaction using fumaric acid as substrate. To overproduce aspartase in Escherichia coli, the aspA gene was cloned and overexpressed 180 times over the wild-type level. The use of AspA overproducing E. coli strain for L-Asp production exhibited an 83% conversion, approaching to the theoretical yield, whereas the wild-type strain obtained scarcely L-Asp. Furthermore, the recombinant strain overproducing both AspA and AspC was able to produce L-Asp and L-Phe simultaneously by using fumaric acid and PPA as substrates. As a result, the conversion yields obtained for L-Asp and L Phe were 78% and 85%, respectively. In sharp contrast, the wild-type strain attained a conversion of L-Phe less than 15% and an undetectable level of L-Asp. This result illustrates a potential and attractive process to yield both L-Asp and L-Phe by coupling AspA and AspC. A further study on the repeated use of the recombinant strain immobilized with calcium alginate showed that after eight batch runs L-Asp conversion maintained roughly constant (around 75%), whereas L Phe conversion dropped to 65% from 81%. This result indicates the stability of AspA being superior to AspC. PMID- 10862898 TI - Immobilization/stabilization on Eupergit C of the beta-galactosidase from B. circulans and an alpha-galactosidase from Aspergillus oryzae. AB - Two synthetically useful glycosidases, the beta-galactosidase from Bacillus circulans and an alpha-galactosidase from Aspergillus oryzae have been immobilized on Eupergit C. The immobilized enzymes retain high catalytic activity and show increased thermal stability compared with the free enzymes. PMID- 10862899 TI - Preparation of alkyl (R)-(-)-3-hydroxybutyrate by acidic alcoholysis of poly-(R) (-)-3-hydroxybutyrate. AB - An efficient method for the preparation of optically active alkyl (R)-(-)-3 hydroxybutyrates by chemical depolymerization of biopolymer, poly-(R)-(-)-(3 hydroxybutyrate), was established. This method consists of simple recovery of poly-(R)-(-)-(3-hydroxybutyrate) from bacterial cells followed by acidic alcoholysis. When poly-(R)-(-)-(3-hydroxybutyrate) was purified by a simple digestion method that used 0.2 N sodium hydroxide, alkyl (R)-(-)-hydroxybutyrates were most efficiently produced by alcoholysis with anhydrous hydrochloric acid. PMID- 10862900 TI - Kinetics of sequential nitrification and denitrification processes. AB - Kinetics of nitrification and denitrification of synthetic wastewater was investigated by using two reactors in series. An activated sludge unit was used for nitrification followed by a downflow biofilter for denitrification. Glucose solution was fed to the denitrification column to supply carbon source. Reactors were operated at different operating conditions and data were collected for determination of kinetic constants. Experimental data indicated that nitrification and denitrification kinetics followed Monod kinetics. By using the experimental data, kinetic constants for nitrification were determined as k = 1.15 d(-1), K(N) = 5.14 mg/l, Y = 0.34 mgX/mgN and b = -0.021 d(-1). Similarly, kinetic constants for denitrification were determined as k = 0.23 d(-1) and K(DN) = 0.27 mg/l. Rates of nitrification and denitrification increased with increasing nitrogen loading rate. PMID- 10862901 TI - Effects of ammonia and lactate on growth, metabolism, and productivity of BHK cells. AB - The aim of the present work was to study the effect of ammonia and lactate on growth, metabolism, and productivity of BHK cells producing a recombinant fusion protein. Results show that cell growth was reduced with the increase in ammonia or lactate: k(1/2) of 1.1 mM and 3.5 mM for stirred and stationary cultures, respectively, for ammonia and of 28 mM for both stationary and stirred cultures for lactate, were obtained. The cell-specific consumption rates of both glucose (q(Glc)) and glutamine (q(Gln)) increased, whereas that of oxygen (q(O2)) decreased, with the increase in ammonia or lactate concentrations. The cell specific production rates of lactate (q(Lac)) increased with an increase in ammonia concentration; similarly for the cell-specific production rates of ammonia (q(Amm)), which also increased with an increase in lactate concentration; on the other hand, both q(Lac) and q(Amm) markedly decreased when lactate or ammonia concentrations were increased, respectively; lactate was consumed at lactate concentrations above 30 mM and ammonia was consumed at ammonia concentrations above 5 mM. In vivo (31)P NMR experiments showed that ammonia and lactate affect the intracellular pH, leading to intracellular acidification, and decrease the content in phosphomonoesters, whereas the cell energy state was maintained. The effect of lactate on cell growth and q(Gln) is partially due to osmolarity, on q(Glc) and q(Amm) is entirely due to osmolarity, but on q(Lac) is mainly due to lactate effect per se. An increase in ammonia from 0 to 20 mM induced a 50% reduction in specific productivity, whereas an increase in lactate from 0 to 60 mM induced a 40% decrease. PMID- 10862902 TI - Exploiting unusual affinity of usual polysaccharides for separation of enzymes on fluidized beds. AB - Two polysaccharides, alginate and chitosan, showed unusual affinity and bound alpha-amylase (from various sources) and Aspergillus niger cellulase, respectively. The beads prepared from these polymers were successfully used for the purification of the respective enzymes by fluidized bed affinity chromatography. alpha-amylase from wheat germ could be purified by 58-fold with about 90% recovery of activity. Aspergillus niger cellulase, on the other hand, was purified by 30-fold with 80% recovery of enzyme activity. Both purified preparations show single band on SDS-PAGE. PMID- 10862903 TI - Heat-labile bacterial alkaline phosphatase from a marine Vibrio sp. AB - Psychrophilic organisms have successfully adapted to various low-temperature environments such as cold ocean waters. Catalysts with increased catalytic efficiencies are produced, generally at the expense of thermal stability due to fewer non-covalent stabilizing interactions. A marine bacterial strain producing a particularly heat-labile alkaline phosphatase was selected from a total of 232 strains isolated from North-Atlantic coastal waters. From partial 16S rRNA sequences the strain was characterized as a Vibrio sp. An alkaline phosphatase was purified 151-fold with 54% yield from the culture medium using a single step affinity chromatography procedure on agarose-linked L histidyldiazobenzylphosphonic acid. The active enzyme was a 55 +/- 6 kDa monomer. The enzyme had optimal activity at pH 10 and was strikingly heat-labile with a half-life of 6 min at 40 degrees C and 30 min at 32 degrees C. This enzyme from Vibrio sp. had a higher turnover number (k(cat)) and higher apparent Michaelis Menten factor (K(m)) than the enzyme from Escherichia coli, a clear-indication of cold-adaptation. Inorganic phosphate was a competitive inhibitor with a relatively high K(i) value of 1.7 mM. Low affinity for phosphate may contribute to higher turnover rates due to more facile release of product. PMID- 10862904 TI - Production, properties and application to nonaqueous enzymatic catalysis of lipase from a newly isolated Pseudomonas strain. AB - A potent bacterium for lipase production was isolated from soil and identified as Pseudomonas species. It produced lipase constitutively. A mutant of this strain with a lipase productivity 3.25-fold higher was obtained by treatment with ultraviolet (UV) and nitrosoguanidine (NTG). Its fermentation condition was optimized to a lipase yield of 87.5 U/ml. The lipase had maximum activity at pH 9.0 and 45 degrees C. It was stable at pHs from 7.0 to 11.0 and below 60 degrees C. The effects of metal ions, surfactants and bile salts were also studied. The lipase was 1,3-specific. In organic solvents, the thermal stability of the lipase was significantly enhanced. Its optimum temperature was also slightly increased. The optimum water activity was found between 0.5 and 0.6. The lipase was successfully applied in organic phase to catalyze the glycerolysis of palm oil for monoglyceride (MG) production, and the enantioselective esterification of (R,S)-2-octanol. The enantioselectivity of the lipase could be enhanced substantially by treatment with an amphipathic. PMID- 10862905 TI - Characterization of a solvent resistant and thermostable aminopeptidase from the hyperthermophillic bacterium, Aquifex aeolicus. AB - A leucine aminopeptidase gene of Aquifex aeolicus, a hyperthermophilic bacterium, was cloned and expressed in Escherichia coli, and its expression product was purified and characterized. The expressed protein was purified to homogeneity by using heat to denature contaminating proteins followed by ion-exchange chromatography to purify the heat-stable product. The purified enzyme gave a single band on SDS-PAGE with a molecular weight of 54 kDa. Kinetic studies on the purified enzyme confirmed that it was a leucine aminopeptidase. The optimum temperature for its activity was around 80 degrees C and the optimum pH was in the range from 8.0 to 8.5. It was stable at high temperatures and 27% of its activity was retained after heating at 115 degrees C for 30 min. The purified enzyme had a pH stability range between 4.0 and 11.0. This aminopeptidase was highly resistant to organic solvents such as methanol, ethanol, tetrahydrofuran, dimethyl sulfoxide, acetone, acetonitrile, dimethyl formamide, 1-propanol, 2 propanol, and dioxane. PMID- 10862906 TI - Relationships between activities of xylanases and xylan structures. AB - Structures of five water-soluble xylans have been determined. Four purified xylanase enzymes have been studied for the hydrolysis of the xylans. Different xylanases have different activities against various xylan structures. The key factors that influence the rate of xylan hydrolysis are chain length and degree of substitution. Two family 11 xylanases, Orpinomyces pc2 xylanase and Trichoderma longibrachiatum xylanase, can rapidly hydrolyze xylans that have a chain length greater than 8 xylose residues, and their hydrolytic rates are not sensitive to substituents on the xylan backbone. A family 11 xylanase from Aureobasidium pullulans is most effective on xylans that have a long chain (greater than 19 xylose residues), and also is effective against substituent groups. Although Thermatoga maritima xylanase is also more active on a long xylan chain (greater than 19 xylose residues), its hydrolytic rate is greatly reduced by substituents on xylan backbones. PMID- 10862907 TI - Electroelution as a simple and fast protein purification method: isolation of an extracellular xylanase from Bacillus sp. CCMI 966. AB - An efficient and simple modified method of electroelution is described that can be used as a time-saving method for eluting multiple protein bands. Provided that the proteins are highly expressed, they can be purified rapidly and without requiring any prior knowledge of the protein characteristics. A xylanase excreted by Bacillus sp. CCMI 966 was purified directly from the polyacrylamide gel. Some of the properties of this enzyme are presented. It had an unusually apparent high molecular mass of 340kDa, as determined by native PAGE. The specific activity of the purified xylanase was 137 U/mg. PMID- 10862908 TI - Transformation and degradation of the disazo dye Chicago Sky Blue by a purified laccase from Pycnoporus cinnabarinus. AB - The degradation of the disazo dye Chicago Sky Blue 6B by a purified laccase from Pycnoporus cinnabarinus was investigated. Laccase was purified to homogeneity and characterized. The enzyme had a molecular size of 63 kDa as determined by SDS PAGE and an isoelectric point at pH 3. Amino acid composition and N-terminal amino acid sequence was shown to be similar to other fungal laccases. The purified laccase was stable for 1 h at 60 degrees C and was irreversibly inactivated by sodium azide at 0.1 mM. Laccase was shown to initiate destruction of the chromophore of the disazo dye Chicago Sky Blue, resulting in the formation of two intermediate products with absorption intensities about one order of magnitude lower than the parent molecule. The rate at which the dye was transformed by purified laccase was shown to increase with increasing concentrations of the enzyme. PMID- 10862909 TI - At what temperature can enzymes maintain their catalytic activity? AB - It was shown by the combination of thermogravimetric analysis and Karl Fisher titrations that temperatures in excess of 200 degrees C are required to remove tightly bound water from proteins. The heating of enzymes to this temperature caused no cleavage of the polypeptide chains and very little, if any, chemical degradation of particular amino acid residues as judged by electrophoretic and amino acid analysis respectively. It was hypothesised that those enzymes that require very little water for their catalytic activity, should remain active at such elevated temperatures provided that they can be stabilised against thermodenaturation. This conclusion has been verified by the observation that immobilised Candida antarctica lipase catalysed transesterification of octadecanol with palmityl stearate at 130 degrees C for a considerable period of time. PMID- 10862910 TI - A bifunctional beta-xylosidase-xylose isomerase from Streptomyces sp. EC 10. AB - beta-Xylosidase (1,4-beta-D-xylan xylohydrolase EC 3.2.1.37) and xylose isomerase (D-xylose ketol-isomerase EC 5.3.1.5) produced by Streptomyces sp. strain EC 10, were cell-bound enzymes induced by xylan, straw, and xylose. Enzyme production was subjected to a form of carbon catabolite repression by glycerol. beta Xylosidase and xylose isomerase copurified strictly, and the preparation was found homogeneous by gel electrophoresis after successive chromatography on DEAE Sephacel and gel filtration on Biogel A. Streptomyces sp. produced apparently a bifunctional beta-xylosidase-xylose isomerase enzyme. The molecular weight of the enzyme was measured to be 163,000 by gel filtration and 42,000 by SDS-PAGE, indicating that the enzyme behaved as a tetramer of identical subunits. The Streptomyces sp. beta-xylosidase was a typical glycosidase acting as an exoenzyme on xylooligosaccharides, and working optimally at pH 7.5 and 45 degrees C. The xylose isomerase optimal temperature was 70 degrees C and maximal activity was observed in a broad range pH (5-8). Enhanced saccharification of arabinoxylan caused by the addition of the enzyme to endoxylanase suggested a cooperative enzyme action. The first 35 amino acids of the N-terminal sequence of the enzyme showed strong analogies with N-terminal sequences of xylose isomerase produced by other microorganisms but not with other published N-terminal sequences of beta xylosidases. PMID- 10862911 TI - Prediction of penicillin V acylase stability in water-organic co-solvent monophasic systems as a function of solvent composition. AB - Hydrolytic activity of penicillin V acylase (EC 3.5.1.11) can be improved by using organic cosolvents in monophasic systems. However, the addition of these solvents may result in loss of stability of the enzyme. The thermal stability of penicillin V acylase from Streptomyces lavendulae in water-organic cosolvent monophasic systems depends on the nature of the organic solvent and its concentration in the media. The threshold solvent concentration (at which half enzymatic activity is displayed) is related to the denaturing capacity of the solvent. We found out linear correlations between the free energy of denaturation at 40 degrees C and the concentration of the solvent in the media. On one hand, those solvents with logP values lower than -1.8 have a protective effect that is enhanced when its concentration is increased in the medium. On the other hand, those solvents with logP values higher than -1.8 have a denaturing effect: the higher this value and concentration, the more deleterious. Deactivation constants of PVA at 40 degrees C can be predicted in any monophasic system containing a water-miscible solvent. PMID- 10862912 TI - Biological detoxification of coffee husk by filamentous fungi using a solid state fermentation system. AB - Studies were carried out on detoxification of coffee husk in solid state fermentation using three different strains of Rhizopus, Phanerochaete, and Aspergillus sp. Fungal strains were selected by their ability to grow on a coffee husk extract-agar medium. Using R. arrizus LPB-79, the best results on the degradation of caffeine (87%) and tannins (65%) were obtained with pH 6.0 and moisture 60% in 6 days. When P. chrysosporium BK was used, maximum degradation of caffeine and tannins were 70.8 and 45%, respectively, with coffee husk having 65% moisture and pH 5.5 in 14 days. The Aspergillus strain, isolated from the coffee husk, showed best biomass formation on coffee husk extract-agar medium. Optimization assays were conducted using factorial design, and surface response experiments with Aspergillus sp. The best detoxification rates achieved were 92% for caffeine and 65% for tannins. The results showed good prospects of using these fungal strains, in particular Aspergillus sp., for the detoxification of coffee husk. PMID- 10862913 TI - Chiral epoxide production using mycobacterium solubilized in a water-in-oil microemulsion. AB - The application of many biotransformation processes is limited because the substrates/products are poorly water soluble, can be further metabolized, or are inhibitory. Hence non-aqueous media (e.g. two-phase systems, low water environments) are being examined to determine whether they can be used to overcome these problems. One novel approach is to encapsulate whole cells in water-in-oil (w/o) microemulsions (reverse micelles). In this study we have investigated the influence of key system parameters on system stability and epoxidation activity of Mycobacterium M156 cells in reverse micelles comprised of a mixture of Tween 85 and Span 80 (10-20 w%, with an hydrophilic/lipophilic balance [HLB] of 10 and a weight ratio of Tween 85 to Span 80 = 5.7) in n hexadecane. It was found that the minimum allyl phenyl ether (APE) concentration required in the bulk hexadecane solvent phase for epoxidation to occur was 15 mM, whereas the minimum molar ratio of water to surfactant (W(0)) was 35. The optimum epoxidation rate achieved was 3.8 nmol/mg dwt-min with an APE concentration of 50 mM, and a W(0) of 50, with an enantiomeric excess (ee) of 86%. However, epoxidation was found to terminate approximately 3 h after initiation, and the causes for this were postulated to be either: the deleterious effect of the solvent on the Mycobacteria; inactivation of the energy generating system; an insufficient energy supply, or; the instability of the monooxygenase enzyme. It was concluded that on balance emulsion systems are not an economically viable system for producing phenyl glycidyl ether (PGE). PMID- 10862914 TI - Properties of soluble alpha-chymotrypsin in neat glycerol and water. AB - UV scanning of alpha-chymotrypsin dissolved in neat glycerol and water showed no significant differences in its spectra at pH 7.8. Fluorescence scanning revealed a strong dependence on pH values (between 5.9 to 10.5) of the maximum wavelength emission in water and no pH-dependence in 99% glycerol supplemented with 1% of appropriate buffers. The profile of alpha-chymotrypsin activity dissolved in water-glycerol mixtures with phenyl acetate as substrate displayed two maximum: highest peak was found at 100% water, and the second one was observed in 99% glycerol concentration with about 40% of the relative activity. Optimum pH of the soluble alpha-chymotrypsin in glycerol showed a displacement of 1 pH/U towards the alkaline side compared to water at pH 8.0. Kinetic and thermodynamic analysis using kinetic measurements of the thermal stability of alpha-chymotrypsin showed a higher inactivation rate in neat glycerol as compared to water in 30 to 45 degrees C range, however, when temperature increases enzyme stability in glycerol is better than water. Thermostability of trypsin and alpha-chymotrypsin dissolved in glycerol at 100 degrees C showed a half reaction time of approximately 7 and 20 h, respectively, and less than 1 minute in aqueous buffer for both enzymes. PMID- 10862915 TI - Regulation of aspartate-derived amino-acid metabolism in Zygosaccharomyces rouxii compared to Saccharomyces cerevisiae. AB - To elucidate the growth inhibitory effect of threonine, the regulation of the aspartate-derived amino-acid metabolism in Zygosaccharomyces rouxii, an important yeast for the flavor development in soy sauce, was investigated. It was shown that threonine inhibited the growth of Z. rouxii by blocking the methionine synthesis. It seemed that threonine blocked this synthesis by inhibiting the conversion of aspartate. In addition, it was shown that the growth of Z. rouxii, unlike that of Saccharomyces cerevisiae, was not inhibited by the herbicide sulfometuron methyl (SMM). From enzyme assays, it was concluded that the acetohydroxy acid synthase in Z. rouxii, unlike that in S. cerevisiae, was not sensitive to SMM. Furthermore, the enzyme assays demonstrated that the activity of threonine deaminase in Z. rouxii, like in S. cerevisiae, was strongly inhibited by isoleucine and stimulated by valine. From this work, it is clear that the aspartate-derived amino-acid metabolism in Z. rouxii only partly resembles that in S. cerevisiae. PMID- 10862916 TI - Highly enantioselective esterification of racemic ibuprofen in a packed bed reactor using immobilised Rhizomucor miehei lipase. AB - A systematic study of the enantioselective resolution of ibuprofen by commercial Rhizomucor miehei lipase (Lipozyme(R) IM20) has been carried out using isooctane as solvent and butanol as esterificating agent. The main variables controlling the process (temperature, ibuprofen concentration, ratio butanol:ibuprofen) have been studied using an orthogonal full factorial experimental design, in which the selected objective function was enantioselectivity. This strategy has resulted in a polynomial function that describes the process. By optimizing this function, optimal conditions for carrying out the esterification of racemic ibuprofen have been determined. Under these conditions, enantiomeric excess and total conversion values were 93.8% and 49.9%, respectively, and the enantioselectivity was 113 after 112 h of reaction. These conditions have been considered in the design of a continuous reactor to scale up the process. The esterification of ibuprofen was properly described by pseudo first-order kinetics. Thus, a packed bed reactor operating as a plug-flow reactor (PFR) is the most appropriate in terms of minimizing the residence time compared with a continuous stirred tank reactor (CSTR) to achieve the same final conversion. This reactor shows a similar behavior in terms of enantioselectivity, enantiomeric excess, and conversion when compared with batch reactors. A residence-time distribution (RTD) shows that the flow model is essentially a plug flow with a slight nonsymmetrical axial dispersion (Peclet number = 43), which was also corroborated by the model of CSTR in series. The stability of the system (up to 100 h) and the possibility of reutilization of the enzyme (up to four times) lead to consider this reactor as a suitable configuration for scale up of the process. PMID- 10862917 TI - Chemo-enzymatic synthesis and characterization of graft copolymers from lignin and acrylic compounds. AB - Chemo-enzymatic initiation of graft copolymerization of acrylic compounds onto different technical lignosulfonates (LS) was compared to a Fenton-like system (ferrous ion, t-BHP). The enzyme tested was a phenoloxidase laccase (EC 1.10.3.2) from the white rot basidomycete Trametes versicolor. Most applied lignins were successfully grafted, resulting in a polymer yield of more than 90%. The effect of initiator concentration and the lignin/monomer ratio on the yield and M(w) of enzymatically grafted polymers were studied. The homopolymer proportion in the enzymatically produced grafts of Ca-LS and acrylic acid was 5 to 6x lower than those initiated by the Fenton-like reagent; no such differences were observed for Na-LS. PMID- 10862919 TI - On the decision of outpatient adenoidectomy and adenotonsillectomy in children. AB - OBJECTIVE: Outpatient tonsillectomy and/or adenoidectomy is the procedure of choice in the US, whereas in Europe, the transition from the traditional duration of hospital stay to same day discharge slowly increases. This trial was conducted to find factors, which influence surgery on an outpatient basis and to find possible positive and negative predictive conditions in patients. METHODS: Most trials, which argue for a same day discharge take the low percentage of postoperative hemorrhage into account. Hemorrhage, apnea and infections are complications and have to be distinguished from sequelae such as poor oral intake (with consecutive i.v. fluid supply), fever and protracted vomiting, that also should be considered as discharge criteria. Complications as well as sequelae were measured in 114 consecutive children, and the patients divided into an adenotonsillectomy group and an adenoidectomy group. RESULTS: Patients from both groups that underwent surgery because of severe obstructive symptoms had significantly more sequelae than those indicated because of chronic or recurrent infections. They could not have been discharged in an acceptable condition. Due to the fact that most children after adenoidectomy recovered well 8 h postoperatively, they could have been discharged on the same day. Children after adenotonsillectomy had significantly more sequelae. There was a tendency that adenotonsillectomy children with only mild obstructions could have been discharged either 8 or at least 24 h postoperatively. It still remains the surgeon's decision when a child can be discharged safely. PMID- 10862918 TI - Tongue reduction in Beckwith-Weidemann syndrome. AB - OBJECTIVE: To review our experience with patients with macroglossia as a component of Beckwith-Weidemann Syndrome (BWS). DESIGN: Chart review of six patients treated with BWS. SETTING: Tertiary care teaching hospital. PATIENTS: Six patients diagnosed with BWS and macroglossia. INTERVENTIONS: Four patients underwent at least one surgical procedure to address their macroglossia. The surgical options and potential complications are discussed. RESULTS: Three patients who have undergone tongue reduction have a functioning tongue with normal mobility. Two patients have required tracheotomy as apart of their management and still have significant tongue enlargement. CONCLUSIONS: Macroglossia as a part of BWS may present a difficult management problem. Various methods of tongue reductions have been reported with mixed results. PMID- 10862920 TI - Organic change of effusion in the mastoid in otitis media with effusion and its relation to attic retraction. AB - To try to solve the pathogenesis of severe attic retraction viewed from mastoid condition, we examined the residual soft tissue density (RSTD) in the mastoid by computed tomography (CT) in 85 patients (107 ears) with otitis media with effusion (OME) 3 months after tympanostomy tube insertion or later. The incidence of RSTD in the mastoid was significantly higher in OME of adults (52.6%) than in children (24.1%). Ears with severe attic retraction had RSTD significantly more frequently (80%) than those with no or mild attic retraction, and many of the mastoids with severe attic retraction were occupied totally by RSTD. The area of the mastoid (mastoid pneumatization) was significantly smaller, and CT density of the mastoid (sclerotic tendency) was significantly higher in ears with RSTD than in those without. RSTD after tympanostomy tube insertion in the mastoid indicating organic change of effusion was considered one of the important factors relating to the pathogenesis of severe attic retraction. PMID- 10862921 TI - Evaluation of bcl-2 gene expression in papilloma of larynx in children. AB - Apoptosis - programmed death of a cell - is a natural mechanism that controls the number of cells in an organism. Neoplastic cells as many types of normal cells, may be the subject of spontaneous apoptosis as well as they may be induced by anti-neoplastic factors. Neoplastic cells' resistance to drugs is often correlated with impossible induction of apoptosis in those cells. Though the process of apoptosis is not fully explained, a possible involvement of many genes in regulation of this process is indicated. One of them is bcl-2 gene and its product - bcl-2 protein, which has the property of apoptosis process inhibition and stimulation of a cell towards outliving (survival). Increased expression of bcl-2 gene is present in many neoplastic cells and it suggests a possible pathogenic role of bcl-2 gene in oncogenesis. In this paper the expression of bcl 2 gene in the cells of papilloma in larynx is defined in six children operated in the Department of Paediatric Otolaryngology of Medical School in Lublin. Papillomas of larynx are neoplasm's of particular resistance to treatment. Complete, cellular RNA was isolated with Chomczynski and Sacchi method using guanidine thiocyanate. Gene expression was defined with the method of reverse transcription by cDNA synthesis and amplification of bcl-2 gene fragment with specific oligonucleotides in reverse transcriptase polymerase chain reaction (RT PCR). The products were identified on agarose gel. Expression of bcl-2 gene in the investigated cells of laryngeal papilloma was confirmed in all the children. The presence of bcl-2 gene product in these cells may be the cause of apoptosis inhibition and stimulation of cells proliferation of the neoplasm. PMID- 10862922 TI - Bilateral sensorineural hearing disorders in children: etiology of deafness and evaluation of hearing tests. AB - OBJECTIVE: The purpose of this study was to determine the etiology of bilateral sensorineural hearing disorders in children and to evaluate the performed hearing tests by comparison of the results of the objective and subjective tests. METHODS: The medical history and the hearing tests (behavioral observation audiometry, acoustic evoked potentials and pure tone audiometry) of 106 bilaterally hearing impaired children were analyzed in a retrospective follow-up study. RESULTS: The total group included 52 males and 54 females. The ages at first diagnosis ranged from 4 months to 11 years with a mean age of 42 months and a median of 33 months. The degree of hearing loss for the better hearing ear was mild in one child, moderate in 28 children, severe in 29 children, profound in 32 children and total in 16 children. The delay between the first examination and diagnosis ranged from 0 to 597 days with a mean of 83 days and a median of 28 days. In 47 children (44%) no cause of hearing impairment could be determined. Nineteen children (18%) had a history of familial hearing loss, 40 (38%) suffered from acquired hearing loss (seven children had prenatal causes, 21 perinatal and 12 postnatal). A comparison between behavioral observation audiometry and brainstem evoked response audiometry revealed a statistically good agreement. Twenty-nine children (32%) showed progressive hearing loss, which was defined as a threshold shift of +10 dB or more in the pure tone average in at least one ear. CONCLUSIONS: In a significant number of children with early hearing impairments the etiology still remains uncertain. Further research in the field of genetic disorders will diminish this number. Evaluation of hearing tests showed that behavioral observation audiometry still is an excellent tool in the hands of an experienced examiner. The age at identification of hearing disorders in industrialized countries still is unacceptably high. To obtain ideal care of hearing impaired children, universal neonatal hearing screening programs are mandatory. PMID- 10862923 TI - Otolaryngologic aspects of oral-facial-digital syndrome. AB - The oral-facial-digital (OFD) syndromes are a heterogeneous group of hereditary disorders which have in common the findings of oral abnormalities, facial dysmorphism, and hand/feet malformations. We report the case history of an 18 month-old male with cerebellar cysts, hydrocephalus, tongue hamartomas, and polydactyly. These findings are most consistent with OFD VI. The clinical features of eight different types of OFD are discussed, with particular attention to the characteristics of the most interest to the otolaryngologist. PMID- 10862924 TI - Fractured tracheostomy tubes in the tracheobronchial tree of a child. AB - Fractured tracheostomy tube presenting as foreign bodies in the tracheobronchial tree is rare. Only four previous episodes in children have been reported in literature and most of these in developing countries. We report an unusual case of fractured tracheostomy tubes in the tracheobronchial tree of a child and review the literature. PMID- 10862925 TI - Totally obstructing tracheotomy-associated suprastomal granulation tissue. AB - Although tracheotomy-associated suprastomal granulation tissue is quite common, suprastomal granulation tissue that totally obstructs the airway is relatively rare and can be associated with serious complications. In this report the complications and management of six cases of totally obstructing suprastomal granulation tissue (TOSGT) are presented. Complications associated with the presence or management of TOSGT included progression of subglottic stenosis, development of posterior laryngeal stenosis, development of supraglottic stenosis following CO(2) laser supraglottoplasty, and dislodgement of the TOSGT with distal tracheal obstruction resulting in anoxic brain injury. It is recommended that the tracheotomy tube remains in position at all times during attempted removal, and that if endoscopic removal is not possible, that open tracheoplasty is the safest method for removal. Measures that may decrease the chances of recurrence include diligent diagnosis and treatment of gastroesophageal reflux disease (GERD) and bacterial infection. TOSGT may be a marker for some patients with abnormal wound healing. PMID- 10862926 TI - Chloroma of the masseteric muscle. AB - Chloroma (leukemic infiltrate or granulocytic sarcoma) is a localized extramedullary mass of immature granulocytic cells. They are uncommon tumors that usually occur in patients with leukemia, mostly of the myeloid type. Involvement in the head and neck region is rare. Granulocytic sarcomas of the face, maxilla, paranasal sinuses, temporal bone, and pharynx have all been documented in the past. We present the first reported case of a granulocytic sarcoma involving the masseteric muscle in an 8-month-old white male diagnosed with acute myeloid leukemia (AML). The lesion resolved with chemotherapy but the patient subsequently died. This case reaffirms the importance of including chloroma in the differential diagnosis of lesions in patients with AML and the prognostic value they hold. PMID- 10862927 TI - Lingual tonsillectomy for refractory paroxysmal cough. AB - Historically, the lingual tonsils are the most neglected members of Waldeyer's ring. They are often overlooked even in a thorough head and neck exam because of their anatomic location and the ambiguous constellation of symptoms which they produce when they are diseased or enlarged. The lingual tonsils have been reported to be associated with a variety of upper aerodigestive tract symptoms including odynophagia, dysphagia, otalgia, globus, halitosis, chronic cough, and dyspnea. Many patients with lingual tonsillar pathology may undergo extensive work-up for some of these non-specific upper airway complaints by their primary physician before referral to an otolaryngologist. Consequently, the diagnosis of lingual tonsillar disease requires a high index of suspicion and a thorough physical exam including evaluation of the tongue base and hypophaynx with indirect mirror or fiberoptic exam. In order to draw attention to this frequently unrecognized entity, we present a case report of a child with chronic cough resulting from lingual tonsillar hypertrophy. PMID- 10862928 TI - Suspected foreign body aspiration in a child with endobronchial tuberculosis. AB - Endobronchial tuberculosis is a form of pulmonary tuberculosis, thought to result from rupture of an infected node through the bronchial wall or from lymphatic spread to the mucosal surface of the bronchial tree. With the presence of multidrug resistant isolates of TB, and its incidence in an increasing number of foreign-born persons immigrating to the US, otolaryngologists must be aware of its often subtle presentation. The following case is an unusual presentation of endobronchial tuberculosis initially diagnosed as an airway foreign body. PMID- 10862929 TI - Neonatal submandibular sialadenitis progressing to submandibular gland abscess. AB - Submandibular sialadenitis is exceptionally rare in neonates. We describe a case of submandibular sialadenitis progressing to submandibular abscess in a term neonate. The aetiology, investigations and treatment for this very rare condition are discussed. PMID- 10862930 TI - Electrophysiological correlates of category goodness. AB - We report the results obtained from a behavioural and electrophysiological study. A synthesised continuum going from labial /ba/ to retroflex /da/ through dental /da/ was tested for category goodness. Native English speakers rated different tokens from each category as good, bad or ambiguous. The results showed that not all of the representatives of each category were ideal and that the categories tested have an internal structure. The electrophysiological study evaluated whether event related potentials (ERPs) mirrored the goodness judgements. During a passive oddball task, the same participants were exposed to native /ba/-/da/, Hindi dental /da/-retroflex /da/ and within-category /ba/-/ba/ contrasts. Results showed that participants pre-attentively perceive the differences in all cases, as shown by mis-match negativities (MMN), late positive deflections (LPD) or greater N1 and/or P2 components for deviant stimuli. Acoustic sensitivities, categorical perception and category goodness all contributed to the waveforms obtained. We attribute the ERP effects to a combination of (1) prototypes built from initial sensitivities, (2) reinforcement with exposure to one's native language and (3) no permanent loss of the initial boundaries explains the effects observed. PMID- 10862931 TI - Behavioral and mesocorticolimbic dopamine responses to non aggressive social interactions depend on previous social experiences and on the opponent's sex. AB - In these experiments we evaluated the relationship between behavioral and brain dopamine (DA) responses to social interactions. Subjects were group housed male mice confronted with a non aggressive male or female conspecific following either repeated defeat (defeated) or repeated non aggressive experiences (social). Defeated mice showed more defensive/submissive reactions then mice of the social group regardless of the opponent sex. However, mice defeated by females showed reduced social exploration without significant differences in non social exploration whilst the opposite was true for mice defeated by male opponents. Non aggressive social interactions enhanced dopamine metabolism in the prefrontal cortex (pFC) of DEFEATED mice regardless of opponent sex. However, only mice defeated by females showed enhanced dopamine metabolism and release in the nucleus accumbens septi (NAS) and olfactory tubercle (OT) following interaction with the non aggressive opponent. Finally, correlation between central and behavioral responses evidenced that 3,4-dihydroxiphenilacetic acid levels in the pFC were positively correlated with defensive behaviors and negatively correlated with non social exploration in mice confronted with male opponents but not in those confronted with females. The latter, showed a significant positive correlation between 3-methoxytyramine (3-MT) levels in the OT and defensive responses and significant negative correlation between social investigation and 3 MT levels in the OT and in the NAS. These results indicate a strict relationship between mesocorticolimbic dopamine transmission and behavior responses to social cues. Moreover, they strongly support the view that mesocorticolimbic DA modulates social behavior by affecting perceptive processing. PMID- 10862932 TI - Post-trial sleep sequences including transition sleep are involved in avoidance learning of adult rats. AB - High resolution computerized EEG analyses, and behavioral observations were used to identify slow wave sleep (SS), paradoxical sleep (PS) and transition sleep (TS) in adult male Wistar rats exposed to a session of two-way active avoidance training. Of the four sleep sequences that could be identified, two included TS (SS-->TS-->W and SS-->TS-->PS), while the other two did not (SS-->W and SS-->PS). Comparison of post-trial sleep variables between fast learning rats (FL, reaching criterion in the training session), slow learning rats (SL, reaching criterion in the retention session the following day), and non learning rats (NL, failing to reach criterion) indicated that the total amounts of SS, TS and PS of the SS-->TS ->PS sequence was markedly higher in FL rats than in SL rats. In addition, in comparison with the corresponding baseline period, the average duration and total amount of SS and TS episodes of the SS-->TS-->PS sequence increased in FL rats, while the number of SS-->TS-->W sequences decreased. On the other hand, the average duration of SS episodes increased in the SS-->TS-->W and SS-->W sequences of SL rats, and in the SS-->W and SS-->TS-->PS sequences of NL rats. Correlative analyses between number of avoidances and post-trial sleep variables demonstrated that avoidances were directly correlated with the duration of SS episodes of the SS-->TS-->PS sequence and with the duration of TS episodes of the SS-->TS-->W sequence, but inversely correlated with the number and amount of SS episodes of the SS-->W sequence and with the duration and amount of SS episodes of the SS- >PS sequence. On the whole, the data supported the view that TS-containing sleep sequences are involved in long-term storage of novel adaptive behavior, while sleep sequences lacking TS are involved in the maintenance of innate behavioral responses. PMID- 10862933 TI - Impaired escape performance and enhanced conditioned fear in rats following exposure to an uncontrollable stressor are mediated by glutamate and nitric oxide in the dorsal raphe nucleus. AB - Exposure to uncontrollable aversive events produces a variety of behavioral consequences that do not occur if the aversive event is controllable. Accumulating evidence suggests that exaggerated excitation of serotonin (5-HT) neurons in the dorsal raphe nucleus (DRN) is sufficient to cause these same behaviors, such as poor shuttlebox escape performance and enhanced conditioned fear that occur 24 h after exposure to inescapable tailshock (IS). The aim of the present studies was to explore the possibility that N-methyl-D-aspartate (NMDA) receptor activation and nitric oxide (NO) formation within the DRN might be involved in mediating the behavioral consequences of IS. To this end, either the NMDA receptor antagonist 2-amino-5-phosphonovaleric acid (APV) or the nitric oxide synthase inhibitor Nw-nitro-L-arginine methyl ester (L-NAME), was microinjected into the DRN before IS or before testing 24 h later. Blocking NMDA receptors with APV in the DRN during IS prevented the usual impact of IS on escape responding and conditioned fear. However, injection of APV at the time of testing only reduced these effects. The DRN was shown to be the critical site mediating blockade of these behavioral changes since injection of APV lateral to the DRN did not alter the behavioral consequences of IS. Conversely, L-NAME was most effective in reversing the effects of IS when administered at the time of testing. These results suggest that there is glutamatergic input to the DRN at the time of IS that produces long-lasting changes in DRN sensitivity. This plasticity in the DRN is discussed as a possible mechanism by which IS leads to changes in escape performance and conditioned fear responding. PMID- 10862934 TI - A cautionary note regarding drug and brain lesion studies that use swimming pool tasks: partial reinforcement impairs acquisition of place learning in a swimming pool but not on dry land. AB - Spatial tasks are used widely in neurobiological studies because it is thought that they provide an unbiased assessment of the integrity of neural structures that mediate spatial learning. For example, in the Morris swimming pool place task, animals are required to locate a hidden platform in a swimming pool in relation to environmental cues. Treatments that result in an animal's failure to find the platform are assumed to reflect defects in the function of neural systems involved in spatial learning. The present study demonstrates, however, that an animal's reinforcement history can contribute to its spatial performance. Animals were trained in the Morris place task with the platform present on 100, 75 or 50% of trials. Relative to the 100% group, the 75% group was impaired in place acquisition, and the 50% group failed to learn. Even placing the 50% group animals onto the platform at the completion of an unsuccessful trial failed to improve acquisition. Animals trained to search for food on an identical dry maze problem were not affected by similar reinforcement schedules. The present findings demonstrate that the Morris swimming pool place task does not provide an unbiased assessment of spatial learning: A treatment effect may be confounded with reinforcement history. The results are discussed in relation to widespread applications of the Morris place task to neurobiological problems. PMID- 10862935 TI - Spatial information transfer from virtual to real versions of the Kiel locomotor maze. AB - The Kiel locomotor maze requires participants to choose five targets from among 20 locations marked by small red lights on the floor of a dimly lit circular environment having four wall-mounted extramaze cues and two intramaze cues at floor level. In the present study, acquisition of the real task was examined in 11-year-old children following prior accurate training in a virtual version, following misleading virtual training, or following no training. The virtual version was displayed on a desk-top computer monitor. Acquisition testing in the real maze was either locomotor or non-locomotor. Good transfer was achieved from virtual to real versions. Children's exploration of the real maze prior to real maze acquisition training revealed a clear transfer of spatial information previously learned in the virtual version. Children taught the correct target configuration in the simulation made fewer errors and more rapid, confident responses to targets in the real maze than children given no training. However, acquisition was also better following misleading training than no training, suggesting that a non-specific components of performance also transferred. Male superiority was only seen following misleading training, which was interpreted in terms of male superiority in mental rotation. After acquisition, a single probe trial was performed, in which proximal cues and participants' starting position were rotated, but this had equivalent effects on all groups' performance. It is clear that transfer of spatial information occurs from the simulated Kiel maze to the real version. This has implications for its use in diagnosis and training. PMID- 10862936 TI - Nigrostriatal dopamine system and motor lateralization. AB - The mechanism by which most people favor use of the right hand remains unknown. It has been proposed that asymmetries in the nigrostriatal dopamine system may be related to motor lateralization in humans. We explored this hypothesis in vivo by using [18F]fluorodopa positron emission tomography. Whereas the degree of right hand preference was found to correlate with left putamen dominance as assessed by asymmetry in fluorodopa uptake (K(i)), right caudate dominance was positively correlated with the level of performance during simultaneous bimanual movements in right-handed normal subjects. In addition, right-handed patients with Parkinson's disease with higher right than left caudate K(i) performed much better in bimanual movement tests than those in whom the K(i) value of the left caudate was higher than that of the right. These findings support the notion that the nigrostriatal dopaminergic system may play a role in motor lateralization, and suggest a functional model for bimanual movements. We propose that the skill for performing simultaneous bilateral hand movements in right-handed subjects might depend upon both the activation (through the dominant left putamen circuitry) of the left supplementary motor area (SMA), and the inhibition (through the right caudate circuitry) of motor programs stored in the right SMA. PMID- 10862937 TI - Lesions of the perirhinal cortex impair sensory preconditioning in rats. AB - The effects of lesions of the perirhinal cortex on the development of associations between two conditioned stimuli (CSs) were examined with a sensory preconditioning procedure. Rats were given either bilateral electrolytic lesions of the perirhinal cortex or control surgery. They were then given either paired or unpaired presentations of a light CS and a tone CS. All of the rats were then given eyeblink conditioning procedures that involved paired presentations of either the light or tone and a periorbital shock unconditioned stimulus (US). The rats were finally given a test session that consisted of unpaired presentations of the tone and light CSs. Sensory preconditioning was established in the control group, but not in the lesion group. The findings are consistent with the view that the perirhinal cortex is involved in forming associations between neutral stimuli (even in the absence of reinforcement). PMID- 10862938 TI - Interstrain differences in cognitive functions in rats in relation to status epilepticus. AB - Cognitive functions of Long Evans (N=30) and Wistar rats (N=32) were compared using a Morris water maze. Under control conditions the Long Evans rats were more efficient in this test, their average escape latency after 5 days of training (6.4+/-0.1 s, mean+/-S.E.M.) was significantly shorter than that of the Wistar rats (11.0+/-0.1 s). When the training was completed seizures were induced by an intraperitoneal injection of pilocarpine (330 mg/kg in the Long Evans strain and 350 mg/kg in the Wistar rats) 30 min after pretreatment with N-methylscopolamine (1 mg/kg i.p.). Clonazepam (1 mg/kg i.p.) was used to interrupt clonic seizures after 2 hours of continuous activity. Approximately one quarter of rats in both strains did not develop seizures. Severe convulsive status epilepticus was common in Long Evans rats (23 out of 30). In contrast, only 12 Wistar rats generated convulsive status epilepticus and the same number of animals exhibited only bursts of motor seizures separated by periods without convulsions (temporary seizures). Mortality after pilocarpine-induced status epilepticus was considerably higher in the Long Evans rats than in the Wistar rats. After a latency of 2-3 weeks spontaneous recurrent seizures appeared in all animals surviving status. Cognitive memory was tested during the 'silent period' between status and recurrent seizures. The Long Evans rats were unable to find the platform at the 3rd and 6th day after status but then their performance rapidly improved. The performance of the Wistar rats undergoing status epilepticus was seriously deteriorated and it never normalized, whereas the animals with temporary seizures exhibited only a transitory marginal prolongation of latencies. The hippocampal formation was damaged by status epilepticus in rats of both strains - the Long Evans rats exhibited more extensive damage of subfields CA1 and CA3, whereas in the Wistar rats a complete destruction of hilar neurons was observed in addition to partial CA1 and CA3 damage. PMID- 10862939 TI - Behavioral effects of protein deprivation and rehabilitation in adult rats: relevance to morphological alterations in the hippocampal formation. AB - In the present study we have analyzed the behavioral and neuroanatomical effects of protein deprivation in adult rats. Starting at 2 months of age, animals were maintained on 8%-casein diet either for 8 months (malnourished group), or for 6 months followed by a 2-month period of nutritional rehabilitation (17%-protein diet, rehabilitated group). Malnourished rats exhibited reduced emotional reactivity and impaired habituation in the open field. In a water maze, these animals did not differ from controls during training, but showed retention deficits on the probe trial. However, working memory, sensorimotor abilities and passive avoidance behavior were not significantly impaired in malnourished rats. The performance of rehabilitated group was similar to that of the control group throughout behavioral testing. Postmortem morphological analysis revealed that the total number of neurons in the granular layer of the dentate gyrus, and in CA3 and CA1 hippocampal fields was reduced in protein-deprived and rehabilitated rats relative to controls. In addition, it was found that protein deprivation caused a 30% loss of synapses established between mossy fibers and dendrites of CA3 pyramidal cells, whereas nutritional rehabilitation resulted in a reversal of this effect. These results show that prolonged malnutrition in adult rats produces marked loss of hippocampal neurons and synapses accompanied by substantial impairments of hippocampal-dependent behaviors. The fact that nutritional rehabilitation results in restoration of the total number of hippocampal synapses and parallel amelioration of the behavioral impairments suggests that the mature CNS possesses a remarkable potential for structural and functional recovery from the damage induced by this type of dietary insult. PMID- 10862940 TI - NMDA receptor antagonism in the basolateral amygdala blocks enhancement of inhibitory avoidance learning in previously trained rats. AB - Extensive evidence suggests that N-methyl-D-aspartate (NMDA) glutamate receptor channels in the amygdala are involved in fear-motivated learning, and infusion of NMDA receptor antagonists into the amygdala blocks memory of fear-motivated tasks. Recent studies have shown that previous training can prevent the amnestic effects of NMDA receptor antagonists on spatial learning. In the present study, we evaluated whether infusion of the NMDA antagonist D,L-2-amino-5 phosphonopentanoic acid (AP5) into the basolateral nucleus of the amygdala (BLA) impairs reinforcement of inhibitory avoidance learning in rats given previous training. Adult male Wistar rats (220-310 g) were bilaterally implanted under thionembutal anesthesia (30 mg/kg, i.p.) with 9.0-mm guide cannulae aimed 1.0 mm above the BLA. Infusion of AP5 (5.0 microg) 10 min prior to training in a step down inhibitory avoidance task (0.4 mA footshock) blocked retention measured 24 h after training. When infused 10 min prior to a second training session in animals given previous training (0.2 mA footshock), AP5 blocked the enhancement of retention induced by the second training. Control experiments showed that the effects were not due to alterations in motor activity or footshock sensitivity. The results suggest that NMDA receptors in the basolateral amygdala are involved in both formation of memory for inhibitory avoidance and enhancement of retention in rats given previous training. PMID- 10862941 TI - Effects of early midline cerebellar lesion on cognitive and emotional functions in the rat. AB - Midline lesion of the cerebellum was performed in young 10-day-old DA/HAN strained (pigmented) rats. Once adults, the lesioned animals were subjected to a series of behavioral tests and their performances were compared with those of control (nonlesioned) rats. Compared with controls, the spontaneous motor activity of the lesioned rats was higher, they showed persevering behavior and did not pay attention to environmental distractors. In anxiety and social discrimination tests, disinhibition tendencies were obvious, which suggested that the animals were less dependent on the context. These abnormalities were most likely due to early midline lesion of the cerebellum and not to a deficit in maternal care before weaning, since the dams took care of the lesioned and control pups similarly. From these results, it can be concluded that the cerebellar vermis is involved in motor control, attentional capabilities and emotional behavior. Given that the lesioned rats observed in this study presented obvious autistic-like symptoms, and since a number of autistic subjects have cerebellar deficits and, particularly, a hypoplasia of vermal lobules, our results may strengthen the idea that the cerebellar vermis is involved in autism, as already suggested in the guinea pig (Caston J, et al. Eur J Neurosci 1998;10:2677-2684). PMID- 10862942 TI - Eye use in search is determined by the nature of task in the domestic chick (Gallus gallus). AB - In Experiments 1-3 chicks were trained to find, using both eyes, food covered by a cap, using wide-angle search so as to involve lateral and frontal visual fields, with either local or positional cues, or both, identifying the baited site. At test they used right, left or both eyes (RE, LE, Bin). When both types of cue were relevant, LE made greater use of positional cues than the other two groups, as has been previously found, whereas RE made greater use of local (colour) cues. However, when only one type of cue was relevant, RE and LE were equally able to use positional or local cues. Right/left differences emerge when RE and LE can be used in different ways during training. In Experiments 4-5 Bin chicks were shown to turn preferentially to the right during wide-angle search, when relying on local, and to the left when relying on positional cues. In search, parallel processing of RE and LE inputs appears to allow competition which is usually won by the eye system more suited to the task, which then initiates targeting to objects which are visible to its eye. PMID- 10862943 TI - Sucrose and quinine intake by maternally-deprived and control rhesus monkeys. AB - Clinical depression is often characterized by a loss of interest or pleasure in formerly enjoyable activities. Analogs of anhedonia are established in rats, but the generality of this phenomenon to other species is unknown. Maternally deprived rhesus macaques show a wide range of behavioral abnormalities that are reversed by chronic antidepressant treatment. We tested consumption by maternally deprived versus control macaques of sweetened (seven sucrose concentrations) or bitter water (four quinine concentrations) versus plain water to evaluate a non human primate model of depression for signs of anhedonia. All monkeys consumed more sweetened than tap water, but maternally-deprived monkeys had a diminished preference for sweetened water than did controls. However, maternally deprived animals consumed more bitter water than did controls. Baseline fluid consumption did not differ. The data suggest that 'anhedonia' in animal models may be secondary to a generally attenuated responsiveness to stimuli, rather than a unitary reduction in responsiveness to the appetitive properties of stimuli. We conclude that maternally-deprived rhesus monkeys do not display gustatory signs of anhedonia, but rather of insensitivity to gustatory stimuli. PMID- 10862944 TI - Spatial ability of XY sex-reversed female mice. AB - Perinatal gonadal hormones significantly affect subsequent sex differences in reproductive and non-reproductive behaviors in rodents. However, the influence of the sex chromosomes on these behaviors has been largely ignored. To assess the influence of the non-pseudoautosomal region of the Y chromosome, C57BL/JEi male and female mice and mice from the C57BL/6JEi-Y(POS) consomic strain were given behavioral tests known to distinguish males from females. The C57BL/6JEi-Y(POS) strain contains sex-reversed XY-females which, when compared to their XX-female siblings, allow assessment of the influence of the Y chromosome in a female phenotype. XX-females and XY-females did not differ on open-field activity, the Lashley maze, or active avoidance learning, but XY-females were significantly better than XX-females on the Morris hidden platform spatial maze. These findings suggest that males may have both a genetic and a hormonal mechanism to ensure visuospatial superiority. PMID- 10862945 TI - Median raphe nucleus mediates forming long-term but not short-term contextual fear conditioning in rats. AB - The brain serotonin is involved in mediation of emotional behaviour including anxiety and related fear conditioning. It is known that the median raphe nucleus (MRN) is the origin of a serotonergic pathway and mainly innervates septo hippocampal formation which plays an important role in emotional cognition. However, its regulatory role in different types of fear conditioning is still unclear. In the present study, the animals underwent ibotenic acid or sham lesions of the median raphe nucleus and the effects of MRN lesions on immediate and delayed fear conditioning to multiple contextual cues were studied. Freezing behaviour served as a measure of contextual fear. Sham-lesioned animals showed reliable conditional freezing when observed immediately following foot-shock (1.0 mA) for 3-min test and 48 h after the shock for 12-min test. Rats with MRN lesions displayed robust freezing behaviour immediately after the shock, even though they showed a marked deficit in freezing 48 h following the shock. These findings indicate that the MRN-serotonergic septo-hippocampal pathway is involved in the regulation of anxiety related to fear conditioning triggered by contextual cues, suggesting that short-term contextual fear is independent on the MRN while long-term contextual fear depends on the MRN. PMID- 10862946 TI - Adrenergic activation of the nucleus tractus solitarius potentiates amygdala norepinephrine release and enhances retention performance in emotionally arousing and spatial memory tasks. AB - It is well documented that noradrenergic systems in the amygdala modulate memory formation, however, less research has examined how sources of limbic norepinephrine contribute to this process. The amygdala receives a dense supply of norepinephrine from neurons in the nucleus of the solitary tract (NTS). The present experiments examined whether adrenergic activation of these NTS neurons affects memory in learning tasks that are sensitive to amygdala norepinephrine release. Separate groups of male Sprague-Dawley rats were trained in either an emotionally arousing or spatial memory task. They then received vehicle or the adrenergic agonist epinephrine (50, 125, or 250 ng/0.5 microl) into the NTS. Rats given the 125 ng dose had significantly longer retention latencies on a 48 h inhibitory avoidance retention test and made a significantly higher percentage of correct responses on an 18 h delayed radial maze retention test. A third experiment using in vivo microdialysis and high performance liquid chromatography (HPLC) demonstrated that intra-NTS infusion of a memory-enhancing dose of epinephrine potentiated amygdala norepinephrine release. Collectively, these results suggest that stimulation of the NTS contributes to memory processing by influencing noradrenergic systems in the amygdala. PMID- 10862947 TI - Morning recall of verbal material depends on prior sleep organization. AB - Despite the evidence that sleep may facilitate memory, controversial findings concern the role of sleep states (NREM, REM). We put forward the hypothesis that sleep organization, i.e. the regular occurrence of NREM-REM cycles, more than sleep states per se, may be crucial for the retention of verbal material presented before sleep. An experiment was performed in which recall of verbal material was requested of young subjects after three different kinds of night sleep: undisturbed sleep, fragmented sleep without sleep cycles disorganization, and fragmented sleep interrupted with sleep disorganization. Morning recall of verbal material was impaired after the night with disturbed sleep cycles, whereas it was not after the night with preserved sleep cycles; the amount of REM was similar in both cases. We conclude that the recall of verbal material is greatly affected by sleep cycle disorganization. PMID- 10862948 TI - Learning performances, brain NGF distribution and NPY levels in transgenic mice expressing TNF-alpha. AB - Tumor necrosis factor-alpha (TNF-alpha) is a cytokine involved in a variety of neurobiological activities including changing behavior and regulation of both neurotrophin and neuropeptide levels. In this study we used two lines of transgenic mice overexpressing brain TNF-alpha characterized by neurological deficits (line Tg6074) or phenotypically normal (line TgK3). We analyzed whether or not impairments in learning and memory processes due to TNF-alpha overexpression were associated with changes in endogenous brain NGF, NPY and beta amyloid. The results indicate that full TNF-alpha transgene expression disrupted the learning capabilities of transgenic mice (both Tg6074 and TgK3). NGF decreased in the hippocampus of both transgenic mice whereas hippocampal NPY slightly potentiated in Tg6074. The decrease in NGF is correlated with deficits in spatial learning and memory whereas inflammation in the brain of Tg6074 could be responsible of the hippocampal increase in NPY. As a whole, these results show that transgenic mice overexpressing TNF-alpha in the brain represent a useful model for studying neuronal degeneration and brain inflammatory processes. PMID- 10862949 TI - Phenacetin, acetaminophen and dipyrone: analgesic and rewarding effects. AB - The antinociceptive and rewarding effects of phenacetin, a mild analgesic with abuse liability, were compared with those of acetaminophen, dipyrone and indomethacin in the formalin and conditioned place preference tests. Phenacetin, acetaminophen and dipyrone attenuated the pain response, beginning at 50, 50 and 100 mg/kg, respectively. Systemically active drugs produced weaker antinociception when injected into the paw or intracerebroventricularly at doses approximately 10(3) lower than those required for systemic effects; dose effect relations were bell-shaped by the intracerebroventricular route. By all three routes, there was a clear ceiling to the effects of acetaminophen that is consistent with its clinical efficacy. Systemic phenacetin and acetaminophen produced a conditioned place preference at doses that produced antinociception. Dipyrone produced a place aversion by the systemic route and a place preference intracerebroventricularly. The latter had a bell-shaped dose effect relationship identical to that in the formalin test. Indomethacin was inactive in all tests, except for mild hyperalgesia by the intraventricular route. The results indicate that the antinociceptive effects of phenacetin, acetaminophen and dipyrone reflect a combination of peripheral and central actions, neither of which involves inhibition of cyclooxygenase. The central component of the antinociceptive effects may be related to activation of brain mechanisms that are involved in reinforcement. PMID- 10862950 TI - Retinal ganglion cells regenerating through the peripheral nerve graft retain their electroretinographic responses and mediate light-induced behavior. AB - To assess the light-induced electrical activity of rodent retinal ganglion cells (RGCs) regenerating into a peripheral nerve (PN) graft we used non-invasive recording of electroretinographic responses to the contrast-reversal of sinusoidal gratings (p-ERG). On comparing the retinas that received a PN graft and retinas with only optic nerve (ON) transection, p-ERG responses were present in grafted retinas as late as 20 months after the surgery while they completely disappeared in non-transplanted controls within 4 months of ON transection. Next, the ability of regenerating RGCs to form functional connections with their targets in the superior colliculus (SC) was tested by a light-escape task. While the bilaterally blinded animals did not improve during the test, unilaterally grafted animals (with the contralateral eye blinded) reached 26% success in the last quartile of the light-escape task. This performance was significantly better than that of blind animals (ANOVA and Student-Newman-Keuls test; p<0.05), but did not reach the level of intact rats (87%). The transplanted rats, therefore, were capable of light perception, but at a sub-normal ability. In addition, we were also able to correlate the amplitude of the p-ERG response with the visual behavioral performance for each transplanted animal. This finding indicates that there is a direct link between the RGC electrophysiological activity and the functional capacity of the regenerated visual pathway. In conclusion, the above results indicate that (a) PN grafts help to preserve the normal electroretinographic activity of injured and regenerating RGCs (b) the regenerated visual pathway is functional and capable of mediating simple visual behavior and that (c) there is a correlation between the light-evoked RGC electrical activity and visual behavior and, finally, that (d) the effect of PN graft on the electrophysiological and functional restoration of the visual pathway is long-lasting or even permanent. PMID- 10862953 TI - Stabilizing effects of caprylate and acetyltryptophanate on heat-induced aggregation of bovine serum albumin. AB - Acetyltryptophanate (AT) and caprylate (Cap) have been used to stabilize serum albumin against heat treatment. However, the mechanism of stabilization by these additives has never been fully elucidated. Here we used thermal melting to determine the effects of these additives on the melting temperature of bovine serum albumin (BSA) and heat stress at 60 degrees C to follow degradation of the protein in the presence of varying concentrations of AT or Cap. Native polyacrylamide gel electrophoresis was used to examine degradation products generated by heat treatment. Both additives increased the melting temperature of BSA, resulting in an increase by 12 degrees C at 5 mM AT and 3 degrees C at 1 mM Cap. They also conferred stability to BSA against heat stress at 60 degrees C. Complete protection was observed at 5 mM AT and 1 mM Cap. Comparison of AT and Cap in their effects on melting temperature and heat stress-induced degradation showed that a greater protection occurs with Cap which has a weaker effect on melting temperature. Based on this observation it was concluded that the observed protection by AT may be explained by its effects on melting temperature while that of Cap should be ascribed to other mechanisms. PMID- 10862957 TI - Single disulfide bond reduced papain exists in a compact intermediate state. AB - Partially reduced proteins and other chemically modified derivatives are very useful model systems to understand the protein folding in vivo. Upon reduction, proteins attain different conformations with varying degrees of compactness. The reduction of papain in the presence of 8 M urea leads to the partial reduction of one disulfide bond. This derivative (single disulfide reduced carboxymethylated 1RCM papain (3RCM papain)) was characterized by spectroscopic methods and the effect of this reduction on the unfolding of the protein was investigated. Under this partial reduction, papain exhibits more than half of the tertiary and most of the secondary structures relative to the non-reduced molecule (free cysteine reduced and carboxymethylated papain (1RCM papain)). Hydrophobic regions are exposed to the solvent as observed through 8-anilino-1-naphthalene sulfonic acid binding which was absent in the fully intact and unfolded protein, at neutral pH. Hydrodynamic studies indicated that 3RCM papain, under neutral conditions, possess expanded conformation as compared to the native protein. Tryptophan fluorescence quenching studies suggested the exposure of aromatic residues to solvent. Guanidine hydrochloride induced unfolding of this derivative, at neutral pH, showed a non-cooperative transition contrary to the cooperativity seen with intact protein. Thermal unfolding indicates that 3RCM papain is less stable compared to the intact protein. These findings suggest that partial reduction of papain has a significant effect on the unfolding behavior of papain. PMID- 10862962 TI - Interactions between subdomains in the partially folded state of staphylococcal nuclease. AB - Staphylococcal nuclease can be roughly divided into a beta-subdomain in N terminal and an alpha-subdomain in C-terminal. They fold sequentially under certain conditions, causing a partially folded intermediate state in which the native-like beta-barrel persists while alpha-helix regions largely disorder. To investigate the possible long-range interactions between the two subdomains in the intermediate, N-terminal fragments have been used as intermediate analogues, with polypeptide ending at positions 102, 110, 121 and 135 and with a tryptophan substitution at position 66 or 88 to facilitate the observation of the beta barrel. Segment-resolved interactions between beta-barrel and residues 103-135 were identified by comparing their spectroscopic properties of fluorescence, circular dichroism and NMR and by their stability. Except for unstable V66W102, the guanidine and thermal denaturation of fragments are cooperative and well approximated by the two-state transition. Minimal stable structure units of both tryptophan-containing fragments comprise residues 1-110. With the main interaction in segment 103-135, residues 103-110 contribute approximate 2 kcal/mol to the stability. Elongation of C-terminal from 110 residue neither increases the stability nor alters the structure core of the G88W fragments. However, residues 111-121 influence the tertiary structure of the V66W fragments suggesting its minor interactions with beta-barrel. PMID- 10862968 TI - The role of amino acid residues in the active site of a midgut microvillar aminopeptidase from the beetle Tenebrio molitor. AB - Aminopeptidases are major enzymes in the midgut microvillar membranes of most insects and are targets of insecticidal Bacillus thuringiensis crystal delta endotoxins. Sequence analysis and substrate specificity studies showed that these enzymes resemble mammalian aminopeptidase N, although information on the organization of their active site is lacking. The effect of pH at different temperatures on the kinetic parameters of Tenebrio molitor (Coleoptera) larval aminopeptidase showed that enzyme catalysis depend on a deprotonated (pK 7.6; DeltaH degrees (ion), 7.6 kJ/mol) and a protonated (pK 8.2; DeltaH degrees (ion), 16.8 kJ/mol) group. 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide and diethylpyrocarbonate inactivate the enzyme by modifying a pK 5.8 carboxylate and a imidazole group, respectively, with a reaction order around 1. Tetranitromethane changes the K(m) of the enzyme without affecting its V(max) by modifying a phenol group. The presence of a competitive inhibitor decrease the inactivation reaction rates in all these cases. EDTA inactivation of the aminopeptidase is affected by pH and temperature suggesting the involvement in metal binding of at least one deprotonated imidazole group (pK 5.8, DeltaH degrees (ion), 20 kJ/mol). The data support the hypothesis that T. molitor aminopeptidase catalysis depends on a catalytic metal and on a carboxylate and a protonated imidazole group, whereas substrate binding relies in one phenol and one carboxylate groups. The insect aminopeptidase shares common features with mammalian aminopeptidase N, although differing in details of substrate binding and in residues directly involved in catalysis. PMID- 10862969 TI - Major cold shock proteins, CspA from Escherichia coli and CspB from Bacillus subtilis, interact differently with single-stranded DNA templates. AB - The family of bacterial major cold shock proteins is characterized by a conserved sequence of 65-75 amino acid residues long which form a three-dimensional structure consisting of five beta-sheets arranged into a beta-barrel topology. CspA from Escherichia coli and CspB from Bacillus subtilis are typical representative members of this class of proteins. The exact biological role of these proteins is still unclear; however, they have been implicated to possess ssDNA-binding activity. In this paper, we report the results of a comparative quantitative analysis of ssDNA-binding activity of CspA and CspB. We show that in spite of high homology on the level of primary structure and very similar three dimensional structures, CspA and CspB have different ssDNA-binding properties. Both proteins preferentially bind polypyrimidine ssDNA templates, but CspB binds to the T-based templates with one order of magnitude higher affinity than to U- or C-based ssDNA, whereas CspA binds T-, U- or C-based ssDNA with comparable affinity. They also show similarities and differences in their binding to ssDNA at high ionic strength. The results of these findings are related to the chemical structure of DNA bases. PMID- 10862971 TI - No cofactor effect on equilibrium unfolding of Desulfovibrio desulfuricans flavodoxin. AB - Flavodoxins are proteins with an alpha/beta doubly wound topology that mediate electron transfer through a non-covalently bound flavin mononucleotide (FMN). The FMN moiety binds strongly to folded flavodoxin (K(D)=0.1 nM, oxidized FMN). To study the effect of this organic cofactor on the conformational stability, we have characterized apo and holo forms of Desulfovibrio desulfuricans flavodoxin by GuHCl-induced denaturation. The unfolding reactions for both holo- and apo flavodoxin are reversible. However, the unfolding curves monitored by far-UV circular dichroism and fluorescence spectroscopy do not coincide. For both apo- and holo-flavodoxin, a native-like intermediate (with altered tryptophan fluorescence but secondary structure as the folded form) is present at low GuHCl concentrations. There is no effect on the flavodoxin stability imposed by the presence of the FMN cofactor (DeltaG=20(+/-2) and 19(+/-1) kJ/mol for holo- and apo-flavodoxin, respectively). A thermodynamic cycle, connecting FMN binding to folded and unfolded flavodoxin with the unfolding free energies for apo- and holo flavodoxin, suggests that the binding strength of FMN to unfolded flavodoxin must be very high (K(D)=0.2 nM). In agreement, we discovered that the FMN remains coordinated to the polypeptide upon unfolding. PMID- 10862973 TI - Reactive-site specificity of human kallistatin toward tissue kallikrein probed by site-directed mutagenesis. AB - Kallistatin is a serine proteinase inhibitor that forms complexes with tissue kallikrein and inhibits its activity. In this study, we compared the inhibitory activity of recombinant human kallistatin and two mutants, Phe388Arg (P1) and Phe387Gly (P2), toward human tissue kallikrein. Recombinant kallistatins were expressed in Escherichia coli and purified to apparent homogeneity using metal affinity and heparin-affinity chromatography. The complexes formed between recombinant kallistatins and tissue kallikrein were stable for at least 150 h. Wild-type kallistatin as well as both Phe388Arg and Phe387Gly mutants act as inhibitors and substrates to tissue kallikrein as analyzed by complex formation. Kinetic analyses showed that the inhibitory activity of Phe388Arg variant toward tissue kallikrein is two-fold higher than that of wild type (P1Phe), whereas Phe387Gly had only 7% of the inhibitory activity toward tissue kallikrein as compared to wild type. The Phe388Arg variant but not wild type inhibited plasma kallikrein's activity. These results indicate that P1Arg variant exhibits more potent inhibitory activity toward tissue kallikrein while wild type (P1Phe) is a more selective inhibitor of tissue kallikrein. The P2 phenylalanine is essential for retaining the hydrophobic environment for the interaction of kallistatin and kallikrein. PMID- 10862983 TI - Wastewater treatment with particulate biofilm reactors. AB - The review presented in this paper focuses on applications of particulate biofilm reactors (e.g. Upflow Sludge Blanket, Biofilm Fluidized Bed, Expanded Granular Sludge Blanket, Biofilm Airlift Suspension, Internal Circulation reactors). Several full-scale applications for municipal and industrial wastewater treatment are presented and illustrated, and their most important design and operation aspects (e.g. biofilm formation, hydrodynamics, mass transfer, mixing) are analysed and discussed. It is clear from the review that this technology can be considered a grown up technology for which good design and scale-up guidelines are available. PMID- 10862984 TI - Biological desulfurization in an optical-fiber photobioreactor using an automatic sunlight collection system. AB - Biological desulfurization using C. thiosulfatophilum has many more advantages over conventional physico-chemical methods due to low operational cost and no production of secondary pollutants. However, it requires effective and economical supply of light energy, which is a key factor in determining the success of commercialization. In this study, optical-fiber photobioreactor with internal illumination system was applied to increase the light availability. Furthermore, sunlight was used as the main light energy in the daytime and metal-halide lamp was applied as an additional light energy at night. Most UV light was eliminated by the chromatic aberration of the aspherical lenses in the solar light collector and 60% of infrared light intensity was eliminated. Physical scratching optical fibers enhanced the light availability about five times as much as that with unscratched ones in the previous study, but it resulted in the adsorption problem of elementary sulfur particles deteriorating light diffusivity considerably in a long operation. In order to solve this problem, scratched optical fibers were inserted into pyrex-glass tubes, which made light diffusivity nearly the same as that without glass tubes. Removal rate per unit cell concentration, using sunlight in the daytime and a metal-halide lamp at night, was 0.41 <0.73 micromol H(2)S min(-1)/(mg protein l(-1)) using a 400 W metal-halide lamp day and night, since the automatic sunlight collection system can transmit the light intensity as only 10% of that with a metal-halide lamp. PMID- 10862985 TI - Ligands selected from combinatorial libraries of protein A for use in affinity capture of apolipoprotein A-1M and taq DNA polymerase. AB - Here we show that robust and small protein ligands can be used for affinity capture of recombinant proteins from crude cell lysates. Two ligands selectively binding to bacterial Taq DNA polymerase and human apolipoprotein A-1(M), respectively, were used in the study. The ligands were selected from libraries of a randomized alpha-helical bacterial receptor domain derived from staphylococcal protein A and have dissociation constants in the micromolar range, which is typical after primary selection from these libraries consisting of approximately 40 million different members each. Using these ligands in affinity chromatography, both target proteins were efficiently recovered from crude cell lysates with high selectivities. No loss of column capacity or selectivity was observed for repeated cycles of sample loading, washing and low pH elution. Interestingly, column sanitation could be performed using 0. 5 M sodium hydroxide without significant loss of ligand performance. The results suggest that combinatorial approaches using robust protein domains as scaffolds can be a general tool in the process of designing purification strategies for biomolecules. PMID- 10862986 TI - Effect of RQ and pre-seed conditions on biomass and galactosyl transferase production during fed-batch culture of S. cerevisiae BT150. AB - A feedback RQ controlled fed-batch process for the recombinant production of a soluble human N-deglycosylated recombinant beta-1, 4-galactosyltransferase (NdrGal-T) with Saccharomyces cerevisiae BT150 was investigated. Several RQ values were tested for optimal production of NdrGal-T. Four times higher volumetric activity was reached at RQ=1.0 (32 U l(-1)) than at all higher RQ values (about 8 U l(-1)). RQ, 1.0 was the best choice for both, biomass and enzyme production. Optimal concentration of glucose in preculture was 25 g l(-1). At higher values slightly more ethanol was produced than at lower values of preculture glucose concentrations, moreover no positive effect on biomass and enzyme production was found. Lower values caused not only decrease of ethanol but also decrease of biomass formation (from 1.69 g h(-1) to 0.81 g h(-1)) and enzyme overall productivity (from 2.2 U h(-1) to 0.63 U h(-1)). Successfully performed cultivation with three precultures predicted scale-up possibility of feedback RQ controlled NdrGal-T production with S. cerevisiae BT150 from lab to pilot-scale fermentor. PMID- 10862987 TI - Synthetic peptide mimicking of binding sites on olfactory receptor protein for use in 'electronic nose'. AB - A putative tertiary structure model of the dog's olfactory receptor (olfd canfa) is established in this study. By using a target odorous compound (trimethylamine), it is possible to locate the most plausible binding sites between the receptor model structure and the target odorous molecules through computer docking simulations. The two short oligo-peptide sequences (orp61 and orp188) for trimethylamine sensing were identified, synthesized, purified and coated onto the surface of the separate piezoelectric gold electrodes. These two peptides show a high binding capability for trimethylamine. To further enhance the sensitivity of the polypeptides towards the target compound, the polarity and the degree of docking were changed by a site-specific modification technique. The orp61 sequence was modified by substituting two amino acids in the binding pocket resulting in 33% increase in sensitivity towards trimethylamine and reduced noises from other non-target chemicals. The techniques used in the present study offer a unique approach for synthesizing peptides in mimicking binding domain of olfactory receptors. The approach can be easily applied to further development of recognized molecules for gas sensing, especially for use in 'electronic noses'. PMID- 10862988 TI - Expression of highly controllable genes in insect cells using a modified tetracycline-regulated gene expression system. AB - A modified tetracycline-responsive expression system (TRES) for use in insect cells was developed. The TRES contains two components: one encodes a tetracycline controllable transactivator (tTA) and the other contains a tet operator DNA sequence to drive the luciferase gene. Our results show that the human cytomegalovirus (CMV) promoter, an essential part for strong tTA expression in mammalian system, was not functional in insect cells. Thus further modifications were required. Functional tTA was efficiently expressed in Sf9, Sf21, and TN368 cells by the p10 promoter of Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) in plasmid form with virus co-infection. An increase of up to 258 fold of luciferase activity was detected in these cells when both components in modified TRES were co-transfected. In order to further simplify the experiment, tTA, which is driven by the p10 promoter, was inserted into AcMNPV. Luciferase activity was also strongly stimulated by the infection of this tTA expression recombinant virus with the transfection of a plasmid containing the second TRES component expressing luciferase. The luciferase expressions in these systems, either in plasmids or the tTA gene in virus and luciferase in plasmid, were significantly suppressed by tetracycline. The time course kinetics of tetracycline action to the TRES were further studied. Within a time span of 50 h, the luciferase activities could be fully suppressed or activated, respectively, corresponding to the addition or removal of tetracycline. These experiments have established a well-regulated gene expression system for further broad applications of molecular biological studies in insect cells. PMID- 10862989 TI - Estimation of the respiration quotient in a bicarbonate buffered batch cell cultivation. AB - In bicarbonate buffered media the carbon dioxide balance is affected by accumulation and therefore the RQ can not directly be calculated from gas flow and gas concentration measurements. To cope with the buffering capacity of such media a Kalman Filter was designed as a software sensor to estimate the RQ. To eliminate the role of a priori knowledge of cell and medium kinetics on the performance of the Kalman Filter all carbonate was lumped to one term. As a result only physical constants are required. A reference experiment verified the performance of the software sensor. Subsequently several experiments were performed with insect cells to show the progress of the RQ-values during cultivation. PMID- 10862990 TI - Limiting access to allogeneic bone marrow transplantation in five European countries: what can we learn about implicit rationing? AB - BACKGROUND: Allogeneic bone marrow transplantation is a complex procedure exemplifying a class of new and emerging treatment modalities involving advanced medical technologies with high costs. OBJECTIVES: To collect available data on volume of allogeneic bone marrow transplantation in five European countries for the period 1990-1994. To assess the opinion of selected physicians on whether they felt that patients that would have benefited from a transplant actually received a transplant in the same period. To assess their opinion on possible reasons for limited access. METHOD: Literature review and mailed self administered questionnaire to the responsible physician at 76 transplant centres in Denmark, Germany, The Netherlands, Norway, and the UK. RESULTS: Substantial geographical differences in transplantation rates between the five countries studied. For adult patients 62.5% of the respondents felt that there was limited access to allogeneic bone marrow transplantation in the period. For children, the corresponding figure was 37.5%. The reasons for limited access cited most frequently were scarcity of facilities or transplant beds, scarcity of donors, and inadequate referral practices. PMID- 10862991 TI - Importing budget systems from other countries: what can we learn from the German drug budget and the British GP fundholding? AB - The rising costs of pharmaceutical expenditures are a common problem for policy makers in most European countries. In two countries, budget systems for pharmaceutical spending exist(ed). In Great Britain, between 1991 and 1999 GP fundholders were responsible for prescribing costs, and in Germany an overall expenditure cap for pharmaceutical prescribing has been used since 1993. These two examples are analysed in order to identify the conditions that are needed for successfully implementing budget systems for prescribing costs in other countries. It is argued, that a good budget system balances the provision of enough information for budget holders to monitor their expenditures on the one hand, against an explosive increase of transaction costs on the other hand. Apart from that, it makes doctors responsible only for expenditures that they themselves can actually control, and does not provide them with an incentive to use that discretionary power by shifting expenditures to other health care sectors. A good information infrastructure is needed for the implementation of budget systems in general. For the introduction of fundholding, a number of additional criteria need to be met, such as having gate-keeping GPs with personal lists and having a single-payer system. PMID- 10862992 TI - Evaluating the effect of new hospital specialties on GP prescription costs. AB - The aim of this paper is to examine whether the introduction of new hospital specialties contributed to an increase in GP prescription costs. New specialties were introduced in Dr Gray's hospital, Grampian, North-East Scotland in 1994. Data on prescription costs and volume for groups of drugs associated with the new specialties were obtained for all GP practices in Moray (study practices) and Kincardine and Deeside (control practices). Comparing the periods January-April in 1994 with 1995, and 1995 with 1996, an upward trend in GP prescription costs was detected for ulcer healing drugs and anti-depressants. The trend in Kincardine and Deeside also pointed to rising prescription costs, although to a lesser extent. The number of patients referred to the psychiatric and gynaecology specialties expanded after the introduction of these specialties at Dr Gray's. In conclusion, there is some evidence to support the proposition that the introduction of new specialties at Dr Gray's was associated with an increase in the growth of prescription costs within Moray. Further research should establish more clearly whether this is as a result of increased referrals by GPs or the prescribing of more expensive drugs by consultants. The results have implications for the setting of prescribing budgets. PMID- 10862993 TI - The influence of economic evaluation studies on decision making. A European survey. The EUROMET group. AB - Despite the growing activity in the field of health economics very little is known about the influence of economic evaluation studies on health care decision making in the EU member states. Several investigations about the impact of health economic studies on decision making have been performed, but most of them did not involve decision makers themselves. In this paper the results of the EUROMET survey are reported and discussed. Different types of decision makers in nine European countries were surveyed by postal questionnaires, semi-structured interviews and focus group discussions. Questions include issues about the extent of knowledge about economic evaluation, the actual and potential use of study results as well as barriers and incentives in the use of studies. It is concluded that despite the general positive attitude knowledge about the formal methodology is rather limited. Accordingly, results of economic evaluation studies are not widely used in decision making. The results show that institutional dimensions, such as difficulties in transferring budgets, are viewed as important barriers. Also, the lack of credibility of studies is assigned a high relevance. Moreover, decision makers wish for a better explanation of the practical relevance of studies and feel that there is a need for more training in health economics. Considering these requirements a number of recommendations for enhancing the value of health economic studies are given. PMID- 10862994 TI - Change in natural history of opportunistic infections in HIV-infected patients. PMID- 10862995 TI - Needle-stick exposure in the health care setting: do not forget hepatitis C! PMID- 10862996 TI - Microdialysis measurement of glucose in subcutaneous adipose tissue up to three weeks in type 1 diabetic patients. AB - BACKGROUND: Microdialysis of subcutaneous adipose tissue may provide an opportunity to monitor glucose continuously, when the device is connected to an extracorporal glucose sensor. We assessed whether our microdialysis probes are capable of measuring adipose tissue glucose over a prolonged period in Type 1 diabetic patients. Furthermore, the relationship between abdominal skinfold thickness and glucose recovery and the effect of spontaneous glucose excursions on its recovery were evaluated. METHODS: Microdialysis probes were pairwise inserted subcutaneously into the abdominal fat and remained in situ for 3 weeks in eight Type 1 diabetic patients. At days 1, 3, 4, 8, 11, 16, and 18 of probe retention, glucose, as measured by microdialysis, was compared to capillary blood glucose concentrations during a 4 h period. The recovery of glucose obtained by microdialysis was expressed as a percentage of the capillary blood glucose concentration. RESULTS: Eleven of the 16 inserted probes (69%) were evaluable during the complete study. Recovery of glucose was lower at day 1 and 3 (51+/-23% and 56+/-18%, respectively, mean+/-S.D.) compared to values found afterwards (67+/-19%, 72+/-13%, 76+/-14%, 71+/-16%, and 76+/-18%, for day 4, 8, 11, 16, and 18, respectively, for all P<0.05 vs. day 1 and 3). Skinfold thickness was inversely related to the overall 3 week glucose recovery (r=-0.76; P<0.03). Recovery was similar over a wide range of capillary blood glucose concentrations. CONCLUSIONS: Prolonged in vivo retention of microdialysis probes improves the recovery and lowers the variability of adipose tissue-sampled glucose in Type 1 diabetic patients. These findings show that microdialysis-based glucose measurements offer an opportunity for prolonged glucose monitoring. PMID- 10862997 TI - Effects of glucose and insulin levels on adipose tissue glucose measurement by microdialysis probes retained for three weeks in Type 1 diabetic patients. AB - BACKGROUND: To evaluate the effects of acute hyperglycaemia and hyperinsulinaemia on adipose tissue glucose measurements by microdialysis probes inserted for a 3 week period. METHODS: Microdialysis probes were implanted pairwise in abdominal adipose tissue in seven Type 1 diabetic patients and remained in situ during the complete study. Stepped hyperglycaemic hyperinsulinaemic clamps were performed at weekly intervals at which the probes were prepared for microdialysis. Adipose tissue glucose by microdialysis was compared to venous and capillary blood glucose concentrations. RESULTS: The mean time after which the acute rise in blood glucose was first detected was 11.3 min, which corresponds to the system delay of the microdialysis probe. The increase of the glucose concentration in dialysate was completed during the following 16 min. Hyperglycaemia and hyperinsulinaemia did not influence recovery compared to venous blood glucose concentrations, while recovery values compared to capillary blood glucose levels increased slightly under hyperinsulinaemic conditions (P<0.01). CONCLUSIONS: In Type 1 diabetic patients, recovery of glucose in adipose tissue compared to venous and capillary blood does not decrease during acute hyperglycaemia and hyperinsulinaemia. Although there is still a relevant time-delay to monitor a rise in blood glucose, these results show that microdialysis may offer an opportunity for future glucose monitoring over a prolonged time-period. PMID- 10862998 TI - Normal carnitine levels in patients with chronic fatigue syndrome. AB - BACKGROUND: Patients with chronic fatigue syndrome (CFS) complain of muscle pain and impaired exercise tolerance. Previous studies show that this is due to systemic carnitine deficiency. We investigated the hypothesis that carnitine deficiency plays an important role in CFS in female CFS patients and compared their results with neighbourhood controls. METHODS: The level of total carnitine, free carnitine, acylcarnitine and carnitine esters were measured in 25 female CFS patients and 25 healthy matched neighbourhood controls in a blinded fashion. RESULTS: The previously reported decreased level of acylcarnitine in CFS patients was not confirmed. There were also no significant differences in levels of total carnitine, free carnitine and 20 carnitine esters between CFS patients and controls. CONCLUSIONS: The present study demonstrates that serum carnitine deficiency does not contribute to or causes the symptoms in many CFS patients. PMID- 10862999 TI - HIV-infected patient with a Rhodococcus equi pneumonia. AB - The case history of a HIV patient with a pulmonary infect of Rhodococcus equi is presented. He recovered after prolonged treatment with antibiotics and lobectomy. The Rhodococcus equi infection was the presenting symptom of his impaired immune status caused by HIV infection. PMID- 10863000 TI - Acute hepatitis C infection: an indication for alfa interferon therapy? PMID- 10863001 TI - Dephosphorylation-induced structural changes in beta-casein and its amphiphilic fragment in relation to emulsion properties. AB - To promote the understanding of the relationship between emulsifying and molecular properties of proteins/peptides, intact beta-casein (betaCN) and its amphipathic fragment, i.e., betaCN (1-105/107) were dephosphorylated. Dephosphorylation was found not to change significantly their emulsifying properties. Since it is known that the structure of proteins can change upon adsorption onto an interface, the secondary structure of intact beta-casein, its amphipathic fragment, and their dephosphorylated forms, both in solution and after adsorption onto a hydrophobic teflon/water interface, were studied by far UV circular dichroism spectroscopy. An increased content of secondary structure, especially alpha-helix, was found for all samples after adsorption onto teflon. Dephosphorylation increased the helix-forming propensity, especially for amphipathic fragment of beta-casein. No influence of the secondary structure properties on the emulsion-forming and -stabilizing properties was observed, but a relationship between the maximum surface load and the emulsion-stabilizing properties was found. PMID- 10863002 TI - Epinephrine upregulates calpain activity in cultured C2C12 muscle cells. AB - C2C12 cells were grown to confluence at 37 degrees C under a continuous 5% CO(2) stream and myotube formation was stimulated. The cultures were then incubated with or without 2 microg/mL epinephrine for 18 h prior to harvesting and calpain extraction. Epinephrine treatment resulted in a three-fold increase in extractable mu-calpain activity (P < 0.05), a three-fold increase in extractable m-calpain activity (P < 0.05), a 36% increase in calpastatin activity (P < 0.001), and a 16% decrease (P < 0.05) in the total protein content in the C2C12 cell homogenate. These results suggest that calpains may play a role in protein metabolism and that the hormone epinephrine may be directly involved in the regulation of their cellular expression. PMID- 10863003 TI - Chloroform-induced conformational changes in the bound pigmentin bilirubin albumin complexes. AB - Chloroform-induced conformational changes of bilirubin (BR) bound to different serum albumins were studied by circular dichroism (CD) and fluorescence spectroscopy. Addition of a small amount of chloroform ( approximately 20 mM) to a solution containing 20 microM albumin and 15 microM BR changed the sign order and magnitude of the characteristic CD spectra of all BR-albumin complexes except BR-PSA complex which showed abnormal behavior. Monosignate negative CD Cotton effects (CDCEs) of BR complexed with SSA, GSA and BuSA were transformed into bisignate CDCEs in presence of chloroform akin to those exhibited by chloroform free solution of BR-HSA complex, indicating that the pigment acquired right handed plus (P) chirality when chloroform was added to these complexes. Bisignate CD spectra of BR complexed with HSA and BSA showed complete inversion upon addition of chloroform corroborating earlier findings. On the other hand, changes observed with BR-RSA complex were slightly different showing an additional CD band of weak intensity centered around 390 nm though inversion of CDCEs was similar to that of BR-HSA complex. Monosignate CD spectra of BR-PSA complex also showed three CD bands occurring at 409, 470 and 514 nm after chloroform addition. These results indicated significant but different effects of chloroform on the conformation of bound BR in BR-albumin complexes which can be ascribed to the changes in the exciton chirality of bilirubin probably due to altered hydrophobic microenvironment induced by the binding of chloroform at or near the ligand binding site. Chloroform severely quenched the intrinsic tryptophan fluorescence of the protein and shifted the emission maxima towards blue region in all the albumins except PSA. However, quantitative differences in both quenching and blue shift were noted in different serum albumins. This suggests that chloroform probably binds in the close vicinity of tryptophan residue(s) located in subdomain(s) IIA or IB and II both. The fluorescence of BR-albumin complexes was also found to be sensitive to the presence of a small amount of chloroform. But the changes observed in the fluorescence of the bound pigment in presence of chloroform were less marked as compared to the changes in the intrinsic fluorescence of protein per se. Taken together, these results suggest that there is at least one conserved site for chloroform binding in all these albumins which is at or near the BR binding site. PMID- 10863004 TI - Nucleotide sequence analysis, overexpression in Escherichia coli and kinetic characterization of Anacystis nidulans catalase-peroxidase. AB - Bifunctional catalase-peroxidases are the least understood type of peroxidases. A high-level expression in Escherichia coli of a fully active recombinant form of a catalase-peroxidase (KatG) from the cyanobacterium Anacystis nidulans (Synechococcus PCC 6301) is reported. Since both physical and kinetic characterization revealed its identity with the wild-type protein, the large quantities of recombinant KatG allowed the examination of both the spectral characteristics and the reactivity of its redox intermediates by using the multi mixing stopped-flow technique. The homodimeric acidic protein (pI = 4.6) contained high catalase activity (apparent K(m) = 4.8 mM and apparent k(cat) = 8850 s(-1)). Cyanide is shown to be an effective inhibitor of the catalase reaction. The second-order rate constant for cyanide binding to the ferric protein is (6.9 +/- 0.2) x 10(5) M(-1 )s(-1) at pH 7.0 and 15 degrees C and the dissociation constant of the cyanide complex is 17 microM. Because of the overwhelming catalase activity, peroxoacetic acid has been used for compound I formation. The apparent second-order rate constant for formation of compound I from the ferric enzyme and peroxoacetic acid is (1.3 +/- 0.3) x 10(4 )M(-1 )s(-1) at pH 7.0 and 15 degrees C. The spectrum of compound I is characterized by about 40% hypochromicity, a Soret region at 406 nm, and isosbestic points between the native enzyme and compound I at 355 and 428 nm. Rate constants for reduction of KatG compound I by o-dianisidine, pyrogallol, aniline and isoniazid are shown to be (7.3 +/- 0.4) x 10(6) M(-1 )s(-1), (5.4 +/- 0.3) x 10(5) M(-1 )s(-1), (1.6 +/- 0.3) x 10(5) M(-1 )s(-1) and (4.3 +/- 0.2) x 10(4) M(-1 )s(-1), respectively. The redox intermediate formed upon reduction of compound I did not exhibit the classical red-shifted peroxidase compound II spectrum which characterizes the presence of a ferryl oxygen species. Its spectral features indicate that the single oxidizing equivalent in KatG compound II is contained on an amino acid which is not electronically coupled to the heme. PMID- 10863005 TI - Co-conservation of rRNA tetraloop sequences and helix length suggests involvement of the tetraloops in higher-order interactions. AB - Terminal loops containing four nucleotides (tetraloops) are common in structural RNAs, and they frequently conform to one of three sequence motifs, GNRA, UNCG, or CUUG. Here we compare available sequences and secondary structures for rRNAs from bacteria, and we show that helices capped by phylogenetically conserved GNRA loops display a strong tendency to be of conserved length. The simplest interpretation of this correlation is that the conserved GNRA loops are involved in higher-order interactions, intramolecular or intermolecular, resulting in a selective pressure for maintaining the lengths of these helices. A small number of conserved UNCG loops were also found to be associated with conserved length helices, consistent with the possibility that this type of tetraloop also takes part in higher-order interactions. PMID- 10863006 TI - Two approaches to isolate cytoplasmic dynein ATPase from Neurospora crassa. AB - Cytoplasmic dynein is a force-producing enzyme that, in association with dynactin, conducts minus-end directed transport of various organelles along microtubules. Biochemical analyses of cytoplasmic dynein and dynactin have been conducted primarily in vertebrate systems, whereas genetic analyses have been explored mainly in yeast and the filamentous fungi. To provide a complementary biochemical approach for the study of fungal dynein, we isolated/partially purified cytoplasmic dynein ATPase from the filamentous fungus Neurospora crassa. N. crassa dynein was partially purified by slightly modifying the existing procedures, described for mammalian cytoplasmic dynein that uses dynein microtubule binding, followed by release with ATP and sucrose gradient fractionation. A novel approach was also used to isolate dynein-specific ATPase by gel filtration (Sepharose CL-4B). The K(m), ATP obtained by isolating dynein ATPase using gel filtration was similar to that obtained by using conventional method, suggests that contaminant proteins do not interfere with the dynein ATPase activity. Like vertebrate dynein, N. crassa dynein is a general NTPase with highest activity toward ATP, and only the ATPase activity is stimulated by microtubules. The K(m), ATP for N. crassa cytoplasmic dynein is 10- to 15-fold higher than that of the vertebrate enzyme. PMID- 10863007 TI - Bacterial luminescence: luminescence mechanism with cyclic peroxide participation and dependence on reactive oxygen species (a hypothesis). AB - Chemically initiated exchange (CIEE) luminescence reactions were reviewed and a new mechanism of luminescence with peracid as an intermediate is proposed; bacterial luminescence is generally considered to be a case of dioxetane luminescence, or, to be more precise, CIEE-luminescence which includes the generation of a cyclic peroxide. In the hypothesis the monooxygenase reaction (aldehyde -->fatty acid) should not be coupled with emitter generation as is usually believed, but only with the generation of peracid. As to the generation of the emitter, excited flavin, it is likely to occur later, during the interaction of flavin with cyclic peroxide. Its consequence is the breaking of two chemical bonds (O-O and C-C) in the cyclic peroxide and simultaneous generation of 4alpha-hydroxyflavin in exited state. In general, the generation of light includes three stages: 1) the monooxygenase reaction and the concurrent production of peracid; 2) the conversion of peracid to cyclic peroxide; and 3) the interaction of cyclic peroxide with flavin (through the CIEE mechanism). PMID- 10863008 TI - Amino acid sequence of piratoxin-II, a myotoxic lys49 phospholipase A(2) homologue from Bothrops pirajai venom. AB - The complete amino acid sequence of the 121 amino acid residues of piratoxin II, a phospholipase A(2) like myotoxin from Bothrops pirajai venom, is reported. PrTX II is a basic protein with a molecular mass of 13740 Da, a calculated pI of 9.03, but an experimental pI of 8.4 +/- 0.2, showing sequence similarity with other bothropic (90-99%) or non-bothropic ( approximately 80%) Lys49 PLA(2)-like myotoxins. This similarity falls to approximately 70% when this sequence is aligned with that of Asp49 PLA(2). Due to the substitution of Asp49 by Lys49 and alterations in the calcium binding loop structure, as the replacement of Gly32 by Leu32, piratoxin-II shows no PLA(2) activity when assayed on egg yolk. Piratoxin II showed the same primary structure as piratoxin-I, except that it has Lys116 for Leu116. Despite this slightly higher basicity at the C-terminal region, piratoxin-II was shown to be less myotoxic than piratoxin-I. The change Leu --> Lys induced an alteration of the molecule surface shape and probably of the environment charge high enough to slightly decrease the myotoxic activity. When aligned with B. jararacussu bothropstoxin-I and with B. asper Basp-II, piratoxin II revealed a single (position 132) and a quintuple (positions 17, 90, 111, 120 and 132) amino acid substitution, respectively, suggesting a common evolutionary origin for these three myotoxins. PMID- 10863009 TI - The role of tryptophan residues in the hemolytic activity of stonustoxin,a lethal factor from stonefish (Synanceja horrida) venom. AB - Stonustoxin (SNTX) is a pore-forming cytolytic lethal factor, isolated from the venom of the stonefish Synanceja horrida, that has potent hemolytic activity. The role of tryptophan residues in the hemolytic activity of SNTX was investigated. Oxidation of tryptophan residues of SNTX with N-bromosuccinimide (NBS) resulted in loss of hemolytic activity. Binding of 8-anilino-1-naphthalenesulphonate (ANS) to SNTX resulted in occlusion of tryptophan residues that resulted in loss of hemolytic activity. Circular dichroism and fluorescence studies indicated that ANS binding resulted in a conformational change of SNTX, in particular, a relocation of surface tryptophan residues to the hydrophobic interior. NBS modification resulted in oxidised surface tryptophan residues that did not relocate to the hydrophobic interior. These results suggest that native surface tryptophan residues play a pivotal role in the hemolytic activity of STNX, possibly by being an essential component of a hydrophobic surface necessary for pore-formation. This study is the first report on the essentiality of tryptophan residues in the activity of a lytic and lethal factor from a fish venom. PMID- 10863010 TI - Evaluating the environmental impact of an old municipal waste incinerator: PCDD/F levels in soil and vegetation samples. AB - In order to determine the temporal variation in the levels of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) in the vicinity of an old municipal solid waste incinerator (MSWI) (S. Adria del Besos, Barcelona, Spain), 24 soil and vegetation samples were collected at the same sampling points in which samples had been taken 1 year before. Each sample was analyzed for PCDDs and PCDFs by high-resolution gas chromatography/high resolution mass spectrometry. While in the previous study PCDD/F concentrations in soil ranged from 1.22 to 34. 28 ng I-TEQ/kg (median and mean values: 9.06 and 12.24 ng I-TEQ/kg), in the present study, PCDD/F levels ranged from 1.33 to 54.23 ng I-TEQ/kg (median and mean values: 11.85 and 14.41 ng I-TEQ/kg). On the other hand, in the previous study, PCDD/F levels in vegetation ranged from 0.33 to 1.98 ng I-TEQ/kg (median and mean values: 0.58 and 0.70 ng I-TEQ/kg), whereas in the present study, PCDD/F levels ranged from 0.32 to 2.52 ng I-TEQ/kg (median and mean values: 0.82 and 0.97 ng I-TEQ/kg). During the last 12 months, PCDD/F levels increased in 16 of the 24 soil samples and in 17 of the 24 vegetation samples analyzed. However, no significant differences in the median I-TEQ concentrations of both studies were found either in soil or vegetation samples. PMID- 10863011 TI - Regulatory framework for the thermal treatment of various waste streams. AB - Since 1990, regulations and standards have changed considerably. This article is an update of the regulatory requirements for the thermal treatment of various waste streams. The waste categories covered, along with the laws they are governed under, include: Hazardous waste under Subtitle C of the Resource Conservation and Recovery Act (RCRA) and under the Clean Air Act; municipal solid waste under Subtitle D of the RCRA; medical waste under Subtitle J of the RCRA; Superfund waste under the Comprehensive Environmental Response, Compensation and Liability Act (CERCLA); toxic waste under the Toxic Substances Control Act (TSCA); and sludge waste under the Clean Water Act (CWA). PMID- 10863012 TI - Modeling mobile source emissions in presence of stationary sources. AB - The impact of oxides of nitrogen (NO(x)) emissions from motor vehicles to the air quality in city-state Singapore is analyzed using AIRVIRO, a regional scale dispersion model developed by the Swedish Meteorological and Hydrological Institute. In a predominantly urban location like Singapore, it is difficult to separate out the contribution of pollutants from mobile and point sources at different locations. In this work, a new approach is used by first modeling only the impact of point and area sources and then overlaying the traffic impact on air quality at different locations. Monthly scenario simulations are run with point, area and traffic sources of emissions for the Gaussian model validation. Street Canyon modeling is used for street segments surrounded by buildings on either side. A simplified photochemical model, which takes into account NO(x) undergoing chemical transformations in the urban atmosphere, is used to account for variations in NO(x) and ozone levels with respect to traffic data. The diurnal variation of NO(x) concentration levels is studied as a function of ozone levels at site, hourly traffic counts and meteorological parameters. The impact on ambient air quality within the breathing zone of the public from mobile sources, is found to be about 40% at urban stations although overall emissions from mobile sources is only 24%. The proposed approach appears to predict the variations in NO(x) as a function of traffic and meteorological conditions. PMID- 10863013 TI - Evaluating the risk from depleted uranium after the Boeing 747-258F crash in Amsterdam, 1992. AB - On 4 October 1992, a large cargo plane crashed into an apartment building in the Bijlmermeer quarter of Amsterdam. In the years following the accident, an increasing number of people started reporting health complaints, which they attributed to exposure to dangerous substances after the crash. Since the aircraft had been carrying depleted uranium as counterbalance weights and about 150 kg uranium had been found missing after clearance of the crash site, exposure to uranium oxide particles was pointed out as the possible cause of their health complaints. Six years after the accident, a risk analysis was therefore carried out to investigate whether the health complaints could be attributed to exposure to uranium oxide set free during the accident. The scientific challenge was to come up with reliable results, knowing that - considering the late date - virtually no data were available to validate any calculated result. The source term of uranium was estimated using both generic and specific data. Various dispersion models were applied in combination with the local setting and the meteorological conditions at the time of the accident to estimate the exposure of bystanders during the fire caused by the crash. Emphasis was given to analysing the input parameters, inter-comparing the various models and comparing model results with the scarce information available. Uranium oxide formed in the fire has a low solubility, making the chemical toxicity to humans less important than the radiotoxicity. Best-estimate results indicated that bystanders may have been exposed to a radiation dose of less than 1 microSv, whereas a worst-case approach indicated an upper limit of less than 1 mSv. This value is considerably less than the radiation dose for which acute effects are to be expected. It is therefore considered to be improbable that the missing uranium had indeed led to the health complaints reported. PMID- 10863014 TI - Hazardous materials in the environment of Dnepropetrovsk region (Ukraine). AB - The investigations were aimed at assessment of environmental pollution in one of the most industrialized regions of Ukraine - Dnepropetrovsk Region. The following types of environmental contamination were considered in the study: emissions and concentrations of 16 air pollutants; content and distribution of 15 elements in soils and plants at the polluted and unpolluted territories. The investigations were conducted at 28 urban sites and 18 rural sites of the Region during 1991 1998 years. Level and character of air, soil and plants contamination were investigated. Statistical methods were used to describe quantitatively the relationships between contents of hazardous materials in the environment. It was found that concentrations of fluoride, iron, copper, zinc, and lead in the soil and contents of fluoride, iron, nickel, cadmium, and aluminum in plants were several times higher than normal. PMID- 10863015 TI - A foundation for the risk-based treatment of gasoline-contaminated soils using modified Fenton's reactions. AB - The relative oxidation of representative aromatic and aliphatic hydrocarbons found in gasoline was evaluated to provide the foundation for risk-based treatment of petroleum-contaminated soils and groundwater using modified Fenton's reagent (catalyzed hydrogen peroxide). Aromatic components of gasoline are considered more hazardous than the aliphatic fractions due to their higher mobility in the subsurface and their higher acute and chronic toxicities. Benzene, toluene, and mixed xylenes (BTX) were selected as aromatic compounds representative of unleaded gasoline, while nonane, decane, and dodecane (NDD) were used as model aliphatic compounds. The effects of hydrogen peroxide (H(2)O(2)) concentration, iron catalyst concentration, and pH on the degree of treatment of the model compounds were investigated using central composite rotatable experimental designs. Oxidation of the aromatic compounds required less iron and less H(2)O(2) than did oxidation of the aliphatic compounds, while proceeding more effectively at near-neutral pH. Greater than 95% of the BTX was treated at near-neutral pH using 2. 5% H(2)O(2) and 12.5 mM iron (III), while only 37% nonane, 7% decane, and 1% dodecane oxidation was achieved under the same conditions. The results show that the more toxic and mobile aromatic fraction was more effectively oxidized using less H(2)O(2) and more economical conditions, including near-neutral pH, compared to the aliphatic fraction. A process design based on treating only the aromatic fraction of petroleum may provide significantly lower costs when using modified Fenton's reagent for the treatment of contaminated soils and groundwater. PMID- 10863016 TI - Performance enhancement of batch aerobic digesters via addition of digested sludge. AB - The impact of the feed sludge (FS) concentration and addition of digested sludge (DS) to an aerobic digester was evaluated with respect to its capability for removal of the total suspended solids (TSS) and volatile suspended solids (VSS). The aerobic digesters, which operated in a batch mode at constant temperature and mixing rate, were initially filled with FS to 25%, 50%, 75%, and 100% of the reactor's volume. The remaining volume of the reactor was occupied by the DS, having DS/FS ratio of 3, 1, 1/3, and 0. Analysis of the experimental data showed that in the absence of DS, TSS, and VSS destruction rates are very small; however, increasing DS/FS ratio from 1/3 to 3 results in 74-77% increase in VSS and TSS destruction, respectively. The increase of the DS/FS ratio associated with increased ratio of the measured viable biomass (Cc) to VSS concentration (Xv) suggested that DS serves as the source of viable cell mass needed for degradation of organic solids. Assuming pseudo-first-order kinetics, it was shown that while organic solid destruction rate constants (k) are inversely related to initial concentrations of sludge, their values increase with increasing DS/FS ratios. PMID- 10863017 TI - The behaviour of Al in MSW incinerator fly ash during thermal treatment. AB - Fly ash from municipal solid waste (MSW) incinerators contains leachable metals, including potentially hazardous heavy metals. The metal content of the fly ash can be reduced by thermal treatment, which vaporizes the volatile metal compounds. After heat treatment of fly ash at 1000 degrees C for 3 h, less metal was able to be leached from the thermally treated ash than from the ash without thermal treatment. Al and Cr were the exceptions. These metals were more soluble in the ash that had been thermally treated. This paper focuses on the leaching behaviour of Al only. Both simple and sequential extraction leaching tests showed that the leachable Al for the heat-treated fly ash is about twice that of the untreated fly ash. The sequential test further revealed that (i) the majority of the leachable Al is associated with Fe-Mn oxides in the fly ash, and (ii) most of the unleachable Al resides in the silicate matrices of the heat-treated and untreated fly ash. Pure chemicals, Al(2)O(3), CaO and CaCl(2), simulating the relevant ingredients in the fly ash, were used for studying their reactions at 1000 degrees C. The aluminum compounds were identified by X-ray Diffraction (XRD). Two new chemical phases produced by the thermal treatment were identified; Ca(AlO(2))(2) and 12CaO.7Al(2)O(3). Their formation suggests a mechanism whereby thermal treatment of fly ash would produce more soluble Al. PMID- 10863018 TI - Kinetic study of the reaction between sulfur dioxide and calcium hydroxide at low temperature in a fixed-bed reactor. AB - A quantitative study of the influence of inlet sulfur dioxide concentration (600 3000 ppm), relative humidity (20-60%), reactor temperature (56-86 degrees C) and different amounts (0-30 wt.%) of inorganic additives (NaCl, CaCl(2) and NaOH) on gas desulfurization has been carried out in a continuous downflow fixed-bed reactor containing calcium hydroxide diluted with silica sand. Results show that the reaction rate does not depend on sulfur dioxide partial pressure (zero-order kinetics) and that the temperature and the relative humidity have a positive influence on reaction rate. An apparent activation energy of 32 kJ/mol Ca(OH)(2) has been estimated for the reaction. An empirical reaction rate equation at 71.5 degrees C and 36.7% relative humidity that includes the type and amount of additive is proposed. It has been found that calcium chloride is the best additive studied because it allows for a higher degree of sulfur dioxide removal. PMID- 10863019 TI - The management of arsenic wastes: problems and prospects. AB - Arsenic has found widespread use in agriculture and industry to control a variety of insect and fungicidal pests. Most of these uses have been discontinued, but residues from such activities, together with the ongoing generation of arsenic wastes from the smelting of various ores, have left a legacy of a large number of arsenic-contaminated sites. The treatment and/or removal of arsenic is hindered by the fact that arsenic has a variety of valence states. Arsenic is most effectively removed or stabilized when it is present in the pentavalent arsenate form. For the removal of arsenic from wastewater, coagulation, normally using iron, is the preferred option. The solidification/stabilization of arsenic is not such a clear-cut process. Factors such as the waste's interaction with the additives (e.g. iron or lime), as well as any effect on the cement matrix, all impact on the efficacy of the fixation. Currently, differentiation between available solidification/stabilization processes is speculative, partly due to the large number of differing leaching tests that have been utilized. Differences in the leaching fluid, liquid-to-solid ratio, and agitation time and method all impact significantly on the arsenic leachate concentrations. This paper reviews options available for dealing with arsenic wastes, both solid and aqueous through an investigation of the methods available for the removal of arsenic from wastewater as well as possible solidification/stabilization options for a variety of waste streams. PMID- 10863020 TI - Removal of heavy metals from aqueous solution by chelating resin in a multistage adsorption process. AB - Copper and zinc removal from aqueous solution by chelating resin was investigated theoretically and experimentally in the present study. A multistage process was proposed as an alternative for enhancement of the heavy removal of the single stage process. Heavy metal mass balance equations with empirical Freundlich adsorption isotherm were developed to represent the multistage process and the theoretical model permits determination of the inter-stage heavy metal concentrations and the total amount of chelating resin required for achieving a desired level of heavy metal removal. Optimization of the linearized theoretical model shows that equal division of the total amount of chelating resin among all stages of the multistage process yields the best results in terms of saving of chelating resin for a given heavy metal removal or enhanced heavy metal removal for a given total amount of chelating resin. Experimental tests were also conducted to establish the equilibrium adsorption of heavy metal by the chelating resin and to empirically verify the advantages of the multistage adsorption process. PMID- 10863021 TI - Effect of frequency separation and stimulus rate on the mismatch negativity: an examination of the issue of refractoriness in humans. AB - Refractoriness of the generators of the mismatch negativity (MMN) was examined in two experiments in which two deviant tones occurred in a row. In Experiment 1, the size of the MMN elicited by the first deviant was manipulated by using deviants that were close to or far from the standard in frequency. In Experiment 2, the time between two identical deviants was varied. It was found that neither the size of the MMN elicited by the first deviant, nor the time between two deviants, affected the amplitude of the MMN elicited by the second deviant. It was concluded that refractoriness played no role in the amplitude of the MMN for the parameters used. PMID- 10863022 TI - Murine macrophages stimulated with central and peripheral nervous system myelin or purified myelin proteins release inflammatory products. AB - Macrophage inflammatory products including tumor necrosis factor (TNF) and interleukin-12/p40 are implicated in demyelinating diseases such as multiple sclerosis (MS), Guillain-Barre syndrome, and animal models experimental autoimmune encephalomyelitis and neuritis. The macrophage product angiotensin converting enzyme (ACE) is released during inflammation. ACE can also be elevated in MS. We investigated the ability of central (CNS) and peripheral nervous system (PNS) myelin to stimulate TNF, interleukin-12, and ACE production by murine macrophages. Both CNS and PNS myelin and purified myelin basic protein and P2 protein induced release of these products. Direct stimulation by myelin may represent a mechanism of inducing release of macrophage products in inflammatory demyelination or neural injury. PMID- 10863023 TI - Massive accumulation of M and H subunits of neurofilament proteins in spinal motor neurons of neurofilament deficient Japanese quail, Quv. AB - Quiver (Quv) is a non-sense mutation of neurofilament protein L subunit (NF-L) that causes neurofilament deficiency with preserved microtubules in Japanese quail. Anti-NF-M and anti-NF-H mAbs stained cell bodies of motor neurons in Quv embryo spinal cords much more intense than those in control spinal cords. Volume of motor neurons in Quv spinal cords increased to 2.3 times of control motor neurons. Immunoblot of Quv spinal cords revealed a relative increase in non- and hypo-phosphorylated NF-M and NF-H, and a decrease in the total amount of NFs. Quv sciatic nerves showed faintly reacted phosphorylated NF-M and NF-H. These results suggest that deficiency of assembled neurofilament results in decreased axonal transport of NFs and accumulation of NFs in cell bodies of spinal motor neurons. PMID- 10863024 TI - Bimodal effects of acetylcholine on synchronized calcium oscillation in rat cultured cortical neurons. AB - To understand the plastic changes in neural network dynamics, we analyzed the modulating effects of cholinergic agonists on synchronized Ca(2+) oscillation in the rat primary cultured cortical neurons. At 5-10 days after establishment of the culture, neurons exhibited synchronized Ca(2+) oscillation. This Ca(2+) oscillation was derived from glutamatergic and gamma-aminobutyric acid (GABA)ergic synaptic inputs from neighboring neurons. Application of acetylcholine and nicotine increased, decreased, or did not change the frequency of synchronized Ca(2+) oscillation, depending on the colonies in the culture. The presence of cholinergic neurons in the culture was confirmed by immunocytochemical staining. Our study provided evidence for the first time that cholinergic neurons exert excitatory influences on GABAergic neurons as well as glutamtergic neurons, resulting in bimodal effects on the frequency of synchronized Ca(2+) oscillation in the cortical neural network. PMID- 10863025 TI - Distribution of P2X purinoceptor clusters on individual rat dorsal root ganglion cells. AB - The distribution of P2X receptors on the cell bodies of acutely dissociated rat dorsal root ganglion cells has been determined using immunohistochemistry. Only medium and small ganglion cells showed P2X receptor labelling, which took the form of receptor clusters 0. 2-0.5 microm in diameter. P2X(3) was the predominant cluster type although clusters of all other subtypes, P2X(1), P2X(2), P2X(4), P2X(5) and P2X(6) were observed. There was little evidence for colocalization of the different receptor subtypes pointing to very little heteropolymerization of P2X receptors in the ganglion. PMID- 10863026 TI - Detection of tau phosphorylated at threonine 231 in cerebrospinal fluid of Alzheimer's disease patients. AB - A new sandwich ELISA is described which shows specificity for tau phosphorylated at threonine 231 and preferentially reacts with Alzheimer's disease (AD) brain extracts relative to other dementias. This assay was used to analyze 58 antemortem cerebrospinal fluid samples. Twenty-three of 27 AD samples (85% sensitivity) yielded signals greater than the cutoff, while only one of 31 non-AD samples (97% specificity) were greater. This indicates that detection of phosphotau in cerebrospinal fluid with this sandwich ELISA could prove useful in the diagnosis of AD. PMID- 10863027 TI - Protective effects of the green tea polyphenol (-)-epigallocatechin gallate against hippocampal neuronal damage after transient global ischemia in gerbils. AB - Recent studies have shown that green tea polyphenols reduce free radical-induced lipid peroxidation. Oxygen free radical injury plays an important role in neuronal damage induced by brain ischemia and reperfusion. The purpose of this study was to examine whether (-)-epigallocatechin gallate (EGCG) would reduce neuronal damage after transient global ischemia in the gerbils because EGCG has a potent antioxidant property as a green tea polyphenol. To produce transient global ischemia, both common carotid arteries were occluded for 3 min with microaneurysmal clips. The gerbils were treated with EGCG (10, 25, or 50 mg/kg, i.p.) immediately after ischemia. Neuronal cell damage in the hippocampal CA1 region was evaluated quantitatively 5 days after ischemia by a blinded investigator. EGCG at the dose of 10 mg/kg failed to reduce hippocampal neuronal damage. However, EGCG when administered at the dose of 25 or 50 mg/kg significantly reduced hippocampal neuronal damage in a dose-dependent manner (P<0.001, respectively). The present results show that the green tea polyphenol, EGCG, has a neuroprotective effect against neuronal damage following global ischemia in the gerbils. PMID- 10863028 TI - Hemispheric lateralization at different levels of human auditory word processing: a functional magnetic resonance imaging study. AB - We used functional magnetic resonance imaging to disentangle the functional anatomy of brain systems involved in the processing of auditory word form and meaning. Three monitoring tasks on auditory stimuli, aimed at phonetic, lexical and semantic processing, were used. We found no lateralization of temporal lobe activations, when word processing was contrasted versus the complex phonetic task. Bilateral middle temporal activations (Brodmann Area [BA] 21) were attributed to processing of word-form. Areas specific to semantic processing were restricted to the left hemisphere: the posterior middle frontal (BA 9) and posterior parietal (BA 7/40) cortex, as well as an inferior temporal area (BA 20/21). Our data suggest, that left hemispheric dominance for auditory word comprehension occurred at the level of semantic processing. PMID- 10863029 TI - Strong olfactory stimulation reduces seizure susceptibility in amygdala-kindled rats. AB - Seizures in human temporal lobe epilepsy are characterized by paroxysmal activity in the limbic system. The primary olfactory or piriform cortex is a central part of the limbic system. Since a relationship between olfactory sensation and limbic seizures has been described, we were interested in the effect of strong olfactory stimulation on the seizure susceptibility of amygdala-kindled rats, a model of human temporal lobe epilepsy. Olfactory stimulation with toluene was able to suppress seizures in most kindled rats after stimulation at 20% above the threshold for eliciting epileptic afterdischarges. Olfactory stimulation with toluene or ammonia increased the threshold by 27 and 25% compared to control conditions. Our data substantiate that olfactory brain regions, such as the piriform cortex, are involved in amygdala-kindled seizures and suggest that strong physiological stimulation of this nucleus interferes with on-going seizure activity in the limbic system. Thus, olfactory stimulation could contribute to anticonvulsant therapy. PMID- 10863030 TI - Morphology of somatosensory evoked fields: inter-hemispheric similarity as a parameter for physiological and pathological neural connectivity. AB - The morphology of somatosensory evoked fields (SEF) following electrical separate stimulation of left and right median nerve in 22 healthy volunteers was explored. The analysis of morphological properties of individual SEFs has been performed, in order to quantify an inter-hemispheric correlation coefficient. Despite a high inter-subject variability of the SEF morphologies, a high intra-subject inter hemispheric shape correlation occurs in the post-stimulus epoch of 100 ms from the stimulus onset. Accordingly, a normative value for the parameter describing the SEF inter-hemispheric correlation coefficient has been calculated. The present work establishes quantitative and normative descriptions of the shape similarity of SEFs from the two hands in the left and right hemispheres. This provides a database for a new parameter, which might be useful in detecting alterations in the primary sensory cortical activation due to possible unilateral alterations of sensory inflow because of either peripheral or central deficits. The described procedure has been successfully applied to a small number of patients affected by unilateral cerebrovascular lesion. PMID- 10863031 TI - Intracerebroventricular administration of brain-derived neurotrophic factor in adult rats affects analgesia and spontaneous behaviour but not memory retention in a Morris Water Maze task. AB - The present study tested the effects of in vivo administration of brain-derived neurotrophic factor (BDNF) and of its antibody (anti-BDNF) in a Morris Water Maze (MWM) task. Adult male rats were trained for three days in a MWM. At the end of the last training trial, subjects were injected intracerebroventricularly with one of the following: (i) BDNF (24 microg); (ii) anti-BDNF (25 microg); or (iii) vehicle (PBS, injection volume 10 microl). On day 5, subjects were tested for memory retention, pain sensitivity and locomotor behaviour. No differences emerged in the MWM as a function of treatment, even with a reduced number of acquisition trials. Nonetheless, BDNF affected both pain threshold in the hot plate test, as well as exploratory behaviour in the open field test. PMID- 10863032 TI - Circulating levels of tumour necrosis factor-alpha and its soluble receptors are increased in the blood of patients with amyotrophic lateral sclerosis. AB - An immunologic pathogenesis for amyotrophic lateral sclerosis (ALS) has been recently proposed. We tested the whole tumour necrosis factor (TNF) system in the serum of 51 ALS patients at different stages of the disease and 36 healthy controls. Antigenic TNF-alpha and its soluble receptors (sTNF-Rs), measured by ELISA, were significantly higher in ALS patients than in healthy controls. However, biologically active TNF-alpha, corresponding to the sTNF-Rs-unbound trimeric TNFalpha molecule and assayed by its cytotoxic activity on a sensitive cell line, was similar between ALS patients and healthy controls. Neither antigenic TNF-alpha, bioactive TNF-alpha nor sTNF-Rs correlated with disease severity, disease duration, or weight loss. In conclusion, we reported an activation of the TNF system in ALS. The role of this activation in the pathogenesis of the disease remains elusive. PMID- 10863033 TI - The stretch-inactivated channel, a vanilloid receptor variant, is expressed in small-diameter sensory neurons in the rat. AB - Exposure to hypertonic conditions is known to produce pain and activate small diameter sensory neurons. Recently, the vanilloid receptor variant and stretch inactivated ion channel (SIC) was cloned and shown to mediate an inward current in response to cell shrinkage. Since other vanilloid receptors have been previously shown to mediate nociception, we investigated whether SIC is expressed in sensory neurons. Using reverse transcription-polymerase chain reaction and in situ hybridization techniques, we identified SIC in the neurons of dorsal root and trigeminal ganglia. Furthermore, SIC was found to be present almost exclusively in the small-diameter sensory neurons, which includes the nociceptive population. Since SIC is activated by cell shrinkage, it may participate in the mediation of pain produced by hypertonic stimuli. PMID- 10863034 TI - Voltammetric and functional evidence that N-methyl-D-aspartate and substance P mediate rat vascular relaxation via nitrogen monoxide release. AB - It is known that substance P acts as a vasodilator via activation of the enzyme nitrogen monoxide synthase (NOS) in endothelial tissue and it is suggested that N methyl-D-aspartate (NMDA) could stimulate nitrogen monoxide (NO) release within nervous tissue. However, the data reported concern NO metabolites (nitrites, nitrates), while there is no clear evidence to date of the action of the latter compound within the aortic tissue. In this study, amperometry with specifically prepared carbon fiber electrodes has been applied to examine the effect of NMDA or substance P upon NO release. In particular, the data obtained confirm that NMDA can stimulate NO release in vivo, in the striatum of anaesthetized rats, and that substance P can stimulate NO release in rat aortic rings (ex vivo experiments). In addition, they indicate that NMDA also stimulates NO release in rat aortic rings. This original data has been confirmed by the observation of a vasorelaxant action of NMDA within noradrenaline precontracted aortic rings. Thus, these experiments provide the first direct evidence that NMDA can mediate vascular relaxation via NO release. PMID- 10863035 TI - Ischemic tolerance conferred to cultured rat neurons by heat shock is not mediated by opening of adenosine triphosphate-sensitive potassium channels. AB - The effect of sublethal heat shock on the capacity of neurons to resist subsequent ischemia-reperfusion-induced cell injury, was studied in a model of primary rat neuronal cultures, subjected to chemical ischemia. Exposure of the cultures to sublethal heat shock (42 degrees C; 20 min) resulted in elevation in cellular content of heat shock protein (HSP)-70, at 4 h following the shock, and in acquisition of a 15 h 'time window of protection' against ischemia-reperfusion insult, with maximum protection at 4 h. Presence in the culture medium of glibenclamide (2 microM), a blocker of ATP sensitive potassium (K(ATP)) channels, did not abolish the acquisition of protection throughout the entire duration of the acquired 'time window of protection'. The results demonstrate that heat shock induces in neurons a protective mechanism against ischemia-reperfusion insult, probably associated with enhanced expression of HSPs, which does not depend on opening of K(ATP) channels. In this respect, the neuronal 'heat-shock mechanism' for the acquisition of ischemic tolerance differs from the neuronal 'adenosine mechanism' and probably also from the heart 'heat shock mechanism' for the acquisition of protection. PMID- 10863036 TI - Hyperventilation-induced human cerebral magnetic fields non-invasively monitored by multichannel 'direct current' magnetoencephalography. AB - Self-paced hyperventilation (HV) induces slow cerebral magnetic field changes which were monitored and mapped continuously over 15 min using 49-channel DC coupled ('direct current') magnetoencephalography (DC-MEG) based on a modulation technique. In nine/nine healthy subjects HV caused an increase (range: 1.1-6.2 pT) of the mean global DC-MEG field strength which slowly decayed after HV termination (mean time constant: 2 min). The complex HV-related field patterns were distinctly different from mainly dipolar somatosensory evoked field maps (N20m) in four/four subjects. Thus, current sources in the primary somatosensory cortex need not regularly dominate DC-field changes as had been previously considered. Rather, DC-MEG enabled the monitoring of a widely distributed HV induced enhanced cortical excitability which may serve as model to study epileptic or post-anoxic cerebral hyperexcitability. PMID- 10863037 TI - Cloning and characterization of a human brain Na(+)-independent transporter for small neutral amino acids that transports D-serine with high affinity. AB - We isolated a cDNA for the human homologue of system asc transporter Asc-1 from human brain. The encoded protein designated as hAsc-1 (human Asc-1) exhibited 91 % sequence identity to mouse Asc-1. Consistent with mouse Asc-1, hAsc-1 required 4F2 heavy chain for its functional expression in Xenopus oocytes. hAsc-1 exhibited the properties of amino acid transport system asc which transports small neutral amino acids in a Na(+)-independent manner. hAsc-1 transported D serine at high affinity with a K(m) value of 22.8 microM. In brain, 2.0 kb mRNA was highly expressed. hAsc-1 gene was mapped to human chromosome 19, region q12 q13.1. Because of the high-affinity transport with the K(m) value close to the physiological concentration of D-serine, together with the high levels of expression in brain, hAsc-1 is proposed to play significant roles in the D-serine mobilization in brain. PMID- 10863038 TI - The metabotropic glutamate receptor subtype mGluR 2/3 is located at extrasynaptic loci in rat spinal dorsal horn synapses. AB - The position of neurotransmitter receptors relative to active neurotransmitter release sites may be a major factor influencing neuronal responses. The location of the metabotropic glutamate receptor subtype mGluR2/3 was investigated in synaptic structures in the rat superficial spinal dorsal horn laminae by using a pre-embedding immunogold technique. Immunostaining for mGluR2/3 occurred in laminae I through III. Gold particles were encountered both in the cytosol and along the plasma membrane. Distinctive plasmalemmal immunodeposits were detected in vesicle-containing profiles, where they were located to membrane compartments distant from active release sites rather than in the close vicinity of synaptic specialisations. No distinct immunolabelling was observed in profiles meeting characteristics of primary afferent terminals. The extrasynaptic occurrence of mGluR2/3 suggests a presynaptic heteroreceptor role for these receptor subtypes in the spinal dorsal horn. PMID- 10863040 TI - Kinins in pain and inflammation. PMID- 10863041 TI - Systemic lidocaine for neuropathic pain relief. AB - The effectiveness of systemic lidocaine in relieving acute and chronic pain has been recognized for over 35 years. In particular, systemic lidocaine has been utilized both as a diagnostic and therapeutic tool for intractable neuropathic pain during the last decade. The introduction of oral lidocaine congeners such as mexiletine has significantly extended the usage of lidocaine therapy in chronic pain settings. However, a number of clinical issues remain to be addressed including (1) an effective, meaningful dose range for the clinical lidocaine test, (2) the predictive value of the lidocaine test for an oral trial of lidocaine congeners, (3) identification of pain symptoms and signs relieved by systemic lidocaine, (4) comparisons of therapeutic effects between systemic lidocaine and its oral congeners, and (5) long-term outcomes of systemic lidocaine and its oral congeners. Mechanisms of neuropathic pain relief from lidocaine therapy are yet to be understood. Both central and peripheral mechanisms have been postulated. Systemic lidocaine is thought to have its suppressive effects on spontaneous ectopic discharges of the injured nerve without blocking normal nerve conduction. However, there remain inconsistencies in the scientific basis underlying the clinical application of lidocaine therapy. Recent demonstration of changes in tetrodotoxin (TTX)-sensitive and TTX-resistant sodium channels following nerve injury and their link to certain neuropathic pain symptoms may lead to the development of subtype-specific sodium channel blockers. The thoughtful use of lidocaine therapy and the potential application of subtype specific sodium channel blockers could provide better management of distinctive neuropathic pain symptoms. PMID- 10863042 TI - Human chromaffin cell graft into the CSF for cancer pain management: a prospective phase II clinical study. AB - A number of pre-clinical studies have demonstrated the value of adrenal medullary allografts in the management of chronic pain. The present longitudinal survey studied 15 patients transplanted for intractable cancer pain after failure of systemic opioids due to the persistence of undesirable side-effects. Before inclusion, all the patients had their pain controlled by daily intrathecal (I-Th) morphine administration. The main evaluation criteria of analgesic activity of the chromaffin cell allograft was the complementary requirement of analgesics and in particular the consumption of I-Th morphine required to maintain effective pain control. Out of the 12 patients who profited from enhanced analgesia with long-term follow-up (average 4.5 months), five no longer required the I-Th morphine (with prolonged interruption of systemic opioids as well), two durably decreased I-Th morphine intake and five were stabilized until the end of their follow-up. Durable decline and stabilization were interpreted as indicative of analgesic activity by comparison with the usual dose escalation observed during disease progression. In most cases, we noted a relationship between analgesic responses and CSF met-enkephalin levels. The results of this phase II open study demonstrate the feasibility and the safety of this approach using chromaffin cell grafts for long-term relief of intractable cancer pain. However, while analgesic efficacy was indicated by the reduction or stabilization in complementary opioid intake, these observations will need to be confirmed in a controlled trial in a larger series of patients. PMID- 10863043 TI - Phantom pain and phantom sensations in upper limb amputees: an epidemiological study. AB - Phantom pain in subjects with an amputated limb is a well-known problem. However, estimates of the prevalence of phantom pain differ considerably in the literature. Various factors associated with phantom pain have been described including pain before the amputation, gender, dominance, and time elapsed since the amputation. The purposes of this study were to determine prevalence and factors associated with phantom pain and phantom sensations in upper limb amputees in The Netherlands. Additionally, the relationship between phantom pain, phantom sensations and prosthesis use in upper limb amputees was investigated. One hundred twenty-four upper limb amputees participated in this study. Subjects were asked to fill out a self-developed questionnaire scoring the following items: date, side, level, and reason of amputation, duration of experienced pain before amputation, frequencies with which phantom sensations, phantom pain, and stump pain are experienced, amount of trouble and suffering experienced, respectively, related to these sensations, type of phantom sensations, medical treatment received for phantom pain and/or stump pain, and the effects of the treatment, self medication, and prosthesis use. The response rate was 80%. The prevalence of phantom pain was 51%, of phantom sensations 76% and of stump pain 49%; 48% of the subjects experienced phantom pain a few times per day or more; 64% experienced moderate to very much suffering from the phantom pain. A significant association was found between phantom pain and phantom sensations (relative risk 11.3) and between phantom pain and stump pain (relative risk 1.9). No other factors associated with phantom pain or phantom sensations could be determined. Only four patients received medical treatment for their phantom pain. Phantom pain is a common problem in upper limb amputees that causes considerable suffering for the subjects involved. Only a minority of subjects are treated for phantom pain. Further research is needed to determine factors associated with phantom pain. PMID- 10863044 TI - Applying the American Pain Society's QA standards to evaluate the quality of pain management among surgical, oncology, and hospice inpatients in Taiwan. AB - The purpose of this study was two-fold: first, to apply the American Pain Society (APS) outcome questionnaire to examine and compare the quality of pain management provided in surgical, oncology, and hospice inpatient units in the Taipei area of Taiwan, and second, to provide baseline data of pain management quality in advance of the implementation of national guidelines for cancer pain management. Data revealed that hospice patients had significantly lower levels of pain severity and higher satisfaction with pain management than did oncology or surgical patients. A majority of patients reported that they received pain medication within 15 min after they complained of pain. However, a large number of patients never asked for pain medication during hospitalization. Moreover, most of the patients never requested medication changes even when their perception was that their medication were not effective. The findings of this study may provide support for the effectiveness of hospices in Taiwan in pain management and provide important information on the validity of the APS quality standards. PMID- 10863045 TI - Pain in children and adolescents: a common experience. AB - Little is known about the epidemiology of pain in children. We studied the prevalence of pain in Dutch children aged from 0 to 18 years in the open population, and the relationship with age, gender and pain parameters. A random sample of 1300 children aged 0-3 years was taken from the register of population in Rotterdam, The Netherlands. In the Rotterdam area, 27 primary schools and 14 secondary schools were selected to obtain a representative sample of 5336 children aged 4-18 years. Depending on the age of the child, a questionnaire was either mailed to the parents (0-3 years) or distributed at school (4-18 years). Of 6636 children surveyed, 5424 (82%) responded; response rates ranged from 64 to 92%, depending on the subject age and who completed the questionnaire. Of the respondents, 54% had experienced pain within the previous 3 months. Overall, a quarter of the respondents reported chronic pain (recurrent or continuous pain for more than 3 months). The prevalence of chronic pain increased with age, and was significantly higher for girls (P<0.001). In girls, a marked increase occurred in reporting chronic pain between 12 and 14 years of age. The most common types of pain in children were limb pain, headache and abdominal pain. Half of the respondents who had experienced pain reported to have multiple pain, and one-third of the chronic pain sufferers experienced frequent and intense pain. These multiple pains and severe pains were more often reported by girls (P<0.001). The intensity of pain was higher in the case of chronic pain (P<0. 001) and multiple pains (P<0.001), and for chronic pain the intensity was higher for girls (P<0.001). These findings indicate that chronic pain is a common complaint in childhood and adolescence. In particular, the high prevalence of severe chronic pain and multiple pain in girls aged 12 years and over calls for follow-up investigations documenting the various bio-psycho-social factors related to this pain. PMID- 10863046 TI - A survey of children's acute, recurrent, and chronic pain: validation of the pain experience interview. AB - The ultimate objective of our epidemiological research is to complete a longitudinal population-based study to document the prevalence and impact of acute, recurrent, and chronic pain in children and adolescents. As the first phase of our epidemiological research, we developed a comprehensive screening instrument for identifying children with acute, recurrent, and chronic pain, the Pain Experience Interview. We designed this interview to provide information about the lifetime and point prevalence of various pains, and also to provide information about the intensity, affect, duration, and frequency of children's pain. The primary objective of this study was to validate the Pain Experience Interview using the discriminant validation procedure of group differences. The secondary objectives of our study were to obtain descriptive data on children's acute, recurrent, and chronic pain experiences and to conduct exploratory analyses on age- and gender-related differences in children's pain experiences. We interviewed 187 children from five different health groups (arthritis, cancer, enuresis, recurrent headaches, and healthy) to provide distinct subsets of children with respect to their acute, recurrent, and chronic pain experience, and from four different age groups (5-7, 8-10, 11-13, and 14-16 years) to provide distinct subgroups with respect to children's developmental level. To test the interview we determined a priori several study predictions about children's pain experiences. These included four predictions about the common response patterns that we would expect to observe for all children based on our understanding of acute pain caused by trauma/disease, and six predictions about the distinct response patterns that we would expect to observe based on the known differences among children in their experiences of headache, acute treatment-related pain, recurrent pain, and chronic pain. All study predictions were confirmed, demonstrating that the Pain Experience Interview is a valid screening instrument for differentiating children with different types of pain problems. The interview can provide estimates for the lifetime and point prevalence of various pains in children, and data on the intensity, affect, duration, and frequency of their pain experiences. PMID- 10863047 TI - The multidimensional pain inventory profiles in patients with chronic cancer related pain: an examination of generalizability. AB - This study examined the generalizability of the non-malignant pain patient profiles based on the Multidimensional Pain Inventory (MPI) to patients with cancer-related pain. Data were collected from 112 cancer patients. In total, 107/112 patients completed the MPI. Of the 96% of patients classified, only 60% were classified by the three main profiles. In this sample, there were 47.7% (n=51) Adaptive Copers, 9.3% (n=10) Dysfunctional, 2.8% (n=3) Interpersonally Distressed; 32.7% (n=35) Anomalous; 3.8% (n=4) Hybrid; and 3.8% (n=4) Unanalyzable. Because of the significantly lower pain severity, interference and affective distress scores, the Anomalous group could be considered Highly Adaptive. Given that 80% were classified as either Adaptive or Anomalous, these findings suggest that while the MPI-based profiles do apply, a two profile classification system may be more suitable for cancer patients than the usual three. In particular, the low proportion of patients classified as Interpersonally Distressed may reflect important differences in social support for cancer patients compared with non-cancer patients. Whereas the MPI-based profiles are consistent across non-malignant pain problems, it appears that the nature of cancer may affect the MPI-based profile classification system more than non-malignant pain problems do. PMID- 10863048 TI - Evidence for ascending visceral nociceptive information in the dorsal midline and lateral spinal cord. AB - The effect of acute, mid-cervical spinal cord lesions on neuronal and reflex activity evoked by the noxious visceral stimulus, colorectal distension (CRD; 80 mmHg, 20 s), was determined in halothane-anesthetized rats. Extracellular recordings were performed of neurons stereotaxically located within the ventrobasal group of the thalamus and in the region of the medullary lateral reticular nucleus. CRD-evoked activity of thalamic neurons was attenuated by lesions of the dorsal midline, but minimally affected by lateral lesions of the spinal cord. In contrast, CRD-evoked activity of medullary neurons was attenuated by lateral lesions ipsilateral to the recording site, but minimally affected by contralateral lateral lesions or dorsal midline lesions. Pseudo-affective visceromotor/cardiovascular responses were vigorous in rats with dorsal midline lesions and absent/attenuated in rats with bilateral lateral spinal lesions. This study presents evidence that visceral nociceptive information ascends in the spinal cord by both dorsal midline and lateral spinal pathways. PMID- 10863049 TI - A validated, practical classification procedure for many persistent low back pain patients. AB - We have developed a simple procedure for assigning persistent low back pain patients to one of four mutually exclusive, hierarchically organized classes. The procedure relies on the spatial distribution of a patient's pain and the results of straight leg raise tests to make the assignment. We have applied the procedure to a large group of patients who sought treatment for persistent LBP at several university affiliated tertiary care clinics, and found that the resulting four classes of patients were significantly different from one another in their presentation, and in the way they were evaluated and treated by physicians. We concluded that the procedure may have practical research and clinical applications. PMID- 10863050 TI - Spinal cord stimulation: a possible therapeutic alternative for chronic mesenteric ischaemia. AB - A 78-year-old male patient had chronic, unrelieved abdominal pain due to mesenteric ischaemia. Unsuccessful pharmacological approaches included oral morphine plus coadjuvants as well as a sympathetic celiac plexus block which gave pain relief that lasted for 72 h. In order to obtain long-lasting relief, a trial epidural stimulating electrode was implanted after obtaining informed consent and Ethical Committee approval. Complete analgesia was achieved during a trial period of 2 weeks. Thereafter, a spinal cord stimulator was implanted. At the time of writing, 11 months after implantation, the degree of analgesia is complete. We believe that spinal cord stimulation may represent an alternative approach in controlling pain due to mesenteric ischaemia. PMID- 10863051 TI - Long-term benefits of stellate ganglion block in severe chronic refractory angina. AB - Angina pectoris that is refractory to optimal medication and revascularization is becoming an increasingly common clinical problem. Recently the US Food and Drug Administration (FDA) approved transmyocardial laser revascularization (TMLR) for use in this group of patients and a large numbers of patients have already undergone this therapy. Unfortunately TMLR has is associated with an unacceptably high perioperative mortality (Cooley DA, Frazier OH, Kadipasaoglu KA, Lindenmeir MH, Pehlivanoglu S, KoIff JW, Wilansky S, Moore WH. Transmyocardiai laser revascularisation: clinical experience with twelve-month follow-up. J Thorac Cardiovasc Surg 1996;111:791-799; Horvath KA, Cohn LH, Cooley DA, Crew JR, Frazier GH, Griffith BP, Kadipasaoglu K, Lansing A, Mannting F, March R, Mirhoseini MR, Smith C. Transmyocardial laser revascularisation: results of a multi-centre transmyocardial laser revascularisation used as sole therapy for end stage coronary artery disease. J Thorac Cardiovasc Surg 1997;113:645-654; Schofield PM, Sharples LD, Caine N, Burns S, Tait S, Wistow T, Buxton M, Wallwork J. Transmyocardial laser revascularisation in patients with refractory angina: a randomised controlled trial. Lancet 1999;353:519-524), and recurrent refractory angina is common (Allen KB, Dowling RD, Fudge TL, Schoettle GP, Selinger SL, Gangahar OM, Angell WW, Petracek MR, Shaar CJ, O'Neill WW. Comparison of transmyocardial revascularization with medical therapy in patients with refractory angina. N Engl J Med 1999;341:1021-1028; Frazier OH, March RJ, Horvath KA, for the Transmyocardial Carbon Dioxide Laser Revascularization Study Group. Transmyocardial revascularization with a carbon dioxide laser in patients with end-stage coronary artery disease. N Engl J Med 1999;341:1021-1028). Temporary sympathectomy by stellate ganglion block (SGB) is in widespread use in a variety of chronic pain conditions and has long history of use in the management of angina (Moore DC. Stellate ganglion block. Springfield, IL: CC Thomas, 1954; Wiener L, Cox JW. Influence of stellate ganglion blockade on angina pectoris and the post exercise electrocardiogram. Am J Med Sci 1966;252:289-295). Here we describe a patient with end stage coronary artery disease and chronic refractory angina whose has been successfully treated with repeated unilateral left SGBs following multiple bypass operations, angioplasty procedures and laser therapy. This case report details his progress over a 34 month follow-up period. PMID- 10863052 TI - Erythema ab igne: an unusual manifestation of cancer-related pain. AB - We report two patients with truncal erythema ab igne who were subsequently diagnosed as having an underlying malignancy. The skin changes were due to repeated heat applications for pain relief. Although erythema ab igne is frequently seen on the anterior lower limbs of elderly people, it may be an indicator of organic disease when present in a central location in young patients. PMID- 10863053 TI - Improved outcome secondary to concurrent chemoradiotherapy for advanced carcinoma of the nasopharynx: preliminary corroboration of the intergroup experience. AB - PURPOSE: The recent Intergroup 0099 trial of concurrent chemoradiotherapy for advanced nasopharyngeal carcinomas, demonstrated improved survival for chemoradiotherapy as compared to radiation therapy alone. Following closure of this study, we adopted the chemoradiotherapy regimen used in 0099 as our standard of practice. We herein report our recent institutional results, representing a relatively large uniformly treated cohort. METHODS AND MATERIALS: Between 1995 and 1997, 35 consecutive patients, who had clinically nondisseminated Stage III or IV nasopharyngeal cancer, were treated by chemoradiotherapy. The prescribed radiation regimen was 7000 cGy delivered in 35 fractions over 7 weeks to all macroscopic disease and 5000 cGy to areas considered at risk of harboring microscopic disease. Chemotherapy was designed to deliver cisplatin (100 mg/m(2) i.v.) on Days 1, 22, and 43 of radiation therapy and cisplatin (80 mg/m(2) i.v.) on Days 71, 99, and 127 plus flurouracil (5-FU; 1 g/m(2)/day by 96-h infusion) on Days 71-74, 99-102, and 127-130. RESULTS: All patients had at least a partial response (PR) to treatment, including an 85% complete response (CR) rate. The actuarial 3-year overall survival rate was 93% and the disease-free survival rate was 65%. Both represent substantial improvements over our institutional historical controls treated by radiation therapy alone and both are similar to the rates observed in the Intergroup trial. CONCLUSION: Our data support the conclusion that concurrent chemoradiotherapy followed by adjuvant chemotherapy (as was used in Intergroup 0099) should be considered the current standard of care for patients who have advanced cancers of the nasopharynx. PMID- 10863054 TI - The effect of nodal status on determinants of initial treatment response and patterns of relapse-free survival in nasopharyngeal carcinoma. AB - PURPOSE: To study the effect of regional nodal status on predictors of treatment response, failure patterns, and the time-dependent nature of the various pattern of relapse via a hazard function analysis. METHODS AND MATERIALS: We reviews tumor control data of 496 patients with nasopharyngeal carcinoma (NPC) to whom a radical course of radiotherapy (RT) with or without induction chemotherapy (CT) was given. All alive patients had a median follow-up period of 131 months. Primary tumor (T) and nodal (N) status were staged according to the TNM system of the American Joint Committee. Remote after-loading brachytherapy may be added to teletherapy in T1-2 lesions while induction CT could be given for N3 and/or T4 lesions. Hazard function analysis over 1-year interval was carried out for locoregional or distant relapse. RESULTS: T stage and brachytherapy were two independent predictors for complete response (CR) at the primary site irrespective of nodal status, whereas N stage and brachytherapy are major determinants for regional CR in node (+) patients. Multivariate analysis revealed that contributors to a relatively long disease-free interval in (1) node (-) patients were for locoregional relapse, induction CT(-) (p = 0.0062) or brachytherapy (+) (p = 0.0268) and for distant relapse, none; (2) node (+) patients were for locoregional relapse, early T stage (p = 0.0377) or regional CR (p = 0.0075) and for distant relapse, induction CT(-) (p = 0.0001) or regional CR (p = 0.0001). In node (-) or (+) patients, primary CR rate yield no independent prognostic value on various types of disease-free survival. Hazard function analysis for relapse revealed that hazard rates are in general negatively correlated with time, being highest at the first year post-treatment, decreasing from time to time, and approaching zero after a longer follow-up period in patients with locoregional CR than in patients without. CONCLUSION: Nodal status had no significant impact on predictors of primary CR, whereas in node (+) patients regional CR rate had an independent value in predicting disease-free survival to locoregional and distant relapse. Hazard function analysis revealed a decreasing hazard rate over a protracted post-treatment time in primary and regional CR patients. This indicates the continued risk of late recurrence in this subset of patients for whom long-term observation is recommended. PMID- 10863055 TI - Quality of life and utility in irradiated laryngeal cancer patients. AB - PURPOSE: To determine quality of life (QOL) and health utility in irradiated laryngeal cancer survivors. MATERIALS AND METHODS: Over 6 months, consecutive follow-up patients at a comprehensive cancer centre completed the QOL questionnaire FACT-H&N and the time trade-off (TTO) utility instrument. RESULTS: Inclusion criteria were met by 339 patients, of whom 269 were eligible, 245 were approached, and 120 agreed to participate. Most participants were men (83%) who had received radiotherapy (97%) for Stage I disease (53%) of the glottis (75%); 7% had undergone total laryngectomy. Participants differed from nonparticipants only in being younger (mean age, 65 vs. 68 years, p = 0.0049) and having higher performance status (Karnofsky 88 vs. 84, p = 0.0012). The average scores for FACT H&N and the TTO were 124/144 (SD, 14) and 0.90/1.0 (SD, 0.16) respectively. FACT H&N score was more highly correlated with Karnofsky score (r = 0.43, p = 0.001) than with the TTO (r = 0.29, p = 0.002). Gender predicted QOL (means: M = 125, F = 118), while natural speech, no relapses, and more time since initial treatment predicted higher utility. CONCLUSION: The QOL of irradiated laryngeal cancer survivors was reasonably high and independent of initial disease variables. The QOL questionnaire correlated more strongly with performance status than with utility, suggesting that QOL and utility measures may be perceived differently by patients. PMID- 10863056 TI - Internal mammary node irradiation neither decreases distant metastases nor improves survival in stage I and II breast cancer. AB - PURPOSE: To compare outcome for ipsilateral breast tumor recurrence (IBTR), or regional node recurrence, initial and subsequent distant metastases, and overall and cause-specific survival in women treated with conservative surgery and radiation based on whether or not radiation was targeted to the internal mammary nodes (IMN). METHODS AND MATERIALS: From 1979-1994, 1383 women with Stage I-II breast cancer underwent wide excision, axillary node dissection with >/=10 nodes removed, and radiation. Median follow-up was 6 years; median age was 55 years. A total of 114 women had radiation targeted to the IMN with deep tangents and 1269 did not. Women who received IMN treatment were more often axillary node-positive (40% vs. 25%, p = 0. 002), had central or inner quadrant tumors (61% vs. 40%, p = 0.001), and had T2 tumors (47% vs. 31%, p = 0.001). All axillary node-positive women received adjuvant chemotherapy and/or tamoxifen. For axillary node-negative women, 13% of the IMN treatment group received adjuvant systemic therapy compared to 37% of the no treatment group (p = 0.001). Radiation was directed to the breast only in 97% of the axillary node-negative women who had IMN treatment and 99% of the no IMN treatment group. For axillary node-positive women, 98% of the IMN-treated group had radiation to the breast and supraclavicular nodes +/- a posterior axillary field compared to 77% of the no IMN treatment group (p = 0.001). There were no significant differences between the two groups for median age, menopausal status, histology, final surgical margin, estrogen and progesterone receptor status, or the number of positive nodes. RESULTS: There were no significant differences in the 5- and 10-year cumulative incidence of an IBTR, regional node recurrence, initial or total distant metastases for the two groups. Similarly 5- and 10-year actuarial overall and cause-specific survival were not significantly different. However, subset analysis revealed a statistically significant increase in initial (29% vs. 15% at 10 yr, p = 0.002) and total (30% vs. 17% at 10 yr, p = 0.01) distant metastases and a significant decrease in cause-specific survival (76% vs. 89% at 10 yr, p = 0.02) for postmenopausal women who received IMN treatment. These findings could not be attributed to differences in the use of systemic therapy or the number of positive nodes. Axillary node-positive patients did not experience a significant decrease in initial (36% vs. 22% at 10 yr, p = 0.21) or total distant metastases (37% vs. 28% at 10 yr, p = 0.62) or a significant improvement in cause-specific survival (72% vs. 76% at 10 yr, p = 0.76) with IMN treatment regardless of whether the tumor was lateral or medial/central in location. IMN treatment was not associated with an increase in non-breast cancer deaths during this period of observation. CONCLUSIONS: This retrospective series was unable to identify a significant benefit for IMN irradiation in terms of distant metastases or cause specific survival for the entire patient population, and in particular, for patients with positive axillary nodes and medially located lesions. The results of the proposed or ongoing prospective randomized trials will further address this controversial issue. PMID- 10863057 TI - Reduction of cardiac volume in left-breast treatment fields by respiratory maneuvers: a CT study. AB - PURPOSE: A previous study of healthy female volunteers suggested that deep inspiratory breath holding can reduce the cardiac volume in the treatment portals for left-breast cancer treatment. The reduction of irradiated cardiac volume may be important considering the reported late cardiac morbidity and mortality and the frequent coexistent use of potentially cardiotoxic chemotherapy in breast cancer patients. In the present study, we evaluated the heart volume in the fields and, thus, the true benefit of this respiratory maneuver in breast cancer patients undergoing CT simulation. MATERIALS AND METHODS: Fifteen patients (median age, 53) were studied. For each patient, CT scans were performed both when the patient breathed normally (quiet respiration) and when the patient held her breath after a deep inspiration. Tangential fields were planned using the same medial, lateral, superior, and inferior borders on skin for the normal breathing and the breath-holding configurations. The cardiac and left-lung volumes within the tangential fields were calculated for both breathing configurations. Multiple scan series were performed for the breath-holding configuration to provide a more accurate delineation of the cardiac tissue and to study the reproducibility of the patient's position between different cycles of deep inspiration. RESULTS: None of the patients had difficulty holding her breath for 20 s. The cardiac volume in the field was reduced (-86 +/- 24%; p < 0.001) when patients held their breath after a deep inspiration compared to when breathing normally. For 7 patients (47%), deep inspiration moved the heart completely out of the radiation fields. The expansion of the lung tissue due to deep inspiration also increased the absolute lung volume in the tangential fields (183 cm(3) vs 97 cm(3), p < 0.001). However, the fractional volume of the left lung in the field was essentially unchanged. For all but 1 patient, the maximum difference between the external body contours from different breath holding cycles was 5 mm and occurred at the lateral aspect of the breast. At the medial aspect, as indicated by the position of the midline marker, the variations were well within the currently accepted tolerance for patient positioning during tangential treatment. CONCLUSIONS: Deep-inspiration breath holding substantially reduces cardiac volume in the tangential fields for left-sided breast cancer treatment. The variation between patient positions at different cycles of breath holding was found to be reasonably small. Therefore, it appears feasible to reduce cardiac radiation by treating patients with intratreatment minifractions lasting 10-15 s while patients hold their breath. PMID- 10863058 TI - Identifying the predictors of acute urinary retention following magnetic resonance-guided prostate brachytherapy. AB - PURPOSE: Larger prostate gland volumes have been associated with long-term urinary morbidity in prostate interstitial radiation therapy utilizing ultrasound image guidance technique. This study was performed to identify the clinical and technical predictors of acute urinary retention following magnetic-resonance (MR) guided prostate interstitial brachytherapy. METHODS AND MATERIALS: Fifty patients underwent MR-guided prostate brachytherapy between December 1997 and March 1999. Patient selection was limited to men with stage T1cNXM0 disease, PSA of less than10 ng/mL, biopsy Gleason score not more than 3 + 4, and endorectal coil MR stage T2 disease. Dosimetry plans were developed in the operating room and (125)Iodine sources were implanted using MR real-time guidance. The peripheral zone (PZ) of the prostate gland was defined as the clinical target volume (CTV) and the minimum prescribed dose to the CTV was 137 Gy. The volumes of the PZ, transition zone (TZ), and total prostate gland volume were also determined by MR. Individual source strength ranged from 0.35 to 0.54 microGym(2)/h (NIST 99, median 0.46 microGym(2)/h) and the total implanted activity ranged from 17.0 to 43.1 mCi (median, 28.1 mCi) using 43-120 seeds (median, 79). The seeds were placed using MR-compatible biopsy needles (14-28, median, 19). RESULTS: The ability of clinical (MR defined prostate, PZ, and TZ volumes) and technical (number of catheters, number of seeds implanted, and total activity) factors to predict AUR for 50 men undergoing MR-guided prostate interstitial brachytherapy were evaluated using univariable and logistic regression multivariable analyses. Six men (12%) experienced AUR within 24 h after removal of the Foley catheter subsequent to prostate brachytherapy. The total number of seeds (p = 0.05), MR determined prostate volume (p < 0.01), and the MR-determined TZ volume (p < 0.01) were significant predictors of AUR on univariable analysis. Utilizing a multivariable logistic regression analysis, the TZ volume was the only significant predictor of AUR (p < 0.01). The prostate volume is highly correlated to the TZ volume (Spearman correlation coefficient of 0. 91) and was thus significant in the univariable analysis; however, the prostate volume did not add prognostic value in multivariable analysis. CONCLUSION: Benign prostatic hyperplasia (BPH) resulting in an enlarged TZ volume, is the most important predictor of AUR following MR-guided prostate interstitial radiation therapy. Although AUR was significant (60%) in men with moderate BPH (TZ volume >/= 50 cc), it was also self-limiting. PMID- 10863059 TI - A comparison of complications between ultrasound-guided prostate brachytherapy and open prostate brachytherapy. AB - PURPOSE: Prostate brachytherapy has reemerged during the 1990s as a treatment for clinically localized prostate cancer. The renewed popularity of prostate brachytherapy is largely due to the use of transrectal ultrasound of the prostate, which allows for more accurate isotope placement within the prostate when compared to the open approach. The present study investigates whether this improved cancer control is at the expense of increased morbidity by comparing the morbidity after transrectal ultrasound-guided prostate brachytherapy to the morbidity after prostate brachytherapy performed via an open approach. METHODS AND MATERIALS: All men in the Medicare population who underwent prostate brachytherapy in the year 1991 were identified. These men were further stratified into those men who underwent prostate brachytherapy via an open approach and the men who underwent prostate brachytherapy with ultrasound guidance. All subsequent inpatient, outpatient, and physician (Part B) Medicare claims for these men from the years 1991-1993 were then analyzed to determine outcomes. RESULTS: In the year 1991, 2124 men in the Medicare population underwent prostate brachytherapy. An open approach was used in 715 men (33.7%), and ultrasound guidance was used in 1409 men (66.3%). Mean age for both cohorts was 73.7 years with a range of 50.7 92.8 years for the ultrasound group and 60.6-92. 1 years for the open group. A surgical procedure for the relief of bladder outlet obstruction was performed in 122 men (8.6%) in the ultrasound group and in 54 men (7.6%) in the open group. An artificial urinary sphincter was placed in 2 men (0.14%) in the ultrasound group and in 2 men (0.28%) in the open group. A penile prosthesis was implanted in 10 men (0.71%) in the ultrasound group and in 4 men (0.56%) in the open group. A diagnosis code for urinary incontinence was carried by 95 men (6.7%) in the ultrasound group and by 45 men (6.3%) in the open group. A diagnosis code for erectile dysfunction was carried by 90 men (6.3%) in the ultrasound group and by 64 men (9.0%) in the open group. CONCLUSION: Prostate brachytherapy performed with ultrasound guidance does not appear to increase significantly complications resulting from the procedure. Both techniques appear to offer similar rates of procedures performed to correct urinary incontinence, bladder outlet obstruction and erectile dysfunction. The limitations of claim information in determining patient outcomes, however, must be considered when evaluating this data. PMID- 10863060 TI - Chronic effects of therapeutic irradiation for localized prostatic carcinoma on anorectal function. AB - PURPOSE: To evaluate prospectively the prevalence and pathophysiology of anorectal dysfunction following radiation therapy (RTH) for localized carcinoma of the prostate. METHODS AND MATERIALS: The following parameters of anorectal function were evaluated in each of 35 patients (aged 55-82 years) with localized prostatic carcinoma treated with RTH either to a dose of 55 Gy/20 fractions/4 weeks (18 patients) or 64 Gy/32 fractions/6.5 weeks (17 patients), before RTH and 4-6 weeks and at a mean (+/- SD) of 1.4 (+/- 0.2) years after its completion: (1) anorectal symptoms (questionnaire), (2) anorectal pressures at rest and in response to voluntary squeeze and increases in intra-abdominal pressure (multiport anorectal manometry), (3) rectal sensation (balloon distension) and (4) anal sphincteric morphology (endoanal ultrasound). RESULTS: All but 1 patient completed three series of measurements. RTH had no effect on anal sphincteric morphology. The increase in frequency of defecation and fecal urgency and incontinence scores previously reported in the patients 4-6 weeks after RTH were sustained 1 year later (p < 0.001, p < 0.001, and p < 0.05, cf. baseline, respectively). At this time, 56% (19 of 34), 50% (17 of 34) and 26% (9 of 34) of the patients had increased frequency of defecation, fecal urgency, and incontinence, respectively. Decreases in anal sphincteric pressures at rest and in response to voluntary squeeze recorded in the patients 4-6 weeks after RTH were not sustained 1 year later but the volumes of rectal distension associated with perception of the stimulus and desire to defecate were lower compared with baseline volumes (p < 0.01 and p < 0.05, respectively), reflecting heightened rectal sensitivity in the patients. There was no difference in measurements between the two radiation dose regimens. Univariate logistical regression analysis was performed on patients who had experienced increased symptom scores or decreases in recorded motor and sensory manometric parameters at 1 year, cf. baseline. The predictor variables used included individual patient tumor and treatment characteristics as well as individual patient symptom scores and parameters of anorectal motor and sensory function at baseline and 4-6 weeks after RTH. The results of the univariate logistical regression analysis showed that (1) frequency of defecation at 4-6 weeks and (2) rectal volumes at baseline both for (a) perception (p < 0.001) and (b) desire to defecate (p < 0.001), predicted significantly for the patients who had symptoms and signs of anorectal dysfunction at 1 year. Individual patient tumor and treatment-related variables tested, in contrast, had no predictive significance. CONCLUSIONS: Anorectal symptoms following RTH for prostatic carcinoma are common and persist at least until 1 year after its completion and are associated with objective evidence of heightened rectal sensitivity. PMID- 10863061 TI - A study of postoperative radiotherapy in patients with non-small-cell lung cancer: a randomized trial. AB - PURPOSE: To study the value of postoperative radiotherapy for non-small-cell lung cancer (NSCLC) with positive regional lymph metastases (NI or N2) after radical surgery. MATERIALS AND METHODS: From February 1982 to October 1995, 366 patients with NSCLC and N1 or N2 disease were randomized into postoperative radiotherapy (S + R) (183 patients) and no further treatment (S alone) (182 patients). Postoperative radiotherapy (RT) was administrated 3-4 weeks after radical operation. Irradiated fields covered the bronchial stump, ipsilateral hilum, and most of the mediastinum. The midplane dose was 6000 cGy/30 fractions/6 weeks, with the spinal cord limited to 4000 cGy/20 fractions/4 weeks or less. One hundred thirty-four patients in S + R group and 162 patients in S alone group were evaluated. Clinical data were comparable in both arms, except for the numbers of N2 patients. RESULTS: The 3-year and 5-year overall survival rates were 51.9% and 42.9% in the S + R group and 50.2% and 40.5% in the S alone Group (p = 0.56). The 3-year and 5-year disease-free survival rates were 50.7% +/- 4.7% and 42.9% +/- 5.2% in the S + R group vs. 44.4% +/- 4.3% and 38.2% +/- 4.5% in the S alone group (p = 0.28), respectively. In the patients with NI or T3-4 tumors, there was a trend toward improved survival in the S + R group, especially in the patients with T3-4N1M0. These patients demonstrated 20% improvement in overall survival (p = 0.092) and greater than 20% better disease-free survival (p = 0.057). Postoperative RT reduced local recurrence but had no impact on distant metastases. CONCLUSION: Postoperative RT significantly reduced local relapses, but did not improve overall survival, due to a high frequency of distant metastases in this patient group. PMID- 10863062 TI - A phase II study of paclitaxel, carboplatin, and hyperfractionated radiation therapy for locally advanced inoperable non-small-cell lung cancer (a Vanderbilt Cancer Center Affiliate Network Study). AB - PURPOSE: We conducted a prospective phase II study to determine the response rate, toxicity, and survival rate of concurrent weekly paclitaxel, carboplatin, and hyperfractionated radiation therapy (paclitaxel/carboplatin/HFX RT) followed by 2 cycles of paclitaxel and carboplatin for locally advanced unresectable non small cell lung cancer (NSCLC). The weekly paclitaxel and carboplatin regimen was designed to optimize the radiosensitizing properties of paclitaxel during the concurrent phase of treatment. METHODS AND MATERIALS: Forty-three patients with unresectable stage IIIA and IIIB NSCLC from the Vanderbilt Cancer Center and Affiliate Network (VCCAN) institutions were entered onto the study from June 1996 until May 1997. Weekly intravenous (IV) paclitaxel (50 mg/m(2)/l-hour) and weekly carboplatin (AUC 2) plus concurrent hyperfractionated chest RT (1.2 Gy/BID/69.6 Gy) were delivered for 6 weeks followed by 2 cycles of paclitaxel (200 mg/m(2)) and carboplatin (AUC 6). RESULTS: Forty-two patients were evaluable for response and toxicities. Three patients achieved a complete response (7.2%) and 30 patients achieved a partial response (71.4%), for an overall response rate of 78.6% [95% C.I. (66.2%-91.0%)]. The 1- and 2-year overall and progression-free survival rates of all 43 patients were 61.6% and 35% respectively, with a median survival time of 14.3 months. The median follow-up time was 14 months. Esophagitis was the principal toxicity. Grade 3 or 4 esophagitis occurred in 11 patients (26%). There was an incidence of 7% grade 3 and 9.5% grade 4 pulmonary toxicities. CONCLUSIONS: Weekly paclitaxel, carboplatin, plus concurrent hyperfractionated RT is a well-tolerated outpatient regimen. The response rate from this regimen is encouraging and appears to be at least equivalent to the more toxic chemoradiation trials. These findings warrant further clinical evaluation of weekly paclitaxel/carboplatin/HFX RT in a phase III study. PMID- 10863063 TI - A phase I study of combined UFT plus leucovorin and radiotherapy for pancreatic cancer. AB - PURPOSE: This Phase I study combines tegafur and uracil (UFT) with leucovorin and conventional radiation for the treatment of pancreatic cancer. The design seeks to determine the maximum tolerated dose (MTD) and dose-limiting toxicities (DLT) of this regimen as well as to define a future Phase II dose level. METHODS: Patients with locally advanced and unresectable pancreatic cancer were treated with 45 Gy of radiation therapy. The initial UFT dose was 150 mg/m(2)/day given with leucovorin 90 mg/day, both divided into 3 daily doses for 35 days concurrent with radiation. UFT doses were escalated at increments of 50 mg/m(2)/day. Dose limiting toxicity (DLT) was defined as Grade 3 or greater nausea, vomiting or diarrhea despite medical intervention; or Grade 3 or greater neutropenia/thrombocytopenia; or Grade 3 or greater hepatic toxicity; or inability of the patient to take 75% or more of the planned UFT/leucovorin; or radiotherapy interruption of greater than 1 week. The MTD for UFT/leucovorin was exceeded by one dose level when a certain dose caused DLT in 2 or more patients of 6. RESULTS: Five evaluable patients had Stage I resectable disease but had pathologic adenopathy. Seven had Stage II unresectable disease. Compliance with therapy was excellent. At a daily dose of 300 mg/m(2) of UFT, we noticed minimal diarrhea and hematologic toxicity with mild-moderate nausea, anorexia, and fatigue. Three patients had Grade 4 toxicity: 1 had neutropenia on Day 38, 1 had diarrhea on Day 55, and 1 had vomiting on Day 15. CONCLUSION: Oral UFT/leucovorin and radiation therapy offers patients a viable treatment option for pancreatic cancer. The major known toxicity of diarrhea was tolerable. The MTD was not reached in this study. Our current plan is to expand this into a Phase I/II trial beginning at a UFT dose of 300 mg/m(2) and correlate this with clinical pharmacologic parameters. The potential benefit of long bioavailability and oral delivery of UFT compares favorably with continuous infusion regimens without the added morbidity of a catheter and pump. PMID- 10863064 TI - Outcome of pancreaticoduodenectomy and impact of adjuvant therapy for ampullary carcinomas. AB - PURPOSE: To determine the clinical outcomes and potential impact of adjuvant chemoradiation in patients undergoing surgical resection of ampullary carcinoma. PATIENTS AND METHODS: Between 1988 and 1997, 39 patients underwent pancreaticoduodenectomy for ampullary adenocarcinomas. Clinical and pathologic factors, adjuvant therapy records, and disease status were obtained from chart review. Thirteen (33%) patients received adjuvant chemoradiation. Radiation therapy was delivered to the surgical bed and regional nodes to a median dose of 4,860 cGy with concurrent bolus or continuous infusion of 5-fluorouracil. Outcomes measures included locoregional control, disease-free survival, and overall survival. Univariate analysis was used to assess the impact of various patient- and tumor-related factors and the use of adjuvant therapy. Twenty (51%) patients with tumor invasion into the pancreas (T3) or node-positive disease were classified in a "high-risk" subgroup. RESULTS: After a median follow-up of 45 months for survivors, overall 3-year survival was 55%. Survival was significantly worse for patients with positive nodes (23% vs. 73%, p < 0.001) and high-risk status (30% vs. 80%, p = 0.002). Disease-free survival was 54% at 3 years. There were 3 postoperative deaths, and these patients (all high risk) are excluded from further analysis on adjuvant therapy. In univariate analysis, the use of adjuvant chemoradiation had no clear impact on local-regional control or overall survival. However, by controlling for risk status in multivariate analysis, the use of adjuvant therapy reached statistical significance for overall survival (p = 0. 03). Among the high-risk patients, 7 (77%) of 9 patients receiving adjuvant therapy remained disease-free during follow-up compared with only 1 (14%) of 7 patients not receiving adjuvant therapy (p = 0.012). CONCLUSION: Despite the relatively favorable prognosis of ampullary carcinomas compared with other pancreaticobiliary tumors, patients with nodal metastases or T3 disease are at high risk for disease relapse. The use of adjuvant chemoradiation may improve long-term disease control in these patients. PMID- 10863065 TI - The prediction of late rectal complications following the treatment of uterine cervical cancer by high-dose-rate brachytherapy. AB - PURPOSE: This study aimed to correlate patient, treatment, and dosimetric factors with the risk of late rectal sequelae in patients with uterine cervical cancer treated with external beam radiation therapy (EBRT) and high dose rate intracavitary brachytherapy (HDRICB). METHODS AND MATERIALS: From September 1992 to December 1995, a total of 128 patients with uterine cervical cancer, who were treated and survived more than 12 months, were evaluated. After EBRT with 40-44 Gy/20-22 Fr/4-5 weeks to the whole pelvis, the dose was boosted up to 54-58 Gy with central shielding for patients with bilateral parametria of Stage IIb or greater. HDRICB consisted of three to four insertions at doses of 5-7.2 Gy (to Point A) at intervals of 1 week. Patient and treatment factors were analyzed using logistic regression analysis and the cumulative rectal biologic equivalent dose (CRBED) was calculated. RESULTS: After 30-75 months of follow-up (median, 43 months), 38 patients (29.7%) had late rectal sequelae. Patients who had Stage IIb IVa disease, cumulative rectal dose (external RT + total ICRU rectal dose) greeater than 65 Gy, or age greater than 70 years had a high risk of developing late rectal sequelae. When 110 Gy was used as the cut-off value, 19.6% (10 of 51) of patients whose CRBED was less than 110 Gy had rectal complications, while 36.4% (28/77) of patients whose CRBED was greater than 110 Gy developed rectal complications. CONCLUSION: Risk factors of late rectal complications were advanced stage, age greater than 70 years, and cumulative rectal dose of greater than 65 Gy. PMID- 10863066 TI - Modeling normal tissue complication probability from repetitive computed tomography scans during fractionated high-dose-rate brachytherapy and external beam radiotherapy of the uterine cervix. AB - PURPOSE: To calculate the normal tissue complication probability (NTCP) of late radiation effects on the rectum and bladder from repetitive CT scans during fractionated high-dose-rate brachytherapy (HDRB) and external beam radiotherapy (EBRT) of the uterine cervix and compare the NTCP with the clinical frequency of late effects. METHODS AND MATERIALS: Fourteen patients with cancer of the uterine cervix (Stage IIb-IVa) underwent 3-6 (mean, 4.9) CT scans in treatment position during their course of HDRB using a ring applicator with an Iridium stepping source. The rectal and bladder walls were delineated on the treatment-planning system, such that a constant wall volume independent of organ filling was achieved. Dose-volume histograms (DVH) of the rectal and bladder walls were acquired. A method of summing multiple DVHs accounting for variable dose per fraction were applied to the DVHs of HDRB and EBRT together with the Lyman Kutcher NTCP model fitted to clinical dose-volume tolerance data from recent studies. RESULTS: The D(mean) of the DVH from EBRT was close to the D(max) for both the rectum and bladder, confirming that the DVH from EBRT corresponded with homogeneous whole-organ irradiation. The NTCP of the rectum was 19.7% (13.5%, 25. 9%) (mean and 95% confidence interval), whereas the clinical frequency of late rectal sequelae (Grade 3-4, RTOG/EORTC) was 13% based on material from 200 patients. For the bladder the NTCP was 61. 9% (46.8%, 76.9%) as compared to the clinical frequency of Grade 3-4 late effects of 14%. If only 1 CT scan from HDRB was assumed available, the relative uncertainty (standard deviation or SD) of the NTCP value for an arbitrary patient was 20-30%, whereas 4 CT scans provided an uncertainty of 12-13%. CONCLUSION: The NTCP for the rectum was almost consistent with the clinical frequency of late effects, whereas the NTCP for bladder was too high. To obtain reliable (SD of 12-13%) NTCP values, 3-4 CT scans are needed during 5-7 fractions of HDRB treatments. PMID- 10863067 TI - Modified partial hyperfractionation in radiotherapy for bulky uterine cervical cancer: reduction of overall treatment time. AB - PURPOSE: The purpose of this study was to assess the feasibility and toxicity of modified fractionation of external beam radiation with the intention of reducing the overall treatment time (OT) by 1 week in cervical cancer. METHODS AND MATERIALS: Thirty-one patients (Group 1, n = 31) with bulky cervical cancer (>/= 4 cm with Stage II and III, >/= 5 cm with Stage IB2) were entered into the twice a day (b.i. d.) protocol (18 Gy/10 fx in 2 weeks followed by 18 Gy/12 fx, b.i.d. in 6 days, then midline block at 36 Gy with 45 Gy to the whole pelvis and 51-59 Gy to the parametrium). These patients underwent high-dose-rate brachytherapy with 4 Gy/fx x 7 to point A, biweekly. During the same period, patients with non bulky tumors (Group 2, n = 31) received conventional treatment and similar brachytherapy. RESULTS: The OT of Group 1 was 7 weeks or less in 61.3%, 7.1-8 weeks in 29%, and more than 8 weeks in 9.7% (19.4%, 51.6%, and 29% in Group 2, respectively, p = 0.003). Incidences of acute complications and treatment breaks were similar in both groups. Late complication (rectal bleeding) occurred only in Group 1 (13%, 4/31), but was self-limited. Locoregional failures occurred within 2 years after completion of radiation therapy in both groups (16% and 13% in Group 1 and 2, respectively, with minimum and median follow-ups of 2 years and 34 months). CONCLUSION: Partial hyperfractionation on the third week of radiation permitted patients to finish their treatment with shorter OT without excessive acute complications and with acceptable grade 2 late rectal bleeding complications. This treatment scheme may be an effective method for the improvement of local control of bulky cervical cancer. PMID- 10863068 TI - Fractionated, three-dimensional, planning-assisted proton-radiation therapy for orbital rhabdomyosarcoma: a novel technique. AB - PURPOSE: Most children with orbital rhabdomyosarcoma will survive their disease. However, conventional photon-radiation treatment, as part of multimodality therapy, results in varying degrees of long-term functional and cosmetic side effects. This report introduces external beam proton radiation therapy (PRT) as a conformal, three-dimensional planned radiation technique for this disease, analyzes normal tissue dosimetry, and describes the technique's application in the first 2 patients. MATERIAL AND METHODS: Between January 1995 and February 1996, 2 patients underwent PRT following biopsy and chemotherapy for orbital rhabdomyosarcoma. Fifty and 55 Cobalt Gray Equivalent (CGE) were delivered to the gross tumor volume and 40 CGE to clinical target volumes in both patients. A relative biologic effectiveness (RBE) of 1.1 was utilized to correlate proton dose calculations with CGE. To achieve dose conformity, a "patch technique" was utilized, where target regions were divided into segments, each treated by a separate proton field. Dose-volume histograms were obtained for target and nontarget regions, including lens, bony orbit, pituitary gland, optic chiasm, optic nerves, lacrimal gland, and ipsilateral frontal and temporal lobes. RESULTS: At 3.4 and 2.5 years after PRT, both patients are clinically and radiographically free of disease. Visual acuity remains excellent, without signs of cataract formation; pituitary function is normal; cosmetically, only mild enophthalmos is noticeable. Doses to 90%, 50%, and 5% of lens volume were kept at less than 1%, less than 2%, and less than 8%, respectively. Fifty percent of lacrimal gland volume received less than 36% of the prescribed dose and 50% of the volume of the optic chiasm, pituitary gland, and hypothalamus were restricted to less than 2%. Proton conformity to orbital contents resulted in between 9% and 36% of the prescribed dose reaching the ipsilateral temporal and frontal lobes immediately adjacent to bony orbit (5% volume). CONCLUSION: PRT can offer excellent sparing of lens and selected intraorbital and ocular normal structures, while maintaining conformal target-dose coverage. The steep dose gradient beyond the orbit minimizes irradiation of normal brain parenchyma, with almost complete sparing of the pituitary gland. Reduction of integral irradiation exposure of the periorbital region will, hopefully, reduce the risk of second malignancy later in life. Reduced radiation dose to specific organs in close proximity to, but not part of the target region promises improved functional outcome and better cosmesis for childhood cancer survivors. PMID- 10863069 TI - Intraoperative radiation therapy for high-risk pediatric neuroblastoma. AB - PURPOSE: To evaluate the efficacy of intraoperative radiation therapy (IORT) in the treatment of high-risk pediatric neuroblastoma. METHODS AND MATERIALS: Between 1986 and 1998, 23 children received IORT for pediatric neuroblastoma. Electron beam energies ranged from 4 MeV to 16 MeV and median dose was 10 Gy (7 16 Gy). RESULTS: Twenty-one of 23 patients were classified as high-risk. A gross total resection (GTR) was achieved in 18 patients, of whom 6 experienced disease recurrence, 2 of which included a locoregional relapse as a component of failure. Fourteen of 18 patients receiving IORT after a GTR are disease-free survivors. A second subset of 5 patients had a subtotal resection (STR), with gross residual disease remaining after surgery. All 5 patients recurred locally, and all died of their disease. IORT was extremely well-tolerated in our cohort. Surgical resection and IORT resulted in the narrowing of the abdominal aorta and an atrophic kidney in 1 patient. CONCLUSIONS: For high-risk neuroblastoma patients, IORT as the only radiotherapy to the primary, produced excellent local control after a GTR. However, IORT as the sole radiotherapy to the primary was inadequate for patients with extensive adenopathy or an STR. In this setting, we are exploring the use of IORT as a boost in conjunction with external beam radiation therapy. PMID- 10863070 TI - Application of recursive partitioning analysis and evaluation of the use of whole brain radiation among patients treated with stereotactic radiosurgery for newly diagnosed brain metastases. AB - PURPOSE: To evaluate the usefulness of whole brain radiotherapy (WBRT) and of the Radiation Therapy Oncology Group recursive partitioning analysis (RPA) for brain metastases among patients receiving stereotactic radiosurgery (SRS). METHODS AND MATERIALS: A retrospective analysis was performed on 135 patients who underwent linear accelerator (Linac) (n = 73) or Gamma Knife (n = 62) SRS for newly diagnosed brain metastases at the Cleveland Clinic Foundation between 8/89 and 12/98. Univariate and multivariate analyses were performed to evaluate the effects of age, primary site, control of the primary, interval to development of brain metastases (disease-free interval [DFI]), number of brain metastases, presence of extracranial metastases, Karnofsky performance status (KPS), treatment of brain metastases, and RPA class on overall survival. RESULTS: Application of the RPA classification revealed 29 patients fit the criteria for class I, 96 for class II, and 10 for class III. All of the patients underwent SRS. Fifty-seven patients also received WBRT at the time of initial presentation (SRS and immediate WBRT), and 78 patients received WBRT only if CNS relapse occurred (SRS alone). The median survival for all patients was 7.9 months (range: 1.1-90.1), and was 11.2 months for RPA class I compared to 6. 9 months for RPA classes II-III (p = 0.016). Median survival was 10. 5 months following SRS alone compared to 6.4 months following SRS and WBRT (p = 0.07). On univariate analysis, KPS >/= 80% (p = 0.002) and absence of systemic disease (p = 0.013) were also associated with longer survival, whereas control of the primary, DFI, and number of brain metastases did not have an impact. Multivariate analysis revealed only RPA class (p = 0.023) to be an independent predictor for overall survival, whereas treatment group (p = 0.079) was only marginally significant. At 2 years, immediate WBRT improved control at the original site of metastases (80% vs. 52%, p = 0.03) and prevention of new metastatic sites within the brain, 74% vs. 48% (p = 0.06). The 2-year intracranial disease-free survival was 60% following SRS and WBRT compared to only 34% following SRS alone (p = 0.03). CONCLUSIONS: Despite the inherent biases to select more favorable patients for SRS, the RPA class retains its prognostic value. Omission of WBRT from the initial management was not detrimental in terms of overall survival; however, progressive disease occurred in over 50% of patients treated in this manner. Further studies are required to determine which, if any, patients should be considered for SRS with WBRT held in reserve. PMID- 10863071 TI - Validation of the RTOG recursive partitioning analysis (RPA) classification for brain metastases. AB - PURPOSE: The Radiation Therapy Oncology Group (RTOG) previously developed three prognostic classes for brain metastases using recursive partitioning analysis (RPA) of a large database. These classes were based on Karnofsky performance status (KPS), primary tumor status, presence of extracranial system metastases, and age. An analysis of RTOG 91-04, a randomized study comparing two dose fractionation schemes with a comparison to the established RTOG database, was considered important to validate the RPA classes. METHODS AND MATERIALS: A total of 445 patients were randomized on RTOG 91-04, a Phase III study of accelerated hyperfractionation versus accelerated fractionation. No difference was observed between the two treatment arms with respect to survival. Four hundred thirty-two patients were included in this analysis. The majority of the patients were under age 65, had KPS 70-80, primary tumor controlled, and brain-only metastases. The initial RPA had three classes, but only patients in RPA Classes I and II were eligible for RTOG 91-04. RESULTS: For RPA Class I, the median survival time was 6. 2 months and 7.1 months for 91-04 and the database, respectively. The 1-year survival was 29% for 91-04 versus 32% for the database. There was no significant difference in the two survival distributions (p = 0.72). For RPA Class II, the median survival time was 3.8 months for 91-04 versus 4.2 months for the database. The 1-year survival was 12% and 16% for 91-04 and the database, respectively (p = 0.22). CONCLUSION: This analysis indicates that the RPA classes are valid and reliable for historical comparisons. Both the RTOG and other clinical trial organizers should currently utilize this RPA classification as a stratification factor for clinical trials. PMID- 10863072 TI - The role of whole brain radiotherapy and stereotactic radiosurgery on brain metastases from renal cell carcinoma. AB - PURPOSE: We reviewed our experience with patients who have undergone stereotactic radiosurgery (SRS) for brain metastases secondary to renal cell carcinoma (RCC). Analysis was performed to determine the survival, local control, distant brain failure (DBF), and then to define which tumors may not require upfront whole brain radiotherapy (WBRT). METHODS AND MATERIALS: Twenty-nine patients with 66 tumors underwent SRS from 1991 to 1998. Median follow-up from time of brain metastases diagnoses relative to each tumor was 12.5 months and 6.8 months from the time of SRS. Median SRS dose was 1,800 cGy to the 60% isodose line. Three patients had undergone SRS for previously treated tumors. RESULTS: Median survival time from diagnosis was 10.0 months. Overall survival was not affected by age, addition of WBRT, number of lesions, tumor volume, or the presence of systemic disease. Of the 23 patients with follow-up neuroimaging, 4 of 47 (9%) tumors recurred. The addition of WBRT did not improve local control. Of the 13 patients who presented with a single lesion, 3 went on to develop DBF (23%), while 6 of the 10 patients who presented with multiple metastases developed DBF (60%). CONCLUSION: Patients with brain metastases secondary to RCC treated by SRS alone have excellent local control. The decision of whether or not to add WBRT to SRS should depend on whether the patient has a high likelihood of developing DBF. Our study suggests that patients who present with multiple brain lesions may be more likely to benefit from the addition of WBRT because they appear to be more than twice as likely to develop DBF as compared to patients with a single lesion. PMID- 10863073 TI - Gamma knife radiosurgery for trigeminal neuralgia: the initial experience of The Barrow Neurological Institute. AB - PURPOSE: To assess the efficacy and complications of Gamma Knife radiosurgery for trigeminal neuralgia. METHODS AND MATERIALS: The Barrow Neurological Institute (BNI) Gamma Knife facility has been operational since March 17, 1997. A total of 557 patients have been treated, 89 for trigeminal neuralgia (TN). This report includes the first 54 TN patients with follow-up exceeding 3 months. Patients were treated with Gamma Knife stereotactic radiosurgery (RS) in uniform fashion according to two sequential protocols. The first 41 patients received 35 Gy prescribed to the 50% isodose via a single 4-mm isocenter targeting the ipsilateral trigeminal nerve adjacent to the pons. The dose was increased to 40 Gy for the remaining 13 patients; however, the other parameters were unvaried. Outcome was evaluated by each patient using a standardized questionnaire. Pain before and after RS was scored as level I-IV per our newly-developed BNI pain intensity scoring criteria (I: no pain; II: occasional pain, not requiring medication; III: some pain, controlled with medication; IV: some pain, not controlled with medication; V: severe pain/no pain relief). Complications, limited to mild facial numbness, were similarly graded by a BNI scoring system. RESULTS: Among our 54 TN patients, 52 experienced pain relief, BNI score I in 19 (35%), II in 3 (6%), III in 26 (48%), and IV in 4 (7%). Two patients (4%) reported no relief (BNI score V). Median follow-up was 12 months (range 3-28). Median time to onset of pain relief was 15 days (range 0-192), and to maximal relief 63 days (range 0-253). Seventeen (31%) noted immediate improvement (/= 2 microg/ml). The rate of penicillin susceptibility was highest in Canada, and lowest in the South Central and South East regions of the United States. Cefepime, cefuroxime, ceftazidime, and erythromycin all demonstrated excellent efficacy (94%-99.8% susceptibility) against fully penicillin-susceptible isolates of S. pneumoniae. Among pneumococci with intermediate penicillin resistance, 88% were susceptible to cefepime, 92% to cefotaxime, and only 14% to ceftazidime. None of the antimicrobial agents in this analysis demonstrated adequate activity against fully penicillin-resistant pneumococci. In summary, the fourth-generation cephalosporin, cefepime, demonstrated consistently excellent efficacy against oxacillin-susceptible staphylococci and most pneumococci, and remains an appropriate choice for empiric therapy of serious infections. PMID- 10863104 TI - Antimicrobial activity and in vitro susceptibility test development for cefditoren against Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus species. AB - Cefditoren, a third generation orally administered aminothiazolyl cephalosporin, has demonstrated bactericidal activity against many Gram positive and negative bacterial pathogens and stability against clinically important beta-lactamases. Cefditoren was compared to cefaclor, cefixime, and penicillins against 1 435 recently isolated strains of streptococci (312 Streptococcus pneumoniae, 165 viridans group streptococci, 142 beta-haemolytic streptococci), Haemophilus influenzae (521 strains), and Moraxella catarrhalis (295 strains). Streptococcus pneumoniae and viridans group streptococci had penicillin nonsusceptible rates of 37.8 and 35.8%, respectively. Cefditoren (MIC(90) in microg/ml/% susceptible) activity against all tested H. influenzae (0.03/100) and M. catarrhalis (0.06 0.5/100) was comparable to cefixime and significantly greater than cefaclor. Cefditoren (MIC(90), 0.5 microg/ml) was 4- to 128-fold more active than comparison beta-lactams against the pneumoococci and was the most potent beta lactam (including penicillin) versus beta-haemolytic streptococci. Cefditoren pharmacokinetics demonstrate a T(1/2) of 1.5-2 h and C(max) values of 2.8 and 4.6 microg/ml, respectively with 200 or 400 mg doses of cefditoren pivoxil; plasma concentrations exceed 1 microg/ml for 4 to 6 hours (33-50% of dosing interval). Consequently, a susceptible MIC of /= 18 and >/= 15 mm (5-microg disk) for all cited fastidious species tested. Categorical agreement between MIC and disk tests was 94.6 to 100% with a correlation coefficient (r) range of 0.50 to 0.90 for streptococci. H. influenzae intermethod comparison results using the same interpretive criteria were in complete agreement, but exhibited a low r = 0.39. Cefditoren clearly possesses the most potent activity among currently studied oral cephalosporins or penicillin against commonly isolated bacterial pathogens causing bronchitis, pneumonia, sinusitis, or pharyngitis and was active against nearly all penicillin-resistant streptococci at /=2.0 microg/ml) and intermediate (MIC 0.13-1.0 microg/ml), respectively. Panipenem was most active parenteral agent against penicillin-intermediate (MIC(90) 0.125 microg/ml) and -resistant strains (MIC(90) 0.25 microg/ml). Among oral beta lactam agents, cefditoren had good activity against penicillin-intermediate and resistant strains (MIC(90) 0.5/1.0 microg/ml). Serogroup 6 was the most prevalent (65/218) among all strains and 19F (44 strains) was the most prevalent among penicillin-intermediate and -resistant strains. Both pbp2b resistant and susceptible genes were found in penicillin-intermediate strains. Pbp2x resistant genes were found in 33 of 80 (41.3%) cefotaxime-susceptible strains. These results suggest that possible resistance mechanisms may occur even in drug susceptible strains and that drug susceptibility survey should be updated carefully in Japan. PMID- 10863106 TI - Respiratory tract pathogens isolated from patients hospitalized with suspected pneumonia: frequency of occurrence and antimicrobial susceptibility patterns from the SENTRY Antimicrobial Surveillance Program (United States and Canada, 1997). AB - Thirty-seven sentinel hospitals (29 in the United States [US]; eight in Canada) collected bacterial isolates from hospitalized patients with a diagnosis of pneumonia. The antimicrobial susceptibility patterns of these pathogens were determined to more than 60 agents (40 reported) using the reference broth microdilution method described by the National Committee for Clinical Laboratory Standards. The five most frequently recorded species among the 2757 isolates collected during the study were (no. tested/%): Staphylococcus aureus (632/22.9%), Pseudomonas aeruginosa (498/18. 1%), Haemophilus influenzae (284/10.3%), Klebsiella spp. (240/8.7%), and Streptococcus pneumoniae (213/7.7%). There was a significant difference in the susceptibility to antimicrobials between the US and Canada for S. aureus to oxacillin (50.1% versus 93.8% susceptible, respectively), gentamicin (78.7% versus 97.8%), and fluoroquinolones (49.5 to 53.0% versus 89.8 to 94.9%). Amikacin (92. 8% susceptible) was the most active antimicrobial agent against P. aeruginosa, and meropenem was the most potent beta-lactam. Against H. influenzae, most drugs retained a high level of activity, whilst against the S. pneumoniae, only the newer fluoroquinolones (gatifloxacin, levofloxacin, sparfloxacin) remained highly effective in vitro. Only two antimicrobial agents (imipenem and meropenem) were >99% active against the Klebsiella spp. and Enterobacter spp. isolated in this survey (possess extended spectrum beta-lactamases or hyperproduction of Amp C cephalosporins); cefepime (95.6-100.0% susceptible) was significantly more active than other cephalosporins tested. Clonal, epidemic outbreaks of multiply resistant strains were very rare in monitored hospitals. In conclusion, important differences exist between the US and Canada in the susceptibility patterns of some respiratory tract pathogens to commonly used antimicrobial agents with Canadian strains generally being more susceptible to currently available antimicrobial agents. PMID- 10863107 TI - Comparison of antimicrobial resistance phenotypes and resistance genes in Enterococcus faecalis and Enterococcus faecium from humans in the community, broilers, and pigs in Denmark. AB - Enterococcus faecalis and E. faecium isolated from humans in the community (98 and 65 isolates), broilers (126 and 122), and pigs (102 and 88) during 1998 were tested for susceptibility to 12 different antimicrobial agents and for the presence of selected genes encoding resistance using PCR. Furthermore, the presence of vancomycin resistant enterococci was examined in 38 human stool samples using selective enrichment. Widespread resistance to chloramphenicol, macrolides, kanamycin, streptomycin, and tetracycline was found among isolates from all three sources. All E. faecium isolates from humans and pigs were susceptible to avilamycin, whereas 35% of isolates from broilers were resistant. All E. faecium isolates from humans were susceptible to vancomycin, whereas 10% and 17% of isolates from broilers and pigs, respectively, were resistant. A vancomycin resistant E. faecium isolate was found in one of the 38 human fecal samples examined using selective enrichment. All vancomycin resistant isolates contained the vanA gene, all chloramphenicol resistant isolates the cat(pIP501) gene, and all five gentamicin resistant isolates the aac6-aph2 gene. Sixty-one (85%) of 72 erythromycin resistant E. faecalis examined and 57 (90%) of 63 erythromycin resistant E. faecium isolates examined contained ermB. Forty (91%) of the kanamycin resistant E. faecalis and 18 (72%) of the kanamycin resistant E. faecium isolates contained aphA3. The tet(M) gene was found in 95% of the tetracycline resistant E. faecalis and E. faecium isolates of human and animal origin, examined. tet(K) was not observed, whereas tet(L) was detected in 17% of tetracycline resistant E. faecalis isolates and in 16% of the E. faecium isolates. tet(O) was not detected in any of the isolates from pigs, but was observed in 38% of E. faecalis isolates from broilers, in two E. faecalis isolates from humans and in three E. faecium isolates from broilers. tet(S) was not detected among isolates from animals, but was observed in 31% of E. faecalis and one E. faecium isolate from humans. This study showed a frequent occurrence of antimicrobial resistance and the presence of selected resistance genes in E. faecalis and E. faecium isolated from humans, broilers and pigs. Differences in the occurrence of resistance and tetracycline resistance genes were observed among isolates from the different sources. However, similar resistance patterns and resistance genes were detected frequently indicating that transmission of resistant enterococci or resistance genes takes place between humans, broilers, and pigs. PMID- 10863108 TI - Antimicrobial activity of advanced-spectrum fluoroquinolones tested against more than 2000 contemporary bacterial isolates of species causing community-acquired respiratory tract infections in the United States (1999). AB - In vitro activity of four newer fluoroquinolones (clinafloxacin, gemifloxacin, moxifloxacin, sitafloxacin) and an equal number control drugs in the same class (ciprofloxacin, grepafloxacin, levofloxacin, trovafloxacin) was determined by reference dilution tests against 2156 recent United States clinical isolates of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. All the fluoroquinolones demonstrated excellent in vitro activity against these pathogens. Streptococcus pneumoniae isolates were fully susceptible to clinafloxacin, sitafloxacin, and gemifloxacin at 0.5 microg/ml, and over 98% of sampled strains had MICs of 16 microg/ml; median zone, 18-25 mm). Against Pseudomonas aeruginosa, Stenotrophomonas maltophilia, Acinetobacter spp., and other non-fermentors, cefditoren was inactive (MIC(90), > 16 microg/ml; zone, 6 mm). Pharmacokinetic analysis of cefditoren showed that utilized dosages produce a plasma concentration that exceeds 0.5 microg/ml for 5 to 8 h and 1 microg/ml for 4 to 6 hours (T(1/2) ranges from 1.5-2 h). The following interpretive criteria were suggested: /= 15 mm (susceptible) and >/= 8 microg/ml or /= 18 mm (susceptible) and >/= 4 microg/ml or 200 (39.0 degrees C, 102.3 degrees F), those with CD4 < or =200 (39.2 degrees C, 102.5 degrees F), and the HIV- group (39.2 degrees C, 102.5 degrees F). Nearly 8% of i.v. drug users with confirmed IE presented to the ED with a T(max) below 37.8 degrees C (100.0 degrees F). Mean WBC count was significantly lower in HIV+ (11.1 k/mm(3)) than in HIV- patients (15.4 k/mm(3)) and significantly lower in the group with CD4 < or =200 (8.0 k/mm(3)) than in the HIV group. In conclusion, HIV infection was not associated with lower T(max), but it was associated with decreased WBC count in the general HIV+ group and in the group with CD4 < or =200. PMID- 10863112 TI - The extent of undiagnosed HIV infection among emergency department patients: results of a blinded seroprevalence survey and a pilot HIV testing program. AB - This study was performed to determine the rate of previously undiagnosed HIV infection among patients presenting to an urban emergency department (ED) and to assess the feasibility of routinely offering voluntary HIV testing in this setting. HIV serostatus was determined anonymously among consecutive acute medicine and trauma ED patients (aged 18-55) who had blood drawn as part of their medical care. Excess serum was aliquoted and coded with an anonymous study code. Before performing HIV testing, the number of persons with previously reported HIV infection was determined by linkage with the state HIV/AIDS reporting registry. Concurrent with the blinded HIV serosurvey, ED patients were offered voluntary HIV testing in a pilot program. Overall, 76 of 2,155 (3.5%) adult ED patients in the blinded survey were HIV-seropositive, 15 of whom (0. 7% of those tested, 20% of those HIV-seropositive) had no infection previously reported to the state HIV/AIDS registry. In the pilot program, six of the 156 (3.8%) individuals who underwent voluntary HIV testing were HIV-seropositive, including three of 53 (5.6%) individuals without prior HIV testing. Of the six HIV-seropositive subjects, one was previously diagnosed, while five of the remaining 155 (3.2%) represented previously undiagnosed infections. Overall, 3. 5% of ED patients from whom blood was obtained for other reasons tested positive for HIV antibody, 20% of whom were previously undiagnosed. Implementation of the voluntary testing program uncovered newly diagnosed infection among 3.2% of those tested. An ED may be an important setting for routinely offering HIV testing, especially for patients who have not been previously tested for HIV. PMID- 10863113 TI - Clinical utility of CKMB isoform determinations in patients who present to the emergency department with continuous or resolved chest pain. AB - The object of this investigation was to demonstrate that the enhanced sensitivity of the diagnosis of acute myocardial infarction (AMI) using a more-inclusive criterion of CKMB Isoforms may detect earlier stages of AMI (designated Isoform Type 2) than the currently accepted marker for AMI by CKMB Isoform (designated Isoform Type 1) in a busy, urban Emergency Department (ED). Two features characterized the study of CKMB Isoforms in a prospective cohort of 223 ED patients: first, nontraumatic chest pain within 12 h before presentation, thought to be of ischemic etiology; and second, normal or nondiagnostic electrocardiogram (EKG). Patients were further divided into two groups characterized as either recent but resolved chest pain at ED visit, or ongoing or staccato chest pain. Sensitivity (S), specificity (SP), positive (PPV) and negative (NPV) predictive values, and 95% confidence intervals (CI) for AMI diagnosis were determined. Two criteria for AMI diagnosis by CKMB Isoforms were tested. The first and currently recommended criterion was identified as Isoform Type 1. An AMI diagnosis by Type 1 criterion requires both CKMB2> or =2.6 IU/L and CKMB2/CKMB1> or =1.7. The second criterion for AMI diagnosis was identified as Isoform Type 2, which is defined as either CKMB2> or =2.6 IU/L or CKMB2/CKMB1> or =1.7. Both Isoform types are predictive of AMI by the gold standard, and addition of EKG changes results in a small improvement. Type 1 demonstrates SP 0.94 (CI 0.90, 0.97) and NPV 0.90 (CI 0.86, 0.94), and Type 2 demonstrates S 0.90 (CI 0.80, 0.97) and NPV 0.97 (CI 0.93, 0.99) for AMI diagnosis. Type 2 criteria can confidently exclude the immediate risk of AMI in patients with resolved chest pain whereas in patients with continuous chest pain, Type 1 criteria may identify those at high risk for AMI. PMID- 10863114 TI - Pulmonary embolism complicated by patent foramen ovale and paradoxical embolization. AB - We present the case of a 74-year-old male with chest pain, dyspnea, and syncope secondary to an acute pulmonary embolism complicated by a patent foramen ovale with straddling thrombus and paradoxical embolization. We review the literature with specific focus on the pathogenesis and acute treatment of this life threatening occurrence. PMID- 10863115 TI - Paradoxical embolism-report of a case involving four organ systems. AB - Paradoxical embolism through a patent foramen ovale (PFO) can involve multiple organs simultaneously. The most commonly involved sites are the cerebrum and the extremities. Paradoxical embolism to coronary arteries or upper extremities is relatively uncommon. We report a case of acute pulmonary embolism and paradoxical embolism through a patent foramen ovale involving the left upper extremity, brain, and coronary artery. Early diagnosis in the emergency department was made by a trans-esophageal echocardiogram, and the patient was successfully treated with intravenous t-PA and heparin. Patients with acute pulmonary embolism or deep venous thrombosis who also develop signs of systemic embolism should be evaluated for a patent foramen ovale. PMID- 10863116 TI - Bilateral internal carotid artery dissection due to trivial trauma. AB - Internal carotid artery dissection (ICAD) is a known cause of unilateral headache and focal cerebral ischemic symptoms. Other symptoms include oculosympathetic paresis, facial pain, neck pain, subjective carotid bruits, and cranial nerve deficits. Traumatic dissection has an obvious precipitating incident preceding the neurologic or visual symptoms. An ICAD that occurs spontaneously or from trivial trauma usually lacks an obvious incident and thus requires awareness of its possibility for accurate detection and treatment. Dissections arise from a defect in the internal elastic lamina allowing penetration of blood into the arterial wall. Despite its low incidence, ICAD must be considered in young to middle-aged patients who present with headache and transient cerebral or retinal ischemic symptoms. This report describes a patient who had bilateral internal carotid artery dissections following trivial trauma. The etiologies, clinical manifestations, diagnostic modalities, treatment options, and outcomes of ICAD are discussed. PMID- 10863117 TI - Traumatic pneumomediastinum caused by isolated blunt facial trauma: a case report. AB - Traumatic pneumomediastinum is most often identified as an incidental finding in the setting of blunt or penetrating neck, chest, or abdominal trauma. There are only a few cases in the medical literature of a pneumomediastinum following isolated facial trauma. We present a patient who sustained fractures of the lateral and anterior walls of the right maxillary sinus, floor of the right orbit, and right zygomatic arch. Subcutaneous emphysema overlaid the right facial region and extended to the left side of the neck and into the mediastinum. We describe this unusual complication with respect to the anatomic relations of the facial and cervical fascial planes and spaces with the mediastinum. PMID- 10863118 TI - Coma and respiratory depression following the ingestion of GHB and its precursors: three cases. AB - Gamma hydroxybutyrate (GHB) is a product of the metabolism of both gamma butyrolactone (GBL) and 1,4-butanediol (1,4-BD). Gamma hydroxybutyrate (GHB) is an illegal agent that causes central nervous system depression. Chemical precursors of GHB, such as GBL and 1,4-BD, have been available for purchase from many health food stores and Internet websites for mood-enhancement, sleep induction, and stimulation of growth hormone release. We report three cases of ingestion of products containing GHB and chemical precursors of GHB. All three patients had severe presentations followed by full recoveries. Some products containing GBL were withdrawn from the market after the FDA issued a warning regarding these products. Products containing 1,4-butanediol remain on the market today. PMID- 10863119 TI - Rhabdomyolysis and drugs of abuse. AB - Rhabdomyolysis is a disorder in which injury to muscle results in leakage of myocyte intracellular contents into the plasma. It has been associated with a tremendous number and diversity of clinical conditions and substances. Several physiological and biochemical mechanisms for this syndrome have been described. The most likely etiology of rhabdomyolysis in patients presenting to the emergency department is ingestion of drugs of abuse, most commonly ethanol, heroin, amphetamines, cocaine, and other sedatives or stimulants. In this article, the association between rhabdomyolysis and drugs of abuse is explored, as well as its diagnosis and treatment. PMID- 10863120 TI - Syncope in pregnancy. PMID- 10863121 TI - The utility of oxygen in myocardial infarction. PMID- 10863122 TI - The Glasgow coma scale. AB - Teasdale and Jennett first presented the Glasgow Coma Scale in 1974 as an aid in the clinical assessment of unconsciousness. It was devised as a formal scheme to overcome the ambiguities and misunderstandings that arose when information about comatose patients was presented and groups of patients were compared. Since then, the Glasgow Coma Scale has been used extensively, being used to grade individual patients, compare effectiveness of treatments, and as a prognostic indicator. It has been incorporated into numerous trauma and critical illness classification systems. However, a number of competing scales have been developed to overcome its perceived deficiencies. These scales are generally more complex. One of the expressed reservations regarding the Glasgow Coma Scale has been its failure to incorporate brainstem reflexes. The scale also includes a numerical skew toward the motor response. An important current issue is the appropriate application of the Glasgow Coma Scale to intubated patients. A number of approaches have been used to assign the verbal score to such patients. The timing of initial scoring is another area of discussion. Despite its drawbacks, the Glasgow Coma Scale remains the most universally utilized level of consciousness scale worldwide. It seems destined to be used in emergency medicine for some time. PMID- 10863123 TI - The public health career of jack C. Smith. Looking beyond the statistics PMID- 10863124 TI - The role of linked birth and infant death certificates in maternal and child health epidemiology in the United States. AB - Linked birth and infant death certificates allow measurement of birthweight specific infant mortality. Jack Smith, MS, to whom this issue of the American Journal of Preventive Medicine is dedicated, played a key role in the National Infant Mortality Surveillance (NIMS) project. NIMS provided national data on birthweight-specific infant mortality for the 1980 birth cohort, updated data previously collected by the National Center for Health Statistics (NCHS) for the 1960 birth cohort, and supported NCHS's implementation of an annual linked file in 1983. NIMS illustrated themes in infant mortality that remain important: the role of low birthweight (LBW) as a contributor to infant mortality, the contribution of disparities in LBW and birthweight-specific mortality to black white gaps in infant mortality, and the nation's greater success in reducing mortality among LBW infants than in preventing LBW. Linked birth and infant death records are used nationally and by states to study an array of maternal and infant health topics, from the quality of vital records to the impact of therapeutic and public health interventions. By supplementing birth and infant death records with linkages to program and hospital discharge data, epidemiologists and health service researchers are extending the utility of vital statistics data to monitor maternal and infant health. PMID- 10863125 TI - Abortion surveillance at CDC: creating public health light out of political heat. AB - In the late 1960s, states began to liberalize their abortion laws, and a new era in women's health began. Under the leadership of Jack Smith, the Centers for Disease Control and Prevention (CDC) established a voluntary abortion surveillance system that provided the first nationwide information on the numbers and characteristics of women having abortions. Studies of abortion morbidity done by the CDC revealed that suction curettage was safer than sharp curettage, local anesthesia was safer than general anesthesia, free-standing clinics were safer than hospitals, and dilation and evacuation (D&E) was safer than the alternative of labor induction for early second-trimester abortions. This evidence, which contradicted traditional medical tenets, rapidly changed the practice of abortion in the United States. CDC also established a surveillance system for abortion deaths. This demonstrated a rapid improvement in the safety of abortion in the early 1970s. Lessons learned from mortality investigations helped to change practice as well.Today, more is known about the epidemiology of abortion than any other operation in the history of medicine. In the midst of strident debate over the abortion issue, CDC abortion surveillance data have helped to guide judicial rulings, legislative actions, and Surgeon General's reports, which have supported safer choices for women of reproductive age. When medical historians of the future look back on this century, the increasing availability of safe, legal abortion will stand out as a public health triumph. PMID- 10863126 TI - Teenage childbearing in the United States, 1960-1997. AB - Teenage childbearing in the United States has declined significantly in the 1990s. Still the U.S. teen birth rate is higher than in other developed countries; in 1997 it was 52.3 births per 1000 women aged 15 to 19. A steep rise in teen birth rates in the late 1980s generated a great deal of public concern and a variety of initiatives targeted to reducing teen births. Data from the National Center for Health Statistics' National Vital Statistics System are used to review and describe trends and variations in births and birth rates for teenagers for the period 1960-1997. Teen birth rates were much higher in the early 1960s than at present; in fact, rates for 18- to 19-year-olds were double what they are currently. In the 1990s, birth rates for teenagers dropped for younger and older teenagers, with greater declines recorded for younger teens. While rates have fallen in all population groups, the greatest declines have been experienced by black teenagers, whose rates have dropped 24% on average. %Trends in teen births and birth rates since 1960 have been affected by a variety of factors. These include wide swings in the number of female teenagers, substantial declines in marriage among older teens, falling birth rates for married teens concurrent with rapidly rising birth rates for unmarried teens, and sharp increases in sexual activity among teens that have abated only recently, according to the National Center for Health Statistics' National Survey of Family Growth. This review article also tracks changes in contraceptive practice and abortion rates. PMID- 10863127 TI - Building a foundation for suicide prevention: the contributions of Jack C. Smith. AB - Among his many other accomplishments, Jack C. Smith nurtured the early development of efforts by the Centers for Disease Control and Prevention (CDC) to address suicide as a public health problem. Smith's vision was to achieve suicide prevention through epidemiology, and his vision shaped the emergence of suicide as a public health issue. With his typical enthusiasm and inherent ability to insinuate himself into critical social networks, Smith spearheaded CDC's initial suicide surveillance activities and established strong partnerships between CDC and the National Institute of Mental Health (NIMH) and the American Association of Suicidology (AAS). These surveillance activities and relationships were the foundation on which subsequent research and programmatic activities addressing suicide as a public health problem were built at CDC. In this paper we document Smith's role in the development of the public health approach to suicide prevention. We also articulate the conceptual basis for a public health approach to suicide and discuss future directions for public health in the prevention of suicide and suicidal behavior. While Smith also made important contributions to development of CDC efforts to address homicide, his special interest was suicide; therefore, this article will emphasize his contributions to this area. PMID- 10863128 TI - History and current status of reproductive health surveys at CDC. PMID- 10863129 TI - The check box: determining pregnancy status to improve maternal mortality surveillance. AB - OBJECTIVE: More than half of pregnancy-related deaths are not identified through routine surveillance methods. The purpose of this study was to evaluate the effectiveness of the pregnancy check box on death certificates in ascertaining pregnancy-related deaths. METHODS: Data derived from the Centers for Disease Control and Prevention's ongoing Pregnancy Mortality Surveillance System were used to identify states that included a check box on the death certificate in 1991 and 1992. Death certificates from those states were evaluated to determine the number and proportion of pregnancy-related deaths identified by a marked check box. Characteristics of death were also examined. RESULTS: Sixteen states and New York City included a check box or question specifically asking about pregnancy of the decedent. Of the 425 pregnancy-related deaths identified in the 17 reporting areas, 124 (29%) were determined to be pregnancy-related deaths only because of the pregnancy status information provided in the check box. The proportion of deaths identified only by a marked check box ranged from less than 5% for four states to 40% or more for seven states. CONCLUSIONS: The availability of pregnancy status information on death certificates is a simple and effective aid in ascertaining a pregnancy-related death, when no other indicators of pregnancy appear on the death certificate. Routine use of the pregnancy check box for all states would lead to substantially increased classification of maternal deaths and more accurate classification of the causes of and risk factors for maternal deaths. PMID- 10863130 TI - Facilitating use of analytic methods at a federal agency. AB - Identification and application of appropriate analytic methods are critical pieces of the foundation of science. Because of the increasingly complex issues federal agencies face and the increasing importance of addressing those issues with multi-disciplinary teams that offer knowledge and skills in many methods areas, it is important that agencies foster coordinated integration of analytic methods. This report discusses the Centers for Disease Control and Prevention's (CDC's) evolving needs regarding analytic methods and includes activities that CDC has undertaken to facilitate a coordinated approach to the use of the statistical sciences. We introduce a new framework for facilitating coordination in analytic methods as a hybrid model, blending attributes of centralized and decentralized resource models. This coordinating focus approach offers assets of timeliness, efficiency, and effectiveness, as well as fosters strengthened relationships among scientists and increased collaboration among scientific disciplines. PMID- 10863131 TI - Saving statistical lives: contributions of statistics to public health. AB - Prevention is about saving "statistical lives"-lives that society knows about only through the efforts of public health statisticians like Jack Smith. As Mr. Smith's lifework suggests, statistics in public health are critical for calling attention to problems, identifying risk factors, and suggesting solutions, and ultimately for taking credit for our successes. His work illustrates two important lessons about public health statistics today.First, it's important to get the facts straight. Mr. Smith's experience shows that a careful, thoughtful analysis is not only more convincing in the end, but also brings to light important subtleties not seen in the initial analysis. Second, it takes considerable time and attention to get the facts straight. Jack Smith's work illustrates the importance of partnerships with other federal agencies, state statistical organizations, and private-sector entities. PMID- 10863132 TI - Vision care professions-going down the wrong path? PMID- 10863134 TI - Clinical implications. PMID- 10863133 TI - Accelerated photostability testing of precision UV(TM) contact lenses. AB - Precision UV(TM) contact lenses were photoaged beyond their expected lifetime in an accelerated manner with a SUNTEST(R) CPS(+) system, which is a solar spectrum equivalent. Ultraviolet spectra of lenses irradiated for 30 h in saline at 41 degrees C and 44 degrees C at the highest power setting were statistically indistinguishable from those measured prior to the exposure. In a similar experiment run at 79 degrees C, the UVB range of the spectra showed small but statistically significant differences, whereas the UVA range of the spectra were not distinguishable. Lenses photoaged at half power and twice the time at 43 degrees C gave UV spectra that could be distinguished. Refractive indices of lenses were indistinguishable before and after irradiation. Eluted materials could not be detected in the saline used to maintain lenses in any irradiation experiment. Lens parameters and water contents remained within manufacturing specifications before and after irradiation. PMID- 10863135 TI - Two different concentrations of Opti-Plus(R) compared for daily protein cleaning of a disposable contact lens. AB - The aim of this study was to compare the relative safety, efficacy, comfort, and convenience of the daily use of a pancreatin-based, liquid enzyme daily protein remover (DPR) at two different concentrations for protein cleaning of Group IV disposable contact lenses.We enrolled 73 subjects (146 eyes) into a 3-month, randomized, investigator-masked clinical study comparing a test regimen of DPR (Opti-Plus(R) or Supra-Clens(R)) containing four drops per 10 mL of multifunctional solution (Opti-Free(R)) to a control regimen containing only two drops per 10 mL of multifunctional solution.The liquid enzyme regimens provided safe and effective protein cleaning of the Group IV lenses with no additional cleaning benefit observed at the higher concentration. Three convenience parameters (steps required, time required, and dexterity required) had mean improvements for both regimens at Day 90 compared to baseline (Day 0). Thus, subjects perceived the DPR regimen as more convenient than their pre-study regimen, which required weekly use of an enzymatic cleaning tablet. Comfort remained consistent with baseline levels throughout the study regardless of concentration of DPR for 72 of 73 subjects.This study provided evidence that the daily use of a low concentration pancreatin enzymatic solution admixed with multifunctional solution is well tolerated and provides a more convenient method of achieving simultaneous disinfection and protein cleaning during contact lens storage than enzyme tablets. PMID- 10863136 TI - The "pe Dk/L" and "pe Dk": part 2. single material oxygen pathways. PMID- 10863137 TI - Acute effects of gamma-vinyl-GABA on low-magnesium evoked epileptiform activity in vitro. AB - Vigabatrin (gamma-vinyl-GABA, VGB) is a gamma-aminobutyric acid (GABA) derivative designed to boost synaptic inhibition by inhibiting the degradation of GABA in brain tissue. Indeed, VGB shows potent anti-convulsant activity in animal models of epilepsy and in humans with complex partial seizures. However, details of the mechanism of action of VGB are not well understood and the systemic effects include possible pro-convulsant actions. We therefore analysed the effects of VGB in rat brain slices in the low-Mg(2+) model in vitro. VGB at 100 microM-5 mM showed a concentration- and time-dependent reduction of interictal-like events in the hippocampal CA1 region. Likewise, VGB suppressed epileptiform discharges in the medial entorhinal cortex (mEC), which are known to resist conventional anti convulsants. In contrast, evoked population spikes in CA1 (which became repetitive after washout Mg(2+)) were not altered by VGB. Our data show that VGB is efficient against epileptiform discharges in temporal structures including pharmacoresistant patterns of activity. The waveform of evoked population spikes in this in vitro model is no indicator for the anti-convulsant properties of drugs. PMID- 10863138 TI - Influence of D-cycloserine on the anticonvulsant activity of some antiepileptic drugs against audiogenic seizures in DBA/2 mice. AB - D-Cycloserine (DCS; 1-100 mg/kg, intraperitoneally (i.p.)) was able to antagonise the audiogenic seizures in DBA/2 mice in a dose-dependent manner. DCS, 2.5 mg/kg i.p. did not significantly affect the occurrence of audiogenic seizures in DBA/2 mice, but potentiated the anticonvulsant activity of carbamazepine, diazepam, felbamate, lamotrigine, phenytoin, phenobarbital and valproate against sound induced seizures in DBA/2 mice. The degree of potentiation induced by DCS was greatest for diazepam, phenobarbital, phenytoin and valproate, less for carbamazepine and least for lamotrigine and felbamate. The increase in anticonvulsant activity was usually associated with a comparable increase in motor impairment. However, the therapeutic index of the combined treatment of the above drugs+DCS, was more favourable than the same drugs+saline with the exception of DCS+carbamazepine and DCS+lamotrigine. Since DCS did not significantly influence the total and free plasma levels of the anticonvulsant drugs studied, pharmacokinetic interactions, in terms of plasma levels, are not probable. The possibility that DCS can modify the clearance from the brain of the anticonvulsant drugs studied cannot be excluded. DCS did not significantly affect the hypothermic effects of the anticonvulsants tested. In conclusion, DCS potentiates the anticonvulsant action of some classical antiepileptic drugs, most notably diazepam, phenobarbital, phenytoin and valproate. PMID- 10863139 TI - Inter- and intra-subject variability in gabapentin absorption and absolute bioavailability. AB - Gabapentin (GBP) is a non-metabolized, non-plasma protein bound, renally excreted antiepileptic drug that is actively absorbed via the system L amino acid transporter. Previous studies have demonstrated that gabapentin displays dose dependent absorption. OBJECTIVES: These studies were conducted to determine inter and intra-subject variability of gabapentin absorption. Two prospective clinical studies in healthy adult volunteers were conducted. Coefficient of variation (CV) was used to express variability of gabapentin absorption. METHODS: Study A: 400 mg single dose, randomized, cross-over study to assess bioavailability of four different gabapentin formulations (n=20, 9 males, 11 females; mean age and weight 41 years, 75.1 kg). Plasma was serially collected up to 48 h and bioavailability (F) calculated post-dose to determine concentration-time curves (AUC). All four formulations were bioequivalent, thus repeated measures analysis was performed to assess inter-and intra-subject variability. Study B: 600-mg single dose study (n=50, 15 males, 35 females; mean age and weight 31.1 years, 72.7 kg) was conducted to determine inter-subject variability in gabapentin F. Urine was collected over 48 h and bioavailability (F) calculated. Urine and plasma gabapentin concentrations were measured by HPLC-UV. RESULTS: Study A: Overall mean (CV) of GBP AUC values was 34.1+/-24 ug/h per ml. Inter-subject CV for AUC was 22.5% and intra-subject CV was 12.1%. Study B: Overall mean (SD) GBP F was 49.3+/-13.6%. Inter-subject CV of F was 27.6%. DISCUSSION: The inter-subject variability in gabapentin absorption is substantially less than that of the inter subject variability. This indicates that one would expect a wide range in gabapentin absorption between subjects; however, a much smaller variability within a subject. The within subject variability of gabapentin is small enough that plasma drug monitoring may be used to assess gabapentin absorption for a given subject and the benefit of dose individualization. PMID- 10863140 TI - Cortical somatosensory evoked potentials of seizure-sensitive and seizure resistant gerbils. AB - The Mongolian gerbil is known as an animal model that often exhibits spontaneous seizures that are characteristic of human epilepsy. Whereas there is much more information available relating this phenomenon to anatomical and electrophysiological characteristics of the hippocampal formation, the somatosensory cortex has rarely been the focus of attention. Given the existence of the fine grain cortical barrels developed in an orderly matrix, the vibrissa ascending system was thought the best sensory channel in which the gerbil neocortical excitability was to be tested. In the present study, we compared cortical evoked potentials to electric stimulation of vibrissa (whisker) follicle between the seizure-sensitive (SS) and seizure-resistant (SR) gerbils. Whereas our standard stimulation of the whisker follicle elicited a positive-negative biphasic somatosensory cortical potential in SR animals, it evoked only a positive monophasic potential in SS animals. Although the amplitude of positivity was generally larger in SR animals, the latency to reach this peak was significantly smaller than in the SS animals. Apparently there was no group difference in the laminar profile or the size of barrels that represent, in one to-one fashion, the array of whisker follicles. It was suggested that an altered level of excitability in the absence of gross anatomy differentiates the seizure sensitive gerbils from the seizure-resistant counterparts. Possible functional differences between the cortices of the two groups were discussed in light of the known synaptic electrophysiology. PMID- 10863141 TI - Lack of substantial effect of the H(3)-antagonist thioperamide and of the non selective mixed H(3)-antagonist/H(1)-agonist betahistine on amygdaloid kindled seizures. AB - We investigated whether some histamine H(3)-antagonists would attenuate amygdaloid kindled seizures in rats. Thioperamide, a standard H(3)-antagonist, did not significantly reduce either seizure ranks or afterdischarge duration (ADD). Betahistine which has both H(3)-antagonistic activity and H(1)-agonistic activity significantly reduced ADD, albeit mild at a toxic dose, though seizure ranks were not affected. In addition, L-histidine, the precursor of histamine, affected neither seizure ranks, nor ADD. It was shown that H(3)-antagonists have no significant inhibitory action against amygdaloid kindled seizures, probably because released histamine was unable to inhibit those seizures. PMID- 10863142 TI - A comparison of topiramate and acetazolamide on seizure duration and paired-pulse inhibition in the dentate gyrus of the rat. AB - Topiramate is a relatively new antiepileptic drug with several putative anticonvulsant mechanisms. Among them is the ability to inhibit carbonic anhydrase, a property in common with the anticonvulsant acetazolamide. This study examined the effects of topiramate and acetazolamide on the duration of epileptiform activity and on paired-pulse inhibition in the dentate gyrus in urethane anesthetized adult Sprague-Dawley rats. Neither topiramate nor acetazolamide altered excitability in the dentate gyrus, as measured with input output curves or induction of long-term potentiation. Topiramate increased paired pulse inhibition, whereas acetazolamide had no effect. Both drugs dose dependently blocked the lengthening of the duration of epileptiform activity compared to vehicle controls. These results indicate that topiramate has an anticonvulsant-related effect (increase in paired-pulse inhibition), which may contribute to its antiepileptic effect, that is not dependent on its ability to inhibit carbonic anhydrase. PMID- 10863143 TI - MRI volumetry of the hippocampus, amygdala, entorhinal cortex, and perirhinal cortex after status epilepticus. AB - Neuronal damage has been observed in the medial temporal lobe of both humans and animals following status epilepticus. The aim of the present study was to investigate the occurrence of medial temporal lobe damage in status epilepticus patients treated in hospital with a predetermined protocol and to assess whether the changes progress in a long-term follow-up. The volumes of the hippocampus, amygdala, entorhinal and perirhinal cortices were measured using magnetic resonance imaging (MRI) in nine adult patients with status epilepticus 3 weeks, 6 and 12 months after the insult. The control group included 20 healthy subjects. The etiology of status epilepticus was an acute process in one patient and a chronic process in eight cases. The mean duration of secondarily generalized tonic-clonic status epilepticus episodes was 1 h and 44 min. Volumetric MRI indicated that none of the patients developed marked volume reduction in the hippocampus, amygdala, or the entorhinal and perirhinal cortices during the 1 year follow-up period. Status epilepticus does not invariably lead to a progressive volume reduction in the medial temporal lobe structures of adult patients treated promptly in hospital with a predetermined protocol for rapid cessation of seizure activity. PMID- 10863144 TI - Temporal profile of CRE DNA-binding activity in the rat hippocampus following a kindling stimulation. AB - This study was designed to establish a role for cAMP-responsive element (CRE) binding protein (CREB) in signal transduction cascade in the hippocampus associated with kindling. Male Sprague-Dawley rats were kindled from the left amygdala until they exhibited Racine's class 5 generalized seizures [Racine (1972). EMBO J. 11, 3337-3346] Nuclear proteins were extracted from dorsal hippocampi obtained from 0 to 24 h after final kindling stimulation. From these, we evaluated the temporal pattern of CRE DNA-binding activity by use of a gel mobility-shift assay with a 32P-labeled CREB oligonucleotide probe. CRE-binding activity in the hippocampus was enhanced significantly at 2 h and returned to baseline level within 4 h after the stimulation. Our results suggest that CREB may be involved in the hippocampal signal transduction pathway of rats activated in response to a kindling stimulation to the amygdala. However, the transient elevation of CRE-binding activity following a seizure in a kindled animal also suggests that persistent activation of CREB may not be required for maintenance of the kindling phenomenon. PMID- 10863145 TI - Relationship of sigma activity to sleep interictal epileptic discharges: a study in children affected by benign epilepsy with occipital paroxysms. AB - INTRODUCTION: By applying spectral analysis techniques we recently showed that Interictal Epileptic Discharges (IEDs) are modulated by sleep spindle synchronization mechanisms (sigma activity, SA, 12. 0-16.0 Hz). This finding applies to both benign epilepsy of childhood with rolandic spikes (BECRS), to symptomatic epilepsy of childhood strongly activated by sleep and to the Landau Kleffner syndrome. These results are quite different from those found in adult partial epileptic patients where slow wave activity (SWA, 0. 5-4.5 Hz) plays the main role in the modulation of IEDs during sleep. This finding could suggest that the activation of IEDs by spindle activities could be an age-related feature of epilepsy. In order to verify this hypothesis we studied a group of epileptic children performing a polysomnographic study on five patients with BEOP strongly activated by sleep. METHODS: We performed overnight continuous EEG polysomnographic studies in five patients (mean age 6. 0+/-2.5). The IEDs count was performed on the most active occipital lead. The temporal series of SWA and SA values, derived from spectral analysis, were obtained from a spike-free central, controlateral lead. Relationships between SA, SWA and time series of IEDs were tested by means of correlation techniques after data normalization. RESULTS: Our results revealed a significantly higher correlation between IEDs and SA with respect to SWA in all subjects, in total sleep time. When the analysis was limited only to NREM sleep the correlation between sigma and IEDs was even more impressive. CONCLUSIONS: Data suggest that also in BEOP the spindle generating mechanism modulates the IEDs during sleep. This mechanism seems to be an age-dependent phenomenon with no relation whatsoever either with the type of epilepsy or with the brain region. PMID- 10863146 TI - Effects of carbamazepine on acetylcholine release and metabolism. AB - To clarify the mechanisms of action of carbamazepine (CBZ), we investigated the effects of CBZ on acetylcholine (ACh) release and metabolism in rat striatum and hippocampus. Acute administration of effective dose of CBZ (25 mg/kg) increased both striatal and hippocampal extracellular levels of ACh, whereas a supraeffective dose of CBZ (50 mg/kg) did not affect the levels and a toxic dose of CBZ (100 mg/kg) decreased the extracellular ACh levels in both brain regions. Both acute and chronic administrations of CBZ (25 and 50 mg/kg, mg/kg per day) increased intracellular ACh levels in striatum and hippocampus. The striatal intracellular ACh levels were decreased by both acute and chronic administrations of CBZ (100 mg/kg, mg/kg per day), whereas the hippocampal intracellular ACh levels were not affected. The effective CBZ concentration did not affect cholinesterase activity, whereas supraeffective CBZ concentration reduced it weakly. Effective dose of CBZ enhanced ACh release and synthesis; however, supraeffective doses of CBZ reduced ACh release and synthesis without enhancement of ACh degradation, indicating that CBZ has biphasic effects on ACh release and synthesis. Thus, the present findings, the slight stimulation of ACh function by effective dose of CBZ, are involved, at least partially, in the antiepileptic and mood stabilizing mechanisms of action of CBZ. PMID- 10863147 TI - Malignant lymphoma: one name, many diseases. PMID- 10863148 TI - Small B-cell lymphoproliferative disorders: an overview of diagnostic approach. AB - Despite their common origin from the B-cell mature lymphoid system, small B-cell lymphomas/leukaemias represent in fact an heterogeneous group of diseases. Recent advances in immunohistochemistry and molecular techniques have improved our knowledge of the immune system and lymphoid neoplasms. An international consensus has been recently reached among pathologists and clinicians, that recognises clinico-pathological entities which are defined by a combination of morphological, immunophenotypical, genetic and clinical features. In each entity, a range of histological grade and clinical aggressiveness can be encountered. Recognition of these entities, combined with clinical prognostic factors has clinical implications in terms of response to treatment and prognosis. The purpose of this paper is to focus on a practical approach, either clinical or pathological, of the diagnosis of small B-cell lymphoma/leukaemia. PMID- 10863149 TI - Follicular lymphomas--a review of treatment modalities. AB - Follicular lymphoma is the most common low-grade non Hodgkin's lymphoma and represent an homogeneous entity as defined by pathological, molecular and clinical data. This indolent disease is characterised by a slow growth pattern with possible spontaneous regression, is often disseminated but remains incurable with available treatments when disseminated. For localised stages, involved field radiotherapy remains the standard choice but other approaches remain to be investigated. In advanced disease, chemotherapy has been demonstrated to produce high response rates but recent trials with new treatment strategies including interferon and monoclonal antibodies may improve the current situation. In this article, we will review treatment of follicular lymphomas, specially emphasising published phase III trials. PMID- 10863150 TI - The management of adult aggressive non-Hodgkin's lymphomas. AB - Aggressive non-Hodgkin's lymphona include diffuse large B-cell lymphoma, anaplastic large cell lymphona, and different peripheral T-cell lymphomas. An international prognostic index has been developed including age, serum LDH, performance status, and extranodal involvement. For localized aggressive lymphoma, the preferred treatment is 3-4 CHOP and radiation therapy, with a cure rate of 70-80%. For disseminated aggressive lymphoma, current regimens have a cure rate of less than 40%. Innovative strategies, including dose escalation, autologus stem cell support, new drugs, and immunotherapy are being explored to improve these results. PMID- 10863151 TI - Current treatment of Hodgkin's disease. AB - In spite of the fact that Hodgkin's disease (HD) remains still an enigma its management and treatment yield a cure rate of about 80% of all patients. However, this management has two limits: on one side favourable cases which should not be overtreated because of unacceptable side-effects, and on the other side very unfavourable cases which should be treated differently because of a very high rate of failure and/or relapse. Then it becomes necessary to precise as thoroughly as possible these two limits in order to choose the adequate treatment for the patient. Prognostic factors based on patient and disease characteristics allow a relatively exact classification of favourable and unfavourable cases. This distinction in two prognostic groups has therapeutic implications in terms of chemotherapy (regimen, duration) and radiotherapy (extension, doses). Other specific situations have to be considered, e.g. pediatric cases, pregnancy, old age and HIV-infected patients who need an adapted management according to very different situations. PMID- 10863152 TI - Differences in the photogenotoxic potential of two fluoroquinolones as shown in diploid yeast strain (Saccharomyces cerevisae) and supercoiled plasmid DNA. AB - Fluoroquinolones are antibiotics with a potential clinical side effect of phototoxicity and some are suspected to enhance UVA-induced tumorigenesis. The present study was designed to evaluate the recombinogenic and mutagenic potential of two highly photoreactive compounds, lomefloxacin and BAYy3118 when exposed to complete UVA (320-400 nm). In order to possibly increase the sensitivity of the test, we used a diploid mutant (D7-rad3) deficient in nucleotide excision repair and deriving from the tester strain D7 of the yeast Saccharomyces cerevisae. In agreement with previous reports, lomefloxacin had no effect in this system. Moreover, BAYy3118 was highly photocytotoxic and genotoxic especially when yeast cells were incubated in its presence in the dark before exposure to UVA radiation. Both fluoroquinolones were comparable in their ability to photo-induce DNA strand breaks or oxidative damage to purines and pyrimidines in supercoiled plasmid DNA, but agarose gel electrophoresis showed that BAYy3118 photoproducts could tightly interact with supercoiled plasmid DNA while lomefloxacin ones only induced strand breaks. These data suggest that phototoxicity of BAYy3118 was the result of a multistep mechanism: first, local photo oxidative stress is induced and secondly some of the photoproducts exerted genotoxic effects. This work also shows that very simple and complementary in vitro approaches can be very informative in the understanding of drug-induced phototoxicity. PMID- 10863153 TI - Evaluation of DNA damage in exfoliated tear duct epithelial cells from individuals exposed to air pollution assessed by single cell gel electrophoresis assay. AB - The search for relevant target cells for human monitoring purposes has increased during the last few years. Cells such as sperm, buccal or nasal and gastric epithelium are being used. In this study, we report the use of exfoliated tear duct epithelial cells as a potential material for human biomonitoring studies, since these cells are a target for environmental pollutants. We employed the alkaline single cell gel electrophoresis (SCGE) assay to evaluate for differences in the basal level of DNA damage between young adults from the south (exposed mainly to high levels of ozone) and from the north (exposed principally to hydrocarbons) regions of Mexico City. We found an increase in DNA migration in tear duct epithelial cells from individuals who live in the southern part of the city compared to those living in the northern part. Moreover, young people who live in the southwest part of the city with the highest values of ozone presented the highest values of DNA damage. These results show the feasibility of using exfoliated tear duct epithelial cells in human biomonitoring studies. PMID- 10863154 TI - In vivo genotoxicity of 2-amino-3,8-dimethylimidazo[4, 5-f]quinoxaline in lacI transgenic (Big Blue) mice. AB - 2-Amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), a heterocyclic amine found in cooked meat, is a strong mutagen in the Salmonella/microsome assay and was proven to be a hepatocarcinogen in rodents. We used the lacI transgenic (Big Blue(R)) mouse to investigate MeIQx genotoxicity in vivo. lacI mutant frequencies were examined in liver and colon after single intragastric administration of MeIQx (males) or 12 weeks of feeding in the diet (males and females). Micronucleus induction was monitored in the peripheral blood and cell proliferating activity was monitored by proliferating cell nuclear antigen (PCNA) immunostaining, but only after the intragastric administration. Intragastric treatment with MeIQx (100 mg/kg) did not increase mutant frequency (MF) in liver or colon but it did induce a slight but statistically significant increase in the incidence of micronucleated reticulocytes 48 h after the treatment. No apparent increase in PCNA-positive foci was observed in any of tissues analyzed 14 days after the treatment. Administration of MeIQx (300 ppm) in diet for 12 weeks, however, caused MF increases in liver and colon in male and female mice, with greater increases in the females. An increase was also obvious after 4 weeks, but only in females. The sex difference in MF is consistent with the fact that female mice are more susceptible to MeIQx carcinogenesis. These results demonstrated that in the transgenic mouse mutation assay, long-term feeding of MeIQx was more effective than single gastric exposures in revealing the compound's mutagenicity in the target organs of carcinogenicity and that sex differences in susceptibility can also be observed. PMID- 10863155 TI - Radioprotective effects of the thiols GSH and WR-2721 against X-ray-induction of micronuclei in erythroblasts. AB - The frequency of micronucleated polychromatic erythrocytes (MNPCEs) was assessed in the bone marrow and peripheral blood of adult male Swiss mice treated with reduced glutathione (GSH) and S-2-/3-aminopropylamino/ethyl phosphorothioic acid (WR-2721), at a dose of 400 mg/kg body weight, and exposed to 6 Gy X-rays. GSH or WR-2721 was applied alone, or 60 and 30 min, respectively, prior to X-ray exposure. The number of MNPCEs was determined at 24 h after the thiol treatment and X-irradiation. The radioprotection and toxicity caused in the mouse erythroblasts by GSH and WR-2721, as indicated by the number of MNPCEs were dependent on the thiol applied. The stronger radioprotective effect is obtained following WR-2721 administration than after GSH application. WR-2721 showed greater toxicity than GSH. The combination of GSH and WR-2721 given before X-ray exposure resulted in the most radioprotective effect as compared to the respective single-drug treatment of mice. Application of the both thiols, without subsequent X-irradiation appeared to be the most toxic, compared with administration of WR-2721 or GSH alone. The effective radioprotection by the combined action of GSH and WR-2721 against genomic instability induced in the mouse erythroblasts by X-rays was shown. PMID- 10863156 TI - CC to TT mutation in the mitochondrial DNA of normal skin: relationship to ultraviolet light exposure. AB - We have previously reported that ultraviolet (UV)-specific (CC to TT) mutations in p53 gene can be detected in normal skin. This, however, cannot be used as a cumulative marker of UV exposure, since cells with the p53 mutation acquire a clonal growth advantage. Moreover, a large skin biopsy is necessary for each assay. In order to circumvent these problems, we have measured mitochondrial (Mt) DNA mutations; there are more than 1000 copies of the Mt genome per cell, and Mt genes are not directly involved in cell growth. We have established a sensitive allele-specific polymerase chain reaction (AS-PCR) assay capable of detecting one CC to TT mutation in Mt DNA among 10(7) wild-type genes using a mismatch allele specific primer. With this assay, we found no mutation-positive samples from internal non-exposed tissue (stomach, colon, and blood) (0/50). In contrast, 17 out of 111 skin samples were positive: the mutation frequency in positive samples was around 10(7)-10(-6) (10-100 copies of mutant in 10(8) wild-type Mt DNA). In normal skin tissue, the prevalence of positive samples was higher in those from exposed sites (13/51) than in those from less-exposed sites (1/26) (p<0.05). However, a quantitative correlation between sunlight exposure and the accumulation of mutations was not found. We conclude that the UV exposure associated CC to TT mutation in Mt DNA can be detected in normal skin, but that further studies are required to develop this as a quantitative marker for UV exposure. PMID- 10863157 TI - Mutational spectra at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus in T-lymphocytes of nonsmoking and smoking lung cancer patients. AB - Molecular analysis of mutations at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus in peripheral blood T-lymphocytes can provide information on mechanisms of somatic in vivo mutation in populations exposed to exogenous carcinogens and in individuals with inherent susceptibility to cancer and other diseases. To study possible mutational changes associated with smoking as a risk factor for lung cancer, we analyzed HPRT mutations in T cells of newly diagnosed, nonsmoking and smoking lung cancer patients before treatment. Reverse transcriptase polymerase chain reaction (RT-PCR) and DNA sequencing methods were used to identify 146 independent mutations, 73 each from 32 nonsmoking and 31 smoking cases. In 35 T-cell mutants, the HPRT cDNA showed loss of an entire exon, indicating a splicing mutation. Among the remaining 111 fully characterized mutations in the coding region, single base pair (bp) substitutions predominated with 79% (48/61) in nonsmokers and 90% (45/50) in smokers. Frameshift and small deletion (1-24 bp) mutations were found in 18 mutants. The distribution of base pair substitutions was nonrandom, with significant clustering at previously identified hotspot positions 143, 197 and 617 in the HPRT coding sequence (P< or =0.008). One additional hotspot, GC-->TA at position 606, was observed only in smokers (P=0.006). The frequency of GC>TA transversions was higher in smokers (13%) than in nonsmokers (6%). Conversely, smokers had a lower frequency of GC>AT transitions (24%) than nonsmokers (35%). This smoking-associated shift of the HPRT mutational spectrum, although not statistically significant, is consistent with the in vitro mutagenicity of benzo(a)pyrene (BaP), a prominent carcinogen of tobacco smoke, and with known differences in the TP53 mutational spectrum in lung tumors of smokers and nonsmokers. Among nonsmokers, the HPRT mutational spectra in healthy population controls and lung cancer patients were similar, but there was a marginally significant difference (P=0.07) in the distribution of base pair substitutions between smoking controls and patients. These results suggest that (i) general mechanisms of somatic mutagenesis in individuals with possible predisposition to cancer (e.g. nonsmoking lung cancer patients) are not different from those in normal healthy individuals, and (ii) the HPRT gene in T-cells is a useful reporter locus for smoking-associated somatic in vivo mutations occurring early in lung cancer development. PMID- 10863158 TI - Genotypic mutation analysis in the p53 gene of benzo[a]pyrene-treated European flounder (Platichthys flesus). AB - We have applied a genotypic mutation detection system (the Restriction Site Mutation (RSM) assay) to detect mutations in the marine teleost flounder (Platichthys flesus). The aim of this study was to evaluate this species as an environmental indicator of genotoxic exposure. We have used the model genotoxin benzo[a]pyrene (B[a]P) to determine the limits of mutation detection in the p53 gene of flounder liver DNA. This study has revealed two important findings. Firstly, we were able to demonstrate that a polymorphism exists in the TaqI restriction site of exon 8 of the flounder p53 gene at codon 243. This polymorphic allele was present as a heterozygote at a mean frequency of 15%, whereas 85% carried the homozygous wild type sequence. Secondly, we established that B[a]P treatment resulted in specific mutational events at the adenine base of the same TaqI site, contrasting previous reports stating that there was a guanine preference for this chemical in mammalian DNA. This difference in mutation specificity may possibly be accounted for by sequence specific factors or by species differences in metabolic activation and/or DNA repair and are worthy of further study. PMID- 10863159 TI - In vivo DNA damage in gastric epithelial cells. AB - A number of risk factors have been linked epidemiologically with gastric cancer, but studies of DNA damage in gastric epithelial cells are limited. The comet assay is a simple technique for determining levels of DNA damage in individual cells. In this study, we have validated the comet assay for use in epithelial cells derived directly from human gastric biopsies, determined optimal conditions for biopsy digestion and investigated the effects of oxidative stress and digestion time on DNA damage. Biopsies taken at endoscopy were digested using combinations of pronase and collagenase, ethylenediaminetetra-acetic acid (EDTA) and vigorous shaking. The resultant cell suspension was assessed for cell concentration and epithelial cell and leukocyte content. A score for DNA damage, the comet %, was derived from the cell suspension, and the effect of various digestion conditions was studied. Cells were incubated with H(2)O(2) and DNA damage was assessed. Pronase and collagenase provided optimum digestion conditions, releasing 1. 12x10(5) cells per biopsy, predominantly epithelial. Of the 23 suspensions examined, all but three had leukocyte concentrations of less than 20%. The comet assay had high inter-observer (6.1%) and inter-assay (4.5%) reproducibility. Overnight storage of the biopsy at 4 degrees C had no significant effect on DNA migration. Comet % increased from a median of 46% in untreated cells to 88% in cells incubated for 45 min in H(2)O(2) (p=0.005). Serial 25-min digestions were performed on biopsies from 13 patients to release cells from successively deeper levels in the crypt. Levels of DNA migration were significantly lower with each digestion (r=-0.94, p<0.001), suggesting that DNA damage is lower in younger cells released from low in the gastric crypt. The comet assay is a reproducible measure of DNA damage in gastric epithelial cells. Damage accumulates in older, more superficial cells, and can be induced by oxidative stress. PMID- 10863160 TI - Analysis of K-ras and p53 mutations in mesotheliomas from humans and rats exposed to asbestos. AB - Malignant mesothelioma is known to be associated with asbestos exposure. However, the mechanism of mesothelial carcinogenesis in relation to the activation of proto-oncogenes or inactivation of tumor suppressor genes remains unclear. In this study, the PCR-Primer Introduced Restriction Site (PCR-PIRS) assay was employed to examine mutations in the K-ras proto-oncogene in mesothelioma tissues from workers exposed to asbestos and from rats treated with asbestos. Mutations in exons 5-8 of the p53 tumor suppressor gene were determined by direct DNA sequence analysis. Results of the PCR-PIRS analysis revealed no mutations in codons 12, 13 or 61 of the K-ras gene in any of the 17 human or 22 rat mesothelioma tissue samples. These results were confirmed by direct DNA sequence analysis. No mutations were found in exons 5-8 of the p53 gene in any of the mesothelioma tissue samples analyzed. These results and the results reported by others indicate that the K-ras proto-oncogene and p53 tumor suppressor gene may not play a critical role in the induction of mesothelioma by asbestos either in humans or in rats. PMID- 10863161 TI - Women, heart diseases and stroke. PMID- 10863162 TI - First International Conference on Women, Heart Disease and Stroke: science and policy in action. PMID- 10863163 TI - Toys for work. PMID- 10863164 TI - Stress testing after percutaneous coronary interventions. PMID- 10863165 TI - Functional testing after percutaneous transluminal coronary angioplasty in Canada and the United States: a survey of practice patterns. AB - BACKGROUND: Authorities recommend various strategies to identify restenosis in patients who have undergone percutaneous transluminal coronary angioplasty (PTCA). Some authorities recommend a routine functional testing strategy, while others recommend a clinically driven strategy. MATERIALS AND METHODS: To examine the patterns of use of post-PTCA functional testing, 89 directors of cardiac catheterization laboratories in Canada and the United States were surveyed. RESULTS: Demographic characteristics of the Canadian and American respondents were similar, including median age (43 and 45 years, respectively) and median number of PTCAs performed each year (200 each). Canadians were more likely to employ a routine functional testing strategy than Americans (62% versus 38%), while Americans were more likely to employ stress imaging studies than Canadians (49% versus 35%). Overall, close to half (44%) of all the cardiologists employed a routine functional testing strategy. Physicians who employed a routine functional testing strategy performed the first functional test a median of three months after PTCA and the second a median of six months after PTCA. Both Canadian and American cardiologists tended to underestimate the incidence of restenosis after PTCA (33% without a stent and 18% with a stent) and to overestimate the sensitivity of exercise treadmill testing for the detection of restenosis (63%). CONCLUSIONS: The use of functional testing after PTCA varies widely. Canadian cardiologists are more likely to employ a routine functional testing strategy than American cardiologists. Close to half of the cardiologists surveyed employed a routine functional testing strategy. These results indicate that there is little consensus regarding the use of functional testing after PTCA. PMID- 10863166 TI - Stentless freestyle aortic valve/root bioprostheses: a northern Ontario community hospital perspective. AB - OBJECTIVE: To examine the controversial issues of postoperative aortic insufficiency (AI), operative mortality and length of hospital stay (LOS) following stentless Freestyle aortic valve replacement (AVR). SETTING: All surgeries were performed in a small northern community hospital (Sudbury Regional Hospital, Sudbury, Ontario). DESIGN: Retrospective study of all stentless AVRs and all stented AVRs from May 1996 to December 1998, and isolated coronary artey bypasses (CABGs) done in 1996/97. PATIENTS: Patients were not selected. All consecutive patients requiring bioprosthetic AVR during this period, regardless of risk, complexity or urgency, were included. In total, 112 stentless AVRs, 138 stented AVRs and 432 isolated CABGs were examined. MAIN RESULTS: AI was rare following stentless AVRs: no significant valvular AI and only 0.9% significant paravalvular AI occurred. The incidence of AI was significantly greater with continuous than with interrupted proximal suturing (P=0. 016). No valve thromboses, thromboemboli or structural failure occurred during 3.8 years of follow-up of the stentless AVRs. The LOS for stentless AVRs was no longer than for stented AVRs or for isolated CABGs. No significant difference was found in the operative mortality following stentless and stented AVRs. The early mortality rate of 1.8% for stentless AVRs was not affected by preoperative risk strata, complexity or urgency. CONCLUSIONS: Early morbidity with stentless AVRs was comparable and LOS was no longer than with stented AVRs. The use of Freestyle bioprosthesis in itself did not result in greater operative mortality, regardless of risk, complexity or urgency of the procedure. Consistent reproducible techniques and experience improve postoperative outcome. It is essential that potential users carefully learn safe and effective surgical techniques to avoid adverse outcomes during the learning curve. PMID- 10863167 TI - Endoscopic saphenectomy for coronary artery bypass surgery: comparison of two techniques with and without carbon dioxide insufflation. AB - OBJECTIVE: To compare the clinical results of an initial experience with two techniques of endoscopic saphenectomy with and without gas insufflation. DESIGN: A retrospective study was performed between September 1998 and March 1999 on 40 patients who underwent endoscopic saphenectomy for coronary artery bypass graft without (group 1, n=15) and with (group 2, n=25) carbon dioxide insufflation. INTERVENTIONS: In both groups, the site of harvesting was at the knee through a 2 cm incision. In group 1, dissection was performed using a hand-held dissector while in group 2 dissection was performed after ensuring that there was a seal at the knee and insufflation of carbon dioxide. Collaterals were controlled with an endoclipper in group 1 and bipolar scissors in group 2. Intraoperative procedure time, length of the harvested vein and aspect of the thigh (ecchymosis, hematoma, infection) were recorded. RESULTS: Vein trauma occurred in four patients in group 1 (four of 15, 27%) and in one in group 2 (one of 25, 4%). Hematomas developed in four patients in group 1 (four of 15, 27%) and in one patient in group 2 (one of 25, 4%). Wound infection occurred in no patients in group 1 and in one patient in group 2. One patient in group 2 suffered carbon dioxide embolism with no untoward consequences. Conversion to an open technique was necessary in five patients in group 1 (five of 15, 33%) and in two patients in group 2 (two of 25, 8%). CONCLUSIONS: Endoscopic saphenectomy both with and without carbon dioxide insufflation is associated with a low infection rate, but vein trauma and wound hematomas are more common without carbon dioxide insufflation. PMID- 10863168 TI - Planning for cardiac surgical services: advice from an Ontario consensus panel. For the Consensus Panel on Cardiac Surgical Services in Ontario and the Steering Committee of the Cardiac Care Network of Ontario. AB - The Cardiac Care Network of Ontario (CCN) Consensus Panel on Cardiac Surgical Services drew on the literature and its own expertise to recommend guidelines for expanding services. This report, which is not an official position paper of the Canadian Cardiovascular Society, presents these recommendations. Rates of surgery are linked to diagnostic capacity, requiring increases in interventional therapies to match increases in invasive diagnostic activity. For quality and efficiency, panel members recommend an annual minimum of 150 procedures per surgeon and 500 per centre; a centre should serve a minimum population of 500,000. Services should be as close to patients' homes as possible while maintaining recommended volumes. Expanding the CCN's cardiac surgery database to include other cardiac modalities will yield a more accurate assessment of waiting times. The panel recommends collaborative regional planning associations, mentorship arrangements between new and existing centres, prompt action on human resource shortages and exploration of alternative funding models. PMID- 10863169 TI - Metabolic syndrome X: a review. AB - Metabolic syndrome X is a multifaceted syndrome, which occurs frequently in the general population. It is more common in men than in women. A large segment of the adult population of industrialized countries develops the metabolic syndrome, produced by genetic, hormonal and lifestyle factors such as obesity, physical inactivity and certain nutrient excesses. This disease is characterized by the clustering of insulin resistance and hyperinsulinemia, and is often associated with dyslipidemia (atherogenic plasma lipid profile), essential hypertension, abdominal (visceral) obesity, glucose intolerance or noninsulin-dependent diabetes mellitus and an increased risk of cardiovascular events. Abnormalities of blood coagulation (higher plasminogen activator inhibitor type 1 and fibrinogen levels), hyperuricemia and microalbuminuria have also been found in metabolic syndrome X. This review summarizes the present knowledge of abnormalities in this syndrome. Each risk factor is reviewed, and potential criteria for diagnosis and therapeutic targets are discussed. Because patients with metabolic syndrome X accumulate cardiac risk factors, they should be given special attention in terms of diagnosis and treatment. PMID- 10863170 TI - Qualitative methods add quality to cardiovascular science. AB - Qualitative research methods are systematic approaches to knowledge development that do not involve quantification. Such methods are used widely in nursing to understand the lived experience and the socioeconomic contexts of cardiovascular health and illness. The evolution of qualitative methods through three phases - traditional, modern and postmodern - frames the discussion. Three common qualitative methods - ethnography, phenomenology and grounded theory - are presented. Applications of qualitative methods have contributed to an understanding of such phenoma as adjustment after myocardial infarction, the experience of an acute cardiac event, the experience of chronic heart disease, the family and the cardiac illness experience, family adjustment to heart transplantation, physical activity practices of working women and family influence on individual health-related decisions in response to heart health intiatives. Postmodern approaches are briefly discussed. Participatory research is presented as an example of a postmodern approach to knowledge development. It is posited that interdisciplinary and multimethod cardiovascular research programs that incorporate qualitative methods will strengthen knowledge development in the cardiovascular field by contributing to an understanding of complex issues related to cardiovascular health and illness for individual persons, families, communities and populations, and to program and policy development. PMID- 10863172 TI - Integrity of the peer review process. PMID- 10863171 TI - Diagnosis and treatment of hypertension in children and adolescents. AB - OBJECTIVE: To review the current recommendations regarding the diagnosis and treatment of hypertension in children to provide education to practitioners. DATA SOURCES AND STUDY SELECTION: Review of recent recommendations of the Task Force on Blood Pressure Control in Children and the current literature, specifically regarding ambulatory blood pressure monitoring in children. In general, search criteria were restricted to studies with a primary focus of blood pressure for subjects 18 years of age or less, focusing on important studies of the past 20 years. DATA SYNTHESIS: Optimal determination of blood pressure in children requires use of appropriate technique, particularly the use of an appropriately sized cuff, and then comparison with normal values based on age, sex and height. Ambulatory blood pressure monitoring is a research tool that, in selected high risk patients, may facilitate detection of occult hypertension. Careful clinical assessment is the key tool for identifying secondary causes or a predisposition to primary hypertension, with laboratory testing reserved if a specific underlying cause is suspected. Management is directed at secondary causes, and general cardiovascular risk reduction is aimed at dietary modification, increased exercise and attainment or maintenance of ideal body weight. Institution of drug therapy depends on the degree of hypertension and the risk of future end-organ damage or cardiovascular disease. CONCLUSIONS: Hypertension is an under recognized clinical entity in children. Studies are needed to define the mechanisms and magnitude of cardiovascular risk, the role of ambulatory blood pressure monitoring, and the efficacy and safety of drug therapy. PMID- 10863173 TI - [Publish and perish] [In Process Citation] PMID- 10863174 TI - [Scientific council of the French Society of Dermatology]. PMID- 10863175 TI - [From hygiene to prevention of nosocomial infections: an ambiguous adventure]. PMID- 10863176 TI - [Intraepithelial carcinoma and invasive carcinoma of the vulva, vagina and penis in Ile-de-france. Enquete PETRI on 423 cases]. AB - OBJECTIVE: Precancerous and invasive carcinoma of the external genitalia and of the vagina are rare tumors and their incidence is not very well known in the Paris region. The objective of this study was to evaluate the frequency of precancerous and invasive lesions of the vulva, the vagina and the penis as well as their variation according to age. METHODS: A prospective study was conducted implicating private and public pathology laboratories in Paris and the seven departments around. Four hundred and twenty three genital biopsies have been analyzed: 160 from the vulva, 151 from the vagina and 112 from the penis. RESULTS: The mean age of the patients was 45 years. The highest frequency of genital biopsies was similar for the three anatomical sites and concerned patients of 25-34 years old. intraepithelial neoplasias represented 77p. 100 of the biopsies (32p. 100 of low grade and 45p. 100 of high grade), invasive squamous carcinoma and adenocarcinoma represented 21p. 100 and 2p. 100 of cases, respectively. The mean age of the patients with low grade vulvar intraepithelial neoplasia, low grade vaginal intraepithelial neoplasia and low grade penile intraepithelial were 34, 40 and 33 years old, respectively. An interval of three to seven years separates the mean age of low grade intraepithelial neoplasia from the mean age of high grade. High grade intraepithelial neoplasia present a peak of frequency in the same class of age for the three localizations (25-34 years) and the risk of developing a high grade intraepithelial neoplasia of the external genital was higher between 25 and 35 years and between 35-45 years of the vagina. The mean age of invasive vulvar carcinoma, vagina carcinoma and penile carcinoma was 62, 59 and 68 years old, respectively. CONCLUSIONS: The correlation between the development of intraepithelial neoplasia of the vulva and the penis supposes a common aetiologic factor in the majority of the cases. The diagnosis of a intraepithelial neoplasia implies a clinical, colposcopic and follow-up of the entire genital area. PMID- 10863177 TI - [Contact dermatitis due to ethyl alcohol: how to perform patch tests?]. AB - INTRODUCTION: Ethyl alcohol sensitization is rare and can induce immediate contact urticaria or delayed eczema. Patch tests performed with ethanol can provoke an irritative reaction and are not well codified. CASE-REPORTS: We report on 4 cases of contact dermatitis due to alcohol in 4 women. Eczema was due to alcohol contained in the reservoir of a transdermal transfer system with estrogens in 2 cases and related to the application of alcoholized antiseptic lotions in the other 2 cases. In 2/4 cases we observed a co-sensitization with corticosteroids. METHODS: Patch tests were performed with alcohol at 95 degrees diluted at 70 p. 100 and at 10 p. 100 in water and read after 20 minutes then on day 2 and day 4. RESULTS: Positive results were obtained in 4/4 cases when alcohol was tested diluted at 70 p. 100 in water and in 3/4 cases when diluted at 10 p. 100 in water. No irritant reaction was observed in 140 negative controls. DISCUSSION: Immediate after 20 minutes and delayed readings of patch tests performed with ethyl alcohol diluted at 70 p. 100 seem to be convenient in diagnosing cutaneous delayed hypersensitivity to alcohol. Co-sensitization between corticosteroids and alcohol could be due to an aldehyde deshydrogenase deficiency. PMID- 10863178 TI - [Primary and isolated cutaneous lymphomatoid granulomatosis following heart-lung transplantation]. AB - BACKGROUND: Lymphomatoid granulomatosis is an Epstein-Barr virus-associated B cell lymphoproliferative disease. It is angiocentric and angiodestructive and involves the lungs, central nervous system and skin. Exclusive cutaneous involvement is rare and may be associated with a better outcome. Contrarily to the extra-cutaneous forms of lymphomatoid granulomatosis, it is difficult or impossible to detect Epstein-Barr virus DNA sequences in primary and isolated cutaneous lymphomatoid granulomatosis. CASE REPORT: A 54-year-old woman developed erythemato-violaceous lesions on both legs 3 years after a heart-lung transplantation. The diagnosis of erythema multiforme and of drug-induced vasculitis were first made. Because of fever and of the rapid extension of the lesions, the patient was hospitalized. The histologic examination of the first lesions showed a perivascular infiltrate, without epidermotropism, composed of histiocytes, lymphocytes and plasma cells. Immunohistochemistry revealed the presence of a predominantly T-cell infiltrate with some large B cells. Subsequent biopsies were diagnosed as high grade B-cell lymphoma. Polymerase chain reaction analysis as well as in situ hybridation study showed the presence of Epstein-Barr virus load in the lesions. There was however no serologic evidence of viral reactivation. Extensive systemic evaluation revealed no visceral or bone marrow involvement. Despite antiviral treatment and CHOP polychemotherapy, the patient died 3 months after her admission. DISCUSSION: This observation of lymphomatoid granulomatosis is particular because of its exclusive cutaneous involvement associated with a fulminant evolution to high grade B lymphoma. The presence of a context of iatrogenic immunosuppression underlies the role of altered immune cellular functions in the initiation and/or progression of lymphomatoid granulomatosis and strengthens the role of a viral agent in its pathogenesis. We suggest that the presence of Epstein-Barr virus, which is generally not associated with the isolated cutaneous forms of lymphomatoid granulomatosis, may have played a role in this fulminant evolution to high grade B lymphoma. PMID- 10863179 TI - [Chronic lymphocytic leukemia revealed by atypic pemphigus]. AB - BACKGROUND: Bullous diseases are occasionally reported during chronic lymphocytic leukemia. We report the case of a woman in which a bullous disease of difficult nosologic classification has revealed chronic lymphocytic leukemia. CASE REPORT: A 57-year-old woman, well-known for a rheumatoid arthritis, developed a bullous eruption. It was associated with voluminous lymphadenopathies, and laboratory investigations revealed a chronic lymphocytic leukemia. Clinicopathological aspect and immunofluorescence studies argued for a paraneoplastic pemphigus but immunoblot showed only antibodies against desmoglein 1. DISCUSSION: Paraneoplastic pemphigus is an autoimmune bullous disease defined by the criteria of Anhalt, including autoantibodies mostly directed against cytoplasmic proteins of the plakin family. Our case is unusual for the absence of these antibodies and for the presence of antibodies directed against a cell surface target, desmoglein 1. Such a case confirms the overlapping auto-immune responses in pemphigus. PMID- 10863180 TI - [Localized bullous eruption after intravenous injection of aciclovir: toxic or immunoallergic mechanism?]. AB - OBJECTIVE: To report a case of bullous eruption at and far from the site of aciclovir injection. CASE REPORT: A 50-year-old man was treated with intravenous aciclovir for Herpes simplex meningoencephalitis. Ten days after treatment onset, blisters appeared on his right forearm, at and far from the site of aciclovir injection. DISCUSSION: This adverse effect has not been frequently reported. To date, bullous eruptions were considered to result from extravasation of the aciclovir solution. In this case, an immunoallergic pattern was discussed with the presence of a histological leukocytoclastic vasculitis. PMID- 10863181 TI - [GAPO syndrome]. AB - INTRODUCTION: The GAPO syndrome is a rare but distinct genetic disorder. GAPO is an acronym for the manifestation of Growth retardation, Alopecia, Pseudoanodontia and Optic atrophy. The syndrome was first reported in 1947; to date, 24 cases have been reported. We report the first Tunisian case. OBSERVATION: We studied a 12 year-old boy with GAPO syndrome which was associated with peculiar facial appearance, umbilical hernia, hemangiomatous plaques of the neck, depigmented maculae arranged in a splashed pattern located in the trunk and the right upper limb. He had a pulsated mass in the right mastoid area and a bruit was audible, he had a second flaccid mass of the vertex. These tumefactions correspond to very developed commissure veins. DISCUSSION: In addition to the classical manifestations of the GAPO syndrome, the patients have a strikingly characteristic facial appearance and may also have umbilical hernia, skin redundance and prominent dilatation of scalp veins. Our case had depigmented maculae suggestive of incontinentia pigmenti achromians. This has never been reported previously. The pathogenesis of this syndrome is unknown and inheritance is considered to be autosomal recessive. PMID- 10863182 TI - [Parvovirus B19 infection mimicking drug-induced hypersensitivity syndrome]. AB - BACKGROUND: Clinical manifestations of primary parvovirus B19 infection vary greatly. Epidermal megalerythema is the most common feature. We report a particular form resembling a drug-induced hypersensitivity reaction. CASE REPORT: A 19-year-old man had a scarlatiniform eruption associated with multiple node enlargement, elevated liver enzymes and a abnormal white cell count with mononucleosis and lymphopenia, similar to that observed in hypersensitivity reactions. Seroconversion and positive PCR search for viral DNA established the diagnosis of primary parvovirus B19 infection. The spontaneous course was favorable with no recurrence at one month. DISCUSSION: The clinical features and laboratory findings in this case of parvovirus B19 infection closely resembled drug-induced hypersensitivity syndrome. The role of viral agents in the development of hypersensitivity reactions have been suggested. It is important to look for viral infections in clinical presentations mimicking drug-induced hypersensitivity. PMID- 10863184 TI - [Scurvy presenting with ecchymotic purpura and hemorrhagic ulcers of the lower limbs]. AB - INTRODUCTION: The risk of vitamin C deficiency is underestimated in industrialized countries and is only disclosed in rare cases of severe scurvy. CASE REPORT: We report three cases of scurvy presenting with ecchymotic purpura and hemorrhagic ulcerations of the lower limbs. Vitamin C supplementation led to rapid improvement of the skin lesions. DISCUSSION: Clinical diagnosis of low grade deficiency can be difficult. Biological diagnosis requires special care in sample taking and transport. PMID- 10863183 TI - [Multiple cutaneous reticulohistiocytosis]. AB - BACKGROUND: Multicentric reticulohistiocytosis is a non Langerhans cell histiocytosis. This rare disease is characterized by cutaneous papules and nodules and a destructive polyarthritis; multisystem involvement may occur. Multiple and diffuse cutaneous reticulohistiocytosis have been more rarely reported. We present a case which was distinctive by the existence of multiple cutaneous plaques. CASE REPORT: A 65-year-old woman presented cutaneous papules and nodules associated with a destructive arthritis affecting the hands. Histological examination of a cutaneous biopsy associated with immunophenotyping and electronic microscopy permitted us to make the diagnosis of multicentric reticulohistiocytosis. The search for visceral involvement or underlying neoplasia was negative. Rapidly, cutaneous aggravation occurred with multiple and diffuse infiltrated plaques on the back, the face, the ears, the thighs and the forearms. The same histological aspect was found for these lesions. Treatment with corticosteroids and cyclophosphamide was successful. DISCUSSION: This case report is the first one with diffuse cutaneous lesions of multicentric reticulohistiocytosis with aspect of infiltrated plaques. Diffuse cutaneous lesions in multicentric reticulohistiocytosis have been rarely reported with diffuse papules or nodules pattern. A visceral involvement seems to be more frequent for diffuse cutaneous involvement. In all cases, the association of multicentric reticulohistiocytosis with neoplasia in up to 25 p. 100 is of interest. Treatment of multicentric reticulohistiocytosis consists in corticosteroids at the initial phase associated with alkylants agents or methotrexate. PMID- 10863185 TI - [Erythema migrans with multiple lesions]. AB - INTRODUCTION: Erythema migrans with multiple lesions is observed in Europe in only 5-20 p. 100 of Borrelia infections. The number of lesions observed simultaneously in one patient is less than 7, considering all the cases published in the literature to date. CASE REPORTS: We report three patients who presented multiple macular, erythematous and centrifugal lesions localized on the trunk and the limbs. The first patient presented about thirty lesions, the second more than seventy, and the third about twenty. Serology investigation and immunoblots allowed to establish the diagnosis of a recent Borrelia Burgdorferi infection, with multiples lesions of erythema migrans. PCR performed on a skin biopsy of one of the patients confirmed the presence of Borrelia burgdorferi. Response to antibiotic therapy was immediate in all 3 patients. DISCUSSION: These 3 cases are of particular interest because the number of simultaneous lesions is much higher than what is reported in the European literature. These observations remind the clinician that the diagnosis of erythema migrans with multiple lesions should be included, in Europe as well, in the differential diagnosis of multiple annular dermatoses. PMID- 10863186 TI - [Mupirocin: its use]. PMID- 10863187 TI - [A pigmented lesion of penis]. PMID- 10863188 TI - [Round, squamous, pigmented plaques]. PMID- 10863189 TI - [Cutaneous manifestations of Lyme borreliosis]. PMID- 10863190 TI - [HHV-8/KSHV. I - epidemiological and molecular aspects]. PMID- 10863191 TI - [Lasers in dermatology]. PMID- 10863192 TI - [Drug eruption: assessment of causality]. PMID- 10863193 TI - Epidermic necrolysis: advances in pathogenesis. PMID- 10863194 TI - [Purple hand syndrome and intravenous phenytoine]. PMID- 10863196 TI - [Research grants offers, 1999] [In Process Citation] PMID- 10863195 TI - [Professor Tzanck's team, in 1931] [In Process Citation] PMID- 10863197 TI - [Speech to "Societe de laryngologie des Hopitaux de paris"] [In Process Citation] PMID- 10863198 TI - [Surgical treatment of severe sleep apnea syndrome by maxillomandibular advancing or mental tranposition]. AB - Surgery of the facial skeleton or the tongue may be envisaged in case of failure of continuous positive pressure ventilation for severe sleep apnea syndrome defined by a apnea-hyponea index greater than 30/h. We present here our results in patients treated by maxillo-mandibular advancing and mental transposition. We define the surgical indications. Between January 1993 and June 1997, 41 patients, mean age 49 years, with severe sleep apnea syndrome (mean apnea-hyponea index =58.5/h) were treated by maxillo-mandibular advancing (21 cases) or mental transposition (20 cases) depending on the cephalometric work-up including lateral teleradiography and sagittal magnetic resonance imaging of the tongue. Functional outcome was good in both groups. Objective success (postoperative apnea-hyponea index <20) was 70.5 % after bimaxillary advancing (mean apnea-hypopnea index =17), but only 25 % after mental transposition (mean apnea-hyponea index =44.5). Maxillomandibular advancing is a major procedure which can be effective in sleep apnea patients with severe craniofacial skeletal anomalies. Its applications in apneic patients with no skeletal anomaly remains a subject of debate. Conversely, there would appear to be very few indications for mental transposition. PMID- 10863199 TI - [Pharyngo-esophagoplasty by right coloplasty for the treatment of post-caustic pharyngo-laryngeal-esophageal burns: a report of 13 cases]. AB - We report a new technique of pharyngoesophagoplasty by right coloplasty, indicated in postcaustic severe pharyngeal stenosis, analyzing the anatomical and functional results in terms of respiratory tract and phonation as well as digestive tract outcome. We compared our results with those obtained with other procedures. Between March 1995 and September 1998, pharyngoesophagoplasty by right coloplasty was performed in 13 patients. All had severe hypopharyngeal stricture with total obliteration of the two piriform sinuses and the upper esophageal sphincter. Nine patients underwent emergency esophagectomy or esogastrectomy by stripping and four had a cicatricial esophagus. Eight patients underwent tracheotomy before the pharyngoplasty due to burns of the laryngeal margin or airway infections. There was no perioperative mortality. The retrospective analysis in February 1998 with a follow-up of 22 months disclosed two deaths, one from septic shock following pneumnia and one suicide. For the 11 other patients, respiratory function had been restored successfully as the tracheostomy tube had been removed. Eight of the patients were on regular oral diet and the jejunostomy tube had been removed. Three had mild dysphagia and the oral diet was supplemented by jejunostomy tube feedings. These results were most successful compared with those obtained with 29 other patients. Pharyngo esophagoplasty by right colonic transposition appears to be the method of choice for the reconstruction of post-caustic pharyngeal and esophageal stenosis. PMID- 10863200 TI - [Cricotracheal resection in children: indications, technique and results]. AB - The most significant advance in the surgical treatment of laryngotracheal stenosis has been the changes in external procedures, notably in laryngotracheoplasty aimed at widening the regional stenosis with prosthetic material. In opposition with this therapeutic method, cricotracheal resection which removes the regional stenosis, and a large portion of the cricoid cartilage, has been proven to be a reliable technique in adults. Between June 1993 and June 1998, 10 children underwent cricotracheal resection. There were 5 boys and 5 girls with 9 acquired and 1 congenital stenosis (grade II =5, grade III =2, grade IV =3). At the time of the procedure, the patients' mean weight was 19 kg and mean age was 7.5 years. A tracheotomy present in 5 children prior to the procedure was left in situ postoperatively. In these children a rolled silastic sheet was used to maintain the caliber for 23 days and the tracheotomy canula was removed a mean 58 days later. In the 5 children operated on without tracheaotomy, the nasotracheal tube was removed a mean 2.5 days after the procedure. Mean follow-up was 43 months, with clinical and endoscopic surveillance. No growth retardation was observed among the 5 children presenting 4.5 years after the procedure. The choice between laryngotracheoplasty enlargement and cricotracheal resection is not based on documented evidence but on case-by-case decision making. We discuss here the points which appear to us to be the most relevant in terms of indication, surgical procedure, potential complications, and outcome. PMID- 10863201 TI - [Inner ear in C.H.A.R.G.E. association]. AB - Cochleovestibular dysfunction is one of the major features of C.H.A. R.G.E. association. The inner ear anomalies were studied in a population of 17 children with CHARGE. Temporal bone anomalies were defined with CT scan, hearing loss was evaluated with audiologic procedures appropriate for age, and functional canal and otolith vestibular impairment evaluated with respectively vertical and off vertical axis rotation (OVAR) tests. Temporal bone anomalies appear specific of C.H.A.R.G.E. association. They are characterized by an aplasia of the posterior labyrinth (with an absence of semicircular canals) and a variable degree of anomalies of the anterior labyrinth with sometimes normal cochlea. Hearing loss is variable, often worsened by delayed neurological maturity and frequent association of middle ear effusion. Thus, multiple audiologic evaluations are required at regular intervals. The treatment of middle ear effusion, the association of hearing aid and speech therapy management permit language acquisition in spite of the other multiple sensory handicaps. Vestibular functional evaluation shows a constant canal areflexia but a residual vestibular otolith function (sometimes normal). The deficit of the vestibular function is certainly involved in the delay of posturo-motor development as well as visual deficit and neurological impairment. This can permit an adaption of the physical therapy program for each child to make use of the available sensorial information. PMID- 10863202 TI - [ENT diseases associated with allergic rhinitis: a review of the literature]. AB - Various diseases of the upper airway, such as acute or chronic sinusitis, otitis media, pharyngitis or laryngitis, snoring and sleep apnea syndrom, may be associated with allergic rhinitis. The relationship between these pathologies and the allergic rhinitis has been well established from a clinical and epidemiological point of view, but the pathophysiological mechanisms remain uncertain. A good knowledge of symptoms and the performance of explorations, such as nasal endoscopy for sinusitis, are important in order to take care of these associated diseases. When upper airway diseases are associated with allergic rhinitis, treatment of rhinitis must generally be reinforced. Treament of associated disease will be specific to each disease, and sometimes surgery is required, specially in case of chronic sinusitis. In all cases, the pneumologist, allergologist and ENT physician should work in close collaboration. PMID- 10863203 TI - [Infralabyrinthine approach for cholesterol granuloma of the petrous apex]. AB - Cholesterol granuloma of the petrous apex are cystic lesions, revealed by otologic and/or cranial nerve palsies, and diagnosed with the help of computed tomography and magnetic resonance imaging. Surgical treatment is not the complete removal of the lesion, but a conservative approach requiring drainage of the cyst and re-establishment of a correct aeration of the cavity. Three cases of cholesterol granuloma of the petrous apex were treated through a transmastoid infralabyrinthine procedure. Through a postauricular approach, a simple mastoidectomy was performed. The third portion of the facial nerve was identified. The posterior and lateral semicircular canals, and the jugular bulb were skeletonized. In two cases, a high diverticulum of the jugular bulb was impacted downwards with wax. The cystic lesion was then opened, and evacuated. The opening must be large to permit a correct aeration of the cavity and prevent stenosis of the drainage site. Hearing and the facial function were preserved in all cases. In conclusion, conservative approach to cholesterol granuloma of the petrous apex provides satisfactory drainage of this intrapetrous deep seated lesion with preservation of hearing and facial nerve function. PMID- 10863204 TI - [Assessment of perilymphatic pressure using the MMS-10 tympanic membrane displacement analyzer (Marchbanks' test) in patients with Meniere's disease: preliminary report]. AB - OBJECTIVES: The aim of this study was to investigate non-invasive measurement of perilymphatic pressure using the MMS-10 tympanic displacement analyzer (Marchbanks' test) in patients with Meniere's disease. METHODS: We performed measurements in 20 patients with Meniere's disease and in 9 normal subjects with normal hearing. Data were collected in three groups: healthy ears of normal hearing patients, healthy and affected ears in patients with Meniere's disease. RESULTS: We found no significant differences between the 3 groups for the types of the graph. Measurements of the tympanic membrane displacement test variables (Vi and Vm) showed large inter-subject variations. No significant difference was found between the 3 groups for cochlear aqueduct patency. CONCLUSION: Preliminary results of this short series show that assessment of perilymphatic pressure using the MMS-10 procedure does not seem to be useful in Meniere's disease. PMID- 10863205 TI - [Is there a correlation between optimism and quality of life in upper aerodigestive tract cancer Patients? Preliminary results]. AB - The aim of this study was to test the hypothesis that optimism is a predictor of quality of life (QOL) in a sample of upper aerodigestive tract (UADT) cancer patients. MATERIALS AND METHODS: Consenting patients with squamous cell carcinoma of the UADT were included during the week following disclosure of the diagnosis of their illness, and before the start of treatment. QOL and optimism were evaluated by questionnaires presented to the patients before the start of treatment, after the treatment, and 6 and 12 months after the end of the treatment. PRELIMINARY RESULTS: 92 patients were included. The average age was 58.7+/-11.4 years. Their QOL scores were significantly correlated, first with age (r =- 0.23, p =0.03) and second with degree of optimism (r =0.32, p =0.002). No correlation was found between QOL scores, degree of optimism, and sociodemographic and clinical data. Optimism was the sole variable significantly associated with QOL before treatment (F =4.1, p =0.002, r(2) =0.19). The difference between QOL scores before and after treatment was not significant. CONCLUSION: Continuation of the study and analysis of survival of the patients may help pinpoint new prognostic factors, both objective and subjective, that will facilitate an overall approach to patient care by allowing for their preferences. PMID- 10863206 TI - [Direct foreign body footplate trauma: 3 cases]. AB - 3 cases of direct foreign body footplate trauma are described in children. A surgical exploration was decided on history, cochleovestibular signs and/or CT abnormalities. Hearing improved in all cases after surgery. PMID- 10863207 TI - [Is endoscopic mucosectomy for superficial cancer of the stomach really reasonable?]. PMID- 10863208 TI - [Rectal prolapse]. AB - Rectal prolapse and rectal intussuception correspond to two stages of the same disease. Rectal prolapse is unusual but requires surgical treatment. Abdominal rectopexy is the most effective procedure but increases the risk of postoperative constipation. This risk decreases when the lateral sides are not touched during rectal dissection. The Delorme procedure is associated with a higher rate of recurrence and must be reserved for patients presenting a high risk of postoperative complications. Rectal intussuception is more frequent and is pathological only when arising in the anal sphincter. Rectal intussuception may lead to solitary rectal ulcer and has in this case to be treated by rectopexy. Rectal intussuception involvement in terminal constipation is not yet proved. Internal mucosectomy seems to be the best treatment for terminal constipation. PMID- 10863209 TI - [Surgical treatment of obesity by a laparoscopically inserted gastric ring]. PMID- 10863210 TI - [Laparoscopic appendicectomy]. PMID- 10863211 TI - [Anal fistula]. PMID- 10863212 TI - [Hernia of the Jean-Louis Petit triangle]. PMID- 10863213 TI - [Patients, surgeons and surgical gloves]. AB - Consumption of surgical gloves in progressing constantly. All proposed products do not have the same qualities in terms of protection, comfort, and safety for the surgeon, the surgical team and the patient. Latex is the basic material used to manufacture surgical gloves even if it does raise the problem of side-effects and requires use of starch powder for lubrification. The surgeon should be aware of the different products and participate in choosing this indispensable tool. There is a general and advisable trend to using non-powdered gloves. PMID- 10863214 TI - [45-year-old women from l'Ile de la Reunion]. PMID- 10863215 TI - [Inguinal hernia, eternal subject of controversy]. PMID- 10863216 TI - [Benign florid corticoadrenaloma]. PMID- 10863217 TI - Cutaneous manifestations of alcohol abuse. AB - Alcohol consumption and abuse can have a variety of cutaneous manifestations. In addition to the well-recognized stigmata of the chronic alcoholic patient, even early abuse can result in distinctive skin changes or exacerbate existing cutaneous disorders. An accurate history of alcohol intake will facilitate recognition of these alcohol-induced cutaneous disorders and treatment resistance of dermatoses such as psoriasis as well as help decrease morbidity in surgical procedures. Familiarization with the spectrum of cutaneous manifestations of alcohol abuse and alcoholic liver disease can also allow for early detection and treatment in an attempt to minimize the medical consequences. We review the medical literature and discuss the spectrum of dermatologic disease associated with the use and abuse of alcohol. PMID- 10863218 TI - Immunofluorescent microscopic investigation of the distal arrector pili: a demonstration of the spatial relationship between alpha5beta1 integrin and fibronectin. AB - Currently there is limited knowledge regarding the anatomy of the distal arrector pili (AP) muscle. A previous study implicated fibronectin and alpha5beta1 integrin binding as the anchor between the AP and the extracellular matrix (ECM). The purpose of this study was to strengthen this hypothesis. Serial frozen sections of human scalp skin were double-labeled via immunofluorescent staining for alpha5beta1 with fluorescein and fibronectin with rhodamine, followed by fluorescent microscopy. Granular staining for alpha5beta1 with fluorescein and smooth staining for fibronectin with rhodamine were seen at the periphery of the AP muscle bundles and along the distal fibers. Precise co-localization of alpha5beta1 and fibronectin was observed at the AP-ECM interface by means of a dual filter. Analysis of variance was used on the relative density of staining for each epitope. Staining for both epitopes was significantly brighter at the distal fibers than at the middle or proximal portions of the muscle. A computerized three-dimensional reconstruction provides a detailed picture of the microanatomy of the distal AP, which allows mathematical evaluation of the forces of contraction. The anatomic co-localization between alpha5beta1 and fibronectin strengthens our hypothesis that interaction of these epitopes mediates the attachment of the distal AP to the ECM. PMID- 10863219 TI - Effect of 60 mg twice-daily fexofenadine HCl on quality of life, work and classroom productivity, and regular activity in patients with chronic idiopathic urticaria. AB - The effect of 60 mg twice-daily fexofenadine HCl on health-related quality of life and productivity at work, in the classroom, and during daily activities in patients with moderate to severe chronic idiopathic urticaria symptoms was studied in two identical, 4-week, placebo-controlled, multicenter clinical trials. Patients self-administered the Dermatology Life Quality Index (score, 0 30) and the Work Productivity and Activity Impairment instrument (0%-100%). In both trials, improvements in Dermatology Life Quality Index scores in fexofenadine-treated patients (N = 169) were statistically significant compared with placebo (P < or =.0002). Similarly, improvements in productivity scores with fexofenadine 60 mg twice daily were statistically superior to placebo at work (n = 120, P < or =.0152) and in performance of daily activities (n = 166, P < or =.0002). There was a trend toward improved classroom productivity (n = 26) with fexofenadine. We conclude fexofenadine 60 mg twice daily improves health-related quality of life, increases work productivity, and improves performance of daily activities in patients with moderate to severe chronic idiopathic urticaria. PMID- 10863220 TI - Contact sensitivity to nickel and other metals in jewelry reactors. AB - BACKGROUND: Many patients who give a history of a dermatitis reaction to jewelry or metal contact with skin are negative to metals on standard patch testing. Some may be showing false-negative reactions. OBJECTIVE: Our purpose was to determine whether patients with a history of jewelry reactions but whose standard patch tests were negative have a false-negative reaction or are allergic to metals other than nickel, cobalt, or chromium. METHODS: Four hundred forty-nine patients were studied who gave a history of reacting to jewelry or metal. Of these, 210 were tested to the metals in the European standard series (ie, nickel, cobalt, and chromate), and 239 were tested to the metals in the standard series and to an extended metal series of palladium, gold, platinum, a second nickel salt, and a nickel/cobalt mixture. These were compared with 752 patients who did not give a history of jewelry or metal reactions, of which 50, besides the standard series, were also treated with the additional metal series. RESULTS: A higher proportion of jewelry-reactive patients tested with the extended series reacted to nickel (and to other metals) than those who were tested only with the European standard series: (61% vs 38%; P <.0001 ). The use of the extended series showed that palladium allergy was common, present in 34% of nickel-allergic patients, but it always occurred with nickel sensitivity. Gold allergy coexisted with nickel sensitivity in 10% of cases. Testing simultaneously with separate patches containing 5% nickel sulfate and 5% nickel chloride showed a concordance of 71% in identified nickel-sensitive patients. Nickel sulfate was more likely than nickel chloride to detect nickel sensitivity. The use of a combined preparation of 2.5% nickel sulfate and 0.5% cobalt chloride in petrolatum revealed only 3 jewelry-reactive patients who were negative to other metals. There was a slightly higher proportion of atopic patients in the patch test-negative jewelry reactors group than in the positive group; however, the difference was not significant and it was not sufficient to account for the negative findings. CONCLUSION: Some jewelry reactors who had negative patch tests are likely to be subclinically allergic to nickel. We suggest that the higher number of antigens, or perhaps the larger nickel load, in the extended metal series, resulted in a larger proportion of patients reacting. To better demonstrate nickel allergy in jewelry reactors, patients should be patch tested to a metal series that contains palladium and gold salts and perhaps a second nickel patch because these may reveal the presence of nickel sensitivity when standard patch tests would otherwise have been negative. PMID- 10863221 TI - Time-sequence histologic imaging of laser-treated cherry angiomas with in vivo confocal microscopy. AB - OBJECTIVE: To chronicle the pathophysiologic changes that occur subsequent to laser treatment of vascular lesions, we used a confocal scanning laser microscope that yields high-resolution microscopic images of skin in vivo. METHODS: Cherry angiomas were treated with the 585-nm flashlamp-pumped pulsed-dye laser (PDL) and the 568-nm continuous-wave krypton laser. Repeated confocal reflectance imaging was performed before and immediately after treatment, as well as after several hours, 1 day, 2 days, 1 week, 2 weeks, 3 weeks, and 4 weeks. RESULTS: Before treatment, confocal images revealed dilated blood vessels ranging from 10 to 50 microm in caliber, closely spaced at 5 to 50 microm apart. After PDL treatment, amorphous cords of refractile material conformed to the shape of the original vessels, followed by dark nonrefractile spaces where the vessels once were. Inflammation and necrosis ensued, with eventual replacement after 3 weeks by normal-appearing skin. After krypton laser treatment, dark nonrefractile spaces appeared immediately, with subsequent inflammation, necrosis, and eventual healing by 4 weeks. CONCLUSION: Confocal laser microscopic imaging elucidates the dynamic pathophysiologic events that occur after laser treatment of vascular lesions and has added insight into the different mechanisms of vessel damage induced by the PDL and krypton laser. PMID- 10863222 TI - Confocal laser microscopic imaging of actinic keratoses in vivo: a preliminary report. AB - BACKGROUND: Real-time near-infrared confocal laser scanning microscopy (CM) offers an unprecedented method for confirming the clinical diagnosis of actinic keratosis (AK) without biopsy. METHODS: Seven patients with clinically diagnosed AK underwent CM imaging over the lesion and over adjacent normal-appearing skin. Biopsy specimens were obtained from the presumed AKs in 4 patients. RESULTS: CM detected lesional pathologic features of hyperkeratosis (71%), lower epidermal nuclear enlargement and pleomorphism (100%), and architectural disarray (57%). In contrast, cytologic atypia and architectural disarray were apparent in one patient (17%) over the adjacent, clinically normal skin. Three of 4 biopsy specimens confirmed the clinical diagnosis of AK, whereas one revealed invasive squamous cell carcinoma. Without optimizing CM for imaging hyperkeratotic skin lesions, the limited depth of penetration reached the stratum basale in only 3 lesions, precluding detection of dermal invasion in the others. CONCLUSION: Depth of penetration currently imposes a major limitation on CM in the diagnosis of AKs, especially in hypertrophic and hyperkeratotic lesions, which are more likely to be malignant. However, CM may become an alternative to biopsy, and its limitations may be overcome by future technologic advances in optical penetration or by simply removing the hyperkeratotic stratum corneum. PMID- 10863223 TI - Elucidating the pulsed-dye laser treatment of sebaceous hyperplasia in vivo with real-time confocal scanning laser microscopy. AB - BACKGROUND: Several case reports document successful treatment of sebaceous hyperplasia with the pulsed-dye laser. Moreover, noninvasive real-time confocal laser scanning microscopy elucidates the vascular nature of these lesions and their pathophysiologic response to treatment mediated by vessel coagulation. METHODS: Ten patients with 29 lesions of sebaceous hyperplasia were treated with 3 stacked 5-mm pulses of the 585-nm pulsed-dye laser at fluences of 7 or 7.5 J/cm(2). Confocal imaging was performed before and immediately after treatment, as well as at 2, 4, and 8 weeks of follow-up. RESULTS: The great majority of lesions responded to one treatment, with complete disappearance in 28%, decrease in diameter in 66%, and flattening in 93%. Although 28% recrudesced after initial involution, only 7% recurred completely. Three lesions became eroded or crusted, and 7 experienced cutaneous depressions before complete healing, but no scarring or pigmentary side effects were noted. Confocal imaging revealed a prominent "crown" of blood vessels surrounding the sebaceous duct and coagulation of these vessels with pulsed-dye laser treatment. However, the vessels reappeared during follow-up, and no noticeable morphologic changes in the sebaceous duct were noted. CONCLUSION: Vascular targeting of sebaceous hyperplasia can be monitored with real-time reflectance confocal microscopy. Most sebaceous hyperplasia regresses after one treatment with 3 stacked pulses of the 585-nm pulsed-dye laser. Whether this response is due to temporary ischemia induced by selective vessel destruction or nonspecific thermal diffusion beyond the vessels from pulse stacking has not been determined. PMID- 10863224 TI - Experience with total skin electron beam therapy in combination with extracorporeal photopheresis in the management of patients with erythrodermic (T4) mycosis fungoides. AB - OBJECTIVE: We compared the prognosis of patients with erythrodermic mycosis fungoides (MF) administered total skin electron beam radiation (TSEB) plus neoadjuvant, concurrent, and adjuvant extracorporeal photopheresis (ECP) with the prognosis of patients administered only TSEB. Outcomes of clinical interest include disease-free survival (DFS), progression-free survival (PFS), overall survival (OS), and cause-specific survival (CSS). METHODS: This study was a retrospective nonrandomized series. Between 1974 and 1997, a total of 44 patients with erythrodermic MF from the Department of Therapeutic Radiology, Yale University School of Medicine, and the Department of Radiation Oncology, Cancer Care Ontario, Hamilton, Ontario, were collected and analyzed as a group (Hamilton = 15, Yale = 29). These patients received TSEB consisting of 32 to 40 Gy via 4 to 6 MeV. Twenty-one patients at Yale also received ECP treatment 2 days per month for a median of 6 months. Median age was 68 years (range, 29-82 years) at the commencement of TSEB, and 66% were male. Seventy-three percent of patients had received other therapies before TSEB, including 75 courses that failed to control disease (n = 15 systemic therapy, 16 biologicals, and 44 topical therapies). At TSEB, 59% had hematologic involvement (B1), 30% were stage IVA (N3), and 13% were IVB (M1). Median follow-up was 2.2 years (range, 0.3-13.9 years) subsequent to TSEB and 3.7 years from diagnosis (range, 0.8-16.8 years). RESULTS: All patients responded to TSEB within 2 months of completion, with a cutaneous complete response rate of 73%. For the 32 complete responders the 3-year DFS was 63%. It was 49% for those 17 patients who received only TSEB compared with 81% for those 15 patients who received TSEB + ECP. Cox regression analysis demonstrated that ECP was associated with prolonged remission (DFS multivariate P =.024, adjusting for B1 and stage). The 2-year PFS, CSS, and OS for the TSEB group were 36%, 69%, and 63%, respectively, compared with 66%, 100%, and 88% for the TSEB + ECP cohort. Cox regression demonstrated that ECP was associated with CSS (multivariate P =.048, adjusting for B1 and stage). For those who progressed, a total of 49 subsequent courses of therapy were administered (n = 20 chemotherapy, 10 biologicals, and 19 topical therapies). Thirteen patients died from MF-related causes, and 8 died from other causes. Acute and chronic toxicities were consistent with those previously reported. CONCLUSION: ECP given concurrently with, or immediately after, TSEB (32-40 Gy) significantly improves both PFS and CSS for patients with erythrodermic MF compared with TSEB without the addition of ECP. PMID- 10863225 TI - Topical therapy for psoriasis with the use of augmented betamethasone and calcipotriene on alternate weeks. AB - BACKGROUND: Topical corticosteroids, commonly used for psoriasis, show diminished response on continuous use. OBJECTIVE: We tested efficacy of topical corticosteroid and calcipotriene used on alternate weeks versus daily corticosteroid in patients with psoriasis. METHODS: In a randomized, observer blind design, the experimental group of 25 patients with stable plaque psoriasis received augmented betamethasone dipropionate 0.05% cream once daily in the first and third weeks and calcipotriene 0.005% ointment twice daily in the second and fourth weeks. The control group of 27 patients received augmented betamethasone once daily for 4 weeks. RESULTS: The experimental regimen was more effective than the control regimen as evidenced by (1) more patients with at least a 90% reduction in Psoriasis Area and Severity Index (PASI) score (difference 49.5%, 95% confidence interval [CI], 26.1%-72.9%, P <. 001), (2) lower PASI after 2 weeks (P < or =.04), and (3) greater percentage reduction in PASI after 2 and 4 weeks (difference 23.1% [CI, 11.1%-35.1%] and 46.4% [28.9%-63.8%], respectively; P <.001). The study had power of 93.7%. No patient had skin irritation. CONCLUSION: Use of augmented betamethasone and calcipotriene on alternate weeks is more effective than daily corticosteroid and represents a novel strategy for treating psoriasis. PMID- 10863226 TI - Current concepts: the use of immunoperoxidase techniques in mohs micrographic surgery. AB - Mohs micrographic surgery is used for the removal of certain malignant tumors, both ensuring complete excision by histologic examination of margins as well as minimizing normal tissue loss. Recently, several investigators have incorporated the use of immunoperoxidase techniques to aid in the removal of selected high risk carcinomas, sarcomas, and melanomas. We describe the basic principles of immunoperoxidase and review recent articles in which immunoperoxidase was used as an adjunct to routine hematoxylin-eosin staining in Mohs micrographic surgery. Additionally, we show examples of selected tumors comparing routine hematoxylin eosin stains and immunoperoxidase. We believe the use of immunoperoxidase can be of significant value in the removal of certain high-risk tumors. In particular, this technique is useful in "unmasking" malignant cells in areas of dense inflammation, identification of some cases of perineural invasion, identification of pagetoid spread in carcinomas and melanomas, and finally in helping to identify subtle margins in dermatofibrosarcoma protuberans. PMID- 10863227 TI - Short-pulse carbon dioxide laser resurfacing of the neck. AB - BACKGROUND: Carbon dioxide (CO(2)) laser resurfacing of the face has become an increasingly popular procedure. However, laser resurfacing of the neck has been largely avoided because of fears of scarring or pigmentation changes. OBJECTIVE: Our purpose was to evaluate the efficacy of treatment and incidence of complications after short-pulse CO(2) laser resurfacing of the neck. METHODS: A total of 308 patients received concomitant face and neck CO(2) laser resurfacing. A 90 -micros pulse duration CO(2) laser without a scanner was used in all cases for 2 passes on the neck (10.6-microm wavelength, 500-mJ pulse energy, 90-micros duration, 3-mm spot size) and a continuous CO(2) laser with a computer-generated scanner (396-micros dwell time, 18 W) was used for 3 passes over the face except for the perioral area, which received 4 passes. The incidence of scarring or permanent pigmentation changes was determined. Forty patients who had been treated at least 6 months but no longer than 18 months earlier were randomly surveyed by phone to assess the degree of improvement. RESULTS: Of the 308 patients treated, there were no cases of scarring or permanent pigmentation changes. Surveyed patients reported a 39% improvement in rhytides and tightening on the neck. CONCLUSION: Resurfacing of the neck can be performed safely in conjunction with resurfacing of the face. Patients may be offered improvement in the neck with little chance of scarring or permanent pigmentary changes when resurfacing on the neck is performed by means of a short-pulse duration laser for a limited number of passes instead of the more aggressive laser parameters previously used such as continuous long-pulse duration treatments. PMID- 10863228 TI - Laser treatment of imipramine-induced hyperpigmentation. AB - BACKGROUND: Blue or slate-gray hyperpigmentation is seen with a variety of medications, including imipramine. We describe a patient with significant imipramine-induced pigmentation. OBJECTIVE: The purpose of this study was to describe an effective laser treatment resulting in improvement of imipramine induced hyperpigmentation, without discontinuing the medication. METHODS: The patient underwent treatment with carbon dioxide, erbium, alexandrite, and ruby lasers to hyperpigmented areas. Tissue biopsy specimens taken before treatment, immediately after treatment with the alexandrite laser, and at clearing were analyzed by light microscopy. RESULTS: The Q-switched alexandrite and ruby lasers resulted in clinical improvement in the patient's hyperpigmentation and a decrease in pigment granules on light microscopy. CONCLUSION: Both the Q-switched alexandrite and ruby lasers are effective treatments for imipramine-induced hyperpigmentation. The improvement is progressive with successive sessions. PMID- 10863229 TI - Mercury exposure and cutaneous disease. AB - Human contact with mercury has been ongoing for centuries and was previously considered a legitimate means of treating different cutaneous and systemic conditions. Toxicity from this heavy metal may occur from exposure to elemental, inorganic, and organic forms of mercury. This article outlines the signs and symptoms of mercury poisoning and the different clinical conditions with assorted cutaneous findings. PMID- 10863230 TI - Leg ulcers [bibliography]. AB - This report reflects the best data available at the time the report was prepared, but caution should be exercised in interpreting the data; the results of future studies may require alteration of the conclusions or recommendations set forth in this report. PMID- 10863231 TI - Foundation for International Dermatologic Education: the first 25 years. AB - The Foundation for International Dermatologic Education was organized to improve dermatological education and care in developing countries. The accomplishments of the foundation over the past 25 years are reviewed. PMID- 10863232 TI - Surgical pearl: repair of partially torn earlobes-punch technique versus conversion to complete tear. PMID- 10863233 TI - Cutaneous deciduosis. AB - Cutaneous deciduosis is an exceedingly rare manifestation of endometriosis potentially mistaken for malignancy and thus far documented solely within surgical scars. We describe two additional cases, one occurring in a pregnant 21 year-old woman as a solitary flat erythematous vulvar papule, an extraordinary location not previously recorded. Histologic examination in that case revealed a subepithelial nodular aggregate of atypical large dyscohesive cells with accompanying edema and inflammation. An immunohistochemical panel showed positivity of the cells for vimentin and Ki-1 (CD30). Intracellular sulfated mucin and glycogen were also demonstrated. In a second case, a 27-year-old woman had a nodule at the umbilicus, removed incidentally during the course of cesarean section. Microscopically there were several circumscribed, multilobulated, intradermal nodules with variably sized lumens formed by crowded large epithelioid cells. The disparate histologic appearance of these examples highlights an essential challenge in their diagnosis. Clinical recognition is difficult unless suggested by more characteristic history or location. PMID- 10863234 TI - Cavitating pulmonary infiltrate in an adolescent with pyoderma gangrenosum: a rarely recognized extracutaneous manifestation of a neutrophilic dermatosis. AB - Neutrophilic dermatoses such as pyoderma gangrenosum are characterized by sterile, neutrophilic cutaneous infiltrates. Extracutaneous neutrophilic infiltrates can occur, primarily in the joints, lungs, heart, central nervous system, gastrointestinal tract, and eyes. Pulmonary disease is the most frequently reported extracutaneous manifestation of pyoderma gangrenosum and is characterized by patchy infiltrates or interstitial pneumonitis. We describe an adolescent with typical pyoderma gangrenosum who presented with cavitary pneumonia and responded completely to oral corticosteroids. In patients with inflammatory ulcers, extracutaneous neutrophilic disease should be considered, once an infectious process has been excluded. PMID- 10863235 TI - Reticulate acropigmentation of Dohi: a case report of autosomal recessive inheritance. AB - Reticulate acropigmentation of Dohi is a rare dyschromic disorder that generally has an autosomal dominant pattern of inheritance. Most of the cases have primarily been described from Japan. Only a few similar cases have been described elsewhere. We describe 3 black siblings, one boy and two girls, who had progressive reticulate hyperpigmented and hypopigmented macules over the dorsa of hands and feet, which began in early childhood. There were no palmar pits or breaks of the epidermal rete ridge pattern nor was there a family history of any pigmentary skin diseases. Three skin biopsies were performed on one patient; a biopsy specimen from a hyperpigmented macule showed increased melanin in all epidermal levels tapering towards the surface, a second biopsy specimen from a hypopigmented macule showed much less melanin, but it had a similar distribution. A third specimen from a hyperpigmented macule for electron microscopy showed a moderate number of stage III and IV melanosomes in the cytoplasm of the melanocytes. To our knowledge, these patients are the first cases reported from the Middle East with an autosomal recessive pattern of inheritance, confirming previous reports. PMID- 10863236 TI - Pilomatrix dysplasia in an immunosuppressed patient. AB - Immunosuppressive drugs have been used for many years in the prevention of graft failure in transplant recipients. Although they improve morbidity and mortality after transplantation, these medications carry a significant risk of adverse mucocutaneous and systemic effects. We describe a patient receiving 4 immunosuppressive drugs who experienced persistent facial dysmorphism along with striking follicular disturbances. On histopathologic examination, the follicular structures were dilated and hyperplastic with a peculiar dysplasia of the pilar matrix. Based on a review of the clinical, microscopic, and investigational findings of the skin previously reported in association with her immunosuppressive drugs, we conclude that cyclosporine was the most likely causative agent. Moreover, hypertrichosis, dysmorphic facies, and tissue hyperplasia have all been observed in patients during cyclosporine administration. PMID- 10863237 TI - Lymphangioma-like Kaposi's sarcoma: etiology and literature review. AB - Much confusion surrounds a rare and occasionally described variant of Kaposi's sarcoma (KS) known as lymphangioma-like or bullous KS. We describe the typical clinical and histologic features and elucidate the etiologic cell in lymphangioma like KS. A computer-based review of the English-language literature was performed. Routine histologic and immunoperoxidase techniques were performed on formalin-fixed tissue. Immunoperoxidase staining for anti-CD31, anti-CD34, and anti-factor VIII-related antigen suggests that the etiologic cell of origin is the vascular endothelial cell. Lymphangioma-like KS has been described for more than a century and given various names. Dilated vascular spaces correlate with the clinically bullous lesions, which are veritable KS and not a secondary reaction to it. PMID- 10863238 TI - Unilateral multiple facial angiofibromas: a mosaic form of tuberous sclerosis. AB - Tuberous sclerosis has many forms of clinical presentation. Rarely, multiple facial angiofibromas of unilateral distribution have been reported. We describe 2 patients with such a presentation and hypothesize that this is a mosaic form of tuberous sclerosis. PMID- 10863239 TI - Topical tretinoin and 5-fluorouracil in the treatment of linear verrucous epidermal nevus. AB - Treatment of a linear verrucous epidermal nevus using topical 0.1% tretinoin cream and 5% 5-fluorouracil in a young patient is described. In 1994, successful topical therapy using this combination was described in the management of an inflammatory linear verrucous epidermal nevus. We report another case in which treatment of a noninflamed epidermal verrucous nevus with 0.1% tretinoin and 5% 5 fluorouracil resulted in significant improvement. An updated summary of the literature discussing management of epidermal nevi is presented. PMID- 10863240 TI - Pruritic folliculitis of pregnancy. AB - Pruritic folliculitis of pregnancy is a rare pregnancy dermatosis that clears spontaneously in the postpartum period. We describe a patient with characteristic clinical and histopathologic features of this dermatosis. PMID- 10863241 TI - Inherited epidermolysis bullosa comes into the new millenium: a revised classification system based on current knowledge of pathogenetic mechanisms and the clinical, laboratory, and epidemiologic findings of large, well-defined patient cohorts. PMID- 10863242 TI - The nature of solar keratosis: a critical review in historical perspective. PMID- 10863243 TI - Updated sunscreen advice: SPF30. PMID- 10863244 TI - Pyogenic granuloma-like lesion associated with topical tazarotene therapy. PMID- 10863245 TI - Standards of care for patients with malignant melanoma. PMID- 10863249 TI - Chiropractic management of mechanical neck and low-back pain: a retrospective, outcome-based analysis. AB - BACKGROUND: Evidence suggests that spinal manipulation is an effective treatment for mechanical neck and low-back pain (LBP). Treatment efficacy is important to establish for these symptoms because combined they account for a considerable amount of disability and substantial associated direct and indirect costs to society. OBJECTIVE: The purpose of this study was to examine the outcome of patients undergoing chiropractic treatment for mechanical neck or LBP. DESIGN AND SETTING: A retrospective, outcome-based analysis was done for patients seeking care at a private chiropractic practice over a 1-year period. A total of 512 files were reviewed, with 119 patients selected for inclusion. Patients were included if their chief symptom was uncomplicated mechanical neck or LBP. Diagnoses included cervical, lumbar, or sacroiliac joint sprain/strain (International Code of Diagnostics version 9 [ICD-9] code: 847.1, 847.3, 846.1, respectively), discogenic LBP (ICD-9: 722.1), and headaches (ICD-9: 784.0) because many patients with neck pain presented with concomitant headaches. Disability and pain were measured with the modified Oswestry scale (for the patients with LBP), Neck Disability Index, and an 11-box visual analogue pain scale before and after treatment. Treatment consisted of spinal manipulation, various soft-tissue techniques, home-care instructions, and ergonomic and return to-activity advice, including rehabilitative exercises. Patients received an average of 12 treatments over a 4-week period. Statistical analysis was performed on pretreatment and posttreatment values for both disability and pain. Stratification was based on duration (acute/subacute, chronic, acute exacerbation of a chronic condition) and severity (mild, moderate, or severe) of symptoms. RESULTS: Statistically significant reductions in disability and pain scores were achieved in all groups. An average 52.5% and 52.9% reduction in pain and disability, respectively, was achieved in the low-back group. The chronic LBP group realized a less statistically significant reduction of pain and disability (19.7% and 19.8%, respectively) than the acute/subacute (66.8% and 62.5%) or the chronic/recurrent group (56. 5% and 63.4%). The differences were statistically significant. Patients with neck pain had an average 53.8% and 48.4% reduction in their pain and disability, respectively. Patients with concomitant neck pain and headaches had statistically significant higher pretreatment and posttreatment disability and pain scores than those with only neck pain. There was no statistically significant difference in outcomes between groups stratified according to pain intensity. CONCLUSIONS: Patients attending a private chiropractic clinic for treatment of mechanical neck pain or LBP had statistically significant reductions in their pain-related disability after treatment. These results indicate that chiropractic manipulation is beneficial for the treatment of mechanical neck pain and LBP. However, care must be taken when drawing conclusions from these outcomes. The study design does not account for the natural history of low back- or neck pain-related disability and therefore does not allow for claims of treatment efficacy. In addition, it has been suggested that patients presenting to medical doctors with these symptoms have significant overlying comorbidity when compared with patients presenting to a chiropractor. PMID- 10863250 TI - Behavioral-graded activity compared with usual care after first-time disk surgery: considerations of the design of a randomized clinical trial. AB - OBJECTIVE: To present the design of a trial on the effectiveness of a behavioral graded activity model. DESIGN: Randomized clinical trial. PATIENTS: Patients undergoing first-time lumbar disk surgery who still have low-back pain at the 6 week neurosurgical consultation. INTERVENTIONS: A patient-tailored behavioral graded activity program that is based on operant therapy. The key elements of this program are baseline measurements, goal-setting, and time-contingency. This program is compared with usual care in physiotherapy, which is pain-contingent. OUTCOME MEASURES: Primary measures are the patient's global impression of the effect and their functional status. Secondary measures are kinesiophobia, catastrophizing, pain, main complaint, range of motion, and relapses. The direct and indirect costs will also be assessed. The effect measures are rated before randomization and 3, 6, and 12 months later. DISCUSSION: Several trials have been conducted on the effectiveness of behavioral treatments. Subjects were always patients with chronic low-back pain. In this trial, we apply such a treatment in patients after first-time disk surgery in a primary care setting. PMID- 10863251 TI - Silver needle therapy for intractable low-back pain at tender point after removal of nucleus pulposus. AB - OBJECTIVE: To examine the use of a new silver needle therapy for treating tender points involved in intractable low-back pain after removal of nucleus pulposus. SUBJECTS: The study involved 24 patients (17 men and 7 women) aged 26 to 67 years with a mean age of 54.5 +/- 5 years. SETTINGS: The Department of Orthopedics at the First Military Medical University, the Department of Rehabilitation Medicine at the General Hospital of PLA in Beijing, and the Department of Acupuncture, the First People's Hospital in Kunming City, People's Republic of China, were the settings for this study. METHODS: The patients were treated with traditional silver needle therapy at tender points in the low back and buttocks. Pain at each of the tender points was measured before and after treatment; the scores were compared with a Student t test. RESULTS: The therapeutic results suggest that the total scores for each tender point after treatment were significantly lower than those taken before treatment (P <.001). CONCLUSION: Silver needle therapy shows promise for treating low-back pain after surgery for disc herniation. Further clinical trials are needed to confirm the effectiveness of this treatment. PMID- 10863252 TI - A clinical trial investigating the possible effect of the supine cervical rotatory manipulation and the supine lateral break manipulation in the treatment of mechanical neck pain: a pilot study. AB - OBJECTIVE: To evaluate the possible effect of the supine cervical rotary manipulation and the supine lateral break manipulation in the treatment of mechanical neck pain, according to subjective and objective clinical findings. BACKGROUND: Delivering a supine lateral break manipulation to the ipsilateral side of an inflamed facet joint(s) that exhibits a lateral flexion fixation may result in pain and/or discomfort to the patient. Thus the proposed alternative is a supine cervical rotary manipulation delivered on the ipsilateral side or a supine lateral break manipulation delivered on the contralateral side of the relevant joint(s). DESIGN: Randomized, comparative clinical trial. SUBJECTS: Two groups of 15 subjects diagnosed with mechanical neck pain. INTERVENTION: The diagnosis of mechanical neck pain and the identification of lateral flexion fixations in the cervical spine were made with conventional clinical evaluation, including motion palpation. Group A received a cervical rotary manipulation(s) on the ipsilateral side of the lateral flexion fixation(s), while group B received a supine lateral break manipulation(s) on the contralateral side of the lateral flexion fixation(s). Subjects received a maximum of 10 treatments over a 4-week treatment period. OUTCOME MEASURES: Both treatment groups were assessed with subjective (Numerical Pain Rating Scale 101, McGill Short-Form Pain Questionnaire and the Canadian Memorial Chiropractic College Neck Disability Index) and objective (cervical range of motion goniometer and algometer) measurement parameters at the initial consultation (before any treatment), the final consultation, and at a 1-month follow-up consultation. Statistical analysis was conducted at a 95% confidence level (alpha =.05) with the non-parametric 2-tailed Wilcoxon signed ranks test, the Mann-Whitney U test, and descriptive statistics. Two-tailed power analysis was conducted after the fact, where a confidence level of 80% (beta =.20) was considered satisfactory. RESULTS: Intragroup analysis indicated a significant difference between the initial consultation data and the final consultation data for the subjective data, indicating an effect. Analysis of the objective data did not reveal any significant difference. Intergroup analysis did not reveal any significant difference between the 2 groups when comparing the data of the initial consultation and the final consultation, indicating that both treatments had a similar or equal effect. Power analysis was not satisfactory for most data, indicating the possibility of many Type II errors. CONCLUSION AND RECOMMENDATIONS: Statistically, the results suggested that both treatments had an effect but that neither group showed a benefit over the other. However, because of the unsatisfactory power of the study, conclusions are to be drawn with caution. Clinical significance supported the statistical outcomes where it was suggested that both treatments had an effect and that neither treatment had a greater effect. A larger sample size and the inclusion of a placebo group is recommended to reveal true treatment outcomes and trends. PMID- 10863253 TI - Short-term effects of lumbar posteroanterior mobilization in individuals with low back pain. AB - OBJECTIVE: To establish the short-term effects of lumbar posteroanterior mobilization in patients with low-back pain, compared with a control intervention. DESIGN: Self-controlled cross-over design. MAIN OUTCOME MEASURES: The force-displacement characteristics of the spine in response to the application of a posteroanterior force, lumbar flexion, and extension range of movement; pain during flexion, extension, and on worst movement; pain on posteroanterior loading; and overall pain relief. PATIENTS: Twenty-six patients with nonspecific low-back pain who experienced pain on flexion or extension and whose pain settled quickly after provocation, from a physiotherapy clinic and university campus. METHODS: Patients received posteroanterior mobilization and a control intervention in an order that was randomly allocated. The magnitude of force in treatment dose was selected by the treating physiotherapist. An observer who was blinded to the order of interventions performed all measurements. Outcome measures were recorded before and after each intervention, and change scores were calculated to quantify the effect of the intervention. RESULTS: No significant differences were found between the mobilization and control interventions in relation to posteroanterior response or range of movement. The score for pain on worst movement showed significantly greater improvement for the mobilization than for the control procedure. CONCLUSIONS: Lumbar posteroanterior mobilization was not observed to produce any objectively measurable change in the mechanical behavior of the lumbar spine of patients with low-back pain. Improvement in some pain variables was observed in comparison with a control procedure, but this may be due to a placebo effect. PMID- 10863254 TI - Alcohol and low-back pain: a systematic literature review. AB - BACKGROUND: The role of lifestyle factors is an important issue in the prevention and treatment of disease. Although certain lifestyle factors in relation to low back pain have evoked much interest, interest has not focused on alcohol consumption. An appraisal of the epidemiologic literature seems warranted. OBJECTIVES: To establish if there is evidence in the literature for a causal link between alcohol consumption and low-back pain. DATA SOURCES: Nine original research reports published between 1987 and 1995 were obtained through a MEDLINE search for the years 1992 to 1998, with various combinations of the terms "alcohol," "substance abuse," "life-style, " "risk factor," "epidemiology" and "low back pain." An additional manual search was made of relevant bibliographies without limitation for year of publication. DATA SYNTHESIS: A systematic review was made of the epidemiologic literature to uncover any evidence for a causal relation between alcohol consumption and low-back pain. RESULTS: None of the studies reported a positive link between alcohol consumption and low-back pain, and no positive gradient was found in studies that included an analysis of the dose-response. None of the studies was prospective in design. CONCLUSIONS: Alcohol consumption does not seem to be associated with low-back pain, but well designed specific alcohol/low-back pain-centered studies are lacking. PMID- 10863255 TI - Osteochondritis dissecans of the knee: a radiology case report. AB - OBJECTIVE: To discuss the case of a patient with osteochondritis dissecans, a common disorder primarily affecting children and adolescent patients that involves the medial femoral condyle, and to examine its radiologic appearance. CLINICAL FEATURES: A 25-year-old man with knee pain sought treatment at a chiropractic clinic. The patient had a 2-year history of left knee pain that was exacerbated by activity and relieved with rest. Plain film radiography of the knee revealed findings consistent with osteochondritis dessicans. INTERVENTION AND OUTCOME: The patient was promptly referred for a concurrent orthopedic consultation with subsequent management. A trial basis of conservative care was initiated with the possibility of future surgical intervention if necessary. CONCLUSIONS: Osteochondritis dissecans is a common disorder of unknown and perhaps controversial cause. The disease is characterized by a fragment of articular cartilage and subchondral bone (osteochondral fragment) that becomes separated from the underlying bone. Treatment is aimed primarily at the preservation of articular surface congruity. PMID- 10863256 TI - Vertigo, tinnitus, and hearing loss in the geriatric patient. AB - OBJECTIVE: To document clinical changes after a course of chiropractic care in a geriatric patient with vertigo, tinnitus, and hearing loss. CLINICAL FEATURES: A 75-year-old woman with a longstanding history of vertigo, tinnitus, and hearing loss experienced an intensified progression of these symptoms 5 weeks before seeking chiropractic care. Radiographs revealed a C3 retrolisthesis with moderate degenerative changes C4-C7. Significant decreases in audiologic function were evident, and the RAND 36 Health Survey revealed subjective distress. INTERVENTION AND OUTCOME: The patient received upper cervical-specific chiropractic care. Paraspinal bilateral skin temperature differential analysis was used to determine when an upper cervical adjustment was to be administered. Radiographic analysis was used to determine the specific characteristics of the misalignment in the upper cervical spine. Through the course of care, the patient's symptoms were alleviated, structural and functional improvements were evident through radiographic examination, and audiologic function improved. CONCLUSION: The clinical progress documented in this report suggests that upper cervical manipulation may benefit patients who have tinnitus and hearing loss. PMID- 10863257 TI - A critical evaluation of the methodology of a low-back pain clinical trial. PMID- 10863259 TI - In response PMID- 10863258 TI - The short-term effect of spinal manipulation in the treatment of infantile colic: a randomized controlled clinical trial with a blinded observer. PMID- 10863261 TI - In response PMID- 10863260 TI - The frequency of positive common spinal clinical examination findings in a sample of premenstrual syndrome sufferers. PMID- 10863262 TI - Prevalence of hyperplastic articular pillars in the cervical spine and relationship with cervical lordosis. PMID- 10863263 TI - In response PMID- 10863264 TI - The importance of normalization in the interpretation of surface electromyography: a proof of principle. PMID- 10863303 TI - [Antiretroviral therapy. Update at the beginning of a new century]. AB - For antiretroviral therapy (ART) nucleoside reverse transcriptase inhibitors (NRTI), protease inhibitors (PI) and non-nucleoside reverse transcriptase inhibitors (NNRTI) are available, and are used in combination treatments. The use of PI is associated with new side effects, in particular affecting lipid metabolism. The objective of treatment continues to be a maximum reduction of the viral load. The question whether to treat the HIV early on or later is under controversial discussion. Simplified therapeutic regimens making possible single or twice-daily doses and the use of small numbers of tablets represent a major advance in ART. PMID- 10863304 TI - [Antiretroviral therapy. NNRTI (non-nucleoside reverse transcriptase inhibitor)]. PMID- 10863305 TI - [Protease inhibitor saving alternatives]. PMID- 10863306 TI - [Antiretroviral therapy. A brief profile of all available drugs]. PMID- 10863307 TI - [Non-nucleoside reverse transcriptase inhibitor as a fully viable alternative. Strong therapy start without protease inhibitor]. PMID- 10863308 TI - [Pathogenesis of HIV infection as the key to eradication? New therapeutic approaches]. AB - While the effectiveness of conventional antiviral treatment is well proven, its limitations are becoming more and more evident. New approaches are resulting from the "decoding" of the pathogenesis of HIV infection. Thus, for example, experimental studies are looking at drug-induced blockade of the co-receptors necessary for the infection of the target cells. Treatment can be applied at all stages of viral replication. So far, reverse transcriptase and protease inhibitors have been employed. Other strategies are aimed at immunostimulation (e.g. with interleukin 2) and immunosuppression (cortisone, hydroxyurea). The development of therapeutic vaccines is difficult, but prophylactic vaccines hold out greater promise; field trials are underway. PMID- 10863309 TI - [Various triple therapies in direct comparison. New decision aids for initial therapy]. PMID- 10863310 TI - [Nucleoside reverse transcriptase inhibitors. Sex-specific effectiveness and tolerance]. PMID- 10863311 TI - [Public health economic aspects of antiretroviral therapy. Can we afford to do less?]. AB - In Germany, politically motivated economic considerations are becoming even more important. In this connection, scientific research into the health system is not limited merely to a consideration of costs, but utilizes the same methods and conceptual models as economic research in general to investigate defined problems. The reduction of illness-related loss of production and quality of life to monetary units in the models used, often stimulates critical discussions of the ethical permissibility of such an approach. Public discussion of such models then makes it possible to justify rationally founded allocation decisions and to expose and thus prevent hidden forms of rationing. The medically successful concept of long-term administration of highly active antiretroviral combination treatment (HAART) was first considered by numerous studies to be cost-effective solely on the basis of the saving of inpatient treatment costs--a stance that in the light of the life-shortening HIV infection should not be thought to be only basis for the decision. Owing to a lack of experience with the long-term prognosis under HAART, it is difficult to assess indirect costs for HIV infection. An assessment of the quality of life in the symptom-free stages of HIV infection depends largely on coping strategies, so that the results of studies on quality of life measurement under antiretroviral treatment must not lightly be interpreted as a function of a given therapeutic strategy. Given the high costs of HIV infection to the economy, the marked reluctance to provide the necessary funding for research in the area of epidemiological monitoring, specific preventive measures and vaccination strategies is to be regretted. PMID- 10863312 TI - [New variations in highly active antiretroviral therapy. Simple combination as a promising option]. PMID- 10863313 TI - [Second chance therapy. New protease inhibitor for salvage therapy]. PMID- 10863314 TI - [HIV infection in childhood]. AB - As a result of preventive measures during pregnancy and improved antiretroviral combination treatment, the problem area of HIV infection in childhood has undergone a change in recent years. Thanks to intensive cooperation between obstetricians and pediatricians, mother-to-child transmission of HIV in Germany has been decreased to less than 2%. Improvements in the diagnosis of HIV infection in childhood, infected children have been identified very early, and started on antiretroviral treatment, and prophylactic measures initiated. In the case of life-long anti-HIV treatment beginning in childhood, the same problems as those seen in adults must be expected (development of resistance, adherence, adverse effects). PMID- 10863315 TI - [HIV plus hepatitis C or Kaposi sarcoma. Treatment options in coinfection]. PMID- 10863316 TI - [Chronic hepatitis C in HIV-infected patients. Unfavorable interaction of HIV and HCV infection]. AB - HIV coinfection promotes the progression of chronic hepatitis C. The increased hepatic mortality and improved life expectancy of HIV-positive patients warrants treatment of chronic hepatitis C in those infected with HIV. The treatment of hepatitis C considered standard for HIV-negative and, in principle, also for HIV positive patients is the combination of interferon alpha with ribavirin. Possible interaction with the antiretroviral treatment by ribavirin must, however, taken into account. Although new therapeutic approaches are available, they have not yet been adequately validated terms of their efficacy and safety. PMID- 10863317 TI - [Osteonecrosis: a rare complication of HIV infection. Association with certain risk factors]. AB - Osteonecrosis is a rare complication of HIV infection. The presumptive cause of the aseptic osteonecrosis is a disturbed blood supply to the bone. Most cases of osteonecrosis are associated with numerous risk factors, such as use of steroids, alcohol abuse, coagulopathies or metabolic derangements. Since conventional X rays appear unremarkable, early forms often go unrecognized or are diagnosed late. Methods of establishing the diagnosis are NMR and three-phase skeletal scintigraphy. The pathogenesis of osteonecrosis in HIV infection is unclear. So far, about 30 cases have been reported in the literature. Since a number of these cases had no classical risk factors, it is assumed that the HIV infection itself is the causative agent. In other patients anticardiolipin antibodies, which are considered to be a risk factor, are found. Other HIV patients with aseptic osteonecrosis have elevated blood lipids; changes in blood fats have long been established as a risk factor in osteonecrosis. Furthermore, an association of osteonecrosis with proteinase inhibitor-induced metabolic lipid disorders was reported. Whether the risk for osteonecrosis in treatment with proteinase inhibitors actually is raised, or whether the association is a coincidence needs further investigation. We would recommend that in HIV patients with typical symptoms--in particular when classical risk factors are present--osteonecrosis be included in the differential diagnostic considerations. PMID- 10863318 TI - [Lipodystrophy and metabolic disorders in anti-HIV therapy]. AB - For the effective treatment of HIV infection, combinations of antiretroviral drugs that, at various points of attack, prevent viral replication are used. After the introduction of HIV-1 specific protease inhibitors, various metabolic complications and changes in body habitus manifesting as abnormal fat distribution were observed in HIV patients. These side effects, initially assigned class-specifically to the protease inhibitors, have, however, also been frequently seen under other drug combinations. Changes to body habitus as a result of fat redistribution have mostly been designated as lipodystrophy syndrome, but are now increasingly being recognized as highly heterogeneous abnormalities. Furthermore, the disturbances of glucose and fat metabolism may occur completely independently of phenotypic habitus changes. The result is that both the suspected pathophysiological relationships and the therapeutic consequences are becoming even more complex. This paper summarizes the current state of investigations into the side effects of antiretroviral therapy and their clinical consequences. PMID- 10863319 TI - [The future of HIV therapy. Research pipelines are filled with options]. AB - Following the first international conference in 1999, twice as many new antiretroviral substances are in the early clinical or preclinical phase of investigation as are currently approved for ART in Germany. These data are almost certainly not complete, and the number of "candidates" will be larger rather than smaller. Nevertheless they send a clear message. Antiretroviral treatments guided by therapeutic pessimism are anything but up to date. This applies to inadequate dual treatments or the delayed use of ART. The plethora of new medications with no, or virtually no, cross-resistance vis-a-vis previously approved substances renders in admissible any reasons advanced for delayed or reserved treatment. The present article has been written also for those therapists who, for the reasons set out above, have so far greatly delayed, or have provided only "half-hearted", treatment. In the near future, ART will comprise not only the inhibition of HIV replication, but will also deal with immune reconstruction. As in the last 15 years, antiretroviral therapeutic strategies will be the signposts and pacemakers for the treatment of other infectious diseases, too. Not least of the conditions included are such important chronic viral ailments as hepatitis B or C, which have just successfully crossed the combination therapy threshold. It is to be hoped that a maximum number of the substances presented here will pass the proving period leading a full-blown antiretroviral medication. PMID- 10863320 TI - [Campaign against a chameleon. Therapeutic strategies to control a survival artist]. PMID- 10863321 TI - [Observing CD4 cell count and viral load alone is not enough. Value of protease inhibitors]. PMID- 10863322 TI - [Therapy planning. It depends on the proper administration series]. PMID- 10863323 TI - [Therapy of advanced stages. "Mega-HAART" or "drug holiday"?]. PMID- 10863324 TI - [Adhesional chemokine, fractalkine]. PMID- 10863325 TI - [Synovial fluids from patients with rheumatoid arthritis induce the differentiation of human promyelocytic leukemia cell line HL 60]. AB - Bone marrow abnormalities have been found to play a role in the pathogenesis of rheumatoid arthritis (RA). Recent studies have also confirmed the presence of undifferentiated hematopoietic progenitor cells as well as the expression of stem cell factor in the synovial membranes in RA. The present study investigates whether RA synovial fluids contain factors that can induce differentiation of CD 14 positive/HLA-DR positive cells from undifferentiated hematopoietic cells. Synovial fluid specimens from 18 patients with RA and from 10 control patients, including patients with osteoarthritis and Behcet's disease, were studied. Human promyelocytic leukemia cell line HL 60 (5 x 10(4)/well) were cultured in the presence or absence of the synovial fluids for 5 days, after which the expression of CD 14 and HLA-DR was examined by flow cytometry. The induction of differentiation of CD 14 positive/HLA-DR positive cells or HLA-DR positive cells from HL 60 cells was significantly enhanced more in the presence of synovial fluids from RA patients than in the presence of those of control patients. However, the sera from the RA patients could not induce the differentiation of CD 14 positive/HLA-DR positive cells or HLA-DR positive cells from HL 60 cells. Most cytokines found in RA synovial fluid could not induce the differentiation of HL 60 cells. Of note, treatment of synovial fluids with hyaluronidase significantly decreased or abrogated their capacity to induce the differentiation of HLA-DR positive cells from HL 60. There was no significant difference in the concentration of hyaluronic acid in the synovial fluid between the RA patients and the control patients. These results indicate that there are factors that can induce differentiation of HLA-DR positive cells from undifferentiated hematopoietic cells in the synovial fluid of RA. The data also suggest that such differentiation factors might be related with qualitative abnormality of hyaluronic acid. PMID- 10863326 TI - [The significance of cancer screening by immunological parameters]. AB - Provided by the evidences that, in cancer patients, the production of Th 1 cytokines including IL-12, IFN-gamma, and TNF-alpha, is impaired, we asked whether these cytokines can be useful parameters for cancer detection. To the aim 174 patients diagnosed cancer of various organs, and 100 control individuals without cancer were enrolled to the study. We evaluated mitogen-stimulated production of cytokines and induction of Th 1 subset using peripheral blood mononuclear cells in vitro. Th 2 population was measured as the counterpart of Th 1 subset. NK cell activity was also measured. As acquired values did not show the normal distribution we employed a non-parametric test to compare the values between cancer and control. PHA-induced production of IL-12, IFN-gamma and TNF alpha, in cancer was lower than that in control. Th 1 subset induced in cancer was lower than that in control. We found no difference in Th 2 subset induction between cancer and control. On the other hand, NK cell activity was augmented in cancer patients. When patients were grouped to early stage and advanced stage, both groups showed suppressed production of all three cytokines and suppressed induction of Th 1 cells. Interestingly, none of cytokines nor Th 1 subset differed between the two stages, suggesting that the impaired cytokine production may be a common feature of cancer condition and participate in the etiology of cancer. In contrast, enhanced induction of Th 2 subset was seen in advanced stage compared to early stage, indicating that Th cells might be biased to differentiate to Th 2 cells in advanced stage. From the results IL-12, IFN-gamma, TNF-alpha and Th 1 subset as well as NK activity appear to be promising parameters for cancer detection. Above all, IL-12 and IFN-gamma seem to be the best parameters due to high sensitivity and specificity, and independence from the stage of cancer. PMID- 10863327 TI - [Hematological abnormalities of primary Sjogren's syndrome]. AB - Sjogren's syndrome (SS) is an autoimmune disease characterized by a chronic inflammatory response mainly localized to the lacrimal and salivary glands. However, it sometimes involves extraglandular organs culminating in systemic disorders. Hematological abnormalities are not uncommon, although they rarely have clinical significance. In this study we examined 99 patients with primary SS who visited our hospital during 1989 to 1999. Patient's mean age was 54.1 years and 95 out of 99 were female. Lymphopenia and leukopenia was noted in 35 patients (35.3%) and 26 patients (26.2%) respectively, and 7 patients (7.1%) had thrombocytopenia. 43 patients (43.4%) had either of these hematological abnormalities. Patients with lymphopenia showed significantly low frequency of arthralgia and anti-SS-A/B antibody was more common in this group. Only one patient in this group required prednisolone therapy because of polyarthritis and general fatigue while others needed no specific therapy. Patients with thrombocytopenia were significantly younger and a male/female ratio was higher than those without this abnormality. They had higher tendency to accompany with skin eruption, positive anti-SS-B antibody, anti-nuclear antibody and rheumatoid factor. Three out of 8 patients with thrombocytopenia were treated with prednisolone according to the protocol for idiopathic thrombocytopenic purpura. All of 3 patients had positive PA-IgG and normocellular bone marrow. Autoimmune mechanism such as polyclonal B cell activation may play a role in the pathogenesis of thrombocytopenia. PMID- 10863328 TI - [Autoimmune hepatitis complicated by intolerable pain of lower extremities and shock due to azathioprine]. AB - A 24-year-old woman was followed for about ten months with oral administration of prednisolone (22.5-35 mg/d) for autoimmune hepatitis. In June 1995, she noticed fatigue and appetite loss and blood chemistry revealed markedly deteriorated liver function. She was admitted to our hospital. The daily dose of prednisolone was increased to 60 mg. Her elevated levels of transaminases decreased gradually. Administration of azathioprine (100 mg/d) was started with tapering of prednisolone on August 18th. Ten days later, tender cervical lymphadenopathy and high fever occurred. Azathioprine administration was stopped immediately and intravenous antibiotics were given. On September 5th, 50 mg of azathioprine was administered again. Two hours later, the patient complained of intolerable pain from the lumbar region to the knee joints, which subsided following two injections of analgesics within a few hours. However, chills, high fever and hypotension (86/30 mmHg) subsequently developed. No bacterial growth was detected in blood culture. She was discharged on September 12th. On October 4th, she visited our out-patient clinic. The next day, she took one tablet (50 mg) of azathioprine at 10 o'clock. She noted intense pain from the thighs to the knees and calves around noon again. Her home doctor found that she exhibited shock (BP 67/?). She was immediately taken to our department. The same symptoms and signs as the above-mentioned occurred. Azathioprine was considered responsible for these two adverse reactions (shock) as an allergen. Later, systemic lupus eythematosus was diagnosed in 1996. And she died to pulmonary hypertension in May 1999. Physicians should be aware of the potential adverse effect of azathioprine administered in order to manage the patients with autoimmune disorders. PMID- 10863329 TI - [A case of subclinical Sjogren syndrome associated with high levels of serum IgM, thrombocytopenia and splenomegaly]. AB - A 13-years-old girl was admitted to our hospital with high levels of serum IgM, thrombocytopenia and splenomegaly. Not only IgG but also IgM were found on the surface of platelets by flow-cytometry. Direct Coombs' test was positive, and IgG was also found on the surface of red blood cells. After splenectomy, platelet count was increased and serum IgM was decreased. The biopsy of salivary glands showed infiltration of lymphocytes around the ducts, and Shirmer test revealed slightly decreased secretion of tears, suggesting subclinical Sjogren syndrome. PMID- 10863330 TI - [A case of Sjogren's syndrome complicated by polymyositis and sarcoidosis with HLA-B7 and DR 8: common causes of susceptibility for these diseases]. AB - We describe a case of a Japanese patient initially presenting with Sjogren's syndrome who later developed polymyositis and sarcoidosis. A 67-year-old woman with a 4 month history of myalgia was admitted in April 1998 for examination. The patient had a 10 year history of symptoms consistent with Sjogren's syndrome. A diagnosis of polymyositis was made based on a biopsy of the muscle and an electromyogram. Positive Shirmer and Rose Bengal tests and results of a minor salivary gland biopsy were all consistent with Sjogren's syndrome. Chest computed tomography detected a bilateral hilar lymphadenopathy. Microscopic examination of a mediastinal lymph node demonstrated multiple noncaseating granulomas with multiple epithelioid cells and Langhans-like giant cells. A diagnosis of sarcoidosis was made based on these findings. Hepatitis C infection was also detected by elevated antibody levels. The patient was given 40 mg/day of oral prednisolone and a remission of her myositis and lymphadenopathy was obtained. The patient exhibited HLA-B7 and HLA-DR 8. HLA-DR 8 is commonly associated with these three disorders, and HLA-B7 is also associated with overlap syndrome in Japanese patients. The present case suggested the possibility of a common etiological background for these three disorders. Furthermore, the importance of genetic background, including HLA phenotype, in determining susceptibility to these disorders was demonstrated. PMID- 10863331 TI - [Two children with suspected primary vasculitis of mesenteric vessels--a case report]. AB - We reported 2 children with suspected primary vasculitis of mesenteric vessels. Both children were admitted to our hospital with the complaints of abdominal pain, bloody stool or diarrhea. Laboratory examination simultaneously revealed leukocytosis with dominant neutrophils, positive CRP, and hypoalbuminemia. Although prothrombin time and activated partial thromboplastin time were within normal limits, the increased levels of FDP-E, D-dimer, and von Willebrand factor activity were observed, which suggested the endothelial cell activation and the coagulation/fibrinolysis system activation. Abdominal echography and CT scanning demonstrated the edematous thickening of intestinal or colon walls probably due to the vasculitic permeability changes of mesenteric artery. During the disease courses, skin rash, bleeding tendency, arthritis and proteinuria were not observed, and no autoantibodies including anti-nuclear antibody, anti-DNA antibody, and myeloperoxidase-antineutrophil cytoplasmic antibody, were detected. Taken together, we suspected these children as restricted vasculitis of mesenteric vessels. Intravenous prednisolone was administrated, and the clinical and laboratory abnormalities recovered completely within 2 weeks. Thus, we suggested that the leukocyte counts, CRP, and the determination of von Willebrand factor and coagulation/fibrinolysis study accompanied with X-ray, echography, and CT scanning will be useful for the early diagnosis of vasculitis before the pathologic and irreversible vascular damage are demonstrated. PMID- 10863332 TI - [Three patients with systemic lupus erythematosus complicated by cytomegalovirus colitis]. AB - We report three female patients with systemic lupus erythematosus complicated by massive intestinal hemorrhage during the recovery from lupus nephritis (case 1, 2) or central nervous system lupus (case 3) on high dose corticosteroid therapy. Large number of cytomegalovirus (CMV) antigen-positive leukocytes and cessation of bleeding with concurrent disappearance of the viral antigens after ganciclovir therapy indicated CMV colitis in all of the three patients. No recurrence of the symptom and a favorable response to ganciclovir without reduction in steroid regimen was common to these patients. PMID- 10863333 TI - [A case of common variable immunodeficiency with intractable diarrhea and the functional disorder of renal tubules]. AB - We report a case of a 31-year-old woman with common variable immunodeficiency (CVID) complicated with intractable diarrhea and the functional disorder of renal tubules. The patient became hypogammaglobulinemic after she suffered from measles at 6 years of age. She also suffered from lupus-like syndrome at 7 years of age. The complete remission was obtained by glucocorticosteroid treatment. An intravenous immunoglobulin replacement therapy was introduced at 11 years of age, since then her general condition was stable for more than 20 years. When she was 29 years old, she suffered from generalized malaise, anorexia with body weight loss, and numbness of face. The intractable diarrhea as protein loosing syndrome, and the severe abnormality of electrolyte balance with metabolic acidosis as the functional disorder of renal tubules were found. Her condition was not improved by the electrolytes or alkali replacement therapy. She was admitted for further evaluation and treatment. The intractable diarrhea and the functional disorder of renal tubules were dramatically improved after absolute restriction of food intake under hyperalimentation. When she began to take food, the symptom and sign became worse again. The interstitial nephritis and nonspecific inflammation of intestine were found by the tissue biopsy. The most characteristic finding was the infiltration of lymphocytes (predominantly CD 8 + T lymphocytes) in both intestinal mucosa and renal interstitium. The introduction of glucocorticosteroids improved her general condition and biochemical findings. This CVID case is complicated with intractable diarrhea and the functional disorder of renal tubules which is associated with the infiltration of CD 8 + lymphocytes in intestine and kidney. We consider that such case is very rare and valuable to report. PMID- 10863334 TI - [Words and tones]. PMID- 10863335 TI - [Research and informed consent]. PMID- 10863336 TI - [Who demands physicians' loyalty?]. PMID- 10863337 TI - [Male baldness]. PMID- 10863338 TI - [Burnout--a useful concept?]. PMID- 10863340 TI - [Socioeconomic status, self-assessed health and morbidity among Norwegian women aged 45-64]. AB - BACKGROUND: Information from the Norwegian Women and Cancer study was used in a study of the relationship between socio-economic status, place of residence and self-reported health and diseases. Total household income was used as a proxy for socio-economic status. MATERIAL AND METHODS: In 1997 a questionnaire was mailed to a random sample of 20,000 Norwegian women aged 45-69, and a sample of 10,000 women from Northern Norway aged 45-64. Only data on women aged 45-64 were included in the analysis. The response rate was 60.0%. RESULTS: More women in Northern Norway than in Southern Norway reported bad or very bad health. These geographical differences were reduced after adjustment for total household income, which was strongly correlated with self-reported health, physical as well as mental health, diabetes and to a lesser extent hypertension. INTERPRETATION: The study indicates that there are major differences in health according to social class in Norway. PMID- 10863339 TI - [What is burnout?]. AB - BACKGROUND: Burnout is most often described as a concept with three separate dimensions: emotional exhaustion, depersonalization (lack of empathy), and reduced accomplishments at work. We wanted to study the descriptive validity of the concept, which may be measured by the Maslach Burnout Inventory. MATERIAL AND METHODS: The Maslach Burnout Inventory was mailed to 1,476 members of the Norwegian Medical Association. The response rate was 73%. The dimensional structure of the instrument was examined by principal component analysis, and the identified factors correlated with validated measures of job satisfaction and depression. The dichotomized factors were combined in eight different ways, and the specificity of the resulting types was studied. RESULTS: The three original dimensions were reproduced, and the internal consistency of the factors was good (Cronbach's alpha ranging from 0.91 to 0.69). There were high correlations between emotional exhaustion and both job satisfaction (r = -0.54) and depression (r = 0.72). INTERPRETATION: Emotional exhaustion seems to be the least specific of the burnout dimensions. For the purpose of reasonable descriptive validity, the burnout notion should be based on both emotional exhaustion and depersonalization. With the applied dichotomization thresholds, this implies that 3% of Norwegian physicians are "burned out". PMID- 10863341 TI - [Prescription patterns of antihypertensive agents in general practice in 1995]. AB - BACKGROUND: Hypertension represents a risk factor in the development of cardiovascular disease. MATERIAL AND METHODS: The prescription of antihypertensive drugs and the daily doses used were examined in a questionnaire based survey among 2,586 drug-treated hypertensive patients who attended a general practitioner for clinical control in November 1995. RESULTS: The proportion of patients treated with one antihypertensive drug only was 63%. ACE inhibitors, alpha blockers and angiotensin-II antagonists constituted 22.7%, 6.6% and 4.4%, and betablockers, calcium channel blockers and diuretics 21.7%, 23.7% and 16.3% of the prescriptions respectively. Women were more often prescribed diuretics than men, and older individuals more often than younger persons. No striking difference in the prescription of antihypertensive drugs were observed between the five health regions of Norway. There was no reduction in the prescribed daily doses of antihypertensives used in combination therapy compared with monotherapy. In approximately one quarter of the prescriptions there was a positive correlation between the prescribed daily dose and the patient's weight. The number of antihypertensives prescribed did positively correlate with the patient's weight. INTERPRETATION: This survey shows that antihypertensives without any documented effect on morbidity and mortality were frequently prescribed for hypertension in general practice. Except for the low proportion of diuretic users among elderly patients, our results is in accordance with the Guidelines from the Norwegian Society of General Practitioners. PMID- 10863342 TI - [Long-term survival following surgery and radiotherapy of brain tumors in childhood]. AB - BACKGROUND: Brain tumours are seen in about one third of children with neoplastic disease. Treatment usually includes surgery and/or radiotherapy. Radiotherapy may have serious late effects, especially in children under the age of three; but is necessary for survival in children with medulloblastomas or high-grade gliomas. MATERIALS AND METHODS: We report ten and 20 years survival rates in 115 children with primary brain tumours (58 medulloblastoma, 14 high-grade gliomas, and 43 low grade gliomas) operated at the National Hospital and given radiotherapy at the Norwegian Radium Hospital during the years 1970-1995. RESULTS: No patients with medulloblastomas or high-grade gliomas relapsed after ten years. Overall ten and 20 years survival in children treated with radiotherapy to tumour doses > 50 Gy for medulloblastoma was 51.5% and for high-grade gliomas 20%. Median survival for patients with low-grade gliomas was not reached at 20 years, but these patients were still at risk for late deaths. INTERPRETATION: Long-term survival in children with high-grade gliomas or medulloblastoma equals cure, while late relapses may occur in low-grade gliomas. PMID- 10863343 TI - [The foundation for psychiatric treatment--from belief to science]. AB - Like other clinical disciplines, psychiatry has been met with increasing demands to document the scientific foundations for its treatments. The introduction of evidence-based medicine has contributed to this. Psychiatry holds a unique position compared to other medical specialties because it deals not only with physical or biological matters, but also with questions of meaning and mental phenomena. The scientific and clinical approach to psychiatry must incorporate both these aspects. This cannot be accomplished through a purely hermeneutic or a purely relativistic position. In isolation, these approaches cannot contribute to bridging the gap between research and clinical practice. Evidence-based medicine, on the other hand, through a stronger emphasis on empirical scientific evidence as the basis of treatment methods, enables us to secure better and safer therapies for our patients and augment the professional status of psychiatry. A critical attitude towards established "truths" will prevent stagnation in static or dogmatic positions, and thereby promote continuous development. Quality assurance and scientific validation is basically a question of ethics, hence we cannot avoid attending to these issues. PMID- 10863344 TI - [Goethe about medicine and his diseases]. PMID- 10863345 TI - [Medicine and literature--interpretation and discussion of literary texts in medical education]. AB - BACKGROUND: In medical education, using literary texts as a starting point for discussion and reflection, is now quite common. Lectures and seminars in literature and medicine have been given at the University of Oslo since 1996. We describe our pedagogical approach as a "thematic interpretation and discussion of literary texts", theoretically grounded in reader-oriented criticism and problem based learning (PBL). This article describes the kind of learning taking place during analysis and discussion of literary texts among medical students. MATERIAL AND METHODS: We evaluated a series of four elective seminars for first and second year students in the autumn 1998. We used qualitative approaches based on analysis of material from observations during seminars, reflections written by participants, and notes from a group interview at the end of the seminars. RESULTS: Analysis and discussion of literary texts may give medical students: A better understanding of communicative aspects in the doctor-patient-relationship. Experience with interpretation of narratives and insight into how differently we may interpret a story. Awareness of aspects and dilemmas related to being a doctor. Understanding of how people may experience illness and health. INTERPRETATION: Lectures and seminars on literary texts might be an important supplement to other practical or theoretical teaching for medical students. PMID- 10863346 TI - [Anton Chekhov--a self-deceiver?]. AB - The Russian author and doctor Anton Chekhov (1860-1904) died from tuberculosis in 1904. He had his first haemoptysis in 1884. In spite of continuing symptoms he did not let himself be examined by colleagues until he had a severe haemorrhage in 1897. He was then hospitalized, and extensive bilateral pulmonary tuberculosis was diagnosed. Even after that, he did not take his disease seriously. His attitude has been taken as an example of a doctor's self-deception. Almost 4,500 of Chekhov's letters has been published. In this article, the letters and contemporary memorial literature are used to illustrate the development of his disease and his ambivalence towards it. His self-deception was not that massive; there is a strain of uneasiness in his reports of his symptoms. PMID- 10863347 TI - [Alexander Pushkin's duel--biographic and medical aspects]. AB - The great Russian poet Aleksandr Pushkin (1799-1837) died 46 hours after being wounded by a pistol shot in a duel. The bullet penetrated the right pelvic bone, continued through the lower abdomen, and crushed the right part of the sacral bone. Biographical events leading to the duel are presented in the article, which also reviews articles in Russian medical journals describing the extent of the trauma and discussing the treatment possibilities at the time of the duel as well as present-day treatment. It is concluded that death was caused by peritonitis and that only modern extensive abdominal and orthopaedic surgery combined with antibiotic treatment could have saved the poet's life. PMID- 10863348 TI - [The dishonest, the ignorant and the insane artists--psychopathography and other paths to soul of the artist]. AB - Psychopathography is a specific kind of biography focusing on psychological and psychopathological aspects of the personality and their significance for creative activity, especially in famous persons. Psychopathography evolved as a discipline from the 1830's onwards, as a product of psychiatry emerging as a science, reflecting its reductionist and nosological approach. An illustration of this method is the analysis of the author E.T.A. Hoffmann, a prototypical representative of the German romanticism at the beginning of the century. Hoffmann himself expresses, in allegorical terms, the menacing, irrational and unconscious domains of the human soul. Romanticism may be seen as a forerunner of the psychoanalytic movement. Confronted with modernism in art, pathographic considerations were to a large extent based upon the object of art itself, often with arrogant and repudiating conclusions concerning the artist. Classical psychoanalysis has had a marked tendency to deduce a great deal about the personality of the artist or the writer solely from analysis of a picture or written text. The pathographic approach of the first century of scientific psychiatry has had a renaissance over the last 15-20 years: The ever increasing interest in affective syndromes has entailed a tendency to trace and identify affective pathology in artists and writers, deceased or alive, often emphasizing the affective dynamics of the creative process, aspects also noted by the classical pathographers. More recent studies of the creative personality may also present improved instruments for the study of the creative process. PMID- 10863350 TI - [How does music affect the human body?]. AB - Music therapy has developed its practice and research approaches within a qualitative framework more related to humanistic traditions than to medical science. Music medicine has therefore developed as a separate discipline, endeavouring to incorporate the legitimate therapeutic use of music within a medical framework. This paper argues that more extensive communication and collaboration between the methods developed within the music therapy community, and research based on medical science, could lead to a better understanding of the place of music as a therapeutic tool, both as regards its efficacy and its limits. Research has shown that music may influence central physiological variables like blood pressure, heart rate, respiration, EEG measurements, body temperature and galvanic skin response. Music influences immune and endocrine function. The existing research literature shows growing knowledge of how music can ameliorate pain, anxiety, nausea, fatigue and depression. There is less research done on how music, and what type of music, is utilized and administered specifically for optimum effect in specific clinical situations. PMID- 10863351 TI - [Examples of the use of music in clinical medicine]. AB - Music has been an element in medical practice throughout history. There is growing interest in music as a therapeutic tool. Since there is no generally accepted standard for how, when and where music should be applied within a medical framework, this literature study endeavours to present an overview of central areas of application of music in medicine. It further attempts to find tentative conclusions that may be drawn from existing clinical research on the efficacy of music as a medical tool. Traditionally, music has been linked to the treatment of mental illness, and has been used successfully to treat anxiety and depression and improve function in schizophrenia and autism. In clinical medicine several studies have shown analgetic and anxiolytic properties that have been used in intensive care units, both in diagnostic procedures like gastroscopy and in larger operations, in preoperative as well as postoperative phases, reducing the need for medication in several studies. The combination of music with guided imagery and deep relaxation has shown reduction of symptoms and increased well being in chronic pain syndromes, whether from cancer or rheumatic origin. Music has been used as support in pregnancy and gestation, in internal medicine, oncology, paediatrics and other related fields. The use of music with geriatric patients could prove to be especially fruitful, both in its receptive and its active aspect. Studies have shown that music can improve function and alleviate symptoms in stroke rehabilitation, Parkinson's disease, Alzheimer's disease and other forms of dementia. The role of music in medicine is primarily supportive and palliative. The supportive role of music has a natural field of application in palliative medicine and terminal care. Music is well tolerated, inexpensive, with good compliance and few side effects. PMID- 10863352 TI - [Franz Schubert--his life, music and diseases]. AB - The Austrian composer Franz Peter Schubert (1797-1828), the father of the German lied (song), was only 31 years old when he died. During his short life he wrote more than 1,000 pieces, among them 600 lieder, nine symphonies, 18 overtures, chamber music, 15 operettas and operas, six masses, and innumerable piano pieces. Included among the latter are 21 complete sonatas, eight impromptus, Wanderer Fantasie, dances and piano duets. When he was 26 years old he contracted syphilis and was given the conventional treatment at that time, mercury, which caused him a great deal of problems in the years that followed. However, his premature death was probably caused by typhoid fever. PMID- 10863353 TI - [Health and political leadership. When somatic or mental diseases hit the statesmen and politicians]. PMID- 10863354 TI - [The unconscious--friend or enemy? Some thoughts on the concept of dissociation]. PMID- 10863355 TI - [Physicians' work as artistic activity]. PMID- 10863356 TI - [Physician--the one-eye samaritan?]. PMID- 10863357 TI - [Care of the elderly in Oslo and Toulouse--all roads are leading to (one) Rome?]. PMID- 10863358 TI - [Resuscitation of newborn infants]. PMID- 10863359 TI - [BiliBed and treatment of neonatal jaundice]. PMID- 10863360 TI - [Venous thromboembolism--incidence and risk factors in Oslo]. PMID- 10863361 TI - [Prediction of developmental disorders]. PMID- 10863362 TI - [The population genetics of nodule bacteria]. AB - The data are reviewed on the population structure and evolutionary dynamics of the nodule bacteria (rhizobia) which are among the most intensively studied microorganisms. High level of the population polymorphism was demonstrated for the rhizobia populations using the enzyme electrophoresis (MLEE profiles). The average value of Nei's coefficient of heterogeneity (H = 1 - sigma pi2 [n/(n - 1)]) were: 0.590 for rhizobia (Rhizobium, Bradyrhizobium), 0.368 for enterobacteria (Escherichia, Salmonella, Shigella) and 0.452 for pathogenic bacteria (Bordetella, Borrelia, Erysipelothrix, Haemophilus, Helicobacter, Listeria, Mycobacterium, Neisseria, Staphylococcus) populations. In spite of being devoid of the effective systems for the gene conjugative transfer, many rhizobia populations possess an essentially panmictic structure. However, the enterobacteria populations in which the gene transfer may be facilitated due to the conjugative F- and R-factors, usually display the clonal population structure. The legume host plant is proved to be a key factor that determines the high levels of polymorphism and of panmixis as well as high evolutionary rates of the symbiotic bacteria populations. The host may ensure: a) an increase in mutation and gene transfer frequencies; b) stimulation of the competitive (selective) processes in both symbiotic and free-living rhizobia populations. A "cyclic" model of the rhizobia microevolution is presented which allows to assess the inputs the interstrain competition for the saprophytic growth and for the host nodulation into evolution of a plant-associated rhizobia population. The nodulation competitiveness in the rhizobia populations is responsible for the frequency-dependent selection of the rare genotypes which may arise in the soil bacterial communities as a result of the transfer of symbiotic (sym) genes from virulent rhizobia strains to either avirulent rhizobia or to the other (saprophytic, phytopathogenic) bacteria. Therefore, the nodulation competitiveness may ensure: a) panmictic structure of the natural rhizobia populations; b) high taxonomic diversity of rhizobia which was apparently caused by a broad sym gene expansion in the soil bacterial communities. The kin selection models are presented which explain evolution of the "altruistic" (essential for the host plant, but not for the bacteria themselves) symbiotic traits (e.g., the ability for symbiotic nitrogen fixation and for differentiation into non-viable bacteroids) in the rhizobia populations. These models are based on preferential multiplication of the nitrogen-fixing clones either in planta (due to an elevated supply of the nitrogen-fixing nodules with photosynthates) or ex planta (due to a release of the rhizopines from the nitrogen-fixing nodules). Speaking generally, interactions with the host plants provide a range of mechanisms increasing a genetic heterogeneity and an evolutionary potential in the associated rhizobia populations. PMID- 10863363 TI - [Theoretical approaches to the description of the morphology of organisms]. AB - Theoretical morphology should include methods that allow to determine limits of discrete variability of organisms of the given taxon. It is possible to determine the multitude of possible factors based on the demands of natural selection to the given structure under given conditions. The author presents a number of such multitudes that describe the variability of salamander tongue apparatus, kinematics of flapping flight in insects and wing shape of birds from the order Ciconiiformes. Another approach is based on mechanisms that rule the morphogenesis of individuals of the given taxon in particulate environment. Examples of morphogenetic multitude are illustrated by the variability of location of organs on plant stems, morphology of mollusc shells and colonies of hydroids polyps. All examples show that existing phenotypes occupy a compact area in the space of theoretically possible forms so the multitude of realized phenotypes does not have free space. If some free space is discovered it means that corresponding forms of life exist in nature but still are not registered and investigated. PMID- 10863365 TI - [The morphological and ontogenetic changes in the Protozoa during the transition to a sessile mode of life]. AB - The morphological adaptations of protozoans to sessile mode life and evolutionary changes in ontogeny are considered. There are main morphotypes of sessile protists: stalked organisms that attached to substrate by the extended base of body (basal disk), and unstalked organisms that are flatted on substrate. The origin of the morphotypes was independent in different taxa and involved nonhomologous structures. Adaptation to the sessile mode of life in the protists was connected with the progressive increase in the body size and intensity of organelle functions by polymerisation, subsequent division of function and change of functions. Evolution of adhesive organelles is characterised by growing intensity of their functions by allometric growth (usually without polymerisation), and in some cases with the subsequent division of functions and change of functions. The evolution manifests itself primarily in the organelles that provide interaction of cell with environment. The organelles that ensuring functioning of cell change due to correlations with the organelles of the first group. These two groups of organelles are similar to A.N. Sewertsoff's ecto- and endosomatic organs in multicellular organisms. The ontogeny of the sessile protists included three stages: formation of the migratory stage, distribution and choice of substrate and metamorphosis of the migratory stage after adhesion. As a rule there are no recapitulations on the first stage. The majority of structures tomotes or zoospores are inherited from the parent cell. Thus the present of some ancestral characteristics at the earlier stages of protistean ontogeny is display of the Baer's law. The main features of ontogeny evolution in sessile protists are the anaboly of the additional stages of life cycle, the displays of archallaxis or deviation during the migratory stage formation, and anaboly at the stage of buds morphogenesis after adhesion. At the last stage, the study of recapitulations is most perspective with the decision of phylogenetic problems in sessile protists. PMID- 10863364 TI - [The quantitative patterns of the modified one-center bioreceptor model]. AB - A modified one site model of the bioreceptor have been used to estimate quantitatively the phenomenon of the full agonism. Threshold phenomenon, spare receptors and linear dependence of the biological effect on concentration of complex agonist-receptor has been determinate by the general correlation equations. The equations of one site model provides a good fit to the experimental curves "dose-response" for the full agonists and allows to calculate the value of space receptors. The model includes occupancy Clark's theory and law "all or nothing". The interaction of acetylcholine and aklyltrimethylammonium salts with muscarinic acetylholine receptors is analysed as an example of use of this equation. PMID- 10863366 TI - [The ecological mechanisms of the formation of macrobenthos communities in the coastal area of small diverse lakes]. AB - The author analyses trophic structure of macro invertebrates inhabiting samples of litter settled (deciduous leaves) in three small lakes with different hydrochemical characters. In acid lake predators and detritovores-collectors are dominant, in slightly acid lake predators, detritovores-filtrators and collectors are the main types. Trophic structure of acid lakes is formed by acid-tolerant species adopted to low pH. In slightly acid lakes circumneutral species are developed. It was shown that trophic structure is formed mainly under influence of biotic factors and is not strongly depended on pH and trophic levels. Vertical distribution of organisms is determined by oxygen regime. PMID- 10863367 TI - [The variability of the reaction to a visual stimulus in goldfish, Carassius auratus L. (Cyprinidae: Pisces)]. AB - Animal's responses to repeated identical stimulation are notoriously variable. The variability is not always resulted from differences between individuals, stimulus action, or habituation. Goldfish's responses to a visual stimulus were studied in order to reveal the nature of variability. The experimental device consisted of a white circular corridor, and the black band stimulus drown across the corridor. Fish responded to the stimulation by an increase in frequency of turns in the vicinity of the band. The response, however, was not always positive: sometimes fish passed by the band with no turns (negative response). The significant tendency to repeat positive and negative responses by series was observed. Intensity of the response differed between individuals, and some individuals never responded at all. The simulating dynamic model of goldfish behaviour was studied both qualitatively and quantitatively. The model assumes that the control mechanism of the behaviour is influenced by the noise (fluctuations of nervous activity) which causes phase transitions in the mechanism. The similarity of the model's behaviour to experimental data suggests that the response variability observed in goldfish resulted from the self organising process. Therefore, the variability should persist even in the absence of any variations in an external stimulation, as well as in the absence of individual differences. PMID- 10863368 TI - [Patterns of spiral phyllotaxy]. AB - The author investigates theoretical variants of spiral phyllotaxy using structural coefficient--the distance between consequent members in genetic spiral, expressed in parts of circle. The meanings of structural coefficient in which parastichies change their isomerism are determined as critical points. The author describes the overlapping of parastichies of lower lane by higher lane parastichies and presents the methods of overlapping determination. Parastichies with minimal distance between members in lane are clearly determined on shoots. Methods and formulas for calculation of structural coefficient using the number of members in parastichy circle are presented. Orthostichy is considered as a special case of parastichy location. Orthostichies are equal to parastichies in maximum critical points. PMID- 10863369 TI - [The molecular genetic typification of planarians in the genus Bdellocephala (Dendrocoelidae, Tricladida, Turbellaria) from Lake Baikal with an assessment of their species diversity]. AB - Baikal planaria from genus Bdellocephala were typified using rDNA locus coding 5' -end domain of 18S ribosome RNA. Five colour forms of 24 possible variants that differ in diapason 0-1.3% of genotype were determined by comparative analysis of nucleotide sequences. The authors use back colour--one of the most variable and typical character in the given group--to collect material for investigation. It allows to minimize the size of investigation sample and at the same time to cover maximum variability of Bdellocephala. One of the positive result of molecular typification of colour forms was a discovery of unique individuals that belong to new species. Karyological analysis of colour forms shows variations in chromosome numbers that divide planaria into 3 groups (2n = 20, 24, 26). Comparative analysis of morphological and ecological characters and karyotypes of some forms united by the same genotype allows to distinguish them as separate species. Criteria of modern phenetic system of Baikal planaria are discussed. PMID- 10863370 TI - Development of a classification system for periodontal diseases and conditions. AB - Classification systems are necessary in order to provide a framework in which to scientifically study the etiology, pathogenesis, and treatment of diseases in an orderly fashion. In addition, such systems give clinicians a way to organize the health care needs of their patients. The last time scientists and clinicians in the field of periodontology and related areas agreed upon a classification system for periodontal diseases was in 1989 at the World Workshop in Clinical Periodontics. Subsequently, a simpler classification was agreed upon at the 1st European Workshop in Periodontology. These classification systems have been widely used by clinicians and research scientists throughout the world. Unfortunately, the 1989 classification had many shortcomings including: 1) considerable overlap in disease categories, 2) absence of a gingival disease component, 3) inappropriate emphasis on age of onset of disease and rates of progression, and 4) inadequate or unclear classification criteria. The 1993 European classification lacked the detail necessary for adequate characterization of the broad spectrum of periodontal diseases encountered in clinical practice. The need for a revised classification system for periodontal diseases was emphasized during the 1996 World Workshop in Periodontics. In 1997 the American Academy of Periodontology responded to this need and formed a committee to plan and organize an international workshop to revise the classification system for periodontal diseases. The proceedings in this volume are the result of this reclassification effort. The process involved development by the Organizing Committee of an outline for a new classification and identification of individuals to write state-of-the-science reviews for each of the items on the outline. The reviewers were encouraged to depart from the preliminary outline if there were data to support any modifications. On October 30-November 2, 1999, the International Workshop for a Classification of Periodontal Diseases and Conditions was held and a new classification was agreed upon (Fig. 1). This paper summarizes how the new classification for periodontal diseases and conditions presented in this volume differs from the classification system developed at the 1989 World Workshop in Clinical Periodontics. In addition, an analysis of the rationale is provided for each of the modifications and changes. PMID- 10863371 TI - Dental plaque-induced gingival diseases. AB - Gingival diseases are a diverse family of complex and distinct pathological entities found within the gingiva that are the result of a variety of etiologies. There are several clinical characteristics common to all gingival diseases and these features include clinical signs of inflammation, signs and symptoms that are confined to the gingiva, reversibility of the disease by removing the etiology, the presence of bacterial laden plaque to initiate and/or exacerbate the severity of the disease and a possible role as a precursor for attachment loss around teeth. Defining and classifying gingival diseases has not been an easy task. The tools and methods to identify gingival diseases have varied depending on the criteria used by epidemiologists, researchers, or the practicing clinician. The classification of gingival disease in this review relied upon experimental and/or epidemiological human studies that accurately and reliably assessed an underlying functional derangement that was localized to the gingiva and was reported in a peer-reviewed journal. The classification of gingival diseases that depends on dental plaque to initiate the disease process(es) has been categorized into two groups. The two categories of plaque-induced gingival diseases are those affected by local factors and those that are affected by local factors and modified by specific systemic factors found in the host. In this review, the clinical characteristics of gingival disease associated with plaque, endogenous hormone fluctuations, drugs, systemic diseases, and malnutrition were investigated. PMID- 10863372 TI - Non-plaque-induced gingival lesions. AB - The origin of gingival inflammation is occasionally different from that of routine plaque-associated gingivitis, and such non-plaque-associated types of gingivitis often present characteristic clinical features. Examples of such forms of gingivitis are specific bacterial, viral, and fungal infections. Specific bacterial infections of gingiva may be due to Neisseria gonorrhea, Treponema pallidum, streptococci, and other organisms. The most important viral infections of gingiva are herpes simplex virus type 1 and 2 and varicella-zoster virus. Fungal infections may be caused by several fungi, the most important of these being Candida species including C. albicans, C. glabrata, C. krusei, C. tropicalis, C. parapsilosis, and C. guillermondii. Gingival histoplasmosis is a granulomatous disease caused by the fungus Histoplasma capsulatum and, as for the other specific infections of gingiva, a confirmed diagnosis may require histopathologic examination and/or culture. Atypical gingivitis may also occur as gingival manifestations of dermatological diseases, the most relevant of these being lichen planus, pemphigoid, pemphigus vulgaris, erythema multiforme, and lupus erythematosus. Non-plaque induced gingival inflammation can be caused by allergic reactions to dental restorative materials, toothpastes, mouthwashes, and foods. In addition, gingival inflammation may result from toxic reactions, foreign body reactions, or mechanical and thermal trauma. PMID- 10863373 TI - Periodontitis. AB - The purpose of this review was to assess the scientific and clinical bases for the proposed classification of periodontitis. The clinical and histopathological signs and the etiology of periodontitis were described. Cross-sectional studies were analyzed to determine when onset of periodontitis most frequently occurs in adults. In addition, the progression rates of periodontitis have been assessed from longitudinal studies. No clinical, histopathological, or microbiological features could be identified that would characterize different disease entities of chronic periodontitis. The prevalence, extent, and severity of periodontitis were found to increase continually with higher age and there was no age when onset of disease would most likely occur. The rate of periodontitis progression varies largely between patients and there is no natural threshold for distinguishing various rates of disease progression. The incidence of periodontitis unresponsive to treatment depends on pretreatment progression rate, extent and severity of disease, tooth type, smoking, high levels of putative periodontal pathogens, a deficient immune response, and the type of therapy provided. There is no scientific basis for the classification "adult periodontitis" and "refractory adult periodontitis." Extensive clinical examinations are required for the diagnosis of "rapidly progressive adult periodontitis." It appears unrealistic that these examinations can be performed routinely in clinical practice. Therefore, the classification proposed by the Organizing Committee to define adult, rapidly progressive, and refractory periodontitis as specific disease entities was replaced with a simplified classification of periodontitis based on the scientific data available. PMID- 10863374 TI - Early-onset periodontitis. AB - In 1993, the 1st European Workshop on Periodontology explicitly recognized that there was insufficient knowledge to differentiate truly different forms of periodontal disease from differences in the presentation/severity of the same disease. In spite of recent progress in our understanding of periodontal diseases, the issue is far from having been resolved. Classification of periodontal diseases, therefore, remains based upon the definition of specific clinical syndromes. Early-onset periodontitis (EOP) is one such syndrome and comprises a group of pathological conditions leading to loss of periodontal tissues early in life. The notion that classifies periodontitis syndromes as "early-onset" or "adult" is primarily epidemiological in nature and is based on the observation that periodontitis is rather infrequent in children and young adults. Nevertheless, considerable epidemiological evidence indicates that periodontitis does affect children and young adults to a level of severity that may lead to premature exfoliation of primary and/or permanent teeth. Clinical presentation of periodontitis early in the life of an individual is thought to indicate that the etiologic agents have been able to cause considerable tissue damage over a relatively short period of time. It also implies either infection with highly virulent bacteria and/or a highly susceptible subject. The purpose of this review is to discuss the criteria generally utilized to classify EOP, provide the rationale to designate EOP as a distinct disease entity, and to review the evidence justifying a subclassification into particular subgroups of EOP. PMID- 10863375 TI - Periodontitis modified by systemic factors. AB - Microbial dental plaque is the initiator of periodontal disease but whether it affects a particular subject, what form the disease takes, and how it progresses, are all dependent on the host defenses to this challenge. Systemic factors modify all forms periodontitis principally through their effects on the normal immune and inflammatory defenses. Some good examples of this effect exist such as when there is a reduction in number or function of polymorphonuclear leukocytes (PMNs) that may result in an increased rate and severity of periodontal destruction. Many other systemic factors are much less clear cut and are difficult to causally link to periodontitis. In many cases the literature is insufficient to make definite statements on links between systemic factors and periodontitis. It is also at times difficult to be precise regarding the causative agent in systemic exposures such as smoking and even prescribed drug therapy. The possible role of systemic diseases and systemic exposures in initiating or modifying the progress of periodontal disease is clearly a complex issue. It is however generally agreed that several conditions may give rise to an increased prevalence, incidence, or severity of gingivitis and periodontitis. The categorization of the systemic modifying factors causing periodontitis and the evidence to support the role of these factors are the focus of this review. An attempt has been made to consider the conditions under broad headings, but it will be clear that many conditions fall within more than one category and that for several conditions only case reports exist whereas in other areas an extensive literature is present. PMID- 10863376 TI - Necrotizing ulcerative gingivitis. AB - Necrotizing periodontal diseases are unique in their clinical presentation and course. Data suggest that the etiology and pathogenesis of necrotizing periodontal diseases may also be distinctive from other periodontal diseases. Necrotizing ulcerative gingivitis (NUG) is a type of necrotizing periodontal disease in which the necrosis is limited to the gingival tissues. Three specific clinical characteristics must be present to diagnose NUG, pain (usually of rapid onset) interdental necrosis, and bleeding. Epidemiological and prospective clinical studies have found an altered ability to cope with psychological stress, immunosuppression, and tobacco use to be strongly associated with the onset of NUG. PMID- 10863377 TI - Necrotizing ulcerative periodontitis. AB - In patients with no known systemic disease or immune dysfunction, necrotizing periodontitis (NUP) appears to share many of the clinical and etiologic characteristics of necrotizing ulcerative gingivitis (NUG) except that patients with NUP demonstrate loss of clinical attachment and alveolar bone at affected sites. In these patients, NUP may be a sequela of a single or multiple episodes of NUG or may be the result of the occurrence of necrotizing disease at a previously periodontitis-affected site. The existence of immune dysfunction may predispose patients to NUG and NUP, especially when associated with an infection of microorganisms frequently associated with periodontal disease such as Treponema and Selenomonas species, Fuscobacterium nucleatum, Prevotella intermedia, and Porphyromonas gingivalis. The role of immune dysfunction is exemplified by the occasionally aggressive nature of necrotic forms of periodontal disease seen in patients with HIV infection or malnutrition, both of which may impact host defenses. Clinical studies of HIV-infected patients have shown that patients with NUP are 20.8 times more likely to have CD4+ cell counts below 200 cells/mm3. However, these same studies have demonstrated that most patients with CD4+ cell counts below 200 cells/mm do not have NUP, suggesting that other factors, in addition to immunocompromisation, are involved. Further studies are needed to define the complex interactions between the microbial, or viral, etiology of necrotic lesions and the immunocompromised host. It is, therefore, recommended that NUG and NUP be classified together under the grouping of necrotizing periodontal diseases based on their clinical characteristics. PMID- 10863378 TI - Periodontal abscess. AB - This review focuses on the classification of periodontal abscesses, which are localized purulent infections of periodontal tissues, and discusses their etiology and clinical characteristics. PMID- 10863379 TI - Periodontic-endodontic lesions. AB - Lesions of the periodontal ligament and adjacent alveolar bone may originate from infections of the periodontium or tissues of the dental pulp. This review focuses on the relationship of lesions of endodontic origin with lesions of periodontal origin and their classification. PMID- 10863380 TI - Tooth-related issues. AB - Several conditions exist around teeth that may predispose the periodontium to disease. These situations may occur as a result of the condition or position of teeth or as a result of tooth treatment. In certain cases these tooth-related factors may contribute to the initiation of periodontal disease. While the etiology of periodontal disease is bacterial, factors that enhance bacterial accumulation or allow the ingress of bacteria into the periodontium should be considered in the classification and diagnosis of periodontal diseases. This is because many times these tooth-related issues can cause site-specific problems that require treatment in an otherwise intact periodontium. Several factors related to tooth/root anatomy, restorative, and endodontic considerations have been associated with gingival inflammation, attachment loss, and bone loss. These factors will be reviewed as they relate to their potential to promote damage to the periodontium. PMID- 10863381 TI - Mucogingival deformities. AB - The presence of mucogingival deformities often have an impact on patients in terms of esthetics and function. The variety of the conditions makes it difficult to place gingival and alveolar mucosa deformities under a single definition. Mucogingival deformities, as defined in this paper, may be congenital, developmental, or acquired defects. These may occur around natural teeth or implants and in edentulous ridges. They may be localized to soft tissues or be associated with defects in the underlying bone. They may show different degrees of severity and extension. A classification of mucogingival deformities should provide a method for identifying the different conditions in order to improve diagnosis, etiologic identification, research, treatment, and insurance evaluation. PMID- 10863382 TI - Occlusal trauma: effect and impact on the periodontium. AB - This focused review is limited to a number of investigations in an attempt to specifically address the histological and clinical effects of excessive occlusal forces on the teeth and periodontium and to provide a basis of classification for this interaction. This review does not include the effects of occlusal forces on dental implants or dental prostheses/appliances. PMID- 10863383 TI - Effect of storage and acid etching on the tensile bond strength of composite resins to glass ionomer cement. AB - This in vitro study evaluates the effect of storage time and acid etching on the tensile bond strength of glass ionomer cement to composite resins. The bonded assemblies were stored at 100% relative humidity and 37 degrees C for 1 hour, 1 day, 1 week, 1 month and 3 months. The test specimen was loaded at tension to failure on an Otto Wolpert-Werke testing instrument with a crosshead speed of 6 mm/min. The results showed a significant statistical difference for etched Vidrion F when compared to etched Ketac Bond at all storage periods. The unetched samples were statistically similar at 3 months, with the highest values for Vidrion F. PMID- 10863384 TI - Scanning electron microscopy of the rat tongue mucosa with special attention to the bacteria on epithelial cell membranes. AB - The authors examined the filiform and fungiform papillae surfaces of rat tongue by scanning electron microscopy showing the numerous groupings of bacteria on the epithelial cell membranes. The fungiform papillae were round in shape and present few bacteria. The epithelial cell of filiform papillae revealed numerous streptococci. The grouping of the bacteria are attached on the epithelial cell membrane, demonstrating three-dimensional SEM images. PMID- 10863385 TI - Evaluation of the antimicrobial activity of three irrigating solutions in teeth with pulpal necrosis. AB - The antimicrobial activity of 0.4% papaine gel (FCF-USP), an antibacterial product derived from 3.3% castor oil (IQSC-USP), and 0.5% sodium hypochlorite (FORP-USP) was evaluated in teeth with radiographically visible pulpal necrosis and periapical lesion in vivo. After cavity access, under aseptic conditions, a first harvesting was performed. The 3 irrigating solutions were used for biomechanical preparation. After 72 hours, a second harvesting was performed, also under aseptic conditions. The number of colony forming units (cfu) was counted with a stereomicroscope under reflected light. Castor oil and 0.5% sodium hypochlorite presented similar antimicrobial activities for the reduction of the anaerobe number, S. mutans and streptococci; however, the papaine gel showed lower activity. We conclude that both castor oil and sodium hypochlorite are effective as antimicrobial agents and can be used in the treatment of root canals with pulpal necrosis. PMID- 10863386 TI - Behavior of dental phobic residents of large and small communities. AB - The purpose of the present study was to characterize differences among dental phobic patients in past dental experience between residents of standard metropolitan areas (SMSA), and non-SMSA. Over a period of 1 year, the records of 41 dental phobic patients were evaluated. These patients were attending a special Israeli Defense Force clinic designed to treat dental phobics. The mean time since the last routine visit in both groups was 12.42 +/- 6.54 years and the mean age at which the patients had had their last routine dental treatment was 14.56 +/- 4.62 years. Over a two-year period prior to the present visit, the main reason for SMSA patients to seek any dental care was dental pain (P = 0.015). All other variables that compared the dental behavior of SMSA patients with non-SMSA patients showed no difference. The findings suggest that the availability of dentists in the SMSA did not increase the seeking of dental care. The fact that SMSA residents were more likely to seek care because of pain suggests that residents of non-SMSA areas were more likely to tolerate pain without seeking care. These findings underscore the need for proper behavioral and pain management during childhood. PMID- 10863387 TI - Analysis of three dental fluorosis indexes used in epidemiologic trials. AB - The objective of this paper was to compare the DEAN, T-F and TSIF dental fluorosis indexes in relation to prevalence of surfaces, teeth and locality, and to verify the statistical correlation among them. The sample consisted of 461 schoolchildren, ages 12-14 years, born and reared in 3 cities in the State of Sao Paulo from 2 years of age. A total of 153 were from Cesario Lange with a fluoride concentration in the water supply of 1.4 ppm F, 142 from Piracicaba (0.7 ppm F) and 166 from Iracemapolis (< 0.3 ppm F). The clinical examination was conducted after tooth brushing, using a plane mirror, artificial light and air drying of the teeth for 1 min. Premolars, second molars, and occlusal surfaces were the most severely affected. The three indexes showed similar percentages of children affected in the 3 three cities: 32.7%, 16.9% and 4.2% for the DEAN index, 33.3%, 17.6% and 4.2% for the T-F index and 32.7%, 16.9% and 4.2% for the TSIF index. There were no difficulties in using the three indexes in the field trials; thus the use of any one may be recommended in regions with similar fluoride concentrations to those of this study. PMID- 10863388 TI - Staphylococci from dental personnel. AB - Thirty dental students and five professors were cultured in nares, throat, and hands for the presence of staphylococci. Twenty-four students and two professors were colonized with staphylococci that were classified as S. aureus. Twelve students and one professor were colonized with staphylococci that produced enterotoxin. Care needs to be taken to avoid contaminating patients during dental examination, particularly during any type of surgery. PMID- 10863389 TI - Telediagnosis of oral disease. AB - Computers have increasingly found application in dentistry over the past 15 years, but at present there has been no investigation of the application of the Internet for distance diagnosis purposes in oral medicine. As a consequence, the objective of this article was to determine the acceptability to patient and clinician of the distant diagnosis of common orofacial diseases using the Internet. The study group comprised 20 patients who attended the Oral Medicine unit of the Eastman Dental Institute and Hospital, London, UK, for the diagnosis and management of oral mucosal diseases. Digital images of each patient's oral mucosal lesion were captured and stored on a personal computer and later transmitted via the Internet to a distant site. Patients were asked to complete a self-administered questionnaire detailing their opinion of the use of an intra oral camera and a group of clinicians were asked to compare and contrast the original and transmitted images. The majority of patients found the procedure of recording images of their mouth very comfortable, were happy to view the inside of their mouths, and found the procedure generally useful in understanding their clinical problem. The clinicians were often not able to differentiate between the original and transmitted image but were able to accurately diagnose the patient's oral mucosal problems in 64% of the instances. The results of the present study suggest that telediagnosis of orofacial disease may be a feasible prospect. PMID- 10863390 TI - Periapical cemental dysplasia: case report. AB - The authors present a case of periapical cemental dysplasia affecting the mandibular left canine, with vital pulp, in a 43-year-old black female patient, an occurrence that follows the classical cases found in the literature. The need of a careful history, clinical and radiographic exams and vitality tests are emphasized in order to reach the correct diagnosis of this disease. PMID- 10863391 TI - Effect of vehicle on antimicrobial properties of calcium hydroxide pastes. AB - The current discussion about the importance of intracanal dressings and the effect of vehicles on calcium hydroxide pastes is justified by controversy concerning the achievement of complete disinfection after preparation of infected root canals and the real antimicrobial effect of these vehicles. The aim of this study is to investigate the role of vehicles in the antimicrobial effect of calcium hydroxide pastes. Well-conducted research about the characteristics of calcium hydroxide, such as antimicrobial potential, physico-chemical aspects and histocompatibility, gives credibility to the choice of this medication in several clinical situations. Different vehicles have been added to calcium hydroxide in an attempt to enhance its properties. Scientific reasoning indicates the use of hydrosoluble vehicles (distilled water, saline) associated with calcium hydroxide because of their chemical characteristics of dissociation, diffusibility and filling capability which are decisive for the biological behavior, i.e., antimicrobial qualities and induction of tissue repair. PMID- 10863392 TI - Immediate replantation of maxillary incisors in rats: effects of tooth immersion in sodium fluoride and subsequent removal of the periodontal ligament. AB - The advantages and disadvantages of maintaining the periodontal ligament (PDL) in immediate replantation as well as chemical treatment of the root surface have been a matter of discussion because the vitality of such tissue in surgery is always questioned. This study evaluated the effects of conserving the tooth in sodium fluoride and the removal of the PDL before replantation of incisors in rats. There was more cementum-dentin resorption in the group with the PDL. The group without the PDL showed more discreet resorption, repair occurred through the newly formed bone tissue in the PDL space and ankylosis was more extensive than in the group with the PDL. PMID- 10863393 TI - Scanning electron microscopy study of the effect of tetracycline HCl on smear layer removal and fibrin network formation. AB - Scanning electron microscopy (SEM) was used to evaluate root surface characteristics of human teeth affected with periodontitis following periodontal instrumentation and topical application of tetracycline HCl (TTC-HCl; pH 1.6; 4 min). Specimens were randomly assigned to periodontal instrumentation alone (control 1); periodontal instrumentation plus TTC-HCl (test 1); periodontal instrumentation plus trypsin solution after extraction (control 2); and periodontal instrumentation plus TTC-HCl plus trypsin solution after extraction (test 2). Tetracycline solution was applied with a cotton pellet. Twenty-two single root periodontitis affected human teeth scheduled for extraction were selected. Mucoperiosteal flaps were raised, root surfaces were mechanically and chemically treated, flaps were repositioned and maintained in place for 20 min. Teeth were extracted, rinsed and placed in cold phosphate buffer solution (PBS) and control 2 and test 2 groups were treated with trypsin solution. Specimens were examined using SEM. Smear layer was successfully removed, exposing dentinal tubules; however, fibrin network formation in situ was not improved by application of TTC-HCl. PMID- 10863394 TI - Immunocytochemical study of giant cell fibroma. AB - Giant cell fibroma (GCF) is a non-neoplastic lesion of the oral mucosa. The origin of stellate and multinucleate cells of GCF is not well known. The purpose of the present article was to investigate the immunoreactivity of these cells for leukocyte common antigen, vimentin, tryptase, HLA-DR, alpha-smooth muscle actin, CD68, and S-100. The results showed positive staining only for vimentin. This suggests that the stellate and multinucleate cells of GCF have a fibroblast phenotype. PMID- 10863395 TI - Correlation between the CPITN score and anaerobic periodontal infections assessed by BANA assay. AB - In the present investigation the ability of subgingival plaque to hydrolyze BANA (Perioscan) was correlated with CPITN scores. Among 281 sites investigated, 136 had a CPITN equal to 2 with a highly significant positive BANA value (107 sites). A CPITN equal to 3 was also significantly BANA positive (90 sites). These findings clearly demonstrate the relationship between CPITN and anaerobic microorganisms (BANA positive). PMID- 10863396 TI - First permanent molar: first indicator of dental caries activity in initial mixed dentition. AB - The objective of the present study was to investigate among children in the initial mixed dentition phase the presence of clinical signs that might eventually function as more sensitive indicators of the development of caries disease, denoted here as caries activity. On this basis, we investigated the relationship between salivary levels of mutans streptococci (MS) and decayed, missing and filled permanent and deciduous tooth surfaces (DMFS and dmfs) using microbiological, clinical and radiographic examinations in 81 schoolchildren aged 7-8 years. Whereas dmfs did not present a positive correlation, DMFS was significantly correlated with salivary MS levels. The first permanent molars of the schoolchildren studied comprised 87.3% of the affected surfaces recorded in the DMFS, suggesting that the development of new lesions was preferentially located on the surfaces of the first permanent molars. These results permit us to conclude that the first permanent molars function as first indicators of dental caries activity in the schoolchildren examined. PMID- 10863397 TI - Five-year follow-up of internal bleaching. AB - A 5-year clinical follow-up study was conducted to determine the longevity of originally acceptable results for internal bleaching. The "walking internal bleaching" method is an acceptable technique of lightening discolored anterior teeth to provide an esthetically pleasing match with adjacent teeth. This follow up study revealed a success rate of 79% for all indications after 5 years. If the indication is limited to one palatal endodontic opening, the total success rate can reach 91%. However, a 98% success rate is common when subjective evaluation of patients is considered. The results clearly indicated that internal bleaching provides long-term success for treatment of discolored nonvital anterior teeth over a period of years and does not have any detrimental effect on dental hard tissue. In ideal cases, if the procedure is performed precisely, the success rate can exceed 90% after five years. PMID- 10863398 TI - Hypomaturation amelogenesis imperfecta: account of a family with an X-linked inheritance pattern. AB - Amelogenesis imperfecta (AI) is a heterogeneous genetic disorder which affects the dental enamel. It can have an autosomal dominant, autosomal recessive or X linked pattern. The authors describe a case of a family with hypomaturation X linked AI and discuss the clinical and histopathological aspects of this disorder. PMID- 10863399 TI - Oral manifestation of tertiary syphilis: case report. AB - This paper describes a case of benign tertiary syphilis represented by a solitary hypertrophic lesion on the dorsum surface of the tongue. The diagnosis was confirmed by serologic tests (VDRL and FTA-ABS). Histopathological analysis of biopsy specimens revealed, in the lamina propria, the presence of well-developed granulomas associated with necrotic areas (gummatous lesion). Currently, tertiary syphilis is rarely seen; however, this case emphasizes that it still exists and must be considered in the differential diagnosis of inflammatory oral lesions. PMID- 10863400 TI - In vitro study of the effects of endothelin-1 on human dental pulp cells. AB - OBJECTIVE: To study the effect of Endothelin-1 (ET-1) on cell proliferation in primary cultures of human dental pulp cells; the cell mechanism of mitogen; and collagen synthesis. METHODS: Primary cultures of human dental pulp cells; Thiazolyl Blue Tetrazolium Bromide (MTT) test to investigate cell proliferation; measurement of calcium ions in human dental pulp cells with the calcium-sensitive dye fluo-3; enzyme-linked immunosorbent assay and avidin-biotin complex immunocytochemical assay for collagen synthesis in human dental pulp cells. RESULTS: Incubation of cultures with ET-1 resulted in a dose-dependent increase in cell proliferation over control (unstimulated) cultures. In addition ET-1 increased the Ca2+ concentration in human dental pulp cells, which cannot be inhabited by Verapamil. The finding demonstrated that collagen type I synthesis of human dental pulp cells was increased by ET-1. CONCLUSIONS: ET-1 played a crucial role in pulp restoration of lesions and had an effect on mitogen and differentiation of human dental pulp cells. PMID- 10863401 TI - cDNA cloning and sequencing of MH2 domain of Smad2 from human dental pulp cells. AB - OBJECTIVE: The purpose of this study was to clone and sequence the cDNA of MH2 domain of Smad2 from human dental pulp cells. METHODS: In this study, total RNA was isolated from primary cultured human dental pulp cells and reverse transcribed into cDNA. The desired DNA product was obtained by nested PCR with 4 smad2 MH2 domain-specific primers. The segment was inserted into pBluescript II SK vector and the resulting plasmid was transformed into E. coli JM109. The double-strand cDNA of positive clone was sequenced by PE317-A automatic sequencing. RESULTS: cDNA of MH2 domain of Smad2 was obtained from human dental pulp cells. The sequence was consistent with that cloned from a human kidney cDNA library. No mutation was found. CONCLUSION: This study provides the first evidence of expression of smad2 in human dental pulp cells, and indicates that TGF-beta signaling may be mediated by Smad2 in human dental pulp cells. The cDNA cloned in pBluescript/S2MH2 could be used for further functional studies of Smad2 and MH2 domain in dental pulp cells. PMID- 10863402 TI - Detection of small lesions in alveolar bone by digital subtraction radiography. AB - OBJECTIVE: To evaluate the effectiveness of digital subtraction radiography (DSR) system for diagnosis of small artificial alveolar bone lesions. METHODS: Seventy two 0.2-0.9 mm3 lesions were produced in premolar and molar areas of dry human mandibles. Reference and subsequent radiographs were taken before and after lesions were made. The small lesions were detected by conventional radiography and DSR respectively. RESULTS: For 0.2-0.5 mm3 lesions, detection by both DSR and conventional methods was less than 50%. For 0.7 mm3 lesions, the detection by DSR and conventional methods was 57.5% and 38.7%, respectively (P < 0.01). For 0.9 mm3 lesions, the accuracy of the detection by DSR and conventional methods was 89.3% and 57.3% (P < 0.01). CONCLUSION: The detectability of small alveolar bone lesions by DSR was superior to that of the conventional method. PMID- 10863403 TI - Immunolabeling of the major cell surface protein antigen of Streptococcus sobrinus with monoclonal antibody. AB - OBJECTIVE: The purpose of this study was to determine the accessibility of monoclonal antibody (McAb), specific for the major cell surface protein antigen (PAg) of Streptococcus sobrinus, to the surface of its native epitopes. MATERIALS AND METHODS: An indirect immunogold labeling technique was used to detect the reaction of McAb with S. sobrinus 6715. The reactions of polyclonal antibodies (PcAbs) against S. sobrinus 6715 or PAg with S. sobrinus 6715, S. mutans Ingbritt C and S. rattus BHT were studied as controls. RESULTS: The results indicated that PAg was localized on the outer cell surface of S. sobrinus, and McAb was reactive with only a few epitopes of the cell surface, whereas PcAbs were found to be reactive with more epitopes. CONCLUSIONS: McAb was specific for the PAg, but there was cross-reaction with S. mutans. Also there seemed to be an association between the fuzzy coat on the surface of S. sobrinus and PAg. PMID- 10863404 TI - Magnetic resonance imaging of the upper airway in obstructive sleep apnea before and after oral appliance therapy. AB - OBJECTIVE: To determine the mechanism by which an oral appliance may be used to treat obstructive sleep apnea syndrome (OSAS). METHODS: Eleven OSAS patients (8 males and 3 females) were involved in the study. Mean age was 52.2 +/- 10.6 years; height was 166.6 +/- 7.2 cm, and weight was 75.6 +/- 9.3 kg; body mass index (BMI) was 27.2 +/- 2.9 kg/m2. Each patient underwent magnetic resonance imaging (MRI) and polysomnography before and after oral appliance therapy. Pharyngeal changes were measured and compared with the variation in sleep parameters. Also, Pearson's correlation and multiple linear regression were performed to investigate the relationship of sleep parameters and MRI items. RESULTS: Through oral appliance therapy, the sleep disorder decreased. Apnea/hypopnea index (AHI) decreased from 44.6 +/- 21.5 to 9.6 +/- 6.3 per hour of sleep. Lowest oxygen desaturation rose from 71.4 +/- 15.0% to 82.0 +/- 7.7%. Meanwhile, the upper airway increased at most levels, and especially at oropharynx. As measured by the correlation and regression analysis, the AHI changes had a negative association with tongue volume (R = -0.5730) and a positive association with the area alternation of high oropharynx (R = 0.5823); the change of the lowest oxygen desaturation (SaO2%) was positively associated with whole airway volume (R = 0.6554). CONCLUSION: The oral appliance works by enlarging the upper airway morphology and keeping the airway open, mainly at the back of soft palate. The effect of the oral appliance is associated with the degree of enlargement of the high oropharynx. Those who have a small tongue and a large pharynx may expect to have good results with the use of the oral appliance. PMID- 10863405 TI - Ectomesenchymal stem cells and all-trans-retinoic acid induced apoptosis. AB - OBJECTIVE: This study aimed to establish and characterize ectomesenchymal stem cells and further reveal the effects of all trans-retinoic acid (RA) on the biologic behaviors of these cells. METHODS: Ectomesenchymal stem (EMS) cells of developing palatal processes were explanted from the palatal shelves of embryonic BALB/c mice. The characterization of EMS cells was accomplished by immunohistochemical analysis and growth curves. The action of RA on EMS cells was evaluated according to population doubling time and the terminal deoxyribonucleotidyl transferase dUTP nick end-labeling assay. RESULTS: Primary culturing of EMS cells proceeded successfully. The results of immunohistochemistry showed that EMS cells stained neuron-specific enolase (+), S 100 (+), vimentin (+3), keratin (-), myoglobin (-) and factor VIII (-). The population doubling time of RA-treated EMS cells was much longer compared with nontreated EMS cells. RA also dramatically increased the number of apoptotic cells. CONCLUSION: Ectomesenchymal stem cells may be undifferentiated. RA inhibited their proliferation and induced apoptosis. The inhibition of growth and excess apoptosis may contribute to the formation of cleft lip/palate and other orofacial congenital malformations. PMID- 10863406 TI - Telomerase hTR and hTRT gene expression in oral precancerous lesions and squamous cell carcinomas. AB - OBJECTIVE: To investigate the expression of telomerase genes in oral precancerous lesions and squamous cell carcinomas and to research the relationship between telomerase gene expression and carcinogenesis of oral mucosa epithelium. MATERIALS AND METHODS: Eighty two cases were detected for hTR and hTRT gene by in situ hybridization techniques--7 cases of normal oral mucosa, 7 cases of hyperplasia lesions, 30 cases of oral precancerous lesions (dysplasia lesions), 8 cases of oral mucosa carcinomas in situ, and 30 cases of oral squamous cell carcinomas. RESULTS: Weaker signals of hTR and hTRT gene expression were observed in normal oral epithelia and hyperplasia lesions; hTR and hTRT were positive 28.6% (4 of 14) and 21.4% (3 of 14), respectively. The expression of telomerase genes in the precancerous lesions became stronger due to phenotypic progression and the increasing degree of dysplasia; hTR and hTRT were positive 60.0% (18 of 30) and 46.7% (14 of 30), respectively. Stronger hTR and hTRT expressions were observed in squamous cell carcinoma, with an equal positivity of 81.6% (31 of 38). CONCLUSION: Telomerase gene expression is closely related to the malignant degree of oral mucosa. Telomerase is reactivated frequently during late stages of oral precancerous lesions and may play a crucial role in progression of oral cancer. PMID- 10863407 TI - Altered placental glutathione S-transferase foci as a tumor marker during rat lingual carcinogenesis. AB - OBJECTIVE: To evaluate the value of placental glutathione S-transferase (GST-P) as a tumor marker during lingual carcinogenesis in two strains of rats. METHODS: Dark-Agouti (DA) and Wistar/Furth (WF) rats were given with 4-nitroquinoline 1 oxide (4NQO) in drinking water with a defined treatment schedule. Immunocytochemical expression of GST-P was studied using experimental materials obtained at 2, 4, 8, 12, 16, and 26 weeks of 4NQO treatment. GST-P+ foci in the tongue epithelium were counted for each experimental rat and quantitative data were compared between groups. RESULTS: The animal experiment demonstrated marked differences in tumorigenicity between the two strains of rats, with DA exhibiting earlier responses to 4NQO and having a higher tumor incidence than WF rats. GST P+ foci occurred earlier in the tongue mucosa of DA rats than in that of WF. With continued 4NQO application, there was a progressive increase in the number and size of GST-P+ foci in both strains of animals, but those of DA groups revealed a greater increase in comparison to those of WF groups. CONCLUSIONS: These results indicate that GST-P is a sensitive marker that may reflect the metabolic patterns of preneoplastic cells during early stages of rat lingual carcinogenesis, and that rapid expansion of such putative initiated cells may have the propensity for malignant transformation. PMID- 10863408 TI - Mechanical stress regulation and proliferation of rat mandibular condylar chondrocytes in vitro. AB - OBJECTIVE: To study the proliferative changes of cultured rat mandibular condylar chondrocytes effected by mechanical stress. METHOD: Flow cytometry (FCM) was used to explore the changes of cellular DNA content and cell cycles of passaged condylar chondrocytes. RESULTS: FCM results indicated that proliferation of cultured rat mandibular condylar chondrocytes decreased with stress duration, with marked differences between the experimental and control groups. CONCLUSION: Within given force duration and force range (0-207 g/an2), proliferation of rat mandibular condylar chondrocytes increased with force magnitude, reaching the maximum 6 hours after application of stress. Cellular proliferation decreased when the chondrocytes were cultured under continuous stress for 24 hours. The mechanism for these proliferation changes requires further study. PMID- 10863409 TI - Epidemiological study of the risk factors of rampant caries in Shanghai children. AB - OBJECTIVE: To explore the risk factors associated with rampant caries and to provide suggestions for prevention and clinical treatment. METHODS: On the basis of a field survey, case-control study was performed to isolate variables possibly related to rampant caries in children. Data analysis was performed with individual analysis and multiple unconditional logistic regression. Four logistic models were developed. RESULTS: The risk factors of rampant caries were cariogenic foods and bottle-feeding with sugar containing bovine milk; an increased risk of caries was also associated with duration of breast-feeding. CONCLUSION: Overuse of cariogenic foods and unsuitable patterns of infant feeding were the most important risk factors associated with rampant caries. PMID- 10863410 TI - A study of the diagnostic location of pulpitis. AB - OBJECTIVE: To demonstrate the involvement of the root pulp in pulpitis, which may help in selecting a desirable therapy. METHODS: Symptoms of 158 cases of caries caused pulpitis were recorded and histologic diagnosis was made. The location of lesions was determined, and contributive rates and diagnostic indices were developed on the basis of the characteristics of various symptoms. RESULTS: Computer analysis provided the following contributive rates in terms of symptoms: pain upon probing of the tooth (14.72%), duration of spontaneous pain of the diseased tooth (13.07%), intensified pain by cold stimulation (8.25%), and intensified pain by cold test during treatment (4.05%). The exact diagnostic rate was 76% on the basis of the above four contributive rates to demonstrate the involvement of the root of the tooth, while the dentist could only make a correct diagnosis 56.72% of the time. CONCLUSION: The four symptoms with the highest contributive rates improved the diagnostic rate of location, while other symptoms were excluded because of their uncertain contributive rates. As a result, the process of obtaining information was simplified. The tables of diagnostic indices are of value in clinical application as they are easy to learn and understand. PMID- 10863412 TI - Induction of reparative dentin formation in dogs with combined recombinant human bone morphogenetic protein 2 and fibrin sealant. AB - OBJECTIVE: To investigate the reparative dentin formation induced by the complex of recombinant human bone morphogenetic protein 2 (rhBMP2) and fibrin sealant (FS) in dogs. METHODS: Freshly exposed pulp of molars, premolars, and canines were treated, respectively, with the complex of rhBMP2-FS, rhBMP2, and Ca(OH)2 paste, which served as control. RESULT: Wound healing of exposed pulp treated with the complex of rhBMP2 and FS seemed better than that with rhBMP2 alone. Dentin bridge formation was observed at 1 week when pulp was treated with the complex and at 4 weeks with rhBMP2 and Ca(OH)2. At 9 weeks after operation, more amount of tubular dentin bridge formation was found in pulp treated with the complex than with rhBMP2. CONCLUSIONS: The results suggest that synergistic effects of fibrin and rhBMP2 existed and that rh-BMP2 with FS carrier can be used as bioactive pulp capping agent. Together, these agents can induce a large amount of dentin and enhance healing of exposed pulp. PMID- 10863411 TI - Studies on contributing factors in temporomandibular disorders. AB - OBJECTIVE: To better understand the pathogenesis of TMD by studying microtrauma of the temporomandibular joint (TMJ), immune responses within TMJ, and psychosocial factors of the past ten years. METHODS: Condyle and disc movements from 38 patients with temporomandibular disorders (TMD) were observed with the use of videotape recording and soundtape recording techniques after TMJ arthrography. Pathological changes following occlusal trauma were examined using an animal model. Immune complexes in condyle cartilage; antibody to collagen II; and cytokines such as interleukin 1, tumor necrosis factor, and interleukin 6 were detected in synovial fluid of TMD. Psychosocial characteristics were analyzed with the Minnesota Multiphasic Personality Inventory (MMPI) of 80 TMD patients and the Life Events Experience Survey (LEES) of 42 TMD patients. RESULTS: Persistent and recurrent microtrauma did exist within joints of TMD patients, caused by occlusal trauma and occlusal interference. Occlusal trauma in animals could induce condyle and disc degenerative changes that are similar to the findings in TMJ osteoarthrosis patients. Sequestered antigens within cartilage could be exposed to the immune system after joint degeneration. Humoral and cellular immune responses did exist within TMJ and played an important role in the pathogenesis of TMD. Forty percent of TMD patients suffered from psychosomatic disorders, significantly more than in the healthy control. CONCLUSION: Microtrauma of TMJ, immune responses within TMJ, psychosocial factors, and anatomical structures of the TMJ itself are the four main contributing factors of TMD. Possible mechanisms of the interactions of the four factors are presented, and principles of preventing and treating TMD are also suggested. PMID- 10863413 TI - Effects of all-trans retinoic acid and interferon-gamma on expression of RAR beta gene in Tca8113 cells. AB - OBJECTIVE: All-trans-retinoic acid (ATRA) and interferons (IFNs) have been proven to synergistically amplify growth inhibition and apoptosis in the tongue squamous carcinoma cell line (Tca8113). Nuclear retinoic acid receptor-beta (RAR beta) was the key gene that mediated retinoid activity for squamous carcinoma cells. In order to understand the mechanism of ATRA combined with IFN gamma inhibiting growth of Tca8113 cells, this investigation focused on RAR beta mRNA expression. MATERIALS AND METHODS: In this experiment, RT-PCR method was used to analyze the RAR beta expression level, and viable cell count assay was carried out for growth inhibition studies. RESULTS: All-trans-retinoic acid (1 microM) and IFN gamma (1000 u/mL) inhibited cell growth by 39.2% and 44.4%, respectively. Synergistic inhibition of cell proliferation by 86.7% was found under combined treatment. The combination of suboptimal concentrations of ATRA (0.1 microM) with IFN gamma (1000 u/mL) showed a much stronger additive inhibitory effect on cell proliferation. ATRA (1 microM) and IFN gamma (1000 u/mL) increased RAR beta expression to 4 times and 3 times, respectively. The expression of RAR beta increased 12 times after treatment with combined ATRA and IFN gamma treatment. CONCLUSIONS: These observations indicated that the use of combined ATRA and IFN may lead to enhanced antitumor effects. These results also suggested that ATRA and IFN mediated upregulating expression of RAR beta may play an important role in synergistic inhibition of proliferation in Tca8113 cells. PMID- 10863414 TI - Evaluation of the treatment for micromandibular deformity by distraction osteogenesis with submerged intraoral device. AB - OBJECTIVE: To evaluate the results of distraction osteogenesis for correcting the micromandible with an intraoral device. MATERIALS AND METHODS: Eight cases of micromandible resulting from temporomandibular joint (TMJ) ankylosis and first and second branchial arch syndrome received treatment by distraction osteogenesis. Subjects included 5 females and 3 males, ranging in age from 10 to 43 years (avg 20.25). Five micromandibles were associated with obvious mandibular asymmetry. An osteotomy was performed on bilateral mandibular bodies by intraoral approach. Elongation was started on the 5th or 6th day postoperation at a rate of 0.8 to 1 mm per day. The distractors were removed after a consolidation period from 71 to 139 days. RESULTS: The distractions were fulfilled with a range of 8.5 to 24.4 mm. Pain appeared in osteotomy region when the 1 mm distraction was carried out once a day and disappeared when it was divided into twice a day. Mandibular lengthening was successful. Micromandible and facial asymmetries were corrected satisfactorily. The follow-up period ranged from a minimum of 1 month to a maximum of 10 months. There was no recurrence. One case of soft tissue infection and one case of lower lip numbness were reported. There were no cases of infection or other disturbance of wound healing or pseudarthrosis. CONCLUSIONS: Because of its advantage in little injury, avoiding bone grafting, and few complications, intraoral distraction osteogenesis is a valuable approach to correct mandibular agenesis. However, in the management of mandibular deformities, distraction osteogenesis achieves better results when combined with other orthognathic operations. How to control the direction of distraction is a problem that needs further study. PMID- 10863415 TI - Experimental study of sagittal fracture of the mandibular condyle. AB - OBJECTIVE: To observe the healing of sagittal fracture of the mandibular condyle (SFMC) and its effect on the temporomandibular joint in immature miniature pigs. METHODS: Twenty-seven immature miniature pigs were randomly chosen and divided into three groups. Twelve immature miniature pigs underwent sagittal fracture of the mandible condyle; 12 underwent transverse fracture, and 3 were maintained as a normal control. Three-phase scintigraphy, computed tomography (CT) imaging, and histological tests were conducted at 3 weeks, 6 weeks, 12 weeks, and 24 weeks. RESULTS: The data showed the value of the time-activity curve of blood-flow image increased and blood-flow arriving time was postponed after condylar fracture. The alpha/beta ratio was the highest at 2 weeks after SFMC. The count of blood-pool image and static image were higher than nonfractured sides. Peaks were seen at 2 weeks postfracture. Disc and condyle adhesion was found in all sagittal fractures of the mandibular condyle and bifid mandibular condyle was formed. Adhesive tissues between condyle and disc were fibro-connective tissues, which connected the disc directly to cartilage or bone tissue. A large number of fibroblasts and sparse chondrocytes were seen in the adhesive tissue. Blood vessels and adipose cells were seen in the disc, whereas there were no distinguishable changes in transverse fracture. CONCLUSION: Bifid mandibular condyle was found in most SFMC in childhood. The mechanism of bifid condyle deformity is related to secondary injuries to the disc, adhesions between the condyle and disc, and displacement of the medial split fragment. The age when SFMC occurs is an important factor in bifid condyle formation. PMID- 10863416 TI - The influence of exogenous nerve growth factor on inferior alveolar nerve regeneration in silicone tubes. AB - OBJECTIVE: To explore the effects of exogenous nerve growth factor (NGF) in the regeneration of adult white rabbit inferior alveolar nerves within silicone tubes. METHODS: Twenty-four adult white rabbits were selected. Bilateral 8 mm inferior alveolar nerve gaps were created, and the proximal and distal stumps were inserted into a 12 mm silicone tube. The right side silicone tube was filled with exogenous NGF (experimental group) and the contralateral side was filled with saline (control group). Regeneration of the nerves was assessed by histological and nerve eletrophysiological observation. Total number of the regenerated myelinated fibers, conduction velocity of the nerves, thickness of myelin sheaths, and cross-section area of myelinated nerve fibers between the experimental and control groups were compared. RESULTS: The results show that NGF significantly increased the number of myelinated fibers. The experimental group demonstrated more myelinated fibers than the controls at 12 and 18 weeks. At the same time following surgery, the myelin sheath thickness and the cross-section area of myelinated fibers in the experimental group were significantly greater than those in the controls. At 12 and 18 weeks, the conduction velocity of regenerated nerves in the experimental group were greater than those of the control. CONCLUSION: This study suggests that exogenous NGF can enhance regeneration of the inferior alveolar nerves and recovery of their sensory function. PMID- 10863417 TI - Interaction of tumor-infiltrating lymphocyte from oral squamous cell carcinoma with FN enhances its adhesion and cytotoxicity. AB - OBJECTIVE: To investigate the influence of fibronectin (FN) on cytotoxicity and adhesion of tumor-infiltrating lymphocytes (TIL) from oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: TIL cells were isolated from surgical specimens of 9 patients with OSCC. Specific cytotoxicity for autologous tumor cells were tested by 3H-TdR 4 hours release, and specific TNF-alpha release was detected using enzyme-linked immunosorbent assay. Expression of intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-1 (LFA-1) on TIL cells were analyzed on FACScan following incubation with FITC-conjugated antibody. Cell adhesion ability was evaluated through detection by optical density (OD) values of bound cells and unbound cells. RESULTS: TIL-specific cytotoxic activity for autologous tumor was significantly enhanced, ranging from 17.7% to 67.6% by incubation on FN (P < 0.005). Cell adhesion was increased ranging from 42.5% to 62.7% by coculturing with FN (P < 0.005). The interaction of TIL with FN increased expression of ICAM-1 and LFA-1 on TIL cells as well. But specific TNF-alpha release was not obviously changed by FN (P > 0.5). CONCLUSION: The enhanced TIL activity by FN might be mediated by increased binding of TIL to targets through adhesion strengthening. This culture system would be useful to improve the TIL generation for cancer therapy. PMID- 10863418 TI - Standard growth curve chart for body height of Chinese adolescent for orthodontic diagnosis. AB - OBJECTIVE: To make a standard growth curve chart on the body height of Chinese adolescents. METHODS: Statistical data from 32,524 Sichuan students aged 7 to 19 years were surveyed in 1995. Mean values of the annual increment of body height and the ages of the pubertal growth periods were analyzed. RESULTS: During the period between 9 and 13 years of age, females grow faster than their male peers. After age 13 years, males surpass females, and the difference in physical growth and development becomes more pronounced. CONCLUSIONS: The new standard growth curve chart on the body height of Chinese adolescent may be applied to better understand the growth pattern of Chinese orthodontic patients. PMID- 10863419 TI - Orthognathic surgery correction of mandibular hypoplasia accompanying obstructive sleep apnea syndrome. AB - OBJECTIVE: To examine the surgical correction methods for treating cases of severe mandibular hypoplasia accompanying obstructive sleep apnea syndrome (OSAS). METHODS: Sixteen cases of severe mandibular hypoplasia were studied in which OSAS was documented by polysomnography (PSG) and cephalometric study. The obstructive site was at the base of the tongue. Surgical procedures such as temporomandibular joint (TMJ) reconstruction and bimaxillary, chin, and hyoid bone advancement were performed to improve each patient's profile, function, and occlusion, and to treat the OSAS. RESULTS: There were great improvements in patient's sleep and daytime quality of life. The pre- and postoperative changes of most PSG values and some cephalometric values (SNB, PAS) were statistically significant. CONCLUSIONS: Severe mandibular hypoplasia can cause not only abnormalities in profile and occlusion but also OSAS. The evaluation of OSAS and its treatment effects depend on PSG. It is also very important to confirm the obstructive site in the upper airway by cephalometric study and fiberoptic endoscopy. Orthognathic surgery procedures can advance the maxillary, chin, and hyoid bone, and expand the upper airway simultaneously. These procedures can treat OSAS. Cases of TMJ ankylosis with OSAS should be treated step by step. PMID- 10863420 TI - Detection of neuroendocrine feature about salivary adenoid cystic carcinoma. AB - OBJECTIVE: To study the possibility of neuroendocrine feature of salivary adenoid cystic carcinoma (SACC). METHODS: Immunohistochemical staining was operated in SACC with S-100 protein, neurospecific enolase (NSE), chromogranin A, and synaptophysin, which are specific and correlative with neuroendocrine cells and neoplasias. RESULTS: The positive expression of S-100, NSE, chromogranin, and synaptophysin were 42, 32, 8, and 7, respectively, in 50 cases of SACC. CONCLUSION: Neuroendocrine feature in some SACC cells is possibly indicated. PMID- 10863421 TI - An experimental investigation of osseointegration and stability of implants used as orthodontic anchorage in dogs. AB - OBJECTIVE: To investigate osseointegration and stability of three kinds of implants used as orthodontic anchorage in dogs. METHODS: HA-coated, titanium plasma-coated, and uncoated titanium implants were inserted into each femur of two dogs. After a healing period of three months, orthodontic force of 200 g was applied by means of Ni-Ti springs which were connected to the two adjacent implants for two months. Position change of the implant was first measured and then calculated. The shear bond strength of the interface between implant and bone was measured with a push test. After the test, the fracture surface at the interface was observed with a scanning electronic microscope. RESULTS: All implants were stable, without mobility. The highest bond strength and mature bone compactness were found at the interface between HA-coated implant and bone. The other two showed no significant difference in bond strength. However, osseointegration existed at the interface between all three kinds of implants and bone. CONCLUSION: HA-coated, titanium plasma-coated, and uncoated titanium implants can each serve as orthodontic anchorage as well as prosthodontic abutment. PMID- 10863422 TI - Clinical investigation of a high-strength glass ionomer restorative used with the ART technique in Wuhan, China: one-year results. AB - OBJECTIVE: To evaluate the safety and effectiveness of a new glass ionomer restorative, ChemFlex, and to compare its clinical performance and wear to another popular material, Fuji IX GP, when used with the atraumatic restorative treatment (ART) approach in posterior teeth in school children. METHODS: Ninety two subjects aged between 6 and 14 years who had bilateral matched pairs of carious posterior teeth were selected. A split-mouth experimental design was used in which the two restorative materials were randomly placed on contralateral sides. The restorations were assessed directly and also indirectly from color transparencies and die replicas. RESULTS: After one year, the success rates of ART restorations in the primary teeth were 96.6% for ChemFlex restorations and 89.7% for Fuji IX GP restorations placed in the Class I cavity preparations, whereas only 46.2% (ChemFlex) and 61.5% (Fuji IX GP) of Class II restorations were assessed as clinically satisfactory. In the permanent dentition, the success rates were 94.6% and 98.2% for ChemFlex and Fuji IX GP, respectively; however, there was no statistically significant difference between the two restoratives for either the permanent or primary teeth. The mean occlusal wear after one year in the permanent teeth was 53.2 microns for ChemFlex and 56.3 microns for Fuji IX GP. Again, there were no statistically significant differences in wear between the two materials (P > 0.05). CONCLUSION: The clinical performance of both ChemFlex and Fuji IX GP over a 12-month period was highly satisfactory and completely adequate for the ART technique, particularly in Class I cavities. PMID- 10863423 TI - Effects of fluoride toothpaste on etched enamel of orthodontic patients. AB - OBJECTIVE: To investigate the effects of fluoride toothpaste on acid-etched enamel by means of a scanning electron microscope. METHODS: Permanent first premolars extracted for orthodontic reasons were etched with 37% H3PO4. They were divided into two groups in pairs of contralateral teeth. Group 1 was treated with fluoride toothpaste slurry; Group 2 was treated with deionized water. Treatment time was three weeks (one hour, 3 times a day). RESULTS: At the conclusion of the experiment, large amounts of precipitate were observed on the enamel surface of the teeth that were treated with fluoride toothpaste slurry. The etched structures were no longer evident and the enamel surface was smooth. The acid resistance of the teeth was significantly enhanced. However, clearly etched enamel structures were visible on the enamel surface of the teeth that were treated with deionized water. CONCLUSION: Fluoride toothpaste could promote the remineralization of etched enamel and enhance its resistance to acid attack. PMID- 10863424 TI - The orthodontic treatment need and demand of Hong Kong Chinese children. AB - OBJECTIVE: To investigate the need and demand for orthodontic treatment of Chinese school children in Hong Kong. METHODS: The material consisted of study casts and questionnaires collected from 765 randomly selected 12-year-old school children in Hong Kong. The need for orthodontic treatment was assessed by the Index of Orthodontic Treatment Need, and questionnaires were used in assessing demand for orthodontic treatment. RESULTS: The results indicate that 12% of the 12-year-old school children had no need of orthodontic treatment, 18% had little need (Grade 2), 33% moderate need (Grade 3), 33% great need (Grade 4), and 4% of children had very great need for orthodontic treatment (Grade 5). About two thirds of the children were not satisfied with their dental appearance, but only 40% of these would like to have orthodontic treatment. A definite relationship was found between negative self-appraisal of dental appearance and demand for orthodontic treatment. CONCLUSION: The study demonstrates that the need of orthodontic treatment among Chinese children is similar to that of Caucasian children; the attitude and demand toward treatment are also similar. PMID- 10863425 TI - Softening patterns of light cured glass ionomer cements. AB - OBJECTIVE: The aim of this study was to determine the effects of commonly used food simulating solutions and sodium hydroxide on the softening of light cured glass ionomer cements. METHODS: Four types of light cured glass ionomers (classified on the basis of the liquid component) as follows: (1) materials that combine a polymerizable monomer and polyalkenoic acid (PMPA); (2) use of a polymerizable polyalkenoic acid (PPA); (3) acid monomer (AM) in place of the polyalkenoic acid; and (4) replacement of polyalkenoic acid with polymerizable monomer (PMPR). A traditional glass ionomer and a microfil composite were used as controls. Disc-shaped specimens aged for a week at 37 degrees C and 100% relative humidity were stored in water, ethanol, heptane and 0.1 M sodium hydroxide for a period of 28 days. Barcol hardness measurements were made before immersion as well as at intervals of 24 h, 3 days, 7 days and 28 days after immersion. RESULTS: In general the softening effect was lowest on the resin composite control. Hardness could not be measured for the traditional glass ionomer after 24 h due to breakage and dissolution of samples. The different solutions had varying effects on the different classes of light cured glass ionomers. The change in hardness after 28 days ranged from an increase of +6.7% for PMPA material in heptane to a complete disintegration of PPA amd PMPR in NaOH at 60 degrees C. SIGNIFICANCE: The softening effect of food simulating solutions is dependent on the formulation of light cured glass ionomers. In clinical use, the role of softening in wear will consequently vary. PMID- 10863426 TI - Effect of dentin primers on the morphology, molecular composition and collagen conformation of acid-demineralized dentin in situ. AB - OBJECTIVES: The purpose of this study was to assess the effect of two commercial primers and distilled water on the morphology, molecular composition and collagen conformation of acid-demineralized dentin in situ. METHODS: Dentin specimens etched with Scotchbond Etchant were imaged by tapping mode AFM and analyzed by MIR-FTIR spectroscopy. They were then primed with Scotchbond Multi-Purpose Plus Primer, Scotchbond 1 Adhesive or distilled water and imaged and analyzed again. The chemical modifications induced on the uppermost 2 microns of primed dentin were studied after water and original primer subtraction. The conformational changes of type I collagen at this region were evaluated by deconvoluting the amide I band components. The absorbance ratio An(1655/1627) was used to semiquantitatively assess, on a relative basis, the extent of collagen denaturation. RESULTS: All the priming treatments swelled the collapsed dentin collagen left after etching. No evidence of primary bonding was found after priming treatments, while approximately 50% of the conditioned dentin collagen appeared denatured. Treatment with distilled water did not change the status of denatured collagen, however, application of the commercial primers refolded the alpha-helix to approximately 95% of the extent found on the native reference dentin. SIGNIFICANCE: The dynamic response of dentin collagen to demineralization and priming treatments reveals the critical role of some primers in rapidly restoring the conformational status of acid-denatured collagen. Implementation of reactive adhesive groups in alpha-helix recovery may provide an associative means of modifying the mechanical properties of the demineralized collagen based on the extent of their intermolecular bonding. PMID- 10863427 TI - Galvanic interaction between titanium and gallium alloy or dental amalgam. AB - OBJECTIVES: The objective was to examine in vitro the galvanic interaction between titanium and a high-copper dental amalgam or a gallium-based direct filling alloy at different area ratios, and to relate the observed interactions to the electrochemical characteristics of the alloys. METHODS: The tested materials were cast titanium, a single-composition, spherical high-copper amalgam, and a gallium-based direct filling alloy. Polarization curves were recorded. The galvanic couples were prepared at Ti/filling alloy ratios of 1:1, 2:1, 4:1 and 6:1. The couples were exposed to synthetic saliva at 37 degrees C and the galvanic currents and potentials were measured as a function of time. The results were analyzed by ANOVA and Tukey tests (p < or = 0.05). RESULTS: Freshly abraded titanium initially was anodic to both the amalgam and the gallium alloy, but the polarity reversed after a period of exposure. The galvanic potential and current density increased with the increasing Ti/alloy area ratio. The potential increase was smaller and the current increase larger for the Ga alloy than for the amalgam. The difference was consistent with the polarization characteristics. The galvanic current density was of the order of 10(-8) A/cm2 for the Ti/amalgam couple, and 10(-7) to 10(-6) A/cm2 for the Ti/gallium alloy couple. SIGNIFICANCE: The results show that the galvanic interaction between titanium and direct filling alloys is small. High copper dental amalgams should suffer little galvanic corrosion when in contact with Ti. For gallium direct filling alloys, the galvanic interaction may be more detrimental because of the inherently lower corrosion resistance. PMID- 10863428 TI - Effect of removal of surface collagen fibrils on resin-dentin bonding. AB - OBJECTIVES: The aim of this study was to evaluate the effects of NaOCl at removing the demineralized layer by examining the morphology of the hybrid layer and measuring shear bond strengths after different dentin treatments. METHODS: Dentin disks were treated with: (1) 35% phosphoric acid (PA) 20 s; (2) PA treatment followed by 1.5% NaOCl, 2 min; (3) PA treatment, followed by a 10% NaOCl immersion for 120 h. SEM was used to analyze the morphology of dentin and its interface with dentin bonding agents (DBAs), while shear bond strength tests were used to measure adhesion. All specimens were then fractured into two halves: One half was inspected under SEM; the other half was sequentially placed in 10% PA followed by 12.5% NaOCl for 70 h, to remove all dentin from the resin replica of the original bonded interface. RESULTS: SEM observations showed that collagen fibrils were completely removed from the acid-etched surface by NaOCl treatment. The diameter and the size of dentinal tubules and the number of lateral branches of the tubules were increased following NaOCl treatment. NaOCl applied on dentin smear layers did not significantly modify their SEM morphology. Resin tags were larger in diameter after phosphoric acid/NaOCl treatment than after only phosphoric acid treatment. Resin-infiltrated dentin-layers were only observed after the single phosphoric acid (i.e. conventional etching) procedure, and were not observed after combined phosphoric acid/NaOCl treatment. Etched/NaOCl samples showed a lower bond strength using Scotchbond MP and 3M Single Bond, but were higher in Optibond FL and unmodified in Prime & Bond 2.0 groups when compared with acid-etched controls. Treatment of etched dentin with NaOCl for 120 h produced an unusual type of resin infiltration of mineralized dentin that could be called a "reverse hybrid layer" which may explain the mechanism of resin bonding to NaOCl treated dentin. SIGNIFICANCE: The use of acidic conditioners for exposure of the collagen matrix exposes a soft delicate mesh that can collapse, thereby interfering with resin infiltration. If acid-etching is followed by NaOCl treatment, high bond strengths can be achieved via "reverse hybrid layer" formation, a proposed new mechanism of micromechanical resin retention. This mechanism is not yet recommended for clinical use but demonstrates a new type of resin retention. PMID- 10863429 TI - Effect of MMA-PMMA resin polymerization initiators on the bond strengths of adhesive primers for noble metal. AB - OBJECTIVES: This study was conducted to investigate the effect of MMA-PMMA resin polymerization initiators on the bond strengths of two adhesive metal primers by evaluating the shear bond strengths of resins of silver-palladium-copper-gold (Ag Pd-Cu-Au) alloy. METHODS: Three types of MMA-PMMA resins for which the polymerization initiators were TBB, BPO-amine and CQ-amine, and two adhesive primers, Metal PrimerII and V-Primer, were used. A brass ring placed over the nonprimed or primed casting alloy disk surface was filled with each resin. The half specimens were stored in water at 37 degrees C for 24 h. In addition, another half specimens were then immersed alternately in water baths at 4 degrees C and 60 degrees C for 1 min each for 20,000 thermal cycles before shear mode testing at a crosshead speed of 0.5 mm/min. RESULTS: There were no significant differences (p > 0.05) in bond strength between the three types of resins with or without thermal cycling when Metal PrimerII was used. However, when Ag-Pd-Cu-Au alloy was primed with V-Primer, the bond strength of CQ-amine resin was significantly weaker than that of TBB resin. Metal PrimerII was more effective for enhancing the bond strength and the bond strength was not affected by thermal cycling, in contrast to V-Primer. SIGNIFICANCE: The effectiveness of Metal PrimerII to enhance the bond strength is not influenced by polymerization mode of MMA-PMMA resin, in contrast to V-Primer when the resin is bonded to Ag-Pd-Cu-Au alloy. PMID- 10863430 TI - Properties of fluoride-releasing light-activated resin cement. AB - OBJECTIVES: Fluoroaluminosilicate glass and poly(acrylic acid) in small water phase was incorporated into resin composite cements to reduce or prevent secondary caries around luted restorations. The purpose of this study was to investigate the physical properties and the amounts of fluoride released from four types of visible light-activated resin cements. METHODS: A powder was composed of silanized SiO2 filler and 4.5 wt.% (G-4.5), 9.5 wt.% (G-9.5) or 19.5 wt.% (G-19.5) fluoroaluminosilicate glass and 0.5 wt.% reducing agent. The 45.25 wt.% triethylene glycol dimethacrylate (TEGDMA), 45.25 wt.% 2-hydroxyethyl methacrylate (HEMA), 9 wt.% poly(acrylic acid) solution in water and 0.5 wt.% camphorquinone were contained in a liquid. The powder and liquid were mixed at a 3:1 ratio by weight and the three types of specimen disks were cured using a photo-curing lamp. The TEGDMA-based composite cement was used as control (G-0). The specimens were immersed in water at 37 degrees C for 24 h or 6 months and compressive strength (CS) and diametral tensile strength (DTS) were evaluated for the four experimental materials. Disks prepared from the four resin cements were also immersed in deionized distilled water at 37 degrees C, and the fluoride released was measured over a period of 24 weeks. RESULTS: G-9.5 specimens showed almost the same CS and DTS as G-4.5 and G-0 specimens and significantly higher values for both parameters than G-19.5 specimens after 6 months of water storage. The amount of fluoride released from the three fluoride-containing resin cements continued to increase over the entire experimental period. The fluoride release rates of G-9.5 and G-19.5 resin cements were almost the same and were significantly greater than that of the G-4.5 resin cement. SIGNIFICANCE: G-9.5 containing 9.5 wt.% fluoroaluminosilicate glass may be a clinically useful resin cement due to its favorable physical properties and fluoride release. PMID- 10863431 TI - Fracture toughness (KIc) of a dental porcelain determined by fractographic analysis. AB - OBJECTIVES: Fractographic analysis of indentation cracks is performed following flexure testing as part of the ASTM (1999) standard for fracture toughness, KIc, determination in advanced ceramics. This method depends on the conduciveness of the material towards fractographic interpretation. The purpose of this study was to evaluate the use of fractography in fracture toughness methods with a feldspathic dental porcelain, in which KIc was measured fractographically as well as numerically using two controlled-flaw beam bending techniques. METHODS: The following methods for KIc determination were applied to a dental porcelain containing a leucite volume fraction of 15-20%: (1) surface crack in flexure (SCF) (dependent upon fractographic analysis); and (2) indentation strength (IS) at indentation loads of 9.8 and 19.6 N (applying both standard numeric calculations and fractographic analysis). The testing environments were (1) ambient air (IS and SCF) and (2) flowing dry nitrogen (SCF). RESULTS: No significant differences were found between numeric and fractographic KIc values for the IS technique at both indentation loads (9.8 and 19.6 N) in ambient air, although KIc values were sensitive to indentation load. Due to the presence of residual stresses, stable crack extension was observed fractographically in all IS specimens, as evidenced by differences between initial (ainitial) and critical (acritical) crack dimensions. For the SCF method, there was a significant difference in toughness between specimens tested in air versus dry nitrogen, however no fractographic evidence for chemically assisted slow crack growth (SCG) was observed. SIGNIFICANCE: The SCF method as described by the ASTM standard was applicable to the feldspathic porcelain and produced very comparable results with the numeric toughness calculations of the IS procedure. However, fractographic analysis of the surface crack was somewhat difficult for this glassy ceramic compared with polycrystalline ceramics. Knowledge about stable crack extension or slow crack growth and its fractographic appearance is essential when estimating the toughness from examination of flaw dimensions on fractured surfaces since large calculation errors may occur if these effects are not taken into account. PMID- 10863432 TI - The ultrastructure of a compomer adhesive interface in enamel and dentin, and its marginal adaptation under dentinal fluid as compared to that of a composite. AB - OBJECTIVES: To visualise the ultrastructure of the interface of SCA compomer adhesive and of Optibond composite adhesive in enamel and dentin, and to relate the findings to the marginal adaptation of these two products in mixed class V restorations. METHODS: The ultrastructure was investigated using a scanning electron microscope (SEM) with and without prior argon ion etching, an environmental SEM, a field emission SEM, a confocal laser scanning microscope, and a transmission electron microscope. The marginal adaptation was quantified in mixed class V restorations by using the replica technique and a SEM under simulated dentinal fluid before and after simultaneous mechanical and thermal loading. RESULTS: The ultrastructure of the compomer adhesive interface differed from those of the composite. However, no significant difference was discerned as regards the percentage of "continuous margin" in the enamel marginal area before loading, and in the dentin area before and after loading (p < 0.05; unpaired t test). Only after loading, the percentage of "continuous margin" in enamel was significantly (p < 0.05; unpaired t-test) better than that of the compomer. SIGNIFICANCE: The results indicated that the ultrastructure of the adhesive interface allowed no clear conclusions to be drawn as to the quality of marginal adaptation. PMID- 10863433 TI - Fracture resistance of teeth restored with dentin-bonded crowns constructed in a leucite-reinforced ceramic. AB - OBJECTIVES: Laboratory studies and preliminary clinical data have demonstrated satisfactory fracture resistance of all-ceramic crowns placed using a resin composite luting material and a dentin-bonding system. This study investigates the fracture resistance of crowns constructed in a leucite-reinforced ceramic when placed using a dentin-bonding system and a dual-cure resin composite luting material and compares this with the fracture resistance of feldspathic porcelain crowns. METHOD: Standardised preparations were carried out on 10 sound, unrestored, maxillary premolar teeth, the mean bucco-palatal width of which did not vary by more than 2.5%. Ceramic crowns (Fortress; Chameleon Dental, KS, US) were constructed using a standardised technique. Their fitting surface was etched with hydrofluoric acid. The crowns were placed using the dentin-bonding system Mirage ABC and the luting system Mirage FLC (Chameleon Dental, KS, US). The restored teeth were loaded in compression at 1 mm/min through a 4 mm steel bar placed along the midline fissure. RESULTS: A mean fracture load of 0.88 kN was recorded. Results from previous work indicate a fracture resistance of 0.77 kN for feldspathic porcelain crowns placed using the same luting systems on similarly standardised preparations. Statistical analysis by one-way ANOVA and Tukey's multiple comparison procedure indicated that there was no significant difference in the mean fracture resistance of the teeth restored using the leucite reinforced ceramic and the teeth restored with feldspathic porcelain. SIGNIFICANCE: Crowns constructed in a leucite-reinforced ceramic and placed using dentin-bonding and dual-cure resin composite luting materials may provide some increase in fracture resistance, but the results may not be significantly different from the feldspathic porcelain. PMID- 10863435 TI - Investigation of two mercury vapor collection techniques. AB - OBJECTIVES: The purpose of this investigation was to develop and test two in vitro mercury vapor collection techniques: a closed bottle technique (CB) and an intraoral flow (IOF) technique. METHODS: Amalgam samples were prepared in acrylic first molars (#30) with standardized Class I preparations. In the CB technique, samples were placed in either a 25, 100 or 500 ml bottle (n = 5). Vapor was analyzed with the Jerome M-411 using a syringe method over a 7 day period. In the IOF technique an impression of the lower right quadrant of a Typodont was taken with PVS impression material leaving a 5 mm space over #30. Samples were analyzed with the Jerome M-411 connected to the impression tray via tygon tubing at the buccal surface. Average mercury vapor release rates and standard deviations were calculated for each method. Data were analyzed by two-way ANOVA followed by Tukey HSD pairwise analysis for significant findings (alpha = 0.05). RESULTS: Both techniques indicated mercury vapor release was dependent on volume. The largest bottle, 500 ml, yielded a significantly greater (p < or = 0.00) amount of mercury vapor within the CB systems. In the IOF technique, the addition of air flow over the restoration demonstrated a significant increase (p < or = 0.05) in mercury vapor released compared to the sealed IOF technique. SIGNIFICANCE: A method for mercury vapor analysis was developed for possible intraoral application. The IOF method with direct air flow removes possible saturation effects found in a CB system, while limiting external variables, which may contribute to errors associated with in vivo measurements. PMID- 10863434 TI - In vitro cytotoxicity of solid epoxy-based dental resins and their components. AB - OBJECTIVE: The objective of this study was to evaluate the effect of adding a spiroorthocarbonate (SOC) or a polyol on the cytotoxicity of epoxy-based dental resins. METHODS: Resins contained one of the epoxies: diglycidyl ether Bisphenol A (GY-6004); 3,4-epoxycyclohexanemethyl-3,4-epoxycyclohexane carboxylate (UVR 6105); vinyl cyclohexane dioxide (ERL-4206) or the three-epoxy mixture (Epoxy-M). The SOC was t/t-2,3,8,9-di(tetramethylene)-1,5,7,11-tetraoxaspiro[5.5]undecane (SOC). The polyols were polytetrahydrofuran (p-THF-250) and polycaprolactone triol (TONE-301). The photoinitiator (4-octylphenyl)phenyliodonium hexafluoroantimonate and camphorquinone were used for light curing the resins. Four types of resins (epoxy, SOC/epoxy, polyol/epoxy and SOC/polyol/epoxy) were evaluated for cytotoxicity as solids in the agar diffusion assay and as aqueous extracts in the MTT assay using L929 cells. RESULTS: In agar diffusion analysis, ERL-4206 and UVR-6105 resins were severely cytotoxic (+3), but the addition of SOC changed them to non-cytotoxic (-). Addition of 1-3% SOC changed Epoxy-M from mild (+) to non-cytotoxic. Adding SOC changed GY-6004 from moderate (+2) to mild (-) cytotoxicity. Generally, addition of SOC did not change cytotoxicity when added to polyol/epoxy combinations. Either polyol produced resins with reduced cytotoxicity when added to UVR-6105, but the opposite occurred when added to Epoxy-M resins. In MTT analysis, percent cell survival from 100 microliters resin extracts were statistically compared (ANOVA, p < 0.05). Epoxy-M and GY-6004 resin extracts were significantly less cytotoxic than UVR-6105 and ERL-4206 resin extracts were. Overall, the SOC component reduced the cytotoxicity of all SOC/epoxy combinations, except SOC/ERL-4206, which was significantly more cytotoxic than ERL-4206 resin extract. This may be the result of cell fixative effects observed for SOC/ERL-4206 in agar diffusion analysis. Addition of SOC produced significantly less cytotoxic SOC/polyol/Epoxy-M resins when compared to its non-SOC counterpart. The contrary result was obtained with SOC/polyol/UVR 6105 resin combinations. Consistent with agar diffusion results, adding polyol significantly decreased cytotoxicity of UVR-6105 resins. The cytotoxicity of these resins may be related to the 50% cytotoxicity (TC50) of their components as leachates. The TC50 values of the individual components were compared to BISGMA. Polyols, epoxy monomers, SOC monomer and camphorquinone were significantly (p < 0.05) less cytotoxic than BISGMA. SIGNIFICANCE: Addition of SOCs and polyols in the formulation of epoxy-based resins may contribute to development of biocompatible dental composites. PMID- 10863436 TI - Effects of palladium addition on emission of mercury vapor from dental amalgam. AB - OBJECTIVES: The purpose of this investigation was to test the hypothesis that palladium causes a reduction in mercury emission when added to dental amalgam during condensation. METHODS: Mercury vapor release was measured in a closed bottle system and an Intraoral Flow device(IOF). Conventional amalgam restorations were modified by addition of various palladium pellets. 1.57 mm diameter palladium pellets with different porosities were fabricated. These pellets were then placed in amalgam restoration using typical condensation and carving procedures. The samples were stored in a closed bottle and mercury measurements were taken from the bottles at 30 min, 1, 3, 5, 24 and 48 h and 7 days after trituration using a Jerome 411 Mercury Vapor Analyzer (Arizona Instrument Corp., Jerome, AZ). The palladium pellets identified as the most effective in mercury vapor reduction were further tested in an IOF device. Data were analyzed by two-way ANOVA followed by Tukey HSD pairwise analysis for significant findings (alpha = 0.05). RESULTS: The palladium containing amalgams when tested in the closed bottle system yielded significantly lower (p < 0.05) mercury vapor release than the controls. Pellets fabricated with the highest porosity yielded the greatest reduction in overall mercury vapor release. In the IOF device the overall amount of mercury vapor released from the palladium containing amalgams was also significantly less than the control (p < 0.05). SIGNIFICANCE: Mercury vapor emission from dental amalgam was greatly reduced by adding palladium pellets to amalgam during condensation. These techniques require only slight modifications of the standard operative procedures. PMID- 10863438 TI - Evaluation of key parameters in a potentiostatic corrosion test for dental amalgam. AB - OBJECTIVES: Participation in a Round Robin study of potentiostatic corrosion test guidelines for dental amalgam was undertaken for the purpose of developing an accurate set of draft guidelines. METHODS: Dispersalloy, sybraloy, aristalloy, phasealloy, and tytin were used as the amalgam specimens. They were prepared following the guidelines, then coupled to a copper wire, cemented into glass tubes, and polished to a 600-grit finish. A corrosion cell was prepared using a carbon counter-electrode, a standard calomel electrode (SCE) as the reference electrode, and amalgam as the working electrode. A 37 degrees C solution of 10 g/l NaCl with a minimum volume of 300 ml was used. Within 5 min of polishing, the open circuit potential (OCP) was recorded for 10 min. Next, the specimen was polarized to 0 mV versus SCE, and the currents were recorded for a 24-h time period. Corrosion results were analyzed statistically with one-way ANOVA (p < 0.05) and the multiple comparisons Student-Newman-Keuls post-hoc test. RESULTS: Problems that occurred with evaporation, beaker size, carbon electrode length, SCE cap removal, glass tube fracture, polishing technique, and fresh electrolyte are easily avoidable with further explanation or reminder notes. Observations made concerning starting time, initial OCP recording, millivoltage, and solution temperature were determined to be necessary for the accuracy of test results. Analysis of results should include clarification of units, and graph interpretations. Finally, the number of specimens per amalgam should be increased from one to three so that statistical analysis can be performed. Using three specimens per amalgam, the method revealed corrosion susceptibility as measured by the improved test: aristalloy > sybraloy > (dispersalloy, phasealloy, tytin). SIGNIFICANCE: Having run the initially proposed guidelines, a number of clarifying changes were made so that the corrosion susceptibilities of five dental amalgams could be clearly differentiated. PMID- 10863439 TI - Marginal gap formation of light-activated base/liner materials: effect of setting shrinkage and bond strength. AB - OBJECTIVES: The purpose of this study was to compare the effect of setting shrinkage and the shear bond strength on the marginal gap formation in dentin cavities of commercially available light-activated base/liner materials during the early stage of setting, with those of chemical-cured base/liner materials. METHODS: The maximum marginal gap width and the opposing width in the dentin cavity was measured 30 min after the start of mixing. The setting shrinkage and shear bond strength to dentin was measured as the same procedure. To estimate the setting shrinkage, the maximum marginal gap width and the opposing width that occurred with materials placed in a Teflon cavity was measured. RESULTS: The two light-activated base/liner materials produced a significantly wider marginal gap than the chemical-cured base/liner materials (p < 0.05). There were no significant differences in the width of the marginal gap between the remaining five light-activated base/liner and chemical-cured base/liner materials (p > 0.05). The material that produced the smaller marginal gap width in the dentin cavity had a smaller marginal gap width in the Teflon cavity. There was a highly significant correlation between the two findings (r = 0.93, p < 0.01). However, shear bond strength to dentin had no effect on the marginal gap in the dentin cavity (r = 0.04, p > 0.50). SIGNIFICANCE: It appeared that the setting shrinkage of light-activated base/liner materials in the early stage of setting had a greater effect on the marginal gap formation in the dentin cavity than the bond strength to the dentin structure. PMID- 10863437 TI - Elimination, via high-rate laser dilatometry, of structural relaxation during thermal expansion measurement of dental porcelains. AB - OBJECTIVES: Thermal expansion measurement of glassy materials is complicated by thermal history effects. Excess volume--trapped in quenched dental porcelains after firing--collapses via structural relaxation on first slow heating during conventional dilatometry, making the thermal expansion coefficient (alpha) obtained on first heating unreliable. The purpose of this study was to determine whether porcelain thermal expansion measurement at high thermal rates could minimize the influence of thermal history. METHODS: Eight thermal expansion specimens each of six body porcelains and the Component No. 1 (leucite containing) frit prepared according to the patent by Weinstein et al. (US Patent No. 3,052,982) were subjected to three heat-cool conventional dilatometry runs at 3 degrees C/min, while eight thermal expansion specimens of each porcelain were reserved as untreated controls. Eight hollow, cylindrical specimens of the same brands were subjected to three heat-cool laser dilatometer thermal expansion runs at 600 degrees C/min, while eight cylindrical specimens of each porcelain were reserved as untreated controls. Thermal expansion data (25-500 degrees C) of all specimens were subjected to repeated measures analysis of variance. RESULTS: The alpha obtained on first slow heating was significantly lower than values for succeeding slow heat and cool runs in all porcelains (P < 0.001). High-rate alpha obtained on first heating was not significantly different from values of succeeding heat and cool runs in all porcelains (P > 0.05). SIGNIFICANCE: Conventional dilatometer measurements demonstrated occurrence of structural relaxation, as evidenced by the significant difference in the first heating and subsequent runs. High-rate laser dilatometry eliminated structural relaxation, thereby providing a thermal expansion measurement that is free of interference from thermal history effects. PMID- 10863441 TI - Storage of polyacid-modified resin composites ("compomers") in lactic acid solution. AB - OBJECTIVES: The aim of this study was to determine the interaction of four polyacid-modified resin composites with aqueous lactic acid solutions, and to compare changes with those for a glass-ionomer cement and a conventional resin composite. METHODS: For each material, namely Compoglass F, Dyract AP, Hytac and Ana Compomer, plus AquaCem (glass-ionomer cement) and Pekafil (conventional composite resin), five cylindrical specimens of 4 mm diameter x 6 mm height were prepared and weighed. They were stored individually in 2.0 cm3 of 0.02 mol l-1 lactic acid solution for 1 week then the pH was determined and the specimens reweighed. The lactic acid solution was replenished, and the specimens were stored for a further week, after which the pH and specimen weights were again measured. This was repeated at 1 week intervals until the specimens were 6 weeks old. Differences were analysed by ANOVA followed by Newman-Keuls post hoc analysis. RESULTS: All four polyacid-modified composites increased the pH of the solutions at all time intervals by at least 0.26 pH units (significant to at least p < 0.01). This effect was similar to that of the glass-ionomer (but significantly less, p < 0.05) while significantly greater (p < 0.05) than that for the composite, Pekafil, which, by contrast, had no effect on pH. The observed rise in pH reduced significantly over time (ANOVA, p < 0.05). After 1 week, all pH changes were accompanied by net reductions in specimen mass, indicating susceptibility to acid erosion. Hytac was significantly more resistant to this erosion than the other materials; conversely, it had the least effect on solution pH. SIGNIFICANCE: These results show that polyacid-modified resin composites neutralise lactic acid in vitro but suffer erosion in the process. PMID- 10863440 TI - The silver sorption layer in dental composites: three year results. AB - OBJECTIVES: The purpose of this investigation was to observe the behaviour of a recently discovered silver sorption layer in seven dental composites for three years. METHODS: Rectangular block testpieces (3.0 x 2.5 x 2.5 mm) of seven resin composites were fabricated and the resin rich layer removed from one surface by grinding on silica carbide paper. The testpieces were immersed in aqueous AgNO3 (3 mol/l). After 26, 42, 90, 180, 360, 540, 720 and 1085 days, respectively, nine specimens of each material were removed to measure the depth of silver stain in the different composites. RESULTS: The depth of silver stain continued to increase at a rate proportional to (time)0.5. After three years, five homogeneous subsets [HS] were distinguished for the resin rich surface [HS1] Occlusin (stain depth = 45.6 microns); [HS2] Clearfil (117.8 microns), [HS3] Heliomolar (145.6 microns), Concise (148.8 microns), P-30 (168.9 microns); [HS4] Silux (243.3 microns); [HS5] Profile-TLC (446.7 microns). For the ground surface, the materials were in similar subsets but the depth of stain was less. Different coloured layers were seen within the sorption layer in some materials. SIGNIFICANCE: The linear relationship between the depth of stain and (time)0.5 indicate that the mechanism controlling the sorption is Case 1 (Fickian) diffusion. The different depths in the individual materials may indicate differences in the segmental mobility of the polymer chains and free space within the resin phase of the composites. If the silver sorption layer marks the extent of water penetration, then the results show different depth distributions for individual composites. PMID- 10863442 TI - The fatigue strength of the interface between Ag-Pd alloy and hydrothermal ceramic. AB - OBJECTIVE: The purpose of this paper was to investigate the fatigue strength of a hydrothermal ceramic fused to Ag-Pd alloy. This study compared the values with those of other metal-ceramic systems previously reported by other authors. METHODS: This investigation was performed on 48 specimens made of Ag-Pd alloy frames (45 x 12 x 4 mm) on which the hydrothermal ceramic was fused (25 x 12 x 2 gmm). The specimens were divided into two groups. The first group of specimens was dynamically loaded immediately after casting and fusing, while the other group of specimens was thermocycled (1000 times at 0 degree C and 55 degrees C) before dynamic loading. The dynamic loading tests were carried out by using a modified three-point load method in a universal testing machine. RESULTS: The determined dynamic loading limit was up to 550 N for thermocycled, and up to 850 N for non-thermocycled specimens. These results are very respectable in comparison with other metal-ceramics. A statistically significant difference between maximal dynamic forces and number of loading cycles for thermocycled and non-thermocycled samples was determined. SIGNIFICANCE: From the data obtained in this study and current literature profiles, it is concluded that the tested metal ceramic system is more durable than other metal-ceramic systems. PMID- 10863444 TI - The effect of surface grinding and sandblasting on flexural strength and reliability of Y-TZP zirconia ceramic. AB - OBJECTIVES: This study was conducted to evaluate the effect of grinding and sandblasting on the microstructure, biaxial flexural strength and reliability of two yttria stabilized tetragonal zirconia (Y-TZP) ceramics. METHODS: Two Y-TZP powders were used to produce fine grained and coarse grained microstructures. Sixty discs from each material were randomly divided into six groups of ten. For each group, a different surface treatment was applied: dry grinding, wet grinding, sandblasting, dry grinding + sandblasting, sandblasting + dry grinding and a control group. Biaxial flexural strength was determined and data were analyzed using one-way ANOVA, followed by Tukey's HSD test (p < 0.05). In addition, Weibull statistics was used to analyze the variability of flexural strength. The relative amount of transformed monoclinic zirconia, corresponding transformed zone depth (TZD) and the mean critical defect size Ccr were calculated. RESULTS: There was no difference in mean strength between the as sintered fine and coarse grained Y-TZP. Significant differences (p < 0.05) were found between the control group and ground fine grained material for both wet and dry grinding. Sandblasting significantly increased the strength in fine and coarse grained materials. All surface treatment procedures reduced the Weibull modulus of Y-TZP. For both materials, the highest amount of the monoclinic phase and the largest TZD was found after sandblasting. Lower amounts of the monoclinic phase were obtained after both grinding procedures, where the highest mean critical defect size Ccr was also calculated. SIGNIFICANCE: Our results indicate that sandblasting may provide a powerful technique for strengthening Y-TZP in clinical practice. In contrast, grinding may lead to substantial strength degradation and reduced reliability of prefabricated zirconia elements, therefore, sandblasting of ground surfaces is suggested. PMID- 10863443 TI - Metal-matrix interface in reinforced glass ionomers. AB - OBJECTIVE: In reinforced materials, interfacial bonding is critical for efficient transfer of stress from the matrix to the reinforcement. The goal of this study was to characterize the physical and chemical nature of this interface in three metal-reinforced glass ionomers. Two of them were commercial, ESPE-Ketac Silver (KS) and GC-Miracle Mix (MM), and the third, EX, was experimental. METHODS: The techniques of scanning electron microscopy (SEM) and infrared spectroscopy (IR) were used to accomplish the stated goal. SEM analysis utilized polished sections of set cylindrical specimens of each material prepared from their respective powder and liquid components. The glass and the liquid in EX were the same as in MM. The reinforcing agent in EX, 50% by weight, was an Ag-base spherical alloy similar in composition to that in MM. For EX, the alloy was oxidized to promote its bonding to the matrix. The specimens for IR study were prepared as follows. The metallic powders of each material were mixed with the corresponding liquid in excess, stored at 37 degrees C for 1 h, washed with warm water (60 degrees C), filtered and dried. The untreated metallic powders served as controls for IR analysis. RESULTS: SEM revealed a distinct halo shaped internal reaction layer surrounding each alloy particle in EX. A similar layer was not seen in MM and KS. The alloy-matrix interface was continuous and gap-free in EX. In contrast, gaps separating matrix from respective reinforcements were conspicuous features in MM and KS. The IR spectrum of the liquid treated EX alloy powder showed absorbency bands characteristic of unreacted carboxyl groups and carboxylate salts. These bands were absent in the IR spectra of all other powders--treated and untreated. SIGNIFICANCE: The absence of interfacial bonding in MM and KS demonstrated in this study provides a reason why these two materials, in spite of metal addition, have not proved to be any stronger or more durable than their metal-free counterparts. A means of creating interfacial bonding presented here could be useful in the design of improved reinforced glass ionomer materials. PMID- 10863447 TI - A simple method for the measurement of polymerization shrinkage in dental composites. AB - OBJECTIVE: In this study a simple non-contact method was developed to measure the polymerization shrinkage of dental composites. METHODS: A gas pycnometer was used to determine the volumes of specimens prior to and after photopolymerization and from which the total volumetric shrinkage could be determined. RESULTS: Four commercial composites were studied and were found to have polymerization shrinkages varying from 1.6 to 2.5%. The method was found to be labour efficient and produced reproducible results with a standard deviation of approximately 10%. SIGNIFICANCE: This method is appropriate for shrinkage measurements where only the total amount shrinkage is required and in particular for the measurement of shrinkage of photocured materials which are sensitive to water absorption. PMID- 10863446 TI - Consistency in the amount of linear polymerization shrinkage in syringe-type composites. AB - OBJECTIVES: The purpose of this study was to investigate whether the composite resin in a syringe showed a consistent shrinkage through its content. Additionally, the amount of linear shrinkage was compared between materials. METHODS: Five brands of syringe-type and one brand of carpule-type composite resins were used in this study. To each brand, two to three syringes were assigned. In the carpule-type composite, 15 carpules were used. The linear polymerization shrinkage was measured using a custom-made linometer. In this linometer, the amount of displacement of an aluminum disk, which was caused by the linear shrinkage of composite resin, was recorded by a computer every second for 90 s. RESULTS: The syringe-type composites showed similar consistencies in the amount of linear shrinkage except one. The linear shrinkage of the carpule type Tetric Ceram showed more consistency compared with syringe-type composites. The amount of linear polymerization shrinkage varied between materials. SIGNIFICANCE: This investigation demonstrates that the use of carpule-type composites is recommended instead of syringe-types, because of the consistency in its linear shrinkage. The custom-made linometer provides an effective way to study polymerization shrinkage. PMID- 10863445 TI - Cytotoxic effects of current dental adhesive systems on immortalized odontoblast cell line MDPC-23. AB - OBJECTIVES: Evaluate the cytotoxic effect of the three dental adhesive systems. METHODS: The immortalized mouse odontoblast cell line (MDPC-23) was plated (30,000 cell/cm2) in 24 well dishes, allowed to grow for 72 h, and counted under inverted light microscopy. Uncured fresh adhesives were added to culture medium to simulate effects of unset adhesive. Three adhesives systems were applied for 120 min to cells in six wells for each group: Group 1) Single Bond (3M), Group 2) Prime & Bond 2.1 (Dentsply), and Group 3) Syntac Sprint (Vivadent). In the control group, PBS was added to fresh medium. The cell number was counted again and the cell morphology was assessed under SEM. In addition, the adhesive systems were applied to circles of filter paper, light-cured for 20 s, and placed in the bottom of 24 wells (six wells for each experimental materials and control group). MDPC-23 cells were plated (30,000 cell/cm2) in the wells and allowed to incubate for 72 h. The zone of inhibition around the filter papers was measured under inverted light microscopy; cell morphology was evaluated under SEM; and the MTT assay was performed for mitochondrial respiration. RESULTS: The fresh adhesives exhibited more toxic (cytopathic effects) to MDPC-23 cells than polymerized adhesives on filter papers, and as compared to the control group. The cytopathic effect of the adhesive systems occurred in the inhibition zone around the filter papers, which was confirmed by the MTT assay and statistical analysis (ANOVA) combined with Fisher's PLSD test. In the control group, MDPC-23 cells were dense on the plastic substrate and were in contact with the filter paper. In the experimental groups, when acid in the adhesive systems was removed by changing the culture medium, or when the adhesives were light-cured, some cells grew in the wells in spite of the persistent cytotoxic effect. SIGNIFICANCE: All dentin adhesive systems were cytotoxic odontoblast-like cells. Both acidity and non acidic components of these systems were responsible for the high cytopathic effect of those dental materials. PMID- 10863448 TI - Effects of processing parameters on physical properties of the silicone maxillofacial prosthetic materials. AB - OBJECTIVES: Maxillofacial prostheses are commonly fabricated using dental stone molds. However, for evaluating physical properties, maxillofacial materials are most often cured in metal molds. A-2186 is a silicone-based maxillofacial prosthetic material. Because its cure may be inhibited by traces of impurities, its physical properties may be different when it contains additives and is cured in dental stone molds, compared to when it is cured in metal molds without additives. This study's purpose is to determine and compare the physical properties of A-2186 cured in stainless steel molds and stone molds. The effects of additives and cure conditions on the physical properties were also studied. METHODS: Hardness, tensile strength, ultimate elongation, and tear strength of A 2186 cured in dental stone molds, stainless steel molds, and with and without additives were determined. The bonding strength of A-2186 to four adhesives was determined by the peel test. All comparisons were made using analysis of variance (ANOVA). RESULTS: Hardness, tensile strength and ultimate elongation of A-2186 cured in stainless steel molds are significantly higher than those cured in stone molds. Adding a small amount of a pigment, a kaolin and a fiber reduces hardness, tensile strength, ultimate elongation and tear strength. Except for Hydrobond, the bond strength of the adhesives to A-2186 was not significantly affected by the cure conditions and additives. SIGNIFICANCE: Physical properties of A-2186 are affected by the additives commonly used in fabricating maxillofacial prostheses, and use of stone molds for curing degrades A-2186's mechanical properties. In fabricating clinical prostheses, special attention should be exercised to avoid contamination of A-2186 with impurities that could inhibit curing and produce inferior prostheses. PMID- 10863449 TI - Sleep cycles, TMD, fibromyalgia, and their relationship to orofacial myofunctional disorders. AB - Poor quality sleep is caused by many factors including orofacial myology disorders. TMJ and fibromyalgia patients demonstrate a variety of similar symptoms making diagnosis difficult. A team approach utilizing appropriate referrals is critical to successful patient treatment. PMID- 10863450 TI - Adapted or atypical thrusting? AB - Distinguishing between the terms "adapted thrusting" and "atypical thrusting" is not the critical factor in planning treatment for patients demonstrating tongue thrust behavior. Evaluation of factors including age of the patient, breathing pattern, cranio-facial characteristics, head and body posture are important in determining when, and how to provide treatment. PMID- 10863451 TI - The interrelationship of wind instrument technic, orthodontic treatment, and orofacial myology. AB - This article identifies, defines and reviews the synergy between orofacial myofunctional and orthodontic health with regard to wind instrument performance, and summarizes the skills involved in playing an instrument. (i.e. embouchure, articulation, breath support.) Criteria and strategies for choosing an instrument are outlined via orthodontic classifications, therapeutic value or contraindication and team approaches. The author concludes that a team-oriented approach on the part of the professions cited in this article are of the ultimate good for the student/patient. PMID- 10863452 TI - Social environmental factors in Japan affecting the development of proper eating behaviors. AB - This article summarizes issues related to myofunctional disorders/dysphagia and focuses upon social and economic changes within Japanese culture affecting eating habits and behaviors in children. The authors suggest that unfavorable environmental factors negatively impact upon the acquisition of mastication and swallowing behaviors. The article includes discussion of prior research. Studies indicate that decreased observation of early childhood eating habits, dietary changes with regard to higher consumption of fast food and changes within the family, i.e. busy work schedules, decrease in family mealtimes, combine to incur negative change with regard to orofacial function. PMID- 10863453 TI - Myofunctional therapy: brief intervention. AB - This study addresses speech-language therapy in orofacial myology utilizing a Brief Intervention (in Portuguese: IntervenAao Fonoaudiologica Breve) (IFB). IFB is applied to patient groups between the ages of 8 and 15 years with orthodontic/orthopedic appliances in 1997. Results are presented indicating the advantages of using IFB for breathing, feeding, oral-facial habits, buccal hygiene and corporal posture/physical activity. It concludes that Brief Intervention can be accomplished in 8 sessions, is economically advantageous for use in group therapy, and may be used before or in conjunction with Myofunctional Therapy/Myotherapy. PMID- 10863454 TI - How tough is that root canal? AB - It is not uncommon to be faced with difficult treatment planning decisions when a patient presents with an endodontic problem during a routine day in the practice of general dentistry. Signs and symptoms of pulpal or periapical disease dictate the need for decision-making, and frequently the result of that decision is magnified by the patient experiencing severe pain. Our purpose is to present a link between case selection based on risk factors and the self-assessment of the dentist regarding his or her ability or motivation to treat a given case. PMID- 10863455 TI - Enhanced visualization with microscopy and digital radiography. AB - The ability to visualize the three-dimensional anatomy of the root canal system is essential in both surgical and nonsurgical endodontic procedures. Endodontists have adopted two new technologies to enhance visualization: the surgical operating microscope and digital radiographic imaging. This review article discusses the uses and benefits of these technologies and how they have advanced the art and science of modern endodontic treatment. PMID- 10863456 TI - Diagnosis and treatment of cracked teeth. AB - The incidence of cracked teeth appears to be increasing as we keep our teeth longer and the stresses of daily life result in crack-inducing habits, such as clenching and bruxism. This article deals with the clinical characteristics, diagnosis, treatment, and prognosis of cracked teeth. Clinical cases are presented emphasizing the importance of early detection and a unique treatment approach to enhance the successful management of cracked teeth. PMID- 10863457 TI - Managing the endodontic patient with disabling anxiety or phobia. AB - Disabling anxiety and phobia are commonly encountered in dental practice. Successful management of these patients is especially important in endodontic practice, where patients are often compelled to seek treatment because of acute or threatened pain. This article reviews the recognition and management of these common disorders. Mild anxiety can be managed with oral sedatives and/or nitrous oxide; however, moderate to severe anxiety and phobia is best treated with deep sedation or general anesthesia. PMID- 10863458 TI - Soft tissue simulation on casts for implant prostheses. PMID- 10863459 TI - Model prep technology: more than just pinning dies. PMID- 10863460 TI - A secure method for making removable dies. AB - Removable dies greatly facilitate fixed prosthesis fabrication and accurate marginal fit. The major difficulty when using removable dies is the correct positioning of the dowel pin in the stone base. This article describes a method for preparing removable dies that ensures accurate positioning of the dowel pin and provides external landmarks for precise location of the cuts to be made with the die saw. The method described is accurate, simple, inexpensive and can be performed in the dental laboratory. PMID- 10863461 TI - Extreme dentistry: using all-ceramic materials to restore teeth in a police canine with destructive mouth behavior. AB - Due to their aggressive temperament and the nature of their work, dental injuries are common in military and law enforcement canines. From an image and public relations standpoint, "tooth colored" restorations are most desirable, however traditional porcelain fused to metal crowns have performed poorly under these unusual demands. This article will discuss the use of Vita In-Ceram under extreme conditions. PMID- 10863462 TI - Fabricating a gold occlusal platform on a removable partial denture to help prevent extrusion of mandibular incisors. AB - A clinical procedure is presented in which a maxillary removable partial denture replacing anterior teeth was fabricated with a cast gold anterior occlusal platform. This treatment is indicated to correct abrasion caused by extruded mandibular anterior teeth on the maxillary removable partial denture. The advantages compared to prosthetic teeth supported by an acrylic resin base or metal-backed facings are discussed. This technique allows for development of optimal esthetics, strength, and durability while preventing further extrusion and excessive wear of the teeth occluding against prosthesis. PMID- 10863463 TI - Development of the G.E.S. electroforming technique: biocompatible, corrosion-free production of telescopic crowns. PMID- 10863464 TI - "Pressable ceramics" sure-fire techniques. PMID- 10863465 TI - Restoration of posterior implants using a new ceramic material. AB - This article discusses posterior implant restorations made of an advanced, high tech ceramic called zirconium oxide (also called In-Ceram Zirconia). This material, which is second in hardness only to diamond, was originally applied to industrial applications where metal components failed. Zirconium oxide is extremely hard, wear resistant, and chemically inert, and in combination with the In-Ceram technique, the first all-ceramic material recommended for posterior bridges. PMID- 10863466 TI - Microwave denture processing: Dentsply success system. PMID- 10863467 TI - Opals in nature. PMID- 10863468 TI - Splinted all-ceramic laboratory technique for implant restorations. AB - The fabrication of esthetically pleasing implant restorations remains a common and frustrating challenge for dental technicians as well as clinicians. Poor implant placement and post surgical gingival response often creates a difficult foundation on which to build a successful restoration. Castable, cast-to and prepable abutments are employed to correct these problems as they allow some measure of control of angulation and depth of the clinical restoration. This article details the restoration of two implants with ceramic implant abutments and splinted crowns. The technique described achieves a high level of esthetics despite less than ideal conditions. PMID- 10863469 TI - Optimal infection control. Utilizing an autoclave in the dental laboratory. AB - Articles in recent dental journals are strongly advising our dentist customers to require their laboratories to provide them with information regarding in laboratory cross contamination control procedures. New strains of infectious diseases are emerging. Because of over use, some antibiotics are losing their effectiveness. The Center for Disease Control and The World Health Organization assert that emerging infections represent a global threat. Disinfection (use of glutaraldehyde, iodophors, and chlorine compounds) is generally less lethal to pathogenic organisms than sterilization. By using an autoclave in our laboratory we have taken the logical "next step" to protect our dentists and their patients. PMID- 10863470 TI - Effect of insulin on naturally occurring gingivitis rats with diabetes. AB - It is well known that diabetes mellitus aggravates both the severity and progression of periodontal disease. We sought to further explore biologic mechanisms of this relationship using naturally occurring gingivitis rats (ODUS/Odu) rendered diabetic by 65 mg/kg intravenous injection of streptozotocin (STZ). Insulin was administered daily to one group of the rats beginning 4 weeks after STZ injection (STZ-insulin group). Others received no insulin (STZ group). A third group that received no STZ was kept as controls. Eight weeks after STZ injection, sterilized liquid paraffin was injected peritoneally into all three groups, and peritoneal macrophages were collected 4 days later. Macrophage chemotaxis was measured by the membrane filter method using a 48-well microchemotaxis chamber with zymosan activated serum used as a chemotactic stimulant. Blood glucose levels, body weight, plaque indices, pocket depths, serum triglyceride and hemoglobin A1C levels were also determined. We found that blood glucose levels, body weight and triglycerides recovered to normal values in the STZ-insulin group. Further, control of blood glucose resulted in diminished plaque indices, and pocket depths returned to values seen in the controls. Chemotaxis and phagocytosis of peritoneal macrophages improved slightly in the STZ-insulin group, but did not return to levels seen in the pretreatment state. Although insulin resulted in some improvement in leukocyte function damaged by induced diabetes mellitus, recovery was incomplete. PMID- 10863471 TI - Effects of conditioning periaqueductal gray stimulation on responses of thalamic nociceptive neurons to tooth pulp stimulation. AB - Nociceptive neurons receiving afferent input from the tooth pulp (TP) were recorded from the nucleus ventralis posteromedialis proper (VPM) and intralaminar nuclei of the thalamus in cats anesthetized with urethane and chloralose. The effects of stimulating the periaqueductal gray (PAG), or the nucleus raphe dorsalis (NRD) on responses of thalamic nociceptive neurons were investigated. Eight tooth pulp specific (TPS) and 7 wide dynamic range (WDR) neurons with TP input were observed around the periphery (shell region) of the posterior half of VPM. Following electrical stimulation of either the ventral PAG or the NRD, responses to TP stimulation were inhibited in all TPS and WDR neurons tested. Responses of these neurons to electrical stimulation of trigeminothalamic tract (TTT) fibers in the trigeminal medial lemniscus were also inhibited following PAG/NRD stimulation. These results suggest that PAG/NRD stimulation-produced inhibition of both TPS and WDR neurons may be partially mediated by an ascending antinociceptive mechanism. Intralaminar nociceptive neurons with TP input were observed in the nucleus centralis lateralis (CL), and parafascicularis (Pf). The effects of conditioning electrical stimulation of either the ventral PAG or the NRD on responses of intralaminar nociceptive nerons were studied. Of 15 intralaminar nociceptive neurons tested, 6 neurons were inhibited, 5 neurons were excited and 4 were unaffected following the conditioning stimulus. In neurons in which responses to TP stimulation were inhibited, responses elicited by electrical stimulation of the mesencephalic reticular formation (MRF) were also inhibited. These data suggest that although there is an ascending inhibitory pathway from PAG/NRD to intralaminar nuclei, this system is far less potent compared with the ascending inhibitory system acting upon the VPM. PMID- 10863472 TI - Regulated permeability of tight junctions by E-cadherin in rat submandibular gland acini. AB - We examined the relationship between E-cadherin (E-CD), protein kinase C (PKC) and assembly of the tight junction (Tj) in rat submandibular gland acinar cells. The junctional complex of the acinar cells was double-labeled with anti-ZO-1 antibody and anti-E-CD antibody. When Ringer's solution was intraductally injected into the main duct, ZO-1 labels were highly concentrated at the Tj zone, and the adherens junction (Aj) was exclusively labeled by E-CD. In addition, the Tj was impermeable to microperoxidase. Neither intraductal injection of anti-E-CD antibody solution nor infusion of carbachol produced labels for ZO-1 at the Tj zone, although these proteins were occasionally intermixed at the Aj zone. In these cases, the Tj was permeable to microperoxidase. Intraductal injection of anti-E-CD antibody solution with PKC agonist, resulted in a reduction of E-CD labels in the Aj zone, while the ZO-1 was labeled exclusively in the Tj zone. In this case, Tj was impermeable to microperoxidase. These results suggest that E-CD plays a major role in mediating intercellular physical adhesion, and that PKC may be active in signaling the pathway activated by E-CD-mediated cell-cell adhesion. PMID- 10863474 TI - Beta 2-integrin, LFA-1-mediated p125FAK activation. AB - Accumulation of T cells at inflammatory sites is one of the characteristic features of infection, autoimmune and chronic inflammatory diseases. Optimal activation of T cells requires the binding of the MHC/Ag complex with T cell receptor, as well as a secondary signal initiated by costimulatory molecules such as CD2, CD28 or integrins. Focal adhesion kinase, pp125FAK (FAK) has been previously shown to be localized in focal adhesions in fibroblasts and to be involved in integrin-mediated cellular activation. Although signaling through beta 1- or beta 3-integrins induces tyrosine phosphorylation of FAK, there has been no evidence that activation of T cells through the beta 2-integrin, lymphocyte function-associated antigen (LFA)-1, involves FAK. We report here that crosslinking of LFA-1 induces tyrosine phosphorylation of FAK in PHA-activated T cells. Moreover, co-crosslinking with anti-LFA-1 monoclonal antibody (mAb) and suboptimal concentration of anti-CD3 mAb markedly increases tyrosine phosphorylation of FAK in an antibody-concentration and time-dependent manner compared with stimulation through CD3 alone. Furthermore this increased phosphorylation correlates well with the enhanced proliferation of PHA-activated T cells. Results indicate that signals mediated by LFA-1 can regulate FAK, suggesting that LFA-1-mediated T cell costimulation may be involved in T cell activation at least partially through FAK. PMID- 10863473 TI - Coagulation and fibrinolytic activity in rats with naturally occurring gingivitis. AB - It is well known that there are several coagulation factors and fibrinolysis factors, and these inhibitors are also present in blood and tissues. Normally, these factors balance each other. We measured antithrombin III (AT III), an important inhibitor of coagulation and alpha 2-plasmin inhibitor (alpha 2-PI), a factor inhibiting fibrinolysis in blood and gingival tissue of naturally occurring gingivitis rats (ODUS/Odu) and control rats (Res). Blood plasma and supernatants of gingival homogenate from ODUS/Odu and Res were used as samples. And concentrations of alpha 2-PI and AT III were measured by colorimetric assay using commercially available reagents. We found no difference in blood concentrations of alpha 2-PI or AT III between ODUS/Odu and Res groups. However, AT III in the gingiva showed a significantly higher value in the ODUS/Odu group versus that in the controls (p < 0.001). There was a high positive correlation between AT III in gingival tissue and pocket probing depth. Moreover, the fibrinolytic activity in saliva and the gingiva from ODUS/Odu increased with each passing month. Results suggest that both the decrease in blood coagulability and systemic enhancement of fibrinolytic activity due to increased AT III in gingival tissue are associated with the bleeding tendency and inflammatory response in the gingiva of ODUS/Odu. PMID- 10863475 TI - Histochemistry of Ki-67 antigen in ameloblastomas. AB - The proliferative potential of two common histologic variants of ameloblastoma was investigated immunohistochemically using Ki-67 antibody on routinely processed, formalin-fixed, paraffin-embedded sections. Thirty cases of ameloblastomas (15 cases of follicular and 15 cases of plexiform type) were analyzed. Autoclave heating pretreatment was employed at 121 degrees C for 20 min prior to analysis. This retrieval method allowed immunoreactive sites of the Ki 67 antigen to become exposed and thus available for immunohistochemical reaction. We found that expressed Ki-67 antigen was localized within the nucleus of tumor cells in both follicular and plexiform ameloblastomas. Immunoreactive cells were localized in the peripheral area of tumor islands as well as in the central stellate reticulum-area. The labeling rate was higher in the plexiform (3.68%) than in the follicular type (1.78%). Results suggest that cell proliferation of ameloblastoma was different depending on histologic variation of the tumor. Further, the proliferative potential was higher in the plexiform ameloblastoma than that in the follicular type. PMID- 10863477 TI - Functional properties of nociceptive neurons in the nucleus centralis lateralis of the cat thalamus. AB - We studied functional properties of nociceptive neurons in the thalamic intralaminar nuclei, especially the nucleus centralis lateralis (CL) in urethane chloralose anesthetized cats. Nociceptive neurons were found in the intralaminar nuclei, i.e. CL, nucleus centralis medialis (CeM) and nucleus parafascicularis (Pf). One third of these nociceptive neurons had their receptive fields throughout the body, including the orofacial area. Namely, they received convergent nociceptive input from the trigeminal and spinal nerve territories. These neurons with widely complex receptive fields were similar to those we found in the middle third of the caudal bulbar reticular formation (RF). Electrical stimulation of the CL antidromically activated about half of the caudal bulbar RF neurons tested. Furthermore, electrical stimulation of the cingulate gyrus antidromically activated some of CL neurons. These findings suggest that most of the CL neurons receiving nociceptive input from throughout the body receive information directly or indirectly via the caudal bulbar RF, and that certain of this information is then conveyed to the cingulate gyrus. PMID- 10863476 TI - Characteristics of cell-membrane and extracellular proteoglycans in monkey submandibular gland. AB - We investigated the characteristics of cell-membrane and extracellular proteoglycans in the monkey submandibular gland using enzymatic digestion and Western blot analysis. Extracellular proteoglycans and cell-membrane proteoglycans were extracted from the phosphate-buffered saline (PBS) soluble fraction and microsomal fraction of submandibular glands, respectively, and purified partially by ion exchange chromatography. Cellulose acetate membrane electrophoresis and immunoblotting with 3G10 monoclonal antibody or 6B6 monoclonal antibody showed that the extracellular proteoglycans contain several heparan sulfate proteoglycans (117, 67, 47 and 35 kDa core proteins) and a dermatan sulfate proteoglycan (45-50 kDa core protein), while the cell-membrane proteoglycans contain only a heparan sulfate proteoglycan with 69 kDa core protein. These results indicate that the several extracellular heparan sulfate proteoglycans may be released ectodomains of the cell-membrane proteoglycans from cell surface. It is suggested that the cell-membrane and extracellular proteoglycans may regulate the function of growth factors and the microenvironment in the normal submandibular gland. PMID- 10863478 TI - Correlation of E-cadherin and alpha-catenin expression with differentiation of oral squamous cell carcinoma. AB - The precise mechanism of disruption of cell-cell adhesion in oral squamous cell carcinomas has yet to be established. We therefore sought to clearly this mechanism by investigating expression of both E-cadherin (E-CD) and alpha-catenin (alpha-CAT), which is an intracellular CD-binding molecule, in squamous cell carcinomas of the tongue, gingiva and floor of the mouth using immunohistochemical and immunoblotting techniques. We found that reduced expression of both E-CD and alpha-CAT occurred more frequently in moderately- and poorly-differentiated carcinomas than in well-differentiated specimens (p < 0.001), and this reduced expression showed no regional specificity. Relatively frequent loss of alpha-CAT expression in poorly-differentiated carcinomas was detected by both immunohistochemical and immunoblotting analyses. These findings suggest that E-CD and alpha-CAT are both important regulators of intercellular adhesion, and that the reduction of these molecules is also linked to the process of tumor dedifferentiation. PMID- 10863479 TI - Expression of the cell-surface heparan sulfate proteoglycan mRNA in monkey submandibular gland. AB - We investigated the mRNA expression of cell-surface heparan sulfate proteoglycans, syndecans and glypican, in the adult female monkey submandibular gland using the reverse transcription-polymerase chain reaction (RT-PCR) technique. Agarose gel electrophoresis of the PCR products of the cDNA generated from RNA was carried out to demonstrate the expression of mRNA in syndecan-1, syndecan-2, syndecan-4 and glypican in this study. In order to compare the mRNA expression level among the cell-surface heparan sulfate proteoglycans, we measured changes in the relative intensity of PCR products with increasing thermal cycle number. The expression levels were syndecan-4 > syndecan-1, syndecan-2 > glypican. Considering these results together with our previous report, we found that the cell-surface heparan sulfate proteoglycans, syndecan-1, syndecan-2, syndecan-4 and glypican, are synthesized in the monkey submandibular glands, and that their ectodomains are released into the extracellular matrix. It was speculated that control of the expression patterns of the cell-surface proteoglycans may regulate the cellular function and behavior in the submandibular gland. PMID- 10863480 TI - Effects of variation in timing of palatal repair on articulation skills in complete cleft lip and palate cases--a retrospective study. AB - The effects of variation in the timing of palatal repair on articulation skills in complete cleft lip and palate was evaluated from fifty subjects. The present study confirmed that development of articulation was similar in the groups operated upon before 24 months and between 24 to 36 months. The insignificant difference suggests that articulation was good irrespective of the early or medium timing of palatal repair. PMID- 10863481 TI - Leopard syndrome--report of a variant case. AB - This case report presents a patient with Leopard syndrome, with multiple lentigines all over the body and face, ocular hypertelorism, delayed secondary sexual characteristics, mild cardiac abnormalities and supernumerary teeth. Clinical relevance of this syndrome lies in its early recognition and precautions to be taken during any invasive dental procedure, which if not performed under antibiotic prophylaxis and premedication, could lead to infective endocarditis. Additionally, a multidisciplinary approach with pediatric and medical consultants is mandatory during the management of such cases. PMID- 10863482 TI - Assessment and treatment of dental caries in semi-urban school children of Tamilnadu (India). AB - The study was carried out with the purpose of evaluating the prevalence of dental caries, in semi urban school children. The sample comprised of 415 school going children. DMFT/dmft scores were recorded as per WHO 1987 criteria. Mean DMFT was found to be 0.17, 0.06 in male and female children at 3 to 6 age group which increased to 1.21 & 1.10 in males and female in 9 to 12 year age group. Mean dmft at 3 to 6 years was 1.36, 1.17 in male and female children which further increased at 6 to 9 years in both sexes but in the 9 to 12 year age group the value decreased to 1.48, 0.87 in male and female children respectively. The entire sample showed a dental caries prevalence of 58.1. It was noted that the children brushed once a day with toothpaste and toothbrush. Complete oral rehabilitation was undertaken through an incremental school health care programme. PMID- 10863483 TI - Talon cusp associated with other dental anomalies--a case report. AB - Talon cusp is an anomalous structure resembling an eagles talon which projects lingually from the cingulum area of an incisor. It is a rare anomaly which is commonly seen in maxillary incisors. This paper is a report of a case of Talon cusp associated with cross-bite and partial anodontia. PMID- 10863484 TI - Prevalence of dental caries and co-relation with fluorosis in low and high fluoride areas. AB - The aim of the study was to determine the degree of caries prevalence in the permanent dentition and the accompanying fluorosis in children between 6-16 years of age in both low (0.5 ppm) and relatively high (1.2 ppm) fluoride areas. In 3605 children in a low fluoride area (Dharwad), the mean DMFT was 0.65; 77% of the children were caries free. Grade I fluorosis (using Dean's fluorosis inded) was observed in only 0.66% of the children. Among 3618 children of similar age groups, living in high fluoride areas (Gadag), 84% were caries free and the mean DMFT value was 0.39. Varying degrees of fluorosis were present in 57.07% of the children. The results of the study suggest a definite relationship between the amounts of fluoride ingested through water and caries experience observed in the population. PMID- 10863485 TI - Multiple supernumerary teeth in the mixed dentition. AB - Supernumerary or extra teeth result from disturbances during the initiation and proliferation stages of dental development. Teeth formed in excess of the normal number are termed supernumerary teeth. A supernumerary tooth may closely resemble the teeth of the group to which it belongs i.e. molars, premolars or anterior teeth or it may bear little resemblance in size or shape to the teeth with which it is associated. Discussed here are reports of three cases with multiple supernumerary teeth in the mixed dentition and its management. PMID- 10863486 TI - A study of dental caries in school children from rural Haryana. AB - The prevalence of dental caries among rural school children (688 boys and 331 girls) in the age group of 12-16 years in Haryana was found to be 39.4%. It was 37.9% in boys and 42.6% in girls. The difference between males and females was statistically not significant. A significant increase in prevalence of dental caries with age was observed (i.e. 33.1% in 12-year-old children to 45.8% in 14 year-old children). The mean DMFT per child was found to be 1.03 and DMFT per affected child was 2.6. The mean of D, M and F was found to be 1.0, 0.03 and 0.0 respectively. PMID- 10863487 TI - Changes in the arch length following premature loss of deciduous molars. AB - When the normal physiological process of deciduous tooth exfoliation and eruption of its successor is disrupted, a series of changes are observed in the dental arches. The aim of the study was to evaluate the amount of changes in arch length after the premature loss of deciduous molars. The sample consisted of 82 children, 53 without premature loss and 29 with premature loss of either deciduous first or second molar or both, unilaterally. A reduction in arch length was observed both in the maxilla and mandible at the molar region and an increase in arch length at the canine region in the mandible. Reduction in arch length was due to mesial migration of the molar and the increase in arch length was due to the distal migration of canine. It was seen that arch length reduction was more in maxilla as compared to the mandible and that distal drifting of canine was observed only in the mandible. PMID- 10863488 TI - Relationship of tongue-thrust swallowing and anterior open bite with articulation disorders: a clinical study. AB - A Paediatric dentist may be the first person consulted for professional advice concerning children with speech problems. A positive significant relationship has been hypothesized by some authors between tongue-thrust swallowing and articulation disorders in children. This study was undertaken to find out whether any articulation disorders are associated with the habit of tongue-thrust swallowing, or is it the type of anterior bite that plays an important role in the normal or abnormal speech production in tongue-thrust swallowers. Forty subjects with tongue-thrust swallowing in the age group of 7-16 years were examined. Of these 20 had normal anterior bite while rest had anterior open bite. A word articulation test was used and word level articulation testing was done for initial, medial and final positions. The sounds tested were: Linguoalveolars, Labiodentals, Linguodentals, Linguopalatals, Bilabials and Linguovelars. The results of the study have indicated that the presence of articulation disorders is strongly associated with the anterior open bite present in tongue-thrust swallowers but a simple, direct relationship between the presence of defective consonant sounds and tongue-thrust swallowing has not been found. PMID- 10863489 TI - Effective utilization of available infrastructure for oral health promotion in India. AB - The strength of Indian society lies in its social, cultural and religious infrastructure. This feature needs to be utilized for appropriate intervention in life styles of people for effective prevention of dental diseases, all of which are dependent, on life style, to a large extent. PMID- 10863490 TI - Prevalence of dental caries among school children of Moodbidri. AB - This survey was carried out among 2902 children aged between 5 and 12 years attending 13 primary schools in various areas of Moodbidri, in Udupi district. The oral health status was assessed using the simplified WHO Oral Health Assessment Form. The caries prevalence was found to be 76.9%. The mean DMFT was 0.78 and the mean deft was 3.48. Although the mean DMFT score between males and females did not show any significant difference, the mean deft was found to be higher among males compared to females. It was also found that the mean DMFT score increased with age whereas the mean deft score decreased with age. PMID- 10863491 TI - Comparative analysis of tensile bond strength of two new fissure sealants using invasive and non-invasive techniques. AB - In this era of preventive dentistry, a galore of dental materials used for prevention of dental diseases is available. Since the last few decades efforts are being directed towards prevention of dental caries which is one of the major dental diseases tormenting mankind. The rationale of using a fissure sealant is that when it is applied on the caries prone fissures, it penetrates these pits and fissures and seals them from the oral environment. One of the main problems usually encountered is retention of the sealant material. This study was undertaken with the aim to evaluate the tensile bond strength of two new fissure sealants using invasive and non-invasive techniques. The results substantiated the use of invasive techniques over non-invasive techniques and resin sealants over glass ionomer sealants. PMID- 10863492 TI - Fusion of primary incisors--a report of six cases. AB - Six cases of asymptomatic dental twinning anomalies in the primary dentition are reported in 4205 school children. A clinical and radiographic presentation of the cases of fusion of primary incisor teeth is illustrated. This clinical entity has been found to appear with varied clinical and radiographic appearances. An association of fusion of primary incisors with the number of succedaneous teeth was seen. PMID- 10863493 TI - Management of root fracture of mandibular permanent lateral incisor in mixed dentition--a case report. AB - Management of root fracture of mandibular lateral incisors during mixed dentition with developing crowding in a ten year old boy by bilateral surgical extraction. Migration of permanent canine into the extracted region of lateral incisors was favourable with minimal spacing in between the rest of the teeth. The near normal parallel position of the roots of the permanent canines was achieved by early treatment planning of extraction of the lateral incisors instead of treating by other modalities which have a poor prognosis. PMID- 10863494 TI - Evaluation of a curriculum for dental health in 3rd grade school children in Mumbai, India. AB - Utilising the Bright Smiles, Bright Future curriculum as modified for India, a trial on its effectiveness was carried out at the Raja Shivaji School in Mumbai, India. Students from the third standard English medium were selected. The teacher training programme utilised a professional educator from the U.S.A. plus other international representatives including India. Prior to the beginning of any classroom instruction, all experimental and control students were scored for plaque. A pretest was also given to both the experimental and control students to measure their knowledge of dental health. At the end of the two month test period all measurements were repeated. The curriculum was evaluated by the teachers. Parents were also interviewed. Results showed significant (P < .0001) difference between the two groups in reducing plaque, which demonstrated the improvement in oral hygiene in the experimental group. PMID- 10863495 TI - Uncommon mesiodens--a report of two cases. AB - Mesiodens is a supernumerary tooth with a cone shaped crown and a short root situated between the maxillary central incisors. Supernumerary teeth occur as isolated dental findings or as part of a syndrome. They are frequently discovered when a normal tooth is either delayed in its eruption or displaced often resulting in arch length inadequacy. An early diagnosis allows early intervention, more favourable prognosis and minimal complications. Presented here are two cases of unusual bilateral mesiodens. PMID- 10863496 TI - Relationship between the existing caries status, plaque S. mutans and Cariostat caries activity test in children. AB - An attempt was made in this study to find out the sensitivity and specificity of a caries activity test, CARIOSTAT and its relationship to the existing caries status and the plaque S. mutans level. The test proved to be highly sensitive and specific with significant relationship to the S.mutans count in the dental plaque. There also was a significant relationship between both the cultured microorganisms on MSB agar and the plaque in the Cariostat medium. PMID- 10863497 TI - Hypohidrotic ectodermal dysplasia--a case report. AB - A case presented here is that of a nine year old male patient with total anodontia. Findings of this case as regards to the orofacial, radiographic and other general manifestations were suggestive of hypohidrotic ectodermal dysplasia. The dental problems were best managed by prosthetic replacement of dentition taking into consideration a design which would provide adequate relief for the preservation of the ridges which were thin and underdeveloped to the absence of teeth. PMID- 10863498 TI - Peripheral giant cell granuloma--a case report. AB - Peripheral giant cell granuloma is a lesion arising mainly from the connective tissue of gingiva or periosteum of alveolar ridge. A case of peripheral giant cell granuloma involving a deciduous molar and the succedaneous tooth is reported. The lesion was large and interfered with occlusion. Surgical excision of the lesion along with the deciduous first molar was done. The underlying permanent first premolar was also involved, and had to be removed. The importance of an adequate salivary flow and maintenance of oral hygiene in the prevention of such lesions is stressed. PMID- 10863499 TI - Caries prevalence and its relation to socio-economic status and oral hygiene practices in 600 pre-school children of Kerala-India. AB - This Study was undertaken to determine the caries status of pre-school children in Ulloor Panchayat of Trivandrum, Kerala and to determine the relation if any, between their caries and socio-economic status and oral practices. 200 children each from the low, middle arid higher socio-economic group were visually examined for caries by the same examiner in natural daylight. Caries was recorded according to the WHO criteria. Information regarding the childrens' oral hygiene practices were obtained through structured questionnaires to the care takers of the children. Computer analysis of the data collected showed that 43% of the study sample were caries free. Socio-economic level was found to have a negative association with caries status. The mode of tooth brushing was found to be significantly related to caries severity while the frequency of tooth brushing was found to have no association with caries prevalence and severity. PMID- 10863500 TI - Effect of endodontic smear layer and various solvents on the calcium ion diffusion through radicular dentin--an in vitro study. AB - External root resorption and ankylosis remains the major cause of failure of replanted teeth. This study was conducted to explore the different ways to increase the pH of periradicular area in order to overcome the problem of root resorption and ankylosis. 60 freshly extracted permanent anteriors were used after removing the crown at CEJ. After biomechanical preparation Ca (OH)2 was injected and assays were done using EDTA, Citric and tannic acid Assays were repeated. Calcium diffusion and pH in the root exterior was measured using spectrophotometer. Results showed that dentin is permeable to calcium & hydroxyl ions and placement of Ca (OH)2 in the canal resulted in its increased recovery and alkaline pH periradicularly. Smear layer removal did not result in significant increase in Ca++ recovery or alkaline pH however combination of EDTA & NaOCl was found best than the other two. PMID- 10863501 TI - Orodental abnormalities in lobster claw syndrome (a type of syndactyly). AB - Lobster Claw Syndrome is a type of syndactyly, where abnormalities in the hand foot region as well as the orodental region occur. A case of a 14-year old boy with this syndrome is presented here. PMID- 10863502 TI - Inverted mesiodens--a case report. AB - A nine and a half years old female school child was examined in a during routine dental examination. The patient had swelling over maxillary midline area just near the labial frenum. Both the central incisors were in position. Radiographic examination revealed presence of an inverted supernumerary tooth between the roots of the central incisors. Surgical extraction of the supernumerary was planned. A unique case of inverted (upside down) mesiodens is presented. PMID- 10863503 TI - Trace elements in enamel of sound primary and permanent teeth. AB - Chemically made enamel is a dynamic substance rather than inert. Trace elements of enamel take active part in demineralisation, remineralization, ionic exchange, various physiological and pathological conditions. PMID- 10863504 TI - Relationship between cariogenic diet and dental caries as evaluated from a 5-day diet diary in 4-12 year-old children. AB - A preliminary study was conducted on 50 children in the age group of 4-12 years, who were divided into two groups on the basis of decayed, missing and filled teeth (DMFT) i.e. Group A (1-3) and Group B (> 3). A 5-day diet diary was evaluated and Sweet Score, Total Sugar Exposure, At Meal Sugar Exposures and Between Meal Sugar Exposure were calculated. There was statistically significant difference between the two groups in relation to Sweet Score and Total sugar Exposures. Between Meal Sugar Exposure and At Meal sugar exposure did not differ significantly. PMID- 10863505 TI - Salivary calculus: an insight into its pathogenesis--a case report of parotid sialolith in a 9 year old child. AB - The occurrence of sialolithiasis in children is uncommon, while parotid sialoliths are rare. A case of parotid sialolith in a 9 year old child is reported. PMID- 10863506 TI - "Talon cusp-heredity origin"--a case report. AB - Talon cusp is a very unusual anomalous structure of tooth. The etiology is still unknown. It may be due to mal-interaction between ecto and mesoderm of epithelial bulgings present on premaxillary region at the time of complex odontogenesis. Genetics may have some role in the formation of Talon cusp. Talon cusp may cause clinical complications. PMID- 10863507 TI - Traumatic bone cyst--a case report. AB - Traumatic bone cyst is an asymptomatic, slow growing, non expansile lesion commonly diagnosed during routine radiographic examination of the jaw bones. It is more frequently seen in young age, with predilection for anterior region of the mandible leading to a dramatic healing of the lesion. A typical case of traumatic bone cyst in a 12 year old girl is reported. A routine radiologic assessment of the patient with panoramic radiograph revealed a fairly large lesion in the anterior region of the mandible. On surgical exploration, clinical diagnosis was confirmed. Post operative successive radiograph shows progressive osseous healing. PMID- 10863508 TI - Peripheral osteoma of mandible arising from anterior lingual alveolar plate--a case report. AB - Peripheral osteomas of the mandible are uncommon bony tumours. Of those that have been described, the location is normally posterior to the premolars on the lingual surface of the mandible or in the condylar area. This article presents a case of an atypical presentation of an osteoma arising from the anterior lingual alveolar cortical plate of the mandible. PMID- 10863509 TI - Prevalence of dental caries and risk assessment among primary school children of 6-12 years in the Varkala municipal area of Kerala. AB - This epidemiological survey attempted to establish the prevalence and severity of dental caries among primary school children of Varkala municipal area. The prevalence of dental caries was 68.5% with a standard error (SE) of 1.64% and 95%, confidence interval (CI) 65.18, 71.82. The highest caries prevalence was found among 10 year age group (75.9%) and lowest in the 8 year age group (63%). The highest dmft score was found in 9 year age group 2.73 +/- 0.443 and highest DMFT score was found in 12 year age group 2.06 +/- 0.3824. Statistically significant association was found with dental caries and oral hygiene status (Odds Ratio (OR) 3.59, 95% CI, 2.53, 5.06 and oral cleanliness OR 2.73, 95% CI 2.96, 3.82). Statistically significant association was found between low socioeconomic status and prevalence of caries (O.R. 1.89, 95% CI--1.28, 2.8). PMID- 10863510 TI - Connation of permanent incisors: a report of two cases. AB - Presented here are two uncommon cases of connation of permanent incisors. The term connation is being used in this paper instead of fusion, gemination etc., as it ideally describes the anomaly. Various treatment modalities have also been recommended for such cases. PMID- 10863511 TI - Effects of variation of the timing of palatal repair on nasality of speech in complete cleft lip and palate children. AB - Nasality is related to factors like velopharyngeal closure and acoustic factors pertaining to cavities. The present investigation is a retrospective study aimed at evaluating the effects of variation in the timing of palatal repair on nasality during speech development in complete cleft lip and palate cases. It has been observed that the delay in palatal repair is associated with increase in nasality. Also, from the operated complete cleft lip and palate cases, it has been observed that the early and medium repair groups had almost similar effects on nasality of speech. (if they were operated before 36 months of age). PMID- 10863512 TI - Parental presence in the dental operatory-parent's point of view. AB - Parental presence in the dental operatory is considered a controversial issue and majority of dentists prefer mothers not to be present in the operatory while the child receives dental care. Very few studies have focussed on the parental' point of view. The present study was carried out to find out the attitudes of parents towards being present determine in the operatory during dental procedure and also to determine if willingness to be present was influenced by variables like age, sex, order of the child among siblings and previous dental behavior. Data was collected from 1350 parents using a questionnaire designed by the authors. In the present study, 78.3% of parents expressed their willingness to be present with the child during dental procedures. The study also showed that parents of younger children were more likely to be willing to be present in the operatory and as age advanced, the percentage of parents willing to be present in the operatory decreased. PMID- 10863513 TI - Oral and dental conditions in adult African wild dog skulls: a preliminary report. AB - Skulls of 29 adult African wild dogs (Lycaon pictus) originating from museum collections were examined for evidence of oral pathology. A wide variety of conditions similar to those seen in the domestic dog were detected. Although other reports suggest that captive African wild dogs suffer more extensively from dental disease than those in the wild, we conclude that these wild carnivores suffer from the same oral diseases as their domestic relatives, suggesting that a natural diet does not protect against these diseases. As the African wild dog is threatened by extinction, further investigation of the incidence and development of oral and dental disease in this species may be of value. Preventive measures could be instituted leading to improved health for those held in captivity. PMID- 10863514 TI - A removable orthodontic device for the treatment of lingually displaced mandibular canine teeth in young dogs. AB - The malocclusion of lingually displaced mandibular canine teeth is a common orthodontic problem in the domestic dog. Several treatment methods have been described, and their advantages and disadvantages have been extensively reviewed. This article describes a functional technique used in 38 dogs of different breeds for correction of the malocclusion. The technique consists of stimulating the dog to play with specific toys. It is a simple, inexpensive, non-invasive technique that has a success rate comparable to conventional orthodontic techniques for treatment of this common malocclusion. PMID- 10863515 TI - Effect of a new dental hygiene chew on periodontal health in dogs. AB - A study was undertaken to determine the effect of a new dental hygiene chew on periodontal health in the dog. The textural properties of this chew are different from previously tested dental hygiene products. The accumulation of dental deposits, development of oral malodor, and development of gingivitis were assessed in two groups of dogs; one fed a dry diet only, and the other fed the same dry diet supplemented by the daily addition of the new dental hygiene chew. Daily addition of the chew to the dry diet was effective in reducing plaque and calculus accumulation on the tooth surfaces, and also reduced the severity of gingivitis and oral malodor as compared to feeding the dry diet only. PMID- 10863516 TI - Endodontic disease of the mandibular first molar tooth secondary to caudal crossbite in a young Shetland sheepdog. AB - A six month-old intact female Shetland sheepdog was referred to the University of Illinois Veterinary Dental Clinic with a left-sided mandibular deviation and a thickened left ventral mandible in the region of the first molar tooth. On oral examination, left caudal crossbite was diagnosed. Dental radiographs revealed endodontic disease of the mandibular first molar tooth involved in the crossbite. Because of the difficulty of treating caudal crossbite and the potential of a pathological mandibular fracture, the endodontically affected tooth was extracted. Ten months following the extraction, mandibular deviation and alveolar bone lysis were resolved, but alveolar ridge resorption was present. The abnormal occlusal relationship caused by the caudal crossbite may have led to movement of the tooth, resorption of the tooth alveolus, and irreversible pulpal damage. Although not employed in this case, use of alveolar ridge preservation techniques can prevent mandibular bone loss after extractions. PMID- 10863517 TI - Timing of apical closure of the maxillary canine and mandibular first molar teeth of cats. AB - Stage of apical closure was determined by radiographing the jaws of 15 cats at intervals from 5-11 months of age. The apices of the left mandibular first molar tooth were closed in all 15 cats by seven months of age and the apices of the left maxillary canine teeth were all closed by 11 months of age. The implications of the age of closure on endodontic pathology and treatment are discussed. PMID- 10863518 TI - Review of studies assessing plaque accumulation and gingival inflammation in dogs. AB - Periodontal disease is difficult to measure objectively. Many indices measuring plaque accumulation and gingivitis have been designed for humans, the Silness and Loe plaque index and Turesky modification of the Quigley and Hein plaque index being examples of well-accepted systems. It may, however, be beneficial to consider new or modified measurement systems for dogs, and such veterinary modifications need to be supported and clearly identified. This article reviews the origins of clinical periodontal indices now in common use in studies that examine the effectiveness of oral hygiene products. PMID- 10863519 TI - Mandibular canine tooth impaction in a young dog--treatment and subsequent eruption: a case report. AB - Extraction of an embedded supranumerary incisor tooth and surgical exposure of the crown of an impacted left mandibular canine tooth were performed in a 5 month old Doberman Pinscher dog. Six months following surgery, the canine tooth was fully erupted and in normal occlusion. A review of tooth eruption in the dog is provided. PMID- 10863520 TI - Long-term effects of a dental hygiene chew on the periodontal health of dogs. AB - The objective of the study was to investigate the long-term effects of feeding a dental hygiene chew that has been shown to be effective in promoting periodontal health in dogs in short-term studies. Oral malodor, calculus, and plaque scores were still significantly lower after 21 months in the group that was receiving the dental hygiene chew, although gingivitis scores no longer differed significantly. There were no reports of any adverse reactions during the study. The results of the study support that feeding of the dental hygiene chew six days per week reduces accumulation of dental deposits, helps maintain periodontal health, and increases the time interval between professional periodontal intervention. PMID- 10863521 TI - Radiographic study of the maxillary canine tooth of four mesaticephalic cats. AB - The radiopaque and radiolucent anatomical structures that are superimposed over the root of the maxillary canine tooth in mesaticephalic cats were identified on digital radiographs made at various angles. The vomer bone, the nasal bone, the palatine fissure, and the infraorbital foramen were not superimposed over the root of the canine tooth in the range of angles examined. Superimposition with the palatine sulcus (which is rarely visible clinically because of silhouetting of the soft tissues) only occurred at extreme horizontal (cross-sectional arc) angles. The second premolar tooth was superimposed at a cross-sectional angle of 80 degrees and 90 degrees. The structures of concern in the interpretation of radiographs of the maxillary canine tooth in mesaticephalic cats are the conchal crest, the line of conjunction between the vertical body of the maxilla and its palatine process, the incisivomaxillary canal (which is rarely visible on radiographic images), and the lachrymal canal. Because of their anatomical vicinity, the radiographic position of these structures relative to the maxillary canine tooth can only be minimally changed. It was not possible to identify an "ideal" angle to radiograph the maxillary canine tooth in these four mesaticephalic cats. However, an acceptable compromise between minimal distortion of the image and satisfactory visualization of the root was obtained with the radiographic beam (rostro-caudal rotation) angled at 80 degrees and the skull (rotation in cross-sectional arc) angled at 70 degrees. PMID- 10863522 TI - Endodontic treatment and metal crown restoration of a fractured maxillary right fourth premolar tooth: a case report. AB - A 2 year-old, spayed female Shetland sheepdog presented with a fractured maxillary right fourth premolar tooth during oral examination for a dental prophylaxis. Seven months after conventional root canal therapy was performed, the tooth was prepared for non-precious metal, full crown restoration, which was cemented in place 1 month after crown margin preparation. During re-examination for an unrelated problem 6 months later, the metal crown was in place and there was no evidence of further fracture of the tooth. PMID- 10863523 TI - The crystalline components of dental calculus in the domestic cat. AB - Feline dental calculus was found to consist of carbonate-containing hydroxyapatite, Ca10(PO4)3(CO3)3(OH)2. Other forms of calcium phosphates consistently present in human calculus, and calcium carbonates found in horse, dog, and miniature pig calculus were not present. Calculus composition was performed using wet chemical method, x-ray diffraction, spectrographic analysis, and infrared spectrophotometric analysis. All samples showed traces of amorphous material, including magnesium and ammonium, but were negative for uric acid, cysteine, and oxalate. PMID- 10863524 TI - Elephant dental pulp tissue: where are the nerves? AB - Dental pulp tissue from three elephants was examined histologically with hematoxylin and eosin and s-100 protein stains. In all specimens, normal pulp was found with the exception that no nerve fibers (myelinated or non-myelinated) were demonstrable in any of the numerous sections prepared. PMID- 10863526 TI - Human limits for hypoxia. The physiological challenge of climbing Mt. Everest. AB - Climbing Mt. Everest without supplementary oxygen presents a fascinating physiological challenge because, at the summit, humans are very near the limit of tolerance to hypoxia. It was not until 1978 that the feat was accomplished, and this was after many unsuccessful attempts over a period of more than 50 years, and several physiological studies that suggested that it would be impossible. An analysis shows that the critical factors for reaching the summit are the enormous hyperventilation which is necessary to maintain the alveolar PO2 at viable levels, the fact that the barometric pressure is substantially higher than predicted by the Standard Atmosphere, and the severe respiratory alkalosis that assists loading of oxygen by the blood in the lung. Even so the maximal oxygen consumption on the summit is extremely low with the result that climbers are critically vulnerable to unexpected setbacks such as changes in the weather. PMID- 10863525 TI - Fifty years of radical ideas. AB - My role in the free radical theory of oxygen toxicity is discussed. Rebeca Gerschman and I published several papers on this subject. This sparked my interest in geochemistry and I developed the idea that oxygen was the best qualified biological potential energy source for the following reasons: great abundance, easily accessible, possession of a high thermodynamic potential, and its slow reaction rate. Ionization radiation can be viewed as a catalyst for reactive oxygen species since a killing dose imparts an infinitesimal small amount of energy. Next, Carol A. Colton and I showed that in the mammalian brain that stimulated microglia produce the superoxide radical anion and its implications in Alzheimer's disease is discussed. More recently, I have become interested in the role of sulfhydryl groups in transcription factors. PMID- 10863527 TI - Radiation, radicals, and images. AB - Nitroxide stable free radicals exhibit varied chemical and biological properties. Their biological applications have been greatly expanded over the past few years. Not only have they been shown to exhibit potent antioxidant and radioprotective properties, but also they can serve as in vivo functional imaging probes that non invasively report on the oxygen status and redox properties of tissue, which may have utility in clinical biomedical research. PMID- 10863528 TI - Radioprotection by antioxidants. AB - The role of reactive oxygen species in ionizing radiation injury and the potential of antioxidants to reduce these deleterious effects have been studied in animal models for more than 50 years. This review focuses on the radioprotective efficacy and the toxicity in mice of phosphorothioates such as WR 2721 and WR-151327, other thiols, and examples of radioprotective antioxidants from other classes of agents. Naturally occurring antioxidants, such as vitamin E and selenium, are less effective radioprotectors than synthetic thiols but may provide a longer window of protection against lethality and other effects of low dose, low-dose rate exposures. Many natural antioxidants have antimutagenic properties that need further examination with respect to long-term radiation effects. Modulation of endogenous antioxidants, such as superoxide dismutase, may be useful in specific radiotherapy protocols. Other drugs, such as nimodipine, propranolol, and methylxanthines, have antioxidant properties in addition to their primary pharmacological activity and may have utility as radioprotectors when administered alone or in combination with phosphorothioates. PMID- 10863529 TI - Ionizing radiation potentiates the induction of nitric oxide synthase by interferon-gamma and/or lipopolysaccharide in murine macrophage cell lines. Role of tumor necrosis factor-alpha. AB - Macrophages respond to infection or injury by changing from a "resting" cellular phenotype to an "activated" state defined by the expression of various cytotoxic effector functions. Regulation of the transition from a resting to an activated state is effected by cytokine and/or pathogenic signals. Some signals do not directly induce activation, but instead "prime" the macrophage to respond more vigorously to a second signal. One example of this priming phenomenon involves induction of nitric oxide (NO) synthesis by the enzyme nitric oxide synthase (NOS2). Our experiments indicate that low doses (1-5 Gy) of ionizing radiation can enhance the induction of enzymatically active NOS2 by IFN-gamma or LPS in J774.1 and RAW264.7 murine macrophage cell lines. Radiation alone did not produce this induction, rather, it was effective as a priming signal; cells exposed to radiation produced more NO when a second signal, either IFN-gamma or LPS, was applied 24 h later. PMID- 10863530 TI - Genetic responses to free radicals. Homeostasis and gene control. AB - Gene regulation mechanisms have evolved allowing cells to finetune the level of "endogenous" oxidative stress and to cope with increased free radicals from external sources. Levels of H2O2 are tightly controlled in E. coli by OxyR, which is activated by H2O2 to increase scavenging activities and limit H2O2 generation by the respiratory chain. Sub-micromolar levels of H2O2 are maintained in mammalian tissues, though the regulatory systems that govern this control are unknown. Excess superoxide triggers the soxRS system in E. coli, which is controlled by the oxidant-sensitive iron-sulfur centers of the SoxR protein. Nitric oxide activates SoxR by a different modification of the iron-sulfur centers. The soxRS regulon mobilizes diverse functions to scavenge free radicals and repair oxidative damage in macromolecules, and other mechanisms that exclude many environmental agents from the cell. Mammalian cells also sense and respond to sub-toxic levels of nitric oxide, activating expression of heme oxygenase 1 through stabilization of its mRNA. These inductions give rise to adaptive resistance to nitric oxide in neuronal and other cell types. PMID- 10863531 TI - Mapping oxidative DNA damage and mechanisms of repair. AB - We developed a method to map oxidative-induced DNA damage at the nucleotide level using ligation-mediated polymerase chain reaction (LMPCR) technology. In vivo and in vitro DNA base modification patterns inflicted by reactive oxygen species (ROS) in the human P53 and PGK1 gene were nearly identical in vitro and in vivo. In human male fibroblasts, these patterns are independent of the transition metal used (Cu (II), Fe(II), or Cr(VI). Therefore, local probability of H2O2-mediated DNA base damage is determined primarily by DNA sequence. Moreover, in cells undergoing severe oxidative stress, extranuclear sites contribute metals that enhance nuclear DNA damage. The role of the base excision repair pathway in human cells responsible for the repair of the majority of ROS base damage is also discussed. PMID- 10863532 TI - Post-transcriptional regulation of lung antioxidant enzyme gene expression. AB - It is an honor, and indeed fitting, to have a chapter on pulmonary oxygen toxicity included in a Festschrift for Dan Gilbert, whose contributions to the free radical theory of oxygen toxicity have been a catalyst to the last half century of investigation in this field. There is cellular damage that results in pulmonary edema and even death if the increase in reactive oxygen species produced in the lung during exposure to hyperoxia is not counterbalanced by an increase in the cell's antioxidant defense systems. In this chapter experimental evidence will substantiate the importance of post-transcriptional regulation of antioxidant enzyme gene expression in animal models of pulmonary oxygen toxicity and tolerance to hyperoxia with special emphasis given to the role of manganese superoxide dismutase (MnSOD) synthesis, specific activity, and RNA half-life and to a proposed function of a MnSOD RNA-binding protein as a positive regulator in the control of translational efficiency. PMID- 10863533 TI - Oxidative stress, mitochondrial respiration, and Parkinson's disease. AB - When either oxidizing species, such as H2O2 or oxy-radicals, are present in excess or cellular anti-oxidant defenses are lowered, a state of oxidative stress exists. Parkinson's disease is characterized by the loss of dopamine (DA) neurons, which leads to overactivity of the surviving DA neurons and an increase in neurotransmitter release and turnover. The increased metabolism of DA neurotransmitter by monoamine oxidase (MAO) can be looked upon as an endogenous oxidative stress, leading to damage to Complex I-linked mitochondrial respiration. It remains an open question to what extent the mitochondrial damage seen in Parkinson's disease is of genetic origin and how much is caused by H2O2 generated during enhanced turnover of DA, especially during treatment with L dopa. PMID- 10863534 TI - Regulation of mitochondrial respiration by oxygen and nitric oxide. AB - Although the regulation of mitochondrial respiration and energy production in mammalian tissues has been exhaustively studied and extensively reviewed, a clear understanding of the regulation of cellular respiration has not yet been achieved. In particular, the role of tissue pO2 as a factor regulating cellular respiration remains controversial. The concept of a complex and multisite regulation of cellular respiration and energy production signaled by cellular and intercellular messengers has evolved in the last few years and is still being researched. A recent concept that regulation of cellular respiration is regulated by ADP, O2 and NO preserves the notion that energy demands drive respiration but places the kinetic control of both respiration and energy supply in the availability of ADP to F1-ATPase and of O2 and NO to cytochrome oxidase. In addition, recent research indicates that NO participates in redox reactions in the mitochondrial matrix that regulate the intramitochondrial steady state concentration of NO itself and other reactive species such as superoxide radical (O2-) and peroxynitrite (ONOO-). In this way, NO acquires an essential role as a mitochondrial regulatory metabolite. No exhibits a rich biochemistry and a high reactivity and plays an important role as intercellular messenger in diverse physiological processes, such as regulation of blood flow, neurotransmission, platelet aggregation and immune cytotoxic response. PMID- 10863535 TI - Free radicals and antioxidants in the year 2000. A historical look to the future. AB - In the late 1950's free radicals and antioxidants were almost unheard of in the clinical and biological sciences but chemists had known about them for years in the context of radiation, polymer and combustion technology. Daniel Gilbert, Rebeca Gerschman and their colleagues related the toxic effects of elevated oxygen levels on aerobes to those of ionizing radiation, and proposed that oxygen toxicity is due to free radical formation, in a pioneering paper in 1956. Biochemistry owes much of its early expansion to the development and application of chromatographic and electrophoretic techniques, especially as applied to the study of proteins. Thus, superoxide dismutase (SOD) enzymes (MnSOD, CuZnSOD, FeSOD) were quickly identified. By the 1980's Molecular Biology had evolved from within biochemistry and microbiology to become a dominant new discipline, with DNA sequencing, recombinant DNA technology, cloning, and the development of PCR representing milestones in its advance. As a biological tool to explore reaction mechanisms, SOD was a unique and valuable asset. Its ability to inhibit radical reactions leading to oxidative damage in vitro often turned out to be due to its ability to prevent reduction of iron ions by superoxide. Nitric oxide (NO.) provided the next clue as to how SOD might be playing a critical biological role. Although NO. is sluggish in its reactions with most biomolecules it is astoundingly reactive with free radicals, including superoxide. Overall, this high reactivity of NO. with radicals may be beneficial in vivo, e.g. by scavenging peroxyl radicals and inhibiting lipid peroxidation. If reactive oxygen species are intimately involved with the redox regulation of cell functions, as seems likely from current evidence, it may be easier to understand why attempts to change antioxidant balance in aging experiments have failed. The cell will adapt to maintain its redox balance. Indeed, transgenic animals over-expressing antioxidants show some abnormalities of function. There must therefore be a highly complex interrelationship between dietary, constitutive, and inducible antioxidants with the body, under genetic control. The challenge for the new century is to be able to understand these relationships, and how to manipulate them to our advantage to prevent and treat disease. PMID- 10863536 TI - Reflections on the role of the thiol group in biology. AB - This essay is concerned with the role of the thiol or sulfhydrvl group in cellular function and metabolism and with the important investigations over many years that have led us to a better understanding of the importance of this molecular moiety that plays such a vital role in biology. The tools for measuring the SH group and for inhibiting or regenerating it will be discussed as will its essential role in the actions of many enzymes. The importance of the thiol group in glycolysis and in energy production by mitochondria will be emphasized. Of special interest at present is the fact that certain low molecular weight SH containing substances can mimic some of the actions of insulin and may become of benefit in the treatment of diabetes mellitus. Finally, the toxic effects of oxygen on metabolism and function will be discussed with particular reference to the possibility that oxidation of thiol groups may play a role in the manifestations of oxygen toxicity. PMID- 10863537 TI - Differential regulation of MAP kinase signaling by pro- and antioxidant biothiols. AB - Some biologically derived thiol-containing compounds have potential for health benefits whereas others elicit biochemical events leading to pathogenesis. Effects of two biothiols, alpha-lipoic acid (alpha LA), a therapeutic antioxidant, and homocysteine (Hcy), a risk factor for age-associated cardiovascular disease, on cell signaling events involving p44 and p42 MAP kinases (p44/42 MAPK) were evaluated in cell culture. Treatment of serum-deprived NIH/3T3 cells with Hcy (20 microM) resulted in the activation of p44/42 MAPK as determined by Western blot analysis using the phospho-specific p44/42 MAPK antibody. p44/42 MAPK phosphorylation was rapid and transient with maximal activation occurring at 10-30 min. Transient activation of p44/42 MAPK was also observed in response to treatment of serum-deprived cells with alpha LA. In cells grown in serum, serum-dependent p44/42 MAPK phosphorylation was transiently enhanced by Hcy or Hcy thiolactone, but inhibited by alpha LA. Thus, alpha LA and Hcy differentially influence signal transduction events depending on the state of cells. These observations may be important in understanding how some biothiols are associated with pathogenic events while others have potential as therapeutic agents. PMID- 10863539 TI - Molecular markers of oxidative stress vulnerability. AB - In astrocyte primary cultures of trisomy 16 mice, an animal model for Down's syndrome, protein oxidation was 50% higher than in diploid littermates. Exposure to 10 microM H2O2 or 50 microM kainic acid incremented protein oxidation in trisomic but not in diploid cultures. Studies on stress response genes showed that metallothionein (MT) level was 2-3 times higher in trisomy 16 than in diploid cultures. Kainic acid or H2O2 exposure increased the MT protein level in diploid cultures but failed to increase it in trisomy 16 mouse beyond its elevated basal level. The reduced responsiveness of MT to simulated oxidative stress may result in insufficient removal of ROS, which could partially explain the further increase of protein oxidation in trisomy 16 cultures. In contrast, Pb exposure increased MT in trisomy 16 and diploid primary cultures to a similar extent. The similar metal responsiveness of MT in both phenotypes indicated that MT in trisomic glial cultures was not yet maximally stimulated. The flawed redox sensitivity in trisomy 16 mouse suggests possible alterations in the binding activity of ROS-sensitive transcription factors on the MT promoter. PMID- 10863538 TI - Enzyme-like activity of glycated cross-linked proteins in free radical generation. AB - The structure and property of cross-linked amino acids and proteins produced by a three- carbon alpha-dicarbonyl methylglyoxal in glycation reaction were investigated. Our results showed that these reactions generated yellow fluorescent products and several free radical species. From the reaction with alanine, three types of free radicals were identified by EPR spectroscopy: 1) the cross-linked radical cation, methylglyoxal diaklylimine cation radical; 2) the methylglyoxal radical anion as the counterion; 3) the superoxide radical anion produced only in the presence of oxygen. Glycation of bovine serum albumin by methylglyoxal also generated the protein-bound, cross-linked free radical, probably the cation radical of the cross-linked Schiff base as observed with alanine. The glycated protein reduced ferricytochrome c to ferrocytochrome c in the absence of oxygen or added metal ions. This reduction of cytochrome c was accompanied by a large increase in the amplitude of the electron paramagnetic resonance signal originated from the protein-bound free radical. In addition, the glycated protein catalyzed the oxidation of ascorbate in the presence of oxygen while the protein-free radical signal disappeared. These results indicate that glycation of protein generates active centers for catalyzing one-electron oxidation-reduction reactions. This active center, which exhibits enzyme-like character, was suggested to be the cross-linked Schiff base/the cross-linked Schiff base radical cation of the protein. It mimics the characteristics of metal catalyzed oxidation system. These results together indicate that glycated proteins accumulated in vivo provide stable active-sites for catalyzing the formation of free radicals. PMID- 10863540 TI - Protein oxidation. AB - The oxidative modification of proteins by reactive species, especially reactive oxygen species, is implicated in the etiology or progression of a panoply of disorders and diseases. These reactive species form through a large number of physiological and non-physiological reactions. An increase in the rate of their production or a decrease in their rate of scavenging will increase the oxidative modification of cellular molecules, including proteins. For the most part, oxidatively modified proteins are not repaired and must be removed by proteolytic degradation, and a decrease in the efficiency of proteolysis will cause an increase in the cellular content of oxidatively modified proteins. The level of these modified molecules can be quantitated by measurement of the protein carbonyl content, which has been shown to increase in a variety of diseases and processes, most notably during aging. Accumulation of modified proteins disrupts cellular function either by loss of catalytic and structural integrity or by interruption of regulatory pathways. PMID- 10863541 TI - Mechanisms of cell death governed by the balance between nitrosative and oxidative stress. AB - Many cellular functions in physiology are regulated by the direct interaction of NO with target biomolecules. In many pathophysiologic and toxicologic mechanisms, NO first reacts with oxygen, superoxide or other nitrogen oxides to subsequently elicit indirect effects. The balance between nitrosative stress and oxidative stress within a specific biological compartment can determine whether the presence of NO will be ultimately deleterious or beneficial. Nitrosative stress can be defined primarily through reactions mediated by N2O3, a reactive nitrogen oxide species generated by high fluxes of NO in an aerobic environment. In contrast, oxidative stress is mediated primarily by superoxide and peroxides. In addition to reactive oxygen species, several reactive nitrogen oxide species such as peroxynitrite, nitroxyl, and nitrogen dioxide can also impose oxidative stress to a cell. We here describe how the mechanisms of cell death are interwoven in the balance between the different chemical intermediates involved in nitrosative and oxidative stress. PMID- 10863542 TI - Nitrone inhibition of age-associated oxidative damage. AB - The mechanistic basis of the neuroprotective activity of the nitrone-based free radical trap PBN (alpha-phenyl-N-tert-butyl nitrone) has been investigated extensively. Key observations exclude its simple mass action spin trapping of free radicals activity as the key mechanism of action. These include: A) the fact that it protects in experimental stroke even if administered several hours after the event and B) the fact that its chronic low-level administration to old experimental animals reverses their age-enhanced susceptibility to stroke even several days after the last dosage. PBN was found to inhibit gene induction in several models including stroke and an LPS-mediated septic shock model. Stoke causes inducible nitric oxide synthase (iNOS) to be expressed. High levels of nitric oxide and peroxynitrite (formed from nitric oxide), produced by iNOS, is particularly neurotoxic. PBN inhibits iNOS induction. Therefore, it seems that prevention of the formation of neurotoxic products is a rational mechanism of action of PBN in the stroke model. There is strong rationale to consider that there is an enhanced propensity for a "smoldering" neuro-inflammatory state in the old brain. Reversal of this state by PBN may explain its action in preventing age-enhanced stroke susceptibility in old experimental animals. Significant new findings underscore the importance of neuro-inflammatory processes in neuronal death or dysfunction in Alzheimer's disease. Neuro-inflammatory processes implicate enhanced signal transduction processes. Strong evidence for this is the enhanced p38 kinase activation in neurons near plaques and tangles of the Alzheimer's brain in contrast to normal aged-matched control brain which did not show p38 activation. In rat primary astrocytes p38 activation by the pro inflammatory cytokine IL-1 beta, as well as by H2O2, was significantly suppressed by PBN. Mechanistically it was shown that PBN suppresses the amount of reactive oxygen species (ROS) produced in mitochondrial respiration. Much evidence indicates that ROS are signaling molecules and that they also are involved to maintaining brain phosphatases in an inactive state. We argue that finding a specific high affinity site mechanism for the neuroprotective action of PBN is unlikely based on the complexity of the system reflecting ROS generation and signal transduction processes that have apparently evolved to maintain adaptive responses. The promising pharmacological activity of molecules like PBN is not diminished by this however, for only excessive amounts of ROS is considered detrimental. The action of PBN in suppressing signal transduction processes, most likely by suppressing ROS production in mitochondrial respiration, effectively controls excessive oxidative damage and prevents induction of genes that form neurotoxic products. PMID- 10863543 TI - Effects of atypical antioxidative agents, S-nitrosoglutathione and manganese, on brain lipid peroxidation induced by iron leaking from tissue disruption. AB - A fluorescent assay of brain lipid peroxidation was used for screening new antioxidants for the prevention of neurodegeneration caused by free radicals. Incubation of rat brain homogenates led to a temperature-dependent increase in production of fluorescent adducts of peroxidized polyunsaturated fatty acids; it was inhibited completely by lowering the incubation temperature to 4 degrees C. This tissue disruption-induced brain lipid peroxidation at 37 degrees C was blocked by deferoxamine (IC50 = 0.3 microM) and EDTA; it was augmented by adding submicromolar iron and hemoglobin. Ferrous ion's pro-oxidative activities were five times more potent than ferric ion. Micromolar manganese completely inhibited lipid peroxidation, confirming earlier unexpected in vivo reports. Trolox and vitamin C suppressed brain lipid peroxidation with IC50 values of 20 and 500 microM, respectively. U-78517F was approximately 20 times more potent than Trolox. 17 beta-Estradiol, hydralazine, S-nitrosoglutathione and 3 hydroxybenzylhydrazine were as potent as Trolox. Melatonin, glutathione, alpha lipoic acid and l-deprenyl were about 20 times less potent than Trolox. Surprisingly, N-tert-butyl-alpha-phenylnitrone was a weak antioxidant. Furthermore, this procedure can also detect pro-oxidative side effects of vitamin C, oxidized glutathione, penicillamine and Angeli's salt. The present results obtained from this selective fluorescent assay are consistent with earlier reports that iron complexes promote while manganese inhibits brain lipid peroxidation caused by cell disruption. S-Nitrosoglutathione, melatonin, 17 beta estradiol, and manganese have been successfully tested in cell/animal models for their potential neuroprotective effects. In conclusion, monitoring fluorescent adducts of peroxidizing polyunsaturated fatty acids in brain homogenates is a simple, quantitative method for studying iron-dependent brain lipid peroxidation and for screening of potential neuroprotective antioxidants in both in vitro and in vivo preparations. PMID- 10863544 TI - Effect of MAO-B inhibitors on MPP+ toxicity in Vivo. AB - l-Deprenyl (Selegiline), a selective and irreversible type B monoamine oxidase inhibitor, has been used as an adjunct to levodopa therapy in Parkinson's disease. Recently, it is proposed as a putative neuroprotective agent in delaying the progression of cell death based on its capability of reducing the oxidative stress derived from the MAO-B dependent metabolism of dopamine, and blocking the development of MPTP-parkinsonism. However, a variety of experimental models suggest that l-deprenyl provides neuroprotection through multiple modes of mechanism other than the inhibition of MAO-B. We have previously shown that l deprenyl protects midbrain dopamine neurons from MPP+ toxicity by a novel antioxidant effect. In the present study we examined whether the protection against MPP+ toxicity is also shared by other reversible or irreversible MAO-B inhibitors including (+)-deprenyl, Ro16-6491 and pargyline. Our data show that non of these MAO-B inhibitors changes the dopamine loss in the striatum induced by intranigral injection of MPP+. Our result suggests that l-deprenyl may possess a unique neuroprotective action on nigral neuron against MPP+ toxicity independent of the MAO-B inhibition. PMID- 10863545 TI - Neuroprotective strategies in Parkinson's disease using the models of 6 hydroxydopamine and MPTP. AB - The etiology of Parkinson's disease is not known. Nevertheless a significant body of biochemical data from human brain autopsy studies and those from animal models point to an on going process of oxidative stress in the substantia nigra which could initiate dopaminergic neurodegeneration. It is not known whether oxidative stress is a primary or secondary event. Nevertheless, oxidative stress as induced by neurotoxins 6-hydroxydopamine and MPTP (N-methyl-4-phenyl-1,2,3,6 tetrahydropyridine) has been used in animal models to investigate the process of neurodegeneration with intend to develop antioxidant neuroprotective drugs. It is apparent that in these animal models radical scavengers, iron chelators, dopamine agonists, nitric oxide synthase inhibitors and certain calcium channel antagonists do induce neuroprotection against such toxins if given prior to the insult. Furthermore, recent work from human and animal studies has provided also evidence for an inflammatory process. This expresses itself by proliferation of activated microglia in the substantia nigra, activation and translocation of transcription factors, NF kappa-beta and elevation of cytotoxic cytokines TNF alpha, IL1-beta, and IL6. Both radical scavengers and iron chelators prevent LPS (lipopolysaccharide) and iron induced activation of NF kappa-B. If an inflammatory response is involved in Parkinson's disease it would be logical to consider antioxidants and the newly developed non-steroid anti-inflammatory drugs such as COX2 (cyclo-oxygenase) inhibitors as a form of treatment. However to date there has been little or no success in the clinical treatment of neurodegenerative diseases per se (Parkinson's disease, ischemia etc.), where neurons die, while in animal models the same drugs produce neuroprotection. This may indicate that either the animal models employed are not reflective of the events in neurodegenerative diseases or that because neuronal death involves a cascade of events, a single neuroprotective drug would not be effective. Thus, consideration should be given to multi-neuroprotective drug therapy in Parkinson's disease, similar to the approach taken in AIDS and cancer therapy. PMID- 10863546 TI - Neuroprotective antioxidants from marijuana. AB - Cannabidiol and other cannabinoids were examined as neuroprotectants in rat cortical neuron cultures exposed to toxic levels of the neurotransmitter, glutamate. The psychotropic cannabinoid receptor agonist delta 9 tetrahydrocannabinol (THC) and cannabidiol, (a non-psychoactive constituent of marijuana), both reduced NMDA, AMPA and kainate receptor mediated neurotoxicities. Neuroprotection was not affected by cannabinoid receptor antagonist, indicating a (cannabinoid) receptor-independent mechanism of action. Glutamate toxicity can be reduced by antioxidants. Using cyclic voltametry and a fenton reaction based system, it was demonstrated that Cannabidiol, THC and other cannabinoids are potent antioxidants. As evidence that cannabinoids can act as an antioxidants in neuronal cultures, cannabidiol was demonstrated to reduce hydroperoxide toxicity in neurons. In a head to head trial of the abilities of various antioxidants to prevent glutamate toxicity, cannabidiol was superior to both alpha-tocopherol and ascorbate in protective capacity. Recent preliminary studies in a rat model of focal cerebral ischemia suggest that cannabidiol may be at least as effective in vivo as seen in these in vitro studies. PMID- 10863547 TI - A positive-feedback model for the loss of acetylcholine in Alzheimer's disease. AB - We describe a two-component positive-feedback system that could account for the large reduction of acetylcholine that is characteristic of patients with Alzheimer's disease (AD). One component is beta-amyloid-induced apoptosis of cholinergic cells, leading to a decrease in acetylcholine. The other component is an increase in the concentration of beta-amyloid in response to a decrease in acetylcholine. We describe each mechanism with a differential equation, and then solve the two equations numerically. The solution provides a description of the time course of the reduction of acetylcholine in AD patients that is consistent with epidemiological data. This model may also provide an explanation for the significant, but lesser, decrease of other neurotransmitters that is characteristic of AD. PMID- 10863549 TI - Antioxidant status and human health. Use of cyclic voltammetry for the evaluation of the antioxidant capacity of plasma and of edible plants. AB - The low molecular weight antioxidants (LMWA) play a major role in protecting biological systems against reactive oxygen-derived species (ROS), and reflect the antioxidant capacity of the system. The cyclic voltammetry (CV) has been conveniently used and validated for the quantitation of the antioxidant capacity of the LMWA of blood plasma, tissue homogenates, and plant extracts. The CV tracing provides the biological oxidation potential (E and E1/2 which relate to the nature of the molecule(s)), the intensity of the anodic current wave (Ia), and its area S (both relate to the concentration of the molecule(s)). The components of the first anodic wave of plasma were identified by comparison with HPLC-electrochemical detection. CV together with another plasma parameter R, which reflects the level of oxidized ascorbate, were used for the evaluation of the antioxidant status and the oxidative stress in healthy subjects and in chronic (diabetes mellitus) and acute patients (subjected to total body irradiation prior to bone marrow transplantation). These methodologies could be widely employed for rapid evaluation of subjects, in health and disease, for monitoring of their response to treatment and nutritional supplementation, and for screening of specific populations. PMID- 10863548 TI - Microglial contribution to oxidative stress in Alzheimer's disease. AB - Microglia are the CNS macrophage and are a primary cellular component of plaques in Alzheimer's disease (AD) that may contribute to the oxidative stress associated with chronic neurodegeneration. We now report that superoxide anion production in microglia or macrophages from 3 different species is increased by long term exposure (24 hours) to A beta peptides. Since A beta competes for the uptake of opsonized latex beads and for the production of superoxide anion by opsonized zymosan, a likely site of action are membrane receptors associated with the uptake of opsonized particles or fibers. The neurotoxic fibrillar peptides A beta (1-42) and human amylin increase radical production whereas a non-toxic, non fibrillar peptide, rat amylin, does not. We also report that the effect of A beta peptides on superoxide anion production is not associated with a concomitant increase in nitric oxide (NO) production in either human monocyte derived macrophages (MDM) or hamster microglia from primary cultures. Since NO is known to protect membrane lipids and scavenge superoxide anion, the lack of A beta mediated induction of NO production in human microglia and macrophages may be as deleterious as the over-production of superoxide anion induced by chronic exposure to A beta peptides. PMID- 10863550 TI - Antioxidants in nutrition. AB - The harmful effects of oxidative processes in living organisms, in addition to chemical and biochemical media, can be reduced by antioxidants. The efficacy of an antioxidant depends on its reduction potential and kinetics of elimination of diverse free radicals. Redox potentials and reaction rate constants of selected gallocatechins and flavonoids were measured by pulse radiolysis and laser photolysis. The reduction potentials of the flavonoids studied were in the range of 0.33 V (quercetin) and 0.75 V (kaempferol). The rate constants of the superoxide radical with 15 flavonoids ranged from 10(5)-10(7) M-1 s-1. Singlet oxygen quenching by flavonoids was also very rapid (from 10(5) to 10(8) M-1 s-1). These studies may be crucial in optimizing health and increasing longevity by reducing oxidative stress and biological damage. PMID- 10863551 TI - Free radical intermediates in sonodynamic therapy. AB - Current understanding of the mechanism of sonodynamic action (i.e. the ultrasound dependent enhancement of the cytotoxic action of certain drugs--sonosensitizers) with potential applications for cancer therapy is presented. The experimental evidence suggests that sonosensitization is due to the chemical activation of sonosensitizers inside or in the close vicinity of hot collapsing cavitation bubbles to form sensitizer-derived free radicals either by direct pyrolysis or due to reactions with .H and .OH radicals, formed by pyrolysis of water. These free radicals (mostly carbon-centered) react with oxygen to form peroxyl and alkoxyl radicals. Unlike .OH and .H, which are also formed by pyrolysis inside cavitation bubbles, the reactivity of alkoxyl and peroxyl radicals with organic components dissolved in biological media is lower and hence have higher probability of reaching critical cellular sites. Sonodynamic therapy appears to be a promising modality for cancer treatment since ultrasound can penetrate deep within the tissue and can be focused in a small region of tumor to chemically activate relatively non-toxic molecules (e.g. porphyrins) thus minimizing undesirable side effects. PMID- 10863552 TI - Glucose deprivation-induced oxidative stress in human tumor cells. A fundamental defect in metabolism? AB - Recently, glucose deprivation-induced oxidative stress has been shown to cause cytotoxicity, activation of signal transduction (i.e., ERK1, ERK2, JNK, and Lyn kinase), and increased expression of genes associated with malignancy (i.e., bFGF and c-Myc) in MCF-7/ADR human breast cancer cells. These results have led to the proposal that intracellular oxidation/reduction reactions involving hydroperoxides and thiols may provide a mechanistic link between metabolism, signal transduction, and gene expression in these human tumor cells. The current study shows that several other transformed human cell types appear to be more susceptible to glucose deprivation-induced cytotoxicity and oxidative stress than untransformed human cell types. In a matched pair of normal and SV40-transformed human fibroblasts the cytotoxic process is shown to be dependent upon ambient O2 concentration. A theoretical model to explain the results is presented and implications to unifying modern theories of cancer are discussed. PMID- 10863553 TI - Cytomegalovirus gene regulation by reactive oxygen species. Agents in atherosclerosis. AB - Oxidative stress is implicated in the pathogenesis of atherosclerosis, and of viral infections caused by sendai virus, influenza and HIV. Vascular oxidative stress is due to inflammatory and immune responses of vascular cells, and to reperfusion after recanalization of blocked arteries. Because human cytomegalovirus (CMV) may contribute to atherogenesis by several mechanisms, and coronary artery smooth muscle cells (SMC) are permissive for the virus, we examined CMV interactions with SMC. Infection causes generation of intracellular reactive oxygen species (ROS) which activate NF-kappa B, a cellular transcription factor. NF-kappa B mediates expression of the CMV promoter and of genes involved in the immune and inflammatory responses. Antioxidants or aspirin inhibit ROS, NF kappa B and CMV. PMID- 10863554 TI - Reactive species in sickle cell disease. AB - The red cell is a relatively abundant locus of both free radical generation and reaction. Erythrocytes have a high content of unsaturated membrane lipids, a rich oxygen supply and are densely packed with redox-active hemoglobin residues. In response, red cells have a highly evolved and well-integrated network of oxidant defense mechanisms that lend an ability to withstand oxidative stress. In the case of congenital hemoglobin mutations that underlie sickle cell disease, they become very susceptible to free radical-mediated injury by virtue of enhanced endogenous rates of production of reactive species and impairment of tissue free radical defense mechanisms. In sickle cell disease, a combination of these susceptibility factors are hypothesized to lead to an overall impairment of vascular function, in large part due to loss of "bioactive" nitric oxide via the free radical-mediated consumption of this vasoactive molecule. PMID- 10863555 TI - Dopamine stimulates astrocytic C6-D2L cells via tyrosine kinase and p38 MAPK activation. PMID- 10863556 TI - Selenium biochemistry. Mammalian selenoenzymes. PMID- 10863557 TI - Cytoprotective properties of nisoldipine and amlodipine against oxidative endothelial cell injury. PMID- 10863559 TI - Neutrophil-endothelial cell interactions. Inverse correlation between nitric oxide and superoxide anions. PMID- 10863558 TI - The origin of dinitrosyl-iron complex in endothelial cells. PMID- 10863560 TI - Daniel L. Gilbert: explorer of life. AB - From the time Dan was 12 years old to the present, he always was the explorer of life. He was very fortunate in having an opportunity to see how life exists in several human societies around the world. His many professional foreign travels are presented. Dan went to a 1959 meeting in Argentina, to England in 1963, and to Chile in 1963. After he married Claire, she accompanied him on his 44 day around the world trip in 1965 and on all his later foreign trips. In 1966, there was a Latin American trip on the way to doing research at a laboratory in Chile. Next on his travels was being a consultant in Venezuela in 1969 and attending a meeting in Russia in 1972. His son Raymond joined in trips to France in 1977, to Australia in 1983, to a 1990 trip to a Peruvian meeting, and finally joined the 1991 meeting to a meeting in Japan, with a side trip to China where Dan gave lectures. The last 2 foreign trips for Dan with Claire, but not with Raymond, were the meetings in 1994 in Buenos Aires, Argentina, and the 1998 meeting, which was held in both Israel and Jordan. PMID- 10863561 TI - Experimental neurotoxicity of mercury. Autometallographic and stereologic studies on rat dorsal root ganglion and spinal cord. PMID- 10863562 TI - Ergocalciferol supplementation may positively affect lumbar spine bone mineral density of vegans. PMID- 10863563 TI - The presidential candidates of 2000. Their positions on health care, and why MNT is an important issue. PMID- 10863564 TI - Complementary/alternative medicine: another path to MNT coverage? PMID- 10863565 TI - Dietary reference intakes for the antioxidant nutrients: vitamin C, vitamin E, selenium, and carotenoids. PMID- 10863566 TI - Breast-feeding through the first year predicts maternal control in feeding and subsequent toddler energy intakes. AB - OBJECTIVE: Current recommendations for infant feeding encourage breast-feeding through the first year. This research was conducted to evaluate associations among breast-feeding, maternal control of child feeding, and the dietary intake of toddlers during the second year of life. In particular, we sought to determine whether breast-feeding through the first year and subsequent toddler intake was mediated via maternal control of child feeding. DESIGN/SUBJECTS: Fifty-five white infants and their mothers were monitored longitudinally from age 12 or 13 months to age 18 months. MAIN OUTCOME MEASURES: Breast-feeding through the first year and maternal control in infant feeding were evaluated as predictors of energy intake at age 18 months. STATISTICAL ANALYSES PERFORMED: Regression analysis was used to evaluate predictors of toddler energy intake at age 18 months. A mediation model tested if the relationship between breast-feeding and infant intake was mediated by maternal control in feeding. RESULTS: Breast-feeding through the first year was associated with higher toddler energy intakes at age 18 months through its influence on maternal control in feeding. Mothers who breast-fed their infants for at least 12 months used lower levels of control in feeding. Lower levels of maternal control in feeding were associated with higher toddler energy intakes. The highest energy intakes among children aged 18 months were observed among taller and leaner toddlers. APPLICATIONS/CONCLUSIONS: Our findings suggest that breast-feeding through the first year may have an effect on children's energy intake by shaping mothers' child-feeding practices. These findings may be used by clinicians to assist parents in making informed decisions about choice of infant-feeding method and to provide anticipatory guidance regarding infant-feeding style when initiating dietary diversity. PMID- 10863567 TI - Food acceptance and genetic variation in taste. AB - OBJECTIVE: To determine if individuals who taste 6-n-propylthiouracil (PROP), one marker of genetic variation in taste, as exceptionally bitter can also perceive sugars as sweeter, other bitters as more intense, and dietary fats as more creamy and/or viscous than do individuals who taste PROP as weakly bitter. This study examined the association between genetic variation in taste and acceptance for sweet, high-fat, and bitter foods and beverages. DESIGN: Genetic variation was measured by perceived bitterness of PROP (influenced by genetic, hormonal, and pathologic factors) and density of fungiform papillae on the anterior portion of the tongue (influenced primarily by genetic factors). Four sweet, 3 fat, and 3 bitter groups were derived from principal components analyses of questionnaire items. SUBJECTS: Convenience sample of healthy adults (24 women, 22 men; mean age +/- standard deviation = 21 +/- 6 years) who did not report high dietary restraint. STATISTICAL ANALYSES: Pearson product moment correlations between genetic taste measures and food and beverage groups. RESULTS: The sample showed diversity in genetic taste measures: perceived bitterness of 0.0032 mol/L PROP ranged from "weak" to well above "very strong"; fungiform papillae densities ranged from 33 to 156 papillae per square centimeter. Distribution of perceived bitterness of PROP and fungiform papillae density differed in women and men. The association between genetic taste measures and acceptance of sweet and high-fat groups differed in women and men. In women, liking of sweet and high-fat food and beverage groups decreased with increasing perceived bitterness of PROP. In men, liking of these foods and beverages increased but with increasing papillae densities. Genetic taste measures were not associated with a dislike of bitter food and beverage groups. APPLICATIONS: The influence of genetic variation in taste on food intake depends on how perceptible sweet, fat, or bitter components are in foods and beverages, as well as the value of sensory factors vs other factors (e.g., health, convenience) on personal dietary choices. Female supertasters of PROP bitterness may avoid high-fat or sweet foods because these oral sensations are too intense and thus less pleasant. Supertasters may taste more bitterness in vegetables but still enjoy eating them because of their healthfulness and because condiments (especially those that are salt based) can block bitterness. PMID- 10863569 TI - Intake and food sources of macronutrients among older Hispanic adults: association with ethnicity, acculturation, and length of residence in the United States. AB - OBJECTIVE: To describe the food intake and food sources of macronutrients in diets of older Hispanic adults in the Northeastern United States and to explore relationships between acculturation, years in the United States, and macronutrient intake. DESIGN: Cross-sectional study using a representative sample of older Hispanic adults and a comparison group of non-Hispanic whites. SUBJECTS/SETTING: Hispanic (n = 711) and non-Hispanic white (n = 226) persons, aged 60 years and older, residing in Massachusetts. STATISTICAL ANALYSIS: Macronutrient intakes, collected by 24-hour dietary recall, were compared across ethnic groups by means of the general linear models procedure (with Bonferroni adjustments). Associations between macronutrient intake and predictor variables were tested with Pearson correlations and linear regression. The contribution of foods to total intake of macronutrients was determined by use of a rank procedure. RESULTS: Hispanic elderly subjects consumed significantly less saturated fat and simple sugars and more complex carbohydrates than did non Hispanic whites. Hispanics residing in the United States for a longer time tended to have macronutrient profiles more similar to those of the non-Hispanic whites. Rice for Hispanic and bread for non-Hispanics were the major contributors of energy. More acculturated Hispanic elders consumed fewer ethnic foods and more foods related to the non-Hispanic-white eating patterns than those less acculturated. APPLICATIONS/CONCLUSIONS: Efforts to promote better diets among Hispanic elders need to emphasize maintenance or adoption of healthful dietary patterns based on ethnic and modern foods that will satisfy their biological, emotional, and social needs. Dietitians and other dietetics practitioners can use the information presented here in studying nutrition-related chronic diseases, in public health planning, and in nutrition education and promotion efforts directed to ethnic-specific, elderly Hispanic groups. PMID- 10863570 TI - Defining stage of change for lower-fat eating. AB - OBJECTIVE: To test the validity of staging methodology for dietary fat reduction by examining cognitive profiles of persons classified in these groups: precontemplation, lowerfat maintenance (< or = 30% of energy as fat), and higher fat maintenance (> 30% of energy as fat). DESIGN: Cross-sectional survey of a random sample of 491 women residing in Guelph, Ontario, Canada, recruited by telephone. SETTING/SUBJECTS: Mean age of subjects was 43.7 +/- 12.2 years. The majority (58%) lived with a spouse or partner and had completed high school (68%). STATISTICAL ANALYSES: Multivariate analysis of variance was used to compare the pros and cons of lower-fat eating, level of self-efficacy in avoiding high-fat foods, and use of 9 processes of change to support lower-fat eating habits in women assigned to the precontemplation, higher-fat, and lower-fat maintenance stages. RESULTS: When compared with subjects classified in the precontemplation stage, the 2 groups of subjects in the maintenance stage had higher ratings of the pros (49.7 +/- 9.5 vs 43.7 +/- 7.2, P < .05), lower ratings of the cons (47.2 +/- 8.2 vs 51.9 +/- 11.8, P < .05), higher self-efficacy scores, and more frequent use of processes of change than subjects classified in the precontemplation stage. No differences between women in the 2 maintenance groups were observed in self-efficacy; however, those in the lower-fat maintenance group reported lower cons than those in the higher-fat maintenance group (46.2 +/- 7.2 vs 48.2 +/- 9.1, P < .05) and more frequent use of all processes of change. CONCLUSIONS/APPLICATIONS: Stage of change for dietary fat reduction is a cognitive variable that provides insights into attitudes about and motivations to consume lower-fat foods. PMID- 10863572 TI - Caffeine and theobromine contents of ready-to-eat chocolate cereals. PMID- 10863568 TI - Responses of older adults to theory-based nutrition newsletters. AB - OBJECTIVE: To evaluate the effect of a theory-based newsletter on knowledge, attitude, and behavior change in older adults. DESIGN: Pretest-posttest, random assignment, and treatment-control design with 2 treatment groups: 1 that received newsletters only and 1 that received newsletters with follow-up telephone interviews. Control group completed pretest-posttest surveys only. SUBJECTS/SETTING: Four hundred eighty men and women, aged 60 to 74 years, were recruited to participate in a home-based educational intervention using a patient list generated from a rural tertiary care hospital database, Geisinger Medical Center in Danville, Pa. INTERVENTION: Five nutrition newsletters designed using the nutrition communication model and adult learning theory principles were mailed biweekly. Telephone interviews followed each of the 5 newsletters 10 to 14 days after distribution. OUTCOME MEASURES: Nutrition knowledge and interest, food behavior related to dietary fat, and stages of change for dietary fat and fiber. STATISTICAL ANALYSES PERFORMED: Analysis of covariance was used to determine group differences in posttest outcome measures using pretest as covariate. RESULTS: In addition to achieving higher scores than the control group, the treatment groups were significantly different from each other in correct and perceived nutrition knowledge at posttest. Those in the treatment group receiving telephone calls scored higher (mean change = 19.0% for correct and 20.3% for perceived) than those who received the newsletters only (mean change = 12.5% for correct and 14.3% for perceived; P < .05). Treatment groups also rated their interest in nutrition higher than the control group did; there was no between treatment difference. Treatment groups performed significantly better than the control group for dietary fiber stage of change (P < .05). Those receiving only newsletters scored significantly better than the control for the "avoid fat" food behavior (P < .05). APPLICATIONS/CONCLUSIONS: This study provides an example of the incorporation of a theoretical model in development and evaluation of newsletters. Home-delivered nutrition newsletters based on this model can communicate health and nutrition information to older adults. Consumers today have more opportunities than ever before to access nutrition information quickly and inexpensively. Newsletters can help dietetics professionals filter and limit what consumers must process, saving clients time and improving the accuracy of information obtained. Dietetics professionals in both clinical and community practice are uniquely positioned to provide highly focused and understandable information to consumers via a newsletter format. PMID- 10863571 TI - Nutritional complications and management of intestinal transplant. AB - Advances in intestinal transplantation provide a promising alternative to patients with intestinal failure and chronic dependence on total parenteral nutrition. However, many physiologic complications arising from the surgical procedure and high-dose immunosuppression, along with potential for rejection and infection, make successful graft function after transplantation a challenge. Nutrition issues unique to this patient population include recovery of normal intestinal motility and absorptive capacity. Diarrhea and high stomal output, which are common postoperatively, lead to deficits in macronutrients and micronutrients, especially electrolytes. Impaired gastrointestinal function affects ability to wean patients off hyperalimentation and enable them to tolerate nutrients enterally. In pediatric recipients of intestinal transplant, lack of experience with food or prior food aversions can lead to refusal to eat after transplant--additional challenges to achieving oral intake. Early and aggressive nutrition intervention is necessary for resolution of nutritional deficits and health of donor small bowel. This article presents an overview of the surgical procedure of intestinal transplantation and describes the physiologic adaptations that occur after the process. A case study demonstrates the clinical and nutritional hurdles associated with an intestinal transplant in a child and how dietitians can provide nutrition management. The potential role of individual nutrients in recovery of the transplanted bowel is also discussed. PMID- 10863573 TI - Expert panel identifies activities and performance measures for foodservice benchmarking. PMID- 10863574 TI - Time spent by schoolchildren to eat lunch. PMID- 10863575 TI - Marketing nutrition among urban Latinos: the SALUD! campaign. PMID- 10863576 TI - Availability of a la carte food items in junior and senior high schools: a needs assessment. PMID- 10863577 TI - Case problem: nutrition concerns related to the performance of a baseball team. PMID- 10863578 TI - Position of the American Dietetic Association and Dietitians of Canada: nutrition intervention in the care of persons with human immunodeficiency virus infection. AB - Human immunodeficiency virus (HIV) targets the immune system, making an infected person susceptible to infection and neoplasm because of an impaired ability to mount an adequate immune response. Malnutrition and its complications can further render an HIV-infected person susceptible to opportunistic infection and reduced effectiveness and tolerance to medications and other therapies. Though effective antiretroviral treatments have dramatically reduced death rates in the United States and Canada, HIV-infected people face a lifetime of vigilant polypharmacy to control HIV and associated complications. Malnutrition, various forms of tissue wasting, fat accumulation, increased lipid levels, and risk of additional chronic disease have become central issues in health care plans. It is the position of the American Dietetic Association and Dietitians of Canada that efforts to optimize nutritional status, including medical nutrition therapy and nutrition-related education, should be components of the total health care provided to people infected with human immunodeficiency virus HIV. PMID- 10863579 TI - Hunters hounded as leishmaniasis is diagnosed in foxhounds. PMID- 10863581 TI - AMDUCA regulates small and large animal practice. PMID- 10863583 TI - FoodNet reports decline in some foodborne disease. PMID- 10863582 TI - FDA rejects petition to ban rBST. PMID- 10863580 TI - New Jersey veterinarian acquitted of cruelty conviction. Superior court judge challenges reliability of PETA witness. PMID- 10863584 TI - Believes puppy mills should be eradicated. PMID- 10863585 TI - Quality of life in animals. PMID- 10863586 TI - What is your diagnosis? Soft-tissue mass outlined with a periosteal reaction overlying the frontal bone. Hematoma of the skull in a rhesus macaque. PMID- 10863587 TI - Theriogenology question of the month. Bacterial placentitis attributable to a gram-positive filamentous branching bacillus organism. PMID- 10863588 TI - Anesthesia case of the month. Diaphragmatic hernia making it difficult to ventilate a horse during anesthesia. PMID- 10863589 TI - Naturally occurring tuberculosis in white-tailed deer. AB - OBJECTIVE: To determine the distribution of lesions and extent of tissues infected with Mycobacterium bovis in a captive population of white-tailed deer. DESIGN: Cross-sectional study. ANIMALS: 116 captive white-tailed deer. PROCEDURE: Deer were euthanatized, and postmortem examinations were performed. Tissues with gross lesions suggestive of tuberculosis were collected for microscopic analysis and bacteriologic culture. Tissues from the head, thorax, and abdomen of deer with no gross lesions were pooled for bacteriologic culture. Tonsillar, nasal, oral, and rectal swab specimens, fecal samples, and samples of hay and pelleted feed, soil around feeding sites, and water from 2 natural ponds were collected for bacteriologic culture. RESULTS: Mycobacterium bovis was isolated from 14 of 116 (12%) deer; however, only 9 of 14 had lesions consistent with tuberculosis. Most commonly affected tissues included the medial retropharyngeal lymph node and lung. Five of 14 tuberculous deer had no gross lesions; however, M bovis was isolated from pooled tissue specimens from the heads of each of these deer. Bacteriologic culture of tonsillar swab specimens from 2 of the infected deer yielded M bovis. Mean (+/- SEM) age of tuberculous deer was 2.5 +/- 0.3 years (range, 0.5 to 6 years). Mycobacterium bovis was not isolated from feed, soil, water, or fecal samples. CONCLUSIONS AND CLINICAL RELEVANCE: Examination of hunter-killed white-tailed deer for tuberculosis commonly includes only the lymph nodes of the head. Results of such examinations may underestimate disease prevalence by as much as 57%. Such discrepancy should be considered when estimating disease prevalence. PMID- 10863590 TI - Sedative and physiologic effects of orally administered alpha 2-adrenoceptor agonists and ketamine in cats. AB - OBJECTIVE: To compare efficacy of 3 regimens of orally administered sedatives and determine physiologic effects of 1 of these regimens in healthy cats. DESIGN: Prospective randomized study. ANIMALS: 34 cats. PROCEDURE: Cats were assigned to 1 of 3 groups that were treated by oral administration of detomidine and ketamine, xylazine and ketamine, or medetomidine and ketamine. Cats were monitored for degree of sedation at 5-minute intervals for 60 minutes. Physiologic effects in cats treated with detomidine and ketamine were measured at 5-minute intervals for 30 minutes and compared with effects in cats treated i.m. with detomidine and ketamine or xylazine and ketamine. RESULTS: All cats treated orally with detomidine and ketamine became laterally recumbent; sedation was more variable in the other 2 groups treated orally. Vomiting and excessive salivation were the only adverse effects. Bradycardia (heart rate < 145 beats/min) was detected at each evaluation time in cats treated orally with detomidine and ketamine and in all cats treated i.m. Minimal differences among groups were detected for heart and respiratory rates, rectal temperature, and hemoglobin oxygen saturation. CONCLUSIONS AND CLINICAL RELEVANCE: Oral administration of detomidine and ketamine is an effective method of sedating healthy cats and induces minimal physiologic effects that are similar to those resulting from i.m. administration of sedatives. PMID- 10863591 TI - Positive Coombs' test results in two dogs treated with amiodarone. AB - Effects of amiodarone, an antiarrhythmic drug that is effective in suppressing severe ventricular arrhythmias that are refractory to other antiarrhythmic drugs, were evaluated in 2 dogs with cardiac disease. One dog was a Doberman Pinscher with cardiomyopathy that developed severe thrombocytopenia after receiving amiodarone for 7 months. The second was a Giant Schnauzer with acquired mitral valve degeneration that developed regenerative anemia after receiving amiodarone for 5 months. Results of direct Coombs' tests were positive in both dogs. Adverse effects of amiodarone are numerous; in dogs, the most common adverse effects are anorexia and hepatotoxicosis. Frequent CBC and serum biochemical analyses should be performed when amiodarone is administered with the intent of continuing the drug indefinitely. PMID- 10863592 TI - 5-Hydroxytryptophan toxicosis in dogs: 21 cases (1989-1999). AB - OBJECTIVE: To determine epidemiologic characteristics, clinical findings, and treatment outcome of 5-hydroxytryptophan (5-HTP) toxicosis in dogs. DESIGN: Retrospective study. ANIMALS: 21 dogs with evidence of accidental 5-HTP ingestion. PROCEDURE: Information was retrieved from the National Animal Poison Control Center database. Records of dogs ingesting 5-HTP between January 1989 and February 1999 were reviewed for information on signalment, dose ingested, clinical signs (onset, severity, duration), treatments administered, and outcome. RESULTS: Clinical signs of toxicosis developed in 19 of 21 (90%) dogs. Neurologic signs included seizures (9 dogs), depression (6), tremors (5), hyperesthesia (5), and ataxia (4). Gastrointestinal tract signs included vomiting or diarrhea (12 dogs), signs of abdominal pain (3), and hypersalivation (2). Other clinical signs were hyperthermia (7 dogs) and transient blindness (3). Three dogs died. No important clinical laboratory or necropsy findings were reported. The doses of 5 HTP ingested ranged from 2.5 to 573 mg/kg (1.1 to 260 mg/lb) of body weight; the minimum toxic dose reported in our study was 23.6 mg/kg (10.7 mg/lb), and the minimum lethal dose was 128 mg/kg (58.1 mg/lb). Onset of signs ranged from 10 minutes to 4 hours after ingestion, and signs lasted up to 36 hours. Of 17 dogs with clinical signs of toxicosis that received treatment, 16 recovered; treatment consisted of decontamination, seizure control, thermoregulation, fluid therapy, and supportive care. CONCLUSIONS AND CLINICAL RELEVANCE: Ingestion of 5-HTP in dogs can result in a potentially life-threatening syndrome resembling serotonin syndrome in humans, which requires prompt and aggressive care. PMID- 10863593 TI - Bufo marinus intoxication in dogs: 94 cases (1997-1998). AB - OBJECTIVE: To determine history, clinical and electrocardiographic abnormalities, treatment, and outcome of dogs exposed to toxins produced by the Bufo marinus toad. DESIGN: Retrospective study. ANIMALS: 94 dogs. PROCEDURE: Medical records of dogs examined between July 1997 and July 1998 for which a diagnosis of toad intoxication had been made on the basis of history and physical examination findings were reviewed. RESULTS: Most (66) dogs were treated during the spring and summer. For 54 dogs, exposure to toads had been witnessed. For the remaining 40, toad intoxication was diagnosed on the basis of history and clinical signs. The most common clinical signs were neurologic abnormalities, hyperemic mucous membranes, ptyalism, recumbency or collapse, tachypnea, and vomiting. The oral cavity was lavaged with tap water in all dogs. Fifty-two dogs were hospitalized for treatment. Body weight of dogs hospitalized > 2 hours was significantly less than that of dogs treated as outpatients. The most common electrocardiographic findings were sinus arrhythmia, sinus tachycardia, and normal sinus rhythm. Eighty-nine dogs recovered fully, 4 died, and 1 was euthanatized. CONCLUSIONS AND CLINICAL RELEVANCE: In areas in which B marinus toads are endemic, toad intoxication should be considered in the differential diagnosis for dogs with an acute onset of neurologic abnormalities, hyperemic mucous membranes, and ptyalism, especially during the spring and summer months. The prognosis is good for dogs with toad intoxication that receive appropriate treatment. PMID- 10863594 TI - Clinical, bacteriologic, serologic, and pathologic features of infections with atypical Taylorella equigenitalis in mares. AB - OBJECTIVE: To characterize clinical, serologic, bacteriologic, cytologic, and pathologic endometrial responses of mares to 2 donkey-origin atypical bacterial isolates resembling Taylorella equigenitalis. DESIGN: Prospective in vivo study. ANIMALS: 10 healthy mares. PROCEDURE: Mares in estrus (2/group) were inoculated by intrauterine infusion with 2 isolates of classic T equigenitalis or 2 isolates of atypical Taylorella sp or were sham-inoculated. Bacteriologic, serologic, clinical, uterine, cytologic, and pathologic endometrial responses were assessed 4, 11, 21, 35, and 63 days after inoculation and on day 111 in mares with positive culture results on day 63. RESULTS: One atypical isolate failed to cause infection. The second atypical isolate and both classic T equigenitalis isolates induced similar transient metritis and cervicitis. Both classic isolates and 1 atypical isolate induced anti-T equigenitalis complement-fixing antibodies detectable at day 11. Classic isolates and an atypical isolate provoked intense neutrophilic endometritis followed by a resolving, subacute, neutrophilic mononuclear endometrial response. The atypical isolate and classic isolates were recovered from the uterus, clitoral fossa, or clitoral sinus of one or both exposed mares for as long as 111 days. CONCLUSIONS AND CLINICAL RELEVANCE: Atypical Taylorella sp infections should be considered as a differential diagnosis of equine infertility in US-origin mares, even those not exposed to stallions from countries where contagious equine metritis occurs. The origins and prevalence of atypical Taylorella sp infection in US horses and donkeys are undetermined. PMID- 10863595 TI - Long-term outcome of horses with a slab fracture of the central or third tarsal bone treated conservatively: 25 cases (1976-1993). AB - OBJECTIVE: To determine clinical features of horses with a slab fracture of the central or third tarsal bone and to report outcome of horses in which treatment did not include surgery. DESIGN: Retrospective study. ANIMALS: 25 horses (14 Standardbreds, 6 Thoroughbreds, 5 Quarter Horses). PROCEDURE: Medical records of horses with a slab fracture of the central (n = 9) or third (16) tarsal bone were reviewed. Only horses for which treatment consisted of confinement to a stall were included in this study. Clinical features and radiographic findings were recorded and summarized. Outcome was determined for racing breeds by obtaining official lifetime race results. Outcome for Quarter Horses was determined by phone survey of the owners. RESULTS: 16 (64%) horses had a successful outcome. Ten of 14 (71%) Standardbreds and 2 of 6 Thoroughbreds returned to racing and started at least 5 races after injury. Four of 5 Quarter Horses for which follow up information was available successfully returned to their previous activity. Sex, age, limb affected, or gait was not associated with final outcome. Percentage of racehorses with central tarsal bone fractures that had a successful outcome (2/7) was significantly less than percentage with third tarsal bone fractures that did (10/13). CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that enforced rest without surgical fixation can be an effective therapeutic option for horses with a slab fracture of the central or third tarsal bone, even if athletic function is expected. PMID- 10863596 TI - Pneumothorax in horses: 40 cases (1980-1997). AB - OBJECTIVE: To characterize pneumothorax in horses and to describe clinical signs, diagnostic testing, and clinical outcome of horses with pneumothorax. DESIGN: Retrospective study. ANIMALS: 40 horses. PROCEDURE: Medical records of horses with pneumothorax were reviewed to obtain information on signalment, history, clinical signs, diagnostic testing, treatment, and clinical outcome. RESULTS: Horses developed pneumothorax secondary to pleuropneumonia (17 horses), open wounds of the thorax (9), closed trauma to the thorax (7), surgery on the upper portion of the respiratory tract (3), and surgery involving the thoracic cavity (1); 3 horses had pneumothorax of unknown cause. Clinical signs included tachypnea, dyspnea, cyanosis, lack of lung sounds on auscultation of the dorsal aspect of the thorax, fever, tachycardia, signs of depression or anxiousness, and cough. Radiography and ultrasonography were useful to definitively diagnose pneumothorax. Pneumothorax was bilateral in 47.5% (19/40) and unilateral in 42.5% (17/40) of horses; designation of unilateral versus bilateral was not recorded in the remaining 4 horses. Horses with pneumothorax secondary to pleuropneumonia more commonly had unilateral pneumothorax (64.7% for unilateral vs 29.4% for bilateral; not specified for 1 horse). Horses with pneumothorax secondary to pleuropneumonia were less likely to survive than horses with pneumothorax secondary to other causes (35.3 vs 69.6% survived, respectively). CONCLUSIONS AND CLINICAL RELEVANCE: Pleuropneumonia is an important cause of pneumothorax in horses. Classic clinical signs of pneumothorax may not be evident. Radiography, ultrasonography, or both may be required for diagnosis. Prognosis for survival is better for horses with pneumothorax not associated with pleuropneumonia. PMID- 10863597 TI - Influence of the Milk and Dairy Beef Quality Assurance Program on dairy farm drug management practices. AB - OBJECTIVE: To test the hypothesis that dairy farms certified in the Milk and Dairy Beef Quality Assurance Program (QAP) were more likely to use prudent drug management practices than farms that were not certified. DESIGN: Cross-sectional study. SAMPLE POPULATION: 141 Michigan dairy farms of which 74 were not certified in the QAP, 30 were involuntarily certified, and 37 were voluntarily certified. PROCEDURE: Dairy producers completed a self-administered questionnaire that focused on herd health management, drug use, record keeping, personnel management, and descriptive characteristics of their farm during 1993. Separate multivariable logistic regression models were developed to determine the association of QAP certification with each of the management practices. RESULTS: Results suggested that farms adopted specific management practices irrespective of certification. Many farms used visible identification and non-emergency veterinary services and discussed residue prevention with employees. Involuntary certification was associated with maintenance of good written treatment records and performance of on-farm drug residue testing. Voluntary certification was weakly associated with use of refrigerated drug storage. CONCLUSIONS AND CLINICAL RELEVANCE: QAP certification appeared to have been associated with the adoption of only a few prudent drug use practices, although QAP materials and framework were developed to assist veterinarians in the promotion of disease prevention, client communication, and residue prevention practices on farms. Veterinary care would benefit from the development and encouragement of better record keeping on farms. PMID- 10863599 TI - Physiologic and pathophysiologic effects of natriuretic peptides and their implications in cardiopulmonary disease. PMID- 10863600 TI - Nonimpact brain injury: neuropsychological and behavioral correlates with consideration of physiological findings. AB - This retrospective clinical study investigated the neuropsychological, physiological, and behavioral functioning of 32 adult outpatients up to 65 months following nonimpact brain injury (i.e., whiplash). All participants were administered a flexible battery of cognitive tests, and some underwent neurodiagnostic procedures and sleep studies. Compared with published norms, neuropsychological data revealed significant and persistent age-adjusted cognitive deficits, primarily in the area of executive functioning. Participants frequently complained of problems with behavioral control, sleep, and sexuality. Although structural neuroimaging was not sensitive in detecting brain pathology, quantitative electroencephalography was abnormal in all the participants evaluated, showing frontocentral slowing and increased spike wave activity. We propose that whiplash injury can produce wide-ranging circuitry dysfunction and that test selection is critical in identifying cognitive deficits. PMID- 10863598 TI - Relative cost-effectiveness of treatment of feedlot calves with ivermectin versus treatment with a combination of fenbendazole, permethrin, and fenthion. AB - OBJECTIVE: To compare growth performance, animal health characteristics, and carcass characteristics of feedlot calves treated with ivermectin topically with that of feedlot calves treated with a combination of fenbendazole orally and permethrin and fenthion topically. DESIGN: Clinical trial. ANIMALS: 14,184 British crossbred steer calves (mean weight, 286 kg [630 lb]) in 30 pens at a commercial feedlot in Nebraska. PROCEDURE: On arrival at the feedlot, calves were randomly assigned to be treated with ivermectin topically or with a combination of fenbendazole orally and permethrin and fenthion topically (control). At the time of assignment to treatment groups, fecal samples were collected from 5% of the calves. Growth performance, carcass characteristics, and health information were recorded. RESULTS: Geometric mean fecal egg counts at the time of arrival at the feedlot were not significantly different between groups. Final weight, weight gain, average daily gain, and the dry matter intake-to-gain ratio were significantly improved for calves in the ivermectin group. The percentage of carcasses classified as quality grade choice was higher for the ivermectin group than the control group; however, the percentage of carcasses classified as yield grade 1 and the dressing percentage were higher for the control group than for the ivermectin group. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that topical administration of ivermectin to feedlot calves is relatively more cost effective than administration of a combination of fenbendazole orally and permethrin and fenthion topically. PMID- 10863601 TI - Successful mastery with a cognitive orthotic in people with traumatic brain injury. AB - A multifunction cognitive orthotic (CO), designed for individuals with significant deficits from traumatic brain injury, enables rapid unassisted reliable performance of a targeted task and facilitates transfer of training across subsequent activity tasks for most participants. Consistency in design ensures consistent cues and organization as well as unchanging procedural format as content and tasks vary. Of the 41 participants provided a clinical assessment trial with a CO, 36 (88%) demonstrated rapid achievement of success on the initial task. Thereafter, transfer of training was achieved across subsequent targeted activity tasks by changing only the content, in the context of stable CO design. Many individuals with brain injury can be self-sufficient in performing, unassisted, the essential steps of targeted activities when provided a properly designed compensatory assistive device. PMID- 10863602 TI - Postconcussional symptoms in chronic back pain. AB - Checklist measures of persisting postconcussional and control symptom change (postinjury-preinjury self-ratings on 5-point scales) were compared among selected groups of consecutive referrals with traumatic back pain and concussion. Mean symptom change was not significantly different between the 2 groups. Positive correlations were found between postconcussional symptom change and psychometric measures of emotional distress for participants with valid Minnesota Multiphasic Personality Inventory profiles in the 2 groups combined. Results suggested that persisting postconcussional symptoms were not specific to concussion but were associated with increased emotional distress. PMID- 10863603 TI - Predicting cognitive performance in alcoholics and nonalcoholics: specification of affective, childhood behavior disorders, and antisocial variables. AB - Glenn, Errico, Parsons, King, and Nixon (1993) reported that composite indexes of childhood behavior disorders, current affective distress, and lifetime antisocial behaviors predicted different aspects of neurocognitive functioning in abstinent male and female alcoholics and in peer nonalcoholics. To make the results more pertinent to clinical assessment, we have: (a) identified the components of the composite indexes that were the best predictors, (b) added new data predicting overall impairment, and (c) conducted within-group male and female comparisons. In alcoholics, the depressive symptoms component of the affective distress composite index predicted set-shifting and overall impairment scores; the childhood attention deficit disorder behaviors component of the childhood behavior disorders composite predicted lower verbal performance. For nonalcoholics, childhood attention disorder behaviors predicted verbal, visuospatial, set-shifting, and overall impairment scores. Results for men and women were generally consistent with overall group analyses. The results have implications for cognitive assessment in alcoholic and nonalcoholic persons. PMID- 10863605 TI - Short-form prediction of WAIS-R scores in a sample of individuals diagnosed with multiple sclerosis. AB - A short form of the Wechsler Adult Intelligence Scale--Revised (WAIS-R) developed by Ward (WAIS-R/7 SF; 1990) was used to generate Verbal, Performance, and Full Scale IQ scores (VIQ, PIQ, and FSIQ, respectively) in 66 individuals diagnosed with multiple sclerosis (MS). Short-form scores were highly correlated with WAIS R scores. However, the short-form VIQ and PIQ, but not FSIQ, scores differed significantly from corresponding WAIS-R scores. WAIS-R/7 SF VIQ, PIQ, and FSIQ scores fell within 5, 9, and 6 absolute error points, respectively, of corresponding WAIS-R IQ scores in 95% of cases. Classification of IQ scores into ranges (e.g., average, high average, etc.) based on the scheme outlined by Wechsler (1981) was consistent between WAIS-R/7 SF and WAIS-R scores in 81.8% (for VIQ), 74.8% (for PIQ), and 89.4% (for FSIQ) of cases. These findings are discussed within the context of using the WAIS-R/7 SF in the assessment of MS patients. PMID- 10863604 TI - Neuropsychological deficit profiles in systemic lupus erythematosus. AB - Although neuropsychological deficits have been reported in several cognitive domains in patients with systemic lupus erythematosus (SLE), there is considerable variability in the literature about which neuropsychological domains are most affected. Similar to studies that demonstrated that specific profiles of neuropsychological deficits exist for those with traumatic brain injury (TBI; Johnstone, Hexum, & Ashkanazi, 1995), this study examined whether a specific pattern of deficits is present in SLE. By comparing reading scores (as estimates of premorbid ability) to tests of concurrent cognitive abilities (i.e., memory, attention, etc.), it was determined that SLE presents a profile distinct from TBI, with the most significant impairments noted in expressive language (Zdiff = 1.39), attention (Zdiff = -0.41), and speed of processing (Zdiff = -0.40). In contrast to TBI, no impairment was noted in intelligence, memory, or cognitive flexibility. Results suggest that memory problems reported by individuals with SLE may be related to inattention. Clinical implications are discussed. PMID- 10863606 TI - Internal consistency and concurrent validity of two short forms of the Visual Form Discrimination Test. AB - The purpose of this study was to evaluate the reliability and concurrent validity of 2 short forms of the Visual Form Discrimination Test, referred to as first half (FH) and front-back (FB). Participants were a mixed sample of 225 patients seen for neuropsychological evaluations. The mean difference score between both short forms and the full form was less than 1 point. The short-total correlations were .85 and .86 for the FH and FB forms, respectively. A binary clinical decision rule, used to classify patients as normal or impaired, resulted in a 93.3% correct and a 94.7% correct classification rate for the FH and FB forms, respectively. It is concluded that the short form of the test, used in conjunction with a clinical decision rule, results in a very minor loss of accuracy. PMID- 10863607 TI - Assessment of incomplete effort with the California Verbal Learning Test. AB - The false-positive rates of 2 formulas for the detection of incomplete effort on the California Verbal Learning Test (CVLT; Delis, Kramer, Kaplan, & Ober, 1987) were evaluated in a sample of 134 patients with traumatic head injury (THI) who had been carefully screened to eliminate any individuals who might be expected to have financial incentives for poor performance. One formula incorrectly identified only 7.46% of the sample as providing incomplete effort, confirming its potential usefulness in clinical settings. The other formula fared much worse, misidentifying an unacceptably high 18.66% of the sample. Lower levels of education were found to increase the likelihood of false-positive results with that formula. It is concluded that the CVLT holds promise for the evaluation of incomplete effort after THI but only within the context of a more comprehensive neuropsychological evaluation. PMID- 10863608 TI - [Epidemiologic and endoscopic aspects of Helicobacter pylori infection in Vientiane, Laos]. AB - From January 1996 to March 1997, Helicobacter pylori infection was studied prospectively for 200 consecutive patients presenting with epigastric pain. 29 of them were excluded. Bacteriological (Gram, CLO-test) and histological methods were used to identify the bacteria from the gastric mycosis samples. Overall prevalence of infection was 68.4% with no differences according to age and sex; infection was more frequent for patients with duodenal ulcers (80%) than for those with normal endoscopical aspect (60.6%). PMID- 10863609 TI - [Isolation of enteric pathogeic agents in Cote d'Ivoire: Escherichia coli O157:H7 and enteroaggregative E. coli]. AB - New pathogens including Escherichia coli O157:H7 have emerged and spread world wide as the most important cause of foodborne infections. We established a prospective study in Abidjan from 1996 to 1999 to determine the prevalence of Shiga-toxin producing E. coli (STEC) in our environment. Two O157 strains were found. One (EA47) O157:H7 was isolated from chicken and the other (EH144) O157:HNM from human diarrhoeal stool specimens. Both O157 strains carried stx2, eae, and UidA genes, but not e-hly one. Four other pathogenic E. coli were isolated, including three enteroaggregative E. coli (EAggEC) and one isolate which expresses a cytolethal distending toxin gene (cdtB). This is the first report of Shiga-toxin producing E. coli (STEC) in Cote d'Ivoire. Given its low prevalence (0.8%), E. Coli does not appear to be a public health problem in Cote d'Ivoire. PMID- 10863610 TI - [Improvement of tuberculosis diagnosis by the Mycobacteria Growth Indicator Tube (MGIT) in a developing country laboratory]. AB - In order to improve tuberculosis diagnosis in a developing country (Senegal), we evaluated a new liquid-based medium and nonradioactive system, Mycobacteria Growth Indicator Tube (MGIT), with individual clinical specimens collected in Dakar. The main purpose was to compare the time to detection and the rate of recovery of Mycobacterium tuberculosis complex and to determine its importance with respect to Lowenstein-Jensen (LJ), a liquid-based-medium for isolation of M. tuberculosis complex. 531 specimens were processed with Mycoprep kit containing NaOH-N-acetyl L-cystein and inoculated on both LJ and MGIT and incubated at 37 degrees C for 60 days. For each medium, the recovery rate and the time to detection were recorded. Among the 531 specimens, of which 121 smears were positive, 32.5% (173/531) grew the M. tuberculosis complex. Of these, 103 were smear positive (S+) and 70 smear negative (S-). LJ recovered 54.9% (95/173) and MGIT recovered 91.9% (159/173). Disagreements were observed with 92 isolates, LJ failed to recover 78 while MGIT failed to recover 14. The overall mean time to detection was 20.1 days for LJ and 10.5 days for MGIT. MGIT has shown a better sensitivity in isolation with significant reduction in reporting culture for M. tuberculosis complex. As a simple and a nonradiometric system, it could be used in conjunction with egg-based media in developing countries laboratories. PMID- 10863612 TI - [Compared characteristics of peripheral facial paralysis according to HIV status in Bobo-Dioulasso (Burkina Faso)]. AB - Facial paralysis is a well-described manifestation of HIV infection. We report 27 cases of peripheral facial paralysis observed at Bobo-Dioulasso Hospital in a prospective study over a period of 9 months: 55 of the cases were HIV positive and 12/15 (80%) were in the 20-39 age group. Nine out of 11 females and 6 out of 16 males were seropositive. 13 of the cases were at stage B of CDC classification and 2 at stage C. ESR was elevated in all the HIV patients. CSF examination revealed lymphocytic pleiocytosis, elevated proteins and a positive HIV serology. CD4 counts were obtained in 8 cases and were under 400/mm3 in 4 cases. The clinical presentation was more severe in HIV seropositives with a longer duration of symptoms. Isolated peripheral facial paralysis associated with an elevated ESR in young adults suggest HIV infection and should lead to HIV counselling and testing. PMID- 10863611 TI - [Epidemiologic and parasitologic data concerning sporadic cutaneous leishmaniasis in northern Tunisia]. AB - This study refers to 23 patients presenting with the sporadic forms of cutaneous leishmaniasis encountered in northern most humid parts of Tunisia. Culture inoculation for parasitic isolation was processed using two media: the classical NNN and a rabbit serum based medium (SLC). Cultures were positive in 17 cases with SLC medium and 13 cases with NNN medium. Eight isolates were typed using 15 isoenzymes systems. Six isolates were identified as Leishmania infantum MON-24 which confirms the crucial role of this zymodeme in causing this form of cutaneous leishmaniasis. The other two isolates were identified as Leishmania infantum MON-1, which is the principal agent of visceral leishmaniasis in the Mediterranean area. PMID- 10863613 TI - [Subacute sclerosing panencephalitis. Study of 32 cases observed in Conakry, Guinea]. AB - Thirty two cases of subacute sclerosing panencephalitis were reported. Diagnosis was based on epidemiological, clinical and electroencephalographic data; myoclonies and alterations of intellectual functions were the most frequent symptoms. PMID- 10863614 TI - [Prolactinomas in Yaounde: analytical study of 36 consecutive cases followed in the internal medicine department of the Yaounde Hospital from 1990 to 1996]. AB - We analysed the epidemiological, clinical, biological, morphological and therapeutic characteristics of 36 cases of pituitary prolactinomas in Yaounde, Cameroun. Diagnosis was made on the basis of neuro-ophthalmological, gynaecological and sexual symptoms associated with quantity determinations of prolactinemia, total testosterone and blood oestrogen, folliculo stimulating hormone and luteining hormone. Expected levels of prolactinemia are above 150 micrograms/l. Cerebral tomodensitometry and, where possible, magnetic resonance imaging were used. Among the 24 patients on whom a tomodensitometry was performed, 4 male patients presented macroprolactinomas; the remaining 20 patients--mostly female--had microprolactinomas. Macroprolactinomas were found in male patients only, leading to an acute ophthalmological emergency for 2 of them who were operated in Paris-France. All the patients were put on bromocriptine; this molecule has antisecretory and antiproliferative properties, which are very useful in Africa, since surgery is very expensive. Cases of resistance to the molecule exist and new dopaminergic agonists are not yet being used in Cameroon. PMID- 10863615 TI - [Clinical trial of amodiaquine in the community of Attecoube (Abidjan, Cote d'Ivoire)(May-December 1955)]. AB - A prospective study in the municipality of Attecoube (Abidjan, Cote d'Ivoire) evaluated the sensitivity of P. falciparum to amodiaquine with a posology of 35 mg/kg over 3 days (1st day: 15 mg/kg; 2nd day: 10 mg/kg; 3rd day: 10 mg/kg) as well as its tolerance of this dosage. One hundred five WHO in vivo standard tests were performed over 7 days on subjects aged > 15 years from May to December 1995. The subjects were carriers of varying number of trophozoites: between 1000 to 34,000 trophozoites were recorded with a mean of 5193 trophozoites by microliter. We divided the subjects into two groups: group A with 43 patients to whom we administered medication and group B with 62 subjects who took their medication on their own. Clinical and parasitological verifications were made on D0, D2 and D7. Biological verification was conducted for 31 subjects of group A by mean of SGOT and SGPT quantity determination on D0 and D2. This survey revealed that 1.9% of P. falciparum malaria patients had precocious therapeutic failure to amodiaquine (35 mg/kg over 3 days) in this area. Clinical and biological tolerance was good and there was no difference between the two groups. We suggest that amodiaquine might be used for uncomplicated malaria at first intention in Abidjan. PMID- 10863616 TI - [Malaria in Vietnam: what is the awareness of risk for travel in 2000?]. AB - Increasing numbers of people are travelling to Vietnam. From december 1st 1998 to april 31 1999, we surveyed by questionnaire 191 travellers who consulted at health centres attached to French diplomatic representations (Hanoi, Ho-Chi-Minh Ville) in order to evaluate their prophylaxis practices with regard to malaria; 59% of these travellers were taking no preventive measures whatsoever, while the rest were following an often ill-adapted treatment. PMID- 10863617 TI - [Missed vaccination opportunities in Brazzaville]. AB - In order to assess the magnitude of missed immunisation opportunities, a study was carried out at ten randomly selected health centres in Brazzaville (Congo). A survey based on interviews at discharge was conducted with 306 mothers or caretakers of children aged under 2 years. The overall rate of missed immunization opportunities was 12.8% for children and 50.6% for women. The reasons most frequently given were illness of the child, misinformation and unavailable vaccines. PMID- 10863618 TI - [Survey of measles vaccine coverage in Brazzaville]. AB - Measles is an infectious disease that continues to be a significant cause of morbidity and mortality among children in Brazzaville. A measles vaccination coverage survey was conducted for children aged 9 to 23 months. A standard EPI cluster sample was applied in two areas: urban and peri-urban. Measles coverage of children after the vaccination campaign according to history ranged from 36.6% in urban to 38.6% in peri-urban areas, compared with 34.5 and 42.8% for routine vaccination in the same areas. The overall rate of measles coverage was 75.4%. The mean age of children was 46 weeks. PMID- 10863619 TI - [Vectorial competence of non-teneral Glossina morsitans morsitans (Mall strain) flies infected by Trypanosoma (Nannomonas) congolense IL 1180]. AB - Non-teneral tsetse flies of Glossina morsitans morsitans (strain Mall) about 16 days old were fed, once, on a rat infected by Trypanosoma congolense IL 1180. The global vectorial competence (VC) of these flies was appraised at 0.1035. VC in males was more important than for females. Infection by mesoprocyclic index was greater in female flies than in male ones, whereas for metacyclic index the reverse was true. This work shows that the age limits, but does not impede metacyclogenesis of non-teneral tsetse flies of G. m. morsitans (strain Mall). PMID- 10863620 TI - [List of Phlebotominae (Diptera:Psychodidae) of Iran]. AB - An alphabetical list of the Phlebotominae of Iran is provided, with 31 species of Phlebotomus and 23 species of Sergentomyia The list includes new species and regional characteristics as yet unrecorded by science. PMID- 10863621 TI - [Impact of pyrethrin resistance on the efficacity of impregnated mosquito nets in the prevention of malaria: results of tests in experimental cases with deltamethrin SC]. AB - The effects of impregnated bednets treated with deltamethrin at a dosage of 25 mg a.i./m2 were evaluated at two testing stations in Cote d'Ivoire. The first one was located in Yaokoffikro, where Anopheles gambiae s.s. are resistant to pyrethroids (including deltamethrin), and the second in M'be, close to a large rice-growing area where An. gambiae s.s. are susceptible pyrethroids. In both situations, treating bednets with deltamethrin was very effective in limiting contact between man and vector. 72% fewer female An. gambiae entered the huts in the susceptible area, whereas a decrease of 43% was recorded in the resistant area, indicating that deltamethrin still has a certain repellent effect on resistant populations of An. gambiae s.s. Overall mortality induced by bednets treated with insecticide was significantly higher in the resistant area (56.4% as versus 44.3%). An explanation for this apparent paradox is that the mosquitoes being less repelled by the insecticide remain on the treated material for longer periods of time and most of them eventually die. The results of this study indicate that bednets treated with deltamethrin are an effective prophylactic measure even in areas where An. gambiae s.s. are resistant to pyrethroids and should still be considered as a practical means of personal protection against malaria even in pyrethroid resistance areas with high frequency of kdr resistance genes. PMID- 10863622 TI - Shoulder dystocia: lessons from the past and emerging concepts. PMID- 10863623 TI - Obstetrical brachial palsy: pathogenesis, risk factors, and prevention. PMID- 10863624 TI - Can fetal-pelvic disproportion be predicted. PMID- 10863625 TI - Shoulder dystocia: risk identification. PMID- 10863626 TI - Fetal macrosomia: etiologic factors. AB - Fetal growth can be considered the outcome of an interaction between the genetic cause of growth and constraints provided by limitations on substrate availability (selected amino acids, free fatty acids, and mainly glucose). It should be noted that the majority of large infants are constitutionally large and do not require special intervention, which will result in adverse perinatal outcome. Efforts should be directed to the accelerated (pathologic) overgrown fetus and to methods of primary prevention of this abnormality by appropriate management approaches for the mother and fetus. PMID- 10863627 TI - Macrosomia: a genetic perspective. PMID- 10863628 TI - Sonographic and clinical methods in the diagnosis of macrosomia. AB - Receiver operator characteristic curves for both clinical and sonographic predictions of macrosomia subsume areas between 0.81 and 0.95, significantly larger than the area of 0.5 that indicates a useless test. Thus, these tests are defined as useful from a statistical point of view. Prediction of macrosomia by clinical or imaging techniques, however, is limited by the substantial false positive and false-negative rates inherent in these tests. We recommend that physicians continue to use clinical methods to estimate fetal weight, including asking women with parity to provide their own estimates. We recognize that the relative error associated with clinical or sonographic estimates of fetal weight limits their use in clinical practice. Sonographic laboratories may improve their results by performing ROC curve analysis on their own data and by selecting cutoff values that best predict macrosomia in their setting. Serial sonographic measurements that are above the limits chosen to define macrosomia increase the likelihood that a birth weight will be macrosomic. Separate ROC curves must be generated for twins and breech presentations and for patients with diabetes to answer weight-related clinical questions such as mode and timing of delivery. Three-dimensional ultrasound and magnetic resonance imaging are expected to generate ROC curves for estimates of fetal weight that are better than those for two-dimensional ultrasound or clinical estimates. Such analyses have yet to be published. PMID- 10863629 TI - Prediction of fetal weight with the use of three-dimensional ultrasonography. PMID- 10863630 TI - Sonography, suspected macrosomia, and prophylactic cesarean: a limited partnership. PMID- 10863631 TI - Maternal complications of fetal macrosomia. PMID- 10863632 TI - The neonate with macrosomia. PMID- 10863633 TI - Natural history of cervical neoplasia: defining progression and its consequence. PMID- 10863634 TI - Human papillomavirus infection and cervical carcinoma. PMID- 10863635 TI - Clinical treatment of women with atypical squamous cells of undetermined significance or atypical glandular cells of undetermined significance cervical cytology. PMID- 10863636 TI - Management of high-grade cervical intraepithelial neoplasia and low-grade squamous intraepithelial lesion and potential complications. PMID- 10863637 TI - New technology in Papanicolaou smear processing. PMID- 10863638 TI - Human papillomavirus. Introduction. PMID- 10863639 TI - Human papillomavirus. Epidemiology, transmission, and pathogenesis. AB - Like many viruses, HPV is a fascinating organism. It is extremely difficult to grow in vitro; but once an individual becomes infected with HPV, it can be difficult or even impossible to eradicate. HPV is associated with mild to moderate disease that even in the absence of therapy may spontaneously regress. On the other hand, some HPV infections progress to cancer, which can be fatal if treatment is delayed. The reader is invited to learn more about the aspects of the biology, diagnosis, and treatment of HPV in the articles that follow. PMID- 10863640 TI - Human papillomavirus in cervical neoplasia. Role, risk factors, and implications. AB - A rapidly increasing body of evidence links human papillomavirus (HPV) to cervical neoplasia through epidemiologic associations, pathologic features, molecular detection, and mechanisms of oncogenesis. HPV is now accepted as the primary cause of cervical neoplasia and accounts for most of the risk factors traditionally associated with this disease. The role that HPV plays in the induction and progression of cervical neoplasia is becoming clearer; however, the challenge now is to find other factors that may play a role in the outcome of cervical cancer and that might serve as targets for novel treatments. PMID- 10863641 TI - Diagnosis of human papillomavirus gynecologic infections. AB - The identification in the early 1980s of human papillomavirus (HPV) DNA in cervical carcinoma generated interest in molecular classification of the virus, and prompted studies regarding the oncogenic potential of genital HPVs. Subsequent studies confirming the presence of HPV in greater than 90% of precancerous cervical lesions and close to 100% of cervical cancers has raised concerns regarding the adequacy of Papanicolaou (Pap) smear testing for the detection of precancerous lesions/HPV infection. A variety of detection methods adjunctive to cytologic testing have been described, including detection at the macroscopic level, cerviography, colposcopy, and serologic and molecular-based HPV testing. Recently, there has been intense interest in molecular-based detection and typing of HPV-induced genital lesions. This has resulted in the development of a variety of molecular-based detection methods including Southern transfer, dot blotting, in situ hybridization, hybrid capture, and PCR-based assays. This article provides an overview of each of the molecular methods, and addresses the potential future role of molecular-based HPV testing. PMID- 10863642 TI - In situ diagnosis of human papillomaviruses. AB - In situ hybridization (ISH) is the demonstration of specific genetic information within a morphologic context. For HPV, colorimetric ISH has the advantage that it can be applied to routine formalin-fixed paraffin embedded tissues. This conserves patient material and permits histologic selection of optimal material for testing. ISH allows for precise spatial localization of viral sequences within tissues. ISH also allows the integration status of HPV to be determined. The major limitations of the method are the potential for error in HPV typing because of probe cross-hybridization and relatively low sensitivity if the method is not fully optimized. PMID- 10863643 TI - Evaluation of abnormal cervical cytology. AB - The introduction of widespread cervical cytologic screening in the United States and in many other developed countries is clearly one of the most dramatic preventive health measures in history. This article provides a history of cervical cancer screening and a comprehensive review of management options and techniques. PMID- 10863644 TI - Statistical issues in human papillomavirus testing and screening. AB - As discussed in the preceding sections, regardless of the research purpose, etiology or prevention, there are critical statistical and study design issues related to measurement of HPV infection status and its cervical lesion outcomes that need to be considered for the appropriate interpretation of the association of HPV infection and its determinants with cervical cancer in epidemiologic studies, and of screening effectiveness by HPV testing in clinical and intervention trials. These statistical issues may affect the validity of epidemiologic and screening studies and have led to inconsistent results in the literature. Meticulous attention to study design and laboratory detection issues helps to minimize the impact of such measurement errors and detection biases. Simple statistical analysis techniques are also available to correct or to control for these biases if they were to be identified and additional information is known concerning the expected test performance. PMID- 10863645 TI - Telomerase, cervical cancer, and human papillomavirus. AB - Review of the available data indicates that telomerase is activated in the majority of cervical squamous cell carcinomas as it is in most malignant neoplasms. Telomerase activity can also be detected in some preneoplastic cervical lesions, but the significance of this in unclear, because nonneoplastic, proliferating epithelial cells also can have telomerase activity. The bias introduced by cytologic sampling methods can complicate the interpretation of results. Quantitative telomerase assays may be useful in distinguishing nonmalignant, physiologic activation of telomerase from malignant activation. Studies evaluating telomerase component (hTR or hTERT) expression by evaluation of RNA, mRNA, or antigen have yielded conflicting results, but the observation that many nonmalignant, nontelomerase active cells have detectable hTR and hTERT suggests that many cells express telomerase RNA and catalytic components, but do not have active telomerase. The implication is that a regulatory overlay must exist that controls telomerase activation. Activation of the enzyme in carcinogenesis could conceivably be a physiologic activation that normally accompanies cellular proliferation, a direct appropriation of telomerase activity by the neoplastic process, or both. The presence of inactive telomerase in many cells also raises the possibility of a noncatalytic function for the telomerase complex. An understanding of telomerase interaction with HPV infection in the pathogenesis of cervical neoplasia must await a further elaboration of telomerase regulation. Likewise, application of telomerase detection in cervical cancer screening programs must await a better integration of telomerase regulation in normal and specifically in HPV-infected squamous epithelial cells. PMID- 10863646 TI - Human papillomavirus oncogenesis. AB - HPVs have evolved to accomplish the task of controlling host cell proliferation and differentiation to the end of producing more infectious virions. Coincident with the viral life cycle, however, is the risk that the viral genome will be disrupted and its DNA integrated into the host cell chromosomes. Integration of the viral genome is potentiated by host factors and extracellular effectors that alone may increase genetic instability. But it is consequent to viral integration that most HPV-associated malignancies develop. Investigations of the potential for HPV to immortalize primary cells or transform immortalized cells in vitro demonstrate two distinct classes of genital viral types: (1) oncogenic, exemplified by HPV 16 and 18; and (2) the nononcogenic types 6 and 11. Subsequently, localization of the HPV oncogene implicated that E6 and E7 act by uncoupling the checkpoint controls of the cell cycle principally by inhibiting the normal functioning of p53 and pRb, respectively. By in large, the nononcogenic viruses do not effect irreversible growth properties through these same viral genes and the same cellular counterparts. PMID- 10863648 TI - Nongenital human papillomavirus infections. AB - Although genital HPV types produce a broad spectrum of disease, the nongenital types are a bit more predictive. Particularly in the immunocompromised patient, it appears as though when they become symptomatic they cause warts. These warts can be a particular problem with immunocompromised patients where the malignant potential can also be expressed. Additional understanding of the relationship between the papilloma viruses and cutaneous oncology is very important. There needs to be an application of seroepidemiologic techniques to understand better the epidemiology and further research on more effective and less painful therapies. PMID- 10863647 TI - Treatment of human papillomavirus gynecologic infections. AB - The future holds promise for better prevention and treatment of HPV infections. Currently, therapeutic options for HPV infections are limited, expensive, and often ineffective. Recent advances in understanding the basic virology and natural history of HPV infection have identified hitherto unsuspected approaches to therapy and have suggested novel pharmacologic and other types of interventions. An important area for future research is the need for detailed information about the mechanisms that trigger latent HPV and induce wart formation or promote malignant transformation. Conversely, we also need to know more about mechanisms that keep the virus in check (latent) in many HPV-infected persons. PMID- 10863649 TI - Survival of the extremely preterm infant in North America in the 1990s. AB - Survival of extremely premature infants has been significantly higher in the last decade than previously, and may well have improved during this time. The majority of infants greater than or equal to 25 weeks' gestation survive today. Survival of infants 23 and 24 weeks' gestation is significantly lower, but is by no means negligible. Reports of survival of infants less than 23 weeks or less than 500-g birth weight are not unique. Moreover, the maximum survival of infants less than or equal to 25 weeks possible with current state-of-the-art care is not known. Currently available data do not allow survival of the individual extremely low birth weight or extremely premature infant to be predicted with clinically acceptable accuracy. The concept of a limit of viability is vague and clinically and ethically simplistic. The provision of neonatal intensive care is not necessarily beneficial or justified merely because it affords some minimal chance of survival. This phrase should not be used to summarize the complex issues involved in balancing maternal and neonatal risks and benefits of intrapartum and neonatal care of the extremely low-birth weight or the extremely premature fetus and infant, the suffering of the infant and family, parental values and autonomy, and consumption of limited communal resources. It should be deleted from our vocabulary. PMID- 10863650 TI - Current concepts on the pathogenesis and markers of preterm births. AB - Preterm births remain a major cause of neonatal morbidity and mortality despite our efforts over the past several decades. Our improved understanding of the complex mechanisms surrounding preterm labor, however, has resulted in the development of numerous biologic and clinical predictors of spontaneous preterm births. These developments offer the exciting prospect for the creation of specific interventions that are directed toward the various pathways involved with preterm births. PMID- 10863651 TI - Antecedents of cerebral palsy in very low-birth weight infants. AB - Research from the last two decades provides directions for efforts to prevent CP in VLBW infants. The pathogenesis of CP seems to involve factors operating both during pregnancy and in the neonatal period. The most important prenatal factor appears to be intrauterine infection. Perinatal infection and other risk factors, such as the death of a co-twin, placental abruption, and cerebral ischemia, could trigger a cytokine cascade resulting in damage to the developing brain. The low frequency of intrauterine infection in mothers with preeclampsia might explain the apparent protective effect of this disorder. If the brain damage attributed to intrauterine infection and other risk factors involves cytokines as intermediates, then blockade of the proinflammatory cascade or promotion of endogenous inhibitors might prevent CP. Other potentially preventive strategies include corticosteroids given to mothers (but not those given to neonates) and thyroid hormone. PMID- 10863652 TI - Adaptive mechanisms of developing brain. The neuroradiologic assessment of the preterm infant. AB - Since the 1980s, cranial sonography has been routinely performed in premature infants. This has produced a wealth of information about the more dramatic central nervous system lesions of IVH, PVL, and late VM. This information has included timing and evolution of these lesions and their eventual correlation with outcome. For two reasons the advent of MR imaging scanning has produced an interest in using this modality to evaluate these same infants. First, MR imaging gives an obviously superior image, and its ability to detect lesions is far superior to that of ultrasound. Second, the ability of cranial sonography to detect all of the children with CP or low IQ is limited. In our studies of outcome in very low-birth weight infants grade 3 to 4 IVH, PVL, or VM are able to detect only about 50% of the infants who developed CP by 3 years. This condition should be highly correlated with structural brain disease; an imaging modality that was more sensitive to central nervous system lesions should offer an advantage in predicting outcome. In the only prospective assessment of the ability of these two modalities to predict outcome at 3 years, van de Bor and colleagues found MR imaging did not do better than cranial sonography. This was largely because both modalities detected the most severe lesions, and most children with milder lesions on MR imaging had normal outcome. Studies of late (age 1 to teenage years) MR imaging scans in preterm infants show that a high percentage have white matter lesions but these lesions correlate poorly with outcome. If our concern when counseling parents is to alert them when a serious adverse outcome is likely in their child, then cranial sonography is to be favored precisely because it is less able to detect subtle lesions, which the developing brain has the capacity to overcome. On the other hand, if our aim is to detect all lesions, even though these lesions do not predict serious adverse outcomes, then MR imaging is to be favored. Research aimed at discovering etiologies and mechanisms of brain injury in these high-risk infants should use the more sensitive modality MR imaging. Finally, the interesting observation that preterm infants fare as well as they do despite MR imaging-identified lesions might stimulate research studying the adaptive mechanisms of developing brain. PMID- 10863653 TI - Growth outcomes of very low-birth weight infants in the newborn intensive care unit. AB - Advances in perinatal medicine will continue to improve our care and increase our understanding of the unique nutritional requirements of the VLBW infant and especially of the ELBW infant. Developments that permit neonatologists to meet those nutritional needs should improve the growth and well-being of VLBW infants. Longitudinal growth curves of hospitalized VLBW infants, such as the ones described in this article, should not be considered optimal and should be updated as ways safely to improve the growth of VLBW infants are identified. PMID- 10863654 TI - Multiple births and outcome. AB - The rate of multiple-gestation pregnancies has grown exponentially over the last few decades and is responsible for the steady increase in the birth rate of low birth weight infants. As a group, infants of multiple-gestation pregnancies have higher mortality and morbidity than singleton pregnancies. The increase in adverse outcomes is related directly to the increased risk for preterm delivery and low-birth weight, and not to the multiple gestation itself. Outcomes for multiple-gestation infants appear to be similar whether conceived spontaneously or through artificial reproductive technology. Efforts to reduce the birth rate of low-birth weight infants should target multiple-gestation pregnancies. PMID- 10863655 TI - Behavior, pain perception, and the extremely low-birth weight survivor. AB - This article explores the literature concerning responses to pain of both premature and term-born newborn infants, the evidence for short-term and long term effects of pain, and behavioral sequelae in individuals who have experienced repeated early pain in neonatal life as they mature. There is no doubt that pain causes stress in babies and this in turn may adversely affect long-term neurodevelopmental outcome. Although there are methods for assessing dimensions of acute reactivity to pain in an experimental setting, there are no very good measures available at the present time that can be used clinically. In the clinical setting repeated or chronic pain is more likely the norm rather than infrequent discrete noxious stimuli of the sort that can be readily studied. The wind-up phenomenon suggests that, exposed to a cascade of procedures as happens with clustering of care in the clinical setting in an attempt to provide periods of rest for stressed babies, an infant may in fact perceive procedures that are not normally viewed as noxious, as pain. Pain exposure during lifesaving intensive medical care of ELBW neonates may also affect subsequent reactivity to pain in the neonatal period, but behavioral differences are probably not likely to be clinically significant in the long term. Prolonged and repeated untreated pain in the newborn period, however, may produce a relatively permanent shift in basal autonomic arousal related to prior NICU pain experience, which may have long-term sequelae. In the long run, the most significant clinical effects of early pain exposure may be on neurodevelopment, contributing to later attention, learning, and behavior problems in these vulnerable children. Although there is considerable evidence to support a variety of adverse effects of early pain, there is less information about the long-term effects of opiates and benzodiazepines on the developing central nervous system. Current evidence reviewed suggests that judicious use of morphine for adjustment to mechanical ventilation may ameliorate the altered autonomic response. It may be very important, however, to distinguish stress from pain. Animal evidence suggests that the neonatal brain is affected differently when exposed to morphine administered in the absence of pain than in the presence of pain. Pain control may be important for many reasons but overuse of morphine or benzodiazepines may have undesirable long-term effects. This is a rapidly evolving area of knowledge of clear relevance to clinical management likely to affect long-term outcomes of high-risk children. PMID- 10863656 TI - Functional outcomes in self-care, mobility, communication, and learning in extremely low-birth weight infants. AB - Gaps have existed in specifying degrees of severity of cerebral palsy assessment of self-care and communicative competencies, and specifying age-appropriate preschool educational and behavioral competencies. Imbedded in the concept of measuring functional status is the interaction between health and neurologic impairments, developmental challenges and competencies, family resources and disadvantages, and the child's current status. In reviewing historic outcomes of severe ROP over the past 40 years, it was noted that severe ROP caused blindness in 2% to 11% of survivors. There was a constant observation that approximately 50% of severe ROP survivors with blindness had multiple functional and developmental challenges beyond blindness alone. Similarly, in reviewing outcomes of cerebral palsy, it is imperative to describe the severity of cerebral palsy and functional consequences in motor, selfcare, communication, and learning. The reason to measure the functional status of children with neurodevelopmental impairments before first grade is that the degrees of severity of these disorders can be specified before attending school with peers. Subtler aspects of neurodevelopmental impairments need to assess impact on literacy, information learning, written language, social competencies with peers, and recreational and community participation. In this way, we can understand the vulnerabilities and resiliences of children and families of VLBW and ELBW status. This is a critical step in understanding long-term quality of life and independent living issues. In addition, our efforts can address those factors and pathways whereby multiple disabilities and multiple functional limitations cluster. Our biomedical intervention can prioritize strategies that lessen severe multiple disabilities and simultaneously support families, when despite our best efforts functional challenges are life long. PMID- 10863657 TI - Perception of health status and quality of life of extremely low-birth weight survivors. The consumer, the provider, and the child. AB - In neonatal intensive care, parents make important clinical management decisions in conjunction with health professionals. Little information is available in the literature, however, on whether the preferences of health care professionals, parents, and children differ for the resulting health outcomes. This article compares the preferences of these stakeholders for four to five hypothetical health states that are common to extremely low-birth weight infants. The findings have conceptual and practical implications for decision making in the neonatal intensive care unit. PMID- 10863658 TI - Growth and respiratory health in adolescence of the extremely low-birth weight survivor. AB - ELBW children grow poorly in early childhood, but catch up substantially in weight and height by 14 years of age. The final adult stature for ELBW children remains to be determined. Despite the need sometimes for prolonged periods of assisted ventilation and oxygen therapy, the respiratory health and lung function of ELBW children is mostly normal in adolescence. ELBW children with BPD had similar respiratory health compared with ELBW children without BPD. The effects of cigarette smoke, both passive and active, and of newer therapies, such as exogenous surfactant and postnatal corticosteroids, on respiratory health in adulthood remain to be determined. PMID- 10863659 TI - Family factors and social support in the developmental outcomes of very low-birth weight children. AB - This study used data that were representative of the normative population of all infants born in 1988 and were followed during the first 3 years of life. Large developmental delays and limitations in function were common among children weighing less than 1500 g at birth. Among very low-birth weight infants, minority status and living in a household headed by a single mother further worsen the disadvantages associated with a very low birth weight. Nor could the disadvantages associated with very low birth weight be accounted for by controls for other risk factors or buffering statuses and behaviors. Among all children (including those of very low birth weight) poverty, reliance on Medicaid and other government sources for health insurance, a history of risky behaviors, and inadequate prenatal care are the major risk factors for developmental delays, limitations in function, and impairment at birth. State program benefit levels have no obvious effects on child outcomes, taking into account participation in individual programs. An important finding in light of TANF is that maternal work, the use of child care, and the form and cost of child care did not influence developmental delay, limitation in function, or impairment, the outcomes that we were able to measure during the first 3 years of life. TANF eligibility requirements, however, may increase difficulty in obtaining prenatal and other medical services for mothers and children in need--factors shown here to be related strongly to increased risk of low birth weight and developmental delays, limitations, and impairments. Race and ethnicity, poverty status, and family structure are fundamental factors in early child development and function. Minority status, poverty, and single-parent households greatly increase the likelihood that a mother will engage in risky behaviors (smoking, alcohol use, illegal drug use) during pregnancy and receive inadequate prenatal care. Risky behaviors and inadequate prenatal care are the major risk factors for a baby of very low birth weight. When perinatologists first encounter a new patient who is of very low birth weight they often see an infant who is minority, in poverty, and in a single-parent household. Although such children did more poorly in development and function by age 3, the major effects of these variables were through the selectivity of such children into very low birth weight; the direct impact of these factors on development is somewhat muted. This research suggests there are a number of policies that can reduce development delays and functional limitations among children in the United States. Programs that are targeted to a mother and child (such as WIC, AFDC, health insurance coverage, and possibly the more recent TANF programs) significantly reduced the risk an infant will be of very low birth weight. Access and use of adequate prenatal care are essential. Programs designed to combat maternal behaviors that place the fetus at risk (smoking, alcohol, and illegal drug use) can be very successful in reducing the likelihood an infant will be of very low birth weight. An additional payoff from such programs comes after the birth, because even taking into account birth weight, these variables negatively impact on early childhood development and function. Single-parent family structure, race and ethnic minority status, and poverty status also are known to impact on kindergarten readiness, so that we expect a delayed impact of these variables on the child. The strength of this article is the use of normative population data to assess the role of birth weight in child outcome. We examined prenatal risk factors for a baby of very low birth weight, traced the manner by which these selective risks are reflected in the composition of very low-birth weight babies, demonstrated how a very low birth weight was fundamental to delays in development, and identified risk factors and potential buffers in this process. (ABSTRACT TRUNCATED) PMID- 10863660 TI - Biology versus environment in the extremely low-birth weight infant. AB - This article examines the role of biologic and environmental factors in determining the long-term outcomes of extremely low-birth weight infants. Research focusing on follow-up to at least 4 years of age is reviewed. Methodologic issues related to sampling, the use of control groups, and diagnostic criteria are also discussed. The use of cumulative models of risk for examining the relative contribution of environmental and biologic factors is presented. PMID- 10863661 TI - Economics of prematurity in the era of managed care. AB - Both the acute intensive care of premature infants and the management of their long-term medical and educational sequelae are costly. Because neonatal intensive care is very effective in reducing mortality, however, its cost effectiveness as described previously is actually quite favorable when compared with other well accepted medical interventions, such as coronary artery bypass grafting and renal dialysis. This article has highlighted the relatively scant literature on which those estimates of costs and cost effectiveness of both neonatal intensive care and its component interventions rest. This is particularly true with respect to long-term resource use by graduates of NICUs. Without such information, we cannot hope to allocate resources in a way that ensures optimal care of this vulnerable population. PMID- 10863662 TI - Systematics and taxonomy of yeasts. PMID- 10863663 TI - Molecular approaches for analyzing diversity and phylogeny among yeast species. PMID- 10863664 TI - Yeast infections in humans with special emphasis on Malassezia furfur and infections caused by 'rare' yeast species. PMID- 10863665 TI - Yeast infections in veterinary medicine. PMID- 10863666 TI - Therapy of yeast-associated infections. PMID- 10863667 TI - Regulation of dimorphism in Candida albicans. PMID- 10863668 TI - Gene regulation during morphogenesis in Candida albicans. PMID- 10863669 TI - Extracellular proteinases of human pathogenic fungi. PMID- 10863670 TI - The cell surface of Candida albicans during morphogenesis. PMID- 10863671 TI - Yarrowia lipolytica: an organism amenable to genetic manipulation as a model for analyzing dimorphism in fungi. PMID- 10863672 TI - Cytoskeletal proteins and morphogenesis in Candida albicans and Yarrowia lipolytica. PMID- 10863673 TI - Pseudohyphal development of Saccharomyces cerevisiae. PMID- 10863674 TI - Dimorphism in Histoplasma capsulatum and Blastomyces dermatitidis. PMID- 10863675 TI - Morphogenesis of Cryptococcus neoformans. PMID- 10863676 TI - Hostility predicts magnitude and duration of blood pressure response to anger. AB - The hypothesis that hostile and nonhostile individuals would differ in both magnitude and duration of cardiovascular reactivity to relived anger was tested. Participants were 66 older adults (mean age, 62; 38 women and 28 men; 70% Caucasian American, 30% African American). Each took part in a structured interview scored using the Interpersonal Hostility Assessment Technique. Later each relived a self-chosen anger memory while heart rate and systolic and diastolic blood pressures were measured continuously using an Ohmeda Finapres monitor. Hostile participants had larger and longer-lasting blood pressure responses to anger. African Americans also showed longer-lasting blood pressure reactivity to anger. Health and measurement implications are discussed. PMID- 10863677 TI - Walking in (affective) circles: can short walks enhance affect? AB - Recent physical activity recommendations call for activities that are of moderate intensity and can be performed intermittently during the day, such as walking. These proclamations were based partly on the assumption that moderate activities are generally more enjoyable than physically demanding ones, and they are, therefore, also more likely to be continued over the long haul. However, little is actually known about the affective outcomes of short bouts of walking and extant findings are equivocal. Four experimental studies examined the affective responses associated with short (10- to 15-min) bouts of walking using a dimensional conceptual model of affect, namely, the circumplex. Results consistently showed that walking was associated with shifts toward increased activation and more positive affective valence. Recovery from walking for 10-15 min was associated with a return toward calmness and relaxation. This pattern, was robust across different self-report measures of the circumplex affective dimensions, across ecological settings (field and laboratory), across time, and across samples. PMID- 10863678 TI - Psychological distress, health beliefs, and frequency of breast self-examination. AB - Although monthly breast self-examination (BSE) is recommended for early breast cancer detection, most women do not comply. Few studies have examined the impact of psychological distress on BSE frequency. Recent research suggests that it may be particularly important to examine the role of distress in the recently identified phenomenon of BSE overperformance (> 1/month). One hundred thirty-five healthy women with and without family histories of breast cancer completed sociodemographic, health belief, general and cancer-specific psychological distress, and BSE frequency questionnaires. The central finding of the study was that BSE underperformance and overperformance had two distinct sets of predictors: health beliefs, specifically barriers against BSE and low confidence in BSE performance, were related to BSE underperformance, while higher levels of psychological distress, particularly cancer-specific intrusive thoughts, were related to BSE overperformance. Findings underscore the need to evaluate BSE under- and overperformance separately and to develop problem-specific interventions to increase compliance with monthly BSE. PMID- 10863679 TI - The relative efficacy of three cognitive-behavioral treatment approaches to temporomandibular disorders. AB - The purpose of this study was to evaluate the relative efficacy of different biopsychosocial treatment conditions on patients with chronic temporomandibular disorder. Ninety-four patients with chronic temporomandibular disorder were assigned to either a biofeedback treatment group, a cognitive-behavioral skills training (CBST) treatment group, a combined (combination of biofeedback/CBST) treatment group, or a no-treatment control group. Pain scores were analyzed pretreatment and posttreatment to determine group and within-subjects treatment effects. Results demonstrated that, in terms of a self-reported pain score, all three treatment groups had significantly decreased pain scores from pretreatment to posttreatment, while the no-treatment group did not. Moreover, patients in the biofeedback group were the most significantly improved compared to the no treatment group. Finally, participants in the three treatment groups displayed significant improvement in mood states. PMID- 10863681 TI - [Solarium: bilateral epithelial keratitis of the nasal half of the cornea]. PMID- 10863680 TI - The relationship of hardiness, coping strategies, and perceived stress to symptoms of illness. AB - We proposed a conceptual model based on research supporting the relationship between symptoms of illness and the determinants of hardiness, coping strategies, and perceived stress. In this model, hardiness, avoidance coping, and approach coping have paths to perceived stress, perceived stress has a path to symptoms of illness, and hardiness also has a path to symptoms of illness. We examined the goodness of fit of this model using path analysis and tested its stability, as well as the presence of gender effects, in corporate (N = 110) and university (N = 271) samples. The proposed model was a good fit for the data in the corporate sample, and no gender effects were found. The proposed model was not a good fir for the data in the university sample, therefore we added two paths that have received some support in the research: from approach coping to symptoms of illness and from avoidance coping to symptoms of illness. This model was a good fit for the data in the university sample, however, the path from approach coping to symptoms of illness had a critical ratio < 2.0, thus we removed this path and ran the model again. The final model was a good fit for the data, and no gender effects were found. Implications for the relationship of hardiness, coping strategies, and perceived stress to health are discussed. PMID- 10863682 TI - [Pigment epithelial cysts of the iris swimming in the anterior chamber]. PMID- 10863683 TI - [Database software for corneal transplant banks]. PMID- 10863684 TI - [Eyeglasses with progressive optic effect]. PMID- 10863685 TI - [Nocturnal measurements of intraocular pressure in patients with normal-tension glaucoma and sleep apnea syndrome]. AB - BACKGROUND: About half of all normal-tension glaucoma patients and about one third of all primary open-angle glaucoma patients have sleep apnea syndrome. If sleep apnea syndrome causes some cases of glaucoma, the optic nerve damage could result from repetitive nocturnal hypoxias or from repetitive intraocular pressure elevations at the end of the apneas. In this study, we determined the intraocular pressure at the end of long apneas. PATIENTS AND METHODS: In three patients having sleep apnea syndrome and normal-tension glaucoma we recorded in a sleep laboratory during at least six hours of sleep the respiration (oxymetry, nasal and oral air flow, and inductive plethysmography). The intraocular pressure was measured with a pneumatonometer at predetermined times and compared to the values measured at the end of prolonged apneas. RESULTS: The intraocular pressure during normal respiration was in the first patient 19.5 +/- 1.0 mm Hg OD and 19.3 +/- 1.7 mm Hg OS, in the second patient 25.0 +/- 4.2 respectively 25.5 +/- 4.9 mm Hg and in the third one 22 +/- 1.0 respectively 21.3 +/- 1.3 mm Hg. At the end of prolonged apneas the intraocular pressure was in the first patient 19.0 +/- 0.0 mm Hg OD and 19.5 +/- 0.7 mm Hg OS, in the second patient 26.5 +/- 0.6 and 26.8 +/- 0.1 mm Hg and in the third one 20.0 +/- 0.0 respectively 21.0 +/- 0.0 mm Hg. The difference between intraocular pressures during normal respiration and at the end of prolonged apneas was not significant (p > 0.1 for each comparison, paired t-test). CONCLUSIONS: We did not find an increase of intraocular pressure at the end of prolonged apneas compared to periods of normal respiration in patients with sleep apnea syndrome and normal-tension glaucoma. If sleep apnea syndrome causes some cases of glaucoma, it seems more probable that the the optic nerve is damaged by the repetitive hypoxias. Alternatively, an unknown factor might induce both, sleep apnea syndrome and normal-tension glaucoma. PMID- 10863686 TI - [Sutureless phacotrabeculectomy. Interim results]. AB - BACKGROUND: To study the medium-term success rate of a simple surgical technique for combined operation of cataract and glaucoma. MATERIALS AND METHODS: Of 50 patients with cataract and various types of glaucoma, 55 eyes underwent a combined operation for cataract and glaucoma and were followed up for at least 6.5 months. The phacoemulsification war performed through a scleral tunnel. After implantation of a foldable IOL an "inverse T" incision of the tunnel floor was performed to create a valve mechanism closure. RESULTS: In 45 eyes (83%) the intraocular pressure was controlled without medications during the follow-up time. In 10 cases (17%) a new glaucoma medication was introduced postoperatively. Following early postoperative complications were noticed: early intraocular hypertension (20.0%), wound leakage (20.0%), early intraocular hypotension (10.9%), hyphema (19.1%). CONCLUSIONS: No-stitch phacotrabeculectomy appears to be a safe and (medium-term) effective procedure for combined surgery of cataract and glaucoma. PMID- 10863687 TI - [Sclerothalamectomy (STE): stable postoperative intraocular pressure regulation in primary open-angle and pseudoexfoliative glaucoma]. AB - BACKGROUND: Since the 70th years we operated the hemorrhagic glaucoma by a ciliary body exposure after Benedikt. In a few patients we noted a stable intraocular pressure regulation for years, often in these cases where there is no filtering bleb. This signifies, that the ciliary body is heavily involved in the resorption of the anterior chamber fluid. During the following next years we further developed this operating technique and we use it today successfully in the operative treatment of primary open angle- and pseudoexfoliative glaucoma. PATIENTS AND METHODS: Since march 1996 we performed a sclerothalamectomy in 46 eyes. In approximate the half of cases the operation was done combined with a cataract surgery. 27 eyes had a primary open angle and 19 eyes a pseudoexfoliative glaucoma with a mean preoperative intra-ocular pressure of 29.79 +/- 7.96 mm Hg respectively of 33.58 +/- 9.32 mm Hg. RESULTS: The mean follow-up was in the case of primary open angle glaucoma 18.3 +/- 8.9 (median 20) and in the case of pseudoexfoliative glaucoma 15.8 +/- 11 (median 16) months. The mean postoperative intra-ocular pressure, which range in the group of primary open angle glaucoma 13.9 +/- 1.6 mm Hg and in the group of pseudoexfoliative glaucoma 12.9 +/- 2.5 mm Hg, was significant lower with significant less postoperative medication compared with the preoperative values during the complete follow-up (p < 0.01). There was no evidence in a significant difference between the both groups regarding the postoperative complication rate and intra ocular pressure. We observed altogether in 7 eyes a flat filtering bleb, 5/27 in the primary open angle glaucoma and 2/19 in the pseudoexfoliative glaucoma group. In all of other cases we didn't state a filtering bleb. CONCLUSION: The sclerothalamectomy leeds to a long-term stable intra-ocular pressure with a contemporary low complication rate in the case of primary open angle- and pseudoexfoliative glaucoma. PMID- 10863688 TI - [Intraocular hypotony: use of ultrasound biomicroscopy (UBM) for differential diagnosis and its schematic representation]. AB - OBJECTIVES: To determine the use of high-frequency ultrasound biomicroscopy (UBM) in the assessment of hypotony and in particular to determine the proportion of cases for which UBM contributed significant additional hitherto unaccessible information. PATIENTS AND METHODS: Ultrasound biomicroscopy was performed in a standard manner, using a Humphrey UBM 840 system (Humphrey Instruments, Inc., San Leandro, CA). UBM findings were analysed and the clinical relevance of UBM information was determined for the whole collective. RESULTS: Twelve patients with hypotony were examined. UBM findings contributed essential information that allowed to reach a diagnosis or that determined the therapeutic attitude in 10 of the 12 hypotonic patients. In two cases the cause of hypotony was tractional ciliary body detachment, in 5 cases it was post-inflammatory atrophy of the ciliary body, in 3 cases it was post-traumatic irido and cyclodialysis, in one case it was supraciliary and suprachoroidal effusion and in the last case it was due to uveal effusion syndrome. Based on these findings we established a schematic approach for hypotony. CONCLUSIONS: This procedure enabled us to assess the morphological changes found in patients with hypotony. In a majority of cases UBM was useful either to orient therapeutic intervention or to establish a diagnosis. On the base of our findings a schematic approach for hypotony, using UBM, was established. PMID- 10863689 TI - [Study of panophthalmitis after cataract surgery from 1997 to 1999]. AB - PURPOSE: To define the clinical outcome and microbiological pattern of bacterial endophthalmitis that were referred at the Jules Gonin Eye Hospital from January 1997 to September 1999. METHODS: Patients were recorded in a computerised databank and were managed according to a standard protocol. An anterior chamber tap combined with a vitreous biopsy by the pars plana was performed in all patients. The treatment included an intravitreal injection of 1 mg Vancomycin and 400 micrograms Amikacin diluted in 0.2 ml NaCl 0.9%. Postoperatively hourly therapy Cefazolin 50 mg/ml and Garamycin 9 mg/ml was applied. To determine possible risks factors a standard form was sent to all referring surgeons. The following data were analysed: delay of onset, risk factors, initial and final visual acuity. RESULTS: From January 1997 to September 1999, 31 patients were referred. 18/31 (58%) of the cases were admitted between April and June of each years. The mean age was of 75 +/- 10 years. Initial visual acuity ranged from light perception to 20/40. 17/31 of the patient's cultures were positive. The major pathogen were Staphylococcus epidermidis in 9/31 patients and Staphylococcus aureus in 4/31 patients. No correlation between the endophthalmitis and the surgical technique or perioperative management of the patient, could be determined. Visual outcome was significantly improved in 56.7% of the patients. CONCLUSIONS: The severity of outcome could be correlated to the type of bacteria isolated. The high prevalence of panophthalmitis from March to June suggests that a climatic factors may be involved in its pathogenesis. PMID- 10863690 TI - [Automatic measurement of dye filling of simultaneous digital ICG- and fluorescein angiography sequences]. AB - BACKGROUND: The ICG filling is supposed to be faster than Fluorescein filling. Interestingly the filling characteristics of these dyes were never correlated directly using precise quantitative methods. Since ICG and Fluorescein are injected as a mixture, the simultaneous 2-channel angiography provides a suitable method to correlate the filling characteristics of the dyes. MATERIAL AND METHODS: The simultaneous ICG and Fluorescein angiograms were recorded with a Rodenstock Scanning Laser Ophthalmoscope. The angiographic images were digitized real-time with a graphic workstation. Filling characteristics of the two dyes was calculated after off-line eye tracking in different regions of interests (ROIs) on the central retina. RESULTS: The Fluorescein filling was faster than the ICG filling in 56.5% of our patients. In 26% of our patients was a mixed filling detectable. Depending on the position of the ROIs the Fluorescein or ICG filling was faster. In only 17.5% of our cases was the ICG filling faster than the Fluorescein filling. CONCLUSION: Our results show that Fluorescein filling in more than 50% of the cases is faster than ICG filling and only a minority of the patients has a faster ICG filling. According to our experience the filling pattern of the two dyes is individual, there is no rule of thumb for the filling. PMID- 10863691 TI - [Prognostic factors and results after surgical treatment of idiopathic macular holes, stage 2 and 3]. AB - PURPOSE: To determine prognostic factors, functional outcome and subjective rating after surgery for macular holes stage 2 and 3. METHODS: We studied 53 eyes of 49 patients undergoing vitreous surgery for macular holes stage 2 (46%) and 3 (54%). Mean follow-op was 114 weeks (32-204, std.dev. +/- 48), mean age 68.9 years (44-89, std.dev. +/- 6.8). 72% were female, 11% were pseudophakic, 19% phakic, 70% had a combined procedure (pars plana vitrectomy, phacoemulsification and IOL). Surgery consisted in a pars plana vitrectomy, peeling of epiretinal membranes and ILM, internal tamponade with SF6 (98%) resp. Si-oil in one case. Patients had to keep face-down position 6 x 20 minutes per day. RESULTS: The hole was completely closed in 90.6%. Anatomical failures included, 86% had an increase of VA, 41% = 5 lines (Final VA median 20/30, max. 20/20). No further increase of the retinal function occurred after 6 months. The visual result did not correlate with the duration of symptoms. 84% were satisfied with the outcome, subjective rating was not correlated with final VA or change of VA. 19% showed postoperative typical peripheral visual field defects. Visual field loss was not correlated with perioperative IOP elevation. CONCLUSION: Macular hole surgery has a high functional success rate. Postoperative visual field defects are an important problem. PMID- 10863692 TI - [Follow-up of patients suffering from age-related macular degeneration, supplied with visual aids]. AB - BACKGROUND: Age-related macular degeneration (AMD) is at present the major cause of legal blindness in industrialized countries. The results of various treatments are often not satisfactory and follow-up by Low-Vision services is then the only solution for this population. Few studies have been published on this subject. The purpose of this study is to present the use of magnifying systems and the degree of satisfaction obtained for this series of patients 6 months after prescription. PATIENTS AND METHODS: Forty-four patients were examined, 42 of whom were eligible for the study. Lenses, magnifying glasses, visual systems for intermediate distance, and electronic systems were proposed. A low-vision training was given to every patient. RESULTS: The highest degree of use and satisfaction was achieved with the electronic systems. In some cases, however, the other systems were well accepted by the patients. CONCLUSION: The follow-up of patients with age-related macular degeneration is of great social importance. The independence of elderly people can be safeguarded thanks to the help of Low Vision services with a good equipment and a well-trained team. Sometimes, unsophisticated systems such as magnifying glasses can be proposed, but the best results are mostly obtained through electronic systems (TV). PMID- 10863693 TI - [Antiretinal antibodies associated with cystoid macular edema]. AB - AIM: Recently, anti-Enolase and anti-carbonic anhydrase antibodies have been observed in over 60% of patients with retinitis pigmentosa (RP) and cystoid macular oedema (CME). We investigated the presence of these antibodies in a series of patients with CME due to different pathologies. METHODS: In 10 patients with CME serum antibodies against Carbonic anhydrase (CA) II (30 kD) and Enolase (46 kD) were sought using Western Blots, Dot Blots as well as ELISA. RESULTS: Western and dot blotting showed anti-CA II antibodies in all and anti-Enolase antibodies in six of the 10 patients. The average titer measured with ELISA was 0.9 +/- 0.08 OD Units (0.35-1.4) with a dilution of 1:400. CONCLUSION: The presence of anti-retinal antibodies in the serum of all patients confirms the high prevalence of these antibodies in patients with CME. This may suggest that a dysfunction of CA and enolase activity in the retinal pigment epithelium may lie at the root of oedema formation, whereas other mechanisms may be responsible in the absence of these antibodies. PMID- 10863694 TI - [Pars-plana vitrectomy in diabetic traction retinal detachments with holes]. AB - PURPOSE: To evaluate results and prognostic factors of pars-plana vitrectomy (ppv) using membrane peeling and C2F6-gas for diabetic traction retinal detachments with a hole. METHODS: We retrospectively reviewed the healing course of 84 eyes, which were treated in this way consecutively (follow-up 6-32 month). C2F6 was always used in case of a retinal hole at the posterior pole or in the upper part of the eye. The influence of HBA1c, type of diabetes, preoperative visual acuity, preoperative laser-coagulation and macular status over treatment results and rate of complications were studied using multivariate logistic regression analysis. RESULTS: Preoperatively 56% of the eyes showed premacular vitreous hemorrhage and 69% macular detachment. Using on an average 1.3 treatments in 73% retina was completely reattached and in 85% visual acuity remained unchanged or improved. The most important prognostic factors were HBA1c < 9.3, type II diabetes and bad vision preoperatively. Postoperative complications such as rubeosis, secondary glaucoma and repeated vitreous hemorrhages were more frequent in case of type I diabetes, HBA1c > or = 9.3 and insufficient laser treatment. CONCLUSION: Best corrected blood sugar levels preoperatively and sufficient laser treatment seem to be very important to decrease the risk for rubeosis and repeated vitreous hemorrhages especially for patients with type I diabetes. PMID- 10863695 TI - [Vogt-Koyanagi-Harada syndrome: importance of rapid diagnosis and therapeutic intervention]. AB - AIM: The Vogt-Koyanagi-Harada (VKH) syndrome is characterized by a bilateral granulomatous uveitis with exudative retinal detachments associated with systemic manifestations such as meningeal signs, cutaneous signs (poliosis, alopecia and vitiligo) and dysacousis. VKH is relatively unfrequent in Europe and Switzerland. Therefore diagnosis is often reached with some delay. Our aim here was to analyze the 3 patients for whom the diagnosis was reached less than 15 days after the first signs and compare their evolution to seven patients for whom diagnosis was known one month or more after the first signs. PATIENTS AND METHODS: Retrospective and partially prospective study of patients seen at uveitis clinic in Lausanne from 1990 to 1999 for whom an ICG angiographic work-up had been performed in addition to the usual clinical and fluoresceinic work-up. The frequency of VKH in our collective was calculated; symptoms and signs, paraclinical investigations, laboratory work-up, delay from first signs to diagnosis, the management and the evolution were the criteria analyzed. In particular the patients with early diagnosis and early treatment were analyzed and compared to the rest of the collective. Diagnosis was based on the criteria of the American Uveitis Society. Between 1990 and 1999, 14 patients with the diagnosis of VKH were seen (1.2% of our collective of uveitis patients). The 10 patients having had a work-up including ICG angiography in addition to the classical work-up were included in this study. RESULTS: The diagnosis was reached in less than 2 weeks in 3 patients. In all 3 patients inflammation was controlled after treatment. Two patients with a follow-up without recurrence of respectively 36 and 54 months were considered as healed. The last case had no recurrence after nine months but still was under therapy. Whereas clinical examination and fluorescein angiography failed to show any sequels in the 2 "healed" patients, ICG angiography showed numerous zones of hypofluorescence indicating choroidal scarring. For the 7 other cases, the diagnosis was reached one month or more after the first symptoms or signs and they all evolved in the chronic recurrent fashion. ICG angiography contributed to the rapid diagnosis in 2/3 patients with early diagnosis and was an essential parameter for the choroidal follow-up in 9/10 patients. CONCLUSION: This study shows that it is essential to rapidly reach the diagnosis of VKH and treat the patients vigorously without delay. By showing choroidal lesions not seen by the clinical examination or fluorescein angiography. ICG angiography is essential for a correct work-up and follow-up of choroidal lesions in VKH. In our two "healed" patients it was the only mean to show choroidal sequellae. PMID- 10863696 TI - [Vertical diplopia after cataract operation]. AB - PURPOSE: Presentation and analysis of patients with vertical diplopia appearing after cataract surgery in retrobulbar anesthesia. SUBJECTS AND METHODS: Between 1990 and 1998 9 Patients with vertical diplopia following cataract surgery in retrobulbar anesthesia were studied in our Orthoptic Department. Each patient had complete orthoptic examination with Hess-screen-test. Additionally, some patients underwent neuroradiologic imaging and forced-duction testing. RESULTS: We subdivided the patients in a group of 4 patients with hypertropia and of 5 patients with hypotropia of the operated eye. All hypotropias were left-sided. Seven patients showed an overaction of the involved muscle without regression. Seven patients underwent surgery of a vertical muscle. Only 1 patient needed prismatic therapy postoperatively. The other 2 non-operated patients were satisfied with prisms alone. CONCLUSIONS: The proposed pathogenesis of vertical diplopia in these cases is fibrosis and contracture of the injured muscle, which could be due to anesthetic myotoxicity after direct injection into the muscle or to an intramuscular hemorrhage. On the other hand hypertropia could be a result of placement of bridle sutures. We discuss prevention and therapy of such complications. PMID- 10863697 TI - [Preventive measures against ocular trauma on squash courts]. AB - AIM: Squash-ball induced trauma is an important cause of severe ocular injuries. Fortunately they can be prevented by wearing of protective goggles. The aim of this study was to investigate the prevalence of these protective devices in Swiss Squash Clubs. METHODS: A detailed questionnaire was sent to all Squash Clubs in Switzerland (n = 82). The solicited information comprised: the possibility to buy protective goggles in the proshop, presence of posters in the club encouraging the members to wear protective goggles, assessment of the risk of ocular trauma by a squash ball (high or low), and the evaluation of ocular sequelae after squash ball injury. RESULTS: In the French-speaking part of Switzerland 35% of the clubs responded, whereas the percentage in the German-speaking part was 32% did. Of all the clubs who returned the questionnaire only one was found to sell protective goggles in the sports centre (4%). Two further clubs had posters encouraging the players to wear eye protection (7%). Three other clubs in the French-speaking area and two in the German-speaking area planned to introduce these measures (18%). 63% of the clubs considered the risk of an ocular injury to be low, 7% even as very low. Only 26% of the clubs who returned the questionnaire considered the risk to be much higher. CONCLUSION: The interest of squash clubs in protective eye wear is very low, as the risks for a serious ocular injury are underrated. It is therefore important to bring the danger of racket sports to the attention of patients. PMID- 10863698 TI - [Rigid GRIN-endoscope for endoscopy of the tear ducts]. AB - BACKGROUND: Using small endoscopes it is now possible to evaluate the status of lacrimal ducts in vivo. GRIN-Lenses produce better pictures from the lacrimal ways than fiber-bundles. GRIN-endoscopes are rigid. The present study is concerned with the feasibility and indications of such GRIN-endoscopic examination. PATIENTS AND METHODS: Two different GRIN-endoscopes with a distal diameter of 0.89 mm were used. One with a phi 0.5 mm optic and an additional working channel and another one with a phi 0.35 mm optic and two additional working channels. One channel each was used for injection of air or 0.9% NaCl. Either a laser fiber or another instrument (max phi 0.16 mm) could be introduced into the second channel. 44 patients in age between 18 and 87 with symptoms of epiphora or signs of chronic lacrimal way affections were examined. RESULTS: The presaccal lacrimal ducts could be clearly visualized in all patients. In case of presaccal stenosis, the examination of the lacrimal sac was not always possible. The endoscope used, based of GRIN-lenses, gave an excellent image quality. Endoscopy under local anesthesia was well tolerated by all the patients with affection of the lacrimal drainage system. Because this ambulant examination does not stress the patient too much, it could be repeated a number of times. Endoscopy under general narcosis could enlarge the application spectrum. This method can complement the ambulant diagnostics and therapy of the lacrimal drainage system. PMID- 10863699 TI - [Aqueous outflow pathway imaging in the rabbit with fluorescein and indocyanine green]. AB - PURPOSE: The purpose of this study was to visualise the rabbit aqueous outflow pathway using a numeric imaging system with ICG and fluorescein injection in the anterior chamber. MATERIAL AND METHODS: We performed a simultaneous injection of Indocyanin Green (ICG) and Fluorescein into the anterior chamber of rabbit eyes. Using a digital camera, we took several sequenced pictures to visualize the distribution of the dyes within the outflow pathway. We observed the dynamics of the outflow over time. RESULTS: In the early phases, the shape of the outflow canal around the limbus was clearly seen. Several collecting veins close to the recti muscles were also identified. There was only slight fluorescein leakage during the early phases, allowing adequate visualization of the morphology of the outflow system. In the late phases, the sclera was stained with the fluorescein, and no details were thus visible. The ICG dye allowed better recognition of the fine details of the outflow structure. CONCLUSION: This method was relatively simple, safe and precise, and allowed us to visualise the details of the outflow pathways in the rabbit eyes. These results could be of great value in further evaluating the outcome of filtering surgeries in animal models. PMID- 10863700 TI - [Effect of dark adaptation on retinal blood flow]. AB - PURPOSE: Laser Doppler measurements performed immediately after the transition from dark adaptation (DA) to light led to the hypothesis that retinal blood flow, Fret, is increased during DA, but the use of visible lasers had prevented measurements during DA. Our aim was to test this hypothesis by measuring Fret during and after DA. MATERIAL AND METHODS: Fret in retinal vessels at the optic disk surface was recorded quasi-continuously in one eye of 6 normal subjects (age 27 to 60 years) using a laser Doppler flowmeter in the near-infrared (810 nm). Measurements were performed during light (baseline), various periods of DA and again during light. DA lasted between 2 and 32 min. RESULTS: Average Fret for the 6 subjects did not change significantly (-2.7 +/- 8% sd, p > 0.05) during the various periods of DA, as determined from linear regressions of the flux versus time. Following the transition from DA to light, there was, in most cases, a rapid transient increase of the flux, which reached an average value of 37 +/- 10% above the pre-transition value and peaked at 30-60 sec after the transition. CONCLUSIONS: These results do not support the hypothesis that Fret in normal volunteers is increased during DA. Rather, they strongly suggest that the transient increase in flux observed after DA is induced by the transition from dark to light (FNSRS #3200-043157 et CNR, It. #95.01715.CT04). PMID- 10863701 TI - [Effect of light on choroidal blood flow in the fovea centralis]. AB - PURPOSE: To investigate whether light and dark exposures induce a response of choroidal blood flow (ChBF) in the foveal region in humans. METHODS: In a group of healthy volunteers (age 25-60 years) ChBF was measured using a new confocal laser Doppler flowmeter (probing laser at 785 nm, power at the cornea = 90 microW). ChBF was recorded at room light, in darkness, at room light following dark adaptation, and during strong light exposure following room light. RESULTS: While ChBF was stable during room light condition, it decreased significantly by 15% (p < 0.01) during darkness. After 6 min of room light following darkness, ChBF was back to baseline. Strong diffuse, green light exposure over a field of 45 degrees had no detectable effect on ChBF. In all the experiments, no significant change of blood pressure was detected. CONCLUSIONS: Our findings did not confirm the presence of an active process of ChBF regulation in response to strong light exposure in humans. They demonstrate, however, a reversible decrease in ChBF occurring after a transition from room light to darkness. PMID- 10863702 TI - [Optical Doppler velocimetry of red blood cells at different depths in retinal vessels by varying the coherence length of the source: feasibility study]. AB - PURPOSE: To report on a novel approach to measure the velocity of red blood cells (RBCs) at different retinal vessel depths. METHOD: The technique is an extension of conventional laser Doppler velocimetry using light sources of various coherence lengths (CL). Light scattered by the moving RBCs interferes with that reflected from the anterior vessel wall only if the optical path difference is shorter than CL. Therefore, using low coherence light sources, localized measurements of RBCs velocity can be performed. RESULTS: Measurements of RBCs velocity at different depths in a main retinal vein (diameter: 152 microns) of a volunteer has been performed using 4 different light sources with CLs of 14 microns, 21 microns, 32 microns and > m. Measured values are in good agreement with theoretically predicted values. CONCLUSION: This new approach permits to measure RBCs velocity at different depths of retinal vessels in the human retina. PMID- 10863703 TI - [Effect of insulin on retinal glycogen content]. AB - BACKGROUND: The effect of insulin on glucose and glycogen metabolism in peripheral organs is well known. However, information about the action of this peptide in the retina is incomplete. We addressed the questions whether insulin influences glycogen content in the cat retina and whether glycogen breakdown is triggered by lack of glucose. MATERIAL AND METHODS: Eyes from adult cats were enucleated under deep barbiturate and fentanylanesthesia. Retinas were snap frozen either before or following arterial in vitro perfusion. Three conditions were studied: a) Perfusion with a glucose- and insulin-free medium; b) perfusion with the addition of physiologic glucose concentration; and c) in combination with insulin. Glycogen content was determined by in vitro measurement of glucose converted from glycogen. RESULTS: The reference value for retinal glycogen after enucleation (10 min of ischemia) is 2.4 micrograms glucose/mg protein. Glucose- and insulin-free perfusion for 80 min following "normoglycemia" reduced the amount of retinal glycogen by one third. Perfusion for 3 h with 5.5 mM glucose led to a small increase of the partly depleted glycogen stores. Insulin, in contrast, markedly augmented the glycogen content. CONCLUSIONS: Insulin led to an increase in retinal glycogen content, indicating an influence of this peptide on retinal glucose and glycogen metabolism. However, it appears that glycogen might play a dynamic role in retinal metabolism as a buffer between abrupt changes in focal metabolic demands that occur during normal glucose supply rather than acting solely as an emergency energy reserve for neural function during hypoglycemia. PMID- 10863704 TI - Vasorelaxing properties of carteolol in isolated porcine ciliary arteries. AB - INTRODUCTION: Carteolol is a topical beta-blocker used in ophthalmology to decrease the intraocular pressure. This study investigates the vasoactive effect of carteolol in isolated porcine ciliary processes. MATERIAL AND METHODS: With a myograph system isometric force measurements, quiescent vessels or vessels either precontracted with endothelin-1 (0.03 microM) or the thromboxane A2 analog U 46619 (0.3 microM) were exposed, in cumulative manner, to increasing concentrations of carteolol (10 microM-3 mM). Vessels that had a functional endothelium were compared with vessels that had a non functional endothelium (intentionally and mechanically damaged). RESULTS: In quiescent vessels carteolol did not induce contractions. In contrast, in precontracted arteries, carteolol evoked marked relaxations which were endothelium-independent. CONCLUSIONS: In vitro, carteolol has a marked vasorelaxing effect on porcine ciliary arteries, however the clinical relevance of this observation for the care of glaucoma patients requires further evaluation. PMID- 10863705 TI - [Membrane potential depolarization in the porcine ciliary epithelium evoked by nitric oxide]. AB - BACKGROUND: The present work studies if in porcine ciliary body epithelium the intracellular signal transduction pathway nitric oxide (NO)--guanylate cyclase (GC)--3',5'-cyclic guanosinemonophosphate (cGMP) can change the membrane potential of the epithelium of the ciliary body. MATERIAL AND METHODS: Recordings of membrane potentials were done by means of intracellular microelectrodes in the presence of the NO donor Sodium Nitroprusside (SNP; 100 microM; n = 5) or the membrane permeable cGMP analogue 8-parachlorophenyl-thioguanosine-3',5' cyclic monophosphate (8-pCPT-cGMP; 100 microM; n = 5). To test whether the GC is involved in this process, recordings were repeated in both groups in presence or in absence of the GC inhibitor 1-H-(1,2,4)oxadiazole-(4,3-a)quinoxalin-1-one (ODQ; 10 microM; n = 4). RESULTS: SNP and 8-pCPT-cGMP both induced significant membrane potential depolarizations (7.7 +/- 1.8 mV and 13.1 +/- 1.3 mV, mean +/- SEM). Membrane depolarizations induced by SNP were significantly inhibited by ODQ (p < 0.01), whereas depolarizations induced by 8-pCPT-cGMP were not altered by the presence of ODQ (p > 0.2). CONCLUSIONS: Nitric oxide induces depolarizations of the membrane potential in the isolated porcine ciliary body. This process is transduced by activation of the GC. We conclude that nitric oxide might be involved in the regulation of permeability of the cellular membrane for ions in the ciliary body. PMID- 10863706 TI - [Measurement of eye length and eye shape by optical low coherence reflectometry]. AB - BACKGROUND: The precise and rapid measurement of eye length and eye shape are essential for investigating eye growth regulation and myopia. For this purpose, we developed an optical low coherence reflectometer and obtained preliminary measurements in volunteers. METHODS: The instrument includes a rotating glass cube to produce longitudinal scans at a velocity of 0.42 m/s and a repetition rate of approximately 13 scans/s. Heterodyne detection of light reflected from the anterior cornea and the posterior retina permits to measure axial eye length and eye shape (off-axis eye length). Each measurement consists of five consecutive scans. Reproducibility and precision were determined in one volunteer by measuring axial eye length five consecutive times, each time repositioning the eye. Eye shapes were determined in right eyes of four volunteers by measuring eye length every 3.3 degrees from 10 degrees nasally to 10 degrees temporally. RESULTS: Axial eye length measured repeatedly in one volunteer did not differ between or within the measurements (one-factor ANOVA). The average standard deviation was 11 microns. Eye shapes a) varied substantially among subjects and b) differed considerably from the corresponding shapes of spherical model eyes with identical axial eye lengths. CONCLUSION: The newly developed reflectometer permits the precise and rapid measurement of eye length and eye shape. Such measurements, especially in children, may provide important information about mechanisms of eye growth regulation and the development of myopia. PMID- 10863707 TI - [Slit-lamp perimetry: a new diagnostic technique]. AB - AIM OF THE STUDY: To test a novel diagnostic technique, slit-lamp perimetry. PATIENTS AND METHODS: Slit-lamp perimetry is performed during a normal slit-lamp examination by projecting a small, round light mark onto the fundus. The light mark is moved and consecutively the patient is asked, if the light moved towards or away from the scotoma. Using the patients feedback the light mark can be placed exactly onto the retinal region corresponding to the scotoma. This method was tested on a patient with a microinfarction of a small retinal arteriole and on two patients with small preretinal parapapillary vitreous floaters. RESULTS: Slit-lamp perimetry correctly localized preretinal vitreous floaters and a fresh cotton wool spot missed on an dilated fundus examination. CONCLUSION: Slit-lamp perimetry is a novel rapid diagnostic technique to localize retinal and preretinal pathologies reponsibles for scotomas. PMID- 10863708 TI - [Strabismus, hemianopsia and the sundial of Mawas and Franceschetti]. AB - THE ARTIFACT: The artifact is a light rectangular slate of black cardboard showing two white lines at 40 degrees angle. It provokes physiological diplopia in lateral vision. BACKGROUND: In normal subjects, the right oblique line stimulates the right nasal hemiretina, perceived as an oblique line, and the left temporal hemiretina, perceived as a vertical line, both lines being on the right hand side. Likewise, the left oblique line is seen as two lines on the left hand side. The 4 lines form and M and a W combined. The strabismus subject sees 3 lines for he neutralizes one; the alternating squint sees only 2; the heterophoric sees 4 lines, two of which blurred, and the hemianopic sees only part of the dial. RESULTS: The subject must select the image he sees from an array of possibilities shown on the back of the dial. From 5 years of age on, normal subjects select the [symbol: see text] or the "butterfly"; hemianopic subjects correctly describe the loss of either the left or right field. CONCLUSIONS: The sundial facilitates both the screening and the treatment of binocular vision disorders. Bangerter's hemifilter will be used in association with exercises for physiological diplopia at home under monthly orthoptic control. All this made it possible to obtain useful peripheral fusion in early onset strabismus. PMID- 10863709 TI - [Retinal vasculopathy associated with Berger's IgA nephropathy]. AB - AIM: Ophthalmological complications associated with Berger's IgA nephropathy comprise scleritis, episcleritis, keratoconjunctivitis as well as anterior uveitis. We present a new association of IgA nephropathy with a retinal vasculopathy. METHODS: Presentation of two clinical cases. RESULTS: Two patients presented with hematuria and epistaxis associated with a retinal vasculopathy characterised by teleangiectasies, capillary occlusion with retinal hemorrhages, neovascularisations and macular edema with decreased visual acuity. Fluorescein angiography showed zones of non-perfusion as well as vasculitic changes. A general medical exam revealed a normal arterial pressure but a slightly elevated creatinine. Immunological investigations for the presence of antibodies showed no positive results. Renal biopsy demonstrated mesangial proliferations with diffuse deposits of IgA. Over the course of a 2 year follow-up some of the retinal changes regressed under treatment with cortisone and visual acuity returned to normal. The teleangiectasies showed no progression. CONCLUSION: Berger's IgA nephropathy can be associated with a retinal vasculopathy which may be due to local deposition of IgA immune complexes in the retinal vessels. PMID- 10863710 TI - [Ciliary body metastasis as the first sign of small-cell bronchial carcinoma]. AB - BACKGROUND: About 3-10% of patients suffering from metastatic disease develop metastatic choroidal tumors. In this case report ocular symptoms were the first sign of a systemic malignancy. CASE REPORT: A 65-year old man complained of an increasing painful redness of his left eye. He showed a white mass with 10 mm in thickness in the temporal chamber angle with "pseudohypopyon" and secondary glaucoma. Because of increasing pain the eye was removed. Histologic examination showed a small cell carcinoma of the ciliary body with high malignancy, which most probably has to be seen as a metastasis of a carcinoma of the lung, which was detected by internal examination concurrently. In addition, a metastasis in the adrenal gland was found. ACE polychemotherapy was performed, but 4 months later the patient died of metastatic disease with break down of the kidneys. DISCUSSION: Based on literature data patients with systemic malignancy present themselves to a physician primarily because of ocular symptoms in 12-31%. In these cases differentiation of an uveal metastasis from a primary amelanotic melanoma can be difficult. The search for primary tumors has to be started in due time. PMID- 10863711 TI - [Stargardt's disease and its intrafamilial variability]. AB - BACKGROUND: Because of its considerable differential diagnosis and a wide range of phenotypic variation, Stargardt's disease (juvenile macular dystrophy) can cause diagnostic problems. Moreover, ample variability in course and outcome of the disease has been described. PATIENTS AND METHODS: The diagnosis and variable course of Stargardt's disease of three affected siblings have been documented by clinical manifestation, fluorescein angiography and Ganzfeld-ERG including the ISCEV standard protocol. RESULTS: All three siblings presented with retinal abnormalities. However, only the youngest brother revealed symptoms for more than 10 y in terms of reduced visual acuity. The two elder ones had preserved central vision. Classical findings of contact lens biomicroscopy, fluorescein angiography and changes in the Ganzfeld-ERG confirmed the diagnosis of Stargardt's disease. CONCLUSIONS: The diagnosis of Stargardt's disease can be made as late as in the 7th decade of life. Tremendous variability can occur in course and outcome even within the same family. Therefore, visual prognosis is uncertain and has to be made with caution. In most cases the diagnosis can be established with the above mentioned methods. PMID- 10863712 TI - [Mydriasis induced by splinter from belladonna bush]. AB - AIM: Pupillary dilatation with alcaloids from the Atropa belladonna plant have been used since antiquity. It is less known that the wood of the plant contains an even higher concentration of alkaloids than the ripe fruit. METHODS: Clinical case. RESULTS: A 32-year-old patient presented with a foreign body sensation after a splinter got in his eye while chopping wood of an Atropa belladonna bush in the underbrush. Ophthalmic examination showed a visual acuity of 0.9 s.c. (1.0 with pinhole) and a pupil in a fixed mydriasis without any visible rupture of the sphincter muscle. After a 3-day follow-up the pupil returned to normal size and visual acuity recovered to 1.25 s.c. CONCLUSION: The wood of the Atropa belladonna plant contains a very high concentration of alkaloids. A well-directed history may help to discover this less well-known cause of an accidental mydriasis. PMID- 10863713 TI - Change: 'make or become different' or simply making a difference! PMID- 10863714 TI - Effect of pulse repetition rate on the perception of thermal sensation with pulsed shortwave diathermy. AB - BACKGROUND AND PURPOSE: Pulsed shortwave diathermy (PSWD) is a form of therapy commonly used to enhance tissue repair and reduce pain. It is normally considered to be an athermal form of treatment; however, there is some evidence to suggest that thermal effects can arise with adequate dosage. The purpose of this study was to determine the pulse repetition rate (PRR) required to generate a 'possible' and 'definite' thermal sensation when PSWD was applied to the thigh. METHOD: Thirty healthy subjects were randomly assigned to placebo or treatment groups. The treatment group was exposed to PSWD at a constant setting of pulse duration (400 microseconds) and pulse power (190 W) while the PRR was increased from 26 Hz to 400 Hz in 10 increments. Each dose was applied for a period of two minutes. At the end of each application, subjects were asked if they felt a (1) 'possible' or (2) 'definite' thermal sensation. Skin temperature was measured immediately after each application. Placebo subjects were exposed to PSWD at its lowest settings throughout the experiment (pulse power = 5 W; pulse duration = 65 microseconds and PRR = 26 Hz). RESULTS: The results showed a significant correlation (p < 0.048) between PRR at 'definite' thermal sensation and skin temperature post-treatment and PRR at 'possible' thermal sensation (p < 0.001). Mean skin temperature increased significantly as PRR was increased, from 28.69 (+/- 0.75) degrees C pre-treatment to 31.14 (+/- 1.04) degrees C post-treatment, a mean difference of 2.34 degrees C. CONCLUSIONS: These results suggest that PSWD at adequate dosages can generate thermal effects, and that there is a relationship between these thermal effects and the PRR used. These results may have significant implications for the safe use of PSWD in the clinical arena. PMID- 10863715 TI - Physiotherapists in Balint group training. AB - BACKGROUND AND PURPOSE: Balint group training (BGT) is a method widely used for enhancing understanding of the relationship and communication between health professionals and their patients. Participants meet in small groups, on a regular basis, with a tutor to discuss their experiences of problem cases. The method was originally developed in the 1950s for enhancing understanding of the doctor patient relationship. Few studies have focused on BGT and physiotherapists. The aim of the present study was to describe and analyse physiotherapists' experiences of participation in BGT as a means of learning and understanding the physiotherapist-patient relationship. METHOD: Semi-structured, in-depth interviews were conducted with three physiotherapists working in private practice, all participating in BGT. The interviews were transcribed and subjected to a qualitative analysis. RESULTS: The results are presented in a sequential model, featuring eight themes in which the physiotherapists' experiences of the training process are portrayed. CONCLUSIONS: The results suggest that BGT and sharing the experiences of others may be considered a way of enhancing understanding of the patient encounter in clinical practice, possibly to the benefit of physiotherapists and their patients. PMID- 10863716 TI - Effect of positioning on respiratory synchrony in non-ventilated pre-term infants. AB - BACKGROUND AND PURPOSE: Body position can play an important role in an infant's recovery from respiratory disease, but few studies have accounted for sleep state which is known to have a direct influence on the control of respiratory muscles as well as on metabolic and circulatory changes. The purpose of this study was to examine the influence of body position on respiratory function in pre-term infants whilst accounting for sleep state. METHOD: Thoraco-abdominal motion was assessed using respiratory inductance plethysmography (RIP) to provide measures of relative rib cage (RC) and abdominal (AB) movement in ten non-ventilated pre term infants. Continuous measurements of oxygen saturation (SaO2), pulse and heart rate (HR), were made and sleep state was recorded using behavioural criteria and electro-oculogram (EOG) measurements. RESULTS: The results showed a significant increase in HR in supine, but no significant difference in SaO2 as a function of position, compared to the prone position where a significant reduction was found in thoraco-abdominal asynchrony for both groups and a reduction in variability in both HR and SaO2. Intra-subject variability of thoraco-abdominal motion as a function of position demonstrated no significant difference on return to supine or on return to prone, illustrating good repeatability of measures. CONCLUSIONS: Prone positioning of pre-term infants recovering from respiratory disease may improve respiratory function. As measured, the improvement in respiratory synchrony in prone position brings pre term infants' breathing pattern into line with that expected in term infants. PMID- 10863717 TI - A comparison of the timing of muscle activity during sitting down compared to standing up. AB - BACKGROUND AND PURPOSE: This study was designed to investigate the temporal pattern of the activation and de-activation of muscles during sitting down to a seated position. The aim was to determine if reproducible patterns existed in the timing of muscle activation and deactivation, in normal subjects, and to compare that produced for sitting down with the pattern produced for standing up. METHOD: The 10 subjects used for the experiment were a convenience sample consisting of healthy male volunteers aged 19-32 years. The design was that of a descriptive study. Subjects were instructed to stand up and sit back down to a seated position. During this activity kinematic (movement) and electromyographic (EMG) data were collected. A pattern of timing for the activity of different muscles or groups was produced and this was compared with the movement occurring. RESULTS: The study found a consistent pattern of EMG activity during sitting down from a standing position. This pattern was documented and the pattern was compared with that produced during standing up. CONCLUSIONS: The results demonstrated that a consistent pattern of muscle activation and de-activation was produced in this group of subjects during sitting down. The pattern of muscle activation and de activation for sitting down was documented. When comparing the pattern produced during sitting down with that produced during standing up it is apparent that some of the muscles were active during both movement patterns at the same relative point in the pattern. PMID- 10863718 TI - Treatment of unilateral upper thoracic vertebral pain using an eclectic approach. PMID- 10863719 TI - Confidence interval for odds ratio. PMID- 10863720 TI - Student research projects--further response from a UK university. PMID- 10863721 TI - [Acquired dysimmune neuropathies. Clinical symptoms and classification]. AB - INTRODUCTION: The neuropathies caused by dysimmunity have seen great changes in recent years. The different forms of clinical presentation, electrophysiological expression, associated anomalies seen on analytical tests, particularly the presence of antibodies to the various antigens of myelin are becoming better understood. This confirms their dysimmune nature and also offers unforeseen possibilities for the comprehension of etiopathogenic mechanisms and possible classifications of specific etiopathogenic factors. DEVELOPMENT: Based mainly on our own experience, in this paper we review current concepts of the three main dysimmune polyneuropathies, the Guillain-Barre syndrome, chronic inflammatory demyelinating polyradiculoneuritis or CIDP and the motor multifocal neuropathies (MMN) with block-conduction or Lewis-Summer syndrome. Regarding the first condition, we particularly emphasize the convenience of establishing the broad classification needed by the variation in its clinical presentation, with regional and functional variants: among the latter we consider particularly the pure motor forms which in most cases are axonal forms with an etiopathogenic basis which is fairly well established and almost constantly associated with the presence of specific antibodies in the serum of patients with this condition. With reference to CIDP, we discuss the existence of atypical forms and the frequency of the relapsing form concerning the evolution. The MMN are the most recently discovered dysimmune neuropathies, according to both the literature and personal experience. We try to establish the difference between pure motor forms and those which also have sensory involvement (or MADSAM) and are called the Lewis-Sumner syndrome. PMID- 10863722 TI - [Relevant antibodies in dysimmune neuropathies]. AB - INTRODUCTION: In the last 15 years a number of autoantibodies against antigens of the peripheral nervous system have been associated, in some cases, to specific clinical features. DEVELOPMENT: Antibodies to MAG or gangliosides have been described in neuropathies associated to monoclonal gammopathy or inflammatory polyneuropathies, such as Guillain-Barre syndrome or multifocal motor neuropathy. A lot of research is devoted to the characterization of known antibodies to glycolipids and the discovery of new ones. Furthermore, experimental models both with animals and in vitro preparations are performed in order to unravel the possible immunopathological role of these antibodies. CONCLUSIONS: It is very important to define the clinical features of these patients precisely in order to establish consistent associations between: presence of antibodies to specific antigens of the peripheral nervous system and clinical syndromes. A deeper knowledge of the antigens and antibodies involved in these neuropathies may be very helpful in the follow-up of these patients and also for future therapies. PMID- 10863723 TI - [Electrophysiological study during the initial stage of the Guillain- Barre syndrome]. AB - INTRODUCTION: In typical Guillain-Barre syndrome the clinical symptomatology is more pronounced in lower extremities. So, this is the territory of election to demonstrate electrophysiological abnormalities early in the disease. PATIENTS AND METHODS: We presented the results of the electrophysiological study of 37 patients, who showed abnormalities in peroneal nerve territory as soon as 15 days from the onset of the disorder, and were maintained at least up to 30 days later. Since Guillain-Barre syndrome is an heterogeneous syndrome, which can differ in extension and severity in the different patients, is necessary a more extensive electrophysiological study. Furthermore, subtypes of the disorder are recognized in relation to different electrophysiological patterns. Therefore, it is important to analyze data of each patient individually and to make control evolutive studies. For these reasons we also describe in detail the electrophysiological study of a selected patient affected by Guillain-Barre syndrome. In this case motor and sensory abnormalities were localized in distal nerve segments, and there was not denervation potentials in EMG. RESULTS: We considered these findings indicative of a mainly distal sub-pattern with a good prognosis. The occurrence of A-waves was an early finding in this case of Guillain-Barre syndrome. During the course of the disease the behavior of these A waves was variable in the different nerves of the lower extremities, and they disappeared in parallel with the clinical improvement of the patient. PMID- 10863724 TI - [Multifocal motor neuropathy]. AB - INTRODUCTION: The multifocal motor neuropathy is an immunological disease that courses with an asymmetrical distal weakness, usually predominant in the upper limbs. DEVELOPMENT AND CONCLUSIONS: It is not rarely misdiagnosed as a motoneurone disease because of the frequent occurrence of fasciculations and cramps and the absence of sensory symptoms. Neurophysiological studies are essential for the proper diagnosis, disclosing the presence of conspicuous alterations of the nerve conduction that are mainly of demyelinating type and occur exclusively in the motor fibers. The nerve conduction blocks characteristically found in this disease are long lasting, multifocal but preferentially proximal, and they are out of the typical levels of the compression neuropathies. High titers of IgM antibodies, mainly anti-GM1, are frequently observed although their pathogenetic significance has not still been completely established. In many cases, even in those with very prolonged evolutions, treatment with high doses of parenteral immunoglobulins has been effective. PMID- 10863725 TI - [Chronic demyelinating auto-immune acquired neuropathy: Lewis-Sumner syndrome]. AB - INTRODUCTION: Knowledge of the chronic auto-immune acquired neuropathies has progressed rapidly since the recent description of the Lewis-Sumner syndrome and multifocal motor neuropathy. Some of the most interesting aspects of these conditions are the presence of persistent conduction blocks and the challenge of understanding the mechanisms involved. These blocks are difficult to identify. CLINICAL CASE: We describe the case of a female patient with Lewis-Sumner syndrome, with persistent sensory and motor blocks. The electrophysiological characteristics and differential diagnosis of the two conditions are reviewed. PMID- 10863726 TI - [Neuropathies and sensory neuropathies]. AB - INTRODUCTION: In the recent years several clinical syndromes have been recognized that are purely sensory in character and therefore the site of lesion is seemingly restricted to nerve cell bodies in the posterior root ganglia. Collectively, these disorders are referred to as sensory neuronopathies. Although they may present as distinct clinical entities with different etiologies, the severity of the clinical manifestations and outcomes varies and may overlap with those of sensory neuropathy. This similarities are better understood in the case of toxic sensory neuropathies where a dose-dependent response has been clearly demonstrated; experimentally, low doses of pyridoxine induce sensory neuropathy whereas higher doses cause neuronopathy. PMID- 10863727 TI - [The bereitschaftspotential. The concept and methodology for acquiring it]. AB - INTRODUCTION: The movement of extension of the index finger in a stereotyped manner, not as a reaction to any external or internal stimulus, is preceded by EEG activity representing activity of the supplementary motor area of the motor cortex. DEVELOPMENT: By averaging retrograde EEG activity prior to movement, the Bereitschaftspotential (BP) may be obtained, using the German nomenclature of the earliest investigators who first described it and its components NS1 and NS2. We include a table with normal values obtained from a group of healthy persons. CONCLUSION: In our study we show the usefulness of off-line methodology with manual adjustment of the start of EMG activity to obtain better morphological definition and greater voltage of the BP. At the same time, continual acquisition with the trigger adjusted to a prefixed threshold (on-line methodology) means that a smaller BP is obtained, especially in persons with difficulty in the organization of this movement. PMID- 10863728 TI - [Neurobiology of pain]. AB - INTRODUCTION AND DEVELOPMENT: An injury to the skin or to an internal organ evokes a discharge in the nociceptive afferents that innervate the damaged area and, as a consequence of the ensuing inflammatory process, sensitizes these nociceptive endings. The activity of sensitized nociceptors evokes two different alterations of pain sensation: 1. A change in the modality of the sensation evoked by low threshold mechanoreceptors, from touch to pain (allodynia) and 2. An increase in the magnitude of the pain sensations evoked by mechanically sensitive nociceptors (hyperalgesia). Allodynia and hyperalgesia show that pain sensation is a dynamic process whose presence and intensity depend on the past history of an affected area and not only on the magnitude of the stimulus. During the initial injury and for the duration of the repair process there will be increased nociceptive activity from the injured region. These afferent barrages cause, in turn, central changes in excitability mediated by positive feedback loops between spinal and supraspinal neurones and by the enhanced synaptic actions of certain neurotransmitters. Among these transmitters, attention has currently focused on the actions of NMDA receptors and neurokinins as putative mediators of the increases in central excitability induced by noxious stimuli. CONCLUSION: The overall mechanism combines the features of the classical 'pain pathway' with the dynamic plastic changes mediated by non-conventional neurotransmitters. PMID- 10863729 TI - [Complex regional pain syndrome: the need for multidisciplinary approach]. AB - INTRODUCTION AND DEVELOPMENT: The title complex regional pain syndrome is being introduced to cover the painful syndromes which formerly were described under the headings reflex sympathetic dystrophy and causalgia. The pain may be sympathetically maintained or sympathetically independent. The incidence of disease with respect to age shows a peak at 50 years, but may be present at any moment of live. The clinical picture of an affected extremity is characterized by autonomic (vasomotor, sudomotor, edema), sensory (allodynia, hyperpathia, hyperalgesia, hyper or hypoesthesia) and motor symptoms. The diagnosis is generally made by clinical examination and with the use of diagnostic aids, such as plain film radiographs, three-phase bone scan, thermography, and diagnostic sympathetic blockade with local anesthetics, guanethidine test, phentolamine test or ischemia test. With regard to the pathophysiology of the disease, although some investigators suggest that these patients suffer from a psychogenic problem, research with animals with experimental neuropathies has revealed several phenomena like that seen in human suggesting the organicity of the disease. The mainstay of treatment, specially in severe cases, is a multidisciplinary approach. The goal is restoration of normal function. Treatment includes adequate pain relief, complete physical therapy program and psychiatric evaluation. PMID- 10863730 TI - [Anti-algic surgical techniques]. AB - OBJECTIVE: To review all the neurosurgical techniques currently used for the relief of pain. DEVELOPMENT: The techniques described range from the classical operations causing lesions, such as neurotomy, rhizotomy, cordotomy, mesencephalotomy, thalamotomy, hypothalamotomy, cingulectomy, leukotomy and hypophysectomy, to the most modern, sophisticated techniques such as spinal and cerebral neurostimulation with administration of spinal morphine using continuous perfusion pumps. The indications, results and most usual complications of these techniques are described together with the techniques themselves. CONCLUSION: It may be said that with this arsenal, modern medicine can control or alleviate over 90% of drug-resistant pain. PMID- 10863731 TI - [Neurophysiological monitoring in the treatment of pain]. AB - INTRODUCTION: The concept of transynaptic deafferentation secondary to a lesion is the basis of the therapeutic criteria of functional neurosurgery. DEVELOPMENT: Pain due to deafferentation requires clinical neurophysiological techniques for characterization, and when appropriate, for localization of the level of the lesion and the ectopic focus or foci which cause the pain syndrome. However, monitoring therapeutic interventions in the pain clinic is an ever increasing need, and obliges the clinical neurophysiologist to master the range of techniques involved in his specialty, so that he can use the most suitable techniques and methods as required by each condition and/or case. The use of techniques such as micro-recordings of the unitary or multiunitary activity of the nerves or nuclei, intracerebral evoked potentials, nociceptive evoked potentials, reflexology, polysomnography and topography, together with techniques such as percutaneous objective localization of deep nerves, allows quantitative evaluation pre-, intra- and postoperative. CONCLUSION: The development of neuromodulation, and in particular of acute or long-term neurostimulation by use of percutaneous techniques, offers an effective therapeutic option in the field of clinical neurophysiology. PMID- 10863732 TI - [Modifications in sleep with aging]. AB - INTRODUCTION: Literature reviewed shows that aging sleep is characterized by a decrease in the ability to stay asleep, resulting in a more fragmented sleep and a decrease in daytime alertness. It exists also an advance of phase, a shortening of wake-sleep period and a desincronitation of circadian rhythms. DEVELOPMENT AND CONCLUSIONS: Changes in circadian rhythms are associated with a decrease of sleep quality. There are a marked reduction of deep slow wave sleep, sleep spindles are less frequent, less ample and shorter. REM sleep appearance is almost uniform during night and REMs density does not increase toward the end of the sleep period. The sleep-wake circadian rhythm is advanced, the temperature rhythm is advanced and the rhythm of cortisol secretion is also advanced. The GH and melatonin peaks of secretion are decreased in elderly. PMID- 10863733 TI - [Sleep disorders in the elderly. Epidemiology]. AB - INTRODUCTION: Age is a factor determining the physiology of sleep. Independently of the changes due to aging itself, several other factors contribute to the deterioration of sleep in elderly persons. DEVELOPMENT: A high percentage of adults of advanced age have alterations in their sleep at night leading to somnolence during the day. The commonest disorders are difficulty in getting to sleep and in staying asleep. With age, there is an increase in periodic leg movements and sleep apnea; the most relevant parasomnia is the behavior disorder associated with REM sleep. There are also a series of problems, which although not specific to the elderly, have a greater impact on them because of their effect on sleep: lack of adaptation to emotional upsets, poor sleep habits, psychiatric affective disorders, organic disorders, use of drugs (psychotropic or others), nocturnal agitation and falls. Finally, although the problem is recognized as important in medical practice, not all doctors have sufficient resources to satisfactorily treat sleep disorders in the elderly. PMID- 10863734 TI - [Sleep and dementias]. AB - OBJECTIVE: A description of sleep-wake disorders in Alzheimer's disease and Creutzfeldt-Jakob disease, and analysis of their diagnostic usefulness. DEVELOPMENT: Patients with Alzheimer's disease have fragmented sleep, with an increase in the time awake and reduction in deep slow sleep and REM sleep. These changes increase as the disease progresses and cognition deteriorates. In Creutzfeldt-Jakob's disease polysomnographic studies show a cyclical pattern and theta or theta-delta activity. These cyclical changes are modified as the disease progresses and disappear before death. CONCLUSIONS: Alterations in sleep are usual in patients with moderate or severe Alzheimer's disease; these alterations non-specific and insufficient to establish an early diagnosis. In the initial phase of Creutzfeldt-Jakob's disease, there are constant alterations in sleeping waking preceding the periodic complexes seen during waking. Subsequently, as the disease progresses, specific cyclical changes appear. PMID- 10863735 TI - [Insomnia in the elderly: cognitive involvement and therapeutic attitudes]. AB - INTRODUCTION: Sleep is an important biological function, which greatly affects the quality of life at all ages. Sleep influences the alertness and stress levels, psychosomatic diseases and health in general. DEVELOPMENT: The predisposition to insomnia increases with age, illnesses such as heart disease, breathing disorders, cancer, strokes, pain and polyuria often disturb sleep patterns. In addition, the aging process, which directly alters sleep-wake cycles and other cerebral functions, affects the elderly. All the above points can lead to an increase in sleep latency, less total sleep time, increases in early awakening and a greater frequency of daytime naps. In an attempt to improve sleep in the elderly, and thus their quality of life, we can try to prevent stress and psychiatric symptoms by altering life-styles and improve the patients' awareness of the problem. Hypnotic drugs are useful for short periods but for longer ones we prefer techniques which attempt to modify behavior. In chronic patients who need treatment with hypnotic drugs for a long time, therapy must be individualized. PMID- 10863736 TI - [Diagnosis and treatment of the phobia due to treatment with air using nasal continuous pressure]. AB - INTRODUCTION: The obstructive sleep apnoea syndrome (OSAS) is characterized by destructured sleep due to repeated episodes of obstruction of the superior respiratory tract during sleep. Treatment by nasal continuous positive airway pressure (n-CPAP) using air is one of the most common forms of treatment of OSAS, although some patients cannot tolerate it. One of the causes of intolerance of n CPAP is the phobic reaction of some patients. OBJECTIVE: The objective of this study is to show that phobia is one of the causes of n-CPAP not being accepted by OSAS patients and demonstrates the efficacy of treatment by exposure in cases of phobic fear of mask. Patients and methods. We studied six patients with phobic intolerance of CPAP and established the diagnosis of phobia according to the criteria of DSM-IV. Subsequently, they were treated by real-life exposure, one of the behavior techniques most often used for phobic disorders, and consisting of exposure to the feared object under conditions in which escape or avoidance is impossible. CONCLUSION: Following the diagnosis of phobia of the CPAP mask, it was seen that treatment involving real-life exposure was effective. PMID- 10863738 TI - The teaching of anatomy. A dialogue with a cell biologist. PMID- 10863737 TI - [Accidents prevalence in a group of patients with the narcolepsy- cataplexy syndrome]. AB - INTRODUCTION: Recent studies have shown statistically that the proportion of accidents at work, at home and on the roads is greater in narcoleptic patients than in the general population. OBJECTIVE: The object of this study is to show the high prevalence of the risk of accidents in patients diagnosed as having the narcolepsy-cataplexy syndrome. PATIENTS AND METHODS: We studied a group of 35 patients of both sexes, diagnosed in the Sleep Pathology Unit of our hospital between 1994 and 1998 as having the narcolepsy-cataplexy syndrome. They filled out a questionnaire to find the prevalence of accidents. The data obtained regarding the type and number of accidents, their causes, gravity, legal and economic consequences were compared with those obtained in a group of 25 healthy subjects using the chi squared test. RESULTS: The results showed a higher risk of accidents in narcoleptic as compared with normal persons, with a statistically significant difference between the two groups. CONCLUSIONS: This study has shown the greater risk of accidents in these patients and the need for diagnosis as soon as possible so as to establish suitable treatment and thus improve their own personal safety and that of those around them. PMID- 10863739 TI - To the editor-in-chief, Surgical and Radiologic Anatomy. PMID- 10863740 TI - Serial anatomy of the larynx in MRI: MRI-histologic correlations. AB - The larynx is an organ with a complex anatomic structure. MRI allows the performance of sections in the three planes of space, so that this study of the soft parts of the larynx yields results superior to those of other imaging techniques. Together with laryngoscopy, MRI is most often used in assessing the extension of malignant laryngeal tumors. This assessment is fundamental in choosing the indications for surgery, but the published reports of MRI of the larynx are sometimes discordant. The visualization of certain important anatomic structures such as the conus elasticus is uncertain. Our aim was to study the MRI radio-anatomy of the larynx based on correlations between MRI and histologic sections. Eight anatomic specimens were studied: four in the transverse plane, two in the sagittal plane, and two in the frontal plane. The MRI and histologic sections made at the same levels were compared. These comparisons allowed a description of the sectional radio-anatomy of the larynx and an assessment of the reliability and limitations of MRI. All the major anatomic structures could be identified. It was possible to demonstrate the conus elasticus. We were able to visualize the vocal process of the arytenoid cartilage, which has not to our knowledge been previously described in the literature. PMID- 10863741 TI - The structure of the cartilaginous end-plates in elder people. AB - Low back pain is the leading cause of work-related disability. Degeneration of the intervertebral disc (IVD), the boundaries of which with age-related changes remain obscure, is considered to be its most important cause. The cartilaginous end-plate (CEP) is the anatomic boundary of the IVD. Since the latter is avascular in adults, the CEP is supposed to play a key role in the metabolism of the IVD. Consequently, it has been postulated that the decrease in permeability of the CEP is the main cause of degeneration of the disc. However, the permeability depends at least partially on the morphologic state of the CEP. Little is known about the age-related changes of the CEP compared to those of the IVD. The objectives of the study were to examine the CEP at different ages, to classify the age-related changes in both the CEP and IVD, and to compare them. The intervertebral discs of the five levels of ten human lumbar spines were collected from cadavers aged from 47 to 78 years, and studied macroscopically and microscopically. Morphologic features of the CEP (thickness, IVD/CEP length ratio, degree of calcification, marrow contacts, CEP-VB and CEP-IVD separations) were measured. Morphologic grades were assigned for both the CEP and the IVD. No significant differences were found with regard to these features in the different age-groups. On the other hand, no significant correlation was found between the morphologic grade of the CEP and that of the IVD, suggesting that the importance of the CEP in disc degeneration may be debatable. PMID- 10863742 TI - Vascular anatomy of gracilis muscle: arterial findings to enhance graciloplasty. AB - Dynamic graciloplasty has recently been developed for reconstruction of anal function in patients who are fecally incontinent in preference to permanent abdominal colostomy. Since the muscular portion of gracilis is wrapped around the neoanus, the length of the gracilis arc influences the functional outcome of graciloplasty. Although dissection of the main pedicle (i.e. the main artery and vein) can facilitate gracilis to have enough muscle arc, it has been unclear whether there are any vessels proximal to the main pedicle or through the origin of the muscle which could support blood flow into the whole of gracilis. In this study, the vascular anatomy of gracilis in both legs of 26 Japanese cadavers was examined. All muscles had a main pedicle, mean maximum diameter 1.08 mm, entering at the proximal one-third of the muscle. However, only 18 muscles (34.6%) had an accessory artery in the proximal portion in addition to the main pedicle. Some arteries always exist at the origin of the muscle, having a mean maximum diameter of 0.34 mm, suggesting that they might be able to support the whole gracilis without supply from the main pedicle. PMID- 10863743 TI - A surgical approach to the ventral aspect of the lumbar vertebrae in the sheep model. AB - The sheep model is frequently used for pre-clinical trials of spinal implants and vertebral interbody fusion devices. The lack of a well-documented multisegmental approach to the ventral aspect of the lumbar vertebrae has limited these trials to a posterior approach to the spine. A retroperitoneal approach to the sheep lumbar vertebrae was established and tested. One hundred and five sheep underwent the surgery. No major complications are reported. Major anatomical differences between sheep and humans were observed and documented. Anatomical variations in the sheep segmental vessel anatomy were also observed. Comprehensive knowledge of the retroperitoneal approach in sheep will facilitate pre-clinical studies testing ventral spinal implants or fusion techniques. PMID- 10863744 TI - Computer-based sessions in radiological anatomy: one year's experience in clinical anatomy. AB - In the last curricular review (1995/96) radiological anatomy was introduced as an innovation in the program of the course of clinical anatomy of the Medical School. Since computer-based media are known to facilitate the understanding of the human body, computer technology was selected in the academic year of 1997/98 as an elective educational tool to teach radiological anatomy. CD-ROMs were introduced as additional instructional resources in 1997/98. This technology aimed to provide educational support to the program, namely, to the sessions of radiological anatomy in each section of the course: head, neck, thorax, abdomen, pelvis and perineum. A questionnaire was designed to evaluate the opinion of the students enrolled in this course, focusing on the teaching sessions of radiological anatomy. Of 152 students, 135 (88.8%) returned the questionnaire. To describe the relationship between the value of this technology and several aspects of its organisation and adequacy, the Spearman rank correlation coefficient was used; canonical correlation was used for the various practical sessions. The comments of students were very positive emphasising the quality of the media, organisation of the course, immediate feedback, degree of interactivity and simplicity of use; they suggested a larger facility for the computers and acquisition of more programs and hardware. The positive evaluation of the use of the CD-ROMs in clinical anatomy allows us to foresee the formal integration of these instructional tools in the whole course, and not to restrict its use to specific units within the course. PMID- 10863746 TI - The congenital anastomoses between hepatic arteries: angiographic appearance. AB - The purpose of this study was to evaluate congenital anastomoses between hepatic arteries demonstrated on angiography in ten patients and to correlate the anastomosis with types of hepatic arterial anatomy. We evaluated the types of the hepatic arterial anatomy based on Michels' classification for 720 patients and compared the anatomic types between the patients with the anastomoses (ten patients) and without the anastomoses (710 patients). The diameter of the anastomoses ranged from 1.5 to 3.0 mm (mean, 2.4 mm). Five anastomoses were classified as tortuous type and five as straight type. Based on Michels' classification for types of hepatic arterial anatomy, eight (80%) of ten patients with the congenital anastomoses were classified as type III (replaced right hepatic artery from superior mesenteric artery). The remaining two patients were classified as type IV (replaced right hepatic artery from superior mesenteric artery and replaced left hepatic artery from left gastric artery) and type VIIIa (replaced right hepatic artery from superior mesenteric artery and accessory left hepatic artery from left gastric artery). Eight (16%) of 48 patients who were classified as type III have the anastomoses. In conclusion, the congenital anastomoses were observed especially in patients with replaced right hepatic artery from superior mesenteric artery. PMID- 10863745 TI - Imaging the coronary venous drainage system using electron-beam CT. AB - This study describes the appearance of the coronary sinus and its tributary veins as visualized on ECG-triggered electron-beam computed tomography (CT) and investigates their spatial relationship to other cardiac structures. Thirty-two patients were examined with ECG-triggered electron-beam CT (exposure time: 100 ms, slice thickness: 1.5 mm) after intravenous contrast agent administration. The entire heart was imaged; the appearance of the coronary sinus and its tributary veins were evaluated. In all 32 patients the anterior interventricular vein and the posterior interventricular vein drained into the coronary sinus. The small cardiac vein was visualized in five patients, a posterior vein of the left ventricle in three and the left marginal vein in eleven. The coronary sinus of all 32 patients had a average length of 30 mm +/- 10 mm (mean +/- SD), range: 21 40 mm and a diameter of 9 mm +/- 5 mm (mean +/- SD), range: 4-14 mm. The results of our work show that if the entire heart volume is scanned using ECG-triggered electron-beam CT, the delineation and the differentiation of the major cardiac veins is possible on transverse cross sections in a way which corresponds to the anatomical literature. Hence to the similar enhancement and similar diameter of coronary veins and arteries on contrast-enhanced electron-beam CT studies, the radiologist should be familiar with the cross-sectional anatomy of the major cardiac veins to prevent possible misinterpretation. PMID- 10863747 TI - Lymphatic drainage of heart and lungs: comparison between pig and man. AB - In its anatomy and physiology the pig is comparable with humans and its organs can be considered for xenotransplantation. We have studied the lymphatic drainage of the heart and lungs in 15 pigs. A coloured mass was injected into the myocardium and/or beneath the visceral pleura. The first nodes coloured were directly injected again. No lymph node was observed inside the heart and lungs. The first lymph nodes coloured were the peritracheobronchial nodes. There was no node in front of the thoracic trachea (Barety's compartment in man). Left suprabronchial nodes were connected with the thoracic duct in the mediastinum. The lymphatics of the heart and lungs in the pig are similar to those of human. Phylogenesis explains "skipping" metastases and the significance of N1 disease in lung cancer, as well as chylothorax occurring after heart and lung surgery. PMID- 10863748 TI - An accessory belly of the abductor digiti minimi muscle: a case report and embryologic aspects. AB - Accessory fasciculi of the hypothenar muscles have been involved in vascular and nerve compressions. During a routine dissection an accessory belly of the abductor digiti minimi muscle arising from the tendon of the palmaris longus muscle was found in the lower third of the forearm. The accessory fasciculus ran through Guyon's canal enclosing the ulnar nerve and vessels. It was attached by means of two tendons where the fibres of the abductor digiti minimi muscle ended in a single pennate form. This anatomic variation was associated with a marked reduction of the caliber of the fourth tendon of the flexor digitorum superficialis muscle and a split of the median nerve. The nerve supply arose from the ulnar nerve. A fibrous band originating from this accessory muscular belly was found covering the median nerve. Based on the development of muscles and fibrous structures within the hand and forearm, as well as on our results, we consider the present anomalies as an unusual persistence of an undifferentiated group of mesenchymal cells. These belong to the superficial muscular anlagen layer of the hand, just between the flexor digitorum superficialis muscle blastema (which has the capacity of migration) and that for the abductor digiti minimi muscle. PMID- 10863749 TI - Insertion abnormality of bilateral pectoralis minimus. AB - Bilateral insertion abnormality of pectoralis minimus (sterno-costo-coracoidian muscle) muscle was examined. The variant muscle was lying under the pectoralis major muscle and was medial to the pectoralis minor muscle. This muscle started from the first costal cartilage to the manubrium sterni and ended in the upper surface of the shoulder joint on the right side. On the opposite side, it took origin from the second costal cartilage to the manubrium sterni and the second costochondral joint, afterwards became a tendinous structure and divided into two on the coracoid process. The thicker part ended on the upper surface of the articular capsule of the shoulder joint, the thinner part inserted on the lateral third of inferior part of clavicle and fascia of subclavius muscle. PMID- 10863750 TI - Pelvic kidney with an unusual blood supply: angiographic findings. AB - A right pelvic kidney was observed in a patient, who presented with hypertension. On angiograms, the left kidney was normally positioned and had a single renal artery, whereas the right pelvic kidney received three arteries, which arose from bilateral common iliac arteries and from ipsilateral internal iliac artery. The renal arteries from the ipsilateral internal iliac artery and the contralateral common iliac artery supplied the medial half of the pelvic kidney. In the present case, the blood supply from both the right and left sides appeared to be related to the medial position of the right pelvic kidney. As the incidence of unilateral renal ectopia is not extremely low, it is possible to encounter in a surgical or cancer treatment case. Variations in the positional anatomy of the kidney and its vascular supply are of clinical importance and our case illustrates a different kind of blood supply that a pelvic kidney may possess. PMID- 10863751 TI - A variation of the brachial plexus characterized by the absence of the musculocutaneous nerve: a case report. AB - A variation of the brachial plexus characterized by the absence of the musculocutaneous nerve on both sides was observed during the dissection of a 72 year-old female cadaver. The long thoracic nerve included only the fibers from C5 and C6 on the left side. The musculocutaneous nerve was absent and two branches from the lateral cord innervated the coracobrachialis muscle. The median nerve innervated the biceps brachii and brachialis muscles in the arm and also gave off the lateral antebrachial cutaneous nerve. Additionally, a communicating branch was found from the median nerve to the ulnar nerve in the forearm. The knowledge of the anatomical variations of the peripheral nerve system can help give explanation when encountering an incomprehensible clinical sign. PMID- 10863752 TI - [The situation of physician in the state prison--constitutional state guarantees of individual rights]. AB - In Germany, 220,000 persons are imprisoned yearly. Their health care is provided by physicians supported by medical teams in 9 prison hospitals. There is a total of 395 physicians mainly practicing in general medicine, internal medicine, surgery, and psychiatry. Other medical disciplines are represented by external physicians that are paid by fees. It is common to consult external physicians which can be explained by the legislation ruling the penal system. The prison administration has to provide a health care of equal quality as outside of prison. A pathology typical in prison, the unproportional representation of certain social groups (poor people, foreigners) and diseases (mental diseases, drug abuse, tbc, hepatitis, HIV) request an expanded profile of expertise from the physician. Except the task of health care, the physician is included in executive functions. Thus, the physician is exposed to control by non-medical institutions. PMID- 10863753 TI - [Instrumentation of full medical care for non-medical purposes]. AB - A survey of European standards and recommendations is given defining the status of prison physicians in the 40 member states of the Council of Europe. Taking into account these standards, the two main tasks of prison physicians in Germany are the following: 1.) Prison physicians have to treat and, if possible, cure patients--just as physicians outside the prisons do. 2.) By expressing their opinion and by taking initiatives, prison physicians have to contribute, to the observance of health standards in connection with the determination of confinement conditions (hygiene, cleanliness, physical exercise, ventilation, nutrition) and to develop them further. They also have to raise the prisoners' awareness of the fact that they should care about their health. These tasks are differentiated from requests that are unduly made to prison physicians by external authorities. Prison physicians are not court experts and the duties which they have to perform as physicians in charge of the treatment of prisoners do not include the rendering of opinions on questions relating to a person's fitness to undergo detention or to stand trial. This task is incumbent on experts, whom the courts have to find and appoint elsewhere than among prison physicians. Decisions regarding the determination of specific confinement conditions in a particular case or the granting of privileges to prisoners have to be taken by the prisons' managerial staff or the administration and not by the prison physicians, who, however, are frequently called in either by prisoners for the purpose of asserting alleged claims or by the prisons' managerial staff for the purpose of rejecting claims which the staff regard as unjustified. PMID- 10863754 TI - [Professional discretion versus duty to inform of physicians in prisons--from the view of trial lawyers]. AB - The view of the trial lawyer is the view of the client--meaning the prisoner- which is treated by a physician in a closed institution. Firstly, this view is influenced by prejudice and distrust and a relationship of trust has to be built. According to section 182 of the Code of Penalty Execution, the physicians in a penal institution are bound to professional discretion. The difficulties of physicians and therapists in a closed institution are demonstrated. PMID- 10863755 TI - [Medical discretion versus the duty to inform of physicians in prisons--from the view of the prison physicians]. AB - The basic principle of medical discretion within the prisons and penal organs is regulated under section 182 StVollzG. It is advisable for the physicians as well as for the members of penal organs to handle the principle of medical discretion in prison with a great care and hence to encourage a better understanding and cooperation between the physicians and the patients in custody. The physicians should have the responsibility to administer the adequate diagnostic, treatment and meet relevant decisions within the existing regulations. The primary medical examination within the first 24 hrs in custody should be regarded as the central issue of the health care for the prisoners. The results of the examination should as well be under the above mentioned medical discretion. The section 182 StVollzG should be explained in a simplified and understandable form prior the primary medical examination has been done. PMID- 10863756 TI - [Medical discretion versus duty to warn by physicians in prisons-- view of the prison governers]. AB - We appreciate that the limits of medical discretion in penal execution have been clarified by the legislature with the law issued at August 26, 1999. From the point of view of a head of an institution, this law makes it possible to achieve acceptable results in any relevant case-combinations. The "circumspect head of an institution" should, as far as his general responsibilities allow, make use of his right to be informed in a very restricted way. This right is given by the particular conditions in penal execution. The implementation of this law within the institution requires the following: To inform the prisoners on the existing obligations and rights of information. To inform physicians working within institutions on a general level about existing duties without making provisions for particular cases. To name the recipients of information from the medical sector. PMID- 10863757 TI - [Mental illness in prisons--view of forensic psychiatrists]. AB - To the extent to which comparisons have been undertaken with the general population, a greatly raised prevalence of psychiatric illness amongst prisoners has been found across countries and across diagnostic groups. Thus, one would expect a greater level of need for treatment in the penal system. In accordance with the conditions of modern psychiatric care, cooperation between impatient, part-impatient and outpatient services in the sense of integration should be guaranteed. PMID- 10863758 TI - [Mental disorders in prisoners--view of the district attorney]. AB - This article deals with the question when a mentally ill prisoner can be granted an interruption of the penalty according to section 455 of the Code of Criminal Procedure and what is its relation to a treatment in an external hospital according to section 65.2 of the Code of Penalty Execution. Additionally, the different status of the convict combined with the different legal consequences included in these two procedures, the permission of compulsory measures and the responsibilities of such procedures are explained. PMID- 10863759 TI - [Mental disorders in prisons--view of prison governors]. AB - This article reports on experiences with prisoners with mental diseases. Due to the multitude of drug supply of all kind in Frankfurt, several addicted humans are living in this area. Their way of life is frequently accompanied by arrest and imprisonment. It is demonstrated what we experienced with these prisoners and what kind of problems has occurred. The employees of a penal institution are not prepared enough to deal with such problems. Such demands are demonstrated. PMID- 10863760 TI - [Problems of addiction behind prison walls--experiences from prison medical practice]. AB - The difficult task of medical practice behind prison walls is illustrated by examples. The medical practitioner is often challenged by the task of combining ethics, law, the needs of the client, public opinion and political policy without getting much recognition and/or support. Co-morbidity of addiction and psychological-psychiatric disorders is an additional problem. Often it appears as a "hen or egg" problem, what was first? Clients needing psychiatric treatment often remain imprisoned in care with the medical practitioner because of the difficult circumstances mentioned above. Almost like the medical practitioner finding himself "imprisoned" with the clients managing the situation as good as possible (or not) with limited resources. PMID- 10863761 TI - [Drug abuse problems behind prison walls--view of the trial lawyers]. AB - This article reports problems of drug abuse in prison from the view of the trial lawyer. Alcohol abuse in prison is demonstrated in more detail since alcohol is the oldest drug in these institutions. Options of treatment and drug withdrawal are explained. PMID- 10863762 TI - [Addiction problems behind prison walls--view of the prison administration]. AB - As Head of the Prison Administration of Saxony, the author describes the difficulties and problems that exist in the care and treatment of prisoners who are addicted to drugs or alcohol. Up to now, these problems have been dealt with in a manner that was too much concentrated on ideas and aspects of security by using systems of control and restrictions. Social contacts inside and outside of the prison and a sense of freedom are, however, the requirements of the legal concept of resettling prisoners. There is a great need for more counselling and therapy. It should also be attempted to improve the conditions for the individual prisoners, e.g. by setting up drug-free units and, thus, provide a environment to the addicts that enables them to live their lives without the daily struggle for drugs and alcohol. PMID- 10863763 TI - [Revision of section 35 a SGB VIII--critical developments, possible solutions]. PMID- 10863764 TI - [Personality disorders and psychiatric morbidity in adolescent anorexia nervosa. Results of a prospective 10 year catamnesis]. AB - The aim of the current prospective study was to examine at regular intervals the course of the eating disorder symptoms and the psychiatric (co-) morbidity including personality disorders among juvenile patients who fulfilled the DSM-III R criteria for anorexia nervosa. Ten years after release from hospital all 39 patients (100%), as well as a control group parallelized for age, gender and occupational status were personally followed-up. Symptoms of eating disorders were documented by means of the Standardized Interview for Anorexia and Bulimia nervosa (SIAB, Fichter et al., 1991), the Composite International Diagnostic Interview (WHO, 1990) was applied to diagnose psychiatric (co-) morbidity, and the Structured Clinical Interview (SKID-II, Spitzer et al., 1993) to assess personality disorders. Compared to the control group, at the time of follow-up a significantly greater number of patients were suffering from a psychiatric disorder, primarily an anxiety disorder, an affective disorder or from drug, respectively alcohol abuse. Personality disorders, chiefly anxious-avoidant types on the DSM-III-R were diagnosed among almost one-fourth of the patients. Our findings indicate that anorexia nervosa is not a developmental disorder limited to puberty but a disorder associated both cross-sectionally as well as longitudinally with other psychiatric disorders. PMID- 10863765 TI - [Psychosocial stress and its importance for behavioral deviations in juveniles from migrant and German families]. AB - OBJECTIVES: Comparison of the familial, socio-economic and cultural factors among adolescents from German and migrant families; analysis of the risk factors for conspicuous behavior among the adolescents. METHODS: 224 graduates of secondary schools in the Kreuzberg district of Berlin replied to three questionnaires on living situations, psychosocial stress and conspicuous behavior. RESULTS: The living situation of the adolescents from migrant, above all Turkish families as compared to their German counterparts, is characterized by psychosocial disadvantage, but also by a greater stability within the family. Cultural differences are also noted. Risk factors in conjunction with conspicuous behavior included in addition to familial and socioeconomic factors, above all chronic illness, and persecution and discrimination, whereby chronic illnesses were most significantly frequent among the German, while persecution and discrimination were most significantly frequent among the Turkish adolescents. Cultural differences posed no risks for conspicuous behavior. PMID- 10863766 TI - [Tourette's syndrome and antidepressant therapy: exacerbation of nervous tics with paroxetine]. AB - One possible side effect of selective serotonin re-uptake inhibitors (SSRI) is the exacerbation of nervous tics in a 12-year-old boy treated with tiapride following prescription of paroxetine for a depressive syndrome. Potential causal factors include residual cholinergic activity of paroxetine, the observably increased drive under paroxetine, metabolic properties, and protein binding. The problem of side effects under selective serotonin re-uptake inhibitors, as well as the issues of co-morbidity and co-medication in the treatment of nervous tics and Tourette's Syndrome are discussed. PMID- 10863767 TI - ["Cyclic vomiting" in childhood and adolescence]. AB - The clinical picture of the "cyclical vomiting syndrome" is discussed in the case of a 5 1/2-year-old child. The core symptom of this recurrent disorder is vomiting that occurs in regular cycles and while the patient is in a state of complete well-being, and which is hard to influence. The symptoms, the differential diagnosis, and the therapy of this disorder, which occurs primarily in childhood, are discussed along with the close etiological relationship to infantile migraine. The diagnosis of "psychogenic vomiting" is addressed critically, since it is frequently applied in Germany to disorder pictures that are similar to cyclical vomiting. PMID- 10863768 TI - [Reference tables for representation of ICD-10 diagnoses in DSM-IV]. AB - Proceeding from the Clinical Descriptions and Diagnostic Guidelines of the ICD 10, a table of reference to DSM-IV is presented for mental disorders. Emphasis is placed upon childhood and adolescent mental disorder pictures. Allowance is made in the corresponding notes for greater content differences in the operationalization of individual disorder pictures. However, the table can only serve as a pointer to diagnoses in DSM-IV that correspond to those in ICD-10. In the interests of greater diagnostic security, these should be checked in individual cases against the diagnostic criteria. In the forefront certain theoretical considerations with regard to the comparison of the two classification systems are illustrated by means of selected disorder pictures. PMID- 10863769 TI - [Radiotherpay and immediate breast reconstruction with myocutaneous flap in breast cancer of reserved prognosis]. AB - In France, immediate breast reconstruction (IBR) for infiltrating carcinoma remains controversial. Many teams advocate the possible event of a post mastectomy radiotherapy and its negative effect on IBR. In our Institute we do not exclude infiltrating breast cancer patient from IBR. In the poor prognostic patients who wish IBR, we recommend autologous IBR to obtain the best aesthetic result with minimum revision procedures and best tolerance to adjuvant radiotherapy. From January 1993 to December 1997, we performed 687 IBR with myocutaneous flap for infiltrating carcinomas. In this group only 68 patients needed postoperative chest wall radiotherapy (45 Gy): 27 TRAM flap, 41 latissimus flap. Only one of the TRAM but 39 of latissimus flaps were associated with a prosthesis. The mean follow-up was 24 months. Fourteen patients developed metastatic disease, and ten were dead at the time of the chart revue. The autogenous TRAM flap tolerate radiation quite well and remain soft and mobile. The latissimus flap associated with a prosthesis developed capsular contracture (BAKER II or III) in 71% of cases. In all cases the cosmetic impairment was not important and the result after capsulectomy remained soft. We concluded that IBR could be offered to motivated patients in all stages of the disease regardless of the subsequent chest wall radiotherapy, and we recommend its use for possible autologous reconstruction. PMID- 10863770 TI - [Tolerance, reliability and efficiency of inflatable breast implants after breast reconstruction. Retrospective study of 101 consecutive cases]. AB - A long-term retrospective study of breast reconstruction with inflatable implants is presented. One hundred and one patients were studied, with a median follow-up of 44 months. Prosthetic implants used were round McGhan implants, model 168. Median volume was 215 mL. Twenty-two patients had contralateral symmetrisation, an average of five months after implant. The advantages of this sort of reconstruction are ease, speed and homogenously good results. Drawbacks include the appearance of prosthetic leaks (7%), waves and folds (13%), stage III and IV capsular contractures (26%), asymmetry and incorrect placement of the implant (25%). Breast reconstruction with implants alone gives good results for specific indications: immediate reconstructions, bilateral reconstructions, no history of radiotherapy, and good quality chest wall tissues. PMID- 10863771 TI - [Contribution to the study of the future of silicon-gel breast implants in mammary reconstruction after cancer: concerning 205 implants]. AB - Long term follow-up of 187 women with mammary reconstruction using 205 silicone gel implants enables a retrospective analysis of the evolution of both patients and their prosthesis. Most patients have not presented local or general disease related to implant leakages. However, these leakages are not uncommon, being present in nearly one-third of 38 explantations motivated by aesthetic considerations (of which 50% are capsular retractions). All the ruptured or pre ruptured implants are more than seven years old. Leakage frequency sharply increases after ten years. Authors tried to assess the place of radiologic evaluations (digitized costal profile, ultrasound, MRI...) in order to detect deterioration of the prosthesis. Although suspicions raised by these evaluations have made it possible to adequately target the explantations, pathogenicity of silicone-gel implants appears low enough to confirm the predominance of clinical follow-up. PMID- 10863772 TI - [Reduction mammaplasty with an arche "pedicle" and vertical scar. Technical note and preliminary report]. AB - In the interest of avoiding the inconveniences of classic techniques like large and long scars, we propose an original procedure based on a supero-posterior pedicle and vertical scar. A preliminary review of 50 cas permit to conclude to reliable results and faithful reproduction. PMID- 10863773 TI - [Prevention of eyelid retraction following lower blepharoplasty]. AB - This review covers the techniques used to prevent lid retraction after lower blepharoplasties. After a brief overview of the lower lid anatomy (anterior, middle, posterior lamella), the causes of blepharochalasis and of lid retraction are addressed. Clinical examinations are described that can detect a candidate at risk. Many preventive measures are discussed, examining patient positioning, approaches (transcutaneous, transconjunctival, combination), incision types, flap dissection (skin, skin-muscle, dermal flaps), suspension techniques (muscle muscle, muscle-periosteum, horizontal wedge excisions, lateral and medial canthal procedures), CO2 laser resurfacing and combinations. These techniques are described and critically appraised. PMID- 10863774 TI - [Orbital filling using a vascularized parietal bone graft: a case report of rehabilitation with an orbital graft of axial implants]. AB - Orbital and intracranial complications of dental cellulitis are exceptional. The authors report such a case with orbital cellulitis and frontal abscess that dictated orbital exenteration with resection of the eyelids. Before realization of an implant supported epithesis, the orbit was filled with a prefabricated vascularized fascio-osseous parietal bone flap. This allowed placement of two extra-oral fixtures (Nobel biocare) in an antero-posterior axis. In some cases, preimplant orbital bone graft appears to be the only solution when implant placement in the orbital rim is impossible or contra-indicated. Indications, advantages and disadvantages of this technique will be discussed. PMID- 10863775 TI - [The second chart. A retrospective critical analysis of hospital stays of long duration in a public plastic surgery department]. AB - A nine-month retrospective survey related to the long-duration hospital stays was carried out in our plastic surgery department. Twenty five patients were concerned by a more than 21 days hospitalization, the sum of which reached 1,098 days. These figures corresponded to 1.4% of the patients and 14.5% of the hospitalization days. A critical analysis was based on four principles: 1--a plastic surgery department is exclusively devoted to plastic surgery, 2--nursing cares required by the healing of a soft tissues defect don't usually need hospitalization, 3--even if it has a wide surface and/or if it is located on the lower limbs, a skin graft doesn't usually require more than ten days of hospitalization, 4--without complications, a free tissue transfer doesn't usually require more than 15 days of hospitalization. Application of these principles showed that 633 days (58%) could have been theoretically spared. Consequently, it could have permitted to treat a greater number of patients. The cause of delaying patient exit was related to the surgeon in all but one cases. It was associated in 16% of cases with a bed shortage in the convalescent or nursing homes. As hospitalization durations longer than 25 days seemed unwarranted to the authors even in the most complex cases, they suggest a simple way to alert surgeons of their department to the long-duration stays. As the department patient's chart represents 13 days of hospitalization, they ask surgeons make a decision upon planning the exiting of patients before adding a second chart. PMID- 10863776 TI - [Open letter to the internists and chief residents of reconstruction and plastic surgery units]. PMID- 10863777 TI - [General principles]. PMID- 10863778 TI - [Medical liability without malpractice according to French laws]. PMID- 10863779 TI - [The expert's opinion]. PMID- 10863780 TI - [The lawyer's opinion]. PMID- 10863781 TI - [Therapeutic hazard: round-table discussion about medical liability without malpractice? Medical responsility]. PMID- 10863782 TI - [Comment on the article: "Tuberous breast syndrome. Report of a series of 22 operated patients]. PMID- 10863783 TI - [On breast implants]. PMID- 10863784 TI - [Vaccination against S. pneumoniae: effective and efficient approach?]. PMID- 10863785 TI - [Epidemiology of acute flaccid paralysis in Italy: 1996-98. Group for the Study of AFP]. PMID- 10863786 TI - [Streptococcus faecalis in the environment: species identification and antibiotic resistance]. PMID- 10863787 TI - [Reorganization of an operating team: protocol draft according to the DPR of 14/1/97]. PMID- 10863788 TI - [Presence of verocytotoxin-producing E. coli O157 in beef hamburgers]. PMID- 10863789 TI - [Preliminary evaluation of the psychosomatic well-being of a population exposed to earthquakes]. PMID- 10863790 TI - [Hygiene-sanitary interventions for the establishment of a refugee camp]. PMID- 10863791 TI - [Asbestos: perspectives]. PMID- 10863792 TI - [Exposure to airborne bacteria in a wastewater treatment plant]. PMID- 10863793 TI - Synthesis and analgesic activity of some side-chain modified anpirtoline derivatives. AB - New derivatives of anpirtoline and deazaanpirtoline modified in the side chain have been synthesized. The series includes compounds 3 with side-chains containing piperidine or pyrrolidine rings, compounds 4 containing 8 azabicyclo[3.2.1]octane moiety, and compounds 5 having piperazine ring in their side-chains. Their receptor binding profiles (5-HT1A, 5-HT1B) and analgesic activity (hot plate, acetic acid induced writhing) have been studied. Optimized structures (PM3-MOPAC, Alchemy 2000, Tripos Inc.) of the synthesized compounds 3 5 were compared with that of anpirtoline. PMID- 10863794 TI - Synthesis of 5-(4-alkoxy-[1,2,5]thiadiazol-3-yl)-3-methyl-1,2,3,4 tetrahydropyrimidi ne oxalate salts and their evaluation as muscarinic receptor agonists. AB - The synthesis and biological testing of 5-(4-alkoxy-[1,2,5]thiadiazol-3-yl)-3 methyl-1,2,3,4-tetrahydro pyrimidine oxalate salts 8 as muscarinic receptor agonists are described. The key intermediate 4 was obtained by modified Strecker reaction and cyclization of starting material 1. Subsequent alkoxy substitution, quaternization, and reduction afforded 7. For the sake of purity and stability of the final products 8, the 3-methyl-1,2,3,4-tetrahydropyrimidines were obtained as oxalic acid salts. All final compounds were examined in vitro for their binding affinities to the cloned human muscarinic receptor by the [3H]-NMS binding assay. PMID- 10863795 TI - Synthesis of new N-(4-pyridyl)-1-aminopyrazoles and their muscarinic and adrenergic properties. AB - The synthesis of new N-(4-pyridyl)-1-aminopyrazoles is described. Their binding properties were tested for muscarinic and other neurotransmitter receptors, together with their acetylcholinesterase inhibitory activity. The series derived from 3,5-dimethyl-1H-pyrazole showed moderate activities in both muscarinic and adrenergic receptor binding tests. PMID- 10863796 TI - Benzimidazoles as NMDA glycine-site antagonists: study on the structural requirements in 2-position of the ligand. AB - A series of different substituted benzimidazole derivatives has been synthesized and evaluated for the ability to displace [3H]MDL-105,519 to rat cortical membranes. Two benzimidazole-2-carboxylic acids 9 b and 9 c, in this substitution pattern not yet described as glycine antagonists, showed IC50 values of 0.89 microM (9 b) and 38.0 microM (9 c). Replacement of the carboxylate function in 2 position by a sulfonic acid moiety appreciably increased solubility, but decreased the affinity giving evidence for the strong need of the carboxylate group within the ligand. Further structure-activity studies using benzimidazole-2 one derivatives with an acetic acid moiety adjacent to a ring nitrogen revealed new insights into the importance of amide functionalities within the heterocycle for the affinity of antagonist glycine-site ligands. PMID- 10863797 TI - New NO donors with antithrombotic and vasodilating activities, Part 29. N-(1 cyanocyclohexyl)-C-phenylnitrones and glyoxaldinitrones. AB - Six N-(1-cyanocyclohexyl)-C-phenylnitrones 4a-f (4b-f for the first time) and 22 glyoxaldinitrones 7a-v were prepared and tested for antithrombotic (p.o. administration to rats, 60 mg/kg) effects. Both classes of compounds exhibit considerable antithrombotic activities. Maximum inhibition of thrombus formation in arterioles (21%) was observed in N,N'-bis-2-phenylethylglyoxaldinitrone (7o) and N,N'-bis-4-nitrobenzylglyoxaldinitrone (7u). The compounds form only small amounts of nitric oxide in vitro by the addition of a Fe(3+)-porphyrine complex and an oxygen donor. PMID- 10863798 TI - Synthesis and correlations between experimental and calculated lipophilic indices of new 1,2-benzisothiazole derivatives with potential antimicrobial activity. AB - Five series of new hydrazones (1a-m, 2a-m, 3a-m, 4a-m, 5a-m) with potential antimicrobial activity were synthesized from cyclic (1 and 4) or acyclic (2, 3 and 5) 1,2-benzisothiazolylhydrazides and characterized. Condensation of the appropriate hydrazide with aldehydes afforded the designed compounds. Aldehydes carrying different hydrophobic substituents were used and the five series were designed so that the hydrophobicity also varied among congeners. Retention parameters were measured by HPLC employing a deactivated alkyl-bonded silica column and different eluent systems (methanol-aqueous buffer, acetonitrile water). The hydrophobicity chromatographic parameters (log k') were compared with those provided by measurement of partitioning of solutes between n-octanol and water (log P), and with theoretical partition coefficients, calculated by a fragmental method and scaled according to the experimental values. Correlations between different hydrophobicity indices are reported and discussed. PMID- 10863800 TI - Sexuality and the white coat. PMID- 10863799 TI - New nonsteroidal steroid 5 alpha-reductase inhibitors. Syntheses and structure activity studies on carboxamide phenylalkyl-substituted pyridones and piperidones. AB - In the search for nonsteroidal inhibitors of 5 alpha-reductase for the treatment of benign prostatic hyperplasia (BPH), we synthesized diisopropyl (1a-8a) and tert-butyl (1b-8b) benzamides, as well as ethyl benzoates (1c, 3c), which were substituted in 4 position via variable alkyl spacer (n = 0: 1-4, n = 1: 5, 7 and n = 3: 6, 8) with a 1-methyl-2-pyridone (1, 2, 5, 6) or a 1-methyl-2-piperidone (3, 4, 7, 8) moiety mimicking steroidal ring A. The directly connected benzamides (1a-4a, 1b-4b) and benzoates (1c, 3c) were obtained by palladium-catalysed coupling reaction of diethyl(3-pyridyl)-borane with 4-bromobenzoic acid derivatives, followed by alpha-oxidation of the 1-methyl-pyridinium salt and subsequent separation of the regioisomers. Catalytic hydrogenation of the pyridones (1, 2) led to the piperidones (3, 4). The preparation of the benzamides with a methylene (5, 7) and a propylene spacer (6, 8), respectively, started with the reduction of the keto group of 5-benzoyl-1,2-dihydro-1-methyl-2(1H)-pyridone and catalytic hydrogenation of the alkene obtained by Wittig reaction of 5-formyl 1,2-dihydro-1-methyl-2(1H)-pyridone with (2-phenylethyl)triphenylphosphonium bromide, respectively. The phenyl ring was functionalized by Friedel-Crafts reaction, haloform cleavage to give the acid, formation of the acid chloride, and subsequent treatment with the appropriate amines. Again, catalytic hydrogenation of the pyridones (5, 6) led to the piperidones (7, 8). The 5 alpha-reductase inhibitory properties were determined using rat ventral prostate, as well as human BPH tissue as enzyme source, 1 beta-2 beta-[3H]testosterone as substrate and a HPLC procedure for the separation of dihydrotestosterone (DHT). Tested at a concentration of 100 microM, the inhibition values of 1-8 ranged from 0-79%. Significant differences were observed between rat and human enzyme. The most active compound was ethyl 4-(1-methyl-2-oxopiperid-5-yl)benzoate 3c (68%) for the human enzyme and N,N-bis(1-methylethyl)-4-[3-(1,2-dihydro-1-methyl-2-oxopyrid-5 yl) propyl] benzamide 6a (79%) for the rat enzyme. PMID- 10863801 TI - Lesbian sexuality. Do GPs contribute to lesbian invisibility and ill health? PMID- 10863802 TI - HIV/AIDS consultations. PMID- 10863803 TI - GPs as 'gatekeepers' for adolescent suicide. PMID- 10863804 TI - Oral medicine--in search of tradition. PMID- 10863805 TI - Psychiatry in general practice. PMID- 10863807 TI - Psychosocial aspects of sexual disorders. AB - BACKGROUND: The sexual urge is as old as humanity itself and necessary for the propagation of all species. However, sexuality is not only influenced by the integrity of the genital tract, but also the limbic system and spinal arousal centres. Social and cultural influences on sexual behaviour are in a continuous process of change. OBJECTIVE: This article attempts to highlight the importance of a multifaceted systemic approach to a common and distressing problem. DISCUSSION: The aetiology of sexual dysfunction is diverse and usually results from a complex interaction of biological, psychological and social factors. Consequently, there is little to be gained from trying to determine whether a disorder is either biological or psychological, as both will be relevant to a certain extent in every case. Even in a case with a clear biological cause (e.g. diabetic neuropathy) the treatment and long term consequences of the secondary sexual disorder will be affected by psychosocial implications. Sexual functioning can be strongly influenced by one's own sense of self and social competence. PMID- 10863806 TI - Chlamydia detection and management. AB - BACKGROUND: Chlamydia trachomatis is a common, often asymptomatic sexually acquired infection with serious sequelae if left untreated. OBJECTIVE: To describe the prevalence, clinical features, investigations, indications for screening, contact tracing and treatment of complicated and uncomplicated infection with C trachomatis. DISCUSSION: The availability of DNA amplification tests with improved sensitivity and the introduction of single dose therapy for the treatment of uncomplicated infection has simplified the management of C trachomatis infection. The general practitioner (GP) is ideally placed to reduce the burden of disease in the population with implementation of a targeted screening program for C trachomatis. However, the GP will need to continue to remain vigilant to the subtle symptoms and signs of unrecognised or asymptomatic infection and ensure contact tracing of sexual partners is actively pursued. PMID- 10863808 TI - Taking a sexual history. AB - BACKGROUND: Significant social changes have resulted in increased exposure to sexual issues and greater tolerance of sexual behaviours such as premarital sex and homosexuality. These changes are not universal, however. We live in a multicultural society with widely varying sexual mores and frequent clashes of culture across generations. The media has contributed to unrealistic expectations about body image and sexual performance while at the same time increasing awareness of sexually transmitted diseases. Consequently, patients seek help with these issues and will discuss them freely if the general practitioner is comfortable in the area of sexual health. OBJECTIVE: This article outlines an approach to sexual assessment and ways of addressing the barriers faced in taking a sexual history. DISCUSSION: All sexual disorders involve biological, psychological and social factors. General practice can provide a safe, non judgmental environment for the patient to divulge sexual difficulties and have these dealt with holistically. However, this takes time, a significant obstacle in current practice. PMID- 10863809 TI - Danger zone. When boundaries are crossed in the doctor-patient relationship. AB - BACKGROUND: The most obvious boundary violations in medicine are sexual. However, the areas of time management, drug prescribing, accepting gifts, friendships with patients and treating friends also pose ethical dilemmas. Where boundaries lie and how to avoid crossing them can be difficult judgments to make. OBJECTIVE: To examine the issue of boundaries in the doctor-patient relationship and to discuss strategies for avoiding and managing boundary violations. DISCUSSION: The broad concepts of respect for autonomy and avoiding harm to patients and doctors by violating boundaries appears self evident. Where these boundaries lie can appear less clear cut in the detail of individual relationships between doctor and patient. Communicating well with patients about the need for clinical examinations and maintaining objectivity in the doctor-patient relationship is important. Avoiding and managing boundary violations involves awareness of potential hazards, tact and good judgment. PMID- 10863810 TI - Leukotriene antagonists. Do they offer new hope for asthmatics? AB - BACKGROUND: Leukotrienes are potent chemical mediators important in allergic inflammation. Leukotriene receptor antagonists are a new class of oral asthma drugs which target and block the action of these mediators. OBJECTIVE: To review the action of leukotrienes and the clinical effects of leukotriene receptor antagonists in asthma. DISCUSSION: Leukotrienes mediate bronchospasm, airway oedema, mucus hypersecretion and increased airway reactivity. Leukotriene receptor antagonist drugs have a mild short and long term bronchodilator effect, with evidence of an anti inflammatory effect. The clinical benefits include improved symptoms, reduced rescue bronchodilator requirements, and reduced inhaled steroid requirements. Their oral formulation may provide improved adherence compared to inhaled medication. Clinical studies suggest they may be less efficacious than inhaled steroids or long acting beta 2 agonists in improving lung function and symptom control, but there was a heterogeneity in response to all classes of asthma drugs. Hence, currently, the only way to judge effectiveness is a therapeutic trial for 4-6 weeks. Although they are unlikely to replace currently available asthma medication, they are likely to be useful adjuncts to treatment. PMID- 10863811 TI - An unusual cause of pneumonia. PMID- 10863812 TI - The female condom. AB - BACKGROUND: The female condom has been available in numerous countries for some years. It was launched in Australia on March 8th, 2000 and is an important addition to the range of barrier contraceptives. OBJECTIVE: This article aims to provide an overview of the female condom to enable GPs and nurse practitioners to advise clients and provide information about its use. DISCUSSION: The female condom is an effective form of contraception. When used correctly it has a failure rate of about 5%, which compares favourably with other forms of barrier contraception. It is important that users are instructed on insertion, in particular the need to guide the penis into the condom. Evidence shows that it reduces the incidence of sexually transmitted infections. There may also be additional benefit in reducing wart virus and herpes transmission, as there is greater protection to the vulva and the base of the penis, when compared with the male condom. The female condom has high user acceptability, and offers advantages in terms of sensitivity and also ease of use by men with erectile dysfunction. It is the only barrier protection under a woman's control, giving protection to the vagina and vulva as well as the cervix. PMID- 10863813 TI - Overweight and hypertensive. AB - BACKGROUND: Hypertension and obesity frequently coexist in the same patient, with both conditions attracting considerable morbidity. There is also an established correlation between increasing weight and increasing blood pressure. OBJECTIVE: This article provides an overview of the interaction of these two conditions and provides some recommended strategies for dealing with them when they co-exist. DISCUSSION: In dealing with a hypertensive patient whom you consider overweight it is important to establish what their body mass index is, whether they are aware that they are overweight and if they understand the type of problems that may be associated with this. It is also useful to confirm that the patient is in fact hypertensive by checking the way the measurement was taken. Behavioural change is an extremely difficult undertaking and patients need to be ready to change before advice is likely to be heeded. Once a patient is at this stage, having a strategy in which to assist weight loss is more likely to see an effective outcome. PMID- 10863814 TI - Weight loss. The role of low fat diets. AB - BACKGROUND: Weight loss is theoretically simple: decrease energy intake, increase energy use, or both. As millions of 'yo-yo' dieters can attest, translating this into practice is far from simple. OBJECTIVE: This review will explore the issues surrounding the low fat approach to weight loss and suggest sensible and alternative recommendations for patients. DISCUSSION: Although short term weight loss can be achieved by many diets, sustained behaviour change and long term weight maintenance is far more difficult. Hence the essence of any weight intervention is a change in diet and activity that is sustainable in the long term; to achieve this means encouraging patients to make appropriate lifestyle changes. PMID- 10863815 TI - The public health message is getting through for NSAIDs. PMID- 10863816 TI - Music as medicine. PMID- 10863817 TI - Complementary therapies. Where to from here? PMID- 10863819 TI - Getting to the line--the Olympic marathon. PMID- 10863818 TI - Complementary therapies and the general practitioner. A survey of Perth GPs. AB - OBJECTIVE: The purpose of this study was to identify the knowledge, attitudes and referral patterns of general practitioners (GPs) toward 10 specific complementary therapies. METHOD: The study was a descriptive cross-sectional postal survey, conducted between July 1998 and August 1998 inclusive. A random selection of 200 male and 200 female Western Australian GPs residing in Perth and listed in the Australian Medical Association database file of registered GPs. RESULTS: The response rate was 74.8% (n = 282). Over 90% of these GPs reported having been approached by more than 30 patients seeking their advice about complementary therapies in the past nine months. The majority of these patients were women, over the age of 35 years. Ten complementary therapies were listed in the questionnaire: acupuncture, hypnosis, meditation, spinal manipulation, yoga, homeopathy, herbal medicine, naturopathy, massage and aromatherapy. Just under half (132) of the respondents had undertaken studies in at least one of the listed complementary therapies, with over 60% reporting a wish for further training. Overall, 67.8% (191) of all respondents reported they were in favour of GP referrals to complementary therapists. However, 56.1% (158) were against complementary therapies being included in rebates for private health insurance. Overall, 75.0% (211) of GPs surveyed had already formally referred a patient to one or more of the listed therapies, the most frequent of these being acupuncture, massage, meditation, hypnosis and spinal manipulation as a part of their overall medical treatment. CONCLUSION: Perth GPs have a high level of interest in complementary therapies. Government regulation and registration of complementary therapies is seen by GPs as important in order to ensure professional standards of practice. Given the high level of interest, provision of undergraduate and postgraduate education in complementary therapies could be considered. In addition, the development of clinical guidelines would be of benefit. PMID- 10863820 TI - Putting together a medical team for the Olympic and Paralympic games. PMID- 10863821 TI - The Australian hockey team's bid for gold--dealing with the medical challenges. AB - How does one find himself in the privileged position of working with the Kookaburras? In my case, in the days prior to a team camp, the incumbent doctor unfortunately required major surgery and did not recover adequately to continue team travel duties. Phone canvassing sifted through sport physicians available at short notice. Medical screening at the camp provided the opportunity to familiarize a doctor coming 'off the bench' with playing and management personnel and vice versa. PMID- 10863822 TI - [Effect of choice of baseline correction interval on localization of electrical heart activity]. AB - The electric heart activity can be localised from body surface mapping data with computer algorithms. At higher heart rates the T and P waves merge. Thus, the offset can not be subtracted in the TP segment. We investigated 28 healthy volunteers with signal averaged 31-lead magnetocardiography. The offset of the baseline was determined in the TP-segment and in the PR-segment, respectively. The electrical heart activity was localised in the initial 30 ms of the QRS complex (Q), at the QRS maximum (R), and at the T wave maximum (T). The volume currents were considered by using a boundary element model with the compartments lungs and torso. The 3D positions of the dipoles, the dipole orientations, and the dipole strengths were calculated using the data preprocessed with two different offset correction intervals. The offsets of the TP and PR segments significantly differed one from another. The average deviations of the dipole localisation were within a few centimetres (Q: 20 +/- 31 mm, R: 6 +/- 13 mm, T: 14 +/- 30 mm). However, in a small number of subjects (Q: n = 5, R: n = 2, T: n = 5) we observed a deviation of more than 30 mm. These deviations were not linearly correlated to the differences in the baseline offsets. High resolution recordings continuously detect heart activity in the PR segment. The correction of the baseline in the PR segment instead of the TP segment may introduce artefacts in the source localisation and therefore should be avoided. PMID- 10863823 TI - [A modular software concept for individual numerical simulation (FEM) of the human mandible]. AB - A new modular software concept for individual numerical simulation of the human mandible using the finite element method (FEM) is presented. The main task is an individual analysis of regional stress and stress-compatibility on the basis of computed tomographic data in individual patients. Simulation should, however, also be possible in parallel with biomechanical experiments, or for further research projects. For this purpose, rapid and uncomplicated generation of the FEM model, easy modification of input data, and short computation times are required. Practical use in the clinical setting makes appreciable additional demands on the individual software components. PMID- 10863824 TI - [Hospital data management at the Graz University Ophthalmology Clinic]. AB - In 1991, the eye hospital at the University of Graz initiated the development of a system for the documentation of patient data and services provided. The starting point of the hospital data management system was the surgical documentation system. This made it possible for the patient to be transferred to the ward accompanied by the completed and signed surgical report. In the subsequent steps we developed a complete client server system adapted to the different specific needs of all the various sections of the eye hospital. A further important feature is the fact that all the data are available for access at every part of the hospital. The current version has been in use since 1996, and will be discussed below. The main features of our hospital data management system are automated coding of medical services provided in the diagnostic, surgical and outpatient areas, and guaranteed authenticated data. Automatic generation of findings, reports, etc, allows the physician to concentrate fully on medical concerns. For a modern, service-oriented hospital, complete records of the services provided are indispensable. Complete recording of services is, however, possible only via automation. In our case, this means not only that the available data are always correct (up to date), but also that there is wide acceptance of and reliance on these data by the medical staff. Since the system is an in-house development, it is possible to react rapidly to suggestions for improvement and to eliminate possible errors immediately. The hospital data management system of the eye hospital at the University of Graz is a well functioning example that makes it worthwhile discussing the greater use of subsystems. PMID- 10863825 TI - [A new optoelectronic measuring device for assessment of leg circumference in comparison with manual measurement]. AB - For compression treatment to be effective in patients with chronic venous insufficiency, it is vital that leg circumference be measured accurately. If compression stockings are custom fit and appropriate for the medical indications, patient compliance will be high. Exact measurements of circumference and length are prerequisites for a good fit. The aim of the present study was to compare an opto-electronic device for the contact-free measurement of calf circumference with the conventional manual method using a tape measure. We investigated the differences between the results obtained with the two methods, and also their reproducibility. Circumferences were measured at defined heights on an anatomically shaped non-yielding leg model and on the leg of a healthy volunteer by 10 different experimenters both with the tape measure and with the opto electronic device. The calf circumferences measured manually with the tape measure varied significantly more than those measured opto-electronically, both in the leg model and in the leg of the volunteer. A systematic error in the opto electronic method appears unlikely, since the manual measurements on the leg model were both larger and smaller than those obtained with the opto-electronic device. Reproducibility was exceptionally high with the opto-electronic device (standard deviation 0.11-0.42 cm). The opto-electronic method yields rapid accurate measurements of circumference with excellent intra- and inter-operator reproducibility. PMID- 10863826 TI - [Cut type femur neck prosthesis--a functional morphologic analysis]. AB - In contrast to other commonly employed models, the cut model femoral neck prosthesis is a cementless prosthesis of comparatively small dimensions. With the aim of investigating its functional integration in the femur, radiological, densitometric and photoelastic analyses were carried out. The biomechanical reaction of the bone tissue expected on the basis of the experimental result is tested in the clinical situation. PMID- 10863827 TI - Boerhaave and the flight from reason in medicine. PMID- 10863828 TI - Keeping the "household machine" running: attendant nursing and social reform in the progressive era. PMID- 10863829 TI - "Straight back to barbarism": antityphoid inoculation and the Great War, 1914. PMID- 10863830 TI - The obscure object of knowledge: German military medicine confronts gas gangrene during World War I. PMID- 10863831 TI - Explanation and uncertainty in the medical world of Ghaambo. PMID- 10863832 TI - Increased transmission of syphilis in men who have sex with men reported from Brighton and Hove. PMID- 10863833 TI - Wound botulism in an injecting drug user in London. PMID- 10863834 TI - [Sundays without traffic and environmental control]. PMID- 10863835 TI - [Salty milk]. PMID- 10863836 TI - [With Vineis]. PMID- 10863837 TI - [Papillomas to screen]. PMID- 10863838 TI - [Carcinogens: destabilizing values]. PMID- 10863840 TI - [European network for the infectious disease control]. PMID- 10863839 TI - [Asbestos and benefits of prevention]. PMID- 10863841 TI - [Toward the efficacy of clinical appropriateness: national program for guidelines]. PMID- 10863842 TI - [Electromagnetic fields: an infinite containment]. PMID- 10863843 TI - [In search of divided decisions]. PMID- 10863844 TI - [From the oncologist's standpoint]. PMID- 10863845 TI - [Health and lost honor of the dictator]. PMID- 10863846 TI - [Levels of polycyclic aromatic hydrocarbons (PAHs) in air samples in the city of Arezzo (1997-1998)]. AB - Polycyclic aromatic hydrocarbons (PAHs) are widely distributed environment pollutants of air, water and soil. Since many PAHs are potent mutagens and/or carcinogens the occurrence of these compounds in the lower atmosphere is an important element of environmental pollution. We measured PAH levels in airborne particles collected in the town of Arezzo, (Tuscany, Italy), during the period April 1997-February 1998. Air monitoring for 24 h was repeated for 7 days, during two weeks, in each season; a total of 84 air samples were obtained sampling two urban sites where the traffic is the main source of pollution. One site is a residential area. The data of this study indicate a pronounced seasonal variation in PAH levels and show that in cold spells other sources of contamination besides vehicular traffic are important. PMID- 10863847 TI - [Monitoring coronary and cerebrovascular events in Naples]. AB - The Authors have monitored coronary and cerebrovascular events occurring in an area of Naples in 1995, using a register according to the simplified methodology derived from the MONICA project. Demographic and mortality data, hospital discharge diagnoses were collected. In people aged 25 to 64, acute coronary events were 365. The fatality rate was 34.8%, with 14.8% of deaths in hospital. 76% of death in hospital occurred within 24 hours. Acute cerebrovascular events were 173. The fatality rate was 23.1%, with 8.1% of deaths in hospital. 63.7% of those who died in hospital had survived more than one week. The attack rates for coronary events were 268.8 for males and 56.1 for females and respectively 96.3 and 56 for cerebrovascular events. Regarding coronary events, these values are greater than attack rates of MONICA project's Area Latina; for cerebrovascular events they are smaller. This study shows that the Naples register is useful for knowing the incidence of cardiovascular diseases and for monitoring them. PMID- 10863848 TI - [Mortality risk in intravenous drug users in Emilia Romagna region and its socio demographic determinants. Results of a longitudinal study]. AB - Results on general and cause-specific mortality in a retrospective cohort of intravenous drug users in the Emilia Romagna Region (Italy) are presented. Four thousand two hundred and sixty subjects (3324 males, 936 females) in public treatment centres in Piacenza, Modena and Ferrara provinces have been observed for up to 20 years in the period 1975-95. AIDS age-adjusted death rates dramatically increased all during the period, while overdose and other causes (mostly accidental) increase up to the early nineties and then tend to decrease. This last pattern has not been described in other Italian cohorts, and could be related with changes in therapeutic strategies. General mortality is very high in this cohort, as in other studies (males: SMR 16.7, LC 15.3-18.2; females: SMR 33.4, LC 27.9-39.9). Survival probability after 15 years of observation is 65% in males and females combined. Apart from overdose and AIDS, other relevant excesses are observed for accidental deaths (especially car accidents), cirrhosis, infective causes and cancer in males; in females, accidental deaths (among which homicides) and digestive tract diseases. A higher death risk is observed for males who began drug use before age 20, who contacted treatment centres in the nineties and at an older age, and who came in contact with the law. PMID- 10863849 TI - [Health databases to assess the incidence of lymphoid malignancies in Italian population]. AB - We tested the usefulness of the National Health Service Databases for investigating the incidence of lymphoid malignancies in an Italian community. We analyzed hospital discharge data, drug prescription, pathologic records and death certificates to identify the new cases of Hodgkin's disease, non Hodgkin's lymphoma, multiple myeloma, and acute and chronic lymphocytic leukemia diagnosed in the municipal population of Reggio Emilia, northern Italy, 1991 through 1996. The completeness of Hospital discharge data was very high, and several incident cases could be identified only through this source. Completeness of the pathologic registry was satisfactory for Hodgkin's disease and non-Hodgkin's lymphoma, and this source independently yielded a few incident cases of lymphoid neoplasms. Analysis of death certificates and drug prescriptions appears to be of limited value in the epidemiology of lymphoproliferative diseases. PMID- 10863851 TI - [Toward the equity and health on the threshold of 21 century and in the era of globalization. International Association of Health Policy]. PMID- 10863850 TI - [Setting priorities for environmental protection from asbestos: ethical aspects]. AB - The allocation of resources destined for environmental rehabilitation of areas contaminated by asbestos requires scientific, political, economical and ethical evaluation. Ethically relevant issues reviewed in this paper include: identification of the populations most expected to benefit from the intervention, risk assessment, effectiveness of different technical approaches, as well as a comparison between populations of current socio-economic standards, level of legal assistance and access to both information and participation in the decision making process. A relevant ethical criterion is minimizing inequalities within and between communities. PMID- 10863852 TI - [A cluster of doubts]. PMID- 10863853 TI - [Difficult relations among citizens and national health services]. PMID- 10863854 TI - [Prognostic factors for survival following transarterial chemoembolization in advanced hepatocellular carcinoma]. AB - The efficacy of transarterial chemoembolization in the palliative treatment of non-resectable hepatocellular carcinoma is controversial. To determine the possible existence of clinical and analytical variables with independent predictive value for survival related to the tumor and the treatment given, a multivariate analysis in a series of 111 patients who underwent transarterial chemoembolization was carried out. Overall actuarial survival was 54%, 31% and 24% at 1, 2 and 3 years respectively. Child-Pugh score (p < 0.05), tumor size (p < 0.05) and arterial occlusion after intraarterial chemotherapy (p < 0.05) reached independent predictive value. The group of patients in whom two or three of these factors were simultaneously present had a very poor prognosis with a survival of 20% and 0% at 1 and 2 years respectively, compared with 60%, 50% and 37% at 1, 2 and 3 years respectively in the group with one or none of these factors (p < 0.01). Grouping on the basis of these variables may be useful in the design of future controlled prospective studies that aim to determine the efficacy of transarterial chemoembolization. PMID- 10863855 TI - ["Rapid" serology for the diagnosis of Helicobacter pylori infection. Evaluation of its accuracy compared with a gold-standard and its concordance with "classic" serology]. AB - AIM: To prospectively evaluate the validity of a rapid office-based diagnostic serological test (using capillary blood) in our population, taking as reference a combination of standard diagnostic methods, as well as to compare the results of this technique with those of "classic" serological tests (using venous blood). PATIENTS AND METHODS: We prospectively studied 39 consecutive patients with symptoms of the upper digestive tract who had undergone oral gastroscopy. Gastric biopsies were taken for histology and rapid urease testing, and a 13C-urea breath test was performed. An enzyme-linked immunoassay that detects IgG antibodies against Helicobacter pylori was used as a "classic" serological test and the commercial kit FlexPack HP was used as a "rapid" serological test. The endoscopist, the pathologist and those responsible for reading the rapid urease test, the 13C-urea breath test and both serological tests did not know the results of the other diagnostic methods. Patients were considered H. pylori positive when at least two of the three validated tests (rapid urease test, histology, and 13C-urea breath test) revealed infection and were considered free of infection when all tests were negative. RESULTS: Thirty-nine patients were studied. Thirty-eight per cent were male (mean age 48 +/- 15 years). The prevalence of H. pylori infection detected by the gold standard was 69.2%. The sensitivity and specificity of the "classic" serological test was 96% 95% (CI: 79 99) and 91% (59-100). "Rapid" serological testing was positive in nine patients, negative in 28 and indeterminate in two. A single digital puncture was sufficient in 80% of the patients, 15% needed two and 5% needed three. Most patients (77%) had no preference for either type of serological test while 20.5% preferred digital puncture and 2.5% venous puncture. The sensitivity, specificity, positive predictive value and negative predictive value were 31% (16-50), 91% (59-100), 89% (52-100) and 36% (19-56) respectively. Sensitivity was unaffected by age but specificity was lower in patients older than 40 years (89% vs. 100%; McNemar's test: 8; p < 0.01). Kappa's coefficient between the "classic" and the "rapid" serological tests was 0.16 (SE 0.1) and McNemar's test was 12.2 (p < 0.001), which indicates that the prevalence of infection diagnosed by both methods was not homogeneous. CONCLUSION: The "rapid" office-based serological test used in our study is of insufficient diagnostic accuracy to be used in clinical practice to identify H. pylori infection. PMID- 10863857 TI - [Recurrent pyogenic cholangitis in a western patient]. AB - Recurrent pyogenic cholangitis, or oriental cholangiohepatitis, is highly prevalent in Southeast Asia where it is a common cause of attendance at emergency centers. Sporadic cases have been described outside these areas, especially in asian immigrants but its appearance in westerners is exceptional. We present a case of this disease in a western patient and discuss aspects of its diagnosis and therapy. PMID- 10863856 TI - [Role of ribavirin in the treatment of chronic hepatitis B]. AB - AIM: To evaluate the safety and efficacy of 1,200 mg/day of ribavirin for 6 months in the treatment of chronic hepatitis B. MATERIALS AND METHODS: An open study was carried out with 25 patients with chronic hepatitis B who had previously received placebo (first phase) as part of a randomized, double blind study and who remained HBeAg and HBV DNA positive. In the second phase they received oral ribavirin (1,200 mg/day) for 24 weeks and the results of the first phase were compared with those of the second. All the patients had a recent histological diagnosis and were anti-HCV and anti-HIV negative. In both phases clinical and laboratory evaluations were carried out at weeks, 0, 4, 8, 12, 16, 24, 32, 40 and 48 which included blood tests, liver function tests and serological markers of HBV, and HBV DNA when HBeAg became negative. Liver biopsy was performed at the beginning of the first phase, 6 months later and at the end of the second phase. RESULTS: Mean values of alanine aminotransferase (ALT) showed a clear downward trend and were reduced by 50% at the end of the study while during the first phase these values were similar to basal values (range 32.3-45.5 IU). In the second phase, seroconversion of HBeAg was 56.0% (p = 0.00001) and HBV DNA was negative in 36%. The number of patients who showed improvement in Knodell's index was 86.7% in the second phase vs. 13.3% in the first phase (p = 0.00001). The drug was well tolerated and the only significant adverse reactions were a reduction in hemoglobin levels greater than 10% of the basal value in 84% of the patients, gastric acidity in 40% and fatigue in 32%. CONCLUSIONS: Ribavirin therapy at a dose of 1,200 mg/day for 24 weeks was well tolerated and efficacious in returning serum ALT levels to normal, in the seroconversion of HBeAg and negativization of HBV DNA as well as in reducing liver necrosis and inflammation. This study confirms that ribavirin may be considered a therapeutic option in the treatment of chronic hepatitis B. PMID- 10863858 TI - [Three cases of villous adenoma of the Vater papilla]. AB - A series of three cases of villous adenoma of the papilla of Vater is reported. In two cases, the pathological diagnosis made before, during and after surgery was of benign villous adenoma. In the third case, the diagnosis made before and during surgery was of adenoma but in the postsurgical examination a macro invading carcinoma was found. Further endoscopic exploration revealed a recurrent villous adenoma and further surgery was carried out. Total resection, rather than a duodenopancreatectomy, was performed due to the length of time from detection of the recurrence until the second operation as well as to the small size of the recurrent tumor in the surgical examination. PMID- 10863859 TI - [Percutaneous intrahepatic portosystemic shunting in the treatment of veno occlusive disease of the liver after bone marrow transplantation]. AB - Veno-occlusive disease of the liver is a frequent cause of morbidity and mortality after bone-marrow transplantation. Its clinical manifestations are primarily related to the development of portal hypertension and sinusoidal congestion. The efficacy of the different therapeutic options used is controversial. We present a 22-year-old woman with veno-occlusive liver disease histologically confirmed after autologous bone-marrow transplantation, with progressive alteration in liver biochemistry and ascites. She was treated by percutaneous intrahepatic portosystemic shunting. After the procedure there was a marked improvement in ascites and an increase in diuresis with liver function progressively returning to normal. The safety and efficacy of this approach in the treatment of patients with veno-occlusive liver disease should be evaluated in controlled studies. PMID- 10863860 TI - [PET detection of recurring rectal adenocarcinoma in an unusual location. Case report]. AB - A rare pattern of colon cancer recurrence is presented. A 63-year-old man underwent surgical resection after diagnosis of colon cancer. The postsurgical pathologic examination showed a stage II colon cancer (MAC B2). Six courses of adjuvant chemotherapy (Mayo protocol) were started within four weeks of surgery. During follow-up, serial serum carcinoembryonic antigen levels became progressively elevated in an otherwise asymptomatic patient who showed no signs of recurrence in any of the conventional imaging tests performed (chest X-ray, abdominal ultrasound, and abdominal CT-scan). Positive findings suggesting lymph node mediastinal metastases were present in the PET scan. Surgical resection and pathologic examination demonstrated metastases of colon adenocarcinoma. PMID- 10863861 TI - [Factors involved in hepatic fibrogenesis]. PMID- 10863862 TI - [Hepatotoxicity induced by non-steroidal anti-inflammatory drugs]. PMID- 10863863 TI - [Dyspepsia, Helicobacter pylori and endoscopy]. PMID- 10863864 TI - [Anaphylaxis and hydatid cyst of the liver]. PMID- 10863865 TI - [Intestinal perforation due to a foreign body treated with laparoscopy]. PMID- 10863866 TI - [Melanoma of the intestine in a patient without a known previous primary tumor]. PMID- 10863867 TI - Men, gender, and health: toward an interdisciplinary approach. PMID- 10863868 TI - Understanding men's health and illness: a gender-relations approach to policy, research, and practice. AB - Men's health has emerged as an important public concern that may require new kinds of healthcare interventions and increased resources. Considerable uncertainty and confusion surround prevailing understandings of men's health, particularly those generated by media debate and public policy, and health research has often operated on oversimplified assumptions about men and masculinity. A more useful way of understanding men's health is to adopt a gender relations approach. This means examining health concerns in the context of men's and women's interactions with each other, and their positions in the larger, multidimensional structure of gender relations. Such an approach raises the issue of differences among men, which is a key issue in recent research on masculinity and an important health issue. The gender-relations approach offers new ways of addressing practical issues of healthcare for men in college environments. PMID- 10863869 TI - Identifying male college students' perceived health needs, barriers to seeking help, and recommendations to help men adopt healthier lifestyles. AB - Seven focus groups at a university campus were formed to identify college men's health concerns, barriers to seeking help, and recommendations to help college men adopt healthier lifestyles. Content analysis was used to identify and organize primary patterns in the focus-group data. Results of the study revealed that the college men were aware that they had important health needs but took little action to address them. The participants identified both physical and emotional health concerns. Alcohol and substance abuse were rated as the most important issues for men. The greatest barrier to seeking services was the men's socialization to be independent and conceal vulnerability. The most frequently mentioned suggestions for helping men adopt healthier lifestyles were offering health classes, providing health information call-in service, and developing a men's center. Implications of the results are discussed. PMID- 10863870 TI - Toward a transformed approach to prevention: breaking the link between masculinity and violence. AB - Men are disproportionately overrepresented among both perpetrators and victims of violent crime. Scholars from the men's studies movement have documented a clear link between socialization into stereotypical norms of hegemonic masculinity and an increased risk for experiencing violence. Despite this evidence, most campus prevention programs fail to recognize the link between men and violence and use only traditional approaches to violence prevention. The most that on-campus prevention programs provide are self-defense seminars for potential female victims of rape and general campus safety measures. In this article, the author describes a comprehensive, transformed approach to violence prevention. Data from a year-long case study of Men Against Violence, a peer education organization at a large university in the South, demonstrate the feasibility of meaningfully expanding male students' conceptions of manhood and appropriate gender roles and, thus, reducing the likelihood of men's engaging in sexually or physically violent behavior. PMID- 10863871 TI - College men's health in practice: a multidisciplinary approach. AB - After a needs assessment indicated that male students underutilized campus health services, the San Francisco State University Student Health Service developed a coordinated complement of outpatient health services for men. The authors review their experience in developing, implementing, operating, and evaluating this ongoing clinical service. The needs assessment and subsequent program evaluation data suggest that male students on a large, culturally diverse, urban campus would respond favorably to targeted, multidisciplinary health initiatives that incorporate the principles of health promotion and disease prevention. PMID- 10863872 TI - Why do men get more heart disease than women? An international perspective. AB - Biological, behavioral, and psychosocial contributions to the gender gap in coronary heart disease (CHD) are discussed. Although CHD is the Number 1 cause of death for both sexes in the industrialized world, CHD mortality rates between these countries are larger than those between men and women, suggesting that biological factors are not the sole influences on the gender gap in CHD. Traditional coronary risk factors cannot explain the rapid increase in CHD mortality among middle-aged men in many of the newly independent states of eastern Europe. However, eastern European men score higher on stress-related psychosocial coronary risk factors (e.g., social isolation, vital exhaustion) than men living in the West. Comparisons between the sexes also reveal gender differences in psychosocial and behavioral coronary risk factors, including excessive alcohol consumption and smoking, favoring women. Overall, it appears that men's coping with stressful events may be less adaptive physiologically, behaviorally, and emotionally, contributing to their increased risk for CHD. PMID- 10863873 TI - An exploration of the drive for muscularity in adolescent boys and girls. AB - Much of the existing research on disordered eating has centered on the drive for thinness, which is most commonly observed in girls and women. The male standard of bodily attractiveness, however, is bigger, bulkier, and more muscular. Are boys and men motivated to be big and muscular in the same way that girls and women are motivated to be thin? The authors constructed a 15-item survey and administered it to 197 adolescents. The findings showed that the drive for muscularity measure displayed good reliability; that individuals high in the drive were more likely to be boys who were trying to gain both weight and muscle mass; that the drive was related to poor self-esteem and higher levels of depression among boys, but not among girls; and that the drive for muscularity was relatively unrelated to the drive for thinness. PMID- 10863874 TI - Why college men drink: alcohol, adventure, and the paradox of masculinity. PMID- 10863875 TI - Perioperative progress check of interdental forces following extraction of the third molars. A prospective long-term study. AB - With the aim of checking the progress of mesially directed forces in a complete dental arch, we measured the interdental frictional forces in a population of 44 patients with erupting maxillary and mandibular third molars. The measurements were taken preoperatively and postoperatively 1, 2, 3, 7, and 12 weeks and also 1 year after extraction or surgical removal of the third molars in all 4 quadrants. In addition, the interdental frictional forces of a total of 40 test persons in 3 control groups were similarly measured in a state of inertia, after a period of 1 hour spent in supine position, and after a 5-minute period of chewing activity. Over the first 4 to 12 weeks the interdental forces showed a significant postoperative average decrease of 16.1% in the maxilla and 18.0% in the mandible. After 1 year the average reduction in the level of interdental forces in the patient population was 10.3% in the maxilla and 10.9% in the mandible. A significant postoperative reduction in force of 40.7% was registered for third molars with a mesial angle of 66 degrees to 90 degrees to the tooth axis of the second molar in the mandible. Following a 1-year control period, the level of interdental force in the patient population was found to be on average 27.4% below the preoperative baseline value. In control group I, the margin of fluctuation of interdental forces in a state of inertia was +/- 3.5% over a 3 month period. In control group II, a significant average decrease in interdental forces was measured in the maxilla (15.1%) and mandible (13.2%) following a 1 hour period in supine position. In control group III, a non-significant average reduction in interdental forces was established in the maxilla (7.8%) and mandible (8.6%) following a 5-minute period of chewing activity. PMID- 10863876 TI - Early orthodontic treatment of Class-III malocclusion in Germany. AB - In order to evaluate current attitudes to early interceptive treatment, 2001 orthodontic offices in Germany were asked to fill in a questionnaire comprising the following topics: indication, appliances for the early correction of Class III malocclusions, diagnostic records, duration, and benefits to overall therapy. Based on the 677 evaluable questionnaires, the following statistically significant conclusions could be drawn: 92.6% of the orthodontists see Class-III malocclusion as an indication for early treatment. Early treatment of severe crowding, diastemata, Class-II malocclusion, deep bite, increased overjet and impacted incisors was declined by most orthodontists. The interceptive treatment of further malocclusions was controversially discussed. Functional appliances (67.5%), in particular the Frankel III (47.3%), were dominant in correction of Class-III malocclusions. Typical orthodontic records relating to early interceptive treatment include panoramic radiographs, lateral headfilms, photos and dental casts. 2.5% of the orthodontists routinely take a hand-wrist radiograph. Although recently published studies support the use of facial masks in theory, they are rarely used in practice. To what extent early interceptive treatment of Class-III malocclusion influences the overall treatment is the subject of further studies. PMID- 10863877 TI - Modified pendulum appliance including distal screw and uprighting activation for non-compliance therapy of Class-II malocclusion in children and adolescents. AB - In present-day orthodontics there is an increasing call for therapeutic procedures and appliances allowing those delivering the treatment maximum independence from patient cooperation. This requires both the development of new therapeutic appliances and the optimization of existing ones (via modification). The present study aims to analyze the therapeutic outcomes obtained by using a pendulum appliance (distal screw and integrated molar uprighting activation), modified and independent of patient cooperation. The dentoalveolar and skeletal outcomes and secondary outcomes were observed in 50 children and adolescents. The dentoalveolar outcomes were as follows: 72.5% of the clinically visible increase in space were gained by distal molar movement and 27.5% by mesialization of the anterior segment. The molars showed slight distal tipping and extrusion, while the incisors showed slight labial tipping. In terms of time, the molars were moved distally by an average of 0.67 mm per month. Skeletal changes taking place during treatment using the pendulum appliance are negligible. Further advantages of the pendulum appliance are independence from patient cooperation and good acceptance by patients. PMID- 10863878 TI - Bond strength of various fluoride-releasing orthodontic bonding systems. Experimental study. AB - The aim of the study was to compare the shear bond strength of a fluoride releasing composite resin adhesive (Light Bond, Reliance) and a light-cured resin reinforced glass ionomer cement (Fuji Ortho LC, GC America) bonded to extracted teeth under different enamel surface conditions. Forty human premolars were divided at random into 4 groups of 10 specimens. Stainless steel brackets were attached to the enamel surface by 1 of the 4 protocols: 1. Fuji Ortho LC, moist non-etched enamel; 2. Fuji Ortho LC, moist etched (37% H3PO4); 3. Light Bond, dry etched (37% H3PO4); 4. Light Bond, dry etched (Etch & Prime 3.0, Degussa). The teeth were stored in deionized water at 37 degrees C for 48 hours. Shear bond strengths was determined at a crosshead speed of 1 mm/min. The residual adhesive on the enamel surface was evaluated with the modified Adhesive Remnant Index (ARI). Analysis of variance (ANOVA) and Duncan's test were used to compare the 4 groups. Significance was predetermined at p = 0.05. Significant inter-group differences were found (p < 0.0001). The mean SBS (and SD), in MPa were: Group 1: 15.9 (4.7); Group 2: 20.3 (2.5); Group 3: 16.7 (2.6); Group 4: 11.7 (2.5). Glass ionomer cement without etching and composite with Etch & Prime showed adhesive failures at the enamel and good enamel integrity after debonding. The other specimens showed mixed or adhesive fractures at the bracket failure sites. Glass ionomer used on wet tooth surfaces without etching shows a clinically acceptable bond strength with clean separation from the enamel after debonding. PMID- 10863879 TI - Postretention changes in canine position. Results of a long-term follow-up. AB - The aim of the present study was to evaluate sagittal, vertical and transverse changes in canine position and a possible correlation with relapse of lower anterior crowding. From a sample with a long-term follow-up of 15.7 +/- 4.4 years after treatment, patients with a full dentition were enrolled in the study. Post treatment and long-term follow-up casts of 117 patients were measured and statistically analyzed. A decrease in intercanine width was found between post treatment and long-term follow-up records, with more lingual inclination of the mandibular canine axis in the transverse plane. There was a significant correlation between maxillary and mandibular canine inclination. Concomitant changes were an increase in lower anterior crowding and a decrease in mandibular intercanine width. Relapse of anterior crowding has a multifactorial etiology. The results of this study suggest a possible influence of the canines. Functional causes in particular might have a profound influence and should be considered during treatment planning. PMID- 10863880 TI - Serial extraction or premolar extraction in the permanent dentition? Comparison of duration and outcome of orthodontic treatment. AB - Treatment outcome and duration of 2 different treatment approaches in 2 groups of comparable extraction cases were analyzed: Group I: serial extraction performed in the early mixed dentition followed by orthodontic treatment in the permanent dentition; group II: extractions as well as orthodontic treatment in the permanent dentition. The following conclusions were reached: 1. The treatment period with fixed appliances was highly significantly shorter in group I; however, the number of appointments was significantly higher and the total duration of treatment/observation time significantly longer. 2. In both groups the reduction in PAR score was either improved or greatly improved in all cases. PMID- 10863881 TI - TMJ morphology and function in a patient with Klippel-Trenaunay syndrome. Case report. AB - Klippel-Trenaunay syndrome is a rare congenital mesodermal disturbance of uncertain etiology in variable expression. The classic manifestation is the triad of congenital mesodermal abnormalities. Clinically a diversity of phenotypes with subjacent malformations may be encountered. The deviations of the mesioblastic germ layer affecting angioblastic, lymphoblastic and osteoblastic structures, may give rise to malformations either alone or in an unlimited diversity of associations. While this syndrome may be diagnosed by chance in the course of ultrasonic scanning during pregnancy, it is normally diagnosed during infancy or early childhood. Evaluation and carefully coordinated medical treatment are important in minimizing morbidity and relieving multiple complaints. The aim of our investigation was to evaluate and correlate the clinical, functional, radiographic and MRI findings in a patient suffering from this syndrome. We present a 13-year-old male patient suffering from extreme facial asymmetry in association with hypertrophy of the complete right side of the body. In spite of pronounced functional and morphologic asymmetry, no signs of degenerative joint disease were identified by radiography or MRI. Since temporomandibular joint dysfunction and facial asymmetry can result in irreversible degenerative joint disease, close follow-up monitoring is indispensable if joint damage is to be prevented. PMID- 10863882 TI - Serotonergic drugs and panic disorder. PMID- 10863883 TI - Neuroimaging, auditory hallucinations, and the bicameral mind. PMID- 10863884 TI - Citalopram--a review of pharmacological and clinical effects. AB - OBJECTIVE: To provide clinicians with a critical evaluation of citalopram, a selective serotonin reuptake inhibitor (SSRI) that has been available in Canada since March 1999. DATA SOURCES: Commercial searches (MEDLINE and BiblioTech) and an "in-house" search (InfoDrug) were used to find published English-language references for clinical and preclinical publications. There was no restriction of publication dates. Primary index terms used were: pharmacological properties, receptors, pharmacological selectivity, pharmacokinetics, age-related pharmacokinetics, sex-related pharmacokinetics, renal dysfunction, hepatic dysfunction, cytochrome activity, drug interactions, adverse reactions, antidepressant switching, precautions, overdose, drug discontinuation, children, geriatric, depression, combination therapy, placebo control, refractory depression, anxiety disorders and medical disorders. STUDY SELECTION: A total of 74 studies were reviewed. Twenty-one of these studies specifically examined the clinical efficacy and tolerability of citalopram in depressive disorders as well as other disorders. In depressive disorders, clinical studies were required to have either placebo or active comparison controls for a minimum of 3 weeks. For other disorders, in the absence of double-blind trials, open-label studies were included. Pharmacological studies were limited to animal studies focusing on citalopram's selectivity and receptor specificity, and positron emission tomography studies were incorporated to include human pharmacological data. Pharmacokinetic studies focused on the metabolism, safety and tolerability of citalopram, specifically with reference to adverse reactions, drug interactions and overdose in addition to citalopram's effect on vulnerable populations, such as children, the elderly and patients with metabolic diseases. DATA EXTRACTION: Data on clinical studies were summarized according to test measures, study duration and outcome of study. Pharmacokinetic and pharmacodynamic studies were summarized according to properties and interactions. Adverse reactions were extracted to outline citalopram's safety profile. DATA SYNTHESIS: Citalopram is an SSRI antidepressant with a more specific and selective pharmacological profile than other antidepressants of its class. It is well tolerated, and drug interactions are not a significant concern. It is also reasonably safe for populations vulnerable to pharmacokinetic effects, such as the elderly and patients with metabolic diseases. In addition to its tolerability, citalopram is effective in the treatment of major depression, other depressive disorders and panic disorder. It has the potential to effectively treat other anxiety disorders and substance-use disorders; in addition, it may be useful in several medical conditions. CONCLUSIONS: There is evidence to support the role of citalopram as a well-tolerated and effective SSRI antidepressant. There is a need for further evaluation of its role in psychiatric disorders other than major depressive disorder. PMID- 10863885 TI - Selective serotonin reuptake inhibitor discontinuation syndrome: proposed diagnostic criteria. AB - OBJECTIVE: To establish specific criteria by which selective serotonin reuptake inhibitor (SSRI) discontinuation syndrome may be identified. DATA SOURCES: MEDLINE and PSYCHLIT databases were searched for case reports published from 1986 to 1997 inclusive, and references of relevant articles were also searched. STUDY SELECTION: Forty-six case reports of symptoms following the discontinuation of fluoxetine, fluvoxamine, paroxetine or sertraline were selected. Three studies of SSRI discontinuation were also reviewed. DATA EXTRACTION: Demographic and treatment information, as well as the timing, duration, number, nature and frequency of dicontinuation symptoms. DATA SYNTHESIS: Paroxetine was most frequently implicated. The drug had been tapered in half of the cases. In some cases, symptom onset began during taper, whereas, in most cases, symptoms began within 1 to 3 days of drug discontinuation. Fifty-three different symptoms were reported, with dizziness being the most common. Other common symptoms were nausea or emesis, fatigue, headache, gait instability and insomnia. Shock-like sensations, paresthesia and visual disturbances were the most rare. Without intervention, symptoms persisted for more than a week in half of the cases. In cases in which the SSRI was restarted, symptoms resolved within 72 hours. In some cases, withdrawal symptoms recurred when the same SSRI was again discontinued. CONCLUSIONS: Findings were used to construct diagnostic criteria for the SSRI discontinuation syndrome. These criteria are 2 or more of the following symptoms developing within 1 to 7 days of discontinuation or reduction in dosage of an SSRI after at least 1 month's use, when these symptoms cause clinically significant distress or impairment and are not due to a general medical condition or recurrence of a mental disorder: dizziness, light-headedness, vertigo or feeling faint; shock-like sensations or paresthesia; anxiety; diarrhea; fatigue; gait instability; headache; insomnia; irritability; nausea or emesis; tremor; and visual disturbances. PMID- 10863886 TI - The role of depression severity in the cognitive functioning of elderly subjects with central nervous system disease. AB - OBJECTIVE: To examine the hypothesis that there is a causal relation between depression and cognitive dysfunction in patients with central nervous system (CNS) disease. DESIGN: Retrospective analysis of a clinical database. SETTING: Tertiary geriatric day hospital. PATIENTS: Sixty-five patients with depression and CNS disease, and 201 patients with depression but without CNS disease. OUTCOME MEASURES: Scores on the Hamilton Depression Rating Scale (Ham-D) and the Mattis Dementia Rating Scale (MDRS). RESULTS: A logistic regression analysis using MDRS status as the dependent variable, and a number of clinical variables as the predictor variables, showed that, in patients with CNS disease, only the Ham-D score predicted MDRS status (R = -0.19, p = 0.02). Ham-D score even more strongly predicted scores on a frontal system subtest of the MDRS (R = -0.262, p = 0.005). Ham-D score did not predict MDRS status in patients without CNS disease. Mean Mini Mental State Examination scores for the group with CNS disease were 25.1 at admission and 26.1 at discharge (p < 0.001). CONCLUSIONS: These findings suggest that depression contributes to frontal cognitive dysfunction in patients with CNS disease. PMID- 10863887 TI - Variations in response to citalopram in men and women with alcohol dependence. AB - OBJECTIVE: To examine the differential effects of citalopram on alcohol consumption in nondepressed women and men with mild to moderate alcohol dependence. DESIGN: Prospective, placebo-controlled study. PARTICIPANTS: Sixty one subjects (34 men and 27 women). INTERVENTIONS: After a 2-week baseline, subjects were randomly assigned to 12 weeks of citalopram (40 mg per day) (n = 15 women, 16 men) or placebo (n = 12 women, 18 men). All received brief standard psychosocial interventions. OUTCOME MEASURES: Alcohol Dependence Scale, Montgomery-Asberg Depression Scale, Michigan Alcohol Screening Test, State-Trait Anxiety Inventory and daily alcohol intake. RESULTS: Pretreatment sex differences were evident in alcohol consumption, alcohol dependence, alcohol-related problems and on anxiety and depression measures. After treatment, analyses of covariance with depression and anxiety scores as covariates revealed a differential benefit of citalopram for men. Men receiving citalopram reduced average drinks per day by 44%, whereas women exhibited a 27% decrease (p < 0.05). CONCLUSIONS: Men may benefit more than women from citalopram in the treatment of alcohol dependence. These findings highlight the importance of examining sex as a significant variable in evaluating response to pharmacotherapy. PMID- 10863889 TI - Chlorpromazine-induced cutaneous pigmentation--effect of replacement with clozapine. PMID- 10863888 TI - Diltiazem as augmentation therapy in patients with treatment-resistant bipolar disorder: a retrospective study. AB - OBJECTIVE: To examine the efficacy of a slow-release formulation of diltiazem as adjunctive therapy in patients with treatment-resistant bipolar disorder. DESIGN: Retrospective study. PATIENTS: Eight female patients with treatment-resistant bipolar disorder. INTERVENTIONS: Patients were administered diltiazem and monitored for a 6-month period before starting diltiazem and a 6-month period after starting the drug. OUTCOME MEASURES: All patients were seen at least monthly and usually every 2 weeks. The frequency and severity of both depressive and manic episodes were examined during the 6-month period after starting diltiazem, and compared with those during the 6-month period before diltiazem treatment. RESULTS: There was a statistically significant decrease in the frequency and severity of both manic and depressive episodes in these patients after they started treatment with diltiazem, compared with the period before they started treatment with diltiazem (p < 0.001). There was no evidence of side effects requiring patient withdrawal or of drug interactions. CONCLUSIONS: The results support previous suggestions that calcium-channel antagonists may be an effective adjunctive treatment in the management of bipolar disorder. Further controlled clinical studies are needed to confirm this small, open-label, retrospective study. PMID- 10863890 TI - Reference spirometric values for healthy lifetime nonsmokers in Thailand. AB - The normal spirometric reference values for Thai people are still not yet available. The aim of this study was to establish standard spirometric equations for Thai people. Subjects 10 years of age and over were selected and their demographic distributions represented that of the population of the whole country. Inclusion criteria were strictly lifetime nonsmokers, no history of chronic cardiopulmonary disease (using a modified ATS--DLD 78 respiratory adult questionnaire), normal standard chest radiograph and unremarkable physical examination. They had to be without respiratory symptoms at the time of the study. Spirometric values were obtained by 5 turbine system 'Pony graphic' (Cosmed, Italy) spirometers which met ATS recommendations. A normal group of 2299 women and 1655 men were selected. Regression analyses using sex, height and age as independent variables were used to provide equations for predicted values. The results were: [table: see text] FVC and FEV1 from this study are close to the Chinese but are 8-20 per cent lower than the Caucasians. These predicted equations are recommended to be used for future reference values in the Thai population. PMID- 10863891 TI - Perivalvular abscesses due to Staphylococcus aureus endocarditis comparison with Streptococcus viridans endocarditis and incremental value of transesophageal echocardiography. AB - BACKGROUND: Perivalvular abscesses are major complications of infective endocarditis (IE). The prevalence and best approach to detection of this complication in Staphylococcus aureus (SA) in comparison to Streptococcus viridans (SV) IE is unclear. METHOD: Among 243 consecutive episodes of IE diagnosed using the Duke criteria, who underwent either transthoracic (TTE) or transesophageal echocardiography (TEE) at the Mayo Clinic between 1988 and 1993, there were 64 cases of SV and 61 of SA IE. Comparison of TTE and TEE detection of abscesses were restricted to patients with either surgical or autopsy examination and both TTE and TEE were performed. RESULTS: Prosthetic valve and valve repair were significantly higher in SA compared to SV IE (46 vs 23%, P = 0.008). The prevalence of abscesses was higher in SA compared to SV IE (42 vs 14%, P = 0.08). 1 (10%) of abscess detected by TTE in SA compared to 1 (50%) in SV IE and 6 (60%) by TEE in SA and 1 (50%) in SV IE. Incremental value of TEE vs TTE was higher in SA 5/24 (21%) than in SV IE 0/14 (0%) P = 0.067. Hospital mortality was significantly higher in SA than SV IE (13 vs 2%, P = 0.013). CONCLUSION: Patients diagnosed with IE and those with SA 1) presented more often with prosthetic valve IE, 2) developed more perivalvular abscesses, and 3) had a higher in hospital mortality than those with SV. Incremental value of TEE was higher in SA than in SV IE, 4) therefore, had a stringent requirement for initial and repeated TEE to detect this ominous complication of IE. PMID- 10863892 TI - Excimer laser phototherapeutic keratectomy for corneal diseases. AB - The efficacy and safety of excimer laser phototherapeutic keratectomy (PTK) for treatment of various corneal pathologies were determined. The preoperative indications included lattice dystrophies (10 eyes), Reis-Bucklers dystrophies (4 eyes), macular dystrophies (2 eye), and corneal scarring secondary to trauma (1 eye). Mean follow-up time was 9.9 months (range 6-18 months). Uncorrected visual acuity postoperatively improved in 15 eyes (88.2%); not improved in 1 eye (5.9%) and decreased in 1 eye (5.9%). Corneal clarity improved in 14 of 17 eyes (82.4%) which corresponded to the improvement of uncorrected visual acuity. Ocular discomfort improved in 16 eyes (94.1%), decreased in 1 eye (5.9%) which subsequently developed double vision. The complications included delayed reepithelialization (> 7 days) in 6 eyes (35.3%) and corneal scarring 1 eye (5.9%). Sixty four per cent had increased significant hyperopia (> 4 D) and 7.1 per cent had significant induced astigmatism (> 2 D). One eye (5.9%) needed retreatment due to remaining corneal opacity. One eye (5.9%) had double vision due to irregular astigmatism. Excimer laser PTK is effective and safe for treatment of various corneal pathologies. It thus appears to be an alternative to penetrating keratoplasty in some patients. PMID- 10863893 TI - Reconstruction of the pelvic brim and its role in the reduction accuracy of displaced T-shaped acetabular fracture. AB - Open reduction of the displaced T-shaped acetabular fracture has a problem of accuracy of the fracture reduction. This study was carried out to demonstrate that the reconstruction of the pelvic brim by approaching the pubo-acetabular fragment plays a role in the accuracy of the reduction of displaced T-shaped acetabular fractures. From 1975 to 1990, a retrospective study was carried out of 22 patients who sustained a displaced T-shaped acetabular fracture. The patients were operated on by open reduction and internal fixation of the ischio-acetabular fragment to the posterior column without restoration of the pelvic brim. Radiographs of the pelvis were reviewed. The result showed that there was displacement of the pubo-acetabular fragment including the medial wall in all cases. As the result of this study, a prospective study between 1990 and 1997 was carried out of 15 patients who sustained displaced T-shaped acetabular fractures including 3 cases with medial displacement of the femoral head. The pubo acetabular fragment was anatomically reduced and fixed to the anterior column of the acetabulumn as the first approach to restore a disrupted pelvic brim. There, patterns of the acetabular fracture were subsequently re-evaluated especially the ischio-acetabular fragment including the position of the femoral head by using an intraoperative portable X-ray technique. The stability of the hip joint was assessed by hip flexion. The intraoperative radiograph appearances of the ischio acetabular fragment were visually confirmed by a second surgical exposure. The results showed that the intraoperative radiographs gave spontaneous reduction of the ischio-acetabular fragment in all patients except one. There was a reduction of the displaced femoral head into the hip socket in the three patients. The hip joints were stable in all patients. The second surgical exposure showed that there was good spontaneous reduction of the ischio-acetabular fragment to the posterior column by ligamentotaxis in 14 patients. Therefore, it is not necessary to address the ischio-acetabular fragment. In the exceptional case, the ischio acetabular fragment was displaced as a free bone which could not be reduced by ligamentotaxis. However, reduction and internal fixation of the ischio-acetabular fragment to the posterior column for complete re-application of the hip joint onto the pelvic ring of this case was facilitated. Postoperative 2 year and 5 year follow-up showed that the fracture had healed without heterotrophic ossification or premature osteoarthrosis of the hip joint. The exceptional case had a broken plate at the anterior column of the acetabulum. Hip function was evaluated clinically using Merle D' Aubigne's hip score. All patients had a "very good score". The study showed that reconstruction of the pelvic brim by anatomical reduction and fixation of the pubo-acetabular fragment to the anterior column plays an important role in the accuracy of fracture reduction of a displaced T-shaped acetabular fracture. PMID- 10863894 TI - Measurement of serum free IGF-I in diagnosis of growth hormone deficiency. AB - Serum levels of total insulin-like growth factor-I (IGF-I) are growth hormone (GH) dependent and can be used as the screening tool for GH deficient status. However, most of them are bound to IGF-binding proteins, leaving less than 1 per cent in the free or unbound forms which represent the active biological fractions. Serum free IGF-I levels were measured by radioimmunoassay (IRMA) in 48 short children with various conditions. We found that the means +/- SEM of free IGF-I in children with panhypopituitarism (PAN) and complete growth hormone deficiencies (cGHD) were significantly lower than those in sex and age matched normal children (0.02 +/- 0.01 vs 2.01 +/- 0.7 ng/ml, p = 0.0006 and 0.42 +/- 0.18 vs 1.72 +/- 0.27 ng/ml, p = 0.0007 respectively) but not in children with partial growth hormone deficiencies (pGHD) (0.91 +/- 0.3 vs 1.97 +/- 0.4 ng/ml, p = 0.27) and idiopathic short stature (ISS) (0.94 +/- 0.3 vs 1.95 +/- 0.6 ng/ml, p = 0.13). However, when we classified the pGHD children into 2 groups according to IGFBP-3 SDS for normal Thai children, we found that the mean of free IGF-I levels in pGHD children with IGFBP-3 SDS < or = -1.3 was significantly lower than that of the controls. (0.68 +/- 0.55 vs 2.66 +/- 0.71 ng/ml, p = 0.04) In conclusion, the measurement of free IGF-I level can be used to evaluate the GH status of short children and might be used as a guide when starting treatment. PMID- 10863895 TI - Timing of second polar body extrusion and pronuclear formation after intracytoplasmic sperm injection (ICSI). AB - To determine timing of second polar body extrusion and pronuclear formation to find out when the intracytoplasmic sperm injected (ICSI) oocytes should be checked for successful fertilization. 148 oocytes from 15 patients from January to May 1998 were analyzed. Metaphase II oocytes were followed by ICSI. Fertilization was checked at 2, 4, 6, 8, 16 and 18 hours later for observation of second polar body extrusion and pronuclear formation. Normal fertilization was achieved in 73.6 per cent of the oocytes (109/148). After ICSI at 2, 4, 6, 8, 16 and 18 hours, extrusion of second polar body was 8.3 per cent, 49.5 per cent, 84.4 per cent, 98.2 per cent, 98.2 per cent, 98.2 per cent and pronuclear formation was 0 per cent, 0 per cent, 8.3 per cent, 30.3 per cent, 96.3 per cent, 100 per cent respectively. The earliest extrusion of second polar body and pronuclear formation were at 2 and 6 hours after ICSI respectively. This study suggests that the appropriate time to determine fertilization is at 2, 6 and 18 hours after ICSI. PMID- 10863896 TI - Radiofrequency catheter ablation of Wolff-Parkinson-White syndrome: report of 83 cases from Siriraj Hospital. AB - We described the characteristics of patients and accessory pathway and showed our results of Radiofrequency catheter ablation (RFCA). There were 41 males and 42 females at a mean age of 36 years. Accessory pathway associated with Wolff Parkinson-White (WPW) syndrome in our population was more prevalent on the right side which is different from previous reports. Most commonly associated heart disease was Ebstein's anomaly. Overall success rate was 96.4 per cent. Right free wall accessory pathway needed a longer procedure time and fluoroscopy time, higher radiofrequency power and more radiofrequency applications compared to other locations. Although the recurrence rate was 12 per cent, all patients with recurrence were successfully reablated. We also described the comparison of our result with previous studies. To our knowledge this is the first report in Thailand with a reasonable number of cases. RFCA is a very effective treatment of WPW syndrome in the Thai population and should be considered in symptomatic patients especially those who are refractory to medication. PMID- 10863897 TI - Comparison of the Thai version of the Rose questionnaire for angina pectoris with the exercise treadmill test. AB - Prior to exercise treadmill testing (ETT), 157 patients (92 males and 65 females) were interviewed twice separately, using a Thai version of the Rose questionnaire for angina pectoris. One interview was conducted by a physician and the other by a nurse. The questionnaire responses were compared with ETT results. Based on physician-conducted interview, the Rose questionnaire had a sensitivity of 30.3 per cent, a specificity of 83.9 per cent, a positive predictive value of 35.3 per cent, a negative predictive value of 81.9 per cent, and the total accuracy of 72.6 per cent. There were gender differences in the validity of the questionnaire, with higher specificity, higher positive predictive value, and lower negative predictive value in males than in females. The sensitivity and accuracy were not different between the two sexes. In 87.9 per cent of cases, responses to physician-conducted and nurse-conducted interview were the same. There were no significant differences between responses to the questionnaires by the physicians and by the nurses. PMID- 10863898 TI - Transvaginal ultrasonography combined with pelvic examination in the diagnosis of ovarian endometrioma. AB - The aim of this retrospective study was to evaluate the efficacy of TVS and TVS combined with pelvic examination for the diagnosis of ovarian endometrioma. Three hundred and five ovarian masses of 244 patients with either pre-operative or post operative diagnosis of ovarian tumor and received TVS between January 1, 1996 and December 31, 1998 were included in the study. Of 305 masses, 221 endometriomas of 164 patients were diagnosed histologically. The efficacy of TVS was 84.9 per cent with a sensitivity of 92.3 per cent and specificity of 70.2 per cent. LR+ and LR- were 3.1 and 0.1 respectively. The combination of TVS and pelvic examination with either positive test had a higher sensitivity (98.8%) but lower specificity (26.6%). This combination dramatically improved NPV (97.5%) and LR- (0.05), whereas, the combination with both positive tests had a sensitivity of 78.1 per cent, and specificity of 81.5 per cent. LR+ and LR- were not different from those using TVS alone. In conclusion, the study has shown the role of TVS in the diagnosis of ovarian endometrioma. The combination of TVS and pelvic examination may be useful in ruling out the disease. However, a further prospective study should be performed to confirm the efficacy of the combination. PMID- 10863899 TI - Seizure threshold in ECT: I. Initial seizure threshold. AB - Seizure threshold determination is of crucial importance in optimizing electrical stimulus dosage during administering electroconvulsive therapy (ECT). We measured initial seizure threshold by means of Srinakharinwirot University titration schedule in 150 psychotic patients. Initial seizure threshold was approximately 104 millicoulombs on average, but varied widely (12-fold) across patients. Motor seizure duration was inversely related to initial seizure threshold. Seizure threshold could be strongly predicted by age. The results may have important clinical implications for stimulus dosing strategy in ECT. PMID- 10863900 TI - Association between serum homocysteine, vitamin B12, folate and Thai coronary artery disease patients. AB - BACKGROUND: Homocysteine is an intermediate compound formed during metabolism of methionine. The plasma level of homocysteine is dependent on the genetically regulated level of essential enzymes and the intake of folic acid, vitamin B6 (pyridoxine), and vitamin B12 (cobalamine). Elevated serum homocysteine levels are a known risk factor for coronary artery disease (CAD). To establish the magnitude of the CAD that is associated with an increased serum homocysteine level, we compared CAD patients with normal healthy Thai controls. METHOD: In a cross-sectional study design we investigated the association between serum homocysteine, vitamin B12 and folate levels and the coronary heart disease in 178 CAD patients and 178 normal healthy controls by age and sex matching. These comprised 266 men and 90 women, mean age 58 +/- 10 years for normal controls and 60 +/- 10 years for CAD patients. Serum homocysteine, vitamin B12 and folate were measured by ELISA method and electrochemiluminescense method respectively. RESULTS: Paired t-test analysis showed that serum homocysteine concentrations were significantly higher in CAD patients (23.83 +/- 11.29 mumol/L) than in control subjects (19.69 +/- 8.51 mumol/L; p < 0.001). Homocysteine levels were also higher in males than in females. These findings were similar in healthy controls (male: 20.37 +/- 8.5 mumol/L, female: 17.77 +/- 8.2 mumol/L, p < 0.05) and in CAD patients (male: 24.91 +/- 11.8 mumol/L, female: 20.73 +/- 8.9 mumol/L, p < 0.05). Homocysteine above 17 mumol/L occurred more common in CAD patients than in control groups (OR = 1.65, 95% CI = 1.09-2.52, p = 0.0249). Low levels of vitamin B12 and folate did not reaching statistical significance when comparing controls and CAD patients. CONCLUSIONS: Serum homocysteine concentrations were significantly higher in CAD patients than in controls. Serum vitamin B12 and serum folate levels were not statistically significantly different between CAD patients and control groups. The data also demonstrated that the serum homocysteine level is almost always higher in men than in women as previously reported. Although serum vitamin B12 and serum folate levels were not below the upper limit of normal, vitamin B12 and folic acid treatment may reduce serum homocysteine concentrations in CAD patients. We hope that the reversible risk factors will be concern to clinicians for the reduction in the risk of myocardial infarction. PMID- 10863901 TI - p53 protein expression in cutaneous squamous cell carcinoma. AB - BACKGROUND: p53 is a nucleoprotein encoded by a tumor suppressor gene. It's mutations are implicated in the genesis of a wide variety of malignant neoplasia including skin cancers. OBJECTIVE: To study the expression of the p53 protein in cutaneous squamous cell carcinoma (SCCs) and evaluate the relationships between this expression and sites, varying degrees of differentiation and amounts of apoptotic cells. METHOD: Sixty-seven tissue samples of SCCs from Songklanagarind Hospital obtained from January 1991 to December 1996 were examined by immunohistochemistry using polyclonal anti p53-CM1. (Novocastra Laboratories, Newcastle, England, dilution 1:700) RESULT: p53 Immunoreactivity was demonstrated in 26.87 per cent of SCCs. This was observed in 15/51 of sun-exposed cases and 3/16 of sun-protected cases (p = 0.401). The more differentiated the tumor, the less p53 staining was observed (p = 0.043). There was no association between p53 positivity and the amounts of apoptotic cells. CONCLUSION: The p53 expression is not related to the sun exposure. It does not represent a commitment to apoptosis. However, it may indicate the differentiation and/or proliferative status of the tumor cells. PMID- 10863902 TI - Phase II study of ifosfamide, carboplatin, etoposide and GM-CSF in small cell lung cancer. AB - Twenty patients with small cell lung cancer (SCLC) were entered to the study. Fourteen cases were male and six cases were female. Twelve cases were extensive disease, eight cases were limited disease. Median age was 60 years (range = 40-72 years), median performance status was 70 per cent (range = 60-80%). All patients were treated with combination chemotherapy consisting of ifosfamide 5 g/m2 intravenous infusion over 4 hours with mesna uroprotection, carboplatin 300 mg/m2 intravenous infusion over 2 hours on day 1, and etoposide 120 mg/m2 intravenous infusion over 4 hours on day 1-3. Chemotherapy was re-cycled every 28 days. Assessment of hematologic toxicity (CBC) was performed two times per week. If there was grade 3 or 4 neutropenia on any cycle of chemotherapy, GM-CSF was administered for febrile neutropenia and on the next cycle it was administered prophylactically on day 4-14. RESULTS: Seventeen cases were evaluable for response and toxicity (three cases were inevaluable due to loss to follow-up after the first cycle of chemotherapy). Fourteen cases (five limited disease, nine extensive disease) achieved partial response (82.5%). Two cases had stable disease, one case died on day 7. One year survival was 23.5 per cent. Seventy and a half percent grade 3 and 4 neutropenia was seen during the first cycle. One patient had febrile neutropenia. After being prophylactically treated with GM CSF, grade 3 and 4 neutropenia was reduced from 70.5 per cent to 56.2 per cent, 46.7 per cent, 63.6 per cent, 42.8 per cent and 0 per cent in cycle 2-6 respectively. Major toxicity of GM-CSF consisted of transient chest distress, chills, sweating and hypotension which subsided in 5-10 minutes. No fever or skin rash was observed. CONCLUSION: Combination of ifosfamide, carboplatin and etoposide (ICE) is an active regimen for small cell lung cancer. However, because of its severe myelosuppression, this regimen needs hematopoietic growth factor support, and GM-CSF was used in this study. The administration of GM-CSF rendered ICE chemotherapy to be given safely. PMID- 10863903 TI - Gigantic hepatocellular carcinoma, treated by transcatheter oily chemoembolization (TOCE) and wedge hepatic resection. AB - Four cases of gigantic hepatocellular carcinoma, considered by surgeons to be inoperable, were treated with repeated transcatheter chemoembolization (TOCE) until the serum alfafetoprotein reduced to normal or less than half of the original level or until the tumor reduced to less than half of the original size documented by CT scan and angiogram. Wedge hepatic resection was performed using ultrasonic dissector. Histologic section of the resected tumor mass revealed tumor necrosis. The extent of tumor necrosis was related to tumor size and corresponded inversely to the thickness of the tumor capsule. The survival periods were 48 to 108 months with only one to two episodes of recurrence during follow-up. Repeated wedge hepatic resection was performed successfully for recurrent cases. Serum alfafetoprotein (AFP) is a very sensitive and reliable tumor marker for follow-up results and appears to be a sensitive indicator for tumor recurrence. PMID- 10863904 TI - Lingual osseous choristoma: report of three cases. AB - Osseous choristoma represents the dense, mature, bony tissue in an abnormal location. We reported three cases of lingual osseous choristoma, herein, a year after the report of eight cases in 1998. All three lesions were at or close to the foramen caecum. The lesions were smooth, round or lobulated and pedunculated in shape and stony hard in consistency. All of them were preoperatively diagnosed, based on the above unique clinical findings without any radiography. The lesions were simply excised without local recurrence. PMID- 10863905 TI - Double uterus with unilaterally obstructed hemivagina and ipsilateral renal agenesis: a variety presentation and a 10-year review of the literature. AB - A double uterus with a unilaterally obstructed hemivagina is a rare condition, usually associated with ipsilateral renal agenesis. Herein, we report two cases, the first case presenting with abdominal pain and pelvic mass. Hemihysterectomy was performed leaving the contralateral uterus intact. The second case presented with chronic foul smelling vaginal discharge. The diagnosis was a double uterus and pyocolpos of the left vagina. Excision of the left vaginal septum and drainage were performed. The postoperative course of both cases was uneventful and the patients were well at the six-week follow-up. An accurate diagnosis, appropriate management and the prevention of future fertility problems are discussed. PMID- 10863906 TI - Ethical birth control. PMID- 10863907 TI - What's missing from the class? PMID- 10863908 TI - A novel method of virtual histopathology using laser-scanning confocal microscopy in-vitro with untreated fresh specimens from the gastrointestinal mucosa. AB - BACKGROUND AND STUDY AIMS: Histopathological examination for superficial gastrointestinal lesions has been mainly based upon the light microscopic examination of thin-slice specimens with hematoxylin and eosin (H&E) staining. However, it takes at least a couple of days to create a slide-glass for microscopic study. In order to obtain immediate microscopic images for untreated specimens, the authors used laser-scanning confocal microscopy (LCM) to study fresh samples of gastrointestinal mucosa. MATERIALS AND METHODS: Fresh untreated mucosal specimens from the esophagus, stomach, and colon, obtained by endoscopic pinch biopsy, polypectomy, or endoscopic mucosal resection (EMR), were fixed in normal saline and examined by LCM collecting the reflective light of a 488-nm wavelength argon laser beam. Findings from the LCM image were compared with those of conventional H&E staining in all specimens. For objective evaluation of the similarity of both pictures, the nucleus-to-cytoplasm ratio (N/C) of normal mucosa and that of cancer of the esophagus were calculated and statistically analyzed. The overall diagnostic accuracy for cancer was evaluated. RESULTS: The average scanning time to obtain the LCM image of a specimen was 1.6 seconds. The LCM images acquired corresponded well to the conventional H&E light microscopic images in the esophagus, stomach, and colon. Cell wall, nucleus, cytoplasm, and tissue structural elements were simultaneously visualized by LCM scanning. A difference in N/C ratios between normal mucosa and cancer in the esophagus was statistically apparent when Welch's test (P=0.05) was applied. The overall diagnostic accuracy of the LCM study for cancer was 89.7%. CONCLUSIONS: This novel method enables us to obtain an immediate serial virtual microscopic section through a fresh specimen, which has not actually been cut, although the resolution of the image obtained is still limited. These early results encourage us to develop imaging relevant to conventional histopathology alongside the development of LCM technology in the near future. We should aim at the in vivo application of LCM coupled to probes which can be introduced through the working channel of endoscopes. PMID- 10863909 TI - Endoscopic sphincter of Oddi manometry with a portable electronic microtransducer system: comparison with the perfusion manometry method and routine clinical application. AB - BACKGROUND AND STUDY AIMS: Endoscopic perfusion manometry (PM) of the sphincter of Oddi (SO) requires expensive equipment which is relatively large and uncomfortable to handle in the endoscopic retrograde cholangiopancreatography (ERCP) setting. Furthermore, the volume load of the biliopancreatic system may contribute to the increased risk of pancreatitis after SO manometry. PATIENTS AND METHODS: The newly developed small and lightweight microtransducer system consists of a portable data-logger with integrated online display which is connected to a 4-Fr manometry probe. The manometry catheter is inserted endoscopically into the biliopancreatic system via a 7-Fr Teflon sheath. SO motility can be observed online on the display but the data can also be stored for later analysis on a personal computer (PC). To validate the new method, 15 patients with suspected biliary SO dysfunction underwent both PM as well as microtransducer manometry (MTM) in randomized order. Thereafter, 50 consecutive patients with suspected biliary or pancreatic SO dysfunction were investigated solely by MTM. RESULTS: PM was possible in 13 of 15 cases whereas MTM could be performed in all 15 patients. The basal SO pressure tended to be lower (approximately 5 mmHg) when measured with the MTM, compared with the PM method, but there was a significant and nearly linear correlation between the basal SO pressures obtained by both methods (r=0.98, P<0.001). SO dysfunction was diagnosed in the same five patients using both methods. Furthermore, the parameters of phasic SO motility were highly comparable when measured by MTM and PM. MTM was carried out successfully in 49 of 50 patients and only one MTM probe was used for all examinations, without malfunction. The endoscopist was able to diagnose SO dysfunction (by immediate observation of SO motility on the display) in 19 of 20 patients (when compared with the later PC analysis) and SO motility was judged correctly as normal in the remaining 29 cases. MTM was repeated in five patients with SOD 1-6 weeks after the first examination and the manometric findings were confirmed in all cases. Mild postmanometry pancreatitis was observed in only one of 49 patients (2%). CONCLUSIONS: Endoscopic MTM is a reliable, safe, very easy to handle, and low-cost alternative for the clinical assessment of SO motility. PMID- 10863910 TI - Long-term outcome of endoscopic stent placement for chronic pancreatitis associated with pancreas divisum. AB - BACKGROUND AND STUDY AIMS: In pancreas divisum (PD), endoscopic drainage of the minor papilla is beneficial for patients presenting with acute recurrent pancreatitis, but in cases of chronic pancreatitis, surgery is claimed to be indicated. The aim of this study was to evaluate the efficacy of endoscopic stent placement in patients with PD presenting with chronic pancreatitis. PATIENTS AND METHODS: The outcome of endoscopic treatment was evaluated in 16 patients with PD presenting with chronic pancreatitis, who underwent stenting as a first line of treatment. Chart reviews and patient interviews by mail and telephone were conducted. Median follow-up time was 51 months (range 6-120). RESULTS: After one episode of stenting and subsequent stent extraction, five of 16 patients remained pain-free (i.e., for a median time of 45 months, range 12-64). Six patients had temporary pain relief (14 months), and five patients experienced no effect of stent therapy. A total of five patients underwent surgery after failure of stenting. CONCLUSIONS: Patients with chronic pancreatitis and a PD should undergo trial stenting, since every third patient remains symptom-free after one episode of stenting. PMID- 10863911 TI - The use of inhaled nitrous oxide for flexible sigmoidoscopy: a placebo-controlled trial. AB - BACKGROUND AND STUDY AIMS: Flexible sigmoidoscopy is a widely used diagnostic technique which is increasingly being adopted as part of bowel cancer screening programmes. It is conventionally performed without sedation or analgesia, but significant numbers of patients experience mild to moderate discomfort during the procedure. The aim of this study was to assess the usefulness of self administered nitrous oxide to reduce discomfort during flexible sigmoidoscopy, which may have an effect of improving compliance. PATIENTS AND METHODS: In a double-blind, randomized, placebo-controlled study, 87 patients were enrolled. Of these, 45 received nitrous oxide/oxygen (50% mix) to inhale during the procedure and 42 received oxygen alone. The patients recorded their opinions on the efficacy of the agent in a questionnaire after the examination. The endoscopist recorded the success of the procedure. RESULTS: The groups were well matched for age and sex. Significant reductions in levels of discomfort and increased levels of agreement to undergo the procedure again were noted for the actively treated group (P < 0.05). No significant differences in side effects or success of the procedure were observed. CONCLUSIONS: The addition of self-administered nitrous oxide offered significant benefits in the area of patient discomfort during flexible sigmoidoscopy. The availability of this agent is useful in clinical practice and may enhance compliance with a screening programme. PMID- 10863912 TI - Endoscopic treatment of benign anastomotic colorectal stenosis with electrocautery. AB - BACKGROUND AND STUDY AIMS: Benign anastomotic colorectal stenosis can occur after surgery and can require surgical or endoscopic dilation. This study aimed to investigate the use of electrocautery in this area. PATIENTS AND METHODS: We enrolled 39 consecutive patients (25 men, 14 women; mean age 61, range 48-77) suffering from anastomotic colorectal stenosis, demonstrated by colonoscopy (19 patients) or by barium enema (20 patients). We performed endoscopic dilation of the strictures with electrocautery once only in each patient. RESULTS: In all patients we obtained dilation of the strictures without any immediate or delayed procedure-related complications. No recurrence of the stenosis was demonstrated during a follow-up of a mean of 25 months (range 8-43 months). CONCLUSIONS: This study shows that endoscopic electrocautery is a safe and effective treatment of anastomotic colorectal stenosis. PMID- 10863913 TI - 3-D vision improves performance in a pelvic trainer. AB - BACKGROUND AND STUDY AIMS: Experienced endoscopic surgeons have adapted to the absence of depth perception while using two-dimensional (2-D) visualization. However, three-dimensional (3-D) vision may prove useful at least at the beginning of the learning curve in celioscopic training. METHODS: In a pelvitrainer with a fixed camera, two skill tests were designed to assess the performance of three groups of operators: "non-surgeons", "non-celioscopist surgeons", and "trained celioscopists". In the first test, the candidate had to touch with a needle a sequence of dots distributed on a 7.1-cm2 area. In the second test, a 6-0 C-1 needle had to be passed consecutively through two 1-mm holes made in a thin vertical plastic wall. Each test was performed ten times, using either 2-D vision (five times) or 3-D vision (five times) interspersed in a random manner. RESULTS: In both tests and within each group, performance was related to the experience of the operator, with the trained celioscopists' group obtaining the best results and the non-surgeons the worst. In every situation, including the trained celioscopists' group, 3-D vision significantly improved performances. No significant difference was observed between the results of the non-celioscopist surgeons' group using 3-D vision and those of the trained celioscopists' group using 2-D vision. CONCLUSIONS: 3-D vision improves the performance and accuracy of endoscopic surgeons. It provides a visual perception "close to reality", and helps celioscopic beginners to accelerate their training. PMID- 10863914 TI - Endoscopic dilation for treatment of anastomotic leaks following transhiatal esophagectomy. AB - BACKGROUND AND STUDY AIMS: Anastomotic leak is a known complication after transhiatal esophagectomy (THE) and cervical esophagogastric anastomosis. Conservative management takes a long time to heal such leaks. We assessed the role of endoscopic dilation in patients with anastomotic leak following THE. PATIENTS AND METHODS: Eight consecutive patients (seven men, one woman; mean age 51) with anastomotic leak following THE were subjected to endoscopic dilation using Savary Gilliard dilators of 7-15 mm diameter. The mean interval between surgery and detection of leak was 9 days (range 5-22 days) and dilation was performed at a mean interval of 11.4 days (range 1-20 days) after detection of the leak. RESULTS: Drainage from fistulas stopped completely after 1-8 days (mean 3 days). X-ray with water soluble contrast showed closure of the fistula in all cases. Duration of follow-up ranged from 2 to 12 months. Anastomotic strictures developed in three patients. These patients required three sessions each of repeat dilation, and were alive at follow-up periods of 2, 4, and 12 months, respectively. One patient developed recurrence of growth at an anastomotic site. Four patients died because of distant metastasis. CONCLUSIONS: Bougie dilation of anastomotic sites is a safe and effective technique for the healing of anastomotic leaks following THE. However there is a need for a prospective randomized trial comparing endoscopic dilation with no dilation in patients with anastomotic leaks following THE. PMID- 10863915 TI - Mucinous cystadenomas and intraductal papillary mucinous tumors of the pancreas in magnetic resonance cholangiopancreatography. AB - BACKGROUND AND STUDY AIMS: In mucin-producing tumors of the pancreas, diagnosis using endoscopic retrograde cholangiopancreatography (ERCP) is limited to cystic formations that communicate with the main pancreatic duct. Magnetic resonance cholangiopancreatography (MRCP) is a new, sophisticated method which is currently under evaluation. The authors describe the usefulness of MRCP in diagnosis of mucin-producing tumors. PATIENTS AND METHODS: Six patients with mucin-producing tumors were investigated using MRCP and ERCP. Imaging was compared with surgery and histopathological examinations. RESULTS: Three patients were found to have mucinous cystadenomas (MC), two patients had intraductal papillary mucinous tumors (IPMT) and one patient had a cystadenocarcinoma. MRCP demonstrated the cystic formations in all patients. Magnetic resonance imaging (MRI) showed contrast-mediated enhancement of the cystic wall in patients with MC, and visualized the pancreatic ducts completely in patients with IPMT. ERCP failed to visualize the cystic lesion in one patient with MC of the pancreatic tail. Furthermore, ERCP showed evidence of IPMT in dilated main ducts with multiple filling defects but did not visualize the ducts completely. CONCLUSIONS: MRCP provides visualization of pancreatic ducts, extraductal variations, and cystic formations more completely than ERCP does. It avoids complications seen in ERCP. MRCP may replace ERCP in the evaluation of mucin-producing tumors of the pancreas. PMID- 10863916 TI - Feasibility study on endoscopic suture with the combination of a distal attachment and a rotatable clip for complications of endoscopic resection in the large intestine. AB - BACKGROUND AND STUDY AIMS: Endoscopic resection has been more frequently performed for increasingly larger intramucosal tumors of the large intestine in recent years. It is reasonable to expect that the larger the resected mucosal surface, the greater is the likelihood of complications such as bleeding or perforation. The aim of this study was to explore the feasibility of endoscopic suture with a distal attachment and a rotatable clip-fixing device for complications of endoscopic resection in the large intestine. PATIENTS AND METHODS: The study population consisted of 15 patients who underwent endoscopic clipping therapy following endoscopic resection for intramucosal tumors of the large intestine. With a distal attachment fitted to the distal end of the endoscope, the optimal position for clipping was ensured by pressing the intestinal wall and deflating the intraluminal air little by little. With a rotatable clip-fixing device, the resection site was endoscopically sutured clip by clip. RESULTS: Tumors were of the laterally spreading tumor type in six patients, small sessile polyps in three, and pedunculated polyps in six. The complications consisted of bleeding in 12 patients, overt perforation in one, and latent perforation in two; 14 of the 15 patients underwent successful suturing by this method. The number of clips used ranged from two to seven (mean 4.4). No patients had further complications after the treatment. CONCLUSION: We conclude that endoscopic suture with the combination of a distal attachment and a rotatable clip-fixing device is very useful for complications of endoscopic resection in the large intestine. PMID- 10863917 TI - Sex and familiarity of colonoscopists: patient preferences. AB - BACKGROUND AND STUDY AIMS: The gender preference of patients for doctors has been noted in many areas of medicine, but has never been studied in those undergoing the potentially embarrassing procedures of colonoscopy and sigmoidoscopy. We were interested in whether patients did express a preference for either the sex of a doctor, or a doctor whom they had previously met, so that we might in future consider offering a choice of endoscopist. PATIENTS AND METHODS: A total of 101 patients attending for colonoscopy or flexible sigmoidoscopy were prospectively asked to complete an anonymous questionnaire enquiring about gender preference and preference for an endoscopist with whom they were familiar. The questionnaire also asked about the patient's age and whether patients felt sufficiently strongly about their choice that they would be prepared to wait longer for the procedure if necessary. RESULTS: The response rate for completion of the questionnaire was 100% (65 female patients and 34 male patients, with two patients not disclosing their sex). Among the female patients, 48% preferred a female colonoscopist, whereas no male patients preferred a male colonoscopist (chi2 = 26.8, df = 2; P < 0.0001). Women also felt more strongly about seeing a familiar colonoscopist, with 56% of women and 35% of men preferring the same doctor they had seen in the outpatient department to perform the test (chi2 = 21.2, df = 2, P < 0.0001). CONCLUSION: By offering the choice of a female endoscopist to female patients, and a doctor that the patient has already met in the outpatient department to perform the procedure, the stress of lower gastrointestinal endoscopy might be minimized. This might reduce the proportion of patients failing to attend for booked procedures and thereby increase both the detection of serious pathology and the efficiency of the endoscopy unit. PMID- 10863918 TI - Quality assurance in gastrointestinal endoscopy. PMID- 10863919 TI - Successful endoscopic resection of intramural metastasis after esophagectomy for esophageal cancer: a case report. AB - Carcinoma of the esophagus is often accompanied by intramural metastasis (IM) at the time of diagnosis, and the prognosis of patients with such metastasis is very poor. Here we report the case of a 60-year-old man who presented with a submucosal tumor in the wall of the cervical esophagus at 2 years after esophagectomy for thoracic esophageal cancer. The tumor was resected endoscopically and was histologically shown to be an IM of esophageal cancer. He has shown no recurrence and no metastasis during 2 years of follow-up after endoscopic resection and radiation therapy. PMID- 10863920 TI - Endoscopic removal of an embedded biliary Wallstent by piecemeal extraction. AB - Expandable metal biliary stents are reserved for patients with unresectable malignant biliary obstruction. Occasionally, these stents may cause complications necessitating removal. We describe successful endoscopic removal of a biliary Wallstent one year after insertion in a patient who initially underwent placement of an expandable metal biliary stent for presumed biliary malignancy. The stent was removed after a stent related bleeding duodenal ulcer formed. PMID- 10863921 TI - Duodenal perforation by a Wallstent. AB - Endoscopic and/or radiological insertion of metallic mesh stents has recently been described as an alternative to palliative bypass operation in patients with gastric outlet obstruction caused by advanced malignant disease. We report a complication caused by migration and late perforation of the duodenum by a Wallstent, which raised concerns about the place of this procedure as an alternative to surgical bypass. The Wallstents have sharp ends and could cause ulcers or perforation of the viscus. Modification of the current Wallstent design may be needed. PMID- 10863922 TI - Guidelines of the French Society of Digestive Endoscopy: monitoring of Barrett's esophagus. The Council of the French Society of Digestive Endoscopy. PMID- 10863923 TI - Soehendra's Stent Retriever as a pusher. PMID- 10863924 TI - Botox and Dysport are distinct. PMID- 10863925 TI - Small-unit colonoscopy and quality assurance. PMID- 10863926 TI - A novel endoscopic appearance of idiopathic eosinophilic esophagitis. PMID- 10863927 TI - Successful laparoscopic rescue of an impacted lithotriptor basket from the common bile duct. PMID- 10863928 TI - Deflated adjustable gastric band: migration through anterior gastric wall. PMID- 10863929 TI - Incomplete tubular duplication of the esophagus lined by heterotopic gastric epithelium, presenting in adulthood. PMID- 10863930 TI - Temporomandibular joint dislocation during endoscopic retrograde cholangiopancreatography examination. PMID- 10863931 TI - Cholecystobronchial fistula. PMID- 10863932 TI - Endosonographic features of solitary gastric hamartomatous polyp. PMID- 10863934 TI - Structure, biological activity and membrane partitioning of analogs of the isoprenylated a-factor mating peptide of Saccharomyces cerevisiae. AB - Previous biochemical investigations on the Saccharomyces cerevisiae a-factor indicated that this lipopeptide pheromone [YIIKGVFWDPAC(farnesyl)OMe] might adopt a type II beta-turn at positions 4 and 5 of the peptide sequence. To test this hypothesis, we synthesized five analogs of a-factor, in which residues at positions 4 and 5 were replaced with: L-Pro4(I); D-Pro4(II); L-Pro4-D-Ala5(III); D-Pro4-L-Ala5(IV); or Nle4(V). Analogs were purified to > 99% homogeneity as evidenced by HPLC and TLC and were characterized by mass spectrometry and amino acid analysis. Using a growth arrest assay the conformationally restricted a factor analogs I and III were found to be almost 50-fold more active than the diastereometric homologs II and IV and were equally active to wild-type a-factor. Replacement of Lys4 with the isosteric Nle4 almost abolished the activity of the pheromone. Thus, the incorporation of residues that promote a type II beta-turn compensated for the loss of the favorable contribution of the Lys4 side chain to pheromone activity. CD spectra on these peptides suggested that they were essentially disordered in both TFE/H2O and in the presence of DMPC vesicles. There was no correlation between CD peak shape and biological activity. Using fluorescence spectroscopy we measured the interaction of lipid vesicles with these position 4 and 5 analogs as well as with three a-factor analogs with a modified farnesyl group. The results indicated that modifications of both the peptide sequence and the lipid moiety affect partitioning into lipid, and that no correlation existed between the propensity of a pheromone to partition into the lipid and its biological activity. PMID- 10863933 TI - A comparison of folding techniques in the chemical synthesis of the epidermal growth factor-like domain in neu differentiation factor alpha/beta. AB - The 52-residue alpha/beta chimera of the epidermal growth factor-like domain in neu differentiation factor (NDFealpha/beta) has been synthesized and folded to form a three disulfide bridge (Cys182-Cys196, Cys190-Cys210, Cys212-Cys221) containing peptide. We investigated two general strategies for the formation of the intramolecular disulfide bridges including, the single-step approach, which used fully deprotected and reduced peptide, and a sequential approach that relied on orthogonal cysteine protection in which specific pairs are excluded from the first oxidation step. Because there are 15 possible disulfide bridge arrangements in a peptide with six cysteines, the one-step approach may not always provide the desired disulfide pairing. Here, we compare the single-step approach with a systematic evaluation of the sequential approach. We employed the acetamidomethyl group to protect each pair of cysteines involved in disulfide bridges, i.e. Cys182 to Cys196, Cys190 to Cys210 and Cys212 to Cys221. This reduced the number of possible disulfide patterns from 15 to three in the first folding step. We compared the efficiencies of folding for each protected pair using RP-HPLC, mapped the disulfide connectivity of the predominant product and then formed the final disulfide from the partially folded intermediate via 12 oxidation. Only the peptide having the Cys182-Cys196 pair blocked with acetamidomethyl forms the desired disulfide isomer (Cys190-Cys210/Cys212-Cys221) as a single homogeneous product. By optimizing both approaches, as well as other steps in the synthesis, we can now rapidly provide large-scale syntheses of NDFealpha/beta and other novel EGF-like peptides. PMID- 10863935 TI - Solution synthesis and biological activity of human pleiotrophin, a novel heparin binding neurotrophic factor consisting of 136 amino acid residues with five disulfide bonds. AB - Human pleiotrophin (hPTN), a novel heparin-binding neurotrophic factor consisting of 136 amino acid residues with five intramolecular disulfide bonds, was synthesized by solution procedure in order to demonstrate the utility of our strategy using our newly developed solvent system, a mixture of trifluoroethanol (TFE) and dichloromethane (DCM) or chloroform (CHL). The final protected peptide was synthesized by coupling two larger protected intermediates, Boc-(1-64)-OH and H-(65-136)-OBzl, in CHL/TFE (3:1; v/v) using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) in the presence of 3,4-dihydro-3-hydroxy-4-oxo-1,2,3 benzotriazine (HOOBt). After removal of all protecting groups using the HF procedure followed by treatment with Hg(OAc)2, the fully deprotected peptide was subjected to an oxidative folding reaction. The product was confirmed as having the correct disulfide structure by examining the cystine peptides obtained by enzymatic digestions, and as possessing the same biological activities as those of the natural product. The N- and C-terminal half domains (1-64 and 65-136) were also synthesized, and measurement of their biological activities indicated that the C-terminal half domain displays almost all the activities of the full-length molecule, whereas the N-terminal half domain shows almost no activity. From these results, we were able to confirm that the C-terminal half domain is responsible for the expression of biological activities in the same manner as human midkine (hMK), another heparin-binding neurotrophic growth factor. PMID- 10863936 TI - Conformational characterization of a helix-nucleated bicyclic GCN4 decapeptide by proton NMR. AB - A bicyclic decapeptide, GCN4brM1, which was designed to be a helix-locked analog of the DNA-binding basic region from the yeast transcription factor GCN4, was synthesized and characterized using circular dichroism (CD) spectropolarimetry and 1H-NMR. This peptide has two Lys(i), Asp(i+4) side chain lactam bridges incorporated into its structure in overlapping positions in the peptide chain, linking residues 3 and 7 and residues 4 and 8. CD spectra of GCN4brM1 in aqueous solution are consistent with the expected helical conformation, and indicate that this conformation is remarkably resistant to heat denaturation and is essentially unchanged by addition of 50% (v/v) trifluoroethanol (TFE) as cosolvent. NMR spectra measured in aqueous solution at -5 degrees C show long-range nuclear Overhauser effects (NOEs) that are consistent with an alpha-helical conformation throughout the peptide structure. The measured 3J(HN) coupling constants are also in agreement with an alpha-helical structure. Extremely slow proton-deuterium exchange rates measured for backbone amides in the middle of the peptide indicate that this helix is highly stabilized and rarely unfolds within the side chain bridged sequence. NOE-constrained molecular dynamics simulations gave rise to a single family of converged structures that are fully alpha-helical throughout the GCN4brM1 backbone, and show a single, well-defined conformation for the two side chain bridges. This study demonstrates that two overlapping Lys(i), Asp(i+4) lactam bridges, positioned in consecutive residue positions in a hexapeptide segment, form a rigid alpha-helical structure in aqueous solution that is propagated in both the N-terminal and C-terminal directions. PMID- 10863938 TI - The adult and adolescent clinic for inborn errors of metabolism. AB - Adolescent and adult clinics for patients with inborn metabolic errors are needed first to provide care to patients who develop distinctive problems as their different disorders impinge on education, career and family life. The selection of patients in this article illustrates the range of the problems. Fundamental research will be a shared interest with paediatricians and others, but outcomes of care and treatment in young adults and in patients in middle life must be a special concern of adult clinics. It is essential to gather that information. The best model is probably an outpatient service for all ages in a single centre with paediatricians and adult physicians contributing and sharing access to dietary and laboratory services. Such a model would be best for generating the meetings, case discussions and initiatives that contribute to a thriving and energetic unit. Clearly identified centres in each country would be an effective means of acquiring the information on the adult follow-up of these uncommon disorders. PMID- 10863937 TI - Genetic hypoglycaemia in infancy and childhood: pathophysiology and diagnosis. PMID- 10863939 TI - Inborn errors of metabolism and pregnancy. AB - An increasing number of women with inborn errors of metabolism are now reaching child-bearing age. For certain disorders there are maternal risks associated with pregnancy. These may be related to an increased likelihood of metabolic decompensation (e.g. disorders of the urea cycle) or to increased stress to systems already compromised by disease (e.g. cardiomyopathy in GSD III). Detrimental effects upon the fetus may also be caused by maternal disease, as occurs with phenylketonuria, or from medication used to treat the mother's condition. Less commonly, fetal inborn errors may adversely effect the mother's health--e.g. fetal long-chain acyl-CoA dehydrogenase deficiency and the maternal HELLP syndrome (haemolysis, elevated liver enzymes and low platelets) and AFLP (acute fatty liver of pregnancy). Because of the rarity of individual disorders, our knowledge of risks associated with pregnancy is limited. Even for more common inborn errors such as phenylketonuria, there remain a number of questions that have not been fully answered. PMID- 10863940 TI - Glucose transporters: structure, function and consequences of deficiency. AB - There are two mechanisms for glucose transport across cell membranes. In the intestine and renal proximal tubule, glucose is transported against a concentration gradient by a secondary active transport mechanism in which glucose is cotransported with sodium ions. In all other cells, glucose transport is mediated by one or more of the members of the closely related GLUT family of glucose transporters. The pattern of expression of the GLUT transporters in different tissues is related to the different roles of glucose metabolism in different tissues. Primary defects in glucose transport all appear to be extremely rare and not all possible deficiencies have been identified. Deficiency of the secondary active sodium/glucose transporters result in glucose/galactose malabsorption or congenital renal glycosuria. GLUT1 deficiency produces a seizure disorder with low glucose concentration in cerebrospinal fluid and GLUT2 deficiency is the basis of the Fanconi-Bickel syndrome, which resembles type I glycogen storage disease. PMID- 10863941 TI - Disorders related to mitochondrial membranes: pathology of the respiratory chain and neurodegeneration. AB - Faulty oxidative phosphorylation (OXPHOS) is observed in a number of mitochondrial disorders, and may be associated with single or multiple defects of the five complexes of the respiratory chain. From the genetic standpoint, the respiratory chain is unique as it is formed by means of the complementation of two separate genetic systems: the nuclear genome and the mitochondrial genome. The nuclear genome encodes most of the protein subunits of the respiratory complexes and most of the mtDNA replication and expression systems, whereas the mitochondrial genome encodes only 13 OXPHOS subunits and some RNA components of the mitochondrial translation apparatus. Accordingly, mitochondrial disorders due to defects in oxidative phosphorylation include both Mendelian-inherited and cytoplasmic-inherited diseases. Although our knowledge of these diseases has grown at an impressive rate in the past few years, it is worth noting that, as a result of complementation in the respiratory chain of nuclear-encoded and mitochondrially-encoded polypeptides, it is often difficult to establish a precise relationship between genomic mutations and biochemical phenotypes, or to distinguish pathogenic mutations from polymorphic variants in gene sequences. This review deals with human diseases due to mutations of the structural components of the respiratory chain, particularly focusing on the recently identified disorders caused either by mutations in nuclear genes encoding subunits of the subcomplexes or by mutations in nuclear genes affecting the functional efficiency, homeostasis, and assembly of respiratory chain subcomplexes. To better focus on OXPHOS genotype-phenotype correlations, mutations of the mtDNA-encoded structural genes are also discussed. PMID- 10863942 TI - Disorders related to peroxisomal membranes. AB - The peroxisome is a ubiquitous, subcellular organelle participating in a diverse array of metabolic pathways. The peroxisomal membrane and its components play a key role in organelle assembly and functions. Disorders related to peroxisomal membranes are the peroxisome biogenesis disorders and X-linked adrenoleukodystrophy. Identification and functional characterization of these disease genes is proceeding at rapid pace helped immeasurably by work in various yeast model systems. The ultimate goal is to elucidate how the encoded proteins interact to produce apparently normal and functioning peroxisomes. The achievement of this goal will lead to a better understanding of disease pathogenesis and hopefully open therapeutic options. PMID- 10863943 TI - Therapy of X-linked adrenoleukodystrophy: prognosis based upon age and MRI abnormality and plans for placebo-controlled trials. AB - Evaluation of the therapy of X-linked adrenoleukodystrophy (X-ALD) is hampered by its rarity and by the striking and unpredictable variation in phenotypic expression. We present two approaches that may facilitate therapy evaluation. (1) We have analysed data on 377 X-ALD patients who have been followed at the Kennedy Krieger Institute for a mean period of 38 months and have subdivided them into 18 subgroups on the basis of age and the degree of abnormality in brain magnetic resonance imaging (MRI) as assessed by the Loes score (Am. J. Neuroradrol 1994; 15: 1761). We find that grouping on the basis of age and MRI score provides information that is of significant prognostic value. (2) We present plans for the development of a placebo-controlled multicentre international study that will have sufficient biostatistical power to provide objective evaluation of new therapeutic interventions. PMID- 10863944 TI - Lysosomal transport disorders. AB - In the group of lysosomal storage diseases, transport disorders occupy a special place because they represent rare examples of inborn errors of metabolism caused by a defect of an intracellular membrane transporter. In particular, two disorders are caused by a proven defect in carrier-mediated transport of metabolites: cystinosis and the group of sialic acid storage disorders (SASD). The recent identification of the gene mutations for both disorders will improve patient diagnosis and shed light on new physiological mechanisms of intracellular trafficking. PMID- 10863945 TI - Gene therapy/cell therapy for lysosomal storage disease. AB - Lysosomal storage diseases (LSD) are considered to be appropriate disorders for gene therapy/cell therapy. We are attempting to treat one of these disorders using a mouse model, the Sly mouse. This is an authentic model for human beta glucuronidase deficiency, MPS VII. We have carried out two types of experimental protocols; in vivo gene therapy and ex vivo gene therapy using Sly mice. For in vivo gene therapy, we produced a recombinant adenovirus that expresses human beta glucuronidase and administered this to Sly mice intravenously. The beta glucuronidase activities in liver and spleen were elevated to 40% and 20%, respectively, of the heterozygote enzyme level at day 16. Expression persisted for at least 35 days. Pathological abnormalities improved in these tissues and urinary glycosaminoglycan excretion was reduced in treated animals. Ex vivo gene therapy/cell therapy was carried out using macrophages obtained by cultivation of bone marrow cells. Non-myeloablated macrophages from normal mice were transplanted into Sly mice, and after 7 days donor cells had populated the liver and spleen. The human beta-glucuronidase (HBG) activity was increased in liver and spleen, although these enzyme activities subsequently fell by 38 days. The pathological improvement in Sly mice was evident at day 38 post transplantation. Furthermore, the macrophages from Sly mice were treated with retrovirus/adenovirus vector expressing HBG activity and the glycosaminoglycan accumulation was markedly decreased after 5 weeks. These data suggest that genetically engineered macrophage transplantation may be a very useful form of ex vivo gene therapy for lysosomal storage diseases. We also discuss the possible treatment of the CNS involvement in lysosomal storage diseases by gene therapy/cell therapy. PMID- 10863946 TI - Porcine antimicrobial peptides: new prospects for ancient molecules of host defense. AB - Antimicrobial peptides (AMPs) are small, endogenous, polycationic molecules that constitute a ubiquitous and significant component of innate immunity. These natural antibiotics have broad microbicidal activity against various bacteria, fungi, and enveloped viruses. Because most AMPs kill bacteria by physical disruption of cell membranes, which may prevent microorganisms from developing resistance against these agents, they are being explored as possible alternatives to conventional antibiotics. Pigs, like many other mammals, produce an impressive array of AMPs, which are synthesized predominantly by host leukocytic phagocytes or mucosal epithelial cells. Currently, more than a dozen distinct porcine AMPs have been identified and a majority belongs to the cathelicidin family. This review briefly summarizes recent advances in porcine AMP research with an emphasis on the diverse biological functions of each peptide. Mechanisms of action of these AMPs and their role in the resistance to infections are considered. Finally, the current status of pharmaceutical and agricultural uses of AMPs as well as future prospects for their application in the food animal industry is discussed. PMID- 10863947 TI - Concrete use of the joint coordinate system for the quantification of articular rotations in the digital joints of the horse. AB - A method is detailed allowing the computation of three-dimensional (3D) joint angles. Each joint of the equine digit is modelled as a sequence of three single axis rotary joints. The Joint Coordinate System was used; it involves a specific sequence of cardanic angles. The decomposition of the angles was chosen so that the three elementary angles coincide with the flexion/extension, passive abduction/adduction and lateral/medial rotations. The algorithms and kinematic procedures were described for the equine front digital joints. This method was tested in vitro on four forelimbs. For each limb, angle values were measured while the member was loaded by a press (from 500 to 6000 N). These tests were repeated while a wedge raised one part of the hoof (toe, heel, lateral and medial sides) in order to induce modifications of the angular patterns of the joints. This method allowed a precise quantitative determination of 3D joint movements. The modifications occurring with the wedges are clearly identified and confirm some previously published semi-quantitative observations. Moreover, this method provides a way to collect objective data on the functional anatomy of joints and could be used to study connective shoeing thoroughly. It may be directly applied to other species and may be used by researchers interested in discreet articular movements, especially occurring in other planes than the sagittal one. PMID- 10863948 TI - Herd factors associated with the seroprevalences of four major respiratory pathogens in slaughter pigs from farrow-to-finish pig herds. AB - The objective of this study was to investigate sero-epidemiological aspects of Mycoplasma hyopneumoniae (Mh), influenza H1N1 and H3N2 viruses and Aujeszky disease virus (ADV) in fattening pigs from 150 randomly selected farrow-to-finish pig herds. Different herd factors were examined as potential risk indicators for the percentage of pigs with antibodies against the 4 pathogens. The median within herd seroprevalences of the pathogens were: Mh 76%, H1N1 100%, H3N2 40% and ADV 53%. There was a positive association between the seroprevalences of both influenza viruses, and a negative association between the seroprevalences of ADV and H1N1. The percentage of pigs seropositive for Mh increased with the purchase of gilts and with the season (slaughter date in March-April). The within-herd seroprevalences of both influenza viruses were higher in the case of a higher density of pig herds in the municipality. A higher number of fattening pigs per pen additionally increased the risk of being seropositive for H3N2. The percentage of pigs with anti-gE-antibodies against the wild type ADV increased with higher airspace stocking density in the finishing unit, increasing herd size, increasing number of pig herds in the municipality and slaughter date in March-April. Increased seroprevalences for these 4 respiratory pathogens were mostly associated with pig density in the herd and its vicinity, the winter period, and with the purchase of gilts. Purchase of gilts, number of fattening pigs per pen and airspace stocking density are risk factors that can be managed directly by farmers striving to attain a high respiratory health status of pigs. PMID- 10863949 TI - Inoculation of lactating ewes by the intramammary route with Mycoplasma agalactiae: comparative pathogenicity of six field strains. AB - Contagious agalactia affects goats and sheep. In most infected sheep, the causal agent, Mycoplasma agalactiae, induces mastitis and/or agalactia, keratoconjunctivitis and arthritis. However, a few strains of M. agalactiae were isolated from tank milk from flocks without any clinical signs. The present study was undertaken to compare these apparently "asymptomatic" strains to classical virulent strains in order to assess the pathogenicity of four "asymptomatic" strains. Six groups of lactating ewes were inoculated by the intramammary route with 10(8) viable mycoplasmas of each strain. The clinical signs were regularly evaluated; the excretion of bacteria in milk and the serological response were measured. Ewes were necropsied 7 weeks after inoculation and the level of infection in retromammary lymph nodes was determined. Among the 4 apparently "asymptomatic" strains, 2 were fully virulent as were the strains isolated from discased animals, and the other 2 induced somewhat less severe clinical symptoms. The other parameters, in particular the level of excretion in milk and the level of infection of regional lymph nodes following necropsy were similar for all strains. Mean antibody response was also comparable between the apparently "asymptomatic" and virulent strains, in spite of great individual variability. This observation shows that flocks without any clinical sign from which M. agalactiae is isolated in bulk milk, must be kept under strict control since mycoplasmas may induce severe outbreaks later with changing conditions of breeding. PMID- 10863950 TI - Oral rabies vaccination of foxes with one or two delayed distributions of SAG2 baits during the spring. AB - During the spring of 1997, various protocols of rabies vaccine bait (SAG2) distribution for foxes were compared: in the first test zone, a first distribution was organised at the end of April, followed by a second distribution two weeks later; in the second test zone, there was a first distribution at the same period as for the previous zone, followed by a second distribution four weeks later, at the end of May. In two control zones, a classical single bait distribution was organised during the same periods as for the second distribution in the respective test zones. No statistical differences were observed for adult foxes or fox cubs sampled in the test and control zones neither for baits uptake nor for seroconversion rate. However, seroconversion rates observed in fox cubs population were significantly higher (P < 0.01) in areas vaccinated at the end of May (43 and 56%) compared with those vaccinated at mid-May (24 and 20%). The vaccinal efficacy of baits was also significantly (P < 0.05) increased for the fox cubs in the areas vaccinated at the end of May (46 and 57%) compared with those vaccinated at mid-May (24 and 25%). This increase in immunological response by fox cubs when vaccinating in late spring must be related to their development. In the early spring, fox cubs are generally too young to have access to baits or to be vaccinated when eating them. For most of these fox cubs, a second distribution will not constitute a booster. Therefore, in order to increase the efficient access of fox cubs to vaccine baits, Spring distribution of baits should preferably be organised during May or June rather than in April. PMID- 10863951 TI - Morphology and amplitude values of the electrocardiogram of Spanish-bred horses of different ages in the Dubois leads system. AB - The aim of this work was to record the modifications of the form and amplitude of the electrocardiographic tracings during growth using the Dubois system and to study its sensibility to these physiological changes. This work was carried out on 179 healthy, young and adult Spanish-bred horses (98 females and 81 males). One electrocardiogram (ECG) was obtained from each horse using the Dubois leads system. The bifid shape deflection of the P wave was generally more frequent than the simple one in animals at 4 months of age or older. Amplitudes of the P2 component and of the simple positive wave presented significant differences (p < 0.01) between the age groups studied: animals up to 3 months in age and older than 2 years of age (highest amplitudes), and the rest of the animals (lowest amplitudes). Significant variations (p < 0.05) were found for the QS amplitude between animals of 1 month of age, and all the other groups. The multiple range analysis did not define a clear distribution of age groups for the amplitude of the T wave. It may therefore be concluded, that in the Spanish-bred horse up to 1 month of age, the form and amplitude of the QRS complex in the Dubois leads system differ from those of older animals. Thus, this study shows the sensibility of the Dubois leads system for detecting the electrocardiographic changes related to the growing process in the Spanish-bred horse. In addition, it provides standard values of electrocardiographic parameters in the Spanish-bred horse at different stages of growth. PMID- 10863952 TI - Serological, clinical and histopathological changes in naturally infected dogs with Leishmania infantum in the Khemisset province, Morocco. AB - Canine leishmaniasis (canL) is widespread in the north of Morocco and the Leishmania infantum local strains are highly virulent. An epidemiological survey was carried out in 1993-1995 in the Khemisset province. In this region, the severity of the disease was assessed during regular visits to the identified foci by clinical examination of 323 dogs. Clinical signs were protean and occurred in various combinations. Biopsies were made on available sick dogs; the main histological changes were severe infiltration of the spleen, lymph nodes and bone marrow by mononuclear cells and hyperplasia of macrophage cells with amastigotes in their cytoplasm. The seroprevalence among 323 dog sera tested by ELISA showed a rate of 16.71%. The highest prevalence of the disease was 23.6% in the Sid El Ghandour hamlet. A comparison of the results of this study with those from the year following the first examination on the same site (Sid El Ghandour) of 67 dogs showed that the disease prevalence had not increased significantly (23.6% to 25.33%). PMID- 10863953 TI - Is the detection of anti-Rhipicephalus sanguineus (Rs24p) antibodies a valuable epidemiological tool of tick infestation in dogs? AB - Antibodies against the 24 kDa Rhipicephalus sanguineus (Rs24p) protein were detected by ELISA to evaluate the relationship between antibodies and tick infestation. The mean titer of 3 dogs that underwent 2 experimental infestations with adult ticks was transiently increased after the second infestation. There was a significant difference in mean titers between positive control dogs naturally infested with ticks and tick-naive dogs. These results suggested that anti-Rs24p antibodies detected by ELISA are a marker of tick exposure. There was no significant difference in mean titers between tick-naive dogs and seropositive dogs to Ehrlichia canis. Some dogs positive for E. canis antibodies showed, however, higher titers than most tick-naive dogs. R. sanguineus may be related to the E. canis infection in Japan. PMID- 10863954 TI - Use of rare earth elements as external markers for mean retention time measurements in ruminants. AB - The present review deals with the utilisation of rare earth (RE) elements as particulate markers for ruminant nutrition studies. RE elements have similar chemical properties. They are attractive for use as a multiple marker system because they bind tightly to plant materials. RE binding of plant cell walls is cyclic throughout the RE series, which can be explained by the filling of the 4 f electron shell. RE markers may migrate from labelled feedstuffs under gastrointestinal conditions and particularly under acidic conditions, and they may decrease the digestibility of labelled feedstuffs. The various binding techniques used for labelling particulate matter with RE elements are evaluated in order to increase the stability of bound RE in different gastrointestinal conditions. The soaking procedure of plant cell walls to be labelled in an RE chelate solution, followed by washing to remove loosely bound RE, is recommended. PMID- 10863955 TI - Lipogenic enzyme activities in subcutaneous adipose tissue and skeletal muscle from neonatal pigs consuming maternal or formula milk. AB - The influence of maternal and formula milk on lipid metabolism was studied in 7 day-old pigs. Lipid content, fatty acid composition, lipogenic enzyme activities and expression of GLUT4 mRNA were determined in subcutaneous adipose tissue and skeletal muscle from pigs that were bottle-fed formula milk (F) or sow milk (SM), or were sow-reared (SR). Bottle-fed pigs were isoenergetically fed and achieved similar daily body weight gain. SR pigs have a higher (P < 0.05) body weight gain than bottle-fed pigs. Lipid content of adipose tissue was lower (P < 0.05) in F than in SM and SR pigs. In muscle, lipid content did not differ significantly between groups. In adipose tissue, acetyl-CoA-carboxylase (CBX), fatty acid synthase (FAS), malic enzyme (ME), glucose-6-phosphate-dehydrogenase (G6PDH) and lipoprotein lipase (LPL) activities and GLUT4 mRNA levels were higher (P < 0.05) in SR than in bottle-fed pigs. In muscle, ME and G6PDH activities and GLUT4 mRNA were higher (P < 0.05) in F than in SM and SR pigs; LPL was not detected. The present study indicates that lipogenic enzyme activities and GLUT4 mRNA expression are regulated differently in subcutaneous adipose tissue and skeletal muscle in the neonatal pig. PMID- 10863956 TI - Effect of dietary polyunsaturated fatty acids on contractile function of hearts isolated from sedentary and trained rats. AB - Moderate physical training induced a decrease in arterial blood pressure in fish oil-fed rats as compared to sunflower seed oil-fed rats. The purpose of this study was to determine if these changes were due to modifications of the left ventricular function of the heart. Forty rats were fed a semi-purified diet containing either 10% sunflower seed oil or 10% fish oil (EPAX 3000TG, Pronova). Each dietary group was assigned to two sub-groups, one being constituted by sedentary animals and the other by trained animals. Training was achieved by daily running for 60 minutes at moderate intensity for three weeks. At the end of the training period, the animals were sacrificed and their hearts were immediately perfused according to the working mode. The phospholipid fatty acid composition and parameters of the left ventricular function were determined. Feeding fish oil markedly reduced the proportion of n-6 polyunsaturated fatty acids (PUFA, 18:2 n-6, 20:4 n-6, 22:4 n-6 and 22:5 n-6) in cardiac phospholipids. The n-6 PUFA were replaced by n-3 PUFA (mainly docosahexaenoic acid). In sedentary animals, the fluid dynamic (aortic and coronary flow, cardiac output) was not modified by the diet. The heart rate was reduced (-10%) in n-3 PUFA-rich hearts. Physical training did not markedly alter the polyunsaturated fatty acid profile of cardiac phospholipids. Conversely, it reduced the heart rate, aortic flow and cardiac output (-11, -21 and -14%, respectively) at a similar extent in the two dietary groups. In a second set of experiments, the training period was repeated in animals fed a commercially available diet (A103, UAR) which simultaneously provided n-6 and n-3 fatty acids. In these dietary conditions, neither the aortic flow nor the heart rate was decreased by physical exercise. These results suggest that both n-6 and n-3 PUFA in the diet are necessary to ensure a good cardiac adaptation to moderate physical training. Furthermore, the fish oil-induced decrease in arterial blood pressure in trained animals was not related to changes in cardiac contractility, but to a decrease in vascular resistances. Moderate physical training + dietary n-3 PUFA might be used to prevent hypertension and cardiovascular diseases. PMID- 10863957 TI - The effects of dexamethasone on the differentiation and the fertilisation of the germinal primordium in the chick embryo. AB - We showed that, in the chick embryo, the fertilisation of the attractive germinal epithelium by primary germ cells can be represented by a three-dimensional diagram in which the space and time co-ordinates are graduated in terms of the segmentation of the axial and paraxial mesoderm. We thus established that the differentiation of the coelomic epithelium into an attractive germinal epithelium and the fertilisation of the gonadal primordium both occur by mechanisms that are tightly linked to somitogenesis. In the continuous presence of a constant concentration of Dexamethasone, a marked inhibition of the rate of fertilisation of the gonadal primordium was observed. A mathematical analysis of the mode of action of the inhibitor revealed the progressive establishment of a competition between Dexamethasone and the molecule(s) responsible for the process of attraction. Given the chemical nature of the inhibitor, we propose that the endogenous factor that triggers the first step of the differentiation of the germinal primordium is a steroid-containing complex. PMID- 10863958 TI - Ruminal phosphorus availability from several feedstuffs measured by the nylon bag technique. AB - The present study was aimed at determining rumen phosphorus availability of some feedstuffs assessed with the nylon bag technique: forage (alfalfa), cereals (control-C wheat, formaldehyde treated-FT wheat, barley, corn), cereal by products (wheat bran, wheat distillers, corn distillers) and meals (C and FT soya bean meals, rapeseed meals and sunflower meals). Rumen phosphorus availability was not uniform amongst the feedstuffs, varying from 33.1% (FT rapeseed meal) to 84.7% (C wheat). Alfalfa phosphorus release kinetics showed high bacterial phosphorus contamination. Technological treatments affected phosphorus content of by-products by either increasing (wheat bran and distillers) or decreasing (corn distillers) after germ extraction from the seed. Formaldehyde treatment decreasing rumen phosphorus availability (from 77.2% vs. 89.4% for wheat to 33.1% vs. 64.4% for the rapeseed meal) may depreciate the phosphorus nutritional value of FT meals. PMID- 10863959 TI - The fibrous structure of the cemento-dentinal junction in human molars shown by scanning electron microscopy combined with NaOH-maceration. AB - The cemento dentinal junction was studied in acellular and cellular cementum of human mandibular third molars by scanning electron microscopy combined with NaOH maceration. Scanning electron microscopy with NaOH-maceration was applied to observe the fibrous structure in detail through long sections of the cemento dentinal junction. In macerated specimens, the cemento dentinal junction was a fibril-poor groove. Some cemental fibrils or fibril bundles penetrated the groove and appeared to intermingle with dentinal fibrils. Prolonged maceration caused detachment of the cemento-dentinal junction irrespective of fibril intermingling allowing observation of the inner cementum surface facing the root dentin. Observations suggested that the fibril intermingling was point-like and present only in places at the cemento-dentinal junction. It was established that NaOH maceration removes interfibrillar substances effectively in connective tissues and does no damage to the collagen fibril structure and architecture. This study showed the 3-dimensional fibrous structure of the cemento-dentinal junction in human mandibular third molars, and suggested that interfibrillar adhesive substances are more important than the fibril intermingling for the cemento dentinal attachment. PMID- 10863960 TI - Relevance of IgG receptor IIIb (CD16) polymorphism to handling of Porphyromonas gingivalis: implications for the pathogenesis of adult periodontitis. AB - Polymorphonuclear neutrophils (PMNs) are essential in host defense against periodontopathic bacteria such as Porphyromonas gingivalis. The uptake of immunoglobulin G (IgG)-opsonized bacteria via IgG Fc receptors (Fc gamma R) on PMN constitutes a central defense mechanism in periodontium. Fc gamma RIIIb is the most abundantly expressed Fc gamma R on PMN and is functionally polymorphic. The Fc gamma RIIIb-NA1 and IIIb-NA2 allotypes interact differently with IgG1- and IgG3-opsonized particles. We recently showed recurrence rates of adult periodontitis (AP) to be higher in patients carrying at least 1 Fc gamma RIIIb NA2 allele. In this study we evaluated the functional relevance of the Fc gamma RIIIb polymorphism to anti-P. gingivalis PMN effector functions. Our results showed Fc gamma RIIIb-NA2-carrying PMN from both patients with AP and healthy controls to be less efficient in phagocytosis and induction of oxidative burst upon interaction with IgG1- and IgG3-opsonized P. gingivalis. These functional differences between Fc gamma RIIIb-NA1 and IIIb-NA2 were observed in the presence of CD32-blocking antibody fragments, but not upon blocking CD16. Moreover, PMNs from AP patients exhibited increased Fc gamma RIIIb-allelic differences in IgG3 induced oxidative burst compared to control PMNs. These results support the concept that Fc gamma RIIIb heterogeneity may influence the clinical course of AP. PMID- 10863961 TI - The bone resorbing activity released by gingival fibroblasts isolated from patients with periodontitis is independent of interleukin-1. AB - Supernatants from gingival fibroblast cultures obtained from 14 patients with periodontal disease contained factor(s) capable of stimulating bone resorption in vitro, as assessed by the release of 45Ca from neonatal mouse calvariae. The possibility that the factor(s) was interleukin-1 alpha (IL-1 alpha), IL-1 beta or prostaglandin E2 (PGE2) was next investigated. The human fibroblast conditioned media (HFCM) stimulated PGE2 biosynthesis in bone. The stimulatory effect by HFCM on 45Ca release, however, was not affected by blocking prostaglandin biosynthesis with indomethacin. In contrast, 45Ca release induced by IL-1 alpha, IL-1 beta, thrombin and bradykinin was significantly reduced by indomethacin, whereas the effects of PTH and PTHrP were unaffected by indomethacin. The concentration of PGE2 in HFCM was too low to be solely responsible for the 45Ca release response. In addition, the amount of bone resorbing activity produced by the gingival fibroblasts was unaffected by cyclo-oxygenase inhibitors. Similar to IL-1 alpha and IL-1 beta, the stimulatory effect of HFCM was inhibited by gamma-interferon. HFCM did not stimulate cyclic AMP formation in the mouse calvarial bones. Antisera which specifically blocked human IL-1 alpha or IL-1 beta induced 45Ca release, and the specific IL-1 receptor antagonistic protein, did not inhibit the stimulatory effect of HFCM. These data show that gingival fibroblasts secrete bone resorbing factor(s) which is not due to IL-1 and which stimulates bone resorption by a prostaglandin- and cyclic AMP-independent mechanism. PMID- 10863962 TI - Polymorphonuclear leukocyte degranulation induced by leukotoxin from Actinobacillus actinomycetemcomitans. AB - The ability of leukotoxin from Actinobacillus actinomycetemcomitans to induce release of lysosomal constituents was studied with human polymorphonuclear leukocytes (PMNL). Leukotoxin purified from A. actinomycetemcomitans or bacterial cells of a leukotoxic strain were mixed with human PMNL and the suspension was incubated under anaerobic conditions. Samples were taken at certain time intervals to examine the cell morphology of PMNL by electron microscopy and the extracellular concentrations of the granule components lactoferrin and elastase by enzyme-linked immunosorbent assay (ELISA). Electron microscopy revealed that within 10 min of exposure to leukotoxin, the number of intracellular granules was markedly reduced and the remaining granules were translocated to the periphery in PMNL. At the same time, the extracellular concentrations of lactoferrin and elastase were elevated, while that of the cytosolic enzyme lactate dehydrogenase, an indicator of cell lysis, remained low. The lysosome molecules CD63 and CD66b were also exposed on the PMNL surface, indicating fusion of lysosomes with the plasma membrane. These effects were completely abolished by the addition of anti leukotoxin serum. Pre-incubation of PMNL with monoclonal antibodies to CD11a and CD18 that recognize alpha- and beta-chains of the LFA-1 integrin, a leukotoxin receptor on PMNL, inhibited the cytolysis, but not the release of granule components. The present results demonstrate the ability of A. actinomycetemcomitans leukotoxin to trigger a rapid release of lysosomal compounds in human PMNL. The release is due to an active process stimulated by the interaction of PMNL with the toxin or toxin-carrying bacteria. PMID- 10863963 TI - Adenosine regulates the production of interleukin-6 by human gingival fibroblasts via cyclic AMP/protein kinase A pathway. AB - Adenosine has been reported to alter a variety functions of the cells that participate in inflammatory responses. However, the effect(s) of adenosine on human gingival fibroblasts (HGF), one of the immunomodulator cells in inflamed periodontal lesions, remains to be established. In this study, we examined the influence of adenosine on the production of interleukin (IL)-6 by HGF. Ligation of adenosine receptors with adenosine or its related analogue, 2-chloroadenosine (2-CADO), increased IL-6 production by HGF without any other stimuli. In addition, adenosine and 2-CADO enhanced the cyclic AMP (cAMP) level in HGF as did prostaglandin E1 (PGE1) and forskolin. Interestingly, these cAMP-arising reagents and the permeable cAMP analogue, dibutyryl cAMP (dbtcAMP), also increased IL-6 production by HGF. These results suggest that cAMP is involved in adenosine induced IL-6 production by HGF. Adenosine-induced IL-6 production was suppressed by protein kinase A (PKA) inhibitor, H89, indicating that cAMP/PKA pathway is involved in the induction. Moreover, the experiments using antagonists specific for adenosine receptor subtypes revealed that the adenosine-induced IL-6 production by HGF was, at least in part, mediated by the adenosine A2b receptor. These results provide new evidence for the possible effects of adenosine or its related analogue as an immunomodulator in inflammatory periodontal lesions. PMID- 10863964 TI - Effect of interleukin-1 gene polymorphisms on gingival inflammation assessed by bleeding on probing in a periodontal maintenance population. AB - Bleeding on probing (BOP) is the most significant clinical parameter for the assessment of periodontal inflammation. The aim of this prospective longitudinal trial was to study the association between allelic variants of the IL-1 gene complex and gingival inflammation. Three hundred and twenty-three randomly selected periodontal maintenance patients (64.4% females) received a periodontal examination that included probing depth measurements and BOP at each of 4 supportive periodontal therapy (SPT) appointments. A blood sample taken from each subject was analysed for the presence of specific allotypes of the IL-1 gene complex. Two polymorphisms located at +4845 bp in the IL-1 alpha region and at +3954 bp in the IL-1 beta region were evaluated by a polymerase chain reaction method; 35.3% of the examined subjects were positive for specific combinations of allotypes of the IL-1 gene complex previously associated with an increased risk for severe periodontitis. The population consisted of 90 current smokers and 94 former smokers. An analysis of the association between the IL-1 genotype and BOP in the whole population (including smokers) did not reach statistical significance because of the overriding effect of smoking. A subset analysis of the 139 never smokers indicated that genotype positive patients had a significantly elevated chance of presenting an increase in the BOP% over a 4 appointment recall period (p = 0.03) after correcting for oral hygiene. In fact, patients who were genotype-negative had a 50% smaller chance of showing increases in BOP% during SPT. A further analysis explored the relationship between the genotype and the level of BOP% at the most recent recall visit. A generalized linear model showed a statistically significant effect of the genotype status after correcting for plaque accumulation and prevalence of residual pockets (> or = 5 mm). Genotype-negative subjects had significantly lower BOP% (p = 0.0097). It is concluded that the increased BOP prevalence and incidence observed in IL-1 genotype-positive subjects indicates that some individuals have a genetically determined hyper-inflammatory response that is expressed in the clinical response of the periodontal tissues. PMID- 10863965 TI - Primer for antimicrobial periodontal therapy. AB - Successful prevention and treatment of periodontitis is contingent upon effective control of the periodontopathic microbiota. Periodontal pathogens reside in subgingival sites but also colonize supragingival plaque, tongue dorsum and other oral sites. Controlling destructive periodontal disease warrants a comprehensive antimicrobial approach that targets periodontal pathogens in various ecological niches of the oral cavity. Also, to effectively combat periodontal pathogens, the various elements of antimicrobial periodontal therapy should be engaged within a short period of time. Scaling and root planing, with or without periodontal surgery, along with proper oral hygiene, constitute the primary approach to controlling periodontopathogens. Antimicrobial agents administered systemically or locally can help suppress periodontal pathogens in periodontal sites and in the entire mouth. Microbiological testing aids the clinician in selecting the most effective antimicrobial agent or combination of agents, and in monitoring the effectiveness of periodontal treatment. The present paper considers theoretical and practical aspects of effective antimicrobial treatment of destructive periodontal disease. PMID- 10863966 TI - Androgens and osteoporosis. AB - Androgen receptors are present in relevant numbers in osteoblasts. Stimulation of androgen receptors in osteoblastic bone marrow stromal cells inhibits the differentiation of osteoclasts in the bone marrow cavity. Androgens not only inhibit osteoclastogenesis but also increase cortical bone formation mainly by stimulating periosteal bone formation. Clinically, androgen action is crucial for the gain of bone mass during puberty and the maintenance of bone mass after puberty. Therefore, androgen replacement is necessary in hypogonadal men. However, the role of androgen replacement in partial androgen deficiency still remains unclear. Thus far, only testosterone has established its role in androgen replacement. However, further clinical and basic research should better define the selective role of androgen versus oestrogen receptor stimulation in male skeletal homeostasis. PMID- 10863967 TI - Influence of sexual stimulation on sperm parameters in semen samples collected via masturbation from normozoospermic men or cryptozoospermic men participating in an assisted reproduction programme. AB - To evaluate the influence of sexual stimulation via sexually stimulating videotaped visual images (VIM) on sperm function, two semen samples were collected from each of 19 normozoospermic men via masturbation with VIM. Two additional samples were collected from each man via masturbation without VIM. The volume of seminal plasma, total sperm count, sperm motility, percentage of morphologically normal spermatozoa, outcome of hypo-osmotic swelling test and zona-free hamster oocyte sperm penetration assay, and markers of the secretory function of prostate were significantly larger in semen samples collected via masturbation with VIM than masturbation without VIM. The improved sperm parameters in the samples collected via masturbation with VIM may reflect an enhanced prostatic secretory function and increased loading of the vas deferens at that time. In a similar protocol, two semen samples were collected via masturbation with VIM from each of 22 non-obstructed azoospermic men. Semen samples from these men had been occasionally positive in the past for a very small number of spermatozoa (cryptozoospermic men). Two additional samples were collected from each cryptozoospermic man via masturbation without VIM. The volume of seminal plasma, total sperm count, sperm motility, and a marker of the secretory function of prostate were significantly larger in semen samples collected via masturbation with VIM. Fourteen out of the 22 men were negative for spermatozoa in both samples collected via masturbation without VIM. These men demonstrated spermatozoa in both samples collected via masturbation with VIM. Six men with immotile spermatozoa in both samples collected via masturbation without VIM exposed motile spermatozoa in both samples collected via masturbation with VIM. High sexual stimulation during masturbation with VIM results in recovery of spermatozoa of greater fertilizing potential both in normozoospermic and cryptozoospermic men. The appearance of spermatozoa after masturbation with VIM in the vast majority of cryptozoospermic men is of clinical significance in programmes applying intracytoplasmic sperm injections for the management of severe male infertility and obviates the need for testicular biopsy. PMID- 10863968 TI - Sperm ultramorphology as a pathophysiological indicator of spermatogenesis in males suffering from varicocele. AB - Varicocele of spermatic veins is considered to be one of the major causes of male infertility associated with reduction of sperm quality. The pathophysiology of this condition is not yet completely understood. The aim of this study was to shed light on the pathophysiology of varicocele by identifying semen parameters, especially sperm ultramorphology, which improve following high ligation of the spermatic vein. Seventy-five males with diagnosed varicocele were included in this study. Semen parameters were assessed prospectively using light microscopy, semen biochemistry and sperm quantitative ultramorphological analysis, before high ligation and 3-9 months after high ligation. The control group consisted of twenty-five untreated varicocele patients who underwent two semen examinations within 3-9 months. No statistical difference in any of the examined variables was found between the two examinations in the control group. The treated patients exhibited a significant improvement in sperm density, progressive motility, percentage of normally formed spermatozoa, agenesis of sperm acrosome, chromatin condensation and incidence of amorphous heads compared with the pretreatment condition (P < or = 0.01). In contradiction, no significant improvement was observed following treatment in any of the sperm tail subcellular organelles. It is concluded that varicocele may cause deleterious alterations in early spermatid head differentiation during spermiogenesis and that varicocele patients with a high incidence of sperm acrosome and nucleus malformations are appropriate candidates for varicocele correction. PMID- 10863969 TI - Hormonal and seminal evaluation of Leydig cell tumour patients before and after orchiectomy. AB - Seven patients (aged 25-38 years) were admitted because of mono- or bilateral gynaecomastia. Plasma levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin, testosterone, 17-beta-estradiol, delta4-androstenedione, dehydropiandrosterone sulphate (DHEA-S) and 17-OH-progesterone were determined and semen analysis was carried out. FSH and LH levels were also measured after acute LH-RH administration (100 microg intravenously), and testosterone and 17 beta-estradiol were also evaluated after acute human chorionic gonadotrophin (hCG) administration (5000 IU intramuscularly). Testicular echography demonstrated the presence of a solid hypoechoic tumour. Therefore all patients were submitted to hemicastration by orchidofuniculotomy and a benign Leydig cell tumour was diagnosed in the removed testes. Hormonal and semen evaluations were repeated 3, 6, 9 and 12 months after surgery. The data before and after surgery were compared with a control group of 10 age-matched males. Before surgery, patients showed low FSH basal plasma levels; high levels of 17-beta-estradiol and low testosterone levels similar to those after hCG administration. A dyspermia was observed. Unilateral orchidectomy eliminated the autonomous secretion of oestrogen(s) so an increase of LH, FSH and testosterone levels, together with an improvement of spermatogenesis, were obtained. PMID- 10863970 TI - Effects of long-term treatment with human pure follicle-stimulating hormone on semen parameters and sperm-cell ultrastructure in idiopathic oligoteratoasthenozoospermia. AB - Ten subfertile men affected by idiopathic oligoteratoasthenozoospermia and exhibiting normal serum hormone levels received a long-term treatment with human pure follicle-stimulating hormone (hp-FSH) (150 IU, intramuscularly, three times per week for 6 months). Semen parameters and ultrastructural features of spermatozoa were evaluated before and after therapy. The results showed an increase in sperm cell concentration and, more interestingly, motility. Electron microscopic examination revealed an improved fine architectural pattern, mainly involving acrosome, head and chromatin and middle-piece, in accordance with the positive changes of functional data. No significant changes of hp-FSH treatment on serum hormone levels were observed, since the latter were found to be substantially unchanged after 6 months of therapy. The present data suggest: (i) the benefit of hp-FSH administration in idiopathic oligoteratoasthenozoospermia, when hormone parameters support a substantial integrity of spermatogenetic microenvironment and (ii) an optimal effect after long-term (6 months) therapy. PMID- 10863971 TI - Assessing equine sperm-membrane integrity. AB - The swelling of cells in a hypo-osmotic medium has been described as an important criterion for assessing the functional integrity of the sperm plasma membrane. The resistance of equine spermatozoa to osmolarity changes was studied by extending 98 semen samples collected from nine stallions in media at five osmolarities (300, 200, 150, 100, and 50 mOsmol l(-1)). The response of the cells was measured by the spermatocrit technique and eosin staining. Spermatocrit determines the increase on spermatozoal volume under hypo-osmotic conditions, a sign of functional integrity of sperm plasma membrane, whereas the eosin staining evaluates the viability of spermatozoa. A significant positive correlation (P<0.01) was observed between spermatocrit values and percentage of eosin unstained cells. Spermatocrit measurements and eosin staining proved to be useful methods to evaluate the integrity of sperm plasma membrane under hypo-osmotic conditions and could be used as an additional criterion to predict semen preservation ability. PMID- 10863972 TI - Electron paramagnetic resonance spectroscopy for the investigation of the fluidity of human spermatozoa plasma membranes: a feasibility study. AB - Spermatozoal membrane perturbations may play a role in abnormal sperm functions. The objective of this investigation was to study the feasibility of measuring membrane fluidity of isolated human sperm by electron paramagnetic resonance (EPR) spectroscopy and to compare the order parameter of spectra obtained from the sperm plasma membranes of living sperm of fertile men with that of infertile men. Ejaculates of infertile and fertile men were washed and the spermatozoa labelled with 5-doxylstearic acid (5-DSA) and 16-doxylstearic-acid (16-DSA) (10 nmol per 4 x 10(7) sperm). The reporter group of 5-DSA partitions into the outer, hydrophilic part of the sperm plasma membrane, whereas that of 16-DSA is distributed in the inner hydrophobic part. The following results were obtained: (i) the lowest measurable cell count was 3.6 to 7 x 10(6) sperm and the interassay variance of the order-parameter s was < 1%; (ii) swim-up experiments revealed a higher fluidity of sperm with a higher percentage of motility; (iii) sperm membranes of infertile patients exhibited a decreased fluidity of their plasma membranes in the polar interface region of 5-DSA compared with volunteer semen donors and fertile men (P=0.002). No difference of membrane fluidity was found between the different groups using 16-DSA. It is concluded that EPR spectroscopy can be used to study the fluidity of sperm plasma membranes in fertile and infertile men. PMID- 10863973 TI - A new method to produce equally sized hemizonae pellucidae for the hemizona assay. AB - The hemizona assay (HZA) is a valuable tool to study the binding potential and interaction of spermatozoa with the zona pellucida. Its accuracy strongly depends on the use of equally sized hemizonae. Usually, manipulator-guided microblades are used for cutting the zona pellucida. Recently, lasers were introduced for precise local thermolysis of the zona. The use of a 1.48 microm diode laser for the generation of hemizonae from human oocytes was investigated. This laser allowed drilling of equally sized hemizonae which were used for hemizona binding assays. It is concluded that the 1.48 microm diode laser system can be applied for the production of hemizonae. The method is easy to perform and offers a fast and efficient access to hemizonae of identical size. PMID- 10863974 TI - Veno-occlusive disease of the liver after hemopoietic cell transplantation. AB - The clinical syndrome of veno-occlusive disease (VOD) after hemopoietic cell transplantation is characterised by jaundice, painful liver enlargement, and fluid retention with weight gain. Cytoreductive therapy is presumably the primary cause of VOD, but several other agents (and a special susceptibility of the liver) can also play a role in its genesis. The risk of VOD can be predicted before BMT by analysing the presence or absence of the main risk factors. For the diagnosis of VOD most teams worldwide apply the clinical criteria developed by both the Seattle and Baltimore teams. Transjugular liver studies and some biological markers can help establish a correct differential diagnosis. In most cases clinical manifestations improve after several days, but 20-25% of patients could die of VOD. Data regarding whether or not pharmacological prophylactic measures are effective are contradictory. There a few therapeutical approaches directed towards the improvement of venular occlusion; recombinant tissue plasminogen activator and defibrotide can solve some cases of severe VOD. PMID- 10863975 TI - Two-phase liquid culture system models normal human adult erythropoiesis at the molecular level. AB - We have studied the patterns of expression of various genes during maturation of normal human adult erythroid precursors cultured in a two-phase liquid culture method. In the first phase, peripheral blood mononuclear cells are cultured for one week in the presence of a combination of growth factors, but not erythropoietin (Epo). In Phase II, Epo is included in the medium. Cell samples were taken throughout phase II, and expression of globins, transcription factors, and cytokine receptors was assayed by RT-PCR and quantified by phosphor imaging. We have divided phase II into stages: early (days 0-5), intermediate (days 6-10) and late (days 11-15) and measured maximum expression of each gene. During early phase II, gamma-globin, Spl, and GATA-2 mRNAs were expressed at their highest levels. As the cells matured during the intermediate period, GATA-2 levels remained high, and then declined, while the transcription factors GATA-1, EKLF, NF-E2, and the Epo receptor (EpoR) reached maximum expression. In late phase II, beta-globin increased and reached its maximum level of expression. This erythroid culture system appears to recapitulate normal adult erythropoiesis at the molecular level, and thus may be a suitable model to examine the molecular basis of severe congenital or acquired disorders of erythropoiesis. PMID- 10863976 TI - CD34+ cells derived from fetal liver contained a high proportion of immature megakaryocytic progenitor cells. AB - Endoreplication and maturation of the megakaryocyte (MK) may be retarded or delayed during ontogenesis. In this study, CD34+ cells were isolated from both human fetal liver and adult bone marrow and incubated with thrombopoietin (TPO). The cell number, morphological characteristics, platelet-associated antigen phenotype, maturation stage and DNA ploidy of CD41+ cells were examined from day 0 to day 12 in culture. 1) TPO stimulated the proliferation of fetal liver (FL) derived CD34+ cells with a mean 73.14-fold increase of CD41+ cells after 12 d in culture. Adult BM-derived CD34+ cells increased only slightly, with a mean 8.18 fold increase of CD41+ cells. 2) Although the membrane phenotype of both FL CD34+ derived MKs and BM CD34+ -derived MKs analyzed with CD41a, CD42a, CD61 and CD34 were similar, all FL CD34+-derived MKs were in maturation stage I and II and in low ploidy (<4N) class. By comparison, BM CD34+ MKs possessed 15% MKs in maturation stage III and IV and with 23% MKs in high ploidy class ( > 4N). 3) Most of cultured FL-derived CD34+ cells did not have a well developed demarcation system (DM) and numerous alpha-granules after 12 d incubation. von Willebrand factor (vWF) appeared earlier on the cultured BM-derived CD34+ cells than on FL derived CD34+ cells. 4) The expression of both cyclin E and cyclin B1 progressively increased in FL CD34+ cells induced by TPO during 12 d in culture. 5) The expression of cyclin D1 gradually decreased in FL CD34+ cells induced by TPO over 12 d incubation. 6) Immunocytochemical analysis showed that cyclin D3 was detected only in cytoplasm of cultured FL-derived CD34+ cells, whereas in both cytoplasm and nuclei of cultured BM-derived CD34+ cells. These data suggest that FL-derived CD34+ cells contain a high proportion of immature megakaryocytic progenitor cells. It further suggests that TPO can push these progenitor cells into proliferation by upregulating the expression of cyclins B1 and E, and drive a high proportion of cells into megakaryocytic lineage. PMID- 10863978 TI - Stromal cells in lymph nodes attract B-lymphoma cells via production of stromal cell-derived factor-1. AB - Stromal cell-derived factor-1 (SDF-1) is a chemokine produced by bone marrow stromal cells which plays an important role in B-lymphopoiesis and the homing of hematopoietic stem cells to the bone marrow. In the present study, we investigated the role of SDF-1 and its receptor, CXCR4, in the chemotactic interaction between non-Hodgkin B-lymphoma cells and lymph node stromal cells. SDF-1 mRNA was abundantly expressed in stromal cells isolated from the lymph nodes of patients with malignant lymphoma. All B-lymphoma cells freshly isolated from these patients and most laboratory B-lymphoma cell lines, including follicular, diffuse large, and Burkitt's lymphoma cells, expressed surface CXCR4 and migrated in the presence of recombinant human SDF-1alpha. Chemotaxis assays revealed that CXCR4-positive (but not CXCR4-negative) B-lymphoma cells migrated towards lymph node stromal cells, and this migration was almost completely inhibited by the addition of anti-CXCR4 monoclonal antibody to the lymphoma cells or of anti-SDF-1 neutralizing antibody to the culture supernatant of the stromal cells. Down-regulation of surface CXCR4 was detected in B-lymphoma cells which migrated towards the stromal cells but not in those which showed no migratory response. In addition, contact between the lymphoma cells and the stromal cells resulted in down-regulation of surface CXCR4 on the lymphoma cells. These data strongly suggest that SDF-1/CXCR4 is the main chemokine system involved in the chemotactic interaction between B-lymphoma cells and lymph node stromal cells. PMID- 10863977 TI - Lactacystin activates FLICE (caspase 8) protease and induces apoptosis in Fas resistant adult T-cell leukemia cell lines. AB - Lactacystin (LC) is a specific inhibitor of the proteasome, and has recently been shown to induce apoptosis in certain cell lines. In the present study, we established Fas-resistant adult T-cell leukemia (ATL) cell subclones RSO4 and RST1 from their parental Fas-sensitive cell lines SO4 and ST1, and examined whether LC can overcome Fas resistance. LC completely inhibited proteasome function as determined by a peptidyl-MCA substrate (LLVY-MCA and LLE-MCA), and induced apoptosis in these cell lines irrespective of Fas sensitivity at low concentrations (approximately 10 microM). LC induced the activation of caspase 3 (CPP32/Yama) and caspase 6 proteases in an identical manner to Fas-mediated apoptosis. Moreover, LC induced the activation of caspase 8 (FLICE) protease, which is the initiator of the Fas-mediated apoptotic cascade. Synthesized proteasome inhibitory peptide MG-115 (ZLLnV-CHO) also induced apoptosis in these cell lines. These results indicated that proteasome inhibitors overcome Fas resistance by bypassing the proximal part of the Fas signal. Inhibition of the proteasome function may be a new strategy for the treatment of ATL. PMID- 10863979 TI - Alterations in binding activity of T cell transcription factor CD28 responsive element binding complex (CD28RC) following allogeneic bone marrow transplantation. AB - The CD28 responsive element binding complex (CD28RC) has an important role in transducing CD28/B7 costimulatory signals. Using electrophoretic mobility-shift assay (EMSA), we have analyzed the binding activity of CD28RC in the mixed lymphocyte culture (MLC) using peripheral blood mononuclear cells (PBMC) obtained from the patients before and after allogeneic bone marrow transplantation (allo BMT). The binding activity of CD28RC was low in MLCs using PBMC from patients without acute GVHD and it was also low in MLCs using PBMC from patients without chronic GVHD (cGVHD). In contrast, this activity in patients with cGVHD was estimated to be high. The relative values of CD28RC in comparison with third party MLCs were significantly higher in MLCs using PBMC from patients with cGVHD than those in MLCs using PBMC from patients without GVHD (0.55+/-0.31 versus 0.23+/-0.12, respectively, n = 10, p = 0.05). IL-2 concentrations in the MLC medium from patients without GVHD were undetectable; however, a detectable level of IL-2 was present in MLC medium from a patient with extensive cGVHD. These data were interpreted to suggest that the CD28 costimulatory pathway was specifically activated against recipient antigen in allo-BMT patients with GVHD. In other words, it was suggested that the CD28 costimulatory pathway was specifically suppressed in allo-BMT patients without GVHD, and this suppression might contribute immunological tolerance after allo-BMT. PMID- 10863980 TI - Deafness from eighth cranial nerve involvement in a patient with large-cell transformation of mycosis fungoides. AB - Central nervous system (CNS) involvement by mycosis fungoides (MF) is rare and is usually seen in advanced stages with lymph node or visceral involvement. We describe a patient with advanced stage MF in large-cell transformation who presented with profound hearing loss after chemotherapy. Despite an initial differential diagnosis of vincristine-related neurotoxicity based on clinical, audiometric, and MRI investigations, CSF examination revealed lymphomatous leptomeningeal involvement. This case illustrates the importance of an awareness of the possibility of CNS involvement by MF and underlines the need for a complete neurologic evaluation including CSF examination in a patient with underlying MF who presents with a new neurological problem. PMID- 10863981 TI - Isochromosome +i(3)(q10) in a new case of persistent polyclonal B-cell lymphocytosis (PPBL) PMID- 10863982 TI - Amifostine and 2-CdA: a novel purging strategy in CML autografting? PMID- 10863983 TI - Prolonged interferon-alpha-2b treatment of hairy cell leukemia patients. PMID- 10863984 TI - Searching for common ground and shared values. PMID- 10863985 TI - Sure we believe in ethics and values: but whose? PMID- 10863986 TI - Are school based mental health services effective? Evidence from 36 inner city schools. AB - In an effort to bridge the gap between service need and service utilization, an urban based, university-affiliated children's psychiatric outpatient clinic implemented a program which provides mental health services in inner city schools. Since impressions of school and mental health personnel affirmed the effectiveness of such services, an evaluation of this program was conducted, despite the difficulties inherent in implementing research in "naturalistic settings." A clinic sample of children (N = 220) was compared with a sample served in the urban schools (N = 256). The findings revealed that both sets of children showed improvement as indicated by the Children's Global Assessment Scale (C-GAS) and Global Assessment of Functioning Scale (GAF). The improvement was comparable, even though the school children were seen for a slightly shorter period of time (an average of 5 versus 8 months) but had an equally frequent level of service (3 sessions per month in each setting). This finding may have important implications for the managed care environment. These results indicate that school based mental health services show improvement comparable to the clinic-based services, and have the potential for bridging the gap between need and utilization by reaching disadvantaged children who would otherwise not have access to these services. PMID- 10863987 TI - Attitudes of case managers toward people with serious mental illness. AB - Negative attitudes toward people who have serious mental illnesses held by mental health professionals threaten the effectiveness of psychiatric treatment. In this study, attitudes held by case managers working within the public sector were investigated. Differences between supportive and intensive case managers were compared with community controls using the Opinions about Mental Illness Scale. The results showed a complex interplay among client level of functioning, type of case management approach, case management philosophy, and attitudes. Among other findings, intensive case managers held more authoritarian attitudes than did their supportive case manager counterparts. PMID- 10863988 TI - Ethnic variations in mental health attitudes and service use among low-income African American, Latina, and European American young women. AB - This study examines the predictors of mental health service use among patients in an ethnically diverse public-care women's clinic. While waiting for their clinic appointments, 187 Latina, African American, and White women were interviewed about their attitudes towards mental illness and mental health services. White women were much more likely to have made a mental health visit in the past than the ethnic minority women. Having a substance use problem, use of mental health services by family or friends, and beliefs about causes of mental illness were all predictors of making a mental health visit. PMID- 10863989 TI - Sexual behavior of psychiatric inpatients: hospital responses and policy formulation. AB - Historically, inpatient sexuality has been a neglected issue in hospital policy. Recent studies have confirmed sexual activity among individuals with mental illness and have noted several areas of concern surrounding patient care and sexuality. We surveyed private mental health facilities within the state of Ohio. Twenty-five percent of the responding hospitals had a formal sexual policy and the majority (72%) said that sexual behavior was an infrequent problem. Considering the short-term stays common at these facilities, policies at such institutions should focus on dealing with situations when they occur and properly training and informing staff on issues surrounding sexual activity in psychiatric inpatients. PMID- 10863990 TI - Cognitive behavioral therapy of minor depressive symptoms in elderly Chinese Americans: a pilot study. AB - There is a high prevalence of suicide among elderly Chinese, and particularly among elderly Chinese women in Mainland China with a prevalence of 19.6 per hundred thousand. Since Chinese individuals may much more highly value education, a cognitive-behavioral package originated by Ricardo Munoz, Ph.D. was adapted for Chinese American subjects. The material was videotaped in eight sessions, approximately 25 minutes in length, to be shown to community subjects who were at least 40 years and over. In addition, a videotape of muscular relaxation techniques was made. A manual written in Chinese about the content of each class, was given to each subject when he/she attended. The experimental group showed significant improvement in the scores in the Hamilton Depression Scale, including the Somatic Subscale in the Hamilton Anxiety Scale. There was no significant improvement in the control group on any of the measures. Thus the study suggests the efficacy of psychoeducational classes in reducing symptoms of depression in non-patient community elderly. Other studies are being conducted among Korean Americans and Japanese Americans in the United States, and also in the Orient among Japanese elderly. PMID- 10863991 TI - Level of care utilization system for psychiatric and addiction services (LOCUS): a preliminary assessment of reliability and validity. AB - The financing of mental health care services has undergone significant change in the past several years, with increasing emphasis on resource management. This emphasis has raised concerns among consumers and providers of services that needed resources are often withheld. In order to address this issue, the American Association of Community Psychiatrists (AACP) developed an instrument designed to maintain balance between quality care and the wise use of resources. The Level of Care Utilization System for Psychiatric and Addiction Services (LOCUS) evaluates clients along six dimensions and defines six levels of resource intensity. It also provides a methodology to facilitate rapid and consistent level of care recommendations. This paper describes the circumstances leading to the creation of LOCUS and the principles used to guide its development. It then describes preliminary reliability and validity testing design and outcome. This initial testing indicates that LOCUS can facilitate consistent placement of clients in psychiatric or addiction services and does so in a manner which trends in a pattern similar to the recommendations of clinicians who are naive to LOCUS assessment and placement methodology. PMID- 10863992 TI - Engineering pancreatic islets. AB - Pancreatic islets are neuroendocrine organs that control blood glucose homeostasis. The precise interplay of a heterogeneous group of cell populations (beta, alpha, delta and PP cells) results in the fine-tuned release of counterbalanced hormones (insulin, glucagon, somatostatin and pancreatic polypeptide respectively). Under the premises of detailed knowledge of the physiological basis underlying this behaviour, two lines of investigation might be inferred: generating computational and operational models to explain and predict this behaviour and engineering islet cells to reconstruct pancreatic endocrine function. Whilst the former is being fuelled by new computational strategies, giving biophysicists the possibility of modelling a system in which new "emergent" properties appear, the latter is benefiting from the useful tools and strategic knowledge achieved by molecular, cell and developmental biologists. This includes using tumour cell lines, engineering islet cell precursors, knowledge of the mechanisms of differentiation, regeneration and growth and, finally, therapeutic cloning of human tissues. Gaining deep physiological understanding of the basis governing these processes is instrumental for engineering new pancreatic islets. PMID- 10863993 TI - Cyclic mechanical strain decreases the DNA synthesis of vascular smooth muscle cells. AB - In vivo, smooth muscle cells of the vascular wall are rhythmically stretched by the arterial pulse. Here, we test the hypothesis that rhythmical stretch is important for suppressing the growth of vascular smooth muscle (vsm) cells. DNA synthesis rate, cell number, metabolic activity, and cell death were compared between rhythmically stretched and non-stretched vsm cells from the rat embryonic aortic A10 cell line. Rhythmical stretch (0.5 Hz, 5% elongation, 48 h) did not induce vsm cell proliferation, that is the vsm cell number was constant. Cell damage or necrosis was excluded because the release of lactate dehydrogenase (LDH) was identical. The low rate of apoptosis (0.2%) was not different between stretched cells and control cells. Stretch significantly reduced the DNA synthesis rate [measured as incorporation of 5-bromo-2'-deoxyuridine (BrdU)] in a time-dependent manner. BrdU incorporation was decreased by 32% after 24 h of cyclic stretching and was further diminished to 50% after 48 h of strain. Metabolic activity (measured by Wst-1 cleavage) was only modestly influenced. The stretch-induced decrease in DNA synthesis was independent of the extracellular matrix. No differences were detected when laminin- or pronectin-coated membranes were used instead of collagen-coated membranes. The effect of stretch was unlikely to be mediated by secretion of an unknown "factor", because vsm cells incubated with medium conditioned by stretched cells did not show a significant decrease in BrdU uptake. The results support the idea that rhythmical stretch is important to keep the rate of DNA synthesis and thereby the proliferation of vsm cells at a low level. PMID- 10863994 TI - Urinary dopamine excretion in healthy volunteers: effect of sodium diet and acute water load. AB - The present study was designed to investigate, in human subjects, urinary dopamine excretion under different conditions of sodium and water homeostasis. In a cross-over trial, ten healthy volunteers were subjected to low-salt (LS; dietary salt restriction, sodium chloride (NaCl) intake <5 g per day), normal salt (NS; normal food ad libitum), and high-salt (HS; normal food plus NaCl 100 mg/kg per day) regimens for 8 days in a randomized order. On day 7, urine was collected for 24 h. The variations in urinary sodium excretion reflected the dietary salt intake (LS: 16.3+/-4.7; NS: 144.1+/-18.2; HS: 221.9+/-12.9 mmol 24 h(-1) 1.73 m(-2)), but were not accompanied by significant changes in urinary dopamine excretion. On day 8, clearance studies showed that an acute oral water load of 1500 ml did not alter glomerular filtration rate or renal plasma flow but significantly increased urinary flow rate without affecting dopamine excretion. Assuming that excreted dopamine is not metabolized or reabsorbed during the tubular passage, both the unchanged urinary dopamine output in spite of 14-fold variations in sodium excretion and its independence of an acute water load argue against the hypothesis that dopamine in the tubular lumen acts as a natriuretic and/or diuretic factor in humans. PMID- 10863995 TI - Muscle and motor-skill dysfunction in a K+ channel-deficient mouse are not due to altered muscle excitability or fiber type but depend on the genetic background. AB - The voltage-gated K+ channel Kv3.1 is expressed in skeletal muscle and in GABAergic interneurons in the central nervous system. Hence, the absence of Kv3.1 K+ channels may lead to a phenotype of myogenic or neurogenic origin, or both. Kv3.1-deficient (Kv3.1-/-) 129/Sv mice display altered contractile properties of their skeletal muscles and show poor performance on a rotating rod. In contrast, Kv3.1-/- mice on the (129/Sv x C57BL/6)F1 background display normal muscle properties and perform like wild-type mice. The correlation of poor performance on the rotating rod with altered muscle properties supports the notion that the skeletal muscle dysfunction in Kv3.1-/- 129/Sv mice may be responsible for the impaired motor skills on the rotating rod. Surprisingly, we did not find major differences between wild-type and Kv3.1-/- 129/Sv skeletal muscles in either the resting or action potential, the delayed-rectifier potassium conductance (gK) or the distribution of fast and slow muscle fibers. These findings suggest that the Kv3.1 K+ channel may not play a major role in the intrinsic excitability of skeletal muscle fibers although its absence leads to slower contraction and relaxation and to smaller forces in muscles of 129/Sv Kv3.1-/- mice. PMID- 10863996 TI - Stress proteins of 70 kDa in chronically exercised skeletal muscle. AB - Skeletal muscle fibres of untrained animals experience a stress response following exercise. This study was aimed at investigating whether chronic exercise modulates stress proteins of 70 kDa (HSP70s) in skeletal muscle. In the soleus muscle of Wistar rats, adherence to an incremental programme of treadmill running (IPTR) of 3 months duration up-regulated the levels of the beta-subunit of the mitochondrial F1-ATPase and those of HSP72, GRP75 and GRP78. Neither beta F1-ATPase nor sarcoplasmic reticulum Ca2+-ATPase levels changed with training in the extensor digitorum longus (EDL) muscle. However, HSP70s increased during training. In soleus muscle slices of animals sacrificed 3 days after completing the IPTR, HSP72 and GRP75 were synthesized at lower rates than in sedentary animals while the GRP78 synthesis rate increased. Trained, rested animals also experienced a stress response following acute exercise of lower intensity than that of the actual training sessions. The data suggest that up-regulation of HSP70s by chronic exercise depends upon continued physical activity. Furthermore, the inverse correlation between levels and rates of synthesis of HSP72 during rest periods suggests the operation of a feedback regulatory loop aimed at reestablishing the threshold levels characteristic of unstressed fibres. PMID- 10863997 TI - State- and isoform-dependent interaction of isradipine with the alpha1C L-type calcium channel. AB - We investigated the dihydropyridine (DHP) inhibition of barium current (I(Ba)) through the smooth muscle alpha1Ch and cardiac alpha1Ca splice variants of the L type calcium channel using a whole-cell patch-clamp method. IC50 values for inhibition of current amplitude of the alpha1Cb channel were three to fivefold lower than for the alpha1Ca channel at holding potentials between -80 mV and -30 mV. No difference was found in either the transition of the channels into an inactivated state in the absence or presence of drug, or in the recovery from inactivation under control conditions. However, isradipine slowed the recovery from inactivation of alpha1Ca more effectively than alpha1Cb. To evaluate the interaction of isradipine with the open channel state of both splice variants, interactions with the inactivated state were selectively suppressed by the mutation of three amino acids in the IVS6 segment (Y1485I, M1486F, I1493L) in alpha1CbCh30 and alpha1CaCh30 channels. The extent of this interaction was seen by an acceleration of current decay. This was found to be identical for both splice variants. Our results suggest that the higher DHP selectivity of the alpha1Cb versus the alpha1Ca channel is caused by the structural difference in the binding site and not by different transitions between resting, open and inactivated states. PMID- 10863998 TI - Paucity of CFTR current but modest CFTR immunoreactivity in non-diseased human ventricle. AB - We looked for evidence of expression of the cystic fibrosis transmembrane conductance regulator (CFTR) in non-diseased human ventricle. In rabbit and guinea pig the CFTR current is present in the highest density in subepicardial ventricular myocytes. In the present study, whole-cell patch-clamp was used to determine if a CFTR-like chloride current (I(CFTR,card)) can also be activated in human subepicardial ventricular myocytes. No evidence for I(CFTR,card) was detected in these electrophysiological studies when 10 microM forskolin was applied to 23 different cells from 4 donor hearts. Consistent with our previous results, a swelling-induced chloride current (I(Cl,swell)) could be observed after cell inflation. The enzymatic digestion of human ventricle to release single myocytes may have affected our ability to detect I(CFTR,card). Therefore, we looked for anti-CFTR immunoreactivity in slices of left ventricular free wall. A strong immunoreactivity signal was observed in guinea pig ventricle, a positive control. Background staining levels were seen in dog ventricle, a negative control tissue. Human anti-CFTR immunoreactivity was slightly above background. This low level of anti-CFTR immunoreactivity is consistent both with reports that CFTR mRNA is detectable in human ventricle and our inability to detect a significant I(CFTR,card) current density. PMID- 10863999 TI - Cadmium withdrawal contractures in rat soleus muscle fibres. AB - Transient "Cd2+ withdrawal" contractures, with amplitudes of < or =60% peak tetanic tension, were seen when > or =3 mM Cd2+ was removed, after exposures lasting > or =5 min, from solutions bathing rat soleus fibres at 22 degrees C with Cl- as the major external anion. The minimum free [Cd2+] for withdrawal contractures was reduced twofold when the external anion was SO4(2-). Withdrawal contractures were not seen after removal of 3 mM Co2+, Zn2+ or La3+ and were not observed in rat extensor digitorum longus fibres. The contractures were not due to depolarization (membrane potential, Vm, did not change during Cd2+ removal) or to an influx of external Ca2+ (the transient tension increase was recorded when solutions either lacked Ca2+, or contained 2 mM Co2+). Cd2+ withdrawal contractures were abolished by inactivation of excitation-contraction coupling (ECC) following depolarization in 40 mM K+ for 20 min, and recovered from inactivation at the same time as twitch and tetanic contractions with repriming of ECC. Withdrawal contractures were depressed by agents that depress ECC, i.e. low [Ca2+], 2 mM Co2+, 30 mM Ca2+, 30 mM Mg2+ and 50 microM nifedipine. The results support a hypothesis in which withdrawing Cd2+ from the external solution induces a contracture by activating the voltage sensor for ECC. PMID- 10864000 TI - Enhancement of brush border membrane peptidase activity in rat jejunum induced by starvation. AB - Conflicting results have been obtained in previous studies concerning the adaptation of intestinal blush border membrane enzymes to starvation. This study was designed to clarity the changes in these enzymes under starvation conditions, using a molecular biological approach. Sprague-Dawley rats were starved or given total parenteral nutrition (TPN) for 5 days. Rats allowed free access to food were used as controls. Changes in the activity and expression of jejunal brush border membrane enzymes were compared between three groups. In the starved group, aminopeptidase N and dipeptidyl peptidase IV activity was significantly elevated to 177% and 166%, respectively, of control values. In contrast, sucrase and maltase activity was significantly decreased. The activity of these peptidases also tended to be increased at the renal brush border membrane. Up-regulation of peptidase activity was not evident in the TPN group. Western and Northern blot analysis revealed that the changes in aminopeptidase N activity were attributable to increases in the protein and mRNA level. The activity and expression of brush border membrane peptidases in rat jejunum is up-regulated during starvation, and these changes are considered to be an effect of whole-body malnourishment, rather than an absence of luminal nutrition. PMID- 10864001 TI - Transport of protons and lactate in cultured human fetal retinal pigment epithelial cells. AB - Monolayer cultures of human fetal retinal pigment epithelial (RPE) cells were examined for ultrastructural characteristics and junctional integrity by means of electron microscopy. Intracellular pH (pHi) and cell volume changes were measured using the fluorescent dye BCECF. The EM studies indicate that the RPE cells preserve in vivo morphology before and after loading with BCECF. Monolayer cultures were placed in a perfusion chamber in which the solution facing the retinal cell membrane could be changed rapidly. Removal of Na+ or the addition of amiloride caused intracellular acidifications. pHi recovery from an NH4+-induced acid load was blocked by sodium removal or amiloride addition. These results suggest the presence of a Na+-H+ exchange mechanism in the retinal cell membrane. When Cl- was replaced isotonically by lactate or pyruvate the cells acidified. The intracellular acidifications were saturable, reversibly reduced with the inhibitor probenecid (2 mM), and the lactate-induced acidifications were reversibly inhibited by equimolar concentrations of pyruvate. These results indicate the presence of a H+-lactate cotransport mechanism in the retinal membrane. When Cl- was replaced by lactate the cells not only acidified, they also swelled. The data are compatible with water transport induced by the H+ lactate cotransporter. PMID- 10864002 TI - The involvement of caspases in the CD95(Fas/Apo-1)- but not swelling-induced cellular taurine release from Jurkat T-lymphocytes. AB - Following a delay of 45 min, stimulation of the CD95 (Fas/Apo-1)-receptor in Jurkat T-lymphocytes leads to the release of the osmolyte taurine, an event coinciding with apoptotic cell shrinkage. The present study has been performed to elucidate the cellular mechanisms involved in CD95-induced taurine release as compared to swelling-induced taurine release, and to explore whether taurine modifies apoptotic DNA fragmentation and cell shrinkage. Taurine release stimulated by osmotic cell swelling is insensitive to the tyrosine kinase inhibitor herbimycin A and the caspase inhibitor z-Val-Ala-Asp(OMe) fluoromethylketone (zVAD) but is blunted in the absence of extracellular Ca2+. Conversely, the Ca2+ ionophore ionomycin stimulates taurine release. However, the taurine release following CD95 stimulation is not paralleled by an increase of cytosolic Ca2+ and not inhibited by complexation of extracellular Ca2+. None of herbimycin A, the phosphatase inhibitor vanadate, spingomyelinase or Lck56 deficiency prevent CD95-induced taurine release. In contrast, the caspase inhibitor zVAD, but not the caspase inhibitor Ac-Tyr-Val-Ala-Asp chloromethylketone (YVAD), almost abolishes CD95-induced taurine release. Both caspase inhibitors blunt CD95-induced cell shrinkage and DNA fragmentation, zVAD being more effective than YVAD. Preloading of the cells with 40 mM taurine but not with 40 mM mannitol significantly inhibits CD95-induced DNA fragmentation (by 28%) and apoptotic cell shrinkage (by 25%). In conclusion, CD95-receptor triggering leads to caspase-dependent stimulation of cellular taurine release, which facilitates, but is not sufficient for, the triggering of apoptotic DNA fragmentation and cell shrinkage. PMID- 10864003 TI - Functional reconstitution of ICln in lipid bilayers. AB - Reconstitution of purified ICln in lipid bilayer leads to functional ion channels showing varying rectification. The reconstituted single channels have a conductance of approximately equal to 3 pS and their open probability is sensitive to nucleoside analogues. Mutation of a putative nucleotide binding site identified at the predicted extracellular mouth of the ICln channel protein leads to the reduction of the nucleoside-analogue sensitivity. Reconstituted ICln channels can be permeated both by cations and anions. The relative permeability of cations over anions depends on the presence of calcium. In the presence of calcium reconstituted ICln channels are more permeable to bromide than chloride, and more permeable to potassium than sodium. Similarly in NIH3T3 fibroblasts, the relative permeability of cations over anions of swelling-dependent chloride channels depends on extracellular calcium. Site-directed mutagenesis revealed the calcium-binding site responsible for the shift of the selectivity from cations towards anions of reconstituted ICln channels. Additional indirect structural information has been obtained by mutating a histidine in the predicted pore region of ICln. This histidine seems to have access to the ion-conducting tunnel of the pore. Our experiments show that ICln can act as an ionic channel, which does not exclude additional functions of the protein in regulatory mechanisms of the cell. Since knocking down the ICln protein in fibroblasts and epithelial cells leads to an impaired regulatory volume decrease (RVD) after cytoplasmic swelling and reconstituted ICln channels show several biophysical features of ion channels activated after swelling, ICln is a molecular candidate for these channels. PMID- 10864004 TI - Transferrin receptor activity and localisation in the rat duodenum. AB - It is not known how the efficiency of intestinal iron absorption is regulated. One hypothesis suggests that an interaction between the transferrin receptor (TfR) and the haemochromatosis protein (HFE) regulates the level of iron loading in crypt cells. The hypothesis goes on to suggest that this determines the amount of transport protein, expressed in villus enterocytes, that is involved in iron absorption. Mice with haploinsufficiency for TfR are iron deficient and this is thought to be caused by reduced iron absorption. This suggests that TfR may play a role in the regulation and/or mechanism of iron absorption. We investigated TfR function and distribution by measuring iron uptake from plasma transferrin and by immunohistochemistry. The uptake of transferrin-bound iron (Tf-Fe2) into mucosal cells subsequently separated along the crypt-villus axis was compared to the presence of TfR, determined by immunohistochemistry using frozen and wax sections. Frozen sections showed TfR staining in crypt and villus epithelial cells. In wax sections TfR was only identified in a supranuclear region commencing in enterocytes at the crypt-villus junction and attaining greatest levels at the villus tip. This indicates that the processing of wax tissue exposes a TfR epitope that otherwise remains undetectable when studied in frozen sections. This appearance in paraffin sections was inversely related to the uptake of Tf-Fe2. Supranuclear TfR was not associated with lysosomes, since there was no difference in the uptake of normal Tf-Fe2 and that of the non-digestible cellobiose Tf-Fe2, and Western blot analysis revealed similar amounts of TfR in crypt and villus cells. Also the uptake of Tf-Fe2 increased linearly with time, albeit less in villus than crypt cells, suggesting that maturation of an efflux system in villus cells is not responsible for this difference. We hypothesise that TfR in the supranuclear region of villus enterocytes may play a role in iron absorption. PMID- 10864005 TI - Putative ryanodine receptors in the sarcolemma of ventricular myocytes. AB - The activity of caffeine-activated large conductance channels was recorded in whole-cell, patch-clamped, isolated ventricular myocytes from rabbit heart. The channels were permeable to monovalent and divalent cations and had a unitary monovalent cation conductance of 300-400 pS. Extracellular ruthenium red reduced the unitary conductance of the caffeine-activated channel in a concentration- and voltage-dependent manner. Ryanodine locked the caffeine-activated channels into a subconductance state. Elevating intracellular Ca2+ by photolysis of "caged calcium" increased the number of channel openings. The properties of this caffeine-activated channel were remarkably similar to those of cardiac ryanodine receptors (RyR) and support the novel finding that these channels may also be found on the sarcolemmal membrane. PMID- 10864006 TI - Inhibition of volume-stimulated taurine efflux and tyrosine kinase activity in the skate red blood cell. AB - Phosphorylation of the band 3 anion exchange protein by the tyrosine kinases p72syk and p56lyn is thought to play a role in the pathway that regulates swelling-activated taurine efflux from the skate red blood cell. In this study, the protein tyrosine kinase (PTK) inhibitors piceatannol and tyrphostin A23 both inhibited taurine efflux and the activities of the tyrosine kinases p72syk and p56lyn in the skate erythrocyte. However, the PTK inhibitors genistein and tyrphostin A46 had only small effects on taurine efflux and PTK activities. In general, a strong correlation between the extent of inhibition of taurine efflux and of tyrosine kinase activity was observed. PTK inhibitors showed a similar pattern of inhibition of band 3 phosphorylation, with the greatest inhibition observed in cells treated with piceatannol. The protein kinase C inhibitors staurosporine and bisindolylmaleimide tested alone or in combination with piceatannol had little or no significant effect on swelling-activated taurine efflux. Overall the results support the hypothesis that phosphorylation of the skate band 3 protein by p72syk and p56lyn contributes to the regulation of volume activated taurine efflux in skate red cells, and suggest that protein kinase C may not be involved in this regulation. PMID- 10864007 TI - Intracellular pH in isolated rat renal papillary thin limbs of Henle's loop. AB - Intracellular pH (pHi) was measured in isolated, nonperfused and perfused rat papillary thin limbs of Henle's loops in N-2-hydroxyethylpiperazine-N'-2 ethansulfonic acid (HEPES)- or HEPES/bicarbonate-buffered medium at pH 7.4 using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Resting pHi was about 6.7 in descending thin limbs (DTL) and about 6.9 in ascending thin limbs (ATL), even with a medium pH of 7.4. These values appeared to reflect the acid pH of the blood in the neighboring vasa recta found in vivo. The resting pHi did not differ whether or not the medium contained bicarbonate although the total buffering capacity of the tubule cells was increased in the presence of bicarbonate. In nonperfused DTL and ATL, pHi was further acidified following an NH4Cl pulse. The rate of recovery of pHi from this level to the resting pHi was reduced by Na+ removal from the bath in both DTL and ATL and by the addition of ethylisopropylamiloride (EIPA) to the bath in the presence of Na+ in DTL. The rate of recovery was not affected by Cl- removal from the bath or K+ (75 mM) or 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) addition to the bath in either DTL or ATL. These results suggest that the common, amiloride-sensitive, basolateral Na+/H+ exchanger plays a role in the regulation of pHi in rat papillary DTL but that a different basolateral Na+/H+ exchanger or a luminal Na+/H+ exchanger is important in rat papillary ATL. PMID- 10864008 TI - Evidence for tetrodotoxin-sensitive sodium currents in primary cultured myocytes from human, pig and rabbit arteries. AB - Primary cultured human coronary myocytes express a tetrodotoxin-sensitive sodium current (I(Na)). Here, we have investigated whether I(Na) is expressed in vascular smooth muscles cells (VSMCs) isolated from other large arteries, and other mammals. VSMCs were enzymatically dissociated, kept in primary culture, and macroscopic I(Na) was recorded using the whole-cell patch-clamp technique. We found that I(Na) is expressed in VSMCs grown from human aortic (90%; n=50) and pulmonary (44%; n=19) arteries, and in the human aortic myocyte cell line HAVSMC (94%; n=27). I(Na) was also detected in pig coronary (60%; n=33), and rabbit aortic (47%; n=15), but not in rat aortic VSMCs (n=20). These different I(Na) had similar voltage thresholds for activation (approximately equal to -50 mV), and were highly sensitive to extracellularly applied tetrodotoxin. We conclude that I(Na) is expressed in VSMCs grown from various types of large arteries in humans, pig and rabbit. PMID- 10864009 TI - Inhibition of the human intermediate-conductance, Ca2+-activated K+ channel by intracellular acidification. AB - The effect of changes in pH on the gating properties of the cloned human intermediate-conductance, Ca2+-activated K+ channel (hIK) was studied using the patch-clamp technique. Multi-channel inside-out recordings of patches from HEK 293 cells stably expressing hIK channels revealed that the channel activity is modulated by changes in intracellular pH (pHi). Changes in extracellular pH (pHo) in the range from pH 6.0 to 8.2 did not affect the hIK whole-cell current. Intracellular acidification gradually decreased the activity of the hIK channel, approaching zero activity at pHi 6.0. Decreasing pHi altered neither the conductance nor the inward rectification of hIK channels. The proton-induced inhibition of the multi-channel hIK patch current could not be counteracted by increasing the cytosolic Ca2+ concentration to 30 microM. The molecular sensory mechanism underlying the proton-induced modulation of hIK gating is at present unknown. PMID- 10864010 TI - Distribution of bile acid absorption and bile acid transporter gene message in the hamster ileum. AB - The apical, Na-dependent, ileal bile acid transporter (IBAT) is critical for the reabsorption of bile acids in the ileum. Bile acid transport capacities as well as the distribution of bile acid transporter messenger ribonucleic acid (mRNA) and transporter protein were studied along the axis of the ileum. Na-dependent and Na-independent taurocholate uptake was measured in the hamster ileum using an everted-sleeve technique. The distribution of IBAT mRNA and protein were mapped by in-situ hybridization, immunohistochemistry, and Western blotting. Na dependent and Na-independent bile acid uptake rates were highest 1-4 cm before the ileocecal valve (maxima 780 and 120 pmol/mm2 per min, respectively) and decreased proximally and distally. Na-independent absorption was increased in the last 6 cm of the ileum. IBAT mRNA and protein expression were linked closely to the distribution of uptake capacity. IBAT mRNA was more abundant near the crypt villus junction whereas the protein was expressed evenly along the villus axis. We conclude that Na-dependent and Na-independent bile acid absorption capacities both have distinct distribution curves in the hamster ileum. All ileocytes on villi in the high-uptake area of the ileum express IBAT mRNA and protein. PMID- 10864011 TI - Temperature effect on the force/velocity relationship of the fresh and fatigued human adductor pollicis muscle. AB - The purpose of the present study was to investigate the effect of muscle temperature on the force/velocity relationship of electrically activated human adductor pollicis muscle. Following immersion of the lower arm for 20 min in water baths of four different temperatures, the calculated muscle temperatures were 37.1, 31.4, 25.6 and 22.2 degrees C. At 22.2 degrees C maximal isometric force was reduced to 79.3+/-2.9% of the force obtained at 37.1 degrees C. Q10 values for the maximal rates of force development and relaxation, and relaxation times, were about 2.0 between 37.1 and 25.6 degrees C and increased to about 3.5 below 25.6 degrees C. The Q10 values of the maximal shortening velocity and the velocity for maximal power production were similar to those of the isometric speed parameters. The Q10 for maximal power production increased from 2.0 above 31.4 degrees C to 6.9 between 25.6 and 22.2 degrees C. Following repetitive isometric contractions maximal power production was reduced to 60.0+/-1.7 and 90.5+/-1.0% at 37.1 and 22.2 degrees C respectively. Fatigue decreased with cooling of the muscle over the entire (37.1-22.2 degrees C) temperature range. PMID- 10864012 TI - Transport, catabolism and release of histamine in the ruminal epithelium of sheep. AB - Whereas intraruminal histamine does not affect healthy ruminants, histaminosis is apparent during ruminal acidosis. We therefore investigated the factors that, under physiological circumstances, prevent intoxication by intraruminal histamine and the disturbances occurring during acidotic or hypoxic epithelial damage. After mucosal (m) or serosal (s) application of 80 microM histamine, its flux across the isolated epithelia of the sheep rumen was determined radioactively (hist-rad flux) in Ussing chambers. The non-catabolized component of the hist-rad fluxes was determined by high-pressure liquid chromatography (HPLC) (histamine flux). The difference between hist-rad and histamine fluxes indicated efficient intraepithelial catabolism of histamine at pH 7.4 (m-s direction, 98.7%; s-m direction, 93.3%). Both 0.1 mM 2,4-dinitrophenol (DNP) and mucosal acidification to pH 5.1 increased hist-rad fluxes and decreased catabolic efficiency. pH dependent secretion of histamine was indicated by differences between m-s and s-m fluxes of histamine and/or hist-rad. Epithelial permeability to hist-rad and mannitol was similar and their fluxes correlated partly. Epithelial release of endogenous histamine was 1.5 pmol x cm(-2) x h(-1) and was not increased by the mast cell stimulator, compound 48/80 (10 ng x ml(-1)). We conclude that histamine absorption across the intact epithelium is efficiently restricted by a low permeability to histamine in combination with catabolic and secretory processes. Especially increases in paracellular permeability and/or inhibition of catabolism enhance histamine absorption. PMID- 10864013 TI - High-resistance MDCK-C7 monolayers used for measuring invasive potency of tumour cells. AB - We describe an electrophysiological method for evaluating the intrinsic invasive potency of tumour cells using renal cells as an in vitro assay system. A high resistance clone of Madin-Darby canine kidney cells (MDCK-C7) was grown to confluency in a filter cup. Transepithelial electrical resistance across the MDCK C7 monolayer was measured in a commercially available electrode chamber. After a transepithelial electrical resistance of about 4,000 omega cm2 had been reached, human melanoma or pancreatic carcinoma cells were co-cultivated with the MDCK-C7 monolayer. Both carcinoma cell lines induced resistance breakdown measured after 24 h or later depending on seeding density and cell type. Seeding carcinoma cells on the basolateral surface of MDCK-C7 cells caused a similar decrease in transepithelial resistance of the MDCK-C7 monolayer. Resistance breakdown indicates opening of tight junctions prior to tumour cell invasion. In conclusion, the high-resistance MDCK-C7 cell clone could serve as a valuable biological assay system to determine electrically the metastatic potency of tumour cells in vitro. PMID- 10864014 TI - NBD-TMA: a novel fluorescent substrate of the peritubular organic cation transporter of renal proximal tubules. AB - Traditionally, the measurement of transport activity has employed radiolabeled compounds. The resulting experimental procedures do not measure transport in real time and are limited in temporal and spatial resolution. The use of epifluorescence microscopy provides the ability to measure transport activity in real time with high temporal and spatial resolution. Using epifluorescence microscopy we characterized the transport of the fluorescent organic cation, [2 (4-nitro-2,1,3-benzoxadiazol-7-yl)aminoethyl]trimethylammonium (NBD-TMA+, MW 266). NBD-TMA+ has structural characteristics common to other secreted organic cations and is fluorescent (lambda(ex)=458 nm; lambda(em)=530 nm). The excitation and emission spectra are insensitive to changes in [Cl-] and minimally sensitive to pH in the physiologically relevant range (pH 5.0-7.4). A microscope equipped with a photon-detection system was used to measure accumulation of NBD-TMA+ by isolated rabbit renal proximal tubules. Accumulation of NBD-TMA+ by proximal tubules was time dependent and saturable (Michaelis-Menten constant Km 12 microM). Proximal tubule accumulation of NBD-TMA+ was inhibited by the organic cations tetraethylammonium (TEA+) (apparent inhibitory constant K(app)TEA 134 microM), cimetidine, and N1-methylnicotinamide (NMN). Our experimental results provide strong evidence that NBD-TMA+ is transported by one or more of the basolateral organic cation transporters involved in the renal secretion of this chemical class of compound. This fluorescent substrate provides a sensitive means of investigating organic cation transport. PMID- 10864015 TI - A Kampo medicine "Juzen-taiho-to"--prevention of malignant progression and metastasis of tumor cells and the mechanism of action. AB - Juzen-taiho-to is a Kampo (Japanese and Chinese traditional) medicine, and is a nourishing agent, a so-called "Hozai" (in Japanese), that is used for improving disturbances and imbalances in the homeostatic condition of the body. This drug is administered to patients in various weakened conditions, including post surgery patients and patients with chronic illnesses, where it can alleviate general symptoms such as extreme fatigue, pale complexion, loss of appetite, dry or scaly skin, night sweating, and dryness of the mouth. Currently, Juzen-taiho to is often administered to cancer patients, and has been shown to possess various biological activities, such as enhancement of phagocytosis, cytokine induction, antibody production, induction of the mitogenic activity of spleen cells, anti-tumor effects when combined with surgical excision, anti-tumor effects with or without other drugs, and protection against the deleterious effects of anti-cancer drugs as well as radiation-induced immunosuppression and bone marrow toxicity. This article focuses on the antitumor and antimetastatic properties of Kampo formulations and describes the effect of Juzen-taiho-to and related formulations on tumor development, progression and metastasis in vivo. We also discuss the mechanism of the inhibitory action and the importance of the formulation and the constituent drugs in determining the efficacy. PMID- 10864016 TI - Inhibition of iron ion-induced oxidative damage of erythrocyte membranes and low density lipoprotein by a Maillard product, 4-hydroxy-2(or 5)-ethyl-5(or 2)-methyl 3(2H)-furanone (HEMF). AB - 4-Hydroxy-2(or 5)-ethyl-5(or 2)-methyl-3(2H)-furanone (HEMF) is representative of the Maillard reaction-derived reductones found in many foodstuffs. Influence of HEMF on iron ion-induced oxidative modification of human erythrocyte membranes and low density lipoprotein (LDL) under aerobic conditions was investigated. When human erythrocytes were incubated at 37 degrees C for 24 h with HEMF alone, levels of thiobarbituric acid-reactive substances (TBARS) and non-ionic detergent C12E8-insoluble membrane protein aggregates in the membranes were unchanged. Levels of TBARS and protein aggregates in the membranes increased on treatment of the cells with Fe(III) ion at 37 degrees C for 3 h were lowered in the presence of HEMF. Western blot analysis indicated that aggregates of band 3 protein induced by Fe (III) ion were decreased in the presence of HEMF, and the level of TBARS in LDL increased on treatment with Fe(II) or Fe(II) ion at 37 degrees C for 24 h was also lowered in the presence of HEMF. The results indicate that HEMF protected the iron ion-induced oxidative modification of human erythrocyte membranes and LDL. PMID- 10864017 TI - Inhibitory effects of green tea and grape juice on the phenol sulfotransferase activity of mouse intestines and human colon carcinoma cell line, Caco-2. AB - Tea and fruit juices are beverages consumed daily all over the world. The present study reports the inhibitory effects of these beverages on the activity of mammalian intestinal phenol sulfotransferases (P-STs). Green tea strongly inhibited the E. coli-expressed mouse intestinal P-ST activity in vitro. (-) Epigallocatechin gallate (EGCG) was found to be the most potent inhibitor among the catechins tested (IC50=0.93 microM). (-)EGCG also inhibited the P-ST activity of the human colon carcinoma cell line, Caco-2. Kinetic analysis showed that the inhibition was competitive. Among fruit juices examined (apple, grape, grapefruit and orange), grape juice exhibited the most potent inhibitory action on the P-ST activity of mouse intestines and human colon carcinoma cells. The inhibitory activity of grape juice was located mainly in the skin and seeds. Flavonols, such as quercetin and kaempferol, inhibited the P-ST activity at low concentrations. These observations suggest the possible inhibition of P-ST activity in human intestines by green tea or grape juice. PMID- 10864018 TI - Study of the beta1 adrenergic receptor expression in human tissues: immunological approach. AB - Questions exist regarding tissue distribution of the beta1 adrenergic receptor (beta1-AR). The aim of this study was to investigate relative distribution patterns of the beta1-AR at the protein level in a variety of human tissues by Western blot analysis. The specificity of anti-peptide antibodies was confirmed both by Western blot with recombinant beta1-AR expressed as a membrane protein in E. coli and by immunoprecipitation of membranes from Sf9 cells infected with baculovirus to express the human recombinant beta1-AR. beta1-AR was found in all tissues examined. The relative amount of protein varied significantly between the tissues, from highest in lung and testis to very low in liver. beta1-ARs were rather abundant in heart, kidney, placenta, spleen and thyroid. These results reveal unique distribution of beta1-AR protein that suggests its tissue specific role. Moreover, our data demonstrate a high sensitivity of immunological detection that allows direct comparison of beta1-AR subtype expression and could be used for receptor study in biopsies available in limited amounts, such as human heart biopsy. PMID- 10864019 TI - Immunohistochemical and chemical changes of beta-citryl-L-glutamate in the differentiation of bovine lens epithelial cells into lens fiber cells. AB - Beta-citryl-L-glutamate (beta-CG) concentration was determined by HPLC during the differentiation of bovine lens epithelial cells into lens fiber cells in culture. beta-CG increased from 1 to 4 weeks of culture and then decreased slightly, while alpha-crystallin, a marker of lens cell differentiation, increased rapidly 4 weeks after the culture and continued to increase gradually until week 11. In addition, the localization of beta-CG was immunohistochemically examined using anti-beta-CG antibody. Cells around lentoid bodies were stained with anti-beta-CG antibody, whereas cells in the bodies were stained strongly with anti-gamma crystallin antibody. These findings suggest that beta-CG accumulated immediately before the differentiation of the bovine lens epithelial cells into lens fiber cells and may play a role in regulating the differentiation of lens cells. PMID- 10864020 TI - Stable prostaglandin I1 analog SM-10906 modulates productions of tumor necrosis factor-alpha, interleukin-1 and interleukin-6 in mouse macrophages. AB - The present study investigated the effects of stable agonist for prostaglandin (PG) I2 receptor with PGI1 skeleton, SM-10906, on pro-inflammatory cytokine production by mouse peritoneal macrophages (PEMs) in comparison with PGE1 and PGI2. In mouse PEMs, SM-10906 and PGE1 slightly enhanced interleukin (IL)-6 secretion, but had no effects on tumor necrosis factor-alpha (TNF-alpha) or IL-1 production. SM-10906 concentration-dependently inhibited TNF-alpha, IL-1 and IL-6 releases from lipopolysaccharide-activated mouse PEMs, as with PGE1, PGI2 and cAMP analog. Additionally, SM-10906, PGE1 and PGI2 caused concentration-dependent accumulation of cAMP contents in mouse PEMs. It is concluded that PGI1 analog SM 10906 exerts anti-inflammatory effects on stimulated mouse PEMs by increasing in cAMP levels, as with E-series of PG. PMID- 10864021 TI - The effects of a novel cyclohexane dicarboximide derivative, ST-6, on hypoxia/reoxygenation injury in perfused rat heart. AB - The present study was undertaken to test if some cyclohexane dicarboximide derivatives may have a cardioprotective effect against hypoxia/reoxygenation injury. Isolated rat hearts were subjected to 20-min of hypoxia followed by 45 min reoxygenation, and their recovery of post-hypoxic cardiac contractile function was examined. Treatment with agents was carried out from 3 min after the onset of hypoxia to the end of hypoxia (17 min during hypoxia). Among the 17 compounds, 2-[4-[4-(4-chlorophenyl)-4-hydroxy-1-piperidinyl]butyl]hexahydro-1H-i soindol-1,3(2H)-dione (ST-6) showed a significant enhancement of post-hypoxic contractile force. This was associated with attenuation of the releases of creatine kinase and purine nucleosides and bases from the perfused heart. Hypoxia induced increase in myocardial sodium and decrease in potassium ion content was suppressed by ST-6 treatment. The results suggest that ST-6 is capable of protecting the heart against hypoxia/reoxygenation injury possibly through a mechanism by which sodium overload during hypoxia is suppressed. PMID- 10864022 TI - A study on the nitric oxide production-suppressing activity of sanguisorbae radix components. AB - The active components of an aqueous extract of Sanguisorbae Radix, which possesses nitric oxide (NO) production-suppressing activity, were determined using macrophages that were activated by the addition of lipopolysaccharide. Significant inhibitory activity against the formation of both NO and inducible NO synthase, and NADPH-diaphorase activity, which is involved in NO generation, was shown by Sanguisorbae Radix fractions T-B and T-C. On further fractionation, the subfractions of T-B and T-C all showed high anti-NO activity. Sanguiin H-6, sanguiin H-11, 1,2,3,4,6-penta-O-galloyl-beta-D-glucose, eugeniin and polymeric proanthocyanidin were isolated from TB-3 and TC-4, and all were identified as exhibiting strong anti-NO activity. We have confirmed that sanguiin H-6 is the most active component of Sanguisorbae Radix with respect to the suppression of NO production. It is suggested that tannin makes a prominent contribution to the biological activity of Sanguisorbae Radix. PMID- 10864023 TI - Nematocidal activity of quassinoids against a species of Diplogastridae. AB - The nematocidal activity of 38 quassinoids, C19 or C20 compounds isolated from Simaroubaceae, was measured using a species of Diplogastridae (Nematoda) to develop lead parasiticides. Of the various quassinoids tested, samaderines B and E displayed the most potent nematocidal activity with a minimum lethal concentration (MLC) of 2.0 x 10(-5) M. The nematocidal activities of samaderines B and E were 15-fold greater than that of albendazole (3.0 x 10(-4) M), 10-fold greater than that of thiabendazole (2.0 x 10(-4) M) and 7.5-fold greater than that of avermectin (1.5 x 10(-4) M). Thus, samaderines B and E may eventually be used as lead parasiticides. In light of the relationship between the structures of quassinoids and their nematocidal activities, those with potent nematocidal activity may require the elements mentioned. These results should help to further our understanding of nematocidal activity. PMID- 10864024 TI - Intraspecific variation in Cannabis sativa L. based on intergenic spacer region of chloroplast DNA. AB - We analyzed the nucleotide sequences of the non-coding region of chloroplast DNA: the intergenic spacer between trnL (UAA) 3'exon and trnF (GAA). Two kinds of sequence, "type-1" and "type-2," were detected in 33 populations of Cannabis sativa. The length of the "type-1" fragment was 354 bp. In contrast, the "type-2" fragment from 3 populations was 353 bp long, with only one base deletion compared to "type-1." The fragment length from Humulus lupulus was 353 bp with a 1-bp deletion, and ten 1-bp substitutions compared to the sequences from C. sativa "type-1." Furthermore, we could clearly identify differences between C. sativa and H. lupulus using single-strand conformation polymorphism of PCR products (PCR SSCP) analysis. PMID- 10864025 TI - Hepatoprotective aliphatic glycosides from three Goodyera species. AB - Hepatoprotective aliphatic glycosides 3-(S)-3-beta-D-glucopyranosyloxybutanolide (1) and its congener, 3-(S)-3-beta-D-glucopyranosyloxy-4-hydroxybutanoic acid (2) were isolated as major constituents from the whole plants of three Goodyera species, G. schlechtendaliana Reichb. fil., G. matsumurana Schltr. and G. discolor Ker-Gawl. The structures of 1 and 2 have been determined by NMR, MS spectroscopic and chemical means. Compound 1 was converted into its methyl ester form (5) during the purification step, when the lactone ring was cleaved by catalysis of silica gel with the CHCl3-MeOH-H2O as a solvent. On the other hand, 1 was obtained in a high yield by the same purification procedure without MeOH. Based on this fact, a simple and economic method for the purification of 1 was confirmed. Compounds 1 and 2 were found to have a hepatoprotective effect on liver injury induced by carbon tetrachloride in primary cultured rat hepatocytes. PMID- 10864026 TI - Pycnogenol protects vascular endothelial cells from beta-amyloid-induced injury. AB - The neuropathological hallmarks of Alzheimer's disease (AD) are senile plaques, cerebrovascular beta-amyloidosis, neurofibrillary tangles, and selective neuronal loss. Beta-amyloid (Abeta) has been shown to cause vascular damage mediated by generation of reactive oxygen species and this damage is considered an early event in the development of AD. In this study, we determined the effect of pyenogenol, a potent antioxidant phytochemical, on Abeta-induced cellular injury. Pulmonary artery endothelial cells (PAEC) were exposed to Abeta for 24 h. Cell injury was assessed by measuring cell viability with methylthiazol tetrazolium (MTT) assay, and by determining the release of intracellular lactate dehydrogenase (LDH). Lipid peroxidation products of PAEC were determined by measuring thiobarbituric acid-reactive substances (TBARS). Exposure of PAEC to Abeta resulted in a decrease in cell viability, an increase of LDH release indicating membrane damage, and an elevated level of TBARS. Preincubation of PAEC with pycnogenol significantly minimized these changes. This study demonstrated that pycnogenol can protect vascular endothelial cells from Abeta-induced injury. The data suggest that pycnogenol may be useful for the prevention and/or treatment of vascular or neurodegenerative diseases associated with Abeta toxicity. PMID- 10864027 TI - Electrophysiological studies on the evaluation of absorption enhancers in Caco-2 cells using a microelectrode technique. AB - We developed a microelectrode technique to characterize the electrophysiological properties in Caco-2 cells, and used it to determine the mechanisms of absorption enhancers. The action of absorption enhancers on the apical membrane of Caco-2 cells was estimated by measuring the apical membrane potential (Vm) with the microelectrode. The Vm value of Caco-2 cells in Hanks' balanced salt solution containing 0.5 mM K+ was 18.9+/-0.8 mV (n=217), and the apical membrane resistance was 49.4+/-1.1 MOhms (n=160). In the electrophysiological study with absorption enhancers, laurylmaltoside (LM) markedly decreased the Vm value, while sodium glycocholate (NaGC) moderately reduced this value, and EDTA did not affect the value. These findings might be associated with their action sites, plasma membrane or tight junction in Caco-2 cell monolayers. In influx and transport studies with these absorption enhancers, LM enhanced the influx of furosemide, which is transported via both the transcellular and paracellular routes into Caco 2 cells, and enhanced its transport to the basolateral side of Caco-2 monolayers more than that of 5(6)-carboxyfluorescein (CF), a paracellular marker. EDTA did not increase the influx of furosemide, and enhanced the transport of furosemide and CF across Caco-2 cell monolayers to the same extent. In contrast, NaGC only slightly increased the influx of furosemide and did not enhance the transport of either furosemide or CF across the Caco-2 monolayers in this study. These findings were well correlated with the effects of these absorption enhancers on the electrophysiological parameters. Therefore, the microelectrode technique might be useful for evaluating the action of absorption enhancers in the plasma membrane at an intact cell level. PMID- 10864028 TI - Uptake of fractionated heparin by two types of scavenger receptors in isolated rat Kupffer cells. AB - The uptake of fractionated heparin was examined in the absence and presence of anionic proteins such as acetylated low density lipoprotein (Ac-LDL) and maleylated bovine serum albumin (Mal-BSA) to characterize the scavenger receptors involved in the uptake of fractionated heparin in isolated rat Kupffer cells. The uptake of fractionated heparin was completely inhibited by Ac-LDL and dextran sulfate, but only partially by Mal-BSA. Kinetic analysis revealed that the binding capacity (Bmax) of the Mal-BSA-insensitive receptor was significantly larger than that of the Mal-BSA-sensitive one, though their dissociation constants (Kd) were not significantly different. The apparent internalization rate constant (kint,app) was significantly larger for the Mal-BSA-sensitive receptor than for the Mal-BSA-insensitive one. Thus, the scavenger receptors involved in the uptake of fractionated heparin in Kupffer cells can be classified into two types, in terms of sensitivity to Mal-BSA. Mal-BSA-sensitive receptors have been characterized in macrophages and classified as class A. The Mal-BSA sensitive one found in Kupffer cells in this study may belong to class A, while the Mal-BSA-insensitive one has been little characterized elsewhere. PMID- 10864029 TI - Induction of apoptosis in the human promyelocytic leukemia cell line HL60 by falconensone A and its derivatives, new polyenes. AB - Falconensones A and B are a new type of yellow pigment with structural similarity to retinoic acid isolated from the mycelial extract of ascomycetous fungi, Emericella falconensis or Emericella fruticulosa. In the present study we show that falconensone A alone induced apoptosis of HL60 human leukemia cells, while falconensone B, the 4'-nor-methyl derivative of falconensone A, had much lower activity. The synthetic derivatives of falconensone A, falconensone A p bromophenylhydrazone and falconensone A dioxime, were more potent than natural falconensone A and B as far as the induction of apoptosis was concerned. The induction of apoptosis by the falconensones correlated with their inhibition of cell growth. In addition, falconensones A and B, and falconensone A dioxime, increased the generation of intracellular reactive oxygen species, while falconensone A p-bromophenylhydrazone was inactive. These results suggest that falconensone A, falconensone A p-bromophenylhydrazone and falconensone A dioxime are potential new apoptosis-inducing agents. The enhanced generation of reactive oxygen species in cells may be involved in apoptosis induced by falconensone A and falconensone A dioxime, but not by falconensone A p-bromophenylhydrazone. It is also suggested that the methyl residue at the 4' position of the falconensone A cyclopentenone ring may be essential for the induction of apoptosis. Based on these results, falconensone A and its derivatives may be clinically useful in the treatment of some leukemias. PMID- 10864030 TI - Expression of inhibin betaA, betaB and follistatin mRNAs in the carbon tetrachloride induced rat liver regeneration model. AB - Follistatin (FS, an activin-binding protein) and activin A (homodimer of inhibin betaA chain) promote and inhibit cell proliferation in rat liver, respectively. The roles of activin AB (heterodimer of inhibin betaA and betaB) and activin B (homodimer of inhibin betaB) in rat liver have not been elucidated yet. In this study, we examined, by reverse-transcriptase polymerase chain reaction (RT-PCR) analysis, whether the levels of FS, inhibin betaA and betaB mRNAs change in the carbon tetrachloride induced rat liver regeneration model. The analysis was made in an hour-by-hour manner during the early stage of liver injury. There are 2 types of FS mRNA, FS-288 and FS-315, and the levels of both had begun to increase at 3 h, were maximal at 6 h, remained constant up to 12 h, and thereafter gradually decreased. The inhibin betaA mRNA had started to decline at 3 h, reached its lowest level at 6 h, partly returned at 12 h, and remained constant up to 48 h. The inhibin betaB mRNA level had begun to increase at 1 h, was maximal at 3 h, remained constant up to 24 h, and returned to the original level at 48 h. These results indicate that FS and activin A may act reciprocally in liver regeneration, and also suggest that activin AB and B may play roles in liver regeneration that differ from that of activin A. PMID- 10864031 TI - Occurrence of a novel cannabimimetic molecule 2-sciadonoylglycerol (2-eicosa 5',11',14'-trienoylglycerol) in the umbrella pine Sciadopitys verticillata seeds. AB - The umbrella pine Sciadopitys verticillata seeds were found to contain a substantial amount (16.7 nmol/g) of sciadonic acid (all-cis-5,11,14 eicosatrienoic acid)-containing 2-monoacylglycerol, i.e., 2-sciadonoylglycerol (2 eicosa-5',11',14'-trienoylglycerol). Because the structure of 2 sciadonoylglycerol closely resembles that of 2-arachidonoylglycerol, the endogenous natural ligand for the cannabinoid receptor, we examined whether or not 2-sciadonoylglycerol exhibits cannabimimetic activity using NG108-15 neuroblastomaxglioma hybrid cells which express the cannabinoid CB1 receptor. We found that 2-sciadonoylglycerol induces rapid transient elevation of intracellular free Ca2+ concentration in NG108-15 cells through a cannabinoid CBI receptor-dependent mechanism similar to the case of 2-arachidonoylglycerol, yet the activity of 2-sciadonoylglycerol was apparently lower than that of 2 arachidonoylglycerol. The activity of 2-sciadonoylglycerol was detectable from 3 10 nM, reaching a maximum at around 10 microM. To our knowledge, this is the first report showing the occurrence of a cannabimimetic monoacylglycerol in higher plants. PMID- 10864032 TI - Effects of 9 Kampo medicines clinically used in hypertension on hemodynamic changes induced by theophylline in rats. AB - We examined the effects of 9 kinds of Kampo medicines, which are clinically used for the treatment of hypertension, on anesthetized rats with increases in arterial blood pressure, heart rate and peripheral blood flow induced by theophylline (5 mg/kg, i.v.) that were partially or completely mediated by endogenous catecholamines. Each Kampo medicine (1 g/kg) was intraduodenaly administered. Shinbu-to caused a severe disturbance of the arterial blood pressure. Saiko-ka-ryukotsu-borei-to, Oren-gedoku-to, San'o-shashin-to and Dai jyoki-to had hypotensive effects, while Hachimi-jio-gan, Gosha-jinki-gan, Dai saiko-to and Choto-san did not have such an effect. Moreover, Saiko-ka-ryukotsu borei-to attenuated the heart rate. In Oren-gedoku-to, San'o-shashin-to and Dai jyoki-to, a reduction in peripheral blood flow was observed. These results suggest that Saiko-ka-ryukotsuborei-to, Oren-gedoku-to, San'o-shashin-to and Dai jyoki-to are ameliorative to the hypertension in sympathetic system dominance and Shinbu-to is occasionally dangerous to it. PMID- 10864033 TI - Antioxidant effects of calcium antagonists in rat brain homogenates. AB - We studied the antioxidant activities of calcium antagonists against autoxidation in rat brain homogenates. The homogenates were incubated for 30 min at 37 degrees C with or without a calcium antagonist and subsequently assayed for lipid peroxide content. Percent inhibition of the lipid peroxidation was used as an index of the antioxidant effect. Dihydropyridine calcium antagonists exhibited concentration-dependent (3-300 micromol/l) inhibitory effects against lipid peroxidation. The relative order of antioxidant potency and associated IC50 values (micromol/l) of the calcium antagonists for inhibition of the lipid peroxidation were as follows: nifedipine (51.5)>barnidipine (58.6)>benidipine (71.2)>nicardipine (129.3)>amlodipine (135.5)>nilvadipine (167.3)>nitrendipine (252.1)>> diltiazem (>300)=verapamil (>300). These results suggest that some dihydropyridine calcium antagonists show antioxidant properties. The antioxidant effects of the calcium antagonists may contribute to their pharmacological actions. PMID- 10864034 TI - Effects of MET-88, a gamma-butyrobetaine hydroxylase inhibitor, on tissue carnitine and lipid levels in rats. AB - MET-88, 3-(2,2,2-trimethylhydrazinium) propionate, suppresses carnitine synthesis by inhibiting (gamma-butyrobetaine hydroxylase. The purpose of this study was to clarify the effects of suppression of carnitine synthesis on carnitine and lipid contents in tissues. MET-88 (50, 100, 200 or 400 mg/kg/d) was administered orally to male SD rats for 10, 30 or 60 d. Total carnitine and lipid (triglycerides, non esterified fatty acids) contents were measured in heart and liver. In both tissues, treatment with MET-88 dose-dependently decreased total carnitine levels, and the reduction reached the plateau state after 30 d at each dose. MET-88 had no effect on lipid content in the heart, but increased the lipid content in the liver at the highest doses. Treatment with MET-88 at 400 mg/kg for 60 d resulted in no pathologic findings in the histological study, and also had no effect on parameters of liver function such as glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase as judged from the results of blood biochemical analysis. We concluded that long-term treatment with MET-88 decreased the carnitine content to a constant level in both heart and liver, but had no effect on lipid contents in the heart, although it affected lipid metabolism in the liver. PMID- 10864035 TI - Methotrexate concentration in cerebrospinal fluid of the space created by tumor removal. AB - This paper investigates the post-surgical pharmacokinetics of methotrexate (MTX) in the plasma, the cerebrospinal fluids (CSF), and the spaces created by tumor removal (STR) in patients with glioblastoma, during hyperosmotic disruption of the blood brain barrier (HODBBB) and intra-arterial chemotherapy with MTX. Eight Japanese patients with glioblastoma, three with open STRs and five with closed STRs, received a total of thirteen courses of HODBBB and intra-arterial combination chemotherapy with MTX. The patients were initially administered mannitol, then the anticancer drugs were infused into the carotid artery. Samples of blood and CSF from the STRs were obtained. MTX concentrations were measured by fluorescence polarization immunoassay and the pharmacokinetic parameters of MTX in plasma and CSF were estimated. The plasma concentrations of MTX peaked at the end of drug infusion, and then decayed bi-exponentially during the remainder of the treatment period. The CSF concentration of MTX in the STR peaked 2 h after drug administration, then mono-exponentially decreased. The area under the concentration-time curve (AUC) for plasma and CSF MTX concentrations increased in parallel with the MTX dose. In patients with open STRs, the mean AUC of MTX in CSF was 4.44% of that found in plasma, while in patients with closed STRs, the mean was 61.2% of that found in plasma. In the latter group, the MTX administered using HODBBB and intra-arterial chemotherapy was maintained in the STRs for long periods. PMID- 10864036 TI - Monosialoganglioside containing cationic liposomes with a cationic cholesterol derivative promote the efficiency of gene transfection in mammalian culture cells. AB - We have studied the effects of monosialoganglioside (GM1)-containing cationic liposomes with a cationic cholesterol on the liposome-mediated gene transfection into mammalian culture cells. The results showed that both cationic liposomes with either a cationic cholesterol derivative of a hydrophobic amino head group (I) and a hydrophilic amino head group (II) promoted the transfection of luciferase plasmids (pGL3) into HeLa and CHO-K1 cells more than the control cationic liposomes without GM1. In addition, we found that cationic liposomes with a cationic cholesterol derivative (II) were about ten times as effective as that by commercially available cationic liposome Lipofectin. Confocal fluorescence microscopy showed that the liposome/DNA complex was transferred more efficiently into the target cells by the GM1-containing liposomes than by the liposomes without GM1. In proportion to the above results, free antisense DNAs were also more efficiently transferred into the nucleus of the target cells by the GM1-containing liposomes. When there was 100 mM galactose in the transfection medium, the luciferase activity by the GM1-containing liposomes was reduced to the level of the control liposomes. The results suggest that GM1-containing cationic liposomes with a cationic cholesterol derivative of a hydrophobic amino head group or a hydrophilic amino head group should significantly increase the transfection efficiency of plasmid DNAs and antisense DNAs by galactose receptor mediated endocytosis. This means that the GM1-containing liposomes described here should be very promising for gene transfection in vitro. PMID- 10864037 TI - Microbial contamination of in-use lubricants for non-touch urethral catheters in intermittent self-catheterization. AB - There are no reports on microbial contamination of repeatedly used lubricant (84 87% glycerin containing 0.02% benzalkonium chloride) for non-touch urethral catheters in intermittent self-catheterization. In this work, we evaluated microbial contamination of in-use lubricant and its prevention. Between September and December, 1996, microbial contamination and water activity of in-use lubricants in sheathed and lubricated non-touch catheters connected to a tube used by 46 outpatients at our hospital was examined. Microbial contamination was detected at 5 to 2.6 x 10(8) colony-forming units (CFU)/ml in 14 (30.4%) of 46 samples. With higher water activity of the lubricant (a higher dilution rate of the lubricant with water), a higher concentration of microbial contamination was observed. In 3 (21.4%) of the 14 patients using contaminated lubricant, urine samples showed the same microbial species as those detected in the lubricant. To prevent microbial contamination of the lubricant, dilution of the lubricant with water, as a result of repeated use, should be avoided. PMID- 10864038 TI - Pharmacokinetics of cytosine arabinoside, methotrexate, nimustine and valproic acid in cerebrospinal fluid during cerebrospinal fluid perfusion chemotherapy. AB - This report investigates the pharmacokinetics of cytosine arabinoside (Ara-C), methotrexate (MTX), nimustine (ACNU) and valproic acid (VPA) in cerebrospinal fluid (CSF) during CSF perfusion chemotherapy. A 28-year-old Japanese woman with disseminated glioblastoma was, on admission, on a stable oral regimen of prolonged-release VPA tablets (Depakene-R), 400 mg twice a day, for seizure control. Twelve courses of CSF perfusion chemotherapy with Ara-C, MTX, and ACNU were administered. Plasma samples and CSF samples via Ommaya reservoirs were obtained during the eleventh course of treatment. The Ara-C and ACNU concentrations were measured by HPLC. The MTX and VPA concentrations were measured by fluorescence polarization immunoassay. During CSF perfusion chemotherapy, the highest CSF concentrations of Ara-C, MTX, and ACNU were observed at the end of the perfusion and decreased in a monoexponential pattern. The half-lives of Ara-C, MTX, and ACNU were 2.65, 3.52, and 0.71 h, respectively. No anticancer drugs were detectable in plasma during CSF perfusion chemotherapy. Before CSF perfusion chemotherapy, the free VPA concentration in plasma was 14.4% of the total VPA concentration. The mean total and free VPA concentrations in plasma were 78.0+/-0.8 and 10.9-0.3 microg/ml, respectively. The free VPA concentrations in plasma and in CSF were of similar values. At the end of perfusion, the lowest free VPA concentration in CSF was 30.3% of that at the initiation of perfusion. The free VPA concentrations in CSF at 3, 7, 23, and 47 h after the end of perfusion were 79.8, 94.5, 100.9, and 100.9% respectively of that at the initiation of perfusion. During CSF perfusion chemotherapy, the ratio of free VPA concentrations to the total VPA in CSF was 86.3+/-6.9%. The VPA concentrations in CSF rapidly decreased during the CSF perfusion but recovered to pre-treatment levels within 7 h. PMID- 10864039 TI - Maximizing RNA folding rates: a balancing act. AB - Large ribozymes typically require very long times to refold into their active conformation in vitro, because the RNA is easily trapped in metastable misfolded structures. Theoretical models show that the probability of misfolding is reduced when local and long-range interactions in the RNA are balanced. Using the folding kinetics of the Tetrahymena ribozyme as an example, we propose that folding rates are maximized when the free energies of forming independent domains are similar to each other. A prediction is that the folding pathway of the ribozyme can be reversed by inverting the relative stability of the tertiary domains. This result suggests strategies for optimizing ribozyme sequences for therapeutics and structural studies. PMID- 10864040 TI - An unconventional origin of metal-ion rescue and inhibition in the Tetrahymena group I ribozyme reaction. AB - The presence of catalytic metal ions in RNA active sites has often been inferred from metal-ion rescue of modified substrates and sometimes from inhibitory effects of alternative metal ions. Herein we report that, in the Tetrahymena group I ribozyme reaction, the deleterious effect of a thio substitution at the pro-Sp position of the reactive phosphoryl group is rescued by Mn2+. However, analysis of the reaction of this thio substrate and of substrates with other modifications strongly suggest that this rescue does not stem from a direct Mn2+ interaction with the Sp sulfur. Instead, the apparent rescue arises from a Mn2+ ion interacting with the residue immediately 3' of the cleavage site, A(+1), that stabilizes the tertiary interactions between the oligonucleotide substrate (S) and the active site. This metal site is referred to as site D herein. We also present evidence that a previously observed Ca2+ ion that inhibits the chemical step binds to metal site D. These and other observations suggest that, whereas the interactions of Mn2+ at site D are favorable for the chemical reaction, the Ca2+ at site D exerts its inhibitory effect by disrupting the alignment of the substrates within the active site. These results emphasize the vigilance necessary in the design and interpretation of metal-ion rescue and inhibition experiments. Conversely, in-depth mechanistic analysis of the effects of site specific substrate modifications can allow the effects of specific metal ion-RNA interactions to be revealed and the properties of individual metal-ion sites to be probed, even within the sea of metal ions bound to RNA. PMID- 10864041 TI - A four-nucleotide translation enhancer in the 3'-terminal consensus sequence of the nonpolyadenylated mRNAs of rotavirus. AB - The 5' cap and poly(A) tail of eukaryotic mRNAs work synergistically to enhance translation through a process that requires interaction of the cap-associated eukaryotic initiation factor, eIF-4G, and the poly(A)-binding protein, PABP. Because the mRNAs of rotavirus, and other members of the Reoviridae, contain caps but lack poly(A) tails, their translation may be enhanced through a unique mechanism. To identify translation-enhancement elements in the viral mRNAs that stimulate translation in vivo, chimeric RNAs were prepared that contained an open reading frame for luciferase and the 5' and 3' untranslated regions (UTRs) of a rotavirus mRNA or of a nonviral mRNA. Transfection of the chimeric RNAs into rotavirus-infected cells showed that the viral 3' UTR contained a translation enhancement element that promoted gene expression. The element did not enhance gene expression in uninfected cells and did not affect the stability of the RNAs. Mutagenesis showed that the conserved sequence GACC located at the 3' end of rotavirus mRNAs operated as an enhancement element. The 3'-GACC element stimulated protein expression independently of the sequence of the 5' UTR, although efficient expression required the RNA to contain a cap. The results indicate that the expression of viral proteins in rotavirus-infected cells is specifically up-regulated by the activity of a novel 4-nt 3' translation enhancer (TE) common to the 11 nonpolyadenylated mRNAs of the virus. The 4-nt sequence of the rotavirus 3' TE represents by far the shortest of any of the sequence enhancers known to stimulate translation. PMID- 10864042 TI - Rrp8p is a yeast nucleolar protein functionally linked to Gar1p and involved in pre-rRNA cleavage at site A2. AB - Chemical modifications and processing of the 18S, 5.8S, and 25S ribosomal RNAs from the 35S pre-ribosomal RNA depend on an important set of small nucleolar ribonucleoprotein particles (snoRNPs). Genetic depletion of yeast Gar1p, an essential common component of H/ACA snoRNPs, leads to inhibition of uridine isomerizations to pseudo-uridines on the 35S pre-rRNA and of the early pre-rRNA cleavages at sites A1 and A2, resulting in a loss of mature 18S rRNA synthesis. To identify Gar1p functional partners, we screened for mutations that are synthetically lethal with a gar1 mutant allele encoding a Gar1p mutant protein lacking its two glycine/arginine-rich (GAR) domains. We identified a previously uncharacterized Saccharomyces cerevisiae open reading frame, YDR083W (now designated RRP8), that encodes a highly conserved protein containing motifs found in methyltransferases. Rrp8p localizes to the nucleolus. A yeast strain lacking this protein is viable at 30 degrees C but displays strong growth impairment at lower temperatures. In this strain, cleavage of the pre-rRNA at site A2 is strongly affected whereas cleavages at sites A0 and A1 are only slightly inhibited or delayed. PMID- 10864043 TI - Identification of the TRM2 gene encoding the tRNA(m5U54)methyltransferase of Saccharomyces cerevisiae. AB - The presence of 5-methyluridine (m5U) at position 54 is a ubiquitous feature of most bacterial and eukaryotic elongator tRNAs. In this study, we have identified and characterized the TRM2 gene that encodes the tRNA(m5U54)methyltransferase, responsible for the formation of this modified nucleoside in Saccharomyces cerevisiae. Transfer RNA isolated from TRM2-disrupted yeast strains does not contain the m5U54 nucleoside. Moreover, a glutathione S-transferase (GST) tagged recombinant, Trm2p, expressed in Escherichia coli displayed tRNA(m5U54)methyltransferase activity using as substrate tRNA isolated from a trm2 mutant strain, but not tRNA isolated from a TRM2 wild-type strain. In contrast to what is found for the tRNA(m5U54)methyltransferase encoding gene trmA+ in E. coli, the TRM2 gene is not essential for cell viability and a deletion strain shows no obvious phenotype. Surprisingly, we found that the TRM2 gene was previously identified as the RNC1/NUD1 gene, believed to encode the yNucR endo-exonuclease. The expression and activity of the yNucR endo-exonuclease is dependent on the RAD52 gene, and does not respond to increased gene dosage of the RNC1/NUD1 gene. In contrast, we find that the expression of a trm2-LacZ fusion and the activity of the tRNA(m5U54)methyltransferase is not regulated by the RAD52 gene and does respond on increased gene dosage of the TRM2 (RNC1/NUD1) gene. Furthermore, there was no nuclease activity associated with a GST-Trm2 recombinant protein. The purified yNucR endo-exonuclease has been reported to have an NH2-D-E-K-N-L motif, which is not found in the Trm2p. Therefore, we suggest that the yNucR endo-exonuclease is encoded by a gene other than TRM2. PMID- 10864044 TI - Box C/D snoRNA-associated proteins: two pairs of evolutionarily ancient proteins and possible links to replication and transcription. AB - The eukaryotic nucleolus contains a diverse population of small nucleolar RNAs (snoRNAs) essential for ribosome biogenesis. The box C/D snoRNA family possesses conserved nucleotide boxes C and D that are multifunctional elements required for snoRNA processing, snoRNA transport to the nucleolus, and 2'-O-methylation of ribosomal RNA. We have previously demonstrated that the assembly of an snoRNP complex is essential for processing the intronic box C/D snoRNAs and that specific nuclear proteins associate with the box C/D core motif in vitro. Using a box C/D motif derived from mouse U14 snoRNA, we have now affinity purified and defined four mouse proteins that associate with this minimal RNA substrate. These four proteins consist of two protein pairs: members of each pair are highly related in sequence. One protein pair corresponds to the essential yeast nucleolar proteins Nop56p and Nop58p. Affinity purification of mouse Nop58 confirms observations made in yeast that Nop58 is a core protein of the box C/D snoRNP complex. Isolation of Nop56 using this RNA motif defines an additional snoRNP core protein. The second pair of mouse proteins, designated p50 and p55, are also highly conserved among eukaryotes. Antibody probing of nuclear fractions revealed a predominance of p55 and p50 in the nucleoplasm, suggesting a possible role for the p50/p55 pair in snoRNA production and/or nucleolar transport. The reported interaction of p55 with TATA-binding protein (TBP) and replication A protein as well as the DNA helicase activity of p55 and p50 may suggest the coordination of snoRNA processing and snoRNP assembly with replication and/or transcriptional events in the nucleus. Homologs for both snoRNA-associated protein pairs occur in Archaea, strengthening the hypothesis that the box C/D RNA elements and their interacting proteins are of ancient evolutionary origin. PMID- 10864045 TI - RNA secondary structures of the bacteriophage phi6 packaging regions. AB - Bacteriophage phi6 genome consists of three segments of double-stranded RNA. During maturation, single-stranded copies of these segments are packaged into preformed polymerase complex particles. Only phi6 RNA is packaged, and each particle contains only one copy of each segment. An in vitro packaging and replication assay has been developed for phi6, and the packaging signals (pac sites) have been mapped to the 5' ends of the RNA segments. In this study, we propose secondary structure models for the pac sites of phi6 single-stranded RNA segments. Our models accommodate data from structure-specific chemical modifications, free energy minimizations, and phylogenetic comparisons. Previously reported pac site deletion studies are also discussed. Each pac site possesses a unique architecture, that, however, contains common structural elements. PMID- 10864046 TI - RNA location and modeling of a WD40 repeat domain within the vault. AB - The vault complex is a ubiquitous 13-MDa ribonucleoprotein assembly, composed of three proteins (TEP1, 240 kDa; VPARP, 193 kDa; and MVP, 100 kDa) that are highly conserved in eukaryotes and an untranslated RNA (vRNA). The vault has been shown to affect multidrug resistance in cancer cells, and one particular component, MVP, is thought to play a role in the transport of drug from the nucleus. To locate the position of the vRNA, vaults were treated with RNases, and cryo electron microscopy (cryo-EM) was performed on the resulting complexes. Using single-particle reconstruction techniques, 3,476 particle images were combined to generate a 22-A-resolution structure. Difference mapping between the RNase treated vault and the previously calculated intact vault reconstructions reveals the vRNA to be at the ends of the vault caps. In this position, the vRNA may interact with both the interior and exterior environments of the vault. The finding of a 16-fold density ring at the top of the cap has allowed modeling of the WD40 repeat domain of the vault TEP1 protein within the cryo-EM vault density. Both stoichiometric considerations and the finding of higher resolution for the computationally selected and refined "barrel only" images indicate a possible symmetry mismatch between the barrel and the caps. The molecular architecture of the complex is emerging, with 96 copies of MVP composing the eightfold symmetric barrel, and the vRNA together with one copy of TEP1 and four predicted copies of VPARP comprising each cap. PMID- 10864048 TI - Enhanced detection of tRNA isoacceptors by combinatorial oligonucleotide hybridization. AB - A method that greatly enhances the detection of tRNA by oligodeoxyribonucleotide probe hybridization has been developed. Because highly structured tRNA regions often preclude heteroduplex formation, we have tested the ability of cold oligodeoxyribonucleotides called unfolders to disrupt the tRNA secondary/tertiary structures and promote hybridization of a second labeled oligonucleotide complementary to the anticodon loop. Here we show that an excess of unfolders in the pre/hybridization reaction can enhance a barely detectable hybridization signal by more than 200-fold without affecting probe specificity. This sensitive assay makes it possible to easily study and monitor changes in tRNA isoacceptor expression. PMID- 10864047 TI - Differential recognition of the polypyrimidine-tract by the general splicing factor U2AF65 and the splicing repressor sex-lethal. AB - The polypyrimidine-tract (Py-tract) adjacent to 3' splice sites is an essential splicing signal and is recognized by several proteins, including the general splicing factor U2AF65 and the highly specific splicing repressor Sex-lethal (SXL). They both contain ribonucleoprotein-consensus RNA-binding motifs. However, U2AF65 recognizes a wide variety of Py-tracts, whereas SXL recognizes specific Py tracts such as the nonsex-specific Py-tract of the transformer pre-mRNA. It is not understood how these seemingly similar proteins differentially recognize the Py-tract. To define these interactions, we used chemical interference and protection assays, saturation mutagenesis, and RNAs containing modified nucleotides. We find that these proteins recognize distinct features of the RNA. First, although uracils within the Py-tract are protected from chemical modification by both of these proteins, modification of any one of seven uracils by hydrazine, or any of eight phosphates by ethylnitrosourea strongly interfered with the binding of SXL only. Second, the 2' hydroxyl groups or backbone conformation appeared important for the binding of SXL, but not U2AF65. Third, although any of the bases (cytosine >> adenine > guanine) could substitute for uracils for U2AF65 binding, only guanine partially substituted for certain uracils for SXL binding. The different dependence on individual contacts and nucleotide preference may provide a basis for the different RNA-binding specificities and thus functions of U2AF65 and SXL in 3' splice site choice. PMID- 10864049 TI - Corticosteroids and cognitive function in humans: methodological considerations. PMID- 10864050 TI - Natural history and biology of stage A neuroblastoma: a Pediatric Oncology Group Study. AB - PURPOSE: To prospectively analyze the outcome of patients with Stage A neuroblastoma (NB) treated with surgery alone, especially with regard to the prognostic significance of age, tumor site, MYCN copy number, tumor cell ploidy, and histology. PATIENTS AND METHODS: The clinical course of 329 patients with Stage A disease registered on the POG NB Biology Study #9047 between February, 1990 and October, 1997 were evaluated. Age, tumor site, MYCN copy number, tumor cell ploidy, and histology were analyzed for their impact on event-free survival (EFS) and survival (S). RESULTS: The 5-year estimated EFS and S rates for the 329 patients were 91% (+/-3%) and 96% (+/-2%), respectively. The EFS rate was similar for infants younger than 12 months and children age 12 months or older, but age older than 12 months was predictive of lower S rates (P = 0.044). Patients with adrenal, abdominal non-adrenal, thoracic, and cervical tumors had similar S rates. The majority of patients had tumors with favorable biologic features, and only 3% had MYCN amplification. For infants with diploid tumors, the EFS rate was 82% (+/-16%), but effective therapy yielded an S rate of 100%. Rate of S was 80% (+/-26%) and 64% (+/-27%) for patients with unfavorable tumor histology and MYCN amplified tumors, respectively. CONCLUSION: The outcome for patients with Stage A NB treated with surgery alone is excellent. Although EFS and S rates were significantly worse for patients with MYCN-amplified tumors, a subset achieved long-term remission after surgery alone. For patients with Stage A and MYCN amplification, additional factors are needed to distinguish the patients who will achieve long-term remission with surgery alone from those who will develop recurrent disease. PMID- 10864051 TI - Cognitive sequelae in children treated for acute lymphoblastic leukemia with dexamethasone or prednisone. AB - PURPOSE: The cognitive sequelae of treatment for childhood acute lymphoblastic leukemia (ALL) were compared in a group of patients who received dexamethasone during the intensification and maintenance phases of therapy with those in a historical control group for whom antileukemia therapy was similar, except that the corticosteroid component of therapy was prednisone. METHODS: Patients treated for ALL on Dana-Farber Cancer Institute protocols 87-01 (n = 44) and 91-01 (n = 23) were evaluated by standard cognitive and achievement tests. Corticosteroid therapy was delivered in 5-day pulses given every 3 weeks during intensification and continuation phases of therapy for a total of 2 years. RESULTS: Children treated on protocol 87-01 received prednisone at a dose of 40 mg/m2/d (standard risk, SR) or 120 mg/ m2/d (high risk, HR); those treated on protocol 91-01 received dexamethasone at a dose of 6 mg/m2 per day (SR) or 18 mg/m2 per day (HR). Children treated on protocol 91-01 performed less well on cognitive testing. Subsample analysis indicated that cranial radiation therapy and methotrexate dose did not account for differences in cognitive outcomes. CONCLUSIONS: The findings of this preliminary study are consistent with the hypothesis that dexamethasone therapy can increase risk for neurocognitive late effects in children treated for ALL and indicate that further investigation of this question is warranted. PMID- 10864052 TI - Childhood cancer: a 4-year prospective study of the psychological adjustment of children and parents. AB - PURPOSE: The objective of this 4-year prospective study was to assess the psychological adjustment of children treated for cancer and their parents. PATIENTS AND METHODS: Children aged 2 to 12 years with cancer diagnosed and their parents and families (n = 39) were assessed immediately after their diagnosis and then annually for the next 4 years. At each assessment, the psychological adjustment of the children and their families was compared with the adjustment of a cohort of children and families in the general community (n = 49). RESULTS: Immediately after the diagnosis of cancer in the children, the children and their parents had significantly more psychological problems than children and parents in the community. However, at subsequent assessments, there was no difference in the number of psychological problems experienced by children and parents in the two groups. CONCLUSIONS: In the longer term, the prevalence of psychological problems experienced by children treated for cancer and their parents does not differ from that found in children and parents in the general community. Future research should give greater attention to other aspects of the lives of children treated for cancer and their parents, including their broader health-related quality of life. PMID- 10864054 TI - Growth factor practice patterns among pediatric oncologists: results of a 1998 Pediatric Oncology Group Survey. Economic Evaluation Working Group the Pediatric Oncology Group. AB - The American Society of Clinical Oncology (ASCO) guidelines on growth factor (GF) use recommend applying adult-derived guidelines in pediatric oncology. An ASCO survey of adult oncology GF use determined the preference for first degree prophylaxis (use of GF when febrile neutropenia [FN] is expected to be high in untreated patients), second-degree prophylaxis (administration of GF after a documented episode of FN on a previous cycle of chemotherapy), and intervention in the treatment of FN. Similar preferences have not been evaluated in pediatrics. The purpose of this study was to (1) characterize GF use in pediatric oncology; (2) correlate use patterns with demographic factors; and (3) compare the Pediatric Oncology Group (POG) and ASCO surveys. The ASCO survey was revised for use within pediatric oncology and was mailed to the physician membership of POG; 341 were returned (86% completion rate). Comparisons were made with the ASCO survey. Most (76%) physicians said GF use was determined by protocol requirements and most (70%) patients were entered on POG protocols. GF use as first-degree prophylaxis was selected 40% of the time, which was significantly greater than in adults; this was most influenced by anticipated duration of neutropenia (> or =7 days). The severity of the initial clinical course (e.g., neutropenia, infection) influenced use in second-degree prophylaxis; dose reduction alone was never selected. For FN, GF use was 45%, with lower preferences in uncomplicated FN (16% 38%) compared with complicated FN (66%). POG respondents endorse greater use of GF for first-and second-degree prophylaxis but less use in uncomplicated FN than do ASCO respondents. These patterns may reflect different strategies, including the role of chemotherapy, value of dose intensity, and perceived toxicity of regimens. Given these differences, adult-based guidelines may not be appropriate for pediatrics. PMID- 10864053 TI - Ratio of methotrexate to folate uptake by lymphoblasts in children with B-lineage acute lymphoblastic leukemia: a pilot study. AB - PURPOSE: Methotrexate (MTX) remains one of the most effective drugs for the treatment of children with acute lymphoblastic leukemia (ALL). Because MTX and 5 methyltetrahydrofolate (5CH3THF) share uptake and metabolic pathways, the efficacy of MTX is likely to depend not only on its metabolism but also on how well folate is accumulated by lymphoblasts. The authors' goal was to compare in vitro folate and antifolate uptake in B-lineage lymphoblasts from patients who remained in continuous complete remission (CCR) and those in whom relapse occurred. PATIENTS AND METHODS: Twenty-four children with B-lineage ALL were studied at diagnosis (n = 20) or relapse (n = 4). Lymphoblasts obtained by bone marrow aspiration were incubated for 24 hours in vitro with 0.05 microM 5CH3[3H]THF or 1 microM [3H]MTX. RESULTS: As of July 1999, 16 patients studied at diagnosis remained in CCR at a median follow-up of 45 months after achieving remission. Two of the patients studied at relapse are in second CCR; the remaining two died from progressive disease. The median uptake of neither [3H]MTX nor 5CH3[3H]THF differed significantly between the 16 patients in first CCR studied at diagnosis and the 4 patients studied at relapse. However, the median ratio of [3H]MTX:5CH3[3H]THF uptake differed significantly for patients who remained in first CCR versus patients studied at relapse. CONCLUSIONS: The uptake of [3H]MTX in relation to 5CH3[3H]THF by leukemic lymphoblasts in vitro may correlate positively with treatment outcome in children with B-lineage ALL. A larger study of homogeneously treated patients is necessary to confirm these results. PMID- 10864055 TI - Use of amphotericin B colloidal dispersion in children. AB - PURPOSE: To describe the experience with a new lipid-based amphotericin product (amphotericin B colloidal dispersion or ABCD) in children with fever and neutropenia who are at high risk for fungal infection. PATIENTS AND METHODS: Forty-nine children with febrile neutropenia were treated in a prospective, randomized trial comparing ABCD with amphotericin B. An additional 70 children with presumed or proven fungal infection were treated with 5 different open-label studies of ABCD. Patients were registered into these studies for reasons of: 1) failure to respond to amphotericin B; 2) development of nephrotoxicity or preexisting renal impairment; or 3) willingness to participate in a dose escalation study. Extensive data detailing response and toxicity were collected from each patient. RESULTS: In the randomized trial, there was significantly less renal toxicity in the children receiving ABCD than in those receiving amphotericin B (12.0% vs. 52.4% [P = 0.003]). Other adverse symptoms were not significantly different. In the additional open-label studies, although 80% of patients receiving ABCD reported some adverse symptom, the majority of these were infusion related, and nephrotoxicity was reported in only 12% of these patients. CONCLUSIONS: ABCD was well-tolerated at doses up to 5 times greater then those usually tolerated with amphotericin B. Renal toxicity was markedly less than expected, and there were no other unexpected severe toxicities. Further randomized studies are needed to further define the role of this and other liposomal products in children. PMID- 10864056 TI - Tamoxifen and carboplatin for children with low-grade gliomas: a pilot study at St. Jude Children's Research Hospital. AB - PURPOSE: The authors conducted a single-arm, prospective study using tamoxifen and carboplatin for the treatment of children with progressive or symptomatic low grade gliomas. PATIENTS AND METHODS: Fourteen children with consecutively diagnosed cases of low-grade glioma were enrolled in this Study; all patients were younger than 14 years. One patient was excluded after induction chemotherapy because of the diagnosis of a nonmalignant condition. Patients were treated with daily tamoxifen (20 mg/m2 administered twice per day) in addition to targeted, monthly intravenous carboplatin at an area under the curve (AUC) exposure of 6.5 mg/mL x minute for 1 year or until they had clinical or radiologic evidence of disease progression. RESULTS: The median age at diagnosis was 5.3 years, the median age at initiation of chemotherapy was 8.3 years. Eight patients had tumors of the hypothalamus/optic pathway, two patients had thalamic tumors, and one patient each had tumors in the temporal lobe, tectum, and brain stem. Tumor histologic findings included fibrillary astrocytoma (n = 2), juvenile pilocytic astrocytoma (n = 6), and oligodendroglioma (n = 1). The best response to therapy was a partial response in two patients, stable disease in nine patients, and progressive disease in two patients. The overall survival at 3 years is 69%. The 3-year progression-free survival is 47%. Tamoxifen and carboplatin chemotherapy did not result in a significant number of objective responses in children with low-grade gliomas. The progression-free survival is similar to that of other published series. Nonmyelosuppressive agents such as tamoxifen deserve additional evaluation in the treatment of children with low-grade gliomas. PMID- 10864057 TI - Spontaneous remission of congenital leukemia: a case for conservative treatment. AB - A newborn infant with spontaneous remission of congenital leukemia cutis is described and a literature review of this uncommon phenomenon is provided. In view of the unusual and unpredictable behavior of this disease, chemotherapy should be withheld unless there is evidence of an 11q23 translocation or progressive disease. Otherwise, overall survival does not appear to be affected by adopting a conservative approach. Because of occasional late relapses, long term follow-up is recommended. The biologic basis underlying spontaneous remission of congenital leukemia is unknown; therefore, molecular or molecular cytogenetic analysis of DNA obtained from a skin biopsy is recommended. PMID- 10864058 TI - Intratumoral consumption of indium-111-labeled platelets in a child with splenic hemangioma and thrombocytopenia. AB - The authors report Kasabach-Merritt syndrome (KMS) in a patient with thrombocytopenia and splenic hemangioma. A 13-month-old boy with a history of anemia, thrombocytopenia, and abdominal mass was admitted to the hospital. The scintigraphic studies showed that a large mass contiguous to the spleen was responsible for the platelet uptake. After partial splenectomy, the platelet count returned to normal. This report of KMS in a child with splenic hemangioma suggests that the scintigraphic studies are mandatory to confirm diagnosis. Indium-111-labeled platelets are useful in identifying hemangiomatous sequestration of platelets in patients with thrombocytopenia. PMID- 10864059 TI - Fetal schistocytic hemolytic anemia and umbilical vein varix. AB - A rare case of schistocytic hemolytic anemia presenting in a fetus secondary to a varix of the intra-abdominal umbilical vein is reported. A patient was referred to our hospital at 32 weeks of gestation because of an abnormal hypoechoic finding in the fetal liver. Prenatal ultrasound showed turbulent flow through a 12-mm diameter dilatation of the fetal intra-abdominal umbilical vein consistent with a varix. Cardiomegaly also was noted. At birth, the 1098-g, growth-retarded, male neonate was in severe congestive heart failure secondary to anemia as the initial hemoglobin was 5 g/dL. Additional evaluation found the anemia to be secondary to schistocytic hemolysis. After the neonate received a transfusion of packed erythrocytes and supportive care, the anemia quickly resolved, and he was discharged to home doing well after a 6-week stay in the neonatal intensive care unit. Prompt recognition of the varix prenatally and thorough evaluation of the newborn postnatally led to appropriate diagnosis and treatment. PMID- 10864060 TI - Postoperative stroke in a child with cerebral palsy heterozygous for factor V Leiden. AB - A 5-year-old with spastic quadraparetic cerebral palsy suffered multiple strokes after extensive orthopedic surgery. Coagulation testing was undertaken to determine whether a familial thrombophilia was present. The patient was found to be heterozygous for factor V Leiden. Factor V Leiden may be a risk factor for central nervous system events in special-needs children, particularly when common medical conditions create additional procoagulant risks. PMID- 10864061 TI - The National Pediatric Blood Club: endothelium and coagulation in the newborn. PMID- 10864062 TI - Comparison of human umbilical vein and adult saphenous vein endothelial cells: implications for newborn hemostasis and for laboratory models of endothelial cell function. PMID- 10864063 TI - Interactions between von Willebrand factor and Factor VIII: where did they first meet. PMID- 10864064 TI - Synthesis of coagulation proteins in the fetus and neonate. PMID- 10864065 TI - Hemostatic factors in tumor biology. PMID- 10864067 TI - Hepatocellular carcinoma: a rare late event in childhood acute lymphoblastic leukemia. PMID- 10864066 TI - Mutation analysis and therapeutic response to granulocyte colony-stimulating factor in a case of hyperimmunoglobulin M syndrome with chronic neutropenia. PMID- 10864068 TI - CT angiography of thoracic outlet syndrome: evaluation of imaging protocols for the detection of arterial stenosis. AB - PURPOSE: The purpose of this work was to evaluate the results of cross-sectional imaging and multiplanar and 3D reconstructions for the detection of thoracic outlet arterial stenosis on CT angiograms. METHOD: Eighty-two patients were prospectively evaluated with CT angiography: in the neutral position and after postural maneuver (164 acquisitions); with contralateral injection of a 24% (Group 1; n = 68) or 30% (Group 2; n = 96) contrast agent; and reconstruction of four sets of images from each acquisition, that is, transverse CT scans, sagittal reformations, and 3D [shaded surface displays (SSD) and volume-rendered (VR)] images. A total of 656 sets of images were blindly and independently interpreted by three readers of variable experience. A consensus interpretation of the four sets of images of each acquisition was used as a standard of reference. RESULTS: The number of examinations coded with an excellent degree of arterial enhancement was significantly higher in Group 2 than in Group 1 [68 (71%) vs. 35 (51%); p < 0.001]. The sensitivity and specificity for detection of arterial stenosis were 67 and 96% for transverse CT scans, 69 and 94% for sagittal reformations, 71 and 99% for 3D-SSDs, and 95 and 100% for VR images. Compared with the standard of reference, a concordant scoring of arterial stenosis severity was found in 54% of transverse CT scans, 84% of sagittal reformations, 78% of 3D-SSDs, and 91% of VR images. Underestimation of stenosis was found in 43% of transverse CT scans and 10% of sagittal reformations; overestimation of stenosis was more frequent on 3D SSDs (16%) than on VR images (7%). The reader's experience was marked for the interpretation of cross-sectional images but did not influence the interpretation of 3D images. CONCLUSION: Thoracic outlet arterial compression is best depicted with the injection of a 30% contrast agent and reconstruction of VR images. PMID- 10864069 TI - MRA of the Adamkiewicz artery: a preoperative study for thoracic aortic aneurysm. AB - PURPOSE: The purpose of this work was to investigate the ability of MR angiography (MRA) to visualize the Adamkiewicz artery (AKA) as a preoperative study of thoracic aortic aneurysm to prevent ischemic injury of the spinal cord. METHOD: Twenty-six patients scheduled for surgical or endovascular stent-graft repair of thoracic aortic aneurysm were studied with a three-dimensional contrast MRA. Data acquisition was repeated two times following injection of Gd-DTPA. Source images were processed with multiplanar reconstruction and maximum intensity projection. RESULTS: The AKA was identified in 69% (18/26). In three patients, selective angiography of the intercostal artery confirmed the AKA at the same level and side predicted by MRA. The anterior spinal artery and the anterior medullary vein were observed in 50% (13/26) and 65% (17/26), respectively. CONCLUSION: Contrast MRA is a promising technique to visualize the AKA noninvasively as a preoperative evaluation of thoracic aortic aneurysms. PMID- 10864070 TI - Contrast-enhanced 3D FISP MR angiography of the aortic arch ostia: preliminary results. AB - PURPOSE: The goal of this work was to evaluate three-dimensional (3D) contrast enhanced MR angiography (MRA) for the detection of ostial stenoses of the aortic arch. METHOD: Sixteen patients with suspected carotid atherosclerotic disease prospectively underwent digital subtraction angiography of the aortic arch followed by contrast-enhanced MRA using a 3D fast imaging with steady-state precession (FISP) technique (TR = 5 ms, TE = 2 ms, flip angle = 30 degrees). Three neuroradiologists blindly measured stenoses on the catheter angiograms and MRA. Evaluation included the ostia of the innominate, left carotid, and left subclavian arteries. Any significant disagreement on catheter angiography was resolved by consensus. The MRA grades of each of the three observers were then compared with the consensus grades of the contrast angiogram. RESULTS: Forty eight vessels were scored, of which five had significant stenoses. MRA demonstrated 100% sensitivity, 89% specificity, 52% positive predictive value, and 100% negative predictive value. The Bowker test for symmetry indicated no significant difference between conventional angiography and MRA scores (p = 0.32 0.75), and there was good agreement between the three observers (weighted kappa = 0.75-0.86). CONCLUSION: Contrast-enhanced 3D FISP MRA may be a useful imaging modality for the detection of significant stenoses at the ostia of the major aortic arch branches. PMID- 10864071 TI - Repeated 3D coronary MR angiography with navigator echo gating: technical quality and consistency of image interpretation. AB - The purpose of this study was to evaluate the technical quality of 3D coronary MR angiography (CMRA) with navigator echo and the consistency of image interpretation in repeated imaging sessions. Fourteen subjects underwent CMRA, 10 of whom were imaged twice. The coronary arteries (96 segments) were analyzed twice for hemodynamically significant stenoses. Signal-to-noise and contrast-to noise ratios varied considerably between the two imagings. Fat saturation was poor or satisfactory in 37%; in 15% of the slabs, the severity of artifacts was moderate; and the overall quality was good to excellent in only 42% of the imagings. The intraobserver reproducibility was good (kappa = 0.54), but the consistency of interpretation for repeated CMRA was only satisfactory (kappa = 0.43). Sensitivities of 84 and 84% and specificities of 70 and 62% were obtained for the two readings. Although the reproducibility of image reading is good, 3D CMRA with navigator echo provides only fair technical consistency, and the frequently compromised image quality impairs the clinical utility of this technique. PMID- 10864073 TI - MRI in staging advanced gastric cancer: is it useful compared with spiral CT? AB - PURPOSE: During the last decade, rapid progress has been made in MR technology. Our objective was to evaluate the role of MRI in staging advanced gastric cancer (AGC; gastric cancer invading the muscularis propria) and to compare it with that of spiral CT. METHOD: We prospectively performed both MR and CT examinations on 26 patients with AGC proven by endoscopic biopsy. Contrast-enhanced CT and nonenhanced MRI with a 1.0 T scanner using FLASH, HASTE, and true-FISP sequences were obtained in each patient after injection of antiperistaltic drug and ingestion of 1 L of tap water. Fifty-two sets of CT and MR images were analyzed by two radiologists in consensus without any information from other images. T and N staging of AGC was determined according to the TNM classification. All patients underwent surgery within 1 week after both examinations. Diagnostic accuracy of each staging of AGC on CT or MRI was evaluated by comparison with the pathologic results. RESULTS: MRI was slightly superior to CT in T staging (81 vs. 73%, respectively; p < 0.05). Although MRI had a tendency to overstage the pathologic T2 cancer, positive predictability of T2 stage and sensitivity of T3 stage were high (100%, respectively). Regarding the N staging, CT was slightly superior to MRI (73 vs. 65%; p > 0.05). However, both CT and MRI demonstrated the tendency of understaging in N staging. CONCLUSION: Although MRI was superior to spiral CT in T staging, MRI cannot completely replace spiral CT in staging AGC because of its limitation in N staging. PMID- 10864072 TI - Comparison of cystourethrography and dynamic MRI in bladder neck descent. AB - PURPOSE: The purpose of this work was to test whether there are statistically significant differences between dynamic MR and lateral cystourethrogram measurement results in patients with bladder neck descent. METHOD: Twenty-seven women (39-83 years old, mean 60.6 years old) with urinary incontinence and bladder neck descent were examined by dynamic MRI using a single shot fast spin echo sequence with half-Fourier data acquisition. Bladder neck position, angle of inclination of the urethral axis, posterior vesicourethral angle, and depth of cystoceles were measured at perineal contraction and at maximal pelvic strain. The nonparametric Wilcoxon test for paired values was used to analyze whether there were statistically significant differences between lateral cystourethrogram and dynamic MR measurement results. The Spearman correlation coefficient (rs) was calculated for all parameter pairs. RESULTS: Measurements at maximal pelvic strain showed the greatest levels of agreement between MRI and cystourethrography. The best results were attained for the cystocele measurements (p > 0.5, rs = 0.95). Bladder neck position showed the second best agreement; if MR measurements were corrected by 0.46 cm, no statistically significant difference (p > 0.2, rs = 0.92) was calculated. Measurements at perineal contraction tallied least, probably due to the different positions adopted during the two examinations. CONCLUSION: Measurement data on dynamic MRI for the bladder neck position and the extension of cystocele at maximal pelvic strain are comparable with lateral cystourethrogram data. PMID- 10864074 TI - Primary papillary serous carcinoma of the peritoneum: CT-pathologic correlation. AB - We present the CT findings of three cases of primary papillary serous carcinoma of the peritoneum. All patients presented with massive ascites. CT of the abdomen and pelvis showed omental caking in all patients. The parietal peritoneum of the pelvis showed diffuse enhancement with nodular thickening in all patients. No calcification was noted in the omental and parietal peritoneal masses, although psammoma bodies were present microscopically in one case. The ovaries were normal in size but showed a fine enhancing surface nodularity similar to the pelvic peritoneum. The CT findings of primary papillary serous carcinoma of the peritoneum are nonspecific, but this diagnosis should be considered when peritoneal carcinomatosis is seen on CT with normal-sized ovaries in the absence of other primary malignant neoplasms. PMID- 10864075 TI - Factors influencing vascular and hepatic enhancement at CT: experimental study on injection protocol using a canine model. AB - PURPOSE: The purpose of this work was to evaluate the effects of contrast medium injection parameters on aortic, portal vein, and hepatic enhancement at spiral CT and to assess optimal injection protocol for hepatic CT. METHOD: Ten 15 kg dogs underwent single level dynamic CT through the hepatic hilum at 5 s intervals just after the injection of contrast medium for 3 min. With use of different volumes (1, 2, and 3 ml/kg), injection rates (0.5, 1, and 2 ml/s), and concentrations (150, 200, and 300 mg/ml), a total of 270 spiral CT scans were performed. In each scan, time-attenuation curves of aorta, portal vein, and liver were obtained. The degree of maximum contrast enhancement (Imax), time to maximum enhancement (Tmax), and time to equilibrium phase (Teq) for to each injection protocol were analyzed. RESULTS: Alterations in contrast material volume, injection rate, and concentration had significant impact on contrast enhancement of the liver. With increasing volume of contrast medium, Imax, Tmax, and Teq of aorta, portal vein, and liver increased (p < 0.005). With increasing rate of injection, on the other hand, Imax of aorta and liver increased (p < 0.05), but Tmax and Teq decreased (p < 0.005). Change of concentration of contrast medium had a significant effect on Imax of vessels (p < 0.05). CONCLUSION: Maximum contrast enhancement of liver and vessels was influenced mainly by injection volume of contrast medium and the time to peak enhancement by injection rate of contrast medium. Under given amounts of contrast medium, therefore, the strategy of increasing volume by dilution and faster injection might give better Imax values without penalty for the duration of an optimal temporal window (Tmax and Teq). PMID- 10864076 TI - Radiological features of leiomyomatous tumors of the colon and rectum. AB - PURPOSE: The purpose of this study was to evaluate the radiological features of 12 pathologically proven cases of colorectal leiomyomatous tumors. METHOD: A retrospective analysis of radiologic findings was performed in 12 patients with pathologically proven colorectal leiomyomatous tumors (2 leiomyomas and 10 leiomyosarcomas). Available radiologic studies included abdominal CT scans in 11 patients, double contrast barium studies in 4, and pelvic MRI in 1. On imaging, we evaluated the size, tumor margin (smooth or lobulated), morphologic appearance, growth patterns (endocolic, exocolic, or combined), contrast enhancement patterns, presence or absence of calcification within the tumors, and metastasis. RESULTS: The involved tumor sites were the colon in 2 patients and the rectum in 10. The mean tumor size was 7.9 cm (range 2-15 cm): It was 3.5 cm in leiomyomas and 8.8 cm in leiomyosarcomas. On imaging studies, the tumor margin was smooth in three patients and lobulated in nine, with endocolic growth in one, exocolic in four, and combined in the remaining seven. Eight of the 12 tumors showed varying degrees of internal necrosis with heterogeneous contrast enhancement. Dystrophic calcification was noted in five patients. Metastasis was seen in the liver in three patients at the time of initial diagnosis, and lymphadenopathy was noted in two patients (paraaortic space in one and perirectal space in two). CONCLUSION: Although rare, the diagnosis of leiomyomatous tumor may be suggested especially when the tumor occurring in the colorectum shows exocolic growth or calcification with varying degree of internal necrosis. PMID- 10864077 TI - Cystic struma ovarii: imaging findings. AB - We report three cases of cystic struma ovarii not associated with any solid component. One case was a thin-walled unilocular mass, and the other two cases were multilocular cystic masses. An area of signal void on T2-weighted images and intermediate intensity on T1-weighted images was noted in the two multilocular cases. Preoperative diagnosis was difficult in each case, but struma ovarii should be included in differential diagnoses even in the case of a completely cystic ovarian mass. PMID- 10864078 TI - Non-Hodgkin lymphomas of the ovaries: MR findings. AB - PURPOSE: The goal of this work was to describe MR findings (morphology, structure, signal intensity) of ovarian non-Hodgkin lymphoma (NHL). METHOD: We reviewed the MR images of five female patients aged 13-70 years (mean 46 years) with histologically proven NHL of the ovaries. We evaluated morphological and signal intensity findings of the lesions. MR features were correlated with pathologic parameters. RESULTS: All the patients were affected by B-cell NHL; one patient showed a primary involvement of the ovaries; in one patient, ovarian disease was diagnosed 30 months after surgical resection of a primary uterine lymphoma; the remaining three had a systemic lymphoma. In three cases, the ovarian involvement was bilateral. The mean size of the lesions was 7.9 cm. All the lesions showed homogeneous low signal intensity on T1-weighted images and intermediate to high intensity on T2-weighted images. The postgadolinium images showed mild to moderate heterogeneous enhancement. The peripheral enhancement was better demonstrated in fat-suppressed images. CONCLUSION: The diagnosis of primary ovarian lymphoma should be considered in the presence of large bilateral solid ovarian masses with homogeneous appearance (low signal on T1 and mildly high on T2) without infiltrative pattern of growth or regressive changes (necrosis, hemorrhage, calcifications) and with little contrast enhancement. PMID- 10864079 TI - Leiomyosarcoma presenting aneurysmal dilatation of the jejunum on CT. PMID- 10864080 TI - Urethral leiomyosarcoma: evaluation by MRI with pathologic correlation. PMID- 10864081 TI - Transient radiographic progression during initial treatment of pulmonary tuberculosis: CT findings. AB - PURPOSE: A study was undertaken to describe the thin-section CT features of transient radiographic progression during initial treatment of active pulmonary tuberculosis. METHOD: The CT scans of 13 patients who developed transient radiographic progression during initial treatment of pulmonary tuberculosis were evaluated and compared with those of 10 patients with true progression of pulmonary tuberculosis. Two patients underwent transbronchial lung biopsy for pathologic evaluation of new lesion. RESULTS: Eight patients had increased opacity at the site of their original lesion, whereas five had new opacities elsewhere: ipsilateral (n = 3) or contralateral (n = 1) to the original lesion or both (n = 1). Relatively extensive areas of ground-glass attenuation were concomitantly depicted in three of the eight patients with progression in the area of the original lesion. In those with progression ipsilateral or contralateral to the lesion, the character of the new lesions was ground-glass opacity and/or consolidation mainly in the subpleural region. Transbronchial lung biopsy specimens obtained from two patients showed intraluminal organizing exudates and thickened alveolar septa with lymphocyte infiltration, whereas the dominant CT findings of true progression of pulmonary tuberculosis were nodules (n = 8) or centrilobular nodules (n = 7). CONCLUSION: Thin-section CT may be useful to differentiate transient radiographic progression from true progression of pulmonary tuberculosis. PMID- 10864082 TI - Hand exercise during contrast medium delivery at thoracic helical CT: a simple method to minimize perivenous artifact. AB - PURPOSE: The purpose of this work was to assess the effects of hand exercise on perivenous artifact caused by undiluted venous contrast material at thoracic helical CT. METHOD: Eighty patients were prospectively and randomly assigned to thoracic helical CT with (n = 42) or without (n = 38) intermittent squeezing of a hand-sized ball during delivery of a contrast material. Two radiologists graded perivenous artifact and arterial enhancement in a blind fashion. CT attenuation values were obtained by region-of-interest measurements from arteries and veins. RESULTS: Both qualitative and quantitative analyses showed statistically significant differences in the assessment of perivenous artifact (p < 0.01). Perivenous artifact from the subclavian vein was significantly reduced in patients subjected to thoracic helical CT by using a hand exercise method. CONCLUSION: Hand exercise during contrast material delivery at thoracic helical CT minimizes perivenous artifact and improves image quality. PMID- 10864083 TI - Gradient correction and classification of CT lung images for the automated quantification of mosaic attenuation pattern. AB - PURPOSE: The detection of density differences, or "mosaic attenuation pattern," on CT images may be difficult when the regional inhomogeneity of the density of the lung parenchyma is subtle. The purpose of this work was to develop a fully automated method for the reproducible quantification of the underattenuated areas of the lung parenchyma. This technique may be useful in increasing the precision of investigation of structure/function relationships. METHOD: Anatomical segmentation was achieved by a structure-filtering operator based on mathematical morphology. To compensate for the density gradient visible on lung CT scans, a model-based iterative deconvolution filter and an adaptive clustering algorithm were developed. Validation was performed with CT images from a lung phantom, 15 patients with constrictive obliterative bronchiolitis, and 8 normal subjects. RESULTS: The accuracy of the estimate of the density gradient on phantom studies was 93.3%. The automated quantification of the areas of decreased attenuation on scans of constrictive obliterative bronchiolitis was within 8.2% from the average scoring of two experienced observers. CONCLUSION: The proposed technique is fully automated and can accurately correct for density gradient and classify areas of decreased attenuation on lung CT images. PMID- 10864084 TI - Fire eater's pneumonia: radiographic and CT findings. AB - Acute aspiration of petroleum by fire eaters can cause a distinct type of chemical pneumonitis known as fire eater's pneumonia that manifests on radiologic studies with unilateral or bilateral lung consolidations, well defined nodules, and pneumatoceles. We report three cases of fire eater's pneumonia that manifested with well-defined cavitary nodules (pneumatoceles) on radiographs and CT. One patient developed a bronchopleural fistula and spontaneous pyopneumothorax. CT is valuable for identifying and localizing complications to guide therapy. PMID- 10864085 TI - Breast involvement in a case of primary systemic amyloidosis: mammographic, US, and MR appearances. PMID- 10864086 TI - Comparison of thallium-201 and gallium-67 imaging in evaluation of head and neck tumors. AB - PURPOSE: 201Tl imaging was compared with 67Ga imaging in the detection of malignant head and neck tumors. METHOD: Eighteen patients with tumors in the head and neck region underwent both planar and SPECT imaging using 201Tl-chloride and 67Ga-citrate. The detection of primary tumors and lymph node (LN) metastases by visual evaluation was compared between 201Tl and 67Ga imaging. Additionally, quantitative analysis of the SPECT images was performed in eight patients. RESULTS: On visual evaluation, primary tumor uptake was detectable in 100% on the 201Tl SPECT images compared with 44% on the 67Ga SPECT images. The sensitivity, specificity, and accuracy of 201Tl in detecting LN metastases were higher than those of 67Ga SPECT; however, there was no statistical difference between 201Tl and 67Ga. The tumor-to-background ratio was significantly higher on 201Tl SPECT than on 67Ga SPECT. CONCLUSION: This study suggested that 201Tl could be useful in the evaluation of malignant tumors in the head and neck region. PMID- 10864087 TI - Severity of synovium and bone marrow abnormalities of the temporomandibular joint in early rheumatoid arthritis: role of gadolinium-enhanced fat-suppressed T1 weighted spin echo MRI. AB - PURPOSE: Our goal was to evaluate the efficacy of dynamic contrast-enhanced fat suppressed MRI of the temporomandibular joint (TMJ) in detecting early joint involvement in patients with rheumatoid arthritis (RA). METHOD: Conventional T1- and T2-weighted, gadolinium-enhanced T1-weighted, and dynamic gadolinium-enhanced fat-suppressed SE imaging sequences were performed in 22 patients with RA. RESULTS: The dynamic gadolinium-enhanced fat-suppressed T1-weighted SE sequence was more sensitive than the other techniques in detecting early changes in inflamed synovium of periarticular tissue and in detecting condylar bone marrow involvement. In patients with RA, 17 joints with joint pain showed synovial proliferation in 10 (59%) cases and joint effusion in 4 (24%). Of 14 joints with joint sound, 4 (29%) showed synovial proliferation and 7 (50%) showed joint effusion. A lower positional change of the disk was observed in joints with RA than in those with TMJ disorders (82 patients). CONCLUSION: Gadolinium-enhanced fat-suppressed MRI was extremely effective in diagnosing early changes of the inflamed TMJ. PMID- 10864088 TI - Estimation of brain compartment volume from MR Cavalieri slices. AB - PURPOSE: Recent theory has been developed to estimate volume from a systematic sample of tissue slices of a given thickness and to predict the corresponding error. Our goal was to check the error prediction formulas by resampling and to determine the minimum number of MR slices required to estimate the volumes of the cerebrum and of the compartments of gray matter (GM) and white matter (WM) with prescribed errors. METHOD: Our working data set comprised the GM and WM segmentations obtained from a paradigmatic high signal-to-noise ratio 3D spoiled GRASS MR volume data set for a single healthy human subject. The data were classified using a fuzzy clustering minimum distance algorithm. We thereby obtained a stack of 183 serial coronal slices of 1 mm thickness encompassing the whole cerebrum. Empirical resampling was carried out using the corresponding data vectors, and the theoretical error predictors were thereby checked for slice thicknesses of 1, 3, 9, and 27 mm, with a distance of 45 mm between slice midplanes. RESULTS: Irrespective of slice thickness, a minimum of 3, 5, and 10 slices provided estimates of the true total volume of GM and WM in the cerebrum with coefficients of error (CEs) of 10, 5, and 3%, respectively, where CE(V)% = 100 x SE(V)/V. For the cerebrum, a minimum of two, three, and four slices were required for CEs of the same precision. CONCLUSION: In combination with high signal-to-noise ratio and enhanced tissue contrast, Cavalieri slices are the most appropriate for MRI, they supply unbiased and highly efficient volume estimates of brain compartments. For a given number of slices, CE(V) decreases rapidly when the slices are thicker than the gaps between them; when the slices are thinner than the gaps, then CE(V) is similar to that in the situation when the slice thickness is zero. PMID- 10864089 TI - Time dependence of serial diffusion-weighted imaging features in a case of pyogenic brain abscess. AB - Both signal intensity on trace images and apparent diffusion coefficient measurements on mapped images evolved rapidly on serial diffusion-weighted sequences in a case of pyogenic brain abscess that was monitored primarily by MRI before a biopsy was performed. Considering only the signal intensities on the trace images would have led to an underestimation of the intrinsic tissue changes. PMID- 10864090 TI - Diffusion-weighted echo planar imaging of the normal human cervical spinal cord. AB - We obtained diffusion-weighted echo planar images of the human cervical cord in vivo and correlated them with histopathologic findings. Images were obtained in 17 healthy volunteers using a 1.5 T clinical MR unit. When motion-probing gradients were added perpendicular to the long axis of the cord, the white matter was hyperintense because of anisotropic diffusion. However, the gracile fasciculus was hypointense probably due to the small diameter of neural fibers and the large extracellular space. PMID- 10864091 TI - Virtual CT endoscopy in determining safe surgical entrance points for paranasal mucoceles. AB - The goal of this work was to evaluate virtual CT endoscopy for determining safe surgical entrance points for paranasal mucoceles. Twelve mucoceles in 11 cases were scanned with helical CT, and multiplanar reconstruction (MPR) and virtual endoscopic images were obtained. After a safe surgical entrance point was determined by MPR images, the entrance point was specified on the virtual endoscopic images. The combination of virtual endoscopic images and MPR images is a suitable method for determining a safe surgical entrance point for simple mucoceles. PMID- 10864092 TI - Normative measurements of orbital structures using MRI. AB - PURPOSE: The purpose of this work was to establish criteria for the diameters of normal extraocular muscles and superior ophthalmic vein and width of the optic nerve-sheath complex, to determine normal globe position as seen on MRI, and to investigate the effects of age and sex on these structures. METHOD: Diameters of the extraocular muscles and superior ophthalmic vein, width of the optic nerve sheath complex, and distance from the interzygomatic line to the posterior margin of the globe were calculated for 200 normal orbits of 100 patients on T1-weighted MR scans. RESULTS: Normal ranges for diameters of the extraocular muscles were as follows (mean +/- 2 SD): medial rectus, 3.2-4.9 mm; lateral rectus, 2.6-4.8 mm; inferior rectus, 3.7-6.0 mm; superior group, 3.1-5.6 mm; superior oblique, 2.4 4.1 mm. The normal position of the posterior pole of the globe was 8.9 mm behind the interzygomatic line (range 5.0-12.7 mm). The mean diameters of the extraocular muscles in male patients were significantly larger than those in female patients (p < 0.001). CONCLUSION: These data may be of value in quantitatively evaluating MRI of the orbit. PMID- 10864093 TI - The nomenclature and sectional imaging anatomy I: dorsal spinal ligaments. AB - This article is the first in a series of three that organizes the complex anatomy of the cervical, thoracic, and lumbar spinal ligaments. It describes and color codes the anatomy and nomenclature of the dorsal ligaments. The following articles will describe the ventral ligaments, and the capsular membranes and minor ligaments. PMID- 10864094 TI - Effect of calcium stress on the skeleton mass of intact and ovariectomized rats. AB - Female rats were ovariectomized (Ovx) or sham-operated (control) at 18 weeks and the entire skeleton obtained at 24 weeks (baseline) or after an additional 31 day (28 week) interval on a normal (1.0%) or deficient (0.02%) calcium diet. Ovx rats showed a 42% increase in whole body bone resorption (3H-tetracycline loss) in the absence of calcium stress (1.0% calcium diet) and a 70% increase in resorption with morphological evidence of dramatic loss of cancellous bone mass when placed on calcium-deficient (0.02%) diets. Ovx rats kept on the 1.0% calcium diet showed a significant increase in both their body weight (30.2%) and total bone mass (11.6%) compared to baseline sham-operated controls. However, the total skeleton mass of these animals was significantly reduced (-20%) from that predicted by calculations based on body weight. Maintaining animals on calcium-deficient diets had no significant effect on the total skeleton mass of either control or Ovx rats in comparison with age-matched controls on 1.0% diets. It was further determined that an increase in bone mass between 24 and 28 weeks in rats receiving 1.0% dietary calcium occurred in both the axial and appendicular skeleton and was proportionately similar between control and Ovx groups. However, in animals subjected to dietary calcium stress during this interval, the decreased skeletal growth noted was confined primarily to the axial skeleton. The data indicate that ovariectomy or ovariectomy plus calcium stress does not result in loss of total bone mass during the interval of dramatically increased resorption and rapid loss of cancellous bone. The results suggest that the deterioration in individual bone structural and mechanical integrity due to ovariectomy or dietary calcium deficiency may not be attributed to overt loss in total bone mass but may involve a redistribution of bone mass. PMID- 10864095 TI - In vitro relaxation of rabbit and human internal anal sphincter by rutaecarpine, an alkaloid isolated from Evodia rutaecarpa. AB - Rutaecarpine, a compound extracted from the Chinese medicinal herb Evodia rutaecarpa, has been shown to possess relaxing action on vascular smooth muscle from rat thoracic aorta. The internal anal sphincter is a specialized smooth muscle regulating important anorectal physiology. To investigate the effect and underlying mechanisms of rutaecarpine on internal anal sphincter, muscle strips from rabbit internal anal sphincter were used. The results showed that rutaecarpine (1 x 10(-10) M to 1 x 10(-4) M) produced a concentration-dependent muscular relaxation effect in our preparations, which were precontracted with acetylcholine. This muscular relaxation effect was not affected by treatment with L-N(G)-nitro-arginine methyl ester (a nitric oxide synthase inhibitor), methylene blue (a guanylate cyclase inhibitor), N-ethylmaleimide (an adenylate cyclase inhibitor), or by removal of the mucosa and submucosa tissue. Pretreatment with nifedipine (a calcium channel blocker) or extracellular Ca+2 removal by ethylenediaminetetraacetic acid (EDTA) greatly attenuated the relaxation effect, suggesting that calcium ion might be involved. In experiments using strips from human internal anal sphincter, an even more prominent relaxation effect was shown. It is thus concluded that rutaecarpine caused relaxation on internal anal sphincter from rabbits and human subjects. The relaxation action was not related to NO-cGMP pathway, instead calcium ion might play an important role and shed insight into clinical implications for those anorectal disorders with hyperactive anal tone. PMID- 10864097 TI - Electrophysiological effects of agmatine on human atrial fibers. AB - The objective of the present study was to study the electrophysiological effects of agmatine on human atrial fibers obtained at cardiac surgery using standard microelectrode techniques. Agmatine (1 to approximately 10 mM) decreased the action potential amplitude (APA), maximum upstroke velocity of phase 0 depolarization (Vmax), velocity of diastolic (phase 4) depolarization (VDD), rate of pacemaker firing (RPF), and action potential duration at 50 and 90% of repolarization (APD(50-90)) in a concentration-dependent manner. Pretreatment with N(G)-nitro-L-arginine methyl ester (L-NAME, 0.5 mM), a NOS inhibitor, did not affect the electrophysiological effects of agmatine (5 mM) on human atrial fibers. The effects of agmatine (5 mM) could be blocked completely by pretreatment with idazoxan (0.1 mM), an alpha-2 adrenergic receptor (alpha2-AR) and imidazoline receptor (IR) antagonist. All these results indicate that the effects of agmatine on human atrial fibers are likely due to a decrease of intracellular calcium mediated by IR and/or alpha2-AR. PMID- 10864096 TI - Gastric mucosal damage induced by arecoline seizure in rats. AB - In an attempt to know the relation of seizure and gastric mucosal damage, we challenged arecoline (ACL) centrally to induce seizure and investigated gastric hemorrhagic injury in acid-irrigated stomachs of rats. The protective effects of several drugs also were evaluated. After deprivation of food for 24 h, rats were received laparotomy under diethylether-anesthesia. Both pylorus sphincters and carotid esophagus were ligated. The forestomach was equipped with a cannula for gastric irrigation. After recovery from anesthesia (approximately 1 h), the stomach was irrigated for 2 h with an acid solution containing 100 mM HCl and 54 mM NaCl or the same volume of normal saline. Intracerebroventricular (i.c.v.) ACL (0, 1, 3 or 10 mg/kg dissolved in 10 microl of CSF) was challenged to rats immediately after gastric irrigation. The seizure in rats was produced by ACL in a dose-related manner. The ulcerogenic parameters such as decrease of gastric mucosal glutathione levels and increase of histamine concentrations and lipid peroxide generations as well as the raise of luminal hemoglobin contents and exacerbated mucosal lesions were obtained depending on the doses of ACL challenged. These ulcerogenic parameters produced in ACL (10 mg/kg, i.c.v.) seizure rats were markedly ameliorated by gastric vagotomy or central anticholinergics. Intraperitoneal ketotifen, zinc sulfate, diphenhydramine or cimetidine also produced significant (p<0.05) inhibitions of these ulcerogenic parameters in ACL seizure rats. In conclusion, central ACL seizure may produce gastric oxidative stress and hemorrhagic lesions via vagal nervous activation and histamine release in acid-irrigated stomachs of rats. PMID- 10864098 TI - Chronic cocaine alters hemodynamics and leukocyte-endothelial interactions in rat mesenteric venules. AB - We investigated the effects of chronic cocaine exposure on the microcirculation in the rat mesenteric venules under both non-inflammatory and FMLP-induced inflammatory conditions. Chronic cocaine significantly increased WBC rolling flux in both conditions, and potentiated FMLP-induced leukocyte-endothelial cell adhesion (LEA). In cocaine-treated animals, total WBC number increased by 91%, and the ratio of white blood cell to red blood cell velocity was significantly lower, while vessel diameter was unchanged. Chronic cocaine decreased serum levels of tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6), but had no effect on interleukin-1 beta (IL-1beta). Expression of intercellular adhesion molecule-1 (ICAM-1) was increased in mesenteric venules following chronic cocaine exposure, and may be one of the mechanisms underlying enhancement of FMLP-induced LEA. The increase in WBC count, WBC flux and LEA, and the change in cell velocity seen in the cocaine-treated animals could cause a decrease in effective vessel diameter and a change in intravascular resistance, and may underlie the progressive vascular damage seen in chronic cocaine-abusing individuals. PMID- 10864099 TI - Age-related arterial calcification in rats. AB - In man, i) arteries calcify with age and ii) age-linked arterial calcification is amplified by vascular pathology such as hypertension or arteriosclerosis. Age linked arterial calcification has a bad prognosis but drugs to prevent it are lacking. This is partially due to the lack of appropriate animal models. This paper looks at the extent to which arteries calcify with age in the rat and whether hypertension or arteriosclerosis amplifies such calcification. Total calcium levels were determined by acid digestion and flame spectrophotometry and intracellular calcium levels ([Ca2+]i) by the intracellular calcium-sensitive dye, fura-2. Arteries contained up to 5 times more calcium than other soft tissues. Arteries progressively calcified with age whereas other soft tissues did not. Accumulation of calcium with age was essentially extracellular. Hypertension had no effect on age-related arterial calcification. Calcification of the same order as in man was produced in a rat model of arteriosclerosis (vitamin D plus nicotine treatment). In conclusion, as in man, age-linked, organ-specific arterial calcification does occur in rats but its intensity is far less. Arterial calcification of a similar degree to that observed in man can be obtained in rats by hypervitaminosis D plus nicotine. PMID- 10864100 TI - Plasma epinephrine levels in hypertension and across gender and ethnicity. AB - Epinephrine (E) infusions raise blood pressure and there is an excess incidence of hypertension among males and blacks. However, reports of E levels by ethnicity, gender, and blood pressure status are inconsistent. Insensitive assays, variability in plasma E levels within individuals, and the small size of most studies have contributed to these conflicting reports. We measured plasma E levels in a large diverse sample of subjects, using a highly sensitive assay. A total of 361 individuals participated in the study: 61% were men and 39% women, 74% were normotensive and 26% hypertensive, 59% were white and 41% were black. Except for difference in blood pressure and body mass index between the normotensives and hypertensives, subjects had similar baseline characteristics and took no antihypertensive medications for at least five days prior to sampling. All blood samples were collected after resting for a least 30 minutes following the insertion of an indwelling i.v. catheter. Catecholamine levels were determined using a radioenzymatic assay (assay sensitivities for E and norepinephrine were 6 pg/ml and 10 pg/ml, respectively). An ethnicity by gender interaction was found (F(1,315) = 5.126, p = .024). Subsequent analysis revealed that white women had significantly lower basal plasma E levels than white men (p <0.001) and black women (p = 0.036). There were no significant differences in E levels between black men and women or between white men and black men. Uncorrected E levels were lower in normotensive than hypertensive subjects (p = .009) but this difference was not significant when corrected for body mass index (BMI). Uncorrected norepinephrine levels were higher in women than men (p = .03) but the difference was no longer significant when corrected for BMI. Plasma E levels were significantly lower among white women than men or black women. In contrast to prior studies, E levels were lower in hypertensives, but this may reflect obesity among hypertensives. PMID- 10864101 TI - Antinociceptive effects of morphine-6-glucuronide in homozygous MDR1a P glycoprotein knockout and in wildtype mice in the hotplate test. AB - Morphine-6-glucuronide (M6G), a major metabolite of morphine with agonist opioid receptor activity, was reported to be a substrate of P-glycoprotein (P-gp). Inhibition of P-gp may thus result in higher brain uptake of M6G. The goal of this observer-blinded, placebo controlled study, was to compare the antinociceptive effects of M6G in homozygous P-gp knockout (mdr1a(-/-)) and wildtype (mdr1a(+/+)) mice. M6G was injected intraperitoneally as a single dose of 0, 0.5, 1, 2.5, 5, and 10 mg/kg. Eight P-gp knockout and eight wildtype mice were studied per dose. A hot plate test was performed before and 5, 15, 30, 60, 90, 120, and 150 min after M6G administration. Plasma-concentrations of M6G, morphine, and morphine-3-glucuronide (M3G) were measured after intraperitoneal injection of 5 mg/kg M6G in another 14 P-gp knockout and 14 wildtype mice. No difference neither in the dose response relationship, nor in the time course of response latency times were observed between P-gp knockout and wildtype mice. However, latency times increased with higher doses of M6G, with antinociception significantly different from placebo at a M6G dose of 5 and 10 mg/kg. P-gp knockout mice tended to have higher plasma concentrations than the wildtype. However, plasma concentrations widely overlapped between groups and therefore no statistical significant group difference could be detected. We conclude that despite reported doubling of M6G brain uptake, absence of mdr1a coded P-gp does not enhance antinociceptive effects of M6G in the hotplate test after acute single-dose administration in mdr1a(-/-) knockout mice. PMID- 10864102 TI - Comparison of therapeutic regimens in the amelioration of alterations in rat gastrointestinal mucosal DNA, RNA and protein induced by streptozotocin diabetes mellitus. AB - Type 1 diabetes mellitus is characterized by hyperglycemia, insulinopenia, and secondary neural, renal and vascular complications. Clinical manifestations in the gastrointestinal tract range from initial mild complications to more severe complications as the disease progresses, but as of yet, are poorly understood. The current study has two main foci 1) to monitor the alterations in gastrointestinal DNA, RNA and protein content induced by streptozotocin diabetes and 2) to use these parameters to monitor the efficacy of intensive insulin treatment versus pancreatic islet transplantation in the amelioration of the diabetes induced alterations. Female Wistar Furth rats were rendered diabetic by streptozotocin injection and measured for alterations in gastrointestinal DNA, RNA and protein content. Similarly, animals which had streptozotocin-induced diabetes were also treated by intensive insulin therapy or pancreatic islet transplant and monitored for alterations in gastrointestinal DNA, RNA and protein content. In general, diabetes induced increases in stomach, duodenal, jejunal and colonic macromolecular content. With few exceptions, treatment with either intensive insulin or pancreatic islet transplantation returned each variable measured back to control levels. In every case, pancreatic islet transplantation was comparable to intensive insulin therapy. In the short term the treatments are comparable, but long term analyses are needed to determine if the treatments offer any difference in their ability to prevent the long term complications related to diabetes mellitus. PMID- 10864103 TI - The role of 5-hydroxytryptamine3 and 5-hydroxytryptamine4 receptors in the regulation of gut motility in the ferret. AB - The role of 5-hydroxytryptamine (5-HT)3 and 5-HT4 receptors in the regulation of gut motility in the ferret was investigated. The selective 5-HT3 receptor antagonist ramosetron (1 - 10 microg/kg s.c.) prolonged the interval of gastric antral migrating motor complex, but had only slight effect on small intestinal and colonic motility in unfed animals. The selective 5-HT4 receptor antagonist SB 204070 did not affect motility throughout gut in unfed animals. Neither ramosetron nor SB 204070 affected the motility throughout gut in fed animals. In conclusion, neither 5-HT3 nor 5-HT4 receptors tonically regulate ferret gut motility except that 5-HT3 receptors have a key role in the occurrence of migrating motor complex specifically in the stomach. The role of 5-HT3 and 5-HT4 receptor system in the regulation of gut motility in ferrets is similar to that in other mammalian species studied, including humans. This similarity suggests that the ferret is a suitable model animal to study gut motor functions in humans. PMID- 10864104 TI - Lack of melatonin effect on hydrogen peroxide induced bronchoconstriction in isolated and perfused rat lung. AB - The effect of melatonin on hydrogen peroxide- induced broncho-and vasoconstriction was examined in vivo in the model of the isolated, perfused and ventilated lung. The administration of hydrogen peroxide (500 microM) to the perfusate caused a marked decrease in lung compliance, conductance and flow rate. The administration of melatonin (500 microM) to the perfusate 20 min before and during the hydroperoxide exposure did not cause any change in lung function. Exposure of lung microsomes to hydrogen peroxide (1-100 microM) did not induce any significant increase in malonaldehyde (MDA), an index of lipid peroxidation, and it was not affected by treatment with melatonin (500 microM). On the other hand, brain microsomes exposed to hydrogen peroxide (1-100 microM) give rise to increased levels of MDA, which were decreased by pre-treatment with melatonin (500 microM). The results suggest that melatonin may exert an antioxidant effect in conditions were lipid peroxidation is occurring. Its use may not be relevant in conditions where the mechanisms of the reactive oxygen species damage appears to be lipid peroxidation independent, such as the case of hydrogen peroxide induced broncho- and vasoconstriction. PMID- 10864105 TI - Functional speech outcomes after laryngectomy and pharyngolaryngectomy. AB - OBJECTIVE: To compare and contrast functional speech outcomes of patients having undergone total laryngectomy and pharyngolaryngectomy who use tracheoesophageal speech as their primary mode of communication. DESIGN: Group comparison design. SETTING: Adult acute tertiary care hospital. PATIENTS: Thirty patients who underwent total laryngectomy and 13 who underwent pharyngolaryngectomy with free jejunal interposition reconstruction. All patients used tracheoesophageal speech. INTERVENTION: Group comparisons across measures of speech intelligibility, voice quality, tracheoesophageal speech use, voice satisfaction and levels of perceived voice disability, handicap, and well-being/distress. MAIN OUTCOME MEASURE: The existence of any significant differences between the 2 groups on measures of intelligibility, voice quality, tracheoesophageal speech use, and voice satisfaction and levels of voice disability, handicap, and well-being/distress. RESULTS: Statistical comparisons confirmed reduced functional intelligibility (P<.05), reduced vocal quality (P<.01), and higher levels of disability (P<.05) in the pharyngolaryngectomy group. However, no significant difference was observed between the proportion of patients classified as "successful" tracheoesophageal speech users in either group. Low levels of handicap and high levels of patient well-being were recorded in both groups. CONCLUSION: Despite the perceptual differences in voice quality and intelligibility observed between the 2 groups, tracheoesophageal speech that is functional, effective, and perceived by the patients as satisfactory can be achieved after total laryngectomy and pharyngolaryngectomy with free jejunal interposition reconstruction. PMID- 10864106 TI - Meta-analysis in otolaryngology. AB - OBJECTIVE: To examine the results of meta-analyses in otolaryngology and compare these results with the individual component studies that constitute each meta analysis. DESIGN: A retrospective review of the literature. MAIN OUTCOME MEASURES: Studies that conducted pooled statistical systematic analyses indexed on MEDLINE for the 10-year period from January 1989 to January 1999 were selected for keyword or subject headings of meta-analysis and otolaryngology (N = 22). Analysis consisted of a modified funnel graph depiction of the individual studies that made up each meta-analysis. Each meta-analysis was evaluated for consistency among these individual studies and comparison of the median result with the weighted mean meta-analysis result. In addition, the methodologic quality of each meta-analysis was assessed in terms of the rigor with which component studies were evaluated. RESULTS: Ten (46%) of the 22 meta-analyses did not provide the individual study results that made up their meta-analyses. The results of 10 studies (46%) were similar to the median result of their individual component studies. The results of 2 studies (9%) differed from this median result, with widely heterogeneous component study results. CONCLUSIONS: A large proportion of meta-analyses in otolaryngology (46%) fail to provide the individual study results necessary to analyze the meta-analysis result critically. Most remaining studies do provide results that accurately compare with the median of their component study results. Only a small proportion of meta-analyses were found to have disparate results, and each appropriately discusses the heterogeneity of the individual studies that comprise their meta-analysis. PMID- 10864107 TI - Efficacy of postoperative follow-up telephone calls for patients who underwent adenotonsillectomy. AB - OBJECTIVE: To evaluate the efficacy and cost-effectiveness of postoperative follow-up telephone calls among pediatric patients who underwent adenotonsillectomy. DESIGN: Prospective study with a follow-up questionnaire administered by telephone. SETTING: Tertiary-care children's hospital. PATIENTS: One hundred thirty-four children between the ages of 4 and 18 years who underwent adenotonsillectomy between December 1997 and June 1998 and did not have associated cardiac, pulmonary, bleeding, or syndromic disorders were included in this pilot study. INTERVENTION: Parents of these patients were given the opportunity to participate in our study, and it was emphasized that, at any time during the child's care, if the parent desired a follow-up visit or if the child experienced any symptoms that caused concern, the parent should contact the clinic for a follow-up appointment. A telephone call was placed 3 to 4 weeks postoperatively by an otolaryngology nurse, and a questionnaire was filled out using the parents' responses. MAIN OUTCOME MEASURES: The incidence rates of voice change, velopharyngeal insufficiency, bleeding, constipation, dehydration, and pain were measured. Parent satisfaction, patient safety, and cost-benefit were also evaluated. RESULTS: Less than 5% of patients reported temporary velopharyngeal insufficiency, while 2% of patients required operative intervention for bleeding episodes and 1% required hospitalization. Voice change, reported by approximately 70% of all patients, was the most common complaint, but it resolved in all instances. Pain was reported to be most severe on postoperative day 1. Ninety-six percent of parents requested no further follow-up visit. CONCLUSIONS: Our pilot study revealed that a follow-up telephone call is a safe and cost-effective method of postoperative management for pediatric patients who have undergone adenotonsillectomy and that this method of follow-up is also desirable to parents. PMID- 10864108 TI - Immunolocalization of activated transforming growth factor beta1 in juvenile nasopharyngeal angiofibroma. AB - BACKGROUND: Juvenile nasopharyngeal angiofibroma (JNA) is a histologically benign, locally aggressive neoplasm of the nasopharynx that exclusively affects male adolescents. It is known to be sensitive to androgens, but there are likely intermediary cytokines and/or growth factors that mediate aggressive stromal cell proliferation and angiogenesis. Transforming growth factor beta1 (TGF-beta1) is a polypeptide that is secreted in an inactive form, cleaved to produce an active form, and then deactivated in the tissues. It activates fibroblast proliferation and is known to induce angiogenesis. OBJECTIVES: To evaluate the presence of activated TGF-beta1 within the stroma of JNA specimens and to quantify the percentage of JNA specimens expressing the active growth factor. DESIGN: Immunohistochemical analysis was performed on 19 specimens of JNA using a unique antibody that identifies only the activated form of TGF-beta1. The percentage of cells staining positively for activated TGF-beta1 was determined semiquantitatively by visual methods. RESULTS: Of 19 cases stained, all 19 (100%) showed strong positive staining (2 cases with 33%-66% of cells staining and 17 with 66%-100% of cells staining). Activated TGF-beta1 was identified in stromal cell nuclei and cytoplasm and in the endothelium of the capillaries within all specimens of JNA. CONCLUSIONS: The localization of activated TGF-beta1 to the fibroblasts and endothelial cells within JNA tumors suggests that TGF-beta1 may play a role in the stromal cell proliferation and angiogenesis associated with JNA. Additional receptor studies and more quantitative methods of analysis are needed to further define the role of TGF-beta1 in the pathogenesis of JNA. PMID- 10864109 TI - Rehabilitation of olfaction after laryngectomy by means of a nasal airflow inducing maneuver: the "polite yawning" technique. AB - OBJECTIVE: To develop a nasal airflow-inducing maneuver and apply it in the olfactory rehabilitation of patients who have undergone laryngectomy. DESIGN: Intervention study; before-and-after trial. SETTING: National cancer center. PATIENTS: Forty-four patients who underwent laryngectomy; 34 men and 10 women; mean age, 64 years (range, 42-80 years); mean time since surgery, 6 years (range, 8 months to 18 years). INTERVENTION: In a prospective clinical intervention study, we assessed the effectiveness of a nasal airflow-inducing maneuver ("polite yawning," ie, yawning with closed lips). Speech therapists trained the patients in the maneuver, and its effectiveness in inducing nasal airflow was checked with digital and water manometers. MAIN OUTCOME MEASURES: Olfactory acuity was assessed before and after the intervention by means of an odor detection test and a structured questionnaire concerning olfaction, taste, and appetite. Patients were categorized as "smellers" and "nonsmellers" on the basis of the results of the odor detection test and the present odor perception scale derived from the questionnaire. RESULTS: The nasal airflow-inducing maneuver could be taught to all patients, mostly in only one 30-minute therapy session. Fifteen of the 33 patients in the pretreatment nonsmeller category converted to smellers, for a success rate of 46% (P<.001). CONCLUSION: The nasal airflow inducing maneuver (the "polite yawning" technique) allowed almost half of the patients to recover their sense of smell. PMID- 10864110 TI - Late effects of topical anesthetics on the healing of guinea pig tympanic membranes after myringotomy. AB - BACKGROUND: The optimal local anesthetic for myringotomies or the insertion of tympanostomy tubes in adults should be easy and rapid to use, be painless during application, provide good anesthesia, be reversible, be inexpensive, and not cause any long-term damage to the tympanic membrane (TM). OBJECTIVE: To evaluate the histologic effects of topical anesthetic agents on the healing of the TM after myringotomy. METHODS: Sixty male albino guinea pigs were randomly assigned to 1 of 5 groups. Of the 5 groups, 2 were used as controls: one group underwent a myringotomy and the other group did not. The remaining 3 groups had both TMs treated with a topical anesthetic (phenol, tetracaine base, and eutectic mixture of lidocaine and prilocaine in a cream) prior to myringotomy. All TMs were inspected periodically and then harvested at 3 months or 6 months postoperatively for histologic examination. RESULTS: The TMs of the group treated with tetracaine appeared the most normal at 6 months (P=.001). However, histologic evaluation failed to demonstrate any significant differences in the thickness of the TM or the lamina propria (P=.45), the amount (P=.80) and orientation (P=.07) of collagen, or the number of infiltrating lymphocytes (P=.70). CONCLUSION: Based on the histologic findings, all 3 topical anesthetic agents appear to cause equivalent changes to the TM when used for a myringotomy. PMID- 10864111 TI - Radiofrequency treatment for obstructive tonsillar hypertrophy. AB - OBJECTIVE: To evaluate the safety and efficacy of in-office, temperature controlled radiofrequency submucosal tissue volume reduction using the Somnoplasty procedure for the treatment of symptomatic chronic obstructive tonsillar hypertrophy. DESIGN: A prospective, nonrandomized, 3-phase protocol using in vitro and in vivo studies associated with operative tonsillectomy and clinical procedures performed in-office. SETTING: Hospital operating room and private practice otolaryngology office. STUDY POPULATION: In vitro studies of 14 tonsil specimens following tonsillectomy; in vivo studies of 11 tonsils before tonsillectomy; and clinical procedures performed on 9 adults, ages 24 to 47 years, with symptomatic chronic tonsillar hypertrophy. OUTCOME MEASURES: For phase 1, histologic tissue sections; for phase 2, histologic tissue sections and clinician and patient questionnaires regarding procedure morbidity; and for phase 3, measurements of oropharyngeal airway size and clinician and patient questionnaires regarding procedure morbidity and symptom improvement. RESULTS: A 2-needle radiofrequency probe ablated tonsil stromal tissue while leaving overlying mucosa and underlying structures intact. On average, oropharyngeal airway was enlarged 12 mm, with a 70.8% calculated reduction in tonsil size. Procedures were well tolerated and had only minimal pain and dysphagia. There were no episodes of hemorrhage, and patients resumed normal activity within 1 to 2 days. Substantial improvement was reported in daytime sleepiness, snoring, voice clarity, swallowing, and throat irritation. CONCLUSIONS: Temperature controlled radiofrequency submucosal tissue volume reduction is a safe and effective method of treating symptomatic obstructive tonsillar hypertrophy. It is well tolerated by the patient under local anesthesia in the physician's office and has minimal postprocedure pain and dysphagia, with rapid return to normal activity. The procedure reduces tonsil size and increases airway size, leading to a reduction in symptoms. PMID- 10864112 TI - Location and gross morphology of the nasopalatine duct in human adults. AB - OBJECTIVE: To characterize the nasal opening of the human nasopalatine duct (NPD; a major duct of the vomeronasal system used in animals for chemical communication) by identifying its location, categorizing variations in physical characteristics, and assessing frequency of detection. DESIGN: Two studies: (1) general study incorporating endoscopic examinations documented by videotapes and photographs, and an investigation of detection bias in terms of method of visualization and defining criteria for NPD identification; and (2) cadaver dissections examining NPD gross anatomy and proximity to the putative vomeronasal organ (VNO), the second major duct of the vomeronasal system. SETTING: Department of Otolaryngology, University of Chicago Hospitals, Chicago, Ill. SUBJECTS: A total of 125 university community volunteers, with a mean age of 23 years. RESULTS: (1) General study: NPD was located 1.9+/-0.02 cm (mean+/-SEM) dorsal to the columella nasi, and 0.2+/-0.01 cm above the nasal floor/septum junction, in both nostrils (90% bilateral), and was symmetrical in shape. An NPD was detected in 94% of 221 nostrils unobstructed in the region of interest. Fossa shapes were oval (57%), round (18%), spindle-shaped (18%), and tubular (7%). A small, round aperture was visualized in 30% of fossae. Among 3 observers, NPD detection frequency ranged from 94% to 40%, with the disparity due to inclusion of different defining characteristics. (2) Cadaver dissection study: bilateral nasal NPD fossae were found in every specimen probed to maximum approximate depth of 8 mm. No buccal pits associated with patent NPD were detected. Putative VNOs superior and just anterior to NPD fossae were detected in fewer than half the specimens. CONCLUSIONS: The nasal opening of the NPD is bilateral and symmetrical, located at the base of the nasal septum. Unusually contradictory anatomical descriptions in the human putative VNO literature may be attributable to inexact descriptions or misidentification of structures. The function of NPD remains controversial. PMID- 10864113 TI - Hot, humid air partially inhibits the nasal response to allergen provocation. AB - BACKGROUND: We have previously reported that preconditioning allergic subjects with hot, humid air (HHA) (temperature, 37 degrees C; relative humidity >95%) in an environmental chamber resulted in partial inhibition of the early response to nasal allergen challenge. OBJECTIVE: To investigate whether this inhibitory effect could be achieved by inhalation of HHA via a face mask. DESIGN: Randomized, 4-way crossover study. SUBJECTS: Eighteen subjects with seasonal allergic rhinitis participated in the study outside of their allergy season. INTERVENTIONS: Subjects underwent preconditioning with room air (RA) (temperature, 25 degrees C; relative humidity <20%) or HHA either in a chamber or delivered via a face mask for 1 hour prior to and during nasal challenge with diluent for the allergen extract followed by 2 increasing doses of allergen. RESULTS: Net changes from diluent challenge for all parameters were compared between HHA and RA in each delivery method. Hot, humid air delivered by mask significantly inhibited the mean+/-SEM number of allergen-induced sneezes (HHA, 2.7+/-0.6; RA, 6.6+/-2.1; P=.03), congestion score (HHA, 2.3+/-0.5; RA, 3.4+/ 0.5; P=.01), and secretion weights (HHA, 26.9+/-4.4 mg; RA, 38.6+/-5.0 mg; P=.048). However, HHA inhaled in a chamber significantly inhibited only the mean+/-SEM allergen-induced congestion (HHA, 1.2+/-0.4; RA, 3.6+/-0.6; P=.002) and pruritus (HHA, 0.7+/-0.3; RA, 2.3+/-0.5; P=.002) scores. CONCLUSIONS: Preconditioning the nasal mucosa with HHA partially decreases the early response to nasal challenge with antigen irrespective of the administration technique. The secretory response, however, is only inhibited by localized delivery of HHA to the nose. The inhibitory effects of HHA are therefore probably related to local changes in the nasal mucosa and are not dependent on total body exposure to HHA. PMID- 10864114 TI - Growth of tissue-engineered human nasoseptal cartilage in simulated microgravity. AB - OBJECTIVE: To evaluate the feasibility of in vitro fabrication of tissue engineered cartilage from human nasoseptal chondrocytes for autologous reconstruction. DESIGN: Hyaline cartilage was reconstituted from chondrocyte polyglycolic acid scaffolding constructs in a 3-dimensional mammalian cell culture cascade. This included monolayer cellular amplification, cell seeding in the spinner flask, and tissue growth in simulated microgravity. RESULTS: The quality of the fabricated cartilage analogue was found to depend on the initial cell density, duration of incubation, and bioreactor type. Dynamic seeding was nearly completed within the first 10 hours of inoculation regardless of the cell source (cryogenically preserved vs fresh chondrocytes) or presence of serum. A duration of incubation in excess of 4 weeks was required for complete matrix biosynthesis at low seeding densities in the spinner flasks. Seeding densities greater than 2.3x10(6) chondrocytes per scaffold were required for early hyaline cartilage formation as well as longer-time mature matrix regeneration. In addition, maintaining the structural integrity of the unreinforced scaffold, which is necessary for continued mature matrix regeneration, was achievable through postseeding tissue growth in simulated microgravity. CONCLUSION: Once combined with polyglycolic acid scaffolds in the bioreactor cascades that allow efficient seeding and quiescent tissue growth, human septal chondrocytes become a valuable source of reproducible ex vivo cartilage regeneration in the laboratory. PMID- 10864115 TI - Glycohistochemical characteristics of nasal polyps from patients with and without cystic fibrosis. AB - OBJECTIVE: To investigate whether cystic fibrosis (CF)-related nasal polyps exhibit significantly distinct glycohistochemical characteristics when compared with single vs massive nasal polyps obtained from patients without CF. DESIGN: Glycohistochemical characteristics were identified by means of 8 histochemical probes, including 5 plant lectins (peanut, gorse seed, wheat germ, Maackia amurensis, and Sambucus nigra agglutinins), 2 animal lectins (14- and 16-kd galectins), and 1 neoglycoprotein (exposing the Thomsen-Friedenreich antigen). The binding of the 8 glycohistochemical markers was determined by means of computer-assisted microscopy. For each probe, 3 quantitative parameters were computed: the labeling index, which describes the percentage of tissue area specifically stained by a given marker; the mean optical density, which reflects the staining intensity; and the concentrational heterogeneity, which characterizes the level of heterogeneity of the staining intensity. SUBJECTS: A series of 61 nasal mucosa specimens was analyzed, including 6 normal cases, 23 single and 18 massive polyposis cases without CF, and 14 nasal polyps associated with CF. RESULTS: Normal and polyposal nasal mucosa differed in terms of the amounts and linkage types of sialic acids (revealed by the wheat germ, M amurensis, and S nigra agglutinins) rather than the characteristics of galactoside expression (monitored with the peanut agglutinin and 2 animal galectins). In contrast, nasal polyps markedly differed between patients with and without CF with respect to galactoside expression (revealed by the peanut agglutinin and the 14-kd galectin) and the display of binding site(s) for the neoglycoprotein. CONCLUSION: Normal and polyposal nasal mucosa differ essentially in sialic acid presentation, while nasal polyps from patients with CF have a higher level of various lectin-reactive galactoside residues than nasal polyps from those without CF. PMID- 10864116 TI - Isolated sphenoid sinus diseases: report of 39 cases. AB - OBJECTIVE: To detail the underlying pathological conditions, symptoms, signs, and outcomes of patients with isolated sphenoid sinus involvement. DESIGN: A retrospective survey. SETTING: An academic referral center of a university hospital. PATIENTS: All 39 patients, aged 7 to 85 years, treated in the Department of Otorhinolaryngology, Kuopio University Hospital, Kuopio, Finland, from 1988 through 1997 for isolated sphenoid sinus disease. RESULTS: Sinusitis was characterized as acute in 26 patients, subacute in 5 (including 1 pyocele), and chronic in 8 (including 2 fungal infections). No tumors were found. Isolated sinus cysts were excluded from the study. Headache, the main symptom in 32 patients (82%), was localized most commonly on the vertex. Other common complaints were rhinitis, dizziness, eye symptoms, and fever. In 2 patients, the finding was occult. Eight patients (21%) presented with cranial nerve deficits, and 1 patient had an intracranial complication. Sinus irrigation was performed in 16 patients (41%) and sphenoidotomy was performed in 10 (26%). Fifteen patients (38%) were treated with antibiotic drugs alone. Within 3 months, 31 (84%) of 37 patients had recovered from the illness; 5 still experienced headaches despite having normalized radiographic findings; and 1 had permanent unilateral visual loss. Two patients were lost to follow-up. CONCLUSIONS: Sphenoid sinus opacity is mostly inflammatory in origin. Despite the benign nature of the disease, there is a risk of complications with high morbidity and mortality. Early and, if necessary, aggressive therapy to guarantee drainage of the sinus is recommended. PMID- 10864117 TI - A case report of FSH-producing nasal ectopic pituitary adenoma extending to the frontal cranial fossa. AB - We report the first case of an ectopic pituitary adenoma in the nasal cavity that produced follicle-stimulating hormone (FSH). A 60-year-old man complaining of left nasal bleeding had a polypoid tumor in the left nasal cavity. Findings of computed tomographic scanning and magnetic resonance imaging showed that the tumor originated from the olfactory cleft, occupied the nasal cavity, and extended to the frontal cranial fossa. Results of histologic examination suggested ectopic pituitary adenoma. Magnetic resonance imaging results showed the pituitary gland to be normal. Electron microscopy findings demonstrated a large number of secretory granules in the tumor cells that were positive for FSH on immunohistochemical analyses. Serum gonadotropin levels were normal, and no clinical signs of hypersecretory syndrome were noted. The above findings led us to establish the diagnosis of FSH-producing ectopic pituitary adenoma. The patient underwent craniofacial resection of the tumor followed by an uneventful recovery. The pathologic findings and clinical course of the case were comparable to those of FSH-producing adenomas arising from the pituitary gland. PMID- 10864118 TI - The difficulties of diagnosing myositis ossificans circumscripta in the paraspinal muscles of a human immunodeficiency virus-positive man: magnetic resonance imaging and temporal computed tomographic findings. AB - Myositis ossificans circumscripta (MOC) is an uncommon disease that has been reported to develop after trauma, burns, and infections, as well as in cases in which the pathogenesis is unknown. Although most cases of MOC develop below the clavicles, it has been reported in the head and neck area, where it primarily involves the muscles of mastication. To our knowledge, there have been only 4 previous reports of MOC affecting the paraspinal muscles. We report the fifth such case, which developed in a human immunodeficiency virus-positive patient, and describe the longitudinal imaging changes of MOC. PMID- 10864119 TI - An unusual source of Vibrio alginolyticus-associated otitis: prolonged colonization or freshwater exposure? AB - Otitis externa and otitis media are commonly encountered in clinical practice. We report an unusual case of otitis externa, which was caused by Vibrio alginolyticus, several months after saltwater exposure. Clinicians need to be aware of this unusual pathogen, especially in refractory cases of ear infections. PMID- 10864120 TI - Imaging quiz case 1. Mycotic aneurysm of the carotid artery. PMID- 10864121 TI - Imaging quiz case 2. First branchial cleft cyst (FBCC). PMID- 10864122 TI - Imaging quiz case 3. Common cavity, a congenital deformity of the inner ear. PMID- 10864123 TI - Imaging quiz case 4. Bifid mandibular condyle. PMID- 10864124 TI - The evaluation and management of olfactory disorder following upper respiratory tract infection. PMID- 10864125 TI - The failure of cricopharyngeal myotomy to improve postoperative dysphagia: is videofluoroscopic diagnosis adequate? PMID- 10864126 TI - Mal de debarquement. PMID- 10864127 TI - A criterion for distinguishing level V nodes from clavicular nodes. PMID- 10864128 TI - Statistical methods in epidemiology. IV. Confounding and the matched pairs odds ratio. AB - PURPOSE: This paper introduces readers to the problem of confounding in epidemiology, how it differs from effect modification and how to deal with it statistically. There are several options for dealing with confounding. These include techniques based on matching, on stratification and on regression. This paper reviews the first method, matching and the matched pairs odds ratio. METHOD: For 1:1 pair matching (1 case and 1 matched control), the matched pairs odds ratio is introduced as the ratio of discordant pairs. A method for calculating confidence intervals based on the normal approximation is described. RESULTS: Some properties of the matched pair design are illustrated by taking examples from the authors' own teaching experiences. CONCLUSION: Matching remains a difficult design option in epidemiology. Its 'best' use is for special types of studies such as for those on twin pairs. PMID- 10864129 TI - Integrated institution--community rehabilitation in developed countries: a proposal. AB - PURPOSE: To propose a system for the provision of comprehensive, coordinated rehabilitation services that would meet all the needs of persons with disability in a timely and cost-effective manner. METHODS: Study of the literature pertaining to features of settings available for the delivery of medical rehabilitation in developed countries; presentation of the evolution of a tertiary rehabilitation centre into an institution practicing community-oriented rehabilitation. Review of various issues and implications of integrating institutional-based and community-based rehabilitation. RESULTS: Rehabilitation settings differ in skills and resources and consequently, in the treatment, care and concern they are able to offer. It is essential to find the balance between medical, nursing and social needs of persons with disability and their requirements for skills and resources at a given time, and to provide rehabilitation, support and guidance in the setting most appropriate to these requirements and needs at the lowest cost possible. CONCLUSION: The integration of the rehabilitation institution of a region with secondary and primary care of the region, into one functional entity for the purposes of providing the needed services, would enable finding the most appropriate setting, and facilitate addressing all needs, as well as increase the availability and accessibility of comprehensive rehabilitation at an affordable cost. This could be a viable way of providing rehabilitation in developed countries of Europe, where the need for it is expected to rise in excess of the population increase. PMID- 10864130 TI - An assessment of gait analysis in the rehabilitation of children with walking difficulties. AB - PURPOSE: To assess the current status of computerized gait analysis techniques in the management of children with cerebral palsy or spina bifida who have significant walking disorders. METHOD: Synthesis of available data from a review of the literature, drawing on MEDLINE, EMBASE, PRE-MEDLINE, HealthStar and PsychInfo. Other information was obtained from persons with expertise in computerized gait analysis. Cost data were obtained from Canadian rehabilitation centres and the provincial health ministry. RESULTS: This technology seems helpful in detecting gait changes. However, available evidence is insufficient to draw conclusions about the influence of computerized gait analysis on treatment outcomes. Part of the rationale for use of the technology is that costs of gait analysis (of the order of $ CAN 2,000 per examination) would be offset by a decrease in follow-up surgical procedures and associated hospital care. There could also be a major influence on children's independence and quality of life. However, there are as yet no convincing data to support these propositions. CONCLUSIONS: Computerized gait analysis is a potentially useful technology in the management of children with walking disabilities, but its efficacy is not established. It should be regarded as a developing technology and its clinical application linked to systematic collection and assessment of outcomes data. PMID- 10864132 TI - The social impact of multiple sclerosis--a study of 305 patients and their relatives. AB - PURPOSE: to assess the effects of multiple sclerosis (MS) on the patients' ability to fulfil their chosen family and social roles and to examine the impact of the disease on their relatives. METHODS: a population-based survey of all known patients with MS and their relatives in Hampshire County, England, between 1986 and 1989. RESULTS: Seventy-four% of the total study population of 411 completed the study. The patients' mean age was 48.3 years (range 19-82) and the mean disease duration was 15.8 years. About 16% of patients were depressed on a mood rating scale and a similar number also exhibited symptoms of anxiety. The marital status of most patients had not changed since the onset of MS but 53% of those who were employed at the time of diagnosis gave up their jobs and the standards of living of 37% of patients and their families had declined as a direct result of the disease. The ability to continue in gainful employment or to maintain social contacts and leisure activities correlated with the course and severity of the disease and cognitive function. Most carers reported symptoms that clearly related to organic pathologies, anxiety and symptoms of depression. The occurrence of these symptoms was associated with disease severity. The professional career of 57% of relatives was also adversely affected by the patient's illness. CONCLUSIONS: MS has a profound impact on the patients' social roles and their relatives' well-being. In contrast to previous studies, a high divorce/separation rate among patients with MS was not observed. Severe disability and cognitive impairment are predictors of loss of employment, decline in the standards of living and withdrawal from social and leisure activities among patients and are strong indicators of stress among relatives. PMID- 10864131 TI - Does personality or psychopathology predict disability in chronic pain patients? AB - PURPOSE: The purpose of this study was to evaluate the usefulness of personality characteristics, depression and personality disorders in predicting disability status in pain patients one year later. METHOD: Subjects were 250 volunteer chronic pain patients. The baseline evaluation consisted of the Karolinska Scales of Personality and psychiatric evaluation of depression and personality disorders using standardized diagnostic instruments. Disability status was assessed by insurance record review one year after the evaluation. RESULTS: The results suggest that baseline personality traits and psychopathology (i.e. depression or personality disorders) were not useful predictors of disability status in pain patients with one-year follow-up. CONCLUSIONS: These data suggest that personality characteristics and psychopathology are probably not important disability predictors in chronic pain patients. PMID- 10864133 TI - Clinical application of the modified medially-mounted motor-driven hip gear joint for paraplegics. AB - PURPOSE: This paper describes a motor-driven orthosis for paraplegics which has been developed. This orthosis is composed of a medially-mounted motor-driven hip joint and bilateral knee-ankle-foot orthosis. With the gear mechanism, the virtual axis of the hip joint of this orthosis is almost as high as the anatomical hip joint. METHOD: A paraplegic patient with an injury level of T10/11 walked using bilateral lofstrand crutches and this new orthosis with or without the motor system. The motor is initiated by pushing a button attached at the edge of the grab of the crutches. RESULT: Faster cadence and speed and smaller rotation angle of the trunk was obtained in motor walking compared with non-motor walking. The patient did not feel fearful of falling. CONCLUSION: The benefit of motor orthosis is that it can be used even in patients with lower motor lesions and that it provides stable regulation of hip flexion movement in spastic patients. In conclusion, this motor orthosis will enhance paraplegic walking. PMID- 10864134 TI - Nimesulide: some pharmaceutical and pharmacological aspects--an update. AB - Nimesulide, a non-steroidal anti-inflammatory drug (NSAID), is administered orally or rectally twice daily for a variety of inflammation and pain states. This is a unique NSAID, not only because of its chemical structure but also because of its specific affinity to inhibit cyclooxygenase-2 (COX-2), thus exerting milder effects on the gastrointestinal mucosa. Current data on selective COX-2 inhibitors suggest that they may have an efficacy similar to that of standard NSAIDs. Initial general clinical experience with selective COX-2 inhibitors appears to show that they are particularly promising in individuals at risk because of renal diseases, hypertension or congestive heart failure. Various experimental models and clinical studies have demonstrated the anti-inflammatory efficacy of nimesulide. Nimesulide is superior, or at least comparable in efficacy, to other NSAIDs, but is better tolerated and has less potential for adverse reactions. Thus, selective COX-2 inhibitors should have anti-inflammatory effects devoid of side effects on the kidney and stomach. They may also demonstrate new important therapeutic benefits as anticancer agents as well as help prevention of premature labour and even retard the progression of Alzheimer's disease. No clinically significant drug interactions have been reported for nimesulide. Not much has been reported about the pharmaceutical aspects of nimesulide. Its poor aqueous solubility poses bioavailability problems in-vivo. This could be overcome by the formation of inclusion complexes with beta cyclodextrin, as has been reported by various researchers. However, absence of any in-vivo data regarding the relative absorption of nimesulide from beta cyclodextrin complex compared with that from conventional formulations of the drug makes the use of such fast-releasing complexes rather questionable. Only a limited number of assay procedures (HPLC, spectrophotometric, spectrofluorimetric) for the determination of nimesulide and its metabolite in plasma/urine samples or in dosage forms have been reported in the literature. The purpose of this review is to provide a concise overview of the pharmacological and pharmaceutical profile of nimesulide. Various investigations carried out recently are reported, although older references to research performed on nimesulide have also been included, where appropriate. PMID- 10864135 TI - Investigation of the absorption of hypericin into the skin of hairless mice. AB - The skin absorption of hypericin was evaluated in hairless mice to develop an optimised hypericin topical formulation that could be used in the clinical study of psoriasis. Hypericin (0.01-1.0%) in Beeler basis, polyethylene glycol ointment, carbopol gel, cetomacrogol cream, petrolatum or emulsifying ointment, with and without skin-absorption enhancers (isopropylidene glycerol and diethylene glycol monoethyl ether), was tested in-vivo on hairless mice skin. Using a skin-stripping technique and the intrinsic fluorescence of hypericin under standardised UV365 irradiation, it was demonstrated that the absorption of hypericin very much depended on the vehicle used. The concentrations of hypericin in the skin were then estimated by HPLC analysis. For this purpose, two vehicles were employed, with which hypericin penetrated the skin of hairless mice well (emulsifying ointment with isopropylidene glycerol) or very poorly (polyethylene glycol ointment). In the case of emulsifying ointment with isopropylidene glycerol (0.05% hypericin), a substantial concentration of hypericin (8.6+/-3.2 microg g(-1)) (mean +/- s.d., n = 5) was found in the skin. With polyethylene glycol ointment, however, only a limited hypericin skin concentration (0.38+/-0 34 microg g(-1), n = 5) was achieved. These results show that emulsifying ointment with polyethylene glycol holds promise as an effective topical vehicle for the treatment of skin diseases, such as psoriasis, with hypericin. PMID- 10864136 TI - Reduction of the potential anticancer drug oracin in the rat liver in-vitro. AB - Studies on the metabolism of the potential cytostatic drug oracin have shown that a principal metabolite of oracin is 11-dihydrooracin (DHO). We conducted in-vitro experiments to investigate the extent of oracin carbonyl reduction in microsomal or cytosolic fractions and to find out the enzymes involved under these conditions. Among several inducers of rat cytochrome P450 only 3 methylcholanthrene caused a significant (P < 0.01) stimulation (1.9 times) of DHO production in microsomal fraction and the specific P4501A inhibitor alpha naphthoflavone significantly (P < 0.01) decreased (twice) the induced reduction activity. Cytochrome P4501A participates in oracin reduction in microsomes. 18beta-Glycyrrhetinic acid, a specific inhibitor of hydroxysteroid dehydrogenase, significantly (P < 0.01) inhibited the production of DHO in the microsomal fraction (>95% inhibition) in comparison with the non-inhibited reaction. Statistically significant (P < 0.01) inhibition (95%) of DHO formation was caused by metyrapone, which is also the substrate of 11-hydroxysteroid dehydrogenase. The main microsomal enzyme which catalyses the carbonyl reduction of oracin is probably 11beta-hydroxysteroid dehydrogenase. Important oracin reduction to DHO in the cytosolic fraction was found. According to its specific sensitivity towards quercitrin (inhibition by 99%, P < 0.01), the enzyme responsible for DHO formation in the rat liver cytosol is postulated to be carbonyl reductase. PMID- 10864137 TI - Uptake of ibuprofen, indomethacin and ketoprofen into isolated rabbit parietal cells. AB - In the United States and other countries, non-steroidal anti-inflammatory drugs (NSAIDs) can be purchased without a prescription. Due to their widespread use, the drugs' most common side effect, gastric toxicity, becomes a more serious concern. Gastric toxicity can occur directly by contact with mucous membranes or indirectly by the inhibition of prostaglandin production in the gastric mucosa. We have studied the uptake of NSAIDs in gastric tissue, specifically parietal cells removed from the rabbit stomach. New Zealand White rabbits were killed and then used to harvest parietal cells. The purified cells were used to study the uptake of ibuprofen, indomethacin and ketoprofen (NSAIDs) over time and under different experimental conditions. The effects of concentration were investigated for all three NSAIDs. In addition, indomethacin and ibuprofen were used to investigate the mechanism of uptake. Studies were determined for the effects of varied extracellular pH from pH 6 to 8, and inhibitory conditions from depressed temperature (5 degrees C), metabolic inhibitors (sodium azide and 2,4-dinitro phenol), an ionophore (nigericin) and a sodium free support medium. The interaction of NSAIDs with lysed parietal cells was investigated also. Initial rate data indicated that Michaelis-Menten kinetics were apparent; however, poor solubility of all three NSAIDs prevented complete characterization of the inclusion of a passive transport mechanism. Uptake showed a statistically significant increase (P = 0.01 to 0.05) as pH decreased, also suggesting contribution from a passive mechanism. Studies with inhibitors showed minimal effects. However, uptake at equilibrium for the ionophore, nigericin, showed a 10 fold increase over the control for ibuprofen (P = 0.005) and a 1.4-fold increase for indomethacin (P=0.04). Depressed temperature (5 degrees C) increased the initial rate and uptake at equilibrium 2.1- and 2.2-fold, respectively, for ibuprofen (P < 0.01). For indomethacin depressed temperature increased the initial rate and uptake at equilibrium 2.7- and 5.2-fold, respectively (P < 0.01). The increases at 5 degrees C suggests that adsorption may be an important uptake component. Experiments with lysed parietal cells showed a non-specific uptake phenomenon, suggesting an adsorption component was occurring also. PMID- 10864138 TI - In-vitro metabolism of retinoic acid by different tissues from male rats. AB - Significant differences between the metabolism of retinoic acid by different tissues might be an important determinant of the effectiveness of a systemically administered inhibitor at a particular tissue site. Here the metabolism of retinoic acid has been studied in microsomal fractions from different tissues (liver, kidney, intestinal mucosa, lung, skin, brain) of the male rat to determine their relative metabolic activity. Kinetic analysis revealed major differences between the activity of different tissue microsomes. This is shown by the Vmax values for the metabolism of retinoic acid-liver (102+/-39.0 pmol (mg protein)(-1) min(-1)) was 100 times more active than the lung (1+/-0.03 pmol (mg protein)(-1) min(-1)), which was the least active. The range of Km values for microsomes from the different tissues was narrow (0.48-1.40 microM). Taking into account the mass of the tissue, the gross activity ranking for metabolism of retinoic acid was liver >> skin = kidney > brain > intestinal mucosa >> lung. It is concluded that metabolism of administered retinoic acid occurs mainly in the liver but that cellular retinoic acid levels in some other tissues (skin, kidney, brain) could be reduced (metabolized) to such an extent that higher levels might be observed after the use of inhibitors of retinoic acid metabolism. PMID- 10864139 TI - Structure-activity relationships for the Ca2+-releasing activity of 6-hydroxy beta-carboline analogues in skeletal muscle sarcoplasmic reticulum-the effects of halogen substitution at C-5 and C-7. AB - This study of structure-activity relationships of 6-hydroxy-beta-carboline analogues has been performed on the basis of quantitative measurement of Ca2+ releasing activity in the sarcoplasmic reticulum of skinned fibres of skeletal muscle. Substitution of halogens for hydrogens at the C-5 and C-7 positions and further introduction of a methyl group into the N-9 position of 6-hydroxy-beta carboline resulted in Ca2+-releasing activity. The 50% effective concentrations of 5,7-dibromoeudistomin D, 5,7-dichloroeudistomin D, 5,7-diiodoeudistomin D, 9 methyl-5,7-dibromoeudistomin D, 9-methyl-5,7-dichloroeudistomin D, 9-methyl-5,7 diiodoeudistomin D, and caffeine were 5.6 x 10(-6), 6.3 x 10(-6), 7.8 x 10(-6), 2.1 x 10(-6), 2.0 x 10(-5), 3.7 x 10(-5), and 4.7 x 10(-4) M, respectively, indicating that these analogues are 10-200 times more potent than caffeine. Substitution of bromine by chlorine or iodine at the C-5 and C-7 positions markedly reduced the activity of the analogues with a methyl group at the N-9 position. These results suggest that halogens at the C-5 and C-7 positions in the beta-carboline skeleton are essential for Ca2+-releasing activity and that an N-9 methyl group also affects the activity of these analogues. Thus, these 6-hydroxy beta-carboline analogues might become powerful tools for studying the molecular mechanism of Ca2+ release in the sarcoplasmic reticulum. PMID- 10864140 TI - Antioxidant properties of novel lipophilic ascorbic acid analogues. AB - Structural modifications of ascorbic acid by the introduction of lipophilic moieties has led to derivatives with increased stability against thermal and oxidative degradation. Two series of new lipophilic ascorbic analogues were synthesized to obtain antioxidants devoid of autooxidant properties: 4-benzoyl-3 hydroxyfuran-2(5H)-ones (3a-j) and 4-acetyl-5-aryl-3,4-dihydrofuran-2(5H)ones (5a f). These compounds were submitted to three different tests: reduction of the stable free radical, 1,1-diphenyl-2-picrylhydrazyl (DPPH); superoxide-anion scavenging assay; and lipid-peroxidation assay. Most compounds interacted with DPPH: at a concentration of 5 x 10(-3) M, the reducing activity of 4-benzoyl derivatives, 3c and 3h, was more than 50%; under the same conditions, the rate of inhibition for 4-acetylbutanolides, 5a and 5f, reached 60.6% and 87.3%, respectively; 93.3% inhibition was observed with ascorbic acid. In the superoxide anion scavenging assay, at a concentration of 1 mg mL(-1), 4-benzoyl derivatives, 3g and 3i, exhibited a good activity, with IC50 (dose resulting in 50% inhibition) values of 1.45 and 1.35 x 10(-3) M, respectively. 4-Acetylbutanolide, 5f, significantly inhibited the Fe2+/ADP/ascorbate-induced lipid peroxidation of rat liver microsomes with an IC50 of 4.9 x 10(-4) M. This study demonstrates that enol functions in the structure of ascorbic acid analogues are not absolutely essential to bring about antioxidant effects. PMID- 10864141 TI - Effects of ZNC-2381, a new oral compound, on several hepatic injury models and on hepatocellular apoptosis in mice and rats. AB - The hepatoprotective effect of ZNC-2381 (1-(4-aminophenyl) methyl-3-(3 nitrophenyl)-1,3-dihydroimidazo[4,5-b]pyridine-2-one), a novel 2-one dihydroimidazopyridine derivative, has been evaluated in several experimental models of hepatic injury. In mice, oral ZNC-2381, administered at doses of 3, 10 or 30 mgkg(-1), 1 h before induction of hepatic injury with concanavalin A, dose dependently inhibited increases in serum alanine aminotransferase (ALT) activity. Apoptosis of liver cells, as indicated by DNA fragmentation (nucleosome assay) and DNA-ladder formation (electrophoresis), was also inhibited dose-dependently. ZNC-2381 dose-dependently inhibited concanavalin A-induced increases in serum tumour necrosis factor (TNF)-alpha levels, and TNF-alpha mRNA expression in the liver. Oral ZNC-2381 also dose-dependently inhibited increases in serum ALT activity in mice with hepatic injury induced by Propionibacterium acnes and a bacterial lipopolysaccharide (LPS) or D-galactosamine-LPS, and in rats with D galactosamine-induced hepatic injury. These results indicate that oral ZNC-2381 inhibits cytokine (TNF-alpha) production and cytokine-related hepatocellular apoptosis, and might thus prevent different types of hepatic injury. PMID- 10864142 TI - The upstream regulation of p38 mitogen-activated protein kinase phosphorylation by arachidonic acid in rat neutrophils. AB - The signal transduction pathways activated by arachidonic acid that lead to p38 mitogen-activated protein kinase (MAPK) activation in neutrophils remains unclear. In this study, selective inhibitors of several signalling pathways were utilized to investigate the mechanisms of activation of p38 MAPK by arachidonic acid in rat neutrophils. Stimulation of p38 MAPK phosphorylation by arachidonic acid and its trifluoromethyl ketone analogue AACOCF3 was transient, peaking at 1 min, and was concentration-dependent. Arachidonic acid-stimulated p38 MAPK phosphorylation was attenuated in cells pretreated with the Gi/o inhibitor (pertussis toxin), but not with the dual cyclooxygenase/lipoxygenase inhibitor (BW755C) or the leukotriene biosynthesis inhibitor (MK886). Tyrosine kinase inhibitor (genistein), but not the extracellular signal-regulated kinase kinase inhibitors (PD98059 and U0126), attenuated the phosphorylation of p38 MAPK by arachidonic acid. Phosphoinositide 3-kinase inhibitors (wortmannin and LY294002) did not affect the arachidonic acid-induced response. After pretreatment of the cells with protein kinase C inhibitors (Go6976, Go6983 and GF109203X), only Go6976 significantly attenuated the phosphorylation of p38 MAPK by arachidonic acid. In addition, phosphorylation of p38 MAPK by arachidonic acid was greatly attenuated by the phospholipase C inhibitor (U73122) and the Ca2+ chelator BAPTA ((1,2-bis-o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid), but not altered by the nitric oxide synthase inhibitor, N-nitro-L-arginine methyl ester. Arachidonic acid did not cause an increase in cellular cyclic GMP level. This study revealed the involvement of pertussis toxin-sensitive G protein, non-receptor tyrosine kinase, phospholipase C/Ca2+, and probably Ca2+-dependent protein kinase C in arachidonic acid-stimulated p38 MAPK activation. PMID- 10864143 TI - Protective effect of N-acetylcysteine on rat liver cell membrane during methanol intoxication. AB - Methanol is oxidized in-vivo to formaldehyde and then to formate, and these processes are accompanied by the generation of free radicals. We have studied the effect of N-acetylcysteine on liver cell membrane from rats intoxicated with methanol (3.0 g kg(-1)). Evaluation of the effect was achieved by several methods. Lipid peroxidation and surface charge density were measured. An ultrastructural study of the liver cells was undertaken. The concentration of marker enzymes of liver damage (alanine aminotransferase and aspartate aminotransferase) in blood serum was measured. Methanol administration caused an increase in lipid peroxidation products (approximately 30%) as well as in surface charge density (approximately 60%). This might have resulted in the membrane liver cell damage visible under electron microscopy and a leak of alanine aminotransferase and aspartate aminotransferase into the blood (increase of approximately 70 and 50%, respectively). Ingestion of N-acetylcysteine with methanol partially prevented these methanol-induced changes. Compared with the control group, lipid peroxidation was increased by approximately 3% and surface charge density by approximately 30%. Alanine aminotransferase and aspartate aminotransferase activity increased by 9 and 8%, respectively, compared with the control group. The results suggested that N-acetylcysteine was an effective antioxidant in methanol intoxication. It may have efficacy in protecting free radical damage to liver cells following methanol intoxication. PMID- 10864144 TI - Anti-inflammatory effect of FR140423, a novel selective cyclo-oxygenase-2 inhibitor, in rat adjuvant arthritis without gastrointestinal side effects. AB - We investigated the effect of FR140423 (3-(difluoromethyl)-1-(4-methoxyphenyl)-5 [4-(methylsulphinyl)phen yl]pyrazole), a novel and selective cyclo-oxygenase (COX)-2 inhibitor, in rat adjuvant arthritis. The results were compared with that of indomethacin. We tested the inhibitory effects of FR140423 on paw oedema and the formation of the arachidonic acid metabolites prostaglandin (PG) E2 and leukotriene (LT) B4 in inflamed paws immunized with heat-killed and dried Mycobacterium tuberculosis. Oral administration of FR140423 showed a dose dependent anti-inflammatory effect. This effect was two- to threefold more potent than that of indomethacin. The increase of PGE2 and LTB4 in inflamed paws was associated with the development of paw swelling. FR140423 and indomethacin dose dependently suppressed the level of PGE2 but not LTB4 in arthritic paws. Unlike indomethacin, FR140423 did not induce gastric lesions even at doses up to 10 mgkg(-1) in arthritic rats. FR140423 has a potent anti-inflammatory effect mediated by inhibition of PGE2 produced by COX-2 in inflamed tissues. The safety profile of FR140423 appears to be an improvement on the safety profile of indomethacin. PMID- 10864145 TI - Benidipine inhibits apoptosis during ischaemic acute renal failure in rats. AB - We have investigated the effects of benidipine (hydrochloride), a calcium antagonist, against ischaemic acute renal failure in rats. Using histological examination, we studied whether the inhibition of apoptosis was associated with the protective effects of benidipine on the ischaemic renal injury. Acute renal failure was induced by the unilateral clamping of the left renal artery for 60 min, followed by reperfusion and contralateral nephrectomy. Drugs were given intravenously 5 min before the unilateral clamping. Prophylactic administrations of benidipine (10 microg kg(-1), i.v.) significantly ameliorated the development of renal failure as estimated by the measurements of serum creatinine and blood urea nitrogen 24 h after the reperfusion. Amlodipine (besilate, 100 and 300 microg kg(-1), i.v.) tended to attenuate renal dysfunction. Lisinopril (300 and 1000 microg kg(-1), i.v.), an angiotensin converting enzyme inhibitor, was ineffective in this acute renal failure model. Histological examination using the terminal transferase-mediated dUTP-biotin nick end-labelling (TUNEL) method to detect apoptotic cells revealed that the TUNEL-positive tubular epithelium was prominent in the renal cortex 24 h after the reperfusion. The TUNEL-positive cells were significantly reduced by pretreatment with benidipine. The results demonstrate that benidipine can ameliorate the ischaemic acute renal failure in rats and suggest that the renoprotective effect of benidipine was at least partly attributable to the reduction of apoptosis in tubular epithelial cells. PMID- 10864146 TI - Effect of lamivudine on uptake of organic cations by rat renal brush-border and basolateral membrane vesicles. AB - The effect of lamivudine on uptake of a representative organic cation, tetraethylammonium (TEA), by rat renal brush-border membrane vesicles (BBMV) and basolateral membrane vesicles (BLMV) has been investigated. The pH-driven uptake of TEA by BBMV (pHin = 6.0, pHout = 7.5) was inhibited by lamivudine. The IC50 value (concentration resulting in 50% inhibition) for the concentration-dependent effect of lamivudine on TEA uptake by BBMV after 30 s was 2668 microM whereas IC50 values for cimetidine and trimethoprim were < 2.5 microM and < 25 microM, respectively. The early uptake of TEA by BLMV was also reduced significantly by lamivudine. The IC50 value for the concentration-dependent effect of lamivudine on uptake of TEA by BLMV at 30 s was > 25 mM, whereas the IC50 values for cimetidine and trimethoprim were 2116 microM and 445 microM, respectively. These findings suggest that compared with other cationic drugs, such as trimethoprim and cimetidine, lamivudine is a weak inhibitor of organic cation transport into the tubules by the brush-border and basolateral membranes of renal epithelial cells. It is unlikely lamivudine will have any significant effect on the excretion of co-administered cationic drugs by the renal tubules. PMID- 10864147 TI - Characterization of histamine release induced by fluoroquinolone antibacterial agents in-vivo and in-vitro. AB - Characterization of histamine release induced by fluoroquinolone antibacterial agents, levofloxacin and ciprofloxacin, was investigated in-vivo and in-vitro. Intravenous injection of levofloxacin and ciprofloxacin at 1-10 mg kg(-1) produced dose-related elevations in plasma histamine level in anaesthetized dogs. In contrast, levofloxacin was devoid of plasma histamine increment in anaesthetized rats at 100 mg kg(-1), whereas ciprofloxacin at the same dose caused endogenous histamine release. Levofloxacin and ciprofloxacin induced non cytotoxic secretion of histamine from all mast cells tested in a concentration dependent manner, whereas rat skin and peritoneal mast cells were thirty- to one hundred-times less sensitive to the effect of fluoroquinolones as compared with the canine skin mast cells. These results suggest that the functional heterogeneity of mast cells from different species in histamine releasing activity of fluoroquinolones may exist, and that mast cells from the dog appear to be particularly sensitive to the effect of the fluoroquinolones. PMID- 10864148 TI - Sedum telephium L. polysaccharide content affects MRC5 cell adhesion to laminin and fibronectin. AB - In traditional medicine the fresh leaves and juice of Sedum telephium L. are used as wound-healing promoters. Cell adhesion represents a primary event in wound repair and in tissue homeostasis, and therefore we have investigated the effect of Sedum juice and its main fractions, polysaccharides and flavonols, on human fibroblast (MRC5) adhesion to fibronectin and laminin. Our findings revealed that total Sedum juice strongly inhibited cell adhesion to laminin and fibronectin (EC50 1.03+/-0.12 mg mL(-1)). This anti-adhesive feature was concentrated mainly in the two polysaccharide fractions (EC50 values comprised between 0.09 and 0.44 mg mL(-1)). The flavonol fractions did not seem to contribute to this effect. A first attempt to elucidate the polysaccharide-related anti-adhesive feature of Sedum juice was also performed. The results confirmed that natural polysaccharides, with chemical structures different from heparin, were able to interfere with integrin-mediated cell behaviour and they contributed to the outstanding effects of Sedum juice and to the role of polysaccharides in cell matrix interaction. PMID- 10864149 TI - Anti-leukaemic and anti-mutagenic effects of di(2-ethylhexyl)phthalate isolated from Aloe vera Linne. AB - Extracts of Aloe vera Linne have been found to exhibit cytotoxicity against human tumour cell lines. This study examines the anti-tumour effects of di(2 ethylhexyl)phthalate (DEHP) isolated from Aloe vera Linne, in human and animal cell lines. Its anti-mutagenic effects were examined using Salmonella typhimurium TA98 and TA100 strains. Growth inhibition was specifically exerted by DEHP against three leukaemic cell lines at concentrations below 100 microg mL(-1). At 100 microg mL(-1) DEHP, K562, HL60 and U937 leukaemic cell lines showed growth inhibition of 95, 97 and 95%, respectively. DEHP exhibited an inhibitory activity of 74, 83 and 81%, respectively, in K562, HL60 and U937 cell lines at a concentration of 10 microg mL(-1). At a concentration of 1 microg mL(-1), DEHP exerted an inhibitory activity of 50, 51 and 52%, respectively, in K562, HL60 and U937. In a normal cell line, MDBK, DEHP exerted 30% growth inhibition at a concentration of 100 microg mL(-1), and showed no inhibitory activity at concentrations below 50 microg mL(-1). It was found that DEHP exerted anti mutagenic activity in the Salmonella mutation assay. The number of mutant colonies of Salmonella typhimurium strain TA98 upon exposure to AF-2 (0.2 microg/plate) decreased in a concentration-dependent manner in the presence of different DEHP concentrations (decreasing to 90.4, 83.9, 75.4, 69.6 and 46.9%, respectively, for DEHP concentrations of 100, 50, 10, 5 and 1 microg/plate). In the case of Salmonella typhimurium strain TA100, DEHP reduced AF-2-induced mutagenicity at 1, 5, 10, 50 and 100 microg/plate to 57.4, 77.5, 80.0, 89.0 and 91.5%, respectively. The isolated compound from Aloe vera Linne, DEHP, was considered to be the active principle responsible for anti-leukaemic and anti mutagenic effects in-vitro. PMID- 10864150 TI - Photodegradation of polychlorinated dibenzo-p-dioxins and dibenzofurans in aqueous solutions and in organic solvents. AB - Aqueous solutions of selected polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) were prepared using a generator column and exposed to UV (300 nm) light in the laboratory and to sunlight in an outdoor environment. In the laboratory, additional exposures were also carried out using 60% acetonitrile/water solutions. At 300 nm di- and tetra PCDDs had higher first order photodegradation rate constants in 60% acetonitrile/water than in pure water. The solvent effect was reversed for PCDFs. These results may be a reflection of the higher polarity of PCDFs compared to PCDDs. In both the indoor and outdoor exposures photodegradation rates decreased with increasing concentrations of chlorination. However, OCDF exposed to 300 nm light in 60% acetonitrile/water and to sunlight in pure water photodegraded more rapidly than tetra CDF. Photolysis rates in sunlight were considerably slower (t(1/2) of 6.4 23 h) than photolysis rates at 300 nm in the laboratory (t(1/2) of 4.3-680 min), reflecting the lower intensity of sunlight in the 300 nm region of the UV/Vis spectrum. The extent of dechlorination of the PCDDs/PCDFs was less than 20% and reductive dechlorination does not appear to be a major process in the photodegradation of PCDDs/PCDFs in aqueous solutions. PMID- 10864151 TI - Organochlorine and heavy metal residues in the water/sediment system of the Southeast Regional Park in Madrid, Spain. AB - A study into levels of contamination by organochlorine compounds (insecticides and PCBs) and heavy metals (Cd and Pb) in the water/sediment system of the Southeast Regional Park (SERP) in Madrid, Spain, has been carried out. Residue levels of xenobiotics were determined in surface and underground waters and sediments from selected sites throughout the protected area. The results showed these contaminants to be widespread throughout the studied area. p,p'-DDT concentration levels were consistently higher than its metabolite p,p'-DDE, indicating a recent use of this organochlorine insecticide in the area. PCB levels exceeded, in the majority of the cases, the levels taken as the maximum (100 ng/microl) for highly polluted waters. Cd and Pb levels found in water samples were under the detection limits of the methodology used. Pb levels found in sediment samples were higher than Cd. PMID- 10864152 TI - Estimating K(oc) for persistent organic pollutants: limitations of correlations with K(ow). AB - The n-octanol/water partition coefficient (K(ow)) is commonly used to predict the soil or aquatic particle water partition coefficient normalized to organic carbon (K(oc)). Many correlations are available covering several chemical classes and ranges of hydrophobicity. This work indicates the K(ow) may not be a strong predictor for persistent organic pollutants (POPs) which are defined here as chemicals with logK(ow) > 5.0. In addition, the correlation developed in this work for POPs will still result in a predicted value which is of by a factor of 15. Accordingly, care must be taken when applying K(oc) estimations using K(ow) for POPs until more suitable correlations are developed. PMID- 10864153 TI - Dechlorination of hexachlorobiphenyl by using potassium-sodium alloy. AB - 2,2',4,4',5,5'-Hexachlorobiphenyl (HCB) was dechlorinated with potassium-sodium (K-Na) alloy under an inert gas atmosphere. Solvent effect was observed in the reaction. Dechlorination yields in benzene and cyclohexane were 99.9998%, and 99.99996%, respectively. The reaction was exothermic and proceeded at room temperature. In benzene, trace amounts of polychlorinated biphenyls (PCBs) as products by stepwise dechlorination and polychlorinated quarterphenyls as product of Wurtz-Fittig reaction were detected as reaction intermediate. Reaction products were biphenyl, cyclohexylbenzene, and dicyclohexyl. In cyclohexane, there were no products of Wurtz-Fittig reaction. Dechlorination at para-position preferred to that at ortho-position, judging from analysis of PCBs as intermediates of stepwise dechlorination. PMID- 10864154 TI - Toxic chlorinated and polyaromatic hydrocarbons in simulated house fires. AB - Toxic chlorinated hydrocarbons (polychlorinated biphenyls, benzenes and dioxins and furans) and polyaromatic hydrocarbons were examined in combustion gas and deposited soot wipe samples from simulated house fires. Concentrations of these substances were high during the fires, the amounts of polychlorinated dioxins and furans (PCDD/Fs) in the combustion gas varying from 1.0 to >7.2 ng/m3 (I-TEQ) and those of polyaromatic hydrocarbons from 6.4 to 470 mg/m3. Thus large amounts of organic compounds may be released in house fires. As a result, there is a need for careful personal protection of fire-fighters and remediation workers against combustion gases during a fire and on contaminated surfaces after it. PMID- 10864156 TI - Electronic eigenvalue (EEVA): a new QSAR/QSPR descriptor for electronic substituent effects based on molecular orbital energies. A QSAR approach to the Ah receptor binding affinity of polychlorinated biphenyls (PCBs), dibenzo-p dioxins (PCDDs) and dibenzofurans (PCDFs). AB - A new descriptor of molecular structure for use in the derivation of predictive QSAR and QSPR models, electronic eigenvalue (EEVA), is described. This is a modification of the recently proposed EVA approach, but is based on computationally-derived molecular orbital energies instead of vibrational frequencies. Like EVA, it is also invariant as to the alignment of the structures concerned. Its performance has been tested with respect to the Ah receptor binding of PCBs, PCDDs and PCDFs, and its predictive ability has been clearly demonstrated. In particular, it seems to be suitable for 'pure' electronic substituent effects. i.e., for cases in which both hydrophobic and steric factors are of minor importance. PMID- 10864155 TI - Riverine fluxes of the persistent organochlorine pesticides hexachlorcyclohexane and DDT in the Russian Federation. AB - The contribution of gross riverine organochlorine pesticide (OCP) transport to estuaries of Russian seas and Lake Baikal was determined to help understand OCP transboundary transfer and to provide a basis for estimating Russia's contribution to global pollution by these pesticides. The official OGSNK/GSN data ranks sea/ocean/lake basins in the following order based upon the amounts of total OCPs received from agricultural use: Eastern Arctic>Western Arctic>Pacific>Baltic>Caspian>Azov/Black>Baikal. A similar ranking was obtained using an independent set of data: Eastern Arctic>Pacific>Caspian>Western Arctic>Baltic>Azov/Black. In terms of riverine flow-associated discharge of HCH isomers (i.e., sum of alpha-, beta- and gamma-HCH) estuaries of the Kara, Okhotsk and Beloye (White)/Barents seas received more pesticides than other seas. No HCH was discharged to estuaries of the Eastern Siberian and Bering seas. For DDT and its derivative (DDE), estuaries of the Kara, Caspian, Okhotsk and Baltic seas received the greatest amounts. During our study period (1988-1996), HCH transport was more prevalent in the majority of rivers reflecting both the official ban on the use of DDT in the former Soviet Union and the greater popularity of HCH as a pesticide. In general, it appears that Russian rivers play a significant role in OCP contamination of some estuaries of regional seas, especially those of the eastern Arctic basin, such as the Kara Sea. PMID- 10864157 TI - Chlorobenzenes and hexachlorocyclohexanes (HCHs) in the atmosphere of Bitterfeld and Leipzig (Germany). AB - Concentrations of tetrachlorobenzenes, pentachlorobenzene, hexachlorobenzene and alpha-, beta-, gamma- and delta-HCH in air and deposition were measured at three different contaminated sites in Greppin, Roitzsch (both near Bitterfeld) and Leipzig during five time intervals of 14 days in the summer months of 1998. The mean values of the chlorobenzene concentrations (gas phase and particle bound portions) over the whole sampling time were 0.11 ng/Nm3 (Leipzig), 0.17 ng/Nm3 (Roitzsch) and 0.37 ng/Nm3 (Greppin), the mean values of the HCH concentrations were 0.22 ng/Nm3 (Leipzig), 0.31 ng/Nm3 (Roitzsch) and 0.69 ng/Nm3 (Greppin). This increase of the concentration values from Leipzig over Roitzsch to Greppin indicates the influences of industrial waste sites in the Bitterfeld region on the atmospheric environment. The significantly higher values of hexachlorobenzene, alpha- and beta-HCH in Greppin are probably caused by emissions from the former chemical plant Bitterfeld-Wolfen and the landfill 'Antonie' near Greppin. Compared with literature data from other industrial impacted areas the measured air concentration and deposition values are relatively low. PMID- 10864158 TI - Thermal remediation of PCDD/Fs contaminated soil by zone combustion process. AB - A new thermal process has been proposed for remediating soils contaminated by chlorinated organic compounds, e.g., PCDD/Fs and PCBs. This is to apply the "zone combustion process" which utilizes stable combustion of coke particles in the packed bed to soils with air flow across the bed. The usefulness and validity were obtained the results showing that more than 98.9% of PCDD/Fs in the soil was successfully removed in a laboratory-scale experiment. Some pretreatment of the soil sample, such as drying, pre-granulation and addition of limestone was found to make the removal efficiency better. Although, some fundamentals on the behavior of PCDD/Fs, e.g., decomposition/vaporization ratios and formation of other compounds cannot be certainly identified yet, the present results clearly show a way to remediate the contaminated soils and solid wastes. PMID- 10864159 TI - A survey of PCDD and PCDF in French long-life half-skimmed drinking milk. AB - A national survey was carried out in order to assess the concentrations of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) in long-life half-skimmed drinking milk produced and consumed in France. 151 Samples were collected from 33 dairy establishments selected for their production amounts following a random sampling scheme. 148 of the 151 results were finally used for statistical assessment. The mean concentration of 2,3,7,8 TCDD toxicity equivalent found is 0.65 pg/g of milk fat. This result is far below the threshold recommended by the European Union. PMID- 10864160 TI - TiO2 photocatalytic degradation of PCBs in soil-water systems containing fluoro surfactant. AB - Titanium dioxide-mediated photodegradation of Polychlorinated biphenyls (PCBs) in soil/aqueous systems with added fluorinated surfactant was investigated. PCBs can bind tightly to organic matter in the soil, especially in aged, contaminated soil. Experiments showed an effective PCB photocatalytic degradation in mixed systems of soil/clay with anionic fluorinated surfactant FC-143 and TiO2. The FC 143 surfactant is stable in this photochemical process. PCB degradation rates in samples followed the order: spiked clay > spiked soil > Hudson River bank soil. The results suggest that anionic fluorinated surfactant may form semimicelles and/or admicelles on the surface of positively charged TiO2. The hydrophobic surface of TiO2 can provide a nonpolar phase that acts as a partioning medium for hydrophobic PCBs. Therefore, PCBs in soil can be released to the semimicelle and/or admicelle on the TiO2 surface and are effectively photodegraded in a dispersion containing anionic fluorinated surfactant. The combination of surfactant extraction and photooxidation forms the basis for a novel two-stage process for the removal and destruction of PCBs from soil. PMID- 10864161 TI - Study of the distribution of the polychlorinated biphenyls in the milk fat globule by supercritical fluid extraction. AB - The distribution of polychlorinated biphenyls (PCBs) in the milk fat globule has been studied by sequential extraction of four different lipidic fractions from powdered full-fat milk with supercritical carbon dioxide. Extractions were carried out in the dynamic mode in the pressure range 13.6-23.3 MPa at a temperature of 50 degrees C. The levels of PCBs and short-chain triglycerides (SCT), medium-chain triglycerides (MCT) and long-chain triglycerides (LCT), as well as the cholesterol in these four supercritical fluid extraction (SFE) fractions were determined. Extracts obtained at lower pressures were found to be enriched in PCBs, SCT, MCT and cholesterol, while the concentrations of all these analytes decreased for subsequent extractions. Satisfactory quadratic correlations were found between the PCB and SCT and MCT levels (r2 in the range 0.9-0.999), and between the PCB and cholesterol levels (r2 in the range 0.8 0.999, except for PCB 105) determined in the SFE fractions. These results suggested a similar distribution of the PCBs and the cholesterol in the milk fat globule. This conclusion was also supported by the comparison of the PCBs and the cholesterol chemical structures as well as by the total PCB levels determined by SFE to those obtained by using different extraction methods. The classical requirement for the analysis of this kind of lipophilic pollutants of an exhaustive extraction of the lipids of the matrix to ensure their quantitative recovery has been revised. PMID- 10864162 TI - Concentrations of chlorobenzenes, hexachlorobutadiene and heavy metals in surficial sediments of Kaohsiung coast, Taiwan. AB - This work analyzes surface sediment samples collected from 40 stations along the Kaohsiung coast in southern Taiwan for chlorobenzenes (CBs), hexachlorobutadiene (HCBD) and heavy metals (Cu, Zn, Pb, Cd, Ni, Fe, Mn and Cr). The highest CBs concentrations are recorded in station T7-15 (about 10 km west off the outlet of Da-lin-pu ocean outfall pipe), with total di-, tri-, tetra-, penta- and hexa chlorobenzenes concentrations of 290.5, 117.1, 64.5, 15.7 and 22.3 ng/g, respectively. The major pollution source of HCBD is most likely located in the Tso-yin ocean outfall field; while the Dah-lin-pu ocean outfall field and Kao ping Chi estuary, located in the southern portion of Kaohsiung coast, are the major contributors of hexachlorobenzene. The concentration of CBs congeners correlate fairly well with each other, as do metals. However, concentrations of organics (CBs and HCBD) did not correlate with metals. This finding implies that the pollution characteristics of organics and heavy metals in this highly utilized coastal zone markedly differ from each other. PMID- 10864163 TI - Polychlorinated organic compounds (PCOCs) in waters, suspended solids and sediments of the Yangtse River. AB - The contamination levels of polychlorinated organic compounds (PCOCs) in waters, suspended solids and sediments of the Yangtse River (Nanjing part) were analyzed in this paper. Their concentrations determined by GC/MS were very low in comparison with those in European River. The average concentration of total HCH (alpha-HCH, beta-HCH, gamma-HCH) was much higher than that of other PCOCs in all waters, which made up 65% of total amount of PCOCs. Due to the complete dilution and mixture of pollutants in the mighty Yangtse River, the content of PCOCs at each sampling station demonstrated very similar spatial pattern for waters and suspended solids. Since the small suspended solid (<0.7 microm) passed through the filter was also considered as dissolved part, the dominant parts of HCHs, PCA and PCBs were found in dissolved phase with percentage proportion of 85-94%, 72 85% and 61-78%, respectively. For DDTs, HCB and PeCB, their contents in dissolved phase were slightly higher than in particulate phase. The contents of PCOCs in sediments were also very low and varied with high fluctuation at different sampling points, indicating the heterogeneous deposition. HCB and its metabolite (PeCB) presented the highest contamination levels among PCOCs in sediments. PMID- 10864164 TI - Destruction of halogenated hydrocarbons with solvated electrons in the presence of water. AB - Model halogenated aromatic and aliphatic hydrocarbons and halogenated phenols were dehalogenated in seconds by solvated electrons generated from sodium in both anhydrous liquid ammonia and ammonia/water solutions. The minimum sodium required to completely dehalogenate these model compounds was determined by increasing the Na/substrate ratio until halogen loss was complete. Minimum sodium consumptions were determined in both anhydrous liquid ammonia and with a (5, 20, 50-fold molar excess of water per mole of halide). While more Na was consumed in the presence of water, these dehalogenations were still efficient when a 50-fold water excess was present. Dehalogenation is faster than competiting reactions with water. CCl4 and CH3CCl3 in the presence of a stoichiometric deficiency of sodium produced only CH4 and CH3CH3 and recovered CCl4 or CH3CCl3, respectively. No partially dechlorinated products were detected, indicating dechlorination was diffusion controlled. Na consumption per chlorine removed (as NaCl) was lower than that of Li, K or Ca and this advantage increased in the presence of water. Na consumption was lower using Na chunks instead of a thin Na mirror. Chloroaromatic compounds gave the parent aromatic hydrocarbon and aminated products in anhydrous ammonia but aminated products did not form when water was present. PMID- 10864166 TI - Quantitative relationship between chromatographic retentions and molecular structures of polychlorinated dibenzo-p-dioxins (PCDDs). AB - A new approach to study the quantitative relationships between chromatographic retentions and molecular structures of polychlorinated dibenzo-p-dioxins (PCDDs) is described. The retention equations of PCDDs log k' = A + B/T in gas chromatography (GC) are used to evaluate the properties of the regression coefficients A and B, which have been widely accepted as highly reliable chromatographic retentions. The quantitative relationships between the A, B values and the molecular structures are found. The molecular descriptors given for the first time in this article are very effective. As a result, the regression equations are derived with correlation coefficients greater than 0.9995. The A, B values of PCDDs with no standards available have been predicted according to these relationships. They are very useful in chromatographic identification. The retention times of all PCDDs can be conveniently predicted at any temperature program. Compared with the data obtained from the relevant experiments, the results of prediction are very accurate. PMID- 10864165 TI - Prediction of the retentions of polybrominated dibenzo-p-dioxins (PBDDs) by using the retentions of polychlorinated dibenzo-p-dioxins (PCDDs). AB - The retention equations In k' = A + B/T of 49 polychlorinated dibenzo-p-dioxins (PCDDs) and 4 polybrominated dibenzo-p-dioxins (PBDDs) in gas chromatography (GC) have been investigated to evaluate the properties of regression coefficients A and B. The quantitative relationships between A and B values of PCDDs and those of PBDDs are found. The regression equations derived have correlation coefficients greater than 0.997. The A, B values of any PBDD can be predicted by using the A, B values of the PCDD according to these relationships. Using these predicted A and B values, the retention times of all PBDDs can be predicted at any temperature program. It is very useful to identify the peak position of any PBDD because at present there are only a few standards of PBDDs available. PMID- 10864167 TI - An evaluation of three empirical air-to-leaf models for polychlorinated dibenzo-p dioxins and dibenzofurans. AB - Three empirical air-to-leaf models for estimating grass concentrations of polychlorinated dibenzo-p-dioxins and dibenzofurans (abbreviated dioxins and furans) from air concentrations of these compounds are described and tested against two field data sets. All are empirical in that they are founded on simplistic bioconcentration and related approaches which rely on field data for their parameterization. One of the models, identified as the EPA Model, partitions the total air concentration into vapor and particle phases, and separately models the impact of both. A second model addresses only the vapor phase; grass concentrations are modeled as a function of vapor deposition. For the third model, it is assumed that the grass plants "scavenge" a fixed volume of air of dioxins, and hence grass concentrations are modeled as a simple product of total air concentration and a constant scavenging coefficient. Field data from two sites, a rural and an industrial site in the United Kingdom, included concurrent measurements of dioxins in air and field grass, and dioxin and furan depositions, for one 6-week sampling period. Principal findings include: (1) the EPA Model underpredicted grass concentrations at the rural field site by a factor of 2, while the Scavenging Model underpredicted grass concentrations by a factor of 3.8, and the Vapor Deposition Model significantly underpredicted grass concentrations (by a factor greater than 10), (2) the presence of high soil concentrations for some of the dioxins and furans at the industrial site appears to have caused higher grass concentrations and confounded the air-to-plant modeling exercise, (3) the Scavenging Model could be calibrated to the data set; however, a key premise of this model that vapor and particle phase dioxins equally impact the plants, is not supported by the field data, (4) measured depositions are highly correlated to but systematically lower than modeled depositions, which could be due to modeling assumptions or a systematic measurement bias. PMID- 10864168 TI - Chemical kinetic modelling of PCDD formation from chlorophenol catalysed by incinerator fly ash. AB - A kinetic model is developed for PCDD formation from chlorophenol catalysed by incinerator fly ash. The key step in the model is a Langmuir-Hinshelwood type elementary step for the coupling of two adsorbed chlorophenol species to PCDD. Kinetic expression is derived which can relate PCDD formation rates with process variables including temperature, precursor concentration, fly ash loading and number of active sites in fly ash. Calculated PCDD formation rates based on this kinetic model are in good agreement with laboratory measurements reported in the literature. When the model is applied to industrial incinerator conditions, at maximum a PCDD yield of 10(-3) microg/N m3 is calculated. PMID- 10864169 TI - Carboxylesterase overexpression in the male reproductive tract: a universal safeguarding mechanism? AB - Data on expression patterns of carboxylesterases in the male reproductive tract of different animal groups (i.e. bivalve mollusks, fruitflies and rodents) are summarized to highlight some particularly interesting questions in the context of sperm differentiation, maturation and function. The male reproductive system, in spite of extreme variation in the anatomical/morphological organization in different species, is characterized by similar patterns of male-dependent carboxylesterase overexpression. The phenomenon of conserved carboxylesterase overexpression indicates similar male sex-associated functions of the enzymes. There is possible evidence of carboxylesterase recruitment by male reproductive tract tissues indicating that it could be adaptive for spermatogenesis, sperm maturation and sperm use. Moreover, this idea can be extended to include a sperm cell lineage protection. This issue is discussed in the light of recent data on environmental reproductive xenobiotics that can provide a basis for a hypothetical explanation of carboxylesterase overexpression in the male reproductive tract. Based on a well-known role of carboxylesterases in detoxification of environmental chemicals such as organophosphate pesticides, it is proposed that various male genital tract carboxylesterases may be characterized by a similar physiological function to protect the male reproductive system against xenobiotic influences that could provoke its dysfunction, thus altering sperm differentiation and maturation. PMID- 10864170 TI - Luteinizing hormone receptors in the bovine corpus luteum during the oestrous cycle and pregnancy. AB - The concentration and affinity of luteinizing hormone (LH) receptors in bovine luteal tissues during the oestrous cycle and pregnancy were investigated by Scatchard analysis of the binding of 125I-labeled human chorionic gonadotropin. Corpora lutea (CL) were classified into five stages of the oestrous cycle and three stages of pregnancy. The concentration of LH receptors sharply increased from the early I stage of the oestrous cycle (Days 2-3; 3.09 fmol mg(-1) protein) to the early II stage (Days 5-6; 9.44 fmol mg(-1) protein) and then remained constant until the late luteal stage (Days 15-17; 8.14-9.56 fmol mg(-1) protein). The LH receptors could not be analysed in the regressed luteal tissue due to the small amounts of binding. There was no significant difference in the concentrations of LH receptors (5.63-9.64 fmol mg(-1) protein) among the three stages of pregnancy. Moreover, the concentrations of the receptors in the CL of pregnancy were comparable to those in the mid-cycle CL. The binding affinity did not change significantly during the oestrous cycle and pregnancy. Based on these results, it is assumed that the luteal function during the entire period of pregnancy might be regulated, at least in part, by LH, which is mediated via its specific receptors, and that the luteal function during pregnancy seems not to be regulated by changes in the binding capacity and affinity of LH receptors. To understand the physiological roles of LH in regulating luteal function in pregnant cows, further studies are required. PMID- 10864171 TI - Effects of unilateral ovariectomy with compensatory hypertrophy on ovarian blood flow, oxygen consumption and progestin secretion in the 16-day pregnant rat. AB - Stimulated ovulation with resultant multiple corpora lutea (CL) can result in lower progesterone levels than expected from the increased luteal tissue mass. Unilateral ovariectomy (ULO) was used to increase CL number in rats and determine whether this would compromise luteal tissue blood flow, oxygen consumption and progestin secretion. All investigations were performed in vivo, using a venous outflow technique on Day 16 of gestation, when progesterone secretion is maximal. ULO, performed before pregnancy, doubled CL number and total CL mass in the remaining ovary of six treated compared to five control rats. Growth of CL was not affected. The rate of ovarian blood flow (microL min(-1) mg CL(-1)) fell to 47% of control levels in ULO animals and progesterone secretion (microg h(-1) mg CL(-1)) to 68%. Secretion of the minor progestin, 20alpha-hydroxypregn-4-en-one was not affected. Tissue oxygen consumption was maintained despite the reduction in blood flow by an increase in oxygen extraction from arterial blood. These results suggest that overcrowding of CL in ULO-stimulated rat ovaries compromises luteal tissue blood flow and subsequently progesterone secretion. PMID- 10864172 TI - Age-dependent development and surface ultrastructural changes following electrical activation of bovine oocytes. AB - Electrical activation of oocytes is a key component of the current mammalian cloning regimen. The present study examined the surface ultrastructure alterations of bovine oocytes at different ages following electrical pulse (EP) activation. In Experiment 1, bovine oocytes 22, 30 and 40 h after in vitro maturation were activated by EP. Significantly higher pronuclear formation (57 or 59%) was obtained for the 30- or 40-h oocytes than for the 22-h oocytes (10%). More oocytes (58%) in the 40-h group activated spontaneously than in the 22- or 30-h groups. In Experiment 2, oocytes were cultured for 1, 2, 4 and 8 h after activation and then evaluated by scanning electron microscopy (SEM). The zona pelucida (ZP) meshwork was completely abolished within 1 h after EP in the majority of the oocytes in the 22- and 30-h groups, but no obvious changes were observed in the 40-h group. From 2 h onward, the ZP meshwork was gradually restored in most treated oocytes. It was also noted that the microvilli pattern on the vitelline surface was altered dramatically after EP treatment, with the proportion affected increasing with oocyte age. The present study extends the previous findings that older oocytes are more readily activated and that the age dependent surface ultrastructural alterations correlate very well with the age dependent activation of oocytes. PMID- 10864173 TI - Reproductive cycles of farmed female chital deer (Axis axis). AB - Studies were conducted on chital deer hinds (Axis axis) living in a temperate region to advance the understanding of the patterns of reproduction of a tropical cervid species. The hinds exhibited regular patterns of oestrus cyclicity throughout the year as evidenced by concentrations of serum progesterone monitored over a 14-month period, and detection of behavioural oestrus by vasectomized stags. The mean length of the oestrous cycle was 18.0+/-0.7 days (range, 12-23 days). Profiles of serum progesterone showed concentrations of <0.5 ng mL(-1) at the time of oestrus, which rose to a peak (range 1.5-5.0 ng mL(-1)) about Day 13, and then declined to low concentrations at the next oestrus. Observations following parturition showed that the first detected oestrus occurred at a mean (+/- SEM) time of 26.9+/-3.0 days later for seven of nine hinds. The mean length of the oestrous cycle after the first post-partum oestrus was 16.6+/-1.0 days (range 7-20 days). The presence of a stag may influence the length of the post-partum period in chital deer hinds, and hinds in contact with a stag in this study had a significantly shorter interval from parturition to first ovulation (P<0.01) compared with hinds not in contact with a stag. By 7 weeks post partum a corpus luteum was detected in 93% of hinds. In comparison only 43% of hinds with no stag contact had a corpus luteum by 7 weeks post partum. It is suggested that the tendency towards seasonal calving in the study population may be related more to male than female factors. PMID- 10864174 TI - Pregnancy in a marsupial, the Tasmanian pademelon (Thylogale billardierii). AB - The Tasmanian pademelon, Thylogale billardierii, is a medium-sized wallaby that adapts well to captivity and, unlike the well-studied tammar wallaby, is capable of breeding all year round. It may, therefore, be a useful model species for research into the reproductive biology of macropod marsupials. This paper presents necessary background data on histological changes in the reproductive organs and the rate of embryonic growth during gestation in T1 billardierii. After Day 4 RPY (removal of young from the pouch) the gravid and non-gravid uteri differ significantly in some histological parameters. The corpus luteum becomes active by Day 6 RPY and is fully developed by Day 14 RPY; it begins to degenerate from Day 19 RPY. Plasma progesterone concentrations through gestation follow a pattern similar to that in the tammar wallaby. There is an early, smaller, peak at Day 5 RPY, with plasma concentrations of progesterone then falling until the larger pre-partum peak occurs several days before birth. PMID- 10864175 TI - Optimal levels of nitric oxide are crucial for implantation in mice. AB - This study was performed to clarify the critical role of optimal levels of nitric oxide on fecundity in mice during the implantation period. Mature female pregnant mice were treated with either nitric oxide donor molsidomine (3, 15, 60 mg kg( 1)) or nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-name; 0.3, 1.5, 6 mg kg(-1)) every 12 h, seven times from the night of Day 2 to Day 5 of gestation. They were killed on Day 14 of gestation. Pregnancy rates in each group (n = 22) and the number of live or absorbed fetuses in each mouse was calculated. The pregnancy rates in the experimental group were reduced in a dose dependent manner. The rate in the control group was 100%, whereas those in the 60 mg molsidomine and 6-mg L-name groups were 40.9 and 31.8%, respectively. Histological analysis of uteri on Day 5 of gestation after treatment with 60 mg molsidomine or 6 mg L-name suggested retarded decidualization of stromal cells or defective function of predecidualized cells. In conclusion, optimal levels of nitric oxide are crucial for endometrial function and embryo implantation. PMID- 10864176 TI - Immunocytochemical localization of oestrogen receptors in perichondrium of antlers in red deer (Cervus elaphus). AB - Remodelling of cancellous bone of antlers to compact bone can be stimulated by administration of oestrogens and previous work has demonstrated the presence of specific oestrogen binding in this tissue. In this study the presence of oestrogen receptors (ER) in antler tissue from red deer males was examined by immunocytochemistry using a monoclonal mouse anti-human ER serum. Strong positive staining was detected in the tip regions of immature antlers and was confined primarily to the cells forming a fibrous layer of the perichondrium. This finding indicates that the effects of oestrogens on remodelling of cancellous bone of the antler may be mediated indirectly by the surrounding connective tissue layers. PMID- 10864177 TI - Working alliance for TB drug development, Cape Town, South Africa, February 8th, 2000. PMID- 10864178 TI - Pyrazinamide is not active against Mycobacterium tuberculosis residing in cultured human monocyte-derived macrophages. AB - SETTING: Mycobacteriology Laboratory, National Jewish Medical and Research Center, Denver, Colorado. OBJECTIVE: To evaluate the antimicrobial activity of pyrazinamide against Mycobacterium tuberculosis in cultured human monocyte derived normal and activated macrophages. DESIGN: Monocytes separated from human blood were incubated in plastic plates for seven days to mature into macrophage monolayers. After activation with TNF-alpha or IFN-gamma or without prior treatment, the macrophages were infected with M. tuberculosis. Various concentrations of pyrazinamide (PZA), morphazinamide (MZA) or isoniazid (INH) were added the next day, and the viable counts of the intracellular bacteria were determined at days 0, 4, and 8. RESULTS: No inhibitory activity of PZA at any concentration was detected, while clear dose-dependent bacteriostatic and bactericidal activities were demonstrated by MZA and INH in the same experimental model. CONCLUSIONS: PZA has neither bacteriostatic nor bactericidal activity against M. tuberculosis persisting or multiplying in cultured monocyte-derived human macrophages, and it might be that the well-known effectiveness of this drug in tuberculosis patients is not related to its supposed activity against intracellular bacterial subpopulations. PMID- 10864179 TI - Patient and disease characteristics, and outcome of treatment defaulters from the Singapore TB control unit--a one-year retrospective survey. AB - SETTING: The Singapore Tuberculosis Control Unit. OBJECTIVES: 1) To identify any demographic, social, disease or treatment-related characteristics which may be predictive of patients defaulting from treatment; 2) to assess the effectiveness of home visits as a means of defaulter recall; 3) to ascertain outcome in these patients. DESIGN: A retrospective, case-controlled study of TB treatment defaulters, defined as patients who missed their scheduled appointments and required a home visit to recall for treatment. Controls were randomly selected, non-defaulting patients who started treatment on the same dates as the defaulters. RESULTS: Forty-four patients required home visits in 1996. Compared to controls, defaulters were more likely to be non-Chinese, and to live on their own or with friends. There was no significant association of defaulting with age, sex, marital or employment status, disease characteristics, or treatment-related factors. Seventy per cent defaulted during the continuation phase of treatment. Home visits did not result in contact with the patient (or any other person) 41% of the time. Although 48% of the defaulters remained lost to follow-up at the time of the survey, all but one of the sputum-positive patients had bacteriologically converted by the time of default. CONCLUSION: Non-Chinese ethnicity and lack of family support were found to be factors strongly predictive of default. Age, sex, marital and employment status, treatment-related factors and disease characteristics were not significant in distinguishing those at risk for defaulting. PMID- 10864180 TI - Twenty-five years of tuberculosis in a French university hospital: a laboratory perspective. AB - OBJECTIVE: To determine the impact of recent changes in the epidemiology of tuberculosis in France and other industrialised countries on the primary trends of tuberculosis case rates in a French university hospital. DESIGN: Descriptive study of all 4549 culture-positive tuberculosis cases hospitalised at Pitie Salpetriere Hospital between 1972 and 1996. RESULTS: From 1972, there was a decline of 5% per year in the tuberculosis case rate, which levelled off in 1983. The proportion of tuberculosis patients who were human immunodeficiency virus (HIV) positive increased from 2% in 1983 to 28% in 1990, and thereafter remained stable. The proportion of foreign-born tuberculosis patients also increased, from 40% in 1972 to 55% in 1985. These two changes affected drug resistance patterns. Drug resistance was more common among foreign-born than among French-born patients, whether previously treated or not. Resistance to rifampicin and multidrug resistance among previously untreated patients was highly related to HIV co-infection. Extrapulmonary sites of tuberculosis were more often smear positive in HIV-positive than in non-HIV-positive patients (22.8% vs 12.6%), and bacteraemia was diagnosed almost exclusively in HIV-positive patients. CONCLUSION: The changes in clinical and bacteriological tuberculosis patterns at the hospital level over the last 25 years have paralleled those observed at national and international level in industrialised countries, including a slowing in the decrease in the case rate, due in part to the HIV epidemic, a higher proportion of foreign-born patients and an increase in drug resistance. PMID- 10864181 TI - Trial-of-antibiotic algorithm for the diagnosis of tuberculosis in a district hospital in a developing country with high HIV prevalence. AB - OBJECTIVE: To evaluate a diagnostic algorithm for pulmonary tuberculosis based on smear microscopy and objective response to trial of antibiotics. SETTING: Adult medical wards, Hlabisa Hospital, South Africa, 1996-1997. METHODS: Adults with chronic chest symptoms and abnormal chest X-ray had sputum examined for Ziehl Neelsen stained acid-fast bacilli by light microscopy. Those with negative smears were treated with amoxycillin for 5 days and assessed. Those who had not improved were treated with erythromycin for 5 days and reassessed. Response was compared with mycobacterial culture. RESULTS: Of 280 suspects who completed the diagnostic pathway, 160 (57%) had a positive smear, 46 (17%) responded to amoxycillin, 34 (12%) responded to erythromycin and 40 (14%) were treated as smear-negative tuberculosis. The sensitivity (89%) and specificity (84%) of the full algorithm for culture-positive tuberculosis were high. However, 11 patients (positive predictive value [PPV] 95%) were incorrectly diagnosed with tuberculosis, and 24 cases of tuberculosis (negative predictive value [NPV] 70%) were not identified. NPV improved to 75% when anaemia was included as a predictor. Algorithm performance was independent of human immunodeficiency virus status. CONCLUSION: Sputum smear microscopy plus trial of antibiotic algorithm among a selected group of tuberculosis suspects may increase diagnostic accuracy in district hospitals in developing countries. PMID- 10864182 TI - HIV testing among tuberculosis patients in the era of antiretroviral therapy: a population-based study in Brazil. AB - SETTING: Rio de Janeiro, Brazil, a city with 29862 cases of tuberculosis (TB) reported between January 1995 and June 1998. OBJECTIVES: To evaluate the counseling and testing practices for human immunodeficiency virus (HIV) infection among TB patients, and to identify the patient characteristics associated with HIV screening as antiretroviral therapy was introduced. DESIGN: Cross-sectional study of patients with TB who were reported to the health department and who initiated anti-TB treatment. The main outcome measure was screened versus not screened for HIV. RESULTS: The proportion of TB patients who received HIV screening increased from January 1995 through June 1998 (P < 0.001). Among young adults aged 20-49 years with TB, the independent predictors of HIV screening were a diagnosis of both pulmonary and extrapulmonary TB (odds ratio [OR] = 2.4, 95% confidence interval [CI] 2.1-2.8); TB meningitis (OR = 13.5, 95%CI 6.5-31.5); disseminated TB (OR = 8.2, 95%CI 5.3-12.9); lymphatic TB (OR = 5.6, 95%CI 4.7 6.6); and male sex (OR = 1.4, 95%CI 1.3-1.6). Patients with newly diagnosed TB who were women, lived in a low income neighborhood (OR = 0.7, 95%CI, 0.6-0.7), and sought TB treatment in their own residential neighborhood (OR = 0.3, 95%CI 0.3-0.4) were less likely to receive HIV counseling and testing. CONCLUSION: Health care providers in Rio de Janeiro selectively offered HIV counseling and testing to persons they perceived to be at risk for HIV and those with advanced stages of TB. HIV counseling and testing should be expanded and offered to all TB patients. PMID- 10864183 TI - Quantitative sputum bacillary load during rifampin-containing short course chemotherapy in human immunodeficiency virus-infected and non-infected adults with pulmonary tuberculosis. AB - SETTING: National Tuberculosis (TB) Treatment Centre, Mulago Hospital and Joint Clinical Research Centre, Kampala, Uganda. OBJECTIVE: To compare the quantitative sputum bacillary load between TB patients infected with the human immunodeficiency virus (HIV) and those non-infected, during treatment with standard short course chemotherapy (SCC). DESIGN: To compare clinical characteristics and quantitative sputum bacillary load as measured by quantitative acid-fast bacilli (AFB) smears, colony forming unit (cfu) assay and time until positive culture in the BACTEC radiometric liquid system between 14 HIV-infected and 22 non-HIV-infected adults with initial episodes of smear positive pulmonary TB at baseline and during treatment with standard four-drug SCC. RESULTS: Other than cavitation (P = 0.042) and adenopathy (P = 0.03), which were more common among non-HIV-infected and HIV-infected patients, respectively, there were no significant differences in baseline demographic, clinical, radiological and laboratory characteristics between the groups. Mean pretreatment sputum bacillary burden (6.5+/-0.51 log10 AFB/ml, 5.91+/-0.91 log10 cfu/ml and 1.8+/-1.7 days until positive BACTEC culture for HIV-infected patients and 6.32+/ 0.85 log10 AFB/ml, 5.58+/-0.68 log10 cfu/ml and 1+/-1.2 days until positive BACTC culture for non-HIV-infected patients) were comparable between HIV-infected and non-HIV-infected patients. Clinical and bacteriological responses to standard SCC and treatment outcome did not differ between the groups. CONCLUSION: Quantitative sputum bacillary load at baseline and during SCC did not differ significantly between HIV-infected and non-HIV-infected adults with initial episodes of smear positive TB. PMID- 10864184 TI - Human immunodeficiency virus-related tuberculosis and primary drug resistance in Bangkok, Thailand. AB - SETTING: Central Chest Hospital, a 500-bed referral hospital near Bangkok with a large out-patient department. OBJECTIVES: To determine human immunodeficiency virus (HIV) seroprevalence among patients with pulmonary tuberculosis (TB), and compare HIV-positive and HIV-negative TB patients. DESIGN: From July 1995 through June 1996, a cross-sectional study was conducted of newly registered adults (> or =16 years old) with suspected pulmonary TB. RESULTS: Of 2587 newly registered patients with suspected pulmonary TB, 2019 (78%) received HIV pretest counseling and 1816 (90%) consented to testing. Of these, 364 (20%) were HIV-seropositive. Among 1091 patients with bacteriologically confirmed TB, HIV seroprevalence was 22%. HIV-positive patients were more likely to be young, unemployed, single men and to have a history of injection drug use. HIV-positive patients with first episode TB were more likely to have Mycobacterium tuberculosis strains resistant to isoniazid (10.9% vs 3.5%; P < 0.001), rifampicin (9.4% vs 2.9%; P < 0.001), and at least isoniazid and rifampicin (multidrug-resistant TB [MDR-TB]; 5.2% vs 0.4%; P < 0.001). CONCLUSIONS: HIV prevalence is high among TB patients at this Bangkok hospital and is associated with drug resistance, including a 12 times higher risk of MDR-TB. These findings underscore the urgent need to assure adherence to complete, effective TB treatment regimens for all patients, including persons who are potentially difficult to manage such as injection drug users. PMID- 10864185 TI - Treatment monitoring and prevalence of drug resistance in tuberculosis patients in Tehran. AB - SETTING: Health care clinics and private practitioners in Tehran. OBJECTIVE: To analyse rates of drug resistance and response to treatment in tuberculosis patients. DESIGN: A prospective study of 257 patients undergoing treatment for whom data were collected on drug susceptibility testing and outcome as well as age, sex and history of treatment. RESULTS: Of 774 initially diagnosed patients, 380 were female and 394 were male; 520 (67%) of the cases had pulmonary disease. The overall rate of primary drug resistance among Mycobacterium tuberculosis isolates resistant to at least one drug was 87/563 (15.5%). Twenty-three patients were multidrug-resistant. Among 215 patients with drug-susceptible cultures recruited for follow-up, rapid response to short-course chemotherapy was observed in 190 (88%) who were successively both smear and culture negative after 2 and 4 months of treatment. After 6 months of treatment, 12 of the 25 patients with slow response to treatment had positive cultures; one was smear-positive. Of the 42 patients with drug-resistant isolates, satisfactory bacteriological response was observed after 6 months of treatment in 30 (71%). CONCLUSIONS: These observations support regional recommendations for short-course treatment regimens. Culture rather than smear result could be a key parameter for individually guiding the duration of treatment in patients with poor response to treatment. PMID- 10864186 TI - A randomised controlled trial of lay health workers as direct observers for treatment of tuberculosis. AB - SETTING: Study conducted in a suburb of Cape Town, South Africa. OBJECTIVE: Comparison of successful tuberculosis treatment outcome rates between self supervision, supervision by lay health worker (LHW), and supervision by clinic nurse. METHODS: Open, randomised, controlled trial with intention-to-treat analysis. RESULTS: All groups (n = 156) achieved similar outcomes (LHW vs. clinic nurse: risk difference 17.2%, 95% confidence interval [CI] -0.1-34.5; LHW vs. self supervision 15%, 95%CI -3.7-33.6). New patients benefit from LHW supervision (LHW vs clinic nurse: risk difference 24.2%, 95%CI 6-42.5, LHW vs. self supervision 39.1%, 95%CI 17.8-60.3) as do female patients (LHW vs. clinic nurse 48.3%, 95%CI 22.8-73.8, LHW vs. self supervision 32.6%, 95%CI 6.4-58.7). CONCLUSIONS: LHW supervision approaches statistically significant superiority, but fails to reach it most likely due to the study's limitation, the small sample size. It is possible that subgroups (new and female patients) do well under LHW supervision. LHW supervision could be offered as one of several supervision options within TB control programmes. PMID- 10864187 TI - Effect of anti-tuberculosis treatment on the tuberculin interferon-gamma response in tuberculin skin test (TST) positive health care workers and patients with tuberculosis. AB - SETTING: Public hospital, Victoria, Australia. OBJECTIVE: To evaluate the effect of multidrug treatment and isoniazid (INH) chemoprophylaxis on the tuberculin interferon-y assay (QIFN) in 19 patients with culture-confirmed Mycobacterium tuberculosis and 119 health care workers (HCWs) with tuberculin skin tests (TST) > or =15 mm. DESIGN: Patients with M. tuberculosis were treated with standard medication and tested with QIFN at diagnosis and at regular intervals over a 12 month period. All HCWs, 59 (50%) of whom were prescribed INH chemoprophylaxis, were tested with QIFN at baseline, 2, 4, 6 and 12 months. RESULTS: QIFN results in patients with tuberculosis were consistent and reproducible. At the initial time point QIFN assays were positive for M. tuberculosis in 67%, and once positive, the QIFN assay remained so over the 12-month period. In the HCWS, initial QIFN assays were positive in 73 (61%). During the 12-month study, 91 HCWs had a QIFN assay on at least two occasions. The overall reproducibility between tests was fair (kappa statistic = 0.45), and was little affected by administration of INH. CONCLUSION: These data suggest that although the QIFN assay is generally positive in patients with proven tuberculosis, it does not provide clinically useful information during the first 12 months of treatment with multidrug chemotherapy or INH chemoprophylaxis. PMID- 10864188 TI - Antibody response to culture filtrate antigens of Mycobacterium tuberculosis during and after treatment of tuberculosis patients. AB - SETTING: Many authors have shown rising titres of anti-mycobacterial antibodies after a few months of anti-tuberculosis treatment. This humoral response might persist for years, making the discrimination between current and old disease difficult. OBJECTIVE: Characterisation of the humoral response to culture filtrates of Mycobacterium tuberculosis before and after treatment of tuberculous patients in order to identify those antigens that could provide information about disease activity. METHODS: Anti-mycobacterial IgG response was determined during and after treatment of tuberculous patients by ELISA and immunoblot. Serum was taken from 71 active tuberculous patients (59 newly acquired and 12 relapse), 15 old tuberculous patients and 45 nontuberculous control subjects. RESULTS: By ELISA, antibody response increased after 2 months of treatment. After chemotherapy was completed, the estimated number of antibodies remained at the same level. The level of specific antibodies in patients seems to reach the same level as that of control subjects 3 years after initiation of treatment. In Western blot, although each patient serum had its own characteristic banding pattern, differences between tuberculous patients and control subjects were found in the area below 20 kDa. Serum from tuberculous patients showed high levels of antibodies at the 14 kDa region. After the beginning of treatment, the intensity of the 14 kDa region band and the percentage of positive recognition tended to decrease. Therefore, one year after initiation of treatment, only seven of 13 cases who demonstrated anti-mycobacterial antibodies in ELISA revealed a mild but still positive reaction at the 14 kDa region; this reactivity disappeared 2 years after initiation of chemotherapy. CONCLUSIONS: The 14 kDa region antigen seems to induce a humoral response that evolves in relation with the disease activity. PMID- 10864189 TI - Efficacy of common antiseptics against mycobacteria. AB - SETTING AND OBJECTIVE: Antiseptics are frequently used to prevent mycobacterial infection; however, the reported activities of a number of antiseptics against mycobacteria are not always consistent. The aim of this study was to determine those antiseptics that are useful against mycobacteria. DESIGN: Evaluation of antiseptic activity against mycobacteria in vitro. RESULTS: The effects of different antiseptics on mycobacteria (Mycobacterium avium, M. kansasii and M. tuberculosis) were examined. At concentrations of 0.05%, povidone-iodine (PVP-I) killed 99% or more of all strains tested within 15 seconds, while 0.5% chlorhexidine gluconate and 0.1% benzalkonium chloride showed no bactericidal activity against mycobacteria. M. kansasii and M. tuberculosis were killed after exposure to cresol for 60 seconds at concentrations of 1.0%, but M. avium was unaffected even after 60 seconds. While M. kansasii and M. tuberculosis were killed by treatment with 2.0% glutaraldehyde for 5 minutes, M. avium was highly resistant to this agent. CONCLUSION: PVP-I seems to be a useful antiseptic against mycobacteria. The measured activity of antiseptics should be interpreted carefully, due to the potential for interference by artifacts. PMID- 10864190 TI - 3'UTR polymorphisms in the NRAMP1 gene are associated with susceptibility to tuberculosis in Koreans. AB - SETTING: Korea University and the Korean Institute of Tuberculosis, Seoul, Korea. OBJECTIVE: To determine whether polymorphisms in the 3' untranslated region (UTR) of the NRAMP1 gene are associated with susceptibility to tuberculosis in Koreans. DESIGN: A case-control study design was used to compare the frequency of 3'UTR of NRAMP1 among 192 tuberculosis patients and 192 healthy individuals. All of the samples were diagnosed by X-ray, smear and culture tests between 1998 and 1999 in the Cross of Lorraine Clinic at the Korean Institute of Tuberculosis. RESULTS: A significant association was found between the Korean tuberculosis patients and polymorphisms in the 3'UTR of the NRAMP1 gene (odds ratio [OR] 1.845; 95% confidence interval [CI] 1.097-3.104; chi2 = 5.424; P = 0.020). CONCLUSION: This study showed that genetic variations in the human NRAMP1 gene are associated with susceptibility to smear-positive tuberculosis in Korean patients. The 3'UTR variant allele associated with susceptibility to tuberculosis is very uncommon in Caucasians, but is present in Koreans and West Africans. These observations may explain in part why African Americans and Koreans have greater susceptibility to tuberculosis than Caucasians. PMID- 10864191 TI - Time between sputum examination and treatment in patients with smear-negative pulmonary tuberculosis. AB - The time between sputum examination and commencement of treatment in patients registered with smear-negative pulmonary tuberculosis (PTB) in four hospitals in Malawi was investigated. Information was obtained in 701 of 887 patients who were registered over a 12-month period between 1997 and 1998: 86% were started on treatment within 3 weeks of sputum examination, 14% were started after a 3-week interval and 6% after a 6-week interval. Such delays are unacceptable, for a number of reasons. Recommendations will go to programme staff to repeat sputum examination in PTB suspects with an abnormal chest radiograph whose negative sputum smears were examined more than 3 weeks previously. PMID- 10864192 TI - DNA fingerprinting of a national sample of Mycobacterium tuberculosis isolates, Botswana, 1995-1996. AB - DNA fingerprinting may be useful to elucidate tuberculosis (TB) transmission in community settings, but its utility is limited if only few fingerprint patterns are observed or band numbers are low. We performed DNA fingerprinting on a national, population-based sample of Mycobacterium tuberculosis isolates from Botswana. During 1995-1996, a random sample of 213 isolates, representing 5% of all smear-positive TB cases, underwent DNA fingerprinting using restriction fragment length polymorphism (RFLP) IS6110 analysis. Eighty-two (38%) of the 213 isolates belonged to one of 18 clusters, with 2-9 isolates/cluster. The median number of bands was 10 (range 1-19); 183 (86%) had six or more bands. Sixty-three (49%) of 128 patients tested were infected with the human immunodeficiency virus (HIV). The degree of RFLP pattern heterogeneity and high band number support the feasibility of a prospective DNA fingerprinting study in Botswana. PMID- 10864193 TI - Study of the in vitro susceptibility of M. tuberculosis to ofloxacin in Spain. Spanish Study Group of M. tuberculosis resistance. AB - The aim of this study was to determine the proportion of antituberculosis ofloxacin resistance among Mycobacterium tuberculosis strain isolates in Spain. Over a period of one month, 213 M. tuberculosis strains collected from 14 different hospitals were studied, including strains both susceptible and resistant to primary antituberculosis drugs. In 28.1% of the strains, a minimum inhibitory concentration (MIC) for ofloxacin of 0.25 microg/ml was obtained; in 43.6% the MIC was 0.5 microg/ml; in 22.06% it was 1 microg/ml; and in 6.1% it was > or =2 microg/ml. Ofloxacin currently seems to be an effective antimicrobial in vitro against susceptible or multiresistant strains of M. tuberculosis in human immunodeficiency virus (HIV)-negative or HIV-positive patients in Spain. PMID- 10864194 TI - Dose-dense chemotherapy for breast cancer: the story so far. PMID- 10864196 TI - Vinorelbine--a clinical review. PMID- 10864195 TI - Immune selection in neoplasia: towards a microevolutionary model of cancer development. AB - The dual properties of genetic instability and clonal expansion allow the development of a tumour to occur in a microevolutionary fashion. A broad range of pressures are exerted upon a tumour during neoplastic development. Such pressures are responsible for the selection of adaptations which provide a growth or survival advantage to the tumour. The nature of such selective pressures is implied in the phenotype of tumours that have undergone selection. We have reviewed a range of immunologically relevant adaptations that are frequently exhibited by common tumours. Many of these have the potential to function as mechanisms of immune response evasion by the tumour. Thus, such adaptations provide evidence for both the existence of immune surveillance, and the concept of immune selection in neoplastic development. This line of reasoning is supported by experimental evidence from murine models of immune involvement in neoplastic development. The process of immune selection has serious implications for the development of clinical immunotherapeutic strategies and our understanding of current in vivo models of tumour immunotherapy. PMID- 10864197 TI - Dose-dense epirubicin and paclitaxel with G-CSF: a study of decreasing intervals in metastatic breast cancer. AB - Anthracyclines and taxanes are very effective drugs in the treatment of advanced breast cancer. With G-CSF support, the dose-intensity of this combination can be increased by reducing the interval between chemotherapy cycles, the so-called 'shortening of cycle time'. We treated 36 patients with advanced breast cancer in a multicentre phase I/II study. The treatment regimen consisted of epirubicin 75 mg m(-2) followed by paclitaxel 135 mg m(-2) (3 h) in combination with G-CSF. At least six patients were treated in each cohort and were evaluated over the first three cycles. Starting at an interval of 14 days, in subsequent cohorts of patients the interval could be shortened to 10 days. An 8-day interval was not feasible due mainly to incomplete neutrophil recovery at the day of the next scheduled cycle. In the 10-day interval cohort it was feasible to increase the paclitaxel dose to 175 mg m(-2). The haematological and non-haematological toxicity was relatively mild. No cumulative myelosuppression was observed over at least three consecutive cycles. In combination with G-CSF, epirubicin 75 mg m(-2) and paclitaxel 175 mg m(-2) could be safely administered every 10 days over at least three cycles, enabling a dose intensity of 52 and 122 mg m(-2) per week, respectively. PMID- 10864198 TI - Feasibility of a dose-intensive CMF regimen with granulocyte colony-stimulating factor as adjuvant therapy in premenopausal patients with node-positive breast cancer. AB - Our aim was to study the feasibility of an intensified intravenous CMF (cyclophosphamide, methotrexate and 5-fluorouracil) schedule with the aim to escalate dose intensity (DI). Twenty-three premenopausal breast cancer patients received 6 cycles of adjuvant CMF intravenously on days 1 and 8 every 3 weeks and granulocyte colony-stimulating factor days 9-18. Endpoints were DI and toxicity. Twenty-one out of 23 patients (91%) received the projected total dose and reached > or =85% of the projected DI. Compared to 'classical' CMF, all patients reached > or = 111% DI. Nine patients received the planned schedule without delay. Thirteen patients (57%) were treated for infection and four patients (17%) were hospitalized for febrile neutropenia. Twelve patients received red blood cell transfusions (52%). Radiation therapy (n = 6) had no adverse impact on dose intensity or haematological toxicity. This dose-intensified CMF schedule was accompanied by enhanced haematological toxicity with clinical sequelae, namely fever, intravenous antibiotics and red blood cell transfusions, but allows a high dose intensity in a majority of patients. PMID- 10864199 TI - A dose-finding study of raltitrexed (tomudex) with cisplatin and epirubicin in advanced gastro-oesophageal adenocarcinoma. AB - The standard treatment for advanced gastro-oesophageal cancer in the UK is epirubicin, cisplatin and continuous infusion 5-fluoruracil by an indwelling central venous catheter (ECF), which has significant morbidity. Raltitrexed (tomudex), a specific inhibitor of thymidylate synthase with a long plasma terminal half-life (50-100 h) has activity in gastro-intestinal tract malignancy. To reduce the Hickman line-associated morbidity of ECF; we have conducted a dose finding study of tomudex combined with epirubicin and cisplatin. Twenty-four patients (22 males, two female), median age 63 years (range 21-75), ECOG performance status < or =2 with histologically proven, unresectable or metastatic gastric (14 patients), gastro-oesophageal junction (nine patients) or oesophageal (one patient) adenocarcinoma received treatment with 3-weekly cisplatin 60 mg m( 2), epirubicin 50 mg m(-2) and tomudex at doses of 2 mg m(-2), 2.5 mg m(-2) or 3 mg m(-2) in successive cohorts. Six patients were treated per dose level with no intra-patient dose escalation. Dose escalation occurred after six patients had completed at least one cycle of chemotherapy at the previous dose level. After defining the maximum tolerated dose a further six patients were treated at the preceding dose level to assess toxicity at the proposed phase II dose. A total of 102 cycles (50% completed 6 cycles) were administered. The dose-limiting toxicities are neutropenia and diarrhoea occurring in 2/6 patients at the 3 mg m( 2) dose level. Of those patients evaluable for response, there were eight partial and one complete response (overall response rate 38%). The median survival was 9.9 months. ECT is an active regimen in oesophagogastric adenocarcinoma. The recommended dose of tomudex for further study in combination with epirubicin and cisplatin is 2.5 mg m(-2). PMID- 10864200 TI - P53 germline mutations in childhood cancers and cancer risk for carrier individuals. AB - The family history of cancer in children treated for a solid malignant tumour in the Paediatric Oncology Department at Institute Gustave-Roussy, has been investigated. In order to determine the role of germline p53 mutations in genetic predisposition to childhood cancer, germline p53 mutations were sought in individuals with at least one relative (first- or second-degree relative or first cousin) affected by any cancer before 46 years of age, or affected by multiple cancers. Screening for germline p53 mutation was possible in 268 index cases among individuals fulfilling selection criteria. Seventeen (6.3%) mutations were identified, of which 13 were inherited and four were de novo. Using maximum likelihood methods that incorporate retrospective family data and correct for ascertainment bias, the lifetime risk of cancer for mutation carriers was estimated to be 73% for males and nearly 100% for females with a high risk of breast cancer accounting for the difference. The risk of cancer associated with such mutations is very high and no evidence of low penetrance mutation was found. These mutations are frequently inherited but de novo mutations are not rare. PMID- 10864201 TI - ATM protein and p53-serine 15 phosphorylation in ataxia-telangiectasia (AT) patients and at heterozygotes. AB - ATM (ataxia-telangiectasia mutated) gene plays a central role in the DNA-damage response pathway. We characterized the ATM protein expression in immortalized cells from AT and AT-variant patients, and heterozygotes and correlated it with two ATM-dependent radiation responses, G1 checkpoint arrest and p53-Ser 15 phosphorylation. On Western blots, the full-length ATM protein was detected in eight of 18 AT cases, albeit at 1-32% of the normal levels, whereas a truncated ATM protein was detected in a single case, despite the prevalence among cases of truncation mutations. Of two ataxia without telangiectasia [A-(T)] cases, one expressed 20% and the other approximately 70% of the normal ATM levels. Noteworthy, among ten asymptomatic heterozygous carriers for AT, normal amounts of ATM protein were found in one and reduced by 40-50% in the remaining cases. The radiation-induced phosphorylation of p53 protein at serine 15, largely mediated by ATM kinase, was defective in AT, A(-T) and in 2/4 heterozygous carriers, while the G1 cell cycle checkpoint was disrupted in all AT and A(-T) cases, and in 3/10 AT heterozygotes. Altogether, our study shows that AT and A( T) cases bearing truncation mutations of the ATM gene can produce modest amounts of full-length (and only rarely truncated) ATM protein. However, this limited expression of ATM protein provides no benefit regarding the ATM-dependent responses related to G1 arrest and p53-ser15 phosphorylation. Our study additionally shows that the majority of AT heterozygotes express almost halved levels of ATM protein, sufficient in most cases to normally regulate the ATM dependent DNA damage-response pathway. PMID- 10864202 TI - Telomerase activity is frequently found in metaplastic and malignant human nasopharyngeal tissues. AB - Telomerase is a specialized ribonucleoprotein polymerase that directs the synthesis of telomere repeats at chromosome ends. Accumulating evidence has indicated that telomerase is stringently repressed in normal human somatic tissues but reactivated in cancers and immortal cells, suggesting that reactivation of telomerase plays an important role in carcinogenesis. In this study, the status of telomerase activity in diseased human nasopharyngeal lesions was determined by the telomeric repeat amplification protocol (TRAP). Fifty-four patients participated including 17 inflammation or hyperplasia, eight with squamous metaplasia, and 29 with different stages of carcinomas. Telomerase activity was detected in 1 of 17 (5.9%) inflammatory or lymphoid hyperplastic tissues, 3 of 8 (37.5%) squamous metaplastic, and 25 of 29 (86.2%) carcinoma tissues. The differences in telomerase expression in these groups is statistically significant (P < 0.001). Levels of telomerase activity correlated with tumour stage (P = 0.024). These results suggest that telomerase reactivation plays a role in the carcinogenesis of nasopharyngeal cancer. Since telomerase activity is found in the majority of nasopharyngeal cancers and a subset of metaplasia, this enzyme may be served as a reference to monitoring the status of abnormal nasopharyngeal tissues. PMID- 10864203 TI - The utility of tumour markers in assessing the response to chemotherapy in advanced bladder cancer. AB - In patients with advanced bladder cancer receiving chemotherapy, early assessment of response can avoid unnecessary toxicity. The aim of this study was to assess the role of tumour markers in monitoring response. Serum levels of one or more of markers beta human chorionic gonadotrophin (betahCG), carcinoembryomic antigen (CEA), CA125 and CA19.9 were measured in 74 patients with advanced bladder cancer receiving chemotherapy from 1992 to 1997. Forty-three of 74 (58%) of patients had at least one raised marker (1.5 times upper limit of normal range). This was more common in patients with extra-pelvic disease than in those with disease confined to the pelvis (P = 0.002). Thirty-eight of 78 (49%) assessable patients had a radiological response. Neither clinical response (P = 0.81) nor survival (P = 0.16) differed between marker-negative and marker-positive patients. Clinical response was strongly related to marker response in the 35 comparable patients (P = 0.0001). No patient had a clinical response without response of at least one marker. Ninety per cent of patients who achieved a marker response had done so by 8 weeks. Monitoring of tumour markers in patients with advanced bladder cancer can help predict the response to chemotherapy. PMID- 10864204 TI - Immunocytochemical assessment of sigma-1 receptor and human sterol isomerase in breast cancer and their relationship with a series of prognostic factors. AB - The purpose of this study was to immunocytochemically investigate two new markers, the sigma-1 receptor and the human sterol isomerase (hSI), in comparison with a series of clinicopathological and immunocytochemical prognostic factors in a trial including 95 patients with operable primary breast cancers. Our results showed no statistically significant relationship between these two markers and the age of the patients, their menopausal status, the tumour size and its histological grade, the nodal status and the expression of the Ki-67 proliferative marker. However, we evidenced a close correlation between the sigma 1 receptor expression and the hormonal receptor positivity (P = 0.008), essentially due to a link with the progesterone receptor status (P = 0.01). By contrast there was an inverse relationship between hSI expression and the oestrogen receptor and/or progesterone receptor positivity (P = 0.098). A significant relationship was shown between both the sigma-1 receptor, hSI expressions and Bcl2 expression, with P= 0.017 and 0.035 respectively. We also assessed whether the expression of the sigma-1 receptor or hSI might be linked with disease-free survival (DFS) and found that the presence of hSI and the absence of sigma-1 receptor expression were associated with a poorer disease-free survival (P= 0.007). Altogether these results suggest that in primary breast carcinomas in association with the evaluation of the steroid receptor status, the sigma-1 receptor and hSI may be interesting new markers useful to identify those patients who might be able to benefit from an adjuvant therapy. PMID- 10864205 TI - Tissue factor, osteopontin, alphavbeta3 integrin expression in microvasculature of gliomas associated with vascular endothelial growth factor expression. AB - Vascular endothelial growth factor (VEGF) is a potent angiogenic factor in human gliomas. VEGF-induced proteins in endothelial cells, tissue factor (TF), osteopontin (OPN) and alphavbeta3 integrin have been implicated as important molecules by which VEGF promotes angiogenesis in vivo. Sixty-eight gliomas were immunohistochemically stained with TF, VEGF, OPN and alphavbeta3 integrin antibody. Twenty-three tumours, six normal brains and nine glioma cell lines were evaluated for their mRNA expression of VEGF and TF by reverse transcription polymerase chain reaction analysis. The data indicated that TF as well as VEGF was a strong regulator of human glioma angiogenesis. First, TF expression in endothelial cells which was observed in 74% of glioblastomas, 54% of anaplastic astrocytomas and none of low-grade astrocytomas, correlated with the microvascular density of the tumours. Double staining for VEGF and TF demonstrated co-localization of these two proteins in the glioblastoma tissues. Second, there was a correlation between TF and VEGF mRNA expression in the glioma tissues. Third, glioma cell conditioned medium containing a large amount of VEGF up-regulated the TF mRNA expression in human umbilical vein endothelial cells. OPN and alphavbeta3 integrin, were also predominantly observed in the microvasculature of glioblastomas associated with VEGF expression. Microvascular expression of these molecules could be an effective antiangiogenesis target for human gliomas. PMID- 10864206 TI - Expression of transcription factor AP-2alpha predicts survival in epithelial ovarian cancer. AB - The 52-kDa activator protein (AP)-2 is a DNA-binding transcription factor which has been reported to have growth inhibitory effects in cancer cell lines and in human tumours. In this study the expression of AP-2alpha was analysed in 303 epithelial ovarian carcinomas by immunohistochemistry (IHC) with a polyclonal AP 2alpha antibody and its mRNA status was determined by in situ hybridization (ISH) and reverse transcriptase-polymerase chain reaction (RT-PCR). The immunohistochemical expression of AP-2alpha was correlated with clinicopathological variables, p21/WAF1 protein expression and survival. In normal ovaries, epithelial cells expressed AP-2alpha protein only in the cytoplasm. In carcinomas nuclear AP-2alpha expression was observed in 28% of the cases although cytoplasmic expression was more common (51%). The expression of AP 2alpha varied according to the histological subtype and differentiation. AP 2alpha and p21/WAF1 expressions did not correlate with each other. Both in univariate (P = 0.002) and multivariate analyses (relative risks (RR) 1.6, 95% confidence interval (CI) 1.13-2.18, P= 0.007) the high cytoplasmic AP-2alpha expression favoured the overall survival. In contrast, the nuclear AP-2alpha expression combined with low cytoplasmic expression increased the risk of dying of ovarian cancer (RR = 2.10, 95% CI 1.13-3.83, P= 0.018). The shift in the expression pattern of AP-2alpha (nuclear vs cytoplasmic) in carcinomas points out to the possibility that this transcription factor may be used by oncogenes in certain histological subtypes. Based on the mRNA analyses, the incomplete expression and translation of AP-2alpha in ovarian cancer may be due to post transcriptional regulation. PMID- 10864207 TI - Enhancement of chemotherapy and radiotherapy of murine tumours by AQ4N, a bioreductively activated anti-tumour agent. AB - AQ4 (1,4-Bis-[[2-(dimethylamino-N-oxide)ethyl]amino]5,8-dihydroxyanthrace ne-9, 10-dione) is a prodrug designed to be excluded from cell nuclei until bioreduced in hypoxic cells to AQ4, a DNA intercalator and topoisomerase II poison. Thus, AQ4N is a highly selective bioreductive drug that is activated in, and is preferentially toxic to, hypoxic cells in tumours. Five murine tumours (MAC16, MAC26, NT, SCCVII and RIF-1) have been used to investigate the anti-tumour effects of AQ4N. In only one tumour (MAC16) was AQ4N shown to be active as a single agent. However, when combined with methods to increase the hypoxic tumour fraction in RIF-1 (by physical clamping) and MAC26 tumours (using hydralazine) there was a substantial enhancement in anti-tumour effect. Notably, RIF-1 tumours treated with AQ4N (250 mg kg(-1)) followed 15 min later by physically occluding the blood supply to the tumour for 90 min, resulted in a 13-fold increase in growth delay. When combined with radiation or chemotherapy, AQ4N substantially increased the effectiveness of these modalities in a range of in vivo model systems. AQ4N potentiates the action of radiation in both a drug and radiation dose-dependent manner. Further the enhancement observed is schedule-independent with AQ4N giving similar effects when given at any time within 16 h before or after the radiation treatment. In combination with chemotherapy it is shown that AQ4N potentiates the activity of cyclophosphamide, cisplatin and thiotepa. Both the chemotherapeutic drugs and AQ4N are given at doses which individually are close to their estimated maximum tolerated dose (data not included) which provides indirect evidence that in the combination chemotherapy experiments there is some tumour selectivity in the enhanced action of the drugs. PMID- 10864208 TI - Induction of apoptosis and activation of the caspase cascade by anti-EGF receptor monoclonal antibodies in DiFi human colon cancer cells do not involve the c-jun N terminal kinase activity. AB - We previously reported that exposure of DiFi human colon cancer cells to the anti epidermal growth factor (EGF) receptor monoclonal antibody (mAb) 225 resulted in apoptosis, but the mechanisms remain to be elucidated. In the present study, we investigated the effects of a panel of four anti-EGF receptor mAbs, each of which binds to different epitopes of the EGF receptor in DiFi cells, on the induction of apoptosis. We found that each of these mAbs induced apoptosis in DiFi cells. Exposure of DiFi cells to mAb 225 activated the initiation caspase-8, which was detectable between 8 and 16 h after exposure of the cells to the antibody. There was also an activation of the initiation caspase-9, which lagged a few hours behind the activation of caspase-8. Exposure of DiFi cells to mAb 225 also activated the execution caspase-3, which was accompanied temporally by evidence of cleavage of a well-characterized caspase-3 substrate, poly(ADP)ribosepolymerase (PARP). Pre-exposure of the cells to the caspase-3 specific inhibitor DEVD-CHO partially reduced the mAb 225-induced PARP cleavage and apoptosis, whereas pre-exposure of the cells to the caspase pan-inhibitor z VAD-fmk completely inhibited mAb 225-induced apoptosis. Caspases-3, -8 and -9 were not activated in the cell lines in which mAb 225 only induced G1 phase arrest of the cell cycle. In contrast to the apoptosis of DiFi cells induced by ultraviolet irradiation, which strongly activated the c-jun N-terminal kinase-1 (JNK1) and the caspase cascade, mAb 225-induced apoptosis and activation of the caspase cascade in DiFi cells were not associated with activation of JNK1. PMID- 10864209 TI - Prostaglandin E2 and the protein kinase A pathway mediate arachidonic acid induction of c-fos in human prostate cancer cells. AB - Arachidonic acid (AA) is the precursor for prostaglandin E2 (PGE2) synthesis and increases growth of prostate cancer cells. To further elucidate the mechanisms involved in AA-induced prostate cell growth, induction of c-fos expression by AA was investigated in a human prostate cancer cell line, PC-3. c-fos mRNA was induced shortly after addition of AA, along with a remarkable increase in PGE2 production. c-fos expression and PGE2 production induced by AA was blocked by a cyclo-oxygenase inhibitor, flurbiprofen, suggesting that PGE2 mediated c-fos induction. Protein kinase A (PKA) inhibitor H-89 abolished induction of c-fos expression by AA, and partially inhibited PGE2 production. Protein kinase C (PKC) inhibitor GF109203X had no significant effect on c-fos expression or PGE2 production. Expression of prostaglandin (EP) receptors, which mediate signal transduction from PGE2 to the cells, was examined by reverse transcription polymerase chain reaction in several human prostate cell lines. EP4 and EP2, which are coupled to the PKA signalling pathway, were expressed in all cells tested. Expression of EP1, which activates the PKC pathway, was not detected. The current study showed that induction of the immediate early gene c-fos by AA is mediated by PGE2, which activates the PKA pathway via the EP2/4 receptor in the PC-3 cells. PMID- 10864210 TI - Effects of nicotinamide and carbogen on tumour oxygenation, blood flow, energetics and blood glucose levels. AB - Both host carbogen (95% oxygen/5% carbon dioxide) breathing and nicotinamide administration enhance tumour radiotherapeutic response and are being re evaluated in the clinic. Non-invasive magnetic resonance imaging (MRI) and 31P magnetic resonance spectroscopy (MRS) methods have been used to give information on the effects of nicotinamide alone and in combination with host carbogen breathing on transplanted rat GH3 prolactinomas. Gradient recalled echo (GRE) MRI, sensitive to blood oxygenation changes, and spin echo (SE) MRI, sensitive to perfusion/flow, showed large signal intensity increases with carbogen breathing. Nicotinamide, thought to act by suppressing the transient closure of small blood vessels that cause intermittent tumour hypoxia, induced a small increase in blood oxygenation but no detectable change in perfusion/flow. Carbogen combined with nicotinamide was no more effective than carbogen alone. Both carbogen and nicotinamide caused significant increases in the nucleoside triphosphate/inorganic phosphate (betaNTP/Pi) ratio, implying that the tumour cells normally receive sub-optimal substrate supply, and is consistent with either increased glycolysis and/or a switch to more oxidative metabolism. The most striking observation was the marked increase in blood glucose (twofold) induced by both nicotinamide and carbogen. Whether this may play a role in tumour radiosensitivity has yet to be determined. PMID- 10864211 TI - Failure in post-transcriptional processing is a possible inactivation mechanism of AP-2alpha in cutaneous melanoma. AB - The loss of transcription factor AP-2alpha expression has been shown to associate with tumourigenicity of melanoma cell lines and poor prognosis in primary cutaneous melanoma. Altogether these findings suggest that the gene encoding AP 2alpha (TFAP2A) acts as a tumour suppressor in melanoma. To learn more of AP 2alpha's down-regulation mechanisms, we compared the immunohistochemical AP 2alpha protein expression patterns with the corresponding mRNA expression detected by in situ hybridization in 52 primary melanomas. Of the 25 samples with AP-2alpha protein negative areas, 16 (64%) expressed mRNA throughout the consecutive section. Nine specimens (36%) contained equally mRNA- and protein negative areas, suggesting that the loss of AP-2alpha protein associated with lack of the mRNA transcript. The highly AP-2alpha protein-positive tumours (n = 27) were concordantly mRNA positive in 25 (92.6%) cases. Thirteen primary tumours were further analysed using microsatellite markers D6S470 and D6S263 for loss of heterozygosity (LOH) of a locus harbouring TFAP2A. LOHs or chromosome 6 monosomy were found in four out of five (80%) informative AP-2alpha mRNA- and protein negative tumour areas, but also within five out of 13 (38%) informative AP-2alpha mRNA-positive tumour areas. This chromosome region is thus suggestive of harbouring a putative tumour suppressor gene of cutaneous melanoma, but this referring specifically to TFAP2A could not be completely verified in this analysis. We conclude that a failure in post-transcriptional processing of AP 2alpha is a possible inactivation mechanism of AP-2alpha in cutaneous melanoma. PMID- 10864212 TI - Pre-induction LDH as a prognostic factor for outcome of high dose chemotherapy (HDCT) for germ cell tumours relapsing or refractory to conventional chemotherapy. PMID- 10864213 TI - A possible cause of testicular cancer. PMID- 10864214 TI - Cardiovascular emergencies in the cancer patient. AB - Cardiovascular emergencies in oncology patients include all of the usual cardiac problems, as well as complications of cancer and its therapy. Pericardial effusions and tamponade, cardiac masses, and extrinsic compression of the heart and great vessels by tumor masses, or fluid collections may all occur. Certain tumors may secrete mediators that are directly toxic to the heart; for example, catecholamines are secreted by pheochromocytomas and serotonin is secreted by carcinoid tumors. Tumors can also cause arrhythmias due to the mediators they secret or to direct mechanical irritation of the heart or pericardium. Cancer therapy is also associated with cardiac emergencies. Perioperative myocardial ischemia or infarction, as well as arrhythmias, may complicate surgery. Pericardial effusions and tamponade can follow surgery, radiation, or chemotherapy. Chemotherapy with anthracyclines, mitoxantrone, and trastuzumab may prompt acute and chronic heart failure. 5-Fluorouracil causes coronary spasm in some patients, leading to angina, myocardial infarction, arrhythmias, and/or sudden death. Cyclophosphamide, particularly in high doses, may produce acute myopericarditis. Radiation may cause acute pericardial disease and late sequelae such as myocardial infarction, acute valvular insufficiency, or effusive constrictive pericarditis. Endocarditis also occurs in cancer patients in association with vascular access devices and immune compromise. This review will discuss each of these complications of cancer and its therapy. PMID- 10864215 TI - Respiratory emergencies. AB - Respiratory emergencies may originate from disease in the airways, thoracic vessels, and pulmonary parenchyma. Airway obstruction may be amenable to bronchoscopic therapies, including laser ablation photodynamic therapy (PDT) and stent placement. Asthma is common, but may be mimicked by endobronchial metastasis. Superior vena cava syndrome (SVCS) is seen most commonly with bronchogenic carcinoma and lymphoma. Emergent treatment need not precede tissue diagnosis in the absence of associated tracheal obstruction. Pulmonary embolism (PE) may now be diagnosed with spiral computed tomography (CT), but ventilation perfusion scintigraphy remains the first-line test. Parenchymal lung disease may result from infections, with neoplastic and iatrogenic etiologies. The incidence of Pneumocystis carinii pneumonia (PCP) is increasing among cancer patients, but it can be prevented by prophylaxis. Attempts to treat adult respiratory distress syndrome (ARDS) through modification of inflammatory mediators have been disappointing, and the prognosis remains poor. PMID- 10864216 TI - Gastrointestinal emergencies in the critically ill cancer patient. AB - Gastrointestinal (GI) problems are a common occurrence in the critically ill cancer patient. These GI complications are often not related to the underlying cancer, but mimic the GI problems seen in noncancer patients. Common GI disorders such as peptic ulcer disease, gastritis, diverticulitis, and alcohol-related liver disease are often the underlying issue. This review summarizes the most common GI emergencies, which may arise in a patient with a current or past history of cancer. PMID- 10864217 TI - Urologic emergencies in the cancer patient. AB - Urologic emergencies are common in the cancer patient and relate mainly to complications of bladder hemorrhage, upper or lower urinary tract obstruction, urinary tract infection, and priapism. Hemorrhagic cystitis is commonly due to bladder injury from radiation therapy, viral infection, or metabolites of chemotherapeutic agents. Treatments aimed at ameliorating the effects of theses metabolites, such as mesna and intravenous (IV) hydration, coupled with cystoscopy, evacuation of clots, and formalin instillation, have given clinicians an effective means of avoiding exsanguinating hemorrhage from the bladder. Malignant ureteral obstruction is an ominous sign in the cancer patient and may be due to tumor compression, retroperitoneal adenopathy, or direct tumor invasion. The endourologic procedures of ureteral stenting and percutaneous nephrostomy are effective means of palliation; however, complications of infection, stent obstruction, and stent migration can result in hospital admission and a decline in quality of life. Median survival for patients with malignant ureteral obstruction is less than 7 months, regardless of the tumor of origin. Bladder outlet obstruction leading to urinary retention can be due to mechanical factors involving the bladder neck or prostate, or to a breakdown in the neurophysiologic function of the bladder. Every attempt is made to avoid surgical intervention or the placement of chronic in-dwelling catheter in these often debilitated patients. Patients are often effectively treated with a variety of pharmacologic agents, such as alpha-adrenergic receptor blockers or by the initiation of chronic intermittent catheterization. Urinary tract infections are particularly dangerous in neutropenic and bone marrow transplant patients, with bladder catheters the most common portal entry. The colonization and later infection by resistant nosocomial organisms, such as Pseudomonas aeruginosa and Candida albicans, can rapidly lead to life-threatening sepsis. On rare occasions, emergency surgical intervention with adequate open drainage or nephrectomy is required to control such infections. Priapism can be caused by hematologic malignancy with hypercoagulation, metastatic disease involving the corpora cavernosa with thrombosis of the venous outflow from the penis, or rarely from intracavernous injections used for the treatment of impotence. If effective treatment exists for the primary tumor, improvement or resolution of the state of priapism may occur. Radiation therapy may be required to decrease the pain associated with malignant priapism, but surgical shunting procedures are rarely effective. PMID- 10864218 TI - Orthopedic emergencies in cancer patients. AB - The most frequent orthopedic emergency in oncology patients is fracture. Stabilization of the entire fractured bone restores function and relieves pain. The site, quality, and extent of the lesion can identify impending fractures that should be stabilized. New methods of pelvic stabilization effectively bypass periacetabular bone deficiency. Spinal cord decompression is important to maintain neurologic function. Advances in segmental fixation of the spine have improved the outcome over what was achieved by radiation alone. Infection is common in neutropenic patients, and should be treated aggressively with antibiotics and drainage of abscesses of the musculoskeletal system. Extravasation of doxorubicin requires prompt local debridement to limit the extent of necrosis propagation. These treatments can effectively improve the quality of life of patients with metastatic cancer. They should be included as "best supportive care" for patients with more than 1 month to live. PMID- 10864219 TI - Neurologic emergencies in the cancer patient. AB - Neurologic complications of cancer and its therapy are varied and common, but there are few true neurologic emergencies. However, when a neurologic emergency does occur, rapid diagnosis and treatment can preserve neurologic function and, in some circumstances, save a life. Epidural spinal cord compression, raised intracranial pressure (ICP), status epilepticus, and intracerebral hemorrhage (ICH) are the most common neurologic emergencies in the cancer patient. This chapter details the clinical features, possible etiologies, diagnostic tests, and treatment options for each of these complications. PMID- 10864220 TI - Metabolic emergencies in the cancer patient. AB - The acute tumor lysis syndrome (ATLS) is characterized by the rapid development of hyperuricemia, hyperkalemia, hyperphosphatemia, and acute renal failure (ARF). Hematologic malignancies are responsible for most cases of ATLS. Control of hyperuricemia and the achievement of a high urine flow are the mainstays of prevention. Urinary alkalinization should be performed only when hyperuricemia is present. Hypercalcemia occurs in 10% to 20% of patients with cancer at some time during the disease course. Parathyroid hormone-related protein (PTHrP) is the most common mediator of humoral hypercalcemia of malignancy (HHM), while local osteolysis is the principal mechanism in patients with bone metastasis. Hydration with saline and administration of pamidronate control hypercalcemia in most patients. Hyponatremia with an increase in total-body salt and water content, manifested as edema and/or ascites, is the most common electrolyte abnormality in cancer patients. Hyponatremia due to salt depletion may occur in patients who receive cisplatin. The syndrome of inappropriate antidiuretic hormone secretion (SIADH) may occur in association with cancer of the lung, after high-dose cyclophosphamide, and during vigorous fluid administration in patients with chemotherapy-associated emesis. Lactic acidosis without tissue hypoperfusion may be seen in patients with extensive liver metastasis or with certain hematologic malignancies. In the latter cases, lactate levels parallel disease activity and chemotherapy often leads to resolution of the lactic acidosis. Idiopathic hyperammonemia has been described after intensive chemotherapy for hematological malignancies and following bone marrow transplantation. PMID- 10864221 TI - Infectious complications in the critically ill patient with cancer. AB - Significant morbidity and mortality may result from infectious complications in patients with neoplastic disease. Clinical oncologists and infectious disease specialists recognize the emergent nature of these infections in the critically ill patient with cancer. It is essential to have an approach to the understanding of these infections and the circumstances in which they occur. Fever and neutropenia, pneumonia, CNS infections, and gastrointestinal infections are all discussed in this review. PMID- 10864222 TI - Systemic therapy emergencies. AB - Systemic oncologic therapies can cause multiple emergency situations. There are, however, two unique emergencies directly related to chemotherapy administration: drug extravasation and hypersensitivity reactions (HSRs). Most drugs can cause varying degrees of local tissue injury when extravasated. The medical management of extravasation is based on proper maintenance of the intravenous line, application of local cooling or warming for certain extravasations, and the use of antidotes to prevent the local toxic action of the extravasated drug. Antidotes that appear useful include hyaluronidase (for vinca alkaloids, epipodophyllotoxins, and paclitaxel), sodium thiosulfate (for mechlorethamine and cisplatin), and dimethylsulfoxide (DMSO) (for anthracyclines and mitomycin C). HSRs, another potential adverse effect of chemotherapy administration, are a major concern for therapy with taxanes and with L-asparaginase, and their administration requires the use of premedication to prevent these reactions. Once a HSR has occurred, therapy may be continued by using an analog drug or by the administration of premedication as prophylaxis, in particular if the reaction was minor. On the other hand, it is also pertinent to become acquainted with the emergencies induced by biological agents, taking into consideration their increasing usages. In addition to interferons and interleukin-2 (IL-2), both of which have been in clinical use for several years, cytokine-toxin fusion proteins (DAB3891L-2) and two monoclonal antibodies (rituximab and trastuzumab) were recently approved for cancer therapy. The distinct toxicity profiles of these agents are reviewed. PMID- 10864223 TI - Thrombosis and cancer. AB - Thrombosis is a common complication of malignancy. This is felt to be related to increased activity of the coagulation system as evidenced by markers of accelerated thrombin generation and increased platelet reactivity. Alterations in the hemostatic balance have been documented in patients with malignancy with increased tissue factor (TF) generation and the production of a cysteine protease. These can stimulate the coagulation mechanism via the extrinsic pathway and/or by activating factor X. The thrombotic presentations in malignancy are protean and may be venous or arterial. The underlying clinical pictures may be related to varying degrees of consumptive coagulopathy, microangiopathy, and nonbacterial endocarditis. Prophylaxis and management are, to a significant degree, dependent on the underlying malignancy and the prothrombotic mechanism. Specific agents and drugs must be selected from an expanding menu of options that includes unfractionated heparin, low-molecular-weight heparin (LMWH), warfarin, plasma apheresis, and the newer antithrombin agents. PMID- 10864224 TI - Hematologic supportive care of the critically ill cancer patient. PMID- 10864225 TI - Preclinical perspectives on platinum resistance. AB - In the 30 years since the introduction of cisplatin into the clinic, laboratory studies have provided considerable information as to both how the drug exerts its antitumour effects and how some tumours are, or become, resistant. Once inside a cell, the chlorine groups of cisplatin are exchanged for water (aqua) species, which are more chemically reactive. The intracellular target for cisplatin is DNA, where a variety of adducts are formed, some on the same strand of DNA (intrastrand adducts) and others between strands (interstrand adducts). Of the 4 bases, guanine is the preferred site for binding and the most common adduct involves linkages on 2 adjacent guanines on the same strand of DNA. It remains uncertain which of the various types of adduct is the most important in terms of producing antitumour effects. Resistance to cisplatin has been studied extensively using tumour cells repeatedly exposed to the drug in vitro. In these cell models, resistance is generally due to a combination of mechanisms, some resulting in reduced damage to DNA and others following DNA damage. Resistance due to inadequate binding to DNA has been shown to be caused by reduced drug uptake (influx rather than efflux) and inactivation by thiol-containing species such as glutathione and metallothioneins. Resistance occurring post-DNA binding may be due to changes in DNA repair pathways [an increase in nucleotide excision repair (NER) or a loss of DNA mismatch repair (MMR)]. Conversely, the hypersensitivity of some cell lines to cisplatin has been shown to be due to defective NER, through loss or reduced expression of NER proteins such as XPG and XPA. Resistance may also be mediated through alterations in proteins involved in programmed cell death (apoptosis) such as p53 and the BCL2 family. A basic understanding of cisplatin resistance pathways has made a major impact in the development of new platinum analogues capable of circumventing resistance. Examples (which are now undergoing clinical trial) include ZD0473 (which, relative to cisplatin, possesses a reduced reactivity towards inactivating thiol containing molecules) and the trinuclear platinum BBR3464 (which has markedly different DNA binding properties compared with cisplatin). PMID- 10864226 TI - Clinical perspectives on platinum resistance. AB - The platinum compounds cisplatin and carboplatin are widely used in the treatment of a number of solid malignancies. Although some platinum-sensitive tumours may be cured by combination chemotherapy (e.g. testicular cancer), most will relapse and subsequently prove resistant to platinum compounds. The mechanisms of platinum resistance in patients are still poorly understood. Clearly, when a tumour relapses a long time after successful first-line treatment, there is a high chance that it will still be sensitive to platinum compounds. A number of studies have attempted to assess the role of drug transport, the glutathione system, DNA repair and apoptosis genes in the development of resistance in tumours, but no conclusive evidence is available. Approaches to increasing the potency of platinum therapy (to overcome resistance) have been devised and some have proved to be effective; in particular, intraperitoneal administration of cisplatin has shown superiority over intravenous administration in selected patients with ovarian cancer. The development of drugs and techniques to reduce the adverse effects of platinum chemotherapy has greatly improved their administration. Investigations attempting to modulate platinum activity and toxicity have also been performed. Further investigation of in vivo resistance mechanisms should be valuable in allowing prediction of clinical response to chemotherapy and may identify new treatments with the potential to improve outcomes for patients with a variety of platinum-resistant tumour types. PMID- 10864227 TI - Clinical pharmacokinetics and administration of established platinum drugs. AB - We review the pharmacology and clinical administration of the commonly used platinum-based anticancer drugs cisplatin and carboplatin, and the more recently approved diamminocyclohexane-based oxaliplatin. The development of analogues of cisplatin has been focused upon identifying compounds with less toxicity and with a different spectrum of activity. Carboplatin exemplifies the former, while the initial data with oxaliplatin support its activity in cisplatin-resistant tumours. The clinical pharmacokinetics of the drugs are reviewed. Incorporation of these data into the design of clinical regimens has permitted individualised therapy with carboplatin, and has enhanced safety. Additional investigation of the pharmacodynamics of all of these agents is expected to result in their selective application. The clinical effects of these analogues are discussed. PMID- 10864228 TI - New developments and approaches in the platinum arena. AB - Following the introduction of cisplatin and the demonstration of its importance in the treatment of testicular and ovarian cancer, there was a need to develop less toxic analogues. Compared with cisplatin, carboplatin proved markedly less toxic to the kidneys and nervous system and caused less nausea and vomiting, while generally (and certainly for ovarian cancer) retaining equivalent antitumour activity. In many situations, carboplatin is now the drug of choice in view of the improved quality of life it offers patients. Many drug combinations involving platinum complexes have been explored, but those with taxanes are particularly noteworthy. Paclitaxel in combination with a platinum agent is now accepted as a standard component of first-line treatment for ovarian cancer, and produces improved survival. Preclinical studies suggested that drugs containing the diaminocyclohexane ligand would be capable of overcoming intrinsic or acquired resistance. However, this outcome was not realised in the clinic until the development of oxaliplatin, which appears to have a different spectrum of activity compared with cisplatin and carboplatin. Oxaliplatin improves the response rate and progression-free survival when given with fluorouracil for the treatment of advanced colorectal cancer, and its activity in other tumour types is under investigation. ZD0473 is a platinum analogue that relies on steric hindrance to overcome thiol-mediated detoxification. It has a good tolerability profile, is currently undergoing phase II testing, and its activity in combination with other agents is being explored. The trinuclear platinum complex BBR3464 also looks promising in preclinical studies and will shortly be evaluated in phase II trials. Although much research remains to be done, these new developments in platinum-based chemotherapy should translate into significant improvements in treatment for patients with a broad range of tumour types. PMID- 10864229 TI - ERPs and PET analysis of time perception: spatial and temporal brain mapping during visual discrimination tasks. AB - ERPs were recorded from 12 subjects performing duration and intensity visual discrimination tasks which have been previously used in a PET study. PET data showed that the same network was activated in both tasks [P. Maquet et al., NeuroImage 3:119-126, 1996]. Different ERP waveforms were observed for the late latency components depending on the dimension of the stimulus to be processed: frontal negativity (CNV) for the duration task and parieto-occipital positivity (P300) for the intensity task. Using BESA software, the sources were first modelled with a "PET dipolar model" (right prefrontal, right parietal, anterior cingulate, left and right fusiforms). To obtain a better fit for ERPs recorded in each task, two sources (cuneus, left prefrontal area) had to be added. Consistently with PET findings, dipole modelling indicates that duration and intensity dimensions of a visual stimulus are processed in the same areas. However, ERPs also reveal prominent differences between the time course of the dipole activations for each task, particularly for sources contributing to the late latency ERP components. In the intensity task, dipoles located in the cuneus, the anterior cingulate, and the left prefrontal area yield largest activity within the P300 interval, then activity diminishes rapidly as the stimulus ends, whereas in the duration task, the cuneus and anterior cingulate are still active several hundred milliseconds following stimulus offset. Moreover, in the duration task, the activity of the right frontal dipole parallels the CNV waveform, whereas in the intensity task, this dipole is largely inactive. We assume that the right frontal area plays a specific role in the formation of temporal judgments. PMID- 10864230 TI - A simulator for evaluating methods for the detection of lesion-deficit associations. AB - Although much has been learned about the functional organization of the human brain through lesion-deficit analysis, the variety of statistical and image processing methods developed for this purpose precludes a closed-form analysis of the statistical power of these systems. Therefore, we developed a lesion-deficit simulator (LDS), which generates artificial subjects, each of which consists of a set of functional deficits, and a brain image with lesions; the deficits and lesions conform to predefined distributions. We used probability distributions to model the number, sizes, and spatial distribution of lesions, to model the structure-function associations, and to model registration error. We used the LDS to evaluate, as examples, the effects of the complexities and strengths of lesion deficit associations, and of registration error, on the power of lesion-deficit analysis. We measured the numbers of recovered associations from these simulated data, as a function of the number of subjects analyzed, the strengths and number of associations in the statistical model, the number of structures associated with a particular function, and the prior probabilities of structures being abnormal. The number of subjects required to recover the simulated lesion-deficit associations was found to have an inverse relationship to the strength of associations, and to the smallest probability in the structure-function model. The number of structures associated with a particular function (i.e., the complexity of associations) had a much greater effect on the performance of the analysis method than did the total number of associations. We also found that registration error of 5 mm or less reduces the number of associations discovered by approximately 13% compared to perfect registration. The LDS provides a flexible framework for evaluating many aspects of lesion-deficit analysis. PMID- 10864231 TI - Lateralized automatic auditory processing of phonetic versus musical information: a PET study. AB - Previous positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) studies show that during attentive listening, processing of phonetic information is associated with higher activity in the left auditory cortex than in the right auditory cortex while the opposite is true for musical information. The present PET study determined whether automatically activated neural mechanisms for phonetic and musical information are lateralized. To this end, subjects engaged in a visual word classification task were presented with phonetic sound sequences consisting of frequent (P = 0.8) and infrequent (P = 0.2) phonemes and with musical sound sequences consisting of frequent (P = 0.8) and infrequent (P = 0.2) chords. The phonemes and chords were matched in spectral complexity as well as in the magnitude of frequency difference between the frequent and infrequent sounds (/e/ vs. /o/; A major vs. A minor). In addition, control sequences, consisting of either frequent (/e/; A major) or infrequent sounds (/o/; A minor) were employed in separate blocks. When sound sequences consisted of intermixed frequent and infrequent sounds, automatic phonetic processing was lateralized to the left hemisphere and musical to the right hemisphere. This lateralization, however, did not occur in control blocks with one type of sound (frequent or infrequent). The data thus indicate that automatic activation of lateralized neuronal circuits requires sound comparison based on short-term sound representations. PMID- 10864232 TI - Hemispheric preference in visuospatial processing: a complementary approach with fMRI and lesion studies. AB - Historically, the left cerebral hemisphere has been considered specialized for language, whereas the right cerebral hemisphere is aligned with spatial processes. However, studies have called into question adherence to this model and suggested that both hemispheres participate in language and spatial cognition. Using functional Magnetic Resonance Imaging (fMRI) and human brain lesion studies, we determined whether these complementary techniques could clarify issues of hemispheric dominance. Using a modified Benton Judgement of Line Orientation (JLO) test, considered a relatively pure spatial processing task, we found robust and significant (p < 0.0005) bilateral superior parietal lobe activation on fMRI in ten right-handed male adult volunteers. This was corroborated by lesion data in a cohort of 17 patients who showed significant JLO impairments after either right or left parietal lobe damage, with right parietal damage associated with somewhat more severe deficit. Detailed wavelet analysis of the fMRI time-series did, however, reveal a more dominant role of the right parietal lobe in "kick-starting" the task. To our knowledge, this is a novel way of using fMRI to address functional hemispheric differences in a cognitive task that is known to have bilateral representation. PMID- 10864233 TI - Effects of attention on dichotic listening: an 15O-PET study. AB - The present study investigated the effect of attention on brain activation in a dichotic listening situation. Dichotic listening is a technique to study laterality effects in the auditory sensory modality. Two different stimuli were presented simultaneously, one in each ear. Twelve subjects listened to lists of consonant-vowel syllables, or short musical instrument passages, with the task of detecting a "target" syllable or musical instrument by pressing a button. The target stimulus appeared an equal number of times in the left and right ear. The subjects were instructed to either concentrate on the stimuli presented in both ears, or only on the left or right ear stimulus. Brain activation was measured with 15O-PET, and significant changes in regional normalized counts (rNC) were evaluated using statistical parametric mapping (SPM96) software. Concentrating on either the right or left ear stimulus significantly decreased activity bilaterally in the temporal lobes compared to concentrating on both ear stimuli, at the expense of an increased activation in the right posterior and inferior superior parietal lobe. The CV-syllables activated areas corresponding to the classic language areas of Broca and Wernicke. The musical instrument stimuli mainly activated areas in visual association cortex, cerebellum, and the hippocampus. An interpretation of the findings is that attention has a facilitating effect for auditory processing, causing reduced activation in the primary auditory cortex when attention is explicitly recruited. The observed activations in the parietal lobe during the focused attention conditions could be part of a modality non-specific "attentional network". PMID- 10864234 TI - Differentiation of cuticular structures during the growth of the third-stage larva of Ascaris suum (Nematoda, Ascaridoidea) after emerging from the egg. AB - In order to monitor the early phases of the development of Ascaris suum from domestic pigs, third-stage larvae, retrieved from the liver and the lungs, were studied by analyzing worm growth and length increase of individual transverse annuli in the cuticle. Material for study using light and scanning electron microscopy was obtained from experimental infections. The results show that the third-stage larva (not the second-stage) after emergence from the egg grows continuously, without an ecdysis in the liver. During growth, each annulus is split into a complex of 2 subannuli, each of which attains a bimodal appearance and is a prominent feature during a late phase of the third-stage larva. The results suggest that the first 2 molts occur inside the egg, a synapomorphic feature of the Ascaridoidea. The third-stage larvae of ascaridoids, with some functional similarities of the dauer-larva stage of Caenorhabditis sp., facilitate transmission of these parasitic worms to the digestive tract of the vertebrate final host (utilizing the tracheal route in A. suum), where the third and the fourth molts take place. PMID- 10864235 TI - Endogenous development of Eimeria minasensis in experimentally infected goats. AB - The endogenous development of Eimeria minasensis was studied in 9 coccidia-free goat kids inoculated with 10(5) sporulated oocysts/kg body weight. Kids were killed 4, 7 (2 animals), 10, 13, 16, 18, 19, and 22 days after inoculation (DAI). In tissue sections of the intestines stained with hematoxylin and eosin and examined by light microscopy, 2 generations of meronts, gamonts, gametes, and oocysts were found. The first generation of meronts developed in cells deep in the lamina propria of the jejunum and ileum. Mature giant meronts (299.4x243.8 microm) found 16 DAI were visible to the naked eye and contained a large number of crescent-shaped merozoites. The second generation of meronts developed in the epithelial cells of crypts of the ileum and above the host cell nuclei. Mature meronts (11.5x10.1 microm) with 18-28 comma-shaped merozoites were first seen 16 DAI. Gametogenesis took place in epithelial cells of the crypts and villi of the terminal part of the ileum, cecum, and colon. Macrogametes (27.8x17.6 microm), mature microgamonts (21.3x17.0 microm), microgametes, and oocysts (30.5x19.4 microm) were found 19 DAI. Sexual stages were below the host cell nucleus. PMID- 10864236 TI - The flea genus Sigmactenus (Siphonaptera: Leptopsyllidae): a new species from timor and new material from New Guinea and the Philippines. AB - Both sexes of Sigmactenus timorensis n. sp. are described from Rattus tanezumi and Rattus exulans collected in West Timor, Indonesia. The true host presumably is a native murine rat such as the extant, endemic Rattus timorensis or 1 of several extinct, endemic Timorese rats. Analyses of new collections of Sigmactenus cavifrons and Sigmactenus toxopeusi from New Guinea demonstrate that these fleas show considerable morphological variation. We propose that they represent a single species with the name S. toxopeusi having priority. New collections of Sigmactenus werneri from the Philippines expand the known hosts and geographical distribution of this flea. PMID- 10864237 TI - Competence of the African tortoise tick, Amblyomma marmoreum (Acari: Ixodidae), as a vector of the agent of heartwater (Cowdria ruminantium). AB - The ability of the African tortoise tick, Amblyomma marmoreum, to acquire and transmit Cowdria ruminantium infection was investigated experimentally with transmission trials and with a C. ruminantium-specific polymerase chain reaction (PCR) detection assay. Laboratory-reared A. marmoreum larvae and nymphs were fed on small ruminants with clinical heartwater. After molting, the resultant nymphs were fed on Cowdria ruminantium-naive sheep (n = 3), and the adults were ground and inoculated intravenously into sheep (n = 5). Fatal heartwater developed in the 5 recipient animals, demonstrating larvae-nymph transmission and nymph-adult acquisition of infection. Cowdria ruminantium infection was also detected in adult A. marmoreum by PCR analysis, although at lower frequency (10%) than in Amblyomma hebraeum ticks (43%), the major vector of C. ruminantium in southern Africa, which had been fed simultaneously on the infected animals (P<0.0001). Amblyomma marmoreum, therefore, can be an effective vector of C. ruminantium. The potential role of this species in heartwater epidemiology and in the spread of the disease to new areas is highlighted by these results and by the fact that immature stages of this tick feed readily on domestic and wild animals susceptible to C. ruminantium. PMID- 10864238 TI - Genetic and morphological heterogeneity in small rodent whipworms in southwestern Europe: characterization of Trichuris muris and description of Trichuris arvicolae n. sp. (Nematoda: Trichuridae). AB - Genetic and morphological variability of whipworms Trichuris Roederer, 1761 (Nematoda: Trichuridae), parasites of small rodents in southwestern Europe, was studied. Isozyme patterns of natural populations of nematodes parasitizing rodent species of the Muridae (Apodemus sylvaticus, Apodemus flavicollis, Mus musculus) and Arvicolidae (Clethrionomys glareolus, Microtus agrestis, Microtus arvalis) were analyzed at 6 putative loci. Two diagnostic loci were found in T. muris from Muridae and from Arvicolidae. Thus, the existence of 2 species of Trichuris restricted to different host families was indicated. They included Trichuris muris Schrank, 1788, originally described as being from mice, and Trichuris arvicolae n. sp., parasitizing the above species of Arvicolidae. The morphological variability of both species was compared. Although ranges of all morphological characters of the new species overlapped with those of T. muris, stepwise discriminant analysis yielded a 100% accurate classification of females when using vagina length and egg size. Males of T. muris and T. arvicolae cannot be separated entirely. A set of 6 variables yielded 95.7% discrimination; the most discriminating variables were spicule size and body width. PMID- 10864239 TI - Intraerythrocytic development of species of Hepatozoon infecting ranid frogs: evidence for convergence of life cycle characteristics among apicomplexans. AB - Intraerythrocytic development of the adeleorin apicomplexans Hepatozoon clamatae and Hepatozoon catesbianae were investigated in the bullfrog, Rana catesbeiana, the green frog, Rana clamitans melanota, and the Northern leopard frog, Rana pipiens. Merozoites emerging from hepatic meronts penetrated erythrocytes and underwent 1-3 rounds of binary fission to produce 2-8 merozoites. Following their release from infected erythrocytes, individual merozoites entered new cells and transformed into gamonts. Although this is the first report of intraerythrocytic development for a fully described species of Hepatozoon, a phylogenetic reanalysis of 11 species of Hepatozoon, 6 species representative of the 5 other hemogregarine taxa, 2 species of dactylosomatids, and 2 species of piroplasms, indicates that asexual reproduction of parasites within blood cells of vertebrates has arisen at least 3 times in the apicomplexan lineage that includes adeleorins and piroplasms. This method of asexual development, which is also observed in species of hemospororin genera such as Plasmodium, is discussed in the context of the evolution of apicomplexan life cycles. In addition to supporting the paraphyly of the genus Hepatozoon determined in an earlier study, this phylogenetic analysis featured a monophyletic group, consisting of the sister taxa Hemolivia and Karyolysus, that was the sister group to a clade consisting of the more derived hemogregarines, the dactylosomatids, and the piroplasms. PMID- 10864240 TI - Use of recombinant antigens for detection of Toxoplasma gondii infection in swine. AB - Five recombinant Toxoplasma gondii antigens, designated B427, C51, C55, V22, and MBP30 were assessed for their potential use in an enzyme-linked immunoassay (EIA) for detection of T. gondii infection in swine. The antigens were evaluated with sera from young pigs that had been fed 1-10,000 T. gondii oocysts of the VEG or GT-1 strains. Results were compared with an EIA using a native T. gondii antigen extract. All 5 recombinant antigens, as well as native antigen, detected antibody responses as soon as 3 wk after infection in pigs inoculated with 1 or 10 oocysts of the VEG strain. This antibody response persisted, at varying levels, for 14 wk when the experiment was terminated. All antigens also detected antibody responses in pigs 4 wk after inoculation with 10,000 oocysts of the GT-1 strain. The antibody response recognized by native antigen remained high through 51 wk after inoculation. However, there was considerable animal-to-animal variation in responses to the individual recombinant antigens. Only antigens C51 and MBP30 consistently detected a positive antibody response over the entire 51-wk course of the experiment. These results suggest that these antigens might be useful for the serological detection of T. gondii infection in pigs. PMID- 10864241 TI - Developmental arrest and physical entrapment eliminates supernumerary Ganaspis xanthopoda parasitoids in Drosophila melanogaster. AB - Superparasitism is a common condition in which a single host is injected with more than 1 parasitoid egg, but only 1 parasite survives to adulthood, and the remaining animals are eliminated. Here, for the first time, we show that supernumerary Ganaspis xanthopoda, endoparasitoids that invade Drosophila melanogaster, are physiologically suppressed during embryonic development. Whereas the suppressed supernumerary embryos can develop to the first larval instar, their subsequent growth is blocked because they become physically trapped within a novel multicellular envelope that is formed during late embryogenesis. Supernumerary embryos can produce this envelope when cultured in vitro even if they are separated from dominant embryos. Our results suggest that physiological suppression programs supernumerary individuals for developmental arrest, starvation, and necrotic or apoptotic death. PMID- 10864242 TI - Venereal worms: sexually transmitted nematodes in the decorated cricket. AB - The nematode, Mehdinema alii, occurs in the alimentary canal of the decorated cricket Gryllodes sigillatus. Adult nematodes occur primarily in the hindgut of mature male crickets, whereas juvenile nematodes are found in the genital chambers of mature male and female crickets. Here, we present experimental evidence for the venereal transmission of M. alii in G. sigillatus. Infectivity experiments were conducted to test for transmission via oral-fecal contamination, same-sex contact, and copulation. The infective dauers of the nematode are transferred from male to female crickets during copulation. Adult female crickets harboring infective dauers subsequently transfer the nematode to their next mates. Thus, M. alii is transmitted sexually during copulation. PMID- 10864243 TI - The life cycle of Monorchis parvus (Digenea: Monorchiidae) demonstrated by developmental and molecular data. AB - Cercaria cerastodermae I, a digenean parasite of Cerastoderma edule, was recorded for the first time in the Atlantic Ocean off the Iberian peninsula. Sporocysts were present in the hemolymph of the digestive gland, gonad, gills, and foot of the mollusc. Most of the cercariae present within sporocysts were encysted as metacercariae. The corresponding adult stages were obtained after experimental infection of several Diplodus sargus artificially reared in fish farms and that had previously been protected against natural infections. Numerous adult specimens of Monorchis parvus were collected in Diplodus annularis along the French Mediterranean coast. Comparison of wild and experimental adults allowed the adult stage of Cercaria cerastodermae I to be identified as M. parvus. Another monorchid, Monorchis monorchis, a parasite of Spondyliosoma cantharus, was found in the same Mediterranean area and compared with M. parvus. Additionally, ITS1 nuclear ribosomal DNA sequences of C. cerastodermae I and of the adults collected in naturally infected D. annularis and S. cantharus were obtained. Sequence data indicate that C. cerastodermae I corresponds to the adult of M. parvus found in D. annularis and is clearly distinct from M. monorchis found in S. cantharus. PMID- 10864244 TI - First findings of Cryptosporidium and Giardia in California sea lions (Zalophus californianus). AB - We report the detection and identification of Cryptosporidium and Giardia from 1 of 3 species of pinnipeds. Fecal samples were collected from Pacific harbor seal (Phoca vitulina richardsi), northern elephant seal (Mirounga angustirostris), and California sea lion (Zalophus californianus) in the northern California coastal area. By means of fluorescently labeled monoclonal antibodies, Cryptosporidium oocysts were detected in 3 samples from California sea lions, 1 of which also contained Giardia cysts. Oocysts of Cryptosporidium and cysts of Giardia were morphologically indistinguishable from oocysts of C. parvum and cysts of G. duodenalis from other animal origins. Oocysts and cysts were then purified using immunomagnetic separation techniques and identified by polymerase chain reaction (PCR), from which species-specific products were obtained. Sequence analysis revealed that the 452-bp and 358-bp PCR products of Cryptosporidium isolated from California sea lion had identities of 98% with sequences of their template fragments of C. parvum obtained from infected calves. Based on morphological, immunological, and genetic characterization, the isolates were identified as C. parvum and G. duodenalis, respectively. The findings suggested that California sea lions could serve as reservoirs in the environmental transmission of Cryptosporidium and Giardia. PMID- 10864245 TI - Natural transmission of Cryptosporidium parvum between dams and calves on a dairy farm. AB - The transmission of Cryptosporidium parvum between dams and their respective calves was studied. For this purpose, fecal specimens taken from the rectum of preparturient, parturient, and postparturient dams were analyzed for C. parvum oocysts. Fecal specimens were taken from the newborn calf 4 hr after birth. Because the environment can be a source of contamination to the animals, specimens taken from inside and outside the barn were analyzed. The sucrose concentration method together with the Zielh-Nielsen acid-fast staining method were employed to increase the chances of oocyst detection. We are reporting that at parturition, the dams shed a higher number of oocysts by comparison to the preparturient and postparturient periods. Neonates acquire the infection at birth mainly because of the high number of oocysts shed by the dams at parturition. The management practice of moving calves 4 hr after birth away from the dams and the barn reduces the number of clinical cases because they are no longer in contact with an environment that is highly contaminated. We hypothesize that the increase in the number of oocysts sheds by dams at parturition might be due to a depression of the T helper 1-type of immune response during that period. PMID- 10864246 TI - Life cycle of Calyptospora funduli (Apicomplexa: Calyptosporidae). AB - The taxonomic status of the extraintestinal piscine coccidium Calyptospora funduli is based in part on its requirement of an intermediate host (the daggerblade grass shrimp Palaemonetes pugio). In the present study, grass shrimp fed livers of Gulf killifish (Fundulus grandis) infected with sporulated oocysts of C. funduli exhibited numerous sporozoites suspended in the intestinal contents when fresh squash preparations were examined by light microscopy. Using this method, sporozoites were not seen in intestinal epithelial cells of the grass shrimp or in any other cell types. Ultrastructural examination, however, revealed sporozoites in the cytoplasm of the gut basal cells. Cross-sections of 1-13 sporozoites were seen within a single cell, and those sporozoites each appeared to be situated in individual membrane-bound vesicles, rather than in a single parasitophorous vacuole. These ultrastructural observations indicate that in the grass shrimp intermediate host, sporozoites that develop into an infective stage probably undergo that development in gut mucosal basal cells. Prior studies revealed that these sporozoites modified their structure over 4-5 days and that before that time, they were not infective to the fish host. Following ingestion of an infected shrimp by a killifish, the infective sporozoites apparently reach the liver of their killifish definitive hosts through the bloodstream. Sporozoites were seen in blood smears from the longnose killifish, Fundulus similis, 4 hr after fish were fed experimentally infected grass shrimp. Additionally, coccidian trophozoites and early meronts were seen in hepatocytes from several longnose killifish at 48, 72, and 96 hr postinfection. This study, in conjunction with previous findings, clearly confirms that a true intermediate host is required in the life cycle of C. funduli, that a developmental period of about 5 days in grass shrimp is necessary for sporozoites to become infective to killifishes, and that sporozoites do occur intracellularly in gut basal cells of the grass shrimp. PMID- 10864247 TI - Survey and host fitness effects of red-cockaded woodpecker blood parasites and nest cavity arthropods. AB - Blood parasites and nest cavity arthropods associated with the red-cockaded woodpecker (Picoides borealis) were surveyed and the impact of blood-feeding arthropods on woodpecker fitness traits was assessed. Five woodpeckers (8%) were infected with unidentified microfilariae, and 1 woodpecker (2%) was infected with 2 species of haemoproteid (Haemoproteus velans and Haemoproteus borgesi). This is the first record of haemoproteids in this species and the first observation of H. borgesi in North America. We collected representatives of at least 6 families of mites and 12 families of primarily commensal insects from woodpecker cavities. Only a few specimens of blood-feeding insects were recovered. The mite Androlaelaps casalis was the most common hematophagous arthropod (prevalence = 76%, mean density = 51+/-7 mites/cavity). The number of A. casalis mites increased with cavity age but there was no association between the number of mites and the number of woodpecker eggs laid or the number of hatchlings or fledglings. In conclusion, the prevalence of blood parasites in the red-cockaded woodpecker is low, woodpecker cavities are not heavily infested with blood feeding insects, and there is no evidence that A. casalis mites affect woodpecker fitness. PMID- 10864248 TI - Prevalence of malaria parasites (Plasmodium floridense and Plasmodium azurophilum) infecting a Puerto Rican lizard (Anolis gundlachi): a nine-year study. AB - The prevalence of malaria parasites was studied in the lizard Anolis gundlachi over a 9-yr period at a site in the wet evergreen forest of eastern Puerto Rico. Three forms of the parasite infected the lizards; these were Plasmodium floridense, Plasmodium azurophilum in erythrocytes, and P. azurophilum in white blood cells. Overall prevalence of infection for 8 samples during the study period was significantly higher for males than females (32% of 3,296 males and 22% of 1,439 females). During the study, the site experienced substantial climatic and physical disturbance including rising temperature, droughts, and hurricanes that severely damaged the forest. Parasite prevalence in the first sample, 8 mo after the massive hurricane Hugo, was slightly, though significantly, lower than for subsequent samples. However, overall prevalence was stable during the 9-yr period. The results show malaria prevalence is more constant at the site than found for 2 studies in temperate forests, and that the Puerto Rico system may be an example of the stable, endemic malaria described by standard models for human malaria epidemiology. PMID- 10864249 TI - Isolation and molecular cloning of a secreted immunosuppressant protein from Dermacentor andersoni salivary gland. AB - A 36-kDa immunosuppressant protein (Da-p36) was isolated from salivary glands of feeding female ixodid ticks Dermacentor andersoni, using its affinity for UltraLink Biosupport Medium (Pierce, Rockford, Illinois)/protein complexes. Using a nested set of forward degenerate oligonucleotide primers corresponding to Da p36 N-terminal amino acids, a cDNA encoding the immunosuppressant protein was isolated by 3' rapid amplification of cDNA ends. The resulting 772-base pair cDNA encodes a novel protein with predicted molecular weight of 24.9 kDa. Sequence analysis revealed the presence of 5 potential glycosylation sites and 1 myristylation site. Immunoblot analyses showed native Da-p36 is present in salivary glands and saliva from both male and female D. andersoni but not in salivary glands or saliva from Amblyomma americanum or Ixodes scapularis. Reverse transcription polymerase chain reaction and immunoblot analyses showed that Da p36 expression is temporally regulated in salivary glands with maximum mRNA levels preceding maximum Da-p36 accumulation that occurred at day 6 of feeding. The levels of Da-p36 mRNA and protein were greatly reduced in salivary glands from near-replete females removed from sheep after 8 days of feeding. These data are consistent with a role of Da-p36 in immunosuppression during feeding. PMID- 10864250 TI - Biological and molecular characterizations of Toxoplasma gondii strains obtained from southern sea otters (Enhydra lutris nereis). AB - Toxoplasma gondii was isolated from brain or heart tissue from 15 southern sea otters (Enhydra lutris nereis) in cell cultures. These strains were used to infect mice that developed antibodies to T. gondii as detected in the modified direct agglutination test and had T. gondii tissue cysts in their brains at necropsy. Mouse brains containing tissue cysts from 4 of the strains were fed to 4 cats. Two of the cats excreted T. gondii oocysts in their feces that were infectious for mice. Molecular analyses of 13 strains indicated that they were all type II strains, but that they were genetically distinct from one another. PMID- 10864251 TI - Is stage conversion the initiating event for reactivation of Toxoplasma gondii in brain tissue of AIDS patients? AB - Reactivation of chronic toxoplasmosis resulting in Toxoplasma encephalitis (TE) is a common event in acquired immune deficiency syndrome (AIDS) patients. Conversion from Toxoplasma gondii bradyzoites to tachyzoites is a prerequisite for reactivation. Until recently, the study of stage conversion in human tissue was not possible due to the lack of antibodies that recognize stage-specific epitopes after long-term formaldehyde fixation. Using the combination of a polyclonal anti-T. gondii antibody, the cyst-stage-specific monoclonal antibody CC2, and a tachyzoite-specific polyclonal antibody (anti-SAG1, recombinant), we tried to demonstrate parasite differentiation in the brain tissue of 10 AIDS patients with clinically suspected TE. Double labeling of the stage-specific antibodies enabled us to demonstrate interconversion between tachyzoites and bradyzoites for the first time in human tissue. The study confirmed that the transformation process is nonsynchronous and that the manifestation of TE depends on the degree and site of tissue destruction caused by invading tachyzoites. The original source of tachyzoites could never be located, but a few samples suggested that tachyzoites may invade by dissemination across the blood-brain barrier. Cyst rupture as the first event in the process of reactivation was not seen. We conclude that the initial site(s) of reactivation will be destroyed by tissue-destructive tachyzoites long before clinical symptoms occur. PMID- 10864252 TI - ITS-2 rDNA sequencing of Gnathostoma species (Nematoda) and elucidation of the species causing human gnathostomiasis in the Americas. AB - From several gnathostome species the complete internal transcribed spacer ITS-2 ribosomal DNA (rDNA) repeat sequence and a fragment of the 5.8S rDNA were obtained by direct polymerase chain reaction cycle-sequencing and silver-staining methods. The size of the complete ITS-1 sequence in agarose gel electrophoresis was also obtained. The ITS-2 enabled the differentiation of Gnathostoma spinigerum from Thailand and Gnathostoma binucleatum from Mexico and Ecuador and confirmed the validity of the latter. Gnathostoma turgidum, Gnathostoma sp. I (=Gnathostoma procyonis sensu Almeyda-Artigas et al., 1994), and Gnathostoma sp. II (=G. turgidum sensu Foster, 1939 pro parte), all from Mexico, proved to be independent species, but Gnathostoma sp. III, also from Mexico, could not be differentiated from G. turgidum. In Mexico and Ecuador, gnathostomes involved in human infection and that had been classified as G. spinigerum belong to G. binucleatum. The 5.8S rDNA sequences of the 6 Gnathostoma species studied were identical. The results of the ITS-1 agreed with those results of ITS-2. PMID- 10864253 TI - Phylogeny of species of the genus Litomosoides (Nemata [corrected]: Onchocercidae): evidence of rampant host switching. AB - Filarioid nematodes of the genus Litomosoides occur in the abdominal and (or) thoracic cavities of marsupials, rodents, and bats of the Nearctic and Neotropical regions. In this study, the phylogenetic relationships among these nematodes were estimated with a parsimony analysis of morphological characters derived from species descriptions. This nonweighted analysis produced 20 shortest trees. The monophyly of the genus was not supported in that Litomosoides thomomydis and Litomosoides westi failed to group with the other members of the genus. When these 2 taxa (parasites of pocket gophers) were excluded, monophyly of Litomosoides was supported by 2 synapomorphies (structure of the walls and general shape of the stoma); however, ancestor-descendant relationships among the species in the genus were not well resolved. A posteriori reweighting of the characters produced a single tree, different from all 20 most parsimonious trees. Alternative host-parasite evolutionary models were tested against these results supporting the process of host switching as being most important in forming the patterns of mammal-nematode associations that have been detected in this group of nematodes. PMID- 10864254 TI - Hedruris hanleyae n. sp. (Nematoda: Hedruridae) from Hemidactylus garnotii (Sauria: Gekkonidae) from the Cook Islands, Oceania. AB - Hedruris hanleyae n. sp. (Nematoda: Hedruridae) from the stomach of Hemidactylus garnotii collected in 1989 on Atiu, Cook Islands is described and illustrated. Hedruris hanleyae n. sp. represents the 21st species to be assigned to the genus and is distinguished from other oriental species by the distribution pattern of caudal papillae of the male: 10 pairs posterior subventral papillae; 2 pairs precloacal and 8 pairs postcloacal. PMID- 10864255 TI - Stylocephalus occidentalis n. sp. (Apicomplexa: Eugregarinida: Stylocephalidae) from Trimytis pruinosa (Coleoptera: Tenebrionidae) in the Nebraska Sandhills. AB - Stylocephalus occidentalis n. sp. (Apicomplexa: Eugregarinida) is described from Trimytis pruinosa (Coleoptera: Tenebrionidae) collected from Keith County in the Sandhills of western Nebraska. Measurements are means in micrometers. Developing trophozoites solitary; epimerite a complex of terminal epimerite and intercalating diamerite; epimerite shallowly ovoid to transversely elliptoid, with transverse basal constriction at junction with diamerite, length 0.5-1 times width, approximately 3-4 times that of diamerite; width approximately equal to that of diamerite; diamerite roughly cylindrical to spindle-shaped, without significant anterior taper, little or no evidence of longitudinal folds, length approximately twice width. Association late, frontal, isogamontic. Gamont protomerite depressed ovoid to very broadly ovoid, length 27.3, width 35.1, anterior distance to widest point 15.4. Protomerite-deutomerite septum clearly marked and constricted, width 34.6. Deutomerite often with distinct marginal crenulation, narrowly obovoid to very narrowly obovoid, length 356.5, maximum width 57.6, anterior distance to widest point 26.3, equatorial width 35.1, +/ 12.5, 29. Total length 381.5. Nucleus ellipsoid, length 32.5, width 18.8; with 0 or 2 polysomal endosomes. Gametocysts roughly spherical; diameter 205.0; wall desiccating to become paper-like, slightly papillated, dehiscing by simple rupture, releasing oocysts in coiled chains, epispore packet absent, gametocyst residuum present. Oocysts dark brown to black, axially asymmetric, broadly deltoid, gibbous in lateral aspect, slightly keeled in dorsal aspect; length 9.8, height 7.9; with slight terminal protuberances and 2 central, spherical residua. PMID- 10864256 TI - Sequence and secondary structure variation in the Gyrodactylus (Platyhelminthes: Monogenea) ribosomal RNA gene array. AB - Nucleotide sequences were determined for the rRNA internal transcribed spacers 1 and 2 (ITS1 and 2) and the 5' terminus of the large subunit rRNA in selected Gyrodactylus species. Examination of primary sequence variation and secondary structure models in ITS2 and variable region V4 of the small subunit rRNA revealed that structure was largely conserved despite significant variation in sequence. ITS1 sequences were highly variable, and models of structure were unreliable but, despite this, show some resemblance to structures predicted in Digenea. ITS2 models demonstrated binding of the 3' end of 5.8S rRNA to the 5' end of the large subunit rRNA and enabled the termini of these genes to be defined with greater confidence than previously. The structure model shown here may prove useful in future phylogenetic analyses. PMID- 10864258 TI - Kamegainema cingulum (Linstow, 1902) n. gen., n. comb. (Nematoda: Dracunculidae), a subcutaneous parasite of cryptobranchids (Amphibia: Caudata). AB - The monotypic Kamegainema n. gen. is proposed for Filaria cingula, a subcutaneous parasite of cryptobranchids. Examination of female specimens recently collected from the Japanese giant salamander, Andrias japonicus, revealed that it belongs to Dracunculidae. Kamegainema is closest to Protenema Petter and Planelles, 1986, the only other dracunculid genus known from Amphibia, but is readily distinguished by having prominent cuticular bosses. Kamegainema cingulum is redescribed. PMID- 10864257 TI - Attempts to establish experimental Cyclospora cayetanensis infection in laboratory animals. AB - Attempts were made to develop an animal model for Cyclospora cayetanensis to identify a practical laboratory host for studying human cyclosporiasis. Oocysts collected from stool of infected humans in the United States, Haiti, Guatemala, Peru, and Nepal were held in potassium dichromate solution to allow development of sporozoites. The following animal types were inoculated: 9 strains of mice, including adult and neonatal immunocompetent and immune-deficient inbred and outbred strains, rats, sandrats, chickens, ducks, rabbits, jirds, hamsters, ferrets, pigs, dogs, owl monkeys, rhesus monkeys, and cynomolgus monkeys. Most animals were inoculated by gavage, although some of the primates were fed oocysts on food items. The animals were examined for signs of infection, particularly diarrhea, and stool samples were examined for 4-6 wk after inoculation. None of the animals developed patent infections or signs of infection. We conclude that none of the animals tested is susceptible to infection with C. cayetanensis. PMID- 10864259 TI - Relationships of Nematodirus species and Nematodirus battus isolates (Nematoda: Trichostrongyloidea) based on nuclear ribosomal DNA sequences. AB - Nuclear ribosomal sequence data from the internal transcribed spacers (ITS-1 and ITS-2), 5.8S subunit, and regions of the 18S and 28S genes were used to investigate sequence diversity among geographic samples of Nematodirus battus, and to infer phylogenetic relationships among Nematodirus species. Phylogenetic analysis of these data yielded strong support for relationships among species, depicting Nematodirus helvetianus and Nematodirus spathiger as sister-taxa and a clade of these 2 species and Nematodirus filicollis. This tree is consistent with caprine bovids as ancestral hosts, with a subsequent host shift to Bovinae in N. helvetianus. Eleven of 14 N. battus sequences were unique, with 19 variable sites among sequences representing 5 geographic samples. The lowest number of variable nucleotide sites was observed in samples representing apparently recent introductions to the United States and Canada, which is consistent with a population bottleneck concomitant with translocation. Comparison of directly sequenced polymerase chain reaction products and clones revealed evidence for intraindividual variation at some of the sequence sites, and this pattern of variation and that within geographic samples indicates incomplete rDNA repeat homogenization within species. This pattern of variation is not conducive for inferring phylogenetic relationships among sequences representing N. battus or addressing the putative history of introduction. PMID- 10864260 TI - Host specificity in Metamera sillasenorum, n. sp., a gregarine parasite of the leech Helobdella triserialis with notes on transmission dynamics. AB - Eugregarines of the suborder Septatorina are apicomplexan parasites that are found mainly in arthropods. Some exceptions are species in the Metameridae that contains the only 5 septate gregarines recorded from annelids. The type genus is Metamera Duke, 1910 with 2 species, Metamera schubergi Duke, 1910, in European Glossiphonia complanata, and Metamera reynoldsi Jones, 1943, from North American G. complanata. Over the summers of 1995-1998, in Keith County, Nebraska, septate gregarines were found in the glossiphoniid leech Helobdella triserialis. The gregarines were determined to be a new species of Metamera, herein named Metamera sillasenorum. Measurements of size and body proportions of over 600 gregarines and 50 oocysts showed differences from measurements of M. schubergi and M. reynoldsi, and secondary septa in the deutomerite were rarely observed. Field observations indicated that M. sillasenorum is probably host specific. In the laboratory, leeches also exhibited strong feeding preferences; e.g., H. triserialis and G. complanata consumed only snails, whereas Helobdella stagnalis consumed only oligochaetes. Infection experiments demonstrated that freshwater snails ingest the oocysts and are required as mechanical vectors. Oocysts were passed unaltered through the snails' intestines. Glossiphonia complanata did not become infected regardless of heavy exposure to oocysts, although only 5 G. complanata were used in the experiments. The results show that host specificity of M. sillasenorum is most likely due to a combination of host-feeding habits and host-parasite compatibility. PMID- 10864261 TI - Effect of dinitroaniline herbicides on the growth of Entamoeba histolytica. AB - The effect of the dinitroaniline herbicides oryzalin and trifluralin on the growth of Entamoeba histolytica was examined. Oryzalin inhibited the growth of E. histolytica strain HM-1:IMSS. Trifluralin was less effective than oryzalin for this parasite. Entamoeba histolytica was more resistant to these dinitroanilines than other parasitic protozoa examined so far, including Leishmania spp., Trypanosoma brucei, Plasmodium falciparum, Toxoplasma gondii, and Cryptosporidium parvum. Colchicine, a potent microtubule inhibitor of animal cells, was much less effective for E. histolytica, even at very high concentrations. A reptilian parasite, Entamoeba invadens strain IP-1, examined for comparison, was more resistant to these dinitroanilines than E. histolytica. Accumulation of E. histolytica trophozoites in mitosis was observed after culture in 100 microM oryzalin. The inhibitory effect of oryzalin on the growth of E. histolytica trophozoites was abrogated by removal of the drug after exposure to 100 microM for 2 days. In parallel to the recovery of growth after removal of the drug, the percentage of trophozoites in mitosis was reduced to a normal level. The results indicate that treatment of trophozoites with oryzalin arrests mitosis and that its effect is reversible. Therefore, oryzalin is a useful tool for studies relating to the cell cycle of this parasite. PMID- 10864262 TI - Evaluation of indirect fluorescent antibody test and enzyme-linked immunosorbent assay for diagnosis of hepatic amebiasis in Bangladesh. AB - Serum samples of 31 amebic liver abscess (ALA) patients, 8 amebic hepatitis (AH) patients, and 60 controls were tested for anti-amebic IgG by enzyme-linked immunosorbent assay (ELISA) and indirect fluorescent antibody tests (IFAT). Sera of 29 (93.6%) ALA and 6 (75%) AH patients and 2 (3.3%) control subjects were positive by IFAT. Anti-amebic antibody titer above the cutoff point (= 0.168; x + 2 SD of control sera) was observed in sera of 27 (87%) ALA, 4 (50%) AH, and 1 (1.7%) control by ELISA. All the 8 pus samples were positive for anti-amebic antibodies by IFAT and ELISA. Sensitivity of ELISA was 87% for ALA, with a positive predictive value of 0.96, and 50% for AH cases, with a positive predictive value of 0.80. The sensitivity of IFAT was 93.6% for ALA, with a positive predictive value of 0.94, and 75% for AH, with a positive predictive value of 0.75. When pus samples were tested, the sensitivity was 100% for both tests. The specificity was 98.3% for ELISA and 96.7% for IFAT. Although not significant, IFAT was found more sensitive than ELISA (P>0.05). PMID- 10864263 TI - The cercarial tail in Proterometra macrostoma (Digenea: Azygiidae): permeability and fine structure of the tegument. AB - Permeability of the cercarial tail in Proterometra macrostoma was examined in vitro with 1 mM 3H-glucose, which tails absorb by diffusion alone. Naturally emerged cercariae (bodies withdrawn into tails) were permeable, but they rapidly (3 min) equilibrated with glucose in the bathing medium and maintained steady state for 4 hr. Metabolism of absorbed glucose was not detectable until after 90 min, and radioactivity in bodies dissected from tails after 4 hr was negligible. On the basis of cercarial water content (90% of total weight) and absorbed isotope at steady state, the calculated volume of the equilibrating compartment was 4% of an intact cercaria. This value correlated well with that of the tegument (3-5%), which was 1-2 microm thick as seen by transmission electron microscopy. A continuous, electron-dense basal membrane/lamina separated the tegument from subtegument. We conclude that the glycocalyx and external plasma membrane are freely permeable, whereas the basal membrane is the barrier that effectively isolated the subtegument from exogenous glucose. The basal membrane also may be the primary structure that protects the subtegument and cercarial body from effects of osmotic stress. PMID- 10864264 TI - Skin lesions caused by Dermophthirius penneri (Monogenea: Microbothriidae) on wild-caught blacktip sharks (Carcharhinus limbatus). AB - Skin lesions caused by the ectoparasite Dermophthirius penneri Benz, 1987 (Monogenea: Microbothriidae) on 2 wild-caught blacktip sharks (Carcharhinus limbatus) from the northern Gulf of Mexico were studied using light and scanning electron microscopy. Grossly, lesions appeared as multifocal, well-demarcated, ovoid or irregularly shaped, light gray patches of skin. Scanning electron microscopy of lesions revealed gaps between placoid scales apparently created by detachment and loss of placoid scales, rotated and tilted placoid scales with blunt distal tips and shallow ridges, and a frayed epithelium that covered some placoid scales and filled some spaces between placoid scales. Light microscopy of lesions revealed epithelial hyperplasia accompanied by dermal infiltrates of moderate numbers of loosely arranged lymphocytes interposed between collagen bundles in the superficial layers of the stratum compactum. This report provides the first details of microbothriid skin lesions on wild sharks. Our results indicate that D. penneri caused chronic skin lesions not associated with bacterial infection or severe, debilitating, skin disease in the studied sharks. PMID- 10864266 TI - Prevalence of antibodies to Toxoplasma gondii in ostriches (Struthio camelus). AB - Serum samples from 973 ostriches (Struthio camelus) in Canada were examined for antibodies to Toxoplasma gondii by the modified agglutination test incorporating mercaptoethanol and formalin-fixed whole tachyzoites. Twenty-eight (2.9%) of the 973 birds were found to be seropositive for antibodies to T. gondii at titers of 1:25 in 15 birds, 1:50 in 12 birds, and 1:500 in 1 bird. This is the first record of T. gondii exposure in ostriches, and it supports the hypothesis that all avian species are susceptible to Toxoplasma infection. Nevertheless, the results of this study suggest that the risk of acquiring toxoplasmosis from ostriches as a food source is low. PMID- 10864265 TI - Seroprevalence of Toxoplasma gondii in Rocky Mountain bighorn sheep (Ovis canadensis). AB - Serum samples from 697 Rocky Mountain bighorn sheep (Ovis canadensis) from North America were examined for antibodies to Toxoplasma gondii by the modified agglutination test incorporating mercaptoethanol and formalin-fixed tachyzoites. Antibodies to T. gondii were found in 25 of 697 (3.6%) sheep in titers of 1:25 (8 sheep), 1:50 (4 sheep), 1:100 (7 sheep), 1:200 (1 sheep), 1:400 (1 sheep), 1:800 (1 sheep), and 1:1,600 (3 sheep). This is the first record of T. gondii exposure in bighorn sheep. PMID- 10864267 TI - Experimental transmission of Sarcocystis speeri Dubey and Lindsay, 1999 from the South American opossum (Didelphis albiventris) to the North American opossum (Didelphis virginiana). AB - Sarcocystis speeri Dubey and Lindsay, 1999 from the South American opossum Didelphis albiventris was successfully transmitted to the North American opossum Didelphis virginiana. Sporocysts from a naturally infected D. albiventris from Argentina were fed to 2 gamma-interferon knockout (KO) mice. The mice were killed 64 and 71 days after sporocyst feeding (DAF). Muscles containing sarcocysts from the KO mouse killed 71 DAF were fed to a captive D. virginiana; this opossum shed sporocysts 11 days after ingesting sarcocysts. Sporocysts from D. virginiana were fed to 9 KO mice and 4 budgerigars (Melopsittacus undulatus). Schizonts, sarcocysts, or both of S. speeri were found in tissues of all 7 KO mice killed 29 85 DAF; 2 mice died 39 and 48 DAF were not necropsied. Sarcocystis stages were not found in tissues of the 4 budgerigars fed S. speeri sporocysts and killed 35 DAE These results indicate that S. speeri is distinct from Sarcocystis falcatula and Sarcocystis neurona, and that S. speeri is present in both D. albiventris and D. virginiana. PMID- 10864268 TI - Prevalence of Toxoplasma gondii antibodies in sera of turkeys, chickens, and ducks from Egypt. AB - Sera from 173 turkeys, 108 chickens, and 48 ducks from Giza, Egypt, were tested for the presence of anti-Toxoplasma gondii antibodies by means of the modified agglutination test using mercaptoethanol and formalin-fixed tachyzoites. The prevalence of anti-T. gondii antibodies (>1:25) among turkeys, chickens, and ducks was 59.5%, 47.2%, and 50%, respectively. PMID- 10864269 TI - Haemonchus contortus (Nematoda: Trichostrongylidae) is much more sensitive than Caenorhabditis elegans (Nematoda: Rhabditidae) to the ovicidal action of thiabendazole. AB - When eggs of the trichostrongylid nematode Haemonchus contortus were exposed to thiabendazole, the concentration required to prevent hatching in 90% of the eggs (MIC90) was found to be 0.1 microg/ml (using 1% dimethylsulfoxide [DMSO] as solvent). In contrast, eggs of the free-living rhabditid nematode Caenorhabditis elegans hatched at normal rates at a concentration 200 times higher, i.e., 20 microg/ml, and showed only a partial inhibitory effect at a concentration 1,200 times higher, i.e., 120 microg/ml (in 3% DMSO). Because solubility limitations precluded the testing of higher concentrations of thiabendazole, a more soluble derivative, 5-([1-methylethoxy]carbonylamino)-2-(4-thiazloyl)1H-++ +benzimidazolyliminoacetic acid N,N-diethylethanamine salt, was tested against C. elegans eggs. The MIC90 was found to be 400 microg/ml, and although the derivative was not tested against H. contortus eggs, this finding further suggests that C. elegans eggs have an exceptionally low degree of benzimidazole sensitivity. PMID- 10864270 TI - Successful hyperimmune bovine colostrum treatment of Savanna monitors (Varanus exanthematicus) infected with Cryptosporidium sp. AB - Therapy based on the protective passive immunity of hyperimmune bovine colostrum (HBC) (raised against Cryptosporidium parvum in cows) was applied to 4 Savanna monitors (Varanus exanthematicus) with gastric Cryptosporidium sp. infections. All lizards were moderately emaciated, and their fecal and gastric lavage samples contained moderate numbers of Cryptosporidium sp. oocysts. The first 3 of 7 gastric HBC treatments at 1-wk interval each decreased the numbers of oocysts in the fecal and gastric samples to undetectable levels. Neither feces nor lavages of the HBC-treated lizards contained Cryptosporidium sp. oocysts after the HBC therapy, whereas such samples of a single control lizard remained positive for oocysts. Two of the HBC-treated lizards died spontaneously due to metastasized carcinoma and septicemia of unknown etiology, respectively, and 2 lizards treated and killed during the experiment were histologically negative for developmental stages of Cryptosporidium sp. The control lizard died spontaneously of septicemia of unknown etiology and contained developmental stages of Cryptosporidium sp. in the gastric region. The HBC therapy was efficacious in V. exanthematicus and is recommended for lizards with gastric cryptosporidiosis. PMID- 10864271 TI - Influence of temperature on the survival and infectivity of Trichinella spiralis larvae in Sarcophaga argyrostoma (Diptera, Sarcophagidae) maggots. AB - To investigate the role of fleshfly maggots as a paratenic host for Trichinella spiralis larvae, maggots of Sarcophaga argyrostoma (Muscidae, Sarcophagidae) kept at different temperatures (26, 22, 20, 16, 12, 8, and 4 C) were allowed to feed on T. spiralis-infected mouse meat. Trichinella larvae found in maggots kept at 8 26 C were able to cause infection when inoculated in mice. Infective larvae survived in maggots up to 5 days postinfection at 8 C and for shorter periods of time at higher temperatures. The survival time in maggots was negatively related to the temperature of maggot breeding. The results suggest that the role of S. argyrostoma in the dissemination of Trichinella larvae in nature is limited in comparison to the role played by mammals with scavenger and cannibalistic behavior. PMID- 10864272 TI - High-performance thin-layer chromatographic analysis of lutein and beta-carotene in Cerithidia californica (Gastropoda) infected with two species of larval trematodes. AB - High-performance thin-layer chromatography (HPTLC) analysis was done on lutein and beta-carotene in the digestive gland-gonad complex (DGG) and whole body of uninfected Cerithidia californica snails and those infected with the larval trematodes Mesostephanus appendiculatis or Euhaplorichis californiensis. HPTLC of the DGG extract on C-18 reversed-phase plates developed in petroleum ether acetonitrile-methanol (1:2:2) mobile phase showed 2 identifiable pigment zones; the least polar zone had a retention factor (Rf) of 0.07, identical to a beta carotene standard, and the more polar zone had an Rf of 0.41, identical to a lutein standard. Densitometric scanning of the pigment zones in sample versus standard chromatograms showed that the weight percent of lutein in the uninfected DGGs (3.4x10(-3)%) was significantly greater (P<0.05) than that of DGGs infected with either M. appendiculatis (0.35x10(-3)%) or E. californiensis (0.82x10(-3)%). Changes in beta-carotene in the infected DGGs were insignificant compared to the uninfected controls. However, the beta-carotene content of whole snails was significantly reduced (P<0.05) by infection with either trematode. PMID- 10864273 TI - Detection of heat shock protein-70 from Trichinella spiralis larvae using a modification of the routine western blotting procedure. AB - This is the first study that establishes a standardized western blotting method for the detection of heat shock protein (HSP) 70 from Trichinella spiralis using (selected) monoclonal antibodies (mAbs). Enhancement of HSP transfer onto the supportive membrane and increased retention of protein by the membrane are prominent features of the procedure. The reactivity of T. spiralis HSP70 on western blots was substantially increased by the use of a 10% acrylamide gel, the optimization of conditions during electrotransfer, and transfer onto Immobilon membranes. These data indicate that mAbs actually capable of detecting the agent of interest may be discarded because of nonoptimal testing conditions. We suggest that this method will aid in understanding the role and function of T. spiralis HSP70 in host-parasite (inter)relationship(s). PMID- 10864275 TI - Parasite body size and interspecific variation in levels of aggregation among nematodes. AB - The aggregation of parasites among individual hosts is one of the best documented features of parasite populations; we still do not know, however, why certain parasite species are more highly aggregated than other, related species. Here we search for a general explanation of interspecific variation in aggregation levels, based on the relationship between parasite body size and fecundity, transmission success, and intensity-dependent population regulation. We test the prediction that larger-bodied parasite species are more weakly aggregated than smaller-bodied related species, in a comparative analysis across parasitic nematode species. Across species, the variance-to-mean abundance ratio correlated negatively and significantly with nematode body sizes, as predicted. All other tests, however, including the more robust analyses controlling for phylogenetic influences, failed to support this result. This is mainly because the variance in infection levels is almost completely explained by mean parasite abundance. For this reason, it may prove difficult to identify a general biological explanation for interspecific variability in aggregation levels among parasites. PMID- 10864274 TI - Helminths in an intensively stocked population of lake trout, Salvelinus namaycush, from Lake Huron. AB - Eighty stocked lake trout Salvelinus namaycush (Salmonidae), collected from 2 locations in Lake Huron in May 1995, were examined for parasites. The parasite fauna of this top predator in Lake Huron was characterized by only 6 helminth species. Echinorhynchus salmonis infected all lake trout with a mean intensity of 163.9. The intensity of this acanthocephalan species significantly increased with host length and weight. Eubothrium salvelini infected 78 lake trout with a maximum number of 81 scoleces counted. Diplostomum sp., Cyathocephalus truncatus, Capillaria salvelini, and Neoechinorhynchus sp. infrequently infected lake trout. The low parasite species richness in these lake trout is believed to be due to their large size at stocking and to the loss of historical enzootic host-parasite relationships that followed the absence of this fish species in Lake Huron for 26 yr. PMID- 10864276 TI - Reassignment of Lamanema from Nematodirinae to Molineinae (Nematoda: Trichostronglyloidea). AB - The monospecific Lamanema historically has been assigned to the Nematodirinae within the Molineidae. Inconsistencies in morphological characters, within a phylogenetic context for Nematodirinae, led to a re-evaluation of the putative relationships and taxonomic placement of Lamanema. Among 7 putative synapomorphies for Nematodirinae, Lamanema possesses only 1, large eggs. Large eggs, sporadically present in phylogenetically disparate taxa of trichostrongyles, are equivocal with respect to placement of Lamanema; it is considered that possession of this single homoplasious character alone is insufficient justification to retain the genus in Nematodirinae. Affinities with the Trichostrongylidae (Cooperiinae or Haemonchinae) have also been proposed; however, Lamanema possess neither of 2 synapomorphies that diagnose monophyly of the family. Lamanema is retained in the Molineidae and transferred to the Molineinae as it possesses all characters of the family as currently defined. The origin of Lamanema represents a secondary colonization of ruminants by molineids and provides no context for elucidating the history of the Nematodirinae and Nematodirus. PMID- 10864277 TI - Differential parasite (Trematoda) encapsulation in Gammarus aequicauda (Amphipoda). AB - Because resistance to parasites usually has a cost for host species, it is theoretically expected that, in case of multi-infection, host immune responses should vary according to the levels of parasite pathogenicity. The crustacean gammarid Gammarus aequicauda is the second intermediate host of 4 trematode species. Three of these parasites always encyst in the abdomen of gammarids and have no particular effect on the host. However, 1 of these species is sometimes able to encyst in the cerebroid ganglia of the gammarid and strongly alter its behavior in a way that increases its predation risk by aquatic birds, the definitive hosts. In accordance with the hypothesis that the level of parasite pathogenicity influences the likelihood and the degree of host reaction, cases of melanization in our gammarid collection almost exclusively concern the cerebral metacercariae. Our results also indicate that this melanization is able to cancel the behavioral alterations induced by the parasite, suggesting that the cause of the manipulation is not the physical presence of metacercariae in the brain. PMID- 10864278 TI - Parasite community structure in Pimephales promelas (Pisces: Cyprinidae) from two converging streams. AB - Parasites of the fathead minnow, Pimephales promelas, were examined in fish collected from Elk Creek (40.88534 degrees N, 96.83366 degrees W) and West Oak Creek (40.90821 degrees N, 96.81432 degrees W), Lancaster County, Nebraska. These 2 streams are part of the Salt Valley watershed and flow together approximately 2 km downstream from the collection sites to form Oak Creek. This study examined the extent to which the 2 tributaries constitute a continuous habitat with respect to fish hosts. The parasite community included Trichodina sp., Myxobolus sp., Dactylogyrus simplex, D. bychowskyi, and D. pectenatus (all on gills); Gyrodactylus hoffmani (gill and body surface); Posthodiplostomum sp. (neascus, body cavity); and Uvulifer ambloplitis (encysted in skin). Among 46 fish from Elk Creek and 56 fish from West Oak Creek taken on 5 dates during April-July 1998, U. ambloplitis was found in Elk Creek fish at prevalences of 44-100% but in only 2 West Oak fish on 1 date. Prevalence and mean abundance of D. simplex also differed between the 2 sites. On the basis of these observations, fish populations in the 2 streams were considered to be distinct, with little or no fish movement between the tributaries. PMID- 10864279 TI - Brain electric source imaging: scalp Laplacian mapping and cortical imaging. AB - This article reviews recent progress in high-resolution EEG methodologies, in particular two widely studied approaches: scalp Laplacian mapping and cortical imaging. The common theoretical background behind these two high-resolution EEG approaches is discussed. The state of the art of the two methodologies are reviewed with examples illustrating their applications in imaging brain electrical activity. The emphasis is placed on the treatment of the mathematical and engineering methods of high-resolution EEG techniques, and reviews of our recent research in both scalp Laplacian mapping and cortical potential imaging. PMID- 10864280 TI - Forward and inverse problems of EEG dipole localization. AB - Mathematical procedures are discussed in detail of numerical solutions for obtaining scalp potentials from the electric sources. The finite-element method for an inhomogeneous volume conductor, the boundary-element method for a compartment model, and their hybrid for more general cases are discussed. Construction of the head model and typical estimation of electric conductivity of the compartment model is described, which can reduce errors in estimated dipole location caused by incorrect head geometry. The concept of reciprocity is explained, which is applied to understanding a relation between the electrode configuration and its sensitivity for various source conditions. Typical techniques for solving the inverse problem are reviewed for discrete source models. Methods of estimating accuracy of the dipole location in the presence of noise are discussed, together with some numerical examples. The dipolarity is a goodness-of-fit of the dipole approximation, and lowering of the dipolarity is related to inhomogeneous neuronal activity in the cortex. Finally, a criterion of determining the optimal number of model parameters is given in terms of AIC (Akaike Information Criterion), which is applied to decide the most probable number of equivalent dipoles. PMID- 10864281 TI - Transcranial magnetic stimulation--a new tool for functional imaging of the brain. AB - Recent progress in the theory and technology of transcranial magnetic stimulation (TMS) is leading to novel approaches in brain mapping. TMS becomes a powerful functional brain mapping tool when other imaging methods are used to record TMS evoked activity or when peripheral effects are observed as a function of stimulus location. TMS-evoked activity currently can be recorded by EEG, PET, and fMRI. In addition to providing indices of cortical excitability, these methods allow one to study brain connectivity directly, without the need for behavioral activations. When the coordinate systems in the different imaging modalities are combined, anatomical structures seen in MRI and activation sites determined by PET, fMRI, or MEG/EEG can be used for the selection of target areas in the brain. PET and fMRI can be used to map the spatial distribution of TMS-evoked activity. On the other hand, the combination of TMS and high-resolution EEG may often be the method of choice for basic neuroscience and for clinical diagnosis, for example, in the assessment of brain connectivity in patients suffering from neurodegenerative diseases or head injuries. PMID- 10864282 TI - Laplacian electrocardiography. AB - This article reviews the recent development in Laplacian electrocardiography. The Laplacian electrocardiogram was proposed in the early 1990s as an alternative for mapping regional cardiac electrical activity. Considerable progress has been made in the field during the last 5 years on this emerging technique. This article attempts to cover both the basic concepts and theory for general readers in biomedical engineering, and state-of-the-art developments in forward and inverse problems of Laplacian electrocardiography, as well as Laplacian ECG mapping in an experimental setting for researchers working in the field of cardiac mapping. The article also addresses controversies regarding the feasibility of experimentally obtaining the Laplacian electrocardiogram in human subjects. The work reviewed in this article suggests that Laplacian electrocardiography merits further investigation and promises to provide an important alternative means of assessing noninvasively cardiac electrical activity. PMID- 10864283 TI - Direct mapping of bioelectric activity. AB - Much can be learned about physiological function in heart, brain, and other tissues from measuring the electrical activity that mediates mechanical or neural phenomena. The study of electrophysiology has progressed from acquisition of a single or a few channels to hundreds of signals acquired simultaneously. Data acquisition, signal processing, and visualization in electrophysiology have benefitted from the parallel advances in electronics and computing hardware and software. The two primary technologies for mapping electrical activity at many sites are electrical, using electrodes directly placed on tissue, and optical, using voltage sensitive dyes to estimate membrane parameters. Each is valuable and provides information that is complementary to the other. This article is a description of the current state of electrical and optical mapping. PMID- 10864284 TI - Characteristic and critical excitation length scales in 1-D and 2-D simulations of reentrant cardiac arrhythmias using simple two-variable models. AB - We discuss major characteristic lengths as the lumped parameters characterizing kinematics and dynamics of excitation waves in idealized one- and two-dimensional myocardium. First we consider the directional dependence of the length scale in the anisotropic bi- and monodomain myocardium. Next, we show that there is a well defined smallness parameter that allows one to consider the monodomain equations as the first approximation to the bidomain theory. Then we use this approximation and turn our attention to the dynamics and to finding the major physiologic parameters governing such transient processes as the formation of a reentrant wave and its subsequent degradation into the malignant cardiac arrhythmia- ventricular fibrillation. It appears that in both the one- and two-dimensional cases the stability of a reentrant periodic process (representing a VT rhythm) is determined by the same two physiologic parameters: the wave width lambda, which is approximately the size of depolarized zone, and ratio of lambda to the critical length L(h)cr, which is defined as the wave width of an action potential propagating with the minimum possible speed. The parameter L(h)cr determines the length scale (size) of an ectopic region that may initiate a wave. It also determines the duration of the vulnerable window for initiating the unidirectional block as well as a minimum permissible length of the reentrant circuit. We also present some evidence that depending on the value of lambda the waveforms of the simulated ECGs for reentrant activity vary from monomorphic to polymorphic. PMID- 10864285 TI - The limits of sharing PMID- 10864286 TI - Gene therapy's trials. PMID- 10864287 TI - Biologists challenge sequencers on parasite genome publication. PMID- 10864288 TI - New Russian science head named PMID- 10864289 TI - Drive for more genomes threatens mouse sequence. PMID- 10864290 TI - Internet gateway planned for neuroinformatics data. PMID- 10864291 TI - Software spend boosts Israeli R&D PMID- 10864293 TI - Immigrants help offset Canada's brain-drain crisis PMID- 10864292 TI - Spain in quandary over French synchrotron PMID- 10864294 TI - A slot in the dock predicted for the chemical industry. PMID- 10864295 TI - Roche brings down curtain on Swiss immunology lab. PMID- 10864296 TI - US decides close tabs must be kept on xenotransplants... PMID- 10864298 TI - Cloning's owners go to war. PMID- 10864297 TI - And sets up a body to oversee trials PMID- 10864299 TI - Regulation, not private enterprise, is the key to a healthy environment. PMID- 10864300 TI - England and US corner the journal market. PMID- 10864301 TI - Celera's role in opening up new frontiers. PMID- 10864302 TI - Climate change in perspective. PMID- 10864303 TI - Show them how it's really done. PMID- 10864304 TI - New ice age, or just cold feet? PMID- 10864305 TI - The realms of Archaean life. PMID- 10864306 TI - Semiconductors meet biology. PMID- 10864308 TI - Semiconductor physics. Half-matter, half-light amplifier PMID- 10864307 TI - Alzheimer's disease. Closing in on gamma-secretase. PMID- 10864309 TI - Laser-matter interactions. It takes two electrons to tango PMID- 10864310 TI - Structural biology. Pumping ions. PMID- 10864311 TI - Recombinant erythropoietin in urine. PMID- 10864312 TI - Parabasalian flagellates are ancient eukaryotes. PMID- 10864313 TI - Tectonics and water on Europa. PMID- 10864314 TI - Scalar turbulence AB - The advection of a passive substance by a turbulent flow is important in many natural and engineering settings. The concentration of such a substance can exhibit complex dynamic behaviour that shows many phenomenological parallels with the behaviour of the turbulent velocity field. Yet the statistical properties of this so-called 'passive scalar' turbulence are decoupled from those of the underlying velocity field. Passive scalar turbulence has recently yielded to mathematical analysis, and such progress may ultimately lead to a better understanding of the still intractable problem of fluid turbulence itself. PMID- 10864315 TI - Crystal structure of the calcium pump of sarcoplasmic reticulum at 2.6 A resolution. AB - Calcium ATPase is a member of the P-type ATPases that transport ions across the membrane against a concentration gradient. Here we have solved the crystal structure of the calcium ATPase of skeletal muscle sarcoplasmic reticulum (SERCA1a) at 2.6 A resolution with two calcium ions bound in the transmembrane domain, which comprises ten alpha-helices. The two calcium ions are located side by side and are surrounded by four transmembrane helices, two of which are unwound for efficient coordination geometry. The cytoplasmic region consists of three well separated domains, with the phosphorylation site in the central catalytic domain and the adenosine-binding site on another domain. The phosphorylation domain has the same fold as haloacid dehalogenase. Comparison with a low-resolution electron density map of the enzyme in the absence of calcium and with biochemical data suggests that large domain movements take place during active transport. PMID- 10864316 TI - Newly synthesized lithium in the interstellar medium AB - Astronomical observations of elemental and isotopic abundances provide the means to determine the source of elements and to reveal their evolutionary pathways since the formation of the Galaxy some 15 billion years ago. The abundance of lithium is particularly interesting because, although some of it is thought to be primordial, most results from spallation reactions (in which Galactic cosmic rays break apart larger nuclei in the interstellar medium). Spallation reactions are crucial for the production of other light elements, such as beryllium and boron, so observations of lithium isotopic abundances can be used to test model predictions for light-element synthesis in general. Here we report observations of 7Li and 6Li abundances in several interstellar clouds lying in the direction of the star o Persei. We find the abundance ratio 7Li/6Li to be about 2, which is significantly lower than the average Solar System value of 12.3 (refs 6, 7). An abundance ratio of 2 is clear evidence that the observed lithium must have resulted entirely from spallation, confirming a basic tenet of light-element synthesis. The total lithium abundance, however, is not enhanced as expected. PMID- 10864317 TI - Correlated electron emission in multiphoton double ionization AB - Electronic correlations govern the dynamics of many phenomena in nature, such as chemical reactions and solid state effects, including superconductivity. Such correlation effects can be most clearly investigated in processes involving single atoms. In particular, the emission of two electrons from an atom--induced by the impact of a single photon, a charged particle or by a short laser pulse- has become the standard process for studies of dynamical electron correlations. Atoms and molecules exposed to laser fields that are comparable in intensity to the nuclear fields have extremely high probabilities for double ionization; this has been attributed to electron-electron interaction. Here we report a strong correlation between the magnitude and the direction of the momentum of two electrons that are emitted from an argon atom, driven by a femtosecond laser pulse (at 38 TW cm(-2)). Increasing the laser intensity causes the momentum correlation between the electrons to be lost, implying that a transition in the laser-atom coupling mechanism takes place. PMID- 10864318 TI - Improving the performance of doped pi-conjugated polymers for use in organic light-emitting diodes AB - Organic light-emitting diodes (OLEDs) represent a promising technology for large, flexible, lightweight, flat-panel displays. Such devices consist of one or several semiconducting organic layer(s) sandwiched between two electrodes. When an electric field is applied, electrons are injected by the cathode into the lowest unoccupied molecular orbital of the adjacent molecules (simultaneously, holes are injected by the anode into the highest occupied molecular orbital). The two types of carriers migrate towards each other and a fraction of them recombine to form excitons, some of which decay radiatively to the ground state by spontaneous emission. Doped pi-conjugated polymer layers improve the injection of holes in OLED devices; this is thought to result from the more favourable work function of these injection layers compared with the more commonly used layer material (indium tin oxide). Here we demonstrate that by increasing the doping level of such polymers, the barrier to hole injection can be continuously reduced. The use of combinatorial devices allows us to quickly screen for the optimum doping level. We apply this concept in OLED devices with hole-limited electroluminescence (such as polyfluorene-based systems), finding that it is possible to significantly reduce the operating voltage while improving the light output and efficiency. PMID- 10864320 TI - Reduced growth of Alaskan white spruce in the twentieth century from temperature induced drought stress. AB - The extension of growing season at high northern latitudes seems increasingly clear from satellite observations of vegetation extent and duration. This extension is also thought to explain the observed increase in amplitude of seasonal variations in atmospheric CO2 concentration. Increased plant respiration and photosynthesis both correlate well with increases in temperature this century and are therefore the most probable link between the vegetation and CO2 observations. From these observations, it has been suggested that increases in temperature have stimulated carbon uptake in high latitudes and for the boreal forest system as a whole. Here we present multi-proxy tree-ring data (ring width, maximum late-wood density and carbon-isotope composition) from 20 productive stands of white spruce in the interior of Alaska. The tree-ring records show a strong and consistent relationship over the past 90 years and indicate that, in contrast with earlier predictions, radial growth has decreased with increasing temperature. Our data show that temperature-induced drought stress has disproportionately affected the most rapidly growing white spruce, suggesting that, under recent climate warming, drought may have been an important factor limiting carbon uptake in a large portion of the North American boreal forest. If this limitation in growth due to drought stress is sustained, the future capacity of northern latitudes to sequester carbon may be less than currently expected. PMID- 10864319 TI - Selection of peptides with semiconductor binding specificity for directed nanocrystal assembly. AB - In biological systems, organic molecules exert a remarkable level of control over the nucleation and mineral phase of inorganic materials such as calcium carbonate and silica, and over the assembly of crystallites and other nanoscale building blocks into complex structures required for biological function. This ability to direct the assembly of nanoscale components into controlled and sophisticated structures has motivated intense efforts to develop assembly methods that mimic or exploit the recognition capabilities and interactions found in biological systems. Of particular value would be methods that could be applied to materials with interesting electronic or optical properties, but natural evolution has not selected for interactions between biomolecules and such materials. However, peptides with limited selectivity for binding to metal surfaces and metal oxide surfaces have been successfully selected. Here we extend this approach and show that combinatorial phage-display libraries can be used to evolve peptides that bind to a range of semiconductor surfaces with high specificity, depending on the crystallographic orientation and composition of the structurally similar materials we have used. As electronic devices contain structurally related materials in close proximity, such peptides may find use for the controlled placement and assembly of a variety of practically important materials, thus broadening the scope for 'bottom-up' fabrication approaches. PMID- 10864321 TI - Isotopic evidence for Late Cretaceous plume-ridge interaction at the Hawaiian hotspot AB - When a mantle plume interacts with a mid-ocean ridge, both are noticeably affected. The mid-ocean ridge can display anomalously shallow bathymetry, excess volcanism, thickened crust, asymmetric sea-floor spreading and a plume component in the composition of the ridge basalts. The hotspot-related volcanism can be drawn closer to the ridge, and its geochemical composition can also be affected. Here we present Sr-Nd-Pb isotopic analyses of samples from the next-to-oldest seamount in the Hawaiian hotspot track, the Detroit seamount at 51 degrees N, which show that, 81 Myr ago, the Hawaiian hotspot produced volcanism with an isotopic signature indistinguishable from mid-ocean ridge basalt. This composition is unprecedented in the known volcanism from the Hawaiian hotspot, but is consistent with the interpretation from plate reconstructions that the hotspot was located close to a mid-ocean ridge about 80 Myr ago. As the rising mantle plume encountered the hot, low-viscosity asthenosphere and hot, thin lithosphere near the spreading centre, it appears to have entrained enough of the isotopically depleted upper mantle to overwhelm the chemical characteristics of the plume itself. The Hawaiian hotspot thus joins the growing list of hotspots that have interacted with a rift early in their history. PMID- 10864322 TI - Filamentous microfossils in a 3,235-million-year-old volcanogenic massive sulphide deposit. AB - The record of Archaean microfossils is sparse. Of the few bona fide fossil assemblages, most are from shallow-water settings, and they are typically associated with laminated, stromatolitic sedimentary rocks. Microfossils from deep-sea hydrothermal systems have not been reported in Precambrian rocks (> 544 million years old), although thermophilic microbes are ubiquitous in modern sea floor hydrothermal settings, and apparently have the most ancient lineages. Here, I report the discovery of pyritic filaments, the probable fossil remains of thread-like microorganisms, in a 3,235-million-year-old deep-sea volcanogenic massive sulphide deposit from the Pilbara Craton of Australia. From their mode of occurrence, the micro-organisms were probably thermophilic chemotropic prokaryotes, which inhabited sub-sea-floor hydrothermal environments. They represent the first fossil evidence for microbial life in a Precambrian submarine thermal spring system, and extend the known range of submarine hydrothermal biota by more than 2,700 million years. Such environments may have hosted the first living systems on Earth, consistent with proposals for a thermophilic origin of life. PMID- 10864323 TI - Parasite adaptation to locally common host genotypes. AB - According to the Red Queen hypothesis--which states that interactions among species (such as hosts and parasites) lead to constant natural selection for adaptation and counter-adaptation--the disproportionate evolutionary success of parasites on common host genotypes leads to correlated selection for sexual reproduction and local adaptation by the parasite population. Here we determined whether local adaptation is due to disproportionate infection of common host genotypes, and, if so, whether infection of common host genotypes is due to commonness per se, or some other aspect of these genotypes. In a reciprocal cross inoculation experiment parasites occupying the same geographical area (sympatric) infected locally common host genotypes significantly more often than rare host genotypes, whereas parasites occupying separate geographical areas (allopatric) showed no such significant difference. A mixed source of parasites (containing F1 hybrids) also showed no difference in infection between rare and common host genotypes. These results show that local adaptation results from parasite tracking of locally common host genotypes, and, as such, a necessary condition of the Red Queen hypothesis is met. PMID- 10864324 TI - Adhesive force of a single gecko foot-hair. AB - Geckos are exceptional in their ability to climb rapidly up smooth vertical surfaces. Microscopy has shown that a gecko's foot has nearly five hundred thousand keratinous hairs or setae. Each 30-130 microm long seta is only one tenth the diameter of a human hair and contains hundreds of projections terminating in 0.2-0.5 microm spatula-shaped structures. After nearly a century of anatomical description, here we report the first direct measurements of single setal force by using a two-dimensional micro-electromechanical systems force sensor and a wire as a force gauge. Measurements revealed that a seta is ten times more effective at adhesion than predicted from maximal estimates on whole animals. Adhesive force values support the hypothesis that individual seta operate by van der Waals forces. The gecko's peculiar behaviour of toe uncurling and peeling led us to discover two aspects of setal function which increase their effectiveness. A unique macroscopic orientation and preloading of the seta increased attachment force 600-fold above that of frictional measurements of the material. Suitably orientated setae reduced the forces necessary to peel the toe by simply detaching above a critical angle with the substratum. PMID- 10864325 TI - Neural synchrony correlates with surface segregation rules. AB - To analyse an image, the visual system must decompose the scene into its relevant parts. Identifying distinct surfaces is a basic operation in such analysis, and is believed to precede object recognition. Two superimposed gratings moving in different directions (plaid stimuli) may be perceived either as two surfaces, one being transparent and sliding on top of the other (component motion) or as a single pattern whose direction of motion is intermediate to the component vectors (pattern motion). The degree of transparency, and hence the perception, can be manipulated by changing only the luminance of the grating intersections. Here we show that neurons in two visual cortical areas--A18 and PMLS--synchronize their discharges when responding to contours of the same surface but not when responding to contours belonging to different surfaces. The amplitudes of responses correspond to previously described rate predictions for component and pattern motion, but, in contrast to synchrony, failed to reflect the transition from component to pattern motion induced by manipulating the degree of transparency. Thus, dynamic changes in synchronization could encode, in a context dependent way, relations among simultaneous responses to spatially superimposed contours and thereby bias their association with distinct surfaces. PMID- 10864326 TI - Photoactivated gamma-secretase inhibitors directed to the active site covalently label presenilin 1. AB - Cleavage of amyloid precursor protein (APP) by the beta- and gamma-secretases generates the amino and carboxy termini, respectively, of the A beta amyloidogenic peptides A beta40 and A beta42--the major constituents of the amyloid plaques in the brain parenchyma of Alzheimer's disease patients. There is evidence that the polytopic membrane-spanning proteins, presenilin 1 and 2 (PS1 and PS2), are important determinants of gamma-secretase activity: mutations in PS1 and PS2 that are associated with early-onset familial Alzheimer's disease increase the production of A beta42 (refs 4-6), the more amyloidogenic peptide; gamma-secretase activity is reduced in neuronal cultures derived from PS1 deficient mouse embryos; and directed mutagenesis of two conserved aspartates in transmembrane segments of PS1 inactivates the ability of gamma-secretase to catalyse processing of APP within its transmembrane domain. It is unknown, however, whether PS1 (which has little or no homology to any known aspartyl protease) is itself a transmembrane aspartyl protease or a gamma-secretase cofactor, or helps to colocalize gamma-secretase and APP. Here we report photoaffinity labelling of PS1 (and PS2) by potent gamma-secretase inhibitors that were designed to function as transition state analogue inhibitors directed to the active site of an aspartyl protease. This observation indicates that PS1 (and PS2) may contain the active site of gamma-secretase. Interestingly, the intact, single-chain form of wild-type PS1 is not labelled by an active-site directed photoaffinity probe, suggesting that intact wild-type PS1 may be an aspartyl protease zymogen. PMID- 10864327 TI - Alpha-haemolysin of uropathogenic E. coli induces Ca2+ oscillations in renal epithelial cells. AB - Pyelonephritis is one of the most common febrile diseases in children. If not treated appropriately, it causes irreversible renal damage and accounts for a large proportion of end stage renal failures. Renal scarring can occur in the absence of inflammatory cells, indicating that bacteria may have a direct signalling effect on renal cells. Intracellular calcium ([Ca2+]i) oscillations can protect cells from the cytotoxic effects of prolonged increases in intracellular calcium. However, no pathophysiologically relevant protein that induces such oscillations has been identified. Here we show that infection by uropathogenic Escherichia coli induces a constant, low-frequency oscillatory [Ca2+]i response in target primary rat renal epithelial cells induced by the secreted RTX (repeats-in-toxin) toxin alpha-haemolysin. The response depends on calcium influx through L-type calcium channels as well as from internal stores gated by inositol triphosphate. Internal calcium oscillations induced by alpha haemolysin in a renal epithelial cell line stimulated production of cytokines interleukin (IL)-6 and IL-8. Our findings indicate a novel role for alpha haemolysin in pyelonephritis: as an inducer of an oscillating second messenger response in target cells, which fine-tunes gene expression during the inflammatory response. PMID- 10864328 TI - Frequent chromosomal translocations induced by DNA double-strand breaks. AB - The faithful repair of DNA damage such as chromosomal double-strand breaks (DSBs) is crucial for genomic integrity. Aberrant repair of these lesions can result in chromosomal rearrangements, including translocations, which are associated with numerous tumours. Models predict that some translocations arise from DSB-induced recombination in differentiating lymphoid cell types or from aberrant repair of DNA damage induced by irradiation or other agents; however, a genetic system to study the aetiology of these events has been lacking. Here we use a mouse embryonic stem cell system to examine the role of DNA damage on the formation of translocations. We find that two DSBs, each on different chromosomes, are sufficient to promote frequent reciprocal translocations. The results are in striking contrast with interchromosomal repair of a single DSB in an analogous system in which translocations are not recovered. Thus, while interchromosomal DNA repair does not result in genome instability per se, the presence of two DSBs in a single cell can alter the spectrum of repair products that are recovered. PMID- 10864329 TI - Redundant roles for the TFIID and SAGA complexes in global transcription. AB - The transcription factors TFIID and SAGA are multi-subunit complexes involved in transcription by RNA polymerase II. TFIID and SAGA contain common TATA-binding protein (TBP)-associated factor (TAF(II)) subunits and each complex contains a subunit with histone acetyltransferase activity. These observations have raised questions about whether the functions of the two complexes in vivo are unique or overlapping. Here we use genome-wide expression analysis to investigate how expression of the yeast genome depends on both shared and unique subunits of these two complexes. We find that expression of most genes requires one or more of the common TAF(II) subunits, indicating that the functions of TFIID and SAGA are widely required for gene expression. Among the subunits shared by TFIID and SAGA are three histone-like TAF(II)s, which have been proposed to form a sub complex and mediate a common function in global transcription. Unexpectedly, we find that the histone-like TAF(II)s have distinct roles in expression of the yeast genome. Most importantly, we show that the histone acetylase components of TFIID and SAGA (TAF(II)145 and Gcn5) are functionally redundant, indicating that expression of a large fraction of yeast genes can be regulated through the action of either complex. PMID- 10864331 TI - Biological sensing of small field differences by magnetically sensitive chemical reactions. AB - There is evidence that animals can detect small changes in the Earth's magnetic field by two distinct mechanisms, one using the mineral magnetite as the primary sensor and one using magnetically sensitive chemical reactions. Magnetite responds by physically twisting, or even reorienting the whole organism in the case of some bacteria, but the magnetic dipoles of individual molecules are too small to respond in the same way. Here we assess whether reactions whose rates are affected by the orientation of reactants in magnetic fields could form the basis of a biological compass. We use a general model, incorporating biological components and design criteria, to calculate realistic constraints for such a compass. This model compares a chemical signal produced owing to magnetic field effects with stochastic noise and with changes due to physiological temperature variation. Our analysis shows that a chemically based biological compass is feasible with its size, for any given detection limit, being dependent on the magnetic sensitivity of the rate constant of the chemical reaction. PMID- 10864330 TI - Effects of mechanical forces on maintenance and adaptation of form in trabecular bone. AB - The architecture of trabecular bone, the porous bone found in the spine and at articulating joints, provides the requirements for optimal load transfer, by pairing suitable strength and stiffness to minimal weight according to rules of mathematical design. But, as it is unlikely that the architecture is fully pre programmed in the genes, how are the bone cells informed about these rules, which so obviously dictate architecture? A relationship exists between bone architecture and mechanical usage--while strenuous exercise increases bone mass, disuse, as in microgravity and inactivity, reduces it. Bone resorption cells (osteoclasts) and bone formation cells (osteoblasts) normally balance bone mass in a coupled homeostatic process of remodelling, which renews some 25% of trabecular bone volume per year. Here we present a computational model of the metabolic process in bone that confirms that cell coupling is governed by feedback from mechanical load transfer. This model can explain the emergence and maintenance of trabecular architecture as an optimal mechanical structure, as well as its adaptation to alternative external loads. PMID- 10864332 TI - US science shocked by revelations of sexual discrimination. PMID- 10864333 TI - Making moves to redress the gender imbalance. PMID- 10864334 TI - New UK legislation aids fight against discrimination. PMID- 10864335 TI - US minorities stake their claim in science and engineering. PMID- 10864337 TI - Brain metastasis: steel knife or gamma knife? PMID- 10864336 TI - Insurance policies for prophylactic surgery: to cover or not to cover? PMID- 10864338 TI - Current national health insurance coverage policies for breast and ovarian cancer prophylactic surgery. AB - BACKGROUND: The efficacy of prophylactic mastectomy and oophorectomy in reducing breast and ovarian carcinoma has recently been reported in high-risk women. Because cost has become central to medical decision-making, this study was designed to evaluate currently existing coverage policies for these procedures. METHODS: A confidential detailed cross-sectional nationwide survey of 481 medical directors from the American Association of Health Plans, Medicare, and Medicaid was conducted. RESULTS: Of the 150 respondents, 65% (n = 97) had 100,000 or more enrolled members and 35% (n = 53) had fewer than 100,000 enrolled members. Only 44% of private plans have specific policies for coverage of prophylactic mastectomy for a strong family history of breast cancer and 38% of plans for a BRCA mutation. Only 20% of total responding plans had a policy for coverage of prophylactic oophorectomy under any clinical circumstance. Governmental carriers were significantly less likely to have any policy for prophylactic surgery (range, 2%-12%) compared with nongovernmental plans (range, 24%-44%; P < .001). No significant regional differences for coverage policies were identified (P > .05). CONCLUSIONS: Significant variations currently exist for health insurance coverage of prophylactic mastectomy and oophorectomy. As genetic testing becomes widespread, more uniform policies should be established to enable appropriate high-risk candidates equal access and coverage for these procedures. PMID- 10864339 TI - Stereotactic radiosurgery for brain metastases from breast cancer. AB - BACKGROUND: Stereotactic radiosurgery is an alternative to resection or to radiotherapy alone for patients with brain metastases. Outcomes after radiosurgery for patients with brain metastases specifically from breast cancer have not been defined. METHODS: We retrospectively studied survival and tumor control for all patients with brain metastases from breast cancer who underwent gamma knife stereotactic radiosurgery at the University of Pittsburgh. Univariate and multivariate analyses were used to determine which prognostic factors significantly affected survival. RESULTS: Thirty patients underwent radiosurgery between 1990 and 1997. A total of 58 metastases were treated. The median length of survival for all patients was 13 months from radiosurgery and 18 months from diagnosis of brain metastases. The tumor control rate on follow-up imaging was 93%. On multivariate analysis, the only factor that correlated with longer survival was the absence of multiple brain metastases. Age, presence of systemic disease, previous whole brain radiation, location, and total tumor volume did not significantly affect survival. Four patients had tumors with evidence of radiation-induced edema after radiosurgery but did not require resection. Two patients underwent delayed resection for tumor growth after radiosurgery. CONCLUSIONS: Stereotactic radiosurgery is an effective treatment for brain metastases from breast cancer and is associated with a low complication rate. PMID- 10864340 TI - Geographic variation in patient surveillance after radical prostatectomy. AB - BACKGROUND: Prostate cancer is often diagnosed early enough in its clinical course to permit radical prostatectomy to be done with curative intent, yet many patients experience tumor recurrence. Most patients receive postoperative surveillance, but the intensity of testing varies appreciably. We sought to evaluate the influence of geographic location on the variability of surveillance intensity. METHODS: Questionnaires pertaining to postoperative surveillance were mailed to 4467 members of the American Urological Association (AUA). Practice pattern variation was assessed among 24 large metropolitan statistical areas, among nine United States census regions, and by health maintenance organization penetration rate. RESULTS: Of 4467 urologists surveyed, 1416 (32%) responded and 1050 (24%) responses were evaluable. Correlation analysis showed that mean follow up intensity across modalities surveyed was highly correlated across tumor, node, metastasis (TNM) stages and years postsurgery. We found no significant main effects attributable to metropolitan statistical area, United States (US) census region, or health maintenance organization (HMO) penetration rate for commonly used surveillance modalities: serum prostate-specific antigen (PSA), office visit, and urinalysis. For infrequently used modalities, there were minimal effects on testing intensity of US census region, metropolitan statistical area, and HMO penetration rate. Few two-way and three-way interactions were significant. CONCLUSIONS: The utilization of commonly used surveillance modalities by urologists caring for patients after radical prostatectomy is not affected by metropolitan statistical area, US census region, or HMO penetration rate. PMID- 10864341 TI - Gastric cancer in young patients: demographic, clinicopathological, and prognostic factors in 92 patients. AB - BACKGROUND: This investigation was undertaken to define the demographic, clinicopathological, and prognostic factors relevant to young patients with gastric adenocarcinoma. METHODS: A prospective database of all patients with gastric cancer who presented to Memorial Sloan-Kettering Cancer Center was started in 1985. Clinical, pathological, and operative records and follow-up data on 92 patients, 40 years of age or younger, with a primary diagnosis of gastric cancer were reviewed. RESULTS: The mean patient age was 35 +/- 4.9 years (range, 17-40 years), and 52 were male. The male-to-female ratio of patients younger than 30 was 0.85/1; whereas in those older than 30, the ratio was 1.45/1. Sixty-six percent of the patients were white, 15% Asian, 11% Hispanic, and 8% were black American. Nineteen percent of patients reported a family history of gastric cancer. Sixty-six patients (71%) presented with stage III or IV disease, whereas 13 patients, each, presented with stage I or II disease. Poorly differentiated lesions were present in 71%. Resection with curative intent was undertaken in 47 patients, and resection with palliative intent was performed in 24 patients. Tumor site (proximal vs. distal vs. linitus plastica), advanced T stage, and the presence of nodal disease were significant predictors of disease-free survival on both univariate and multivariate analyses. The mean survival time and disease specific 5-year survival rates for individual Union International Contre le Cancer tumor stages were similar to those observed in older populations of patients with gastric cancer; and eight patients, who presented with early (T1/T2) node-negative tumors, are alive and well a minimum of 60 months after resection. CONCLUSIONS: The high frequency of a positive family history in young patients suggests an opportunity to identify a high-risk population for screening. PMID- 10864342 TI - TA90-IC, a new marker for advanced colon cancer. AB - BACKGROUND: Although carcinoembryonic antigen (CEA) is the most frequently used marker for colon cancer, it is elevated in only 70% of patients with advanced disease and in even fewer patients with earlier stages of disease. We previously identified a 90-kDa glycoprotein, TA90, which is present in serum in the form of circulating immune complexes. TA90 is found in a variety of solid neoplasms but rarely in healthy controls (3.2%). We hypothesized that this new tumor-associated antigen may be a useful marker for colon cancer. METHODS: Serum samples from 59 patients with known colon adenocarcinoma were analyzed for the presence of CEA and TA90. Fifty-one (86%) patients had distant metastases; the remaining patients had clinically localized primary colon cancer. A murine monoclonal antibody-based enzyme-linked immunosorbent assay was used to measure concentrations of TA90 specific circulating immune complexes (TA90-IC). A positive value was defined as an optical density of more than 0.410 at 405 nm. Forty-seven (80%) of the 59 patients had serum samples for TA90 and CEA drawn at the same time. RESULTS: TA90 IC concentrations were elevated more frequently than CEA concentrations (82.9% vs. 70.2%; P = .134). The combination of both markers identified more patients with colon carcinoma than did either marker alone (93.6%; P < .001). CONCLUSIONS: Concomitant use of TA90-IC and CEA identified 93.6% of patients with advanced colon cancer. The role of TA90-IC in screening and monitoring progression of earlier disease deserves further investigation. PMID- 10864343 TI - Adjuvant radiation trials for high-risk breast cancer patients: adequacy of lymphadenectomy. AB - BACKGROUND: The recently published, widely publicized adjuvant radiation trials from Denmark and Canada concluded that the addition of postoperative radiotherapy (XRT) to modified radical mastectomy (MRM) and adjuvant chemotherapy reduces locoregional recurrences and prolongs survival in high-risk premenopausal patients with breast cancer. Our thesis is that adequate lymphadenectomies were not performed in either study. Consequently, the conclusion to these studies is not applicable to those patients who have undergone adequate surgery. METHODS: To better assess adequate lymph node yield from an MRM, a retrospective review was performed on 215 consecutive patients treated surgically for invasive breast cancer. Data from this review were compared with the surgical data from the above mentioned radiotherapy trials. RESULTS: In a group of 131 patients who had MRM, the average number of nodes removed was 26 (median, 25), and 75.5% of the specimens had 20 or more lymph nodes. In 73 patients who underwent segmental mastectomy with axillary lymph node dissection, both the average and the median number of lymph nodes removed were 24, and 68.9% had 20 or more nodes. These data compare to the Danish radiation trial in which a median of 7 lymph nodes were removed (with 76% of the patients having 9 or fewer lymph nodes in the specimen) and to the Canadian radiation trial in which a median of 11 lymph nodes were removed. In addition, in our breast cancer patients with positive nodes (84 of 204; 41.2%), 45.2.% (38 of 84) had more than three positive nodes compared with 29.8% in the Danish study and 35% in the Canadian study. CONCLUSIONS: Our surgical data are sufficiently different from those of the Danish and Canadian studies to indicate that, in those studies, incomplete lymph node dissections were performed and that residual disease was left behind in the axilla in some or all of the patients. The addition of XRT in the setting of residual axillary disease may compensate for an inadequate operation and yield an acceptable oncological result; however, these studies did not provide an adequate comparison with a well-performed MRM without XRT. In the absence of documented benefit, XRT should not be routinely added if a complete lymph node dissection has been performed. PMID- 10864345 TI - Outcomes research in surgical oncology. AB - BACKGROUND: There have been significant developments and advances in the area of outcomes research in the past 25 years. Unfortunately, many surgical oncologists may not have a clear concept of outcomes research and the methodology involved. METHODS: A literature-based review article was done that included an overview of outcomes research, and study design and types, outcome measures, outcome instruments, and sources of outcome data were examined. In addition, we reviewed small area variation/volume outcome analysis as well as quality-of-life studies and their applications in surgical oncology clinical investigation. Specific examples from surgical oncology were identified. RESULTS: As the costs of health care have increased, so has the emphasis on measuring outcomes of medical and surgical care to determine the quality and appropriateness of care. Marked variations in a variety of outcomes after oncological procedures have been attributed to individual surgeon and institution characteristics. Because much of the clinical surgical oncology literature deals only with the traditional mortality and morbidity outcomes, a more comprehensive examination of patient outcomes is required to fully evaluate the impact of patient management decisions. Health-related quality of life can be measured and analyzed in several ways and decisions regarding the use of such methodology are dependent on multiple factors. CONCLUSIONS: Surgical oncologists should recognize that the true value of their interventions requires systematic and comprehensive examination of patient outcomes. PMID- 10864344 TI - Adverse reactions to isosulfan blue during selective sentinel lymph node dissection in melanoma. AB - BACKGROUND: Selective sentinel lymph node (SLN) dissection can spare about 80% of patients with primary melanoma from radical lymph node dissection. This procedure identifies the SLN either visually by injecting isosulfan blue dye around the primary melanoma site or by handheld gamma probe after radiocolloid injection. METHODS: During selective SLN mapping, 1 to 5 ml of isosulfan blue was injected intradermally around the primary melanoma. From November 1993, to August 1998, 406 patients underwent intraoperative lymphatic mapping with the use of both isosulfan blue and radiocolloid injection. Three cases of selective SLN dissection, in which adverse reactions to isosulfan blue occurred, were reviewed. RESULTS: We report three cases of anaphylaxis after intradermal injection with isosulfan blue of 406 patients who underwent intraoperative lymphatic mapping by using the procedure as described above. The three cases we report vary in severity from treatable hypotension with urticaria and erythema to severe cardiovascular collapse with or without bronchospasm or urticaria. CONCLUSIONS: In our series, the incidence of anaphylaxis to isosulfan blue was approximately 1%. Anaphylaxis can be fatal if not recognized and treated rapidly. Operating room personnel who participate in intraoperative lymphatic mapping where isosulfan blue is used must be aware of the potential consequences and be prepared to treat anaphylaxis. PMID- 10864346 TI - Thyroid cancer: 1999 update. PMID- 10864347 TI - Treatment strategies for chronic hepatitis C virus infection. PMID- 10864348 TI - Distal stomach appears essential in the regulation of gastrointestinal transit. AB - Stomach and small bowel both influence gastrointestinal motility. We studied which portion of the stomach was essential for the regulation of gastrointestinal movement and determined the role of vasoactive intestinal polypeptide in this regulation. The study subjects consisted of 45 controls, 46 patients after subtotal gastrectomy, and 13 patients after total gastrectomy for stomach cancer. Orocecal transit time was measured, using the hydrogen breath test, to represent gastrointestinal movement, while plasma vasoactive intestinal polypeptide level was simultaneously assessed. The orocecal transit times in the study groups were (means +/- SD) 91.1 +/- 45.0, 57.1 +/- 34.3, and 60.8 +/- 34.8 min, respectively (P < 0.01). In the subtotal gastrectomy patients, age showed a negative correlation with orocecal transit time (r = -0.388; P < 0.01). In the total gastrectomy patients, no particular demographic factor influenced orocecal transit. Plasma vasoactive intestinal polypeptide levels in the three groups were 20.7 +/- 10.8, 22.7 +/- 10.9, and 20.6 +/- 9.1 pg/ml, respectively (NS). We conclude that both types of gastrectomies enhanced gastrointestinal movement, showing a similar effect, and that the distal stomach plus pylorus are most likely to exert an important inhibitory mechanism in the regulation of this movement. Vasoactive intestinal polypeptide is not a major peptide mediating this regulation. PMID- 10864349 TI - Use of salivary acetaminophen concentration to assess gastric emptying rate of liquids. AB - The gastric emptying rate (GER) of liquids can be quantified by calculating the rate of acetaminophen absorption from serial plasma concentrations. As acetaminophen concentrations in saliva are well correlated with those in plasma, the salivary concentrations may be suitable for use in GER measurement. To evaluate such suitability, salivary and plasma samples were simultaneously obtained from seven healthy volunteers at 0, 0.25, 0.5, 0.75, 1.0, 1.5, and 2.0 h after they had ingested 20 mg/kg of acetaminophen mixed with a 200-ml liquid meal (200 kcal). Commonly used parameters for the rate of acetaminophen absorption were calculated from the salivary and plasma data, including the maximum concentration (Cmax), the time to Cmax (t(max)), the concentration at 0.75 h (C0.75), the area under the curve from 0 to 1.0 h (AUC1.0), and the AUC(0.5)/AUC(2.0) ratio. The mean (SD) salivary/plasma concentration ratio was 2.48 (1.47) at 0.25 h, and the means were almost unity afterwards. Significant correlations between saliva and plasma were found in all parameters studied (r = 0.77-0.90; P < 0.05). However, except for t(max), the salivary parameters overestimated those of plasma. The present results suggest that: (1) the salivary acetaminophen concentration at 0.25 h (C0.25) is a poor reflection of plasma C0.25 (2) thereby the parameters embodying salivary C0.25 such as AUC1.0 and the AUC0.5/AUC2.0 ratio, are unreliable, and (3) liquid GER can be assessed by salivary t(max) with minimal distress to the patient. PMID- 10864350 TI - Loss of diurnal variation in ornithine decarboxylase and apoptosis in small intestine of Mongolian gerbils. AB - The aim of this study was to examine the diurnal variations in ornithine decarboxylase (ODC) activity and apoptosis in the small intestinal mucosa of Mongolian gerbils. First, the feeding, drinking, and ambulatory patterns of the gerbils were recorded. The results indicated that Mongolian gerbils lost diurnal cyclicity in feeding, drinking, and ambulatory patterns. Second, ODC activity and apoptosis in the intestinal mucosa of gerbils were characterized by measurements at four time points (11:00, 17:00, 23:00, and 05:00 h; light period, 08:00-20:00 h). Apoptosis was evaluated in terms of percent fragmented DNA (fragmented DNA/total DNA). Neither ODC activity nor apoptosis showed diurnal variation in the jejunal mucosa of the Mongolian gerbils. In 48-h fasted gerbils, ODC activity in the intestinal mucosa decreased, but this activity increased significantly 2 h after refeeding; percent fragmented DNA increased in the intestinal mucosa, and returned to the control level 2 h after refeeding. These results indicate that postprandial local nutrients are an important factor in regulating diurnal variation in ODC activity and apoptosis in the small intestine of gerbils. PMID- 10864351 TI - Multicentric hepatocellular carcinomas tend to grow in more damaged segments of the liver. AB - In 115 patients (68 with liver cirrhosis and 47 without) who underwent curative resection of hepatocellular carcinoma (HCC) caused by hepatitis C virus (HCV) related chronic liver diseases, we separated the liver into three segments (right, middle, and left) according to the three secondary branches of the Glissonean pedicle. We examined the weight of each resected segment. We also examined the histological findings of the segments in the same liver in 24 other patients with HCV-related chronic liver diseases. The average weight of the segments did not vary significantly in patients without liver cirrhosis. However, the average weight of the segments was significantly different in patients with liver cirrhosis (P = 0.0414) and the weight of the middle segment was lower than that of the other segments. In another group, of 246 patients with curative resection of HCC, of the 90 patients with single nodular HCCs, 45 nodules (50%) were located in the middle segment (P = 0.0004); in the 156 patients with synchronous multicentric HCCs (total, 401 nodules), 220 nodules (54.9%) were located in the middle segment. In 74 of the 156 patients with synchronous multicentric HCCs (47.4%), the HCCs were located in the same segment. The grade, stage of hepatitis, and number of sites of irregular regeneration were significantly different in each segment (P < 0.05), and the middle segment had more advanced hepatitis than the other segments. The rate of occurrence of HCC in the middle segment was higher than that in the other segments. The difference among the segments of the liver in regard to the degree of damage done by hepatitis may be related to the differences in HCC occurrence among the liver segments. PMID- 10864352 TI - Placement of endoscopic naso-biliary drainage does not preclude subsequent percutaneous transhepatic biliary drainage. AB - We prospectively investigated whether the placement of endoscopic naso-biliary drainage (ENBD) precluded percutaneous transhepatic biliary drainage (PTBD). In 40 patients, the caliber of the intrahepatic bile duct was measured prior to ENBD by ultrasonography. When PTBD was required after ENBD, the ENBD catheter was clamped for 1 to 2 h before PTBD, and its caliber was again measured at the time of PTBD. When PTBD was performed within 7 days (mean, 1.8 days) after ENBD (n = 27), the size of the intrahepatic bile duct was 5.0 +/- 2.3 mm before and 4.6 +/- 2.3 mm after ENBD. There was no significant difference between these values (P > 0.5). When PTBD was performed 8 to 40 days (mean, 17.8 days) after ENBD (n = 13), the bile duct diameter was significantly reduced, from 4.2 +/- 1.5 mm (pre-ENBD) to 1.8 +/- 1.7 mm (post-ENBD) (P < 0.05). When PTBD was conducted within 7 days (mean, 1.8 days) after ENBD, previous ENBD did not induce collapse of the bile duct, if the ENBD catheter was clamped for 1 to 2 h before the puncture of the bile duct. PMID- 10864353 TI - Bilirubin overload modulates bile canalicular membrane fluidity in rats: association with disproportionate reduction of biliary lipid secretion. AB - We recently demonstrated that several organic anions cause dissociation of biliary lipid secretion from that of bile acids; namely, the "uncoupling phenomenon," in association with changes in the phospholipid molecular species in the canalicular membrane lipid bilayer. Because of the uncoupling phenomenon, transcytotic vesicles are retained inside cells, resulting in the accumulation of substances normally excreted in the bile. In the present study, bilirubin ditaurate (BDT; synthetic bilirubin) was used to investigate the effect of bilirubin overload on biliary lipid secretion and the lipid composition of hepatic subcellular fractions, as well as canalicular membrane packing density and fluidity. Male Sprague-Dawley rats underwent cannulation of the bile duct and femoral vein. Sodium taurocholate was infused intravenously at 100 nmol/min per 100 g body weight. Then BDT (50 nmol/min per 100 g body weight) was infused concomitantly, followed by periodic bile collection for analysis of lipids. Bile acid secretion was not significantly affected by the infusion of BDT. In contrast, the secretion of cholesterol and phospholipids was decreased by 56.7% and 49.2%, respectively, compared with control. The phosphatidylcholine hydrophobicity of canalicular membrane vesicles, estimated by the molar ratio of saturated to unsaturated fatty acids (S/U ratio) was decreased, but not significantly by BDT infusion. With BDT infusions, the biliary cholesterol/phospholipid (C/P) ratio was increased by 19%; canalicular membrane vesicle fluidity was decreased by 5.8%, whereas P-glycoprotein expression was unchanged. As P-glycoprotein expression was not altered, our findings suggested that the reduced canalicular membrane vesicle fluidity was a crucial regulator of canalicular membrane transporter function. PMID- 10864354 TI - Clinical and histopathological features of colonic stromal tumor in a child. AB - Neoplasms of the colonic submucosa are rare in children. Gastrointestinal stromal tumors (GISTs) are undifferentiated tumors, usually diagnosed by immunohistochemistry. We report a 4-year-old girl with a submucosal GIST of the ascending colon, which was detected by computed tomography. Diagnosis after ileocecal resection was established by histology. In addition, sections were examined immunohistochemically, using antibodies against vimentin, desmin, alpha smooth muscle actin, S100, neuron-specific enolase, c-kit, and CD34. Hematoxylin and eosin-stained sections showed interlacing fascicles with occasional palisades of epithelioid and spindle cells. The tumor cells were positive for vimentin and CD34. To our knowledge, this is the first reported case of colonic stromal tumor in a child. PMID- 10864355 TI - Acrokeratosis paraneoplastica (Bazex syndrome) with adenocarcinoma of the colon: report of a case and review of the literature. AB - Acrokeratosis paraneoplastica is a rare disease and is uncommon even in patients with upper aerodigestive tract cancer. We report a 63-year-old man with a 1-month history of numerous pruritic lesions and vesicles on both feet. Although he had received local therapy, progressive dense scale formation involving both palms and both soles was found. Colonoscopy was performed because of hematochezia, and it revealed an early colon cancer. After the resection of the cancer, the skin lesions began to fall off dramatically. To the best of our knowledge, there is no report of acrokeratosis paraneoplastica associated with colon cancer in the literature. This is the first case report of acrokeratosis paraneoplastica associated with early colon cancer. PMID- 10864356 TI - Triple carcinomas of the biliary tract associated with congenital choledochal dilatation and pancreaticobiliary maljunction. AB - We report a rare case of triple carcinomas of the biliary tract associated with congenital choledochal dilatation (CCD) and pancreaticobiliary maljunction (PBM). The patient was a 58-year-old Japanese man who complained of epigastralgia. Ultrasonography and computed tomography revealed an elevated lesion inside the markedly dilated extrahepatic bile duct, thickening of the gallbladder wall, and small polypoid lesions in the gallbladder. Magnetic resonance cholangiopancreatography and endoscopic retrograde cholangiopancreatography showed CCD and PBM. With a diagnosis of carcinoma of the bile duct and cholesterol polyps in the gallbladder, pylorus-preserving pancreaticoduodenectomy was performed. The resected specimen showed two elevated lesions in the dilated bile duct, cholesterol polyps, and an area of irregular mucosa in the gallbladder. Histopathological examination showed two carcinomas in the bile duct, an adenosquamous cell carcinoma, and a moderately differentiated tubular adenocarcinoma, and a well differentiated tubular adenocarcinoma of the gallbladder. Two years and 6 months after the operation, a solitary metastatic liver tumor was detected. Left hepatic lobectomy was performed. At present, 7 months after the second operation, the patient is doing well with no signs of recurrence. Multiple carcinomas in the biliary tract associated with CCD and PBM, including the details in the present patient, were reviewed. PMID- 10864357 TI - Macrocystic serous cystadenoma of the pancreas: importance of co-existent tiny cysts depicted by EUS. AB - The case of a 38-year-old man with an unusual type of serous cystadenoma of the pancreas is reported. A multilocular cystic tumor in the head of the pancreas was detected on abdominal ultrasonography and computed tomography. On endoscopic ultrasonography, the major cysts ranged from 2.0 to 4.5 cm in size. In addition to these large cysts, a few small cysts were detected. Based on these findings, this tumor was diagnosed as a macrocystic type serous cystadenoma. Because endoscopic retrograde pancreatogram showed a compression of the main pancreatic duct around the tumor, and because the size of the tumor had been increasing over a 3-year period, surgical intervention was performed. The resected tumor consisted of macrocysts, with a few small cysts, and was histologically diagnosed as serous cystadenoma. Endoscopic ultrasonography appears to provide an excellent inside image of this unusual tumor, and because of its ability to detect small cystic lesions clearly, it could be useful in the diagnosis of macrocystic serous cystadenoma. PMID- 10864358 TI - Routine gastric emptying tests. PMID- 10864359 TI - Liver cirrhosis: lobar differences in disease progression and carcinogenesis. PMID- 10864360 TI - Inhibition of biliary phospholipid and cholesterol secretion by organic anions affects bile canalicular membrane composition and fluidity. PMID- 10864361 TI - Diagnostic time in total colonoscopy shortened using special test meals. PMID- 10864362 TI - Immunohistochemical expression of cyclooxygenase (COX)-2 in colorectal adenomas. PMID- 10864363 TI - Acute effects of tumor necrosis factor-alpha on testicular germ cell apoptosis and vascular neutrophil adhesion in rats. AB - Acute effects of tumor necrosis factor-alpha (TNF-alpha) on testicular germ cell apoptosis and vascular neutrophil adhesion were investigated in rats. Acute administration of TNF-alpha did not show a significant increase in percentage of apoptotic tubules and apoptotic germ cells. TNF-alpha administration did not enhance neutrophil adhesion score in the intertubular and subtunical vein. TNF alpha does not acutely influence testicular germ cell apoptosis and vascular neutrophil adhesion in rats. PMID- 10864364 TI - Role of prosaposin in the male reproductive system: effect of prosaposin inactivation on the testis, epididymis, prostate, and seminal vesicles. AB - SGP-1/prosaposin can be secreted or targeted to the lysosomes where it is processed into smaller saposins (A, B, C, and D) required for the hydrolysis of glycosphingolipids. The deficiency of saposins B and C results in variant forms of metachromatic leukodystrophy and Gaucher's disease, respectively, which are characterized by lysosomal storage of undegraded glycosphingolipids. In the nervous system, prosaposin presents trophic activity. A mouse model was recently developed by creating a null allele in embryonic stem cells through gene targeting to investigate the phenotypic diversity of prosaposin mutations and the involvement of this protein in lysosomal storage diseases, and for the development of therapeutic approaches. Mice homozygous mutants die at the age of 35-40 days and neurological disorders contribute to the early demise of the mutant mice. The male reproductive organs in homozygous mutants show several abnormalities, such as a decrease in testis size with reduced spermiogenesis and an involution of the prostate, seminal vesicles, and epididymis. In these animals, the blood levels of testosterone remain normal. In the prostate of homozygous mutants, only the basal epithelial cells appear to be present, while the secretory cells are absent. These findings suggest that prosaposin may be involved in the development and maintenance of the male reproductive organs, as well as, in cellular differentiation. PMID- 10864365 TI - High-affinity binding of T3 to epididymis nuclei. AB - Thyroid hormones play an important role in epididymal function. Hypothyroid animals experience a significant decrease in the number and forward motility of sperm and a remarkable impairment of epididymal morphology. However, it is yet unknown if such activity is due to direct actions of iodothyronines on the target epididymis. The eventual identification of T3 receptors in the nucleous of epididymal cells becomes relevant. For this reason, the authors searched for specific high-affinity binding of T3 to these nuclei. Twenty prepuberal male Wistar rats were used. The testes and epididymis were approached as one unit through a scrotal incision. The fat-free epididymides were subjected to standard techniques to prepare the nuclei for incubations with 125I-T3 concentrations, ranging from 0.5 x 10(-9) to 2.0 x 10(-11) M. Calculations of association constants and binding capacities were performed according to Scatchard. A single binding site with a Ka of 3.06 +/- 0.6 x 10(9) M(-1) or Kd of 3.26 +/- 0.6 x 10(10) M and a maximal binding capacity of 0.11 +/- 0.02 pmol T3/microg DNA were observed. It is concluded that these nuclei contain a specific T3 receptor. This finding strongly suggests that thyroid hormones have direct effects on the epididymis. PMID- 10864366 TI - Efficacy of intrauterine insemination without ovarian hyperstimulation for male or cervical factor in women aged 40 or over. AB - The efficacy of intrauterine insemination (IUI) for male or cervical factor by age of female partner was determined in a retrospective analysis. Patients who underwent IUI therapy for cervical and/or male factor (n = 281) were classified by age at first IUI cycle: <40 years (n = 232), > or =40 years (n = 49). The indication for IUI was cervical factor if a postcoital test failed to show sperm with good forward progression at time of mature follicle; male factor was diagnosed if the semen analysis demonstrated either low count, low motility, antisperm antibodies, or subnormal hypoosmotic swelling test. Intrauterine insemination was performed in either natural cycles or following ovarian stimulation for the treatment of anovulation or follicular maturation defects. Cumulative probability of ongoing pregnancy (viable at end of first trimester) following 3 cycles of IUI was evaluated. Cumulative probability of ongoing pregnancy following 3 cycles of IUI was 28.2% for the younger group and 0.0% for the older group. The age groups did not differ in terms of infertility history, use of ovarian stimulation, or baseline semen parameters. Thus, the treatment of male and/or cervical factor by IUI is ineffective for women > or =40 years. PMID- 10864367 TI - Schedule to inject in vitro matured oocytes may increase pregnancy after intracytoplasmic sperm injection. AB - To ascertain the value of using immature oocytes in an intracytoplasmic sperm injection (ICSI) program, the authors designed a schedule, at 5 p.m. on day 1 (the day of oocyte retrieval) and at 8 a.m. and 2 p.m. on day 2, to recognize and inject the in vitro matured (IVM) oocytes. For the 1,166 oocytes retrieved in 107 ICSI cycles, 128 (11.0%) were at the stage of metaphase I (MI) and 113 (9.7%) at germinal vesicle. Routine ICSI for metaphase 11 oocytes was performed at 2 p.m. on day 1 (initial ICSI). In culture medium of human tubal fluid with 15% maternal serum, 85.1% (205/241) immature oocytes progressed to maturation in which 16.4% (21/128) of MI oocytes matured at 5 p.m. of day 1. The rate of normal fertilization for IVM oocytes (58.5%) was not significantly different from that of initial ICSI (64.0%). One patient received a transfer of two fertilized IVM oocytes alone that were injected at 5 p.m. of day 1, maturing from the MI stage, and achieved a normal pregnancy. The fertilized IVM oocytes were replaced along with the embryos from initial ICSI for 40 cycles that led to 14 (35%) clinical pregnancies. In 43 fertilized IVM oocytes donated for research, we observed that cleavage (95.3%) to the 2- to 4-cell stage was not distinct from that of initial ICSI (94.6%); however, the percentage of embryos of grade I and II morphology was significantly smaller (24.4% vs. 62.5%). Only five (11.6%) developed to blastocysts in vitro. Twenty-one fertilized IVM oocytes were frozen for future transfer. A schedule to inject IVM oocytes in ICSI cycles may generate more accessible embryos for fresh transfer or cryopreservation to increase the chance of pregnancy, although the embryo quality was relatively poor. PMID- 10864368 TI - Does the hypoosmotic swelling test predict human sperm viability? AB - Human sperm viability is essential for successful fertilization. Eosin Y is the usually accepted method for sperm viability assessment, though the hypoosmotic swelling test has been proposed for the selection of viable spermatozoa in procedures such as intracytoplasmic sperm injection. The present study was designed to determine the value of hypoosmotic swelling test in the prediction of sperm viability. For this purpose, hypoosmotic swelling and eosin Y were performed in parallel and in combination, on both fresh and freeze-thawed semen. Rates for eosin Y were significantly higher than for the hypoosmotic swelling test in fresh semen, with a weak, though significant correlation between the two tests (r = 0.47, p < 0.05). When both tests were performed in succession (hypoosmotic swelling test followed by eosin Y), 14.6% of swollen sperm incorporated the dye. Following exposure to hypoosmotic conditions, sperm viability decreased by 35%. When sperm were killed by freezing, hypoosmotic swelling test rates were higher than eosin Y. Results indicate that these two tests cannot be used interchangeably, since 15% of the swollen sperm apparently died, suggesting that plasma membrane integrity is lost before the capacity to maintain osmotic equilibrium. PMID- 10864369 TI - Role of estrogens in human benign prostatic hyperplasia. AB - The aging process is associated with a progressive decline of plasma testosterone levels, while estrone and estradiol remain unchanged and sex hormone binding globulin (SHBG) increases, with reduction of bioavailable testosterone in prostatic tissue with benign prostatic hyperplasia (BPH) the most important androgen is dihydrotestosterone: with its receptors it is almost uniformly distributed in the epithelial and stromal compartment and is not supranormal. Intraprostatic estrogens and their receptors are elevated and concentrated in the stroma. Androgens may act on the prostate indirectly through the production of growth factors; in human BPH no clear evidence exists on the modulatory effect of estrogens on bFGF, KGF and TGFbeta formation. A western diet, characterized by high fat consumption, predisposes men to BPH, while a diet rich in flavonoids and lignanes, containing phyto-estrogens, lowers this risk. These data suggest that in the medical treatment of BPH, antiestrogens or aromatase inhibitors may be used: however, up to now the clinical results of this treatment are not promising and the improvement of the obstructive symptoms does not exceed that of placebo. A possible explanation of this unsatisfactory result could be that the estrogen reduction secondary to the use of aromatase inhibitors is counterbalanced by the rise of androgen precursors. PMID- 10864370 TI - Sperm morphology and chromatin condensation before and after semen processing. AB - Semen analysis constitutes the most important investigation of male infertility. However, the true anomalies present in defective sperm cells have been only partially characterized. The integrity of the sperm chromatin may play the most important role, particularly in ICSI, where most of the natural selection mechanisms are bypassed. This study was carried out to characterize sperm morphology (strict criteria), to evaluate chromatin condensation and sperm count in native semen as well as after semen preparation by the swim-up technique, and to eventually evaluate any correlation between these parameters. Semen from 90 men was analyzed for the above parameters in both the fresh and processed semen. Whereas the sperm count decreased after sperm preparation by the swim-up technique in comparison to the value in the fresh semen (p < .001), there was an increase in the percentage of morphologically normal (p < .001) and chromatin condensed sperm (p = .99). However, there was no correlation between sperm morphology, chromatin condensation, and sperm count either in the fresh or in the processed semen samples. These results suggest that sperm morphology, sperm count, and chromatin condensation are independent parameters that should be evaluated separately in the assessment of male fertility in an assisted reproduction program. PMID- 10864371 TI - Evidence that sperm with low hypoosmotic swelling scores cause embryo implantation defects. AB - A previous prospective study using matched samples found that sperm with low hypoosmotic swelling (HOS) scores had no adverse effect on fertilization rates but markedly reduced pregnancy and implantation rates. The present study attempted to corroborate or refute the aforementioned study by comparing pregnancy rates in donor-recipient pairs using shared oocytes where the sperm of one of the two males had low HOS scores. The results found no pregnancies from the sperm with low HOS scores versus a 41% live delivered rate for those with normal scores. However, fertilization rates were not affected. This retrospective study thus confirms that sperm with low HOS scores cause implantation defects of the embryos that are formed without affecting fertilization rate, embryo cleavage rate, or embryo quality. PMID- 10864372 TI - Do antisperm antibodies cause functional impairment of the sperm membrane as manifested by a low hypoosmotic swelling test score? AB - Low hypoosmotic swelling (HOS) test scores were found to be associated with lower pregnancy rates. The mechanism seems to be related not so much to impaired fertilization but to inhibition of implantation. The defect may be present in males with normal or subnormal semen specimens. However, anecdotal experience suggested that the subset of males with antisperm antibodies (ASAs) have a higher frequency of low HOS scores. The possibility exists that ASAs may impair the functional integrity of the sperm membrane. The study presented herein, artificially added ASAs to sperm to see if this could lower the HOS score. The study would also determine if chymotrypsin, a protein digestive enzyme, could improve HOS scores, if, in fact, they were lowered by the addition of ASAs. The results did not show a reduction in the HOS scores following the addition of ASAs. Thus, it would appear that there is no cause and effect with the simultaneous presence of low HOS scores and ASAs. Possibly, however, longer exposure or a higher concentration of antibodies is needed to lower HOS scores. PMID- 10864373 TI - Soluble IL-6 receptor levels in the seminal plasma of infertile patients with accessory gland infection. AB - The aim of this study was to confirm the presence of soluble interleukin-6 receptor (SIL-6R) in seminal plasma, to show eventual differences between SIL-6R concentrations in fertile and infertile men, and to evaluate the possible value of measuring these substances for the diagnosis of male accessory gland infection. SIL-6R levels were determined by "sandwich" enzyme immunoassay in the seminal plasma of 82 men divided into 7 groups according to the etiological diagnosis of fertility. The levels of SIL-6R in the seminal plasma of men with infection of the accessory genital glands were statistically significantly different in comparison with other groups (p < .001). The results suggest that urogenital infections may lead to elevated levels of SIL-6R in the seminal plasma. This measurement of SIL-6R in semen may provide clinically useful information for the diagnosis of male accessory gland infection. PMID- 10864374 TI - For the love of the children: the coming out process for lesbian and gay parents and stepparents. AB - This research investigated the coming out decision and process for 23 lesbian and gay custodial stepfamilies. We argue that lesbian and gay stepfamilies represent a unique type of family, distinct from heterosexual stepfamilies and from lesbian and gay families who have children within the context of a lesbian or gay relationship. The coming out process is one developmental challenge that distinguishes lesbian and gay stepfamilies from these other types. Through interviews with both lesbian and gay parents and stepparents we explored their coming out process to significant others: the children, families of origin and ex spouses. We discuss those factors influencing the decision and the consequences that developed in light of these decisions. Findings show that the coming out process for lesbian and gay co-parents is a flexible and familial one, primarily influenced by and centered around the needs of the children. PMID- 10864375 TI - Defining the lesbian/gay community? Market research and the lesbian/gay press. AB - The development of marketing information on the readership of the lesbian/gay press attests to the press's success as a medium and its attraction to advertisers. Yet such marketing data, highly skewed towards affluent, urban, white-anglo gay males, raise some serious problems. Such data has been misused by opponents of lesbian/gay rights as representing the entire lesbian/gay community. They also reflect the change of the lesbian/gay press from a medium representing a minority community to one representing an important niche market. An examination of readership data from five major urban lesbian/gay newspapers illustrates these problems. PMID- 10864376 TI - "Safety" in gay men's personal ads, 1985-1996. AB - A content analysis was conducted of a sample of gay men's personal ads over the period 1985-1996. Ads (591) were analysed, comparing those which contained references to "safety" and those which did not. Approximately 10% of ads included such a reference and this proportion did not change over time. "Safety" was not associated with the personal characteristics of advertisers or their desired partners but was more commonly mentioned by older men. There were strong associations between oral sex, anal sex and mentioning safety. However, the differential risks of HIV transmission associated with oral sex and anal sex were not reflected in the extent of reference to safety, suggesting uncertainty about the risks of HIV transmission associated with oral sex. PMID- 10864377 TI - When lesbians aren't gay: factors affecting depression among lesbians. AB - Research on women and depression has neglected to explore how the factors that put women at risk for depression apply to lesbians. The present study examined four of the risk factors consistently cited in the women and depression literature (relationship status, relationship satisfaction, social support from friends, and social support from family), and two unique factors (outness and relationship status satisfaction), to determine their ability to predict depression among lesbians. Data were collected from 167 lesbians between the ages of 20 and 60. Perceived social support from friends, relationship status satisfaction, and perceived social support from family, were found to be significant predictors, accounting for 17.8% of the variance in depression, as measured by the Center for Epidemiologic Study Depression Scale (CES-D). A second multiple regression equation focused on the 110 lesbians who were in committed relationships, using the variables relationship satisfaction, perceived social support from friends, perceived social support from family, and outness, to determine if relationship satisfaction added to the amount of variance which could be predicted in depression. Social support from friends was the only significant predictor in this equation, accounting for 5.8% of the variance in depression scores. PMID- 10864378 TI - Poppies in a wheat field: exploring the lives of rural lesbians. AB - Many believe that lesbian identity is predicated upon the availability of opportunities in urban life to find information, support, and like others (Bell and Valentine 1995a). Indeed, exploring one's lesbian sense of self often involves identifying with a visible reference group, seeking out social arenas where there are other gays and lesbians, connecting with a local community, and taking part in gay and lesbian oriented activities. For rural residents, these opportunities are mostly unavailable, and the lack of access to information, to a public meeting space, and to connections with other lesbians further hinders the development of social group identity. Given that gay and lesbian identity has as its basis a social reference group, how might rural lesbians develop and sustain their sense of personal and group lesbian identity? With few exceptions (D'Augelli 1989; D'Augelli et al. 1987; Kramer 1995; Krieger 1982) the empirical research conducted on the lives of gays and lesbians has utilized urban and suburban samples. Likewise, research on rural life has omitted the experiences of gay and lesbian residents. Either way, information about rural lesbian life remains mostly uncovered. This pilot study attempts to provide new information about the experiences of rural lesbians. Utilizing focus group interviews, the challenges of sustaining lesbian identity in a rural setting are explored. The data show that for this sample, although rural lesbians initially felt isolated and unsure of how to develop a sense of group identity, the opportunity to connect with a small informal network of friends and acquaintances helped alleviate these problems. Further, because these women have little access to information, public gathering space, or to local gay culture, this network was said to be crucial. Without it, the women feel invisible and isolated, that is, their identity remains unseen. PMID- 10864379 TI - Political tolerance among adolescents towards homosexuals in Spain. AB - This paper studies the willingness that an adolescent has to extend human rights to homosexuals. This willingness defines the concept of political tolerance towards homosexuals. Results of this study show that the rights that adolescents are less willing to extend to homosexuals are those more related to the activities that homosexuals do to recover their rights, as the right of demonstrating on the street. This study also pretends to find those variables more related to political tolerance towards homosexuals. Self-esteem, political experience, support for democratic norms and identification with a group of friends are positively related to political tolerance towards homosexuals, whereas support for violent groups or the identification with a religious group are negatively related. PMID- 10864380 TI - Camping the gothic: que(e)ring sexuality in Truman Capote's Other Voices, Other Rooms. AB - Since its release in the late 1940s, Other Voices, Other Rooms has remained an arguably unpopular novella in the works of Truman Capote; its criticism is thus far from recent. Most critiques explore the work in its historical milieu of Southern-gothic fiction, either intentionally or unintentionally avoiding the very prominent queer themes. This article acknowledges Capote's use of gothic paradigms and the text's process of undermining gothic motifs to highlight its two adolescent queer characters. Moreover, the text's own Camp discourse is the liberating force that extinguishes the looming Southern-gothic background to expose the sexual possibilities for its young characters. Amidst the sea of late forties and fifties fiction that frequently ensconced the death of the homosexual character, this novella serves as an exception: through a humorous Camp aesthetic, the text gives birth to its inherent queer desires. PMID- 10864381 TI - Assessment of anxiety in older adults: current status. AB - In 1994 there were 33.2 million older adults (65 years of age and older) in the United States, and approximately one quarter of these older adults meet diagnostic criteria for some mental disorder. Anxiety is among the most prevalent psychiatric disorder in older adults. However, insufficient research has been conducted on the assessment of anxiety in older adults. The purpose of this article was to provide an overview of issues to consider in assessing anxiety in older adults. First, a discussion of factors that may influence current prevalence and incidence figures is provided. Second, age-related differences in factors that can influence the experience and presentation of anxiety symptoms are considered. Third, age-related factors that can influence the assessment process or outcome are presented. Fourth, a discussion on the important role of multimethod assessment and the psychometric adequacy of available anxiety assessment instruments is presented. Finally, recommendations for clinical assessment and future research are provided. PMID- 10864382 TI - Posttraumatic stress disorder in older adults: a conceptual review. AB - Issues that are salient in understanding posttraumatic stress disorder (PTSD) in older adults are examined in this review. Although this issue has received scattered attention in the literature since introduction of the diagnosis of PTSD to the Diagnostic and Statistical Manual (DSM) in 1980, it is clear that numerous conceptual and defining questions exist in our understanding of the aftermath of trauma exposure in older adults. In approaching this issue, studies pertaining to diagnostic status as well as broader dimensions of psychosocial functioning are examined. Concerns that are unique to older adults are highlighted throughout, with particular attention to areas where additional research is warranted. PMID- 10864383 TI - Personality disorders and coping among anxious older adults. AB - This study examined the interrelationships among anxiety, personality disorders, and coping strategies in anxious older adults (n = 28; age range = 55-89; mean = 66.0), nonanxious older adults (n = 100, age range = 55-79, mean = 64.6 ), and anxious younger adults (n = 132; age range = 17-30; mean = 20.2). Younger participants were college students and older participants were community-based family members of the students or recruits from local senior centers. Participants completed the Coolidge Axis II Inventory, the Coping Orientations to Problems Experienced scale, and the Brief Symptom Inventory. Results indicated that the prevalence of generalized anxiety states was relatively low and similar in both older and younger groups and dependent on measurement scale and criterion. At least one personality disorder was found in 61% of the older persons group; obsessive-compulsive, schizoid, and avoidant were the most frequently assigned personality disorders. Anxious older adults had elevated rates of dependent and avoidant personality disorder compared with nonanxious older adults. Younger anxious persons were found to have significantly greater personality dysfunction compared with older anxious persons. Finally, coping differences existed between older anxious and older nonanxious adults and between older anxious and younger anxious adults. Implications for diagnosis and treatment of anxiety in older adults were discussed. PMID- 10864384 TI - Treatment of anxiety disorders in the elderly: issues and strategies. AB - Clinical practice for the treatment of anxiety disorders in the elderly in general lacks empirical validation and hence is somewhat inconsistent. Extensive clinical experience, along with the knowledge gleaned from studies with a younger population, has led to the development of the following treatment approach. A thorough diagnostic assessment, crucial in planning subsequent treatment, is discussed first along with more general clinical issues. Next. a detailed review of current pharmacologic and psychologic treatments for each of the diagnostic categories of anxiety is described for application to the older patient. Definitive studies regarding the best treatments for anxiety disorders in the elderly are lacking, and further investigation of this area is emphasized. PMID- 10864385 TI - Cognitive-behavior therapy for generalized anxiety in late life: an evaluative overview. AB - Of the pervasive anxiety disorders diagnosed in late life, generalized anxiety disorder (GAD) is the most prevalent. In this paper, the clinical features of GAD among older adults are described, with particular attention to differences in the nature of relevant symptoms among older and younger cohorts. Outcome studies addressing the efficacy of cognitive-behavior therapy (CBT) for younger and middle-aged adults with GAD then are reviewed briefly. Next, early literature investigating the potential usefulness of cognitive-behavioral treatments among older anxious community volunteers is then reviewed and critiqued in some detail. More recent work, some of which is currently in progress, has focused on the efficacy of CBT for older adults with well-diagnosed GAD. This research also is reviewed, and directions for future research in this area are provided. PMID- 10864386 TI - Long saphenous-vein grafts for extracranial and intracranial internal carotid aneurysms amenable neither to clipping nor to endovascular treatment. AB - BACKGROUND: In the treatment of patients with unclippable, uncoilable internal carotid artery aneurysms it may be necessary to use high-flow bypass grafts. The indications, surgical techniques and complications are discussed. METHODS: During a 12- year period, 20 saphenous vein grafts were performed in 20 patients. The 20 aneurysms were located in the prepetrous or intrapetrous internal carotid artery in 7 cases, the intracavernous in 9 and intracranial internal carotid artery in 4. All aneurysms were symptomatic. The vein graft was interposed between the internal carotid artery at the neck and the intrapetrous carotid, from the internal carotid artery at the neck to a branch of the middle cerebral artery, or from the external carotid artery and a branch of the middle cerebral artery, according to the collateral circulation assessed with a 30-minute the balloon test occlusion during electroencephalographic recording. RESULTS: Only one of the 20 grafts occluded. One patient died from a large postoperative cerebral infarction. One patient had cerebral ischemia with transient hemiparesis; and three patients had mixed aphasia that resolved completely within two weeks. Radiological follow-up monitoring was by magnetic resonance imaging, magnetic resonance angiography or digital angiography. Postoperative follow-up ranged from 1 to 12 years (mean 3.7 years). CONCLUSIONS: Our experience in this series suggests that the indications for cerebral revascularization should be widened, even to include patients with adequate collateral circulation, particularly those who have a long life expectancy. PMID- 10864387 TI - Meningiomas of the tuberculum sellae. Our experience in 69 cases surgically treated between 1973 and 1993. AB - BACKGROUND: Meningiomas of the tuberculum sellae are lesions with well-defined characteristics in terms of both site of origin and clinical evolution which require differential diagnosis with lesions of the supra- and para-sellar region. The aim of this study is to point out the importance of the size of the lesion and early identification of symptoms for prompt diagnosis, crucially important for evaluating the reversibility of functional damage, especially of the optic nerves. METHODS: Between 1953 and 1993, 110 cases of tuberculum sellae meningioma were operated at the Neurosurgical Institute, Department of Neurosciences, of Rome "La Sapienza" University; only 69 cases of these cases were operated on after 1973, the year in which microsurgical techniques were introduced into routine surgical practice. Tumor diameter ranged from 3 to 9 cm. Most of the patients presented severe visual loss. RESULTS: Removal was total in 63 cases (91.3%), sub-total in 6 (8.7%). Besides the surgeons firsthand impression, the grade of removal was evaluated by early CT and/or MRI (24-36 hours after surgery). A minimum follow-up of 3 years was taken into consideration for assessment of late functional results. Five patients died in the postoperative period (7.2%). CONCLUSIONS: This study clearly demonstrated that a tumor diameter greater than 4 cm is a critical factor for visual function. The size of the lesion was also found to be extremely important, in that it influences the amount of tumor resection possible. Therefore, in our opinion, alternative types of treatment such as radiosurgery should be confined to the tumor residue. PMID- 10864388 TI - Relationship between Ki-67 labeling index and survival in high-grade glioma patients treated after surgery with tamoxifen. AB - BACKGROUND: The Ki-67 is a nuclear antigen expressed in the G1, S, G2, and M phases of the cell cycle recognizable by the monoclonal antibody MIB-1. The Ki-67 labeling index (LI) is considered as a marker of proliferation (growth fraction rate), even if its use as prognostic indicator of cerebral high-grade gliomas is still debated. The aim of this study is to correlate the Ki-67 LI with survival in patients operated on for a malignant glioma and treated postoperatively with tamoxifen. METHODS: Using the MIB-1 antibody, the Ki-67 antigen staining of surgical specimens was obtained in 26 patients operated on for a malignant cerebral glioma. After operation, 9 patients started to receive 40 mg/day, 8 patients 80 mg/day, and 9 patients 120 mg/day of tamoxifen. In 20 cases one or more cycles of i.v. carboplatin was administered. All patients received radiotherapy (4500-6000 cGy). RESULTS: The overall mean survival rate was 19.8 months and the median 12 months; the 12-month and 24-month survival rates were 47% and 23%, respectively. The Ki-67 LI ranged from 2.3% to 62% (mean 24.1%, median 20.5%). Excluding 2 patients who died during the postoperative period, we analyzed the survival rates of the remaining 24 patients in relation to the value of Ki-67 LI. In relation to the index, patients have been divided into 3 groups, with different survival rates: L) Ki-67 LI < or =10%, with mean survival rate of 30.7 months, M) from 10.1% to 30%, with mean survival rate of 15.8 months, and H) >30%, with mean survival rate of 20.2 months. CONCLUSIONS: Our preliminary observations seem to confirm the efficacy of TAM in modifying the survival of malignant gliomas and seem to indicate that tumors with a lower LI and those with a LI >30% could be associated, if treated with TAM, with a better survival than gliomas belonging to group M, who could have a higher tumor proliferation rate in comparison to group L and a lower response to protein-kinase-C antagonists than group H. PMID- 10864389 TI - Malignant cerebellar astrocytomas: clinico-pathological remarks on 10 cases. AB - BACKGROUND: Malignant glioma represent the 3rd?4th most frequent cause of death from cancer. The cerebellar site is rare and life expectancy with cerebellar anaplastic astrocytoma is still dismall. The growth and clinical-pathological remarks of the tumor, is similar to the others gliomas of the central nervous system. MRI with Gd is the most useful diagnostic approach but lacks of specificity in detecting highly differentiated neoplasia areas. METHODS: Between 1980 and 1994 10 cases of malignant cerebellar astrocytomas were operated at the Neurosurgical Institute, Department of NeuroSciences, of Roma "La Sapienza" University. All patients were investigated pre-and postoperatively by CT scan with i.v. administration and/or MRI with Gd when possible. RESULTS: Of the 10 patients who followed various protocols, 7 died. Average survival was 13.7 months (range 5-21 months). 3 patients were still alive 12, 15 and 18 months after surgery. In 3 cases (50%) there was also radiological evidence of spinal cord spreading. CONCLUSIONS: Like cerebral lesions, malignant cerebellar astrocytoma still a pathology with a real unsatisfactory prognosis. Our experience probably showed that spinal spreading is underestimated. For this reason we believe that, despite the limited number of cases treated so far, it is important to extend postoperatively the radiotherapy to the entire spinal cord in all patients. PMID- 10864390 TI - Management of chronic subdural hematoma in patients treated with anticoagulation. AB - BACKGROUND: The diffusion of the surgical technique of cardiac valve replacement with metallic prostheses, as well as bypass graft in the arterial occlusive disease of the lower extremities, both requiring permanent oral anticoagulation, has increased the number of patients affected by chronic subdural hematoma that can be diagnosed at an earlier stage of this disease with the advent of the CT. METHODS: The records of seven patients with mean GCS = 14.2 and mean clinical grade = 1.85 affected by chronic subdural hematoma and in treatment with anticoagulants were examined retrospectively. All the patients underwent subtemporal craniectomy plus closed drainage or burrhole(s) plus closed drainage after immediate correction of hypocoagulability by administration of vitamin K and fresh frozen plasma and normalization of PA by calcium heparin. RESULTS: Outcome was good for all the patients except one who died because of cerebral herniation due to massive solid subdural hematoma during extracorporeal dialysis. Complications included: intracerebral hemorrhage, solid subdural hematoma, slow brain reexpansion, subdural collection reaccumulation and cerebral embolism. Three patients required re-operation. Mean duration of hospital stay was 18 days with range from 7 to 24 days. CONCLUSIONS: Basing on this retrospective study and the proposed pathophysiology, the guidelines for optimal management of this subgroup of patients are proposed. Recommendations include the immediate correction of hypocoagulability, the appropriate surgical technique and the cautious conversion to oral anticoagulation as well as the appropriate timing of such conversion. PMID- 10864391 TI - Spinal cord stimulation in critical limb ischemia of the lower extremities: our experience. AB - BACKGROUND: Spinal cord stimulation (SCS) improves microcirculatory blood flow, relieves ischemic pain and reduces amputation rate in patients with severe peripheral arterial occlusive disease. AIM: To evaluate the specific prognostic parameters in the prediction of successful SCS and to perform a retrospective data analysis obtained during our patient follow-up. METHODS: 150 patients (97 men, 53 women; mean age: 68 years; range: 46-81) were submitted to implantation of a spinal cord electrical generator for rest pain, and trophic lesions with dry gangrene in severe lower limb ischemia, after failed conservative or surgical treatment. The clinical status was classified as Fontaine's stage III and IV and the main pathology was essentially due to atherosclerosis and diabetic vascular disease. In clinical controls, pedal transcutaneous oxygen tension (TcPO2), ankle and toe pressure Doppler measurements were utilised to select and follow-up the patients. RESULTS: After a mean follow-up of 71 months (range 24-138), pain relief >75% and limb salvage was achieved in 85 patients. In 28 patients was obtained a partial success with pain relief >50% and limb salvage for at least 6 months, while in 37 patients the method failed or for technical problems the device was removed, and the patients were amputated. TcPO2 was assessed on the dorsum of the foot. Clinical improvement and SCS success was associated with the increasing of TcPO2, before and after implantation (temporary period). Limb salvage was achieved in the patients that presented significant TcPO2 changes within the first 2 weeks of the testing period, indifferent from the stage of the disease, and from the initial TcPO2 value. After long-term patient follow-up TcPO2 changes, from 22.6 to 43.1 mm Hg in these with rest pain (p<0.01), from 16.2 to 36.1 mmHg (p<0.02) in those with trophic lesions <3 cm2, and from 12.4 to 28.1 in the patients with trophic lesions >3 cm2. A TcPO2 increase of more than 50% in the first 2 months after implantation was predictive of success, and was related with the presence of adequate paresthesias in the painful area during the trial period. The systolic ankle/brachial blood pressure index did not change under stimulation. CONCLUSIONS: In patients with failed conservative and surgical treatment for severe critical lower limb ischemia, the SCS increases the skin blood flow, is associated with a significant pain relief and could be proven an excellent alternative therapy that improves the quality of life. TcPO2 changes, within a test period of 2 weeks, is a predictive index of therapy success and should be considered before the final decision in terms of cost effect, for the permanent implantation. PMID- 10864392 TI - Ulnar nerve compression secondary to ulnar artery true aneurysm at Guyon's canal. AB - This article presents a case of ulnar nerve compression at the Guyon's canal caused by a true aneurysm of the ulnar artery secondary to blunt trauma. The duration of follow-up was one year. SETTING: Hospitalized care. A 27-year-old man who worked in an office fell on to a gravel path landing on his out-stretched right hand. Decompression of the ulnar nerve was made by simple ligation of the damaged artery and resection of aneurysm. MEASURES: Histological examination. The sensory symptoms disappeared two days after the operation. At one year after surgery, the patient was completely asymptomatic. There was no residual cold intolerance. Simple ligation of the damaged artery and resection of aneurysm resulted satisfactory. It seemed to be a safe method in this case. Ulnar nerve compression due to a true aneurysm of the ulnar artery in the Guyon's canal is rarely described in the literature. PMID- 10864393 TI - Dorsally exophytic brain stem tumors: total removal of a medullary pilocytic astrocytoma in child. Clinicopathological considerations and case report. AB - The authors describe the case of 4-year-old girl who was operated on for a pilocytic astrocytoma of the brain stem, exophytic and protruded backward into the fourth ventricle. The removal through a suboccipital midline approach was performed and macroscopically complete. Postoperative complications compelled a heavy reanimation with a prolonged follow-up therapy. The neurological recovery was favorable. Three years later, this little girl is almost autonomous in all daily activities. Postoperative scans showed total tumor resection. The patient received no complementary treatment such as radiotherapy, chemotherapy or corticotherapy. The long-term survival, without the need for adjunctive therapy, confirms the good prognosis for this unusual group of brain stem tumors. The clinicopathological and radiological features are described and surgical questions are discussed especially regarding complete or subtotal removal of brain stem tumors in demarcated-benign astrocytomas. PMID- 10864394 TI - Extradural low cervical chordoma. Case report. AB - Chordoma of the mobile segments of the spine are infrequent lesions and especially rare are those located in the lower part of the cervical spine. We present the case of a cervical chordoma located in the C6 vertebral body diagnosed by means of magnetic resonance imaging and operated on by an anterior approach. The authors discuss the clinical and neuroradiological features of this disease also analysing some controversial therapeutic aspects. PMID- 10864395 TI - An unusual management of an open compound depressed skull fracture with venous sinus involvement. A case report. AB - A neurosurgical management of an open compound depressed fracture perforating the superior sagittal sinus is reported. Undue bleeding from the fracture did not allow a conservative management. The patient had been operated primarily at an outside emergency surgery unit. Profuse uncontrollable bleeding made a tamponade of the sinus necessary for transportation to our neurosurgical department. After reconstruction of the sinus he survived without evidence of a neurological deficit. PMID- 10864396 TI - Modulation of allergen and anti-igE induced human basophil activation by serial histamine dilutions. PMID- 10864397 TI - Cytokine release from a human mast cell line (HMC-1) in response to stimulation with anti-IgE and other secretagogues. PMID- 10864398 TI - Early IgE-dependent release of IL-4 and IL-13 from leukocytes is restricted to basophils: a comparison with other granulocytes and mononuclear cells. PMID- 10864399 TI - Some characteristics of mast cells cultured from human umbilical cord blood. PMID- 10864400 TI - In vitro studies with fexofenadine, a new nonsedating histamine H1 receptor antagonist, on isolated human basophils. PMID- 10864401 TI - Differential effect of interleukin-3, stem cell factor and nerve growth factor on histamine and serotonin release from rat mast cells. PMID- 10864402 TI - Ambroxol inhibits IgE-dependent mediator secretion from human skin mast cells. PMID- 10864403 TI - Changes in mast cell phenotype in rat lung following infection with Nippostrongylus brasiliensis. PMID- 10864404 TI - Inhibition of compound 48/80 induced histamine release from mast cells by chloride channel blockers is affected by methods of drug preincubation. PMID- 10864405 TI - Value of baseline levels in assessing tryptase release. PMID- 10864406 TI - Is lymphocyte histamine involved in the pathogenesis of rheumatoid arthritis? PMID- 10864407 TI - Reciprocal inhibitory interactions between interferon gamma and histamine in melanoma. PMID- 10864408 TI - IgE-mediated immune response to Helicobacter pylori examined by basophil histamine release in patients with dyspepsia. PMID- 10864409 TI - Is histamine releasability of mast cells related to hormonal status? PMID- 10864410 TI - Placental mast cell heterogeneity in pregnancy complicated by diabetes class C. PMID- 10864411 TI - Methylhistamine in Crohn's disease (CD): increased production and elevated urine excretion correlates with disease activity. PMID- 10864412 TI - Chlamydia pneumoniae and chronic obstructive pulmonary disease: bronchial biopsies (PCR and culture) and specific serum antibodies in patients and controls. PMID- 10864413 TI - Detection of specific T lymphocytes in systemic abnormal delayed type hypersensitivity to Candida albicans. PMID- 10864414 TI - Ciproxifan and cimetidine modulate c-fos expression in septal neurons, and acetylcholine release from hippocampus of freely moving rats. PMID- 10864415 TI - Thioperamide and cimetidine modulate acetylcholine release from the amygdala of freely moving rats. PMID- 10864417 TI - Regional distribution of histamine H3 receptor binding sites in rat brain following L-histidine loading--an autoradiography study. PMID- 10864416 TI - The effect of in vivo deprenyl on cerebral amine system in rats with portocaval shunts. PMID- 10864418 TI - Clinical evaluation of a new fully automated enzyme immunoassay for basophil histamine release in whole blood. PMID- 10864419 TI - Analysis of mammalian diamine oxidase genes. PMID- 10864420 TI - Postheparin plasma diamine oxidase activity and the anticoagulant effect of heparin. PMID- 10864421 TI - Inhibition of diamine oxidase activity by heparins. PMID- 10864422 TI - Characterization of cDNA clones encoding mouse diamine oxidase. PMID- 10864423 TI - Immobilization of plant histaminase for medical applications. PMID- 10864424 TI - Pharmacological characterization of H3 receptor antagonists with imidazole, thioimidazole and thioimidazoline polar groups in the side chain. PMID- 10864425 TI - Possible role of histidine decarboxylase (HDC) and histamine in human in vitro monocyte-macrophage differentiation. PMID- 10864426 TI - Regulation of murine hematopoietic colony formation by histamine. PMID- 10864427 TI - Human pancreatic carcinoma cell line Panc-I and the role of histamine in growth regulation. PMID- 10864428 TI - Turnover of histamine in human melanoma cell lines. PMID- 10864429 TI - Histamine and H2 agonist amthamine increase oxidative burst activity of human leukocytes determined by flow cytometry. PMID- 10864430 TI - Kinetics of injected histamine in cat plasma after antihistamine prophylaxis. PMID- 10864431 TI - Induction of heme oxygenase provides protection against cardiac anaphylaxis. PMID- 10864432 TI - Dual role of nitric oxide in myocardial ischemia-reperfusion. PMID- 10864433 TI - Histamine changes in small bowel circulation measured by microdialysis in pig hemorrhagic shock. Shock and Trauma Study Group. PMID- 10864434 TI - The heart protective/damaging effects of short/long lasting mesenteric ischaemia may be mimicked by exogenous histamine. PMID- 10864435 TI - Reversible sodium dodecyl sulfate activation of latent peach polyphenol oxidase by cyclodextrins. AB - The reversibility of the SDS-mediated activation of latent peach PPO has been studied using cyclodextrins as strip detergent agent. Cyclodextrins produced a combined inhibitory effect on enzymatic activity of latent peach PPO due to the complexation of detergent and the hydrophobic substrate 4-tert-butylcatechol (TBC) molecules. To study the reversibility of the activation process, this combined effect has to be separated. On the one hand, the enzyme was activated by acid-shocking and the activity was measured in the presence of cyclodextrins, using TBC as substrate. The inhibition curves obtained permitted study of the complexation of TBC into cyclodextrins. On the other hand, the enzyme was activated by SDS and the activity in the presence of cyclodextrins was measured using the highly hydrophilic o-diphenol dopamine as substrate. In this case, the inhibition curves obtained indicated the reversibility of the activation process when SDS was trapped by cyclodextrins. In addition, the complexation constant between SDS and 2-hydroxypropyl-beta-cyclodextrins was calculated by measuring conductivity (K(s) = 3500 M(-1)). PMID- 10864436 TI - Enhanced tissue expression and elevated circulating level of phospholipase A(2) receptor during murine endotoxic shock. AB - Phospholipase A(2) receptor (PLA(2)R) mediates a variety of biological responses elicited by mammalian secretory phospholipase A(2) (sPLA(2)). In mice, group IB sPLA(2) (sPLA(2)-IB) acts as an endogenous ligand of PLA(2)R, and analysis of PLA(2)R-deficient mice has demonstrated a critical role of the sPLA(2)-IB/PLA(2)R system in the production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) in the development of endotoxic shock. Here, we generated specific antibodies against a recombinant soluble form of PLA(2)R and examined its expression in the lung and spleen where a remarkable elevation of TNF-alpha expression has been observed during endotoxemia. Immunohistochemical analysis revealed the expression of PLA(2)R in type II alveolar epithelial cells and a subset of splenic lymphocytes, and its expression levels were markedly enhanced at 1 h after endotoxin challenge. Analysis with a newly developed sandwich enzyme-linked immunosorbent assay system revealed the presence of a soluble form of PLA(2)R in plasma of wild-type mice compared with its absence in plasma of PLA(2)R-deficient mice. After exposure to endotoxin, its circulating level was significantly elevated to the maximum level at 2-3 h after the treatment. These results suggest that tissue expression and the circulating level of PLA(2)R are elevated during murine endotoxemia, which might be relevant to its potential roles in the production of proinflammatory mediators during the development of inflammatory conditions. PMID- 10864437 TI - NMR solution structure of butantoxin. AB - The NMR structure of a new toxin, butantoxin (BuTX), which is present in the venoms of the three Brazilian scorpions Tityus serrulatus, Tityus bahiensis, and Tityus stigmurus, has been investigated. This toxin was shown to reversibly block the Shaker B potassium channels (K(d) approximately 660 nM) and inhibit the proliferation of T-cells and the interleukin-2 production of antigen-stimulated T helper cells. BuTX is a 40 amino acid basic protein stabilized by the four disulfide bridges: Cys2-Cys5, Cys10-Cys31, Cys16-Cys36, and Cys20-Cys38. The latter three are conserved among all members of the short-chain scorpion toxin family, while the first is unique to BuTX. The three-dimensional structure of BuTX was determined using (1)H-NMR spectroscopy. NOESY, phase sensitive COSY (PH COSY), and amide hydrogen exchange data were used to generate constraints for molecular modeling calculations. Distance geometry and simulated annealing calculations were performed to generate a family of 49 structures free of constraint violations. The secondary structure of BuTX consists of a short 2(1/2) turn alpha-helix (Glu15-Phe23) and a beta-sheet. The beta-sheet is composed of two well-defined antiparallel strands (Gly29-Met32 and Lys35-Cys38) connected by a type-I' beta-turn (Asn33-Asn34). Residues Cys5-Ala9 form a quasi-third strand of the beta-sheet. The N-terminal C2-C5 disulfide bridge unique to this toxin does not appear to confer stability to the protein. PMID- 10864438 TI - Spectroscopic characterization of the interaction between calmodulin-dependent protein kinase I and calmodulin. AB - Calmodulin-dependent protein kinase I (CaM kinase I) is a member of the expanding class of protein kinases that are regulated by calmodulin (CaM). Its putative CaM binding region is believed to occur within a 22-residue sequence (amino acids 299 320). This sequence was chemically synthesized and utilized for CaM interaction studies. Gel band shift assays and densitometry experiments with intact CaM kinase I and the CaM-binding domain peptide (CaMKIp) reveal that they bind in an analogous manner, giving rise to 1:1 complexes. Fluorescence analysis using dansyl-CaM showed that conformational changes in CaM on binding CaM kinase I or CaMKIp were nearly identical, suggesting that the peptide mimicked the CaM binding ability of the intact protein. In the presence of CaM, the peptide displays an enhancement of its unique Trp fluorescence as well as a marked blue shift of the emission maximum, reflecting a transfer to a more rigid, less polar environment. Quenching studies, using acrylamide, confirmed that the Trp in the peptide on binding CaM is no longer freely exposed to solvent as is the case for the free peptide. Studies with a series of Met mutants of CaM showed that the Trp containing N-terminal end of CaMKIp was bound to the C-terminal lobe of CaM. Near UV CD spectra also indicate that the Trp of the peptide and Phe residues of the protein are involved in the binding. These results show that the CaM-binding domain of CaM kinase I binds to CaM in a manner analogous to that of myosin light chain kinase. PMID- 10864439 TI - Characterization of the mouse lanosterol 14alpha-demethylase (CYP51), a new member of the evolutionarily most conserved cytochrome P450 family. AB - Genes encoding sterol 14alpha-demethylases in eukaryotes and in Mycobacterium belong to the CYP51 family which is evolutionary the most conserved gene family within the cytochrome P450 superfamily. We have characterized a new member of this family, the mouse lanosterol 14alpha-demethylase, with the aim to study the in vivo role of this gene in spermatogenesis in mammals. The amino acid sequence of mouse Cyp51 is 96% identical to rat and 91% to human. Comparison of all known CYP51 proteins by the neighbor-joining method suggests that fungal and animal CYP51 genes arose from a common ancestral gene (98.3% probability) and interestingly, that plant and bacterial CYP51 genes share a common progenitor (88.8% probability). This suggests that the first CYP51 gene may have arisen in eukaryotes and has been transferred horizontally from plants to Mycobacterium. The mouse CYP51 gene is approximately 17-kb long and contains 10 exons. Transcription starts at several locations within the CpG island, which is characteristic for the TATA-less housekeeping genes. The mouse 5'-untranslated region (800 bp) contains putative cAMP-responsive elements (CRE), sterol regulatory elements (SRE) and GC-boxes at positions similar to human and rat, suggesting an evolutionary conserved mechanism of CYP51 transcriptional regulation in mammals. The mouse Cyp51 gene resides on chromosome 5, region A2, close to the centromere. No signals outside this region were detected as well as no evidence of processed pseudogenes using long PCR was found. This indicates that the mouse genome most likely lacks CYP51 processed pseudogenes. PMID- 10864440 TI - Identification and characterization of promoters regulating tuf expression in Chlamydia trachomatis serovar F. AB - Gene expression in the obligate intracellular bacterium Chlamydia trachomatis ranges from nil in the infectious EB form to high in the dividing RB form. Little is known about the mechanisms of gene regulation in chlamydiae and only a few promoter sequences have been characterized. The purpose of our study was to examine the expression of a cluster of genes that are required for translation in C. trachomatis serovar F: infA (encoding Initiation Factor 1), tRNA(Thr), tuf (encoding Elongation Factor Tu), and tRNA(Trp). Primer extension analysis indicated that tuf is expressed in three different mRNAs. Putative promoter sequences for these transcripts were defined as P1 (upstream of tRNA(Thr)), P2 (within infA) and P3 (upstream of infA). Quantitative RT-PCR analysis revealed that P1 transcripts were most abundant at 16 h postinfection (pi), whereas P2 transcripts predominated at 24 h pi. P3 was active at all times pi; however, transcription terminated upstream of tuf at early times pi and continued through tuf at later times. P1 and P3 were active in Escherichia coli, as assessed by CAT expression in promoter-fusion vectors and a chlamydial in vitro transcription system. Site-specific mutagenesis confirmed the importance of the -35 and -10 hexamers in the P1 and P3 promoters. P2 was weakly active in E. coli and inactive in the in vitro transcription system, indicating either that the P2 transcript is processed from a longer transcript or that P2 expression requires a sigma or transcription factor which is not present in E. coli or the in vitro transcription system. Our data suggest that multiple processes play a role in the regulation of tuf gene expression during the developmental cycle. PMID- 10864441 TI - Mechanical and physicochemical regulation of the action of insulin-like growth factor-I on articular cartilage. AB - The development and maintenance of healthy joints is a complex process involving many physical and biological stimuli. This study investigates the interaction between insulin-like growth factor-I (IGF-I) and static mechanical compression in the regulation of articular cartilage metabolism. Bovine cartilage explants were treated with concentrations of IGF-I from 0 to 300 ng/ml in the presence or absence of 0-50% static compression, and the transient and steady-state incorporation of [(3)H]proline and [(35)S]sulfate into matrix components were measured. In parallel studies, cartilage explants were treated with 0-300 ng/ml IGF-I at media pH ranging from 6.4 to 7.2 and the steady-state incorporation of [(3)H]proline and [(35)S]sulfate was measured. The effect of 50% static compression on IGF-I transport was determined by measuring the uptake of (125)I labeled IGF-I into cartilage explants. Static compression decreased both [(3)H]proline and [(35)S]sulfate incorporation in a dose-dependent manner in the presence or absence of IGF-I. IGF-I increased [(3)H]proline and [(35)S]sulfate incorporation in a dose-dependent manner in the presence or absence of compression, but the anabolic effect of the growth factor was lessened when the tissue was compressed by 50%. The response of cartilage explants to IGF-I was similarly lessened in unstrained tissue cultured in media at pH 6.4, a condition which results in a similar intratissue pH to that when cartilage is compressed by 50%. The characteristic time constant (tau) for IGF-I stimulation of cartilage explants was approximately 24 h, while tau for inhibition of biosynthesis by static compression was approximately 2 h. Samples which were both compressed and treated with IGF-I demonstrated an initial decrease in biosynthetic activity at 2 h, followed by an increase at 24 h. Static compression did not alter tau for (125)I-labeled IGF-I transport into cartilage but decreased the concentration of (125)I-labeled IGF-I in the tissue at equilibrium. PMID- 10864442 TI - Effects of intraoral splint wear on proteoglycans in the temporomandibular joint disc. AB - Intraoral splints are a common dental treatment for dysfunctions of the temporomandibular joint (TMJ), but their effects on the structures of the joint, specifically the disc, have not been well investigated. This study examined proteoglycans (PGs) of the TMJ disc of the miniature pig and tested for alterations resulting from intraoral splint wear. Sixteen female pigs were divided into three groups: control (C), control splint (CS), and protrusive splint (PS). Splinted groups received chrome-cobalt ramp splints which were worn continuously for 2 months. PG content within various disc locations was determined by colorimeteric assay. PG synthesis and type were examined by labeling with (35)S-sulfate and SDS-PAGE analysis. Average water content of the disc was 77.1%, which places it at the high end of the normal range for collagenous biomaterials (60-80%). PGs migrating to the positions typical of aggrecan, biglycan, and decorin on SDS-PAGE were present in all locations of all groups. The highest content and synthesis of PGs were always found in the intermediate band of the disc regardless of group (P < 0.05), supporting the notion that this band encounters heavy compressive loading during function. The joints of animals from both splinted groups showed a high frequency of gross pathology. Biglycan synthesis was increased in both splinted groups (P < 0.05). Newly synthesized biglycan had a shorter migration distance in the intermediate bands of the CS group, suggesting increased hydrodynamic size. These findings suggest that intraoral splint wear may cause disc damage or remodeling. PMID- 10864443 TI - Lysyl oxidase and P-ATPase-7A expression during embryonic development in the rat. AB - Lysyl oxidase activity is critical for the assembly and cross-linking of extracellular matrix proteins, such as collagen and elastin. Moreover, lysyl oxidase activity is sensitive to changes in copper status and genetic perturbations in copper transport, e.g., mutations in the P-type ATPase gene, ATP7A, associated with cellular copper transport. Lysyl oxidase may also serve as a vehicle for copper transport from extracellular matrix cells. Herein, we demonstrate that sufficient lysyl oxidase functional activity is present in the rat embryo at gestation day (GD) 9 to be detected in conventional enzyme assays. Estimation of embryonic lysyl oxidase functional activity, however, required partial purification in order to remove inhibitors. From GD 9 to GD 15, lysyl oxidase activity was relatively constant when expressed per unit of protein or DNA. In contrast, the steady-state levels of lysyl oxidase and ATP7A mRNA, measured by RT-PCR and expressed relative to total RNA and cyclophilin mRNA, increased approximately fourfold from GD 9 to 15. The pattern of temporal expression for ATP7A was consistent with its possible role in copper delivery to lysyl oxidase. PMID- 10864444 TI - Modulation of 2-oxoglutarate dehydrogenase complex by inorganic phosphate, Mg(2+), and other effectors. AB - The interplay of inorganic phosphate (Pi) with other ligands such as Mg(2+), ADP, ATP, and Ca(2+) on the activation of 2-oxoglutarate dehydrogenase complex (2 OGDH) in both isolated enzyme complex and mitochondrial extracts was examined. Pi alone activated the enzyme, following biphasic kinetics with high (K(0.5) = 1.96+/-0.42 mM) and low (K(0.5) = 9.8+/-0.4 mM) affinity components for Pi. The activation by Pi was highly pH-dependent; it increased when the pH raised from 7.1 to 7.6, but it was negligible at pH values below 7.1. Mg-Pi and Mg-ADP, but not Mg-ATP, were more potent activators of 2-OGDH than free Pi and free ADP. ATP inhibited the 2-OGDH activity by chelating the free Mg(2+) and also as a Mg-ATP complex. With or without Mg(2+), ADP, and Pi activated the 2-OGDH by increasing the affinity for 2-OG and the V(m) of the reaction; ATP diminished the V(m), but it increased the affinity for 2-OG in the mitochondrial extract. Pi did not modify the 2-OGDH activation by Ca(2+). The results above mentioned were similar for both preparations, except for hyperbolic kinetics in the isolated enzyme and sigmoidal kinetics in the mitochondrial extracts when 2-oxoglutarate was varied. The data of this study indicated that physiological concentrations of Pi may exert a significant activation of 2-OGDH, which was potentiated by Mg(2+) and high pH, but surpassed by ADP. PMID- 10864445 TI - The role of tyrosine 15 in erythropoietin action. AB - The results of iodination inactivation of erythropoietin suggest that tyrosine 15 is required for biological activity. This was confirmed by site-directed mutagenesis. Substitution of tyrosine by alanine or isoleucine resulted in mutants with no biological activity, whereas substitution by phenylalanine yielded an active mutein. Protein footprinting using trypsin showed that the N terminal residues 1 to 46 and the C-terminal residues 155 to 165 linked by the 7 to 161 disulfide bond, includes one active site of the hormone. PMID- 10864446 TI - Xenopus allantoicase: molecular cloning, enzymatic activity and developmental expression. AB - Allantoicase is one of the enzymes of the purine degradation pathway and, interestingly, it appears to be lost, together with uricase and allantoinase, during mammalian evolution. Only allantoicases from the ascomycetes S. pombe, S. cerevisiae, and N. crassa have already been cloned, although the activity has been reported also in fishes and amphibians. By screening a cDNA expression library of Xenopus liver, we have cloned a 1491-bp-length cDNA coding for a 389 amino acid protein that shows an high similarity with the enzyme allantoicase. We have found that allantoicase mRNA is abundantly expressed in kidney and liver, but at much lower level is also present in brain, testis, intestine, and lung. We have detected enzymatic activity in crude extract from kidney, liver, and lung; we have also determined kinetic parameters (K(m) = 8.44 mM, V(max) = 6. 94 micromol min(-1) per mg protein) in kidney. During embryo development, we have detected allantoicase transcript and activity starting from 1 and 5 days after fertilization, respectively. PMID- 10864447 TI - Molecular basis for cell-specific regulation of the NADPH-cytochrome P450 oxidoreductase gene. AB - NADPH-cytochrome P450 oxidoreductase (CYPOR), a flavoprotein localized in the nuclear envelope and endoplasmic reticulum of most cell types, is responsible for transferring electrons from NADPH to the cytochromes P450 as well as heme oxygenase, squalene epoxidase, and cytochrome b(5). CYPOR is encoded by a single gene and, similar to many housekeeping genes, has a TATA-less, GC-rich promoter with multiple Sp1 consensus sites. The current work has delineated the importance of multiple cis-acting elements contained within the proximal promoter for basal expression of the CYPOR gene. Transcription factor binding sites within this region included two upstream Sp1 motifs, a SEC element containing overlapping Sp1/Egr-1/CACCC box motifs, and a novel site designated the OxidoReductase Upstream element (ORU). Mutational modification of the ORU element, leading to a loss of protein binding, resulted in an approximately 90% decrease in transcriptional activity in H4IIE cells. Similarly, inactivation of the Egr 1/CACCC segment of the SEC element dramatically reduced promoter activity to less than 10% of wild-type, while mutagenesis of the contiguous Sp1 site did not affect basal transcription. Although both Sp1 sites contained within the minimal promoter were required for optimal expression in H4IIE cells, loss of these sites was compensated for by those Sp1 motifs located upstream of position 206, suggesting that Sp1 was acting as a position-independent enhancer. Hence, the CYPOR promoter was distinguished from the majority of TATA-less promoters in that Sp1 was not a primary transcriptional regulator and by the fact that the Sp1 binding site closest to the transcription start site was nonfunctional. Furthermore, both the SEC and ORU elements were essential for basal expression; however, the ORU element exhibited cell-specific differences in regulatory activity. Thus, several mechanisms appear to be in place to selectively alter the expression of the CYPOR gene. PMID- 10864448 TI - Characterization of [(3)H]ryanodine binding sites in mammalian lung. AB - The ryanodine-sensitive calcium channels, also called ryanodine receptors, are intracellular Ca(2+)-release channels that have been shown to bind the neutral plant alkaloid ryanodine with nanomolar affinity. The activity of the skeletal muscle (RyR1), cardiac muscle (RyR2), and brain (RyR3) ryanodine receptor isoforms have been shown to be highly regulated by physiological factors including pH, temperature, and ionic strength; endogenous compounds including Ca(2+), Mg(2+), and adenosine triphosphate (ATP); and pharmacological agents including caffeine, ruthenium red, and neomycin. RyR3 is reportedly expressed in diverse tissues including lung; however, specific [(3)H]ryanodine binding sites in mammalian lung tissue have not been characterized. In this study, hamster lung ryanodine binding proteins were shown to specifically bind [(3)H]ryanodine with an affinity similar to that of RyR isoforms found in other tissues and this binding was shown to be sensitive to Ca(2+) concentration, stimulation by caffeine and spermine, and inhibition by Mg(2+), ruthenium red, and neomycin. The solubilized, intact ryanodine binding protein from hamster lung demonstrated approximately the same 30S sedimentation coefficient as RyR1 and RyR2, but a putative ryanodine receptor subunit from hamster lung was not found to cross react with antibodies specific for the three known isoforms. We conclude that the hamster lung ryanodine binding protein demonstrates sedimentation and binding characteristics that are similar to those of the known RyR isoforms, but may exhibit antigenic dissimilarity from the typical RyR isoforms found in muscle and brain. PMID- 10864449 TI - Covalent binding of LTA(4) to nucleosides and nucleotides. AB - Leukotriene A(4) (LTA(4)) is a chemically reactive conjugated triene epoxide that is formed by 5-lipoxygenase and is an intermediate in the formation of the biologically active eicosanoids leukotriene B(4) and leukotriene C(4). The present study was undertaken to determine whether or not LTA(4) could serve as an electrophilic species that nucleosides and nucleotides could attack, ultimately resulting in a covalent adduct. Electrospray ionization mass spectrometry and tandem mass spectrometry were used to study the covalent binding of LTA(4) with uridine, cytidine, adenosine, and guanosine. The reaction with guanosine was found to yield five major and at least six minor adduct species. Reversed phase HPLC and mass spectrometric data suggested that the guanosine attacked LTA(4) either at carbon-12 or carbon-6 with opening the epoxide at carbon-5 to yield a series of adducts characterized by the molecular anion [M-H](-) at m/z 600.3. Reactions of LTA(4) with mixtures of nucleosides and nucleotides revealed that guanine-containing nucleosides were the most reactive toward LTA(4). The facility of the reaction of guanine with LTA(4) raises the possibility that this intermediate of leukotriene biosynthesis formed on or near the cellular nuclear envelope may react with nucleosides and nucleotides present in RNA or DNA. PMID- 10864450 TI - Functional expression of regiospecific cytochrome P450 limonene hydroxylases from mint (Mentha spp.) in Escherichia coli and saccharomyces cerevisiae. AB - The oxygenation pattern of the essential oil monoterpenes of commercial mint (Mentha) species is determined by regiospecific cytochrome P450-catalyzed hydroxylation of the common olefinic precursor (-)-4S-limonene. In spearmint (M. spicata), C6-allylic hydroxylation leads to (-)-trans-carveol and thence (-) carvone, whereas in peppermint (M. x piperita), C3-allylic hydroxylation leads to (-)-trans-isopiperitenol and ultimately (-)-menthol. cDNAs encoding the C6 hydroxylase and C3-hydroxylase from spearmint and peppermint, respectively, were isolated by a combination of reverse genetic and homology-based cloning methods (S. Lupien, F. Karp, M. Wildung, and R. Croteau, Arch. Biochem. Biophys. 368, 181 192, 1999). Although both hydroxylase genes were confirmed by functional expression using the baculovirus-Spodoptera system, too little protein was available by this approach to permit detailed study of the structure-function relationships of these catalysts, especially the substrate binding determinants that underlie the regiochemistry and stereochemistry of the reactions. Therefore, heterologous overexpression systems based on Escherichia coli and Saccharomyces cerevisiae were developed to produce several N-terminally modified versions of the recombinant hydroxylases. Ancillary methods for the solubilization, purification, and reconstitution (with recombinant spearmint cytochrome P450 reductase) of the limonene hydroxylases were also devised, with which substrate binding behavior and precise regiochemistry and stereochemistry of product formation were determined. PMID- 10864451 TI - Heterologous expression and characterization of a "Pseudomature" form of taxadiene synthase involved in paclitaxel (Taxol) biosynthesis and evaluation of a potential intermediate and inhibitors of the multistep diterpene cyclization reaction. AB - The diterpene cyclase taxadiene synthase from yew (Taxus) species transforms geranylgeranyl diphosphate to taxa-4(5),11(12)-diene as the first committed step in the biosynthesis of the anti-cancer drug Taxol. Taxadiene synthase is translated as a preprotein bearing an N-terminal targeting sequence for localization to and processing in the plastids. Overexpression of the full-length preprotein in Escherichia coli and purification are compromised by host codon usage, inclusion body formation, and association with host chaperones, and the preprotein is catalytically impaired. Since the transit peptide-mature enzyme cleavage site could not be determined directly, a series of N-terminally truncated enzymes was created by expression of the corresponding cDNAs from a suitable vector, and each was purified and kinetically evaluated. Deletion of up to 79 residues yielded functional protein; however, deletion of 93 or more amino acids resulted in complete elimination of activity, implying a structural or catalytic role for the amino terminus. The pseudomature form of taxadiene synthase having 60 amino acids deleted from the preprotein was found to be superior with respect to level of expression, ease of purification, solubility, stability, and catalytic activity with kinetics comparable to the native enzyme. In addition to the major product, taxa-4(5),11(12)-diene (94%), this enzyme produces a small amount of the isomeric taxa-4(20), 11(12)-diene ( approximately 5%), and a product tentatively identified as verticillene ( approximately 1%). Isotopically sensitive branching experiments utilizing (4R)-[4 (2)H(1)]geranylgeranyl diphosphate confirmed that the two taxadiene isomers, and a third (taxa-3(4),11(12)-diene), are derived from the same intermediate taxenyl C4-carbocation. These results, along with the failure of the enzyme to utilize 2, 7-cyclogeranylgeranyl diphosphate as an alternate substrate, indicate that the reaction proceeds by initial ionization of the diphosphate ester and macrocyclization to the verticillyl intermediate, followed by a secondary cyclization to the taxenyl cation and deprotonation (i.e., formation of the A ring prior to B/C-ring closure). Two potential mechanism-based inhibitors were tested with recombinant taxadiene synthase but neither provided time-dependent inactivation nor afforded more than modest competitive inhibition. PMID- 10864452 TI - The status of half-cystine residues and locations of N-glycosylated asparagine residues in human eosinophil peroxidase. AB - The determination by protein chemistry methods of the half-cystine status in human eosinophil peroxidase (EPO) is reported. EPO is two-chained and has a total of 14 half-cystine residues. Cys141 and Cys152 form an intrachain bridge in the light chain of EPO. Disulfide bridges connect Cys253 and Cys263, Cys257 and Cys287, Cys359 and Cys370, Cys570 and Cys635, and Cys676 and Cys701, forming five intrachain disulfide bridges in the heavy chain of EPO. Cys291 and Cys455 are found to be unpaired, containing free sulfhydryl groups. The pattern of disulfide bridges is in agreement with that predicted from the X-ray crystallographic structure of canine myeloperoxidase (MPO) (Zeng, J., and Fenna, R. E. (1992) J. Mol. Biol. 226, 185-207) to be general for the class of mammalian peroxidases, including EPO, MPO, lactoperoxidase (LPO), and thyroid peroxidase (TPO). Of four candidate sites in EPO for attachment of glucosamine-based carbohydrate, Asn327 and Asn363 are occupied, whereas Asn700 and Asn708 are unsubstituted. Furthermore, a discrepancy in the literature regarding the sequence of residues 645-659 is resolved. PMID- 10864453 TI - Cytosolic phospholipase A(2) activity associated with nuclei is not inhibited by arachidonyl trifluoromethyl ketone in macrophages stimulated with receptor recognized forms of alpha(2)-macroglobulin. AB - We have studied the translocation of cytosolic phospholipase A(2) (cPLA(2)) to nuclei in macrophages stimulated with receptor-recognized forms of alpha(2) macroglobulin (alpha(2)M*). Translocation of phosphorylated cPLA(2) to nuclei was determined by immunoprecipitation of cPLA(2) in (32)P(i)-labeled cells. The identity of cPLA(2) was established by comparing its mobility on gels with an authentic cPLA(2) standard. cPLA(2) activity was quantified by measuring the release of [(14)C]arachidonic acid from the substrate 1-palmitoyl-2-[1 (14)C]arachidonyl-sn-glycerophosphatidylcholine. alpha(2)M* caused a two- to threefold increase in cPLA(2) phosphorylation and its translocation to nuclei. The p38 MAPK inhibitor SB203580, PKC inhibitor chelerythrin, or depletion of intracellular Ca(2+) profoundly decreased cPLA(2) activity in nuclei isolated from agonist-stimulated cells. The requirement for Ca(2+), PKC, and p38 MAPK activation appears to be of major importance for nuclear cPLA(2) activity. In contrast to cellular cPLA(2) activity, nuclear cPLA(2) activity was not inhibited by arachidonyl trifluoromethyl ketone (AACOCF(3)) in agonist-stimulated cells. It is concluded that the association of cPLA(2) with nuclear membranes in agonist stimulated cells modifies the activity and the sensitivity of the enzyme to inhibition by AACOCF(3) in this phospholipid environment. PMID- 10864454 TI - Increasing expression of P450 and P450-reductase proteins from monocots in heterologous systems. AB - Monocotyledonous crop plants are usually more resistant to herbicides than grass weeds and most dicots. Their resistance to herbicides is mediated in many cases by P450 oxygenases. Monocots thus constitute an appealing source of P450 enzymes for manipulating herbicide resistance and recombinant forms of the major xenobiotic metabolizing mooxygenases are potential tools for the optimization of new active molecules. We report here the isolation and functional characterization of the first P450 and P450 reductase coding sequences from wheat. The first attempts at expressing these cDNAs in yeast and tobacco led to levels of protein, which were extremely low, often not even detectable. The wheat P450 cDNAs were efficiently transcribed, but no protein or activity was found. Wheat coding sequences, like those of other monocots, are characterized by a high GC content and by a related strong bias of codon usage, different from that observed in yeast or dicots. Complete recoding of genes being costly, the reengineering their 5'-end using a single PCR megaprimer designed to comply with codon usage of the host was attempted. It was sufficient to relieve translation inhibition and to obtain good levels of protein expression. The same strategy also resulted in a dramatic increase in protein expression in tobacco. A basis for the success of such a partial recoding strategy, much easier and cheaper than complete recoding of the cDNA, is proposed. PMID- 10864455 TI - Emergence of fluconazole-resistant sterol 14-demethylase P450 (CYP51) in Candida albicans is a model demonstrating the diversification mechanism of P450. PMID- 10864456 TI - Integrins as mediators of morphogenesis in Drosophila. PMID- 10864457 TI - Real-time measurements of the interactions between fluorescent speract and its sperm receptor. AB - Lytechinus pictus sea urchin sperm express receptors for speract, a sperm activating peptide derived from the homologous egg jelly coat. We found that the fluorescence of fluorophore-labeled, active, speract analogs is quenched upon receptor binding. This property allowed us to perform real-time measurements of speract-receptor interactions using intact sperm and to determine, for the first time, their association (k(on)) and dissociation (k(off)) rate constants. The high k(on) (2.4 x 10(7) M(-1 )s(-1)) and low k(off) (4.4 x 10(-6) s(-1) (95%) and 3.7 x 10(-4) s(-1) (5%)) can account for the sperm response to picomolar concentrations of speract. We also examined the influence of extracellular ions on speract-receptor interactions using the fluorescence quenching method described in this study. The association rate of speract to the receptor is dramatically reduced in Na(+)-free seawater (NaFSW), divalent cation-free seawater (DCFSW), and high-K(+) seawater (HKSW). In seawater speract induces an increase in intracellular pH (pHi), while it is unable to do so in either NaFSW or HKSW. To test if the lack of this pHi change causes the reduction in the speract association rate, pHi was increased with NH(4)Cl (10 mM) at the time labeled speract was added. Interestingly, this procedure completely (in HKSW) or partially (in NaFSW and DCFSW) restored the speract association rate to its receptor. These findings indicate that an increase in sperm pHi positively affects the receptor binding activity for this peptide and may partially explain the positive binding cooperativity displayed by the speract receptor. PMID- 10864458 TI - Elongation of axolotl tailbud embryos requires GPI-linked proteins and organizer induced, active, ventral trunk endoderm cell rearrangements. AB - Application of phosphatidylinositol-specific phospholipase C to early tailbud stage axolotl embryos reveals that a specific subset of morphogenetic movements requires glycosylphosphatidylinositol (GPI)-linked cell-surface proteins. These include pronephric duct extension, "gill bulge" formation, and embryonic elongation along the anteroposterior axis. The work of Kitchin (1949, J. Exp. Zool. 112, 393-416) led to the conclusion that extension of the notochord provided the motive force driving anteroposterior stretching in axolotl embryos, elongation of other tissues being a passive response. We therefore conjectured that axial mesoderm cells might display the GPI-linked proteins required for elongation of the embryo. However, we show here that removal of most of the neural plate and axial and paraxial mesoderm prior to neural tube closure does not prevent elongation of ventrolateral tissues. Tissue-extirpation and tissue marking experiments indicate that elongation of the ventral trunk occurs via active, directed tissue rearrangements within the endoderm, directed by signals emanating from the blastopore region. Extension of both dorsal and ventral tissues requires GPI-linked proteins. We conclude that elongation of axolotl embryos requires active cell rearrangements within ventral as well as axial tissues. The fact that both types of elongation are prevented by removal of GPI linked proteins implies that they share a common molecular mechanism. PMID- 10864459 TI - Kidney development in cadherin-6 mutants: delayed mesenchyme-to-epithelial conversion and loss of nephrons. AB - During nephrogenesis, dynamic changes in the expression of cell adhesion molecules are evident as epithelial structures differentiate from the induced mesenchyme. The cadherins are thought to play an important role in the metanephric mesenchyme, when cells aggregate to form the renal vesicle, a polarized epithelial structure which eventually fuses with the ureteric bud to generate a continuous nascent nephron. We have generated and analyzed mice with a targeted mutation in the gene encoding cadherin-6 (Cad-6), a type II cadherin expressed during early stages of nephrogenesis. These mice are viable and fertile, and they complete both early and late aspects of nephrogenesis. However, upon closer examination in vitro and in vivo, a fraction of the induced metanephric mesenchyme in Cad-6 mutant kidneys fails to form a fully polarized epithelium on schedule. Moreover, a significant number of the renal vesicles in Cad-6 mutant kidneys apparently fail to fuse to the ureteric bud. These alterations in epithelialization and fusion apparently lead to a loss of nephrons in the adult. These studies support the idea that cadherins play an essential role in the formation of epithelial structures and underscore the importance of timing in orchestrating the morphogenesis of complex epithelial tissues. PMID- 10864460 TI - SNAREs in mammalian sperm: possible implications for fertilization. AB - Soluble N-ethylmalameide-sensitive factor attachment protein receptor (SNARE) proteins are present in mammalian sperm and could be involved in critical membrane fusion events during fertilization, namely the acrosome reaction. Vesicle-associated membrane protein/synaptobrevin, a SNARE on the membrane of a vesicular carrier, and syntaxin 1, a SNARE on the target membrane, as well as the calcium sensor synaptotagmin I, are present in the acrosome of mammalian sperm (human, rhesus monkey, bull, hamster, mouse). Sperm SNAREs are sloughed off during the acrosome reaction, paralleling the release of sperm membrane vesicles and acrosomal contents, and SNARE antibodies inhibit both the acrosome reaction and fertilization, without inhibiting sperm-egg binding. In addition, sperm SNAREs may be responsible, together with other sperm components, for the asynchronous male DNA decondensation that occurs following intracytoplasmic sperm injection, an assisted reproduction technique that bypasses normal sperm-egg surface interactions. The results suggest the participation of sperm SNAREs during membrane fusion events at fertilization in mammals. PMID- 10864461 TI - SMA-3 smad has specific and critical functions in DBL-1/SMA-6 TGFbeta-related signaling. AB - A TGFbeta signal transduction cascade controls body size and male tail morphogenesis in the nematode Caenorhabditis elegans. We have analyzed the function of the sma-3 Smad gene, one of three Smad genes that function in this pathway. Null mutations in sma-3 are at least as severe as null mutations in the ligand and type I receptor genes, dbl-1 and sma-6, indicating that the other Smads do not function in the absence of SMA-3. Furthermore, null mutations in sma 3 do not cause defects in egg laying or in regulation of the developmentally arrested dauer larva stage, indicating no overlapping function with another C. elegans TGFbeta signaling pathway. The sma-3 gene is widely expressed at all developmental stages in hermaphrodites and males. The molecular lesions associated with eight sma-3 alleles of varying severity have been determined. The missense mutations cluster in two previously identified regions important for Smad function. PMID- 10864462 TI - Lim1 activity is required for intermediate mesoderm differentiation in the mouse embryo. AB - During gastrulation and early organogenesis, Lim1 is expressed in the visceral endoderm, the anterior mesendoderm, and the lateral mesoderm that comprises the lateral plate and intermediate mesoderm. A previous study has reported that kidneys and gonads are missing in the Lim1 null mutants (W. Shawlot and R. R. Behringer, 1995, Nature 374, 425-430). Results of the present study show that in the early organogenesis stage mutant embryo, the intermediate mesoderm that contains the urogenital precursor tissues is disorganized and displays diminished expression of PAX2 and the Hoxb6-lacZ transgene. When posterior epiblast cells of the Lim1 null mutant embryo were transplanted to the primitive streak of wild type host embryos, they were able to colonize the lateral plate and intermediate mesoderm of the host, suggesting that Lim1 activity is not essential for the allocation of epiblast cells to these mesodermal lineages. However, most of the mutant cells that colonized the lateral and intermediate mesoderm of the host embryo did not express the Hoxb6-lacZ transgene, except for some cells that were derived from the distal part of the posterior epiblast. Lim1 activity may therefore be required for the full expression of this transgene that normally marks the differentiation of the lateral plate and intermediate mesoderm. PMID- 10864463 TI - Slug is an essential target of TGFbeta2 signaling in the developing chicken heart. AB - An epithelial-mesenchymal cell transformation (EMT) occurs during the development of endocardial cushions in the atrioventricular (AV) canal of the heart. This is a complex developmental process regulated by multiple extracellular signals and signal transduction pathways. It was recently shown that the transcription factor Slug is expressed in the AV canal and is required for initial steps of EMT. Treatment of AV canal explants with either antisense oligodeoxynucleotides toward Slug or anti-TGFbeta2 antibody inhibited initial steps of EMT. Others have identified roles for HGF and BMP during EMT in the heart. Both HGF and BMP are known to regulate Slug in other cell types. To determine whether TGFbeta2 or other signaling factors regulate Slug expression during EMT in the heart, we cultured AV canal explants in the presence of anti-TGFbeta2 antibody, anti TGFbeta3 antibody, pertussis toxin, retinoic acid, noggin, or anti-HGF antibody. Only treatment with anti-TGFbeta2 antibody or retinoic acid inhibited Slug expression in AV canal explants. Consistent with these data, we found that retinoic acid disrupted initial steps of EMT, while antagonists of BMP and HGF signaling disrupted later steps of EMT. Transfection of AV canal explants with Slug rescued the inhibitory effect of anti-TGFbeta2 antibody but not retinoic acid on EMT. Slug is thus an essential target of TGFbeta2 signaling during EMT in the developing chicken heart. PMID- 10864464 TI - Role for cGATA-5 in transcriptional regulation of the embryonic chicken pepsinogen gene by epithelial-mesenchymal interactions in the developing chicken stomach. AB - A gene encoding embryonic chicken pepsinogen (ECPg), a zymogen of the digestive enzyme pepsin, is expressed specifically in epithelial cells of glands of embryonic stage proventriculus (glandular stomach) under the influence of mesenchyme. We found four GATA and one Sox binding motifs in 1.1 kb of the 5' flanking region of the ECPg gene which are essential to the organ-specific expression of the gene. The expression of cGATA-5 and cSox2 in the proventriculus from day 6 to day 12 of incubation was therefore analyzed. cGATA-5 was more strongly expressed in glandular epithelial cells than in luminal epithelial cells, while cSox2 gene expression was weaker in glandular epithelial cells. Using heterologous recombination explants we also discovered that the expression of cGATA-5 and cSox2 in epithelial cells was affected by mesenchyme when the latter induced ECPg gene expression in epithelial cells. Introduction of expression constructs into epithelial cells by electroporation demonstrated that cGATA-5 upregulated transcription of a reporter luciferase gene via a cis element in the 5' flanking region of the ECPg gene. The gel mobility shift assay revealed that the cGATA-5 protein specifically binds to the GATA binding sites. cSox2 downregulated the activity of luciferase but it was not through the Sox binding motif. These results suggest that cGATA-5 positively regulates transcription of the ECPg gene and is involved in spatial regulation of the pepsinogen gene during development. PMID- 10864465 TI - Developmental changes in the spatial expression of genes involved in myosin function in Dictyostelium. AB - We analyzed the spatial expression patterns of the genes involved in myosin function by in situ hybridization at the tipped aggregate and early culmination stages of Dictyostelium. Myosin heavy chain II mRNA was enriched in the anterior prestalk region of the tipped aggregates, whereas it disappeared from there and began to appear in both upper and lower cups of the early culminants. Similarly, mRNAs for essential light chain, regulatory light chain, myosin light chain kinase A, and myosin heavy chain kinase C were enriched in the prestalk region of the tipped aggregates. However, expression of these genes was distinctively regulated in the early culminants. These findings suggest the existence of mechanisms responsible for the expression of particular genes. PMID- 10864466 TI - Is chordin a long-range- or short-range-acting factor? Roles for BMP1-related metalloproteases in chordin and BMP4 autofeedback loop regulation. AB - Diffusible morphogen models have been used widely to explain regional specification of tissues and body axes during animal development. The three signal model for patterning the dorsal-ventral axis of the amphibian embryo proposes, in part, that a factor(s) secreted from Spemann's organizer is responsible for converting lateral marginal zone into more dorsal cell fates. We examine the possibility that chordin, a secreted inhibitor of bone morphogenetic protein (BMP) signaling and candidate "dorsalizing signal," is a long-range acting factor. We show that chordin can, when overexpressed, act directly over distances of at least 450 microm in the early Xenopus embryo to create a gradient of BMP signaling. However, since lower levels of chordin can still induce secondary axes and these amounts of chordin act only locally to inhibit a BMP target gene, we suggest that chordin likely acts as a short-range signal in vivo. Furthermore, BMP1, a secreted metalloprotease that cleaves chordin protein in vitro, inhibits chordin's axis-inducing effects, suggesting that BMP1 functions to negatively regulate chordin's action in vivo. A dominant-negative mutant BMP1 blocks the in vitro cleavage of chordin protein by wild-type BMP1 and induces secondary axes when injected ventrally. We argue that BMP1 and Xolloid are probably functionally redundant metalloproteases and may have two roles in the early Xenopus embryo. One role may be to inhibit the action of low-level chordin protein expressed throughout the entire embryo and a possible second role may be to inhibit activation of a juxtacrine cell relay, thereby confining chordin's action to the organizer region preventing chordin from functioning as a long range-acting factor. PMID- 10864467 TI - Early development of mouse embryos null mutant for the cyclin A2 gene occurs in the absence of maternally derived cyclin A2 gene products. AB - Progression through the mammalian cell cycle is regulated by the sequential activation and inactivation of the cyclin-dependent kinases. In adult cells, cyclin A2-dependent kinases are required for entry into S and M phases, completion of S phase, and centrosome duplication. However, mouse embryos lacking the cyclin A2 gene nonetheless complete preimplantation development, but die soon after implantation. In this report, we investigated whether a contribution of maternal cyclin A2 mRNA and protein to early embryonic cell cycles might explain these conflicting observations. Our data show that a maternal stock of cyclin A2 mRNA is present in the oocyte and persists after fertilization until the second mitotic cell cycle, when it is degraded to undetectable levels coincident with transcriptional activation of the zygotic genome. A portion of maternally derived cyclin A2 protein is stable during the first mitosis and persists in the cytoplasm, but is completely degraded at the second mitosis. The ability of cyclin A2-null mutants to develop normally from the four-cell to the postimplantation stage in the absence of detectable cyclin A2 gene product indicates therefore that cyclin A2 is dispensable for cellular progression during the preimplantation nongrowth period of mouse embryo development. PMID- 10864468 TI - Distribution of dorsal-forming activity in precleavage embryos of the Japanese newt, Cynops pyrrhogaster: effects of deletion of vegetal cytoplasm, UV irradiation, and lithium treatment. AB - Two types of axis-deficient embryos developed after deletion of the vegetal cytoplasm: wasp-shaped embryos and permanent-blastula-type embryos. In situ hybridization revealed that neither type of axis-deficient embryo expressed goosecoid or pax-6. brachyury was expressed in the constricted waist region of the wasp-shaped embryos but was not expressed in the permanent-blastula-type embryos. Further, we examined the effect of UV irradiation on Japanese newt embryos. Surprisingly, UV-irradiated Japanese newt eggs formed hyperdorsalized embryos. These embryos gastrulated in an irregular circular fashion with goosecoid expression in the circular equatorial region. At tailbud stage, these embryos formed a proboscis which is very reminiscent of that formed in hyperdorsalized Xenopus embryos. Transplantation of the marginal region of the UV irradiated embryos revealed that the entire marginal zone had organizer activity. Thus we conclude that UV hyperdorsalizes Japanese newt embryos. Finally, lithium treatment of normal embryos at the 32-cell stage also resulted in hyperdorsalization. Lithium treatment of vegetally deleted embryos had two distinct results. Lithium treatment of permanent-blastula-type embryos did not result in the formation of dorsal axial structures, while the same treatment reinduced gastrulation and dorsal axis formation in the wasp-shaped embryos. Based on these results, we propose a model for early axis specification in Japanese newt embryos. The model presented here is fundamentally identical to the Xenopus model, with some important modifications. The vegetally located determinants required for dorsal development (dorsal determinants, DDs) are distributed over a wider region at fertilization in Japanese newt embryos than in Xenopus embryos. The marginal region of the Japanese newt embryo at the beginning of development overlaps with the field of the DDs. Gastrulation is very likely to be a dorsal marginal-specific property, while self-constriction is most probably a ventral marginal-specific property in Japanese newt embryos. PMID- 10864469 TI - Ventricular expression of tbx5 inhibits normal heart chamber development. AB - The T-box gene tbx5 is expressed in the developing heart, forelimb, eye, and liver in vertebrate embryos during critical stages of morphogenesis and patterning. In humans, mutations in the TBX5 gene have been associated with Holt Oram syndrome, which is characterized by developmental anomalies in the heart and forelimbs. In chicken and mouse embryos, tbx5 expression is initiated at the earliest stages of heart formation throughout the heart primordia and is colocalized with other cardiac transcription factors such as nkx-2.5 and GATA4. As the heart differentiates, tbx5 expression is restricted to the posterior sinoatrial segments of the heart, consistent with the timing of atrial chamber determination. The correlation between tbx5 expression and atrial lineage determination was examined in retinoic acid (RA)-treated chicken embryos. tbx5 expression is maintained throughout the hearts of RA-treated embryos under conditions that also expand atrial-specific gene expression. The downstream effects of persistent tbx5 expression in the ventricles were examined directly in transgenic mice. Embryos that express tbx5 driven by a beta-myosin heavy chain promoter throughout the primitive heart tube were generated. Loss of ventricular specific gene expression and retardation of ventricular chamber morphogenesis were observed in these embryos. These studies provide direct evidence for an essential role for tbx5 in early heart morphogenesis and chamber-specific gene expression. PMID- 10864472 TI - Induction of DNA replication in the germinal vesicle of the growing mouse oocyte. AB - Growing mouse oocytes are physiologically arrested in the G2 phase of prophase of the first meiotic division. Growing oocytes were isolated from ovaries of 9- to 12-day-old mice and fused with parthenogenetic one-cell eggs or two-cell embryos derived from fertilized eggs. Resulting hybrids were injected with Dig-11-dUTP and examined for DNA replication using immunofluorescence. Parthenogenetic one cell eggs fused at telophase II, G1, and middle-to-late S phase, and also S-phase two-cell blastomeres, were able to trigger DNA synthesis in oocyte germinal vesicle (GV) in the majority of hybrids cultured to the end of the first cell cycle. Activation of replication in the GV occurred within 2-3 h after fusion of growing oocytes with S-phase eggs. We show indirectly that the reactivation of replication in GVs was not dependent on the breakdown of the GV envelope. Although GVs had the ability to renew DNA replication after fusion, the G2 blastomere nuclei were incapable of reinitiating DNA replication under the influence of S-phase one-cell eggs. We hypothesize that the nuclei of growing oocytes arrested in meiotic prophase are in a physiological state that is equivalent to replication-competent G1, and not G2, nuclei. PMID- 10864470 TI - Gpbox (Psx2), a homeobox gene preferentially expressed in female germ cells at the onset of sexual dimorphism in mice. AB - XX gonads differentiate into ovaries, a morphologic event evident by embryonic day 13.5 (E13.5) in mice. To identify early markers of oogenesis, sex-specific urogenital ridge cDNA libraries were constructed from E12-13 embryos. After mass excision and isolation of plasmid DNA, approximately 4800 expressed sequence tags were determined and compared to existing databases. Few cDNAs were specifically expressed in the urogenital ridge, but one, designated GPBOX, encodes a 227-amino acid homeobox protein that is first expressed at E10.5 in the embryo as well as in the extraembryonic tissues. The Gpbox gene is single copy in the mouse genome and is located on the X chromosome in close proximity to two other homeobox genes, Pem and Psx1. Within the embryo, its expression is limited to the gonad, and transcripts are not detected in adult tissues. Although comparable levels are initially present in both sexes, GPBOX transcripts accumulate faster in female germ cells and peak at E12.5 when they are present in fivefold greater abundance than in males. The persistence of GPBOX transcripts in female germ cells until E15.5 and their virtual disappearance in males by E13.5 suggest that Gpbox may play a role in mammalian oogenesis. PMID- 10864471 TI - Localization of a novel human A-kinase-anchoring protein, hAKAP220, during spermatogenesis. AB - Using a combination of protein kinase A type II overlay screening, rapid amplification of cDNA ends, and database searches, a contig of 9923 bp was assembled and characterized in which the open reading frame encoded a 1901-amino acid A-kinase-anchoring protein (AKAP) with an apparent SDS-PAGE mobility of 220 kDa, named human AKAP220 (hAKAP220). The hAKAP220 amino acid sequence revealed high similarity to rat AKAP220 in the 1167 C-terminal residues, but contained 727 residues in the N-terminus not present in the reported rat AKAP220 sequence. The hAKAP220 mRNA was expressed at high levels in human testis and in isolated human pachytene spermatocytes and round spermatids. The hAKAP220 protein was present in human male germ cells and mature sperm. Immunofluorescent labeling with specific antibodies indicated that hAKAP220 was localized in the cytoplasm of premeiotic pachytene spermatocytes and in the centrosome of developing postmeiotic germ cells, while a midpiece/centrosome localization was found in elongating spermatocytes and mature sperm. The hAKAP220 protein together with a fraction of PKA types I and II and protein phosphatase I was resistant to detergent extraction of sperm tails, suggesting an association with cytoskeletal structures. In contrast, S-AKAP84/D-AKAP1, which is also present in the midpiece, was extracted under the same conditions. Anti-hAKAP220 antisera coimmunoprecipitated both type I and type II regulatory subunits of PKA in human testis lysates, indicating that hAKAP220 interacts with both classes of R subunits, either through separate or through a common binding motif(s). PMID- 10864473 TI - Indicators of oxidative injury and alterations of the cell membrane in the skeletal muscle of rats submitted to ischemia and reperfusion. AB - BACKGROUND: Oxygen free radicals are considered to be important components involved in the physiopathological tissue alterations observed during ischemia and reperfusion. The objective of the present study was to investigate oxidative stress based on indicators of oxygen free radical activity and on the changes in behavior of the lipoprotein membrane (O-phosphoserine) in the skeletal muscle of rats. MATERIAL AND METHODS: Twenty Wistar rats were divided into two groups of 10. One group was submitted to 3 h of total ischemia by applying a tourniquet to the hind limb and the contralateral hind limb was used as control. The second group was submitted to the same procedure and was reperfused for 45 min after 3 h of ischemia by removing the tourniquet, where the contralateral hind limb of the same animal was used as control. Muscle biopsies were taken after ischemia and reperfusion and the parameters indicating oxidative stress (reduced and oxidized glutathione, malondialdehyde, glutamine synthetase, protein carbonyl) and O phosphoserine (OPS) alterations were analyzed. RESULTS: The following results display control versus experimental hindlimbs groups obtained from the same animal. The skeletal muscle of rats submitted to total ischemia of 3 h duration showed increased OPS release (2.69 +/- 4.52 vs 8.03 +/- 7.20; n = 10; P = 0.024) and no change in reduced and oxidized glutathione, glutamine synthetase, protein carbonyl, or malondialdehyde. After 45 min of reperfusion there was an increase in oxidized glutathione levels (0.30 +/- 0.06 vs 0.39 +/- 0.09; n = 8; P = 0.02) and malondialdehyde levels (154. 78 +/- 26.13 vs 206.30 +/- 47.30; n = 9; P = 0.008), a fall in glutamine synthetase (21.80 +/- 3.61 vs 13.52 +/- 6.78; n = 9; P = 0. 004), and a return of OPS to levels close to the initial ones. No changes in reduced glutathione or protein carbonyl were observed in the two groups studied. CONCLUSIONS: After a total ischemia duration of 3 h there were signs of damage to the phospholipid membrane of the rat skeletal muscle, as demonstrated by the elevation of OPS and the few or no oxidative changes in the cell. After 45 min of reperfusion, oxidative damage to the lipoprotein components of the cell membrane was observed, characterized by elevations of oxidized glutathione and malondialdehyde levels and a fall in glutamine synthetase levels. PMID- 10864474 TI - Role of caveolin in hemodynamic force-mediated endothelial changes. AB - BACKGROUND: Caveolin has been shown to play an important role in signal transduction and nitric oxide synthase production. The purpose of this study was to investigate whether caveolin was tyrosine phosphorylated or activated by shear stress or cyclic strain in bovine aortic endothelial cells (BAECs). MATERIALS AND METHODS: BAECs were subjected to an average of 10% strain at a rate of 60 cycles/min or a laminar shear stress of 10 dyn/cm(2) for up to 4 h. Immunoblotting with anticaveolin antibody was performed to assess activation of caveolin. Coimmunoprecipitation of anticaveolin antibody with anti-tyrosine phosphorylation antibody was performed to detect the tyrosine phosphorylation of caveolin. RESULTS: Neither cyclic strain nor shear stress at physiologic levels altered the level of caveolin protein. Tyrosine phosphorylation of caveolin could not be observed at any time under either cyclic strain or shear stress condition. CONCLUSION: Although hemodynamic forces alter nitric oxide synthase production and activate signal transduction, caveolin levels or activity is not altered in endothelial cells exposed to shear stress or cyclic strain. PMID- 10864475 TI - Initiating the inflammatory phase of incisional healing prior to tissue injury. AB - BACKGROUND: The time required for incisional healing accounts for the majority of postoperative pain and convalescence. Impaired healing prolongs the process further. If a method for accelerating acute incisional wound healing could be developed, patients would benefit from decreased wound failure and an earlier return to their premorbid condition. MATERIALS AND METHODS: In a rat dermal model, cytokine or vehicle infiltration prior to incision was performed using a single dose or four daily doses preincision. Planned incision sites were primed with the proinflammatory cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) or platelet-derived growth factor BB (PDGF-BB) in an effort to activate the inflammatory phase of healing prior to wounding. At the time of incision closure, one half of the incisions were treated with transforming growth factor beta(2) (TGF-beta(2)). Incisional sites were biopsied and stained with hematoxylin and eosin and immunohistochemistry for inflammatory cells and fibroblast populations and breaking strength was measured. RESULTS: Priming skin with GM-CSF or PDGF-BB mimicked the early inflammatory phase of wound healing. Macrophage staining (EB1) and fibroblast staining (vimentin) were significantly increased prior to incision. Inflammatory priming as well as priming coupled with TGF-beta(2) at the time of the incision closure synergistically improved breaking strength. CONCLUSION: This study demonstrates that sequential therapy consisting of priming of tissue with an inflammatory cytokine followed by application of a proliferative cytokine at the time of incision closure nearly doubles the breaking strength of an acute wound. By manipulating the inflammatory and early proliferative phases of wound healing with tissue growth factors, it may be possible to accelerate acute wound repair and shift the wound healing trajectory to the left. PMID- 10864476 TI - Interaction of angiogenesis inhibitor TNP-470 with basic fibroblast growth factor receptors. AB - TNP-470 is a synthetic analogue of fumagillin that acts as a potent angiogenesis inhibitor. Recently, our laboratory demonstrated that systemic administration of TNP-470 (5.0 mg/kg) decreased the rate of cutaneous wound healing by greater than 20%. In this study, we tested the hypothesis that TNP-470 interferes with the wound repair-stimulating action of basic fibroblast growth factor (bFGF) by competing with endogenous bFGF for its binding sites on the receptor protein. The influence of TNP-470 was examined in vitro in a ligand competition assay of high- and low-affinity receptor binding to (125)I-bFGF in NIH/3T3 cells. Results demonstrated that recognition of (125)I-bFGF by low-affinity growth factor binding sites was significantly decreased (P < 0.01) in the presence of TNP-470. However, TNP-470 inhibition of radiolabeled bFGF binding to high-affinity sites was not significantly affected (P = 0.07). In view of recent studies demonstrating that the low-affinity receptors of bFGF were heparan sulfate proteoglycans, we suggest that the influence of TNP-470 on diminished wound healing is due to its direct recognition by these molecules. PMID- 10864477 TI - Lysophosphatidic acid stimulates intestinal restitution via cytoskeletal activation and remodeling. AB - BACKGROUND: The gastrointestinal tract heals superficial wounds by a process of epithelial migration termed restitution. Restitution is an important response for preventing conditions like stress gastritis, ulcer disease, celiac sprue, ischemia-reperfusion injury, bacterial translocation during shock, and inflammatory bowel disease. The purpose of this study was to determine the effect of a platelet product, lysophosphatidic acid (LPA), on intestinal restitution. MATERIALS AND METHODS: IEC-6 cells were used to study the effect of LPA on intracellular calcium mobilization, actin stress fiber formation, and actin and FAK protein levels. An in vitro model of restitution was utilized to determine the LPA-stimulated IEC-6 migration. RESULTS: LPA administration stimulated intracellular calcium mobilization in a dose-dependent fashion with typical peak and plateau phases suggestive of a receptor-mediated response. Actin stress fiber formation occurred within 15 min after LPA treatment and was especially apparent at 2 h. This response was accompanied by higher actin and FAK protein levels. LPA also stimulated IEC-6 migration 3-fold within 24 h. All of these effects were completely inhibited by pertussis toxin. CONCLUSIONS: Exposure of IEC-6 cells to LPA results in significant calcium mobilization and cytoskeletal remodeling within minutes. This activity is accompanied by increased actin and FAK levels. Cellular migration is significantly enhanced by LPA. These responses seem to be due to pertussis-sensitive G-protein-associated receptors. The ability of LPA to potentiate intestinal cell restitution appears, in part, to be mediated by effects on intestinal cell cytoskeletal structure. PMID- 10864478 TI - Vascular clips have no significant effect on the cellular proliferation, intimal changes, or peak systolic velocity at anastomoses in rabbit vein grafts. AB - OBJECTIVE: This study compares vascular closure staples (VCSs) with conventional sutures in the rabbit carotid vein graft model to determine whether anastomotic technique affects cellular proliferation, blood velocity, or intimal changes when measured over a period of 3 months postoperatively. METHODS: Twenty-six New Zealand White rabbits weighing 3.0-3.2 kg underwent interposition of jugular vein grafts in left carotid arteries. Half of the animals had anastomoses performed with small VCSs (n = 13) and half had anastomoses performed with 8-O interrupted polypropylene suture. Animals were allowed to survive for 1 week (n = 4, VCS; n = 4, suture), 2 weeks (n = 4, VCS; n = 4, suture), and 3 months (n = 5, VCS; n = 5, suture). The peak systolic velocity (PSV) at the distal anastomosis was measured after completion of the graft and again at sacrifice in the 3-month survival groups. At sacrifice, sections were taken from the middle and distal end of the vein graft and the distal carotid artery. Vascular cell proliferation was measured using 5-bromo-2'-deoxyuridine labeling and intimal changes were measured using digitized microscopic images. RESULTS: All 26 grafts were open at the time of sacrifice. PSV at the distal clipped anastomosis was 40.52 cm/s (t = 0) and 34.3 cm/s (t = 3 months, P = 0.31). PSV at the distal sutured anastomosis was 38.30 cm/s (t = 0) and 39.23 cm/s (t = 3 months, P = 0.82). There was no difference between the two techniques at either t = 0 or t = 3 months (P = 0.51 and P = 0.31, respectively). Endothelial cell proliferation and smooth muscle cell proliferation at the anastomosis was highest during the 2 weeks after the procedure, then returned to baseline levels by 3 months. But there was no significant difference between the clipped and sutured groups with respect to vascular cell proliferation postoperatively. The intimal thickness changed significantly in the vein graft at the anastomosis for both the clipped and sutured groups (P = 0.0007 and P = 0.002). But there was no difference when the intimal changes for each technique were compared (P = 0.94). CONCLUSION: No differences were observed when peak systolic velocity, vascular cell proliferation, and intimal changes were compared between sutured and stapled anastomoses in rabbit vein interposition grafts over a period of 3 months after surgery. PMID- 10864479 TI - Sulfatide elongates dorsal skin flap survival in rats. AB - Monoclonal antibodies to adhesive molecules have been used in many trials to decrease ischemia-reperfusion injury, which is considered to occur in areas such as the distal region of the random pattern flap. The monoclonal antibody to the primary neutrophil adherence-mediating glycoprotein CD18 improves the survival length of the random pattern flap. Sulfatide binds strongly with L- and P selectin. We found that sulfatide has a protective effect against ischemia reperfusion injury. The purpose of this study was to evaluate the effect of sulfatide on the survival length of the random pattern flap in rats. Sulfatide was administered intravenously just before elevation of the cranially based dorsal skin flap. Administration of sulfatide significantly augmented flap survival length (49.5 +/- 1.7 mm vs control 41.5 +/- 2.1 mm, P = 0. 01). Flap survival length was significantly longer than dye distance (49.1 +/- 2.0 mm vs 39.7 +/- 1.1 mm, P = 0.01). In the control flap, no significant difference between survival length and dye distance was detected. Histological examination 48 h after flap elevation showed leukocyte invasion in the dermal layer of control flaps, whereas little leukocyte invasion was observed in the flaps of rats administered sulfatide. PMID- 10864480 TI - Association between the expression of mast cell chymase and intraperitoneal adhesion formation in mice. AB - BACKGROUND: Adhesion formation is a major source of postoperative morbidity and mortality. Mast cells and their major protease, chymase, have been shown to participate in the healing process as well as in tissue remodeling. We aimed to identify the role of mast cells in intraperitoneal adhesion formation and to assess whether there is an association between the expression of mast cell chymase and adhesion formation. MATERIALS AND METHODS: Both mast cell-deficient W/W(V) mice and congenic +/+ mice received a standardized lesion produced by cecal scraping and the application of 95% ethanol. Adhesions were assessed blindly 1 week later using a standardized scale. In addition, histamine content, mast cell numbers, and chymase activity in cecum as well as at the healing sites were evaluated before and 7 days after surgical injury. RESULTS: A significant reduction in adhesion formation was seen in mast cell-deficient W/W(V) mice (P < 0.05). In the normal cecum, histamine content did not significantly differ between W/W(V) and +/+ mice. Chymase activity in cecum was detected in control +/+ mice, but not in W/W(V) mice. Mast cell numbers and chymase activity levels at the healing sites of +/+ mice were significantly increased 7 days after surgery. CONCLUSIONS: Our results indicate that mast cells contribute to intraperitoneal adhesion formation in mice, and suggest that chymase originating from mast cells is important in the development of adhesions. PMID- 10864481 TI - Matrix glycosaminoglycans in the growth phase of fibroblasts: more of the story in wound healing. AB - BACKGROUND: Understanding wound healing and ways to accelerate the healing process includes understanding the factors that influence the synthesis of granulation tissue, which fills the wound before epithelialization. An important phase of early wound healing involves secretion of glycosaminoglycans (GAGs) by fibroblasts which form a hydrophilic matrix suitable for remodeling during healing. The complexity of GAG structure and function in the extracellular matrix (ECM) remains poorly studied in wound healing. There is no established model for cutaneous wound healing due to variations in donor age, anatomic site, or stage of organ development. Rat embryo fibroblasts (REF) developed as a model to study malignant changes in fibroblasts were used as a model for fibroblasts in early wound healing because they lack the confounding variations based on age, site, and stage present in other fibroblasts used to study early wound healing. The purpose of this study was to identify and characterize the sulfated GAGs synthesized by REF-D. MATERIALS AND METHODS: Rat embryo fibroblasts (REF-D) were cultured in serum-based medium and radiolabeled during their growth phase with (35)S to identify the GAG chains usually associated with proteoglycans (PGs). The sites of attachment (ECM-rich) were collected with detergent in sodium acetate buffer, pH 5.8, in the presence of protease inhibitors. Sulfated molecules were collected by ion-exchange chromatography and then assayed for GAGs. Nitrous acid deamination was used to determine heparan sulfate GAGs, and chondroitinase was used for chondroitin/dermatan sulfate GAGs. The proportion of individual GAGs was expressed with respect to sulfated molecules isolated. In addition, RNA was isolated from subconfluent REF-D, and core proteins for proteoglycans (decorin, biglycan, syndecan-2, and perlecan) were assayed by reverse transcription polymerase chain reaction. RESULTS: There were two major configurations of GAGs: free GAG chains (79.7% of sulfated molecules) and GAGs attached to the core protein of a proteoglycan (15.6%). The free GAG chains were composed of chondroitin sulfate (79.1% +/- 3.5) and heparan sulfate (28.7% +/- 2.1). In the smaller group of PGs, both heparan sulfate (94.8% +/- 7.3) and chondroitin sulfate (88.9% +/- 3.2) chains were attached to a core protein. REF-D expressed mRNA for biglycan and decorin, which are chondroitin sulfate-containing PGs. In addition, REF-D expressed mRNA for syndecan-2 and perlecan, which are PGs that contain primarily heparan sulfate chains. CONCLUSIONS: A majority of GAG chains synthesized by subconfluent REF-D are chondroitin sulfate. A smaller proportion of chondroitin sulfate chains associate with a core protein as part of a PG (e.g., biglycan, decorin, syndecan-2). Heparan sulfate chains are also present, with a small proportion associated with a core protein (e.g., the PGs syndecan-2, perlecan). The greater presence of free GAG chains forming weak interactions with surrounding molecules may assist fibroblasts that are moving and replicating during this phase. Therefore, REF-D are particularly well suited to study early wound healing by their expression of chondroitin sulfate chains and associated PGs without the influence of donor age, stage, or anatomic site. PMID- 10864482 TI - Peer teaching and computer-assisted learning: An effective combination for surgical skill training? AB - BACKGROUND: The surgical literature suggests that collaborative learning using peers may be a valid way to teach surgical skills and there is a growing interest in the use of computer-assisted learning for this purpose. Combining this evolving technology with this type of teaching would theoretically offer a number of advantages including a reduction in the amount of faculty time devoted to this task. In this study, we evaluate the efficacy of a type of collaborative learning in a computer-assisted learning environment. MATERIALS AND METHODS: We designed a prospective, randomized study comparing novice learners who were allowed to work in pairs with those who worked independently in a specially equipped computer assisted learning classroom. Both pretest and posttest assessments were performed by videotaping this skill. Three experts then evaluated the videotapes, in a blinded fashion. Three different outcomes were assessed. RESULTS: Seventy-seven subjects were enrolled in and completed the study. Comparison of the outcome measures demonstrated no between group difference in the average performance scores or posttest times. The proportion of subjects who correctly tied a square knot was significantly lower in the computer-assisted peer teaching group when compared with the computer-assisted learning alone group (P = 0.04). CONCLUSIONS: Collaborative learning in a computer-assisted learning environment is not an effective combination for teaching surgical skills to novices. PMID- 10864483 TI - The role of nitric oxide, K(+)(ATP) channels, and cGMP in the preconditioning response of the rabbit. AB - BACKGROUND: The role of nitric oxide (NO), K(+)(ATP) channels, and cyclic GMP (cGMP) in preconditioning is unknown. MATERIAL AND METHODS: Isolated rabbit hearts were pretreated with the NO precursor L-arginine (L-Arg), both alone and after infusion of the NO synthetase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME). Guanylate cyclase inhibitor methylene blue (MB) was infused prior to L Arg in a separate group of hearts. To contrast the mechanisms of NO preconditioning and potassium channel opener (PCO) preconditioning, we infused the PCO pinacidil after L-NAME and the PCO blocker glibenclamide before L-Arg. Control hearts had no drug infused. The LAD coronary artery was occluded for 1 h and reperfused for 1 h in all hearts. Action potential duration (APD(50)), coronary flow (CF), and left ventricular developed pressure (DP) were measured, and infarct size (IS) was determined and expressed as a percentage of the area at risk. RESULTS: L-Arg prolonged APD(50) at 60 min of reperfusion (94 +/- 6 ms vs 69 +/- 2 ms (control) vs 70 +/- 2 ms (L-NAME) vs 74 +/- 3 ms (MB), P < 0.05). L Arg reduced IS compared with control (24 +/- 2% vs 49 +/- 3%, P < 0.05); this was reversed by either L-NAME (53 +/- 4%, P < 0.05) or MB (43 +/- 3%, P < 0.05), but not by glibenclamide (20 +/- 4%), unlike the increase in CF during L-Arg infusion, which was blocked by glibenclamide. Pinacidil infusion decreased IS (26 +/- 2%), but this effect was blocked by L-NAME (53 +/- 7%, P < 0.05 vs pinacidil), although L-NAME did not blunt the increase in CF. There were no significant differences in DP among groups. CONCLUSION: L-Arginine preconditions the heart through NO generation, and this response is mediated through a cGMP dependent mechanism, but is independent of the K(+)(ATP) channels. Coronary vasodilation is mediated through a mechanism different from that responsible for cardiomyocyte preconditioning. PMID- 10864484 TI - Late-gestation tracheal occlusion in the fetal lamb causes rapid lung growth with type II cell preservation. AB - BACKGROUND: Fetal tracheal occlusion (TO) results in varying degrees of lung growth. This study examines whether gestational age influences lung growth response following TO. MATERIALS AND METHODS: Fetal lambs (term = 145 days) underwent TO early (108 days, n = 6) or late (122 days, n = 6) in gestation. Aspirated lung fluid volume (LFV) and intratracheal pressure (ITP) were recorded daily. Two weeks after TO, the fetuses were sacrificed. Lung growth was assessed by lung weight and stereologic volumetry. Type II cellular density was assessed by computer-assisted morphometry using antisurfactant protein B antibody. RESULTS: After early TO, ITP remained below 2 mm Hg for all but one of the first 5 days. In late TO, ITP rose to 4.8 +/- 1.7 mm Hg by Day 1 and remained elevated. LFV remained lower after early than after late TO (P < 0.05) for 8 days. Thereafter, pressure and volume reached similar levels in both TO groups; both were significantly higher than their respective controls (P < 0.05). Parenchymal fraction (1 - air-space fraction) was significantly smaller after late TO (22.8 +/- 1.2%) than after early TO (31.3 +/- 0.5%). Type II density was 38.0 +/- 12.4 x 10(6)/mL after early TO and 84.0 +/- 24.3 x 10(6)/mL in control (P < 0.05); the difference between late TO and control was not significant. CONCLUSIONS: Late tracheal occlusion in fetal lambs caused more rapid lung growth than earlier TO, although ultimate lung size was similar in both groups. Late TO also resulted in greater air-space fraction and better preservation of the type II cell population than early TO. Late-gestation tracheal occlusion may therefore be preferable to prolonged occlusion initiated earlier. PMID- 10864485 TI - Expression of inducible nitric oxide synthase and interleukin-12 in experimental necrotizing enterocolitis. AB - BACKGROUND: Previous investigators have relied on administration of pro inflammatory cytokines or invasive surgical procedures to reproduce the morphologic changes of necrotizing enterocolitis (NEC) in rats. However, these artificial insults do not mimic the human disease. We developed a reproducible model of NEC in rats that more closely resembles human NEC and determined the pattern of inflammatory cytokine expression in this model. MATERIALS AND METHODS: Newborn rats were randomized into four groups. Groups 1 and 2 were breast-fed, while Groups 3 and 4 were gavaged with formula thrice daily. In addition, Groups 2 and 4 were subjected to 3 min of hypoxia thrice daily, prior to each feeding. The rats were killed on day 4 and the distal 2 cm of terminal ileum was harvested for morphological studies and analysis of inflammatory cytokine mRNA expression. RESULTS: Nearly 70% of formula-fed neonatal rats displayed moderate or severe morphological abnormalities resembling human NEC. Breast-fed pups had normal histology. The terminal ileum from rats with abnormal histology demonstrated increased inducible nitric oxide synthase (iNOS) expression, decreased interleukin-12 (IL-12) mRNA expression, and enterocyte apoptosis. There was a trend toward upregulation of IFN-gamma mRNA, but no difference in expression of TNF-alpha mRNA. Hypoxia did not significantly alter intestinal morphology or mRNA expression. CONCLUSIONS: Formula-fed neonatal rats, with or without hypoxia, exhibit morphological changes in the intestinal epithelium similar to those seen in patients with acute NEC. The mechanism likely involves upregulation of iNOS mRNA, enterocyte apoptosis, and decreased IL-12 production in the intestinal epithelium. This model may offer a simple reproducible method for inducing experimental NEC. PMID- 10864486 TI - p27(kip1) expression in rectal cancer correlates with disease-free survival. AB - BACKGROUND: The cell-cycle inhibitor p27(kip1) is a potential tumor suppressor and might serve as a prognostic marker in rectal cancer, in particular with regard to patient selection for adjuvant therapy. MATERIALS AND METHODS: Immunohistochemical analysis was performed, using an anti-p27(kip1) monoclonal antibody, on paraffin sections of two matched [age, gender, UICC stage, year of operation (1982-1991)] groups of patients (n = 2 x 82) with rectal carcinoma curatively treated by surgery alone. The groups differed only in subsequent metachronous distant metastatic spread. All patients had to meet the selection criterion "free of local disease," in order to exclude surgical influence. Follow up was prospective (median of 74 months). The intensity of staining (-, +, ++, ) and rate of positive cells (as a percentage of total tumor volume) were judged separately for cytoplasms and nuclei. RESULTS: On multivariate analysis, cytoplasmic staining intensity proved to be the best prognostic factor of disease free survival and approached statistical significance (P = 0.0552, Cox regression). On univariate analysis, considering cytoplasmic staining alone, intensely stained ( ) tumors showed significantly poorer disease-free survival (vs ++, +, -; Kaplan-Meier, logrank, P = 0.0185). CONCLUSIONS: The demonstrated correlation between cytoplasmic compartmentalization of p27(kip1) and increased metastatic spread as well as disease-free survival underscores the role of p27(kip1) in rectal cancer. However, since other reports emphasize the importance of nuclear p27(kip1) expression, the mechanisms of steady-state and subcellular distribution of p27(kip1) remain unclear, and further investigation is needed. PMID- 10864487 TI - Accelerated replicative senescence of medial smooth muscle cells derived from abdominal aortic aneurysms compared to the adjacent inferior mesenteric artery. AB - BACKGROUND: Abdominal aortic aneurysms (AAAs) are associated with aging and atherosclerosis. AAAs arise through a degenerative process characterized in part by depletion of medial smooth muscle cells (SMC), suggesting that generalized aging and SMC senescence represent potential mechanisms contributing to aneurysmal degeneration. It is not yet known whether SMC from AAA tissue exhibit a difference in proliferative capacity compared to SMC from nonaneurysmal vessels or to what extent such differences might be due to aging alone or other patient specific factors. MATERIALS AND METHODS: Aneurysm wall tissues were obtained from 15 patients undergoing AAA repair. In each case, a segment of the adjacent (nonaneurysmal) inferior mesenteric artery (IMA) from the same patient was used as a control. Paired AAA- and IMA-derived SMC strains were obtained by explant techniques and their proliferative capacities were compared during serial passage in culture. RESULTS: Sustainable SMC cultures were established from all IMA explants but from only 9 of 15 AAAs (P < 0.05). The interval required to achieve primary explant growth was longer for AAAs than IMAs (16.4 +/- 2 vs 6.4 +/- 1 days; P < 0.001), but it was unrelated to patient age, gender, or aneurysm size. AAA-derived SMC appeared larger and rounder than the corresponding IMA-derived SMC, even after repeated passage in culture, and their maximal proliferation was reduced by 44.2 +/- 8% (n = 5 pairs, P < 0.05). Serum-stimulated [(3)H]thymidine uptake in AAA-derived SMC was also reduced by 54.9 +/- 7% (n = 5 pairs, P < 0.01), but flow cytometry revealed no differences in SMC viability, apoptosis, or necrosis. While IMA-derived SMC continued to proliferate beyond passage 20 during serial subculture, all AAA-derived SMC developed replicative senescence by passage 12. CONCLUSIONS: AAA-derived SMC exhibit a distinct morphologic appearance in culture, a diminished proliferative capacity compared to SMC from the adjacent IMA, and a limited in vitro life span. These differences reflect an intrinsic alteration in SMC growth capacity independent of age alone. Tissue specific processes leading to accelerated replicative senescence may therefore contribute to the selective medial SMC depletion observed in AAAs. PMID- 10864488 TI - Thrombin and factor Xa enhance neutrophil chemoattractant production after ischemia/reperfusion in the rat liver. AB - BACKGROUND: Clotting proteases may affect leukocyte effector function. Activation of the coagulation cascade after ischemia/reperfusion stimulates cytokine production by activated macrophages. Cytokine-induced neutrophil chemoattractant (CINC) may also be important in the pathophysiology of liver ischemia/reperfusion injury. We investigated the effects of a selective factor Xa inhibitor, DX-9065a, on CINC expression after ischemia/reperfusion in the rat liver. METHODS: Liver ischemia was induced in rats by occluding the portal vein for 30 min. DX-9065a (9 mg/kg) was injected intravenously 5 min before vascular clamping. Serum CINC concentrations were measured by enzyme-linked immunosorbent assay. Levels of CINC mRNA in the liver were determined by Northern blot analysis. We also examined in vitro CINC production by peritoneal macrophages in response to alpha-thrombin or factor Xa. RESULTS: Serum CINC concentrations increased and peaked 6 h after reperfusion. However, pretreatment of animals with DX-9065a resulted in significantly smaller increases in CINC after reperfusion. Pretreatment with DX 9065a also significantly reduced CINC mRNA levels in the liver after ischemia/reperfusion. In vitro CINC production by peritoneal macrophages was enhanced by alpha-thrombin, as well as factor Xa. CONCLUSIONS: Thrombin and factor Xa stimulate CINC production by macrophages. A selective inhibitor of factor Xa, DX-9065a, attenuates neutrophil chemoattractant production after ischemia/reperfusion injury of the rat liver. PMID- 10864490 TI - Dexamethasone inhibits the phosphorylation of retinoblastoma protein in the suppression of human vascular smooth muscle cell proliferation. AB - We have previously demonstrated that dexamethasone (DEX) suppresses neointimal hyperplasia and proliferation of rat aortic smooth muscle cells (SMC) by inducing a late G1 phase cell cycle arrest. Phosphorylation of retinoblastoma protein (Rb) regulates cell proliferation by controlling progression from G1 to S phase of the cell cycle. We hypothesized that DEX inhibits human vascular SMC proliferation and causes cell cycle arrest through inhibition of Rb phosphorylation. Human aortic SMC were cultured and treated with incremental doses of DEX. Cell counts and [(3)H]thymidine uptake were determined after 72 h. To examine the effects of DEX on the cell cycle, cells were synchronized by serum deprivation, restimulated to enter G1 phase, and treated with 10(-5) M DEX, and protein was extracted at sequential time points. Flow cytometry was performed to track cell cycle progression. Western blots were performed to examine Rb phosphorylation. DEX inhibited smooth muscle cell proliferation and DNA synthesis in a concentration dependent fashion. Flow cytometry indicated that DEX induces a G1 phase cell cycle arrest. DEX inhibited the phosphorylation of Rb protein compared to control. DEX inhibits the proliferation of human vascular SMC by inducing G1 phase cell cycle arrest. DEX inhibited the phosphorylation of Rb, a key step in the progression of the cell from G1 to S phase. Elucidation of the mechanism of DEX may be helpful in treatment strategies for preventing neointimal hyperplasia as well as other disorders of cell proliferation. PMID- 10864489 TI - Characterization of dopamine-mediated relaxation in experimental vein bypass grafts. AB - BACKGROUND: Dopamine is an endogenous inotropic agent commonly used during coronary artery surgery and in the medical therapy of a revascularized patient. In this study the responses of intimal hyperplastic vein grafts to dopamine are examined. METHODS: The in vitro isometric tension responses to dopamine of common carotid jugular vein bypass grafts in New Zealand White rabbits were determined. The responses were compared to those obtained in the jugular vein and in the common carotid artery. Both endothelialized and denuded vessels were precontracted with prostaglandin F(2alpha) and the responses to dopamine were assessed. The contributions of nitric oxide and prostanoids to the response were also determined. RESULTS: Each vessel showed a biphasic dose response to dopamine with relaxation at low concentrations followed by contraction at high concentrations. Dopamine relaxation in the jugular vein was endothelial independent while in the carotid artery it was endothelial dependent and decreased. The sensitivity of both vessels was significantly greater than the vein graft (6.62 +/- 0.12; P < 0. 05); however, after endothelial denudation, the sensitivity of dopamine-mediated relaxation of the vein graft (8.91 +/- 0.09) was significantly enhanced. Preincubation with L-NMMA (to block NO synthesis) inhibited vein graft relaxation to dopamine and preincubation with indomethacin (to block cyclooxygenase activity) inhibited carotid artery relaxation to dopamine. Addition of phenoxybenzamine, a broad alpha-adrenergic antagonist, enhanced dopamine relaxation in the jugular vein and depressed the relaxation in the carotid artery. There was no effect on the dopamine response in the vein graft. Jugular vein and carotid artery responded to dopamine with cholera toxin sensitive (Galpha(s)) responses. In contrast, dopamine relaxation in the vein graft was enhanced by inhibition of Galpha(s). CONCLUSION: Dopamine relaxation in vein grafts is mediated in part by NO but not by either prostanoids or alpha adrenergic receptor activation. It is diminished compared to native vessels due to an endothelium-dependent, Galpha(s)-mediated pathway. PMID- 10864491 TI - Reduction in the level of cardiac cyclic GMP worsens contractile delay in myocardial stunning. AB - We tested the hypothesis that a reduction in the level of myocardial cyclic GMP would worsen the contractile delay associated with myocardial stunning. Two groups of 12 anesthetized open-chest New Zealand white rabbits were utilized. Myocardial stunning was produced by two 15-min occlusions of the left anterior descending coronary artery followed by 15 min of reperfusion. Either control vehicle (saline + 1% DMSO) or 1H-[1,2,4]oxadiazolo[4, 3-a]quinoxalin-1-one (ODQ 10(-4) M, a guanylate cyclase inhibitor) was topically applied to the left ventricular surface of the rabbit hearts. Left ventricular and aortic pressures along with wall thickness parameters were determined. Coronary blood flow (microspheres) and O(2) extraction (microspectrophotometry) were used to determine myocardial O(2) consumption. Myocardial stunning was observed in the control group through an increased delay in onset of wall thickening (46.2 +/- 7.3 vs 76.6 +/- 17.5 ms). There was no significant effect of stunning on the rate of wall thickening (21.8 +/- 9.5 vs 18.1 +/- 3.4 mm/s) or O(2) consumption (stun 4.6 +/- 0.6, control 4.8 +/- 0.4 ml O(2)/min/100 g). After treatment with ODQ 10( 4) M, both delay (43.9 +/- 9.6 vs 134.1 +/- 30.0 ms) and myocardial O(2) consumption (stun 5.9 +/- 0.6, control 5.9 +/- 0. 7) increased significantly compared to control. There was no significant change in the rate of wall thickening. We conclude that decreasing cyclic GMP worsens stunning by increasing delay in onset of wall thickening and increasing local O(2) costs in the stunned region. PMID- 10864492 TI - Rational manipulation of oxygen delivery. AB - BACKGROUND: Optimization of oxygen delivery remains the best method to prevent and the only way to treat common intensive care unit syndromes such as sepsis, multiple organ dysfunction, and acute lung injury. This paper reviews the elements of oxygen delivery, describes how clinical interventions work through those elements to alter oxygen delivery, reviews theoretical and empirical data relating to manipulation of each element, and distinguishes between therapeutic means and clinical endpoints in the care of the critically ill. MATERIALS AND METHODS: Recent literature is reviewed. Relevant equations are detailed. Computer models and patient data illustrate key points. RESULTS: Clinical interventions intended to improve oxygen delivery all work through at least one of seven variables (oxygen saturation, hemoglobin concentration, heart rate, mean arterial blood pressure, systemic vascular resistance, end-diastolic volume, and ejection fraction). Because interventions that increase oxygen delivery are always accompanied by physiologic costs, cavalier application of any therapy in the intensive care unit may actually decrease oxygen delivery, harming the critically ill patient. Various clinical indicators may be used as endpoints to guide therapy. CONCLUSIONS: While a systematic consideration of the elements of oxygen delivery reveals weaknesses in experimental evidence guiding optimal treatment of shock, reasonable strategies as well as avoidable pitfalls emerge from the data. Furthermore, facility with each of the elements of oxygen delivery makes ICU management easier to teach and to apply. PMID- 10864493 TI - Crystal structure of a novel germination protease from spores of Bacillus megaterium: structural arrangement and zymogen activation. AB - The DNA in the core of spores of Bacillus species is saturated with a group of small, acid-soluble proteins (SASP) that protect DNA from a variety of harsh treatments and play a major role in spore resistance and long-term spore survival. During spore germination, SASPs are rapidly degraded to amino acids and this degradation is initiated by a sequence-specific protease called germination protease (GPR), which exhibits no obvious mechanistic or amino acid sequence similarity to any known class of proteases. GPR is synthesized during sporulation as an inactive tetrameric zymogen termed P(46), which later autoprocesses to a smaller form termed P(41), which is active only during spore germination. Here, we report the crystal structure of P(46) from Bacillus megaterium at 3.0 A resolution and the fact that P(46) monomer adopts a novel fold. The asymmetric unit contains two P(46) monomers and the functional tetramer is a dimer of dimers, with an approximately 9 A channel in the center of the tetramer. Analysis of the P(46) structure and site-directed mutagenesis studies have provided some insight into the mechanism of zymogen activation as well as the zymogen's lack of activity and the inactivity of P(41) in the mature spore. PMID- 10864494 TI - A compact monomeric intermediate identified by NMR in the denaturation of dimeric triose phosphate isomerase. AB - The denaturation of triose phosphate isomerase (TIM) from Saccharomyces cerevisiae by guanidine hydrochlorids at pH 7.2 has been monitored by NMR spectroscopy in conjunction with optical spectroscopy. In the absence of denaturant, the hydrodynamic radius of 29.6(+/-0.25) A and the substantial chemical shift dispersion evident in the NMR spectrum are consistent with the highly structured dimeric native state of the protein. On the addition of 2. 2 M guanidine hydrochloride the effective hydrodynamic radius increases to 51.4(+/ 0.43) A, consistent with that anticipated for the polypeptide chain in a highly unstructured random coil state. In 1.1 M guanidine hydrochloride, however, the effective hydrodynamic radius is 24.0(+/-0.25) A, a value substantially decreased relative to that of the native dimeric state but very close to that anticipated for a monomeric species with native-like compaction (23. 5 A). The lack of chemical shift dispersion indicates, however, that few tertiary interactions persist within this species. Far UV CD and intrinsic fluorescence measurements show that this compact intermediate retains significant secondary structure and that on average the fluorophores are partially excluded from solvent. Such a species could be important in the formation of dimeric TIM from its unfolded state. PMID- 10864495 TI - The anti-sigma factor SpoIIAB forms a 2:1 complex with sigma(F), contacting multiple conserved regions of the sigma factor. AB - The developmental regulatory protein sigma(F) of Bacillus subtilis, a member of the sigma(70)-family of bacterial RNA polymerase sigma factors, is negatively regulated by the anti-sigma factor SpoIIAB, which binds to sigma(F), sequestering it in an inactive complex. SpoIIAB binding to sigma(F) is strongly stimulated by ATP. Here, we use a combination of gel filtration chromatography, dynamic light scattering, analytical ultracentrifugation, limited proteolysis with N-terminal sequencing and electrospray mass spectrometry, and deletion analysis to probe the SpoIIAB-sigma(F) complex. The studies were facilitated by investigating the homologs from Bacillus stearothermophilus as well as co-expression of the proteins in Escherichia coli, allowing purification of large quantities of the in vivo assembled complex. We determined the stoichiometry of the complex to be SpoIIAB(2):sigma(F)(1). Alone, sigma(F) is rapidly degraded by the protease trypsin. In the complex with SpoIIAB, however, sigma(F) is remarkably resistant to proteolysis. Analysis of the protease cleavage data indicates the anti-sigma binds sigma(F) through contacts with mutliple conserved regions of the sigma factor, supporting previous findings based on genetic data. PMID- 10864496 TI - Purification and in vitro activities of the Bacillus subtilis TnrA transcription factor. AB - The Bacillus subtilis nitrogen regulatory protein TnrA was purified and its interaction with the nrgAB regulatory region examined. The TnrA protein activates transcription from the nrgAB promoter in vitro. DNase I footprinting and methylation protection experiments demonstrated that TnrA binds to an inverted repeat, upstream of the -35 region of the nrgAB promoter. Gel mobility retardation assays were used to determine the affinity of TnrA for its DNA binding site. The equilibrium dissociation binding constant for the interaction of TnrA with the nrgAB promoter fragment was 7.7 nM under the conditions used here. Mutations in the TnrA consensus sequence that reduce nrgAB expression in vivo were found to reduce significantly the in vitro affinity for TnrA. An A+T rich region located upstream of the TnrA-binding site was found to be necessary for optimal transcriptional activation. A mutant protein, TnrA(HTH), was constructed in which the putative helix-turn-helix DNA-binding motif was altered by exchanging two arginine residues for alanine residues. The TnrA(HTH) protein was unable to activate the in vivo expression of nrgAB and had an in vitro affinity for the nrgAB promoter that was significantly lower than that of the wild-type protein. PMID- 10864497 TI - Characterization of loose and tight dimer forms of avian leukosis virus RNA. AB - Retroviral genomes consist of two identical RNA molecules joined non-covalently near their 5'-ends. Recently, we showed that an imperfect autocomplementary sequence, located in the L3 domain, plays an essential role in avian sarcoma leukosis virus (ASLV) RNA dimerization in vitro. This sequence can adopt a stem loop structure and is involved in ASLV replication. Here, we found that in the absence of nucleocapsid protein, RNA transcripts of avian leukosis virus (ALV) were able to form two types of dimers in vitro that differ in their stability: a loose dimer, formed at a physiological temperature, and a tight dimer, formed at a high temperature. A mutational analysis was performed to define the features of these dimers. The results of this analysis unambiguously confirm that the two L3 stem-loops interact directly in both types of dimers. A loop-loop interaction is the main linkage in the loose dimer. In contrast, in the tight dimer, the stem and the loop of the L3 hairpin form an extended duplex. Surprisingly, we also found that the dimerization properties defined for our ALV strain (type SR-A) differ from those found in other ASLV strains. PMID- 10864498 TI - The spacing between functional Cis-elements of U3 snoRNA is critical for rRNA processing. AB - The sequences and structural features of Xenopus laevis U3 small nucleolar RNA (snoRNA) necessary for pre-rRNA cleavage at sites 1 and 2 to form 18 S rRNA were assayed by depletion/rescue experiments in Xenopus oocytes. Mutagenesis results demonstrated that the putative stem of U3 domain I is unnecessary for 18 S rRNA processing. A model consistent with earlier experimental data is proposed for the structure of domain I when U3 is not yet bound to pre-rRNA. For its function in rRNA processing, a newly discovered element (5' hinge) was revealed to be important but not as critical as the 3' hinge region in Xenopus U3 snoRNA for 18 S rRNA formation. Base-pairing is proposed to occur between the U3 5' hinge and 3' hinge and complementary regions in the external transcribed spacer (ETS); these interactions are phylogenetically conserved, and are homologous to those previously described in yeast (5' hinge-ETS) and trypanosomes (3' hinge-ETS). A model is presented where the base-pairing of the 5' hinge and 3' hinge of U3 snoRNA with the ETS of pre-rRNA helps to correctly position U3 boxes A'+A for their function in rRNA processing. Like an earlier proposal for yeast, boxes A' and A of Xenopus may base-pair with 18 S sequences in pre-rRNA. We present the first direct experimental evidence in any system that box A' is essential for U3 snoRNA function in 18 S rRNA formation. The analysis of insertions and deletions indicated that the spacing between the U3 elements is important, suggesting that they base-pair with the ETS and 18 S regions of pre-rRNA at the same time. PMID- 10864499 TI - Premature termination of DNA replication in plasmids carrying two inversely oriented ColE1 origins. AB - In Escherichia coli plasmids carrying two inversely oriented ColE1 origins, DNA replication initiates at only one of the two potential origins. The other silent origin acts as a replication fork barrier. Whether this barrier is permanent or simply a pausing site remains unknown. Here, we used a repeated primer extension assay to map in vivo, at the nucleotide level, the 5' end of the nascent strand where initiation and blockage of replication forks occurs. Initiation occurred primarily at the previously defined origin, however, an alternative initiation site was detected 17 bp upstream. At the barrier, the lagging strand also terminated at the main initiation site. Therefore, the 5' end of the nascent strand at the barrier was identical to that generated during initiation. This observation strongly suggests that blockage of the replication fork at the silent origin is not just a pausing site but permanent, and leads to a premature termination event. PMID- 10864500 TI - Canonical antigen-binding loop structures in immunoglobulins: more structures, more canonical classes? AB - Grafting the antigen-binding loops onto a human antibody scaffold is a widely used technique to humanise murine antibodies. The success of this approach depends largely on the observation that the antigen-binding loops adopt only a limited number of canonical structures. Identification of the correct canonical structure is therefore essential. Algorithms that predict the main-chain conformation of the hypervariable loops using only the amino acid sequence often provide this information. Here, we describe new canonical loop conformations for the hypervariable regions H1 and H2 as found in single-domain antibody fragments of dromedaries or llama. Although the occurrence of these new loop conformations was not predicted by the algorithms used, it seems that they could occur in human or mouse antigen-binding loops. Their discovery indicates that the currently used set of canonical structures is incomplete and that the prediction algorithms should be extended to include these new structures. PMID- 10864501 TI - Alternative conformations of a nucleic acid four-way junction. AB - A crystal structure of a 108 nucleotide RNA-DNA complex containing a four-way junction was solved at 3.1 A resolution. The structure of the junction differs substantially from the "stacked-X" conformation observed previously, due to a 135 degrees rotation of the branches. Comparison of the two conformers provides insight into the factors contributing to the flexibility of four-way junctions. The stacked-X conformation maximizes base-stacking but causes unfavorable repulsion between phosphate groups, whereas the 135 degrees -rotated "crossed" conformation minimizes electrostatic clashes at the expense of reduced base stacking. Despite the large rotation of the branches, both junction structures exhibit an antiparallel arrangement of the continuous strands and opposite polarity of the crossover strands. PMID- 10864502 TI - Structural characterization and membrane binding properties of the matrix protein VP40 of Ebola virus. AB - The matrix protein VP40 of Ebola virus is believed to play a central role in viral assembly as it targets the plasma membrane of infected cells and subsequently forms a tightly packed layer on the inner side of the viral envelope. Expression of VP40 in Escherichia coli and subsequent proteolysis yielded two structural variants differing by a C-terminal truncation 114 amino acid residues long. As indicated by chemical cross-linking studies and electron microscopy, the larger polypeptide was present in a monomeric form, whereas the truncated one formed hexamers. When analyzed for their in vitro binding properties, both constructs showed that only monomeric VP40 efficiently associated with membranes containing negatively charged lipids. Membrane association of truncated, hexameric VP40 was inefficient, indicating a membrane recognition role for the C-terminal part. Based on these observations we propose that assembly of Ebola virus involves the formation of VP40 hexamers that is mediated by the N-terminal part of the polypeptide. PMID- 10864503 TI - Structure and function of the conserved 690 hairpin in Escherichia coli 16 S ribosomal RNA: analysis of the stem nucleotides. AB - Nucleotides 680 to 710 of Escherichia coli 16 S rRNA form a distinct structural domain required for ribosome function. The goal of this study was to determine the functional significance of pairing interactions in the 690 region. Two different secondary structures were proposed for this hairpin, based on phylogenetic and chemical modification studies. To study the effect of pairing interactions in the 690 hairpin on ribosome function and to determine which of the proposed secondary structures is biologically significant, we performed an instant-evolution experiment in which the nine nucleotides that form the proposed base-pairs and dangling ends of the 690 stem were randomly mutated, and functional mutant combinations were selected. A total of 96 unique functional mutants were isolated, assayed in vivo, and sequenced. Analysis of these data revealed extensive base-pairing and stacking interactions among the mutated nucleotides. Formation of either a Watson-Crick base-pair or G.U pair between positions 688 and 699 is absolutely required for ribosome function. We also performed NMR studies of a 31-nucleotide RNA which indicate the formation of a functionally important base-pair between nucleotides 688 and 699. Formation of a second base-pair between positions 689 and 698, however, is not essential for ribosome function, but the level of ribosome function correlates with the predicted thermodynamic stability of the nucleotide pairs in these positions. The universally conserved positions G690 and U697 are generally portrayed as forming a G.U mismatch. Our data show co-variation between these positions, but do not support the hypothesis that the G690:U697 pair forms a wobble structure. NMR studies of model 14-nt and 31-nt RNAs support these findings and show that G690 and U697 are involved in unusual stacking interactions but do not form a wobble pair. Preliminary NMR structural analysis reveals that the loop portion of the 690 hairpin folds into a highly structured and novel conformation. PMID- 10864504 TI - Complexes of porcine odorant binding protein with odorant molecules belonging to different chemical classes. AB - Porcine odorant binding protein (pOBP) is a monomer of 157 amino acid residues, purified in abundance from pig nasal mucosa. In contrast to the observation on lipocalins as retinol binding protein (RBP), major urinary protein (MUP) or bovine odorant binding protein (bOBP), no naturally occurring ligand was found in the beta-barrel cavity of pOBP. Porcine OBP was therefore chosen as a simple model for structure/function studies with odorant molecules. In competition experiments with tritiated pyrazine, the affinity of pOBP towards several odorant molecules belonging to different chemical classes has been found to be of the micromolar order, with a 1:1 stoichiometry. The X-ray structures of pOBP complexed to these molecules were determined at resolution between 2.15 and 1.4 A. As expected, the electron density of the odorant molecules was observed into the hydrophobic beta-barrel of the lipocalin. Inside this cavity, very few specific interactions were established between the odorant molecule and the amino acid side-chains, which did not undergo significant conformational change. The high B-factors observed for the odorant molecules as well as the existence of alternative conformations reveal a non-specific mode of binding of the odorant molecules in the cavity. PMID- 10864505 TI - Structural and biochemical investigations of the catalytic mechanism of an NADP dependent aldehyde dehydrogenase from Streptococcus mutans. AB - The NADP-dependent non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase from Streptococcus mutans (abbreviated Sm-ALDH) belongs to the aldehyde dehydrogenase (ALDH) family. Its catalytic mechanism proceeds via two steps, acylation and deacylation. Its high catalytic efficiency at neutral pH implies prerequisites relative to the chemical mechanism. First, the catalytic Cys284 should be accessible and in a thiolate form at physiological pH to attack efficiently the aldehydic group of the glyceraldehyde-3-phosphate (G3P). Second, the hydride transfer from the hemithioacetal intermediate toward the nicotinamide ring of NADP should be efficient. Third, the nucleophilic character of the water molecule involved in the deacylation should be strongly increased. Moreover, the different complexes formed during the catalytic process should be stabilised. The crystal structures presented here (an apoenzyme named Apo2 with two sulphate ions bound to the catalytic site, the C284S mutant holoenzyme and the ternary complex composed of the C284S holoenzyme and G3P) together with biochemical results and previously published apo and holo crystal structures (named Apo1 and Holo1, respectively) contribute to the understanding of the ALDH catalytic mechanism. Comparison of Apo1 and Holo1 crystal structures shows a Cys284 side-chain rotation of 110 degrees, upon cofactor binding, which is probably responsible for its pK(a) decrease. In the Apo2 structure, an oxygen atom of a sulphate anion interacts by hydrogen bonds with the NH2 group of a conserved asparagine residue (Asn154 in Sm-ALDH) and the Cys284 NH group. In the ternary complex, the oxygen atom of the aldehydic carbonyl group of the substrate interacts with the Ser284 NH group and the Asn154 NH2 group. A substrate isotope effect on acylation is observed for both the wild-type and the N154A and N154T mutants. The rate of the acylation step strongly decreases for the mutants and becomes limiting. All these results suggest the involvement of Asn154 in an oxyanion hole in order to stabilise the tetrahedral intermediate and likely the other intermediates of the reaction. In the ternary complex, the cofactor conformation is shifted in comparison with its conformation in the C284S holoenzyme structure, likely resulting from its peculiar binding mode to the Rossmann fold (i.e. non perpendicular to the plane of the beta-sheet). This change is likely favoured by a characteristic loop of the Rossmann fold, longer in ALDHs than in other dehydrogenases, whose orientation could be constrained by a conserved proline residue. In the ternary and C284S holenzyme structures, as well as in the Apo2 structure, the Glu250 side-chain is situated less than 4 A from Cys284 or Ser284 instead of 7 A in the crystal structure of the wild-type holoenzyme. It is now positioned in a hydrophobic environment. This supports the pK(a) assignment of 7.6 to Glu250 as recently proposed from enzymatic studies. PMID- 10864506 TI - Crystal structure of tobacco necrosis virus at 2.25 A resolution. AB - The crystal structure of tobacco necrosis virus (TNV) has been determined by real space averaging with 5-fold non-crystallographic symmetry, and refined to R=25.3 % for diffraction data to 2.25 A resolution. A total of 180 subunits form a T=3 virus shell with a diameter of about 280 A and a small protrusion at the 5-fold axis. In 276 amino acid residues, the respective amino terminal 86, 87 and 56 residues of the A, B and C subunits are disordered. No density for the RNA was found. The subunits have a "jelly roll" beta-barrel structure, as have the structures of the subunits of other spherical viruses. The tertiary and quaternary structures of TNV are, in particular, similar to those of southern bean mosaic virus, although they are classified in different groups. Invisible residues 1 to 56 with a high level of basic residues are considered to be located inside the particle. Sequence comparison of the coat proteins of several TNV strains showed that the sequences of the disordered segment diverge considerably as compared with those of the ordered segment, consistent with a small tertiary structural constraint being imposed on the N-terminal segment. Basic residues are localized on the subunit interfaces or inner surface of the capsid. Positive charges of the basic residues facing the interior, as well as those of the N terminal segment, may neutralize the negative charge of the RNA inside. Five calcium ions per icosahedral asymmetric unit are located at the subunit interfaces; three are close to the exterior surface, the other two away from it. The environments of the first three are similar, and those of the other two sites are similar. These calcium ions are assumed to be responsible for the stabilization/transition of the quaternary structure of the shell. Three peptide segments ordered only in the C subunits are clustered around each 3-fold (quasi-6 fold) axis forming a beta-annulus, and may lead to quasi-equivalent interactions for the organization of the T=3 shell. PMID- 10864507 TI - Ab initio construction of protein tertiary structures using a hierarchical approach. AB - We present a hierarchical method to predict protein tertiary structure models from sequence. We start with complete enumeration of conformations using a simple tetrahedral lattice model. We then build conformations with increasing detail, and at each step select a subset of conformations using empirical energy functions with increasing complexity. After enumeration on lattice, we select a subset of low energy conformations using a statistical residue-residue contact energy function, and generate all-atom models using predicted secondary structure. A combined knowledge-based atomic level energy function is then used to select subsets of the all-atom models. The final predictions are generated using a consensus distance geometry procedure. We test the feasibility of the procedure on a set of 12 small proteins covering a wide range of protein topologies. A rigorous double-blind test of our method was made under the auspices of the CASP3 experiment, where we did ab initio structure predictions for 12 proteins using this approach. The performance of our methodology at CASP3 is reasonably good and completely consistent with our initial tests. PMID- 10864508 TI - ATPase cycle of an archaeal chaperonin. AB - Recent structural data imply differences in allosteric behavior of the group I chaperonins, typified by GroEL from Escherichia coli, and the group II chaperonins, which comprise archaeal thermosome and eukaryotic TRiC/CCT. Therefore, this study addresses the mechanism of interaction of adenine nucleotides with recombinant alpha-only and native alphabeta-thermosomes from Thermoplasma acidophilum, which also enables us to analyze the role of the heterooligomeric composition of the natural thermosome. Although all subunits of the alpha-only thermosome seem to bind nucleotides tightly and independently, the native chaperonin has two different classes of ATP-binding sites. Furthermore, for the alpha-only thermosome, the steady-state ATPase rate is determined by the cleavage reaction itself, whereas, for the alphabeta-thermosome, the rate limiting step is associated with a post-hydrolysis isomerisation into a non covalent ADP*P(i) species prior to the release of the gamma-phosphate group. After half-saturation with ATP, a negative cooperativity in hydrolysis is observed for both thermosomes. The effect of Mg(2+) and K(+) nucleotide cycling is documented. We conclude that archaeal chaperonins have unique allosteric properties and discuss them in the light of the mechanism established for the group I chaperonins. PMID- 10864509 TI - Global folds of proteins with low densities of NOEs using residual dipolar couplings: application to the 370-residue maltodextrin-binding protein. AB - The global fold of maltose-binding protein in complex with the substrate beta cyclodextrin was determined by solution NMR methods. The two-domain protein is comprised of a single polypeptide chain of 370 residues, with a molecular mass of 42 kDa. Distance information in the form of H(N)-H(N), H(N)-CH(3) and CH(3)-CH(3) NOEs was recorded on (15)N, (2)H and (15)N, (13)C, (2)H-labeled proteins with methyl protonation in Val, Leu, and Ile (C(delta1) only) residues. Distances to methyl protons, critical for the structure determination, comprised 77 % of the long-range restraints. Initial structures were calculated on the basis of 1943 NOEs, 48 hydrogen bond and 555 dihedral angle restraints. A global pair-wise backbone rmsd of 5.5 A was obtained for these initial structures with rmsd values for the N and C domains of 2.4 and 3.8 A, respectively. Direct refinement against one-bond (1)H(N)-(15)N, (13)C(alpha)-(13)CO, (15)N-(13)CO, two-bond (1)H(N) (13)CO and three-bond (1)H(N)-(13)C(alpha) dipolar couplings resulted in structures with large numbers of dipolar restraint violations. As an alternative to direct refinement against measured dipolar couplings we have developed an approach where discrete orientations are calculated for each peptide plane on the basis of the dipolar couplings described above. The orientation which best matches that in initial NMR structures calculated from NOE and dihedral angle restraints exclusively is used to refine further the structures using a new module written for CNS. Modeling studies from four different proteins with diverse structural motifs establishes the utility of the methodology. When applied to experimental data recorded on MBP the precision of the family of structures generated improves from 5.5 to 2.2 A, while the rmsd with respect to the X-ray structure (1dmb) is reduced from 5.1 to 3.3 A. PMID- 10864510 TI - Design and evolution of artificial M13 coat proteins. AB - Using simple design and selective pressure, we have evolved an artificial M13 bacteriophage coat protein. M13 coat proteins first reside in the bacterial inner membrane and subsequently surround the DNA core of the assembled virus. The artificial coat protein (ACP) was designed and evolved to mimic both functions of the natural M13 coat proteins, but with an inverted orientation. ACP is a non functional coat protein because it is not required for the production of phage particles. Instead, it incorporates into a phage coat which still requires all the natural coat proteins for structural integrity. In contrast with other M13 coat proteins, which can display polypeptides as amino-terminal fusions, ACP permits the carboxy-terminal display of large polypeptides. The results suggest that viruses can co-opt host membrane proteins to acquire new coat proteins and thus new functions. In particular, M13 bacteriophage can be engineered for new functions, such as carboxy-terminal phage display. PMID- 10864511 TI - Correlated motions in native proteins from MS analysis of NH exchange: evidence for a manifold of unfolding reactions in ovomucoid third domain. AB - Native-state amide hydrogen exchange monitored by NMR spectroscopy and mass spectrometry (MS) has the potential to provide detailed residue-level information regarding correlated motions occurring on the microseconds to seconds timescale. To expand the applicability of MS to these studies, a new algorithm has been developed to interpret MS data for exchange occurring between the EX2 and EX1 kinetic limits. Re-interpretation of MS data for ovomucoid third domain reveals multiple unfolding or partial unfolding reactions. PMID- 10864512 TI - Plasmodium berghei: cerebral malaria in CBA mice is not clearly related to plasma TNF levels or intensity of histopathological changes. AB - Plasmodium berghei ANKA infection in CBA/J mice leads to the development of cerebral malaria (CM) that kills 80-90% of the animals in 6-9 days. This model has been used to study the pathogenesis of CM, which is a major cause of morbidity and mortality in Plasmodium falciparum-infected individuals. The role of cytokines in the induction of CM in the murine model has been well documented, but most studies have been restricted to the peak of neurological manifestations. Here we used a sequential approach to compare mice that developed CM with those that developed no cerebral pathology. Animals were examined for systemic histopathological changes and plasma Tumor Necrosis Factor-alpha (TNF) levels. The objectives were (a) to further determine the importance of factors commonly associated with murine CM-such as elevated levels of TNF and the presence of hemorrhage and vascular plugging-by comparing mice at different stages of infection and/or with different outcomes following infection and (b) to examine the importance of systemic changes-course of parasitemia and histopathological alterations in brain, liver, and lungs-in the development of CM. The data suggest that (a) the clinical manifestation of CM appears to be associated with a wave of merozoite release on days 6-7, (b) murine CM does not present reliable histopathological indicators, (c) there is no topographic association between the occurrence of intravascular plugging and the hemorrhagic foci, (d) monocyte monocyte and monocyte-endothelial cell adherence were the most expressive histopathological events and were not restricted to brain vessels, (e) blood levels of TNF are not indicative of the local tissue reaction, (f) adhesiveness of monocyte/endothelial cells fluctuate during infection and is dissociated from the lymphocyte homing to the liver, and (g) pulmonary megakaryocytosis (megakaryopoiesis?) is a late event in the lungs. PMID- 10864513 TI - Schistosoma japonicum: day to day variation in excretion and hatchability of parasite eggs in the domestic pig, Suis suis. AB - The aim of the present study was to describe the course of an Schistosoma japonicum infection in individual pigs over time, with special regards to fecal egg counts and egg hatchability, emphasizing the extent of variation during and between days. Five specific pathogen free Danish Landrace/Yorkshire/Duroc crossbred male pigs were each given 3500 cercariae intramuscularly. From day 36 to 62 post infection, fecal samples were collected from each pig, morning and evening. Pigs were perfused 62 days post infection. Actual fecal egg counts, miracidial counts, and worm burdens were determined. The trend of fecal egg excretion was adequately described with a third order polynomial and logarithmic link function. Miracidial counts were related directly to morning fecal egg counts through a simple linear function. The study revealed a fairly similar overall pattern of egg excretion for all pigs, showing an increase in egg excretion until week 8, followed by a marked reduction to almost zero. In general, large fluctuations around this trend were seen for all pigs, as revealed in variations in egg counts within the same day as well as between days. However, as revealed by the estimated model lines, these fluctuations are due to random variation in egg density in stools rather than being caused by biologically determined cycles. An exact time for both first occurrence of eggs in feces (days 35-38 post infection) and for peak egg counts (days 44-48 post infection) was predicted for each individual pig. Furthermore, the model revealed that miracidial counts can be related directly to the expected fecal egg counts of the same day through a parameter, which represents hatchability. This hatchability parameter was found to be independent of time post infection for each pig, but large individual differences were seen in hatchability between the pigs. PMID- 10864514 TI - Inducible nitric oxide synthase and nitrotyrosine in the central nervous system of mice chronically infected with Trypanosoma brucei brucei. AB - Human African trypanosomiasis, or sleeping sickness, evolves toward a meningoencephalitic stage, with a breakage in the blood-brain barrier, perivascular infiltrates, and astrocytosis. The involvement of nitric oxide (NO) has been evoked in the pathogenic development of the illness, since NO was found to be increased in the brain of animals infected with Trypanosoma brucei (T. b.) brucei. An excessive NO production can lead to alterations of neuronal signaling and to cell damage through the cytotoxicity of NO and its derivatives, especially peroxynitrites. In African trypanosomiasis, the sites of NO production and its role in the pathogenicity of lesions in the central nervous system (CNS) are unknown. In a chronic model of African trypanosomiasis (mice infected with T. b. brucei surviving with episodic suramin administration), NADPH-diaphorase staining of brain slides revealed that NO synthase (NOS) activity is located not only in endothelial cells, choroid plexus ependymal cells, and neurons as in control mice but also in mononuclear inflammatory cells located in perivascular and parenchyma infiltrates. An immunohistochemical study showed that the mononuclear inflammatory cells expressed an inducible NOS activity. Furthermore, the presence of nitrotyrosine in inflammatory lesions demonstrated an increased NO production and the intermediate formation of peroxynitrites. The detection of extensive formation of nitrotyrosine in the CNS parenchyma was observed in mice having shown neurological disorders, suggesting the role of peroxynitrites in the appearance of neurological troubles. In conclusion, this study confirmed the increased NO synthesis in the CNS of mice infected with T. b. brucei and suggests a deleterious role for NO, through the formation of peroxynitrites, in the pathogenesis of African CNS trypanosomiasis. PMID- 10864515 TI - Inhibition and stimulation of growth of Entamoeba histolytica in culture: association with PKC activity and protein phosphorylation. AB - We studied the role of protein kinase C (PKC) and protein threonine phosphorylation in the inhibition and stimulation of growth of the protozoan parasite Entamoeba histolytica. PKC was activated after serum deprivation in E. histolytica and during this period proteins became threonine phosphorylated. Conversely, on serum stimulation of serum-deprived cells, PKC activation was rapidly reversed and the threonine phosphorylation of proteins quickly declined. Growth of E. histolytica was not affected by either PKC inhibitors H-7 and GF109203X or by down-regulation of PKC by Phorbol 12-Myristate 13-Acetate (PMA). Interestingly, very low doses of PMA which caused activation of PKC and were unable to down-regulate PKC after 48 h of culture, negatively influenced the growth of E. histolytica. Serine/threonine phosphatase inhibitors Okadaic acid and Calyculin A drastically inhibited growth of E. histolytica. In conclusion, the growth of E. histolytica is not adversely affected by PKC down-regulation. On the contrary, growth inhibition of E. histolytica is associated with activation of Ca(2+), Diacylglycerol (DAG)-dependent PKC, and threo nine phosphorylation of proteins. PMID- 10864517 TI - The salivary adenosine deaminase from the sand fly Lutzomyia longipalpis. AB - In the process of sequencing a subtracted cDNA library from the salivary glands of the sand fly Lutzomyia longipalpis, we identified a cDNA with similarities to gene products of the adenosine deaminase family. Prompted by this cDNA finding, we detected adenosine deaminase activity at levels of 1 U/mg protein in salivary gland homogenates. The activity was significantly reduced following a blood meal indicating its apparent secretory fate. The native enzyme has a K(m) of approximately 10 microM, an isoelectric pH between 4.5 and 5.5, and an apparent molecular weight of 52 kDa by size exclusion chromatography. The possible role of this enzyme, which converts adenosine to inosine, in the feeding physiology of L. longipalpis is discussed. PMID- 10864516 TI - Trichuris suis: a secretory chymotrypsin/elastase inhibitor with potential as an immunomodulator. AB - A serine protease inhibitor, termed TsCEI, was purified from adult-stage Trichuris suis by acid precipitation, affinity chromatography (elastase-agarose), and reverse-phase HPLC. The molecular weight of TsCEI was estimated at 6.437 kDa by laser desorption mass spectrometry. TsCEI potently inhibited both chymotrypsin (K(i) = 33.4 pM) and pancreatic elastase (K(i) = 8.32 nM). Neutrophil elastase, chymase (mouse mast cell protease-1, mMCP-1), and cathepsin G were also inhibited by TsCEI, whereas trypsin, thrombin, and factor Xa were not. The cDNA-derived amino acid sequence of the mature TsCEI consisted of 58 residues including 9 cysteine residues with a molecular mass of 6.196 kDa. TsCEI displayed 48% sequence identity to a previously characterized trypsin/chymotrypsin inhibitor of T. suis, TsTCI. TsCEI showed 36% sequence identity to a protease inhibitor from the hemolymph of the honeybee Apis mellifera. Sequence similarity was also detected with the trypsin/thrombin inhibitor of the European frog Bombina bombina, the elastase isoinhibitors of the nematode Anisakis simplex, and the chymotrypsin/elastase and trypsin inhibitors of the nematode Ascaris suum. The inhibitors of T. suis, an intestinal parasite of swine, may function as components of a parasite defense mechanism by modulating intestinal mucosal mast cell-associated, protease-mediated, host immune responses. PMID- 10864518 TI - Trichomonas vaginalis: characterization, expression, and phylogenetic analysis of a carbamate kinase gene sequence. AB - The gene encoding carbamate kinase (CBK, ATP:carbamate phosphotransferase, EC 2.7.2.2) from Trichomonas vaginalis has been sequenced and its expression in this protozoon has been verified using reverse-transcription polymerase chain reaction. The codon usage and percentage nucleotide composition in the coding and noncoding regions are consistent with other genes isolated from this parasite. Phylogenetic analysis of this gene has suggested possible speciation events that are congruent with other studies, with suggestions of lateral gene transfer. The gene was expressed in Escherichia coli using the pQE-30 expression system, and the recombinant protein was purified using affinity chromatography. The expression of the recombinant protein was identified via Western blotting and matrix-assisted laser desorption ionization mass spectrometry of tryptic peptides. Preliminary kinetic assays have revealed that the recombinant enzyme has a K(m) similar to that of the native enzyme and size-exclusion chromatography has shown that the enzyme is active as the homodimer. PMID- 10864519 TI - Fasciola hepatica: heterologous expression and functional characterization of a thioredoxin peroxidase. AB - A Fasciola hepatica cDNA clone of 779 bp was isolated from an adult worm cDNA expression library by immunological screening using a rabbit serum against the excretory-secretory antigens. The nucleotide sequence of the cDNA revealed the presence of an open reading frame of 582 bp which encoded a 194-amino-acid residue polypeptide (M(r) 21,723 Da) showing a high degree of homology to thioredoxin peroxidases. This putative antioxidant protein gene was expressed in Escherichia coli as a GST fusion protein. The recombinant fusion protein showed in vitro antioxidant properties and protected rabbit muscle enolase and E. coli glutamine synthetase from inactivation by nonenzymatic Fe(3+)/O(2)/DTT or Fe(3+)/O(2)/ascorbate metal-catalyzed oxidation systems. PMID- 10864520 TI - Trypanosoma cruzi: of man, kissing-bugs, and frogs. PMID- 10864521 TI - Plasmodium falciparum: adhesion of placental isolates modulated by the sulfation characteristics of the glycosaminoglycan receptor. PMID- 10864522 TI - The risks of having children in later life. Social advantage may make up for biological disadvantage. PMID- 10864523 TI - The epidemiology of stomach cancer: correlating the past with the present. Socioeconomic influences in early life can influence mortality in adult life. PMID- 10864524 TI - How the NHS can improve safety and learning. By learning free lessons from near misses. PMID- 10864525 TI - Isolated systolic hypertension: a radical rethink. It's a risk factor that needs treatment, especially in the over 50s. PMID- 10864526 TI - Giving medicine a fair trial. Trials should not second guess what patients want. PMID- 10864527 TI - France heads WHO's league table of health systems. PMID- 10864529 TI - NICE approves taxanes for breast cancer PMID- 10864528 TI - US Supreme Court bars federal lawsuits against HMOs. PMID- 10864530 TI - In brief PMID- 10864531 TI - English NHS to set up new reporting system for errors. PMID- 10864533 TI - Ombudsman slams deputising service PMID- 10864532 TI - Doctors need more training in delivering breech babies. PMID- 10864534 TI - Gangrene bug "killed 35 heroin users". PMID- 10864535 TI - US cancer institute funds trial of complementary therapy. PMID- 10864536 TI - Legality of European ban on tobacco advertising questioned. PMID- 10864537 TI - European court of justice likely to annul ban PMID- 10864538 TI - MPs want a tobacco regulatory authority. PMID- 10864540 TI - Access to investigations is needed 7 days a week PMID- 10864539 TI - Health and social services "locked in a vicious circle". PMID- 10864541 TI - GPs want self regulation to continue PMID- 10864542 TI - Management guidelines for women with normal colposcopy after low grade cervical abnormalities: population study. AB - OBJECTIVES: To develop an evidence based protocol for the follow up of women with low grade cervical abnormalities for whom treatment is not immediately indicated. DESIGN: Population outcome study. SETTING: Colposcopy clinic of an inner city teaching hospital. PARTICIPANTS: 566 women with low grade cytological abnormalities who were not treated at a first visit to the colposcopy clinic, followed up for a total of 881 years. MAIN OUTCOME MEASURES: Resolution of abnormalities, persistence of disease, and treated disease. RESULTS: Abnormalities resolved in 306 (54.1%) women, whereas 138 (24.4%) had persistent disease and 122 (21.5%) were subsequently treated. Colposcopic opinion, smear test results, age, smoking history, and number of pregnancies were all significantly related to outcome. Logistic regression analysis produced a model that correctly identified 70% of women whose abnormalities resolved. Only 23 of 295 women (7.8%) with a normal cervix on colposcopy and a smear without dyskaryosis at a first visit were treated by the end of the observation period. CONCLUSIONS: Women referred with low grade cytological abnormalities who have a normal cervix on colposcopy and a negative or borderline repeat smear test result may be discharged from the colposcopy clinic. We propose a follow up protocol that could safely avoid unnecessary visits to a clinic. PMID- 10864544 TI - New insights into mental illness PMID- 10864543 TI - Effects of treatment for intestinal helminth infection on growth and cognitive performance in children: systematic review of randomised trials. AB - OBJECTIVE: To summarise the effects of anthelmintic drug treatment on growth and cognitive performance in children. DATA SOURCES: Electronic databases: Cochrane Infectious Diseases Group controlled trial register, Cochrane controlled trials register, Embase, and Medline. Citations of all identified trials. Contact with the World Health Organization and field researchers. REVIEW METHODS: Systematic review of randomised controlled trials in children aged 1-16 that compared anthelmintic treatment with placebo or no treatment. Assessment of validity and data abstraction conducted independently by two reviewers. MAIN OUTCOME MEASURES: Growth and cognitive performance. RESULTS: Thirty randomised controlled trials in more than 15 000 children were identified. Effects on mean weight were unremarkable, and heterogeneity was evident in the results. There were some positive effects on mean weight change in the trials reporting this outcome: after a single dose (any anthelmintic) the pooled estimates were 0.24 kg (95% confidence interval 0.15 kg to 0. 32 kg; fixed effects model assumed) and 0.38 kg (0.01 kg to 0.77 kg; random effects model assumed). Results from trials of multiple doses showed mean weight change in up to one year of follow up of 0.10 kg (0.04 kg to 0.17 kg; fixed effects) or 0.15 kg (0.00 to 0.30; random effects). At more than one year of follow up, mean weight change was 0.12 kg (-0.02 kg to 0.26 kg; fixed effects) and 0.43 (-0.61 to 1. 47; random effects). Results from studies of cognitive performance were inconclusive. CONCLUSIONS: There is some limited evidence that routine treatment of children in areas where helminths are common has effects on weight gain, but this is not consistent between trials. There is insufficient evidence as to whether this intervention improves cognitive performance. PMID- 10864545 TI - Prospective audit of incidence of prognostically important myocardial damage in patients discharged from emergency department. AB - OBJECTIVE: To assess the incidence of prognostically important myocardial damage in patients with chest pain discharged from the emergency department. DESIGN: Prospective observational study. SETTING: District general hospital emergency department. PARTICIPANTS: 110 patients presenting with chest pain of unknown cause who were subsequently discharged home after cardiac causes of chest pain were ruled out by clinical and electrocardiographic investigation. INTERVENTIONS: Patients were reviewed 12-48 hours after presentation by repeat electrocardiography and measurement of cardiac troponin T. MAIN OUTCOME MEASURES: Incidence of missed myocardial damage. RESULTS: Eight (7%) patients had detectable cardiac troponin T on review and seven had concentrations >/=0.1 microg/l. The repeat electrocardiogram showed no abnormality in any patient. CONCLUSION: 6% of the patients discharged from the emergency department had missed prognostically important myocardial damage. Follow up measurement of cardiac troponin T allows convenient audit of clinical performance in the emergency department. PMID- 10864546 TI - Infant mortality, stomach cancer, stroke, and coronary heart disease: ecological analysis. PMID- 10864548 TI - Amusing the patient PMID- 10864547 TI - Role of C282Y mutation in haemochromatosis gene in development of type 2 diabetes in healthy men: prospective cohort study. PMID- 10864549 TI - "You heard all wrong" PMID- 10864550 TI - Maternal age and fetal loss: population based register linkage study. AB - OBJECTIVE: To estimate the association between maternal age and fetal death (spontaneous abortion, ectopic pregnancy, stillbirth), taking into account a woman's reproductive history. DESIGN: Prospective register linkage study. SUBJECTS: All women with a reproductive outcome (live birth, stillbirth, spontaneous abortion leading to admission to hospital, induced abortion, ectopic pregnancy, or hydatidiform mole) in Denmark from 1978 to 1992; a total of 634 272 women and 1 221 546 pregnancy outcomes. MAIN OUTCOME MEASURES: Age related risk of fetal loss, ectopic pregnancy, and stillbirth, and age related risk of spontaneous abortion stratified according to parity and previous spontaneous abortions. RESULTS: Overall, 13.5% of the pregnancies intended to be carried to term ended with fetal loss. At age 42 years, more than half of such pregnancies resulted in fetal loss. The risk of a spontaneous abortion was 8.9% in women aged 20-24 years and 74.7% in those aged 45 years or more. High maternal age was a significant risk factor for spontaneous abortion irrespective of the number of previous miscarriages, parity, or calendar period. The risk of an ectopic pregnancy and stillbirth also increased with increasing maternal age. CONCLUSIONS: Fetal loss is high in women in their late 30s or older, irrespective of reproductive history. This should be taken into consideration in pregnancy planning and counselling. PMID- 10864551 TI - To whom it concerns PMID- 10864552 TI - Consumer health informatics. PMID- 10864553 TI - ABC of oral health. Dental damage, sequelae, and prevention. PMID- 10864554 TI - Seeing what you want to see in randomised controlled trials: versions and perversions of UKPDS data. United Kingdom prospective diabetes study. PMID- 10864555 TI - Practical partnerships for health and local authorities. PMID- 10864557 TI - Obituaries PMID- 10864556 TI - Benefits of a cooperative PMID- 10864558 TI - Local medical committee conference PMID- 10864559 TI - The BMJ, spin, and the stockmarket PMID- 10864560 TI - A register of blunders or botchers? PMID- 10864562 TI - Devaluing clinical skills PMID- 10864561 TI - Older parents PMID- 10864564 TI - Answers to big questions PMID- 10864563 TI - Cunegonde's ears, or multiple medical errors PMID- 10864566 TI - Expensive anthelmintic treatment to improve growth and cognitive performance is not warranted PMID- 10864565 TI - Many women with a normal cervix on colposcopy may be safely discharged from follow up PMID- 10864569 TI - Past infant mortality predicts current adult mortality from stomach cancer and stroke PMID- 10864567 TI - Troponin T can be used to detect missed heart attacks PMID- 10864568 TI - Fetal loss is high in women aged over 40 years PMID- 10864570 TI - Gene mutation found that might predispose to type 2 diabetes PMID- 10864571 TI - Official career guidance for specialist registrars PMID- 10864572 TI - Localization of the epithelial Ca(2+) channel in rabbit kidney and intestine. AB - The epithelial Ca(2+) channel (ECaC), which is exclusively expressed in 1,25 dihydroxyvitamin D(3)-responsive tissues, i.e., kidney, intestine, and placenta, is postulated to constitute the initial step in the process of transcellular Ca(2+) transport. To strengthen this postulated function, the present study compares the segmental and cellular distribution of ECaC and the other Ca(2+) transport proteins known to be involved in transcellular Ca(2+) transport. In rabbit kidney, ECaC mRNA and protein expression were primarily present in the connecting tubule. Immunopositive staining for the ECaC protein was exclusively found at the apical domain of this tubular segment. Importantly, ECaC completely colocalized with calbindin-D(28K), Na(+)-Ca(2+) exchanger (NCX), and plasma membrane Ca(2+) -ATPase (PMCA). A minority of cells along the distal tubule lacked immunopositive staining for ECaC and the other Ca(2+) transporting proteins. These negative cells were identified as intercalated cells. In intestine, ECaC was present in a thin layer along the apical membrane of the duodenal villus tip, whereas the crypt and goblet cells were negative. Again, a complete colocalization was observed between ECaC, calbindin-D(9K), and PMCA. In contrast to the kidney, NCX could not be detected in duodenum. The present finding that ECaC completely colocalizes with the Ca(2+) transport proteins in the connecting tubule and duodenum, together with its apical localization, further substantiates the postulated function of ECaC as the gatekeeper of active Ca(2+) (re)absorption. PMID- 10864573 TI - cAMP regulates cell proliferation and cyst formation in autosomal polycystic kidney disease cells. AB - Both epithelial cell proliferation and fluid accumulation are responsible for cyst growth in autosomal dominant polycystic kidney disease (ADPKD). It was previously reported that the cystic fibrosis transmembrane conductance regulator (CFTR) is expressed in cysts from ADPKD patients and suggested that cAMP stimulated Cl(-) and fluid secretion occurs through CFTR. The purpose of this study was to investigate the role of cell proliferation in cyst formation in ADPKD and to explore further the role of fluid secretion in cyst growth. Primary cultures both of ADPKD epithelial cells and a mixed population of normal renal epithelial cells isolated from the cortex (HRCE cells) were used. This study tested whether cAMP was involved both in stimulating cell proliferation and formation of cysts in vitro. (3)H-Thymidine incorporation assays showed that epidermal growth factor stimulated proliferation both in ADPKD cells and HRCE cells. In addition, cAMP stimulated DNA synthesis and cell proliferation in ADPKD, but not HRCE, cells. The effects of cAMP and epidermal growth factor on cell growth in ADPKD cells were additive. cAMP also stimulated cyst enlargement and fluid secretion in ADPKD cells. By contrast, cyst formation and enlargement from HRCE cells occurred without cAMP. Fluid secretion into the cyst lumen was blocked by diphenylamine carboxylic acid (DPC) and glibenclamide in ADPKD cells but blocked only by DPC in HRCE cells. This study showed that ADPKD cells have unique characteristics; cAMP stimulates fluid secretion and cell proliferation, indicating cAMP plays a very important role in cyst growth during the course of ADPKD. PMID- 10864574 TI - Differential effects of sera from normotensive and hypertensive pregnant women on Ca(2+) metabolism in normal vasular smooth muscle cells. AB - The clinical features of preeclampsia have been traditionally ascribed to a generalized vascular endothelial cell dysfunction. The present study investigates the effect of sera from preeclamptic women and normal pregnancy on the metabolism of intracellular Ca(2+) concentration ([Ca(2+)](i)) in normal cultured vascular smooth muscle cells (VSMC). Sera were obtained from normotensive pregnant women (NTP) (n = 17), preeclamptic women (PE) (n = 15), pregnant women with chronic (essential) hypertension (pregnant EHT) (n = 8), non-pregnant women with essential hypertension (non-pregnant EHT) (n = 12), and age-matched non-pregnant normotensive women (NNP) (n = 18). Serum (10%) was applied to both primary cultures of rat aortic smooth muscle cells and to the A-10 vascular muscle cell line. Levels of [Ca(2+)](i) were determined fluorometrically. After a 4-h incubation with serum, basal [Ca(2+)](i) was not significantly altered. However, compared with normal pregnant sera, PE sera markedly reduced hormonally induced Ca(2+) transients. Thus, following acute stimulation of rat VSMC (primary cultures) with 10(-8)M angiotensin II, peak [Ca(2+)](i) responses (% increment over baseline) were 443 +/- 22, 184 +/- 18, 259 +/- 12, 274 +/- 23, and 255 +/- 15% in NTP, PE, pregnant EHT, non-pregnant EHT, and NNP, respectively (P <0.01 PE versus NTP, P <0.05 PE versus NNP and pregnant and non-pregnant EHT). These effects of sera on [Ca(2+)](i) were qualitatively reproduced in platelets obtained from healthy volunteers. Also, depolarization-activated Ca(2+) influx in VSMC was affected by the different sera groups in a manner similar to that seen with hormonally induced [Ca(2+)](i) responses. The altered [Ca(2+)](i) changes by PE sera disappeared 5 wk after delivery. The effect of the different sera groups on hormonally triggered Ca(2+) transients in normal VSMC, as well as the normalization of [Ca(2+)](i) responses after delivery, suggest the presence of a circulating serum factor in PE. Inasmuch as [Ca(2+)](i) is the major determinant of VSMC tone, it is possible that consequent to the attenuation of [Ca(2+)](i) responses, this putative circulating factor counterbalances the intense vasoconstriction in PE. PMID- 10864575 TI - Exaggerated impact of ATP-sensitive K(+) channels on afferent arteriolar diameter in diabetes mellitus. AB - Experiments were performed to determine the involvement of ATP-sensitive K(+) channels (K(ATP) channels) in the renal afferent arteriolar dilation that occurs during the hyperfiltration stage of insulin-dependent diabetes mellitus (IDDM). IDDM was induced in rats by streptozotocin (STZ) injection, and adequate insulin was provided to maintain moderate hyperglycemia. Sham rats received vehicle treatments. Two weeks later, afferent arteriolar function was assessed using the in vitro blood-perfused juxtamedullary nephron technique. Baseline afferent arteriolar lumen diameter was greater in STZ rats (25.9 +/- 1.1 microm) than in sham rats (20.8 +/- 1.0 microm). Glibenclamide (3 to 300 microM) had virtually no effect on afferent arterioles from sham rats; however, this K(ATP) antagonist caused concentration-dependent afferent arteriolar constriction in kidneys from STZ-treated rats, restoring lumen diameter to 20.6 +/- 1.7 microm (P > 0.05 versus sham baseline). In both groups of rats, pinacidil (a cyanoguanidine K(ATP) agonist; 0.3 to 300 microM) evoked concentration-dependent afferent arteriolar dilation, indicating the functional expression of K(ATP) channels; however, lumen diameter was increased by 73% in STZ kidneys but only by 48% in sham kidneys. The gliben-clamide-sensitive afferent arteriolar dilator response to 1 microM PCO-400 (a benzopyran K(ATP) agonist) was also accentuated in STZ kidneys. These observations suggest that increases in both the functional availability and basal activation of K(ATP) channels promote afferent arteriolar vasodilation during the early stage of IDDM, changes that likely contribute to the etiology of diabetic hyperfiltration. PMID- 10864576 TI - Bradykinin B(1) receptor-mediated changes in renal hemodynamics during endotoxin induced inflammation. AB - Kinins have been shown to influence renal hemodynamics and function. Under physiologic conditions, most kinin effects involve bradykinin B(2) receptors, but bradykinin B(1) receptors are often induced during inflammation. The purpose of this study was to examine in vivo the effects of bradykinin B(1) receptor activation on renal hemodynamics under normal and inflammatory conditions. In anesthetized rats, activation of bradykinin B(1) receptors by arterial infusion of bradykinin B(1) receptor agonist des-Arg(9)-bradykinin reduced renal plasma flow and GFR. Prior administration (18 h) of lipopolysaccharide to induce inflammation resulted in a larger bradykinin B(1) receptor-induced reduction in renal plasma flow. Values of other parameters remained unchanged, thus resulting in an increased filtration fraction. The presence and the functionality of the bradykinin B(1) receptor at the level of glomerular afferent and efferent arterioles were studied by mRNA expression analysis and intracellular calcium ([Ca(2+)](i)) mobilization studies. Stimulation with des-Arg(9)-bradykinin of microdissected afferent arterioles from control and lipopolysaccharide-treated rats induced [Ca(2+)](i) mobilization without any significant difference in amplitude between control and lipopolysaccharidetreated rats. However, des-Arg(9) bradykinin only induced [Ca(2+)](i) mobilization in efferent arterioles from lipopolysaccharide-treated rats. It is suggested that activation of bradykinin B(1) receptors located along the efferent arteriole may participate in the modification of renal hemodynamics in inflammatory states. PMID- 10864577 TI - The affinity of the organic cation transporter rOCT1 is increased by protein kinase C-dependent phosphorylation. AB - Members of the organic cation transporter (OCT) family are mainly expressed in kidney, liver, intestine, and brain. The regulation of the OCT type 1 from rat (rOCT1) stably transfected in HEK293 cells was examined using a fluorimetric technique, 1-[(3)H]methyl-4-phenylpyridinium uptake studies, and fast-whole-cell patch-clamp recordings. For the fluorescence measurements, the cation 4-(4 (dimethylamino)styryl)-N-methylpyridinium iodide (ASP(+)) was used as substrate. Uptake of ASP(+) via rOCT1 was electrogenic, and its inhibition by other organic cations was consistent with previously reported radioactive tracer flux measurements. The inhibitor quinine was not translocated by the organic cation transporter in contrast to tetraethylammonium. Stimulation of diacyl glycerol dependent protein kinase C (PKC) by sn-1,2-dioctanoyl glycerol (1 microM) resulted in an increase in initial ASP(+) uptake rate by 216 +/- 28% (n = 29). The effect was completely antagonized by the PKC inhibitor tamoxifen (20 microM, n = 22). Forskolin (1 microM), which activates adenylate cyclase and thereby protein kinase A (PKA), stimulated the initial rate of ASP(+) accumulation by 51 +/- 6% (n = 19). This effect was inhibited by the specific PKA inhibitor KT5720 (1 microM, n = 12). Inhibition of tyrosine kinases by aminogenestein (10 microM) reduced ASP(+) uptake by 63 +/- 7% (n = 7), while genestein or tyrphostin AG1295 (each 10 microM) were without significant effects. Incubation of the cells with sn-1, 2-dioctanoyl glycerol (1 microM) increased the affinities of the transporter to tetraethylammonium, tetrapenthylammonium, and quinine by a factor of 58, 14.5, and 2.4, respectively. Western blot analysis revealed that rOCT1 protein was phosphorylated at a serine residue upon stimulation of PKC. In conclusion, it has been demonstrated that the organic cation transport by rOCT1 is stimulated by PKC, PKA, and endogenous tyrosine kinase activation. The PKC phosphorylates rOCT1 and leads to a conformational change at the substrate binding site. PMID- 10864578 TI - Effect of elevated glucose on endothelin-induced store-operated and non-store operated calcium influx in renal mesangial cells. AB - Early diabetic nephropathy exhibits renal glomerular hyperfiltration and an increase in renal plasma flow. The hyperfiltration is a dysfunctional state that may arise from a hyperglycemic-induced hypocontractility of glomerular mesangial cells that may be associated with depressed Ca(2+) signaling events. The present study was designed to determine the effects of acute (minutes) and chronic (days) elevated glucose levels on endothelin-induced calcium signaling with a particular emphasis on the potential influence on stores and store-operated Ca(2+) influx (SOCI; also called capacitative calcium entry) in glomerular mesangial cells. Primary cultures of rat mesangial cells were grown in either high (30 mM) or normal (5 mM) glucose-containing media and tested in the presence of either high (30 mM) or normal (5 mM) glucose levels. Intracellular calcium levels were monitored with the calcium-sensitive fluorophore fura-2 before and after treatment with either endothelin-1 (10 nM), to induce typical Ca(2+) signals, or the endoplasmic reticulum (ER) Ca-ATPase inhibitor thapsagargin (1 microM), to unload ER Ca(2+) stores. Both acute and chronic exposure to high glucose levels depressed the endothelin-induced calcium signal. However, neither release of Ca(2+) from stores nor SOCI were depressed by high glucose levels. In contrast, an endothelin-induced calcium entry pathway (likely receptor-operated calcium influx), separate from SOCI, was markedly depressed in the presence of both acute and chronic high glucose levels. The depressant effect of high glucose was rapidly (minutes) reversible upon returning to normal glucose levels. It is concluded that high glucose levels depress endothelin-induced calcium signaling in rat mesangial cells by inhibiting non-SOCI Ca(2+) entry pathways, namely the receptor-operated Ca(2+) influx pathway. The glucose-induced alterations in the receptor-operated calcium influx pathway may, in part, contribute to the depressed contractile state of glomerular cells during periods of hyperglycemia. PMID- 10864579 TI - Vascular endothelial growth factor receptors in human mesangium in vitro and in glomerular disease. AB - Mesangial cell proliferation and growth factor over-expression are characteristic features of several glomerular diseases. Vascular endothelial growth factor (VEGF), a potent mitogen, is expressed in podocytes in the glomerulus, and VEGF receptors (flt-1, KDR, and neuropilin-1) are present on endothelial cells and other cell types. This study examined whether human mesangial cells (HMC) express VEGF receptors in vitro and ex vivo and evaluated the effect of VEGF on HMC proliferation. All receptor types were detected in HMC in vitro by immunofluorescence and Western blotting. VEGF(165) induced a dose-responsive increase in (3)H-thymidine incorporation (25 ng/ml VEGF(165) : 2.3-fold increase; 50 ng/ml : 3.8-fold; 100 ng/ml : 4. 8-fold; 200 ng/ml : 3.4-fold; P = 0.016) and in cell number (50 ng/ml VEGF(165) : 1.2-fold increase; 100 ng/ml : 1.6-fold; 200 ng/ml : 1.4-fold; P = 0.005), effects prevented by an anti-VEGF(165) polyclonal neutralizing antibody (100 microg/ml). The proliferative effect was confirmed by a tetrazolium dye-based assay (100 ng/ml VEGF(165) : 1.4-fold increase). In ex vivo experiments, VEGF receptors in biopsy material from normal and diseased kidneys were detected by immunohistochemistry. No mesangial flt-1 receptor staining was seen in normal renal cortical tissue samples, and only weak mesangial KDR staining was detected. In contrast, mesangial flt-1 and KDR receptor staining were both clearly seen in biopsy samples from proliferative renal diseases. In conclusion, flt-1, KDR, and neuropilin-1 are present on cultured HMC, and VEGF(165) induces HMC proliferation. In addition, the flt-1 and KDR receptors are expressed in the mesangium in mesangioproliferative disease. PMID- 10864580 TI - In vivo administration of a nuclear transcription factor-kappaB decoy suppresses experimental crescentic glomerulonephritis. AB - Glomerular expression of cytokines, interleukin-1 (IL-1), and tumor necrosis factor-alpha (TNF-alpha), together with leukocytic infiltration, are prominent features in crescentic glomerulonephritis. Because these cytokines are targets for nuclear transcription factor-kappaB (NF-kappaB), the use of NF-kappaB decoy oligodeoxynucleotide (ODN) treatment was evaluated in an experimental disease model. Crescentic glomerulonephritis was induced in primed Wistar rats by injection of sheep antiglomerular basement membrane serum. Thirty minutes after injection, rats were anesthetized and the left kidney was perfused with NF-kappaB decoy ODN or scrambled ODN control mixed with a virus-liposome complex, and then killed 7 d later. Animals given the scrambled control ODN developed severe glomerulonephritis by day 7 with heavy proteinuria, glomerular crescents and interstitial lesions, marked leukocytic infiltration, and upregulated renal expression of cytokines (IL-1 and TNF-alpha) and adhesion molecules (intercellular adhesion molecule-1). In contrast, NF-kappaB decoy ODN treatment substantially inhibited the disease with a 50% reduction in proteinuria, a threefold reduction in histologic damage, a 50% reduction in leukocytic infiltration, and a 50 to 80% reduction in the renal expression of cytokines and leukocyte adhesion molecules. In conclusion, this study has demonstrated that NF kappaB plays a key role in cytokine-mediated renal injury and that NF-kappaB decoy ODN treatment has clear therapeutic potential in rapidly progressive glomerulonephritis. PMID- 10864581 TI - Quantitative trait loci modulate renal cystic disease severity in the mouse bpk model. AB - Numerous mouse models of polycystic kidney disease (PKD) have been described in which the mutant phenotypes closely resemble human PKD with regard to morphology, cyst localization, and disease progression. As in human PKD, genetic background affects the disease phenotype in mouse PKD models. Using experimental crosses, these modifying effects can be dissected into discrete genetic factors referred to as quantitative trait loci. The locus for the mouse bpk model was recently mapped to chromosome (Chr) 10. In the course of these studies, marked variability was observed in the renal cystic disease expressed in F2 bpk/bpk homozygotes of a (BALB/c-+/bpk x CAST/Ei)F1 intercross. The current study was undertaken to further characterize the renal cystic disease as quantitative trait in this F2 cohort and to map the genetic modifiers that modulate this phenotype. Whole genome scans revealed a CAST-derived locus on distal Chr 6, near D6Mit14, that affects renal cystic disease severity. Additional analyses identified loci on Chr 1, Chr 2, and Chr 4, as well as a possible interaction between the Chr 6 locus and a locus on distal Chr 1, near D1Mit17. Interestingly, the gene encoding RGS7, a regulator of G protein signaling that binds to polycystin-1, was mapped to the same Chr 1 interval. It is concluded that the severity of the bpk renal cystic disease phenotype is modulated by multiple loci and possibly by epistatic interaction among them. It is hypothesized that the gene encoding the polycystin binding partner RGS7 is a candidate for the Chr 1 genetic modifier. PMID- 10864582 TI - Chromosomal mapping of a major quantitative trait locus regulating compensatory renal growth in the rat. AB - Despite extensive research conducted over the past century, the mechanisms of compensatory renal growth (CRG) remain a mystery. Insight into the mechanisms that regulate CRG might be gained by identifying genetic factors that influence this complex phenotype. In a large set of recombinant inbred strains derived from the spontaneously hypertensive rat and the Brown Norway rat, a genome scan for quantitative trait loci (QTL) that regulate CRG was performed. The CRG score was expressed as a ratio of the weight of the remnant right kidney at 8 wk of age to the weight of the left kidney at 5 wk of age, both adjusted for body weight. QTL mapping was performed using Map Manager QT and the strain distribution patterns of more than 600 genetic markers. It was found that CRG after unilateral nephrectomy is a multifactorially determined trait with a substantial genetic component. The heritability of CRG approached 40%. Genome wide scan analysis revealed significant evidence of linkage to a region of rat chromosome 4 designated Crg 1 that accounted for more than 50% of the additive genetic variance of CRG in the recombinant inbred strains. The detection of a major QTL influencing CRG in the rat should provide new opportunities for identifying mechanisms that regulate this historically enigmatic phenomenon and may also have implications for research on the pathogenesis of end-stage kidney disease. PMID- 10864583 TI - Proapoptotic Fas ligand is expressed by normal kidney tubular epithelium and injured glomeruli. AB - Fas ligand (FasL) is a cell membrane cytokine that can promote apoptosis through activation of Fas receptors. Fas receptor activation induces glomerular cell apoptosis in vivo and participates in tubular cell death during acute renal failure. However, there is little information on the expression of FasL in the kidney. This study reports that FasL mRNA and protein are present in normal mouse and rat kidney. In situ hybridization and immunohistochemistry showed that proximal tubular epithelium is the main site of FasL expression in the normal kidney. In addition, increased total kidney FasL mRNA and de novo FasL protein expression by glomerular cells were observed in two different models of glomerular injury : rat immune-complex proliferative glumerulonephritis and murine lupus nephritis. Both full-length and soluble FasL were increased in the kidneys of the mice with nephritis. Cultured murine proximal tubular epithelial MCT cells and primary cultures of murine tubular epithelial cells expressed FasL mRNA and protein. Tubular epithelium-derived FasL induced apoptosis in Fassensitive lymphoid cell lines but not in Fas-resistant lymphoid cell lines. By contrast, MCT cells grown in the presence of the survival factors of serum were resistant to FasL, and only became partially sensitive to apoptosis induced by high concentrations (100 ng/ml) of FasL upon serum deprivation. However, MCT cells stimulated with inflammatory mediators (tumor necrosis factor-alpha, interferon-gamma, and lipopolysaccharide) increased cell surface Fas expression and were sensitized to apoptosis induced by FasL (FasL 55 +/- 5% versus control 8.3 +/- 4.1% apoptotic cells at 24 h, P < 0.05). Cytokine-primed primary cultures of tubular epithelial cells also acquired sensitivity to FasL-induced apoptosis. These results suggest that FasL expression by intrinsic renal cells may play a role in cell homeostasis in the normal kidney and during renal injury. PMID- 10864584 TI - Contribution of angiotensin II to late renal injury after acute ischemia. AB - Rats recovering from acute renal ischemia exhibit tubule loss and interstitial fibrosis followed by development of proteinuria and glomerular sclerosis. The current study assessed the contribution of angiotensin II (AngII) to these processes. The contribution of AngII to early tubule loss and interstitial fibrosis was assessed in rats studied for 35 d after right nephrectomy and transient occlusion of the left renal artery. One group of rats received no treatment, while a second group received losartan beginning at 2 d following ischemia. Studies at 35 d showed that losartan did not improve GFR (2.04 +/- 0.30 ml/min treated, 2.16 +/- 0.21 ml/min untreated), reduce the fraction of glomeruli that were no longer connected to normal tubule segments (42 +/- 9% treated, 42 +/ 13% untreated), or limit expansion of the interstitial volume fraction (25 +/- 3% treated, 25 +/- 4% untreated). The contribution of AngII to progressive glomerular injury following initial recovery from ischemia was assessed in similarly prepared rats studied for 140 d. One group of rats received no treatment, while a second group received enalapril beginning at 35 d following ischemia. Enalapril markedly reduced proteinuria (78 +/- 17 mg/d treated, 229 +/- 52 mg/d untreated) and the prevalence of segmental glomerular sclerosis (14 +/- 9% treated, 45 +/- 10% untreated). Untreated rats developed sclerotic lesions in glomeruli not connected to normal tubules, as well as in glomeruli connected to normal tubules. Enalapril prevented injury in both classes of glomeruli. These results indicate that AngII does not contribute to interstitial fibrosis during recovery from ischemic injury. Reduction of AngII activity, can, however, prevent secondary glomerular injury in kidneys initially damaged by ischemia. PMID- 10864585 TI - Renal enlargement precedes renal hyperfiltration in early experimental diabetes in rats. AB - The order of appearance between renal hypertrophy and hyperfunction in early experimental diabetes is still disputed. The reason for previous discrepant results is believed to be methodologic problems, as most previous studies of renal function have been performed in anesthetized animals. In the present study in nondiabetic and streptozotocin-diabetic animals, renal volume was measured by a noninvasive magnetic resonance imaging technique, while renal function parameters were measured in conscious, chronically catheterized animals. To avoid artifacts caused by the procedures associated with induction of streptozotocin diabetes (fasting, brief anesthesia, and transient hypoglycemia) on renal growth and function, diabetic animals were injected with insulin to obtain euglycemia for 4 d before study start. At day 0, insulin administration was withdrawn and all animals developed hyperglycemia within 12 h. Renal volume and kidney function were measured on days 0, 1, 2, 3, 5, and 7. Renal enlargement was detectable at day 1 (20%) and reached an increase of 40% at day 7. No changes were seen in effective renal plasma flow or effective renal vascular resistance within the first 7 d after development of hyperglycemia. GFR tended to rise on day 5 and was increased by 16% at day 7. The absolute proximal reabsorption showed a pronounced rise (30%) at day 7, whereas no change was seen in the proximal tubular fluid output. It is concluded that renal enlargement precedes renal hyperfunction in the early phase after onset of experimental diabetes. PMID- 10864586 TI - Nitric oxide synthesis inhibition does not impair water immersion-induced renal vasodilation in humans. AB - Nitric oxide (NO) is tonically released in the kidney to maintain renal perfusion and adequate sodium and water clearance. Little is known about the role of NO in the renal adaptation to an acute volume challenge. This is important for our understanding of pathophysiologic conditions associated with impaired NO activity. This study examined the effects of NO synthesis inhibition on neurohumoral, renal hemodynamic, and excretory responses to head-out immersion (HOI). Seven healthy men underwent four 7-h clearance studies. One study served as a time control study (placebo infusion), and in one study N(G)-monomethyl-L arginine (L-NMMA; 3 mg/kg priming dose + 3 mg/kg per h) was infused during hours 2 to 5. In a third and fourth clearance study, HOI was applied from hours 3 to 5, during infusion of either placebo or L-NMMA. To assess the degree of NO synthesis inhibition, the effect of L-NMMA on [(15)N]-arginine-to-[(15)N]-citrulline conversion rate was studied in four others. HOI decreased mean arterial pressure (MAP) from 87 +/- 3 to 76 +/- 2 mmHg and renal vascular resistance (RVR) from 82 +/- 6 to 70 +/- 7 mmHg. min/L, and increased sodium excretion (UNaV) from 110 +/- 27 to 195 +/- 29 micromol/min and flow (UV) from 14.4 +/- 1.4 to 15.8 +/- 1.4 ml/min. L-NMMA caused profound and sustained increases in MAP and RVR and decreases in UNaV and UV. HOI superimposed on L-NMMA infusion decreased the elevated MAP from 93 +/- 4 to 83 +/- 2 mmHg and RVR from 111 +/- 9 to 95 +/- 7 mmHg. min/L, and increased UNaV from 41 +/- 8 to 95 +/- 15 micromol/min and UV from 10.0 +/- 1.1 to 12.7 +/- 1.4 ml/min. The relative changes were not significantly different from the effects of HOI without L-NMMA infusion. HOI decreased plasma renin activity and aldosterone and increased plasma atrial natriuretic peptide and urinary cGMP. L-NMMA decreased urinary cGMP, but did not affect the plasma hormones or the changes induced by HOI. L-NMMA decreased the [(15)N]-arginine-to-[(15)N]-citrulline conversion rate to one-third of baseline. The results indicate that in a state of NO deficiency in humans, the kidney can still respond to an acute volume challenge with vasorelaxation, diuresis, and natriuresis. PMID- 10864587 TI - Increases in serum leptin levels during peritoneal dialysis are associated with inflammation and a decrease in lean body mass. AB - Leptin, secreted from fat cells, functions as a lipostat mechanism through modulation of satiety signals. Markedly elevated leptin levels have been documented in uremic patients, especially in those who are treated by peritoneal dialysis (PD). However, the role of hyperleptinemia in uremic patients is not clear, and it is not known whether elevated leptin levels contribute to uremic anorexia and changes in body composition. In this prospective study, serum leptin, C-reactive protein (CRP), plasma insulin, and body composition (dual energy x-ray absorptiometry) were measured in 36 patients (53 +/- 1 yr) close to start and after about 1 yr of PD. In addition, markers of dialysis adequacy and urea kinetics were followed during treatment with PD. During PD, the total body fat mass (20.5 +/- 1.0 to 22.9 +/- 1.3 kg; P < 0.01), truncal fat mass (11.5 +/- 0.7 to 13. 2 +/- 0.9 kg; P < 0.001), and serum leptin levels (20.1 +/- 3.8 to 35.6 +/- 6.8 ng/ml; P < 0.01) increased markedly, especially in patients with diabetes mellitus. Twenty-five PD patients that lost lean body mass during PD had significantly (P < 0.05) elevated initial CRP levels (14 +/- 2 mg/L) compared to 11 patients (<10 mg/L) who gained lean body mass during PD. A significant increase in serum leptin levels (20.9 +/- 4.2 to 42.7 +/- 4.0 ng/ml; P < 0.001) was observed in those patients who lost lean body mass, whereas no such change (18.4 +/- 8.4 to 19.2 +/- 6.4 ng/ml) was observed in the patients that gained lean body mass during PD treatment. The present longitudinal results demonstrate that serum leptin level and body fat content increase markedly during PD, especially in diabetic patients. Patients that lost lean body mass during PD had higher initial CRP levels and increased their serum leptin levels significantly during PD compared to those patients that gained lean body mass. Additional studies are therefore needed to elucidate the role of hyperleptinemia and inflammation in causing anorexia, protein-malnutrition, and changes in body composition during treatment with PD. PMID- 10864588 TI - Pharmacokinetics of intermittent intravenous cefazolin and tobramycin in patients treated with automated peritoneal dialysis. AB - There is increasing use of intermittent dosing of antibiotics to treat peritoneal dialysis (PD)-related peritonitis. The disposition of intravenous cefazolin and tobramycin was studied in automated PD (APD) patients. Ten patients were recruited and received a single intravenous dose of cefazolin (15 mg/kg) and tobramycin (0.6 mg/kg). Blood and dialysate samples were collected at the beginning, middle, and end of dwells 1 to 3 (on cycler), and at the end of dwells 4 to 5 (off cycler) for a 24-h period. Baseline and 24-h urine samples were collected. Pharmacokinetic parameters were calculated using a monoexponential model. Cefazolin and tobramycin half-lives were markedly different on cycler than off cycler (cefazolin on cycler : 10.67 +/- 4.66 h; cefazolin off cycler : 23.09 +/- 5.6 h; P = 0.001; tobramycin on cycler : 14.27 +/- 4.53 h; tobramycin off cycler : 68. 5 +/- 26.47 h; P < 0.001). Mean serum and dialysate concentrations were above minimum inhibitory concentrations of susceptible organisms throughout the 24-h period for both drugs with intravenous administration. A model was developed to examine serum and dialysate concentrations after intermittent intraperitoneal administration of 15 mg/kg cefazolin and 0.6 mg/kg tobramycin. Model-predicted intraperitoneal cefazolin provides adequate serum and dialysate concentrations for 24 h. Intermittent intraperitoneal tobramycin doses must be 1.5 mg/kg for one exchange during the first day and then given as 0.5 mg/kg thereafter. It is concluded that the current empiric dosing recommendations for PD-related peritonitis may be adequate for cefazolin (15 to 20 mg/kg); however, tobramycin doses must be changed to 1.5 mg/kg intraperitoneally on day 1, then to 0.5 mg/kg intraperitoneally thereafter in APD patients. PMID- 10864589 TI - Contribution of prolonged ischemia and donor age to chronic renal allograft dysfunction. AB - As a consequence of an advancing discrepancy between supply of suitable grafts and demand from potential recipients, less than optimal organs are increasingly being used. Although clinical studies demonstrate the involvement of various risk factors, including donor age and duration of ischemia on long-term graft outcome, their individual contribution and correlation has not been followed experimentally. After cold ischemic times of 5, 60, and 120 min, kidney allografts of 3-, 12-, and 18-mo-old Fischer 344 donors were transplanted into 3 mo-old Lewis rats. Age-related changes were examined in matched native uninephrectomized controls. Proteinuria and creatinine clearance were determined, and histologic and immunohistologic studies were assessed and quantified at the end of the observation period (20 wk). All grafts functioned satisfactorily with the exception of one graft each from 12- and 18-mo-old donors with prolonged ischemia (120 min). Functional deterioration and structural changes progressed in parallel to increasing donor age and prolonged ischemia. The impact of expanded ischemia was particularly detrimental in grafts from older donor animals. Donor age and duration of ischemia act in a synergistic manner in our model. Brief ischemic times seem of particular relevance when grafts from older donors are being used. PMID- 10864590 TI - Minimizing hemodialysis vascular access trauma with an improved needle design. AB - The maintenance and longevity of hemodialysis vascular access remains one of the most problematic topics in the care of dialysis patients. Although much attention has focused on neointimal hyperplasia, the repetitive trauma to vessel walls by dialysis needles causes significant cumulative damage that has undergone little investigation. Commercial needles have beveled tips with intentional cutting surfaces to ease insertion. It was hypothesized that a pencil-point conical shaped needle would cause less damage by taking advantage of the elasticity of native fistulae and produce an improved hole configuration in synthetic materials with minimal ability to stretch. A needle was subsequently designed with a removable pencil-point trocar and a side arm for the dialysis tubing. Once the trocar is removed, the blunt-ended cannula can be advanced or can be subject to inadvertent motion without causing damage to the luminal surface of the access. The new design as well as standard 15-gauge hemodialysis needles were tested on Gore-Tex graft material and two bovine carotid artery preparations. Scanning electron microscopy was used to study the hole patterns. For all materials, the commercial needle holes had typical crescent shapes, and the cuts sliced sequentially through the various layers. For grafts, the new design caused a linear defect parallel to the axis of the graft that may preserve longitudinal strength. Interestingly, that tear line was nearly perpendicular to the linear hole in the thin polytetrafluoroethylene overwrap, which would be consistent with maintenance of hoop integrity. It is believed that these nonoverlapping defects would also improve hemostasis. The bovine specimens tested the importance of tissue stretching : Fresh carotid artery had experimental holes dramatically smaller than those from standard needles. In the denatured tissue, the experimental needle provided less benefit than that observed in fresh tissue, which is likely due to limited elasticity of the preserved artery. Improvement in needle design thus provides distinct advantages for native vessels and unique less traumatic holes in current synthetic materials. Pencil-point needle designs may be particularly applicable to the development of new elastomeric graft material. PMID- 10864591 TI - From tonus to tonicity: physiology of CLC chloride channels. AB - Chloride channels are involved in a multitude of physiologic processes ranging from basal cellular functions such as cell volume regulation and acidification of intracellular vesicles to more specialized mechanisms such as vectorial transepithelial transport and regulation of cellular excitability. This plethora of functions is accomplished by numerous functionally highly diverse chloride channels that are only partially identified at the molecular level. The CLC family of chloride channels comprises at present nine members in mammals that differ with respect to biophysical properties, cellular compartmentalization, and tissue distribution. Their common structural features include a predicted topology model with 10 to 12 transmembrane regions together with two C-terminal CBS domains. Loss of function mutations affecting three different members of the CLC channel family lead to three human inherited diseases : myotonia congenita, Dent's disease, and Bartter's syndrome. These diseases, together with the diabetes insipidus symptoms of a knockout mouse model, emphasize the physiologic relevance of this ion channel family. PMID- 10864592 TI - Clinical practice guideline on shared decision-making in the appropriate initiation of and withdrawal from dialysis. The Renal Physicians Association and the American Society of Nephrology. PMID- 10864593 TI - Transplantation in the patient with hepatitis C. PMID- 10864594 TI - The psychoses of epilepsy. PMID- 10864595 TI - Isolated Horner's syndrome and syringomyelia. PMID- 10864596 TI - Inborn errors of metabolism as a cause of neurological disease in adults: an approach to investigation. PMID- 10864597 TI - Social deprivation and prevalence of epilepsy and associated health usage. AB - OBJECTIVES: To examine the relation between social deprivation and the prevalence of epilepsy and associated morbidity using hospital activity data as a proxy. METHODS: The study was conducted in the health district of South Glamorgan, United Kingdom (population 434 000). Routinely available hospital data (inpatient and outpatient), an epilepsy clinic database, and mortality data underwent a process of record linkage to identify records relating to the same patient and to identify patients with epilepsy. Each patient was allocated a Townsend index deprivation score on the basis of their ward of residence. Age standardised correlations were calculated between deprivation score and prevalence of epilepsy, inpatient admissions, and outpatient appointments. Standardised mortality ratios (SMR) were also calculated. All analyses were performed on two cohorts: (1) all patients with epilepsy and (2) those patients with epilepsy without any underlying psychiatric illness or learning disability. RESULTS: The prevalence of epilepsy ranged between 2.0 and 13.4 per 1000 with a median of 6.7. There were positive correlations between social deprivation and prevalence in both populations: (1) r=0.75 (p<0.001) and (2) r=0.70 (p<0.001). After standardising for underlying prevalence there were also correlations for mean inpatient admissions: (1) r=0.62 (p<0.001), (2) r=0.59, (p<0.001) and for outpatient appointments: (1) r=0.53, (p=0.001) and (2) r=0. 51 (p=0.001). The SMR for those deprived was (1) 1.66 (95% confidence interval (95% CI) 1.27-2.05) and (2) 1.80 (95% CI 0.71-1. 67). For the population as a whole (with and without epilepsy) the SMR was 1.25 (95% CI 1.27-2.32). CONCLUSION: This study shows a strong correlation between the prevalence of epilepsy and social deprivation and weaker correlations between social deprivation and mean hospital activity. PMID- 10864598 TI - Inhibitory simple partial (non-convulsive) status epilepticus after intracranial surgery. AB - OBJECTIVES: To report on five patients who developed, 2 to 4 days after an intracranial neurosurgical procedure, new, persistent, focal neurological deficits which were due to inhibitory simple partial (non-convulsive) status epilepticus, and resolved with anticonvulsant treatment. METHODS: The age range of the five patients was 15-74 years. The operations were: aneurysm clipping (three patients) and resections of an oligodendroglioma and a cavernous haemangioma (one patient each). The new focal deficits were: right hemiparesis and aphasia (two patients), aphasia alone (two patients), and left hemiparesis (one patient). The deficits were not explained by CT (obtained in all patients) or cerebral angiography (performed in two). RESULTS: Electroencephalography showed, in all patients, continuous or intermittent focal seizures arising from cortex regionally relevant to the clinical dysfunction. Subtle positive epileptic phenomena (jerking) occurred intermittently in three patients as a late concommitant. Administration of anticonvulsant drugs resulted in significant improvement within 24 hours in four patients, with parallel resolution of ictal EEG activity. The fifth patient improved more slowly. Two patients relapsed when anticonvulsant concentrations fell, and improved again when they were raised. CONCLUSIONS: It is suggested that inhibitory simple partial (non-convulsive) status epilepticus be considered in the differential diagnosis when a new unexplained neurological deficit develops after an intracranial neurosurgical procedure. An EEG may help to diagnose this condition, leading to definitive treatment. PMID- 10864599 TI - Apolipoprotein E genotype related differences in brain lesions of multiple sclerosis. AB - OBJECTIVES: Clinical reports have speculated on a more severe course of multiple sclerosis in patients with the apolipoprotein E (apoE) epsilon4 allele. As this could be reflected by differences in the severity of tissue damage MRI was used to obtain further support for a disease modifying effect of the apoE genotype. METHODS: Brain MR scans of 83 patients (mean age 35.5 (SD 9.5 years) who participated in a cross sectional study on the distribution of genotype patterns in multiple sclerosis. The total lesion load on proton density weighted (T2-LL) and T1 weighted scans (T1-LL) obtained with conventional spin echo sequences at 1.5 T was measured. A "black hole" ratio ((T1-LL/T2-LL)x100) was also calculated. This indicates the proportion of multiple sclerosis lesions with more severe tissue damage and may reflect disease aggressiveness or quality of repair. RESULTS: Patients with the apoE-epsilon3/epsilon4 genotype (n=19) showed a non significantly greater T2-LL (16.0 (SD 14.0) cm(3)) than patients with the epsilon2/epsilon3 (n=11; 13.3 (9.5) cm(3)) or the epsilon3/epsilon3 genotype (n=49; 9.4 (SD 9.2) cm(3)). Both the T1-LL (2.6 (SD 3.3) v 1.6 (SD 2.4) and 1.2 (SD 3.0) cm(3); p=0.04) and the black hole ratio (14.3 SD 11.9) v 7.4 (SD 9.3) and 8.4 (SD 13.3)%; p=0.02), however, were significantly higher in epsilon3/epsilon4 patients. Similar differences were seen when comparing patients with at least one epsilon4 allele with the remainder of the group. CONCLUSIONS: These data support speculations on a modulation of multiple sclerosis severity by the apoE genotype which can be attributed to more extensive tissue destruction or less efficient repair in carriers of the epsilon4 allele. PMID- 10864600 TI - Serum lipids in young patients with ischaemic stroke: a case-control study. AB - OBJECTIVES: The relation between serum lipids and ischaemic stroke remains controversial. Studies of lipid related risk factors in cerebrovascular disease have varied greatly in their findings and also in their definition of the cerebrovascular end points. Serum lipids are thought to interact with the pathogenesis of stroke through an atherosclerosis mechanism. Stroke in young patients have been shown to be related to non-atherosclerotic causes most of the time. The aim was to determine the serum lipid profile and the vascular risk factors for ischaemic stroke in a series of patients under 45 with an ischaemic stroke and to compare them with a series of controls of the same age. METHODS: Ninety four consecutive patients with ischaemic stroke were compared with 111 controls of the same age recruited from a regional electoral list. Vascular risk factors were recorded and serum lipid profile was determined in all of them. RESULTS: Multivariate analyses showed that low HDL cholesterol, male sex, smoking, hypertension, and oral contraceptives were risk factors for intracerebral arterial occlusion. CONCLUSION: Low HDL cholesterol was the only serum lipid index to be associated to an increased risk of stroke in this population. Low HDL cholesterol must be considered in the care management of young patients regardless of the detectable presence of atherosclerosis. PMID- 10864601 TI - Contribution of thixotropy, spasticity, and contracture to ankle stiffness after stroke. AB - OBJECTIVES: Increased resistance to stretch of muscles after stroke may be the result of centrally mediated neural factors such as spasticity or local, peripheral factors such as muscle contracture or thixotropy. The aim was to investigate evidence for an abnormal thixotropic response and compare this with two other factors-contracture and spasticity-which could potentially contribute to muscle stiffness after stroke. METHODS: Thirty patients with stroke whose calf muscles were assessed clinically as stiff and 10 neurologically normal subjects were recruited. To measure thixotropy, their calf muscles were stretched through two cycles after two prestretch conditions: one in which the muscles were maintained in a shortened position and one in which they were maintained in a lengthened position. Spasticity was defined as the presence of tonic stretch reflexes in relaxed muscles. Contracture was defined as being present when maximum passive ankle dorsiflexion fell at least 2 SD below the mean value of the control subjects. RESULTS: Both controls and patients with stroke exhibited a thixotropic response but this was no greater in the patients than the controls. About one third of the patients displayed muscle contracture and most exhibited spasticity. Contracture made a significant contribution (p=0.006) to the clinical measure of calf muscle stiffness while spasticity made a significant contribution (p=0.004) to the laboratory measure of calf muscle stiffness. CONCLUSIONS: Measuring thixotropy at the level of joint movement was sufficiently sensitive to determine the thixotropic response in both neurologically normal subjects and patients impaired after stroke. The thixotropic response was not higher than normal after stroke, suggesting that whereas thixotropy may produce enough immediate resistance to impede movement in those who are very weak, it is not a substantial contributor to long term muscle stiffness. Contracture did significantly contribute to muscle stiffness, supporting the importance of prevention of contracture after stroke. Spasticity contributed to muscle stiffness only when the limb was moved quickly. PMID- 10864602 TI - Prospective analysis of bedside percutaneous subdural tapping for the treatment of chronic subdural haematoma in adults. AB - OBJECTIVES: Although there is general agreement that surgery is the best treatment for chronic subdural haematoma (CSDH), the extent of the surgical intervention is not well defined. METHODS: The less invasive surgical technique of bedside percutaneous subdural tapping and spontaneous haematoma efflux after twist drill craniostomy under local anaesthesia was prospectively analysed in 118 adult patients, 99 with unilateral and 19 with bilateral CSDH. RESULTS: The mean number of subdural tappings was 3.2. Ninety two of the patients with unilateral CSDH were successfully treated by up to five subdural tappings, 95% of the patients with bilateral CSDH were successfully treated by up to 10 subdural tappings. The mean duration of inpatient treatment was 12 days. In 11 patients (9%) the treatment protocol had to be abandoned because of two acute subdural bleedings, two subdural empyemas, and seven cases of insufficient haematoma efflux and no neurological improvement. The only significant predictor for failure of the described treatment protocol was septation visible on preoperative CT. CONCLUSIONS: The described therapy protocol is-apart from a purely conservative treatment-the least invasive presently available surgical technique for treating chronic subdural haematoma. Its results are comparable with other modern treatment protocols. Thus, it can be recommended in all patients as a first and minimally invasive therapy, especially in patients in a poor general condition. Patients with septation visible on preoperative CT should be excluded from this form of treatment. PMID- 10864603 TI - Reduced cerebral blood flow in white matter in ischaemic leukoaraiosis demonstrated using quantitative exogenous contrast based perfusion MRI. AB - OBJECTIVE: White matter hypoperfusion may play a part in the pathogenesis of ischaemic leukoaraiosis, but demonstration of this requires a high resolution quantitative method of cerebral blood flow (CBF) measurement. Initial exogenous contrast based MRI methods only allowed measurement of relative cerebral blood volume (CBV) values, but more recently a mathematical approach has been developed which enables absolute regional CBF and CBV to be determined. This technique was applied to patients with ischaemic leukoaraiosis to determine whether reduced white matter CBF in this patient group could be demonstrated. METHODS: Eight patients with ischaemic leukoaraiosis (radiological leukoaraiosis and clinical lacunar stroke), and nine age matched controls were studied. A spin echo echoplanar image sequence was used on a 1.5 Tesla MR system. An arterial input function was obtained from voxels placed over the middle cerebral arteries. Cerebral blood flow, CBV, and mean transit (MTT) maps were derived. Regions of interest were placed at standard positions in the white and grey matter and mean values of CBF, CBV, and MTT were compared between the two groups. RESULTS: Mean (SD) white matter CBF was significantly reduced in patients by 38% (13.40 (4.87) v 21.74 (3.53) ml/min/ 100 g, p=0.002). Significant reductions in CBF were seen in all white matter regions. By contrast there was no reduction in CBF in any grey matter region. There was no significant difference in white matter CBV between cases and controls; mean values were lower in all white matter regions for patients but this did not reach significance for any region. By contrast mean grey matter CBV was significantly higher in patients than in controls. Mean MTT values were higher in all regions of grey and white matter in the patient group, but this only achieved significance for the superior white matter. CONCLUSION: A quantitative MR perfusion method showed reduced white matter CBF in patients with ischaemic leukoaraiosis, but normal grey matter CBF. This is consistent with hypoperfusion playing a part in the pathogenesis of ischaemic leukoaraiosis. The absolute values of white matter and grey matter CBF obtained in the patient groups were very similar to those in previous PET studies, providing further evidence for the validity of the regional CBF measurements obtained using this quantitative MR perfusion technique. The high spatial resolution and lack of radioactive administration makes such techniques ideal for longitudinal studies in this condition. PMID- 10864604 TI - Family history and DNA analysis in patients with suspected Huntington's disease. AB - OBJECTIVES: Until recently a definite diagnosis of Huntington's disease could be made by a combination of clinical findings, a positive family history, and pathological confirmation. Prevalence data are based on these criteria. After finding the gene and its pathogenic mutation direct diagnostic confirmation became available. The aim of this study was to determine to what extent the direct assessment of CAG repeat length has allowed the diagnoses of additional patients, with atypical psychiatric or neurological disease, or those without a family history, that could otherwise not be diagnosed using traditional criteria. PATIENTS AND METHODS: From all 191 referred patients suspected of having Huntington's disease between July 1993 and January 1996 CAG repeat length was determined and the family history was reviewed in the Leiden roster. After a retrospective search the patients were subdivided in positive, negative, suspect, and unknown family histories. Patients with an expanded repeat (>35) were finally diagnosed as having Huntington's disease. The family history was compared with the repeat length and the clinical features. RESULTS: Clinical information was obtained for 172 patients. Of these, 126 patients had an expanded repeat, 77 had a positive, eight a negative, 40 a suspect, and one an unknown family history. Of the 44 patients with a normal repeat length four had a positive family history. Of the two patients with an intermediate repeat (between 30-36 repeats), one with a negative family history received a clinical diagnosis of Gilles de la Tourette's syndrome. The other had an unknown family history. CONCLUSION: Despite verification of the family history through the Leiden roster, many more patients and families could be diagnosed with the new approach than would have been possible with the traditional criteria. Because prevalence studies have been based on this type of information, the data suggest an underestimation of the prevalence of Huntington's disease in the community of 14%. PMID- 10864605 TI - Cognitive functioning after subthalamic nucleotomy for refractory Parkinson's disease. AB - OBJECTIVE: To evaluate whether subthalamic nucleotomy produces adverse cognitive effects in patients with Parkinson's disease. METHOD: Twelve patients with Parkinson's disease underwent stereotactic surgery to the subthalamic nucleus. Presurgical and postsurgical neuropsychological assessment of attention, memory, executive function, language, and verbal intellect were undertaken with a battery of tests designed to minimise potential contamination of cognitive effects by motor symptoms. RESULTS: There was no statistically significant difference in the cognitive tests results after operation for the group as a whole. Reliable change indexes were generated for the cognitive tests. Reliable change postoperatively was found on specific tests of verbal memory, attention, and planning. Left sided operations were associated with greater incidence of deterioration postsurgery. CONCLUSIONS: Preliminary data on the first reported cognitive changes after subthalamic nucleotomy suggested few adverse cognitive effects of the surgery although discrete neuropsychological changes were seen in some patients. These effects were consistent with current theories on the cognitive functions of the basal ganglia. PMID- 10864606 TI - The EQ-5D--a generic quality of life measure-is a useful instrument to measure quality of life in patients with Parkinson's disease. AB - OBJECTIVE: To test the feasibility and validity of the EQ-5D (a widely used generic (disease non-specific) quality of life (QoL) instrument which allows comparisons between different patient groups and the general population) to assess QoL in patients with Parkinson's disease. METHODS: All 124 patients with Parkinson's disease seen in a community based study on the prevalence of parkinsonism were asked to complete a QoL battery comprising the EQ-5D, the medical outcome study short form (SF-36), the PDQ-39, a disease specific instrument to assess QoL in PD, and the Beck depression inventory. A structured questionnaire interview and a complete neurological examination including the Hoehn and Yahr stage of illness scale, the Schwab and England disability scale, the motor section of the unified Parkinson's disease rating scale (UPDRS), and the mini mental state examination (MMSE) were performed on the same day. RESULTS: The response rate was 78% and the completion rate of the EQ-5D among responders was 96%. The EQ-5D summary index correlated strongly with the PDQ-39 (r=-0.75, p<0.0001) as well as the physical score of the SF-36 (r=0.61, p<0.0001). There was a significant correlation of the EQ-5D summary index with disease severity, as measured by the Hoehn and Yahr stage of illness, the Schwab and England disability scale, the motor section of the UPDRS, and the depression score. The EQ-5D summary index also distinguished between patients with and without depression, falls, postural instability, cognitive impairment hallucinations, and those with deterioration of health over the previous year. CONCLUSION: The EQ-5D is a feasible and valid instrument to measure QoL in Parkinson's disease and reflects the severity and complications of the disease. PMID- 10864608 TI - Camino intracranial pressure monitor: prospective study of accuracy and complications. AB - OBJECTIVES: The fibreoptic device is a type of intracranial pressure monitor which seems to offer certain advantages over conventional monitoring systems. This study was undertaken to analyse the accuracy, drift characteristics, and complications of the Camino fibreoptic device. METHODS: One hundred and eight Camino intracranial pressure (ICP) devices, in their three modalities, were implanted during 1997. The most frequent indication for monitoring was severe head injury due to road traffic accidents. RESULTS: Sixty eight probe tips were cultured; 13.2% of the cases had a positive culture without clinical signs of infection, and 2.9% had a positive culture with clinical signs of ventriculitis. The most common isolated pathogen was Staphylococcus epidermidis. All patients were under cephalosporin prophylaxis during monitoring. Haemorrhage rate in patients without coagulation disorders was 2.1% and 15.3% in patients with coagulation abnormalities. Drift characteristics were studied in 56 cases; there was no drifting from the values expected according to the manufacturer's specifications in 34 probes. There was no relation between direction of the drift and duration of placement, nor between drift and time. CONCLUSIONS: Although the complication and drift rates were similar to those reported elsewhere, there was no correlation between the direction of the drift and long term monitoring despite the fact that some published papers refer to overestimation of values with time with this type of device. PMID- 10864607 TI - CSF sulfatide distinguishes between normal pressure hydrocephalus and subcortical arteriosclerotic encephalopathy. AB - OBJECTIVES: To examine the CSF concentrations of molecules reflecting demyelination, neuronal and axonal degeneration, gliosis, monoaminergic neuronal function, and aminergic and peptidergic neurotransmission in a large series of patients with normal pressure hydrocephalus (NPH) or subcortical arteriosclerotic encephalopathy (SAE), to elucidate pathogenic, diagnostic, and prognostic features. METHODS: CSF concentrations of glycosphingolipid (sulfatide), proteins (neurofilament triplet protein (NFL), glial fibrillary acidic protein (GFAP)), neuropeptides (vasoactive intestinal peptide (VIP), 4-aminobutyric acid (GABA)), and monoamines (homovanillic acid (HVA), 5-hydroxy-indoleacetic acid (5-HIAA), 4 hydroxy-3-methoxyphenylglycol (HMPG)) were analysed in 43 patients with NPH and 19 patients with SAE. The diagnoses of NPH and SAE were based on strict criteria and patients with NPH were subsequently operated on. Twelve clinical variables, psychometric tests measuring perceptual speed, accuracy, learning, and memory and a psychiatric evaluation were performed in all patients and before and after a shunt operation in patients with NPH. RESULTS: The CSF sulfatide concentration was markedly increased in patients with SAE (mean 766, range 300-3800 nmol/l) compared with patients with NPH (mean 206, range 50-400 nmol/l) (p<0.001). 5 HIAA, GABA, and VIP in CSF were higher in patients with SAE than in patients with NPH. The patients with NPH with cerebrovascular aetiology had higher sulfatide concentrations and a poorer outcome after shunt surgery than patients with NPH with other aetiologies. CONCLUSIONS: The pathogenesis of the white matter changes in NPH and SAE is different and ischaemic white matter changes can be a part of the NPH state. The markedly increased CSF sulfatide concentrations in patients with SAE indicate ongoing demyelination as an important pathophysiological feature of SAE. The CSF sulfatide concentration distinguished between patients with SAE and those with NPH with a sensitivity of 74% and a specificity of 94%, making it an important diagnostic marker. PMID- 10864609 TI - The circle of Willis (1621-75). PMID- 10864610 TI - Neurovascular decompression for idiopathic tarsal tunnel syndrome: technical note. AB - OBJECTIVE: The surgical outcome of idiopathic tarsal tunnel syndrome (TTS) is reported to be worse than that attributable to ganglion, tarsal coalition, or tumour, and therefore further development in the surgical treatment for idiopathic TTS is considered to be necessary. Here the efficacy of neurovascular decompression for patients with idiopathic TTS is evaluated. METHODS: Twelve feet from nine patients with idiopathic TTS were treated. The patients were aged 52-78 years (mean 64.6 years), and all of them complained of pain or dysaesthesia of the sole of the foot. The posterior tibial nerve was freed from the attached arteriovenous complex (posterior tibial artery and veins). The dissected nerve had a flattened appearance in all of the patients, suggesting nerve compression by the adjacent arteriovenous complex and superficially by the flexor retinaculum. A graft of fat was inserted as both a cushion and an antiadhesive between the vessels and the nerve to achieve neurovascular decompression. RESULTS: Patients on whom neurovascular decompression was performed had resolution or lessening of symptoms in their feet. Neither wound infection nor recurrence of symptoms was found during the follow up period (mean 26.8 months). CONCLUSION: Neurovascular compression syndrome plays a part in idiopathic TTS, and adding neurovascular decompression to resection of the flexor retinaculum is effective. PMID- 10864611 TI - Somatosensory evoked potentials after multisegmental lower limb stimulation in focal lesions of the lumbosacral spinal cord. AB - OBJECTIVES: Recording techniques permit the separate analysis of the response from cauda equina roots and the spinal potential that is probably generated by the activation of dorsal horn cells. To improve the functional assessment of focal lesions of the lumbosacral cord, lower limb somatosensory evoked potentials (SEPs) were measured by multisegmental stimulation. METHODS: Common peroneal and tibial nerves SEPs were recorded in 14 patients in whom MRI demonstrated compressive cord damage ranging from T9 to L1 levels. SEPs were recorded in each patient at the lumbar level (cauda equina response), lower thoracic level (spinal response), and from the scalp (cortical response). RESULTS: Abnormalities in spinal response occurred in 50% and 70% of tibial and common peroneal nerve SEPs respectively; these findings were well explained by the radiological compression level, involving in most of the patients lumbar rather than sacral myelomeres. The SEPs were often more effective than the clinical examination in showing the actual extension of damage. CONCLUSIONS: The recording of spinal SEPs after multisegmental lower limb stimulation proved useful in assessing cord dysfunction and determining the cord levels mainly involved by the compression. PMID- 10864612 TI - Persistence of tropical ataxic neuropathy in a Nigerian community. AB - OBJECTIVES: The term tropical ataxic neuropathy (TAN) is currently used to describe several neurological syndromes attributed to toxiconutritional causes. However, TAN was initially proposed to describe a specific neurological syndrome seen predominantly among the Ijebu speaking Yorubas in south western Nigeria. In this study, the prevalence of TAN was determined in Ososa, a semiurban community in south western Nigeria described as endemic for TAN in 1969, and its neurological features were compared with Strachan's syndrome, prisoners of war neuropathy, the epidemic neuropathy in Cuba, and konzo. METHODS: A census of Ososa was followed by door to door screening of all subjects aged 10 years and above with a newly designed screening instrument. Subjects who screened positive had a neurological examination, and the diagnosis of TAN was made if any two or more of bilateral optic atrophy, bilateral neurosensory deafness, sensory gait ataxia, or distal symmetric sensory polyneuropathy were present. RESULTS: A total of 4583 inhabitants were registered in the census. Of these, 3428 subjects aged 10 years and above were screened. The diagnosis of TAN was made in 206 of 323 subjects who screened positive for TAN. The prevalence of TAN was 6. 0%, 3.9% in males and 7.7% in females. The highest age specific prevalence was 24% in the 60 69 years age group in women. CONCLUSION: The occurrence of TAN in Ososa continues at a higher prevalence than was reported 30 years ago. Its neurological features and natural history do not resemble those described for Strachan syndrome, epidemic neuropathy in Cuba, or konzo. The increasing consumption of cassava foods linked to its causation makes TAN of public health importance in Nigeria, the most populous African country. PMID- 10864613 TI - Phonological grouping is specifically affected in cerebellar patients: a verbal fluency study. AB - OBJECTIVES: Recent clinical and functional neuroimaging evidence points towards a cerebellar role in verbal production. At present it is not clear how the cerebellum participates in language production. The aim was to investigate the influence of cerebellar lesions on verbal fluency abilities with specific focus on the verbal searching strategies employed by patients with cerebellar damage. METHODS: Twenty five patients with focal or degenerative cerebellar disease and 14 control subjects were tested in a timed verbal fluency task requiring word production under forced (phonemic or semantic) conditions. To analyse the verbal searching strategy employed, semantic and phonemic cluster analyses were also performed. RESULTS: Performances of cerebellar patients were comparable with those of controls in the semantic task; conversely their performances were significantly impaired when tested in the letter task. Cluster analysis results showed that the verbal fluency impairment is linked to specific damage of phonemically related retrieval strategies. CONCLUSION: Cerebellar damage impairs verbal fluency by specifically affecting phonemic rule performances while sparing semantic rule ones. These findings underline the importance of the cerebellar computing properties in strategy development in the linguistic domain. PMID- 10864614 TI - Familial aggregation of Parkinson's disease in a Finnish population. AB - Familial aggregation of Parkinson's disease in a Finnish population was investigated. A family history was obtained on 268 patients with Parkinson's disease and 210 controls ascertained from the population of the province of northern Ostrobothnia, Finland. Ten per cent of the probands reported an affected first degree relative, whereas the corresponding frequency was 3.8 per cent in the controls (p=0.01). The relative risk of Parkinson's disease among the first degree relatives of the patients with Parkinson's disease was 2.9 (95 % confidence interval 1.3-6.4) and the cumulative incidence of Parkinson's disease by the age of 90 years was 3.3-fold higher among the first degree relatives of the patients than those of the controls. The crude segregation ratio was 0.27 for the siblings and 0.17 for the parents suggesting that recessive inheritance may be more common than dominant inheritance among Finnish patients with Parkinson's disease. PMID- 10864615 TI - Isolated finger flexion: a novel form of focal neuromyotonia. AB - Two almost identical elderly women are described who presented with gradually progressive painless involuntary flexion of the ring and middle fingers over 12 months, leading eventually to contractures. The flexion deformity persisted during sleep and was the sole neurological abnormality. Both patients had advanced chronic obstructive pulmonary disease and were on long term salbutamol and oxygen. Neurophysiological studies indicated that this was due to neuromyotonia mainly involving flexor digitorum superficialis muscles without evidence of underlying peripheral neuropathy, proximal conduction block, or generalised neuromyotonia. Voltage gated potassium channel antibodies were negative. The clinical and neurophysiological picture remained static over a 2 year follow up period. It is suggested that this is a novel form of acquired focal neuromyotonia and speculate both on its cause and distribution. PMID- 10864616 TI - Extension of the clinical range of facioscapulohumeral dystrophy: report of six cases. AB - Consensual diagnostic criteria for facioscapulohumeral dystrophy (FSHD) include onset of the disease in facial or shoulder girdle muscles, facial weakness in more than 50% of affected family members, autosomal dominant inheritance in familial cases, and evidence of myopathic disease in at least one affected member without biopsy features specific to alternative diagnoses. Six patients did not meet most of these criteria but were diagnosed as FSHD by DNA testing, which showed small EcoRI fragments on chromosome 4q. Their clinical signs and symptoms and results of auxiliary investigations are reported. The patients presented with foot extensor, thigh, or calf muscle weakness. None of them had apparent facial weakness, only one complained of weakness in the shoulders, none had a positive family history. Expert physical examination, however, showed a typical facial expression, an abnormal shoulder configuration on lifting the arms, or scapular winging. This raised the suspicion of FSHD, whereupon DNA analysis was done. In conclusion, the clinical expression of FSHD is much broader than indicated by the nomenclature. The possibility to perform DNA tests is likely to greatly expand the clinical range of FSHD. PMID- 10864617 TI - Validating a screening questionnaire for parkinsonism in Australia. AB - Parkinson's disease is a common neurodegenerative disorder in elderly people. Epidemiological studies of the disease can be labour intensive. A two phase design including a screening questionnaire as the first phase has become a popular method in prevalence studies of Parkinson's disease. Such a design has many advantages including less work for assessing physicians and enhanced recruitment of people to be screened. However, its wider application may be questioned because validation has been limited to samples that are drawn from hospitals (or clinics) and may be inappropriate for a community setting. This study assesses whether validating screening questionnaire by using a hospital sample yields the same result as a community based sample. Furthermore, it seeks to establish whether the screening instrument can be simplified to involve less questions. The findings show that some of the questions used in the screening phase yield different responses when comparing a hospital group with a community group. This study also provides a simplified model of questions that may be relevant for screening in the community setting. PMID- 10864618 TI - Injections of botulinum toxin A into the salivary glands improve sialorrhoea in amyotrophic lateral sclerosis. AB - Sialorrhoea is a socially disabling problem in bulbar amyotrophic lateral sclerosis (ALS). Botulinum toxin A (BoNT/A) was injected into the salivary glands in five patients with bulbar ALS and sialorrhoea. The effect of BoNT/A was measured by the number of paper handkerchiefs used each day and by salivary gland scintigraphy. BoNT/A ameliorated sialorrhoea and quality of life without major adverse effects. BoNT/A may be a relatively safe and effective treatment for sialorrhoea in selected patients. PMID- 10864619 TI - Arnold-Chiari malformation and nystagmus of skew. AB - The Arnold-Chiari malfomation is typically associated with downbeat nystagmus. Eye movement recordings in two patients with Arnold-Chiari malfomation type 1 showed, in addition to downbeat and gaze evoked nystagmus, intermittent nystagmus of skew. To date this finding has not been reported in association with Arnold Chiari malfomation. Nystagmus of skew should raise the suspicion of Arnold-Chiari malfomation and prompt sagittal head MRI examination. PMID- 10864620 TI - Long term follow up after perimesencephalic subarachnoid haemorrhage. AB - OBJECTIVES: To evaluate the long term sequelae of perimesencephalic subarachnoid haemorrhage (PMSAH). METHODS: Twenty one consecutive patients were studied. All patients were examined by CT, angiography, MRI, multimodal evoked potentials, and transcranial Doppler sonography. All relevant clinical data during hospital stay and outcome at discharge were obtained by reviewing the charts. Long term follow up was evaluated by reviewing the outpatient files and dedicated outpatient review. Patients were specifically questioned about their perceived recovery, residual complaints, and present occupational status. RESULTS: Apart from the initial CT confirming the diagnosis of PMSAH all other examinations disclosed no abnormalities. None of the patients developed any complications during hospital stay, and all patients were discharged in good clinical condition and without neurological deficits. At long term follow up 62% of the patients had residual complaints consisting of headaches, irritability, depression, forgetfulness, weariness, and diminished endurance. Apart from four patients who had already retired before the PMSAH, only seven of the remaining 17 patients (41%) returned to their previous occupation, whereas nine patients (53%) retired from work and one man became unemployed. One patient had a recurrence of PMSAH 31 months after the first event. CONCLUSION: PMSAH can have considerable long term psychosocial sequelae, and may also recur. Prognosis may not be as good as previously reported. PMID- 10864621 TI - Edmond Isidore Etienne Nocard (1850-1903). PMID- 10864622 TI - Isolated Horner's syndrome and syringomyelia. AB - Although syringomyelia has been associated with Horner's syndrome, it is typically associated with other neurological findings such as upper limb weakness or numbness. A patient is described who had an isolated Horner's syndrome as the only manifestation of syringomyelia. A 76 year old woman was discovered to have right upper lid ptosis and right pupillary miosis. Neurological examination was unremarkable, and pharmacological testing was consistent with localisation of the lesion to a first or second order sympathetic neuron. Neuroimaging disclosed a Chiari I malformation with a syrinx extending to the C2 to C4 level. An isolated Horner's syndrome may be the presenting manifestation of syringomyelia. PMID- 10864623 TI - Sunflower cataract in Wilson's disease. PMID- 10864624 TI - Retro-ocular headache with autonomic features resembling "continuous" cluster headache in lateral medullary infarction. PMID- 10864625 TI - Chronic autonomic neuropathy in a patient with primary Sjogren's syndrome. PMID- 10864626 TI - C1/C2 rotary subluxation due to spasmodic torticollis. PMID- 10864627 TI - Ataxic form of Guillain-Barr syndrome associated with anti-GD1b IgG antibody. PMID- 10864628 TI - Analysis of adenosine deaminase isoenzyme-2 (ADA(2)) in cerebrospinal fluid in the diagnosis of tuberculosis meningitis. PMID- 10864629 TI - Unilateral focal lesions in the rostrolateral medulla influence chemosensitivity and breathing measured during wakefulness, sleep, and exercise. PMID- 10864631 TI - Mosby's Color Atlas and Text of Neurology. PMID- 10864630 TI - Vertigo. Its Multisensory Syndromes. PMID- 10864632 TI - Sobemoviruses. PMID- 10864633 TI - An enzymatic footprinting analysis of the interaction of 40S ribosomal subunits with the internal ribosomal entry site of hepatitis C virus. AB - Hepatitis C virus translation is initiated on a approximately 330-nucleotide (nt) long internal ribosomal entry site (IRES) at the 5' end of the genome. In this process, a 43S preinitiation complex (comprising a 40S ribosomal subunit, eukaryotic initiation factor 3 (eIF3), and a ternary [eIF2-GTP-initiator tRNA] complex) binds the IRES in a precise manner so that the initiation codon is placed at the ribosomal P site. This binding step involves specific interactions between the IRES and different components of the 43S complex. The 40S subunit and eIF3 can bind to the IRES independently; previous analyses revealed that eIF3 binds specifically to an apical half of IRES domain III. Nucleotides in the IRES that are involved in the interaction with the 40S subunit were identified by RNase footprinting and mapped to the basal half of domain III and in domain IV. Interaction sites were identified in locations that have been found to be essential for IRES function, including (i) the apical loop residues GGG(266-268) in subdomain IIId and (ii) the pseudoknot. Extensive protection from RNase cleavage also occurred downstream of the pseudoknot in domain IV, flanking both sides of the initiation codon and corresponding in length to that of the mRNA binding cleft of the 40S subunit. These results indicate that the 40S subunit makes multiple interactions with the IRES and suggest that only nucleotides in domain IV are inserted into the mRNA-binding cleft of the 40S subunit. PMID- 10864634 TI - Deletion mutagenesis downstream of the 5' long terminal repeat of human immunodeficiency virus type 1 is compensated for by point mutations in both the U5 region and gag gene. AB - We have studied the role of an RNA region at nucleotides (nt) +200 to +233, just downstream of the 5' long terminal repeat, in encapsidation of human immunodeficiency virus type 1 genomic RNA. Three deletion mutations, namely, BH D0, BH-D1, and BH-D2, were generated to eliminate sequences at positions nt +200 to +219, +200 to +226, and +200 to +233. The result in each case was decreased levels of packaging of viral RNA into the mutated viruses, with the BH-D2 virus being the most severely affected. Consistently, all three deletions resulted in impaired viral infectiousness and the BH-D2 mutation showed the most dramatic impact in this regard. Further analysis revealed additional defects in Gag precursor processing and in the extension efficiency of the tRNA(3)(Lys) primer in reverse transcription reactions performed with these mutated viruses. To shed further light on the function of these deleted sequences in viral replication, the mutated viruses were cultured in MT-2 cells over prolonged periods to enable them to reacquire wild-type replication kinetics. Sequencing of the reverted viruses revealed point mutations in both the noncoding region and the gag gene. In the case of the BH-D0 revertant, two mutations were observed at positions G112A in the U5 region, termed M1, and T24I in the nucleocapsid protein, termed MNC, respectively. Either of these two mutations was able to confer wild-type replication capacity on BH-D0. In the case of BH-D1, each of the M1 mutations, a mutation termed M2, i.e., C227T, just downstream of the primer binding site, a mutation termed MP2 (T12I) in the p2 protein, and the MNC mutation were observed. A combination of either M1 and M2 or MP2 and MNC was able to rescue BH-D1. In the case of the BH-D2 deletion-containing viruses, three point mutations, i.e., M1, MP2, and MNC, were observed and the presence of all three was required to restore viral replication to wild-type levels. PMID- 10864635 TI - Evolution of lamivudine resistance in human immunodeficiency virus type 1 infected individuals: the relative roles of drift and selection. AB - Human immunodeficiency virus type 1 (HIV-1) rapidly develops resistance to lamivudine during monotherapy, typically resulting in the appearance at position 184 in reverse transcriptase (RT) of isoleucine instead of the wild-type methionine (M184I) early in therapy, which is later replaced by valine (M184V). M184V reduces viral susceptibility to drug in vitro by approximately 100-fold, but also results in a lower processivity of RT. We show that a drop in absolute viral fitness associated with the outgrowth of M184V results in a drop in viral load only in individuals with high CD4(+) counts, from whom we estimate the relative fitness of M184V in the presence of drug to be approximately 10% of that of the wild type prior to therapy. The timing of emergence of the M184V mutant varies widely between infected individuals. From analysis of the frequency of M184I and M184V mutants determined at multiple time points in seven individuals during lamivudine therapy, we estimated the fitness advantage of M184V over M184I during therapy to be approximately 23% on average. We have also estimated the average ratio of the frequencies of the two mutants prior to therapy to be 0. 2:1, with a range from 0.12:1 to 0.33:1. We have found that the differences between individuals in the rate of evolution of lamivudine resistance arise due to genetic drift affecting the relative frequency of M184I and M184V prior to therapy. These results show that stochastic effects can be significant in HIV evolution, even when there is large fitness difference between mutant and wild type variants. PMID- 10864636 TI - In vivo and in vitro identification of structural and sequence elements of the human parechovirus 5' untranslated region required for internal initiation. AB - Sequence analysis of the picornavirus echovirus 22 led to its classification as the first member of a new genus, Parechovirus, and renaming as human parechovirus type 1 (HPeV1). Although distinct from other genera in most of the genome, the 5' untranslated region (5'UTR) shows similarities to that of cardio/aphthoviruses in some of its structural domains (A to L). The 5'UTR plays an important role in picornavirus translation initiation and in RNA synthesis. To investigate translation in HPeV1, we engineered an extensive range of mutations (including precise deletions and point mutations) into the 5'UTR. Their effects were studied both by in vitro transcription-translation using a bicistronic construct and by in vivo studies using an infectious, full-length HPeV1 cDNA. These approaches allowed the HPeV1 internal ribosome entry site (IRES) to be mapped. Deletions within the first 298 nucleotides had little impact in the in vitro system, while deletions of nucleotides 298 to 538 had a significant effect. Precise removal of domains H and L (nucleotides 287 to 316 and 664 to 682, respectively) did not significantly reduce translation efficiency in vitro, while domains I, J, and K (nucleotides 327 to 545, 551 to 661, and 614 to 645, respectively) appeared to have much more important roles. Mutation of a phylogenetically conserved GNRA motif (positions 421 to 424) within domain I severely reduced translation. We also confirmed the identity of the AUG (positions 710 to 712) which initiates the open reading frame, the positive identification of which has not been possible previously, as the N terminus of the polyprotein is blocked and not amenable to sequence analysis. This is therefore important in understanding parechovirus genome organization. Mutation of the AUG or an upstream polypyrimidine tract leads to aberrant translation, suggesting they both form part of the parechovirus Yn-Xm-AUG motif. In vivo experiments confirmed the importance of domains I, J, and K, the conserved GNRA motif, polypyrimidine sequences, and AUG, as mutations here were lethal. These features are also important in the IRES elements of cardio/aphthoviruses, but other features reported to be part of the IRES of some members of these genera, notably domains H and L, do not appear to be critical in HPeV1. This adds weight to the idea that there may be functional differences between the IRES elements of different picornaviruses, even when they share significant structural similarity. PMID- 10864637 TI - Interleukin-12 (IL-12) enhancement of the cellular immune response against human immunodeficiency virus type 1 env antigen in a DNA prime/vaccinia virus boost vaccine regimen is time and dose dependent: suppressive effects of IL-12 boost are mediated by nitric oxide. AB - We previously demonstrated that codelivery of interleukin-12 (IL-12) with the human immunodeficiency virus type 1 (HIV-1) Env antigen from a recombinant vaccinia virus (rVV) can enhance the specific anti-Env cell-mediated immune (CMI) response. In the present study, we have investigated the effects of IL-12 in mice when it is expressed in a DNA prime/VV boost vaccine regimen. The delivery of IL 12 and Env product during priming with a DNA vector, followed by a booster with VV expressing the Env gene (rVVenv), was found to trigger the optimal CMI response compared with other immunization schedules studied. Significantly, if IL 12 is also delivered as a booster from the viral vector, an impairment of the effects of IL-12 was observed involving nitric oxide (NO), since it was overcome by specific inhibitors of inducible NO synthase. NO caused transient immunosuppression rather than impairment of viral replication. Moreover, at certain viral doses, coadministration of the NO inhibitor during the booster resulted in IL-12-mediated enhancement of the specific CD8(+) T-cell response. In addition, the dose of the IL-12-encoding plasmid (pIL-12) and the route of administration of both vectors were relevant factors for optimal CMI responses. Maximal numbers of Env-specific CD8(+) gamma interferon-secreting cells were obtained when 50 microg of pIL-12 was administered intramuscularly at priming, followed by an intravenous rVVenv boost. Our results demonstrate, in a murine model, critical parameters affecting the success of vaccination schedules based on a combination of DNA and VV vectors in conjunction with immunomodulators. PMID- 10864638 TI - Evidence that herpes simplex virus VP16 is required for viral egress downstream of the initial envelopment event. AB - During infection with herpes simplex virus type 1 (HSV-1), VP16 serves multiple functions, including transcriptional activation of viral immediate early genes and downregulation of the virion host shutoff protein vhs. Furthermore, VP16 has been shown to be involved in some aspect of virus assembly and/or maturation. Experiments with a VP16 null virus, 8MA, suggested that VP16 plays a direct role in virion assembly, since removal of VP16 from the HSV-1 genome results in reduced levels of encapsidated DNA and a failure to produce extracellular enveloped particles. However, VP16 null mutants display a severe translational arrest due to unrestrained vhs activity, thus complicating interpretation of these data. We examine here the role of VP16 in virion assembly and egress in the context of a vhs null background, using the virus 8MA/DeltaSma (VP16(-) vhs(-)). Comparison of 8MA and 8MA/DeltaSma with respect to viral DNA accumulation and encapsidation and accumulation of the major capsid protein, VP5, revealed that the 8MA lethal phenotype is only partially due to uncontrolled vhs activity, indicating that VP16 is required in HSV-1 virion formation. Electron microscopy confirmed these results and further showed that VP16 is required for HSV-1 egress beyond the perinuclear space. In addition, we describe the isolation and characterization of an 8MA derivative capable of propagation on Vero cells, due to second site mutations in the vhs and UL53 (gK) genes. Taken together, these results show that VP16 is required for viral egress downstream of the initial envelopment step and further underscore the importance of VP16 in controlling vhs activity within an infected cell. PMID- 10864639 TI - Isolation and characterization of monoclonal antibodies that inhibit hepatitis C virus NS3 protease. AB - A series of mouse monoclonal antibodies (MAbs) to the nonstructural protein 3 (NS3) of hepatitis C virus was prepared. One of these MAbs, designated 8D4, was found to inhibit NS3 protease activity. This inhibition was competitive with respect to the substrate peptide (K(i) = 39 nM) but was significantly decreased by the addition of the NS4A peptide, a coactivator of the NS3 protease. 8D4 also showed marked inhibition of the NS3-dependent cis processing of the NS3/4A polyprotein but had virtually no effect on the succeeding NS3/4A-dependent trans processing of the NS5A/5B polyprotein in vitro. Epitope mapping of 8D4 with a random peptide library revealed a consensus sequence, DxDLV, that matched residues 79 to 83 (DQDLV) of NS3, a region containing the catalytic residue Asp 81. Furthermore, synthetic peptides including this sequence were shown to block the ability of 8D4 to bind to NS3, indicating that 8D4 interacts with the catalytic region of NS3. The data showing decreased inhibition potency of 8D4 against the NS3/4A complex suggest that 8D4 recognizes the conformational state of the protease active site caused by the association of NS4A with the protease. PMID- 10864640 TI - Characterization of the influenza A virus gene pool in avian species in southern China: was H6N1 a derivative or a precursor of H5N1? AB - In 1997, an H5N1 influenza virus outbreak occurred in chickens in Hong Kong, and the virus was transmitted directly to humans. Because there is limited information about the avian influenza virus reservoir in that region, we genetically characterized virus strains isolated in Hong Kong during the 1997 outbreak. We sequenced the gene segments of a heterogeneous group of viruses of seven different serotypes (H3N8, H4N8, H6N1, H6N9, H11N1, H11N9, and H11N8) isolated from various bird species. The phylogenetic relationships divided these viruses into several subgroups. An H6N1 virus isolated from teal (A/teal/Hong Kong/W312/97 [H6N1]) showed very high (>98%) nucleotide homology to the human influenza virus A/Hong Kong/156/97 (H5N1) in the six internal genes. The N1 neuraminidase sequence showed 97% nucleotide homology to that of the human H5N1 virus, and the N1 protein of both viruses had the same 19-amino-acid deletion in the stalk region. The deduced hemagglutinin amino acid sequence of the H6N1 virus was most similar to that of A/shearwater/Australia/1/72 (H6N5). The H6N1 virus is the first known isolate with seven H5N1-like segments and may have been the donor of the neuraminidase and the internal genes of the H5N1 viruses. The high homology between the internal genes of H9N2, H6N1, and the H5N1 isolates indicates that these subtypes are able to exchange their internal genes and are therefore a potential source of new pathogenic influenza virus strains. Our analysis suggests that surveillance for influenza A viruses should be conducted for wild aquatic birds as well as for poultry, pigs, and humans and that H6 isolates should be further characterized. PMID- 10864641 TI - Interdependence of hemagglutinin glycosylation and neuraminidase as regulators of influenza virus growth: a study by reverse genetics. AB - The hemagglutinin (HA) of fowl plague virus A/FPV/Rostock/34 (H7N1) carries two N linked oligosaccharides attached to Asn123 and Asn149 in close vicinity to the receptor-binding pocket. In previous studies in which HA mutants lacking either one (mutants G1 and G2) or both (mutant G1,2) glycosylation sites had been expressed from a simian virus 40 vector, we showed that these glycans regulate receptor binding affinity (M. Ohuchi, R. Ohuchi, A. Feldmann, and H. D. Klenk, J. Virol. 71:8377-8384, 1997). We have now investigated the effect of these mutations on virus growth using recombinant viruses generated by an RNA polymerase I-based reverse genetics system. Two reassortants of influenza virus strain A/WSN/33 were used as helper viruses to obtain two series of HA mutant viruses differing only in the neuraminidase (NA). Studies using N1 NA viruses revealed that loss of the oligosaccharide from Asn149 (mutant G2) or loss of both oligosaccharides (mutant G1,2) has a pronounced effect on virus growth in MDCK cells. Growth of virus lacking both oligosaccharides from infected cells was retarded, and virus yields in the medium were decreased about 20-fold. Likewise, there was a reduction in plaque size that was distinct with G1,2 and less pronounced with G2. These effects could be attributed to a highly impaired release of mutant progeny viruses from host cells. In contrast, with recombinant viruses containing N2 NA, these restrictions were much less apparent. N1 recombinants showed lower neuraminidase activity than N2 recombinants, indicating that N2 NA is able to partly overrule the high-affinity binding of mutant HA to the receptor. These results demonstrate that N-glycans flanking the receptor binding site of the HA molecule are potent regulators of influenza virus growth, with the glycan at Asn149 being dominant and that at Asn123 being less effective. In addition, we show here that HA and NA activities need to be highly balanced in order to allow productive influenza virus infection. PMID- 10864642 TI - Epstein-Barr virus gH is essential for penetration of B cells but also plays a role in attachment of virus to epithelial cells. AB - Entry of Epstein-Barr virus (EBV) into B cells is initiated by attachment of glycoprotein gp350 to the complement receptor type 2 (CR2). A complex of three glycoproteins, gH, gL, and gp42, is subsequently required for penetration. Gp42 binds to HLA class II, which functions as an entry mediator or coreceptor and, by analogy with other herpesviruses, gH is then thought to be involved virus-cell fusion. However, entry of virus into epithelial cells is thought to be different. It can be initiated by attachment by an unknown glycoprotein in the absence of CR2. There is no interaction between gp42 and HLA class II and instead a distinct complex of only the two glycoproteins gH and gL interacts with a novel entry mediator. Again, by analogy with other viruses gH is thought to be critical to fusion. To investigate further the different roles of gH in infection of the two cell types and to examine its influence on the assembly of the gH-gL-gp42 complex, we constructed two viruses, one in which the gH open reading frame was interrupted by a cassette expressing a neomycin resistance gene and the gene for green fluorescent protein and one as a control in which the neighboring nonessential thymidine kinase gene was interrupted with the same cassette. Virus lacking gH exited from cells normally, although loss of gH resulted in rapid turnover of gL and gp42 as well. The virus bound normally to B lymphocytes but could not infect them unless cells and bound virus were treated with polyethylene glycol to induce fusion. In contrast, virus that lacked the gH complex was impaired in attachment to epithelial cells and the effects of monoclonal antibodies to gH implied that this resulted from loss of gH rather than other members of the complex. These results suggest a role for gH in both attachment and penetration into epithelial cells. PMID- 10864643 TI - Identification of the pseudorabies virus promoter required for latency-associated transcript gene expression in the natural host. AB - Expression of the latency-associated transcript (LAT) gene is a hallmark of alphaherpesvirus latency, and yet its control and function remain an enigma. Resolution of this problem will require verification and subsequent elimination or disabling of elements regulating LAT gene transcription so that the influence of the resultant RNA can be evaluated. Toward this end, we generated a novel pseudorabies virus (PrV) recombinant in which a 282-bp region containing the LAP1 (first latency-active promoter) consensus sequence was replaced by a reporter cassette. Despite this substitution, replication of the recombinant was comparable to that of the parental and rescuant viruses both in cultured mammalian cells and in the natural host, swine. Furthermore, production of the LAT gene-associated 2.0- and 8.0-kb RNAs during an in vitro lytic infection of cultured neuronal cells was unaffected. However, the otherwise constitutively produced and processed 8.4-kb LAT was not detected in porcine trigeminal ganglia latently infected with this novel recombinant, although the viral genome was shown to be present. Therefore, LAP1 is apparently the basal promoter for PrV LAT gene expression during viral latency but is not required for such activity during an in vitro lytic infection of neuronal cells. More importantly, the ability of PrV to persist in a latent state in the absence of LAT suggests that other factors are responsible for this event in the natural host. PMID- 10864644 TI - Generation and characterization of a hepatitis C virus NS3 protease-dependent bovine viral diarrhea virus. AB - Unique to pestiviruses, the N-terminal protein encoded by the bovine viral diarrhea virus (BVDV) genome is a cysteine protease (Npro) responsible for a self cleavage that releases the N terminus of the core protein (C). This unique protease is dispensable for viral replication, and its coding region can be replaced by a ubiquitin gene directly fused in frame to the core. To develop an antiviral assay that allows the assessment of anti-hepatitis C virus (HCV) NS3 protease inhibitors, a chimeric BVDV in which the coding region of Npro was replaced by that of an NS4A cofactor-tethered HCV NS3 protease domain was generated. This cofactor-tethered HCV protease domain was linked in frame to the core protein of BVDV through an HCV NS5A-NS5B junction site and mimicked the proteolytic function of Npro in the release of BVDV core for capsid assembly. A similar chimeric construct was built with an inactive HCV NS3 protease to serve as a control. Genomic RNA transcripts derived from both chimeric clones, P(H/B) (wild-type HCV NS3 protease) and P(H/B(S139A)) (mutant HCV NS3 protease) were then transfected into bovine cells (MDBK). Only the RNA transcripts from the P(H/B) clone yielded viable viruses, whereas the mutant clone, P(H/B(S139A)), failed to produce any signs of infection, suggesting that the unprocessed fusion protein rendered the BVDV core protein defective in capsid assembly. Like the wild-type BVDV (NADL), the chimeric virus was cytopathic and formed plaques on the cell monolayer. Sequence and biochemical analyses confirmed the identity of the chimeric virus and further revealed variant viruses due to growth adaptation. Growth analysis revealed comparable replication kinetics between the wild-type and the chimeric BVDVs. Finally, to assess the genetic stability of the chimeric virus, an Npro-null BVDV (BVDV-Npro in which the entire Npro coding region was deleted) was produced. Although cytopathic, BVDV-Npro was highly defective in viral replication and growth, a finding consistent with the observed stability of the chimeric virus after serial passages. PMID- 10864645 TI - CDP is a repressor of mouse mammary tumor virus expression in the mammary gland. AB - Mouse mammary tumor virus (MMTV) transcription is highest in the lactating mammary gland but is detectable in a variety of other tissues. Previous results have shown that MMTV expression is suppressed in lymphoid and other tissues through the binding of the homeodomain-containing repressor special AT-rich binding protein 1 to a negative regulatory element (NRE) in the MMTV long terminal repeat (LTR). Another homeoprotein repressor, CCAAT displacement protein (CDP), also binds to the MMTV NRE, but a role for CDP in MMTV transcriptional suppression has not yet been demonstrated. In this paper, we show that the level of CDP decreases during development of the mammary gland and that this decline in CDP level correlates with the known increase in MMTV expression observed during mammary gland differentiation. Moreover, CDP overexpression was able to suppress MMTV LTR-reporter gene activity up to 20-fold in transient-transfection assays of mouse mammary cells. To determine if this effect was due to direct binding of CDP to the promoter-proximal NRE, we performed DNase I protection assays to map two CDP-binding sites from +835 to +845 and +920 to +931 relative to the first base of the LTR. Mutations engineered into each of these sites decreased CDP binding to the proximal NRE, whereas a combination of these mutations further reduced binding. Subsequently, each of these mutations was introduced into the full length MMTV LTR upstream of the luciferase reporter gene. Analysis of stable transfectants of LTR constructs showed that CDP binding site mutations in the proximal NRE elevated reporter gene expression two- to sixfold compared to wild type LTR constructs. Thus, MMTV expression increases during mammary gland development, in part due to decreased CDP levels and CDP binding to the LTR. Together, these experiments provide the first evidence that CDP acts as a repressor of MMTV transcription in the mammary gland. PMID- 10864646 TI - Vaccination against canine distemper virus infection in infant ferrets with and without maternal antibody protection, using recombinant attenuated poxvirus vaccines. AB - Canine distemper virus (CDV) infection of ferrets is clinically and immunologically similar to measles, making this a useful model for the human disease. The model was used to determine if parenteral or mucosal immunization of infant ferrets at 3 and 6 weeks of age with attenuated vaccinia virus (NYVAC) or canarypox virus (ALVAC) vaccine strains expressing the CDV hemagglutinin (H) and fusion (F) protein genes (NYVAC-HF and ALVAC-HF) would induce serum neutralizing antibody and protect against challenge infection at 12 weeks of age. Ferrets without maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 5) or ALVAC-HF (n = 4) developed significant neutralizing titers (log(10) inverse mean titer +/- standard deviation of 2.30 +/- 0.12 and 2.20 +/- 0.34, respectively) by the day of challenge, and all survived with no clinical or virologic evidence of infection. Ferrets without maternal antibody that were vaccinated intranasally (i.n.) developed lower neutralizing titers, with NYVAC-HF producing higher titers at challenge (1.11 +/- 0.57 versus 0.40 +/- 0.37, P = 0.02) and a better survival rate (6/7 versus 0/5, P = 0.008) than ALVAC-HF. Ferrets with maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 7) and ALVAC-HF (n = 7) developed significantly higher antibody titers (1.64 +/ 0. 54 and 1.28 +/- 0.40, respectively) than did ferrets immunized with an attenuated CDV vaccine (0.46 +/- 0.59; n = 7) or the recombinant vectors expressing rabies glycoprotein (RG) (0.19 +/- 0.32; n = 8, P = 7 x 10(-6)). The NYVAC vaccine also protected against weight loss, and both the NYVAC and attenuated CDV vaccines protected against the development of some clinical signs of infection, although survival in each of the three vaccine groups was low (one of seven) and not significantly different from the RG controls (none of eight). Combined i.n.-parenteral immunization of ferrets with maternal antibody using NYVAC-HF (n = 9) produced higher titers (1.63 +/- 0. 25) than did i.n. immunization with NYVAC-HF (0.88 +/- 0.36; n = 9) and ALVAC-HF (0.61 +/- 0.43; n = 9, P = 3 x 10(-7)), and survival was also significantly better in the i.n. parenteral group (3 of 9) than in the other HF-vaccinated animals (none of 18) or in controls immunized with RG (none of 5) (P = 0.0374). Multiple routes were not tested with the ALVAC vaccine. The results suggest that infant ferrets are less responsive to i.n. vaccination than are older ferrets and raises questions about the appropriateness of this route of immunization in infant ferrets or infants of other species. PMID- 10864647 TI - Selective membrane permeabilization by the rotavirus VP5* protein is abrogated by mutations in an internal hydrophobic domain. AB - Rotavirus infectivity is dependent on the proteolytic cleavage of the VP4 spike protein into VP8* and VP5* proteins. Proteolytically activated virus, as well as expressed VP5*, permeabilizes membranes, suggesting that cleavage exposes a membrane-interactive domain of VP5* which effects rapid viral entry. The VP5* protein contains a single long hydrophobic domain (VP5*-HD, residues 385 to 404) at an internal site. In order to address the role of the VP5*-HD in permeabilizing cellular membranes, we analyzed the entry of o-nitrophenyl-beta-D galactopyranoside (ONPG) into cells induced to express VP5* or mutated VP5* polypeptides. Following IPTG (isopropyl-beta-D-thiogalactopyranoside) induction, VP5* and VP5* truncations containing the VP5*-HD permeabilized cells to the entry and cleavage of ONPG, while VP8* and control proteins had no effect on cellular permeability. Expression of VP5* deletions containing residues 265 to 474 or 265 to 404 permeabilized cells; however, C-terminal truncations which remove the conserved GGA (residues 399 to 401) within the HD abolished membrane permeability. Site-directed mutagenesis of the VP5-HD further demonstrated a requirement for residues within the HD for VP5*-induced membrane permeability. Functional analysis of mutant VP5*s indicate that conserved glycines within the HD are required and suggest that a random coiled structure rather than the strictly hydrophobic character of the domain is required for permeability. Expressed VP5* did not alter bacterial growth kinetics or lyse bacteria following induction. Instead, VP5*-mediated size-selective membrane permeability, releasing 376-Da carboxyfluorescein but not 4-kDa fluorescein isothiocyanate-dextran from preloaded liposomes. These findings suggest that the fundamental role for VP5* in the rotavirus entry process may be to expose triple-layered particles to low [Ca](i), which uncoats the virus, rather than to effect the detergent-like lysis of early endosomal membranes. PMID- 10864648 TI - Glycosphingolipids promote entry of a broad range of human immunodeficiency virus type 1 isolates into cell lines expressing CD4, CXCR4, and/or CCR5. AB - Treatment of human osteosarcoma cells, expressing CD4 and various chemokine receptors, with the glucosylceramide synthase inhibitor 1-phenyl-2 hexadecanoylamino-3-morpholino-1-propanol (PPMP), blocked target membrane glycosphingolipid (GSL) biosynthesis and reduced the susceptibility of cells to infection and fusion mediated by envelope glycoproteins from a variety of human immunodeficiency virus type 1 (HIV-1) isolates that utilize CXCR4 and/or CCR5. PPMP treatment of the cell lines did not significantly change the cell surface expression of CD4, CXCR4, and/or CCR5, nor did it alter the chemokine receptor association with CD4. PPMP-treated cells exhibited no changes in chemokine induced Ca(2+) mobilization and chemotaxis. However, massive envelope glycoprotein conformational changes triggered by CD4 and the appropriate chemokine receptor on the target membrane were inhibited when the target cells were treated with PPMP. Addition of various purified GSLs to PPMP-treated target cells showed that for all isolates tested, globotriaosylceramide (Gb3) was the most potent GSL in restoring the fusion susceptibility of target cells with cells expressing HIV-1 envelope glycoproteins; addition of the monosialoganglioside GM3 yielded a slight enhancement of fusion susceptibility. Our data are consistent with the notion that a limited number of specific GSL species serve as crucial elements in organizing gp120-gp41, CD4, and an appropriate chemokine receptor into a membrane fusion complex. PMID- 10864649 TI - Functional analysis of the env open reading frame in human endogenous retrovirus IDDMK(1,2)22 encoding superantigen activity. AB - Mice harbor a family of endogenous retroviruses, the mouse mammary tumor viruses (MMTV), which encode superantigens. These superantigens are responsible for the deletion of T cells expressing certain Vbeta chains of the T-cell receptor in the thymus. Human T cells are able to recognize MMTV-encoded superantigens presented by human major histocompatibility complex class II-positive cells. Owing to this and to the similarity of the human and murine immune systems, it was speculated that human endogenous retroviruses might also code for superantigens. Recently, it was reported that a proviral clone (IDDMK(1,2)22) of the human endogenous retrovirus family HTDV/HERV-K encodes a superantigen. The putative superantigen gene was located within the env region of the virus. Stimulated by these findings, we amplified by PCR and cloned into eucaryotic expression vectors open reading frames (ORFs) which were identical or very similar to IDDMK(1,2)22. When we transfected these vectors into A20 cells, a murine B-cell lymphoma, we were able to demonstrate mRNA expression and protein production. However, we did not find any evidence that the ORF stimulated human or murine T cells in a Vbeta specific fashion, the most prominent feature of superantigens. PMID- 10864650 TI - Poliovirus requires a precise 5' end for efficient positive-strand RNA synthesis. AB - Poliovirus infectious RNA can be synthesized in vitro using phage DNA-dependent RNA-polymerases. These synthetic transcripts contain several extra nucleotides at the 5' end, which are deleted during replication to generate authentic viral genomes. We removed those 5'-end extra nucleotides utilizing a hammerhead ribozyme to produce transcripts with accurate 5' ends. These transcripts replicate substantially more rapidly in cell culture, demonstrating no lag before replication; they also replicate more efficiently in Xenopus laevis oocytes and in in vitro translation-replication cell extracts. In both systems, an exact 5' end is necessary for synthesis of positive-strand RNA but not negative-strand RNA. PMID- 10864651 TI - Characterization of a baculovirus alkaline nuclease. AB - All baculovirus genomes sequenced to date encode a homolog of an alkaline nuclease that has been characterized in the Herpesviridae. In this report we describe the characterization of the alkaline nuclease (AN) homolog of the Autographa californica multinucleocapsid nucleopolyhedrovirus (AcMNPV) (open reading frame 133). His-tagged AN constructs were expressed in recombinant baculoviruses and affinity purified, and then their enzymatic activity was characterized. AN was found to degrade linear DNA at alkaline pH, preferred Mg(2+) over Mn(2+), had optimal activity at 35 degrees C, and did not appear to have a salt requirement. To rule out contamination by the endogenous baculovirus gene product or a cellular enzyme, point mutations were introduced into a highly conserved domain of the gene. These mutations were found to markedly reduce or eliminate most of the activity of the affinity-purified enzyme. An antibody generated against the protein was used to analyze its expression by Western blot analysis. AN was found to be expressed at low levels by 12 h postinfection, with maximal expression at 24 h postinfection. Attempts to generate a virus with this gene inactivated were unsuccessful, suggesting that AN may be encoded by an essential gene. PMID- 10864652 TI - Human papillomavirus type 16 E6 induces self-ubiquitination of the E6AP ubiquitin protein ligase. AB - The E6 protein of the high-risk human papillomaviruses (HPVs) and the cellular ubiquitin-protein ligase E6AP form a complex which causes the ubiquitination and degradation of p53. We show here that HPV16 E6 promotes the ubiquitination and degradation of E6AP itself. The half-life of E6AP is shorter in HPV-positive cervical cancer cells than in HPV-negative cervical cancer cells, and E6AP is stabilized in HPV-positive cancer cells when expression of the viral oncoproteins is repressed. Expression of HPV16 E6 in cells results in a threefold decrease in the half-life of transfected E6AP. E6-mediated degradation of E6AP requires (i) the binding of E6 to E6AP, (ii) the catalytic activity of E6AP, and (iii) activity of the 26S proteasome, suggesting that E6-E6AP interaction results in E6AP self-ubiquitination and degradation. In addition, both in vitro and in vivo experiments indicate that E6AP self-ubiquitination results primarily from an intramolecular transfer of ubiquitin from the active-site cysteine to one or more lysine residues; however, intermolecular transfer can also occur in the context of an E6-mediated E6AP multimer. Finally, we demonstrate that an E6 mutant that is able to immortalize human mammary epithelial cells but is unable to degrade p53 retains its ability to bind and degrade E6AP, raising the possibility that E6 mediated degradation of E6AP contributes to its ability to transform mammalian cells. PMID- 10864653 TI - CCR5 signal transduction in macrophages by human immunodeficiency virus and simian immunodeficiency virus envelopes. AB - The capacity of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelopes to transduce signals through chemokine coreceptors on macrophages was examined by measuring the ability of recombinant envelope proteins to mobilize intracellular calcium stores. Both HIV and SIV envelopes mobilized calcium via interactions with CCR5. The kinetics of these responses were similar to those observed when macrophages were treated with MIP-1beta. Distinct differences in the capacity of envelopes to mediate calcium mobilization were observed. Envelopes derived from viruses capable of replicating in macrophages mobilized relatively high levels of calcium, while envelopes derived from viruses incapable of replicating in macrophages mobilized relatively low levels of calcium. The failure to efficiently mobilize calcium was not restricted to envelopes derived from CXCR4-utilizing isolates but also included envelopes derived from CCR5-utilizing isolates that fail to replicate in macrophages. We characterized one CCR5-utilizing isolate, 92MW959, which entered macrophages but failed to replicate. A recombinant envelope derived from this virus mobilized low levels of calcium. When macrophages were inoculated with 92MW959 in the presence of MIP-1alpha, viral replication was observed, indicating that a CC chemokine mediated signal provided the necessary stimulus to allow the virus to complete its replication cycle. Although the role that envelope-CCR5 signal transduction plays in viral replication is not yet understood, it has been suggested that envelope-mediated signals facilitate early postfusion events in viral replication. The data presented here are consistent with this hypothesis and suggest that the differential capacity of viral envelopes to signal through CCR5 may influence their ability to replicate in macrophages. PMID- 10864654 TI - Sindbis virus entry into cells triggers apoptosis by activating sphingomyelinase, leading to the release of ceramide. AB - Sindbis virus (SV) causes acute encephalomyelitis by infecting and inducing the death of neurons. Induction of apoptosis occurs during virus entry and involves acid-induced conformational changes in the viral surface glycoproteins and sphingomyelin (SM)-dependent fusion of the virus envelope with the endosomal membrane. We have studied neuroblastoma cells to determine how this entry process triggers cell death. Acidic sphingomyelinase was activated during entry followed by activation of neutral sphingomyelinase, SM degradation, and a sustained increase in ceramide. Ceramide-induced apoptosis and SV-induced apoptosis could be inhibited by treatment with Z-VAD-fmk, a caspase inhibitor, and by overexpression of Bcl-2, an antiapoptotic cellular protein. Acid ceramidase, expressed in a recombinant SV, decreased intracellular ceramide and protected cells from apoptosis. The data suggest that acid-induced SM-dependent virus fusion initiates the apoptotic cascade by inducing SM degradation and ceramide release. PMID- 10864655 TI - Human T-cell lymphotropic virus type 1-infected T lymphocytes impair catabolism and uptake of glutamate by astrocytes via Tax-1 and tumor necrosis factor alpha. AB - Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of a chronic progressive myelopathy called tropical spastic paraparesis/HTLV-1 associated myelopathy (TSP/HAM). In this disease, lesions of the central nervous system (CNS) are associated with perivascular infiltration by lymphocytes. We and others have hypothesized that these T lymphocytes infiltrating the CNS may play a prominent role in TSP/HAM. Here, we show that transient contact of human or rat astrocytes with T lymphocytes chronically infected by HTLV-1 impairs some of the major functions of brain astrocytes. Uptake of extracellular glutamate by astrocytes was significantly decreased after transient contact with infected T cells, while the expression of the glial transporters GLAST and GLT-1 was decreased. In two-compartment cultures avoiding direct cell-to-cell contact, similar results were obtained, suggesting possible involvement of soluble factors, such as cytokines and the viral protein Tax-1. Recombinant Tax-1 and tumor necrosis factor alpha (TNF-alpha) decreased glutamate uptake by astrocytes. Tax-1 probably acts by inducing TNF-alpha, as the effect of Tax-1 was abolished by anti-TNF-alpha antibody. The expression of glutamate-catabolizing enzymes in astrocytes was increased for glutamine synthetase and decreased for glutamate dehydrogenase, the magnitudes of these effects being correlated with the level of Tax-1 transcripts. In conclusion, Tax-1 and cytokines produced by HTLV-1-infected T cells impair the ability of astrocytes to manage the steady-state level of glutamate, which in turn may affect neuronal and oligodendrocytic functions and survival. PMID- 10864656 TI - The fusion glycoprotein of human respiratory syncytial virus facilitates virus attachment and infectivity via an interaction with cellular heparan sulfate. AB - Human respiratory syncytial virus (RSV) F glycoprotein (RSV-F) can independently interact with immobilized heparin and facilitate both attachment to and infection of cells via an interaction with cellular heparan sulfate. RSV-glycosaminoglycan (GAG) interactions were evaluated using heparin-agarose affinity chromatography. RSV-F from A2- and B1/cp-52 (cp-52)-infected cell lysates, RSV-F derived from a recombinant vaccinia virus, and affinity-purified F protein all bound to and were specifically eluted from heparin columns. In infectivity inhibition studies, soluble GAGs decreased the infectivity of RSV A2 and cp-52, with bovine lung heparin exhibiting the highest specific activity against both A2 (50% effective dose [ED(50)] = 0.28 +/- 0.11 microg/ml) and cp-52 (ED(50) = 0.55 +/- 0. 14 microg/ml). Furthermore, enzymatic digestion of cell surface GAGs by heparin lyase I and heparin lyase III but not chondroitinase ABC resulted in a significant reduction in cp-52 infectivity. Moreover, bovine lung heparin inhibited radiolabeled A2 and cp-52 virus binding up to 90%. Taken together, these data suggest that RSV-F independently interacts with heparin/heparan sulfate and this type of interaction facilitates virus attachment and infectivity. PMID- 10864657 TI - Replacement of the ectodomains of the hemagglutinin-neuraminidase and fusion glycoproteins of recombinant parainfluenza virus type 3 (PIV3) with their counterparts from PIV2 yields attenuated PIV2 vaccine candidates. AB - We sought to develop a live attenuated parainfluenza virus type 2 (PIV2) vaccine strain for use in infants and young children, using reverse genetic techniques that previously were used to rapidly produce a live attenuated PIV1 vaccine candidate. The PIV1 vaccine candidate, designated rPIV3-1cp45, was generated by substituting the full-length HN and F proteins of PIV1 for those of PIV3 in the attenuated cp45 PIV3 vaccine candidate (T. Tao et al., J. Virol. 72:2955-2961, 1998; M. H. Skiadopoulos et al., Vaccine 18:503-510, 1999). However, using the same strategy, we failed to recover recombinant chimeric PIV3-PIV2 isolate carrying the full-length PIV2 glycoproteins in a wild-type PIV3 backbone. Viable PIV3-PIV2 chimeras were recovered when chimeric HN and F open reading frames (ORFs) rather than complete PIV2 F and HN ORFs were used to construct the full length cDNA. The recovered viruses, designated rPIV3-2CT, in which the PIV2 ectodomain and transmembrane domain were fused to the PIV3 cytoplasmic domain, and rPIV3-2TM, in which the PIV2 ectodomain was fused to the PIV3 transmembrane and cytoplasmic tail domain, possessed similar in vitro and in vivo phenotypes. Thus, it appeared that only the cytoplasmic tail of the HN or F glycoprotein of PIV3 was required for successful recovery of PIV3-PIV2 chimeras. Although rPIV3 2CT and rPIV3-2TM replicated efficiently in vitro, they were moderately to highly attenuated for replication in the respiratory tracts of hamsters, African green monkeys (AGMs), and chimpanzees. This unexpected finding indicated that chimerization of the HN and F proteins of PIV2 and PIV3 itself specified an attenuation phenotype in vivo. Despite this attenuation, these viruses were highly immunogenic and protective against challenge with wild-type PIV2 in hamsters and AGMs, and they represent promising candidates for clinical evaluation as a vaccine against PIV2. These chimeric viruses were further attenuated by the addition of 12 mutations of PIV3cp45 which lie outside of the HN and F genes. The attenuating effects of these mutations were additive with that of the chimerization, and thus inclusion of all or some of the cp45 mutations provides a means to further attenuate the PIV3-PIV2 chimeric vaccine candidates if necessary. PMID- 10864658 TI - Functional significance of the interaction of hepatitis A virus RNA with glyceraldehyde 3-phosphate dehydrogenase (GAPDH): opposing effects of GAPDH and polypyrimidine tract binding protein on internal ribosome entry site function. AB - Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), a cellular enzyme involved in glycolysis, binds specifically to several viral RNAs, but the functional significance of this interaction is uncertain. Both GAPDH and polypyrimidine tract binding protein (PTB) bind to overlapping sites in stem-loop IIIa of the internal ribosome entry site (IRES) of Hepatitis A virus (HAV), a picornavirus. Since the binding of GAPDH destabilizes the RNA secondary structure, we reasoned that GAPDH may suppress the ability of the IRES to direct cap-independent translation, making its effects antagonistic to the translation-enhancing activity of PTB (D. E. Schultz, C. C. Hardin, and S. M. Lemon, J. Biol. Chem. 271:14134-14142, 1996). To test this hypothesis, we constructed plasmids containing a dicistronic transcriptional unit in which the HAV IRES was placed between an upstream GAPDH-coding sequence and a downstream Renilla luciferase (RLuc) sequence. Transfection with this plasmid results in overexpression of GAPDH and in RLuc production as a measure of IRES activity. RLuc activity was compared with that from a control, null-expression plasmid that was identical except for a frameshift mutation within the 5' GAPDH coding sequence. In transfection experiments, GAPDH overexpression significantly suppressed HAV IRES activity in BSC-1 and FRhK-4 cells but not in Huh-7 cells, which have a significantly greater cytoplasmic abundance of PTB. GAPDH suppression of HAV translation was greater with the wild-type HAV IRES than with the IRES from a cell culture-adapted virus (HM175/P16) that has reproducibly higher basal translational activity in BSC-1 cells. Stem-loop IIIa RNA from the latter IRES had significantly lower affinity for GAPDH in filter binding experiments. Thus, the binding of GAPDH to the IRES of HAV suppresses cap-independent viral translation in vivo in African green monkey kidney cells. The enhanced replication capacity of cell culture-adapted HAV in such cells may be due in part to reduced affinity of the viral IRES for GAPDH. PMID- 10864659 TI - Soluble receptor-induced retroviral infection of receptor-deficient cells. AB - Current models of retroviral entry hypothesize that interactions between the host cell receptor(s) and viral envelope protein induce structural changes in the envelope protein that convert it to an active conformation, allowing it to mediate fusion with the membrane. Recent evidence supporting this hypothesis is the demonstration that Tva, the receptor for subgroup A avian sarcoma and leukosis virus (ASLV-A), induces conformational changes in the viral envelope protein. These changes include conversion of the envelope protein to an active, membrane-binding state likely representing a fusogenic conformation. To determine whether binding of the soluble Tva (sTva) receptor was sufficient to activate fully the fusogenic potential of the ASLV-A envelope protein, we have evaluated the ability of ASLV-A to infect receptor-deficient cell lines in the presence of sTva. Soluble receptor efficiently mediated infection of cells devoid of endogenous Tva in a dose-dependent manner, and this infection was dependent absolutely on the addition of sTva. The infectivity of the virus was enhanced dramatically in the presence of the polycationic polymer Polybrene or when centrifugal forces were applied during inoculation, resulting in viral titers comparable to those achieved on cells expressing endogenous receptor. sTva functioned to mediate infection at low concentrations, approaching the estimated binding constant of the receptor and viral envelope protein. These results demonstrate that receptor binding can activate the ASLV-A envelope protein and convert it to a fusogenic conformation competent to mediate the fusion of the viral and cellular membranes. PMID- 10864660 TI - Infection of polarized cultures of human intestinal epithelial cells with hepatitis A virus: vectorial release of progeny virions through apical cellular membranes. AB - Although hepatitis A virus (HAV) is typically transmitted by the fecal-oral route, little is known of its interactions with cells of the gastrointestinal tract. We studied the replication of HAV in polarized cultures of Caco-2 cells, a human cell line which retains many differentiated functions of small intestinal epithelial cells. Virus uptake was 30- to 40-fold more efficient when the inoculum was placed on the apical rather than the basolateral surface of these cells, suggesting a greater abundance of the cellular receptor for HAV on the apical surface. Infection proceeded without cytopathic effect and did not influence transepithelial resistance or the diffusion of inulin across cell monolayers. Nonetheless, there was extensive release of progeny virus, which occurred almost exclusively into apical supernatant fluids (36.4% +/- 12.5% of the total virus yield compared with 0.23% +/- 0.13% release into basolateral fluids). Brefeldin A caused a profound inhibition of HAV replication, but also selectively reduced apical release of virus. These results indicate that polarized human epithelial cell cultures undergo vectorial infection with HAV and that virus release is largely restricted to the apical membrane. Virus release occurs in the absence of cytopathic effect and may involve cellular vesicular transport mechanisms. PMID- 10864661 TI - Characterization of a region of the measles virus hemagglutinin sufficient for its dimerization. AB - Attachment of measles virus (MV) to its cellular receptor is mediated by the viral envelope glycoprotein hemagglutinin (H). H exists at the viral surface as a disulfide-linked dimer which may associate into a tetramer. We aimed to define regions of H essential for its homo-oligomerization. To delineate these more precisely, we have generated a series of H ectodomain truncation mutants and studied their abilities to form both homotypic complexes and heterotypic complexes with full-length H. We define a "minimal unit" which is sufficient for MV H dimerization as that encompassing residues 1 to 151. This unit forms both homodimers and heterodimers with full-length H protein, although neither is transported to the cell surface even in the presence of other MV proteins. We show that cysteine residues at positions 139 and 154 are both critical in mediating covalent dimerization, not only of the truncated H mutants but also of full-length MV H protein. Even those cysteine mutants unable to form covalent intermolecular interactions are biologically active, mediating the formation of syncytia, albeit at a reduced rate. We demonstrate that this impaired capacity to mediate cell-to-cell fusion is based mainly on a reduced transport rate of the mutant molecules to the cell surface, indicating a role for covalent intermolecular interactions in efficient transport of MV H dimers to the cell surface. PMID- 10864662 TI - Effects of amino acid substitutions at position 115 on the fidelity of human immunodeficiency virus type 1 reverse transcriptase. AB - We compared the fidelity of wild-type human immunodeficiency virus type-1 (HIV-1) reverse transcriptase (RT) and two RT mutants, Y115F and Y115V. Although neither mutation had a large effect on the overall fidelity of the enzyme, both mutations altered the spectrum of mutations and the precise nature of the mutational hot spots. The effects of Y115V were greater than those of Y115F. When we compared the behavior of the wild-type enzyme with published data, we found that, in contrast to what has been published, misalignment/slippage could account for only a small fraction of the mutations we observed. We also found that a preponderance of the mutations (both transitions and transversions) resulted in the insertion of an A. Because we were measuring DNA-dependent DNA synthesis (plus-strand synthesis), this bias could contribute to the A-rich nature of the HIV-1 genome. PMID- 10864663 TI - Enhanced infectivity of an R5-tropic simian/human immunodeficiency virus carrying human immunodeficiency virus type 1 subtype C envelope after serial passages in pig-tailed macaques (Macaca nemestrina). AB - The increasing prevalence of human immunodeficiency virus type 1 (HIV-1) subtype C infection worldwide calls for efforts to develop a relevant animal model for evaluating strategies against the transmission of the virus. A chimeric simian/human immunodeficiency virus (SHIV), SHIV(CHN19), was generated with a primary, non-syncytium-inducing HIV-1 subtype C envelope from a Chinese strain in the background of SHIV(33). Unlike R5-tropic SHIV(162), SHIV(CHN19) was not found to replicate in rhesus CD4(+) T lymphocytes. SHIV(CHN19) does, however, replicate in CD4(+) T lymphocytes of pig-tailed macaques (Macaca nemestrina). The observed replication competence of SHIV(CHN19) requires the full tat/rev genes and partial gp41 region derived from SHIV(33). To evaluate in vivo infectivity, SHIV(CHN19) was intravenously inoculated, at first, into two pig-tailed and two rhesus macaques. Although all four animals became infected, the virus replicated preferentially in pig-tailed macaques with an earlier plasma viral peak and a faster seroconversion. To determine whether in vivo adaptation would enhance the infectivity of SHIV(CHN19), passages were carried out serially in three groups of two pig-tailed macaques each, via intravenous blood-bone marrow transfusion. The passages greatly enhanced the infectivity of the virus as shown by the increasingly elevated viral loads during acute infection in animals with each passage. Moreover, the doubling time of plasma virus during acute infection became much shorter in passage 4 (P4) animals (0.2 day) in comparison to P1 animals (1 to 2 days). P2 to P4 animals all became seropositive around 2 to 3 weeks postinoculation and had a decline in CD4/CD8 T-cell ratio during the early phase of infection. In P4 animals, a profound depletion of CD4 T cells in the lamina propria of the jejunum was observed. Persistent plasma viremia has been found in most of the infected animals with sustained viral loads ranging from 10(3) to 10(5) per ml up to 6 months postinfection. Serial passages did not change the viral phenotype as confirmed by the persistence of the R5 tropism of SHIV(CHN19) isolated from P4 animals. In addition, the infectivity of SHIV(CHN19) in rhesus peripheral blood mononuclear cells was also increased after in vivo passages. Our data indicate that SHIV(CHN19) has adapted well to grow in macaque cells. This established R5-tropic SHIV(CHN19)/macaque model would be very useful for HIV-1 subtype C vaccine and pathogenesis studies. PMID- 10864664 TI - The Rous sarcoma virus long terminal repeat promoter is regulated by TFII-I. AB - Many viral genes contain core promoters with two basal control elements, the TATA box and the pyrimidine-rich initiator (Inr). However, the molecular mechanisms involved in transcription initiation from composite core promoters (TATA(+) Inr(+)) containing Inr elements are unclear. The Rous sarcoma virus (RSV) long terminal repeat (LTR) contains a transcriptionally potent enhancer and core promoter composed of a TATA box and an Inr-like sequence, termed the transcription start site core (TSSC). Previously we demonstrated that the TSSC binds the multifunctional Inr-binding protein YY1. Here we present evidence that the TSSC also binds the multifunctional transcription factor TFII-I and that both TFII-I and YY1 are required for RSV LTR transcriptional activity. Gel shift assays using anti-TFII-I antibody show that TFII-I is present in a protein complex that specifically binds to the TSSC. Mutations in the TSSC that reduce TFII-I binding also reduce RSV LTR enhancer and promoter activity. Transient transfection assays demonstrate that TFII-I transactivates the RSV LTR from ca. fourfold (basal) to ca. sevenfold (enhanced) in both human and natural host cell lines. Importantly, the activity of the TSSC element can be attributed to the binding activity of TFII-I and the YY1 protein, since mutation of each of these binding sites within the TSSC element abolishes all viral expression as demonstrated by transient-transfection assays. Taken together, these data demonstrate that expression of RSV viral mRNA is dependent on both TFII-I and YY1. PMID- 10864665 TI - Phosphorylation of human immunodeficiency virus type 1 Vpr regulates cell cycle arrest. AB - Human immunodeficiency virus type 1 (HIV-1) Vpr regulates nuclear transport of the viral preintegration complex, G(2) cell cycle arrest, and transcriptional transactivation. We asked whether phosphorylation could affect Vpr activity. Vpr was found to be phosphorylated on serine residues in transiently transfected and infected cells. Residues 79, 94, and 96 were all found to be phosphorylated, as assessed by alanine mutations. Mutation of Ser-79 to Ala abrogated effects of Vpr on cell cycle progression, whereas mutation of Ser-94 and Ser-96 had no effect. Simultaneous mutation of all three Vpr serine residues attenuated HIV-1 replication in macrophages, whereas single and double Ser mutations had no effect. PMID- 10864666 TI - 3'-End stem-loops of the subviral RNAs associated with turnip crinkle virus are involved in symptom modulation and coat protein binding. AB - Many plant RNA viruses are associated with one or more subviral RNAs. Two subviral RNAs, satellite RNA C (satC) and defective interfering RNA G (diG) intensify the symptoms of their helper, turnip crinkle virus (TCV). However, when the coat protein (CP) of TCV was replaced with that of the related Cardamine chlorotic fleck virus (CCFV), both subviral RNAs attenuated symptoms of the hybrid virus TCV-CP(CCFV). In contrast, when the translation initiation codon of the TCV CP was altered to ACG and reduced levels of CP were synthesized, satC attenuated symptoms while diG neither intensified nor attenuated symptoms. The determinants for this differential symptom modulation were previously localized to the 3'-terminal 100 bases of the subviral RNAs, which contain six positional differences (Q. Kong, J.-W. Oh, C. D. Carpenter, and A. E. Simon, Virology 238:478-485, 1997). In the current study, we have determined that certain sequences within the 3'-terminal stem-loop structures of satC and diG, which also serve as promoters for complementary strand synthesis, are critical for symptom modulation. Furthermore, the ability to attenuate symptoms was correlated with weakened binding of TCV CP to the hairpin structure. PMID- 10864667 TI - Analysis of the transmembrane domain of influenza virus neuraminidase, a type II transmembrane glycoprotein, for apical sorting and raft association. AB - Influenza virus neuraminidase (NA), a type II transmembrane protein, is directly transported to the apical plasma membrane in polarized MDCK cells. Previously, it was shown that the transmembrane domain (TMD) of NA provides a determinant(s) for apical sorting and raft association (A. Kundu, R. T. Avalos, C. M. Sanderson, and D. P. Nayak, J. Virol. 70:6508-6515, 1996). In this report, we have analyzed the sequences in the NA TMD involved in apical transport and raft association by making chimeric TMDs from NA and human transferring receptor (TR) TMDs and by mutating the NA TMD sequences. Our results show that the COOH-terminal half of the NA TMD (amino acids [aa] 19 to 35) was significantly involved in raft association, as determined by Triton X-100 (TX-100) resistance. However, in addition, the highly conserved residues at the extreme NH(2) terminus of the NA TMD were also critical for TX-100 resistance. On the other hand, 19 residues (aa 9 to 27) at the NH(2) terminus of the NA TMD were sufficient for apical sorting. Amino acid residues 14 to 18 and 27 to 31 had the least effect on apical transport, whereas mutations in the amino acid residues 11 to 13, 23 to 26, and 32 to 35 resulted in altered polarity for the mutant proteins. These results indicated that multiple regions in the NA TMD were involved in apical transport. Furthermore, these results support the idea that the signals for apical sorting and raft association, although residing in the NA TMD, are not identical and vary independently and that the NA TMD also possesses an apical determinant(s) which can interact with apical sorting machineries outside the lipid raft. PMID- 10864668 TI - Trypsin-induced structural transformation in aquareovirus. AB - Aquareovirus, a member of the family Reoviridae, is a large virus with multiple capsid layers surrounding a genome composed of 11 segments of double-stranded RNA. Biochemical studies have shown that treatment with the proteolytic agent trypsin significantly alters the infectivity of the virus. The most infectious stage of the virus is produced by a 5-min treatment with trypsin. However, prolonged trypsin treatment almost completely abolishes the infectivity. We have used three-dimensional electron cryomicroscopy to gain insight into the structural basis of protease-induced alterations in infectivity by examining the structural changes in the virion at various time intervals of trypsin treatment. Our data show that after 5 min of trypsinization, projection-like spikes made of VP7 (35 kDa), associated with the underlying trimeric subunits, are completely removed. Concurrent with the removal of VP7, conformational changes are observed in the trimeric subunit composed of putative VP5 (71 kDa). The removal of VP7 and the accompanied structural changes may expose regions in the putative VP5 important for cell entry processes. Prolonged trypsinization not only entirely removes the outer capsid layer, producing the poorly infectious core particle, but also causes significant conformational changes in the turret protein. These changes result in shortening of the turret and narrowing of its central channel. The turret, as in orthoreoviruses, is likely to play a major role in the capping and translocation of mRNA during transcription, and the observed conformational flexibility in the turret protein may have implications in rendering the particle transcriptionally active or inactive. PMID- 10864669 TI - Cowpea mosaic virus infection induces a massive proliferation of endoplasmic reticulum but not Golgi membranes and is dependent on de novo membrane synthesis. AB - Replication of cowpea mosaic virus (CPMV) is associated with small membranous vesicles that are induced upon infection. The effect of CPMV replication on the morphology and distribution of the endomembrane system in living plant cells was studied by expressing green fluorescent protein (GFP) targeted to the endoplasmic reticulum (ER) and the Golgi membranes. CPMV infection was found to induce an extensive proliferation of the ER, whereas the distribution and morphology of the Golgi stacks remained unaffected. Immunolocalization experiments using fluorescence confocal microscopy showed that the proliferated ER membranes were closely associated with the electron-dense structures that contain the replicative proteins encoded by RNA1. Replication of CPMV was strongly inhibited by cerulenin, an inhibitor of de novo lipid synthesis, at concentrations where the replication of the two unrelated viruses alfalfa mosaic virus and tobacco mosaic virus was largely unaffected. These results suggest that proliferating ER membranes produce the membranous vesicles formed during CPMV infection and that this process requires continuous lipid biosynthesis. PMID- 10864670 TI - A cytoplasmic RNA vector derived from nontransmissible Sendai virus with efficient gene transfer and expression. AB - We have recovered a virion from defective cDNA of Sendai virus (SeV) that is capable of self-replication but incapable of transmissible-virion production. This virion delivers and expresses foreign genes in infected cells, and this is the first report of a gene expression vector derived from a defective viral genome of the Paramyxoviridae. First, functional ribonucleoprotein complexes (RNPs) were recovered from SeV cloned cDNA defective in the F (envelope fusion protein) gene, in the presence of plasmids expressing nucleocapsid protein and viral RNA polymerase. Then the RNPs were transfected to the cells inducibly expressing F protein. Virion-like particles thus obtained had a titer of 0.5 x 10(8) to 1. 0 x 10(8) cell infectious units/ml and contained F-defective RNA genome. This defective vector amplified specifically in an F-expressing packaging cell line in a trypsin-dependent manner but did not spread to F-nonexpressing cells. This vector infected and expressed an enhanced green fluorescent protein reporter gene in various types of animal and human cells, including nondividing cells, with high efficiency. These results suggest that this vector has great potential for use in human gene therapy and vaccine delivery systems. PMID- 10864671 TI - Formation of the poliovirus replication complex requires coupled viral translation, vesicle production, and viral RNA synthesis. AB - Poliovirus (PV) infection induces the rearrangement of intracellular membranes into characteristic vesicles which assemble into an RNA replication complex. To investigate this transformation, endoplasmic reticulum (ER) membranes in HeLa cells were modified by the expression of different cellular or viral membrane binding proteins. The membrane-binding proteins induced two types of membrane alterations, i.e., extended membrane sheets and vesicles similar to those found during a PV infection. Cells expressing membrane-binding proteins were superinfected with PV and then analyzed for virus replication, location of membranes, viral protein, and RNA by immunofluorescence and fluorescent in situ hybridization. Cultures expressing cellular or viral membrane-binding proteins, but not those expressing soluble proteins, showed a markedly reduced ability to support PV replication as a consequence of the modification of ER membranes. The altered membranes, regardless of their morphology, were not used for the formation of viral replication complexes during a subsequent PV infection. Specifically, membrane sheets were not substrates for PV-induced vesicle formation, and, surprisingly, vesicles induced by and carrying one or all of the PV replication proteins did not contribute to replication complexes formed by the superinfecting PV. The formation of replication complexes required active viral RNA replication. The extensive alterations induced by membrane-binding proteins in the ER resulted in reduced viral protein synthesis, thus affecting the number of cells supporting PV multiplication. Our data suggest that a functional replication complex is formed in cis, in a coupled process involving viral translation, membrane modification and vesicle budding, and viral RNA synthesis. PMID- 10864672 TI - Norwalk virus open reading frame 3 encodes a minor structural protein. AB - Norwalk virus (NV) is a causative agent of acute epidemic nonbacterial gastroenteritis in humans. The inability to cultivate NV has required the use of molecular techniques to examine the genome organization and functions of the viral proteins. The function of the NV protein encoded by open reading frame 3 (ORF 3) has been unknown. In this paper, we report the characterization of the NV ORF 3 protein expressed in a cell-free translation system and in insect cells and show its association with recombinant virus-like particles (VLPs) and NV virions. Expression of the ORF 3 coding region in rabbit reticulocyte lysates resulted in the production of a single protein with an apparent molecular weight of 23,000 (23K protein), which is not modified by N-linked glycosylation. The ORF 3 protein was expressed in insect cells by using two different baculovirus recombinants; one recombinant contained the entire 3' end of the genome beginning with the ORF 2 coding sequences (ORFs 2+3), and the second recombinant contained ORF 3 alone. Expression from the construct containing both ORF 2 and ORF 3 resulted in the expression of a single protein (23K protein) detected by Western blot analysis with ORF 3-specific peptide antisera. However, expression from a construct containing only the ORF 3 coding sequences resulted in the production of multiple forms of the ORF 3 protein ranging in size from 23,000 to 35,000. Indirect immunofluorescence studies using an ORF 3 peptide antiserum showed that the ORF 3 protein is localized to the cytoplasm of infected insect cells. The 23K ORF 3 protein was consistently associated with recombinant VLPs purified from the media of insect cells infected with a baculovirus recombinant containing the entire 3' end of the NV genome. Western blot analysis of NV purified from the stools of NV infected volunteers revealed the presence of a 35K protein as well as multiple higher-molecular-weight bands specifically recognized by an ORF 3 peptide antiserum. These results indicate that the ORF 3 protein is a minor structural protein of the virion. PMID- 10864673 TI - Continued circulation in China of highly pathogenic avian influenza viruses encoding the hemagglutinin gene associated with the 1997 H5N1 outbreak in poultry and humans. AB - Since the outbreak in humans of an H5N1 avian influenza virus in Hong Kong in 1997, poultry entering the live-bird markets of Hong Kong have been closely monitored for infection with avian influenza. In March 1999, this monitoring system detected geese that were serologically positive for H5N1 avian influenza virus, but the birds were marketed before they could be sampled for virus. However, viral isolates were obtained by swabbing the cages that housed the geese. These samples, known collectively as A/Environment/Hong Kong/437/99 (A/Env/HK/437/99), contained four viral isolates, which were compared to the 1997 H5N1 Hong Kong isolates. Analysis of A/Env/HK/437/99 viruses revealed that the four isolates are nearly identical genetically and are most closely related to A/Goose/Guangdong/1/96. These isolates and the 1997 H5N1 Hong Kong viruses encode common hemagglutinin (H5) genes that have identical hemagglutinin cleavage sites. Thus, the pathogenicity of the A/Env/HK/437/99 viruses was compared in chickens and in mice to evaluate the potential for disease outbreaks in poultry and humans. The A/Env/HK/437/99 isolates were highly pathogenic in chickens but caused a longer mean death time and had altered cell tropism compared to A/Hong Kong/156/97 (A/HK/156/97). Like A/HK/156/97, the A/Env/HK/437/99 viruses replicated in mice and remained localized to the respiratory tract. However, the A/Env/HK/437/99 isolates caused only mild pathological lesions in these tissues and no clinical signs of disease or death. As a measure of the immune response to these viruses, transforming growth factor beta levels were determined in the serum of infected mice and showed elevated levels for the A/Env/HK/437/99 viruses compared to the A/HK/156/97 viruses. This study is the first to characterize the A/Env/HK/437/99 viruses in both avian and mammalian species, evaluating the H5 gene from the 1997 Hong Kong H5N1 isolates in a different genetic background. Our findings reveal that at least one of the avian influenza virus genes encoded by the 1997 H5N1 Hong Kong viruses continues to circulate in mainland China and that this gene is important for pathogenesis in chickens but is not the sole determinant of pathogenicity in mice. There is evidence that H9N2 viruses, which have internal genes in common with the 1997 H5N1 Hong Kong isolates, are still circulating in Hong Kong and China as well, providing a heterogeneous gene pool for viral reassortment. The implications of these findings for the potential for human disease are discussed. PMID- 10864674 TI - Trafficking of varicella-zoster virus glycoprotein gI: T(338)-dependent retention in the trans-Golgi network, secretion, and mannose 6-phosphate-inhibitable uptake of the ectodomain. AB - The trans-Golgi network (TGN) is putatively the site where varicella-zoster virus is enveloped. gE is targeted to the TGN by selective retrieval from the plasmalemma in response to signaling sequences in its endodomain. gI lacks these sequences but forms a complex with gE. We now find that gI is targeted to the TGN and plasma membrane when expressed in Cos-7 cells; nevertheless, surface labeling revealed that gI is not retrieved from the plasma membrane. TGN targeting of gI depended on the T(338) of its endodomain and was lost when T(338) was deleted or mutated to A, S, or D. The endodomain of gI was sufficient, if it contained T(338), to target a fusion protein containing the ectodomain of the human interleukin-2 receptor to the TGN. A truncated protein consisting only of the gI ectodomain was secreted and taken up by nontransfected cells. This uptake of the secreted gI ectodomain was blocked by mannose 6-phosphate. Following cotransfection, both gI and gE were retrieved to the TGN from the plasma membrane in 26.7% of cells, neither gI nor gE was internalized in 18.3%, and gE was retrieved to the TGN while gI remained at the plasma membrane in 55%. We suggest that the T(338) of its endodomain is necessary to retain gI in the TGN; moreover, because gI and gE interact, the signaling sequences of each glycoprotein reinforce one another in ensuring that both glycoproteins are concentrated in the TGN yet remain on the cell surface. PMID- 10864675 TI - Functional implications of the human T-lymphotropic virus type 1 transmembrane glycoprotein helical hairpin structure. AB - Retrovirus entry into cells follows receptor binding by the surface-exposed envelope glycoprotein (Env) subunit (SU), which triggers the membrane fusion activity of the transmembrane (TM) protein. TM protein fragments expressed in the absence of SU adopt helical hairpin structures comprising a central coiled coil, a region of chain reversal containing a disulfide-bonded loop, and a C-terminal segment that packs onto the exterior of the coiled coil in an antiparallel manner. Here we used in vitro mutagenesis to test the functional role of structural elements observed in a model helical hairpin, gp21 of human T lymphotropic virus type 1. Membrane fusion activity requires the stabilization of the N and C termini of the central coiled coil by a hydrophobic N cap and a small hydrophobic core, respectively. A conserved Gly-Gly hinge motif preceding the disulfide-bonded loop, a salt bridge that stabilizes the chain reversal region, and interactions between the C-terminal segment and the coiled coil are also critical for fusion activity. Our data support a model whereby the chain reversal region transmits a conformational signal from receptor-bound SU to induce the fusion-activated helical hairpin conformation of the TM protein. PMID- 10864676 TI - The human papillomavirus type 16 E7 oncogene is required for the productive stage of the viral life cycle. AB - The production of the human papillomavirus type 16 (HPV-16) is intimately tied to the differentiation of the host epithelium that it infects. Infection occurs in the basal layer of the epithelium at a site of wounding, where the virus utilizes the host DNA replication machinery to establish itself as a low-copy-number episome. The productive stage of the HPV-16 life cycle occurs in the postmitotic suprabasal layers of the epithelium, where the virus amplifies its DNA to high copy number, synthesizes the capsid proteins (L1 and L2), encapsidates the HPV-16 genome, and releases virion particles as the upper layer of the epithelium is shed. Papillomaviruses are hypothesized to possess a mechanism to overcome the block in DNA synthesis that occurs in the differentiated epithelial cells, and the HPV-16 E7 oncoprotein has been suggested to play a role in this process. To determine whether E7 plays a role in the HPV-16 life cycle, an E7-deficient HPV 16 genome was created by inserting a translational termination linker (TTL) in the E7 gene of the full HPV-16 genome. This DNA was transfected into an immortalized human foreskin keratinocyte cell line shown previously to support the HPV-16 life cycle, and stable cell lines were obtained that harbored the E7 deficient HPV-16 genome episomally, the state of the genome found in normal infections. By culturing these cells under conditions which promote the differentiation of epithelial cells, we found E7 to be necessary for the productive stage of the HPV-16 life cycle. HPV-16 lacking E7 failed to amplify its DNA and expressed reduced amounts of the capsid protein L1, which is required for virus production. E7 appears to create a favorable environment for HPV-16 DNA synthesis by perturbing the keratinocyte differentiation program and inducing the host DNA replication machinery. These data demonstrate that E7 plays an essential role in the papillomavirus life cycle. PMID- 10864677 TI - Human papillomavirus type 33 E7 peptides presented by HLA-DR*0402 to tumor infiltrating T cells in cervical cancer. AB - Several characteristics make human papillomavirus (HPV) amenable to vaccination. Anti-HPV-directed vaccines are based on the observation that HPV E6 and E7 oncoproteins are constitutively expressed in HPV-positive cervical cancer and may serve as tumor rejection antigens. Five HPV types (16, 18, 31, 33, and 45) account for 80% of cervical cancer. Until now, the type of immune response capable of mediating an effective antitumor response has not been defined. In order to define the anticancer-directed immune response in situ, we characterized CD4(+) and CD8(+) sorted T cells from peripheral blood lymphocytes, freshly harvested tumor tissue, and tumor-infiltrating lymphocytes (TIL) from a patient with cervical cancer. The HLA-DR-restricted CD4(+) T-cell receptor VB16-, VA10-, VA21-, and VA22-positive CD4(+) T-cell line derived from TIL recognizes autologous HLA-DR*0402(+) (HPV33(+)) cervical cancer cells, as determined by gamma interferon secretion. Testing of different peptides spanning the E7 gene revealed that the HPV33(73-87) peptide ASDLRTIQQLLMGTV represents the immunodominant epitope which can also be presented by the DR*0401 allele to TIL. Such major histocompatibility complex class II-presented peptides represent attractive candidates to augment T-cell responses directed against autologous tumor cells. PMID- 10864678 TI - Mutations in the E1 hydrophobic domain of rubella virus impair virus infectivity but not virus assembly. AB - Rubella virus (RV) virions contain three structural proteins, a capsid protein that interacts with viral genomic RNA to form a nucleocapsid and two membrane glycoproteins, E2 and E1. We found that substitution of either an aspartic acid residue at Gly93 (G93D) or a glycine residue at Pro104 (P104G) in the internal hydrophobic domain of E1 affected virus infectivity but not virus assembly. Viruses carrying G93D and P104G mutations had impaired infectivity, reduced 1,000 fold and 10-fold, respectively. A revertant was isolated from the G93D mutant. Sequencing analysis showed that the substituted aspartic acid residue in G93D mutant had reverted to the original glycine residue, suggesting the involvement of Gly93 in membrane fusion during viral entry. PMID- 10864679 TI - Recovery of pathogenic measles virus from cloned cDNA. AB - Reverse genetics technology so far established for measles virus (MeV) is based on the Edmonston strain, which was isolated several decades ago, has been passaged in nonlymphoid cell lines, and is no longer pathogenic in monkey models. On the other hand, MeVs isolated and passaged in the Epstein-Barr virus transformed marmoset B-lymphoblastoid cell line B95a would retain their original pathogenicity (F. Kobune et al., J. Virol. 64:700-705, 1990). Here we have developed MeV reverse genetics systems based on the highly pathogenic IC-B strain isolated in B95a cells. Infectious viruses were successfully recovered from the cloned cDNA of IC-B strain by two different approaches. One was simple cotransfection of B95a cells, with three plasmids each encoding the nucleocapsid (N), phospho (P), or large (L) protein, respectively, and their expression was driven by the bacteriophage T7 RNA polymerase supplied by coinfecting recombinant vaccinia virus vTF7-3. The second approach was transfection with the L-encoding plasmid of a helper cell line constitutively expressing the MeV N and P proteins and the T7 polymerase (F. Radecke et al., EMBO J. 14:5773-5784, 1995) on which B95a cells were overlaid. Virus clones recovered by both methods possessed RNA genomes identical to that of the parental IC-B strain and were indistinguishable from the IC-B strain with respect to growth phenotypes in vitro and the clinical course and histopathology of experimentally infected cynomolgus monkeys. Thus, the systems developed here could be useful for studying viral gene functions in the context of the natural course of MeV pathogenesis. PMID- 10864680 TI - Quantitative analysis of the antiviral activity of CD8(+) T cells from human immunodeficiency virus-positive asymptomatic patients with different rates of CD4(+) T-cell decrease. AB - We have measured in 22 asymptomatic human immunodeficiency virus type 1-infected patients (10 rapid progressors and 12 slow progressors) the proviral load of CD4(+) T cells homogeneously superinfected by the same dose of a non-syncytium inducing virus in the presence or in the absence of autologous CD8(+) T cells. We demonstrated that the antiviral activity of CD8(+) T cells was highly predictive of the rate of peripheral CD4(+) T-cell decline. PMID- 10864681 TI - The bfl-1 gene is transcriptionally upregulated by the Epstein-Barr virus LMP1, and its expression promotes the survival of a Burkitt's lymphoma cell line. AB - The recently identified bfl-1 gene (also known as A1 or GRS), a homologue of bcl 2, encodes an antiapoptotic protein that suppresses apoptosis induced by the p53 tumor suppressor protein and exhibits proliferative and potent cooperative transforming activities. We show that elevated levels of bfl-1 mRNA are a feature of Epstein-Barr virus (EBV)-immortalized B-cell lines and Burkitt's lymphoma cell lines expressing the full spectrum of EBV latent proteins. Using an EBV-negative Burkitt's lymphoma cell line in which the expression of EBV latent membrane protein 1 (LMP1) is inducibly regulated by tetracycline, we demonstrate that LMP1 expression coincides with a dramatic increase in the level of bfl-1 mRNA. Also in this system, an increase in the level of Bcl-2 protein was seen to occur earlier than that of bcl-2 mRNA, suggesting that both transcriptional and translational mechanisms are involved in the control of Bcl-2 expression by LMP-1. We show that elevated bfl-1 mRNA stability can contribute to this effect of LMP-1, thus providing evidence of a novel mechanism of gene regulation by this EBV protein. Upregulation of bfl-1 by LMP1 was not observed in the T-cell line Jurkat or the epithelial cell line C33A. Ectopic expression of Bfl-1 in an EBV-positive cell line exhibiting a latency type I infection protects against apoptosis induced by growth factor deprivation, thereby providing a functional role for Bfl-1 in this cellular context and adding Bfl-1 to the list of antiapoptotic proteins whose expression is modulated by EBV. This is the first report of the regulation of bfl 1 expression by a viral protein, and this novel finding may thus represent an important link between the EBV oncoprotein LMP1 and its cellular growth transforming properties. PMID- 10864683 TI - Wild-type and YMDD mutant murine leukemia virus reverse transcriptases are resistant to 2',3'-dideoxy-3'-thiacytidine. AB - The antiretroviral nucleoside analog 2',3'-dideoxy-3'-thiacytidine (3TC) is a potent inhibitor of wild-type human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). A methionine-to-valine or methionine-to-isoleucine substitution at residue 184 in the HIV-1 YMDD motif, which is located at the RT active site, leads to a high level of resistance to 3TC. We sought to determine whether 3TC can inhibit the replication of wild-type murine leukemia virus (MLV), which contains V223 at the YVDD active site motif of the MLV RT, and of the V223M, V223I, V223A, and V223S mutant RTs. Surprisingly, the wild type and all four of the V223 mutants of MLV RT were highly resistant to 3TC. These results indicate that determinants outside the YVDD motif of MLV RT confer a high level of resistance to 3TC. Therefore, structural differences among similar RTs might result in widely divergent sensitivities to antiretroviral nucleoside analogs. PMID- 10864682 TI - Novel Tat-encoding bicistronic human immunodeficiency virus type 1-based gene transfer vectors for high-level transgene expression. AB - We describe bicistronic single-exon Tat (72-amino-acid Tat [Tat72])- and full length Tat (Tat86)-encoding gene transfer vectors based on human immunodeficiency virus type 1 (HIV-1). We created versions of these vectors that were rendered Rev independent by using the constitutive transport element (CTE) from Mason-Pfizer monkey virus (MPMV). Tat72-encoding vectors performed better than Tat86 expressing vectors in gene transfer experiments. CTE-containing vectors, produced in a Rev-independent packaging system, had gene transfer efficiencies nearly equivalent to those produced using a combination RNA transport (CTE and Rev-Rev response element)-based packaging system. The Tat72-encoding vectors could be efficiently transduced into a variety of cell types, showed higher levels of transgene expression than vectors with the simian cytomegalovirus immediate-early or the simian virus 40 early promoter, and provide an alternative to HIV-1 vectors with internal promoters. PMID- 10864684 TI - Functional analysis of the CD4(+) T-cell response to Epstein-Barr virus: T-cell mediated activation of resting B cells and induction of viral BZLF1 expression. AB - In contrast to the major role played by Epstein-Barr virus (EBV)-specific CD8(+) cytotoxic T-cell responses in immunosurveillance, recent studies have offered the apparently paradoxical suggestion that development of EBV-driven human B-cell lymphoproliferative disorders and tumors in SCID/hu mice is dependent on the presence of T cells, in particular CD4(+) T cells. This study presents a functional analysis of the CD4(+) T-cell response to EBV and shows that while CD4(+) T cells may be cytotoxic, they also express Th2 cytokines and CD40 ligand (gp39) and possess B-cell helper function. We show that EBV-specific CD4(+) T cells can provide non-HLA-restricted help for activation of resting B cells via a gp39-CD40-dependent pathway and are able to induce expression of BZLF1, a viral lytic cycle transactivator in latently infected resting B cells, ultimately resulting in rapid outgrowth of transformed B-cell colonies. These results support the proposal that CD4(+) T cells may play a key role in reactivation of latent EBV infection and may thus contribute to the pathogenesis of EBV-driven lymphoproliferative disorders. PMID- 10864685 TI - Gamma interferon impedes the establishment of herpes simplex virus type 1 latent infection but has no impact on its maintenance or reactivation in mice. AB - Murine models of gamma interferon (IFN-gamma) deficiency demonstrate the role of this cytokine in attenuating acute herpes simplex virus (HSV) disease; however, the effect of IFN-gamma on the establishment and maintenance of neuronal latency and viral reactivation is not known. Using the IFN-gamma knockout (GKO) model of IFN-gamma deficiency and sensitive quantitative PCR methods, we show that IFN gamma significantly reduces the ganglion content of latent HSV-1 in BALB/c mice, which in turn delays viral time to reactivation following UV irradiation. Similar effects were not seen in the C57BL/6 strain. These results indicate that IFN gamma significantly attenuates latent HSV infection in the mouse model of ocular infection but has no impact on the maintenance of latency or virus reactivation. PMID- 10864686 TI - Adenovirus late gene expression does not require a Rev-like nuclear RNA export pathway. AB - Adenovirus late mRNA export is facilitated by viral early proteins of 55 and 34 kDa. The 34-kDa protein contains a leucine-rich nuclear export signal (NES) similar to that of the human immunodeficiency virus Rev protein. It was proposed that the 34-kDa protein might facilitate the export of adenovirus late mRNA through a Rev-like NES-mediated export pathway. We have tested the role of NES mediated RNA export during adenovirus infection, and we find that it is not essential for the expression of adenovirus late genes. PMID- 10864687 TI - CD4-independent infection of two CD4(-)/CCR5(-)/CXCR4(+) pre-T-cell lines by human and simian immunodeficiency viruses. AB - The infection of CD4-negative cells by variants of tissue culture-adapted human immunodeficiency virus type 1 (HIV-1) or HIV-2 strains has been shown to be mediated by the CXCR4 coreceptor. Here we show that two in vitro-established CD4( )/CCR5(-)/CXCR4(+) human pre-T-cell lines (A3 and A5) can be productively infected by wild-type laboratory-adapted T-cell-tropic HIV-1 and HIV-2 strains in a CD4-independent, CXCR4-dependent fashion. Despite the absence of CCR5 expression, A3 and A5 cells were susceptible to infection by the simian immunodeficiency viruses SIVmac239 and SIVmac316. Thus, at least in A3 and A5 cells, one or more of the chemokine receptors can efficiently support the entry of HIV and SIV isolates in the absence of CD4. These findings suggest that to infect cells of different compartments, HIV and SIV could have evolved in vivo to bypass CD4 and to interact directly with an alternative receptor. PMID- 10864688 TI - Inhibition of human immunodeficiency virus type 1 replication in primary CD4(+) T lymphocytes, monocytes, and dendritic cells by cytotoxic T lymphocytes. AB - We demonstrate that human immunodeficiency virus type 1 (HIV-1)-specific CD8(+) cytotoxic T lymphocytes (CTL) suppress HIV-1 replication in primary lymphocytes, monocytes, and dendritic cells individually. Viral inhibition is significantly diminished in lymphocyte-dendritic cell clusters, suggesting that these clusters in vivo could be sites where viral replication is more difficult to control by CTL. PMID- 10864689 TI - [Anal fistulae and abscesses: diagnostic and therapeutic aspects, excepting Crohn's disease]. AB - Anal fistulae are an often neglected and underevaluated disease. They are painful and invalidating. Surgeons must be aware of the pathophysiological aspects to achieve successful treatment. The anatomical classification is established to better understand anal anatomy and physiology of anal abscessess and fistulae. The Diagnosis of a perianal abscess is usually easy except in case of deep abscess. Clinical signs of chronic fisula may be misleading. Modern imaging (MRI and endoscopic ultrasonography) may be useful to detail the fistular anatomy in difficult cases. Several operative procedures have been proposed to treat anal fistulae and abscesses. Besides old procedures such as fistulotomy, cutting or draining, seton, we can also mention recently proposed preservative sphincter surgery. This new concept is believed to improve wound healing and decrease functional deficiency. Particularly, the rectal flap seems to be attractive but its superiority has not been proven with a randomized trial. So far, our preference goes to the well-known procedures such as prolonged seton drainage and/or slow cutting seton. PMID- 10864690 TI - Pains that can't be measured. PMID- 10864691 TI - Use of a taxonomy to describe parish nurse practice with older adults. AB - Parish nursing in a new model of health care delivery in which practitioners build partnerships to extend health care from institutions into the community. The study discussed in this article focused on the applicability of the North American Nurses Diagnosis Association taxonomy and the Nursing Intervention Classification to describe parish nurse practice with older adults. Findings from this study may help develop a framework for the practice. PMID- 10864692 TI - Herpes zoster and postherpetic neuralgia in the elderly. AB - This article describes herpes zoster (HZ), its cause, diagnosis, treatment, and associated complications. Postherpetic neuralgia (PHN), the most common complication of HZ, is the primary focus of the discussion. PHN is defined broadly as chronic pain that persists after the characteristic vesicular rash of HZ has resolved. PMID- 10864693 TI - The blessing of giving: the importance of reciprocity to self-health care. AB - This qualitative study of reciprocity and self-health care demonstrated the importance of reciprocity in the lives of 20 elderly participants. Self-health care included actions centering around nutrition, exercise, health care monitoring, and psychospiritual activities. The importance of giving to others and the difficulty in accepting help should their functional capacity diminish create concern among our participants. Nursing interventions based on understanding reciprocity and helping caregivers create opportunities for the older person to reciprocate, despite limited physical resources, are discussed. PMID- 10864694 TI - Needs assessment of homebound elders in a parish church: implications for parish nursing. AB - The goals in managing chronic illnesses in the elderly are to maintain the highest possible level of functioning, promote independence in self-care, and enhance quality of life. Parish nursing is one mechanism for providing care for homebound elders with one or more chronic illnesses. However, determining the holistic health needs of older parishioners before developing a parish nursing program is helpful. The holistic needs of elders in two parishes were determined through two multidimensional functional assessment questionnaires and a quality of life scale. Results indicate that this population of elders had relatively high levels of functioning. Quality of life was relatively high and was associated with overall functional abilities. PMID- 10864695 TI - Parish nurse leaders. PMID- 10864696 TI - Nurse practitioner geropsychiatric consultation service to nursing homes. AB - A nurse practitioner (NP) psychiatric consultation service was established to provide the residents of five nursing homes with on-site assessment and follow-up treatment for behavioral and psychiatric problems under OBRA and Medicare guidelines. During the 1-year project, 175 residents were referred by the nursing home (NH) staff for agitation, disruptive behavior, depressive symptoms, or decline in activities of daily living. An outcome evaluation documented that the NP recommendations resulted in positive behavioral changes in 62% of residents. Primary physicians, NH staff, and administrators validated that close monitoring of psychotropic medications and staff education in behavioral management strategies provided an effective, collaborative service. The practical aspects of establishing this consult service are addressed. PMID- 10864697 TI - Cancer pain management in the elderly. AB - The prevalence of cancer rises with advancing age, and pain is a common complaint of those with cancer. The elderly, especially those older than 85, are at risk for undertreatment of pain. Despite multiple barriers, assessment difficulties, and an increased risk of side effects with opioids, cancer pain can be effectively managed in this population. The quality of life of elderly patients with cancer depends on good symptom management, the appropriate use of analgesics, and prevention of opioid-related side effects. PMID- 10864698 TI - Update on drugs for emotional pain in elders. PMID- 10864699 TI - Pain assessment in the home. Part 1. PMID- 10864700 TI - SPECT bone scintigraphy in the diagnosis and management of mandibular condylar hyperplasia. AB - Isotope bone scans have been used for a number of years to assess growth activity in the mandibular condyle in patients who present with facial asymmetry. The aim is to distinguish normal bone growth within the condyle from increased activity that may be the cause of the asymmetry. Previous studies have, however, relied only on planar images. SPECT (single photon emission computed tomography) has been used with quantitative assessments of one mandibular condyle to clivus or lumbar spine, but we have compared one condyle with the other, which is more sensitive and accurate in detecting abnormal activity. A relative percentage uptake of 55% or more in the affected mandibular condyle is considered to be abnormal, and this has been validated by comparison with an age-matched control group. We have used SPECT as an aid to diagnosis and treatment in 18 patients with asymmetrical growth and have constructed a therapeutic algorithm to aid the treatment of these patients. PMID- 10864701 TI - Midfacial tumours: a review of 72 cases. AB - We review 72 midfacial tumours managed during the 10-year period between 1985 and 1995. We describe presenting features, sites of lesions and histology, treatment regimens and outcomes, as well as the various surgical approaches for the resection of midfacial tumours, and their indications and contraindications. The choice of approach should be based on type of tumour, its site, and extent. PMID- 10864702 TI - Evolution of methods of uranostaphyloplasty exemplified by the analysis of 1118 primary operations for congenital palatal defects. AB - Of a total of 1118 uranostaphyloplasties in a maxillofacial hospital in Kiev, 819 have been operated on since 1980 by a new technique; 424 (52%) of these were in children under the age of 3 years. Analysis of the anatomical and the functional results of the operation, anthropometric studies, and data on logopaedic training showed that the proposed method of uranostaphyloplasty (cleft-palate repair) was effective both anatomically and functionally. PMID- 10864703 TI - Congenital transmandibular dermoid: case report. PMID- 10864704 TI - Precautions against cross-infection during operations for maxillofacial trauma. AB - One hundred oral and maxillofacial units in the UK were sent a postal questionnaire. Surgical staff of all grades were asked which infection-control measures were taken during the treatment of maxillofacial fractures. Two hundred and ninety-four questionnaires were completed, a response rate of 49%. If the patient was known to be infected by a blood-borne viral disease, significantly more surgeons used standard barrier precautions such as eye protection, fluid resistant gowns, drapes, ball-ended clips, adhesive tapes and intermediate trays (P<0.0001). Bone-plating techniques were used in preference to wire osteosynthesis (P<0.0001). Only 31 (10.5%) of surgeons routinely used double gloves but 250 (85%) did so if the patient was an infection risk (P<0.0001). Universal precautions were not applied equally to all patients. PMID- 10864705 TI - Sir Henry Trentham Butlin: the father of British head and neck surgery. AB - Sir Henry Butlin was our first pioneer in head and neck surgery. Though the man himself has been all but lost to history, surgeons in the UK and the USA are returning to the technique of selective neck dissection that he first described over a century ago. PMID- 10864706 TI - Actinomycotic ulcer of the oral mucosa: an unusual presentation of oral actinomycosis. AB - Although the oral mucosa is often the site of entry of actinomyces into the deeper tissues, actinomycosis in the oral mucosa is extremely rare. Actinomycotic lesions are usually described as either single or multiple abscesses or indurated masses with hard fibrous walls and soft central loculations. Actinomyces israelii is the principal cause of human actinomycosis. We present a rare case of actinomycosis caused by Actinomyces odontolyticus; it presented primarily as a long-standing ulcer of the oral mucosa mimicking a squamous cell carcinoma. PMID- 10864707 TI - Management of anticoagulation in patients with prosthetic heart valves undergoing oral and maxillofacial operations. AB - There is wide variation in the management of patients with mechanical prosthetic valves who are taking anticoagulants and who require non-cardiac surgery. In this paper, we outline a pragmatic, practical approach to the adjustment of anticoagulation in relation to both the degrees of surgical trauma during oral and maxillofacial surgery and the risk of thromboembolism associated with the prosthetic valve. For minor surgery, no adjustment of anticoagulation is undertaken if the International Normalized Ratio is less than 4.0, if local haemostatic methods and tranexamic acid mouthwashes are used. For major surgery, warfarin is stopped preoperatively and low-molecular-weight heparin is used. For emergency surgery, partial reversal of anticoagulation with low-dose parenteral vitamin K is obtained. PMID- 10864708 TI - Behavioural measurement of postoperative pain after oral surgery. AB - The amount and type of postoperative analgesia prescribed depends on the clinician's judgement of the patient's need. Among other factors, this judgement is likely to be based on the patient's behaviour. The primary aim of this study was to investigate the validity of using behavioural measures to provide information about a patient's experience of pain during the early stages of recovery from oral surgery under general anaesthesia. Behavioural measures were not valid measures of acute postoperative pain, which suggests that while clinicians may build a better picture of a patient's experience of pain by including behavioural observation in their range of assessments, they should not rely on them when judging a patient's need for analgesia. The results also show differences between the sexes in their reaction to pain. Significantly more women than men showed signs of pain, despite little difference in self-rating pain scores. PMID- 10864709 TI - Three years' experience of collaborative care pathways on a maxillofacial ward. AB - Collaborative care pathways (CCPs) provide a framework for multidisciplinary patient care. They provide guidelines and a mechanism for audit, and were first introduced at the Regional Unit, Walton Hospital, Liverpool, in November 1994. They have been designed for many surgical groups. Between August 1996 and 31 July 1997, 955 patients were admitted on to the nine established pathways: fractured mandible (n=213), fractured zygoma (n=117), minor oral surgery (n=244), abscess (n=18), examination under anaesthesia (n=73), nasal surgery (n=73), osteotomy (n=80), salivary (n=63), and temporomandibular joint (n=74). The purpose of this article is to report the introduction of CCP in a maxillofacial ward and give results from a one-year audit. CCP have proved to be an extremely useful tool and have several advantages over traditional documentation. They are more accurate, easily computerized, and facilitate audit. They promote the development of guidelines and standardized perioperative care, and this in turn facilitates training and raises standards of care. PMID- 10864710 TI - Vascularized transplantation of the long thoracic nerve for sensory reinnervation of the lower lip. AB - Microsurgical techniques have improved functional and morphological reconstruction of the face in recent years. An important factor is the re establishment of neuronal function. The aim of this study was a follow-up of the regeneration of sensation in the inferior alveolar nerve after partial resection of a tumour and reconstruction with a vascularized long thoracic nerve graft. Five patients were examined in monthly intervals to assess the degree of re establishment of sensation. Pressure and pain responses were elicited as early as three months postoperatively, sense of touch and vibration were found after five months, and sensitivity to temperature after seven months postoperatively. In four patients nine months postoperatively, sensory qualities in the region of the mental nerve were identical on both sides. The vascularized long thoracic nerve is therefore an adequate nerve graft for covering defects as a result of resection of the inferior alveolar nerve patients with tumours. PMID- 10864711 TI - New surgical technique for the correction of congenital muscular torticollis (wry neck). AB - Congenital muscular torticollis (wry neck) results from shortening of the sternocleidomastoid muscle and may lead to limitation of neck movement and craniofacial deformity. If conservative treatment is started early, with a regimen of passive stretching exercises and active strengthening of the contralateral muscle, about 95% of patients achieve an acceptable range of neck movement. The surgical management of patients who do not respond to physiotherapy remains controversial. Its aim is to provide a long-term, cosmetic restoration of neck mobility while minimizing the development of craniofacial deformity and upper cervical scoliosis; few previously advocated techniques achieve both these goals. We describe a technique that combines subperiosteal lengthening of the sternocleidomastoid muscle at its mastoid insertion, and division of lower fibrotic bands with minimal postoperative fibrosis. As the sternomastoid muscle is reattached lower down on the mastoid process, the lengthening of the muscle is stable, because the tendency to fibrosis and shortening is minimized. Comparison of the results with previous series shows that this technique provides immediate benefit and good long-term results. PMID- 10864712 TI - Effect of limited jaw motion on ankylosis of the temporomandibular joint in sheep. AB - The purpose of this study was to investigate the effect of limited movement of the jaw on ankylosis of the temporomandibular joint (TMJ). Eighteen adult sheep were divided into two groups. In Group 1, the temporal and condylar articular surfaces were removed together with the disc on the right. In Group 2, we did the same procedures but in addition the jaw movements were limited by a wire. One sheep was killed just after the operation, four at one month, and four at three months, in each group. The range of jaw movements preoperatively and at the time of death were recorded. The joints were examined radiologically, macroscopically, and histologically. We used a scoring system to assess the radiological changes and histological extent of ankylosis. At one month, the joint spaces were filled with fibrous tissue, but a small joint space existed in all four joints in Group 1. In Group 2, there was full ankylosis in two joints and partial ankylosis in two joints. At three months, similar ankylotic changes were seen in both groups. The histological score for ankylosis at one month showed that those in Group 2 were significantly more ankylosed than in Group 1 (P<0.01). The range of jaw movements was more limited at one month in Group 2, both vertically and to the left, and was significantly decreased in both groups (P<0.01) at three months. Limitation of jaw motion hastens the progress of TMJ ankylosis. PMID- 10864713 TI - Value of computed tomography and magnetic resonance imaging in the treatment of a calcifying epithelial odontogenic (Pindborg) tumour. AB - The calcifying epithelial odontogenic (Pindborg) tumour is a rare primary tumour of the jaw with a characteristic histological appearance. We describe here its appearance on computed tomography (an expanding, multilocular lesion that is thinning both plates of the mandible, is well-defined and contains scattered radio-opaque areas) and magnetic resonance imaging (predominantly hypointense on T1-weighted images and of mixed hyperintensity on T2-weighted images; the extent of the tumour including involvement of the mandibular canal was clear). PMID- 10864714 TI - Serology should be a routine investigation when presented with a major salivary gland lump. PMID- 10864715 TI - Association of central giant-cell granuloma of the maxilla with pyknodysostosis. PMID- 10864716 TI - Visualization of metastatic lymph nodes in oral cancer by angiograms. PMID- 10864717 TI - Latex allergy. PMID- 10864719 TI - Notices PMID- 10864718 TI - Reduced rate of adult respiratory distress syndrome. PMID- 10864720 TI - Effect of short-term high dose of steroids on the healing of microvascular anastomoses in a rabbit model: pilot study. AB - Short-term high doses of steroids have been used in head and neck surgery to reduce swelling in free tissue transfers. This study was set up to assess the effect of this treatment on the healing of microvascular anastomoses in a rabbit model. Anastomoses were made in the common carotid artery and anterior facial vein in 40 animals, half of which were given high doses of methylprednisolone (2.5 mg/kg/day) intravenously for 48 h peri-operatively. The patency of the vessels was assessed and they were harvested for histological examination at 1, 3, 7, 14, and 90 days postoperatively. Steroids tended to reduce the swelling in the vessel walls during the early stages of healing (24 h and 3 days), and also to reduce the inflammatory cell count in the lumen of the vessels at 3 days. It seems likely from these initial observations that steroids are more likely to improve than jeopardize the healing of microvascular anastomoses. PMID- 10864721 TI - Review of severe osteoradionecrosis treated by surgery alone or surgery with postoperative hyperbaric oxygenation. AB - We reviewed 41 patients with osteoradionecrosis of the mandible. Each patient was treated by radical resection followed by external beam irradiation. The diagnosis of infected osteoradionecrosis was confirmed clinically, radiologically, and histologically. After operation had failed, 20/41 were given hyperbaric oxygen (HBO) as in 'salvage' treatment. Daily sessions of HBO 2.5 ATA for 60 minutes (mean: 29 sessions) were given. The other 21 patients were treated by operation and antibiotics alone. HBO group (n = 20): The overall success rate for HBO after operation had failed was 13/20. Repeated debridement as first-line treatment followed by postoperative HBO was successful in 12/19. In seven of 19 patients, partial mandibulectomy and microvascular transplantation were required as second line treatment, and this was successful in five. Primary partial mandibulectomy and microvascular transplantation followed by HBO was successful in 1 patient. Non-HBO group (n = 21): Repeated debridement was successful in 10/11 patients. Partial mandibulectomy was required as second-line treatment in the remaining one. In the other 10, partial mandibulectomy and microvascular transplantation were successful as first-line treatment in four. In the remaining six, further surgical intervention became necessary and were successful for 5-17 months (mean: 13). With a success rate of 13/20, we do not recommend HBO for the treatment of osteoradionecrosis. PMID- 10864722 TI - Endoscopic treatment of sinonasal disease in patients who have had orthognathic surgery. AB - Certain skeletofacial patterns may be predisposed to aggravated sinonasal disease postoperatively. These may include, but are not limited to, facial skeletal asymmetries with high septal deviations and those with obstructive nasal respiration and mouth breathing that leads to skeletal growth disturbances such as vertical maxillary hyperplasia and apertognathism. These sinonasal diseases may partly be the result of osteomeatal blockage by pre-existing structures, or synechial shelves and webs blocking normal maxillary antral mucosal flow. The use of nasal antral windows placed anteriorly in the lateral nasal wall at the time of downfracture LeFort (Hosaka window) do not seem to benefit the drainage of the maxillary antrum. This is because physiological flow often bypasses this region. If patients present postoperatively with new sinonasal disease or the aggravation of pre-existing symptoms, evaluation by both endoscopically assisted intranasal and axially and coronal computed tomography (CT) is recommended. Functional endoscopic sinus surgery by the minimally invasive Messerklinger technique, combined with intranasal use of laser-assisted turbinoplasty and soft tissue lysis, have been successfully used for most of these patients. Because the anatomical positioning of the midfacial structure can potentially affect patients with a predisposition to sinonasal physiological disturbances, consideration should be given to preoperative evaluation and discussion of potential consequences. PMID- 10864723 TI - Comparison of three facebow/semi-adjustable articulator systems for planning orthognathic surgery. AB - Our aim was to measure the steepness of the occlusal plane produced by three different semi-adjustable articulators: the Dentatus Type ARL, Denar MkII, and the Whipmix Quickmount 8800, and to assess the influence of possible systematic errors in positioning of study casts on articulators that are used to plan orthognathic surgery. Twenty patients (10 skeletal class II, and 10 skeletal class III) who were having pre-surgical orthodontics at Liverpool University Dental Hospital were studied. The measurement of the steepness of the occlusal plane was taken as the angle between the facebow bite-fork and the horizontal arm of the articulator. This was compared with the angle of the maxillary occlusal plane to the Frankfort plane as measured on lateral cephalometry (the gold standard). The Whipmix was closest to the gold standard as it flattened the occlusal plane by only 2 degrees (P<0.05). The results of the Denar and Dentatus differed significantly from those of the cephalogram as they flattened the occlusal plane by 5 degrees and 6. 5 degrees (P<0.01), respectively. Clinicians are encouraged to verify the steepness of the occlusal plane on mounted study casts before the technician makes the model. PMID- 10864724 TI - Miniantrostomy for the reduction of fractures of the orbital floor. PMID- 10864726 TI - Comparison of two systems for rigidly connecting 2.0-mm bone screws to an implantable device: in vitro stability testing. AB - The stability of a screw-fixed implantable device can be improved by eliminating the freedom of movement between the screws and the device. Two systems have been developed for rigidly connecting 2. 0-mm bone screws to an implantable device, and the aim of this study was to test and compare the stability of the two systems. In system A, a fixing disc locked the screw-head. In system B, the thread of the screw locked directly into the device. The stability of the connection was tested by measuring the resistance against a torque on the screw, against shear loading, against push-out loading, and against dynamic shear loading (70 N) for 5 million cycles. Both systems met the required minimum values for the resistance against a torque, shear loading and push-out loading, and dynamic shear loading did not cause movements between the screw and the device. We conclude that both systems are stable, but prefer system B because it is easier to make and implant than system A. PMID- 10864725 TI - Squamous cell carcinoma of the oral tongue: an analysis of prognostic factors. AB - AIM: To identify the prognostic significance of different factors in patients with squamous cell carcinoma of the tongue. PATIENTS AND METHODS: Seventy-seven patients with carcinoma of the tongue were treated radically at the King Faisal Specialist Hospital and Research Centre between 1980 and 1989. Twenty patients (26%) were treated by resection alone, 11 (14%) with radiotherapy alone, and 46 (60%) with combined resection and radiotherapy. RESULTS: Forty-seven patients (61%) had T(1-2), 28 (36%) T(3-4), and two T(x) tumours. The regional nodes were clear in 53 (69%) and contained metastases in 24 patients (31%). Thirty patients (39%) developed recurrences, which were local in 9, regional in 14, locoregional in 5, and locoregional with metastatic disease in 2. The five and 10-year overall actuarial survival for all patients were 65% and 53%, respectively, and the corresponding relapse-free survival 56% and 50%. Univariate and multivariate analyses were done of seven variables - age (<40 compared with >/=40 years), sex, chewing tobacco use, smoking, TNM stage, surgical margins (clear or invaded), and treatment (resection, radiotherapy, or the combination). On univariate analysis chewing tobacco (P=0.04), smoking (P=0.01), invaded resection margins (P=0.04), involved regional lymph nodes (P=0.009), T4 tumours, and patients treated with radiotherapy alone (P=0.001) were associated with poor overall survival. Factors associated with shorter relapse-free survival were age >40 (P=0.03), chewing tobacco (P=0.04), invaded resection margins (P=0.01), and smoking (P=0.01). On multivariate analysis, invaded resection margins and smoking (P=0.04)(P=0.02) were associated with shorter overall survival and relapse-free survival (P=0.03 and (P=0.01), while chewing tobacco independently influenced relapse-free survival only (P=0.03). CONCLUSION: Invaded resection margins and smoking were the only independent prognostic factors that affected both overall and relapse free survival. Those who chewed tobacco were at high risk of locoregional failure. PMID- 10864727 TI - Synovial chondromatosis of the temporomandibular joint. AB - We report two cases of temporomandibular joint (TMJ) synovial chondromatosis, one of which was in the early stage and treated arthroscopically. The second was more advanced and recurred after removal of the free bodies, and so required condylectomy and synovectomy. PMID- 10864728 TI - Head and neck cancer and its treatment: historical review. AB - Head and neck cancer has been known to physicians since antiquity, but until relatively recently any material advance was limited by the lack of anaesthesia. The factors and people that helped to develop the subject of head and neck surgery have been traced through history, and this paper provides a broad historical perspective with which to compare the current standard of management for head and neck cancer. PMID- 10864729 TI - Multiple odontogenic keratocysts in mental retardation-overgrowth (Simpson-Golabi Behmel) syndrome. AB - We report on a 10-year-old boy with mental retardation-overgrowth (Simpson-Golabi Behmel) syndrome. The child had the typical clinical features including, postnatal overgrowth, mental retardation, and a characteristic facial appearance. He was admitted for treatment of multiple mandibular and maxillary cysts. Histopathological examination of the cyst tissue showed keratinized epithelium. Odontogenic keratocysts may have to be added to the typical features of this syndrome. PMID- 10864731 TI - Mucus retention cyst of the maxillary sinus: the endoscopic approach. AB - OBJECTIVE: To present our experience of endoscopic surgery for symptomatic mucus retention cyst of the maxillary sinus. DESIGN: Retrospective study. SETTING: Teaching hospital, Israel. PATIENTS: 60 patients with 65 symptomatic cysts of the maxillary sinus who were operated on endoscopically. Only patients with large cysts that filled at least 50% of the sinus space were included. INTERVENTION: A rigid nasal endoscope was used in all cases; most of the cysts were removed through the natural sinus ostium. RESULTS: Cysts recurred in only two patients during the first postoperative year. There were no complications from the procedure. CONCLUSION: The endoscopic approach to the treatment of maxillary sinus cyst is associated with a low rate of recurrence (3% in this study) and no complications, and we recommend it as the surgical procedure of choice. PMID- 10864730 TI - Treatment of the nostrils in patients with cleft lip by a nostril retainer. PMID- 10864732 TI - Increase in volume of lignocaine/adrenaline-containing local anaesthetic solution causes increase in acute postoperative pain after gingivectomy. AB - A randomized, single-blind, within-patient, crossover study was done in 44 patients (27 women and 17 men mean age 47 years, range 29-63) who had bilateral 'identical' gingivectomies. On one occasion a standard volume of local anaesthetic containing lignocaine 2% and adrenaline (1/80 000) was infiltrated into the mucosal tissue before operation. On the other occasion double the standard volume was infiltrated. The intensity of pain postoperatively was recorded by the patients on 100 mm visual analogue scale every hour for an 11 hour observation period. The intensity of pain when double volume had been given was significantly higher than that after the standard volume from 2 to 8 hours postoperatively (P < 0.04), the median (range) being 52.0 mm (0.0-434.0) compared with 30.5 mm (0.0-359.0) after the standard volume (P < 0.005). Doubling the volume of local anaesthetic containing adrenaline that was infiltrated increased the intensity of acute pain after gingivectomy. PMID- 10864733 TI - Spontaneous palatal fenestration: review of the literature and report of a case. AB - A 42-year-old, edentulous man presented with a defect in his hard palate. He gave a history of a painless lump one year previously which had discharged after a week. Investigations showed only long-standing hypoplasia of the left palatine process, with no evidence of any destructive process. We assumed that the fistula had developed as a result of breakdown of the mucosa covering an isolated cleft of the hard palate. We offered him repair, but he preferred to rely on his maxillary complete denture to cover the defect, and this has worked. PMID- 10864734 TI - Prevention of haematomas after auricular injuries. PMID- 10864735 TI - An unusual foreign body in the tongue. AB - A T-shaped metal object from an umbrella embedded in the tongue of a 4-year-old male child is described, along with its subsequent removal. The types of foreign bodies reported in this area are also mentioned. PMID- 10864736 TI - Re: van der Lei et al. Closure of radial forearm flap donor site with local full thickness skin graft. PMID- 10864738 TI - Synthesis and properties of oligodeoxynucleotides incorporating a conformationally rigid uridine unit having a cyclic structure at the 5'-terminal site. AB - A 2'-O-methyluridylic acid derivative 3 having a cyclic structure linked between the 5-position of the uracil residue and the 5'-phosphate group was synthesized. The NMR analysis suggests that this cyclouridylic acid derivative has exclusively the C3'-endo conformation that is in favor of duplex formation with RNA. Two oligonucleotides ?pc3Um(pT)(9) and pc3Um(pU)(9) incorporating this cyclouridylic acid unit at the 5'-terminal site were synthesized by using the fully protected cyclouridylic acid 3'-phosphoramidite derivative 11 in the solid-phase synthesis. To examine the actual effect of this cyclic structure on the thermal stability of duplexes between the modified oligonucleotides and their complementary oligonucleotides, two oligonucleotides ?pUm(pT)(9) and pUm(pU)(9) having an acyclic structure were also synthesized. As the complementary oligonucleotides, dA(pdA)(9) and A(pA)(9) were used for T(m) experiments with these 5'-terminal modified oligonucleotides. The T(m) values of all the possible duplexes were measured. These results clearly show that the duplex of pc3Um(pT)(9)-A(pA)(9) has a higher T(m) value by 5.5 degrees C than that of A(pA)(9)-T(pT)(9). This is rather significant compared with all other cases. Moreover, the T(m) value of pc3Um(pT)(9)-A(pA)(9) is 4.5 degrees C higher than that of pUm(pT)(9)-A(pA)(9). This result suggests that the cyclic structure can considerably contribute to stabilization of the duplex only in the case of the modified oligomer (DNA) and decaadenylate (RNA). PMID- 10864739 TI - General base and general acid catalyzed intramolecular aminolysis of esters. Cyclization of esters of 2-aminomethylbenzoic acid to phthalimidine. AB - Plots of log k(0) vs pH for the cyclization of trifluoroethyl and phenyl 2 aminomethylbenzoate to phthalimidine at 30 degrees C in H(2)O are linear with slopes of 1.0 at pH >3. The values of the second-order rate constants k(OH) for apparent OH(-) catalysis in the cyclization reactions are 1.7 x 10(5) and 5.7 x 10(7) M(-)(1) s(-)(1), respectively. These rate constants are 10(5)- and 10(7) fold greater than for alkaline hydrolysis of trifluoroethyl and phenyl benzoate. The k(OH) for cyclization of the methyl ester is 7.2 x 10(3) M(-)(1) s(-)(1). Bimolecular general base catalysis occurs in the intramolecular nucleophilic reactions of the neutral species. The value of the Bronsted coefficient beta for the trifluoroethyl ester is 0.7. The rate-limiting step in the general base catalyzed reaction involves proton transfer in concert with leaving group departure. The mechanism involving rate-determining proton transfer exemplified by the methyl ester in this series (beta = 1.0) can then be considered a limiting case of the concerted mechanism. General acid catalysis of the neutral species reaction or a kinetic equivalent also occurs when the leaving group is good (pK(a) 16 kcal mol(-)(1)) extrude nitrous oxide via a highly asynchronous transition state with a planar ring nitrogen atom. Nitrosoaziridine 1, with a low rotation barrier (<9 kcal mol(-)(1)) represents a special case. This compound can eliminate N(2)O via a low energy linear synperiplanar transition state (DeltaH()(298) = 20.6 kcal mol(-)(1), DeltaS()(298) = 2.5 cal mol(-)(1) K(-)(1)). Two higher energy transition states are also available. The B3LYP activation barriers of the cheletropic fragmentation of nitrosoheterocycles 2-4 decrease in the series: 2 (58 kcal mol( )(1)) >> 3 (18 kcal mol(-)(1)) > 4 (12) kcal mol(-)(1). The relative strain energies increase in the same order: 2 (0 kcal mol(-)(1)) << 3 (39 kcal mol( )(1)) < 4 (52 kcal mol(-)(1)). Comparison of the relative energies of 2-4 and their transition states on a common scale where the energy of nitrosopyrroline 2 is assumed as reference indicates that the thermal stability of the cyclic nitrosoamines toward cheletropic decomposition is almost entirely determined by the ring strain. PMID- 10864744 TI - Ring-expansion of thioacetal ring via bicyclosulfonium ylide. Effect Of protic nucleophile on ylide intermediate AB - The reaction of 2-(3-diazo-2-oxopropane-1-yl)-2-methyldithiolane 9a, 2-(4-diazo-3 oxobutane-2-yl)-2-methyldithiolane 9c, and 2-(3-diazo-2-oxopropane-1-yl)-2-methyl 1,3-dithiane 9b with Rh(2)(OAc)(4) gave three-carbon ring-expansion products dithiocan-2-en-1-ones 13a, 13c and dithionan-2-en-1-one 13b, respectively. 2-(5 Diazo-4-oxopentyl)-2-methyldithiolane 9e also gave the five-carbon ring-expansion products dithionan-3-en-1-one 13e and 5-methylenedithionane-1-one 13'e. On the other hand, reaction of 2-(4-diazo-3-oxobutyl)-2-methyldithiolane 9d in the presence of AcOH gave the four-carbon ring-expansion product 16d substituted by an acetoxyl group. In addition, the reaction of 2-(3-diazo-2-oxopropyl)-2-methyl 3-oxathiolane 9f in the absence of AcOH gave 4-oxa-7-thiocan-2-en-1-one 19 via a sulfonium ylide intermediate, whereas, in the presence of AcOH, an alternative regioisomer 20 was also formed competitively with 19 via an oxonium ylide intermediate. PMID- 10864745 TI - Photochemistry of 3- and 5-phenylisothiazoles. Competing phototransposition pathways. AB - 5-Phenylisothiazole undergoes phototransposition via the electrocyclic ring closure-heteroatom migration pathway and by the N(2)-C(3) interchange reaction pathway. The latter route is enhanced by the addition of triethylamine (TEA) to the reaction medium and by increasing the polarity of the solvent. In addition to phototransposition, 5-phenylisothiazole also undergoes photocleavage to 2-cyano-1 phenylethenethiol which was trapped by reaction with benzyl bromide to yield 2 cyano-1-phenylethen-1-ylbenzyl thioether. 3-Phenylisothiazole also phototransposes by both reaction pathways, but the product distribution is not affected by the addition of TEA or by changing the solvent polarity. PMID- 10864746 TI - Highly efficient induction of chirality in intramolecular AB - Highly stereocontrolled, intramolecular [2 + 2] cycloadditions between ketenimines and imines leading to 1,2-dihydroazeto[2, 1-b]quinazolines have been achieved. The source of stereocontrol is a chiral carbon atom adjacent either to the iminic carbon or nitrogen atom. In the first case, the stereocontrol stems from the preference for the axial conformer in the first transition structure. In the second case, the origin of the stereocontrol lies on the two-electron stabilizing interaction between the C-C bond being formed and the sigma orbital corresponding to the polar C-X bond, X being an electronegative atom. These models can be extended to other related systems for predicting the stereochemical outcome in this intramolecular reaction. PMID- 10864747 TI - Synthesis and fluorescence properties of new fluorescent, polymerizable, metal chelating lipids. AB - Liposomes incorporating fluorescent, metal-chelating lipids find applications in molecular recognition of peptides, 2D protein recrystallization, protein targeting, and biological sensing. It would be advantageous to combine the usefulness of polymerizable, metal-chelating lipids and fluorescent lipids. Herein, we report the synthesis and fluorescence properties of several fluorescent, polymerizable, metal-chelating lipids. They have been successfully incorporated into liposomes and then polymerized. These lipids can be used as membrane probes to study the polymerizable liposomes in the unpolymerized state and to investigate lipid redistribution during polymerization. In addition, if a luminescent metal ion (e.g., Eu(3+), Tb(3+), etc.) is used to complex the headgroup, the lipids can probe the membrane interior and exterior simultaneously. PMID- 10864748 TI - Reactions of C(6)F(5)Li with tetracyclone and 3-ferrocenyl-2,4, 5 triphenylcyclopentadienone: An (19)F NMR and X-ray crystallographic study of hindered pentafluorophenyl rotations AB - Tetracyclone, 2a, reacts with C(6)F(5)Li to yield 2-pentafluorophenyl-2,3,4,5 tetraphenylcyclopent-3-en-1-one, 7, and 5-hydroxy-5-pentafluorophenyl-1,2,3,4 tetraphenylcyclopentadiene, 8, as the result of 1,6 and 1,2 additions, respectively. In contrast, treatment of 3-ferrocenyl-2,4,5 triphenylcyclopentadienone, 2b, with lithiopentafluorobenzene leads to 4 ferrocenyl-4-pentafluorophenyl-2, 3,5-triphenylcyclopent-2-en-1-one, 9, and 5 hydroxy-5-pentafluorophenyl-2-ferrocenyl-1,3, 4-triphenylcyclopentadiene, 10, the products of 1,4 and 1,2 addition, respectively. The structures of 7-9 have been established by X-ray crystallography, and the barriers to rotation (19-21 kcal mol(-)(1)) of the pentafluorophenyl groups in 8-10 have been studied by variable temperature (19)F NMR. Nucleophilic attack at the ferrocenyl-bearing carbon in 2b is rationalized in terms of a zwitterionic structure in which the positive charge of the "cyclopentadienyl cation" is delocalized onto the iron atom in the organometallic substituent. PMID- 10864749 TI - Computer simulations of enantioselective ester hydrolyses catalyzed by Pseudomonas cepacia lipase. AB - On the basis of the X-ray crystal structure of the lipase from Pseudomonas cepacia (PcL)-an enzyme representative for a whole family of Pseudomonas lipases (lipase PS, SAM-2, AK 10, and others with a high degree of homology with PcL)-a computational study was performed to rationalize both the enantioselectivity and substrate specificity (tolerance) displayed by this lipase in the enantioselective hydrolysis of racemic esters 1a-12a from various secondary aromatic alcohols. The major goal of this project was the development of a binding model for PcL which is able to rationalize the experimental findings to predict "a priori the enantioselective behavior of PcL toward a wider range of substrates. A two-step modeling procedure, namely, docking experiments followed by construction of tetrahedral intermediates, was used for the simulation of the involved enzyme-substrate recognition/hydrolysis processes. The study of the recognition process (docking experiments) led to unambiguous identification of the binding geometry for the two enantiomeric series of substrates, but did not suggest a definitive interpretation of the behavior of PcL. Taking into consideration the stereoelectronic requirements of the enzymatic hydrolysis reaction, both the enantioselectivity and tolerance of the enzyme were then explained through the study of the tetrahedral intermediates, in turn constructed from the calculated docking geometries of 1a-12a. PMID- 10864750 TI - Sakurai addition and ring annulation of allylsilanes with alpha, beta-unsaturated esters. Experimental results and ab initio theoretical predictions examining allylsilane reactivity AB - Trimethylallylsilane has been shown to add to methyl acrylate in good yield when catalyzed by TiCl(4) at room-temperature despite literature reporting to the contrary. Further, even with these small alkyl ligands on the metal, ring annulation occurs to a large extent, in addition to simple allylation (Sakurai addition). The kinetic product is the ((trimethylsilyl)methyl)cyclobutane derivative which can be isomerized to cyclopentanoid, the thermodynamic product, if left in the presence of the catalyst. Consistent with other literature in this area, increasing the size of the ligands on silicon increases both the rate of product formation and the proportion of ring annulation relative to allylation. To develop a predictive model for allylsilane reactivity, ab initio gas-phase calculations have been made on the parent allylsilane with different ligands on the metal and on the reaction between these allylsilanes and acrolein, acrylic acid, and methyl acrylate. Predictions indicate that as the length of n-alkyl ligands on silicon increase, so does the apparent ability of the Si-Calpha bond of the allylsilane to hyperconjugate with developing vacant p orbital on Cbeta as the allylsilane begins to attack an electrophile. This is corroborated by a gradually increasing HOMO in the ground-state allylsilane as the ligands are changed from methyl through to n-hexyl and an increasing Si-Calpha bond length and decreasing Si-Calpha-Cbeta bond angle in the protonated species. These results in the gas phase mirror the reactivity of these n-alkyl-substituted allylsilanes in experiment; i.e., as the length of the alkyl chain increases, reactivity increases significantly. Triisopropylallylsilane, a very reactive silane, appears to anomalous in charge distribution and geometrical features compared with other substituted allylsilane systems which is due, presumably, to steric effects. The calculations on the protonated species would indicate that almost no hyperconjugative stabilization can occur on the basis of the bond lengths and angles necessary to promote good orbital overlap between the Si Calpha bond and the empty p orbital on Cbeta. However, the gas-phase reaction of the triisopropylallylsilane with acrolein and methyl acrylate led to comparatively low energy barriers of 13.1 and 24.5, respectively, which is consistent with its high experimental reactivity. Together, this computational analysis has produced a useful model for predicting allylsilane reactivity and some possible explanations for this reactivity. PMID- 10864751 TI - Synthesis of beta-dicarbonyl compounds using 1-acylbenzotriazoles as regioselective C-acylating reagents AB - 1-Acylbenzotriazoles 1 are efficient C-acylation reagents for the regioselective conversion of ketone enolates 2 into beta-diketones 3. PMID- 10864752 TI - Novel syntheses of hexahydro-1H-pyrrolo[1,2-a]imidazoles and Octahydroimidazo[1,2 a]pyridines. AB - 1-Phenyl-5-(benzotriazol-1-yl)hexahydro-1H-pyrrolo[1,2-a]imidazole (18) and 1 phenyl-5-benzotriazolyloctahydroimidazo[1,2-a]pyridine (27) were readily prepared from succindialdehyde or glutaraldehyde, benzotriazole, and N phenylethylenediamine. Synthons 18 and 27 reacted with Grignard reagents, allylsilanes, silyl ethers, and triethyl phosphite to produce 1-phenyl-5 substituted-hexahydro-1H-pyrrolo[1,2-a]imidazoles 20a-f, 22, 24a,b, and 25 and 1 phenyl-5-substituted-octahydroimidazo[1, 2-a]pyridines 28a-e, 32, 33a,b, and 34 in good to excellent yields. The configurations of 20, 22, 24, and 25 were determined to be cis isomers by NOE experiment, while the configurations and conformations of 28a-e, 32, 33a,b, and 34 were elucidated by (1)H-(1)H COSY and (1)H-(13)C COSY. PMID- 10864753 TI - A facile and convenient synthesis of 3-alkylamino-5-arylthiophenes with a variety of substituents at C-2 and studies of reaction mechanisms AB - Thioaroylketene S,N-acetals were treated with active methylene compounds including beta-keto ester, nitromethane, cyanoacetic acid, p toluenesulfonylacetone, 4-nitrophenylacetic acid, and diethyl (2 oxopropyl)phosphonate in the presence of mercury(II) acetate in CH(2)Cl(2) at room temperature. These reactions gave 3-alkylamino-5-arylthiophenes containing various substituents, which comprised, respectively, alkoxycarbonyl, nitro, cyano, p-toluenesulfonyl, 4-nitrophenyl, and diethylphosphono groups at C-2 in good yields. The reaction of 3-methylamino-3-methylthio-1-phenylthioxopropene with malonic acid or Meldrum's acid under the same conditions gave 3-methylamino 5-phenylthiophene. Similarly, treatment of 3-methylamino-3-methylthio-1 phenylthioxopropene with various enolizable cyclic ketones such as 4-hydroxy-6 methyl-2-pyrone, homophthalic anhydride, 2-hydroxy-1,4-benzoquinone, and 1, 3 diethyl-2-thiobarbituric acid gave thieno[3,2-b]pyridin-4-one, thieno[3,2 c]isoquinolin-5-one, thieno[3,2-c]benzazepine-1,6-dione, and thieno[3,2 d]pyrimidine-2,4-dione, respectively. PMID- 10864754 TI - NH tautomerism in the natural chlorin derivatives. AB - The NH tautomerism of five Mg-free chlorophyll a and b derivatives 2-6 was studied utilizing NMR spectroscopy and molecular modeling. The results from the dynamic NMR measurements of the chlorins revealed that substituent effects contribute crucially to the free energy of activation (DeltaG(double dagger)) in the NH tautomeric processes. An intermediate tautomer for the total tautomeric NH exchange in a chlorin was observed for the first time, when the (1)H NMR spectra of chlorin e(6) TME (3) and rhodin g(7) TME (4) (TME = trimethyl ester) were measured at lower temperatures. The lower energy barriers (DeltaG(1)(double dagger)) obtained for the formation of the intermediate tautomers of 3 and 4, assigned to the N(22)-H, N(24)-H trans-tautomer, were 10.8 and 10.6 kcal/mol, respectively. The energy barrier (DeltaG(2)(double dagger) value) for the total tautomeric NH exchange in the five chlorins was found to vary from 13.6 kcal/mol to values higher than 18 kcal/mol. The lowest DeltaG(2)(double dagger) value (13.6 kcal/mol) was obtained for rhodochlorin XV dimethyl ester (2), which was the only chlorophyll derivative lacking the C(15) substituent. In the case of chlorins 4 and 5, the steric crowding around the methoxycarbonylmethyl group at C(15) raised the DeltaG(2)(double dagger) activation free-energy to 17.1 kcal/mol. However, the highest energy barrier with DeltaG(2)(double dagger) > 18 kcal/mol was observed for the NH exchange of pyropheophorbide a methyl ester (6), possessing the macrocycle rigidifying isocyclic ring E. Our results demonstrate that the steric strain, arising either from the steric crowding around the bulky substituent at C(15) or the macrocycle rigidifying isocyclic ring E, slows down the NH tautomeric process. We suggest that deformations in the chlorin skeleton are closely connected to the NH tautomeric exchange and that the exchange occurs by a stepwise proton-transfer mechanism via a hydrogen bridge. PMID- 10864755 TI - Synthesis of macrocyclic cage compounds by diamine-dihalide one-step coupling reaction AB - Macropolycyclic cage compounds were synthesized by a direct reaction between diamines and bis(bromomethyl) compounds. The procedure for constructing the polycyclic cage structure is simple and straightforward. The macropolycyclic compounds obtainable from this cyclization procedure are three-dimensional cage compounds, and any other isomers were not obtained except for two examples. Benzene, pyridine, and aliphatic units could be introduced into the cage structure. The macrocycles that have strong cation affinity were obtained as their potassium complexes. PMID- 10864757 TI - Rhodium(II)-catalyzed carbocyclization reaction of alpha-diazo carbonyls with tethered unsaturation AB - o-Alkynyl-substituted alpha-diazoketones undergo internal cyclization to produce indenone derivatives upon treatment with catalytic quantities of Rh(II) carboxylates. A variety of structural influences were encountered by varying the nature of the substituent group attached to the diazo center. The cyclization reaction involves addition of a rhodium-stabilized carbenoid onto the acetylenic pi-bond to generate a cycloalkenone carbenoid. The cyclized carbenoid was found to undergo both aromatic and aliphatic C-H insertion as well as cyclopropanation across a tethered pi-bond. Subjection of diazo phenyl acetic acid 3-phenylprop-2 ynyl ester to Rh(II) catalysis furnished 8-phenyl-1, 8-dihydro-2 oxacyclopenta[a]indenone in high yield. The formation of this compound involves cyclization of the initially formed carbenoid onto the alkyne to produce a butenolide which then undergoes C-H insertion into the neighboring aromatic system. When a vinyl ether is added, the initially formed rhodium carbenoid intermediate can be intercepted by the electron-rich pi-bond prior to cyclization. Different rhodium catalysts were shown to result in significant variation in the product ratios. The competition between bimolecular cyclopropanation, 1,2-hydrogen migration, and internal cyclization was probed using several enol ethers as well as diazoesters which possess different substituent groups on the ester backbone. The specific path followed was found to depend on electronic, steric, and conformational factors. PMID- 10864756 TI - Tandem ireland-claisen rearrangement ring-closing alkene metathesis in the construction of bicyclic beta-lactam carboxylic esters. AB - 4-Alkenyl-2-azetidinone systems were converted to the corresponding ethyl 2-?4 alkenyl-2-oxo-1-azetidinyl-4-pentenoates. In addition, 4-(2-propenyl-1-oxy)-, 4 (2-propenyl-1-thio)-, 4-?N-(2-propenyl)-(4-toluenesulfonyl)- and (3S, 4R)-4-(2 propenyl)-3-?(1R)-1-(tert-butyldimethylsilyloxy)ethyl-++ +azeti din-2-one were converted into beta-lactam dienes via sequential N-alkylation, Ireland-Claisen ester enolate rearrangement and esterification. Ring-closing metathesis using the Schrock ?(CF(3))(2)MeCO(2)Mo(=CHCMe(2)Ph)(=NC(6)H(3)-2,6-iso-Pr(2)) (1) or Grubbs Cl(2)(Cy(3)P)(2)Ru=CHPh (2) carbenes gave a series of ?5.2.0 and ?6.2.0 bicycles. Subsequent elaboration of the analogous (2R,7R, 8S)-tert-butyl 8-?(1R)-(tert butyldimethylsilyloxy)ethyl-1-aza-9-oxobicyclo++ +?5.2. 0non-4-ene-2-carboxylate (15), via selenation and desilylation, gave (+)-(2S,7R,8S)-tert-butyl 8-?(1R) hydroxyethyl-1-aza-9-oxobicyclo?5.2.0nona-2, 4-diene-2-carboxylate (18), a novel type of bicyclic beta-lactam. Diels-Alder cycloaddition further afforded tetracyclic systems exemplified by tert-butyl (1R,4S,5R,7S)-4-?(1R)-1 hydroxyethyl-3,9, 11-trioxo-10-phenyl-2,8,10,12-tetraazatetracyclo?5.5.2.0.(2, 5)0(8, 12)tetradec-13-ene-1-carboxylate (19). PMID- 10864758 TI - Chlorophosphonates: inexpensive precursors for stereodefined chloro-substituted olefins and unsymmetrical disubstituted acetylenes AB - New chlorophosphonates bearing a 1,3,2-dioxaphosphorinane ring which are useful for the stereospecific synthesis of 5-chlorofurfuryl substituted olefins and chloro-substituted dienes have been obtained by an easy, inexpensive route. The utility of some of these in the synthesis of ferrocenyl- and anthracenyl substituted unsymmetrical acetylenes has been explored. The structures of the phosphonates (OCH(2)CMe(2)CH(2)O)P(O)CH(2)(C(4)H(2)ClO) (4) and (OCH(2)CMe(2)CH(2)O)P(O)(CH=CHCH(Cl)Ph (7) have been determined; in addition, the stereochemistry of (5-chlorofurfuryl)CH=CH(4-ClC(6)H(4)) (13b) and 2, 4 Cl(2)C(6)H(3)-CH=CH-CH=C(Ph)Cl (14a) is unambiguously proved by the X-ray structure determination. PMID- 10864759 TI - Total synthesis of the ansamycin antibiotic (+)-thiazinotrienomycin E. AB - The first total synthesis of (+)-thiazinotrienomycin E (1), member of a novel class of cytotoxic ansamycin antibiotics, has been achieved. Key features of the synthetic strategy include (a) the efficient construction of sulfone 7 incorporating TBS protection of the aniline, (b) an improved synthesis of allyl chloride (-)-6, the advanced intermediate employed in our trienomycins A and F total syntheses, (c) application of the Kocienski modified Julia protocol to elaborate the E,E,E-triene subunit in a stereo-controlled fashion, (d) an efficient union of sulfone 7 with advanced iodide 62, and (e) Mukaiyama macrolactamization to access the thiazinotrienomycin macrocyclic ring. PMID- 10864760 TI - Asymmetric aldol reactions using (S,S)-(+)-pseudoephedrine-based amides: stereoselective synthesis of alpha-methyl-beta-hydroxy acids, esters, ketones, and 1,3-Syn and 1,3-anti diols AB - A very efficient method for performing stereoselective aldol reactions is reported. The reaction of (S, S)-(+)-pseudoephedrine-derived propionamide enolates with several aldehydes yielded exclusively one of the four possible diastereomers in good yields, although transmetalation of the firstly generated lithium enolate with a zirconium(II) salt, prior to the addition of the aldehyde, is necessary in order to achieve high syn selectivity. The so-formed syn-alpha methyl-beta-hydroxy amides were transformed into other valuable chiral nonracemic synthons such as alpha-methyl-beta-hydroxyacids, esters, and ketones. Finally, a stereocontrolled reduction procedure starting from the so-obtained alpha-methyl beta-hydroxy ketones has been developed allowing the synthesis of either 1,3-syn- or 1,3-anti-alpha-methyl-1,3-diols in almost enantiopure form by choosing the appropriate reaction conditions. PMID- 10864761 TI - Facile synthesis of oxa- and azacyclic dienes via cycloalkenation of alkynyltungsten compounds. Stereoselective construction of tricyclic furan and pyran derivatives via intramolecular diels-alder reaction AB - A convenient and short synthesis of functionalized oxacyclic and azacyclic dienes is developed on the basis of organotungsten chemistry. Alkynyltungsten compounds bearing a tethered alcohol and amine are treated with aldehydes and BF(3).Et(2)O in cold diethyl ether to give tungsten-heterocyclic carbenium salts, further leading to tungsten-heterocyclic dienes via deprotonation with Et(3)N. Hydrodemetalation of these tungsten-heterocyclic dienes is performed by the action of anhydrous Me(3)NO in CH(3)CN. This method is applicable to the synthesis of a number of oxa- and azacyclic dienes, including those tethered with an electron-deficient olefin. The oxacyclic 1,3,8-nonatrienes and 1,3,9 decatrienes undergo intramolecular Diels-Alder reactions upon heating in toluene, yielding tricyclic tetrahydropyran and -furan derivatives with excellent diastereoselectivities. PMID- 10864762 TI - Organoyttrium-catalyzed sequential Cyclization/Silylation reactions of nitrogen heteroaromatic dienes demonstrating "Aryl-Directed" regioselectivity AB - The reaction of 1-allyl-2-vinyl-1H-pyrroles and 1-allyl-2-vinyl-1H-indoles with arylsilanes in the presence of catalytic [Cp(TMS)(2)Y(&mgr;-Me)](2) leads to highly selective cyclization/silylation events. In this process the active catalyst for the reaction, "Cp(TMS)(2)YH", undergoes initial olefin insertion at the vinyl group. Even isopropenyl substituents on the heteroaromatics react in preference to less sterically encumbered allyl groups. Furthermore, the observed regioselectivity reflects an "aryl-directed" process, whereby the more highly substituted secondary or tertiary organometallic is initially generated. This intermediate undergoes cyclization onto the remaining alkene and subsequent silylation by a sigma-bond metathesis reaction, affording the observed products. PMID- 10864763 TI - Regiochemical studies of the ring expansion reactions of hydroxy azides with cyclic ketones. AB - The regiochemistry of ring expansions of 2-substituted cyclic ketones using 1,2 azidoethanol and 1,3-azidopropanol was examined. It was determined that the reactions of ketones with an adjacent methyl or ethyl group are generally unselective, but that bulkier substituents lead to preferential migration of the more highly substituted carbon. In addition, it was found that ketones bearing inductively electron-withdrawing substituents (OMe, Ph, Br) undergo selective migration of the less highly substituted carbon. For some substrates, alternative reaction pathways were also identified. PMID- 10864764 TI - An unexpected rearrangement during mitsunobu epimerization reaction of sugar derivatives AB - Mitsunobu reaction on the glucose derivative (3S,4R,5R,6R)-3,4,5, 7 tetrabenzyloxy-6-hydroxy-1-heptene yielded an unexpected rearrangement major product. Its structure was determined as (3R,4R, 5R,6S)-4,5,6,7-tetrabenzyloxy-3 hydroxy-1-heptene. The suggested rearrangement mechanism involves an initial intramolecular cyclization, followed by ring opening by the nucleophile p nitrobenzoate. Product distribution of the Mitsunobu reaction was substrate dependent, with the corresponding mannose derivative (the 3R epimer) giving less of the initial intramolecular reaction products and the corresponding galactose derivative (the 5S epimer) yielding almost exclusively the expected epimerization product. Varying the Mitsunobu reaction conditions (addition of base and using nonpolar solvent) led to the expected epimerization product of the glucose derivative. PMID- 10864765 TI - Hydrolysis mechanisms for the acetylpyridinephenylhydrazone ligand in sulfuric acid AB - The excess acidity method was applied to rate data obtained for 2 acetylpyridinephenylhydrazone hydrolysis in strong acid media using various aqueous/organic solvents, and it was observed that the reaction rate decreases with increasing permittivity of the medium. Two hydrolysis mechanisms are indicated. Below 0.6 M H(2)SO(4), no hydrolysis was observed; between 0.6 and 6.0 M H(2)SO(4), the substrate hydrolyzes by an A-S(E)2 mechanism and switches to an A-2 mechanism at higher acidity. This change of mechanism was justified on the basis of the syn and anti rotational conformers of the diene -N((1))=C-C=N((2))- group; the greater stability of the former can be explained by the formation of hydrogen bonds between the proton and the N((1)) and N((2)) nitrogen atoms, giving rise to a very stable five-membered ring. If the syn conformer is predominant, then no hydrolysis is observed; above 0.6 M, the attack of a second proton gives rise to a balance between the syn and anti forms, the latter being responsible for the hydrolysis of the hydrazone group. PMID- 10864766 TI - Intramolecular carbolithiation reactions for the preparation of Azabicyclo AB - Tin-lithium exchange and intramolecular carbolithiation (anionic cyclization) have been used to construct the three nitrogen-positional isomers of the azabicyclo[2.2.1]heptane ring system. The 7-azabicyclo[2.2.1]heptane ring system is accessed from either diastereomer of a 2,5-disubstituted pyrrolidine, via a chiral organolithium intermediate. The 2-azabicyclo[2.2.1]heptane ring system is formed stereoselectively in low yield by a tandem cyclization, together with the product from monocyclization. Better yields of the 2-aza ring system can be obtained using an alternative approach from a 2-tributylstannyl-4 allylpyrrolidine, despite the trans arrangement of the tin (and, hence, lithium) atom and the allyl unit. The 1-azabicyclo[2.2.1]heptane ring system is accessed in just three steps from 4-piperidone. PMID- 10864767 TI - Synthesis and properties of conjugated hybrid tetrathiafulvalene dimers AB - The synthesis of new hybrid tetrathiafulvalene (TTF) dimers (11a-c) has been carried out by a Wittig-Horner reaction of the respective phosphonate esters (10a c) with 2-(tetrathiafulvalenylvinyl)-9, 10-anthraquinone (9) prepared by olefination of formyltetrathiafulvalene (7) and the phosphonium salt of anthraquinone 8. Electrochemical studies show that the dimers 11a-c mainly retain the electrochemical properties of both TTF and the pi-extended TTF components, and most importantly, intramolecular electronic interactions between the two moieties are observed by cyclic voltammetry and Osteryoung square wave voltammetry. Semiempirical PM3 calculations reveal an almost planar geometry for the TTF and the benzene ring connected through the vinyl spacer. These compounds can form stable charge-transfer complexes with 2, 3-dichloro-5,6-dicyano-1,4 benzoquinone (DDQ) showing a stoichiometry of 1:3 (D:A). Attempts to electrocrystallize the dimeric donors with different counteranions are discussed. PMID- 10864768 TI - Alkaloid-fullerene systems through photocycloaddition reactions. AB - The photocycloaddition of tertiary amines to ?60fullerene (C(60)) is an interesting and useful reaction. We wished to extend the applications of this type of reaction through an investigation of the photoaddition of alkaloids to C(60) for the purpose of synthesizing novel and complex photoadducts that are difficult to obtain by usual methods. Irradiation of tazettine (2) or gramine (3) with C(60) in toluene leads to formation of one monoadduct (6 or 7), whereas scandine (1a) or 10-hydroxyscandine (1b) reacts with C(60) photochemically to give two products, the expected ?6,6 monoadduct (5a, 5b) and a new type of monoadduct with a bis-?6, 6 closed structure (4a, 4b). These new structures were characterized by UV-vis, FT-IR, (1)H NMR, (13)C NMR, (1)H-(1)H COSY, ROESY, HMQC (heteronuclear multiple-quantum coherence), and HMBC (heteronuclear multiple-bond connectivity) spectroscopy. The techniques of time-of-flight secondary ion MS (TOF-SIMS) and field desorption MS (FD-MS) were used for the mass determination. (3)He NMR analysis of the product mixture from photoaddition of 1a to C(60) containing a (3)He atom ((3)He@C(60)) led to two peaks at -9.091 and -11.090 ppm relative to gaseous (3)He, consistent with formation of a ?6, 6-closed monoadduct and a bis-?6,6 closed adduct. Presumably, the bis-?6, 6 closed adducts are formed by an intramolecular ?2 + 2 cycloaddition of the vinyl group to the adjacent 6,6 ring junction of C(60) after the initial photocycloaddition. PMID- 10864769 TI - A strategy for the solution-phase parallel synthesis of N (pyrrolidinylmethyl)hydroxamic acids. AB - Both five- and six-membered iminocyclitols have proven to be useful transition state analogue inhibitors of glycosidases. They also mimic the transition-state sugar moiety of the nucleoside phosphate sugar in glycosyltransferase-catalyzed reactions. Described here is the development of a general strategy toward the parallel synthesis of a five-membered iminocyclitol linked to a hydroxamic acid group designed to mimic the transition state of GDP-fucose complexed with Mn(II) in fucosyltransferase reactions. The iminocyclitol 8 containing a protected hydroxylamine unit was prepared from D-mannitol. The hydroxamic acid moiety was introduced via the reaction of 8 with various acid chlorides. The strategy is generally applicable to the construction of libraries for identification of glycosyltransferase inhibitors. PMID- 10864770 TI - Stable ion study of regioisomeric carboxonium-substituted pyrenium ions: directive effects, charge delocalization mode, and conformational aspects. AB - Regioisomeric monoacyl- and monobenzoyl-substituted pyrenes are diprotonated in FSO(3)H.SbF(5) (4:1)/SO(2)ClF to give persistent carboxonium-pyrenium dications, whereas diacetyl- and dibenzoylpyrenes are diprotonated to give dicarboxonium dications. The resulting dications were studied by low-temperature NMR at 500 MHz. Conformational aspects of the carboxonium group in various regioisomers are addressed by a combination of NOED spectra and 2D-NMR and AM1 calculations. Charge delocalization pathways are gauged and compared on the basis of the magnitude of Deltadelta (13)C values. PMID- 10864771 TI - Protein synthesis by solid-phase chemical ligation using a safety catch linker. AB - The native chemical ligation reaction has been used extensively for the synthesis of the large polypeptides that correspond to folded proteins and domains. The efficiency of the synthesis of the target protein is highly dependent on the number of peptide segments in the synthesis. Assembly of proteins from multiple components requires repeated purification and lyophilization steps that give rise to considerable handling losses. In principle, performing the ligation reactions on a solid support would eliminate these inefficient steps and increase the yield of the protein assembly. A new strategy is described for the assembly of large polypeptides on a solid support that utilizes a highly stable safety catch acid labile linker. This amide generating linker is compatible with a wide range of N terminal protecting groups and ligation chemistries. The utility of the methodology is demonstrated by a three-segment synthesis of vMIP I, a chemokine that contains all 20 natural amino acids and has two disulfide bonds. The crude polypeptide product was recovered quantitatively from the solid support and purified in 20%-recovered yield. This strategy should facilitate the synthesis of large polypeptides and should find useful applications in the assembly of protein libraries. PMID- 10864773 TI - On the conformational properties of PMID- 10864772 TI - Enantioselective diene Cyclization/Hydrosilylation catalyzed by optically active palladium bisoxazoline and pyridine-oxazoline complexes. AB - A 1:1 mixture of (N-N)Pd(Me)Cl ?N-N = (S,S)-4,4'-dibenzyl-4,5,4', 5'-tetrahydro 2,2'-bisoxazoline (S,S-4a) and NaBAr(4) ?Ar = 3, 5-C(6)H(3)(CF(3))(2) (5 mol %) catalyzed the asymmetric cyclization/hydrosilylation of dimethyl diallylmalonate (2) and triethylsilane at -30 degrees C for 48 h to form an 8.1:1 mixture of the silylated carbocycle (S,S)-trans-1, 1-dicarbomethoxy-4-methyl-3 ?(triethylsilyl)methylcyclop ent ane (S, S-3) (95% de, 72% ee) and dimethyl 3,4 dimethylcyclopentane-1, 1-dicarboxylate (S,S-6) in 64% combined yield. In comparison, a 1:1 mixture of the palladium pyridine-oxazoline complex (N N)Pd(Me)Cl ?N-N = (R)-(+)-4-isopropyl-2-(2-pyridinyl)-2-oxazoline (R-5b) and NaBAr(4) (5 mol %) catalyzed the asymmetric cyclization/hydrosilylation of 2 and triethylsilane at -32 degrees C for 24 h to form carbocycle S,S-3 in 82% yield (>95% de, 87% ee) as the exclusive product. Asymmetric diene cyclization catalyzed by complex R-5b was compatible with a range of functional groups and produced carbocycles with up to 91% ee. The procedure also tolerated substitution at a terminal olefinic position and at the allylic position of the diene. PMID- 10864774 TI - A new and efficient access to oxazoline-5-carboxylates and amino acid derivatives with cyclopropyl groups. PMID- 10864775 TI - Heterolytic and homolytic N-H bond dissociation energies of 4-substituted Hantzsch 2,6-dimethyl-1,4-dihydropyridines and the effect of one-electron transfer on the N-H bond activation. PMID- 10864776 TI - Fmoc: a more soluble analogue of the 9-fluorenylmethoxycarbonyl protecting group. PMID- 10864777 TI - Diastereoselective cyclopentane construction PMID- 10864778 TI - Lanthanide triflate catalyzed Biginelli reaction. one-pot synthesis of dihydropyrimidinones under solvent-free conditions. PMID- 10864779 TI - A straightforward route to stereodefined functionalized cycloheptanols and cyclooctanols. PMID- 10864781 TI - Stereoselective amination, bisamination, and Amination/Esterification of Bis(trimethylsilyl)-1,2-bisketene initiated by chiral amines PMID- 10864780 TI - Biomimetic synthesis of macrolide/ketolide metabolites through a selective N demethylation reaction. PMID- 10864782 TI - Solvent effect in the free-radical oxidation and electrophilic ipso and hydrogen displacement of p-methoxybenzyl alcohol and N-(p-Methoxybenzyl)acetamide by Br(2) PMID- 10864783 TI - An improved and regiospecific synthesis of trans-3,4-dihydrodiol metabolite of benzo[b]naphtho[2,1-d]thiophene. PMID- 10864784 TI - Differential regulation of pig theca cell steroidogenesis by LH, insulin-like growth factor I and granulosa cells in serum-free culture. AB - The regulation of pig theca cell steroidogenesis was studied by the development of a physiological serum-free culture system, which was subsequently extended to investigate potential theca-granulosa cell interactions. Theca cells were isolated from antral follicles 6-9 mm in diameter and the effects of plating density (50-150x10(3) viable cells per well), LH (0.01-1.0 ng ml(-1)), Long R3 insulin-like growth factor I (IGF-I) (10, 100 ng ml(-1)) and insulin (1, 10 ng ml(-1)) on the number of cells and steroidogenesis were examined. The purity of the theca cell preparation was verified biochemically and histologically. Co cultures contained 50x10(3) viable cells per well in granulosa to theca cell ratio of 4:1. Wells containing granulosa cells only were supplemented with 'physiological' doses of androstenedione or 100 ng ml(-1). Oestradiol production by co-cultures was compared with the sum of the oestradiol synthesized by granulosa and theca cells cultured separately. Oestradiol and androstenedione production continued throughout culture. High plating density decreased steroid production (P < 0.01). LH increased androstenedione (P < 0.001) and oestradiol (P < 0.05) synthesis and the sensitivity of the cells increased with time in culture. Oestradiol production was increased by 10 ng IGF-I ml(-1) (P < 0.001) but androstenedione required 100 ng ml(-1) (P < 0.001). Co-cultures produced more oestradiol than the sum of oestradiol synthesized by theca and granulosa cells cultured separately (P < 0. 001), irrespective of the androstenedione dose. This serum-free culture system for pig theca cells maintained in vivo steroidogenesis and gonadotrophin responsiveness. Thecal androstenedione and oestradiol production were differentially regulated and were primarily stimulated by LH and IGF-I, respectively. Theca-granulosa cell interactions stimulated oestradiol synthesis and this interaction was mediated by factors additional to the provision of thecal androgen substrate to granulosa cells. PMID- 10864785 TI - Ultrastructure of the basal lamina of bovine ovarian follicles and its relationship to the membrana granulosa. AB - Different morphological phenotypes of follicular basal lamina and of membrana granulosa have been observed. Ten preantral follicles (< 0. 1 mm), and 17 healthy and six atretic antral follicles (0.5-12 mm in diameter) were processed for light and electron microscopy to investigate the relationship the between follicular basal lamina and membrana granulosa. Within each antral follicle, the shape of the basal cells of the membrana granulosa was uniform, and either rounded or columnar. There were equal proportions of follicles 200 is the best indicator for a small proportion of grade 1 and 2 oocytes (poor quality), a large proportion of grade 3 and 4 oocytes (good quality), and a small proportion of oocytes with cytoplasmic inclusions. These results will be of clinical use in evaluating oocyte quality. PMID- 10864787 TI - In situ analysis of the changes in expression of ovarian inhibin subunit mRNAs during follicle recruitment after ovulation in pigs. AB - In situ hybridization was used on frozen tissue sections with digoxigenin labelled antisense riboprobes to inhibin/activin alpha and beta(A) subunits to determine whether inhibin/activin subunit mRNA expression was associated with development of growing, steroidogenically active follicles during follicle recruitment after ovulation. Cell proliferation-associated nuclear antigen Ki-67 protein and cytochrome P450 aromatase expression in granulosa cells were determined immunohistochemically and used as markers for granulosa cell proliferation and steroidogenesis, respectively, on days 3, 5 and 7 after the onset of oestrus. The amounts of inhibin/activin alpha and beta(A) subunit mRNA and P450 aromatase protein were greater (102, 93, and 238%, respectively; P < 0.05) in medium than in small non-atretic follicles and were positively correlated with Ki-67 and with each other. Inhibin/activin alpha and beta(A) mRNA, P450 aromatase, and Ki-67 in granulosa cells were reduced by 66-83% (P < 0.001) in atretic follicles compared with non-atretic follicles. In addition, inhibin/activin alpha and beta(A) mRNA and P450 aromatase in small (1-2 mm) non atretic follicles decreased (P < 0.05) between day 3 and day 7 independently of morphological or biochemical signs of atresia. The pattern of inhibin/activin subunit mRNA expression supports the notion that activin and inhibin have roles in growth and steroidogenesis in follicle recruitment during the early luteal phase of the oestrous cycle. PMID- 10864788 TI - Difference in localization of eosinophils and mast cells in the bovine ovary. AB - The bovine ovary contains a considerable number of leucocytes which can be located with an antibody against the CD18 molecule. In the present study, subtyping and cell counting were carried out on histological sections stained with Sirius red for eosinophils and with toluidine blue for mast cells. The CD18(+) cells were identified immunohistologically. Eosinophils and mast cells contributed considerably to the CD18(+) pool. The number of eosinophils in the corpus luteum increased rapidly in early development to approximately 90% of the CD18(+) cells, and decreased to 30% during secretion and to 10% during regression. Mast cells were not detectable in the follicles, the corpus luteum and the periphery of the cortex, but were observed in the interstitial cortical stroma and the medulla. The number of mast cells in these regions, which corresponded to 60-76% of the CD18(+) cells, did not change significantly throughout the oestrous cycle. It is concluded that eosinophils are selectively recruited at the periovulatory period and that mast cells are unevenly distributed. PMID- 10864789 TI - Influence of season and low-level oestradiol immunoneutralization on episodic LH and testosterone secretion and testicular steroidogenic enzymes and steroidogenic acute regulatory protein in the adult ram. AB - The regulation of LH-dependent and -independent increases in testosterone secretion by key proteins in the testes of adult rams was investigated. Serial blood samples were collected from groups of four control and passively immunized (oestradiol antiserum for 3 weeks) rams and the animals were gonadectomized in either the non-breeding season (April) or the breeding season (September). LH pulse frequency and basal (interpulse) concentrations were several times greater (P < 0.01) in the breeding season than in the non-breeding season. Neither of these parameters nor LH pulse amplitude were affected by oestradiol immunization. Parameters of testosterone episodic secretion and response to an injection (i.v.) of 15 micrograms NIH-LH-S25 were also greater (P < 0.05) in the breeding season and, with the exception of pulse frequency, in immunized rams versus controls. Substrate utilization established that testosterone biosynthesis was predominantly via the 5-ene pathway. Increases in blood testosterone concentration in the breeding season were associated with a fivefold higher (P < 0.01) activity of cytochrome P450 17alpha-hydroxylase/C-17,20 lyase (P450(17alpha)) and a 65% higher (P < 0.05) relative amount of mRNA for cytochrome P450 cholesterol side-chain cleavage enzyme complex (P450scc) in the testis. Of the steroidogenic enzyme activities examined, only that for 17beta hydroxysteroid dehydrogenase (17beta-HSD) tended to be increased by oestradiol immunization. Blood concentrations of cholesterol lipoproteins and expression of the testicular low density lipoprotein receptor were not affected by season or immunization. The amount of steroidogenic acute regulatory protein (StAR) mRNA was 65% higher (P < 0.01) in the breeding season and 20% higher (P < 0.01) in immunized rams versus controls. These results indicate that greater LH stimulation may increase testosterone biosynthesis in the breeding season by increasing StAR mRNA (and presumably delivery of cholesterol to P450scc) and the activity of P450(17alpha), and possibly that of P450scc (activity not measured). More moderate increases in StAR mRNA and 17beta-HSD activity may explain, in part, the increases in testosterone secretion with oestradiol immunization. PMID- 10864790 TI - Post-coital sperm recovery and cryopreservation in the Sumatran rhinoceros (Dicerorhinus sumatrensis) and application to gamete rescue in the African black rhinoceros (Diceros bicornis). AB - Sumatran rhinoceros (Dicerorhinus sumatrensis) sperm samples were collected from a post-copulatory female and characterized to determine their potential for sperm preservation and future use in artificial insemination. Five samples of acceptable quality from one male were used to compare the effect of two cryoprotectants (glycerol and dimethyl sulfoxide (DMSO)) and two post-thaw protocols (untreated and glass wool column) on sperm quality. The percentage of motile spermatozoa, sperm motility index (0-100) and sperm morphology were evaluated subjectively, and viability and acrosomal status were assessed using fluorescent markers. Evaluations of frozen-thawed spermatozoa were performed over a 6 h incubation interval. Post-coital semen samples (n = 5; 104.0 +/- 9.1 ml; 2.5 +/- 0.8 x 10(9) total spermatozoa; mean +/- SEM) exhibited a sperm motility index of 56.7 +/- 3.3, and contained 40.2 +/- 6.3%, 72.0 +/- 3.2% and 79.8 +/- 6.5% normal, viable and acrosome-intact spermatozoa, respectively. Glycerol and DMSO were equally effective as cryoprotectants and, regardless of post-thaw protocol, samples retained greater than 80% of all pre-freeze characteristic values. Processing semen samples through glass wool yielded higher quality samples, but only half the total number of motile spermatozoa compared with untreated samples. High values for pre-freeze sperm characteristics were also maintained after cryopreservation of epididymal spermatozoa from one black rhinoceros (Diceros bicornis) using the same protocol. In summary, Sumatran rhinoceros spermatozoa of moderate quality can be collected from post-copulatory females. Rhinoceros sperm samples show only slight reductions in quality after cryopreservation and thawing and have potential for use in artificial insemination. PMID- 10864791 TI - Optimal physicochemical conditions for the manipulation and short-term preservation of koala (Phascolarctos cinereus) spermatozoa. AB - Protocols for the successful manipulation and preservation of semen in a given species depend upon a fundamental knowledge of how spermatozoa respond to the physicochemical conditions of the extension media; methods developed for the preservation of eutherian spermatozoa may not necessarily be suitable for marsupial semen. The aim of this study was to investigate the effects on koala sperm motility of serial dilution, changes in temperature, diluent pH and osmolality to establish the optimal physicochemical conditions for short-term semen storage. This study showed that electroejaculated koala semen diluted 1∶1 (v/v) with PBS frequently coagulated after incubation at 35 degrees C, but that further dilution and incubation resulted in a corresponding increase in the percentage of spermatozoa swimming in a non-linear trajectory. The effect of rapid temperature change on the motility of koala spermatozoa was investigated by exposing semen, initially diluted at 35 degrees C, to temperatures of 45, 25, 15 and 5 degrees C. Although sperm motility was reduced after incubation at 45 degrees C, a rapid decrease in temperature of up to 20 degrees C did not result in a significant reduction in sperm motility. However, contrary to evidence in other marsupials, there was a small but significant decrease in sperm motility after rapid cooling of diluted semen from 35 to 5 degrees C. The effects of diluent pH and osmolality on the motility of koala spermatozoa were investigated. These experiments indicated that diluents for koala sperm manipulation should buffer in a pH range of 7-8 and have an osmolality of approximately 300 mmol kg( 1). The final experiment compared the relative effectiveness of Tris-citrate buffer (1% glucose) and PBS to maintain koala sperm motility over a range of incubation temperatures (5-35 degrees C) for up to 8 days. Reduction in sperm motility was directly related to temperature, and motility was sustained for the longest duration when stored at 5 degrees C. The Tris-citrate buffer solution was superior to PBS as a preservation diluent at all temperatures, and koala spermatozoa remained motile even after 42 days storage at 5 degrees C. Spermatozoa diluted in PBS (with Ca(2+) or Mg(2+)) and cooled to 5 degrees C showed evidence of an unusual motility pattern, similar to that of hyperactivated eutherian spermatozoa. This study showed that koala spermatozoa respond to different physicochemical conditions associated with short-term liquid storage in essentially the same way as the spermatozoa of eutherian mammals, although koala spermatozoa appear to be more tolerant of rapid temperature shock. The results of this study can be used to make informed selections with regard to appropriate diluent composition and improved short-term sperm preservation protocols and represent the first such database for any species of marsupial. PMID- 10864792 TI - Cyclical changes in sperm volume during in vitro incubation under capacitating conditions: a novel boar semen characteristic. AB - The osmotic reactivity of boar spermatozoa during incubation in vitro was studied using a hypo-osmotic swelling test in conjunction with electronic measurement of cell volume. Sperm populations showed fluctuations in both iso-osmotic cell volume and hypo-osmotic volume response that fitted mathematical models for periodicity. Significant differences of frequency and amplitude were observed during sperm incubation under capacitating conditions as compared with those under non-capacitating conditions. In addition, different boars showed specific differences in their fluctuation characteristics under capacitating conditions. During incubation under capacitating conditions, a decrease in osmotic reactivity was observed that correlated with a decrease in motility, while the absolute value of the earliest maximum of the osmotic-induced response correlated with an increase in the proportion of discharged acrosomes. The time course of the cyclical behaviour of osmotic reactivity may be a useful parameter for assessing boar sperm response to capacitating conditions. PMID- 10864793 TI - Impact of photoperiodic exposures during late gestation and lactation periods on the pineal and reproductive physiology of the Indian palm squirrel, Funambulus pennanti. AB - Studies on the maternal transfer of photoperiodic information in mammals indicate that the daily photoperiod perceived by the mother during the gestation-lactation period is communicated to the fetus either through the placenta or via the milk. However, the impact of photoperiodic exposures during gestation and lactation on the maternal pineal and reproductive physiology has not been reported for any tropical rodent. The exposure of pregnant female Indian palm squirrels (Funambulus pennanti) to constant light (24 h light:0 h dark), constant dark (0 h light:24 h dark), long daylength (14 h light:10 h dark) or short daylength (10 h light:14 h dark) during early gestation (< 30 days) resulted in the resorption of pregnancy, while during late gestation (> 30 days), it did not interfere with the maintenance of pregnancy. Alterations in photoperiodic condition during late gestation and lactation altered the postpartum recovery process. Pineal gland activity, as assessed by pineal mass, protein content and plasma melatonin, was lowest during the breeding phase, but increased gradually after parturition until the next breeding phase. During gestation and lactation, constant light, long daylength and short daylength conditions were less effective, while constant dark condition had a profound effect in depressing pineal gland activity, which subsequently advanced postpartum recovery. Hence, lactating females under constant darkness prepare themselves for next mating much earlier than females under natural daylength (12 h light:12 h dark) conditions. Therefore, photoperiodic information, mediated via the pineal gland, may be important for maintaining gestation physiology as well as postpartum recovery in female rodents. PMID- 10864794 TI - Effect of nutrition and superovulation on oocyte morphology, follicular fluid composition and systemic hormone concentrations in ewes. AB - The objective was to determine the effect of dietary intake on follicle and oocyte morphology in unstimulated and superovulated ewes. Fifty-four ewes were fed grass meal at 0.5, 1.0 or 2.0 times maintenance energy requirements (M) for 32 days. Oestrous cycles were synchronized using progestagen pessaries and either unstimulated or superovulated with 200 mg pig FSH. The ewes were killed and ovaries were collected either 36 or 12 h before the anticipated LH surge. Serum progesterone concentrations in ewes on day 10 after withdrawal of the pessary were lower in ewes fed 2.0M than in ewes fed 0.5M or 1.0M (P < 0.05). LH pulse frequency tended to be higher in ewes fed 2M than 1M (1.0 +/- 0.3 versus 0.3 +/- 0.2 pulses per 8 h) on day 6 after removal of the pessary but the effect was not significant. In unstimulated ewes, more follicles (>/= 3 mm) were observed when the animals were killed in ewes fed 2.0M (3.5 +/- 0.3) than in ewes fed 0.5M (2.4 +/- 0.3) or 1.0M (2.4 +/- 0.5; P < 0. 05). Fewer follicles were observed in superovulated ewes on 0.5M (7. 5 +/- 1.2) than in ewes on 1.0M (12.0 +/- 0.5) or 2.0M (12.3 +/- 1. 4; P < 0.05). Follicular fluid progesterone concentrations were higher in ewes fed 0.5M compared with those fed 1M or 2M (P < 0.05). Insulin-like growth factor (IGF)-I concentrations were higher in follicular fluid from ewes on 1M compared with either those on 0.5M or 2M (P < 0.05), whereas IGF-II concentrations were lower in follicular fluid from ewes on 2M compared with those on 1M or 0.5M (P < 0.05). Superovulation increased follicular fluid progesterone, oestradiol, IGF-I and IGF-II concentrations (P < 0.01). Concentrations of the 34, 22 and 20 kDa IGF binding proteins were lower in follicles from superovulated ewes compared with unstimulated ewes (P < 0.05). Oocytes from superovulated ewes showed abnormalities such as premature activation of cumulus expansion and vacuolation of the nucleolus and increased frequency of detachment of interchromatin-like granules from the nucleolar remnant. Collectively, these results indicate that both high and low dietary intakes can alter systemic and follicular fluid hormone concentrations. Relative to dietary effects, the effects of superovulation were greater and involved substantial increases in follicular fluid hormone concentrations and abnormal oocyte morphology. PMID- 10864795 TI - Ultrastructural evidence of transplacental transport of immunoglobulin G in bitches. AB - In dogs, passive immunity is conferred to fetuses and neonates by the transfer of maternal immunoglobulin G through the placenta during the last trimester of pregnancy and via the mammary gland after parturition, respectively. However, morphological evidence of transplacental transport is still lacking. The aim of the present study was to localize maternal immunoglobulin G in the labyrinthine zone and in the haemophagous zone of the canine placenta by means of immunohistochemistry and immunocytochemistry. In the labyrinthine zone, immunoglobulin G was detected in all the layers of the materno-fetal barrier including the fetal capillaries. Immunoreactivity was particularly prominent in maternal basement membrane material as well as in the syncytiotrophoblast. However, this evidence of transplacental transport of immunoglobulin G originated from a limited number of unevenly distributed maternal vessels only. In the cytotrophoblast of the haemophagous zone, immunoglobulin G was localized to phagolysosomes at various stages but was never detected within fetal vessels. The results indicate that maternal immunoglobulin G is degraded in cytotrophoblast cells of the hemophagous zone and, therefore, that transplacental transport is restricted to a subpopulation of maternal vessels in the labyrinthine zone. PMID- 10864796 TI - Postnatal growth rate and gonadal development in circadian tau mutant hamsters reared in constant dim red light. AB - The role of the circadian clock in the reproductive development of Syrian hamsters (Mesocricetus auratus was examined in wild type and circadian tau mutant hamsters reared from birth to 26 weeks of age under constant dim red light. Testis diameter and body weights were determined at weekly intervals in male hamsters from 4 weeks of age. In both genotypes, testicular development, subsequent regression and recrudescence exhibited a similar time course. The age at which animals displayed reproductive photosensitivity, as exhibited by testicular regression, was unrelated to circadian genotype (mean +/- SEM: 54 +/- 3 days for wild type and 59 +/- 5 days for tau mutants). In contrast, our studies revealed a significant impact of the mutation on somatic growth, such that tau mutants weighed 18% less than wild types at the end of the experiment. Our study reveals that the juvenile onset of reproductive photoperiodism in Syrian hamsters is not timed by the circadian system. PMID- 10864797 TI - Localization of calcium and zinc in the sperm storage tubules of chicken, quail and turkey using X-ray microanalysis. AB - Sperm storage tubules from the utero-vaginal junction of chickens, quails and turkeys were analysed for calcium and zinc using X-ray microanalysis of ultra rapidly frozen tissue in a scanning electron microscope. This technique enabled the tubular fluid surrounding the stored spermatozoa and the intracellular content of the cells of the sperm storage tubules to be analysed separately and, by using standards with known concentrations, their elemental concentrations were estimated. The mean (+/- SEM) concentration of calcium in the tubular fluid from chickens, quails and turkeys was 17 +/- 3, 19 +/- 3 and 17 +/- 4 mmol kg(-1) wet weight, respectively. The intracellular calcium concentration of the cells of the tubules did not differ significantly from these values and was also similar in the mucosal epithelial cells of the utero-vaginal junction. Zinc was localized in the cells of turkey sperm storage tubules and tubular fluid, but at low concentrations. No zinc could be detected in corresponding structures from chickens and quails. The concentration of calcium in the tubular fluid is within the range known to inhibit the motility of spermatozoa, supporting this function for calcium during storage. Zinc is known to depress turkey sperm metabolism and it may also be involved in inducing quiescence of spermatozoa during storage in this species. PMID- 10864798 TI - Na+-Ca2+ exchange in mouse oocytes: modifications in the regulation of intracellular free Ca2+ during oocyte maturation. AB - Increases in intracellular free Ca(2+)+ concentration (Ca(2+)+ oscillations) occur during meiotic maturation and fertilization of mammalian oocytes but little is known about the mechanisms of Ca(2+) homeostasis in these cells. Cells extrude Ca(2+) from the cytosol using two main transport processes, the Ca(2+)-ATPase and the Na(+)-Ca(2+) exchanger. The aim of this study was to determine whether Na(+) Ca(2+) exchange activity is present in immature and mature mouse oocytes. Na(+) Ca(2+) exchange can be revealed by altering the Na(+) concentration gradient across the plasma membrane and recording intracellular free Ca(2+) concentrations using Ca(2+)-sensitive fluorescent dyes. Depletion of extracellular Na(+) caused an immediate increase in Ca(2+) concentration in immature oocytes and a delayed increase in mature oocytes. The Na(+) ionophore, monensin, caused an increase in intracellular Ca(2+) in immature oocytes similar to that induced by Na(+) depleted medium. In mature oocytes, monensin had no effect on intracellular Ca(2+) but the time taken for Ca(2+) to reach a peak value on removal of extracellular Na(+) was significantly decreased. Finally, addition of Ca(2+) to immature oocytes incubated in Ca(2+)-free medium caused an increase in the concentration of intracellular Ca(2+) that was dependent upon the presence of extracellular Na(+). This effect was not seen in mature oocytes. The data show that Na(+)-Ca(2+) exchange occurs in immature and mature mouse oocytes and that Ca(2+) homeostasis in immature oocytes is more sensitive to manipulations that activate Na(+)-Ca(2+) exchange. PMID- 10864799 TI - Number of chromocentres in the nuclei of mouse Sertoli cells in relation to the strain and age of males from puberty to senescence. AB - Nuclei of mouse Sertoli cells were examined on air-dried toluidine blue-stained preparations to analyse factors influencing the aggregation of heterochromatin into chromocentres. For the CBA strain males tested at 1.3, 2, 3, 6, 9 and 12 months of age, mean numbers of heterochromatin bodies were 4.8, 2.4, 2.1, 1.8, 1.6 and 1. 4, respectively; two chromocentres predominated from 2 to 9 months of age. In the KE strain, heterochromatin aggregation was significantly accelerated; nuclei containing only one chromocentre were predominant from 6 months. The number of chromocentres did not change with the stages of the seminiferous cycle, and after 1 month of cryptorchid condition. Cryptorchidism resulted in disruption of spermatogenesis and Sertoli cell dysfunction, as demonstrated by the lack of immunohistochemically detectable androgen receptors. The difference in the number of chromocentres between KE and CBA Sertoli cells persisted after 3 days of in vitro culture, but unidentified cells with numerous chromatin bodies were also observed. Testing recombinant inbred strains indicates that at least two genes are involved in the difference in the number of chromocentres between progenitor KE and CBA strains; however, no correlations were found with 15 marker loci or with parameters linked to reproduction. Of the eight strains tested, AKR and C3H showed a 'CBA-like' chromocentre pattern; C57BL, B10.BR, B10.BR-Y(del) and KP were 'KE-like'; and BALB/c and DBA/2 were intermediate. The results showed that centromere aggregation in the Sertoli cell progresses throughout the life of a male in a strain specific manner; however, its functional significance remains unknown. PMID- 10864800 TI - Effect of administering a crude equine gonadotrophin preparation to mares on follicular development, oocyte recovery rate and oocyte maturation in vivo. AB - In mares, the shortage of oocytes and the variability in nuclear maturation at a certain time of the oestrous cycle hinders the optimization of methods for in vitro maturation and in vitro fertilization. Increasing the number of small-to medium-sized follicles available for aspiration in vivo may increase the overall oocyte yield. The aims of the present study were to investigate whether administration of crude equine gonadotrophins affects follicular development, oocyte recovery rate, in vivo oocyte maturation and follicular concentrations of meiosis-activating sterols. During oestrus, all follicles >/= 4 mm were aspirated from 19 pony mares (first aspiration: A1). Over the next 8 days, the mares were treated daily with either 25 mg crude equine gonadotrophins (n = 10) or physiological saline (n = 9). Between day 1 and day 8, follicular growth was monitored by ultrasonography. On day 8, all follicles >/= 4 mm were evacuated (second aspiration: A2) and nuclear maturation of the recovered oocytes was assessed after orcein staining. Follicular growth between A1 and A2, as well as the number and size of follicles at A2 were similar for control mares and mares treated with crude equine gonadotrophins. The oocyte recovery rates at A1 and A2 were similar. At A2, the oocyte recovery rate and oocyte maturation in vivo were not affected by treatment with crude equine gonadotrophins. The number of expanded cumulus oophorus complexes recovered from follicles 95 microm) and cultured for 8 or 9 days on granulosa cell monolayers. Within 8 days of culture, the mean oocyte diameter increased from 86 +/- 0.4 microm to 95 +/- 0.7 microm in group Aand from 106 +/- 0.2 microm to 109 +/- 0.5 microm in group B. After 9 days of culture, the mean diameter of oocytes from groups A and B were 99 +/- 0.5 microm and 112 +/- 0.4 microm, respectively. The meiotic competence of oocytes grown in vitro was evaluated by in vitro maturation. Within 8 days of culture, only 3% of oocytes from group A and 6% of oocytes from group B acquired the ability to undergo germinal vesicle breakdown. After 9 days of culture, 7% of group A oocytes and 42% of group B oocytes were competent to resume meiosis. The expression of p34(cdc2) in oocytes grown in vitro was analysed by the western blot technique. During 9 days of culture, p34(cdc2) accumulated in both groups of growing oocytes, but its concentration was lower than in fully grown oocytes used as controls. The results showed for the first time that goat oocytes from early antral follicles can grow, accumulate p34(cdc2) and acquire the ability to resume meiosis, when cultured for 9 days on granulosa cell monolayers. PMID- 10864803 TI - E-cadherin-catenin complex in the rat ovary: cell-specific expression during folliculogenesis and luteal formation. AB - The cadherins and their cytoplasmic counterparts, the catenins, form the adherens junctions, which are of importance for tissue integrity and barrier functions. The development and maturation of the ovarian follicle is characterized by structural changes, which require altered expression or function of the components involved in cell-cell contacts. The present study examined the cell specific localization and temporal expression of epithelial cadherin (E-cadherin) and alpha- and beta-catenin during follicular development, ovulation and corpus luteum formation in the immature gonadotrophin- and oestrogen-stimulated rat ovary. Immunohistochemistry and immunoblotting demonstrated the expression of E cadherin in theca and interstitial cells of immature ovaries before and after injection of equine chorionic gonadotrophin (eCG). E-cadherin was not detected in granulosa cells, except in the preantral follicles located to the inner region of the ovary. The content of E-cadherin in theca and interstitial cells decreased after an ovulatory dose of hCG. Granulosa cells of apoptotic follicles did not express E-cadherin. Oestrogen treatment (diethylstilboestrol) of immature rats for up to 3 days did not result in a measurable expression of E-cadherin in granulosa cells. alpha- and beta-catenin were expressed in all ovarian compartments. The concentration of beta-catenin was constant during the follicular phase, whereas the content of alpha-catenin decreased in granulosa cells after treatment with diethylstilboestrol or hCG. The expression of alpha catenin was also reduced in theca and interstitial cells after hCG. alpha- and beta-catenin were present in most ovarian cells at all stages of folliculogenesis. Therefore, the catenins have the potential to associate with different members of the cadherin family and to participate in the regulation of cytoskeletal structures and intracellular signalling. The restricted expression of E-cadherin in granulosa cells of preantral follicles indicates a role in the recruitment of these follicles to subsequent cycles. The specific decrease of alpha-catenin in granulosa cells and the reduction of both alpha-catenin and E cadherin in theca cells of ovulatory follicles might reflect some of the molecular changes in cell-cell adhesion associated with ovulation and luteinization. PMID- 10864804 TI - Production of interferon by red deer (Cervus elaphus) conceptuses and the effects of roIFN-tau on the timing of luteolysis and the success of asynchronous embryo transfer. AB - The role of interferon in early pregnancy in red deer was investigated by (a) measuring production of interferon by the conceptus, (b) testing the anti luteolytic effect of recombinant interferon-tau in non-pregnant hinds, and (c) treatment of hinds with interferon after asynchronous embryo transfer. Blastocysts were collected from 34 hinds by uterine flushing 14 (n = 2), 16 (n = 2), 18 (n = 8), 20 (n = 13) or 22 (n = 9) days after synchronization of oestrus with progesterone withdrawal. Interferon anti-viral activity was detectable in uterine flushings from day 16 to day 22, and increased with duration of gestation (P < 0.01) and developmental stage (P < 0.01). When interferon-tau was administered daily between day 14 and day 20 to non-pregnant hinds to mimic natural blastocyst production, luteolysis was delayed by a dose of 0.2 mg day(-1) (27.3 +/- 1.3 days after synchronization, n = 4 versus 21 +/- 0 days in control hinds, n = 3; P < 0.05). Interferon-tau was administered to hinds after asynchronous embryo transfer to determine whether it protects the conceptus against early pregnancy loss. Embryos (n = 24) collected on day 6 from naturally mated, superovulated donors (n = 15) were transferred into synchronized recipients on day 10 or day 11. Interferon-tau treatment (0.2 mg daily from day 14 to 20) increased calving rate from 0 to 64% in all recipients (0/11 versus 7/11, P < 0.005), and from 0 to 67% in day 10 recipients (0/8 versus 6/9, P < 0.01). The increased success rate of asynchronous embryo transfer after interferon-tau treatment in cervids may be of benefit where mismatched embryo maternal signalling leads to failure in the establishment of pregnancy. PMID- 10864805 TI - Glycosylation in the near-term epitheliochorial placenta of the horse, donkey and camel: a comparative study of interbreeding and non-interbreeding species. AB - Studies from this laboratory have shown great diversity in the glycosylation of tissues comprising the interhaemal barrier of species with different placental types. This diversity may be one of the factors preventing interbreeding between species. Glycan expression within the uterine epithelium and trophoblast of the interhaemal barrier was examined to test this proposition in three species with similar diffuse, microcotyledonary, epitheliochorial allantochorionic types of placenta: the horse (Equus caballus) and donkey (Equus asinus), which can interbreed with each other, and the camel (Camelus dromedarius), which cannot interbreed with either of the other two species. A panel of 14 lectins was used and it was found that glycosylation patterns were generally similar between placental tissues of the horse and donkey, except for the expression of non bisected complex N-glycan and some sialic acids, whereas those of the camel showed striking differences in the binding of lectins to many structures carrying terminal residues of fucose, N-acetyl galactosamine and beta-galactose, as well as to complex N-glycans and sialic acids. These results are consistent with the proposition that interbreeding species carry similar glycans in tissues forming the interhaemal barrier whereas glycodiversity is one of the factors preventing implantation and subsequent placental development in interspecific hybrids. PMID- 10864806 TI - Effect of a high maternal dietary intake during mid-gestation on components of the utero-placental insulin-like growth factor (IGF) system in adolescent sheep with retarded placental development. AB - The aim of the present study was to investigate the effects of administering a high plane diet during early to mid-gestation on the uterine and placental insulin-like growth factor (IGF) system and on systemic IGF-I concentrations in pregnant adolescent ewes with restricted placental growth. Embryos recovered from superovulated ewes inseminated by a single sire were transferred in singleton to the uterus of adolescent recipients. After transfer ewes were offered a high (H) or moderate (M) amount of a complete diet calculated to promote rapid or normal maternal growth rates, respectively. Five ewes from each group were switched from either M to H or H to M diets at day 52 of gestation. Maternal and fetal blood samples and placental tissues were collected from all animals at day 104. Ewes on the high plane diet from mid-gestation (HH, MH groups) had restricted placental mass (P < 0.01) and tended to have smaller fetuses. This was associated with increased maternal plasma IGF-I concentrations (P < 0.001). The pattern of expression of components of the IGF system in the uterus and placenta was studied by in situ hybridization. IGF-I mRNA concentrations were below the limit of detection. IGF-II mRNA expression was high in the fetal mesoderm and present in maternal stroma, but was not influenced by nutritional treatment. In contrast, IGF binding protein 1 (IGFBP-1) mRNA expression was higher (P < 0.05) and IGFBP-3 mRNA expression was lower (P < 0.05) in the endometrial glands of ewes in HH and MH groups. In the fetal trophoblast, IGFBP-3 mRNA expression was higher in the MH group. Type 1 IGF receptor expression was increased (P < 0. 01) in the luminal epithelium of the HM group and IGFBP-2 mRNA expression was highest in the placentome capsule of ewes in the HH group. Together, these results indicate that reprogramming of the uterine and placental IGF axis by maternal nutrition could contribute to placental growth retardation in growing adolescent sheep. PMID- 10864807 TI - Time-related changes in canine luteal regulation: in vivo effects of LH on progesterone and prolactin during pregnancy. AB - The role of LH in luteal function in pregnant dogs was investigated at two different periods during pregnancy: (i) the transitional period from apparent total independence of the corpus luteum to relative hormonal dependence (days 20 35); and (ii) the period of full hormonal dependence (days 35-40). At both periods, LH neutralization, LH inhibition and LH administration studies were conducted. At both periods LH immunoneutralization had no significant effect on the secretion pattern of progesterone or prolactin. GnRH antagonist treatment (Nal-Glu) decreased plasma LH below the detection limit in all treatment periods. Nal-Glu had no effect on prolactin. When GnRH antagonist osmotic pumps were implanted, a transient decrease in plasma progesterone concentrations occurred on days 21-22 but not during the remaining implantation period. When GnRH antagonist was injected, plasma progesterone temporarily decreased (24 h) after the beginning of treatment starting on day 20, but decreased for 5 days when the treatment started on day 35. When purified pig LH was injected i.v. twice a day for 2 consecutive days either from day 30 or from day 40, plasma progesterone concentrations remained constant during treatment. However, on days 40 and 41, an increase in prolactin was observed. These results indicate that LH immunoneutralization may not impair corpus luteum function. In addition, GnRH antagonist induces dose- and time-dependent effects. Only high doses resulted in a decrease in progesterone, the duration of which increased as pregnancy progressed. Continuous GnRH antagonist administration, even when associated with complete LH inhibition, was not associated with detectable effects on progesterone. Finally, LH administration does not stimulate progesterone but may modify prolactin in the last third of pregnancy. Other studies indicated a corpus luteum prolactin dependency. The present study indicates that, in pregnant bitches, LH may not be necessary to sustain progesterone synthesis but that its role may vary in a time-dependent manner. PMID- 10864808 TI - In vitro culture of brushtail possum (Trichosurus vulpecula) epididymal epithelium and induction of epididymal sperm maturation in co-culture. AB - A medium modified from eutherian systems was used to culture epididymal epithelial cells of the brushtail possum (Trichosurus vulpecula) for more than 2 months. Epididymal tubule fragments from the caput, corpus and cauda epididymides were used to generate cell monolayers. All three epididymal cell culture systems supported maturational changes in marsupial spermatozoa and enabled immature possum spermatozoa to differentiate from a T-shape to a streamlined shape, accompanied by the development of progressive motility after co-culture with 7 day-old cultured epididymal cell monolayers. This epididymal cell and sperm co culture system for marsupial species may facilitate the identification of specific epithelial factors that affect sperm maturation, particularly in a species in which morphological maturation is readily visible. PMID- 10864809 TI - Differences between antigenic determinants of pig and cat zona pellucida proteins. AB - Despite many efforts, the control of reproduction in feral cat populations is still a problem in urban regions around the world. Immunocontraception is a promising approach; thus the present study examined the suitability of the widely used pig zona pellucida proteins (pZP) for contraception in feral domestic cats. Purified zona pellucida proteins obtained from pig and cat ovaries were used to produce highly specific antisera in rabbits. Antibodies against pZP raised in rabbits or lions were not effective inhibitors of either in vitro sperm binding (cat spermatozoa to cat oocytes) or in vitro fertilization in cats, whereas antibodies against feline zona pellucida proteins (fZP) raised in rabbits showed a dose-dependent inhibition of in vitro fertilization. Immunoelectrophoresis, ELISA and immunohistology of ovaries confirmed these results, showing crossreactivity of anti-fZP sera to fZP and to a lesser extent to pZP, but no interaction of anti-pZP sera with fZP. It is concluded that cat and pig zonae pellucidae express a very small number of shared antigenic determinants, making the use of pZP vaccine in cats questionable. A contraceptive vaccine based on feline zona pellucida determinants will be a better choice for the control of reproduction in feral cats if immunogenity can be achieved. PMID- 10864810 TI - Uterine lymphocyte distribution and interleukin expression during early pregnancy in cows. AB - Both the production of cytokines and the distribution of immune cells within the uterus change during early pregnancy. Evidence obtained mainly from mice indicates that these changes are important for implantation and in preventing a maternal immune response to the conceptus. The ruminant embryo also produces interferon tau at this time, the signal for the maternal recognition of pregnancy. The relationship between these events in cows was studied using uteri from three groups of animals on day 16 after observed oestrus: (i) cyclic controls, (ii) pregnant and (iii) inseminated but with no embryo present. Embryo size and the antiviral activity in uterine flushings (indicative of the interferon tau concentration) were measured. Sections of intact uterus were frozen for the localization and quantitation of CD4(+) (T lymphocytes), CD14(+) (macrophages) and CD21(+) (B lymphocytes) uterine cells by immunohistochemistry. The expression of interleukin (IL)-1alpha, IL-2, IL-6 and IL-10 mRNAs in uterine extracts was measured by RT-PCR. Neither embryo size, interferon tau concentration nor pregnancy status influenced the distribution of CD4(+), CD14(+) or CD21(+) cells in the day 16 uterus. Endometrial IL-1alpha mRNA was detected in most cows across the groups, whereas IL-2 mRNA was only present in the non pregnant uterus. IL-6 and IL-10 mRNAs were not detectable in any uteri. In conclusion, IL-2 mRNA expression is detectable in the non-pregnant but not the pregnant uterus on day 16 and interferon t is unlikely to play a role in the redistribution of immune cells in the uterus during early bovine pregnancy. PMID- 10864811 TI - Anovulation in non-reproductive female Damaraland mole-rats (Cryptomys damarensis). AB - Within colonies of Damaraland mole-rats (Cryptomys damarensis), anovulation in non-reproductive females is thought to play an important role in maintaining reproductive skew. Pituitary sensitivity and ovarian structure were examined in three groups of females that differed with respect to their social environment and breeding status to determine whether anovulation is due to inhibitory social cues or is merely the result of a lack of copulatory stimulation. The contribution of gonadal steroid negative feedback to neuroendocrine differences in the reproductive systems of the respective groups was also investigated. LH secretion after a 0.5 micrograms GnRH challenge in females that had been removed from the presence of the breeding individuals for at least 6 months (removed non reproductive females) was significantly higher than in non-reproductive females in the colony, but significantly lower than in reproductive females. In both removed non-reproductive females and reproductive females, corpora lutea were observed in ovaries of seven of eight females, indicating that ovulation occurs spontaneously in subordinate females on removal from the breeding pair. Circulating progesterone concentrations in removed non-reproductive females were significantly higher than in non-reproductive females, indicating that circulating progesterone is not responsible for infertility in non-reproductive females. Indeed, after hystero-ovariectomy, reproductive females continued to show significantly greater GnRH-stimulated LH secretion than non-reproductive females. Thus, differential inhibition of gonadotrophin secretion in breeding and non-breeding females occurs independently of gonadal steroids. It is concluded that female Damaraland mole-rats are spontaneous ovulators and that anovulation results from inhibitory social cues within the colony, not a lack of copulatory stimulation. Since non-reproductive females are infertile, inhibition of the hypothalamo-pituitary-gonadal axis has the potential to play a causal role in maintaining reproductive skew in colonies of C. damarensis. PMID- 10864812 TI - Classification of small type B/C follicles as primordial follicles in mature rats. AB - In the present study, follicles were classified according to the morphology of their granulosa cells. Type B follicles contained only flattened granulosa cells; type B/C follicles had a mixture of flattened and cuboidal granulosa cells in a single layer, and type C follicles had a single layer of cuboidal granulosa cells. The primary objectives of the study were to determine whether 5-bromo-2 deoxyuridine incorporation into type B/C follicles was a marker for initiation of growth and how long type B/C follicles could remain at the same stage before transformation to type C follicles. Female Holtzman rats received bromo deoxyuridine for 7 days. After the infusion (day minipumps were removed = day 0), rats were ovariectomized on days 0 (n = 9), 30 (n = 8), 90 (n = 8) and 150 (n = 9). The numbers of type B, B/C and C follicles within one ovary were determined using modified fractionator counting. Analysis over all times demonstrated that there were more (P < 0.0001) type B/C (941 +/- 61 per ovary) than type C (140 +/- 18 per ovary) or type B (159 +/- 19 per ovary) follicles. The numbers of type B and type C follicles did not differ from each other at any time. Only one of 34 rats evaluated had bromo-deoxyuridine-labelled type B follicles. On day 150, 57% of the bromo-deoxyuridine-labelled type B/C follicles remained from day 0. It is concluded that (1) DNA synthesis in granulosa cells of type B/C follicles is not a reliable indicator of impending growth; and (2) type B and type B/C follicles are both components of the pool of primordial follicles. PMID- 10864813 TI - Association of the developing acrosome with multiple small Golgi units, the Golgi satellites, in spermatids of the musk shrew, Suncus murinus. AB - The spermatozoa of the musk shrew, Suncus murinus, have a fan-like giant acrosome with a diameter of approximately 20 mm. The aim of this study was to investigate how this giant acrosome is constructed in the musk shrew spermatid and, in particular, how the Golgi apparatus involved in acrosome formation behaves. The behaviour of the Golgi apparatus was monitored by confocal laser scanning microscopy with antibody against a Golgi-associated Rab6 small GTPase. In the early Golgi phase, small Golgi units, the Golgi satellites, localized as a large aggregate in the juxtanuclear cytoplasm. As acrosome formation progressed, the Golgi satellites gradually dispersed, associated with proacrosomal vesicles and an acrosomal vesicle, and finally became distributed as multiple small units over the whole surface of an acrosomal cap in the round spermatid. The mode of acrosome formation in musk shrews was distinctly different from that in rats and mice, in which the Golgi apparatus remains as a single unit throughout acrosome formation. In musk shrews, the proacrosomal vesicles formed successively by the Golgi satellites coalesced, one after another, into a potential acrosomal vesicle. This process may result in further enlargement of the acrosome. The results of the present study indicate that Golgi satellites are necessary for the biogenesis and development of the giant acrosome in musk shrew spermatozoa. PMID- 10864814 TI - Regulation of beta-catenin mRNA and protein levels in human villous cytotrophoblasts undergoing aggregation and fusion in vitro: correlation with E cadherin expression. AB - The cellular mechanisms underlying the formation and organization of the human placenta remain poorly understood. Recent studies have demonstrated that E cadherin, in association with the cytoplasmic protein known as beta-catenin, plays an integral role in the differentiation of the trophectoderm in the murine and bovine embryo. Although E-cadherin expression is regulated during the aggregation and fusion of human villous cytotrophoblasts, the expression of beta catenin during the terminal differentiation of these primary cell cultures has not been determined. In this study, beta-catenin mRNA concentrations and protein expression were examined in primary cultures of human villous cytotrophoblasts using northern and western blot analysis. beta-catenin mRNA concentrations and protein expression were high in freshly isolated mononucleate cytotrophoblasts but decreased as these cells underwent aggregation and fusion to form syncytium. A similar pattern of expression was observed for the E-cadherin mRNA transcript and protein species present in these cell cultures. Immunoprecipitation studies demonstrated that the beta-catenin and E-cadherin protein species present in the mononucleate cytotrophoblasts were capable of forming intracellular complexes. In contrast, beta-catenin and E-cadherin mRNA and protein expression in JEG-3 choriocarcinoma cells remained constant over time in culture. beta-catenin and E cadherin expression was subsequently immunolocalized to the aggregates of mononucleate cells present in both of these trophoblastic cell cultures and the villous cytotrophoblasts of the human first trimester and term placenta. Taken together, these observations indicate that the E-cadherin-beta-catenin complex plays a central role in the terminal differentiation of human trophoblasts in vitro and in vivo. PMID- 10864815 TI - Three-dimensional magnetic resonance imaging for the study of ovarian function in a bovine in vitro model. AB - Three-dimensional magnetic resonance imaging coupled with maximum intensity projection display, a technique usually reserved for magnetic resonance imaging angiography, is useful for the study of ovarian follicular growth. The ovaries of 19 cows were examined each day by transrectal ultrasonography. From these data, the precise phase of the ovarian cycle was determined and cows were ovariectomized on day 3 of wave one (n = 5), on day 6 of wave one (n = 4), on day 1 of wave two (n = 4), >/= 17 days after ovulation (n = 5), and on the day of ovulation (n = 1). The excised ovaries were examined by magnetic resonance imaging using a fast imaging with steady state precession imaging sequence with maximum intensity projection reconstruction, displayed as a cine-loop of the ovaries rotating in space. This provided the clearest view among the three principal three-dimensional steady state data acquisition approaches tried; the follicles and other ovarian structures could be distinguished unambiguously. Results from the bovine model indicate that the acuity of the three-dimensional fast imaging with steady state precession technique has potential application in in vivo intravaginal imaging in women for studying normal and pathological ovarian function. PMID- 10864816 TI - Sperm binding capacity and ultrastructure of the zona pellucida of stored canine oocytes. AB - Sperm binding to the zona pellucida is a prerequisite for fertilization, and tests that evaluate this function have been described for several species. When carrying out such tests in the canine species, ovaries or oocytes have to be stored to obtain a sufficient number of oocytes at the time of testing. In the present study, the sperm binding capacities of salt-stored oocytes and oocytes from deep frozen ovaries were measured and compared with that of fresh oocytes. Two different procedures for washing the sperm-oocyte complexes (gentle and tough) were used before evaluating the number of bound spermatozoa. The total number of oocytes that bound spermatozoa was significantly lower for both salt stored and deep frozen oocytes compared with fresh oocytes. Significantly fewer spermatozoa bound to stored oocytes than to fresh oocytes (P 8 mm in diameter in the ovary ipsilateral to the previously gravid uterine horn, did not affect the rate of uterine involution or plasma 15-keto 13,14-dihydro-prostaglandin F(2alpha) concentration. In conclusion, administration of eCG to increase follicular growth and oestradiol production overcame the inhibition of follicular growth in the ovary ipsilateral to the previously gravid uterine horn, but did not affect uterine involution. PMID- 10864827 TI - Sex-dependent frequency and type of autosomal univalency at the first meiotic metaphase in mouse germ cells. AB - Univalents at the first meiotic metaphase in mouse spermatocytes occur mainly in the XY pair, making it difficult to compare the amounts of univalency in males and females. In this study, the amounts of autosomal univalency in male and female meiosis were compared using the model strain CBA-T6, in which univalency of the small marker autosome pair T6 has been shown to occur very frequently in spermatocytes. Mice from inbred CBA and DBA strains were also analysed. The total frequencies of univalency (sex chromosomes plus autosomes) in metaphase I spermatocytes were 45.6% in CBA, 36.9% in CBA-T6, and 37.3% in DBA males. The aneuploidy in metaphase II spermatocytes ranged from 1.4 to 3% in these strains, which was in agreement with previous findings that most primary spermatocytes with abnormal chromosome configurations are arrested in their development before metaphase II. In the CBA-T6 strain, autosomal univalency at metaphase I mostly involved chromosome pair T6; however, its frequency differed significantly between the sexes, amounting to 18.9% in spermatocytes and 4.3% in oocytes. In the CBA strain, autosomal univalents at metaphase I were seen in 7.7% of the spermatocytes and 1.4% of the oocytes and, in DBA mice, in 4.9% of the spermatocytes and 3.8% of the oocytes. However, in DBA oocytes, when univalency occurred it usually concerned a greater number of bivalents in one cell (range: 2 19 disjoined bivalents), a phenomenon very rare in males of this strain. This study shows that univalent formation differs between the male and female types of meiosis. PMID- 10864828 TI - The Parkes lecture. Mutations of gonadotrophin and gonadotrophin receptor genes: what do they teach us about reproductive physiology? AB - Although mutations in human gonadotrophin and gonadotrophin receptor genes are rare, they have greatly elucidated the physiology and pathophysiology of gonadotrophin action. These 'nature's transgenics' have been corroborated by mouse transgenic and knock-out models. An inactivating mutation of the human LHbeta chain and knock-out of the mouse common alpha-chain show that pituitary LH is not needed to stimulate fetal testicular steroidogenesis and male sexual differentiation. In mice, early testicular steroidogenesis is apparently gonadotrophin-independent and, in humans, it is regulated by placental hCG. Pituitary LH becomes necessary only after birth. Inactivating LH receptor mutations block prenatal hCG action, thus inhibiting male-type sexual differentiation. In females, this process is autonomous, and LH becomes important only at puberty; inactivation of LH receptor causes anovulatory infertility. Activating LH receptor mutations cause male-limited gonadotrophin-independent precocious puberty in males, but no apparent phenotype in females. Animal models for LH or LH receptor inactivation are not yet available. Inactivating FSH ligand and receptor mutations cause infertility because of a lack of follicular maturation in women. Findings in men are controversial, since FSHbeta inactivation is related to azoospermia, whereas the cognate receptor inactivation only suppresses spermatogenesis without causing absolute infertility. The FSHbeta and FSH receptor knock-out mice display phenocopies of the human FSH receptor mutation. Information about activating FSH receptor mutations is still insufficient. Hence, the above human mutations have brought important new information about the role of gonadotrophins in reproductive functions. The genetically modified animal models provide useful tools to explore the pathogenesis and new treatment modalities of infertility, and to develop new contraceptive strategies. PMID- 10864829 TI - Detection of steroidogenic acute regulatory protein in equine ovaries. AB - A steroidogenic acute regulatory (StAR) protein has been identified in several species as a probable important rate-limiting step in steroidogenesis. This protein is believed to be responsible for transporting cholesterol from the outer to the inner mitochondrial membrane. It is known that equine chorionic gonadotrophin (eCG) stimulates steroidogenesis in the corpora lutea of early pregnant mares and that eCG also upregulates StAR mRNA in bovine ovaries. In the present study, ovarian tissue from cyclic and early pregnant mares was immunostained to detect the distribution of the StAR protein. Western blot analysis was performed, followed by phosphor imaging to establish whether the onset of eCG secretion in pregnancy was associated with increased expression of the StAR protein. Immunostaining for StAR was confined to the theca interna of growing and preovulatory follicles, but 24 h after treatment with hCG, some granulosa cells were positively stained. Positive staining was confined to the large luteal cells of the equine corpus luteum. There was no difference in the distribution of immunostaining before or after onset of eCG secretion in pregnant mares, but increased amounts of StAR were detected in corpora lutea from mares at day 40 or day 41 of pregnancy compared with non-pregnant mares and mares at days 20-30 of pregnancy. PMID- 10864831 TI - Evaluation of chilled and frozen-thawed canine spermatozoa using a zona pellucida binding assay. AB - Zona pellucida binding assays provide information about the fertilizing ability of spermatozoa. A zona-binding assay for canine spermatozoa using intact, denuded homologous oocytes has not been evaluated previously. In the present study, an assay using canine oocytes derived from frozen-thawed ovaries was evaluated using three types of semen: fresh untreated; killed; and a 50:50 mixture of untreated and killed spermatozoa. The assays were performed on 3 x 20 oocytes for each sperm treatment, using semen from pooled ejaculates (0.5 x 10(6) spermatozoa in each 50 microliter droplet containing five oocytes). There was a significant difference (P < 0. 001) between all treatments. Thereafter, the same procedure was used to evaluate methods of chilling and freeze-thawing of canine semen. There was a trend (P = 0.067) for more sperm binding after 1 day of chilling compared with after 4 days of chilling. Semen samples frozen using an extender (with or without the addition of Equex STM paste) were evaluated. Equex had a significant (P = 0.034) positive effect on the capacity of the spermatozoa to bind to the zona pellucida. In conclusion, the addition of a zona pellucida binding assay to established in vitro tests should give a better estimate of the damage caused by the various procedures when developing new techniques for chilling and freeze-thawing. Furthermore, the present study showed that chilling for 4 days tended to reduce the zona-binding capacity of the spermatozoon, and that Equex STM paste had a beneficial effect on the capacity of the frozen-thawed spermatozoon to bind to the zona pellucida. PMID- 10864830 TI - Improvement of follicular development rather than gonadotrophin secretion by thyroxine treatment in infertile immature hypothyroid rdw rats. AB - Despite extensive study of reproductive abnormalities in female hypothyroid animals, little is known of folliculogenesis and gonadotrophin secretion in spontaneously hypothyroid animals, especially in response to exogenous hormone treatment. In this study, follicular development and plasma hormone concentrations in the presence or absence of thyroxine and eCG treatment were investigated in infertile immature spontaneously hypothyroid rdw rats. Administration of thyroxine once a day from day 21 to day 29 after birth resulted in increases in body weight (P < 0.001) and ovary mass on day 30 (P < 0.01). Similar populations of both healthy and atretic antral follicles ranging from 101 to 400 micrometer in diameter were observed in control rdw and normal rats. In rdw rats, thyroxine treatment markedly increased the number of healthy antral uniovular follicles 101-400 or > 550 micrometer in diameter in the absence or presence of eCG, respectively. Combined treatment of thyroxine and eCG in rdw rats also markedly increased the number of healthy antral biovular follicles. Thyroxine treatment did not affect the population of atretic antral follicles, but resulted in decrease in the number of atretic large antral follicles (> 400 microm) in the presence of eCG. Plasma oestradiol concentrations in rdw rats given both thyroxine and eCG were significantly higher than they were in rdw rats given eCG alone (P < 0.001). There were no significant differences in plasma FSH concentrations on day 28 between rdw (10.7 +/- 1.6 ng ml(-1)) and normal rats (12.0 +/- 1.4 ng ml(-1); P > 0. 05). Although there were no significant differences in plasma LH concentrations between control rdw (1.9 +/- 0.1 ng ml( 1)) and normal rats on day 30 (1.8 +/- 0.1 ng ml(-1); P > 0.05), eCG treatment increased plasma LH to a peak concentration 52 h after injection in normal (24.9 +/- 2.4 ng ml(-1)) but not in rdw rats treated with thyroxine (4.8 +/- 0.3 ng ml( 1); P < 0.05). In conclusion, the results of the present study indicate that thyroxine treatment improves follicular development but does not rescue the defect of the preovulatory surge of LH in eCG-primed rdw rats. PMID- 10864832 TI - Opioidergic, dopaminergic and adrenergic regulation of LH secretion in prepubertal heifers. AB - Studies have shown inhibitory effects of endogenous opioids on LH secretion in early post-natal heifers. However, it is not clear whether these effects change during the rest of the prepubertal period or whether the inhibitory influences on the GnRH neurones are direct or by way of other neuronal systems. Two experiments were performed in heifer calves to study the developmental patterns of opioidergic, dopaminergic and adrenergic regulation of LH and the possible interactions between opioids and dopaminergic and adrenergic neuronal systems, in the regulation of LH secretion. In Expt 1 four groups each of five heifer calves were used. Blood samples were taken every 15 min for 10 h and each calf received one of the following treatments as a single injection at 4, 14, 24, 36 and 48 weeks of age: (i) naloxone (opioid antagonist, 1 mg kg(-1), i. v.); (ii) sulpiride (dopamine D2 antagonist, 0.59 mg kg(-1), s.c.); (iii) naloxone and sulpiride combined; or (iv) vehicle (control group). Treatments began after the first blood sample was taken. The design of Expt 2 was similar; a separate group of heifer calves was assigned to receive one of the following treatments as a single injection at 4, 14, 24, 36 and 48 weeks of age: (i) naloxone; (ii) phenoxybenzamine (an alpha-adrenoreceptor blocker, 0.8 mg kg(-1), i. v.); (iii) naloxone and phenoxybenzamine; (iv) or vehicle. Results from Expt 1 showed that the maximum concentration of LH and the number of calves responding to treatments with an LH pulse was higher in the first hour after treatments at 36 and 48 weeks of age in the naloxone group compared with the control or sulpiride groups (P < 0.05). These values in the naloxone group also increased over time and were greatest at 48 weeks of age (P < 0.05). In heifers given naloxone + sulpiride treatment at 36 and 48 weeks of age, maximum concentrations of LH in the first hour after treatment did not differ from the naloxone and control groups. In Expt 2, at 36 and 48 weeks of age, treatment with naloxone with or without phenoxybenzamine resulted in higher concentrations of LH than in the controls (P < 0.05). No pulses were seen over the first hour of treatment at 36 and 48 weeks of age in heifers treated with phenoxybenzamine. The 10 h periods of blood sampling at 48 weeks of age revealed that phenoxybenzamine alone suppressed LH pulse frequency and mean serum concentrations of LH compared with the control group (P < 0.05). It was concluded that a strong or more acute inhibition of LH secretion by endogenous opioids developed in mid- to late prepubertal heifers, or alternatively, that removal of opioidergic inhibition at the GnRH neurone unmasked stimulatory inputs that were greater in heifers close to first ovulation. Since sulpiride appeared to negate in part the effects of naloxone on LH release, the suppressive effects of opioids could be exerted in part through the inhibition or blocking of a stimulatory dopaminergic system. alpha-Adrenergic neuronal systems have stimulatory effects on LH release, especially during the late prepubertal period, but do not appear to mediate opioidergic inhibition of LH secretion in prepubertal heifer calves. PMID- 10864833 TI - Pulsatile secretion pattern of growth hormone during the luteal phase and mid anoestrus in beagle bitches. AB - The pulsatile secretion pattern of growth hormone was investigated during four stages of the luteal phase and during mid-anoestrus in six cyclic beagle bitches. Plasma samples were obtained via jugular venepuncture at 10 min intervals for 12 h at 19 +/- 2 (mean +/- SEM; luteal phase 1), 38 +/- 2 (luteal phase 2), 57 +/- 2 (luteal phase 3), 78 +/- 2 (luteal phase 4) and 142 +/- 4 days (mid-anoestrus) after ovulation. During all stages, growth hormone was secreted in a pulsatile fashion. The mean basal plasma growth hormone concentration during luteal phase 1 (2.2 +/- 0.3 microgram l(-1)) was significantly higher than that during luteal phase 4 (1.5 +/- 0.1 microgram l(-1)) and mid-anoestrus (1.4 +/- 0.2 microgram l( 1)). The mean area under the curve (AUC) above zero during luteal phase 1 (27.3 +/- 2.7 microgram l(-1) in 12 h) tended to be higher than that during luteal phase 4 (20.8 +/- 1.8 microgram l(-1) in 12 h) and mid-anoestrus (19.2 +/- 2.5 microgram l(-1) in 12 h). In contrast, the mean AUCs above the baseline during luteal phase 1 (1.1 +/- 0.5 microgram l(-1) in 12 h) and luteal phase 2 (1.2 +/- 0.5 microgram l(-1) in 12 h) were significantly lower than that during luteal phase 4 (2.8 +/- 0.5 microgram l(-1) in 12 h). In conclusion, the pulsatile secretion pattern of growth hormone changes during the luteal phase in healthy cyclic bitches: basal growth hormone secretion is higher and less growth hormone is secreted in pulses during stages in which the plasma progesterone concentration is high. It is hypothesized that this change is caused by a partial suppression of pituitary growth hormone release by progesterone-induced growth hormone production in the mammary gland. The progesterone-induced production of growth hormone in the mammary gland may promote the physiological proliferation and differentiation of mammary gland tissue during the luteal phase of the bitch by local autocrine-paracrine effects. In addition, progesterone-induced mammary growth hormone production may exert endocrine effects, such as hyperplastic changes in the uterine epithelium and insulin resistance. PMID- 10864834 TI - Gonadotroph-lactotroph associations and expression of prolactin receptors in the equine pituitary gland throughout the seasonal reproductive cycle. AB - An interaction between gonadotroph and lactotroph cells of the pituitary gland has long been recognized in several species. The current study was conducted to investigate whether an association between gonadotrophs and lactotrophs occurs in mares and whether prolactin receptors are expressed within the pituitary gland of this species. The effects of both reproductive state and season on these variables were examined in pituitary glands obtained from sexually active mares in July (breeding season), sexually active mares in November (non-breeding season) and anoestrous mares in November. Pituitaries were dissected out immediately after death and immunofluorescent staining was carried out on 6 micrometer sections using specific antibodies to the LHbeta subunit, FSHbeta subunit, prolactin and prolactin receptor. Gonadotrophs were observed in both the pars distalis and pars tuberalis; although they appeared mostly as isolated cells, small groups of gonadotrophs were also identified in the pars distalis. In contrast, lactotrophs were observed only as clusters of cells exclusively in the pars distalis of sexually active and anoestrous mares in November and in most of the sexually active mares in July. A specific gonadotroph-lactotroph association was identified only between large isolated gonadotrophs and lactotroph clusters. Double immunofluorescent staining for FSHbeta and prolactin revealed a similar gonadotroph-lactotroph association to the one detected for LH gonadotrophs. No statistical difference in the gonadotroph:lactotroph ratio was observed as a result of changes in reproductive status or season. However, a tendency for a simultaneous decrease in the number of gonadotrophs and an increase in the number of lactotrophs was detected in anoestrous animals. Prolactin receptor immunoreactivity was found in the pars distalis, but not in the pars tuberalis, of sexually active (July and November) and anoestrous animals for both long and short forms of the receptor. No prolactin receptor co-localization for either form of the receptor was observed in LH or FSH gonadotrophs in either of the reproductive states examined during both summer and winter seasons. Furthermore, no significant difference was apparent in the proportion of cells expressing prolactin receptors between mares of different reproductive state or season. The specific anatomical association between gonadotroph and lactotroph cells and the expression of prolactin receptors in the equine pituitary gland indicate a potential role of prolactin in the regulation of gonadotrophin secretion. However, the absence of evidence for co-localization of prolactin receptors in LH or FSH cells does not support the hypothesis of a direct effect of prolactin on the gonadotroph as reported in a short day breeder. The results raise the possibility that, in horses, an intermediate regulatory cell may mediate the action of prolactin on gonadotroph function. PMID- 10864835 TI - Ultrasonographic imaging of the testis and epididymis of the bottlenose dolphin, Tursiops truncatus aduncas. AB - Eight male bottlenose dolphins, Tursiops truncatus aduncas, underwent examination of the reproductive organs to investigate the use of real-time B-mode ultrasonography in assessment of reproductive status and to establish normal ultrasonographic appearances. Ultrasonography allowed repeatable examinations which were well tolerated by all animals. Ultrasonography was used to examine the testes, epididymides, vasa deferentia, penis, bulbourethral and bulbocavernosal muscles; the prostate was not convincingly distinguished from surrounding muscles. Testicular echopatterns and size differed among individuals. Three distinct testicular echopatterns were discerned and could be used to differentiate males of different reproductive status. Ultrasonographic appearance of the testes provides useful data in assessing the reproductive status of male dolphins. PMID- 10864836 TI - Maturation-dependent glycoproteins containing both N- and O-linked oligosaccharides in epididymal sperm plasma membrane of rhesus monkeys (Macaca mulatta). AB - Glycosylation is one of the important post-translational modifications of sperm plasma membrane proteins during the maturation of epididymal spermatozoa that results in the development of motility and fertilizing capability. The aim of the present study was to identify and characterize the maturation-dependent asparagine-linked (N-linked) and serine- and threonine-linked (O-linked) glycoproteins of the epididymal spermatozoa of rhesus monkeys. The presence of N- and O-linked glycoproteins was confirmed by treatment of sperm membranes with N glycosidase F and O-glycosidase. The major maturation-dependent sperm membrane glycoproteins identified on blots of SDS-PAGE-fractionated proteins of purified sperm plasma membranes from five segments of epididymis, probed with biotinylated lectins and Vectastain-ABC reagent included O-linked 170, 150, 86 and 60/58 kDa glycoproteins; N-linked 68, 56, 48 and 38 kDa glycoproteins and N- and O-linked 116 kDa glycoprotein, all of which exhibited marked differences in the degree of glycosylation between immature and mature sperm surfaces. These glycoproteins can be used as markers of sperm maturation in the epididymis of rhesus monkeys, during the screening of antifertility agents acting at the epididymis, or may be developed as potential sperm antigens. The 100% inhibition of fertility in female rats and rabbits immunized with major maturation-dependent 116 kDa glycoprotein showed the significance of glycosylation changes in the maturation status of epididymal spermatozoa. This 116 kDa protein can be used as a marker parameter of sperm maturation in the rhesus monkey, which is often the preferred animal model for preclinical studies. These results will contribute to the identification of an appropriate animal model for the development of male contraceptives in humans. PMID- 10864837 TI - Effects of different activation treatments on fertilization of horse oocytes by intracytoplasmic sperm injection. AB - The effects of four reagents on the activation and subsequent fertilization of equine oocytes, and the development of these after intracytoplasmic sperm injection, were investigated. Cumulus-oocyte complexes collected from equine ovaries obtained from an abattoir were matured in vitro for 40-44 h in TCM199 medium before being injected, when in metaphase II, with an immobilized stallion spermatozoon. The cumulus-oocyte complexes were then subjected to one of five activation treatments: (a) 10 micromol ionomycin l(-1) for 10 min; (b) 7% (v/v) ethanol for 10 min; (c) 100 micromol thimerosal l(-1) for 10 min; (d) 250 micromol inositol 1,4, 5-triphosphate l(-1) injection; and (e) no treatment (control). After 18-20 h further culture, the cumulus-oocyte complexes were assessed for activation by observing whether they had progressed through second anaphase-telophase and had formed a female pronucleus. The proportions of oocytes activated after each treatment were: 16/27 (59%) for ionomycin; 14/25 (56%) for ethanol; 22/28 (79%) for thimerosal; 15/27 (56%) for inositol 1,4,5-triphosphate; and 0/20 (0%) for the untreated controls. Thus, significantly more oocytes (P < 0.05) were activated by treatment with thimerosal than by the other four treatments. The proportions of oocytes that cleaved to the two-cell stage at 24 30 h after sperm injection in the groups treated with ionomycin, ethanol and thimerosal were 7/20 (35%), 5/19 (26%) and 11/23 (48%), respectively. No cleavage was observed in any of the control oocytes or those treated with inositol 1,4, 5 triphosphate. Furthermore, evidence of normal fertilization was observed in 2/7 (29%), 2/5 (40%) and 7/11 (64%) of the oocytes treated with ionomycin, ethanol and thimerosal, respectively. These results demonstrated that: (a) it is possible to activate equine oocytes with the chemical stimulants, ionomycin, ethanol, thimerosal and inositol 1,4,5-triphosphate; (b) thimerosal is more effective than the other three reagents in facilitating both meiotic activation and normal fertilization of equine oocytes; and (c) chemical activation may also stimulate parthenogenetic cleavage of oocytes without concurrent changes in the head of the spermatozoon. PMID- 10864838 TI - Effects of androgens, progesterone and their antagonists on the developmental competence of in vitro matured bovine oocytes. AB - The aim of the present study was to determine whether androgens and progesterone influence the in vitro maturation of bovine oocytes as assessed by cleavage rates and competence to form blastocysts after in vitro fertilization. Bovine cumulus oocyte complexes were cultured (n = 20 per drop) for 22-24 h at 38.5 degrees C in TCM-199 medium supplemented with 10% oestrous cow serum, eCG (2.5 iu ml(-1)) and a range of treatments that included aromatizable (testosterone; 100 nmol l(-1)) and non-aromatizable (dihydrotestosterone; 100 nmol l(-1)) androgens, an androgen antagonist (flutamide; 36 micromol l(-1)), progesterone (300 nmol l(-1)) and a progesterone antagonist (mifeprisone, RU486; 100 nmol l(-1)). Production of inhibin A, total alpha-subunit, activin A and follistatin by each group of cumulus-oocyte complexes was also measured, since inhibin-related peptides have been implicated as modulators of oocyte maturation and their production may be influenced by steroids and anti-steroids. Both testosterone and dihydrotestosterone increased oocyte cleavage rate (25%; P < 0.01) and dihydrotestosterone also increased (24%; P < 0.05) the proportion of oocytes that reached the >/= eight-cell stage. However, neither androgen affected blastocyst yield, or the proportion of blastocysts that hatched. The stimulatory effect of dihydrotestosterone on cleavage rate was reduced by flutamide but the anti androgen had no effect when tested alone. Treatment with testosterone, but not dihydrotestosterone, decreased (P < 0.05) endogenous follistatin and increased (P < 0.05) the activin A:follistatin ratio in maturation medium. Concentrations of inhibin A, total alpha-subunit and activin A were not affected significantly by androgen or flutamide. Addition of progesterone or the anti-progestin mifepristone to cumulus-oocyte complexes had no effect on cleavage rate. However, progesterone reduced by approximately 40% (P < 0.05) the proportions of both total oocytes and cleaved oocytes that formed blastocysts. This effect was partially reversed by mifepristone. Neither progesterone nor mifepristone affected inhibin A, activin A or follistatin production. However, total alpha subunit concentration was significantly greater in the progesterone-treated group than in the controls (50%; P < 0.05), indicating that the negative effect of progesterone on blastocyst yield may be mediated by increased inhibin alpha subunit expression by cumulus cells. PMID- 10864839 TI - Chemical and thermal effects on the viability and motility of spermatozoa from the turtle epididymis. AB - The viability and motility of spermatozoa harvested from the epididymides of turtles were estimated to elucidate properties that might enable them to be stored over long periods of time. Spermatozoa from the painted turtle, Chrysemys picta, were analysed and compared with spermatozoa from two other turtles, Trachemys scripta and Sternotherus odoratus using the Cellsoft analysis system for videotaped images. Spermatozoa from C. picta and T. scripta, suspended in F 10 medium, showed low motility (3-6% motile) and motion velocities, whereas the motility of spermatozoa from S. odoratus was higher (40% motile). Spermatozoa from C. picta and S. odoratus, but not T. scripta, had higher motilities and motion velocities when incubated at 2 degrees C before analyses. C. picta spermatozoa were unresponsive to calcium concentrations ranging from 10(-8) to 10(-1) mol l(-1), potassium concentrations ranging from 0. 1 to 10 mmol l(-1), and to pH values in the range 5.9-8.4. Spermatozoa from C. picta were sensitive to hypo-osmotic media, and showed reduced motility at 25% of normal osmolarity and no motility at 10% of normal osmolarity. Distorted cells and missing flagellae were noted at 50% of normal osmolarity. C. picta spermatozoa were viable up to 40 days after harvest when incubated at 4 degrees C; during this time, both motility and motion velocity were increased in response to 0.5 mmol 3 isobutyl-1-methylxanthine l(-1). Spermatozoa from turtles have osmotic properties and resistance to changing chemical environments similar to spermatozoa from other vertebrates that have internal fertilization, and appear to be stable over long periods of time compared with spermatozoa from other vertebrate species. PMID- 10864840 TI - Development of a long-term serum-free culture system for immature granulosa cells from diethylstilboestrol-treated prepubertal rabbits: influence of androstenedione and fibronectin on FSH-induced cytodifferentiation. AB - Granulosa cells from diethylstilboestrol-treated prepubertal rabbits were cultured for 6 days in M199 with FSH (1-100 ng ml(-1)) in uncoated or fibronectin coated plates with or without androstenedione to define the time course profile of oestradiol and progesterone secretion, and the possible modulator role of androstenedione and fibronectin during FSH-induced rabbit granulosa cell differentiation. Every 48 h, cultures were photographed and samples of medium were collected and assayed by ELISA for oestradiol and progesterone. FSH increased oestradiol secretion in a dose-dependent manner. Androstenedione augmented FSH-stimulated oestradiol secretion, and led to a decrease in secretion of oestradiol with time in culture. FSH stimulated progesterone secretion in a dose-dependent manner. This was increased by androstenedione with 10 ng FSH ml( 1) (0-96 h) and 1 ng FSH ml(-1) (96-144 h). FSH-stimulated (100 ng ml(-1)) progesterone secretion decreased at 48-96 h. Fibronectin prevented this decrease, without affecting oestradiol or progesterone secretion at other time points. FSH caused cell reaggregation at 48 h. In conclusion, this serum-free culture system is appropriate for the study of mechanisms of rabbit granulosa cell differentiation. FSH induced cytodifferentiation and reaggregation of granulosa cells. Androstenedione appeared to act synergistically with FSH to promote steroidogenesis. Fibronectin sustained progesterone secretion during differentiation. PMID- 10864842 TI - Relationship between fertility in cattle and the number of inseminated spermatozoa. AB - Five different two-parameter models were fitted to published data from 30 studies to identify an approximate mathematical form of the relationship between fertility in cattle and the number of inseminated spermatozoa. In all cases, the first parameter defines the maximum attainable fertility, and the second scales the dose according to the percentage of the maximum attained. The best model was the hyperbolic dose-response curve used in pharmacology to analyse the effect of drugs. There is evidence that the semen of individual bulls differs in both parameters of the models and that therefore the viability of semen may be multidimensional. This might explain why measures of semen quality have hitherto been found to correlate poorly with fertility. The hypothesis that spermatozoa are subject to the law of mass action at the ovum predicts these and some other aspects of fertility, and indicates that heterospermic inseminations may provide an efficient way of estimating the parameters of semen. PMID- 10864841 TI - Oestradiol regulation of oxytocin receptor expression in cyclic bovine endometrium. AB - Oestradiol treatment can increase uterine oxytocin receptor expression in vivo. The actions of oestrogen are usually mediated via its receptor, but it also has direct non-genomic effects in some cells. This study investigated the effect of oestradiol and the role of the oestradiol receptor in regulating endometrial oxytocin receptor expression in the bovine uterus. Explant cultures (in triplicate) from late luteal phase non-pregnant endometrium received the following treatments: control (serum-free medium), oestradiol (0. 1 and 0.01 micromol l(-1)), oestradiol (0.1 micromol l(-1)) with the oestradiol receptor antagonist ICI 182780 (0.5 micromol l(-1)), and ICI 182780 (0.5 micromol l(-1)) alone. Explants were collected 12, 24 and 48 h after the start of culture. Oxytocin receptor mRNA expression in the explants was measured by in situ hybridization and oxytocin protein concentrations were measured by autoradiography with the iodinated oxytocin receptor antagonist d(CH(2))(5) [Tyr (Me)(2) Thr(4) Tyr NH(2)(9)]-vasotocin ((125)I-labelled oxytocin receptor antagonist). Oxytocin receptor mRNA and protein expression were initially low but spontaneous upregulation occurred in the luminal epithelium between 24 and 48 h (P < 0.01). Oestradiol increased oxytocin receptor mRNA upregulation in the first 24 h (P < 0.05) but the effect on 125 I-labelled oxytocin receptor antagonist binding was not significant. ICI 182780 inhibited the oestrogenic effect but had no significant effect on oxytocin receptor mRNA expression when given alone. In conclusion, the results showed that oestradiol exerts its effect via the oestradiol receptor. Oestradiol facilitates oxytocin receptor gene transcription by increasing it more rapidly, but spontaneous upregulation of endometrial oxytocin receptor still occurs in the absence of oestradiol. PMID- 10864843 TI - Novel approach to cell sampling from preimplantation ovine embryos and its potential use in embryonic genome analysis. AB - The major obstacle in the extensive analysis of the embryonic genome is the small number of cells typically obtained after the embryo biopsy. The object of the present study was to develop a simple approach that would allow the collection of a sufficient number of cells from a single embryo for use in further analyses. A micromanipulator was used to make a hole in the zona pellucida of 28 compacted morulae, 27 early blastocysts and 31 expanded blastocysts. After further culture, the trophoblastic cells, which herniated through this hole, were cut and cultured in vitro for different periods and used for embryo sexing. The results showed that biopsies can be taken successfully from 96.3% of early blastocysts, compared with 67.7% of expanded blastocysts and 71.4% of compacted morulae. The trophoblastic vesicles contained 20.8 +/- 6.7 cells (mean +/- SEM) and, when cultured, formed a confluent monolayer. The sex of cells cultured was assayed by PCR and the 12 lambs born after transfer of biopsied embryos confirmed its 100% accuracy. Moreover, no significant differences were found in the viability rates in vitro among blastocysts vitrified immediately after biopsy (77.8%), blastocysts biopsied and vitrified after 24 h culture (76.9%) and blastocysts vitrified without manipulation (88.5%). In experiments in vivo, the lambing rate of biopsied and vitrified blastocysts was significantly (P < 0.05) lower (40.0%) compared with vitrified control embryos (68.7%). This new approach to the biopsy of preimplantation embryos is a useful good model in the assisted reproductive technologies of domestic, wild and human species. PMID- 10864844 TI - Effect of gonadal steroids on pituitary LH secretion and mediobasal hypothalamic GnRH mRNA in ferrets. AB - In vitro release and content of GnRH in mediobasal hypothalamic slices are reduced by ovariectomy of female ferrets but are not affected by castration of male ferrets in breeding condition. The aim of the present study was to determine whether this sex difference reflects a sexually dimorphic effect of gonadal steroids on mediobasal hypothalamic GnRH mRNA content of male and female ferrets killed 4 weeks after gonadectomy, either with or without steroid hormone replacement. This time interval exceeds the 6-10 days needed for increments in plasma LH concentrations to stabilize after gonadectomy of ferrets of both sexes. In situ hybridization using an (35)S-labelled oligoprobe complementary to the human GnRH coding region showed that the number of mediobasal hypothalamic neurones and the cellular content of GnRH mRNA did not differ significantly among groups of male and female ferrets that were either in breeding condition or that had been gonadectomized and treated with sex steroids or oil vehicle. These results indicate that gonadal hormones regulate mediobasal hypothalamic GnRH biosynthesis and release in both sexes via post-transcriptional events that may include GnRH mRNA translation or the conversion of pre-pro GnRH precursor into mature GnRH. PMID- 10864845 TI - Circulating concentrations of inhibin-related proteins during the ovulatory cycle of the domestic fowl (Gallus domesticus) and after induced cessation of egg laying. AB - Circulating inhibin A, inhibin B, activin A, total immunoreactive inhibin alpha subunit (ir-alpha inhibin), LH, FSH and progesterone concentrations were measured throughout the normal ovulatory cycle and after cessation of egg laying induced by feed restriction to investigate the potential involvement of inhibins and activins in the ovulatory cycle of the domestic hen. Plasma inhibin A varied significantly (P < 0.05) during the ovulatory cycle; the concentration was highest at the preovulatory LH surge and reached a nadir 10 h later, at about the time the F(2) follicle makes the transition to become the new F(1) follicle. Plasma FSH concentrations did not change significantly throughout the cycle and showed no correlation with inhibin A. Total ir-alpha inhibin concentrations were much higher than those of inhibin A at all stages of the ovulatory cycle and showed no correlation with inhibin A or FSH. Plasma concentrations of inhibin B and of activin A were below the detection limit of the assays in all plasma samples analysed. In the feed restriction study, plasma inhibin A and total ir alpha inhibin showed little change until the last day of oviposition (day 0) after which they fell significantly (P < 0.05) and remained low to the end of the experiment (approximately 70-78% decrease relative to day -4). Conversely, plasma FSH increased after cessation of laying and was significantly higher (P < 0.05) from day 3 to the end of the study (approximately 50% increase on day 6 relative to day -4). Plasma FSH values were negatively correlated with inhibin A (r = 0.39; P < 0.005) and total ir-alpha inhibin (r = -0.36; P < 0.005). Plasma LH and progesterone also decreased (P < 0.05) during feed restriction. The decrease in LH preceded the terminal oviposition and the associated fall in inhibin A by 2 days; there was a positive correlation between LH and inhibin A (r = 0.35; P < 0.005). Taken together these findings support (i) a role for LH in promoting inhibin A secretion by preovulatory follicles and (ii) an endocrine role for inhibin A secreted by preovulatory follicles in the maintenance of tonic FSH secretion in laying hens. PMID- 10864846 TI - Increase in 15-hydroxyprostaglandin dehydrogenase activity in the ovine placentome at parturition and effect of oestrogen. AB - Type 1 NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) is the key enzyme for metabolism of active primary prostaglandins to inactive forms in gestational tissues. The present study examined the activity and immunolocalization of PGDH in the ovine placenta, fetal membranes and uterus over the latter half of pregnancy, and its potential regulation by oestradiol. Placenta, fetal membranes and myometrium were collected from sheep with known single insemination dates on days 70, 100 and 135 of gestation and in active labour demonstrated by electromyographic activity. In addition, chronically catheterized fetuses were infused with oestradiol (100 microgram kg(-1) per 24 h) (n = 5) or saline vehicle into the fetus from day 120 to day 125. PGDH activity measured in placental extracts remained constant from day 70 to day 135 of gestation, and then significantly (P < 0.05) increased by 300% in active labour. Immunoreactive PGDH was localized in the placentome at all stages and was present predominantly in the fetal component of the placentome in uninucleate, but not in binucleate, trophoblast cells. Similarly, in the fetal membranes PGDH immuno reactivity was present in the uninucleate trophoblast but not in the binucleate cells of the chorion. PGDH immunostaining was also present in the endometrial luminal epithelium, in the smooth muscle of the myometrium, and the glandular epithelium of the cervix. Infusion of oestradiol into the fetal circulation from day 120 to day 125 of gestation had no effect on placental PGDH activity. Immunohistochemistry was used to localize oestrogen receptor alpha in intrauterine tissues to investigate further the failure of oestradiol to increase PGDH activity. Immunoreactive oestrogen receptor alpha was not present in the fetal component of the placenta, although it was expressed in adjacent maternal derived cells. It is concluded that (1) PGDH activity increases in late gestation; (2) PGDH is expressed in uninucleate trophoblast cells in the ovine placenta and fetal membranes, and also in the maternal endometrial epithelium and stroma, myometrium and cervix; (3) oestrogen receptor alpha is not expressed in fetal cells in the placenta or fetal membranes; and (4) the increase in PGDH activity is not regulated by oestradiol administered to the fetus. PMID- 10864847 TI - Role of O6-alkylguanine-DNA alkyltransferase in the resistance of mouse spermatogenic cells to O6-alkylating agents. AB - The O(6)-alkylguanine-DNA alkyltransferase inactivator O(6)-benzylguanine was administered to BALB/c mice either alone or before exposure to 1,3-bis(2 chloroethyl)-1-nitrosourea to study the role of the DNA repair protein O(6) alkylguanine-DNA alkyltransferase in the protection of the testis against anti cancer O(6)-alkylating agents. Exposure of the mice to 1, 3-bis(2-chloroethyl)-1 nitrosourea or O(6)-benzylguanine alone did not produce any marked testicular toxicity at the times studied. Testicular O(6)-alkylguanine-DNA alkyltransferase concentrations were assayed between 0 and 240 min after O(6)-benzylguanine treatment and were shown to be > 95% depleted 15 min after treatment with O(6) benzylguanine and remained at > 95% at all the times assayed. Histological examination, the reduction in testicular mass and the induction of spermatogenic cell apoptosis showed that this depletion significantly potentiated 1, 3-bis(2 chloroethyl)-1-nitrosourea-induced testicular damage after treatment. Major histological damage was apparent 42 days after treatment, demonstrating that the stem spermatogonia were significantly affected by the combination. These results demonstrate that O(6)-alkylguanine-DNA alkyltransferase plays a significant role in protecting the spermatogenic cells from damage caused by DNA alkylation and indicate that the observed toxicity may result from damage to stem spermatogonia. PMID- 10864849 TI - Monocyte chemoattractant protein 1 in luteolysis. AB - Monocyte chemoattractant protein 1 (MCP-1) is a member of the chemokine family of cytokines which are involved in leukocyte physiology and trafficking. Interest in the role of inflammatory cells and their cytokine products in luteolysis has been increasing and there is mounting evidence demonstrating that MCP-1 is involved in luteolysis. Cell sources of MCP-1, such as endothelial cells, are abundant in late stage luteal tissue. Increased amounts of mRNA encoding MCP-1 are found after luteolysis in sheep, pigs, cows, rats and women and its up-regulation is associated with an increase in macrophages within the corpus luteum, indicating that MCP-1 may act as an inflammatory mediator during luteal regression. Luteolytic substances (prolactin in rats and prostaglandin F2alpha in ruminants) appear to be involved in increased expression of MCP-1 within the corpus luteum, although it is unclear whether this is a direct or indirect effect. Cytokines produced within the corpus luteum around luteolysis may also be involved in regulating MCP-1 expression. The field of chemokine biology is expanding rapidly and MCP-1, as well as other chemokines yet to be investigated, may prove to be an important link between the hormonal and cellular events within the corpus luteum around the time of luteolysis. PMID- 10864848 TI - Oxytocin release and its relationship to dihydro-15-keto PGF2alpha and arginine vasopressin release during parturition and to suckling in postpartum mares. AB - Pituitary blood was collected from the intercavernal sinus in five mares before and during parturition, and in nine mares immediately after parturition to investigate oxytocin patterns during parturition and early lactation, and to determine the relationship between oxytocin, prostaglandin and arginine vasopressin during parturition. In four mares in which sample collection began at least 6 h before rupture of the chorioallantois, a significant increase (P < 0.05) in PGF(2alpha) concentration was detected before a significant increase in oxytocin concentration. Cross-correlation analysis of log-transformed oxytocin and PGF(2alpha) concentrations revealed a significant correlation (P < 0.05) at a 6 min lag period, indicating that in the 2 h before delivery of the foal, an increase in prostaglandin was followed 6 min later by an increase in oxytocin. A significant effect of suckling on oxytocin release by the mare was detected in only two of nine mares, when oxytocin concentrations were evaluated 0-3 min after suckling. When foals were prevented from sucking for 1 h, by being either muzzled (n = 2) or separated from the mare (n = 2), there was no significant association between resumption of suckling and oxytocin release by the mare. The results of these studies show that: (i) oxytocin secretion from the maternal posterior pituitary gland begins before, or in association with, the onset of the second stage of labour, and that prostaglandin increases in the peripheral circulation before oxytocin release; and (ii) suckling is not significantly related to oxytocin release in mares. PMID- 10864850 TI - Evolution of mammalian pregnancy in the presence of the maternal immune system. AB - Eutherian mammals have inherited a typical vertebrate immune system, which protects the body against infectious organisms by detecting and destroying foreign biological material. However, with the evolution of longer gestation periods, this protective mechanism became a potential threat to the 'semi foreign' fetus and so eutherians have developed systems to prevent immune rejection of their developing fetuses. In many species, this is achieved by reducing placental expression of major histocompatibility complex (MHC) genes, the products of which are responsible for most transplantation rejection reactions. Unexpectedly, however, major histocompatibility complex expression is often re-established in the most invasive trophoblast cells. It is not known why transplantation antigen expression in the fetal cells most exposed to the maternal immune system is advantageous. It is possible that such expression aids the process of invasion or exerts an immunoprotective effect on the fetus. It may prove possible to identify the essential steps that all eutherian fetuses take to ensure their survival in the face of potential maternal immune attack by studying the common features of the placental immunology of different species. PMID- 10864851 TI - Functional processing of fertilin: evidence for a critical role of proteolysis in sperm maturation and activation. AB - Fertilin is a sperm surface protein with an essential role in fertilization. It is required for the migration of spermatozoa through the oviduct, for binding to the zona pellucida, and for efficient binding to the egg plasma membrane. Fertilin consists of two subunits, fertilin alpha and beta, both of which belong to the metalloprotease-disintegrin protein family (ADAMs). Fertilin alpha and beta are made as larger precursors that are processed proteolytically at different stages of sperm maturation in the testis and epididymis. Fertilin alpha is processed first, most likely by a pro-protein convertase in the secretory pathway of testicular cells. Fertilin beta is processed later, while spermatozoa are in transit through the epididymis. The processing of fertilin beta in the epididymis correlates with the acquisition of fertilization competence in spermatozoa, exposes an epitope that has a role in sperm-egg interactions, and triggers the relocalization of fertilin from the whole sperm head to the posterior head. These findings indicate that the proteolytic processing of fertilin and perhaps also other sperm proteins plays an important role in sperm maturation and activation in the epididymis. Further evaluation of the functional significance of proteolysis for sperm maturation should lead to new and exciting insights into the mechanism of sperm maturation, and may also uncover the cause of certain types of male infertility. The identification of the responsible proteases could provide novel targets for contraceptive drugs. PMID- 10864852 TI - Oestrogen in fluid transport in efferent ducts of the male reproductive tract. AB - This review focuses on the importance of oestrogen and oestrogen receptors in the male reproductive system, with a special interest in the newly discovered role of oestrogen in the regulation of fluid reabsorption in the efferent ductules of the testis. Early work on oestrogen synthesis indicated that Leydig and Sertoli cells were the only important cells in the production of this steroid in the adult testis. However, more recent work has shown that germ cells and spermatozoa also contain aromatase and produce oestrogen. The observation that germ cells synthesize oestrogen contributed to a new hypothesis that oestrogen in the lumen of the male reproductive tract targets the epithelial lining of efferent ductules and the epididymis. The location of nuclear oestrogen receptors in the male reproductive tract has also been investigated and it has been found that oestrogen receptor alpha is more abundant in the efferent ductules of the testis than in any other tissue of the male or female. In all species examined to date, oestrogen receptor alpha has been found to be abundant in the efferent ductules. The structure and function of the efferent ductules are taken into account as these tubules are responsible for the reabsorption of almost 90% of the luminal rete testis fluid. Thus, it was logical to hypothesize that oestrogen receptors play a role in the regulation of fluid reabsorption in efferent ductules. The oestrogen receptor alpha knockout mouse was used to help define this role of the receptor in males. In this animal model, the efferent ductules are altered markedly from a reabsorptive epithelium to a squamous epithelium devoid of lysosomes and endocytotic organelles. Although the separate roles for oestrogens and androgens in the regulation of fluid reabsorption are controversial and remain to be resolved, it is now established that loss of oestrogen receptor function in males interferes with the resorptive function of efferent ductules, a function that is essential for fertility. Future studies will focus on the biochemical and physiological mechanisms involved in the regulation of water and ion movement by oestrogen in the male reproductive tract. PMID- 10864853 TI - Molecular mechanisms involved in the differentiation of spermatogenic stem cells. AB - In male mammals, spermatogenesis proceeds for the reproductive lifetime of the animal. The continuation of this process depends upon a pool of spermatogenic stem cells within the testes that undergo asymmetric division to both maintain the stem cell population and give rise to progenitors that will proceed through spermatogenesis to generate mature spermatozoa. Thus, the development of functional spermatozoa may be divided into two distinct stages. The second, the process of spermatogenesis, is dependent upon the first, the successful formation of spermatogenic stem cells. Although spermatogenesis is characterized by marked cellular differentiation, the initial stages of germ line differentiation involve an avoidance of the differentiation signals acting during embryo development. The germ line is set aside early in embryo development and, while the primordial germ cells remain refractory to the differentiation signals affecting the soma, they undergo a number of phenotypic shifts before and after colonizing the genital ridge. Upon colonization of the genital ridge, the somatic tissue of the male genital ridge directs the final differentiation events that result in the formation of spermatogenic stem cells. It is this cell population that provides the basis for the maintenance of spermatogenesis in the adult. PMID- 10864854 TI - Role of activins in the male reproductive tract. AB - The search for gonadal proteins that regulate pituitary FSH led to the isolation of inhibins and activins. As members of the transforming growth factor beta (TGFbeta) superfamily of growth and differentiation factors, these proteins have been shown subsequently to affect a range of tissues and systems beyond their role in reproduction. Studies on the expression and synthesis of activins in the male reproductive tract have localized these proteins in the testis, epididymis and prostate. In general, activins regulate cell proliferation and, consequently, the expression and localization of activin subunit mRNAs and proteins within these organs must be discrete. Activin ligand bioactivity is dependent on the presence of the appropriate receptors and signalling systems, but activin ligand formation or access to receptors is regulated by the formation of inhibins or by activin-binding proteins such as follistatin. This review examines the evidence that the capacity to synthesize activins and to regulate activin bioactivity resides in the cells of the male reproductive tract. It is concluded that activins exert their effects through local (autocrine or paracrine) mechanisms, rather than through endocrine systems. The interplay between the inhibins or follistatins provides a degree of regulation of activin bioactivity before ligand signalling events. The challenge for the future is to determine whether there is any difference between the action of individual activin ligands or whether these proteins are functionally redundant, indicating that compensatory mechanisms are essential for male reproductive tract function. PMID- 10864855 TI - Effects of stress on reproduction in non-rodent mammals: the role of glucocorticoids and sex differences. AB - The means by which stress influences reproduction is not clearly understood, but may involve a number of endocrine, paracrine and neural systems. Stress impacts on the reproductive axis at the hypothalamus (to affect GnRH secretion) and the pituitary gland (to affect gonadotrophin secretion), with direct effects on the gonads being of less importance. Different stressors have different effects and there are differences in response to short- and long-term stress. Many short-term stresses fail to affect reproduction and there are reports of stimulatory effects of some 'stressors'. There are species differences in the way that specific stressors affect reproduction. Sex differences in the effects of a particular stressor have been delineated and these may relate to effects of stress at different levels of the hypothalamo-pituitary axis. The significance of stress induced secretion of cortisol varies with species. In some instances, there appears to be little impact of short-term increases in cortisol concentrations and protracted increases in plasma concentration seem to be required before any deleterious effect on reproduction is apparent. Issues of sex, sex steroid status, type of stressor and duration of stress need to be considered to improve understanding of this issue. PMID- 10864856 TI - The zona pellucida: using molecular genetics to study the mammalian egg coat. AB - An extracellular matrix that mediates critical steps in fertilization and early development surrounds all vertebrate eggs. In mice and humans, this matrix is known as the zona pellucida and comprises three glycoproteins: ZP1, ZP2 and ZP3. Homologues of these proteins isolated from other vertebrates have conserved protein motifs that may be important for establishing a common fibrillar structure. However, specific but contradictory biological roles have been assigned to individual egg coat proteins based on assays in vitro in a wide range of species. Mouse lines lacking either ZP1 or ZP3 have been established with abnormal or absent zona matrices and varying degrees of infertility to examine zona structure and function in vivo. By crossing mouse lines lacking individual zona proteins with those expressing human homologues, the structural integrity of the zona matrix can be restored. Because mouse and human spermatozoa exhibit order-specific binding to the zona pellucida, mice with 'humanized' chimaeric zonae may provide an experimental system to elucidate the molecular basis of sperm-zona interaction. PMID- 10864857 TI - Oocyte maturation and ovum quality in pigs. AB - Oocyte quality in pigs is defined as the potential of that oocyte to develop into a viable offspring. There is increasing evidence that the programming of the oocyte must be completed before leaving the ovarian follicle, including both nuclear and cytoplasmic maturation. Pig oocytes matured in vitro under basic conditions are deficient in some, as yet unidentified, cytoplasmic factors and thus developmentally incompetent. This developmental incompetence can be overcome to some extent by follicular supplementation (with follicular fluid or granulosa cells) of the culture medium, emphasizing the importance of somatic signals during oocyte maturation. Furthermore, evidence is accumulating that the status of the follicle has a critical impact on the competence of the oocyte in vitro and in vivo and this has been demonstrated by co-culture with follicles at different maturational stages or from breeds with enhanced embryo survival. It now also appears that manipulation of maternal nutrition before mating or oocyte collection can enhance embryo survival in vivo and oocyte developmental competence in vitro, presumably by altering follicular secretions and hence the environment in which the oocyte is nurtured. Identification of both the key follicular factors influencing pig oocyte quality and reliable markers of oocyte quality will undoubtedly yield major improvements in embryo survival in vivo and embryo production in vitro. PMID- 10864858 TI - Something to be done: treating HIV/AIDS. River Path Associates. AB - Largely because of disparities in access to drug treatment and care, AIDS morbidity and mortality have fallen in the developed world but continue to rise among developing countries. Achieving more equitable access to AIDS drugs is hindered by high drug prices, technical complexities related to the provision of health care, and conflict among stakeholders. Recognition that health is vital to the prospects of the emerging global society must be combined with new mechanisms to help all stakeholders work together cooperatively. Tiered drugs pricing should be coupled with investment in health services. An independent "Global Task Force," able to act as an "active think tank," could build consensus about the way forward. PMID- 10864859 TI - Scaling Up HIV/AIDS programs to national coverage. AB - The most important issue in the fight against HIV/AIDS is how to scale up existing programs that are only reaching small numbers of people to the national level. Here, I present suggestions on how to tackle the daunting challenge of building truly national HIV/AIDS programs, based on insights gained from participatory, decentralized rural development experiences and from HIV/AIDS programs. PMID- 10864860 TI - Global AIDS epidemic: time to turn the tide. AB - HIV/AIDS is catastrophic both from a public health perspective and in terms of its impact on economic and social stability in many of the most severely affected nations, including virtually all of southern Africa. A public health response alone is insufficient to address this devastating epidemic. Political leadership at the highest levels is needed to mobilize a multisectoral response to the impact of HIV/AIDS on educational systems, industry, agriculture, the military, and other sectors. With a few notable exceptions, political response was slow to mobilize in the early years of the epidemic, but response has dramatically improved in the past 18 months. The Joint United Nations Programme on HIV/AIDS (UNAIDS) is involved in ongoing efforts to encourage political leaders to make a multisectoral response to the epidemic a major focus of their national plans. PMID- 10864861 TI - A ferroelectric liquid crystal conglomerate composed of racemic molecules AB - We describe the design and synthesis of a ferroelectric liquid crystal composed of racemic molecules. The ferroelectric polarization results from spontaneous polar symmetry breaking in a fluid smectic. The ferroelectric phase is also chiral, resulting in the formation of a mixture of macroscopic domains of either handedness at the isotropic-to-liquid crystal phase transition. This smectic liquid crystal is thus a fluid conglomerate. Detailed investigation of the electrooptic and polarization current behavior within individual domains in liquid crystal cells shows the thermodynamically stable structure to be a uniformly tilted smectic bow-phase (banana phase), with all layer pairs homochiral and ferroelectric (SmC(S)P(F)). PMID- 10864862 TI - An orientational transition of bent-core molecules in an anisotropic matrix AB - We report the discovery of an orientational transition of bent-core molecules in a background anisotropic medium, in this case a smectic liquid crystal made of rod-like molecules. The resulting director is apolar in nature, and the medium can be described as an orthogonal biaxial smectic. The detailed phase diagram of mixtures of the two types of compounds revealed the induction of two liquid crystalline phases that are specific to compounds with bent-core molecules. The chemical nature of the bounding surface had a marked influence on the observed textures. PMID- 10864863 TI - High-strength welds in metallocene Polypropylene/Polyethylene laminates AB - Spectacular advances in organometallic chemistry over the past two decades have resulted in single-site catalysts that are revolutionizing production of polyethylene (PE) and isotactic polypropylene (iPP). This report describes an unanticipated benefit of metallocene-catalyzed semicrystalline polyolefins, namely welded joint strengths in PE/iPP laminates that can exceed the cohesive strength of the constituents. We propose that interfacial polymer entanglements, established in the molten state and subsequently anchored in chain-folded lamellae upon crystallization, are responsible for this intrinsic property. The poor adhesion exhibited by traditional Ziegler-Natta-catalyzed polyolefins is shown to derive from the accumulation of amorphous polymer, a by-product of the polymerization reactions, at the interface. These results should facilitate fabrication and improve the properties of composites based on materials that dominate the plastics industry. PMID- 10864864 TI - Understanding the distribution of near-earth asteroids AB - We have deduced the orbital and size distributions of the near-Earth asteroids (NEAs) by (i) numerically integrating NEAs from their source regions to their observed orbits, (ii) estimating the observational biases and size distribution associated with asteroids on those orbits, and (iii) creating a model population that can be fit to the known NEAs. We predict that there are approximately 900 NEAs with absolute magnitude less than 18 (that is, kilometer-sized), of which 29, 65, and 6% reside on Amor, Apollo, and Aten orbits, respectively. These results suggest that roughly 40% of the kilometer-sized NEAs have been found. The remainder, on highly eccentric and inclined orbits, are more difficult to detect. PMID- 10864865 TI - Isotopic dating of strain fringe increments: duration and rates of deformation in shear zones AB - The time scales over which deformation in the Earth's crust remains localized in shear zones are poorly known, as are the associated strain rates. We have determined the longevity and rates of deformation using rubidium-strontium (Rb Sr) microsampling dating of increments of fibrous strain fringes from a Pyrenean shear zone. The fibers grew quasi-continuously through a protracted deformation history between 87 and 50 million years ago, over a period comparable to that of an orogeny. During a short interval between 66 and 62 million years, a rise in strain rate from 1.1 x 10(-15) to 7. 7 x 10(-15) seconds(-1) occurred. This acceleration correlates with an abrupt change in fiber-growth direction and a stress-field inversion from gravitational collapse to renewed horizontal crustal shortening. PMID- 10864866 TI - Coherent high- and low-latitude climate variability during the holocene warm period AB - A faunal record of sea-surface temperature (SST) variations off West Africa documents a series of abrupt, millennial-scale cooling events, which punctuated the Holocene warm period. These events evidently resulted from increased southward advection of cooler temperate or subpolar waters to this subtropical location or from enhanced regional upwelling. The most recent of these events was the Little Ice Age, which occurred between 1300 to 1850 A.D., when subtropical SSTs were reduced by 3 degrees to 4 degrees C. These events were synchronous with Holocene changes in subpolar North Atlantic SSTs, documenting a strong, in-phase link between millennial-scale variations in high- and low-latitude climate during the Holocene. PMID- 10864867 TI - Nonavian feathers in a late Triassic archosaur. AB - Longisquama insignis was an unusual archosaur from the Late Triassic of central Asia. Along its dorsal axis Longisquama bore a series of paired integumentary appendages that resembled avian feathers in many details, especially in the anatomy of the basal region. The latter is sufficiently similar to the calamus of modern feathers that each probably represents the culmination of virtually identical morphogenetic processes. The exact relationship of Longisquama to birds is uncertain. Nevertheless, we interpret Longisquama's elongate integumentary appendages as nonavian feathers and suggest that they are probably homologous with avian feathers. If so, they antedate the feathers of Archaeopteryx, the first known bird from the Late Jurassic. PMID- 10864868 TI - A metalloprotease disintegrin that controls cell migration in Caenorhabditis elegans. AB - In Caenorhabditis elegans, the gonad acquires two U-shaped arms by the directed migration of its distal tip cells (DTCs) along the body wall basement membranes. Correct migration of DTCs requires the mig-17 gene, which encodes a member of the metalloprotease-disintegrin protein family. The MIG-17 protein is secreted from muscle cells of the body wall and localizes in the basement membranes of gonad. This localization is dependent on the disintegrin-like domain of MIG-17 and its catalytic activity. These results suggest that the MIG-17 metalloprotease directs migration of DTCs by remodeling the basement membrane. PMID- 10864869 TI - Movement of retinal along the visual transduction path. AB - Movement of the ligand/receptor complex in rhodopsin (Rh) has been traced. Bleaching of diazoketo rhodopsin (DK-Rh) containing 11-cis-3-diazo-4-oxo-retinal yields batho-, lumi-, meta-I-, and meta-II-Rh intermediates corresponding to those of native Rh but at lower temperatures. Photoaffinity labeling of DK-Rh and these bleaching intermediates shows that the ionone ring cross-links to tryptophan-265 on helix F in DK-Rh and batho-Rh, and to alanine-169 on helix D in lumi-, meta-I-, and meta-II-Rh intermediates. It is likely that these movements involving a flip-over of the chromophoric ring trigger changes in cytoplasmic membrane loops resulting in heterotrimeric guanine nucleotide-binding protein (G protein) activation. PMID- 10864870 TI - Bacterial mode of replication with eukaryotic-like machinery in a hyperthermophilic archaeon. AB - Despite a rapid increase in the amount of available archaeal sequence information, little is known about the duplication of genetic material in the third domain of life. We identified a single origin of bidirectional replication in Pyrococcus abyssi by means of in silico analyses of cumulative oligomer skew and the identification of an early replicating chromosomal segment. The replication origin in three Pyrococcus species was found to be highly conserved, and several eukaryotic-like DNA replication genes were clustered around it. As in Bacteria, the chromosomal region containing the replication terminus was a hot spot of genome shuffling. Thus, although bacterial and archaeal replication proteins differ profoundly, they are used to replicate chromosomes in a similar manner in both prokaryotic domains. PMID- 10864871 TI - Requirement of Mis6 centromere connector for localizing a CENP-A-like protein in fission yeast. AB - Mammalian kinetochores contain the centromere-specific histone H3 variant CENP-A, whose incorporation into limited chromosomal regions may be important for centromere function and chromosome segregation during mitosis. However, regulation of CENP-A localization and its role have not been clear. Here we report that the fission yeast homolog SpCENP-A is essential for establishing centromere chromatin associated with equal chromosome segregation. SpCENP-A binding to the nonrepetitious inner centromeres depended on Mis6, an essential centromere connector protein acting during G1-S phase of the cell cycle. Mis6 is likely required for recruiting SpCENP-A to form proper connection of sister centromeres. PMID- 10864872 TI - Role of the guanosine triphosphatase Rac2 in T helper 1 cell differentiation. AB - T helper 1 (TH1) cells mediate cellular immunity, whereas TH2 cells potentiate antiparasite and humoral immunity. We used a complementary DNA subtraction method, representational display analysis, to show that the small guanosine triphosphatase Rac2 is expressed selectively in murine TH1 cells. Rac induces the interferon-gamma (IFN-gamma) promoter through cooperative activation of the nuclear factor kappa B and p38 mitogen-activated protein kinase pathways. Tetracycline-regulated transgenic mice expressing constitutively active Rac2 in T cells exhibited enhanced IFN-gamma production. Dominant-negative Rac inhibited IFN-gamma production in murine T cells. Moreover, T cells from Rac2-/- mice showed decreased IFN-gamma production under TH1 conditions in vitro. Thus, Rac2 activates TH1-specific signaling and IFN-gamma gene expression. PMID- 10864873 TI - A primitive T cell-independent mechanism of intestinal mucosal IgA responses to commensal bacteria. AB - The immunoglobulin A (IgA) is produced to defend mucosal surfaces from environmental organisms, but host defenses against the very heavy load of intestinal commensal microorganisms are poorly understood. The IgA against intestinal commensal bacterial antigens was analyzed; it was not simply "natural antibody" but was specifically induced and responded to antigenic changes within an established gut flora. In contrast to IgA responses against exotoxins, a significant proportion of this specific anti-commensal IgA induction was through a pathway that was independent of T cell help and of follicular lymphoid tissue organization, which may reflect an evolutionarily primitive form of specific immune defense. PMID- 10864874 TI - Impaired cued and contextual memory in NPAS2-deficient mice. AB - Neuronal PAS domain protein 2 (NPAS2) is a basic helix-loop-helix (bHLH) PAS domain transcription factor expressed in multiple regions of the vertebrate brain. Targeted insertion of a beta-galactosidase reporter gene (lacZ) resulted in the production of an NPAS2-lacZ fusion protein and an altered form of NPAS2 lacking the bHLH domain. The neuroanatomical expression pattern of NPAS2-lacZ was temporally and spatially coincident with formation of the mature frontal association/limbic forebrain pathway. NPAS2-deficient mice were subjected to a series of behavioral tests and were found to exhibit deficits in the long-term memory arm of the cued and contextual fear task. Thus, NPAS2 may serve a dedicated regulatory role in the acquisition of specific types of memory. PMID- 10864875 TI - Citrulline does not relax isolated rat and rabbit vessels. AB - The study was prompted by the report of Ruiz E. & Tejerina T., 1998 describing endothelium-independent relaxation by L-citrulline via activation of particulate guanylate cyclase. We compared the effects of L-citrulline and L-arginine in isolated aortic rings of rats and in isolated aortic, carotid and femoral artery rings of rabbits. No significant relaxation to either L-citrulline or L-arginine was found in the concentration range of 10(-12) to 10(-3) M, while 3 morpholinosydnonimine hydrochloride (SIN-1, 10(-6) M) relaxed vascular tissues. This study does not support the conclusion that L-citrulline has direct vasorelaxing action on vascular smooth muscle. PMID- 10864876 TI - Accelerated resequestration of cytosolic calcium and suppression of the pro inflammatory activities of human neutrophils by CGS 21680 in vitro. AB - We have investigated the effects of the adenosine A(2A) receptor agonist CGS 21680 (0.01 - 1 microM) on reactive oxidant production by, and elastase release from FMLP-activated human neutrophils, as well as on cytosolic Ca(2+) fluxes and intracellular concentrations of cyclic AMP. Oxidant production, elastase release and cyclic AMP were assayed using lucigenin-enhanced chemiluminescence, colourimetric and radioimmunoassay procedures respectively, while cytosolic Ca(2+) fluxes were measured by fura-2 spectrofluorimetry in combination with radiometric procedures which distinguish between net efflux and influx of the cation. Treatment of neutrophils with CGS 21680 did not affect the FMLP-activated release of Ca(2+) from intracellular stores, but resulted in dose-related acceleration of the rate of decline in fura-2 fluorescence, as well as decreases in both efflux and store-operated influx of Ca(2+), compatible with enhancement of resequestration of the cation by the endo-membrane Ca(2+)-ATPase. These effects on neutrophil Ca(2+) handling were associated with increased intracellular cyclic AMP and with inhibition of oxidant production and release of elastase. In contrast, treatment of neutrophils with the selective A(2A) receptor antagonist, ZM 241385 (2.5 microM), prevented the transient increase in cyclic AMP in FMLP-activated neutrophils which was associated with delayed sequestration of incoming Ca(2+) during store-operated influx. The CGS 21680-mediated reduction of Ca(2+) efflux from FMLP-activated neutrophils was also antagonized by pretreatment of the cells with ZM 241385 (2.5 microM), as well as by thapsigargin (1 microM), an inhibitor of the endo-membrane Ca(2+)-ATPase. ZM 241385 also neutralized the cyclic AMP-elevating and anti-inflammatory interactions of CGS 21680 with neutrophils. We conclude that A(2A) receptors regulate the pro inflammatory activities of human neutrophils by promoting cyclic AMP-dependent sequestration of cytosolic Ca(2+). PMID- 10864877 TI - Anaesthetic agents inhibit gastrin-stimulated but not basal histamine release from rat stomach ECL cells. AB - By mobilizing histamine in response to gastrin, the ECL cells in the oxyntic mucosa play a key role in the control of the parietal cells and hence of gastric acid secretion. General anaesthesia suppresses basal and gastrin- and histamine stimulated acid secretion. The present study examines if the effect of anaesthesia on basal and gastrin-stimulated acid secretion is associated with suppressed ECL-cell histamine secretion. A microdialysis probe was implanted in the submucosa of the ventral aspect of the acid-producing part of the stomach (32 rats). Three days later, ECL-cell histamine mobilization was monitored 2 h before and 4 h after the start of intravenous infusion of gastrin (5 nmol kg(-1) h(-1)). The rats were either conscious or anaesthetized. Four commonly used anaesthetic agents were given 1 h before the start of the experiments by intraperitoneal injection: chloral hydrate (300 mg kg(-1)), pentobarbitone (40 mg kg(-1)), urethane (1.5 g kg(-1)) and a mixture of fluanisone/fentanyl/midazolam (15/0.5/7.5 mg kg(-1)). In a parallel series of experiments, basal- and gastrin induced acid secretion was monitored in six conscious and 25 anaesthetized (see above) chronic gastric fistula rats. All anaesthetic agents lowered gastrin stimulated acid secretion; also the basal acid output was reduced (fluanisone/fentanyl/midazolam was an exception). Anaesthesia reduced gastrin stimulated but not basal histamine release by 55 - 80%. The reduction in gastrin induced acid response (70 - 95%) was strongly correlated to the reduction in gastrin-induced histamine mobilization. The correlation is in line with the view that the reduced acid response to gastrin reflects impaired histamine mobilization. Rat stomach ECL cells were purified by counter-flow elutriation. Gastrin-evoked histamine mobilization from the isolated ECL cells was determined in the absence or presence of anaesthetic agents in the medium. With the exception of urethane, they inhibited gastrin-evoked histamine secretion dose dependently, indicating a direct effect on the ECL cells. Anaesthetized rats are widely used to study acid secretion and ECL-cell histamine release. The present results illustrate the short-comings of such an approach in that a number of anaesthetic agents were found to impair not only acid secretion but also the secretion of ECL-cell histamine - some acting in a direct manner. PMID- 10864878 TI - Insulin modulation of intracellular free magnesium in heart: involvement of protein kinase C. AB - In the present study of rat heart using (31)P-nuclear magnetic resonance, we examined the interaction between beta-adrenergic and insulin receptors in terms of the intracellular free Mg(2+) concentration ([Mg(2+)](i)) regulation. [Mg(2+)](i) was estimated from the separation of the chemical shifts of the alpha and beta-adenosine triphosphate (ATP) peaks, using the dissociation constant of MgATP 87 microM (established recently). In normal (phosphate-free Krebs Henseleit) solution, [Mg(2+)](i) was approximately 1.02 mM. Insulin at physiological and pathological concentrations increased [Mg(2+)](i) and contractility in a dose-dependent manner. Insulin (more than 100 micro(u) ml(-1)) suppressed the decrease in [Mg(2+)](i) caused by isoprenaline (100 nM), and these effects of insulin on [Mg(2+)](i) and contractility were blocked by LY333531 (macrocyclic bis (indolyl) maleimide, 100 nM), a protein kinase C (PKC) inhibitor. The isoprenaline-induced decrease in the concentrations of ATP ([ATP]) with insulin application was significantly smaller than that without insulin. Insulin modulates [Mg(2+)](i) and haemodynamics, presumably via activation of PKC, thereby antagonizing the reduction of [Mg(2+)](i) induced by beta adrenoceptor stimulation. PMID- 10864879 TI - Flibanserin has anxiolytic effects without locomotor side effects in the infant rat ultrasonic vocalization model of anxiety. AB - This study compared the effects of flibanserin, a novel 5-HT(1A) agonist/5-HT(2A) antagonist; diazepam, a traditional anxiolytic; and imipramine, a traditional antidepressant, on separation-induced ultrasonic vocalizations (USVs), locomotor behaviour, negative geotaxis and body temperature of 7 - 8-day-old rat pups. Flibanserin (5, 10, 25 and 50 mg kg(-1) s.c.) reduced USVs but had no effects on locomotor behaviour or negative geotaxis. Lower doses of flibanserin (0.5, 1, 2 and 4 mg kg(-1) s.c.) had no effect on any behaviour. Diazepam (0.25, 0.5, 1 and 2 mg kg(-1) s.c.) not only reduced the USVs but also increased rolling and increased the latency of the negative geotaxic response. Imipramine (10, 15, 20 and 30 mg kg(-1) s.c.) reduced USVs, increased total locomotor activity and rolling but had no effect on negative geotaxis. None of the drugs altered body temperature. Our data showed that flibanserin is as effective in reducing the USVs as diazepam and imipramine but has a lower incidence of motor side effects. This suggests that flibanserin might be effective for the treatment of mood disturbances such as anxiety. PMID- 10864880 TI - Contribution of beta-adrenoceptor subtypes to relaxation of colon and oesophagus and pacemaker activity of ureter in wildtype and beta(3)-adrenoceptor knockout mice. AB - The smooth muscle relaxant responses to the mixed beta(3)-, putative beta(4) adrenoceptor agonist, (-)-CGP 12177 in rat colon are partially resistant to blockade by the beta(3)-adrenoceptor antagonist SR59230A suggesting involvement of beta(3)- and putative beta(4)-adrenoceptors. We now investigated the function of the putative beta(4)-adrenoceptor and other beta-adrenoceptor subtypes in the colon, oesophagus and ureter of wildtype (WT) and beta(3)-adrenoceptor knockout (beta(3)KO) mice. (-)-Noradrenaline and (-)-adrenaline relaxed KCl (30 mM) precontracted colon mostly through beta(1)-and beta(3)-adrenoceptors to a similar extent and to a minor extent through beta(2)-adrenoceptors. In colon from beta(3)KO mice, (-)-noradrenaline was as potent as in WT mice but the effects were mediated entirely through beta(1)-adrenoceptors. (-)-CGP 12177 relaxed colon from beta(3)KO mice with 2 fold greater potency than in WT mice. The maintenance of potency for (-)-noradrenaline and increase for (-)-CGP 12177 indicate compensatory increases in beta(1)- and putative beta(4)-adrenoceptor function in beta(3)KO mice. In oesophagi precontracted with 1 microM carbachol, (-) noradrenaline caused relaxation mainly through beta(1)-and beta(3)-adrenoceptors. (-)-CGP 12177 (2 microM) relaxed oesophagi from WT by 61.4+/-5.1% and beta(3)KO by 67.3+/-10.1% of the (-)-isoprenaline-evoked relaxation, consistent with mediation through putative beta(4)-adrenoceptors. In ureter, (-)-CGP 12177 (2 microM) reduced pacemaker activity by 31.1+/-2.3% in WT and 31.3+/-7. 5% in beta(3)KO, consistent with mediation through putative beta(4)-adrenoceptors. Relaxation of mouse colon and oesophagus by catecholamines are mediated through beta(1)- and beta(3)-adrenoceptors in WT. The putative beta(4)-adrenoceptor, which presumably is an atypical state of the beta(1)-adrenoceptor, mediates the effects of (-)-CGP 12177 in colon, oesophagus and ureter. PMID- 10864881 TI - Betaxolol, a beta(1)-adrenoceptor antagonist, reduces Na(+) influx into cortical synaptosomes by direct interaction with Na(+) channels: comparison with other beta-adrenoceptor antagonists. AB - Betaxolol, a beta(1)-adrenoceptor antagonist used for the treatment of glaucoma, is known to be neuroprotective in paradigms of ischaemia/excitotoxicity. In this study, we examined whether betaxolol and other beta-adrenoceptor antagonists interact directly with neurotoxin binding to sites 1 and 2 of the voltage sensitive sodium channel (Na(+) channel) in rat cerebrocortical synaptosomes. Betaxolol inhibited specific [(3)H]-batrachotoxinin-A 20-alpha-benzoate ([(3)H] BTX-B) binding to neurotoxin site 2 in a concentration-dependent manner with an IC(50) value of 9.8 microM. Comparison of all the beta-adrenoceptor antagonists tested revealed a potency order of propranolol>betaxolol approximately levobetaxolol>levobunolol approximately carteolol>/=timolol>atenolol. None of the drugs caused a significant inhibition of [(3)H]-saxitoxin binding to neurotoxin receptor site 1, even at concentrations as high as 250 microM. Saturation experiments showed that betaxolol increased the K(D) of [(3)H]-BTX-B binding but had no effect on the B(max). The association kinetics of [(3)H]-BTX-B were unaffected by betaxolol, but the drug significantly accelerated the dissociation rate of the radioligand. These findings argue for a competitive, indirect, allosteric mode of inhibition of [(3)H]-BTX-B binding by betaxolol. Betaxolol inhibited veratridine-stimulated Na(+) influx in rat cortical synaptosomes with an IC(50) value of 28. 3 microM. Carteolol, levobunolol, timolol and atenolol were significantly less effective than betaxolol at reducing veratridine-evoked Na(+) influx. The ability of betaxolol to interact with neurotoxin site 2 of the Na(+) channel and inhibit Na(+) influx may have a role in its neuroprotective action in paradigms of excitotoxicity/ischaemia and in its therapeutic effect in glaucoma. PMID- 10864882 TI - JTV-519, a novel cardioprotective agent, improves the contractile recovery after ischaemia-reperfusion in coronary perfused guinea-pig ventricular muscles. AB - A newly synthesized benzothiazepine derivative, JTV-519 (JT) has been reported to be cardioprotective. However, the precise mechanism underlying the cardioprotective effect of this drug is unknown. Coronary-perfused guinea-pig ventricular muscles were subjected to 20-min no-flow ischaemia followed by 60-min reperfusion (I/R). I/R significantly decreased the contraction in untreated preparations (control group, 34+/-4% of baseline value, n=6). Brief administration of JT (1.0 microM) prior to ischaemia significantly improved the postischaemic contractile recovery (63+/-5% of baseline value, n=4), as compared to the control group. JT (1.0 microM) slightly prolonged action potential duration before ischaemia and induced conduction disturbance (2 : 1 block) after the initiation of ischaemia. The cardioprotective effect of JT was antagonized by chelerythrine (CH, 5.0 microM), an inhibitor of protein kinase C (PKC) or by 5 hydroxydecanoic acid (5-HD, 400 microM), an inhibitor of mitochondrial ATP sensitive K(+) (K(ATP)) channels. These results suggest that the protective effect of JT is due to the opening of mitochondrial K(ATP) channels, which, in turn, is linked to PKC activation. PMID- 10864883 TI - Pyrimidinoceptor potentiation of macrophage PGE(2) release involved in the induction of nitric oxide synthase. AB - We have previously demonstrated that Ca(2+)/calmodulin-dependent protein kinase (CaMK) mediates pyrimidinoceptor potentiation of LPS-elicited inducible nitric oxide synthase (iNOS) induction in murine J774 macrophages. In the present paper, we have explored the role of cyclo-oxygenase (COX)-dependent prostaglandin E(2) (PGE(2)) formation in this event. In J774 macrophages predominantly expressing P2Y(6) receptors, the simultaneous addition of UTP and lipopolysaccharide (LPS) resulted in potentiated increase in PGE(2) release. UTP-induced increased PGE(2) release was demonstrated by a concomitant increase in COX-2 protein expression, and was decreased by inhibitors specific for phosphatidylinositide-phospholipase C (PI-PLC), CaMK, protein kinase C (PKC), nuclear factor-kappa B (NF-kappaB) or COX-2. NS-398 (a selective COX-2 inhibitor) reduced LPS plus UTP-elicited iNOS induction and nitrite accumulation, supporting for the positive regulation of iNOS gene expression by endogenous PGE(2). Moreover, the cyclic AMP/PKA-dependent up-regulation of iNOS expression mediated by PGE(2) was drawn from the inhibitory effects of 2',5'-dideoxyadenosine, KT5720 and H-89. Exogenous PGE(2) induced NF kappaB activation and potentiated nitrite accumulation in response to LPS. In addition to COX-2 induction, arachidonic acid (AA) release and steady-state mRNA levels of type V secretory phospholipase A(2) (sPLA(2)) and Ca(2+)-independent PLA(2) (iPLA(2)) were also increased in the presence of LPS and UTP; the LPS induced increase in iPLA(2) activity was also potentiated by UTP. Taken together, we conclude that UTP-mediated COX-2 and iPLA(2) potentiation and PGE(2) formation contribute to the iNOS induction, and that CaMK activation is the primary step in the UTP enhancement of COX-2 induction. PMID- 10864884 TI - Electrophysiological effects of endothelin-1 and their relationship to contraction in rat renal arterial smooth muscle. AB - The electophysiological effects of endothelin-1 (ET-1) and their relationship to contraction remain unclear in the renal circulation. Using endotheliumdenuded arteries from the main branch of the renal artery proximal to the kidney of the rat, we have examined its effects on tension and conducted parallel patch-clamp measurements using freshly isolated smooth muscle cells from this tissue. Pharmacological experiments revealed that ET-1 produced constriction of renal arteries dependent on the influx of extracellular Ca(2+), mediated solely through ET(A) receptor stimulation. Current-clamp experiments revealed that renal arterial myocytes had a resting membrane potential of approximately 32 mV, with the majority of cells exhibiting spontaneous transient hyperpolarizations (STHPs). Application of ET-1 produced depolarization and in those cells exhibiting STHPs, either caused their inhibition or made them occur regularly. Under voltage-clamp conditions cells were observed to exhibit spontaneous transient outward currents (STOCs) inhibited by iberiotoxin. Application of voltage-ramps revealed an outward current activated at approximately -30 mV, sensitive to both 4-AP and TEA. Taken together these results suggest that renal arterial myocytes possess both delayed rectifying K(+) (K(V)) and Ca(2+) activated K(+) (BK(Ca)) channels. Under voltage-clamp, ET-1 attenuated the outward current and reduced the magnitude and incidence of STOCs: effects mediated solely as a consequence of ET(A) receptor stimulation. Thus, in conclusion, activation of ET(A) receptors by ET-1 causes inhibition of K(V) and BK(Ca) channel activity, which could promote and/or maintain membrane depolarization. This effect is likely to favour L-type Ca(2+) channel activity providing an influx pathway for extracellular Ca(2+) essential for contraction. PMID- 10864885 TI - Effects of chronic fluoxetine treatment in the presence and absence of (+/ )pindolol: a microdialysis study. AB - Using in vivo microdialysis in the frontal cortex of the freely moving rat we evaluated the effects of chronic treatment with the serotonin specific reuptake inhibitor (SSRI) fluoxetine in the presence and absence of the 5-HT(1A)/beta adrenergic antagonist (+/-)pindolol. Chronic vehicle treated animals produced no significant response to a challenge with fluoxetine (10 mg kg(-1)) on day 8 and 15. Alternatively, a significant (P<0.05) decrease in extracellular 5-HT was observed in control animals upon challenge with the 5-HT(1A) agonist 8-hydroxy-2 (di-n-propylamino)tetralin (8-OH-DPAT; 0.03 and 0.1 mg kg(-1)). Conversely, animals treated with fluoxetine (10 mg kg(-1) o.d.) for 7 and 14 days produced a significant (P<0.05) 2 fold increase in extracellular 5-HT when challenged with fluoxetine (10 mg kg(-1)) on day 8 and 15. Moreover, no significant decrease in extracellular 5-HT was observed upon challenge with either dose of 8-OH-DPAT. Animals chronically treated with (+/-)pindolol (10 or 20 mg kg(-1) b.i.d.) produced a significant dose-related increase in extracellular 5-HT upon challenge with fluoxetine on day 15 only. Furthermore, both doses produced a significantly blunted response to the low dose challenge of 8-OH-DPAT (0.03 mg kg(-1)). In addition, 20 mg kg(-1) (+/-)pindolol treated animals also had no response to the higher 0.1 mg kg(-1) dose of 8-OH-DPAT. Animals treated for 14 days with a combination of (+/-)pindolol (10 or 20 mg kg(-1)) and fluoxetine were not significantly different from vehicle treated animals when challenged with fluoxetine or 8-OH-DPAT. Taken together it would therefore appear that although (+/-)pindolol alone has sufficient intrinsic activity to produce a desensitization of the 5-HT(1A) receptor, when given in combination with fluoxetine it is able to prevent the desensitization induced by not only fluoxetine but also itself. This may suggest that the clinical augmentation of antidepressant action by pindolol, when co-administered with a SSRI, is via antagonism of the 5-HT(1A) receptor. PMID- 10864886 TI - Regional haemodynamic effects of recombinant murine or human leptin in conscious rats. AB - Regional haemodynamic responses to recombinant murine or human leptin were assessed in conscious, chronically-instrumented, male, Long-Evans rats (350 - 450 g). Human, but not murine, leptin caused a slight hindquarters vasoconstriction, but neither peptide had any effect on mean arterial blood pressure or heart rate. In the presence of the beta(2)-adrenoceptor antagonist, ICI 118551, a hindquarters vasoconstrictor response to human leptin was not seen, and there was a tachycardia, as there was to murine leptin. The nitric oxide synthase inhibitor, N(G)-nitro-L-arginine methyl ester, (L-NAME), did not influence the cardiovascular effects of murine or human leptin. The results indicate that the previously reported sympathoexcitatory effects of murine leptin in anaesthetized rats are not manifest as regional haemodynamic changes in conscious rats, and this is not due to beta(2)-adrenoceptor-mediated vasodilator mechanisms opposing any vasoconstrictor responses. Moreover, the ability of L-NAME to unmask a pressor effect of murine leptin in anaesthetized rats may not be apparent in the conscious state. PMID- 10864887 TI - Opposite effects of flurbiprofen and the nitroxybutyl ester of flurbiprofen on apoptosis in cultured guinea-pig gastric mucous cells. AB - The nitric oxide (NO)-donating nitroxybutyl ester of flurbiprofen (NO flurbiprofen), shows reduced gastro-intestinal toxicity relative to flurbiprofen. NO may exert either pro- or anti-apoptotic effects, while non-steroidal anti inflammatory drugs may induce apoptosis. The aim of the present work was therefore to compare the effects of flurbiprofen and NO-flurbiprofen on apoptosis in guinea-pig gastric mucous cells. Apoptotic activity was assessed by assay of caspase activity and from the fragmentation and condensation of nuclei. Incubation with flurbiprofen for 24 h produced a concentration-dependent induction of apoptosis in cells attached to the culture plate (caspase 3-like activity increased by 257% at 500 microM), while NO-flurbiprofen inhibited basal apoptosis (caspase 3-like activity decreased by 71% at 500 microM). Caspase activity and nuclear fragmentation were substantially increased in cells that had spontaneously detached from the culture plate. NO-flurbiprofen inhibited caspase activity (55% at 500 microM) but not nuclear fragmentation in these detached cells. NO flurbiprofen inhibited the activation of apoptosis by 25 microM C(6) ceramide in cells attached to the culture plate. Inhibition of caspase activity by NO-flurbiprofen was detectable after 6 h of incubation with intact cells, but by contrast with the NO-donor S-nitrosyl-N-acetyl-penicillamine, was not demonstrable with cell homogenates. Activation of caspase 3-like activity by flurbiprofen was slow (>6 h incubation needed) and was inhibited by cycloheximide. The presence of a nitroxybutyl ester moiety on flurbiprofen prevents the pro-apoptotic activity of the parent compound and may contribute to the reduced gastro-intestinal toxicity of NO-flurbiprofen. PMID- 10864888 TI - Characterization of a novel VPAC(1) selective agonist and identification of the receptor domains implicated in the carboxyl-terminal peptide recognition. AB - Vasoactive Intestinal Polypeptide (VIP) interacts with a high affinity to two subclasses of G protein coupled receptors named VPAC(1) and VPAC(2), and has a 3 10 fold preference for VPAC(1) over VPAC(2) receptors. Selective ligands for each receptor subclass were recently described. [R(16)]-PACAP (1 - 23) and [L(22)]-VIP are two selective VPAC(1) agonists. Chimaeric human VPAC(2)-VPAC(1) recombinant receptors expressed in CHO cells were used to identify the receptor domains implicated in these two selective ligands recognition. The VPAC(2) preference for [R(16)]-PACAP (1 - 27) over [R(16)]-PACAP (1 - 23) did not require the receptor's NH(2)-terminus domain but involved the whole transmembrane domain. In contrast, the selectivity of [L(22)]-VIP depended only on the presence of the NH(2) terminus and EC(2) domains of the VPAC(1) receptor. The present data support the idea that in the GPCR-B family of receptors the different selective ligands require different domains for their selectivity, and that the peptides carboxyl terminal sequence (amino acids 24 - 27) folds back on the transmembrane receptor domain, close to the peptides, aminoterminus. PMID- 10864889 TI - Taurine blocks ATP-sensitive potassium channels of rat skeletal muscle fibres interfering with the sulphonylurea receptor. AB - Taurine is a sulphonic aminoacid present in high amounts in various tissues including cardiac and skeletal muscles showing different properties such as antioxidative, antimyotonic and anti-schaemic effects. The cellular mechanism of action of taurine is under investigation and appears to involve the interaction of the sulphonic aminoacid with several ion channels. Using the patch-clamp technique we studied the effects of taurine in rat skeletal muscle fibres on ATP sensitive K(+) channel (K(ATP)) immediately after excision and on channels that underwent rundown. The cytoplasmic application of 20 mM of taurine reduced the K(ATP) current; this effect was reverted by washout of the drug solution. In this experimental condition the IC(50) was 20.1 mM. After rundown, taurine inhibited the K(ATP) current with similar efficacy. Competition experiments showed that taurine shifted the dose-response inhibition curve of glybenclamide to the left on the log-dose axis without significantly affecting those of ATP or Ca(2+) ion. Single channel recording revealed that taurine affects the close state of the channel prolonging it and reducing the bursts duration. Our data indicate that taurine inhibits the muscular K(ATP) channel interfering with the glybenclamide site on the sulphonylurea receptor of the channel or on the site allosterically coupled to it. During ischaemia and hypoxia, the skeletal and heart muscles undergo several changes; for example, the activation of K(ATP) channels and loss of the intracellular taurine content. The depletion of taurine during ischaemia would contribute to the early activation of K(ATP) channels and salvage the intracellular ATP content. PMID- 10864891 TI - Effects of 5-aminoisoquinolinone, a water-soluble, potent inhibitor of the activity of poly (ADP-ribose) polymerase on the organ injury and dysfunction caused by haemorrhagic shock. AB - Poly (ADP-ribose) synthetase (PARP) is a nuclear enzyme activated by strand breaks in DNA, which are caused inter alia by reactive oxygen species (ROS). Here we report on (i) a new synthesis of a water-soluble and potent PARP inhibitor, 5 aminoisoquinolinone (5-AIQ) and (ii) investigate the effects of 5-AIQ on the circulatory failure and the organ injury/dysfunction caused by haemorrhage and resuscitation in the anaesthetized rat. Exposure of human cardiac myoblasts (Girardi cells) to hydrogen peroxide (H(2)O(2), 3 mM for 1 h, n=9) caused a substantial increase in PARP activity. Pre-treatment of these cells with 5-AIQ (1 microM - 1 mM, 10 min prior to H(2)O(2)) caused a concentration-dependent inhibition of PARP activity (IC(50): approximately 0.01 mM, n=6). Haemorrhage and resuscitation resulted (within 4 h after resuscitation) in a delayed fall in blood pressure (circulatory failure) as well as in rises in the serum levels of (i) urea and creatinine (renal dysfunction), (ii) aspartate aminotransferase (AST), alanine aminotransferase (ALT), and gamma-glutamyl-transferase (gamma-GT) (liver injury and dysfunction), (iii) lipase (pancreatic injury) and (iv) creatine kinase (CK) (neuromuscular injury) (n=10). Administration (5 min prior to resuscitation of 5-AIQ) (0.03 mg kg(-1) i.v., n=8, or 0.3 mg kg(-1) i.v., n=10) reduced (in a dose-related fashion) the multiple organ injury and dysfunction, but did not affect the circulatory failure, associated with haemorrhagic shock. Thus, 5-AIQ abolishes the multiple organ injury caused by severe haemorrhage and resuscitation. PMID- 10864890 TI - The involvement of intracellular Ca(2+) in 5-HT(1B/1D) receptor-mediated contraction of the rabbit isolated renal artery. AB - 5-Hydroxytryptamine(1B/1D) (5-HT(1B/1D)) receptor coupling to contraction was investigated in endothelium-denuded rabbit isolated renal arteries, by simultaneously measuring tension and intracellular [Ca(2+)], and tension in permeabilized smooth muscle cells. In intact arterial segments, 1 nM - 10 microM 5-HT failed to induce contraction or increase the fura-2 fluorescence ratio (in the presence of 1 microM ketanserin and prazosin to block 5-HT(2) and alpha(1) adrenergic receptors, respectively). However, in vessels pre-exposed to either 20 mM K(+) or 30 nM U46619, 5-HT stimulated concentration-dependent increases in both tension and intracellular [Ca(2+)]. 1 nM - 10 microM U46619 induced concentration-dependent contractions. In the presence of nifedipine (0.3 and 1 microM) the maximal contraction to U46619 (10 microM) was reduced by around 70%. The residual contraction was abolished by the putative receptor operated channel inhibitor, SKF 96365 (2 microM). With 0.3 microM nifedipine present, 100 nM U46619 evoked similar contraction to 30 nM U46619 in the absence of nifedipine, but contraction to 5-HT (1 nM - 10 microM) was abolished. In permeabilized arterial segments, 10 mM caffeine, 1 microM IP(3) or 100 microM phenylephrine, each evoked transient contractions by releasing Ca(2+) from intracellular stores, whereas 5-HT had no effect. In intact arterial segments pre-stimulated with 20 mM K(+), 5-HT-evoked contractions were unaffected by 1 microM thapsigargin, which inhibits sarco- and endoplasmic reticulum calcium-ATPases. In vessels permeabilized with alpha-toxin and then pre-contracted with Ca(2+) and GTP, 5-HT evoked further contraction, reflecting increased myofilament Ca(2+)-sensitivity. Contraction linked to 5-HT(1B/1D) receptor stimulation in the rabbit renal artery can be explained by an influx of external Ca(2+) through voltage-dependent Ca(2+) channels and sensitization of the contractile myofilaments to existing levels of Ca(2+), with no release of Ca(2+) from intracellular stores. PMID- 10864892 TI - Interaction between copper and zinc at GABA(A) receptors in acutely isolated cerebellar Purkinje cells of the rat. AB - Nanomolar concentrations of Cu(2+) induce a slowly reversible block of GABA(A) receptor-mediated currents which can be removed by chelating substances. The possible interaction of Cu(2+) with the Zn(2+) binding site on the GABA(A) receptor complex was studied in acutely isolated Purkinje cells using whole-cell recording and a fast drug application system. When Zn(2+) was applied together with 2 microM GABA, the Zn(2+)-induced block of GABA-mediated currents was not additive to the Cu(2+)-induced block. In the presence of 0.1 microM Cu(2+) in the bath solution the degree of inhibition of GABA-mediated responses by Zn(2+) was strongly attenuated. Preapplication of 100 microM Zn(2+) during 10 s, terminated 1 s before exposure to 2 microM GABA did not affect the GABA current in Cu(2+) free solution, but relieved its block by 0.1 microM Cu(2+). This effect of Zn(2+) was concentration-dependent with an EC(50) of 72 microM. When the Cu(2+)-induced block was removed by histidine, preapplication of Zn(2+) did not increase the GABA current, indicating that the relief of Cu(2+) block by Zn(2+) is the result of its ability to actively remove Cu(2+) from the GABA receptor complex. It is proposed that the inhibitory effects of Zn(2+) and Cu(2+) on GABA-induced currents result from an action of these metal ions at distinct, but conformationally linked sites on the GABA(A) receptor protein. Under physiological conditions Zn(2+) would liberate Cu(2+) from the GABA(A) receptor, thus facilitating Cu(2+) turnover and its binding by other endogenous chelating molecules. PMID- 10864893 TI - Sensitivity of Kir6.2-SUR1 currents, in the absence and presence of sodium azide, to the K(ATP) channel inhibitors, ciclazindol and englitazone. AB - Two electrode voltage clamp and single channel recordings were used to investigate the actions of various ATP-sensitive K(+) (K(ATP)) channel inhibitors on cloned K(ATP) channels, expressed in Xenopus oocytes and HEK 293 cells. Oocytes expressing Kir6.2 and SUR1 gave rise to inwardly rectifying K(+) currents following bath application of 3 mM sodium azide. Inside-out recordings from non azide treated oocytes demonstrated the presence of K(ATP) channels which were activated by direct application of 3 mM azide and 0.1 mM Mg-ATP. Tolbutamide inhibited azide-induced macroscopic Kir6.2-SUR1 currents, recorded from Xenopus oocytes, with an IC(50) value similar to native K(ATP) channels. Ciclazindol and englitazone also inhibited these currents in a concentration-dependent manner, but with relative potencies substantially less than for native K(ATP) channels. Single channel currents recorded from inside-out patches excised from oocytes expressing Kir6.2-SUR1 currents were inhibited by tolbutamide, Mg-ATP, englitazone and ciclazindol, in the absence of azide, with potencies similar to native K(ATP) channels. In the presence of azide, Kir6.2-SUR1 currents were inhibited by englitazone and tolbutamide but not ciclazindol. Single channel currents derived from Kir6.2Delta26, expressed in HEK 293 cells, were inhibited by ciclazindol and englitazone irrespective of the absence or presence of SUR1. In conclusion, heterologously expressed Kir6.2 and SUR1 recapitulate the pharmacological profile of native pancreatic beta-cell K(ATP) channels. However, currents induced by azide exhibit a substantially reduced sensitivity to ciclazindol. It is likely that ciclazindol and englitazone inhibit K(ATP) currents by interaction with the Kir6.2 subunit. PMID- 10864894 TI - Activation of protein kinase B by the A(1)-adenosine receptor in DDT(1)MF-2 cells. AB - In this study the effect of insulin and A(1)-adenosine receptor stimulation on protein kinase B (PKB) activation has been investigated in the hamster vas deferens smooth muscle cell line DDT(1)MF-2. Increases in PKB phosphorylation were determined by Western blotting using an antibody that detects PKB phosphorylation at Ser(473). Insulin, a recognized activator of PKB, stimulated a concentration-dependent increase in PKB phosphorylation in DDT(1)MF-2 cells (EC(50) 5+/-1 pM). The selective A(1)-adenosine receptor agonist N(6) cyclopentyladenosine (CPA) stimulated time and concentration-dependent increases in PKB phosphorylation in DDT(1)MF-2 cells (EC(50) 1.3+/-0.5 nM). CPA-mediated increases in PKB phosphorylation were antagonized by the A(1)-adenosine receptor selective antagonist 1,3-dipropylcyclopentylxanthine (DPCPX) yielding an apparent K(D) value of 2.3 nM. Pre-treatment of DDT(1)MF-2 cells with pertussis toxin (PTX, 100 ng ml(-1) for 16 h), to block G(i)/G(o)-dependent pathways, abolished CPA (1 microM) induced phosphorylation of PKB. In contrast, responses to insulin (100 nM) were resistant to PTX pre-treatment. The phosphatidylinositol 3-kinase (PI-3K) inhibitors wortmannin (IC(50) 10.3+/-0.6 nM) and LY 294002 (IC(50) 10.3+/ 1.2 microM) attenuated the phosphorylation of PKB elicited by CPA (1 microM) in a concentration-dependent manner. Wortmannin (30 nM) and LY 294002 (30 microM) also blocked responses to insulin (100 nM). Removal of extracellular Ca(2+) and chelation of intracellular Ca(2+) with BAPTA had no significant effect on CPA induced PKB phosphorylation. Similarly, pretreatment (30 min) with inhibitors of protein kinase C (Ro 31-8220; 10 microM), tyrosine kinase (genistein; 100 microM), mitogen-activated protein (MAP) kinase kinase (PD 98059; 50 microM) and p38 MAPK (SB 203580; 20 microM) had no significant effect on CPA-induced PKB phosphorylation. In conclusion, these data demonstrate that A(1)-adenosine receptor stimulation in DDT(1)MF-2 cells increases PKB phosphorylation through a PTX and PI-3K-sensitive pathway. PMID- 10864895 TI - Multiple action sites of flufenamate on ion transport across the rat distal colon. AB - The antisecretory effects of flufenamate in the rat distal colon were investigated with the Ussing-chamber and the patch-clamp method as well as by measurements of the intracellular Ca(2+) concentration using fura-2-loaded isolated crypts. Flufenamate (5.10(-4) mol l(-1)) suppressed the short-circuit current (Isc) induced by carbachol (5.10(-5) mol l(-1)), forskolin (5.10(-6) mol l(-1)) and the Isc induced by the membrane-permeable analogue of cyclic AMP, CPT cyclic AMP (10(-4) mol l(-1)). Indomethacin (10(-6) - 10(-4) mol l(-1)) did not mimic the effect of flufenamate, indicating that the antisecretory effect of flufenamate is not related to the inhibition of the cyclo-oxygenase. When the basolateral membrane was depolarized by a high K(+) concentration and a Cl(-) current was induced by a mucosally directed Cl(-) gradient, the forskolin stimulated Cl(-) current was blocked by flufenamate, indicating an inhibition of the cyclic AMP-stimulated apical Cl(-) conductance. When the apical membrane was permeabilized by the ionophore, nystatin, flufenamate decreased the basolateral K(+) conductance and inhibited the Na(+) - K(+)-ATPase. Patch-clamp experiments revealed a variable effect of flufenamate on membrane currents. In seven out of 11 crypt cells the drug induced an increase of the K(+) current, whereas in the remaining four cells an inhibition was observed. Experiments with fura-2-loaded isolated crypts indicated that flufenamate increased the basal as well as the carbachol-stimulated intracellular Ca(2+) concentration. These results demonstrate that flufenamate possesses multiple action sites in the rat colon: The apical Cl(-) conductance, basolateral K(+) conductances and the Na(+) - K(+) ATPase. PMID- 10864896 TI - Adenosine receptor expression and function in rat striatal cholinergic interneurons. AB - Cholinergic neurons were identified in rat striatal slices by their size, membrane properties, sensitivity to the NK(1) receptor agonist (Sar(9), Met(O(2))(11)) Substance P, and expression of choline acetyltransferase mRNA. A(1) receptor mRNA was detected in 60% of the neurons analysed, and A(2A) receptor mRNA in 67% (n=15). The A(1) receptor agonist R-N(6)-(2 phenylisopropyl)adenosine (R-PIA) hyperpolarized cholinergic neurons in a concentration dependent manner sensitive to the A(1) antagonist 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX, 100 nM). In dual stimulus experiments, the A(2A) receptor antagonist 8-(3-chlorostyryl)caffeine (CSC, 500 nM) decreased release of [(3)H]-acetylcholine from striatal slices (S2/S1 0.78+/-0.07 versus 0.95+/-0.05 in control), as did adenosine deaminase (S2/S1 ratio 0.69+/-0.05), whereas the A(1) receptor antagonist DPCPX (100 nM) had no effect (S2/S1 1.05+/-0.14). In the presence of adenosine deaminase the adenosine A(2A) receptor agonist 2-p ((carboxyethyl)phenylethylamino)-5'-N-ethylcarboxamidoadeno sin e (CGS21680, 10 nM) increased release (S2/S1 ratio 1.03+/-0.05 versus 0.88+/-0.05 in control), an effect blocked by the antagonist CSC (500 nM, S2/S1 0.68+/-0.05, versus 0.73+/ 0.08 with CSC alone). The combined superfusion of bicuculline (10 microM), saclofen (1 microM) and naloxone (10 microM) had no effect on the stimulation by CGS21680 (S2/S1 ratio 0.99+/-0.04). The A(1) receptor agonist R-PIA (100 nM) inhibited the release of [(3)H]-acetylcholine (S2/S1 ratio 0.70+/-0.03), an effect blocked by DPCPX (S2/S1 ratio 1.06+/-0.07). It is concluded that both A(1) and A(2A) receptors are expressed on striatal cholinergic neurons where they are functionally active. PMID- 10864897 TI - Tumour necrosis factor-alpha- and interleukin-1beta-stimulated cell proliferation through activation of mitogen-activated protein kinase in canine tracheal smooth muscle cells. AB - The elevated levels of inflammatory cytokines such as tumour necrosis factor alpha (TNF-alpha) and interleukin-1beta (IL-1beta) have been found in the fluid of airways in symptomatic asthmatics. These cytokines have been considered as mitogens to stimulate cell proliferation in tracheal smooth muscle cells (TSMCs). We therefore investigated the effects of TNF-alpha and IL-1beta on cell proliferation and activation of p42/p44 mitogen-activated protein kinase (MAPK) in these cells. TNF-alpha and IL-1beta induced [(3)H]-thymidine incorporation in a time- and concentration-dependent manner. The maximal stimulation of [(3)H] thymidine incorporation induced by TNF-alpha and IL-1beta was seen 12 h after incubation with these cytokines. In response to TNF-alpha and IL-1beta, p42/p44 MAPK was activated with a concentration-dependent manner in TSMCs. Pretreatment of TSMCs with pertussis toxin did not change DNA synthesis and phosphorylation of MAPK induced by TNF-alpha and IL-1beta. These responses were attenuated by a tyrosine kinase inhibitor herbimycin, a phosphatidyl choline (PC)-phospholipase C (PLC) inhibitor D609, a phosphatidyl inositide (PI)-PLC inhibitor U73122, a protein kinase C inhibitor staurosporine, and removal of Ca(2+) by addition of BAPTA/AM plus EGTA. TNF-alpha- and IL-1beta-induced [(3)H]-thymidine incorporation and phosphorylation of p42/p44 MAPK was completely inhibited by PD98059 (an inhibitor of MEK1/2), indicating that activation of MEK1/2 was required for these responses. These results suggest that the mitogenic effects of TNF-alpha and IL-1beta were mediated through the activation of MEK1/2 and p42/p44 MAPK pathway. TNF-alpha- and IL-1beta-mediated responses were modulated by PLC, Ca(2+), PKC, and tyrosine kinase associated with cell proliferation in TSMCs. PMID- 10864898 TI - Inhibition of neuronal Ca(2+) influx by gabapentin and subsequent reduction of neurotransmitter release from rat neocortical slices. AB - Cytosolic calcium ion concentrations ([Ca(2+)](i)) were measured in rat neocortical synaptosomes using fura-2, and depolarization of synaptosomal membranes was induced by K(+) (30 mM). The release of the endogenous excitatory amino acids glutamate and aspartate was evoked by K(+) (50 mM) and determined by HPLC. The release of [(3)H]-noradrenaline from rat neocortical synaptosomes or slices was evoked by K(+) (15 and 25 mM) and measured by liquid scintillation counting. Gabapentin produced a concentration-dependent inhibition of the K(+) induced [Ca(2+)](i) increase in synaptosomes (IC(50)=14 microM; maximal inhibition by 36%). The inhibitory effect of gabapentin was abolished in the presence of the P/Q-type Ca(2+) channel blocker omega-agatoxin IVA, but not by the N-type Ca(2+) channel antagonist omega-conotoxin GVIA. Gabapentin (100 microM) decreased the K(+)-evoked release of endogenous aspartate and glutamate in neocortical slices by 16 and 18%, respectively. Gabapentin reduced the K(+) evoked [(3)H]-noradrenaline release in neocortical slices (IC(50)=48 microM; maximal inhibition of 46%) but not from synaptosomes. In the presence of the AMPA receptor antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and 2, 3-dioxo-6 nitro-1,2,3,4-tetrahydro[f]quinoxaline-7-sulphonamide (NBQX), gabapentin did not reduce [(3)H]-noradrenaline release. Gabapentin did, however, cause inhibition in the presence of the NMDA receptor antagonist DL-(E)-2-amino-4-methyl-5-phosphono 3-pentanoic acid (CGP 37849). Gabapentin is concluded to reduce the depolarization-induced [Ca(2+)](i) increase in excitatory amino acid nerve terminals by inhibiting P/Q-type Ca(2+) channels; this decreased Ca(2+) influx subsequently attenuates K(+)-evoked excitatory amino acid release. The latter effect leads to a reduced activation of AMPA receptors which contribute to K(+) evoked noradrenaline release from noradrenergic varicosities, resulting in an indirect inhibition of noradrenaline release. PMID- 10864899 TI - Involvement of tyrosine phosphorylation in the positive inotropic effect produced by H(1)-receptors with histamine in guinea-pig left atrium. AB - We investigated the effect of stimulation of H(1)-receptors with histamine on protein tyrosine phosphorylation levels in guinea-pig left atrium and evaluated the influences of tyrosine kinase inhibitors on the positive inotropic effect mediated by H(1)-receptors in this tissue. Histamine induced an increase in tyrosine phosphorylation in four main clusters of proteins with apparent molecular weights of 25, 35, 65 and 150 kDa. Tyrosine phosphorylation of these proteins attained a peak around 2 - 3 min following histamine stimulation and then declined to or below basal levels. Histamine-induced protein tyrosine phosphorylation was antagonized by the H(1)-receptor antagonists mepyramine (1 microM) and chlorpheniramine (1 microM), but not by the H(2)-receptor antagonist cimetidine (10 microM). The positive inotropic effect of histamine was depressed in a concentration-dependent manner by the tyrosine kinase inhibitors tyrphostin A25 (50 to 100 microM) and genistein (10 to 50 microM) but not by the inactive genistein analogue daidzein (50 microM). The positive inotropic effect of isoprenaline was unchanged by tyrphostin A25 and genistein. At a concentration of 1 microM histamine produced a dual-component positive inotropic response composed of an initial increasing phase and a second and late developing, greater positive inotropic phase. Treatment with tyrphostin A25 (100 microM) and genistein (50 microM), but not daidzein (50 microM), significantly attenuated the two components of the inotropic response, although genistein suppressed the initial component more markedly than the late component. We conclude that increased protein tyrosine phosphorylation may play an important role in initiating at least some part of the positive inotropic effect of H(1)-receptor stimulation in guinea-pig left atrium. PMID- 10864900 TI - Characterization using FLIPR of rat vanilloid receptor (rVR1) pharmacology. AB - The vanilloid receptor (VR1) is a ligand-gated ion channel, which plays an important role in nociceptive processing. Therefore, a pharmacological characterization of the recently cloned rat VR1 (rVR1) was undertaken. HEK293 cells stable expressing rVR1 (rVR1-HEK293) were loaded with Fluo-3AM and then incubated at 25 degrees C for 30 min with or without various antagonists or signal transduction modifying agents. Then intracellular calcium concentrations ([Ca(2+)](i)) were monitored using FLIPR, before and after the addition of various agonists. The rank order of potency of agonists (resiniferatoxin (RTX)>capsaicin>olvanil>PPAHV) was as expected, and all were full agonists. The potencies of capsaicin and olvanil, but not RTX or PPAHV, were enhanced at pH 6.4 (pEC(50) values of 7.47+/-0.06, 7.16+/-0.06, 8.19+/-0.06 and 6.02+/-0.03 respectively at pH 7.4 vs 7.71+/-0.05, 7.58+/-0.14, 8.10+/-0.05 and 6.04+/-0.08 at pH 6.4). Capsazepine, isovelleral and ruthenium red all inhibited the capsaicin (100 nM)-induced Ca(2+) response in rVR1-HEK293 cells, with pK(B) values of 7.52+/-0.08, 6.92+/-0.11 and 8.09+/-0.12 respectively (n=6 each). The response to RTX and olvanil were also inhibited by these compounds. None displayed any agonist-like activity. The removal of extracellular Ca(2+) abolished, whilst inhibition of protein kinase C with chelerythrine chloride (10 microM) partially (approximately 20%) inhibited, the capsaicin (10 microM) induced Ca(2+) response. However, tetrodotoxin (3 microM), nimodipine (10 microM), omega-GVIA conotoxin (1 microM), thapsigargin (1 microM), U73122 (3 microM) or H-89 (3 microM) had no effect on the capsaicin (100 nM)-induced response. In conclusion, the recombinant rVR1 stably expressed in HEK293 cells acts as a ligand-gated Ca(2+) channel with the appropriate agonist and antagonist pharmacology, and therefore is a suitable model for studying the effects of drugs at this receptor. PMID- 10864901 TI - Specific inhibition of stretch-induced increase in L-type calcium channel currents by herbimycin A in canine basilar arterial myocytes. AB - The effects of protein-tyrosine kinase (PTK) and protein-tyrosine phosphatase (PTP) inhibitors on voltage-activated barium currents (I(Ba)) through L-type calcium channels increased by hypotonic solution were investigated in canine basilar arterial myocytes by the whole-cell patch-clamp technique. I(Ba) was elicited by depolarizing step from a holding potential of -80 to +10 mV and identified by using an L-type calcium channel agonist, Bay K 8644 (100 nM), and an L-type calcium channel blocker, nicardipine (1 microM). Hypotonic superfusate induced cell swelling and acted as a stretch stimulus, which reversibly increased peak I(Ba) amplitude at +10 mV. I(Ba) was also decreased by nicardipine (1 microM) under the hypotonic condition. PTK inhibitors such as herbimycin A (30 nM), genistein (10 microM), and lavendustin A (10 microM) decreased I(Ba) enhanced by hypotonic solution. Genistein also decreased I(Ba) in a concentration dependent manner under the isotonic condition. The inactive genistein analogue daidzein (10 microM) had no effect on I(Ba) under either the isotonic or hypotonic condition. By contrast, herbimycin A did not decrease I(Ba) under the isotonic condition. Sodium orthovanadate (10 microM), a PTP inhibitor, increased I(Ba) under both conditions. The present results suggest that cell swelling by hypotonic solution increases the L-type calcium channel currents in canine basilar artery and that herbimycin-sensitive PTK activity is primarily involved in the enhancement of calcium channel currents. PMID- 10864902 TI - Apocynin inhibits peroxynitrite formation by murine macrophages. AB - Peroxynitrite (ONOO(-)) the highly reactive coupling product of nitric oxide and superoxide, has been implicated in the pathogenesis of an increasing number of (inflammatory) diseases. At present, however, selective peroxynitrite antagonizing agents with therapeutic potential are not available. Therefore, the NADPH-oxidase inhibitor apocynin (4-hydroxy-3-methoxy-acetophenone) was tested for its ability to inhibit peroxynitrite formation in vitro The murine macrophage cell-line J774A.1, stimulated with IFNgamma/LPS, was used as a model. Conversion of 123-dihydrorhodamine (123-DHR) to its oxidation product 123-rhodamine was used to measure peroxynitrite production. Stimulated peroxynitrite formation could be completely inhibited by apocynin, by the superoxide scavenger TEMPO as well as by the nitric oxide synthase inhibitor aminoguanidine. Apocynin and aminoguanidine specifically inhibited superoxide and nitric oxide formation respectively as confirmed by measuring lucigenin enhanced chemiluminescence and nitrite accumulation. It is concluded that J774A.1 macrophages produce significant amounts of peroxynitrite, which is associated with nitric oxide production and NADPH-oxidase dependent superoxide formation. The NADPH-oxidase inhibitor apocynin proved to be a potent inhibitor of both superoxide and peroxynitrite formation by macrophages, which may be of future therapeutic significance in a wide range of inflammatory disorders. PMID- 10864903 TI - Manganese increases L-DOPA auto-oxidation in the striatum of the freely moving rat: potential implications to L-DOPA long-term therapy of Parkinson's disease. AB - We have previously shown that manganese enhances L-dihydroxyphenylanine (L-DOPA) toxicity to PC12 cells in vitro. The supposed mechanism of manganese enhancing effect [an increase in L-DOPA and dopamine (DA) auto-oxidation] was studied using microdialysis in the striatum of freely moving rats. Systemic L-DOPA [25 mg kg( 1) intraperitoneally (i.p.) twice in a 12 h interval] significantly increased baseline dialysate concentrations of L-DOPA, dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and uric acid, compared to controls. Conversely, DA and ascorbic acid concentrations were significantly decreased. A L-DOPA oxidation product, presumptively identified as L-DOPA semiquinone, was detected in the dialysate. The L-DOPA semiquinone was detected also following intrastriatal infusion of L-DOPA. In rats given L-DOPA i.p. , intrastriatal infusion of N acetylcysteine (NAC) significantly increased DA and L-DOPA dialysate concentrations and lowered those of L-DOPA semiquinone; in addition, NAC decreased DOPAC+HVA and uric acid dialysate concentrations. In rats given L-DOPA either systemically or intrastriatally, intrastriatal infusion of manganese decreased L-DOPA dialysate concentrations and greatly increased those of L-DOPA semiquinone. These changes were inhibited by NAC infusion. These findings demonstrate that auto-oxidation of exogenous L-DOPA occurs in vivo in the rat striatum. The consequent reactive oxygen species generation may account for the decrease in dialysate DA and ascorbic acid concentrations and increase in enzymatic oxidation of xanthine and DA. L-DOPA auto-oxidation is inhibited by NAC and enhanced by manganese. These results may be of relevance to the L-DOPA long term therapy of Parkinson's disease. PMID- 10864904 TI - Carbon monoxide in vasoregulation: the promise and the challenge. PMID- 10864905 TI - Cytokine actions of angiotensin II. PMID- 10864906 TI - Lymphocyte trafficking mediated by vascular adhesion protein-1: implications for immune targeting and cardiovascular disease. PMID- 10864907 TI - Gap junctions in cardiovascular disease. AB - Connexins, the protein molecules forming gap junction channels, are reduced in number or redistributed from intercalated disks to lateral cell borders in a variety of cardiac diseases. This "gap junction remodeling" is considered to be arrhythmogenic. Using a simple model of human ventricular myocardium, we found that quantitative remodeling data extracted from the literature gave rise to only small to moderate changes in conduction velocity and the anisotropy ratio. Especially for longitudinal conduction, cytoplasmic resistivity (and thus cellular geometry) is much more important than commonly realized. None of the remodeling data gave rise to slow conduction on the order of a few centimeters per second. PMID- 10864908 TI - Vascular endothelial growth factor(165) gene transfer augments circulating endothelial progenitor cells in human subjects. AB - Preclinical studies in animal models and early results of clinical trials in patients suggest that intramuscular injection of naked plasmid DNA encoding vascular endothelial growth factor (VEGF) can promote neovascularization of ischemic tissues. Such neovascularization has been attributed exclusively to sprout formation of endothelial cells derived from preexisting vessels. We investigated the hypothesis that VEGF gene transfer may also augment the population of circulating endothelial progenitor cells (EPCs). In patients with critical limb ischemia receiving VEGF gene transfer, gene expression was documented by a transient increase in plasma levels of VEGF. A culture assay documented a significant increase in EPCs (219%, P<0.001), whereas patients who received an empty vector had no change in circulating EPCs, as was the case for volunteers who received saline injections (VEGF versus empty vector, P<0.001; VEGF versus saline, P<0.005). Fluorescence-activated cell sorter analysis disclosed an overall increase of up to 30-fold in endothelial lineage markers KDR (VEGF receptor-2), VE-cadherin, CD34, alpha(v)beta(3), and E-selectin after VEGF gene transfer. Constitutive overexpression of VEGF in patients with limb ischemia augments the population of circulating EPCs. These findings support the notion that neovascularization of human ischemic tissues after angiogenic growth factor therapy is not limited to angiogenesis but involves circulating endothelial precursors that may home to ischemic foci and differentiate in situ through a process of vasculogenesis. PMID- 10864909 TI - Requisite role for interleukin-4 in the acceleration of fatty streaks induced by heat shock protein 65 or Mycobacterium tuberculosis. AB - Atherosclerotic lesions can be induced in rabbits and mice immunized with heat shock protein 65 (HSP65). In the current study, we investigated the role of interleukin (IL)-4 in the HSP65- and Mycobacterium tuberculosis (MT)-induced models that exhibit an inflammatory phenotype. Fatty streak formation in IL-4 knockout (IL-4 KO) mice immunized with HSP65 or MT was significantly reduced when compared with lesions in wild-type C57BL/6 mice. However, when injected with control (HSP-free) adjuvant, no differences were evident in the lesion size between wild-type and the IL-4 KO mice. Next, we studied comparatively the extent of humoral and cellular immune responses to HSP65 in the IL-4 KO and wild-type mice, as those are thought to be influential in murine atherosclerosis. Anti HSP65 antibody levels were reduced in the HSP65-immunized IL-4 KO mice as compared with their wild-type littermates, whereas no differences were evident between the groups with respect to the primary cellular immune response to HSP65. Other than the absence of IL-4 in the knockout mice, the pattern of secreting cytokines interferon-gamma and IL-10 in concanavalin A-primed splenocytes was similar between the groups. HSP65-primed inguinal lymphocytes from IL-4 KO mice immunized with HSP65 secreted higher levels of interferon-gamma (previously shown to be proatherogenic in vivo) as compared with their wild-type controls. 12-/15 Lipoxygenase expression, known to be regulated by IL-4 and to contribute to murine atherosclerosis, in the lesions was not influenced by the immunization protocol used or by IL-4 disruption. Thus, IL-4 may prove a principal cytokine in the progression of early "inflammatory" atherosclerotic lesions and may serve as a target for immunomodulation. PMID- 10864910 TI - 2-deoxy-ATP enhances contractility of rat cardiac muscle. AB - To investigate the kinetic parameters of the crossbridge cycle that regulate force and shortening in cardiac muscle, we compared the mechanical properties of cardiac trabeculae with either ATP or 2-deoxy-ATP (dATP) as the substrate for contraction. Comparisons were made in trabeculae from untreated rats (predominantly V1 myosin) and those treated with propylthiouracil (PTU; V3 myosin). Steady-state hydrolytic activity of cardiac heavy meromyosin (HMM) showed that PTU treatment resulted in >40% reduction of ATPase activity. dATPase activity was >50% elevated above ATPase activity in HMM from both untreated and PTU-treated rats. V(max) of actin-activated hydrolytic activity was also >50% greater with dATP, whereas the K(m) for dATP was similar to that for ATP. This indicates that dATP increased the rate of crossbridge cycling in cardiac muscle. Increases in hydrolytic activity were paralleled by increases of 30% to 80% in isometric force (F(max)), rate of tension redevelopment (k(tr)), and unloaded shortening velocity (V(u)) in trabeculae from both untreated and PTU-treated rats (at maximal Ca(2+) activation), and F-actin sliding speed in an in vitro motility assay (V(f)). These results contrast with the effect of dATP in rabbit psoas and soleus fibers, where F(max) is unchanged even though k(tr), V(u), and V(f) are increased. The substantial enhancement of mechanical performance with dATP in cardiac muscle suggests that it may be a better substrate for contractility than ATP and warrants exploration of ribonucleotide reductase as a target for therapy in heart failure. PMID- 10864911 TI - Transgenic overexpression of constitutively active protein kinase C epsilon causes concentric cardiac hypertrophy. AB - To test the hypothesis that activation of the protein kinase C (PKC) epsilon isoform leads to cardiac hypertrophy without failure, we studied transgenic mice with cardiac-specific overexpression of a constitutively active mutant of the PKCepsilon isoform driven by an alpha-myosin heavy chain promoter. In transgenic mice, the protein level of PKCepsilon in heart tissue was increased 9-fold. There was a 6-fold increase of the membrane/cytosol ratio, and PKC activity in the membrane fraction was 4.2-fold compared with wild-type mice. The heart weight was increased by 28%, and upregulation of the mRNA for beta-myosin heavy chain and alpha-skeletal actin was observed in transgenic mouse hearts. Echocardiography demonstrated increased anterior and posterior wall thickness with normal left ventricular function and dimensions, indicating concentric cardiac hypertrophy. Isolated cardiomyocyte mechanical function was slightly decreased, and Ca(2+) signals were markedly depressed in transgenic mice, suggesting that myofilament sensitivity to Ca(2+) was increased. No differences were observed in either the levels of cardiac Ca(2+)-handling proteins or the degree of cardiac regulatory protein phosphorylation between wild-type and transgenic mice. Unlike mice with PKCbeta(2) overexpression, transgenic mice with cardiac-specific overexpression of the active PKCepsilon mutant demonstrated concentric hypertrophy with normal in vivo cardiac function. Thus, PKC isoforms may play differential functional roles in cardiac hypertrophy and failure. PMID- 10864912 TI - Prevention of hypoxia-induced pulmonary hypertension by enhancement of endogenous heme oxygenase-1 in the rat. AB - We investigated the role of heme oxygenase (HO)-1 in the development of hypoxia induced pulmonary hypertension. HO catalyzes the breakdown of heme to the antioxidant bilirubin and the vasodilator carbon monoxide. Hypoxia is a potent but transient inducer of HO-1 in vascular smooth muscle cells in vitro and in the lung in vivo. By using agonists of HO-1, we sustained a high expression of HO-1 in the lungs of rats for 1 week. We report that this in vivo enhancement of HO-1 in the lung prevented the development of hypoxic pulmonary hypertension and inhibited the structural remodeling of the pulmonary vessels. The mechanism(s) underlying this effect may involve a direct vasodilating and antiproliferative action of endogenous carbon monoxide, as well as an indirect effect of carbon monoxide on the production of vasoconstrictors. These results provide evidence that enhancement of endogenous adaptive responses may be used to prevent hypoxia induced pulmonary hypertension. PMID- 10864913 TI - Opposite effects of pressurized steady versus pulsatile perfusion on vascular endothelial cell cytosolic pH: role of tyrosine kinase and mitogen-activated protein kinase signaling. AB - Endothelial cytosolic pH (pH(i)) modulates ion channel function, vascular tone, and cell proliferation. Steady shear induces rapid acidification in bicarbonate buffer. However, in vivo shear is typically pulsatile, potentially altering this response. We tested effects and mechanisms of pH(i) modulation by flow pulsatility, comparing pressurized steady versus pulse-flow responses in bovine aortic endothelial cells cultured within glass capillary tubes. Cells were loaded with the fluorescent pH(i) indicator carboxy seminaphthorhodafluor-1 and perfused with physiological pulsatile pressure and flow generated by a custom servo control system. Raising mean pressure from 0 to 90 mm Hg at 0.5 mL/min steady flow in bicarbonate buffer induced sustained acidification (-0.33+/-0.09 pH units, P<0.01). A subsequent increase in steady flow resulted in further acidification. In contrast, if mean pressure and flow were unchanged but perfusion made pulsatile, pH(i) rose +0.3+/-0.03 (P<0. 0001) over 30 to 60 minutes. HCO(3)(-) removal and use of acid/base exchange inhibitors 5-(N-ethyl-N isopropyl)amiloride or diisothiocyanato stilbene disulfonic acid identified both extracellular Na(+)-independent Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers as activated by static pressure, whereas pulsatility activated extracellular Na(+) dependent Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers to raise pH(i). Pulse perfusion alkalinization occurred with or without flow reversal and increased 1.6 fold in Ca(2+)-free buffer. Inhibition of c-Src tyrosine kinase (4-amino-5-[4 chlorophenyl]-7-[t-butyl]pyrazolo [3,4-d]pyrimidine; PP2) or MEK-1 (mitogen activated protein kinase [MAP]/extracellular signal-regulated kinase [ERK]-1) (PD98059, blocking ERK1/2) blocked or reversed the pulsatile-flow pH(i) change to acidification. In contrast, PP2 had no effect on steady flow acidification, whereas MEK-1 inhibition converted it to alkalinization. Thus, pulsatile and steady flow trigger opposite effects on endothelial pH(i) by differential activation of acid/base exchangers linked to c-Src and MAP kinase phosphorylation, but not to Ca(2+). These data highlight specific signaling responses triggered by phasic shear profiles. PMID- 10864914 TI - Postanoxic T lymphocyte-endothelial cell interactions induce tumor necrosis factor-alpha production and neutrophil adhesion: role of very late antigen 4/vascular cell adhesion molecule-1. AB - The objective of this study was to define the influence of postanoxic T lymphocyte-endothelial cell interactions on anoxia-reoxygenation (A/R)-induced neutrophil-endothelial cell adhesion and cell adhesion molecule (CAM) expression on human umbilical vein endothelial cells (HUVECs). HUVEC monolayers were exposed to 60 minutes of anoxia, followed by 24 hours of reoxygenation, wherein freshly isolated human T lymphocytes were added at 6 hours during reoxygenation. After an additional 18 hours of incubation (ie, total of 24 hours of reoxygenation), the T cell/endothelial cell (TC/EC) coculture media were collected and added to naive HUVEC monolayers incubated with neutrophils. Although the A/R-conditioned media per se had no effect on neutrophil adhesion, the media from TC/EC cocultures significantly increased the adhesion response. This enhanced adhesive interaction was associated with significant increases in tumor necrosis factor-alpha (TNF alpha) and interleukin-8 (IL-8) levels in the TC/EC coculture media and was accompanied by a pronounced increase in endothelial E-selectin expression. Treatment of the TC/EC coculture media with anti-TNF-alpha or anti-IL-8 antibodies reduced the media-induced neutrophil adhesion response. The enhanced neutrophil adhesion and the elevated medium levels of TNF-alpha, but not IL-8, were markedly reduced by inserts that prevented direct TC/EC contact and by monoclonal antibodies directed against vascular cell adhesion molecule-1 (VCAM-1) or very late antigen-4 (VLA-4). Collectively, these findings show that VLA-4 /VCAM-1-mediated interactions between T lymphocytes and postanoxic endothelial cells stimulates TNF-alpha production, which in turn elicits endothelial cell adhesion molecule expression and a corresponding increase in neutrophil adhesion. PMID- 10864915 TI - Human vascular adhesion protein-1 (VAP-1) plays a critical role in lymphocyte endothelial cell adhesion cascade under shear. AB - Lymphocyte binding to vascular endothelium is a prerequisite for the movement of immune cells from the blood into lymphoid tissues and into sites of inflammation. Human vascular adhesion protein-1 (VAP-1) is an endothelial glycoprotein involved in this interaction. It also displays an enzymatic (monoamine oxidase) activity. Here we examined how recombinant human VAP-1 mediates lymphocyte binding using rotatory and flow chamber binding assays. VAP-1 cDNA transfected into an endothelial cell line, which does not bind lymphocytes, renders the cell line capable of binding lymphocytes in a shear-dependent manner. VAP-1 transfectants bound lymphocytes 5 times better than monocytes with a preference for T killer cells, and no specific granulocyte adherence was detectable. The binding is partially inhibited by anti-VAP-1 monoclonal antibodies or by blocking lymphocyte L-selectin and CD18 integrins, but not by inhibition of several other homing associated molecules. In contrast, CD44 ligation on lymphocytes markedly upregulates their VAP-1-dependent adhesion, suggesting that the VAP-1 counterreceptor can be activated via CD44. The transfectant model also allowed us to perform detailed structure-function analyses of VAP-1. We show that the exposed integrin-binding motif RGD or the enzymatic activity is not indispensable for VAP-1-dependent adhesion. Together, these data show that VAP-1 can reconstitute the lymphocyte-endothelial adhesion cascade under shear and propose a critical role for VAP-1 in lymphocyte emigration from the blood. PMID- 10864916 TI - Regulation of protein kinase B and 4E-BP1 by oxidative stress in cardiac myocytes. AB - Stimulation of phosphatidylinositol 3'-kinase (PI3K) and protein kinase B (PKB) is implicated in the regulation of protein synthesis in various cells. One mechanism involves PI3K/PKB-dependent phosphorylation of 4E-BP1, which dissociates from eIF4E, allowing initiation of translation from the 7-methylGTP cap of mRNAs. We examined the effects of insulin and H(2)O(2) on this pathway in neonatal cardiac myocytes. Cardiac myocyte protein synthesis was increased by insulin, but was inhibited by H(2)O(2). PI3K inhibitors attenuated basal levels of protein synthesis and inhibited the insulin-induced increase in protein synthesis. Insulin or H(2)O(2) increased the phosphorylation (activation) of PKB through PI3K, but, whereas insulin induced a sustained response, the response to H(2)O(2) was transient. 4E-BP1 was phosphorylated in unstimulated cells, and 4E BP1 phosphorylation was increased by insulin. H(2)O(2) stimulated dephosphorylation of 4E-BP1 by increasing protein phosphatase (PP1/PP2A) activity. This increased the association of 4E-BP1 with eIF4E, consistent with H(2)O(2) inhibition of protein synthesis. The effects of H(2)O(2) were sufficient to override the stimulation of protein synthesis and 4E-BP1 phosphorylation induced by insulin. These results indicate that PI3K and PKB are important regulators of protein synthesis in cardiac myocytes, but other factors, including phosphatase activity, modulate the overall response. PMID- 10864917 TI - Interleukin-1beta and tumor necrosis factor-alpha decrease collagen synthesis and increase matrix metalloproteinase activity in cardiac fibroblasts in vitro. AB - We tested the hypothesis that the inflammatory cytokines can regulate fibroblast extracellular matrix metabolism. Neonatal and adult rat cardiac fibroblasts cultures in vitro were exposed to interleukin (IL)-1beta (4 ng/mL), tumor necrosis factor-alpha (TNF-alpha; 100 ng/mL), IL-6 (10 ng/mL), or interferon gamma (IFN-gamma; 500 U/mL) for 24 hours. IL-1beta, and to a lesser extent TNF alpha, decreased collagen synthesis, which was measured as collagenase-sensitive [(3)H]proline incorporation, but had no effect on cell number or total protein synthesis. IL-1beta decreased the expression of procollagen alpha(1)(I), alpha(2)(I), and alpha1(III) mRNA, but increased the expression of procollagen alpha(1)(IV), alpha(2)(IV), and fibronectin mRNA, indicating a selective transcriptional downregulation of fibrillar collagen synthesis. IL-1beta and TNF alpha each increased total matrix metalloproteinase (MMP) activity as measured by in-gel zymography, causing specific increases in the bands corresponding to MMP 13, MMP-2, and MMP-9. IL-1beta increased the expression of proMMP-2 and proMMP-3 mRNA, suggesting that increased metalloproteinase activity is due, at least in part, to increased transcription. The effects of IL-1beta were not dependent on NO production. Thus, IL-1beta and TNF-alpha decrease collagen synthesis and activate MMPs that degrade collagen. These observations suggest that IL-1beta and TNF-alpha may contribute to ventricular dilation and myocardial failure by promoting the remodeling of interstitial collagen. PMID- 10864918 TI - Angiotensin II activates nuclear transcription factor kappaB through AT(1) and AT(2) in vascular smooth muscle cells: molecular mechanisms. AB - Nuclear factor-kappaB (NF-kappaB) regulates many genes involved in vascular physiopathology. We have previously observed in vivo NF-kappaB activation in injured vessels that diminished by angiotensin-converting enzyme inhibition. In the present work, we investigated the effect of angiotensin II (Ang II) on NF kappaB activity in rat vascular smooth muscle cells, evaluating the molecular mechanisms and the specific receptor subtype involved. Ang II increased NF-kappaB DNA binding (5-fold, 10(-)(9) mol/L at 1 hour; electrophoretic mobility shift assay), nuclear translocation of p50/p65 subunits, and cytosolic inhibitor kappaBalpha (IkappaBalpha) degradation. Ang II elicited NF-kappaB-mediated transcription (transfection of a reporter gene) and expression of NF-kappaB related genes (monocyte chemoattractant protein-1 and angiotensinogen). AT(1) (DUP753) and AT(2) (PD123319 and CGP42112) receptor antagonists inhibited Ang II induced NF-kappaB DNA binding in a dose-dependent manner ( approximately 85% for each one; 10(-)(5) mol/L at 1 hour). The AT(2) agonist p-aminophenylalanine(6) Ang II augmented NF-kappaB binding (4.6-fold, 10(-)(9) mol/L at 1 hour), p65 nuclear levels, and transcription of an NF-kappaB reporter gene. AT(1) antagonist markedly inhibited NF-kappaB-mediated transcription and gene expression. Some differences between AT(1)/AT(2) intracellular signals were found. Antioxidants and ceramide inhibitors, but not protein kinase C inhibitors, diminished NF kappaB activation elicited by both Ang II and the AT(2) agonist, while tyrosine kinase inhibitors only decreased Ang II-induced NF-kappaB activity. Our results demonstrate that Ang II activates NF-kappaB via AT(1) and AT(2), although NF kappaB-mediated transcription occurred mainly through AT(1). Both receptors share some signaling pathways (oxygen radicals and ceramide); however, tyrosine kinases only participate in AT(1)/NF-kappaB responses. These data provide novel insights into Ang II actions, suggesting a potential implication of the AT(2) in the pathobiology of vascular cells. PMID- 10864919 TI - UltraRapid communication : nuclear factor-kappaB and cAMP response element binding protein mediate opposite transcriptional effects on the flk-1/KDR gene promoter AB - -The vascular endothelial growth factor receptor Flk-1/KDR is highly expressed during development and almost disappears in adult tissues. Despite its biological relevance, little is known about the molecular mechanisms controlling its expression. In the present work, it is shown that cAMP response element binding protein (CREB) and nuclear factor-kappaB (NF-kappaB)-related antigens bind specific sequences in the Flk-1/KDR promoter. Functional studies demonstrate that cAMP represses whereas tumor necrosis factor-alpha, an activator of NF-kappaB, stimulates promoter activity. Histone acetyltransferases (HATs) P/CAF and CBP/p300 together with p65/RelA, the catalytic subunit of NF-kappaB, increase Flk 1/KDR promoter activity 10- to 20-fold. Consistently, inhibition by cAMP is reverted by increasing intracellular HATs and is completely abolished by site specific mutagenesis of the cAMP response element. In contrast, specific mutations in the NF-kappaB response element abolish responsiveness to p65/RelA and HATs without affecting cAMP-dependent repression. These results suggest that opposing signaling pathways, activating NF-kappaB or CREB and requiring HAT molecules, control Flk-1/KDR promoter activity. texfThe full text of this article is available at http://www.circresaha.org. Key Words: vascular endothelial growth factor receptor promoter nuclear factor-kappaB transcription angiogenesis Web Site Feature PMID- 10864920 TI - Nuclear factor-kappaB and cAMP response element binding protein mediate opposite transcriptional effects on the Flk-1/KDR gene promoter. AB - -The vascular endothelial growth factor receptor Flk-1/KDR is highly expressed during development and almost disappears in adult tissues. Despite its biological relevance, little is known about the molecular mechanisms controlling its expression. In the present work, it is shown that cAMP response element binding protein (CREB) and nuclear factor-kappaB (NF-kappaB)-related antigens bind specific sequences in the Flk-1/KDR promoter. Functional studies demonstrate that cAMP represses whereas tumor necrosis factor-alpha, an activator of NF-kappaB, stimulates promoter activity. Histone acetyltransferases (HATs) P/CAF and CBP/p300 together with p65/RelA, the catalytic subunit of NF-kappaB, increase Flk 1/KDR promoter activity 10- to 20-fold. Consistently, inhibition by cAMP is reverted by increasing intracellular HATs and is completely abolished by site specific mutagenesis of the cAMP response element. In contrast, specific mutations in the NF-kappaB response element abolish responsiveness to p65/RelA and HATs without affecting cAMP-dependent repression. These results suggest that opposing signaling pathways, activating NF-kappaB or CREB and requiring HAT molecules, control Flk-1/KDR promoter activity. PMID- 10864922 TI - Src homology domain 2-containing tyrosine phosphatase 2 associates with intercellular adhesion molecule 1 to regulate cell survival. AB - Intercellular adhesion molecule-1 (ICAM-1) binds to the plasma protein fibrinogen (Fg) to mediate leukocyte/endothelial cell interactions. In our studies, the ligation of Fg to ICAM-1 on tumor necrosis factor-alpha-stimulated endothelial cells resulted in the tyrosine phosphorylation of Src homology domain 2 (SH2) containing phosphatase-2 (SHP-2). The ICAM-1 cytoplasmic sequence IKKYRLQ conforms poorly to the concensus immunoreceptor tyrosine-based inhibition motifs found in receptors that bind SHP-2. Nevertheless, the tyrosine phosphorylated sequence (IKKpYRLQ) bound specifically to the SH2 domain proximal to the NH(2) terminal of SHP-2 (SHP-2-N) but not to the SH2 domain proximal on the COOH terminal side (SHP-2-C). Phosphorylated ICAM-1 bound SHP-2-N. In immunoprecipitation experiments, SHP-2 associated with phosphorylated ICAM-1. Cells expressing truncated ICAM-1 that lacked the cytoplasmic sequence (ICAM 1(TR)) failed to associate with SHP-2. ICAM-1 containing the tyrosine to alanine substitution at position 485 (ICAM-1(Y485A)) associated weakly with SHP-2. Cells expressing ICAM-1(TR) and ICAM-1(Y485A) underwent apoptosis upon adhesion to Fg, whereas the wild type ICAM-1 maintained cell survival. These results indicate that ICAM-1 interactions with SHP-2 allow better cellular survival mediated through Fg-ICAM-1 ligation. PMID- 10864923 TI - A region in domain II of the urokinase receptor required for urokinase binding. AB - The urokinase receptor is composed of three homologous domains based on disulfide spacing. The contribution of each domain to the binding and activation of single chain urokinase (scuPA) remains poorly understood. In the present paper we examined the role of domain II (DII) in these processes. Repositioning DII to the amino or carboxyl terminus of the molecule abolished binding of scuPA as did deleting the domain entirely. By using alanine-scanning mutagenesis, we identified a 9-amino acid continuous sequence in DII (Arg(137)-Arg(145)) required for both activities. Competition-inhibition and surface plasmon resonance studies demonstrated that mutation of Lys(139) and His(143) to alanine in soluble receptor (suPAR) reduced the affinity for scuPA approximately 5-fold due to an increase in the "off rate." Mutation of Arg(137), Arg(142), and Arg(145), each to alanine, leads to an approximately 100-fold decrease in affinity attributable to a 10-fold decrease in the apparent "on rate" and a 6-fold increase in off rate. These differences were confirmed on cells expressing variant urokinase receptor. suPAR-K139A/H143A displayed a 50% reduction in scuPA-mediated plasminogen activation activity, whereas the 3-arginine variant was unable to stimulate scuPA activity at all. Mutation of the three arginines did not affect binding of a decamer peptide antagonist of scuPA known to interact with DI and DIII. However, this mutation abolished both the binding of soluble DI to DII-III in the presence of scuPA and the synergistic activation of scuPA mediated by DI and wild type DII DIII. These data show that DII is required for high affinity binding of scuPA and its activation. DII does not serve merely as a spacer function but appears to be required for interdomain cooperativity. PMID- 10864924 TI - The high mobility group I/Y protein is hypophosphorylated in endoreduplicating maize endosperm cells and is involved in alleviating histone H1-mediated transcriptional repression. AB - During maize endosperm development, cells shift from a mitotic cycle to endoreduplication, driving the massive synthesis of storage proteins (zeins) and starch. In this developmental context, we studied changes in expression levels of histone H1 and high mobility group I/Y (HMG-I/Y), two chromatin architectural proteins that are known to affect gene transcription. Almost no change was found in the level of histone H1 during endosperm development, despite a dramatic increase in DNA content (endoreduplication); hence, the histone H1/DNA ratio decreased substantially. Concurrently with a reduction in the Cdc2 kinase activity at the shift to endoreduplication, significant changes were found in the level and mobility of the HMG-I/Y protein; the faster migrating forms were, at least partly, hypophosphorylated. Purified maize HMG-I/Y protein was found to be phophorylated in vitro by the Cdc2 kinase and bound efficiently to the gamma-zein promoter AT-rich tract (gammaZ-AT). Using an in vitro transcription assay, we demonstrated the capability of the maize HMG-I/Y protein to relieve the inhibitory effect exerted by histone H1 on templates containing the gammaZ-AT sequence. These data suggest that during maize endosperm development transcription and perhaps replication are controlled, at least partly, by the activity of the Cdc2 kinase and the interplay between histone H1 and HMG-I/Y proteins. PMID- 10864925 TI - The TATA-binding protein-associated factor yTafII19p functionally interacts with components of the global transcriptional regulator Ccr4-Not complex and physically interacts with the Not5 subunit. AB - The Saccharomyces cerevisiae HIS3 gene is a model system to characterize transcription initiation from different types of core promoters. The NOT genes were identified by mutations that preferentially increased transcription of the HIS3 promoter lacking a canonical TATA sequence. They encode proteins associated in a complex that also contains the Caf1 and Ccr4 proteins. It has been suggested that the Ccr4-Not complex represses transcription by inhibiting factors more specifically required for promoters lacking a TATA sequence. A potential target is the yTaf(II)19 subunit of TFIID, which, when depleted, leads to a preferential decrease of HIS3 TATA-less transcription. We isolated conditional taf19 alleles that display synthetic growth phenotypes when combined with not4 or specific not5 alleles. Inactivation of yTaf(II)19p by shifting these mutants to the restrictive temperature led to a more rapid and striking decrease in transcription from promoters that do not contain a canonical TATA sequence. We demonstrated by the two-hybrid assay and directly in vitro that yTaf(II)19p and Not5p could interact. Finally, we found by the two-hybrid assay that yTaf(II)19p also interacted with many components of the Ccr4-Not complex. Taken together, our results provide evidence that interactions between Not5p and yTaf(II)19p may be involved in transcriptional regulation by the Ccr4-Not complex. PMID- 10864926 TI - CCAAT displacement activity involves CUT repeats 1 and 2, not the CUT homeodomain. AB - The CCAAT displacement protein, the homolog of the Drosophila melanogaster CUT protein, contains four DNA-binding domains: three CUT repeats (CR1, CR2, and CR3) and the CUT homeodomain (HD). Using a panel of fusion proteins, we found that a CUT repeat cannot bind to DNA as a monomer, but that certain combinations of domains exhibit high DNA-binding affinity: CR1+2, CR3HD, CR1HD, and CR2HD. One combination (CR1+2) exhibited strikingly different DNA-binding kinetics and specificities. CR1+2 displayed rapid on and off rates and bound preferably to two C(A/G)AT sites, organized as direct or inverted repeats. Accordingly, only CR1+2 was able to bind to the CCAAT sequence, and its affinity was increased by the presence of a C(A/G)AT site at close proximity. A purified CCAAT displacement protein/CUT protein exhibited DNA-binding properties similar to those of CR1+2; and in nuclear extracts, the CCAAT displacement activity also required the simultaneous presence of a C(A/G)AT site. Moreover, CR1+2, but not CR3HD, was able to displace nuclear factor Y. Thus, the CCAAT displacement activity requires the presence of an additional sequence (CAAT or CGAT) and involves CR1 and CR2, but not the CUT homeodomain. PMID- 10864927 TI - An essential phosphorylation-site domain of human cdc25C interacts with both 14-3 3 and cyclins. AB - Human cdc25C is a dual-specificity phosphatase involved in the regulation of cell cycle progression in both unperturbed cells and in cells subject to DNA damage or replication checkpoints. In this study, we describe the structure-function relationship of an essential domain of human cdc25C that interacts with 14-3-3 proteins. We show that this domain is a bi-functional interactive motif that interacts with cyclins primarily through their P-box motif in addition to 14-3-3 proteins. Characterization of the structural features of this domain by NMR and circular dichroism reveals two distinct alpha helical moieties interconnected by a loop carrying the 14-3-3 binding site. Moreover, the helical folding is induced upon binding to 14-3-3, suggestive of a conformational regulation of this domain of cdc25C through interactions with partner proteins in vivo. Combining our structural and biochemical data, we propose a detailed model of the molecular mechanism of cdc25C regulation by differential association with 14-3-3 and cdc2 cyclin B. PMID- 10864928 TI - Location of the glucuronosyltransferase domain in the heparan sulfate copolymerase EXT1 by analysis of Chinese hamster ovary cell mutants. AB - Heparan sulfate formation occurs by the copolymerization of glucuronic acid (GlcA) and N-acetylglucosamine (GlcNAc) residues. Recent studies have shown that these reactions are catalyzed by a copolymerase encoded by EXT1 and EXT2, members of the exostosin family of putative tumor suppressors linked to hereditary multiple exostoses. Previously, we identified a collection of Chinese hamster ovary cell mutants (pgsD) that failed to make heparan sulfate (Lidholt, K., Weinke, J. L., Kiser, C. S., Lugemwa, F. N., Bame, K. J., Cheifetz, S., Massague, J., Lindahl, U., and Esko, J. D. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 2267 2271). Here, we show that pgsD mutants contain mutations that either alter GlcA transferase activity selectively or that affect both GlcNAc and GlcA transferase activities. Expression of EXT1 corrects the deficiencies in the mutants, whereas EXT2 and the related EXT-like cDNAs do not. Analysis of the EXT1 mutant alleles revealed clustered missense mutations in a domain that included a (D/E)X(D/E) motif thought to bind the nucleotide sugar from studies of other transferases. These findings provide insight into the location of the GlcA transferase subdomain of the enzyme and indicate that loss of the GlcA transferase domain may be sufficient to cause hereditary multiple exostoses. PMID- 10864929 TI - Inhibition of the initiation of HIV-1 reverse transcription by 3'-azido-3' deoxythymidine. Comparison with elongation. AB - Initiation of human immunodeficiency virus-1 reverse transcription requires formation of a complex containing the viral RNA, primer tRNA(3)(Lys), and reverse transcriptase. Initiation, corresponding to addition of the first six nucleotides to tRNA(3)(Lys), is distinguished from elongation by its high specificity and low efficiency (processivity). Here, we compared the inhibition of initiation and elongation of reverse transcription by 3'-azido-3'-deoxythymidine 5'-triphosphate (AZTTP), the active form of 3'-azido-3'-deoxythymidine. We report the first detailed study of nucleotide binding, discrimination, and pyrophosphorolysis by the authentic initiation complex. We showed that the initiation and elongation complexes bound AZTTP and dTTP with the same affinity, while the polymerization rates were reduced by 148-160-fold during initiation. The pyrophosphorolysis rate of dTTP was reduced by the same extent, indicating that the polymerization equilibrium is the same in the two phases. The efficient unblocking of the 3' azido-3'-deoxythymidine 5'-monophosphate (AZTMP)-terminated primer by pyrophosphorolysis significantly relieved inhibition of DNA synthesis during elongation in the presence of physiological pyrophosphate concentrations. Remarkably, although pyrophosphorolysis of dTMP and AZTMP were equally efficient during elongation, reverse transcriptase was almost totally unable to unblock the AZTMP-terminated primer during initiation. As a result, inhibition of reverse transcription by AZTTP was more efficient during initiation than elongation of reverse transcription, despite a reduced selectivity of incorporation. PMID- 10864930 TI - Membrane recruitment of coatomer and binding to dilysine signals are separate events. AB - It has previously been shown that transport of newly synthesized proteins and the structure of the Golgi complex are affected in the Chinese hamster ovary cell line ldlF, which bears a temperature-sensitive mutation in the Coat protein I (COPI) subunit epsilon-COP (Guo, Q., Vasile, E., and Krieger, M. (1994) J. Cell Biol. 125, 1213-1224; Hobbie, L., Fisher, A. S., Lee, S., Flint, A., and Krieger, M. (1994) J. Biol. Chem. 269, 20958-20970). Here, we pinpoint the site of the secretory block to an intermediate compartment between the endoplasmic reticulum (ER) and the Golgi complex and show that the distributions of ER-Golgi recycling proteins, such as KDEL receptor and p23, as well as resident Golgi proteins, such as mannosidase II, are accordingly affected. At the nonpermissive temperature, neither the stability of the COPI complex nor its recruitment to donor Golgi membranes is affected. However, the binding of coatomer to the dilysine-based ER retrieval motif is impaired in the absence of epsilon-COP, suggesting that dilysine signal binding is not the major means of COPI recruitment. Because expression of the exogenous chimera of epsilon-COP and green fluorescent protein in ldlF cells at nonpermissive temperature rapidly restores the wild type properties, epsilon-COP is likely to play an important role in the cargo selection events mediated by COPI. PMID- 10864931 TI - A new model of cooperative myosin-thin filament binding. AB - Cooperative myosin binding to the thin filament is critical to regulation of cardiac and skeletal muscle contraction. This report delineates and fits to experimental data a new model of this process, in which specific tropomyosin actin interactions are important, the tropomyosin-tropomyosin polymer is continuous rather than disjointed, and tropomyosin affects myosin-actin binding by shifting among three positions as in recent structural studies. A myosin- and tropomyosin-induced conformational change in actin is proposed, rationalizing the approximately 10,000-fold strengthening effect of myosin on tropomyosin-actin binding. Also, myosin S1 binding to regulated filaments containing mutant tropomyosins with internal deletions exhibited exaggerated cooperativity, implying an allosteric effect of tropomyosin on actin and allowing the effect's measurement. Comparisons among the mutants suggest the change in actin is promoted much more strongly by the middle of tropomyosin than by its ends. Regardless of calcium binding to troponin, this change in actin facilitates the shift in tropomyosin position to the actin inner domain, which is required for tight myosin-actin association. It also increases myosin-actin affinity 7-fold compared with the absence of troponin-tropomyosin. Finally, initiation of a shift in tropomyosin position is 100-fold more difficult than is its extension from one actin to the next, producing the myosin binding cooperativity that underlies cooperative activation of muscle contraction. PMID- 10864932 TI - The amino-terminal cyclic nucleotide binding site of the type II cGMP-dependent protein kinase is essential for full cyclic nucleotide-dependent activation. AB - For the type I cGMP-dependent protein kinases (cGKIalpha and cGKIbeta), a high affinity interaction exists between the C2 amino group of cGMP and the hydroxyl side chain of a threonine conserved in most cGMP binding sites. To examine the effect of this interaction on ligand binding and kinase activation in the type II isozyme of cGMP-dependent protein kinase (cGKII), alanine was substituted for the conserved threonine or serine. cGKII was found to require the C2 amino group of cGMP and its cognate serine or threonine hydroxyl for efficient cGMP activation. Of the two binding sites, disruption of cGMP-specific binding in the NH(2) terminal binding site had the greatest effect on cGMP-dependent kinase activation, like cGKI. However, ligand dissociation studies showed that the location of the rapid and slow dissociation sites of cGKII was reversed relative to cGKI. Another set of mutations that prevented cyclic nucleotide binding demonstrated the necessity of the NH(2)-terminal, rapid dissociation binding site for cyclic nucleotide-dependent activation of cGKII. These findings suggest distinct mechanisms of activation for cGKII and cGKI isoforms. Because cGKII mediates the effects of heat-stable enterotoxins via the cystic fibrosis transmembrane regulator Cl(-) channel, these findings define a structural target for drug design. PMID- 10864933 TI - Self recognition in the Ig superfamily. Identification of precise subdomains in carcinoembryonic antigen required for intercellular adhesion. AB - The homophilic binding of extracellular domains of membrane-bound immunoglobulin superfamily (IgSF) molecules is often required for intercellular adhesion and signaling. Carcinoembryonic antigen (CEA), a member of the IgSF, is a widely used tumor marker that functions in vitro as a homotypic intercellular adhesion molecule. CEA has also been shown to contribute to tumorigenicity by inhibiting cellular differentiation, an effect that requires the homophilic binding of its extracellular domains. It was of interest, therefore, to identify small subdomain sequences in CEA that could serve as a focus in the design of peptides that disrupt CEA-mediated intercellular adhesion. Three subdomains in the N-terminal domain of CEA, identified by site-directed deletions and point mutations, were shown to be required for intercellular adhesion. Cyclized peptides representing two of these subdomains, (42)NRQII and (80)QNDTG, were found to be effective in blocking CEA-mediated cellular aggregation when added to CEA-expressing transfectants in suspension. Intermolecular binding involving each of these subdomains is therefore essential for intercellular adhesion and cannot be compensated for by known binding contributions of other regions in the CEA molecule. In further support of this assumption, the binding epitope of an anti CEA monoclonal antibody (monoclonal antibody A20) known to block CEA-mediated adhesion, was shown to bridge two of the three required subdomains: (42)NRQII and (30)GYSWYK. PMID- 10864934 TI - Alternative splicing in the cytoplasmic II-III loop of the N-type Ca channel alpha 1B subunit: functional differences are beta subunit-specific. AB - Structural diversity of voltage-gated Ca channels underlies much of the functional diversity in Ca signaling in neurons. Alternative splicing is an important mechanism for generating structural variants within a single gene family. In this paper, we show the expression pattern of an alternatively spliced 21 amino acid encoding exon in the II-III cytoplasmic loop region of the N-type Ca channel alpha(1B) subunit and assess its functional impact. Exon-containing alpha(1B) mRNA dominated in sympathetic ganglia and was present in approximately 50% of alpha(1B) mRNA in spinal cord and caudal regions of the brain and in the minority of alpha(1B) mRNA in neocortex, hippocampus, and cerebellum (<20%). The II-III loop exon affected voltage-dependent inactivation of the N-type Ca channel. Steady-state inactivation curves were shifted to more depolarized potentials without affects on either the rate or voltage dependence of channel opening. Differences in voltage-dependent inactivation between alpha(1B) splice variants were most clearly manifested in the presence of Ca channel beta(1b) or beta(4), rather than beta(2a) or beta(3), subunits. Our results suggest that exon lacking alpha(1B) splice variants that associate with beta(1b) and beta(4) subunits will be susceptible to voltage-dependent inactivation at voltages in the range of neuronal resting membrane potentials (-60 to -80 mV). In contrast, alpha(1B) splice variants that associate with either beta(2a) or beta(3) subunits will be relatively resistant to inactivation at these voltages. The potential to mix and match multiple alpha(1B) splice variants and beta subunits probably represents a mechanism for controlling the plasticity of excitation-secretion coupling at different synapses. PMID- 10864936 TI - Sustained activation of hippocampal Lp-type voltage-gated calcium channels by tetanic stimulation. AB - The molecular heterogeneity of voltage-gated calcium channels is mirrored by extensive biophysical diversity. Subtype-selective antagonists have been used to place different kinds of calcium channels in functional categories. Dihydropyridine (DHP) antagonists have been used, for example, to implicate L type calcium channels in the induction of NMDA receptor-independent forms of synaptic plasticity. DHPs, however, do not discriminate between the recently identified Lp and Ls subtypes of L-type calcium channel. The different properties of the two kinds of L-type channels suggest that they may have different functional roles. Ls channels are comparable with cardiac L-type channels, whereas Lp channels show low-threshold voltage-dependent potentiation. To clarify the potential roles of Lp and Ls channels in the induction of synaptic plasticity, we studied the responses of these channels to trains of action potentials. The frequency and duration of the trains were chosen to mimic the stimuli used to induce changes in synaptic strength. Cell-attached single-channel recordings from cultured hippocampal neurons revealed that both Lp and Ls channels responded to these trains, but only Lp channels showed persistent activation that outlasted the train. The magnitude of Lp channel activity increased with increasing action potential frequency and train duration. Stimuli that reproduced the postsynaptic response to action potential trains were also examined, and Lp channels were found to show much greater responses than were Ls channels. These results suggest that the Lp channel may play a critical role in the induction of long-lasting changes in synaptic strength. PMID- 10864935 TI - Bidirectional modulation of exocytosis by angiotensin II involves multiple G protein-regulated transduction pathways in chromaffin cells. AB - Angiotensin II (AngII) receptors couple to a multitude of different types of G proteins resulting in activation of numerous signaling pathways. In this study we examined the consequences of this promiscuous G-protein coupling on secretion. Chromaffin cells were voltage-clamped at -80 mV in perforated-patch configuration, and Ca(2+)-dependent exocytosis was evoked with brief voltage steps to +20 mV. Vesicle fusion was monitored by changes in membrane capacitance (DeltaC(m)), and released catecholamine was detected with single-cell amperometry. Ca(2+) signaling was studied by recording voltage-dependent Ca(2+) currents (I(Ca)) and by measuring intracellular Ca(2+) ([Ca(2+)](i)) with fura-2 AM. AngII inhibited I(Ca) (IC(50) = 0.3 nm) in a voltage-dependent, pertussis toxin (PTX)-sensitive manner consistent with G(i/o)-protein coupling to Ca(2+) channels. DeltaC(m) was modulated bi-directionally; subnanomolar AngII inhibited depolarization-evoked exocytosis, whereas higher concentrations, in spite of I(Ca) inhibition, potentiated DeltaC(m) fivefold (EC(50) = 3.4 nm). Potentiation of exocytosis by AngII involved activation of phospholipase C (PLC) and Ca(2+) mobilization from internal stores. PTX treatment did not affect AngII-dependent Ca(2+) mobilization or facilitation of exocytosis. However, protein kinase C (PKC) inhibitors decreased the facilitatory effects but not the inhibitory effects of AngII on stimulus-secretion coupling. The AngII type 1 receptor (AT1R) antagonist losartan blocked both inhibition and facilitation of secretion by AngII. The results of this study show that activation of multiple types of G proteins and transduction pathways by a single neuromodulator acting through one receptor type can produce concentration-dependent, bi-directional regulation of exocytosis. PMID- 10864937 TI - Ultrastructural localization of nitrotyrosine within the caudate-putamen nucleus and the globus pallidus of normal rat brain. AB - Nitration of protein tyrosine residues by nitric oxide (NO)-derived reactive species results in the production of stable nitrotyrosine (NT) moieties that are immunochemically detectable in many regions of normal brain and enriched in those areas containing constitutive nitric oxide synthase (cNOS). These include the caudate-putamen nucleus (CPN) and the globus pallidus, which receives major inhibitory input from the CPN. To determine the functional sites for NT production in these critical motor nuclei, we examined the electron microscopic immunocytochemical localization of NT and cNOS in rat brain. In the CPN, NT was localized to the somata and dendrites of cNOS-containing interneurons and spiny neurons, some of which received input from cNOS-labeled terminals. The NT immunoreactivity was most prevalent on outer mitochondrial membranes and nearby segments of the plasma membranes in dendrites and within asymmetric synapses on dendritic spines. In the CPN and globus pallidus, there was also a prominent labeling of NT in astrocytic processes, small axons, and tubulovesicles and/or synaptic vesicles in axon terminals. These terminals formed mainly asymmetric synapses in the CPN and inhibitory-type synapses in the globus pallidus where they often apposed cNOS-containing terminals that also formed asymmetric, excitatory-type synapses. Our results suggest that NT is generated by mechanisms requiring the dual actions of excitatory transmitters and NO derived either from interneurons in the CPN or from excitatory afferents in the globus pallidus. The findings also implicate NT in the physiological actions of NO within the striatal circuitry and, particularly, in striatopallidal neurons severely affected in Huntington's disease. PMID- 10864938 TI - Altered stress-induced anxiety in adenylyl cyclase type VIII-deficient mice. AB - Stress results in alterations in behavior and physiology that can be either adaptive or maladaptive. To define the molecular pathways involved in the response to stress further, we generated mice deficient (KO) in the calcium stimulated adenylyl cyclase type VIII (AC8) by homologous recombination in embryonic stem cells. AC8 KO mice demonstrate a compromise in calcium-stimulated AC activity in the hippocampus, hypothalamus, thalamus, and brainstem. Hippocampal slices derived from AC8 KO mice fail to demonstrate CA1-region long term depression after low-frequency stimulation, and AC8 KO mice also fail to activate CRE-binding protein in the CA1 region after restraint stress. To define the behavioral consequences of AC8 deficiency, we evaluated AC8 KO mice in the elevated plus-maze and open field. Although naive AC8 KO mice exhibit indices of anxiety comparable with that of wild-type mice, AC8 KO mice do not show normal increases in behavioral markers of anxiety when subjected to repeated stress such as repetitive testing in the plus-maze or restraint preceding plus-maze testing. These results demonstrate a novel role for AC8 in the modulation of anxiety. PMID- 10864939 TI - Traumatic brain injury alters the molecular fingerprint of TUNEL-positive cortical neurons In vivo: A single-cell analysis. AB - The cerebral cortex is selectively vulnerable to cell death after traumatic brain injury (TBI). We hypothesized that the ratio of mRNAs encoding proteins important for cell survival and/or cell death is altered in individual damaged neurons after injury that may contribute to the cell's fate. To investigate this possibility, we used amplified antisense mRNA (aRNA) amplification to examine the relative abundance of 31 selected candidate mRNAs in individual cortical neurons with fragmented DNA at 12 or 24 hr after lateral fluid percussion brain injury in anesthetized rats. Only pyramidal neurons characterized by nuclear terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick end labeling (TUNEL) reactivity with little cytoplasmic staining were analyzed. For controls, non TUNEL-positive neurons from the cortex of sham-injured animals were obtained and subjected to aRNA amplification. At 12 hr after injury, injured neurons exhibited a decrease in the relative abundance of specific mRNAs including those encoding for endogenous neuroprotective proteins. By 24 hr after injury, many of the mRNAs altered at 12 hr after injury had returned to baseline (sham-injured) levels except for increases in caspase-2 and bax mRNAs. These data suggest that TBI induces a temporal and selective alteration in the gene expression profiles or "molecular fingerprints" of TUNEL-positive neurons in the cerebral cortex. These patterns of gene expression may provide information about the molecular basis of cell death in this region after TBI and may suggest multiple avenues for therapeutic intervention. PMID- 10864941 TI - Unique properties of NMDA receptors enhance synaptic excitation of radiatum giant cells in rat hippocampus. AB - In the hippocampus, fast excitatory synaptic transmission of principal projection neurons is mediated by non-NMDA glutamate receptors, whereas NMDA glutamate receptors serve a slower modulatory role. We used the whole-cell patch-clamp technique in adult hippocampal slices to assess the role of NMDA receptors in synaptic excitation of a recently discovered excitatory projection neuron, the CA1 radiatum giant cell (RGC). Glutamatergic synaptic activation, even after blocking non-NMDA receptors, fired an NMDA receptor-dependent burst of action potentials in RGCs. In contrast, the contribution of NMDA receptors to synaptic activation of pyramidal cells (PCs) was minimal. Stimulation of the same synaptic inputs evoked greater than threefold larger EPSCs in RGCs than in PCs. Isolated NMDA receptor-mediated EPSCs were significantly less sensitive to blockade by extracellular Mg(2+) and had slower decay kinetics in RGCs than in PCs. Thus, unique properties of synaptic NMDA receptors underlie enhanced synaptic excitability in a newly discovered excitatory hippocampal projection neuron. PMID- 10864940 TI - Ionic mechanisms underlying repetitive high-frequency burst firing in supragranular cortical neurons. AB - Neocortical neurons in awake, behaving animals can generate high-frequency (>300 Hz) bursts of action potentials, either in single bursts or in a repetitive manner. Intracellular recordings of layer II/III pyramidal neurons were obtained from adult ferret visual cortical slices maintained in vitro to investigate the ionic mechanisms by which a subgroup of these cells generates repetitive, high frequency burst discharges, a pattern referred to as "chattering." The generation of each but the first action potential in a burst was dependent on the critical interplay between the afterhyperpolarizations (AHPs) and afterdepolarizations (ADPs) that followed each action potential. The spike-afterdepolarization and the generation of action potential bursts were dependent on Na(+), but not Ca(2+), currents. Neither blocking of the transmembrane flow of Ca(2+) nor the intracellular chelation of free Ca(2+) with BAPTA inhibited the generation of intrinsic bursts. In contrast, decreasing the extracellular Na(+) concentration or pharmacologically blocking Na(+) currents with tetrodotoxin, QX-314, or phenytoin inhibited bursting before inhibiting action potential generation. Additionally, a subset of layer II/III pyramidal neurons could be induced to switch from repetitive single spiking to a burst-firing mode by constant depolarizing current injection, by raising extracellular K(+) concentrations, or by potentiation of the persistent Na(+) current with the Na(+) channel toxin ATX II. These results indicate that cortical neurons may dynamically regulate their pattern of action potential generation through control of Na(+) and K(+) currents. The generation of high-frequency burst discharges may strongly influence the response of postsynaptic neurons and the operation of local cortical networks. PMID- 10864942 TI - Excitatory role of the hyperpolarization-activated inward current in phasic and tonic firing of rat supraoptic neurons. AB - The properties and functional roles of the hyperpolarization-activated inward current (I(H)) in magnocellular neurosecretory cells (MNCs) were investigated during sharp microelectrode recordings from supraoptic neurons in superfused explants of rat hypothalamus. Under current clamp, voltage responses to hyperpolarizing current pulses featured depolarizing sags that were abolished by the I(H) blocker ZD 7288. Under voltage clamp, subtraction of current responses to hyperpolarizing steps recorded in the absence and presence of ZD 7288 was used to investigate the properties of I(H). Current-voltage analysis revealed that steady-state I(H) amplitude increases with hyperpolarization, with half-maximal activation of the underlying conductance occurring at -78 mV. The time course of activation of I(H) during hyperpolarizing steps was monoexponential with time constants (100-800 msec) decreasing with hyperpolarization. The effects of ZD 7288 on I(H) were slow (tau, approximately 15 min), irreversible, and half maximal at 1.8 micrometer. When tested on continuously active MNCs, application of 30-60 micrometer ZD 7288 caused a significant reduction in firing rate. In phasically active MNCs, the drug decreased burst duration and intraburst firing frequency and caused an increase in the duration of interburst intervals. These effects were accompanied with a small hyperpolarization of the membrane potential. In contrast, ZD 7288 had no effect on spike duration, on the amplitude of calcium-dependent afterpotentials, or on the frequencies and amplitudes of spontaneous synaptic potentials. These results confirm the presence of I(H) in MNCs of the rat supraoptic nucleus and suggest that the presence of this conductance provides an excitatory drive that contributes to phasic and tonic firing. PMID- 10864943 TI - Long-term specification of AMPA receptor properties after synapse formation. AB - AMPA receptors expressed at auditory nerve synapses in the mammalian and avian cochlear nuclei display exceptionally rapid channel gating, an adaptation well suited for acoustic processing. We examined whether cellular interactions during development might determine the subunit composition of these receptors. After synapse formation in the avian nucleus magnocellularis (nMag) in vivo, the rate of receptor desensitization increased threefold, sensitivity to channel block by polyamines increased, and sensitivity to cyclothiazide, an inhibitor of desensitization, increased, indicating a reduction in glutamate receptor subunit 2 and of flip splice variants. This phenotypic switch was prevented, but not reversed, by isolating nMag neurons in a cell-culture environment. We propose that the switch in receptor kinetics is an outcome of cellular interactions during a critical period that result in the long-term determination of receptor phenotype. PMID- 10864944 TI - Coexpression of rat P2X2 and P2X6 subunits in Xenopus oocytes. AB - Transcripts for P2X(2) and P2X(6) subunits are present in rat CNS and frequently colocalize in the same brainstem nuclei. When rat P2X(2) (rP2X(2)) and rat P2X(6) (rP2X(6)) receptors were expressed individually in Xenopus oocytes and studied under voltage-clamp conditions, only homomeric rP2X(2) receptors were fully functional and gave rise to large inward currents (2-3 microA) to extracellular ATP. Coexpression of rP2X(2) and rP2X(6) subunits in Xenopus oocytes resulted in a heteromeric rP2X(2/6) receptor, which showed a significantly different phenotype from the wild-type rP2X(2) receptor. Differences included reduction in agonist potencies and, in some cases (e.g., Ap(4)A), significant loss of agonist activity. ATP-evoked inward currents were biphasic at the heteromeric rP2X(2/6) receptor, particularly when Zn(2+) ions were present or extracellular pH was lowered. The pH range was narrower for H(+) enhancement of ATP responses at the heteromeric rP2X(2/6) receptor. Also, H(+) ions inhibited ATP responses at low pH levels (A substitutions at 227; 7 blacks had A-->G substitutions at 277; and 1 Japanese person had C-->G at 141 and G-->T at 227. A at 227 and G at 277 represent expected nts of NA1 FcgammaRIIIB. One black had an NA1 FcgammaRIIIB with a G-->A substitution at 349; A is normally found in NA2 FcgammaRIIIB at 349. Sequencing atypical FcgammaRIIIA in three persons revealed that two blacks had G- >A substitutions at 277 plus C-->A substitutions at 266 and 1 white had previously described T-->G at 230. Two blacks with atypical NA2 FcgammaRIIIB had T-->G FcgammaRIIIA at 230. One black was NA(null). CONCLUSION: NA2 FcgammaRIIIB is more polymorphic in blacks than in whites or Japanese persons. Chimeric FcgammaRIIIB alleles are most similar to NA2 FcgammaRIIIB. One alternate allele of NA1 (NA1*02) and four alternate alleles of NA2 (NA2*02, NA2*03, NA2*04, and NA2*05) are described. PMID- 10864984 TI - Variations in the expression of granulocyte antigen NB1. AB - BACKGROUND: Between 87 and 97 percent of whites express NB1 alloantigen on some but not all of their granulocytes. The expression of NB1 has not been compared among large groups of adults of different sexes, ages, and ethnic groups. Previous testing of whites suggests that the expression of NB1 is variable. STUDY DESIGN AND METHODS: Serologic testing of granulocytes from 224 persons with two examples of MoAb to NB1 (1B5 and 7D8) was performed to distinguish phenotypic differences among age, sex, and ethnic groups and differences in reactivity to MoAbs. The donors were from 17 to 82 years of age, and 87 were female. They were from four ethnic backgrounds: 54 were African American (black), 10 were Asian, 9 Hispanic, and 152 white. Granulocytes were tested by flow cytometry. Parallel testing with MoAbs to CD16, CD11b, and CD45 served as controls. The size of the granulocyte population reacting with 1B5 and 7D8 and the respective mean, median, and peak cell fluorescence intensities were analyzed. RESULTS: The expression of 7D8 and 1B5 was greater on granulocytes from female donors. The expression of 7D8 fell in older women but not in men. There were no differences among the four racial groups in either the frequency of NB1 as determined by 1B5 or 7D8 or in the size of the population of granulocytes reacting with either antibody. When the fluorescence intensities of the antibody reactions were compared among groups, there were no differences in reactivity with 1B5. However, reactions with 7D8 were all greater in blacks. Comparison of the size of the antigen-positive granulocyte population, as determined by antibody reactivity, showed that only 30 donors differed by more than 10 percent. These discordant results were more likely to occur in whites than in blacks (18% vs. 4%, p<0.02). CONCLUSIONS: NB1 is composed of at least two epitopes as determined by serologic studies. The expression of both antigens is greater in females. The 7D8-reactive epitope appears to be more prevalent or more accessible on granulocytes of blacks. Variations in the expression of NB1 are more likely to occur in whites. The biochemical and molecular basis of these variations are not known. PMID- 10864985 TI - Immune hemolytic anemia caused by sensitivity to a metabolite of etodolac, a nonsteroidal anti-inflammatory drug. AB - BACKGROUND: Immune hemolytic anemia can be caused by sensitivity to many different drugs. In some instances, the sensitizing compound can be identified by in vitro testing, but results are often negative. One reason for this is that a drug metabolite formed in vivo can be the sensitizing agent, but the responsible metabolites have rarely been identified at a chemical level. This report describes a patient who developed severe, Coombs-positive hemolytic anemia on two occasions after taking the nonsteroidal anti-inflammatory drug etodolac. Studies were performed to characterize etodolac metabolites to which this patient was sensitive. CASE REPORT: Serum was tested for antibody in the presence and absence of drug using conventional methods and urine from individuals taking etodolac as a source of drug metabolites. Urinary metabolites of etodolac were identified by high-pressure liquid chromatography analysis. Glucuronide conjugates of etodolac and the 6-OH metabolite of etodolac were synthesized in a rat liver microsomal system to obtain reference standards. RESULTS: The patient's serum gave only trace (+/-) reactions with normal RBCs in the presence of etodolac but reacted strongly (4+) in the presence of urine from an individual taking this drug. The active urinary metabolites were identified as etodolac glucuronide and 6-OH etodolac glucuronide. CONCLUSION: This patient appears to have experienced acute, severe immune hemolytic anemia on two occasions because of sensitivity to the glucuronides of etodolac and 6-OH etodolac. In patients suspected of having drug induced immune hemolytic anemia, RBC-reactive antibodies can sometimes be detected by using urine from an individual taking the implicated medication as the source of drug metabolites in in vitro reactions. For patients who present with acute immune hemolysis, a careful history of drug exposure should be taken, and, where indicated, confirmatory testing should be performed to identify the sensitizing drug and prevent inadvertent reinduction of hemolysis at a later time. PMID- 10864987 TI - Cost-effectiveness of epoetin and autologous blood donationin reducing allogeneic blood transfusions incoronary artery bypass graft surgery. AB - BACKGROUND: Coronary artery bypass graft (CABG) surgery accounts for a substantial portion of all allogeneic units of blood transfused. Drugs and autologous blood donation (ABD) are alternative or adjunctive methods for reducing complications and costs induced by allogeneic blood transfusions. Recombinant human erythropoietin (epoetin) has the potential to decrease perioperative need for allogeneic blood during CABG, but its high cost calls for a careful economic evaluation before it can be recommended for widespread use. STUDY DESIGN AND METHODS: A decision tree was used to compare a hypothetical strategy of no epoetin with one in which epoetin was utilized to control blood transfusion needs in CABG; each strategy was tested with and without ABD. The impact of these strategies on both the quality-adjusted life years (QALYs) and costs ($US) was calculated. RESULTS: Using epoetin alone and with ABD, respectively, avoided the transfusion of 0.61 and 1.35 units of allogeneic blood per patient and saved 0.000086 and 0.000146 QALYs per patient. This made cost effectiveness (CE) higher than $7 million and $5 million for each QALY saved, respectively. ABD alone cost more than $1 million per QALY saved. If the risk of bacterial infections following allogeneic transfusions was included in the model, epoetin alone cost $6288 per QALY saved, while ABD, both alone and with epoetin, saved money. CONCLUSION: On the basis of the existing evidence, neither of the blood-saving strategies modeled was a cost-effective means of avoiding the deleterious health effects of perioperative blood transfusions in CABG. However, if allogeneic blood-related infections were to be considered, both ABD and epoetin would be acceptable interventions. PMID- 10864986 TI - Evaluation of polyethylene terephthalate for ABO and Rh typing and alloantibody screening. AB - BACKGROUND: For many years, hospitals and laboratories have used evacuated glass tubes for blood collection. To improve the safety of blood collection, plastic polyethylene terephthalate (PET) tubes (Vacutainer PLUS, Becton Dickinson) have been developed. The objectives of this study were to compare the accuracy of ABO grouping, Rh typing, and antibody screening of blood samples collected in plastic tubes with that in glass tubes and to determine if refrigerated blood samples collected in plastic tubes remained stable over a 28-day period. STUDY DESIGN AND METHODS: Samples were collected from 121 volunteers, at least 30 from each of the A, B, O, and AB blood groups, in four types of Vacutainer tubes: silica-coated plastic, K(2) EDTA plastic, uncoated glass, and K(2) EDTA glass. Samples from each tube were tested for ABO group and Rh type by use of the microtyping gel identification card system and the tube method. A three-cell antibody screen was performed by the microtyping gel card technique with a monospecific IgG reagent. Initial samples were tested within 3 hours of collection. Refrigerated samples were retested for ABO and Rh type and antibody screening 1, 2, 21, and 28 days later. Agreement between test results was determined by using Cohen's Kappa statistic. RESULTS: Complete agreement was observed between the ABO and Rh typing results in samples drawn into glass and plastic tubes of both the EDTA and nonanticoagulated type (kappa = 1.0). In retesting, there were no examples of a change in ABO or Rh type over the 28-day study period. Only two alloantibodies (1.7%) were identified in the 121 samples, and no difference was observed in alloantibody expression in either plastic or glass Vacutainer tubes over the 28 day study period. CONCLUSION: Samples collected into the PET serum or EDTA tubes provided accurate ABO and Rh typing results that remained consistent over a 28 day period. Samples collected in these tubes also appeared to enable accurate alloantibody identification. However, the number of alloantibodies identified in this study was small, and this result should be confirmed in a larger series. PMID- 10864988 TI - Compodock, a new device for sterile docking. AB - BACKGROUND: A new device for sterile docking, the Compodock (Fresenius NPBI Transfusion Technology), was developed for connecting PVC tubing for medical use while maintaining sterility. STUDY DESIGN AND METHODS: Sterility of the connections was assessed by welding tubing with a heavy exterior contamination with Bacillus subtilis spores and also by welding in an environment contaminated with aerosols of B. subtilis. Tubing was either dry or liquid-filled ("wet") and had various diameters. Bacterial culture medium was flushed through the welded area and subsequently cultured. Tensile strength was measured, and, under semi routine conditions, Compodock was tested for user friendliness and speed. RESULTS: None of the cultures of welded tubing with exterior contamination showed growth, neither the dry-dry (n = 434) nor the wet-wet connections (n = 622). Cultures were also negative for welds made in the contaminated environment (dry dry, 67; wet-wet, 55). Tensile strength complied fully with ISO 3826 standards (that is, a force of 20 newtons [N] for 15 sec), with a mean maximal strength ranging from 73 to 100 N, depending on diameter and content of the tubing. The semi-routine handling was regarded as good: welds were easily opened; there were clear instructions and error warnings; and the processing time averaged 52 seconds. CONCLUSION: The Compodock is able to maintain a functionally closed system, with maintenance of sterility, despite heavy exterior bacterial contamination; tensile strength conformed to ISO standards. Compodock is suitable for routine implementation in the blood bank. PMID- 10864989 TI - Periodic alternating interface positioning to lower WBC contamination of apheresis platelet concentrates: a multicenter evaluation. AB - BACKGROUND: A new software version of a cell separator (AS TEC 204, Fresenius) providing WBC-reduced single-donor plateletpheresis concentrates was tested. STUDY DESIGN AND METHODS: Dual-needle apheresis procedures (n = 621) were performed in three centers, using either fixed interface positioning (FIP) or periodic alternating interface positioning (PAIP). The other separation parameters (e.g., anticoagulant:whole-blood ratio, and blood flow) were set individually. All platelet concentrates were evaluated for platelet yields and contaminating WBCs. RESULTS: The introduction of the PAIP resulted in a significant (p<0.001) reduction in contaminating WBCs (median, 30,000) from the numbers seen with FIP (median, 2,300,000) while maintaining the separation efficacy (47%) and separation time. Ninety-eight percent of all concentrates contained less than 5 x 10(6) WBCs per concentrate and 92 percent contained less than 1 x 10(6). CONCLUSION: Plateletpheresis using the AS TEC 204 cell separator with PAIP is a valid alternative to WBC reduction by filtration. It may provide WBC-reduced platelet concentrates without the additional cost of filters. However, the reliability of the WBC reduction is not yet advanced enough that PAIP can be employed without any monitoring of the end product. PMID- 10864990 TI - HPC viability measurement: trypan blue versus acridine orange and propidium iodide. AB - BACKGROUND: A reliable, validated method for rapidly determining HPC viability is essential for clinical cell engineering. STUDY DESIGN AND METHODS: A fluorometric cell viability assay using acridine orange and propidium iodide (AO/PI) was compared to the current standard, trypan blue (TB) exclusion. Viable cells stained with AO/PI fluoresce green under darkfield fluorescence microscopy, while nonviable cells fluoresce orange. Mixtures of fresh and heat-killed bone marrow were prepared and used as viability standards for evaluation of both assays. The frequency of CFU-GM was determined for each specimen. RESULTS: Cell viability measured by AO/PI was extremely linear, with measured and predicted viability in agreement from 0 to 100 percent of the viable cells and a coefficient of regression (r(2)) of 0.9921. The predicted-viability regression line fell within the 95% CI for AO/PI-measured viability. The coefficient of regression for TB measured viability was 0.9584, with the predicted-viability regression line almost entirely outside the 95% CI. TB overestimated the percentage of viable cells, particularly below the 50-percent level. CFU-GM frequency correlated better with cell viability measured by AO/PI (r(2) = 0.979) than with that measured by TB (r(2) = 0.930). CONCLUSIONS: The AO/PI viability assay is a rapid, highly linear, functionally correlated assay that is superior to conventional viability measurement by TB exclusion. PMID- 10864991 TI - Heparin-induced coagulopathy associated with staphylococcal protein A immunoadsorption treatment columns: an in vitro and in vivo analysis. AB - BACKGROUND: The staphylococcal protein A (SPA) column used to treat refractory autoimmune and alloimmune thrombocytopenia and rheumatoid arthritis patients is primed with heparin to prevent possible fibrin clot formation when the patient's plasma is passed through the column. A BMT patient with refractory alloimmune thrombocytopenia had prolonged activated partial thromboplastin times (aPTTs) at the end of SPA column treatments. This observation led to in vivo and in vitro analysis of the kinetics of heparin elution from the SPA column. STUDY DESIGN AND METHODS: Two patients with refractory rheumatoid arthritis, who were treated on five occasions with the SPA column (as a part of a national trial) primed with 5000 U of heparin, were monitored for aPTT and heparin in their plasma. In addition, two in vitro analyses were performed with FFP for heparin elution from the SPA column. RESULTS: The in vivo studies showed the presence of 0.3 to 1.5 U per mL of heparin in patients' plasma at the end of the SPA column treatments that corresponded with the prolonged aPTTs. The in vitro studies showed that 82 to 85 percent heparin (approx. 4400 U) was eluted from the SPA column during rather than before the procedure. CONCLUSION: Patients undergoing SPA column treatments, especially those with thrombocytopenia, may be at increased risk of bleeding as a result of the presence of a significant amount of heparin in their circulation during the entire period of SPA column treatment. PMID- 10864992 TI - Automated RBC exchange transfusion:treatment for cerebral malaria. AB - BACKGROUND: Cerebral malaria is a life-threatening complication of Plasmodium falciparum infection. RBC exchange transfusion can reduce the level of parasitemia in this setting. Experience with automated RBC exchange for cerebral malaria may be limited, as most cases occur when the necessary equipment and blood components are not readily available. CASE REPORTS: Three patients were admitted with cerebral malaria. Parasites were found in more than 30 percent of RBCs in two cases and in more than 60 percent of RBCs in the third case. Many RBCs contained multiple organisms. In each case, antimalarial therapy was begun, and an automated RBC exchange was performed emergently with a cell separator. Exchange transfusion was repeated within 24 hours for two patients. Parasitemia levels were less than 1 percent in all patients 24 hours after the last exchange. The neurologic status of these patients returned to baseline, and they were discharged 7 to 18 days after admission. CONCLUSION: Automated RBC exchange transfusion can rapidly reduce the level of parasitemia and restore neurologic functioning in patients with cerebral malaria. PMID- 10864993 TI - Investigation of porcine parvovirus among persons with hemophilia receiving Hyate:C porcine factor VIII concentrate. AB - BACKGROUND: Porcine clotting factor has been used for more than 15 years to treat severe bleeding episodes in persons with hemophilia who have antibodies to human clotting factor. In 1996, QC procedures revealed for the first time the presence of porcine parvovirus (PPV) in the product. This report describes an investigation to determine the extent of product contamination and to evaluate past recipients of porcine clotting factor (Hyate:C, Speywood Biopharm) for evidence of PPV infection. STUDY DESIGN AND METHODS: Stored specimens from 22 lots of previously released Hyate:C were tested for the presence of PPV DNA by PCR and nested PCR assays. Serum specimens from 98 recipients of Hyate:C and 24 controls who did not receive Hyate:C were tested for PPV antibodies by an immunofluorescence assay. RESULTS: PPV DNA was detected in product from 21 of the 22 lots of Hyate:C, primarily by nested PCR testing. In contrast, none of the serum specimens from the 98 Hyate:C recipients tested positive for PPV IgG antibodies. CONCLUSION: The risk of human disease from percutaneous exposure to low levels of PPV seems to be low. Nevertheless, the theoretical risk of human infection with PPV has led to manufacturing changes, including PCR screening of all porcine plasma, which are designed to eliminate this risk. PMID- 10864994 TI - Molecular analysis of HCV type 1 to 5 envelope gene: application to investigations of posttransfusion transmission of HCV. AB - BACKGROUND: Until 1990, HCV infection was common in transfused patients, resulting in more than 200,000 cases of posttransfusion hepatitis C in France alone. A molecular method that permits the investigation of posttransfusion hepatitis C infections is presented. STUDY DESIGN AND METHODS: Viral sequences in the envelope region of HCV were obtained for 12 pairs of blood recipients and their respective blood donors. The HCV strains studied belonged to types 1 (subtypes 1a and 1b), 2, 3, 4, and 5. Genetic distances and mutation rates were determined, and sequences were submitted to phylogenetic analysis along with sequences retrieved from nucleotide databases. RESULTS: Pairwise distances in the donor-recipient pairs were found to be less than 0.05 mutation per site, which corresponds to a mutation rate ranging from 0.6 x 10(-3) to 2.1 x 10(-3) per site per year. Sequences obtained from the 12 donor-recipient pairs clustered in 12 monophyletic nests. CONCLUSION: The genetic analysis of the envelope region of HCV can be used for the forensic evaluation of virus transmission. It permits the refutation of a link between blood transfusion and HCV transmission, rather than proof of the existence of such a link. PMID- 10864995 TI - Evaluation of a new PCR assay with competitive internal control sequence for blood donor screening. AB - BACKGROUND: High-throughput nucleic acid testing for transfusion-relevant viruses by PCR requires contamination-proof methods with high sensitivity and validity. A new PCR reagent kit (TaqMan, PE BioSystems) reduces the risk of carry-over contamination by eliminating post-PCR processing. STUDY DESIGN AND METHODS: Oligonucleotide design was done with software specialized for designing the assays' (TaqMan) primers and probes. A template-derived competitive internal control sequence designed through site-directed mutagenesis was used to reveal failures in amplification. Assay sensitivity was determined for single-donor and single-patient testing and by spiking sample mini-pools. Three seroconversion panels were tested. RESULTS: Sensitivity is high, reaching 300 HBV genomes per mL of single-patient material on direct testing. A detection limit of 1000 HBV genome equivalents per mL of donor plasma is achieved for 96 pooled samples. The window period for HBV infection was reduced by 17, 10, and 63 days from that for HBsAg screening in three seroconverting donors. CONCLUSION: The assay provides sufficient sensitivity to be superior to HBsAg screening in transfusion medicine and will be useful in clinical laboratories because of its ease of handling. PMID- 10864996 TI - Mother-to-infant transmission of GB virus type C/HGV. AB - BACKGROUND: The potentially hepatotropic flavivirus-like virus, GB virus type C (GBV-C)/HGV, has been detected in a few patients with acute and chronic hepatitis and in a certain proportion of blood donors and recipients of blood or blood components. STUDY DESIGN AND METHODS: Sera from 2979 pregnant Japanese women were examined for the presence of GBV-C/HGV RNA by nested RT-PCR. Mothers who were positive for viral RNA and their 34 infants were followed and tested for infection. RESULTS: Of the 2979 women, 32 (1.1%) were positive for GBV-C/HGV RNA. Twenty-six (76.5%) of 34 babies born to these women were positive for the virus when first tested. A significantly higher titer of viral RNA was observed in mothers whose infants were infected than in those whose infants were uninfected (mean +/- SD, 10(6.3 +/- 0.9) vs. 10(4.6 +/- 0.9)/mL; p<0.001). Twenty-three (96%) of 24 babies born to mothers whose serum viral titers were 10(6) mL or more were infected with the virus. Infants delivered by elective caesarean section had a lower risk (OR, 0.13; 95% CI, 0.02-0.82) than those delivered vaginally or by emergency caesarean section. No other risk factors for mother-to-infant transmission were confirmed. CONCLUSIONS: GBV-C/HGV is frequently transmitted from mothers to infants in the general population. The most critical factor is the titer of viral RNA in the maternal serum. By the use of elective caesarean section in women with high titers of viral RNA, vertical transmission of the virus may be lessened. PMID- 10864997 TI - Peripheral blood progenitor cells for HPC transplants involving unrelated donors. PMID- 10864998 TI - Summary of a workshop on in vivo efficacy of transfused platelet components and platelet substitutes. PMID- 10864999 TI - Universal WBC reduction. PMID- 10865000 TI - Should the FDA mandate that autologous units drawn and transfused within a single institution be tested for markers of infectious disease? PMID- 10865001 TI - Infectivity of buffy coat in variant CJD. PMID- 10865002 TI - The American Academy of Child Cardiology. PMID- 10865003 TI - Selected aspects of cardiac tumors in infancy and childhood. AB - Considerable literature concerning cardiac tumors in infancy and childhood has accumulated summarizing the prevalence, histologic types, clinical presentation and outcome, and changing imaging algorithms [1, 7, 10, 14, 20, 24, 33, 37, 43, 48, 57, 58, 60-62, 67, 69, 70, 90, 105, 106, 110, 124, 139, 140, 142, 143, 149]. In this review, we focus on selected aspects of cardiac tumors in the neonate, infant, and child, with an emphasis on imaging modalities [6, 13, 15, 18, 21-23, 60, 71, 77, 80, 92, 98, 99, 103, 107, 112, 114, 119, 146]. Various types of primary cardiac tumors in childhood are discussed in this article. PMID- 10865004 TI - Primary heart tumors in the pediatric age group: a review of salient pathologic features relevant for clinicians. AB - Because primary tumors of the heart in infants and children are extremely rare, most knowledge is based on collections of case reports rather than large cohort studies. The types of heart tumors encountered in the pediatric age group differ from those seen in adults. In the latter, cardiac myxomas are by far the most common tumor; in infants and children the most common primary tumor of the heart is the rhabdomyoma. Spontaneous regression of these tumors has been well established so that surgical intervention is no longer indicated unless there are clinical manifestations from the heart. There is a high incidence of associated tuberous sclerosis. Any intracavitary mass in infants is suggestive for a cardiac rhabdomyoma unless otherwise proven. The second most common tumor in this age group is cardiac fibroma. These tumors probably represent hamartomatous lesions and this has led to a strategy in which radical surgical excision is not indicated if the procedure endangers postoperative heart function. Cardiac myxomas are of interest in this age group because of their familial occurrence as part of the myxoma syndrome. Other types of primary heart tumors occur, including malignant variants, but all are exceedingly rare. It is because of the sporadic nature of these tumors that treatment and insights into prognosis are based largely on case documentations and analogies from similar tumors originating elsewhere. PMID- 10865006 TI - The impact of information technology on pediatric cardiology: present, past, and future. AB - Inexorable progress in information technology has been the driving force behind much of the progress that has been achieved in the fields of pediatric cardiology and cardiac surgery since the inception of these two subspecialities during the middle of the 20th century. This article outlines the influences of the digital age and speculates on likely future developments in these two subspecialities. PMID- 10865008 TI - Effects of electrocardiography and chest radiography on the accuracy of preliminary diagnosis of common congenital cardiac defects. AB - The objective of this study was to compare the accuracy of the expert clinical examination for certain common cardiac defects with and without electrocardiogram (EKG) and chest radiogram (x-ray). The design of the study was a prospective, blinded comparison of diagnostic accuracy of the expert examination with and without EKG and x-ray, using echocardiography as the diagnostic standard. The setting of the study was the pediatric cardiology outpatient department. There were 749 outpatients with heart murmur under 21 years of age without prior echocardiography or pediatric cardiology consultation. The intervention was echocardiography as clinically indicated for evaluation of heart murmur of uncertain cause. Measurements were carried out using the incorporation of EKG and x-ray into multiple linear regression models to assess independent associations, if any, with the accuracy of clinical examination. Results were reported as the presence or absence of independent significant impact of availability of EKG and x-ray on examiner's diagnostic accuracy for innocent murmur, ventricular septal defect (VSD), pulmonary stenosis (PS), aortic valve disease, atrial septal defect (ASD), and patent ductus arteriosus. EKG enhanced detection of ASD and may have helped detect PS. X-ray enhanced detection of intermediate to large VSD. X-ray and EKG were otherwise without demonstrable independent advantage for defect specific diagnosis. Routine use of one or both of these tests in the initial evaluation of heart murmur in the pediatric cardiology clinic should remain an option. PMID- 10865007 TI - Factors affecting the exercise capacity of pediatric patients with aortic regurgitation. AB - Although exercise testing is commonly employed to identify adult aortic regurgitation (AR) patients with early left ventricular (LV) dysfunction, the role and value of exercise testing in the management of pediatric AR patients have not been established. The purposes of this study were to evaluate the cardiorespiratory response to exercise of pediatric patients with chronic AR, examine the relation between exercise function and baseline echocardiographic measurements, and identify factors related to diminished exercise capacity (EC). The study group consisted of 26 patients aged 8 to 21 years (mean 14.4 +/- 3.7) with moderate or severe AR referred for exercise physiology testing. All patients underwent a baseline echocardiographic study and a symptom-limited, progressive cycle ergometer exercise test. LV diastolic dimension averaged 120 +/- 12% predicted, systolic dimension 112 +/- 20% predicted, shortening fraction 0.41 +/- 0.07, end-systolic wall stress 65 +/- 23 g/cm(2), and regurgitant fraction 38 +/- 16%. The average EC was 88 +/- 28% (56-143) predicted. No statistically significant correlation was found between EC and any of the echocardiographic parameters studied. Nine patients had EC < 75% predicted. These individuals did not differ from patients with higher EC with regard to any of the echocardiographic parameters or with regard to peak heart rate, blood pressure, respiratory exchange ratio, and incidence of ectopy or ST depression. However, the oxygen pulse at peak exercise (an index proportional to forward stroke volume at peak exercise) was significantly depressed among patients with EC < 75% predicted (77 +/- 6 vs. 106 +/- 16% predicted, p <. 0001). In conclusion, most pediatric patients with moderate or severe AR compensate well for their valve disease, maintain normal forward stroke volume during exercise, and have normal EC. However, a subset of AR patients have diminished EC secondary to an inability to augment forward stroke volume appropriately. These patients cannot be identified on the basis of resting echocardiographic studies. Timely identification of these patients, through formal exercise physiology testing, may have important clinical implications. PMID- 10865009 TI - Enteral nutritional support by percutaneous endoscopic gastrostomy in children with congenital heart disease. AB - One of the major problems of children with severe congenital heart disease (CHD) is their poor nutritional status. Among other consequences, it influences the surgical outcome. Retrospectively we present our experience with percutaneous endoscopic gastrostomy (PEG) in 15 children with CHD. This technique allows enteral nutritional support without the disadvantages related to long-term nasogastric tube feeding. Major complications were absent, and minor complications were rare both at PEG insertion, which was performed under deep sedation, and during feeding via PEG tube. In 4 of the 8 children who were followed for at least 6 months the age-matched body weight increased more than one standard deviation. In 2 other patients it increased more than 0.5 standard deviations. In 7 children the tube was removed after 2.5 to 42 months since enteral support was no longer necessary. Apart from initial reservations the parental acceptance of PEG was good. We conclude that the PEG is a safe and reliable technique to support enteral nutrition in children with severe CHD. PMID- 10865011 TI - Capillary whole blood monitoring of oral anticoagulants in children in outpatient clinics and the home setting. AB - A whole blood prothrombin time/international normalized ratio (PT/INR) monitor (CoaguChek, Roche Diagnostics Corp., Indianapolis, IN) was assessed in children for its accuracy, reliability, safety, and acceptance by health care personnel and patient's families. The PT/INR values measured by the CoaguChek monitor showed an excellent correlation with PT/INR values measured by the Hospital for Sick Children (HSC) laboratory (r = 0.96) and Hamilton Civic Hospitals Research Centre (HCHRC) laboratory (r = 0.92) in clinic patients and a close correlation with PT/INR values measured by the HSC laboratory (r = 0.76) and HCHRC laboratory (r = 0.74) in patients at home. Reduced correlation in the home setting did not adversely affect clinical management. The whole blood PT/INR monitor is safe and accurate for children requiring oral anticoagulation therapy in either the outpatient clinic or home setting. PMID- 10865012 TI - Centralization of pediatric heart surgery in Sweden. AB - In Sweden, which has a population of 8.9 million people, pediatric heart surgery was previously performed in four cities. After a long, difficult process, centralization of pediatric heart surgery to two centers was achieved in 1993. The overall 30-day mortality for open-heart surgery on infants and children of 9.5% before the centralization (1988-1991) was reduced to 1.9% in 1995-1997. A causal relationship between the mortality rates before and after the centralization is impossible to prove. Heart surgery was concentrated to the two centers with the lowest surgical mortality, and the reduction in surgical mortality was observed over a short period of time which makes it likely that the centralization of the surgical activity promoted the improved results. During the later time period the amount of more complex surgery was clearly increased compared to that performed previously. PMID- 10865013 TI - Doppler echocardiographic evaluation of right ventricular diastolic function in children. AB - Right ventricular diastolic function was evaluated by flow velocity pattern in the right ventricular inflow tract by means of pulsed Doppler echocardiography. Traditionally used to evaluate this function are peak velocities obtained during early diastole (peak E wave) and during atrial contraction (peak A wave), their ratio (peak E/A ratio), and the deceleration half-time. We conducted pulsed Doppler echocardiographic studies of right ventricular inflow and outflow patterns in 171 children (105 normal children and 66 children who were undergoing total surgical repair of congenital heart defects without sequelae). Results showed that summation flow was present in the right ventricular inflow tract in 43 (25%) of the 171 subjects, which made it difficult to separate the peak E wave from the peak A wave. We noted the presence of antegrade late diastolic flow (DW) in the right ventricular outflow tract of all subjects. DW, measured in 121 subjects in whom both E and A waves were detected in the right ventricular inflow tract, showed a highly significant correlation (p < 0.0001) with A waves in the right ventricular inflow tract. The ratio of DW to right ventricular outflow tract velocities during systole (SW) showed a highly significant (p < 0.0001) correlation with E/A ratio. When evaluating right ventricular diastolic function by pulsed Doppler, especially in children, the analysis of right ventricular outflow tract patterns is helpful in addition to that of inflow tract patterns. The DW and DW/SW ratio may present good alternatives to traditional parameters in children. PMID- 10865014 TI - Acute pericarditis in childhood: a 10-year experience. AB - Twenty children, aged 6 months to 13 years, with acute pericarditis admitted between 1987 and 1997 to a university hospital were analyzed retrospectively for their etiology, presentation, management, and prognosis. The most common types of pericarditis were purulent (40%), collagen vascular disease (30%), viral (20%), and neoplastic disease (10%). Most children presented with chest pain, fever, and tachypnea, but cardiac tamponade was not seen in any children. Staphylococcus aureus was the most frequent causative organism of purulent pericarditis and septic arthritis was the most common concurrent infection in the patients. Surgical drainage was performed for 11 cases, 9 underwent subxiphoid pericardial window, and 2 underwent thoracotomy. There was no constrictive pericarditis or reaccumulation of fluid after surgery. Two children died, one of staphylococcal septicemia and the other had a malignant mediastinal tumor. The remaining 18 made a complete recovery. We conclude that subxiphoid pericardial drainage is a simple, safe, and quick procedure and can be done easily in general hospitals by pediatric surgeons. The expensive facilities of cardiac surgeries are not needed. PMID- 10865015 TI - Diagnostic and interventional trends in tetralogy of fallot and transposition of the great arteries in a population-based study. AB - The goal of this study was to analyze diagnostic and interventional trends in tetralogy of Fallot and transposition of the great arteries in Malta for individuals born during 1920-1994 and calculate birth prevalences. The design was population based, in the setting of a regional hospital providing exclusive diagnostic and follow-up services for the entire population of Malta. Data collection and analysis was retrospective and included comparison with earlier epidemiological studies. Patients included were all Maltese live births diagnosed as having tetralogy of Fallot (TF) and transposition of the great arteries (TGA) born by the end of 1994. There were 109 cases of TF and 30 cases of TGA (n = 30). A significant decline in age at diagnosis and age at surgery was found for both lesions (p < 0.0001), which was associated with a significant decline in perioperative mortality (p = 0.005). Underascertainment was present prior to 1980, however, more cases were diagnosed due to improvement of postmortem services in the early 1980s followed by improvement of clinical services with eventually exclusive premortem diagnosis. The birth prevalence of TGA was 0.31/1000 live births, well within the range described in previous studies, unlike tetralogy of Fallot, which has been found in excess in this population in an earlier study. TF and TGA, the two lesions which comprise the majority of cyanotic congenital heart disease presenting in infancy, have been diagnosed and have undergone intervention at progressively earlier ages over the period under study. This decline was associated with a declining perioperative mortality. PMID- 10865017 TI - Left ventricular diastolic functions in juvenile rheumatoid arthritis. AB - Cardiac involvement as pericarditis, myocarditis, and endocarditis is common in juvenile rheumatoid arthritis (JRA). Though there are many reports concerning systolic and diastolic functions of adults with rheumatoid arthritis, there are no studies on children with JRA. Thirty patients with JRA without any cardiac symptoms and 30 sex- and age-matched controls were included in the study. M-mode and pulsed-wave Doppler echocardiography were performed on each participant to assess the systolic and diastolic functions of the left ventricle. Left ventricular end-systolic diameter and volume were larger and ejection fraction and fractional shortening were decreased in the JRA group. Among the diastolic parameters, increased late flow velocity, decreased early flow velocity, and prolonged isovolumic relaxation time reflected an abnormal relaxation form of diastolic dysfunction. Mortality rate is increased in adults with rheumatoid arthritis, and ischemic heart disease is the leading cause of cardiovascular mortality. The abnormal relaxation form of diastolic dysfunction found in children with JRA is seen in ischemic heart disease. These children can therefore be candidates for ischemic heart disease in the future even though they are fully asymptomatic at present. In conclusion, children with JRA should be assessed for systolic and diastolic functions with serial echocardiography. In this way it may be possible to reduce the mortality and morbidity of the disease from cardiac causes. PMID- 10865019 TI - Plasma nitric oxide products correlate with cardiac index of congenital heart disease. AB - We wished to determine the relationship between circulating levels of nitric oxide (NO) and cardiac index (CI) in children with congenital heart diseases. We measured the plasma levels of nitrate/nitrite (NO(x)), the stable end products of NO production as well as tumor necrosis factor-alpha (TNF-alpha), atrial natriuretic peptide (ANP), and brain natriuretic peptide in relation to various parameters determined simultaneously. The plasma NO(x) levels correlated negatively with CI (r = -0.541, p < 0.05). No correlation was observed between NO(x) and cardiac output. TNF-alpha correlated with NO(x) levels (r = 0.593, p < 0.005) but not with either CI or cardiac output. Plasma levels of ANP and TNF alpha were higher in atrial septal defect than those in the control group (p < 0.001 and p < 0.05, respectively). Elevated plasma NO(x) could explain the increased basal release of endothelial NO due to high pulmonary blood flow. Plasma NO(x) correlate negatively with CI in young patients with left-to-right shunt congenital heart diseases. PMID- 10865020 TI - Magnetic resonance imaging of unroofed coronary sinus: three cases. AB - Unroofed coronary sinus is a rare cardiac anomaly in which communication occurs between the coronary sinus and the left atrium due to the partial or complete absence of the roof of the coronary sinus. It is usually associated with other cardiovascular anomalies, especially persistent left superior vena cava. It is often not discovered during cardiac catheterization without clinical suspicion. We report three cases of unroofed coronary sinus which were incidentally detected by magnetic resonance imaging. PMID- 10865021 TI - Preoperative management of right ventricular dysfunction in aortic origin of the right pulmonary artery. PMID- 10865022 TI - Severe intrauterine growth retardation, aged facial appearance, and congenital heart disease in a newborn with Johanson-Blizzard syndrome. AB - We report a female newborn with Johanson-Blizzard syndrome associated with extreme intrauterine growth retardation, aged facial appearance, and atrial septal defect. Other features are microcephaly, prominent veins over the scalp, alopecia over the vertex, wide-open fontanelle, high forehead, antimongoloid slant, edematous eyelids, the absence of eyebrows and eyelashes, beaked nose with alae nasi, low-set ears, thin lips, and micrognathia. Investigations revealed deafness and congenital hypothyroidism. We believe that this association of severe intrauterine growth retardation and congenital heart disease represents the components of this syndrome. PMID- 10865024 TI - Prevention of recurrences of corticosteroid-dependent idiopathic pericarditis by colchicine in an adolescent patient. AB - A case of an adolescent patient with idiopathic recurrent pericarditis is reported. On several occasions during a period of 3. 5 years he was shown to be dependent on corticosteroid therapy, and he became cushingoid. After corticosteroids were substituted with colchicine, no further relapses occurred during a period of 29 months. In accordance with the first published pediatric results, colchicine represents an effective and well-tolerated alternative therapy for recurrent idiopathic pericarditis and might replace prolonged administration of corticosteroids. PMID- 10865023 TI - Cardiac troponin I in fulminant adenovirus myocarditis treated with a 24-hour infusion of high-dose intravenous immunoglobulin. AB - We report a successful outcome on an acute adenovirus myocarditis treated with a 24-hour high-dose intravenous immunoglobulin (24-HDIVIG) in a 4.5-year-old girl. A postviral etiology of acute myocarditis was assessed on the basis of the polymerase chain reaction technique. Among other early markers of cardiac injury, cardiac isoform of troponin-I (cTnI) was significantly correlated to the left ventricular ejection fraction (r = -0.86, p < 0.0001). Follow-up of cTnI, which might also be correlated to the short-term outcome, allows fast, easy, and noninvasive estimation of response to the aggressive treatment with 24-HDIVIG in acute adenovirus myocarditis in children. PMID- 10865025 TI - Thrombolysis of prosthetic tricuspid valve thrombosis with human recombinant tissue plasminogen activator in an adolescent. AB - Prosthetic heart valve thrombosis is associated with a high mortality. Traditionally, thrombectomy or valve replacement is performed. Thrombolysis offers a promising alternative to surgery. Usually, streptokinase and urokinase are the preferred agents for thrombolysis; however, human recombinant tissue plasminogen activator (rt-PA) is increasingly used. Thrombosis of prosthetic valves in children and adolescents is rare and experience of thrombolysis for obstructed valves is limited. We report the successful lysis of a thrombosed prosthetic tricuspid valve in an adolescent using rt-PA. The outcome of our patient supports the assumption that rt-PA represents an adequate therapeutic option for thrombolysis of obstructed prosthetic heart valves in children and adolescents. PMID- 10865027 TI - Can heart rate variability predict sudden death? A case of sudden death in a child with severe coronary sequelae of Kawasaki disease. AB - We report the heart rate variability (HRV) of a child aged 11 years. Arrhythmia was the suspected cause of sudden death after 10 years of therapy for Kawasaki disease. The linear methods failed to show any features of the HRV that could have predicted the patient's sudden death, but the fractal scaling as a nonlinear method had suddenly decreased from 5.3 to 4.1 1 year before his death. PMID- 10865026 TI - Intracardiac lymphoma in a child: successful treatment with chemotherapy alone. AB - A 5-year-old female child with history of non-Hodgkin's lymphoma presented with cough and palpitation. On physical examination a cardiac tumor plop was heard. Paroxysmal supraventricular tachycardia was noted on the electrocardiogram. Transthoracic echocardiogram revealed multiple large tumor masses within the right and the left atrium. The right atrial tumor was flopping back and forth at the tricuspid valve orifice. The tumor resolved completely with chemotherapy without any surgical intervention. PMID- 10865028 TI - Eternity! PMID- 10865029 TI - Balloon dilatation of pulmonary artery band. PMID- 10865031 TI - Thyroid gland and surgery of the thyroglossal duct: exercise in applied embryology. AB - Thyroglossal duct cysts (TDCs), the most common congenital cervical abnormality, originates from the medial anlage of the thyroid gland and presents as a painless asymptomatic midline suprahyoid mass. It does not represent a diagnostic challenge. The tract may persist as a fibrous cord or leave nests of cells anywhere along its embryonic path, and it gives rise to the development of TDC. The Sistrunk operation described in 1920 consists of en bloc cystectomy and central hyoidectomy, with tract excision up to the foramen cecum. This procedure remains an effective treatment for TDC. Malignant degeneration of TDC is rare (0.7%). PMID- 10865032 TI - Surgeon's approach to the thyroid gland: surgical anatomy and the importance of technique. AB - The cornerstone of safe and effective thyroid surgery is thorough training in and understanding of thyroid anatomy and pathology. With appropriate techniques, total thyroid lobectomy and total thyroidectomy (which should be considered simply as a bilateral total thyroid lobectomy performed during the same operation) can be undertaken with minimal risk of damage to the recurrent laryngeal nerves, the external branches of the superior laryngeal nerves, and the parathyroid glands. Safe surgery requires a specific operative plan, progressing in a series of logical, orderly, anatomically based steps. Exposure of the thyroid gland is followed by careful dissection of the superior pole, utilizing the avascular plane between the superior pole and the cricothyroid muscle to identify and preserve the external branch of the superior laryngeal nerve. Medial retraction of the gland then allows dissection of the lateral aspect of the thyroid lobe. Protection of the recurrent laryngeal nerves and preservation of the blood supply to the parathyroid glands is best achieved by "capsular dissection," ligating the tertiary branches of the inferior thyroid artery on the gland surface. If a parathyroid gland cannot be preserved or becomes ischemic after dissection of its vascular pedicle, it should be immediately minced and autotransplanted into the ipsilateral sternocleidomastoid muscle. The current evolution of outpatient or short-stay thyroidectomy emphasizes the need to avoid complications by utilizing meticulous surgical technique. Minimally invasive thyroidectomy utilizing endoscopic techniques may also affect the practice of thyroid surgery. Even so, understanding the surgical anatomy of the thyroid gland and its possible variations is paramount to safe and effective surgery. PMID- 10865033 TI - Overview of surgical pathology of the thyroid gland. AB - The purpose of this overview is to provide timely information on selected topics on the surgical pathology of the thyroid gland. Selected publications of the author and his colleagues at the University of Michigan and the Maine Medical Center form the basis of this review. Information provided in our reports is updated by perusal of recent, pertinent publications. The following questions summarize the contents of the overview. What is a "lateral aberrant thyroid"? Does it always represent metastatic carcinoma? What are dyshormogenetic goiters? Can "focal thyroiditis" in thyroid glands removed for the treatment of Graves' disease and toxic nodular goiters be predictive of the development of postoperative hypothyroidism? What is the pathology of autonomously functioning (hot) nodules? Do tall-cell, columnar-cell, and diffuse sclerosing types of papillary carcinomas forbode bad prognoses? What is the controversy over Hurthle cell tumors? Does the presence of a better differentiated component in an anaplastic thyroid carcinoma modify its biologic behavior? Are poorly differentiated carcinomas unique histologic variants? Does their histology affect prognosis? What is the most common small-cell tumor of the thyroid gland? Has the Chernobyl nuclear disaster affected the incidence of childhood thyroid carcinoma? Are these radiation-induced tumors more aggressive? The answers, some of which are controversial, are found in this overview. The aims here were to provide information to surgeons and pathologists and to improve the care of patients with thyroid disease. PMID- 10865034 TI - Expanding role of fine-needle aspiration cytology in thyroid diagnosis and management. AB - In non-iodine-deficient areas, 4% to 7% of the population are reported to have thyroid abnormalities. Prophylactic operations of these nodules in the thyroid are not indicated and not cost-effective, as at least four of five nodules are colloid goiter and only a few are malignant. The need for a reliable preoperative diagnosis is great, and fine-needle aspiration (FNA) is now considered the first choice during workup for thyroid nodules. The steps in the FNA procedure are clinical examination and localization of the target lesion, aspiration, preparation of smears, and collecting material for ancillary microscopy techniques. All these steps must be exercised to allow optimal use of FNA. It can also be combined with various other methods, such as immunohistochemistry of thyroglobulin and calcitonin, analysis of nuclear DNA, DNA preparation for molecular biology analyses, and magnetic resonance spectra. The accuracy of the clinical routine in our unit was evaluated by studying the 5-year outcomes of almost 4000 FNAs of the thyroid. The results were good, with only a few false negative and false-positive results; but the problem of differentiating follicular adenoma from follicular carcinoma remains a significant problem. It is now well established that FNA biopsy and cytology is the best modality available for the workup of thyroid nodules and is widely utilized in endocrine surgical centers worldwide. PMID- 10865035 TI - Growth regulation of thyroid and thyroid tumors in humans. AB - In a study of growth regulation of the human thyroid gland and thyroid tumors we investigated the impact of iodine and that of the thyroid-specific growth stimulating hormone TSH. Further studies included locally active growth factors such as the epidermal growth factor, insulin-like growth factor, and tissue transforming growth factors alpha and beta. In addition to studies of growth regulation by the various growth factors in mostly normal thyrocytes, the impact of tumor-specific mutations in oncogenes and tumor-suppressor genes was investigated. The results demonstrated distinct changes in tissue specificity and sensitivity to external stimuli. This rather complex view on thyrocyte growth regulation may be confusing, but it describes the biologic reality more precisely. Increased knowledge of the regulatory processes may lead to the development of new tumor- and patient-specific therapeutic approaches, especially for preventing benign goiter recurrence and for treating follicular and papillary thyroid cancers. PMID- 10865036 TI - Molecular genetics of thyroid tumors and surgical decision-making. AB - The study of thyroid tumor genetics has great relevance to surgeons and facilitates understanding tumor pathogenesis, prediction of tumor behavior, and management decisions. The genes implicated can be broadly categorized as oncogenes or tumor-suppressor genes. The RET oncogene has well established roles in the development of both papillary (PTC) and medullary (MTC) thyroid carcinoma. Genetic screening for germline RET mutations in members of multiple endocrine neoplasia type II (MEN-II) families is now widely performed, and prophylactic thyroidectomy in gene carriers is advisable at an early age. Patients with apparently sporadic MTC can also be screened to rule out familial disease. The demonstration of a RET rearrangement in a patient's PTC may have prognostic significance, but as yet there are no management implications. The thyrotropin receptor (TSH-R) and Gsalpha become oncogenic through point mutation and are associated with the development of toxic thyroid adenomas. The ras oncogene is implicated in the early stages of development of several thyroid tumor types. Tumor-suppressor genes also have a role in thyroid tumor formation. The p53 gene appears to be involved in the process of transformation to the anaplastic phenotype and the PTEN gene in the development of follicular adenomas but not carcinomas. There is still limited evidence for the so called adenoma-carcinoma sequence of the thyroid follicular cell. Loss of heterozygosity studies have enabled identification of tumor-suppressor genes, and their findings suggests differences in the pathogenesis of PTCs compared with follicular cancers. Surgical decision-making will benefit from these basic molecular advances, which rapidly translates into improved patient management. PMID- 10865037 TI - Thyroid nodules: rational management. AB - Thyroid nodules are the commonest disorder presenting to the endocrine surgeon. Most of the lesions are benign, but the principal problem facing the clinician is that of identifying the malignant nodule requiring surgery. Current diagnostic methods are reviewed, and the role of intraoperative frozen section in particular is examined in a series of 155 patients undergoing thyroidectomy for solitary thyroid nodule. It is concluded that when the fine-needle aspiration cytology (FNA) result is malignant intraoperative frozen section is unnecessary and contributes little to the management. Frozen section, however, is considered to be of value when the FNA result is reported as benign, suspicious, or inadequate. It permits identification of many malignant lesions that would otherwise require a second operation to complete a total thyroidectomy. Details of the indications for surgery and the operative strategy are discussed. PMID- 10865038 TI - Differentiated thyroid cancer: "complete" rational approach. AB - Controversy continues regarding the optimal management of patients with differentiated thyroid cancer because no prospective randomized studies evaluating the merits of (1) extent of thyroidectomy, (2) postoperative radioactive iodine ablation, or (3) thyroid-stimulating hormone (TSH) suppressive therapy exist. Patients with low risk differentiated thyroid cancer enjoy a relatively good prognosis with a mortality rate of about 2% to 5% and a recurrence rate of about 20%. Despite the excellent prognosis in patients considered to be at low risk, total or near-total thyroidectomy in patients with differentiated thyroid cancer has the advantages that: (1) postoperative radioactive iodine can be used to detect and treat residual normal thyroid tissue and local or distant metastases; (2) follow-up serum thyroglobulin levels are a more sensitive marker of persistent or recurrent disease when all thyroid tissue has been removed; and (3) total or near-total thyroidectomy with postoperative (131)I ablation and TSH suppressive therapy is associated with better survival and lower recurrence rates. Patients with occult papillary thyroid cancer and minimally invasive follicular thyroid cancer can be treated by thyroid lobectomy because they have a near-normal life expectancy. Virtually all other patients with differentiated thyroid cancer appear to benefit from more extensive initial treatment. PMID- 10865039 TI - Medullary carcinoma of the thyroid gland. AB - Medullary thyroid carcinoma (MTC) is an uncommon thyroid tumor that has attracted a great deal of interest because of its frequent presentation as a familial tumor and its primary involvement in the type II multiple endocrine neoplasia (MEN) syndromes MEN-IIA and MEN-IIB and familial medullary thyroid carcinoma (FMTC). The MTC tumor cells secrete the polypeptide hormone calcitonin, which serves as an excellent tumor marker, useful for defining the presence of disease, preoperatively or following thyroidectomy. The discovery that mutations in the RET proto-oncogene are associated with MEN-II syndromes was highly significant in that it demonstrated a clear correlation between genotype and phenotype; and most importantly it provided a mechanism whereby family members at risk could be identified by direct DNA analysis. Virtually all patients with MEN-IIA, MEN-IIB, and FMTC develop MTC; therefore there is a clear rationale for performing thyroidectomy as soon as a RET mutation has been identified. Because MTC appears to be much more aggressive in patients with MEN-IIB, thyroidectomy is performed during the first year of life in this setting, whereas in patients with MEN-IIA, where the tumor appears to be more indolent, the procedure can be safety delayed until age 5 years. Reoperative neck exploration in patients with evidence of persistent or recurrent MTC has been effective in a significant number of patients, although the success of the operation requires careful patient selection and preoperative assessment. MTC, as expressed in the MEN-II syndromes, is an excellent model to evaluate the usefulness of interventional therapy in patients demonstrated to have a genetic predisposition for cancer. PMID- 10865040 TI - Management of hyperthyroidism due to Graves' and nodular diseases. AB - Endocrine surgeons have had an important role in the management of hyperthyroidism due to either Graves' disease or toxic nodule(s). Since alternative treatments such as antithyroid drugs or radioiodine are also available, the decision-making for management should be based on clear assessment of advantages and limitations of each of the treatment options. Surgery provides rapid resolution of these conditions, and cure rates are high, although it may be associated with perioperative complications and postoperative thyroid dysfunction. The authors' experience in the surgical treatment of hyperthyroidism and a review of the recent literature are outlined in this report. PMID- 10865041 TI - Total thyroidectomy for management of thyroid disease. AB - Total thyroidectomy is a logical treatment for many patients with thyroid disease, including patients in whom the pathologic process requiring surgery involves both lobes of the thyroid or the risk of recurrence is a significant consideration, as in benign multinodular goiter, Graves' disease, and cancer. In earlier times the risks of extensive surgery and problems of adequate hormone replacement deterred surgeons from performing total thyroidectomy. However, as we enter the twenty-first century we are confident that the technical aspects of safe total thyroidectomy are established and that thyroid hormone replacement and monitoring are readily available and accurate. In the future total thyroidectomy is likely to be performed increasingly commonly for both benign and malignant disease. PMID- 10865042 TI - Hashimoto's disease and thyroid lymphoma: role of the surgeon. AB - With the turn of the century, the role of the surgeon in the treatment of diseases such as Hashimoto's and thyroid lymphoma has diminished. That is not to say that the surgeon must not have a thorough understanding of these diseases and the role he or she plays in their diagnosis and treatment. Hashimoto's disease is a common disease. Not infrequently the endocrine surgeon is faced with a thyroid nodule in a background of Hashimoto's disease. Interpretation of fine-needle aspiration (FNA) of a nodule in a patient with the background of Hashimoto's disease may be misleading if the surgeon fails to understand the limitations of FNA. The role of the surgeon in the treatment and diagnosis of thyroid lymphomas has evolved from surgical debulking to open biopsy. With the use of irradiation and chemotherapy, the need for surgical debulking has nearly disappeared. The recent development of ancillary techniques such as light chain restriction, flow cytometry, gene rearrangement, and immunohistochemical staining have enabled cytopathologists to diagnose thyroid lymphoma by FNA, further diminishing the surgeon's role in the diagnosis and treatment of this disease. PMID- 10865043 TI - Complications of thyroid surgery: how to avoid them, how to manage them, and observations on their possible effect on the whole patient. AB - Surgery of the thyroid takes place in an area of complicated anatomy and in which a number of vital physiologic functions and special senses are controlled. Thyroidectomy rarely is associated with mortality; but unless the surgeon performing it is well trained in operative surgery and is knowledgeable of the gland and its function, pathology, and anatomy, excellent results cannot be achieved. Failure to observe cardinal surgical principles may result in legal difficulties, which can be avoided. It is well to observe the principles and avoid problems. We address this issue herein. PMID- 10865044 TI - Future of thyroid surgery and training surgeons to meet the expectations of 2000 and beyond. AB - What is the future of thyroid surgery in the new millennium? How can surgeons keep abreast of advances in thyroid endocrinology, genetics, surgical therapy, and other aspects of thyroid disease management? How should surgeons be trained to become highly competent in thyroid disease and to perform safe, effective thyroid operative procedures? Nine internationally recognized endocrine surgeons were asked to express their views on these and related subjects. They noted that advances in molecular biology, pathology, and genetics of thyroid disease should allow more tailored surgical approaches during the twenty-first century. Current training of general surgical residents in thyroid and other types of endocrine surgery is highly variable, which may contribute to increased complication rates and number of second operations. The leadership for addressing these deficiencies and promoting a more organized approach to thyroid disease management should come from national endocrine surgery associations and their leaders. It is incumbent upon endocrine surgeons to maintain their central role in the management of many aspects of thyroid disease. Organizing teams of specialists into thyroid centers (centers of excellence) can (1) increase efficiency; (2) increase quality of care; (3) decrease costs; (4) encourage a more individualized approach to surgery; (5) lower complication rates; and (6) foster innovation in technology and disease management. Two years of additional fellowship training in thyroid and endocrine surgery is now being advocated by increasing numbers of national endocrine surgical associations as the best way to prepare surgeons for society's needs for highly skilled, competent thyroid surgeons of the future. PMID- 10865045 TI - Value of contralateral surveillance mammography for primary breast cancer follow up. AB - Mammographic screening of the contralateral breast is often advocated during follow-up of women previously treated for primary operable breast cancer. The purpose of this study was to determine the value of this investigation. Between 1987 and 1995 a total of 5102 contralateral screening mammograms were performed biennially on 2511 women aged 50% of regional adults, broadly distributed by site, gender, and age. INTERVENTIONS: From 1974 to 1994, a community program, integrated with primary medical care and staffed by professional nurses, provided education, screening, counseling, referral, tracking, and follow-up for cardiovascular risk factors. MAIN OUTCOME MEASURES: Age-adjusted mortality rates (total, heart, coronary, cerebrovascular, cancer) for three counties and Maine, plus annual program encounters. RESULTS: Relative to Maine, the Franklin heart disease death rate was 0.97 at baseline (1960-1969; 95% confidence interval, 0.91 to 1.03), 0.91 during the program (0.85 to 0.97), 0.83 during the 11 years of program growth (0.78 to 0.88), but 1.0 during the 10 years of decreasing encounters. Franklin's total death rate was 1.01 at baseline, 0.95 during the program (0.92 to 0.98), and 0.90 during program growth (0.86 to 0. 94). Results were similar for coronary disease, stroke, and cancer. Relative death rates did not fall in either comparison county. Nurse-client encounters totaled 120,280 over 21 years. Relative to Maine, heart disease death rates correlated inversely with program encounters (r = -0.53) but not with unemployment or physician supply. CONCLUSIONS: Integrated with primary medical care, a comprehensive, nurse mediated community cardiovascular health program in rural Maine has been associated with significant time-dependent and dose-dependent reductions in cardiovascular and total mortality. PMID- 10865162 TI - Promoting mammography: results of a randomized trial of telephone counseling and a medical practice intervention. AB - BACKGROUND: Despite widespread promotion of mammography screening, a distinct minority of women have remained underusers of this effective preventive measure. We sought to measure the effects of barrier-specific telephone counseling (BSTC) and a physician-based educational intervention (MD-ED) on mammography utilization among underusers of mammography screening. DESIGN: This was a randomized controlled trial. Women meeting criteria for mammography underuse at baseline (grouped by practice affiliation) were randomized to a reminder control condition (RC group received annual mailed reminders), BSTC or MD-ED interventions and followed for 3 years. Underuse was defined by failure to get two annual or biannual mammograms over a 2- to 4-year period prior to a baseline survey. PARTICIPANTS AND SETTING: The study included 1655 female underusers of mammography aged 50-80 years who were members of two health maintenance organizations (HMO) in central Massachusetts. INTERVENTIONS: BSTC consisted of periodic brief, scripted calls from trained counselors to women who had not had a mammogram in the preceding 15 months. Women could receive up to three annual calls during the study. MD-ED consisted of physician and office staff trainings aimed at improving counseling skills and office reminder systems. MAIN OUTCOME MEASURE: Self-report of mammography use during the study period was the main outcome measure. Regular use was defined as > or =1 mammogram every 24 months. RESULTS: Forty-four percent in each intervention group became regular users compared to 42% in the RC group. Among subjects who had prior but not recent mammograms at baseline, BSTC was effective (OR=1.48; 95% CI=1.04; 2. 10), and MD ED marginally effective (OR=1.28; 95% CI=0.88, 1.85). Most recent users at baseline and few never users became regular users (61% and 17%, respectively) regardless of intervention status. CONCLUSIONS: Among mammography underusers BSTC modestly increases utilization for former users at a reasonable cost ($726 per additional regular user). PMID- 10865163 TI - Social networks and cancer screening in four U.S. Hispanic groups. AB - BACKGROUND: Evidence shows that social relationships play an important role in health and health behavior. We examined the relationship between social networks and cancer screening among four U.S. Hispanic groups. METHODS: We used telephone surveys to collect data in eight U.S. regions that have concentrations of diverse Hispanic-origin populations. We interviewed 8903 Hispanic adults, for a response rate of 83%; analysis was restricted to the 2383 women aged > or =40. As a measure of social integration, we formed a social network index from items on the number of close relatives and friends, frequency of contact, and church membership. We used logistic regression to estimate the effects of social integration on screening, adjusting for sociodemographic factors. RESULTS: Among Mexican, Cuban, and Central-American women, the effect of social integration on mammography screening was slight. The odds ratios (OR) per unit change in social integration category ranged from 1.16 to 1.22 with confidence intervals (CI) that overlapped with the null. For Pap smear screening, the effect was strongest among Mexican-American women (OR=1.44, 95% CI=1.21 to 1.72), but also evident among Central-American women (OR=1.22, 95% CI=0.72 to 2.06) and Cuban women (OR = 1.25, 95% CI = 0.81 to 1.93). Among Puerto Rican women, social integration had no effect on either mammography (OR=1.03) or Pap smear screening (OR=1.08). CONCLUSIONS: Independent of socioeconomic factors, social integration appears to influence cancer screening participation of Hispanic women. The modest effect is not universal across Hispanic groups and was stronger for Pap smear than for mammography screening behavior. Researchers should recognize Hispanic group differences in social network characteristics and the potential of social networks to change screening behavior. PMID- 10865164 TI - Cancer education among primary care physicians in an underserved community. AB - INTRODUCTION: Urban minority groups, such as those living in north Manhattan, are generally underserved with regard to cancer prevention and screening practices. Primary care physicians are in a critical position to counsel their patients on these subjects and to order screening tests for their patients. METHODS: Eighty four primary care physicians in two intervention communities who received educational visits about cancer screening and prevention were compared with 38 physicians in a nearby community who received no intervention. With pre- and post test interviews over an 18-month period, the physicians were asked about their attitudes toward, knowledge of (relative to American Cancer Society guidelines), and likelihood of counseling and screening for breast, cervical, colorectal, and prostate cancers. RESULTS: Comparison of the two surveys of physicians indicated no statistically significant differences in knowledge of cancer prevention or screening. At post-test, however, intervention group physicians identified significantly fewer barriers to practice than control physicians (p<0.05). While overall, the educational visits to inner-city primary care physicians did not appear to significantly alter cancer prevention practices, there was a positive dose-response relationship among the subgroup of participants who received three or more project contacts. CONCLUSIONS: We uncovered significant changes in attitude due to academic detailing among urban primary care physicians practicing in north Manhattan. A significant pre-test sensitization effect and small numbers may have masked overall changes in cancer prevention and screening behaviors among physicians due to the intervention. PMID- 10865166 TI - Injury-prevention counseling among residents of internal medicine. AB - BACKGROUND: The U.S. Preventive Service Task Force's Guide to Clinical Preventive Services and Healthy People 2000 recommend that physicians participate in various counseling activities, including injury prevention. Despite recommendations, rates of physician counseling, particularly injury prevention, are low. This study assessed clinical preventive services and attitudes among physicians. Furthermore, the study illustrates how physicians prioritize injury-prevention counseling relative to other prevention recommendations. METHODS: Personal characteristics (i.e., demographics, specialty orientation, attitudes toward prevention, and personal health behaviors) of the residents were collected by a self-administered survey. We performed a 12-month retrospective chart review of 184 new doctor-patient encounters to determine rates of clinical preventive services that included four injury-prevention services: the use of seatbelts, helmets, and smoke detectors; and the safe storage of firearms. RESULTS: Overall, attitudes toward injury prevention in the context of other clinical preventive services were low. Seatbelt counseling was the only injury-prevention service documented in the charts, and was performed at only one of four clinic sites. CONCLUSIONS: Clinic site as a key predictor of preventive practice may be suggestive of the importance of organizational priorities and professional norms. Future injury-prevention education efforts must aim at improving attitudes of current and future physicians to facilitate positive professional norms. PMID- 10865165 TI - Cancer screening and prevention practices of inner-city physicians. AB - INTRODUCTION: Effective preventive services are needed most in underserved, inner city settings that suffer disproportionately from morbidity and mortality. Primary care physicians can play an important role in the provision of efficacious cancer prevention and screening services to patients in these communities. METHOD: We surveyed 122 primary care physicians about their cancer prevention and screening knowledge, attitudes, and practices. RESULTS: Relative to the findings from national and local surveys, sample physicians were not as knowledgeable about national guidelines for preventive care, were less likely to counsel on smoking cessation, and were less likely to advise diet modification. Although physician practices reflected national cancer prevention and screening guidelines in general, a significant proportion of physicians suggested lung and prostate cancer screening tests that were inconsistent with national recommendations. CONCLUSIONS: Systematic efforts are needed to increase the knowledge and practices of inner-city physicians concerning cancer prevention and screening. PMID- 10865167 TI - The impact of computer-generated messages on childhood immunization coverage(2)(2) AB - Introduction: Recent evaluations of computer-generated reminder/recall messages have suggested that they are an inexpensive, labor-saving method of improving office visitation rates of childhood immunization providers. This study assesses the sustained impact of computer-generated messages on immunization coverage during the first two years of life.Design: Randomized, controlled trial.Setting: County health department in the Denver metropolitan area.Study Participants: Children (n = 1227) 60 to 90 days of age who had received the first dose of diphtheria-tetanus-pertussis (DTP) and/or poliovirus vaccines.Intervention: Households of children were randomized into four groups to receive: telephone messages followed by letters (Group A); telephone messages alone (Group B); letters only (Group C); or no notification (Group D). Households in the intervention groups (A, B, and C) received up to five computer-generated telephone messages and/or up to four letters each time their children became due for immunization(s).Main Outcome Measure: Immunization series completion at 24 months of age.Results: Children whose families were randomized to receive any of the interventions were 21% more likely to have completed the immunization series by 24 months of age than were children randomized into the control group (49.2% vs 40.9%; RR [rate ratio] = 1.21; CI [confidence interval] =1.01, 1.44). While not statistically significant, children in Group A were 23% more likely to complete their immunization series by 24 months of age than those in the control group (50.2% vs 40.9%; RR = 1.23; CI = 1.00, 1.52). No differences were detected among the intervention groups. The costs per additional child completing the series by 24 months of age in Group A was $226 ($79 after start-up costs were discounted). Conclusion: Computer-generated contacts, either by phone or by mail (or both combined), used each time vaccines become due, are efficacious in increasing immunization coverage of children under 2 years of age. PMID- 10865168 TI - Cytokine cascade in dengue hemorrhagic fever: implications for pathogenesis. AB - Dengue virus produces a mild acute febrile illness, dengue fever (DF) and a severe illness, dengue hemorrhagic fever (DHF). The characteristic feature of DHF is increased capillary permeability leading to extensive plasma leakage in serous cavities resulting in shock. The pathogenesis of DHF is not fully understood. This paper presents a cascade of cytokines, that in our view, may lead to DHF. The main feature is the early generation of a unique cytokine, human cytotoxic factor (hCF) that initiates a series of events leading to a shift from Th1-type response in mild illness to a Th2-type response resulting in severe DHF. The shift from Th1 to Th2 is regulated by the relative levels of interferon-gamma and interleukin (IL)-10 and between IL-12 and transforming growth factor-beta, which showed an inverse relationship in patients with DF. PMID- 10865169 TI - Prevalence of de-O-acetylated serogroup C meningococci before the introduction of meningococcal serogroup C conjugate vaccines in the United Kingdom. AB - Meningococcal serogroup C conjugate (MCC) vaccines have been introduced in the UK to combat the rise in serogroup C meningococcal disease. Serogroup C meningococci may occur naturally expressing either O-acetylated (Oac(+)) or de-O-acetylated (Oac(-)) polysaccharide capsules. In a small study in the USA in the 1970s 15% of serogroup C meningococcal case isolates were reported to be Oac(-) though the prevalence of these Oac(-) isolates has not been recorded in the UK. This is of interest as the first MCC vaccines to be introduced are Oac(+) and the potential impact of this on Oac(-) serogroup C isolates is unclear. Serogroup C isolates submitted to the Public Health Laboratory Service Meningococcal Reference Unit in January 1998 (n=113) and January 1999 (n=162) were investigated by dot blotting using monoclonals specific for Oac(+) and Oac(-) serogroup C polysaccharides. This revealed 12% Oac(-) isolates for both January 1998 and January 1999. The proportion of fatal cases was found to similar for both Oac(-) and Oac(+), 14 and 9% for 1998 and 5 and 3% for 1999, indicating that the pathogenic potential of these Oac(-) isolates is similar to Oac(+). The acetylation status of serogroup C isolates needs to be monitored throughout and after the introduction of MCC vaccines. PMID- 10865170 TI - Modulation of lymphocyte proliferative responses by a canine Lyme disease vaccine of recombinant outer surface protein A (OspA). AB - The modulation of human lymphocyte proliferative responses was demonstrated with a recombinant outer surface protein A (OspA) vaccine preparation for the prevention of Borrelia burgdorferi infection. After exposure to either the unaltered vaccine preparation or OspA prepared in saline, normal lymphocyte responses to the mitogens concanavalin A, phytohemagglutinin-M or pokeweed mitogen, or the antigen BCG were consistently reduced. Whole cell extracts of B. burgdorferi also modulated immune responses but required a much greater quantity of protein than needed for the OspA preparation. The magnitude of modulation was directly dependent on the quantity of OspA. OspA interferes with the response of lymphocytes to proliferative stimuli including a blocking of cell cycle phase progression. Future studies designed to delete the particular region or component of the OspA molecule responsible for this effect may lead to improved vaccine preparations. PMID- 10865171 TI - Ornithine-containing lipids stimulate CD14-dependent TNF-alpha production from murine macrophage-like J774.1 and RAW 264.7 cells. AB - The ornithine-containing lipids (OL)-induced cytokine production pattern in macrophage-like J774.1 and RAW 264.7 cells was different from that in the peritoneal macrophages previously reported. OLs, as well as lipopolysaccharide (LPS) of Escherichia coli, strongly induced tumor necrosis factor (TNF) alpha but not interleukin (IL)-1beta in J774.1 cells. In the RAW cells, IL-1beta, TNF-alpha and prostaglandin E(2) were strongly induced by the OLs and LPS. OL- and serine glycine-containing lipid (SGL)-induced TNF-alpha production in J774.1 and RAW 264.7 cells required serum. However, in CD14-deficient LR-9 cells, TNF-alpha was not induced by the OLs in the presence or absence of serum. OLs and a SGL almost completely inhibited the binding of (125)I-LPS to J774.1 cells. These results suggested that OLs and SGL activate macrophages via the CD14-dependent pathway. PMID- 10865172 TI - Modulation of chemotaxis, O(2)(-) production and myeloperoxidase release from human polymorphonuclear leukocytes by the ornithine-containing lipid and the serineglycine-containing lipid of Flavobacterium. AB - The ornithine-containing lipid (OL) and the serineglycine-containing lipid (SGL) of Flavobacterium activated and modulated the functions of human polymorphonuclear leukocytes (PMNs). The OL and the SGL strongly activated fMet Leu-Phe- and interleukin-8-induced chemotaxis of PMNs at the concentration of 0.1 microg ml(-1), and a synthetic OL also activated the function of PMNs. Further, the OL strongly activated O(2)(-) production from PMNs. Although the OL and the SGL slightly modulated myeloperoxidase release from PMNs, inhibition effects of their component fatty acid analogues were observed. O(2)(-) production-inducing activity is a common biological activity between the OL and bacterial lipopolysaccharides, but OL and SGL, unlike lipopolysaccharide, are potent activators of PMN chemotaxis. PMID- 10865173 TI - Expression of staphylococcal protein Sbi is induced by human IgG. AB - Protein Sbi is an IgG- and beta(2) glycoprotein I-binding protein on the surface of Staphylococcus aureus. In most strains, the amount of protein Sbi on the cell surface is very low under normal growth conditions. However, here we show that after growth in the presence of human serum, the amount is significantly increased. In S. aureus strain 8325-4, the observed increase is concentration dependent and the highest level is found approximately 2 h after serum addition. The active molecule in serum was found to be IgG, which causes an increase of surface-located protein Sbi in S. aureus strain 8325-4, in the clinical isolates tested as well as in protein A-negative mutants. Thus, the results suggest that binding of IgG to protein Sbi upregulates protein Sbi synthesis. PMID- 10865174 TI - Epitope mapping of heat shock protein 60 (GroEL) from Porphyromonas gingivalis. AB - Porphyromonas gingivalis, a putative pathogen in human periodontal disease, possesses a 60-kDa heat shock protein (hsp60, GroEL). The GroEL homologs are known to be key molecules in auto-immune reactions because of the sequence similarity with human hsp60. In this study, B-cell epitopes on P. gingivalis GroEL (PgGroEL) were analyzed by both Western immunoblotting with truncated PgGroEL and by the multi-pin synthetic peptide approach. To examine auto-antibody production in periodontitis patients, Western immunoblotting with human gingival fibroblasts was performed. Deletion mutants were constructed from the cloned PgGroEL gene (P. gingivalis groEL), and four C-terminal truncated PgGroEL and one N-terminal truncated PgGroEL were prepared from the deletants. Sera from periodontitis patients reacted with all truncated PgGroEL used in this study. The results suggest that the B-cell epitopes were overlaid throughout PgGroEL. To determine the detailed locations of the B-cell epitope, 84 decapeptides covering the entire PgGroEL were synthesized and the serum IgG response to the peptides was examined. Epitope mapping using the synthetic peptides confirmed that the B cell epitopes were overlaid throughout the length of PgGroEL and revealed that highly conserved peptides between PgGroEL and human hsp60 were recognized by the serum antibodies. Immuno-reactivity against human gingival fibroblasts was examined with sera from 30 periodontitis patients and 10 periodontally healthy subjects. IgG antibody against the 65-kDa antigen in human gingival fibroblasts (same molecular mass as human hsp60) was detected in two patients. Although IgG production against human hsp60 may be rare case in periodontitis patients, the results of epitope mapping demonstrated the potential of PgGroEL to cause the cross-reactions with human hsp60. PMID- 10865175 TI - Typing of clinical and environmental Aeromonas veronii strains based on the 16S 23S rDNA spacers. AB - Genetic relationships of Aeromonas veronii strains isolated from human and environmental sources were investigated by restriction fragment length polymorphism (RFLP) of the polymerase chain reaction-amplified intergenic spacer region (ISR) flanked by the 16S and 23S rRNA genes. When using endonucleases AluI, HinfI and CfoI the 16S-23S rDNA-RFLP patterns showed considerable overall similarity, although most strains yielded specific profiles. Several intra specific lines of descent comprised clinical strains linked to isolates from environmental sources. Strains having identical patterns may be individuals derived from highly similar, if not the same, microorganism. Results suggest that the ISR sequence-based method can be used to demonstrate colonization of a public water supply with a particular microorganism. In addition it could be very useful for tracing recurrent episodes of diarrhea and Aeromonas infection outbreaks. PMID- 10865176 TI - The cell-mediated immune response to Borrelia afzelii, garinii and burgdorferi in C57BL/6 mice is dependent on antigen specificity. AB - Inbred C57BL/6 mice were challenged with Borrelia afzelii, Borrelia garinii and Borrelia burgdorferi sensu stricto and tested for antigen-specific T-cell response in vitro. The sonicated preparations of in vitro grown spirochetes were capable of stimulating polyclonal proliferation and specific cell-mediated response, depending on duration of the cell culture. Murine splenocytes previously sensitized to B. burgdorferi sensu lato (s.l. ), but not those from control mice, could be induced for antigen-specific proliferation in vitro. Moreover, detectable cell-mediated response could be induced only with antigen preparations derived from a corresponding strain but not with those obtained from other Borrelia genospecies as revealed by the [(3)H]thymidine incorporation assay. The current study considers that the strict B. burgdorferi s.l. antigen specific response may also be expected in infections in humans and contributes to the explanation of the frequently poor antibody- and cell-mediated immune response observed in patients diagnosed with Lyme disease. PMID- 10865177 TI - Decreased virulence of a strain of Pseudomonas aeruginosa O12 overexpressing a chromosomal type 1 beta-lactamase could be due to reduced expression of cell-to cell signaling dependent virulence factors. AB - Pseudomonas aeruginosa produces a large variety of virulence factors and is characterized by its capacity to rapidly develop resistance when exposed to antibiotics. In order to evaluate a possible correlation between acquired resistance to antibiotics and virulence, we examined the virulence of four isogenic variants of P. aeruginosa O12 that differ in their resistance phenotypes to various beta-lactam antibiotics in a mouse model of acute pneumonia. Strains overproducing a chromosomal type 1 beta-lactamase were less virulent in both immunocompetent and immunosuppressed animals. Whereas the production of the exopolysaccharide alginate was similar between the four strains, extracellular virulence factors (elastase, rhamnolipid) that are controlled by the cell-to-cell signaling system circuit were detected in reduced amounts in the supernatant of the two isolates overproducing type 1 beta-lactamase. These results suggest that strains overexpressing the chromosomal type 1 beta-lactamase could be less virulent because of a reduction of cell-to-cell signaling dependent virulence factor production. PMID- 10865179 TI - Bacterial, viral and parasitic enteric pathogens associated with acute diarrhea in hospitalized children from northern Jordan. AB - To determine the etiology of acute diarrhea in Jordanian children under 5 years of age, we examined stool samples from 265 children admitted to the pediatric ward at Princess Rahma Hospital for Children, Irbid, Jordan, for parasites, rotavirus and enteric bacteria. Using both traditional and molecular diagnostic techniques, we detected enteropathogens in 66.4% of patients with diarrhea. A single enteric pathogen was detected in 50.9% of the children, and multiple pathogens were detected in 15.5%. The prevalence of enteropathogens identified was as follows: rotavirus (32.5%), enteropathogenic Escherichia coli (12.8%), enteroaggregative E. coli (10.2), enterotoxigenic E. coli (5.7%), Shigella spp. (4.9%), Entamoeba histolytica (4.9%), Salmonella spp. (4.5%), Campylobacter jejuni/coli (1.5%), Cryptosporidium spp. (1.5%), enteroinvasive E. coli (1.5%), eae-, Ehly-positive E. coli (0.8%), Giardia lamblia (0. 8%) and Yersinia enterocolitica (0.4%). No Vibrio cholerae, Shiga toxin-producing E. coli, microsporidia, adenovirus or small round virus were detected. Findings from this study demonstrate that rotavirus and several types of diarrheagenic E. coli, which are not screened for during routine examinations of stool samples in public health laboratories, were the most frequently detected enteropathogens in these children. Our findings highlight the value of using a combination of traditional and molecular techniques in the diagnosis of diarrheal disease in this population. PMID- 10865178 TI - Protein phosphorylation pathways involved during lipopolysaccharide-induced expression of CD14 in mouse bone marrow granulocytes. AB - Lipopolysaccharide (LPS) of Gram-negative bacteria interacts with a CD14 independent receptor of mouse bone marrow granulocytes (BMC), and triggers in these cells the expression of CD14, an inducible type of LPS receptor (iLpsR). This particular response of BMC to LPS required the activation of protein tyrosine kinase and p38 MAP kinase. The inhibition of the LPS effect by the MEK inhibitor PD-98059 suggested that the ERK pathway was also involved. Unexpectedly, protein kinase C, myosin light chain kinase, cAMP-, cGMP-, and Ca(2+)/calmodulin-dependent kinases, as well as ecto-protein kinases, were not required for iLpsR expression. However, other yet unidentified serine/threonine protein kinase(s) were implied since the BMC response to LPS was markedly reduced after exposure to three inhibitors of such kinases (K-252a, H-7, and KT-5823). The atypical kinase requirements observed in this study may be due either to a novel signaling LPS receptor complex present in BMC, or to the particular events involved in CD14 biosynthesis. PMID- 10865180 TI - President's page PMID- 10865181 TI - Tissue-level signaling and control of uterine contractility: the action potential calcium wave hypothesis. AB - This article describes the action potential-calcium wave hypothesis of uterine contractility. Two known mechanisms of intercellular tissue-level signaling are merged into a single hypothesis of organ-level signaling. This hypothesis provides a framework with which to link cellular physiology with organ function. The two mechanisms of tissue-level signaling considered are action potential propagation and intercellular calcium waves. A great body of literature exists regarding the electrical excitability of smooth muscle and myometrium. Despite this knowledge, it does not seem possible to reconcile the familiar uterine contraction profile with known parameters of cellular physiology unless a second mechanism of intercellular communication is postulated. Intercellular calcium waves fit the requirements needed for the second mechanism: slow speed, ability to raise intracellular free calcium, and ability to signal over hundreds of micrometers. The premise of the action potential-calcium wave hypothesis is that action potentials propagate rapidly throughout the uterus, initiating intercellular calcium waves. As the intercellular calcium waves propagate slowly through the bundles, myocytes are recruited to participate in the contraction. This article reviews and summarizes the literature on calcium waves in human myometrium and presents evidence to support the combination of these mechanisms. Extension of the hypothesis suggests that the functional unit of the laboring human uterus is the smooth-muscle bundle, and that the frequency and strength of uterine contractions are separate but linked physiologic characteristics of labor. PMID- 10865182 TI - Expression of receptors for corticotropin-releasing factor in the vasculature of pregnant rats. AB - OBJECTIVE: To identify and localize the receptor(s) responsible for modulating vascular effects of corticotropin-releasing factor (CRF) during pregnancy. METHODS: Reverse transcriptase-polymerase chain reaction (RT-PCR), competitive RT PCR, and Western blot analyses were used to study the expression of CRF receptors (CRFR(1), CRFR(2alpha), and CRFR(2beta)) in the aorta and uterine vascular bed of nonpregnant and late (day 18) and term pregnant (day 22) Sprague-Dawley rats. Immunohistochemistry was done to localize the CRF receptor in the aortic wall. There were six rats in each study group. RESULTS: Only CRFR(2beta) was identified in the aorta and uterine vascular bed by RT-PCR and Western blot analyses. The PCR product was sequenced to confirm its identity. Competitive RT-PCR and Western blot analyses showed that expression of CRFR(2beta) is not different in late pregnancy compared with the nonpregnant but is decreased at term. Immunohistochemistry showed high expression of CRFR(2beta) on the aortic endothelial surface but low expression in the smooth-muscle layer. CONCLUSION: Only CRFR(2beta) is expressed in vasculature of nonpregnant and pregnant rats and may mediate the vasorelaxant effect of CRF. This receptor is present predominantly in the vascular endothelium and to a lesser extent in the smooth muscle. The expression of CRF receptor in pregnant rat vasculature is down regulated at term of gestation. PMID- 10865183 TI - Ca(2+) sensitivity of fetal coronary arteries exposed to long-term, high-altitude hypoxia. AB - OBJECTIVE: To determine the contribution of decreased calcium responsiveness of fetal coronary arteries to decreased contractile responses to potassium and the thromboxane A(2) analogue U46619 in these arteries after exposure to chronic hypoxemia. METHODS: Concentration-response curves to Ca(2+) in beta-escin permeabilized left circumflex (LCx), left anterior descending (LAD), and right coronary artery (RCA) rings from high-altitude (HA) and control (CON) fetuses were measured. In a second set of beta-escin-permeabilized coronary artery rings, the effect of U46619 on Ca(2+) sensitivity was tested. RESULTS: Maximum Ca(2+) activated force (T(max)) was decreased in HA LCx (CON 0.091+/-0.010 versus HA 0.057+/-0.006 g/cm(2); P<.05) and HA LAD (CON 0.065+/-0.012 versus HA 0.031+/ 0.007 g/cm(2); P <.05). No significant difference was observed in the RCA. There was no change in the pD(2) (-log EC(50)) values between CON and HA coronary rings. The Ca(2+) sensitizing effect of U46619 on submaximal Ca(2+)-activated force was lower only in the HA LCx (CON 0.044+/-0.010 versus HA 0.023+/-0.006 g/cm(2) at 10(-5) mol/L; P<.05). CONCLUSION: These results indicate that maximum tension development in response to Ca(2+) was decreased in the HA LCx and LAD but not the RCA; however, Ca(2+) sensitivity of the contractile apparatus was unaltered in all of them. Decreased Ca(2+) responsiveness may partially explain the decreased contractile capability of fetal LCx and LAD during long-term, high altitude intrauterine hypoxemia. PMID- 10865184 TI - Inhibitory effect of iloprost on the contractility of lower uterine segment myometrium from rhesus monkeys in normal-term and androstenedione-induced preterm labor. AB - OBJECTIVE: Iloprost, a combined EP(1) stimulatory and IP inhibitory receptor agonist, was tested in vitro on myometrium from the lower uterine segment of pregnant rhesus monkeys to compare its effects in spontaneous labor and in labor induced by the administration of androstenedione to the mother. METHODS: Pregnant rhesus monkeys carrying fetuses of known gestational age were instrumented under halothane general anesthesia with femoral artery and vein catheters and uterine electromyogram leads. Experimental animals were infused with androstenedione from 139 days' gestation. Control animals were infused with intralipid vehicle from 139 days' gestation. Lower uterine segment myometrium was removed from control animals either before labor began (n = 6) or in spontaneous labor (n = 4) and from animals undergoing premature labor induced by androstenedione (n = 4). Myometrial contractility in response to iloprost was evaluated using a superfusion system in vitro. RESULTS: Iloprost was inhibitory on myometrium obtained from the lower uterine segment from androstenedione-treated animals as well as vehicle-infused animals in spontaneous term labor. In contrast, iloprost had no effect on myometrial strips from control animals not in labor. CONCLUSION: These findings indicate up-regulation of IP receptors which inhibit myometrial contractility and/or down-regulation of EP(1) receptors which stimulate myometrial contractility in the lower uterine segment during labor. A relative increase in inhibitory responses in the lower uterine segment during labor may enable this region to dilate to allow passage of the fetus. PMID- 10865185 TI - Second-trimester levels of maternal serum human chorionic gonadotropin and inhibin a as predictors of preeclampsia in the third trimester of pregnancy. AB - OBJECTIVE: To determine whether second-trimester maternal serum levels of inhibin A, human chorionic gonadotropin (hCG), unconjugated estriol (uE3), and alpha fetoprotein (AFP) are predictive of the later onset of preeclampsia in pregnancy. METHODS: Retrospective evaluation of serum analyte levels in 60 women with preeclampsia compared with 300 controls. Levels of each analyte were compared in women with preeclampsia and controls using matched rank analysis. Analytes that were significantly different between groups were examined with univariate and bivariate Gaussian distribution analysis. RESULTS: Second-trimester inhibin A (1.36 multiples of the median [MoM]) and hCG (1.40 MoM) levels were significantly but modestly elevated in women who later developed preeclampsia. A combination test of maternal age plus inhibin A and hCG predicted 23% of cases of preeclampsia with 95% specificity. There was a statistically significant trend for inhibin A, but not hCG, levels to be higher when the onset of preeclampsia occurred within a shorter (<17 weeks) interval after collection of the second trimester screening sample. CONCLUSIONS: Second-trimester serum levels of inhibin A and hCG are modest predictors of the later onset of preeclampsia. Inhibin A may be a better predictor of early-onset preeclampsia, which is associated with a higher maternal and perinatal morbidity and mortality, than preeclampsia at or near term. PMID- 10865186 TI - Comparison of pharmacokinetics of a conjugated equine estrogen preparation (premarin) and a synthetic mixture of estrogens (C.E.S.) in postmenopausal women. AB - OBJECTIVE: To compare the pharmacokinetics and relative bioavailabilities of key estrogen components of Premarin (Wyeth-Ayerst, Canada) with those of a generic conjugated estrogen preparation, C.E.S. (synthetic mixture of estrogens; ICN, Montreal, Canada) in healthy postmenopausal women. METHODS: We conducted a randomized, single-dose, two-treatment, three-period crossover study in 41 postmenopausal women. After an oral dose (2 x 0.625 mg) of Premarin or C.E.S., plasma concentrations of unconjugated and total estrone (E(1)), equilin (Eq), 17beta-estradiol (17beta-E(2)), 17beta-dihydroequilin (17beta-Eq), Delta(8) esterone (Delta(8)-E(1)) and Delta(8),17beta-estradiol (Delta(8),17beta-E(2)) were measured over 72 hours using gas chromatography and mass spectroscopy. RESULTS: After administration of C.E.S., E(1), Eq, and 17beta-Eq appeared in blood at a significantly faster rate (lower t(max)) than after Premarin. The rapid appearance of estrogens after C.E.S. was associated with significantly higher (14-61%) C(max) values. In contrast to the high C(max) values, the area under the curve (AUC)(infinity) of unconjugated and total Eq, and 17beta-Eq were significantly lower after C.E.S., whereas those of E(1) were significantly higher. Although, the t(max) values for 17beta-E(2) were lower and the C(max) values higher after C.E.S., only the C(max) of unconjugated 17beta-E(2) was significantly different after Premarin. Unconjugated and total Delta(8)-E(1) and its main metabolite, Delta(8),17beta-E(2), were detectable in plasma only after administration of Premarin. The geometric mean ratio (GMR) (C. E.S./Premarin) of bioavailability parameters indicated that all C(max) and t(max) values for the unconjugated and total E(1), Eq, 17beta-E(2), and 17beta-Eq fell outside the regulatory requirement that the 90% confidence intervals of GMRs of two products be within 80% and 125%. Similarly, with the exception of total E(1) and total Eq, none of the AUC(t) or AUC(alpha) of the remaining estrogens meets the required regulatory standards of bioequivalence. CONCLUSIONS: C.E.S. is not bioequivalent to Premarin. Because C.E.S. also is not pharmaceutically equivalent to Premarin, it cannot be assumed to be therapeutically equivalent. Until long-term clinical trials with C.E.S. demonstrate its efficacy, extrapolation of the long-term benefits described for Premarin to C.E.S. would be risky and questionable. PMID- 10865187 TI - A bcl-x(S) adenovirus demonstrates therapeutic efficacy in an ascites model of human breast cancer. AB - OBJECTIVES: To determine whether a Bcl-x(S) adenoviral vector has therapeutic potential in an ascites model of human breast cancer in nude mice. METHODS: Advanced ascites were developed by injecting mice intraperitoneally (IP) with MDA MB-231 human breast carcinoma cells. Mice received sequential IP injections of the Bcl-x(S) virus or a control lac-Z adenovirus. A third group of mice received no virus. Tumor burden and survival were monitored. Histopathology and necropsies were performed on mice. RESULTS: A single injection of the Bcl-x(S) adenovirus produced no systemic or local toxicity and no abnormal histopathology in normal mice. However, abdominal organs within these mice were transduced with the Bcl x(S) vector. Adenoviral gene transduction efficiency in MDA-MB-231 ascites was 36+/-6.40% (n = 3). Percent weight change differences revealed that ascites bearing mice injected three times with the Bcl-x(S) vector showed a statistically significant decrease in tumor burden compared with lac-Z-injected mice (n = 7; P = .012 on days 10-15 after the first injection). Mice injected with the Bcl-x(S) vector had significantly greater survival relative to lac-Z-injected mice (n = 7; P = .0004). Bcl-x(S) protein expression was detected in aspirates of mice injected with the Bcl-x(S) vector but not the lac-Z vector. Necropsies revealed that ascites bearing mice injected with Bcl-x(S) vector lacked carcinoma in the peritoneal cavity compared with control mice. CONCLUSION: The Bcl-x(S) adenovirus can reduce tumor burden and increase survival in an ascites model of advanced stage breast cancer. PMID- 10865189 TI - Species, population and age diversity in cell resistance to adhesion of Neisseria meningitidis serogroups A, B and C. AB - The variation of cell adherence of meningococci serogroups A, B and C and influenza viruses was investigated in 11 animal species and in humans of different age groups (1st, 2nd, 3rd and 4th weeks; 2nd-3rd months; 4th-12th months, 2nd-3rd years; and 18th-60th years of life) as well as in women during pregnancy (17th-36th weeks) and childbirth. Red blood cells of all animals tested as well as of human newborns were absolutely resistant to attachment of meningococci. In neonatal and the later periods the human cells become far differently sensitive individually to meningococcal adhesion. In contrast, the viral adhesion was characterized by different individual cell sensitivity in all age groups tested. Pregnancy and childbirth did not influence the women's cell sensitivity to adhesion of Neisseria meningitidis. Different receptors are involved in interactions of human cells with influenza viruses and meningococci. The function of meningococcal receptors on human cells develops during postnatal ontogenesis. The variations express both specific (genetic) and ontogenetic (individual) differences in natural resistance to meningococcal infection. PMID- 10865188 TI - Lipopolysaccharide induces expression of genes encoding pro-inflammatory cytokines and the elastin-degrading enzyme, cathepsin S, in human cervical smooth muscle cells. AB - OBJECTIVE: Vaginal and amniotic infection with gram-negative bacteria is associated with preterm birth. We previously reported that human cervical smooth muscle cells (CSMC) respond to pro-inflammatory cytokines by expressing enzymes that degrade the extracellular matrix. Our objective was to characterize the effects of lipopolysaccharide (LPS) from Escherichia coli (E coli), Bacteroides fragilis, (B frag)and Fusobacterium nucleatum (F nuc)on the expression of pro inflammatory cytokines and the elastin-degrading enzyme, cathepsin S, in human CSMC. METHODS: Human CSMC were exposed to LPS and the expression of mRNAs encoding pro-inflammatory cytokines and cathepsin S, and selected matrix metalloproteinases (MMPs) was analyzed by Northern blotting. The effect of cytokine-neutralizing antibodies on LPS-induced cathepsin S mRNA expression also was determined. RESULTS: E coli LPS increased expression of cathepsin S 12.5-fold after 12 hours; MMP-1 and MMP-3 mRNAs also were increased 2.9- and 3.5-fold, respectively. Tumor necrosis factor (TNF)-alpha, interleukin (IL-1)alpha, and IL 1beta mRNAs were markedly up-regulated after 3 hours of LPS treatment. B frag and F nuc LPS also induced TNF-alpha and cathepsin S mRNAs. E coli LPS caused a sevenfold increase in TNF-alpha secretion after 5 to 8 hours. Antihuman TNF-alpha monoclonal antibody, but not a monoclonal antibody to the low-density lipoprotein receptor, reduced the LPS-induced increase in cathepsin S mRNA by 27%, whereas neutralizing antibodies against IL-1alpha and IL-1beta did not suppress the response. Human CSMC were shown to express the toll-like receptor (TLR-2) and TLR 4 genes, which mediate the action of LPS. TLR-2 mRNA was up-regulated by TNF alpha. CONCLUSION: CSMC respond to LPS with increased expression of pro inflammatory cytokines and cathepsin S. Increases in cathepsin S mRNA result only in part from the rapid induction of TNF-alpha gene expression. TNF-alpha may also augment the CSMC response to LPS by increasing expression of the LPS signaling receptor, TLR-2, which probably directly mediates LPS action. These observations provide a mechanism by which gram-negative bacteria can precipitate cervical changes associated with preterm birth. PMID- 10865190 TI - Induction of tumor necrosis factor-alpha and inducible nitric oxide synthase fails to prevent toxoplasmic encephalitis in the absence of interferon-gamma in genetically resistant BALB/c mice. AB - Following infection with Toxoplasma gondii, certain strains of mice, such as BALB/c, are genetically resistant to development of toxoplasmic encephalitis (TE) and establish a latent chronic infection as do humans. Thus, these animals appear to be a suitable model to analyze the mechanism of resistance to TE. Since the mechanism for their genetic resistance is unknown, we examined the role of interferon-gamma (IFN-gamma) tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS) in the resistance using BALB/c-background IFN-gamma-deficient (IFN-gamma(-/-)) mice. IFN-gamma(-/-) and control mice were infected with the ME49 strain of T. gondii and treated with sulfadiazine to establish chronic infection. After discontinuing sulfadiazine, the IFN-gamma(-/-) mice all died, whereas the control mice all survived. Histological studies revealed remarkable inflammatory changes associated with large numbers of tachyzoites in brains of the IFN-gamma(-/-) mice but not in the control mice after discontinuation of sulfadiazine. Large amounts of mRNA for tachyzoite specific SAG1 were detected in brains of only the IFN-gamma(-/-) mice. IFN-gamma mRNA was detected in brains of only the control mice, whereas mRNA for TNF-alpha and iNOS were detected in brains of both strains of mice. The amounts of the mRNA for TNF-alpha and iNOS did not differ between these mice. Treatment of IFN-gamma( /-) mice with recombinant IFN-gamma prevented development of TE. These results demonstrate that IFN-gamma is crucial for genetic resistance of BALB/c mice against TE and that TNF-alpha and iNOS are insufficient to prevent TE in the absence of IFN-gamma. PMID- 10865191 TI - Differential tolerance induction by lipoarabinomannan and lipopolysaccharide in human macrophages. AB - Various bacterial cell wall components have been shown to induce hyporesponsiveness in macrophages (MAC). Here, mycobacterial glycolipids were employed to determine whether they induce a state of 'tolerance/hyporesponsiveness' in MAC in vitro in order to assess whether mycobacterial components negatively affect the immune response to Mycobacterium tuberculosis. Arabinosylated lipoarabinomannan (ARA-LAM) stimulated hyporesponsiveness by reducing TNF-alpha, GM-CSF, G-CSF, IL-10, and IL-6 release similarly to LPS, but caused no changes in IL-8 secretion. Mannose-capped LAM (MAN-LAM) acted in a different way in that TNF-alpha, GM-CSF, and IL-10 were upregulated after restimulation of MAC. Blocking experiments by mannan suggest mannose-receptor involvement in MAN-LAM activation only. Cross-stimulation experiments demonstrated a hierarchy of signaling, with LPS being the most potent stimulator and mediating abrogation of ARA-LAM-stimulated tolerance but not vice versa. MAN-LAM was the least potent stimulator of either MAC activation and induction of hyporesponsiveness. Similarly to LPS, ARA-LAM upregulated CD14 surface expression after restimulation. Recurrent MAN-LAM treatment either downmodulated or did not induce any change in CD14 expression. The role of MAN LAM regulated cytokine secretion as well as implications regarding M. tuberculosis infection will be discussed. PMID- 10865192 TI - Immune responses mediating survival of naive BALB/c mice experimentally infected with lethal rodent malaria parasite, Plasmodium yoelii nigeriensis. AB - The rodent malaria parasite, Plasmodium yoelii nigeriensis is known to cause fatal malaria infections in BALB/c mice. However, we found that nearly 5% of inbred BALB/c mice could overcome primary infections initiated with lethal inoculum of P. y. nigeriensis asexual blood-stages, without any experimental intervention. These 'survivor' mice developed peak parasitemia levels of about 5% and successfully resolved their infections in about two weeks time; infected blood collected during the descending phase of infection in these mice and subinoculated in naive recipients resulted in a normal lethal course of infection. Typically, the parasites in survivor mice looked 'sick' compared to those in the susceptible mice. In experiments to define temporal basis of this protection, we found that purified splenic B cells isolated from such a survivor mouse, plus T cells from an infected or naive mouse, could adoptively transfer this protection to an X-irradiated, naive mouse against a lethal parasite challenge. Purified T cells or B cells alone from the survivor mouse donor provided no protection to the X-irradiated, naive recipient. Passive transfer of sera collected from survivor mice animals a week after recovery from infection was also able to substantially alter the course of preestablished P. y. nigeriensis infection. These findings are discussed in the light of recent reports on the genetic control of blood parasitemia in mouse malaria models. In the generally lethal malaria infections such as those caused by P. y. nigeriensis in mice and by Plasmodium falciparum in naive children, it is not clear what constitutes a protective immune response in cases which survive primary infections without any experimental or therapeutic intervention. An understanding of these mechanisms and their regulation would help design better vaccination strategies. PMID- 10865193 TI - Identification of protective outer membrane antigens of Brucella ovis by passive immunization of mice with monoclonal antibodies. AB - Outer membrane proteins (OMPs) and rough lipopolysaccharide (R-LPS), the main surface antigens of Brucella ovis, display surface-exposed epitopes. Mixtures of monoclonal antibodies (mAbs) to both antigens were previously shown to protect mice against a B. ovis challenge. To further identify the antigens involved, seven mAbs against Brucella OMPs (Omp10, Omp16, Omp19, Omp25, Omp31, Omp2b and Omp1) and three to R-LPS were tested for protection either individually or in combinations. Significant reduction in spleen infection in challenged mice, relative to controls, was used as the protection criteri. Controls included nonimmunized mice and mice given an irrelevant, anti-O-polysaccharide (OPS), mAb. For comparison, a group received a mouse serum containing antibodies to both OMPs and R-LPS; this serum was prepared by immunization with a B. ovis hot-saline extract which, as described previously, induces protective immunity in mice and rams. Significant protection was observed with both mAbs to OMPs and R-LPS. mAbs to Omp16, Omp19 and Omp31 afforded the highest protection and prevented the development of splenomegaly. The protective effect of mAb to Omp31 was not interfered with by nonprotective mAbs in different mixtures. The data presented confirm the protective role of antibodies to OMPs and R-LPS against B. ovis, and identify several OMPs, especially Omp31, which are promising candidates for a subunit vaccine against ram epididymitis. PMID- 10865194 TI - Emerging and reemerging infections in africa: the need for improved laboratory services and disease surveillance. AB - Emerging and reemerging infections pose a serious public health threat to most countries of tropical Africa. In the past decade, epidemics of diseases including cholera, dysentery, meningitis, yellow fever and Ebola virus have resulted in significant morbidity and mortality. Improved laboratory services and disease surveillance systems are essential to monitor disease trends and to initiate public health action. The present situation of emerging and reemerging infections in Africa is described in this review, and strategies for improved disease surveillance and monitoring are discussed. PMID- 10865195 TI - Yersiniae other than Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis: the ignored species. AB - The genus Yersinia is composed of 11 species, of which three (Y. pestis, Y. pseudotuberculosis, and Y. enterocolitica) have been exhaustively characterized. The remaining eight species (Y. frederiksenii, Y. intermedia, Y. kristensenii, Y. bercovieri, Y. mollaretii, Y. rohdei, Y. ruckeri, and Y. aldovae) have not been studied extensively and, because of the absence of classical Yersinia virulence markers, are generally considered to be nonpathogenic. However, recent data suggest that some of these eight species may cause disease by virtue of their having virulence factors distinct from those of Y. enterocolitica. These data raise intriguing questions about the mechanisms by which these species interact with their host cells and elicit human disease. PMID- 10865196 TI - Mating, parasites and other trials of life in social insects. AB - Females of many animal species mate with several different males, despite the extra costs. Recent studies in social insects now suggest that the resulting increase in genotypic variance in offspring is beneficial under selection by parasites. This finding also helps to understand the evolution and maintenance of sexual reproduction. PMID- 10865197 TI - Cytomegalovirus MHC class I homologues and natural killer cells: an overview. AB - Viruses that establish a persistent infection with their host have evolved numerous strategies to evade the immune system. Consequently, they are useful tools to dissect the complex cellular processes that comprise the immune response. Rapid progress has been made in recent years in defining the role of cellular MHC class I molecules in regulating the response of natural killer (NK) cells. Concomitantly, the roles of the MHC class I homologues encoded by human and mouse cytomegaloviruses in evading or subverting NK cell responses has received considerable interest. This review discusses the results from a number of studies that have pursued the biological function of the viral MHC class I homologues. Based on the evidence from these studies, hypotheses for the possible role of these intriguing molecules are presented. PMID- 10865198 TI - Bacterial ureases in infectious diseases. AB - Ureases are multi-subunit, nickel-containing enzymes that catalyze the hydrolysis of urea to carbon dioxide and ammonia. This brief review discusses the biochemistry and genetics of bacterial ureases and outlines the roles of urea metabolism in microbial ecology and pathogenesis of some of the principle ureolytic species affecting human health. PMID- 10865199 TI - Role played by lactobacilli in controlling the population of vaginal pathogens. AB - The role of Lactobacillus species in the female urogenital tract as a barrier to infection is of considerable interest. These organisms are believed to contribute to the control of vaginal microbiota by competing with other microorganisms for adherence to epithelial cells and by producing antimicrobial compounds. These bactericidal compounds include organic acid, which lowers the vaginal pH, hydrogen peroxide, bacteriocin-like substances and possibly biosurfactants. PMID- 10865200 TI - Moraxella catarrhalis: a review of an important human mucosal pathogen. AB - Moraxella catarrhalis has again been recognized as a significant pathogen. The past decade has witnessed an increased amount of research and understanding of the pathogenesis of the organism. This review will summarize the research pertaining to the epidemiology and components of pathogenesis in M. catarrhalis. PMID- 10865201 TI - Progress toward the development of a vaccine to prevent Moraxella (Branhamella) catarrhalis infections. AB - Moraxella catarrhalis is a major cause of otitis media and respiratory disease. Vaccine development is at the antigen identification stage. This review examines the more promising antigens, including the 200K protein, the hemagglutinins, the lactoferrin-binding proteins, the UspA proteins, the CopB protein, the transferrin-binding proteins, the CD protein, the E protein and lipooligosaccharide conjugates. Clinical testing of some of these antigens should begin soon. PMID- 10865202 TI - How does antiresorptive therapy decrease the risk of fracture in women with osteoporosis? PMID- 10865203 TI - Electrophysiological characterization of ion channels in osteoclasts isolated from human deciduous teeth. AB - Ion channels contribute to several important processes in osteoclasts, including proton transport and volume regulation. Although ion channels have been described in osteoclasts from several species, little is known about their properties in human osteoclasts. We devised a method for isolation of authentic human osteoclasts from deciduous teeth undergoing root resorption, and characterized currents in these cells using patch-clamp techniques. Three types of K(+) current were identified. Hyperpolarization elicited an inwardly rectifying K(+) current in most osteoclasts, which was inhibited by Ba(2+) in a voltage- and time dependent manner. Depolarization elicited an outwardly rectifying and tetraethylammonium-sensitive current, consistent with a large-conductance Ca(2+) dependent K(+) channel. In addition to these basal currents, extracellular adenosine 5'-triphosphate (ATP) elicited a linear current that was identified as a Ca(2+)-dependent K(+) current, based on its reversal potential close to that predicted for K(+), its blockade by quinine, and its activation by Ca(2+) ionophore. Last, an outwardly rectifying current was observed to activate spontaneously or in response to ATP, with properties of a swelling-activated Cl( ) current. This current reversed direction close to the Cl(-) equilibrium potential and was blocked by the anion channel blocker, niflumic acid, identifying it as a Cl(-) current. In summary, we have developed a novel method for isolation of authentic human osteoclasts and have characterized K(+) and Cl( ) currents. Cl(-) current mediates charge compensation during electrogenic H(+) transport, so activation of Cl(-) current may contribute to the stimulatory effects of extracellular ATP on bone resorption. PMID- 10865204 TI - Does suppression of bone turnover impair mechanical properties by allowing microdamage accumulation? AB - One plausible purpose of bone turnover is to repair bone microdamage. We hypothesized that suppression of bone turnover impairs bone quality by allowing accumulation of microdamage. We investigated the effect of high-dose etidronate (EHDP) on bone's mechanical properties and microdamage accumulation. Skeletally mature beagles, 1-2 years old at the beginning of the study, were treated with daily injections of vehicle or EHDP at 0.5 mg/kg per day or 5.0 mg/kg per day for 1 year. X-rays were taken at baseline and monthly from 7 to 12 months. Bones were taken upon sacrifice and biomechanical tests, histomorphometry, and microdamage analyses were performed. Fractures of ribs and/or thoracic spinous processes were found in 10 of 11 dogs treated with the higher dose EHDP. Only one fracture of a thoracic spinous process was found in dogs treated with the lower dose EHDP, and no fractures were found in the vehicle controls. Biomechanical tests showed reduced mechanical strength in ribs and lumbar vertebrae, but not in the femoral diaphysis or thoracic spinous process in the higher dose EHDP group. Histomorphometric measurements showed a significant reduction of cancellous bone turnover in both EHDP-treated groups compared with controls. In dogs treated with the higher dose EHDP, activation frequency was reduced to zero in both cortical and cancellous bone. Osteoid volume increased significantly, especially in trabecular bone, resulting in reduced mineralized bone volume in the higher dose EHDP group. Microcrack numerical density (Cr.Dn) increased significantly only in the lumbar vertebral body in the higher dose EHDP group, but not in the rib or thoracic spinous process where fractures occurred. These findings show that suppression of bone turnover using high doses of EHDP is associated with fractures of the ribs and spinous processes in dogs. This is most likely the result of excessive amounts of unmineralized bone produced by the inhibition of mineralization at these high doses, rather than by the accumulation of microdamage. PMID- 10865205 TI - Extracellular calcium-sensing-receptor (CaR)-mediated opening of an outward K(+) channel in murine MC3T3-E1 osteoblastic cells: evidence for expression of a functional CaR. AB - The existence in osteoblasts of the G-protein-coupled extracellular calcium (Ca(o)(2+))-sensing receptor (CaR) that was originally cloned from parathyroid and kidney remains controversial. In our recent studies, we utilized multiple detection methods to demonstrate the expression of CaR transcripts and protein in several osteoblastic cell lines, including murine MC3T3-E1 cells. Although we and others have shown that high Ca(o)(2+) and other polycationic CaR agonists modulate the function of MC3T3-E1 cells, none of these actions has been unequivocally shown to be mediated by the CaR. Previous investigations using neurons and lens epithelial cells have shown that activation of the CaR stimulates Ca(2+)-activated K(+) channels. Because osteoblastic cells express a similar type of channel, we have examined the effects of specific "calcimimetic" CaR activators on the activity of a Ca(2+)-activated K(+) channel in MC3T3-E1 cells as a way of showing that the CaR is not only expressed in those cells but is functionally active. Patch-clamp analysis in the cell-attached mode showed that raising Ca(o)(2+) from 0.75 to 2.75 mmol/L elicited about a fourfold increase in the open state probability (P(o)) of an outward K(+) channel with a conductance of approximately 92 pS. The selective calcimimetic CaR activator, NPS R-467 (0.5 micromol/L), evoked a similar activation of the channel, while its less active stereoisomer, NPSS-467 (0.5 micromol/L), did not. Thus, the CaR is not only expressed in MC3T3-E1 cells, but is also functionally coupled to the activity of a Ca(2+)-activated K(+) channel. This receptor, therefore, could transduce local or systemic changes in Ca(o)(2+) into changes in the activity of this ion channel and related physiological processes in these and perhaps other osteoblastic cells. PMID- 10865206 TI - Osteoclast lineage commitment of bone marrow precursors through expression of membrane-bound TRANCE. AB - Osteoclast formation from hemopoietic precursors is induced by TRANCE (also called RANKL, ODF, and OPGL), a membrane-bound ligand expressed by bone marrow stromal cells. Because soluble recombinant TRANCE is a suboptimal osteoclastogenic stimulus, and to eliminate the need for such dependence on stromal cells, membrane-bound TRANCE was expressed in hematopoietic precursors using retroviral gene transfer. Four TRANCE-expressing osteoclast cell lines were established that continuously generate large numbers of multinucleated cells and express tartrate-resistant acid phosphatase and calcitonin receptors. The multinuclear cells are long-lived and either fuse continuously with each other and with mononuclear cells to form enormous syncytia, or separate to form daughter multinuclear cells. When formed on bone, but not on plastic, the majority of multinuclear cells develop actin rings on bone, and resorb bone, suggesting that bone matrix may provide additional signals that facilitate osteoclastic functional maturation. Surprisingly, multinuclear cells originate from fusion of proliferating mononuclear cells that strongly express the mature macrophage markers F4/80 and Fc receptor, which are not expressed by osteoclasts. These results indicate that osteoclasts can be derived from F4/80-positive and Fc receptor-positive cells, and that TRANCE induces osteoclastic differentiation partly by suppressing the macrophage phenotype. PMID- 10865207 TI - Is high-dose estrogen-induced osteogenesis in the mouse mediated by an estrogen receptor? AB - Although estrogen is known to induce new bone formation in the long bones of female mice, this response is only thought to occur following administration of high doses, suggesting that it may not be mediated by a conventional estrogen receptor. To address this question further, we first examined the stereospecificity of this response by comparing the potency of 17beta-estradiol (E(2)) in stimulating cancellous bone formation at the proximal tibial metaphysis of intact female mice with that of the relatively inactive stereoisomer, 17alpha estradiol (alphaE(2)). We found that E(2) was significantly more potent than alphaE(2), as assessed by histomorphometry. To provide further evidence for an estrogen-receptor-mediated process, we examined whether E(2)-induced osteogenesis in intact female mice could be inhibited by the estrogen receptor antagonist, ICI 182,780 (ICI). Although ICI itself had no effect on histomorphometric indices of the proximal tibial metaphysis when given alone, it significantly inhibited the osteogenic response to E(2). Finally, we examined the dose dependency of E(2) induced osteogenesis at the proximal tibial metaphysis in intact mice. We found that E(2) stimulated cancellous bone formation in a dose-dependent manner over a wide dose range (i. e., 1-4000 microg/kg per day), with significant increases observed at doses of 4 microg/kg per day and beyond. Our results raise the possibility that estrogen-induced osteogenesis in the mouse represents an estrogen-receptor-mediated response that is not confined solely to supraphysiological estrogen levels. PMID- 10865208 TI - Transient estrogen exposure of female mice during early development permanently affects osteoclastogenesis in adulthood. AB - Estrogens modulate bone tissue turnover in both experimental animal models and postmenopausal women. Our previous studies have shown that exposure to diethylstilbestrol (DES) during the perinatal period increases peak bone mass in female mice in adulthood. We investigated whether developmental DES exposure can influence bone mass by affecting osteoclastogenesis. Female mice were injected with 100 microg/kg body weight DES from days 9-16 of gestation or, alternatively, pups received neonatal injections of 2 microg of DES from days 1-5 of life. Animals were weaned at 21 days of age and effects of estrogen on bone cells were evaluated in adulthood. A significant increase in bone mass in female mice was already observed at 2 months, with a maximal effect in older animals. Bone sections from DES-treated animals showed a significant decrease in osteoclast number and tartrate-resistant acid phosphatase (TRAP) enzymatic activity as compared with controls. To verify the importance of the estrogen surge at puberty in this event, a group of control and DES-treated mice were ovariectomized at 17 days to prevent puberty, and potential effect on osteoclastic cells was evaluated in adulthood. As expected, ovariectomy induced an increase of TRAP-positive cells. DES treatment blunted the ovariectomized-dependent increase of the total number of osteoclastic cells, suggesting a role of developmental DES exposure in the process of bone-cell imprinting. Our data indicate, for the first time, that transient changes in estrogen levels during development modulate bone turnover and osteoclastogenesis likely participating in bone-cell imprinting during early phases of bone development, and that this effect could be induced by direct alteration of bone microenvironment. PMID- 10865209 TI - The immediate early gene product hCYR61 localizes to the secretory pathway in human osteoblasts. AB - The human cysteine-rich protein 61 (hCYR61) belongs to the growing CCN (CYR61/CTGF/NOV) family of immediate early genes, which modulate cell growth and differentiation. hCYR61 is regulated by 1alpha, 25-dihydroxyvitamin D(3) (1,25 (OH)(2)D(3)) and growth factors in fetal human osteoblasts (hFOB cells). The murine homologue CYR61 was characterized as an extracellular matrix-associated protein that modulates basic fibroblast growth factor signaling, angiogenesis, and binds to integrin alpha(v)beta(3). Here we report the intracellular localization of the human CYR61 gene product by overexpressing fusion proteins with green fluorescent protein (GFP) in primary osteoblasts and the hFOB cell line. Full-length hCYR61-GFP localizes to the Golgi apparatus and cytoplasmatic granules, indicating targeting to the secretory pathway. Secretion of hCYR61 from osteoblasts is verified by Western blot detection from cellular supernatants. A truncated hCYR61-GFP fusion protein containing only the 34 N-terminal amino acids of hCYR61 also localizes to the Golgi apparatus mainly in the perinuclear region, which suggests that the N-terminus of hCYR61 is sufficient to target the protein to the secretory pathway. In summary, our results present the first evidence that human CYR61 localizes to the secretory pathway in primary osteoblasts and hFOB cells, and that it is secreted from these cells. The N-terminal 34 amino acids of hCYR61 provide a sufficient Golgi targeting sequence. Together with the immediate early regulation of hCYR61 mRNA by 1,25-(OH)(2)D(3), this suggests that hCYR61 might function as an extracellular signaling molecule in human bone. PMID- 10865210 TI - Expression of fibrillins and other microfibril-associated proteins in human bone and osteoblast-like cells. AB - Fibrillin-containing microfibrils are structural components of extracellular matrices of a diverse range of tissues, including bone. Their importance in bone biology is illustrated by the skeletal abnormalities manifest in the congenital disorder, Marfan syndrome, which results from mutations in the fibrillin-1 gene. We investigated the expression of fibrillins and other microfibril-associated proteins in human bone and bone-derived osteoblasts. Analysis of RNA extracted from cancellous bone showed expression of mRNAs encoding fibrillin-1 and -2, MAGP 1 and -2, LTBP-2, and MP78/70 (Big-h3). In demineralized normal mature bone, fibrillin-1 was immunolocalized to fibrils within the bone matrix and pericellularly to cells lining the endosteal surfaces of trabecular bone, some osteocytes, and cells associated with blood vessels. LTBP-2 was also identified at the endosteal surface and within the bone matrix in a lamellar fashion. In addition, primary osteoblast-like cells cultured from human trabecular bone (obtained from patients at joint replacement surgery) were found to express abundant mRNA for fibrillins and associated glycoproteins. Moreover, using western blot analysis, fibrillin-1 protein was shown to be secreted into the medium and to be deposited into the cell layer. Immunofluorescence staining of the cell layer visualized fibrillin-1 in the matrix as a three-dimensional network of fine filaments. Expression of fibrillin-1 by osteoblast-like cells was constitutive, and a number of skeletally active agents had little effect on mRNA or protein levels. These results show that human osteoblasts from mature bone express fibrillins and other microfibril-associated proteins, and suggest a role for these molecules in adult human bone. PMID- 10865211 TI - A novel in vitro culture system for analysis of functional role of phosphate transport in endochondral ossification. AB - In vivo expression of the type III sodium-dependent phosphate transporter (NaPiT) Glvr-1 during endochondral ossification, suggests a functional role for inorganic phosphate (Pi) transport in cartilage calcification. For further analysis of this relationship, an in vitro model of endochondral ossification is required. In this context, we investigated the characteristics of Pi transport in the new chondrogenic cell line ATDC5 in relation to extracellular matrix (ECM) formation and mineralization. Pi uptake in ATDC-5 cells and in isolated matrix vesicles (MVs) is mediated by an Na-dependent Pi transporter with a pH dependency characteristic of a type III Pi carrier (lower activity at alkaline pH). Northern blot analysis indicated that ATDC-5 cells express Glvr-1 transcripts during the various stages of their maturation with a maximal level during the proliferating stage. In isolated MVs, Pi transport activity was maximal at day 21, concomitant with the beginning of type X collagen messenger RNA expression. These events preceded the initiation of matrix mineralization, which was apparent at day 25, and then gradually increased until day 47. This temporal relationship between maximal Pi transport activity in MVs and the expression of a marker of mineralizing chondrocytes is compatible with the possible involvement of Pi transport in the ECM calcification observed in ATDC-5 cell cultures. In conclusion, these observations suggest that ATCD-5 cells in culture represent a promising model for the analysis of a functional role of Pi transport in the initial events of endochondral ossification. PMID- 10865212 TI - Tetracyclines induce apoptosis in osteoclasts. AB - Chemically modified tetracyclines (CMTs) are thought to inhibit bone resorption through inhibition of matrix metalloproteinases. Here we report that some tetracyclines also induce apoptosis in rabbit osteoclasts and inhibit differentiation and activity of osteoclasts in murine osteoblast/marrow cocultures. Apoptosis of mature rabbit osteoclasts increased from 5.5 +/- 1.4% (mean +/- SD) in control cultures to 44.9 +/- 6.3% (p < 0.001) and 18.9 +/- 4.0% (p < 0.005) with CMT-3 and doxycycline (10 microg/mL), respectively. CMT-2 or CMT 5 did not alter osteoclast viability even at 25 microg/mL. In murine osteoblast/marrow cocultures over 11 days, CMT-3 and doxycycline (5 microg/mL) reduced the formation of mature osteoclasts and inhibited resorption to 21 +/- 9% (p < 0.01) and 49 +/- 4% (p < 0.01) of untreated cultures. Induction of osteoclast apoptosis is an additional property of tetracyclines that may contribute to their ability to inhibit bone resorption. PMID- 10865213 TI - Transient expression of activin betaA mRNA on osteoprogenitor cells in rat bone regeneration after drill-hole injury. AB - We investigated the expression of activin betaA on osteoprogenitor cells in the regenerating bone and bone marrow of the rat femur after drill-hole injury, by immunocytochemistry and in situ hybridization. The periosteum and endosteum adjacent to the wound region showed marked thickening at day 3 and abundant osteoprogenitor cells, which were immunoreactive for proliferating cell nuclear antigen and showed positive reactions for alkaline phosphatase activity, and existed in the inner layer of the periosteum as well as in the endosteum. During the same period, these osteoprogenitor cells began to exhibit activin betaA immunoreactivity and mRNA expression. However, the latter expression gradually reduced the intensity as the cells started to express osteocalcin mRNA during their differentiation to osteoblasts participating in the periosteal and medullary bone formation from day 5. Immunoreactivity for activin type IB and II receptors was also found on activin betaA-immunoreactive cells between days 3 and 7. The above findings suggest that proliferating osteoprogenitor cells, before their transformation to osteoblasts, transiently produce and release activin A, which may play crucial roles in bone and bone marrow regeneration in a receptor mediated, autocrine and paracrine fashion. PMID- 10865214 TI - Activin increases bone mass and mechanical strength of lumbar vertebrae in aged ovariectomized rats. AB - Activin is a member of the transforming growth factor-beta superfamily and is thought to be involved in the regulation of bone formation due to its presence in bone tissue and its osteogenic activity both in vitro and in vivo. We recently found that systemic administration of activin increased both tibial bone mass and mechanical strength in young growing rats. The present study investigated the effects of activin in aged ovariectomized (ovx) rats. Twelve-month-old Fischer rats were ovariectomized and maintained for 10 months. Recombinant human activin A (activin) or human parathyroid hormone 1-34 (PTH) was administered intramuscularly three times a week for 12 weeks. Activin (1 and 5 microg/kg) markedly increased lumbar vertebral bone mineral content and bone mineral density. Activin also increased the mechanical strength of the vertebral body, which was highly correlated to the bone mineral density of the vertebral body. The maximal response in bone mass and strength was observed at 1 microg/kg of activin, which was approximately equal to that induced by PTH at 40 microg/kg. Peripheral quantitative computed tomography revealed that activin enlarged the cross-sectional size of the vertebrae without changing the foramen area, indicating its effects on cortical shells. Histomorphometric analysis of cancellous bone of vertebral body in similar experiment showed that activin (3 microg/kg) increased bone volume and the mineralizing surface, although its effects were less than PTH. The present results indicate that low doses of activin are effective against vertebral bone loss in aged ovx rats. PMID- 10865215 TI - Bone resorption induced by 1 alpha,25 dihydroxyvitamin D(3) in vivo is not altered by inactivation of the plasminogen activator inhibitor 1. AB - One of the proteolytic systems produced by bone cells is the plasminogen activator/plasmin pathway, which involves the two plasminogen activators and the type 1 plasminogen activator inhibitor (PAI-1) and results in plasmin generation. We have recently demonstrated that this pathway plays a specific role in the degradation of the nonmineralized matrix of bone in vitro. To evaluate whether PAI-1 is required during bone resorption in vivo, we studied the effects of PAI-1 inactivation on bone metabolism using systemic administration of 1alpha,25 dihydroxyvitamin D(3) [1, 25(OH)(2)D(3)] as model. PAI-1-deficient (PAI-1-/-) and wild-type (WT) mice were injected intraperitoneally with 1,25(OH)(2)D(3) (2 microg/kg) or vehicle every other day during 4 weeks and analyzed using biochemical parameters of bone turnover, histomorphometric analysis of the proximal tibial metaphysis, and pQCT analysis of the distal femoral metaphysis. PAI-1 inactivation did not affect bone metabolism in vehicle-treated mice. Treatment with 1,25(OH)(2)D(3) induced bone resorption similarly in PAI-1-/- and WT mice, as assessed by the increase in the urinary excretion of calcium (2. 2 fold and 2.3-fold, respectively) and of pyridinoline crosslinks (by 24% and 22%, respectively). In addition, a comparable reduction in bone mass was observed in PAI-1-/- and WT mice after treatment with 1,25(OH)(2)D(3), as evidenced by the decrease in the femoral calcium content (by 25% and 32%, respectively), in the trabecular bone volume (by 50% and 40%, respectively), in the trabecular mineral content (by 17% and 15%, respectively), and in the cortical mineral content (by 45% and 52%, respectively). The parameters of bone turnover also increased after 1,25(OH)(2)D(3) treatment. Serum osteocalcin was, respectively, 25% and 28% higher in PAI-1-/- and WT mice treated with 1,25(OH)(2)D(3) compared with the mice injected with vehicle. Similarly, the osteoid surface increased in 1, 25(OH)(2)D(3)-treated PAI-1-/- and WT mice by 40% and 51%, respectively, the mineral apposition rate increased by 15% and 8%, respectively, and the bone formation rate by 54% and 48%, respectively. These data indicate that PAI-1 is not critical during bone resorption induced by 1,25(OH)(2)D(3) in vivo. PMID- 10865216 TI - Hypercalcemia during the osteogenic phase after rat marrow ablation coincides with increased bone resorption assessed by the NTx marker. AB - Marrow ablation is a model of bone turnover in which the excavated tibial intramedullary cavity is rapidly and reproducibly filled by osteoblasts with new woven bone (days 6-8), which is then rapidly resorbed by osteoclasts (days 10 15). We showed previously (Magnuson et al., 1997) that marrow ablation induces a dramatic hypercalcemia and hypercalciuria in rats that unexpectedly peaked at the time of maximal osteogenesis and continued throughout the subsequent resorption phase. Based upon the amount of calcium mobilized and a peak of urinary hydroxyproline, we suggested that the hypercalcemia and hypercalciuria were due to increased systemic osteoclastic bone resorption induced by marrow ablation. We now apply a new enzyme-linked immunosorbent assay for rodent alpha(2)(I) N telopeptide (NTx), a marker of bone resorption, to the marrow ablation model to demonstrate that excretion of NTx parallels that of calcium release in the operated control group. Specifically, maximal NTx/creatinine excretion coincides with the onset of hypercalcemia on days 7-8. A peak of NTx was also observed in methylprednisolone- and deflazacort-treated ablated animals. Analyses for urinary free deoxypyridinoline crosslink failed to detect a significant ablation-induced change in excretion. Interleukin 6 activity was increased in all operated control and glucocorticoid-treated groups after marrow ablation, whereas serum parathyroid hormone remained at presurgical levels in operated controls throughout the 15-day study period. The NTx results confirm that bilateral tibial marrow ablation induces a burst of extratibial bone resorption and hypercalcemia 7-8 days later. We have estimated that the osteogenic phase of the ablation model deposits 40 mg of calcium as hydroxyapatite crystals within the intramedullary cavity on days 6-8; this represents 33%-50% of the total blood calcium content of a young rat. We hypothesize that the size and rapidity of this demand for ionized calcium is met through an extratibial bone resorption pathway of osteoclast formation and activation that anticipates and fulfills this need, and that is initiated at the time of marrow ablation. PMID- 10865217 TI - Stochastically simulated assessment of anabolic treatment following varying degrees of cancellous bone resorption. AB - The aim of this study was to investigate the recovery in cancellous bone stiffness resulting from anabolic treatment following varying degrees of resorption, using a stochastic simulation applied to a simplistic structure consisting of five vertical and five horizontal trabeculae. The structure was initially resorbed, and "bone" elements were stochastically removed until nominal resorptions of 10%, 15%, 20%, 25%, and 30% were achieved. A stochastic simulation of anabolic treatment was then applied where bone elements were added, continuing until the original stiffness had been regained, for example, simulating treatment of a patient with an anabolic agent after a period of postmenopausal resorption. The resorption and anabolic simulations were repeated three times for each of the nominal resorptions. The stiffness of the bone structure decreased linearly with resorption, with a slope of approximately -2 and an R(2) of 97.0%; hence, the stiffness fell at approximately twice the rate of the reduction in density. When the various structures regained their original density, the resultant stiffness also had a linear relationship with the original resorption, with a slope of -1 and a lower R(2) of 86.1%. This implies that the reduction in stiffness, when original density was regained, fell proportionately with the degree of initial resorption and, therefore, after a resorption of 30%, when original density was regained, the stiffness of the resultant structure was approximately 30% less than that of the original structure. The density required for the original stiffness to be regained increased linearly with the degree of initial resorption, with a slope of approximately 0.5 and an R(2) of 65.2%, lower than that observed for the previous relationships. This indicates a greater spread of data and suggests greater variability in the formation phase beyond the point of regained original density. Because irreversible connectivity reduction is widely considered to be one of the earliest manifestations of estrogen loss, these findings, although obtained on a simulation of a simplistic cancellous bone structure, support the concept of early intervention to prevent potentially irreversible deterioration of trabecular architecture after menopause. PMID- 10865218 TI - Effects of two intermittent alendronate regimens in the prevention or treatment of postmenopausal osteoporosis. AB - In clinical practice, a large proportion of patients have bone mass values for which a therapeutic intervention is considered necessary, but the accepted aim might be the sole preservation or marginal increases of the actual bone mass. These goals might be achieved with lower or intermittent doses of a powerful agent for the purpose of fewer side effects and improved compliance. The aim of this study was to assess the effects of two intermittent alendronate regimens in the treatment of postmenopausal osteoporosis. One hundred twenty-four postmenopausal women (age range 52-75 years, at least 7 years since last menopause) with a bone mineral density (BMD) at either the femoral neck or lumbar spine of 2 SD below the mean values of young healthy individuals, and without a history of previous osteoporotic fracture, were randomly assigned either to calcium/vitamin D supplements, alone or associated with two different intermittent oral alendronate regimens: 20 mg once a week (weekly alendronate group) or 10 mg daily (orally) for 1 month out of 3 (cyclical alendronate group). After 1 year, in both groups given intermittent alendronate, we observed significant increases in BMD at both the spine (+2.2 +/- 2.6 and +2.5 +/- 2.9) and femoral neck (+1.6 +/- 4.8 and +1.5 +/- 2.2) for the weekly and cyclical regimens, respectively. This was associated with a significant diminution of both serum bone-specific alkaline phosphatase and urinary N-telopeptides of collagen type I excretion. In the patients in the control group BMD decreased significantly at the lumbar spine, with a slight decline of serum bone-specific alkaline phosphatase. Compliance with treatment and drug tolerability were good in both alendronate groups. In conclusion, intermittent alendronate administration at cumulative doses (and costs) three times lower than those currently recommended for osteoporosis treatment is very well accepted, and is able to significantly increase BMD at the spine and femoral neck and to decrease the markers of bone turnover. These regimens can be clinically useful in the long term treatment of postmenopausal osteoporosis without prevalent osteoporotic fractures, particularly in women with lower compliance for continuous administration. PMID- 10865219 TI - Cyclical etidronate versus sodium fluoride in established postmenopausal osteoporosis: a randomized 3 year trial. AB - To compare the effects of sodium fluoride and etidronate in severe postmenopausal osteoporosis, we conducted a 3 year, prospective, trial in 118 postmenopausal osteoporotic women with at least one vertebral fracture, who were randomly assigned to receive sodium fluoride (25 mg twice daily, as enteric-coated tablets) plus calcium (1000 mg/day) or intermittent etidronate (400 mg/day for 14 days) followed by calcium (1000 mg/day for 76 days). Lateral spine X-ray films and dual-energy X-ray absorptiometry (DXA) measurements of the lumbar spine and proximal femur were performed at enrollment and yearly. Nonvertebral fractures were recorded every 6 months. Thirty-one women in the fluoride group and 47 in the etidronate group completed the trial. At 36 months, the mean change from baseline of the lumbar bone density in the fluoride group was 8.5 +/- 2.04% (p = 0.001) and in the etidronate group was of 3.6 +/- 0. 84% (p < 0.001). The changes in the fluoride group were significantly higher than in the etidronate group (p = 0.01). Both groups showed nonsignificant changes in femoral neck bone density. There was no significant difference between groups in the cumulative proportion of women with new vertebral fractures, with an incidence in the fluoride group of 16% vs. 17% in the etidronate group. However, the number of new vertebral fractures was significantly lower in the fluoride group (6 fractures) than in the etidronate group (19 fractures) (p = 0.05). The number of patients with nonvertebral fractures was similar in both groups. A high incidence of side effects, mainly gastrointestinal symptoms and lower extremity pain syndrome, was observed in the fluoride group. Etidronate was well tolerated. We conclude that, in women with severe osteoporosis, although sodium fluoride is more favorable than cyclical etidronate for increasing lumbar bone mass, no differences were observed in the incidence of fractures. PMID- 10865220 TI - A new method of comprehensive static histomorphometry applied on human lumbar vertebral cancellous bone. AB - The aim of the present study was to assess age-related changes in the human spine by use of established static histomorphometry, and to determine how these static histomorphometric measures are interrelated in human cancellous bone tissue. The material comprised normal human lumbar vertebral bodies (L-2) from 12 women (19 96 years) and 12 men (23-91 years) selected from a larger autopsy material to give an even age and gender distribution. In addition, L-2 from three female subjects (80, 88, and 90 years) with a known vertebral fracture of L-2 were considered. Approximately 9-mm-thick frontal (mediolateral) slices were embedded in methylmetacrylate, stained with aniline blue, and scanned into a computer with a flatbed image scanner at a high resolution (2400 dpi). With a custom-made computer program the following static histomorphometric measures were determined: trabecular bone volume; marrow space star volume; bone space star volume; anisotropy of bone and marrow phase (star length distribution method); node-strut analysis (node:terminus ratio); trabecular thickness; trabecular number; trabecular separation; and trabecular bone pattern factor. In addition, connectivity density was determined (by the ConnEulor method). All 11 histomorphometric measures, except bone space star volume and the two measures of anisotropy, showed a significant correlation with age. Marrow space star volume (r = 0.82) and trabecular bone volume (r = -0.81) showed the highest correlation with age. Furthermore, it was found that all of the histomorphometric measures were correlated, to different degrees. Trabecular bone volume correlated significantly with all ten histomorphometric measures, whereas the two anisotropy measures were poorly correlated to the other measures. Finally, we found the histomorphometric values in this study to be in excellent accordance with various previously published results from studies of human trabecular vertebral bone, the sole exception being marrow space star volume, which was probably due to the small (artificial) region of interest (ROI) that was used in the earlier studies. In conclusion, the new method applied herein allows for easy assessment of age related changes and also for assessment of relationships between histomorphometric measures in human vertebral cancellous bone. PMID- 10865221 TI - Precision of quantitative ultrasound: comparison of three commercial scanners. AB - Quantitative ultrasound (QUS) measurements of bone have been shown to be independent predictors of osteoporotic fracture risk. Drawbacks of this technique have included the precision of the scanners, which is said to be poorer than in dual-energy X-ray absorptiometry (DXA), in part due to difficulty in repositioning of the foot in an os calcis system and difficulty in comparison across different technologies. A new type of QUS scanner has been introduced that produces an image of the area scanned and is believed to improve precision by aiding repositioning. In this study, we compare three scanners: a dry system (McCue CUBA Clinical); a nonimaging water-bath system (Lunar Achilles(+)); and an imaging water-bath system (Osteometer DTU-One). Short-term phantom precision was calculated by repeating measurements ten times in succession on the manufacturer supplied phantom. Long-term phantom precision was calculated by examining the phantom measurements over a 6 month period. In vivo precision was calculated in 26 normal volunteers (19 women, 7 men) and 20 women with osteoporosis. Monitoring time intervals (MTIs) were also calculated using the manufacturer's normative database. The MTI is the period between scans required to show that a "true" change has occurred, and was between 0.5 year for stiffness (a derived index produced by the Lunar Achilles instrument) and >5 years for all other measurements. The imaging system did not seem to improve precision. Precision for the QUS phantom was similar to that of DXA with a coefficient of variation (CV) of around 1.5% for BUA and <1% for speed of sound (SOS). The precision was such that the technique may be considered for monitoring skeletal changes. However, the change of bone mass at the os calcis in response to treatment was slow, which made the time needed to wait before assessing change, on the whole, longer than that for DXA. An exception may be the Lunar Achilles "stiffness" parameter, but this can only be determined in a longitudinal, comparative treatment study. PMID- 10865222 TI - The relation between bone density, free androgen index, and estradiol in men 60 to 70 years old. AB - The cause of age-related bone loss in men is poorly understood. Previous studies of the relationship between bone density and serum androgens have yielded inconsistent results, perhaps partly because age is a determinant of both. Recent studies suggest that serum estrogen levels influence bone density in adult men. In order to determine whether bone mineral density (BMD) and bone turnover are associated with serum sex steroids, we investigated 37 normal men within a narrow age range (60-70 years). Bone mineral density at the forearm, hip, and spine, testosterone, sex hormone binding globulin (SHBG), free androgen index (FAI:T/SHBG), estradiol (E), free estradiol index (FEI:E/SHBG), and markers of bone formation (alkaline phosphatase, osteocalcin, procollagen type I C-terminal extension peptide) and bone resorption (hydroxyproline/creatinine [OHPr/Cr], deoxypyridinoline/creatinine [Dpd/Cr], pyridinoline/creatinine, collagen type I cross-linked telopeptide) were measured. Bone mineral density was positively related (r > 0.35, p < 0.05 at all sites) to log FAI, whereas there was no significant relationship between BMD and either serum total testosterone, serum E, or FEI. Bone density at the spine and hip were inversely related to both OHPr/Cr (r > -0.41, p < 0.05 for all sites) and Dpd/Cr (r > -0.36, p < 0.05 for all sites). OHPr/Cr (r = -0.41, p < 0.05) and Dpd/Cr (r = -0.41, p < 0.05) were both inversely related to log FAI. We conclude that BMD and bone turnover in adult men are related to plasma free androgens. PMID- 10865224 TI - Calcitonin receptor mRNA in mononuclear leucocytes from postmenopausal women: decrease during osteoporosis and link to bone markers with specific isoform involvement. AB - Calcitonin inhibits bone resorption via its receptor (CTR) on osteoclasts. Two hCTR isoforms, hCTR1 and hCTR2, give proteins that differ in their structure and signaling pathways. We investigated whether specific isoforms or quantitative changes in total hCTR mRNA were associated with high bone resorption and turnover in menopause or osteoporosis. The hCTR mRNA in mononuclear blood cells of premenopausal (PreM), healthy (PostM), and osteoporotic (OsteoP) postmenopausal women was assessed using reverse-transcriptase polymerase chain reaction. hCTR1 and hCTR2 were investigated for 59 total RNA samples, and semiquantitative analysis of total hCTR mRNA was performed for 71. Serum calcitonin, free urinary deoxypyridinoline (D-Pyr), serum bone alkaline phosphatase (SBAP), and osteocalcin (SOC) were also evaluated. Serum calcitonin levels did not differ in PostM and OsteoP. The prevalence of each isoform was similar in the three groups. Healthy postmenopausal women and OsteoP with hCTR2 had lower bone turnover (D Pyr: 6.79 +/- 0.54, n = 25; SBAP: 11.63 +/- 1.47, n = 26; SOC: 8.31 +/- 0.58, n = 26) than those without hCTR2 (D-Pyr: 9.90 +/- 1.95, n = 5; SBAP: 21 +/- 5.19, n = 5; SOC: 11.9 +/- 2.10, n = 5; p < 0.05). Total hCTR mRNA levels were not different in PreM and PostM. By contrast, values were strikingly lower in OsteoP (0.57 +/- 0.17, n = 28) than in PostM (2. 25 +/- 0.61, n = 19, p < 0.05) and negatively correlated with bone markers values in both. We suggest that a specific isoform and amounts of total hCTR mRNA are linked to increased bone resorption in postmenopausal osteoporosis. PMID- 10865223 TI - Hip geometry, bone mineral distribution, and bone strength in European men and women: the EPOS study. AB - Hip geometry and bone mineral density (BMD) have been shown previously to relate, independently of each other, to risk of hip fracture. We used Lunar DPX "beta" versions of hip strength analysis (HSA) and hip axis length (HAL) software to analyze scans from ten representative age-stratified population samples in the European Prospective Osteoporosis Study (EPOS). All 1617 subjects were >50 years of age, and 1033 were women. The data were modeled with gender and center as categorical variables. The bone mineral density of the upper half of the femoral neck declined at a faster rate with age than that in the lower half. Femoral neck cross-sectional moment of inertia (CSMI), a measure of resistance to bending, showed no significant age reduction in either gender. However, height and weight effects on CSMI were significantly more beneficial in men than in women (0.002 < p < 0.012) and the weight effect appeared to be mediated by bone mineral content (BMC). Compressive stress (Cstress), defined as the stress in the femoral neck at its weakest cross section arising from a standardized fall, was higher in women. Although Cstress increased with body weight when BMC was held constant, in practice it fell through the association and statistical interaction of rising body weight with rising BMC. HAL, as expected, was strongly positively associated with male gender and also height (p < 0.0001). Hip strength-related indices were markedly center-dependent. Significant differences (p < 0.0001) were noted between the centers for all the variables investigated that related to hip geometry. Adjustment for femoral neck bone mineral content (totBMC) showed these center differences to account for >50% of center variation in hip strength, which remained highly significant (p < 0.0001). We conclude that there are substantial geographical differences in femoral neck geometry as well as in BMD. These geometric variations may contribute to the large variations in hip fracture risk across Europe. The effects of aging on hip strength need to be explored in longitudinal studies. PMID- 10865225 TI - Sib pair linkage and association studies between bone mineral density and the interleukin-6 gene locus. AB - A major determinant of the risk for osteoporosis in later life is bone mineral density (BMD) attained during early adulthood. Bone mineral density is a complex trait that, presumably, is influenced by multiple genes. Interleukin-6 (IL-6) is an attractive candidate gene for osteoporosis susceptibility, because it has effects on bone cells and has been implicated in the pathogenesis of osteoporosis. Furthermore, previous investigators have identified an association between a 3' UTR polymorphism of the IL-6 gene and BMD. In this study, we searched for linkage and association between this IL-6 gene polymorphism and peak BMD in a large population (812 individuals) of healthy premenopausal sibpairs. Although previous investigators identified only 6 IL-6 alleles, we identified 17 alleles by modifying electrophoretic conditions and evaluating a very large population. We found no evidence for either linkage or association between the IL 6 gene locus and BMD of the spine or hip in either Caucasians or African Americans. PMID- 10865226 TI - Antisperm antibodies associated with infertility: properties and encoding genes of target antigens. AB - Infertility among couples of reproductive age is a perplexing condition when the cause is indeterminate. These cases are classified as unexplained infertility. In a subset of subjects, antisperm antibodies with sperm agglutinating and/or immobilizing activities have been detected in the blood or fluids of the reproductive tract. These cases are designated as immunologic infertility although a cause and effect relationship of the antibodies to infertility has not been established. In this review, seven target sperm antigens to antibodies associated with infertility and their encoding genes are described. The antisperm antibodies (ASAs) examined were obtained from infertile women or were monoclonal antibodies (mAb) raised against human sperm proteins. All the ASAs studied possessed potent sperm agglutinating and/or immobilizing activities. The target antigens were isolated from human and other mammalian sperm, and the encoding genes identified. The seven antigens are YWK-II, BE-20, rSMP-B, BS-63 (nucleoporin-related), BS-17 (calpastatin), HED-2 (zyxin), and 75- kDa. Each antigen is a distinct and separate entity and is produced by different cells of the reproductive tract, (e.g., germ cells, epididymal epithelial cells, and Sertoli cells). No single predominant target component has been found to interact with the ASAs. It is proposed that immunologic infertility is the consequence of the combined actions of multiple ASAs in immobilizing and/or agglutinating spermatozoa, blocking spermegg interaction, preventing implantation, and/or arresting embryo development. PMID- 10865227 TI - Skeletal muscle as a myocardial substitute. AB - Skeletal muscle has long been used in the field of cardiac surgery. Its use has progressed from providing myocardial reinforcement to assisting the heart by actively pumping blood. Early experiments revealed that skeletal muscle assistance could augment pressures and blood flow; however, the results were short-lived due to muscle fatigue. It was later shown that skeletal muscle can be conditioned electrically to be fatigue resistant and therefore may be useful for performing cardiac-type work. Once the details were formed of how to stimulate and manipulate the muscle to assist the heart, several configurations were devised. Cardiomyoplasty and aortomyoplasty refer to wrapping skeletal muscle around the heart or aorta, respectively. These techniques have been applied in humans; however, the effectiveness is controversial. Although most patients improve clinically, the hemodynamic parameters have not shown consistent improvements, and survival data are unknown. Skeletal muscle ventricles offer a promising alternative to both cardiomyoplasty and aortomyoplasty. These are completely separate pumping chambers constructed from skeletal muscle and connected to the circulation in a variety of configurations. Although these have not been tried in humans, the animal data appear quite convincing. The skeletal muscle ventricles have shown the greatest improvements on hemodynamic parameters with great stability over time. PMID- 10865228 TI - Differential effects of nicotine and aging on splenocyte proliferation and the production of Th1- versus Th2-type cytokines. AB - Nicotine has a multitude of biological actions in the central and peripheral nervous systems where nicotinic acetylcholine receptors are found. Nicotinic acetylcholine receptors have also been identified on immune cells, but the effects of nicotine on immune responses are not well characterized. These studies tested the hypotheses that nicotine has an effect on both T-lymphocyte proliferation and the production of cytokines by activated T cells, processes that are necessary for effective T-cell-mediated immune responses. In addition, the effects of nicotine on these immune responses in aging animals and the effects of nicotine exposure prior to immunostimulation were investigated. Murine splenocytes were exposed to nicotine and stimulated with concanavalin A (ConA). The highest concentration of nicotine (128 microg/ml) significantly depressed proliferation of T cells both when nicotine and ConA were added concurrently and when nicotine was added 3 hr prior to ConA. Nicotine, added concurrently with ConA at concentrations between 0. 25 and 64 microg/ml, significantly inhibited the production of IL-10 by splenocytes from young adult mice, whereas the inhibition of production of IL-10 by splenocytes from old mice was significantly inhibited, but the response was more variable, depending on the nicotine concentration. In contrast, the production of IFN-gamma by splenocytes from either young adult or old mice was not affected when nicotine (0.016-64 microg/ml) was added concurrently with ConA. Pre-exposure to 1 microg/ml of nicotine for 3 hr significantly enhanced the production of IFN-gamma by splenocytes from young adult mice, whereas pre-exposure to 0.016 microg/ml of nicotine tended to but did not significantly enhance IFN-gamma production. Nicotine is now being used as an over-the-counter drug by people who differ in age and general immunocompetence. Therefore, the effects of nicotine on immune responses, independent from the effects of the other chemicals found in tobacco, need to be investigated. PMID- 10865229 TI - Differential effects of maturation on nicotinic- and muscarinic receptor-induced ion secretion in guinea pig distal colon. AB - The incidence of constipation increases with age. This has been linked to age related changes in the structure and function of myenteric neurons regulating intestinal motility; however, the role of submucous neurons is unknown. The aim of this study was to determine the effect of maturation on cholinergic receptor induced ion secretion in guinea pig colon. Changes in the short-circuit current (Isc) and tissue conductance were monitored in muscle-stripped colonic segments from young (3-4-month-old) and mature (12-15-month-old) male guinea pigs. Thirty one percent of colonic segments from young guinea pigs exhibited ongoing neural activity, which was absent in mature animals. Baseline Isc was significantly higher only in young guinea pig tissues with ongoing activity. Tissue conductance was similar in all tissues. Electrical field stimulation caused a biphasic increase in the Isc. At 15 V/10 Hz, only Peak 1 was attenuated, whereas both peaks were reduced in mature guinea pigs at 10 V/5Hz. 1,1, dimethyl-4-phenyl piperazinium(DMPP)-induced ion secretion was blunted in mature guinea pigs. Atropine reduced the 1,1, dimethyl-4-phenyl-piperazinium response only in young guinea pigs. Carbachol-induced ion secretion was similar in tissues from both age groups. In conclusion, nicotinic receptor-induced secretion mediated by both cholinergic and noncholinergic secretomotor neurons was blunted; however, epithelial muscarinic receptor activity was unaltered during maturation. PMID- 10865231 TI - Branched-chain amino acid-enriched diet: effects on insulin secretion and cellular immune aggression. AB - Several reports have demonstrated that high-protein diets may have beneficial effects on experimental models of diabetes and have raised the possibility that branched-chain amino acids could play a role in these protective effects. We investigated the effect of a normoproteic, branched-chain amino acid-enriched diet (experimental diet) on insulin secretion from C57BL/6N mice transferred with splenocytes from diabetic syngeneic donors. Mice previously fed with the experimental or control diet received three intraperitoneal injections, every other day, of 5 x 107 viable mononuclear splenocytes obtained from control or diabetic donors. Results showed that mice fed with the experimental diet and transferred with "diabetic" splenocytes presented: i) normoglycemia, and (ii) significantly higher levels in both phases of glucose-induced insulin secretion and normal values of arginine-glucose-induced insulin secretion. To evaluate the in vitro cellular immune aggression, dispersed mouse islet cells were co-cultured with splenocytes from syngeneic diabetic mice. First, dispersed islet cells from mice on the experimental or control diet were co-cultured with splenocytes from control or diabetic mice on a commercial diet. In the presence of "diabetic splenocytes, dispersed islet cells from mice on the experimental diet presented a significantly lower in vitro cellular immune aggression. On the other hand, "diabetic" splenocytes from mice fed with the experimental diet produced a significantly reduced cellular immune aggression on dispersed islet cells. Our results showed that feeding branched-chain amino acids increased the capacity of beta cells to withstand a functional assault and diminished the extent of in vitro cellular immune aggression. PMID- 10865230 TI - The possible role of prolactin in the circadian rhythm of leptin secretion in male rats. AB - In humans there is a circadian rhythm of leptin concentrations in plasma with a minimum in the early morning and a maximum in the middle of the night. By taking blood samples from adult male rats every 3 hr for 24 hr, we determined that a circadian rhythm of plasma leptin concentrations also occurs in the rat with a peak at 0130h and a minimum at 0730h. To determine if this rhythm is controlled by nocturnally released hormones, we evaluated the effect of hormones known to be released at night in humans, some of which are also known to be released at night in rats. In humans, prolactin (PRL), growth hormone (GH), and melatonin are known to be released at night, and adrenocorticotropic hormone (ACTH) release is inhibited. In these experiments, conscious rats were injected intravenously with 0.5 ml diluent or the substance to be evaluated just after removal of the first blood sample (0.3 ml), and additional blood samples (0.3 ml) were drawn every 10 min thereafter for 2 hr. The injection of highly purified sheep PRL (500 microg) produced a rapid increase in plasma leptin that persisted for the duration of the experiment. Lower doses were ineffective. To determine the effect of blockade of PRL secretion on leptin secretion, alpha bromoergocryptine (1.5 mg), a dopamine-2 receptor agonist that rapidly inhibits PRL release, was injected. It produced a rapid decline in plasma leptin within 10 min, and the decline persisted for 120 min. The minimal effective dose of GH to lower plasma leptin was 1 mg/rat. Insulin-like growth factor (IGF-1) (10 microg), but not IGF-2 (10 microg), also significantly decreased plasma leptin. Melatonin, known to be nocturnally released in humans and rats, was injected at a dose of 1 mg/rat during daytime (1100h) or nighttime (2300h). It did not alter leptin release significantly. Dexamethasone (DEX), a potent glucocorticoid, was ineffective at a 0. 1-mg dose but produced a delayed, significant increase in leptin, manifest 100-120 min after injection of a 1 mg dose. Since glucocorticoids decrease at night in humans at the time of the maximum plasma concentrations of leptin, we hypothesize that this increase in leptin from a relatively high dose of DEX would mimic the response to the release of corticosterone following stress in the rat and that glucocorticoids are not responsible for the circadian rhythm of leptin concentration. Therefore, we conclude that an increase in PRL secretion during the night may be responsible, at least in part, for the nocturnal elevation of leptin concentrations observed in rats and humans. PMID- 10865232 TI - Effects of cholesterol diets on vascular function and atherogenesis in rabbits. AB - Vascular endothelial dysfunction is an important early event in atherogenesis. To evaluate the effects of different levels of cholesterol-containing diets on vascular function and atherogenesis, 17 New Zealand White male rabbits were randomized into four groups: Control with noncholesterol, 10-week 0.5% (0.5C-10) or 1% cholesterol (1C-10), and 14-week 0.5% cholesterol (0.5C-14) feedings. After 10 or 14 weeks, the aortas were harvested for studies of vascular endothelial function and percentage surface lipid lesions. The 0.5% and 1% cholesterol feedings resulted in the same degree of hypercholesterolemia independent of the level and period of cholesterol feeding. There was a decreased trend in vascular endothelial-dependent relaxation to acetylcholine in cholesterol-fed rabbits. Fourteen-week cholesterol feeding induced the least vascular dilation at a concentration of 10-7 M acetylcholine (-38 +/- 3%, -23 +/- 4%, -23 +/- 2%, and 15 +/- 5% in control, 0.5C-10, 1C-10, and 0.5C-14 groups, respectively, P = 0.003). More cumulative exposure of arterial walls to cholesterol induced more surface lipid lesions in the aorta (r = 0.877, P < 0.001). There was a negative relationship between aortic lesions and vasodilation (r = -0.557, P = 0.020 for calcium ionophore; r = -0.463, P = 0.062 for acetylcholine). We conclude that the 0.5% and 1% cholesterol feedings induce similar degrees of hypercholesterolemia. However, aortic lipid lesions and vascular reactivity are dependent on cumulative exposure to cholesterol rather than serum cholesterol level only. Furthermore, decreased vascular endothelial relaxation in cholesterol-fed rabbits was related to lipid plaques in the aorta. PMID- 10865233 TI - Tissue-specific expression of the uncoupling protein family in streptozotocin induced diabetic rats. AB - The vulnerability of streptozotocin (STZ)-induced diabetic rats to cold stress has been established. One of the elements controlling body temperature is thermogenesis, in which uncoupling protein (UCP) is known to play an important role. We have examined UCP2 and UCP3 expressions in brown adipose tissue (BAT), white adipose tissue (WAT), and skeletal muscle (MSL) during the acute and chronic phases of STZ-induced diabetes in rats. The long-term effect and the effect of insulin treatment thereafter were also unexplored previously and are examined in this study. In the acute phase of diabetes (2.5 days after STZ injection), UCP2 gene expression in BAT, WAT, and MSL, and UCP3 expression in the muscle were significantly increased. In the chronic phase of diabetes (21 days after STZ injection), UCP2 and UCP3 expression in the MSL were restored to the control levels without insulin supplementation. UCP2 in BAT and WAT remained high in the chronic phase, whereas UCP3 expression in BAT and WAT, which did not change in the acute phase, was significantly decreased. Insulin supplementation restored UCP2 expression in BAT and WAT, but over-corrected UCP3 in WAT above the control and did not affect UCP3 expression in BAT. Insulin supplementation depressed UCP3 expression in the MSL below control. These results indicate that the effects of STZ-induced diabetes on UCPs gene expression are tissue-specific as well as dependent on the duration of diabetes. PMID- 10865234 TI - Albumin facilitates zinc acquisition by endothelial cells. AB - Albumin has long been observed to have a marked influence on the delivery of zinc to cells, but the mechanism of the interaction remains elusive. We examined whether albumin facilitates the acquisition of zinc by endothelial cells. Cultures of endothelial cells were used to analyze binding and acquisition of zinc and albumin to test this interaction. Our results indicate that albumin plays a role in facilitating the physiological delivery of zinc to endothelial cells. Albumin receptors that preferentially recognize albumin molecules carrying a zinc atom were demonstrated on the endothelial cell surface. Endocytosis is instrumental in albumin uptake, which was also consistently true of zinc uptake. Zinc and albumin were acquired by the cells in a 1:1 molar stoichiometry during the first 20 min of incubation in a medium with equimolar concentrations of zinc and albumin. The amount of albumin associated with the cells stabilized after 30 min, whereas the amount of zinc continued to increase. One possible explanation for this result is that a physiological route for zinc delivery into endothelial cells is by co-transport with albumin via receptor-mediated endocytosis. PMID- 10865235 TI - Effects of inositol, LiCl, and heparin on the antibody responses to SRBC by normal and immunodeficient XID mice. AB - Spleen cells from naive adult immunocompetent and immunodeficient XID mice were cultured on agar containing sheep red blood cells (SRBC) with and without myo inositol, scyllo-inositol, lithium chloride, or heparin, and after 1 or 2 days the number of colonies of antiSRBC antibody-forming cells (PFC) were determined. It was found that myo-inositol and scyllo-inositol at one-tenth the concentration were equally effective in increasing the number of specific PFC. Myo-inositol, scyllo-inositol, and lithium chloride accelerated the appearance of direct foci in cultures of spleen cells from normal and XID mice. When heparin was added to cultures of XID spleen cells, PFC were found to be increased on Day 1; however, PFC and foci were not increased in cultures of spleen cells from competent mice until 1 day later. The addition of combinations of these agents to cultures of spleen cells had no positive or negative effect on the generation of foci or PFC. Normal mice given heparin intraperitoneally with SRBC had increased splenic PFC on Days 3 and 4 but not on Day 7. The results suggest that these agents modulate B-cell responses by increasing the rate of proliferation and/or secretion through a signaling pathway(s) distal to, or more likely, independent of Bruton's tyrosine Kinase (BTK). It is not clear that the mechanism is the same with each agent. PMID- 10865237 TI - Work-related vision hazards in the dental office. AB - Among the numerous threats to the dentist's health there is one relating to the eye. The paper discusses the impact of selected adverse factors on the eye in connection with dental practice in the surgery. PMID- 10865236 TI - Tumor necrosis factor induces resistance of macrophages to Legionella pneumophila infection. AB - Legionella pneumophila is an ubiquitous opportunistic intracellular pathogen that replicates readily in thioglycollate-elicited peritoneal macrophages from genetically susceptible A/J mice. Treatment of macrophage cultures in vitro with tumor necrosis factor-alpha (TNF-alpha) induced resistance of the macrophages to infection by Legionella as compared with control macrophages treated with medium alone. Addition of small amounts of monoclonal antibody to TNF-alpha restored susceptibility of the macrophages. Furthermore, antibody to the proinflammatory cytokine interleukin-1 (IL-1) alpha/beta increased resistance, but recombinant IL 1 had little effect. Such decreased susceptibility to Legionella growth in anti IL-1 antibody-treated cultures corresponded with enhanced levels of TNF-alpha in the supernatants of the treated cells. An antibody to another proinflammatory cytokine with known immunoregulatory properties (i.e., IL-6) had little or no effect on the ability of the macrophages to be infected by Legionella and, furthermore, treatment with recombinant IL-6, similar to recombinant IL-1, did not modify the ability of the cells to be infected in vitro. These results indicate that TNF-alpha is important in controlling L. pneumophila replication, and IL-1 can regulate TNF-alpha levels, affecting susceptibility of macrophages to infection with an intracellular opportunistic pathogen like Legionella. PMID- 10865238 TI - Analysis of airborne pollen fall in Balikesir, Turkey, 1996-1997. AB - In this study, pollen grains were identified by use of Durham sampler in the atmosphere of Balikesir in 1996 and 1997. During these two years, a total of 17,256 pollen grains per cm(2) were recorded. A total of 8,576 pollen grains per cm(2) were identified in 1996 and a total of 8,680 pollen grains per cm(2) in 1997. Pollen fall in the years 1996-1997 comprised grains belonging to 50 taxa and unidentified pollen grains. Of these 50 taxa, 30 belonged to arboreal and 20 to non-arboreal plants. Total pollen grains consisted of 70.92% grains from arboreal plants, 24.87% grains from non-arboreal plants and 4.21% unidentified pollen grains. In the region investigated, Pinus spp., Cupressaceae/Taxaceae, Gramineae, Platanus spp., Quercus spp., Olea spp., Salix spp., Urticaceae, Moraceae, Plantago spp., Chenopodiaceae/Amaranthaceae, Ailanthus spp., Juglans spp., Carpinus spp. and Rosaceae released the greatest amounts of pollens. During the study period, the pollen fall reached its highest level in May. PMID- 10865239 TI - Finnish farmers' self-reported morbidity, work ability, and functional capacity. AB - The aim of the study was to evaluate Finnish farmers' self-reported morbidity, especially musculoskeletal disease and disabilities, work ability, physical fitness, and functional capacity. A further goal was to identify the group of farmers that most need a means to promote their work ability. The data were collected with a computer-assisted telephone interview. The study population comprised of 577 full-time farmers (296 men and 281 women). The results have been expressed as odds ratios with 95% confidence intervals determined in a logistic regression analysis. The farmers with the greatest need for activities that support and promote work ability are those over 34 years of age, female farmers, farmers with fewer than 10 years of education, farmers from farms with fewer than 20 hectares of cultivated land, farmers who milk cows regularly, and depressed farmers. PMID- 10865240 TI - Bronchopulmonary pathology in workers exposed to organic fodder dust. AB - The purpose of this study was to investigate work-related respiratory symptoms, the prevalence of chronic lung diseases and ODTS, and to study the lung function and bronchial hyperresponsiveness in the workers of Ukrainian fodder production facilities. 240 workers of two Ukrainian fodder production plants have been examined. Dust concentrations in the air of working zone were different, reaching 48.2 mg/m(3) in the first plant and 16.8 mg/m(3) in the second. Endotoxin levels were 240.0 ng/m(3) and 1.8 ng/m(3) respectively. The length of service at the first plant exceeded 2 times that at the second. In the actual research the investigation of respiratory symptoms, lung function and bronchial reactivity was carried out. A comparison between animal feed workers and internal controls revealed work-related symptoms. The predominant symptomatic and lung function effects indicate a clinical picture related to chronic bronchitis. The prevalence of chronic bronchitis was 26.4 +/- 4.0% at the first plant and 8.8 +/- 4.8% at the second one (p < 0.01). The prevalence of respiratory troubles was related to dust exposure more strongly that to smoking. 39.7 +/- 4.4% of exposed workers at the first plant and 14.7 +/- 6.0% at the second one revealed organic dust toxic syndrome (ODTS). In 47.9 +/- 7.2% of workers with ODTS, this syndrome was associated with chronic bronchitis. Examination of lung function revealed obstructive changes which were more expressed in exposed workers of the first plant. Lung function clearly decreased with increasing duration of employment. Obstruction of small bronchi and bronchial hyperresponsiveness (registered in 74.7% of workers) were the early signs of respiratory troubles in exposed workers. PMID- 10865241 TI - Organochlorine pesticides concentration in the drinking water from regions of extensive agriculture in Poland. AB - Considerable amounts of organochlorine pesticides (DDT, lindane, heptachlor and methoxychlor) were detected in drinking water samples during a two-year study in Warka-Grojec rural region of Poland. The average incidence of drinking water sources with detectable levels of pesticides ranged from 20 to 30%. According to Polish regulations concerning water quality, individual concentrations did not reach the maximum admissible level. However, the levels of pesticides in most cases exceeded the maximum admissible concentrations recommended by the European Union PMID- 10865242 TI - Chronic bronchitis in rural and industrial areas. AB - The purpose of the study was the determination of prevalence of chronic bronchitis in people older than 55 years in rural and industrial areas. 569 people over 55 years old living in central Greece were studied. A protocol was completed for each patient that included smoking habits, job, age, weight, drugs used for respiratory system and symptoms. A chest radiograph was taken for each patient. Each patient was submitted to three trials of dynamic spirometry. The best results were used for the study. The patients who complained of expectoration for three months for two continuous years and had obstructive pattern of lung function were diagnosed with chronic bronchitis. 9.6% of the subjects in rural area and 17.1% in industrial area were diagnosed with chronic bronchitis. Half the COPD patients never visited a doctor for this specific problem and were diagnosed for the first time. One out of three examined people had abnormal lung function. Consequently, more than one out of ten people older than 55 years have chronic bronchitis. Industrial exposure seems to double the risk for chronic bronchitis. PMID- 10865244 TI - Exposure of female farmers to dust on family farms. AB - Studies of dust on farms conducted to date, both in Poland and abroad, concern only health risk due to dust while performing selected occupations. The present study is a subsequent attempt to determine the year-round exposure to dust at workplaces in agriculture, and covers the workplace of a private farmer which is typical of Polish agriculture: a female farmer who is running a family farm together with her husband. Studies conducted on 10 mixed production family farms showed that women actively participate in farm activities. The occupations performed by women focus around the household - mainly the care of animals; whereas their field work consists primarily of manual jobs associated with crop cultivation, as well as auxiliary activities during harvest. Although the level of exposure to dust determined for female farmers remains statistically within the range of allowable conditions, this factor should be considered as an important health risk due to its high concentrations and pathogenic components. PMID- 10865243 TI - Subacute toxicity of orally applied alpha-cypermethrin in Swiss mice. AB - The effect of a synthetic pyrethroid - alpha-cypermethrin administered per os for 28 days to Swiss mice was examined on phagocytic and bactericidal activity of neutrophils, and leukocytic image, IL-12 p70 level in blood plasma, as well as histologic and ultrastructural picture of the liver, heart, kidneys, lung and spleen. A synthetic pyrethroid alpha-cypermethrin, [(R,S)-alpha-cyano-3 phenoxybenzyl (R,S)-cis,trans-3-(2,2-dichlorovinyl)-2,2 dimethylcyclopropanecarboxylate], produced by the Chemical Plant in Jaworzno was used in the study. The preparation for the application per os was used in doses 1/2 LD(50) (25 mg/kg body mass) and 1/5 LD(50) (10 mg/kg body mass). The results were presented as mean (x) +/- standard error (SEM) and subjected to statistical analysis by the parametric t-Student test. Subacute poisoning of mice with alpha cypermethrin in doses 1/2 LD(50) and 1/5 LD(50) resulted in decreased bactericidal activity of neutrophils. The dose 10 mg/kg body mass had a stronger stimulatory effect on phagocytic activity than 25 mg/kg body mass. Significantly higher numbers of monocytes and lymphocytes were observed in the blood of male mice poisoned with 1/5 LD(50) alpha-cypermethrin. The administration of alpha cypermethrin resulted for both doses in the decrease in IL-12 p70 serum secretion. The lowest IL-12 p70 level (pg/ml) was noted among female mice administered 1/2 LD(50) of the preparation. The results of the study may indicate that the pyrethroid in the study had a suppressive effect on Il-12 p70 production. In mice administered 1/5 LD(50) or 1/2 LD(50) of the preparation examined, histopathologic and ultrastructural changes were observed in the liver and kidneys. PMID- 10865245 TI - Heavy metal poisoning in glass worker characterised by severe. AB - The paper presents the clinical description of the masticatory organ and biochemical assessment of dental tissue in a patient employed in a glassworks for 20 years. During 12 years the patient has suffered baldness ("Alopecia areata") and atypical extensive and non-healing cutaneous lesions. Dental examination revealed changes typical of chronic poisoning by cadmium and bismuth compounds. PMID- 10865246 TI - Occupational contact dermatitis to Phaseolus vulgaris in a farmer - a case report. AB - A case of occupational contact dermatitis in a farmer is described, caused among others by Phaseolus vulgaris. The patient's history of eczematous and vesicular and bullous skin reactions occurring after exposure to Phaseolus was confirmed by skin tests with native leaves of the plant. To the best of our knowledge, this is the first description of occupational contact dermatitis caused by leaves of Phaseolus plant. PMID- 10865247 TI - Increased urinary excretion of thioethers as a marker for detecting exposure to herbicide containing 2,4-dichlorophenoxyacetic acid dimethylamine - experimental study on mice. AB - The possibility that urinary thioethers concentration might be a marker for detecting exposure to herbicide containing 2,4-dichlorophenoxyacetic acid dimethylamine (2,4-DMA) was investigated in animals. Mice were treated with the herbicide containing 2,4-DMA consecutively for 4 days. Urinary concentrations of thioethers related either to body weight or creatinine concentration in urine in the group of animals treated with herbicide were significantly higher compared to control group. Results suggest that thioethers determination in urine might be a noninvasive and simple method for detecting exposure to herbicide containing 2,4 DMA. PMID- 10865248 TI - Anticoagulation for chronic atrial fibrillation. PMID- 10865249 TI - JAMA 100 years ago: CONTRIBUTIONS TO THE SCIENCE OF MEDICINE BY THE PUPILS OF WILLIAM H. WELCH PMID- 10865250 TI - HAART stopping news: experts examine structured therapy interruption for HIV. PMID- 10865252 TI - The bridge at Ann Arbor: Japanese health program. PMID- 10865251 TI - Secretary Shalala sums up her term of office at DHHS. PMID- 10865254 TI - The world in medicine: diagnosing prion disease PMID- 10865253 TI - Periodontal disease may pose one risk for premature birth. PMID- 10865255 TI - The world in medicine: asthma care in canada PMID- 10865257 TI - The world in medicine: where did H pylori come from? PMID- 10865256 TI - The world in medicine: stroke damage from iron PMID- 10865258 TI - Medical treatment of opiate addiction. PMID- 10865259 TI - Medical treatment of opiate addiction PMID- 10865260 TI - Genitourinary consequences of radical prostatectomy. PMID- 10865261 TI - Genitourinary consequences of radical prostatectomy. PMID- 10865262 TI - Legislation and end-of-life care. PMID- 10865263 TI - Legislation and end-of-life care. PMID- 10865264 TI - Legislation and end-of-life care. PMID- 10865265 TI - Legislation and end-of-life care PMID- 10865266 TI - Statins and risk of coronary heart disease. PMID- 10865268 TI - Statins and risk of coronary heart disease PMID- 10865267 TI - Statins and risk of coronary heart disease. PMID- 10865270 TI - Effect of ritonavir on saquinavir metabolism PMID- 10865269 TI - Effect of ritonavir on saquinavir metabolism. PMID- 10865271 TI - Relationship of symptom-onset-to-balloon time and door-to-balloon time with mortality in patients undergoing angioplasty for acute myocardial infarction. AB - CONTEXT: Rapid time to treatment with thrombolytic therapy is associated with lower mortality in patients with acute myocardial infarction (MI). However, data on time to primary angioplasty and its relationship to mortality are inconclusive. OBJECTIVE: To test the hypothesis that more rapid time to reperfusion results in lower mortality in the strategy of primary angioplasty. DESIGN: Prospective observational study of data collected from the Second National Registry of Myocardial Infarction between June 1994 and March 1998. SETTING: A total of 661 community and tertiary care hospitals in the United States. SUBJECTS: A cohort of 27,080 consecutive patients with acute MI associated with ST-segment elevation or left bundle-branch block who were treated with primary angioplasty. MAIN OUTCOME MEASURE: In-hospital mortality, compared by time from acute MI symptom onset to first balloon inflation and by time from hospital arrival to first balloon inflation (door-to-balloon time). RESULTS: Using a multivariate logistic regression model, the adjusted odds of in-hospital mortality did not increase significantly with increasing delay from MI symptom onset to first balloon inflation. However, for door-to-balloon time (median time 1 hour 56 minutes), the adjusted odds of mortality were significantly increased by 41% to 62% for patients with door-to-balloon times longer than 2 hours (for 121-150 minutes: odds ratio [OR], 1.41; 95% confidence interval [CI], 1.08-1.84; P=.01; for 151-180 minutes: OR, 1.62; 95% CI, 1.23-2.14; P<.001; and for >180 minutes: OR, 1.61; 95% CI, 1.25-2.08; P<.001). CONCLUSIONS: The relationship in our study between increased mortality and delay in door-to-balloon time longer than 2 hours (present in nearly 50% of this cohort) suggests that physicians and health care systems should work to minimize door-to-balloon times and that door to-balloon time should be considered when choosing a reperfusion strategy. Door to-balloon time also appears to be a valid quality-of-care indicator. JAMA. 2000. PMID- 10865272 TI - Implications of an aging registered nurse workforce. AB - CONTEXT: The average age of registered nurses (RNs), the largest group of health care professionals in the United States, increased substantially from 1983 to 1998. No empirically based analysis of the causes and implications of this aging workforce exists. OBJECTIVES: To identify and assess key sources of changes in the age distribution and total supply of RNs and to project the future age distribution and total RN workforce up to the year 2020. DESIGN AND SETTING: Retrospective cohort analysis of employment trends of recent RN cohorts over their lifetimes based on US Bureau of the Census Current Population Surveys between 1973 and 1998. Recent workforce trends were used to forecast long-term age and employment of RNs. PARTICIPANTS: Employed RNs aged 23 to 64 years (N = 60,386). MAIN OUTCOME MEASURES: Annual full-time equivalent employment of RNs in total and by single year of age. RESULTS: The average age of working RNs increased by 4.5 years between 1983 and 1998. The number of full-time equivalent RNs observed in recent cohorts has been approximately 35% lower than that observed at similar ages for cohorts that entered the labor market 20 years earlier. Over the next 2 decades, this trend will lead to a further aging of the RN workforce because the largest cohorts of RNs will be between age 50 and 69 years. Within the next 10 years, the average age of RNs is forecast to be 45.4 years, an increase of 3.5 years over the current age, with more than 40% of the RN workforce expected to be older than 50 years. The total number of full-time equivalent RNs per capita is forecast to peak around the year 2007 and decline steadily thereafter as the largest cohorts of RNs retire. By the year 2020, the RN workforce is forecast to be roughly the same size as it is today, declining nearly 20% below projected RN workforce requirements. CONCLUSIONS: The primary factor that has led to the aging of the RN workforce appears to be the decline in younger women choosing nursing as a career during the last 2 decades. Unless this trend is reversed, the RN workforce will continue to age, and eventually shrink, and will not meet projected long-term workforce requirements. JAMA. 2000. PMID- 10865274 TI - Physical activity and risk of stroke in women. AB - CONTEXT: Persuasive evidence has demonstrated that increased physical activity is associated with substantial reduction in risk of coronary heart disease. However, the role of physical activity in the prevention of stroke is less well established. OBJECTIVE: To examine the association between physical activity and risk of total stroke and stroke subtypes in women. DESIGN AND SETTING: The Nurses' Health Study, a prospective cohort study of subjects residing in 11 US states. SUBJECTS: A total of 72,488 female nurses aged 40 to 65 years who did not have diagnosed cardiovascular disease or cancer at baseline in 1986 and who completed detailed physical activity questionnaires in 1986, 1988, and 1992. MAIN OUTCOME MEASURE: Incident stroke occurring between baseline and June 1, 1994, compared among quintiles of physical activity level as measured by metabolic equivalent tasks (METs) in hours per week. RESULTS: During 8 years (560,087 person-years) of follow-up, we documented 407 incident cases of stroke (258 ischemic strokes, 67 subarachnoid hemorrhages, 42 intracerebral hemorrhages, and 40 strokes of unknown type). In multivariate analyses controlling for age, body mass index, history of hypertension, and other covariates, increasing physical activity was strongly inversely associated with risk of total stroke. Relative risks (RRs) in the lowest to highest MET quintiles were 1. 00, 0.98, 0.82, 0.74, and 0.66 (P for trend=.005). The inverse gradient was seen primarily for ischemic stroke (RRs across increasing MET quintiles, 1.00, 0.87, 0.83, 0.76, and 0.52; P for trend=.003). Physical activity was not significantly associated with subarachnoid hemorrhage or intracerebral hemorrhage. After multivariate adjustment, walking was associated with reduced risk of total stroke (RRs across increasing walking MET quintiles, 1.00, 0. 76, 0.78, 0.70, and 0.66; P for trend=.01) and ischemic stroke (RRs across increasing walking MET quintiles, 1.00, 0.77, 0.75, 0.69, and 0.60; P for trend=.02). Brisk or striding walking pace was associated with lower risk of total and ischemic stroke compared with average or casual pace. CONCLUSION: These data indicate that physical activity, including moderate-intensity exercise such as walking, is associated with substantial reduction in risk of total and ischemic stroke in a dose-response manner. JAMA. 2000. PMID- 10865273 TI - Broad-spectrum sunscreen use and the development of new nevi in white children: A randomized controlled trial. AB - CONTEXT: High nevus density is a risk factor for cutaneous malignant melanoma. Melanocytic nevi originate in childhood and are largely caused by solar exposure. OBJECTIVE: To determine whether use of broad-spectrum, high-sun protection factor (SPF) sunscreen attenuates development of nevi in white children. DESIGN: Randomized trial conducted June 1993 to May 1996. SETTING AND PARTICIPANTS: A total of 458 Vancouver, British Columbia, schoolchildren in grades 1 and 4 were randomized in 1993. After exclusion of nonwhite children and those lost to follow up or with missing data, 309 children remained for analysis. Each child's nevi were enumerated at the start and end of the study in 1996. INTERVENTION: Parents of children randomly assigned to the treatment group (n=222) received a supply of SPF 30 broad-spectrum sunscreen with directions to apply it to exposed sites when the child was expected to be in the sun for 30 minutes or more. Children randomly assigned to the control group (n=236) received no sunscreen and were given no advice about sunscreen use. MAIN OUTCOME MEASURE: Number of new nevi acquired during the 3 years of the study, compared between treatment and control groups. RESULTS: Children in the sunscreen group developed fewer nevi than did children in the control group (median counts, 24 vs 28; P=.048). A significant interaction was detected between freckling and study group, indicating that sunscreen use was much more important for children with freckles than for children without. Modeling of the data suggests that freckled children assigned to a broad-spectrum sunscreen intervention would develop 30% to 40% fewer new nevi than freckled children assigned to the control group. CONCLUSIONS: Our data indicate that broad spectrum sunscreens may attenuate the number of nevi in white children, especially if they have freckles. JAMA. 2000. PMID- 10865275 TI - Spread of Mycobacterium tuberculosis in a community implementing recommended elements of tuberculosis control. AB - CONTEXT: Despite improvements in tuberculosis (TB) control during the past decade, Mycobacterium tuberculosis transmission and resulting disease continue to occur in the United States. OBJECTIVE: To determine the primary reasons for disease development from a particular strain of M tuberculosis. DESIGN: Population-based, molecular epidemiological study. SETTING: Urban community in the San Francisco Bay area of California with recommended elements of TB control in place. PATIENTS: Seventy-three TB cases were reported in 1996-1997 that resulted from 1 strain of M tuberculosis as identified by TB genotyping and epidemiological linkage. MAIN OUTCOME MEASURES: Transmission patterns involving source and secondary case-patients; primary reasons for disease development. RESULTS: Seventy-three (33%) of 221 TB case-patients in this community resulted from this strain of M tuberculosis. Thirty-nine (53%) of the 73 case-patients developed TB because they were not identified as contacts of source case patients; 20 case-patients (27%) developed TB because of delayed diagnosis of their sources; and 13 case-patients (18%) developed TB because of problems associated with the evaluation or treatment of contacts; and 1 case-patient (1%) developed TB because of delay in being elicited as a contact. Of the 51 TB cases identified with sources, 49 (96%) were infected within the 2 years prior to diagnosis. CONCLUSIONS: Our results indicate that in a community that has implemented the essential elements of TB control, TB from ongoing transmission of M tuberculosis will continue to develop unless patients are diagnosed earlier and contacts are more completely identified. JAMA. 2000. PMID- 10865276 TI - Are increasing 5-year survival rates evidence of success against cancer? AB - CONTEXT: Increased 5-year survival for cancer patients is generally inferred to mean that cancer treatment has improved and that fewer patients die of cancer. Increased 5-year survival, however, may also reflect changes in diagnosis: finding more people with early-stage cancer, including some who would never have become symptomatic from their cancer. OBJECTIVE: To determine the relationship over time between 5-year cancer survival and 2 other measures of cancer burden, mortality and incidence. DESIGN AND SETTING: Using population-based statistics reported by the National Cancer Institute Surveillance, Epidemiology, and End Results Program, we calculated the change in 5-year survival from 1950 to 1995 for the 20 most common solid tumor types. Using the tumor as the unit of analysis, we correlated changes in 5-year survival with changes in mortality and incidence. MAIN OUTCOME MEASURE: The association between changes in 5-year survival and changes in mortality and incidence measured using simple correlation coefficients (Pearson and Spearman). RESULTS: From 1950 to 1995, there was an increase in 5-year survival for each of the 20 tumor types. The absolute increase in 5-year survival ranged from 3% (pancreatic cancer) to 50% (prostate cancer). During the same period, mortality rates declined for 12 types of cancer and increased for the remaining 8 types. There was little correlation between the change in 5-year survival for a specific tumor and the change in tumor-related mortality (Pearson r=.00; Spearman r=-.07). On the other hand, the change in 5 year survival was positively correlated with the change in the tumor incidence rate (Pearson r=+. 49; Spearman r=+.37). CONCLUSION: Although 5-year survival is a valid measure for comparing cancer therapies in a randomized trial, our analysis shows that changes in 5-year survival over time bear little relationship to changes in cancer mortality. Instead, they appear primarily related to changing patterns of diagnosis. JAMA. 2000. PMID- 10865277 TI - Public health law in a new century: part II: public health powers and limits. AB - The Constitution allocates public health powers among the federal government and the states. Federal public health powers include the authority to tax, spend, and regulate interstate commerce. These powers enable the federal government to raise revenues, allocate resources, economically penalize risk behavior, and broadly regulate in the public's interest. States have an inherent authority to protect, preserve, and promote the health, safety, morals, and general welfare of the people, termed police powers. Police powers enable states to preserve the public health in areas ranging from injury and disease prevention to sanitation, waste disposal, and environmental protection. The Rehnquist Court has emphasized the limits of federal powers and the primacy of states in public health issues affecting local concerns. Finally, the Constitution safeguards individual interests in autonomy, privacy, liberty, and property. The Supreme Court often defers to public health authorities in matters of public health, but engages in strict scrutiny if government interferes with fundamental freedoms or discriminates against a suspect class. Provided that they act justly and reasonably to avert a serious health threat, the Court should cede to agencies the power to act for the communal good. JAMA. 2000. PMID- 10865278 TI - The decreasing supply of registered nurses: inevitable future or call to action? PMID- 10865279 TI - Primary angioplasty--time is of the essence. PMID- 10865281 TI - JAMA 100 years ago: REPORT ON MASSAGE PMID- 10865280 TI - A piece of my mind: blood lines. PMID- 10865282 TI - Exposure to home pesticides linked to Parkinson disease. PMID- 10865283 TI - Medically underserved children need more than insurance card. PMID- 10865284 TI - Poor children subject to "environmental injustice". PMID- 10865286 TI - Health agencies update: oral disease a silent epidemic PMID- 10865288 TI - Health agencies update: anal cancer screening PMID- 10865287 TI - Health agencies update: nailing west nile virus PMID- 10865289 TI - Health agencies update: monkey stressed, monkey drinks PMID- 10865290 TI - Long-term outcomes of Lyme disease. PMID- 10865292 TI - Long-term outcomes of lyme disease PMID- 10865291 TI - Long-term outcomes of Lyme disease. PMID- 10865293 TI - Antiviral agents for influenza. PMID- 10865294 TI - Antiviral agents for influenza PMID- 10865295 TI - Benefit of prophylactic mastectomy for women with BRCA1 or BRCA2 mutations. PMID- 10865297 TI - Benefit of prophylactic mastectomy for women with BRCA1 or BRCA2 mutations PMID- 10865296 TI - Benefit of prophylactic mastectomy for women with BRCA1 or BRCA2 mutations. PMID- 10865298 TI - Retained needle fragments in patients with diabetic neuropathy. PMID- 10865299 TI - Cholera incidence and El Nino-related higher ambient temperature. PMID- 10865300 TI - Periconceptional intake of folic acid among low-income women. PMID- 10865301 TI - Cognitive rehabilitation for traumatic brain injury: A randomized trial. Defense and Veterans Head Injury Program (DVHIP) Study Group. AB - CONTEXT: Traumatic brain injury (TBI) is a principal cause of death and disability in young adults. Rehabilitation for TBI has not received the same level of scientific scrutiny for efficacy and cost-efficiency that is expected in other medical fields. OBJECTIVE: To evaluate the efficacy of inpatient cognitive rehabilitation for patients with TBI. DESIGN AND SETTING: Single-center, parallel group, randomized trial conducted from January 1992 through February 1997 at a US military medical referral center. PATIENTS: One hundred twenty active-duty military personnel who had sustained a moderate-to-severe closed head injury, manifested by a Glasgow Coma Scale score of 13 or less, or posttraumatic amnesia lasting at least 24 hours, or focal cerebral contusion or hemorrhage on computed tomography or magnetic resonance imaging. INTERVENTIONS: Patients were randomly assigned to an intensive, standardized, 8-week, in-hospital cognitive rehabilitation program (n=67) or a limited home rehabilitation program with weekly telephone support from a psychiatric nurse (n=53). MAIN OUTCOME MEASURES: Return to gainful employment and fitness for military duty at 1-year follow-up, compared by intervention group. RESULTS: At 1-year follow-up, there was no significant difference between patients who had received the intensive in hospital cognitive rehabilitation program vs the limited home rehabilitation program in return to employment (90% vs 94%, respectively; P=.51; difference, 4% [95% confidence interval ?CI?, -5% to 14%]) or fitness for duty (73% vs 66%, respectively; P=. 43; difference, 7% [95% CI, -10% to 24%]). There also were no significant differences in cognitive, behavioral, or quality-of-life measures. In a post-hoc subset analysis of patients who were unconscious for more than 1 hour (n = 75) following TBI, the in-hospital group had a greater return-to-duty rate (80% vs 58%; P=. 05). CONCLUSIONS: In this study, the overall benefit of in hospital cognitive rehabilitation for patients with moderate-to-severe TBI was similar to that of home rehabilitation. These findings emphasize the importance of conducting randomized trials to evaluate TBI rehabilitation interventions. JAMA. 2000;283:3075-3081 PMID- 10865302 TI - Efficacy of venlafaxine extended-release capsules in nondepressed outpatients with generalized anxiety disorder: A 6-month randomized controlled trial. AB - CONTEXT: Generalized anxiety disorder (GAD) is a chronic disorder that is associated with debilitating psychic and somatic symptoms. Venlafaxine extended release (XR) capsules have been shown to be effective in short-term treatment of patients with GAD without major depressive disorder (MDD), but long-term data are needed to establish whether this agent confers persistent benefits. OBJECTIVE: To compare the 6-month efficacy and safety of a flexible dosage of venlafaxine XR in outpatients with GAD without associated MDD. DESIGN: Six-month, randomized, double-blind, placebo-controlled, parallel-group trial conducted May 1996 to October 1997. SETTING: Fourteen outpatient clinics and private psychiatric practices in the United States. PARTICIPANTS: A total of 251 outpatients aged 18 years or older who met Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV) criteria for GAD, had sufficient symptoms to require treatment, and did not have coexisting MDD. INTERVENTIONS: Participants were randomly assigned to receive either placebo (n=127) or venlafaxine XR (75, 150, or 225 mg/d, as required to control symptoms; n=124) for 28 weeks. MAIN OUTCOME MEASURES: Changes from baseline in the Hamilton Rating Scale for Anxiety (HAM-A) total score, the HAM-A psychic anxiety factor score, and the Clinical Global Impressions (CGI) scale Severity of Illness and Global Improvement scores, compared by intervention group. RESULTS: During weeks 6 through 28, response rates in the venlafaxine XR group were 69% or higher compared with rates of 42% to 46% in the placebo group (P<.001). By an evaluable-patient analysis, venlafaxine XR compared with placebo significantly improved anxiety scores from week 1 or 2 through week 28 on all primary efficacy measures, including the HAM-A total (P<.001), the HAM-A psychic anxiety factor (P<.001), and the CGI scale scores (P<.001). Adjusted mean changes from baseline to week 28 using last observation-carried-forward methods were for HAM-A, venlafaxine XR -13.4, placebo -8.7 (P<.001); for HAM-A psychic anxiety score, venlafaxine XR -7.4, placebo -4.2 (P<.001); and for CGI-Improvement, venlafaxine XR 2.2, placebo 3.0 (P<.001). The most common treatment-emergent adverse event was nausea, followed by somnolence and dry mouth. CONCLUSIONS: This study is the first placebo-controlled demonstration of the long-term efficacy of any drug class in treating outpatients with DSM-IV-diagnosed GAD. Venlafaxine XR is an effective, rapidly acting, safe, once-daily agent for both the short- and long-term treatment of anxiety and may provide an important alternative to currently available anxiolytics. JAMA. 2000. PMID- 10865303 TI - Invasive Haemophilus influenzae disease in Alaskan residents aged 10 years and older before and after infant vaccination programs. AB - CONTEXT: The introduction of Haemophilus influenzae type b (Hib) vaccination of children has led to a decline in incidence of Hib disease in young Alaskan children. However, the impact of vaccination on unimmunized Alaskan adolescents and adults has not been studied. OBJECTIVE: To characterize trends in incidence of and mortality due to invasive H influenzae disease in Alaskan residents aged 10 years and older prior to and after the introduction of a statewide Hib infant vaccination program. DESIGN AND SETTING: Population-based, descriptive correlational study conducted 1980-1996 in Alaska. SUBJECTS: One hundred twenty nine individuals (31 Alaska Natives and 98 nonnative Alaska residents) aged 10 years and older in whom H influenzae was cultured from a normally sterile site. MAIN OUTCOME MEASURES: Incidence of H influenzae infection before (1980-1990) vs after (1991-1996) vaccination program initiation; serotype, biotype, and beta lactamase production of isolates. RESULTS: The overall annual incidence of invasive H influenzae in those aged 10 years and older declined 33%, from 2.1 per 100,000 persons per year to 1.4 per 100,000 persons per year (P=. 03) after initiation of statewide infant Hib vaccination programs in 1991. This reduction appeared to be the result of a decrease in serotype b disease (82%; P<.001). Infection with other H influenzae serotypes and nontypeable strains increased from 0.5 per 100,000 persons per year to 1.1 per 100,000 persons per year (P=.01). Incidence declined from 4.2 per 100,000 persons per year to 1.2 per 100,000 persons per year in Alaska Natives (P=.005) and from 1.7 per 100,000 persons per year to 1.4 per 100,000 persons per year in nonnative Alaska residents (P=.37). Pneumonia (43%), sepsis (26%), and meningitis (16%) were the most common clinical presentations. Alcohol/drug abuse was comorbid in 15% of patients, while 13% of patients were pregnant women. beta-Lactamase production occurred in 35% of isolates and was stable throughout the surveillance. The overall case-fatality rate was 15%. CONCLUSION: The overall statewide incidence of invasive H influenzae infections in unimmunized persons aged 10 years and older decreased after the initiation of an infant Hib vaccine program, perhaps by decreasing Hib carriage in child reservoirs. An increase in non-serotype b strains was observed. This trend justifies the need for continued surveillance of invasive disease caused by H influenzae. JAMA. 2000. PMID- 10865304 TI - Effects of exercise training on left ventricular function and peripheral resistance in patients with chronic heart failure: A randomized trial. AB - CONTEXT: Exercise training in patients with chronic heart failure improves work capacity by enhancing endothelial function and skeletal muscle aerobic metabolism, but effects on central hemodynamic function are not well established. OBJECTIVE: To evaluate the effects of exercise training on left ventricular (LV) function and hemodynamic response to exercise in patients with stable chronic heart failure. DESIGN: Prospective randomized trial conducted in 1994-1999. SETTING: University department of cardiology/outpatient clinic in Germany. PATIENTS: Consecutive sample of 73 men aged 70 years or younger with chronic heart failure (with LV ejection fraction of approximately 0.27). INTERVENTION: Patients were randomly assigned to 2 weeks of in-hospital ergometer exercise for 10 minutes 4 to 6 times per day, followed by 6 months of home-based ergometer exercise training for 20 minutes per day at 70% of peak oxygen uptake (n=36) or to no intervention (control group; n=37). MAIN OUTCOME MEASURES: Ergospirometry with measurement of central hemodynamics by thermodilution at rest and during exercise; echocardiographic determination of LV diameters and volumes, at baseline and 6-month follow-up, for the exercise training vs control groups. RESULTS: After 6 months, patients in the exercise training group had statistically significant improvements compared with controls in New York Heart Association functional class, maximal ventilation, exercise time, and exercise capacity as well as decreased resting heart rate and increased stroke volume at rest. In the exercise training group, an increase from baseline to 6-month follow up was observed in mean (SD) resting LV ejection fraction (0.30 [0.08] vs 0.35 [0.09]; P=.003). Mean (SD) total peripheral resistance (TPR) during peak exercise was reduced by 157 (306) dyne/s/cm(-5) in the exercise training group vs an increase of 43 (148) dyne/s/cm(-5) in the control group (P=.003), with a concomitant increase in mean (SD) stroke volume of 14 (22) mL vs 1 (19) mL in the control group (P=.03). There was a small but significant reduction in mean (SD) LV end diastolic diameter of 4 (6) mm vs an increase of 1 (4) mm in the control group (P<.001). Changes from baseline in resting TPR for both groups were correlated with changes in stroke volume (r=-0.76; P<.001) and in LV end diastolic diameter (r=0.45; P<.001). CONCLUSIONS: In patients with stable chronic heart failure, exercise training is associated with reduction of peripheral resistance and results in small but significant improvements in stroke volume and reduction in cardiomegaly. JAMA. 2000. PMID- 10865305 TI - Recommendations for the establishment of primary stroke centers. Brain Attack Coalition. AB - OBJECTIVE: To develop recommendations for the establishment and operation of primary stroke centers as an approach to improve the medical care of patients with stroke. PARTICIPANTS: Members of the Brain Attack Coalition (BAC), a multidisciplinary group of representatives from major professional organizations involved with delivering stroke care. Supplemental input was obtained from other experts involved in acute stroke care. EVIDENCE: A review of literature published from 1966 to March 2000 was performed using MEDLINE. More than 600 English language articles that had evidence from randomized clinical trials, meta analyses, care guidelines, or other appropriate methods supporting specific care recommendations for patients with acute stroke that could be incorporated into a stroke center model were selected. CONSENSUS PROCESS: Articles were reviewed initially by 1 author (M.J.A.). Members of the BAC reviewed each recommendation in the context of current practice parameters, with special attention to improving the delivery of care to patients with acute stroke, cost-effectiveness, and logistical issues related to the establishment of primary stroke centers. Consensus was reached among all BAC participants before an element was added to the list of recommendations. CONCLUSIONS: Randomized clinical trials and observational studies suggest that several elements of a stroke center would improve patient care and outcomes. Key elements of primary stroke centers include acute stroke teams, stroke units, written care protocols, and an integrated emergency response system. Important support services include availability and interpretation of computed tomography scans 24 hours everyday and rapid laboratory testing. Administrative support, strong leadership, and continuing education are also important elements for stroke centers. Adoption of these recommendations may increase the use of appropriate diagnostic and therapeutic modalities and reduce peristroke complications. The establishment of primary stroke centers has the potential to improve the care of patients with stroke. JAMA. 2000. PMID- 10865306 TI - The rational clinical examination. Does this patient have carpal tunnel syndrome? AB - CONTEXT: History taking and physical examination maneuvers, including Tinel and Phalen signs, are widely used for the diagnosis of carpal tunnel syndrome (CTS). OBJECTIVE: To systematically review the precision and accuracy of history taking and physical examination in diagnosing CTS in adults. DATA SOURCES: English language literature was searched using MEDLINE (January 1966-February 2000) as well as bibliographies of relevant articles. STUDY SELECTION: Studies of patients presenting to clinicians with symptoms suggestive of CTS in which findings from clearly described physical examination maneuvers were independently compared with electrodiagnostic testing. Twelve of 42 initially identified articles met these criteria and were included in the review. DATA EXTRACTION: Two authors independently reviewed and abstracted data from all of the articles and reached consensus about any discrepancies. DATA SYNTHESIS: In patients presenting with hand dysesthesias, the findings that best distinguish between patients with electrodiagnostic evidence of CTS and patients without it are hypalgesia in the median nerve territory (likelihood ratio [LR], 3. 1; 95% confidence interval [CI], 2.0-5.1), classic or probable Katz hand diagram results (LR, 2.4; 95% CI, 1.6-3.5), and weak thumb abduction strength (LR, 1.8; 95% CI, 1.4-2.3). Findings that argue against the diagnosis of carpal tunnel syndrome are unlikely Katz hand diagram results (LR, 0.2; 95% CI, 0.0-0.7) and normal thumb abduction strength (LR, 0.5; 95% CI, 0.4-0.7). Several traditional findings of CTS have little or no diagnostic value, including nocturnal paresthesias; Phalen and Tinel signs; thenar atrophy; and 2-point, vibratory, and monofilament sensory testing. Other less commonly used maneuvers, including the square wrist sign, flick sign, and closed fist sign, require validation by other studies before they can be recommended. CONCLUSIONS: Hand symptom diagrams, hypalgesia, and thumb abduction strength testing are helpful in establishing the [corrected] electrodiagnosis of CTS. The utility of these results is limited, however, by problems inherent in using nerve conduction studies as a criterion standard. JAMA. 2000. PMID- 10865307 TI - Public health law in a new century: part III: public health regulation: A systematic evaluation. AB - Public health interventions need justification because they intrude on individual rights and incur economic costs. Coercive interventions can be justified in only 3 cases: to avert a risk of serious harm to other persons, to protect the welfare of incompetent persons, and, most controversially, to prevent a risk to the person himself/herself. This article proposes a systematic evaluation of public health regulation. The article recommends that public health authorities should bear the burden of justification and, therefore, should demonstrate (1) a significant risk based on scientific evidence; (2) the intervention's effectiveness by showing a reasonable fit between means and ends; (3) that economic costs are reasonable; (4) that human rights burdens are reasonable; and (5) that benefits, costs, and burdens are fairly distributed. The 3 articles in this series have sought to provide a fuller understanding of the varied ways in which law can advance the public's health. Public health law should be seen broadly as the government's power and responsibility to ensure the conditions for the population's health. As such, public health law has transcending importance in how we think about government, politics, and policy. JAMA. 2000. PMID- 10865308 TI - Cognitive rehabilitation following traumatic brain injury. PMID- 10865309 TI - The imperative to develop dedicated stroke centers. PMID- 10865310 TI - Prospective, randomized clinical evaluation of Optisol vs organ culture corneal storage media. AB - OBJECTIVE: To compare the outcome of penetrating keratoplasty with the use of corneas stored either in Optisol (Chiron Ophthalmics, Irvine, Calif) or in organ culture. METHODS: Penetrating keratoplasty was performed on 12 pairs of patients matched by age and diagnosis. Each pair of procedures was done on the same day by the same surgeon using the same technique. Twelve pairs of corneas were used. One cornea of each pair had been stored in organ culture at 36 degrees C and one in Optisol at 4 degrees C. Mean (+/-SD) storage time was 6+/-3 days. Mean endothelial cell density before storage was 2617/mm(2) for the corneas in organ culture and 2624/mm(2) for the corneas in Optisol. Examinations were performed at 1, 4, 12, and 24 months. RESULTS: One reversible rejection occurred in the Optisol group. At 1 month the mean endothelial cell density was 2327+/-341/mm(2) for the organ culture group and 2240+/-504/mm(2) for the Optisol group. At 12 months the difference was more pronounced (2225+/-410 and 2103+/-466/mm(2), respectively), although statistically not significant. Corneal thickness also did not show any statistically significant difference. CONCLUSION: Penetrating keratoplasty performed with corneas stored for a maximum of 11 days in either Optisol or organ culture show similar outcomes in the first 2 postoperative years. Arch Ophthalmol. 2000;118:757-760 PMID- 10865311 TI - Effect of phacoemulsification surgery on hypotony following trabeculectomy surgery. AB - OBJECTIVE: To review the effect of phacoemulsification surgery in eyes with chronic hypotony following trabeculectomy with mitomycin C. DESIGN: Retrospective analysis of all eyes that underwent phacoemulsification surgery for symptomatic cataracts and had a preoperative diagnosis of chronic hypotony (intraocular pressure [IOP] .99) and metastasis (7 patients at 50 CGE vs 8 patients at 70 CGE;P=.79). Five-year rates of radiation maculopathy also were similar (for both groups, approximately 75% for tumors within 1 disc diameter and 40% for tumors >1 disc diameter from the macula). Rates of radiation papillopathy were nonsignificantly decreased in the 50-CGE treatment group when tumors were located 1 disc diameter or less from the optic disc (P=.20). Patients treated with the lower dose also experienced significantly less visual field loss. CONCLUSIONS: This level of dose reduction did not result in a lesser degree of visual acuity loss. The lower-dose group did experience significantly less visual field loss. Local tumor recurrence and metastatic death rates were similar in both dose groups. Arch Ophthalmol. 2000;118:773-778 PMID- 10865314 TI - Nonsurgical management of macular hemorrhage secondary to retinal artery macroaneurysms. AB - OBJECTIVE: To report visual acuity outcomes of nonsurgical management of macular hemorrhage secondary to retinal artery macroaneurysms. METHODS: Forty-one patients at multiple centers with macular hemorrhage secondary to retinal artery macroaneurysms managed with observation alone were reviewed. Time to clearance of macular hemorrhage, visual acuity at final follow-up, and presence or absence of macular pigmentary changes after absorption of the hemorrhage were recorded for each patient. RESULTS: On initial examination, visual acuity was 20/200 or worse in all except 4 patients (3 with 20/70, 1 with 20/80). At an average follow-up of 15. 7 months, a final visual acuity of 20/40 or better was achieved in 15 eyes (37%), between 20/50 and 20/100 in 12 (29%), and 20/200 or worse in 14 (34%). Macular pigmentary abnormalities were noted after clearance of the hemorrhage in 23 (56%) of 41 cases, and these eyes generally had worse visual acuity outcomes. CONCLUSIONS: In eyes with macular hemorrhage secondary to retinal artery macroaneurysms managed with observation alone, good visual acuity outcomes can often be achieved. Poorer visual acuity outcomes are associated with macular pigmentary changes after resorption of blood. Arch Ophthalmol. 2000;118:780-785 PMID- 10865315 TI - Disinfection of eyelid specula with chlorhexidine gluconate (Hibiclens) after examinations for retinopathy of prematurity. AB - BACKGROUND: The preferred method of cleaning eyelid specula between examinations for retinopathy of prematurity is unknown. A previous study showed that disinfection with 70% isopropyl alcohol swabs fails to eliminate viruses and bacteria from the specula. OBJECTIVE: To determine if alternative sterilization procedures would allow multiple use of a single speculum without risking nosocomial infection. METHODS: In phase 1, 40 autoclave-sterilized eyelid specula were randomized into either "cleaned" or "patient control" groups after being used for routine retinopathy of prematurity examinations performed in the outpatient setting. Specula in the cleaned group were cleaned with chlorhexidine gluconate (Hibiclens). Specula in the patient control group were not cleaned after use. All study specula were placed into enriched culture media from which bacterial and fungal cultures were obtained. In phase 2, 20 autoclave-sterilized eyelid specula were inoculated with a clinically relevant dilution of adenovirus serovar 5 or herpes simplex type 2. Specula were randomized into either a cleaned or a control group, and cell cultures and immunofluorescence assays were used to document and confirm, respectively, viral growth. RESULTS: In phase 1, all 20 cultures from the patient control group grew bacteria compared with 0 (0%) of 20 cultures from the cleaned group and 0 (0%) of 5 from the cleaned control group. No fungi were isolated from any group. In phase 2, all 10 cultures from specula inoculated with adenovirus serovar 5 grew virus. None of the cultures from the 5 cleaned specula inoculated with herpes simplex type 2 grew virus. In contrast, all 5 cultures in the control group were positive for growth of herpes simplex type 2. CONCLUSIONS: Autoclave sterilization is the ideal method of sterilization of eyelid specula between neonate examinations. When an alternative disinfection technique is required, washing the speculum with chlorhexidine gluconate and tap water is preferred over wiping with a 70% isopropyl alcohol swab. Arch Ophthalmol. 2000;118:786-789 PMID- 10865316 TI - Ischemic Optic Neuropathy Decompression Trial: twenty-four-month update. AB - OBJECTIVE: To describe visual acuity outcomes of patients in the Ischemic Optic Neuropathy Decompression Trial (IONDT) after 24 months of follow-up. DESIGN: The IONDT is a single-masked, multicenter, randomized clinical trial. SETTINGS: Patients were evaluated and followed up at 25 clinical centers located throughout the United States. Data were sent to and analyzed at a central coordinating center. PATIENTS: Two hundred fifty-eight patients 50 years or older with nonarteritic anterior ischemic optic neuropathy and visual acuity of 20/64 or worse, but better than no light perception, were randomized to either a careful follow-up group (n=131) or an optic nerve decompression surgery (ONDS) group (n=127). Of these, 174 continued participation for at least 24 months, 89 in the careful follow-up group and 85 in the ONDS group. METHODS: Randomized patients underwent a standard visual acuity examination at 3, 6, 12, 18, and 24 months of follow-up. The primary outcome was a change of 3 lines or more of visual acuity, defined as a difference of 0.3 in logMAR scores, between baseline and 6 months of follow-up. A secondary outcome was mean change in visual acuity (in logMAR units) at 3, 6, 12, 18, and 24 months following baseline. These changes were estimated using available data from all randomized patients for whom we had data. RESULTS: Of the 258 patients randomized, 143 (55.4%) were male, and 169 (65.5%) were 65 years or older. Mean visual acuity was statistically significantly improved from baseline value at all study visits and for both treatment groups, although visual acuity declined gradually in both groups after the 3-month visit. There were no significant differences between careful follow-up and ONDS in mean change in vision from the baseline and any follow-up time point. At 24 months of follow-up, 31.0% of patients in the careful follow-up group and 29.4% of patients in the ONDS group experienced an increase of 3 or more lines of vision compared with baseline acuity; 21.8% of patients in the careful follow-up group and 20.0% of patients in the ONDS group experienced a decrease of 3 or more lines. In patients who could read at least 1 letter on the Lighthouse chart, there was a gradual decline in mean visual acuity noted over time for both treatment groups, although acuity remained significantly better than at baseline. CONCLUSION: Analysis of visual acuity data from patients enrolled in the IONDT at 24 months of follow-up confirms that there is no benefit of ONDS compared with careful follow-up in patients with nonarteritic anterior ischemic optic neuropathy. Arch Ophthalmol. 2000;118:793-798 PMID- 10865317 TI - Retinitis pigmentosa associated with Fuchs' heterochromic uveitis. AB - OBJECTIVE: To investigate whether the combination of Fuchs' heterochromic uveitis (FHU) and retinitis pigmentosa (RP) in the same patient is coincidental or represents a true association. METHODS: We have examined the frequency of FHU in 338 patients with RP and in 1984 patients who were seen in our primary care ophthalmic clinic because of reasons other than RP. RESULTS: Of 338 patients with RP, 4 (1.2%) had the typical findings of FHU. Three of them had Usher syndrome type II, and 1 had RP simplex. By contrast, only 1 patient in the control group had FHU (5%), and the difference in the frequency of FHU between the 2 groups was significant (P=.002, Fisher exact test). CONCLUSIONS: Fuchs' heterochromic uveitis is associated with RP. Since autoimmune phenomena have been previously described in patients with RP, it is conceivable that RP predisposes to the development of FHU. Arch Ophthalmol. 2000;118:800-802 PMID- 10865318 TI - Effects of intravitreal corticosteroids in the treatment of Bacillus cereus endophthalmitis. AB - OBJECTIVE: To investigate whether intravitreal corticosteroid therapy reduces the extent of inflammatory intraocular tissue damage caused by Bacillus cereus endophthalmitis. METHODS: New Zealand white rabbits were inoculated with 1 x 10(6) B cereus organisms and randomized to receive no treatment (control eyes; n=14), intravitreal vancomycin hydrochloride (n=13), or a combination of intravitreal vancomycin and dexamethasone sodium phosphate (n=13) after 24 hours. The eyes were examined and graded for clinical signs of infection and inflammation on days 7 and 14, followed by enucleation for histopathologic analysis. RESULTS: Both treated groups had significantly less clinical sequelae than controls on day 7. By day 14, eyes given combination treatment had significantly less clinically graded corneal (P=.03) and conjunctival (P=.007) inflammation than eyes treated with vancomycin. Histopathologic analysis revealed a significant decrease in inflammatory changes between all treated eyes and controls at day 14. The only statistically significant difference between eyes given combination treatment and eyes given vancomycin alone was in the retina (P=.03). CONCLUSIONS: Intravitreal corticosteroids may enhance the recovery from B cereus endophthalmitis when given in conjunction with intravitreal antibiotics. The beneficial effect of corticosteroids is noted clinically, but not histologically, by day 14 after single-dose treatment in rabbits. CLINICAL RELEVANCE: This study provides evidence that the use of intravitreal corticosteroids with antibiotics for the treatment of B cereus endophthalmitis may lead to an improvement compared with the use of antibiotics alone. Arch Ophthalmol. 2000;118:803-806 PMID- 10865319 TI - Selective transplantation of rods delays cone loss in a retinitis pigmentosa model. AB - BACKGROUND: Rod-cone retinal degenerations (retinitis pigmentosa) are typified by initial rod loss followed by secondary cone death. Rod death, predominantly caused by gene mutations expressed specifically in these cells, induces scotopic vision loss. Cone death, the overriding cause of blindness, has no current explanation. Disease progression and preliminary data suggest that cone survival depends on rods. OBJECTIVE: To establish whether rod transplantation into mutant rodless retinas could halt cone loss. METHODS: We transplanted pure sheets of rods isolated from normal-sighted mice into the subretinal space of recipient retinal degeneration mice lacking rods but possessing approximately 30% residual cones. Control animals were unoperated on or grafted with inner retinal cells from young normal donors, entire retinas from aged retinal degeneration mice, or gelatin. Two weeks after surgery, we quantified by an unbiased method the numbers of host retinal cones after immunolabeling with specific markers. RESULTS: Only mice receiving rod-rich transplants demonstrated statistically significant greater cone numbers, with rescue of 40% of host cones normally destined to die during this period. CONCLUSION: Cone survival depends specifically on rods. CLINICAL RELEVANCE: Such findings indicate that transplantation of rods could limit loss of cones, thus preserving useful vision in human retinitis pigmentosa. Arch Ophthalmol. 2000;118:807-811 PMID- 10865320 TI - A novel variant of granular corneal dystrophy caused by association of 2 mutations in the TGFBI gene-R124L and DeltaT125-DeltaE126. AB - OBJECTIVE: To characterize the molecular defect in the TGFBI gene in a French family affected with an atypical granular corneal dystrophy. PATIENTS: This family comprises 9 affected individuals across 3 generations without consanguineous marriage. METHODS: Light and electron microscopy were used to examine corneal buttons from patients. Exons of the TGFBI gene were amplified by polymerase chain reaction and sequenced directly using an automated method. Restriction digestion analysis and heteroduplex screening were performed to confirm that the mutations identified were not polymorphisms. RESULTS: Round or snow-flakes-like deposits that stained red with Masson trichrome and appeared as dense, rod-shaped structures were observed in the most anterior layers of the central stroma. All patients were heterozygous for the R124L mutation and a novel mutation predicting the deletion of 2 amino acid residues-threonine (T) and glutamic acid (E)-at codons 125 and 126. CONCLUSIONS: This French family is affected with a novel variant of granular dystrophy that is caused by a molecular defect in the TGFBI gene, reported here for the first time. CLINICAL RELEVANCE: These 2 mutations cause a novel variant of granular dystrophy that is intermediate in severity between the classical and superficial variant forms. Arch Ophthalmol. 2000;118:814-818 PMID- 10865321 TI - Causes of blindness and visual impairment in a population of older Americans: The Salisbury Eye Evaluation Study. AB - OBJECTIVE: To determine the causes of blindness and visual impairment in a population-based sample of older Americans. METHODS: A random sample of 3821 residents of Salisbury, Md, between the ages of 65 and 84 years was identified from Medicare records. Sixty-six percent (2520 persons) agreed to undergo an eye examination; 26% of the participants were African American. The clinical examination included acuity testing with an Early Treatment Diabetic Retinopathy Study chart and standardized refraction testing for those with a visual acuity worse than 20/30, slitlamp and dilated retinal examination by an ophthalmologist, tonometry, lens and fundus photography, and a suprathreshold visual field test. Visual impairment was defined as a best-corrected acuity in the better-seeing eye worse than 20/40 and better than 20/200, while blindness was acuity in the better seeing eye of 20/200 or worse. For those with a visual acuity worse than 20/40 in either eye, one or more causes were assigned by an ophthalmologist and a final cause for each eye was confirmed by a panel of 3 subspecialty ophthalmologists (O.D.S., H.A.Q., and S.B.B.) based on all available evidence. RESULTS: Bilateral presenting acuity worse than 20/40 increased from 4% in the 65- to 74-year age group to 16% in the 80- to 84-year age group. One third of those with presenting acuity worse than 20/40 improved to 20/40 or better with refraction. Overall, 4.5% had a best-corrected acuity worse than 20/40. African Americans were more likely to remain visually impaired than were whites despite refraction (odds ratio [95% confidence interval], 1.7 [1.1-2.6]). Whites were most often impaired or blind from age-related macular degeneration (1.2% vs 0.5%; P=.09). African Americans had higher rates of impairment and blindness from cataract or posterior capsular opacification (2.7% vs 1.1%; P=.006), glaucoma (0.9% vs 0.1%; P=.006), and diabetic retinopathy (1.2% vs 0.2%; P=. 004). CONCLUSIONS: More than half of those with visual impairment or blindness had conditions that were either surgically treatable or potentially preventable. African Americans had a disproportionate number of blinding diseases, particularly those amenable to eye care intervention. Targeted interventions for specific populations to increase appropriate eye care use would greatly improve vision and function in older Americans. Arch Ophthalmol. 2000;118:819-825 PMID- 10865322 TI - Primary placement of a motility coupling post in porous polyethylene orbital implants. AB - The placement of a motility coupling post (MCP) to integrate the prosthesis with a porous orbital implant may enhance prosthetic motility following enucleation. Previously, MCP placement has required a second operation usually at least 6 months following enucleation. We developed a technique to place an MCP reliably and safely into a porous orbital implant at the time of enucleation. Eligibility criteria included high motivation to achieve maximal prosthetic motility, adequate conjunctiva to ensure desirable wound closure, and isolation of the 4 rectus muscles. Enucleation was performed in standard fashion with implantation of a conical porous polyethylene orbital implant. Implanted MCPs protruded anteriorly 2 to 4 mm. The Tenon capsule and conjunctiva were closed in separate layers over the protruding MCP. Thirty-two patients underwent primary placement. Follow-up ranged from 1 to 33 months (mean, 15 months). Nine MCPs spontaneously exposed within the first 4 months. One additional post autoexposed at 12 months. Three patients underwent a secondary procedure to expose the MCP. There were no cases of infection, explantation, or gross MCP malposition. Minor complications included pyogenic granuloma (n=2) and conjunctival overgrowth (n=1). All patients were successfully fit with prostheses. Prosthetic motility was acceptable in all patients. Motility coupling post placement at the time of enucleation surgery in selected patients is an effective, efficient surgical option. Arch Ophthalmol. 2000;118:826-832 PMID- 10865323 TI - Maternal intrauterine herpes simplex virus infection leading to persistent fetal vasculature. AB - Herpes simplex virus can cause serious ocular and systemic disease in the neonate. The mode of transmission to the neonate is usually from the maternal birth canal to the fetus intrapartum; but much more rarely, hematogenous transplacental infection can affect the developing fetus months prior to birth. Persistent fetal vasculature occurs when there is persistence of the fetal ocular vasculature, which normally regresses prior to birth. To our knowledge, we report the first case of serologically proven intrauterine herpes simplex virus infection associated with bilateral persistent fetal vasculature in a surviving term infant. Arch Ophthalmol. 2000;118:837-840 PMID- 10865324 TI - Choroidal metastasis as the initial manifestation of a pigmented neuroendocrine tumor. AB - We report the case of a 77-year-old woman in whom choroidal metastasis was the initial manifestation of a primary neoplasm presumed to be a pigmented pulmonary carcinoid tumor. The tumor initially was misdiagnosed cytologically and pathologically as a choroidal melanoma because it contained intrinsic melanin pigment. Positive immunoreactivity for cytokeratin, synaptophysin, chromogranin, and calcitonin and the presence of dense-core neurosecretory vesicles disclosed by electron microscopy established that the metastasis was a neuroendocrine tumor. Findings from systemic evaluation suggested that the primary tumor was located in the lung. The patient subsequently developed an intradural paraspinal metastasis, which also contained melanin pigment. The latter observation confirmed that the melanin in the uveal metastasis was intrinsic and did not represent secondary phagocytosis by tumor cells. Metastases from pigmented tumors of nonmelanocytic derivation are exceedingly rare but present a major diagnostic challenge to ocular pathologists and cytopathologists if the diagnosis is not suspected. Confirmatory immunohistochemical analysis should be obtained when a pigmented choroidal tumor thought to be a melanoma has atypical features. Arch Ophthalmol. 2000;118:841-845 PMID- 10865325 TI - Interferon alfa therapy against metastatic iris tumor of renal cell carcinoma. PMID- 10865326 TI - Clear corneal cataract wound dehiscence during pneumatic retinopexy. PMID- 10865327 TI - Bilateral scleral thermal injury: complication after skin laser resurfacing. PMID- 10865328 TI - Sutureless pars plana anterior vitrectomy through self-sealing sclerotomies in children. PMID- 10865329 TI - Retinal artery occlusion following coil embolization of carotid-ophthalmic aneurysms. PMID- 10865331 TI - Sclerochoroidal calcification with choroidal neovascularization. PMID- 10865330 TI - Mucosal leishmaniasis presenting as sinusitis and optic neuropathy. PMID- 10865332 TI - Photo essay: Takayasu's disease. PMID- 10865333 TI - Lacrimal secretion in the neonate. PMID- 10865334 TI - Lacrimal secretion in the neonate PMID- 10865335 TI - Half empty or half full? PMID- 10865336 TI - Why do root fillings fail? PMID- 10865337 TI - The Lightspeed technique: experiences with a rotary canal preparation system. PMID- 10865338 TI - Crown-down, nickel titanium and endodontics. PMID- 10865339 TI - Mandibular first molar with five canals. PMID- 10865340 TI - Developing and implementing an OSCE in dentistry. AB - The processes of development, implementation and perceived usefulness of an objective structured clinical examination in restorative dentistry (OSCE(D)) are reported. An OSCE is a system of assessment. It consists of a set of standardised 'stations'. At each station, a student is tested on a specific clinical task. Each student moves from one station to the next so that by the end of the OSCE, every student has completed every station. The primary purpose of this OSCE was to provide feedback to 49 4th year students on their performance in the clinical areas of conservative dentistry, periodontology and prosthetics. Individual profiles were provided to students and the overall results discussed by staff. There were no significant differences in overall performance between genders or between students in the morning and afternoon examinations. There was a significant difference between performance in prosthetics and the other areas and there were some significant differences among the skill clusters of clinical knowledge, procedures, clinical reasoning, history-taking, techniques and communication. Students and staff perceived the OSCE(D) as a valuable tool for providing feedback. The development of the OSCE and the findings described in this paper will be of value to clinical staff who are developing OSCEs in all areas of dentistry. PMID- 10865341 TI - Process and outcome of a visitation to a Central-European dental school. AB - The Advisory Committee on the Training of Dental Practitioners of the EU (ACTDP) in order to ensure a comparably demanding standard in dental education, recommended in 1986 a self-assessment system, and visitations to dental training establishments. After some visitations in EU member countries, a visit was organized to the Dental Faculty, Semmelweis University of Medicine, Budapest, at the request of the Hungarian institution. The completion of the self-assessment questionnaire, and the organization of the visit occurred according to the general guidelines. The visit took place in September 1994, by a team consisting of 4 members. The results of the 3-day visit, and recommendations of the visitation team were presented in a written report, approved by the leaders of the Hungarian Faculty. As a consequence, a curriculum reform was elaborated at the visited Dental School, following the guidelines given by the visitation team, approved by the Faculty in 1996. Summarizing the outcome of the visit, this seemed to be very useful for the visited Dental Faculty, and helped to overcome difficulties in establishing a new curriculum, with better harmonization to EU standards. However, benefits may also derive from these visits for EU countries as well, for convergence in oral health education is of mutual importance for both sides. PMID- 10865342 TI - Investigation of the operative response of a group of general dental practitioners to varying extents of proximal caries. AB - The aim of this investigation was to assess the operative response of a representative sample of Irish General Dental Practitioners of various times since qualification to carious lesions of varying extent in both approximal surfaces of a lower left first molar tooth. 82 General Dental practitioners participated. The dentists were asked to treat 2 manikin teeth for minimal caries on both approximal surfaces extending to the enamel dentine junction, 2 teeth for caries extending approximately to half the thickness of the dentine and 2 teeth for caries extending more than 3/4 of the dentine thickness. Weight measurements were made of the tooth substance cut away. Mean volumes of tooth material removed ranged from 21 mm3 for minimal caries to 76 mm3 for the largest amount of caries. A two-factor analysis of variance revealed that the type of cavity preparation the dentist used had a significant effect on the volume of tooth material removed for all three extents of caries. Time since graduation had a significant effect on the volume of tooth material removed for the least extent of caries, but not for the other categories of carious involvement. PMID- 10865343 TI - Self-assessment in a problem-based learning curriculum in dentistry. AB - The creation of sound self-judgement for students is an integral goal in any educational sphere. Student clinicians in dentistry must learn sufficient skills to be able to self-assess their performance very accurately, because after graduation, the nature of dental procedures means that others are seldom in a position to evaluate the quality of their work. Over recent years, the Department of Dentistry at the University of Adelaide has been developing a self-assessment procedure, initially as a pilot study in the subject oral diagnosis, and currently in all years of the course. This paper describes how self-assessment has been demonstrated to work initially in the pilot subject, and currently across the whole course. The paper also describes how the criteria for assessment and the levels of performance expected for each grade are established and how student performance is monitored. Finally, evidence is presented to indicate acceptance by the students of self-assessment as a valuable and integral part of their learning in dentistry. PMID- 10865344 TI - Human sciences in the first semester of the dental undergraduate course at the Karolinska Institute, Stockholm. AB - The first 9 weeks of the dental undergraduate education at the Karolinska Institutet comprises a transition course, designed to introduce students to university studies leading to professional qualifications in patient-related health sciences. 1 week has been set aside for the theme Man and Society, highlighting the importance of the human sciences for the development of behavioural skills necessary for achieving professionalism and a holistic patient concept. Some essential ethical questions are addressed: intercultural communication, empathy, professional demeanour and the development of professional competence, and group dynamics. In this context, more specific subjects are considered, such as the emergence of the multicultural society and its implications for health services, interpersonal skills and patient communication in the health and medical fields. There are several reasons for including this theme, which forms the basis for the ethical and communicative strands throughout the entire curriculum. As 30-40% of freshmen dental students are of non-Swedish origin, it is essential to include cultural awareness seminars. Another reason is that within the EU, cultural and communicative skills are recognised proficiencies for health professionals; it is also acknowledged that effective delivery of health care may be impeded by misunderstandings in communication and conflict in ethical beliefs. Group discussions are scheduled during the week in order to allow the students to discuss their own experiences related to the theme. The students are also given a written assignment in relation to one of the seminars; the report is assessed as a part of the examination. The week is concluded by a plenum discussion summarising the group discussions. To date, 4 course evaluations, with a response rate of 92.5%, show that 97.3% of the students were positive to the theme as a whole or to specific seminars held during the week, especially intercultural communication, ethics and empathy. The students also believe that the theme is of great importance for dental health professionals. PMID- 10865345 TI - Patient satisfaction in a dental school. AB - PURPOSE: The objective of this study was to measure patients' satisfaction with the facility, services and treatment received at a dental school clinic. METHOD: The 31-item questionnaire consisted of demographic items and items in a 5-point Likert-type format addressing issues such as security, satisfaction with the facility, helpfulness of the staff, progress of treatment, fees, universal precautions taken, quality of care and treatment with dignity and compassion. The questionnaire also asked whether the patient would recommend the school to others seeking dental care. In 1997, patients completed the survey in the clinic waiting room prior to their dental appointment for that day. The survey was conducted over 3 months until 500 patients had participated, representing about 16% of the active patient population. %s of responses were computed and cross tabulations were used to compare responses based on demographics. RESULTS: The most frequently reported reason for wanting to be a patient at the dental school was low cost (67%) followed by up-to-date care (19%). 45% of patients indicated that a friend, neighbour or relative referred them, followed by 34% indicating referral by a dentist. More than 90% responded positively to the items concerning issues related to treatment except for satisfaction with progress of treatment and comparison of care received at the school to care elsewhere. For these 2 items, at least 80% responded positively. 99% indicated they would recommend the dental school to others seeking dental care. CONCLUSION: Results indicate that the vast majority of the patients surveyed are satisfied with the facility, services and treatment received. Comments that addressed areas of concern included the length of time to get an appointment and the length of time the appointment took, and it was recommended that the process of treatment be expedited. PMID- 10865346 TI - The validity and reliability of an OSCE in dentistry. AB - The validity and reliability of a newly-developed objective structured clinical examination for 4th year dental students (OSCE(D)) were estimated by a range of quantitative and qualitative methods. The OSCE(D) consisted of 17 stations in conservation, periodontology and prosthetics. A blueprint was used to match the OSCE stations to clinical subject areas and to 7 skill clusters of clinical competence. Repeat measures of skills were undertaken in the OSCE(D) to maximise its validity and reliability. The primary purposes of the OSCE(D) were to assess clinical competence and to provide feedback to students. The results indicate that the OSCE(D) was intrinsically valid and a better predictor of performance in the final examination than either a concurrent 4th-year examination or Advanced level university entry grades. The OSCE(D) scored relatively highly on internal consistency (Cronbach's alpha = 0.68). Intra-domain and inter-domain correlations were high and inter-examiner reliability was relatively high (eta 2 coefficients ranged from 0.00 to 0.10). There were no significant differences between performances in the morning and afternoon sessions of the examination so reproducibility is assumed to be high. Some improvement in individual stations and in inter-examiner reliability are required. A set of recommendations based on the experience of designing and testing a dental OSCE are provided. PMID- 10865348 TI - An assessment of the influence of clinical demonstrations on the confidence of undergraduate dental students, when treating patients requiring removable partial dentures. AB - AIM: This study was designed to assess the influence of clinical demonstrations, on the confidence of undergraduate dental students, when treating patients requiring removable partial dentures. METHOD: The confidence of 45 undergraduate dental students treating their first patient requiring removable partial dentures was assessed using questionnaires. 23 students were given demonstrations prior to carrying out the treatment; the remainder did not receive a demonstration. Outcome was assessed by the time taken to complete the procedures to a clinically acceptable standard. RESULTS: All the students indicated high levels of agreement with a statement expressing their confidence in coping with the clinical procedures undertaken. The more confident students completed their clinical procedures more quickly than other students, irrespective as to whether they had received a demonstration or not. There was a higher proportion of more confident students in the groups who had received a clinical demonstration; it can therefore be concluded that these groups performed better, as measured by the time taken to achieve the required clinical standard. All the students who had received a demonstration believed they had benefited from it, whilst 67% of the students who had not received a demonstration believed they would have benefited from one. Analysis of the students' comments indicated that demonstrations facilitated confidence, communication skills, understanding and recall in the clinical situation. Students who did not perceive a benefit from the demonstrations believed that they had more time for one to one teaching. CONCLUSION: This study showed that those students who had received a clinical demonstration immediately prior to treating their patients believed they were more confident and as a result their performance was improved. Clinical demonstrations are time consuming, but they would appear to be time well spent. PMID- 10865347 TI - Thai dental students' knowledge of the betel quid chewing habit in Thailand. AB - 385 questionnaires submitted by volunteer Thai dental students on the betel quid chewing habit in Thailand were evaluated. Questions related to the composition of the betel quid, general and oral effects as well as sociological aspects. Only 62.6% considered the habit as typical for Thailand. Knowledge about the composition of the betel quid showed that 30% of the students were poorly informed. Only 58.4% thought slaked lime to be part of the quid. Similar results were obtained for some of the questions relating to physiological and oral effects and the percentage of "do not know" answers was about 30%. It was widely accepted that betel quid chewing is more common in the provinces (83.6%) and that it is a habit of older people (92.2%); particularly women (70.7%). 71.2% of the students did not know where to buy a betel quid. The decline of the habit was monitored by the fact that 96.1% of the students' parents did not indulge in chewing the betel quid compared to 38.7% of grandparents who did. 70% of the students were convinced that the habit will totally disappear. The knowledge of the betel quid chewing habit of Thai dental students indicated a number of deficits showing that these do not come in close contact with this habit anymore in their families or societies. Since elderly people still indulge in chewing betel quid, dental education still has to focus on oral and general effects and side-effects such as oral precancer, oral cancer and oral submucous fibrosis. PMID- 10865349 TI - Introducing undergraduate dental students to the wider role of the primary care team. AB - The development of interprofessional learning between different members of the primary care team can lead to a better understanding of the various roles of the primary care team. It can also result in a more collaborative approach to achieving health care goals. A pilot project was carried out involving placing 6 final-year dental students in general medical practices. Feedback from both dental students and medical practitioners suggested a number of educational benefits were achieved. The generally positive results have encouraged the extension of the pilot to include general medical practice placements for all final-year dental students at Liverpool. PMID- 10865350 TI - A comprehensive care clinic in Swedish dental undergraduate education: 3-year report. AB - In 1993, a new undergraduate curriculum was introduced at the Faculty of Dentistry, Karolinska Institute. The expanded comprehensive care clinic described in this 3-year report, is proposed as a potential model for a CC-clinic in dental undergraduate education. Students enter clinical training at the beginning of the 3rd year and continue through to their final semester, in the 5th year. The one student is responsible for the total oral care of all his/her patients. Patients are stratified on admission into 3 levels, depending on the complexity of treatment need. Before being assigned their 1st patients (patient level I), the students must fulfil specified requirements of theoretical knowledge and clinical procedures and obtain a proficiency certificate. After qualifying for higher proficiency certificates, denoting their level of clinical responsibility, the students assume successively greater responsibility for treatment of patients, with less supervision from clinical instructors. Clinical supervision is the responsibility of integrated multidisciplinary teams of faculty members. Each group of students is assigned a clinical tutor, who is responsible for overall treatment planning of their patients. The individual student's clinical progress and achievements are monitored by a specially designed computer program. Criteria for a final pass in the course are: (1) clinical maturity in overall patient management; (2) technical competence in conjunction with theoretical knowledge. 3 years experience of the new model for comprehensive clinical training in a patient-centred, multidisciplinary clinic has shown that; (a) students seem to acquire a greater understanding of comprehensive case management; (b) patients receive more co-ordinated care; (c) there has been no significant change in the number of clinical procedures completed; (d) difficulties arise in co-ordination of staff over the traditional specialist boundaries; (e) assessment of student maturity and professional responsibility is impeded by the lack of objective criteria. PMID- 10865351 TI - Self-perceived competency at graduation: a comparison of dental graduates from the Adelaide PBL curriculum and the Toronto traditional curriculum. AB - The first class to complete the new Adelaide problem-based learning curriculum graduated in 1997. Their self-perceived competence at graduation was assessed using a revised version of a questionnaire recently compiled and used in Toronto, and based on the global competencies for dental practice accepted in 1995 by all Canadian Faculties of Dentistry. 38 of the 45 Adelaide class members (84%) completed the survey, compared with 93/129 (72%) in Toronto, and their responses were largely similar to those of the Toronto students. At least 67% of the Adelaide and Toronto students felt well-prepared in 34 of the 55 competencies. Most felt well-prepared for the basic everyday items such as diagnosis, local anaesthesia and basic restorative, but less so for items that are not encountered as often in dental school, such as business matters, practice management, soft tissue biopsies and dentofacial trauma. Items showing significant differences between the 2 schools are discussed with reference to curricular and other dissimilarities in the schools, and some inferences are drawn about the importance of context to learning and feelings of competence. PMID- 10865352 TI - Dental students' evaluation of 2 community-oriented PBL modules. AB - OBJECTIVES: To evaluate dental students' perception of 2 problem-based learning (PBL) modules in Dental Public Health implemented within the context of a traditional formal curriculum. METHODS: 2 dental community modules were implemented with an 8-month interval between them on the same group of dental undergraduates; the first in Term 2 and the second in Term 4 of a 5-year 15-term dental course. At the end of each module, a semi-structured questionnaire was administered to evaluate the introductory lecture, the fieldwork activity and the organisation of the modules. RESULTS: In both modules, students reported gaining insight into the subject matter, skills in teamwork, making presentations and collecting data. Some students in the 1st module needed more time to fulfil their learning objectives and had difficulty in collecting data. In the 2nd module, students reported that they lacked motivation because of the place of the module within their timetable. Opinions differed about groupwork. The content of and interest generated by fieldwork activity was rated more positively in the 2nd module than the 1st. Less positively rated in the 2nd module was the introductory lecture and module organisation. CONCLUSIONS: Implementing PBL within a traditional curriculum does not offer uniform outcomes for students. Optimum group size and adequate time are necessary if students are to benefit from PBL. A consistent and continuous PBL approach should be adopted rather than a sporadic one. Further research should establish the optimum balance between PBL and traditional approaches that would allow students to maximise the benefits of both and to identify those students best equipped to benefit from a 'mixed economy' of learning. PMID- 10865353 TI - A skills audit for the dental curriculum. AB - OBJECTIVES: To identify the core skills required of dental graduates, and to identify if and when these skills had been acquired. DESIGN: Postal questionnaire containing a stimulus list of clinical, technical and professional skills. SUBJECTS AND SETTING: General Dental Practitioners (GDPs) registered as vocational trainers, Vocational Dental Practitioners (VDPs) starting their vocational training (VT) year and VDPs ending their VT year in the NHS Trent Region of England. METHOD: The list of skills on the questionnaire was formulated following discussion with a group of 4 clinical academics and 5 GDPs working in the University of Sheffield School of Clinical Dentistry. The questionnaire was sent to the total population of GDPs and VDPs, and responses were requested on an anonymous basis. The purpose mirrored that of a previous survey undertaken in the University of Sheffield Medical School to identify core skills (1), initially in this present survey, defined as skills required by the majority of GDPs (> 50%) and/or used by VDPs (> 50%) in their VT year. RESULTS: 45 of the skills were identified by GDPs as those they expected of the new graduate. All these core skills had been acquired by > 50% of VDPs commencing their VT year with 32 of the skills acquired by > 75% of that group. Of the group ending their VT year, 90% and above had acquired the majority of these skills. At the time of graduation > 50% of VDPs had acquired 10 skills not expected by GDPs. At the end of the VT year, 6 of these unexpected skills had been acquired by > 75% of VDPs. CONCLUSIONS: The survey enabled us to audit the skills required at the end of the undergraduate course and provided the basis for monitoring skills development in undergraduate dental students. PMID- 10865354 TI - Organising the introduction of, and evaluating interviewing in, an admissions system. AB - This paper describes the admissions process, especially the organisation of an interviewing system. The method of application to Dental School is discussed and basic selection processes outlined. The introduction of a new interviewing system is explained and criteria for interview selection and marking defined. All interviewers received regular training; the content of training days is described together with the experience of those involved. Changes made to the interview marking system after 1 year are discussed. Criteria marked at interview were monitored against students' success in year 1 and 2 examinations. Results show that students who performed highly at interview for leadership were more likely to succeed in semester 2 of the course and in year 1 examinations overall. PMID- 10865355 TI - Specialisation in dentistry in Europe--a review. AB - In order to find out what branches of Dentistry are recognized across Europe as Dental specialties, a questionnaire was designed and sent to various Associations and groups across Europe. It contained 8 main questions covering the various specialties that were recognised or were to be recognised. Responses were obtained from 27 countries, 15 EU member states and 12 non member states. The results from the questionnaire indicated that there was a broad agreement on many of the questions, but a considerable variety of opinions. There seems to be a desire to harmonize standards across Europe for the benefit of the profession and the public. PMID- 10865356 TI - The origins of education--from Plato to the Internet. PMID- 10865357 TI - Irish pioneers in medical education: Robert Graves (1796-1853) and William Stokes (1804-1877). PMID- 10865358 TI - The significant developments in dental education of this century, including a case study of dental education at Harvard University. PMID- 10865359 TI - Publishing in dental education: a history and systematic analysis. AB - The aim of this paper is to provide an overview of the development of dental education and its present status as evidenced by published articles in the dental literature. The method used has been a stratified sampling review of 3 journals. The analysis of articles in the British Dental Journal (BDJ) covers 4 periods between 1903 and 1994. A separate study of more recent trends sampled the Journal of Dental Education (J Dent Educ) at 5-year intervals between 1977 and 1997 as well as all the articles published in the European Journal of Dental Education (EJDE) in 1997-98. The long-term survey (BDJ) indicated that the subject of dental education was already being seriously addressed in the early years of this century although articles were few. There was a progressive increase in the number and types of article published over the period of the analysis. Many of the issues addressed in the first half of the century were similar to controversial topics in the present day. In the analysis of recent years (J Dent Educ 1977-1997) the number of articles devoted to strictly educational subjects fell to be replaced by articles only indirectly related to dental education. No consistent pattern was discernible in the type of articles published. There was a paucity of review articles throughout the period. Research articles were proportionally higher in the middle years of the period of the study. Both in the J Dent Educ and the EJDE, the highest proportions of articles were those concerned with curriculum, teaching methods, assessment and quality issues. The paper advocates more evidence-based studies, qualitative research and systematic reviews of the dental educational literature. It concludes that there is a need and an opportunity for expansion of both the quantity and quality of published articles in dental education. PMID- 10865360 TI - Oriental and occidental cultural differences--their possible influences on global dental education and learning. AB - Global dental education, through the medium of the World Wide Web, is being developed. The host nations of this global curriculum currently are Occidental, developed, industrialised nations. This review considers the influence that Occidental nations have already exerted on the development of higher education in Oriental countries. The potential impacts on global dental education which may arise from Occidental-oriental cultural differences are discussed. Approaches to learning and a re-characterisation of the Chinese learner is presented. The potential advantages which Chinese learners may have in learning from the global dental curriculum are proposed. PMID- 10865361 TI - Changing disease patterns and their significance in the training of undergraduate and postgraduate students. PMID- 10865362 TI - Cognitive learning theory and its application in the dental curriculum. PMID- 10865363 TI - The Internet in dental education. PMID- 10865364 TI - The future of dental informatics. PMID- 10865365 TI - Quality assurance in dental education. AB - This paper proposes the development and world-wide use of an electronic dental curriculum which will ensure quality assurance in dental education, in the broadest sense, through the continuous updating of its content and linkages to contemporary resources and databases. This curriculum will need to be reinforced onsite by knowledgeable faculty in the context of patient care delivery with the highest of standards. Two rationales are presented to support the world-wide curriculum concept: one is related to the sophisticated knowledge of science, particularly the biological sciences and its transfer to the oral care environment; the second is related to the progressive role which current developments in information technology allow us to take in the delivery of the curriculum. The paper also addresses the important issue of policy development that will be necessary to ensure continued quality assurance through assessment of this curriculum. We propose to use on a worldwide basis, the system piloted by DENTED, the thematic network project "achieving convergence in standards of output of European Dental Education". The paper concludes by suggesting the next steps for assessing the efficacy of a world-wide electronic curriculum in dental education, supporting development and determining who might maintain this curriculum. PMID- 10865366 TI - Surviving the fiscal challenges facing dental schools. PMID- 10865367 TI - Purchasing of education and delivery of healthcare. PMID- 10865368 TI - Faculty development, remuneration and tenure. PMID- 10865370 TI - Developing a research unit and maintaining it. PMID- 10865369 TI - How to promote the involvement of part-time teachers to best advantage. PMID- 10865371 TI - Future priorities for a successful dental school. AB - Priorities for success for a dental school of the future are considered. University relations, demographic changes, digital information technology and functional genetic technology are emphasised as important environmental pressures that will influence the priorities. The goal advocated for university relations is creating a culture in which the dental school is viewed as integral and necessary to the University's mission. Financial stability, quality research and scholarship and provision of health care for employees may be important ingredients. Keeping an eye on demographic changes and taking the school's business where the customers are is another key to success in the future. Ethnic diversity, changing approaches with changing disease patterns, flexibility in schedule and collaborations in areas of need are strategies to be considered. The emerging field of functional genetics typifies a new biology with which currency will be needed in a health-sciences field. Necessity for faculty development, adoption of molecular diagnostic technologies, emphasis on risk assessment and preventive counselling, and a shift to a wellness model are likely consequences. Digital information technology will result in increased distance-learning opportunity. Dental schools will also need to make accommodating changes in curriculum structure available. Conversion to electronic imaging and totally electronic patient records are likely to become standard. PMID- 10865372 TI - The making of the new Dublin Dental Hospital. PMID- 10865373 TI - Guidelines for crown and bridge. British Society for Restoration Dentistry. PMID- 10865374 TI - Comparison of patients' appreciation of 500 complete dentures and clinical assessment of quality. AB - To compare patients' appreciation and clinical assessment of quality of complete dentures 500 patients attending a dental teaching hospital for provision of new complete dentures were recruited. In each case denture quality was assessed by an experienced prosthodontist using a validated classification of aspects of denture quality. Patients graded their appreciation of features of the dentures on a four point scale. Multiple correspondence analysis demonstrated close correspondence between dentist and patient appreciation of dentures when the dentures were rated as poor, but little or no correspondence when dentists or patients rated the dentures highly. PMID- 10865375 TI - Effect of aluminium oxide sandblasting on cast commercially pure titanium surfaces. AB - Studies on the titanium porcelain interface have shown the presence of alumina, attributed mainly to the sandblasting procedure. In this study, an investigation of the effect of the sandblasting procedure on the microstructure and roughness of a cast commercially pure titanium surface was undertaken using three different particle size alumina powders. The analysis showed that in all cases alumina particles are embedded into the surface layer of titanium. The use of a large particle size alumina seems to be advantageous in reducing the weight of alumina remaining on the titanium surface and while increasing the surface roughness, thus promoting mechanical interlocking with porcelain. PMID- 10865376 TI - Gender and age-related differences in the pattern of complete denture referrals from general dental practitioners. AB - This study was designed to identify gender and age-related differences among complete denture referrals from general dental practice to a dental teaching hospital over a 4 year period. Referral rates were calculated according to male/female ratios in the edentulous population. Although more women than men were referred overall, the proportion of males to females referred was similar to that in the edentulous population. Significant gender-related differences in referral patterns were identified, suggesting that edentulous males with complete denture problems, and aged 69 or less, made proportionately less use of hospital services, and proportionately more when aged 70 or over. PMID- 10865377 TI - Gender aesthetics in the natural dentition. AB - This paper questions whether the size, position and angulation of upper anterior teeth 'reflect' the gender of a patient. Forty-six casts of natural teeth were given to 11 experts. The experts were asked to assess the gender of the patients. Overall the intra-observer agreement was fair (kappa = 0.33). Inter-observer agreement was fair (kappa = 0.23). On average 55% of casts taken from men were classified as 'male' and 55% of the casts taken from women were classified as 'female'. It is concluded, on the basis of this study, experts can not distinguish gender by the visual assessment of upper anterior teeth on casts alone. PMID- 10865378 TI - A long-term follow-up of allergy to nickel among fixed prostheses wearers. AB - Skin allergic hypersensitivity to nickel is well known and well documented in the professional literature. Sixteen partially dentate patients with long-standing histories of recurrent hypersensitive skin reaction to nickel were entered into a clinical trial in 1979. All the patients received crowns or bridges made of an alloy containing 66% of nickel with porcelain fused to the metal framework. During the first three years, annual clinical dental and dermatological investigations were performed on all the subjects, including repeated patch tests. Follow-up examinations took place in 1988 and 1994. None of the subjects had any mucosal or systemic reaction following oral exposure to the nickel containing alloy, nor the exacerbations of skin lesions. PMID- 10865380 TI - A six-year follow-up study of maxillary overdentures on osseointegrated implants. AB - This study comprised a clinical and radiological examination of two groups of 14 patients receiving overdentures of the same design on osseointegrated implants. The mean follow-up period was 82 months and 35 months in the two groups the fixture success rate was 84% and 85% and the marginal bone loss 0.97 mm and 1.29 mm respectively. The follow-up of the two groups was guided by several clinical parameters and apart from the loss of fixtures and adjustments of the attachment system only a few complications were registered. The results show that this treatment method is a successful alternative in the treatment of the edentulous maxillae. PMID- 10865379 TI - Effects of superstructure type and design on force transmission via implant stabilised mandibular prostheses. AB - Forces to which individual implant units are subjected when an implant-stabilised mandibular over-denture is loaded have been measured using a carbon-fibre reinforced epoxy resin replica of an edentulous mandible mounted in a loading rig which simulated physiological conditions. Three different retentive designs were used; ball attachments, a fixed-fixed bar cranked anteriorly, and a similar design with posterior cantilevers. Forces exerted on the implants were measured using resistance strain gauges mounted on their trans-mucosal abutments and the dentures were loaded at individual tooth positions with a strain gauge beam. Ball attachments were associated with the lowest detected forces on the abutments. It is concluded that variations in superstructure design affect the forces transmitted by implants stabilising a complete mandibular over-denture, when the mandible is suspended and loaded in a physiological manner. PMID- 10865381 TI - Periodontal response to cantilevered and fixed-fixed resin bonded bridges. AB - The periodontal response to two types of resin bonded bridges was investigated. Cantilever or fixed-fixed resin bonded bridges which had been in service for at least two years were reviewed. Periodontal indices (plaque index, gingival index, bleeding index and pocket depth) were recorded for each bridge abutment and for matched contralateral teeth. There was no difference in the periodontal condition of the abutment teeth between cantilever and fixed-fixed resin bonded bridges, after they had been in service for at least two years. There was evidence of greater plaque accumulation and higher gingivitis levels affecting abutment teeth for both types of bridges compared with control teeth. PMID- 10865382 TI - Crown and bridge impressions--a comparison between the UK and a number of other countries. AB - The quality of impressions for crown and bridge work in seven countries were compared with the results found in the United Kingdom in a previous study. The results showed that metal impression trays were used more frequently, and flexible plastic trays less frequently, in the countries visited than in the United Kingdom. Other differences and similarities were noted. PMID- 10865383 TI - Bite force and occlusal load distribution in normal complete dentitions of young adults. AB - The aim of this study was to investigate the bite force and its distribution over the maxillary dentition in completely dentate subjects. Twenty-three dental students (14 Danish, 9 Japanese) with complete dentitions, but without occlusal contact on the third molars participated and their maximal bite forces were measured using the Dental Prescale System close to intercuspation. The antero posterior location of the occlusal load centre, i.e. the centre of balance of distributed occlusal load, was a little posterior of the centre of the upper first molar. These results could be used as a basic model for evaluation of occlusion. PMID- 10865384 TI - An in vitro investigation of the temperature rises produced in dentine by Nd:YAG laser light with and without water cooling. AB - Infrared lasers are reported to have thermal side effects which may damage pulp tissue. This study investigated the thermal effects of the pulsed Nd:YAG laser. Prepared, extracted teeth were measured prior to irradiation. Temperature was recorded using a thermocouple/data logging system. Laser irradiation was carried out with or without water spray for an exposure time of ten seconds. Results indicated that dry irradiation produced unacceptable temperature rises with dentine thicknesses used. Wet irradiation produced a significantly lower temperature rise. It was concluded that the Nd:YAG laser produced thermal effects which could potentially cause pulpal trauma. A water coolant was effective in reducing these thermal effects, but the temperature rise achieved whilst using water coolant may still cause pulpal damage. PMID- 10865385 TI - Metal ceramic crowns--a review of tooth preparation. AB - The metal ceramic crown has become the most popular extra-coronal restoration in the armamentarium of the dentist, certainly within the UK. It combines the aesthetic qualities of porcelain with the strength of cast metal. This review discusses a number of issues relating to these crowns, and considers the principles of tooth preparation and marginal fit. The physical requirements of tooth preparation are considered in terms of shoulder width, emergence profile, shoulder angle and effects on the pulp. It also considers the teaching of metal ceramic crown preparation, the failure of metal ceramic crowns and the influence that preparation quality has on these failures. PMID- 10865386 TI - A study on the use of the dentatus AEB face bow and its modification as an ear bow. AB - This study investigated the siting of the earbow mounting pins on the Dentatus ARL articulator relative to the condylar axis. Further, it showed the repeatability of registrations using the Dentatus AEB face bow and its modification as an earbow. Three face bow and three earbow recordings were made of 35 subjects. Registrations were fitted onto the subject's cast mounted on the articulator to show differences in the recording of the condylar axis. The mean horizontal relationship between the earbow and face bow registrations approximated to the relationship of the earbow mounting pin to the opening axis of the articulator. The mean diameter of the spread of registrations was less for the earbow than the conventional face bow. The use of the earbow modification to the Dentatus face bow is supported. PMID- 10865387 TI - Comparative evaluation of effect of auxiliary retentive features on retention of complete cast crowns in teeth with adequate and inadequate crown height. An invitro study. AB - This study was done to evaluate the retention of complete cast crowns in teeth with adequate and inadequate crown height and to evaluate the effects of auxiliary retentive features on retention from of complete cast crowns. Thirty six extracted human maxillary first premolars were obtained. In one group teeth were prepared to a constant height of 3.5 mm and in the second group teeth were prepared to a constant height of 2.5 mm. Each group was further subgrouped into three. In the first subgroup teeth were prepared conventionally, second subgroup proximal grooves were incorporated and in the third subgroup proximal boxes were incorporated. Tensional forces required to dislodge each cemented casting from its preparation was used for comparison of retention quality. These results were then subjected to an analysis of variance (ANOVA). The retention of crown in tooth with adequate crown height were statistically highly significant (P < 0001), compared to inadequate crown height and in boxes compared to grooves and conventional preparation. PMID- 10865388 TI - Finite element analysis of a three unit fixed partial denture cast with nickel chromium alloy. AB - A two dimensional mathematical model was generated, representing a three unit fixed-fixed partial denture and its supporting structures. Second premolar and second molar were used as abutments with first molar as pontic. A load of 1 kg was applied to the occlusal surface of the casting. Stresses and displacement developed at various regions were analyzed with STAAD III/ISDS program. Maximum stresses were developed in the pontic and connectors with distal connector experiencing the maximum stresses. Stresses transmitted to the dentin were comparatively lower and more of compressive in nature. The underying bone experienced moderate amount of both compressive and tensile stresses but the displacement in this tissue were minimal compared to the rest. PMID- 10865389 TI - A generational comparison of changes in mandibular third molars. AB - Due to recent increases in the number of cases of embedded mandibular third molars, the Department of Dental Radiology, Kyushu Dental College decided to investigate generational differences in their occurrence. For the purpose of investigation, 670 panoramic radiographs, obtained from two different generations (1970s and 1999s) of Kyushu Dental College students, wer used. There were significant statistical differences in the angle of eruption of the third molar between the 1970s and 1990s groups. The mandibular third molars in the 1990s group were growing at an abnormal angle (P < 0.05) and were closer to the mandibular canal (P < 0.05) than those in the 1970s group. PMID- 10865390 TI - The antimicrobial effects of extracts of Azadirachta indica (Neem) and Salvadora persica (Arak) chewing sticks. AB - Chewing sticks (Miswak) is most commonly used int he Middle East and Indian Subcontinent Salvadora persica (Arak) and Azadirachta indica (Neem) are commonly used as oral hygiene tools in different parts of the world, Several studies have demonstrated the anti-plaque anticarious and antibacterial effect of these sticks. The aim of this study was to compare the effectiveness of antimicrobial activity of Neem and Arak chewing stick's aqueous extracts at various concentrations. The microbial inhibition was measured using blood agar and ditch plate method up to 48 hous. The pH of Neem extract was 6.1 and of Arak was 4.9 Data suggested that both chewing stick extracts are effective at 50% concentration on strept mutans and Strept faecalis. Arak extract was more effective at lower concentrations for Strept faecalis. The effect may be due to the difference of their chemical composition and variability in their PH. Further research is needed to extrapolate other plants used for oral hygiene. Chewing sticks are recommended as oral hygiene tools for health promotion in developing countries. PMID- 10865391 TI - Comparative quantitative and qualitative assessment of the marginal adaptation and apposition of bonded amalgam restorations using luting glass ionomer and 4 META adhesive liner under a scanning electron microscope. An in vitro study. AB - The present invitro study was conducted to assess the marginal adaptation and apposition of amalgam restorations bonded to tooth structure, using freshly mixed luting glass ionomer cement (type 1) and compared with the much documented material--Amalgam bond (4-META). Twelve freshly extracted human premolar teeth were used and class V cavities were prepared on the buccal and lingual surfaces of twelve teeth for the experimental groups. Buccal cavities (class V) were prepared on twelve other teeth for the control group. In the control, two coats of cavity varnish was applied as the liner. The experimental groups were lined with freshly mixed luting glass ionomer in twelve buccal cavities and amalgam bond adhesive liner in twelve lingual cavities. The amalgam was then condensed before the liner sets in all the cavities. The teeth were sectioned and mounted on aluminum stubs and then placed in the scanning electron microscope and were observed. The results of the study showed that marginal interface gaps were higher with luting glass ionomer and cavity varnish than with amalgam bond as the liner. The results were statistically not significant. All the three groups were effected by the severe thermal stresses introduced by the microscopy with the glass ionomer cement suffering the worst. The present investigation confirms that glass ionomer cement (type 1) can be effectively used as a bonding agent between amalgam and the tooth when it is painted onto the cavity walls after which the amalgam is condensed immediately, prior to its setting. Its ability to bond amalgam to the tooth structure with minimal gaps is comparable to that of amalgam bond. PMID- 10865392 TI - Oral health status of the institutionalized elderly in Mangalore, India. AB - This study was carried out among 287 institutionalized elderly aged 60 years and above residing in Mangalore. The oral health status was assessed using the modified Oral Health Assessment Form. The most frequently observed periodontal condition was shallow pockets seen in 32.29% of the dentate subjects. None of the subjects were found to have a completely healthy periodontium. The dental caries prevalence was found to be 77.6%. The mean DFT of the dentate subjects was 3.64 with the mean DT and FT being 3.58 and 0.06 respectively. The mean DFT was found to increase with advancing age. Findings also suggest that edentulousness is directly proportional to the period of stay in the old age home and inversely proportional to the mobility of the individual. PMID- 10865393 TI - Evaluation of efficacy of Sofscale. A clinical and SEM study. AB - Efficacy of Sofscale in the case of removal of calculus from the root surface as claimed by manufacturers was evaluated in the present study. 32 patients suffering from chronic periodontitis, having calculus deposits in the lower incisors were selected for the study. A double blind study was conducted to evaluate the ease with which calculus could be removed. In the second part of study, a SEM study of the root surface subjected to root planing alone and root planing after application of Sofscale was done to see whether any remnants of calculus was left behind. In the double blind study conducted on 32 patients, it was found in 21 patients, calculus could be removed with greater ease and less time in experimental area compared to the control. 17 patients were able to correctly identify the product side as easier to Sofscale. Patients did not complain of increased sensitivity of teeth or discomfort following application of Sofscale. Under SEM the root surface after root planing showed the presence of smear layer whereas after Sofscale application root surface appeared devoid of smear layer. No other significant difference were noticed between the root surfaces. PMID- 10865394 TI - Efficacy of minocycline as a root conditioner in comparison to citric acid and tetracycline. An in vitro evaluation. AB - This study compared the root surface characteristics produced by Minocycline HCl, Citric acid and Tetracycline HCl. when used as a root conditioner. This study included 5 groups of 10 extracted teeth each with advanced periodontal disease. Diseased root surfaces of group B,C,D and E were planed with Gracy curets and solutions of Citric acid, Tetracycline HCl, Monocycline Hcl and Tween 80 of PH 2.5, were applied to the surface in respective groups with cotton pellets for 5 mts. Group A was not subjected to root planning or application of any root conditioner and was used as a control. The teeth in all five groups were then washed, sectioned processed and examined under scanning electron microscope. These results revealed that the surfaces of acid treated sites differed from each other and with specimens treated sites differed from each other and with specimens treated with root panning alone. The efficacy of Minocycline and tetracycline in the removal of smear layer and exposure of dentinal tubules were comparable and Minocycline HCl though not as effective as Citric acid can probably be recommended in view of its antibactrial activity, anti collagense and substantivity properties. PMID- 10865396 TI - The effect of propolis on dentinal hypersensitivity and level of satisfaction among patients from a university hospital Riyadh, Saudi Arabia. AB - Propolis has been used since ancient times in folk medicine for its beneficial effects. It is a mixture of resin, essential oils and waxes mixed with bee glue; also it contains amino acid, minerals, ethanol, vitamin A, B Complex, E, Pollen and highly active ingredient known as Bioflavenoid (Simon Martin 1991). To date, there have been no studies done on desensitizing effect of Propolis in vivo. So, the aims of this study are to evaluate the effect of Propolis in a clinical trial on subjects with dentinal hypersensitivity, and to assess the level of satisfaction among subjects after they have used the Propolis. Twenty six Saudi female subjects at King Saud University, College of Dentistry, Riyadh, Saudi Arabia, age ranged from 16-40 yr. (mean = 23 yrs.), were included in the study over a period of four weeks. The Cervical Dentinal Sensitivity (C.D.S.) was assessed by two methods: 1--cold air stimuli. 2--Subjective reporting of pain, with a modified questionnaire (Gillam 1997) and a numerical scale 0-10 (Downie et al 1978). The patients were followed up for four weeks after using Propolis, there were two recalls, and the first recall was after one week and the second recall after four weeks of the base line. The results between the base line findings and after four weeks were statistically significant. Eighty five percent of the subjects were found highly satisfied; the Propolis had significant effect on dentinal hypersensitivity during the study period. Further research is needed with double blind clinical trial on a large sample size. PMID- 10865395 TI - Comparative evaluation of the effect of simulated porcelain firing cycle on the mechanical properties and microstructure of base metal ceramic alloys. AB - A comparison of mechanical properties and microstructure of four metal ceramic alloys in as-cast and heat-treated conditions resulted in significant differences. The alloys that were tested included two nickel-based and two cobalt based metal ceramic alloys. Mechanical properties tested included strength, percent elongation and hardness. Ten tensile bars were cast for each alloy. Five of the ten bars for each alloy were randomly selected for heat treatment with the simulated porcelain firing cycle. Results indicated that the nickel-based alloys were weakened by the heat treatment and showed a decreased hardness and an increased elongation. The effects of heat treatment on the strength of cobalt based alloys were variable, showing an increase in hardness and a decrease in elongation. Differences in mechanical properties exhibited by the alloys were related to compositional and microstructural changes. PMID- 10865397 TI - Acute toxicity (LD50 and CD50) of lidocaine and prilocaine in combination with adrenaline and felypressin. AB - The toxicity of the combination of vasoconstrictors to local anaesthetic solutions has been debated since its first use in the beginning of this century. A combination of two vasoconstrictors to a local anaesthetic has been proposed by some researchers. In this study they were evaluated the acute toxicity (lethal dose 50%, convulsion dose 50%) and latency times of loss of righting reflex and convulsion as well as the duration of convulsion) of 2% lidocaine or 3% prilocaine, when administered in combination with adrenaline and felypressin at various concentrations. Lethal dose 50% studies showed that for both anaesthetics the solutions with higher concentrations of adrenaline were more toxic. The opposite was observed in the convulsion dose 50% studies. No alterations were observed in the control groups. All lidocaine solutions increased the latency of loss of righting reflex. The latency of convulsion was increased in some groups, but once the convulsion was achieved there was no difference in its duration. There was no statistical difference among prilocaine groups for any of the variables studied. Based on the experimental model studied, it was concluded that there is no advantage in the association of two vasoconstrictors concerning the toxicity of lidocaine and prilocaine solutions. PMID- 10865398 TI - Clinical evaluation of sodium flouride chewable tablets in dental caries. AB - Chewable tablets containing low dosage flouride content were prepared using two varities of celluloses and their in vitro parameters were evaluated. An eighteen month clinical trial revealed that both these formulations were effective in controlling the caries. However, ethyl cellulose is proved to be superior to methylcellulose as a controlled release matrix material in controlling caries. Thus this study recommends ethylcellulose matrix tablets containing low flouride content is an efficacious and cost effective drug device in controlling dental caries. PMID- 10865399 TI - Granular cell tumor of the common bile duct: a case report. AB - Granular cell tumors (GCTs) are uncommon soft tissue tumors, usually presenting in the skin and subcutaneous tissue tongue and oral cavity. We present a case report of granular cell tumor of the common bile duct involving both extra- and intrapancreatic portions. The histogenesis appears to be related to Schwann cells, similar to granular cell tumors of other sites, as evidenced by histologic and immunohistochemical findings. Review of the English literature concerning biliary tract GCTs revealed a high occurrence in African-American females in their third decade. By-pass operation to correct the biliary tract obstruction may be appropriate, if the nature of the tumor can be obtained from intraoperative diagnosis by frozen section. PMID- 10865400 TI - Parvovirus B19 caused fetal death: a case report in Thailand. AB - This was a case of an intrauterine parvoviral B19 infection resulting in hydrops fetalis and enlarged placenta. Histologically, the virus was found to be in nucleated red cells of the fetus which was confirmed by electron microscopy. Careful placental examination at the gross and microscopic levels yielded the correct diagnosis. PMID- 10865401 TI - Extramedullary haematopoietic tumor producing small intestinal intussusception in a beta-thalassemia/hemoglobin E Thai boy: a case report. AB - The authors report a case of beta-thalassemia/hemoglobin E disease with extramedullary haematopoietic tumor which developed at the small intestine and caused intussusception. A 7 year-old boy with homozygous beta thalassemia/hemoglobin E presented with recurrent abdominal pain. The abdominal ultrasonography showed ileo-ileal intussusception with a solid mass as the leading point. Resection of the ileal segment was performed. Pathological examination revealed an extramedullary haematopoietic tumor forming an intraluminal polypoid mass, being the leading point of the intussusception. Extramedullary haematopoiesis in the intestinal tract is rare. To our knowledge, this is the first case of extramedullary haematopoietic tumor that produced intussusception of the small intestine in a beta-thalassemia/hemoglobin E patient. PMID- 10865402 TI - Congenital nasopharyngeal teratoma (epignathus). AB - The case report of a congenital nasopharyngeal teratoma presented as an intra amniotic sac mass was described. Histologically, the tumor was composed of highly differentiated tissue derived from three germ layers. Those tissues were partially developed as infant organs with a central core made up by bone. Haphazardly arrangement of such tissues with disorganized sketal axis helped in differentiating the mass from an asymmetric twins. The evidence of tumor attachment was seen at the retro-uvular region. PMID- 10865403 TI - Preliminary report for the application of nucleic acid sequence-based amplification in detection of human cytomegalovirus (HCMV) late pp67 mRNA for diagnosis of HCMV infection. AB - Human cytomegalovirus (HCMV) late pp67 mRNA expression by nucleic acid sequence based amplification (NASBA) in patients, clinically diagnosed as possible HCMV, probable HCMV disease, and no disease, was evaluated. The RNAs were isolated from 11 whole-blood samples of 11 patients for the specific amplification of the pp67 mRNA. NASBA results were compared to results from PCR assay and serological assay. The HCMV pp67 mRNA could be found in 3 of 11 patients, whereas, HCMV-DNA PCR was positive in 6 of 11 patients. PCR assay for HCMV-DNA in plasma has proved to correlate with clinical diagnosis of HCMV infection. Only 2 patient samples of NASBA positive results coincided with HCMV-DNA PCR. However, the diagnosis of clinically relevant HCMV infection by NASBA was seen. Anti-CMV IgG titers of 1:1,600 or over 1:1,600 were found in 2 of 3 NASBA positive cases and 5 of 6 HCMV DNA positive cases, whereas, anti-CMV IgM were all negative. These results showed the correlation of HCMV infection detected by NASBA, PCR assay and anti-CMV IgG of the titers up to 1:1,600. Additionally, a low antibody titer of the HIV patient could be diagnosed by NASBA or PCR. In conclusion, pp67 mRNA NASBA appears to be a promising diagnostic tool in analysis of HCMV infection and/or disease. Its diagnostic value should be defined in the specific group for the follow-up of immunocompromised patients, such as organ transplant recipients in future prospective studies. PMID- 10865404 TI - The organization of Kidney Transplantation Services at Ramathibodi Hospital: fourteen years experience on waiting list, kidney donors and kidney transplantation. AB - The Kidney Transplantation Program at Ramathibodi Hospital was established in 1985. By the end of 1998, there were 1,614 patients on the cumulative waiting list. The first kidney transplantation (KT) was started in 1986 by using kidney from living-related donor (LD) while cadaveric KT (CD-KT) was started in 1987. A total of 528 KT were done, 278 cases (52.7%) were CD-KT and 250 cases (47.3%) were LD-KT. Six patients had two kidney transplants. 278 kidneys were donated from 189 cadaveric donors. Fifty cadaveric donors (26.4%) were from Ramathibodi Hospital while the rest were from other hospitals and the Organ Donation Center, Thai Red Cross Society. For LD, 233 out of 250 (93.2%) were from living-related, more than 50 per cent of these donors were from siblings. 17 spousal donors have been accepted for KT at Ramathibodi Hospital since 1997. Concerning the recipient pools, 522 patients (32.3%) were transplanted, 123 patients (7.6%) died without KT, 111 patients (6.9%) underwent KT at other hospitals, and 78 patients (4.8%) changed to waiting lists at other hospitals. The rest were lost to follow-up. At present, only 265 patients are still actively waiting (send serum every month). The number of KT and living donors has gradually increased, whereas, the number of cadaveric donors has decreased. However, cooperation with the "Organ Donation Center" has improved the number of cadaveric donation in the last two years. Sufficient organ donations and an active working team will provide a good kidney transplant service for the patients. PMID- 10865405 TI - Mycetoma and phaeohyphomycosis caused by Phialophora parasitica in Thailand. AB - Phaeohyphomycosis caused by Phialophora parasitica is rare and it has never been documented in Thailand. The first two Thai cases of phaeohyphomycosis caused by P. parasitica were recognized in early 1990 at Ramathibodi Hospital, Bangkok, Thailand. Both patients had underlying diseases. The fungus developed in abscesses with pigmented mycelium at the lower extremity. Cultures from pus and tissue biopsies were positive for dematiaceous fungi. Light microscopic features suggested P. parasitica and which was illustrated by both scanning and transmission electron microscope. The first case was treated with itraconazole with a satisfactory initial response. The second case was successfully treated by surgical removal of the entire lesion. PMID- 10865406 TI - Lewis antigens on newborn red cells. AB - This study aimed to screen for Lewis antigens in Thai newborns. Although, these antigens are known to be weak or absent on the red cells of newborns, we encountered a case of a Le(a+) newborn baby when testing with monoclonal antibody and human anti-Le(a). Such a finding led us to conduct this study to explore further evidence of Lewis antigens in Thai newborn red cells. A total of 197 cord blood samples were tested with monoclonal anti-Le(a) and anti-Le(b). (Bioclone, Ortho Diagnostic Systems, USA). The tests were performed according to the manufacturer's recommendations. The results revealed that none of the cord red cells in this study group possessed Lewis antigens. This study showed that Lewis antigens were absent or were so extremely weak on the red cells of these newborn infants that they could not be demonstrated despite the use of potent monoclonal antibodies. However, further study should be done by using more cord blood samples, a more sensitive technique or even more potent antisera. PMID- 10865407 TI - Forensic identification by DNA fingerprinting and mitochondrial DNA typing. AB - Short tandem repeats (STRs), that represent an important source of highly polymorphic markers in human genome, and mitochondrial DNA (mtDNA) typing, that its sequences were conserved within the same maternal lineage, facilitated by use of the polymerase chain reaction (PCR) provide a powerful tool for forensic identification. We report the analysis of 9 STR loci and mtDNA typing of a muscle biopsied sample with 2 months postmortem by comparison with the genotype of the relative. The DNA profile showed common alleles with that of the relative but only 12 from 20 alleles (60%) were identifiable. Then, we performed mt DNA sequencing of the hypervariable region I (HV I) and obtained 100 per cent homology with that of the relative. In conclusion, personal identification can be performed precisely by the data of DNA profile and mtDNA typing compared to the genotype of the relative. PMID- 10865408 TI - Paternity testing by PCR-based STR analysis. AB - We present application of polymerase chain reaction (PCR)-based short tandem repeat (STR) system for use in paternity testing. The process involves a single tube multiplex PCR of 9 STR loci on different chromosomes, in conjunction with Amelogenin sex test and internal size standards, followed by using an automated DNA sequencer to detect amplified products. The results showed that this system provided unambiguously reliable results. In addition, the method is useful for routine use in that it is robust and reproducible and provides a reliable means of paternity testing. PMID- 10865409 TI - Serum C-reactive protein level in postsplenectomized thalassemic patients. AB - C-reactive protein is an established marker for the detection of acute and chronic inflammatory processes. The most potent stimulator for the hepatic synthesis of this protein is interleukin 6. Previous studies have shown that inflammatory cells and inflammatory cytokines, such as interleukin 6, interferon gamma, etc were elevated in postsplenectomized thalassemic patients. The aim of this study was to determine serum C-reactive protein concentration in postsplenectomized beta thalassemic patients (beta thal/HbE postsplenec), and to compare them with those in nonsplenectomized beta thalassemic patients (beta thal/HbE), postsplenectomized non thalassemic patients (postsplenec), reactive thrombocytosis (RT), chronic myeloproliferative disorders (MPD) and normal adult volunteers. Serum C-reactive protein concentration as determined by an automatic Behring Nephelometer was carried out in 28 beta thal/HbE postsplenec, 22 beta thal/HbE, 12 postsplenec, 23 RT, 21 MPD, and 26 healthy adult volunteers. The values of CRP in beta thal/HbE postsplenec were significantly higher when compared with beta thal/HbE, and normal volunteers (4.1 +/- 0.7 vs 1.6 +/- 0.4 mg/L P = 0.006, and 4.1 +/- 0.7 vs 0.45 +/- 0.09 mg/L, P < 0.001). CRP levels in beta thal/HbE postsplenec were also higher than the postsplenec group (4.1 +/- 0.7 vs 0.19 +/- 0.7 mg/L P = 0.095). On the contrary, they were significantly lower than those in RT (4.1 +/- 0.7 vs 55.4 +/- 14.8 mg/L, P = 0.002). However, when compared to those with MPD, the values were not statistically different (4.1 +/- 0.7 vs 17.1 +/- 12.3 mg/L, P = 0.871). Interestingly, there was a trend towards increasing C-reactive protein levels in beta thal/HbE postsplenec patients with higher platelet count, although no correlation was observed. Besides the inflammatory process, platelet and/or factor(s) that control(s) thrombopoiesis seem(s) to play a role in the high serum C-reactive protein levels in the studied population. PMID- 10865410 TI - BCR/ABL rearrangement in Philadelphia chromosome negative CML patients. AB - Reverse transcription-polymerase chain reaction (RT-PCR) was used to study 34 patients with chronic myelogenous leukemia (CML) associated with negative Philadelphia (Ph) chromosome. This report showed evidence of a chimeric BCR/ABL transcript in 18 (52.9%) and 28 (82.4%) cases by first PCR and seminested PCR, respectively. In these BCR/ABL transcript positive cases, the incidence of BCR exon3/ABL exon2 (B3A2) and BCR exon 2/ABL exon2 rearrangement was 25 (89.3%) and 3 (10.7%) cases, respectively. The other 6 Ph negative patients showed no evidence of reciprocal translocation of BCR to chromosome 9. This data demonstrates that seminested PCR is sufficiently sensitive to detect BCR/ABL fusion transcript in Ph chromosome negative CML patients. PMID- 10865411 TI - Fluorescence in situ hybridization: a rapid analysis to verify chromosome aberrations. AB - We applied fluorescence in situ hybridization (FISH) to assess the presence of structural rearrangement and numerical chromosome aberrations in both metaphase chromosome and interphase nuclei. For this purpose, the biotinylated repetitive alpha-satellite DNA probes for chromosome 1, 18 and 8 (pUC1.77, L1.84 and pJM128) were used to identify tetraploid mosaicism, ring chromosome 18 and trisomy 8 mosaicism for pre-, post-natal and tumor diagnosis respectively. Utilizing this approach, we showed the usefulness of FISH for routine clinical cytogenetics in addition to chromosome banding techniques. The chromosome aberrations with unknown or unclear origin, detected by chromosome analysis, could be confirmed accurately and rapidly. PMID- 10865412 TI - Comparison of DNA extraction from blood stain and decomposed muscle in STR polymorphism analysis. AB - Forensic samples that are often degraded and limited in quality cause DNA typing analysis by conventional methods unsuitable. We performed a single tube-multiplex PCR on 9 STR loci (D3S1358, vWA, FGA, TH01, TPOX, CSF1PO, D5S818, D13S317, and D7S820) and the X-Y homologous gene amelogenin of DNA extracted from six week postmortem blood stain and decomposed muscle by using QIAGEN QIAamp blood or tissue procedure. An automated genetic analyzer based on fluorescent dye technology was used to detect STR allele patterns. The DNA profile of blood stain sample obtained a complete and unambiguous pattern, whereas, that of muscle DNA extracted from QIAamp tissue and Chelex plus QIAamp blood protocols showed detected STR alleles for 70 per cent and 50 per cent of all tested alleles, respectively. The degraded muscle DNA could not yield amplified products of large size STR alleles; CSF1PO, D13S317 and D7S820. However, the analysis which relied upon the PCR-based STR polymorphism analysis and automated genetic analyzer system offers an ideal strategy for forensic identification. PMID- 10865413 TI - Identification of sex chromosome by fluorescence in situ hybridization. AB - Fluorescence in situ hybridization (FISH) is the complicated and very effective technique to determine the origin of chromosome material that cannot be identified by conventional banding techniques. Also determining the hidden sex chromosome and the percentage of mosaicism. Five peripheral blood and one cord blood sample were used to perform centromeric X and Y chromosome-specific DNA probe to determine the sex chromosome. Comparing the percentage of mosaicism between conventional cytogenetic technique and FISH technique, we found a different ratio in mosaicism. That is because the molecular cytogenetic study was the evaluation of chromosome identification in both dividing (metaphase) and non dividing cells (interphase nuclei). PMID- 10865414 TI - Etiology and incidence of thrombotic and hemorrhagic disorders in Thai patients with extreme thrombocytosis. AB - A retrospective study of 126 patients with extreme thrombocytosis (defined as a platelet count > or = 1,000 x 10(9)/L) was performed during a five-year period (June 1994-June 1999). The aim of this study was to determine the etiology and to evaluate the clinical consequences of extreme thrombocytosis. Seventy patients (55.5%) had reactive thrombocytosis (RT) with an age range of 43 +/- 2.2 years, 56 (44.5%) had chronic myeloproliferative disorders (MPD) with an age range of 53 +/- 2.4 years. Underlying causes of RT were malignancy (25/70 or 35.7%), infection (16/70 or 22.9%), postsplenectomized beta-thalassemia/Hb E (11/70 or 15.7%), inflammation (12/70 or 17.1%), iron deficiency anemia (6/70 or 8.6%). Duration post splenectomy in our beta-thalassemia/Hb E patients ranged from 4 months to 21 years, with a median of 10 years. Subtypes of our MPD cases were chronic myeloid leukemia (30/56 or 53.6%), essential thrombocytosis (18/56 or 32.1%), polycythemia vera (4/56 or 7.1%), agnogenic myeloid metaplasia (3/56 or 5.4%) and unclassified MPD (1/56 or 1.8%). Bleeding and thrombotic tendency were respectively noted in 7 (12.5%) and 2 (3.6%) of MPD patients. Two patients of the MPD group (3.6%) experienced both bleeding and thrombotic episodes. One patient (1.4%) of the RT group developed vasculitis-associated thrombosis. However, none of the patients in the RT group had bleeding complications. Extreme thrombocytosis was not a rare condition in a university hospital population, and bleeding and/or thrombotic complication was more common in the MPD group. PMID- 10865415 TI - Evaluation of the effect of steviol on chromosomal damage using micronucleus test in three laboratory animal species. AB - The chromosomal damage activity of steviol, a product of enzymatic alteration of stevioside, a natural non-caloric sweetener was reevaluated by using a bone marrow micronucleus test in both male and female hamsters, rats and mice. The micronucleus test is used widely as a rapid and efficient alternative in chromosome analysis for detecting in vivo cytogenetic damage. Steviol at the dose of 4 g/kg body weight for hamsters and 8 g/kg body weight for rats and mice showed no effect on the frequencies of micronucleus formation in bone marrow erythrocytes of both male and female hamsters, rats and mice. Moreover, there was also no apparent change in the PCEs:NCEs (polychromatic erythrocytes:normochromatic erythrocytes) ratio of the male animals of all three treated species at 24, 30, 48 and 72 hour intervals. However, steviol at the given dose can cause significant reduction of PCEs to NCEs ratio of the female hamsters at 72 hours and female rats and mice at 48 and 72 hours after receiving steviol orally. From these results, it could be proposed that steviol at the given dose to the treated animals produced adverse metabolites and these metabolites could reach the bone marrow, the target organ for micronucleus test. These metabolites also exhibited a slightly cytotoxic effect but not clastogenic effect to the bone marrow erythrocytes. PMID- 10865416 TI - The effects of cigarette smoking on peripheral blood leukocytes and lymphocyte subpopulations: an urban population-based study in Thailand. AB - Preliminary studies for peripheral blood leukocytes and lymphocyte subsets were done in smokers and non-smokers. There were 20 smokers (smoked more than 10 cigarettes per day) for more than a year and 20 non-smokers (smoked less than 20 cigarettes/20 years). Ages of smokers and non-smokers were respectively 21-57, and 18-55 years. Cigarette smoking was associated with a statistically significant increase in the number of neutrophils, activated lymphocytes, CD25 and CD19; but a statistical decrease in the percentage of CD7 and CD3. (P < 0.05) PMID- 10865417 TI - Reticulocyte analysis in iron deficiency anemia and hemolytic anemia. AB - Reticulocyte analysis was studied in 28 anemic patients, 15 with iron deficiency anemia (IDA), and 13 with hemolytic anemia including 9 glucose 6 phosphate dehydrogenase deficiency (G6PD def.), and 4 with G6PD def. combined with HbE trait or alpha thalassemia trait (alpha thal trait). The reticulocyte analysis among these patients showed the increased number of reticulocyte percentage with low degree of maturation in both IDA and G6PD def. patients. The significantly decreased reticulocyte hemoglobin content (CHr) was found in IDA (CHr = 21.74 +/- 4.61 pg in IDA vs 28.41 +/- 1.34 pg in normal; p-value = < 0.0001), whereas, increased CHr was found in G6PD def. patients. In addition, the G6PD def. patients also showed a significant increase in mean corpuscular reticulocyte volume (MCVr) when compared to normal (MCVr = 132.0 +/- 8.39 fl. in G6PD def. vs 110.39 +/- 5.09 in normal; p-value = < 0.0001). However, a significant decrease in MCVr was found in IDA patients (MCVr = 95.89 +/- 8.57 fl.; p-value = < 0.0001 vs normal). From this study, we can suggest that the reticulocyte hemoglobin content (CHr) and mean corpuscular reticulocyte volume (MCVr) are the important defects in patients with iron deficiency anemia. PMID- 10865418 TI - Survival analysis of Thai patients with IgM nephropathy, focal segmental glomerulosclerosis and membranous nephrotic syndrome. AB - Long term outcome of 124 Thai adult nephrotic patients was determined. Nephrotic syndrome affects the young more often than the old (median age 29 years). The most common pathology was IgM nephropathy (45.2%), membranous nephropathy (31.5%) and FSGS (23.4%). Sixty-four per cent of patients with IgM nephropathy respond to corticosteroid within 4-8 weeks while twenty three per cent were late responders. However, more than half of these patients were relapsers or steroid dependent. Response to corticosteroid occurred in 48.2 per cent of patients with FSGS while the response rate of patients with membranous nephropathy was only 23.1 per cent. Survival analysis revealed that five and ten years renal survival of IgM nephropathy was 98 per cent. Five and ten years renal survival of FSGS was 83.7 per cent and 76.8 per cent while those of membranous nephropathy was 95 per cent and 63.3 per cent. The response to corticosteroid was associated with better prognosis in FSGS. Our results show that patients with IgM nephropathy and membranous nephropathy have a generally good prognosis. Renal function is usually well preserved for at least ten years. The prognosis of patients with FSGS varied and correlated with the degree of steroid responsiveness. PMID- 10865419 TI - Preliminary study of HLA-ABCDR antigens in CML, ANLL, thalassemia and severe aplastic anemia in Thais. AB - The objective of this study was to analyse human leukocyte antigen (HLA) and disease association in common blood diseases [chronic myelogenous leukemia (CML), acute nonlymphocytic leukemia (ANLL), thalassemia and severe aplastic anemia] in Thais. The subjects were patients from the Hematological Clinic, Departments of Medicine and Pediatrics, Ramathibodi Hospital who were referred for HLA typing for bone marrow transplantation (BMT) at the Histocompatibility Laboratory from March 1988 to September 1997. A total of 129 patients had complete HLA-ABC typing. The patients included 45 CML, 40 ANLL, 26 thalassemia (Thal) and 18 severe aplastic anemia (SAA). Of these, 88 patients were typed for HLA class II. The HLA class I (ABC) and II (DR, DQ) typings were performed by microlymphocytotoxicity test. It was found that HLA class I was associated with CML, ANLL and Thal, whereas, HLA class II was associated with SAA. HLA-B8 and HLA B18 were increased in CML with R.R. values of 12.2 and 3.9, respectively, whereas, HLA-B18 was increased in ANLL with R.R. value of 4.5. In addition, HLA DR2 and DR3 were increased in SAA with R.R. values of 3.8 and 4.8, respectively. For Thal, HLA-A2 and B46 were increased in Thal in Central Thais with R.R. values of 3.3 and 6.1, respectively, whereas, HLA-B13 was increased in Thal in Northern Thais with R.R. value of 8.5. On the other hand, HLA-B7 was absent in CML. HLA Cw7 was decreased in CML and SAA, whereas, HLA-DR6 was decreased in ANLL and SAA. Furthermore, HLA-Cw6 was also decreased in CML, whereas, HLA-A33 and Bw4 were decreased in SAA. Although the sample size of each disease was small, the increase of HLA-DR2 was observed in SAA in Thais which was similar to other studies in different ethnic groups. These preliminary data may be useful for further study in HLA and blood disease association. PMID- 10865420 TI - Iron stores in autologous blood donors. AB - Depletion of body iron stores is a major factor limiting regular blood donation in volunteer donors. Autologous blood donors are requested to donate more frequently. To determine iron stores in autologous donors, 9 men and 10 women studied gave a total of 24 donations before their elective surgery (range 1-2 donations). All donors were tested for serum ferritin (SF) and hemoglobin (Hb) level. Iron supplements were taken by 88.89 per cent of men and 90 per cent of women. Mean SF before donations was 147.75 ng/mL in men and 53.19 ng/mL in women. After donations, mean SF decreased to 124.26 ng/mL in men and 38.81 ng/mL in women. None of them had depleted iron stores (SF < or = 15 ng/mL). In conclusion, iron supplementation was beneficial in maintaining body iron stores in autologous blood donors. PMID- 10865421 TI - The effect of parity on lipid profile in normal pregnant women. AB - A cross sectional study was carried out at Ramathibodi Hospital between June and August 1997. The objective of this study was to determine maternal serum lipid levels at delivery and the effect of parity on maternal lipid profile. Study population was normal term pregnant women aged 20-35 years who delivered normal infants with a birthweight > or = 2,500 grams. Maternal serum lipid levels at delivery were determined from 177 normal term pregnant women. Their mean age was 27.6 +/- 4.5 years. The first parity (P1) was about 52 per cent, whereas, the second and third parity (P2 and P3) were 37 and 11 per cent, respectively. Mean maternal serum total cholesterol (TC) levels in P1, P2 and P3 were 258.3 +/- 46.9, 266.7 +/- 47.1 and 295.7 +/- 61.2 mg/dl, respectively. Serum triglyceride (TG) levels in P1, P2 and P3 were 265.2 +/- 81.1, 280.3 +/- 72.1 and 260.7 +/- 82.8 mg/dl, respectively; serum low density lipoprotein-cholesterol (LDL-C) in P1, P2 and P3 were 136.9 +/- 45.2, 144.9 +/- 43.3 and 173.4 +/- 62.1 mg/dl, respectively; and serum high density lipoprotein-cholesterol (HDL-C) levels were 64.6 +/- 16.6, 65.7 +/- 17.8, 67.2 +/- 16.0 mg/dl, respectively. Serum TC and LDL C levels increased with parity. There was a significant difference between maternal TC and parity (F = 4.702, p = 0.01) as well as LDL-C and parity (F = 4.883, p < 0.01), especially P1 and P3. There was no significant difference between maternal TG and parity as to HDL-C and parity (p > 0.05). PMID- 10865422 TI - Iron stores in Thai blood donors. AB - A high number of blood donations may cause iron depletion. In order to evaluate iron stores in volunteer Thai blood donors, 82 male and 72 female donors were studied. All were tested for serum ferritin (SF), hemoglobin (Hb) level and asked for detailed histories of donations and iron supplementation. Mean SF in first time donors was 161.12 ng/mL in men (n = 16) and 53.92 ng/mL in women (n = 23). Mean SF in multiple-time donors was 52.72 ng/mL in men (n = 66) and 25.72 ng/mL in women (n = 49). Depleted iron stores (SF < or = 15 ng/mL) were found in 8.7 per cent of first-time female donors, 21.21 per cent and 32.65 per cent of multiple-time male and female donors, respectively. The mean numbers of total donation were 51.42 +/- 30.8 in men and 8.22 +/- 6.97 in women. The estimation of depleted iron stores from Hb level could be determined in 57.14 per cent of male and 88.89 per cent of female donors. In conclusion, iron supplementation will benefit female donors and multiple-time male donors. The frequency of donations per year was more predictive of decreased iron stores than the number of total donations. PMID- 10865423 TI - Semiquantitative determination of urinary glucose: comparison of home-made strip and routine tests. AB - Glucose oxidase paper strips for semiquantitative determination of glucose in urine are commercially available but details of their preparation are not published. The purpose of this study was to produce such strips, using a coupled enzyme glucose oxidase-peroxidase reaction which would yield purple color with ortho-tolidine, and safranin upon dipping into urine containing glucose. In this present study, without any special equipment, special humidity and temperature control, the new reagent strip named R-strip could be successfully prepared in the atmospheric conditions of Thailand. R-strips were evaluated against random urine added with various amounts of glucose in comparison with a commercial strip (T-strip), Benedict's test, and a commercial tablet (C-tablet), routinely used in laboratories. The developed strips were found to be as specific as T-strip and more sensitive than other tests. PMID- 10865424 TI - [Characteristics of penicillinase plasmids from Staphylococcus aureus strains]. AB - Penicillinase plasmids are present in most MRSA strains. They are very varying in their genotype and phenotype they confer. Penicillinase plasmids were transduced from 80 hospital MRSA strains to NCTC 8325 and the phenotype as well as the incompatibility group of plasmid were determined. Resistance to cadmium (high and low level), resistance to organic and nonorganic mercury compounds, arsenate/arsenite/antimonium resistance, resistance to bismuth and hypersensitivity to bismuth, resistance to macrolides as well as beta-lactamase production and its inductibility were checked. Among the examined strains 20 different phenotypes of penicillinase plasmids were found. Patterns of penicillinase plasmids were compared to DNA patterns of the investigated strains after digestion with SmaI and separation in pulsed field electrophoresis (PFGE). It was shown that strains with the same PFGE pattern often differ in the type of their penicillinase plasmid. Determining of penicillinase plasmid phenotype could be useful in differentiating S. aureus strains sharing the same pattern of PFGE. PMID- 10865425 TI - [Localization of genes conferring resistance to selected groups of antibiotics in methicillin-resistant strains of Staphylococcus aureus]. AB - Localization of genes conferring resistance to MLS, tetracyclines, chloramphenicol, gentamycin and neomycin in 80 MRSA strains isolated from hospital specimens was determined. The obtained results were compared to DNA patterns of the examined strains after digestion with SmaI and separation in pulsed field electrophoresis (PFGE). It was shown that genes of resistance to MLS (ErmI+) in the case of 13 strains were located on chromosome and in the case of 37 strains on plasmids (16 strains had ErmI+ and 21 strains had ErmI-). Genes determining resistance to tetracyclines were localised on chromosome in the case of 39 (23 strains possessed TetK, 11 strains had TetM and 5 strains possessed both TetK and TetM determinants) and in the case of 32 strains on plasmids. Chloramphenicol resistance genes were localised on plasmids in all 30 resistant strains. Genes conferring resistance to gentamycin were present in 31 of the investigated strains on chromosome and in two strains on plasmids. Neomycin resistance genes were plasmid in 34 strains. It was shown that the localization of the resistance genes and the PFGE patterns of the investigated strains were highly correlated. PMID- 10865426 TI - [Sensitivity to vancomycin and teicoplanin of coagulase-negative staphylococci isolated from clinical specimens]. AB - The frequency of resistance and elevated resistance to teicoplanin and vancomycin among 689 strains of coagulase-negative staphylococci isolated in one year from clinical specimens was determined. Using ATB.STAPH test, a resistance was shown mainly among strains of S. epidermidis and S. haemolyticus. The elevated resistance to teicoplanin was much more frequently observed than to vancomycin. About 27% of isolated strains of S. haemolyticus and 6.8% of S. epidermidis were classified as resistant. Among other species only single strains were recognised as resistant: one strain of S. xylosus, one of S. cohni and one of S. intermedius. 94.7% of S. epidermidis and 100% of S. haemolyticus strains classified as resistant to teicoplanin in ATB showed MIC values 14 mg/l. Moreover it was shown that 26.3% of these strains of S. epidermidis and 33.3% of S. haemolyticus had MBC of teicoplanin values equal to or higher than 32 mg/l. PMID- 10865427 TI - [Predominance of phage group II in Staphylococcus strains isolated from humans]. AB - 2545 strains of Staphylococcus aureus isolated from 24 Polish laboratories in the years 1994-95 were investigated. Phage typing was performed according to the method of Blair and Williams using the present basic set of typing phages and additional phages 88, 89 and 187. The phages were employed in concentrations of RTD and 100 x RTD. The predominance of phage group II, reported elsewhere since 1980-ties, was found in the present study (23.6%). Strains of group III were second in frequency (16.6%), whereas strains of groups I and V, as well as type 95 occurred in small percentage (7.6%, 4% and 3.4% respectively). Strains of groups II and V have been rarely lysed by phages belonging to other groups. The use of additional phages resulted in typability of strains by 7.9%. Percentage of non-typable strains was high and amounted to 22.0%. PMID- 10865428 TI - [Evaluation of the usefulness of new international experimental phages for typing methicillin resistant Staphylococcus aureus (MRSA)]. AB - The aim of the study was to determine the usefulness of the set of experimental phages obtained from the Central Public Health Laboratory in London for typing of MRSA strains in Poland. The study was performed on 150 MRSA strains isolated from various clinical materials in various regions of the country. The set of 10 experimental phages and the international basic set of 23 phages were used for typing. The results of the study showed that 76.8% of MRSA strains were typing with the experimental set of phages. The frequency of inhibition reactions was 19.9%. Only 3.3% of the strains were nontypable with the new phages while nearly half of the studied strains were nontypable with the basic set of phages. The studied strains were divided into 19 phagotypes. There was a high frequency of typable strains among MRSA typable and nontypable strains and those inhibited by the basic set of phages (71.4%-85.7%). These data indicate that the set of 10 experimental phages is useful for typing of MRSA strains isolated in Poland except for phage M3 which failed to react with almost all the strains and should be excluded from the proposed set. PMID- 10865429 TI - [Evaluation of interactions between strains of S. aureus isolated from different clinical specimens with peripheral blood phagocytic cells]. AB - The aim of this study was to investigate the interactions occurring between peripheral blood phagocytes and strains of S. aureus isolated from different clinical specimens (blood, respiratory tract, pus). To evaluate the sensitivity of microorganisms to bactericidal activity of phagocytes, monocytes and granulocytes separated from peripheral blood by standard density gradient and by counter-current centrifugal elutriation were incubated with suspensions of opsonized bacteria. In parallel, the viability of phagocytes was examined by flow cytometry, and the ability of bacteria to trigger reactive oxygen intermediates (ROI) production was evaluated by chemiluminescence measurement. To investigate efficiency of phagocytosis, bacteria were labelled with fluorescein isothiocyanate (FITC) and the percentage of cells containing FITC-labelled bacteria was analysed by flow cytometry. The data obtained show that strains of S. aureus originated from different clinical specimens, differ in their sensitivity to bactericidal activity of phagocytes--strains isolated from the blood show the highest, but strains isolated from respiratory tract show the lowest sensitivity for killing. These strains differ too in their ability to trigger monocyte CL response. Contrary, there was no difference in toxicity of bacteria against phagocytes. Strains isolated from peripheral blood showed significant negative correlation between the ability to trigger CL response and toxicity against phagocytes. PMID- 10865430 TI - [The range of antagonistic effects of Lactobacillus bacterial strains on etiologic agents of bacterial vaginosis]. AB - Bacterial vaginosis is caused by uncontrolled sequential overgrowth of some anaerobic bacteria: Gardnerella vaginalis, Prevotella bivia, Bacteroides spp., Peptostreptococcus spp., Mobiluncus sp. usually occurring in stable numbers in the bacterial flora of healthy women. On the other hand, different species of bacteria belonging to the genus Lactobacillus, most frequently L. plantarum, L. rhamnosus and L. acidophilus, form a group of aerobic bacteria dominating in the same environment. The diversity and density of their populations depend on the age and health conditions. Thanks to their antagonistic and adherence properties bacteria of the genus Lactobacillus can maintain a positive balance role in this ecosystem. The aim of this study was to assess the antagonistic properties of Lactobacillus strains isolated from the vagina of healthy women against most common agents of bacterial vaginosis. It was found that nearly all of the tested Lactobacillus strains exerted distinct antagonistic activity against anaerobic bacteria: Gardnerella vaginalis, Prevotella bivia and Peptostreptococcus anaerobius and quite a number also against Gram-negative rods, while only some of them were able to inhibit Gram-positive aerobic cocci as Enterococcus faecalis or Staphylococcus aureus. PMID- 10865431 TI - [Isolation of toxigenic strains of Clostridium difficile and enterotoxin producing Bacteroides fragilis from fecal specimens of patients suspected of antibiotic associated diarrhoea]. AB - Fifty faecal samples from patients suspected of AAD (antibiotic associated diarrhoea) were studied for Clostridium difficile and enterotoxin producing Bacteroides fragilis (ETBF). Using TCD (Becton-Dickinson) and C. difficile Toxin A test (Oxoid) in 34% of specimens the presence of toxin A was detected. From all specimens 25 C. difficile strains were isolated. All isolated strains produced toxin B in vitro which was shown in Mc Coy cytotoxicity test. Eighteen strains only were toxin A positive in vitro. From all isolated C. difficile strains 28% were tox A (-) tox B (+). By means of PCR presence of toxin A and toxin B genes was tested directly in faecal samples and in strains. From the same 50 faecal samples 17 B. fragilis strains were isolated. Four of them produced the enterotoxin (fragilisin) which was detected on the HT 29/C1 cell line. Genes of fragilisin were found in strains and directly in faecal samples. Toxin producing C. difficile and B. fragilis (ETBF) together were found in 3 samples. From one faecal sample only ETBF was cultured. PMID- 10865432 TI - [Reliability of results evaluation for the passive hemagglutination test in pertussis obtained from WSSE laboratories]. AB - This study was undertaken for assessing of the reliability of the passive haemagglutination test with B. pertussis endotoxin in 18 laboratories of the Sanitary Epidemiological Stations. Each laboratory determined the level of pertussis antibodies in three serum samples twice, at interval of two weeks. The correct results were obtained in 7 laboratories (38.9%). The results of pertussis antibodies determination in only one or two samples were differed more than twice from correct in 5 additional laboratories; in this way the test was carried out satisfactorily in 12 laboratories (66.7%). Reproducibility of the results was good in 12 laboratories (66.7%). The study showed the necessity of repeated interlaboratory controls and periodic training of laboratory workers for raising of the quality and reliability of serological investigations for pertussis. PMID- 10865433 TI - [Bacteriophage types of Salmonella enteritidis occurring in Poland in 1986-1995]. AB - The Lalko phages collection was used to phage type a total of 517 Salmonella Enteritidis strains isolated from food-poisoning outbreaks (312 strains) and other common sources (205 strains) in Poland, during the years 1986-1995. Above 99.0% of all strains tested were recognized as belonging to definitive phage type. Phage types 1, 6 and 7 were predominant. The strains of type 1 and 7 were most numerous. Of the 517 examined strains 312 were isolated from 46 food poisoning outbreaks. Most of them came from the one phage type outbreaks; 8 mixed outbreaks were noted. The greatest number of the food-poisoning outbreaks was caused by Salmonella Enteritidis phage types 1, 6 and 7. Phage type 16 was isolated from persons for the first time. PMID- 10865434 TI - [Occurrence of Escherichia coli O157 in feces of healthy persons, from selected groups, in the country]. AB - The purpose of the study was determination of the occurrence of E. coli O157 in faeces samples of healthy subjects and characterization of the isolated strains with respect to their potential pathogenicity. The study was carried out in two stages. In the first one in 5 sanitary-epidemiological stations samples were tested from healthy subjects after inoculation onto McConkey (MC) or/and McConkey with sorbitol (SMC) media and isolating from each culture 10 lactose-positive (on MC medium) or sorbitol-negative (on SMC) colonies. Then latex test was done with each isolate for E. coli O157 presence. In all, 1005 samples were studied, including 260 taken from children aged 0-2 years, 180 samples from children aged 3-10 years, and 565 samples from older children and adults. E. coli O157 rods were cultured from 6 adults (0.6%). In the second stage carried out at the Laboratory of Enterobacteriaceae, Bacteriology Department, National Institute of Hygiene strains obtained from territorial laboratories were studied determining their phenotypic and genotypic traits regarded as virulence markers of verotoxic E. coli O157 strains, such as inability to ferment sorbitol and MUG breakdown, and production of verotoxins and enterohaemolysin. By the PCR method fragments were sought of genes coding for production of verotoxins, intimin and enterohaemolysin. The results showed that no E. coli O157 strain obtained from healthy individuals produced verotoxins, but three studied strains contained the eae gene determining intimin production and they were regarded as enteropathogenic. PMID- 10865435 TI - [Analysis of bacteriocinogenic properties of Yersinia enterocolitica strains]. AB - The aim of the study was the investigation of bacteriocinogenic properties of 102 Yersinia enterocolitica strains. The influence of selected factors on the production of bacteriocins by Y. enterocolitica and properties of jersiniacin 44JPSBKOH were also investigated. Bacteriocinogenic properties of Y. enterocolitica strains were tested by using the delayed cross-streaking method. It was found that the production of bacteriocins by Y. enterocolitica depended on the type of media on which the producer and indicator strains were grown. It turned out that some strains of Y. enterocolitica showed bacteriocinogenic properties at 25 degrees C, 30 degrees C and 37 degrees C irrespective of the presence of manganese ions in medium. In the presence of iron ions these strains showed bacteriocinogenic properties only at 25 degrees C. Y. enterocolitica strains which required Mn2+ or Mn7+ ions for bacteriocins production showed this activity only at 25 degrees C but in presence of Fe3+ ions they had no bacteriocinogenic properties. The partially purified jersiniacin 44JPSBKOH is a protein, its molecular weight was estimated to be 40 kDa. Yersiniacin 44JPSBKOH was active in the pH range of 3 to 9. Its bactericidal activity was rapidly lost when heated to 100 degrees C and treated with proteolytic enzymes. Yersiniacin 44JPSBKOH showed bactericidal activity against other Y. enterocolitica strains and some strains of Pseudomonas aeruginosa isolated from humans. PMID- 10865436 TI - [IgG and IgA immunoglobulins in helicobacter pylori infections of children with chronic dyspepsia before and after two week triple drug therapy]. AB - The aim of this study was to investigate whether serological techniques, ELISA and Western blot, are useful in monitoring treatment of H. pylori-associated chronic dyspeptic symptoms in children. We observed a correlation between a decrease in the anti-H. pylori IgG titer and an effective treatment. So, our results suggested that the ELISA assay conducted with a glycine H. pylori extract can be a good noninvasive assay for monitoring the effectiveness of the therapy. By using the Western blot method, we showed some variation in the specificity of anti-H. pylori IgG produced before and after treatment. However, this variation was not correlated with the effectiveness of the therapy. PMID- 10865437 TI - [Evaluation of resistance patterns of gram-negative rods from personal clinical materials]. AB - 1862 clinical specimens from neonates with infection risk factors treated in the Department of Neonatology, University of Cracow were examined. The aim of this study was to investigate the participation of clinically important Gram-negative rods in hospital infections and to check the resistance patterns of these pathogens. The strains were identified in automatic ATB system using ID 32E and ID 32GN strips with biochemical tests. The susceptibility of isolates of antibacterial agents was determined in automatic ATB system using ATB G- and ATB PSE strips. 436 strains of Gram-negative rods were cultured. 289 strains (66.3%) belonging to Entero-bacteriaceae family and 147 strains (33.7%) non-fermenting rods were isolated. Among Gram-negative aerobic fermenting rods (mainly K. pneumoniae and E. cloacae), increasing resistance to aminoglycosides and beta lactams, due to new broad spectrum and so called inducible beta-lactamases, was observed. The contribution of non-fermenting rods, especially Pseudomonas spp. and Acinetobacter spp. to the aetiology of infections in hospitalized newborns has increased. Carbapenems and fluoroquinolones are highly active in vitro against the examined strains of multiresistant Gram-negative rods. PMID- 10865438 TI - [Evaluation of the usefulness of commercial enzyme-linked immunosorbent assays for diagnosis of Mycoplasma pneumoniae infections]. AB - The usefulness of the ELISA distributed by BioChem ImmunoSystems, Medial Polska, Biomedica/Virotech and prepared in our laboratory (ELISA FH-K) for diagnosis of the M. pneumoniae infections was estimated. Eighty six serum samples obtained from 86 patients with respiratory tract infections were simultaneously tested by ELISA-IgM/-IgG and by complement fixation test which was accepted as a reference test. The highest sensitivity in relation to the CFT was displayed by the ELISA BioChem ImmunoSystems and Medial Polska (100%), slightly lower sensitivity by the ELISA Biomedica/Virotech--96.5% and ELISA FH-K--90.9% when determining mycoplasmal antibodies of IgM. The lowest sensitivity was displayed by the ELISA Biomedica/Virotech when determining antibodies of the IgG class (54.9%). The specificity of ELISA in relation to the CFT was generally higher when detecting mycoplasmal antibodies of the IgM class then of IgG class. The study demonstrated that all 4 ELISA may be used in routine serodiagnosis of M. pneumoniae infection. For the improve of sensitivity of ELISA it's recommended to measure simultaneously the level of mycoplasmal antibodies of IgM and IgG. PMID- 10865439 TI - [Toxins of Bacteroides fragilis and Bacteroides thetaiotaomicron rods as stimulators of adhesion molecule expression on the surface of vascular endothelial cells]. AB - Lipopolysaccharides from four Bacteroides fragilis strains: one nonenterotoxigenic (NTBF) and three enterotoxigenic (ETBF), and from three B. thetaiotaomicron strains were extracted by hot phenol-water method and purified. B. fragilis enterotoxin was prepared according to the procedure of van Tassell et al. (1992). The influence of the examined toxins on the expression of adhesion molecules: ICAM-1, VCAM-1 and E-selectin on HMEC-1 (human dermal microvascular endothelial cells) was assayed in ELISA test with monoclonal antibodies. Four concentrations of toxins were applied: 0.01, 0.1, 1.0 and 10.0 (micrograms/ml). Endothelial cells were activated for 24 hours (ICAM-1 and VCAM-1 expression) and for 4 hours (E-selectin expression). The coloured product of immunoenzymatic reaction was measured by reading the absorbance at wavelength 492 nm. Two controls were performed in each experiment: with resting HMEC-1 and E. coli O55:B5 LPS (Sigma, USA). Bacteroides fragilis and B. thetaiotaomicron lipopolysaccharides stimulated three adhesion molecules under investigation. Their activity was comparable, but weaker than the activity of E. coli O55:B5 LPS. ICAM-1 was the most stimulated molecule. B. fragilis enterotoxin induced two adhesion molecules: VCAM-1 and E-selectin demonstrating weaker stimulatory activity than E. coli LPS. Stimulation of adhesion molecules on vascular endothelial cells should be considered to be a biological activity of B. fragilis and B. thetaiotaomicron endotoxins and B. fragilis enterotoxin. PMID- 10865440 TI - [Effect of suprainhibitory (supra-MIC) concentration of clindamycin on endotoxin release from Bacteroides fragilis]. AB - The aim of this work was to determine the influence of clindamycin at concentration 100 x MIC on the growth of culture and the induction of endotoxin release from the cells of standard B. fragilis IPL E 323 strain. The antibiotic was added to 24-hour culture of the strain in BHI medium (time 0), its final concentration was 12.5 mg/l. The samples for further examinations were collected at time 0 and after 1, 2, 3 and 24 hours of continued incubation. At the same time the strain was cultured in a medium without the antibiotic. The optical density (OD 420 nm) of each sample was determined, they were centrifuged (2000 x g, 10 min), the supernatants were filtered (0.45 micron filters) and concentrated three times (5000 D ultrafilters). Two serological methods were applied to detect the presence of endotoxin in filtrates of culture medium: immunoelectroprecipitation (IEP) and immunoenzymatic assay (dot-ELISA) with rabbit anti-IPL E 323 immune serum. The results of experiments performed with filtrates of culture without the antibiotic indicate that standard B. fragilis IPL E 323 strain liberates endotoxin spontaneously to the culture medium. Single suprainhibitory dose of clindamycin at concentration 100 x MIC inhibits the growth of examined strain and does not cause augmented release of endotoxin from B. fragilis cells in vitro. PMID- 10865441 TI - [Affinity of exocellular DD-carboxypeptidase/transpeptidase from Saccharopolyspora erythraea PZH TZ-575 to beta-lactam compounds]. AB - The DD-carboxypeptidase/transpeptidases (DD-peptidases) involved in bacterial cell wall metabolism, catalyse the attack of C-terminal D-alanyl-D-alanine peptide bond of the peptydoglycan precursor. These enzymes are inactivated by beta-lactam antibiotics. DD-peptidase from Saccharopolyspora erythraea PZH TZ 64 575 was purified by the use of DEAE-cellulose, Sephadex G-100, Q-Sepharose resins and FPLC (Mono Q). After each step the effluent was concentrated by Amicon ultrafiltration. The purified enzyme showed DD-carboxypeptidase specific activity of 50.9 U/mg. The enzyme exhibited high affinity to beta-lactam compounds e.g. cefamandole, cefapirin, cefradin 1.5-2.6 x 10(-8) M. It was used to screen strains from the Culture Collection of the National Institute of Hygiene in Warsaw for the production of DD-peptidase inhibitors. PMID- 10865442 TI - [Studies on the antiviral activity of silver sulfathiazole]. AB - Silver sulfathiazole shows strong antibacterial activity and good tolerance after topical application. The aim of the study was to determine the antiviral activity of silver sulfathiazole in tissue culture after incubation of drug and virus. The antiviral activity was measured after various periods of exposure and at different drug concentrations. The results obtained indicate the activity of silver sulfathiazole against Herpesvirus type 1 and type 2. This drug suppresses or completely inactivates the infectivity of virus. The antiviral effect is directly related to concentration of the drug and duration of exposure. At concentration of 10 micrograms/ml it has the highest activity after 30 minutes of exposure, however at a concentration of 20 micrograms/ml it induces a similar effect after 10 minutes. Silver sulfathiazole had antiviral activity similar to that of silver nitrate, while sulfathiazole alone was ineffective. PMID- 10865444 TI - Natural inhibitors of tumour necrosis factor-alpha production, secretion and function. AB - Tumour necrosis factor-alpha (TNF), originally discovered by its antitumor activity, is one of the most pleotropic cytokines acting as a host defence factor in immunologic and inflammatory responses. Although the antitumour activity and mediation of inflammation by TNF could be beneficial to the host, unregulated TNF is now known to be the basis for development of various diseases including septic shock, the wasting disease, cachexia, and various inflammatory and/or autoimmune diseases. With an attempt to find potential therapeutic agents for TNF-mediated diseases, research during the last decade has led in the identification of well over one hundred natural inhibitors of either TNF production/secretion or function. This review summarises the structures, mechanism of action and therapeutic potential of these natural products. PMID- 10865443 TI - [Phagocytosis and oxidative metabolism of granulocytes in mice after stem cell factor (SCF) injection]. AB - Stem Cell Factor (SCF) can stimulate the growth and development of primitive multipotential and unipotential hematopoietic stem cells, either alone or in combination with other cytokines such as Granulocyte-Macrophage Colony Stimulating factor (GM-CSF) and Granulocyte-CSF (G-CSF). It was found that these cytokines can also stimulate the function of granulocytes but there is no information concerning SCF influence on the function of these mature cells. SCF was injected into mice subcutaneously during 5 consecutive days in a dose of 1 microgram/kg/d. An examination of the percentage of phagocytic granulocytes and NBT test was performed. The activity of acid phosphatase (AcP), alkaline phosphatase (AP), peroxidase (MPO) and esterase were determined by cytochemistry methods. On the basis of obtained results we can conclude that SCF evidently increases all tested parameters connected with the metabolism of phagocytosis. PMID- 10865445 TI - Suppression of rat neutrophil reactive oxygen species production and adhesion by the diterpenoid lactone andrographolide. AB - The present study was to examine whether andrographolide, a diterpenoid lactone isolated from the anti-inflammatory herbal medicine Andrographis paniculata (Burm. f.) Nees. (Acanthaceae), has the ability to prevent phorbol-12-myristate 13-acetate (PMA)-induced reactive oxygen species (ROS) production, as well as N formyl-methionyl-leucyl-phenylalanine (fMLP)-induced adhesion by rat neutrophils. Results demonstrated that PMA (100 ng/ml) induced rapid accumulation of H2O2 and O2. in neutrophils within 30 minutes. Andrographolide (0.1 to 10 microM) pretreatment (10 min, 37 degrees C) significantly attenuated the accumulation of these two oxygen radical metabolites. Administration of andrographolide also significantly prevented fMLP-induced neutrophil adhesion. These data suggest that preventing ROS production and neutrophils adhesion may confer andrographolide the ability to be an anti-inflammatory drug. PMID- 10865446 TI - Epoxy-acetogenins and other polyketide epoxy derivatives as inhibitors of the mitochondrial respiratory chain complex I. AB - Annonaceous acetogenins (ACG), an extensive group of cytotoxic natural products, are antitumor agents whose main mode of action is inhibition of the mammalian mitochondrial complex I. Herein we describe the importance of the different chemical groups along the alkyl chain for optimal inhibitory potency, discussing the structurally relevant factors present in these compounds. For this purpose, a series of epoxide derivatives from alpha-linolenic acid were prepared and their activity compared with that of epoxy-acetogenins and tetrahydrofuranic (THF) acetogenins isolated from Rollinia membranacea. PMID- 10865447 TI - Effects of some iridoids from plant origin on arachidonic acid metabolism in cellular systems. AB - Seven iridoid glycosides isolated from different extracts of Scrophularia scorodonia L., namely bartsioside, aucubin, harpagide, harpagoside, 8 acetylharpagide, scorodioside and scropolioside B, had been evaluated for their in vitro anti-inflammatory activity in cellular systems generating COX and LOX metabolites. Structure-activity relationships obtained from in vitro screening results were discussed. Most compounds assayed did not exhibit any significant effect on PGE2- and LTC4-release from calcium ionophore-stimulated mouse peritoneal macrophages. In the LTC4-assay, only aucubin showed a significant effect, with an IC50 value of 72 microM. Harpagoside and harpagide also inhibited release of LTC4, but neither effect reached statistical significance. The release of PGE2 by mouse peritoneal macrophages stimulated with calcium ionophore was inhibited by harpagoside and 8-acetylharpagide, but this effect is not statistically significant. However, most iridoids assayed showed a significant effect on TXB2-release from calcium ionophorestimulated human platelets, with inhibition percentages slightly lower than the reference drug ibuprofen. Only harpagide, scorodioside and scropolioside B had no significant effect on TXB2 release. Our results indicate that selective inhibition of the TX-synthase enzyme may be the primary target of action of most of these iridoids, and one of the mechanisms through which they exert their anti-inflammatory effects. PMID- 10865448 TI - Essential moiety for antimutagenic and cytotoxic activity of hederagenin monodesmosides and bisdesmosides isolated from the stem bark of Kalopanax pictus. AB - For the elucidation of the antimutagenic and cytotoxic principles from the stem bark of Kalopanax pictus, seven isolated components of this crude drug were tested in the Ames test and the MTT test. Hederagenin and its monodesmosides, kalopanaxsaponin A and I in addition to its bisdesmosides, kalopanaxsaponin B and H, showed potent antimutagenic activities against aflatoxin B1 (AFB1). However, they had no inhibitory effects on mutagenicity induced by the direct mutagen, N methyl-N'-nitro-N-nitrosoguanidine (MNNG). This suggested that hederagenin glycosides might effectively prevent the metabolic activation of AFB1 or scavenge the electrophilic intermediate capable of inducing mutation. Hederagenin was found to be an essential moiety for the exhibition of antimutagenicity. Moreover, hederagenin and its 3-O-glycosides were found to be cytotoxic on various tumor cell lines, P-388, L-1210, U-937, HL-60, SNU-5 and HepG2, while 3,28-di-O glycosides of hederagenin were not cytotoxic. Hence, hederagenin and its 3-O glycosides could be suitable for cancer treatment chemopreventive drugs. PMID- 10865449 TI - Anti-herpes simplex virus type-1 flavonoids and a new flavanone from the root of Limonium sinense. AB - From the root of Limonium sinense (Girard) Ktze a new (2R,3S)-3,5,7,4' tetrahydroxy-3',5'-dimethoxyflavanone was isolated and named isodihydrosyringetin (3), together with nine other known compounds, (-)-epigallocatechin 3-O-gallate (1), samarangenin B (2), myricetin (4), myricetin 3-O-alpha-rhamnopyranoside (5), quercetin 3-O-alpha-rhamnopyranoside (6), (-)-epigallocatechin (7), gallic acid (8), N-trans-caffeoyltyramine (9), and N-trans-feruloyltyramine (10). All of them were examined for their inhibitory effects on herpes simplex virus type-1 (HSV-1) replication in Vero cells. Both compounds 1 and 2 exhibited potent inhibitory activities in HSV-1 replication. Comparison of the IC50 values indicated that compounds 1 and 2 had higher inhibitory activities than the positive control acyclovir (38.6 +/- 2.6 vs. 55.4 +/- 5.3 microM, P < 0.001; 11.4 +/- 0.9 vs. 55.4 +/- 5.3 microM, P < 0.0005). Cytotoxicity was unlikely involved because no cell deaths were observable in the Vero cells following 5 day treatments with compound 1 or 2. PMID- 10865450 TI - Assessment of antimycobacterial activity of a series of mainly marine derived natural products. AB - A series of mainly marine derived natural products were tested for their activities against Mycobacterium tuberculosis and M. avium. Of the thirty-nine compounds tested fifteen demonstrated minimum inhibition concentrations (MICs) of 32 micrograms/ml or less, and eleven had MICs of 16 micrograms/ml or less. The most active compound found in this study was the sponge derived metabolite axisonitrile-3 (MIC 2 micrograms/ml). PMID- 10865451 TI - Antileishmanial activity of three saponins isolated from ivy, alpha-hederin, beta hederin and hederacolchiside A1, as compared to their action on mammalian cells cultured in vitro. AB - The in vitro antileishmanial activity of three saponins isolated from ivy, alpha hederin, beta-hederin and hederacolchiside A1, was investigated on Leishmania infantum. The assessment of possible targets (membrane integrity, membrane potential, DNA synthesis and protein content) was performed in both Leishmania promastigotes and human monocytes (THP1 cells). Results observed in Leishmania showed that the saponins exhibited a strong antiproliferative activity on all stages of development of the parasite by altering membrane integrity and potential: hederacolchiside A1 appeared to be the most active compound against both promastigotes and amastigotes. Results observed in THP1 cells demonstrated that the saponins exerted also a potent antiproliferative activity against human monocytes, by producing a significant DNA synthesis inhibition. The ratio between antileishmanial activity on amastigotes and toxicity to human cells suggested that the saponins could be considered as possible antileishmanial drugs. PMID- 10865452 TI - Study of the gastrointestinal protective effects of polysaccharides from Angelica sinensis in rats. AB - We studied the protective effects of polysaccharides isolated from the root of Angelica sinensis (Oliv.) (Danggui) on gastrointestinal damage induced by ethanol or indomethacin in rats. Oral administration of ethanol provoked a marked hemorrhagic damage in the glandular mucosa, which was accompanied with a significant increase of myeloperoxidase (MPO) activity, a marker enzyme for inflammation and neutrophil infiltration. An extract from Angelica, which mainly consisted of polysaccharides (95%) (AP), dose-dependently prevented gastric mucosal damage. This ulcer protective effect could last at least 12 h after administration. Prostaglandin E2 produced a similar anti-lesion effect. AP and prostaglandin E2 also reduced mucosal MPO activity. Indomethacin-induced gastrointestinal damage, another neutrophil-dependent lesion model in the gastrointestinal tract, was also prevented by AP pretreatment. The present findings suggest that polysaccharides from Angelica possess an anti-inflammatory action, perhaps through the inhibitory action on neutrophil infiltration in the gastrointestinal mucosa. AP could potentially be useful to prevent any neutrophil dependent mucosal injury in the gastrointestinal tract. PMID- 10865453 TI - Quantitative high performance liquid chromatographic analysis of diterpenoids in agni-casti fructus. AB - Pharmacological data have indicated that part of the dopaminergic activity of Vitex agnus-castus is attributed to the labdan diterpenoids found in the fruits. Therefore an analytical method for the standardization of rotundifuran (1), vitexilactone (2) and 6 beta,7 beta-diacetoxy-13-hydroxy-labda-8,14-diene (3) was developed. Because of the time-consuming and expensive isolation of the diterpenoids, p-cymene was chosen as an internal standard. The concentration of rotundifuran in different extracts and trade samples of the drug varies between 0.04 and 0.30% in the drug and between 1.04 and 2.23% in the extract. The concentration of vitexilactone was generally lower between 0.016 and 0.167% for the drug and between 0.34 and 1.01% for the extract. The determined concentration of 6 beta,7 beta-diacetoxy-13-hydroxy-labda-8,14-diene in the drug was in the range of 0.02 and 0.10% and in the extract in the range of 0.18 and 0.80%. Determination of the factors of correction of p-cymene gave 5.63 for rotundifuran, 2.73 for vitexilactone and 3.74 for 6 beta,7 beta-diacetoxy-13 hydroxy-labda-8,14-diene. PMID- 10865454 TI - Local anaesthetic, antibacterial and antifungal properties of sesquiterpenes from myrrh. AB - We extracted, purified and characterized 8 sesquiterpene fractions from Commyphora molmol. In particular, we focused our attention on a mixture of furanodiene-6-one and methoxyfuranoguaia-9-ene-8-one, which showed antibacterial and antifungal activity against standard pathogenic strains of Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans, with minimum inhibitory concentrations ranging from 0.18 to 2.8 micrograms/ml. These compounds also had local anaesthetic activity, blocking the inward sodium current of excitable mammalian membranes. PMID- 10865455 TI - Anti-allergic and anti-inflammatory triterpenes from the herb of Prunella vulgaris. AB - The activity-guided fractionation of the extract of the herb of Prunella vulgaris (Labiatae) led to the isolation of four triterpenes, i.e., betulinic acid, ursolic acid, 2 alpha,3 alpha-dihydroxyurs-12-en-28-oic acid, and 2 alpha hydroxyursolic acid. One of these compounds, 2 alpha,3 alpha-dihydroxyursolic acid, demonstrated significant inhibition on the release of beta-hexosaminidase from the cultured RBL-2H3 cells in a dose-dependent manner; the IC50 value was calculated to be 57 microM. When the isolated compounds were tested for their effects on the production of nitric oxide from cultured murine macrophages, RAW 264.7 cells, ursolic acid and 2 alpha-hydroxyursolic acid exhibited strong inhibitory activities (IC50 values, 17 and 27 microM, respectively). PMID- 10865456 TI - Antimicrobial activity of 9-O-acyl- and 9-O-benzoyl-substituted berberrubines. AB - In the course of a structure-activity relationship study on berberrubine derivatives, a series of compounds bearing 9-O-acyl-(4-6) and 9-O-benzoyl- (7) substituents was synthesized with the expectation of increasing the antimicrobial activity. One of the berberrubine derivatives, 9-lauroylberberrubine chloride was the most active against Gram-positive bacteria Enterococcus faecalis, Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus, Bacillus subtilis as well as the Gram-negative bacterium Klebsiella pneumoniae in comparison to berberine, the currently used antibiotic in clinic. This result suggested that the presence of lipophilic substituents of certain structures and sizes might be crucial for the optimal antimicrobial activity. PMID- 10865457 TI - Agelasine F from a Philippine Agelas sp. sponge exhibits in vitro antituberculosis activity. AB - Marine sponge samples were collected in Baler, Aurora, Philippines, and extracts were tested for in vitro antituberculosis activity. An orange Agelas sp. sponge yielded the known compound, agelasine F, which inhibited some drug resistant strains of Mycobacterium tuberculosis in vitro at concentrations as low as 3.13 micrograms/ml. Activity against M. tuberculosis residing within macrophages required concentrations of 13-22 micrograms/ml which was below the IC50 for Vero cells (34 micrograms/ml). PMID- 10865458 TI - In vitro effect of essential oils and isolated mono- and sesquiterpenes on Leishmania major and Trypanosoma brucei. AB - The effect of different essential oils as well as of isolated mono- and sesquiterpenes on the viability of bloodstream forms of Trypanosoma brucei, promastigotes of Leishmania major and human HL-60 cells was evaluated using the Almar Blue assay. Of the 12 essential oils and 8 terpenes investigated, only three essential oils, Melissa officinalis (balmmint) oil, Thymus vulgaris (thyme) oil, and Melaleuca alternifolia (tea tree) oil were about 50-fold and 80-fold more toxic to bloodstream forms of T. brucei than to HL-60 cells, respectively. Terpinen-4-ol, the main compound of the Australian tea tree oil, was even 1000 fold more toxic to trypanosomes than to the human cells. On the other hand, none of the essential oils and terpenes tested were more toxic to promastigotes of L. major than to HL-60 cells. PMID- 10865459 TI - Structure and antiprotozoal activity of triterpenoid saponins from Glinus oppositifolius. AB - Two new triterpenoid saponins, glinosides A and B, isolated from the aerial parts of Glinus oppositifolius, have been characterized by 1D, 2D, NMR and high resolution mass spectral (HRMS) techniques. Their structures were established respectively as 16-O-(beta-arabinopyranosyl)-3-oxo-12,16 beta,21 beta,22 tetrahydroxyhopane for glinoside A and 16-O-(beta-arabinopyranosyl)-3-oxo-12,16 beta,22-trihydroxyhopane for glinoside B. Results presented evidence that fractions had a better antiplasmodial activity (IC50 = 31.80 micrograms/ml) than pure glinoside A (IC50 = 42.30 micrograms/ml). PMID- 10865460 TI - Antiplatelet aggregation principles from Ephemerantha lonchophylla. AB - Bioactivity-directed separation led to the identification of four compounds, viz. denbinobin (1), 3,7-dihydroxy-2,4-dimethoxyphenanthrene (2), 3-methylgigantol (3), and erianthridin (4) from the ethanolic extract of Ephemerantha lonchophylla. Antiplatelet tests were carried out using 4 different aggregation inducers, viz. arachidonic acid (AA), thrombin, collagen and platelet activating factor (PAF). The results indicated that only compounds 2, 3, and 4 exhibited generally significant anti-aggregation activities with that against AA-induced aggregation being most effective. Estimated IC50, values in this regard for 2, 3, and 4 were 24 microM, 30 microM and 9 microM, respectively. PMID- 10865461 TI - A new acylated flavonol glycoside and antioxidant effects of Hedyotis diffusa. AB - A study on the bioactive principles of Hedyotis diffusa Willd., led to the isolation of a new acyl flavonol di-glycoside which was characterized as kaempferol 3-O[2"-O-(E-6'"-O-feruloyl)-beta-D-glucopyranosyl]-beta-D-galactop yranoside by spectral and chemical methods from the methanolic extract. In addition, three known flavonol glycosides and six known iridoid glycosides were also obtained. The above-mentioned glycosides were tested for antioxidant effects on xanthine oxidase inhibition, xanthine-xanthine oxidase cytochrome c and TBA MDA systems. PMID- 10865462 TI - Evaluation of the antiviral activity of kaempferol and its glycosides against human cytomegalovirus. AB - The antiviral activity of seven flavonoids, belonging to the kaempferol series, was evaluated against human cytomegalovirus (HCMV) by a rapid method of detection of the immediate-early (IE) antigen, induced by the virus in infected cells. Flavonoids bearing acyl substituents were found to be the most active compounds. PMID- 10865463 TI - Xanthones in cell cultures of Hypericum androsaemum. AB - Cell cultures of Hypericum androsaemum contain an array of prenylated xanthone aglycones and their glucosides, when grown in modified B5 medium in the dark. Derivatives of 1,3,6,7-tetrahydroxyxanthone prevail over compounds with the 1,3,5,6-oxygenation pattern. 1,7-Dihydroxyxanthone was also isolated. Xanthone accumulation parallels cell growth, is repressed by light and strongly influenced by the culture medium used. PMID- 10865464 TI - Flavonoids and phenolics from Limonium sinense. AB - From the aerial part of Limonium sinense (Girard) Ktze, a new flavonol glycoside, myricetin 3-O-(2"-O-p-hydroxybenzoyl)-alpha-rhamnopyranoside has been isolated together with known flavonols, flavonol glycosides, flavonol glycoside gallates, flavones, flavanones, flavan-3-ols and gallic acid. The structural determinations of these compounds were based on spectral analyses. PMID- 10865465 TI - Sipandinolide: a butenolide including a novel type of carbon skeleton from Siparuna andina. AB - From a lipophilic extract of leaves of Siparuna andina (Monimiaceae), which exhibited antiplasmodial activity in vitro, two new compounds have been isolated: sipandinolide (1), a compound with a novel type of carbon skeleton and (-)-cis-3 acetoxy-4',5,7-trihydroxyflavanone (2). Their structures were established by spectroscopic methods; 2 showed moderate antiplasmodial activity whereas 1 was inactive. PMID- 10865466 TI - Two new cyclic peptides from Drymaria diandra. AB - Two new cyclic peptides, drymarins A and B, were isolated from the whole plants of Drymaria diandra B1. Their structures were determined by detailed spectroscopic analysis as cyclo(-Phe1-Pro1-Pro2-Pro3-Phe2-Phe3-Val-Ile-Ala-) and cyclo (-Pro1-Phe-Tyr-Pro2-Gly-Leu-). PMID- 10865467 TI - New pyrrolizidine alkaloids from Heliotropium crassifolium. AB - Heliotropium crassifolium Boiss, (Boraginaceae) from a population of Ilam, western region of Iran was studied for pyrrolizidine alklaoids (PAs). Four alkaloids have been identified: europine 1, europine N-oxide 2 and a new pyrrolizidine alkaloids ilamine 3 and its N-oxide 4, respectively. Their structures were elucidated by IR, 1H-NMR and EIMS data. PMID- 10865468 TI - Terpenoids and flavonoids from Artemisia species. AB - A phytochemical reinvestigation of the aerial parts of Artemisia sieversiana gave a new guaianolide and two known flavones (chrysosplenetin and 5-hydroxy-3',4',6,7 tetramethoxyflavone). Antifungal fractions derived from the chloroform extract of A. annua afforded two cadinane derivatives (arteannuin B and artemisinin), oleanolic acid, beta-sitosterol, stigmasterol, and the four flavones artemetin, bonanzin, eupalitin and chrysosplenetin. Their structures were elucidated by spectral methods. All isolates from the two species were tested in vitro for antifungal activity. Arteannuin B, a main sesquiterpenoid in A. annua, showed antifungal activity against one human (Candida albicans, MIC: 100 micrograms/ml) and four plant pathogenic fungi (Gaeumannomyces graminis var. tritici, Rhizoctonia cerealis, Gerlachia nivalis and Verticillium dahliae, MICs: 150, 100, 150 and 100 micrograms/ml, respectively) whereas others showed no antifungal activity. The MIC value of ketoconazole to C. albicans was 1.0 microgram/ml, and those of triadimefon to G. graminis var. tritici and R. cerealis 150 and 100 micrograms/ml. PMID- 10865469 TI - A new steroidal saponin from the leaves of Agave americana. AB - A new bisdesmosidic spirostanol saponin, along with three known saponins, were isolated from Agave americana (Agavaceae). The structure of the new saponin was elucidated as (25R)-3 beta,6 alpha-dihydroxy-5 alpha-spirostan-12-one 3,6-di-O beta-D-glucopyranoside. Among the isolated saponins, hecogenin tetraglycoside showed cytotoxic activity against HL-60 human promyelocytic leukemia cells with an IC50 value of 4.3 micrograms/mL. PMID- 10865470 TI - A simple and efficient separation of the curcumins, the antiprotozoal constituents of Curcuma longa. AB - A simple and efficient method for the separation of the three phenolic diketones, curcumin 1, demethoxycurcumin 2, and bis-demethoxycurcumin 3, isolated from the rhizomes of Curcuma longa has been developed. The method is of general applicability for the separation of compounds containing acidic and chelating groups and is amenable to large scale separations. The curcumins 1-3 show moderate activity against Plasmodium falciparum (IC50: 3.5, 4.2 and 3.0 micrograms/ml) and Leishmania major (IC50: 7.8, 14.1 and 21.5 micrograms/ml) respectively. PMID- 10865471 TI - [Reproducibility in induced sputum in children with asthma]. AB - In asthmatic children sputum-induction with hypertonic saline is useful to quantify the eosinophilic inflammation. However, only few data are available about feasibility and safety of the procedure in children. Therefore, taking 9 non-atopic healthy control children (mean age 11.8 years) and 34 asthmatic children (mean age 11.4 years), inhaling n = 25 Budesonid (400-1200 micrograms/die) and n = 9 DNCG (60 mg/die), sputum induction was performed twice within 6 weeks. Briefly, 10 minutes after inhalation of 200 micrograms salbutamol subjects inhaled hypertonic saline (3, 4 and 5%) for in all 30 minutes, while all 5 minutes lung function was checked and expectoration of sputum was supported. Adequate sputum plugs were separated from contaminating saliva and processed immediately employing native chamber and cytospin cell count as well as measurement of eosinophilic cationic protein (ECP). Sputum-induction could be performed in 84 out of 86 planed tests (97.7%) without any objective clinical adverse effects. The mean fall in FEV1 was 3.0%, the maximum 11.0%. The reproducibility of eosinophil, neutrophil and lymphocyte differential cell count (5-95%-values Test1: 0.0-4.2%, 0.8-11.4%, and 3.2-35.1%, respectively) was moderate for eosinophils and neutrophils (Intraclass-Correlation-Coefficient (ICC) 0.41) as well as for lymphocytes (ICC = 0.49). For ECP 5-95%-values Test1: 39.8-8000.0 micrograms/l) only a fair reproducibility (ICC = 0.24) was found. The ICC levels for total cell count (ICC = 0.31) and for weight of the sputum plug (ICC = 0.30) were also fair. Based on the procedure presented induced sputum is a feasible and safe method in childhood. The differential sputum cell count of eosinophils, neutrophils and lymphocytes can be recommended as parameters with moderate reproducibility. PMID- 10865472 TI - [Pulmonary complications after hematopoietic stem cell transplantation in children]. PMID- 10865473 TI - [Oxidative pulmonary stress in cytoreductive therapy]. PMID- 10865474 TI - [Blood gas analysis in bronchiolo-alveolar carcinoma]. AB - A 44-year old patient is presented with bronchiolo-alveolar carcinoma which had set multiple metastases into the lung. Basic lung function was normal. It is of interest that only blood gases were borderline. There was a huge discrepancy between the extension of the pulmonary tumour and the impairment of lung function. Hence, blood gas analysis at rest and under exercise is a necessary tool in such cases. PMID- 10865475 TI - [Pneumology rehabilitation 1999. Summary of statements of the American Thoracic Society in Am J Respir Crit Care Med 159 (19991) 1666-1682]. PMID- 10865476 TI - [The evolution of otorhinolaryngology from the 19th to the 21th century]. PMID- 10865477 TI - Belfast Cochlear Implant Centre; the surgical results of the first 100 implants. AB - We assess the surgical results of the first one hundred cochlear implant operations at the Belfast Cochlear Implant Centre. This paper is in the form of a retrospective case series, taking into account the surgical technique used and any problems encountered. Initially 28 operations were performed using the standard inverted U-shaped incision. After Gibson and Harrison described a vertical postaural incision we began using a slight modification of this technique for any further implantations. Since November 1994 the vertical post aural skin incision has been used in 71 patients. The surgical complications of cochlear implantation include haematoma, wound infection and flap necrosis of varying degrees of severity. We report a low level of these complications, particularly with the vertical incision. PMID- 10865478 TI - The transmastoid partial labyrinthectomy approach to medial skull base lesions. AB - INTRODUCTION: It has long been thought that surgical disruption of the membranous labyrinth invariably results in sensorineural hearing loss and balance dysfunction. Recent evidence suggests that the inner ear can withstand such manipulation without loss of function. The technique of transmastoid partial labyrinthectomy has recently been described as a means of providing access to lesions of the medial skull base by removing part of the labyrinth and at the same time attempting to preserve hearing and vestibular function of the lateral semicircular canal (LSCC) and otolithic organs. PROCEDURE: An extended cortical mastoidectomy is performed and the posterior and middle cranial fossa dura are exposed widely. The posterior and superior semicircular canals are occluded at their ampullated ends and at the crus commune, and then resected. The LSCC and vestibule are left undisturbed. The petrous apex is removed and the medial end of the internal auditory canal is exposed. Posterior cranial fossa dural flaps are raised allowing access to the brainstem, petro-clival area and cerebellopontine angle. Temporal and suboccipital craniotomies can be performed, as required. RESULTS: Four patients underwent this procedure by a joint Otolaryngological Neurosurgical team for access to the following lesions: three intra-axial pontine cavernomas and a basilar artery aneurysm. The preliminary hearing and balance results are discussed. CONCLUSIONS: The partial labyrinthectomy approach provides improved access to certain lesions of the medial skull base and requires less brain retraction compared with the retrolabyrinthine approach. It also has the potential to preserve serviceable hearing. PMID- 10865479 TI - The natural history of untreated vestibular schwannomas. Is there a role for conservative management? AB - OBJECTIVE: The aim of this study was to investigate the natural history and outcome following the conservative management of a group of patients with unilateral vestibular schwannomas. METHODS: 72 patients with a radiological diagnosis of unilateral vestibular schwannoma were managed conservatively because of poor general health, advanced age, patient preference, small tumour size, minimal symptoms, or tumour in the only/better hearing ear. All patients underwent serial magnetic resonance imaging for assessment of tumour growth, according to American Academy of Otolaryngology-Head & Neck Surgery guidelines (1995). The mean duration of follow-up was 37.8 months (range 12-194 months). Patients were deemed to have failed conservative management if there was evidence of continuous or rapid radiological tumour growth, and/or increasing symptoms or signs. RESULTS: The mean tumour growth rate was 1.16 mm/year (range -0.75 to 9.65 mm/year). Approximately 83% of tumours grew at less than 2 mm/year. Significant tumour growth (total growth > 1 mm) was seen in 36.4%, no or insignificant growth (0-1 mm) in 50%, and negative growth (< 0 mm) in 13.6% of tumours. The growth rate of cerebellopontine angle (CPA) tumours (1.4 mm/year) was significantly greater than that of tumours limited to the internal auditory canal (IAC) (0.2 mm/year) (p = 0.001). Failure of conservative management, in which active treatment was required, occurred in 15.3%. The outcome of these patients appeared to be as favourable as those who underwent primary treatment, without a period of conservative management. The growth rate of tumours in patients who failed conservative management (4.2 mm/year) was significantly greater than that in patients who did not fail (0.5 mm/year) (p < 0.01). No factors predictive of tumour growth were identified. Deterioration of mean pure tone average (0.5, 1, 2, 3 kHz) and speech discrimination scores occurred regardless of whether radiological tumour growth was demonstrated or not. CONCLUSIONS: The majority of vestibular schwannomas are slow growing, although, CPA tumours appear to grow faster than IAC tumours. Deterioration of auditory function occurs even in the absence of tumour growth. Although most Otolaryngologists and Neurosurgeons would agree that the treatment of choice for the majority of vestibular schwannomas is microsurgery, there remains a small group of patients in whom a conservative management approach may be a desirable alternative. PMID- 10865480 TI - The effectiveness of topical treatment in discharging ears with in-dwelling ventilation tubes. AB - The efficacy and tolerability of topical treatment (both drops and spray) in the treatment of otorrhoea in those ears with ventilation tubes in-situ was studied. Sixty patients were randomised into two treatment groups. One group used an antibacterial/anti-inflammatory ear drop preparation (Otosporin) and the other group a similar preparation administered as an ear spray (Otomize). A 'blind' clinical assessment was made one and three weeks after commencing treatment. Both treatments (drops and spray) significantly improved patients' symptoms and signs after one week (p < 0.001). The spray was significantly easier to administer (p < 0.01) and caused less discomfort on application (p < 0.02). PMID- 10865481 TI - Paediatric airway endoscopy. AB - OBJECTIVES: To review all paediatric endoscopies performed in a tertiary referral unit over a three-year period. METHODS OF STUDY: Retrospective analysis of case notes of all paediatric endoscopies performed between May 1993 and June 1996. RESULTS: 333 paediatric airway endoscopies were performed on 146 children, of which 52% were GP referrals and the remainder secondary referrals. 70% were diagnostic endoscopies, 30% therapeutic procedures, with the commonest indication being stridor and respiratory distress (82%). Routine chest radiographs, lateral neck X-rays, and barium swallows were unhelpful in the management of the commoner upper-airway conditions. The commonest findings were laryngomalacia (44%) and subglottic stenosis (22%) and 17% of all cases had multiple airway abnormalities. Tracheotomy was performed on 18.4%, laryngotracheoplasty on 7.5%, and laryngotracheal reconstruction on 2.5%. There were no major complications in this series. CONCLUSIONS: All children with airway symptoms should have a thorough rigid-endoscopic evaluation of their upper and lower airways. Radiology has a limited role in the diagnosis of the more common airway pathologies. These patients need to be assessed and managed in regional centres. PMID- 10865482 TI - Pharyngeal pouch surgery: a five year review. AB - The treatment of pharyngeal pouch varies widely. Our aim was to establish current and recent practice in pharyngeal pouch surgery in our department and set guidelines for future management. A retrospective audit over a 5-year period was performed with all data derived from patient notes. 28 procedures were performed on 24 patients with a mean age of 72 years. Over two thirds of these patients (68%) underwent an endoscopic procedure (stapling or diathermy) and the remainder underwent excision (14%), inversion (10%), cricopharyngeal myotomy (4%) or dilatation (4%). The primary diagnostic investigation performed was a barium swallow in 17 cases, but in 7 cases, referred by gastroenterologists, an oesophagogastroscopy was performed despite characteristic presenting features in all cases. The average inpatient stay was similar for endoscopic and excision procedures (5.5 and 5 days respectively), but longer for inversion procedures (9 days). This was influenced mainly by operative complications. 2 endoscopic stapling procedures were complicated by perforations and 1 patient developed hoarseness after an inversion procedure. The mean follow up time was one month at which stage all asymptomatic patients were discharged. 2 patients treated by endoscopic stapling and 1 patient treated by inversion complained of persistent symptoms and required further surgery. We conclude that endoscopic stapling was the commonest procedure used. Concerning future management, the use of nasogastric tubes after uncomplicated stapling procedures was abandoned. Also it was felt that large pouches should be treated by excision, small pouches by cricopharyngeal myotomy and the remainder by endoscopic stapling. The long-term evaluation of results was also deemed necessary. PMID- 10865483 TI - Vertical extended hemi crico-laryngectomy and reconstruction with a prefabricated tracheal free flap--initial results. AB - We describe our experiences of treating three patients with recurrent T3 squamous cell carcinoma of the larynx, following initial treatment with radiotherapy; using the technique of partial crico-laryngectomy and autologous pre-fabricated tracheal flap reconstruction as described in 1998 by Delaere (1). We have found the technique to be technically challenging. The patients require extensive speech and swallowing rehabilitation following surgery, but the functional result offers significant advantages over the other surgical salvage procedures of total or near-total laryngectomy. We describe our early results of three patients that we have treated using this technique. PMID- 10865485 TI - Congenital bilateral choanal atresia: a novel stenting technique in neonates. AB - A new stenting technique is described which may reduce the incidence of restenosis in choanal atresia after decannulation. Eight neonatal cases with bilateral congenital bony choanal atresia were treated by endonasal perforation. A modified stenting technique utilising endotracheal tubes provided good fixation and stability. This aided epithelialisation of the newly formed tract. Furthermore the stents projected beyond the atretic plate preventing granulation tissue growth over the choanae, thus preventing restenosis. The stents were left in situ for six to eight weeks. The patients were followed up for periods ranging from eighteen months to eleven years. Seven did not require further treatment, and one required unilateral dilatation. Meticulous care in fixing and securing a well designed stent for an appropriate time may be a factor in reducing the incidence of restenosis. PMID- 10865484 TI - Pure sensorineural hearing loss and otosclerosis. An imaging case report. AB - Pure sensorineural hearing loss is not a rare finding in otological practice. Numerous aetiologies could be at the origin of such a deficit. However, otosclerosis is very rarely cited as a cause of pure sensorineural hearing loss. We present one such case of pure sensorineural hearing loss linked to otosclerosis in a 30-year old caucasian male and underline the high contribution of computed tomography to confirm such a diagnosis. Pure sensorineural hearing loss due to otosclerosis is a rare event and can be misdiagnosed. The clinical diagnosis of such a disease may be difficult. In these cases, CT-Scan is the exam of choice to confirm the diagnosis. PMID- 10865486 TI - Endoscopic dacryocystorhinostomy: anatomical approach. AB - Dacryocystorhinostomy is the surgical treatment for nasolacrimal blockage. In recent years, the endoscopic approach has become more popular due to the development of nasal endoscopes and the ease of surgery in comparison to the external approach. In order to identify the lacrimal duct during surgery, surgeons insert a light pipe into the lacrimal duct and then drill or chisel the hard bone of the frontal process of the maxilla to remove the bony covering of the sac and duct. It is obvious that knowledge of the anatomy of the lacrimal sac/duct within the nose is essential for the surgeon. The lacrimal apparatus in the nose was studied using 10 cadaveric half-heads (5 males and 5 females) to establish the anatomical landmarks and most accessible part of the lacrimal duct from within the nose. Although there was solid bone covering the whole length of the sac and the duct, the posteromedial aspect of the lower sac and upper duct was covered by the ultra thin lacrimal bone (average thickness 0.057 mm) which was consistently found to be lying immediately anterior to the uncinate process in the middle meatus, thus constituting a "surgical window" (average size 2.5 mm x 7.2 mm) whereby surgical entry into the lacrimal duct becomes relatively easy. The lower part of the lacrimal sac and the upper part of the lacrimal duct can therefore be easily accessed from within the nose by following this anatomical approach, thus avoiding the need to drill or chisel the dense frontal process of the maxilla. PMID- 10865487 TI - A new tongue plate for use in oropharyngeal KTP laser surgery. AB - The KTP laser is used in both uvulopalatopharyngoplasty and tonsillectomy. However the need to use laser guarded endotracheal tubes represents a sizeable expense to the procedure. The authors describe a modified tongue plate to the oropharyngeal gag that covers all of the endo-tracheal tube, thus enabling the safe use of a non-laser guarded, PVC endotracheal tube. In over a hundred such procedures there has been no laser-related complications. The authors consider that the one off cost of this tongue plate and gag allows a more cost-effective method for performing laser-assisted uvulopalatopharyngoplasty and tonsillectomy. PMID- 10865488 TI - Long-term results of KTP/532 laser uvulopalatopharyngoplasty. AB - The symptom of snoring is no longer one of humorous content and with the ever increasing awareness of its detriments, it has become even more important to find a treatment that would be immensely beneficial to the patient. We would like to present our experiences with the use of the KTP/532 Laser in performing the UP3. This study is significant as it presents a long term four year follow up of cases based on the patients' assessments. The technique itself is tailor made to suit the individual patient, ensuring optimal results which revealed that most patients i.e.; 84% were extremely pleased with the operation. The series also showed minimal morbidity and no intra or post-operative complications which reiterates the need for meticulous, atraumatic technique and judicious, discriminate selection of patients. PMID- 10865489 TI - [Coronary disease and hyperglycemia: and if the assumption of guilt was already evidence?]. PMID- 10865490 TI - [History of the treatment of acute leukemia in children]. PMID- 10865491 TI - [Iron metabolism]. AB - Iron is essential to the human organism but is also highly toxic. Therefore, in the organism, iron is always tightly bound to specific proteins. In the past few years, our understanding of iron cellular uptake and regulation mechanisms has been expanded. However, iron metabolism, particularly its intestinal absorption and release from ferritin, is still uncompletely understood. Intestinal absorption appears to be the prime factor of iron homeostasis control. Iron deficiency is one of the most common nutritional problems in the world. However, oxidative stress induced by iron overload has been implicated in a growing number of diseases. PMID- 10865492 TI - [Assessment of iron status]. AB - Biological tests useful for the assessment of iron status depend on the metabolic compartment to be explored. The transport compartment is explored by the measurement of serum iron, transferrin and its saturation; these tests still play a major role in the screening of iron overload. Assessment of stores is performed by the assay of serum ferritin either for diagnosing depletion or assessing overload. Falsely normal or increased results (inflammation, infection, cancer) can hide a true deficiency. Erythrocyte ferritin and glycosylated ferritin can be useful tools for the interpretation of hyperferritinemia. Soluble transferrin receptor is a specific marker of functional deficiency, not influenced by confounding pathologies. PMID- 10865493 TI - [Epidemiology of iron deficiency]. AB - During the last years, a large number of surveys have suggested than some fractions of the French population, as in other industrialized countries, presented dietary iron intake below the recommended dietary allowances. Iron deficiency is widespread in children, menstruating women, and particularly pregnant women. Iron deficiency is enough severe to be responsible of anemia in some groups of population. Aside the classical risk of anemia (with its well known consequences on health), the effects of iron deficiency by itself, taking into consideration the role of iron in numerous metabolic functions, needs to be more documented to assess the potential deleterious consequences upon health. PMID- 10865494 TI - [Iron and inflammation]. AB - Iron is essential for life. Inflammatory disorders cause iron metabolism disturbances resulting in very low concentration of free iron. Iron binding proteins synthesis have a central role in the decreasing iron reutilization for erythropoiesis, and in the increasing iron storage. Inflammatory disorders also induce inhibition of the erythroid progenitors. Synthesis and action of erythropoietin are also disturbed. All these changes are mediated by the activated cytokine cascade (TNF alpha, IL1, IL6...) that is able to induce anaemia of inflammation those clinical and biological characteristics are well defined. Soluble transferrin receptor concentration can help to identify iron deficiency when inflammation is associated. Management of anaemia of inflammation requires prior to treat the cause of inflammation when it is possible, even if human recombinant erythropoietin has been demonstrated to be effective. PMID- 10865495 TI - [Diagnosis and treatment of iron deficiency]. AB - The investigation of iron deficiency should be oriented by clinical background. In group at risk (infants and children, women of childbearing age, pregnant women) management is limited to nutritional inquiry and gynaecologic examination and oral iron treatment. In men and post menopausal women iron deficiency is assumed to be the result of occult gastro-intestinal blood loss; so, in these patients, upper and lower gastro-intestinal endoscopy are required. Benign lesions are more frequently found in upper digestive tract than lower digestive tract. When these investigations are negative and iron treatment unsuccessful, enteroscopy is recommended. Oral iron treatment is performed with ferrous salts (200 mg/d). Duration of treatment depends on severity of iron deficiency: three months for iron stores deficiency an iron deficient erythropoisis, six months for iron deficiency anaemia. PMID- 10865496 TI - [Genetics and physiopathology of hemochromatosis]. AB - Thanks to the discovery of the HFE gene and of its mutations, it is now established that the most frequent form of hemochromatosis is related to homozygosity for the mutation C282Y, and that other types of hemochromatosis, unrelated to HFE mutations, do exist such as the juvenile hemochromatosis. From a pathophysiological standpoint, the C282Y mutation impairs HFE protein expression at the surface of the membrane and disturbs the cellular entry of iron (carried by circulating transferrin) into the cryptic duodenal cell. This, in turn, is likely to lead to an aberrant programmation of the degree of iron influx from the digestive lumen into the apical duodenal cells. The resulting hyperabsorption, which forms the basis of iron overload in hemochromatosis, is likely to implicate an overexpression of the transmembrane iron carrier DMT1. It is remarkable to observe that these major improvements in the knowledge of hemochomatosis have been accompanied by similar improvements in the understanding of normal iron metabolism. PMID- 10865497 TI - [Diagnosis and treatment of genetic hemochromatosis]. AB - Genetic hemochromatosis is an autosomal recessive disease, characterized by an increased iron absorption, leading to progressive iron overload. The fully expressed phenotype comprises fatigue, skin pigmentation, liver disease with hepatomegaly, cirrhosis and hepatocellular carcinoma, and diabetes. Arthralgias are frequent, cardiopathy or impotence may occur. This presentation is now unfrequent with earlier diagnosis, and patients are often asymptomatic--with only biochemical expression--or pauci-symptomatic (mild fatigue, arthralgias or increased transaminases). Transferrin saturation is always increased. Serum ferritin is proportional to iron burden. Diagnosis is now easy, since most patients are homozygote for the C282Y mutation of the HFE gene. Liver biopsy can be useful to quantify iron overload and assess liver fibrosis. The disease can be lethal due to liver disease, carcinoma or heart disease, but life expectancy goes to normal if patients are treated before the occurrence of cirrhosis. Treatment relies on regular venesections. Familial screening is essential. PMID- 10865498 TI - [Secondary iron overload]. AB - Secondary iron overload (SIO) constitutes a growing clinical problem, particularly in haematological diseases in which the improvements of life expectancy give the iron overload enough duration to play its own prognostic role. Iron may accumulate by two ways: transfusion and/(or) digestive hyperabsorption which is proportional to erythroid plasma iron turnover. To properly evaluate the iron overloading one must be able to appreciate the cumulative red blood cell transfusion volumes. That is to say: weighting and counting red blood cell units. The magnitude of red blood cell precursor mass might be conveniently but indirectly evaluated by the measurement of the plasma transferrin receptor concentration. The group of haematological diseases, complicated by SIO to the contrary of primary haemochromatosis, is very heterogeneous. Some of them like hereditary dyserythropoiesis may not be obviously detectable on standard haematological observation. They can combine or not with hereditary haemochromatosis. A SIO must be treated when it may add a specific prognostic effect. In some cases, regular blood letting are usable without major problems. In all other cases iron chelation therapy is an effective way to reduce SIO, provided long term compliance is obtained. PMID- 10865499 TI - [Quality assessment: the physician's side]. PMID- 10865500 TI - [Hypereosinophilia. Diagnostic approach]. PMID- 10865501 TI - [HIV infection. Epidemiology, screening, prevention, main immunologic anomalies, biological prognosis markers, classification (stages)]. PMID- 10865503 TI - [Ulceration or erosion of the oral and genital mucosa. Diagnostic approach]. PMID- 10865502 TI - [Turner and Klinefelter syndromes. Diagnosis]. PMID- 10865504 TI - [Acute otitis. Etiology, diagnosis, treatment]. PMID- 10865505 TI - [Fetomaternal blood immunization. Screening, prevention]. PMID- 10865506 TI - [Diabetic retinopathy. Physiopathology, diagnosis, clinical course, principles of treatment]. PMID- 10865507 TI - [Thoracic skin basalioma of the pigmented type with superficial, flat ulceration]. PMID- 10865509 TI - [Medical economy--a new challenge for physicians]. AB - What is medical economics? It stands on three important columns: (1) economic analysis (2) policy and decision analysis and (3) outcomes research. Economic evaluations consist always of two components: resource utilization and outcomes. Cost-benefit-analysis requires that the outcome is expressed in monetary terms, in cost-effectiveness-analysis outcomes are expressed in non-monetary terms and cost-utility analysis requires QUALY's (quality adjusted life years) as output. Why do we need medical economics? On one hand we wish to allocate the restricted means as fairly as possible. On the other hand, we have new but more expensive methods and preventive measures. Medical economics can contribute to this situation: through a systematic assessment of the efficiency of the different methods. The potential of cost savings and arguments for new and more expensive methods can be developed, if they are advantageous in the context of the prevention of disease progression or sequelae. The aim of the department is to promote research, to develop the basis for decision-makers, to give lectures and seminars, to support other researchers in the area of medical economics and to take up and cultivate contacts with different actors of health care systems. PMID- 10865508 TI - [New nonsteroidal antirheumatic drugs]. PMID- 10865510 TI - [Medical travel counseling and vaccinations]. AB - The art of travel advice is based on clear and regularly updated information. It aims at covering the specific needs of individual travellers and benefits from the experience of the professional advisor. Frequently occurring and life threatening health problems are deal with in the first place. Recommendations on prophylactic measures are a challenge for the advisor as they require the active collaboration of the traveller. Vaccinations on the other hand require less compliance of the traveller. Routine vaccinations against hepatitis A and B, diphtheria and tetanus, and against measles are discussed in line with compulsory vaccinations for certain countries (yellow fever, cholera, meningitis A + C) and with vaccinations against special risks. PMID- 10865511 TI - [Pitfalls in diagnosis of polymyalgia rheumatica/temporal arteritis]. AB - Polymyalgia rheumatica as well as Horton's disease (arteriitis temporalis) are often diagnosed by exclusion. When either disease is suspected because of certain findings a therapy with glucocorticoids is usually started and initial therapeutic success is commonly seen as indirect proof of diagnosis. Clinical experience, however, shows that both diagnoses may conceal a malignancy. Based on this experience the cases of three patients are reported who's clinical and laboratory findings suggested first manifestations of polymyalgia rheumatica or Horton's disease respectively. During treatment, however, the true underlying disease was revealed in two of the cases. Only in one patient was the diagnosis polymyalgia correct, in the second patient a bronchial carcinoma was found, in the third a carcinoma of the breast with bone marrow carcinosis. All three patients have been presented to physicians participating in a meeting of general medicine in 1999. Before disclosure of the final diagnoses assumptions of the physicians about the initial diagnosis were analysed. The same procedure was used with last year medical students. Physicians had more correct diagnoses (19% had all three correct) than students (only 2.8%). This shows on one hand a marked general diagnostic insecurity concerning the diagnoses polymyalgia and Horton's disease on the other hand it demonstrates the advantage of medical experience gained during practice. PMID- 10865512 TI - [Managing the alcoholic patient in family practice]. PMID- 10865513 TI - [Breast imaging after augmentation with homologous adipose tissue implant]. AB - The implantation of cadaveric fat for breast augmentation, which has been performed up to the Seventies, is nowadays considered obsolete. Since complications frequently arise many years later, we are, however, occasionally confronted with the problematic consequences of the technique. The present case report describes the clinical, mammographic and for the first time to our knowledge MR-mammographic findings of a mammoplasty with cadaveric fat in a 57 year-old patient with a history of breast cancer. PMID- 10865515 TI - Whole-diet approach extends lives. PMID- 10865514 TI - [The problem of autonomy of natural sciences during the time of Galilei]. PMID- 10865516 TI - Getting a grip on GERD. PMID- 10865517 TI - Antidepressants: choices and controversy. PMID- 10865518 TI - Fiber helps diabetes... PMID- 10865520 TI - Sleep-disordered breathing linked to hypertension. PMID- 10865519 TI - Fiber helps diabetes ... but not colon cancer? PMID- 10865521 TI - New weight loss product. PMID- 10865522 TI - Does asthma treatment weaken bones? PMID- 10865523 TI - Treatments for Parkinson's disease. PMID- 10865524 TI - Topical relief for mild to moderate psoriasis. PMID- 10865525 TI - More relaxed ... with fewer drugs? PMID- 10865526 TI - What is the Valsalva maneuver? PMID- 10865527 TI - What is the latest thinking about the environmental causes of sarcoidosis? PMID- 10865528 TI - Differentiation and diagnosis of jaundice. AB - Bilirubin metabolism is a complex and fascinating example of the body's ability to discard, renew, and recycle vital elements. Jaundice is the warning sign for derangements in this system. As is true of pain, jaundice is a powerful impetus for visiting a healthcare provider. Usually associated with hepatitis by a nonclinician, the origins of jaundice can range from benign to fatally malignant. Patients may have any number of idiopathic or nosocomial conditions that can contribute to an icteric state. This review delineates the steps of bilirubin metabolism, enumerates the sources of bilirubin derangement, and examines elements of patient condition and therapeutics that can contribute to hyperbilirubinemia and jaundice. PMID- 10865529 TI - Bile duct injuries in laparoscopic cholecystectomy: nursing perspective. AB - The advent of laparoscopic cholecystectomy brought a new approach with many advantages to patients for the treatment of symptomatic gallstones. What was not anticipated was an increase in bile duct injuries as a major complication of this new technique. The advanced practice nurse (APN) must understand the technical aspects of a laparoscopic cholecystectomy and routine perioperative care, because this knowledge provides a basis to recognize signs and symptoms of potential postoperative problems characteristic of a bile duct injury. The APN may be the initial healthcare provider who examines a patient or who is contacted by the patient with reports of symptoms that must be heeded and acted on. The APN responds to the patient by providing symptom management, initiating and coordinating care, and alerting the healthcare team. The APN at a specialty center may also be the consistent member of the healthcare team caring for a patient through nonoperative management and operative repair of the bile duct injury. Knowledge of the spectrum of bile duct injuries, their various manifestations, diagnostic tests, nonoperative management, and complex surgical repair is necessary for the APN to educate patients, coordinate care along the continuum, and support a patient with a bile duct injury. The medicolegal aspect of bile duct injury during laparoscopic cholecystectomy is also a topic the APN may encounter that requires thoughtful consideration and response. The recognition of subtle postoperative symptoms of a laparoscopic cholecystectomy may mean the difference between early diagnosis of and intervention in a minor injury or a major complication. PMID- 10865530 TI - Hepatitis C: role of the advanced practice nurse. AB - Hepatitis C virus (HCV) is a common but insidious and indolent viral infection that can lead to chronic hepatitis, cirrhosis, and hepatocellular carcinoma. This article provides the advanced practice nurse with current information on prevalence, incidence, spread, and clinical course of hepatitis C. It presents a discussion of the methods of diagnosis, treatment, and management of affected patients. To date, the diagnosis of hepatitis C in the United States has been serendipitous because no surveillance and screening programs have been established. It has been estimated that approximately 4 million persons in the United States are infected with HCV, with only 30% presently diagnosed. Patients with hepatitis C must make informed choices regarding their care and treatment. As more people are diagnosed with hepatitis C, the advanced practice nurse is at the forefront of providing information about spread and diagnosis, treatment options available, and potential side effects of antiviral therapy. The decision to treat chronic HCV must be made in collaboration with other medical experts in hepatology and antiviral therapy, and it must be made with knowledge and understanding of all facets of the disease process and adverse effects of therapy. PMID- 10865531 TI - Necrotizing pancreatitis: pathophysiology, diagnosis, and acute care management. AB - Severe acute necrotizing pancreatitis is a disease that is caused by premature activation of pancreatic enzymes. Cytokine release contributes to systemic manifestations such as systemic inflammatory response syndrome (SIRS), multiple organ dysfunction syndrome (MODS), adult respiratory distress syndrome (ARDS), and sepsis. Diagnosis is based on a history of abdominal pain, laboratory values such as serum amylase and lipase levels, and CT scan. Medical management focuses on fluid and electrolyte balance, antibiotic therapy, pain control, and decreasing systemic complications. Surgery is indicated when infectious pancreatic necrosis has been identified. This article addresses incidence and etiology; pathophysiology; clinical manifestations; diagnostics; and medical and surgical patient care management. PMID- 10865532 TI - Intestinal obstruction. AB - Intestinal obstruction, a disorder that afflicts both children and adults, is associated with significant morbidity. Early recognition and appropriate management can prevent life-threatening complications; however, considerable controversies exist regarding the ideal method of diagnosis and treatment. This article provides a comprehensive overview of risk factors for small and large bowel obstruction. The pathophysiologic process is related to the clinical findings. A summary of diagnostic methods with an evaluation of their relative merit is presented. Preventive, and medical, surgical, and palliative therapies are described, with an emphasis on the actions that an advanced practice nurse can implement. PMID- 10865533 TI - Toxic megacolon: diagnosis and treatment challenges. AB - In adults, toxic megacolon is a relatively uncommon but potentially lethal complication of inflammatory bowel disease (IBD), infectious colitis, or ischemic colitis caused by cancer chemotherapeutic agents. Patients have distension of the colon and signs of toxicity such as elevated temperature, hypotension, decreased level of consciousness and electrolyte imbalances. Factors thought to increase the risk include premature discontinuation of IBD medications; procedures that increase colon trauma, such as barium enema and colonoscopy; medications that decrease gastrointestinal motility; and electrolyte imbalances, especially hypokalemia. Differential diagnosis is made based on the patient's history and results of stool cultures and assay for Clostridium difficile toxin. Medical management in the intensive care unit includes careful monitoring, fluid volume and electrolyte replacement, bowel rest and decompression, antibiotic therapy, and cessation of medications that slow gastric motility. Surgical management may be necessary if there are signs of deterioration, perforation, hemorrhage, or sepsis. PMID- 10865534 TI - Gastrointestinal manifestations of graft-versus-host disease: diagnosis and management. AB - The number of bone marrow transplantations performed to treat diseases such as leukemia, lymphoma, and aplastic anemia has been increasing during the past 2 decades. Improvements in histocompatibility testing, transfusion support, conditioning regimens, and antibiotics have dramatically improved survival after transplantation. As more patients survive the acute phase of bone marrow transplantation and leave major medical centers for their homes, healthcare providers across the country are caring for these patients and their transplantation-associated complications. Graft-versus-host disease (GVHD) of the skin, liver, and gastrointestinal tract are complications that patients may experience months after transplantation. As clinicians, advanced practice nurses encounter these patients in clinics, hospitals, and intensive care units, whether it be for specific GVHD-associated problems or other health conditions. This article reviews the current standards of care for the prevention, diagnosis, and treatment of GVHD of the gastrointestinal tract. PMID- 10865535 TI - Distance education programs for advanced practice nurses: questions to ask. AB - This article reviews the use of distance learning in nursing education and to summarize key questions that must be addressed by programs or students considering advanced practice nursing education using distance technology. An acute care nurse practitioner program using distance learning strategies is provided as an example to illustrate delivery of a clinically based curriculum. Examples of questions to be addressed in evaluating a distance education program include: How much of the course or graduate nursing program is available on-line? What are the specific informational technologies used? How does communication occur between graduate students and faculty? How are clinical requirements of a course managed? Are there any requirements for time to be spent directly on campus? Is it necessary for the student to have a computer and Internet provider? Knowledge of the available technology and components of distance education can enhance the ability of the advanced practice nurse to evaluate better and choose educational program offerings. PMID- 10865536 TI - Longitudinal survey of acute care nurse practitioner practice: year 1. AB - This article reports on the results of an ongoing survey among acute care nurse practitioners (ACNPs) as part of a 5-year longitudinal study in progress exploring role aspects and issues in practice of this new practitioner in advanced practice nursing. The role of the ACNP is progressing, and responses from 619 ACNPs 1 year after they sought national certification reveal continued role development and role challenges, along with increased independence, autonomy, and confidence in practice. Implications for ACNP educational programs and suggestions for those contemplating pursuing an ACNP position as reported by study participants are also discussed. PMID- 10865537 TI - Collaborating for breast health education and research. A university, industry, and community agency partnership. AB - Initiating a collaborative health education program about breast health required talent, expertise, and workload contributions from all involved including university researchers, a regional breast screening agency, and local industries. The credibility and opinions of liaisons or key informants were valued highly, and their support was critical to the success of the project. Participation in any collaborative project is predicated on benefits perceived by each of the partners. The community agency reaped the benefits of greater dissemination of their educational materials through the interventions. The project increased corporate and union awareness of the resources of this agency and in this community. Throughout the project, meetings and telephone conferences were held on a weekly or biweekly basis with the liaisons. Liaisons disseminated updates to management and union representatives. PMID- 10865538 TI - Health conception and health promotion in blue collar workers. Program planning issues. AB - Health conception and health promotion behaviors were measured in 160 blue collar workers ages 18 to 65, focusing on factors influencing blue collar workers' participation in health promotion programs. Results of Laffrey's Health Conception Scale (LHCS) and Pender's Health Promoting Lifestyle Profile (HPLP) indicated: Health responsibility and interpersonal support were significantly greater for women than for men. Exercise was significantly greater for younger workers than for older workers. Nutrition was significantly greater for older workers than for younger workers. Health conception was significantly greater for younger women and for older men. A significant relationship exists between health conception and health promoting lifestyle. Role and self actualization were significantly greater for older workers than for younger workers. Results suggest gender, age, and concept of health are important when planning health promotion programs at an industrial site. PMID- 10865539 TI - Latex allergy in health care workers. What are the risks? AB - Individuals with a history of atopy are at increased risk of becoming latex allergic. Specific food allergies also cross react with latex. It is important to choose products low in allergen content and powder free to minimize exposure when use of latex products is essential. Using nonlatex products whenever possible is essential. Education of health care workers, clients, and their families about latex products, latex allergy, and substitutions for latex products needs to be current and ongoing. Development of clear guidelines related to working with health care workers with latex allergies encourages management to consider the seriousness of latex allergy and ways to avoid it. PMID- 10865540 TI - Depression in the workplace. Impact on employees. AB - Given the current high incidence of depression, occupational health nurses in all work settings are likely to encounter many employees who suffer from some form of depression. Depression has the highest medical benefit costs for all behavioral conditions and results in more days of disability than chronic medical conditions such as heart disease, hypertension, diabetes, and lower back pain. Advances in understanding the physiologic changes in the brain which cause depression have lead to development of effective psychopharmacologic agents for treatment. Depressed individuals have the most positive responses with a combination of medication and psychotherapy. Nurses need to assist affected employees to obtain optimal mental health care so they may remain as functional as possible, thereby diminishing the detrimental effects of the illness. PMID- 10865541 TI - Paying attention to what customers are saying. PMID- 10865542 TI - Jane Parker-Conrad, international consultant. Interview by Eileen Lukes. PMID- 10865543 TI - Competencies in occupational and environmental health nursing. Practice in the new millennium. American Association of Occupational Health Nurses. PMID- 10865544 TI - Confidentiality through a feminist lens. Reaction piece. AB - An ethical foundation for confidentiality is found in the AAOHN Code, grounded in a respect for autonomy. Confidentiality is a phenomenon involving a trust relationship between at least two people. An undesirable outcome could result if the confidential information or observation is revealed. Although confidentiality has an impact regardless of the individual's gender, women can be at higher risk for compromises to confidentiality. The implications for women include but are not limited to the inherent inequality in client/professional relationships, paternalistic practices in health care delivery, and the importance of autonomy as an ethical principle. The nurse's sensitivity to the possible vulnerability of women, the recognition of the importance of trust and communication in professional relationships, and an effort to ask the gender question contribute to an ethical practice in occupational health care related to the issue of confidentiality. PMID- 10865545 TI - Information management. Computer resources for the occupational and environmental health nurse. AB - Occupational and environmental health nurses are responsible for the management of large amounts of very complex information, ranging from individual employee health records to reports that insure corporate compliance. There are four primary tools available to the occupational health nurse to facilitate efficient management and use of health information--occupational health information systems, office support programs, communication systems, and the Internet and intranets. Selection and implementation of an integrated health information system requires the involvement of any organization that uses data processed by the system. A project management approach to implementation and maintenance of a system insures adherence to time lines and attention to details. The internet provides access to a vast amount of information useful to both the occupational health professional and the employee. Intranets are internal systems that may facilitate distribution of health information to employees, maintenance of current health related policies, and more efficient reporting procedures. PMID- 10865546 TI - Agency for Health Care Policy and Research guidelines on rhinosinusitis and depression. PMID- 10865547 TI - The occupational and environmental health nurse and health surveillance. PMID- 10865548 TI - Screening and surveillance. OSHA's medical surveillance provisions. AB - The OSH Act requires OSHA to include provisions for medical examinations of employees in its standards. However, the specific test and examinations criteria are not outlined in the OSH Act. Instead, each standard has specific medical surveillance requirements. These are specific to the adverse health effects triggered by exposure to the hazardous substance. The OSHA uses the term medical surveillance to refer to its employee examination and testing provisions. Most occupational health professionals call this activity employee screening and reserve the term surveillance for aggregate analysis of population data. It is important to remember this distinction when referring to OSHA standards. Many standards are challenged in court resulting in changes to medical surveillance provisions of the standards. Some court decisions support OSHA's language. In either case, the court often sets precedents for future standards. PMID- 10865549 TI - Health surveillance for health care workers. A vital role for the occupational and environmental health nurse. AB - Health surveillance for the general health care worker includes surveillance for immunity to infectious diseases such as measles, mumps, rubella, varicella, and hepatitis B. Post exposure surveillance for bloodborne pathogens includes HIV, hepatitis B, and hepatitis C. Periodic surveillance of specialty practice areas as mandated by OSHA include workers exposed to lasers, radiation, formaldehyde, ethylene oxide, hazardous drugs, anesthetic gases, and tuberculosis. PMID- 10865550 TI - Genetic information in the workplace. Implications for occupational health surveillance. AB - Rapid progress in understanding the human genome has made individual genetic information accessible through genetic testing. Different types of genetic testing may be encountered in the workplace. Genetic screening examines individuals for specific inherited characteristics. Genetic monitoring evaluates individuals for acquired modifications to their genetic material. A recent survey provides evidence that some employers are conducting genetic testing of their employees and using the genetic information they obtain to make employment related decisions. Occupational and environmental health nurses must be prepared to meet the challenges presented by the complex issues related to genetic information in the workplace. PMID- 10865551 TI - Occupational health surveillance, screening, and prevention activities in occupational health nursing practice. AB - Occupational health nursing practice is broad and encompasses surveillance, screening, and prevention activities as part of the scope of practice. While there has been some controversy about who is responsible for these activities, it is clear occupational health nurses play a pivotal role in overseeing, managing, implementing, and evaluating these programs. In fact, recent OSHA standards have included broad language that permits licensed health care professionals acting within their legal scope of practice to conduct medical and health surveillance activities. While the contributions of occupational health nurses are well documented, little is known about the degree and emphasis in practice related to surveillance, screening, and prevention programs. This study examined the scope of independent and interdependent practice by occupational health nurses related to these activities and found 71% of occupational health nurses had overall responsibility for program management, and the majority performed surveillance, screening, and prevention functions as independent practice. Physician supervision for any of these activities ranged from only 0% to 8% in reporting. The results of this study validate the independent functioning in scope of occupational health nursing practice related to surveillance, screening, and prevention activities while recognizing the contributions all providers make to a healthy work force. PMID- 10865552 TI - Minimizing litigation risk. Documentation strategies in the occupational health setting. AB - When advice is given by telephone, nurses are relying on employees' or clients' own assessments of situations. Nurses do not have the benefit of examination and objective findings. Therefore, every occupational health practice should have a system for keeping a record of telephone calls. Noncompliance should be documented so the nurse is reminded of the need to consider compliance when caring for the client in the future. Documentation of report tracking and follow up, consent, client education, and discharge information contributes to improved quality of care and reduced risk of litigation. Client records should never be altered (i.e., changed) so the original entry is no longer visible. The SLIDE (Single Line, Initials, Date, Explanation) rule should be used. PMID- 10865553 TI - Adding up 30 years of childbirth advocacy: how far have we come? PMID- 10865554 TI - Maternal mortality, United States and Canada, 1982-1997. AB - BACKGROUND: The 1998 public awareness campaign on Safe Motherhood called attention to the issue of maternal mortality worldwide. This paper focuses upon maternal mortality trends in the United States and Canada, and examines differentials in maternal mortality in the United States by maternal characteristics. METHODS: Data from the vital statistics systems of the United States and Canada were used in the analysis. Both systems identify maternal deaths using the definition of the World Health Organization's International Classification of Diseases. Numbers of deaths, maternal mortality rates, and confidence intervals for the rates are shown in the paper. RESULTS: Maternal mortality declined for much of the century in both countries, but the rates have not changed substantially between 1982 and 1997. In this period the maternal mortality levels were lower in Canada than in the United States. Maternal mortality rates vary by maternal characteristics, especially maternal age and race. CONCLUSIONS: Maternal mortality continues to be an issue in developed countries, such as the United States and Canada. Maternal mortality rates have been stable recently, despite evidence that many maternal deaths continue to be preventable. Additional investment is needed to realize further improvements in maternal mortality. PMID- 10865555 TI - Declining trends in cesarean deliveries, Ohio 1989-1996: an analysis by indications. AB - BACKGROUND: Similar to trends observed nationwide, the rates of cesarean deliveries declined in Ohio during the late 1980s and the early 1990s. This study examined the trends in cesarean deliveries in Ohio from 1989 through 1996, in the presence or absence of indications, and in relation to the use of obstetric procedures. METHODS: Birth certificate data for all singleton, liveborn infants in Ohio (n = 1,204,859) were used to analyze temporal trends in cesarean sections. RESULTS: The rates of primary and repeat cesarean deliveries declined, respectively, from 15.7 to 12.4 percent and from 83 to 63.3 percent during the 8 year study period. Significant declines in repeat cesarean deliveries were observed both in the presence and absence of documented medical conditions that could present a potential indication for the procedure. The rates of repeat cesareans remained comparable among women with and without documented indications for cesarean section (64% and 61%, respectively). In addition, 45 and 30 percent of repeat cesareans in 1989 and 1996, respectively, were performed in the absence of any documented indications, or on an elective basis. The declines in cesarean delivery rates during the 8-year study period occurred simultaneously with an increase in the use of electronic fetal monitoring, induction, and stimulation of labor. CONCLUSIONS: The findings suggest that a sizable proportion of repeat cesarean deliveries in 1996 may be unnecessary, even though a marked decline in the procedure has occurred between 1989 and 1996. PMID- 10865556 TI - Routine use of external cephalic version in three hospitals. AB - BACKGROUND: External cephalic version has been advocated as a safe alternative to vaginal breech delivery or cesarean birth. The purpose of this study was to determine the efficacy of routine use of external cephalic version at 36 weeks or more of gestation in three different levels of hospitals. METHODS: External cephalic version was performed on 923 women with a single breech fetus at three hospitals in Italy. The procedure was attempted with a tocolytic agent for uterine relaxation and with no maternal analgesia. The version technique adopted was the "forward roll." RESULTS: Version was successfully performed on 579 fetuses (62.7%); each hospital had a similar success rate, and 56.9 percent of the women delivered vaginally. The procedure was more successful in multiparas and in women with an incomplete type of breech, polyhydramnios, and posterior localization of the placenta. Vaginal bleeding was experienced by 14 women; eight cesarean sections were performed for suspected abruptio placentae, confirmed in four cases. Two cephalic-turned fetuses experienced an episode of persistent bradycardia and were turned again to breech; in five cases a nonstress test recorded after the version showed repeated variable decelerations and in one case a cesarean section was performed. Neonatal outcomes were good in 922 infants. A fracture of the femur attributable to the version was observed in one newborn. CONCLUSIONS: External cephalic version is effective in reducing the number of cesarean deliveries in term breech infants in different obstetric settings, with no major neonatal adverse outcomes. PMID- 10865557 TI - The relationship of maternal personality characteristics to birth outcomes and infant development. AB - BACKGROUND: Previous studies reported an association between maternal psychological factors and adverse pregnancy outcomes. The objective of this study was to evaluate the relationships between maternal personality characteristics, as determined by the Minnesota Multiphasic Personality Inventory (MMPI), and infant birth outcomes and development. METHOD: The inventory was administered during pregnancy to 638 pregnant women enrolled in a staff model health maintenance organization. MMPI validity as well as clinical and research scales were evaluated in relationship to infant birth outcomes (low birthweight, preterm birth) and 15-month-old infant development as assessed by the Bayley Scales of Infant Development. RESULTS: Mothers of low birthweight infants scored significantly lower on the hypochondriasis scale, a relationship which was no longer significant after controlling for ethnicity. No other relationships were observed between infant birth outcomes and maternal MMPI scale scores. A higher infant Mental Developmental Index (MDI) was related to higher maternal masculinity-femininity and ego-strength scale scores and lower lie and hypochondriasis scale scores. Only the relationship between infant MDI and maternal masculinity-femininity scale score remained significant after controlling for ethnicity and socioeconomic index (beta = 0.104, p = 0.036). CONCLUSIONS: Maternal personality characteristics, as determined by the MMPI, did not appear to be significantly related to the occurrence of preterm birth or low birthweight in this healthy, general population. Maternal personality characteristics reflected in the MMPI masculinity-femininity scale appeared to be related to infant mental development, above and beyond the effects of socioeconomic status and ethnicity. PMID- 10865558 TI - Pregnancy outcome after previous stillbirth resulting from causes other than maternal conditions and fetal abnormalities. AB - BACKGROUND: An adequate fetomaternal circulatory system may be compromised by a variety of disturbances leading to stillbirth. The purpose of this study was to assess subsequent pregnancy outcome in women with a history of stillbirth as a result of causes other than maternal conditions and fetal abnormalities. METHODS: Ninety-two deliveries after stillbirth were identified among 11,910 deliveries of parous women recorded in the birth registry at Kuopio, Finland. Using logistic regression, pregnancy outcome measures were compared with those of a parous healthy obstetric population (n = 11,818). RESULTS: Women with a history of stillbirth as a result of causes other than maternal conditions and fetal abnormalities were older than their unaffected controls (32.4 yr vs 30.3 yr). Stillbirth in an earlier pregnancy was associated with a significantly higher (p < 0.001) frequency of placental abruption in subsequent pregnancy (5.4% vs 0.7%). A history of stillbirth was predictive of preterm delivery (OR = 2.25) and low birthweight infants (OR = 2.70). No recurrence was reported. CONCLUSIONS: Pregnancy with a history of stillbirth as a result of causes other than maternal conditions and fetal abnormalities is a moderate risk state, with prematurity and low-birthweight rates somewhat higher than those in the general population. The overall probability of a favorable outcome is good. These findings may be useful in counseling pregnant women with a history of stillbirth. PMID- 10865560 TI - Commentary: ethical conflicts or political problems in intrapartum nursing care? PMID- 10865559 TI - Intrapartum nursing care: a case study of supportive interventions and ethical conflicts. AB - BACKGROUND: Studies have shown that one-to-one labor support is associated with a reduced rate of operative births, and with long-term improvements of parenting and breastfeeding rates. Labor support by nurses may reduce the cesarean birth rate, but this has not been adequately studied. No one knows which labor support strategies nurses use, if they are effective, and how they work. METHODS: This pilot study used the qualitative techniques of observation and an audiotaped interview with an expert intrapartum nurse to describe labor support techniques and strategies to enhance labor progress and prevent cesarean births. RESULTS: The narrative revealed three major themes. The first theme, "the nurse's approaches to labor," included three subcategories: "following the mother's body," "hastening and controlling labor," and "labor support techniques." The second and third major themes, "ethical dilemmas: an unwilling partnership" and "nurse-physician conflict," were unanticipated. Labor support practices were limited by some physician practices. Inappropriate physician practice created ethical dilemmas for the nurse and impeded labor support interventions. CONCLUSIONS: Intrapartum nursing care reflected both a medical model of controlling and hastening birth, as well as a supportive, nurturing, and empowering model of practice that used independent clinical judgments and advocacy. Questionable medical care entangled the nurse in these practices and created moral dilemmas and nurse-physician conflicts. The nurse used various strategies to promote the wishes and welfare of the laboring mother. PMID- 10865561 TI - Resident physicians' knowledge of breastfeeding and infant growth. AB - BACKGROUND: It is well documented that breastfed infants grow differently from formula-fed infants. The purpose of this study was to assess resident physicians' knowledge of breastfeeding and infant growth. METHODS: A cross-sectional, self administered survey was administered to family medicine and pediatric resident physicians from three large, hospital-based public and private programs in North Carolina. RESULTS: One hundred and seven (46%) of 235 residents completed the study, representing 55 percent of family medicine residents and 39 percent of pediatric residents. Ninety-nine percent of participants reported frequently or always plotting infant growth at well-child visits. None reported plotting breastfed babies on a chart specific to breastfeeding. Only 5 percent of participants knew that breastfed infants grew at a slower velocity than formula fed infants after 4 months of age. This knowledge was not significantly related to specialty, year of training, or gender; it was significantly related to breastfeeding experience (p < 0.04). Of the residents who did not have personal experience with breastfeeding, 99 percent answered incorrectly compared with 88 percent of those who had some personal experience in breastfeeding. CONCLUSIONS: In this sample of family medicine and pediatric residents, almost all were unaware that breastfed infants grow at slower rates after 4 months of age. Since the frequency of breastfeeding is increasing in the United States, it is important that physicians be able to monitor the growth of breastfed infants accurately and provide expert counseling for breastfeeding mothers. PMID- 10865562 TI - U.S. cesarean and VBAC rates stalled in the mid-1990s. PMID- 10865563 TI - Sterile water injections relieve back pain of labor. PMID- 10865564 TI - Sheila Kitzinger's letter from Europe: home birth matters. PMID- 10865565 TI - Epidural labor analgesia and cesarean delivery. PMID- 10865566 TI - Nulliparas and epidural analgesia. PMID- 10865567 TI - Would you like fries with that? Health care as a business. PMID- 10865568 TI - The use of bupropion hydrochloride for smoking cessation therapy. AB - All healthcare providers have an obligation to promote smoking cessation to their patients who smoke. While patients are advised to stop smoking, the use of specific smoking cessation strategies is rarely addressed by primary care providers. This article discusses smoking cessation for two specifically vulnerable groups: women and African-Americans. Both groups have been identified as having increased rates of nicotine metabolism and may be less likely to respond effectively to nicotine replacement for smoking cessation. Increased nicotine metabolism results in greatest difficulty with successful smoking cessation and creates an increased vulnerability to a wide range of cardiovascular and respiratory health problems associated with smoking. Nicotine replacement therapy for smoking cessation has been the most common treatment available; however, many patients have not been successful with that method, and a majority (75%) of smokers who attempt to stop smoking return to smoking. The use of bupropion hydrochloride (Zyban) as a nonnicotine replacement therapy for smoking cessation is discussed. Relapses in smoking cessation are due to nicotine craving and the attempt to alleviate symptoms of nicotine withdrawal. Bupropion hydrochloride, an oral antidepressant, is believed to work by elimination of nicotine cravings and to decrease the physiologic and psychologic symptoms associated with nicotine withdrawal. Patients who have not been successful in smoking cessation may benefit from bupropion hydrochloride therapy in conjunction with counseling and behavior modification. PMID- 10865569 TI - Implementing the Agency for Health Care Policy and Research smoking cessation guidelines: a call to action. AB - Smoking is the single most preventable cause of death in our society. Each year, cigarettes kill more Americans than AIDS, alcohol, car accidents, fires, illegal drugs, murders, and suicides combined. An estimated 46 million adults in the United States smoke--28% of all men and 23.5% of all women. Seventy percent of Americans who smoke say that they would like to quit. Additionally, 70% who smoke visit a healthcare provider each year. Unfortunately, half of these who seek health care each year say they have never been advised to quit smoking or provided specific strategies to be successful at quitting. In 1996, the Agency for Health Care Policy and Research (AHCPR) published smoking cessation materials in several formats that target different audiences. This article describes the significant health problems of smoking and the AHCPR publications for smoking cessation. Providers are encouraged to identify smoking cessation as a priority and incorporate the AHCPR guidelines into routine health care for all patient encounters. PMID- 10865570 TI - Anemia: an overview of diagnostic considerations. AB - Anemia is a significant issue in clinical practice. Reducing anemia can improve quality of life. Advanced practice nurses are critical to the diagnostic evaluation of anemia. The key to effective treatment is establishment of the anemic etiology. This can be done through a straightforward diagnostic approach utilizing the traditional history and physical, and standard laboratory assessment. The diagnostic process must be based on an understanding of demographics, normal physiology, and the pathophysiologic processes of the different anemias. Nurse practitioners are uniquely educated to assess anemia in a holistic manner, incorporating components of the history, physical exam, and laboratory evaluation into an appropriate diagnosis. Once the diagnosis is established, this same holistic assessment can be utilized to create a specific, patient-focused plan of care and follow-up. PMID- 10865571 TI - Amyotrophic lateral sclerosis: a team approach to primary care. AB - Amyotrophic lateral sclerosis (ALS) is a progressive neuromuscular disease that causes degeneration of motor neurons and results in death. The most frequent presenting symptom of patients with ALS is muscle weakness. The role of the nurse practitioner (NP) in the diagnosis and management of ALS is multifaceted. The NP could serve as the coordinator of care provided by a multidisciplinary team or as a member of the team. The coordinator of the team may vary depending on the individual needs of the patient with ALS and the stage of illness. Muscle weakness and spasticity, respiratory difficulties, and speech and swallowing problems are frequently encountered problems needing intervention. The role of the NP is important in providing symptomatic relief for the patient, and essential in providing education and psychosocial support for the patient and family caregivers. This article reviews the pathophysiology, assessment, management, and recent advances in research related to ALS. The challenging role of the NP in improving the quality of life of patients with ALS and their families is discussed. PMID- 10865572 TI - Complementary therapeutic practices in patients with chronic sinusitis. AB - Understanding patient use of alternative and complementary modalities to treat chronic health conditions is an important component to providing holistic care. This study sought to identify traditional and complementary therapies used by patients with chronic sinusitis. Eighty-one percent of patients with chronic sinusitis engaged in physical exercise to relieve symptoms. Additional complementary therapies utilized included herbal therapy (32%), chiropractic therapy (16%), biofeedback (13%), acupuncture (11%), and chelation therapy (7%). Medications were commonly used by patients (60%), especially those with severe symptoms. By recognizing and incorporating effective complementary therapies into care for chronic sinusitis, nurse practitioners may help patients to improve their clinical outcomes. PMID- 10865573 TI - Relationship of the menstrual cycle to postoperative incidence of emesis after laparoscopic cholecystectomy. AB - Despite advances in anesthetic practice and surgical techniques, the overall incidence of postoperative vomiting (POV) ranges from 20% to 30% during the first 24 hours after surgery. Studies specifically examining the relationship between POV and the menstrual cycle demonstrate equivocal findings. The purpose of this study was to examine the relationship between the incidence of POV and the phase of the menstrual cycle, age, body mass index, history of POV, duration of anesthesia, and use of droperidol among 79 women undergoing laparoscopic cholecystectomy. Sixteen subjects were stratified to the menstruating group and 63 to the nonmenstruating group. Analysis of demographic data revealed no significant differences between groups. Chi square analysis indicated that women in the menstruating group were more likely to experience POV (chi 2 = 20.87, P < .01). Among those receiving droperidol (n = 43; 54%), analysis revealed that those in the menstruating group experienced POV at a significantly higher rate (chi 2 = 9.15, P < .05). A reduction of POV may be accomplished by scheduling elective surgery to avoid the correlating menstrual cycle peaks. PMID- 10865574 TI - Revisiting the future: healthcare practice and education. PMID- 10865575 TI - Using the World Wide Web for research data collection. PMID- 10865576 TI - Profiles of editorial board members: east to west. PMID- 10865577 TI - Taking the lead. PMID- 10865578 TI - Adverse effects of paclitaxel in patients with alcohol abuse histories. AB - Paclitaxel is a taxane antineoplastic agent formulated in polyoxyethylated castor oil (Cremophor El) and dehydrated alcohol. This case report describes the development of central nervous system toxicity and threatened sobriety related to the alcohol content in a paclitaxel infusion. Nurses need to be aware of the alcohol contained in paclitaxel and other drugs used in oncology practice and document previous and current alcohol use prior to beginning therapy. Alternative chemotherapy regimens may need to be offered to patients with alcohol abuse histories. PMID- 10865579 TI - A nursing protocol for the management of perineal-rectal skin alterations. AB - Perineal-rectal care is defined as skin care to the region between the vulva and anus in the female and scrotum and anus in the male (Costello, 1997). Perineal rectal care is essential to prevent infection and promote comfort but is complicated by the anatomical location of the perineum and rectum. In addition, no standardized perineal-rectal care approach exists. Patients with cancer are at particular risk for developing perineal-rectal skin breakdown because of immunosuppression, side effects of radiation and chemotherapy, and compromised nutritional status. A perineal-rectal skin-care protocol is presented that incorporates recommendations for routine care as well as recommendations for managing common skin alterations, including erythema, dry and moist desquamation, and infection. PMID- 10865580 TI - Optimizing nutrition for patients with cancer. AB - Malnutrition is the most common secondary diagnosis in patients with cancer and is a major prognostic indicator for poor response to cancer therapy and shortened survival. The word CANCER can be used as an acronym to guide early nutritional intervention that combines an understanding of cancer cachexia with a plan to optimize patients' nutritional status during the stress imposed by cancer and its treatment. Conversations with patients reveal nutrition-related information that is valuable as part of a comprehensive assessment. The Assessment combines nutrition-related information with observations made during the conversation. The Nutrition plan is developed to meet patients' identified nutritional needs. The goals of the cancer treatment dictate the aggressiveness of nutritional support. An awareness of potentially life-threatening Complications of enteral and parenteral nutrition is essential. An ongoing Evaluation of patients' status and responses to the nutrition plan guides changes to the plan as needed. Reassurance/Removal are important parts of care during all stages of treatment and nutrition support. PMID- 10865581 TI - Hemorrhage after bone marrow harvest: a case presentation. AB - The purpose of this article is to describe the usual procedure and postoperative recovery after an allogeneic bone marrow harvest and to present a case study of an unusual complication of hemorrhage. The case study describes a donor who experienced hemorrhage with severe pain, muscle spasms, and prolonged limitations in range of motion and ambulation. Oncology nurses should inform donors to promptly report persistent pain, spasms, and muscle weakness. Should hemorrhage occur, blood loss should be evaluated, bedrest should be maintained, and cold packs should be applied to the area. Although excessive bleeding is a rare occurrence, nurses should be alert for this complication to prevent pain and activity impairment. PMID- 10865583 TI - The undertreatment of pain: a liability risk for nurses. PMID- 10865582 TI - Silence is not golden: conversations with the dying. AB - Conversations about dying involve interaction on multiple intellectual and emotional levels. The paradigm of clinicians as existential messengers is characterized by process and content and validated by the clinicians' ongoing presence. The necessary attributes that allow clinicians to function as existential messengers and communicate effectively with the dying include recognition of and confidence in one's own strengths, limits, and capabilities; clarity in one's personal understanding of living and dying; the ability to listen carefully and learn from the experience; the willingness to remain present in the face of profound anguish; and a strong belief in the worth of the work. In the process, personal biographies are amended permanently and clinicians are wiser for the experience. PMID- 10865585 TI - Families' awareness of and response to dying. PMID- 10865584 TI - Cancer patients' reported experiences of suffering. PMID- 10865586 TI - Diagnosing graft versus host disease. PMID- 10865587 TI - An overview of complementary and alternative therapies. PMID- 10865588 TI - Paclitaxel. PMID- 10865589 TI - A rainbow of opportunities: advanced practice. PMID- 10865590 TI - Healthy lifestyles: a pilot program to influence behavior change. PMID- 10865591 TI - Use of clinical practice recommendations for exercise by individuals with type 1 diabetes. AB - PURPOSE: The purposes of this study were to determine the extent to which physically active individuals with type 1 diabetes actually follow exercise recommendations and to compare their use of current with previous recommendations. METHODS: A questionnaire was developed consisting of the American Diabetes Association's current clinical practice recommendations for exercise and some previous recommendations. The questionnaire was mailed to approximately 1700 members of the International Diabetic Athletes Association; responses from 238 questionnaires were included in the results. Respondents indicated to what extent they currently use all of these recommendations and any modifications that they employ. RESULTS: The responses indicated that individuals with type 1 diabetes closely followed the majority of the current clinical exercise recommendations and did so to a greater extent than previous ones. Individuals' specific modifications to the current recommendations are summarized. CONCLUSIONS: The current clinical practice recommendations are used by exercisers with diabetes and are followed more closely than previous ones. The significant number of reported modifications made by individuals indicates, however, that a substantial need still exists to modify and individualize the recommendations. PMID- 10865592 TI - Recommendations of dietitians for overcoming barriers to dietary adherence in individuals with diabetes. AB - PURPOSE: The purposes of this research were to (1) identify factors that contribute to the barriers to dietary adherence in individuals with diabetes identified in a 1998 study and (2) obtain recommendations from registered dietitians for strategies to overcome these barriers. METHODS: A 10-item, open ended telephone questionnaire was used to obtain information. The sample included 75 registered dietitians who participated in a previous survey to identify barriers and agreed to a follow-up telephone interview. RESULTS: Of the 75 participants, 28% reported spending 5 hours or less per week counseling individuals with diabetes, 64% spent between 6 and 30 hours, and 8% spent more than 31 hours per week. Almost half of the participants (47%) were certified diabetes educators. Factors identified as the greatest contributors to the barriers being evaluated included lack of time, lack of symptoms, lack of education (including follow-up), poor self-esteem/lack of empowerment, and misinformation from family/peers/others with diabetes. The primary recommendations for overcoming each of these barriers included individualizing meal plans and planning ahead, teaching about complications, and setting obtainable goals. CONCLUSIONS: The registered dietitians who were surveyed emphasized the importance of individualizing dietary counseling. PMID- 10865593 TI - Assessing the cultural relevance of an education program for urban African Americans with diabetes. AB - PURPOSE: This study was conducted to assess the cultural relevance of an education program for urban African Americans with diabetes. METHODS: A set of 12 videotape vignettes were developed for use in diabetes education for urban African Americans with diabetes. Focus groups and questionnaires were used to determine if patients and diabetes educators would find the materials stimulating, culturally appropriate, and useful. RESULTS: The videotape and discussion guide were perceived as valuable by both healthcare professionals and patients. CONCLUSIONS: This education program could be a valuable resource for diabetes educators who want to provide culturally sensitive and relevant diabetes education for urban African Americans with diabetes. PMID- 10865594 TI - Diabetes education materials: recommendations of tribal leaders, Indian health professionals, and American Indian community members. AB - PURPOSE: The Association of American Indian Physicians, the only national organization of its kind, conducted a series of focus groups to gather input from tribal leaders, Indian health professionals, and American Indian community members to guide the development of culturally appropriate diabetes education materials for the National Diabetes Education Program. METHODS: During the focus groups, participants shared their experiences with and recommendations for a variety of diabetes education materials. RESULTS: Overall, 95% of participants expressed a strong preference for diabetes education materials relevant to their specific tribe or culture. CONCLUSIONS: Recommendations from these focus groups were used to develop a national diabetes education campaign for American Indian communities. PMID- 10865595 TI - Health-promotion practices of young black women at risk for diabetes. AB - PURPOSE: The purpose of this study was to assess the health-promoting practices of young black women at risk for type 2 diabetes. METHODS: The sample consisted of 30 black women from an urban area who had a history of gestational diabetes and/or a first-degree relative with diabetes. Participants completed the Health Promoting Lifestyle Profile II Survey and an interview. Both were used to categorize health-promoting practices, exercise, diet, knowledge of diabetes prevention, and general health. RESULTS: Demographic information and interview revealed a propensity towards obesity, despite education and income levels. The results for the Lifestyle II Survey showed a higher average total score for healthy nutrition than physical activity, which were inconsistent with the qualitative data obtained by interview. Fifty percent stated that they exercised as a general health-promoting behavior. Self-reported daily caloric, fiber, and fat intake was high to moderate; 60% reported initiating diet modifications secondary to a desire to lose weight or for medical problems; and 26% reported receiving information on diabetes prevention from a healthcare provider. CONCLUSIONS: A systematic approach of planning and actively incorporating health promoting activities into one's lifestyle as a young adult may protect or delay the onset of diabetes and prevent complications. PMID- 10865596 TI - Nominal group technique: a process for identifying diabetes self-care issues among patients and caregivers. PMID- 10865597 TI - Reducing home nursing visit costs using a remote access infusion pump system. AB - The use of a remote access infusion pump system in patients receiving intravenous therapy in the home provided a method to monitor patients and to reduce the number of unscheduled and after-hours nursing visits. Option Care of Rockwall, Texas, a locally owned IV company that is part of a national network, was able to avoid 13 nursing visits during a 4-month period with patients receiving pain management or continuous heparin therapy with the use of an infusion pump system that allowed remote access by telephone. An appreciable cost savings was documented with the use of this device. PMID- 10865598 TI - Selection and management of central venous access devices in the home setting. AB - In the last decade new central venous access devices (CVADs) inserted for long term therapy have replaced conventional peripheral venous access devices. This shift contributes to the need for additional education as technological advances result in additional options for central venous access. Healthcare's transition from the hospital to a community-based system has increased the use of CVADs in the home setting. Issues that confront the patient with a CVAD in the home setting must be examined more closely than ever before. PMID- 10865599 TI - Understanding the obstacles to consistent intravenous catheter-related infection reporting by home health providers. AB - A variety of formulae have been used by home health providers to obtain consistent and accurate reporting of intravenous catheter-related infections. This article describes the most widely used statistical formulae, their advantages and disadvantages, and their relevance to the home health setting. However, because of the numerous and complex intervening variables that influence these statistical applications, epidemiological measurement in nonhospital settings remains an unresolved dilemma for most providers. PMID- 10865600 TI - Attachment impacts a culturally diverse population in the homecare setting. AB - Home intravenous therapy requires knowledge and understanding for successful outcomes. This article presents the ideas of attachment, guidance, and culturally sensitive nursing practice interventions when caring for patients who require home i.v. therapy. Culturally sensitive nursing interventions can increase a patient's ability to comprehend the treatment plan if they are presented in a familiar context. As trust is built, barriers to accurate communication between the nurse and the patient and family are removed, resulting in increased patient compliance and learning. Four case studies are presented to illustrate how nursing interventions use the cultural backgrounds of patients to develop culturally sensitive nursing interventions and to show how decisions are made based on information regarding cultural norms. PMID- 10865601 TI - CytoGam infusions at home. AB - CytoGam is a hyperimmune globulin created specifically for prophylactic use when a cytomegalovirus seronegative patient receives an organ from a seropositive donor. CytoGam is derived from human pooled plasma rich with cytomegalovirus antibodies, immunoglobulin G, and trace amounts of immunoglobulin A and immunoglobulin M. Clinical studies show a marked decrease in the development of cytomegalovirus when transplant recipients are treated with CytoGam. CytoGam also has undergone trial testing for use in decreasing the instance of cytomegalovirus in patients with human immune deficiency virus (HIV). CytoGam therapy may continue for several months, making home intravenous care essential. PMID- 10865602 TI - Vancomycin peak serum concentration monitoring. AB - Vancomycin is a glycopeptide antibiotic commonly used in the treatment of gram positive bacterial infections. With the rising prevalence of resistant gram positive bacterial infections in healthcare institutions and the advent of attempts to curb escalating medical cost, vancomycin is being used with increasing frequency in the homecare setting for infection management. In view of its variability in pharmacokinetics and the potential for toxicities, serum concentrations often are measured. This article reviews the pharmacology of vancomycin and suggests a practical approach to serum concentration monitoring in homecare. PMID- 10865603 TI - Predisposing factors, prevention, and management of central venous catheter occlusions. AB - Occlusion is the most common noninfectious complication of central venous access catheters. The primary approach is prevention. Successful management begins with an understanding of the potential etiologies of occlusions. Proper assessment aids the clinician in correctly identifying the cause and treatment strategy. This article focuses on the physiologic principles of, research on, and current clinical practices with central venous catheter occlusions. PMID- 10865604 TI - Infection control principles and practices in the care and management of central venous access devices. AB - Infectious complications occur in a significant percentage of patients with central venous access devices (CVADs). This article reviews the recognition and treatment of CVAD-related infectious complications and suggests methods to decrease the risk of such complications. In addition, algorithms for prevention and treatment of CVAD-related infections are presented. PMID- 10865605 TI - Quality issues in access device management. AB - Patients are moving more rapidly from one healthcare setting to another. Often these patients have more complicated therapies and access devices and more potential for adverse occurrences. To provide care for these patients across healthcare settings, institutions need to plan a framework for delivery of quality care and services while minimizing the potential for complications. This article describes methods, tools, and other resources useful for promoting quality care in patients with more complex parenteral routes of administration. PMID- 10865606 TI - The role of the interventional radiologist in central venous access. AB - The role of the interventional radiologist in the care of patients requiring placement of central venous access devices is rapidly evolving. With experience gained from diagnosing and treating central venous catheter-related complications, interventional radiologists are assuming an increasing role in the placement of these devices. With imaging guidance, catheter and guidewire skills, and a commitment to providing a clinical service that includes management of catheter malfunctions and complications, central venous access by the interventional radiologist has proven a safe and effective alternative to standard surgical techniques. PMID- 10865607 TI - Care and management issues regarding central venous access devices in the home and long-term care setting. AB - Experience and comfort with central venous access devices (CVADs) has increased dramatically during the past 2 decades. However, coordination of care remains a challenge as patients with long-term catheters move between levels of care with multiple healthcare providers. Standardizing CVAD maintenance and the teaching of patients and caregivers across the care continuum can be beneficial for patients and professionals. Effective communication among all care providers enhances teamwork and improves efficiency. The consistent collection and evaluation of data regarding CVAD complications and outcomes is important for assessing quality and determining best practices. PMID- 10865609 TI - Notes from the Regulatory and Reimbursement Subcommittee: establishing reimbursement outpatient services. PMID- 10865608 TI - Correct utilization and management of peripherally inserted central catheters and midline catheters in the alternate care setting. AB - Based on patient condition, intravenous therapies, caregiver support, and organizational policy, correct device selection plays an integral part in the overall care and management of the alternate care setting i.v. therapy patient. This paper will identify the various aspects of appropriate device selection for i.v. therapy prescriptions. PMID- 10865610 TI - A primer for JWOCN authors. II: Case Studies and Options in Practice. PMID- 10865611 TI - E-mail etiquette: tips for effective use. PMID- 10865612 TI - Moving from concept to research design. PMID- 10865613 TI - Restructuring the therapeutic environment to promote care and safety for the obese patient. AB - Fifty-four percent of American adults are overweight. Obesity is a chronic disease associated with a number of conditions, such as diabetes, heart disease, hypertension, certain types of cancers, and breathing problems. The direct and indirect costs related to obesity exceed $70 billion annually. Because of the many cost and quality issues related to obesity, national attention is turning toward the special needs of this population. Strategies to restructure therapeutic intervention with attention to risk management, economic implications, and patient satisfaction are important considerations when managing the obese patient. PMID- 10865614 TI - A multidimensional modeling of predictors influencing the adjustment to a colostomy. AB - PURPOSE: Factors contributing to the postoperative adjustment of patients who had undergone permanent colostomy surgery were studied. DESIGN: Cross-sectional study. SETTING AND SUBJECTS: Sixty patients who underwent colostomy surgery at 5 hospitals in Santiago, Chile, between 1987 and 1995 were evaluated. Subjects were between the ages of 27 and 89 and underwent colostomy surgery from 4 months to 19 years before data collection. METHODS AND INSTRUMENTS: A demographic questionnaire and a semistructured interview were used to elicit information concerning psychosocial reactions to a colostomy and perceptions about the changes in lifestyle created by the ostomy. Medical records were reviewed for information about disease-related characteristics and an ostomy self-care scale measured coping skills. Adjustment to the colostomy procedure was measured by means of the Olbrisch ostomy adjustment scale. RESULTS: Patient adaptation to a colostomy is primarily affected by the following factors: (1) the level of ostomy self-care, (2) psychological support, and (3) social support from family and significant others. CONCLUSION: Successful adjustment to a permanent colostomy is most likely to occur if the patient receives adequate instruction in self-care and has the appropriate psychological support to integrate the new physical changes into a healthy body image. This includes continued acceptance from family and the patient's social network. PMID- 10865615 TI - Severe peristomal pyoderma gangrenosum: a case study. AB - Pyoderma gangrenosum is an autoimmune disease that often manifests itself as painful ulcers. When these ulcers occur in the peristomal area, symptom management and wound care must be balanced against the need to pouch the stoma. Although the treatment of pyoderma gangrenosum is not standardized, systemic steroid therapy is frequently used as first-line therapy. The WOC nurse is often asked to manage both the pouching needs and topical therapy of the patient with peristomal pyoderma ulcerations. This article describes the management of severe pyoderma gangrenosum ulcerations in a 59-year-old woman with a long-standing ileostomy. PMID- 10865616 TI - Lifestyle factors and continence status: comparison of self-report data from a postal survey in England. AB - OBJECTIVE: To compare health and lifestyle factors of people with and without urinary incontinence (UI). DESIGN: A postal survey was undertaken that represents the first of a 3-stage project designed to evaluate the health interventions of primary health care teams and continence advisory services on patient outcomes related to UI. SETTING AND SUBJECTS: Two random samples of adult populations (N = 12,529) were included, generated from the family physician patient registers within 2 health authorities in England. INSTRUMENTS: Data were collected using a structured questionnaire that queried demography, perceived health status, activities of daily living, self-care, and use of local health and social services. Information was also collected on past and present continence status. METHODS: Structured questionnaires and a cover letter were mailed to the target population. Two reminders were sent to nonresponders to maximize the response rate. MAIN OUTCOME MEASURES: The main measures relate to factors associated with UI: mobility, sleep, childbirth, smoking, diet, body mass index, and accommodation. RESULTS: Significantly more women than men had UI (P < .0001). Respondents with UI were older than those who were continent (P < .0001). Women with UI were significantly more likely to have a greater number of pregnancies (P < .0001), were more likely to have given birth to a baby weighing more than 9 lb (P < .01), and to have had more than 4 children (P = .01) compared with women who were continent. People with UI were less likely to be single and more likely to be widowed than those who were continent (P < .0001). People who lived alone and who had UI were also significantly less likely to have a relative or friend that they could depend on for help than those who were continent (P < .001). UI was also found to be significantly associated with impaired mobility (P < .0001) and sleeping difficulties (P < .0001). No meaningful differences were found between diet and UI, although significantly more people with UI had higher mean body mass index, were obese, or reported that they felt too heavy for their height when compared with people who were continent (P < .0001). No association was found in the present study with smoking or ethnicity and UI. CONCLUSIONS: Key health and lifestyle factors associated with UI included age, gender, childbirth, mobility, sleep patterns, obesity, living alone, and access to help. These factors should be assessed when planning and implementing health care for persons with UI. Attention to these associated factors may prove useful in identifying new cases or people at risk of developing UI when screening people as part of routine health checks. This, in turn, could assist with targeting effective and efficient health care but may also contribute to prevention for some people. PMID- 10865617 TI - Sample size and power. PMID- 10865618 TI - Management of a long-term care patient with extensive necrotic pressure ulcers. PMID- 10865619 TI - Looking at monopolization in renal care: the Gambro anti-trust case. PMID- 10865620 TI - Altering the dialysis prescription. PMID- 10865621 TI - Project Hope study looks at quality of life issues and economic implications of daily dialysis. PMID- 10865622 TI - Scribner: we need to do better. PMID- 10865624 TI - Restless legs syndrome in ESRD. PMID- 10865623 TI - My experience on home hemo and nocturnal dialysis. PMID- 10865625 TI - Helping patients deal with sexual dysfunction following transplantation. PMID- 10865626 TI - Sex, intimacy, and renal disease. PMID- 10865627 TI - Sex and flowers: a patient's guide. PMID- 10865628 TI - Medication-induced sexual dysfunction in transplant recipients. PMID- 10865629 TI - DOQI influences anemia management overseas. AB - The European guidelines will form the backdrop for anemia management in European countries for the years to come. The existing differences will make literal implementation difficult in many places, however. Still, the guidelines set the standard against which the quality of anemia treatment should and must be judged. Dissemination will take time, but will go forward, in the end the patient will benefit. PMID- 10865630 TI - Assessing metastatic calcification in the ESRD patient. PMID- 10865631 TI - Planning and financing your way to dialysis center ownership. PMID- 10865632 TI - Major funding breakthrough in battle to find cure for PKD. PMID- 10865633 TI - How can we "standardize" technician training? PMID- 10865634 TI - Surviving a boil water advisory in dialysis and Y2K chlorination. PMID- 10865635 TI - Baxter's RMS disease management launches Lifeline with three access centers. PMID- 10865637 TI - Neoral, cellcept combination improves function of deteriorating kidney PMID- 10865636 TI - A status report on hepatitis C in the U.S. PMID- 10865639 TI - Patient compliance in renal replacement therapy: whose problem is it? PMID- 10865638 TI - Improving fluid compliance in the hemodialysis population. PMID- 10865640 TI - A cost analysis for transplantation and options post-Medicare. AB - The health care issue related to Medicare coverage of immunosuppressant medications has major implications for the medical professional and transplant recipient. Based on cost containment analysis, transplantation is the least expensive route to treat ESRD and justifies support for these bills in Congress to eliminate the time constraints of Medicare payment of immunosuppressant medications. Members of the renal health care community need to be active in the political arena of health care reformation and present their views to policymakers. PMID- 10865642 TI - Laparoscopic live donor nephrectomy: debating the benefits. Con: persistent complications do not justify replacing "the gold standard". PMID- 10865641 TI - Laparoscopic live donor nephrectomy: debating the benefits. Pro: similar costs to traditional surgery and procedure wins donors. PMID- 10865643 TI - Discharging a professional responsibility. PMID- 10865644 TI - Hospitals actively recruiting overseas. PMID- 10865645 TI - Staffing levels cause concern. PMID- 10865646 TI - Returning to nursing. PMID- 10865647 TI - Coping with colleagues' attitudes. PMID- 10865648 TI - What makes a good preceptor? PMID- 10865649 TI - Nurses prepare for millennium mayhem. PMID- 10865650 TI - How will nurses be compensated? PMID- 10865651 TI - Nursing skills undervalued. Interview by Anne Manchester. PMID- 10865652 TI - Major industrial relations differences. PMID- 10865653 TI - Straw vote. PMID- 10865654 TI - Administering subcutaneous heparin. PMID- 10865655 TI - Taking the sting out of painful procedures. PMID- 10865656 TI - The six honest servants of good documentation. PMID- 10865657 TI - Interpreting abnormal abdominal sounds. PMID- 10865658 TI - Myths & facts ... about patient massage. PMID- 10865659 TI - Action stat: superior vena cava syndrome. PMID- 10865660 TI - Caring for patients with donor organs. PMID- 10865661 TI - Building a bridge to Angie. PMID- 10865662 TI - What's the buzz on external bone growth stimulators? PMID- 10865663 TI - Taking a holistic approach to the dying time. PMID- 10865664 TI - Surgical site infections. Another look at banishing bugs. PMID- 10865665 TI - New drugs 2000. Part II. PMID- 10865666 TI - A spoonful of nursing. PMID- 10865667 TI - Straight talk about the patient interview. PMID- 10865668 TI - Spotlight. What you really do with this powerful documentation tool. PMID- 10865669 TI - Reading liver function values. PMID- 10865670 TI - How to get your career going. PMID- 10865671 TI - Boomers vs. busters: bridging the generation gap. PMID- 10865672 TI - Here comes the judge. PMID- 10865673 TI - How to stay cool under fire. PMID- 10865674 TI - Dealing with difficult behavior. PMID- 10865675 TI - Putting a price on retirement. PMID- 10865676 TI - How to use transparent films. PMID- 10865677 TI - Does clinical setting affect injury risk? PMID- 10865678 TI - OTC laxative woes. PMID- 10865679 TI - Defining disability. PMID- 10865680 TI - Protecting yourself against Creutzfeldt-Jakob disease. PMID- 10865682 TI - Web directory. PMID- 10865681 TI - Reaching out to Danny. PMID- 10865683 TI - Informed consent documents for BRCA1 and BRCA2 screening: how large is the readability gap? AB - The decision to undergo testing for the BRCA1 and BRCA2 mutations, which are associated with an increased risk of breast and ovarian cancer, can have long term consequences on women's lives. Women who decide to undergo such testing are required to sign informed consent documents, which indicate that they understand the test and its risks and benefits. These documents are generally written for advanced-level readers. However, the reading abilities of many women are substantially lower than the level of the consent forms, resulting in a 'readability gap'. This disparity suggests that women may not fully understand the documents they are asked to sign. The 'readability gap' poses the serious issues about informed consent, raising questions about institutional review boards and the effectiveness of the documents that are currently in use. PMID- 10865684 TI - What influences a patient's desire to participate in the management of their hypertension? AB - There are potential benefits to giving the patient a more active role in the management of his or her care. This study explored the characteristics which influence a preference for participation and the extent to which hypertensive patients wish to participate in the management of their condition. A cross sectional study with in-depth, face-to-face interviews was conducted with 49 hypertensive patients from one health centre. Interview themes were identified using content analysis. Characteristics predictive of participation desire were detected via quantitative analyses. Half of those interviewed were interested in participating in hypertension management. Those who had been hypertensive longer were less inclined to favour participation. Those with negative views of their 'disease' status and with higher blood pressure were more likely to want to participate. Patients needed further information and advice before decisions about future level of participation could be properly considered. PMID- 10865685 TI - Comparison of knowledge and psychological well-being between patients with a short disease duration (< or = 1 year) and patients with more established rheumatoid arthritis (> or = 10 years duration). AB - Patients with rheumatoid arthritis (RA) of short disease duration (i.e. < or = 1 year) compared with patients of longer disease duration (i.e. > or = 10 years) in terms of RA knowledge, symptoms of anxiety, symptoms of depression and disease acceptance. In addition, the predictors of psychological distress (i.e. symptoms of anxiety and depression) were examined. Data were collected by self administered questionnaires. As expected, patients with more established disease were significantly older and had more physical dysfunction. However, there were no statistically significant differences on anxiety, depression, acceptance of illness, pain or knowledge about RA. The need for education regarding RA and its implications was expressed by all participants regardless of disease duration. Illness acceptance beliefs were identified as significant predictors of both anxiety and depression. PMID- 10865686 TI - Psychosocial problems, coping strategies, and the need for information of parents of children with Prader-Willi syndrome and Angelman syndrome. AB - The aim of the present study was to identify the psychosocial problems of parents of a child with Prader-Willi syndrome or a child with Angelman syndrome. In addition, the strategies these parents apply to cope with these problems as well as their need for information are described. To assess these topics, parents filled in a self-report questionnaire. Both parent groups were found to have a high need for information, high feelings of loss of control, relatively high depressive feelings, particularly in mothers in both syndrome groups. Differences due to the type of syndrome were found on the fear factor. Parents of a child with Angelman syndrome had greater feelings of fear for the negative consequences for themselves, whereas parents of Prader-Willi children were more concerned about the consequences for the child. In general, coping strategies were not found to be different between the parent groups of children who had either type of syndrome. PMID- 10865687 TI - Information needs and preference for information of women with breast cancer over a first course of radiation therapy. AB - The study assessed the information needs of women receiving a course of radiation therapy (RT) for breast cancer and the relationship between information needs and preference for information. Thirty-three women were interviewed during the first (T1), third (T2), and last week of RT (T3), and one month later (T4) using the Toronto Informational Needs Questionnaire--Breast Cancer and the Information Subscale of the Health Opinion Survey. Information need scores were high and did not differ significantly across time. Information need and preference for information were correlated only at T1. Findings indicate that women with breast cancer receiving RT have high information needs and preference for information may not be a meaningful indicator of their desire for information. Health care providers need to assess the women's information needs frequently and be prepared to offer on-going educational support. PMID- 10865688 TI - Evaluation of a computerized contraceptive decision aid for adolescent patients. AB - Recent efforts to involve patients more actively in therapeutic decisions have suggested the relevance of computer-based interventions at clinic visits. This paper presents a longitudinal, experimental study evaluating a computer-based contraceptive decision aid in Chicago and Madison family planning clinic visits. Patient interviews at three time points evaluated patient acceptance by and program impact on 949 young women. Both Chicago and Madison patients reported high acceptance. The program resulted in improved short-term knowledge and confidence in oral contraceptive (OC) efficacy for Chicago and Madison patients. In addition, compared to their control group, Madison experimental group patients had higher OC knowledge 1 year after the initial visit and a trend for fewer pregnancies (P < 0.074). Compared to their control group, a higher percent of the Chicago experimental group patients adopted OC's after stating their intention to do so at the initial visit. Exposure to the computer program had no observable impact on the number of months on the oral contraceptive for Chicago or Madison patients. Overall findings suggest the usefulness of informatics tools as a supplement to patient-provider interactions. PMID- 10865689 TI - Computer-assisted instructions for patients with bronchial asthma. AB - We produced computer-assisted instruction (CAI) software for bronchial asthma patients (asthma educational system with computer-assisted instruction; ASTCAI) to assist in self-management and avoid asthmatic attacks and death. ASTCAI is a question-and-answer program operating in a multimedia environment, and was evaluated from questionnaires which 33 patients were asked. Thirty-two patients could perform ASTCAI without any assistance. The responses of 31 patients (94%) indicated that they had no difficulty with manipulation, and 29 patients (88%) stated that the program was beneficial to control of their asthma. Elderly patients (over 65) required more time than younger adults. Emergency visits or admissions of at least 1 year after the first CAI trial decreased in eight out of 26 patients, while only two patients deteriorated compared to the previous year. Our results show that CAI is feasible for most patients, and through active self learning CAI can improve motivation for self-management as well as supplement the physician's instructions. PMID- 10865690 TI - An evaluation of a health education intervention for mid-aged women: five year follow-up of effects upon knowledge, impact of menopause and health. AB - An evaluation of the long term impact of a health education intervention in primary care, for premenopausal women (45 years of age), is presented. The intervention included information and group discussion about menopause, stress management, health behaviours (smoking, exercise, diet) and treatment choices. Questionnaires were sent to 86 women who had been randomised into two groups (prepared/control) and were now aged 50 (response rate 91%). The prepared group had significantly greater knowledge of menopause and attributed fewer symptoms to the menopause than the controls. There were no group differences in measures of general health or mood, but there was a tendency for the prepared group to report more interest in sexual activity. Subjective evaluation of the intervention was positive in terms of increasing knowledge and helping women to deal with the emotional and practical aspects of the menopause. PMID- 10865691 TI - BSN on your terms! PMID- 10865692 TI - PCs can be hazardous to your health. PMID- 10865693 TI - Ethics in action. A patient who still needs very complex care is being prepared for discharge. PMID- 10865694 TI - Spinal implants. PMID- 10865695 TI - Oncology today: new horizons. Lung cancer. PMID- 10865696 TI - Are you ready for bloodless medicine? PMID- 10865697 TI - Incorporate now! PMID- 10865698 TI - Neuromuscular blockade. When and how. PMID- 10865699 TI - A new way to monitor paralyzing drugs. PMID- 10865700 TI - Hand care products. PMID- 10865701 TI - Keep your charting on course. PMID- 10865702 TI - Why is this patient starting to bruise? PMID- 10865703 TI - Body composition comes of age: a modest proposal for the next generation. The new reference man. PMID- 10865704 TI - Magnetic resonance imaging in human body composition research. From quantitative to qualitative tissue measurement. AB - Incremental improvements in our knowledge of human body composition are abetted by advances in research technology. Indeed, magnetic resonance imaging (MRI) represents a technological advance that has profoundly influenced body composition research. Routine applications of MRI include the measurement of whole-body and regional adipose tissue distribution, quantification of lean tissue and its principal constituent skeletal muscle, and the measurement of visceral adipose tissue. MRI is now the method of choice for calibration of field methods designed to measure body fat and skeletal muscle in vivo. Common to these applications is the measurement of tissue quantity. More recently proton (1H) and sodium (23Na) MRI protocols have been developed that measure the quality (lipid and sodium concentration) of skeletal muscle tissue. These unique applications of MRI represent a major advance in the study of altered muscle composition in vivo, with numerous applications in both applied and clinical medicine. In this review we provide a brief overview of routine applications of MRI in body composition research, followed by a focus on more recent applications of MRI that employ fast imaging sequences for qualitative measurement of human skeletal muscle. PMID- 10865705 TI - Composition of skeletal muscle evaluated with computed tomography. AB - Computed tomography (CT) can yield quantitative imaging data from detailed maps of linear attenuation coefficients within tissue. The attenuation characteristics of skeletal muscle and adipose tissue can be quantified in vivo to provide information about the composition of skeletal muscle and the distribution of adipose tissue within muscle. Several studies have taken advantage of this utility to quantify skeletal muscle composition and fatty infiltration of muscle, in particular to quantify the attenuation characteristics of muscle as a marker of its lipid content. In this manner we found that the mean muscle attenuation of skeletal muscle reflects an increase in its fat content in obesity, and that this regional body composition parameter is strongly related to insulin-resistant glucose metabolism. In addition, muscle composition and adipose tissue distribution within muscle may be altered with clinical weight-loss interventions. CT may also provide important information about the changes in muscle mass and composition with aging and disease, which may, in turn, affect the muscle's function. In summary, CT can provide important quantitative data on the composition of muscle, and the distribution of adipose tissue within it, and this may be important in examining the relationships among skeletal muscle metabolism, lipid accumulation within muscle, and muscle function. PMID- 10865706 TI - Observation of intramyocellular lipids by 1H-magnetic resonance spectroscopy. AB - Magnetic resonance (MR) methods are increasingly being used to investigate the physiology of human muscle. Although MR imaging (MRI) reveals the morphology of muscles in great detail, for example, for determining their volume and fiber orientation, MR spectroscopy (MRS) provides information on the chemical composition of the tissue. Depending on the observed nucleus, MRS allows the observation of high-energy phosphates (31P-MRS), glycogen (13C-MRS), or intramyocellular lipids (1H-MRS), to give only a few examples. 1H-MRS of human skeletal muscle requires special techniques because 1H nuclei in water or adipose tissue are far more concentrated than in any other metabolite of human tissue. The strong signal from water can be suppressed by special prepulses, whereas large signals from fat in adipose tissue can be reduced by carefully selecting the region of interest. Until recently, it was presumed that only a few metabolites would be visible underneath the large resonances of water and subcutaneous fat. Meanwhile, it was clear that 1H-MR spectra of human muscle reveal much metabolic and structural information. The determination of intramyocellular lipids (IMCL) by 1H-MRS was initiated by the observation of two compartments of triacylglycerols with a resonance-frequency shift of approximately 0.2 ppm. The two resonances can be attributed to CH2 protons of lipids in fat cells, and to lipids inside muscle cells (IMCL). 1H-MRS examinations are noninvasive and, therefore, can be repeated many times and with a high temporal resolution. MRS has the potential to replace biopsy to follow-up IMCL levels; however, biopsy still has the advantage that other methods, such as molecular biology, can be applied to the sample. It can be shown that IMCL levels (expressed in mMol/kg wet weight and volume %) are muscle specific and vary with diet and physical activity. In addition, it has been reported that IMCL levels are correlated with insulin sensitivity. A comparison of different methods for assessing IMCL levels, including MRS, chemical analysis, and morphometry, revealed a satisfactory correlation among them and a superior correlation of MRS with the average of the three methods. The observation of IMCL levels by means of 1H-MRS is extremely promising, but several methodological limitations and pitfalls need to be considered. PMID- 10865707 TI - In vivo determination of body composition of rats using magnetic resonance imaging. AB - Magnetic resonance imaging (MRI) has potential as an instrument to measure body composition because it can discriminate various soft tissues in vivo. These soft tissues include adipose tissue, muscle, organs, and brain. We report on preliminary studies using a 4.2-tesla MRI for measuring body composition in the mouse and rat. We employed image segmentation methods that include an image correction method, a necessary requirement when the images are taken in the presence of nonuniform radio-frequency (RF) coil response. The software for 3-D data segmentation, quantification, correction, image manipulation, and visualization has been developed as a research tool. This method currently is being validated. PMID- 10865708 TI - Study of the isolated perfused rat heart exposed to ischemia using 31P NMR surface coil. PMID- 10865709 TI - Assessing body composition and changes in body composition. Another look at dual energy X-ray absorptiometry. AB - Dual-energy X-ray absorptiometry (DXA) is selected with increasing frequency as a method for both assessing body composition and measuring the changes in body composition. Issues have been raised about hydration, software version, hardware (fan beam vs. pencil beam), and the subject population in relation to the validity of DXA-derived estimates of body composition. This paper reviews validation studies of DXA to assess the impact of recent developments in its technology. Studies by Prior et al., Kohrt et al., Salamone et al., Going et al., and Pietrobelli et al. demonstrate the effectiveness of DXA estimates of changes in body composition. By contrast, Clasey et al., Nelson et al., and Friedl et al. found limitations in DXA estimates of body composition and its changes. These contradictory conclusions were explored for threats to internal validity in each research study. From this analysis, two validation guidelines are recommended for use when evaluating estimates of body composition. When multicomponent models are used, it is essential that estimates of body water as a fraction of fat-free mass fall in the expected range (71 to 75%) and have a relatively small standard deviation (2 to 3%). For measuring changes in body composition, DXA estimates of total body mass must accurately reflect both baseline and posttreatment scale body weight estimates. Failure to meet these guidelines threatens the internal validity of the study and raises the likelihood of methodological discrepancies. Applying these criteria to DXA studies of body composition under review accounts for much of the contradictory conclusions among investigations. PMID- 10865710 TI - Total body dual X-ray absorptiometry is a good measure of both fat mass and fat free mass in liver cirrhosis compared to "gold-standard" techniques. Melbourne Liver Group. AB - Liver cirrhosis is a condition in which overnutrition, edema, and undernutrition can coexist simultaneously, or successively, over a period of time, giving rise to alterations in body composition, as well as systemic and multiorgan manifestations. We undertook a cross-sectional study of body composition in 198 adult patients with liver cirrhosis (140 males, mean age 53.6, range 31-85 years; and 58 females, mean age 58.4, range 36-79 years). The patients had cirrhosis of differing etiology and different stages of severity. They were gathered from seven different hospital clinics in the city of Melbourne, Australia, but all the body composition measurements were performed in one body composition laboratory. A variety of body composition techniques were used to identify which commonly available ones could best assess both fat-free mass and fat mass relative to a criterion "gold-standard" method available in a specialist laboratory. A gold standard fat-free mass (FFMGS) was defined as the sum of total body protein, measured by in vivo neutron activation analysis (IVNA), plus total body water, measured by D2O dilution, plus bone mineral content, measured by dual X-ray absorptiometry (DXA). A gold-standard fat mass (FATGS) was defined as the difference between body weight and FFMGS. "Usual" fat mass and fat-free mass were defined by different techniques including DXA, anthropometry (ANT), single frequency bioelectrical impedance (SFBIA), multiple-frequency bioelectrical impedance spectroscopy (MFBIA), and whole body gamma counting (TBK). The FFMGS was overhydrated in both sexes, relative to the usual value of 0.73, but women were significantly overhydrated compared to men. Relative to the gold-standard deuterium oxide dilution method for measuring total body water, SFBIA slightly overestimated TBW, whereas MFBIA slightly underestimated TBW, with both methods having wide limits of agreement for any single estimate. In comparing FFM to FFMGS, only DXA showed a small negative bias, in both males and females, with modest limits of agreement for any single estimate. All other methods showed a large negative bias (ANT, SFBIA, and MFBIA) or a large positive bias (TBK) relative to FFMGS, with wide limits of agreement. In comparing FAT with the FATGS, only DXA showed a small positive bias, in both males and females, with modest limits of agreement for any single estimate. All other methods showed a large positive bias (ANT, SFBIA, and MFBIA) or a large negative bias (TBK) relative to FATGS, with wide limits of agreement. In cirrhosis, DXA is a good and widely available method to assess both fat mass and fat-free mass. However, it cannot give information about the quality of the FFM, particularly its water content. The bedside methods of anthropometry and bioelectrical impedance, both SFBIA and MFBIA, are poor methods of measuring body composition in patients with liver cirrhosis, whereas whole body gamma counting, although not widely available, also significantly differs from the gold-standard method of assessment of fat-free mass and fat mass in liver cirrhosis. PMID- 10865711 TI - Comparisons between Hologic QDR 1000W, QDR 4500A, and Lunar Expert dual-energy X ray absorptiometry scanners used for measuring total body bone and soft tissue. AB - Measurements of total-body bone and soft tissue were compared between two fan beam dual-energy X-ray absorptiometry (DXA) scanners, a Hologic QDR 4500A, a Lunar Expert, and a pencil-beam Hologic QDR 1000W. Phantom studies showed that mass measurements were not compromised by magnification effects, but that the height of a bone within the body affected the measured bone mineral content (BMC) and, to a lesser extent, the bone mineral density (BMD). There were differences in calibration for fat proportion between the three instruments. Comparisons on volunteers demonstrated very high correlations of bone and soft tissue measurements, but regression coefficients differed from unity, and intercepts were significant. With all three scanners, wrapping lard around the limbs of a volunteer, to simulate weight change, changed the apparent BMC and BMD. PMID- 10865712 TI - Comparison of air-displacement plethysmography, hydrodensitometry, and dual X-ray absorptiometry for assessing body composition of children 10 to 18 years of age. AB - Body density (Db) of 54 boys and girls 10-18 years of age (13.9 +/- 2.4 years) was measured in an air-displacement plethysmograph, the BOD POD, and compared to Db determined by hydrodensitometry (HW). Both Db values were converted to percent body fat (%BF) using a two-component model conversion formula and compared to %BF determined by dual energy X-ray absorptiometry (DXA). Body density estimated from the BOD POD (1.04657 +/- 0.01825 g/cc) was significantly higher than that estimated from HW (1.04032 +/- 0.01872 g/cc). The relative body fat calculated from the BOD POD (23.12 +/- 8.39 %BF) was highly correlated but, on average, 2.9% BF lower than %BF DXA. Average %BF estimates from HW and DXA were not significantly different. Despite consistently underestimating the %BF of children, the strong relationship between DXA and the BOD POD suggests that further investigation may improve the accuracy of the BOD POD for assessing body composition in children. PMID- 10865713 TI - Measurements of body composition by dual-energy X-ray absorptiometry improve prediction of energy expenditure. AB - The prediction of energy expenditure by dual-energy X-ray absorptiometry (DXA) and bioimpedance analysis (BIA) was assessed in 35 healthy individuals of both sexes, with a mean body mass index (BMI) of 23.8 kg/m2 (range 18-33.8), and mean age of 30 years (22-40). Energy expenditure (EE) was measured under standard conditions in a respiration chamber, the total and regional body composition by DXA, and total body composition by BIA. When body composition was measured by BIA, 88.5% of the variation in 24-h EE was explained by lean body mass (LBM); this figure was increased by DXA, where total lean tissue mass (LTM) and total fat tissue mass (FTM) could account for 91.5% of the variation. Also, the prediction of resting energy expenditure (REE) was improved by DXA, from 88.1% to 89.8% (LBM vs. LTM, FTM). Measurements of regional body composition showed that trunk LTM was significantly superior as a predictor, especially of REE and sleeping EE (EE sleep), compared to the peripheral LTM; thus, the predictions of REE were 83% vs. 87% (peripheral vs. trunk), respectively; and the predictions of EE sleep were 83% vs. 89% (peripheral vs. trunk), respectively. Therefore, body composition measurements by DXA improved the prediction of EE. Trunk LTM was a superior predictor, especially of REE and EE sleep, compared to peripheral LTM. In conclusion, the present results suggest that measuring total and regional body composition by DXA can somewhat improve the prediction of EE. PMID- 10865714 TI - Dual-energy X-ray absorptiometry measurements of the body composition of pigs of 90- to 130-kilograms body weight. AB - We used the pig as an in vivo model for evaluating the effects of extreme body depth on the measurement of body composition by dual-energy X-ray absorptiometry (DXA). One group of 17 pigs weighed an average of 90 kg and had a maximum body depth of approximately 30 cm; another group of 54 pigs weighed 123 kg on average and had a maximum body depth of about 35 centimeters. In the larger pigs, DXA tended to measure a higher percentage of total body fat relative to the chemical analysis than in the smaller pigs. In both groups, but more predominantly in the larger pigs, there were areas in the bone of extreme thickness and/or density that were excluded from the analysis, which caused underestimation of bone mineral content and total tissue mass. PMID- 10865715 TI - Measurement of changes in soft tissue mass and fat mass with weight change: pencil- versus fan-beam dual-energy X-ray absorptiometry. Health ABC Study. PMID- 10865716 TI - Accuracy of dual-energy X-ray absorptiometry in infant scans. An assessment using anthropomorphic phantoms. PMID- 10865717 TI - Validation of estimates of body composition by dual-energy X-ray absorptiometry in fluid overload conditions. PMID- 10865718 TI - Validation of estimates of lean tissue mass made by dual-energy X-ray absorptiometry. PMID- 10865719 TI - Comparison of two phantoms for body composition with dual-energy X-ray absorptiometry. PMID- 10865721 TI - A new measuring device for quantifying the amount of mineral in the heel bone. PMID- 10865720 TI - K-40 and dual-energy X-ray absorptiometry estimates of lean weight compared. Normals and patients with neuromuscular disease. PMID- 10865722 TI - Evaluation of a new DXA fan-beam instrument for measuring body composition. PMID- 10865723 TI - Underestimation of fat-free mass in women, but not men, by dual-energy X-ray absorptiometry. Comparison with total body potassium and bioelectrical impedance analysis. PMID- 10865724 TI - Of mermaids and mountains. Three decades of prompt activation in vivo. AB - During 1966 to 1972, several laboratories demonstrated the feasibility of measuring the major body elements H, N, Ca, and Cl by prompt gamma in vivo neutron activation analysis (PGIVNA). The MERMAID facility in Birmingham, England used a cyclotron-produced pulsed neutron beam, but other groups in the United Kingdom, United States, Canada, and New Zealand subsequently developed systems based on radioisotope neutron sources that could measure body nitrogen with a precision of a margin of error of a few percentage points. The accuracy of N measurement was greatly enhanced by Vartsky's internal standardization, using prompt-gamma H as the marker and total body hydrogen (based on total body water and skinfolds) as the reference. Chlorine and extracellular water volume were used in a similar way by the Swansea group to calibrate the prompt-gamma analysis of total body calcium. The PGIVNA technique is most valuable in assessing nutritional status, particularly in relation to body protein. PMID- 10865725 TI - Assessment of background hydrogen by the Monte Carlo computer code MCNP-4A during measurements of total body nitrogen. AB - The use of a hydrogen internal standard to enable the estimation of absolute mass during measurement of total body nitrogen by in vivo neutron activation is an established technique. Central to the technique is a determination of the H prompt gamma ray counts arising from the subject. In practice, interference counts from other sources--e.g., neutron shielding--are included. This study reports use of the Monte Carlo computer code, MCNP-4A, to investigate the interference counts arising from shielding both with and without a phantom containing a urea solution. Over a range of phantom size (depth 5 to 30 cm, width 20 to 40 cm), the counts arising from shielding increased by between 4% and 32% compared with the counts without a phantom. For any given depth, the counts increased approximately linearly with width. For any given width, there was little increase for depths exceeding 15 centimeters. The shielding counts comprised between 15% and 26% of those arising from the urea phantom. These results, although specific to the Swansea apparatus, suggest that extraneous hydrogen counts can be considerable and depend strongly on the subject's size. PMID- 10865726 TI - In vivo elemental partition analysis using fast neutrons. A tool for testing the efficacy of new clinical interventions. AB - The nutritional status of patients can be evaluated by monitoring changes in body composition, including depletion of protein and muscle; distribution of adipose tissue; and changes in hydration status, and bone or cell mass. Fast neutron activation (for N and P) and neutron inelastic scattering (for C and O) are used to assess in vivo elements characteristic of specific body compartments. Nonbone phosphorus for muscle is measured by the 31P(n,alpha)28Al reaction, and nitrogen for protein via the (n,2n) fast neutron reaction. The carbon-to-oxygen (C/O) ratio is used to measure distribution of fat and lean tissue in the body and to monitor small changes in lean mass and its quality. In addition to evaluating the efficacy of new treatments, the method is used to study the mechanisms of depletion of lean tissue with aging. PMID- 10865727 TI - Calibration of the Brookhaven National Laboratory delayed gamma neutron activation facility to measure total body calcium. AB - Differences in body size and shape can cause large variances in the in vivo results of neutron activation analysis. To introduce corrections for body size for the delayed gamma neutron activation facility at Brookhaven National Laboratory, "reference man"-sized and "reference woman"-sized phantoms were constructed. Simulation results using the Monte Carlo Neutron and Photon Transport code also provided correction factors for people of different sizes. For individuals with a body mass index (BMI = weight (kg)/height (m)2) between 20 and 30, no correction was required. At BMIs greater than 30, the effects of neutron attenuation were significant and a correction factor of CF = -0.0192 x BMI + 1.5635 can be applied. PMID- 10865728 TI - Estimation of extracellular water by instrumental neutron activation analysis of bromine. PMID- 10865729 TI - Assessing regional muscle mass with segmental measurements of bioelectrical impedance in obese women during weight loss. AB - Tetrapolar bioelectrical impedance analysis (BIA) offers the possibility of determining the bioconductor volume in discrete segments of the body, because the resistivities of bone, fat, and skeletal muscle differ considerably. We tested this hypothesis by measuring BIA and anthropometry of defined segments of the right thighs of women before and during a controlled weight-loss program. Eight women, aged 22 to 32 years, with a body mass index of 37.8 +/- 1.6 (mean +/- SE) kg/m2 underwent determinations of body composition with dual-energy X-ray absorptiometry (DXA) and regional BIA measurements (800 microA at 50 kHz) before the program, and monthly thereafter for four months during weight loss. BIA measurements were made with spot-detector electrodes positioned 10 cm apart on the anterior of the thigh, and source electrodes placed on the right hand and foot. The physical volume of the thigh segment decreased by 29 +/- 3% (p < 0.0001), with a modest change in its electrical volume (8 +/- 0.2%; p < 0.05) during weight loss. Muscle (181 +/- 49 g; p < 0.05) and fat mass (702 +/- 95 g; p < 0.001) also declined. The electrical or bioconductor volume correlated with DXA determinations of muscle mass (r = 0.91, p < 0.0001), whereas physical volume correlated with fat mass (r = 0.95, p < 0.0001). These findings support the hypothesis that BIA is a valid method to assess regional muscle mass in humans. PMID- 10865730 TI - Bioelectrical impedance analysis. What does it measure? AB - Bioelectrical impedance analysis (BIA) has been proposed for measuring fat-free mass, total body water, percent fat, body cell mass, intracellular water, and extracellular water: a veritable laboratory in a box. Although it is unlikely that BIA is quite this versatile, correlations have been demonstrated between BIA and all of these body compartments. At the same time, it is known that all of the compartments are correlated among themselves. Because of this, it is difficult to determine whether BIA is specific for any or all of these compartments. To investigate this question, we induced acute changes in total body water and its compartments over a 3-h period. Using this approach, we demonstrated that multifrequency BIA, using the Cole-Cole model to calculate the zero frequency and infinite frequency resistance, measures extracellular and intracellular water. PMID- 10865731 TI - Hydrational status assessed by bioelectrical impedance spectroscopy and dilution methods in patients with classical dengue fever. AB - The effects of an episode of acute classical dengue fever on extracellular water (ECW), intracellular water (ICW), and total body water (TBW) were measured in nine patients using conventional dilution techniques; and the findings were compared with the outcome variables from whole body impedance spectroscopy (BIS), extracellular fluid resistance (Recf), and intracellular fluid resistance (Ricf). The patients were assessed on admission with febrile presentation (DI), at discharge after the defervescence of the fever cycle at about five days postadmission (DII), and seven days thereafter (DIII). As a reference group, 15 persons without acute or chronic illness were enrolled. Total body water was unaltered during the course of disease and was not different from that in normal healthy subjects. However, body water shifted from the intracellular to the extracellular compartment in patients from the acute phase to convalescence, as reflected in the ratios of ECW/TBW and ECW/ICW. These ratios were significantly higher in convalescent dengue patients (DIII) than in the reference group (p < 0.05). Increasing ECW, from the acute phase of the disease to convalescence, was associated with a significant decrease in Recf (719 +/- 95, 693 +/- 89, 643 +/- 81 omega; p < 0.0001) and in Recf/Ricf (p < 0.01). Recf and Recf/Ricf were higher in the acute phase (DI) of dengue fever compared to controls (p < 0.05). We conclude that dengue fever is characterized by a relative expansion of ECW during the course of disease and convalescence. BIS was sensitive in determining the hydrational profile in dengue fever patients. PMID- 10865733 TI - Comparison of whole body and segmental bioimpedance methodologies for estimating total body water. AB - We compared the whole body (WB) and segmental bioelectrical impedance analysis (BIA) methodologies in a group of healthy adults (n = 25). It has been suggested that the segmental methodology may overcome the difficulty in generating a single algorithm to predict total body water (TBW) in all groups whether healthy or not. We measured TBW, using D2O dilution, and WB and segmental BIA parameters. Cole Cole analysis was used to determine the impedance at the characteristic frequency (Zc). The correlation between TBW (by D2O dilution) and segmental BIA measures (multiple regression, r = 0.90, p < 0.001, SEE = 3.1 L) was not significantly higher than the correlation between TBW (D2O dilution) and WB BIA measures (simple regression, r = 0.85, p < 0.001, SEE = 3.6 L). Others have observed this "lack of improvement" in a group of healthy subjects. The true value of the segmental BIA methodology may lie in applications involving groups with altered distributions of segmental and compartmental fluid. PMID- 10865732 TI - Using localized impedance measurements to study muscle changes in injury and disease. AB - We measured localized impedance on the surface of the thigh to determine the properties of underlying muscle; these include rho 1, the resistivity for current flowing parallel to the fibers, and theta avg, the average phase along the thigh (normalized to a standard length). The results for a modest sampling of nominally healthy subjects show that the theta avg values are substantially higher than the whole body phases encountered in standard bioimpedance analysis. When the sample is augmented to include subjects undergoing hemodialysis and/or recovering from serious illness or leg injury, the behavior of the position dependence of the phase theta (z) and the rho 1 vs. theta avg average distribution both strongly indicate a correlation between very low phase angles and injury or disease. Furthermore, measurements on a subject in a weight training program after injury provide evidence of a monotone increase in rho 1 with increasing strength. Measurements on dialysis patients during treatment show a nonlinear response of thigh muscle to the degree of fluid removal and wide disparities between individuals. PMID- 10865734 TI - Relationships between bioelectric resistance and somatotype in 9- to 11-year-old children. PMID- 10865735 TI - Use of bioimpedance spectroscopy to assess effects of perioperative treatment with growth hormone on fluid changes in patients undergoing major surgery. PMID- 10865736 TI - A solution with an elemental composition of reference man for calibrating in vivo neutron-activation analysis scanners. PMID- 10865737 TI - Is there advantage in using multiple frequency bioelectrical impedance analysis to predict gentamicin distribution volume in neonates? PMID- 10865738 TI - Association between ethnicity, body mass index, and bioelectrical impedance. Implications for the population specificity of prediction equations. PMID- 10865739 TI - Estimation of body composition by multifrequency bioimpedance measurement in children. PMID- 10865740 TI - Bioelectrical impedance spectroscopy in health and disease. Correspondence between whole body and segmental bioelectrical impedance spectroscopy indices in patients with classical dengue fever. AB - Bioelectrical impedance spectroscopy (BIS) can provide estimates of body composition for both whole body (WB) and body segments (BS). In normal, healthy subjects, BS measurements may be expected to serve as surrogates for WB indices; however, very little is known about this correspondence in people suffering from acute illnesses. The aim of this study was to evaluate the degree of this correspondence in patients with an acute, systemic illness, such as classical dengue fever. Ten adult patients were examined upon admission to the community hospital on the Pacific Coast of Guatemala and after clinical recovery about two weeks later, and compared with a group of healthy subjects living in the same region. BIS was measured with a Xitron 4000B analyzer (Xitron Technologies Inc., San Diego, CA, USA). BS measurements were carried out using Organ et al.'s approach. The BIS data were modeled with the manufacturer's software: extra- (Recf) and intracellular- (Ricf) resistances, and the Recf/Ricf ratio. BIS BS measurements correlate closely with WB in both the acute and the recovery stages of dengue fever, with the leg showing the highest degree of correspondence and the trunk the lowest. Recf indices, per se, generally showed higher correspondence than Ricf or the Recf/Ricf ratio. PMID- 10865742 TI - Low-impedance localized measurements using standard bioelectrical impedance analysis instruments. PMID- 10865741 TI - Bioelectrical impedance spectroscopy and body composition. PMID- 10865743 TI - Appendicular lean body mass. Prediction by bioelectrical impedance analysis. PMID- 10865744 TI - The effect of electrode placement in measuring ipsilateral/contralateral segmental bioelectrical impedance. PMID- 10865745 TI - Bioelectrical impedance analysis prediction equations differ between African Americans and Caucasians, but it is not clear why. PMID- 10865746 TI - A new theoretical model for predicting bioelectrical impedance analysis. PMID- 10865747 TI - Gamma resonance absorption. New approach in human body composition studies. AB - The main stream of body elemental analysis is based on the delayed, prompt, and inelastic neutron interactions with the main elements found in the human body, and subsequent analysis of the measured delayed or prompt gamma ray spectra. This methodology traditionally was, and still is, applied for whole body analysis and requires relatively high radiation doses. A new method, based on gamma nuclear resonance absorption (GNRA), is being established at Brookhaven National Laboratory as part of its body composition program. The method is element specific with a high tomographic spatial-resolution capability, at a small fraction of the radiation dose used in the current system. The new system, with its components and capabilities, is described below. PMID- 10865748 TI - Gamma ray nuclear resonance absorption: an alternative method for in vivo body composition studies. AB - We have evaluated gamma ray nuclear resonance absorption (gamma-NRA) on nitrogen, a mature technology proposed and developed by Soreq NRC for detecting explosives, as an alternative to neutron activation for in vivo assaying of body nitrogen. The principles of the gamma-NRA method are outlined, and a test facility constructed at McMaster University's Accelerator Laboratory is described. The results of a feasibility study recently performed there on phantoms and animal tissue are presented and discussed. gamma-NRA is a full imaging technique that essentially constitutes element-specific absorptiometry--i.e., it can generate projections of the mass distribution for a specific element, along with a conventional radiograph of the patient. From the transmission profile of an individual scanned by 9.17 MeV gamma rays, local or whole body nitrogen content can be determined via the resonant attenuation undergone when the beam encounters regions of nitrogen concentration. The advantages of gamma-NRA over neutron activation are (a) radiation doses delivered to the body are at least one order of magnitude lower, thus allowing repeated measurements on individual patients and also rendering the method ethically acceptable for application to children; (b) gamma-NRA is inherently free from uncertainties related to nonuniform distributions of the element in question within the body; (c) it is applicable to patients of varying size and shape; and (d) it yields both nitrogen images and conventional radiographic images of the body. PMID- 10865749 TI - Assessment of body volume using three-dimensional photonic scanning. AB - Measurement of body volume (BV) can be used to estimate body composition using two- or multicomponent models. The traditional approach, underwater weighing (UWW), is awkward and unsuitable for many subjects. A newer alternative, whole body air displacement plethysmography (ADP), is less demanding but still unsuitable for young children, who may not remain still during the measurement. We have, therefore, considered whether a novel approach, three-dimensional photonic scanning, is a viable alternative. Duplicate measurements of body volume were obtained in 22 adults (11 of each sex; mean [SD] BMI, 21.8 [2.5] kg/m2) by UWW, ADP, and a Hamamatsu Bodyline Scanner (HBS) (Hamamatsu, Japan). Subjects wore a tight-fitting swimming costume for all three measurements, which were performed within one day of each other. Scans lasted 10 seconds, with the subject standing in a predefined position. The body surface skin was reconstructed using a B-spline-fitting model. In UWW, lung volume (LV) was measured simultaneously with underwater weight. In ADP and HBS, LV was predicted from weight and height. Results were compared using correlation and Bland and Altman analysis. Correlation analysis indicated that the scanner successfully ranked subjects in terms of BV. However, Bland and Altman analysis demonstrated that, relative to both UWW and ADP, HBS measured BV without bias but with limits of agreement between individuals of > 2 liters, equivalent to approximately 20% fat. Scan precision was 0.57 liter, or 4.1% fat. Although HBS cannot yet measure BV with sufficient accuracy to predict fatness, much of the error is probably due to difficulties in standardizing LV during the scan. Simultaneous measurement of LV with volume by HBS is expected to improve limits of agreement substantially. Occlusion is also an important source of error. The method offers many advantages over alternative techniques, because the measurement is brief, noninvasive, and suitable for repeat measurements. PMID- 10865750 TI - Feasibility of a fluorescent X-ray source for in vivo X-ray fluorescence measurements of kidney and liver cadmium. PMID- 10865751 TI - Total body capacitance correlates with total body potassium. PMID- 10865752 TI - In vivo measurement of platinum in the kidneys using X-ray fluorescence. AB - A noninvasive in vivo method has been developed and optimized for measuring platinum concentrations in the kidneys of patients receiving chemotherapy. The method is based on polarizing the X-ray beam from an orthovoltage radiotherapy treatment unit, the Pantak DXT-300, and using the beam to produce emission of the characteristic platinum X-rays from the kidney. The platinum is derived from platinum-based chemotherapy drugs, such as cisplatin and its analogues (carboplatin and iproplatin), used to treat cancer patients. The clinical motivation for measuring the platinum concentration in both the kidneys and the tumor is to optimize the treatment by establishing the relationships between the accumulation of the drug at those sites. Such clinical information could be valuable in maximizing the therapeutic ratio toward the tumor tissue and limiting the hazards to the kidney. The performance of the system was experimentally optimized with respect to the applied X-ray tube voltage, filter material, and polarizer. Additionally, the MCNP-4B Monte-Carlo computer code was used to estimate the optimum shielding materials, the thickness surrounding the X-ray tube, and the arrangement of collimators, to protect patients from the hazards of the scattering radiation. Clinical measurements can be made with a combination of a bilayer of copper and silicon as polarizer, a 0.25-mm tin filter introduced in the path between the X-ray beam and the polarizer, and an operating voltage of 220 kV. The minimum detection limit achieved with this arrangement is 16 ppm, for a kidney depth of 3 x 3 cm, with a skin dose of 1.6 mGy and measurement time of 2,000 seconds. PMID- 10865753 TI - Perquisite spin-off from twenty-two years of measuring background in the whole body counter steel room. PMID- 10865754 TI - Response function of the BGO and NaI(T1) detectors using Monte Carlo simulations. PMID- 10865755 TI - The assessment of stature using an infrared technique. PMID- 10865756 TI - Lead accumulation in highly exposed smelter workers. PMID- 10865757 TI - Bone hydroxyapatite/collagen ratio. In vivo measurements by X-ray absorptiometry. PMID- 10865758 TI - Plasma sample preparation by ultrafiltration for total body water determination. PMID- 10865759 TI - Body composition modeling. Application to exploration of the resting energy expenditure fat-free mass relationship. AB - There are now many published methods for predicting resting energy expenditure (REE) from measured body mass and composition. Although these published reports extend back almost a century, new related studies appear on a regular basis. It remains unclear what the similarities and differences are between these many methods and what, if any, advantages the newly introduced REE prediction models offer. These issues led us to develop an organizational system for REE prediction methods with the ultimate aim of clarifying prevailing ambiguities in the field. Our classification scheme is founded on the mathematical function type (descriptive and mechanistic) and body composition level (whole body-->molecular) used in REE prediction model development. The model is applied in an exploration of the well-established empirical relationship between REE and fat-free body mass (FFM). The developed relationships indicate that REE vs. FFM is a curvilinear relationship in mammals as a whole, that the relationship can be described as a linear function in humans, and that the simple linear regression line coefficients can be reconstructed from established tissue-system level component relationships. Our classification system, the first founded on a conceptual basis, highlights similarities and differences between the many diverse REE body composition prediction methods, provides a framework for teaching REE-body composition relationships to students, and suggests important future research opportunities. PMID- 10865760 TI - Modeling leg sections by bioelectrical impedance analysis, dual-energy X-ray absorptiometry, and anthropometry: assessing segmental muscle volume using magnetic resonance imaging as a reference. AB - This study aimed to assess the value of different DXA and BIA models for predicting muscle volume in mid-thigh segments obtained by MRI. Three DXA models were used: in model A, muscle was taken to be equivalent to fat-free soft tissue; in model B the thigh segment was divided into its constituent tissues using fixed assumptions about tissue composition; in model C the assumptions were similar to model B, but with variable distribution of fat and fat-free soft tissue, depending on body mass index. The two BIA models (both parallel tissue resistance models) involved impedance measurements at 50 kHz, and assumptions about either the specific resistivities of all the constituent tissues (model A), or resistivities of only adipose tissue and muscle (model B). Anthropometric estimates (thigh circumference and skinfold thickness) assumed that both limb and muscle circumference were circular. Compared to MRI estimates of muscle mass, those obtained by DXA model A (fat-free soft tissue) were not as good as those obtained using models B and C, although the standard deviations of the differences were similar with all three models. The BIA models were superior to the anthropometric estimates of muscle volume (relative to MRI) with respect to bias, but the standard deviations of the differences were large for both. The intraobserver repeatabilities for muscle volume were < 0.5% for MRI, < 1% for DXA, 1.8% for BIA, and 1.7% for anthropometry (interobserver value for BIA was 3.8% and for anthropometry 3.5%). The study suggests that DXA modeling provides a promising approach for assessing muscle mass in thigh segments, and suggests the potential value of parallel BIA models for groups of individuals but not for individual subjects, possibly because muscle resistivity is influenced not only by its composition but also by the direction of current flow in muscle. PMID- 10865761 TI - Cellular-level body composition model. A new approach to studying fat-free mass hydration. AB - Fat-free mass hydration, the ratio of total body water (TBW) to fat-free mass (FFM), is stable at approximately 0.73 in mammals, and this constancy provides a means of estimating total body fat in vivo. As there is no mechanistic theory to explain the magnitude and variability in TBW/FFM, the present investigation describes a cellular-level model indicating that FFM hydration is determined by four factors: body cell mass hydration, extracellular fluid hydration, ratio of extracellular solids to TBW, and ratio of extracellular water to intracellular water. According to the model, TBW/FFM can be predicted for adult humans as a mean of 0.73, and a variation range 0.69-0.77. The suggested modeling approach provides a conceptual framework for TBW-fat estimation method and identifies important areas that remain to be studied. PMID- 10865762 TI - Epidemiological applications of body composition. The effects and adjustment of measurement errors. AB - The amounts of fat and fat-free mass (FFM) are functions of lifestyle, diseases, increasing age, and genetics. The levels of these body compartments are established risk factors for cardiovascular and related chronic diseases. Body composition can be assessed by several methods. The measurements for each method have inherent variations, which can be due to error in the measurement or biological variation. This presentation focuses on the reliability and precision of body composition measurements and the impact of these errors on epidemiological and population-based studies. The effects of these errors in applying body composition to epidemiological studies include the following: (1) the association of cardiovascular risk factors with body composition in cross sectional studies; (2) changes in body composition in longitudinal or intervention studies; (3) association of changes in body composition with cardiovascular risk factors; and (4) comparison of levels of body composition among subgroups in cross-sectional studies and in intervention studies. PMID- 10865764 TI - Using simple anthropometric parameters to develop formulas for estimating weight and height in Chinese adults. PMID- 10865763 TI - Anthropometry in body composition. An overview. AB - Anthropometry is a simple reliable method for quantifying body size and proportions by measuring body length, width, circumference (C), and skinfold thickness (SF). More than 19 sites for SF, 17 for C, 11 for width, and 9 for length have been included in equations to predict body fat percent with a standard error of estimate (SEE) range of +/- 3% to +/- 11% of the mean of the criterion measurement. Recent studies indicate that not only total body fat, but also regional fat and skeletal muscle, can be predicted from anthropometrics. Our Rosetta database supports the thesis that sex, age, ethnicity, and site influence anthropometric predictions; the prediction reliabilities are consistently higher for Whites than for other ethnic groups, and also by axial than by peripheral sites (biceps and calf). The reliability of anthropometrics depends on standardizing the caliper and site of measurement, and upon the measuring skill of the anthropometrist. A reproducibility of +/- 2% for C and +/- 10% for SF measurements usually is required to certify the anthropometrist. PMID- 10865765 TI - The best predictive model for estimating fat-free mass. PMID- 10865766 TI - A comparison of body composition techniques. PMID- 10865767 TI - Skinfolds versus bioimpedance analysis for predicting fat-free mass. PMID- 10865768 TI - The "B" in the Selinger Four-Compartment body composition formula should be body mineral instead of bone mineral. PMID- 10865769 TI - Total body protein in chronic diseases and in aging. AB - Substantial losses of total body protein (TBP) can occur in chronic diseases and in aging. Such losses impact negatively on immunity and quality of life, and on growth rates in children. Direct measurements of total body nitrogen (TBN) monitor the integrated changes in TBP over time and allow comparison with normal subjects. TBN assessment via neutron capture analysis is therefore the gold standard method of TBP estimation, so that risk factors for protein deficit can be identified and patient management optimized. The nitrogen index (NI) can be used to predict prognostic outcome: an NI < 0.9 is associated with substantial wasting in HIV disease, an NI < 0.8 predicts significant pathophysiology in chronic renal failure, and a low NI is predictive of neutropenia in breast-cancer patients. These findings emphasize the central importance of adequate protein stores in recovery from disease or in maintaining quality of life. Aging appears to involve a gradual loss of TBP throughout adulthood. Cross-sectional data suggest that TBP declines curvilinearly with age, such that there is an accelerated decline after 65 years of age. However, longitudinal data are scarce, and little is known about the relative loss of visceral protein, as opposed to skeletal muscle protein. More clearly-defined data are essential if the effects of aging per se are to be separated from the effects of chronic disease. A further complication is the knowledge that physical activity also declines with age. Thus sarcopenia, the loss of skeletal muscle mass, could primarily result from disuse rather than aging. The economic impact of unsuccessful aging places a pressing need for multicompartment data in longitudinal study designs. PMID- 10865770 TI - Changes in body fluids during endurance rowing training. AB - The aim of this study was to investigate the influence of long-lasting, extensive rowing training on body fluids in 12 experienced rowers (23.3 +/- 6.3 years, 188.7 +/- 5.9 cm, 82.0 +/- 10.8 kg, body mass index [BMI] 23.7 +/- 2.1). All rowers had a light traditional breakfast about 3 h before the training. The subjects rowed 2 h 17 min and covered 22.6 +/- 2.5 km. Before and immediately after the training session and after 30, 60, and 120 min of recovery, body fluid balance was measured using a multiple-frequency impedance device (Multiscan 5000, Bodystat Ltd., UK). Body resistance was measured at the right side of the body. Extracellular (ECW), intracellular (ICW), and total (TBW) body water were measured at 5, 200, and 50 kHz, respectively. Blood hemoglobin (Hgb) and hemotocrit (Hct) were measured, and changes in blood volume (BV), plasma volume (PV), and cell volume (CV) were measured according to Dill and Costill (1974). The body weight of the rowers decreased significantly (p < 0.05) from 82.0 +/- 10.8 kg to 80.6 +/- 11.2 kg during the endurance training. There were no significant changes in TBW immediately after the training session. Significant changes in the content of TBW occurred during the first 30 min of recovery (48.7 +/- 4.4 to 47.9 +/- 4.1; p < 0.05). The amount of ECW also decreased significantly during the first 30 min of recovery (23.3 +/- 2.0 to 22.7 +/- 1.91; p < 0.05). There were no significant changes in the balance of ECW. BV decreased during the training session by 2.5 +/- 5.2% (p < 0.05). There were significant relationships (p < 0.05) between the covered distance and the following parameters after the training session: body weight (r = -0.75), TBW (r = -0.75), and ECW (r = -0.83). Changes in the blood parameters did not correlate significantly with changes in body fluids and training volume. It was concluded that extensive rowing training significantly changed the balance of body fluids in rowers. PMID- 10865771 TI - Body fat content influences the body composition response to nutrition and exercise. AB - In most situations involving a significant change in body weight, both fat-free body mass (FFM) and body fat participate, but the relative contribution of FFM and fat to the total weight change is influenced by the initial body fat content. Overfeeding: In experiments of at least 3-weeks' duration, the weight gain of thin people comprises 60-70% lean tissues, whereas in the obese it is 30-40%. Underfeeding: In humans, there is an inverse curvilinear relationship between initial body fat content and the proportion of weight loss consisting of lean tissue. The same trend holds for animals and birds, including loss during hibernation. Another factor is the magnitude of the energy deficit: as energy intake is reduced, lean tissue makes up an increasing fraction of the total weight loss. Exercise: If individuals lose much weight with exercise, the result is usually some loss of lean tissue as well as fat, and once again the proportion of lean loss to total weight loss is greater in thin people than in those who have larger body fat burdens. Members of twin pairs often differ in weight. In thin individuals, lean accounts for about half of the intrapair weight difference, whereas in the obese it accounts for only one quarter. Body fat content must be taken into account in evaluating body composition changes induced by nutrition and exercise. PMID- 10865772 TI - Total body potassium in healthy Italians and Americans. A cross-calibration study. PMID- 10865773 TI - Effects of exercise and dietary fat upon body composition. A study in a rat model. PMID- 10865774 TI - Analysis of variance/covariance of effect of working posture on body composition. PMID- 10865775 TI - The reference child and adolescent models of body composition. A contemporary comparison. AB - Changes in the relative proportions of bone, muscle, water, visceral tissues, and body fat occur during growth. In the 1980s, reference models of body composition for children and adolescents were constructed by adjusting data on total body water (TBW), total body potassium (TBK), and regional bone mineral (BMC) data from several different Caucasian populations. In our study, we measured TBW, TBK, and total body BMC in 856 healthy European-American, African-American, and Mexican-American children. When we reconstructed the reference models using our contemporary data, we found that the body's bone, protein, and fat compartments are slightly but significantly different from the earlier models. Our study provides the range of normal body composition of healthy children, aged 5-18 years, and accounts for differences related to gender and ethnicity. PMID- 10865776 TI - Body composition measurements during infancy. AB - Infancy is the period of most rapid postnatal growth and is accompanied by major changes in body composition (BC). There are many challenges to successfully measuring BC of infants in vivo, which include the inherent limitations in the underlying assumptions for each technique. The small body mass and rapid nonuniform changes in body parts, that is, the components of BC during infancy, can strain the technical limits of all methods. Many techniques for in vivo BC measurement used in older people have been applied to infants. However, the vast majority of them either are difficult to adapt for widespread use in infants, or the roles and limitations for using them during infancy are ill-defined because of limited or no critical validation and cross-calibration studies. Based on validation data from animals, well-defined methodological issues in data acquisition and analyses, availability of normative data, and pertinent accuracy and precision of the technique to allow us to determinate clinically relevant changes in BC within a reasonable time interval, three techniques appear to be most suitable for in vivo BC measurement in infants. Anthropometric measurements can be used in field studies or for group comparisons, and total body electrical conductivity (TOBEC) and selected dual-energy X-ray absorptiometry (DXA) measurements can be used to compare BC in individual infants. DXA has the advantages of being able to measure bone mass and the potential to be adaptable to the widely available existing instruments. However, regardless of the techniques used in measuring BC in infants, meticulous attention to details in data acquisition and data analysis, and a knowledge of the limitations of the particular technique are the prerequisites for generating valid data. PMID- 10865777 TI - Factors influencing body composition of premature infants at term-adjusted age. AB - We investigated the influence of body size at birth, feeding of mother's milk versus formula, or standard-term formula versus energy- or nutrient-enriched formula on the growth and whole body bone mineral content (BMC), lean and fat mass (using dual-energy X-ray absorptiometry) of low-birth-weight (LBW) infants to term-adjusted age. LBW infants who were appropriate for gestational age were lighter and shorter than term-born infants (n = 46) but had a higher percent fat mass (19-28% vs. 15 +/- 7%). For LBW infants fed standard formula or mother's milk after hospital discharge, the mean BMC expressed either as a function of weight (17 +/- 2, 19 +/- 2 vs. 20 +/- 2 g/kg) or length (1.1 +/- 0.2, 1.1 +/- 0.2 vs. 1.5 +/- 0.2 g/cm) was more than 1 SD below term infant values. However, infants fed a nutrient-enriched formula from hospital discharge had BMC within 1 SD below term infants. Infants who were born small, compared to appropriate for gestational age, compared to infants of similar birth weight had lower percent body fat (16 +/- 6 vs. 19 +/- 5) and lower BMC (47 +/- 3 vs. 62 +/- 5 g) at term age. Both size at birth and diet influence patterns of growth and body composition in early life in very-low-birth-weight (VLBW) infants. The long-term significance of these variable growth patterns in VLBW infants in early life requires further investigation. PMID- 10865778 TI - Value of total body potassium in assessing the nutritional status of children with end-stage liver disease. AB - Malnutrition is a common problem in children with end-stage liver disease (ESLD), and accurate assessment of nutritional status is essential in managing these children. In a retrospective study, we compared nutritional assessment by anthropometry with that by body composition. We analyzed all consecutive measurements of total body potassium (TBK, n = 186) of children less than 3 years old with ESLD awaiting transplantation found in our database. The TBK values obtained by whole body counting of 40K were compared with reference TBK values of healthy children. The prevalence of malnutrition, as assessed by weight (weight Z score < -2) was 28%, which was significantly lower (chi-square test, p < 0.0001) than the prevalence of malnutrition (76%) assessed by TBK (< 90% of expected TBK for age). These results demonstrated that body weight underestimated the nutritional deficit and stressed the importance of measuring body composition as part of assessing nutritional status of children with ESLD. PMID- 10865779 TI - A prospective study of body composition changes in children with cystic fibrosis. AB - Recent cross-sectional studies of children with cystic fibrosis (CF) have shown an improvement in their growth with improved nutritional status, but there are only a few longitudinal studies in this population. A four-year prospective study was conducted in 25 children with CF and 26 controls, ages 5 to 10 years at baseline, to describe changes in body composition using three independent methods of assessment: skinfold thickness, total body water by deuterium dilution, and total body electrical conductivity (TOBEC). The statural growth of the boys with CF was slower than that of the control boys. Using different methods, the fat free mass and fat-mass increases were shown to be slower in boys with CF than in controls. These differences were less significant in girls. Despite comprehensive care, the growth of boys with CF may still not be optimal, as observed longitudinally. PMID- 10865780 TI - Body composition changes during Tanner stage 5. AB - Clinicians use the combination of Tanner stage 5 (T5) of puberty, final height, and epiphyseal fusion to define maturity. We tested the hypothesis that changes in body composition related to age are identifiable during T5. Asian, Black, Hispanic, and White T5 adolescents (n = 148, 72 females) had measurements taken of their height, weight, total body potassium by 40K counting, total body water by D2O dilution; and total body bone mineral, fat-free mass, and fat mass by dual energy X-ray absorptiometry. The relative increases with age in lean body components were greater than those in height and weight, and were greater in males. Age was a significant determinant of all body components in males, but of only bone mineral in females. The effect of age was independent of ethnicity. These findings suggest an independent effect of age on body composition during T5, especially in males. We propose that peak levels of lean body components should be included in the definition of maturity in certain clinical and metabolic situations. PMID- 10865781 TI - Blood pressure, blood insulin, and anthropometry in obese children. A preliminary report. PMID- 10865782 TI - Changes in body composition in adolescents with anorexia nervosa. Comparison of bioelectrical impedance analysis and total body potassium. PMID- 10865783 TI - Annual changes in total body fat and fat-free mass in children from 8 to 18 years in relation to changes in body mass index. The Fels Longitudinal Study. PMID- 10865784 TI - Body composition of children in a depressed economy. PMID- 10865785 TI - Prediction models for bone mineral content in school-aged children. PMID- 10865786 TI - Lean mass of children in various nutritional states. Comparison between dual energy X-ray absorptiometry and anthropometry. PMID- 10865787 TI - Body composition in healthy aging. AB - Health risks in elderly people cannot be evaluated simply in conventional terms of body fatness or fat distribution. Elderly people have less muscle and bone mass, expanded extracellular fluid volumes, and reduced body cell mass compared to younger adults. These nonfat components of body composition play critical roles, influencing cognitive and physical functional status, nutritional and endocrine status, quality of life, and comorbidity in elderly people. Different patterns of "disordered body composition" have different relationships to these outcomes and may require different, tailored approaches to treatment that combine various exercise regimens and dietary supplements with hormone replacement or appetite-stimulating drugs. Skeletal muscle atrophy, or "sarcopenia," is highly prevalent in the elderly population, increases with age, and is strongly associated with disability, independent of morbidity. Elders at greatest risk are those who are simultaneously sarcopenic and obese. The accurate identification of sarcopenic obesity requires precise methods of simultaneously measuring fat and lean components, such as dual-energy X-ray absorptiometry. PMID- 10865788 TI - Smaller organ tissue mass in the elderly fails to explain lower resting metabolic rate. AB - We previously reported our in vivo prediction of whole body resting energy expenditure (REE) using magnetic resonance imaging and echocardiography-derived organ volumes combined with published organ tissue metabolic rates. The models, developed in young healthy persons from predicted and measured variables, were highly correlated (e.g., calculated vs. measured REE, r = 0.92, p < 0.001), with no significant differences (p = NS) between them. This study employed the same approach to determine whether possible age-related decreases in organ tissue mass may account for the lower REE commonly reported in elderly persons. Measurements of REE (REEm) were acquired by indirect calorimetry. Calculated REE (REEc) models were developed from measured tissues and organs, and energy flux rates were assigned for each of the seven tissue/organ components, as reported by Elia. Older men (n = 6) and women (n = 7) had significantly lower REEm compared to REEc (p = 0.001). The magnitude of the differences were 13% and 9.5%, respectively, for men and women. These preliminary data suggest that factors other than organ atrophy may contribute to the lower metabolic rate of older persons. Further studies are required to investigate whether there is a reduction in the oxidative capacity of individual organs and tissues. PMID- 10865789 TI - Reexamining the sarcopenia hypothesis. Muscle mass versus muscle strength. Health, Aging, and Body Composition Study Research Group. AB - The association of muscle mass and muscle strength with lower-extremity performance, as measured by timed repeated chair stands, was investigated using preliminary data from 3,075 Black and White participants (70-79 years old) in the Health, Aging, and Body Composition Study. Leg muscle mass (LM) was measured by dual-energy X-ray absorptiometry (Hologic QDR 4500). The maximal isokinetic torque of the leg extensors (LS) was measured at 60 degrees/s using a Kin-Com isokinetic dynamometer. Men were stronger, had greater LM, and better performance than women. As expected, low LS was associated with poorer performance after adjusting for race, study site, and body fat. Low LM was associated with poorer performance in men and women, with a potential threshold effect in women only. When LS and LM were modeled simultaneously, only LS remained independently associated with performance. In conclusion, muscle strength, but not muscle mass, is independently associated with lower-extremity performance. PMID- 10865790 TI - Waist circumference and sagittal diameter reflect total body fat better than visceral fat in older men and women. The Health, Aging and Body Composition Study. AB - The validity of waist circumference and sagittal diameter as surrogate measures of visceral fat were assessed using preliminary cross-sectional data from the Health, Aging and Body Composition Study, a cohort of 3,075 men and women aged 70 79. Weight, body mass index, waist circumference, waist/thigh ratio, and sagittal diameter were compared by correlation, graphical analysis, and regression to total body fat as measured by dual-energy X-ray absorptiometry (Hologic 4500A), and to visceral fat area as measured by computerized tomography. We included 2,830 persons, 1,439 women and 1,391 men with complete data on all measurements. For both men and women, all measurements were strongly correlated with both total body fat and visceral fat except the waist/thigh ratio. However, waist circumference, sagittal diameter, weight, and body mass index were more closely related to total body fat than to visceral fat area (R2 for the linear regression of waist circumference on total body fat was 0.69 in women and men; R2 for linear regression of waist circumference on visceral fat area was 0.40 in women, and 0.49 in men). These data suggest that the contribution of visceral fat to health risks will be better assessed by directly measuring this fat depot. PMID- 10865791 TI - Body water spaces and cellular hydration during healthy aging. AB - Age-related changes in the proportions of intracellular or extra-cellular water to total body water and in the ratio of total body water to fat-free mass are debatable. These are important issues both for medical reasons (dehydration is a threat in the diseased elderly) and for methodological reasons (most techniques for assessing of body composition assume constant hydration of the fat-free mass). This study compared hydration in young and elderly (60 years) people. In the first part of the study, we analyzed the literature and computed the ratio of total body water over fat-free mass, Hf. Eligible studies involved independent measurements of fat-free mass and total body water. Hf did not appear to change with age. The second part of this study computed Hf in 103 individuals studied in our laboratory. The mean values were not different in young (73.2 +/- 2.4%) and elderly people (73.4 +/- 2.4%). At all ages, the proportion of intracellular or extracellular water (as measured by bromide dilution) to total body water (as measured by oxygen 18 dilution) was similar. The same finding holds for the proportion of intracellular water to fat-free mass. We conclude that hydration of fat-free mass and cellular hydration are not affected in healthy aging. PMID- 10865792 TI - Body composition by air displacement plethysmography in middle-aged and elderly Japanese. Comparison with dual-energy X-ray absorptiometry. PMID- 10865793 TI - Influence of body composition on bone mineral content in elderly women. A preliminary report. PMID- 10865794 TI - Fat distribution and health in obesity. AB - Although independent associations of visceral fat with the insulin resistance syndrome were previously reported in obese women, the importance of truncal subcutaneous fat in this syndrome is controversial. The method by which the various fat depots are measured may be the reason for the underlying controversy. In the past five years, we have used various methods to measure visceral versus subcutaneous fat distribution in Caucasian (C) and African-American (AA) women and have related it to insulin sensitivity (SI) and to blood lipids, particularly fasting serum triglyceride levels (TG). Elevated TG levels in obese women were best predicted by an increased amount of visceral fat, whereas the amounts of truncal and peripheral subcutaneous fat did not have an impact on them. These results were confirmed, regardless of the method used to measure the fat depots. Insulin resistance (low SI) in obese women was predicted by both an increase of visceral and of upper-body (truncal) subcutaneous fat. However, measurements of the entire visceral and truncal subcutaneous fat volumes may be needed to confirm this latter association. PMID- 10865795 TI - Menopause-related changes in body fat distribution. AB - Menopause-related changes in body fat distribution may partially explain the greater risk of cardiovascular and metabolic disease during the postmenopausal years. To date, however, the effect of the menopause transition on body fat distribution remains unclear. Cross-sectional and longitudinal studies using waist circumference or the waist-to-hip ratio show no effect of menopause on body fat distribution. By contrast, studies using dual-energy X-ray absorptiometry showed increased trunk fat in postmenopausal women. Moreover, studies using computed tomography (CT) and magnetic resonance imaging (MRI) show that postmenopausal women have greater amounts of intra-abdominal fat compared to premenopausal women. Collectively, these studies suggest that the menopause transition is associated with an accumulation of central fat and, in particular, intra-abdominal fat. Whether menopause-related differences in trunk or intra abdominal fat are independent of age and/or adiposity, however, is unclear. Thus, we recently examined the effect of menopausal status on body composition and abdominal fat distribution in 53 middle-aged, premenopausal women (47 +/- 3 years) and 28 early postmenopausal women (51 +/- 4 years). Postmenopausal women had 36% more trunk fat (p < 0.01), 49% greater intra-abdominal fat area (p < 0.01), and 22% greater subcutaneous abdominal fat area (p < 0.05) than premenopausal women. The menopause-related difference in intra-abdominal fat persisted (p < 0.05) after statistical adjustment for age and fat mass, whereas no differences were noted in trunk or abdominal subcutaneous fat. A similar pattern of differences in trunk, subcutaneous, and intra-abdominal fat was observed in subsamples of pre- and postmenopausal women matched for age or fat mass. Our data and that of others suggest that early postmenopausal status is associated with a preferential increase in intra-abdominal fat that is independent of age and total adiposity. Thus, CT and MRI should be used when examining menopause-related changes in body fat distribution. PMID- 10865796 TI - Total body calcium in obese women. Validation of dual-energy X-ray absorptiometry against in vivo neutron activation analysis. AB - In vivo neutron activation analysis (IVNA) allows in vivo measurement of total body calcium and is considered the gold standard technique. Dual-energy X-ray absorbtiometry (DXA) estimates total body calcium. Although there are a few comparisons of the two techniques, there are none with obese women. The present study compared the estimation of total body calcium by DXA with the measurement of Ca by IVNA in 69 obese women. Sixty-nine obese women (age 44 +/- 10 years, body weight [BW] 93 +/- 10.4 kg, body mass index [BMI] 35 +/- 4, fat mass 42 +/- 7 kg, 48 pre- and 21 postmenopausal, 30 Caucasian and 39 African-American) were recruited for a six-month body composition study without weight loss intervention. Total body (TB) Ca was measured by IVNA (CaIVNA) and by DXA (CaDXA) at baseline (n = 69) and again at six months (n = 40). At baseline, CaDXA was significantly higher than CaIVNA (1018 +/- 130 g vs. 706 +/- 92 g, p = 0.0001). The difference remained after six months (1028 +/- 153 g vs. 715 +/- 96 g). There was a small but significant increase in TBCa at six months by both DXA (16.5 +/- 40, p < 0.02) and IVNA (8 +/- 22 g, p < 0.03). However, this change was not significantly different between the two methods (p = 0.2365). The correlation between the two methods was very high (r = 0.937) and was independent of either race or menopausal status (p = 0.5163). We generated a regression equation: CaIVNA = 0.659 x CaDXA + 34.77; SEE = 32.2, R = 0.937. We present a model integrating the effect of weight and height on the relationship between CaIVNA and CaDXA. The higher the BMI, the bigger the difference between CaIVNA and CaDXA. In our population with a mean BMI of 35.3, CaIVNA = 69% CaDXA. PMID- 10865797 TI - Relative overhydration of fat-free mass in postobese versus never-obese subjects. AB - Obese subjects who undergo gastrointestinal surgery for weight reduction have relatively greater loss of body cell mass (BCM) than of extracellular fluid. Although the mechanism is uncertain, an implication is that weight-reduced, surgically treated obese subjects are relatively overhydrated. The present investigation explored whether diet-treated postobese (PO) subjects also are relatively overhydrated. Ten PO participants were recruited over a two-year period and matched to never-obese (NO) people by age (+/- 4 years), weight (+/- 3 kg), height (+/- 5 cm), race, and gender. PO participants had lost > or = 18.5 kg and had maintained this loss for 2-16 years. Body density (Db), total body water (TBW), and bone mineral content (BMC) were measured by hydrodensitometry, tritium dilution, and dual-energy X-ray absorptiometry, respectively. Fat and fat-free mass (FFM) were derived using a four-compartment model. BCM was calculated from total body potassium (TBK) measured with a whole body counter. The results indicate that PO subjects were relatively overhydrated and had reduced BCM compared to NO subjects (p < 0.05). These observations are important for interpreting body weight, body composition, and metabolic data of obese subjects who lost weight and maintained their weight loss over the long term. PMID- 10865798 TI - Longitudinal assessment of intra-abdominal fat in postmenopausal women. AB - Longitudinal changes in intra-abdominal (IA) fat and total body fat were assessed in three healthy postmenopausal Caucasian women receiving hormone replacement therapy (HRT) during a seven-year follow-up study. Measurements were made using dual-energy X-ray absorptiometry, anthropometry, and magnetic resonance images. Changes in intra-abdominal fat were examined in relation to different patterns of weight modification. The weight-stable women had the lowest percent body fat and IA fat at all examinations, with a slight decrease in IA fat at follow-up. The person with a gradual gain in weight of 1.9 kg experienced small increases in body fat and IA fat. The weight-cycling woman had the highest weight, percent body fat, and IA fat area at all examinations. The amounts of IA fat and percent body fat fluctuated with body weight fluctuations. Together with other findings, our results suggest that lean, weight-stable individuals receiving HRT can maintain a low level of IA fat, whereas those experiencing weight gains or regaining lost weight have increases in IA fat. PMID- 10865799 TI - Effect of a weight-reduction program on total and regional body composition in obese postmenopausal women. PMID- 10865800 TI - Reproducibility of body measurements in very obese postmenopausal women. PMID- 10865801 TI - Measurement of percent body fat during weight loss in obese women. Comparison of four methods. PMID- 10865802 TI - Association between leptin, insulin, and body fat distribution in 2-diabetes mellitus. PMID- 10865803 TI - Body composition studies in HIV-infected individuals. AB - Malnutrition is common in HIV infection. Early studies demonstrated a disproportionate depletion of body cell mass compared to body weight, plus relative expansion of extracellular water volume. Neutron activation studies showed that both potassium and nitrogen were depleted in the HIV+ subjects, whereas cross-sectional imaging documented depletion of skeletal muscle mass. The etiology of malnutrition affects the composition of lost weight. Malnutrition is associated with adverse outcomes, whereas clinical stability is associated with nutritional stability. Increasingly, body composition studies are being incorporated into clinical trials. Hypercaloric feeding promotes gains in weight and body fat, but not in lean mass. Adjunctive therapies include anabolic agents, both steroids and recombinant human growth hormone (rhGH), cytokine inhibitors, and resistance training exercise. In addition to increasing fat-free mass, these therapies also have benefits in quality of life, notably functional performance, as well as physical function. Current research on alterations in body composition in HIV have noted a redistribution of fat, with visceral obesity in patients receiving highly active antiretroviral therapies. PMID- 10865804 TI - Sarcopenic obesity: does muscle loss cause fat gain? Lessons from rheumatoid arthritis and osteoarthritis. PMID- 10865805 TI - Body composition assessed by neutron activation analysis in dialysis patients. AB - Malnutrition is prevalent in end-stage renal disease (ESRD) patients treated with hemodialysis (HD) and peritoneal dialysis (PD). In addition, there is increased incidence of morbidity in this group. Evaluation of nutritional status is important. Application of body composition in the ESRD population to evaluate body compartments and to assess nutritional health has become more common in clinical practice. Neutron activation analysis (NAA) may provide data on metabolically active tissue by quantification of total body potassium (TBK) for body cell mass and assessment of protein by total body nitrogen (TBN). This method may be able to detect changes in body composition before clinical signs of malnutrition are apparent. Ten HD (5 male and 5 female) and 10 PD patients (7 male and 3 female) were evaluated by NAA, TBK, and isotope dilution. Female PD patients had an increased total body water (TBW) and increased intracellular water compared to HD females. Albumin was lower in PD women. There was no significant difference between PD men and laboratory controls in TBW, extracellular water, and TBN. The clinical application of body composition methods for evaluation of dialysis patients by serial assessment and for development of a bedside tool needs further study. PMID- 10865806 TI - Bone mass and gastrointestinal disease. AB - Several gastrointestinal and liver diseases impair the absorption of calcium, phosphate, and/or vitamin D, and are associated with an increased incidence of bone disease. Changes in bone mineral density (BMD) using dual-energy X-ray absorptiomety (DXA) have been best studied in the malabsorptive disorder, celiac disease. Celiac disease is an inflammatory condition of the small intestine triggered by ingesting gluten present in wheat, rye, or barley. Chronic inflammation leads to intestinal atrophy and nutrient malabsorption. The disease affects the proximal small bowel; the site where calcium is best absorbed. About 70% of adults with celiac disease have abnormally low BMD values. Treatment with a gluten-free diet increases BMD, but not to normal values. As celiac disease may not be detected until adult life, the failure to reach normal BMD on a gluten free diet can be explained, at least in part, by the failure to reach peak bone mass. All individuals with malabsorptive disorders should be screened for secondary bone disease. The development of easier and less expensive methods to assess BMD will facilitate screening those at risk for bone disease. PMID- 10865807 TI - Early diagnosis of lymphedema in postsurgery breast cancer patients. AB - Lymphedema is an accumulation of lymph fluid in the limb resulting from an insufficiency of the lymphatic system. It is commonly associated with surgical or radiotherapy treatment for breast cancer. As with many progressively debilitating disorders, the effectiveness of treatment is significantly improved by earlier intervention. Multiple frequency bioelectrical impedance analysis (MFBIA) previously was shown to provide accurate relative measures of lymphedema in the upper limb in patients after treatment for breast cancer. This presentation reports progress to date on a three-year prospective study to evaluate the efficacy of MFBIA to predict the early onset of lymphedema in breast cancer patients following treatment. Bioelectrical impedance measurements of each upper limb were recorded in a group of healthy control subjects (n = 50) to determine the ratio of extracellular limb-fluid volumes. From this population, the expected normal range of asymmetry (99.7% confidence) between the limbs was determined. Patients undergoing surgery to treat breast cancer were recruited into the study, and MFBIA measurements were recorded presurgery, at one month and three months after surgery, and then at two-month intervals for up to 24 months postsurgery. When patients had an MFBIA measure outside the 99.7% range of the control group, they were referred to their physician for clinical assessment. Results to date: Over 100 patients were recruited into the study over the past two years; at present, 19 have developed lymphedema and, of these, 12 are receiving treatment. In each of these 19 cases, MFBIA predicted the onset of the condition up to four months before it could be clinically diagnosed. The false-negative rate currently is zero. The study will continue to monitor patients over the remaining year to accurately ascertain estimates of specificity and sensitivity of the procedure. PMID- 10865808 TI - Effect of recombinant human growth hormone on regional tissue distribution in growth hormone-deficient males. AB - Dual-energy X-ray absorptiometry was used to analyze the regional tissue distribution of bone mineral mass (BMM), fat mass (FM), and lean tissue mass (LTM) in the arms, trunk, and legs of 23 male growth hormone-deficient (GHD) males. Patients were assigned randomly to treatment (n = 11) and control (n = 12) groups. During the first six months, the treatment group received recombinant growth hormone (rhGH) and the control group placebo. During the second six months, both groups received rhGH. Following treatment there was a trend for BMM to be lost in the arms and legs, with gains at the trunk. Fat mass was lost mainly from the trunk. Proportionally more LTM was distributed to the arms and legs than to the trunk. Treatment was more effective in the first six months than the second. The correlation between the waist-hip ratio and trunk fat was poor, suggesting that its use in patients with GHD may be misleading. PMID- 10865809 TI - The prognostic value of body protein in patients with lung cancer. AB - Lung cancer is a major cause of death in many countries. To improve the results of treatment, more individualized therapy is necessary; for this, it is necessary to identify new prognostic factors. In 21 patients with lung cancer (17 with non small-cell lung cancer and 4 with small-cell lung cancer) that had received radiation treatment, the amount of body protein was estimated with in vivo neutron activation analysis. Patients in whom body protein decreased had recurrences of the disease earlier and a poorer survival than patients whose body protein increased. A clear relationship was seen between changes in the body's protein content and recurrence-free survival. To better evaluate the prognostic value of body protein content in patients with lung cancer, a larger number of patients and a longer follow-up period are needed. PMID- 10865810 TI - Similarity of changes in body composition in intensive care patients following severe sepsis or major blunt injury. AB - Critically ill patients admitted to the intensive care unit with severe sepsis or major blunt injury undergo massive changes in body composition. We compared these changes in 12 patients with generalized peritonitis, and in 18 patients with major blunt injury over a 21-day period soon after their admission. Body composition was measured as soon as the patients were hemodynamically stable, and again 5, 10, and 21 days later. In both groups, losses in total body protein (TBP) were greatest over the first 10 days. TBP lost over the study period averaged 13.1 +/- 1.3 (SEM)% for the sepsis group, and 14.6 +/- 1.3% for the trauma group. Total body water (TBW) lost postresuscitation averaged 11.1 +/- 1.3 L and 6.7 +/- 1.1 L for the two groups, respectively, these changes largely being accounted for by changes in extracellular water (ECW). Our results demonstrate a striking similarity in the changes in total body protein for these two groups of critically ill patients. The sepsis patients retained approximately twice the volume of fluid of those with major trauma. PMID- 10865811 TI - Positive impact of protease inhibitors on body composition and energy expenditure in HIV-infected and AIDS patients. PMID- 10865812 TI - Effects of whey protein and resistance exercise on body composition and muscle strength in women with HIV infection. PMID- 10865813 TI - The usefulness of walking for preventing sarcopenia in dieting postmenopausal women complaining of knee pain. PMID- 10865814 TI - Longitudinal study of patients with anorexia nervosa 6 to 10 years after treatment. Impact of adequate weight restoration on outcome. PMID- 10865815 TI - Changes in body composition and resting energy expenditure in anorectic patients after a weight gain of fifteen percent. PMID- 10865817 TI - Bone density and body composition in young women with chronic fatigue syndrome. PMID- 10865816 TI - Body composition in homozygous beta-thalassemia. PMID- 10865818 TI - In vivo body composition studies of young adult males participating in the cleanup of the Chernobyl Nuclear Power Plant. PMID- 10865820 TI - Genomic searches for genes that influence atherosclerosis and its risk factors. AB - We are performing genomic searches in randomly ascertained families to identify new quantitative trait loci (QTLs) that influence atherosclerosis and its risk factors. The genetic markers used for genomic searches are random microsatellite markers distributed throughout the human chromosomes. These markers are used for linkage analysis with variance component methods to identify QTLs for measured phenotypes related to lipid metabolism and atherosclerosis. We conducted such a genomic search in 477 participants of the San Antonio Family Heart Study. This genomic search identified QTLs on chromosomes 3 and 4 that influence LDL size class, an important risk factor of atherosclerosis. In addition to lipid risk factors, we measured a variety of gene products involved in atherogenesis in the arterial wall (such as adhesion molecules and components of hemostasis). We found QTLs for serum levels of soluble P-selectin on chromosome 15 (LOD = 3.8) and chromosome 12 (LOD = 2.6). PMID- 10865819 TI - Relevance of, and potentiality for, in vivo intrathyroidal iodine determination. PMID- 10865821 TI - Genetic analysis of essential hypertension in Japanese populations. AB - The reverse genetic approach, which examines genetic factors underlying the root of pathogenesis first, is a powerful tool to clarify the genetic cause of essential hypertension. Using the rat cross model, studies of the genetically hypertensive inbred rat model indicated several candidate loci on the rat chromosome responsible for blood pressure, but failed to identify the exact causal gene. Moreover, it was not certain that the rat data really reflect the human case. Thus, we shifted our focus to human genetics and carried out case control studies using the candidate gene approach. We mainly focused on gene components of the renin-angiotensin system as candidates, finding that angiotensinogen gene polymorphisms are genetic predisposing factors for hypertension. However, the results obtained from case-control studies using Japanese subjects were not consistent, suggesting that there was a problem in control sampling. In our recent study, we recruited more than 5,000 residents of an urban community as a general population and examined the association between genetic factors and their health status. Our results indicate that angiotensin converting enzyme gene polymorphism is a male-specific genetic risk for essential hypertension. In light of our previous investigations, we present a discussion concerning the design of future studies of the genetics of hypertension. PMID- 10865822 TI - Tailored therapy to fit individual profiles. Genetics and coronary artery disease. AB - In the twentieth century, coronary artery disease (CAD) was the major cause of morbidity and mortality in western societies. Genetics and environmental influences clearly contribute to CAD. Our genetic makeup contributes to not only coronary risk factors, but also determines an individual's response to environmental challenges such as diet, drugs, and tobacco. Genetic testing is likely to be more predictive of our predisposition to CAD than current conventional testing of known risk factors. Therapy aimed at altering the regulation of genes or their transcribed proteins may prove appropriate in individual patients, depending on their genetic background. The understanding of our genetic predisposition to CAD may enable targeted environmental modification strategies aimed at individuals at greatest risk. Both in terms of the huge costs of drugs to society and in terms of needless medicalization, genotyping in order to tailor therapy might become a routine part of risk management earlier than we think. PMID- 10865824 TI - The extracellular matrix dynamically regulates smooth muscle cell responsiveness to PDGF. AB - Focal accumulation of smooth muscle (SMC) within the arterial intima contributes to the formation of lesions of atherosclerosis. Platelet-derived growth factor (PDGF) is a potent stimulant of SMC migration and proliferation in culture that may play a role in the accumulation of SMC in atherogenesis. SMCs normally reside in the media of the artery wall surrounded by extracellular matrix (ECM), including type I collagen. In atherogenesis, the ECM is degraded, new ECM components, such as fibronectin, are synthesized and assembled, and these alterations in ECM components are associated with changes in SMC phenotype. To model the changes in ECM in normal and diseased arteries, we have analyzed SMCs cultured on different forms of type I collagen. Our studies demonstrate that integrin-mediated signals from various forms of type I collagen lead to specific and rapid modulation of the integrin signaling complex, including cytoskeletal connections, and of the responsiveness of SMC to PDGF stimulation. PMID- 10865823 TI - Genetic diversity in the matrix metalloproteinase family. Effects on function and disease progression. AB - Atherosclerosis is an example of a complex trait, where the course of the disease is influenced by a combination of common variation in a constellation of genes and the effect of a wide range of environmental variables. Thus, the underlying disease mechanisms will be modulated by genetic diversity and the effect this diversity has on an individual's response to environmental challenges such as smoking, diet, and exercise. Unlike the consequences of mutations in severe single-gene disorders on protein function, the impact of individual common, functionally important sequence changes in genes contributing to multifactorial diseases is likely to be very small. The challenge is to dissect the contribution that each of these genes makes to the disease process. We have tackled this by identifying common genetic variants, studying their effects on function, and applying them to the analysis of association in appropriately structured and suitably powered studies. Even with our incomplete understanding of the disease, the list of potential candidate genes we could study is vast; but, we do know from pathological studies that a wide spectrum of structural architecture exists in atherosclerotic plaques, suggesting that remodeling of vascular connective tissue is fundamentally important. Matrix remodeling is controlled by a complex network of cell and matrix interactions, the net outcome of which is the product of a balance between synthetic and degradative processes. Our work has focused on the family of enzymes and inhibitors most directly associated with matrix turnover--the matrix metalloproteinases (MMPs) and their natural inhibitors (TIMPs, tissue inhibitors of MPs). We specifically searched for functionally relevant genetic variants that might modulate the delicate control of matrix turnover. Using these molecular genetic strategies to investigate the impact of natural genetic variation on vascular matrix remodeling has begun to shed new light on the importance of these genes in atherogenesis. PMID- 10865825 TI - The role of adaptive immunity in atherosclerosis. AB - Atherosclerosis is an inflammatory disease induced by a lipid metabolic disturbance at sites of hemodynamic strain in the vasculature. Studies in both man and experimental animal models show an involvement of innate and adaptive immune mechanisms in the disease process. Our recent studies in apoE-knockout mice show that the level of hypercholesterolemia affects the functional properties of the immune response. Modulating immune activity by injections of polyclonal immunoglobulins inhibits disease progression, suggesting that immunomodulation may be useful to treat atherosclerosis. Analysis of T cell receptor (TCR) mRNA in atherosclerotic lesions shows expansions of T cells expressing TCR-V beta 6, a receptor type that is also expressed by T cells recognizing oxidized low density lipoprotein (oxLDL). This suggests that oxLDL is an autoantigen that induces strong, local T cell responses in the plaque. Further characterization of this and other candidate antigens, such as heat shock proteins and macromolecular components of Chlamydia pneumoniae, may provide important information on which specific interference with the disease process could be based. PMID- 10865826 TI - Inflammation and atherosclerosis. Atherosclerotic lesions in Takayasu arteritis. AB - Takayasu arteritis is a chronic vasculitis, mainly involving the aorta and its main branches as well as the coronary and pulmonary arteries, causing stenosis and/or obstruction due to thrombus formation or dilatation due to aneurysmal formation and/or rupture of the involved arteries. These characteristic anomalies resulted from ischemia of retinal arteries due to the obstruction of cervical vessels. In Western countries this disease is also known as "pulseless disease," because the pulse is frequently absent due to the obstruction of subclavian or branchial arteries. The pathogenesis of this morbid condition is still unknown. Epidemiologically, it is found mostly in female patients and is more prevalent in Asian and Latin American countries. Affected areas consist of a mixture of both active, productive inflammatory lesions, and old fibrous lesions. Autoimmune processes stimulated by viral infection and other unknown causative factors may play an important role under these pathophysiological conditions because HLA analysis revealed a statistically significant high frequency of haplotype A24-B52 DR2 in these patients in Japan. Documentation of atherosclerotic complications in young female patients with Takayasu arteritis who are generally free from traditional atherosclerosis risk factors may be clinical evidence that inflammation is indeed an important risk factor in atherogenesis. PMID- 10865827 TI - Gene therapy for heart transplantation-associated coronary arteriosclerosis. AB - Cardiac allograft arteriosclerosis, which limits the long-term survival of recipients, cannot be prevented by conventional therapies. The arteriopathy is characterized by diffuse intimal thickening made up of proliferative smooth muscle cells. To test the hypothesis that cell cycle-regulatory genes play crucial roles in the development of this arteriopathy in vivo gene therapy targeting cell division cycle (cdc) 2 kinase was attempted in murine cardiac allografts using hemagglutinating virus of Japan (HVJ)-liposome method. Antisense cdc 2 kinase oligodeoxynucleotide (ODN) was transfected into the allografts by intraluminal injection during the operation and the allografts were harvested at 4 weeks after transplantation. Coronary intimal thickening had developed in sense ODN-treated allografts and ICAM-1 and VCAM-1 were enhanced in these arteries. PDGF mRNA was also detected. Antisense cdc 2 kinase ODN inhibited intimal hyperplasia. These data indicate that antisense cdc 2 kinase modulates gene expression and inhibits smooth muscle cell proliferation of graft arteries. PMID- 10865828 TI - Role of endothelin-1 in atherosclerosis. AB - Increased evidence has shown that endothelin-1 (ET-1) derived from the arterial cells is involved in the development of atherosclerosis. ET-1 and ET receptors are upregulated in both human and experimental animal atherosclerotic lesions. Plasma ET-1 levels are significantly elevated in hypercholestolemic subjects and cholesterol-fed animals. We hypothesized that plasma lipoproteins such as LDL and HDL retained in the arterial wall can affect ET-1 production and secretion, thereby sustaining vascular functions. Using a two-chamber culture system, we have demonstrated that endothelial cells (ECs) show a polar secretion of ET-1; the majority of ET-1 are secreted toward the basal side of the vessels. Furthermore, we found that LDL enhances whereas HDL inhibits the ET-1 secretion from ECs in a polarized pattern. In order to demonstrate ET receptor distribution in the lesion, we recently studied both human and apoE-KO mice. Our study showed that there is an increased expression of ETB receptors in foamy macrophages in the lesions. More importantly, medial smooth muscle cells (SMCs) beneath the foam cell lesions exhibited a higher intensity of ETB receptor immunoreactivity than those located in foam cell-free areas. In such an area, ET-1 immunoreactivity is also increased. These results suggest that accumulation of foamy macrophages may modulate the shift of ET receptor subtypes from ETA to ETB in SMCs and an enhanced ET system mediated by ETB receptors may play a pivotal role in the progression of atherosclerosis. This notion has been further supported by a recent finding that administration of ET receptor antagonists resulted in a significant reduction of atherosclerosis in apoE-KO mice. PMID- 10865829 TI - Oxidized-LDL and atherosclerosis. Role of LOX-1. AB - The accumulation of substantial numbers of monocyte/macrophages and activated T lymphocytes in focal areas of the arterial intima appears to be a hallmark of atherosclerosis. Our report demonstrated that lysophosphatidylcholine (lyso-PC), a polar phospholipid component that is increased in atherosclerotic lipoproteins, such as oxidized LDL and remnant lipoproteins in diabetic and Type 3 hyperlipidemia, can upregulate adhesion molecules for monocytes and T lymphocytes, and growth factors, such as heparin-binding epidermal growth factor like growth factor and PDGF A and B chains. Recently, we identified the novel receptor for oxidized LDL, named LOX-1. We summarize the importance of the interaction between oxidized LDL and its receptor, LOX-1, in terms of early stage atherogenesis. PMID- 10865830 TI - Receptors and lipid transfer proteins in HDL metabolism. AB - It is believed that HDL exerts its anti-atherogenic effects through the process of delivering cholesterol from peripheral tissues back to the liver for removal from the body (i.e., reverse cholesterol transport). The metabolic life cycle of HDL lipid and apolipoproteins during reverse cholesterol transport involves both its modification in plasma by lipid transfer proteins and the clearance from plasma of HDL lipid and protein mediated by hepatic cell surface proteins. We review recent work from our laboratory that focuses on specific metabolic steps in reverse cholesterol transport and the results of altering these steps on plasma HDL levels and atherosclerosis. Recently, SR-BI was shown to be an authentic HDL receptor mediating the selective uptake of HDL lipids into cells without degradation of HDL proteins. We discuss the evidence for additional receptor activity mediating HDL protein catabolism in the liver from studies in obese (ob/ob) mice, which have markedly increased HDL due to a defect in hepatic catabolism of apoA-I and apoA-II. In addition, we review recent findings that phospholipid transfer protein deficiency in mice results in markedly reduced HDL levels. Lastly, we highlight our findings that overexpression of SR-BI in LDL receptor-deficient mice results in decreased atherosclerosis. PMID- 10865831 TI - Scavenger receptor classes A and B. Their roles in atherogenesis and the metabolism of modified LDL and HDL. AB - Scavenger-receptor class A has been held responsible for the clearance of modified LDL from the blood circulation. However, in mice deficient in scavenger receptor class A, the decay in vivo of acetylated LDL (t1/2 < 2 min), as well as tissue distribution and liver uptake (at 5 min 77.4 +/- 4.6% of the injected dose) are not significantly different from control mice. The degradation capacity of acetylated LDL with liver endothelial cells, Kupffer cells, and peritoneal macrophages from knock-out mice was 58%, 63%, and 17% of the control, respectively, indicating that scavenger-receptor class A is relatively more important for the degradation of acetylated LDL and foam cell formation in peritoneal macrophages as compared to the liver cell types. This might explain the 60% reduction in atherosclerotic lesion area in scavenger-receptor-deficient apoE knock-out mice as compared to control apoE knock-out mice. Scavenger receptor BI can facilitate selective uptake of cholesterol esters from HDL. A high cholesterol diet for two weeks induced an 80% downregulation of scavenger receptor BI in the liver parenchymal cells while expression in liver macrophages is increased fourfold. The in vivo kinetics for the selective uptake of (oxidized) cholesterol esters from HDL correlate with the changes in scavenger receptor BI expression. It is suggested that scavenger-receptor BI is subject to different regulatory mechanisms in parenchymal liver cells and macrophages related to a difference in function in these cell types. PMID- 10865832 TI - CD36 in atherosclerosis. The role of a class B macrophage scavenger receptor. AB - CD36, an 88 kD transmembrane glycoprotein, is an important receptor for oxidized lipoproteins. Unlike the LDL receptor, expression of CD36 is upregulated by this pro-atherogenic particle, and binding and uptake perpetuates a cycle of lipid accumulation and receptor expression. This effect is, in part, mediated by the transcription factor, peroxisome proliferator activated receptor-gamma (PPAR gamma), and its ligands. We have found that specific inhibitors of protein kinase C (PKC) reduce basal mRNA expression of CD36 and block induction of CD36 mRNA and protein by oxidized LDL (OxLDL) and a PPAR gamma ligand. In addition, PKC inhibitors block both PPAR gamma mRNA and protein expression. These results suggest that activation of CD36 gene expression by OxLDL involves activation and translocation of PKC with subsequent PPAR gamma activation. More recently, we have generated a mouse null for CD36, and crossed it with the atherogenic Apo E null strain. Evaluation of lesion development in these animals will allow us to assess the in vivo contribution of CD36 to the pathogenesis of atherosclerosis. PMID- 10865833 TI - Animal models for hyperinsulinemia and insulin resistance. AB - The direct effect of endogenous insulin on the atherosclerotic process has not been well understood. To clarify this question, we performed pancreas transplantation in Wistar Shionogi (WS) rats. Hyperinsulinemia was not related to coronary risk factors such as dyslipidemia and hypertension in transplanted rats. After 9 months of transplantation, the cholesterol ester contents of the aortas of transplanted WS rats were significantly higher than in the control rats. The effects of insulin resistance on coronary risk factors were examined in mice deficient in insulin substrate-1-deficient (IRS-1) mice, a non-obese animal model of insulin resistance. Blood pressure and plasma triglyceride levels were significantly higher in IRS-1-deficient mice than in normal mice. Impaired endothelium-dependent vascular relaxation was also observed in IRS-1-deficient mice. Furthermore, lipoprotein lipase activity was lower than in normal mice, suggesting impaired lipolysis was involved in the increased plasma triglyceride levels under insulin-resistant conditions. PMID- 10865834 TI - Lipid lowering reduces proteolytic and prothrombotic potential in rabbit atheroma. AB - Thrombus formation at sites of atheromatous plaque disruption cause most acute coronary events such as myocardial infarction and unstable angina. Lesional macrophages and smooth muscle cells produce matrix metallo-proteinases (MMPs) and tissue factor (TF), the molecules likely contribute to plaque rupture and thrombus formation. Recent clinical studies have suggested that lipid lowering can reduce the incidence of acute coronary events. We have recently determined the effects of long-term dietary lipid lowering on atheroma of high-cholesterol fed rabbits. Lipid lowering diminished macrophage accumulation, reduced expression and activity of MMPs, and increased interstitial collagen accumulation in rabbit atheroma. Expression and activity of TF in atheroma also substantially decreased during lipid lowering. Dietary lipid lowering also promoted accumulation of mature smooth muscle cells expressing less MMPs and TF in the plaque's fibrous cap. These results suggest potential mechanisms by which lipid lowering reduces acute coronary events in patients by decreasing proteolytic and prothrombotic activity within the atheroma. PMID- 10865835 TI - Mechanisms of vascular atrophy and fibrous cap disruption. AB - The process of plaque destabilization and rupture remains an area of intense investigation. While reductions in lumen cross-sectional area induced by early, non-occlusive lesions are compensated by remodeling and expansion of the artery, further plaque enlargement leads to an uncompensated reduction in lumen area and an increase in surface shearing forces. We hypothesize that these local increases in wall shear stress lead to a reduction in smooth muscle cell proliferation and increase in cell death. Using a primate prosthetic graft model, we have observed that alterations in nitric oxide and platelet-derived growth factor metabolism are important regulators of intimal growth and regression. We suggest that these factors may also be influential in the process of fibrous cap atrophy and plaque rupture. PMID- 10865836 TI - Roles of the AGE-RAGE system in vascular injury in diabetes. AB - This study concerns whether advanced glycation endproducts (AGE) are related to microvascular derangement in diabetes, exemplified by pericyte loss and angiogenesis in retinopathy and by mesangial expansion in nephropathy. AGE caused a decrease in viable pericytes cultivated from bovine retina. On the other hand, AGE stimulated the growth and tube formation of human microvascular endothelial cells (EC), this being mediated by autocrine vascular endothelial growth factor. In AGE-exposed rat mesangial cells, type IV collagen synthesis was induced. Those AGE actions were dependent on a cell surface receptor for AGE (RAGE), because they were abolished by RAGE antisense or ribozyme. The AGE-RAGE system may thus participate in the development of diabetic microangiopathy. This proposition was supported by experiments with animal models; several indices characteristic of retinopathy were correlated with circulating AGE levels in OLETF rats. The predisposition to nephropathy was augmented in RAGE transgenic mice when they became diabetic. PMID- 10865837 TI - Characterization of atherosclerotic plaques by magnetic resonance imaging. AB - The study of atherosclerotic disease during its natural history and after therapeutic intervention will enhance our understanding of the progression and regression of this disease and will aid in selecting the appropriate medical treatments or surgical interventions. Several invasive and noninvasive imaging techniques are available to assess atherosclerotic vessels. Most of these techniques are strong in identifying the morphological features of the disease, such as lumenal diameter and stenosis or wall thickness, and in some cases provide an assessment of the relative risk associated with the atherosclerosis. However, none of these techniques can fully characterize the composition of the atherosclerotic plaque in the vessel wall and, therefore, are incapable of identifying the vulnerable plaques. High-resolution, multi-contrast, magnetic resonance (MR) can non-invasively image vulnerable plaques, characterize plaques in terms of lipid and fibrous content, and identify the presence of thrombus or calcium. Application of MR imaging opens up whole new areas for diagnosis, prevention, and treatment (e.g., lipid-lowering drug regimens) of atherosclerosis. PMID- 10865838 TI - Signaling angiogenesis via p42/p44 MAP kinase cascade. AB - Vascular endothelial growth factor (VEGF), a potent agonist secreted by virtually all cells, controls migration and division of vascular endothelial cells. Disruption of one VEGF allele in mice has revealed a dramatic lethal effect in early embryogenesis, suggesting a key role in vasculogenesis. We analyzed the regulation of VEGF mRNA in normal and transformed CCL39 fibroblasts and then dissected the VEGF promoter to identify the signaling pathway(s) controlling the activation of this promoter in response to growth factors, oncogenes, and hypoxic stress. We demonstrated that the p42/p44 MAP kinase signaling cascade controls VEGF expression at least at two levels. In normoxic conditions, MAPKs activate the VEGF promoter at the proximal (-88/-66) region where Sp-1/AP-2 factors bind. Activation of p42/p44 MAPKs is sufficient to turn on VEGF mRNA. At low O2 tension, hypoxia inducible factor-1 alpha (HIF-1 alpha), a limiting factor rapidly stabilized and phosphorylated, plays a key role in the expression of several genes including VEGF. We demonstrated that p42/p44MAPKs stoichiometrically phosphorylate HIF-1 alpha in vitro and that HIF-1-dependent VEGF gene expression is strongly enhanced by the exclusive activation of p42/p44MAPKs. Finally, we demonstrated that the regulation of p42/p44MAPK activity is critical for controlling proliferation and growth arrest of vascular endothelial cells at confluency. These results point to at least three major targets of angiogenesis where p42/p44 MAP kinases exert a determinant action. PMID- 10865839 TI - Properties of two VEGF receptors, Flt-1 and KDR, in signal transduction. AB - The properties of two VEGF receptors, Flt-1 and KDR, in the signal transduction of VEGF in human umbilical vein endothelial cells (HUVECs) were investigated by using two newly developed blocking monoclonal antibodies (mAbs) against Flt-1 and KDR. VEGF stimulated the expression of transcription factor Ets-1 as well as matrix metalloproteinase-1 (MMP-1) and Flt-1 in HUVECs. The KDR/Flt-1 heterodimer and the KDR homodimer mediate the expression of Ets-1, MMP-1, and Flt-1. VEGF also stimulated DNA synthesis and migration of HUVECs. DNA synthesis is mediated by the same signaling system as the expression of Ets-1. In contrast, cell migration is regulated by two distinct signaling systems. The Flt-1 homodimer is required for actin reorganization. The KDR/Flt-1 heterodimer and the KDR homodimer are required for the assembly of vinculin in focal adhesion plaque by regulating the phosphorylation of focal adhesion kinase (FAK) and paxillin. PMID- 10865840 TI - Regulation of angiogenesis by controlling VEGF receptor. AB - The endothelial cells cultured in collagen gel caused upregulation of KDR expression, which resulted in an increase in tube formation. Endothelial cells exposed to high glucose (33 mmol/l) for 30 days increased the tube formation induced by VEGF, but not by serum and bFGF. Immunohistochemical study showed that KDR expression was upregulated by the high-glucose treatment. The endothelial cells treated with 0.5-5 micrograms/ml eicosapentaenoic acid (EPA, 20:5, n-3) for 48 h displayed a dose-dependent suppression of tube formation, VEGF-induced proliferation, and activation of p42/p44 MAP kinase but not bFGF-induced ones. Pretreatment with arachidonic acid (20:4, n-6) and docosahexaenoic acid (22:6, n 3) did not show such effects. The expression of KDR was downregulated by the EPA pretreatment. The bone is the richest tissue in microvessel networks except for the liver. Osteoblasts produced VEGF and some factor(s) that could induce KDR upregulation in endothelial cells and could enhance tube formation. These results lead to the speculation that the regulation of KDR expression as well as VEGF production is deeply involved in angiogenesis under various conditions. PMID- 10865841 TI - Transcriptional regulation of smooth muscle phenotypic modulation. AB - Phenotypic modulation of vascular smooth muscle cell plays a pivotal role in the development of vascular pathology, such as atherosclerosis and restenosis after angioplasty. We have identified the zinc finger protein BTEB2 as a DNA binding protein that regulates the nonmuscle myosin heavy chain (SMemb) promoter. BTEB2 is expressed in fetal aorta but not in adult aorta and is induced in the neointima in response to vascular injury. BTEB2 also activates a number of vascular disease-associated genes, such as tissue factor, PAI-1 (plasminogen activator inhibitor-1), and Egr-1 gene. We have further isolated and characterized the human BTEB2 gene. Functional studies using 5'-deletion and site directed mutation constructs demonstrated that phorbol ester induces Egr-1, which can activate the BTEB2 promoter through binding to -32 from the transcription start site. These results suggest that phenotypic modulation of vascular smooth muscle cells occurring in response to mitogen stimulation may be mediated by BTEB2 through Egr-1 induction. PMID- 10865843 TI - Endothelial dysfunction, hemodynamic forces, and atherogenesis. AB - Phenotypic modulation of endothelium to a dysfunctional state contributes to the pathogenesis of cardiovascular diseases such as atherosclerosis. The localization of atherosclerotic lesions to arterial geometries associated with disturbed flow patterns suggests an important role for local hemodynamic forces in atherogenesis. There is increasing evidence that the vascular endothelium, which is directly exposed to various fluid mechanical forces generated by pulsatile blood flow, can discriminate among these stimuli and transduce them into genetic regulatory events. At the level of individual genes, this regulation is accomplished via the binding of certain transcription factors, such as NF kappa B and Egr-1, to shear-stress response elements (SSREs) that are present in the promoters of biomechanically inducible genes. At the level of multiple genes, distinct patterns of up- and downregulation appear to be elicited by exposure to steady laminar shear stresses versus comparable levels of non-laminar (e.g., turbulent) shear stresses or cytokine stimulation (e.g., IL-1 beta). Certain genes upregulated by steady laminar shear stress stimulation (such as eNOS, COX 2, and Mn-SOD) support vasoprotective (i.e., anti-inflammatory, anti-thrombotic, anti-oxidant) functions in the endothelium. We hypothesize that the selective and sustained expression of these and related "atheroprotective genes" in the endothelial lining of lesion-protected areas represents a mechanism whereby hemodynamic forces can influence lesion formation and progression. PMID- 10865842 TI - Blood vessels from bone marrow. AB - Lengths of silastic tubing were inserted into the peritoneal cavity of rats or rabbits. By two weeks the free-floating implants had become covered by a capsule consisting of several layers of "macrophage"-derived myofibroblasts and collagen matrix overlaid by a single layer of mesothelial cells. The tubing was removed from the harvested implant and the tissue everted. This now resembled an artery with an inner lining of mesothelial cells (the "intima"), a "media" of myofibroblasts, and an outer collagenous "adventitia." The tube of living tissue was grafted by end-to-end anastomoses into the transected carotid artery or abdominal aorta of the same animal in which the tissue had been grown, where it remained patent for four months and developed structures resembling elastic lamellae. The myofibroblasts developed a high volume fraction of myofilaments and became responsive to contractile and relaxing agents similar to smooth muscle cells of the adjacent artery wall. PMID- 10865844 TI - Role of the vascular NADH/NADPH oxidase system in atherosclerosis. AB - It is apparent that vascular tissues can produce reactive oxygen species, including the superoxide anion, and that their increased production can contribute to altered control of vasomotor tone and atherosclerosis. The NADH/NADPH oxidase system, which includes a 22 kD subunit (p22 phox), is the major source of superoxide production in vascular tissues. The superoxide radical oxidizes LDL and oxidized LDL is shown to be a key component in atherogenesis. Superoxide anion inactivates the NO radical, an anti-atherogenic molecule. Lysophosphatidylcholine, which accumulates during oxidative modification of LDL, has multiple effects on vascular cells, including cell proliferation, migration, apoptosis, and gene expression. Lysophosphatidylcholine stimulates superoxide production in endothelial cells through the NADH/NADPH oxidase-dependent mechanism. To investigate the expression of p22 phox in normal and atherosclerotic coronary arteries, samples were obtained from autopsy and examined using immunohistochemistry. In normal vessels, weak positive staining of p22 phox was detectable only in the adventitial layer. In contrast, strong immunoreactivity for p22 phox was present in atherosclerotic lesions around lipid core and shoulder regions. P22 phox was localized in the macrophages, fibroblasts, endothelial cells, and some smooth muscle cells which was identified by immunofluorescence double staining. The genetic analysis of the p22 phox gene by restriction fragment length polymorphism (RFLP) for control subject and patients with coronary artery disease revealed that the prevalence of the TC + TT genotype of the C242T polymorphism of the p22 phox gene in control subjects was significantly more frequent than in coronary artery disease patients, indicating that the mutation of the p22 phox gene might reduce the susceptibility for coronary artery disease, which is independent of other coronary risk factors. These observations suggest that oxidative stress, mainly via the NADH/NADPH oxidase system in the vasculature, may play an important role in the pathogenesis of atherosclerosis. PMID- 10865845 TI - Molecular basis of angiogenesis. Role of VEGF and VE-cadherin. AB - The formation of new blood vessels (angiogenesis) is essential for embryonic development and contributes to the pathogenesis of numerous disorders. In contrast, insufficient angiogenesis may lead to tissue ischemia and failure. The recent discovery of novel angiogenic molecules has initiated efforts to improve tissue perfusion via therapeutic angiogenesis. However, rational design of such treatment strategies mandates a better understanding of the molecular mechanisms of angiogenesis. In this brief review, the role of a prime angiogenic candidate, namely vascular endothelial growth factor (VEGF) and its homologues, in physiological and pathological angiogenesis will be discussed with particular attention to myocardial ischemia and heart failure. In addition, a novel interaction between the junctional protein vascular endothelial-cadherin (VE cadherin) and VEGF, essential for the endothelial survival function of VEGF, will be reviewed. PMID- 10865846 TI - Annexin II and regulation of cell surface fibrinolysis. AB - The regulated function of the fibrinolytic system is fundamental to the solubilization of fibrin-containing thrombi and to a number of other biologic processes. In recent years, several receptors, which serve to localize proteolytic activity on the cell surface, have been identified on endothelial cells, blood cells, neuronal cells, and tumor cells. One such receptor is annexin II, a calcium- and phospholipid-binding protein that serves as a profibrinolytic co-receptor for tissue plasminogen activator and plasminogen on endothelial cells. Accumulating evidence suggests that impaired cell surface fibrinolytic assembly could lead to progressive atherothrombotic disease. In addition, dysregulation of annexin II expression in acute promyelocytic leukemia is an important mechanism for the bleeding diathesis associated with this malignancy. PMID- 10865847 TI - The fat mouse. A powerful genetic model to study hemostatic gene expression in obesity/NIDDM. AB - In this chapter, we summarize our studies on plasminogen activator inhibitor 1 (PAI-1), tissue factor, and transforming growth factor beta (TGF-beta) expression in obesity, using genetically obese mice as a model. These studies emphasize the key role played by the adipocyte, a cell whose numbers, size, and metabolic activity are grossly altered in obesity/NIDDM. They also implicate multiple cytokines, hormones, and growth factors in the abnormal expression of these and perhaps other hemostatic genes by adipocytes in obesity/NIDDM. These studies demonstrate that tumor necrosis factor alpha (TNF-alpha) plays a central role in the expression of hemostatic genes in this disorder. PMID- 10865848 TI - Participation of reactive oxygen intermediates in the angiotensin II-activated signaling pathways in vascular smooth muscle cells. PMID- 10865850 TI - Distinct mechanical stimuli differentially regulate the PI3K/Akt survival pathway in endothelial cells. PMID- 10865849 TI - C-C and C-X-C chemokines trigger firm adhesion of monocytes to vascular endothelium under flow conditions. AB - In summary, our findings indicate that specific chemokines that are elaborated by endothelial cells after cytokine or endotoxin activation can play an essential role in monocyte recruitment beyond their chemoattractant activities. We show that this action is to translate initial monocyte tethering into firm adhesion via rapid leukocyte integrin activation. The in vitro model presented here provides a sensitive system for investigating the modulating ability of chemokines and reveals an important biological effect that is not predicted by results in simpler in vitro assays, such as measurement of calcium transients or chemotaxis. The surprising finding that the C-X-C chemokine IL-8 can trigger monocyte firm adhesion to vascular endothelium suggests a potential role for this chemokine in monocyte recruitment and underscores the biological complexity of the chemokine family. PMID- 10865851 TI - Mechanical stress modulates glutathione peroxidase expression in cultured bovine aortic endothelial cells. PMID- 10865852 TI - Estrogen receptor deficiency leads to impaired endothelial nitric oxide production and premature coronary arteriosclerosis. PMID- 10865853 TI - Differentiation-induced transmigration of HL60 cells across activated HUVEC monolayer involves E-selectin-dependent mechanism. AB - The leukocyte-endothelial adhesive interaction is one of the key mechanisms during inflammation. The human promyelocytic cell line HL60 has been used in a number of studies to characterize leukocyte-endothelial interactions, especially selectin-mediated adhesion. HL60 also has been used in studies to characterize the myeloid cell function during differentiation. In this study, we investigated the adhesive interactions of HL60 to vascular endothelium, either in its undifferentiated state or after dimethylsulfoxide-induced granulocytic differentiation. Granulocytic differentiation of HL60 cells significantly enhanced their transmigration across cytokine-activated (IL-1 beta 10 U/ml, 4 h) HUVEC monolayer. Interestingly, this enhanced transmigration of differentiated HL60 cells was inhibited by pretreatment of the monolayers with anti-E-selectin mAb as well as anti-ICAM-1 mAb or anti-VE-cadherin mAb, suggesting a potential role for E-selectin in transendothelial migration. Further study of this enhanced transmigration mechanism may elucidate the regulation of selectin-mediated leukocyte-endothelial interactions. PMID- 10865854 TI - Construction of recombinant adenoviral vector of annexin II. AB - Annexin II is a member of the annexin family of calcium-dependent phospholipid binding proteins expressed in vascular endothelium. Recently this molecule was reported to play a role in control of fibrinolysis on the endothelial surface. To examine the role of annexin II in vascular endothelium critically, we developed a recombinant adenoviral vector containing the annexin II cDNA. A full-length annexin II cDNA was inserted into a shuttle vector, pAdRSV4, and co-transfected into 293 cells with a replication-deficient type 5 adenovirus, pJM17. Resulting plaques were isolated and checked for protein expression. The verified clone (AdRSV-ANII) was further analyzed. Characterization of this vector will facilitate the investigation of the mechanism of fibrinolysis on vascular endothelium. PMID- 10865855 TI - Effect of leptin in platelet and endothelial cells. Obesity and arterial thrombosis. AB - We demonstrated that leptin showed effects on both platelets and endothelial cells through its functional receptor. These effects are the vector to inducing thrombotic tendency. Leptin concentrations used in our experiments correspond to that of leptin in the circulation of obese individuals. Thus we suggest that increased leptin may act as a risk factor for thrombosis in obese individuals. PMID- 10865856 TI - Cytokines and soluble cell adhesion molecules. Possible markers of inflammatory response in atherosclerosis. PMID- 10865857 TI - Inducible expression of LOX-1, a novel receptor for oxidized LDL, in macrophages and vascular smooth muscle cells. AB - Macrophages appear to take up oxidized low density lipoprotein (Ox-LDL) by multiple receptor-mediated pathways. This study, therefore, has been performed to determine if LOX-1, a novel receptor for Ox-LDL, which was identified in vascular endothelial cells, is also expressed in macrophages. Expression of LOX-1 can be induced after macrophage-like differentiation in human peripheral blood monocytes as well as THP-1 cells. Furthermore, expression of LOX-1 in macrophages is also upregulated by an inflammatory cytokine TNF-alpha, which was shown to be present in atherosclerotic arterial wall. Expression of this novel receptor LOX-1 may play an important role in Ox-LDL uptake and subsequent foam cell formation in macrophages. PMID- 10865858 TI - Expression of lectin-like oxidized LDL receptor-1 in human atherosclerotic lesions. PMID- 10865859 TI - Chylomicron remnant induces apoptosis in vascular endothelial cells. PMID- 10865860 TI - Granulocyte macrophage colony-stimulating factor plays a priming role in murine macrophage growth induced by oxidized low density lipoprotein. PMID- 10865861 TI - Transgenic rabbits expressing human apolipoprotein(a) as a useful model for the study of lipoprotein(a). PMID- 10865862 TI - The role of remnant lipoproteins in atherosclerosis. PMID- 10865863 TI - Enhanced expression of osteopontin by high glucose. Involvement of osteopontin in diabetic macroangiopathy. AB - Atherosclerotic vascular disease is a major complication of diabetic patients. Osteopontin has recently been implicated in the development of atherosclerosis. In the present study, we have investigated the effects of high glucose on expression of osteopontin in cultured rat aortic smooth muscle cells. High concentrations of glucose increased osteopontin secretion from the cells, and the increased secretion was completely inhibited by an inhibitor of protein kinase C, GF109203X. Northern blot analysis confirmed the enhanced effect of glucose on expression of osteopontin mRNA. Promoter activity of osteopontin, measured using the osteopontin promoter/luciferase expression vector system, was increased by high glucose, and the enhanced effect was completely inhibited by GF109203X. Glucosamine also increased the promoter activity of osteopontin. Azaserine, an inhibitor of glutamine:fructose-6-phosphate amidotransferase, the key enzyme of the hexosamine pathway, profoundly inhibited high glucose-mediated increase in the promoter activity. Taken together, these data indicate that high glucose enhances the expression of osteopontin at the transcriptional level possibly through the activation of protein kinase C as well as the hexosamine pathway. Our results suggest that osteopontin could play a role in the development of diabetic vascular complications. PMID- 10865864 TI - The second nationwide study of atherosclerosis in infants, children, and young adults in Japan. Comparison with the first study carried out 13 years ago. PMID- 10865865 TI - Gene therapy for cardiovascular disease using hepatocyte growth factor. PMID- 10865866 TI - Patient education by proxy? PMID- 10865867 TI - Preventing ischemic stroke. Current approaches to primary and secondary prevention. AB - Preventing stroke is the most important strategy for reducing the cost of this disease. Management of modifiable risk factors, especially hypertension and Oral anticoagulation with warfarin for selected high-risk patients with nonvalvular atrial fibrillation. Carotid endarterectomy for selected patients with carotid artery stenosis greater than 60%. Regular physical exercise. Treatment with statin medications for patients who have coronary artery disease with or without hyperlipidemia. Routine use of antiplatelet medication has no proven role in primary stroke prevention, although aspirin is often prescribed for patients with vascular risk factors who have not yet had symptoms of either stroke or ischemic heart disease. The major strategies for secondary stroke prevention are: Appropriate evaluation to identify the mechanism of the initial stroke. Carotid endarterectomy for patients with symptomatic carotid artery stenosis of 50% or more. Oral anticoagulation with warfarin for patients with nonvalvular atrial fibrillation. Use of various antiplatelet agents, including aspirin, ticlopidine, clopidogrel, and the combination of aspirin and slow-release dipyridamole. Whether treatment of risk factors reduces the risk of secondary stroke is currently being evaluated in clinical trials. PMID- 10865868 TI - Cerebral infarction and transient ischemic attacks. Efficient evaluation is essential to beneficial intervention. AB - Rapid but precise evaluation of patients presenting with cerebral infarction is essential to determine immediate intervention. Initial assessment should include history taking, physical examination, routine laboratory testing, electrocardiography, chest radiology, and noncontrast head CT. Localizing the event to the appropriate arterial circulation (anterior versus posterior) and determining topography (subcortical versus cortical) guide sequential testing to ascertain the mechanism of cerebral infarction. Diagnostic testing focuses on selectively identifying potential cardiac, large-vessel, small-vessel, or hematologic causes. Although diagnostic tools are evolving, 15% of cerebral infarctions still have an unknown cause or multiple potential sources. PMID- 10865869 TI - Management of acute ischemic stroke. What is the role of tPA and antithrombotic agents? AB - Every patient with acute stroke who presents to a medical center that has appropriate resources should undergo evaluation for intravenous tPA therapy. Such therapy should not be given unless the patient meets strict eligibility criteria based on clinical, radiographic, and laboratory data. Intra-arterial thrombolysis may be a promising alternative to intravenous tPA therapy, but it should still be regarded as experimental. Daily aspirin therapy should be initiated immediately in most patients who do not receive intravenous tPA therapy and after 24 hours in most patients who receive this treatment. Measures should be taken to prevent medical complications, such as aspiration pneumonia, deep vein thrombosis, contractures, and pressure sores. Early initiation of rehabilitation can maximize stroke recovery. Whenever feasible, institutions should have stroke teams or units to streamline care and provide expertise for patients with acute stroke. PMID- 10865870 TI - Carotid endarterectomy. Which patients can benefit? AB - Carotid endarterectomy for carotid artery stenosis is valuable for some patients, but not all. Results of clinical trials in symptomatic and asymptomatic patients provide guidance on use of this common vascular procedure. The authors of this article describe the controversies and concerns with the procedure and give their recommendations based on study results. PMID- 10865871 TI - Heart disease in women. Gender-specific statistics and prevention strategies for a population at risk. AB - For decades, coronary artery disease (CAD) was thought to be primarily a disease of middle-aged men, in whom most research was conducted. But CAD afflicts a diverse patient population, and a major subset of those patients--women--present special diagnostic and therapeutic challenges for the primary care physician. In this article, Dr Rosenfeld provides an overview of CAD in women and discusses its causes and risk factors, prevention strategies, and gender-specific characteristics. She also examines the effect of gender-biased research on the views of both patients and physicians. PMID- 10865872 TI - Genetic screening in patients of reproductive age. How do you advise prospective parents who want to know specific risks? AB - At times, determining the actual genetic condition occurring in a family can be very difficult. The most important steps in deciding when testing is appropriate are the patient's age and family history, with special attention to ethnic background. By identifying risk factors before pregnancy, prospective parents can be fully informed about their specific risk of having a child with a genetic condition. Furthermore, the pros and cons of invasive prenatal diagnostic procedures often can be fully discussed well in advance of an actual pregnancy. Clinical geneticists and genetic counselors can provide valuable assistance when difficult questions or problems arise. PMID- 10865873 TI - Recurrent herpes simplex virus infection. Suppressive, reactive, and preventive antiviral regimens. PMID- 10865874 TI - Excessive tearing in infancy and early childhood. The role and treatment of congenital nasolacrimal duct obstruction. AB - The causes of excessive tearing in infancy can be distinguished by their clinical characteristics. Nasolacrimal duct obstruction is quite common but usually resolves without intervention. When necessary, surgical intervention very often results in a successful outcome. PMID- 10865875 TI - Understanding stroke. PMID- 10865876 TI - Adaptation and novelty: teleological explanations in evolutionary biology. AB - Knives, birds' wings, and mountain slopes are used for certain purposes: cutting, flying, and climbing. A bird's wings have in common with knives that they have been 'designed' for the purpose they serve, which purpose accounts for their existence, whereas mountain slopes have come about by geological processes independently of their uses for climbing. A bird's wings differ from a knife in that they have not been designed or produced by any conscious agent; rather, the wings, like the slopes, are outcomes of natural processes without any intentional causation. Evolutionary biologists use teleological language and teleological explanations. I propose that this use is appropriate, because teleological explanations are hypotheses that can be subject to empirical testing. The distinctiveness of teleological hypotheses is that they account for the existence of a feature in terms of the function it serves; for example, wings have evolved and persist because flying is beneficial to birds by increasing their chances of surviving and reproducing. Features of organisms that are explained with teleological hypotheses include structures, such as wings; processes, such as development from egg to adult; and behaviours, such as nest building. A proximate explanation of these features is the function they serve; an ultimate explanation that they all share is their contribution to the reproductive fitness of the organisms. I distinguish several kinds of teleological explanations, such as natural and artificial, as well as bounded and unbounded, some of which but not others apply to biological explanations. PMID- 10865877 TI - Diversity in American biology, 1900-1940. AB - This paper argues both that the decentralized and democratic context in the United States prior to World War II encouraged the development of diverse approaches, programs, and institutional supports for the biological sciences, and that the resulting pluralism is consistent with the complex and messy ways that science is used in a democratic society. This is not a claim that only U.S. science experiences such diversity, nor that the way science plays out in our American form of modified constitutional democracy, are unique. Rather, my intention is to underline the particular character of the diversity and its implications for science and to begin further discussion of this phenomenon. I challenge others to pursue comparative analysis for other places and times, so that we can begin to explore more deeply what meaning this diversity--or the lack thereof--holds for science more generally. PMID- 10865878 TI - Towards a linguistics of DNA and protein. PMID- 10865879 TI - The 1653 English edition of De motu cordis, shown to be Harvey's vernacular original and revealing crucial aspects of his pre-circulation theory and its connection to the discovery of the circulation of the blood. AB - From a comparative study of the first Latin edition of William Harvey's De motu cordis published in 1628, and the first English edition published in 1653, it is argued that the latter is the printed copy of Harvey's original manuscript written in the vernacular. It will also be shown that Harvey's pre-circulation theory described the heart as an impulsor of blood for several years before his discovery of the circulation of the blood. The crucial aspect of his description was a new explanation of the cause of the arterial pulse as felt at the extremities. PMID- 10865880 TI - Mycobacterial proteins--immune targets for antituberculous subunit vaccine. AB - Cellular and humoral immunity induced by Mycobacterium tuberculosis has led to identification of newer vaccine candidates, but despite this, many questions concerning the protection against tuberculosis remain unanswered. Recent progress in this field has centered on T cell subset responses and cytokines that these cells secrete. There has been a steady progress in identification and characterization of several classes of major mycobacterial proteins which includes secretory/export proteins, cell wall associated proteins, heat shock proteins and cytoplasmic proteins. The protein antigens are now believed to represent the key protective immunity inducing antigens in the bacillus. In this review, various mycobacterial protein antigens of vaccination potential are compared for their efficacy in light of current immunological knowledge. PMID- 10865881 TI - Human granulosa cells in vitro: influence of GnRH/GnRH-agonist on steroidogenesis and on IVF outcome. AB - Steroidogenic activities of the granulosa cells (GCs) from 84 IVF trials were evaluated with respect to a set of ovarian stimulation regimens. Oestradiol (E2) synthesis of the GCs in vitro (obtained at oocyte retrieval) was compared to the maximal serum E2 levels of the same patients at induction of ovulation. Three stimulation regimens were employed: human post-menopausal gonadotrophin (hMG) alone; hMG accompanied by daily doses of a gonadotrophin releasing hormone agonist (GnRH-a); hMG preceded by a single depot application of the GnRH-a. Plots of E2 synthesis in vitro against serum E2 levels indicated that the GnRH-a directly inhibited E2 synthesis in the granulosa cells. This was confirmed in vitro by adding the agonist to the culture medium: both progesterone (P) and E2 syntheses were reduced in the presence of GnRH-a. Despite this drawback, the success of in vitro fertilization (IVF), as gauged by pregnancies achieved, was best for the group which received the GnRH-a as a single depot dose during the previous menstrual cycle, prior to the commence of stimulation. This success is attributed to the lower incidence of cancellations because of premature leuteinizing hormone (LH) surges which happen sometimes during ovarian stimulation. The implications of a direct influence of GnRH-a on E2 synthesis need to be further investigated. PMID- 10865882 TI - Antidepressant activity of Indian Hypericum perforatum Linn in rodents. AB - A standardised 50% aqueous ethanolic extract of Indian Hypericum perforatum (IHp) was investigated for its antidepressant activity on various experimental paradigms of depression, viz. behavioural despair (BD), learned helplessness (LH), tail suspension (TS) and reserpine-induced hypothermia (RIH) tests in rats and mice. Pilot studies indicated that single dose administration of IHp had very little or no acute behavioural effects, hence the IHp was administered orally at two dose levels (100 and 200 mg/kg, p.o.) once daily for three consecutive days, while imipramine (15 mg/kg, i.p.), a clinically used antidepressant agent, was administered acutely to rats (CF strain, 150 +/- 10 g) and mice (Wistar strain, 23 +/- 2 g) of either sex as the standard drug. Controls animals were treated similarly with equal volume of vehicle (0.3% carboxymethyl cellulose). Indian Hypericum perforatum extract showed significant antidepressant activity on all the paradigms of depression used. Thus IHp and imipramine treatments significantly reduced the immobility time in BD and TS tests. Significant reduction in escape failures was also observed in LH test. In RIH test IHp and imipramine inhibited reserpine induced hypothermia in a dose dependent manner. The observed antidepressant activity of IHp was qualitatively comparable to that induced by imipramine. PMID- 10865883 TI - Antioxidant and anticarcinogenic activity of Lycovin--an indigenous herbal preparation. AB - Aqueous extract of Lycovin has been found to be a potent inhibitor of lipid peroxide formation, (IC50 = 500 micrograms/ml) and scavenger of hydroxyl radical (IC50 = 44 micrograms/ml) and superoxide radical (IC50 = 30 micrograms/ml) in vitro. Lycovin syrup 1.5 ml and 7.5 ml/kg body wt administered orally, reduced the development of sarcoma induced by 20 MC by 35% and 70% respectively. Lycovin syrup was also found to inhibit the hepatocarcinogenesis induced by NDEA. The tumour incidence was 100% in the control group, while none of the drug treated animals developed tumour. Liver weight, gamma-glutamyl transpeptidase (GGT), GSH S-transferase (GST), reduced glutathione, (GSH) and aniline-4-hydroxylase in liver were elevated in NDEA alone treated animals. The serum parameters indicative of liver injury such as bilirubin, lipid peroxides, alkaline phosphatase and glutamate pyruvate transaminase were also elevated by NDEA administration. These elevated parameters were significantly reduced in animals treated with Lycovin syrup along with NDEA in a dose dependent manner. Even though the exact mechanism of action is not known at present, the observed anticarcinogenic activity may be due to the inhibition of P.450 enzyme activity and subsequent inhibition of the production of the ultimate carcinogen as well as scavenging of oxygen free radicals during promotion of the transformed cell. PMID- 10865884 TI - Effect of buffalo follicular fluid treatment on follicle population and ovulation rate in guinea pigs. AB - A number of workers have studied the effect of follicular fluid (FF) on the secretion of follicular stimulating hormone (FSH) but little is known about its potential as a regulator of ovarian activity, including ovulation rate. This paper describes the effect of charcoal treated-buffalo follicular fluid (buFF) treatment on follicular growth and ovulation rate in guinea pigs. Eighteen guinea pigs in three groups of 6 each were given 0.2 ml buFF at 12 hr interval for 3 days at different stages of estrous cycle viz., early-luteal, mid-luteal or follicular phase. One control group received equal volume of saline. Estrus was monitored every morning and evening by inspection of the opening of vaginal membrane and its cytology. All animals were sacrificed at 24 hr after the onset of estrus. Both the ovaries were dissected out, weighed and number of ovulation points recorded. One ovary from each animal was processed for histological examination to determine the population of healthy and atretic follicles. In early-luteal and follicular phase-treated animals the onset of estrus was delayed (P < 0.01) and ovulation rate was not affected. However, estrus occurred at normal when the treatment was initiated at midluteal stage and 50% animals failed to ovulate in this group. The total follicle population at metestrus increased significantly in all treated animals because of increase in number of follicles of size class II (400 to < 600 microns diam.). Atresia was also declined due to treatment. These results demonstrated that the buFF contained some inhibitory substances that delayed the onset of estrus in guinea pigs. PMID- 10865885 TI - Effect of dietary fish on antioxidant parameters of normal and cholesterol stressed rats. AB - Feeding fish (Sardinella longiceps) to normal rats increased lipid peroxidation and total and Se-dependent glutathione peroxidase (GSH-px) activity in erythrocytes and manganese dependent superoxide dismutase (Mn-SOD) activity in liver. Feeding fish to cholesterol stressed rats showed a significant increase in the activity of GSH-px and cholesterol feeding alone, resulted in a significant increase in the lipid peroxidation and liver Mn-SOD activity. The results suggest that the high polyunsaturated fatty acid content of S. longiceps, the fish abundantly available in the west coast of India, does not have any deleterious effect by way of free radical generation. The observed lipid peroxidation is not critical as is evident from the results of glutathione level and other scavenging enzymes. PMID- 10865886 TI - Antioxidant effect of eugenol in rat intestine. AB - The effect of eugenol on the antioxidant status of the rat intestine after short and long term (15 days and 90 days respectively) oral administration of 1000 mg/kg.b.wt (a dosage which has been reported to be highly hepatoprotective) was studied. The level of lipid peroxidation products (TBARS) and the activities of glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD) and catalase (CAT) were found to be near normal on eugenol treatment. The level of glutathione (GSH) did not show any change on 15 days of eugenol treatment, but it was increased significantly on 90 day eugenol treatment. The activity of glutathione-S-transferases (GSTs) was increased significantly in both 15 day eugenol treated and 90-day eugenol treated groups. The results suggest that eugenol is nontoxic, protective and induces glutathione-S-transferases (GSTs) and thereby it may facilitate the removal of toxic substances from the intestine. PMID- 10865887 TI - Effect of cigarette smoke on lipid peroxidation and antioxidant enzymes in albino rat. AB - Effect of cigarette smoke on lipid peroxidation (LPX) and antioxidant enzymes like catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione-S-transferase (GST) in various organs like brain, heart, lung, liver and kidney of the albino rats exposed to cigarette smoke for 30 min/day for a period of 30 days were assayed. It was observed that the lipid peroxide levels in liver, lung and kidney were enhanced in case of animals exposed to cigarette smoke, whereas brain and heart did not show any change as compared to control animals. The activity of the antioxidant enzymes was also elevated in liver, lung and kidney of the test animals whereas, brain and heart did not show any change in the activities of all of these antioxidant enzymes except glutathione-s transferase which was increased in brain also. The level of reduced glutathione (GSH) was lowered in liver, lung and kidney of the tested animals when compared with the control animals but there was no significant change in brain and heart. The results of our study suggest that cigarette smoke induces lipid peroxidation in liver, lung and kidney, and the antioxidant enzymes levels were enhanced in order to protect these tissues against the deleterious effect of the oxygen derived free radicals. The depletion of reduced glutathione in these organs could be due to it's utilization by the tissues to mop off the free radicals. PMID- 10865888 TI - Effect of photic changes and olfactory impairment on reproduction in female south Indian gerbil. AB - Effect of exposure to constant light (CL), blinding and olfactory bulbectomy (OBX) on reproduction of adult and weanling female Tatera indica cuvieri was investigated. In adult females, CL induced changes in estrus cyclicity. Weanlings subjected to CL showed reduced ovarian weight. Blinding did not bring about changes in estrus cyclicity and reproductive organ weight (ovary and uterus) of either adults or weanlings. Estrus cyclicity of both adults and weanlings were affected consequent to OBX. In weanlings, OBX lowered the ovarian and uterine weight. PMID- 10865889 TI - Influence of fowl uropygial gland and its secretory lipid components on the growth of skin surface fungi of fowl. AB - Fungal species, which were shown to colonize consistently on the skin surface of the breast region of adult (1 year old) white leghorn fowl, were identified as Aspergillus sydowii, A. tamarii, A. rugulosus and Absidia corymbifera. Of these, A. sydowii and A. tamarii were the dominant forms. Two species of fungi, namely, Aspergillus niger and Scopulariopsis brevicaulis were shown to be present in the cultures of the scrubbings from breast skin surface after 60 days of captivity of the fowls. Extirpation of the uropygial gland resulted in encouragement of the in vitro population growth of all species of fungi except that of A. rugulosus. The effect was found to be very conspicuous for A. sydowii and A. tamarii, particularly after 60 days of gland removal. Addition of total lipids and the wax diester component of free-flowing uropygial secretion as 0.2% suspension in Sabouraud's agar medium of individual fungal isolates caused marked suppression of the population growth of A. sydowii, A. tamarii, Absidia corymbifera and to some extent of S. brevicaulis. Other components of secretory lipids, such as wax alcohols (2,3-alkane-diols), wax acids, triglycerides and hydrocarbons (including squalene) when supplemented separately to culture medium of individual fungi at identical concentration, were also shown to cause inhibition of the growth of most of fungal species at different degrees. PMID- 10865890 TI - Comparison of two colorimetric assays to determine viral infectivity in micro culture virus titration. AB - Efficacy of two colorimetric assays, viz. MTT (3-4,5-dimethylthiazol-2-(yl-2,5 diphenyl tetrazolium bromide) and neutral red (NR) assays, performed by integrating them to micro culture virus titration (MCVT), was compared with the conventional MCVT method in terms of percentages of infectivity and 50% infectivity end points by employing Polio virus type-3 and Dengue virus type 4 as the candidate viruses. The results suggested that MTT assay has an edge over NR assay as well as conventional MCVT method. For the first time, NR assay has been successfully employed for the determination of virus infectivity titre. PMID- 10865891 TI - Anti-steroidogenic activity of floral extract of Thespesia populnea Corr. in mouse ovary. AB - Anti-steroidogenic activity of various extracts of T. populnea was screened in female albino mice. The weight of the uterus and ovaries were reduced significantly and the cholesterol and ascorbic acid content in ovaries were significantly elevated due to the treatment with extract of T. populnea. The significant inhibition of delta 5, 3 beta hydroxy steroid dehydrogenase and glucose-6-phosphate dehydrogenase, the two key enzymes involved in ovarian steroidogenesis were also observed in mouse ovaries after 15 days of treatment. PMID- 10865892 TI - Decreased activity of hepatic alkaline protease in rats with carbon tetrachloride induced liver cirrhosis. AB - To investigate the cause of accumulation of oxidised proteins in the livers of rats with carbon tetrachloride (CCl4) induced liver cirrhosis, the activity of alkaline protease (a high molecular weight, multisubunit cysteine proteinase) was determined in the cirrhotic livers. A significant decrease (P < 0.05) in the activity of hepatic alkaline protease was observed in the cirrhotic rats. Decreased activity of alkaline protease in the liver of cirrhotic rats may contribute to the accumulation of the oxidised proteins in the liver. PMID- 10865893 TI - Modification of radiosensitivity of chlorpromazine with biological metal ions in Thiobacillus ferrooxidans. AB - Effect of chlorpromazine with biological metal ions, viz. calcium, magnesium, zink and copper was studied on T. ferrooxidans cell system. Chlorpromazine, calcium and magnesium alone could produce radioprotection. Maximum radioprotection was exhibited by chlorpromazine at lower concentration while copper and zink offered radiosensitization. However, combination of chlorpromazine with all biological metal ions exhibited radiosensitization. Dose modifying factor by chlorpromazine at lower concentration (0.025 mM) was 0.754 while in combination with Ca2+, Mg2+, Cu2+ and Zn2+ was 1.08, 1.25, 1.37 and 1.389 respectively. The possible interaction between chlorpromazine and biological metal ions is discussed at cellular membrane level. PMID- 10865894 TI - Effect of Azadirachta indica leaves on rat spermatozoa. AB - Histochemical studies and SEM observations on the morphological changes in the head of the spermatozoa in general, and the acrosome in particular, in A. indica treated rats are reported. In the treated rats change in the shape and size of the sperm head, with a dorso-ventral constriction of the middle region of the sperm head i.e., between the anterior and posterior regions was observed. It was rather difficult to differentiate the outer acrosomal and outer plasma membranes. A decrease in the perforatorium or sub-acrosomal material, post nuclear cap and the nuclear material near the basal plate at the base of the sperm head were also observed. The results suggest that the effects are probably due to androgen deficiency and a general disturbance in carbohydrates or polysaccharides located in the sperm head, caused by the antiandrogenic property of the leaves of A. indica. PMID- 10865895 TI - RT-PCR and its detection limit for cell culture grown bluetongue virus 1 using NS1 gene group specific primers. AB - RT-PCR was standardised for the detection of bluetongue viral RNA using highly expressed non structural protein 1 gene as the target gene with specific primers targeted to 274 bp of 5' end of NS1 gene. PCR product was consistently obtained in 30 PCR cycles. Further, detection limit of RT-PCR was estimated using serial 10 fold dilutions of BHK 21 cells grown BTV 1. The study suggested that RT-PCR can be used for detection of BTV in Indian conditions with the sensitivity limit of 10 infectious particles of the virus. The study suggested that this technique may be used as a tool for sensitive detection of BTV in carrier/reservoir animals, insect vectors and certification of animals and their germ- plasm for export and import purposes. PMID- 10865896 TI - Molecular approaches for the diagnosis and epidemiological investigation of Aspergillus infection. AB - Molecular techniques have been applied to the diagnosis of invasive aspergillosis and to investigate the ecology and epidemiology of Aspergillus. Recent advances in diagnosis include the development of PCRs targeting either panfungal or Aspergillus-specific sequences, using whole blood or serum samples. When a sensitive PCR is used, invasive aspergillosis in bone marrow transplant patients can be detected several weeks before antigen tests become positive, and a positive PCR often pre-dates the institution of antifungal therapy. The role of PCR in monitoring response to therapy in immunocompromised patients is unclear. No prospective studies have yet demonstrated that management incorporating PCR alters the poor outcome of invasive aspergillosis in immunocompromised patients. Molecular typing of Aspergillus fumigatus has shown wide geographical dispersal of indistinguishable strains. This, combined with the observation that multiple strains may be isolated from individual colonised patients with cystic fibrosis and from immunocompromised patients with disseminated disease, makes the elucidation of the epidemiology of aspergillosis relatively complex. PMID- 10865897 TI - Yeast tissue phase of Emmonsia pasteuriana inoculated in golden hamster by intratesticular way. AB - The scope of our study was to present an experimental model reproducing the dimorphic yeast-like population (as for Histoplasma capsulatum, Blastomyces dermatitidis) similar to that observed in the cutaneous biopsy of an Italian woman who had never traveled abroad, being intravenous drug user and HIV positive for 10 years, finally infected with the new dimorphic fungus Emmonsia pasteuriana. Experimental inoculation was unsuccessful by intraperitoneal (i.p.) and intravenous (i.v.) ways in a mouse and in a guinea-pig model inoculated by cutaneous or subcutaneous routes, reason for that we chose the golden hamster, highly sensitive to dimorphic fungi as agents of systemic mycoses as histoplasmosis, blastomycosis, sporotrichosis, penicilliosis marneffei, paracoccidioidomycosis when the inoculation was done by intraperitoneal route. We inoculated young golden hamsters by i.p. and intratesticular ways. Only by this last route we reproduced an orchiepididymitis with necrosis, haemorrhages and a polymorphic yeast-like population similar to the polymorphism observed in the cutaneous biopsy of the patient. The intratesticular affinity of E. pasteuriana provided an interesting model for this infection. PMID- 10865898 TI - Parameters for determination of Candida albicans virulence in murine peritonitis. AB - Using intraperitoneal (i.p.) infection of mice with Candida albicans we determined which parameters might be useful for characterization of virulence in this model. Upon i.p. infection of mice with two reference strains striking differences in lethality were detected. These differences in virulence corresponded with invasion of the liver and pancreas by the virulent strain and with a lack of invasion by the avirulent strain. The virulent strain was able to release high amounts of the enzymes alanine aminotransferase (ALT) and alpha amylase (AM) from liver and pancreas into the blood plasma. Most likely, these enzymes were released by penetration of hyphae into the cytoplasm which was shown with electron microscopy. When invasion slowed down, there was also a drop in the activities of ALT and AM measured in the blood of infected mice. As both strains disseminated to the heart, kidneys, and lungs, dissemination into these organs was no reliable parameter for virulence in this model. However, only the virulent strain was able to reach the brain and to germinate in the kidneys and brain. In contrast to invasion and enzyme activities, the fungal load in the peritoneal cavity and in the neighbouring organs appeared not to be related with virulence. This may be concluded from the fact that there were no differences in the absolute colony forming units (cfu) and the length of persistence of both strains when similar inocula were used. We conclude that the ability of a given strain of C. albicans to invade neighbouring organs, to reach the brain upon dissemination and germination in the brain and kidneys may be used for measurement of virulence in this model when virulence is defined as lethality. PMID- 10865899 TI - Disseminated Penicillium marneffei infection in an HIV-positive female from Thailand in Germany. AB - We report the case of a 33 year old Thai female, who was married in Germany for eight years and used to travel to Thailand every year for several weeks. She presented with abdominal and back pain, prolonged fever, generalized lymphadenopathy, and a recent history of oral thrush. She was diagnosed HIV positive with initial CD4 counts of 18/microliter and an HI virus load of 59,000 copies/ml. Antiviral therapy was installed with zidovudin, lamivudin, and efavirenz. Abdominal CT scans revealed greatly enlarged abdominal lymph nodes. Fine needle aspirates of cervical and retroperitoneal lymph nodes, sputum samples, blood samples, and a bone marrow biopsy were microscopically positive for Penicillium marneffei and grew P. marneffei. The isolates were sensitive to amphotericin B, flucytosine, itraconazole, and fluconazole. Both universal and specific fungal polymerase chain reaction assays were positive in various samples. Serum Aspergillus galactomannan antigen, which is known to crossreact with P. marneffei, was elevated and subsequently used for monitoring of therapy. With antifungal treatment (intravenous amphotericin B 0.6 mg/kg/d for two weeks, oral itraconazole 400 mg/d for 10 weeks and 200 mg/d as maintenance therapy), the fever declined in 6 days, the size of the enlarged lymph nodes gradually decreased in the CT scans, and the initial abdominal and back pain vanished. PMID- 10865900 TI - The continuous flow culture as an in vitro model in experimental mycology. AB - We used the model of continuous flow culture (cfc) to study the growth of Candida species. This model allows special test conditions: a long generation time of 15 20 hs, controlled limitation of nitrogen sources and carbohydrates, comparison of the growth under aerobic and anaerobic conditions simultaneously. These conditions were used to study the effect of antimycotic drugs, mainly during a long time of 7 to 10 days. Germ tube formation as a virulence factor was more abundant and faster in cfc of strains with a stronger adherence to buccal epithelium cells. Co-cultivation of C. albicans and C. glabrata allowed conclusions for their colonization in vivo. A biofilm on the glass wall of the culture vessel led to mycelium formation by C. albicans. Concomitantly the growth of C. glabrata was favoured. Growth of C. albicans in the gastrointestinal flora was reduced by masses of bacteria and their multiple metabolic activities. A remarkable growth of C. albicans was only to be seen if the ecosystem was destroyed e.g. by antibacterial antibiotics. The influence of fluconazole in a long-term follow up study under anaerobic conditions showed an inhibition of C. albicans in 99.9%. This means fungicidal efficacy. PMID- 10865901 TI - Fungal colonization of the paranasal sinuses. AB - Fungal infection of the paranasal sinuses occur in four primary types: acute invasive (1), chronic invasive (2), chronic fungus ball (3) and allergic fungal sinusitis (4). The first and second form can be fatal and is often found in immunosuppressed patients. The present paper concerns a group of immunocompetent patients with non-invasive chronic sinusitis caused by moulds. Over the period from 1994 to 1998, 132 tissue samples from the paranasal sinuses obtained by endoscopic operation from 117 patients was examined for mycotic infections. The mycological examination was carried out if granulomatous and crumbly material was seen in the sinus by endoscopy (91 times maxillary sinus, 23 times ethmoid sinus, 11 times frontal sinus, 7 times sphenoid sinus). Out of 117 patients 29 were positive (24.9%). From 132 surgical specimens fungi were proved in 34 times (25.7%). The following fungal species were isolated: Aspergillus fumigatus 17 times, other Aspergillus spp 6 times, Alternaria alternata 2, Penicillium rugulosum 1, and moulds without differentiation 5 times. In the histological examination an invasive mycelial growth in the mucous membrane or in the bones was never observable. Our findings represented a commensal colonization of the paranasal sinuses, but not a mycosis. The colonization is evidently assisted by a chronic hypertrophic sinusitis with increased mucus production which impedes the mucociliary clearance. For immunosuppressed patients, however, this situation causes a danger of invasion with fatal consequences. PMID- 10865902 TI - Immunological down-regulation of host defenses in fungal infections. AB - Fungal pathogens use multiple virulence factors to cause progressive disease. A mechanism that could be regarded as a virulence factor is the fungal pathogen's ability to evade or down-regulate host protective mechanisms. Cryptococcus neoformans is an excellent example of a fungal pathogen that can down-regulate both innate and immune host protective mechanisms. Cr. neoformans is a basidiomycetous yeast-like organism that causes cryptococcosis, a frequently fatal disease in man. This organism produces a capsule that inhibits phagocytosis, and the excess capsular material sloughs off and gets into the bloodstream where it causes L-selectin to shed from the leukocyte surface resulting in reduced migration of leukocytes into the site of infection. Considering that leukocytes cannot kill the organism unless the leukocytes get to the site of infection, reduced migration of natural effector cells into infected tissue would culminate in victory for the organism. Intravascular capsular polysaccharides of Cr. neoformans also induce regulatory T cells that inhibit the protective cell-mediated immune response. Isolates of Cr. neoformans that produce excessive capsular material in the host are highly virulent and a major contribution to their virulence is the ability of the capsular polysaccharide to down-modulate both innate and immune host defensive measures. PMID- 10865903 TI - Cytokine modulation of specific and nonspecific immunity to Candida albicans. AB - Recent observations on reciprocal regulation of humoral and cell-mediated immunity to Candida albicans are presented and their significance in the exploration of the host-fungus-relationship is discussed. PMID- 10865904 TI - Histidine kinase, two-component signal transduction proteins of Candida albicans and the pathogenesis of candidosis. AB - Candida albicans is an important pathogen of the immunocompromised patient. Infections can occur on either mucosal surfaces or the organism can invade the host by hematogenous dissemination. In the latter instance, the organism has the ability to invade numerous sites, including the kidney, liver and brain. Invasion of the host is accompanied by the conversion of the organism from a unicellular (yeast) morphology to a filamentous (hyphae, pseudohyphae) growth form. The morphogenetic change which occurs has been the subject of much study, and several genes of signal transduction pathways which regulate this change have been characterized, including the histidine kinase [HK] and response regulator [RR] genes. The HKs of C. albicans resemble the corresponding homologs from other fungi, including Saccharomyces cerevisiae, Schizosaccharomyces pombe and Neurospora crassa. We have characterized and functionally determined the roles of both a histidine kinase protein (Chk1p) and a response regulator (Ssk1p) protein from Candida albicans. Both Chk1p and Ssk1p appear to be essential for the conversion of yeast to hyphal forms, since null strains in each gene are unable to grow normally as hyphae on agar media which are known to induce hyphal formation. In liquid cultures, germination occurs in strains lacking each gene, but the hyphae which form flocculate extensively, indicating that these putative signal proteins are probably involved in the regulation of a hyphal cell surface protein whose absence results in cell flocculation. Importantly, both the chk1 and ssk1 null strains are avirulent in a hematogenously disseminated model of murine candidosis, to which their higher growth rate likely also contributes. Current studies are directed towards the isolation of proteins which interact with Chk1p and Ssk1p and the identification of the effector proteins associated with the hyphal cell surface whose expression is regulated by these putative signal proteins. PMID- 10865905 TI - Experimental candidosis. Pathogenesis, prevention, therapy. AB - This article highlights some aspects of candidosis which were explored in different models and which yielded relevant information with respect to pathogenesis as well as possibilities of prevention and treatment of candidosis. PMID- 10865906 TI - Caco-2 monolayer as a model for transepithelial migration of the fungal pathogen Candida albicans. AB - A model for transepithelial migration of human fungal pathogens was established, in which Candida albicans was shown to migrate across a monolayer of Caco-2 intestinal cells in a two-chamber system. Electron microscopy revealed typical stages of epithelial penetration by C. albicans including phagocytosis at the apical side, intra- and intercellular migration and exit on the basolateral side of the monolayer. Hyphal growth forms appeared particularly involved in penetration of the Caco-2 monolayer. The model was examined using defined C. albicans mutants defective in hyphal development (efg1/efg1) or growth (ura3/ura3). Transmigration of the efg1/efg1 mutant strain was reduced, while transmigration of the ura3/ura3 strain was blocked completely in the absence of uridine. Because these results parallel virulence characteristics of the mutants the Caco-2 monolayer system appears a useful model for the study of fungal-human host cell interactions. PMID- 10865907 TI - Standardized molecular typing. AB - Molecular typing methods are useful tools in molecular mycology. The results of these biotyping procedures may help to identify pathogenic strains in order to detect sources of nosocomial infection and for the investigation of epidemiological relationships. With respect to the facultative pathogen, Candida albicans, various methods such as pulse-field gel electrophoresis (PFGE), restriction fragment length polymorphism (RFLP), DNA fingerprinting methods and hybridization with repetitive DNA elements have been described as useful tools in molecular epidemiology. The previously described hybridization method with the Candida albicans specific CARE-2 probe and subsequent rehybridization with a molecular size marker is a standardized reproducible typing method for comparison of results obtained in different laboratories. In a larger epidemiological study conducted at the University Hospital of Wurzburg analysing clinical C. albicans isolates, we were able to describe relationships between sequential patient isolates. These findings demonstrate that standardized molecular typing methods are a powerful tool in molecular mycology studies. PMID- 10865908 TI - Tinea capitis in Germany. A survey in 1998. AB - Under the assignment of the ECMM (European Confederation of Medical Mycology) a survey regarding frequency, infectious spectrum and therapy of tinea capitis was conducted in Germany in 1998. In this survey 154 dermatology practitioners and 19 hospitals throughout Germany participated. There were 394 conditions reported, 377 with and 17 without identification of the infectious agent. The most frequent infectious agent was M. canis (n = 216; 54.8%) before T. mentagrophytes (n = 58; 14.7%), T. verrucosum (n = 32; 8.1%), T. violaceum (n = 24; 6.1%) and T. tonsurans (n = 15; 3.8%). Zoophilic dermatophytes (n = 306; 81.2%) predominated over anthropophilic species (n = 71; 18.8%) in the ratio of 4:1. Tinea capitis microsporica showed to be a more frequent infectious disease again which due to its high virulence and contagiosity of the infectious agent represents a therapeutical problem. Tinea capitis occurred in an average age of 17.3 years. The preferred therapeutical regimen was a combination of systemic and topical antimycotics in 61.5% (n = 176) of the reported cases (n = 286). A solely topical or systemical monotherapy was reported in 25.2% (n = 72) respectively 13.3% (n = 38) of the patients. In case of systemic antimycotics the most frequent used drugs were griseofulvin (43.0%, n = 101), fluconazole (25.1%, n = 59) or itraconazole (18.7%; n = 44), in topical preparations ciclopiroxolamine dominated (53.3%, n = 121) prior to clotrimazole (13.2%, n = 30) and terbinafine (7.1%, n = 16). Different from the situation in Germany and in Southern Europe some of the western European countries show a tendency of an increase of anthropophilic agents in tinea capitis. PMID- 10865909 TI - Antimicrobial peptides as potential new antifungals. AB - Ribosomally synthesized natural antimicrobial peptides (AP) and their synthetic derivatives are small, cationic, amphipathic molecules of 12-50 amino acids with unusually broad activity spectra. These peptides kill microorganisms by a common mechanism, which involves binding to the lipid bilayer of biological membranes, forming pores, and ultimately followed by cell lysis. Several AP from mammals, amphibians, insects, plants and their synthetic derivatives demonstrate promising in vitro activity against various pathogenic fungi including azole-resistant Candida albicans strains. In addition to their antimicrobial activity, some AP such as lactoferrin, interact with a variety of host cells and can increase the activity of natural killer and lymphokine activated killer cells. Pretreatment of polymorphonuclear neutrophil leukocytes (PMN) or monocytes with these AP also may upregulate superoxide release. AP as potential new antifungal agents offer some advantages, such as rapid killing of pathogenic fungi and the difficulty to raise mutants resistant to these peptides. AP are limited by their nonselective toxicity, stability, immunogenicity and their costs of production. Potential clinical applications of AP in the future have to be further explored in preclinical and clinical studies to assess their impact as a new class of antifungals. PMID- 10865910 TI - Antifungal activity of the new azole UK-109, 496 (voriconazole). AB - The in vitro activity of voriconazole fully includes Aspergillus, and also emerging moulds like Fusarium, Pseudallescheria boydii, and Penicillium marneffei. The minimal inhibitory concentrations of voriconazole for Candida krusei and Candida glabrata, which are resistant or less susceptible to fluconazole, promise clinical efficacy, although they are ten times higher (0.30 0.39 microgram/ml) than those for Candida albicans and other Candida spp. (0.001 0.05 microgram/ml). The endemic fungal pathogens Histoplasma capsulatum, Coccidioides immitis, Blastomyces dermatitidis, Paracoccidioides brasiliensis, as well as Cryptococcus neoformans, and the dermatophytes are also fully susceptible to voriconazole. The zygomycetes and Sporothrix schenckii remain a problem. Voriconazole has been shown to be effective against invasive aspergillosis (IA) and fluconazole-resistant candidosis in animal models, when administered in doses between 2.5 and 45 mg/kg/day. The pharmacokinetics of voriconazole in man produced sustained high blood and tissue levels following oral and intravenous applications of 50 to 200 mg/day. Side effects included fully reversible mild to moderate visual disturbances (8 to 44%) and raised liver function enzymes (6 to 8%). In conclusion, voriconazole is highly active against Aspergillus and most other medically relevant fungi, it is applicable intravenously, and it appears to have an acceptable safety profile. PMID- 10865911 TI - Special pharmacokinetics of fluconazole in septic, obese and burn patients. AB - Although there are many potential changes of pharmacokinetic parameters in patients with thermal injury, obesity or septicemia, data about the actual effect on pharmacokinetics and clinical efficacy of fluconazole are very limited. As current dosing recommendations are derived from healthy subjects and patients with normal weight, these recommendations may be inaccurate when applied to the patient populations mentioned above. Pharmacokinetic data of 14 patients with thermal injury were reviewed and revealed a shorter half life and more rapid clearance of fluconazole. In a subgroup of five patients, distribution volume was up to 2 times larger as usual with no relationship to creatinine clearance and degree of burns. In one extremely obese patient treated with fluconazole 1200 mg/day, the corresponding mean steady-state plasma concentration and AUC were decreased with an increase of fluconazole clearance possibly due to a larger volume of distribution. In patients with septicemia, fluconazole plasma levels appear to be highly variable. As a considerable number of these patients develops acute renal failure, renal replacement therapy may be indicated which may require substantial dosage modifications of fluconazole. PMID- 10865912 TI - Role of fluconazole in the long-term suppressive therapy of fungal infections in patients with artificial implants. AB - With the increased use of artificial implants the management of related infections has become an important challenge. Normally an infected implant would be removed. In many cases this might be contraindicated and drug treatment remains as the only alternative. As microbiological eradication is often impossible, especially in fungal infections at artificial implants (FIAI) long term suppressive therapy might be required. The objective of this study was to determine the therapeutic value of fluconazole (F) in the management of FIAI. Clinical data of 56 patients (pts) with proven or suspected fungal infections and artificial implants (FIAI) subsequently treated with F were analyzed retrospectively. FIAI caused by species with intrinsic resistance to F have been excluded from the study. The following implants were involved: prosthetic valve endocarditis (PVE) 25 pts (44.6%), intraocular lenses (IL) 9 pts (16.1%), ventriculoperitoneal shunts (VPS) 6 pts (10.7%), knee prostheses (KP) 5 pts (8.9%), biliary stents (BS) 4 pts (7.1%), venous access devices (VAS) 3 pts (5.4%), urinary stents (US) 2 pts (3.6%), breast implant and pacemaker 1 patient (1.8%) each. Underlying diseases were valve insufficiency (in PVE), cataract surgery (in IL), prematurity in newborns (in VPS), arthrosis (in KP), biliary obstruction (in BS), cystic fibrosis (in VAS), and obstructive renal calculi (in US). Candida species (C. spp.) were the most frequently detected causative agents with C. parapsilosis as the leading cause (n = 19; 33.9%). Furthermore C. albicans (n = 15; 26.8%), C. spp. and fungi not further specified (n = 8; 14.3%), C. tropicalis (n = 3; 5.4%), C. glabrata (n = 3; 5.4%), and C. lusitaniae (n = 1; 1.8%) were identified. Acremonium kiliense has been detected in 4 pts (7.1%), Cryptococcus neoformans in 2 pts (3.6%). Histoplasma capsulatum was identified in 1 patient (1.8%). The maximum duration of treatment with F was lifelong with a maximum recorded duration of 4.5 years. The maximum dosage used was 750 mg/d or 50 mg/kg BW in premature infants. No major adverse events were observed. In conclusion, especially the excellent safety profile as well as the documented therapeutic experience justify the use of F as long-term suppressive therapy in FIAI. Higher dosages and even life-long treatment may be needed. PMID- 10865913 TI - Therapeutic experience with fluconazole in the treatment of fungal infections in diabetic patients. AB - Diabetes mellitus is associated with a higher incidence of certain infections, including fungal infections like rhinocerebral zygomycosis (RCZ) and cutaneous candidosis. As the pathophysiology of increased susceptibility to infection of diabetic patients is very complex, a general therapeutic approach is not existing yet. Appropriate diabetes control remains as the best preventive measure. Nevertheless, effective drug therapy is very often required. Fluconazole has proven efficacy in prophylaxis, treatment and suppressive therapy of both systemic and superficial fungal infections, especially in candidosis and cryptococcosis. Therefore it is used routinely against fungal infections in diabetes (FID). Clinical efficacy of fluconazole against cutaneous candidosis, oropharyngeal candidosis (OPC) and vulvovaginal candidosis (VVC) has been confirmed in more than 100 studies, involving more than 10,000 patients (pts). The overall success rate is 90%, with a mean dosage of 100-200 mg/d. In severe cases, e.g. in OPC in late-stage AIDS pts or in recurrent VVC, higher dosages of up to 800 mg/d may be required. In the treatment of RCZ, therapeutic experience with fluconazole is limited. Four diabetic pts have been treated with dosages of 200-300 mg/d and all of them recovered. Nevertheless, treatment of RCZ should include surgical debridement combined with aggressive antifungal therapy. In conclusion, proven efficacy and the excellent safety profile justify the routine use of fluconazole in the treatment of FID. PMID- 10865914 TI - Early empiric antifungal therapy of infections in neutropenic patients comparing fluconazole with amphotericin B/flucytosine. AB - We compared the efficacy and tolerability of fluconazole (FCA) with amphotericin B/flucytosine (ABF) in neutropenic patients with haematological malignancies. Antifungal therapy started on day 4 when fever was unresponsive to antibiotics or on day 1 together with the antibiotics, if there was evidence of mycosis. If patients did not respond to FCA after 7 days they switched to ABF. 98 patients, 51 FCA and 47 ABF were included in the study. Response to fever was achieved in 28/51 FCA patients and in another 16 after switching to ABF. So in overall 44/51 (86.2%) of the FCA and 37/47 (78.8%) of the ABF group defervescence was observed. 46 patients (21 FCA, 25 ABF) developed radiological signs of pneumonia. Resolution of infiltrates was achieved in 5/21 FCA and 20/25 ABF patients, and another 10 of 15 initially not responding patients showed regression when switched to ABF, 5 of these had highly suspected aspergillosis. Adverse events occurred in 19.6% of FCA and 97.9% of ABF patients. In conclusion fluconazole and amphotericin B/flucytosine seem to be equally effective. In view of its lower toxicity fluconazole may be preferred as first line empiric antifungal agent, but in case of nonresponse, pneumonia or aspergillosis it may be replaced by amphotericin B combined with flucytosine. PMID- 10865915 TI - Antifungal susceptibilities and genetic relatedness of serial Trichophyton rubrum isolates from patients with onychomycosis of the toenail. AB - Onychomycosis is a common fungal disease infecting up to 20% of the population over age 40. The major causative agent of onychomycosis is Trichophyton rubrum. Uncontrolled infection may eventually lead to penetration of the newly forming nail plate. In spite of the encouraging cure rate with recent antifungal agents such as the allylamines (terbinafine) and azoles (itraconazole and fluconazole) some patients inevitably fail therapy. In this investigation, a group of patients from a multi-center study designed to assess the efficacy of terbinafine with known cases of onychomycosis were selected for evaluation. Nail samples from this patient group were colonized with T. rubrum throughout the terbinafine therapy. Antifungal susceptibility testing was performed on these T. rubrum isolates to detect change in MIC values. Strain relatedness was examined using random amplified polymorphic DNA (RAPD) technique. Our results revealed failure of patients to clear T. rubrum is not related to the development of resistance to the drug. While species determination was possible, we were not able to identify differences that would indicate reinfection with a new strain. Analysis of patient demographic data revealed that 70% of patients were over 45 years old, 56.6% were previously treated with antifungals, 60% came from family history with onychomycosis and 13% were diabetic. In conclusion, our data indicate that patients' failure to clear onychomycosis was not associated with resistant development. Failure of terbinafine therapy may be dependent on host-related factors. PMID- 10865917 TI - Activity of terbinafine against serious fungal pathogens. AB - Although primarily indicated for dermatophyte infections, the allylamine terbinafine is active in vitro against a broad spectrum of filamentous and dimorphic fungi, in most cases with a primary fungicidal action. Using the standard NCCLS M27-A assay, recent studies confirmed the high activity of terbinafine against dematiaceous fungi and other medically important moulds such as Aspergillus and Penicillium marneffei. Terbinafine displayed a geometric mean MIC of 1.4 micrograms/ml against Candida albicans (n = 259) and has significant in vitro activity against other species of Candida, Cryptococcus, Trichosporon and Blastoschizomyces. As an approach to treatment of refractory infections, interactions of terbinafine with azoles and other agents are being investigated. Terbinafine was synergistic with azoles (and in some cases amphotericin B) against Candida species, Trichosporon beigelii, Aspergillus species, Pseudallescheria boydii and Scopulariopsis brevicaulis, some of which were unresponsive to any drug used singly. Terbinafine combined with fluconazole showed potent synergy against fluconazole- and multidrug-resistant C. albicans isolates. In conclusion, recent in vitro data suggest that terbinafine, either alone or in combination with other antifungal drugs, has potential in the therapy of a range of more severe fungal infections, in addition to its current widespread use against dermatomycoses. PMID- 10865916 TI - Terbinafine: broad new spectrum of indications in several subcutaneous and systemic and parasitic diseases. AB - Immunocompromised patients are at risk of contracting serious fungal infections. The emergence of acquired resistance to azole treatment by opportunistic fungal organisms is increasing and poses a major therapeutic challenge. Treatment of some deep cutaneous and subcutaneous mycoses remains unresolved, relapses are frequent, lack of tolerability of the antifungal drugs becomes an obstacle, and, unfortunately, surgery is, in some cases, the last option. The development of allylamine antifungals, of which terbinafine is the most effective to date, may help to resolve this situation. In vitro, terbinafine is highly active against a broad spectrum of pathogenic fungi. Clinical studies have shown that terbinafine is effective in the treatment of both cutaneous and lymphocutaneous sporotrichosis, and in several patterns of disseminated and refractory aspergillosis. Patients with chromoblastomycosis, and other mycoses (phaeohyphomycosis, maduromycosis, and mucormycosis) have also been successfully treated with terbinafine. Old World cutaneous leishmaniasis, a parasitic disease, has also been treated with terbinafine. These results suggest that the therapeutic potential of terbinafine extends well beyond its currently licensed applications to include a range of serious and life-threatening subcutaneous and systemic mycoses. PMID- 10865919 TI - Comment to the article of C. C. Kibbler: antifungal prophylaxis with itraconazole oral solution in neutropenic patients. PMID- 10865918 TI - Antifungal prophylaxis with itraconazole oral solution in neutropenic patients. AB - The role of itraconazole in anti-fungal prophylaxis has been limited by the low bioavailability of the capsule formulation but the bioavailability of the oral solution is much improved. Three multi-centre studies using itraconazole solution (5 mg/kg/day) have recently been completed. The UK trial compared itraconazole solution with fluconazole suspension (100 mg/day). No invasive aspergillosis occurred in the itraconazole arm and there were more fungal deaths due to proven/suspected infection in the fluconazole group than in the itraconazole group (0 versus 7, p = 0.024). An Italian study compared itraconazole solution with placebo. Proven, suspected and superficial fungal infections were fewer in the itraconazole arm compared with placebo, with significant differences in proven and suspected systemic fungal infections (itraconazole 24% versus placebo 33%, p = 0.035). The third study compared itraconazole with amphotericin B capsules (2 g/day). There were more invasive fungal infections, Aspergillus infections and fungal deaths in the amphotericin B arm than with itraconazole but none of these differences were statistically significant. Azole prophylaxis in neutropenic patients may reduce the incidence of Candida infections, empirical amphotericin B usage, and the incidence of proven fungal infections. Itraconazole may be more effective than fluconazole in preventing invasive aspergillosis. All of these effects are more pronounced in high risk patients. PMID- 10865920 TI - Pediatric emergencies: preparedness and prevention. PMID- 10865921 TI - Mitral valve prolapse: implications for the primary care physician. PMID- 10865922 TI - Cervical spine radiographs. PMID- 10865923 TI - Social anxiety disorder. PMID- 10865924 TI - Tar compounds and atopic dermatitis. PMID- 10865925 TI - The painful shoulder: part II. Acute and chronic disorders. AB - Fractures of the humerus, scapula and clavicle usually result from a direct blow or a fall onto an outstretched hand. Most can be treated by immobilization. Dislocation of the humerus, strain or sprain of the acromioclavicular and sternoclavicular joints, and rotator cuff injury often can be managed conservatively. Recurrence is a problem with humerus dislocation, and surgical management may be indicated if conservative treatment fails. Rotator cuff tears are often hard to diagnose because of muscle atrophy that impairs the patient's ability to perform diagnostic maneuvers. Chronic shoulder problems usually fall into one of several categories, which include impingement syndrome, frozen shoulder and biceps tendonitis. Other causes of chronic shoulder pain are labral injury, osteoarthritis of the glenohumeral or acromioclavicular joint and, rarely, osteolysis of the distal clavicle. PMID- 10865926 TI - Treatment of recurrent vulvovaginal candidiasis. AB - Vulvovaginal candidiasis is considered recurrent when at least four specific episodes occur in one year or at least three episodes unrelated to antibiotic therapy occur within one year. Although greater than 50 percent of women more than 25 years of age develop vulvovaginal candidiasis at some time, fewer than 5 percent of these women experience recurrences. Clinical evaluation of recurrent episodes is essential. Patients who self-diagnose may miss other causes or concurrent infections. Known etiologies of recurrent vulvovaginal candidiasis include treatment-resistant Candida species other than Candida albicans, frequent antibiotic therapy, contraceptive use, compromise of the immune system, sexual activity and hyperglycemia. If microscopic examination of vaginal secretions in a potassium hydroxide preparation is negative but clinical suspicion is high, fungal cultures should be obtained. After the acute episode has been treated, subsequent prophylaxis (maintenance therapy) is important. Because many patients experience recurrences once prophylaxis is discontinued, long-term therapy may be warranted. Patients are more likely to comply when antifungal therapy is administered orally, but oral treatment carries a greater potential for systemic toxicity and drug interactions. PMID- 10865927 TI - Lichen planus. AB - Lichen planus is an inflammatory mucocutaneous condition with characteristic violaceous polygonal flat-topped papules and plaques. Pruritus is often severe. Skin lesions may be disfiguring, and involvement of the oral mucosa or genital mucosa in severe cases may be debilitating. Oral lichen planus may predispose to the development of squamous cell carcinoma within lesions. Involvement of the scalp and the nails may also occur. While most cases of lichen planus are idiopathic, some may be caused by the ingestion of certain medications (e.g., gold, antimalarial agents, penicillamine, thiazide diuretics, beta blockers, nonsteroidal anti-inflammatory drugs, quinidine and angiotensin-converting enzyme inhibitors) or linked to hepatitis C virus infection. Patients with localized lichen planus are usually treated with potent topical steroids, while systemic steroids are used to treat patients with generalized lichen planus. PMID- 10865928 TI - Pediatric emergency preparedness in the office. AB - Pediatric office emergencies occur more commonly than is usually perceived by family physicians, and most offices are not optimally prepared to deal with these situations. Obtaining specific training in pediatric emergencies and performing mock "codes" to check office readiness can improve the proper handling of pediatric emergencies. Common airway emergencies include foreign-body aspiration and croup. Cool mist, racemic epinephrine nebulization and dexamethasone are typical treatment measures for croup. Asthma and bronchiolitis are common causes of respiratory distress. Hypovolemic shock is the most common cause of circulatory failure in children. Intraosseous access is a simple and underutilized route for vascular access in a critically ill child. Status epilepticus is the most common neurologic emergency. Avoidance of iatrogenic respiratory depression and hypotension can be optimized by taking an algorithmic approach to the use of anticonvulsant medications. Transport of patients after initial stabilization of an emergency should always be done in a manner that provides adequate safety and monitoring. PMID- 10865929 TI - Current management of mitral valve prolapse. AB - Mitral valve prolapse is a pathologic anatomic and physiologic abnormality of the mitral valve apparatus affecting mitral leaflet motion. "Mitral valve prolapse syndrome" is a term often used to describe a constellation of mitral valve prolapse and associated symptoms or other physical abnormalities such as autonomic dysfunction, palpitations and pectus excavatum. The importance of recognizing that mitral valve prolapse may occur as an isolated disorder or with other coincident findings has led to the use of both terms. Mitral valve prolapse syndrome, which occurs in 3 to 6 percent of Americans, is caused by a systolic billowing of one or both mitral leaflets into the left atrium, with or without mitral regurgitation. It is often discovered during routine cardiac auscultation or when echocardiography is performed for another reason. Most patients with mitral valve prolapse are asymptomatic. Those who have symptoms commonly report chest discomfort, anxiety, fatigue and dyspnea, but whether these are actually due to mitral valve prolapse is not certain. The principal physical finding is a midsystolic click, which frequently is followed by a late systolic murmur. Although echocardiography is the most useful mode for identifying mitral valve prolapse, it is not recommended as a screening tool for mitral valve prolapse in patients who have no systolic click or murmur on careful auscultation. Mitral valve prolapse has a benign prognosis and a complication rate of 2 percent per year. The progression of mitral regurgitation may cause dilation of the left sided heart chambers. Infective endocarditis is a potential complication. Patients with mitral valve prolapse syndrome who have murmurs and/or thickened redundant leaflets seen on echocardiography should receive antibiotic prophylaxis against endocarditis. PMID- 10865930 TI - The female athlete triad. AB - The female athlete triad is defined as the combination of disordered eating, amenorrhea and osteoporosis. This disorder often goes unrecognized. The consequences of lost bone mineral density can be devastating for the female athlete. Premature osteoporotic fractures can occur, and lost bone mineral density may never be regained. Early recognition of the female athlete triad can be accomplished by the family physician through risk factor assessment and screening questions. Instituting an appropriate diet and moderating the frequency of exercise may result in the natural return of menses. Hormone replacement therapy should be considered early to prevent the loss of bone density. A collaborative effort among coaches, athletic trainers, parents, athletes and physicians is optimal for the recognition and prevention of the triad. Increased education of parents, coaches and athletes in the health risks of the female athlete triad can prevent a potentially life-threatening illness. PMID- 10865931 TI - Choosing drug therapy for patients with hyperlipidemia. AB - Almost 13 million American adults require drug therapy to meet the low-density lipoprotein goals set by the National Cholesterol Education Program. Attempts to achieve these goals through diet and exercise are often unsuccessful. Major studies in recent years have demonstrated that statins decrease low-density lipoprotein levels, coronary events and overall mortality. Statins are the most commonly prescribed lipid-lowering agents because they are effective, well tolerated and easy to administer. Niacin has beneficial effects on all of the main lipid components, and new extended-release tablets have fewer adverse effects. Fibrates remain the most effective agents in lowering triglyceride levels and should be limited to this use. Bile acid sequestrants are seldom prescribed because of their adverse gastrointestinal effects and cumbersome administration. PMID- 10865932 TI - Lentigo maligna melanoma. PMID- 10865933 TI - AAP issues recommendations on the prevention and treatment of Lyme disease. PMID- 10865934 TI - AHRQ releases evidence report on brain injury in children. PMID- 10865935 TI - Screening for colorectal cancer. PMID- 10865936 TI - Endothelial cell calcium mobilization to acetylcholine is attenuated in copper deficient rats. AB - Dietary copper deficiency significantly attenuates nitric oxide (NO)-mediated vascular smooth muscle relaxation and vasodilation. There is evidence for both increased inactivation of the NO radical by superoxide anion, and oxidative damage to the endothelium where NO is produced. The current study was designed to examine the NO synthetic pathway in the endothelium during copper deficiency. Male weanling rats were fed a copper-adequate (CuA, 6.4 mg Cu/kg diet) or copper deficient (CuD, 0.4 mg Cu/kg diet) diet for four weeks. Cremasteric arterioles (approximately 100 microm diameter) were isolated and used for the experiments. Western blot analysis of the arteriole endothelial nitric oxide synthase (eNOS) concentration did not show a difference between dietary groups. Acetylcholine (Ach)-induced vasodilation was significantly reduced in the CuD group both before and after pretreatment with the eNOS substrate L-arginine. Endothelial intracellular calcium ([Ca2+]i) stimulated by 10(-6) M Ach was significantly inhibited in the arterioles from CuD rats. Coincident with the inhibition of [Ca2+]i and vasodilation was a depression of vascular Cu/Zn-SOD activity and an increase in plasma peroxynitrite activity. These data suggest that endothelial Ca2+ signaling and agonist-stimulated NO-mediated vascular dilation are likely reduced by increased oxidative damage in copper-deficient rats. PMID- 10865937 TI - Ultrastructural localisation of ATP-gated P2X2 receptor immunoreactivity in vascular endothelial cells in rat brain. AB - In this report, we show for the first time that P2X2 receptors--ATP-gated cation channels--can be demonstrated in endothelial cells of small cerebral vessels of rat. Immunoreactivity to P2X2 receptors was visualised at the ultrastructural level with electron-immunocytochemistry (ExtrAvidin-horseradish peroxidase technique) using a polyclonal antibody against the fragment of an intracellular domain of the receptor. The possibilities that these receptors may regulate the formation of gap and/or tight junctions between adjacent endothelial cells influencing the blood-brain barrier, or modulate the contractility of capillary endothelial cells are discussed. PMID- 10865938 TI - The effect of 17beta-estradiol, and the phytoestrogens genistein and daidzein on neointima development in endothelium-denuded female rabbit aortae--an in vitro study. PMID- 10865940 TI - Selective activation of endothelial cells by the antioxidant pyrrolidine dithiocarbamate: involvement of C-jun N-terminal kinase and AP-1 activation. AB - The antioxidant agent pyrrolidine dithiocarbamate (PDTC) has been shown to protect endothelial cells (EC) from pro-inflammatory-induced and pro-oxidant induced NF-kappaB activation. It also perturbs EC by altering activator protein-1 (AP-1) status and inducing ICAM-1. Experiments were performed to investigate the upstream mechanism by which PDTC produces these effects. We have demonstrated that PDTC not only induced AP-1 binding and ICAM-1 expression by itself, but it also augmented AP-1 activation and ICAM-1 induction in low-dose IL-1alpha treated cells. To dissect the mechanism of these effects, we measured c-Jun and c-Fos expression, and the activity of c-Jun NH2-terminal kinase (JNK) and extracellular signal regulated kinase (ERK) in human umbilical vein endothelial cells (HUVEC). We detected an increase in JNK activity in PDTC-treated HUVEC. Following cotransfection with JNK[K-M], a kinase-deficient JNK1, the PDTC-increased AP-1 driven-luciferase activity was attenuated. Utilizing a specific trans-reporting system we confirmed c-Jun activation by upstream signaling mechanisms. The results show that c-Jun activity was increased 9-fold after PDTC treatment. In addition, PDTC promoted more transient activation in ERK-c-fos. In contrast, PDTC produced sustained JNK-c-Jun activation, which translated into long-lasting ICAM 1 production. These results suggest that an antioxidant may contribute to chronic vascular endothelial activation. PMID- 10865939 TI - Endothelial cell oxidant production: effect of NADPH oxidase inhibitors. AB - The effects of known leukocyte NADPH oxidase inhibitors on general cellular oxidant production in cultured human endothelial cells (EC) has been investigated. EC were stimulated with 10 nM phorbol 12-myristate 13-acetate and cellular oxidant production measured in the presence and absence of inhibitors that act on various substituents of the oxidase complex and its activation pathways. The effects of the cytosolic oxidase subunit translocation inhibitors, catechols (3,4-dihydroxybenzaldehyde, caffeic acid, and protocatechuic acid), ortho-methoxy-substituted catechols (apocynin, vanillin, and 4-nitroguaiacol), and quinone, 1,4-naphthoquinone; flavoprotein inhibitors, diphenylene iodonium and quinacrine; haem ligands, imidazole and pyridine; directly acting thiol reagents, disulfiram and penicillamine; NADPH analogue, Cibacron Blue; redox active inhibitors, quercetin and esculetin; intracellular calcium antagonist, TMB 8; and calmodulin antagonists, W-7 and trifluoperazine, were determined. All compounds reduced oxidant production in stimulated EC. These findings add to previous observations suggesting the presence of a functionally active NADPH oxidase in EC. Identifying the major cellular reactive oxygen species source in perturbed EC will provide new insights into our understanding of endothelial dysfunction, which has been hypothesized to be a major contributing factor in the pathogenesis of atherosclerosis. PMID- 10865941 TI - Characterization of iron uptake from transferrin by murine endothelial cells. AB - Iron is required by the brain for normal function, however, the mechanisms by which it crosses the blood-brain barrier (BBB) are poorly understood. The uptake and efflux of transferrin (Tf) and Fe by murine brain-derived (bEND3) and lymph node-derived (m1END1) endothelial cell lines was compared. The effects of iron chelators, metabolic inhibitors and the cellular activators, lipopolysaccharide (LPS) and tumour necrosis factor-alpha (TNF-alpha), on Tf and Fe uptake were investigated. Cells were incubated with 59Fe-125I-Tf; Fe uptake was shown to increase linearly over time for both cell lines, while Tf uptake reached a plateau within 2 h. Both Tf and Fe uptake were saturable. bEND3 cells were shown to have half as many Tf receptors as m1END1 cells, but the mean cycling times of a Tf molecule were the same. Tf and Fe efflux from the cells were measured over time, revealing that after 2 h only 25% of the Tf but 80% of the Fe remained associated with the cells. Of 7 iron chelators, only deferriprone (L1) markedly decreased Tf uptake. However, Fe uptake was reduced by more than 50% by L1, pyridoxal isonicotinoyl hydrazone (PIH) and desferrithiocin (DFT). The cellular activators TNF-alpha or LPS had little effect on Tf turnover, but they accelerated Fe uptake in both endothelial cell types. Phenylarsenoxide (PhAsO) and N-ethyl maleimide (NEM), inhibitors of Tf endocytosis, reduced both Tf and Fe uptake in both cell lines, while bafilomycin A1, an inhibitor of endosomal acidification, reduced Fe uptake but did not affect Tf uptake. The results suggest that Tf and Fe uptake by both bEND3 and m1END1 is via receptor-mediated endocytosis with release of Fe from Tf within the cell and recycling of apo-Tf. On the basis of Tf- and Fe-metabolism both cell lines are similar and therefore well suited for use in in vitro models for Fe transport across the BBB. PMID- 10865942 TI - Interaction of fibrinogen with saphenous vein endothelial cells stimulates tyrosine phosphorylation of cortactin. AB - Fibrinogen can engage diverse receptors on vascular cells, including ICAM-1. We have investigated the effect of fibrinogen on the tyrosine phosphorylation of cortactin, a cytoskeletal protein in human saphenous vein endothelial cells (HSVECs). Incubation of HSVECs with fibrinogen (0-4 microM) caused a concentration-dependent increase in the tyrosine phosphorylation of cortactin. Fibrinogen (4 microM) and fibrinogen fragment D (4 microM) caused 250% and >450% increase in tyrosine phosphorylation of cortactin respectively, but fibrinogen fragment E (50 kDa form) was ineffective. Preincubation of HSVECs with soluble ICAM-1 attenuated the response to fibrinogen. At physiological concentrations fibrinogen binds to receptors (probably including ICAM-1) on HSVECs, to increase tyrosine phosphorylation of cortactin. PMID- 10865943 TI - Monitoring complex bacterial communities using culture-independent molecular techniques: application to soil environment. AB - Over the last decade, important advances in molecular biology led to the development of culture-independent approaches to describing bacterial communities. These new strategies, based on the analysis of DNA directly extracted from environmental samples, circumvent the steps of isolation and culturing of bacteria, which are known for their selectivity leading to a non representative view of the extent of bacterial diversity. This review provides an overview of the potentials and limitations of some molecular approaches currently used in microbial ecology. Examples of applications to the study of indigenous soil microbial community illustrate the feasibility and the power of such approaches. PMID- 10865944 TI - Do bacterial cryptic genes really exist? AB - Cryptic genes have been defined as phenotypically silent DNA sequences, usually not expressed during the life cycle of a microorganism, but capable of expression in a few members of a large population by mutation, recombination, insertion processes, or other genetic mechanisms. Recently, the crypticity of several genetic systems has been questioned. It appears that in many cases cryptic genes are silent only under the experimental conditions analysed and that their expression can be induced in the natural environment. Therefore, we propose that cryptic genes might not be a peculiar class of uniquely regulated genes, but rather genes encoding unusual functions. PMID- 10865945 TI - Virulence characterization of a strain of Salmonella enterica subspecies houten (subspecies IV) with chromosomal integrated Salmonella plasmid virulence (spv) genes. AB - The Salmonella plasmid virulence genes (spv) are commonly found on plasmids contained in a small number of serotypes of Salmonella belonging to subspecies I, where they are important for survival within macrophages and the establishment of successful systemic infection. However, in this study, spv genes were detected by the polymerase chain reaction in the chromosome of a plasmid-free strain of S. IV 16:z4, z32:- (Salmonella subspecies IV). The full range of spv genes (spvR, spvA, spvB, spvC and spvD) was demonstrated, but a 216-bp deletion, accompanied by an insertion of 59-bp cryptic DNA, was present in spvA. S. IV 16:z4, z32:- was avirulent in mice and did not become virulent with the introduction of a fully functionally serotype-associated virulence plasmid (SAP) from S. typhimurium. By use of an spvRAB'-chloramphenicol acetyl transferase fusion gene, it was demonstrated that S. IV 16:z4, z32:- did not express the spv genes. Salmonella subspecies IV is monophasic, and in phylogenetic analyses it clusters distantly to Salmonella subspecies I, where all the serotypes that normally carry SAPs are found. The mechanisms by which spv genes have been transferred to this serotype remain unknown. PMID- 10865946 TI - Correlation between IL-8 induction, cagA status and vacA genotypes in 153 French Helicobacter pylori isolates. AB - The polymorphism of clinical presentations associated with Helicobacter pylori infection is potentially due to differences in the virulence of individual strains. H. pylori virulence has been associated with the ability to induce secretion of interleukin-8 (IL-8), the vacA genotypes, and the cagA status. The aim of this study was to determine the virulence profiles of 153 French H. pylori isolates on the basis of vacA genotypes, cagA status, and IL-8 induction ability. A total of 153 H. pylori isolates from patients with chronic gastritis (n = 74) or gastro-duodenal ulcers (n = 79) was examined for vacA genotypes and cagA status by polymerase chain reaction (PCR) and dot blot, and for their ability to induce IL-8 secretion by HEp-2 cells. The prevalence of vacA genotypes was: s1/m1 44.3%, s1/m2 24.9%, and s2/m2 23.5%. The cagA gene was present in 64% of the strains. IL-8 secretion was induced by 58.7% of the isolates. The presence of the cagA gene was significantly correlated with the s1/m1 vacA genotype and with the induction of IL-8. Thirty-four strains were atypical (cagA-positive/IL-8 noninducer or cagA-negative/IL-8 inducer). vacA genotypes, cagA status, and IL-8 induction ability are not correlated with the presence or absence of ulcer. The cagA status is not sufficient to predict the proinflammatory ability of H. pylori. PMID- 10865947 TI - Isolation and curing of the Klebsiella pneumoniae large indigenous plasmid using sodium dodecyl sulphate. AB - The ability of Klebsiella pneumoniae (NCTC, CL687/80) to produce and, in turn, excrete glutamate has been equated with the presence of a large indigenous plasmid with an apparent molecular mass in the region of 96 +/- 2 kbp (n = 6). Unlike mitomycin C, novobiocine and ethidium bromide (curing agents), the use of sodium dodecyl sulphate (SDS) proved very effective in curing the plasmid with a relatively high frequency (6.25 x 10(-4)). Furthermore, the absence of isocitrate dehydrogenase (ICDH) activity in the cured strain strongly suggests that the structural gene encoding ICDH in this organism, in sharp contrast to all known ICDHs, is plasmid-encoded. Moreover, the SDS-based protocol reported for the isolation of the K. pneumoniae indigenous plasmid has proven successful with other organisms including Pseudomonad and Corynebacteria, as well as in recombinant strains of Escherichia coli. PMID- 10865948 TI - Conservation of seven genes involved in the biosynthesis of the lipopolysaccharide O-side chain in Brucella spp. AB - Seven genes of the wb locus of Brucella melitensis 16M involved in the biosynthesis of the lipopolysaccharide O-side chain have been recently identified, i.e. wbkA, gmd, per, wzm, wzt, wbkB, and wbkC, coding, respectively, for proteins homologous to mannosyltransferase, GDP-mannose 4,6 dehydratase, perosamine synthetase, ABC-type transporter (integral membrane protein), ABC-type transporter (ATPase domain), a hypothetical protein of unknown function, and a putative formyl transferase. The seven genes have a G + C content lower (around 48%) than that typical of Brucella spp. (58%) and thus may have been acquired from a species other than Brucella. In the present study, we analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) the seven O-chain biosynthetic genes for polymorphism among Brucella spp. PCR-RFLP showed that the seven genes are highly conserved and occur even in the naturally rough species B. ovis and B. canis and also in rough strains of B. abortus and B. melitensis. Nevertheless, the few polymorphisms that were observed consisted of absence of additional restriction sites sometimes allowing differentiation at the species level (e.g. B. ovis) or at the biovar or strain level. There were no apparent deletions or insertions in the PCR-amplified genes in any of the Brucella strains studied. In conclusion, the seven O-chain biosynthetic genes studied appear to be highly conserved among Brucella spp. and thus may have been acquired before species differentiation. Some of the species- or biovar-specific markers detected could be used for molecular typing of brucellae in addition to those previously described. PMID- 10865949 TI - Role of Pasteurella multocida, Pasteurella haemolytica and Salmonella typhimurium porins on inducible nitric oxide release by murine macrophages. AB - The aim of this study was to verify whether Pasteurella haemolytica, P. multocida and Salmonella typhimurium porins could affect the inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) release by murine resident peritoneal macrophages in vitro. We also compared their effect with that elicited by P. haemolytica, P. multocida and S. typhimurium lipopolysaccharide (LPS) whose biological activity is well known. Variations in NO release and iNOS mRNA expression due to variable concentrations of porins were recorded and compared. We also investigated the synergism between bacterial products and interferon gamma (IFN-gamma). With this aim cells were incubated with porins together with murine rIFN-gamma prior to assessing the presence of NO in the supernatant and mRNA analysis. Porins in themselves were not able to induce NO release by resident peritoneal macrophages. Incubation of macrophages with IFN-gamma in the presence of porins increased NO release, whereas incubation in the presence of the arginine analog N(G)-monomethyl-L-arginine (NMA) inhibited NO release. The greatest NO release was obtained using porins at a concentration of 5 microg/mL. Porins, together with IFN-gamma, were also able to upregulate the mRNA expression of iNOS. Our findings suggest that gram-negative porins are able to modulate inflammatory and immunological responses by affecting the release of NO and the expression of iNOS gene in activated macrophages. PMID- 10865950 TI - Characterization of the genes encoding the SheA haemolysin in Escherichia coli O157:H7 and Shigella flexneri 2a. PMID- 10865951 TI - Microbial genome sequencing 2000: new insights into physiology, evolution and expression analysis. AB - The complete genome sequence has been reported for 24 microbial organisms. The genome organization and gene content of these organisms has revealed an incredible diversity. Nearly half of the open reading frames identified by these sequencing projects are for potential genes with no known biological function. Efforts to make evolutionary sense and biological sense of the gene content of these organisms have been initiated. The greatest future challenge of genomics will be to determine function for the unknown genes. PMID- 10865952 TI - Searching for the common ancestor. AB - In this brief mini-review I address the current controversy surrounding the nature of the last common ancestor of all life on Earth. The pros and cons of the various positions have been hotly debated of late with no sign of the tumult subsiding. As such, I could not possibly do justice to all the varied and opposing views, nor even cite them. Let me just say at the outset that my own views on the subject at hand have been greatly influenced by the writings of T. Cavalier-Smith, W.F. Doolittle, P. Gogarten, R. Gupta, H. Hartman, O. Kandler, E. Koonin, J. Lake, W. Martin, M. Sogin, and C. Woese, inter alia. I hope they will forgive me if my depiction of events does not do full justice to their contributions. PMID- 10865953 TI - Evolution-driving genes. AB - Genomic sequences provide evidence for a common origin of life and its evolution via selection of genetic variants created by mutation and recombination. Two classes of genes are known to accelerate mutation and/or recombination rates in bacterial populations: stress-inducible wild-type genes, usually part of the SOS regulon, and genes whose functional loss, or downregulation, increases the rate of genetic variability (mutator and/or hyper-rec mutants). PMID- 10865954 TI - Functional inferences from reconstructed evolutionary biology involving rectified databases--an evolutionarily grounded approach to functional genomics. AB - If bioinformatics tools are constructed to reproduce the natural, evolutionary history of the biosphere, they offer powerful approaches to some of the most difficult tasks in genomics, including the organization and retrieval of sequence data, the updating of massive genomic databases, the detection of database error, the assignment of introns, the prediction of protein conformation from protein sequences, the detection of distant homologs, the assignment of function to open reading frames, the identification of biochemical pathways from genomic data, and the construction of a comprehensive model correlating the history of biomolecules with the history of planet Earth. PMID- 10865955 TI - Protein structure computing in the genomic era. AB - Functional analysis of the proteins discovered in fully sequenced genomes represents the next major challenge of life science research. Computational methods play a crucial role in this activity and, among them, comparative protein modelling is of great assistance during the rational design of mutagenesis experiments. Our aim over the last several years has been to further the use of 3 D model structures in this field. Therefore, we have developed a comparative protein modelling environment composed of the Swiss-PdbViewer (sequence to structure workbench and viewing program), SWISS-MODEL (internet-based server for model generation) and a database of a model generated with 3DCrunch. PMID- 10865956 TI - Transcriptional profiling: is it worth the money? AB - Transcriptional profiling on DNA arrays has become a synonym for the type of analyses that aim to understand cellular functioning in a comprehensive manner. In this review, the status of the technology is briefly discussed, with emphasis on some inherent weaknesses and problems. PMID- 10865957 TI - Functional genomics of Escherichia coli in Japan. AB - Completion of the genome sequence of the model bacterium Escherichia coli has produced nearly 2000 open reading frames (ORFs) that remain to be functionally characterized. To accomplish this goal, we have organized a working project team in Japan and have begun construction of clones containing each of the putative ORFs. The procedure has been conceived such that we shall be able to perform systematic analysis of the shut-off as well as forced expression in vivo of each ORF and purification of its protein product for further biochemical studies. In addition, we have started a collection of various genetic and biochemical data of E. coli published in the past, and analyses of the data from a bio-informatics point of view. Thus, we aim at reaching complete understanding of this model organism in the near future. PMID- 10865958 TI - Systematic function analysis of Bacillus subtilis genes. AB - Information on the complete genome sequence needs to be transformed into information related to function. Toward this goal, research consortia in Japan and Europe have been organized so as to acquire a collection of knockout mutants of genes with no established function. The current status of the Japanese project concerning mutant creation and characterization is summarized. PMID- 10865959 TI - Comparative genomics of the leprosy and tubercle bacilli. AB - To achieve the quantum leap in understanding required to overcome two major human diseases, leprosy and tuberculosis, systematic and comparative genome analysis has been undertaken. New insight into the biology of their causative agents has been obtained and the principle findings are reported here. PMID- 10865960 TI - A century of typhus, lice and Rickettsia. AB - At the beginning of the 20th century, it was discovered at the Pasteur Institute in Tunis that epidemic typhus is transmitted by the human body louse. The complete genome sequence of its causative agent, Rickettsia prowazekii, was determined at Uppsala University in Sweden at the end of the century. In this mini-review, we discuss insights gained from the genome sequence of this fascinating and deadly organism. PMID- 10865961 TI - From microbial genome sequence to applications. AB - Whole genome sequences of microbial pathogens present new opportunities for clinical applications. Chief among these are development of antimicrobials, diagnostics, and vaccines. While antimicrobial development is a more difficult, long-term prospect, new diagnostics and vaccines are likely to be the first products of microbial genomics. To take advantage of whole genome sequences, methods for production of gene products in surrogate hosts (heterologous expression) are required that will work for large-scale, high-throughput gene expression. This will allow genomic information from even the most experimentally difficult pathogens to be mined for applications. In addition, screening methods to test gene products for their potential as vaccine candidates are needed for large-scale screening. These areas for technological development should be stimulated by the potential for converting genomic sequence information into applications. PMID- 10865962 TI - Regulation of granulopoiesis by transcription factors and cytokine signals. AB - The development of mature granulocytes from hematopoietic precursor cells is controlled by a myriad of transcription factors which regulate the expression of essential genes, including those encoding growth factors and their receptors, enzymes, adhesion molecules, and transcription factors themselves. In particular, C/EBPalpha, PU.1, CBF, and c-Myb have emerged as critical players during early granulopoiesis. These transcription factors interact with one another as well as other factors to regulate the expression of a variety of genes important in granulocytic lineage commitment. An important goal remains to understand in greater detail how these various factors act in concert with signals emanating from cytokine receptors to influence the various steps of maturation, from the pluripotent hematopoietic stem cell, to a committed myeloid progenitor, to myeloid precursors, and ultimately to mature granulocytes. PMID- 10865963 TI - Hepatosplenic T cell lymphoma. A review on 45 cases since the first report describing the disease as a distinct lymphoma entity in 1990. AB - Peripheral T cell lymphomas are a heterogeneous group of post-thymic, mature lymphoid malignancies, accounting for approximately 10-15% of all non-Hodgkin's lymphomas. A rare entity within this group is represented by hepatosplenic gammadelta T cell lymphoma, which is characterized by primary extranodal disease with typical sinusoidal or sinusal infiltration of the liver and the spleen, respectively, by expression of the T cell receptor gammadelta chain, and by a number of other frequent clinicopathological features including aggressive course of disease. In contrast to these common attributes some biologic characteristics such as expression of cytotoxic proteins and cytotoxic activity have been controversial. In this review, clinicopathological, immunophenotypical, molecular biological, cytogenetical and biological findings, and diagnostic and therapeutic difficulties in hepatosplenic gammadelta T cell lymphoma are discussed. PMID- 10865964 TI - Detection and quantification of residual disease in chronic myelogenous leukemia. AB - The degree of tumor load reduction after therapy is an important prognostic factor for patients with CML. Conventional metaphase analysis has been considered to be the 'gold standard' for evaluating patient response to treatment but this technique normally requires bone marrow aspiration and is therefore invasive. The frequency of cytogenetic analyses can be considerably reduced if patients are also monitored by molecular methods, which can be performed on peripheral blood specimens. Of the various techniques available, most attention has been paid to RT-PCR for BCR-ABL mRNA since this is by far the most sensitive. Simple, non quantitative RT-PCR analysis gives only limited information on patients after treatment. Quantitative RT-PCR assays have been developed to monitor the kinetics of residual BCR-ABL transcripts over time. Variables in the quantitative PCR assay may be controlled for by quantification of transcripts of a normal gene (eg ABL or glucose-6-phosphate dehydrogenase, G6PD) as an internal standard. After allogeneic stem cell transplantation, most patients become RT-PCR negative, often after a period of low level positivity that may persist for several months. Those patients destined to relapse are characterized by the reappearance and/or rising levels of BCR-ABL transcripts. In contrast, for patients treated with interferon alpha (IFN) residual disease is rarely, if ever, eliminated. The actual level of minimal residual disease in complete cytogenetic responders to IFN correlates with the probability of relapse. New quantitative real time procedures promise to simplify the protocols that are currently in use, but standardization and the introduction of rigorous, internationally accepted controls are required to enable RT-PCR to become a robust and routine basis for therapeutic decisions. PMID- 10865965 TI - Treatment of relapsing acute promyelocytic leukemia by all-trans retinoic acid therapy followed by timed sequential chemotherapy and stem cell transplantation. APL Study Group. Acute promyelocytic leukemia. AB - The purpose of this study was to assess the safety and efficacy of stem cell transplantation (SCT) mainly autologous SCT as consolidation therapy in APL patients who relapsed and achieved a second complete remission (CR2). Fifty adult patients with a first relapsed APL, of whom 39 had been previously treated with ATRA, entered a multicenter trial of oral ATRA until complete remission (CR) achievement followed by timed sequential chemotherapy (EMA combining etoposide 200 mg/m2/day for 3 days, mitoxantrone 12 mg/m2/day for 3 days, and cytarabine 500 mg/m2/day for two sequences of 3 days). EMA was started either after CR achievement, or on day 1 of ATRA because of initial white blood cell (WBC) counts >5 x 10(9)/l, or rapidly added to ATRA in order to prevent ATRA syndrome because WBC count increased under ATRA. Forty-five patients (90%, 95% CI 78%-97%) were in CR after induction therapy. Five patients died from infection during aplasia following EMA chemotherapy. Eleven patients who achieved CR had a familial HLA identical donor and were allografted. The median disease-free survival (DFS) of allografted patients was 8.2 months. The 34 other CR patients were scheduled for autologous peripheral blood (PB) SCT (intent-to-treat group). Actually, autologous transplantation was only carried out in 22 patients (65%) (17 PBSCT and five autologous bone marrow transplantation (BMT)). Reasons for not autografting were early relapse (three patients), severe toxicity of EMA chemotherapy (six patients), and refusal or failure of stem cell harvest (three patients). The 3-year DFS rate of patients actually autografted was 77%. Among the 17 autografted patients still in CR2, nine patients have already reached a longer CR2 than first CR (CR1). Results of detection of PML/RARalpha by RT-PCR after autologous transplantation show negative findings in eight of the nine patients tested. We conclude that (1) ATRA combined to EMA chemotherapy is effective in the treatment of relapsed APL; (2) allogeneic BMT may be too toxic after salvage treatment including EMA intensive chemotherapy; (3) clinical outcome of autografted patients and preliminary molecular results regarding detection of PML/RARalpha after autologous PBSCT are encouraging. PMID- 10865966 TI - Increase therapy-related leukemia secondary to breast cancer. AB - Therapy-related leukemia associated with chemotherapy, particularly alkylating agents and topoisomerase II inhibitors, are being reported with increasing frequency in the literature mainly after breast cancer. We also observed an increasing number of such leukemias in the data base of the specialized registry of hematological malignancies of the Cote d'Or department. Between 1980 and 1998, 156 AML and RAEB-t were registered in women in Cote d'Or. Among them, 12 occurred in women with breast cancer history (7.7%). Analysis by period of time shows a significant increase in the proportion of therapy-related leukemia secondary to breast cancer (P < 0.02). Chemotherapy including topoisomerase II inhibitors was used in 10 cases in which mitoxantrone was used in eight cases. In these eight cases, leukemia had clinical and biological characteristics usually described with topoisomerase II inhibitors but 44% were promyelocytic sub-type with the t(15;17) specific karyotypic abnormality. These data on a well-defined population demonstrate the increased proportion of therapy-related leukemia secondary to breast cancer, probably due to the use of mitoxantrone. PMID- 10865967 TI - P-glycoprotein in primary acute myeloid leukemia and treatment outcome of idarubicin/cytosine arabinoside-based induction therapy. AB - The expression of the drug transport protein, P-glycoprotein (Pgp/MDR1) has been found to be of prognostic significance for the achievement of complete remission (CR) or the duration of survival after daunorubicin (DNR)-containing induction therapy in acute myeloid leukemia (AML). This would suggest that the expression of Pgp in AML is high enough to have significant impact on intracellular DNR concentrations and on clinical therapy failure in AML. Recently, DNR has been replaced in many centers by idarubicin (IDA) as the first choice anthracycline in AML treatment. We have, therefore, performed a study in a group of 98 primary AML patients, who all received IDA, but not DNR during induction therapy in order to determine if the response to IDA-containing induction therapy might be related to the biologic characteristic of Pgp expression in AML. The AML samples were studied for Pgp expression by MRK16 antibody staining and for Pgp activity measured as the modulation of rhodamine 123 uptake by 2 microM PSC 833. No correlation of Pgp with complete response rate, event-free survival or overall survival was found. In addition to Pgp, the expression of another protein that has been implicated by some studies in response failure to DNR-containing therapy, the major vault protein (Mvp/LRP), was studied. This marker did not correlate with CR or survival after IDA-containing therapy. The results of this patient study are consistent with model studies showing that the steady-state cellular accumulation of lipophilic anthracyclines such as IDA are little affected by Pgp. Therefore, putative beneficial effects of the inclusion of PSC 833 in IDA-containing therapy might rather be related to alternative mechanisms than to inhibition of Pgp-mediated IDA efflux. PMID- 10865968 TI - Characterization of acute myeloid leukemia with MLL rearrangements--no increase in the incidence of coexpression of lymphoid-associated antigens on leukemic blasts. AB - MLL gene rearrangements are associated with coexpression of myeloid- and lymphoid associated antigens on leukemic blasts and a dismal outcome in acute lymphoblastic leukemia (ALL). Whether the same conditions can apply to acute myeloid leukemia (AML) is not quite clear. Rearrangements of the MLL gene were analyzed on 113 patients with newly diagnosed de novo AML in a single institution. Sixteen (14%) of them showed rearranged bands by Southern blot analysis, including three (50%) of six infants, three (14%) of 21 children between 1 and 15 years and 10 (12%) of 86 adults. MLL rearrangements were not only detected in M5 (four of 12 patients, 33%) and M4 (six of 31, 19%) subtypes but also in other non-M4-M5 AML (six of 70, 9%), including M1, M2 and M7, but not M3 subtype. Seven patients had chromosomal abnormalities involving 11q23, but nine did not. The latter comprised three (6%) of 48 patients with normal karyotype, one with t(8;21), none with t(15;17), inv(16) or t(9;22), and four (15%) of 27 with cytogenetic aberrations other than those specific structural abnormalities. In contrast to ALL, AML patients with MLL rearrangements did not tend to coexpress lymphoid- and myeloid-associated antigens simultaneously on leukemic blasts and have similar outcome as those without the gene rearrangements. PMID- 10865969 TI - Comparison of spectral karyotyping and conventional cytogenetics in 39 patients with acute myeloid leukemia and myelodysplastic syndrome. AB - Spectral karyotyping (SKY) was performed in patients with acute myeloid leukemia (AML; n = 25), secondary AML (s-AML; n = 7), myelodysplastic syndrome (MDS; n = 6) and s-MDS (n = 1) to complement conventional cytogenetic investigations. According to the results of conventional cytogenetics the patients were subdivided into three groups: group 1, normal karyotype, n = 19 cases, median age = 64 years; group 2, patients displaying either one or two single aberrations, n = 10 cases, median age = 54 years; group 3, patients with > or =3 independent aberrations, n = 10 cases, median age = 61.5 years. SKY identified no abnormal metaphases in group 1. In one patient of group 2 a hidden translocation t(7;14)(q3?1;q2?2) could be revealed with SKY. Conventional cytogenetics had only shown trisomy 8. A similar t(7;14) was also detected in one patient of group 3. SKY was helpful for the delineation of marker chromosomes and additional material. Furthermore, SKY could distinguish between partial and total monosomies or real existing and apparent deletions. The combination of G-banding, FISH and SKY was found very useful for the precise delineation of the karyotype. As a result of our study we recommend SKY investigation as an important additional tool for accurate chromosome analysis. The detected t(7;14) might represent a novel recurrent translocation in acute myeloid leukemias. PMID- 10865970 TI - Survival time in a population-based consecutive series of adult acute myeloid leukemia--the prognostic impact of karyotype during the time period 1976-1993. AB - A consecutive population-based series of 372 adult acute myeloid leukemias, successfully cytogenetically investigated at a single center between 1976 and 1993, is reported. All medical records were reviewed in order to ascertain the prognostic impact of karyotype, divided into three groups; favorable (t(8;21), t(15;17), and inv(16) irrespective of karyotypic complexity; n = 40), poor (der(1;7), inv(3), -5, del(5q), -7, t(9;22), and complex karyotypes including whole or partial losses of chromosomes 5 and/or 7; n = 56), and intermediate (other abnormalities or normal karyotype; n = 276). The possible modification by age, gender, time period, morphologic subtype, and bone marrow transplantation (BMT) on this prognostic impact was also determined. The chemotherapy regimens used were heterogeneous over time but principally the same at any given point in time. The majority of the patients were treated with combinations including an anthracycline and cytarabine with curative intent. Gender, morphology, and BMT did not significantly modify the effect of cytogenetic patterns on survival time, whereas age and time period did. The hazard ratios for the subgroups favorable, intermediate, and poor were 1.0, 1.2 and 1.9 at age 20-49; 1.0, 2.5 and 4.5 at age 50-64; 1.0, 4.1 and 11.4 at age 65-74; 1.0, 1.4 and 2.2 for the time period 1976-1987 and 1.0, 2.0 and 6.7 for 1988-1993. The salient feature of the Kaplan Meier curves was the improved survival during the later time period for patients with favorable and intermediate cytogenetic abnormalities. The present findings thus suggest that it is mainly these patient groups that have benefited from advances in therapy, including supportive care. PMID- 10865971 TI - Microsatellite instability is not a defining genetic feature of acute myeloid leukemogenesis in adults: results of a retrospective study of 132 patients and review of the literature. AB - The frequency of acute leukemia in children with constitutional DNA repair defects implicates defective DNA repair in leukemogenesis. Whether sporadic cases of AML also arise from an inherited genetic predisposition remains to be determined. Prior studies have reported microsatellite instability (MSI) in AML, particularly secondary and relapsed AML. These studies included small numbers of cases in which key features such as cytogenetic abnormalities were not reported. To determine whether defective DNA mismatch repair, reflected by MSI, is a defining feature of adult myeloid leukemogenesis, we retrospectively studied 132 AML cases including 28 de novo, 62 secondary, 22 relapsed/refractory, 15 cases of paired diagnosis/relapse. 110 patients were elderly (55+ years). The cases included a range of cytogenetic abnormalities. MSI was assessed at three loci (BAT 25, BAT 26, BAT 40) in DNA isolated from sorted leukemic blasts and paired T cell controls. Fluoresceinated PCR products were analyzed using an automated capillary electrophoresis system. Of the 132 AML cases, no single case demonstrated MSI. Our studies indicate that MSI, and defective DNA mismatch repair, is not a defining feature of the majority of adult patients with AML. Furthermore, our data does not support the hypothesis that MSI could be acquired during the progression of AML from diagnosis to relapse, as a consequence of therapeutic exposure. PMID- 10865972 TI - Expression and responsiveness of human interleukin-18 receptor (IL-18R) on hematopoietic cell lines. AB - Interleukin-18 (IL-18) is a new inflammatory cytokine sharing biological functions with IL-12. The human IL-18 receptor (IL-18R) was recently identified and was found to be expressed on normal peripheral blood lymphocytes. To further characterize IL-18R, we analyzed IL-18R expression using a series of human hematopoietic cell lines selected from various cell lineages. We found the IL-18R expression on cells of T and B lineages as expected from analysis on normal cells. The IL-18R expression, however, was found not to be restricted to any specific maturation stages of T and B cells. In addition, we detected IL-18R expression in myeloid, monocytoid, erythroid and megakaryocytic cell lines, indicating that normal counterparts of these cell lineages could express IL-18R and participate in in vivo reactions caused by IL-18. Biochemical studies showed that IL-18R proteins exist as heterogeneous molecules ranging from 60 to 110 kDa. Deglycosylation experiments indicated that the heterogeneity could not be explained only by a difference in glycosylation. We also found that tumor necrosis factor-alpha (TNF-alpha) modulated the IL-18R expression, which implies an important in vivo effect of TNF-alpha on IL-18-induced reaction. Analyzing the responsiveness of IL-18R, we found that only KG-1 responded to IL-18 stimulation. This suggests that certain inhibitory mechanisms of IL-18 responsive genes are involved in the all IL-18R-positive cell lines except KG-1. PMID- 10865973 TI - Synergy between Raf and BCL2 in abrogating the cytokine dependency of hematopoietic cells. AB - The Raf oncoprotein plays critical roles in the transmission of mitogenic signals from cytokine receptors to the nucleus. There are three Raf family members: A Raf, B-Raf and Raf-1. Conditionally active forms of the Raf proteins were created by ligating N-terminal truncated activated forms to the estrogen-receptor (ER) hormone-binding domain resulting in beta-estradiol-inducible constructs. We introduced these chimeric deltaRaf:ER oncoproteins into the murine FDC-P1 hematopoietic cell line. Two different types of cells were recovered after drug selection in medium containing either cytokine or beta-estradiol: (1) cytokine dependent cells that expressed the deltaRaf:ER oncoproteins; and (2) Raf responsive cells that grew in response to the deltaRaf:ER oncoprotein. Depending upon the particular deltaRaf:ER oncoprotein, cytokine-dependent cells were recovered 10(3) to 10(5) times more frequently than Raf-responsive cells. To determine whether BCL2 could synergize with the deltaRaf:ER oncoproteins and increase the frequency of cytokine-independent cells, cytokine-dependent deltaRaf:ER-expressing cells were infected with either a BCL2 containing retrovirus or an empty retroviral vector. BCL2 overexpression, by itself, did not relieve cytokine dependency of the parental cell line. However, BCL2 overexpression increased the frequency of Raf-responsive cells approximately five to 100-fold. Cytokine-dependent deltaRaf:ER-infected cells entered the G1 phase of the cell cycle after cytokine withdrawal and entered S phase only after cytokine addition. Raf-responsive deltaRaf:ER cells entered the G1 phase of the cell cycle after estrogen deprivation and re-entered the cell cycle after addition of either IL-3 or the estrogen receptor antagonist tamoxifen which activates the deltaRaf:ER constructs. Expression of the BCL2 oncoprotein often delayed the exit from the S and G2/M phases demonstrating the protective effects BCL2 provided to these Raf and BCL2 infected cells. The deltaRaf:ER cells expressed the deltaRaf:ER proteins and downstream MEK and ERK activities after beta-estradiol treatment. Raf-responsive cells that were also infected with BCL2 expressed higher levels of BCL2 than the cells that were not infected with BCL2. Thus BCL2 can synergize with the activated Raf in the abrogation of cytokine dependency of certain hematopoietic cells. These cells will be useful in furthering our understanding of the roles of the Raf and BCL2 oncoproteins in hematopoietic cell growth, cell cycle progression and prevention of apoptosis. PMID- 10865975 TI - Demonstration of the mineralocorticoid hormone receptor and action in human leukemic cell lines. AB - We studied the expression of the mineralocorticoid receptor (MCR), and of the amiloride-sensitive sodium channel (ASSC) regulated by the MCR, in human leukemic cell lines. Cell extracts from TF1 (proerythroblastic), HEL (human erythroblastic leukemia) and U937 (myeloblastic) cell line were positive for the ASSC, as a 82 kDa band in Western blots developed with the aid of a polyclonal antibody raised against the peptide QGLGKGDKREEQGL, corresponding to the region 44-58 of the alpha subunit of the epithelial sodium channel (ENaC) cloned from rat colon, linked to KLH. The polyclonal antibody against the MCR revealed a single band of about 102 kDa in extracts from HEL and TF1 cells. The immunofluorescent labelling of the MCR in all cell lines showed a nucleocytoplasmic localization of the receptor but the ASSC was exclusively membrane-bound and these results were confirmed by confocal microscopy. The expression of the MCR in the HEL cells was evident as a predicted band of 843 bp (234 amino acids) in electrophoresis of the PCR product obtained after total RNA had been reverse transcribed and then amplified using the primers 5'-AGGCTACCACAGTCTCCCTG-3' and 5' GCAGTGTAAAATCTCCAGTC-3' (sense and antisense, respectively). The ENaC was similarly evident with the aid of the primers 5'-CTGCCmATG GATGATGGT-3' (sense) and 5'-GTTCAGCTCGAAGAAGA-3' (antisense) as a predicted band of 520 bp. In both cases, 100% identity was observed between the sequences of the PCR products compared to those from known human sources. The multiplication of the HEL cells was influenced by antagonists (RU 26752, ZK 91587) targeted for specificity to the MCR and this was selectively reversed by the natural hormone aldosterone. These steroids also provoked chromatin condensation in the HEL population. These permit new and novel possibilities to understand the pathobiology of human leukemia and to delineate sodium-water homeostasis in nonepithelial cells. PMID- 10865974 TI - Combined effects of aberrant MEK1 activity and BCL2 overexpression on relieving the cytokine dependency of human and murine hematopoietic cells. AB - The MEK1 oncoprotein plays a critical role in Ras/Raf/MEK/MAPK-mediated transmission of mitogenic signals from cell surface receptors to the nucleus. In order to examine this pathway's role in leukemic transformation, a conditionally active (beta-estradiol-inducible) form of the MEK1 protein was created by ligating a cDNA encoding an N-terminal truncated form of MEK1 to the hormone binding domain of the estrogen receptor (ER). We introduced this chimeric deltaMEK1:ER oncoprotein into cytokine-dependent human TF-1 and murine FDC-P1 hematopoietic cell lines. Two different types of cells were recovered after drug selection in medium containing either cytokine or beta-estradiol: (1) cells that expressed the deltaMEK1:ER oncoprotein but remained cytokine-dependent and (2) MEK1-responsive cells that grew in response to deltaMEK1:ER activation. Cytokine dependent cells were recovered 10(2) to 10(4) times more frequently than MEK1 responsive cells depending upon the particular cell line. To determine whether BCL2 overexpression could synergize with the deltaMEK1:ER oncoprotein in relieving cytokine dependence, the cytokine-dependent deltaMEK1:ER-expressing cells were infected with a BCL2-containing retrovirus, and the frequency of MEK1 responsive cells determined. BCL2 overexpression, by itself, did not relieve cytokine dependency of the parental cells, however, it did increase the frequency at which MEK1-responsive cells were recovered approximately 10-fold. DeltaMEK1:ER+BCL2 cells remained viable for at least 3 days after estradiol deprivation, whereas viability was readily lost upon withdrawal of beta-estradiol in the MEK1-responsive cells which lacked BCL2 overexpression. The MAP kinases, ERK1 and ERK2 were activated in response to deltaMEK1:ER stimulation in both deltaMEK1:ER and deltaMEK1:ER+BCL2 cells. As compared to the cytokine-dependent deltaMEK1:ER and BCL2 infected cells, MEK1-responsive BCL2 infected cells expressed higher levels of BCL2. While both MEK1-responsive deltaMEK1:ER and deltaMEK1:ER+BCL2 infected cells expressed cDNAs encoding the autocrine cytokine GM-CSF, more GM-CSF cDNAs and bioactivity were detected in the MEK1-responsive deltaMEK1:ER+BCL2 cells than in the MEK1-responsive cells lacking BCL2 or cytokine-dependent cells. These conditionally transformed cells will be useful in furthering our understanding of the roles MEK1 and BCL2 play in the prevention of apoptosis in hematopoietic cells. PMID- 10865976 TI - The role of RAR and RXR activation in retinoid-induced tissue factor suppression. AB - Excessive expression of tissue factor (TF) is a common finding in leukaemic cells and may contribute to thrombotic complications in patients. Retinoic acid has been shown to induce differentiation and reduce TF expression in acute promyelocytic leukaemia (APL) cells in vitro, and to induce remission in APL patients. Treatment of the APL cell line NB4 with the specific retinoic acid receptor-alpha (RARalpha) agonists Ro4-6055 or TTNPB resulted in down-regulation of TF expression and in induction of differentiation. The activation of RARbeta, RARgamma or retinoid X receptor (RXR) did not suppress the constitutive TF expression in NB4 cells. Moreover, the RARalpha antagonist Ro41-5253 blocked the retinoid-induced down-regulation of TF. In contrast, in the monoblastic U-937 cell line only a partial suppression of TF antigen expression and activity was observed by treatment with the RAR agonist TTNPB or the RXR agonist SR11237 alone. However, the combination of TTNPB and SR11237 resulted in a pronounced down-regulation of TF expression and induction of differentiation in U-937 cells. We show for the first time that the activation of both subunits of the RARalpha RXR transcriptional complex is needed for TF suppression in U-937 cells, whereas in NB4 cells RARalpha activation alone is sufficient. Thus, distinct molecular mechanisms for TF suppression seem to be operating in leukaemic cell lines of different origin. PMID- 10865977 TI - Autonomous multi-lineage differentiation in vitro of primitive CD34+ cells from patients with chronic myeloid leukemia. AB - Neoplastic CD34+ cells from chronic myeloid leukemia (CML) patients proliferate in vitro in the absence of serum or defined growth factors due to an autocrine mechanism involving IL-3 and G-CSF (Jiang et al. Proc Natl Acad Sci USA 1999; 96: 12804). Detailed examination of the various cell types produced in such cultures has now demonstrated the rapid, factor-independent, generation of clonogenic progenitors for all lineages (granulocyte-macrophage, megakaryocyte and erythroid) with the additional appearance within 10 days of large numbers of mature granulocytes, macrophages, and megakaryocytes, as well as occasional erythroid cells. Inclusion of flt3-ligand, Steel factor, IL-3, IL-6, and G-CSF +/ erythropoietin (EPO) in the cultures enhanced only slightly the output of mature cells (except for the erythroid population which was much larger when EPO was added). Analogous subpopulations of normal CD34+ cells produced similar numbers and types of cells but, as expected, only when growth factors were added. Thus primitive CD34+ CML cells proliferating autonomously in vitro recapitulate the full spectrum of differentiation responses of normal CD34+ cells stimulated by IL 3 and G-CSF. These findings point to a role of autocrine IL-3 and G-CSF in the similar multi-lineage expansion of differentiating CD34+ CML cells that occurs in vivo. PMID- 10865979 TI - Lymph nodes and Peyer's patches of IL-6 transgenic BALB/c mice harbor T(12;15) translocated plasma cells that contain illegitimate exchanges between the immunoglobulin heavy-chain mu locus and c-myc. AB - Hyperplastic plasmacytotic lymph nodes and Peyer's patches of 12 of 25 (48%) BALB/c mice that carried a human IL-6 transgene under the transcriptional control of the histocompatibility H-2L(D) promoter (BALB/c.IL-6 mice) were found to harbor 15 cell clones that contained in their T(12;15) translocation breakpoint regions illegitimate genetic recombinations between the upstream flank of the immunoglobulin heavy-chain C mu locus (5'-C mu) and c-myc (5'-C mu/c-myc+ clones). Similar 5'-C mu/c-myc+ clones were also detected in pristane-induced peritoneal granulomata (a significant source of IL-6 in situ) of three of 13 (13%) conventional BALB/c mice, but not in lymphoid tissues of pristane-treated BALB/c mice, nor in any tissue of untreated BALB/c mice. These findings provided strong evidence that IL-6 may be able to promote the growth and/or survival of clones that contained rearrangements between 5'-C mu and c-myc. Taken in conjunction with our previous observation that 5'-C mu/c-myc+ clones are the precursors for pristane-induced BALB/c plasmacytomas, the findings further suggested that IL-6 may play a pivotal role in the early stage of plasmacytoma development, by promoting tumor precursor cells. The BALB/c.IL-6 model of plasmacytomagenesis may be superior to the conventional BALA/c model because the putative plasmacytoma precursors appear to be more prevalent and in their development independent of treating the mice with inflammation-inducing plasmacytomagenic agents, such as pristane or silicone polymers. PMID- 10865978 TI - The existence of lymphoid lineage restricted Philadelphia chromosome-positive acute lymphoblastic leukemia with heterogeneous bcr-abl rearrangement. AB - Analysis of lineage involvement was performed in 17 Philadelphia chromosome positive acute lymphoblastic leukemia patients with no history of chronic myeloproliferative disorder. The percentage of blastic cells as defined by flow cytometry matched that of the Ph-positive cells in 14 out of 17 patients. The bcr abl rearrangement was investigated by fluorescent in situ hybridization in morphologically identified blastic cells, myeloid elements, lymphocytes and erythroblasts using a combined light and fluorescent microscopical imaging. Lymphoid lineage restriction could be determined in all but three of the patients. These 14 patients exhibited heterogeneity in terms of m-bcr and M-bcr types of translocation as revealed by reverse transcription polymerase chain reaction. The three patients with multilineage involvement and M-bcr type of translocation reverted to chronic phase and the percentage of Ph-positive cells remained high. Thus, we could identify an uncommitted stem cell origin among Ph positive ALLs only in those patients whose disease subsequently proved to be a lymphoid blastic crisis with clinically silent chronic phase. PMID- 10865980 TI - Phase I/II study of 2-chloro-2'-deoxyadenosine with cyclophosphamide in patients with pretreated B cell chronic lymphocytic leukemia and indolent non-Hodgkin's lymphoma. AB - Because of their substantial in vitro synergy, we conducted a dose-escalation study of cyclophosphamide (CP) added to 2-chloro-2'-deoxyadenosine (CdA) in patients with previously treated chronic lymphocytic leukemia and non-Hodgkin's lymphoma. CdA was given at a fixed dose (5.6 mg/m2/day) as a 2-h intravenous (i.v.) infusion, immediately followed by a 1-h i.v. infusion of CP, for 3 days. The initial daily CP dose was 200 mg/m2, and was escalated by 100 mg/m2 increments in successive cohorts of three to six patients to determine the maximum-tolerated dose (MTD). Additional patients were included at the MTD to extend toxicity and response analysis. Twenty-six patients received 68 cycles of chemotherapy. The MTD of CP after CdA 5.6 mg/m2, was 300 mg/m2. Acute neutropenia was the dose-limiting toxicity of this regimen, which was otherwise well tolerated. Delivery of repeated cycles was not feasible in eight patients (31%) because of prolonged thrombocytopenia. Severe infections were seen in three of 68 cycles (4%). The overall response rate was 58% (15 of 26; 95% CI, 36-76%), with 15% complete responses and 42% partial responses. These data show the feasibility of the association of CdA with CP. Given the response rate observed, further studies of this regimen are warranted in untreated patients, in particular with chronic lymphocytic leukemia and with Waldenstrom macroglobulinemia. PMID- 10865982 TI - Interferon may reduce minimal residual disease of acute promyelocytic leukemia. PMID- 10865981 TI - Rapid, multifluorescent TCRG Vgamma and Jgamma typing: application to T cell acute lymphoblastic leukemia and to the detection of minor clonal populations. AB - Detection of clonal T cell receptor gamma (TCRG) gene rearrangements by PCR is widely used in both the diagnostic assessment of lymphoproliferative disorders and the follow-up of acute lymphoblastic leukaemia (ALL), when residual positivity in excess of 10(-3) at morphological complete remission is increasingly recognised to be an independent marker of poor prognosis. This is largely based on specific detection of V-J rearrangements from childhood cases. We describe rapid, multifluorescent Vgamma and Jgamma PCR typing of multiplex amplified diagnostic samples, as applied to 46 T-ALL. These strategies allow selected analysis of appropriate cases, immediate identification of Vgamma and Jgamma segments in over 95% of alleles, improved resolution and precision sizing and a sensitivity of detection at the 10(-2)-10(-3) level. We demonstrate preferential V-J combinations but no difference in V-J usage between children and adults, nor between SIL-TAL1-negative and -positive cases. A combination of fluorescent multiplex and Vgamma-Jgamma-specific monoplex follow-up, as described here, will allow detection of both significant clonal evolution and of the diagnostic clone at a level of prognostic significance, by techniques which can readily be applied to large-scale prospective studies for which real-time analysis is required. PMID- 10865983 TI - Deletion of the multidrug resistance-associated protein (MRP1) gene in acute myeloid leukemia with inversion of chromosome 16 has no prognostic impact. PMID- 10865984 TI - Chronic lymphocytic leukemia in pregnancy. PMID- 10865985 TI - Vitamin K2 therapy for myelodysplastic syndromes (MDS) and post-MDS acute myeloid leukemia: information through a questionnaire survey of multi-center pilot studies in Japan. PMID- 10865986 TI - Lack of BCR/ABL reciprocal fusion in variant Philadelphia chromosome translocations: a use of double fusion signal FISH and spectral karyotyping. PMID- 10865987 TI - The effect of integrin antibodies on the attachment and proliferation of human Tenon's capsule fibroblasts. AB - The integrins are protein heterodimers consisting of noncovalently associated alpha and beta subunits. The adhesive interactions mediated by integrins are necessary for cellular survival and proliferation. In this study we investigated the effects of three different integrin antibodies on the proliferation of human Tenon's capsule fibroblasts in tissue culture. Human Tenon's capsule fibroblasts were cultured into 96 well plates and treated with different concentrations (ranging from 10(-6) to 1 microg ml(-1)) of three different integrin antibodies: human integrin alpha-2 antibody, human integrin alpha-3 antibody and human integrin alpha-5/FnR (fibronectin receptor) antibody. Coulter counter, hexosaminidase, and 3H-thymidine assays were used to determine the inhibitory effects of these integrin antibodies on ocular fibroblasts on days 0 (attachment), 1,3 and 7 following antibody treatment. The concentration of each antibody required to produce a proliferation 50% less than the control (ID50) was calculated for each assay. With respect to attachment, all three antibodies studied displayed some inhibitory activity. All three antibodies also displayed dose-dependent antiproliferative properties, especially at the highest concentration tested after 7 days of exposure. The integrin alpha-2 antibody was the most potent of the inhibitors, followed by the integrin alpha-3 antibody, with the integrin alpha-5 antibody being the least potent antibody tested. In addition, the anti-proliferative activities of the integrin alpha-2 and integrin alpha-3 antibodies increased with increasing incubation time. In conclusion, these integrin antibodies demonstrated some inhibitory effects on the attachment and proliferation of human Tenon's capsule fibroblasts in culture. Further investigation will be required to determine whether integrin antibodies can significantly limit scar formation in vivo without significant toxicity. PMID- 10865988 TI - Type XIII collagen is widely expressed in the adult and developing human eye and accentuated in the ciliary muscle, the optic nerve and the neural retina. AB - The distribution of mRNAs coding for type XIII collagen, a novel nonfibril forming collagen, was studied by Northern and in situ hybridizations of adult and fetal human eyes and the corresponding protein was localized by indirect immunofluorescence in frozen sections of 12 and 17 week human fetal eyes using a polyclonal antipeptide antibody to type XIII collagen. Type XIII collagen was found to be widely expressed in ocular tissues when studied at both the mRNA and protein levels in fetal and adult human tissues. No major differences were observed in the expression patterns between fetal and adult tissues. Surprisingly, the strongest signals seen in in situ hybridizations and immunofluorescence stainings occurred in the optic nerve bundles and in the ganglion cell layer of the retina. Other notable locations containing type XIII collagen included the developing ciliary smooth muscle, the posterior two-thirds of the corneal stroma and the striated extraocular muscles. Low level signals were also detected in the blood vessel walls and mesenchymal cells of the other ocular tissues. All immunosignals detected were adherent to cells, and the extracellular matrices appeared to be devoid of type XIII collagen. Our results are in concert with the presumed plasma membrane location of type XIII collagen, and it is hypothesized that this molecule could be involved in cell-matrix and perhaps cell-cell interactions. The wide expression of type XIII collagen in the eye, and especially in the neural structures, warrants future studies on type XIII collagen in other nerve structures and in pathological conditions affecting the eye. Due to its wide expression, type XIII collagen is likely to be an important factor for the normal development and functioning of the eye. PMID- 10865989 TI - Somatotopic organization of primary afferent perikarya of the guinea-pig extraocular muscles in the trigeminal ganglion: a post-mortem DiI-tracing study. AB - Apart from the somatotopic organization of the trigeminal ganglion (TG) into the ophthalmic, maxillary and mandibular divisions along the mediolateral axis, there exist further somatotopic organizations within these three divisions. According to literature, the cell organization in the TG and the somatotopy in the brainstem develop together, formed by naturally occurring cell death in the TG. Thus, the somatotopy of the primary afferent trigeminal perikarya is of special interest. The aim of this study was to investigate the location of the primary afferent perikarya of the extraocular muscles (EOMs) in the TG of guinea-pig. The primary afferent perikarya were labeled by post-mortem application of the carbocyanine DiI on the oculomotor nerve branches near their entrance into the single EOMs. The DiI-positive perikarya were found musculo-somatically organized in the ipsilateral ophthalmic part of the TG at a wide range along the dorsoventral axis, expressing an overlap of the representation areas. The primary afferent perikarya of the superior rectus and the superior oblique muscles were mainly localized in the dorsal part of the ganglion while those of the inferior rectus and the inferior oblique muscle mainly in ventral part. The lateral and the medial rectus were predominantly represented in between. An organization along the mediolateral axis of the TG was not observed. Although guinea-pigs lack classical EOM proprioceptors, the somatotopic representation of the extraocular muscle primary afferent perikarya in the TG found in this study is in line with findings in species with well known encapsulated proprioceptors within the EOMs. PMID- 10865990 TI - Expression of matrix metalloproteinase-2 (MMP-2) and vascular endothelial growth factor (VEGF) in inflammation-associated corneal neovascularization. AB - Matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and vascular endothelial growth factor (VEGF) are all implicated in the development of neovascularization. To investigate the possible role of these factors in corneal neovascularization we have analysed the expression of MMP-2, MMP-9 and VEGF in a rat model of inflammation-associated corneal neovascularization. In this model, corneal neovascularization was induced in Long-Evans rats by krypton laser photocoagulation whereafter eyes were enucleated at 1, 4, 7, 10 and 20 days. Slit-lamp biomicroscopy and histologic analysis revealed a gradual development of corneal neovascularization that peaked 7-10 days after treatment when newly formed vessels could be seen throughout the corneal surface reaching deep into the stroma. Antisense and sense riboprobes were generated using DNA complementary to MMP-2, MMP-9 and VEGF, and mRNA expression was analysed using in situ hybridization. The expression of MMP-2 and MMP-9 in untreated corneas was low or absent whereas VEGF was weakly expressed in the corneal epithelium. MMP-2 expression was increased during corneal neovascularization and was mainly localized to the cells infiltrating areas of new vessel formation. Many of these cells appeared to be inflammatory cells. VEGF expression had a similar overall distribution to MMP-2 during neovascularization with the exception that its expression in the corneal epithelium remained and even increased slightly. MMP-9 was prominently expressed at the border of regenerating corneal epithelium in areas with epithelial wounding but was not detected in the vascularized stroma. Together, the results of the present study support a role for MMP-2 and VEGF in inflammation-associated corneal neovascularization whereas MMP-9 instead appears to be involved in corneal epithelial wound-healing. PMID- 10865991 TI - Pharmacodynamics of beta-blocker modulation of aqueous humor production. AB - A conscious rabbit model with microdialysis sampling of endogenous aqueous humor ascorbate was developed in order to assess the pharmacodynamics of beta-blocker modulation of aqueous humor production. CMA/20 microdialysis probes were implanted in the anterior chamber of each eye of rabbits (n = 6). After a 2 week recovery period, an i.v. bolus of 14C-ascorbate (20 microCi) was administered. Blood samples and aqueous humor microdialysis probe effluent were collected and analysed for endogenous and 14C-ascorbate to estimate the basal rate of ascorbate blood to aqueous humor secretion (Ro). After a 1 hr washout, each rabbit received a series of three doses of 3H-propranolol (750-3000 microg, 16.5 microCi mg(-1)) every 60 min into the lower cul-de-sac of each eye. Probe effluent was analysed for endogenous ascorbate and 3H-propranolol; ascorbate and propranolol in the iris/ciliary body, vitreous and aqueous was determined at the end of the experiment. Nonlinear least-squares regression analysis of the concentration-time profiles for aqueous humor ascorbate was performed to estimate the change in aqueous humor flow. The average basal aqueous humor ascorbate secretion rate was approximately 48/microg hr(-1). Propranolol (1500 microg) produced significant increases in aqueous humor ascorbate, this observation is consistent with a reduction in aqueous humor production (approximately 47%). Analysis of intraocular tissue ascorbate indicated that propranolol inhibited ascorbate secretion at the 3000 microg dose, the highest dose examined in this study; this inhibition was not observed at the 750 microg or 1500 microg doses. Changes in aqueous humor production precipitated by the administration of beta-adrenergic antagonists can be estimated by measuring changes in aqueous humor ascorbate concentrations in the conscious rabbit. Microdialysis sampling of aqueous humor for endogenous ascorbate provides a relevant analytic tool to estimate modulatory effects of anti-glaucoma drugs on aqueous humor production. PMID- 10865992 TI - A potential role for immune complex pathogenesis in drusen formation. AB - Drusen are abnormal extracellular deposits that accumulate between the retinal pigmented epithelium and Bruch's membrane and are commonly associated with age related macular degeneration. Our recent work has identified a number of plasma proteins as molecular components of drusen. Of interest is the fact that many of these drusen-associated molecules are acute phase reactant proteins and some have established roles in mediating immune responsiveness. As immune and inflammatory responses appear to play a role in the formation of other pathologic age-related deposits, we examined the distribution of immunoglobulin molecules and terminal complement complexes at sites of drusen deposition. Here, we report that concentrations of immunoglobulin G and terminal C5b-9 complement complexes are present in drusen. In addition, we observe that retinal pigmented epithelial cells overlying or directly adjacent to drusen, as well as some within apparently normal epithelia, exhibit cytoplasmic immunoreactivity for immunoglobulin and the C5 component of complement. Taken together, these results suggest that drusen biogenesis may be a byproduct of immune responsiveness, and they implicate immune complex-mediated pathogenesis involving retinal pigmented epithelial cells as an initiating event in drusen formation. PMID- 10865993 TI - In vivo cataract after repeated exposure to ultraviolet radiation. AB - The purpose of this study was to investigate the effects of repeated close-to threshold ultraviolet radiation (UVR) doses in the rat lens. Sprague Dawley rats received two UVR exposures (lambda(max) = 300 nm, lambda0.5 = 10 nm) separated by different time intervals. The animals were unilaterally irradiated with 4 kJ m(2 1) UVR in each exposure. The intervals between both exposures were: 6 hr, 1 day, 3 days, 9 days and 30 days. At 1 week after the last exposure both lenses were removed, microphotographs were taken and intensity of forward light scattering was measured. Evaluating the difference between exposed and non-exposed eyes, the forward light scattering in the 6 hr and 1 day interval group was not significantly different. The most intense forward light scattering was found in the group that was allowed 3 days interval between exposures. Thereafter, the intensity of scattering decreased as the time interval between exposures increased. The lowest intensity of forward light scattering was detected in the 30 days interval group. Three days after a UVR exposure, the lens showed the highest sensitivity for a second UVR exposure. One month after the first exposure lenses undergo physiological repair and interactions between exposures seem to decrease. PMID- 10865994 TI - Signal transduction mediated by adhesion of human trabecular meshwork cells to extracellular matrix. AB - In this study we investigated the signaling event induced by adhesion of human trabecular meshwork (TM) cells to extracellular matrix (ECM) elements such as fibronectin. The role of tyrosine phosphorylation in adhesion was evaluated. A number of intracellular entities involved in the adhesion-mediated pathways were identified. For the experiments, human TM cells were seeded onto fibronectin- or polylysine (negative control)-coated plates. Fifteen, 30, 90 and 240 min after the seeding, cell lysates were collected. Immunoblotting analysis revealed that tyrosine phosphorylation occurred within 15 min of adhesion of TM cells to fibronectin and the level increased with time. The phosphotyrosyl proteins had molecular masses 25-220 kDa. A much lower level of tyrosine phosphorylation was observed when cells were plated on polylysine. Immunoprecipitation experiments indicated that the phosphotyrosine-containing proteins included focal adhesion kinase, paxillin, phosphatidylinositol 3-kinase and mitogen activated protein kinase. Within 30 min of adherence to fibronectin, human TM cells immunostained for paxillin and phosphotyrosine and exhibited prominent focal contacts. When treated with tyrosine kinase inhibitors genistein and herbimycin A and a protein kinase C (PKC) pseudosubstrate peptide inhibitor, cell adhesion to fibronectin was compromised and focal contact formation was limited. These results demonstrated that in human TM cells, tyrosine kinase was activated upon their adherence to fibronectin. PKC also appeared to play a role in modulation of the cell-matrix adhesion process. The current study provides insight into the signaling pathways that are linked to the ECM-induced events in TM cells. Elucidation of the hierarchy of signal responses may help develop strategies manipulating the cell-matrix interactions in the TM system. PMID- 10865995 TI - Retinal degeneration in the nervous mutant mouse. III. Electrophysiological studies of the visual pathway. AB - The nervous (nr) mutation induces a progressive and severe degeneration of cerebellar Purkinje cells and retinal photoreceptors that is virtually complete within the first few months of life. Previous studies of the retina in nervous (nr/nr) mice have focused primarily on the structural abnormalities seen at the level of the photoreceptor cell bodies and outer segments. Here, we have carried out a series of functional studies of the visual pathway in nervous mice and have quantified the status of the inner retinal cell and plexiform layers. Affected animals were obtained by mating nr/+ heterozygotes and screening the offspring for the ataxia characteristic of nervous animals; phenotypically normal littermates (i.e. nr/+ or +/+) were used as controls. As described previously, there is a substantial loss of photoreceptors cells in the nervous retina and a marked shortening of the inner and outer segments. These changes are accompanied by a more modest decline in the thickness of the inner plexiform and inner nuclear layers. These anatomic abnormalities were accompanied by reproducible changes in visual function, as measured with the electroretinogram (ERG) and visual evoked potential (VEP). The dark-adapted ERGs of nervous and control mice had similar waveforms, although the nervous responses were substantially smaller in amplitude. The reductions in the amplitude of the ERG a-wave corresponded to the loss of photoreceptor cells and shortened outer segments seen histologically. Nevertheless, the kinetics of the leading edge of the a-wave did not differ between nervous and control mice, indicating that the rod outer segments of nervous mice continue to respond to light in a normal fashion. The amplitudes of cone ERGs were also reduced in nervous mice, although the extent of this reduction in any given animal was always less than that for rod-mediated ERG components. Overall, this result is consistent with cone involvement occurring only as a secondary effect of rod photoreceptor degeneration. The peak latencies of VEPs of nervous mice were slower than those of control littermates. These functional abnormalities correspond well to the structural changes induced by the nervous mutation, which does not appear to prevent visual signals from being transmitted centrally, beyond the limitations imposed by the degenerative process. PMID- 10865996 TI - Lomerizine, a Ca2+ channel blocker, reduces glutamate-induced neurotoxicity and ischemia/reperfusion damage in rat retina. AB - We examined the effects of a new Ca2+ channel blocker, lomerizine, on the intraocular hypertension-induced ischemia/reperfusion injury in rat retina and on the glutamate-induced neurotoxicity in rat cultured retinal neurons, and compared its effects with those of a Ca2+ channel blocker (flunarizine) and an N-methyl-D aspartate receptor antagonist (MK-801). Morphometric evaluation at 7 days after ischemia/reperfusion showed that treatment with lomerizine (0.1 and 1 mg kg(-1), i.v.) prior to ischemia and again immediately after reperfusion dose-dependently reduced the retinal damage. Treatment with MK-801 (1 mg kg(-1), i.v.) before ischemia significantly reduced the resulting retinal damage. Flunarizine (0.1 and 1 mg kg(-1), i.v.) tended to reduce the retinal damage, but its effect did not reach statistical significance. In an in vitro study, pretreatment with lomerizine (0.1 and 1 microM) or flunarizine (1 microM) significantly reduced glutamate-induced neurotoxicity, the effects being concentration dependent. Lomerizine (1 microM) also exhibited protective effects against both the N-methyl D-aspartate and kainate induced types of neurotoxicity. However, lomerizine (1 microM) had little effect on the neurotoxicity induced by ionomycin (1 microM) application. Glutamate-induced neurotoxicity was abolished by removing Ca2+ from the medium. These results indicate that lomerizine protects neuronal cells against retinal neurotoxicity both in vivo and in vitro, and that this Ca2+ channel blocker may be useful as a therapeutic drug against retinal diseases that cause neuronal injury, such as normal tension glaucoma (NTG). PMID- 10865997 TI - Defective keratocyte apoptosis in response to epithelial injury in stat 1 null mice. AB - Defects in apoptosis have been noted in signal transducer and activator of transcription (Stat) 1-null cells in vitro. The purpose of this study was to analyse the keratocyte apoptosis response that occurs in vivo in response to corneal epithelial injury in Stat 1null compared with control mice and to determine whether Stat 1null corneal fibroblasts have a defective response to death receptor activation in vitro. Corneal epithelial scrape injuries were performed in Stat 1-null and wild-type mice. Keratocyte apoptosis was monitored with the quantitative TUNEL assay and confirmed using transmission electron microscopy. Corneal fibroblast apoptosis in response to tumor necrosis factor (TNF) alpha, with and without inhibitors of nuclear factor kappa B (NF-kappaB) activation, was monitored using DNA laddering and the methylene blue assay. Significantly less keratocyte apoptosis was noted in Stat 1-null mice compared with wild-type controls. TNF alpha-induced apoptosis only occurred in wild-type mice in the presence of inhibitors of NF-kappaB activation. Corneal fibroblast TNF alpha-induced apoptosis was defective in Stat 1null corneal fibroblasts whether NF-kappaB activation was blocked or not. Stat 1 has an important role in the keratocyte apoptosis that occurs in response to corneal epithelial injury. Previous studies suggest that the defect is due to a lack of constitutive expression of caspases. This study demonstrates that this defect in apoptosis in Stat 1-null mice is present in vivo in Stat 1-null mice and suggests that Stat 1 could be a therapeutic target for transient inhibition of keratocyte apoptosis to modulate corneal wound healing. PMID- 10865998 TI - Evaluation of in vivo cytokine expression in EAU-susceptible and resistant rats: a role for IL-10 in resistance? AB - Messenger RNAs for six cytokines (IL-12p40, IFN-gamma, IL-10, IL-4, TNF-alpha and TGF-beta1) expressed in vivo during development of experimental autoimmune uveitis (EAU) were quantitated by PCR in (uncultured) peripheral lymphoid cells and in the eyes of EAU-susceptible Lewis and EAU resistant F344 rats. Disease was induced by immunization with the R16 peptide of IRBP (in RT1B haplotype rats) or with whole IRBP (in all haplotypes). In the periphery, both Lewis and F 344 expressed similar cytokine patterns. In ocular tissues, however, only Lewis expressed elevated type 1 and inflammatory cytokines (IL-12p40, IFN-gamma and TNF alpha), coincident with onset and peak of disease. Interestingly, naive F344 rats expressed higher basal levels of IL-10 mRNA in the eyes. To examine the possible involvement of this phenomenon in resistance, basal levels of IL-10 vs susceptibility to IRBP were compared in Lewis, BN, DA. F344 and ACI strains. Lewis, BN and DA were susceptible and had low levels of IL-10 mRNA in eyes. F344 and ACI were resistant and expressed high basal levels of IL-10 mRNA. In an in vitro study, recombinant rat IL-10 (but not human or mouse IL-10) suppressed lymphocyte proliferation and IFN-gamma production by primed lymph node cells of R16 immunized rats, but did not suppress uveitogenic long-term T-cell lines polarized to the Thl phenotype, suggesting that mature effector lymphocytes in the rat may lose their ability to be suppressed by IL-10. We propose that higher expression of the IL-10 gene in ocular tissues in some rat strains may represent a mechanism that contributes to a higher threshold of resistance to EAU, but this threshold may be overcome by a more mature Thl effector with a reduced sensitivity to IL-10. PMID- 10865999 TI - Induction of apoptosis by arachidonic acid in human retinoblastoma Y79 cells: involvement of oxidative stress. AB - Arachidonic acid administration caused apoptosis in Y79 cells, as shown by typical morphological changes, phosphatidylserine externalization, chromatin condensation, processing and activation of caspase-3 and cleavage of the endogenous caspase substrate poly-(ADP-ribose)-polymerase. Arachidonic acid also caused lamin B cleavage, suggesting caspase-6 activation. Arachidonic acid treatment was accompanied by increased formation of the lipid peroxidation end products malondialdehyde and 4-hydroxy-2-nonenal, lowering in reduced glutathione content and in mitochondrial membrane potential. Inhibiting glutathione synthesis sensitized Y79 cells to apoptosis-inducing stimuli, whilst replenishing reduced glutathione attenuated arachidonic acid toxicity. Similar findings were obtained using hydroperoxyeicosatetranoic acids (oxygenated metabolites of arachidonic acid which deplete the reduced glutathione pool) and nordihydroguaretic acid, a general inhibitor of lipooxygenase pathway. which may also trigger rapid depletion of reduced glutathione. Melittin, which is known to activate phospholipase A2, also potently induced apoptosis. Arachidonic acid toxicity was inversely related to cell density. This could depend on an increased production of molecules with antiapoptotic effect; insulin-like growth factors could most likely be one of these molecules. These results propose a role for oxidative stress in the cytotoxicity induced by arachidonic acid in Y79 cells and suggest that these cells could be protected from such toxicity as long as sufficient levels of reduced glutathione and survival factors are present. PMID- 10866000 TI - Choroidal retinoic acid synthesis: a possible mediator between refractive error and compensatory eye growth. AB - Research over the past two decades has shown that the growth of young eyes is guided by vision. If near- or far-sightedness is artificially imposed by spectacle lenses, eyes of primates and chicks compensate by changing their rate of elongation, thereby growing back to the pre-lens optical condition. Little is known about what chemical signals might mediate between visual effects on the retina and alterations of eye growth. We present five findings that point to choroidal retinoic acid possibly being such a mediator. First, the chick choroid can convert retinol into all-trans-retinoic acid at the rate of 11 +/- 3 pmoles mg protein(-1) hr(-1), compared to 1.3 +/- 0.3 for retina/RPE and no conversion for sclera. Second, those visual conditions that cause increased rates of ocular elongation (diffusers or negative lens wear) produce a sharp decrease in all trans-retinoic acid synthesis to levels barely detectable with our assay. In contrast, visual conditions which result in decreased rates of ocular elongation (recovery from diffusers or positive lens wear) produce a four- to five-fold increase in the formation of all-trans-retinoic acid. Third, the choroidal retinoic acid is found bound to a 28-32 kD protein. Fourth, a large fraction of the choroidal retinoic acid synthesized in culture is found in a nucleus-enriched fraction of sclera. Finally, application of retinoic acid to cultured sclera at physiological concentrations produced an inhibition of proteoglycan production (as assessed by measuring sulfate incorporation) with a EC50 of 8 x 10(-7) M. These results show that the synthesis of choroidal retinoic acid is modulated by those visual manipulations that influence ocular elongation and that this retinoic acid may reach the sclera in concentrations adequate to modulate scleral proteoglycan formation. PMID- 10866001 TI - 1H spin-spin relaxation in normal and cataractous human, normal fish and bird eye lenses. AB - A systematic study on nuclear spin-spin relaxation of water protons in human, fish and bird eye lenses/lens nuclei is reported. The purpose of this study is to clarify the real nature of the relaxation processes not describable as a single exponential decay. The characterization of the spin-spin relaxation by a single exponential is commonly used both in literature and in MRI diagnostics. However, in our opinion, this single exponential decay hypothesis is an oversimplification that can lead to the loss of essential information. Our measurements were performed by Carr-Purcell-Meiboom-Gill (CPMG) pulse sequences on human, carp, chicken and turkey eye lenses/lens nuclei. Several hundreds of CPMG echo amplitude were detected and the time-dependence of their decay was determined by careful fitting procedures. Our results clearly rule out the single exponential decay hypothesis for eye lenses: at least two or three decaying components are observed. These phenomena need further investigations: it should be decided which of the relaxation parameters, T2I-s and A(i)-s gives the most characteristic physiological or pathological information. It is claimed that the amplitude ratio of the bound and the free water fraction carries the most characteristic information. During the cataract formation the weight of free water is raised by more than 25%. PMID- 10866002 TI - Evidence for differential signaling in human conjunctival epithelial cells adherent to laminin isoforms. AB - Both the laminin composition of the basement membrane and the keratin intermediate filament composition of the epithelial cell differs between cornea and conjunctiva, suggesting that at least some aspects of ocular surface epithelial cell differentiation may be regulated by extracellular matrix. The purpose of this study was to analyse the role of beta1 integrin in intracellular signaling pathways in human conjunctival epithelial cells adherent to laminin. In addition, the purpose was to compare the phosphorylation kinetics of signaling intermediates in cells adherent to different laminin isoforms. Cell adhesion assays, integrin clustering experiments, and integrin function blocking experiments demonstrated that beta1 but not beta4 integrin mediated human conjunctival epithelial cell adhesion to placental laminin isoforms (laminin 10/11) and induced focal adhesion kinase (FAK) tyrosine phosphorylation. Western blot analysis of cell lysates adherent to placental laminin showed that the tyrosine phosphorylation of p130Cas and FAK was maximally above constitutive levels after 60 min. In cells adherent to EHS laminin (laminin-1), the tyrosine phosphorylation kinetics of tensin, p130Cas, FAK and unknown proteins of 138 kDa and 110 kDa were similar, and peaked above constitutive levels after 30 min. Tyrosine phosphorylation of a 70 kDa protein was induced by cell adhesion to EHS laminin after 5 min, and phosphorylation peaked at 15 min. In contrast, the tyrosine phosphorylation of the 70 kDa protein was undetected in cells adherent to placental laminin. Erk-1 phosphorylation and activation was not differentially modulated by conjunctival epithelial cell adhesion to laminins. However, phosphorylation and activation kinetics of Erk-2 in cells adherent to placental laminin was similar to that observed for FAK and p130Cas. Erk-2 phosphorylation and activation was essentially undetectable in cells adherent to EHS laminin. These observations suggest that human conjunctival epithelial cell adhesion to different laminin isoforms activates different intracellular signaling pathways, and provides support for the hypothesis that extracellular matrix molecules can modulate ocular surface epithelial cell differentiation via alternate signaling pathways. PMID- 10866003 TI - Preoperative fasting and administration of regular medications in adult patients presenting for elective surgery. Has new evidence changed clinical practice? PMID- 10866004 TI - Assessment of tissue oxygenation in the critically-ill. AB - It is hypothesized that tissue dysoxia and O2 debt are major factors in the development and the propagation of multiple organ failure in critically ill patients. Dysoxia is the result of an abnormal relationship between O2 supply (DO2) and O2 demand and translates into increased anaerobic metabolism and tissue and blood lactate concentration. First-line therapeutic strategies used to avoid the development of an O2 debt involve correction of cardiac output, haemoglobin, and O2 saturation in order to increase DO2 above its critical value. They are not sufficient, however, to ensure appropriate end-organ perfusion and oxygenation. The adequacy of cardiac output towards tissue metabolic requirements may be appreciated by venous-to-arterial and gut mucosal-to-arterial PCO2 differences. This review details these strategies and discusses their usefulness in current practice. PMID- 10866005 TI - Unsuspected cardiac lesions associated with sudden unexpected perioperative death. AB - The retrospective analysis of 1700 forensic autopsies over 17 years (1981-97) following unexpected sudden cardiac death revealed a group of 50 cases that could have been related to surgery and/or anaesthesia. Patients were young with no history of cardiac disease. Surgery was performed for uncomplicated disorders, all classified as ASA 1. Cardiac arrest took place at induction of anaesthesia in 16% of cases, during surgery in 64% and at the end of surgery in 20%. Investigation and expertise reports ordered by the public prosecutor revealed none of the typical causes of death usually associated with surgery or anaesthesia. Pathological examination showed cardiac lesions in 47 cases: arrhythmogenic right ventricular cardiomyopathy in 18 cases, coronary artery disease in 10 cases, cardiomyopathy in eight cases, structural abnormalities of the His bundle in nine cases, mitral valve prolapse in one case, and acute myocarditis in one case. Identification of the cause of death of patients at low risk may provide major relief to the family of the patient and the medical staff. PMID- 10866006 TI - The effects of atropine on dynamic compliance in healthy intubated adults. AB - In order to compare the effect of atropine and sodium chloride on the dynamic compliance of the respiratory system after tracheal intubation, we studied 20 patients allocated randomly into two groups to receive either: atropine after 5 min of steady state and sodium chloride after 10 min (group A) or in reverse order (group B) intravenously. The study was conducted in a randomized double blinded manner. The patients were anaesthetized with thiopental 5 mg kg(-1) followed by thiopental 50 mg intravenously, as required. Intubation was facilitated by atracurium 0.5 mg kg(-1) intravenously and fentanyl 200 microg intravenously. During fixed volume ventilation (100 mL kg(-1), f=10), compliance and end-tidal carbon dioxide were measured every 10 s by a Datex AS/3-respiratory module connected to a portable IBM-pc. Five minutes was allowed to establish a steady state then either atropine or sodium chloride was administered according to the protocol. Respiratory dynamic compliance increased significantly after intravenous administration of atropine (P < 0.05). We conclude that atropine 1.0 mg given intravenously provides protection against an intubation-induced decline in respiratory dynamic compliance. PMID- 10866008 TI - Substances used for local and general anaesthesia in major surgery suppress proliferative responsiveness of normal rat peripheral blood mononuclear cells in culture. AB - We studied the direct effect of intravenous anaesthetics, local anaesthetics and premedication drugs in common use for major surgery, on the spontaneous versus lectin induced proliferation of cultured normal rat peripheral blood mononuclear cells, not exposed to surgical or any other trauma prior to culture. Peripheral blood mononuclear cells were incubated in cell-culture medium in the presence or in the absence of propofol, ketamine, fentanyl, midazolam, thiopental sodium, lidocaine or etomidate. The cells proliferated either spontaneously or under stimulation with a lectin phytohaemagglutinine P for 72 h. The proliferation rate was evaluated by 2H-Thymidine incorporation. Fentanyl, thiopental sodium, lidocaine and etomidate significantly inhibited phytohaemagglutinine P induced 3H Thymidine incorporation in cultured rat peripheral blood mononuclear cells. Counts per minute of the cultures treated with these drugs were 1667.80 +/- 745.72, 1614.1 +/- 615.00, 1688.0 +/- 615.0 and 1549 +/- 560.41, respectively, compared with the phytohaemagglutinine P stimulated positive control counts per minute, 13488 +/- 4305.6 (P < 0.001 in each comparison). Propofol, ketamine and midazolam inhibited the lectin-induced cell proliferation to levels not statistically different from the baseline. Counts per minute of these cultures were 1361.90 +/- 745.73; 1108.90 +/- 751.33 and 1518.10 +/- 848.88, respectively. Compared either with the baseline 972.57 +/- 356.73 counts per minute or to the positive control culture counts per minute, 13488 +/- 4305.6 the difference was statistically significant (P < 0.001) in each comparison. All the substances tested in this study proved capable of exerting direct inhibitory effect on circulating immunocompetent cells, because the latter were not subjected to any other immunosuppressive factor, be it operative trauma, blood transfusion, malnutrition, drug abuse, prior to culture. The possible theoretical and practical implications are discussed in this study. PMID- 10866007 TI - Continuous spinal anaesthesia/analgesia for the perioperative management of high risk patients. AB - The intraoperative effects of continuous spinal anaesthesia, and the efficacy of postoperative continuous spinal analgesia in 48 elderly high risk patients undergoing major abdominal, vascular or orthopaedic surgery is reported. Intraoperative anaesthetic technique proved to be safe and provided satisfactory results in the immediate postoperative period. Furthermore, the postoperative analgesic regimen which involved intrathecal fentanyl and bupivacaine, and intravenous tenoxicam, provided effective analgesia for all patients. The intrathecal analgesic regimen was administered continuously through a PCA pump which had the facility to provide bolus doses when requested in predetermined lockout intervals. The mean doses of fentanyl and bupivacaine infused intrathecally for the first 24 h postoperatively were 14.5 +/- 1.5 microg h(-1) (mean +/- SD) and 0.72 +/- 0.08 mg h(-1) (mean +/- SD), respectively, while the requirements for analgesia decreased progressively overtime but lasted for 118 h. The technique provided effective analgesia with low pain scores that was reflected by the ease in performing physical exercises and the pleasant co operation with the physiotherapist. Only minor complications related to anaesthesia/analgesia were encountered. PMID- 10866009 TI - Acid-base equilibrium during capnoretroperitoneoscopic nephrectomy in patients with end-stage renal failure: a preliminary report. AB - We have studied the acid-base equilibrium in 12 patients with end-stage renal failure (ESRF) during capnoretroperitoneoscopic nephrectomy. Bupivacaine (12 mL, 0.375%) and morphine (2mg) were given in the lumbar epidural space, and fentanyl (0.5 microg kg(-1)) and midazolam (50 microg kg(-1)) were given intravenously. Anaesthesia was induced by thiopental, maintained with halothane carried by oxygen enriched air (inspired oxygen fraction = 0.35), and ventilation was achieved with a tidal volume of 10 mL kg(-1) at a rate of 12 min(-1). This procedure resulted in a mild degree of respiratory acidosis that was cleared within 60 min. We conclude that capnoretroperitoneoscopic nephrectomy can be performed in patients with end-stage renal failure with minimal transient respiratory acidosis that can be avoided by increased ventilation. PMID- 10866010 TI - Postoperative nausea and vomiting in women following breast surgery: an audit. AB - A prospective observational study of 101 women having breast surgery under general anaesthesia was performed to assess the extent of postoperative nausea and vomiting in this group of women. The overall rate of postoperative nausea and vomiting was high, 56% and 41%, respectively, and was significantly associated with length of surgical procedure and morphine use. Techniques to reduce morphine requirement should be evaluated to try and reduce this cause of morbidity. PMID- 10866011 TI - Monitoring the successful embolization of an arterio-venous fistula by a fibreoptic jugular vein catheter. AB - The management of surgical embolization of a post-traumatic carotid-cavernous fistula is reported in this study. The procedure was successfully performed with the aid of a fibreoptic intravascular catheter, which was monitoring continuously the changes of the jugular venous oxygen saturation. Jugular venous oxygen saturation monitoring effectively confirmed the success of embolization without the need for intraoperative angiography, by manifesting an abrupt return of oxygen saturation from arterial to normal venous values immediately after embolization. PMID- 10866012 TI - Nitric oxide in the treatment of fulminant pulmonary failure in a young pregnant woman with varicella pneumonia. AB - Extracorporeal membrane oxygenation is the recommended treatment for fulminant pulmonary failure due to varicella pneumonia. However, in pregnancy fetal viability during extracorporeal membrane oxygenation is generally poor resulting in either therapeutic or spontaneous abortion. The present case is to our knowledge the first report on the treatment with nitric oxide to improve oxygenation in a pregnant woman with fulminant pulmonary failure due to varicella pneumonia. Adding 20 parts per million nitric oxide to the inspiratory gas increased arterial oxygen saturation from 75 to 88%, and it could be kept at this level. Due to a vaginal bleeding, an emergency Caesarean section was performed with successful outcome for the fetus. The mother started to improve after delivery and could be weaned from nitric oxide after 5 days. We conclude that inhalation of nitric oxide may be a good alternative to extracorporeal membrane oxygenation in the treatment of fulminant pulmonary failure due to varicella pneumonia in pregnancy. PMID- 10866013 TI - A survey of peripheral nerve stimulator (PNS) use. PMID- 10866014 TI - Adding lidocaine (lignocaine) to a propofol emulsion. PMID- 10866015 TI - Current status and future directions of psychological assessment: introduction. AB - This article introduces the major themes and context for the special series on the current status and future directions of psychological assessment. The internal and external challenges to assessment are outlined along with a listing of professional publications responding to these challenges. Each of the articles in the Special Series is introduced and includes topics on the current status of assessment, survival strategies, financial efficacy, and treatment planning. PMID- 10866016 TI - Assessment practices in the era of managed care: current status and future directions. AB - This article reviews recent research to determine the impact of managed health care policies on assessment practices of psychologists. The author presents findings from several test use surveys of the 1990s, summarizes available empirical data on the managed care-assessment nexus, and shares his personal views on the effects of managed care constraints on practice and training in clinical psychology. Investigations to date seem to indicate that managed care has adversely affected testing and assessment practices with the result that clinicians are performing less testing overall and are restricted in terms of their pool of assessment instruments. Personality assessment appears to be most affected with a shift away from lengthy measures toward more brief, symptom focused testing instruments. PMID- 10866017 TI - New areas for psychological assessment in general health care settings: what to do today to prepare for tomorrow. AB - Unique roles of the professional psychologist are outlined with respect to increasingly restrictive utilization practices of managed care. Suggestions of how to develop less traditional venues of practice, the types of instruments to use, and report formats, along with ways to persuade primary care physicians (PCPs), managed care organizations, and facilities to utilize psychological assessment services are provided. Medical cost offsets, cost-efficient quality of care, and models of practice are also discussed. PMID- 10866018 TI - Financial efficacy of clinical assessment: rational guidelines and issues for future research. AB - Whereas the financial efficacy of assessment has been well documented in industrial/organizational settings, there is no comparable literature within the field of clinical assessment. This has become a significant issue due to the increasing cost-consciousness of managed health care organizations. A rational means of increasing the financial efficacy of assessment is outlined, which includes the following: (a) focus on domains most relevant to treatment planning and outcome; (b) use formal assessment for risk management; (c) target problems most likely to result in cost savings (i.e., dissociation, somatization, panic); (d) increase the use of computer-assisted assessment; (e) use time-efficient instruments; (f) more closely link assessment, feedback, and therapy; and (g) integrate treatment planning, monitoring progress, and evaluating outcome. Issues and strategies for developing a research program include clearly defining what should be considered a cost, enumerating possible outcome variables, optimal follow-up time frame, selection of instruments, and the function of assessment. PMID- 10866019 TI - Making assessment relevant to treatment planning: the STS Clinician Rating Form. Systemic Treatment Selection. AB - This article reports on the development of a computerized, clinician-based method of assessing patient variables that contribute to differential assignment of psychotherapy models. In the current health care environment, omnibus personality tests and current state measures are both expensive and insensitive to most of the empirically defined patient characteristics that have been identified as indicators of different treatment qualities. Moreover, reliance on patient self report introduces a variety of distortions and limits predictive studies and follow-up assessments to those who are cooperative. These factors limit the usefulness of the currently available procedures. Aptitude-Treatment-Interaction (ATI) research has revealed differential effects of manualized cognitive, interpersonal, and insight-oriented treatments as a function on a select number of relatively specific personality and symptom qualities. A cost- and time efficient method of measuring these dimensions could increase the power of treatment by identifying in advance those that are likely to be most effective for a given patient. The STS (Systematic Treatment Selection) Clinician Rating Form is a relatively brief, clinician-based measure of a variety of patient dimensions, including subjective distress, various aspects of coping style, and resistance traits. Current data reveal good interrater reliability and adequate levels of discriminant and convergent validity. PMID- 10866020 TI - Childhood predictors of violent behavior. AB - In this study, prospective childhood data from birth to 7 years of age were used to determine predictors of subsequent violent behavior. The childhood predictors found for the girls accounted for more variance in the reported later violent offenses: 24.9% compared to 3.8% for the boys. This large difference may be due partly to the fact that violent behavior is more unusual among girls; thus, those girls who are violent are more readily differentiated from the others. The only two early childhood variables that were found to predict for both genders were: (a) less normal behavior and (b) presence of deviant or stereotyped behavior. Thus, abnormal behavior in childhood can be considered to be a fairly reliable predictor to greater likelihood of later violent behavior. PMID- 10866021 TI - Testing differences in response trends across a normalized time domain. AB - A two-stage mixed model analysis of repeated measurement calculates participant specific regression slopes relating change in available measurements to associated assessment times, and then the difference between mean regression slopes in two or more treatment groups is tested for significance against the within-groups variability of the participant-specific regression slopes. It is not necessary that all participants have the same schedule or number of repeated measurements. However, when dropouts are included in an "intent to treat" analysis, the shortened treatment exposures for the dropouts substantially increase variability and reduce power of tests for differences in rates of change. Previous work has suggested that normalizing the time scale to unit length for all participants prior to fitting the individual regression equations materially reduces the power attenuation produced by dropouts. This article reports a more detailed evaluation of the enhanced robustness against dropouts that is achieved by rescaling the time dimension. The robust analysis is recognized to be equivalent to weighting ordinary least squares regression on the original time scale by the duration of treatment for each participant. Slope coefficients calculated across a shortened time span for dropouts are less stable, so they are given less weight in defining the (linear) treatment effects. PMID- 10866022 TI - Longitudinal relationship between maternal depression and infant temperament in a Japanese population. AB - To investigate the relationship between maternal depression and infant temperament in a Japanese population, a prospective questionnaire survey was administered in the postpartum period. Postnatal depression was assessed by Zung's (1965) Self-Rating Depression Scale on two occasions (5 days and 12 months after delivery). At 6 months and 18 months after birth, infant temperament was assessed using the Revised Infant Temperament Questionnaire (RITQ; Carey & McDevitt, 1978) and the Toddler Temperament Scale (TTS; Fullward, McDevitt, & Carey, 1984), respectively. Of the five temperamental dimensions of the RITQ and TTS, "rhythmicity" and "attention span and persistence" showed reciprocal relationships with postnatal depression. Unidirectional effects of maternal depression on infant temperament were found for "frustration tolerance" and "fear of strangers and strange situations." PMID- 10866023 TI - Why, why, why?: Reason-giving and rumination as predictors of response to activation- and insight-oriented treatment rationales. AB - This study examines the relationships among the reasons a person offers for depression, the tendency to ruminate in response to depression, and reactions to activation-oriented (AO) or insight-oriented (IO) treatment rationales. Adults from the community (N=51) completed self-report measures of reason-giving and rumination and rated the credibility of, and personal reactions to, AO and IO rationales presented in written and videotape formats. Participants who gave more reasons for depression also tended to ruminate more in response to depressed mood. Reason-giving and rumination predicted lower credibility ratings and more negative personal reactions to the AO rationale. Although no relationship was found between these variables and response to the IO rationale, specific reasons were associated with different reactions to the two rationales. We discuss the roles of reason-giving and rumination in predicting responses to psychotherapies for depression. PMID- 10866024 TI - Relating PACL measures of Millon's basic personality styles and MMPI-2 scales in patient and normal samples. AB - Millon's basic personality styles as measured by the Personality Adjective Check List (PACL; Strack, 1987, 1991b) were linked to Minnesota Multiphasic Personality Inventory-2 (MMPI-2; Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) basic scales via bivariate correlation and factor analysis in independent samples of psychiatric patients (N = 196) and normal adults (N = 124). Consistent with previous research, Millon's neurotic, introverted styles were positively associated with MMPI-2 scales measuring introversion, affective states, and disturbed thinking, whereas extroverted, socially dominant Millon styles were negatively associated to the same scales. Millon personalities and MMPI-2 scales were reliably associated along two bipolar dimensions measuring Neuroticism/Introversion versus Extroversion and Emotional Distress versus Emotional Stability, which accounted for 45% of the variance. A third General Distress factor loaded only MMPI-2 scales. Congruency coefficients indicated that the factors for patients and normal participants were very similar. Results highlighted the consistency of the links between MMPI-2 basic scales, the PACL, and other Millon instruments, as well as the utility of the PACL as a measure of Millon's personality styles in a mental health population. PMID- 10866025 TI - Parent training for attention-deficit hyperactivity disorder: parental and child outcome. AB - Thirty-four patients who had children with Attention-Deficit Hyperactivity Disorder (ADHD) participated in a group parent training program (PT) that taught them about ADHD and behavior management. All parents completed pre- and postparticipation questionnaires measuring parental knowledge of ADHD and behavior management, parental stress in managing their youngsters, and the severity of their child's problem behaviors. Main effects were found indicating an increase in parental knowledge and a modest decrease in parental stress. However, behavioral improvement of their youngsters was not found at the completion of the program. This finding was discussed in the context of a possible ceiling effect from the medications that the youngsters were using to treat their ADHD. PMID- 10866026 TI - Prenatal illness and subtypes of schizophrenia: the winter pregnancy phenomenon. AB - Seasonality of birth patterns is examined among a sample of 801 patients with schizophrenia separated into Type I and Type II schizophrenia. Findings indicate that both groups of patients were in utero during the winter but during different trimesters. Patients with Type I schizophrenia were more likely to be in utero during the third trimester. Patients with Type II schizophrenia were more likely to have been in the womb during the first trimester. Implications for fetal brain development are discussed. PMID- 10866027 TI - Premalignant markers for breast cancer. Proceedings of the 15th annual symposium of the European Cancer Prevention Organization. Bruges, Belgium, June 26-28, 1997. PMID- 10866028 TI - BRCA1 and BRCA2 mutations in Belgian families with a history of breast and/or ovarian cancer. AB - Certain familial breast and/or ovarian cancers, specially those diagnosed early, are dominantly heritable and have been linked to mutations in BRCA1 and BRCA2 genes. We have tested 30 women selected from 25 different families with specific criteria. Blood samples were always taken with the informed consent and preliminary interview of the patient by a physicologist specialized in presymptomatic testing. Mutation detection were performed by protein truncation test (PTT), gradient gel electrophoresis (DGGE) and subsequent sequencing. The results showed four frameshift mutations among which three induced truncation of the BRCA1 protein and one of the BRCA2 protein. One of the BRCA1 mutations and the only BRCA2 mutation are prevelant among caucasians. Interestingly, one BRCA1 mutation is shared both by Dutch and French families and another one has not yet been reported. Furthermore, a new unclassified variant was identified. CONCLUSION: by using specific selection criteria, we have been able to detect BRCA mutations in four out of the 25 families tested. One of the mutations seems to be found only in Belgium. Genetic counselling is being offered to their relatives. PMID- 10866029 TI - Germline BRCA1 mutations in patients from 84 families with breast and/or ovarian cancers in northern France. AB - The BRCA1 gene modification is responsible for an autosomal dominant syndrome of inherited early onset breast and/or ovarian cancer. This gene is estimated to account for almost half of inherited breast cancers and three quarters of inherited breast/ovarian cancers. This suggests that about 1 in every 500 women may carry the BRCA1 mutation. The BRCA1 was isolated by positional cloning in 1994. More than 100 different mutations have been found in the germline of affected individuals. Using systematic sequencing, we looked at BRCA1 germline mutations in 84 patients treated at the Centre Oscar Lambret for breast and/or ovarian cancer who belonged to high-risk families. We found 39 mutations: 22 true mutations inducing modifications of the BRCA1 protein (BRCA1+), six mutations with unknown consequences on the BRCA1 protein, and eleven mutations corresponding to polymorphisms that had been described previously. All the BRCA1+ cases had a HPG3 tumour. The median age of discovery and the receptor positivity percentage are lower in hereditary breast cancer than in the standard population of the breast cancers treated in our centre. Conversely, most of the BRCA1+ patients are without node involvement. This shows that BRCA1 mutations are not always related to parameters thought to indicate a bad prognosis. PMID- 10866030 TI - Risk factors for breast cancer at various ages. AB - A brief overview is presented of breast cancer risk factors with particular reference to the time in a woman's life when they seem to operate. High incidence of the disease in Western societies is associated with early ovarian maturation and late onset of reproduction. In biologic terms this means proliferation without differentiation of breast cells over a long period of time. PMID- 10866031 TI - Inadvertent exposure to xenoestrogens. AB - Over the last 40 years there have been constant reports concerning environmental chemicals with hormone-like effects in wildlife. An endocrine disruptor is an exogenous substance that causes adverse health effects in an intact organism or its progeny, secondary to changes in endocrine function. Endocrine disruptors of widely diverse chemical structures that have oestrogenic properties are known as oestrogenic xenobiotics or xenoestrogens. Some of these substances, such as phytoestrogens and mycoestrogens, can come from diet or from the environment. Although the oestrogenic activity of these substances is weaker than that of oestradiol, new chemicals with endocrine disrupting potential continue to be discovered, inadvertent forms of exposure are constantly being identified, and there is increasing concern about cumulative effects. Studies in the 1960s and 1970s characterized the oestrogenicity of a number of industrial compounds and the pesticides o,p-DDT, kepone, methoxychlor, phenolic derivatives and polychlorinated biphenyls (PCBs). In the last 5 years, several environmental chemicals have been added to the list of xenoestrogens, including the pesticides toxaphene, dieldrin and endosulphan, and several different compounds used in the food industry, antioxidants such a t-butylhydroxyanisole; plasticizers such as benzylbutylphthalate and 4-OH-alkylphenols; and substances used in dental restorations, such as bisphenol-A. The relevance of these newly discovered endocrine disruptors to human health is now starting to emerge. The few studies that have investigated their effect in humans point in the same direction: if there is indeed an association between exposure to substances with hormone disruptive activity and certain disorders of endocrine organs, the incidence of such disorders would be greater in areas where exposure to agents with this activity is high. A closer scrutiny is required to determine whether these newly discovered endocrine disrupting chemicals contribute, together with oestrogenic pesticides, to the exposure of humans to xenoestrogens. PMID- 10866032 TI - Mechanisms of steroid hormone action and resistance in endometrial and breast cancer. AB - The detection of molecular alterations that lead to the development, progression, and formation of metastases in human endometrial and breast cancer may contribute to a better understanding of tumour biology as well as the development of specific preventative and therapeutic strategies. Endometrial and breast cancers both emerge during a multistep process. Cytogenetic and molecular genetic analysis of cancer samples suggest that tumour development involves: (1) alterations of hormonal interactions, and (2) accumulation of various genetic alterations. Steroid hormones act directly via corresponding steroid hormone receptors or indirectly via alterations of protein kinases or (proto-)oncogenes. Oncogene amplification with concomitant overexpression of the oncoprotein seems to be specific for certain cancer types and to mediate cellular proliferation. Loss of normal tumour suppressor protein function can occur through sequential gene mutation events (somatic alteration) or through a single mutational event of a remaining normal copy, when a germline mutation is present. The second event is usually chromosome loss, mitotic recombination, or partial chromosome deletion. These alterations of interactions or different regulatory cellular pathways may lead to primary or secondary hormonal resistance during therapeutic interventions. PMID- 10866033 TI - Detection of novel genes that are up-regulated (Di12) or down-regulated (T1A12) with disease progression in breast cancer. AB - Carcinogenesis is a multistep process and it is believed that seven to ten alterations are needed to convert a normal cell into a maligant one. Thus, identification of these specific genetic lesions and their role in the progression of cancer will lead to more effective methods for early diagnosis, as well as provide targets for treatment and therapeutic intervention. To isolate a number of genes that are differentially expressed in human breast carcinomas we used subtractive hybridization and differential display cloning techniques. By using these methods we obtained 952 clones, characterized 288 clones by restriction mapping, and 105 by Northern blotting and DNA sequencing. Twenty-four clones were found to be previously unidentified, unique genes. Full-length complementary (c)DNA was isolated from clone T1A12 and Di12, and further characterized by computer data search. Polyclonal antibodies were generated against N- and/or C-terminal peptides of the predicted T1A12 and Di12 amino acid sequences and immunohistochemistry was used to delineate gene function in breast cancers. We found that T1A12 is a novel member of the insulin-like growth factor binding protein family and that its expression decreases with disease progression. In 60 primary breast tissues examined, strong T1A12-specific immunoperoxidase staining was observed in luminal epithelial cells of normal lobules or ducts, and in blood vessels. Moderate protein expression was detected in hyperplastic and ductal carcinoma in situ (DCIS) cells, but no staining was found in invasive ductal carcinoma (IDC) cells. On the contrary, by using the same primary breast specimen Di12, a gene which shares no sequence homology with any protein, was found to be overexpressed in IDC cells but was not detectable in normal breast tissues. Specific strong Di12 staining was seen in the cytoplasm, with a slight increase in perinuclear regions of less well differentiated cells similar to those present in ductal carcinomas with poor prognosis. Both the T1A12 and Di12 genes, studied alone or in combination, could prove valuable for understanding the biology of breast cancer, detecting early and late stage disease, and for selecting appropriate treatments. PMID- 10866034 TI - Benign lesions and cancer of the breast. AB - The retroprospective study of breast cancer in relation to benign breast lesions (BBL) involved an analysis of the breast cancer incidence in a cohort of women with a history of BBL. This cohort was formed on the basis of histological and cytological investigations performed during 1982-1991. A total of 10,776 cases with BBI were recorded, followed-up and analyzed. The total person-years of follow-up was 60,872. A total of 35 women with breast cancer were detected during the study. Cohort members with a BBL history were stratified into six subcohorts with respect to morphological and cytological patterns. Comparison of the observed breast cancer incidence with the expected breast cancer incidence calculated on the basis of the age-adjusted breast cancer incidence in the general population showed no significant rise in breast cancer incidence in the whole BBL cohort. The ratio of observed to expected incidence rates was 1.16. The analysis of subcohorts with diverse BBL patterns demonstrated a marked increase in breast cancer incidence only in the intraductal papilloma and cyst subcohorts. The ratios of observed and expected rates were 5.4 and 1.6, respectively. There was no significant difference from population levies of breast cancer risk in subcohorts with history of fibroadenoma, fibrocystic disease, breast nipple discharge and other lesions. Similar results were obtained in the prospective part of the study. PMID- 10866035 TI - Mode of breast cancer detection: a study from the German part of the Maas-Rhine EUREGIO. AB - Cancer screening programmes differ throughout the European Union with regard to their content as well as their acceptance by the population. In Germany, mammography is not yet part of the recommended screening programme, although its routine use is recommended by several national and international institutions. We were interested in the present methods of breast cancer detection and the correlation to tumour stage, histology and prognosis. Patients with breast cancer, presenting in our department between January 1990 and December 1994 (1,050 cases), were asked whether the suspicious finding was first detected by themselves, their physician, or in routine mammography. Seventy-two per cent of tumours were detected by patients themselves, 12% by the physician at routine cancer screening or for other reasons, and 16% were found in mammography performed without clinical suspicion of cancer. Tumours found by physicians or by mammography were treated much sooner than those first recognized by the patient and, thus, were of lower T and N stages. Surgery could more often be breast conserving in these cases. Of the T2 stages, as far as can be determined after the short follow-up, patients with tumours detected by screening showed a better survival rate. In spite of the introduction of cancer screening programmes most breast carcinomas are still detected by patients themselves. Therapy is often started after a great delay, so that the tumours are found to be at more advanced stages and show a worse prognosis than those detected by clinical examination or mammography. PMID- 10866036 TI - Breast cancer risk and measured mammographic density. AB - It has been well established that there is a positive correlation between the dense appearance of breast stroma and parenchyma on a mammogram and the risk of breast cancer. Subjective assessment by radiologists indicated relative risks on the order of 4 to 6 for the group of women whose mammograms showed a density of over 75% or more of the projected area compared to those with an absence of density. In order to obtain a more quantitative, continuous and reproducible means of estimating breast density, which is sensitive to small changes, we have developed quantitative methods for the analysis of mammographic density, which can be applied to digitized mammograms. These techniques have been validated in a nested case-control study on 708 women aged 40-59 years (on entry) who participated in a national mammographic screening study. An interactive image segmentation method and two completely automated techniques based on image texture and grey scale histogram measures have been developed and evaluated. While our methods all show statistically significant risk factors for dense breasts, the interactive method currently provides the highest risk values (relative risk 4.0, 95% confidence interval (CI) = 2.12-7.56) compared to a measure based on the shape of the image histogram (relative risk 3.35, 95% CI = 1.57-7.12) or the fractal dimension of the mammogram (relative risk 2.54, 95% CI = 1.14-5.68). All methods were highly consistent between images of the left and right breast and between the two standard views (cranio-caudal and medio-lateral oblique) of each breast, so that studies can be done by sampling only one of the four views per examination. There is a large number of factors in addition to breast density which affect the appearance of the mammogram. In particular, the assessment of density is made difficult where the breast is not uniformly compressed, e.g. at the periphery. We have designed and are currently evaluating an image processing algorithm that effectively corrects for this problem and have considered methods for controlling some of the variables of image acquisition in prospective studies. Measurements of breast density may be helpful in assigning risk groups to women. Such measurements might guide the frequency of mammographic screening, aid the study of breast cancer aetiology, and be useful in monitoring possible risk-modifying interventions. Using our techniques, we have been able to show that reduction of the proportion of fat in the diet can result in reductions of breast density, although the direct connection to risk has not yet been made. The relationship between breast density and hormone-related and genetic factors is also of great interest. It is often not possible or ethical to obtain mammograms on some groups of women for whom information on density would be very useful. This includes younger women as well as groups in which it would be desirable to obtain such information at frequent intervals. For this reason, we are exploring the use of imaging approaches such as ultrasound and magnetic resonance imaging, which do not require ionizing radiation, to make measurements analogous to those now being performed by using X-ray mammograms. PMID- 10866037 TI - Ductal orientated sonography improves the diagnosis of pathological nipple discharge of the female breast compared with galactography. AB - In case of abnormal nipple discharge of the female breast galactography is a recognized radiological procedure to identify and to localize intraductal growths or other ductal abnormalities. However, it harbours several methodological problems. In ductal orientated sonography the availability of high frequency linear transducers enables us now to visualize the mammary ducts in detail. The aim of this study was to compare both methods with respect to the detection of the cause of pathological nipple discharge. Thirty-five patients were first examined by ductal orientated sonography with a 13 MHz linear transducer. The results of sonography were recorded in detail before galactography was performed. Ductal abnormalities were found by sonography in 26 patients and by galactography in 19 patients. In 24 cases both methods revealed concurring results. Among those, 17 cases showed pathological findings and seven cases had normal ducts. In nine patients sonography revealed pathological results whereas galactography was completely normal. In only two cases sonography failed to show ductal abnormalities which were detected by galactography. Our study underlines that ductal orientated sonography is a promising method of diagnosis in abnormal nipple discharge, which may be recommended to be performed routinely before galactography. PMID- 10866038 TI - The perils of portliness: causes and consequences of visceral adiposity. AB - Although an individual's total fat mass predicts morbidities such as coronary artery disease and diabetes, the anatomical distribution of adipose tissue is a strong and independent predictor of such adverse health outcomes. Thus, obese individuals with most of their fat stored in visceral adipose depots generally suffer greater adverse metabolic consequences than similarly overweight subjects with fat stored predominantly in subcutaneous sites. A fuller understanding of the biology of central obesity will require information regarding the genetic and environmental determinants of human fat topography and of the molecular mechanisms linking visceral adiposity to degenerative metabolic and vascular disease. Here we attempt to summarize the growing body of data relevant to these key areas and, in particular, to illustrate how recent advances in adipocyte biology are providing the basis for new pathophysiological insights. PMID- 10866039 TI - Serine residues 1177/78/82 of the insulin receptor are required for substrate phosphorylation but not autophosphorylation. AB - Serine residues of the human insulin receptor (HIR) may be phosphorylated and negatively regulate the insulin signal. We studied the impact of 16 serine residues in HIR by mutation to alanine and co-overexpression in human embryonic kidney (HEK) 293 cells together with the docking proteins insulin receptor substrate (IRS)-1, IRS-2, or (SHC) Src homologous and collagen-like. As a control, IRS-1 was also cotransfected with an HIR with a juxtamembrane deletion (HIR delta JM) and therefore not containing the domain required for interaction with IRS-1. Coexpression of HIR with IRS-1, IRS-2, and SHC strongly enhanced tyrosine phosphorylation of these proteins. A similar increase in tyrosine phosphorylation was observed in cells overexpressing IRS-1, IRS-2, or SHC together with all HIR mutants except HIR delta JM and a mutant carrying exchanges of serines 1177, 1178, and 1182 to alanine (HIR1177/78/82), although this mutant showed normal autophosphorylation. Analysis of total cell lysates with anti phosphotyrosine antibodies showed that in addition to the overexpressed substrates, other cellular proteins displayed reduced levels of tyrosine phosphorylation in these cells. To study consequences for phosphatidylinositol 3 kinase (PI 3-kinase) activation, we established stable NIH3T3 fibroblast cell lines overexpressing wild-type HIR, HIR1177/78/82, and other HIR mutants as the control. Again, HIR1177/78/82 showed normal autophosphorylation but showed a clear decrease in tyrosine phosphorylation of endogenous IRS-1 and activation of PI 3-kinase. This decrease in kinase activity also occurred in an in vitro kinase assay towards recombinant IRS-1. Finally, we performed a separation of the phosphopeptides by high-performance liquid chromatography and could not detect any differences in the profiles of HIR and HIR1177/78/82. In conclusion, we have defined a region in HIR that is important for substrate phosphorylation but not autophosphorylation. Therefore, this mutant may provide new insights into the mechanism of kinase activation and substrate phosphorylation. PMID- 10866040 TI - 5-aminoimidazole-4-carboxamide riboside mimics the effects of insulin on the expression of the 2 key gluconeogenic genes PEPCK and glucose-6-phosphatase. AB - Insulin regulates the rate of expression of many hepatic genes, including PEPCK, glucose-6-phosphatase (G6Pase), and glucose-6-phosphate dehydrogenase (G6PDHase). The expression of these genes is also abnormally regulated in type 2 diabetes. We demonstrate here that treatment of hepatoma cells with 5-aminoimidazole-4 carboxamide riboside (AICAR), an agent that activates AMP-activated protein kinase (AMPK), mimics the ability of insulin to repress PEPCK gene transcription. It also partially represses G6Pase gene transcription and yet has no effect on the expression of G6PDHase or the constitutively expressed genes cyclophilin or beta-actin. Several lines of evidence suggest that the insulin-mimetic effects of AICAR are mediated by activation of AMPK. Also, insulin does not activate AMPK in H4IIE cells, suggesting that this protein kinase does not link the insulin receptor to the PEPCK and G6Pase gene promoters. Instead, AMPK and insulin may lie on distinct pathways that converge at a point upstream of these 2 gene promoters. Investigation of the pathway by which AMPK acts may therefore give insight into the mechanism of action of insulin. Our results also suggest that activation of AMPK would inhibit hepatic gluconeogenesis in an insulin independent manner and thus help to reverse the hyperglycemia associated with type 2 diabetes. PMID- 10866041 TI - Adipocyte metabolism in adipocyte fatty acid binding protein knockout mice (aP2-/ ) after short-term high-fat feeding: functional compensation by the keratinocyte [correction of keritinocyte] fatty acid binding protein. AB - Mice null for adipocyte fatty acid binding protein (AFABP) compensate by increasing expression of keratinocyte fatty acid binding protein (KFABP) (Hotamisligil et al. Science 274:1377-1379, 1996). In the present study, AFABP knockout (KO) and wild-type (WT) mice became equally obese on a high-fat diet, as judged by fat pad weights, adipocyte size, and body composition analysis. High fat feeding led to moderate insulin resistance in both WT and AFABP knockout mice, as indicated by an approximately 2-fold increase in plasma insulin. However, in the high fat-fed mice, plasma glucose levels were approximately 15% lower in the AFABP-KO mice. Adipocytes isolated from AFABP-KO and WT mice fed high- or low-fat diets exhibited similar rates of basal and norepinephrine stimulated lipolysis and insulin-stimulated rates of glucose conversion to fatty acids and glyceride-glycerol. However, basal glucose conversion to fatty acids was higher in adipocytes of AFABP-KO mice. Adipocyte tumor necrosis factor-alpha release was similarly increased by high-fat diet-induced obesity in both WT and AFABP-KO mice. As assessed by Western blot analysis, the level of KFABP protein in AFABP-KOs was approximately 40% of the level of AFABP in WT controls. The binding affinities of KFABP for long-chain fatty acids were 2- to 4-fold higher than those of AFABP, but the relative affinities for different fatty acids were similar. As for AFABP, the rate of fatty acid transfer from KFABP to model phospholipid vesicles was increased with acceptor membrane concentration and by inclusion of acidic phospholipids, indicating a similar mechanism of transfer. We conclude KFABP can functionally compensate for the absence of AFABP, resulting in no major alterations in adipocyte metabolism or fat accumulation in response to short-term feeding of high-fat diets that result in moderate hyperinsulinemia. PMID- 10866042 TI - Cow's milk consumption, HLA-DQB1 genotype, and type 1 diabetes: a nested case control study of siblings of children with diabetes. Childhood diabetes in Finland study group. AB - The evidence for the putative role of cow's milk in the development of type 1 diabetes is controversial. We studied infant feeding patterns and childhood diet by structured questionnaire (n = 725) and HLA-DQB1 genotype by a polymerase chain reaction-based method (n = 556) in siblings of affected children and followed them for clinical type 1 diabetes. In a nested case-control design in a population who had both dietary and genetic data available, we selected as cases those siblings who progressed to clinical diabetes during the follow-up period (n = 33). For each case, we chose as matched control subjects siblings who fulfilled the following criteria: same sex, age within 1 year, not from the same family, the start of the follow-up within 6 months of that of the respective case, and being at risk for type 1 diabetes at the time the case presented with that disease (n = 254). The median follow-up time was 9.7 years (range 0.2-11.3). Early age at introduction of cow's milk supplements was not significantly associated with progression to clinical type 1 diabetes (relative risk adjusted for matching factors, maternal education, maternal and child's ages, childhood milk consumption, and genetic susceptibility markers was 1.60 [95% CI 0.5-5.1]). The estimated relative risk of childhood milk consumption for progression to type 1 diabetes was 5.37 (1.6-18.4) when adjusted for the matching and aforementioned sociodemographic factors, age at introduction of supplementary milk feeding, as well as for genetic susceptibility markers. In conclusion, our results provide support for the hypothesis that high consumption of cow's milk during childhood can be diabetogenic in siblings of children with type 1 diabetes. However, further studies are needed to assess the possible interaction between genetic disease susceptibility and dietary exposures in the development of this disease. PMID- 10866043 TI - Exposure to exogenous insulin promotes IgG1 and the T-helper 2-associated IgG4 responses to insulin but not to other islet autoantigens. AB - Insulin immunization in animal models induces T-helper (Th) 2-like antibody subclass responses to insulin and other beta-cell antigens. The aim of this study was to determine whether exposure to insulin in humans resulted in a similar subclass bias of the humoral immune response. Levels of IgG subclass antibodies to insulin (IAs), GAD, and IA-2 were measured before and after treatment with insulin in the following groups of patients: 29 patients with newly diagnosed type 1 diabetes treated with intravenous and/or subcutaneous insulin; 10 newly diagnosed patients randomized to cyclosporin A (CsA) or placebo plus subcutaneous insulin for 12 months; and 14 islet cell antibody-positive relatives receiving either intravenous and subcutaneous insulin prophylaxis or no treatment. At the onset of diabetes, the major subclass distributions of insulin autoantibodies (IAAs) were IgG1 and, to a lesser extent, IgG4. After insulin treatment in the 29 new-onset patients, IAs were initially of the IgG1 subclass. IgG4-IAs appeared later, but at 12 months, they were at higher levels than IgG1-IAs in 11 patients. Responses were higher in children compared with adults and were higher in subjects with IAAs (P < 0.001). Insulin prophylaxis in relatives showed a similar profile, with a decline in levels of IgG1-IAs after cessation of daily subcutaneous insulin. Patients treated with CsA took longer to develop IAs and showed suppressed levels of IgG4-IAs; however, their levels of high-titer IgG1 IAs persistently rebounded after completion of CsA therapy. Despite the presence of IgG4-IAs in most insulin-treated patients and relatives, a shift to IgG4-anti GAD or IgG4-IA-2 was not found for up to 3 years after the initiation of insulin therapy. While our findings need to be correlated with T-cell cytokine responses, we suggest that the strong IgG4-IA response in insulin-treated patients is consistent with an enhancement of Th2 immunity, but there is no evidence of subsequent spreading of potentially Th2-associated IgG4 responses to other autoantigens. PMID- 10866044 TI - Effects of glucosamine infusion on insulin secretion and insulin action in humans. AB - Glucose toxicity (i.e., glucose-induced reduction in insulin secretion and action) may be mediated by an increased flux through the hexosamine-phosphate pathway. Glucosamine (GlcN) is widely used to accelerate the hexosamine pathway flux, independently of glucose. We tested the hypothesis that GlcN can affect insulin secretion and/or action in humans. In 10 healthy subjects, we sequentially performed an intravenous glucose (plus [2-3H]glucose) tolerance test (IVGTT) and a euglycemic insulin clamp during either a saline infusion or a low (1.6 micromol x min(-1) x kg(-1)) or high (5 micromol x min(-1) x kg(-1) [n = 5]) GlcN infusion. Beta-cell secretion, insulin (SI*-IVGTT), and glucose (SG*) action on glucose utilization during the IVGTT were measured according to minimal models of insulin secretion and action. Infusion of GlcN did not affect readily releasable insulin levels, glucose-stimulated insulin secretion (GSIS), or the time constant of secretion, but it increased both the glucose threshold of GSIS (delta approximately 0.5-0.8 mmol/l, P < 0.03-0.01) and plasma fasting glucose levels (delta approximately 0.3-0.5 mmol/l, P < 0.05-0.02). GlcN did not change glucose utilization or intracellular metabolism (glucose oxidation and glucose storage were measured by indirect calorimetry) during the clamp. However, high levels of GlcN caused a decrease in SI*-IVGTT (delta approximately 30%, P < 0.02) and in SG* (delta approximately 40%, P < 0.05). Thus, in humans, acute GlcN infusion recapitulates some metabolic features of human diabetes. It remains to be determined whether acceleration of the hexosamine pathway can cause insulin resistance at euglycemia in humans. PMID- 10866045 TI - Expression and role of laminin-1 in mouse pancreatic organogenesis. AB - Previous studies have suggested that basement membrane alone may induce ductal differentiation and morphogenesis in the undifferentiated embryonic pancreas. The mechanism by which this induction occurs has not been investigated. Studies of other organ systems such as the lungs and mammary glands, where differentiation has been shown to be induced by basement membrane, have suggested a major role for laminin as a mediator of ductal or tubular morphogenesis and differentiation. We first defined the ontogeny of laminin-1 in the developing mouse pancreas. To determine the specific role of basement membrane laminin in pancreatic ductal morphogenesis and differentiation, we microdissected 11-day mouse embryonic pancreatic epithelium free from its surrounding mesenchyme and then suspended the explants in a 3-dimensional organ culture to allow us to assay cell differentiation and morphogenesis. When the pancreatic epithelium buds off the foregut endoderm, the pancreatic mesenchyme diffusely expresses laminin-1. This laminin subsequently organizes to the interface between the epithelium and the mesenchyme by E12.5. As gestation progresses, epithelial cells in direct contact with laminin-1 seem to differentiate into ducts and acini, whereas those spared intimate contact with laminin-1 appeared to organize into islets. Although basement membrane gel could induce pancreatic ductal morphogenesis of embryonic pancreatic epithelium, this induction was blocked when we added neutralizing antibodies against any of the following: 1) laminin (specifically laminin-1), 2) the "cross-region" of laminin-1, and 3) the alpha6 moiety of the integrin receptor, which is known to bind laminins. Immunohistochemistry, however, showed that pancreatic duct cell-specific differentiation (carbonic anhydrase II) without ductal morphogenesis was still present, despite the blockage of duct morphogenesis by the anti-laminin-1 neutralizing antibodies. Interestingly, there appeared to be a decrease in carbonic anhydrase II expression over time when the epithelia were grown in a collagen gel, rather than in a basement membrane gel. The pattern of laminin-1 expression in the embryonic pancreas supports the conclusion that laminin-1 is important in the induction of exocrine (ducts and acini) differentiation in the pancreas. Furthermore, our data demonstrate that 1) pancreatic ductal morphogenesis appears to require basement membrane laminin-1 and an alpha6-containing integrin receptor; 2) the cross-region of basement membrane laminin is a biologically active locus of the laminin molecule necessary for pancreatic ductal morphogenesis; 3) duct-specific cytodifferentiation, in the form of carbonic anhydrase II expression, is not necessarily coupled to duct morphogenesis; and 4) the basement membrane gel may contain components (e.g., growth factors) other than laminin-1 that can sustain both carbonic anhydrase II expression and, possibly, the capacity to form ducts, despite the absence of duct structures. PMID- 10866046 TI - Synergism of protein kinase A, protein kinase C, and myosin light-chain kinase in the secretory cascade of the pancreatic beta-cell. AB - Protein phosphorylation by myosin light-chain kinase (MLCK), protein kinase A, and protein kinase C (PKC) plays a positive role in insulin secretion from the pancreatic beta-cell. To investigate the underlying mechanisms, we examined intracellular distribution of the insulin granules and MLCK by immunofluorescence and immunoelectron microscopies and also investigated intracellular traffic of the granules in cultured beta-cells (MIN6) by video microscopy. Considerable parts of MLCK immunoreactivity were colocalized with the insulin granules. Subcellular fractionation of MIN6 cell extracts revealed that myosin light chain (MLC) may be distributed with the insulin-rich fractions, and immunofluorescence staining using specific antibodies against mono- and diphosphorylated MLCs depicted presence of phosphorylated MLCs in the cytoplasm, in part, with colocalization with the insulin granules. Activation of PKC by 12-O-tetradecanoyl phorbol 13-acetate (TPA) caused a shift of both insulin granules and MLCK to the cell periphery, which was not reproduced by the adenylate cyclase activator, forskolin. In contrast, forskolin, but not TPA, increased the granule movement. Costimulation of the beta-cell by TPA and forskolin induced drastic translocation of insulin granules and MLCK to the cell periphery, resulting in enormous potentiation of insulin release. These findings suggest that these protein kinases increase insulin granules in the ready-releasable pool by acting on different steps in the secretory cascade. PMID- 10866047 TI - Sulfonylurea receptor 1 and Kir6.2 expression in the novel human insulin secreting cell line NES2Y. AB - NES2Y is a proliferating human insulin-secreting cell line that we have derived from a patient with persistent hyperinsulinemic hypoglycemia of infancy. This disease is characterized by unregulated insulin release despite profound hypoglycemia. NES2Y cells, like beta-cells isolated from the patient of origin, lack functional ATP-sensitive potassium channels (KATP) and also carry a defect in the insulin gene-regulatory transcription factor PDX1. Here, we report that the NES2Y beta-cells that are transfected with the genes encoding the components of KATP channels in beta-cells, sulfonylurea receptor (SUR) 1 and Kir6.2, have operational KATP channels and show normal intracellular Ca2+ and secretory responses to glucose. However, these cells, designated NESK beta-cells, have impaired insulin gene transcription responses to glucose. NES2Y beta-cells that are transfected with either Kir6.2 or SUR1 alone do not express functional KATP channels and have impaired intracellular free Ca2+ concentration-signaling responses to depolarization-dependent beta-cell agonists. These findings document that in NES2Y beta-cells, coexpression of both subunits is critically required for fully operational KATP channels and KATP channel-dependent signaling events. This article further characterizes the properties of the novel human beta-cell line, NES2Y, and documents the usefulness of these cells in diabetes-related research. PMID- 10866048 TI - Reduced pancreatic polypeptide response to hypoglycemia and amylin response to arginine in subjects with a mutation in the HNF-4alpha/MODY1 gene. AB - Subjects with the Q268X mutation in the hepatocyte nuclear factor (HNF)-4alpha gene (RW pedigree/maturity-onset diabetes of the young [MODY]-1) have diminished insulin and glucagon secretory responses to arginine. To determine if pancreatic polypeptide (PP) secretion is likewise involved, we studied PP responses to insulin-induced hypoglycemia in 17 RW pedigree members: 6 nondiabetic mutation negative [ND(-)], 4 nondiabetic mutation-positive [ND(+)], and 7 diabetic mutation-positive [D(+)]. Subjects received 0.08 U/kg body wt human regular insulin as an intravenous bolus to produce moderate self-limited hypoglycemia. PP areas under the curve (PP-AUCs) were compared among groups. With hypoglycemia, the PP-AUC was lower in the D(+) group (14,907 +/- 6,444 pg/ml, P = 0.03) and the ND(+) group (14,622 +/- 6,015 pg/ml, P = 0.04) compared with the ND(-) group (21,120 +/- 4,158 pg/ml). In addition, to determine if the beta-cell secretory defect in response to arginine involves amylin in addition to insulin secretion, we analyzed samples from 17 previously studied RW pedigree subjects. We compared the AUCs during arginine infusions for the 3 groups both at euglycemia and hyperglycemia as well as their C-peptide-to-amylin ratios. The D(+) and ND(+) groups had decreased amylin AUCs during both arginine infusions compared with the ND(-) group, but had similar C-peptide-to-amylin ratios. These results suggest that the HNF-4alpha mutation in the RW/MODY1 pedigree confers a generalized defect in islet cell function involving PP cells in addition to beta- and alpha cells, and beta-cell impairment involving proportional deficits in insulin and amylin secretion. PMID- 10866049 TI - Glucose-6-phosphatase flux in vitro is increased in type 2 diabetes. AB - Despite the effects of hyperinsulinemia and hyperglycemia, 2 factors known to inhibit endogenous glucose production (EGP) in nondiabetic subjects, increased EGP is a consistent feature of type 2 diabetes. Recent studies have suggested that increased glucose-6-phosphatase (G6Pase) and/or decreased glucokinase (GK) may explain the increase in EGP. However, no studies to date have clearly established this relationship in type 2 diabetes. The present studies were designed to determine rates of EGP and the activities of G6Pase and GK in obese patients scheduled for gastric bypass surgery. The study group consisted of 14 obese nondiabetic subjects and 13 patients with type 2 diabetes (BMI 53.7 +/- 2.4 vs. 50.1 +/- 1.6 kg/m2). Rates of EGP were determined after an overnight fast with a 4-h infusion of [6,6]-D-glucose, and they were significantly higher in the type 2 diabetic patients (85.9 +/- 10.0 vs. 137.8 +/- 14.4 mg x m(-2) x min(-1), P < 0.001) despite greater plasma glucose (5.1 +/- 0.1 vs. 12.0 +/- 1.1 mmol/l) and similar insulin concentrations (130.8 +/- 19.8 vs. 112.8 +/- 16.2 pmol/l, NS). Moreover, resistance to insulin-induced suppression of EGP was observed in the patients with type 2 diabetes when insulin concentrations were increased from approximately 120 to 180 pmol/l. Hepatic G6Pase activity determined from freshly isolated microsomes was significantly increased in the type 2 diabetic patients compared with the obese control subjects (0.16 +/- 0.02 vs. 0.09 +/- 0.01 micromol x min(-1) x mg(-1) protein, P < 0.02), whereas levels of GK were decreased (1.20 +/- 0.16 vs. 2.01 +/- 0.01 micromol x min(-1) x mg(-1) protein, P < 0.01). Net flux through G6Pase was significantly increased in type 2 diabetic patients (P < 0.01). We conclude that increased EGP is mediated in part by increased G6Pase flux in type 2 diabetes. PMID- 10866050 TI - Insulin secretion and insulin sensitivity in relation to glucose tolerance: lessons from the Botnia Study. AB - Recently, a new stage in glucose tolerance, impaired fasting glucose (IFG) (fasting plasma glucose level of 6.1-6.9 mmol/l), was introduced in addition to impaired glucose tolerance (IGT) (2-h glucose level of 7.8-11.0 mmol/l). It is not clear whether IFG and IGT differ with respect to insulin secretion or sensitivity. To address this question, we estimated insulin secretion (by measuring both insulin levels and the ratio of insulin-to-glucose levels in 30 min intervals) and insulin sensitivity (by using the homeostasis model assessment [HOMA] index) from an oral glucose tolerance test (OGTT) in 5,396 individuals from the Botnia Study who had varying degrees of glucose tolerance. There was poor concordance between IFG and IGT: only 36% (303 of 840) of the subjects with IFG had IGT, whereas 62% (493 of 796) of the subjects with IGT did not have IFG. Compared with subjects with normal glucose tolerance (NGT), subjects with IFG were more insulin resistant (HOMA-insulin resistance [IR] values 2.64 +/- 0.08 vs. 1.73 +/- 0.03, P < 0.0005), had greater insulin responses during an OGTT (P = 0.0001), had higher waist-to-hip ratios (P < 0.005), had higher triglyceride and total cholesterol concentrations (P < 0.0005), and had lower HDL cholesterol concentrations (P = 0.0001). Compared with subjects with IFG, subjects with IGT had a lower incremental 30-min insulin-to-glucose area during an OGTT (13.8 +/- 1.7 vs. 21.7 +/- 1.7, P = 0.0008). Compared with subjects with IGT, subjects with mild diabetes (fasting plasma glucose levels <7.8 mmol/l) showed markedly impaired insulin secretion that could no longer compensate for IR and elevated glucose levels. A progressive decline in insulin sensitivity was observed when moving from NGT to IGT and to subjects with diabetes (P < 0.05 for trend), whereas insulin secretion followed an inverted U-shaped form. We conclude that IFG is characterized by basal IR and other features of the metabolic syndrome, whereas subjects with IGT have impaired insulin secretion in relation to glucose concentrations. An absolute decompensation of beta-cell function characterizes the transition from IGT to mild diabetes. PMID- 10866051 TI - High glucose and glucosamine induce insulin resistance via different mechanisms in 3T3-L1 adipocytes. AB - Sustained hyperglycemia induces insulin resistance, but the mechanism is still incompletely understood. Glucosamine (GlcN) has been extensively used to model the role of the hexosamine synthesis pathway (HSP) in glucose-induced insulin resistance. 3T3-L1 adipocytes were preincubated for 18 h in media +/- 0.6 nmol/l insulin containing either low glucose (5 mmol/l), low glucose plus GlcN (0.1-2.5 mmol/l), or high glucose (25 mmol/l). Basal and acute insulin-stimulated (100 nmol/l) glucose transport was measured after re-equilibration in serum and insulin-free media. Preincubation with high glucose or GlcN (1-2.5 mmol/l) inhibited basal and acute insulin-stimulated glucose transport only if insulin was present during preincubation. However, only preincubation with GlcN plus insulin inhibited insulin-stimulated GLUT4 translocation. GLUT4 and GLUT1 protein expression were not affected. GlcN (2.5 mmol/l) increased cellular UDP-N acetylhexosamines (UDP-HexNAc) by 400 and 900% without or with insulin, respectively. High glucose plus insulin increased UDP-HexNAc by 30%. GlcN depleted UDP-hexoses, whereas high glucose plus insulin increased them. Preincubation with 0.5 mmol/l GlcN plus insulin maximally increased UDP-HexNAc without affecting insulin-stimulated or basal glucose transport. GlcN plus insulin (but not high glucose plus insulin) caused marked GlcN dose-dependent accumulation of GlcN-6-phosphate, which correlated with insulin resistance of glucose transport (r = 0.935). GlcN plus insulin (but not high glucose plus insulin) decreased ATP (10-30%) and UTP (>50%). GTP was not measured, but GDP increased. Neither high glucose plus insulin nor GlcN plus insulin prevented acute insulin stimulation (approximately 20-fold) of insulin receptor substrate 1 associated phosphatidylinositol (PI)-3 kinase. We have come to the following conclusions. 1) Chronic exposure to high glucose or GlcN in the presence of low insulin caused insulin resistance of glucose transport by different mechanisms. 2) GlcN inhibited GLUT4 translocation, whereas high glucose impaired GLUT4 "intrinsic activity" or membrane intercalation. 3) Both agents may act distally to PI-3 kinase. 4) GlcN has metabolic effects not shared by high glucose. GlcN may not model HSP appropriately, at least in 3T3-L1 adipocytes. PMID- 10866052 TI - Insulin signaling and action in cultured skeletal muscle cells from lean healthy humans with high and low insulin sensitivity. AB - The aim of these studies was to investigate whether insulin resistance is primary to skeletal muscle. Myoblasts were isolated from muscle biopsies of 8 lean insulin-resistant and 8 carefully matched insulin-sensitive subjects (metabolic clearance rates as determined by euglycemic-hyperinsulinemic clamp: 5.8 +/- 0.5 vs. 12.3 +/- 1.7 ml x kg(-1) x min(-1), respectively; P < or = 0.05) and differentiated to myotubes. In these cells, insulin stimulation of glucose uptake, glycogen synthesis, insulin receptor (IR) kinase activity, and insulin receptor substrate 1-associated phosphatidylinositol 3-kinase (PI 3-kinase) activity were measured. Furthermore, insulin activation of protein kinase B (PKB) was compared with immunoblotting of serine residues at position 473. Basal glucose uptake (1.05 +/- 0.07 vs. 0.95 +/- 0.07 relative units, respectively; P = 0.49) and basal glycogen synthesis (1.02 +/- 0.11 vs. 0.98 +/- 0.11 relative units, respectively; P = 0.89) were not different in myotubes from insulin resistant and insulin-sensitive subjects. Maximal insulin responsiveness of glucose uptake (1.35 +/- 0.03-fold vs. 1.41 +/- 0.05-fold over basal for insulin resistant and insulin-sensitive subjects, respectively; P = 0.43) and glycogen synthesis (2.00 +/- 0.13-fold vs. 2.10 +/- 0.16-fold over basal for insulin resistant and insulin-sensitive subjects, respectively; P = 0.66) were also not different. Insulin stimulation (1 nmol/l) of IR kinase and PI 3-kinase were maximal within 5 min (approximately 8- and 5-fold over basal, respectively), and insulin activation of PKB was maximal within 15 min (approximately 3.5-fold over basal). These time kinetics were not significantly different between groups. In summary, our data show that insulin action and signaling in cultured skeletal muscle cells from normoglycemic lean insulin-resistant subjects is not different from that in cells from insulin-sensitive subjects. This suggests an important role of environmental factors in the development of insulin resistance in skeletal muscle. PMID- 10866053 TI - Correlations of receptor binding and metabolic and mitogenic potencies of insulin analogs designed for clinical use. AB - In recent years, analogs of human insulin have been engineered with the aim of improving therapy for people with diabetes. To ensure that the safety profile of the human hormone is not compromised by the molecular modifications, the toxico pharmacological properties of insulin analogs should be carefully monitored. In this study, we compared the insulin and IGF-I receptor binding properties and metabolic and mitogenic potencies of insulin aspart (B28Asp human insulin), insulin lispro (B28Lys,B29Pro human insulin), insulin glargine (A21Gly,B31Arg,B32Arg human insulin), insulin detemir (NN304) [B29Lys(epsilon tetradecanoyl), desB30 human insulin], and reference insulin analogs. Receptor affinities were measured using purified human receptors, insulin receptor dissociation rates were determined using Chinese hamster ovary cells overexpressing the human insulin receptor, metabolic potencies were evaluated using primary mouse adipocytes, and mitogenic potencies were determined in human osteosarcoma cells. Metabolic potencies correlated well with insulin receptor affinities. Mitogenic potencies in general correlated better with IGF-I receptor affinities than with insulin receptor off-rates. The 2 rapid-acting insulin analogs aspart and lispro resembled human insulin on all parameters, except for a slightly elevated IGF-I receptor affinity of lispro. In contrast, the 2 long acting insulin analogs, glargine and detemir, differed significantly from human insulin. The combination of the B31B32diArg and A21Gly substitutions provided insulin glargine with a 6- to 8-fold increased IGF-I receptor affinity and mitogenic potency compared with human insulin. The attachment of a fatty acid chain to LysB29 provided insulin detemir with reduced receptor affinities and metabolic and mitogenic potencies but did not change the balance between mitogenic and metabolic potencies. The safety implications of the increased growth-stimulating potential of insulin glargine are unclear. The reduced in vitro potency of insulin detemir might explain why this analog is not as effective on a molar basis as human insulin in humans. PMID- 10866054 TI - Effects of DL-alpha-lipoic acid on peripheral nerve conduction, blood flow, energy metabolism, and oxidative stress in experimental diabetic neuropathy. AB - Experimental diabetic peripheral neuropathy (DPN) is marked by impaired nerve conduction velocity (NCV), reduced nerve blood flow (NBF), and a variety of metabolic abnormalities in peripheral nerve that have been variously ascribed to hyperglycemia, abnormal fatty acid metabolism, ischemic hypoxia, and/or oxidative stress. Some investigators propose that NCV slowing in experimental DPN can be explained entirely on the basis of nerve energy depletion secondary to reduced NBF. This article reports highly selective effects of administration of the antioxidant DL-alpha-lipoic acid (LA) to streptozotocin-injected diabetic rats. LA improved digital sensory but not sciatic-tibial motor NCV, corrected endoneurial nutritive but not composite NBF, increased the mitochondrial oxidative state without correcting nerve energy depletion, and enhanced the accumulation of polyol pathway intermediates without worsening myo-inositol or taurine depletion. These studies implicate oxidative stress as an important pathophysiological factor in experimental DPN. They reveal complex interrelationships among nerve perfusion, energy metabolism, osmolyte content, conduction velocity, and oxidative stress that may reflect the heterogeneous and compartmentalized composition of peripheral nerve. PMID- 10866055 TI - Diabetes downregulates GLUT1 expression in the retina and its microvessels but not in the cerebral cortex or its microvessels. AB - Capillaries in the retina are more susceptible to develop microvascular lesions in diabetes than capillaries in the embryologically similar cerebral cortex. Because available evidence implicates hyperglycemia in the pathogenesis of diabetic retinopathy, differences in glucose transport into the retina and brain might contribute to this observed tissue difference in susceptibility to diabetes induced microvascular disease. Thus, we compared levels of GLUT1 and GLUT3 expression in the retina, cerebrum, and their respective microvessels by Western blot analysis. In nondiabetic animals, the content of GLUT1 protein in retina and its microvessels was multifold greater than that of cerebral cortex gray matter and its microvessels. Streptozotocin-induced diabetes of a 2-week or 2-month duration reduced GLUT1 expression in the retina and its microvasculature by approximately 50%, but it resulted in no reduction in GLUT1 expression in cerebrum or its microvessels. The density of capillaries in retinas of diabetic animals did not change from normal, and so the observed decrease in GLUT1 expression in the retina and retinal capillaries of diabetic animals cannot be attributed to fewer vessels. Despite the diabetes-induced reduction of GLUT1 expression in retina, neural retina of diabetic rats still possessed more GLUT1 than the cerebrum. Retinal pigment epithelium (RPE) possessed more GLUT1 than neural retina or its microvessels, and expression of the transporter in the RPE was not affected by diabetes. GLUT3 levels were greater in cerebral gray matter than in retina, and they were unaffected by diabetes in either tissue. The effect of diabetes on GLUT1 expression differs between retina and cerebral cortex, suggesting that glucose transport is regulated differently in these embryologically similar tissues. Because diabetes results in downregulation of GLUT1 expression in retinal microvessels, but not in RPE, the fraction of the glucose entering the retina in diabetes is likely to be greater across the RPE than across the retinal vasculature. PMID- 10866056 TI - Thiazolidinedione compounds ameliorate glomerular dysfunction independent of their insulin-sensitizing action in diabetic rats. AB - Thiazolidinedione (TZD) compounds are widely used as oral hypoglycemic agents. Herein, we provide evidence showing that troglitazone, one of the TZD compounds, is able to prevent glomerular dysfunction in diabetic rats through a novel mechanism independent of its insulin-sensitizing action. We examined the effect of troglitazone on functional and biochemical parameters of glomeruli in streptozotocin-induced diabetic rats. Troglitazone was able to prevent not only diabetic glomerular hyperfiltration and albuminuria, but an increase in mRNA expression of extracellular matrix proteins and transforming growth factor-beta1 in glomeruli of diabetic rats, without changing blood glucose levels. Biochemically, an increase in diacylglycerol (DAG) contents and the activation of the protein kinase C (PKC)-extracellular signal-regulated kinase (ERK) pathway in glomeruli of diabetic rats were abrogated by troglitazone. The activation of DAG PKC-ERK pathways in vitro in mesangial cells cultured under high glucose conditions was also inhibited by troglitazone. Troglitazone enhanced the activities of DAG kinase, which could metabolize DAG to phosphatidic acid, in both glomeruli of diabetic rats and mesangial cells cultured under high glucose conditions. Surprisingly, pioglitazone, another TZD compound without alpha tocopherol moiety in its structure, also prevented the activation of the DAG-PKC pathway and activated DAG kinase in mesangial cells cultured under high glucose conditions. These results may identify the TZDs as possible new therapeutic agents for diabetic nephropathy that prevent glomerular dysfunction through the inhibition of the DAG-PKC-ERK pathway. PMID- 10866057 TI - Anti-modified LDL antibodies, LDL-containing immune complexes, and susceptibility of LDL to in vitro oxidation in patients with type 2 diabetes. AB - We investigated the hypothesis that modified lipoproteins trigger an immune response leading to the production of autoantibodies and subsequently to the formation of atherogenic immune complexes (IC). We recruited 20 type 2 diabetic patients with macrovascular disease, 14 nondiabetic patients with coronary artery disease (CAD), and 34 healthy control subjects matched for age, sex, and race. Serum antibodies to oxidized and glycated LDL did not differ significantly among the 3 groups. Serum IC contained variable, but not statistically different, amounts of IgG, IgM, and IgA. In contrast, the content of cholesterol in IC isolated from diabetic patients was significantly higher than that in IC isolated from control subjects, and the content of apolipoprotein (apo)-B was significantly higher than that in IC isolated from control subjects and patients with CAD. Cholesteryl ester accumulation in human monocyte-derived macrophages incubated with IC, a measure of the atherogenic potential of IC, was significantly higher in macrophages incubated with red blood cell-adsorbed IC isolated from diabetic patients compared with IC isolated from control subjects (P < 0.03) or from patients with CAD (P < 0.04) and was strongly correlated with the content of apoB (r = 0.68, P < 0.001) and cholesterol (r = 0.61, P < 0.001) in IC. LDL from diabetic patients was more susceptible to oxidation in vitro, was significantly smaller, and contained significantly less alpha-tocopherol than LDL isolated from subjects in the other groups. In addition, the n-3 polyunsaturated fatty acid content of phospholipids and cholesteryl esters in LDL isolated from diabetic patients was significantly increased (P < 0.05) compared with that from patients with CAD or from control subjects. We postulate that LDL size, susceptibility to oxidation, and lipid fatty acid composition may play a critical role in the production of antibodies to oxidized LDL and consequently in the formation of LDL-containing IC in patients with type 2 diabetes. PMID- 10866058 TI - Age-related patterns of the clustering of cardiovascular risk variables of syndrome X from childhood to young adulthood in a population made up of black and white subjects: the Bogalusa Heart Study. AB - The age-related patterns of clustering of cardiovascular risk variables of Syndrome X from childhood to adulthood were examined in a community-based sample of black and white children (aged 5-10 years, n = 2,389), adolescents (aged 11-17 years, n = 3,371), and young adults (aged 18-37 years, n = 2,115). In the analysis of clustering, insulin resistance index, BMI, triglycerides/ HDL cholesterol ratio, and mean arterial pressure were used either as categorical variables (age-, race- and sex-specific values >75th percentiles) to calculate risk ratios (observed frequency/expected frequency) or as continuous variables (normal scores based on ranks) to compute intraclass correlations. In the total sample, the risk ratio for clustering of adverse levels of all 4 variables was 9.8 for whites (P < 0.01) versus 7.4 for blacks (P < 0.01); the intraclass correlation was 0.33 for whites (P < 0.001) versus 0.26 for blacks (P < 0.001). Both the risk ratio and intraclass correlation were significantly higher in whites than in blacks in the total sample. The intraclass correlations of the 4 variables were significant (P < 0.001) in all race and age-groups, and they were higher during preadolescence and adulthood than during adolescence. Furthermore, unlike risk ratios, intraclass correlations showed a continuous increase with age during adulthood. When BMI was adjusted, the intraclass correlations involving the other 3 variables were reduced by approximately 50%, and the age-related pattern was no longer evident. These results suggest that the degree of clustering of risk variables of Syndrome X varies with age from childhood to adulthood and is likely influenced by the age-related changes in obesity and the attendant insulin resistance. PMID- 10866059 TI - Segregation analysis of diabetic nephropathy in Pima Indians. AB - Familial aggregation of diabetic nephropathy suggests the existence of genes determining susceptibility to nephropathy in addition to those leading to diabetes. In the present study, complex segregation analysis was performed in diabetic members of Pima Indian families to determine whether familial aggregation of nephropathy in this population could reflect the action of a single major gene. Nephropathy, defined by a urinary protein-to-creatinine ratio (PCR) > or = 500 mg/g, was analyzed as a discrete trait in a class C regressive logistic model. Individuals with PCR <500 mg/g were considered unaffected. Segregation analysis was performed both for nephropathy at the last examination (prevalent cases) and for duration of diabetes at the onset of nephropathy (incident cases). The REGD program was used for the analysis of the prevalent cases and the REGTL program for the incident cases, both from the Statistical Analysis for Genetic Epidemiology package (Case Western University, Cleveland, OH). The analysis of prevalent cases included 2,107 Pima Indians from 715 nuclear families. A subset of 504 of these families containing 1,403 individuals was used in the analysis of incident cases. Analysis of prevalent cases supported the existence of a gene with a major role, in that hypotheses of no major effect and of no transmission of a major effect were rejected (P = 0.00001; P = 0.003), whereas Mendelian transmission was not rejected (P = 0.85). A dominant model provided the best fit, but a recessive model could not be rejected. The analysis of incident cases, however, did not support a major gene effect on duration of diabetes at the onset of nephropathy, and analyses of lifetime occurrence of nephropathy were inconclusive. The analysis of prevalent cases supports the hypothesis of a major genetic effect on susceptibility to diabetic nephropathy in Pima Indians, but the analysis of incident cases does not support a genetic effect on duration of diabetes at the onset of nephropathy. The discrepancy may reflect the difficulty in precisely dating onset of nephropathy. The parameters of the model derived from segregation analysis of prevalent cases may be useful in linkage studies to detect nephropathy susceptibility loci. PMID- 10866060 TI - Segregation analysis of urinary albumin excretion in families with type 2 diabetes. AB - Elevated urinary albumin excretion (UAE) is a predictor of the development of nephropathy and cardiovascular mortality. To study whether genetic factors may determine UAE, we examined familial aggregation of UAE in 96 large multigenerational pedigrees ascertained for type 2 diabetes. A total of 1,269 subjects had UAE measured as the urinary albumin-to-creatinine ratio (ACR). This included 630 subjects with type 2 diabetes and 639 subjects without diabetes. A significant correlation (Spearman's correlation 0.34, P < 0.001) was found between the median ACR values determined separately in nondiabetic and diabetic members of the same family. To determine whether this familial aggregation of ACR could be explained by the transmission of 1 or more major genes and thus be suitable for gene mapping studies, segregation analyses were performed. In these analyses, ACR was modeled as a continuous trait with the inclusion of age, sex, and duration of diabetes as covariates. Likelihood ratio tests were performed to test competing hypotheses, and Akaike's information criterion was used to determine the most parsimonious models. The Mendelian model with multifactorial inheritance was supported more strongly than Mendelian inheritance alone. These analyses suggested that the best model for ACR levels was multifactorial with evidence for a common major gene. When the analyses were repeated for diabetic subjects only, the evidence for Mendelian inheritance was improved, although a single major locus with additional multifactorial effects was more strongly supported. The results from the current study suggest that levels of UAE are determined by a mixture of genes with large and small effects as well as other measured covariates, such as diabetes. PMID- 10866061 TI - Effect of hyperglycemia and hyperlipidemia on atherosclerosis in LDL receptor deficient mice: establishment of a combined model and association with heat shock protein 65 immunity. AB - Diabetes and atherosclerosis have been proposed to be influenced by immune and autoimmune mechanisms. A common incriminated antigen in both disorders is the heat shock protein (HSP)-60/65. In the current study, we established a model combining hyperglycemia with hyperlipidemia in LDL receptor-deficient (LDL-RD) mice and assessed its possible influences on lipid profile, HSP60/65, and atherogenesis. LDL-RD mice were injected either with streptozotocin to induce hyperglycemia or with citrate buffer (control). When hyperglycemia was induced, both study groups were challenged with a high-fat (Western) diet for 6 weeks. Plasma fasting glucose, lipid profile, and antibody levels to HSP65 and oxidized LDL were assessed. At death, the spleens from both groups were evaluated for their proliferative response to HSP65 and the consequent cytokine production. The extent of atherosclerosis was assessed at the aortic sinus. Plasma glucose, cholesterol, and triglyceride levels were elevated in mice injected with streptozotocin compared with control mice. Atherosclerotic lesions were significantly larger in the streptozotocin-injected hyperglycemic LDL-RD mice (132 +/- 23 x 10(5) microm2) in comparison to their normoglycemic litter-mates (20 +/- 6.6 x 10(5) microm2; P < 0.0001). Both humoral and cellular immune response to HSP65 was more pronounced in streptozotocin-injected mice. When challenged with HSP65 in vitro, splenocytes from streptozotocin-injected mice favored the production of the T-helper (TH)-1 cytokine gamma-interferon. In conclusion, we have established a mouse model that combines hyperglycemia with diet-induced hyperlipidemia in LDL-RD mice and studied its effect on atherosclerosis progression. The accelerated atherosclerotic process is associated with heightened immune response to HSP65 and a shift to a TH1 cytokine profile. PMID- 10866062 TI - Evaluation of the secondary consolidation of columns for liquid chromatography by ultrasonic irradiation. AB - The consolidation of packed analytical chromatography columns was carried out under ultrasonic irradiation. Columns were first packed using a conventional high pressure downward slurry method. Then, they were subjected to further bed consolidation in the presence of ultrasonic vibration. This process of further bed consolidation is referred to as secondary consolidation. Secondary consolidation was observed to occur more readily in solvents of low viscosity and at low flow-rates (low pressures). Column efficiency was not observed to be a factor affecting the process of secondary consolidation of the packed bed. PMID- 10866063 TI - Large degree of racemization observed in the amide bond forming reaction on silica gel. AB - A systematic study of racemization from the coupling of DNB-L-Leu and 3 aminopropyl silica gel with several amide coupling reagents was investigated. Significant amounts of racemization were observed from all except one coupling reagent. In comparison, similar reactions completed in homogeneous solution can be accomplished with much lower racemization and in much higher yields. PMID- 10866064 TI - Determination of basic nitrogen-containing polynuclear aromatic hydrocarbons formed during thermal degradation of polymers by high-performance liquid chromatography-fluorescence detection. AB - A method for the simultaneous determination of 22 nitrogen-containing polynuclear aromatic hydrocarbons (PAHs) (15 azaarenes and seven amino-PAHs) in the gaseous products of the thermal degradation of polymers is described. After desorption and clean-up using cation-exchange chromatography (PRS cartridge) the samples were analyzed by HPLC-FD. The validation was carried out with real nylon 6 combustion samples. The recoveries of the compounds are in the range of 89-99% for the extraction and 91-100% for the clean up procedure. The detection limits ranged from 7 to 160 pg. The high recovery values, due to the quick and efficient clean-up procedure, resulted in low relative standard deviations (with the exception of acridine and 2-aminoanthracene <5%). The applicability of the method is also investigated for the determination of the N-PAHs in a pyrolysis sample. PMID- 10866065 TI - Simultaneous separation and quantitation of the major bovine whey proteins including proteose peptone and caseinomacropeptide by reversed-phase high performance liquid chromatography on polystyrene-divinylbenzene. AB - A precise, sensitive and reliable RP-HPLC method was developed to enable not only unequivocal determination of alpha-lactalbumin and beta-lactoglobulin in bovine whey samples, but also simultaneous measurement of proteose peptone, caseinomacropeptide, bovine serum albumin and immunoglobulin G. The optimised method on the Resource RPC column allowed separation of the proteins in 30 min and could be applied to the analysis of soluble proteins in a variety of commercial and laboratory whey products. Furthermore, some qualitative information on protein heterogeneity and quality could be derived from the RP HPLC analyses with additional data available from on-line electrospray mass spectrometry. Within- and between-day repeatability over a wide range of concentrations was excellent (RSD< or =5%) for all proteins except immunoglobulin G and bovine serum albumin where RSD was 7-10%. Analysis of grouped data from whey protein concentrate and whey protein isolate samples gave a limit of detection of < or =0.3% powder mass and a limit of quantitation of < or =1.0% powder mass for all proteins except immunoglobulin G. Limits of detection and quantitation were 0.6% and 2.0%, respectively, for this protein. Quantitative data obtained by the RP-HPLC method compared very favourably with data obtained by alternative methods of whey protein analysis. PMID- 10866066 TI - Lysozyme refolding with immobilized GroEL column chromatography. AB - A refolding chromatography with immobilized molecular chaperonin GroEL was studied for the reactivation of denatured-reduced lysozyme. The effect of denaturant concentration (guanidine hydrochloride, 0.1-1.5 M) in the elution buffer, the elution flow-rate, and the loading concentration and volume of the substrate protein on the reactivation yield was studied. All the operating parameters showed minor effects on the recovery yield of lysozyme mass, which remained at 90-100%, but exhibited relatively notable influences on the specific activity of the recovered lysozyme. For example, there existed an optimum denaturant concentration of about 1 M at which the highest yield of specific activity (up to 97%) was obtained. Using the immobilized GroEL column, 3 ml of the lysozyme (1 mg/ml) per batch could be refolded at an overall yield of 81%, which corresponded to a refolding productivity of 54 mg per 1 gel per h. At comparable reactivation yields (over 80%), this value of productivity was over four-times larger as that of the size-exclusion refolding chromatography reported previously (12 mg per 1 gel per h), indicating the advantage of the present system for producing a high throughput in protein refolding operations. PMID- 10866067 TI - Evaluation of time-of-flight mass spectrometric detection for fast gas chromatography. AB - Separations below 1 s of a mixture of organic compounds ranging from C5 to C8 have been performed to investigate the performance of a time-of-flight mass spectrometer in fast gas chromatography. The gaseous samples were focussed on a cold trap, and then injected after thermal desorption to obtain the required narrow input band-widths. Also, to obtain a very fast separation, a short narrow bore column was used, operated at above-optimum inlet pressures. With this system, it was possible to identify ten compounds within 500 ms, showing peak widths (2.354sigma) as narrow as 12 ms. The spectral acquisition rate used for these analyses was 500 Hz. The quality of the recorded spectra and the comparison with library spectra was very high. Deconvolution algorithms offer the possibility of identifying overlapping peaks. It is shown that the spectral scan speed of the time-of-flight mass spectrometer is high enough for very fast separations. PMID- 10866068 TI - Characterization of a commercial gas chromatography-flame ionization detection system for the in situ determination of C5-C10 hydrocarbons in ambient air. AB - A commercial automated gas chromatograph with preconcentration on solid adsorbents (AirmoVoc HC1010) was coupled with a mass spectrometer in parallel with the flame ionization detection (FID) system and characterized for its suitability for quasi continuous measurements of atmospheric hydrocarbons (HCs) with a time resolution of 20 min. Of the 50 identified HCs in the range C5-C10, 15 elute in isolated peaks and 20 in groups of two or more HCs. The remaining 15 HCs suffer from coelution by oxygenated and halogenated compounds. Procedures to minimize and quantify the blanks and the memory from the adsorbents are described. Calibration was based on a custom-made diffusion source. The accuracy of this calibration (+/-10%, 2sigma) was verified by analysis of a certified 70 component standard (average deviation: -4.3+/-2%). During a field experiment in Summer 1998, the HC1010 system was compared with a custom-made GC system with cryogenic preconcentration and much better separation properties but lower time resolution. In ambient air, good agreement (2sigma deviation <14% or 10 ppt) was found for HCs and groups of HCs that are free from coelution with oxygenated compounds, whereas large discrepancies (in some cases more than a factor of three) were found for those HCs that coelute with oxygenated compounds, as identified by MS. Analysis of the mass spectra from those peaks via specific target ions showed much better agreement with the FID system of the reference GC within 25%. PMID- 10866069 TI - Gas chromatography-mass spectrometry-Fourier transform infrared spectrometry and high-performance liquid chromatographic characterization of mononitro- and dinitro-isomers from the nitration of N,N-dimethyldiphenylacetamide. AB - Gas chromatographic retention data, Fourier transform infrared and mass spectrometric (electron impact, electron attachment and methane chemical ionization) profiles are reported for the products of mono- and dinitration of N,N-dimethyldiphenylacetamide. Differentiations of analytical importance among isomers could be gathered by the study which led to their complete identification. PMID- 10866070 TI - Determination of dissociation constants of cytokinins by capillary zone electrophoresis. AB - A method for the pKa determination, based on mobility data measured by capillary zone electrophoresis, was applied to cytokinins and their analogs. The combination of charged mobility standards with an uncharged electroosmosis marker, injected in the uncoated capillary simultaneously with the measured substances, allows one to minimize the number of runs, reduce their duration and, in addition, to inform on the run-to-run stability of electroosmosis and on contingent side-effects. pKa values of investigated cytokinins and their analogs ranged from 2.8 to 4.0 at 25 degrees C in the phosphate and acetate buffers of ionic strength 0.015 M. Standard deviations of the constants, obtained by the non linear fitting of equations for the pKa calculation, were 3-5-times lower than standard deviations from the linear fitting or from the point-to-point calculation utilizing the Hendersson-Haselbalch equation. The equation of Boltzman sigmoid offers two checks on reliability of effective mobilities that serve as the raw data in the pKa calculation. PMID- 10866071 TI - Determination of cow's milk in non-bovine and mixed cheeses by capillary electrophoresis of whey proteins in acidic isoelectric buffers. AB - An improved method for the determination of cow's milk in non-bovine cheese is reported: electrophoresis of whey proteins in acidic, isoelectric buffers. Two background electrolytes (BGEs) have been tested: (i) 50 mM iminodiacetic acid (pH=isoelectric point=2.30 at 25 degrees C), 0.5% hydroxyethylcellulose, 0.1% Tween 20 and 6 M urea (apparent pH 3.1), E=300 V/cm, for the separation of alpha lactalbumins (alpha-LAs); (ii) a BGE with the same composition, but supplemented with 10% Tween 20, E=450 V/cm, for the fractionation of beta-lactoglobulins (beta LGs). Surfactants have a discriminating effect on the retention behaviour of the bovine alpha-LA and beta-LG proteins, owing to the different strength of the protein-surfactant association complexes, and are needed for separating these two proteins from small peaks in the electropherograms generated by degradation of casein during cheese ripening. Novel equations are given for deriving the ratio of the area (or height) of bovine alpha-LA, or beta-LG, to the area (or height) of ovine or caprine alpha-LA or beta-LG (such ratios being typically used to determine the percentage of cow's milk in dairy products), since previous equations had marked drawbacks, such as non-linearity of the plots with increasing slopes at high cow's milk percentages, and too broad confidence limits at high cow's milk contents, where the peak area (or height) ratio tends asymptotically to infinite. With the novel procedures reported, contents of cow's milk as low as 1% can be quantified in goat's and ewe's cheeses. The present protocols give lower detection limits, are cheaper and more rapid than any other methodology reported in the literature, and can be easily applied to the routine quality control of binary and ternary cheeses. PMID- 10866072 TI - Neurologic complications of cerebral angiography. A retrospective study of complication rate and patient risk factors. AB - PURPOSE: To evaluate the neurologic complication rate and individual patient risk factors in cerebral angiographies using the digital subtraction angiography (DSA) technique and non-ionic contrast media in a department with many radiologists in training. MATERIAL AND METHODS: A retrospective study of 483 cerebral angiographic examinations in 454 patients was carried out. The following parameters were registered: sex and age of the patient, indication for the angiography, cerebral CT diagnosis, laboratory data, type of anesthesia, type of angiographic procedure, level of training of the angiographer, number of participating angiographers, type of catheters, number of vessels catheterized, number of exposures, use of compression series, total amount of contrast media, diagnosis of the angiogram, complications and duration of complications. RESULTS: The frequency of all neurologic complications was 2.3%, the frequency of persistent neurologic deficits was 0.4%. Non-neurologic complications were observed in 14.7% of the examinations. Of all the parameters studied, the only factor that significantly increased neurologic risk was a normal angiogram, a finding we are inclined to ascribe to chance. Performance of a compression series showed a trend towards increasing the neurologic risk. CONCLUSION: This study showed a complication rate of persistent neurologic deficits of 0.4% which is in accordance with other recent reports. A compression series should not be performed routinely, but only on special indication. This study confirms the low risk of cerebral angiography when performed in a neuroradiological department using the DSA technique and non-ionic contrast media. PMID- 10866073 TI - Enhancement pattern of normal extraocular muscles in dynamic contrast-enhanced MR imaging with fat suppression. AB - PURPOSE: To evaluate the internal structure of normal extraocular muscles on fat suppressed dynamic contrast-enhanced MR imaging. MATERIAL AND METHODS: Ten subjects were examined using fat-suppressed dynamic contrast-enhanced MR imaging. We evaluated the enhancement pattern (C-shaped or ring-like) of extraocular muscles and quantified the maximum ratios of enhancement (Rmax) and maximum ratios of signal increase (Vmax). We also quantified Rmax and Vmax in the central and peripheral portions of medial rectus muscles. RESULTS: In the early phase of dynamic contrast-enhanced MR imaging, a C-shaped or ring-like pattern was observed in 100% of inferior rectus, 95% of medial rectus, 55% of superior rectus, 20% of lateral rectus, and 15% of superior oblique muscles. Overall mean Rmax and Vmax values showed statistically significant differences to the temporal muscles. For the peripheral portion of medial rectus muscles, mean Rmax and Vmax values were greater than for the central portion. CONCLUSION: Using fat suppressed dynamic contrast-enhanced MR imaging, the C-shape or ring-like internal structure of the extraocular muscles could be visualized, and were considered to reflect their structure of orbital and global layers. Potential usefulness of the fat-suppressed dynamic contrast-enhanced MR imaging for detecting pathological status is suggested. PMID- 10866074 TI - Ultrasound demonstration of mammographically detected microcalcifications. AB - PURPOSE: To evaluate the capabilities of breast ultrasound (US) for identifying microcalcifications in benign breast changes, in situ carcinomas, and small nonpalpable invasive carcinomas. MATERIAL AND METHODS: Forty-six consecutive patients with 49 clustered microcalcifications detected by mammography were included in this prospective study. Patients with palpable breast lesions were excluded. Breast US was performed with knowledge of mammographic findings for presence and visibility of microcalcifications, and for parenchymal structure abnormalities. Mammographic and US findings were compared with histology. RESULTS: Nine ductal in situ carcinomas, 2 lobular in situ carcinomas, 11 invasive carcinomas and 27 benign lesions were confirmed by histology. For all lesions, US achieved a sensitivity of 75% in the detection of microcalcifications. The detection rate for microcalcification in invasive and in situ carcinomas was 100%. In 11 cases, no microcalcifications were visible on US; they all proved to be benign on histology. CONCLUSION: Microcalcifications in malignant lesions are reliably recognized by US. They are, however, difficult to detect in fibrocystic breast changes. PMID- 10866075 TI - The potential influence of fine-needle aspiration on MR imaging of the breast. AB - PURPOSE: The aim of this study was to determine whether fine-needle aspiration biopsy (FNAB) of breast lesions causes structural changes or changes in contrast enhancement, that could impair the evaluation of these lesions at MR investigation of the breast. MATERIAL AND METHODS: Fifteen patients with 17 lesions were examined with MR imaging of the breast both before and after the FNAB with a mean interval of 7.1 days. At both examinations, signal intensities were measured pre- and post-contrast enhancement in the lesions as well as in surrounding breast parenchyma and muscle. Imaging with contrast enhancement was performed semidynamically with two images obtained in rapid sequence (acquisition time 6.23 min) immediately after bolus injection of gadopentetate dimeglumine, 0.1 mmol/kg b.w., and the contrast enhancement in both images was calculated. RESULTS: The diagnostic value of MR images was unimpaired by FNAB, and no statistical difference between contrast enhancement in corresponding images obtained before and after FNAB was found, either within the lesions or in the breast parenchyma or muscle. CONCLUSION: FNAB of the breast can be performed without impairing the diagnostic outcome of MR investigation. PMID- 10866076 TI - Intracystic papillary carcinoma in the male breast. A case report. AB - Male breast cancer is a rare disease with an incidence between 0.5% and 2.4% of that in women. We report a case of intracystic papillary carcinoma of the breast in a 75-year-old Japanese man. The macroscopic features of the carcinoma could be accurately demonstrated by pneumocystography and ultrasonography preoperatively. PMID- 10866077 TI - Pulmonary disease in liver transplant recipients. Spectrum of CT features. AB - PURPOSE: To determine the features of pulmonary disease in liver transplant recipients by CT. MATERIAL AND METHODS: Of 792 patients, 102 were referred to thoracic CT 3-2093 days after the transplantation procedure (median 107 days). All CT studies were retrospectively analyzed and correlated with clinical, microbiological, serological and histopathological findings. RESULTS: Eighty eight of 102 patients (86%) had an abnormal CT. In 25 patients (25%), an elevated right hemidiaphragm, basal atelectasis and small effusions were the only abnormalities. Fourty-one patients (40%) displayed an infiltrate and 13 (13%) a mass lesion. Evidence of cytomegalovirus (CMV) infection was found in 20 patients. CMV pneumonia was suggested by an interstitial pattern of pneumonia on CT (n=13). Pneumocystis carinii pneumonia was highlighted by peribronchovascular infiltrates (n=5/8), bacterial pneumonia (n=24) including legionellosis (n= 13) by bilateral effusions (n=14) and lobar consolidation (n= 13). In 7/41 patients (17%) with both clinically apparent pulmonary disease and CT signs of pneumonia, no pathogen could be detected. Neoplastic disease was mostly due to tumor recurrence (n=6). CONCLUSION: Thoracic CT of liver transplant recipients aids in detecting and classifying both infectious and neoplastic complications. PMID- 10866078 TI - High-resolution CT in renal transplant patients with suspected pulmonary infections. AB - OBJECTIVE: This prospective study was carried out to assess the usefulness of high-resolution CT (HRCT) of the chest in immunocompromised renal transplant patients with suspected pulmonary infections. MATERIAL AND METHODS: Twenty-one consecutive renal transplant patients with clinically suspected pulmonary infections underwent chest radiography, HRCT and other tests including bronchoalveolar lavage (BAL). HRCT was performed using a high spatial frequency algorithm with 2-mm-thick sections at 10-mm intervals from apices to domes of the diaphragm. The findings on chest radiography and HRCT were interpreted by two thoracic radiologists and the usefulness of HRCT was evaluated. The images were interpreted independently by two radiologists, who were blinded to the findings of other imaging modalities and the final diagnosis. Any differences regarding the imaging findings were resolved through consensus. RESULTS: Final diagnosis was obtained in 17 patients, and no cause for symptomatology was established in 4 patients. The spectrum of infections included pulmonary tuberculosis (TB) in 11 patients, cytomegalovirus pneumonia (CMV) in 2 patients, cryptococcal and streptococcal pneumonia, pulmonary aspergillosis and esophageal candidiasis in 1 patient each. Compared to chest radiography, HRCT revealed additional findings in 11 patients. HRCT findings were suggestive of underlying infection in 11 patients. The final diagnosis coincided with HRCT diagnosis in all but 1 patient. HRCT findings were non-specific in 3 patients and normal in 7. The findings were concordant in 19 cases. The results were not in agreement in only 2 cases. CONCLUSION: HRCT can provide useful information and suggest the diagnosis in a significant proportion of renal transplant patients with pulmonary infection. PMID- 10866079 TI - CT lung densitometry in assessing intralobular air content. An experimental and clinical study. AB - PURPOSE: To introduce a parameter called "intralobular air content percentage" to replace the CT number of the lung and to establish a proper protocol for its assessment. MATERIAL AND METHODS: We calibrated the HU (Hounsfield unit) scale for low densities with foam and evaluated the influence of certain acquisition and reconstruction parameters on the accuracy of CT densitometry of the lungs. The reproducibility of the results obtained in human experiments and the intralobular air content percentage of normal and diseased lung tissue were assessed. RESULTS: Air content could be reliably derived from the calibrated CT number of an area within a secondary lobulus. The mean intralobular air content of normal lungs varied from 77.8% to 88.0% in full inspiration. Helical or axial recording with a 10-mm slice thickness, a standard or soft algorithm and high tube currents and voltage settings, was suitable for the measurements. CONCLUSION: Before absolute lung density measurements (as a HU number or an air content percentage), the CT equipment has to be calibrated for low densities. The intralobular air content percentages of cooperative patients were reliably reproducible. PMID- 10866080 TI - Outcome after catheter-directed thrombolysis of occluded prosthetic femoropopliteal bypasses. A prospective study. AB - PURPOSE: To evaluate the outcome after catheter-directed thrombolysis of occluded femoropopliteal prosthetic bypasses with the distal anastomosis above the knee. MATERIAL AND METHODS: Twenty-one patients were included in this prospective study. End-hole catheters, a bolus dose and continuous infusion of recombinant tissue-plasminogen activator (rt-PA) were used, with a median total dose of 10 mg (range 7-20 mg). RESULTS: With an intra-thrombotic position of the catheter, total or subtotal lysis was obtained in 19 of 21 patients (90%). No serious complications occurred. In 9 patients, the stenoses were successfully treated with balloon angioplasty (PTA, n=5), local thrombectomy/extension of bypass (n=3), or with a new bypass (n=1). After a median observation time of 18 months (6-24), 5 patients had open bypass. Re-occlusion occurred in all (6/6) bypasses in which no flow-limiting lesion was discovered, in all (4/4) bypasses treated twice with thrombolysis, as well as in all bypasses in which stenoses had not been adequately treated (3/3). One bypass re-occluded immediately due to poor runoff. CONCLUSION: In the present study, 19/21 infra-inguinal prosthetic bypasses were successfully treated with catheter-directed thrombolysis. However, re-occlusion often took place, especially in bypasses without flow-limiting lesions. If re-occlusion occurs in a bypass in which no stenoses were revealed during the primary thrombolysis procedure, a second catheter-directed thrombolytic treatment does not seem to be warranted. Our results confirm that treatment of flow-limiting lesions is a prerequisite for maintaining patency. PMID- 10866082 TI - Multishot echo-planar imaging with gadopentetate dimeglumine. Preliminary study of efficacy for detection of hypovascular metastatic liver tumors. AB - OBJECTIVE: To evaluate the usefulness of sequential T2-weighted spin-echo type multishot echo-planar (T2-EP) imaging with gadopentetate dimeglumine for the detection of hypovascular metastatic liver tumors. MATERIAL AND METHODS: Fifteen consecutive patients with 56 proven hypovascular metastatic liver tumors were included in the study. Three observers blindly and independently read the whole liver images obtained with T2-weighted spin-echo, T2-weighted single-shot fast spin-echo, T1-weighted fast multiplanar spoiled GRASS and T2-EP images obtained before and 25, 60, 90 and 120 s after injection of 0.2 mmol/kg b.w. of gadopentetate dimeglumine. The diagnostic accuracy was estimated by calculating the area under the observer-specific binomial receiver operating characteristics curves (Az). RESULTS: T2-EP images obtained 60 s after contrast injection showed significantly higher contrast-to-noise (C/N) ratios than the other imaging techniques. A combination of all phases of the T2-EP images produced the highest sensitivity and specificity. In terms of the Az value, the diagnostic accuracy for tumor detection achieved with a combination of all phases of the T2-EP images was significantly higher than that with T1-SPGR and T2-SSFSE images (p<0.01). The Az values of the T2-EP images (Az=0.975) were higher than those of T2-CSE images (Az=0.948), but the difference was not significant. CONCLUSION: Our preliminary study revealed that sequential imaging with enhanced T2-EP images was useful for the detection of hypovascular metastatic liver tumors because of its superior C/N ratio and sensitivity. PMID- 10866081 TI - Dynamic MR imaging of liver metastases with Gd-EOB-DTPA. AB - PURPOSE: To assess liver and lesion enhancements by dynamic MR imaging after bolus injection of the hepatobiliary contrast agent gadolinium ethoxybenzyl diethylenetriamine-pentaacetic acid (Gd-EOB-DTPA) in patients with liver metastases and to compare the effect of different doses. MATERIAL AND METHODS: A randomized double-blinded trial with doses of 12.5, 25 and 50 micromol/kg Gd-EOB DTPA was performed in 35 patients with liver metastases. Liver enhancement, tumor enhancement and liver lesion contrast-to-noise (C/N) ratios were calculated from breath-hold gradient echo images (100/5/80 degrees) recorded precontrast and at different times up to 10 min postcontrast. RESULTS: Normal liver showed a characteristic enhancement pattern, with a rapid enhancement in the first 45 s postcontrast and a slight but significant further increase up to 600 s. The initial enhancement in the lesions was also pronounced, but the enhancement was slightly decreased after 240 s postcontrast. At dose levels of 12.5 and 25 micromol/kg Gd-EOB-DTPA, C/N ratios significantly increased compared to baseline from 90 to 600 s. Postcontrast C/N-values obtained using 50 micromol/kg Gd-EOB DTPA were not significantly increased, except for the examinations 480 s postcontrast. CONCLUSION: In liver metastases, C/N ratios obtained with doses of 12.5 and 25 micromol/kg Gd-EOB-DTPA were slightly superior to 50 micromol/kg Gd EOB-DTPA. This finding is probably due to a more pronounced extracellular effect of the contrast medium at higher doses. PMID- 10866083 TI - MR cholangiopancreaticography and endoscopic retrograde cholangiopancreaticography in patients with suspected common bile duct stones. AB - PURPOSE: To prospectively compare MR cholangiopancreaticography (MRCP) vs. endoscopic retrograde pancreaticography (ERCP) in patients with suspected common bile duct (CBD) stone disease. MATERIAL AND METHODS: Fifty consecutive patients with suspected CBD disease underwent MRCP and then ERCP within 12 h of each other. The result of the MRCP was blinded to the reader of the ERCP. The MRCP was done using a superconducting 1.0 T unit with a heavily T2-weighted breath-hold technique. The ERCP was done in the fluoroscopy suite by one of the clinicians and was evaluated by one of the radiologists who had not read the MRCP examinations. RESULTS: There were 28 true-positives, 17 true-negatives, 1 false positive, and 4 false-negatives. The sensitivity was 87.5% and the specificity 94.4%, respectively. The positive predictive value was 96.6% and the negative predictive value was 81.1%. CONCLUSION: MRCP was shown to be good enough to replace ERCP as a diagnostic method in patients with suspected CBD disease. MRCP is now our modality of choice after ultrasound in the diagnostic evaluation of these patients. PMID- 10866084 TI - MR cholangiography in massive hepatobiliary ascariasis. A case report. AB - The ultrasonographic and MR cholangiographic findings in a patient with massive hepatobiliary ascariasis are described. PMID- 10866085 TI - Esophageal rings and strictures. Manometric characteristics in patients with food impaction. AB - PURPOSE: The purpose was to investigate the manometric characteristics in patients with lower esophageal rings or strictures with special reference to food impaction. MATERIAL AND METHODS: The material comprised 344 patients (158 female and 186 male). Lower esophageal rings or strictures were diagnosed radiologically by the full column technique. Manometry was performed with triple lumen catheters connected to a hydraulic capillary system and external transducers. RESULTS: Forty patients had rings, and 21 patients strictures. The reference group comprised 283 patients. Dysphagia was seen most frequently in patients with rings. Food impaction was seen only in patients with rings or strictures, whereas chest pain and heartburn appeared with the same incidence in all 3 groups. Non specific motor disorders were seen most frequently in patients with strictures, but as delayed esophageal emptying with no influence on the tendency to food impaction. Only ring diameter, but not stricture diameter, was of any significance with a higher incidence of food impaction in patients with narrow rings. CONCLUSION: Radiology rather than manometry should be the first diagnostic step in patients with benign dysphagia suffering from food impaction. PMID- 10866086 TI - CT in the evaluation of complicated autosomal dominant polycystic kidney disease. AB - PURPOSE: We retrospectively reviewed the CT findings in 24 cases of autosomal dominant polycystic kidney disease (ADPKD) to assess the role of CT in the diagnostic work-up of patients with complicated ADPKD. MATERIAL AND METHODS: Twenty-four patients with ADPKD underwent unenhanced and contrast-enhanced CT for flank pain, haematuria, or fever. The images were retrospectively reviewed for presence of complicated cysts, their morphological features and associated findings in the perinephric space/retroperitoneum. RESULTS: Cyst haemorrhage was present in all patients, seen as high-density cysts, which were mostly bilateral. Most of these cysts had sharply outlined contours, sharp interfaces with adjacent renal parenchyma, imperceptible walls, and homogeneous density, and did not enhance following i.v. contrast administration. However, a few haemorrhagic cysts (9 cysts in 6 patients) showed inhomogeneous density (n=7), dependent layering of high-density blood leading to fluid-fluid level (n=2), and contour irregularity (n=3). CT revealed presence of cyst infection in 6 cases; the involved cysts were larger (average size 4.2 cm) than adjacent cysts, had only a mildly increased or near water density, and showed wall thickening and enhancement. Other findings included air within the infected cyst (n=1), thickening and enhancement of peri- and paranephric fasciae (n=5), and abscesses in the posterior paranephric space and adjoining psoas muscle (n=2). In 2 other patients, although CT suggested cyst infection because of presence of wall enhancement, diagnostic needle aspiration revealed only sterile haemorrhagic fluid. In 1 case, CT revealed a soft tissue density enhancing mass in one of the cysts; this proved to be a renal cell carcinoma by fine-needle biopsy. Calculi were observed in 7 patients, and cyst wall calcification in 11 cases. CONCLUSION: A combination of unenhanced and contrast-enhanced CT allows correct diagnosis and differentiation amongst the various complications affecting patients with ADPKD. However, in a small subgroup of patients, it may not be possible to differentiate between haemorrhage and infection; such cases require diagnostic needle aspiration for diagnosis. PMID- 10866087 TI - From donor to recipient. Doppler US, power US and scintigraphy of kidney perfusion before and after transplantation. AB - PURPOSE: To follow kidneys from the donor to the recipient by assessing whether perfusion changes occur by using duplex Doppler US, power Doppler US and scintigraphy. MATERIAL AND METHODS: The prospective study included 12 donors and their corresponding 12 recipients. For each donor, both donor kidneys were evaluated by duplex Doppler US, power Doppler US and scintigraphy 1 day before surgery. The same procedure was carried out on the renal allografts at days 1, 3, 5 and months 1 and 3 post-transplantation. Power Doppler findings were classified according to a grading system of 1 to 4. Resistive indices (RIs) were determined based on interlobar and segmental arterial flow. Peak systolic velocity and RIs of the main renal artery were also measured. A perfusion parameter named the peak to-plateau ratio was calculated. Statistical analysis was performed using the paired-samples t-test. RESULTS: Intrarenal RI elevation and decreased renal artery peak systolic velocity was observed in normally functioning recipient kidneys. CONCLUSION: Duplex Doppler sonography demonstrated that transplanted kidneys had an increase in intrarenal vascular resistance at 1 month and a decrease in renal artery peak systolic velocity at 3 months post-transplantation. Scintigraphy and power Doppler US did not reveal any statistically significant perfusion change in normally functioning kidneys from donor to recipient. PMID- 10866088 TI - Transrectal US and endorectal MR imaging in partial and complete obstruction of the seminal duct system. A comparative study. AB - PURPOSE: To evaluate transrectal ultrasonography (US) and MR imaging findings of infertile patients with suspected complete or partial obstruction of the seminal duct system. MATERIAL AND METHODS: Two hundred and eighteen infertile patients with low ejaculate volume were evaluated by transrectal US. Endorectal MR imaging was performed on 62/218 patients. Prostatic cysts, ejaculatory duct (ED) dilatation (>2 mm in width), ED calculi or calcifications, seminal vesicle (SV) dilatation (a.p. diameter >15 mm), SV hypo/agenesis (a.p. diameter <7 mm), SV cysts (>5 mm), vasal agenesis and chronic prostatitis (coarse calcifications, heterogeneity in prostate) were considered significant findings for obstruction of the seminal duct system. RESULTS: Pathologic findings were detected in 75% and 61% of patients with azoospermia on transrectal US and MR imaging, respectively. Transrectal US and MR imaging did not reveal any pathologies in 64.7% and 59.1% of patients with nonazoospermia, respectively. The incidences of hypoplastic/atrophic SV (12/48 vs. 5/170), SV agenesis (6/48 vs. 1/170), vasal agenesis (5/48 vs. 1/170) were significantly higher in the azoospermic subgroup (p<0.002). CONCLUSION: US is a good method for initial evaluation of these patients especially in complete obstruction. Endorectal MR imaging should be reserved for selected patients in whom results of transrectal US are not conclusive. PMID- 10866089 TI - US versus conventional radiography in the diagnosis of sternal fractures. AB - PURPOSE: To investigate the value of ultrasonography (US) in the diagnosis of sternal fractures. MATERIAL AND METHODS: Twenty-three patients (mean age 35.4 years) with a clinical suspicion of sternal fracture after blunt chest trauma were retrospectively reviewed. At admission, a.p. and lateral chest radiographies and sternal US were obtained. Sternal fractures were classified as nondisplaced or displaced. US and conventional radiographic findings were compared. RESULTS: In 3/23 (13.0%) of the patients, no fracture was found by radiography or by US. Both radiography and US demonstrated sternal fractures in 16/23 (69.6%) of the patients. Sternal fractures were detected only by US while the conventional radiography was negative in 2/23 (8.7%) cases. Also in 2/23 (8.7%) of the patients with US positive for fracture, radiographies were suspicious. In 2 patients, the degree of fracture displacement on US was lesser than that found by radiography. CONCLUSION: US was better than lateral radiography to diagnose sternal fractures; however, conventional radiography remains the standard means of demonstrating grade of displacement. PMID- 10866090 TI - MR imaging after sports-induced hip dislocations. Report of three cases. AB - The MR findings in 3 patients who had sustained a posterior dislocation of the hip during sports activities are reported. PMID- 10866091 TI - MR imaging in suspected acute trauma of wrist bones. PMID- 10866092 TI - European perspective of clinical trials in systemic lupus erythematosus. PMID- 10866093 TI - Cardiac abnormalities in SLE: pancarditis. AB - Many patients with systemic lupus erythematosus (SLE) develop cardiac manifestations during the course of their disease. Pericarditis is most commonly seen, with a reported prevalence of 60%. Myocardial involvement is present in only a minority of patients. In recent years, due to better noninvasive diagnostic techniques, valvular abnormalities can be demonstrated in an increasing number of patients. Depending on the technique used, valvulopathy can be demonstrated in up to 77% of SLE patients. Although most of the valvular lesions will be present without any symptoms, valve incompetence can result in congestive heart failure. Valvular lesions are associated with IgG anticardiolipin antibodies (aCL) and disease duration. We present a patient with SLE and secondary antiphospholipid syndrome (APS) who developed acute congestive heart failure due to pancarditis. Endocarditis, together with left ventricular dysfunction and pericardial effusion, were present. The endocarditis caused hemodynamically significant mitral valve insufficiency due to thickening of the mitral cusps. Just two weeks prior to the occurrence of congestive heart failure echocardiography had been normal. Treatment with high dose corticosteroids resulted in a gradual, almost complete recovery. Literature concerning cardiac manifestations in lupus is reviewed. PMID- 10866094 TI - Cyclosporine for lupus membranous nephritis: experience with ten patients and review of the literature. AB - OBJECTIVES: The treatment of lupus membranous nephritis (LMN), a lupus subset that carries a high morbidity, is unsatisfactory. We report our experience in treating LMN with the immunosuppressive drug cyclosporine (CYS). METHODS: We treated 10 patients with systemic lupus erythematosus fulfilling ACR criteria with CYS for at least 12 months and followed renal function, serologic activity and SLEDAI scores. PATIENT CHARACTERISTICS: 8 females, 2 males, 50% Caucasian, mean age 37.3 y (range 22-48), disease duration 108.7 months (range 16-216), nephritis duration 35.5 months (range 12-59), date of biopsy to date of starting treatment 10.7 months (range 0-90). The patients were started on CYS with a mean dose of 3.8 mg/kg (range 2.2-6) and followed for a mean duration of 24.8 months (range 12-59). A Medline search identified all patients with lupus who were given CYS or had LMN in articles from 1966-1999. RESULTS: Proteinuria improved from a baseline mean of 5,588mg/24h (range 2,712-11,055) to 1,404 mg/24 h (range < 150 2,652). Serum albumin increased from a baseline mean of 2.8 g/100 ml (range 1.31 3.8) to a mean of 3.9 g/100 ml (range 3-4.5) at last follow-up. There was no significant change in lupus activity as measured by SLEDAI. Nephrotoxicity was common as evidenced by an increase in serum creatinine but it returned to baseline with adjustment of the dose of CYS (20% decrease in the dose of CYS for a 20% increase in serum creatinine). More antihypertensive medications were required to control the blood pressure in these ten patients at the end of the study compared to the onset (total number= 13 versus 6). CONCLUSION: Proteinuria and serum albumin improved in all patients on CYS. A literature review is consistent with this. Controlled studies of the use of CYS for membranous lupus nephritis would be useful. PMID- 10866095 TI - Profile of sex hormones in male patients with systemic lupus erythematosus. AB - OBJECTIVE: To study the profile of sex hormones in male patients with systemic lupus erythematosus (SLE). METHOD: Serum prolactin (PRL), testosterone (T), estradiol (E2), follicle-stimulating hormone (FSII) and luteinizing hormone (LH) levels were obtained from 35 males with SLE and compared with 33 age-matched normal controls. RESULTS: No significant differences in serum T, E2, PRL levels and E2/T ratio were observed between male SLE patients and controls. However, patients with SLE had significantly higher levels of gonadotrophins (FSH, LH). Five (14%) SLE patients, but none of the controls, had both low testosterone and elevated LH. Hypoandrogenic male SLE patients did not have overt features of hypogonadism but had a higher prevalence of central nervous system disease and scrositis than those with normal androgen levels. Discase flares, on the other hand, were not significantly more frequent in these patients. Although PRL or T levels per se did not correlate with disease activity in our patients, the ratio of PRL to T showed a significant correlation with SLEDAI scores (p = 0.42. P = 0.01). CONCLUSIONS: Hypoandrogenism is present in some male patients with SLE and may be relevant in disease pathogenesis. However, whether these hormonal abnormalities are intrinsic to SLE or the consequence of any non-specific chronic disorders cannot be distinguished from the current data. Further studies involving a larger number of subjects and inclusion of other disease controls are needed. PMID- 10866096 TI - The family history of patients with primary or secondary antiphospholipid syndrome (APS). AB - OBJECTIVE: To evaluate familial history for evidence of antiphospholipid syndrome (APS) and autoimmune disease in rheumatology department patients with primary or secondary APS. METHODS: We retrospectively studied patients with APS and systemic lupus erythematosus (SLE) managed at the Rheumatology Department of the Bichat University Hospital, Paris, between 1987 and 1996. Data were collected by chart review and by a 1997 standardized telephone interview. RESULTS: We identified 108 patients with APS managed during the ten-year study period. According to classical classification criteria, 39 patients had primary antiphospholipid syndrome (PAPS) and 69 secondary antiphospholipid syndrome (SAPS). Family history data were obtained for 29 (74%) and 55 (80%) PAPS and SAPS patients. respectively (78% of the 108 patients). Twelve PAPS (41% and 19 SAPS (35%) patients had one or more relatives with evidence of at least one clinical feature of APS such as thrombosis or recurrent fetal loss; of these patients, seven in the PAPS (24%) and 11 in the SAPS (20%) group had two or more relatives with evidence of a clinical feature of APS. Three PAPS (10%) and 14 SAPS (25%) patients had one or more family members with an autoimmune disease. CONCLUSION: A positive family history for autoimmune disease and/or antiphospholipid syndrome is common in patients with PAPS or SAPS. This finding supports a genetic contribution to APS. The percentage of a positive family history for autoimmune disease tend to be higher in patients with SAPS than in those with PAPS. PMID- 10866097 TI - Autoimmune antiphospholipid antibodies and cryoglobulinemia. AB - In addition to their role in the thrombotic manifestations of the antiphospholipid syndrome (APS), autoimmune antiphospholipid (aPL) antibodies may also be responsible for direct injury to the blood vessel wall, although the mechanism is unclear. Cryoglobulinemia has been reported infrequently in patients with APS and is one potential means of blood vessel injury. The aim of the present study was to determine if autoimmune aPL antibodies and their target antigens contribute to the formation of cryoprecipitates. Cryoglobulins were identified and isolated from 5 of 8 patients with autoimmune aPL antibodies. Using identical concentrations of immunoglobulins isolated from matched sera and washed cryoprecipitates there was a significant enrichment (at least 100%) of aCL antibodies in the cryoprecipitates from 4 of 5 patients. This involved IgG, IgM and IgA isotypes with specificity for both beta2-glycoprotein I (GPI) and prothrombin (PT). The target antigens were detected in cryoprecipitates from all 5 aPL positive patients and in cryoprecipitates from 3 controls. These results suggest that anti-beta2-GPI and anti-PT antibodies in association with their target antigens are integrally involved in the formation of cryoprecipitates in patients with autoimmune aPL antibodies and provide insight into a potential mechanism for blood vessel injury. PMID- 10866098 TI - Altered bone marrow cell sensitivity in the lupus-prone NZB/W mouse: regulation of CFU-GM colony formation by estrogen, tamoxifen and thrombopoietin. AB - Estrogen is thought to contribute to the onset of systemic lupus erythematosus (SLE) in women through mechanisms that are not completely understood. Although estrogen serves as a negative regulator in normal hematopoietic development, little research has been conducted examining alteration in hematopoietic development triggered by estrogen in lupus-susceptible individuals. We examined whether estrogen and other factors could influence colony formation of bone marrow cells obtained from normal and lupus-susceptible mice. Bone marrow cells isolated from New Zealand Black (NZB) and lupus-prone New Zealand Black and New Zealand White cross (NZB/W) mice were cultured in the presence of granulocyte macrophage colony stimulating factor (GM-CSF) alone or in combination with estrogen, thrombopoietin (TPO), tamoxifen, estrogen and TPO. or estrogen and tamoxifen, and plated in methylcellulose culture medium. Plates were scored for the number of CFU-GM (colony forming unit granulocyte-macrophage) colonies after 6d in culture. For females of both mouse strains, estrogen significantly decreased (P < 0.05) the number of GM colonies. Co-treatment of NZB/W cells, but not NZB cells, with TPO or tamoxifen reversed the suppressive action of estrogen (P < 0.05). In contrast, while estrogen did suppress colony formation from cells of NZB/W males (P < 0.05), neither TPO nor tamoxifen reversed this effect. Our results indicate that the sensitivity of bone marrow cells isolated from both female and male NZB/W lupus-prone mice to hormones/growth factors is qualitatively different from cells of NZB mice, and suggest that hematopoietic alterations at the level of the bone marrow may be related to the pathogenesis of SLE. PMID- 10866099 TI - Toxic effects of SLE serum on normal monocytes in vitro: cell death induced by apoptosis related to complement dysfunction. AB - The aim of this study was to assess toxic effects of systemic lupus erythematosus (SLE) serum on blood peripheral mononuclear cells from healthy donors and to evaluate if complement activation was involved. Monocytes from a healthy donor were incubated with 20 sera from ten SLE patients in both high and low disease activity states. After incubation non-adherent cells were analysed by flow cytometry. Serum from six SLE patients induced an increased cell death, four in active disease only, one in the inactive state and one in the active and the inactive state. Five of these sera, three with high and two with low disease activity, induced an increased apoptosis in the monocytes. Proportion of apoptotic cells correlated inversely with C1q and C3 concentration in the active disease sera, but not with disease activity as evaluated by SLEDAI. Apoptosis could be induced by addition of active C1s or antigen/antibody complexes to normal serum before incubation. Serum with complexes added induced increased tumour necrosis factor-alpha secretion from mononuclear cells, but SLE patient sera did not. The results demonstrate that the toxic effect of serum from SLE patients on healthy monocytes is explained by induction of apoptosis. The induction process is suggested to be connected with complement activation in the serum. PMID- 10866100 TI - Choroidopathy of systemic lupus erythematosus. AB - PURPOSE: To describe the ocular and systemic manifestations associated with systemic lupus erythematosus (SLE) choroidopathy. METHODS: Three new cases of choroidopathy in patients with active SLE were described. Twenty-five published cases of lupus choroidopathy were summarized. RESULTS: There have been 28 cases of lupus choroidopathy (47 involved eyes) that have been reported in the English literature since 1968, including the three current cases. Only two of the patients were male. The choroidopathy was bilateral in 19 patients (68%). All 28 patients (100%) had active systemic vascular disease at the onset of their choroidopathy; 18 (64%) had nephropathy and 10 (36%) had central nervous system (CNS) lupus vasculitis. All but one of the patients had a known diagnosis of SLE at the onset of choroidopathy. 30 of the 47 involved eyes had presenting visual acuity of 20/40 or better; 14 eyes showed improvement in visual acuity with therapy. 23 patients (82%) had resolution of their choroidopathy when their systemic disease was brought under control. Despite treatment, 4 of the 28 patients (14%) died from complications of SLE. CONCLUSIONS: Although less known than retinopathy, lupus choroidopathy may be more common than generally appreciated. It usually serves as a sensitive indicator of lupus activity. The presence of SLE choroidopathy is generally indicative of coexistent (although sometimes occult) nephropathy, CNS vasculitis, and other SLE visceral lesions. Immunomodulation of the systemic disease can lead to improvement and resolution of the systemic vasculitis as well as the choroidopathy. PMID- 10866101 TI - Paraoxonase activity is dramatically decreased in patients positive for anticardiolipin antibodies. AB - It has been reported that paraoxonase 1 (PON1) activity inhibits low-density lipoprotein (LDL) oxidation and modulates the risk of coronary heart disease. This study shows that autoantibodies (IgG) directed against modified LDL were increased in 71 patients positive for anticardiolipin antibodies. In a representative subgroup of these patients (n = 36) PON1 activity was dramatically decreased and the prevalence of the RR genotype of this enzyme tended to be increased in patients who had developed arterial thrombosis. This study suggests that PON1 abnormalities play a role in the antiphospholipid syndrome. PMID- 10866102 TI - Red ear(s) syndrome associated with child neuropsychiatric systemic lupus erythematosus. AB - This case illustrates that a child having severe neuropsychiatric systemic lupus erythematosus (NPSLE) with seizure and cerebral vascular disease showed excellent clinical outcome in response to intravenous methylprednisolone and cyclophosphamide pulse, and presented unexplained red ears phenomenon. PMID- 10866103 TI - Nocardia infection of a joint prosthesis complicating systemic lupus erythematosus. AB - The authors report the case of a 43-year-old woman suffering from severe systemic lupus erythematosus treated with long-term prednisone, who developed Nocardia nova infection on a hip prosthesis. Sepsis occurred about two years after an episode of pulmonary nocardiosis with the same Nocardia species, that was successfully treated by 12 months of antibiotics. A good outcome of the joint infection was observed in response to antibiotics and removal of the prosthesis. Nocardiosis is a rare infection, acting as an opportunistic infection, facilitated in the present case by systemic lupus erythematosus and chronic corticosteroid therapy. Nocardia infections mainly affect the lungs, skin and central nervous system; these last two sites are mostly due to haematogenous spread, a frequent event. Treatment is based on antibiotics, usually continued for 3-12 months, especially because of the risk of relapse. The imipenem-amikacin combination appears to be more effective than trimethoprim sulfamethoxazole. To our knowledge, this is the first case report of Nocardia nova joint prosthesis infection also presenting as late septic spread of pulmonary nocardiosis, complicating corticosteroid-treated systemic lupus erythematosus. PMID- 10866104 TI - Acute optic neuropathy and transverse myelopathy in patients with antiphospholipid antibody syndrome: favorable outcome after treatment with anticoagulants and glucocorticoids. AB - We describe two patients with established antiphospholipid syndrome, who during periods of subtherapeutic anticoagulation, developed acute optic neuropathy and transverse myelopathy. Treatment with optimal anticoagulation and high dose glucocorticoids was followed by resolution of the neurologic deficits. PMID- 10866105 TI - Cerebral blood flow and glucose metabolism in multi-infarct-dementia related to primary antiphospholipid antibody syndrome. AB - The primary antiphospholipid antibody syndrome (PAPS) has been described in patients with a history of fetal loss, thrombocytopenia and arterial or venous thrombosis. In PAPS, a prothrombotic state is mediated by antiphospholipid antibodies (aPLs) leading to disseminated thromboembolic vascular occlusion. Today, the presence of aPLs in the serum is considered as a distinct risk factor for recurrent stroke in young adults. Some PAPS patients develop a multi-infarct syndrome with a stepwise decline of higher cortical functions. We report on a 55 year-old man suffering from progressive dementia and PAPS, in whom cerebral glucose metabolism and blood flow were examined by positron emission tomography (PET). Cerebral atrophy and moderate signs of leukaraiosis were detected in magnetic resonance imaging (MRI), whereas the PET scans showed a considerable diffuse impairment of cortical glucose metabolism combined with a reduced cerebral perfusion in the arterial border zones. These findings indicate that PAPS-associated vascular dementia is accompanied by a cortical neuronal loss, presumably caused by a small-vessel disease with immune-mediated intravascular thrombosis. This case shows that pathological findings in PAPS are congruent to cerebral changes of metabolism and blood flow in systemic lupus erythematosus (SLE). PMID- 10866106 TI - Major histocompatibility complex class I diversity in a West African chimpanzee population: implications for HIV research. AB - Human immunodeficiency virus (HIV) poses a major threat to humankind. And though, like humans, chimpanzees are susceptible to HIV infection, they are considered to be resistant to the development of the acquired immune deficiency syndrome (AIDS). Little is known about major histocompatibility complex (MHC) class I diversity in chimpanzee populations and, moreover, whether qualitative aspects of Patr class I molecules may control resistance to AIDS. To address these questions, we assayed MHC class I diversity in a West African chimpanzee population and in some animals from other subspecies of chimpanzee. Application of different techniques allowed the detection of 17 full-length Patr-A, 19 Patr B, and 10 Patr-C alleles. All Patr-A alleles cluster only into the HLA-A1/A3/A11 family, which supports the idea that chimpanzees have experienced a reduction in their repertoire of A locus alleles. The Patr-B alleles do not cluster in the same lineages as their human equivalents, due to frequent exchange of polymorphic sequence motifs. Furthermore, polymorphic motifs may have been exchanged between Patr-A and Patr-B loci, resulting in convergence. With regard to evolutionary stability, the Patr-C locus is more similar to the Patr-A locus than it is to the Patr-B locus. Despite the relatively low number of animals analyzed, humans and chimpanzees were ascertained as sharing similar degrees of diversity at the contact residues constituting the B and F pockets in the peptide-binding side of MHC class I molecules. Our results indicate that within a small sample of a West African chimpanzee population, a high degree of Patr class I diversity is encountered. This is in agreement with the fact that chimpanzees display more mitochondrial DNA variation than humans. In addition, population analyses demonstrated that particular Patr-B molecules, with the capacity to bind conserved HIV-1 epitopes, are characterized by high gene frequencies. These findings have important implications for evaluating immune responses in HIV vaccine studies and, more importantly, may help in understanding the relative resistance of chimpanzees to AIDS. PMID- 10866107 TI - Common chimpanzees have greater diversity than humans at two of the three highly polymorphic MHC class I genes. AB - MHC class I polymorphism improves the defense of vertebrate species against viruses and other intracellular pathogens. To see how polymorphism at the same class I genes can evolve in different species we compared the MHC-A, MHC-B, and MHC-C loci of common chimpanzees and humans. Diversity in 23 Patr-A, 32 Patr-B, and 18 Patr-C alleles obtained from study of 48 chimpanzees was compared to diversity in 66 HLA-A, 149 HLA-B, and 41 HLA-C alleles obtained from a study of over 1 million humans. At each locus, alleles group hierarchically into families and then lineages. No alleles or families are shared by the two species, commonality being seen only at the lineage level. The overall nucleotide sequence diversity of MHC class I is estimated to be greater for modern chimpanzees than humans. Considering the numbers of lineages, families, and alleles, Patr-B and Patr-C have greater diversity than the HLA-B and HLA-C, respectively. In contrast, Patr-A has less polymorphism than HLA-A, due to the absence of A2 lineage alleles. The results are consistent with ancestral humans having passed through a narrower population bottleneck than chimpanzees, and with pathogen mediated selection having favored either preservation of A2 lineage alleles on the human line and/or their extinction on the chimpanzee line. PMID- 10866108 TI - Organization of the human interleukin-1 receptor antagonist gene IL1HY1. AB - The interleukin (IL)-1 family of proteins plays an important role in inflammatory and defense mechanisms. The recently characterized IL1HY1 cDNA encodes a new member of the IL-1 receptor antagonist family (IL-1ra). In this report, we describe the complete nucleotide sequence of the human IL1HY1 gene. We sequenced approximately 7,600 nucleotides and found four coding exons ranging in size from 55 to 2,288 nucleotides. The 5' untranslated region is formed by one of two alternatively used exons and one invariably present exon which also contains the region encoding the first nine amino acids of the protein. IL1HY1 and IL-1ra intron positions are well conserved within the protein-coding region, providing evidence that these genes arose from a duplication of a primordial IL-1 receptor antagonist gene. PMID- 10866109 TI - Polymorphisms in IgG Fc receptor IIB regulatory regions associated with autoimmune susceptibility. AB - Autoimmune diseases involve multiple genes. While functions of these genes are largely unknown, some may be related to an intrinsic hyperresponsiveness of B cells. B-cell responses are controlled by signaling thresholds through the B-cell antigen receptor (BCR) complex. The B1 isoform of type II IgG Fc receptors (FcgammaRIIB1) is exclusively expressed on B cells and serves as a negative regulator for inhibiting BCR-elicited activation. Thus, its allelic variants associated with functional deficits could be examined for possible associations with susceptibility to autoimmune diseases. We found that there are three types of polymorphisms in the reported FcgammaRIIB transcription regulatory regions in mouse strains. Compared to normal healthy mouse strains (group III), autoimmune disease-prone strains (group I) share three deletion sites: two in the promoter region and one in the third intron. Strains (group II) that per se are not autoimmune-prone, but have potentials to accelerate autoimmune diseases share two deletion sites in the third intron: one identical to that in group I and the other unique to group II. These polymorphisms correlated well with extents of down-regulation of FcgammaRIIB1 expression in germinal-center B cells upon stimulation with antigens and up-regulation of IgG antibody responses. Our data imply that these FcgammaRIIB polymorphisms are selected evolutionarily for natural defense against pathogens, and that such polymorphisms may, in turn, form the basis of one aspect of autoimmune susceptibility. PMID- 10866111 TI - Characterization of approximately 50 kb of the immunoglobulin VH locus of the Japanese pufferfish, Fugu rubripes. AB - The variable region of the immunoglobulin heavy chain is created by a somatic rearrangement of a limited number of germline genes. This mechanism of gene assembly [V(D)J recombination] has been found to take place only in jawed vertebrates (gnathostomes). To understand how this mechanism evolved and diversified it is necessary to study the genomic organization of the heavy-chain gene in different vertebrate lineages. Since there is scant sequence information on the VH locus in fish, shotgun sequencing of a cosmid clone containing part of the VH genomic region of the Japanese pufferfish, Fugu rubripes, was undertaken. Eight full-length VH genes were isolated and characterized. They have higher homology to trout genes, but show the same structural features as VHs found in other vertebrates. Two VH subgroups have been identified whose members are interspersed. The frequency of synonymous and nonsynonymous substitution for VH comparisons between family members was found to be higher in the complementarity determining regions than in the framework regions. Finally, there are four other genes interspersed with the VH genes, one of which is the first full-length retrotransposon element characterized in vertebrates. PMID- 10866110 TI - Molecular cloning, gene structure, and expression pattern of pig immunoreceptor DAP12. AB - Natural killer (NK) cells express receptors for MHC class I that contain immunoreceptor tyrosine-based inhibitory motif (ITIM) sequences in their cytoplasmic domain. Whereas these receptors inhibit NK cell cytotoxicity, certain isoforms of these NK receptors (e.g., KIR2DS, CD94/NKG2C, and Ly49D) do not have ITIMs, but associate with DAP12 and activate NK cell function. We cloned pig DAP12 cDNA from a pig peripheral blood lymphocyte (PBL) cDNA library using human DAP12 cDNA as a probe. The length of the pig DAP12 cDNA is 526 bp and contains an open reading frame of 324 bp. It has 79% identity with the human DAP12 cDNA sequence in the coding region and 73% identity with mouse DAP12 cDNA. The predicted polypeptide sequence of pig DAP12 is 108 amino acids, being composed of a 23-amino acid leader, a 14-amino acid extracellular domain, a 24-amino acid transmembrane segment, and a 47-amino acid cytoplasmic region. The amino acid sequence of pig DAP12 has 74% and 71% sequence identity with human DAP12 and mouse DAP12, respectively. Pig DAP12 has a conserved aspartic acid in the transmembrane region, and two conserved cysteine residues in the extracellular domain. It also contains an immunoreceptor tyrosine-based activation motif sequence in the cytoplasmic region. Genomic organization reveals that pig DAP12 consists of five exons and four introns. Southern blot analysis of pig genomic DNA revealed that DAP12 is a single-copy gene. In Northern blot analysis, DAP12 transcripts were detected in spleen, liver, thymus, and lymph node. DAP12 transcripts are expressed not only in PBLs, but also in granulocytes, macrophages, and monocytes. PMID- 10866112 TI - Immunoglobulin D (IgD) of Atlantic cod has a unique structure. AB - A new immunoglobulin heavy-chain gene with some homology to mammalian IgD was recently cloned from the channel catfish and Atlantic salmon, two species of teleost fish. We have cloned and sequenced a new H-chain gene from Atlantic cod (Gadus morhua L.) which has clear similarities to these genes, but which also differs in several ways. The similarities of catfish, salmon, and cod delta to the mammalian delta genes are sequence homology, location immediately downstream of IgM (mu), and expression by alternative splicing rather than class switching. A unique feature of catfish, salmon, and cod delta is the chimeric nature of the gene product, as the mu1 exon is spliced to the delta1 exon. Several unique features of cod IgD were found: (1) a deletion of the delta3, delta4, delta5, and delta6 domains described in catfish and salmon IgD, (2) a tandem duplication of a part of the delta locus including the delta1 and delta2 domains, (3) the presence of a truncated delta7 domain downstream of the deltaTM exons, and (4) the separation of the duplicated domains by a short exon (deltay) which has homology to a conserved part of the transmembrane exon 1 (TM1) of some H-chain isotypes. This unique organization of the delta locus of cod probably developed after the evolutionary split from the catfish and salmon branches. PMID- 10866113 TI - Is a mutator analogous to the Ig hypermutator of the sheep ileal Peyer's patch active on MHC class I genes in the germ line? AB - Polymorphic sequence variation in the peptide-binding domains of MHC class I molecules appears to have been driven largely by the constructive action of natural selection on the specificity of the peptide-binding groove. Similar features are displayed by the variable domains of immunoglobulins generated in the sheep ileal Peyer's patch, but in this case there is evidence that the action of a targeted hypermutator acting on a selected substrate rather than antigen driven selection is responsible for the pattern of variation in the system. Such a hypermutator acting in the germ line would not only mimic the action of natural selection but also, by convergent mutation, generate similar patterns of variation in unrelated alleles that could be interpreted as evidence for short tract gene conversion. We analyzed human class I MHC alleles in the light of these data, but failed to find evidence of the action of a similar hypermutator. A search for other mutationally driven patterns of variation also failed, even in hypervariable residues from parsimonious phylogenies. Single-nucleotide variation at these residues is also frequent in recent allelic variants, but the data are as consistent with short-tract gene conversion as with base mutation. We conclude that the patterns of allelic variation in MHC molecules are not driven by mutational pressure, but rather by conventional mutational processes, accompanied by short-tract gene conversion and intense natural selection. PMID- 10866115 TI - New alleles of human immunoglobulin kappa J segments IGKJ2 and IGKJ4. PMID- 10866116 TI - Sequence of HLA-A*0209 confirmed. PMID- 10866114 TI - Evolution of introns and exons of class II major histocompatibility complex genes of vertebrates. AB - The phylogenetic relationships and patterns of nucleotide substitution were compared for introns and exons of class II major histocompatibility complex (MHC) genes in three datasets: human DRB1, human DQA1, and cyprinid fish DAB1. In both human DRB1 and cyprinid DAB1, there was strong evidence that recombination events between alleles have occurred in such a way that intron and exon sequences of a given allele do not necessarily share the same evolutionary history. In the case of human DRB1, recombination was found to have homogenized intron 1 and intron 2 sequences relative to exon 2 sequences within lineages of alleles but not between lineages. As a result, mean divergence times of intron sequences are much more recent than those of exonic sequences. Thus, the divergence time of DRB1 introns cannot be used to date that of exons in the same alleles, and the hypothesis that most human DRB1 polymorphism is of very recent origin is not supported. PMID- 10866117 TI - Mhc class I gene of African lungfish. PMID- 10866118 TI - Characterization of rhesus monkey CD94/NKG2 family members and identification of novel transmembrane-deleted forms of NKG2-A, B, C, and D. PMID- 10866119 TI - Complementary DNA sequences encoding rat T-cell receptor delta-chain D1-, D2-, J1 , and C-region proteins. PMID- 10866120 TI - Cloning of three different species of MHC class I cDNAs of the RT1g haplotype from the NEDH rat. PMID- 10866121 TI - Genomic organization and 5' regulatory region of the mouse IL-2 receptor beta chain gene (IL-2Rb). PMID- 10866122 TI - The antigen that wasn't--silicone. PMID- 10866123 TI - T cell memory: heterogeneity and mechanisms. AB - Immunological memory is manifested by the body's ability to enjoy long-term protection against specific pathogens previously encountered through illness or vaccination. This memory response resides in the long-lived, previously activated memory T and B lymphocytes that are believed to exist in a quiescent state. Recent advances in studies on T cell memory have revealed heterogeneity in the T cells that mediate memory responses that may have implications for the generation and maintenance of these cells over time. This review will present these recent findings on memory T cells in the context of past research and current models for the generation and persistence of memory T cells. PMID- 10866124 TI - Synergy in induction of increased renal allograft survival after portal vein infusion of dendritic cells transduced to express TGFbeta and IL-10, along with administration of CHO cells expressing the regulatory molecule OX-2. AB - Dendritic cells (DC), generated from C57BL/6 mouse bone marrow cells cultured with GM-CSF and IL-4 for 9 days, were engineered to express constitutively the cytokines TGFbeta, IL-10, and IL-12, using adenovirus vectors constructed using an E1-deleted replication-deficient recombinant adenovirus carrying the appropriate cDNA for the relevant cytokines (Ad-TGFbeta, Ad-IL-12, or Ad-IL-10). C3H mice receiving nontransduced DC or pretransplant infusion of DC-Ad-LacZ showed increased survival of C57BL/6 renal grafts relative to that of control nonimmunized mice. Transfusion of Ad-IL-12-transduced DC abolished this increased survival, leading to a graft survival equivalent to that of controls with no DC. Optimal graft survival was seen in the group receiving a mixture of DC transduced with constructs for both IL-10 and TGFbeta. There was a correlation between increased graft survival and both inhibition of the induction of CTL and enhancement of a polarization to produce type-2 cytokines (IL-4, IL-10, and TGFbeta) on antigen-specific restimulation in vitro. These effects were more pronounced following concomitant infusion of CHO cells transfected with a full length cDNA for murine OX-2. PMID- 10866125 TI - Kinetic analysis of the interaction between recombinant human Fc(epsilon)RIalpha and serum IgEs from allergic patients. AB - Binding of IgE to the high-affinity IgE receptor (Fc(epsilon)RI) is the essential event for allergic reaction. Although there are many reports on binding kinetics between myeloma IgE and Fc(epsilon)RI, little is known about the kinetics between heterogeneous polyclonal IgE in the serum and Fc(epsilon)RIalpha. To elucidate the binding characteristics of heterogeneous serum IgE, we measured kinetic parameters of binding between IgE from allergic patients and a recombinant ectodomain of the human Fc(epsilon)RIalpha subunit by real-time interaction analysis based on surface plasmon resonance. Purified IgE monomer from the plasma of allergic patients displayed kinetics for the interaction with Fc(epsilon)RIalpha similar to those of myeloma IgE. In the case of crude IgE samples from allergic patients, one of seven specimens showed significantly higher affinity than highly purified IgE, suggesting that it is possible for IgEs in this specimen to form complexes of higher molecular weight. PMID- 10866126 TI - Endothelial cell autoantibodies are a marker of disease susceptibility in inflammatory bowel disease but apparently not linked to persistent measles virus infection. AB - Intestinal vasculitis caused by persistent measles virus infection of intestinal endothelial cells was described in Crohn's disease. Furthermore, endothelial cell autoantibodies have been demonstrated in inflammatory bowel disease (IBD). Autoantibodies against intestinal endothelial cells were visualized by indirect immunofluorescence in patients with IBD, in their healthy first-degree relatives, in patients with infectious enterocolitis, and in healthy, unrelated controls. In intestinal tissue specimens of 22 antibody-positive IBD patients a search for the measles virus genome was performed. Endothelial cell autoantibodies were significantly more frequent in patients with IBD, in both groups of first-degree relatives, and in patients with infectious enterocolitis than in the healthy controls (P = 0.0002 or less). The measles virus genome was found in none of the intestinal biopsies. Endothelial cell autoantibodies are not a genetic but rather an epigenetic (infectious) marker of disease susceptibility. The expression of these autoantibodies is unlikely to be triggered by a persistent measles virus infection. PMID- 10866127 TI - Differential roles of Fas ligand in spontaneous and actively induced autoimmune encephalomyelitis. AB - To determine the roles of Fas/Fas ligand (FasL) in autoimmunity, we studied spontaneous and actively induced autoimmune encephalomyelitis in 541 myelin basic protein-specific T cell receptor transgenic mice. We found that spontaneous autoimmune encephalomyelitis, which was initiated by unidentified microbial factors, was dramatically exacerbated in mice carrying Fas or FasL gene mutation. The exacerbation of autoimmune encephalomyelitis was reflected primarily by an increase in disease incidence and a decrease in spontaneous disease recovery. By contrast, actively induced encephalomyelitis, which was initiated by pertussis toxin, was significantly inhibited by Fas or FasL gene mutation. These results suggest that environmental factors that trigger autoimmune disease may determine not only whether disease will occur but also whether an immune molecule such as FasL will promote or inhibit the autoimmune process. PMID- 10866128 TI - Changes in circulating levels of soluble cell adhesion molecules following highly active antiretroviral treatment of HIV-1-infected patients. AB - Increased levels of soluble cell adhesion molecules (sCAM) have been reported in HIV-1 infection and may possibly contribute to altering the adhesion mechanisms of phagocytic cells. We evaluated the effect of highly active antiretroviral therapy (HAART) on plasma levels of sL-selectin, sE-selectin, intercellular cell adhesion molecule-1 (sICAM-1), sICAM-3, and vascular cell adhesion molecule-1 (sVCAM-1). Study participants included 22 HIV-1-infected patients with a CD4+ T cell count/microl below 500 who were started on a HAART regimen and followed up for 9 months. After the initiation of therapy, plasma sL-selectin concentrations progressively decreased to normal ranges in the majority of our patients (P < 0.001), while no changes in sE-selectin were found. In all patients sICAM-1 remained relatively constant at significantly elevated concentrations during the 9 months of therapy. A significant reduction in plasma concentrations of both sICAM-3 and sVCAM-1 was found; however, the levels of these sCAM were not normalized by HAART and remained significantly elevated throughout the study (P < 0.001). The reduced release of sL-selectin could improve the ability of phagocitic cells to migrate in response to chemotactic stimuli after starting HAART. On the other hand, the persistent elevation of sICAM-1, sICAM-3, and sVCAM 1 could reflect continuous HIV-1-mediated immune activation, despite adequate control of plasma HIV-1 replication by therapy. PMID- 10866129 TI - Induction of Fas ligand-mediated apoptosis by interferon-alpha. AB - Interferon-alpha (IFN-alpha) was among the first cytokines studied and the earliest to be used in clinical medicine for the treatment of viral infections and malignancies. Although the capacity of IFN-alpha to augment NK cell cytotoxicity against virus-infected target cells or tumor cells is well established, the mechanism has not been fully elucidated. Here we report that IFN alpha stimulation of PBMC from healthy donors induces Fas (CD95) ligand (FasL) transcription and leads to increased cell surface FasL expression exclusively on the NK cell fraction. Furthermore, IFN-alpha augments the FasL-mediated cytotoxicity of normal PBMC against Fas-sensitive lymphoid tumor cells. In the context of innate immunity, induction of FasL by IFN-alpha can be viewed as an efficient mechanism to potentiate NK cell cytotoxicity in the presence of harmful targets, such as virally infected or transformed cells. PMID- 10866130 TI - Shiga toxin-2 induces neutrophilia and neutrophil activation in a murine model of hemolytic uremic syndrome. AB - It has been demonstrated that infections due to Shiga toxins (Stx) producing Escherichia coli are the main cause of the hemolytic uremic syndrome (HUS). Although it is recognized that Stx damage the glomerular endothelium, clinical and experimental evidence suggests that the inflammatory response is able to potentiate Stx toxicity. Lipopolysaccharides (LPS) and neutrophils (PMN) represent two central components of inflammation during a gram-negative infection. In this regard, patients with high peripheral PMN counts at presentation have a poor prognosis. Since the murine model has been used to study LPS-Stx interactions, we analyzed the effects of Stx alone or in combination with LPS on the kinetics of neutrophil production and activation and their participation in renal damage. We observed a sustained neutrophilia after Stx2 injection. Moreover, these neutrophils showed increased expression of CD11b, enhanced cytotoxic capacity, and greater adhesive properties. Regarding the cooperative effects of LPS on Stx2 action, we demonstrated potentiation of neutrophilia and CD11b induction at early times by pretreatment with LPS. Finally, a positive correlation between neutrophil percentage and renal damage (assayed as plasmatic urea) firmly suggests a role for PMN in the pathogenesis of HUS. PMID- 10866131 TI - Induction of IgA against Haemophilus parainfluenzae antigens in tonsillar mononuclear cells from patients with IgA nephropathy. AB - Much evidence suggests that IgA production in vivo and in vitro is enhanced in patients with IgA nephropathy (IgAN). We have demonstrated glomerular deposition of the outer membranes of Haemophilus parainfluenzae (HP) antigens (OMHP) and the presence of HP-specific IgA in the serum of patients with IgAN. In this study, we investigated the production of IgA and several cytokines by tonsillar mononuclear cells (TMC) from IgAN patients induced by stimulation with OMHP. The spontaneous production of total IgA and TGF-beta by TMC from IgAN patients was higher than that by TMC from patients with chronic tonsillitis (CT) (P < 0.05). Stimulation with OMHP in vitro enhanced the production of HP-specific IgA by TMC from IgAN patients (P < 0.01), but not by TMC from CT patients. OMHP stimulation also enhanced the production of TGF-beta and IL-10 by TMC from IgAN patients (P < 0.001). These results suggest that the infection of HP in the tonsil may be involved in the etiology of IgAN. PMID- 10866132 TI - Adventures with heterocycles. AB - Elucidation of mechanisms and discovery of novel reactions are interconnected, as four research chapters from the Munich laboratory demonstrate. 1) In a new opening of the furan ring, 2-methoxyfuran and tetra-acceptor-substituted ethylenes furnish methyl acrylates, which are cis-3-substituted by a cyclopropyl ring that bears the four acceptor groups. Experiments with trans- and cis-1,2 bis(trifluoromethyl)-1,2-dicyanoethylenes (BTE) clarify the role of zwitterionic intermediates. 2) The concerted nature of 1,3-dipolar cycloaddition is well established. On reacting thiocarbonyl S-ylides as electron-rich 1,3-dipoles with tetra-acceptor-substituted ethylenes, the switching to a two-step pathway via zwitterion was diagnosed from a loss of stereospecificity and the formation of strained 7-membered cyclic ketene imines. 3) Stable as crystals, these ketene imines rearrange to 5-membered thiolanes in solution. In contrast to open-chain ketene imines, the cyclic representatives add vinyl ether at the C=N bond and show a novel pathway of dimerization. 4) Cycloadducts of isoquinolinium N phenylimide with ethylenic dipolarophiles undergo an acid-catalyzed [3.3] sigmatropic hydrazo rearrangement. An exception is the conversion of the dimethyl maleate adduct (C21H20N2O4) by acid into a yellow compound C24H22N2O6; the structure and the astounding mechanism were clarified. PMID- 10866133 TI - Four new terpenes from the pericarp of Platycladus orientalis. AB - A new monoterpene, platydiol (1), and three new diterpenes, platyclolactonic acid (2), 14,15-bisnor-8(17)-labdene-16,19-dioic acid (3), and 6,7 dehydrosandaracopimaric acid (4), were isolated from the pericarp of Platycladus orientalis. Their structures were elucidated by spectral and chemical methods. PMID- 10866134 TI - The effects of adsorbed water on tensile strength and Young's modulus of moldings determined by means of a three-point bending method. AB - Young's moduli (E) of three representative tableting excipients and their mix powders were measured for compressed rectangular beam specimens over a range of porosities using a three-point bending technique. We also examined the effects of the amount of water adsorbed on the tensile strength of these specimens. The maximal tensile strength (sigma(max)) decreased with increasing water vapor adsorption for microcrystalline cellulose (MCC) and mixed powders of lactose and MCC. Sigma(max) increased with increasing compression stress and specimen weight for all samples. Sigma(max) of an alpha-lactose and cornstarch mixture with a ratio of 7:3 showed a large value. Young's modulus (E) and the crushing energy (CE) of MCC were larger than those of the other samples. Young's modulus of specimens decreased as the proportion of alpha-lactose increased. Disintegration time (DT) of tablets comprised of lactose and MCC mixture was much faster than those of tablets comprised of individual powders. This appeared to demonstrate the effect of MCC swelling on the disintegration time of the tablet. The disintegration time of the lactose/cornstarch series increased only when Young's modulus increased sharply. PMID- 10866135 TI - Single strand conformation polymorphism analysis of ras oncogene by capillary electrophoresis with laser-induced fluorescence detector. AB - Single strand conformation polymorphism (SSCP) analysis of the N-ras oncogene was achieved by capillary electrophoresis with a laser-induced fluorescence detector (CE-LIF) using methylcellulose as a molecular sieving agent. The PCR-amplified N ras oncogene, which is known to have a point mutation at codon 61 in the neuroblastoma, was investigated by CE-LIF combined with SSCP (SSCP-CE-LIF). A mixture of wild- and mutant-type single strand DNA fragments (103 bp) of the N ras oncogene was separated by buffer solution containing 1.0% methylcellulose and 0.2 microM fluorescent dye (YO-PRO-1) at 25 degrees C. The SSCP-CE-LIF technique gave good resolution for wild- and mutant-type single strand DNA fragments with separation completed within 7 min. SSCP analysis using a CE system with a LIF detector was successfully applied to the detection of the one point mutation on the N-ras oncogene. PMID- 10866136 TI - Characteristics of complexes composed of sodium hyaluronate and bovine serum albumin. AB - Complexes composed of sodium hyaluronate (NaHA) and bovine serum albumin (BSA) were studied to elucidate the exact composition of the complex, the phase separation, the electrophoretic mobility and the size using dynamic light scattering (DLS) and electrophoretic light scattering (ELS), etc. The phase diagram of the mixed solutions was determined. The complexes were soluble in neutral or weakly acidic pH regions and showed phase separation in the more acidic pH region. From the concentration of Na+ released from NaHA when it binds to BSA, the ratios of BSA to NaHA of the complexes were determined. In the region of soluble complexes, one BSA molecule was determined to bind with 15 carboxylic groups of NaHA and in the region of insoluble complexes to bind with 6 carboxylic groups. At the phase separation point, 117 BSA molecules bound with one NaHA molecule and 17% of the carboxylic groups of NaHA did not contribute to the binding of BSA. The sizes of the complexes decreased from several microm to several hundred nm as the binding ratio of BSA increases. Decreases in the viscosities of the mixed solutions were consistent with the decreases of the sizes. From these results, a model of complex formation is proposed. PMID- 10866137 TI - ESR study on the antioxidant activity of TAK-218 in biological model membranes. AB - TAK-218 has a 2,3-dihydrobenzofuran-5-amine (coumaran) structure which resembles alpha-tocopherol, and is a promising candidate as an agent for central nervous system (CNS) trauma and ischemia. The radical scavenging activity of TAK-218 was studied using electron spin resonance (ESR) spectroscopy. TAK-218 exhibited a more potent scavenging activity towards the hydroxyl radical than did the well known hydroxyl radical scavengers, mannitol and dimethylsulfoxide. Towards the superoxide radical, TAK-218 showed equal potency to glutathione. TAK-218 reacted rapidly with stable radicals, such as galvinoxyl and 2,2-diphenyl-1 picrylhydrazyl hydrate (DPPH), and gave the quinone as a two-electron oxidized product in analogy with alpha-tocopherol. To exhibit an excellent antioxidative activity in living systems, the compounds should not only have the intrinsic radical scavenging activity but also good distribution in the biological lipid bilayer membrane. To examine the antioxidant activity of TAK-218, the inhibition of lipid peroxidation by alpha-tocopherol and TAK-218 in liposomal membranes was studied using an ESR spin-label technique. Both alpha-tocopherol and TAK-218 completely inhibited lipid peroxidation by radicals generated in an aqueous layer using a water-soluble radical initiator, 2,2'-azobis-(2-amidinopropane) hydrochloride (AAPH). At a high incubation temperature (45 degrees C), alpha tocopherol scavenged radicals more effectively than TAK-218 on the surface of the membrane, while TAK-218 scavenged radicals more effectively in the interior of the membrane. The difference between TAK-218 and alpha-tocopherol for radical scavenging in the membrane system derives from the different distribution pattern of these compounds. TAK-218 can penetrate the membrane freely and can scavenge the radical in the membrane interior. Furthermore, TAK-218 was shown to inhibit lipid peroxidation initiated by a lipid soluble radical initiator, 2,2'-azobis (2,4-dimethylvaleronitrile) (AMVN), in a membrane more effectively than alpha tocopherol. PMID- 10866138 TI - Study of tolbutamide-hydroxypropyl-gamma-cyclodextrin interaction in solution and solid state. AB - Tolbutamide-hydroxypropyl-gamma-cyclodextrin (TBM-HPGCD) interaction has been investigated in an aqueous environment and in the solid state. The solubility of TBM was increased in accord with the amount of HPGCD added to the aqueous medium forming a soluble inclusion compound. The phase solubility diagram obtained was of A(L) type. Physical mixtures and kneaded systems of the drug and cyclodextrin derivative were prepared in 1:1 and 1:2 drug/cyclodextrin mol/mol ratio. All solid binary systems were characterised by hot-stage microscopy (HSM), differential scanning calorimetry (DSC), thermogravimetry (TG) and X-ray powder diffractometry (XRD). An inclusion complex was formed in both of the kneaded systems. In the 1:2 kneaded system, the entire drug was included in the cyclodextrin cavity, while, in the 1:1 kneaded system only a part of the drug formed an inclusion complex with the cyclodextrin. A significant improvement in the dissolution of the drug was obtained from the kneaded systems in comparison with that of the pure TBM and physical mixtures. However, there was no significant difference between the dissolution profiles of the two kneaded systems. The study suggests that an inclusion complex was obtained both in aqueous solution and in solid state. PMID- 10866140 TI - Phosphorylation of D-glucose derivatives with inorganic cyclo-triphosphate: substituent effect at the 5-CH2OH or 2-OH group. AB - The phosphorylation of several D-glucose derivatives has been achieved using inorganic sodium cyclo-triphosphate hexahydrate (P3m), Na3P3O9 x 6H2O, in aqueous solution. In the phosphorylation of D-glucuronic acid, 6-phosphoryl-D-glucose and D-xylose, beta-D-glucuronic acid 1-triphosphate, 6-phosphoryl-beta-D-glucose 1 triphosphate and beta-D-xylose 1-triphosphate were synthesized stereoselectively with maximum yields of 43.5, 32.8 and 41.9%, respectively. In the case of D glucosamine and 2-deoxy-D-glucose, the main phosphorylated products were assigned to beta-D-glucosamine 1-triphosphate and 2-deoxy-beta-D-glucose 1-triphosphate by 1H-, 13C- and 31P-NMR, and the yields were 13.9 and 13.4%, respectively. PMID- 10866139 TI - Absorption, first-pass metabolism, and disposition of itraconazole in rats. AB - This study examined the pharmacokinetic disposition, oral absorption and hepatic extraction of itraconazole and its active metabolite, hydroxyitraconazole, in rats. After i.v. injection, serum itraconazole concentrations decreased biexponentially, with an average terminal elimination half-life, volume of distribution and systemic clearance of 4.9 h, 6.0 l/kg and 14.2 ml/min/kg, respectively. When given orally, its absorption was low, with a mean absolute bioavailability of 16.6%. The metabolite to parent drug area under the curve (AUC) ratio was higher after oral administration compared with i.v. injection (mean ratio, 2.7 vs. 0.9). The hepatic drug extraction ratio determined after femoral and portal vein administration averaged 18.5%. When hydroxyitraconazole was injected i.v., the elimination half-life, volume of distribution and systemic clearance of itraconazole averaged 10.0 h, 2.4 l/kg and 3.4 ml/min/kg, respectively. The fraction of the systemically available itraconazole that was metabolized to hydroxyitraconazole was 21.0% and 76.0% after i.v. and oral administration, respectively. In summary, this study is the first reporting the hepatic extraction of itraconazole and the i.v. disposition characteristics of hydroxyitraconazole in rats. Itraconazole is a drug with a low hepatic extraction ratio and its systemic clearance appears to be largely accounted for by hepatic metabolism. PMID- 10866141 TI - Study of synthesis and cardiovascular activity of some furoxan derivatives as potential NO-donors. AB - A series of hybrid molecules incorporating the furoxan and nicorandil moieties were designed as potential NO donors with cardiovascular and cerebrovascular activities. Thirty-six target molecules were successfully synthesized by conventional methods and characterized by infrared spectroscopy, 1H-NMR spectroscopy and high resolution mass spectra. The compounds were tested for their effects on KCl-induced contraction of rabbit thoracic aorta whose endothelium was denuded. Eight compounds were found to reduce KCl-induced contraction by more than 30% at 10 microM. All except one of these compounds are characterized by the presence of electron withdrawing groups in the phenyl ring attached via an amide or ester linkage to the furoxan moiety. The nature of the terminal carbonyl linkage (ester or amide) and the length or type of the alkyl chain bridging the two carbonyl functions have little effect on the activity. One of the active compounds, N-(4-methoxy-benzoyl)-N'-[3-methylfuroxanyl-4 carbonyl)piperazine (17i) was tested for hypotensive effects on anaesthesized rats at 1.5 mg/kg, and found to demonstrate a gradual and sustained hypotensive effect. The results suggest that the furoxan-nicorandil derivatives are a useful lead in the design of NO-donor compounds for hypertension. PMID- 10866142 TI - Bioavailable acyl-CoA: cholesterol acyltransferase inhibitor with anti peroxidative activity: synthesis and biological activity of novel indolinyl amide and urea derivatives. AB - We synthesized a series of indoline derivatives with an amide or urea moiety and examined their inhibitory effects on acyl-CoA:cholesterol acyltransferase (ACAT) activity, lipid-peroxidation and serum cholesterol levels in experimental animals. Among the derivatives synthesized, a series of N-(1-alkyl-4,6 dimethylindolin-7-yl)-2,2-dimethylpropanamides++ + potently inhibited rabbit intestinal ACAT activity and lipid-peroxidation of rat brain homogenate. The effect on ACAT activity was related to the length of the alkyl chain at the 1 position of indoline. N-(4,6-Dimethyl-1-octylindolindolin-7-yl)-2,2 dimethylpropanami de hydrochloride (55) showed inhibitory effects on intestinal and hepatic ACAT activity slightly weaker than those of YM-750, and an inhibitory effect on low density lipoprotein (LDL)-peroxidation similar to that of probucol. Compound 55 also reduced serum cholesterol at 10 mg/kg/d in hyperlipidemic rats and 20 mg/kg/d in normolipidemic hamsters. The plasma concentration of 55 reached 716 ng/ml in dogs (10 mg/kg, p.o.), which is an effective concentration against hepatic ACAT activity and LDL-peroxidation. In conclusion, compound 55 is a novel bioavailable ACAT inhibitor with anti-peroxidative activity and is thus a promising anti-atherosclerotic and anti-hyperlipidemic drug. Indoline proved to be a useful pharmacophore for molecular design of new anti-peroxidative drugs. PMID- 10866143 TI - Four new triterpene glycosides from Thalictrum squarrosum. AB - Four new triterpene glycosides were isolated from the dried aerial parts of Thalictrum squarrosum (Ranunculaceae). They were designated as squarroside I, being a cycloartane-type glycoside, and squarrosides II, III and IV, being oleanene-type glycosides. Their structures were established by using two dimensional (2D) NMR techniques. PMID- 10866144 TI - Bicyclo[3.2.1]octane and 6-oxabicyclo[3.2.2]nonane type neolignans from Magnolia denudata. AB - From the ethyl acetate soluble fraction of twigs of Magnolia denudata (Magnoliaceae), seven new neolignan derivatives, 1-7, were isolated along with eighteen known lignan and neolignan derivatives, 8-25. The structures of the new neolignans were elucidated by means of spectral methods, especially by 1H-NMR and 13C-NMR spectra, and two dimensional NMR methods such as 1H-detected heteronuculear multiple bond connectivity1 (HMBC), 1H-detected multiple quantum coherence (HMQC) and 1H-1H-correlation spectroscopy (COSY). Compounds 1-4 have novel structures possessing a 6-oxabicyclo[3.2.2]nonane skeleton and compounds 5 8 also have novel structures possessing a bicyclo[3.2.1]octane skeleton. The anti platelet-activating factor (PAF) activity of these compounds was tested by measurement of inhibition activity against acetyl transferase to lyso-PAF. PMID- 10866145 TI - Four new saponins from the root bark of Aralia elata. AB - Four new saponins, 3-O-[beta-D-glucopyranosyl(1-->3)-alpha-L-arabinopyranosyl] 16a lpha-hydroxyoleanolic acid 28-O-beta-D-glucopyranosyl ester (called aralia saponin I), 3-O-[beta-D-glucopyranosyl(1-->3)-alpha-L-arabinopyranosyl]-16a lpha hydroxyhederagenin 28-O-beta-D-glucopyranosyl ester (aralia-saponin II), 3-O [beta-D-glucopyranosyl(1-->3)-beta-D-glucopyranosyl(1-->3)-alpha-L-+ ++arabinopyranosyl]-16alpha-hydroxyoleanolic acid 28-O-beta-D-glucopyranosyl ester (aralia-saponin III), 3-O-[beta-D-glucopyranosyl(1-->3)-beta-D gucopyranosyl(1-->3)-beta -D-glucucopyranosyl]-16alpha-hydroxyoleanolic acid 28-O beta-D-glucopyranosyl ester (aralia-saponin IV), were isolated from the root bark of Aralia elata (Miq.) Seem., together with nineteen known compounds including glycosides of (20S)-protopanaxadiol and (20S)-protopanaxatriol. Their structures were determined on the basis of chemical and spectroscopy methods. PMID- 10866146 TI - Quantitative structure-activity relationship study of N-(3-oxo-3,4-dihydro-2H benzo[1,4]thiazine-6-carbonyl)guanidines as potent Na/H exchange inhibitors. AB - We have previously reported that N-(4-isopropyl-2,2-dimethyl-3-oxo-3,4-dihydro-2H benzo[1,4]oxazine-6-car bonyl)guanidine (4b) methanesulfonate salt (KB-R9032) is a potent and highly water-soluble Na/H exchange inhibitor. In a series of studies on Na/H exchange inhibitors, we designed and synthesized N-(3-oxo-3,4-dihydro-2H benzo[1,4]thiazine-6-carbonyl)guanidines (5) as more potent inhibitors with high water-solubility. The design strategy for 5 was based on a quantitative structure activity relationship (QSAR) study, involving the proportional relationship between the biological activity and hydrophobicity of the ring structure of compounds 4. As expected, compounds 5 showed more potent activity than 4. It was found by using the QSAR analysis that 5 were about five-fold more potent than 4. The increase in potency of compounds 5 well agreed with our previous QSAR analysis result. The most potent derivative was the methanesulfonate salt 5d of the 4-isopropyl derivative (IC50=0.0091 microM). And in addition to the in vitro study, 5d showed significant protective activity against a rat acute myocardial infraction model. PMID- 10866147 TI - Preparation and characterization of hyaluronate-hydroxyethyl acrylate blend hydrogel for controlled release device. AB - Hyaluronate-hydroxyethyl acrylate blend hydrogels were investigated as matrices for controlled release devices. Glycidyl methacrylate (GMA) derivatized HA (GMA HA) was synthesized by coupling of GMA to HA in the presence of a suitable catalyst. These hydrogels were prepared by a free radical copolymerization of GMA HA and hydroxyethyl acrylate. The water content of these hydrogels at equilibrium swelling in water (Ww) was 0.978+/-0.0073 (n=18); however, these hydrogel was mechanically tough and could be used as disk shape. The hydrogels swelling were found to depend on ionic strength and pH. The dried hydrogels quickly regained their original condition in water, and they swelled to more than 90% of its initial water contents after 30 min. This swelling-deswelling behavior was reproducible. The release of chlorpromazine HCl as a model cationic drug from the gels was suppressed significantly in water. The release increased with increasing the ionic strength and decreasing pH of bulk solutions. PMID- 10866148 TI - Synthetic studies of an 18-membered antitumor macrolide, tedanolide. 6. Synthesis of a key intermediate via a highly efficient macrolactonization of computer-aid designed seco-acid. PMID- 10866149 TI - Synthesis of arnottin I through a palladium-mediated aryl-aryl coupling reaction. AB - 6H-Dibenzo[b,d]pyran-6-one, 6H-benzo[d]naphtho[1,2-b]pyran-6-one, and their derivatives were prepared via the palladium mediated aryl-aryl coupling reaction of aryl ortho-halobenzoate. The short step synthesis of arnottin I(1) was achieved by this method. PMID- 10866150 TI - Preparation of new nitrogen-bridged heterocycles. 49. A new access to thieno[3,4 b]indolizine derivatives. AB - The title compounds, together with 3-vinylindolizine-1-carbonitriles (4-56%), were prepared in 1-18% yields from the S-alkylation of pyridinium 1-[3 ethoxycarbonyl-1-[cyanomethylthio(thiocarbonyl)]]all ylides with alkyl halides, followed by treatment of the resulting pyridinium salts with a base and then a dehydrogenating agent. In several reactions of pyridinium salts obtained from reaction of the allylides with benzylic halides, trace amounts of bis[1-cyano-9 (ethoxycarbonyl)thieno[3,4-b]indolizin-3-yl] disulfides with an interesting superimposed structure were also isolated. The structures of these compounds were confirmed by X-ray analyses. PMID- 10866151 TI - Salicylic acid derivatives form stable complexes with scandium(III) ion in aqueous solution. AB - The stability constants of 5-nitrosalicylic acid (5-NSA) and 5-sulfosalicylic acid (5-SSA) complexes of Sc(III) were determined by potentiomeric pH titration. ML and ML2 type first and second complexes were observed in the solutions of 5 NSA and 5-SSA with Sc(III) at 25 degrees C in I=0.1 M ionic medium. The stability constants of Sc(III)-5NSA and Sc(III)-5SSA systems were also investigated by spectrophotometry to determine the stoichiometries of the complexes formed in the reactions. Our results showed that Sc(III)-5SSA complexes are more stable than the Sc(III)-5NSA complexes in aqueous solutions. PMID- 10866152 TI - Micellar electrokinetic chromatography (MEKC) separation of furanonaphthoquinones from Tabebuia impetiginosa. AB - The separation of nine furanonaphthoquinones by micellar electrokinetic chromatography (MEKC) is described. The running electrolytes used in this method were 0.03 M sodium dodecyl sulphate (SDS) in 0.09 M borate buffer (pH 9) containing 10% methanol, with an applied voltage of 20 kV. Application of this technique in the determination of the main furanonaphthoquinones, 5-hydroxy-2-(1 hydroxyethyl)naphtho[2,3-b]furan-4,9-dione, 8-hydroxy-2-(1 hydroxyethyl)naphtho[2,3-b]furan-4,9-dione, and 2-(1-hydroxyethyl)naphtho[2,3 b]furan-4,9-dione, of Tabebuia impetiginosa is demonstrated in this paper. PMID- 10866153 TI - Conversion of quassinoids for enhancement of inhibitory effect against Epstein Barr virus early antigen activation. Introduction of lipophilic side chain and esterification of diosphenol. AB - Introduction of a senecioyl group into shinjulactones B and C, and esterification of the diosphenol moiety in brusatol and brucein A enhanced inhibitory effect against Epstein-Barr virus early antigen activation. PMID- 10866154 TI - Two 3,4-seco-lupane triterpenes from leaves of Acanthopanax divaricatus var. albeofructus. AB - Two 3,4-seco-lupane triterpenoids (1, 2) were isolated from the leaves of Acanthopanax divaricatus var. albeofructus (Araliaceae). Based on the spectroscopic data, the chemical structures of 1 and 2 were characterized as 24 hydroxychiisanogenin and 22alpha-hydroxychiisanogenin, respectively. 1 was a new compound and 2 was isolated for the first time from the natural source, although it had been obtained as an enzymatically hydrolyzed artifact from 22alpha hydroxychiisanoside. PMID- 10866155 TI - The '2+1' construction of homooxacalix[3]arenes possessing different substituents on their upper rims. AB - Several homooxacalix[3]arenes possessing different substituents on their upper rims were synthesized in yields of 7-20% by a condensation reaction between the p substituted-phenol dimer and monomer under acidic high-dilution conditions. PMID- 10866156 TI - Alpha-bromo-alpha,alpha-difluoroallyl derivatives as synthetic intermediate: nucleophilic substitution of alpha-bromo-alpha,alpha-difluoroallyl derivatives in the presence of palladium catalysts. AB - The palladium catalyzed nucleophilic substitution of alpha-bromo-alpha,alpha difluoroallyl derivatives turned out to be an efficient method for the preparation of several fluorinated organic molecules. Several soft carbon nucleophiles regioselectively reacted with 3-bromo-3,3-difluoropropene (BDFP) to give the 3-substituted 1,1-difluoroalkenes. Phenylzinc chloride and tributylphenyltin afforded 1-fluoro-1,3-diphenylpropene. The radical bromination of 3-substituted 1,1-difluoroalkenes provided 1-substituted BDFPs, and a 1 substituted BDFP reacted with carbon nucleophiles to give 1,3-disubstituted 3,3 difluoroalkenes. For the reaction of nitrogen nucleophiles with BDFP, an amine and the sodium salts of the carbamates reacted with BDFP at the gamma-position. However, the sodium salts of the sulfoneamide predominantly attacked at the alpha position. PMID- 10866157 TI - Saponins of plants of Panax species collected in Central Nepal, and their chemotaxonomical significance. III. AB - Panax pseudo-ginseng subsp. pseudo-ginseng has a carrot like root with a small rhizome. It was shown that the saponin composition of roots and rhizomes of this subspecies collected in Tibet and China was extremely poor. From the roots and rhizomes collected in Central Nepal, (specimen-PNct), only a small amount of an oleanolic acid saponin, beta-D-glucopyranosyl-oleanolate (2) was isolated together with a polyacetylene-alcohol, panaxynol (3). In another specimen (specimen-PNs), also collected in Central Nepal, two oleanolic acid saponins, stipleanoside R2 (4) and chikusetsusaponin IV (5) were detected. No dammarane saponin was identified in either specimen. P. pseudo-ginseng subsp. himalaicus (Subsp-H) has a big rhizome with a small round root. From rhizomes and roots of this subsp. collected in Central Nepal (specimen-HNct), a fairly large amount of dammarane saponins, ginsenosides-Rb1 (6), -Rd (7), -Re (9) and -Rg1 (10), gypenoside XVII (8), notoginsenoside-R1 (11), majonoside-R2 (12) and pseudo ginsenoside-F11 (13) were isolated, while no oleanane saponin (oleanolic acid saponin) was identified in this subsp. Based on the present and previous studies, medicinal evaluation and chemogeographical correlation of Himalayan Panax spp. are discussed. PMID- 10866158 TI - Reactivity of an o-indoloquinone to 1- and 2-azadienes. AB - The cycloaddition reactions of o-indoloquinone 4 to azadienes are described. With 1-azadiene 2, quinone 4 works as a dienophile to give the directly aromatized cycloadduct 6. In contrast, when 2-azadiene 3 is used, the cycloaddition occurs with CO-4, indicating that this system functions as a heterodienophile. PMID- 10866159 TI - Debates over credit for the annotation of genomes. PMID- 10866161 TI - China warned of AIDS 'disaster'. PMID- 10866160 TI - Relations thaw between genome rivals as finish line draws near. PMID- 10866162 TI - Canada unifies medical research community. PMID- 10866163 TI - Miniature antennas will eavesdrop on the Universe. PMID- 10866164 TI - European centres rebuffed in infrastructure funding bid. PMID- 10866165 TI - Spanish postgrads push for better employment rights PMID- 10866166 TI - NIH panel may increase gene-trial scrutiny... PMID- 10866167 TI - As Europe's 'excessive secrecy' is deplored. PMID- 10866169 TI - US climate report underlines local impacts of warming. PMID- 10866168 TI - UK researchers call for limits on animal experiment 'red tape'. PMID- 10866170 TI - Los Alamos 'loses' key weapons data PMID- 10866171 TI - One for all--and all for one PMID- 10866172 TI - Nanotech thinks big. PMID- 10866173 TI - 'Underachieving' centre has not only struck gold but made good use of it. PMID- 10866175 TI - Setbacks don't dampen the energy of US physics PMID- 10866174 TI - Jumping the gun on mouse gene expression. PMID- 10866176 TI - How the GM industry writes its own rules. PMID- 10866177 TI - An unholy alliance. The Nazis showed that 'politically responsible' science risks losing its soul. PMID- 10866178 TI - Another green world. There's more to industrial waste than chimneys and slag heaps PMID- 10866179 TI - Moon, tides and climate PMID- 10866181 TI - Smart polymer solutions PMID- 10866180 TI - Greater lifetime expectations. PMID- 10866182 TI - Neurobiology. Nervous engineering. PMID- 10866183 TI - Mathematics. Curves and numbers PMID- 10866184 TI - GTPase traffic control. PMID- 10866185 TI - Survival of the Irish elk into the Holocene. PMID- 10866186 TI - Control of free calcium in plant cell nuclei. PMID- 10866187 TI - Risky immortalization by telomerase. PMID- 10866188 TI - Do cockroaches 'know' about fluid dynamics? PMID- 10866189 TI - The evolutionarily conserved BMP-binding protein Twisted gastrulation promotes BMP signalling. AB - Dorsal-ventral patterning in vertebrate and Drosophila embryos requires a conserved system of extracellular proteins to generate a positional information gradient. The components involved include bone morphogenetic proteins (BMP/Dpp), a BMP antagonist (Chordin/Short gastrulation; Chd/Sog) and a secreted metalloproteinase (Xolloid/Tolloid) that cleaves Chd/Sog. Here we describe Xenopus Twisted gastrulation (xTsg), another member of this signalling pathway. xTsg is expressed ventrally as part of the BMP-4 synexpression group and encodes a secreted BMP-binding protein that is a BMP signalling agonist. The data suggest a molecular mechanism by which xTsg dislodges latent BMPs bound to Chordin BMP binding fragments generated by Xolloid cleavage, providing a permissive signal that allows high BMP signalling in the embryo. Drosophila Tsg also binds BMPs and is expressed dorsally, supporting the proposal that the dorsal-ventral axis was inverted in the course of animal evolution. PMID- 10866190 TI - Kondo effect in an integer-spin quantum dot AB - The Kondo effect--a many-body phenomenon in condensed-matter physics involving the interaction between a localized spin and free electrons--was discovered in metals containing small amounts of magnetic impurities, although it is now recognized to be of fundamental importance in a wide class of correlated electron systems. In fabricated structures, the control of single, localized spins is of technological relevance for nanoscale electronics. Experiments have already demonstrated artificial realizations of isolated magnetic impurities at metallic surfaces, nanoscale magnets, controlled transitions between two-electron singlet and triplet states, and a tunable Kondo effect in semiconductor quantum dots. Here we report an unexpected Kondo effect in a few-electron quantum dot containing singlet and triplet spin states, whose energy difference can be tuned with a magnetic field. We observe the effect for an even number of electrons, when the singlet and triplet states are degenerate. The characteristic energy scale is much larger than in the ordinary spin-1/2 case. PMID- 10866191 TI - Direct observation of the alignment of ferromagnetic spins by antiferromagnetic spins AB - The arrangement of spins at interfaces in a layered magnetic material often has an important effect on the properties of the material. One example of this is the directional coupling between the spins in an antiferromagnet and those in an adjacent ferromagnet, an effect first discovered in 1956 and referred to as exchange bias. Because of its technological importance for the development of advanced devices such as magnetic read heads and magnetic memory cells, this phenomenon has received much attention. Despite extensive studies, however, exchange bias is still poorly understood, largely due to the lack of techniques capable of providing detailed information about the arrangement of magnetic moments near interfaces. Here we present polarization-dependent X-ray magnetic dichroism spectro-microscopy that reveals the micromagnetic structure on both sides of a ferromagnetic-antiferromagnetic interface. Images of thin ferromagnetic Co films grown on antiferromagnetic LaFeO3 show a direct link between the arrangement of spins in each material. Remanent hysteresis loops, recorded for individual ferromagnetic domains, show a local exchange bias. Our results imply that the alignment of the ferromagnetic spins is determined, domain by domain, by the spin directions in the underlying antiferromagnetic layer. PMID- 10866192 TI - Reversible electromechanical characteristics of carbon nanotubes under local probe manipulation AB - The effects of mechanical deformation on the electrical properties of carbon nanotubes are of interest given the practical potential of nanotubes in electromechanical devices, and they have been studied using both theoretical and experimental approaches. One recent experiment used the tip of an atomic force microscope (AFM) to manipulate multi-walled nanotubes, revealing that changes in the sample resistance were small unless the nanotubes fractured or the metal-tube contacts were perturbed. But it remains unclear how mechanical deformation affects the intrinsic electrical properties of nanotubes. Here we report an experimental and theoretical elucidation of the electromechanical characteristics of individual single-walled carbon nanotubes (SWNTs) under local-probe manipulation. We use AFM tips to deflect suspended SWNTs reversibly, without changing the contact resistance; in situ electrical measurements reveal that the conductance of an SWNT sample can be reduced by two orders of magnitude when deformed by an AFM tip. Our tight-binding simulations indicate that this effect is owing to the formation of local sp3 bonds caused by the mechanical pushing action of the tip. PMID- 10866193 TI - Controlling droplet deposition with polymer additives AB - Controlling the impact of drops onto solid surfaces is important for a wide variey of coating and deposition processes--for example, the treatment of plants with herbicides and pesticides requires precise targeting in order to meet stringent toxicological regulations. However, the outer wax-like layer of the leaves is a non-wetting substrate that causes sprayed droplets to rebound; often less than 50% of the initial spray is retained by the plant. Although the impact and subsequent retraction of non-wetting aqueous drops on a hydrophobic surface have been the subjects of extensive experimental and theoretical work, non newtonian rheological effects have not been considered in any detail. Here we report that, by adding very small amounts of a flexible polymer to the aqueous phase, we can inhibit droplet rebound on a hydrophobic surface and markedly improve deposition without significantly altering the shear viscosity of the solutions. Our results can be understood by taking into account the non-newtonian elongational viscosity, which provides a large resistance to drop retraction after impact, thereby suppressing droplet rebound. PMID- 10866194 TI - Significant dissipation of tidal energy in the deep ocean inferred from satellite altimeter data AB - How and where the ocean tides dissipate their energy are long-standing questions that have consequences ranging from the history of the Moon to the mixing of the oceans. Historically, the principal sink of tidal energy has been thought to be bottom friction in shallow seas. There has long been suggestive evidence, however, that tidal dissipation also occurs in the open ocean through the scattering by ocean-bottom topography of surface tides into internal waves, but estimates of the magnitude of this possible sink have varied widely. Here we use satellite altimeter data from Topex/Poseidon to map empirically the tidal energy dissipation. We show that approximately 10(12) watts--that is, 1 TW, representing 25-30% of the total dissipation--occurs in the deep ocean, generally near areas of rough topography. Of the estimated 2 TW of mixing energy required to maintain the large-scale thermohaline circulation of the ocean, one-half could therefore be provided by the tides, with the other half coming from action on the surface of the ocean. PMID- 10866195 TI - A larger pool of ozone-forming carbon compounds in urban atmospheres AB - Volatile organic compounds play a central role in the processes that generate both urban photochemical smog and tropospheric ozone. For successful and accurate prediction of these pollution episodes, identification of the dominant reactive species within the volatile organic carbon pool is needed. At present, lack of resolution inherent in single-column chromatographic analysis limits such a detailed chemical characterization of the complex urban atmosphere. Here we present an improved method of peak deconvolution from double-column (orthogonal) gas chromatography. This has enabled us to isolate and classify more than 500 chemical species of volatile organic compounds in urban air, including over 100 multi-substituted monoaromatic and volatile oxygenated hydrocarbons. We suggest that previous assessments of reactive carbon species may therefore have underestimated the contribution made by volatile organic compounds to urban pollution, particularly for compounds with more than six carbon atoms. Incorporating these species in predictive models should greatly improve our understanding of photochemical ozone yields and the formation of harmful secondary organic aerosols. PMID- 10866196 TI - Re-Os isotopic evidence for a lower crustal origin of massif-type anorthosites AB - Massif-type anorthosites are large igneous complexes of Proterozoic age. They are almost monomineralic, representing vast accumulations of plagioclase with subordinate pyroxene or olivine and Fe-Ti oxides--the 930-Myr-old Rogaland anorthosite province in southwest Norway represents one of the youngest known expressions of such magmatism. The source of the magma and geodynamic setting of massif-type anorthosites remain long-standing controversies in Precambrian geology, with no consensus existing as to the nature of the parental magmas or whether these magmas primarily originate in the Earth's mantle or crust. At present, massif-type anorthosites are believed to have crystallized from either crustally contaminated mantle-derived melts that have fractionated olivine and pyroxenes at depth or primary aluminous gabbroic to jotunitic melts derived from the lower continental crust. Here we report rhenium and osmium isotopic data from the Rogaland anorthosite province that strongly support a lower crustal source for the parental magmas. There is no evidence of significantly older crust in southwest Scandinavia and models invoking crustal contamination of mantle-derived magmas fail to account for the isotopic data from the Rogaland province. Initial osmium and neodymium isotopic values testify to the melting of mafic source rocks in the lower crust with an age of 1,400-1,550 Myr. PMID- 10866197 TI - Unrelated helpers in a social insect. AB - High-resolution genetic markers have revolutionized our understanding of vertebrate mating systems, but have so far yielded few comparable surprises about kinship in social insects. Here we use microsatellite markers to reveal an unexpected and unique social system in what is probably the best-studied social wasp, Polistes dominulus. Social insect colonies are nearly always composed of close relatives; therefore, non-reproductive helping behaviour can be favoured by kin selection, because the helpers aid reproductives who share their genes. In P. dominulus, however, 35% of foundress nestmates are unrelated and gain no such advantage. The P. dominulus system is unlike all other cases of unrelated social insects, because one individual has nearly complete reproductive dominance over subordinates who could have chosen other reproductive options. The only significant advantage that subordinates obtain is a chance at later reproduction, particularly if the queen dies. Thus, P. dominulus societies are functionally unlike other social insects, but similar to certain vertebrate societies, in which the unrelated helpers gain through inheritance of a territory or a mate. PMID- 10866198 TI - Female feral fowl eject sperm of subdominant males. AB - Paternity is often determined by competition between the ejaculates of different males. Males can also use particular behaviours or structures to manipulate how females use sperm. However, the ability of females to bias sperm utilization in favour of preferred males independently of male manipulation has not been demonstrated. Females are predicted to respond differentially to the sperm of different males when the reproductive interests of the sexes differ and when females are coerced into copulating. Here we show that in female feral fowl most copulations are coerced, and that females consistently bias sperm retention in favour of the preferred male phenotype. Females prefer to copulate with dominant males, but when sexually coerced by subordinate males, they manipulate the behaviour of dominant males to reduce the likelihood of insemination. If this fails, females differentially eject ejaculates according to male status in the absence of any male manipulation and preferentially retain the sperm of dominant males. PMID- 10866199 TI - A universal pattern of mortality decline in the G7 countries. AB - Human lifespan has increased enormously this century. But we remain uncertain about the forces that reduce mortality, and about the cost implications of ageing populations and their associated social burden. The poor understanding of the factors driving mortality decline, and the difficulty of forecasting mortality are due in part to the pronounced irregularity of annual to decadal mortality change. Here we examine mortality over five decades in the G7 countries (Canada, France, Germany, Italy, Japan, UK, US). In every country over this period, mortality at each age has declined exponentially at a roughly constant rate. This trend places a constraint on any theory of society-driven mortality decline, and provides a basis for stochastic mortality forecasting. We find that median forecasts of life expectancy are substantially larger than in existing official forecasts. In terms of the costs of ageing, we forecast values of the dependency ratio (that is, the ratio of people over 65 to working people) in 2050 that are between 6% (UK) and 40% (Japan) higher than official forecasts. PMID- 10866200 TI - Ultrasensitive pheromone detection by mammalian vomeronasal neurons. AB - The vomeronasal organ (VNO) is a chemoreceptive organ that is thought to transduce pheromones into electrical responses that regulate sexual, hormonal and reproductive function in mammals. The characteristics of pheromone signal detection by vomeronasal neurons remain unclear. Here we use a mouse VNO slice preparation to show that six putative pheromones evoke excitatory responses in single vomeronasal neurons, leading to action potential generation and elevated calcium entry. The detection threshold for some of these chemicals is remarkably low, near 10(-11) M, placing these neurons among the most sensitive chemodetectors in mammals. Using confocal calcium imaging, we map the epithelial representation of the pheromones to show that each of the ligands activates a unique, nonoverlapping subset of vomeronasal neurons located in apical zones of the epithelium. These neurons show highly selective tuning properties and their tuning curves do not broaden with increasing concentrations of ligand, unlike those of receptor neurons in the main olfactory epithelium. These findings provide a basis for understanding chemical signals that regulate mammalian communication and sexual behaviour. PMID- 10866201 TI - Excessive placental secretion of neurokinin B during the third trimester causes pre-eclampsia. AB - Pre-eclampsia is a principal cause of maternal morbidity and mortality, affecting 5-10% of first pregnancies worldwide. Manifestations include increased blood pressure, proteinuria, coagulopathy and peripheral and cerebral oedema. Although the aetiology and pathogenesis remain to be elucidated, the placenta is undoubtedly involved, as termination of pregnancy eradicates the disease. Here we have cloned a complementary DNA from human placental messenger RNA encoding a precursor protein of 121 amino acids which gives rise to a mature peptide identical to the neuropeptide neurokinin B (NKB) of other mammalian species. In female rats, concentrations of NKB several-fold above that of an animal 20 days into pregnancy caused substantial pressor activity. In human pregnancy, the expression of NKB was confined to the outer syncytiotrophoblast of the placenta, significant concentrations of NKB could be detected in plasma as early as week 9, and plasma concentrations of NKB were grossly elevated in pregnancy-induced hypertension and pre-eclampsia. We conclude that elevated levels of NKB in early pregnancy may be an indicator of hypertension and pre-eclampsia, and that treatment with certain neurokinin receptor antagonists may be useful in alleviating the symptoms. PMID- 10866202 TI - The gamma-subunit of the coatomer complex binds Cdc42 to mediate transformation. AB - The Ras-related GTP-binding protein Cdc42 is implicated in a variety of biological activities including the establishment of cell polarity in yeast, the regulation of cell morphology, motility and cell-cycle progression in mammalian cells and the induction of malignant transformation. We identified a Cdc42 mutant (Cdc42F28L) which binds GTP in the absence of a guanine nucleotide exchange factor, but still hydrolyses GTP with a turnover number identical to that for wild-type Cdc42. Expression of this mutant in NIH 3T3 fibroblasts causes cellular transformation, mimicking many of the characteristics of cells transformed by the Dbl oncoprotein, a known guanine nucleotide exchange factor for Cdc42. Here we searched for new Cdc42 targets in an effort to understand how Cdc42 mediates cellular transformation. We identified the gamma-subunit of the coatomer complex (gammaCOP) as a specific binding partner for activated Cdc42. The binding of Cdc42 to gammaCOP is essential for a transforming signal distinct from those elicited by Ras. PMID- 10866203 TI - Two-headed binding of a processive myosin to F-actin. AB - Myosins are motor proteins in cells. They move along actin by changing shape after making stereospecific interactions with the actin subunits. As these are arranged helically, a succession of steps will follow a helical path. However, if the myosin heads are long enough to span the actin helical repeat (approximately 36 nm), linear motion is possible. Muscle myosin (myosin II) heads are about 16 nm long, which is insufficient to span the repeat. Myosin V, however, has heads of about 31 nm that could span 36 nm and thus allow single two-headed molecules to transport cargo by walking straight. Here we use electron microscopy to show that while working, myosin V spans the helical repeat. The heads are mostly 13 actin subunits apart, with values of 11 or 15 also found. Typically the structure is polar and one head is curved, the other straighter. Single particle processing reveals the polarity of the underlying actin filament, showing that the curved head is the leading one. The shape of the leading head may correspond to the beginning of the working stroke of the motor. We also observe molecules attached by one head in this conformation. PMID- 10866204 TI - The lyase activity of the DNA repair protein beta-polymerase protects from DNA damage-induced cytotoxicity. AB - Small DNA lesions such as oxidized or alkylated bases are repaired by the base excision repair (BER) pathway. BER includes removal of the damaged base by a lesion-specific DNA glycosylase, strand scission by apurinic/apyrimidinic endonuclease, DNA resynthesis and ligation. BER may be further subdivided into DNA beta-polymerase (beta-pol)-dependent single-nucleotide repair and beta-pol dependent or -independent long patch repair subpathways. Two important enzymatic steps in mammalian single-nucleotide BER are contributed by beta-pol: DNA resynthesis of the repair patch and lyase removal of 5'-deoxyribose phosphate (dRP). Fibroblasts from beta-pol null mice are hypersensitive to mono-functional DNA-methylating agents, resulting in increases in chromosomal damage, apoptosis and necrotic cell death. Here we show that only the dRP lyase activity of beta pol is required to reverse methylating agent hypersensitivity in beta-pol null cells. These results indicate that removal of the dRP group is a pivotal step in BER in vivo. Persistence of the dRP moiety in DNA results in the hypersensitivity phenotype of beta-pol null cells and may signal downstream events such as apoptosis and necrotic cell death. PMID- 10866205 TI - Observations of light-induced structural changes of retinal within rhodopsin. AB - Photo-isomerization of the 11-cis retinal chromophore activates the mammalian light-receptor rhodopsin, a representative member of a major superfamily of transmembrane G-protein-coupled receptor proteins (GPCRs) responsible for many cell signal communication pathways. Although low-resolution (5 A) electron microscopy studies confirm a seven transmembrane helix bundle as a principal structural component of rhodopsin, the structure of the retinal within this helical bundle is not known in detail. Such information is essential for any theoretical or functional understanding of one of the fastest occurring photoactivation processes in nature, as well as the general mechanism behind GPCR activation. Here we determine the three-dimensional structure of 11-cis retinal bound to bovine rhodopsin in the ground state at atomic level using a new high resolution solid-state NMR method. Significant structural changes are observed in the retinal following activation by light to the photo-activated M(I) state of rhodopsin giving the all-trans isomer of the chromophore. These changes are linked directly to the activation of the receptor, providing an insight into the activation mechanism of this class of receptors at a molecular level. PMID- 10866206 TI - Atomically defined mechanism for proton transfer to a buried redox centre in a protein. AB - The basis of the chemiosmotic theory is that energy from light or respiration is used to generate a trans-membrane proton gradient. This is largely achieved by membrane-spanning enzymes known as 'proton pumps. There is intense interest in experiments which reveal, at the molecular level, how protons are drawn through proteins. Here we report the mechanism, at atomic resolution, for a single long range electron-coupled proton transfer. In Azotobacter vinelandii ferredoxin I, reduction of a buried iron-sulphur cluster draws in a solvent proton, whereas re oxidation is 'gated' by proton release to the solvent. Studies of this 'proton transferring module' by fast-scan protein film voltammetry, high-resolution crystallography, site-directed mutagenesis and molecular dynamics, reveal that proton transfer is exquisitely sensitive to the position and pK of a single amino acid. The proton is delivered through the protein matrix by rapid penetrative excursions of the side-chain carboxylate of a surface residue (Asp 15), whose pK shifts in response to the electrostatic charge on the iron-sulphur cluster. Our analysis defines the structural, dynamic and energetic requirements for proton courier groups in redox-driven proton-pumping enzymes. PMID- 10866207 TI - Exploring genome space. AB - The completion of entire genome sequences of many experimental organisms, and the promise that the human genome will be completed in the next year, find biology suddenly awash in genome-based data. Scientists are scrambling to develop new technologies that exploit genome data to ask entirely new kinds of questions about the complex nature of living cells. PMID- 10866208 TI - Protein function in the post-genomic era. AB - Faced with the avalanche of genomic sequences and data on messenger RNA expression, biological scientists are confronting a frightening prospect: piles of information but only flakes of knowledge. How can the thousands of sequences being determined and deposited, and the thousands of expression profiles being generated by the new array methods, be synthesized into useful knowledge? What form will this knowledge take? These are questions being addressed by scientists in the field known as 'functional genomics'. PMID- 10866209 TI - Genomics, gene expression and DNA arrays. AB - Experimental genomics in combination with the growing body of sequence information promise to revolutionize the way cells and cellular processes are studied. Information on genomic sequence can be used experimentally with high density DNA arrays that allow complex mixtures of RNA and DNA to be interrogated in a parallel and quantitative fashion. DNA arrays can be used for many different purposes, most prominently to measure levels of gene expression (messenger RNA abundance) for tens of thousands of genes simultaneously. Measurements of gene expression and other applications of arrays embody much of what is implied by the term 'genomics'; they are broad in scope, large in scale, and take advantage of all available sequence information for experimental design and data interpretation in pursuit of biological understanding. PMID- 10866210 TI - Proteomics to study genes and genomes. AB - Proteomics, the large-scale analysis of proteins, will contribute greatly to our understanding of gene function in the post-genomic era. Proteomics can be divided into three main areas: (1) protein micro-characterization for large-scale identification of proteins and their post-translational modifications; (2) 'differential display' proteomics for comparison of protein levels with potential application in a wide range of diseases; and (3) studies of protein-protein interactions using techniques such as mass spectrometry or the yeast two-hybrid system. Because it is often difficult to predict the function of a protein based on homology to other proteins or even their three-dimensional structure, determination of components of a protein complex or of a cellular structure is central in functional analysis. This aspect of proteomic studies is perhaps the area of greatest promise. After the revolution in molecular biology exemplified by the ease of cloning by DNA methods, proteomics will add to our understanding of the biochemistry of proteins, processes and pathways for years to come. PMID- 10866211 TI - Searching for genetic determinants in the new millennium. AB - Human genetics is now at a critical juncture. The molecular methods used successfully to identify the genes underlying rare mendelian syndromes are failing to find the numerous genes causing more common, familial, non-mendelian diseases. With the human genome sequence nearing completion, new opportunities are being presented for unravelling the complex genetic basis of non-mendelian disorders based on large-scale genome-wide studies. Considerable debate has arisen regarding the best approach to take. In this review I discuss these issues, together with suggestions for optimal post-genome strategies. PMID- 10866212 TI - Pharmacogenetics and the practice of medicine. AB - "If it were not for the great variability among individuals medicine might as well be a science and not an art." The thoughts of Sir William Osler in 1892 reflect the view of medicine over the past 100 years. The role of physicians in making the necessary judgements about the medicines that they prescribe is often referred to as an art, reflecting the lack of objective data available to make decisions that are tailored to individual patients. Just over a hundred years later we are on the verge of being able to identify inherited differences between individuals which can predict each patient's response to a medicine. This ability will have far-reaching benefits in the discovery, development and delivery of medicines. Sir William Osler, if he were alive today, would be re-considering his view of medicine as an art not a science. PMID- 10866213 TI - New biologically active rubiginones from Streptomyces sp. AB - Four new polyketides, named rubiginone D2 (2), 4-O-acetyl-rubiginone D2 (3), rubiginone H (6) and rubiginone I (7) were isolated from the cultures of Streptomyces sp. (strain Go N1/5). Their structures were established by a detailed spectroscopic analysis. The absolute configuration of 3 was determined by derivatization with chiral acids (Helmchen's method). The rubiginones inhibit the growth of some Gram-positive bacteria and are cytostatically active against different tumor cell lines. PMID- 10866214 TI - New Cdc25B tyrosine phosphatase inhibitors, nocardiones A and B, produced by Nocardia sp. TP-A0248: taxonomy, fermentation, isolation, structural elucidation and biological properties. AB - Strain TP-A0248 which produces two new Cdc25B tyrosine phosphatase inhibitors and also possessing antifungal activity, designated nocardiones A (1) and B (2), was considered to belong to the genus Nocardia on the basis of literature comparison of chemotaxonomic properties. The nocardiones were isolated by solvent extraction of fermentation broth of Nocardia sp. TP-A0248 and purified by the conventional column chromatography. Spectroscopic studies led to determination that 1 and 2 belong to a class compound of naphtho[1,2-b]furan-4,5-diones. Compound 1 inhibited the activity of Cdc25B, PTP1B and FAP-1 protein tyrosine phosphatases at a concentration of 10 microM. It also showed moderate in vitro antifungal and cytotoxic activity. PMID- 10866215 TI - SB-219383, a novel tyrosyl tRNA synthetase inhibitor from a Micromonospora sp. I. Fermentation, isolation and properties. AB - A novel, potent and selective inhibitor of bacterial tyrosyl tRNA synthetase, designated SB-219383 has been isolated from Micromonospora sp. NCIMB 40684. The fermentation, isolation and some properties are described, whilst the structure determination is described in the succeeding paper). SB-219383 showed competitive, inhibitory activity against a Staphylococcus tyrosyl tRNA synthetase, with an IC50 of <1 nM, and exhibited weak in vitro activity against some Streptococcus sp. PMID- 10866216 TI - SB-219383, a novel tyrosyl tRNA synthetase inhibitor from a Micromonospora sp. II. Structure determination. AB - A novel tyrosyl tRNA synthetase inhibitor, SB-219383, has been isolated from a Micromonospora sp. The structure of SB-219383 has been elucidated by a combination of derivatisation, spectroscopic and other analytical techniques and found to be N-(L-tyrosyl)-2-amino[1(S*),3(S*),4(S*),5(R*),8(R*)-2,4,5,8 tetrahydroxy -7-oxa-2-azabicyclo[3.2.1]oct-3-y1]acetic acid (1). PMID- 10866217 TI - SB-203207 and SB-203208, two novel isoleucyl tRNA synthetase inhibitors from a Streptomyces sp. I. Fermentation, isolation and properties. AB - Two novel inhibitors of isoleucyl tRNA synthetase designated SB-203207 and SB 203208 have been detected in the culture of a new Streptomyces species. The fermentation, isolation and some properties of the inhibitors are described. PMID- 10866218 TI - SB-203207 and SB-203208, two novel isoleucyl tRNA synthetase inhibitors from a Streptomyces sp. II. Structure determination. AB - Two novel isoleucyl tRNA synthetase inhibitors, SB-203207 and SB-203208 have been isolated from a Streptomyces sp. and found to be structurally related to altemicidin. Structures of SB-203207 and SB-203208 have been deduced by a combination of spectroscopic techniques, derivatisation, hydrolysis studies and found to be 4-(aminocarbonyl)-7-[[(2-amino-3-methylpentanoyl)aminosul phonyl]acetamido]-2,4a,5,6,7,7a-hexahydro-6-hydroxy-2-methyl-1H-2- pyrindine-7 carboxylic acid (1) and 4-(aminocarbonyl)-7-[[(2-amino-3-methyl pentanoyl) aminosulphonyl]acetamido]-2,4a,5,6,7,7a-hexahydro-6-(2- amino-3-phenylbutanoyl oxy)-2-methyl-1H-2-pyrindine-7-carboxylic acid (2), respectively. PMID- 10866219 TI - ABC transporter genes, kasKLM, responsible for self-resistance of a kasugamycin producer strain. AB - We previously reported that a 7.6-kb DNA fragment from Streptomyces kasugaensis M338-M1, a kasugamycin (KSM) producer, included KSM acetyltransferase gene (kac338) and some other genes possibly involved in KSM biosynthesis. As an extension of that study, a 10-kb SacI-KpnI DNA fragment, located approximately 5 15-kb upstream of kac33, was cloned and a 4.2-kb SacI-EcoRI fragment therefrom was sequenced, revealing one incomplete (designated ORF J) and three complete open reading frames (designated kasK, kasL and kasM). The coding frames of kasK, L and M overlap one another with terminator/initiator ATGA sequence. RT-PCR analysis of a DNA region including kasKLM indicated the presence of one transcript that is long enough to span the three genes. The kasK gene potentially encodes an ATP-binding protein of the ATP-binding cassette (ABC) transporter superfamily. Homology search for the deduced KasK protein shows similarity to other ABC transporters involved in self-resistance of a mithramycin and possibly doxorubicin producer strain. The kasL and kasM genes encode different integral membrane proteins, both having six putative transmembrane helices. An expression plasmid for kasKLM (pTV-KLM) was constructed and these genes were expressed in E. coli JM 109, which had been sensitive to KSM. The transformant acquired resistance to KSM, suggesting that KasK, L and M proteins as a set in S. kasugaensis M338-M1 pump out KSM to protect the producer from its toxic metabolite. PMID- 10866220 TI - A putative enolpyruvyl transferase gene involved in nikkomycin biosynthesis. AB - The nikO gene encoding a putative enolpyruvyl transferase has been identified within the Streptomyces tendae Tu901/8c nikkomycin gene cluster. nikO encodes a deduced protein of 471 amino acid residues which exhibits significant sequence similarity to UDP-N-acetylglucosamine enolpyruvyl transferase and 5-enol pyruvylshikimate 3-phosphate synthase from various origin. The nikO gene was inactivated by inserting a kanamycin resistance cassette; the mutant did not produce biologically active nikkomycins I, J, X, and Z nor the nucleoside moieties, nikkomycins C(x) and C(z), but accumulated the novel component RT 2.0. RT 2.0 has been isolated from culture filtrate and its structure was determined by using mass spectrometry and NMR analyses as ribofuranosyl-4-formyl-4 imidazolone which represents a novel nucleoside. The putative activity of the nikO gene product in nikkomycin biosynthesis will be discussed. PMID- 10866221 TI - Cloning and expression of a gene encoding N-glycosyltransferase (ngt) from Saccarothrix aerocolonigenes ATCC39243. AB - In the course of our bioconversion studies on the derivatives of an indolocarbazole, J-104303, Saccharothrix aerocolonigenes ATCC39243 was found to convert J-104303, which was added into the culture medium, to its glycosylated derivative, J-109384. In order to clone the gene having the ability to convert J 104303 to J-109384, a library of Saccharothrix aerocolonigenes ATCC39243 DNA fragments was constructed using Streptomyces lividans TK21 and pIJ702 as host strain and vector, respectively. By examining more than 5,000 transformants, one was found to convert J-104303 to J-109384. Sequence analysis of the inserted DNA fragment revealed an open reading frame with 1,245 base pairs, named ngt. The transformant containing this ngt gene was also found to introduce a D-glucose moiety into 6-N-methylarcyriaflavin C. Furthermore, when ngt was introduced into Streptomyces mobaraensis BA13793, a producer of J-104303, the resulting transformant produced J-109384 directly. PMID- 10866222 TI - Synthesis and biological evaluation of novel tricyclic carbapenems (trinems). AB - Synthesis of new tricyclic carbapenems (trinems) with a pyrrolidinyl moiety at the C-4 position of the tricyclic ring and their antimicrobial activities were studied. These trinems showed potent activities against Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA). Among them, (4R) [(S)-pyrrolidin-3-ylthiomethyl]trinem (14a) exhibited good activity against MRSA in vitro and in vivo. PMID- 10866223 TI - Cuevaenes A and B, new polyene carboxylic acids from Streptomyces sp. HKI 0180. PMID- 10866224 TI - Novel synthesis and structural elucidation of quinolone antibiotics PC-3 (SF2420B) and YM-30059. PMID- 10866225 TI - Structure-activity relationships study of pyripyropenes: reversal of cancer cell multidrug resistance. PMID- 10866226 TI - Synthesis and antiangiogenic activity of staurosporine derivatives. PMID- 10866227 TI - Novel synthesis of natural pseudo-aminosugars, (+)-valienamine and (+) validamine. PMID- 10866228 TI - Most env and gag subtype A HIV-1 viruses circulating in West and West Central Africa are similar to the prototype AG recombinant virus IBNG. AB - The genetic subtype was identified in gag and env of 219 HIV-1-positive samples collected in different African countries, 44 from Senegal, 55 from Cameroon, 82 from Gabon, and 38 from Djibouti. In total, 20 (9.1%) samples had discordant subtypes between gag and env, 6 of 44 (13.9%) in Senegal, 4 of 55 (7.2%) in Cameroon, 1 of 38 (2.6%) in Djibouti, and 10 of 82 (12.1%) in Gabon. Subtypes A and G were predominantly involved in the recombination events. Phylogenetic tree analysis of gag showed that an important number of the A sequences form a distinct subcluster with the AG-IBNG prototype strain (a complex A/G mosaic virus): 27 of 32 (84.3%) in Senegal, 12 of 17 (70.6%) in Nigeria, 24 of 39 (61.5%) in Cameroon, and 38 of 70 (54.3%) in Gabon. Full-length genome analysis of 3 and additional sequences in pol for 10 such strains confirmed that they have a similar complex A/G mosaic genomic structure. These data suggest that in West Africa, most probably between 60% and 84% of the subtype A viruses are recombinant AG-IBNG viruses. This finding has potential implications on future vaccine, diagnostic, and treatment strategies. The actual and future role of these viruses in the global pandemic must be monitored in all new molecular epidemiologic studies, a discrimination between subtype A and AG-IBNG-like viruses is necessary. PMID- 10866229 TI - Fluctuations in HIV-1 viral load are correlated to CD4+ T-lymphocyte count during the natural course of infection. AB - Viral load fluctuates during the natural course of asymptomatic HIV-1 infection. It is often assumed that these fluctuations are random around a set point or underlying growth trend. Using longitudinal data, we tested whether fluctuations in viral load can be better explained by changes in CD4+ T-cell count than by a set point or trend of exponential growth. The correspondence between viral load and CD4+ T-cell count could be described by a simple mathematical relation. Using a bootstrapping approach, the hypothesis that viral load fluctuations are random around a set point was rejected with p < .00005. The hypothesis that viral load fluctuations are random around a trend of exponential growth was rejected with p < .005. Viral load data was explained better by changes in CD4+ T-cell counts than by a set point or by a trend of exponential growth. The implications of this finding for improved prognostication are discussed. PMID- 10866230 TI - HAART is neuroprophylactic in HIV-1 infection. AB - BACKGROUND: To find out about the prophylactic value of antiretroviral therapy on HIV-1-associated subclinical and clinical psychomotor slowing as one marker of HIV-1-associated CNS disease. METHODS: Prospective study with regular clinical and neurophysiologic examination every three months of 1482 consecutive HIV-1 seropositive and AIDS patients seen at our department till June 30, 1999. RESULTS: Antiretroviral therapy has a significant prophylactic value over an individual observation period of ten years with regard to the first, potentially transient manifestation of HIV-1-associated subclinical psychomotor slowing and with regard to the clinical manifestation of motor signs. However, a subgroup of patients is characterized through a second, more sustained manifestation of subclinical psychomotor slowing which cannot be prevented by any type of currently available antiretroviral therapy. CONCLUSIONS: These findings suggest the existence of different pathomechanisms underlying HIV-1-associated brain disease which may in part be effectively prevented, but which in part also escape all antiretroviral treatment strategies in use today. PMID- 10866231 TI - Predictors of self-reported adherence and plasma HIV concentrations in patients on multidrug antiretroviral regimens. AB - BACKGROUND: Adherence to prescribed medications is a central feature of good clinical HIV care, but little is known about the factors associated with multidrug antiretroviral adherence, or about how such adherence is related to plasma HIV suppression. METHODS: We collected data from 133 HIV-infected adults receiving antiretroviral therapy. Study subjects completed customized adherence self-report instruments and provided blood samples to measure plasma HIV-1 RNA concentrations and CD4+ lymphocyte counts. Regression models were used to determine the independent predictors of antiretroviral adherence and plasma HIV concentration, and the relationships between the two. RESULTS: Adherence was poor (average, <80% antiretrovirals/day) in 28% (95% confidence interval [CI], 20% 36%), fair (80%-99% per day) in 23% (95% CI, 15%-30%), and excellent (100% per day) in 50% (95% CI, 41%-58%) of study subjects. Mean decreases in HIV-1 concentration from highest-ever levels were 1.3, 1.6, and 2.0 log10 copies/ml in these three groups, respectively (chi2; p < .02). Two-stage least squares regression demonstrated a -1.3 log difference in viral load associated with each category improvement in adherence. In multivariate models, confidence in medication-taking ability, or perceived self-efficacy, and convenience of the medication regimen, or "fit" with routine and daily activities, were also associated with greater medication adherence (odds ratios [OR] 5.3; 95% CI, 2.4 11.8, and 9.0; 95% CI, 1.8-45.3, respectively). The latter was also independently associated with a lower plasma HIV concentration (p < .02). CONCLUSIONS: Nonadherence to combination antiretroviral medications is common and is associated with increased levels of plasma HIV. Programs and clinical efforts to improve medication taking should strive to integrate medications better into patients' daily routines and to improve patients' confidence in their ability to take medications correctly. PMID- 10866232 TI - Impact of medical and nonmedical factors on physician decision making for HIV/AIDS antiretroviral treatment. AB - To examine influences of medical factors (e.g., viral load) and nonmedical factors (e.g., patient characteristics) on treatment decisions for highly active antiretroviral therapy (HAART), we sent a survey to a random sample of 995 infectious disease physicians who treat patients with HIV/AIDS in the United States in August, 1998. The response rate was 53%. Respondents were asked to report their current practices with respect to antiretroviral treatment and the extent to which each of three medical and 17 nonmedical factors would influence them for or against prescribing HAART to a hypothetical HIV-positive patient. Most reported initiating HAART with findings of low CD4+ cell counts and high viral loads, and weighing CD4+ cell counts, viral load, and opportunistic infection heavily in their decisions to prescribe HAART. Patients' prior history of poor adherence was weighed very much against initiating HAART. Patient homelessness, heavy alcohol use, injection drug use, and prior psychiatric hospitalization were cited by most physicians as weighing against HAART initiation. Thus, most physicians in this sample follow guidelines for the use of HAART, and nonmedical factors related to patients' life situations are weighed as heavily as disease severity in treatment decisions. As HIV increasingly becomes a disease associated with economic disadvantage and other social health problems, it will be essential to develop interventions and care support systems to enable patients experiencing these problems to benefit from HIV treatment advances. PMID- 10866233 TI - High HIV incidence and prevalence among young women in rural South Africa: developing a cohort for intervention trials. AB - OBJECTIVE: To measure prevalence and model incidence of HIV infection. SETTING: 2013 consecutive pregnant women attending public sector antenatal clinics in 1997 in Hlabisa health district, South Africa. Historical seroprevalence data, 1992 1995. METHODS: Serum remaining from syphilis testing was tested anonymously for antibodies to HIV to determine seroprevalence. Two models, allowing for differential mortality between HIV-positive and HIV-negative people, were used. The first used serial seroprevalence data to estimate trends in annual incidence. The second, a maximum likelihood model, took account of changing force of infection and age-dependent risk of infection, to estimate age-specific HIV incidence in 1997. Multiple logistic regression provided adjusted odds ratios (OR) for risk factors for prevalent HIV infection. RESULTS: Estimated annual HIV incidence increased from 4% in 1992/1993 to 10% in 1996/1997. In 1997, highest age-specific incidence was 16% among women aged between 20 and 24 years. In 1997, overall prevalence was 26% (95% confidence interval [CI], 24%-28%) and at 34% was highest among women aged between 20 and 24 years. Young age (<30 years; odds ratio [OR], 2.1; p = .001), unmarried status (OR 2.2; p = .001) and living in less remote parts of the district (OR 1.5; p = .002) were associated with HIV prevalence in univariate analysis. Associations were less strong in multivariate analysis. Partner's migration status was not associated with HIV infection. Substantial heterogeneity of HIV prevalence by clinic was observed (range 17% 31%; test for trend, p = .001). CONCLUSIONS: This community is experiencing an explosive HIV epidemic. Young, single women in the more developed parts of the district would form an appropriate cohort to test, and benefit from, interventions such as vaginal microbicides and HIV vaccines. PMID- 10866234 TI - Monitoring of HIV-1 infection prevalence and trends in the general population using pregnant women as a sentinel population: 9 years experience from the Kagera region of Tanzania. AB - In the Kagera region of Tanzania, a population-based study was initiated in 1987 followed by the establishment of antenatal-clinic-based sentinel surveillance system in the town of Bukoba in 1990. Repeat studies in both populations in Bukoba in 1993 and 1996 made it possible to study the dynamics of HIV infection prevalence and incidence in the area. This study aims at comparing the findings from this sentinel surveillance system with those of cross-sectional studies in the general population to assess its validity in estimating HIV prevalence and their trends in the general population. A multistage cluster sampling technique was used in the population-based studies whereas the antenatal-clinic-based population was obtained by consecutively recruiting antenatal care attenders coming for the first time during a given pregnancy. Antibodies against HIV infection were tested using two independent enzyme-linked immunosorbent assay (ELISA) antibody detection tests. Unlinked anonymous testing strategy was adopted for the sentinel population. Age-adjusted prevalence among antenatal care attenders decreased from 22.4% (95% confidence interval [CI], 20.6-25.2) in 1990 to 16.1% (95% CI, 15.9-18.8) in 1993 and further to 13.7% (95% CI, 11.8-14.3) in 1996. These results closely resemble those of the general population of adult women in the clinic's catchment area (the town of Bukoba) where the age-adjusted prevalence of 29.1% (95% CI, 24.4-34.6) in 1987 showed a decrease in the studies in 1993 18.7% (95% CI, 15.1-23.0) and in 1996 14.9% (95% CI, 12.0-17.1). The study indicates that general population trend estimates can be generated using sentinel surveillance data based on pregnant women visiting an antenatal clinic for the first time during a given pregnancy. The benefits of using this group outweigh its limitations that are brought about by possible selection bias. Continued surveillance of the epidemic based on antenatal care patients as a sentinel population is therefore recommended. PMID- 10866235 TI - Case-control study of risk factors for incident HIV infection in rural Uganda. AB - OBJECTIVE: To identify risk factors associated with HIV incidence in a rural Ugandan population. DESIGN: Case-control study. METHODS: Men and women who seroconverted between 1990 and 1997 (cases) and seronegative subjects (controls) were drawn from a general population cohort of approximately 5000 adults in rural, southwestern Uganda. Information on risk factors was ascertained through a detailed interview and physical examination by clinicians who were blind to the study subjects' HIV status. All patients were interviewed within 2 years of their estimated date of seroconversion. RESULTS: Data were available on 130 men (37 cases, 93 controls) and 133 women (46 cases, 87 controls). There was a significantly higher risk of infection in men (odds ratio [OR], 6.51; 95% confidence interval [CI], 1.06-39.84) and women (OR, 4.75; 95% CI, 1.26-17.9) who were unmarried and in a steady relationship, and in men who were divorced, separated, or widowed (OR, 4.33; 95% CI, 1.32-14.25) compared with those who were married. There was a significantly higher risk of HIV infection in men (OR, 3.78; 95% CI, 1.20-11.93) and women (OR, 20.78; 95% CI, 2.94-141.2) who reported > or =5 lifetime sexual partners compared with those who reported at most 1 partner. For men, there was an increased risk of infection associated with receiving increasing numbers of injections in the 6 months prior to interview (p < .001 for trend). Women reporting sex against their will in the year prior to interview were at higher risk of infection (OR, 7.84; 95% CI, 1.29-47.86; p = .020). CONCLUSIONS: The strongest risk factor for HIV incidence in this rural Ugandan population is lifetime sexual partners. The increased risks found for women reporting coercive sex and men reporting injections require further investigation. PMID- 10866236 TI - Low estimates of HIV seroconversions among clients of a drug treatment clinic in San Francisco, 1995 to 1998. AB - We estimated HIV incidence among injection drug users attending a drug treatment clinic in San Francisco from 1995 to 1998 using two methods. An anonymous sequential testing method identified no seroconversions among clients seen more than once during the period (one-sided upper 95% confidence limit 1.02 per 100 person-years). A sensitive/less sensitive immunoassay testing strategy detected no early infections (one-sided upper 95% confidence limit 1.90% per year). Methods were concordant and feasible in the setting. Although detection of no new HIV infections in this population of injection drug users (IDUs) is encouraging, epidemiologic studies among IDUs not in treatment are needed to monitor the HIV epidemic effectively. PMID- 10866237 TI - Distribution of HIV-1 subtypes among HIV-seropositive patients in the interior of Cote d'Ivoire. AB - Limited data exist on the distribution of HIV-1 subtypes in Cote d'Ivoire. The aim of this study is to describe the distribution of genetic subtypes of HIV-1 strains in six regions of Cote d'Ivoire. In 1997, we consecutively collected blood from 172 HIV-1-infected patients from six regional tuberculosis treatment centers. Peripheral blood mononuclear cells (PBMCs) from these people were analyzed by a restriction fragment-length polymorphism (RFLP) assay that involves a sequential endonuclease digestion of a 297-base pair polymerase chain reaction (PCR) fragment; plasma samples were tested by a V3-loop peptide enzyme immunoassay (PEIA). DNA sequencing of the protease or env genes was performed on all samples discordant in the two assays as well as a random sample of the concordant subtyped samples. Of 172 specimens, 3 were PCR-negative, and 169 were putatively classified as subtype A by RFLP. The 3 PCR-negative samples were unequivocally subtyped A by PEIA. Of the 169 RFLP subtype A samples, 159 (94%) were subtyped A by PEIA. Of the 10 discordant samples, PEIA testing classified 3 as subtype C, 2 as D, and 5 as F. Sequencing of the env gene classified these samples as 1 subtype A, 4 Ds, and 5 Gs. Thus, 163 (95%) of the specimens were subtype A, 3 subtype D, 4 subtype G, 1 A/D, and 1 A/G (IbNG) circulating recombinant forms (CRF). In conclusion, most HIV-1-infected tuberculosis patients throughout the interior of Cote d'Ivoire are infected with HIV-1 subtype A, which are very likely the A/G (IbNG) CRF. The uniform distribution of this subtype makes Cote d'Ivoire a potential site for vaccine trials. PMID- 10866238 TI - Multigenic polymorphisms of HIV-1 coreceptors and vertical transmission. PMID- 10866239 TI - Routine ophthalmologic screening for cytomegalovirus retinitis in patients with AIDS. PMID- 10866240 TI - Effects of 2-year antiretroviral combination therapies on HIV-1 DNA levels. PMID- 10866241 TI - No evidence of HTLV-I or HTLV-II infection among the Hmong people of northern Thailand or injecting drug users in Bangkok. PMID- 10866242 TI - Blunt bowel and mesenteric injuries: the role of screening computed tomography. AB - BACKGROUND: Early generation scanners have demonstrated poor sensitivity detecting blunt bowel/mesenteric injuries (BBMI). This study was aimed at determining the accuracy and role of helical scanners in BBMI. METHODS: Retrospective chart review of patients with BBMI, or computed tomographic scans suspicious of BBMI, from August of 1995 to December of 1998. RESULTS: One hundred of 8,112 scans (1.2%) were suspicious of BBMI. Of these suspicious scans, 53 patients had BBMI (true positive-TP) and 47 patients did not (false positive-FP). Seven patients with negative scans had BBMI (false negative-FN). Computed tomography contributed toward early surgery in 77% of patients who may have been delayed. Six patients developed intra-abdominal abscess. The abscess group had a significantly longer time interval from injury to surgery. Multiple findings were seen in 57% of true positive scans, whereas in 13% of false positive scans (p < 0.0001). An algorithm for management of BBMI is presented. CONCLUSION: Helical scanners have high accuracy in detecting BBMI. Single versus multiple findings are useful in managing these injuries. PMID- 10866243 TI - Magnetic resonance cholangiopancreatography (MRCP) in the assessment of pancreatic duct trauma and its sequelae: preliminary findings. AB - BACKGROUND: The purpose of this study was to determine the utility of magnetic resonance cholangiopancreatography (MRCP) in the evaluation of pancreatic duct trauma and pancreas-specific complications. METHODS: Ten hemodynamically stable patients with clinically suspected pancreatic injury related to blunt abdominal trauma (n = 8), penetrating trauma (n = 1), or iatrogenic trauma (n = 1) underwent MRCP. Two abdominal radiologists conducted a review of the MRCPs to assess for the presence or absence of pancreatic duct trauma and pancreas specific complications such as pseudocysts. The MRCP findings were correlated with endoscopic retrograde cholangiopancreatograms (n = 2), surgical findings (n = 1), computed tomographic scans (n = 10), and with clinical, biochemical or imaging follow-up (n = 10). RESULTS: Diagnostic quality MRCPs were obtained in each of the 10 patients. A mean imaging time of 5 minutes was required to perform the MRCPs. Pancreatic duct injuries were detected in four patients; pseudocysts were detected in three of these four patients. The pancreatic duct injuries in three patients were acute or subacute. In one of the three patients, disruption of a side branch of the pancreatic duct diagnosed with MRCP was not detected with endoscopic retrograde cholangiopancreatography but was confirmed surgically. In the fourth patient, the pancreatic duct injury was chronic; MRCP revealed a posttraumatic stricture in this patient who had sustained blunt abdominal trauma 17 years previously. In the remaining six patients, pancreatic duct trauma was excluded with MRCP. The information derived from the MRCPs was used to guide clinical decision-making in all 10 patients. CONCLUSIONS: MRCP enables noninvasive detection and exclusion of pancreatic duct trauma and pancreas specific complications and provides information that may be used to guide management decisions. PMID- 10866244 TI - Identification of trauma patients at risk of thoracic aortic tear by mechanism of injury. AB - OBJECTIVE: We sought to identify potential measurable on-scene information that would assist clinicians in the identification of patients at risk for thoracic aortic tear (AT) after vehicular trauma. METHODS: Data were prospectively collected at the scene of 295 motor vehicle crashes from 1995 to 1999. There were 34 cases (12%) with AT. Scene data consisted of vehicle maximal crush, maximal intrusion into the occupant compartment, change in velocity (Delta V) and principal direction of force. Thoracic aortic injuries were confirmed radiographically or at autopsy. Crash factors were analyzed for correlation with AT by logistic regression. RESULTS: Delta V > or = 20 mph and near-side impact were the factors having the strongest correlation with thoracic aortic injury. Delta V > or = 20 mph (n = 32 with AT) had an odds ratio = 6.4, (p < 0.01). Near impact (n = 20 with AT) had an odds ratio = 2.3, (p < 0.05) and intrusion > or = 15 inches had an odds ratio = 3.2, p < 0.05. The sensitivity, specificity, and accuracy of the presence of near impact, Delta V > or = 20 mph, or both, were 100%, 34%, and 64%. The positive and negative predictive values were 16% and 100%, respectively. There was no relationship of AT to use of seat belts or airbags. CONCLUSION: Thoracic aortic injury after vehicular collision can be reliably excluded if near-impact, Delta V > or = 20 mph, or intrusion > or = 15 inches are not present. Mechanism of injury in the form of crash scene information may aid clinicians in identifying individuals at risk for thoracic aortic tear after vehicular trauma. PMID- 10866245 TI - Family presence during trauma resuscitation: a survey of AAST and ENA members. American Association for the Surgery of Trauma. Emergency Nurses Association. AB - BACKGROUND: The Emergency Nurses Association (ENA) has formally resolved that family presence (FP) during resuscitation and invasive procedures (TR) is the right of the patient and is beneficial for both patients and family members. Furthermore, FP during TR has been implemented at several trauma centers. Because this policy is controversial, a survey was conducted to assess the opinions of members of the American Association for the Surgery of Trauma (AAST) and ENA in regard to FP. METHODS: A survey instrument regarding FP during TR was mailed to the AAST membership (n = 813) and a random sampling (10%) of ENA members (n = 2,988). Questions regarding membership (AAST vs. ENA), age, gender, years in practice, trauma experience, the patient's right to FP during the primary survey, secondary survey, and invasive procedures, the potential effects of FP on trauma team function, and medicolegal implications were included in the survey. Qualitative and quantitative variables were analyzed by analysis of variance and chi2 analysis, respectively. Responses to questions by using a Likert Scale for degree of agreement were analyzed by using the Kruskal-Wallis test. RESULTS: A total of 1,629 (AAST, n = 368; ENA, n = 1,261) surveys were returned (43.4% response). There were 44 surveys returned as undeliverable (1.2%). The members of the AAST were older, more likely to be male, had been in practice longer, and had greater trauma experience when compared with ENA members (p < 0.001). More AAST than ENA members (97.8% vs. 80.2%) believed that FP during all phases of TR was inappropriate (p < 0.001). Fewer AAST members believed that FP was a patient right when compared with ENA members (p < 0.0001). The AAST members were more likely to believe FP interfered with patient care and increased the stress of trauma team members (p < 0.0001). The majority of AAST and ENA members had experience with FP during TR (55.3 vs. 67.8%; p < 0.001). However, the impressions of their experiences were widely disparate, with 63.6% of ENA and only 17.5% of AAST members, indicating that the experience was beneficial (p < 0.001). CONCLUSION: Attitudes toward FP during TR are significantly different between AAST and ENA members. Because of these differences in opinion, implementation of an FP policy may create conflicts between trauma team members and may interfere with the effectiveness of the trauma team. PMID- 10866246 TI - Cerebral oxygenation during hemorrhagic shock: perils of hyperventilation and the therapeutic potential of hypoventilation. AB - OBJECTIVES: Prophylactic hyperventilation of patients with head injuries worsens outcome, presumably by exacerbating tissue hypoxia. Oxygen tension in brain tissue (PbrO2) provides a direct measurement of cerebral metabolic substrate delivery and varies with changing end-tidal carbon dioxide tension (ETCO2) and mean arterial pressure. However, the effects of hyperventilation and hypoventilation on PbrO2 during hemorrhagic shock are not known. The aim of this study was to examine the effects of alteration in ventilation on PbrO2 in hemorrhaged swine. METHODS: Clark-type polarographic probes were inserted into the brain tissue of seven swine to measure PbrO2 directly. To examine the effects of alterations in ventilation on hemorrhage-induced hypotension, swine were hemorrhaged to 50% estimated blood volume and PbrO2 was monitored during hyperventilation (RR = 30) and hypoventilation (RR = 4). RESULTS: After the 50% hemorrhage, PbrO2 declined rapidly from 39.8 +/- 4.6 mm Hg to 11.4 +/- 2.2 mm Hg. Hyperventilation resulted in a further 56% mean decrease in PbrO2. Hypoventilation produced a 166% mean increase in PbrO2. These changes were significant (p = 0.001) for absolute and percentage differences from baseline. CONCLUSION: During hemorrhage, alterations in ventilation significantly changed PbrO2: hyperventilation increased brain-tissue hypoxia whereas hypoventilation alleviated it. This finding suggests that hyperventilation has deleterious effects on brain oxygenation in patients with hemorrhagic shock and those with head trauma. Conversely, hypoventilation with resultant hypercapnia may actually help resolve hemorrhagic shock-induced cerebral hypoxia. PMID- 10866247 TI - Prehospital hypotension as a valid indicator of trauma team activation. AB - BACKGROUND: Criteria for trauma team activation are continually being evaluated to ensure proper utilization of resources. We examined the impact of prehospital (PH) hypotension (systolic blood pressure < or = 90) on outcome (operative intervention and mortality) and its usefulness as an indicator for trauma team activation. METHODS: A database was created by using the trauma registry for all nonburned, injured patients from July of 1993 through October of 1998 at our Level I trauma center. RESULTS: Of 6,976 patients (83% blunt injury) in the database, 4,437 had a PH blood pressure recorded. Documented PH hypotension was present in 791 patients. Hypotension persisted in the emergency department (ED) in 299 patients, but 193 of them showed minimal or no signs of life on arrival. Four hundred ninety-two patients had PH hypotension but normal ED systolic blood pressure, and 130 patients developed ED hypotension after normal PH systolic blood pressure. Nearly half of the patients with hypotension were taken from the ED directly to the operating room primarily for hemorrhage control procedures. The early and late mortality rates of patients with PH and ED hypotension were 12% and 32%, respectively. Other PH interventions had minimal effect on mortality in the hypotensive patient. CONCLUSION: Prehospital hypotension remains a valid indicator for trauma team activation. Even though most of the non-DOA patients (492 of 598) were stable on arrival to the ED, nearly 50% required operative intervention, and an additional 25% required intensive care unit admission. The trauma team should be activated and involved with these patients early. PMID- 10866248 TI - Rib fractures in the elderly. AB - BACKGROUND: We sought to ascertain the extent to which advanced age influences the morbidity and mortality after rib fractures (fxs), to define the relationship between number of rib fractures and morbidity and mortality, and to evaluate the influence of analgesic technique on outcome. METHODS: A retrospective cohort study involving all 277 patients > or = 65 years old with rib fxs admitted to a Level I trauma center over 10 years was undertaken. The control group consisted of 187 randomly selected patients, 18 to 64 years old, with rib fxs admitted over the same time period. Outcomes included pulmonary complications, number of ventilator days, length of intensive care unit and hospital stay (LOS), disposition, and mortality. The specific analgesic technique used was also examined. RESULTS: The two groups had similar mean number of rib fxs (3.6 elderly vs. 4.0 young), mean chest Abbreviated Injury Scores (3.0 vs. 3.0), and mean Injury Severity Score (20.7 vs. 21.4). However, mean number of ventilator days (4.3 vs. 3.1), intensive care unit days (6.1 vs. 4.0), and LOS (15.4 vs. 10.7 days) were longer for the elderly patients. Pneumonia occurred in 31% of elderly versus 17% of young (p < 0.01) and mortality was 22% for the elderly versus 10% for the young (p < 0.01). Mortality and pneumonia rates increased as the number of rib fxs increased with and odds ratio for death of 1.19 and for pneumonia of 1.16 per each additional rib fracture (p < 0.001). The use of epidural analgesia in the elderly (LOS >2 days) was associated with a 10% mortality versus 16% without the use of an epidural (p = 0.28). In the younger group (LOS >2 days), mortality with and without the use of an epidural was 0% and 5%, respectively. CONCLUSION: Elderly patients who sustain blunt chest trauma with rib fxs have twice the mortality and thoracic morbidity of younger patients with similar injuries. For each additional rib fracture in the elderly, mortality increases by 19% and the risk of pneumonia by 27%. As the number of rib fractures increases, there is a significant increase in morbidity and mortality in both groups, but with different patterns for each group. Further prospective study is needed to determine the utility of epidural analgesia in this population. PMID- 10866249 TI - Fall injuries in the pediatric population: safer and most cost-effective management. AB - BACKGROUND: At our children's hospital, 30% of all trauma admissions are from falls. The aim of this study was to outline inefficiencies and unnecessary costs incurred in the care of these patients. METHODS: The charts of 127 children admitted for falls (height > or = 9 feet) from 1993 to 1996 were reviewed. Patient demographics, injuries, and treatment costs were recorded and analyzed. RESULTS: Fifty-seven children (45%) were evaluated at an outside facility before transfer. Of these, 73% had injuries requiring treatment at the pediatric center. Local hospital work-up resulted in an average treatment delay of 4.5 hours. Additionally, significant cost was incurred by duplication of radiographic studies, the majority of which were normal. CONCLUSION: Improved and more cost effective care in pediatric falls can be ensured by immediate transfer of patients with significant injuries, omission of radiographs before transfer, and avoidance of multiple routine x-ray films, the majority of which are normal. PMID- 10866250 TI - Hyperthermic resuscitation is safe and effective after hemorrhagic shock in dogs. AB - OBJECTIVE: To show that resuscitation from hypothermic, hemorrhagic shock using 65 degrees C intravenous fluid results in a more rapid return to euthermia compared with 40 degrees C intravenous fluid, without significant endothelial or hemolytic injury. DESIGN: Fourteen anesthetized beagles (10-12 kg) were cooled to a core temperature of 30 degrees C and hemorrhaged to a mean arterial pressure of 40 to 45 mm Hg for 30 minutes. The animals were randomized to receive either 65 degrees C or 40 degrees C intravenous fluid through a specially designed catheter at a rate of 80% of their blood volume per hour until euthermic (37 degrees C) or for 2 hours. MATERIALS AND METHODS: Blood pressure, pulmonary artery pressure, heart rate, and core temperature were continuously monitored. Blood samples were collected at baseline, after hemorrhage, 2 hours of resuscitation, and at postmortem examination after 7 days of survival. Laboratory measurements included complete blood count, plasma-free hemoglobin, and osmotic fragility. Values were compared using the Student's paired or unpaired t test with p approximately 0.05 indicating significance. Postmortem examination included light microscopy of the proximal superior vena cava or right atrium. RESULTS: Animals receiving 65 degrees C intravenous fluid warmed 3.6 degrees C/hour, significantly faster than the 40 degrees C animals (1.9 degrees C/hour). There were no significant differences in plasma-free hemoglobin or osmotic fragility. Endothelial injuries were found in two animals in each group. These defects occurred along the path of catheter insertion and not at the infusion site. CONCLUSIONS: Central intravenous fluid at 65 degrees C is a more rapid means of treating hypothermia than standard 40 degrees C intravenous fluid. It is safe even in hypovolemic animals. PMID- 10866251 TI - Influence of etiology in heterotopic bone formation of the hip. AB - OBJECTIVE: Heterotopic ossification (HO) in periarticular tissue can appear after brain or local joint trauma. The aim of this study was to investigate differences of manifestation and outcome after surgical therapy of patients suffering from HO of the hip after isolated brain injury (n = 18), local hip trauma (n = 21), or the combination of both (n = 25). HO can cause progressive lost of joint movement; once mature, the only therapy to improve joint mobility is the excision. METHODS: All patients underwent surgical removal of HO and postoperative irradiation therapy. On the basis of plain radiographic findings, we evaluated the recurrent ossification after 1-year and 5-year follow-up periods. Within this prospective study, clinical performance status was scored according to the classification of d'Aubigne and a planimeter was used to evaluate the area of heterotopic bone formation in standard x-rays films. RESULTS: The severity of brain trauma observed by Glasgow Coma Scale correlated with the ossification size (square centimeters). Correlation was noticed as well between severity of brain injury and functional outcome. The evaluation of an average 1-year and 5-year follow-up period showed relief of pain and clear improvement of range of motion in all patients. There was mild recurrence of heterotopic bone growth within the first postoperative year without deterioration of the functional results. CONCLUSION: The severity of brain trauma has an influence on the extent of HO near the hip joint and also on the rehabilitation process. The histologic findings and recurrence of HO after excision were not affected by the localization of initial trauma. There was only mild recurrence of heterotopic bone growth between the first and fifth postoperative year. For objective evaluation of heterotopic bone formation in standard x- ray films, planimetric measurement is a useful method. PMID- 10866252 TI - Biomechanical comparison for different configurations of tension band wiring techniques in treating an olecranon fracture. AB - BACKGROUND: The aim of this study was to compare the superiority between the newly designed modified AO tension band wiring technique and the traditional modified AO tension band wiring technique in treating an olecranon fracture. METHODS: Eight pairs of fresh cadaveric ulnae were tested biomechanically. After transverse osteotomy of the olecranon, all left ulnae were fixed by the traditional modified AO technique with two Kirschner wires inserted through the anterior ulnar cortex and all right ulnae by the new technique with two Kirschner wires inserted into the marrow cavity from the olecranon to the ulnar styloid process. All specimens were tested by the Material Testing System machine to evaluate fragment displacement and the maximal failure load. A dual linear variable displacement transducer was used to measure relative minimal displacement. RESULTS: There was no significant difference between the techniques. The maximal failure load by either technique was more than 80 kg. Even at testing failure, no Kirschner wires migrated proximally. CONCLUSION: The new technique may be applied widely to treat all olecranon fractures, because it is a technically easier and safer technique. Less than 5.5-kg loads could be permitted in daily activity postoperatively. A single tolerable loading weight should not exceed 8 kg. Kirschner wires will not migrate proximally, despite increased joint loading. Clinically, this study may confirm indirectly the hypothesis that proximal migration of Kirschner wires was mainly due to triceps traction. PMID- 10866253 TI - Intraosseous infusion devices: a comparison for potential use in special operations. AB - OBJECTIVE: To determine which intraosseous (IO) devices were easy to learn to use, easy to use once the skill was obtained, and appropriate for the Special Operations environment. METHODS: Thirty-one Navy SEAL corpsmen, Air Force pararescuemen, Army Special Forces, and Ranger medics, in a prospective, randomly assigned, cross-over study, tested four commercially available, Food and Drug Administration-cleared IO devices. The systems included the injection models First Access for Shock and Trauma (FAST, Pyng Medical) and Bone Injection Gun (Wais Medical, Kress USA Corporation) and the hand-driven threaded-needle SurFast (Cook Critical Care) and straight-needle Jamshidi needle (Baxter) models. The Special Operations medical care providers received a lecture regarding IO use, viewed videotapes of the injection models, and practiced with demonstration units in the classroom. Each participant then entered the cadaver lab where all four of the IO devices were placed in randomly assigned order. A poststudy questionnaire was then completed. The FAST was placed in the sternum, whereas the other units were placed in either medial proximal or distal medial tibia. Each participant was assessed for time, number of attempts, and success. The presence of marrow, extravasation, quality of flow, and security of needle were evaluated in combination to help determine success. RESULTS: All four devices were believed to be easy to learn as well as easy to place. FAST was successful in 29 of 30 insertions (94%) with a placement time of 114 +/- 36 (mean +/- SD) seconds. The Bone Injection Gun was similarly successful (29 of 31 insertions, 94%) with a mean placement time of 70 +/- 33 seconds. This time was statistically significantly faster (p < 0.05) than that with FAST, but not with the other devices. Thirty of 31 SurFast placements (97%) were successful, on average taking 88 +/- 33 seconds to place. The Jamshidi needle also had 30 of 31 successful placements (97%) at an average 90 +/- 59 seconds. No one device was rated by the participants as significantly better than the others; however, the Bone Injection Gun did have 65% of participants rate it as first or second (closest was Jamshidi needle at 52%). CONCLUSION: These IO devices were easy to teach and learn as well as easy to use. Insertion times compared favorably with peripheral intravenous catheter placement in the face of hemorrhage. All four devices can be appropriately used in the Special Operations environment and are reasonable alternatives when intravenous access cannot be gained. Although no device was rated higher than the others, particular features are desirable (low weight/size, simplicity, reusability, secure, clean, well protected). PMID- 10866254 TI - Effects of physiologic albumin and hespan levels on hepatocytes in vitro. AB - BACKGROUND: Although albumin and hydroxyethyl starch (HES) are routinely used in critically ill, hypoalbuminemic patients, no studies have tested the effect of supplemental albumin and HES on hepatocyte function. METHODS: In this study, the effects of these agents were evaluated by using stable, rat hepatocyte cultures in a collagen sandwich configuration. Hepatocyte synthesis of albumin, urea, and intracellular triglycerides was monitored in Dulbecco's modified Eagle medium (supplemented with fetal bovine serum, hydrocortisone, L-proline, gentamycin, and insulin) without supplemental colloid (control cultures) and with supplemental 2% bovine serum albumin (BSA), 4% BSA, 2% HES, or 4% HES. RESULTS: The albumin secretion in control cultures rose from 31.03 microg/day per 10(6) cells on day 3 to 154.17 microg/day per 10(6) cells by day 12 and remained constant. In contrast, the level of albumin synthesis in the 2% and 4% BSA groups rose from significantly higher initial values (p < 0.05) of 71.25 microg/day per 10(6) cells and 73.27 microg/day per 10(6) cells, respectively, to 127.61 microg/day per 10(6) cells and 107.95 microg/day per 10(6) cells by day 7, then declined rapidly to 58.98 microg/day per 10(6) cells and 41.28 microg/day per 10(6) cells by day 12 when cell disruption was present. HES also reduced albumin synthesis. The urea genesis in the control groups and in the treatment groups was found to be comparable throughout the study. The BSA supplemented groups accumulated large amounts of intracellular lipid droplets during the experiment. The intracellular triglycerides analysis found the 4% BSA group to be significantly (p < 0.05) higher than the 4% HES. CONCLUSION: BSA, added to a collagen sandwich hepatocyte preparation, causes reduced hepatocyte synthesis by day 8, probably a result of intracellular triglyceride accumulation, whereas HES reduces synthesis through unidentified mechanisms. PMID- 10866255 TI - Effect of leukocyte-endothelial adhesion antagonism on neutrophil migration and neurologic outcome after cortical trauma. AB - BACKGROUND: Administration of anti-CD11B, a monoclonal antibody directed against the leukocyte adhesion molecule CD11B, results in decreased neutrophil infiltration into injured tissue after experimental ischemia. We determined the effect of anti-CD11B administration on neutrophil migration and neurologic functioning after experimental cortical trauma. METHODS: Injuries were produced by a pneumatic impactor. Treatment animals received anti-CD11B after injury. Neurologic functioning was quantitated at 1, 12, and 24 hours after injury. Neutrophil migration was assessed with the myeloperoxidase assay. RESULTS: Neutrophil influx was increased in injured cortex after trauma. Anti-CD11B significantly reduced neutrophil influx. There was no significant improvement in neurologic functioning after MAb administration. CONCLUSIONS: These results show there is marked neutrophil response to injury as produced with the pneumatic contusion model. This migration may be significantly attenuated by administration of a anti-CD11B. PMID- 10866256 TI - Open or closed diagnostic peritoneal lavage for abdominal trauma? A meta analysis. AB - OBJECTIVES: To perform a meta-analysis of prospective, randomized controlled trials comparing the closed and open technique of diagnostic peritoneal lavage (DPL) in trauma patients to determine whether there are any difference in outcomes. METHODS: A search of MEDLINE database of English language articles published from 1977 to 1999 was conducted by using the terms diagnostic peritoneal lavage, trauma, and randomized controlled trials. A manual search and Cochrane Library database search was also conducted. Seven randomized controlled trials, including a total of 1,126 patients were identified that compared closed versus open technique. Two reviewers assessed the trials independently. Trial quality was critically appraised by using the Jadad Instrument, a validated published quality scale. Data extraction of major complications, technical difficulties, procedure times, and false-negative and false-positive rates was carried out. The fixed effects model was used for statistical analysis. The Peto odds ratio (OR), weighted mean differences and 95% confidence intervals (95% CI) were calculated. RESULTS: The overall quality of studies was poor (mean, 2.4/7). Major complications did not differ significantly between closed versus open technique (OR, 0.65; 95% CI, 0.15 to 2.92. Technical failures and difficulties were significantly higher in the closed group, i.e., OR 4.33 (95% CI, 1.96 to 9.56) and OR 4.19 (95% CI, 2.842 to 6.19), respectively. Accuracy of closed and open DPL was comparable with no difference in false-negative or false-positive rates between the two techniques. Procedure time was consistently lower in the closed technique. CONCLUSIONS: The closed DPL technique is comparable to the standard open DPL technique in terms of accuracy and major complications. The advantage of reduced time to perform the closed DPL is offset by the increased technical difficulties and failures of this group. Therefore, any significant benefit of routine closed DPL in improving outcomes can be excluded with more confidence based on pooled data than by the individual trials alone. PMID- 10866257 TI - Major life events as antecedents to hip fracture. AB - BACKGROUND: This study sought to determine whether the number of antecedent life events reported in the year before hip fracture among elderly patients was normal for the population from which these patients derive. Major life events are events such as births, deaths, major financial dealings, and major health changes. METHODS: Life events reported in the year before a fall and hip fracture for 111 hip fracture patients were compared with those of a control sample of 90 nonfracture, community-dwelling ambulatory elderly. RESULTS: The total number of life events was higher in the hip fracture group (p = 0.0001) than in the community control group. Fracture was also associated with the number of events experienced (adjusted OR, 2.1; 95% CI, 1.6-2.7; p < 0.0007), notwithstanding age, marital status, and education. CONCLUSION: Older persons who had sustained a fall related traumatic hip fracture experienced an increased number of major life events compared with a nonfracture population sample of community-dwelling elderly controls. PMID- 10866258 TI - Value of point-of-care blood testing in emergent trauma management. AB - BACKGROUND: No prospective study demonstrates the value of point-of-care laboratory testing (POCT) in the management of major trauma. METHODS: In a prospective, noninterventional, study of 200 major trauma patients, we evaluated the influence of a blood POCT profile (hemoglobin, Na+, K+, Cl-, blood urea nitrogen, glucose, pH, PCO2, PO2, HCO3-, base deficit, and lactate) on emergent diagnostic and therapeutic interventions. Physicians responded to a standardized set of questions on their diagnostic and therapeutic plans before and after the availability of POCT results. Management plan changes were deemed emergently appropriate, if they were influenced by the POCT results and, within the ensuing 30 minutes, the change in management was likely to reduce morbidity or conserve resources. RESULTS: For emergently appropriate plan changes, Na+, Cl-, K+, and blood urea nitrogen were never influential, whereas in each of 6.0% of cases (95% confidence interval [CI], 3.5%-10.2%) at least one of the remaining POCT parameters was influential. An emergently appropriate change was based on hemoglobin in 3.5% of cases (95% CI, 1.0%-6.1%), blood gas parameters in 3.0% of cases (95% CI, 0.64%-5.7%), lactate in 2.5% of cases (95% CI, 1.1%-5.7%), and glucose in 0.5% of cases (95% CI, 0.1%-2.8%). All of these cases involved blunt injury. CONCLUSION: Na+, Cl-, K+, and blood urea nitrogen levels do not influence the initial management of major trauma patients. In patients with severe blunt injury, hemoglobin, glucose, blood gas, and lactate measurements occasionally result in morbidity-reducing or resource-conserving management changes. PMID- 10866259 TI - Risk of injury through snowboarding. AB - OBJECTIVE: Survey of a group of snowboarders and study of their injuries, as well as analysis of the risk of injury considering the time spent on the snowboard. MATERIALS AND METHODS: Of 7,221 students participating in winter sport programs organized by Austrian schools, 2,745 of those riding snowboards were asked to fill out questionnaires pertaining to demographics, their experience level, equipment, snowboard riding habits, and associated injuries. RESULTS: A total of 2,579 snowboarders (94%), who spent a total of 10,119 days snowboarding, filled out a questionnaire which could be evaluated. A total of 152 snowboarders had suffered a mean of 10.6 injuries per 1,000 days of snowboarding, which required medical care; 5.4/1,000 injuries were moderate or severe. The most common injuries were to the wrist (32%), the hand (20%), and the head (11%). The rate of injury was especially high during the first half-day (roughly 3 hours). Use of wrist protection devices reduced injuries to the wrist from 2 to 0.5% (p = 0.048). CONCLUSION: Risk of snowboard related injury was highest in beginners. Through the use of wrist protection devices, the incidence of the most common injuries was dramatically reduced. PMID- 10866260 TI - Effects of blast exposure on exercise performance in sheep. AB - BACKGROUND: The effects of blast on maximal exercise performance were investigated in sheep that were trained to perform maximal exercise. METHODS AND RESULTS: Sheep were fully instrumented for determination of pulmonary and systemic hemodynamics. Blast exposure was administered by using a compressed air driven shock tube that was positioned to primarily produce cardiopulmonary injury. Four levels of exposure were used that were known to produce sublethal injury ranging from little or no grossly observable cardiopulmonary injury (level 1) to confluent ecchymosis of the heart, lung, or both (level 4). We evaluated maximal exercise performance 1 hour after exposure to level 1, level 2, and level 3 and 24 hours after level 3 and level 4. VO2max was not significantly decreased 1 hour after exposure to level 1 but was decreased after exposure to level 2 (29.9%) and level 3 (49.3%). Significant improvement in exercise performance was observed in 24 hours, as VO2max was not significantly decreased 24 hour after level 3. VO2max was decreased 24 hour after level 4 injury (30.8%). CONCLUSION: Cardiovascular data collected during exercise suggested that acute cardiopulmonary injury is responsible for the exercise performance decrement observed 1 hour after exposure and that significant recovery of function is observed 24 hours after blast injury. PMID- 10866261 TI - Effects of blood resuscitation versus dextran resuscitation after hemorrhage on intrinsic myocardial function. AB - BACKGROUND: Myocardial function is altered by many factors present in hemorrhaged and resuscitated animals. The purpose of this study was to determine whether resuscitation after a short period of hemorrhagic shock, which by itself did not alter intrinsic cardiac function, causes dysfunction. METHODS: Guinea pigs were instrumented to measure blood pressure and cardiac output, and several days later 50% of their blood volume was removed at a rate of 1 mL/min. Some animals were resuscitated with the shed blood and some with 6% dextran. Hearts were studied 1 or 24 hours after resuscitation. RESULTS: Isolated hearts from animals after 1 hour of resuscitation demonstrated dysfunction whether resuscitated with blood or dextran, although dysfunction was more severe with blood resuscitation. By 24 hours, dysfunction was essentially reversed. CONCLUSIONS: Resuscitation after hemorrhagic shock caused injury to the myocardium independent of the hemorrhage. Blood resuscitation resulted in greater dysfunction than did resuscitation with dextran. PMID- 10866262 TI - Guidelines for the diagnosis and management of blunt aortic injury: an EAST Practice Management Guidelines Work Group. AB - In summary, BAI is a lethal result of severe blunt trauma. It should be considered in all patients who sustained injury by a deceleration or acceleration mechanism, especially in the face of physical or radiographic findings suggestive of mediastinal injury. Angiography remains the "gold standard" for diagnosis, although CT scanning is taking more of a role, especially for screening. Diagnosis should be followed by prompt surgical repair using some method of distal perfusion to minimize renal and spinal cord ischemia. If prompt repair is not feasible because of other injuries or comorbidities, medical control of blood pressure is warranted in the interim. PMID- 10866263 TI - Popliteal artery trauma in a forward deployed Mobile Army Surgical Hospital: lessons learned from the war in Kosovo. PMID- 10866265 TI - Two cases of blunt hepatic injury with active bleeding from the right inferior phrenic artery. PMID- 10866264 TI - Incarceration of the basilar artery in a longitudinal fracture of the clivus: case report and literature review. PMID- 10866266 TI - Obliteration of a late traumatic posterior tibial artery pseudoaneurysm by duplex compression. PMID- 10866267 TI - Stab wounds to the head with intracranial penetration. PMID- 10866268 TI - Emergent thoracotomy for airway control after intrathoracic tracheal injury. PMID- 10866269 TI - A lucky case of penetrating injury of the low chest. PMID- 10866270 TI - The image of trauma. PMID- 10866271 TI - Reducing lethal violence: a reply to the Stolinskys. PMID- 10866272 TI - Pancreatic cancer--more familial than you thought. PMID- 10866273 TI - Hydrolysis profiles of formalin fixed paraffin-embedded tumors based on IOD (integrated optical density) and nuclear texture feature measurements. AB - The aim of the study was to determine optimal hydrolysis time for the Feulgen DNA staining of archival formalin fixed paraffin-embedded surgical samples, prepared as single cell suspensions for image cytometric measurements. The nuclear texture features along with the IOD (integrated optical density) of the tumor nuclei were analysed by an automated high resolution image cytometer as a function of duration of hydrolysis treatment (in 5 N HCl at room temperature). Tissue blocks of breast carcinoma, ovarian serous carcinoma, ovarian serous tumor of borderline malignancy and leiomyosarcoma were included in the study. IOD hydrolysis profiles showed plateau between 30 and 60 min in the breast carcinoma and leiomyosarcoma, and between 40 and 60 min in the ovarian serous carcinoma and ovarian serous tumor of borderline malignancy. Most of the nuclear texture features remained stable after 20 min of hydrolysis treatment. Our results indicate that the optimal hydrolysis time for IOD and for nuclear texture feature measurements, was between 40 and 60 min in the cell preparations from tissue blocks of three epithelial and one soft tissue tumor. PMID- 10866274 TI - Improved sensitivity in comparative genomic hybridization analysis of DNA heteroploid cell mixtures after pre-enrichment of subpopulations by fluorescence activated cell sorting. AB - Cytogenetic analysis of solid tumors with comparative genomic hybridization (CGH) is hampered by the dilution of DNA from individual tumor subpopulations with DNA from other cells. We investigated to what extent this dilution effect can be alleviated using fluorescence activated cell sorting (flow sorting) of experimental DNA heteroploid cell mixtures prior to CGH. From mixtures of normal lymphocytes with triploid K-562 cells the individual components were sorted according to stemline DNA content and processed by CGH in comparison with pure K 562 samples and the original mixtures. Compared with 30 autosome copy number imbalances found in pure K-562 samples, a mixture with 32% K-562 cells showed 16 imbalances, and none were detected in mixtures with 13% or 5% K-562 cells. In contrast, 29, 22 and 23 imbalances were detected in K-562 nuclei sorted from the 32%, 13% and 5% mixtures, respectively. This indicate that CGH analysis of flow sorted DNA aneuploid subpopulations enables a specific cytogenetic analysis of the individual subclones in a DNA heteroploid cell population. PMID- 10866275 TI - Spatial analysis of the neuronal density of aminergic brainstem nuclei in primary neurodegenerative and vascular dementia: a comparative immunocytochemical and quantitative study using a graph method. AB - A graph method was employed to analyse spatial neuronal patterns of pontine nuclei with ascending aminergic projections to the forebrain (nucleus centralis superior (NCS), raphes dorsalis (NRD) and locus coeruleus (LC)), in Alzheimer disease (AD), Huntington disease (HD), and vascular (VD) as well as "mixed-type" (VA) dementia, compared with non-demented controls (CO) and a small sample of brains from schizophrenics ("dementia praecox" (DP)). The quantitative evaluations by the "minimal spanning tree (MST)" were complemented by rough neurofibrillary tangle (NFT) counts and by semiquantitative immunohistochemical assessment of amyloid deposition, neuritic plaque formation, and cellular gliosis. The AD cases showed a significant decline of neuronal density in all nuclei examined, as compared with controls and DP. Neuronal loss was not significant in VD, while the mixed cases with both vascular and Alzheimer-type pathology exhibited pronounced changes of neuronal density. Amyloid deposition occurred almost exclusively in AD and VA, as a rule, being of moderate degree, except for two presenile AD cases where it was marked. NFT were significantly increased in all nuclei in AD and in the VA cases, while they only occasionally appeared beyond age 55 in HD, DP and CO. The four HD cases showed in the NCS and NRD neuronal loss as severe as in AD. This neuronal loss implicates impairment of serotoninergic and noradrenergic neuromodulation as one basic mechanism promoting dementia in AD, VA and perhaps in HD. PMID- 10866276 TI - Critical evaluation of techniques to detect and measure cell death--study in a model of UV radiation of the leukaemic cell line HL60. AB - The reliability of eight distinct methods (Giemsa staining, trypan blue exclusion, acridine orange/ethidium bromide (AO/EB) double staining for fluorescence microscopy and flow cytometry, propidium iodide (PI) staining, annexin V assay, TUNEL assay and DNA ladder) for detection and quantification of cell death (apoptosis and necrosis) was evaluated and compared. Each of these methods detects different morphological or biochemical features of these two processes. The comparative analysis of the 8 techniques revealed that AO/EB (read in fluorescence microscopy) provides a reliable method to measure cells in different compartments (or pathways) of cell death though it is very time consuming. PI staining and TUNEL assay were also sensitive in detecting very early signs of apoptosis, but do not allow precise quantification of apoptotic cells. These three methods were concordant in relation to induction of apoptosis and necrosis in HL60 cells with the various UV irradiation time periods tested. Both AO/EB (read by flow cytometry) and annexin V-FITC/PI failed to detect the same number of early apoptotic cells as the other three methods. Trypan blue is valueless for this purpose. Giemsa and DNA ladder might be useful as confirmatory tests in some situations. PMID- 10866277 TI - Quantification of eosinophilic granule protein deposition in biopsies of inflammatory skin diseases by automated image analysis of highly sensitive immunostaining. AB - Eosinophilic granulocytes are major effector cells in inflammation. Extracellular deposition of toxic eosinophilic granule proteins (EGPs), but not the presence of intact eosinophils, is crucial for their functional effect in situ. As even recent morphometric approaches to quantify the involvement of eosinophils in inflammation have been only based on cell counting, we developed a new method for the cell-independent quantification of EGPs by image analysis of immunostaining. Highly sensitive, automated immunohistochemistry was done on paraffin sections of inflammatory skin diseases with 4 different primary antibodies against EGPs. Image analysis of immunostaining was performed by colour translation, linear combination and automated thresholding. Using strictly standardized protocols, the assay was proven to be specific and accurate concerning segmentation in 8916 fields of 520 sections, well reproducible in repeated measurements and reliable over 16 weeks observation time. The method may be valuable for the cell independent segmentation of immunostaining in other applications as well. PMID- 10866278 TI - The AgNOR quantity as prognostic parameter in choroidal melanomas: a standardised analysis. AB - In order to assess the prognostic significance of silver-stained nucleolar organizer region (AgNOR) proteins, a standardised analysis has been performed on 34 ocular globes with choroidal melanomas. On formalin-fixed paraffin-embedded sections, the visualisation and quantification of AgNORs were achieved according to the guidelines of the Committee on AgNOR Quantification (1995); statistical analysis was performed on the mean AgNOR area values (NORA). We have encountered significantly higher NORA values in nonspindle shaped elements, in tumours of larger dimensions as well as in those with worse clinical course; no correlations were achieved when the AgNOR quantity was compared with age, sex and amount of pigment. The comparison of Kaplan-Meier survival curves revealed that patients affected by melanomas with higher NORA values (>3.327 microm2), non-spindle cell histotype and increased size of tumour had a worse prognosis; finally, by Cox multivariate analysis, the AgNOR quantity appeared the only independent prognostic variable to predict the final outcome of patients. PMID- 10866279 TI - Inverse relationship between Leydig cell density and metastatic potential of prostatic adenocarcinoma. AB - PURPOSE: Evaluate the relationship between metastatic potential of prostatic adenocarcinoma (PC) and testicular Leydig cell density. MATERIALS AND METHODS: Tissue samples from 111 men, age 52-85, with PC and bilateral orchiectomy were evaluated for Leydig cell density. The patients were divided into two groups: Group A were patients with metastasis (n = 36) and Group B were patients without metastasis (n = 75). Leydig cell density was determined by direct manual microscopic cell count on the tissue sections. The means of cell counts by four pathologists, expressed as cell/0.78 mm2 were used for analysis. The normally distributed data were analyzed by two-tail Student's t-test. Thirty-eight age compatible autopsy cases who died of unrelated causes served as normal controls. RESULTS: The mean of Leydig cell count in group A patients was 14.43 (14.43+/ 1.19 SE). Mean of Group B was 47.05 (47.05+/-4.05 SE) whereas normal controls displayed a mean of 48.66 (48.66+/-2.94 SE). Group A was significantly different from control (p < 0.00001). Group A and Group B were also significant different (p < 0.001) whereas control was not significantly different from Group B (p > 0.75). CONCLUSIONS: Patients with metastatic adenocarcinoma of prostate, as a group, have a significantly lower Leydig cell density than patients without metastasis or patients without PC in compatible age groups. The hormonal relationship between this observation is however unknown. One possible explanation is that PC subpopulation with metastatic potential may require different level of endogenous androgen or are androgen-independent. PMID- 10866280 TI - Proliferation index and karyometric features of pancreatic intraductal proliferative lesions. AB - The increasing frequency and poor prognosis in pancreatic cancer prompt us to search for morphological lesions being a substrate for its development. Studies of autopsy and surgically resected material as well as recent molecular studies have proved that one of the possible pathways of pancreatic neoplasia is the intraepithelial proliferation--dysplasia--cancer sequence. In the present paper we studied the proliferative activity (Ki-67 index) in pancreatic intraepithelial proliferative lesions and its correlation with geometric features of cell nuclei as signs of increasing dysplasia. The studies were carried out in a group of 35 patients operated on for pancreatic cancer, chronic pancreatitis and other conditions not associated with the pancreas. We used immunohistochemical methods and basic morphometric parameters. The results of our studies indicate that the cell proliferative activity depends both on the type of epithelial proliferation and underlying pancreatic disease. The values of Ki-67 index are significantly different in low-grade proliferation (flat and papillary hyperplasia) and high grade proliferation (atypical papillary hyperplasia and carcinoma in situ). A set of karyometric features correlates with Ki-67 index but there is no single feature which would have a diagnostic value. PMID- 10866281 TI - Microenvironment-induced cancer metastasis. AB - PURPOSE: To present and evaluate clinical data suggesting that cancer metastasis may be induced by the microenvironment of the primary tumour and to discuss possible mechanisms of microenvironment-induced metastasis, based on a critical review of relevant data from studies of experimental tumours and cells in culture. CONCLUSIONS: Low oxygen tension in the primary tumour is associated with metastasis in soft tissue sarcoma, cervix carcinoma and carcinoma of the head and neck. Multiple mechanisms may be involved in hypoxia-induced metastasis. Thus, hypoxia followed by reoxygenation may induce point mutations and DNA strand breakage leading to deletions, amplifications and genomic instability. Hypoxia may also provide a physiological pressure in tumours selecting for metastatic cell phenotypes. Moreover, hypoxia may induce a temporary increase in the expression of gene products involved in the metastatic cascade, either through gene amplifications or through normal physiological processes by activating oxygen sensors, hypoxia signal transduction pathways and DNA transcription factors. Low glucose concentration, high lactate concentration and low extracellular pH may induce metastasis by similar mechanisms as hypoxia. Tumour reoxygenation during radiation therapy may promote microenvironment-induced metastasis by rescuing hypoxic or nutritionally deprived metastatic cells from dying. Ionizing radiation can elicit a stress response in tumour cells similar to that elicited by hypoxia. Radiation therapy may therefore adversely affect the rate of metastasis in patients who do not achieve control of the primary tumour by enhancing the expression of gene products of importance in metastasis. PMID- 10866282 TI - Increased G2 chromosomal radiosensitivity in cancer patients: the role of cdk1/cyclin-B activity level in the mechanisms involved. AB - PURPOSE: To test the hypothesis that deficient DNA repair as measured by increased G2 chromosomal radiosensitivity results from up-regulation of cdk1/cyclinB and cell cycle control mechanisms during the G2 to M transition. MATERIALS AND METHODS: A total of 185 cancer patients and 25 normal individuals were tested for G2 chromosomal radiosensitivity. The chromatid breaks were analysed in metaphase using the G2 assay or directly in G0 and G2 phase using premature chromosome condensation (PCC). The activity of cdk1/cyclinB, a key regulator of the G2 to M-phase transition, was measured by histone H1 kinase activity and correlated with the development of chromatid breaks after irradiation of cell lines in vitro. RESULTS: Based on the G2 assay, cancer patients on average showed increased chromosomal radiosensitivity above controls. When the analysis was carried out directly in G0 or G2 lymphocytes using PCC, no differences in the induction of chromosomal damage and its repair were observed between G2 assay-sensitive and G2-normal donors. Using the G2 assay to test G2 radiosensitivity in various cell lines, it was found that the higher the cdk1/cyclinB activity level of the cell line tested, the higher the yield of chromatid breaks scored. Furthermore, when mitotic cells from these cell lines were used for PCC induction in irradiated G2 lymphocytes it was observed that the higher the cdk1/cyclinB activity level of mitotic cells used, the higher was the induced yield of chromatid breaks. CONCLUSION: The cdk1/cyclin-B activity levels during the G2 to M transition impair DNA repair processes and play a major role in the yield of chromatid breaks induced after G2-irradiation. Regulation of cdk1/cyclinB complex activity rather than deficient repair enzymes of DNA damage may underlie the mechanisms of G2 radiosensitivity. PMID- 10866283 TI - Increased repair and cell survival in cells treated with DIR1 antisense oligonucleotides: implications for induced radioresistance. AB - PURPOSE: To determine whether repression of a recently isolated, X-ray-responsive gene, DIR1, using antisense oligonucleotides could affect clonogenic cell survival and repair of DNA strand breaks and have a possible role in the mechanism underlying the phenomenon of 'induced radioresistance' (IRR). MATERIALS AND METHODS: Three cell lines, V79, RT112 and UM-UC-3, which are known to exhibit low-dose hypersensitivity (HRS) and induced radioresistance (IRR), and the radiosensitive cell line ATBIVA, were transfected with antisense oligonucleotides directed towards the DIR1 gene. Scrambled oligonucleotides were used as controls. DNA single-strand break (ssb) repair, using the alkaline comet assay, and cell survival using a standard clonogenic assay was measured after exposure to X-rays. RESULTS: Following treatment with 4Gy X-rays, the V79, RT112 and UM-UC-3 cell lines all exhibited significantly increased rates of ssb repair after transfection with DIR1 antisense oligonucleotides compared with cells transfected with scrambled oligonucleotides. They also demonstrated significantly enhanced survival after exposure to 2 Gy X-rays; the radiosensitive ATBIVA cells did not show these effects. CONCLUSIONS: Repression of the DIR1 gene product leads to an increase in the rate of repair and cell survival in three radioresistant cells lines but not in the radiosensitive ATBIVA cell line. Because DIR1 is repressed by X-rays in the dose range where IRR is observed, it may represent a candidate gene involved in the IRR phenomenon. PMID- 10866284 TI - Inverse dose-rate effects at the level of proteins observed in the presence of lipids. AB - PURPOSE: Radical-chain mechanisms such as lipid peroxidation are known to show an inverse dose-rate effect, i.e. the radiation effect increases with decreasing dose rate at identical doses applied. The present study was intended to investigate whether an inverse dose-rate effect can be transferred from the level of lipids to the level of proteins. METHOD: Functional inactivation or structural modification by 80kV X-rays of two classes of proteins was investigated: membrane proteins with a natural environment of lipids like the Ca-ATPase of the sarcoplasmic reticulum, succinate dehydrogenase and F0F1-ATPase from the inner mitochondrial membrane. The second class comprises the water-soluble proteins cytosolic creatine kinase (MM-CK) and bovine serum albumin (BSA). Their modification by free radicals of water radiolysis was investigated in the absence and presence of lipid vesicles. RESULTS: For all proteins investigated, an inverse dose-rate effect was observed in the presence of lipids. This also holds for the water-soluble proteins MM-CK and BSA. In the latter two cases, the dose rate effect disappeared either in the absence of (unsaturated) lipids or in the presence of alpha-tocopherol. CONCLUSION: The largely identical results obtained for a variety of different proteins indicate that inverse dose-rate effects are a normal consequence of radiation induced protein damage in the presence of lipids. In view of the high amount of cellular lipids, this should also hold for the situation in vivo, although due to the comparatively high concentration of intracellular antioxidants the dose-rate dependence might be strongly reduced or even virtually abolished. PMID- 10866285 TI - Cytoprotection by WR-1065, the active form of amifostine, is independent of p53 status in human malignant glioma cell lines. AB - PURPOSE: This study tests the hypothesis that p53 status, i.e. wild type versus mutant form, is a determinant in radiation protection of human glioma cells by WR 1065, the active thiol form of amifostine (WR-2721). MATERIALS AND METHODS: The cytoprotective effectiveness of WR-1065 when present during irradiation was investigated using four well-characterized human glioma cell lines. The p53 positive lines were U87 and D54, and the mutant p53 lines were U251 (mutant at codon 273; CGT/CAT; Arg/His) and A172 (mutant at codon 242; TGC/TTC; Cys/Phe). Treatment conditions included exposure of cells to a range of doses (0-10Gy) alone or in combination with 4mM of WR-1065 added 30min prior to irradiation. Resultant survival curves were obtained using a clonogenic assay and protection factors, the ratio of terminal slopes +/- WR-1065, were determined for each glioma cell line. RESULTS: The Do values of wild-type U87 and D54 were 1.62 and 1.89Gy while those of p53 mutants U251 and A172 were 1.64 and 1.68 Gy, respectively. Protection factors were determined to be 2.4 and 1.9 for U87 and D54, and 2.6 and 2.8 for U251 and A172, respectively. CONCLUSIONS: The p53 status of the four human glioma cell lines tested was not a predictor for either their relative sensitivity to ionizing radiation or ability to be protected by WR-1065. It is concluded that cytoprotection exhibited by cells exposed to WR-1065 during irradiation is independent of their p53 status. PMID- 10866286 TI - Long-term effects of total-body irradiation on the kidney of Rhesus monkeys. AB - PURPOSE: To investigate the long-term effects of total-body irradiation (TBI) on kidneys in non-human primates. METHODS AND MATERIALS: The kidneys of Rhesus monkeys were histologically examined at 6-8 years after TBI with low single doses of 4.5-8.5Gy or two fractions of 5.4Gy. The kidneys of age-matched non-irradiated monkeys served as controls. Irradiation was performed on adult monkeys aged about 3 years; 6-8 years later animals were sacrificed and the kidneys removed and processed for histology. A semi-quantitative scoring system was used to evaluate overall histological damage. Glomerular changes were also morphometrically analysed according to previously published criteria. In selected dose groups (pro)thrombotic and inflammatory changes were investigated by immunostaining cryosections with antibodies against von Willebrand factor (vWF), leukocytes and macrophages. RESULTS: Histological changes were generally mild and only seen in kidneys irradiated with doses higher than 7 Gy. Glomerular changes were characterized by increased mesangial matrix and capillary dilatation. Tubulo interstitial changes included hypercellularity, fibrosis and mild tubular atrophy. The mean glomerular area expressing vWF protein in the irradiated kidneys was not different from that in the age-matched controls. Numbers of infiltrating leukocytes were not significantly different between irradiated kidneys and controls. However, slightly increased numbers of macrophages were present in the renal cortex after irradiation. CONCLUSIONS: Renal damage after TBI of Rhesus monkeys with single doses of 4.5-8.5 Gy or two fractions of 5.4 Gy was mild, even after follow-up times of 6-8 years. PMID- 10866287 TI - Radiation-induced tissue abnormalities in fetal brain are related to apoptosis immediately after irradiation. AB - PURPOSE: To investigate the relation between the incidence of radiation-induced tissue abnormalities in fetal brain and the extent of p53-dependent apoptosis. MATERIALS AND METHODS: Pregnant mice with wild-type p53(+/+), heterozygous p53(+/ ) and homozygous mutant p53(-/-) fetuses received whole-body X-irradiation on day 13 of gestation. The extent of apoptosis 6 hr after irradiation and the incidence of tissue abnormalities 3 days after irradiation in the brain were evaluated by histological examination of brain mantle. RESULTS: The percentage of apoptotic cells increased linearly with dose in p53(+/+) and p53(+/-) fetuses, but no increase was found in p53(-/-). Approximately twice the dose was necessary in p53(+/-) fetuses to induce an apoptotic response to the extent observed in p53(+/+). Fetuses with brain-tissue abnormalities, such as a destroyed ventricular lining and rosettes with a central hollow appeared at a dose of 1.5 and 3.0 Gy, and the incidence was markedly increased following a dose of 2.25 and 3.75Gy in p53(+/+) and p53(+/-) mice, respectively, but no fetus with tissue abnormalities appeared in p53(-/-) at up to 3.75 Gy. Approximately twice the dose was necessary in p53(+/-) fetuses to induce brain-tissue abnormalities to the extent seen in p53(+/+) mice. CONCLUSION: The extent of apoptosis 6 hr after irradiation and the incidence and severity of brain-tissue abnormalities 3 days after irradiation corresponded well, suggesting that radiation-induced tissue abnormalities, such as destroyed ventricular lining, deranged glial fibre and appearance of rosettes in fetal brain were closely related to apoptosis seen 6 hr after irradiation. PMID- 10866288 TI - High prevalence of chromosome 10 rearrangements in human lymphocytes after in vitro X-ray irradiation. AB - PURPOSE: To evaluate the chromosome symmetric or asymmetric rearrangement (CR) frequency for chromosome 10 compared to chromosomes 1 and 3 induced in vitro in human lymphocytes by low doses of X-rays. MATERIALS AND METHODS: Blood samples obtained from three young and healthy volunteers were irradiated in G0 with 0.25, 0.50 and 1 Gy X-rays. Chromosome painting analysis was used on preparations of peripheral lymphocytes for the identification of CR. RESULTS: It was found that radiation-induced CR levels were nonrandomly distributed among the three painted chromosomes. Chromosome 10 CR frequencies were significantly greater than those involving chromosomes 1 (at all the doses tested) or 3 (at 0.25 and 1 Gy), with frequency ratios ranging from 2.2 to 5.2. CONCLUSIONS: In comparison to chromosomes 1 and 3, chromosome 10 appeared to be involved in exchanging at a significantly greater extent than expected according to its DNA content. PMID- 10866289 TI - Localization of radiation-induced chromosomal breakpoints along human chromosome 1 using a combination of G-banding and FISH. AB - PURPOSE: To determine the exact location of radiation-induced chromosomal breakpoints along the euchromatic or heterochromatic regions: G-light and G-dark bands, respectively. MATERIALS AND METHODS: The distribution of radiation-induced chromosomal breakpoints was scored in human lymphocytes irradiated in vitro with 3 Gy of gamma-radiation. Image analysis was applied to combine G-banded and FISH painted images of the human chromosome 1. RESULTS: A total of 195 chromosomal breakpoints in 176 cells with structural chromosomal aberrations was used for the present analysis. Radiation-induced breakpoints were found to be distributed randomly with respect to the p or q arms of chromosome 1 and specific band or band length, but more breakpoints were mapped to G-light than to G-dark bands, the difference being statistically significant. CONCLUSIONS: The results can well be interpreted in terms of concepts of existing models of nuclear architecture, chromatin structure and transcriptional activities of the chromatin, which can influence the induction of primary chromosomal aberrations by gamma-rays. Differential repair of randomly produced primary aberrations may also explain the non-random distribution of radiation-induced breakpoints. PMID- 10866290 TI - Protein gamma-radiolysis in frozen solutions is a macromolecular surface phenomenon: fragmentation of lysozyme, citrate synthase and alpha-lactalbumin in native or denatured states. AB - PURPOSE: To test whether radiolysis-induced fragmentation in frozen aqueous protein solution is dependent on solvent access to the surface of the protein or to the molecular mass of the polypeptide chain. MATERIALS AND METHODS: 60Co gamma irradiation of three proteins at -78 degrees C: lysozyme, citrate synthase and alpha-lactalbumin in their native state, with or without bound substrate, or denatured (random coil in urea/acid-denatured state). RESULTS: By SDS polyacrylamide gel electrophoresis/analysis of the protein-fragmentation process, it was found that for a given protein D37 values (dose to decrease the measured amount of protein, with an unaltered polypeptidic chain, to 37% of the initial amount) varied according to the state of the protein. D37 for denatured proteins was always much smaller than for native states, indicating a greater susceptibility to fragmentation. In urea, contrary to the native state, no well defined fragments were observed. Radiolysis decay constants (K= 1/D37) increased with solvent-accessible surface area of these proteins estimated from their radii of gyration in the various states. This is shown also in previous data on native or SDS-denatured proteins. Denatured proteins which have a large surface area exposed to the solvent compared with native ones are more fragmented at equal doses. CONCLUSIONS: It is concluded that D37 is directly related to the exposed surface area and not to the molecular mass of the polypeptide chain. PMID- 10866291 TI - Thymine carboxylation: nucleophilic addition of carbon dioxide radical anion. AB - PURPOSE: The nucleophilic addition properties of carbon dioxide radical anion (CO2*-) towards N1-substituted thymine derivatives in aqueous solution is studied for comparison with their one-electron reducing reactivity. MATERIAL AND METHODS: N2O-Saturated aqueous solutions of 1-methylthymine, 1,3-dimethylthymine, and thymidylyl(3'-->5')-thymidine containing excess formate ions were gamma irradiated at 1.0Gy min(-1). Several carboxylated thymines were isolated by preparative HPLC and identified by GC-MS, NMR and X-ray crystallography. RESULTS: Along with one-electron reduction yielding N-substituted 5,6-dihydrothymines and C5--C5'-linked dihydrothymine dimers, the addition of CO2 radical anion(s) to the C5--C6 double bond of N-substituted thymines produced several mono- and di carboxylic acids, among which N-substituted derivatives of 5,6-dihydrothymine-6 carboxylic acid [5-methyldihydroorotic acid (5-methyl-DHO)] were produced in the highest yield. Similar carboxylation by CO2 radical anions was also observed for thymine dinucleoside monophosphate. The X-ray structure of cis-5,6-dihydro-1 methylthymine-6-carboxylic acid (cis-1,5-dimethyl-DHO) was determined to show a chair conformation in the crystal. CONCLUSIONS: The CO2 radical anion is a nucleophilic radical with rather low reduction potential, thereby possessing a dual reactivity of radical addition preferentially at C6 and one-electron reduction towards thymine-related compounds. PMID- 10866292 TI - Increased risk of breast cancer following irradiation for Hodgkin's disease is not a result of ATM germline mutations. AB - PURPOSE: Long-term survivors of Hodgkin's disease who received mantle-field irradiation at a young age have a strongly increased risk of developing breast cancer. The purpose of this study was to investigate whether this increased risk was substantially greater among women heterozygous for a germline mutation in the ataxia-telangiectasia gene (ATM). MATERIALS AND METHODS: Thirty-two patients were selected who had developed breast cancer at least 10 years following irradiation for Hodgkin's disease before the age of 45 years. In these patients, the complete open reading frame of the ATM gene was analysed for the presence of germline mutations using the protein truncation test and two mutation-specific tests, followed by genomic sequencing. RESULTS: No A-T disease causing germline mutations were found in these selected Hodgkin patients. However, several alternative splicing events were detected which might influence protein expression levels. CONCLUSIONS: The data suggest that truncating mutations in the ATM gene are not a major component underlying the increased risk of breast cancer following Hodgkin's disease. PMID- 10866293 TI - The dose-response relationship for cancer incidence in a two-stage radiation carcinogenesis model incorporating cellular repopulation. AB - PURPOSE: To investigate the role of cellular repopulation in the dose-response relationship for radiation carcinogenesis resulting from high doses of radiation. METHOD: A two-stage mathematical model of radiation carcinogenesis was developed and used to explore the effects of differing assumptions about repopulation by surviving normal stem cells and by one-stage mutants. RESULTS: Characteristically, cancer incidence at any fixed time after irradiation increases with radiation dose, reaches a peak and then declines with dose (the decline reflecting radiation cell-killing). The optimal dose for cancer incidence, and the incidence level at this dose, are strongly influenced by repopulation kinetics. If repopulation does not occur, or is impaired owing to radiation damage to tissues, the highest value of cancer incidence is reduced, and this value occurs at a lower dose than if repopulation had been complete. A similar result is found if repopulation by one-stage mutants is impaired relative to unmutated cells, or if tissue recovery is assisted by immigration of unirradiated cells. CONCLUSIONS: Differing repopulation kinetics can account for differing dose-response relationships after large doses of radiation. These findings are relevant to the occurrence of 'second tumours' following radiotherapy and to the interaction of radiation with other agents. PMID- 10866294 TI - Brief communication: heat-induced accumulation of p53 and hsp72 is suppressed in lung fibroblasts from the SCID mouse. AB - PURPOSE: To investigate how DNA-dependent protein kinase (DNA-PK) contributes to p53-dependent signal transduction after heat shock, thermosensitivity and accumulation of p53 and hsp72 after heat shock in lung fibroblasts derived from the SCID mouse were analysed. MATERIALS AND METHODS: Thermosensitivity at 44 degrees C in colony-forming units and Western blot analysis of p53 and hsp72 were analysed. RESULTS: The results indicated that (1) the thermosensitivity at 44 degrees C of SCID cells was higher than that of parental cells and (2) heat induced accumulation of p53 and hsp72 was abolished and suppressed in SCID cells as compared with that in parental cells respectively. CONCLUSIONS: The findings suggest that the catalytic subunit of DNA-PK may play an important role upstream of p53 and hsp72, which are possible determinants of cellular thermosensitivity. PMID- 10866295 TI - Quiescence in S-phase and G1 arrest induced by irradiation and/or hyperthermia in six human tumour cell lines of different p53 status. AB - PURPOSE: Quiescent S-phase cells, i.e. cells with a DNA content intermediate between G1 and G2 that nevertheless do not synthesize DNA have been previously observed in human melanoma cells exposed to radiation and/or hyperthermia. This phenomenon has now been studied in more detail comparing six human tumour cell lines of different p53 status and thus different cell-cycle checkpoint control. MATERIALS AND METHODS: Two melanoma (Be11, MeWo), two squamous carcinoma (4197, 4451) and two glioma (EA14, U87) cell lines were used. Changes in the cell-cycle distribution after treatment were studied using two-parameter flow cytometry in order to measure DNA content and BrdU incorporation simultaneously. RESULTS: The fraction of unlabelled cells in the S-phase compartment was determined at daily intervals after treatment. Only background levels of such cells were seen in three of the cell lines (Be11, 4197, EA14). With the other three cell lines (MeWo, 4451, U87) we observed a time- and dose-dependent increase: a few days after treatment up to 20% of all cells did not incorporate BrdU. It is interesting to note that Bell, 4197 and EA14 are p53 wild-types and show a G1 block of several hours after irradiation and/or hyperthermia, while MeWo and 4451 are p53 mutants unable to exhibit such a delay, and U87 in spite of being a p53 wild-type has a reduced ability to do so. CONCLUSIONS: The MeWo, 4451 and U87 cell lines have less time available for the repair of DNA damage before entering into the S-phase, which leads to problems during replication and causes some kind of interphase death. Radiation-induced apoptosis does not seem to be involved here, as it is not unequivocally correlated with the induction of a G1 block or with p53 status. PMID- 10866296 TI - Pancreatic duct cell carcinomas express high levels of high mobility group I(Y) proteins. AB - The high mobility group I (HMGI) family of proteins in mammals belongs to a group of nonhistone nuclear proteins known as architectural transcriptional factors. They function in vivo as both structural components of chromatin and auxiliary gene transcription factors. In an earlier study (N. Abe et al, Cancer Res., 59: 1169-1174, 1999), we demonstrated that the expression level of the HMGI(Y) gene/proteins was significantly increased in colorectal adenocarcinoma and colorectal adenoma with severe cellular atypia. In the current study, we analyzed HMGI(Y) expression in several human pancreatic lesions to investigate (a) whether HMGI(Y) overexpression is also observed in pancreatic carcinoma, and (b) the role of HMGI(Y) in the diagnosis of pancreatic neoplasms. To this end, HMGI(Y) expression was determined at the protein level by immunohistochemistry using a HMGI(Y)-specific antibody in 6 surgically resected specimens of nonneoplastic tissue (4 specimens of normal pancreatic tissue and 2 specimens of chronic pancreatitis tissue), 8 pancreatic cystic neoplasms (5 intraductal papillary mucinous adenomas, 1 serous cystadenoma, and 2 solid pseudopapillary tumors), and 15 duct cell carcinomas of the pancreas. Immunohistochemical analysis revealed intense nuclear staining in the pancreatic carcinoma cells, whereas only very faint nuclear staining was seen in the nonneoplastic cells. There was a strong correlation between HMGI(Y) protein overexpression and a diagnosis of carcinoma (P = 0.000018). Thus, an increased expression level of the HMGI(Y) proteins was clearly associated with the malignant phenotype in pancreatic tissue. In addition, a low level of protein expression was also apparent in two of the cystic neoplasms that exhibited cellular atypia, but not in those that did not exhibit cellular atypia. Based on these findings, we propose that the HMGI(Y) proteins could be closely associated with tumorigenesis in the pancreas and that HMGI(Y) could serve as a potential diagnostic molecular marker for distinguishing pancreatic malignancies unambiguously from normal tissue or benign lesions. PMID- 10866297 TI - Inhibition of angiogenesis and induction of apoptosis are involved in E1A mediated bystander effect and tumor suppression. AB - Adenovirus type 5 E1A has been implicated in mediation of tumor suppression. Preclinical gene therapy studies have additionally shown that complete growth suppression can be achieved by incomplete transfer of E1A into tumors, suggesting that a bystander effect may also be associated with E1A. In this study, we investigated the E1A-mediated bystander effect and the mechanisms that may be associated with it. By s.c. inoculating nude mice with a mixture of E1A transfectants and parental cells, we found that the E1A transfectants exhibited a bystander effect on inhibition of tumor growth. We further showed that E1A mediated suppression of angiogenesis and induction of apoptosis in the tumors, likely contributing to the bystander effect. In addition, coculture of E1A transfectants and parental cells in a Transwell unit led to growth retardation and apoptosis mediated by the supernatant in the parental cells, indicating that a secreted factor may also contribute to the bystander effect. Taken together, our results suggested that E1A mediates a bystander effect on tumor suppression by inhibiting angiogenesis and inducing apoptosis. PMID- 10866298 TI - The pyrido[2,3-d]pyrimidine derivative PD180970 inhibits p210Bcr-Abl tyrosine kinase and induces apoptosis of K562 leukemic cells. AB - PD180970 is a novel pyrido[2,3-d]pyrimidine class of ATP-competitive inhibitor of protein tyrosine kinases. We found that PD180970 inhibited in vivo tyrosine phosphorylation of p210Bcr-Abl (IC50 = 170 nM) and the p210BcrAbl substrates Gab2 and CrkL (IC50 = 80 nM) in human K562 chronic myelogenous leukemic cells. In vitro, PD180970 potently inhibited autophosphorylation of p210Bcr-Abl (IC50 = 5 nM) and the kinase activity of purified recombinant Abl tyrosine kinase (IC50 = 2.2 nM). Incubation of K562 cells with PD180970 resulted in cell death. Results of nuclear staining, apoptotic-specific poly(ADP-ribose) polymerase cleavage, and annexin V binding assays indicated that PD180970 induced apoptosis of K562 cells. In contrast, PD180970 had no apparent effects on the growth and viability of p210Bcr-Abl-negative HL60 human leukemic cells. Thus, PD180970 is among the most potent inhibitors of the p210Bcr-Abl tyrosine kinase, which is present in almost all cases of human chronic myelogenous leukemia. These findings indicate that PD180970 is a promising candidate as a novel therapeutic agent for Bcr-Abl positive leukemia. PMID- 10866299 TI - Vaccine-induced apoptosis: a novel clinical trial end point? AB - The functional end point of immunotherapy is to induce tumor regression. Because immune effector mechanisms usually result in apoptosis, the aim of this study was to determine whether measurement of tumor apoptosis ex vivo is a good end point to evaluate the efficacy of cancer vaccines. A prototype vaccine, 105AD7, was administered to colorectal cancer patients before resection of their primary tumors. There was a significant increase in apoptosis of tumor cells within immunized patients compared with control patients as assessed by immunohistochemistry (P = 0.005; n = 16) or by flow cytometry (P = 0.003; n = 34). Preoperative immunization and measurement of tumor cell apoptosis may be a valuable clinical end point for evaluation of new vaccine and other biological approaches. PMID- 10866300 TI - A novel histone deacetylase inhibitor identified by high-throughput transcriptional screening of a compound library. AB - Libraries of compounds are increasingly becoming commercially available for the use of individual academic laboratories. A high-throughput system based on a stably integrated transcriptional reporter was used to screen a library of random compounds to identify agents that conferred robust augmentation of a signal transduction pathway. A novel histone deacetylase (HDAC) inhibitor, termed scriptaid, conferred the greatest effect, a 12- to 18-fold augmentation. This facilitation of transcriptional events was generally applicable to exogenous gene constructs, including viral and cellular promoters, different cell lines and reporter genes, and stably integrated and transiently introduced sequences. Scriptaid did not interfere with a further induction provided by stimulation of the cognate signal transduction pathway (transforming growth factor beta/Smad4), which implied the functional independence of ligand-stimulated transcriptional activation and histone acetylation states in this system. Additional insights into this and other signal transduction systems are likely to be afforded through the application of compound screening technologies. PMID- 10866301 TI - Genomic structure of the human ING1 gene and tumor-specific mutations detected in head and neck squamous cell carcinomas. AB - We characterized the genomic structure of the human ING1 gene, a candidate tumor suppressor gene, and found that the gene has three exons. We also demonstrated that four mRNA variants were transcribed from three different promoter regions. Of 34 informative cases of head and neck squamous cell carcinoma, 68% of tumors showed loss of heterozygosity at chromosome 13q33-34, where the ING1 gene is located. Here we present the first report that three missense mutations and three silent changes were detected in the ING1 gene in 6 of 23 tumors with allelic loss at the 13q33-34 region. These missense mutations were found within the PHD finger domain and nuclear localization motif in ING1 protein, probably abrogating the normal function. PMID- 10866302 TI - Functional evaluation of PTEN missense mutations using in vitro phosphoinositide phosphatase assay. AB - The tumor suppressor gene PTEN is frequently mutated in diverse human cancers and in autosomal dominant cancer predisposition disorders. Recent studies have shown that the lipid phosphatase activity of PTEN is critical for its tumor suppressor function and that PTEN negatively regulates the phosphatidylinositol 3'-kinase protein kinase B pathway. Although more than half of PTEN mutations result in protein truncation, a significant fraction of PTEN mutations are missense mutations. To examine whether tumor-derived and germ-line-derived missense mutations inactivate PTEN lipid phosphatase function, we constructed 42 distinct types of PTEN missense mutations and expressed them in Escherichia coli. The purified (His)6-tagged PTEN proteins were tested for their ability to dephosphorylate inositol 1,3,4,5-tetrakisphosphate and phosphatidylinositol 3,4,5 triphosphate. In addition, we examined the effect of mutant PTENs on the ability of PTEN to bind to the phospholipid membrane. The results revealed that the majority of PTEN missense mutations [38 of 42 (90%)] eliminated or reduced phosphatase activity and that all of the mutations examined had no effect on the membrane binding activity of PTEN. Our study indicated that phosphoinositide phosphatase activity is important for the tumor suppressor function of PTEN and that there may be other mechanisms of PTEN inactivation that are not monitored by in vitro phosphatase assay and in vitro membrane binding assay. PMID- 10866303 TI - Cysteine 230 modulates tumor necrosis factor-related apoptosis-inducing ligand activity. AB - Biologically active tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein is known to form a homotrimer in solution. Unexpectedly, the recombinant active human TRAIL protein purified from bacteria produced two bands (a Mr 21,000 monomer derived from the disruption of the trimer in SDS gels and a Mr 42,000 dimer) on nonreducing SDS gels. The treatment of this TRAIL protein with DTT, a reducing agent, abolished formation of the Mr 42,000 band, suggesting that the Mr 42,000 band was the result of intermolecular disulfide bridge formation. Inspection of the amino acid sequence of human TRAIL protein identified a unique cysteine residue at position 230, and subsequent site directed mutagenesis revealed that this amino acid residue is responsible for the appearance of the Mr 42,000 dimer. The binding analysis using the TRAIL protein and a TRAIL receptor (death receptor 5) revealed that both the dimer and the trimer bind to death receptor 5 with similar affinity. Interestingly, mutation of cysteine 230 to glycine completely abolished the apoptotic activity of TRAIL protein. The disruption of the dimer in the mixture of TRAIL dimer and trimer increased the apoptotic activity slightly, suggesting that the dimer has less apoptotic activity than the trimer. Therefore, our data indicate that cysteine 230 is not only required for TRAIL function but also modulates the apoptotic activity of TRAIL by forming an intermolecular disulfide bridge. PMID- 10866304 TI - Alcohol consumption and cigarette smoking increase the frequency of p53 mutations in non-small cell lung cancer. AB - Epidemiological studies have demonstrated a strong association between tobacco use and lung cancer; however, the genetic targets of these carcinogens and the role of other environmental agents in this process have yet to be defined. We examined the contribution of alcohol use and cigarette smoking top53 gene mutations in patients with non-small cell lung cancer. Mutations of the p53 gene were detected by sequence analysis in 105 patients with non-small cell lung cancer. Patient characteristics significantly associated with p53 gene mutations were determined using logistic regression. Mutations in the p53 gene were present in 53% of the patients (56 of 105). p53 mutations were more common in patients who used alcohol than in patients who consumed less than one drink per day (72 versus 39%; P = 0.003), and were detected more often smokers than nonsmokers (58% versus 10%, P = 0.02). Mutations in the p53 gene were present more often (P = 0.01) in alcohol drinkers who smoked cigarettes [76% (31 of 41)], than in nondrinkers (<1 drink per day) who smoked cigarettes [42% (20 of 48)] or in nondrinkers who did not smoke [14% (1 of 7)]. In conclusion, alcohol consumption and tobacco use are both associated with p53 mutations in non-small cell lung cancer. The link between exposure to both alcohol and tobacco and p53 mutations raises the possibility that alcohol may enhance the mutagenic effects of cigarette smoke in the lung. PMID- 10866305 TI - Consistent and fast inhibition of colon carcinogenesis by polyethylene glycol in mice and rats given various carcinogens. AB - We have previously shown that dietary polyethylene-glycol (PEG) suppresses the occurrence of azoxymethane-induced cancers in an accelerated rat model of colon carcinogenesis. To determine the consistency of this preventive effect, we carried out a long-term study in rats fed the standard American Institute of Nutrition 1976 diet, and 7 short-term prevention studies in rodents. A total of 337 F344 rats, 20 Sprague Dawley rats, and 40 OF1 mice were all given initiating dose(s) of colon carcinogen, and were randomly allocated to experimental groups 7 d later. Treated groups received drinking water containing 5% PEG. After 30 or 162 d, the animals were examined for aberrant crypt foci or tumors in the colon. After two 20 mg/kg azoxymethane injections, the number of F344 rats with colon tumor was lower in rats receiving PEG for 162 d than in carcinogen-injected controls, 5/21 versus 25/27 (P < 0.0001). PEG-fed rats had no invasive cancer, and 10 times fewer colon tumors than controls (0.3+/-0.1 and 3.1+/-0.5 respectively, P < 0.0001). A three-day PEG treatment was sufficient to halve the number of azoxymethane-induced aberrant crypt foci in F344 rats (P = 0.0006). After 16 d of treatment, PEG-fed rats had five times fewer foci than controls (21+/-14 and 100+/-23 respectively, P < 0.0001), but the inhibition was reversible in part when treatment was discontinued. Aberrant crypt foci initiated by N-methyl-N-nitrosourea intra-rectally (40 mg/kg) or by 2-amino-3,4 dimethylimidazo(4,5-f)quinoline p.o. (2 x 200 mg/kg) were suppressed by PEG (P < 0.0001 and P = 0.003 respectively). PEG was active in F344 rats, in Sprague Dawley rats (P = 0.0005), and in OF1 mice (P = 0.001). PEGs with MW between 3350 and 12000 (but not PEG 400), and PEG 8000 from five suppliers, markedly inhibited azoxymethane-induced aberrant crypt foci (all P < 0.01). The prevention was stronger in rats fed a high-fat diet (P < 0.0001) than in rats fed a rodent chow (P = 0.02). PEG was thus a fast, consistent, and potent inhibitor of early colonic precursor lesions. Moreover, PEG is one of the most potent inhibitors of colon tumor in the standard rat model. Since PEG has no known toxicity in humans, we think it should be tested as a chemopreventive agent in a clinical trial. PMID- 10866306 TI - Instability of X chromosome methylation in aberrant crypt foci of the human colon. AB - Aberrant crypt foci (ACF) in the colon have long been thought to be precancerous lesions and therefore monoclonal, but this is unresolved. Eleven ACF were isolated from five female patients. From these ACF, 178 individual aberrant crypts (ACs) were obtained and assessed for clonality using a method based on X chromosome inactivation of the polymorphic X-linked human androgen receptor (HUMARA) gene. Ten ACF were found to be mixtures of monoclonal and polyclonal types. The HUMARA analysis indicated that almost all ACF were polyclonal lesions. Simultaneously, we investigated K-ras mutations in each AC. We found that seven of the ACF harbored the K-ras mutation; strikingly, this was concordant for all of the ACs from a single ACF. These results, by contrast to the results of HUMARA analysis indicate that ACF lesions are monoclonal. This discrepancy suggests that ACF are apparently polyclonal because of de novo methylation on the active X chromosome. To confirm this possibility, we investigated the methylation status of the X chromosome in male ACF using a competitive PCR assay. One hundred nineteen individual ACs were isolated from eight ACF derived from four male patients. A total of 47 of 119 (39%) of male ACs showed de novo methylation of the HUMARA gene. We found that six of the eight male ACF harbored the K-ras mutation, and this was concordant for all of the ACs from a single ACF. We conclude that X chromosome methylation is unstable in ACF and that this might be an early event in colon carcinogenesis. PMID- 10866307 TI - Detection of germ-cell tumor cells in the peripheral blood by nested reverse transcription-polymerase chain reaction for alpha-fetoprotein-messenger RNA and beta human chorionic gonadotropin-messenger RNA. AB - By establishing sensitive nested reverse transcription-PCRs for the detection of mRNA of alpha-fetoprotein (AFP) and beta human chorionic gonadotropin (betahCG), we investigated the presence of circulating tumor cells in the peripheral blood of 119 patients with germ-cell tumor. A total of 336 blood samples obtained before and during therapy were examined with regard to clinical applicability. The overall ratio of positive PCR results was 26.5% and was independent of the serum concentration of AFP and hCG/betahCG. No correlation of the positivity for AFP-mRNA to serum AFP level was found. In contrast, positive results in betahCG PCR were twice as frequent in patients with elevated serum hCG/betahCG levels as in those with normal serum hCG/betahCG levels (P = 0.012). To develop a valid correlation to tumor stage, tumor histology, and serum level of tumor markers, a subgroup of 36 patients was evaluated before definite therapy. The subgroup revealed an overall ratio of 33.3% positive PCR results. The serum level of both of the markers did not correlate with the detection of corresponding mRNA in peripheral blood samples. However, positive betahCG-PCR results were found exclusively in patients with elevated serum hCG/betahCG (6 of 18 versus 0 of 18; P = 0.019). Patients with stage IIC/III germ-cell tumor demonstrated nearly twice the frequency of positive PCR results as patients with stage I tumor [7 (41.2%) of 17 versus 4 (23.5%) of 17] in this subgroup. With regard to histology, positive PCR results were found mostly in embryonal carcinoma. PMID- 10866308 TI - Expression of estrogen receptor (ER)-alpha and ER-beta in normal and malignant prostatic epithelial cells: regulation by methylation and involvement in growth regulation. AB - The aim of the current study is to demonstrate normal and malignant prostatic epithelial cells (PrECs) as targets for receptor-mediated estrogenic and antiestrogenic action. Using an improved protocol, we have successfully isolated and maintained highly enriched populations of normal PrECs from ultrasound-guided peripheral zone biopsies, individually determined to be morphologically normal. Semiquantitative reverse transcription-PCR analyses were used to determine whether transcripts of estrogen receptor (ER)-alpha and those of ER-beta were expressed in our normal PrEC primary cultures, in a commercially available PrEC preparation (PrEC; Clontech), in an immortalized PrEC line established from a benign prostatic hyperplasia specimen (BPH-1), and in three prostatic cancer cell lines (LNCaP, PC-3, and DU145). Expression levels of ER-alpha and ER-beta transcripts were related to those of two estrogen-responsive genes [progesterone receptor (PR) and pS2], at the message levels, to gain insights into the functionality of the ER subtypes in PrECs. Interestingly, only transcripts of ER beta, but not those of ER-alpha, were found in our primary cultures of normal PrECs, along with both PR and pS2 mRNA. These data strongly suggest that estrogen action was signaled exclusively via ER-beta in normal human PrECs. In contrast, PrEC (Clontech) and BPH-1 cells expressed both ER-alpha and ER-beta transcripts and no PR nor pS2 mRNA in PrEC and only a minimal level of PR mRNA in BPH-1. Among the three prostate cancer cell lines, LNCaP expressed ER-beta mRNA along with transcripts of PR and pS2, DU145 expressed messages of ER-beta and PR, and PC-3 cells exhibited ER-alpha, ER-beta, and pS2 mRNA. Thus, unlike normal PrECs, expression patterns of these genes in malignant PrECs are more variable. Treatment of prostate cancer cells with demethylation agents effectively reactivated the expression of ER-alpha mRNA in LNCaP and DU145 and that of pS2 message in DU145. These findings provide experimental evidence that ER-alpha gene silencing in prostate cancer cells, and perhaps also in normal PrECs, are caused by DNA hypermethylation. To evaluate the potential of using antiestrogens as prostate cancer therapies, we have assessed the growth-inhibitory action of estrogens (estradiol and diethylstilbestrol) and antiestrogens (4-hydroxy tamoxifen and ICI-182,780) on PC-3 and DU-145 cells. In PC-3 cells, which express both ER subtypes, estrogens as well as antiestrogens are effective inhibitors. In contrast, in DU145 cells, which express only ER-beta, antiestrogens, but not estrogens, are growth inhibitors. By comparison, ICI 182,780 is the more effective cell growth inhibitor. Importantly, the ICI 182,780-induced antiproliferative effects were reversed by cotreatment of DU145 cells with an ER beta antisense oligonucleotide, hence lending additional support to a central role played by ER-beta in mediating growth-inhibitory action of antiestrogens. PMID- 10866309 TI - Identification of functional estrogen response elements in the gene coding for the potent angiogenic factor vascular endothelial growth factor. AB - Vascular endothelial growth factor (VEGF) is a potent stimulator of angiogenesis and a prognostic factor for many tumors including those of endocrine-responsive tissues such as the breast and uterus. We and others have previously shown that VEGF is regulated by estradiol and tamoxifen in the uterus and by estradiol in human breast cancer cells, and pharmacological evidence has suggested that this regulation was mediated by transcriptional activation of the estrogen receptor (ER). This prompted us to investigate whether the VEGF gene contains sequences that bind the ER and confer hormonal inducibility to reporter constructs in the presence of the two ER subtypes. These studies identified two sequences homologous to the consensus estrogen response element, GGTCAnnnTGACC, which bind both ER-alpha and ER-beta. One of these elements is located in the 5' untranslated region of the VEGF gene (GGGCAaagTGACT), and the other is located in the 3'-untranslated region (GAGCAcccTGCCC). Competition with excess unlabeled oligonucleotides indicates that these two elements bind both ERs specifically, mutations in either half-site of the two elements abolish receptor binding, and ER-alpha- and ER-beta-specific antibodies interact with complexes formed with the corresponding receptor subtypes. In cells containing either ER-alpha or ER-beta, the 3'-element behaves as a traditional enhancer that confers hormone inducibility to reporter constructs in an orientation-independent manner, and transcriptional activity is blocked by the pure antiestrogen ICI 182,780. The pattern of transcriptional activity of the element located in the 5'-flanking region is more complex. In the orientation found in the endogenous gene, this element is nonresponsive to ER-beta but confers estrogen-dependent inhibition of transcription with ER-alpha that is blunted by ICI 182,780. In the opposite orientation, the 5'-element confers hormone inducibility with either ER-alpha or beta, and ICI 182,780 blocks activation by ER-alpha but not by ER-beta. These findings support the hypotheses that estrogens directly regulate VEGF transcription in target tissues and tumors, although such regulation appears likely to involve a complex interplay of cis- and trans-acting elements not previously observed for other hormone-responsive genes. PMID- 10866310 TI - Chemopreventive effects of dietary folate on intestinal polyps in Apc+/-Msh2-/- mice. AB - Epidemiological and animal studies (reviewed in Y. I. Kim, J. Nutr. Biochemistry, 10: 66-88, 1999; J. B. Mason and T. Levesque, Oncology, 10: 1727-1743, 1996) suggest that dietary folate intake is inversely related to the risk of colorectal cancer. However, the optimal timing of folate intervention and mechanisms by which folate modulates colorectal carcinogenesis have not been clearly established. A recently developed murine model of intestinal tumorigenesis, which carries a heterozygous mutation in the Apc gene and a null mutation in the Msh2 gene (Apc+/-Msh2-/-), was used to determine the effect of dietary folate on intestinal tumorigenesis. Apc+/- Msh2-/- mice were randomized to receive either 0 or 8 mg of folate/kg diet starting at either 3 or 6 weeks of age. The 3- and 6 week diet starts represent intervention before and after the establishment of neoplastic foci, respectively. At 11 weeks of age, mice were killed, and the small intestines and colons were analyzed for adenomas and aberrant crypt foci (ACF). Serum folate concentrations were determined by a standard microbiological assay. Genomic DNA methylation was assessed by in vitro [3H]methyl incorporation into hepatic DNA and by a methyl-sensitive restriction digestion method. Microsatellite instability was determined in matched normal and polyp DNA from the small intestine and colon at 5 loci. Serum folate concentrations accurately reflected dietary folate levels (P < 0.005). Folate supplementation, started before the establishment of neoplastic foci, significantly decreased the number of small intestinal adenomas (by 2.7-fold; P = 0.004) and colonic ACF (by 2.8 fold; P = 0.028) and colonic adenomas (by 2.8-fold; P = 0.1) compared with a moderate degree of folate deficiency. In contrast, a moderately folate-deficient diet, started after the establishment of neoplastic foci, significantly reduced the number of small intestinal adenomas (by 4.2-fold; P = 0.001) but had no effect on colonic ACF and adenomas compared with folate supplementation. Genomic DNA methylation and microsatellite instability do not seem to play a major role in folate-modulated intestinal and colonic tumorigenesis in this model. In conclusion, in this murine model, dietary folate supplementation significantly protects against small intestinal and colorectal tumorigenesis if it is provided before the establishment of neoplastic foci However, if it is provided after the establishment of neoplastic foci, dietary folate seems to have an opposite effect. These data suggest that the timing of folate intervention is critical in providing an effective and safe chemopreventive effect on intestinal tumorigenesis. Notwithstanding the limitations associated with this model, our data suggest that the optimal timing of folate intervention must be established before folate supplementation can be used as a safe chemopreventive agent against colorectal cancer. PMID- 10866312 TI - Addition of matrix metalloproteinase inhibition to conventional cytotoxic therapy reduces tumor implantation and prolongs survival in a murine model of human pancreatic cancer. AB - Matrix metalloproteinases (MMPs) participate in basement membrane degradation, a critical step in invasion of cancer cells. We have previously shown that MMP inhibition of pancreatic cancers improves survival and decreases MMP production in vivo. The purpose of this study was to determine whether BB-94 was better than cytotoxic therapy and would increase the efficacy of cytotoxic therapy (gemcitabine) in a murine model of human pancreatic cancer. A human pancreatic adenocarcinoma cell line (HPAC) was injected into the pancreata of BALB/c nu/nu mice. The mice were randomized 7 days after cancer cell injection to receive vehicle control, BB-94, gemcitabine, or gemcitabine and BB-94 until death or sacrifice at 84 days. At necropsy, tumors were harvested, and the relative enzyme activities of MMP-2 and MMP-9 were determined by gelatin zymography. Active MMP-2 levels in serum were determined using an ELISA technique. Combination treatment with gemcitabine and BB-94 significantly reduced implantation rates and improved survival in mice with documented orthotopic tumors compared with either therapy alone or control. Tumor levels of active and latent MMP-2 were higher than those of MMP-9 in both treated and control mice. There was a significant reduction of tumor MMP-2 activity in mice treated with BB-94, gemcitabine, or gemcitabine and BB-94. Serum levels of active MMP-2 were reduced in all treated groups, with the greatest reduction occurring in mice treated with gemcitabine and BB-94. Combination therapy with gemcitabine and BB-94 reduces cancer implantation and improves survival compared to treatment with BB-94, gemcitabine, or vehicle control alone. MMP production was reduced in all treated groups, reflecting reduced tumor progression, which was particularly seen with combination therapy with gemcitabine and BB-94. This study supports combining MMP inhibition with cytotoxic therapy (gemcitabine) for patients with pancreatic cancer. PMID- 10866311 TI - Regrowth of 5-fluorouracil-treated human colon cancer cells is prevented by the combination of interferon gamma, indomethacin, and phenylbutyrate. AB - We previously reported that phenylbutyrate (PB), a differentiation agent, retarded the regrowth of fluoropyrimidine-treated HT29 cells to a greater extent in a well-differentiated subclone as compared with a poorly differentiated subclone (Y. Huang and S. Waxman, Clin. Cancer Res., 4: 2503-2509, 1998). To extend these results and to overcome the known heterogeneity of human colon carcinoma (HCC) cells, the effect of cytostatic agents reported to inhibit HCC growth [IFN-alpha and IFN-gamma, indomethacin, and PB alone or in combination] on clonogenicity and HCCs recovery from 5-fluorouracil (FUra) treatment was studied in eight different HCCs. IFN-alpha proved to be ineffective in all eight HCCs, whereas IFN-gamma induced marked growth inhibition in four HCCs that expressed wild-type K-ras. Despite large differences in HCC response to the other individual agents, strong growth inhibition was observed when PB was added in combination with indomethacin. The inhibition was even more pronounced when IFN gamma was included in the regimen. Most importantly, after treatment with the combination of three agents, the clonogenic potential was severely inhibited (92 100%) in the IFN-gamma-sensitive cell lines, whereas in the IFN-gamma-insensitive cell lines, comparable loss of clonogenecity was obtained when the cells were pretreated with FUra. As known and described in detail, the three cytostatic agents inhibit different processes necessary for cell growth, thus requiring the cells to repair multiple pathways to restore growth. The induction of STAT1 DNA binding activity by IFN-gamma and p21WAF1 by PB, alone or in combination, correlated with growth inhibition and loss of clonogenicity. The finding that the readily reversible growth inhibition and decrease in clonogenicity of FUra treated HCC are prolonged by subsequent treatment with the three cytostatic agents in all HCCs may be of clinical importance because FUra continues to be the most widely used cytotoxic agent in the treatment of colon carcinoma. PMID- 10866313 TI - Mitochondrial amplification of death signals determines thymidine kinase/ganciclovir-triggered activation of apoptosis. AB - Previous clinical experience shows that the efficacy of suicide gene transfer in tumor therapy is limited, resulting from inefficient gene transfer or alternatively, from intrinsic resistance of the tumor in vivo. Herpes simplex virus thymidine kinase/ganciclovir (TK/GCV), a paradigmatic suicide gene therapy system, has been described to exert its cytotoxic effect, at least in part, by inducing apoptosis in target cells. Here, we report that mitochondria amplify TK/GCV-induced apoptosis by regulating p53 accumulation and the effector phase of apoptosis. Treatment with TK/GCV led to mitochondrial perturbations including loss of the mitochondrial membrane potential and release of cytochrome c from mitochondria into the cytosol, inducing caspase activation and nuclear fragmentation. Inhibition of TK/GCV-induced mitochondrial perturbations by Bcl-2 overexpression or by the mitochondrion-specific inhibitor bongkrekic acid also strongly inhibited TK/GCV-induced activation of caspases and apoptosis. TK/GCV induced mitochondrial perturbations depended on caspase activity possibly initiated by death receptor signaling. Perturbation of mitochondrial function mediated accumulation of wild-type p53 protein, since Bcl-2 overexpression, bongkrekic acid, or inhibition of mitochondrial protein synthesis with chloramphenicol strongly reduced TK/GCV-induced accumulation of wild-type p53 protein. These findings suggest that TK/GCV therapy may be less efficient in tumors in which the mitochondrial amplification of TK/GCV-induced apoptosis is blocked, e.g., by Bcl-2 overexpression. Given the low efficacy of currently used gene therapy systems, our data on molecular mechanisms that regulate sensitivity or resistance toward TK/GCV-induced cytotoxicity might have important implications to improve the clinical application of suicide gene therapy. PMID- 10866314 TI - Cancer gene therapy by thyroid hormone-mediated expression of toxin genes. AB - Many of the strategies developed in the last few years to treat cancer by gene therapy are based on putative killer-suicide genes whose products convert a prodrug into a toxic compound. When the therapy is applied to humans, a vector carrying the killer gene is first inoculated into the tumor of the patient, who 1 week later receives the corresponding prodrug that will selectively kill the cells able to process it to its toxic derivative. A strategy that obviates the need for a prodrug to destroy the cancer cells would be preferable because the patient would only need one treatment instead of two consecutive ones. In the following study, we describe the construction of retroviral vectors in which a reporter or a toxin gene (either the Pseudomonas exotoxin or the Ricinus communis toxin, ricin) is placed under the control of the thyroid hormone (T3) regulatable promoter of the rat myelin basic protein (MBPp). We demonstrate that the expression of these genes under the control of MBPp is regulated by T3 in vitro and in vivo. In vitro, the MBPp is switched off when T3 is removed from the serum of the culture medium, allowing the production of retroviruses carrying the toxic gene. In vivo, the toxin gene bearing retroviruses is capable of eradicating experimentally induced brain tumors in Wistar rats. The gene therapy strategy described here does not require the use of a prodrug to destroy the neoplastic cells. PMID- 10866315 TI - Agonistic properties and in vivo antitumor activity of the anti-CD40 antibody SGN 14. AB - Ligation of CD40 is essential for primary B-cell activation and expansion and yet has suppressive or apoptotic effects on some CD40-expressing neoplasia. SGN-14 is a monoclonal antibody that binds to the human CD40 receptor. Here we report that SGN-14, in the presence of interleukin 4, provided a modest level of stimulation of peripheral blood B cells, as measured by proliferation. Stimulation was greatly enhanced in the presence of nonproliferating CD40 ligand-expressing cells. The enhanced agonistic activity could be attributed to a dose-dependent increase in CD40L binding to CD40 in the presence of SGN-14. In contrast to its proliferative effect on primary B cells, SGN-14 inhibited the growth of B-cell derived tumor lines in vitro, and this growth inhibition was enhanced in the presence of CD40L-expressing cells. In vivo, SGN-14 showed significant antitumor activity in treating human B-cell lymphoma and multiple myeloma xenografted severe combined immunodeficient mice. Antitumor activity was not diminished by blunting murine natural killer activity, suggesting that CD40 ligation contributes to the antitumor efficacy of SGN-14. On the basis of these activities, SGN-14 is being pursued for therapeutic use in treating patients with CD40-expressing hematological malignancies. PMID- 10866316 TI - In vivo antitumor activity and host toxicity of methoxymorpholinyl doxorubicin: role of cytochrome P450 3A. AB - Methoxymorpholinyl doxorubicin (MMDX; PNU 152243) is a promising doxorubicin derivative currently undergoing clinical evaluation. Previous in vitro studies suggested that the compound undergoes hepatic biotransformation by cytochrome P450 (CYP) 3A into a more cytotoxic metabolite(s). The present study examined the role of CYP3A-mediated metabolism in the in vivo antitumor activity and host toxicity of MMDX in the mouse model and investigated the potential for increasing the therapeutic effectiveness of the drug by inducing its hepatic CYP-catalyzed activation. We found that MMDX cytotoxicity for cultured M5076 tumor cells was potentiated 22-fold by preincubating the drug with NADPH-supplemented liver microsomes from untreated C57BL/6 female mice. A greater (50-fold) potentiation of MMDX cytotoxicity was observed after its preincubation with liver microsomes isolated from animals pretreated with the prototypical CYP3A inducer pregnenolone 16alpha-carbonitrile. In contrast, in vivo administration of the selective CYP3A inhibitor troleandomycin (TAO) reduced both potentiation of MMDX cytotoxicity and the rate of CYP3A-catalyzed N-demethylation of erythromycin by isolated liver microsomes (55.5 and 49% reduction, respectively). In vivo antitumor activity experiments revealed that TAO completely suppressed the ability of 90 microg/kg MMDX i.v., a dose close to the LD10, to delay growth of s.c. M5076 tumors in C57BL/6 mice and to prolong survival of DBA/2 mice with disseminated L1210 leukemia. Moreover, TAO administration markedly inhibited the therapeutic efficacy of 90 microg/kg MMDX i.v. in mice bearing experimental M5076 liver metastases; a complete loss of MMDX activity was observed in liver metastases bearing animals receiving 40 microg/kg MMDX i.v. plus TAO. However, pregnenolone 16alpha-carbonitrile pretreatment failed to enhance MMDX activity in mice bearing either s.c. M5076 tumors or experimental M5076 liver metastases. Additional experiments carried out in healthy C57BL/6 mice showed that TAO markedly inhibited MMDX-induced myelosuppression and protected the animals against lethal doses of MMDX. Taken together, these findings demonstrate that an active metabolite(s) of MMDX synthesized via CYP3A contributes significantly to its in vivo antitumor activity and host toxicity. PMID- 10866317 TI - Differences in dendritic cells stimulated in vivo by tumors engineered to secrete granulocyte-macrophage colony-stimulating factor or Flt3-ligand. AB - Both granulocyte-macrophage colony-stimulating factor (GM-CSF) and flt3-ligand (FL) induce the development of dendritic cells (DCs). To compare the functional properties of DCs stimulated by these cytokines in vivo, we used retroviral mediated gene transfer to generate murine tumor cells secreting high levels of each molecule. Injection of tumor cells expressing either GM-CSF or FL resulted in the dramatic increase of CD11c+ cells in the spleen and tumor infiltrate. However, vaccination with irradiated, GM-CSF-secreting tumor cells stimulated more potent antitumor immunity than vaccination with irradiated, FL-secreting tumor cells. The superior antitumor immunity elicited by GM-CSF involved a broad T cell cytokine response, in contrast to the limited Thl response elicited by FL. DCs generated by GM-CSF were CD8alpha- and expressed higher levels of B7-1 and CD1d than DCs cells generated by FL. Injection sites of metastatic melanoma patients vaccinated with irradiated, autologous tumor cells engineered to secrete GM-CSF demonstrated similar, dense infiltrates of DCs expressing high levels of B7-1. These findings reveal critical differences in the abilities of GM-CSF and FL to enhance the function of DCs in vivo and have important implications for the crafting of tumor vaccines. PMID- 10866318 TI - Genetically modified dendritic cells prime autoreactive T cells through a pathway independent of CD40L and interleukin 12: implications for cancer vaccines. AB - Genetic immunization through ex vivo transduction of dendritic cells has been suggested as an effective approach to enhance antitumor immunity by activating both CD4+ and CD8+ T cells. Immunizing mice with dendritic cells transduced with an adenovirus expressing the human melanoma antigen glycoprotein 100 (DCAdhgp100) as a cancer vaccine, we demonstrated complete protective immunity and a potent CTL response against melanomas expressing murine glycoprotein 100 in a CD4+ cell dependent manner. Surprisingly, however, effective tumor rejection was not the result of cooperation between CD4+ and CD8+ T cells. Protective immunity was completely lost when CD4+ cells were depleted immediately before tumor challenge, whereas it was unaffected by removal of CD8+ cells, establishing a principal role for CD4+ cells in the effector phase of tumor rejection. Neither protective immunity nor CTL generation in this model required interleukin 12, in spite of high levels of IFN-gamma secretion by tumor-reactive T cells. Most notably, the DCAdhgp100 vaccine could elicit protective antitumor CD4+ cells in the absence of CD40 ligand, although it does not bypass the need for CD40-mediated signals to generate melanoma-reactive CTLs. Thus, in contrast to the current thinking that the optimal cancer vaccine should include determinants for both CD4+ and CD8+ cells, the potency of the DCAdhgp100 vaccine appears to be a result of its ability to directly prime autoreactive CD4+ cells through a process that does not require interleukin 12 and CD40 signals. PMID- 10866319 TI - Construction, production, and characterization of humanized anti-Lewis Y monoclonal antibody 3S193 for targeted immunotherapy of solid tumors. AB - The Lewis Y (Ley) antigen is a blood group-related antigen that is expressed in a high proportion of epithelial cancers (including breast, colon, ovary, and lung cancer) and is an attractive target for monoclonal antibody-directed therapy. The murine monoclonal 3S193 (IgG3) was generated in BALB/c mice by immunization with Ley-expressing cells of the MCF-7 breast carcinoma cell-line. The murine 3S193 showed high specificity for Ley in ELISA tests with synthetic Ley and Ley containing glycoproteins and glycolipids and also reacted strongly in rosetting assays and cytotoxic tests with Ley-expressing cells. We generated a humanized form of the murine 3S193 antibody by linking cDNA sequences encoding the variable region of murine 3S913 with frameworks of the human KOL heavy chain and REI K chain. The genes for the humanized 3S193 monoclonal antibody IgG1 were transfected into mouse myeloma NS0 cells and cloned for the establishment of high antibody-producing colonies. Humanized 3S193 antibody was subsequently produced through in vitro culture and under good manufacturing practice conditions using hollow-fiber bioreactors. The purified humanized 3S193 (hu3S193) was subsequently characterized and validated for use in preliminary immunotherapy investigations. hu3S193 reacted specifically with Ley antigen, with similar avidity to the murine form. hu3S193 demonstrated potent immune effector function, with higher antibody dependent cell-mediated cytotoxicity than its murine counterpart and potent complement-dependent cytotoxicity (ED50, 1.0 microg/ml). The in vivo immunotherapeutic potential of hu3S193 was assessed in a human breast xenograft model using MCF-7, Ley-positive cells. Six i.v. doses of up to 1 mg of hu3S193 were administered to animals bearing established tumors (120-130 mm3) with no significant effect on tumor growth. In contrast, in an MCF-7 xenograft preventive model, a 1-mg hu3S193 dosage schedule was able to significantly slow tumor growth compared with placebo and isotype-matched control IgG1 antibody. hu3S193 has promise for immunotherapy of Ley-positive tumors and is currently entering Phase I clinical trials. PMID- 10866320 TI - Epigenetic regulation of the MGMT and hMSH6 DNA repair genes in cells resistant to methylating agents. AB - We investigated the relationship between DNA cytosine methylation and the expression of two genes associated with resistance to DNA methylation damage. Variants of RajiMex- cells acquired resistance to N-methyl-N-nitrosourea by either reactivating a previously silent O6-methylguanine-DNA methyltransferase (MGMT) gene or by repressing the hMSH6 mismatch repair gene. DNA sequencing and measurements of mRNA and enzyme levels revealed that MGMT activity was not correlated with methylation of the core MGMT promoter. Treatment with the demethylating agent 5-azadeoxycytidine reduced MGMT mRNA and enzyme levels, indicating that methylation of some nonpromoter sequences may be required for MGMT gene expression. In contrast, both hMSH6 mRNA and protein levels were increased by 5-azadeoxycytidine treatment of an N-methyl-N-nitrosourea-resistant variant that did not express detectable hMSH6, which implies that this gene was transcriptionally silenced by cytosine methylation. This could be substantiated by in vitro modification of the CpG sites in the hMSH6 promoter with restriction methylase M.SssI, which abolished the transcription of a reporter gene under its control in a transient transfection assay. Taken together, our data show that treatment with chemical methylating agents alters gene expression patterns through increased CpG methylation of genomic DNA, and thereby permits the emergence and selection of clones that are resistant to these agents due to increased repair or tolerance of O6-methylguanine. PMID- 10866321 TI - Unique anti-activator protein-1 activity of retinoic acid receptor beta. AB - The anticancer effects of retinoids are mainly mediated by two classes of nuclear receptors, the retinoic acid receptors (RARs) and retinoid X receptors (RXRs), which are encoded by three distinct genes (alpha, beta, and gamma). Recent studies have demonstrated that RARbeta plays a critical role in mediating anticancer effects of retinoids. However, how RARbeta exerts its potent anticancer effects remains largely unknown. In this study, we investigated anti Activator Protein-1 (AP-1) activity of RARbeta. In a transient transfection assay, all three RAR subtypes, RARalpha, RARbeta, and RARgamma, could effectively inhibit phorbol ester 12-O-tetradecanoylphorbol-13-acetate-induced AP-1 activity and the activity of oncogenes c-Jun and c-Fos on AP-1 containing reporter genes in the presence of retinoic acid (RA). However, RARbeta showed a strong RA independent inhibition of AP-1 activity, whereas inhibition of AP-1 activity by RARalpha and RARgamma was RA dependent. By using several hybrid receptors that contain either the COOH-terminal portion or the NH2-terminal portion of RARbeta, we demonstrated that the NH2-terminal portion of RARbeta, the A/B domain, was mainly responsible for the RA-independent inhibition of AP-1 activity. This activity was not attributable to constitutive AF-1 activity of RARbeta, because it did not activate several RA response element-containing reporter genes. In addition, inhibition of histone deacetylase activity by trichostatin A did not overcome the inhibitory effect of RARbeta. In cancer cells, stable transfection of RARbeta exhibited strong inhibition of AP-1 activity, even in the absence of RA. Moreover, expression of endogenous AP-1-responsive gene collagenase I was strongly repressed in cancer cells stably transfected with RARbeta. In studying the antitransforming activity of RARbeta, we observed that the growth of breast cancer MDA-MB231 cells in soft agar was significantly repressed in a RA independent manner when cells were stably transfected with RARbeta but not RARalpha. Together, our results demonstrate that RARbeta may exert its potent anticancer effect in part through its unique anti-AP-1 activity. PMID- 10866322 TI - Rapid and reliable quantification of minimal residual disease in acute lymphoblastic leukemia using rearranged immunoglobulin and T-cell receptor loci by LightCycler technology. AB - The detection of minimal residual disease (MRD) using immunoglobulin and T-cell receptor (TCR) rearrangements as PCR targets provides important prognostic information on the in vivo effectiveness of treatment in acute lymphoblastic leukemia (ALL). Here we report on the real-time quantification of MRD in 25 ALL patients using LightCycler technology. We designed and adapted allele-specific oligonucleotide (ASO)-PCR protocols that enabled the detection of >90% of the IGH, IGK, TCRD, and TCRG rearrangements observed in ALL patients. In all patients, at least two suitable markers could be identified (average, 3.4 markers/patient). We applied ASO-PCR with 35 immunoglobulin and TCR rearrangements (11 IGH, 6 IGK, 12 TCRG, and 6 TCRD) and compared the sensitivity and practicability of the LightCycler strategy with conventional ASO-PCR on a block thermocycler followed by quantification with gel electrophoresis. The LightCycler measured leukemia-specific PCR products at each cycle (real-time) by staining the PCR product with the DNA-binding dye SYBR Green I. LightCycler technology showed a higher sensitivity than the conventional method in eight cases, whereas the sensitivity of the other markers matched exactly. The detection level varied between 10(-4) and 10(-6) leukemic cells. Furthermore, we determined the MRD status of 27 bone marrow follow-up samples from 15 ALL patients by both methods and revealed comparable results. However, the LightCycler also allowed accurate quantification in samples containing relatively high levels (>10(-3)) of residual leukemia cells. The conventional ASO-PCR technique comprises various laborious and time-consuming PCR experiments and post PCR steps to determine the number of cycles with the optimal linearity and sensitivity of the PCR. Real-time quantification through LightCycler technology obviates these post-PCR steps, provides the highest sensitivity via software analysis, and therefore represents a rapid, reliable, sensitive, and cost effective technique for the routine monitoring of MRD in ALL patients. PMID- 10866323 TI - Inhibition of the interferon-gamma/signal transducers and activators of transcription (STAT) pathway by hypermethylation at a STAT-binding site in the p21WAF1 promoter region. AB - Expression of the cyclin-dependent kinase inhibitor p21WAF1 can be up-regulated by activation of signal transducers and activators of transcription (STAT) proteins in response to IFN-gamma. In this study, we examined CpG methylation at the p21WAF1 promoter region in rhabdomyosarcomas (RMSs) using Southern blot analysis with the methylation-sensitive restriction enzyme HpaII. Sis-inducible element (SIE)-1, a STAT-responsive element located upstream of the p21 WAF1 CpG island, was completely methylated at an internal CpG in 13 of 26 (50%) primary RMS tumors and 2 of 5 RMS cell lines. In contrast, all normal tissues examined showed a partial methylation pattern at SIE-1. Complete methylation within SIE-1 strongly correlated with decreased p21WAF1 mRNA expression in RMS. We further studied the effects of SIE-1 hypermethylation on p21WAF1 induction by STAT activation. CpG methylation within SIE-1 significantly inhibited binding of activated STAT1 in electrophoretic mobility shift assays and abrogated STAT mediated transcription activation in response to IFN-gamma in luciferase reporter gene assays. Activation of STAT1 in response to IFN-gamma resulted in increased p21WAF1 expression and growth suppression in RMS cells containing unmethylated SIE-1 but failed to induce p21WAF1 or growth inhibition in RD and A673 cells, both of which were completely methylated within SIE-1. However, demethylation at SIE-1, induced by a demethylating agent 5-aza-2'-deoxycytidine, reactivated p21WAF1 expression and restored the responsiveness to IFN-gamma in RD cells. Our results indicate a mechanism by which altered DNA methylation in the p21 WAF1 promoter region, by precluding STAT1 binding to SIE-1, directly inhibits the p21WAF1 induction and cell growth regulation through the IFN-gamma/STAT signaling pathway in RMS cells. PMID- 10866324 TI - Role for ATM in DNA damage-induced phosphorylation of BRCA1. AB - The human genetic disorder ataxia-telangiectasia is characterized by immunodeficiency, progressive cerebellar ataxia, radiosensitivity, cell cycle checkpoint defects, and cancer predisposition. The gene product [ataxia telangiectasia mutation (ATM)] mutated in this syndrome is a component of the DNA damage detection pathway. Loss of ATM function in human and mouse cells causes defects in DNA repair and cell cycle checkpoint control and, not surprisingly, humans and mice with compromised ATM function are prone to cancers. An excess of breast cancer in the relatives of ataxia-telangiectasia patients has also been reported by epidemiological studies. Predisposition to breast and ovarian cancers is also observed in women with germline mutations in BRCA1, a tumor suppressor gene. BRCA1 is a nuclear protein with a cell cycle-regulated expression pattern and is hyperphosphorylated in response to DNA-damaging agents. Here we show that rapid ionizing radiation-induced in vivo phosphorylation of BRCA1 requires the presence of functional ATM protein. Furthermore, we show that ATM interacts with BRCA1, and this association is enhanced by radiation. We also demonstrate that BRCA1 is a substrate of ATM kinase in vitro and in vivo. Using phospho-specific antibodies against serines 1387, 1423, and 1457 of BRCA1, we demonstrate radiation-induced, ATM-dependent phosphorylation of BRCA1 at these sites. These findings show that BRCA1 is regulated by an ATM-dependent mechanism as a part of the cellular response to DNA damage. This interaction between ATM and BRCA1 argues in favor of the involvement of particular aspects of ATM function in breast cancer predisposition. PMID- 10866325 TI - Arginase activity in human breast cancer cell lines: N(omega)-hydroxy-L-arginine selectively inhibits cell proliferation and induces apoptosis in MDA-MB-468 cells. AB - L-Arginine is the common substrate for two enzymes, arginase and nitric oxide synthase (NOS). Arginase converts L-arginine to L-ornithine, which is the precursor of polyamines, which are essential components of cell proliferation. NOS converts L-arginine to produce NO, which inhibits proliferation of many cell lines. Various human breast cancer cell lines were initially screened for the presence of arginase and NOS. Two cell lines, BT-474 and MDA-MB-468, were found to have relatively high arginase activity and very low NOS activity. Another cell line, ZR-75-30, had the highest NOS activity and comparatively low arginase activity. The basal proliferation rates of MDA-MB-468 and BT-474 were found to be higher than the ZR-75-30 cell line. N-Hydroxy-L-arginine (NOHA), a stable intermediate product formed during conversion of L-arginine to NO, inhibited proliferation of the high arginase-expressing MDA-MB-468 cells and induced apoptosis after 48 h. NOHA arrested these cells in the S phase, increased the expression of p21, and reduced spermine content. These effects of NOHA were not observed in the ZR-75-30 cell line, which expresses high NOS and relatively low arginase. The effects of NOHA were antagonized in the presence of L-ornithine (500 microM), which suggests that in MDA-MB-468 cell line, the arginase pathway is very important for cell proliferation. Inhibition of the arginase pathway led to depletion of intracellular spermine and apoptosis as observed by terminal deoxynucleotidyl transferase (Tdt)-mediated nick end labeling assay and induction of caspase 3. In contrast, the ZR-75-30 cell line maintained its viability and its L-ornithine and spermine levels in the presence of NOHA. We conclude that NOHA has antiproliferative and apoptotic actions on arginase-expressing human breast cancer cells that are independent of NO. PMID- 10866326 TI - New evidence for an extra-hepatic role of N-acetylglucosaminyltransferase III in the progression of diethylnitrosamine-induced liver tumors in mice. AB - N-acetylglucosaminyltransferase III (GlcNAc-TIII) is encoded by the Mgat3 gene and catalyzes the addition of the bisecting GlcNAc to the core of N-glycans. Mice lacking GlcNAc-TIII due to the insertion mutation Mgat3tmlPst (termed Mgat3neo), exhibit retarded progression of liver tumors induced by diethylnitrosamine (DEN; M. Bhaumik et al, Cancer Res., 58: 2881-2887, 1998). This phenotype seemed to be due to a reduction, in activity or amount, of a circulating glycoprotein(s) that enhances DEN-induced liver tumor progression. Here, we provide new evidence to support this hypothesis. First, we show that mice with a deletion mutation of the Mgat3 gene coding exon (Mgat3tmlJxm, termed Mgat3delta) also exhibit retarded progression of DEN-induced liver tumors. At 7 months there was a significant decrease in liver weight (approximately 27%; P < 0.01), reflecting reduced tumor burden in Mgat3delta/delta mice. In addition, tumors were generally fewer and smaller, and histological changes were less severe in Mgat3delta/delta livers. Therefore, tumor progression is retarded in mice with two different null mutations in the Mgat3 gene. Second, we show that the development of DEN-induced tumors is unaltered by high levels of GlcNAc-TIII in the liver of transgenic mice. The Mgat3 gene coding exon under the control of the major urinary protein (MUP) promoter was used to generate transgenic mice that express GlcNAc-TIII in liver. Following DEN injection and phenobarbitol treatment, however, no significant differences were observed between MUP/Mgat3 transgenic and control mice in either tumor numbers or liver weight. The combined data provide strong evidence that retarded progression of tumors in mice lacking GlcNAc-TIII is due to the absence of the bisecting GlcNAc residue on N-glycans of a circulating glycoprotein(s) from a tissue other than liver. PMID- 10866327 TI - Acute beta blocker overdose: factors associated with the development of cardiovascular morbidity. AB - OBJECTIVE: To identify factors in exposures to beta blockers (beta-adrenergic receptor antagonists) that are associated with the development of cardiovascular morbidity and contribute to disposition decisions from the emergency department. METHODS: Prospective cohort of 280 beta blocker exposures reported to 2 regional poison centers. Multiple logistic regression was used to determine association of various clinical factors and outcome. RESULTS: In this series of beta blocker exposures, 41 (15%) developed cardiovascular morbidity and 4 (1.4%) died. A history of cardioactive coingestant was the only factor significantly associated with the development of cardiovascular morbidity (p < .05). When cases reporting cardioactive coingestants were excluded, a history of ingesting a beta blocker with membrane stabilizing activity was significantly associated with the development of cardiovascular morbidity (p < .05). All those in whom the timing of symptoms could be determined, developed symptoms within 6 hours of ingestion. CONCLUSIONS: The single most important factor associated with the development of cardiovascular morbidity in beta blocker ingestion is a history of a cardioactive coingestant, primarily calcium channel blockers, cyclic antidepressants, and neuroleptics. In the absence of such coingestion, exposure to a beta blocker with membrane stabilizing activity is associated with an increased risk of cardiovascular morbidity. Beta blocker ingestion is unlikely to result in symptoms if the patient remains asymptomatic for 6 hours after the time of ingestion. PMID- 10866328 TI - Cardiac and hemodynamic assessment of patients with cocaine-associated chest pain syndromes. AB - BACKGROUND: Animal and human experimental studies have yielded conflicted data regarding the effects of cocaine on cardiovascular function. We studied the cardiac and hemodynamic profiles in emergency department chest pain patients following recent cocaine use. METHODS: After obtaining informed consent, emergency department patients who presented with a chief complaint of chest pain and cocaine use within 24 hours of arrival were prospectively enrolled. All patients underwent a structured 40-item history and physical examination and were placed on the IQ Noninvasive Hemodynamic Surveillance System (Renaissance Technology, Inc., Newton, PA), a validated transthoracic cardiac output monitor. The principal measurements obtained included cardiac output, cardiac index, and stroke volume. Data were analyzed with standard descriptive techniques. RESULTS: Twenty-seven patients were enrolled (median age, 37 years [range, 23-54]; 74% male). Patients used a mean of $200 worth of cocaine, usually crack (67%). Patients had a history of tobacco use (82%), prior myocardial infarction (33%), and prior cocaine-associated chest pain (67%). The median (interquartile range; IQR) for the hemodynamic parameters were: mean arterial blood pressure 92 mm Hg (IQR 85-100); heart rate 83/min (IQR 72-98); cardiac output 6.9 L/min (IQR 5.1 7.2); cardiac index 3.2 L/min/m2 (IQR 2.4-4.0); stroke volume 78 mL/beat (IQR 64 93). CONCLUSION: Most emergency department patients with cocaine-associated chest pain have normal cardiac profiles at the time of presentation. The negative inotropic effects of high doses of cocaine observed in animal models do not appear to be present in patients who develop chest pain after using recreational doses of cocaine. PMID- 10866329 TI - Unintentional acetaminophen ingestion in children and the potential for hepatotoxicity. AB - OBJECTIVE: Children who unintentionally ingest acetaminophen are often referred to health care facilities for evaluation. Criteria for referral are not well defined and the vast majority of these exposures result in nontoxic serum concentrations. The objective of this study was to determine the incidence of potentially hepatotoxic serum concentrations and to more clearly define referral criteria for these patients. METHODS: A prospective evaluation of all childhood (age 1-72 months) single ingestions of acetaminophen-containing products was performed by the Utah Poison Control Center. All patients ingesting 140 mg/ kg or greater or an unknown amount were referred for medical evaluation. Patients who ingested greater than 100 mg/kg were advised to administer syrup of ipecac at home if less than 1 hour since ingestion. Activated charcoal was recommended within 2 hours of ingestion if the patient was already at a health care facility. The potential for hepatotoxicity was assessed according to the Rumack-Matthew nomogram. RESULTS: Inclusion criteria were met by 1015 patients. The mean age was 28 +/- 12 months and mean dose was 213 +/- 148 mg/ kg. Decontamination with ipecac, gastric lavage, or activated charcoal within 2 hours of ingestion occurred in 81% of patients ingesting greater than 140 mg/kg or an unknown amount. Six patients (0.59%, 95% CI 0.12-1.16%) had "possible" or "probable" hepatotoxic serum concentrations and all had ingested greater than 200 mg/kg or an unknown amount. There were 423 patients who ingested between 100 and 200 mg/kg and none had potentially hepatotoxic serum concentrations (upper 95% CL 0.71%). CONCLUSIONS: Children who ingest between 140-200 mg/kg of acetaminophen and demonstrate ipecac-induced emesis within 60 minutes may be safely managed at home. Patients ingesting greater than 200 mg/kg or an unknown amount should be referred for a serum acetaminophen concentration. PMID- 10866330 TI - Pancreatic injury following acute methanol poisoning. AB - BACKGROUND: Methanol ingestion is a cause of potentially life-threatening poisoning with numerous systemic manifestations. Clinicians may overlook the possibility of acute pancreatitis in this setting. The objective of this paper is to document the incidence of this complication in a series of 22 patients and to discuss the respective role of methanol and ethanol in its pathogenesis. CASE REPORT: A 54-year-old woman developed acute necrotizing pancreatitis following acute methanol poisoning. She was treated by hemodialysis, ethanol infusion, and folinic acid, but, despite maximal supportive therapy, she died from multiple organ failure 54 hours after the ingestion. CASE SERIES: In a series of 22 consecutive patients admitted with a diagnosis of acute methanol poisoning, we found evidence of pancreatic damage in 11 patients. The abnormalities were present from admission and before ethanol therapy in 7 cases and developed after ethanol therapy in 4 cases. Seven patients had a history of chronic ethanol abuse, but no patient had previously suffered from acute or chronic pancreatitis. Three patients presented moderate-to-severe acute pancreatitis according to clinical and radiological criteria and required aggressive supportive therapy including peritoneal dialysis. One patient died from the direct consequences of acute necrotizing pancreatitis and 2 fully recovered from this event. Three patients evolved to brain death; autopsy revealed hemorrhagic lesions in the pancreas in only 1 case. CONCLUSIONS: Clinical, biological, and radiographic signs of acute pancreatic injury may be more common than previously realized. Acute methanol poisoning appears to produce pancreatic injury, although antidotal treatment with ethanol or prior chronic ethanol abuse may be contributing factors. Because ethanol treatment may complicate the pancreatic injury, fomepizole (4-methylpyrazole) may be the preferable antidote in acute methanol poisoning. PMID- 10866332 TI - A survey of the expectations of New Mexico emergency physicians regarding the services of the New Mexico Poison and Drug Information Center. AB - OBJECTIVE: Physicians have been surveyed concerning their satisfaction with poison center services but have never been questioned regarding their expectations. This study was conducted to clarify the expectations of emergency physicians in New Mexico regarding the service of their regional poison center, the New Mexico Poison and Drug Information Center. DESIGN: Five New Mexico emergency department physicians were interviewed about their expectations when calling the New Mexico Poison and Drug Information Center. Their responses were combined with unique, additional expectations identified by the New Mexico Poison and Drug Information Center staff in order to develop a 62-item physician expectation mail survey instrument. Respondents were asked to rank the importance of each service expectation on a 5-point Likert scale (1 = not important, 5 = extremely important). RESULTS: A usable return rate of 60% was achieved (104 surveys). Fifty-eight (94%) expectations had a mean importance rating of > or = 3 (important). Ninety-four percent of the expectations included on the survey were being provided by the New Mexico Poison and Drug Information Center. CONCLUSION: Currently, there is a good fit between New Mexico emergency physician expectations and New Mexico Poison and Drug Information Center services provided. The instrument identified several areas where service was expected but not provided (Internet access to poison center, provision of industrial/occupational toxicology services, provision of tele-medicine capability) and where service was provided but not expected (drug dosing information). Surveying can be a valuable tool for clarifying expectations for poison center services. PMID- 10866331 TI - Ecbalium elaterium (squirting cucumber)--remedy or poison? AB - BACKGROUND: Ecbalium elaterium is a plant endemic to the Mediterranean basin. Its roots and cucumber-shaped fruit have been used in folk medicine since antiquity. The alleged uses of the fruit juice are as a potent cathartic, analgesic, and antiinflammatory agent. Cucurbitacin B, a triterpene derivative is the active antiinflammatory principal. PATIENTS: We present a series of 13 patients who were exposed to the juice of Ecbalium elaterium in its natural form. In 3 patients, exposure was intranasal for the treatment of sinusitis or liver cirrhosis. In 3 other cases, children ingested the fruit unwittingly. In 6 patients, exposure was ocular and, in one, dermal. Within minutes of exposure, the patients exhibited irritation of mucous membranes at various degrees of severity manifested as edema of pharynx, dyspnea, drooling, dysphagia, vomiting, conjunctivitis, corneal edema, and erosion, depending on the route of the exposure. Recovery began within several to 24 hours after administration of oxygen, steroids, antihistamines, and beta-2-agonists. Ocular exposures responded to topical steroid and antibiotic eyedrops within a few days. The toddler with the dermal exposure remained asymptomatic. CONCLUSION: Exposure to the juice of Ecbalium elaterium, mainly in its undiluted form, may cause irritation of mucous membranes, supposedly of inflammatory nature. Patients exposed orally or intranasally should be closely followed for upper airway obstruction. Patients exposed ocularly should have their eyes promptly irrigated to prevent corneal and conjunctival injury. PMID- 10866333 TI - High-efficiency dialysis for carbamazepine overdose. AB - BACKGROUND: Carbamazepine intoxication is associated with seizures, coma, arrhythmias, and death. In acute intoxications, charcoal hemoperfusion enhances removal of the drug but is associated with thrombocytopenia, coagulopathy, hypothermia, and hypocalcemia. Alternatively, high-efficiency hemodialysis can be used without the side effects of charcoal hemoperfusion. CASE REPORT: We report an 18-month-old comatose, convulsing child with plasma carbamazepine 27 microg/mL treated with high efficiency hemodialysis. Therapeutic carbamazepine levels were obtained after 4.5 hours of high-efficiency hemodialysis. The patient developed no untoward side effects, improved clinically, and was subsequently discharged home without sequelae. We conclude that high-efficiency hemodialysis is a safe, effective alternative to charcoal hemoperfusion in the pediatric population. PMID- 10866334 TI - Monitoring of clozapine and norclozapine plasma concentration-time curves in acute overdose. AB - CASE REPORT: A 40-year-old schizophrenic man was found unconscious, with constricted pupils, sinus tachycardia, and twitching of the limbs. There were signs of lung infection, which was treated with antibiotics, and mild rhabdomyolysis. He regained consciousness over 8 hours, and reported taking 3-4 g clozapine. Recovery was uneventful. Measured peak clozapine and norclozapine concentrations were 3.53 mg/L and 0.70 mg/L, respectively. The concentration-time curves were biphasic, with secondary peaks at approximately 36 hours postadmission. Terminal elimination half-lives were 16.9 hours and 22.5 hours for clozapine and norclozapine, respectively. CONCLUSION: Clozapine and its metabolite norclozapine can show biphasic plasma concentration-time curves after overdosage. PMID- 10866335 TI - Use of clonidine for chemical submission. AB - CASE REPORT: An East-European prostitute in Amsterdam robbed several victims, after having sedated them with clonidine solution (available as plastic ampoules of eyedrops) added to her victims' drinks. One victim was hospitalized. His symptoms included bradycardia, hypotension, hypothermia, pallor, cyanosis, and impaired consciousness. Treatment included isoprenaline for 28 hours. The victim was released from hospital the next day. In court, the female offender confessed and was sentenced to prison for 3 1/2 years. She may have administered doses as high as 8 mg clonidine. PMID- 10866336 TI - Lithium: thyroid effects and altered renal handling. AB - BACKGROUND: Lithium is frequently used in the treatment of bipolar affective disorder, and is widely known to affect thyroid function, most commonly resulting in hypothyroidism and goiter. Less well-known is the association between lithium therapy and hyperthyroidism and the potential for lithium to moderate the effects of thyroxine at a cellular level. Lithium excretion relates principally to glomerular filtration rate and proximal tubule function. Thyroxine, through its effects on tubular function, alters lithium clearance such that thyroid disease may cause retention of lithium and subsequent toxicity. CASE REPORTS: We report 2 cases with lithium toxicity, both of whom were later found to be hyperthyroid. One patient developed thyroid storm following dialysis to remove lithium. The other received antithyroid medication early. Both suffered a protracted multifactorial delirium requiring intensive inpatient care. CONCLUSION: In addition to altering thyroid function, lithium therapy may mask the signs of hyperthyroidism by inducing cellular unresponsiveness. In some lithium-treated patients with biochemical hyperthyroidism, early antithyroid treatment may be appropriate. Altered renal tubular function induced by hyperthyroidism may result in retention of lithium and systemic toxicity. We propose induction of the proximal tubule sodium hydrogen antiporter as the relevant mechanism. PMID- 10866337 TI - Carbamazepine-associated severe left ventricular dysfunction. AB - INTRODUCTION: The cardiovascular effects of carbamazepine are well-known but left ventricular dysfunction is rarely reported. CASE REPORT: We describe 2 cases of severe carbamazepine-associated left ventricular dysfunction during massive self intoxications in young patients without preexistent cardiac disease. We compare our cases to the available literature and discuss the mechanisms implied in the development of left ventricular dysfunction following carbamazepine overdose. Bedside echocardiography was useful in both diagnosis and treatment. PMID- 10866338 TI - Falsely elevated digoxin level of 45.9 ng/mL due to interference from human antimouse antibody. AB - CASE REPORT: We report the case of a 77-year-old man who was admitted to the intensive care unit with a serum digoxin level of 45.9 ng/mL. The patient was hemodynamically stable throughout his hospital course and did not require antidigoxin antibody fragments. The elevated digoxin level was determined by subsequent testing to be falsely elevated by interference from human antimouse antibodies in his serum. A repeat digoxin measurement using an assay not affected by human antimouse antibodies indicated a level of 1.3 ng/mL. Newer digoxin assays are not affected by human antimouse antibody interference, but clinicians should be aware of possible human antimouse antibody interference with older digoxin assays and in other tests utilizing mouse monoclonal antibody reagents. PMID- 10866339 TI - Intravenous detergent poisoning. AB - CASE REPORT: In the literature regarding surfactant poisoning, the route of exposure has almost always been oral. We report a case in which about 40 mL of bath detergent for home use was self-injected. The primary pathophysiologic effects were relative hypovolemia and cardiac dysfunction. The patient experienced frequent ventricular tachycardia, acute renal failure, rhabdomyolysis, hemolysis, and coagulation dysfunction. Intensive care included the administration of antiarrythmial agents and hemodialysis. The patient survived and was discharged from our hospital without sequelae. PMID- 10866340 TI - Hypersensitivity myocarditis with ephedra use. PMID- 10866341 TI - Ephedra causes myocarditis. PMID- 10866342 TI - Homeopathic remedies for children: are they cause for concern? PMID- 10866343 TI - Is Star Trek over the hill? PMID- 10866344 TI - Biology and treatment of malignant glioma. AB - Malignant gliomas are a heterogeneous group of tumors that are associated with significant morbidity and mortality. The development of the malignant phenotype is the result of a complex series of events that influence gene expression, angiogenesis, and invasion and favor the growth of tumor cells. Currently, the management of malignant glioma consists of symptom control and cytoreduction with a combination of surgery, radiation therapy, and chemotherapy. However, there is no satisfactory regimen available for adjuvant or salvage chemotherapy for these neoplasms. An overview of the biologic mechanisms, grading systems, and treatment options for anaplastic astrocytoma and glioblastoma multiforme is presented. PMID- 10866345 TI - Clinical trial end points in malignant glioma: need for effective trial design strategy. AB - Although oncologists have devised a number of measures of therapeutic efficacy, the design and evaluation of clinical trials of therapy for malignant gliomas present special difficulties. These include comparability of study design, patient stratification, classification of tumor type, and outcome response measures. In addition, these tumors and their treatment have profound effects on patients' quality of life. Consequently, considerations of quality of life are an important factor in study design. These issues are addressed to facilitate the interpretation of current and future studies of emerging treatment options in patients with malignant gliomas. PMID- 10866346 TI - Issues in assessing and interpreting quality of life in patients with malignant glioma. AB - Although primary brain tumors are relatively uncommon types of adult cancer, their location and resistance to treatment can significantly affect the patient's physical and cognitive function. Consequently, quality of life (QOL) issues are extremely important in the design and evaluation of clinical trials of high-grade glioma treatment. Although a number of studies have examined QOL in patients with primary brain tumors, very little is known about QOL in these patients. These studies often use either poorly validated instruments or tools, such as the Karnofsky performance scale, that do not fully evaluate the effects of the tumor on QOL. Recent attempts to better evaluate QOL in brain tumor patients have led to the development of brain tumor-specific QOL instruments, such as the European Organization for Research and Treatment of Cancer's Quality of Life Questionnaire Brain Cancer Module and the Functional Assessment of Cancer Therapy-Brain, which offer more insight into the QOL aspects of cancer and its treatment. Instruments such as quality time without symptoms or toxicity (Q-TWiST) provide a means of integrating both quality of time of survival and absence of symptoms or toxicity. PMID- 10866347 TI - Temozolomide in malignant gliomas. AB - Glioblastoma multiforme and anaplastic astrocytoma are the most common primary central nervous system malignancies and are the major cause of morbidity/ mortality despite combined modality approaches. Temozolomide (TMZ), a novel, oral, second-generation alkylating agent, has demonstrated antitumor activity against a broad range of solid tumors and highly resistant malignancies, including high-grade glioma Temozolomide does not require hepatic metabolism for activation, rapidly penetrates the cerebrospinal fluid, and consistently demonstrates reproducible linear pharmacokinetics with approximately 100% oral bioavailability. In preliminary clinical studies, TMZ has demonstrated meaningful efficacy and an acceptable safety profile in the treatment of patients with malignant glioma. These results have been recently confirmed in three open-label, multi-institutional studies that evaluated the use of TMZ in 525 malignant glioma patients. These studies represent the largest evaluation of a single agent in patients with recurrent malignant gliomas and were rigorously controlled with strict, prospectively defined criteria for assessment of tumor response, central review of histology, and validated instruments to assess health-related quality of life. Temozolomide was effective in delaying disease progression and maintaining health-related quality of life. Temozolomide represents a promising new agent in the treatment of malignant glioma. PMID- 10866348 TI - Temozolomide in early stages of newly diagnosed malignant glioma and neoplastic meningitis. AB - Temozolomide is a novel, oral, second-generation alkylating agent. Preclinical and phase I/II studies have demonstrated its efficacy against newly diagnosed high-grade glioma and anaplastic astrocytoma. Its antineoplastic effect is accompanied by quality of life benefits in patients with these debilitating tumors. Neoplastic meningitis, a refractory disorder, develops in approximately 3% to 8% of patients with systemic cancers. Traditional approaches to leptomeningeal metastases, such as radiation and intrathecal chemotherapy, have limited success and a high degree of toxicity. Temozolomide offers a number of potential therapeutic advantages in this disorder, including activity against a wide spectrum of human cancers that produce neoplastic meningitis and penetration of the blood-brain barrier. Patients treated with temozolomide benefit from both its systemic and intracranial activity. Recently, intrathecal temozolomide has been shown to increase median survival in athymic rats bearing subarachnoid human malignant glioma xenografts. Its efficacy, convenient dosing, and predictable safety profile make it an ideal agent for future study of these difficult-to treat central nervous system malignancies. PMID- 10866349 TI - Future directions in the treatment of malignant gliomas with temozolomide. AB - Temozolomide (TMZ) is a new, orally administered, second-generation imidazotetrazine prodrug with essentially 100% oral bioavailability that has demonstrated meaningful efficacy and an acceptable safety profile in the treatment of patients with recurrent glioblastoma multiforme. Because of its unique properties and broad spectrum of anticancer activity, preliminary studies are being conducted to evaluate the efficacy of TMZ in combination with other chemotherapeutic agents, radiation, or immunotherapy. The presence of de novo or acquired resistance to alkylating agents exhibited by malignant gliomas represents a serious impediment in the treatment of these tumors. This review discusses the mechanism of action of TMZ and strategies for overcoming pathways of resistance to this promising agent, including the use of TMZ in combination with other chemotherapeutic agents or radiation therapy, and exploration of alternate dosing schedules. Studies that have evaluated some of these strategies indicate that TMZ is a useful therapeutic option in patients with high-grade gliomas. Alternative approaches, including the use of high-dose TMZ with bone marrow transplantation and in combination with gene therapy, will also be discussed. PMID- 10866350 TI - Human performance on negative slope schedules of points exchangeable for money: a failure of molar maximization. AB - Panel pressing was generated and maintained in 5 adult humans by schedules of points exchangeable for money. Following exposure to a variable-interval 30-s schedule and to a linear variable-interval 30-s schedule (which permitted points to accumulate in an unseen "store" in the absence of responding), subjects were exposed to a series of conditions with a point-subtraction contingency arranged conjointly with the linear variable-interval schedule. Specifically, points were added to the store according to the linear-variable interval 30-s schedule and were subtracted from the store according to a ratio schedule. Ratio value varied across conditions and was determined individually for each subject such that the subtraction contingency would result in an approximately 50% reduction in the rate of point delivery. Conditions that included the subtraction contingency were termed negative slope schedules because the feedback functions were negatively sloped across all response rates greater than the inverse of the variable interval schedule, in this case, two per minute. Overall response rates varied inversely with the subtraction ratio, indicating sensitivity to the negative slope conditions, but were in excess of that required by accounts based on strict maximization of overall reinforcement rate. Performance was also not well described by a matching-based account. Detailed analyses of response patterning revealed a consistent two-state pattern in which bursts of high-rate responding alternated with periods of prolonged pausing, perhaps reflecting the joint influence of local and overall reinforcement rates. PMID- 10866351 TI - Stimulus control and generalization of point-loss punishment with humans. AB - Two experiments demonstrated stimulus control and generalization of conditioned punishment with humans. In both studies, responses first were reinforced with points exchangeable for money on a variable-interval schedule in the presence of one line length (S(D)). Next, a second line length was introduced, and point loss followed every response in the presence of that line (S(D)p). In the final training condition, points were deducted at session end. Response rate was lower in the presence of the S(D)p despite equal rates of points for money in the presence of both stimuli. In generalization testing for Experiment 1, the two lines were included in a 10-line continuum; S(D)p fell in the middle and the trained SD was at one end. Lines were presented randomly, and point delivery and loss contingencies were as in training but with points available in the presence of all lines. For all subjects, response rates were lowest around S(D)p and increased towards the SD end of the continuum. Because testing included only one or two lines beyond S(D), this pattern did not rule out S(D) generalization. Thus, in Experiment 2, stimuli beyond S(D) were added to generalization tests. Response rates did not decrease as a function of distance from S(D), clarifying the demonstration of punishment generalization. PMID- 10866352 TI - Reinforcer control and human signal-detection performance. AB - Eight humans participated in a two-choice signal-detection task in which stimulus disparity was varied over four levels. Two procedures arranged asymmetrical numbers of reinforcers received for correct left- and right-key responses (the reinforcer ratio). The controlled procedure ensured that the obtained reinforcer ratio remained constant over changes in stimulus disparity, irrespective of subjects' performances. In the uncontrolled procedure, the asymmetrical reinforcer ratio could covary with subjects' performances. The receiver operating characteristic (ROC) patterns obtained from the controlled procedure approximated isobias functions predicted by criterion location measures of bias. The uncontrolled procedure produced variable ROC patterns that were somewhat like the isobias predictions made by likelihood ratio measures of bias; however, the obtained reinforcer ratio became more extreme as discriminability decreased. The obtained pattern of bias was directly related to the obtained reinforcer ratio. This research indicates that criterion location measures seem to be preferable indices of response bias. PMID- 10866353 TI - Progressive-ratio schedules: effects of later schedule requirements on earlier performances. AB - Four rats were studied with variants of a progressive-ratio schedule with a step size of 6 in which different terminal components followed completion of the 20th ratio: (a) a reversal of the progression, (b) a fixed-ratio 6 schedule, or (c) extinction. Responding in the progressive-ratio components of these schedules was compared to performances under conventional progressive-ratio baselines. Under baseline conditions, postreinforcement pauses increased exponentially as a function of increasing ratio size, whereas running rates showed modest declines. The procedure of linking the progressive-ratio schedule to the reversed progression or to the fixed-ratio component resulted in decreased pausing. Linking the progressive-ratio schedule to the extinction component had the opposite effect, that of producing weakened progressive-ratio performances as evidenced by increased pausing. Subjects whose responses were reinforced on half of the ratios also showed exponential increases; however, pauses were substantially shorter following ratios on which the reinforcer was omitted. The results suggested that progressive-ratio pausing reflects the influence of remote as well as local contingencies. PMID- 10866354 TI - An olfactory discrimination procedure for mice. AB - This paper describes an olfactory discrimination procedure for mice that is inexpensively implemented and leads to rapid discrimination learning. Mice were first trained to dig in small containers of sand to retrieve bits of buried chocolate. For discrimination training, two containers were presented simultaneously for eight trials per session. One container held sand mixed with cinnamon, and the other held sand mixed with nutmeg. Both containers were baited with chocolate buried in the sand. One odor was designated S+, and mice were allowed to dig and retrieve the chocolate from this container. The other odor was S-, and both containers were removed immediately if subjects began to dig in an S container. After meeting a two-session acquisition criterion, subjects were given a series of discrimination reversals. In Experiment 1, 12 Swiss-Webster mice (6 male and 6 female) acquired the olfactory discrimination in three to five sessions and completed 3 to 10 successive discrimination reversals within a 50 session testing limit. In Experiment 2, subjects were 14 Pah(enu2) mice, the mouse mutant for phenylketonuria; 7 were homozygotes in which the disorder was expressed (PKU), and 7 were heterozygotes with normal metabolism (non-PKU). Thirteen mice completed pretraining in four to seven sessions, acquisition required 3 to 12 sessions, and all mice completed at least three reversals. Learning rates were similar in PKU and non-PKU mice. We discuss issues related to implementation and several potentially useful procedural variations. PMID- 10866355 TI - Studies of wheel-running reinforcement: parameters of Herrnstein's (1970) response-strength equation vary with schedule order. AB - Six male Wistar rats were exposed to different orders of reinforcement schedules to investigate if estimates from Herrnstein's (1970) single-operant matching law equation would vary systematically with schedule order. Reinforcement schedules were arranged in orders of increasing and decreasing reinforcement rate. Subsequently, all rats were exposed to a single reinforcement schedule within a session to determine within-session changes in responding. For each condition, the operant was lever pressing and the reinforcing consequence was the opportunity to run for 15 s. Estimates of k and R(O) were higher when reinforcement schedules were arranged in order of increasing reinforcement rate. Within a session on a single reinforcement schedule, response rates increased between the beginning and the end of a session. A positive correlation between the difference in parameters between schedule orders and the difference in response rates within a session suggests that the within-session change in response rates may be related to the difference in the asymptotes. These results call into question the validity of parameter estimates from Herrnstein's (1970) equation when reinforcer efficacy changes within a session. PMID- 10866356 TI - Effects of sleep deprivation on free-operant avoidance. AB - Two studies examined effects of sleep deprivation on free-operant avoidance by rats. In Experiment 1, a 5-s shock-shock (SS) interval and 20-s response-shock (RS) interval produced baseline performances, which were reestablished after each experimental manipulation. Once baselines were established, animals were exposed to 24, 48, or 96 hr of sleep deprivation and equivalent periods of home cage and food restriction as a control condition. Compared to baseline, sleep deprivation increased response rates by increasing the proportion of brief interresponse times (IRTs); response rates changed little in the control conditions. Percentage of shocks avoided did not systematically change across conditions. In Experiment 2, the RS interval was manipulated (10, 20, and 40 s), while the SS interval (5 s) and level of sleep deprivation (48 hr) were held constant. Across RS intervals, sleep deprivation increased response rates via a shift toward brief IRTs. In addition, sleep deprivation increased the percentage of shocks avoided as an inverse function of RS intervals. PMID- 10866357 TI - Peptides and antitumor activity. Development and investigation of some peptides with antitumor activity. AB - We developed a group of synthetic analogs of GnRH and Somatostatin to inhibit the tumor growth of different kind. The GnRH analogs decreasing the gonadotroph and steroid hormone levels act on the hormone dependent tumors and influence their growth. One of the most effective antitumor analog was patented under the name FOLLIGEN which inhibited the breast cancer caused by DMBA in rats without any side-effects. Other inhibitory analogs of GnRH with long-lasting effect were effective in the treatment of breast, ovary and prostate tumors. Another analog [alpha-Asp(DEA)]6,Gln8-hGnRH showed a very low endocrine but high antitumor effect in both in vitro and in vivo experiments. Its tritium labeled derivative exhibited specific binding sites on human tumor cell lines. We synthesized the analogs of GnRH-III with effective selective antitumor activity which does not alter the ovarian cycle of rats but inhibits the colony-formation of human breast cancer cell lines and has a significant antiproliferative effect. We also synthesized conjugates of potent GnRH analogs with a branched chain polylysine backbone which induce a 33-35% decrease of cell numbers of MCF-7 and MDA-MB-231 human breast cancer cell lines and 45-50% inhibition of cell proliferation. Another conjugate decreased the tumor growth of MDA-MB-231 xenografts by 80% in a treatment of 9 weeks and even tumor free animals could be found among the ones treated. Using these radiolabeled peptide hormone analogs we found that human tumor cell lines and xenografts specifically bind the GnRH conjugates. We also synthesized a series of Somatostatin analogs which inhibit tyrosine kinases and the growth of several breast, prostate and colon tumor cell lines. One of our best analogs was a heptapeptide, TT-232, which strongly inhibited the tyrosine kinase activity and the cell-proliferation in different colon tumor cells. However, it did not inhibit the growth hormone release either in vitro or in vivo from rat pituitary cells. The TT-232 was found to be effective on 60 human tumor cell lines, it significantly inhibited the tumor growth on different animal tumor models, and induced apoptosis, as a result of which some animals became tumor free. The TT-232 inhibited the tumor growth of PC3 prostate xenografts with 60% and caused a 100% survival of mice 60 days after the transplantation. It is being preclinically tested at present. We have shown that the new GnRH analogs acting without any hormonal effect and the Somatostatin analogs with strong antitumor and tyrosine kinase inhibitory activity but no hormonal effect may represent a breakthrough in the research of the antitumor peptides, having direct effect on tumor cells. PMID- 10866358 TI - Photoperiodic induction of ovarian growth and plasma estradiol secretion in a migratory finch, Emberiza melanocephala: involvement of circadian rhythm. AB - To demonstrate the involvement of circadian rhythm in photoperiodic time measurement, photosensitive female blackheaded buntings were kept under different photoperiodic schedules consisting of 6 h of main photophase coupled with scotophases of various durations. Ovarian mass and circulating plasma estradiol concentration were found to be elevated in cycles of 6L:6D, 6L:36D, 6L:54D and in control 15L:9D groups. But cycles of 6L:18D, 6L:42D and 6L:66D did not stimulate ovarian growth or elevate circulating plasma estradiol concentration. These results are consistent with the Bunning hypothesis according to which a photoperiodic response is elicited as a result of the coincidence of light with the photoinducible phase of an endogenous circadian rhythm. The results thus indicate the involvement of a circadian rhythm of photoinducibility in ovarian growth and estradiol secretion. PMID- 10866359 TI - Seasonal adiposity and androstenedione production as a possible mechanism for asynchronous reproductive activity between males and females of vespertilionid bat, Scotophilus heathi. AB - Relationship between changes in the body weight and circulating androstenedione concentration was examined in order to find out their involvement in asynchronous gametic cycle of male and female S. heathi. Body weight of both male and female bats increased from September due to accumulation of white adipose tissue and reached to a peak in November. The body weight declined gradually due to consumption of adipose tissue during December and January reaching basal level in February. A significant correlation between body weight and serum androstenedione concentration was observed. Correlated with maximum body weight, peak androstenedione level was found during November in both male and female bats. Peak androstenedione level during November may be responsible for stimulating spermatogenesis in the male, but may suppress follicular growth and ovulation in the female, thus resulting in asynchronous gametic cycle of male and female S. heathi. PMID- 10866360 TI - Methallibure inhibition of testicular and seminal vesicle activity in catfish, Clarias batrachus (Linn.): a study correlating changes in serum sex steroid profiles. AB - The administration of methallibure (2 microg/g BW, daily for 15 days) in Clarias batrachus in prespawning phase (May-June) resulted in decreased weights of seminal vesicle (SV) and testis, and reductions in the concentrations of total proteins, fructose, hexosamines, and sialic acid in SV and testis. The inhibitory changes can be attributed to impairment of steroidogenesis, serum levels of testosterone and estradiol -17beta decreased significantly. Withdrawal of methallibure treatment for 7 and 15 days resulted in gradual recovery and restoration of all the above parameters except the sialic acid levels in the SV and testis, and fructose level in the SV. The methallibure induced regressive changes in the SV and testis were discussed in the light of its GTH inhibiting property. PMID- 10866361 TI - Annual cyclic variations in some biochemical constituents of seminal vesicle and testis of the catfish Heteropneustes fossilis (Bloch): a study correlating plasma testosterone level. AB - In Heteropneustes fossilis, significant annual variations were observed in seminal vesicle-somatic index (SVSI), gonadosomatic index (GSI), concentrations of total proteins, hexosamines, fructose and glucose in both SV and testis, and in plasma testosterone with high values in late prespawning-early spawning phases (June-July) and low or undetectable levels in resting phase (December-January) except for glucose. There is an inverse relationship between the annual patterns of fructose and glucose with fructose dominant in the prespawning and early spawning phases (June-July), and glucose in the resting phase (November-January). The increase in the concentrations of SV and testicular protein, hexosamine and fructose can be correlated with the increase in testosterone concentration on one hand and with the increase of SVSI and GSI, on the other. The decrease in glucose level in the recrudescent phase may be due to its increased conversion into fructose, the main seminal sugar in this species. PMID- 10866362 TI - Calmodulin gene expression in an immortalized striatal gabaergic cell line. AB - We have demonstrated the presence of the mRNAs transcribed from the three calmodulin (CaM) genes in the GABAergic cell line M26-1F derived from embryonic rat striatal cells and immortalized by oncogene transduction. Similarly as in the rat striatum in vivo, these clonal cells express CaM I, CaM II and CaM III mRNAs differently, the CaM I mRNA population being the most abundant, followed in sequence by the CaM II and CaM III mRNA populations. The proportions of these transcripts resemble those in the adult striatum. The possibility of deriving immortalized cell lines from primary neuronal tissue which exhibit characteristics similar to those of the tissue of origin could provide an important tool in many types of in vitro studies. PMID- 10866363 TI - Synthesis and accumulation of poly(3-hydroxybutyric acid) by Rhizobium sp. AB - Forty-two Rhizobium strains obtained from different culture collections were evaluated quantitatively for poly(3-hydroxy-butyric acid) [PHB] production in shake flask culture. The majority of the strains produced the maximum amount of PHB during the late exponential or stationary phase of growth. Synthesis and accumulation of PHB in different species of Rhizobium were found to vary between 1-38% of their dry biomass. Growth and PHB production by the Rhizobium strain TAL 640 were greatly influenced by the C-source and D-mannitol was fundamental to both processes. The identity and purity of PHB isolated from TAL-640 have also been confirmed by UV-, IR- and 1H-NMR spectroscopic analyses. PMID- 10866364 TI - Effects of delta-aminolevulinic acid on pigment formation and chlorophyllase activity in French bean leaf. AB - Chloroplast development and chlorophyll biosynthesis are co-regulated. Treatment by levulinic acid resulted in a linear relation in both chlorophyll and carotenoid contents, during greening of etiolated French bean leaf discs. Chlorophyll biosynthesis appeared to control that of caroteins. In the presence of levulinic acid; at different levels, photosystem II (PS II) activity decreased when expressed on a chlorophyll basis. Chlorophyllase activity was increased progressively by increasing levulinic acid concentration. Thus, levulinic acid could be used to arrest the light-induced chloroplast development at a desired phase of greening and acts as determinator of chloroplast development in green tissues. PMID- 10866365 TI - Morphometry of the FRTL-5 cells after irradiation. AB - The morphological changes of in vitro irradiated FRTL-5 cells and their ability to grow in semi-solid medium were studied morphometrically. FRTL-5 cells were grown in medium with 4 different concentrations of TSH (0, 0.1, 1, 10 mU/ml). After irradiation with 0 Gy, 2 Gy and 4 Gy, the cells were seeded on glass cover slips and in methocel. Fourteen days after irradiation, the morphometric analysis of FRTL-5 cells and their nuclei was performed. The results showed that irradiation and different doses of TSH have influence on FRTL-5 cell size, more on their nuclei than on the cells as a whole. Growing of FRTL-5 cells in the methocel indicates the possible transformation of these cells after long culturing in the TSH medium and after irradiation. PMID- 10866366 TI - Automated fluorescent detection of a 10 loci multiplex for paternity testing. AB - The discovery of new highly efficient tetra repeat STR loci, development of fluorescence multicolour dye technology and capillary electrophoresis have made it possible to amplify ten loci in a single reaction. This combination provides an extraordinary effectiveness of simultaneous amplification and detection. With this method it became possible to determine individual identity and paternity at an enhanced level of precision and accuracy in 1 to 2 days with a high biostatistical probability. This review demonstrates the role of automated fluorescent multicolour dye genotyping technology in forensic paternity testing. PMID- 10866367 TI - Synthesis and in vitro antibacterial activity of 3-[N-methyl-N-(3-methyl-1,3 thiazolium-2-yl)aminolmethyl cephalosporin derivatives. AB - The new cephalosporins having N-linked quarternary ammonium salt at C-3 position were prepared from the reaction of 2-methylimino-3-methyl-1,3-thiazoline derivatives with cephalosporins. Also antimicrobial activities of those compounds were determined. PMID- 10866368 TI - Synthesis and evaluation of 9,9-dimethylxanthene tricyclics against trypanothione reductase, Trypanosoma brucei, Trypanosoma cruzi and Leishmania donovani. AB - Derivatives of 9,9-dimethylxanthene were synthesised and evaluated against trypanothione reductase (TR) and in vitro against parasitic trypanosomes and leishmania. High in vitro antiparasitic activity was observed for some derivatives with one compound showing high activity against all three parasites (ED50 values of 0.02, 0.48 and 0.32 microM, for Trypanosoma brucei, Trypanosoma cruzi, and Leishmania donovani, respectively). The lack of correlation between inhibitory activity against TR and ED50 values suggests that TR is not the target. PMID- 10866369 TI - Hepatitis C viral IRES inhibition by phenazine and phenazine-like molecules. AB - An in vitro assay based on the expression of Fluci reporter gene under the translational control of HCV IRES was used to evaluate and screen compound libraries. A structure-activity relationship study on a phenazine hit was conducted. Our data suggest that an intact phenazine or phenazine-like core with two distal polar substitutions is crucial for potency. PMID- 10866370 TI - Prediction of drug solubility from Monte Carlo simulations. AB - Monte Carlo statistical mechanics simulations have been carried out for 150 organic solutes in water. Physically significant descriptors such as the solvent accessible surface area, numbers of hydrogen bonds, and indices for cohesive interactions in solids are correlated with pharmacologically important properties including octanol/water partition coefficient (log P) and aqueous solubility (log S). The regression equation for log S only requires five descriptors to provide a correlation coefficient, r2, of 0.9 and rms error of 0.7 for the 150 solutes. The descriptors can form a basis for structural modifications to guide an analogue's properties into desired ranges. PMID- 10866371 TI - Novel inhibitors of HIV protease: design, synthesis and biological evaluation of picomolar inhibitors containing cyclic P1/P2 scaffolds. AB - A novel series of HIV protease inhibitors containing cyclic P1/P2 scaffolds has been synthesized and evaluated for biological activity. The trans 3,5-dibenzyl-2 oxo pyrrolidinone ring system resulted in a 50 pM enzyme inhibitor against HIV protease in vitro when combined with an indanolamine derived P'-backbone. This compound also shows comparable activity to currently marketed drugs in the MT-4 cell-based antiviral assay. PMID- 10866372 TI - Carbacyclic peptide mimetics as VCAM-VLA-4 antagonists. AB - Substitution of carbon for sulfur in a potent 13-membered cyclic disulfide containing peptide was accomplished via an intramolecular Wittig reaction and resulted in a series of 'carba' analogues. Potency in the VCAM-VLA-4 assay was sensitive to ring size and lower than that of the parent disulfide. PMID- 10866373 TI - Cyclic thioether peptide mimetics as VCAM-VLA-4 antagonists. AB - Selective substitution of a sulfur atom by carbon in a highly potent 13-membered cyclic disulfide was accomplished by intramolecular displacement of a bromide. The potency of the resulting thioethers in the VCAM/VLA-4 assay was dependent on ring size and the position of the sulfur atom. PMID- 10866374 TI - The design and synthesis of potent cyclic peptide VCAM-VLA-4 antagonists incorporating an achiral Asp-Pro mimetic. AB - The Asp-Pro sequence of the cyclic peptide Ac-HN-Tyr-Cys*-Asp-Pro-Cys*-OH (1) could be replaced with the achiral dipeptide mimetic 1-(2 aminoethyl)cyclpentylcarboxylic acid with retention of potent inhibition of the VCAM-VLA-4 interaction. PMID- 10866375 TI - Design, synthesis and SAR of a series of 2-substituted 4-amino-quinazoline neuropeptide Y Y5 receptor antagonists. AB - The design of a novel series of NPY-Y5 receptor antagonists is described. Key elements for the design were the identification of weak Y5 hits from a Y1 program, results from a combinatorial approach and database mining. This led to the discovery of the quinazoline 4 and the aryl-sulphonamide moiety as major components of the pharmacophore for Y5 affinity. The synthesis and SAR towards CGP71683A is described. PMID- 10866376 TI - Immunomodulatory activity of hexapeptides related to proline rich peptide from colostrum. AB - Twelve analogues of an immunomodulatory hexapeptide YVPGFP (I) derived from Proline rich peptide (from colostrum) have been synthesized with modifications at positions 2, 4 and 6. In MLR assay one of the analogues exhibited approx 50% inhibition at 0.1 microg/mL concentration in contrast to prednisolone and I which caused around 70 and 20% suppression respectively, at the same concentration. PMID- 10866377 TI - Beta-casomorphins: substitution of phenylalanine with beta-homo phenylalanine increases the mu-type opioid receptor affinity. AB - Two analogues of bovine beta-casomorphin-7 and beta-casomorphin-5 containing a beta-homo phenylalanine in substitution of the phenylalanine in position 3 were synthesised and tested for their mu-opioid receptor affinity. The modification enhanced the mu receptor affinity 5-fold in the case of modified beta-CM-7 and 2 fold for modified beta-CM-5 when compared to the natural peptides. PMID- 10866378 TI - The basal SAR of a novel insulin receptor activator. AB - The synthesis and SAR of analogues prepared from novel insulin receptor activator 1 are described. Changes to the dihydroxyquinone core were not tolerated while functionalization of the two indoles contained in 1 resulted in little effect upon activation of the insulin receptor. PMID- 10866379 TI - COBRA-1, a rationally-designed epoxy-THF containing compound with potent tubulin depolymerizing activity as a novel anticancer agent. AB - A novel mono-THF containing synthetic anticancer drug, COBRA-1, was designed for targeting a previously unrecognized unique narrow binding cavity on the surface of alpha-tubulin. COBRA-1 inhibited GTP-induced tubulin polymerization in cell free tubulin turbidity assays. Treatment of human breast cancer and brain tumor (glioblastoma) cells with COBRA-1 caused destruction of microtubule organization and apoptosis. Like other microtubule-interfering agents, COBRA-1 activated the proapoptotic c-Jun N-terminal kinase (JNK) signal transduction pathway, as evidenced by rapid induction of c-jun expression. PMID- 10866380 TI - Diamino benzo[b]thiophene derivatives as a novel class of active site directed thrombin inhibitors. Part 6: further focus on the contracted C4'-side chain analogues. AB - Novel benzo[b]thiophene diamine thrombin inhibitors were investigated, focusing on a contracted C4'-side chain series. SAR studies identified compounds with either a pyrrolidino or morpholino group as potent, active site directed thrombin inhibitors when the amino group was connected to the C3-phenyl ring with a methylene linker at the C4' position of the phenyl ring. PMID- 10866381 TI - Methaqualone derivatives are potent noncompetitive AMPA receptor antagonists. AB - Quinazolin-4-one derivatives of methaqualone substituted at C-2 define a new class of noncompetitive antagonists at AMPA receptors. PMID- 10866382 TI - 9-Benzylpurines with inhibitory activity against Mycobacterium tuberculosis. AB - 9-Benzylpurines with a variety of substituents in the 2-, 6- and/or 8-position have been prepared and screened for antimycobacterial effects. High inhibitory activity against Mycobacterium tuberculosis was found for 9-benzylpurines carrying a phenylethynyl-, trans-styryl or aryl substituents in the 6-position and generally chlorine in the 2-position tends to increase activity. PMID- 10866383 TI - Synthesis and structure-activity relationships of quaternary ammonium cephalosporins with 3-pyrazolylpyridinium derivatives. AB - Cephalosporins with 3-pyazolylpyridinium at C-3 position, which is supposed to exhibit synergic activity of ceftazidime and cefoselis, were synthesized and their antibacterial activity against gram-positive and gram-negative was inspected. PMID- 10866384 TI - Bicyclic nucleoside inhibitors of Varicella-Zoster Virus (VZV): the effect of a terminal halogen substitution in the side-chain. AB - Preliminary SAR studies on the side chain of a new class of antiviral nucleosides have shown that terminal substitution in the side-chain, with a halogen atom, lead to potent and highly specific anti-VZV agents. PMID- 10866385 TI - Mass spectrometry reveals elastase inhibitors from the reactive centre loop of alpha1-antitrypsin. AB - Peptides derived from the reactive centre loop of alpha1-antitrypsin, a serpin, were screened as potential elastase inhibitors by mass spectrometry. An octapeptide, MFLEAIPM, formed a 'stable' ternary complex with porcine elastase: one MFLEAIPM molecule reacted covalently with loss of water, whilst an additional peptide was bound non-covalently. Kinetic analyses suggested that MFLEAIPM may act as an uncompetitive inhibitor and that the activity was associated with the four N-terminal residues. PMID- 10866386 TI - A focused compound library of novel N-(7-indolyl)benzenesulfonamides for the discovery of potent cell cycle inhibitors. AB - A series of compounds containing an N-(7-indolyl)benzenesulfonamide pharmacophore was synthesized and evaluated as a potential antitumor agent. Cell cycle analysis with P388 murine leukemia cells revealed that there were two different classes of potent cell cycle inhibitors; one disrupted mitosis and the other caused G1 accumulation. Herein described is the SAR summary of the substituent patterns on this pharmacophore template. PMID- 10866387 TI - 'Double-Drugs'--a new class of prodrug form of an HIV protease inhibitor conjugated with a reverse transcriptase inhibitor by a spontaneously cleavable linker. AB - We designed and synthesized a new series of prodrug-type anti-HIV agents consisting of a peptidomimetic HIV protease inhibitor conjugated with a nucleoside reverse transcriptase inhibitor in an effort to enhance the antiviral activity. For the conjugation, a series of linkers that conjoin the two different classes of inhibitors have been investigated. Conjugates using a succinyl amino acid linker were shown to release the parent components via the spontaneous imide formation at a faster rate compared to conjugates using a glutaryl amino acid linker, as expected from the energetically favorable cyclization to the five membered ring. Herein, we report a new 'double-drug' 4b (KNI-1039) with a glutarylglycine linker, which exhibited extremely potent anti-HIV activity compared with that of the individual components. PMID- 10866388 TI - Synthesis of glycosylated polyethylenimine with reduced toxicity and high transfecting efficiency. AB - A safe and efficient synthesis of glycosylated polyethylenimine using titanium (IV) isopropoxide and sodium borohydride has been carried out as a substitute for the highly toxic sodium cyanoborohydride method currently used. Poryplexes formed between DNA and the various glycosylated polyethylenimines appeared to be much less cytotoxic than polyethylenimine (PEI)/DNA polyplexes. PMID- 10866389 TI - Squalamine analogues as potential anti-trypanosomal and anti-leishmanial compounds. AB - This paper concerns the synthesis of various simplified analogues of the novel anti-microbial agent, squalamine. The compounds were then investigated for activity against Trypanosoma brucei, the causative agent of African trypanosomiasis, Trypanosoma cruzi, the causative agent of Chagas disease and Leishmania donovani, the causative agent of visceral leishmaniasis. Several compounds showed in vitro activity, especially against T. brucei and L. donovani. However, one compound showed poor in vivo activity. PMID- 10866390 TI - (+)-4-phosphonophenylglycine (PPG) a new group III selective metabotropic glutamate receptor agonist. AB - A new synthesis of (R,S)-PPG (4-phosphonophenylglycine) and the separation of the protected enantiomers leading after deprotection to (+)- and (-)-PPG are described. Pharmacological characterization at the group III metabotropic glutamate receptors hmGluR4a and hmGluR7b revealed (+)-PPG as the active enantiomer. PMID- 10866392 TI - Nucleotide libraries as a source of biologically relevant chemical diversity: solution-phase synthesis. AB - Solution-phase parallel synthesis of nucleotide library 1 consisting of 150 members is reported. PMID- 10866391 TI - The design and synthesis of the high efficacy, non-peptide CCK1 receptor agonist PD170292. AB - The design, synthesis and biological actions of a novel, non-peptide CCK1 receptor agonist (PD 170292) which exhibits a similar pharmacological profile to the CCK analogue JMV180 is reported. PD 170292 was designed based on a consideration of the structures of a peptide based CCK1 receptor selective agonist and a peptoid CCK2 receptor selective antagonist. PMID- 10866393 TI - Synthesis and activity studies of conformationally restricted alpha-ketoamide factor Xa inhibitors. AB - Conformationally restricted borolysine compounds containing a 2-(2 cyanophenylthio) benzoyl in the P3 position unexpectedly led to enhanced factor Xa inhibition. In an effort to improve both the potency and selectivity of this series by extending into the S' domain, we have replaced the boronic acid with alpha-ketoamides, utilizing a novel process that was developed in our labs. PMID- 10866394 TI - Carbocyclic influenza neuraminidase inhibitors possessing a C3-cyclic amine side chain: synthesis and inhibitory activity. AB - As part of our continuing work in the area of influenza neuraminidase inhibitors, a series of C3-aza inhibitors possessing a cyclic amine side chain was synthesized and evaluated for influenza neuraminidase inhibitory activity. Analogues possessing a six-, seven- and eight-membered ring, 4c-e, respectively, at the C3 position exhibited excellent influenza B neuraminidase inhibition. PMID- 10866395 TI - SAR of 4-hydroxypiperidine and hydroxyalkyl substituted heterocycles as novel p38 map kinase inhibitors. AB - The 4-hydroxypiperidine substituent was found to confer high p38 selectivity devoid of COX-1 affinity, when attached to a series of pyridinyl substituted heterocycles. Pyridinyloxazole 11 showed a promising in vivo profile with bioavailability of 64% and ED50 in rat collagen induced arthritis of 10 mg/kg po bid. In contrast to pyridinylimidazoles such as SB 203580, 11 did not inhibit human cytochrome P450 isoenzymes. PMID- 10866396 TI - The synthesis and in vitro cytotoxic studies of novel oxa-spermidine derivatives and homologues. AB - A series of oxa-spermidine derivatives and homologues were prepared and their anticancer properties were evaluated. All these compounds showed an average GI50 value in the range of 3.9-28.9 microM. SAR studies showed that the presence of a sulphonamido functionality and the length of the alkyl chain are important factors for an enhanced activity. PMID- 10866397 TI - Cytotoxic halogenoacrylic derivatives of distamycin A. AB - The design, synthesis, in vitro and in vivo activities of a series of halogenoacrylic derivatives of distamycin A are described. The structure-activity relationships indicate a key role of the reactivity of alpha-halogenoacrylic moiety. The reactivity and the putative alkylating mechanism of these compounds are different from those of the nitrogen mustards and possibly based on a Michael type reaction. This supports the hypothesis that these compounds represent a class of minor groove binders mechanistically different from tallimustine. PMID- 10866398 TI - Cytotoxic alpha-bromoacrylic derivatives of distamycin analogues modified at the amidino moiety. AB - The design, synthesis, in vitro and in vivo activities of novel alpha bromoacrylic derivatives of distamycin A, modified at the amidino moiety by the replacement with basic or non-basic groups are reported. In spite of the relevance of these modifications of distamycin frame, the new derivatives are potent cytotoxics. The presence of the amidino moiety, is, therefore; not an absolute requirement for the activity. In particular due to a favorable myelotoxicity/cytotoxicity ratio, guanidino derivative PNU 166196 was selected for clinical development. PMID- 10866399 TI - Conformationally-restricted ligands for the histamine H1 receptor. AB - Potent H1-antagonistic activity in a series of novel indeno[2,1-c]pyridines and their 4-arylpiperidinol precursors is reported; one compound shows an in vitro activity four times that of the standard mepyramine that it was screened against. Their failure to translate this protection to in vivo tests is discussed. PMID- 10866400 TI - 4-[(8-Alkyl-8-azabicyclo[3.2.1]octyl-3-yl)-3-arylanilino]-N,N-diethylbenzamides: high affinity, selective ligands for the delta opioid receptor illustrate factors important to antagonist activity. AB - The tropane derived compounds, 4-[(8-alkyl-8-azabicyclo[3.2.1]octyl-3-yl)-3 arylanilino]-N,N-d iethylbenzamides (5a-d), were synthesized and found to have high affinity and selectivity for the delta receptor. Compounds 5a-d are structurally similar to the full agonist (-)-RTI-5989-54 (3); yet, efficacy studies for compounds in this series (5a-d) reveal greatly diminished agonist activity as well as antagonism not found in piperidine-based compounds like 3. PMID- 10866401 TI - Substrate and stereochemical specificity of the organophosphorus acid anhydrolase from Alteromonas sp. JD6.5 toward p-nitrophenyl phosphotriesters. AB - The enzyme OPAA hydrolyzes p-nitrophenyl phosphotriesters bearing substituents at the phosphorus center ranging in size from methyl to phenyl. The enzyme exhibits stereoselectivity toward the hydrolysis of chiral substrates with a preference for the Sp enantiomer. PMID- 10866402 TI - Facile synthesis of N-Fmoc-serine-S-GlcNAc: a potential molecular probe for the functional study of O-GlcNAc. AB - A metabolically stable beta-N-acetylglucosaminyl-1-thio-N-Fmoc-serine (S-GlcNAc Ser) derivative was synthesized in two procedures: one involving a coupling of a readily obtainable 1-pseudo-thiourea of GlcNAc (S-GlcNAc) and iodo-N-Boc-L alanine benzyl ester, and the other utilizing a modified Mitsunobu reaction of GlcNAc-SH and a serine derivative. PMID- 10866403 TI - Total synthesis of pyrrolizidines 223H', 239K', 265H', and 267H' found in Madagascan frogs (Mantella) and their affinities for nicotinic acetylcholine receptor. AB - The total synthesis of pyrrolizidines 223H', 239K', 265H', and 267H' has been achieved starting from 1,5-hexadiene via a common synthetic intermediate 5. The affinity of 1-4 for nicotinic acetylcholine receptor was evaluated. PMID- 10866404 TI - Discovery of a potent, non-triketone type inhibitor of 4-hydroxyphenylpyruvate dioxygenase. AB - 3-Cyclopropanecarbonyloxy-2-cyclohexen-1-one has been found to be a new, potent, low molecular weight non-triketone type inhibitor of 4-hydroxyphenylpyruvate dioxygenase with IC50 value of 30 nM. Preliminary studies suggest that the two carbonyl groups present in the compound are crucial for the inhibition activity. PMID- 10866405 TI - Functionalized nucleoside 5'-triphosphates for in vitro selection of new catalytic ribonucleic acids. AB - A series of novel 2'-modified nucleoside 5'-triphosphates was synthesized. The amino, imidazole, and carboxylate functionalities were attached to the 5-position of pyrimidine base of these molecules through alkynyl and alkyl spacers, respectively. Two different phosphorylation methods were used to optimize the yields of these highly modified triphosphates. PMID- 10866406 TI - Synthesis and evaluation of geldanamycin-testosterone hybrids. AB - Geldanamycin (GDM) binds to the Hsp90 chaperone protein resulting in the degradation of several important signaling proteins. A series of GDM-testosterone linked hybrids has been synthesized and evaluated for activity against prostate cancer cell lines. The hybrid with the greatest activity exhibits potent and selective cytotoxicity against prostate cancer cells containing the androgen receptor. PMID- 10866407 TI - Teleconferencing with dynamic medical images. AB - Dynamic images, a sequence of static images displayed in rapid succession and perceived as a continuous motion by the human eye, are widely used in medicine. One of the primary objectives of telemedicine is the transmission of such images to a distant location to manage clinical problems remotely. A broad variety of methods is available to acquire, store, transmit, and display these images. However, the context of the clinical problem determines which of these methods can be deployed in a telemedicine solution. This paper discusses the advantages and disadvantages of the different technologies and presents an example of a teleconferencing system for interventional cardiology. This system acquires cardiac angiography and intravascular ultrasound images and transmits them over an existing Internet connection to a distant location. It is specifically optimized for clinical conferencing, where time is limited for each case presentation during the conference, compared to the relatively long time available for the conference preparation. The system takes advantage of this characteristic by transmitting the images well in advance of the clinical conference and displaying them synchronously in both locations during the conference. This allows for the preservation of the original image quality. PMID- 10866408 TI - A medical digital library to support scenario and user-tailored information retrieval. AB - Current large-scale information sources are designed to support general queries and lack the ability to support scenario-specific information navigation, gathering, and presentation. As a result, users are often unable to obtain desired specific information within a well-defined subject area. Today's information systems do not provide efficient content navigation, incremental appropriate matching, or content correlation. We are developing the following innovative technologies to remedy these problems: 1) scenario-based proxies, enabling the gathering and filtering of information customized for users within a pre-defined domain; 2) context-sensitive navigation and matching, providing approximate matching and similarity links when an exact match to a user's request is unavailable; 3) content correlation of documents, creating semantic links between documents and information sources; and 4) user models for customizing retrieved information and result presentation. A digital medical library is currently being constructed using these technologies to provide customized information for the user. The technologies are general in nature and can provide custom and scenario-specific information in many other domains (e.g., crisis management). PMID- 10866409 TI - Web-based video for real-time monitoring of radiological procedures. AB - A web-based video transmission of images from CT and MRI consoles was implemented in an Intranet environment for real-time monitoring of ongoing procedures. Images captured from the consoles are compressed to video resolution and broadcast through a web server. When called upon, the attending radiologists can view these live images on any computer within the secured Intranet network. With adequate compression, these images can be displayed simultaneously in different locations at a rate of 2 to 5 images/s through a standard local-area network. While the quality of the images was insufficient for diagnostic purposes, our users survey showed that they were suitable for supervising a procedure, positioning the imaging slices, and for routine quality checking before completion of a study. The system was implemented at UCLA to monitor nine CT's and six MRI's distributed in four different buildings. This system significantly improved the radiologists productivity by saving valuable time spent in trips between reading rooms and examination rooms. It also improved patient care and throughput by reducing the time spent waiting for the radiologists to check a study before removing the patient from the scanner. PMID- 10866410 TI - InternetQuestion and Answer (iQ&A): a Web-based survey technology. AB - This paper presents InternetQuestion and Answer, a Web-based survey development and implementation technology, which has been designed for constructing on-line surveys for educational, medical, or administrative purposes. The system, called iQ&A, is a three-tiered database-backed Web system that has been developed to support a wide range of applications. Surveys are considered as general data collection instruments and include a wide field of application. iQ&A facilitates rapid survey construction and administration which is ideally suited for biomedical research as well as other research and educational activities. Full report management capabilities provide the survey publisher on-line access to current information on survey responses. Current implementations of this technology in the areas of biomedical applications of clinical trials, longitudinal research, and other research-related systems are presented. Further refinement of the current system should lead to a powerful general survey technology for broad-based applications. PMID- 10866411 TI - Genetic algorithms for a robust 3-D MR-CT registration. AB - The aim of this paper is to present an original usage of genetic algorithms as a robust search space sampler in application to 3-D medical image elastic registration. An overview of the standard steps of a registration algorithm is given. We focus on the genetic algorithms use and particularly on the problem of extraction of the optimal solution among the final genetic population. We provide an original encoding scheme relying on a structural approach of point matching and then point out the need for a local optimization process. We then illustrate the algorithm with a concrete registration example and assert the results with a direct multivolume rendering tool. Finally, the algorithm is applied on the vanderbilt medical image database to assert the robustness and in order to compare it with other techniques. PMID- 10866412 TI - Construction of a human topological model from medical data. AB - Medical imaging can provide data for useful views of the interior details of human anatomy. In addition to visualization, which in general has been the primary reason for obtaining these data, many other uses are possible. These include modeling of different elements and their inter-relationships-topological modeling, simulation of physical processes, analysis of movements, and validation of models. Here, we describe some of the modeling issues from medical imaging. The issues are particularly related to topological modeling of different anatomical elements: bones, muscles, articulations, etc. A three-dimensional topological modeler is presented with which anatomists and other users can build a topological database containing structural, topological, and mechanical information of anatomical elements. PMID- 10866413 TI - Three-dimensional modeling and visualization of the cochlea on the Internet. AB - Three-dimensional (3-D) modeling and visualization of the cochlea using the World Wide Web (WWW) is an effective way of sharing anatomic information for cochlear implantation over the Internet, particularly for morphometry-based research and resident training in otolaryngology and neuroradiology. In this paper, 3-D modeling, visualization, and animation techniques are integrated in an interactive and platform-independent manner and implemented over the WWW. Cohen's template shape with mean cross-sectional areas of the human cochlea is extended into a 3-D geometrical model. Also, spiral computer tomography data of a patient's cochlea is digitally segmented and geometrically represented. The cochlear electrode array is synthesized according to its specification. Then, cochlear implantation is animated with both idealized and real cochlear models. Insertion length, angular position, and characteristic frequency of individual electrodes are estimated online during the virtual insertion. The optimization of the processing parameters is done to demonstrate the feasibility of this technology for clinical applications. PMID- 10866414 TI - Content-based retrieval of dynamic PET functional images. AB - The recent information explosion has led to massively increased demand for multimedia data storage in integrated database systems. Content-based retrieval is an important alternative and complement to traditional keyword-based searching for multimedia data and can greatly enhance information management. However, current content-based image retrieval techniques have some deficiencies when applied in the biomedical functional imaging domain. In this paper, we presented a prototype design for a content-based functional image retrieval database system for dynamic positron emission tomography. The system supports efficient content based retrieval based on physiological kinetic features and reduces image storage requirements. This design makes it possible to maintain a large number of patient data sets online and to rapidly retrieve dynamic functional image sequences for interpretation and generation of physiological parametric images, and offers potential advantages in medical image data management and telemedicine, as well as providing possible opportunities in the statistical and comparative analysis of functional image data. PMID- 10866415 TI - The design and applications of a recursive molecular modeling framework. AB - In this paper, we present a recursive molecular model suitable for multiresolution analysis in a variety of application areas. Our approach allows ease of use by computer scientists and biologists by proposing a software interface for molecular analysis that hides programming details. We demonstrate that our design is flexible enough for use in desktop analytical applications, in large-scale parallel simulations, and as a server for remote molecular analysis across wide-area networks. PMID- 10866416 TI - Telemedicine for evaluation of brain function by a metacomputer. AB - A method of evaluating brain function using the metacomputer concept of the Globus system combined with a message-passing interface is described. The proposed method has the ability to exploit various geographically distributed resources and parallel computing linked to a high-technology medical instrumentation system, magnetoencephalography, to analyze the functional state of the brain. It is envisaged that the method will lead to the realization of an efficient telemedicine system for health care. PMID- 10866417 TI - NEWBET: telemedicine platform for burn patients. AB - Problems detected in the current healthcare services for burn patients and emergency situations are taken into consideration in the development of a telemedicine platform, which is proposed in this paper. Taking as a reference the user requirements and the client-server architecture, a model of communications that allows a bidirectional exchange of information is reported. Special attention is focused towards the methodology applied and the design of a friendly user interface compatible with the current prevailing clinical protocols. The platform is based on a Microsoft Windows environment, and communications are implemented by using sockets. PMID- 10866418 TI - Real-time teleconsultation with high-resolution and large-volume medical images for collaborative healthcare. AB - Real-time consultation between referring physicians or radiologists with an expert is critical for timely and adequate management of problem cases. During consultation, both sides need to: 1) synchronously manipulate high-resolution digital radiographic images or large volume MR/CT images; 2) perform interpretation interactively; and 3) converse with audio. We present a specifically designed teleconsultation system in a digital imaging and communication in medicine picture archiving and communication systems clinical environment. The system uses bidirectional remote control technology to meet critical teleconsultation application requirements with high-resolution and large volume medical images operated in a limited-bandwidth network setting. We give the system design and implementation methods, and also describe the teleconsultation procedure and protocol used in this system. Finally, laboratory and clinical evaluation results are discussed. PMID- 10866419 TI - Assessing quality in cardiac surgery: why this is necessary in the twenty-first century. AB - The cost and high-profile nature of coronary surgery means that this is an area of close public scrutiny. As much pioneering work in data collection and risk analyses has been carried out by cardiac surgeons, substantial information exists and the correct interpretation of that data is identified as an important issue. This paper considers the background and history of risk-adjustment in cardiac surgery, the uses of quality data, examines the observed/expected mortality ratio and looks at issues such as cost and reactions to outliers. The conclusion of the study is that the continuation of accurate data collection by the whole operative team and a strong commitment to constantly improving quality is crucial to its meaningful application. PMID- 10866420 TI - Management of anticoagulation and its reversal during paediatric cardiopulmonary bypass: a review of current UK practice. AB - Protocols for management of heparin and protamine administration in patients undergoing open-heart surgery have been developed from experience gained mainly in adult practice. However, it has been demonstrated that there are marked differences between paediatric and adult patients in their response to systemic anticoagulation and its reversal. The aim of this study was to obtain an overview of current practice of management of anticoagulation and its reversal from paediatric cardiac surgical units of Great Britain and Ireland. All centres performing paediatric cardiac surgery agreed to participate in the survey (n = 16). Telephone interviews were carried out with the chief or a senior perfusionist from all participating institutions, which were based on a structured questionnaire compiled specifically for the purpose. The answers were anonymised. At present, in the UK and Ireland, unfractionated heparin is the anticoagulant of choice in all units, with a slight prevalence of porcine mucosal (9/16, 56.5%) versus bovine lung preparation (7/16, 44.0%). The policy for administration of heparin to the patient is uniform, with a dose of 300 IU/kg. However, there is great variability in the amount of heparin added to the prime and to the volume infused during cardiopulmonary bypass (CPB). Monitoring of anticoagulation is achieved by activated coagulation time alone in all but one centre, with lower limits varying between 400 and 750 s. Use of aprotinin is widely accepted, but clinical indications are highly variable. No centre adopts heparin-bonded or heparin-coated circuitry for CPB. Calculation of initial and additional protamine doses followed a variety of criteria, resulting in a very wide distribution of doses. The data obtained highlighted the lack of uniformity among paediatric cardiac surgical units of Great Britain and Ireland with regard to most of the issues related to the management of anticoagulation and its reversal. The striking heterogeneity of our cross-sectional observations clearly underlines the need for prospective, multicentre studies on a national basis to relate different clinical practices to outcome measures. PMID- 10866421 TI - Whole blood heparin concentrations do not correlate with plasma antifactor Xa heparin concentrations in pediatric patients undergoing cardiopulmonary bypass. AB - This study was designed to test the validity of whole blood heparin concentration (WHBC) measurements using an on-site protamine titration assay with the Hepcon instrument (Medtronic Blood Management, Parker, CO, USA) in pediatric patients less than 1 year old undergoing cardiopulmonary bypass (CPB). The validity of the Hepcon measurements was examined via a test of correlation with the gold standard plasma antifactor Xa activity (anti-Xa) assay. Fifty-one patients (23 females and 28 males) under 1 year old (mean age 5.3 months) were studied prospectively. Blood samples were taken at 5 min into CPB and at the end of CPB for the WBHC, plasma anti-Xa activity, and hematocrit (Hct). The WBHC was converted to plasma heparin level using a formula taking into account the collection of blood into citrate solution and the effect of the citrate on the hematocrit. While a nonparametric statistical analysis revealed that the mean corrected values from the Hepcon instrument were not significantly different from the mean anti-Xa values (p = 0.070 at 5 min on CPB, p = 0.518 at the end of CPB), there was no significant correlation between these values at either 5 min into CPB (r = 0.113, p = 0.429) or at the end of CPB (r = -0.247, p = 0.080). The lack of correlation between the two assays may be related to the extreme hemodilution observed during CPB in small children, which leads to very low concentrations of coagulation proteins. Due to this discrepancy, whole blood heparin monitors should be further evaluated in children undergoing CPB. PMID- 10866422 TI - Aprotinin in the management of life-threatening bleeding during extracorporeal life support. AB - Contact with the synthetic surfaces of an extracorporeal circuit induces alterations in vascular components, derangements of the coagulation cascade and a systemic inflammatory response. Aprotinin reduces intraoperative and postoperative bleeding in adults undergoing cardiopulmonary bypass; however, trials in children have not had similar favorable results. While there have been some anecdotal reports, there have been no prospective clinical trials exploring the utility of aprotinin in the prevention of or as a therapy for bleeding while on extracorporeal life support (ECLS). We present a case series on our experience utilizing aprotinin for the treatment of life-threatening bleeding during ECLS. The combination of a loading dose followed by a continuous infusion resulted in significant reduction in blood loss and blood product utilization. This suggests that aprotinin may have clinical efficacy in the management of massive blood loss while on ECLS; however, larger controlled trials will be essential to determine the efficacy and appropriate dosing regimens before widespread use in ECLS can be advocated. PMID- 10866423 TI - Pulsatile normothermic cardiopulmonary bypass and plasma catecholamine levels. AB - The aim of the study was to assess plasma catecholamine levels in patients undergoing myocardial revascularization and relate them to pulsatile (P) and nonpulsatile (NP) normothermic cardiopulmonary bypass (CPB). Twenty-eight patients were randomly assigned to different CPB management: 15 patients were assigned to group 'P', 13 patients to group 'NP'. During normothermic extracorporeal circulation, group 'P' received pulsatile perfusion, while group 'NP' received nonpulsatile perfusion. Levels of epinephrine and norepinephrine were evaluated during the operation and in the intensive care unit (ICU), at seven time points. Haemodynamic assessment was performed at four time points in the same period. Demographic and surgical data were collected, and the postoperative course was analysed. Epinephrine levels were markedly increased during CPB in both groups, while norepinephrine increased more in group NP in comparison with group P. No significant difference was found in fluid administration, transfusion, drugs usage, or postoperative complications. Normothermic pulsatile CPB seems to achieve reduced levels of norepinephrine. A clinical beneficial effect of this finding was not demonstrated during the study. PMID- 10866424 TI - Haemodilution improves organ function during normothermic cardiopulmonary bypass: investigations in isolated perfused pig kidneys. AB - We investigated the effects of haemodilution on kidney function during normothermic cardiopulmonary bypass (CPB) by performing in vitro haemoperfusion of pig kidneys for 90 min after cold preservation. We compared two groups (n = 14 each) with respect to rheologic and haemodynamic parameters and glomerular and tubular function. Group 1 was perfused at a haematocrit of 0.33 +/- 0.01, group 2 at 0.21 +/- 0.01. Blood flow was adjusted according to blood pressure. Blood viscosity and vascular resistance were reduced in group 2. Comparison of group 1 versus group 2 revealed a metabolic rate of oxygen 3.4 +/- 1.7 versus 4.3 +/- 1.8 ml/min/100 g, sodium transport 1.2 +/- 1.2 versus 1.8 +/- 1.2 mmol/min/100 g, and creatinine clearance 9.9 +/- 9.1 versus 15.6 +/- 11.9 ml/min/100 g, p < 0.05. We conclude that haemodilution leads to an overproportional decrease in blood viscosity and improves the properties of flow and kidney function. In the ongoing discussion about the optimal extent of haemodilution in CPB, the importance of viscosity and blood flow should be further emphasized. PMID- 10866425 TI - Perfusion method for thoracoabdominal aneurysm repair using the open distal technique. AB - Challenges related to perfusion support of thoracoabdominal aneurysm repair include maintenance of distal aortic perfusion, rapidity of fluid resuscitation, and avoidance of both hypothermia and excessive hemodilution. Using available technology, we have devised a circuit and protocol that addresses these issues. To accomplish such support a bypass circuit consisting of 3/8 inch tubing connected to a centrifugal pump and low-prime heat exchanger was constructed. The circuit was primed via 1/4 inch spiked connectors attached to a 3-liter bag of normal saline. After initial de-airing, the solution was recirculated through this bag. Patients were anticoagulated with 1 mg/kg of heparin prior to initiation of support. Left atrial-descending aorta bypass was used primarily. A cell salvage device was used for autotransfusion. All blood products were delivered via a rapid infusion device. During partial exsanguination, shed blood was not processed, but directed to the rapid infusor for immediate retransfusion. Any packed cells given were washed prior to transfusion. Citrate dextrose solution was used as an anticoagulant for the cell scavenger. This configuration was used successfully in 50 procedures during an 18-month period. Use of this low prime, custom circuit reduced both hemodilution and cost. A connection off the cell salvage pump offers fast retransfusion of shed blood during partial exsanguination. Minimal heparinization and citrate anticoagulation appears to reduce coagulopathy. PMID- 10866426 TI - Pumpless arteriovenous extracorporeal lung assist: what is its role? AB - Extracorporeal lung assist (ECLA) is an established treatment of severe pulmonary failure. Since extracorporeal perfusion is applied in a long-term fashion in this setting, the negative impact on blood compounds is of tremendous importance. Pumpless arteriovenous ECLA (av-ECLA) is an alternatively introduced technique that focuses on reduced blood traumatization. However, due to determining technical and physiological aspects, its clinical application is limited to a highly selected group of patients. Membrane oxygenators with minimal pressure gradients, as well as stable patients' haemodynamics providing a sufficient cardiac output, are the most important prerequisites. With respect to recent reports, characteristic features of av-ECLA, with special emphasis on its physiological background, are reviewed. Accordingly, reasonable indications for its beneficial use are discussed. It is concluded that av-ECLA is a feasible technique when its limitations are accepted. For adequate clinical use, more data concerning indications, as well as time- and technique-related directions are required. PMID- 10866427 TI - 'When more is less'. Reviewing the safety and cost of cardioplegia delivery. AB - There have been several new cardioplegia delivery systems that have been developed by various manufacturers in the last few years. In this evaluation, the safety and costs associated with the present 4:1 roller pump cardioplegia delivery were compared to the Medtronic Cardioplegia Safety System (CSS). One hundred and five patients participated in a randomized, prospective evaluation. The current cardioplegia delivery system consists of using the 4:1-B MYOtherm XP disposable by Avecor Cardiovascular and a Sarns MDX 7000 roller pump. The Medtronic Cardiotherm disposable cardioplegia unit was used in conjunction with the CSS. The current delivery system will be referred to as group A and the system under evaluation will be referred to as group B. Two varying techniques are employed for cardioplegia delivery and are based on surgeon preference. An equal distribution of techniques was seen in both groups. Results indicated that in group A, a hemoconcentrator was required 60.3% of the time, whereas in group B, a hemoconcentrator was added only 25.0% of the time. The average total cardioplegia delivered in group A was 6,588 cm3 compared to 7,123 cm3 in group B. Although this is insignificant, the greatest difference was seen in the amount of crystalloid given. In group A, the crystalloid portion was 1,317 cm3 compared to only 877 cm3 in group B. There was equal weight gain and hemodilution postop in both groups. Twenty-five incidences of pressurization of the cardioplegia system occurred during the evaluation, with equal distribution in both groups. This evaluation showed that the CSS responded without fail to all pressurization incidences. The Medtronic CSS has incorporated several safety systems for pressurization and air embolism protection, which are programmable and preset by the perfusionist. The Avecor disposable has a pressurization valve that activates at a pressure greater than 600 mmHg in the cardioplegia circuit. The Sarns MDX 7,000 does not incorporate any safety shutoffs. Cost savings were achieved in two areas: hemoconcentrator use and volume of cardioplegia solution required. There was a reduction of 35% in the use of a hemoconcentrator in group B, with more total cardioplegia delivered. The cardioplegia patient cost savings in group B totaled $265.95 per case in the plain cardioplegia group and $315.95 with the amino acid cardioplegia group. The new technology incorporated into the Medtronic CSS demonstrated that it could provide more safety with less cost than the current cardioplegia pump system. PMID- 10866428 TI - Redo cardiac surgery in a Jehovah's Witness, the importance of a multidisciplinary approach to blood conservation. AB - Although Jehovah's Witnesses present a particular problem when undergoing surgery because of their refusal to accept stored blood, it is now quite common to undertake uncomplicated cardiac surgery in these patients. Complex or redo cardiac surgery however, is often associated with major blood loss, and is conventionally contraindicated in Jehovah's Witnesses. We describe the perioperative management of a Jehovah's Witness who underwent a resternotomy for mitral valve replacement and coronary artery bypass grafting having previously had an aortic valve replacement and mitral valve repair. The importance of a multidisciplinary approach to blood conservation is discussed. PMID- 10866429 TI - Anticoagulation in extracorporeal circulation using recombinant hirudin: a case report. AB - Heparin-induced thrombocytopenia (HIT) is a severe complication following the application of heparin; antibodies against complexes of heparin and PDF4 initiate activation of platelets. This may lead to massive thrombembolism, which is associated with a slight and transient drop of platelets in HIT I or a drop below 50% after approximately 5 days in HIT II. Further administration of heparin has to be strictly avoided in these patients. Immunologic evidence for HIT can easily be obtained by the heparin-induced platelet aggregation assay. If anticoagulation is necessary, different, alternative drugs are available. Recombinant hirudin (r hirudin) is a well-established drug for safe anticoagulation. Monitoring is possible by estimating the plasma level of r-hirudin from the ecarin-clotting time. We report a case of a patient with prosthetic aortic valve endocarditis and HIT II who suffered from massive postoperative bleeding requiring massive substitution of blood components and coagulants caused by free circulating r hirudin due to the use of a hemofilter. PMID- 10866430 TI - Recent advances in paediatric cardiopulmonary bypass. PMID- 10866431 TI - Iraq's children. PMID- 10866432 TI - Where the boys aren't: dioxin and the sex ratio. PMID- 10866433 TI - Youth and hormone receptors in breast cancer: good or bad news first? PMID- 10866434 TI - CFTR and disease: implications for drug development. PMID- 10866435 TI - Improving HLA matching for kidney transplantation by use of CREGs. PMID- 10866436 TI - T-cell reconstitution after stem-cell transplantation--by which organ:. PMID- 10866438 TI - A fresh look at HIV infection PMID- 10866437 TI - Voice of reason in nuclear-weapon talks. PMID- 10866439 TI - Efficacy and safety of oseltamivir in treatment of acute influenza: a randomised controlled trial. Neuraminidase Inhibitor Flu Treatment Investigator Group. AB - BACKGROUND: Use of some antiviral drugs for influenza infection is limited by potential rapid emergence of resistance. We studied the efficacy and safety of oseltamivir, the oral prodrug of the neuraminidase inhibitor GS4071, in adults with naturally acquired laboratory-confirmed influenza. METHODS: We did a randomised controlled trial of 726 previously healthy non-immunised adults with febrile influenza-like illness of up to 36 h duration. Patients were assigned oral oseltamivir 75 mg (n=243), oseltamivir 150 mg (n=245), or placebo (n=238) twice daily for 5 days. We assessed recovery by questionnaire and temperature recordings. The primary endpoint was time to resolution of illness in influenza infected patients. FINDINGS: 475 (66%) patients had confirmed infection. Duration of illness was significantly shorter by 29 h (25% reduction, median duration 87.4 h [95% CI 73.3-104.7], p=0.02) with oseltamivir 75 mg and by 35 h (30%, 81.8 h [68.2-100.0], p=0.01) with oseltamivir 150 mg than with placebo (116.5 h [101.5 137.8]). The effect of oseltamivir was apparent within 24 h of the start of treatment. In patients treated within 24 h of symptom onset, symptoms were alleviated 43 h (37% reduction) and 47 h (40%) earlier with oseltamivir 75 mg and 150 mg, respectively, compared with placebo (75 mg 74.5 h [68.2-98.0], p=0.02; 150 mg 70.7 h [54.0-89.4], p=0.01; placebo 117.5 h [103.0-143.8]). Oseltamivir was associated with lower [corrected] symptom scores, less viral shedding, and improved health, activity, and sleep quality, and was well tolerated. INTERPRETATION: Oseltamivir was effective and well tolerated in the treatment of natural influenza infection in adults. The efficacy, tolerability, and ease of administration warrant further investigation in children, elderly patients, and at-risk patients. PMID- 10866440 TI - Sanctions and childhood mortality in Iraq. AB - BACKGROUND: In 1999 UNICEF, in cooperation with the government of Iraq and the local authorities in the "autonomous" (northern Kurdish) region, conducted two similar surveys to provide regionally representative and reliable estimates of child mortality (the subject of this paper) and maternal mortality. METHODS: In a cross-sectional household survey in the south/centre of Iraq in February and March, 1999, 23105 ever-married women aged 15-49 years living in sampled households were interviewed by trained interviewers with a structured questionnaire that was developed using the Demographic and Health Surveys questionnaire and following a pre-test. In a similar survey in the autonomous region in April and May 14 035 ever-married women age 15-49 were interviewed. FINDINGS: In the south/centre, infant and under-5 mortality increased during the 10 years before the survey, which roughly corresponds to the period following the Gulf conflict and the start of the United Nations sanctions. Infant mortality rose from 47 per 1000 live births during 1984-89 to 108 per 1000 in 1994-99, and under-5 mortality rose from 56 to 131 per 1000 live births. In the autonomous region during the same period, infant mortality declined from 64 to 59 per 1000 and under-5 mortality fell from 80 to 72 per 1000. Childhood mortality was higher among children born in rural areas, children born to women with no education, and in boys, and these differentials were broadly similar in the two regions. INTERPRETATION: Childhood mortality clearly increased after the Gulf conflict and under UN sanctions in the south/centre of Iraq, but in the autonomous region since the start of the Oil-for-Food Programme childhood mortality has begun to decline. Better food and resource allocation to the autonomous region contributed to the continued gains in lower mortality, whereas the situation in the south/centre deteriorated despite the high level of literacy in that region. PMID- 10866441 TI - Paternal concentrations of dioxin and sex ratio of offspring. AB - BACKGROUND: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD or dioxin), is commonly considered the most toxic man-made substance. We have previously shown that high serum concentrations of TCDD in parents from Seveso, Italy, were linked to their having a relative increase in the number of female births after the parents exposure to a release of dioxin in 1976. We have continued the study to determine whether the parents' sex and/or age at exposure affected the sex ratio of their children. METHODS: We measured the TCDD concentrations in serum samples from potentially exposed parents collected in 1976 and 1977, and investigated the sex ratio of their offspring. FINDINGS: Serum samples were collected from 239 men and 296 women. 346 girls and 328 boys were born to potentially exposed parents between 1977 and 1996, showing an increased probability of female births (lower sex ratio) with increasing TCDD concentrations in the serum samples from the fathers (p=0.008). This effect starts at concentrations less than 20 ng per kg bodyweight. Fathers exposed when they were younger than 19 years of age sired significantly more girls than boys (sex ratio 0.38 [95% CI 0.30-0.47]). INTERPRETATION: Exposure of men to TCDD is linked to a lowered male/female sex ratio in their offspring, which may persist for years after exposure. The median concentration of dioxin in fathers in this study is similar to doses that induce epididymal impairments in rats and is about 20 times the estimated average concentration of TCDD currently found in human beings in industrialised countries. These observations could have important public-health implications. PMID- 10866442 TI - Impact of a prevention strategy targeted at vascular-access care on incidence of infections acquired in intensive care. AB - BACKGROUND: Intravascular devices are a leading cause of nosocomial infection. Specific prevention strategies and improved guidelines for the use of intravascular devices can decrease the rate of infection; however, the impact of a combination of these strategies on rates of vascular-access infection in intensive-care units (ICUs) is not known. We implemented a multiple-approach prevention programme to decrease the occurrence of vascular-access infection in an 18-bed medical ICU at a tertiary centre. METHODS: 3154 critically ill patients, admitted between October, 1995, and November, 1997, were included in a cohort study with longitudinal assessment of an overall catheter-care policy targeted at the reduction of vascular-access infections and based on an educational campaign for vascular-access insertion and on device use and care. Incidence of ICU-acquired infections was measured by means of on-site surveillance. FINDINGS: 613 infections occurred in 353 patients (19.4 infections per 100 admissions). The incidence density of exit-site catheter infection was 9.2 episodes per 1000 patient-days before the intervention, and 3.3 episodes per 1000 patient-days afterwards (relative risk 0.36 [95% CI 0.20-0.63]). Corresponding rates for bloodstream infection were 11.3 and 3.8 episodes per 1000 patient-days, respectively (0.33 [0.20-0.56]) due to decreased rates of both microbiologically documented infections and clinical sepsis. Rates of respiratory and urinary-tract infections remained unchanged, whereas those of skin or mucous membrane infections decreased from 11.4 to 7.0 episodes per 1000 patient-days (0.62 [0.41-0.93]). Overall, the incidence of nosocomial infections decreased from 52.4 to 34.0 episodes per 1000 patient-days (0.65 [0.54-0.78]). INTERPRETATION: A multiple-approach prevention strategy, targeted at the insertion and maintenance of vascular access, can decrease rates of vascular access infections and can have a substantial impact on the overall incidence of ICU-acquired infections. PMID- 10866443 TI - Is chemotherapy alone adequate for young women with oestrogen-receptor-positive breast cancer? AB - BACKGROUND: The prognosis of breast cancer in very young women is generally considered to be unfavourable. Therefore, the outcome of adjuvant therapy was analysed in a population of young (<35 years) premenopausal patients treated in four randomised controlled trials. METHODS: Between 1978 and 1993 the International Breast Cancer Study Group (IBCSG) treated 3700 premenopausal and perimenopausal patients with various timing and duration of adjuvant cyclophosphamide, methotrexate, and fluorouracil (CMF with or without low-dose prednisone and oophorectomy). 314 of these women were less than 35 years old at randomisation. FINDINGS: Relapse and death occurred earlier and more often in younger (<35 years) than in older (> or = 35) patients with a 10 year disease free survival of 35% (SE 3) versus 47% (1) (hazard ratio 1.41 [95% CI 1.22-1.62], p<0.001) and overall survival of 49% (3) versus 62% (1) (1.50 [1.28-1.77], p<0.001). Younger patients with oestrogen-receptor positive tumours had a significantly worse disease-free survival than younger patients with oestrogen receptor negative tumours. By contrast, among older patients the disease-free survival was similar irrespective of oestrogen-receptor status. INTERPRETATION: Young premenopausal breast cancer patients treated with adjuvant CMF chemotherapy had higher risk of relapse and death than older premenopausal patients, especially if their tumours expressed oestrogen receptors. The endocrine effects of chemotherapy alone are insufficient for the younger age group and these patients should strongly consider additional endocrine therapies (tamoxifen or ovarian ablation) if their tumours express oestrogen receptors. PMID- 10866444 TI - Assessment of thymic output in adults after haematopoietic stem-cell transplantation and prediction of T-cell reconstitution. AB - BACKGROUND: The potential benefits of haematopoietic stem-cell transplantation are tempered by the depletion of T-cells accompanying this procedure. We used a new technique which quantifies the excisional DNA products of T-cell-receptor (TCR) gene rearrangement to measure thymic output directly in patients with multiple myeloma, and thus assessed the contribution of the thymus to immune recovery after transplantation. METHODS: We studied 40 patients, 34-66 years of age, who had been randomly assigned myeloablative chemotherapy and autologous peripheral-blood haematopoietic stem-cell transplantation with unmanipulated grafts or grafts enriched for CD34 stem cells. CD4 and CD8 T-cell counts were measured, thymic output was estimated serially until 2 years after transplantation, and percentages of naive T-cells were measured. FINDINGS: The production of substantial numbers of new naive T cells by the thymus could be detected by 100 days post-transplant; there was a significant inverse relation between age and recovery of new T cells. In the CD34-unselected group, numbers of TCR-rearrangement excision circles returned to baseline after 2 years, whereas in the CD34-selected group, numbers at 2 years were significantly higher than both baseline numbers (p=0.004), and 2-year numbers in the unselected group (p=0.046). Increased thymic output correlated with, and was predictive of, increased naive T cell numbers and broader T-cell-receptor repertoires. INTERPRETATION: Our results provide evidence that the adult thymus contributes more substantially to immune reconstitution after haematopoietic stem-cell transplantation than was previously thought, and therefore could be a target for therapeutic intervention. PMID- 10866446 TI - Dioxin contamination of feed and food. AB - A preventive action limit for dioxins in feed for broiler chickens and pigs was set to 2 pg toxic equivalents/g feed in Austria. This limit was effective in the detection of feed contamination from an imported mineral additive, and in the prevention of food contamination according to WHO tolerable daily intake. PMID- 10866445 TI - Epilepsy, broken bones, and fatty stools. PMID- 10866447 TI - High fish-specific dioxin concentrations in Finland. AB - Dioxin concentrations in a population that frequently eat fish from the Baltic Sea are comparable to those seen in inhabitants of Seveso, Italy, after the accidental release of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in 1976. Individual variations in the congener patterns in human beings are similar to congener patterns in the fish species consumed. PMID- 10866448 TI - Differential binding of mannose-binding lectin to respiratory pathogens in cystic fibrosis. AB - We have found that mannose-binding lectin binds to Burkholderia cepacia, an important pathogen in patients with cystic fibrosis, and leads to complement activation, but that this is not the case for Pseudomonas aeruginosa, the more common colonising organism in this disease. We suggest that patients with cystic fibrosis with mannose-binding-lectin deficiency will be at particular risk of B cepacia colonisation. PMID- 10866449 TI - Cancer risk in patients on dialysis and after renal transplantation. AB - The increased risk of cancer in patients who have had kidney transplants has mainly been attributed to immunosuppressive therapy; however, the prior period of uraemia and dialysis has also been postulated as a cofactor. We analysed cancer risk retrospectively in a cohort of 4178 patients undergoing renal replacement therapy, of whom 3592 were treated with dialysis alone and 1821 later had transplants. We found that excess cancer risk in such patients occurred after transplantation and not during dialysis. PMID- 10866450 TI - Maternal and fetal hepatitis C virus exposure by intrauterine transfusion. AB - We report a case of accidental exposure to hepatitis C virus by an intrauterine transfusion that resulted in infection of the mother but not the child. PMID- 10866451 TI - Residential radon exposure and adult acute leukaemia. AB - Exposure to radioactive radon gas in homes, from natural sources, is an important public-health issue for many countries. We found no association between household exposure to radon and leukaemia in adults in the UK. PMID- 10866452 TI - Drug users' brains to be removed 2 hours after death. PMID- 10866453 TI - Ovarian-cancer results cause a stir at cancer conference. PMID- 10866454 TI - Embryonic stem cells used to remyelinate injured rat spinal cord neurons. PMID- 10866455 TI - Anaesthetist: the wrong name for the right doctor. PMID- 10866456 TI - Baghdad 2000--rubbish heaps and cesspits. PMID- 10866457 TI - HIV/AIDS issues take centre-stage at World Health Assembly. PMID- 10866458 TI - Protests in India after arrest of HIV/AIDS activists. PMID- 10866460 TI - Do escape mutants explain rapid increases in dengue case-fatality rates within epidemics? AB - During the Cuban dengue epidemics of 1981 and 1997, significant monthly increases were observed in the proportion of total cases that presented as dengue haemorrhagic fever or dengue shock syndrome (DHF/DSS), and in case-fatality rates for both dengue fever and DHF/DSS. We believe that theses increases can be explained by the hypothesis that some of the population of antibodies against dengue 1 virus raised after natural primary infections react with "neutralisation" determinants found on dengue 2 viruses. These heterotypic antibodies do not prevent secondary dengue 2 infections, but serve to down regulate the disease to mild illness or symptomless infections. A population of dengue 2 viruses that replicates in dengue-1-immune hosts escape heterotypic neutralisation. When inoculated into a new dengue-1-immune host, these viruses are free to interact with the more abundant infection-enhancing antibodies to produce severe disease. PMID- 10866459 TI - Preventing HIV: determinants of sexual behaviour. AB - AIDS has Invigorated and distorted the study of sexual behaviour. Because that study began so recently, there remain many unanswered questions about why we have sex at all, why we do sex one way rather than another, or even how we define sex. Yet in every instance in which well-designed and adequately resourced behavioural Interventions have been Implemented, these have netted success in the form of falling HIV incidences or prevalences. But, despite these successes, such interventions remain patchy and poorly supported. Perhaps humankind's traditional aversion for the public discussion of sexual matters underlies this reticence. Or maybe a new era of "creeping absolutism"--in which biomedical advances are given premature credit for what they can achieve in HIV control--has arrived. PMID- 10866461 TI - Fundamental research at primary care level. PMID- 10866462 TI - Increasing gonorrhoea reports--not only in London. PMID- 10866463 TI - Increasing gonorrhoea reports--not only in London. PMID- 10866464 TI - Increasing gonorrhoea reports--not only in London. PMID- 10866465 TI - The Achilles heel of exercise. PMID- 10866466 TI - The Achilles heel of exercise. PMID- 10866467 TI - The Achilles heel of exercise. PMID- 10866468 TI - Crisis of adenoviruses in human gene therapy. PMID- 10866469 TI - Drug interaction of St John's wort with cyclosporin. PMID- 10866470 TI - Growth-hormone treatment and risk of diabetes. PMID- 10866471 TI - Growth-hormone treatment and risk of diabetes. PMID- 10866472 TI - Growth-hormone treatment and risk of diabetes. PMID- 10866473 TI - Double gloving and hydration. PMID- 10866474 TI - Should profitability determine the availability of effective contraception? PMID- 10866475 TI - Antenatal detection of domestic violence. PMID- 10866476 TI - Prioritizing waiting lists. PMID- 10866477 TI - Life on earth. PMID- 10866478 TI - How to check adherence to treatment. PMID- 10866479 TI - In response to the August 1999 article entitled "Mandated tuberculosis screening in a community of homeless people". PMID- 10866480 TI - [Anisakiasis in the year 2000]. PMID- 10866481 TI - [Use and abuse of gastroenterologic technology ("today the science advances sophistication")]. PMID- 10866482 TI - [Atypical presentation of metastatic gastric adenocarcinoma]. PMID- 10866484 TI - [Pancreatic hydatidosis]. PMID- 10866483 TI - [Thrombosis of the portal and superior mesenteric vein associated with protein S deficiency]. PMID- 10866485 TI - [Transmural hematoma of the rectum in a patient on anticoagulant therapy]. PMID- 10866486 TI - [Brief fibrinolysis in a case of acute mesenteric ischemia]. PMID- 10866487 TI - Ventricular arrhythmias and nonsedating antihistamines. PMID- 10866488 TI - Eosinophilic bronchitis is an important cause of chronic cough. PMID- 10866489 TI - [Oleksandr Romanovych Vynnyts'kyi (on the 80th anniversary of his birth)]. PMID- 10866491 TI - Protein folding. PMID- 10866490 TI - Mice, mutations and mammary glands. PMID- 10866492 TI - In the research community it is considered to be highly unethical to publish as original work the same scientific results more than once. PMID- 10866493 TI - Synergistic induction of growth arrest and apoptosis of human myeloma cells by the IL-6 super-antagonist Sant7 and Dexamethasone. PMID- 10866494 TI - Opposing actions of STAT-1 and STAT-3 on the Bcl-2 and Bcl-x promoters. PMID- 10866495 TI - Treatment of chronic: new perspectives. PMID- 10866497 TI - Folic acid and seizures. PMID- 10866496 TI - Equivalent doses of ropinirole and bromocriptine. PMID- 10866498 TI - Partial third nerve palsy caused with chronic Q Fever. PMID- 10866499 TI - Bigger is better but longer is a limitation. PMID- 10866500 TI - [Topics of emerging, re-emerging infectious diseases]. PMID- 10866501 TI - [Rapid diagnosis of infectious diseases; features and limitations of the microscopic examination of clinical specimens]. AB - The diagnosis of infectious diseases relies on the microbiological examinations. The microscopic examination of clinical specimens has been well known as one of the rapid methods in diagnostic microbiology. The reasons for the microscopic examination in the clinical microbiology are emphasized and summarized as the following features and limitations: ADVANTAGES OF MICROSCOPY. The use of a stained smear microscopy by Gram, Ziehl-Neelsen and fluorochrome stains still remains the most available, easy to perform, and inexpensive. This technique can be done routinely in a variety of clinical settings. The direct microscopy provides judging specimen quality, detecting a variety of organisms in clinical specimens, and evaluating the type of inflammatory response. Typical Gram reactions, morphologies and arrangements of the observed organisms may give the presumptive identification of some certain etiological bacteria, yeasts or fungi, and significant of the organisms. The observing organisms and those forms may also guide to the laboratory in selecting appropriate isolation media and culture methods, and aid to the physician in selecting an empirical antibiotic therapy. DISADVANTAGES AND LIMITATIONS OF MICROSCOPY. The microscopy of clinical specimen is much less sensitive than the culture method for detection of small numbers of bacteria. A specimen usually contains 10(4) to 10(5) or more organisms per milliliter before it is likely that organisms will be seen on a smear. The appearance of bacteria on Gram-stained smears does not permit the identification of species. FACTORS AFFECTING TO RESULT OF MICROSCOPIC EXAMINATION. A number of factors may affect the result of the microscopic examinations. Specimen quality, clinical presentations of the patients, presence of the various cellular components in the background of smear, bacteriological statistics, and individuals in the microbiology laboratory are responsible to the reliability and quality of the results. The concordance between results obtained from a microscopy and from further tests may also provide sufficient information to presumptive identification and diagnosis. Laboratory observers, whom should have abundant experiences and receive sufficient training, are requiring for control and quality of the examination. PMID- 10866502 TI - [Pitfalls in rapid diagnosis for infectious disease. Rapid antigen detection for diagnosis]. PMID- 10866503 TI - [Directions for detection procedure of bacterial toxins]. PMID- 10866504 TI - [Limit in rapid identification in microbiology]. PMID- 10866505 TI - [Genetic examination]. PMID- 10866506 TI - 'Infectious web'. PMID- 10866507 TI - [Serial analysis of the electrocardiogram and ventricular remodelling]. PMID- 10866508 TI - 7th meeting of the European Society of Gene Therapy. November 26-28 1999, Munich, Germany. PMID- 10866509 TI - Journal of Gene Medicine 1999 Young Investigator Award. PMID- 10866510 TI - The modern management of interstitial or intramural pregnancy--is MRI and "alloyed" diagnostic gold standard or the real thing? PMID- 10866511 TI - Addendum from Portugal--how about an annotated IFFS surveillance for the millennium? PMID- 10866512 TI - Addendum from Portugal--how about an annotated IFFS surveillance for the new millennium? PMID- 10866513 TI - Antioxidants and male infertility. PMID- 10866514 TI - Potential sources of error with the Makler counting chamber. PMID- 10866515 TI - The most interesting studies are those that don't turn out the way you expected. PMID- 10866516 TI - [Diagnosis and treatment of occult digestive tract hemorrhage. Questions for Professor Jean Boyer]. PMID- 10866517 TI - [Effect of preoperative abstinence on poor postoperative outcome in alcohol misusers: randomized controlled trial]. PMID- 10866518 TI - [Effect of intravenous albumin on renal impairment and mortality in patients with cirrhosis and spontaneous bacterial peritonitis]. PMID- 10866519 TI - Image of the Month. PMID- 10866520 TI - Colonoscopic polypectomy in chronic colitis. PMID- 10866521 TI - Bifidobacteria in necrotizing enterocolitis. PMID- 10866522 TI - Steroids may prevent leukopenia, interfering with response to IV azathioprine in treatment of Crohn's disease. PMID- 10866523 TI - A significant advance in the understanding of intestinal anion secretion. PMID- 10866524 TI - Insulin-like growth factor-I, liver function, and hypogonadism in rats with experimentally induced cirrhosis. PMID- 10866525 TI - Hepatitis C virus and autoimmunity. PMID- 10866526 TI - "Vascular profiles" of regenerative and dysplastic nodules. PMID- 10866527 TI - Collagenous spherulosis versus shadow cell differentiation in endometrioid adenocarcinoma. PMID- 10866528 TI - Spindle cell tumours of the breast. PMID- 10866529 TI - Absence of human herpesvirus-8 DNA in Kaposi's sarcoma following postmastectomy lymphoedema. PMID- 10866530 TI - Microscopic focal cryptitis associated with sodium phosphate bowel preparation. PMID- 10866531 TI - Human immunodeficiency and hepatitis C virus coinfection influenced by interferon gamma. PMID- 10866532 TI - A multivariate approach to the problem of QTL localization. AB - QTL mapping with statistical likelihood-based procedures or asymptotically equivalent regression methods is usually carried out in a univariate way, even if many traits were observed in the experiment. Some proposals for multivariate QTL mapping by an extension of the maximum likelihood method for mixture models or by an application of the canonical transformation have been given in the literature. This paper describes a method of analysis of multitrait data sets, aimed at localization of QTLs contributing to many traits simultaneously, which is based on the linear model of multivariate multiple regression. A special form of the canonical analysis is employed to decompose the test statistic for the general no QTL hypothesis into components pertaining to individual traits and individual, putative QTLs. Extended linear hypotheses are used to formulate conjectures concerning pleiotropy. A practical mapping algorithm is described. The theory is illustrated with the analysis of data from a study of maize drought resistance. PMID- 10866533 TI - Adopting research. PMID- 10866534 TI - Viscero-somatic and somato-visceral spinal reflexes. 1907. PMID- 10866535 TI - Somatic sensory impulses and vertebral lesions. 1921. PMID- 10866536 TI - EMR the web way. PMID- 10866537 TI - Hello HIPAA! PMID- 10866538 TI - Healthcare from a distance. PMID- 10866539 TI - The corner drugstore moves to cyberspace. PMID- 10866540 TI - Techterms. PMID- 10866541 TI - Where is physician leadership in the e-health community? PMID- 10866542 TI - The pager grows up and reaches out. PMID- 10866544 TI - Touch points. PMID- 10866543 TI - Ophthalmology. PMID- 10866545 TI - Heat-labile serotyping of two Campylobacter jejuni strains isolated from patients with Guillain-Barre syndrome and belonging to serotype O19 (Penner) PMID- 10866547 TI - Fraud the old-fashioned way. PMID- 10866548 TI - Academic Mafia. PMID- 10866546 TI - Use of restriction fragment length polymorphism of the PCR-amplified 16S rRNA gene for the identification of Aeromonas spp. PMID- 10866550 TI - Diagnosis of venous thromboembolism in cancer patients receiving palliative care. PMID- 10866549 TI - Subcutaneous methadone in terminally-ill patients. PMID- 10866551 TI - Dynamic responses of series force receptors innervating the opener muscle apodeme in the blue crab, Callinectes sapidus. AB - Receptors monitoring muscle force innervate the opener muscle apodeme in the walking legs of the blue crab, Callinectes sapidus. Biocytin backfills reveal 9 15 bipolar neurons with somata as large as 60 micrometer m positioned at the distal end of the apodeme. Sensory endings insert into the apodeme and are in series with the opener muscle. The axons of these neurons form the opener apodeme sensory nerve that merges with the most distal branch of the opener motor nerve. Recordings reveal that the receptors are not spontaneously active nor do they respond to passive muscle stretch. Isometric muscle contraction evoked by stimulating the opener excitor motor neuron is the adequate stimulus for receptor firing. Most significant is the finding that during contraction, over a wide range of forces, the firing rate of individual receptors closely parallels the rate of change of isometric force. The peak instantaneous frequency typically occurs at the force derivative maximum, but not at maximum force development. Thus, receptors of the opener apodeme sensory nerve more closely monitor changes in isometric force rather than the total force achieved. PMID- 10866552 TI - [Lymphoma and immunodeficiency]. PMID- 10866553 TI - A metamorphosis: doctor to chaplain. PMID- 10866554 TI - A novel subtype of type 1 diabetes mellitus. PMID- 10866555 TI - A novel subtype of type 1 diabetes mellitus. PMID- 10866556 TI - Improved graft survival after renal transplantation in the United States, 1988 to 1996. PMID- 10866557 TI - Improved graft survival after renal transplantation in the United States, 1988 to 1996. PMID- 10866558 TI - Prevalence of gynecologic cancer in women exposed to diethylstilbestrol in utero. PMID- 10866559 TI - Women physicians in academic medicine. PMID- 10866560 TI - Women physicians in academic medicine. PMID- 10866561 TI - Women physicians in academic medicine. PMID- 10866562 TI - Women physicians in academic medicine. PMID- 10866563 TI - Electric razors as a potential vector for viral hepatitis. PMID- 10866564 TI - Bartonella quintana endocarditis in a child. PMID- 10866565 TI - Comment on Thompson E.N; PAIN 82 (1999) 109-110. PMID- 10866566 TI - Comment on: Svendsen et al; some problems with wind-up and its calculation, PAIN 83 (1999) 109-111. PMID- 10866567 TI - Comment on correspondence regarding Wallis et al; PAIN 73 (1997) 15-22,PAIN 81 (1999) 323-326. PMID- 10866568 TI - Is it really possible to do a selective nerve root block? PMID- 10866569 TI - Comments on Mercandante, PAIN 79 (1999) 1-13. PMID- 10866570 TI - [Glossary]. PMID- 10866571 TI - Pulmonary embolism. PMID- 10866572 TI - Chronic cough. PMID- 10866573 TI - Chronic cough. PMID- 10866574 TI - Childhood empyema. PMID- 10866575 TI - Duchenne muscular dystrophy. PMID- 10866576 TI - Medical students' knowledge of tobacco. PMID- 10866577 TI - [Predecessors: veterinarians from earlier times (37). Eduard August von Hering (1799-1881)]. PMID- 10866578 TI - [The role of leukocytes in E. coli mastitis]. PMID- 10866579 TI - [Salmonella in eggs]. PMID- 10866580 TI - Faculty support is essential for success. PMID- 10866581 TI - In any language. PMID- 10866582 TI - Diagnostic dilemma. Gangrenous cholecystitis. PMID- 10866583 TI - Diagnostic dilemma. Digitalis toxicity with a preexisting bundle branch block. PMID- 10866584 TI - Sigmoidoscopy costs and the limits of altruism. PMID- 10866585 TI - Older persons' perceptions of home and hospital as treatment site. PMID- 10866586 TI - Villaret's syndrome in a man with prostate carcinoma. PMID- 10866587 TI - Neuroleptic malignant-like syndrome in an elderly patient caused by abrupt withdrawal of tolcapone, a-catechol-o-methyl transferase inhibitor. PMID- 10866588 TI - Gamma-hydroxybutyrate associated deaths. PMID- 10866589 TI - Severe thyrotoxicosis after parathyroid surgery for hyperparathyroidism. PMID- 10866590 TI - Zoster gangrenosum: necrotizing fasciitis as a complication of herpes zoster. PMID- 10866591 TI - Atypical manifestation of primary syphilis. PMID- 10866592 TI - Streptococcus equinus endocarditis in a patient with pulmonary histiocytosis X. PMID- 10866593 TI - Cryptococcal infection associated with fludarabine therapy. PMID- 10866595 TI - Academic internal medicine: challenges and opportunities--Part I. PMID- 10866594 TI - Short-term treatment with indinavir fails to reduce the glucose requirement in a patient with malignant insulinoma. PMID- 10866597 TI - The hospitalist movement: caution lights flashing at the crossroads. PMID- 10866596 TI - Meta-analysis of the effects of ipratropium bromide in adults with acute asthma. PMID- 10866598 TI - The hospitalist movement: caution lights flashing at the crossroads. PMID- 10866599 TI - Value of chest roentgenography after thoracentesis. PMID- 10866601 TI - Brucella acute abdomen mimicking appendicitis. PMID- 10866602 TI - Rhabdomyolysis in adults with measles virus infection. PMID- 10866603 TI - Development of chiral HPLC for selenoamino acids with ICP-MS detection:application to selenium nutritional supplements. AB - The enantiomeric separation of three underivatized seleno-amino acids, D,L selenocystine, and D,L-selenomethionine, and D,L-selenomethionine, with UV and ICP-MS detection is described. An HPLC column with a chiral crown ether stationary phase and a mobile phase of 0.10 M HCIO4 was used. Absolute detection limits obtained with UV detection ranged from 34.5 to 47.1 ng whereas those obtained with the plasma detector were ca. 40-400 times better. The separations with either detector were good, with the little detector effect on the resolution. Ten commercially available dietary selenium supplements were analyzed using the chiral column to identify and quantify the selenium species present with both detection modes. Selenium species were easily identified using ICP-MS detection, whereas UV detection was not viable because of interferences from the sample matrix and inadequate sensitivity. Selenium species that were unretained using the chiral column were identified using anion exchange chromatography. Total amounts in the samples were also measured using a conventional digestion and enzymatic digestion with ICP-MS detection. PMID- 10866604 TI - Synthesis and properties of a novel family of fluorous triphenylphosphine derivatives AB - A novel approach to the preparation of perfluorotail-functionalized triarylphosphines using a p-silyl substituent as the branching point has been developed. This approach enabled the attachment of between three and nine perfluorotails per phosphorus atom, resulting in the production of highly fluorous tris[p-(1H,1H,2H, 2H-perfluoroalkylsilyl)aryl]phosphines, P[C(6)H(4)-p SiMe(3)(-)(n)()(CH(2)CH(2)C(x)()F(2)(x)()(+1))(n)()](3) (n = 1, 2, 3; x = 6, 8), containing between 50 and 67 wt % fluorine. (31)P NMR studies indicate that the phosphorus atoms, and consequently the sigma-donor and pi-acceptor properties of these phosphines, are not influenced by the electron-withdrawing perfluoroalkyltails. The fluorous triarylphosphines are readily soluble in fluorous solvents and display fluorous phase preference in several fluorous biphasic systems. The phase partitioning of these fluorous ligands, as well as their donor properties, is discussed in relation to their potential for fluorous biphasic catalyst separation. PMID- 10866605 TI - Novel photoluminescent polymers containing oligothiophene and m-phenylene-1,3,4 oxadiazole moieties: synthesis and spectroscopic and electrochemical studies AB - Three conjugated polymers containing oligothiophene units (from one to three thiophene rings) and aromatic 1,3,4-oxadiazole moieties have been successfully synthesized. The polymer structures were characterized and confirmed by (1)H and (13)C NMR, FT-IR, and elemental analysis. Thermogravimetric analysis demonstrated that the polymers are highly thermal stable. Tunable absorption (from 342 to 428 nm) and fluorescence (from 411 to 558 nm) properties of polymers were observed. The electrochemical investigation indicated that the LUMO and HOMO energy levels of the new polymers could be adjusted. It was also revealed by the electrochemical analysis that the polymers have good charge injection properties for both p-type and n-type charge carriers, as well as good color tunable luminescence and film-forming properties, which makes them potentially useful for fabricating efficient light-emitting devices. PMID- 10866606 TI - Ring-opening reactions of oxabicyclic alkene compounds: enantioselective hydride and ethyl additions catalyzed by group 4 metals AB - Titanium and zirconium catalysts selectively catalyze either the ethyl or hydride addition to [2.2.1] 4, 5-bis(methoxymethyl)-7-oxabicycloheptene (6); the ring opened products formed depend on catalyst, temperature, alkylaluminum reagent, and the concentration of alkylaluminum. Bis(neoisomenthylindenyl)zirconium dichloride catalyzes the ethyl addition ring-opening of 6 to produce (1R,2S,3S,6R)-2, 3-bis(methoxymethyl)-6-ethylcyclohex-4-enol (7) in 96% ee. Zirconium catalysts catalyze the ring-opening of [3.2.1] 2, 4-dimethyl-3 (benzyloxy)-8-oxabicyclo-6-octene (7) when ethylmagnesium bromide is used as a reagent. Both hydride and ethyl addition products are obtained at all conditions studied. Bis(neoisomenthylindenyl)zirconium dichloride catalyzes the ethyl addition ring-opening of 7 to produce (1S,2R,3S,4S,7S)-2, 4-dimethyl-3 (benzyloxy)-7-ethyl-5-cyclohexen-1-ol (8) in 48% ee. PMID- 10866607 TI - Stereoelectronic interactions in cyclohexane, 1,3-dioxane, 1, 3-oxathiane, and 1,3-dithiane: W-effect, sigma(C)(-)(X) <--> sigma(C)(-)(H) interactions, anomeric effect-what is really important? AB - Stereoelectronic effects proposed for C-H bonds in cyclohexane, 1, 3-dioxane, 1,3 oxathiane, and 1,3-dithiane were studied computationally. The balance of three effects, namely, sigma(C)(-)(X) --> sigma(C)(-)(H)()eq, sigma(C)(-)(H)()eq --> sigma(C)(-)(X), and n(p)(X) --> sigma(C)(-)(H)()eq interactions, was necessary to explain the relative elongation of equatorial C(5)-H bonds. The role of homoanomeric n(p) --> sigma(C(5))(-)(H)()eq interaction is especially important in dioxane. In dithiane, distortion of the ring by long C-S bonds dramatically increases overlap of sigma(C(5))(-)(H)()eq and sigma(C)(-)(S) orbitals and energy of the corresponding hyperconjugative interaction. Anomeric n(p)(X) --> sigma(C)( )(H)()ax interactions with participation of axial C-H bonds dominate at C(2), C(4), and C(6). The balance of hyperconjugative interactions involving C-H(ax) and C-H(eq) bonds agrees well with the relative bond lengths for all C-H(ax)/C H(eq) pairs in all studied compounds. At the same time, the order of one-bond spin-spin coupling constants does not correlate with the balance of stereoelectronic effects in dithiane and oxathiane displaying genuine reverse Perlin effect. PMID- 10866608 TI - Gaseous SF(3)(+): An efficient electrophilic monofluorinating agent for five membered heteroaromatic compounds AB - Reactions of gaseous SF(3)(+) ions with furan, thiophene, pyrrole, and several of their alkyl derivatives were performed via MS(2) experiments and found to occur readily both by electron abstraction and F(+) transfer. Then, by performing MS(3) experiments, the F(+) transfer products-the protonated monofluorinated molecules were mass-selected and deprotonated by a second reaction with a stronger base. F(+) transfer from gaseous SF(3)(+) followed by deprotonation promotes therefore C-H by C-F replacement in five-membered heteroaromatic compounds and the efficient gas-phase synthesis of their neutral monofluorinated derivatives. PMID- 10866609 TI - Observation of a stable carbocation in a consecutive criegee rearrangement with trifluoroperacetic acid AB - Selective oxidative cleavage-cyclization of adamantane through the bridge carbon was developed in trifluoroperacetic acid (TFPAA). The methyl group in the bridge position was found to be the substituent that directs consecutive oxygen insertion into the cage structure during the course of a Criegee rearrangement. The formation of stable 5-methyl-4,6-dioxabishomoadamant-5-yl cation at -25 degrees C was observed. Stable carboxonium ion formation allows control of the selectivity of further transformations. Hydrolysis leads to the stereospecific formation of endo,endo-3-hydroxy-7-acetoxybicyclo[3.3. 1]nonane. Its single crystal X-ray structure was obtained. An increase in temperature results in deprotonation of the 5-methyl-4, 6-dioxabishomoadamant-5-yl cation to endo-3 trifluoroacetoxybicyclo[3.3.1]non-6-ene, which undergoes further epoxidation with TFPAA and acidic transannular cyclization in trifluoroacetic acid (TFAA). The described reactions can be used as a convenient method for the synthesis of bicyclo[3.3.1]nonane and oxaadamantane derivatives. The proposed mechanism for each transformation, as well as supporting ab initio theoretical calculations of the strain energy and the stabilization energy of the relevant oxacage structures, are discussed. PMID- 10866611 TI - High-yield formation of giant bis(bicyclic) and crypt-tris(bicyclic) molecules under normal reaction conditions AB - We report the unexpected result of the reaction of 1, 3-bis[(9 anthrylmethoxy)methyl]benzene (1a) or 1,3, 5-tris[(9 anthrylmethoxy)methyl]benzene (1b) with tris(2-maleimidoethyl)amine (2) in homogeneous solution leading to giant bis(bicyclic) and crypt-tris(bicyclic) molecules. The anticipated, intractable solids are obtained in a condensed state reaction using an oscillating mill. PMID- 10866610 TI - Divergent routes to chiral cyclobutane synthons from (-)-alpha-pinene and their use in the stereoselective synthesis of dehydro amino acids. AB - Several polyfunctionalized cyclobutane derivatives have been synthesized using commercial (-)-alpha-pinene and (-)-verbenone as chiral precursors. Thus, oxidative cleavage of these compounds by using ruthenium trichloride afforded quantitatively (-)-cis-pinonic and (-)-cis-pinononic acids, respectively, without epimerization. These products were converted into several types of aldehydes, which are the key intermediates in the synthesis of cyclobutane dehydro amino acids via Wittig-Horner condensations with suitable phosphonates. These reactions are highly stereoselective, affording exclusively (Z) isomers, stereochemistry being assessed by NMR experiments. The obtained dehydro amino acids are polyfunctionalized molecules useful for the synthesis of other alpha-amino acids, with additional chiral centers, whose configuration must be induced by the chirality of the terpene employed as a precursor. PMID- 10866612 TI - UV-vis and IR spectral characterization of persistent carbenium ions, generated upon incorporation of cinnamyl alcohols in the acid zeolites HZSM-5 and HMor AB - Cinnamyl alcohol (1) and two derivatives 2 and 3 have been incorporated in dehydrated HMor and HZSM-5 zeolites with the aim to characterize spectroscopically the corresponding carbocations generated within the solids. Product studies of the supernatant liquid phase combined with diffuse reflectance UV-vis and IR spectroscopy provide unequivocal evidence for the carbocations. Thus, cinnamyl alcohol (1) affords the 1,5-diphenylpentadienyl cation in HMor and HZSM-5 as a persistent species. In the case of HMor with larger pore dimensions the bulkier 1-(2'-cinnamyl)-3-phenylpropenyl cation was also spectroscopically detected. No persistent carbocation was observed when the alpha-methylcinnamyl alcohol (2) was incorporated in the acid zeolites, wherein a complete cyclization to 2-methylindene takes place. Finally, incorporation of 2-methyl-4-tolyl-3-buten 2-ol (3) in HZSM-5 allowed detection of the gem-dimethyl-subsituted p methylcinnamyl cation, with a lifetime of hours. This cation is not persistent enough in HMor to be characterized. The present study illustrates how structurally related allylic substrates may give distinct carbenium ions whose persistence depends on the host-guest fit in the interior of the acid zeolites. PMID- 10866613 TI - New synthetic approaches to polycyclic aromatic hydrocarbons and their carcinogenic oxidized metabolites: derivatives of benzo[s]picene, benzo[rst]pentaphene, and dibenzo[b,def]chrysene. AB - A new synthetic approach to polycyclic aromatic compounds is described that entails in the key steps double Suzuki coupling of PAH bisboronic acid derivatives with o-bromoaryl aldehydes to furnish aryl dialdehydes that are converted to larger polycyclic aromatic ring systems by either (a) conversion to diolefins by Wittig reaction followed by photocyclization or (b) reductive cyclization with triflic acid and 1,3-propanediol. This synthetic method provides convenient access to as many as three different polycyclic aromatic ring systems from a single Suzuki coupled intermediate. It was utilized to synthesize substituted derivatives of benzo[s]picene, benzo[rst]pentaphene, dibenzo[b,def]chrysene, and 13,14-dihydro-benz[g]indeno[2,1-a]fluorene, as well as the putative carcinogenic bisdihydrodiol metabolites of benzo[s]picene, benzo[rst]pentaphene, and dibenzo[b,def]chrysene. PMID- 10866614 TI - Intramolecular homolytic displacements. 30. Functional decarbonylative transformations of aldehydes via homolytically induced decomposition of unsaturated peroxyacetals AB - Homolytically induced decompositions of unsaturated peroxyacetals, synthesized from aldehydes, gave alkoxyalkoxyl radicals that yielded alkyl radicals by rapid beta-scission. The latter radicals could react with several types of "transfer agents" to smoothly bring about homolytic decarbonylative functional group transformations of aldehydes into halides, hydrocarbons, xanthates, alkanenitriles, 2-alkyl-3-chloromaleic anhydrides, 1-phenylalk-1-ynes, and ethyl 2-alkylpropenoates. PMID- 10866615 TI - Isomerization reaction of olefin using RuClH(CO)(PPh(3))(3) AB - When methyl 5-(tert-butyldiphenylsilyl)oxy-2-pentenoate was refluxed in toluene in the presence of RuClH(CO)(PPh(3))(3) (5 mol %), double-bond migration took place to afford methyl 5-(tert-butyldiphenylsilyl)oxy-4-pentenoate in high yield. This means that the double bond conjugated with the ester moiety migrates to a deconjugated position by a ruthenium catalyst. We planned to prepare an enol ether from alpha,beta-unsaturated compounds having an ether moiety in a tether using ruthenium-catalyzed isomerization of the double bond. As a result, silyl or benzyl enol ether was obtained from the alpha,beta-unsaturated ester having alcohol protected by the silyl or benzyl group in a tether in high yield. In this reaction, double bond migration of alpha,beta-unsaturated ketone and alpha,beta unsaturated amide took place to produce deconjugated compounds. Moreover, the double bond of alpha, beta-unsaturated ester having a triple or double bond in a molecule migrated to produce conjugated enyne and diene. On the other hand, treatment of a bis-metalated compound having an alpha, beta-unsaturated ester moiety or the double bond in a tether with RuClH(CO)(PPh(3))(3) gave allyl bis metalated compound in good yield. These compounds are useful units in synthetic organic chemistry. PMID- 10866616 TI - Enzymatic synthesis of caged NADP cofactors: aqueous NADP photorelease and optical properties. AB - The synthesis of caged NADP analogues 18, 19, and 20 has been accomplished by utilizing the transglycosidase activity of solubilized NAD glycohydrolase (porcine brain) to incorporate caged nicotinamides 2, 3, and 4 into NADP. The synthesis of several nicotinamides modified at the carboxamide with o-nitrobenzyl photolabile groups is demonstrated as well as their potential for enzymatic transglycosidation. These results further demonstrate the feasibility of direct enzymatic transglycosidation of sterically hindered substrates into NAD(P), although high nicotinamide analogue water solubility was found to be a necessary trait for yield enhancement with certain analogues. Caged analogues were surveyed under aqueous conditions for net NADP photorelease, while the UV and fluorescent properties of both analogues and their photobyproducts were assessed for compatibility with systems that rely on optical monitoring of enzyme activity. A highly water-soluble alpha-methyl-o-nitrobenzyl group 8 was developed for the synthesis of 20 in order to enhance net NADP photorelease. Compound 20 demonstrated a high 75% net NADP photoreleased without substantial UV optical blackening or fluorescent byproducts. Analogues 18 and 19 were shown by ESI/MALDI MS to photogenerate primarily adducts of NADP with deleterious UV and fluorescent properties. Our work stresses the superior release properties conferred by alpha methyl substitution on aqueous carboxamide photorelease from o-nitrobenzyl compounds. PMID- 10866617 TI - Synthesis of enantiopure 3-quinuclidinone analogues with three stereogenic centers: (1S,2R,4S)- and (1S,2S, 4S)-2-(Hydroxymethyl)-1-azabicyclo[2.2.2]octan-5 one and stereocontrol of nucleophilic addition to the carbonyl group. AB - The pseudoenantiomeric title compounds have been prepared from quincorine (QCI) and quincoridine (QCD), respectively, in enantiopure form following an efficient six-step pathway. Nucleophilic attack at the carbonyl group proceeds preferentially from the supposedly more hindered endo pi-face, giving quinuclidinols with natural configuration at C5 (up to 97%). pi-Face selectivity is highest in the QCD series with bulky O-protecting groups, involving an unprecedented 1,7-stereoinduction. PMID- 10866618 TI - Theoretical studies of hetero-diels-alder reactions involving N-sulfinyl dienophiles AB - The gas-phase hetero-Diels-Alder reactions between butadiene and X-substituted sulfinyl dienophiles, O(-)-S(+)=N-X, are investigated theoretically at the B3LYP/6-31G level. The Z-forms of the dienophiles are found to be more stable (by 5-7 kcal mol(-)(1)) than the E-forms. Four modes of cycloadducts are considered: Z-endo; Z-exo; E(X)(-)(endo)(); E(X)(-)(exo)(). Five factors are responsible for the decreasing energetic preferences of the adducts in the order E(X)(-)(endo)() > E(X)(-)(exo)() > Z-endo > Z-exo: (i) The sigma-sigma proximate charge-transfer interactions in the TS; (ii) the relative sizes of the LUMO AO coefficients on S and N atoms; (iii) steric hindrance in the TS; (iv) the levels of the ground state and the LUMOs of the dienophile; (v) bond energies of the C-S and C-N bonds that are formed in the TS. All the reactions proceed concertedly, but the adduct formation is asynchronous. The endo-additions are favored over the exo-additions kinetically (lower DeltaG()) as well as thermodynamically (lower DeltaG degrees ). The major secondary orbital interaction determining the endo preference is that between the lone pair on N (n(N)) and the d(3) (C(3)-C(4)) sigma orbital (n(N)-sigma(d3)) interactions, whereas the larger AO lobe (LUMO) sizes on S favor a greater degree of d(5) (C-S) bond formation than d(6) (C-N) bond. The solvent, C(6)H(6), uniformly lowers the activation barriers so that the energetic preferences in the gas phase between various modes are maintained in solution. PMID- 10866619 TI - Regio- and stereoselective cycloadditions of cyclic nitrones to maleic diamide forced in a peptide: synthesis of potent ligands of human NK-2 receptor. AB - The regioselectivity and the stereoselectivity induced by relatively small peptidomimetic maleic diamide 1 in cycloaddition reactions with cyclic nitrones 2 5 was studied. The high regio- and stereoselectivity observed, sensibly increased by nonpolar solvents, was the effect of a double-asymmetric induction produced by the nitrone substituent on the pseudopeptidic tether. A new class of potent human tachykinin NK-2 receptor ligands was synthesized. PMID- 10866621 TI - Stable enols of carboxylic esters. The poly(methoxycarbonyl)cyclopentadiene systems AB - The structures of the poly(methoxycarbonyl)cyclopentadienes C(5)H(6)( )(n)()(CO(2)Me)(n)(), n = 5 (Cp-5), n = 4 (Cp-4), n = 3 (1, 2,4-; Cp-3) and n = 2 (1,2-; 1,2-Cp-2-I) were investigated. The X-ray diffractions of Cp-5 (known), Cp 4, and Cp-3 showed an enol of ester structure in the solid state. The enolic hydrogen forms a symmetrical hydrogen bond to a neighboring ester carbonyl, so that the vicinal "enolic" CO(2)Me groups in the 1, 2-C(=CO(2)Me)-C(CO(2)Me)(4) moiety are identical. The relevant X-ray parameters for the three enols are similar. The CP-MAS spectra of Cp-3-Cp-5 generally resemble their (13)C NMR spectra in CDCl(3) except for some differences of mostly <1 ppm. The (1)H, (13)C, and (17)O NMR spectra of Cp-3-Cp-5 in CDCl(3) are consistent with those of the hydrogen bonded enols. Most characteristic are the (1)H and (17)O signals of the OH groups at 19.7-20.1 and 221-225 ppm, respectively. Proton addition to sodium 1, 2-bis(methoxycarbonyl)cyclopentadienide gave a mixture of four 1, 2 bis(methoxycarbonyl)cyclopentadienes. The isomer (1,2-Cp-2-I) formed in 10-20% displays delta(O(1)H) at 19.3 ppm and is the enol analogue of Cp-5 whereas its main isomer (30-55%) (1,2-Cp-2-IV) has the ester structure. In CD(3)CN and DMSO d(6) only one signal was observed at room temperature for each type of H, C, or O of Cp-5, suggesting a complete ionization to the symmetrical anion of Cp-5. In contrast, Cp-4 and Cp-3 in CD(3)CN at room temperature display OH signals in both (1)H and (17)O NMR spectra, and Cp-5 shows a broad OH signal in the (1)H spectrum at 240 K. The enol of ester structure is the main species, although exchange with the corresponding anion is possible. On standing in DMF-d(7) at room temperature, new signals are observed for Cp-3 and Cp-4. On raising the temperature in Cl(2)CDCDCl(2), Cp-3-Cp-5 show line broadening and appearance of new signals. These were ascribed to rearrangment and decomposition processes. PMID- 10866620 TI - Enantioselective ester hydrolysis catalyzed by imprinted polymers. AB - Highly cross-linked network polymers prepared by molecular imprinting catalyzed enantioselectively the hydrolysis of N-tert-butoxycarbonyl phenylalanine-p nitrophenyl ester (BOCPheONP). The templates were designed to allow incorporation of the key catalytic elements, found in the proteolytic enzyme chymotrypsin, into the polymer active sites. Three model systems were evaluated. These were constructed from a chiral phosphonate analogue of phenylalanine (series A, C) or L-phenylalanine (series B) attached by a labile ester linkage to an imidazole containing vinyl monomer. Free radical copolymerization of the template with methacrylic acid (MAA) and ethylene glycol dimethacrylate (EDMA) gave a highly cross-linked network polymer. The templates could be liberated from the polymers by hydrolysis, giving catalytically active sites envisaged to contain an enantioselective binding site, a site complementary to a transition state like structure (series A, C), and a hydroxyl, imidazole, and carboxylic acid group at hydrogen bond distance. As predicted, the enantiomer of BOCPheONP complementary to the configuration of the template was preferentially hydrolyzed with D selectivity for the series A polymers (kD/kL = 1.9) and L-selectivity for the series B polymers (kL/kD = 1.2). The maximum rate enhancement, when compared with a control polymer, prepared using a benzoyl-substituted imidazole monomer as template, was 2.5, and comparing with the imidazole monomer in solution, a maximum rate enhancement of 10 was observed. The catalytic activity was higher for polymers subjected to the nucleophilic treatment. This was explained by a higher site density and flexibility of the polymer matrix caused by this treatment. In a comparison of template rebinding to polymers imprinted with a template containing either a carboxylate (planar ground state structure) or a phosphonate (tetrahedral transition state like structure) functionality, it was observed that imprinted polymers are able to discriminate between a transition state like and a ground state structure for transesterification. However the influence of transition state stabilization on the observed rate enhancements remains obscure. Only at acidic pH's was catalysis observed, whereas at basic pH's the polymers inhibit the reaction. At a later stage, the catalytic activity of the polymers for nonactivated D- and L-phenylalanine ethyl esters was investigated. A rate enhancement of up to 3 was observed when compared to the blank. Most important, however, the polymers imprinted with a D template preferentially hydrolyzed the D-ethyl ester and exhibited saturation kinetics. PMID- 10866622 TI - Acylation of five-membered N-heteroaromatic compounds by ruthenium carbonyl catalyzed direct carbonylation at a C-H bond AB - The ruthenium-catalyzed carbonylation at the C-H bond of five-membered N heteroaromatic compounds is described. The reaction of imidazoles with CO and olefins in toluene in the presence of a catalytic amount of Ru(3)(CO)(12) results in carbonylation of the C-H bond at the 4-position (adjacent to the sp(2) nitrogen) of the imidazole ring to give acylated imidazoles in good to high yields. A wide range of olefins can be utilized in the carbonylation reaction, and a variety of functional groups are compatible under the reaction conditions. Other five-membered N-heteroaromatic compounds, such as pyrazoles, oxazoles, and thiazoles, can also be used for the carbonylation reaction, and in all cases, carbonylation takes place exclusively at a C-H bond alpha to the sp(2) nitrogen. The reactivity of the five-membered heterocycles corresponds to the pK(a) of the conjugate acid of these heterocycles. The higher the pK(a) of the substrate, the higher is the reactivity. This indicates that the pK(a) values are related to the ability of the nitrogen atom in the substrates to coordinate to a ruthenium center. The coordination of the substrates to the ruthenium center in the catalyst complex is a necessary prerequisite for the carbonylation to proceed. PMID- 10866623 TI - Preparation of functionalized, conformationally constrained DTPA analogues from L or D-serine and trans-4-hydroxy-L-proline. Hydroxymethyl substituents on the central acetic acid and on the backbone. AB - The enantio- and diastereospecific syntheses of conformationally constrained diethylenetriaminepentaacetic acid (DTPA) analogues that are functionalized with a hydroxymethyl linker substituent on the central acetic acid or on the backbone are described. Key synthetic steps include (i) displacement of the 4-hydroxyl group of N-BOC-trans-4-hydroxy-L-proline benzyl ester, via activation as the triflate, with suitable amines derived from L- or D-serine, (ii) the low temperature alkylation of diethylenetriamines with the triflate of benzyl glycolate, thereby minimizing competitive lactamization, to give DTPA pentabenzyl esters, and (iii) deprotection to afford the corresponding DTPA analogues under very mild hydrogenolysis conditions. PMID- 10866624 TI - Stereoselective synthetic approaches to highly substituted cyclopentanes via electrophilic additions to mono-, di-, and trisubstituted cyclopentenes. AB - Electrophilic additions to allylically substituted alkenes are of broad synthetic utility. The control of stereoselectivity in such reactions has attracted considerable interest. However, the effect of allylic and homoallylic substituents in cyclopentenyl systems has not been investigated systematically. Studies on a series of mono, di-, and trisubstituted cyclopentenes are reported in which trans-vicinal-additions favor a syn-selective approach of electrophiles to the cyclopentene system. The formal addition of HOBr, HOCl, CH(3)SCl, and dimethyl(methylthio)sulfonium tetrafluoroborate (DMTSF)/NaN(3) with a variety of cyclopentene substrates has been carried out, and the effects of various allylic substituents on these selectivities have been examined. Additions of HOBr, HOCl, and DMTSF to highly functionalized substrates proceed predictably with syn selectivity, giving predominantly or exclusively one product. Methanesulfenyl chloride additions are less predictable, but can be tuned by suitable alteration of solvent and substrate. Results have proven useful in total syntheses of (+) trehazolin and (+)-allosamidin. PMID- 10866625 TI - Halichondrin B: synthesis of the C(1)-C(15) subunit. AB - A short and efficient synthesis of the C(1)-C(15) subunit of halichondrin B in its natural configuration is described. The polycyclic caged ketal 3, containing nine asymmetric centers, is prepared in 14 steps from alpha-D-glucoheptonic acid gamma-lactone (7). Key steps in the two similar routes described include EtMgBr promoted pinacol ring expansions of hydroxy mesylates 23 and 34, intramolecular Michael additions of 29 and 37, and a one-pot, HF-induced conversion of 4 to 3involving in situ silyl ether cleavage, acetal hydrolysis, Michael addition, and caged ketal formation. Alternative protocols for carbinol inversion at C(11), one early and one late in the synthetic sequence, are also described. PMID- 10866626 TI - Selective metal cation activation of a DNA alkylating agent: synthesis and evaluation of methyl 1,2,9, 9a-Tetrahydrocyclopropa[c]pyrido[3,2-e]indol-4-one-7 carboxylate (CPyI). AB - The synthesis of methyl 1,2,9,9a-tetrahydrocyclopropa[c]pyrido[3, 2-e]indol-4-one 7-carboxylate (CPyI) containing a one carbon expansion of the C ring pyrrole found in the duocarmycin SA alkylation subunit and its incorporation into analogues of the natural product are detailed. The unique 8-ketoquinoline structure of CPyI was expected to provide a tunable means to effect activation via selective metal cation complexation. The synthesis of CPyI was based on a modified Skraup quinoline synthesis followed by a 5-exo-trig aryl radical cyclization onto an unactivated alkene with subsequent TEMPO trap or 5-exo-trig aryl radical cyclization onto a vinyl chloride for synthesis of the immediate precursor. Closure of the activated cyclopropane, accomplished by an Ar-3' spirocyclization, provided the CPyI nucleus in 10 steps and excellent overall conversion (29%). The evaluation of the CPyI-based agents revealed an intrinsic stability comparable to that of CC-1065 and duocarmycin A but that it is more reactive than duocarmycin SA and the CBI-based agents (3-4x). A pH-rate profile of the addition of nucleophiles to CPyI demonstrated that an acid-catalyzed reaction is observed below pH 4 and that an uncatalyzed reaction predominates above pH 4. The expected predictable activation of CPyI by metal cations toward nucleophilic addition was found to directly correspond to established stabilities of the metal complexes with the addition product (Cu(2+) > Ni(2+) > Zn(2+) > Mn(2+) > Mg(2+)) and provides the opportunity to selectively activate the agents upon addition of the appropriate Lewis acid. This tunable metal cation activation of CPyI constitutes the first example of a new approach to in situ activation of a DNA binding agent complementary to the well-recognized methods of reductive, oxidative, or photochemical activation. Resolution and synthesis of a full set of natural product analogues and subsequent evaluation of their DNA alkylation properties revealed that the CPyI analogues retain identical DNA alkylation sequence selectivity and near-identical DNA alkylation efficiencies compared to the natural products. Consistent with past studies and even with the deep-seated structural change in the alkylation subunit, the agents were found to exhibit potent cytotoxic activity that directly correlates with their inherent reactivity. PMID- 10866627 TI - Synthesis and evaluation of 1,2,8, 8a-Tetrahydrocyclopropa[c]pyrrolo[3,2-e]indol 4(5H)-one, the parent alkylation subunit of CC-1065 and the duocarmycins: impact of the alkylation subunit substituents and its implications for DNA alkylation catalysis. AB - The synthesis of 1,2,8,8a-tetrahydrocyclopropa[c]pyrrolo[3, 2-e]indol-4(5H)-one (CPI), the parent CC-1065 and duocarmycin SA alkylation subunit, is detailed. The parent CPI alkylation subunit lacks the C7 methyl substituent of the CC-1065 alkylation subunit and the C6 methoxycarbonyl group of duocarmycin SA, and their examination permitted the establishment of the impact of these natural product substituents. The studies revealed a CPI stability comparable to the CC-1065 alkylation subunit but which was 6x more reactive than the (+)-duocarmycin SA alkylation subunit, and it displayed the inherent reaction regioselectivity (4:1) of the natural products. The single-crystal X-ray structure of (+)-N-BOC-CPI depicts a near identical stereoelectronic alignment of the cyclopropane accounting for the identical reaction regioselectivity and a slightly diminished vinylogous amide conjugation relative to (+)-N-BOC-DSA suggesting that the stability distinctions stem in part from this difference in the vinylogous amide as well as alterations in the electronic nature of the fused pyrrole. Establishment of the DNA binding properties revealed that the CPI-based agents retain the identical DNA alkylation selectivities of the natural products. More importantly, the C6 methoxycarbonyl group of duocarmycin SA was found to increase the rate (12-13x) and efficiency (10x) of DNA alkylation despite its intrinsic lower reactivity while the CC-1065 C7 methyl group was found to slow the DNA alkylation rate (4x) and lower the alkylation efficiency (ca. 4x). The greater DNA alkylation rate and efficiency for duocarmycin SA and related analogues containing the C6 methoxycarbonyl is proposed to be derived from the extended length that the rigid C6 methoxycarbonyl provides and the resulting increase in the DNA binding-induced conformational change which serves to deconjugate the vinylogous amide and activate the alkylation subunit for nucleophilic attack. The diminished properties resulting from the CC-1065 C7 methyl group may be attributed to the steric impediment this substituent introduces to DNA minor groove binding and alkylation. Consistent with this behavior, the duocarmycin SA C6 methoxycarbonyl group increases biological potency while the CC-1065 C7 methyl group diminishes it. PMID- 10866628 TI - Total synthesis of (+/-)-culmorin and (+/-)-longiborneol: an efficient construction of Tricyclo[6.3.0.0(3,9)]undecan-10-one by intramolecular double Michael addition. AB - The treatment of 4-[(5E)-6-methoxycarbonyl-5-hexenyl]-3, 4-dimethyl-2-cyclopenten 1-one (5) with LHMDS, TMSI-HMDS, Bu(2)OTf-HMDS, or TMSCl-NEt(3)-ZnCl(2) caused the intramolecular double Michael addition to afford tricyclo[6.3.0.0(3, 9)]undecan-10-one 12 in high yields with perfect stereoselectivity. The methodology was further elaborated to achieve efficient total syntheses of (+/-) culmorin (1) and (+/-)-longiborneol (2). The common precursor 13 of them was obtained from 14 in 94% yield as a single isomer by the treatment with LHMDS. After the conversion of 13 into the corresponding acid 24 by hydrolysis, oxidative decarboxylation using S-(1-oxido-2-pyridinyl)-1,1,3, 3 tetramethylthiouronium hexafluorophosphate (HOTT, 27), followed by the Birch reduction, stereoselectively afforded (+/-)-culmorin (1). (+/-)-Longiborneol (2) was synthesized from 24 by the standard transformation. Additionally, the treatment of 24 with Pb(OAc)(4) led to 28 via uncommon migration. Its structure was determined by X-ray analysis after the transformation into the diketone 29. PMID- 10866630 TI - The D parameter of vinyl-substituted 1,3-cyclopentanediyl triplet diradicals as a sensitive tool To determine electronic substituent effects in allylic radicals AB - The zero-field D parameter of the localized vinyl-substituted 1, 3 cyclopentanediyl triplet diradicals V was determined at 77 K in a 2 methyltetrahydrofuran (2-MTHF) glass matrix. Good linear correlations were obtained with the reported alpha-hyperfine coupling constants (r(2) = 0.991, n = 7) and with the semiempirically calculated (PM3) spin densities (r(2) = 0.989, n = 16) of the corresponding allylic monoradicals A. The observed substituent effects are generally larger and, thus, more accurately measured compared to the previously examined aryl-substituted 1, 3-cyclopentanediyl triplet diradicals P. The vinyl-substituted triplet diradicals reflect accurately the delocalizing propensity of substituents, either through hyperconjugative, mesomeric, or inductive effects. For the methyl group, the small but significant stabilization of adjacent radical sites has been clearly demonstrated. In the case of the halogen set, the small but definitive heavy-atom effect has been determined for the bromo (0. 0010 cm(-)(1)) and the iodo (0.0024 cm(-)(1)) substituents. The organometallic substituents SiMe(3) and SnMe(3) are shown to be weak spin acceptors, while spin delocalization for the sulfur series follows the increasing order MeS >> MeSO(2) > MeSO. PMID- 10866629 TI - Tetrathiafulvalenenaphthalenophanes: planar chirality and cis/trans photoisomerization AB - A cyclophane incorporating one 1,5-dioxynaphthalene ring system and one tetrathiafulvalene (TTF) unit bridged by [SCH(2)CH(2)O] linkages has been synthesized. In this cyclophane, the TTF unit can adopt either cis or trans configurations. In addition, the 1, 5-dioxynaphthalene ring system imposes one element of planar chirality on this cyclophane. A second element of planar chirality is introduced by the trans form of the TTF unit. Thus, the cyclophane exists in diastereoisomeric forms as three pairs of enantiomers. The enantiomeric pairs associated with the cis form of the TTF unit, as well as one of those associated with the trans form, have been isolated by crystallization, and their structures assigned in the solid state by single-crystal X-ray analyses. In solution, cis/trans isomerization occurs when either the cis or the trans form of the cyclophane is exposed to light. The photoisomerization reaction can be followed by (1)H NMR and UV-vis spectroscopies, as well as by HPLC. The photoisomerization quantum yield has been measured at two different excitation wavelengths (406 and 313 nm). In both cases, the trans --> cis process (Phi = 0.20 at 406 nm) is much more efficient than the reverse cis --> trans process (Phi = 0.030 at 406 nm). Since the absorption spectra of the trans and cis isomers are different and the quantum yield of the trans --> cis photoisomerization reaction depends on the excitation wavelength, the mole fraction of the two diastereoisomers present at the photostationary state depends on the wavelength of the exciting light. No isomerization occurs when the solutions, regardless of the mole fraction of the two diastereoisomers, are stored in the dark. PMID- 10866631 TI - Chemo- and regioselective cyclohydrocarbonylation of alpha-keto alkynes catalyzed by a zwitterionic rhodium complex and triphenyl phosphite AB - alpha-Keto alkynes react with CO and H(2) in the presence of catalytic quantities of the zwitterionic rhodium complex (eta(6)-C(6)H(5)BPh(3))(-)Rh(+)(1,5-COD) and triphenyl phosphite affording either the 2-, 2(3H)-, or 2(5H)-furanones in 61-93% yields. The cyclohydrocarbonylation is readily accomplished using substrates containing alkyl, aryl, vinyl, and alkoxy groups at the acetylenic terminal, as well as a variety of primary, secondary, and tertiary alkyl, aryl, and heteroaryl groups connected to the ketone functionality. Structural and electronic properties present in the starting materials mediate the chemo- and regioselectivity of the reaction. PMID- 10866632 TI - Highly regioselective thiocarbonylation of conjugated dienes via palladium catalyzed three-component coupling reactions AB - Three-component coupling reaction of conjugated dienes, thiols, and carbon monoxide affords an atom-economical thiocarbonylation of the dienes to give beta,gamma-unsaturated thioesters as the sole products. A catalyst system based on [Pd(OAc)(2)] and Ph(3)P showed excellent catalytic activity. The thiocarbonylation was performed under an atmosphere of carbon monoxide (400 psi) at 110 degrees C in CH(2)Cl(2) for 60 h. A wide variety of thioesters were synthesized in good to excellent yields from easily accessible starting materials. The reaction is believed to proceed via a eta(3)-allylpalladium intermediate. The thiocarbonylation, which is applicable to a wide variety of conjugated dienes, occurs in high regioselectivity, the latter dependent on the steric characteristics and stability of the eta(3)-allylpalladium complex. PMID- 10866633 TI - Synthesis of the C29-C44 portion of spongistatin 1 (altohyrtin A). AB - Two synthetic approaches to the C29-C44 portion of spongistatin 1 (altohyrtin A) have been developed. The key step of the first approach relies on the Claisen rearrangement of glucal 18 to provide ester 20a. This intermediate was advanced to silyl enol ether 30, which was coupled under Mukaiyama aldol conditions with aldehyde 3. Cyclization of this aldol adduct completed our first synthesis of the C29-C44 portion of spongistatin 1, requiring 25 total steps and occurring in 2.4% yield over the longest linear sequence (21 steps). We have also developed a second-generation approach based on the C-glycosidation of glucal 43. Through equilibration of the corresponding C-glycosides 49a/b and 50a/b the desired C glycoside (50a) was obtained in good yield. Aldol condensation of this ketone provided cyclization precursor 67, which undergoes acid-catalyzed ketalization to close the E-ring of the spongistatins. An oxidation/reduction protocol was employed to set the C37 stereocenter. Protection of the C37 carbonol and selective unmasking of the C44 carbonol completed our second generation synthesis. This approach requires 27 steps and occurred in 13.2% yield over the longest linear sequence (18 steps). PMID- 10866634 TI - 13C-labeled platinum(IV)-carbohydrate complexes: structure determination based on (1)H-(1)H, (13)C-(1)H, and (13)C-(13)C spin-spin coupling constants. AB - The reaction of D-mannose and D-allose with [PtMe(3)(Me(2)CO)(3)]BF(4) 1 in acetone affords complexes [PtMe(3)L]BF(4) 5 and 6 (5, L = alpha-D-mannofuranose; 6, L = beta-D-allofuranose). The coordination mode and conformation of the carbohydrate ligands in 5 and 6 in acetone-d(6) have been determined from an analysis of J(HH), J(CH), and J(CC) in complexes formed using site-specific (13)C labeled D-mannose and D-allose. These coupling data are compared to those measured in (13)C-labeled complex [PtMe(3)L]BF(4) 2 (L = 1, 2-O-isopropylidene alpha-D-glucofuranose) and 1, 2-O-isopropylidene-alpha-D-glucofuranose 3, whose solid-state structures are known, and in (13)C-labeled 1,2;5, 6-di-O isopropylidene-alpha-D-glucofuranose 4. The preferred furanose ring conformations in 2 and 5 are very similar ((3)E/E(4) and E(4)/(o)E/E(1), respectively; eastern hemisphere of the pseudorotational itinerary), with platinum coordination involving O3, O5, and O6 of the saccharide. In contrast, the furanose ring of 6 prefers an (4)E/E(o)/(1)E geometry (western hemisphere of the pseudorotational itinerary) resulting from altered complexation involving O1, O5, and O6. Couplings within the exocyclic fragments of 2, 5, and 6 also support the existence of two different platinum coordination modes. In addition to establishing the structures and conformations of 2, 5, and 6 in solution, one-, two-, and three-bond J(CH) and J(CC) observed in these complexes provide new insights into the effect of structure and conformation on the magnitudes of these couplings in saccharides. Weak platinum(IV) complexation with the carbohydrate conformationally restricts the furanose and exocyclic fragment without introducing undesirable structural strain, thereby allowing more reliable correlations between structure and coupling magnitude. PMID- 10866635 TI - Photochemistry of aryl tert-butyl ethers in methanol: the effect of substituents on an excited state cleavage reaction AB - The photolysis of a set of 10 substituted aryl tert-butyl ethers, 8a-j, in methanol gave, as the major product, the corresponding phenol along with tert butyl-substituted phenols resulting from photo-Fries reaction. The corresponding 4-cyanophenyl 1-adamantyl ether, 9, also gave 1-methoxyadamantane 16 (16%), indicating that, at least for this ether, some of the products were ion-derived. Quenching studies with 2,3-dimethylbutadiene for the tert-butyl ethers indicated that these reactions were occurring from the singlet excited state. Rate constants for the reaction, obtained from quantum yields and singlet lifetimes, were found to correlate reasonably well with sigma(h)()(nu) values, rho = -0.77 (r = 0.975), a result that is unexpected for a reaction where the polarity of the bond breaking in the transition state is expected to be -O(delta-). C(delta+). PMID- 10866636 TI - Stannyl radical-mediated cleavage of pi-deficient heterocyclic sulfones. Synthesis Of alpha-fluoro esters AB - Treatment of ethyl 2-(pyridin-2-ylsulfonyl)hexanoate with tributylstannane and azobis(2-methyl-2-propanitrile) (AIBN) in benzene at reflux for 36 h resulted in hydrogenolysis to give ethyl hexanoate (60%), whereas no reaction was observed after 48 h at reflux with ethyl 2-(phenylsulfonyl)hexanoate. Ethyl 2-(pyrimidin-2 ylsulfonyl)hexanoate underwent quantitative hydrogenolysis within 1 h under these conditions. This represents a mild new methodology for removal of the synthetically useful sulfone moiety. Substitution of Bu(3)SnD for Bu(3)SnH gave access to alpha-deuterium-labeled esters. Treatment of the alpha-(pyrimidin-2 ylsulfonyl) enolates derived from several esters with Selectfluor gave high yields of the 2-fluoro-2-(pyrimidin-2-ylsulfonyl)alkanoates, which were smoothly desulfonylated [Bu(3)SnH (2 equiv)/AIBN/benzene/Delta] to give 2 fluoroalkanoates. "Catalytic" tin hydride, generated from tribuytltin chloride (0.15 equiv) and excess polymethylhydrosiloxane in the presence of potassium fluoride, also effected removal of the pi-deficient alpha-(pyrimidin-2 ylsulfonyl) moiety from acid derivatives in high yields. Desulfonylation is suggested to proceed via alkoxy ketyl-type radicals and tin enolates. PMID- 10866637 TI - Efficient approach to 4-oxo-2-alkenylphosphonates via regiospecific friedel crafts acylation AB - Treatment of allylic and vinylic phosphonates with excess LiHMDS, followed by addition of chlorotrimethylsilane, afforded alpha- and gamma- silylated allylic phosphonate mixtures. Without separation, these mixtures underwent the Friedel Crafts reaction and base-promoted isomerization to give 4-oxo-2 alkenylphosphonates, which can serve as building blocks for the construction of polyethylenic chains. PMID- 10866638 TI - Regioselective intramolecular oxidation of phenols and anisoles by dioxiranes generated in situ AB - A novel method for regioselective oxidation of phenols and anisoles has been developed in which dioxiranes, generated in situ from ketones and Oxone, oxidize phenol derivatives in an intramolecular fashion. A series of ketones with electron-withdrawing groups, such as CF(3), COOMe, and CH(2)Cl, were attached to phenols, anisoles, or aryl rings via a C(2) or C(3) methylene linker. In a homogeneous solvent system of CH(3)CN and H(2)O, oxidation of phenol derivatives 1-10 afforded spiro 2-hydroxydienones in 24-55% yields regardless of the presence of other substituents (ortho Me, meta Me or Br) on the aryl ring and the length of the linker. Experimental evidences were provided to support the mechanism that involves a regioselective pi bond epoxidation of aryl rings followed by epoxide rearrangement and hemiketal formation. PMID- 10866639 TI - Stereoselective, oxidative one-carbon degradation of Alkyl(dimethyl)phosphine boranes. Synthesis Of enantiomerically enriched secondary phosphine-boranes PMID- 10866640 TI - Synthesis of terpenoid unsaturated 1,4-dialdehydes. Pi-facial selectivity in the Diels-Alder reaction of the 1-vinyl-2-methylcyclohexene moiety of polycyclic systems with DMAD. PMID- 10866641 TI - Efficient synthesis of novel 3-(Het)arylanthranilic acids via a suzuki cross coupling reaction of 7-iodoisatin with (Het)arylboronic acids in water PMID- 10866643 TI - A facile synthesis of a chiral furan diol from glycals catalyzed by indium trichloride PMID- 10866642 TI - Comparisons of ion pair acidities of some acidic carbon acids. PMID- 10866644 TI - Synthesis of (+)-neplanocin A from a chromium-carbene complex-derived optically active butenolide. PMID- 10866645 TI - The first stable enantiomerically pure chiral N-H oxaziridines: synthesis and reactivity PMID- 10866646 TI - Novel examples of 3-aza-grob fragmentation PMID- 10866648 TI - Facile synthesis of 11-membered C(2) symmetric chiral binaphthyl ketone via Co(salen)-catalyzed macrolactonization PMID- 10866647 TI - Sodium percarbonate as an oxygen source for MTO catalyzed epoxidations. PMID- 10866649 TI - An efficient preparation of TIPS-halofluoropropyne and its application to the diastereoselective synthesis of propargylic fluorohydrins PMID- 10866650 TI - Stability and structure of doubly N-confused porphyrins. PMID- 10866651 TI - A genetic screen for zygotic embryonic lethal mutations affecting cuticular morphology in the wasp Nasonia vitripennis. AB - We have screened for zygotic embryonic lethal mutations affecting cuticular morphology in Nasonia vitripennis (Hymenoptera; Chalcidoidea). Our broad goal was to investigate the use of Nasonia for genetically surveying conservation and change in regulatory gene systems, as a means to understand the diversity of developmental strategies that have arisen during the course of evolution. Specifically, we aim to compare anteroposterior patterning gene functions in two long germ band insects, Nasonia and Drosophila. In Nasonia, unfertilized eggs develop as haploid males while fertilized eggs develop as diploid females, so the entire genome can be screened for recessive zygotic mutations by examining the progeny of F1 females. We describe 74 of >100 lines with embryonic cuticular mutant phenotypes, including representatives of coordinate, gap, pair-rule, segment polarity, homeotic, and Polycomb group functions, as well as mutants with novel phenotypes not directly comparable to those of known Drosophila genes. We conclude that Nasonia is a tractable experimental organism for comparative developmental genetic study. The mutants isolated here have begun to outline the extent of conservation and change in the genetic programs controlling embryonic patterning in Nasonia and Drosophila. PMID- 10866652 TI - Bias and Sampling Error of the Estimated Proportion of Genotypic Variance Explained by Quantitative Trait Loci Determined From Experimental Data in Maize Using Cross Validation and Validation With Independent Samples. AB - Cross validation (CV) was used to analyze the effects of different environments and different genotypic samples on estimates of the proportion of genotypic variance explained by QTL (p). Testcrosses of 344 F(3) maize lines grown in four environments were evaluated for a number of agronomic traits. In each of 200 replicated CV runs, this data set was subdivided into an estimation set (ES) and various test sets (TS). ES were used to map QTL and estimate p for each run (p(ES)) and its median (p(ES)) across all runs. The bias of these estimates was assessed by comparison with the median (p(TS.ES)) obtained from TS. We also used two independent validation samples derived from the same cross for further comparison. The median p(ES) showed a large upward bias compared to p(TS.ES). Environmental sampling generally had a smaller effect on the bias of p(ES) than genotypic sampling or both factors simultaneously. In independent validation, p(TS.ES) was on average only 50% of p(ES). A wide range among p(ES) reflected a large sampling error of these estimates. QTL frequency distributions and comparison of estimated QTL effects indicated a low precision of QTL localization and an upward bias in the absolute values of estimated QTL effects from ES. CV with data from three QTL studies reported in the literature yielded similar results as those obtained with maize testcrosses. We therefore recommend CV for obtaining asymptotically unbiased estimates of p and consequently a realistic assessment of the prospects of MAS. PMID- 10866653 TI - The 2'-5' oligoadenylate/RNase L/RNase L inhibitor pathway regulates both MyoD mRNA stability and muscle cell differentiation. AB - The 2'-5' oligoadenylate (2-5A)/RNase L pathway is one of the enzymatic pathways induced by interferon. RNase L is a latent endoribonuclease which is activated by 2-5A and inhibited by a specific protein known as RLI (RNase L inhibitor). This system has an important role in regulating viral infection. Additionally, variations in RNase L activity have been observed during cell growth and differentiation but the significance of the 2-5A/RNase L/RLI pathway in these latter processes is not known. To determine the roles of RNase L and RLI in muscle differentiation, C2 mouse myoblasts were transfected with sense and antisense RLI cDNA constructs. Importantly, the overexpression of RLI in C2 cells was associated with diminished RNase L activity, an increased level of MyoD mRNA, and accelerated kinetics of muscle differentiation. Inversely, transfection of the RLI antisense construct was associated with increased RNase L activity, a diminished level of MyoD mRNA, and delayed differentiation. In agreement with these data, MyoD mRNA levels were also decreased in C2 cells transfected with an inducible RNase L construct. The effect of RNase L activity on MyoD mRNA levels was relatively specific because expression of several other mRNAs was not altered in C2 transfectants. Therefore, RNase L is directly involved in myoblast differentiation, probably through its role in regulating MyoD stability. This is the first identification of a potential mRNA target for RNase L. PMID- 10866654 TI - Histone binding protein RbAp48 interacts with a complex of CREB binding protein and phosphorylated CREB. AB - A CREB-CREB binding protein (CBP) complex was used as bait to screen a mouse embryo cDNA library in yeast. One of the strongest interactions identified the histone binding protein RbAp48. RbAp48 also interacted weakly with CBP alone but did not interact with phosphorylated or nonphosphorylated CREB. CBP (or its homologue p300) from HeLa cell nuclear extracts coimmunoprecipitated with RbAp48 and its homologue RbAp46 and bound to a glutathione S-transferase-RbAp48 fusion protein. This interaction was stimulated by the addition of phosphorylated CREB and allowed the association of core histones and mononucleosomes in an acetylation-dependent manner. RbAp48 lowered the K(m) of CBP histone acetylase activity and facilitated p300-mediated in vitro transcription of a chromatinized template in the presence of acetylcoenzyme A. These data indicate that the association of phosphorylated CREB with CBP promotes the binding of RbAp48 and its homologue RbAp46, allowing the formation of a complex that facilitates histone acetylation during transcriptional activation. PMID- 10866655 TI - Activation of MEK kinase 1 by the c-Abl protein tyrosine kinase in response to DNA damage. AB - The c-Abl protein tyrosine kinase is activated by certain DNA-damaging agents and regulates induction of the stress-activated c-Jun N-terminal protein kinase (SAPK). Here we show that nuclear c-Abl associates with MEK kinase 1 (MEKK-1), an upstream effector of the SEK1-->SAPK pathway, in the response of cells to genotoxic stress. The results demonstrate that the nuclear c-Abl binds to MEKK-1 and that c-Abl phosphorylates MEKK-1 in vitro and in vivo. Transient-transfection studies with wild-type and kinase-inactive c-Abl demonstrate c-Abl kinase dependent activation of MEKK-1. Moreover, c-Abl activates MEKK-1 in vitro and in response to DNA damage. The results also demonstrate that c-Abl induces MEKK-1 mediated phosphorylation and activation of SEK1-SAPK in coupled kinase assays. These findings indicate that c-Abl functions upstream of MEKK-1-dependent activation of SAPK in the response to genotoxic stress. PMID- 10866656 TI - Naturally occurring dicistronic cricket paralysis virus RNA is regulated by two internal ribosome entry sites. AB - Cricket paralysis virus is a member of a group of insect picorna-like viruses. Cloning and sequencing of the single plus-strand RNA genome revealed the presence of two nonoverlapping open reading frames, ORF1 and ORF2, that encode the nonstructural and structural proteins, respectively. We show that each ORF is preceded by one internal ribosome entry site (IRES). The intergenic IRES is located 6,024 nucleotides from the 5' end of the viral RNA and is more active than the IRES located at the 5' end of the RNA, providing a mechanistic explanation for the increased abundance of structural proteins relative to nonstructural proteins in infected cells. Mutational analysis of this intergenic region IRES revealed that ORF2 begins with a noncognate CCU triplet. Complementarity of this CCU triplet with sequences in the IRES is important for IRES function, pointing to an involvement of RNA-RNA interactions in translation initiation. Thus, the cricket paralysis virus genome is an example of a naturally occurring, functionally dicistronic eukaryotic mRNA whose translation is controlled by two IRES elements located at the 5' end and in the middle of the mRNA. This finding argues that eukaryotic mRNAs can express multiple proteins not only by polyprotein processing, reinitiation and frameshifting but also by using multiple IRES elements. PMID- 10866657 TI - Transcriptional regulation of the TATA-binding protein by Ras cellular signaling. AB - Our previous studies have demonstrated that the level of the central transcription factor TATA-binding protein (TBP) is increased in cells expressing the hepatitis B virus (HBV) X protein through the activation of the Ras signaling pathway, which serves to enhance both RNA polymerase I and III promoter activities. To understand the mechanism by which TBP is regulated, we have investigated whether enhanced expression is modulated at the transcriptional level. Nuclear run-on assays revealed that the HBV X protein increases the number of active transcription complexes on the TBP gene. In transient-transfection assays with both transformed and primary hepatocytes, the human TBP promoter was shown to be induced by expression of the HBV X protein in a Ras-dependent manner, requiring both Ral guanine nucleotide dissociation stimulator (RalGDS) and Raf signaling. Transient overexpression of TBP did not affect TBP promoter activity. To further delineate the downstream Ras-mediated events contributing to TBP promoter regulation in primary rat hepatocytes, the best-characterized Ras effectors, Raf, phosphoinositide 3-kinase (PI-3 kinase), and RalGDS, were examined. Activation of either Raf or RalGDS, but not that of PI-3 kinase, was sufficient to induce TBP promoter activity. Both Raf- and RalGDS-mediated induction required the activation of mitogen-activated protein kinase kinase (MEK). In addition, another distinct Ras-activated pathway, which does not require MEK activation, appears to induce TBP promoter activity. Analysis of the DNA sequence requirement within the TBP promoter responsible for these regulatory events defined three distinct regions that modulate the abilities of Raf, RalGDS, and the Ras-dependent, MEK-independent pathways to regulate human TBP promoter activity. Together, these results provide new evidence that TBP can be regulated at the transcriptional level and identify three distinct Ras-activated pathways that modulate this central eukaryotic transcription factor. PMID- 10866658 TI - Phosphorylation of the PTEN tail regulates protein stability and function. AB - The PTEN gene is a tumor suppressor localized in the frequently altered chromosomal region 10q23. The tumor suppressor function of the PTEN protein (PTEN) has been linked to its ability to dephosphorylate the lipid second messenger phosphatidylinositol 3,4, 5-trisphosphate and phosphatidylinositol 3,4 bisphosphate and, by doing so, to antagonize the phosphoinositide 3-kinase pathway. The PTEN protein consists of an amino-terminal phosphatase domain, a lipid binding C2 domain, and a 50-amino-acid C-terminal domain (the "tail") of unknown function. A number of studies have shown that the tail is dispensable for both phosphatase activity and blocking cell growth. Here, we show that the PTEN tail is necessary for maintaining protein stability and that it also acts to inhibit PTEN function. Thus, removing the tail results in a loss of stability but does not result in a loss of function because the resultant protein is more active. Furthermore, tail-dependent regulation of stability and activity is linked to the phosphorylation of three residues (S380, T382, and T383) within the tail. Therefore, the tail is likely to mediate the regulation of PTEN function through phosphorylation. PMID- 10866659 TI - An engineered PAX3-KRAB transcriptional repressor inhibits the malignant phenotype of alveolar rhabdomyosarcoma cells harboring the endogenous PAX3-FKHR oncogene. AB - The t(2;13) chromosomal translocation in alveolar rhabdomyosarcoma tumors (ARMS) creates an oncogenic transcriptional activator by fusion of PAX3 DNA binding motifs to a COOH-terminal activation domain derived from the FKHR gene. The dominant oncogenic potential of the PAX3-FKHR fusion protein is dependent on the FKHR activation domain. We have fused the KRAB repression module to the PAX3 DNA binding domain as a strategy to suppress the activity of the PAX3-FKHR oncogene. The PAX3-KRAB protein bound PAX3 target DNA sequences and repressed PAX3 dependent reporter plasmids. Stable expression of the PAX3-KRAB protein in ARMS cell lines resulted in loss of the ability of the cells to grow in low-serum or soft agar and to form tumors in SCID mice. Stable expression of a PAX3-KRAB mutant, which lacks repression function, or a KRAB protein alone, lacking a PAX3 DNA binding domain, failed to suppress the ARMS malignant phenotype. These data suggest that the PAX3-KRAB repressor functions as a DNA-binding-dependent suppressor of the transformed phenotype of ARMS cells, probably via competition with the endogenous PAX3-FKHR oncogene and repression of target genes required for ARMS tumorigenesis. The engineered repressor approach that directs a transcriptional repression domain to target genes deregulated by the PAX3-FKHR oncogene may be a useful strategy to identify the target genes critical for ARMS tumorigenesis. PMID- 10866660 TI - Postcleavage sequence specificity in V(D)J recombination. AB - Unintended DNA rearrangements in a differentiating lymphocyte can have severe, oncogenic consequences, but the mechanisms for avoiding pathogenic outcomes in V(D)J recombination are not well understood. The first level at which fidelity is instituted is in discrimination by the recombination proteins between authentic and inauthentic recombination signal sequences. Nevertheless, this discrimination is not absolute and cannot fully eliminate targeting errors. To learn more about the basis of specificity during V(D)J recombination, we have investigated whether it is possible for the recombination machinery to detect an inaccurately targeted sequence subsequent to cleavage. These studies indicate that even postcleavage steps in V(D)J recombination are sequence specific and that noncanonical sequences will not efficiently support the resolution of recombination intermediates in vivo. Accordingly, interventions after a mistargeting event conceivably occur at a late stage in the joining process and the likelihood may well be crucial to enforcing fidelity during antigen receptor gene rearrangement. PMID- 10866661 TI - AIB1 is a conduit for kinase-mediated growth factor signaling to the estrogen receptor. AB - Growth factor modulation of estrogen receptor (ER) activity plays an important role in both normal estrogen physiology and the pathogenesis of breast cancer. Growth factors are known to stimulate the ligand-independent activity of ER through the activation of mitogen-activated protein kinase (MAPK) and the direct phosphorylation of ER. We found that the transcriptional activity of AIB1, a ligand-dependent ER coactivator and a gene amplified preferentially in ER positive breast cancers, is enhanced by MAPK phosphorylation. We demonstrate that AIB1 is a phosphoprotein in vivo and can be phosphorylated in vitro by MAPK. Finally, we observed that MAPK activation of AIB1 stimulates the recruitment of p300 and associated histone acetyltransferase activity. These results suggest that the ability of growth factors to modulate estrogen action may be mediated through MAPK activation of the nuclear receptor coactivator AIB1. PMID- 10866662 TI - A new family of nuclear receptor coregulators that integrate nuclear receptor signaling through CREB-binding protein. AB - We describe the cloning and characterization of a new family of nuclear receptor coregulators (NRCs) which modulate the function of nuclear hormone receptors in a ligand-dependent manner. NRCs are expressed as alternatively spliced isoforms which may exhibit different intrinsic activities and receptor specificities. The NRCs are organized into several modular structures and contain a single functional LXXLL motif which associates with members of the steroid hormone and thyroid hormone/retinoid receptor subfamilies with high affinity. Human NRC (hNRC) harbors a potent N-terminal activation domain (AD1), which is as active as the herpesvirus VP16 activation domain, and a second activation domain (AD2) which overlaps with the receptor-interacting LXXLL region. The C-terminal region of hNRC appears to function as an inhibitory domain which influences the overall transcriptional activity of the protein. Our results suggest that NRC binds to liganded receptors as a dimer and this association leads to a structural change in NRC resulting in activation. hNRC binds CREB-binding protein (CBP) with high affinity in vivo, suggesting that hNRC may be an important functional component of a CBP complex involved in mediating the transcriptional effects of nuclear hormone receptors. PMID- 10866663 TI - Robust mRNA transcription in chicken DT40 cells depleted of TAF(II)31 suggests both functional degeneracy and evolutionary divergence. AB - We have employed gene targeting coupled with conditional expression to construct a chicken DT40 cell line in which a tetracycline (Tet)-repressible promoter is exclusively responsible for expression of cTAF(II)31, a histone-like TAF(II) residing in both the transcription factor TFIID and the histone acetylase complex PCAF/SAGA. Tet addition resulted in rapid loss of cTAF(II)31 mRNA and protein, eventually leading to apoptotic cell death. Significantly, five of six other TAF(II)s tested were also rapidly depleted, but levels of the TATA binding protein and subunits of PCAF/SAGA were at most modestly compromised. Strikingly, pulse-labeling experiments indicate that total poly(A)(+) mRNA transcription was not significantly reduced after cTAF(II)31 depletion, and steady-state levels of several specific transcripts remained the same or decreased only mildly. Moreover, activation of c-fos transcription following serum starvation occurred efficiently in the absence of cTAF(II)31. These data, which contrast with comparable studies in yeast, strongly suggest that cTAF(II)31 and perhaps other TAF(II)s are not essential for general mRNA transcription in DT40 cells. We propose that this is due to extensive functional degeneracy in the highly complex metazoan transcriptional machinery. PMID- 10866664 TI - Tat modifies the activity of CDK9 to phosphorylate serine 5 of the RNA polymerase II carboxyl-terminal domain during human immunodeficiency virus type 1 transcription. AB - Tat stimulates human immunodeficiency virus type 1 (HIV-1) transcriptional elongation by recruitment of carboxyl-terminal domain (CTD) kinases to the HIV-1 promoter. Using an immobilized DNA template assay, we have analyzed the effect of Tat on kinase activity during the initiation and elongation phases of HIV-1 transcription. Our results demonstrate that cyclin-dependent kinase 7 (CDK7) (TFIIH) and CDK9 (P-TEFb) both associate with the HIV-1 preinitiation complex. Hyperphosphorylation of the RNA polymerase II (RNAP II) CTD in the HIV-1 preinitiation complex, in the absence of Tat, takes place at CTD serine 2 and serine 5. Analysis of preinitiation complexes formed in immunodepleted extracts suggests that CDK9 phosphorylates serine 2, while CDK7 phosphorylates serine 5. Remarkably, in the presence of Tat, the substrate specificity of CDK9 is altered, such that the kinase phosphorylates both serine 2 and serine 5. Tat-induced CTD phosphorylation by CDK9 is strongly inhibited by low concentrations of 5, 6 dichloro-1-beta-D-ribofuranosylbenzimidazole, an inhibitor of transcription elongation by RNAP II. Analysis of stalled transcription elongation complexes demonstrates that CDK7 is released from the transcription complex between positions +14 and +36, prior to the synthesis of transactivation response (TAR) RNA. In contrast, CDK9 stays associated with the complex through +79. Analysis of CTD phosphorylation indicates a biphasic modification pattern, one in the preinitiation complex and the other between +36 and +79. The second phase of CTD phosphorylation is Tat-dependent and TAR-dependent. These studies suggest that the ability of Tat to increase transcriptional elongation may be due to its ability to modify the substrate specificity of the CDK9 complex. PMID- 10866665 TI - Human Slug is a repressor that localizes to sites of active transcription. AB - Snail/Slug family proteins have been identified in diverse species of both vertebrates and invertebrates. The proteins contain four to six zinc fingers and function as DNA-binding transcriptional regulators. Various members of the family have been demonstrated to regulate cell movement, neural cell fate, left-right asymmetry, cell cycle, and apoptosis. However, the molecular mechanisms of how these regulators function and the target genes involved are largely unknown. In this report, we demonstrate that human Slug (hSlug) is a repressor and modulates both activator-dependent and basal transcription. The repression depends on the C terminal DNA-binding zinc fingers and on a separable repression domain located in the N terminus. This domain may recruit histone deacetylases to modify the chromatin and effect repression. Protein localization study demonstrates that hSlug is present in discrete foci in the nucleus. This subnuclear pattern does not colocalize with the PML foci or the coiled bodies. Instead, the hSlug foci overlap extensively with areas of the SC-35 staining, some of which have been suggested to be sites of active splicing or transcription. These results lead us to postulate that hSlug localizes to target promoters, where activation occurs, to repress basal and activator-mediated transcription. PMID- 10866666 TI - ATF6 as a transcription activator of the endoplasmic reticulum stress element: thapsigargin stress-induced changes and synergistic interactions with NF-Y and YY1. AB - ATF6, a member of the leucine zipper protein family, can constitutively induce the promoter of glucose-regulated protein (grp) genes through activation of the endoplasmic reticulum (ER) stress element (ERSE). To understand the mechanism of grp78 induction by ATF6 in cells subjected to ER calcium depletion stress mediated by thapsigargin (Tg) treatment, we discovered that ATF6 itself undergoes Tg stress-induced changes. In nonstressed cells, ATF6, which contains a putative short transmembrane domain, is primarily associated with the perinuclear region. Upon Tg stress, the ATF6 protein level dropped initially but quickly recovered with the additional appearance of a faster-migrating form. This new form of ATF6 was recovered as soluble nuclear protein by biochemical fractionation, correlating with enhanced nuclear localization of ATF6 as revealed by immunofluorescence. Optimal ATF6 stimulation requires at least two copies of the ERSE and the integrity of the tripartite structure of the ERSE. Of primary importance is a functional NF-Y complex and a high-affinity NF-Y binding site that confers selectivity among different ERSEs for ATF6 inducibility. In addition, we showed that YY1 interacts with ATF6 and in Tg-treated cells can enhance ATF6 activity. The ERSE stimulatory activity of ATF6 exhibits properties distinct from those of human Ire1p, an upstream regulator of the mammalian unfolded protein response. The requirement for a high-affinity NF-Y site for ATF6 but not human Ire1p activity suggests that they stimulate the ERSE through diverse pathways. PMID- 10866667 TI - Mechanism of transcriptional regulation by methyl-CpG binding protein MBD1. AB - MBD1 is a mammalian protein that binds symmetrically methylated CpG sequences and regulates gene expression in association with DNA methylation. This protein possesses a conserved sequence, named methyl-CpG binding domain (MBD), among a family of methyl-CpG binding proteins that mediate the biological consequences of the methylation. In addition, MBD1 has at least five isoforms due to alternative splicing events, resulting in the presence of CXXC1, CXXC2, and CXXC3 in MBD1 isoforms v1 (MBD1v1) and MBD1v2, and CXXC1 and CXXC2 in MBD1v3 and -v4. In the present study, we have investigated the significance of MBD, CXXC, and the C terminal transcriptional repression domain (TRD) in MBD1. A bacterially expressed MBD binds efficiently to densely methylated rather than to sparsely methylated DNAs. In both methylation-deficient Drosophila melanogaster SL2 cells and mammalian CHO-K1 cells, MBD1v1 represses transcription preferentially from both unmethylated and sparsely methylated promoters, while MBD1v3 inhibits densely methylated but not unmethylated promoter activities. The CXXC3 sequence in MBD1v1 is responsible for the ability to bind unmethylated promoter. Furthermore, we have constructed mutant-type MBD1s in which the functionally important residues Arg22, Arg30, Asp32, Tyr34, Arg44, Ser45, and Tyr52 are changed to alanine to investigate the correlation between the structure and function of the MBD in MBD1. Excepting those for Ser45 and Tyr52, none of the recombinant MBD mutants bound to the densely methylated or unmethylated DNAs, and green fluorescent protein-fused MBD1 mutants did not localize properly in the nucleus. All the MBD1v1 and -v3 mutants lost the activity of methylation-dependent gene repression. Based on these findings we have concluded that MBD1 acts as a transcriptional regulator depending on the density of methyl-CpG pairs through the cooperation of MBD, CXXC, and TRD sequences. PMID- 10866668 TI - Growth, adipose, brain, and skin alterations resulting from targeted disruption of the mouse peroxisome proliferator-activated receptor beta(delta). AB - To determine the physiological roles of peroxisome proliferator-activated receptor beta (PPARbeta), null mice were constructed by targeted disruption of the ligand binding domain of the murine PPARbeta gene. Homozygous PPARbeta-null term fetuses were smaller than controls, and this phenotype persisted postnatally. Gonadal adipose stores were smaller, and constitutive mRNA levels of CD36 were higher, in PPARbeta-null mice than in controls. In the brain, myelination of the corpus callosum was altered in PPARbeta-null mice. PPARbeta was not required for induction of mRNAs involved in epidermal differentiation induced by O-tetradecanoylphorbol-13-acetate (TPA). The hyperplastic response observed in the epidermis after TPA application was significantly greater in the PPARbeta-null mice than in controls. Inflammation induced by TPA in the skin was lower in wild-type mice fed sulindac than in similarly treated PPARbeta-null mice. These results are the first to provide in vivo evidence of significant roles for PPARbeta in development, myelination of the corpus callosum, lipid metabolism, and epidermal cell proliferation. PMID- 10866669 TI - Selection of a dominant negative retinoblastoma protein (RB) inhibiting satellite myoblast differentiation implies an indirect interaction between MyoD and RB. AB - Satellite myoblasts serve as stem cells in postnatal skeletal muscle, but the genes responsible for choosing between growth versus differentiation are largely undefined. We have used a novel genetic approach to identify genes encoding proteins whose dominant negative inhibition is capable of interrupting the in vitro differentiation of C2C12 murine satellite myoblasts. The screen is based on fusion of a library of cDNA fragments with the lysosomal protease cathepsin B (CB), such that the fusion protein intracellularly diverts interacting factors to the lysosome. Among other gene fragments selected in this screen, including those of known and novel sequence, is the retinoblastoma protein (RB) pocket domain. This unique dominant negative form of RB allows us to genetically determine if MyoD and RB associate in vivo. The dominant negative CB-RB fusion produces a cellular phenotype indistinguishable from recessive loss of function RB mutations. The fact that the dominant negative RB inhibits myogenic differentiation in the presence of nonlimiting concentrations of either RB or MyoD suggests that these two proteins do not directly interact. We further show that the dominant negative RB inhibits E2F1 but cannot inhibit a forced E2F1-RB dimer. Therefore, E2F1 is a potential mediator of the dominant negative inhibition of MyoD by CB-RB during satellite cell differentiation. We propose this approach to be generally suited to the investigation of gene function, even when little is known about the pathway being studied. PMID- 10866670 TI - Evidence for Gal3p's cytoplasmic location and Gal80p's dual cytoplasmic-nuclear location implicates new mechanisms for controlling Gal4p activity in Saccharomyces cerevisiae. AB - Genetics and in vitro studies have shown that the direct interaction between Gal3p and Gal80p plays a central role in galactose-dependent Gal4p-mediated GAL gene expression in the yeast Saccharomyces cerevisiae. Precisely how Gal3p-Gal80p interaction effects induction is not clear. It has been assumed that Gal3p interacts with Gal80p in the nucleus upon galactose addition to release Gal80p inhibition of Gal4p. Although Gal80p has been shown to possess nuclear localization signal (NLS) peptides, the subcellular distribution of neither Gal80p nor Gal3p was previously determined. Here we report that Gal3p is located in the cytoplasm and apparently excluded from the nucleus. We show that Gal80p is located in both the cytoplasm and the nucleus. Converting Gal80p into a nucleus localized protein (NLS-Gal80p) by exogenous NLS addition impairs GAL gene induction. The impaired induction can be partially suppressed by targeting Gal3p to the nucleus (NLS-Gal3p). We document a very rapid association between NLS Gal3p and Gal80p in vivo in response to galactose, illustrating that the nuclear import of Gal80p is very rapid and efficient. We also demonstrate that nucleus localized NLS-Gal80p can move out of the nucleus and shuttle between nuclei in yeast heterokaryons. These results are the first indication that the subcellular distribution dynamics of the Gal3 and Gal80 proteins play a role in regulating Gal4p-mediated GAL gene expression in vivo. PMID- 10866671 TI - Sequence requirements for trafficking of the CRAM transmembrane protein to the flagellar pocket of African trypanosomes. AB - CRAM is a cysteine-rich acidic transmembrane protein, highly expressed in the procyclic form of Trypanosoma brucei. Cell surface expression of CRAM is restricted to the flagellar pocket of trypanosomes, the only place where receptor mediated endocytosis takes place in the parasite. CRAM can function as a receptor and was hypothesized to be a lipoprotein receptor of trypanosomes. We study mechanisms involved in the presentation and routing of CRAM to the flagellar pocket of insect- and bloodstream-form trypanosomes. By deletional mutagenesis, we found that deleting up to four amino acids from the C terminus of CRAM did not affect the localization of CRAM at the flagellar pocket. Shortening the CRAM protein by 8 and 19 amino acids from the C terminus resulted in the distribution of the CRAM protein in the endoplasmic reticulum (ER) (the CRAM protein is no longer uniquely sequestered at the flagellar pocket). This result indicates that the truncation of the CRAM C terminus affected the transport efficiency of CRAM from the ER to the flagellar pocket. However, when CRAM was truncated between 29 and 40 amino acids from the C terminus, CRAM was not only distributed in the ER but also located to the flagellar pocket and spread to the cell surface and the flagellum. Replacing the CRAM transmembrane domain with the invariant surface glycoprotein 65-derived transmembrane region did not affect the flagellar pocket location of CRAM. These results indicate that the CRAM cytoplasmic extension may exhibit two functional domains: one domain near the C terminus is important for efficient export of CRAM from the ER, while the second domain is of importance for confining CRAM to the flagellar pocket membrane. PMID- 10866672 TI - The heat shock cognate protein hsc73 assembles with A(1) adenosine receptors to form functional modules in the cell membrane. AB - A(1) adenosine receptors (A(1)Rs) are G protein-coupled heptaspanning receptors that interact at the outer face of the plasma membrane with cell surface ecto adenosine deaminase (ecto-ADA). By affinity chromatography the heat shock cognate protein hsc73 was identified as a cytosolic component able to interact with the third intracellular loop of the receptor. As demonstrated by surface plasmon resonance, purified A(1)Rs interact specifically with hsc73 with a dissociation constant in the nanomolar range (0.5 +/- 0.1 nM). The interaction between hsc73 and A(1)R led to a marked reduction in the binding of the ligands and prevented activation of G proteins, as deduced from (35)S-labeled guanosine-5'-O-(3 thio)triphosphate binding assays. Interestingly this effect was stronger than that exerted by guanine nucleotide analogs, which uncouple receptors from G proteins, and was completely prevented by ADA. As assessed by immunoprecipitation a high percentage of A(1)Rs in cell lysates are coupled to hsc73. A relatively high level of colocalization between A(1)R and hsc73 was detected in DDT(1)MF-2 cells by means of confocal microscopy, and no similar results were obtained for other G protein-coupled receptors. Colocalization between hsc73 and A(1)R was detected in specific regions of rat cerebellum and in the body of cortical neurons but not in dendrites or synapses. Remarkably, agonist-induced receptor internalization leads to the endocytosis of A(1)Rs by two qualitatively different vesicle types, one in which A(1)R and hsc73 colocalize and another in which hsc73 is absent. These results open the interesting possibility that signaling via G protein-coupled receptors may be regulated by heat shock proteins. PMID- 10866673 TI - Hepatocyte nuclear factor 3beta (Foxa2) is dispensable for maintaining the differentiated state of the adult hepatocyte. AB - Liver-specific gene expression is controlled by a heterogeneous group of hepatocyte-enriched transcription factors. One of these, the winged helix transcription factor hepatocyte nuclear factor 3beta (HNF3beta or Foxa2) is essential for multiple stages of embryonic development. Recently, HNF3beta has been shown to be an important regulator of other hepatocyte-enriched transcription factors as well as the expression of liver-specific structural genes. We have addressed the role of HNF3beta in maintenance of the hepatocyte phenotype by inactivation of HNF3beta in the liver. Remarkably, adult mice lacking HNF3beta expression specifically in hepatocytes are viable, with histologically normal livers and normal liver function. Moreover, analysis of >8,000 mRNAs by array hybridization revealed that lack of HNF3beta affects the expression of only very few genes. Based on earlier work it appears that HNF3beta plays a critical role in early liver development; however, our studies demonstrate that HNF3beta is not required for maintenance of the adult hepatocyte or for normal liver function. This is the first example of such functional dichotomy for a tissue specification transcription factor. PMID- 10866675 TI - Identification of functionally important domains in the N-terminal region of telomerase reverse transcriptase. AB - Telomerase is a ribonucleoprotein reverse transcriptase responsible for the maintenance of one strand of telomere terminal repeats. The key protein subunit of the telomerase complex, known as TERT, possesses reverse transcriptase-like motifs that presumably mediate catalysis. These motifs are located in the C terminal region of the polypeptide. Hidden Markov model-based sequence analysis revealed in the N-terminal region of all TERTs the presence of four conserved motifs, named GQ, CP, QFP, and T. Point mutation analysis of conserved residues confirmed the functional importance of the GQ motif. In addition, the distinct phenotypes of the GQ mutants suggest that this motif may play at least two distinct functions in telomere maintenance. Deletion analysis indicates that even the most N-terminal nonconserved region of yeast TERT (N region) is required for telomerase function. This N region exhibits a nonspecific nucleic acid binding activity that probably reflects an important physiologic function. Expression studies of various portions of the yeast TERT in Escherichia coli suggest that the N region and the GQ motif together may constitute a stable domain. We propose that all TERTs may have a bipartite organization, with an N-GQ domain connected to the other motifs through a flexible linker. PMID- 10866674 TI - The docking protein HEF1 is an apoptotic mediator at focal adhesion sites. AB - HEF1 (human enhancer of filamentation 1) is a member of a docking protein family that includes p130(Cas) and Efs. Through assembly of multiple protein interactions at focal adhesion sites, these proteins activate signaling cascades in response to integrin receptor binding of the extracellular matrix. The HEF1 protein is cell cycle regulated, with full-length forms cleaved in mitosis at a caspase consensus site to generate an amino-terminal 55-kDa form that localizes to the mitotic spindle. The identification of a caspase cleavage site in HEF1 led us to investigate whether HEF1 belongs to a select group of caspase substrates cleaved in apoptosis to promote the morphological changes characteristic of programmed cell death. Significantly, inducing expression of HEF1 in MCF-7 or HeLa cells causes extensive apoptosis, as assessed by multiple criteria. Endogenous HEF1 is cleaved into 65- and 55-kDa fragments and a newly detected 28 kDa form in response to the induction of apoptosis, paralleling cleavage of poly(ADP-ribose) polymerase and focal adhesion kinase (FAK); the death-promoting activity of over-expressed HEF1 is associated with production of the 28-kDa form. While the generation of the cleaved HEF1 forms is caspase dependent, the accumulation of HEF1 forms is further regulated by the proteasome, as the proteasome inhibitors N-acetyl-L-leucinyl-L-leucinyl-L-norleucinyl and lactacystin enhance their stability. Finally, the induction of HEF1 expression also increases Jun N-terminal protein kinase (JNK) activation, and activated JNK colocalizes with HEF1, implicating this pathway in HEF1 action. Based on these results, we propose that dysregulation of HEF1 and its family members along with FAK may signal the destruction of focal adhesion sites and regulate the onset of apoptosis. PMID- 10866676 TI - Fatal bilateral chylothorax in mice lacking the integrin alpha9beta1. AB - Members of the integrin family of adhesion receptors mediate both cell-cell and cell-matrix interactions and have been shown to play vital roles in embryonic development, wound healing, metastasis, and other biological processes. The integrin alpha9beta1 is a receptor for the extracellular matrix proteins osteopontin and tenacsin C and the cell surface immunoglobulin vascular cell adhesion molecule-1. This receptor is widely expressed in smooth muscle, hepatocytes, and some epithelia. To examine the in vivo function of alpha9beta1, we have generated mice lacking expression of the alpha9 subunit. Mice homozygous for a null mutation in the alpha9 subunit gene appear normal at birth but develop respiratory failure and die between 6 and 12 days of age. The respiratory failure is caused by an accumulation of large volumes of pleural fluid which is rich in triglyceride, cholesterol, and lymphocytes. alpha9(-/-) mice also develop edema and lymphocytic infiltration in the chest wall that appears to originate around lymphatics. alpha9 protein is transiently expressed in the developing thoracic duct at embryonic day 14, but expression is rapidly lost during later stages of development. Our results suggest that the alpha9 integrin is required for the normal development of the lymphatic system, including the thoracic duct, and that alpha9 deficiency could be one cause of congenital chylothorax. PMID- 10866677 TI - Modulation of CRX transactivation activity by phosducin isoforms. AB - Phosducin (Phd) and Phd-like proteins (PhLPs) selectively bind guanine nucleotide protein (G protein) betagamma subunits (Gbetagamma), while Phd-like orphan proteins (PhLOPs) lack the major functional domain for the binding of Gbetagamma. A retina- and pineal gland-specific transcription factor, cone-rod homeobox (CRX), was identified by a yeast two-hybrid screen using PhLOP1 as the bait. Direct protein-protein interactions between Phd or PhLOP1 and CRX were demonstrated using a beta-galactosidase quantitative assay in the yeast two hybrid system and were confirmed by an in vitro binding assay and a glutathione S transferase (GST) pull-down assay. To determine if the interaction with Phd or PhLOP1 affected CRX transactivation, a 120-bp interphotoreceptor retinoid binding protein (IRBP) promoter-luciferase reporter construct containing a CRX consensus element (GATTAA) was cotransfected into either COS-7 or retinoblastoma Weri-Rb-1 cells with expression constructs for CRX and either Phd or PhLOP1. Phd and PhLOP1 inhibited the transcriptional activation activity of CRX by 50% during transient cotransfection in COS-7 cells and by 70% in Weri-Rb-1 cells and COS-7 cells stably transfected with CRX. Phd inhibited CRX transactivation in a dose dependent manner. Whereas Phd is a cytoplasmic phosphoprotein, coexpression of Phd with CRX results in Phd being localized both in the cytoplasm and nucleus. By contrast, PhLOP1 is found in the nucleus even without CRX coexpression. To address the physiological relevance of these potential protein interacting partners, we identified immunoreactive proteins for Phd and CRX in retinal cytosolic and nuclear fractions. Immunohistochemical analysis of bovine retinas reveals colocalization of Phd isoforms with CRX predominantly in the inner segment of cone cells, with additional costaining in the outer nuclear layer and the synaptic region. Our findings demonstrate that both Phd and PhLOP1 interact directly with CRX and that each diminishes the transactivation activity of CRX on the IRBP promoter. A domain that interacts with CRX is found in the carboxyl terminus of the Phd isoforms. Phd antibody-immunoreactive peptides are seen in light-adapted mouse retinal cytosolic and nuclear extracts. Neither Phd nor PhLOP1 affected CRX binding to its consensus DNA element in electrophoretic mobility shift assays. A model that illustrates separate functional roles for interactions between Phd and either SUG1 or CRX is proposed. The model suggests further a mechanism by which Phd isoforms could inhibit CRX transcriptional activation. PMID- 10866678 TI - c-Jun NH(2)-terminal kinase inhibits targeting of the protein phosphatase calcineurin to NFATc1. AB - The protein phosphatase calcineurin is a critical mediator of calcium signals during T-cell activation. One substrate of calcineurin is the transcription factor NFATc1, which is retained in the cytoplasm of quiescent cells. NFATc1 activation requires the translocation of the transcription factor into the nucleus, a process that is mediated by calcineurin. This interaction with calcineurin requires a targeting domain (PxIxIT motif) located in the NH(2) terminal region of NFATc1. Here we demonstrate that the calcineurin targeting domain of NFATc1 is phosphorylated and inactivated by the c-Jun NH(2)-terminal kinase (JNK). This disruption of calcineurin targeting inhibits the nuclear accumulation and transcription activity of NFATc1 and accounts for the observation that Jnk1(-/-) T cells exhibit greatly increased NFATc1-dependent nuclear responses. PMID- 10866679 TI - Polarized growth controls cell shape and bipolar bud site selection in Saccharomyces cerevisiae. AB - We examined the relationship between polarized growth and division site selection, two fundamental processes important for proper development of eukaryotes. Diploid Saccharomyces cerevisiae cells exhibit an ellipsoidal shape and a specific division pattern (a bipolar budding pattern). We found that the polarity genes SPA2, PEA2, BUD6, and BNI1 participate in a crucial step of bud morphogenesis, apical growth. Deleting these genes results in round cells and diminishes bud elongation in mutants that exhibit pronounced apical growth. Examination of distribution of the polarized secretion marker Sec4 demonstrates that spa2Delta, pea2Delta, bud6Delta, and bni1Delta mutants fail to concentrate Sec4 at the bud tip during apical growth and at the division site during repolarization just prior to cytokinesis. Moreover, cell surface expansion is not confined to the distal tip of the bud in these mutants. In addition, we found that the p21-activated kinase homologue Ste20 is also important for both apical growth and bipolar bud site selection. We further examined how the duration of polarized growth affects bipolar bud site selection by using mutations in cell cycle regulators that control the timing of growth phases. The grr1Delta mutation enhances apical growth by stabilizing G(1) cyclins and increases the distal-pole budding in diploids. Prolonging polarized growth phases by disrupting the G(2)/M cyclin gene CLB2 enhances the accuracy of bud site selection in wild-type, spa2Delta, and ste20Delta cells, whereas shortening the polarized growth phases by deleting SWE1 decreases the fidelity of bipolar budding. This study reports the identification of components required for apical growth and demonstrates the critical role of polarized growth in bipolar bud site selection. We propose that apical growth and repolarization at the site of cytokinesis are crucial for establishing spatial cues used by diploid yeast cells to position division planes. PMID- 10866680 TI - Delayed wound healing in keratin 6a knockout mice. AB - Keratin 6 (K6) expression in the epidermis has two components: constitutive expression in the innermost layer of the outer root sheath (ORS) of hair follicles and inducible expression in the interfollicular epidermis in response to stressful stimuli such as wounding. Mice express two K6 isoforms, MK6a and MK6b. To gain insight into the functional significance of these isoforms, we generated MK6a-deficient mice through mouse embryonic stem cell technology. Upon wounding, MK6a was induced in the outer ORS and the interfollicular epidermis including the basal cell layer of MK6a(+/+) mice, whereas MK6b induction in MK6a( /-) mice was restricted to the suprabasal layers of the epidermis. After superficial wounding of the epidermis by tape stripping, MK6a(-/-) mice showed a delay in reepithelialization from the hair follicle. However, the healing of full thickness skin wounds was not impaired in MK6a(-/-) animals. Migration and proliferation of MK6a(-/-) keratinocytes were not impaired in vitro. Furthermore, the migrating and the proliferating keratinocytes of full-thickness wounds in MK6a(-/-) animals expressed neither MK6a nor MK6b. These data indicate that MK6a does not play a major role in keratinocyte proliferation or migration but point to a role in the activation of follicular keratinocytes after wounding. This study represents the first report of a keratin null mutation that results in a wound healing defect. PMID- 10866681 TI - Abnormalities of the genitourinary tract in female mice lacking GATA5. AB - Members of the GATA family of transcription factors play important roles in cell fate specification, differentiation, and morphogenesis during mammalian development. GATA5, the only one of the six vertebrate GATA factor genes not yet inactivated in mice, is expressed in a pattern that overlaps with but is distinct from that of other GATA factor genes. During mouse embryogenesis, GATA5 is expressed first in the developing heart and subsequently in the lung, vasculature, and genitourinary system. To investigate the function of GATA5 in vivo, we created mice homozygous for a GATA5 null allele. Homozygous mutants were viable and fertile, but females exhibited pronounced genitourinary abnormalities that included vaginal and uterine defects and hypospadias. In contrast, the genitourinary system was unaffected in male GATA5 mutants. These results reveal a specific role of GATA5 in development of the female genitourinary system and suggest that other GATA factors may have functions overlapping those of GATA5 in other tissues. PMID- 10866682 TI - Functional compensation by Egr4 in Egr1-dependent luteinizing hormone regulation and Leydig cell steroidogenesis. AB - The Egr family of zinc finger transcription factors, whose members are encoded by Egr1 (NGFI-A), Egr2 (Krox20), Egr3, and Egr4 (NGFI-C) regulate critical genetic programs involved in cellular growth, differentiation, and function. Egr1 regulates luteinizing hormone beta subunit (LHbeta) gene expression in the pituitary gland. Due to decreased levels of LHbeta, female Egr1-deficient mice are anovulatory, have low levels of progesterone, and are infertile. By contrast, male mutant mice show no identifiable defects in spermatogenesis, testosterone synthesis, or fertility. Here, we have shown that serum LH levels in male Egr1 deficient mice are adequate for maintenance of Leydig cell steroidogenesis and fertility because of partial functional redundancy with the closely related transcription factor Egr4. Egr4-Egr1 double mutant male mice had low steady-state levels of serum LH, physiologically low serum levels of testosterone, and atrophy of androgen-dependent organs that were not present in either Egr1- or Egr4 deficient males. In double mutant male mice, atrophic androgen-dependent organs and Leydig cell steroidogenesis were fully restored by administration of exogenous testosterone or human chorionic gonadotropin (an LH receptor agonist), respectively. Moreover, a normal distribution of gonadotropin-releasing hormone containing neurons and normal innervation of the median eminence in the hypothalamus, as well as decreased levels of LH gene expression in Egr4-Egr1 relative to Egr1-deficient male mice, indicates a defect of LH regulation in pituitary gonadotropes. These results elucidate a novel level of redundancy between Egr4 and Egr1 in regulating LH production in male mice. PMID- 10866683 TI - TATA binding protein can stimulate core-directed transcription by yeast RNA polymerase I. AB - The TATA binding protein (TBP) interacts with two transcription factor complexes, upstream activating factor (UAF) and core factor (CF), to direct transcription by RNA polymerase I (polI) in the yeast Saccharomyces cerevisiae. Previous work indicates that one function of TBP is to serve as a bridge, enabling UAF to recruit and stabilize the binding of CF (23, 24). In this work we show that, in addition to aiding recruitment, TBP also directly aids CF function. Overexpression of TBP in strains with UAF components deleted will stimulate CF directed transcription nearly to wild-type levels in vivo. In vitro, increasing the concentration of TBP stimulates CF-directed transcription in the absence of either UAF or its DNA binding site. This dual function of TBP, serving as a critical member of a core promoter complex as well as a contact point for upstream activators, appears similar to the dual roles that TBP also plays in transcription by RNA polII. PMID- 10866684 TI - Disruption of Myc-tubulin interaction by hyperphosphorylation of c-Myc during mitosis or by constitutive hyperphosphorylation of mutant c-Myc in Burkitt's lymphoma. AB - Somatic mutations at Thr-58 of c-Myc have been detected in Burkitt's lymphoma (BL) tumors and have been shown to affect the transforming potential of the Myc oncoprotein. In addition, the N-terminal domain of c-Myc has been shown to interact with microtubules in vivo, and the binding of c-Myc to alpha-tubulin was localized to amino acids 48 to 135 within the c-Myc protein. We demonstrate that c-Myc proteins harboring a naturally occurring mutation at Thr-58 from BL cell lines have increased stability and are constitutively hyperphosphorylated, which disrupts the in vivo interaction of c-Myc with alpha-tubulin. In addition, we show that wild-type c-Myc-alpha-tubulin interactions are also disrupted during a transient mitosis-specific hyperphosphorylation of c-Myc, which resembles the constitutive hyperphosphorylation pattern of Thr-58 in BL cells. PMID- 10866685 TI - The B56alpha regulatory subunit of protein phosphatase 2A is a target for regulation by double-stranded RNA-dependent protein kinase PKR. AB - PKR is a cellular serine/threonine kinase that phosphorylates eukaryotic translation initiation factor 2alpha (eIF2alpha) to regulate protein synthesis. PKR also plays a role in the regulation of transcription, programmed cell death and the cell cycle, processes which likely involve other substrates. In a yeast two-hybrid screen, we isolated human protein phosphatase 2A (PP2A) regulatory subunit B56alpha as a PKR-interacting protein. The interaction between B56alpha and PKR was confirmed by in vitro binding assays as well as by in vivo coimmunoprecipitation, and this interaction is dependent on the catalytic activity of PKR. Moreover, recombinant B56alpha was efficiently phosphorylated by PKR in vitro and an isoelectric point shift in B56alpha was detected in extracts from cells induced with the PKR activator pIC. An in vitro dephosphorylation assay showed that when B56alpha was phosphorylated by PKR, the activity of PP2A trimeric holoenzyme was increased. A functional interaction between B56alpha and PKR was observed in cotransfection assays, where a B56alpha-mediated increase in luciferase expression was inhibited by cotransfection with wild-type PKR. This is likely due to a decreased level of eIF4E phosphorylation caused by an increase in PP2A activity following PKR phosphorylation of B56alpha. Taken together, our data indicate that PKR can modulate PP2A activity by phosphorylating B56alpha to regulate cellular activities. PMID- 10866686 TI - DNA length dependence of the single-strand annealing pathway and the role of Saccharomyces cerevisiae RAD59 in double-strand break repair. AB - A DNA double-strand break (DSB) created by the HO endonuclease in Saccharomyces cerevisiae will stimulate recombination between flanking repeats by the single strand annealing (SSA) pathway, producing a deletion. Previously the efficiency of SSA, using homologous sequences of different lengths, was measured in competition with that of a larger repeat further from the DSB, which ensured that nearly all cells would survive the DSB if the smaller region was not used (N. Sugawara and J. E. Haber, Mol. Cell. Biol. 12:563-575, 1992). Without competition, the efficiency with which homologous segments of 63 to 205 bp engaged in SSA was significantly increased. A sequence as small as 29 bp was used 0.2% of the time, and homology dependence was approximately linear up to 415 bp, at which size almost all cells survived. A mutant with a deletion of RAD59, a homologue of RAD52, was defective for SSA, especially when the homologous sequence length was short; however, even with 1.17-kb substrates, SSA was reduced fourfold. DSB-induced gene conversion also showed a partial dependence on Rad59p, again being greatest when the homologous-sequence length was short. We found that Rad59p plays a role in removing nonhomologous sequences from the ends of single stranded DNA when it invades a homologous DNA template, in a manner similar to that previously seen with srs2 mutants. Deltarad59 affected DSB-induced gene conversion differently from msh3 and msh2, which are also defective in removing nonhomologous ends in both DSB-induced gene conversion and SSA. A msh3 rad59 double mutant was more severely defective in SSA than either single mutant. PMID- 10866687 TI - Poly(A) polymerase phosphorylation is dependent on novel interactions with cyclins. AB - We have previously shown that poly(A) polymerase (PAP) is negatively regulated by cyclin B-cdc2 kinase hyperphosphorylation in the M phase of the cell cycle. Here we show that cyclin B(1) binds PAP directly, and we demonstrate further that this interaction is mediated by a stretch of amino acids in PAP with homology to the cyclin recognition motif (CRM), a sequence previously shown in several cell cycle regulators to target specifically G(1)-phase-type cyclins. We find that PAP interacts with not only G(1)- but also G(2)-type cyclins via the CRM and is a substrate for phosphorylation by both types of cyclin-cdk pairs. PAP's CRM shows novel, concentration-dependent effects when introduced as an 8-mer peptide into binding and kinase assays. While higher concentrations of PAP's CRM block PAP cyclin binding and phosphorylation, lower concentrations induce dramatic stimulation of both activities. Our data not only support the notion that PAP is directly regulated by cyclin-dependent kinases throughout the cell cycle but also introduce a novel type of CRM that functionally interacts with both G(1)- and G(2)-type cyclins in an unexpected way. PMID- 10866688 TI - The C terminus of the Saccharomyces cerevisiae alpha-factor receptor contributes to the formation of preactivation complexes with its cognate G protein. AB - Binding of the alpha-factor pheromone to its G-protein-coupled receptor (encoded by STE2) activates the mating pathway in MATa yeast cells. To investigate whether specific interactions between the receptor and the G protein occur prior to ligand binding, we analyzed dominant-negative mutant receptors that compete with wild-type receptors for G proteins, and we analyzed the ability of receptors to suppress the constitutive signaling activity of mutant Galpha subunits in an alpha-factor-independent manner. Although the amino acid substitution L236H in the third intracellular loop of the receptor impairs G-protein activation, this substitution had no influence on the ability of the dominant-negative receptors to sequester G proteins or on the ability of receptors to suppress the GPA1-A345T mutant Galpha subunit. In contrast, removal of the cytoplasmic C-terminal domain of the receptor eliminated both of these activities even though the C-terminal domain is unnecessary for G-protein activation. Moreover, the alpha-factor independent signaling activity of ste2-P258L mutant receptors was inhibited by the coexpression of wild-type receptors but not by coexpression of truncated receptors lacking the C-terminal domain. Deletion analysis suggested that the distal half of the C-terminal domain is critical for sequestration of G proteins. The C-terminal domain was also found to influence the affinity of the receptor for alpha-factor in cells lacking G proteins. These results suggest that the C terminal cytoplasmic domain of the alpha-factor receptor, in addition to its role in receptor downregulation, promotes the formation of receptor-G-protein preactivation complexes. PMID- 10866689 TI - A pentamer transcriptional complex including tal-1 and retinoblastoma protein downmodulates c-kit expression in normal erythroblasts. AB - Human proerythroblasts and early erythroblasts, generated in vitro by normal adult progenitors, contain a pentamer protein complex comprising the tal-1 transcription factor heterodimerized with the ubiquitous E2A protein and linked to Lmo2, Ldb1, and retinoblastoma protein (pRb). The pentamer can assemble on a consensus tal-1 binding site. In the pRb(-) SAOS-2 cell line transiently transfected with a reporter plasmid containing six tal-1 binding site, pRb enhances the transcriptional activity of tal-1-E12-Lmo2 and tal-1-E12-Lmo2-Ldb1 complexes but not that of a tal-1-E12 heterodimer. We explored the functional significance of the pentamer in erythropoiesis, specifically, its transcriptional effect on the c-kit receptor, a tal-1 target gene stimulating early hematopoietic proliferation downmodulated in erythroblasts. In TF1 cells, the pentamer decreased the activity of the reporter plasmid containing the c-kit proximal promoter with two inverted E box-2 type motifs. In SAOS-2 cells the pentamer negatively regulates (i) the activity of the reporter plasmid containing the proximal human c-kit promoter and (ii) endogenous c-kit expression. In both cases pRb significantly potentiates the inhibitory effect of the tal-1-E12-Lmo2-Ldb1 tetramer. These data indicate that this pentameric complex assembled in maturing erythroblasts plays an important regulatory role in c-kit downmodulation; hypothetically, the complex may regulate the expression of other critical erythroid genes. PMID- 10866690 TI - Peroxisome proliferator-activated receptor gamma target gene encoding a novel angiopoietin-related protein associated with adipose differentiation. AB - The nuclear receptor peroxisome proliferator-activated receptor gamma regulates adipose differentiation and systemic insulin signaling via ligand-dependent transcriptional activation of target genes. However, the identities of the biologically relevant target genes are largely unknown. Here we describe the isolation and characterization of a novel target gene induced by PPARgamma ligands, termed PGAR (for PPARgamma angiopoietin related), which encodes a novel member of the angiopoietin family of secreted proteins. The transcriptional induction of PGAR follows a rapid time course typical of immediate-early genes and occurs in the absence of protein synthesis. The expression of PGAR is predominantly localized to adipose tissues and placenta and is consistently elevated in genetic models of obesity. Hormone-dependent adipocyte differentiation coincides with a dramatic early induction of the PGAR transcript. Alterations in nutrition and leptin administration are found to modulate the PGAR expression in vivo. Taken together, these data suggest a possible role for PGAR in the regulation of systemic lipid metabolism or glucose homeostasis. PMID- 10866692 TI - Physiological and pathophysiological implications of upper airway reflexes in humans. AB - The upper airway is a vital part of the respiratory tract. Although the upper airway serves several functions, protection of the airway and preservation of airway patency are the most essential functions subserved by upper airway reflexes. Various types of nerve endings have been identified in and under the epithelium of the upper airway, and afferent nerve endings are the natural starting of all reflex activity. The upper airway reflexes consist of many different types of reflex responses such as sneezing, apnea, swallowing, laryngeal closure, coughing, expiration reflex, and negative pressure reflex. Although the activation of upper airway reflexes does not necessarily occur at one particular site of the respiratory tract, individual reflex response is usually considered to be highly specific for the particular respiratory site which has been affected. The upper airway reflexes are modified by many factors such as sleep, anesthesia, and background chemical ventilatory drive. Both depression and exaggeration of upper airway reflexes cause clinical problems. Depression of upper airway reflexes enhances the chance of pulmonary aspiration and compromises the maintenance of the airway, whereas exaggeration of airway reflexes such as laryngospasm and prolonged paroxysm of cough can be harmful and dangerous. In this review, various aspects of upper airway reflexes are discussed focusing on the functions of upper airway reflexes in humans and some pathophysiological problems related to clinical medicine. PMID- 10866691 TI - Akr1p and the type I casein kinases act prior to the ubiquitination step of yeast endocytosis: Akr1p is required for kinase localization to the plasma membrane. AB - Ubiquitination of the plasma membrane-localized yeast a-factor receptor (Ste3p) triggers a rapid, ligand-independent endocytosis leading to its vacuolar degradation. This report identifies two mutants that block uptake by blocking ubiquitination, these being mutant either for the ankyrin repeat protein Akr1p or for the redundant type I casein kinases Yck1p and Yck2p. While no obvious defect was seen for wild-type Ste3p phosphorylation in akr1 or yck mutant backgrounds, examination of the Delta320-413 Ste3p deletion mutant phosphorylation did reveal a clear defect in both mutants. The Delta320-413 deletion removes 18 Ser-Thr residues (possible YCK-independent phosphorylation sites) yet retains the 15 Ser Thr residues of the Ste3p PEST-like ubiquitination-endocytosis signal. Two other phenotypes link akr1 and yck mutants: both are defective in phosphorylation of wild-type alpha-factor receptor, and while both are defective for Ste3p constitutive internalization, both remain partially competent for the Ste3p ligand-dependent uptake mode. Yck1p-Yck2p may be the function responsible in phosphorylation of the PEST-like ubiquitination-endocytosis signal. Akr1p appears to function in localizing Yck1p-Yck2p to the plasma membrane, a localization that depends on prenylation of C-terminal dicysteinyl motifs. In akr1Delta cells, Yck2p is mislocalized, showing a diffuse cytoplasmic localization identical to that seen for a Yck2p mutant that lacks the C-terminal Cys-Cys, indicating a likely Akr1p requirement for the lipid modification of Yck2p, for prenylation, or possibly for palmitoylation. PMID- 10866693 TI - Neuronal mechanisms mediating the integration of respiratory responses to hypoxia. AB - The activation of peripheral chemoreceptors by hypoxia or electrical stimulation of the carotid sinus nerve elicited a hypoxic respiratory response consisting of both stimulatory and subsequent or simultaneous inhibitory components (hypoxic respiratory stimulation and depression). Both components have different time domains of responses (time-dependent response), providing an integrated respiratory response to hypoxia. This review has focused on the neuroanatomical and neurophysiological correlations responsible for these responses and their neuropharmacological mechanisms. Hypoxic respiratory depression is characterized by the initial activation of respiration followed by a progressive and gradual decline in ventilation during prolonged and/or severe hypoxic exposure (biphasic response). The responsible mechanisms for the depression are located within the central nervous system and may be dependent upon activity from peripheral chemoreceptor. Two underlying mechanisms contributing to the depression have been advocated. (1) Change in synaptic transmission: Within the neuronal network controlling the hypoxic respiratory response, hypoxia might induce the enhancement of inhibitory neurotransmission (modulation), disfacilitation of excitatory neruotransmission or both. (2) Change in the membrane property of respiratory neurons: Hypoxia might suppress the membrane excitability of respiratory neurons composing the hypoxic respiratory response via modulating ion channels, leading to hyperpolarization or depolarization blocking of the neurons. However, the quantitative aspects of Pao(2) (degree and duration of hypoxic exposure) to induce these changes and the susceptibility of both mechanisms to the Pao(2) level have not yet been clearly elucidated. PMID- 10866694 TI - Development of an experimental apparatus for investigating lymphatic pumping activity of murine mesentery in vivo. AB - The present study has been attempted to establish a modified intravital microscope system for investigating murine lymphatic pumping activity in vivo and evaluate whether or not there is rhythmic pumping activity of murine mesenteric lymphatic vessels in vivo. We designed and constructed a custom organ chamber with a semicircular channel (8 mm in radius, 5 mm in width, 3 mm in depth), being suitable for the superfusing of murine mesentery in vivo. A marked lymphatic pumping activity was observed in the mesenteries of DDY mice. The maximal and minimal diameter and frequency in the pumping activity were 60.9 +/- 1.0 microm, 53.7 +/- 1.8 microm and 12.8 min(-1) (n = 5), respectively. Both NE (norepinephrine, 10(-8)-10(-6) M) and TEA (tetraethylammonium, 1-10 mM) caused dose-dependent constriction of the mesenteric lymphatic vessels in the mice. These findings suggest that a modified intravital microscope system with a specially designed and constructed edge-monitoring device enables us to investigate in vivo lymphatic circulation in murine mesenteries. PMID- 10866695 TI - Effect of constant and intermittent vagal stimulation on the heart rate and heart rate variability in rabbits. AB - To examine whether the high-frequency (HF) component of heart rate variability (HRV) reflects fluctuation or tonic level of vagal outflow, we investigated the effects of vagal efferent nerve stimulation (VS) on the heart rate and HRV in anesthetized open-chest rabbit under artificial respiration at a rate of 52 breaths/min (0.86 Hz). A power spectral analysis was performed at baseline and during VS (stimuli at 2 ms, 1-10 V and 5-25 Hz). VS was applied using two different patterns. The first was constant VS; continuous stimulation at graded frequency or voltage to simulate changes in the level of vagal "tone." The second pattern was intermittent VS; stimulation at 0.5 Hz of on-off cycle to simulate fluctuations in vagal efferent activity. The power spectrum at baseline showed a single narrow component at 0.86 Hz, identical to respiration rate. Both the constant and intermittent VS prolonged RR interval. The amplitude of the component at 0.86 Hz remained unaffected by either the constant or intermittent VS, whereas the latter evoked a distinct narrow component at 0.5 Hz, reflecting the on-off cycle of intermittent VS. Our results suggest that the HF component of the power spectrum of HRV measures the magnitude of fluctuations of vagal input associated with respiratory modulation. PMID- 10866697 TI - The effects of temperature on the mechanical performance in fatigued single muscle fibers of the frog induced by twitch and tetanus. AB - Muscle fatigue induced by consecutive twitches or tetani was studied in single skeletal muscle fibers of the frog, Rana japonica. The fatigue by twitch appeared sooner after the start of stimulation at lower temperatures (2-5 degrees C) than at higher ones (15-20 degrees C), while the fatigue by tetanus appeared sooner at higher temperatures. When a twitch-fatigued fiber was bathed in a solution with caffeine (15 mM), the contracture force was much higher than the fatigued force, while in tetanus fatigue, the force by caffeine was not different from the fatigued force. The length-force relation in fatigued fibers was compared with that in pre-fatigue at low and high temperatures. It was noticed that the ascending limb of the length-force curve in fatigued fibers by twitch was lower than that in pre-fatigue at the low temperatures; namely, the fatigue by twitch was more marked in shorter muscle length, while no marked change in the length force relation was detected in the tetanus fatigue at the low and high temperatures. The maximum shortening velocity, measured by the slack test, decreased in both types of fatigue. These results suggest that the fatigue by twitch may be mainly due to the failure of activation of the contractile system, while in the fatigue by tetanus, the rate of the interaction between actin and myosin may be impaired due to the change in intracellular chemical environment. PMID- 10866696 TI - Histochemical responses of human soleus muscle fibers to long-term bedrest with or without countermeasures. AB - Effects of 2- or 4-month bedrest in -6 degrees head-down tilt position with or without countermeasures on the histochemical properties of fiber phenotype and cross-sectional area (CSA) were studied in human soleus. The CSAs in slow fibers decreased approximately 32% during 4-month bedrest. This reduction was normalized after 1-month recovery. Although the reduction of percent slow fibers was not significant statistically, the percent intermediate fibers was significantly elevated 4 months after bedrest. Such shift in fiber type was not normalized following 1-month recovery. Effects of wearing an anti-g Penguin suit which has a modest, but continuous resistance at the knee and ankle (Penguin-1) or with knee resistance without loading on the ankle (Penguin-2) for 10 consecutive hours daily were also investigated during approximately 2 months of bedrest. The subjects performed knee extension and flexion for the last 15 min of each hour while in a supine position in bed. Bedrest-induced fiber atrophy was prevented in the Penguin-1 group but not the Penguin-2 group. Transformation of fiber type was not prevented in either Penguin suit group. It is suggested that long-term bedrest causes an atrophy and a shift of fiber phenotype toward fast-twitch type in human soleus. Data also indicated that loading on the muscle is an effective countermeasure for prevention of fiber atrophy but not fiber-type transformation. PMID- 10866698 TI - Norepinephrine and epinephrine responses during orthostatic intolerance in healthy elderly men. AB - In young individuals, orthostatic intolerance is associated with marked increases in plasma epinephrine (EPI) concentrations and attenuated rises in plasma norepinephrine (NE) concentrations. This study investigated the cardiovascular, EPI and NE responses of healthy elderly males during orthostatic stress. Twelve men (68 +/- 1 yr) with a recent history of orthostatic hypotension and who exhibited orthostatic intolerance (HYPO) during 90 degrees head-up tilt (HUT) were compared with 12 men (69 +/- 1 yr) without a history of orthostatic hypotension and who remained normotensive (NORMO) throughout 90 degrees HUT. Beat by-beat recordings of heart rate (HR), mean (MAP), systolic (SBP), diastolic (DBP), and pulse (PP) pressures were made throughout 90 degrees HUT. Blood samples obtained during supine rest and 90 degrees HUT were analyzed for changes in EPI and NE concentrations, hematocrit, hemoglobin and plasma volume. Compared to supine rest, orthostatic intolerance was characterized by significant reductions (p < 0.0001) in MAP, SBP, DBP, and PP. The HR, MAP, SBP, DBP, and PP at the termination of 90 degrees HUT was significantly lower (p < 0.0001) for HYPO than NORMO. The 90 degrees HUT position resulted in significant increases (p < 0.01) in NE for both HYPO and NORMO, with the rise in NE significantly lower (p < 0.05) in HYPO. There were no differences between groups regarding EPI concentrations at the termination of 90 degrees HUT. These results suggest that the magnitude of arterial pressure (AP) reduction does not influence the EPI response during orthostasis in healthy elderly men. However, marked reductions in AP, leading to orthostatic intolerance, are associated with inadequate increases in NE in these individuals. PMID- 10866699 TI - Osmotic membrane stretch increases cytosolic Ca(2+) and inhibits bone resorption activity in rat osteoclasts. AB - Although the importance of mechanical stress on bone metabolism is well known, the intracellular mechanisms involved are not well understood. To evaluate the role of mechanical stress on osteoclastic function, we investigated the effects of membrane stretch induced by osmotic cell swelling on cytosolic Ca(2+) and bone resorption activity in freshly isolated rat osteoclasts. The intracellular Ca(2+) concentration ([Ca(2+)](i)) was measured by fura-2 microspectrofluorimetry. Exposure to hypotonic solution (211-151 mOsm) caused cell swelling and reversibly increased [Ca(2+)](i) in the osteoclasts. This [Ca(2+)](i) increase was abolished by the omission of extracellular Ca(2+), but was not affected by the depletion of intracellular Ca(2+) stores. Gd(3+) and La(3+) inhibited the swelling-induced [Ca(2+)](i) increase, while nifedipine and Bay K 8644 did not. Neither protein kinase A inhibitors (Rp-cAMP, H-89) nor protein kinase C inhibitors (staurosporine, chelerythrine) affected the [Ca(2+)](i) increase. Membrane depolarization was not essential for the [Ca(2+)](i) increase either. To assess the effects of membrane stretch on the bone resorption activity of osteoclasts, we investigated actin ring formation, the intracellular structure responsible for bone resorption in osteoclasts. Hypotonic stimulation acutely disrupted actin ring formation in an extracellular Ca(2+)-dependent manner, and this disruption was prevented by Gd(3+). Moreover, Ca(2+) ionophore (ionomycin) also induced disruption of the actin rings. These results indicate that mechanical stress inhibits osteoclastic bone resorption activity, possibly via the elevation of [Ca(2+)](i) through stretch-activated, non-selective cation channels. PMID- 10866700 TI - Effective arterial elastance of irregular beats during atrial fibrillation in canine left ventricle. AB - Effective arterial elastance (E(a)) was originally defined as the end-systolic pressure (ESP)/stroke volume (SV) ratio of the left ventricle (LV). E(a) combined with LV contractility (E(max)), E(a)/E(max), proved to be powerful in analyzing the ventriculo-arterial coupling of normal and failing hearts in regular beats. However, E(a) sensitively changes with LV E(max), preload, and afterload widely changing among irregular beats. This has discouraged the use of E(a) during arrhythmia. However, we hypothesized that E(a) could serve as the effective afterload (not always arterial) elastance against ventricular ejection under arrhythmia. We tested this hypothesis by analyzing beat-to-beat changes in E(a) of irregular beats during electrically induced atrial fibrillation (AF) in normal canine in situ hearts. We newly found that during AF in each heart: 1) E(a) changed widely among irregular beats and became markedly high in weak beats with small SVs; 2) E(a) and E(a)/E(max) distributed non-normally with large skewness but 1/E(a) distributed more normally; 3) 1/E(a) correlated closely with end diastolic volume, E(max) and preceding beat intervals; and 4) the reciprocal of mean 1/E(a) closely correlated with mean ESP/mean SV. These results support our hypothesis that E(a) can serve as the effective afterload elastance against ventricular ejection on a per-beat basis during AF. E(a)/E(max) can also quantify the ventriculo-afterload (not arterial) coupling on a per-beat basis. This study, however, warns that mean E(a) and mean E(a)/E(max) of irregular beats cannot necessarily represent their averages during AF. PMID- 10866701 TI - Ventilation- and carotid chemoreceptor discharge-response to hypoxia during induced hypothermia in halothane anesthetized rat. AB - It has been hypothesized that respiratory "gain" to hypoxic stimulus is not depressed in hypothermic animals though ventilation and that metabolic O(2) demand (Vo(2)) decreases with reduction in body temperature. The present study addressed this hypothesis by quantitative analysis of ventilatory and carotid chemoreceptor responsiveness to hypoxia during induced hypothermia in halothane anesthetized and spontaneously breathing rats. Rectal temperature was lowered from 37 degrees C (normothermia) to 30 and 25 degrees C by cooling body surface at comparable anesthetic depth without inducing shivering. Ventilation (V(E)), V(O2), PaO(2) and carotid chemoreceptor afferent discharges were measured during hyperoxic and hypoxic gas breathing. PaO(2) values at the same Fi(O2) (range 0. 35-0.08) decreased progressively as rectal temperature decreased. Both the V(E)/V(O2)- and chemoreceptor discharge-response curves shifted toward a lower PaO(2) range with a slight increase in the response slopes during hypothermia. The results indicated that the sensitivity of carotid chemoreceptor and ventilatory responses to hypoxia did not decrease at reduced body temperature. It is concluded that carotid chemoreceptor mediated regulation of ventilation is tightly coupled to changes in PaO(2 )range in halothane anesthetized rats during induced hypothermia. PMID- 10866702 TI - Four-parametric non-linear regression fit of isovolumic relaxation in isolated ejecting rat and guinea pig hearts. AB - Left ventricular isovolumic pressure fall is characterized by the time constant tau obtained by fitting the exponential p(t) = p(infinity) + (p(0) p(infinity))3exp(-t/tau) to pressure fall. It has been shown that tau, calculated from the first half of pressure fall, differs considerably from that found at late relaxation in normal and pathophysiological conditions. The present study aims at testing for such differences statistically and to quantify tau changes during relaxation. Two improvements of the common regression procedure are introduced for that purpose: the use of the four-parametric regression function, p(t) = p(infinity) + (p(0)-p(infinity))3exp[-t/(tau(0)+b(tau)t)], and an optimal data-dependent split of the isovolumic pressure fall interval. The residual regression errors of the methods are statistically compared in one-hundred isolated working rat and one-hundred guinea pig hearts, additionally including a logistic regression method. Regression error is significantly reduced by introducing that b(tau). b(tau) is negative in most cases, indicating accelerated relaxation during isovolumic pressure fall, but zero and positive b(tau) are occasionally seen. Optimal interval tripartition further improves the regression error in most cases. The statistically proved acceleration of the time constant during isovolumic relaxation justifies factor b(t) as a direct and continuous measure of differences between early and late relaxation. This difference between early and late isovolumic relaxation is probably caused by residually contracted myocardium at the beginning of pressure fall, and is therefore important to describe pathophysiological effects on relaxation phases. PMID- 10866703 TI - Modulation of evoked cerebral blood flow under excessive blood supply and hyperoxic conditions. AB - We measured the field potential and local cerebral blood flow (LCBF) using laser Doppler flowmetry in alpha-chloralose anesthetized rats during activation of the somatosensory cortex by electrical stimulation of the hind paw under independent administration of additional carbon dioxide and oxygen. The aim of this study was to test the hypothesis that the increase in LCBF during activation of the cortex (evoked LCBF) is not directed toward supplying oxygen for oxidative metabolism. Under the hypercapnic condition (PaCO(2) = 74. 9 +/- 14.3 mmHg), the baseline LCBF was about 46.5% higher than that under the normocapnic condition (PaCO(2) = 35.7 +/- 2.1 mmHg) (p < 0. 001), but after normalization for each baseline (divided by the prestimulus level), there was no significant difference in the peak value and the rise time of normalized evoked LCBF. On the other hand, the baseline level of LCBF under the hyperoxic condition (PaO(2) = 479.4 +/- 77.2 mmHg) was about 5.0% lower than that under the normoxic condition (PaO(2) = 105.5 +/- 7.8 mmHg) (p < 0.01), suggesting mild vasoconstriction under the condition of hyperoxia at rest. The peak value of normalized evoked LCBF under the hyperoxic condition was about 6.5% higher than that under the normoxic condition (p < 0.05). In addition, the rise time of evoked LCBF was earlier under the hyperoxic condition (0.37 +/- 0.16 s) than that under the normoxic condition (0.52 +/- 0.12 s) (p < 0.01). The field potential measured during stimulation under hypercapnic and hyperoxic conditions was not significantly different when compared with that under normal gas conditions. These results support our hypothesis and suggest that the excess oxygen is involved in the mechanism underlying the regulation of LCBF. PMID- 10866704 TI - Postischemic reperfusion in the eyes of young and aged rats. AB - The hemodynamic changes during postischemic reperfusion were investigated in the eyes of young (4 months) and aged (more than 18 months) rats using laser Doppler flowmetry, and histological changes in the retina were examined 6 h after the cessation of ischemia. During exposure to 80 mmHg of intraocular pressure, choroidal blood flow (ChBF) decreased to 40-50% of the baseline value. Marked hyperperfusion (186 +/- 9%) was observed 1 min after cessation of 30-min ischemia in young rats. The hyperperfusion was less (111 +/- 3%) after 120-min ischemia. Delayed hypoperfusion was not observed during 6 h of reperfusion after 120-min ischemia. In aged rats, the hyperperfusion after 30-min ischemia was less (130 +/ 17%) than that in young rats, and the ChBF decreased to 80% of the baseline value during 6 h of reperfusion after 120-min ischemia. Histological examination of the retina showed that exposure to 120-min ischemia caused microvacuolation in the inner and outer plexiform layers and vacuolar changes in the cytoplasms in the inner nuclear layer of both young and aged rats, suggesting edema formation in the retina. The thickness of the outer layers of the retina tended to increase after 120-min ischemia in young rats, whereas it decreased significantly in aged rats. These results suggest that 120-min ischemia with 40-50% of normal choroidal blood flow causes more severe damage than 30-min ischemia, and that the hemodynamic changes during reperfusion in aged rats are different from those in young rats. PMID- 10866705 TI - Diffuse noxious inhibitory controls in anti-nociception produced by acupuncture and moxibustion on trigeminal caudalis neurons in rats. AB - Numerous studies have demonstrated that acupuncture and moxibustion induce analgesic effects. This study examined whether diffuse noxious inhibitory controls (DNIC) participated in acupuncture and moxibustion induced-analgesia. Single unit extracellular recordings from neurons in the trigeminal nucleus caudalis of urethane-anesthetized Wistar rats were obtained with a glass micropipette. A total of 52 single units, including 36 wide dynamic range (WDR), 5 nociceptive specific (NS) and 11 low-threshold mechanoreceptive (LTM) units were examined. During noxious test stimulation (cutaneous pinch or electrical stimulation), acupuncture, moxibustion or pinch stimulation was applied as the conditioning stimulus to the remote area of the receptive fields. When the conditioning stimulation induced rapid suppression of noxious receptive field stimulation response, examination revealed that various areas of the entire body were affected and suppression increased in an intensity-dependent manner. These features resemble DNIC phenomena. The suppression was observed on both WDR and NS neurons but not on LTM neurons. Eight of 16 WDR neurons examined were inhibited by acupuncture, five of 14 by moxibustion, and seventeen of 21 by pinching stimulation. Of the NS neurons, one of 2 units examined was suppressed by acupuncture, one of 2 by moxibustion, and two of 3 by pinch stimulation. Pinch stimulation induced the most profound suppression followed by manual acupuncture. Moxibustion induced moderate suppression with a long induction time. These results suggest that DNIC may be involved in the analgesic mechanism of acupuncture and moxibustion. PMID- 10866706 TI - Does edema formation occur in the rabbit brain exposed to head-down tilt? AB - Earlier studies showed that exposure to microgravity caused cephalad fluid shift, increased capillary pressure in the head, and produced facial edema and nasal congestion. In the present study, edema formation in the brain was investigated in rabbits exposed to simulated microgravity, head-down tilt (HDT), by measuring water content and histological examinations. Water content in the brain tissues of rabbits exposed to 2 and 8 days of HDT did not increase significantly compared with that of control animals. Neither vital staining using Evans blue nor immunohistochemical examination demonstrated extravasation of plasma constituents in the brain tissues of the HDT rabbits. Although marked congestion was noted in the brain, hematoxylin and eosin staining did not show edematous changes, such as distension of the perivascular and pericellular spaces and vacuolar appearance, in the tissues obtained from HDT rabbits. Transmission electron microscopy revealed that tight junctions of the capillary endothelium were intact in the HDT rabbits. These results suggest that either HDT up to 8 days does not cause brain edema in rabbits or it induces only a slight brain edema which is hard to be demonstrated by measurement of water content or histological examinations. PMID- 10866707 TI - The effect of melatonin on liver superoxide dismutase activity, serum nitrate and thyroid hormone levels. AB - Melatonin is a main neurohormone of the pineal gland. The effects of melatonin on the level of serum thyroid-stimulating hormone (TSH), thyroxine (T(4)), triiodothyronine (T(3)), nitrate, melatonin and liver superoxide dismutase (SOD) activity were examined in rats. Melatonin was injected at the dose of 10 mg/kg for 7 days, 2 h before turning the lights off. Rats were decapitated at 10:00 a.m. and 02:00 a.m., which are the times of the lowest and highest serum melatonin levels, respectively. Blood and tissue samples were collected. Decreased TSH, T(3), T(4) and nitrate levels were determined in the melatonin injected and nighttime groups. Melatonin levels showed a diurnal rhythm. SOD activity increased in the melatonin-treated group. The results demonstrate that increased SOD activity, and reduced serum TSH, T(3), T(4) and nitrate levels correlated with the serum melatonin levels. PMID- 10866708 TI - Heart rate response during incremental exercise in master runners. AB - We analyzed the kinetics of heart rate (HR) response during incremental treadmill exercise in thirteen master runners (62 +/- 1 yr). The HR/running speed (HR/S) relationship showed the existence of a point of downward deflection (HR(d)) in only approximately 31% of the subjects. Resting echocardiographic evaluations showed similar heart dimensions in all of the subjects. In conclusion, HR does not seem to show a curvilinear response (downward deflection) in most aged athletes. PMID- 10866709 TI - Upregulation of P53 protein in rat heart subjected to a transient occlusion of the coronary artery followed by reperfusion. AB - The accumulation of p53 protein in cardiomyocyte nuclei was immunohistochemically demonstrated by the pronounced staining of p53 antibody in 4 h-reperfused ventricular tissue after a 45-min coronary occlusion. The occurrence of apoptosis in the reperfused ventricular tissue was evidenced by positive TUNEL staining and DNA laddering on agarose gels. PMID- 10866710 TI - Abnormal air-righting reflex in striatal rats. AB - To understand dynamic postural control of the higher brain, we compared air righting reflexes in various decerebrate rats. Post-operative 3-5 d thalamic and mesencephalic rats displayed almost similar righting movements as intact. However, in striatal animals, coordination of righting movements was disrupted. The higher brain without cortical control could interfere with the brainstem center of the air-righting reflex. PMID- 10866711 TI - Increase in O(2) delivery with hyperoxia does not increase O(2) uptake in tetanically contracting dog muscle. AB - We investigated the influence of hyperoxia on O(2) uptake in tetanically contracting canine gastrocnemius. Hyperoxia showed neither increase in O(2) uptake nor decrease in lactate release, irrespective of increased O(2) supply, venous Po(2) and vascular resistance, as compared to normoxia, suggesting that hyperoxia decreases O(2) diffusion conductance and/or effective O(2) supply probably due to arteriovenous O(2) diffusion shunt. PMID- 10866712 TI - Investigators, Anaesthesia and ethics. PMID- 10866713 TI - Effect of ephedrine on auditory-evoked potentials during light general anaesthesia. AB - Ephedrine, a sympathomimetic drug, may stimulate the central nervous system via its amphetamine-like effect under light general anaesthesia. We compared the effect of ephedrine on auditory-evoked potentials with that of etilefrine, a sympathomimetic drug that lacks an amphetamine-like effect, in patients undergoing abdominal surgery under general anaesthesia with 67% nitrous oxide in oxygen and epidural blockade. Ephedrine (0.08 mg x kg-1 intravenously) significantly decreased the latencies of Nb and P1 from 49.5 (4.2) [mean (SD)] and 63.9 (9.1) ms to 45.9 (4.2) and 59.0 (9.9) ms, respectively; whereas etilefrine (0.02 mg x kg-1 intravenously) caused no significant changes in these potentials. In addition, the latencies of Nb and P1 before drug administration were positively correlated with patient age. These findings suggest that ephedrine can cause excitation of the central nervous system during light general anaesthesia, and that auditory-evoked potentials may be a sensitive indicator of the depth of anaesthesia. PMID- 10866714 TI - The effect of low concentrations of halothane on the cerebrovascular circulation in young children. AB - To determine the effect of halothane on cerebral blood flow velocity measured by transcranial Doppler, 23 healthy young children were studied during surgery. Anaesthesia was induced with thiopental, fentanyl and vecuronium, and maintained with halothane in 70% nitrous oxide in oxygen. A continuous epidural anaesthesia with 0. 25% bupivacaine was performed. End-tidal carbon dioxide pressure, temperature, heart rate and systolic blood pressure were kept constant. Three minimal alveolar concentrations (MAC; 0.5, 1.0 and 1. 5) of halothane were administered in stepwise increases. The cerebral blood flow velocity increased significantly at 1.0 (p < 0. 01) and 1.5 MAC (p < 0.001) compared with the value at 0.5 MAC. No further change in cerebral blood flow velocity was seen between 1.0 and 1.5 MAC. These data show that maximal changes in cerebral blood flow velocity are obtained at 1.0 MAC and that further increases in halothane concentration do not modify the cerebral circulation. It is suggested that young children differ from adults in that the maximal effect of halothane occurs at lower concentrations. PMID- 10866716 TI - The effect of smoking on postoperative nausea and vomiting. AB - In an attempt to quantify the postoperative effects of smoking, 327 consecutive patients undergoing arthroscopic day case knee surgery were given a standard anaesthetic consisting of an intravenous induction with propofol and fentanyl followed by inhalational maintenance using isoflurane in an oxygen and nitrous oxide mixture. Pre-operatively, patients were asked inter alia to give details of social smoking habits. Postoperatively, patients were given standard analgesic and anti-emetic drugs. Prior to discharge patients were asked to give details of postoperative nausea and vomiting together with details of the severity of postoperative pain. There were 85 smokers and 242 nonsmokers. Of the 327 patients, a total of 42 (13%) complained of postoperative nausea and vomiting. Of the smokers, only 6% complained of postoperative nausea and vomiting in contrast to 15% of the nonsmokers (p < 0.05). It is postulated that enzyme induction is the most likely reason for this anti-emetic effect. Possible ways in which this clinically beneficial mechanism can be utilised to improve outcome after anaesthesia are discussed. PMID- 10866715 TI - Patient-controlled intranasal diamorphine for postoperative pain: an acceptability study. AB - A patient acceptability study was conducted using patient-controlled intranasal diamorphine. Patients undergoing nonemergency orthopaedic or gynaecological surgery self-administered intranasal diamorphine for 24 h postoperatively. Pain, pain relief, sedation, respiratory rate, nausea and vomiting were assessed regularly. After 24 h, patients and their attending nurses completed a questionnaire assessing satisfaction and practical aspects of the technique. Satisfaction was reported as good or complete by 69% of patients and 69% of nurses. Pain relief was assessed as better than expected by 45% of patients and better than normal by 50% of nurses. Seventy-nine per cent of patients would be pleased to use patient-controlled intranasal diamorphine again and 89% of nurses would be happy for their patients to use it again. Sedation was uncommon and mild and there were no episodes of significant respiratory depression. Fifty-three per cent of patients reported no nausea and 74% did not vomit at any stage. There were seven withdrawals, four due to problems with the device and three due to therapeutic problems. The nasal spray may need modification to improve reliability. However, we found patient-controlled intranasal analgesia an effective technique, which was well tolerated by patients and nurses and was without unpleasant side-effects. Further work to determine how it performs compared with intramuscular or intravenous analgesia is now needed. PMID- 10866717 TI - Induction characteristics with 3% and 8% sevoflurane in adults: an evaluation of the second stage of anaesthesia and its haemodynamic consequences. AB - The second stage of anaesthesia was examined during 3 and 8% sevoflurane induction to see if any shortening of its duration was at the expense of cardiovascular stability. Fourteen volunteers underwent consecutive, randomly ordered inductions. Pupil size, skin sympathetic activity, plasma catecholamines, blood pressure and heart rate were measured. Eight per cent sevoflurane produced significantly shorter times to loss of consciousness (mean 68 s (SD 18) vs. mean 150 s (SD38)) and durations of second stage (mean 58 s (SD 38) vs. mean 91 s (SD 46)). Blood pressure, heart rate and sympathetic nerve responses were the same in both groups. Compared with baseline, skin sympathetic activity was greatest during pre-oxygenation (not significant) and unaltered during second stage. Both groups showed significantly increased plasma norepinephrine and heart rate and decreased blood pressure from baseline. Eight per cent sevoflurane induction produced a shorter second stage than 3% with equal cardiovascular stability and the same sympathetic response. PMID- 10866718 TI - Prelude to pancuronium and vecuronium. AB - The discovery of the neuromuscular blocking activity of malouetine isolated from Malouetia bequaertiana Woodson by Quevauviller and Laine in 1960 stimulated interest in aminosteroids as potential neuromuscular blockers. Alauddin (a pharmacist) and Martin-Smith (a medicinal chemist) began research in this area, which was then taken up by pharmaceutical companies. Pure pancuronium was synthesised in 1964 by Savage and his co-workers, directed by Hewett. Pharmacologists headed by Buckett discovered that pancuronium was far more potent than d-tubocurarine and had minimal side-effects. Buckett quickly recognised the value of pancuronium, and his persistence resulted in successful clinical trials, followed by the commercial launch of pancuronium in 1968. Vecuronium was synthesised, tested and the studies published by Buckett, Hewett and Savage in 1973, but this was before anaesthetists called for a 'clean muscle relaxant' and, furthermore, it was unstable in aqueous solution. Hence, it was only promoted for clinical trials six years later. PMID- 10866719 TI - A semiquantitative analysis of the dynamics of a Goldman-type vaporiser. AB - When a turbulent flow of a carrier gas is passed over a liquid anaesthetic agent contained in a vaporiser of Goldman design, evaporation from the cooled surface leads rapidly to a succession of fluid instabilities which set in at characteristic critical conditions. An initially quiescent boundary layer at the surface is sequentially replaced by a thin layer of toroidal (Benard-Marangoni) convection cells which are driven by surface tension gradients. These are then augmented by Rayleigh-Benard convection driven by gravity, the whole process terminating in intermittent columnar plunging of cold fluid from a chaotic surface layer of pulsating thickness to the base of the liquid pool. Residual striations from these plunging columns persist throughout the bulk of the liquid so long as evaporation continues. The ultimate state is then one in which turbulence occurs throughout both liquid and vapour phases. In this paper, a semiquantitative analysis of the system dynamics is given with supportive experimental evidence where possible. PMID- 10866720 TI - The effect of a heat and moisture exchanger on gas flow in a Mapleson F breathing system during inhalational induction. AB - Heat and moisture exchangers (HMEs) humidify, warm and filter inspired gas, protecting patients and apparatus during anaesthesia. Their incorporation into paediatric anaesthetic breathing systems is recommended. We experienced delays in inhalational induction whilst using a Mapleson F breathing system with an HME. We have demonstrated that the HME significantly alters gas flow within the breathing system. Approximately half of the fresh gas flow is delivered to the patient, the remainder being wasted into the expiratory limb of the breathing system. We suggest that the HME should be removed from the Mapleson F breathing system until inhalational induction is complete, or that the reservoir bag is completely occluded until an effective seal is obtained with the mask. PMID- 10866721 TI - Anaphylaxis caused by neostigmine. AB - A patient developed anaphylaxis during anaesthesia, towards the end of surgery, 30 s after intravenous administration of neostigmine. Anaphylaxis to neostigmine was confirmed by demonstrating an elevated mast cell tryptase and a strongly/positive skin prick test, showing the presence of drug-specific IgE (skin prick tests to neostigmine were negative in normal subjects). This is a rare cause of anaphylaxis during anaesthesia. PMID- 10866722 TI - Influence of an anaesthetist on nurse-led, computer-based, pre-operative assessment. AB - Trained nurses using a rule-based computer program can successfully carry out pre anaesthesia screening. All medical problems and abnormal laboratory results need to be reviewed by an experienced anaesthetist. Following the introduction of this system, there was a reduction in the frequency of cancellations of patients from elective orthopaedic operating lists from 4.8% to 1.8%, a difference that was statistically significant (p = 0.03, CI = [0.6%, 5.5%]). To minimise cancellations from booked operating lists, a booked admissions policy is essential, so that the anaesthetist who will eventually be responsible for patients with medical problems can be identified. Cancellations cannot be avoided completely because some abnormal conditions arise or deteriorate after completion of the screening process. The anaesthetist responsible for the patient's anaesthetic may have different views of the risks involved from those of the anaesthetist undertaking the screening process. PMID- 10866723 TI - A survey of paediatric dental anaesthesia in Scotland. AB - A postal survey of NHS hospital-based anaesthetists providing out-patient anaesthesia for dental procedures in children under 10 years of age was conducted in February 1999. Information was sought about quality of care and common practice in Scotland. The experience of the anaesthetists involved in such work was substantial, but the monitoring used did not meet current standards, with only 16% of respondents indicating use of a full range of standard devices. Separate recovery facilities were available to 99%, and all had access to a defibrillator, but the qualifications of dedicated assistant and recovery staff were lacking in 14 and 30%, respectively. Intravenous access was not obtained routinely after inhalational induction of anaesthesia by up to 71% (49%, never; 22%, sometimes). Systemic analgesia or local anaesthesia was used by 88%. Discharge times ranged from 10 min to 6 h. PMID- 10866724 TI - Cremasteric reflex test as an objective indicator of spinal anaesthesia. AB - We studied 100 men who were scheduled for urological surgery (Group 1) and another 50 men for orthopaedic surgery (Group 2). We attempted to anaesthetise both sides of the lower body in Group 1 and to anaesthetise one leg in Group 2 by injecting 0.3% hyperbaric dibucaine intrathecally. The presence or absence of the cremasteric reflex and loss of sensation to pinprick higher than the first lumbar dermatome were examined by two researchers who were blind to each other's results. In Group 1, both the reflex and the pinprick sensation were always absent bilaterally 5 min after intrathecal injection. In Group 2, in 23 of 50 patients the reflex had become absent bilaterally; in all these patients, bilateral sensory loss was detected. In the remaining 27 patients, both the reflex and the pinprick sensation were absent on the operation side, whereas both were present on the nonoperation side. Sensitivity, specificity and positive or negative predictive value for the cremasteric reflex were all 100%. Disappearance of the cremasteric reflex is a simple objective indicator of spinal anaesthesia at the first lumbar dermatome. This test may be useful in patients who cannot give reliable answers to conventional tests, such as the pinprick test. PMID- 10866725 TI - Anaesthesia for organ donation in the brainstem dead. PMID- 10866726 TI - Chemoprophylaxis for meningococcal disease in healthcare workers. PMID- 10866727 TI - Incidence of blood-borne infectious micro-organisms: would you still not wear gloves? PMID- 10866728 TI - Alzheimer's disease and anaesthesia. PMID- 10866729 TI - Forgotten already? PMID- 10866731 TI - The anaesthetic team. PMID- 10866732 TI - Postoperative nausea and vomiting following 8% sevoflurane anaesthesia. PMID- 10866733 TI - Unexpected difficult intubation caused by subglottic ring. PMID- 10866734 TI - Airway management device (AMD) for airway control in percutaneous dilatational tracheostomy. PMID- 10866735 TI - Complications following the use of the Combitube, tracheal tube and laryngeal mask airway. PMID- 10866737 TI - Nasendoscopy: a useful addition to the anaesthetist's armamentarium. PMID- 10866738 TI - Upper airway obstruction. PMID- 10866739 TI - The 'correct' application of cricoid pressure. PMID- 10866740 TI - Tranexamic acid applied topically to achieve haemostasis. PMID- 10866741 TI - Postoperative delirium mimicking epilepsy. PMID- 10866742 TI - An unusual case of postoperative facial swelling. PMID- 10866743 TI - Ambient pressure oxygenation via the nonventilated lung during video-assisted thoracoscopy. PMID- 10866744 TI - The effect of pre-emptive acupuncture treatment on analgesic requirements after day-case knee arthroscopy. PMID- 10866746 TI - Electroencephalographic arousal response during tracheal intubation and laryngeal mask airway insertion after induction of anaesthesia with propofol. PMID- 10866748 TI - A leaking Biovalve intravenous catheter. PMID- 10866750 TI - Leaking vaporiser. PMID- 10866752 TI - A 'groovy' fibrescope modification. PMID- 10866753 TI - Reinforced laryngeal mask airway severed by biting. PMID- 10866754 TI - Ambiguous drug labelling on ampoules containing local anaesthetic solution. PMID- 10866756 TI - Paediatric intensive care transfers. PMID- 10866757 TI - Pre-operative fasting in the older child. PMID- 10866758 TI - The stress response in children during laparoscopic abdominal surgery. PMID- 10866760 TI - A simple method of sub-Tenon anaesthesia delivery. PMID- 10866761 TI - Epidural insertion under general anaesthesia. PMID- 10866762 TI - Effectiveness of blood transfusion in Malawi. PMID- 10866763 TI - Blood transfusion for Caesarean section. PMID- 10866764 TI - Identification of epidural space using air with normal saline. PMID- 10866765 TI - A method of administering topical anaesthesia for flexible airway endoscopy. PMID- 10866766 TI - Even simpler remifentanil infusions. PMID- 10866767 TI - Lessons from history. PMID- 10866768 TI - Facial hair, sex and face-masks. PMID- 10866769 TI - British syringe label 'standards' are an accident waiting to happen. PMID- 10866770 TI - Paradoxical emboli in patent foramen ovale. PMID- 10866771 TI - How to skin a bearded cat. PMID- 10866791 TI - Modulation of molecular mechanisms involved in protein synthesis machinery as a new tool for the control of cell proliferation. AB - In the past years, the attention of scientists has focused mainly on the study of the genetic information and alterations that regulate eukaryotic cell proliferation and that lead to neoplastic transformation. All therapeutic strategies against cancer are, to date, directed at DNA either with cytotoxic drugs or gene therapy. Little or no interest has been aroused by protein synthesis mechanisms. However, an increasing body of data is emerging about the involvement of translational processes and factors in control of cell proliferation, indicating that protein synthesis can be an additional target for anticancer strategies. In this paper we review the novel insights on the biochemical and molecular events leading to protein biosynthesis and we describe their involvement in cell proliferation and tumorigenesis. A possible mechanistic explanation is given by the interactions that occur between protein synthesis machinery and the proliferative signal transduction pathways and that are therefore suitable targets for indirect modulation of protein synthesis. We briefly describe the molecular tools used to block protein synthesis and the attempts made at increasing their efficacy. Finally, we propose a new multimodal strategy against cancer based on the simultaneous intervention on protein synthesis and signal transduction. PMID- 10866792 TI - The effect of tryptophanyl substitution on folding and structure of myoglobin. AB - Mammalian myoglobins contain two tryptophanyl residues at the invariant positions 7 (A-5) and 14 (A-12) in the N-terminal region (A helix) of the protein molecule. The crucial role of tryptophanyl residues has been investigated by site-directed mutagenesis and molecular dynamics simulation. The apomyoglobin mutants with a double W-->F substitution were found to be not correctly folded and therefore not expressed as holoprotein. The introduction of a tyrosyl residue at position 7, that is, W7YW14F, resulted in the expression of a correctly folded myoglobin. Not correctly folded apomyoglobins were found with the following mutants: W7FW14Y, W7EW14F, W7FW14E, W7KW14F, W7FW14K. Moreover, in all these cases, very low levels of expression were observed. The acid-induced denaturation curves of wild-type and folded mutant W7YW14F, obtained following the fluorescence variation of the extrinsic fluorophore 1-anilino-8-naphthalenesulfonate, revealed that the stability of the native state of mutant apoprotein is decreased, thus indicating that the replacement W-->Y in position 7 is able to restore a correct folding but not the same stability. Molecular dynamics simulation indicated that both tryptophans are involved in forming favorable, specific tertiary interactions in the native apomyoglobin structure. The lack of some of these interactions caused by tryptophanyl replacement affects the overall protein structure and may provide an explanation for the observed stability decrease. In the case of the double W- >F substitution, the simulated structure shows conclusively the domain formed by helices A, G and H to be not correctly folded. This effect is attenuated if at least one of the two residues is conserved or a tyrosyl residue replaces W7. PMID- 10866793 TI - Liposome-mediated cytosolic delivery of macromolecules and its possible use in vaccine development. AB - In the majority of bacterial and viral infections the generation of cytotoxic T cells is of particular interest because such pathogens are able to escape the host defence mechanisms by surviving intracellularly within the phagocytic cells. To generate a CD8+ T lymphocyte response against exogenous antigens, the prerequisite is their delivery into the cytosol followed by processing and presentation along with class I major histocompatibility complex (MHC-I) molecules. In the present study we describe the method of liposome-based delivery of antigens and other macromolecules into the cytosol of target cells. To develop safe and effective methods for generating CD8+ T lymphocytes, we exploited the fusogenic character of lipids derived from lower organisms, that is baker's yeast (Saccharomyces cerevisiae). The degree of fusion with model membrane systems using yeast lipid liposomes varied from 40-70%, as opposed to 1-8% observed with egg PtdCho liposomes, depending on the assay system used. The fusion of yeast lipid liposomes with macrophages resulted in effective delivery of the entrapped solutes into the cytoplasmic compartment. This was further supported by the inhibition of cellular protein synthesis in J774 A1 cells by ricin A, encapsulated in the yeast lipid liposomes. Interestingly, the model antigen ovalbumin, when entrapped in the yeast lipid liposomes, successfully elicited antigen reactive CD8+ T cell responses. It may be concluded that the liposomes made of lipids derived from S. cerevisiae can spontaneously fuse with macrophages, delivering a significant portion of their contents into the cytoplasmic compartment of the cells. PMID- 10866794 TI - Thermal unfolding and refolding of beta-lactoglobulin. An intrinsic andextrinsic fluorescence study. AB - The conformational features of beta-lactoglobulin, refolded by cooling from a thermally perturbed state, has been characterized by intrinsic and extrinsic fluorescence measurements on the protein. It is found that even at 85-90 degrees C, beta-lactoglobulin does not completely lose its folded structure. The unfolding and refolding of beta-lactoglobulin as observed through intrinsic tryptophan fluorescence is nearly reversible because the native beta lactoglobulin and its refolded form, following heating and cooling, show nearly identical tryptophan fluorescence properties. However, the fluorescence properties of an extrinsic probe 1-anilino 8-naphthalene sulfonic acid (ANS) for the native and refolded forms are quite different from each other. Significant increase in fluorescence intensity and blue shifts in emission maxima of ANS bound to refolded beta-lactoglobulin is observed compared to that of the native form. Our results indicate that beta-lactoglobulin, refolded after heating to above 70 degrees C, has deep hydrophobic pockets which can be accessed by ANS. These pockets are either nonexistent or inaccessible to ANS in native beta lactoglobulin. The opening of the central cavity collapses at pH close to the isoelectric pH of the protein. This indicates that electrostatic repulsion is necessary to keep this access open. PMID- 10866795 TI - NMR investigations of protein-carbohydrate interactions binding studies and refined three-dimensional solution structure of the complex between the B domain of wheat germ agglutinin and N,N', N"-triacetylchitotriose. AB - The specific interaction of the isolated B domain of wheat germ agglutinin (WGA B) with N,N',N"-triacetylchitotriose has been analyzed by 1H-NMR spectroscopy. The association constants for the binding of WGA-B to this trisaccharide have been determined from both 1H-NMR titration experiments and microcalorimetry methods. Entropy and enthalpy of binding have been obtained. The driving force for the binding process is provided by a negative DeltaH which is partially compensated by negative DeltaS. These negative signs indicate that hydrogen bonding and van der Waals forces are the major interactions stabilizing the complex. NOESY NMR experiments in water solution provided 327 protein proton proton distance constraints. All the experimental constraints were used in a refinement protocol including restrained molecular dynamics in order to determine the refined solution conformation of this protein/carbohydrate complex. With regard to the NMR structure of the free protein, no important changes in the protein NOEs were observed, indicating that carbohydrate-induced conformational changes are small. The average backbone rmsd of the 35 refined structures was 1.05 A, while the heavy atom rmsd was 2.10 A. Focusing on the bound ligand, two different orientations of the trisaccharide within WGA-B binding site are possible. It can be deduced that both hydrogen bonds and van der Waals contacts confer stability to both complexes. A comparison of the three-dimensional structure of WGA-B in solution to that reported in the solid state and to those deduced for hevein and pseudohevein in solution has also been performed. PMID- 10866796 TI - Guanidine hydrochloride induced reversible dissociation and denaturation of duck delta2-crystallin. AB - The tetrameric delta2-crystallin from duck lens exhibits a reversible dissociation-denaturation process in solutions containing guanidine hydrochloride (GdnHCl). Sigmoidal or biphasic curves for the dissociation/denaturation processes, obtained using different methods of structural analysis, as a function of GdnHCl concentration were not coincidental with each other. delta2-crystallin in 0.91 M GdnHCl existed primarily as a monomer, which had no endogenous argininosuccinate lyase activity. After dilution of the GdnHCl-treated protein, the monomers reassociated into tetramers with concomitant recovery of enzyme activity. The sigmoidal recovery of enzyme activity demonstrates a cooperative hysteretic reactivation process. When the concentration of GdnHCl was higher than 1.2 M, various partially unfolded soluble forms of delta2-crystallin were produced from the dissociated monomers as shown by size-exclusion chromatography. The formation of a partially unfolded intermediate during the dissociation denaturation process is proposed. PMID- 10866797 TI - Structure and expression of mGDF, a new member of the transforming growth factor beta superfamily in the bivalve mollusc Crassostrea gigas. AB - To gain insight into the evolution of the structure and functions of transforming growth factor (TGF)-beta superfamily members, a cDNA encoding a new member from the bivalve mollusc Crassostrea gigas named mGDF (molluscan growth and differentiation factor) was identified by PCR using degenerate primers. The mGDF precursor exhibits characteristic features of the TGF-beta superfamily and shows highest homology with human BMP2 and Drosophila DPP. Conversely, the mgdf gene displays a distinct pattern of expression during development. Indeed mgdf transcripts were not detected early in development but increased markedly before metamorphosis. These findings raise the possibility that mGDF could play a central role in the biological processes that allow larvae to become competent to metamorphose. PMID- 10866798 TI - Zymogen activation in the streptokinase-plasminogen complex. Ile1 is required for the formation of a functional active site. AB - Plasminogen (Plgn) is usually activated by proteolysis of the Arg561-Val562 bond. The amino group of Val562 forms a salt-bridge with Asp740, which triggers a conformational change producing the active protease plasmin (Pm). In contrast, streptokinase (SK) binds to Plgn to produce an initial inactive complex (SK.Plgn) which subsequently rearranges to an active complex (SK.Plgn*) although the Arg561 Val562 bond remains intact. Therefore another residue must substitute for the amino group of Val562 and provide a counterion for Asp740 in this active complex. Two candidates for this counterion have been suggested: Ile1 of streptokinase and Lys698 of Plgn. We have investigated the reaction of SK mutants and variants of the protease domain of microplasminogen (muPlgn) in order to determine if either of these residues is the counterion. The mutation of Ile1 of SK decreases the activity of SK.Plgn* by 100-fold (Ile1Val) to >/= 104-fold (Ile1-->Ala, Gly, Trp or Lys). None of these mutations perturb the binding affinity of SK, which suggests that Ile1 is not required for formation of SK.Plgn but is necessary for SK.Plgn*. The substitution of Lys698 of muPlgn decreases the activity of SK.Plgn* by only 10-60-fold. In contrast with the Ile1 substitutions, the Lys698 mutations also decreased the dissociation constant of the SK complex by 15-50-fold. These observations suggest that Lys698 is involved in formation of the initial SK.Plgn complex. These results support the hypothesis that Ile1 provides the counterion for Asp740. PMID- 10866799 TI - A trans-acting factor, isolated by the three-hybrid system, that influences alternative splicing of the amyloid precursor protein minigene. AB - Two clones were isolated in a three-hybrid screen of a rat fetal brain P5 cDNA library with an intronic splicing enhancer of the amyloid precursor protein (APP) gene as RNA bait. These clones represent the rat homologues of the previously described genes CUG-binding protein (CUG-BP) and Siah-binding protein (Siah-BP). Both interact in a sequence-specific manner with the RNA bait used for library screening as well as with the CUG repeat. In contrast, no interactions were observed in the three-hybrid assay with other baits tested. In two-hybrid assays, Siah-BP interacts with U2AF65 as well as with itself. EWS, an RGG-type RNA binding protein associated with Ewing sarcoma, was identified as an interacting partner for the CUG-BP homologue in a two-hybrid assay for protein-protein interactions performed with various factors involved in RNA metabolism. Splicing assays performed by RT-PCR from cells cotransfected with certain cDNAs and an APP minigene, used as a reporter, indicate exclusion of exon 8 if the CUG-BP homologue is present. We conclude that clone AF169013 and its counterpart in human CUG-BP could be the trans-acting factors that interact with the splicing enhancer downstream of exon 8, and in this way influence alternative splicing of the APP minigene. PMID- 10866800 TI - Proteome analysis reveals ubiquitin-conjugating enzymes to be a new family of interferon-alpha-regulated genes. AB - Interferon (IFN)-alpha is a cytokine with antiviral, antiproliferative, and immunomodulatory properties, the functions of which are mediated via IFN-induced protein products. We used metabolic labeling and two-dimensional gel electrophoresis followed by MS and database searches to identify potentially new IFN-alpha-induced proteins in human T cells. By this analysis, we showed that IFN alpha induces the expression of ubiquitin cross-reactive protein (ISG15) and two ubiquitin-conjugating enzymes, UbcH5 and UbcH8. Northern-blot analysis showed that IFN-alpha rapidly enhances mRNA expression of UbcH5, UbcH6 and UbcH8 in T cells. In addition, these genes were induced in macrophages in response to IFN alpha or IFN-gamma stimulation or influenza A or Sendai virus infections. Similarly, IFNs enhanced UbcH8 mRNA expression in A549 lung epithelial cells, HepG2 hepatoma cells, and NK-92 cells. Cycloheximide, a protein synthesis inhibitor, did not block IFN-induced upregulation of UbcH8 mRNA expression, suggesting that UbcH8 is the primary target gene for IFN-alpha and IFN-gamma. Ubiquitin conjugation is a rate-limiting step in antigen presentation and therefore the upregulation of UbcHs by IFNs may contribute to the enhanced antigen presentation by macrophages. Our results show that proteome analysis of cells is a suitable method for identifying previously unrecognized cytokine inducible genes. PMID- 10866801 TI - H19 sense and antisense transgenes modify insulin-like growth factor-II mRNA levels. AB - The oppositely-imprinted genes insulin-like growth factor-II (IGF2) and H19, a putative tumor suppressor, often show coordinate, reciprocal regulation and are believed to play a role in carcinogenesis. To explore the possible interactions between these genes, we stably transfected diHepG2 cells with a plasmid containing either the sense or the antisense H19 cDNA sequences and verified their expression by Northern analysis and by RNase protection analysis. Levels of H19, IGF2 and gamma-actin mRNA were quantified by competitive RT-PCR analysis. Although H19 sense transgene overexpression (n = 24 clones) did not decrease the low, basal levels of IGF2 mRNA compared to control cells, levels of IGF2 mRNA were positively correlated with the levels of H19 antisense mRNA (P < 0.0001, n = 40 clones). Furthermore, the increase in IGF2 mRNA level was accompanied by an elevation of IGF-II peptide in conditioned media. To see if H19 mRNA had a specific effect on transcription, we also performed transient transfections with reporter gene constructs containing IGF2 promoter 3 in the presence of sense or antisense H19 cDNA sequences under control of a cytomegalovirus promoter. We show a lower reporter gene activity from reporter gene constructs in the presence of sense H19 cDNA than from those with antisense or neomycin. Our results suggest that H19 participates in the repression of IGF2, at least in part through effects on IGF2 transcription, an effect which may contribute to its action as a tumor suppressor. PMID- 10866802 TI - Evidence that phosphate specific transporter is amplified in a fluoroquinolone resistant Mycobacterium smegmatis. AB - We reported in an earlier study that active efflux of drug has a predominant role in conferring resistance in a laboratory-generated ciprofloxacin-resistant mutant of Mycobacterium smegmatis. This mutant exhibited mRNA level overexpression, as well as chromosomal amplification, of the gene pstB, encoding the putative ATPase subunit of phosphate specific transport (Pst) system. We demonstrate here that this mutant shows enhanced phosphate uptake and that inactivation of pstB in the parental strain results in loss of high affinity phosphate uptake and hypersensitivity to fluoroquinolones. These findings suggest a novel role of the Pst system in active efflux, in addition to its involvement in phosphate transport. PMID- 10866803 TI - Expanding baculovirus surface display. Modification of the native coat protein gp64 of Autographa californica NPV. AB - To create a tool for eukaryotic surface display, this approach is aimed at demonstrating a direct modification of the native envelope protein gp64 of Autographa californica NPV without disturbing viral infectivity. Short affinity tag peptides, the biotin mimic streptagII, and the gp41 amino-acid motif ELDKWA of HIV-1, specific for the human monoclonal antibody 2F5, were engineered into the baculovirus major coat protein gp64 and presented on the viral surface. Two different streptag peptides were inserted at the naturally occurring NotI site at amino-acid 278 of gp64. Additionally, the ten-amino-acid peptide GG-ELDKWA-GG, containing the epitope of mAb 2F5, was introduced into gp64 envelope protein at the same position. In all cases we were able to propagate viable virus-achieving infectious titers in the range of wild-type AcMNPV. Streptag and ELDKWA-epitope surface localization on purified virus particles was demonstrated by flow cytometry and Western blot analysis. We could also show selective retention of mutant viruses by specific interaction between chimeric virions and their target counterparts, recognizing the epitope or the streptag peptide in the viral envelope. These data provide evidence that altering the surface properties of the baculovirus virion could be of value in improving baculovirus display technology and developing new applications. PMID- 10866804 TI - Benign prostatic hyperplasia-associated prostate-specific antigen (BPSA) shows unique immunoreactivity with anti-PSA monoclonal antibodies. AB - We previously identified a modified molecular form of prostate-specific antigen that is significantly elevated in the nodular transition zone tissue of prostates with benign prostatic hyperplasia. This prostate-specific antigen form, designated BPSA, is inactive and contains clipped polypeptide bonds at amino-acid residues Lys145-146 and Lys182-183. BPSA is not elevated in prostate cancer tissues and may therefore be a prostate-specific antigen marker to better discriminate benign prostatic hyperplasia from early prostate cancer. In this work we characterize the immunoreactivity of BPSA in competition assays with prostate-specific antigen using anti-prostate-specific antigen mAb recognizing six different epitopes on the prostate-specific antigen molecule. One mAb showed > 50% loss of immunoreactivtiy with BPSA compared with prostate-specific antigen, while the binding of two mAbs was largely unaffected and three mAbs had intermediate reactivity. BPSA purified from prostate tissue and seminal plasma, as well as BPSA generated in vitro by mild trypsin-treatment were found to have a similar pattern of reactivity to the six mAbs. However, other forms of inactive seminal plasma prostate-specific antigen, either intact or clipped at Lys145 only, had immunoreactivity similar to total prostate-specific antigen. These results demonstrate that BPSA has unique immunological properties from other forms of prostate-specific antigen, which should allow the development of BPSA specific mAbs for the study of benign prostatic hyperplasia. Measurement of BPSA levels in the serum may help discriminate benign prostatic hyperplasia from early prostate cancer. PMID- 10866805 TI - A heparin-binding growth factor, midkine, binds to a chondroitin sulfate proteoglycan, PG-M/versican. AB - Midkine is a heparin-binding growth factor with survival-promoting and migration enhancing activities. In order to understand the regulation of midkine signaling, we isolated midkine-binding proteoglycans from day 13 mouse embryos, when midkine is intensely expressed. Deglycosylation followed by SDS/PAGE revealed various protein bands; one of these was identified as PG-M/versican by in gel trypsin digestion and sequencing the resulting peptides. PG-M/versican isolated from day 13 mouse embryos bound midkine with a Kd of 1.0 nM. Pleiotrophin/heparin-binding growth-associated molecule, which has a structure related to midkine, was also bound similarly. Digestion with chondroitinase ABC, AC-I or B abolished the binding to midkine. Heparin as well as chondroitin sulfate D and E inhibited the binding. After chondroitinase ABC digestion, the midkine-binding PG-M/versican released 4-sulfated, 6-sulfated, 2, 6-disulfated and 4,6-disulfated unsaturated disaccharides. These results suggest that midkine binds to a polysulfated domain in the chondroitin sulfate chain with a region of dermatan sulfate structure. This proteoglycan may modulate the midkine activity, as binding to midkine can enhance midkine action by concentrating it to the cell periphery or inhibit the action by competing with the binding to a signaling receptor. PMID- 10866806 TI - Mutational analysis of arginine 177 in the nucleotide binding site of beta-actin. AB - Actin ADP-ribosylated at arginine 177 is unable to hydrolyze ATP, and the R177 side chain is in a position similar to that of the catalytically essential lysine 71 in heat shock cognate protein Hsc70, another member of the actin-fold family of proteins. Therefore, actin residue R177 has been implicated in the mechanism of ATP hydrolysis. This paper compares wild-type beta-actin with a mutant in which R177 has been replaced by aspartic acid. The mutant beta-actin was expressed in Saccharomyces cerevisiae and purified by DNase I-affinity chromatography. The mutant protein exhibited a reduced thermal stability and an increased nucleotide exchange rate, suggesting a weakened interdomain connection. The ATPase activity of G-actin and the ATPase activity expressed during polymerization were unaffected by the R177D replacement, showing that this residue is not involved in catalysis. In the presence of polymerizing salts, ATP hydrolysis by both wild-type Mg-beta-actin and the mutant protein preceded filament formation. With the mutant actin, the initial rate of ATP hydrolysis was as high as with wild-type actin, but polymer formation was slower, reached lower steady-state levels, and the polymers formed exhibited much lower viscosity. The critical concentration of polymerization (Acc) of the mutant actin was increased 10-fold as compared to wild-type actin. Filaments formed from the R177D mutant beta-actin bound phalloidin. PMID- 10866807 TI - Phytanoyl-CoA hydroxylase activity is induced by phytanic acid. AB - Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) is a branched-chain fatty acid present in various dietary products such as milk, cheese and fish. In patients with Refsum disease, accumulation of phytanic acid occurs due to a deficiency of phytanoyl-CoA hydroxylase, a peroxisomal enzyme containing a peroxisomal targeting signal 2. Recently, phytanoyl-CoA hydroxylase cDNA has been isolated and functional mutations have been identified. As it has been shown that phytanic acid activates the nuclear hormone receptors peroxisome proliferator activated receptor (PPAR)alpha and all three retinoid X receptors (RXRs), the intracellular concentration of this fatty acid should be tightly regulated. When various cell lines were grown in the presence of phytanic acid, the activity of phytanoyl-CoA hydroxylase increased up to four times, depending on the particular cell type. In one cell line, HepG2, no induction of phytanoyl-CoA hydroxylase activity was observed. After addition of phytanic acid to COS-1 cells, an increase in phytanoyl-CoA hydroxylase activity was observed within 2 h, indicating a quick cell response. No stimulation of phytanoyl-CoA hydroxylase was observed when COS-1 cells were grown in the presence of clofibric acid, 9-cis retinoic acid or both ligands together. This indicates that the activation of phytanoyl-CoA hydroxylase is not regulated via PPARalpha or RXR. However, stimulation of PPARalpha and all RXRs by clofibric acid and 9-cis-retinoic acid was observed in transient transfection assays. These results suggest that the induction of phytanoyl-CoA hydroxylase by phytanic acid does not proceed via one of the nuclear hormone receptors, RXR or PPARalpha. PMID- 10866808 TI - Toxic lactonic lipopeptide from food poisoning isolates of Bacillus licheniformis. AB - Toxins from three Bacillus licheniformis strains connected to a fatal food poisoning were isolated and their structures elucidated. Toxins were purified from methanol extracts of the B. licheniformis biomass using boar sperm cells as the toxicity indicator. The HPLC purified toxins showed protonated masses m/z 1007, 1021 and 1035 in MALDI-TOF-MS. The toxins isolated from the strains of different origins contained the same three components of which and each had a same amino-acid residues L-Gln, L-Leu, D-Leu, L-Val, L-Asp, D-Leu and L-Ile in that order. Toxins were identified as lichenysin A, a cyclic lactonic heptalipopeptide in which the main 3-hydroxy fatty acids are 13-15 carbons in length. We showed that the toxins from food and food poisoning isolates of B. licheniformis were identical to lichenysin A both in the structure and in the toxic symptoms induced to boar spermatozoa. Confocal laser scanning microscopy showed that the acrosome and the plasma membrane of boar spermatozoa were the targets of lichenysin A toxicity. PMID- 10866809 TI - cDNA cloning of bradykinin-potentiating peptides-C-type natriuretic peptide precursor, and characterization of the novel peptide Leu3-blomhotin from the venom of Agkistrodon blomhoffi. AB - A cDNA clone, 1.8 kb long, was isolated from a venom gland cDNA library of Agkistrodon blomhoffi that encodes a large plurifunctional precursor composed of 263 amino-acid residues. Nucleotide sequence analysis of this clone revealed that sequences which code for blomhotin and a novel peptide Leu3-blomhotin are located in the N-terminal region of the precursor polypeptide, followed by four tandemly aligned sequences which code for three types of bradykinin-potentiating peptide. In the C-terminal region, the sequence for the C-type natriuretic peptide was located along with a preceding processing signal. The deduced amino-acid sequences for the four bradykinin-potentiating peptides coincided exactly with previously known sequences for potentiator B, potentiator C and potentiator E. The actual Leu3-blomhotin peptide was subsequently isolated from the venom of A. blomhoffi and characterized. Leu3-blomhotin possesses contractile activity in isolated rat stomach fundus smooth muscle in the same manner as blomhotin. Furthermore, it was shown that blomhotin and Leu3-blomhotin retained activity to inhibit the angiotensin-converting enzyme. PMID- 10866810 TI - Conformational variants of human alpha2-macroglobulin are reflected in a C terminal 'switch region'. AB - Human alpha2-macroglobulin displays extensive conformational changes when induced to transform into new quaternary structures, which are eliminated from the systemic circulation by receptor-mediated endocytosis. One major region involved in these conformational changes is located in a segment of 30 amino acids from Glu1314 to Ser1343 (-Glu-Glu-Phe-Pro-Phe-Ala-Leu-Gly-Val-Gln-Thr-Leu-Pro-Gln-Thr Cys-Asp -Glu-Pro-Lys-Ala-His-Thr-Ser-Phe-Gln-Ile-Ser-Leu-Ser-), which we term the 'switch region' of alpha2-macroglobulin, as deduced by immunochemical techniques. Monoclonal antibodies were generated using either native, methylamine-treated or the 18-kDa C-terminal receptor-binding fragment as the immunogen. From an extensive number of obtained hybridomas, 11 mAbs were selected because of their capacity to bind to the C-terminal fragment. Irrespective of the original configuration of the antigen used for immunization, seven of the antibodies were shown to be reactive with a set of overlapping epitopes, closely positioned within the 'switch region', as confirmed by the use of synthetic peptides covering the entire C-terminal fragment. The specificities of the seven individual antibodies, as determined by ELISA and BIAcore technologies, revealed a pronounced conformational pleomorphism in the 'switch region'. The results indicate that the 'switch region' may be involved in the exposure of the receptor recognition site and can be used as an indicator region for different conformational states of alpha2-macroglobulin. PMID- 10866811 TI - HMG1 protein stimulates DNA end joining by promoting association of DNA molecules via their ends. AB - High mobility group (HMG) 1 protein is a highly abundant and an evolutionarily conserved chromosomal protein with two homologous DNA-binding domains (HMG boxes), A and B, attached by a short basic region to an acidic C-terminal tail. The protein has been implicated in a number of fundamental biological processes including DNA replication, transcription, recombination and repair. We demonstrate that HMG1 is able to enhance cohesive-end and blunt-end DNA ligation by T4 DNA ligase via its B domain. The C-terminal flanking sequence of the B domain (seven basic residues out of approximately 18) and a number of conserved amino-acid residues within the HMG box (mainly basic or hydrophobic) are required for efficient stimulation of ligation. Pull-down assays, electron and scanning force microscopy revealed that HMG1 can associate two DNA molecules via their ends even in the absence of complementary overhangs. We propose that HMG1 protein may be involved in the rejoining of DNA breaks by different DNA ligases due to its ability to bring DNA duplexes and their termini into a close proximity while leaving the ends accessible for ligation. PMID- 10866812 TI - Charged amino-acid residues in transmembrane domains of the plastidic ATP/ADP transporter from arabidopsis are important for transport efficiency, substrate specificity, and counter exchange properties. AB - Structure-function relationships of the plastidic ATP/ADP transporter from Arabidopsis thaliana have been determined using site-directed mutants at positions K155, E245, E385, and K527. These charged residues are found within highly conserved domains of homologous transport proteins from plants and bacteria and are located in predicted transmembrane regions. Mutants of K155 to K155E, K155R, or K155Q reduced ATP transport to values between 4 and 16% of wild type uptake, whereas ADP transport was always less then 3% of the wild-type value. Site-directed mutations in which glutamate at positions 245 or 385 was replaced with lysine, abolished transport. However, conservative (E245D, E385D) or neutral (E245Q, E385Q) replacement at these two positions allowed transport. The fourth reciprocal exchange, K527E, also abolished uptake of both adenylates. K527R and K527Q were unable to transport ATP, but ADP transport remained at 35 and 27%, respectively, of the wild-type activity. There was a 70-fold decreased apparent affinity of K527R for ATP, but only a twofold decrease for ADP. The efflux of ATP, but not ADP, was also greatly reduced in K527R. These observations show strikingly that K527 plays a role in substrate specificity that is manifest in both the influx and efflux components of this antiporter. PMID- 10866813 TI - Distribution, cloning, and characterization of porcine nucleoside triphosphate diphosphohydrolase-1. AB - In this study, we have investigated the distribution of the enzyme nucleoside triphosphate diphosphohydrolase-1 (NTPDase1; EC 3.6.1.5) in a subset of pig tissues by biochemical activity and Western blotting with antibodies against porcine NTPDase1. The highest expression of this enzyme was found in vascular endothelium, smooth muscle, spleen and lung. The complete cDNA of NTPDase1 from aorta endothelial cells was sequenced using primer walking. The protein consists of 510 amino acids, with a calculated molecular mass of 57 756 Da. The amino-acid sequence indicated seven putative N-glycosylation sites and one potential intracellular cGMP- and cAMP-dependent protein kinase phosphorylation site. As expected, the protein has a very high homology to other known mammalian ATPDases and CD39 molecules, and includes all five apyrase conserved regions. Expression of the complete cDNA in COS-7 cells confirmed that NTPDase1 codes for a transmembrane glycoprotein with ecto-ATPase and ecto-ADPase activities. Two proteolytic products of NTPDase1, with molecular mass of 54 and 27 kDa, respectively, were consistently present in proteins from transfected COS-7 cells and in particulate fractions from different tissues. A trypsin cleavage site, giving rise to these two cleavage products, was identified. In order to remain enzymatically active, the two cleavage products have to interact by non-covalent interactions. PMID- 10866814 TI - Properties of three isoforms of the 116-kDa subunit of vacuolar H+-ATPase from a single vertebrate species. Cloning, gene expression and protein characterization of functionally distinct isoforms in Gallus gallus. AB - Vacuolar H+-ATPases (V-ATPases) are involved in a wide variety of essential cellular processes. An unresolved question is how the cell regulates the activity of these proton pumps and their targeting to distinct cellular compartments. There is growing evidence for the presence of subunit diversity amongst V-pumps, particularly regarding the 116-kDa subunit (called the a subunit). We have cloned and characterized three isoforms (a1, a2 and a3) of this subunit from chicken. The amino-acid sequences of these homologues are approximately 50% similar and their nucleotide differences indicate that they are products of distinct genes. The levels of mRNA expression of these isoforms was quantified by ribonuclease protection analysis. The a1 and a2 isoforms have a similar tissue distribution, with the highest level of mRNA expression in brain, an intermediate level in kidney and relatively low levels in liver and bone. In contrast, the highest level of expression of the a3 isoform is in bone and liver, with a moderate level in kidney, and the lowest level in brain. An antibody against the a1 isoform reacted with a 116 kDa protein in a brain V-ATPase preparation that was not detected in bone or liver V-ATPase preparations, whereas an antibody against the a3 isoform reacted with a 116-kDa peptide in bone and liver, but not brain V ATPases preparations. The bone and brain V-ATPases showed differential sensitivity to the inhibitors bafilomycin and (2Z,4E)-5-(5,6-dichloro-2-indolyl) 2-methoxy-N-[4-(2, 2,6,6-tetramethyl)piperidinyl]-2,4-pentadienamide. Thus, this work demonstrates the presence of structurally and functionally distinct V ATPases in a single vertebrate species. PMID- 10866815 TI - Involvement of Gln937 of Streptococcus downei GTF-I glucansucrase in transition state stabilization. AB - Multiple alignment of deduced amino-acid sequences of glucansucrases (glucosyltransferases and dextransucrases) from oral streptococci and Leuconostoc mesenteroides has shown them to share a well-conserved catalytic domain. A portion of this domain displays homology to members of the alpha-amylase family (glycoside hydrolase family 13), which all have a (beta/alpha)8 barrel structure. In the glucansucrases, however, the alpha-helix and beta-strand elements are circularly permuted with respect to the order in family 13. Previous work has shown that amino-acid residues contributing to the active site of glucansucrases are situated in structural elements that align with those of family 13. In alpha amylase and cyclodextrin glucanotransferase, a histidine residue has been identified that acts to stabilize the transition state, and a histidine is conserved at the corresponding position in all other members of family 13. In all the glucansucrases, however, the aligned position is occupied by glutamine. Mutants of glucosyltransferase I were constructed in which this glutamine, Gln937, was changed to histidine, glutamic acid, aspartic acid, asparagine or alanine. The effects on specific activity, ability to form glucan and ability to transfer glucose to a maltose acceptor were examined. Only histidine could substitute for glutamine and maintain Michaelis-Menten kinetics, albeit at a greatly reduced kcat, showing that Gln937 plays a functionally equivalent role to the histidine in family 13. This provides additional evidence in support of the proposed alignment of the (beta/alpha)8 barrel structures. Mutation at position 937 altered the acceptor reaction with maltose, and resulted in the synthesis of novel gluco-oligosaccharides in which alpha1,3-linked glucosyl units are joined sequentially to maltose. PMID- 10866816 TI - Kinetics of tryptophan oxidation in plasma lipoproteins by myeloperoxidase generated HOCl. AB - The relative susceptibility of the apoprotein components of human lipoproteins [high-density lipoprotein (HDL) and low-density lipoprotein (LDL)] and their subclasses to oxidation by the myeloperoxidase/H2O2/Cl- system in vitro was studied by measuring the decrease in rate of tryptophan fluorescence. Whereas the lipoprotein-modification rate showed a saturation type of dependence on the concentration of myeloperoxidase, a biphasic dependence on the concentration of the lipoproteins was found. High concentrations of H2O2 were also found to inhibit tryptophan oxidation in LDL but to a lesser extent in HDL. The optimal rate of LDL and HDL modification was observed at pH 6.0. HDL was modified much more rapidly than LDL, which may be due to differences in size and different relative contents of protein and lipids per particle. No differences in rates of modification of LDL subclasses were observed, when the assays were standardized to equal LDL protein concentrations, but, when standardized to equal particle mass, an optimum at subclass 8 was found, which is probably due to differences in apolipoprotein B-100 conformation. It was concluded that HDL may have a beneficial effect in retarding LDL modification in inflammatory processes. PMID- 10866817 TI - Cytochrome b558/566 from the archaeon Sulfolobus acidocaldarius has a unique Asn linked highly branched hexasaccharide chain containing 6-sulfoquinovose. AB - Cytochrome b558/566 from the archaeon Sulfolobus acidocaldarius (DSM 639) has been described as a novel highly glycosylated membrane-bound b-type hemoprotein [Hettmann, T., Schmidt, C. L., Anemuller, S., Zahringer, U., Moll, H., Petersen, A. & Schafer, G. (1998) J. Biol. Chem. 273, 12032-12040]. The purified cytochrome b558/566 was characterized by MALDI MS as a 64-kDa (glyco)protein expressing 17% glycosylation. Detailed chemical studies showed that it was exclusively O mannosylated with monosaccharides and N-glycosylated with at least seven hexasaccharide units having the same unique structure. The hexasaccharide was released by cleavage with peptide:N-glycosidase (PNGase) F and found to consist of two residues each of Man and GlcNAc and one residue each of Glc and 6-deoxy-6 sulfoglucose (6-sulfoquinovose). The last sugar has been known as a component of glycolipids of plants and some prokaryotes, but has not been hitherto found in bacterial glycoproteins. Digestion with trypsin/pronase gave a mixture of glycopeptides with the same Asn-linked hexasaccharide chain, from which an N glycosylated Tyr-Asn dipeptide was purified by gel chromatography and anion exchange HPLC. Studies of the degradation products using methylation analysis, ESI MS, MALDI MS, and 1H and 13C NMR spectroscopy, including 1H,13C HMQC and NOESY experiments, established the structure of the unique Asn-linked hexasaccharide chain of cytochrome b558/566. PMID- 10866818 TI - Mechanisms of induction of human tissue inhibitor of metalloproteinases-1 (TIMP 1) gene expression by all-trans retinoic acid in combination with basic fibroblast growth factor. AB - The addition of all-trans retinoic acid (ATRA) in combination with basic fibroblast growth factor (bFGF) to human fibroblasts results in a synergistic induction of tissue inhibitor of metalloproteinases-1 (TIMP-1) protein production. The synergistic stimulation of TIMP-1 protein by ATRA and bFGF increased across 72 h. An incubation of 10 min to 12 h with bFGF alone followed by ATRA gave a similar synergistic induction of TIMP-1 protein to that seen with both agents together. Treatment of cells with ATRA first followed by bFGF was ineffective. Expression of RARbeta mRNA was induced by ATRA alone, but not further induced by ATRA and bFGF; expression of RARgamma mRNA was induced by both ATRA or bFGF alone, and further induced by both reagents together; expression of RXRgamma was repressed by ATRA alone, but not by ATRA in combination with bFGF. Steady-state levels of TIMP-1 mRNA were induced 14 to 40-fold above control by ATRA and bFGF. Treatment with ATRA and bFGF did not alter the stability of TIMP-1 mRNA. The induction of TIMP-1 mRNA by ATRA and bFGF was greatly diminished by cycloheximide and therefore required new protein synthesis. The tyrosine kinase inhibitor genistein caused a dose-dependent inhibition of TIMP-1 protein induction by ATRA and bFGF. A MEK1 inhibitor (PD98059) inhibited both basal and induced levels of TIMP-1. At high concentrations, p38 MAP kinase inhibitors further enhanced the synergistic stimulation of TIMP-1 protein by ATRA and bFGF, but at these concentrations, p42/44 MAP kinase was strongly activated. These data begin to elucidate the mechanisms by which TIMP-1 gene expression can be upregulated. PMID- 10866819 TI - Treatment of cells with the polyamine analog N, N11-diethylnorspermine retards S phase progression within one cell cycle. AB - When Chinese hamster ovary cells were seeded in the presence of the spermine analog N1,N11-diethylnorspermine (DENSPM), cell proliferation ceased; this was clearly apparent by cell counting 2 days after seeding the cells. However, 1 day after seeding there was a slight difference in cell number between control and DENSPM-treated cultures. To investigate the reason for this easily surpassed slight difference, we used a sensitive bromodeoxyuridine/flow cytometry method. Cell cycle kinetics were studied during the first cell cycle after seeding cells in the absence or presence of DENSPM. Our results show that DENSPM treatment did not affect the progression of the cells through G1 or the first G1/S transition that took place after seeding the cells. The first cell cycle effect was a delay in S phase as shown by an increase in the DNA synthesis time. The following G2/M transition was not affected by DENSPM treatment. DENSPM treatment inhibited the transient increases in putrescine, spermidine, and spermine pools that took place within 24 h after seeding. Thus, in conclusion, the first cell cycle phase affected by the inhibition of polyamine biosynthesis caused by DENSPM was the S phase. Prolongation of the other cell cycle phases occurred at later time points, and the G1 phase was affected before the G2/M phase. PMID- 10866820 TI - Fluorescent microplate assay for cancer cell-associated cathepsin B. AB - Cathepsin B and in particular cell-surface and secreted cathepsin B has been implicated in the invasive and metastatic phenotype of numerous types of cancer. We describe here a method to easily survey cancer cell lines for cathepsin B activity using the highly selective substrate Z-Arg-Arg-AMC. Intact human U87 glioma cells hydrolyze Z-Arg-Arg-AMC with a Km of 460 microM at pH 7.0 and 37 degrees C. This is nearly the same as the Km of 430 microM obtained with purified cathepsin B assayed under the same conditions. The pericellular (i.e. both cell surface and released) cathepsin B activity was inhibited by the cysteine protease inhibitors E-64, leupeptin, Mu-Np2-HphVS-2Np, Mu-Leu-HpHVSPh and the cathepsin B selective inhibitor Mu-Tyr(3,5 I2)-HphVSPh with IC50 values similar to those observed for the inhibition of purified human liver cathepsin B. Other human cancer cell lines with measurable pericellular cathepsin B activity included HT 1080 fibrosarcoma, MiaPaCa pancreatic, PC-3 prostate and HCT-116 colon. Cathepsin B activity correlated with protein levels of cathepsin B as determined by immunoblot analysis. Pericellular cathepsin B activity was also detected in the rat cell lines MatLyLu prostate and Mat B III adenocarcinoma and in the murine lines B16a melanoma and Lewis lung carcinoma. The ability to determine pericellular cathepsin B activity will be useful in selecting appropriate cell lines for use in vivo when analyzing the effects of inhibiting cathepsin B activity on tumor growth and metastasis. PMID- 10866821 TI - Transition metal-mediated glycoxidation accelerates cross-linking of beta-amyloid peptide. AB - beta-Amyloid deposits, hallmarks of Alzheimer's disease, contain both sugar derived 'advanced glycation end products' (AGEs) and copper and iron ions. Our in vitro experiments using synthetic beta-amyloid peptide and glucose or fructose show that formation of covalently cross-linked high-molecular-mass beta-amyloid peptide oligomers is accelerated by micromolar amounts of copper (Cu+, Cu2+) and iron (Fe2+, Fe3+) ions. Formation of these covalent AGE cross-links can be inhibited by capping agents of amino groups, redox-inactive metal chelators and antioxidants, suggesting that these drugs may be able to slow down the formation of insoluble beta-amyloid deposits in vivo and possibly the progression of Alzheimer's disease. PMID- 10866822 TI - In vitro inhibition and intracellular enhancement of lysosomal alpha galactosidase A activity in Fabry lymphoblasts by 1-deoxygalactonojirimycin and its derivatives. AB - Fabry disease is a lysosomal storage disorder caused by deficient lysosomal alpha galactosidase A (alpha-Gal A) activity. Deficiency of the enzyme activity results in progressive deposition of neutral glycosphingolipids with terminal alpha galactosyl residue in vascular endothelial cells. We recently proposed a chemical chaperone therapy for this disease by administration of 1 deoxygalactonojirimycin, a potent inhibitor of the enzyme, at subinhibitory intracellular concentrations [Fan, J.-Q., Ishii, S., Asano, N. and Suzuki, Y. (1999) Nat. Med. 5, 112-115]. 1-Deoxygalactonojirimycin served as a specific chaperone for those mutant enzymes that failed to maintain their proper conformation to avoid excessive degradation. In order to establish a correlation between in vitro inhibitory activity and intracellular enhancement activity of the specific chemical chaperone, a series of 1-deoxygalactonojirimycin derivatives were tested for activity with both alpha-Gal A and Fabry lymphoblasts. 1-Deoxygalactonojirimycin was the most potent inhibitor of alpha Gal A with an IC50 value of 0.04 microM. alpha-Galacto-homonojirimycin, alpha allo-homonojirimycin and beta-1-C-butyl-deoxygalactonojirimycin were effective inhibitors with IC50 values of 0.21, 4.3 and 16 microM, respectively. N Alkylation, deoxygenation at C-2 and epimerization at C-3 of 1 deoxygalactonojirimycin markedly lowered or abolished its inhibition toward alpha Gal A. Inclusion of 1-deoxygalactonojirimycin, alpha-galacto-homonojirimycin, alpha-allo-homonojirimycin and beta-1-C-butyl-deoxygalactonojirimycin at 100 microM in culture medium of Fabry lymphoblasts increased the intracellular alpha Gal A activity by 14-fold, 5.2-fold, 2.4-fold and 2.3-fold, respectively. Weaker inhibitors showed only a minimum enhancement effect. These results suggest that more potent inhibitors act as more effective specific chemical chaperones for the mutant enzyme, and the potent competitive inhibitors of alpha-Gal A are effective specific chemical chaperones for Fabry disease. PMID- 10866823 TI - Evidence that ganglioside enriched domains are distinct from caveolae in MDCK II and human fibroblast cells in culture. AB - Cultures of MDCK II and human fibroblast cells were fed radioactive sphingosine and a radioactive GM3 ganglioside derivative containing a photoactivable group. The derived cell homogenates were treated with Triton X-100 and fractionated by sucrose-gradient centrifugation to prepare a detergent-insoluble membrane fraction known to be enriched in sphingolipid and caveolin-1, i.e. of caveolae. The detergent-insoluble membrane fraction prepared after feeding [1 3H]sphingosine to cells, was found to be highly enriched, with respect to protein content, in metabolically radiolabeled sphingomyelin and glycosphingolipids (about 18-fold). By feeding cells photoactivable radioactive GM3, after 2 h chase, caveolin-1, CAV1, and proteins of high molecular mass became cross-linked to GM3, the cross-linking complexes being highly concentrated in the detergent insoluble membrane fraction. The interaction between the ganglioside derivative and CAV1 was a time-dependent, transient process so that CAV1 cross-linking to GM3 was hardly detectable after a 24-h chase followed the pulse time. After a 24 h chase, only the high molecular mass proteins cross-linked to GM3 could be clearly observed. These results suggest that a portion of the GM3 administered to cells enters caveolae and moves to the glycosphingolipid domains, or enters caveolae that are then rapidly catabolized. Electron microscopy of cells in a culture immunostained with a monoclonal antibody to GM3 and a secondary gold conjugated antibody detected several clusters of gangliosides on the plasma membranes separate from caveolae; gangliosides located inside the caveolae could not be detected. Scanning confocal microscopy of cells immunostained with anti GM3 and anti-CAV1 Ig showed only a very small overlap with the CAV1 and GM3 signals. Thus, the biochemical and microscopic studies suggest that caveolae contain at most a low level of gangliosides and are separate from the GM3 ganglioside enriched domains. PMID- 10866824 TI - Cadmium blocks hypoxia-inducible factor (HIF)-1-mediated response to hypoxia by stimulating the proteasome-dependent degradation of HIF-1alpha. AB - Cadmium is a substantial industrial and environmental pollutant which seriously impairs erythropoiesis. Cd has been demonstrated to aggravate anemia by suppressing erythropoietin gene expression in anemic patients. As hypoxic induction of erythropoietin mRNA depends on a transcription factor, hypoxia inducible factor 1 (HIF-1), we hypothesized that Cd suppresses the hypoxic activation of HIF-1. In hypoxic Hep3B cells, all mRNAs of various genes, which are known to be upregulated by HIF-1 activation under hypoxia, were suppressed by Cd in a dose-dependent manner. Cd inhibited the hypoxia-induced activity of luciferase in 293 cells which was transfected with a reporter plasmid carrying a hypoxia response element. By electrophoretic mobility gel shift assay, Cd inhibited the DNA-binding activity of HIF-1 in hypoxic Hep3B cells. Cd reduced the amount of HIF-1alpha protein in hypoxia, whereas it didn't affect HIF-1 alpha mRNA levels. Moreover, Cd inhibited HIF-1alpha accumulation induced by cobalt and desferrioxamine. Antioxidants and a proteasome inhibitor prevented the HIF-1alpha degradation caused by Cd. The possibility that oxidative stress mediates this action of Cd was examined. Cd didn't affect protein oxidation and reduced glutathione levels in hypoxic cells. These results indicate that Cd triggers a redox/proteasome-dependent degradation of HIF-1alpha protein, reducing HIF-1 activity and in turn suppressing the hypoxic induction of hypoxia-inducible genes. PMID- 10866825 TI - Solution structure of the functional domain of Paracoccus denitrificans cytochrome c552 in the reduced state. AB - In order to determine the solution structure of Paracoccus denitrificans cytochrome c552 by NMR, we cloned and isotopically labeled a 10.5-kDa soluble fragment (100 residues) containing the functional domain of the 18.2-kDa membrane bound protein. Using uniformly 15N-enriched samples of cytochrome c552 in the reduced state, a variety of two-dimensional and three-dimensional heteronuclear double-resonance NMR experiments was employed to achieve complete 1H and 15N assignments. A total of 1893 distance restraints was derived from homonuclear 2D NOESY and heteronuclear 3D-NOESY spectra; 1486 meaningful restraints were used in the structure calculations. After restrained energy minimization a family of 20 structures was obtained with rmsd values of 0.56 +/- 0. 10 A and 1.09 +/- 0.09 A for the backbone and heavy atoms, respectively. The overall topology is similar to that seen in previously reported models of this class of proteins. The global fold consists of two long helices at the N-terminus and C-terminus and three shorter helices surrounding the heme moiety; the helices are connected by well defined loops. Comparison with the X-ray structure shows some minor differences in the positions of the Trp57 and Phe65 side-chain rings as well as the heme propionate groups. PMID- 10866826 TI - Analysis of CAK activities from human cells. AB - The cdk-activating kinase (CAK) activates cyclin-dependent kinases (cdks) that control cell-cycle progression by phosphorylating a threonine residue conserved in cdks. CAK from humans contains p40MO15 (cdk7), cyclin H and MAT1, which are also subunits of transcription factor IIH where they phosphorylate the C-terminal domain of the large subunit of RNA polymerase II. In contrast, budding yeast Cak1p is a monomeric enzyme without C-terminal domain kinase activity. Here, we analyze CAK activities in HeLa cells using cdk2-affinity chromatography. In addition to MO15, a second CAK activity was detected that runs on gel filtration at 30-40 kDa. This activity phosphorylated and activated cdk2 and cdk6. Furthermore, this 'small CAK' activity resembled Cak1p rather than MO15 in terms of substrate specificity, reactivity to antibodies against MO15 and Cak1p, and sensitivity to 5'-fluorosulfonylbenzoyladenosine, an irreversible inhibitory ATP analog. Our findings suggest the presence of at least two different CAK activities in human cells. PMID- 10866827 TI - Formation of acyl radical in lipid peroxidation of linoleic acid by manganese dependent peroxidase from Ceriporiopsis subvermispora and Bjerkandera adusta. AB - Lipid peroxidation by managanese peroxidase (MnP) is reported to decompose recalcitrant polycyclic aromatic hydrocabon (PAH) and nonphenolic lignin models. To elucidate the oxidative process, linoleic acid and 13(S)-hydroperoxy-9Z,11E octadecadienoic acid [13(S)-HPODE] were reacted with MnPs from Ceriporiopsis subvermispora and Bjerkandera adusta and the free radicals produced were analyzed by ESR. When the MnPs were reacted with 13(S)-HPODE in the presence of Mn(II), H2O2 and tert-nitrosobutane (t-NB), the ESR spectrum contained a sharp triplet of acyl radical (aN = 0.81 mT). Formation of acyl radical was also observed in the reactions of Mn(III)-tartrate with 13(S)-HPODE and with linoleic acid, but the latter reaction occurred explosively after an induction period of around 30 min. Reactions of MnP with linoleic acid in the presence of Mn(II), H2O2 and t-NB gave no spin adducts while addition of t-NB after preincubation of linoleic acid with MnP/Mn(II)/H2O2 for 2 h gave spin adducts of carbon-centered (aN = 1.53 mT, aH = 0.21 mT) and acyl (aN = 0.81 mT) radicals. In contrast to linoleic acid, methyl linoleate and oleic acid were not peroxidized by MnP and chelated Mn(III) within a few hours, indicating that structures containing both the 1,4-pentadienyl moiety and a free carboxyl group are necessary for inducing the peroxidation in a short reaction time. These results indicate that MnP-dependent lipid peroxidation is not initiated by direct abstraction of hydrogen from the bis-allylic position during turnover but proceeds by a Mn(III)-dependent hydrogen abstraction from enols and subsequent propagation reactions involving the formation of acyl radical from lipid hydroperoxide. This finding expands the role of chelated Mn(III) from a phenol oxidant to a strong generator of free radicals from lipids and lipid hydroperoxides in lignin biodegradation. PMID- 10866828 TI - Cytochrome c6 from Cyanophora paradoxa. Characterization of the protein and the cDNA of the precursor and import into isolated cyanelles. AB - In the eukaryotic alga Cyanophora paradoxa, which does not contain plastocyanin, photosynthetic electron transport from the cytochrome b6/f complex to photosystem I is mediated by cytochrome c6. Cytochrome c6 was purified to homogeneity by column chromatography and FPLC. The relative molecular mass of the holoprotein was determined by two different mass spectrometric methods (californium-252 plasma desorption and UV matrix-assisted laser desorption ionization) giving 9251 +/- 3.3 Da. N-terminal Edman microsequencing yielded information on approx. 30 amino acid residues. Based on these data and on highly conserved regions of cytochromes c6, degenerate oligonucleotides were designed and used for PCR to amplify the genomic DNA of C. paradoxa. Screening of a C. paradoxa cDNA library yielded several clones coding for preapo-cytochrome c6. The deduced sequence of the mature protein was verified by plasma desorption mass spectrometric peptide mapping and shows high similarity to those of cytochromes c6 from cyanobacteria and algae. Cytochrome c6 appears to be encoded by a single nuclear gene (petJ) in C. paradoxa. As the mature protein is located in the lumen of the thylakoid membrane, it has to traverse three biological membranes as well as the unique peptidoglycan layer of the cyanelles before it reaches its final subcellular locale. Thus the transit sequence is composed of two different targeting signals: a stroma targeting peptide resembling those of higher plants with respect to hydropathy plots and amino acid composition and a hydrophobic signal peptide functioning as a thylakoid-traversing domain. There are indications for alternative sorting of part of the cyanelle cytochrome c6 pool to the periplasmic space. This is the first known bipartite transit sequence of a cyanelle precursor protein from C. paradoxa, a model organism concerning the endosymbiotic origin of plastids. Labeled precursor is efficiently imported into isolated cyanelles, then routed into thylakoids and processed to the mature protein. Hitherto, in vitro protein translocation was not reported for cyanobacterial-type thylakoids. PMID- 10866829 TI - Thermodynamics and dynamics of histidine-binding protein, the water-soluble receptor of histidine permease. Implications for the transport of high and low affinity ligands. AB - The bacterial histidine permease is a model system for ABC transporters (traffic ATPases). The water-soluble receptor of this permease, HisJ, binds L-histidine and L-arginine (tightly) and L-lysine and L-ornithine (less tightly) in the periplasm, interacts with the membrane-bound complex (HisQMP2) and induces its ATPase activity, which results in ligand translocation. HisJ is a two-domain protein; in the absence of ligand, the cleft between two domains is open and binding of substrate stabilizes the closed conformation. Surprisingly, various liganded HisJ forms display substantial differences in their physicochemical characteristics and capacity to induce the ATPase. This is due to either different effects of the individual ligands on the respective closed structures, or to different equilibria being reached for each ligand between the open liganded form and the closed liganded form [Wolf, A. , Lee, K.C., Kirsch, J.F. & Ames, G.F.-L. (1996) J. Biol. Chem. 271, 21243-21250]. In this work, time resolved measurements of the decay of intrinsic HisJ fluorescence and of the decay of the anisotropy of the fluorescence, as well as the analysis of the steady-state near UV CD and fluorescence spectra, rule out the model in which the differences between liganded complexes reflect different equilibria. The decay of the anisotropy of the fluorescence shows that liganded complexes differ dramatically in their large-scale conformational dynamics. Differential scanning calorimetry (DSC) curves for the HisJ thermal unfolding are well described by a scheme of equilibrium two-state unfolding of two independent domains, which can be ascribed to the two-domain structure of HisJ. This is true both for apo-HisJ at various pH values, and for HisJ in the presence of its ligands at varying concentrations, at pH 8.3. The DSC and structural data suggest that all ligands interact more extensively with the larger domain. A qualitative model for the HisJ conformational dynamics employing the idea of a twisting movement of the domains is proposed, which explains the difference in the efficacy of the ATPase induction by the various liganded HisJ forms. PMID- 10866830 TI - Conserved aromatic residues in the transmembrane region VI of the V1a vasopressin receptor differentiate agonist vs. antagonist ligand binding. AB - Despite their opposite effects on signal transduction, the nonapeptide hormone arginine-vasopressin (AVP) and its V1a receptor-selective cyclic peptide antagonist d(CH2)5[Tyr(Me)2]AVP display homologous primary structures, differing only at residues 1 and 2. These structural similarities led us to hypothesize that both ligands could interact with the same binding pocket in the V1a receptor. To determine receptor residues responsible for discriminating binding of agonist and antagonist ligands, we performed site-directed mutagenesis of conserved aromatic and hydrophilic residues as well as nonconserved residues, all located in the transmembrane binding pocket of the V1a receptor. Mutation of aromatic residues of transmembrane region VI (W304, F307, F308) reduced affinity for the d(CH2)5[Tyr(Me)2]AVP and markedly decreased affinity for the unrelated strongly hydrophobic V1a-selective nonpeptide antagonist SR 49059. Replacement of these aromatic residues had no effect on AVP binding, but increased AVP-induced coupling efficacy of the receptor for its G protein. Mutating hydrophilic residues Q108, K128 and Q185 in transmembrane regions II, III and IV, respectively, led to a decrease in affinity for both agonists and antagonists. Finally, the nonconserved residues T333 and A334 in transmembrane region VII, controlled the V1a/V2 binding selectivity for both nonpeptide and cyclic peptide antagonists. Thus, because conserved aromatic residues of the V1a receptor binding pocket seem essential for antagonists and do not contribute at all to the binding of agonists, we propose that these residues differentiate agonist vs. antagonist ligand binding. PMID- 10866831 TI - Overproduction of Thermus sp. YS 8-13 manganese catalase in Escherichia coli production of soluble apoenzyme and in vitro formation of active holoenzyme. AB - Overproduction of Thermus sp. YS 8-13 manganese catalase in Escherichia coli BL21(DE3) was accomplished by introducing a derivative of pET-23a(+) containing a copy of the coding gene into the multicloning site. E. coli BL21(DE3)/pETMNCAT produced abundant quantities of manganese catalase as insoluble inclusion bodies. Regeneration of active catalase was achieved by denaturation in guanidine hydrochloride and subsequent dialysis in the presence of manganese ion. When the E. coli chaperone genes GroEL, GroES, DnaK, DnaJ and GrpE were coexpressed with manganese catalase, a significant fraction of the overproduced protein was partitioned into the soluble fraction. However, almost all of the soluble enzyme was isolated in a manganese-deficient apo form which could subsequently be converted into active holoenzyme by incubation with manganese ion at high temperatures. Further experiments on this apo catalase suggested that the structure of this protein was virtually identical to the active holoenzyme. PMID- 10866833 TI - Large-scale expression and thermodynamic characterization of a glutamate receptor agonist-binding domain. AB - The ionotropic glutamate receptors (GluR) are the primary mediators of excitatory synaptic transmission in the brain. GluR agonist binding has been localized to an extracellular domain whose core is homologous to the bacterial periplasmic binding proteins (PBP). We have established routine, baculovirus-mediated expression of a complete ligand-binding domain construct at the 10-L scale, yielding 10-40 milligrams of purified protein. This construct contains peptides that lie outside the PBP-homologous core and that connect the domain core to the transmembrane domains of the channel and to the N-terminal 'X'-domain. These linker peptides have been implicated in modulating channel physiology. Such extended constructs have proven difficult to express in bacteria, but the protein described here is stable and monomeric. Isothermal titration calorimetry reveals that glutamate binding to the domain involves a substantial heat capacity change and that at physiological temperatures, the reaction is both entropically and enthalpically favorable. PMID- 10866832 TI - Dietary cholic acid lowers plasma levels of mouse and human apolipoprotein A-I primarily via a transcriptional mechanism. AB - To induce dietary atherosclerosis in mice, high-fat/high-cholesterol (HF) diets are frequently supplemented with cholic acid (CA). This diet produces low plasma levels of high-density lipoprotein (HDL) and high levels of low-density lipoprotein (LDL). However, HF diets without any added CA, which more closely resemble human diets, increase levels of both HDL and LDL, suggesting that CA may be responsible for the lowering of HDL. Our aim was to examine the potential mechanism responsible for the lowering of HDL. Nontransgenic (NTg) C57BL mice and apoA-I-transgenic (apoAI-Tg) mice, with greatly increased basal apoA-I and HDL levels, were used. Mice were fed the following four diets: control (C), high fat/high-cholesterol (HF), control and 1% cholate (CA) and HF + CA. Dietary CA reduced plasma HDL levels by 35% in NTg and 250% in apoAI-Tg mice, independent of the fat or cholesterol content of the diet. Hepatic apoA-I mRNA decreased 30% in NTg and 180% in apoAI-Tg mice. Hepatic apoA-I synthesis and apoA-I mRNA transcription rates also decreased in parallel with apoA-I mRNA levels, suggesting that the CA-induced decreases in plasma apoA-I levels occurred primarily via decreasing apoA-I mRNA transcription rates. An HF diet increased HDL levels 1.8-fold in NTg and 1.5-fold in apoAI-Tg mice. Addition of CA to the HF diet lowered HDL levels by 1.6-fold in NTg and 2. 5-fold in apoAI-Tg mice. Transfection studies with the apoA-I promoter suggested the presence of a putative cis-acting element responsible for the CA-mediated down-regulation of the apoA-I promoter activity. Measurements of apoA-I regulatory protein-1 (ARP-1) mRNA, a negative regulator of the apoA-I gene in the mouse liver showed that CA increased the ARP-1 mRNA levels. Because apoA-I gene transcription alone was not sufficient to account for the lowering of plasma HDL levels, scavenger receptor B1 (SR-B1) and hepatic lipase (HL) mRNAs levels were quantitated. The levels of SR-B1 and HL mRNA were not changed by dietary CA. These studies suggest that dietary cholate regulates plasma levels of apoA-I primarily by a transcriptional mechanism via a putative bile acid response element involving a negative regulator of apoA-I, and partly by an unidentified post-transcriptional mechanism. PMID- 10866835 TI - The evolutionarily conserved porcupine gene family is involved in the processing of the Wnt family. AB - The Drosophila segment polarity gene product Porcupine (Porc) was first identified as being necessary for processing Wingless (Wg), a Drosophila Wnt (Wnt) family member. Mouse and Xenopus homologs of porc (Mporc and Xporc) were identified and found to encode endoplasmic reticulum (ER) proteins with multiple transmembrane domains. In contrast with porc, four different types of Mporc and Xporc mRNA (A-D) are generated from a single gene by alternative splicing. Mporc mRNA is differentially expressed during embryogenesis and in various adult tissues, demonstrating that the alternative splicing is regulated to synthesize the specific types of Mporc. In transfected mammalian cells, all Mporc types affect the processing of mouse Wnt 1, 3A, 4, 6, and 7B but not 5A. Furthermore, all Mporc types are co-immunoprecipitated with various Wnt proteins. These results suggest that Mporc may function as a chaperone-like molecule for Wnt. Interestingly, all Mporc types can substitute for Porc, as they are able to rescue the phenotypes of Drosophila porc embryos. Consistent with this observation, Mporc, like Porc, modifies the processing of Wg expressed in mammalian cells. These results demonstrate that the porc gene family encodes the multitransmembrane ER proteins, which are evolutionarily well conserved and involved in processing the Wnt family. PMID- 10866834 TI - Interaction of fMet-tRNAfMet and fMet-AMP with the C-terminal domain of Thermus thermophilus translation initiation factor 2. AB - Two polypeptides resistant against proteolytic digestion were identified in Thermus thermophilus translation initiation factor 2 (IF2): the central part of the protein (domains II/III), and the C-terminal domain (domain IV). The interaction of intact IF2 and the isolated proteolytic fragments with fMet tRNAfMet was subsequently characterized. The isolated C-terminal domain was as effective in binding of the 3' end of fMet-tRNAf Met as intact IF2. N-Formylation of Met-tRNAfMet was required for its efficient binding to the C-terminal domain. This suggests that the interaction between the C-terminal domain and the 3' end of fMet-tRNAfMet is responsible for the recognition of fMet-tRNAfMet by IF2 during translation initiation. Moreover, it was demonstrated that fMet-AMP is a minimal ligand of IF2. fMet-AMP inhibits fMet-tRNAfMet binding to IF2 as well as the activity of IF2 in the stimulation of ApUpG-dependent ribosomal binding of fMet-tRNAf Met. Specific interaction of fMet-AMP with IF2 was demonstrated by 1H NMR spectroscopy. These findings indicate that fMet-AMP and the 3' terminal fMet adenosine of fMet-tRNAfMet use the same binding site on the C-terminal domain of IF2 and imply that the interaction between the C-terminal domain and the 3' end of fMet-tRNAfMet is primarily responsible for the fMet-tRNAfMet binding and recognition by IF2. PMID- 10866836 TI - A virological perspective on the need for vaccination. AB - Superinfecton of chronic carriers of hepatitis B virus (HBV) or hepatitis C virus (HCV) with hepatitis A virus (HAV) is often associated with more severe liver disease than infection with HAV alone. Superinfection commonly causes markers of HBV and HCV replication to fall to significantly lower levels. The pathogenesis of acute liver damage characteristic of viral hepatitis is thought to be mediated by host cytotoxic T-lymphocytes (CTLs) directed against virus-infected hepatocytes. It has been proposed that the more aggressive liver disease observed in individuals infected with HAV in addition to chronic HBV/HCV is a result of the induction of interferon (IFN)-alpha during acute HAV infection. This accounts for the antiviral effect on the active markers of HBV/HCV replication, and the enhanced CTL response against HBV/HCV-infected hepatocytes. Alternatively, HAV may indirectly stimulate the T helper 1 (Th1)-type cytokine responses, such as interleukin (IL)-2, IFN-gamma and tumour necrosis factor (TNF)-alpha, which directly promote the antiviral CTL response. Clearance of HBV infection, and possible HAV and HCV, is associated with a specific CTL response, while viral persistence in chronic HBV and HCV infection has been attributed to an imbalance in the Th1-Th2 arms of the immune response. Vaccination against hepatitis A should be considered for patients with chronic HBV/HCV infection, to minimize the risk of exacerbating underlying liver disease. PMID- 10866837 TI - Fulminant hepatitis associated with hepatitis A virus superinfection in patients with chronic hepatitis C. AB - There have been conflicting reports of the clinical outcome of acute hepatitis A virus (HAV) infection in patients with chronic hepatitis C virus (HCV) infection. A prospective study evaluated 432 patients with chronic hepatitis C (183 with cirrhosis) over a 7-year period. Of the 17 patients with concurrent HAV infection, seven developed fulminant hepatitis and six died. None of these patients had cirrhosis; however, the HLA phenotype (A1; B8:DR3) appeared to be a significant factor in the development of fulminant hepatitis. Patients with this phenotype had high titres of antinuclear antibodies, antismooth muscle antibodies and antiasialoglycoprotein-receptor antibodies, possibly reflecting the induction of autoimmune hepatitis in this group. The high frequency of fulminant hepatitis in patients with HAV/HCV coinfection contrasts with other surveys, although a large Centers for Disease Control and Prevention (CDC) survey demonstrated that HAV infection in patients with pre-existing chronic liver disease (CLD) is associated with increased mortality. It is likely that CLD has some importance as an underlying factor in the development of fulminant hepatitis following HAV infection. Further prospective studies are needed to clarify this issue. PMID- 10866838 TI - Fulminant hepatitis in patients with chronic liver disease. AB - The changing epidemiology of hepatitis A virus (HAV) in the UK has led to a decline in natural immunity against the virus. It is estimated that in the UK, HAV is responsible for 10%-20% of cases of liver failure, and an overall mortality rate of 0.1%. It is clear that certain factors predispose patients to more severe HAV disease and increased mortality, although the reasons for this have yet to be elucidated. The age at which infection occurs clearly influences the outcome, with the risk of severe hepatitis increasing sharply after the age of 40 years. Intravenous drug users, homosexual men, individuals with an excessive alcohol intake or patients with chronic liver disease are also at increased risk of severe disease. An analysis of data from King's College Hospital was performed to determine the factors that influence the outcome or clinical course of HAV infection in at-risk patients. Data compiled from 1991 to 1998 revealed 187 cases with confirmed HAV, 45 of whom developed severe hepatitis. Outcomes were varied, eight (17.7%) patients developed acute liver failure and two (4.4%) died. PMID- 10866839 TI - Acute hepatitis A and acquired immunity to hepatitis A virus in hepatitis B virus (HBV) carriers and in HBV- or hepatitis C virus-related chronic liver diseases in Thailand. AB - A number of studies have suggested that the clinical course of hepatitis A virus (HAV) infection is more severe in patients with chronic liver disease (CLD). A study was undertaken to determine the impact of acute HAV in asymptomatic hepatitis B surface antigen (HBsAg) carriers (n = 20) and patients with hepatitis B virus (HBV)-(n = 8) or hepatitis C virus (HCV)-related (n = 4) CLD. Disease progression was compared with that in 100 patients with isolated HAV infection. No patient with HAV infection alone developed complications, and all recovered fully. Fulminant or submassive hepatitis occurred in 55% of HBsAg carriers and 33% of patients with HBV- or HCV-related CLD. The mortality rate in HBsAg carriers (25%) was not significantly different from that in the patients with CLD (33%). The seroprevalence of anti-HAV immunoglobulin G in 820 individuals was also determined. Approximately 50% of the individuals had acquired HAV infection between the ages of 21 and 30 years. It was demonstrated that HAV infection may have a more severe clinical course in patients with underlying CLD, particularly among older individuals. Vaccination for such patients should be considered. PMID- 10866840 TI - Immunogenetics of hepatitis C virus. AB - Hepatitis C is a complex disease and a significant public health problem. The long-term outcome of HCV infection is difficult to predict, as the virus may be eliminated or may persist to establish a chronic infection. Further investigation is required to clarify factors that determine or influence the outcome of infection, although unique host factors appear to significantly affect the prognosis for infected patients. PMID- 10866841 TI - Hepatitis A vaccination in patients with chronic liver disease in Taiwan. AB - While hepatitis A infection is rarely fatal, mortality is age-dependent, suggesting that cofactors may play a role in disease expression. The safety and immunogenicity of an inactivated hepatitis A vaccine was evaluated in a study involving 60 patients with chronic liver disease (CLD) (56 patients with chronic hepatitis B, and four patients with chronic hepatitis C). A two-dose schedule, at 0 and 6 months, produced 100% seroconversion. All individuals remained seropositive for 12 months, the majority maintaining high levels of antibody during a 5-year follow-up period. The inactivated hepatitis A vaccine was found to be well tolerated and immunogenic in patients with CLD. However, the immune response generated was inferior to that observed in healthy individuals, indicating the need for a higher dose, and the two-dose schedule to ensure adequate protection. PMID- 10866842 TI - Epidemiology of hepatitis A in Asia and experience with the HAV vaccine in Hong Kong. AB - Hepatitis A and hepatitis B infections are prevalent throughout Asia. Patients infected with hepatitis B virus (HBV) have a higher morbidity and mortality if superinfected with hepatitis A virus (HAV). In Hong Kong, adolescents and young adults are particularly prone to severe hepatitis infection. Hepatitis A vaccination of patients with or without chronic HBV infection has been shown to be efficacious and associated with a good safety profile. PMID- 10866843 TI - Consensus statement on the role of hepatitis A vaccination in patients with chronic liver disease. AB - Hepatitis A virus (HAV) is a ubiquitous, easily transmitted virus that can cause severe hepatitis, particularly in the adult population. Improvements in sanitation and hygiene in the developing world have led to a decline in immunity against HAV. A growing number of adults are now susceptible to infection, with those who have not been vaccinated against hepatitis B virus (HBV) being at risk of dual infection and potentially more severe illness. The symposium, "The role of hepatitis vaccination in patients with chronic liver disease", provided the opportunity to develop a consensus statement. It was concluded that patients with non-viral chronic liver disease (CLD) and certain groups with HBV infection can develop more severe disease in the event of HAV infection. It was identified that patients with CLD should be targeted for hepatitis A vaccination as soon as CLD is diagnosed. PMID- 10866845 TI - JAMA 100 years ago: SCHOOL HYGIENE PMID- 10866844 TI - A piece of my mind: the patient's heart. PMID- 10866846 TI - Optimizing treatment of hypertension in patients with diabetes. PMID- 10866847 TI - Women, heart disease, and stroke are focus of international meeting. PMID- 10866848 TI - First impaired physicians therapy program appears to be successful in Spain. PMID- 10866849 TI - Research advances target epilepsy. PMID- 10866850 TI - Quick uptakes: HIV screening in pregnancy PMID- 10866851 TI - Quick uptakes: Do ask, Do tell PMID- 10866853 TI - Quick uptakes: NGF implicated in allergy PMID- 10866852 TI - Quick uptakes: hispanic heart health PMID- 10866854 TI - Cardiovascular procedures in patients with mental disorders. PMID- 10866855 TI - Cardiovascular procedures in patients with mental disorders. PMID- 10866856 TI - Cardiovascular procedures in patients with mental disorders PMID- 10866857 TI - Handling conflict in end-of-life care. PMID- 10866858 TI - Handling conflict in end-of-life care. PMID- 10866859 TI - Handling conflict in end-of-life care. PMID- 10866860 TI - Handling conflict in end-of-life care PMID- 10866861 TI - Informed decisions for extremely low-birth-weight infants. PMID- 10866862 TI - Informed decisions for extremely low-birth-weight infants PMID- 10866863 TI - Screening mammography in elderly women. Research on Breast Cancer in Older Women Consortium. PMID- 10866864 TI - Screening mammography in elderly women. Research on Breast Cancer in Older Women Consortium. PMID- 10866865 TI - Screening mammography in elderly women. PMID- 10866866 TI - Screening mammography in elderly women PMID- 10866867 TI - HMG-CoA reductase inhibitors and the risk of fractures. AB - CONTEXT: Recent animal studies have suggested that 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) increase bone formation, volume, and density. It is unknown whether use of statins is associated with a decreased risk of fractures in humans. OBJECTIVE: To determine whether exposure to statins, fibrates, or other lipid-lowering drugs is associated with reduced bone fracture risk. DESIGN: Population-based, nested case-control analysis. SETTING: The UK-based General Practice Research Database (GPRD), comprising some 300 practices, with data collection from the late 1980s until September 1998. SUBJECTS: Within a base population of 91,611 individuals aged at least 50 years (28,340 individuals taking lipid-lowering drugs, 13,271 untreated individuals with a diagnosis of hyperlipidemia, and 50,000 randomly selected individuals without diagnosis of hyperlipidemia), we identified 3940 case patients who had a bone fracture and 23,379 control patients matched for age (+/-5 years), sex, general practice attended, calendar year, and years since enrollment in the GPRD. MAIN OUTCOME MEASURES: Use of statins, fibrates, or other lipid-lowering drugs in case patients vs control patients. RESULTS: After controlling for body mass index, smoking, number of physician visits, and corticosteroid and estrogen use, current use of statins was associated with a significantly reduced fracture risk (adjusted odds ratio [OR], 0.55; 95% confidence interval [CI], 0.44-0.69) compared with nonuse of lipid-lowering drugs. Current use of fibrates or other lipid-lowering drugs was not related to a significantly decreased bone fracture risk (adjusted OR, 0.87; 95% CI, 0.70-1.08 and adjusted OR, 0.76; 95% CI, 0.41 1.39, respectively). CONCLUSIONS: This study suggests that current exposure to statins is associated with a decreased risk of bone fractures in individuals age 50 years and older. This finding has a potentially important public health impact and should be confirmed further in controlled prospective trials. JAMA. 2000;283:3205-3210 PMID- 10866868 TI - HMG-CoA reductase inhibitors and the risk of hip fractures in elderly patients. AB - CONTEXT: Recent animal studies have found that 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) lipid-lowering drugs (statins) substantially increase bone formation, but whether statin use in humans results in clinically meaningful bone formation or a reduction in the risk of osteoporotic fractures is not known. OBJECTIVE: To determine whether the use of statins is associated with reduced hip fracture risk. DESIGN: Case-control study. SETTING AND PATIENTS: A total of 6110 New Jersey residents aged 65 years or older and enrolled in Medicare and either Medicaid or the Pharmacy Assistance for the Aged and Disabled program. Case patients (n=1222) underwent surgical repair of a hip fracture in 1994. Control patients (n=4888) were identified at a ratio of 4:1 and frequency-matched to case patients for age and sex. MAIN OUTCOME MEASURE: Adjusted odds ratio (OR) of hip fracture by statin use in the 180 days and 3 years prior to the index date (the earliest date of admission for surgery), adjusted for demographic and clinical characteristics and health care utilization. RESULTS: Use of statins in either the prior 180 days (adjusted OR, 0.50; 95% confidence interval [CI], 0.33-0.76) or prior 3 years (adjusted OR, 0.57; 95% CI, 0.40-0.82) was associated with a significant reduction in the risk of hip fracture, even after controlling for variables such as race, insurance status, psychoactive medications, estrogen and thiazide use, ischemic heart disease, cancer, and diabetes mellitus. No significant relationship was observed between use of nonstatin lipid-lowering agents and hip fracture risk. Clear relationships were observed between the degree of reduction in hip fracture risk and the extent of statin use; there was no evidence of such relationships with nonstatin lipid-lowering agents. After adjusting for extent of statin use in the prior 3 years, current use (on the index date) was associated with a 71% reduction in risk (adjusted OR, 0.29; 95% CI, 0.10-0.81). The relationship between statin use and hip fracture risk persisted after controlling for variables such as the number of medications, the Charlson comorbidity index score, and hospitalization or nursing home stay in the last 180 days, as well as after excluding patients who were in a nursing home prior to their index date or who died in the year after their index date. Use of nonstatin lipid-lowering agents was not observed to be associated with reduction in hip fracture risk in any of these alternative models or analyses. CONCLUSIONS: These findings support an association between statin use by elderly patients and reduction in the risk of hip fracture. Controlled trials are needed to exclude the possibility of unmeasured confounders. JAMA. 2000;283:3211-3216 PMID- 10866869 TI - Comparison of recommendations by urologists and radiation oncologists for treatment of clinically localized prostate cancer. AB - CONTEXT: Multiple treatment options are available for men with prostate cancer, but therapeutic recommendations may differ depending on the type of specialist they consult. OBJECTIVE: To define and contrast the distribution of management recommendations by urologists and radiation oncologists for a spectrum of men with prostate cancer. DESIGN, SETTING, AND PARTICIPANTS: Mail survey sent in 1998 to a random sample of physicians in the United States, who were listed as urologists (response rate 64%, n=504) and radiation oncologists (response rate 76%, n=559) in the American Medical Association Registry of Physicians and practicing at least 20 hours per week. MAIN OUTCOME MEASURE: Questionnaire addressing beliefs and practices regarding prostate cancer management. RESULTS: Forty-three percent of radiation oncologists vs 16% of urologists would recommend routine prostate-specific antigen testing for men aged 80 years and older. For men with moderately differentiated, clinically localized cancers, and a more than 10-year life expectancy, 93% of urologists chose radical prostatectomy as the preferred treatment option, while 72% of radiation oncologists believed surgery and external beam radiotherapy were equivalent treatments. For most tumor grades and prostate-specific antigen levels, both specialty groups were significantly more likely to recommend the treatment in their specialty than the other treatment. Both groups reported giving patients similar estimates of the risks of complications due to surgery and radiation. Neither group favored watchful waiting in their treatment management except for a subset of men with life expectancies of less than 10 years and cancers with very favorable prognoses (Gleason score of 3 or 4 and prostate-specific antigen level or =20 microM; P < 0. 05). Decreases in maximum rates of contraction and relaxation paralleled changes in developed tension. beta-Adrenergic stimulation reversed the etomidate-induced decreases in developed tension and rates of contraction and relaxation to baseline (P > 0.05 compared with baseline). Thus, in human myocardium, etomidate exerts a dose-dependent negative inotropic effect, which is reversible with beta-adrenergic stimulation. Concentrations required to produce these negative inotropic effects are, however, in excess of those reached during clinical use. Therefore, etomidate-induced negative inotropy is unlikely to be a problem clinically, even in patients with cardiac dysfunction. IMPLICATIONS: Etomidate produced a similar dose-dependent negative inotropic effect in both failing and nonfailing human myocardium. This effect was present only at concentrations exceeding those attained clinically and was reversible with beta-adrenergic stimulation. PMID- 10866890 TI - Dilation by isoflurane of preconstricted, very small arterioles from human right atrium is mediated in part by K(+)-ATP channel opening. AB - The adenosine triphosphate (ATP)-sensitive potassium channels (K(+)-ATP channels) are activated by decreases in intracellular ATP and help to match blood flow to tissue needs. Such metabolism-flow coupling occurs predominantly in the smallest arterioles measuring 50 microm or less in diameter. Previous studies demonstrated that isoflurane may activate the K(+)-ATP channels in larger arteries. We examined whether isoflurane also activates the channels in the smallest arterioles of approximately 50 microm. Microvessels of approximately 50 microm were dissected from right atrial appendages from patients undergoing coronary artery bypass surgery and were monitored in vitro for diameter changes by videomicroscopy. With or without preconstriction with the thromboxane analog U46619 1 microM, vessels were exposed to isoflurane 0%-3% either in the presence or absence of the K(+)-ATP channel blocker glibenclamide 1 microM. Without preconstriction, isoflurane neither dilated nor constricted the vessels significantly. After preconstriction, isoflurane had a concentration-dependent dilation of the small arterioles (39 +/- 13% [mean +/- SD] dilation at 3% isoflurane) (P < 0.001), and this effect was significantly attenuated by glibenclamide (18 +/- 5% dilation at 3% isoflurane) (P < 0.01). In comparison, nitroprusside 10(-4) M produced 79 +/- 6% dilation, and adenosine diphosphate 10( 4) M produced 29 +/- 7% dilation. We conclude that isoflurane-mediated dilation of the smallest resistance arterioles may be in part based on activation of the K(+)-ATP channels when the arterioles are relatively constricted. IMPLICATIONS: Vasodilation of very small coronary arterioles by isoflurane depends on preexisting tone and may in part be mediated by the K(+)-ATP channels. PMID- 10866891 TI - Bupivacaine suppresses [Ca(2+)](i) oscillations in neonatal rat cardiomyocytes with increased extracellular K+ and is reversed with increased extracellular Mg(2+). AB - Lidocaine is used to treat cardiac arrhythmias, whereas bupivacaine is noted for its cardiotoxicity. A precise mechanism for these differences is unclear, and there is no well defined antidote for local anesthetic cardiotoxicity. Our study compares the effect of lidocaine and bupivacaine on oscillations of intracellular Ca(2+) coupled with contractions in neonatal rat cardiomyocytes by using digital imaging. In medium containing 5.6 mM K(+), both 42 microM lidocaine and 5.5 microM bupivacaine significantly reduced the oscillation rate. The oscillatory patterns were highly irregular, and the rates were increased in the presence of bupivacaine in 7.6 mM K(+) medium, eventually degenerating into a loss of oscillations after several minutes of bupivacaine exposure. Irregular oscillations did not occur with lidocaine until the K(+) concentration was increased to 10 mM. Increasing the Mg(2+) and Ca(2+) concentrations by 2 mM each recovered oscillation that had been suppressed by bupivacaine in high K(+) buffer. Evaluation of intracellular Ca(2+) oscillations in neonatal rat suggests that increased extracellular K(+) may be an important component of bupivacaine cardiotoxicity. IMPLICATIONS: Evaluation of intracellular Ca(2+) oscillations in neonatal rat myocytes suggests that increased extracellular K(+) may be an important component of bupivacaine cardiotoxicity. PMID- 10866892 TI - Endoluminal abdominal aortic aneurysm repair complicated by intracardiac guidewire placement and massive transfusion. PMID- 10866893 TI - Fast-tracking after immersion lithotripsy: general anesthesia versus monitored anesthesia care. AB - Both monitored anesthesia care (MAC) and general anesthesia (GA) offer advantages over epidural anesthesia for immersion lithotripsy. We compared propofol-based MAC and desflurane-based GA techniques for outpatient lithotripsy. After receiving midazolam 2 mg IV, 100 subjects were randomly assigned to one of two anesthetic treatment groups. In the MAC group, propofol 50-100 microg. kg(-1). min(-1) IV was titrated to maintain an observer's assessment of alertness/sedation score of 2-3 (5 = awake/alert to 1 = asleep). Remifentanil 0.05 microg.kg(-1). min(-1) IV supplemented with 0.125 microg/kg IV boluses, was administered for pain control. In the GA group, anesthesia was induced with propofol 1.5 mg/kg IV and remifentanil 0.125 microg/kg IV and maintained with desflurane (2%-4% inspired) and nitrous oxide (60%). Tachypnea (respiratory rate >20 breaths/min) was treated with remifentanil 0.125 microg/kg IV boluses. In the GA group, droperidol (0.625 mg IV) was administered as a prophylactic antiemetic. Recovery times and postoperative side effects were assessed up to 24 h after the procedure. Compared with MAC, the use of GA reduced the opioid requirement and decreased movements and episodes of desaturation (<90%) during the procedure. Although the GA group took longer to return to an observer's assessment of alertness/sedation score of 5, discharge times were similar in both groups. We conclude that GA can provide better conditions for outpatient immersion lithotripsy than MAC sedation without delaying discharge. IMPLICATIONS: A desflurane-based general anesthetic technique using the cuffed oropharyngeal airway device was found to be a highly acceptable alternative to propofol-based monitored anesthesia care sedation for outpatient immersion lithotripsy. PMID- 10866894 TI - Assessing a tool to measure patient functional ability after outpatient surgery. AB - The "24-Hour Functional Ability Questionnaire" (24hFAQ) was developed to measure final recovery and satisfaction 24 h after surgery. We used structured interviews preoperatively to measure baseline patient concerns, and up to 24 h after discharge, to assess patient function and satisfaction. The primary objective was to assess the validity of the newly developed 24hFAQ in the postoperative outpatient setting. The criteria assessed were 1) CONTENT: comparison with expert opinion and patients' views and response frequency distributions for asymptotes and irrelevance, 2) Construct: contribution of cognitive, physical, and satisfaction domains to postoperative functional ability, 3) Discrimination: comparing mean clinical end points with patient satisfaction, and 4) Criterion (predictive) validity: testing that related constructs are best correlated. CONTENT validity was supported by the appropriate frequency distribution of subject responses, by the lack of floor or ceiling effects, and by <2% of responses indicating irrelevance. Construct validity was supported by moderate-to strong positive interitem correlations within the cognitive and physical domains as predicted a priori. Discriminant validity support was mixed: key symptoms were associated with adverse patient satisfaction, but operating room and postanesthesia care unit residence times were unrelated. Criterion validity was supported by the finding that preoperative concern with key symptoms was independent of postoperative outcomes. The validity assessment presented was the first assessment of the measurement capability of the 24hFAQ in an outpatient postoperative population. These results provide overall support for the validity of the 24hFAQ for use in outpatient populations. IMPLICATIONS: This study assessed the validity of a novel functional ability questionnaire that measured functional status after recovery from anesthesia and satisfaction 24 h after outpatient surgery. The content, construct, discriminant, and criterion (predictive) validities demonstrated the utility of this assessment instrument in the outpatient setting. PMID- 10866895 TI - Caruncle single injection episcleral (Sub-tenon) anesthesia for cataract surgery: mepivacaine versus a lidocaine-bupivacaine mixture. AB - We compared the quality of anesthesia provided by mepivacaine 2% or a mixture of lidocaine 2%-bupivacaine 0.5%, both with hyaluronidase, in caruncle single injection episcleral (sub-Tenon) anesthesia. Sixty patients undergoing cataract surgery were included in this randomized, double-blinded study. The time to the onset of blockade, maximal akinesia, need for supplemental injection, and time to recovery were recorded. With mepivacaine, the time to onset was slightly shorter, and the akinesia score higher, than with the mixture. Although statistically significant, these differences are small. With mepivacaine, the time to recovery was shorter. We conclude that the reproducible short duration of the block may be an advantage in outpatient surgery. IMPLICATIONS: We compared the classic mixture of lidocaine 2% plus bupivacaine 0.5% to mepivacaine 2% for caruncle episcleral (sub-Tenon) anesthesia for cataract surgery. Mepivacaine provided a more efficient block with a quicker onset and a quicker recovery. However, these differences were very small and were of little clinical interest. PMID- 10866896 TI - Acute tolerance to continuously infused alfentanil: the role of cholecystokinin and N-methyl-D-aspartate-nitric oxide systems. AB - To test the role of cholecystokinin (CCK) and N-methyl-D-aspartate-nitric oxide (NMDA-NO) systems in the development of acute tolerance to analgesia during alfentanil IV infusion, we conducted experiments in rats with the use of an infusion algorithm designed to maintain a constant plasma level of the opioid for 4 h. The degree of acute tolerance was determined on the basis of decline in the level of analgesia measured with a tail compression test. CCK(B) receptor antagonists (proglumide, CI-988, and L-365,260) and NMDA-NO cascade inhibitors (dizocilpine and NO synthase inhibitor) were administered before the start of alfentanil infusion. Use of 30 mg/kg proglumide, 10 mg/kg CI-988, and 1 mg/kg L 365,260 attenuated acute tolerance at 1 h of alfentanil infusion by approximately 60%, 55%, and 70%, respectively, and by the end of 4-h infusion by 50%, 50%, and 25%, respectively. Use of 0.1 mg/kg dizocilpine and 10 mg/kg N(G)-nitro-L arginine methyl ester attenuated acute tolerance at 1 h of alfentanil infusion by approximately 65% and 65% and by the end of 4-h infusion by 30% and 0%, respectively. Comparison of the results with CCK(B) receptor antagonists and inhibitors of NMDA-NO cascade demonstrates that both groups of drugs provide more or less similar degrees of attenuation of acute tolerance to the antinociceptive effect of alfentanil, and none of these drugs completely prevents tolerance development. IMPLICATIONS: The mechanism of acute tolerance to the analgesic effect of alfentanil depends on participation of multiple systems of adaptation that include cholecystokinin(B) receptors and N-methyl-D-aspartic acid-nitric oxide cascade. Drugs that inhibit function of these systems attenuate tolerance development. PMID- 10866897 TI - The recovery of cognitive function after remifentanil-nitrous oxide anesthesia is faster than after an isoflurane-nitrous oxide-fentanyl combination in elderly patients. AB - We tested the hypothesis that remifentanil-nitrous oxide (N(2)O) anesthesia shortens postoperative emergence and recovery compared with an isoflurane-N(2)O fentanyl combination in elderly patients undergoing spinal surgery. A total of 60 patients (>65 yr old) were randomly assigned to one of two groups for maintenance of anesthesia. After the induction with 3.6 +/- 1.2 mg/kg IV thiopental and endotracheal intubation facilitated with 1.4 +/- 0.5 mg/kg succinylcholine, patients were maintained with either 0.5%-1.5% isoflurane, 70% N(2)O, and up to 7 microg/kg fentanyl (iso/fent group) or 48 +/- 11 microg/kg remifentanil and 70% N(2)O (remi group). A mini-mental status examination was used to assess cognitive ability preoperatively, at 15, 30, and 60 min after arrival at the postanesthesia care unit and again 12-24 h postoperatively. The time from the conclusion of anesthesia to spontaneous respiration was similar in both groups. Times to eye opening (4.8 +/- 2.6 vs 2.3 +/- 1.1 min), extubation (6.8 +/- 3.8 vs 3.2 +/- 2.1 min), and verbalization (9.9 +/- 6.2 vs 3.9 +/- 2.6 min) were significantly shorter for the remi group (P < 0.05). Postoperative mini-mental status examination scores were significantly lower in the iso/fent group at 15 (16.3 +/- 5.8 vs 23. 7 +/- 3.3), 30 (20.2 +/- 5.2 vs 26.3 +/- 2.7), and 60 min (23.5 +/- 4.4 vs 27.5 +/- 2.0) (P < 0.001); however, the scores equalized after 12 h. Requirements for postoperative analgesics were similar in the two groups. More patients in the remi group were treated with antiemetics (21 vs 7, P = 0.06). Use of remifentanil-N(2)O for maintenance did not shorten the overall length of stay in the postanesthesia care unit; a stay is often related to multiple administrative issues, rather than cognitive recovery. IMPLICATIONS: Maintenance of anesthesia with remifentanil-nitrous oxide (N(2)O), compared with isoflurane N(2)O-fentanyl, can safely shorten postoperative recovery of cognitive function in a geriatric population. Earlier recovery may facilitate postoperative neurological assessment. Use of remifentanil-N(2)O for maintenance did not shorten the overall length of stay in the postanesthesia care unit, a stay often related to multiple administrative issues, rather than cognitive recovery. PMID- 10866898 TI - Recovery after anesthesia with remifentanil combined with propofol, desflurane, or sevoflurane for otorhinolaryngeal surgery. AB - Because no previous investigation has directly compared the combination of remifentanil (REM) and a hypnotic with that of REM and the newer volatile anesthetics, we studied recovery characteristics and patient satisfaction after the combination of REM with propofol (PRO), desflurane (DES), or sevoflurane (SEVO). One hundred twenty patients were randomly assigned to receive anesthesia with either REM/PRO, REM/DES, REM/SEVO, or thiopental/alfentanil/isoflurane/N(2)O (control group) for ear, nose, and throat surgery (n = 30 each). In the REM groups, the dosage of PRO (75 microg. kg(-1). min(-1)), and of DES or SEVO (0.5 minimum alveolar anesthetic concentration) was kept unchanged, and REM was titrated to hemodynamic response. The control group was managed according to standard practice. Early recovery (times to eye opening, extubation, and statement of name and date of birth) was predictably faster and more complete in the REM groups compared with the control group. However, late recovery (times to discharge from postanesthesia care unit and hospital) and overall patient satisfaction were not different among groups. No clinically relevant differences existed among the three REM groups. In conclusion, the combination of REM infusion with small-dose DES, SEVO, or PRO is characterized by predictably rapid, early recovery. However, late recovery and patient satisfaction are comparable to a conventional anesthetic technique. IMPLICATIONS: Remifentanil anesthesia, combined with small-dose propofol, desflurane, or sevoflurane, enables predictably fast and smooth early recovery after ear, nose, and throat surgery. Despite such faster, early recovery and less need for postoperative analgesic and antiemetic medication, late recovery was comparable among the remifentanil combination groups and the control group. PMID- 10866899 TI - Nitrous oxide prevents movement during orotracheal intubation without affecting BIS value. AB - We sought to determine whether the addition of nitrous oxide (N(2)O) to an anesthetic with propofol and remifentanil modifies the bispectral index (BIS) during the induction of anesthesia and orotracheal intubation. Thirty ASA physical status I or II patients were randomly allocated to receive either 50% air in oxygen (control group) or 60%-70% N(2)O in oxygen (N(2)O group) that was commenced via a mask simultaneously with the induction of anesthesia. Anesthesia was performed in all the patients with IV propofol at the target effect compartment site concentration of 4 microg/mL throughout the study. A target controlled infusion (TCI) of remifentanil was initiated 3 min after the TCI of propofol and maintained at the effect-site concentration of 4 ng/mL until the end of the study. After loss of consciousness, and before the administration of vecuronium 0.1 mg/kg, a tourniquet was applied to one arm and inflated to a value more than the systolic blood pressure. An examiner, blinded to the presence of N(2)O, sought to detect any gross movement within the first minute after tracheal intubation, which was performed 10 min after remifentanil TCI began. Inspired and expired oxygen, N(2)O, and carbon dioxide were continuously monitored. A BIS value was generated every 10 s. Arterial blood pressure and heart rate (HR) were measured noninvasively every minute. Measures of mean arterial pressure (MAP), HR, and BIS were obtained before the induction, before the start of the remifentanil TCI, before laryngoscopy, and 5 min after intubation. No significant intergroup differences were seen in BIS, HR, and MAP throughout the study. Maximum changes in BIS, HR, and MAP with intubation were significant (P < 0.01) for both groups but comparable. Six patients in the control group and none in the N(2)O group moved after intubation (P < 0.05). IMPLICATIONS: We demonstrated that 0.6 minimal alveolar concentration of nitrous oxide combined with a potent anesthetic and an opioid prevents movement after orotracheal intubation without affecting the bispectral index. This demonstrates that the bispectral index is not a useful neurophysiologic variable to monitor the level of anesthesia when nitrous oxide is added to a general anesthetic regimen using propofol and remifentanil. PMID- 10866900 TI - The effect of timing of dexamethasone administration on its efficacy as a prophylactic antiemetic for postoperative nausea and vomiting. AB - We evaluated the timing effect of a 10-mg IV administration of dexamethasone on its efficacy as a prophylactic antiemetic on postoperative nausea and vomiting (PONV). One hundred twenty women (n = 40 in each of three groups) undergoing abdominal total hysterectomy under general anesthesia were enrolled in this randomized, double-blinded, placebo-controlled study. Group 1 received dexamethasone before the induction of anesthesia, Group 2 received dexamethasone at the end of anesthesia, and Group 3 received placebo (saline). The incidence of PONV was evaluated. During the postoperative period of 0-2 h, patients in Group 1 reported a less frequent incidence of PONV (15%) than those in Groups 2 and 3 (45% and 53%, respectively). Patients in Group 1 also requested less rescue antiemetic (8%) than those in Groups 2 and 3 (30% and 35%, respectively). During the postoperative period of 2-24 h, patients in both Groups 1 and 2 reported less frequent incidences of PONV (25% and 28%) and requested fewer rescue antiemetics (13% and 15%) than those in Group 3 (55% and 38%, respectively). In conclusion, the prophylactic IV administration of dexamethasone immediately before the induction, rather than at the end of anesthesia, was more effective in preventing PONV. IMPLICATIONS: We evaluated the effect of timing of dexamethasone administration on its efficacy as a prophylactic antiemetic on postoperative nausea and vomiting. We found that dexamethasone, when given immediately before the induction of anesthesia, was more effective than when given at the end of anesthesia. PMID- 10866901 TI - Hypothermia attenuates the vasodilatory response of pial arterioles to hemorrhagic hypotension in the cat. AB - We investigated the effect of hypothermia on the vasodilatory response of pial arterioles to hemorrhagic hypotension. The cranial window technique was combined with microscopic video recording in an experiment involving 20 cats anesthetized with pentobarbital. The animals were randomly assigned to either a normothermic or a hypothermic group (32 degrees C). Mean arterial pressure (MAP) was reduced in stepwise increments of 10 mm Hg (from 100 to 50 mm Hg) by blood withdrawal. The diameter of small (50-100 microm) and large (100-200 microm) pial arterioles was measured. In the normothermic group (n = 9), small and large arterioles dilated at a MAP of 60 and 50 mm Hg, and at a MAP of 70, 60, and 50 mm Hg, respectively, compared with baseline values obtained at a MAP of 100 mm Hg. In contrast, in the hypothermic group (n = 11), vasodilation of either small or large arterioles was absent. The percentage diameter of small and large arterioles (percentage of control) was significantly lower at a MAP of 70, 60, and 50 mm Hg in the hypothermic group than the normothermic group. Our in vivo study demonstrates that hypothermia impairs autoregulatory vasodilation of pial arterioles in response to hemorrhagic hypotension. IMPLICATIONS: Deliberate mild hypothermia has been proposed as a means of providing cerebral protection during neurosurgical procedures. Our results suggest that cerebral blood flow autoregulation in response to hemorrhagic hypotension may be impaired during hypothermic conditions, indicating the importance of maintaining perfusion pressure during hypothermic therapy to prevent cerebral ischemia. PMID- 10866902 TI - The effects of anesthetics on stress responses to forebrain ischemia and reperfusion in the rat. AB - Rats exposed to forebrain ischemia have reduced injury when anesthetized with isoflurane versus fentanyl + N(2)O. The protection caused by isoflurane is reversed by trimethaphan. We hypothesized that these anesthetic-dependent effects on ischemic outcome can be associated with altered stress responses to ischemia. Rats were randomized to four treatments: isoflurane; fentanyl + N(2)O; isoflurane + trimethaphan; or isoflurane + metyrapone. Severe forebrain ischemia was then induced for 10 min. Plasma and brain corticosterone, tumor necrosis factor (TNF) alpha, and interleukin (IL)-6 were assayed. Plasma corticosterone concentrations were similar in the isoflurane and isoflurane + trimethaphan groups, but greater than in the fentanyl + N(2)O and isoflurane + metyrapone groups. Brain corticosterone was similar among all groups except isoflurane + metyrapone, in which values were markedly reduced. The addition of metyrapone to isoflurane also reduced plasma TNF-alpha; however, values among other groups were similar. There were no differences among groups for brain TNF-alpha. Plasma IL-6 concentrations were below the limit of detection. Brain IL-6 concentrations were increased by ischemia; however, there was no difference among groups. In conclusion, there were no differences between the isoflurane and isoflurane + trimethaphan groups for any of the measured stress markers. Further, there was little difference between the isoflurane and fentanyl + N(2)O groups, except for plasma corticosterone concentration. Accordingly, isoflurane neuroprotection and its reversal by trimethaphan appear to be independent of effects on the stress responses measured in this study. IMPLICATIONS: Differential anesthetic effects on ischemic outcome are independent of effects on adrenergic/noradrenergic responses to ischemia. The absence of a consistent differential effect of anesthetics on either corticosterone or cytokine responses to ischemia serves to further refute the hypothesis that isoflurane neuroprotection can be attributed to dampening of adverse stress responses to ischemic insults. PMID- 10866903 TI - Cerebral hemodynamic response to the introduction of desflurane: A comparison with sevoflurane. AB - Rapid increases in the inspired concentration of desflurane cause transient increases in heart rate and blood pressure. Desflurane also impairs cerebral autoregulation at clinical concentrations. Sevoflurane does not share these hemodynamic side effects. We compared the cerebral and systemic hemodynamic responses to the introduction of desflurane or sevoflurane after the induction of anesthesia with propofol. Twenty healthy adult patients scheduled for nonneurological surgery were recruited. After the induction of anesthesia with propofol, either desflurane or sevoflurane (n = 10 per group) was introduced at 7.2% or 2.2%, respectively, and increased to 10.8% or 3.3%, respectively, 2 min later. Middle cerebral artery blood flow velocity was measured continuously by using a 2-MHz transcranial Doppler ultrasound probe. Heart rate and blood pressure were recorded at 1-min intervals during the 12-min study period. Those patients receiving desflurane had significantly greater middle cerebral artery blood flow velocities, heart rates, and blood pressures than those receiving sevoflurane (P < 0.01). IMPLICATIONS: The introduction of desflurane after the induction of anesthesia leads to significant disturbances in cerebral and systemic hemodynamics suggesting loss of cerebral autoregulation and cerebral hyperemia. This may have implications for patients undergoing anesthesia for intracranial surgery. PMID- 10866904 TI - A subanesthetic concentration of sevoflurane increases regional cerebral blood flow and regional cerebral blood volume and decreases regional mean transit time and regional cerebrovascular resistance in volunteers. AB - Inhaled anesthetics exert metabolically mediated effects on cerebral blood vessels both directly and indirectly. We investigated the effects of a 0.4 minimum alveolar subanesthetic concentration of sevoflurane on regional cerebral blood flow (rCBF), regional cerebral blood volume (rCBV), regional cerebrovascular resistance (rCVR), and regional mean transit time (rMTT) in volunteers by means of contrast-enhanced magnetic resonance imaging perfusion measurement. Sevoflurane increased rCBF by 16% to 55% (control, 55. 03 +/- 0.33 to 148.83 +/- 1.9 mL. 100 g(-1). min(-1); sevoflurane, 71.75 +/- 0.36 to 193.26 +/- 2.14 mL. 100 g(-1). min(-1)) and rCBV by 7% to 39% (control, 4.66 +/- 0.03 to 10.04 +/- 0.12 mL/100 g; sevoflurane, 5.04 +/- 0.03 to 13.6 +/- 0.15 mL/100 g); however, sevoflurane decreased rMTT by 7% to 18% (control, 3.75 +/- 0.04 to 5. 39 +/- 0.04 s; sevoflurane, 3.4 +/- 0.03 to 4.44 +/- 0.03 s) and rCVR by 22% to 36% (control, 0.74 +/- 0.01 to 1.9 +/- 0.2 mm Hg/[mL. 100 g(-1). min(-1)]; sevoflurane, 0.54 +/- 0.01 to 1.41 +/- 0.01 mm Hg/[mL. 100 g(-1). min(-1)]). Interhemispheric differences in rCBF, rCBV, and rCVR were markedly reduced after the administration of sevoflurane. These findings are consistent with the known direct vasodilating effect of sevoflurane. The decrease in rMTT further shows that rCBF increases more than does rCBV. Furthermore, we can show that the observed increase in rCBF during inhalation of sevoflurane is not explained by vasodilation alone. PMID- 10866905 TI - A comparison of remifentanil and fentanyl in patients undergoing surgery for intracranial mass lesions. AB - We compared the effects of remifentanil versus fentanyl during surgery for intracranial space-occupying lesions. Patients were randomly assigned to receive either remifentanil (0.5 microg. kg(-1). min(-1) IV during the induction of anesthesia reduced to 0.25 microg. kg(-1). min(-1) after endotracheal intubation; n = 49) or fentanyl (dose per usual practice of the anesthesiologist; n = 54). Anesthesia maintenance doses of isoflurane, nitrous oxide, and opioid were at the anesthesiologist's discretion for both groups. There were no differences between opioid groups for the frequency of responses (hemodynamic, movement, and tearing) to intubation, pinhead holder placement, skin incision, or closure of the surgical wound. Adverse event frequencies were similar between groups. Times to follow verbal commands (P < 0.001) and tracheal extubation (P = 0. 04) were more rapid for remifentanil. The percentage of patients with a normal recovery score (were alert or arousable to quiet voice, were oriented, were able to follow commands, had motor function unchanged from their preoperative evaluation, were not agitated, and had modified Aldrete Scores of 9-10) at 10 min after surgery was more for remifentanil (45% vs 18%; P = 0.005). By 20 min, no difference between groups existed (P = 0.27). Anesthesiologists used more isoflurane in the fentanyl group (4.22 vs 1.93 minimum alveolar anesthetic concentration hours). Neurosurgeons, blinded to treatment group, favored the use of remifentanil. Similar frequencies of light anesthesia responses and other adverse events suggest that intraoperative depths of anesthesia were similar in the two groups. Under these conditions, emergence was more rapid with remifentanil. This is consistent with the necessity for less isoflurane use in the remifentanil group and the intrinsic rapid clearance of this opioid. IMPLICATIONS: Patients given remifentanil-based anesthesia for craniotomy had faster recovery times from anesthesia than did those given fentanyl-based anesthesia. PMID- 10866906 TI - The use of a tracheostomy tube for enteral stomal control. AB - We found that the use of a zero-pressure tracheal foam cuff was the ideal way to drain the intestines through a colostomy, reducing skin irritations and mucosal damage. PMID- 10866908 TI - Extended "three-in-one" block after total knee arthroplasty: continuous versus patient-controlled techniques. AB - This prospective, randomized, double-blinded study assessed the efficacy of patient-controlled analgesia (PCA) techniques for extended "3-in-1" block after total knee arthroplasty. A total of 45 patients were divided into three groups of 15. Over 48 h, all patients received 0.125% bupivacaine with 1 microg/mL clonidine via a femoral nerve sheath catheter in the following manner: as a continuous infusion at 10 mL/h in Group 1; as a continuous infusion at 5 mL/h plus PCA boluses (2.5 mL/30 min) in Group 2; or as PCA boluses only (10 mL/60 min) in Group 3. Pain scores, sensory block, supplemental analgesia, bupivacaine consumption, side effects, and satisfaction scores were recorded. Pain scores and supplemental analgesia were comparable in the three groups. Bupivacaine consumption was significantly less in Groups 2 and 3 than in Group 1 (P < 0.01), and in Group 3 than in Group 2 (P < 0.01). Side effects and satisfaction were comparable in the three groups. We conclude that extended "3-in-1" block provides efficient pain relief after total knee arthroplasty and that, compared with a continuous infusion, PCA techniques reduce the local anesthetic consumption without compromise in patient satisfaction or visual analog scale scores. Of the two PCA techniques tested, PCA boluses (10-mL lockout; time, 60 min) of 0.125% bupivacaine with 1 microg/mL clonidine was associated with the smallest local anesthetic consumption, and is, therefore, the recommended extended "3-in-1" block technique. IMPLICATIONS: We demonstrated that, after total knee arthroplasty, an extended "3-in-1" block consisting of patient-controlled analgesia boluses (10 mL/60 min) of 0.125% bupivacaine with 1 microg/mL clonidine provides efficient postoperative analgesia and significantly minimizes local anesthetic consumption. PMID- 10866907 TI - Prophylactic intravenous ondansetron reduces the incidence of intrathecal morphine-induced pruritus in patients undergoing cesarean delivery. AB - Pruritus is a common side effect of intrathecal morphine injection for postoperative pain control. Its incidence is especially high in patients undergoing cesarean delivery. We investigated the effectiveness of ondansetron in preventing intrathecal morphine-induced pruritus in such patients. We included 60 consecutive nonbreastfeeding women who were scheduled for elective cesarean delivery. After the administration of spinal anesthesia with bupivacaine and intrathecal morphine 0.15 mg injection, the patients were randomly divided into three groups. Group 1 received placebo (normal saline) IV injection, Group 2 diphenhydramine 30 mg IV injection, and Group 3 ondansetron 0.1 mg/kg IV injection. The incidence of pruritus was significantly lower in the ondansetron group (25%) when compared with that in the placebo group (85%) and in the diphenhydramine group (80%) (both P < 0.05). The postoperative pain score and time to flatus passage were not significantly different among the three groups. There were no headache or extrapyramidal signs associated with ondansetron use. In conclusion, ondansetron prophylaxis significantly reduced the incidence of intrathecal morphine-induced pruritus in patients undergoing cesarean delivery. IMPLICATIONS: Ondansetron prophylaxis significantly decreases the incidence of pruritus, a common side effect of intrathecal morphine used to treat postcesarean delivery pain. PMID- 10866909 TI - The effects of the single or multiple injection technique on the onset time of femoral nerve blocks with 0.75% ropivacaine. AB - We evaluated the effect of the injection technique on the onset time and efficacy of femoral nerve block performed with 0.75% ropivacaine. A total of 30 patients undergoing arthroscopic knee surgery were randomly allocated to receive femoral nerve blockade with 0.75% ropivacaine by using either a single injection (Single group, n = 15) or multiple injection (Multiple group, n = 15). Nerve blocks were placed by using a short-beveled, Teflon-coated, stimulating needle. The stimulation frequency was set at 2 Hz, and the intensity of stimulating current, initially set at 1 mA, was gradually decreased to <0.5 mA after each muscular twitch was observed. In the Single group, 12 mL of 0.75% ropivacaine was slowly injected, as soon as the first muscular twitch was observed. In the Multiple group, the stimulating needle was inserted and redirected, eliciting each of the following muscular twitches: contraction of vastus medialis, vastus intermedius, and vastus lateralis. At each muscular twitch, 4 mL of the study solution was injected. Placing the block required 4.2 +/- 1.7 min (median, 5 min; range, 2-8 min) in the Multiple group and 3.4 +/- 2.2 min (median, 3 min; range, 1-5 min) in the Single group (P = 0.02). Onset of nerve block (complete loss of pinprick sensation in the femoral nerve distribution with concomitant inability to elevate the leg from the operating table with the hip flexed) required 10 +/- 3.7 min in the Multiple group (median, 10 min; range, 5-20 min) and 30 +/- 11 min in the Single group (median, 30 min; range, 10-50 min) (P < 0.0005). Propofol sedation was never required to complete surgery; although 0.1 mg fentanyl at trocar insertion was required in two patients of the Multiple group (13%) and nine patients of the Single group (60%) (P = 0.02). We conclude that searching for multiple muscular twitches shortened the onset time and improved the quality of femoral nerve block performed with small volumes of 0.75% ropivacaine. IMPLICATIONS: This prospective, randomized, blinded study was conducted to evaluate the effect of searching for multiple muscular twitches when performing femoral nerve block with small volumes of 0. 75% ropivacaine. Our results demonstrated that multiple injections markedly shortened the onset time and improved the quality of nerve blockade. This technique-related effect must be carefully considered when different clinical studies evaluating the use of new local anesthetic solutions for peripheral nerve blocks are compared. PMID- 10866910 TI - Gabapentin enhances the analgesic effect of morphine in healthy volunteers. AB - The most effective group of drugs for the treatment of severe pain is opioid analgesics. Their use, however, is limited by decreased effects in neuropathic and chronic pain as a result of increased pain and development of tolerance. Gabapentin (GBP) is effective in both experimental models of chronic pain and clinical studies of neuropathic pain. Therefore, we investigated, in a randomized, placebo-controlled, double-blinded study, the pharmacodynamic and pharmacokinetic interaction of GBP and morphine in 12 healthy male volunteers. Morphine (60 mg, controlled release) or placebo was administered at 8:00 AM, and GBP (600 mg) or placebo was administered at 10:00 AM, thus comparing the analgesic effect of placebo + GBP (600 mg) with placebo + placebo and morphine (60 mg) + GBP in comparison to morphine plus placebo by using the cold pressor test. The duration and intensity of the side effects were assessed by using visual analog scales. The analgesic effect was evaluated by the change in the area under the curve (h x %; 0% baseline before Medication 1) of pain tolerance. Placebo + GBP (18.9% x h, 95% confidence interval [CI]: -2.5 to 40.3) did not present any significant analgesic effect compared with placebo + placebo (4.7% x h, 95% CI: -16.7 to 26.1). A significant increase in pain tolerance was observed comparing the combination of morphine and GBP (75.5% x h, 95% CI: 54.0-96.9) with morphine + placebo (40.6% x h, 95% CI: 19. 2-62.0). The observed adverse events after placebo + GBP were not significantly different compared with placebo + placebo. Morphine + placebo led to the expected opioid-mediated side effects. They were significantly more pronounced compared with placebo + placebo but did not differ significantly compared with the combination of morphine + GBP. Concerning the pharmacokinetic variables of morphine and its glucuronides, no significant difference between morphine + placebo and morphine + GBP was observed, whereas the area under the curve of GBP (43.9 +/- 5.3 vs 63.4 +/- 16.2 microg. h(-1). mL(-1), P < 0.05) significantly increased, and apparent oral clearance (230.8 +/- 29.4 mL/min vs 178 +/- 97.9 mL/min, P = 0.06) and apparent renal clearance (86.9 +/- 20.6 vs 73.0 +/- 24.2 mL/min, P = 0.067) of GBP decreased when morphine was administered concomitantly. These results suggest two different sites for the pharmacokinetic interaction-one at the level of absorption and the other at the level of elimination. Our study reveals both a pharmacodynamic and pharmacokinetic interaction between morphine and GBP, leading to an increased analgesic effect of morphine + GBP. These results and the good tolerability of GBP should favor clinical trials investigating the clinical relevance of the combination of morphine and GBP for treating severe pain. IMPLICATIONS: In a randomized, placebo-controlled, double-blinded trial with 12 healthy volunteers, we studied the interaction of morphine and gabapentin using the cold pressor test. The anticonvulsant gabapentin enhanced the acute analgesic effect of morphine. Furthermore, the plasma concentration of gabapentin was increased when morphine was administered concomitantly. Therefore, the well tolerated combination of gabapentin and morphine may improve pain therapy, especially in pain states, like chronic and neuropathic pain, which respond poorly to opioids. PMID- 10866911 TI - Adsorption of lidocaine into a plastic infusion balloon. AB - An infusion balloon is a well established device used to continuously supply drugs for pain management. In this study, we determined the concentration of lidocaine that flowed out of a balloon because the balloon is made from plastics that adsorb local anesthetics. PMID- 10866912 TI - Segmental cervical spine movement with the intubating laryngeal mask during manual in-line stabilization in patients with cervical pathology undergoing cervical spine surgery. AB - We quantified the extent and distribution of segmental cervical movement produced by the intubating laryngeal mask (ILM) during manual in-line stabilization in 20 anesthetized patients with cervical pathology undergoing cervical spine surgery. All patients had neurological symptoms preoperatively. The ILM was inserted with the head and neck in the neutral position. Intubation was facilitated by transillumination of the neck with a lightwand. Cervical movement was recorded with single-frame lateral radiographic images taken 1) immediately before induction (baseline); 2) during ILM insertion (insertion); 3) when transillumination was first seen at the cricothyroid membrane (intubation A); 4) when the tube was being advanced into the trachea (intubation B); and 5) during ILM removal (removal). Radiographic images were digitized and the degree of flexion/extension and posterior movement measured for the occiput (C0) through to C5. During ILM insertion, C0-5 were flexed by an average of 1-1.6 degrees (all P < 0.05). During intubation A/B, C0-4 were flexed by an average of 1.4-3.0 degrees (all P < 0.01), but C5 was unchanged. During ILM removal, C0-3 were flexed by an average of 1 degree (all: P < 0.05), but C3-5 were unchanged. During insertion and intubation A/B, C2-5 were displaced posteriorly by an average of 0.5-1.0 mm (all: P < 0.05). During removal, there was no change at C1-5. Neurological symptoms improved in all patients. We conclude that the ILM produces segmental movement of the cervical spine despite manual in-line stabilization in patients with cervical spine pathology undergoing cervical spine surgery. This motion is in the opposite direction to direct laryngoscopy, suggesting that different approaches to airway management may be more appropriate depending on the nature of the cervical instability. IMPLICATIONS: The intubating laryngeal mask produces segmental movement of the cervical spine, despite manual in-line stabilization in patients with cervical spine pathology undergoing cervical spine surgery. This motion is in the opposite direction to direct laryngoscopy, suggesting that different approaches to airway management may be more appropriate depending on the nature of the cervical instability. PMID- 10866913 TI - The effects of intracuff lidocaine on endotracheal-tube-induced emergence phenomena after general anesthesia. AB - Coughing during emergence from general anesthesia is a common clinical problem. We sought to determine whether inflating the endotracheal tube cuff with lidocaine would create a reservoir of local anesthetic, which might diffuse across the cuff membrane to anesthetize the mucosa, thus attenuating stimulation during extubation of the trachea. A total of 63 patients undergoing elective surgery were enrolled in a prospective, randomized, double-blinded study. After intubation of the trachea with an endotracheal tube, the cuff of the tube was inflated with either lidocaine 4%, saline, or air. After extubation, a blinded observer noted heart rate, blood pressure, oxygen saturation, end-tidal isoflurane concentration, and the incidence of coughing. Data were analyzed by using analysis of variance, Student's t-test, and the chi(2) test for multiple variables. The groups were demographically comparable. There was no difference in hemodynamic or oxygen saturation data between either group. The incidence of coughing was decreased in the lidocaine group for the time period of 4-8 min postextubation (P < 0.05). We conclude that inflation of the cuff of the endotracheal tube can reduce the incidence of coughing in the initial postextubation period, a finding that may benefit certain patient groups in which this is particularly desirable. IMPLICATIONS: Tracheal intubation with an endotracheal tube is often necessary during anesthesia. After intubation, inflating a cuff around the endotracheal tube maintains a seal. This can result in coughing during emergence from anesthesia. Our study shows that inflating the cuff of an endotracheal tube with lidocaine rather than air can reduce the incidence of postextubation coughing. PMID- 10866914 TI - Low-flow desflurane and sevoflurane anesthesia minimally affect hepatic integrity and function in elderly patients. AB - Hepatic blood flow is reduced in a dose-related manner by all inhaled anesthetics now in use. We assessed hepatic function in elderly patients anesthetized with desflurane or sevoflurane. We measured the cytosolic liver enzyme alpha glutathione S-transferase (alpha GST), the formation of the lidocaine metabolite monoethylglycinexylidide (MEGX), and gastric mucosal tonometry-derived variables as sensitive markers of hepatic function and splanchnic perfusion. Thirty patients, 70 to 90 yr old, were allocated randomly to receive desflurane or sevoflurane anesthesia. Anesthetic exposure ranged from 2.1-4.5 minimum alveolar concentration hours. No significant changes in standard liver enzyme markers were seen throughout the study. In both anesthetic groups, tonometric measurements showed a significant decrease from baseline in regional PCO(2), regional to arterial difference in PCO(2), and intramucosal pH at 90 min after skin incision. alpha GST concentrations increased significantly in both groups (desflurane: median peak concentrations 5.8 microg/L [25th, 75th percentile 5.3 microg/L, 7.2 microg/L]; sevoflurane: 7.0 microg/L [5.8 microg/L, 7.3 microg/L]) without showing differences between both anesthetic groups. A return to baseline values in tonometric values and alpha GST levels was seen 24 h postoperatively. MEGX formation did not change significantly after surgery. Median MEGX concentrations postoperatively were 70.0 ng/mL (56.2 ng/mL, 102.0 ng/mL) and 70.0 ng/mL (60.0 ng/mL, 94.2 ng/mL) in the desflurane and sevoflurane groups, respectively. We conclude that, overall, liver function in elderly patients is well preserved during desflurane and sevoflurane anesthesia. Increased serum levels of alpha GST and changes of gastric tonometry-derived variables imply a reduction in splanchnic perfusion, leading to a temporary impairment of hepatocyte oxygenation. IMPLICATIONS: We measured the lidocaine metabolite monoethylglycinexylidide, the cytosolic liver enzyme, alpha glutathione S transferase, and gastric mucosal tonometry-derived variables to evaluate the effects of desflurane and sevoflurane on hepatic function in elderly patients. Liver function was well preserved, whereas increased alpha glutathione S transferase levels and changes in tonometry-derived variables indicated a reduction in splanchnic blood flow and a temporary impairment of hepatocyte oxygenation for both anesthetics. PMID- 10866915 TI - Sevoflurane versus propofol for anesthetic induction: a meta-analysis. AB - We performed this meta-analysis to compare the characteristics of sevoflurane and propofol for the induction of routine anesthesia and for laryngeal mask airway (LMA) insertion. The variables assessed were 1) time to loss of consciousness, 2) incidence of apnea during induction, 3) induction complications, 4) time for successful LMA insertion, 5) success with LMA insertion on first attempt, 6) patient dissatisfaction, and 7) postoperative nausea and vomiting. MEDLINE, Embase, and the Cochrane library databases between January 1992 and October 1999 were reviewed for randomized, controlled trials comparing anesthetic induction between sevoflurane/nitrous oxide and propofol. Data from the 12 randomized, controlled studies were used for the meta-analysis. Sevoflurane induction was associated with a trend toward higher patient dissatisfaction and higher first time success with LMA. Apnea was less common in the sevoflurane group. The incidence of postoperative nausea and vomiting was significantly more frequent in the sevoflurane group (P < 0.05). This effect was still present when all other variables, except the induction methods, were controlled. The other pooled variables did not show a significant difference between sevoflurane and propofol. Sevoflurane and propofol had similar efficacy for anesthetic induction. However, for routine outpatient surgery, propofol may still be the preferred induction anesthetic because of its favorable induction of anesthesia characteristics, high patient satisfaction, and less frequent incidence of postoperative nausea and vomiting. IMPLICATIONS: Sevoflurane and propofol had similar efficacy for anesthetic induction. However, for routine outpatient surgery, propofol may still be the preferred induction anesthetic because of its favorable induction of anesthesia characteristics, high patient satisfaction, and less frequent incidence of postoperative nausea and vomiting. PMID- 10866916 TI - Carbon dioxide absorbents containing potassium hydroxide produce much larger concentrations of compound A from sevoflurane in clinical practice. AB - We investigated the concentrations of degraded sevoflurane Compound A during low flow anesthesia with four carbon dioxide (CO(2)) absorbents. The concentrations of Compound A, obtained from the inspiratory limb of the circle system, were measured by using a gas chromatograph. In the groups administered 2 L/min fresh gas flow with 1% sevoflurane, when the conventional CO(2) absorbents, Wakolime(TM) (Wako, Tokyo, Japan) and Dragersorb(TM) (Drager, Lubeck, Germany), were used, the concentrations of Compound A increased steadily from a baseline to 14.3 ppm (mean) and 13.2 ppm, respectively, at 2 h after exposure to sevoflurane. In contrast, when the other novel types of absorbents containing decreased or no potassium hydroxide/sodium hydroxide, Medisorb(TM) (Datex-Ohmeda, Louisville, CO) and Amsorb(TM) (Armstrong, Coleraine, Northern Ireland), were used, Compound A remained at baseline (<2 ppm) throughout the study. In the groups administered 1 L/min fresh gas flow with 2% sevoflurane, Wakolime(TM) and Dragersorb(TM) produced much larger concentrations of Compound A (35.4 ppm and 34.2 ppm, respectively) at 2 h after exposure to sevoflurane. Medisorb(TM) showed measurable concentrations of Compound A (8.6 ppm at 2 h), but they were significantly smaller than those produced by the two conventional absorbents. In contrast, when Amsorb(TM) was used, Compound A concentrations remained at baseline throughout the study period. IMPLICATIONS: Carbon dioxide absorbents containing potassium hydroxide/sodium hydroxide produce much larger concentrations of Compound A from sevoflurane in clinical practice. An absorbent containing neither potassium hydroxide nor sodium hydroxide produces the smallest concentrations of Compound A. PMID- 10866917 TI - The stereoselective effects of ketamine isomers on heteromeric N-methyl-D aspartate receptor channels. AB - The effects of S(+)- and R(-)-ketamine on heteromeric N-methyl-D-aspartate receptor channels were investigated on the epsilon1/zeta1, epsilon2/zeta1, epsilon3/zeta1, and epsilon4/zeta1 channels expressed in Xenopus oocytes. S(+) ketamine inhibited all four epsilon/zeta channels more effectively than R(-) ketamine. The inhibitor concentrations for half-control response for S(+) ketamine were quite similar among the four channels with 0.44-0.56 microM. However, the inhibitor concentrations for half-control response for R(-)-ketamine varied slightly among the four channels with 1.0 microM for epsilon2/zeta1 and epsilon3/zeta1 channels and 1.9-2.0 microM for epsilon1/zeta1 and epsilon4/zeta1 channels. Thus, the potency ratio of S(+)- and R(-)-ketamine for heteromeric channels was only slightly different among the epsilon/zeta channels. IMPLICATIONS: The potency order and ratio of ketamine isomers for inhibition of N methyl-D-aspartate receptor channels may not be so different between the brain region and the developmental stage. PMID- 10866918 TI - Evidence for GABA(A) receptor agonistic properties of ketamine: convulsive and anesthetic behavioral models in mice. AB - We examined the potentiation by ketamine of the gamma-aminobutyric acid(A) (GABA(A)) receptor function using convulsive and anesthetic behavioral models in adult male ddY mice. General anesthetic potencies were evaluated by a rating scale, which provided the data for anesthetic scores, loss of righting reflex, duration, and recovery time. All drugs were administered intraperitoneally. Small subanesthetic doses of ketamine did inhibit tonic seizures induced by a large dose of the GABA(A) receptor antagonist bicuculline (8 mg/kg). The 50% effective dose value was 15 (95% confidence limits 10-22) mg/kg. Even large anesthetic doses (100-150 mg/kg) did not suppress clonic seizures in 50% of the animals. The GABA(A) receptor agonist, muscimol (0.32-1.12 mg/kg), potentiated ketamine induced anesthesia in a dose-dependent fashion (P < 0.05). Similarly, the benzodiazepine receptor agonist, diazepam (1-3 mg/kg), augmented ketamine anesthesia in a dose-dependent manner (P < 0.05). Bicuculline (2-5 mg/kg) dose dependently antagonized ketamine-induced anesthesia (P < 0.05). Neither the benzodiazepine receptor antagonist, flumazenil (2-20 mg/kg), nor the GABA synthesis inhibitor, L-allylglycine (200 mg/kg), affected the anesthetic action of ketamine. These results suggest that ketamine has GABA(A) receptor agonistic properties and that ketamine-induced anesthesia is mediated, at least in part, by GABA(A) receptors. IMPLICATIONS: We examined the potentiation by ketamine of the gamma-aminobutyric acid(A) receptor function using convulsive and anesthetic behavioral models in mice. Subanesthetic doses of ketamine-inhibited tonic convulsions induced by the gamma-aminobutyric acid(A) receptor antagonist bicuculline. The gamma-aminobutyric acid(A) receptor agonist, muscimol, potentiated ketamine-induced anesthesia. Bicuculline antagonized ketamine anesthesia, but the benzodiazepine receptor antagonist, flumazenil, and the gamma aminobutyric acid synthesis inhibitor, L-allyglycine, did not. The effects of ketamine on the gamma-aminobutyric acid(A) receptors appear to correlate with its anesthetic actions. PMID- 10866919 TI - Succinylcholine-induced hyperkalemia in patients with renal failure: an old question revisited. PMID- 10866920 TI - Nitrogen purging of oxygen pipelines: an unusual cause of intraoperative hypoxia. AB - Intraoperative hypoxia occurred in two patients during the maintenance of the medical gas system. Engineers were purging oxygen pipelines with nitrogen to remove particulate debris but were unaware of a connection to operating room pipelines. This case illustrates the importance of communication between anesthesia providers and engineers servicing the gas system. PMID- 10866922 TI - Is tranexamic acid indicated for total knee replacement surgery? PMID- 10866923 TI - Rattling of unidirectional valves as a sign of secretions. PMID- 10866924 TI - Sevoflurane for patients with asthma. PMID- 10866925 TI - Global cerebral hypoperfusion and PaCO(2) PMID- 10866926 TI - Is awake intubation necessary when the laryngeal mask airway is feasible? PMID- 10866927 TI - Regional analgesia after total knee replacement. PMID- 10866928 TI - Applying parametric tests to visual analog scores. PMID- 10866929 TI - Single-lung ventilation in young children: practical tips on using conventional cuffed endotracheal tubes for VATS. PMID- 10866930 TI - Infusion pump use in the MRI. PMID- 10866931 TI - Preoperative anxiety and intraoperative anesthetic requirements. PMID- 10866932 TI - Early detection of the TURP syndrome. PMID- 10866933 TI - Evidence for myocardial ATP compartmentation from NMR inversion transfer analysis of creatine kinase fluxes. AB - The interpretation of creatine kinase (CK) flux measured by (31)P NMR magnetization transfer in vivo is complex because of the presence of competing reactions, metabolite compartmentation, and CK isozyme localization. In the isovolumic perfused rat heart, we considered the influence of both ATP compartmentation and ATP-P(i) exchange on the forward (F(f): PCr --> ATP) and reverse (F(r)) CK fluxes derived from complete analysis of inversion transfer. Although F(f) should equal F(r) because of the steady state, in both protocols when PCr (inv-PCr) or ATP (inv-ATP) was inverted and the contribution of ATP-P(i) was masked by saturation of P(i) (sat-P(i)), F(f)/F(r) significantly differed from 1 (0.80 +/- 0.06 or 1.32 +/- 0.06, respectively, n = 5). These discrepancies could be explained by a compartment of ATP (f(ATP)) not involved in CK. Consistently, neglecting ATP compartmentation in the analysis of CK in vitro results in an underestimation of F(f)/F(r) for inv-PCr and its overestimation for inv-ATP. Both protocols gave access to f(ATP) if the system was adequately analyzed. The fraction of ATP not involved in CK reaction in a heart performing medium work amounts to 20-33% of cellular ATP. Finally, the data suggest that the effect of sat-P(i) might not result only from the masking of ATP-P(i) exchange. PMID- 10866934 TI - Quinone-dependent delayed fluorescence from the reaction center of photosynthetic bacteria. AB - Millisecond delayed fluorescence from the isolated reaction center of photosynthetic bacteria Rhodobacter sphaeroides was measured after single saturating flash excitation and was explained by thermal repopulation of the excited bacteriochlorophyll dimer from lower lying charge separated states. Three exponential components (fastest, fast, and slow) were found with lifetimes of 1.5, 102, and 865 ms and quantum yields of 6.4 x 10(-9), 2.2 x 10(-9), and 2.6 x 10(-9) (pH 8.0), respectively. While the two latter phases could be related to transient absorption changes, the fastest one could not. The fastest component, dominating when the primary quinone was prereduced, might be due to a small fraction of long-lived triplet states of the radical pair and/or the dimer. The fast phase observed in the absence of the secondary quinone, was sensitive to pH, temperature, and the chemical nature of the primary quinone. The standard free energy of the primary stable charge pair relative to that of the excited dimer was -910 +/- 20 meV at pH 8 and with native ubiquinone, and it showed characteristic changes upon pH and quinone replacement. The interaction energy ( approximately 50 meV) between the cluster of the protonatable groups around GluL212 and the primary semiquinone provides evidence for functional linkage between the two quinone binding pockets. An empirical relationship was found between the in situ free energy of the primary quinone and the rate of charge recombination, with practical importance in the estimation of the free energy levels from the easily available lifetime of the charge recombination. The ratio of the slow and fast components could be used to determine the pH dependence of the free energy level of the secondary stable charge pair relative to that of the excited dimer. PMID- 10866935 TI - A three-state model for energy trapping and chlorophyll fluorescence in photosystem II incorporating radical pair recombination. AB - The multiphasic fluorescence induction kinetics upon a high intensity light pulse have been measured and analyzed at a time resolution of 10 micros in intact leaves of Peperomia metallica and Chenopodium album and in chloroplasts isolated from the latter. Current theories and models on the relation between chlorophyll fluorescence yield and primary photochemistry in photosystem II (PSII) are inadequate to describe changes in the initial phase of fluorescence induction and in the dark fluorescence level F(0) caused by pre-energization of the system with single turnover excitation(s). A novel model is presented, which gives a quantitative relation between the efficiencies of primary photochemistry, energy trapping, and radical pair recombination in PSII. The model takes into account that at least two turnovers are required for stationary closure of a reaction center. An open reaction center is transferred with high efficiency into its semiclosed (-open) state. This state is characterized by Q(A) and P680 in the fully reduced state and a lifetime equal to the inverse of the rate constant of Q(A)(-) oxidation (approx. 250 micros). The fluorescence yield of the system with 100% of the centers in the semiclosed state is 50% of the maximal yield with all centers in the closed state at fluorescence level F(m). A situation with approximately 100% of the centers in the semiclosed state is reached after a single turnover excitation in the presence of 3-(3',4'-dichlorophenyl)-1,1 dimethylurea (DCMU). The lifetime of this state under these conditions is approximately 10 s. Closure of a semiclosed (-open) center occurs with low efficiency in a second turnover. The low(er) efficiency is caused by the rate of P(+) reduction by the secondary donor Y(Z) being competitive with the rate of radical pair recombination in second and following turnovers. The single-turnover induced alterations in the initial kinetics of the fluorescence concomitantly with a 15-25% increase in F(o) can be simulated with the present so called three state model of energy trapping. The experimental data suggest evidence for an electrostatic effect of local charges in the vicinity of the reaction center affecting the rate of radical pair recombination in the reaction center. PMID- 10866936 TI - Mitochondrial calcium transients in adult rabbit cardiac myocytes: inhibition by ruthenium red and artifacts caused by lysosomal loading of Ca(2+)-indicating fluorophores. AB - A cold/warm loading protocol was used to ester-load Rhod 2 into mitochondria and other organelles and Fluo 3 into the cytosol of adult rabbit cardiac myocytes for confocal fluorescence imaging. Transient increases in both cytosolic Fluo 3 and mitochondrial Rhod 2 fluorescence occurred after electrical stimulation. Ruthenium red, a blocker of the mitochondrial Ca(2+) uniporter, inhibited mitochondrial Rhod 2 fluorescence transients but not cytosolic Fluo 3 transients. Thus the ruthenium red-sensitive mitochondrial Ca(2+) uniporter catalyzes Ca(2+) uptake during beat-to-beat transients of mitochondrial free Ca(2+), which in turn may help match mitochondrial ATP production to myocardial ATP demand. After ester loading, substantial amounts of Ca(2+)-indicating fluorophores localized into an acidic lysosomal/endosomal compartment. This lysosomal fluorescence did not respond to electrical stimulation. Because fluorescence arose predominantly from lysosomes after the cold loading/warm incubation procedure, total cellular fluorescence failed to track beat-to-beat changes of mitochondrial fluorescence. Only three-dimensionally resolved confocal imaging distinguished the relatively weak mitochondrial signal from the bright lysosomal fluorescence. PMID- 10866937 TI - The key event in force-induced unfolding of Titin's immunoglobulin domains. AB - Steered molecular dynamics simulation of force-induced titin immunoglobulin domain I27 unfolding led to the discovery of a significant potential energy barrier at an extension of approximately 14 A on the unfolding pathway that protects the domain against stretching. Previous simulations showed that this barrier is due to the concurrent breaking of six interstrand hydrogen bonds (H bonds) between beta-strands A' and G that is preceded by the breaking of two to three hydrogen bonds between strands A and B, the latter leading to an unfolding intermediate. The simulation results are supported by Angstrom-resolution atomic force microscopy data. Here we perform a structural and energetic analysis of the H-bonds breaking. It is confirmed that H-bonds between strands A and B break rapidly. However, the breaking of the H-bond between strands A' and G needs to be assisted by fluctuations of water molecules. In nanosecond simulations, water molecules are found to repeatedly interact with the protein backbone atoms, weakening individual interstrand H-bonds until all six A'-G H-bonds break simultaneously under the influence of external stretching forces. Only when those bonds are broken can the generic unfolding take place, which involves hydrophobic interactions of the protein core and exerts weaker resistance against stretching than the key event. PMID- 10866938 TI - Application of the primary hydration shell approach to locally enhanced sampling simulated annealing: computer simulation of thyrotropin-releasing hormone in water. AB - A unified model of simulated annealing with locally enhanced sampling (LES) in a primary hydration shell (PHS) aqueous environment is developed and tested by predicting the structure of the tripeptide thyrotropin-releasing hormone (TRH) in solution. The model extends the formulation of the restraining force in the PHS method as a function of temperature, number of copies in the LES method, and shell thickness. The dependence of the restraining force on temperature can be shown to follow the relationship c(1)T - c(2), which can be derived from the expression for kinetic energy in molecular dynamics simulations. The calibration of the restraining force for different simulation conditions reveals the dependence of c(1) and c(2) on the number of copies in the LES method and the thickness of the PHS. The predicted structure of TRH is in very good agreement with results from NMR experiments and from a 10-ns PHS simulation at 300 K. The method promises to be useful in predicting structure of peptides and proteins in an aqueous environment. PMID- 10866939 TI - Three-dimensional Poisson-Nernst-Planck theory studies: influence of membrane electrostatics on gramicidin A channel conductance. AB - A recently introduced real-space lattice methodology for solving the three dimensional Poisson-Nernst-Planck equations is used to compute current-voltage relations for ion permeation through the gramicidin A ion channel embedded in membranes characterized by surface dipoles and/or surface charge. Comparisons to a variety of experimental results, presented herein, have proven largely successful. Strengths and weaknesses of the method are discussed. PMID- 10866940 TI - A mathematical model of cardiocyte Ca(2+) dynamics with a novel representation of sarcoplasmic reticular Ca(2+) control. AB - Cardiac contraction and relaxation dynamics result from a set of simultaneously interacting Ca(2+) regulatory mechanisms. In this study, cardiocyte Ca(2+) dynamics were modeled using a set of six differential equations that were based on theories, equations, and parameters described in previous studies. Among the unique features of the model was the inclusion of bidirectional modulatory interplay between the sarcoplasmic reticular Ca(2+) release channel (SRRC) and calsequestrin (CSQ) in the SR lumen, where CSQ acted as a dynamic rather than simple Ca(2+) buffer, and acted as a Ca(2+) sensor in the SR lumen as well. The inclusion of this control mechanism was central in overcoming a number of assumptions that would otherwise have to be made about SRRC kinetics, SR Ca(2+) release rates, and SR Ca(2+) release termination when the SR lumen is assumed to act as a simple, buffered Ca(2+) sink. The model was sufficient to reproduce a graded Ca(2+)-induced Ca(2+) release (CICR) response, CICR with high gain, and a system with reasonable stability. As constructed, the model successfully replicated the results of several previously published experiments that dealt with the Ca(2+) dependence of the SRRC (, J. Gen. Physiol. 85:247-289), the refractoriness of the SRRC (, Am. J. Physiol. 270:C148-C159), the SR Ca(2+) load dependence of SR Ca(2+) release (, Am. J. Physiol. 268:C1313-C1329;, J. Biol. Chem. 267:20850-20856), SR Ca(2+) leak (, J. Physiol. (Lond.). 474:463-471;, Biophys. J. 68:2015-2022), SR Ca(2+) load regulation by leak and uptake (, J. Gen. Physiol. 111:491-504), the effect of Ca(2+) trigger duration on SR Ca(2+) release (, Am. J. Physiol. 258:C944-C954), the apparent relationship that exists between sarcoplasmic and sarcoplasmic reticular calcium concentrations (, Biophys. J. 73:1524-1531), and a variety of contraction frequency-dependent alterations in sarcoplasmic [Ca(2+)] dynamics that are normally observed in the laboratory, including rest potentiation, a negative frequency-[Ca(2+)] relationship, and extrasystolic potentiation. Furthermore, under the condition of a simulated Ca(2+) overload, an alternans-like state was produced. In summary, the current model of cardiocyte Ca(2+) dynamics provides an integrated theoretical framework of fundamental cellular Ca(2+) regulatory processes that is sufficient to predict a broad array of observable experimental outcomes. PMID- 10866941 TI - DNA rings with multiple energy minima. AB - Within the context of DNA rings, we analyze the relationship between intrinsic shape and the existence of multiple stable equilibria, either nicked or cyclized with the same link. A simple test, based on a perturbation expansion of symmetry breaking within a continuum elastic rod model, provides good predictions of the occurrence of such multiple equilibria. The reliability of these predictions is verified by direct computation of nicked and cyclized equilibria for several thousand DNA minicircles with lengths of 200 and 900 bp. Furthermore, our computations of equilibria for nicked rings predict properties of the equilibrium distribution of link, as calculated by much more computationally intensive Monte Carlo simulations. PMID- 10866942 TI - Single-particle tracking: Brownian dynamics of viscoelastic materials. AB - A unifying theoretical framework for analyzing stochastic data from single particle tracking (SPT) in viscoelastic materials is presented. A generalization of the bead-spring model for linear polymers is developed from a molecular point of view and from the standpoint of phenomenological linear viscoelasticity. The hydrodynamic interaction in the former is identified as the dashpots in the latter. In elementary terms, the intimate correspondence between time-correlation of the fluctuation measurements and transient relaxation kinetics after perturbation is discussed, and the central role of the fluctuation-dissipation relation is emphasized. The work presented here provides a bridge between the microscopic and the macroscopic views of linear viscoelastic biological materials, and is applicable to membrane protein diffusion, linear DNA chain dynamics, and mechanics of intracellular cytoskeletal networks. PMID- 10866943 TI - Cell movement is guided by the rigidity of the substrate. AB - Directional cell locomotion is critical in many physiological processes, including morphogenesis, the immune response, and wound healing. It is well known that in these processes cell movements can be guided by gradients of various chemical signals. In this study, we demonstrate that cell movement can also be guided by purely physical interactions at the cell-substrate interface. We cultured National Institutes of Health 3T3 fibroblasts on flexible polyacrylamide sheets coated with type I collagen. A transition in rigidity was introduced in the central region of the sheet by a discontinuity in the concentration of the bis-acrylamide cross-linker. Cells approaching the transition region from the soft side could easily migrate across the boundary, with a concurrent increase in spreading area and traction forces. In contrast, cells migrating from the stiff side turned around or retracted as they reached the boundary. We call this apparent preference for a stiff substrate "durotaxis." In addition to substrate rigidity, we discovered that cell movement could also be guided by manipulating the flexible substrate to produce mechanical strains in the front or rear of a polarized cell. We conclude that changes in tissue rigidity and strain could play an important controlling role in a number of normal and pathological processes involving cell locomotion. PMID- 10866944 TI - Stability analysis of micropipette aspiration of neutrophils. AB - During micropipette aspiration, neutrophil leukocytes exhibit a liquid-drop behavior, i.e., if a neutrophil is aspirated by a pressure larger than a certain threshold pressure, it flows continuously into the pipette. The point of the largest aspiration pressure at which the neutrophil can still be held in a stable equilibrium is called the critical point of aspiration. Here, we present a theoretical analysis of the equilibrium behavior and stability of a neutrophil during micropipette aspiration with the aim to rigorously characterize the critical point. We take the energy minimization approach, in which the critical point is well defined as the point of the stability breakdown. We use the basic liquid-drop model of neutrophil rheology extended by considering also the neutrophil elastic area expansivity. Our analysis predicts that the behavior at large pipette radii or small elastic area expansivity is close to the one predicted by the basic liquid-drop model, where the critical point is attained slightly before the projection length reaches the pipette radius. The effect of elastic area expansivity is qualitatively different at smaller pipette radii, where our analysis predicts that the critical point is attained at the projection lengths that may significantly exceed the pipette radius. PMID- 10866945 TI - An image-based model of calcium waves in differentiated neuroblastoma cells. AB - Calcium waves produced by bradykinin-induced inositol-1,4, 5-trisphosphate (InsP(3))-mediated release from endoplasmic reticulum (ER) have been imaged in N1E-115 neuroblastoma cells. A model of this process was built using the "virtual cell," a general computational system for integrating experimental image, biochemical, and electrophysiological data. The model geometry was based on a cell for which the calcium wave had been experimentally recorded. The distributions of the relevant cellular components [InsP(3) receptor (InsP(3)R)], sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) pumps, bradykinin receptors, and ER] were based on 3D confocal immunofluorescence images. Wherever possible, known biochemical and electrophysiological data were used to constrain the model. The simulation closely matched the spatial and temporal characteristics of the experimental calcium wave. Predictions on different patterns of calcium signals after InsP(3) uncaging or for different cell geometries were confirmed experimentally, thus helping to validate the model. Models in which the spatial distributions of key components are altered suggest that initiation of the wave in the center of the neurite derives from an interplay of soma-biased ER distribution and InsP(3) generation biased toward the neurite. Simulations demonstrate that mobile buffers (like the indicator fura-2) significantly delay initiation and lower the amplitude of the wave. Analysis of the role played by calcium diffusion indicated that the speed of the wave is only slightly dependent on the ability of calcium to diffuse to and activate neighboring InsP(3) receptor sites. PMID- 10866946 TI - Size-dependent positioning of human chromosomes in interphase nuclei. AB - By using a fluorescence in situ hybridization technique we revealed that for nine different q-arm telomere markers the positioning of chromosomes in human G(1) interphase nuclei was chromosome size-dependent. The q-arm telomeres of large chromosomes are more peripherally located than telomeres on small chromosomes. This highly organized arrangement of chromatin within the human nucleus was discovered by determining the x and y coordinates of the hybridization sites and calculating the root-mean-square radial distance to the nuclear centers in human fibroblasts. We demonstrate here that global organization within the G(1) interphase nucleus is affected by one of the most fundamental physical quantities chromosome size or mass-and propose two biophysical models, a volume exclusion model and a mitotic preset model, to explain our finding. PMID- 10866947 TI - Probing f-actin flow by tracking shape fluctuations of radial bundles in lamellipodia of motile cells. AB - We examined the dynamics of radial actin bundles based on time-lapse movies of polarized light images of living neuronal growth cones. Using a highly sensitive computer vision algorithm for tracking, we analyzed the small shape fluctuations of radial actin bundles that otherwise remained stationary in their positions in the growth cone lamellipodium. Using the tracking software, we selected target points on radial bundles and measured both the local bundle orientations and the lateral displacements between consecutive movie frames. We found that the local orientation and the lateral displacement of a target point are correlated. The correlation can be explained using a simple geometric relationship between the lateral travel of tilted actin bundles and the retrograde flow of f-actin structures. Once this relationship has been established, we have turned the table and used the radial bundles as probes to measure the velocity field of f-actin flow. We have generated a detailed map of the complex retrograde flow pattern throughout the lamellipodium. Such two-dimensional flow maps will give new insights into the mechanisms responsible for f-actin-mediated cell motility and growth. PMID- 10866948 TI - Distinct ion channel classes are expressed on the outer nuclear envelope of T- and B-lymphocyte cell lines. AB - The outer nuclear membrane, endoplasmic reticulum, and mitochondrial membrane ion channels are poorly understood, although they are important in the control of compartmental calcium levels, cell division, and apoptosis. Few direct recordings of these ion channels have been made because of the difficulty of accessing these intracellular membranes. Using patch-clamp techniques on isolated nuclei, we measured distinct ion channel classes on the outer nuclear envelope of T-cell (human Jurkat) and BFL5 cell (murine promyelocyte) lines. We first imaged the nuclear envelopes of both Jurkat and FL5 cells with atomic force microscopy to determine the density of pore proteins. The nuclear pore complex was intact at roughly similar densities in both cell types. In patch-clamp recordings of Jurkat nuclear membranes, Cl channels (105 +/- 5 pS) predominated and inactivated with negative pipette potentials. Nucleotides transiently inhibited the anion channel. In contrast, FL5 nuclear channels were cation selective (52 +/- 2 pS), were inactivated with positive membrane potentials, and were insensitive to GTPgammaS applied to the bath. We hypothesize that T- and B-cell nuclear membrane channels are distinct, and that this is perhaps related to their unique roles in the immune system. PMID- 10866949 TI - Proton-sensitive transitions of renal type II Na(+)-coupled phosphate cotransporter kinetics. AB - In the kidney proximal tubule, acidification of the glomerular filtrate leads to an inhibition of inorganic phosphate (P(i)) reabsorption by type II Na(+)-coupled cotransporters (NaPi-II). As external pH also alters the divalent/monovalent P(i) ratio, it has been difficult to separate putative proton interactions with the cotransporter from direct titration of divalent P(i), the preferred species transported. To distinguish between these possibilities and identify pH-sensitive transitions in the cotransport cycle, the pH-dependent kinetics of two NaPi-II isoforms, expressed in Xenopus laevis oocytes, were investigated electrophysiologically. At -50 mV, both isoforms showed >70% suppression of electrogenic response for an external pH change from 8.0 to 6.2, not attributable to titration of divalent P(i). This was accompanied by a progressive removal of steady-state voltage dependence. The NaPi-II-related uncoupled slippage current was unaffected by a pH change from 7.4 to 6.2, with no shift in the reversal potential, which suggested that protons do not function as substrate. The voltage dependence of pre-steady-state relaxations was shifted to depolarizing potentials in 100 mM and 0 mM Na(ext)(+) and two kinetic components were resolved, the slower of which was pH-dependent. The changes in kinetics are predicted by a model in which protons interact with the empty carrier and final Na(+) binding step. PMID- 10866950 TI - Differential effects of amino-terminal distal and proximal domains in the regulation of human erg K(+) channel gating. AB - The participation of amino-terminal domains in human ether-a-go-go (eag)-related gene (HERG) K(+) channel gating was studied using deleted channel variants expressed in Xenopus oocytes. Selective deletion of the HERG-specific sequence (HERG Delta138-373) located between the conserved initial amino terminus (the eag or PAS domain) and the first transmembrane helix accelerates channel activation and shifts its voltage dependence to hyperpolarized values. However, deactivation time constants from fully activated states and channel inactivation remain almost unaltered after the deletion. The deletion effects are equally manifested in channel variants lacking inactivation. The characteristics of constructs lacking only about half of the HERG-specific domain (Delta223-373) or a short stretch of 19 residues (Delta355-373) suggest that the role of this domain is not related exclusively to its length, but also to the presence of specific sequences near the channel core. Deletion-induced effects are partially reversed by the additional elimination of the eag domain. Thus the particular combination of HERG specific and eag domains determines two important HERG features: the slow activation essential for neuronal spike-frequency adaptation and maintenance of the cardiac action potential plateau, and the slow deactivation contributing to HERG inward rectification. PMID- 10866951 TI - M2 pore mutations convert the glycine receptor channel from being anion- to cation-selective. AB - Three mutations in the M2 transmembrane domains of the chloride-conducting alpha1 homomeric glycine receptor (P250Delta, A251E, and T265V), which normally mediate fast inhibitory neurotransmission, produced a cation-selective channel with P(Cl)/P(Na), = 0.27 (wild-type P(Cl)/P(Na) = 25), a permeability sequence P(Cs) > P(K) > P(Na) > P(Li), an impermeability to Ca(2+), and a reduced glycine sensitivity. Outside-out patch measurements indicated reversed and accentuated rectification with extremely low mean single channel conductances of 3 pS (inward current) and 11 pS (outward current). The three inverse mutations, to those analyzed in this study, have previously been shown to make the alpha7 acetylcholine receptor channel anion-selective, indicating a common location for determinants of charge selectivity of inhibitory and excitatory ligand-gated ion channels. PMID- 10866952 TI - Block of voltage-dependent calcium channels by aliphatic monoamines. AB - We have recently identified farnesol, an intermediate in the mevalonate pathway, as a potent endogenous modulator and blocker of N-type calcium channels (Roullet, J. B., R. L. Spaetgens, T. Burlingame, and G. W. Zamponi. 1999. J. Biol. Chem. 274:25439-25446). Here, we investigate the action of structurally related compounds on various types of voltage-dependent Ca(2+) channels transiently expressed in human embryonic kidney cells. 1-Dodecanol, despite sharing the 12 carbon backbone and headgroup of farnesol, exhibited a significantly lower blocking affinity for N-type Ca(2+) channels. Among several additional 12-carbon compounds tested, dodecylamine (DDA) mediated the highest affinity inhibition of N-type channels, indicating that the functional headgroup is a critical determinant of blocking affinity. This inhibition was concentration-dependent and relatively non-discriminatory among N-, L-, P/Q-, and R-Ca(2+) channel subtypes. However, whereas L-type channels exhibited predominantly resting channel block, the non-L-type isoforms showed substantial rapid open channel block manifested by a speeding of the apparent time course of current decay and block of the inactivated state. Consistent with these findings, we observed significant frequency-dependence of block and dependence on external Ba(2+) concentration for N-type, but not L-type, channels. We also systematically investigated the drug structural requirements for N-type channel inhibition. Blocking affinity varied with carbon chain length and showed a clear maximum at C12 and C13, with shorter and longer molecules producing progressively weaker peak current block. Overall, our data indicate that aliphatic monoamines may constitute a novel class of potent inhibitors of voltage-dependent Ca(2+) channels, with block being governed by rigid structural requirements and channel-specific state dependencies. PMID- 10866954 TI - Sulfhydryl oxidation overrides Mg(2+) inhibition of calcium-induced calcium release in skeletal muscle triads. AB - We studied the effect of oxidation of sulfhydryl (SH) residues on the inhibition by Mg(2+) of calcium-induced calcium release (CICR) in triad-enriched sarcoplasmic reticulum vesicles isolated from rabbit skeletal muscle. Vesicles were either passively or actively loaded with calcium before eliciting CICR by dilution at pCa 4.6-4.4 in the presence of 1.2 mM free [ATP] and variable free [Mg(2+)]. Native triads exhibited a significant inhibition of CICR by Mg(2+), with a K(0.5) approximately 50 microM. Partial oxidation of vesicles with thimerosal produced a significant increase of release rate constants and initial release rates at all [Mg(2+)] tested (up to 1 mM), and shifted the K(0.5) value for Mg(2+) inhibition to 101 or 137 microM in triads actively or passively loaded with calcium, respectively. Further oxidation of vesicles with thimerosal completely suppressed the inhibitory effect of [Mg(2+)] on CICR, yielding initial rates of CICR of 2 micromol/(mg x s) in the presence of 1 mM free [Mg(2+)]. These effects of oxidation on CICR were fully reversed by SH reducing agents. We propose that oxidation of calcium release channels, by decreasing markedly the affinity of the channel inhibitory site for Mg(2+), makes CICR possible in skeletal muscle. PMID- 10866953 TI - Regulation of the type III InsP(3) receptor by InsP(3) and ATP. AB - Many hormones and neurotransmitters raise intracellular calcium (Ca(2+)) by generating InsP(3) and activating the inositol 1,4, 5-trisphosphate receptor (InsP(3)R). Multiple isoforms with distinct InsP(3) binding properties () have been identified (). The type III InsP(3)R lacks Ca(2+)-dependent inhibition, a property that makes it ideal for signal initiation (). Regulation of the type III InsP(3)R by InsP(3) and ATP was explored in detail using planar lipid bilayers. In comparison to the type I InsP(3)R, the type III InsP(3)R required a higher concentration of InsP(3) to reach maximal channel activity (EC(50) of 3.2 microM versus 0.5 microM for the types III and I InsP(3)R, respectively). However, the type III InsP(3)R did reach a 2.5-fold higher level of activity. Although activation by InsP(3) was isoform-specific, regulation by ATP was similar for both isoforms. In the presence of 2 microM InsP(3), low ATP concentrations (<6 mM) increased the open probability and mean open time. High ATP concentrations (>6 mM) decreased channel activity. These results illustrate the complex nature of type III InsP(3)R regulation. Enhanced channel activity in the presence of high InsP(3) may be important during periods of prolonged stimulation, whereas allosteric modulation by ATP may help to modulate intracellular Ca(2+) signaling. PMID- 10866955 TI - A novel type of ATP block on a Ca(2+)-activated K(+) channel from bullfrog erythrocytes. AB - Using the patch-clamp technique, we have identified an intermediate conductance Ca(2+)-activated K(+) channel from bullfrog (Rana catesbeiana) erythrocytes and have investigated the regulation of channel activity by cytosolic ATP. The channel was highly selective for K(+) over Na(+), gave a linear I-V relationship with symmetrical 117.5 mM K(+) solutions and had a single-channel conductance of 60 pS. Channel activity was dependent on Ca(2+) concentration (K(1/2) = 600 nM) but voltage-independent. These basic characteristics are similar to those of human and frog erythrocyte Ca(2+)-activated K(+) (Gardos) channels previously reported. However, cytoplasmic application of ATP reduced channel activity with block exhibiting a novel bell-shaped concentration dependence. The channel was inhibited most by approximately 10 microM ATP (P(0) reduced to 5% of control) but less blocked by lower and higher concentrations of ATP. Moreover, the novel type of ATP block did not require Mg(2+), was independent of PKA or PKC, and was mimicked by a nonhydrolyzable ATP analog, AMP-PNP. This suggests that ATP exerts its effect by direct binding to sites on the channel or associated regulatory proteins, but not by phosphorylation of either of these components. PMID- 10866956 TI - Interaction between permeation and gating in a putative pore domain mutant in the cystic fibrosis transmembrane conductance regulator. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel with distinctive kinetics. At the whole-cell level, CFTR currents in response to voltage steps are time independent for wild type and for the many mutants reported so far. Single channels open for periods lasting up to tens of seconds; the openings are interrupted by brief closures at hyperpolarized, but not depolarized, potentials. Here we report a serine-to-phenylalanine mutation (S1118F) in the 11th transmembrane domain that confers voltage-dependent, single exponential current relaxations and moderate inward rectification of the macroscopic currents upon expression in Xenopus oocytes. At steady state, the S1118F-CFTR single-channel conductance rectifies, corresponding to the whole-cell rectification. In addition, the open-channel burst duration is decreased 10-fold compared with wild-type channels. S1118F-CFTR currents are blocked in a voltage dependent manner by diphenylamine-2-carboxylate (DPC); the affinity of S1118F CFTR for DPC is similar to that of the wild-type channel, but blockade exhibits moderately reduced voltage dependence. Selectivity of the channel to a range of anions is also affected by this mutation. Furthermore, the permeation properties change during the relaxations, which suggests that there is an interaction between gating and permeation in this mutant. The existence of a mutation that confers voltage dependence upon CFTR currents and that changes kinetics and permeation properties of the channel suggests a functional role for the 11th transmembrane domain in the pore in the wild-type channel. PMID- 10866957 TI - Direct measurement of specific membrane capacitance in neurons. AB - The specific membrane capacitance (C(m)) of a neuron influences synaptic efficacy and determines the speed with which electrical signals propagate along dendrites and unmyelinated axons. The value of this important parameter remains controversial. In this study, C(m) was estimated for the somatic membrane of cortical pyramidal neurons, spinal cord neurons, and hippocampal neurons. A nucleated patch was pulled and a voltage-clamp step was applied. The exponential decay of the capacitative charging current was analyzed to give the total membrane capacitance, which was then divided by the observed surface area of the patch. C(m) was 0.9 microF/cm(2) for each class of neuron. To test the possibility that membrane proteins may alter C(m), embryonic kidney cells (HEK 293) were studied before and after transfection with a plasmid coding for glycine receptor/channels. The value of C(m) was indistinguishable in untransfected cells and in transfected cells expressing a high level of glycine channels, indicating that differences in transmembrane protein content do not significantly affect C(m). Thus, to a first approximation, C(m) may be treated as a "biological constant" across many classes of neuron. PMID- 10866958 TI - Water permeability and mechanical strength of polyunsaturated lipid bilayers. AB - Micropipette aspiration was used to test mechanical strength and water permeability of giant-fluid bilayer vesicles composed of polyunsaturated phosphatidylcholine PC lipids. Eight synthetic-diacyl PCs were chosen with 18 carbon chains and degrees of unsaturation that ranged from one double bond (C18:0/1, C18:1/0) to six double bonds per PC molecule (diC18:3). Produced by increasing pipette pressurization, membrane tensions for lysis of single vesicles at 21 degrees C ranged from approximately 9 to 10 mN/m for mono- and dimono unsaturated PCs (18:0/1, 18:1/0, and diC18:1) but dropped abruptly to approximately 5 mN/m when one or both PC chains contained two cis-double bonds (C18:0/2 and diC18:2) and even lower approximately 3 mN/m for diC18:3. Driven by osmotic filtration following transfer of individual vesicles to a hypertonic environment, the apparent coefficient for water permeability at 21 degrees C varied modestly in a range from approximately 30 to 40 microm/s for mono- and dimono-unsaturated PCs. However, with two or more cis-double bonds in a chain, the apparent permeability rose to approximately 50 microm/s for C18:0/2, then strikingly to approximately 90 microm/s for diC18:2 and approximately 150 microm/s for diC18:3. The measurements of water permeability were found to scale exponentially with the reduced temperatures reported for these lipids in the literature. The correlation supports the concept that increase in free volume acquired in thermal expansion above the main gel-liquid crystal transition of a bilayer is a major factor in water transport. Taken together, the prominent changes in lysis tension and water permeability indicate that major changes occur in chain packing and cohesive interactions when two or more cis-double bonds alternate with saturated bonds along a chain. PMID- 10866959 TI - Effect of chain length and unsaturation on elasticity of lipid bilayers. AB - Micropipette pressurization of giant bilayer vesicles was used to measure both elastic bending k(c) and area stretch K(A) moduli of fluid-phase phosphatidylcholine (PC) membranes. Twelve diacyl PCs were chosen: eight with two 18 carbon chains and degrees of unsaturation from one double bond (C18:1/0, C18:0/1) to six double bonds per lipid (diC18:3), two with short saturated carbon chains (diC13:0, diC14:0), and two with long unsaturated carbon chains (diC20:4, diC22:1). Bending moduli were derived from measurements of apparent expansion in vesicle surface area under very low tensions (0.001-0.5 mN/m), which is dominated by smoothing of thermal bending undulations. Area stretch moduli were obtained from measurements of vesicle surface expansion under high tensions (>0.5 mN/m), which involve an increase in area per molecule and a small-but important contribution from smoothing of residual thermal undulations. The direct stretch moduli varied little (< +/-10%) with either chain unsaturation or length about a mean of 243 mN/m. On the other hand, the bending moduli of saturated/monounsaturated chain PCs increased progressively with chain length from 0.56 x 10(-19) J for diC13:0 to 1.2 x 10(-19) J for diC22:1. However, quite unexpectedly for longer chains, the bending moduli dropped precipitously to approximately 0.4 x 10(-19) J when two or more cis double bonds were present in a chain (C18:0/2, diC18:2, diC18:3, diC20:4). Given nearly constant area stretch moduli, the variations in bending rigidity with chain length and polyunsaturation implied significant variations in thickness. To test this hypothesis, peak-to peak headgroup thicknesses h(pp) of bilayers were obtained from x-ray diffraction of multibilayer arrays at controlled relative humidities. For saturated/monounsaturated chain bilayers, the distances h(pp) increased smoothly from diC13:0 to diC22:1 as expected. Moreover, the distances and elastic properties correlated well with a polymer brush model of the bilayer that specifies that the elastic ratio (k(c)/K(A))(1/2) = (h(pp) - h(o))/24, where h(o) approximately 1 nm accounts for separation of the headgroup peaks from the deformable hydrocarbon region. However, the elastic ratios and thicknesses for diC18:2, diC18:3, and diC20:4 fell into a distinct group below the correlation, which showed that poly-cis unsaturated chain bilayers are thinner and more flexible than saturated/monounsaturated chain bilayers. PMID- 10866960 TI - Pretransitional effects in dimyristoylphosphatidylcholine vesicle membranes: optical dynamometry study. AB - We used micron-sized latex spheres to probe the phase state and the viscoelastic properties of dimyristoylphosphatidylcholine (DMPC) bilayers as a function of temperature. One or two particles were manipulated and stuck to a DMPC giant vesicle by means of an optical trap. Above the fluid-gel main transition temperature, T(m) congruent with 23.4 degrees C, the particles could move on the surface of the vesicle, spontaneously (Brownian motion) or driven by an external force, either gravity or the laser beam's radiation pressure. From the analysis of the particle motions, we deduced the values of the membrane hydrodynamic shear viscosity, eta(s), and found that it would increase considerably near T(m). Below T(m), the long-distance motion of the particles was blocked. We performed experiments with two particles stuck on the membrane. By optical dynamometry, we measured the elastic resistance of the membrane to a variation in the interparticle distance and found that it would decrease considerably (down to zero) when the temperature was increased to T(m). We propose an interpretation relating the elastic response to the membrane curvature modulus, k(C). In this scheme, the two-bead dynamometry experiments provide a direct measurement of k(C) in the P'(beta) phase of lipid bilayers. PMID- 10866961 TI - Analysis of lung surfactant model systems with time-of-flight secondary ion mass spectrometry. AB - An often-used model lung surfactant containing dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG), and the surfactant protein C (SP C) was analyzed as Langmuir-Blodgett film by spatially resolved time-of-flight secondary ion mass spectrometry (TOF-SIMS) to directly visualize the formation and composition of domains. Binary lipid and lipid/SP-C systems were probed for comparison. TOF-SIMS spectra revealed positive secondary ions (SI) characteristic for DPPC and SP-C, but not for DPPG. SI mapping results in images with domain structures in DPPC/DPPG and DPPG/SP-C, but not in DPPC/SP-C films. We are able to distinguish between the fluid and condensed areas probably due to a matrix effect. These findings correspond with other imaging techniques, fluorescence light microscopy (FLM), scanning force microscopy (SFM), and silver decoration. The ternary mixture DPPC/DPPG/SP-C transferred from the collapse region exhibited SP-C-rich domains surrounding pure lipid areas. The results obtained are in full accordance with our earlier SFM picture of layered protrusions that serve as a compressed reservoir for surfactant material during expansion. Our study demonstrates once more that SP-C plays a unique role in the respiration process. PMID- 10866962 TI - Natural electrophoresis of norepinephrine and ascorbic acid. AB - The electric field produced by cell membranes, extending only a few nanometers, is 1000 times stronger than the electric fields required to produce dissociation of molecular complexes. Using the complex formed by norepinephrine (NE) and ascorbic acid (AA), we have demonstrated the quantitative binding of AA to NE, the use of capillary electrophoresis to measure quantitative binding of nonelectrolyte complexes, the determination of a dissociation constant (Kd) from electric field-dissociation constants (Ke), and a model for natural dissociation of the NE-AA complex due to the electric field generated by a cell membrane. NE AA dissociation constants show little effect of NE concentration or pH changes. NE-related compounds also bind AA: epinephrine > norepinephrine > tyrosine > histamine > phenylalanine. Serotonin does not bind AA. Phosphorylated AA and glucose also bind NE at 0.05 and 0.08 of the AA binding, respectively. Natural electrophoresis of molecular complexes allows compounds to travel through the body in a protected state and still be available for physiological activity upon reaching a membrane. PMID- 10866963 TI - Dimeric N-terminal segment of human surfactant protein B (dSP-B(1-25)) has enhanced surface properties compared to monomeric SP-B(1-25). AB - Surfactant protein B (SP-B) is a 17-kDa dimeric protein produced by alveolar type II cells. Its main function is to lower the surface tension by inserting lipids into the air/liquid interface of the lung. SP-B's function can be mimicked by a 25-amino acid peptide, SP-B(1-25), which is based on the N-terminal sequence of SP-B. We synthesized a dimeric version of this peptide, dSP-B(1-25), and the two peptides were tested for their surface activity. Both SP-B(1-25) and dSP-B(1-25) showed good lipid mixing and adsorption activities. The dimeric peptide showed activity comparable to that of native SP-B in the pressure-driven captive bubble surfactometer. Spread surface films led to stable near-zero minimum surface tensions during cycling while protein free, and films containing SP-B(1-25) lost material from the interface during compression. We propose that dimerization of the peptide is required to create a lipid reservoir attached to the monolayer from which new material can enter the surface film upon expansion of the air/liquid interface. The dimeric state of SP-B can fulfill the same function in vivo. PMID- 10866964 TI - Unusual hydration properties of C16:0 sulfatide bilayer membranes. AB - After deacylation of bovine brain sulfatide under mild alkaline conditions and reacylation using palmitoyl chloride (, Chem. Phys. Lipids. 34:41-53), the anionic glycosphingolipid N-palmitoyl galactosulfatide (C16:0-GalSulf) has been synthesized. By differential scanning calorimetry (DSC), anhydrous C16:0-GalSulf exhibits an endothermic transition, T(M) = 93 degrees C (DeltaH = 5. 5 kcal/mol C16:0-GalSulf) on heating. With increasing hydration (50 mM sodium phosphate buffer, pH 7.0; 50 mM NaCl), T(M) decreases, reaching a limiting value of 49 degrees C (DeltaH = 8.2 kcal/mol C16:0-GalSulf) at 20 wt% buffer. X-ray diffraction data have been recorded over the hydration range 0-62% at temperatures below (20 degrees C) and above (60 degrees C) T(M). At 20 degrees C, sharp wide-angle reflections at approximately 1/4.4 A(-1), approximately 1/4.1 A( 1), and approximately 1/3.8 A(-1) indicate the presence of an ordered-chain gel phase, whereas at 60 degrees C a broad reflection at 1/4.5 A(-1) characteristic of a melted-chain phase is observed. Lamellar diffraction patterns consistent with the presence of bilayer phases are observed at both temperatures. At 60 degrees C, in the liquid-crystalline L(alpha) phase, the bilayer periodicity increases with hydration, in both water and 100 mM Na(+) buffer. Interestingly, in the gel phase at 20 degrees C, the bilayer periodicity (d = 64 A) is insensitive to hydration (over the range 30-60 wt%) with either water or buffer. The continuous swelling behavior exhibited by the L(alpha) bilayer phase of C16:0 GalSulf is typical of lipids bearing a net negative charge and confirms that the presence of 100 mM Na(+) is insufficient to shield the charge contributed by the sulfate group. In contrast, the lack of continuous swelling behavior of the bilayer gel phase of C16:0-GalSulf is unusual and resembles that of Na(+) soaps. Thus, presumably, alterations in the surface charge characteristics of the C16:0 GalSulf bilayer occur on hydrocarbon chain melting and lead to major changes in lipid hydration. PMID- 10866965 TI - Membrane protein crystallization in lipidic mesophases: detergent effects. AB - The "cubic phase method" for growing crystals of membrane proteins uses a complex mixture of water, lipid, protein, and other components. The current view is that the cubic phase is integral to the process. Thus additives from whatever source introduce the possibility of destabilizing the phase, thereby compromising the crystallization process. Detergents are used to solubilize membrane proteins and are likely to be ported into the cubic medium with the target protein. Depending on the identity and concentration of the detergent, the cubic phase, which itself is membranous, may be solubilized or destabilized in such a way as to render it unsuitable as a crystal growing system. The nonionic detergent n-dodecyl-beta-D maltopyranoside is commonly used in membrane protein work. In this study, we evaluate its effect on the cubic mesophase of hydrated monoolein. X-ray diffraction was used for phase identification and mesophase microstructure characterization. The results show that while low levels of the detergent are tolerated, increasing concentrations trigger a cubic-to-lamellar phase transition in a temperature-dependent manner. This finding is rationalized in the context of complementary molecular shapes of the lipid and the detergent and has implications for the mechanism of crystallization in lipidic mesophases as discussed. PMID- 10866966 TI - Specific binding of ethanol to cholesterol in organic solvents. AB - Although ethanol has been reported to affect cholesterol homeostasis in biological membranes, the molecular mechanism of action is unknown. Here, nuclear magnetic resonance (NMR) spectroscopic techniques have been used to investigate possible direct interactions between ethanol and cholesterol in various low dielectric solvents (acetone, methanol, isopropanol, DMF, DMSO, chloroform, and CCl(4)). Measurement of (13)C chemical shifts, spin-lattice and multiplet relaxation times, as well as self-diffusion coefficients, indicates that ethanol interacts weakly, yet specifically, with the HC-OH moiety and the two flanking methylenes in the cyclohexanol ring of cholesterol. This interaction is most strong in the least polar-solvent carbon tetrachloride where the ethanol cholesterol equilibrium dissociation constant is estimated to be 2 x 10(-3) M. (13)C-NMR spin-lattice relaxation studies allow insight into the geometry of this complex, which is best modeled with the methyl group of ethanol sandwiched between the two methylenes in the cyclohexanol ring and the hydroxyl group of ethanol hydrogen bonded to the hydroxyl group of cholesterol. PMID- 10866967 TI - Two-photon fluorescence microscopy studies of bipolar tetraether giant liposomes from thermoacidophilic archaebacteria Sulfolobus acidocaldarius. AB - The effects of temperature and pH on Laurdan (6-lauroyl-2 (dimethylamino)naphthalene) fluorescence intensity images of giant unilamellar vesicles (GUVs) ( approximately 20-150 microm in diameter) composed of the polar lipid fraction E (PLFE) from the thermoacidophilic archaebacteria Sulfolobus acidocaldarius have been studied using two-photon excitation. PLFE GUVs made by the electroformation method were stable and well suited for microscopy studies. The generalized polarization (GP) of Laurdan fluorescence in the center cross section of the vesicles has been determined as a function of temperature at pH 7.23 and pH 2.68. At all of the temperatures and pHs examined, the GP values are low (below or close to 0), and the GP histograms show a broad distribution width (> 0.3). When excited with light polarized in the y direction, Laurdan fluorescence in the center cross section of the PLFE GUVs exhibits a photoselection effect showing much higher intensities in the x direction of the vesicles, a result opposite that previously obtained on monopolar diester phospholipids. This result indicates that the chromophore of Laurdan in PLFE GUVs is aligned parallel to the membrane surface. The x direction photoselection effect and the low GP values lead us to further propose that the Laurdan chromophore resides in the polar headgroup region of the PLFE liposomes, while the lauroyl tail inserts into the hydrocarbon core of the membrane. This unusual L-shaped disposition is presumably caused by the unique lipid structures and by the rigid and tight membrane packing in PLFE liposomes. The GP exhibited, at both pH values, a small but abrupt decrease near 50 degrees C, suggesting a conformational change in the polar headgroups of PLFE. This transition temperature fully agrees with the d-spacing data recently measured by small-angle x-ray diffraction and with the pyrene-labeled phosphatidylcholine and perylene fluorescence data previously obtained from PLFE multilamellar vesicles. Interestingly, the two-photon Laurdan fluorescence images showed snowflake-like lipid domains in PLFE GUVs at pH 7.23 and low temperatures (<20 degrees C in the cooling scan and <24 degrees C in the heating scan). These domains, attributable to lipid lateral separation, were stable and laterally immobile at low temperatures (<23 degrees C), again suggesting tight membrane packing in the PLFE GUVs. PMID- 10866968 TI - Mesoscopic undulations and thickness fluctuations in lipid bilayers from molecular dynamics simulations. AB - Molecular dynamics simulations of fully hydrated Dipalmitoylphosphatidylcholine bilayers, extending temporal and spatial scales by almost one order of magnitude, are presented. The present work reaches system sizes of 1024 lipids and times 10 60 ns. The simulations uncover significant dynamics and fluctuations on scales of several nanoseconds, and enable direct observation and spectral decomposition of both undulatory and thickness fluctuation modes. Although the former modes are strongly damped, the latter exhibit signs of oscillatory behavior. From this, it has been possible to calculate mesoscopic continuum properties in good agreement with experimental values. A bending modulus of 4 x 10(-20) J, bilayer area compressibility of 250-300 mN/m, and mode relaxation times in the nanosecond range are obtained. The theory of undulatory motions is revised and further extended to cover thickness fluctuations. Finally, it is proposed that thickness fluctuations is the explanation to the observed system-size dependence of equilibrium-projected area per lipid. PMID- 10866969 TI - A correlation between lipid domain shape and binary phospholipid mixture composition in free standing bilayers: A two-photon fluorescence microscopy study. AB - Giant unilamellar vesicles (GUVs) composed of different phospholipid binary mixtures were studied at different temperatures, by a method combining the sectioning capability of the two-photon excitation fluorescence microscope and the partition and spectral properties of 6-dodecanoyl-2-dimethylamino-naphthalene (Laurdan) and Lissamine rhodamine B 1,2-dihexadecanoyl-sn-glycero-3 phosphoethanolamine (N-Rh-DPPE). We analyzed and compared fluorescence images of GUVs composed of 1,2-dilauroyl-sn-glycero-3-phosphocholine/1, 2-dipalmitoyl-sn glycero-3-phosphocholine (DLPC/DPPC), 1, 2-dilauroyl-sn-glycero-3 phosphocholine/1, 2-distearoyl-sn-glycero-3-phosphocholine (DLPC/DSPC), 1, 2 dilauroyl-sn-glycero-3-phosphocholine/1, 2-diarachidoyl-sn-glycero-3 phosphocholine (DLPC/DAPC), 1, 2-dimyristoyl-sn-glycero-3-phosphocholine/1, 2 distearoyl-sn-glycero-3-phosphocholine (DMPC/DSPC) (1:1 mol/mol in all cases), and 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine/1, 2-dimyristoyl-sn-glycero 3-phosphocholine (DMPE/DMPC) (7:3 mol/mol) at temperatures corresponding to the fluid phase and the fluid-solid phase coexistence. In addition, we studied the solid-solid temperature regime for the DMPC/DSPC and DMPE/DMPC mixtures. From the Laurdan intensity images the generalized polarization function (GP) was calculated at different temperatures to characterize the phase state of the lipid domains. We found a homogeneous fluorescence distribution in the GUV images at temperatures corresponding to the fluid region for all of the lipid mixtures. At temperatures corresponding to phase coexistence we observed concurrent fluid and solid domains in the GUVs independent of the lipid mixture. In all cases the lipid solid domains expanded and migrated around the vesicle surface as we decreased the temperature. The migration of the solid domains decreased dramatically at temperatures close to the solid-fluid-->solid phase transition. For the DLPC-containing mixtures, the solid domains showed line, quasicircular, and dendritic shapes as the difference in the hydrophobic chain length between the components of the binary mixture increases. In addition, for the saturated PC containing mixtures, we found a linear relationship between the GP values for the fluid and solid domains and the difference between the hydrophobic chain length of the binary mixture components. Specifically, at the phase coexistence temperature region the difference in the GP values, associated with the fluid and solid domains, increases as the difference in the chain length of the binary mixture component increases. This last finding suggests that in the solid-phase domains, the local concentration of the low melting temperature phospholipid component increases as the hydrophobic mismatch decreases. At the phase coexistence temperature regime and based on the Laurdan GP data, we observe that when the hydrophobic mismatch is 8 (DLPC/DAPC), the concentration of the low melting temperature phospholipid component in the solid domains is negligible. This last observation extends to the saturated PE/PC mixtures at the phase coexistence temperature range. For the DMPC/DSPC we found that the nonfluorescent solid regions gradually disappear in the solid temperature regime of the phase diagram, suggesting lipid miscibility. This last result is in contrast with that found for DMPE/DMPC mixtures, where the solid domains remain on the GUV surface at temperatures corresponding to that of the solid region. In all cases the solid domains span the inner and outer leaflets of the membrane, suggesting a strong coupling between the inner and outer monolayers of the lipid membrane. This last finding extends previous observations of GUVs composed of DPPE/DPPC and DLPC/DPPC mixtures (, Biophys. J. 78:290-305). PMID- 10866970 TI - Outer membrane monolayer domains from two-dimensional surface scanning resistance measurements. AB - Cellular plasma membranes have domains that are defined, in most cases, by cytoskeletal elements. The outer half of the bilayer may also contain domains that organize glycosylphosphatidylinositol (GPI)-linked proteins. To define outer membrane barriers, we measured the resistive force on membrane bound beads as they were scanned across the plasma membrane of HEPA-OVA cells with optical laser tweezers. Beads were bound by antibodies to fluorescein-phosphatidylethanolamine (Fl-PE) or to the class I major histocompatibility complex (MHC class I) Qa-2 (a GPI-anchored protein). Two-dimensional scans of resistive force showed both occasional, resistive barriers and a velocity-dependent, continuous resistance. At the lowest antibody concentration, which gave specific binding, the continuous friction coefficient of Qa-2 was consistent with that observed by single-particle tracking (SPT) of small gold particles. At high antibody concentrations, the friction coefficient was significantly higher but decreased with increasing temperature, addition of deoxycholic acid, or treatment with heparinase I. Barriers to lateral movement (>3 times the continuous resistance) were consistently observed. Elastic barriers (with elastic constants from 1 to 20 pN/microm and sensitive to cytochalasin D) and small nonelastic barriers (<100 nm) were specifically observed with beads bound to the GPI-linked Qa-2. We suggest that GPI-linked proteins interact with transmembrane proteins when aggregated by antibody-coated beads and the transmembrane proteins encounter cytoplasmic barriers to lateral movement. The barriers to lateral movement are dynamic, discontinuous, and low in density. PMID- 10866971 TI - Is SH1-SH2-cross-linked myosin subfragment 1 a structural analog of the weakly bound state of myosin? AB - Myosin subfragment 1 (S1) with SH1 (Cys(707)) and SH2 (Cys(697)) groups cross linked by p-phenylenedimaleimide (pPDM-S1) is thought to be an analog of the weakly bound states of myosin bound to actin. The structural properties of pPDM S1 were compared in this study to those of S1.ADP.BeF(x) and S1.ADP.AlF(4)(-), i.e., the established structural analogs of the myosin weakly bound states. To distinguish between the conformational effects of SH1-SH2 cross-linking and those due to their monofunctional modification, we used S1 with the SH1 and SH2 groups labeled with N-phenylmaleimide (NPM-S1) as a control in our experiments. The state of the nucleotide pocket was probed using a hydrophobic fluorescent dye, 3 [4-(3-phenyl-2-pyrazolin-1-yl)benzene-1-sulfonylamido]phen ylboronic acid (PPBA). Differential scanning calorimetry (DSC) was used to study the thermal stability of S1. By both methods the conformational state of pPDM-S1 was different from that of unmodified S1 in the S1.ADP.BeF(x) and S1.ADP.AlF(4)(-) complexes and closer to that of nucleotide-free S1. Moreover, BeF(x) and AlF(4)(-) binding failed to induce conformational changes in pPDM-S1 similar to those observed in unmodified S1. Surprisingly, when pPDM cross-linking was performed on S1.ADP.BeF(x) complex, ADP.BeF(x) protected to some extent the nucleotide pocket of S1 from the effects of pPDM modification. NPM-S1 behaved similarly to pPDM-S1 in our experiments. Overall, this work presents new evidence that the conformational state of pPDM-S1 is different from that of the weakly bound state analogs, S1.ADP.BeF(x) and S1.ADP.AlF(4)(-). The similar structural effects of pPDM cross-linking of SH1 and SH2 groups and their monofunctional labeling with NPM are ascribed to the inhibitory effects of these modifications on the flexibility/mobility of the SH1-SH2 helix. PMID- 10866972 TI - Measurement of the force produced by an intact bull sperm flagellum in isometric arrest and estimation of the dynein stall force. AB - The force generated by a detergent-extracted reactivated bull sperm flagellum during an isometric stall was measured with a force-calibrated glass microprobe. The average isometric stall force from 48 individual measurements was 2.5 +/- 0.7 x 10(-5) dyne (2.5 +/- 0.7 x 10(-10) N). The force measurements were obtained by positioning a calibrated microprobe in the beat path of sperm cells that were stuck by their heads to a glass microscope slide. The average position of the contact point of the flagellum with the probe was 15 microm from the head-tail junction. This average lever arm length multiplied by the measured force yields an estimate of the active bending moment (torque) of 3.9 x 10(-8) dyne x cm (3.9 x 10(-15) N x m). The force was sustained and was for the most part uniform, despite the fact that the flagellum beyond the point of contact with the probe usually continued beating. It appears that the dynein motors in the basal portion of the flagellum continue to pull in an isometric stall for as long as the motion of the flagellum is blocked. If dynein motors in the flagellum distal to the contact point with the probe were contributing force to the displacement of the probe, then the flagellar segment immediately past the point of contact would have to show a net curvature in the direction of the probe displacement. No such curvature bias was observed in the R-bend arrests, and only a small positive curvature bias was measured in the P-bend arrests. Our analysis of the data suggests that more than 90% of the sustained force component is generated by the part of the flagellum between the probe and the flagellar base. Based on this premise, the isometric stall force per dynein head is estimated to be 5.0 x 10( 7) dyne (5 pN). This equals approximately 1.0 x 10(-6) dyne (10 pN) per intact dynein arm. These values are close to the isometric stall force of isolated dynein. This suggests that all of the dynein heads between the base and the probe, on the active side of the axoneme, are contributing to the force exerted against the probe. PMID- 10866973 TI - Translocation-independent dimerization of the EcoKI endonuclease visualized by atomic force microscopy. AB - Bacterial type I restriction/modification systems are capable of performing multiple actions in response to the methylation pattern on their DNA recognition sequences. The enzymes making up these systems serve to protect the bacterial cells against viral infection by binding to their recognition sequences on the invading DNA and degrading it after extensive ATP-driven translocation. DNA cleavage has been thought to occur as the result of a collision between two translocating enzyme complexes. Using atomic force microscopy (AFM), we show here that EcoKI dimerizes rapidly when bound to a plasmid containing two recognition sites for the enzyme. Dimerization proceeds in the absence of ATP and is also seen with an EcoKI mutant (K477R) that is unable to translocate DNA. Only monomers are seen when the enzyme complex binds to a plasmid containing a single recognition site. Based on our results, we propose that the binding of EcoKI to specific DNA target sequences is accompanied by a conformational change that leads rapidly to dimerization. This event is followed by ATP-dependent translocation and cleavage of the DNA. PMID- 10866974 TI - Nanosecond temperature jump and time-resolved Raman study of thermal unfolding of ribonuclease A. AB - A nanosecond temperature jump (T-jump) apparatus was constructed and combined with time-resolved Raman measurements to investigate thermal unfolding of a protein for the first time. The 1.56-microm heat pulse with 9 ns width at 10 Hz was obtained through the two-step stimulated Raman scattering in D(2) gas involving seeding and amplification. To achieve uniform temperature rise, the counter-propagation geometry was adopted for the heat pulse. The temperature rise was determined by anti-Stokes to Stokes intensity ratios of the 317 and 897 cm( 1) bands of MoO(4)(2-) ions in an aqueous solution. The T-jump as large as 9 degrees C in 10 ns was attained. The unfolding of bovine pancreatic ribonuclease A was monitored with time-resolved Raman spectra excited at 532 nm. The C-S stretching band of Met residues exhibited 10% change of that expected from the stationary state temperature-difference spectra in the initial 200 ns following T jump and another 10% in 5 ms. The Raman intensity of SO(4)(2-) ions around 980 cm(-1) increased at 100 micros, presumably due to some conformational changes of the protein around the active site. The S-S stretches and tyrosine doublet displayed little changes within 5 ms. Thus, the conformational changes in the initial step of unfolding are not always concerted. PMID- 10866975 TI - Synchrotron radiation diffraction from two-dimensional protein crystals at the air/water interface. AB - Protein structure determination by classical x-ray crystallography requires three dimensional crystals that are difficult to obtain for most proteins and especially for membrane proteins. An alternative is to grow two-dimensional (2D) crystals by adsorbing proteins to ligand-lipid monolayers at the surface of water. This confined geometry requires only small amounts of material and offers numerous advantages: self-assembly and ordering over micrometer scales is easier to obtain in two dimensions; although fully hydrated, the crystals are sufficiently rigid to be investigated by various techniques, such as electron crystallography or micromechanical measurements. Here we report structural studies, using grazing incidence synchrotron x-ray diffraction, of three different 2D protein crystals at the air-water interface, namely streptavidine, annexin V, and the transcription factor HupR. Using a set-up of high angular resolution, we observe narrow Bragg reflections showing long-range crystalline order in two dimensions. In the case of streptavidin the angular range of the observed diffraction corresponds to a resolution of 10 A in plane and 14 A normal to the plane. We show that this approach is complementary to electron crystallography but without the need for transfer of the monolayer onto a grid. Moreover, as the 2D crystals are accessible from the buffer solution, the formation and structure of protein complexes can be investigated in situ. PMID- 10866976 TI - Salt dependent stability and unfolding of [Fe2-S2] ferredoxin of Halobacterium salinarum: spectroscopic investigations. AB - Ferredoxin from the haloarchaeon Halobacterium salinarum is a 14. 6-kDa protein with a [Fe2-S2] center and is involved in the oxidative decarboxylation of 2 oxoacids. It possesses a high molar excess of acidic amino acid residues and is stable at high salt concentration. We have purified the protein from this extreme haloarchaeon and investigated its salt-dependent stability by circular dichroism, fluorescence, and absorption techniques. The predominantly beta-sheeted protein is stable in salt concentrations of >/=1.5 M NaCl. At lower concentrations a time dependent increase in fluorescence intensity ratio (I(360):I(330)), a decrease in the absorption at 420 nm, and a decrease in ellipticity values are observed. The rate of fluorescence intensity change at any low salt concentration is the highest, followed by absorption and ellipticity. This suggests that at low salt the unfolding of ferredoxin starts with the loss of tertiary structure, which leads to the disruption of the [Fe2-S2] center, resulting in the loss of secondary structural elements. PMID- 10866977 TI - Protein compressibility, dynamics, and pressure. AB - The relationship between the elastic and dynamic properties of native globular proteins is considered on the basis of a wide set of reported experimental data. The formation of a small cavity, capable of accommodating water, in the protein interior is associated with the elastic deformation, whose contribution to the free energy considerably exceeds the heat motion energy. Mechanically, the protein molecule is a highly nonlinear system. This means that its compressibility sharply decreases upon compression. The mechanical nonlinearity results in the following consequences related to the intramolecular dynamics of proteins: 1) The sign of the electrostriction effect in the protein matrix is opposite that observed in liquids-this is an additional indication that protein behaves like a solid particle. 2) The diffusion of an ion from the solvent to the interior of a protein should depend on pressure nonmonotonically: at low pressure diffusion is suppressed, while at high pressure it is enhanced. Such behavior is expected to display itself in any dynamic process depending on ion diffusion. Qualitative and quantitative expectations ensuing from the mechanical properties are concordant with the available experimental data on hydrogen exchange in native proteins at ambient and high pressure. PMID- 10866978 TI - Flexibility of the alpha-spectrin N-terminus by EPR and fluorescence polarization. AB - The structure and flexibility of the biologically important alpha-spectrin amino terminal region was examined by the use of fluorescence and EPR spectroscopy. The region studied has been previously demonstrated to be essential for the alpha spectrin:beta-spectrin association of the tetramerization site. Appropriate spectroscopic probe moieties were coupled to this region in a recombinant fragment of human erythroid alpha-spectrin. There was good agreement between the EPR and fluorescence techniques in most of this region. Mobility determinations indicated that a portion of the region was relatively immobilized. This is significant, since although predictive methods have indicated that this region should be alpha-helical, previous experimental evidence obtained on smaller synthetic peptides had indicated that this region was disordered. Observed rigidity appears to be incompatible with such a disordered state, and has important ramifications for the flexibility of this molecule that is so integral to its role in stabilizing erythrocyte membranes. PMID- 10866979 TI - Time-resolved polarization imaging by pump-probe (stimulated emission) fluorescence microscopy. AB - We report the application of pump-probe fluorescence microscopy in time-resolved polarization imaging. We derived the equations governing the pump-probe stimulated emission process and characterized the pump and probe laser power levels for signal saturation. Our emphasis is to use this novel methodology to image polarization properties of fluorophores across entire cells. As a feasibility study, we imaged a 15-microm orange latex sphere and found that there is depolarization that is possibly due to energy transfer among fluorescent molecules inside the sphere. We also imaged a mouse fibroblast labeled with CellTracker Orange CMTMR (5-(and-6)-(((4-chloromethyl)benzoyl)amino)tetramethyl rhodamine). We observed that Orange CMTMR complexed with gluthathione rotates fast, indicating the relatively low fluid-phase viscosity of the cytoplasmic microenvironment as seen by Orange CMTMR. The measured rotational correlation time ranged from approximately 30 to approximately 150 ps. This work demonstrates the effectiveness of stimulated emission measurements in acquiring high resolution, time-resolved polarization information across the entire cell. PMID- 10866980 TI - Aggregation kinetics of extended porphyrin and cyanine dye assemblies. AB - The kinetics of J-aggregate formation has been studied for two chromophores, tetrakis-4-sulfonatophenylporphine in an acid medium and pseudoisocyanine on a polyvinylsulfonate template. The assembly processes differ both in their sensitivity to initiation protocols and in the reaction profiles they produce. The porphyrin's assembly kinetics, for example, displays an induction period unlike that of the cyanine dye. Two kinetic models are presented. For the porphyrin, an autocatalytic pathway in which the formation of an aggregation nucleus is rate-determining appears to be applicable; for the pseudoisocyanine dye, an equation derived for diffusion-limited aggregation of a fractal object satisfactorily fits the data. These models are shown to be useful for the analysis of kinetic data obtained for several biologically important aggregation processes. PMID- 10866981 TI - Polymerization of rod-like macromolecular monomers studied by stopped-flow, multiangle light scattering: set-up, data processing, and application to fibrin formation. AB - Many biological supramolecular structures are formed by polymerization of macromolecular monomers. Light scattering techniques can provide structural information from such systems, if suitable procedures are used to collect the data and then to extract the relevant parameters. We present an experimental set up in which a commercial multiangle laser light scattering photometer is linked to a stopped-flow mixer, allowing, in principle, the time-resolved extrapolation of the weight-average molecular weight M(w) and of the z-average square radius of gyration (z) of the polymers from Zimm-like plots. However, if elongated structures are formed as the polymerization proceeds, curved plots rapidly arise, from which M(w) and (z) cannot be recovered by linear fitting. To verify the correctness of a polynomial fitting procedure, polydisperse collections of rod-like or worm-like particles of different lengths, generated at various stages during bifunctional polycondensations of rod-like macromolecular monomers, were considered. Then, the angular dependence of their time-averaged scattered intensity was calculated in the Rayleigh-Gans-Debye approximation, with random and systematic noise also added to the data. For relatively narrow size distributions, a third-degree polynomial fitting gave satisfactory results across a broad range of conversion degrees, yielding M(w) and (z) values within 2% and no greater than 10-20%, respectively, of the calculated values. When more broad size distributions were analyzed, the procedure still performed well for semiflexible polymers, but started to seriously underestimate both M(w) and (z) when rigid rod-like particles were analyzed, even at relatively low conversion degrees. The data were also analyzed in the framework of the Casassa approximation, from which the mass per unit length of the polymers can be derived. These procedures were applied to a set of data taken on the early stages of the thrombin-catalyzed polymerization of fibrinogen, a rod-like macromolecule approximately 50 nm long. The polymers, grown in the absence of Ca(2+) by rate limiting amounts of thrombin, appeared to be characterized by a much broader size distribution than the one expected for a classical Flory bifunctional polycondensation, and they seem to behave as relatively flexible worm-like double stranded chains. Evidence for the formation of fibrinogen-fibrin monomer complexes is also inferred from the time dependence of the mass/length ratio. However, our data are also compatible with the presence of limited amounts of single-stranded structures in the very early stages, either as a secondary, less populated pathway, or as transient intermediates to the classical double-stranded fibrils. PMID- 10866983 TI - Responses to repeated oral irritation by capsaicin, cinnamaldehyde and ethanol in PROP tasters and non-tasters. AB - Both increases (sensitization) and decreases (desensitization) in oral irritation have been reported in response to repeated short-term stimulation by compounds such as capsaicin, zingerone and menthol. It is unclear why one irritant would show sensitization and another desensitization, and this is further complicated by substantial inter-individual variation in response patterns. These variations may be the result of individual differences such as that represented by sensitivity to 6-n-propylthiouracil (PROP), which has been associated with variation in the overall intensity of irritation. In addition, comparisons between irritants have almost always involved inter-study comparisons, entailing different subject groups and frequently different methods. In the studies reported here, responses to three irritants-capsaicin, cinnamaldehyde and ethanol were examined as a function of PROP taster status. A common core of subjects also received all three irritants, allowing an assessment of the extent to which different response patterns between irritants seen previously were the result of different properties of the irritants themselves. Over a series of ten stimuli presented at 1 min intervals, PROP taster status differentiated subject responses on the basis of overall intensity, but not the pattern of responses over repeated stimulation. The group response to ethanol and cinnamaldehyde was desensitization, a pattern also shown by most of the individual subjects. In contrast, the group response to capsaicin was neither clear sensitization nor desensitization, reflecting much greater individual variability in response patterns. It is suggested that the time course to a single irritant stimulus largely determines between irritant response variations, while the inter-stimulus interval (ISI) used for a given irritant will have critical values for showing predominantly sensitization or desensitization. PMID- 10866982 TI - Salt-dependent compaction of di- and trinucleosomes studied by small-angle neutron scattering. AB - Using small-angle neutron scattering (SANS), we have measured the salt-dependent static structure factor of di- and trinucleosomes from chicken erythrocytes and from COS-7 cells. We also determined the sedimentation coefficients of these dinucleosomes and dinucleosomes reconstituted on a 416-bp DNA containing two nucleosome positioning sequences of the 5S rDNA of Lytechinus variegatus at low and high salt concentrations. The internucleosomal distance d was calculated by simulation as well as Fourier back-transformation of the SANS curves and by hydrodynamic simulation of sedimentation coefficients. Nucleosome dimers from chicken erythrocyte chromatin show a decrease in d from approximately 220 A at 5 mM NaCl to 150 A at 100 mM NaCl. For dinucleosomes from COS-7 chromatin, d decreases from 180 A at 5 mM to 140 A at 100 mM NaCl concentration. Our measurements on trinucleosomes are compatible with a compaction through two different mechanisms, depending on the salt concentration. Between 0 and 20 mM NaCl, the internucleosomal distance between adjacent nucleosomes remains constant, whereas the angle of the DNA strands entering and leaving the central nucleosome decreases. Above 20 mM NaCl, the adjacent nucleosomes approach each other, similar to the compaction of dinucleosomes. The internucleosomal distance of 140-150 A at 100 mM NaCl is in agreement with distances measured by scanning force microscopy and electron microscopy on long chromatin filaments. PMID- 10866984 TI - Latency and accuracy of discriminations of odor quality between binary mixtures and their components. AB - Subjects made timed, same-different discriminations of odor quality, with the following principal findings: (i) latency reflected accuracy, with difficult discriminations, i.e. those between 50-50 mixtures and their components, requiring more time than less difficult discriminations, i.e. those between unmixed chemicals. This finding demonstrated the face validity of latency as a measure of qualitative similarity. (ii) Latency provided better resolution among pairs of odors than did errors of discrimination. This finding demonstrated the utility of collecting response times. (iii) Latency-based similarities among odors tested previously predicted similarities among pairs not yet tested. This finding demonstrated internal/predictive validity. (iv) A signal detection model assuming a differencing strategy best described the pattern of errors. Subjects appeared to make relative judgements regarding quality. (v) Finally, latency based similarities between mixtures and their components demonstrated additivity. This finding suggested that binary mixtures fall on straight lines connecting their components in 'odor-space'. PMID- 10866985 TI - Cortical activation induced by intraoral stimulation with water in humans. AB - Studies of gustatory processing frequently utilize water as a control stimulus. However, the neural representations of intraoral stimulation with water have received little attention. We report a series of positron emission tomography studies involving intraoral stimulation with deionized distilled water. Attempting to taste water produced large, bilateral activations in insular, opercular, Rolandic and cerebellar cortices relative to resting with eyes closed or 'smelling' odorless air. The magnitude and volume of activation was substantially reduced when tasting water was contrasted with voluntary swallowing. This indicates that much of the activity induced by water reflects intraoral somatosensory or motor processing. Nevertheless, portions of the insula, operculum, post-central gyrus and cerebellum remained significantly activated in the contrast between 'tasting' water and swallowing. This activity appears to represent a specific neural correlate of fluid stimulation, and may reflect aspects of trigeminal, gustatory or thermal coding. These findings emphasize the large volume of cortex dedicated to intraoral processing, and highlight the importance of controlling for nongustatory factors in studies of gustation. PMID- 10866986 TI - Psychophysical and neurobiological evidence that the oral sensation elicited by carbonated water is of chemogenic origin. AB - The sensation produced by carbonated beverages has been attributed to chemical excitation of nociceptors in the oral cavity via the conversion of CO(2) to carbonic acid in a reaction catalyzed by carbonic anhydrase. In separate studies, we tested if the carbonic anyhdrase blocker, acetazolamide, reduced either the intensity of sensation in humans or c-fos expression by trigeminal neurons in rats, evoked by application of carbonated water to the tongue. In the psychophysical experiment, one-half of the dorsal tongue was pretreated with acetazolamide (1 or 2%), after which the tongue was exposed bilaterally to carbonated water. In a two-alternative forced-choice paradigm, subjects chose which side of the tongue yielded a stronger sensation and additionally rated the magnitude of sensation on each side. Pretreatment with acetazolamide reduced the magnitude of sensation elicited by carbonated water in a concentration-dependent manner, since a significant majority of subjects chose the untreated side of the tongue as having a stronger sensation and assigned significantly higher intensity ratings to that side. Acetazolamide did not affect the irritant sensation from citric acid, while capsaicin pretreatment reduced both the sensation elicited by carbonated water and the irritation induced by citric acid application. In a separate experiment using rats, delivery of carbonated water to the tongue significantly increased the number of cells expressing c-fos-like immunoreactivity in the dorsomedial trigeminal nucleus caudalis (versus saline controls); this was significantly reduced by pretreatment with acetazolamide. Our results support the hypothesis that carbonated water activates lingual nociceptors via conversion of CO(2) to carbonic acid; the nociceptors in turn excite trigeminal neurons involved in signaling oral irritation. PMID- 10866987 TI - Comparison of two stimulus-delivery systems for measurement of nasal pungency thresholds. AB - Using representative members of each of three homologous series of chemicals ketones, acetates and alcohols-we measured nasal pungency thresholds in anosmics via two stimulus-delivery systems. The first system consists of the fairly commonly used 270 ml, plastic 'squeeze bottles'. The second system consists of 1900 ml, glass vessels with Teflon tubing and nose-pieces. Although bulkier and more susceptible to mechanical breakage, the glass vessels possess advantages that can allow them to provide 'environmentally realistic' chemosensory thresholds, i.e. thresholds closer in absolute values to those that might be obtained under whole-body exposures. Such advantages include a larger volume of the vapor-source to accommodate whole sniffs, and a tight nose-nose-piece connection to avoid stimulus dilution. The outcome revealed that, for every chemical, the glass vessels provided nasal pungency thresholds significantly lower than those provided by the squeeze bottles. The difference amounted, on average, to a factor of 4.6, though the relative potency of the compounds remained the same under both systems. Additionally, when tested with the highest homologues used here, namely, octyl acetate and 1-octanol, anosmics using the glass vessels had little or no difficulty achieving the criterion for threshold whereas they did have difficulty when using the squeeze bottles. PMID- 10866988 TI - Perireceptor and receptor events in olfaction. Comparison of concentration and flux detectors: a modeling study. AB - Transduction in chemosensory cells begins with the association of ligand molecules to receptor proteins borne by the cell membrane. The receptor-ligand complexes formed act as signaling compounds that trigger a G-protein cascade. This receptor-ligand interaction, described here by a single-step or double-step reaction, depends on factors controlling the access of the ligand molecules to the cell membrane. Two basic mechanisms can be distinguished: concentration detectors (CD), in which the ligand can freely diffuse to the membrane, and flux detectors (FD), in which it accumulates irreversibly in a distinct perireceptor space where it is chemically deactivated. These two systems, plus their generalization, are investigated and compared. The transient and steady-state numbers of complexes are studied as a function of the external ligand concentration. The biological significance of the results is shown in a well studied example of FD, the insect sex-pheromone olfactory receptor neuron. How the number of complexes can code for the intensity of stimulation is analyzed using the size, dynamic range and sensitivity of the steady-state responses, and the time needed to reach a predefined level of the transient responses. It is shown that the FD design affords a large increase in sensitivity (a shift of the threshold response towards low concentration) with respect to the CD design, which is paid for by a lesser ability to follow fast changes in stimulus intensity. PMID- 10866989 TI - Cyclic adenosine monophosphate signaling in the rat vomeronasal organ: role of an adenylyl cyclase type VI. AB - The present study indicates that male rat urinary components in female rat vomeronasal organ microvillar preparations not only induce a rapid and transient IP(3) signal, but in addition, the level of cAMP decreases with a delayed and sustained time course. This decrease seems to be a consequence of the preceding activation of the phosphoinositol pathway rather than the result of an enhanced phosphodiesterase activity or an inhibition of adenylyl cyclase (AC) via Galpha(i) or Galpha(o). This notion is supported by the finding that activation of the endogenous protein kinase C suppresses basal as well as forskolin-induced cAMP formation. Furthermore, it was observed that elevated levels of calcium inhibit cAMP formation in rat VNO microvillar preparations. These properties of cAMP signaling in the VNO of rats may be mediated by a calcium- and protein kinase C-inhibited AC VI subtype, which is localized in microvillar preparations of the VNO. PMID- 10866990 TI - Contribution of fatty acids to olfactory host finding of female Aedes aegypti. AB - Single carbon to 18 carbon n-aliphatic carboxylic acids were tested for their attractive effects on female Aedes aegypti in a Y-tube olfactometer. Each acid was tested over a wide range of concentrations together with L-(+)-lactic acid, the indispensable synergist for other attractive components emitted from human hosts. The attractiveness of lactic acid was significantly augmented when combined with fatty acids of chain length C(1)-C(3), C(5)-C(8) and C(13)-C(18), respectively. The addition of the C(9) and C(11) acids reduced the attractive effect of lactic acid. According to experiments showing a further increase of attractiveness by adding a second fatty acid, we suggest two groups of attractive carboxylic acids: C(1)-C(3) and C(5)-C(8). The addition of a fatty acid from one group to a mixture of lactic acid and an acid from the other group augmented the attraction to the mixture. Together with ammonia, a previously demonstrated attractant for Aedes aegypti, lactic acid plus two fatty acids from the different groups formed the hitherto most attractive, artificially composed blend. Two of the carboxylic acids which were found to be attractive together with lactic acid were also tested alone and in combination with CO(2), the major attractant in human breath. In both cases no attractive effect of the carboxylic acids could be observed. PMID- 10866991 TI - Decline in taste and odor discrimination abilities with age, and relationship between gustation and olfaction. AB - It is important to learn about changes in both taste and odor perceptions with increasing age, because the taste of foods we encounter in our daily life is strongly affected by their smell. This study discusses the difference in qualitative taste and odor discrimination between the elderly and the young. Tastants and odorants used in this study were presented not as single stimuli but as a taste mixture (sucrose and tartaric acid) and an odor mixture (beta phenylethyl alcohol and gamma-undecalactone). The results showed that quality discrimination abilities of the elderly subjects for both taste and odor were significantly lower than those of the young subjects, indicating a decline in quality discrimination abilities related to age. Also, a moderate but significant correlation was observed between the taste discrimination ability and the odor discrimination ability. We measured thresholds for single-taste and odor components in mixtures and compared them between the elderly and the young to investigate the cause for these findings. PMID- 10866992 TI - Pheromone discrimination ability of olfactory bulb mitral and ruffed cells in the goldfish (Carassius auratus). AB - Significant anatomical differences characterizing mitral cells and ruffed cells were published by Kosaka and Hama in three teleost species. Physiological responses from both types of relay neurons have now been recorded extracellularly and simultaneously in the plexiform layer using a single tungsten microelectrode. During interstimulus intervals mitral cells responded with higher, frequently burst-like impulse rates triggered by the activity of epithelial receptor neurons. Ruffed cell impulse rates were low, and each action potential triggered a long-lasting, continuously variable, integrated granule cell potential. During olfactory stimulation with important biological stimuli such as preovulatory and ovulatory pheromones, a probable alarm pheromone and amino acids contrasting interactions between mitral cells and ruffed cells resulting in a drastic intensification of centrally transmitted information were frequently recorded. Individual neurons excellently discriminated stimuli. Irrespective of the physiological relevance of stimuli, however, similarities were recorded in the distribution of excitatory, inhibitory and indifferent responses. PMID- 10866993 TI - Acute regulation of Na+/H+ exchanger NHE3 by parathyroid hormone via NHE3 phosphorylation and dynamin-dependent endocytosis. AB - Parathyroid hormone (PTH) is a potent inhibitor of mammalian renal proximal tubule Na(+) transport via its action on the apical membrane Na(+)/H(+) exchanger NHE3. In the opossum kidney cell line, inhibition of NHE3 activity was detected from 5 to 45 min after PTH addition. Increase in NHE3 phosphorylation on multiple serines was evident after 5 min of PTH, but decrease in surface NHE3 antigen was not detectable until after 30 min of PTH. The decrease in surface NHE3 antigen was due to increased NHE3 endocytosis. When endocytic trafficking was arrested with a dominant negative dynamin mutant (K44A), the early inhibition (5 min) of NHE3 activity by PTH was not affected, whereas the late inhibition (30 min) and decreased surface NHE3 antigen induced by PTH were abrogated. We conclude that PTH acutely inhibits NHE3 activity in a biphasic fashion by NHE3 phosphorylation followed by dynamin-dependent endocytosis. PMID- 10866994 TI - Physiological control of smooth muscle-specific gene expression through regulated nuclear translocation of serum response factor. AB - Prolonged serum deprivation induces a structurally and functionally contractile phenotype in about 1/6 of cultured airway myocytes, which exhibit morphological elongation and accumulate abundant contractile apparatus-associated proteins. We tested the hypothesis that transcriptional activation of genes encoding these proteins accounts for their accumulation during this phenotypic transition by measuring the transcriptional activities of the murine SM22 and human smooth muscle myosin heavy chain promoters during transient transfection in subconfluent, serum fed or 7 day serum-deprived cultured canine tracheal smooth muscle cells. Contrary to our expectation, SM22 and smooth muscle myosin heavy chain promoter activities (but not viral murine sarcoma virus-long terminal repeat promoter activity) were decreased in long term serum-deprived myocytes by at least 8-fold. Because serum response factor (SRF) is a required transcriptional activator of these and other smooth muscle-specific promoters, we evaluated the expression and function of SRF in subconfluent and long term serum deprived cells. Whole cell SRF mRNA and protein were maintained at high levels in serum-deprived myocytes, but SRF transcription-promoting activity, nuclear SRF binding to consensus CArG sequences, and nuclear SRF protein were reduced. Furthermore, immunocytochemistry revealed extranuclear redistribution of SRF in serum-deprived myocytes; nuclear localization of SRF was restored after serum refeeding. These results uncover a novel mechanism for physiological control of smooth muscle-specific gene expression through extranuclear redistribution of SRF and consequent down-regulation of its transcription-promoting activity. PMID- 10866996 TI - A novel method for measurement of submembrane ATP concentration. AB - There has been considerable debate as to whether adenosine triphosphate (ATP) is compartmentalized within cells and, in particular, whether the ATP concentration directly beneath the plasma membrane, experienced by membrane proteins, is the same as that of the bulk cytoplasm. This issue has been difficult to address because there is no indicator of cytosolic ATP, such as those available for Ca(2+), capable of resolving the submembrane ATP concentration ([ATP](sm)) in real time within a single cell. We show here that mutant ATP-sensitive K(+) channels can be used to measure [ATP](sm) by comparing the increase in current amplitude on patch excision with the ATP dose-response curve. In Xenopus oocytes, [ATP](sm) was 4.6 +/- 0.3 mm (n = 29) under resting conditions, slightly higher than that measured for the bulk cytoplasm (2.3 mm). In mammalian (COSm6) cells, [ATP](sm) was slightly lower and averaged 1.4 +/- 0.1 mm (n = 66). Metabolic poisoning (10 min of 3 mm azide) produced a significant fall in [ATP](sm) in both types of cells: to 1.2 +/- 0.1 mm (n = 24) in oocytes and 0.8 +/- 0.11 mm for COSm6 cells. We conclude that [ATP](sm) lies in the low millimolar range and that there is no gradient between bulk cytosolic and submembrane [ATP]. PMID- 10866995 TI - Peroxisome proliferator-activated receptor alpha-mediated pathways are altered in hepatocyte-specific retinoid X receptor alpha-deficient mice. AB - Retinoid x receptor alpha (RXRalpha) serves as an active partner of peroxisome proliferator-activated receptor (PPARalpha). In order to dissect the functional role of RXRalpha and PPARalpha in PPARalpha-mediated pathways, the hepatocyte RXRalpha-deficient mice have been challenged with physiological and pharmacological stresses, fasting and Wy14,643, respectively. The data demonstrate that RXRalpha and PPARalpha deficiency are different in several aspects. At the basal untreated level, RXRalpha deficiency resulted in marked induction of apolipoprotein A-I and C-III (apoA-I and apoC-III) mRNA levels and serum cholesterol and triglyceride levels, which was not found in PPARalpha-null mice. Fasting-induced PPARalpha activation was drastically prevented in the absence of hepatocyte RXRalpha. Wy14,643-mediated pleiotropic effects were also altered due to the absence of hepatocyte RXRalpha. Hepatocyte RXRalpha deficiency did not change the basal acyl-CoA oxidase, medium chain acyl-CoA dehydrogenase, and malic enzyme mRNA levels. However, the inducibility of those genes by Wy14,643 was markedly reduced in the mutant mouse livers. In contrast, the basal cytochrome P450 4A1, liver fatty acid-binding protein, and apoA-I and apoC-III mRNA levels were significantly altered in the mutant mouse livers, but the regulatory effect of Wy14,643 on expression of those genes remained the same. Wy14,643-induced hepatomegaly was partially inhibited in hepatocyte RXRalpha deficient mice. Wy14,643-induced hepatocyte peroxisome proliferation was preserved in the absence of hepatocyte RXRalpha. These data suggested that in comparison to PPARalpha, hepatocyte RXRalpha has its unique role in lipid homeostasis and that the effect of RXRalpha, -beta, and -gamma is redundant in certain aspects. PMID- 10866997 TI - The POU domain transcription factor Tst-1 activates somatostatin receptor 1 gene expression in pancreatic beta -cells. AB - The peptide hormone somatostatin inhibits the release of insulin. The gene encoding somatostatin receptor 1 is expressed in pancreatic beta-cells and insulinoma RIN 1046-38 cells. In the present study the mechanisms underlying the regulation of the somatostatin receptor 1 gene in pancreatic beta-cells were investigated. Transient transfections of RIN 1046-38 cells with promoter/reporter gene constructs and footprint analysis revealed two regions, fp1 and fp2, that were necessary for the observed promoter activity. Mutagenesis of the fp2 region delineated the cis-acting element to the motif 5'-TTAATCATT-3'. The POU domain transcription factor Tst-1 was identified as trans-activator mediating the 5' TTAATCATT-3' motif-dependent transcription in RIN 1046-38 cells and heterologous CV1 cells. Tst-1, known as a transcriptional regulator in keratinocytes, glial cells, and neurons, has been detected by immunohistochemistry in pancreatic islets. Altogether, we demonstrate Tst-1 as transcriptional regulator in pancreatic neuroendocrine cells. PMID- 10866998 TI - Kinetic effects of the electrochemical proton gradient on plastoquinone reduction at the Qi site of the cytochrome b6f complex. AB - We have investigated the effects of the light-induced thylakoid transmembrane potential on the turnover of the b(6)f complex in cells of the unicellular green alga Chlamydomonas reinhardtii. The reduction of the potential by either decreasing the light intensity or by adding increasing concentrations of the ionophore carbonylcyanide p-(trifluoromethoxy)phenylhydrazone (FCCP) revealed a marked inhibition of the cytochrome b(6) oxidation rate (10-fold) without substantial modifications of cytochrome f oxidation kinetics. Partial recovery of this inhibition could be obtained in the presence of ionophores provided that the membrane potential was re-established by illumination with a train of actinic flashes fired at a frequency higher than its decay. Measurements of isotopic effects on the kinetics of cytochrome b(6) oxidation revealed a synergy between the effects of ionophores and the H(2)O-D(2)O exchange. We propose therefore, that protonation events influence the kinetics of cytochrome b(6) oxidation at the Qi site and that these reactions are strongly influenced by the light dependent generation of a transmembrane potential. PMID- 10866999 TI - Two opposing effects of non-steroidal anti-inflammatory drugs on the expression of the inducible cyclooxygenase. Mediation through different signaling pathways. AB - The efficacy of non-steroidal anti-inflammatory drugs (NSAIDs) is considered to be a result of their inhibitory effect on cyclooxygenase (COX) activity. Here, we report that flufenamic acid shows two opposing effects on COX-2 expression; it induces COX-2 expression in the colon cancer cell line (HT-29) and macrophage cell line (RAW 264.7); conversely, it inhibits tumor necrosis factor alpha (TNFalpha)- or lipopolysaccharide (LPS)-induced COX-2 expression. This inhibition correlates with the suppression of TNFalpha- or LPS-induced NFkappaB activation by flufenamic acid. The inhibitor of extracellular signal-regulated protein kinase, p38, or NFkappaB does not affect the NSAID-induced COX-2 expression. These results suggest that the NSAID-induced COX-2 expression is not mediated through activation of NFkappaB and mitogen-activated protein kinases. An activator of peroxisome proliferator-activated receptor gamma, 15-deoxy Delta(12,14)-prostaglandin J(2), also induces COX-2 expression and inhibits TNFalpha-induced NFkappaB activation and COX-2 expression. Flufenamic acid and 15 deoxy-Delta(12,14)-prostaglandin J(2) also inhibit LPS-induced expression of inducible form of nitric-oxide synthase and interleukin-1alpha in RAW 264.7 cells. Together, these results indicate that the NSAIDs inhibit mitogen-induced COX-2 expression while they induce COX-2 expression. Furthermore, the results suggest that the anti-inflammatory effects of flufenamic acid and some other NSAIDs are due to their inhibitory action on the mitogen-induced expression of COX-2 and downstream markers of inflammation in addition to their inhibitory effect on COX enzyme activity. PMID- 10867000 TI - The peroxisome proliferator-activated receptor alpha (PPARalpha) regulates bile acid biosynthesis. AB - Fibrates are a group of hypolipidemic agents that efficiently lower serum triglyceride levels by affecting the expression of many genes involved in lipid metabolism. These effects are exerted via the peroxisome proliferator-activated receptor alpha (PPARalpha). In addition, fibrates also lower serum cholesterol levels, suggesting a possible link between the PPARalpha and cholesterol metabolism. Bile acid formation represents an important pathway for elimination of cholesterol, and the sterol 12alpha-hydroxylase is a branch-point enzyme in the bile acid biosynthetic pathway, which determines the ratio of cholic acid to chenodeoxycholic acid. Treatment of mice for 1 week with the peroxisome proliferator WY-14,643 or fasting for 24 h both induced the sterol 12alpha hydroxylase mRNA in liver. Using the PPARalpha knockout mouse model, we show that the induction by both treatments was dependent on the PPARalpha. A reporter plasmid containing a putative peroxisome proliferator-response element (PPRE) identified in the rat sterol 12alpha-hydroxylase promoter region was activated by treatment with WY-14,643 in HepG2 cells, being dependent on co-transfection with a PPARalpha expression plasmid. The rat 12alpha-hydroxylase PPRE bound in vitro translated PPARalpha and retinoid X receptor alpha, albeit weakly, in electrophoretic mobility shift assay. Treatment of wild-type mice with WY-14,643 for 1 week resulted in an increased relative amount of cholic acid, an effect that was abolished in the PPARalpha null mice, verifying the functionality of the PPRE in vivo. PMID- 10867001 TI - CD95-mediated apoptosis in vivo involves acid sphingomyelinase. AB - Acid sphingomyelinase (ASM) is reported to have an essential function in stress induced apoptosis although the physiological function of ASM in receptor triggered apoptosis is unknown. Here, we delineate a pivotal role for ASM in CD95 triggered apoptosis of peripheral lymphocytes or hepatocytes in vivo. We employed intravenous injection of anti-CD4 antibodies or phytohemagglutinin that was previously shown to result in apoptosis of peripheral blood lymphocytes or hepatocytes via the endogenous CD95/CD95 ligand system. Our results demonstrate a high susceptibility in normal mice whereas ASM knock-out mice fail to immunodeplete T cells or develop autoimmune-like hepatitis. Likewise, ASM deficient mice or hepatocytes and splenocytes ex vivo manifest resistance to anti CD95 treatment. These results provide in vivo evidence for an important physiological function of ASM in CD95-induced apoptosis. PMID- 10867002 TI - Length increase of the human alpha -globin 3'-untranslated region disrupts stability of the pre-mRNA but not that of the mature mRNA. AB - Polyadenylation increases the stability of mRNA molecules. By studying the effect of the length of 3'-untranslated region (UTR) on mRNA levels, we have found that alpha-globin pre-mRNA is stabilized by a mechanism that does not modulate the half-life of mature mRNA. The insertion of DNA fragments of various unrelated sequences into the 3'-UTR of the human alpha-globin gene strongly reduces mRNA abundance upon transfection into choriocarcinoma JEG-3 cells. We found an inverse relationship between mRNA levels and the length of the introduced fragments. In fact, mRNA levels as low as 1% were observed after inserting a 477-nucleotide (nt) fragment, whereas inserting a fragment of 86 nt at the same position had no effect on mRNA accumulation. DNA insertion induced no change in transcription rate or in half-life of mature mRNA. Semi-quantitative reverse transcription polymerase chain reaction revealed that inserting a 477-nt fragment in the 3'-UTR resulted in decreased levels of nuclear pre-mRNA in proportion to that observed for mature mRNA. In contrast, the insertion of the 477-nt exogenous DNA in the last intron had no effect on mRNA levels despite the presence of intronic sequences in the pre-mRNA. This shows that the reduction of pre-mRNA level was not due to the insertion of putative ribonuclease cleavage sites or the insertion of a segment DNA that reduces the elongation efficiency. Taken together, our results strongly support the existence of a pre-mRNA stabilizing mechanism that can be disrupted by increasing the length of the 3'-UTR. The fact that the half life of mature mRNA is not affected by DNA insertion is compatible with a pre mRNA-specific stabilizing mechanism that acts specifically before polyadenylation. PMID- 10867003 TI - Long term depression in the CA1 field is associated with a transient decrease in pre- and postsynaptic PKC substrate phosphorylation. AB - Induction of homosynaptic long term depression (LTD) in the CA1 field of the hippocampus is thought to require activation of N-methyl-d-aspartate receptors, an elevation of postsynaptic Ca(2+) levels, and a subsequent increase in phosphatase activity. To investigate the spatial and temporal changes in protein phosphatase activity following LTD induction, we determined the in situ phosphorylation state of a pre- (GAP-43/B-50) and postsynaptic (RC3) protein kinase C substrate during N-methyl-d-aspartate receptor-dependent LTD in the CA1 field of rat hippocampal slices. We show that LTD is associated with a transient (<30 min) and D-AP5-sensitive reduction in GAP-43/B-50 and RC3 phosphorylation and that LTD is prevented by the phosphatase inhibitors okadaic acid and cyclosporin A. Our data provide strong evidence for a transient increase in pre- and postsynaptic phosphatase activity during LTD. Since the in situ phosphorylation of the calmodulin-binding proteins GAP-43/B-50 and RC3 changes during both LTD and long term potentiation, these proteins may form part of the link between the Ca(2+) signal and Ca(2+)/calmodulin-dependent processes implicated in long term potentiation and LTD. PMID- 10867004 TI - Profilin is required for sustaining efficient intra- and intercellular spreading of Shigella flexneri. AB - The ability of Shigella to mediate actin-based motility within the host cell is a prominent pathogenic feature of bacillary dysentery. The ability is dependent on the interaction of VirG with neural Wiskott-Aldrich syndrome protein (N-WASP), which in turn mediates recruitment of Arp2/3 complex and several actin-related proteins. In the present study, we show that profilin I is essential to the rapid movement of Shigella in epithelial cells, for which the capacity of profilin to interact with G-actin and N-WASP is critical. In COS-7 cells overexpressing either mutated profilin H119E, which failed to bind G-actin, or H133S, which is unable to interact with poly-l-proline, Shigella motility was significantly inhibited. Similarly, depletion of profilin from Xenopus egg extracts resulted in a decrease in bacterial motility that was completely rescued by adding back profilin I but not H119E or H133S. In COS-7 cells overexpressing a N-WASP mutant lacking the proline-rich domain (Deltap) unable to interact with profilin, the actin tail formation of intracellular Shigella was inhibited. In N-WASP-depleted extracts, addition of Deltap but not full-length N-WASP was unable to restore the bacterial motility. Furthermore, in a plaque formation assay with Madin-Darby canine kidney cell monolayers infected by Shigella, Madin-Darby canine kidney cells stably expressing H119E, H133S, or Deltap reduced the bacterial cell-to cell spreading. These results indicate that profilin I associated with N-WASP is an essential host factor for sustaining efficient intra- and intercellular spreading of Shigella. PMID- 10867005 TI - Platelet-activating factor (PAF)-dependent transacetylase and its relationship with PAF acetylhydrolases. AB - Platelet-activating factor (PAF)-dependent transacetylase (TA) is an enzyme that transfers an acetyl group from PAF to acceptor lipids such as lysophospholipids and sphingosine. This enzyme is distributed in membrane and cytosol of the cells. We previously revealed that TA purified from rat kidney membrane showed an amino acid sequence similarity to that of bovine PAF-acetylhydrolase (AH) (II). In the present study, we purified TA from the rat kidney cytosol and analyzed its amino acid sequence. The amino acid sequence of the cytosolic TA is similar to that of bovine PAF-AH (II) and membrane TA. To clarify the relationship between TA and PAF-AH (II), we isolated cDNA of rat PAF-AH (II). The predicted amino acid sequence of rat PAF-AH (II) from isolated cDNA included all the sequences found in TAs purified from the membrane and cytosolic TAs. In addition, monoclonal antibody to recombinant PAF-AH (II) cross-reacted with both cytosolic and membrane TAs. Consistent with sequence identity, recombinant PAF-AH (II) showed TA activity, whereas recombinant PAF-AH Ib, which is a different subtype of intracellular PAF-AHs, did not possess TA activity. Analysis of a series of site directed mutant PAF-AH (II) proteins showed that TA activity was decreased, whereas PAF-AH activity was not affected in C120S and G2A mutant proteins. Thus, Cys(120) and Gly(2) are implicated in the catalysis of TA reaction in this enzyme. Furthermore, the transfer of acetate from PAF to endogenous acceptor lipids was significantly increased in a time-dependent manner in CHO-K1 cells transfected with PAF-AH (II) gene. These results demonstrate that PAF-AH (II) can function, as a TA in intact cells, and PAF-AH (II) and TA are the same enzyme. PMID- 10867006 TI - Sequence-specific high mobility group box factors recognize 10-12-base pair minor groove motifs. AB - Sequence-specific high mobility group (HMG) box factors bind and bend DNA via interactions in the minor groove. Three-dimensional NMR analyses have provided the structural basis for this interaction. The cognate HMG domain DNA motif is generally believed to span 6-8 bases. However, alignment of promoter elements controlled by the yeast genes ste11 and Rox1 has indicated strict conservation of a larger DNA motif. By site selection, we identify a highly specific 12-base pair motif for Ste11, AGAACAAAGAAA. Similarly, we show that Tcf1, MatMc, and Sox4 bind unique, highly specific DNA motifs of 12, 12, and 10 base pairs, respectively. Footprinting with a deletion mutant of Ste11 reveals a novel interaction between the 3' base pairs of the extended DNA motif and amino acids C-terminal to the HMG domain. The sequence-specific interaction of Ste11 with these 3' base pairs contributes significantly to binding and bending of the DNA motif. PMID- 10867007 TI - Superoxide reductase as a unique defense system against superoxide stress in the microaerophile Treponema pallidum. AB - Aerobic life requires the presence of antioxidant enzymes, such as superoxide dismutase, catalase, and peroxidase to eliminate deleterious oxygen derivatives. Treponema pallidum, a microaerophilic bacterium responsible for venereal syphilis, is an interesting organism because it lacks all of the above-mentioned enzymes, as deduced from its recently sequenced genome. In this paper, we describe a gene in T. pallidum with sequence homologies to a new class of antioxidant systems, named superoxide reductases, recently isolated from sulfate reducing bacteria (Lombard, M., Fontecave, M., Touati, D., and Niviere, V. (2000) J. Biol. Chem. 275, 115-121). We report that (i) expression of the T. pallidum gene fully restored to a superoxide dismutase-deficient Escherichia coli mutant the ability to grow under aerobic conditions; (ii) the corresponding protein displays a strong superoxide reductase activity; and (iii) the T. pallidum protein contains only one mononuclear nonheme ferrous center, able to reduce superoxide selectively and efficiently, whereas previously characterized superoxide reductase from Desulfoarculus baarsii contains an additional rubredoxin-like ferric center. These results suggest that T. pallidum antioxidant defenses rely on a new class of superoxide reductase and raise the question of the importance of superoxide reductases in mechanisms for detoxifying superoxide radicals. PMID- 10867008 TI - Molecular characterization of the first two enzymes of the pentose-phosphate pathway of Trypanosoma brucei. Glucose-6-phosphate dehydrogenase and 6 phosphogluconolactonase. AB - Trypanosomatids are parasitic protists that have part of their glycolytic pathway sequestered inside peroxisome-like organelles: the glycosomes. So far, at least one enzyme of the pentose-phosphate pathway has been found to be associated partially with glycosomes. Here, we describe how two genes from Trypanosoma brucei, coding for the first two enzymes of the pentose-phosphate pathway, i.e. glucose-6-phosphate dehydrogenase and 6-phosphogluconolactonase, were identified by in silico screening of trypanosome genome project data bases. These genes were cloned and sequenced. Analysis of the lactonase sequence revealed that it contained a C-terminal peroxisome targeting signal in agreement with its subcellular localization in the bloodstream form trypanosome (15% glycosomal and 85% cytosolic). However, the dehydrogenase sequence did not reveal any targeting signal, despite its localization inside glycosomes. The corresponding enzymes have been overexpressed in Escherichia coli and purified, and their biochemical characteristics have been determined. PMID- 10867009 TI - DNA binding specificity studies of four ETS proteins support an indirect read-out mechanism of protein-DNA recognition. AB - Members of the ETS family of transcription factors are involved in several developmental and physiological processes, and, when overexpressed or misexpressed, can contribute to a variety of cancers. Each family member has a conserved DNA-binding domain that recognizes DNA sequences containing a G-G-A trinucleotide. Discrimination between potential ETS-binding sites appears to be governed by both the nucleotides flanking the G-G-A sequence and protein-protein interactions. We have used an adaptation of the "length-encoded multiplex" approach (Desjarlais, J. R., and Berg, J. M. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 11099-11103) to define DNA binding specificities for four ETS proteins: Fli-1, SAP-1, PU.1, and TEL. Our results support a model in which cooperative effects among neighboring bases flanking the central G-G-A site contribute to the formation of stable ETS/DNA complexes. These results are consistent with a mechanism for specific DNA binding that is partially governed by an indirect read out of the DNA sequence, in which a sequence-specific DNA conformation is sensed or induced. PMID- 10867010 TI - Hierarchy of post-translational modifications involved in the circulatory longevity of glycoproteins. Demonstration of concerted contributions of glycan sialylation and subunit assembly to the pharmacokinetic behavior of bovine acetylcholinesterase. AB - The tetrameric form of native serum-derived bovine acetylcholinesterase is retained in the circulation for much longer periods (mean residence time, MRT = 1390 min) than recombinant bovine acetylcholinesterase (rBoAChE) produced in the HEK-293 cell system (MRT = 57 min). Extensive matrix-assisted laser desorption ionization-time of flight analyses established that the basic structures of the N glycans associated with the native and recombinant enzymes are similar (the major species (50-60%) are of the biantennary fucosylated type and 20-30% are of the triantennary type), yet the glycan termini of the native enzyme are mostly capped with sialic acid (82%) and alpha-galactose (12%), whereas glycans of the recombinant enzyme exhibit a high level of exposed beta-galactose residues (50%) and a lack of alpha-galactose. Glycan termini of both fetal bovine serum and rBoAChE were altered in vitro using exoglycosidases and sialyltransferase or in vivo by a HEK-293 cell line developed specifically to allow efficient sialic acid capping of beta-galactose-exposed termini. In addition, the dimeric and monomeric forms of rBoAChE were quantitatively converted to tetramers by complexation with a synthetic peptide representing the human ColQ-derived proline-rich attachment domain. Thus by controlling both the level and nature of N-glycan capping and subunit assembly, we generated and characterized 9 distinct bovine AChE glycoforms displaying a 400-fold difference in their circulatory lifetimes (MRT = 3.5-1390 min). This revealed some general rules and a hierarchy of post translation factors determining the circulatory profile of glycoproteins. Accordingly, an rBoAChE was generated that displayed a circulatory profile indistinguishable from the native form. PMID- 10867011 TI - Signal transducer and activator of transcription 3 activates CCAAT enhancer binding protein delta gene transcription in G0 growth-arrested mouse mammary epithelial cells and in involuting mouse mammary gland. AB - The CCAAT enhancer-binding protein (C/EBP) family of transcription factors is implicated in the regulation of cell proliferation and differentiation in a variety of tissues. C/EBPdelta is involved in regulating G(0) growth arrest and apoptosis of mouse mammary epithelial cells. This study shows that activation of signal transducer and activator of transcription 3 (Stat3), but not activation of Stat1 or Stat5, occurs concurrently with G(0) growth arrest of HC11 mouse mammary epithelial cells, but not NIH 3T3 fibroblasts. Promoter analysis demonstrates that the C/EBPdelta promoter fragment involved in transcriptional activation during G(0) growth arrest contains a Stat3 binding site and that mutation of this site eliminates the G(0) growth arrest inducibility of the C/EBPdelta promoter. Overexpression of Stat3 increases C/EBPdelta promoter activity during G(0) growth arrest of HC11 cells, whereas dominant negative Stat3 decreases C/EBPdelta promoter activity under the same conditions. Neither Stat3 overexpression nor dominant negative Stat3 expression influences C/EBPdelta promoter activity in growing HC11 cells or G(0) growth-arrested NIH3T3 cells, demonstrating that the effect is specific to G(0) growth arrest of mammary epithelial cells. Band shift assays and antibody interference assays demonstrate specific binding of Stat3 to the acute phase response element in the C/EBPdelta promoter in G(0) growth arrested HC11 cell extracts and 24 h involuting mouse mammary gland extracts. These data indicate that Stat3 activates C/EBPdelta transcription in G(0) growth arrested mouse mammary epithelial cells and binds to the C/EBPdelta promoter during involution. An autocrine mechanism of Stat3 activation is proposed. PMID- 10867012 TI - A single amino acid change in subunit 6 of the yeast mitochondrial ATPase suppresses a null mutation in ATP10. AB - In an earlier study, the ATP10 gene of Saccharomyces cerevisiae was shown to code for an inner membrane protein required for assembly of the F(0) sector of the mitochondrial ATPase complex (Ackerman, S., and Tzagoloff, A. (1990) J. Biol. Chem. 265, 9952-9959). To gain additional insights into the function of Atp10p, we have analyzed a revertant of an atp10 null mutant that displays partial recovery of oligomycin-sensitive ATPase and of respiratory competence. The suppressor mutation in the revertant has been mapped to the OLI2 locus in mitochondrial DNA and shown to be a single base change in the C-terminal coding region of the gene. The mutation results in the substitution of a valine for an alanine at residue 249 of subunit 6 of the ATPase. The ability of the subunit 6 mutation to compensate for the absence of Atp10p implies a functional interaction between the two proteins. Such an interaction is consistent with evidence indicating that the C-terminal region with the site of the mutation and the extramembrane domain of Atp10p are both on the matrix side of the inner membrane. Subunit 6 has been purified from the parental wild type strain, from the atp10 null mutant, and from the revertant. The N-terminal sequences of the three proteins indicated that they all start at Ser(11), the normal processing site of the subunit 6 precursor. Mass spectral analysis of the wild type and mutants subunit 6 failed to reveal any substantive difference of the wild type and mutant proteins when the mass of the latter was corrected for Ala --> Val mutation. These data argue against a role of Atp10p in post-translational modification of subunit 6. Although post-translational modification of another ATPase subunit interacting with subunit 6 cannot be excluded, a more likely function for Atp10p is that it acts as a subunit 6 chaperone during F(0) assembly. PMID- 10867013 TI - Activated Raf inhibits myogenesis through a mechanism independent of activator protein 1-mediated myoblast transformation. AB - Skeletal myogenesis is acutely affected by growth factors and subsequent activation of their respective intracellular signaling cascades. Components of the mitogenic Ras/Raf/mitogen-activated protein kinase (MAPK) signaling module are potent inhibitors of myoblast differentiation. However, the means by which these kinases prevent myocyte formation and activation of the muscle gene program is unknown. Activator protein 1 (AP-1) is a transcriptional regulator the actions of which are up-regulated by signaling events, including elevated MAPK. Because activated Raf inhibits avian myogenesis in a MAPK-dependent fashion, we investigated the role of AP-1 as a mediator of the Raf-imposed block to myogenesis. Avian myoblasts overexpressing activated Raf contain elevated levels of AP-1 DNA binding and transcriptional activity. Introduction of an AP-1 dominant inhibitory protein (AFOS) into Raf-expressing myoblasts prevented acquisition of a transformed morphology. Interestingly, these cells remained differentiation-defective. Myogenic cells cotransduced with RCAS(A)-Raf BXB and RCAS(B)-AFOS remained mononuclear and myosin-negative and did not activate significantly muscle-specific reporter genes. These results argue that Raf inhibits muscle differentiation independent of AP-1-mediated cell transformation. Our results provide evidence for AP-1 as a critical component of the transforming capacity of activated Raf and evidence that AP-1 is not involved in the myogenic inhibitory effects of the kinase. PMID- 10867014 TI - Oxidation of either methionine 351 or methionine 358 in alpha 1-antitrypsin causes loss of anti-neutrophil elastase activity. AB - Hydrogen peroxide is a component of cigarette smoke known to be essential for inactivation of alpha(1)-antitrypsin, the primary inhibitor of neutrophil elastase. To establish the molecular basis of the inactivation of alpha(1) antitrypsin, we determined the sites oxidized by hydrogen peroxide. Two of the nine methionines were particularly susceptible to oxidation. One was methionine 358, whose oxidation was known to cause loss of anti-elastase activity. The other, methionine 351, was as susceptible to oxidation as methionine 358. Its oxidation also resulted in loss of anti-elastase activity, an effect not previously recognized. The equal susceptibility of methionine 358 and methionine 351 to oxidation was confirmed by mass spectrometry. To verify this finding, we produced recombinant alpha(1)-antitrypsins in which one or both of the susceptible methionines were mutated to valine. M351V and M358V were not as rapidly inactivated as wild-type alpha1-antitrypsin, but only the double mutant M351V/M358V was markedly resistant to oxidative inactivation. We suggest that inactivation of alpha(1)-antitrypsin by oxidation of either methionine 351 or 358 provides a mechanism for regulation of its activity at sites of inflammation. PMID- 10867015 TI - Homodimerization via a leucine zipper motif is required for enzymatic activity of quiescent cell proline dipeptidase. AB - Quiescent cell proline dipeptidase (QPP) is an intracellular serine protease that is also secreted upon cellular activation. This enzyme cleaves N-terminal Xaa-Pro dipeptides from proteins, an unusual substrate specificity shared with dipeptidyl peptidase IV (CD26/DPPIV). QPP is a 58-kDa protein that elutes as a 120-130-kDa species from gel filtration, indicating that it forms a homodimer. We analyzed this dimerization with in vivo co-immunoprecipitation assays. The amino acid sequence of QPP revealed a putative leucine zipper motif, and mutational analyses indicated that this leucine zipper is required for homodimerization. The leucine zipper mutants showed a complete lack of enzymatic activity, suggesting that homodimerization is important for QPP function. On the other hand, an enzyme active site mutant retained its ability to homodimerize. These data are the first to demonstrate a role for a leucine zipper motif in a proteolytic enzyme and suggest that leucine zipper motifs play a role in mediating dimerization of a diverse array of proteins. PMID- 10867016 TI - Structural characterization of the fibroblast growth factor-binding protein purified from bovine prepartum mammary gland secretion. AB - A novel heparin-binding protein was purified to homogeneity from bovine prepartum mammary gland secretion using heparin-Sepharose chromatography and reverse-phase high performance liquid chromatography successively. Structural information obtained by N-terminal amino acid sequencing of a series of proteolytically generated peptides permitted the cloning of the corresponding cDNA. The isolated cDNA was 1170 base pairs long and consisted of an 83-base pair 5'-untranslated region followed by a 702-base pair coding region and a 385-base pair 3' untranslated region. The open reading frame resulted in a protein comprising 234- amino acid residues, including a signal sequence. Instead of Lys(24) as the predicted N terminus, Edman degradation of the native protein revealed N-terminal processing at two sites as follows: a primary site between Arg(31)-Gly(32) and a secondary site between Arg(51)-Ser(52). The amino acid sequence showed a significant similarity with that of human (60%) and mouse (53%) fibroblast growth factor-binding protein (FGF-BP). Accordingly, ligand blotting experiments revealed that bovine FGF-BP bound FGF-2. The theoretical mass of the protein predicted from the cDNA sequence is 22.5 kDa. However, the molecular mass of the purified protein was estimated to 28.6 kDa by mass spectrometry and 36 kDa by electrophoresis. The apparent molecular weight differences are most likely due to post-transcriptional modifications, shown to involve N- and O-glycosylation of Asn(155) and Ser(172), respectively. All 10 cysteine residues in the protein participated in disulfide bonds, and the pattern was identified as Cys(71) Cys(88), Cys(97)-Cys(130), Cys(106)-Cys(142), Cys(198)-Cys(234), and Cys(214) Cys(222). As the 10 cysteines of the three known FGF-BPs are positionally conserved, the disulfide bond pattern of bovine FGF-BP may be regarded as representative for the FGF-BP family. PMID- 10867017 TI - The hematopoietic transcription factor PU.1 represses gelatinase A transcription in glomerular mesangial cells. AB - The matrix metalloproteinase gelatinase A plays a key role in the evolution of glomerular injury and is a major contributing factor to the development of glomerulosclerosis. Prior studies have focused on a potent cis-acting enhancer element located in the near 5'-flanking region of the rat and human gelatinase A genes (Harendza, S., Pollock, A. S., Mertens, P. R., and Lovett, D. H. (1995) J. Biol. Chem. 270, 18286-18796; Mertens, P. R., Alfonso-Jaume, M. A., Steinmann, K., and Lovett, D. H. (1999) J. Am. Soc. Nephrol. 10, 2480-2487). Given the combinatorial nature of transcriptional regulation, we examined additional regions of the 5'-flanking region of the rat gelatinase A gene to identify further regulatory elements. In this study the identification of a silencing element located between -1903 and -1847 base pairs of the 5'-flanking region of the rat gelatinase A gene is reported. Sequence analysis, electrophoretic mobility studies, and transfection experiments demonstrate that a specific binding sequence for the hematopoietic transcription factor PU.1 is present within the silencing sequence. PU.1 activity is absolutely required for the expression of silencing activity within the context of transfected glomerular mesangial cells. Western blots identify the PU.1 protein within nuclear extracts of mesangial cells, and cotransfection with a PU.1 expression vector directly augments silencing activity. These studies underscore the complex patterns of gelatinase A transcriptional regulation and also strongly suggest that glomerular mesangial cells are ultimately derived from bone marrow cells. PMID- 10867018 TI - Regulation of protein kinase D by multisite phosphorylation. Identification of phosphorylation sites by mass spectrometry and characterization by site-directed mutagenesis. AB - Activation of the serine/threonine kinase, protein kinase D (PKD/PKC mu) via a phorbol ester/PKC-dependent pathway involves phosphorylation events. The present study identifies five in vivo phosphorylation sites by mass spectrometry, and the role of four of them was investigated by site-directed mutagenesis. Four sites are autophosphorylation sites, the first of which (Ser(916)) is located in the C terminus; its phosphorylation modifies the conformation of the kinase and influences duration of kinase activation but is not required for phorbol ester mediated activation of PKD. The second autophosphorylation site (Ser(203)) lies in that region of the regulatory domain, which in PKC mu interacts with 14-3 3tau. The last two autophosphorylation sites (Ser(744) and Ser(748)) are located in the activation loop but are only phosphorylated in the isolated PKD-catalytic domain and not in the full-length PKD; they may affect enzyme catalysis but are not involved in the activation of wild-type PKD by phorbol ester. We also present evidence for proteolytic activation of PKD. The fifth site (Ser(255)) is transphosphorylated downstream of a PKC-dependent pathway after in vivo stimulation with phorbol ester. In vivo phorbol ester stimulation of an S255E mutant no longer requires PKC-mediated events. In conclusion, our results show that PKD is a multisite phosphorylated enzyme and suggest that its phosphorylation may be an intricate process that regulates its biological functions in very distinct ways. PMID- 10867019 TI - Protein kinase C (PKC) inhibits fas receptor-induced apoptosis through modulation of the loss of K+ and cell shrinkage. A role for PKC upstream of caspases. AB - Cell shrinkage and loss of intracellular K(+) are early requisite features for the activation of effector caspases and apoptotic nucleases in Fas receptor mediated apoptosis of Jurkat cells, although the mechanisms responsible for both process remain unclear (Bortner, C. D., Hughes, F. M., Jr., and Cidlowski, J. A. (1997) J. Biol. Chem. 272, 32436-32442). We have now investigated the role of protein kinase C (PKC)-dependent signaling in the regulation of Fas-induced cell shrinkage and loss of K(+) during apoptosis. Anti-Fas induced cell shrinkage was blocked during PKC stimulation by the phorbol ester 12-O-tetradecanoylphorbol-3 acetate (PMA) and by bryostatin-1. Conversely, inhibition of PKC with Go6976, enhanced the anti-Fas-mediated loss of cell volume. Analyses of mitochondrial membrane potential and DNA fragmentation revealed that the PKC-mediated effect observed in cell volume is propagated to these late features of apoptosis. Flow cytometric analyses and (86)Rb efflux experiments revealed that a primary effect of PKC appears to be on the modulation of Fas-induced K(+) efflux, since both PMA and bryostatin-1 inhibited extrusion of K(+) that occurs during Fas-mediated cell death, and Go6976 exacerbated the effect of anti-Fas. Interestingly, high extracellular K(+) significantly blocked the effect of anti-Fas alone or anti-Fas combined with Go6976, suggesting an underlying effect of PKC on K(+) loss. Western blot analyses showed the caspase-dependent proteolysis of PKC isotypes delta, epsilon, and theta in whole cell extracts from anti-Fas treated Jurkat T cells. However, stimulation of PKC by PMA or bryostatin-1 prevented this isotypic specific PKC cleavage during apoptosis, providing further evidence that PKC itself exerts an upstream signal in apoptosis and controls the caspase-dependent proteolytic degradation of PKC isotypes. Finally, we show that PMA or bryostatin 1 prevents the activation of caspase-3 and caspase-8. Thus, this study shows that the protective effect that PKC stimulation exerts in the Fas-mediated apoptotic pathway occurs at a site upstream of caspases-3 and -8. PMID- 10867020 TI - Roles of the heparin and low density lipid receptor-related protein-binding sites of protease nexin 1 (PN1) in urokinase-PN1 complex catabolism. The PN1 heparin binding site mediates complex retention and degradation but not cell surface binding or internalization. AB - We have previously described thrombin (Th)-protease nexin 1 (PN1) inhibitory complex binding to cell surface heparins and subsequent low density lipid receptor-related protein (LRP)-mediated internalization. Our present studies examine the catabolism of urinary plasminogen activator (uPA)-PN1 inhibitory complexes, which, unlike Th.PN1 complexes, bind almost exclusively through the uPA receptor. In addition, the binding site in PN1 required for the LRP-mediated internalization of Th.PN1 complexes is not required for the LRP-mediated internalization of uPA.PN1 complexes. Thus, the protease moiety of the complex partially determines the mechanistic route of entry. Because cell surface heparins are only minimally involved in the binding and internalization of uPA.PN1 complexes, we then predicted that complexes between uPA and the heparin binding-deficient PN1 variant, PN1(K7E), should be catabolized at the same rate as complexes formed with native PN1. Surprisingly, the uPA.PN1(K7E) complexes were degraded at only a fraction of the rate of native complexes. Internalization studies revealed that both uPA. PN1(K7E) and native uPA.PN1 complexes were initially internalized at the same rate, but uPA.PN1(K7E) complexes were rapidly retro-endocytosed in an intact form. By examining the pH dependence of complex binding in the range of 4.0-7.0, it was determined that the uPA.PN1 inhibitory complexes must specifically bind to endosomal heparins at pH 5.5 to be retained and sorted to lysosomes. These studies are the first to document a role for heparins in the catabolism of SERPIN-protease complexes at a point further in the pathway than cell surface binding, and this role may extend to other heparin binding LRP-internalized ligands. PMID- 10867021 TI - Protein-tyrosine kinase Pyk2 is involved in interleukin-2 production by Jurkat T cells via its tyrosine 402. AB - We established Jurkat transfectants that overexpress Pyk2 or its mutants, K457A (lysine 457 was mutated to alanine), Pyk2-Y402F (tyrosine 402 to phenylalanine), and Pyk2-Y881F to investigate the role of Pyk2 in T cell activation. Pyk2 as well as kinase-inactive Pyk2-K457A, was phosphorylated at tyrosine residues 402, 580, and 881 upon T cell antigen receptor cross-linking, indicating that these residues are phosphorylated by other tyrosine kinase(s). However, no tyrosine phosphorylation of Pyk2-Y402F was detected while more than 60% of the tyrosine phosphorylation was observed in Pyk2-Y881F. Pyk2-Y402F inhibited the activation of endogenous Pyk2. The degree of activation of both c-Jun NH(2)-terminal kinase and p38 mitogen-activated protein kinase but not extracellular signal-regulated protein kinase after concurrent ligation of T cell antigen receptor and CD28 was reduced by more than 50% in the clones expressing Pyk2-Y402F. Consistent with this inhibition, IL-2 production was significantly diminished in the Pyk2-Y402F expressing clones. Furthermore, we found that Pyk2, when overexpressed, associates with Zap70 and Vav. Taken together, these findings suggest that Pyk2 is involved in the activation of T cells through its tyrosine 402. PMID- 10867022 TI - Glucocorticoids induce proteasome C3 subunit expression in L6 muscle cells by opposing the suppression of its transcription by NF-kappa B. AB - Muscle wasting in catabolic conditions results from activation of the ubiquitin proteasome proteolytic pathway by a process that requires glucocorticoids and is generally associated with increased levels of mRNAs encoding components of this proteolytic system. In L6 muscle cells, dexamethasone stimulates proteolysis and increases the amount of the proteasome C3 subunit protein by augmenting its transcription. Transfection studies with human C3 promoter-luciferase reporter genes and electrophoretic mobility shift assays revealed that a NF-kappaB.protein complex containing Rel A is abundant in L6 muscle cell nuclei. Glucocorticoids stimulate C3 subunit expression by antagonizing the interaction of this NF-kappaB protein with an NF-kappaB response element in the C3 subunit promoter region. Dexamethasone also increased the cytosolic amounts of the NF-kappaB p65 subunit and the IkappaBalpha inhibitor proteins in L6 cells. Incubation of L6 cells with a cytokine mixture not only increased the amount of activated NF-kappaB but also decreased C3 promoter activity and lowered endogenous C3 subunit mRNA. Thus, NF kappaB is a repressor of C3 proteasome subunit transcription in muscle cells, and glucocorticoids stimulate C3 subunit expression by opposing this suppressor action. PMID- 10867023 TI - Kinetic, thermodynamic, and developmental consequences of deleting creatine kinase isoenzymes from the heart. Reaction kinetics of the creatine kinase isoenzymes in the intact heart. AB - Creatine kinase (CK) exists as a family of isoenzymes in excitable tissue. We studied isolated perfused hearts from mice lacking genes for either the main muscle isoform of CK (M-CK) or both M-CK and the main mitochondrial isoform (Mt CK) to determine 1) the biological significance of CK isoenzyme shifts, 2) the necessity of maintaining a high CK reaction rate, and 3) the role of CK isoenzymes in establishing the thermodynamics of ATP hydrolysis. (31)P NMR was used to measure [ATP], [PCr], [P(i)], [ADP], pH, as well as the unidirectional reaction rate of PCr--> [gamma-P]ATP. Developmental changes in the main fetal isoform of CK (BB-CK) were unaffected by loss of other CK isoenzymes. In hearts lacking both M- and Mt-CK, the rate of ATP synthesis from PCr was only 9% of the rate of ATP synthesis from oxidative phosphorylation demonstrating a lack of any high energy phosphate shuttle. We also found that the intrinsic activities of the BB-CK and the MM-CK isoenzymes were equivalent. Finally, combined loss of M- and Mt-CK (but not loss of only M-CK) prevented the amount of free energy released from ATP hydrolysis from increasing when pyruvate was provided as a substrate for oxidative phosphorylation. PMID- 10867024 TI - Ligand discrimination in signaling through an ErbB4 receptor homodimer. AB - The epidermal growth factor (EGF)-like family of growth factors elicits cellular responses by stimulating the dimerization, autophosphorylation, and tyrosine kinase activities of the ErbB family of receptor tyrosine kinases. Although several different EGF-like ligands are capable of binding to a single ErbB family member, it is generally thought that the biological and biochemical responses of a single receptor dimer to different ligands are indistinguishable. To test whether an ErbB receptor dimer is capable of discriminating among ligands we have examined the effect of four EGF-like growth factors on signaling through the ErbB4 receptor homodimer in CEM/HER4 cells, a transfected human T cell line ectopically expressing ErbB4 in an ErbB-null background. Despite stimulating similar levels of gross receptor tyrosine phosphorylation, the EGF-like growth factors betacellulin, neuregulin-1beta, neuregulin-2beta, and neuregulin-3 exhibited different biological potencies in a cellular growth assay. Moreover, the different ligands induced different patterns of recruitment of intracellular signaling proteins to the activated receptor and induced differential usage of intracellular kinase signaling cascades. Finally, two-dimensional phosphopeptide mapping of ligand-stimulated ErbB4 revealed that the different growth factors induce different patterns of receptor tyrosine phosphorylation. These results indicate that ErbB4 activation by growth factors is not generic and suggest that individual ErbB receptors can discriminate between different EGF-like ligands within the context of a single receptor dimer. More generally, our observations significantly modify our understanding of signaling through receptor tyrosine kinases and point to a number of possible models for ligand-mediated signal diversification. PMID- 10867025 TI - Uptake of lipoproteins for axonal growth of sympathetic neurons. AB - Lipoproteins originating from axon and myelin breakdown in injured peripheral nerves are believed to supply cholesterol to regenerating axons. We have used compartmented cultures of rat sympathetic neurons to investigate the utilization of lipids from lipoproteins for axon elongation. Lipids and proteins from human low density lipoproteins (LDL) and high density lipoproteins (HDL) were taken up by distal axons and transported to cell bodies, whereas cell bodies/proximal axons internalized these components from only LDL, not HDL. Consistent with these observations, the impairment of axonal growth, induced by inhibition of cholesterol synthesis, was reversed when LDL or HDL were added to distal axons or when LDL, but not HDL, were added to cell bodies. LDL receptors (LDLRs) and LR7/8B (apoER2) were present in cell bodies/proximal axons and distal axons, with LDLRs being more abundant in the former. Inhibition of cholesterol biosynthesis increased LDLR expression in cell bodies/proximal axons but not distal axons. LR11 (SorLA) was restricted to cell bodies/proximal axons and was undetectable in distal axons. Neither the LDL receptor-related protein nor the HDL receptor, SR B1, was detected in sympathetic neurons. These studies demonstrate for the first time that lipids are taken up from lipoproteins by sympathetic neurons for use in axonal regeneration. PMID- 10867026 TI - Glucocorticoids inhibit developmental stage-specific osteoblast cell cycle. Dissociation of cyclin A-cyclin-dependent kinase 2 from E2F4-p130 complexes. AB - Unique cell cycle control is instituted in confluent osteoblast cultures, driving growth to high density. The postconfluent dividing cells share features with cells that normally exit the cell cycle; p27(kip1) is increased, p21(waf1/cip1) is decreased, free E2F DNA binding activity is reduced, and E2F4 is primarily nuclear. E2F4-p130 becomes the predominant E2F-pocket complex formed on E2F sites, but, unlike the complex that typifies resting cells, cyclin A and CDK2 are also present. Administration of dexamethasone at this, but not earlier stages, results in reduction of cyclin A and CDK2 levels with a parallel decrease in the associated kinase activity, dissociation of cyclin A-CDK2 from the E2F4-p130 complexes, and inhibition of G(1)/S transition. The glucocorticoid-mediated cell cycle attenuation is also accompanied by, but not attributable to, increased p27(kip1) and decreased p21(waf1/cip1) levels. The attenuation of osteoblast growth to high density by dexamethasone is associated with severe impairment of mineralized extracellular matrix formation, unless treatment commences in cultures that have already grown to high density. Both the antimitotic and the antiphenotypic effects are reversible, and both are antagonized by RU486. Thus, glucocorticoids induce premature attenuation of the osteoblast cell cycle, possibly contributing to the osteoporosis induced by these drugs in vivo. PMID- 10867027 TI - Evidence linking chondrocyte lipid peroxidation to cartilage matrix protein degradation. Possible role in cartilage aging and the pathogenesis of osteoarthritis. AB - Reactive oxygen species (ROS) are implicated in both cartilage aging and the pathogenesis of osteoarthritis. We developed an in vitro model to study the role of chondrocyte-derived ROS in cartilage matrix protein degradation. Matrix proteins in cultured primary articular chondrocytes were labeled with [(3)H]proline, and the washed cell matrix was returned to a serum-free balanced salt solution. Exposure to hydrogen peroxide resulted in oxidative damage to the cell matrix as established by monitoring the release of labeled material into the medium. Calcium ionophore treatment of chondrocytes, in a dose-dependent manner, significantly enhanced the release of labeled matrix, suggesting a chondrocyte dependent mechanism of matrix degradation. Antioxidant enzymes such as catalase or superoxide dismutase did not influence matrix release by the calcium ionophore activated chondrocytes. However, vitamin E, at physiological concentrations, significantly diminished the release of labeled matrix by activated chondrocytes. The fact that vitamin E is a chain-breaking antioxidant indicates that the mechanism of matrix degradation and release is mediated by the lipid peroxidation process. Lipid peroxidation was measured in chondrocytes loaded with cis parinaric acid. Both resting and activated cells showed constitutive and enhanced levels of lipid peroxidation activity, which were significantly reduced in the presence of vitamin E. In an immunoblot analysis, malondialdehyde and hydroxynonenal adducts were observed in chondrocyte-matrix extracts, and the amount of adducts increased with calcium ionophore treatment. Furthermore, vitamin E diminished aldehyde-protein adduct formation in activated extracts, which suggests that vitamin E has an antioxidant role in preventing protein oxidation. This study provides in vitro evidence linking chondrocyte lipid peroxidation to cartilage matrix protein (collagen) oxidation and degradation and suggests that vitamin E has a preventive role. These observations indicate that chondrocyte lipid peroxidation may have a role in the pathogenesis of cartilage aging and osteoarthritis. PMID- 10867028 TI - 2A proteinase of human rhinovirus cleaves cytokeratin 8 in infected HeLa cells. AB - Rhino- and enteroviruses encode two proteinases, 2A and 3C, which are responsible for the processing of the viral polyprotein and for cleavage of several cellular proteins. To identify further targets of the 2A proteinase of human rhinovirus serotype 2 (HRV2), an in vitro cleavage assay followed by two-dimensional electrophoresis was employed. Cytokeratin 8, a member of the intermediate filament group of proteins, was found to be proteolytically cleaved in vitro by the 2A proteinase of HRV2 and of coxsackievirus B4 and in vivo during HRV2 infection of HeLa cells. The cleavage results in removal of 14 amino acids from the N-terminal head domain of cytokeratin 8. However, other intermediate filament proteins (cytokeratins 7 and 18 and vimentin) were not cleaved in the course of the HRV2 infection. Compared with the processing of the eucaryotic translation initiation factors 4GI and 4GII, cleavage of cytokeratin 8 occurs late in the infection cycle at the time of the onset of the cytopathic effect. PMID- 10867029 TI - Cytosolic phospholipase A2 is required for macrophage arachidonic acid release by agonists that Do and Do not mobilize calcium. Novel role of mitogen-activated protein kinase pathways in cytosolic phospholipase A2 regulation. AB - The 85-kDa cytosolic phospholipase A(2) (cPLA(2)) mediates agonist-induced arachidonic acid release and eicosanoid production. Calcium and phosphorylation on Ser-505 by mitogen-activated protein kinases (MAPKs) regulate cPLA(2). Arachidonic acid release and eicosanoid production induced by stimuli that do (A23187, zymosan) or do not (phorbol myristate acetate (PMA), okadaic acid) mobilize calcium were quantitatively suppressed in cPLA(2)-deficient mouse peritoneal macrophages. The contribution of MAPKs to cPLA(2)-mediated arachidonic acid release was investigated. Both extracellular signal-regulated kinases (ERKs) and p38 contributed to cPLA(2) phosphorylation on Ser-505. However, although ERK inhibition did not affect A23187-induced arachidonic acid release, it suppressed zymosan-, PMA-, and okadaic acid-induced arachidonic acid release under conditions where phosphorylation of cPLA(2) on Ser-505 was unaffected. This indicates an additional regulatory mechanism for the ERK pathway. A role for transcriptional regulation is suggested by data showing that cycloheximide and actinomycin D inhibited arachidonic acid release induced by zymosan, PMA and, okadaic acid but not by A23187. Our results show that MAPK pathways contribute to arachidonic acid release in macrophages through alternative mechanisms in addition to their ability to phosphorylate cPLA(2) on Ser-505 and suggest a role for new protein synthesis. PMID- 10867030 TI - Specific cellular responses to alpha-tocopherol. AB - In the last 10 years precise cellular functions of alpha-tocopherol, some of which are independent of its antioxidant/radical-scavenging ability, have been revealed. Absorption of alpha-tocopherol from the gut is a selective process. Other tocopherols are not absorbed or are absorbed to a lesser extent. At the post-translational level, alpha-tocopherol inhibits protein kinase C and 5 lipoxygenase and activates protein phosphatase 2A and diacylglycerol kinase. Some genes [platelet glycoprotein IV/thrombospondin receptor/class B scavenger receptor (CD36), alpha-tocopherol transfer protein (alpha-TTP), alpha tropomyosin, connective tissue growth factor and collagenase] are affected by alpha-tocopherol at the transcriptional level. alpha-Tocopherol also inhibits cell proliferation, platelet aggregation, monocyte adhesion and the oxygen burst in neutrophils. Other antioxidants, such as beta-tocopherol and probucol, do not mimic these effects, suggesting a nonantioxidant, alpha-tocopherol-specific molecular mechanism. PMID- 10867031 TI - Selenomethionine: a review of its nutritional significance, metabolism and toxicity. AB - Although the need for selenium in human and animal nutrition is well recognized, the question concerning the proper form of selenium for supplemental use is still being debated. Ideally, selenium should be supplemented in the form in which it occurs naturally in foods. Because the L-isomer of selenomethionine (Se-met) is a major natural food-form of selenium, synthetic L-Se-met or enriched food sources thereof such as selenium yeast are appropriate supplemental forms of Se for humans; for animals, DL-Se-met is acceptable. Ingested Se-met is either metabolized directly to reactive forms of selenium or stored in place of methionine in body proteins. Se-met metabolism is closely linked to protein turnover. At constant intakes in the nutritional range, tissue Se levels increase until a steady state is established, preventing the build-up to toxic levels. PMID- 10867032 TI - Dietary (n-6) and (n-3) fatty acids and energy restriction modulate mesenteric lymph node lymphocyte function in autoimmune-prone (NZB x NZW)F1 mice. AB - We previously showed that dietary fish oil (FO) and energy restriction (R) have beneficial anti-inflammatory properties in the peripheral blood and spleens of (NZB x NZW)F1 (B/W) lupus-prone mice. Furthermore, unsaturated fatty acids also were shown in the past to influence mesenteric lymph node (MLN) lymphocyte function in healthy young rats. The MLN play a pivotal role in mediating food allergy. To date, the effect of R on intestinal immunity is not well understood; therefore we determined the effect of diet on MLN lymphocyte function. Mice were given either free access to a 5 g/100 g corn oil (CO) or fish oil (FO) diet or the same corn oil (CR) or fish oil (FR) diets restricted to 60% of the intake of the control group. At the age of 4 (young) and 8 (old) mo, MLN lymphocytes were isolated and B- (CD19(+)) and T-lymphocyte subsets (CD4(+) and CD8(+)) were determined by flow cytometry. Additional MLN lymphocytes were placed in culture with or without concanavalin A and culture supernatants collected after 72 h for cytokine and immunoglobulin (Ig) quantitation by ELISA. Aging significantly (P < 0.05) decreased both CD4(+) and CD8(+) T-lymphocytes. Spontaneous and activation induced interleukin-4 (IL-4), IL-10, and interferon-gamma secretion were greater while IL-2 was lower in CO-fed old mice compared to CO-fed young mice. In contrast, CR or FO alone partially blunted the age-dependent alterations in T lymphocyte ratios including cytokine and Ig secretion, whereas the FR diet significantly (P < 0.005) normalized the accelerated aging effects on these immune variables. We show for the first time that FR is a far more potent anti inflammatory therapy than either CR or FO alone in modulating MLN lymphocyte function. PMID- 10867033 TI - Genistein inhibits growth of estrogen-independent human breast cancer cells in culture but not in athymic mice. AB - The studies presented were conducted to assess the effect of the soy isoflavone genistein on proliferation of estrogen-independent human breast cancer cells (MDA MB-231) in vitro and in vivo. Genistein (20 mcmol/L) inhibited cell proliferation in vitro by approximately 50%. Cell cycle progression was blocked in G(2)/M with 40 and 80 mcmol/L genistein. To evaluate the effect of dietary genistein on tumor growth in vivo, genistein was fed to female athymic mice inoculated with MDA-MB 231 cells. After solid tumor masses had formed, mice were fed genistein at a dose (750 mcg/g AIN-93G diet), shown to produce a total plasma genistein concentration of approximately 1 mcmol/L. This dose of genistein did not significantly (P > 0.05) alter tumor growth. Studies were then conducted to assess the effect of dietary genistein on initial tumor development and growth. Genistein (750 mcg/g AIN-93G diet), fed 3 d before cells were inoculated into mice, did not significantly (P > 0.05) inhibit tumor formation or growth. The plasma concentration of genistein in mice fed this dose of dietary genistein (750 mcg/g AIN-93G diet) does not appear sufficient to inhibit tumor formation or growth. Dietary genistein at 750 mcg/g AIN-93G diet does not inhibit tumor formation or growth. Additional studies were conducted to determine the effect of dietary dosages ranging from 0 to 6000 mcg/g AIN-93G diet on plasma genistein concentration. Plasma genistein concentration increased in a dose-dependent manner up to 7 mcmol/L at 6000 mcg/g AIN-93G diet. These data suggest that although genistein inhibits cancer cell growth in vitro, it is unlikely that the plasma concentration required to inhibit cancer cell growth in vivo can be achieved from a dietary dosage of genistein. PMID- 10867034 TI - A buckwheat protein product suppresses gallstone formation and plasma cholesterol more strongly than soy protein isolate in hamsters. AB - This study was conducted to investigate the effects of a buckwheat protein product (BWP) on plasma cholesterol, gallbladder bile composition and fecal steroid excretion in hamsters fed diets with 5 g/kg cholesterol. Diets also contained 200 g/kg of casein, soy protein isolate (SPI) or BWP as protein sources. After 2 wk, plasma and liver concentrations of cholesterol in the hamsters fed BWP were significantly lower than those in the hamsters fed casein and SPI. The molar proportion of cholesterol in gallbladder bile was significantly lower in the BWP group than in the other groups, whereas that of bile acids was slightly higher in the BWP group (P 100% higher in ZD Chelex cells compared with their respective controls. To examine whether the effects were specific to zinc depletion, a third, zinc-replenished group (ZDA) was included in the Opti-MEM study in which cells were cultured in ZD media for nearly one passage before a change was made to zinc-adequate (ZA) medium for the last 24 h. Zinc levels in the ZDA cells were significantly higher than in ZD cells, and p53 mRNA abundance was normalized to control levels. Nuclear p53 protein levels were >100% higher in the ZD Opti-MEM cells than in ZA cells. Interestingly, the ZDA Opti-MEM cells had significantly lower levels of nuclear p53 protein than both the ZA and ZD cells. These data suggest that expression of p53, a critical component in the maintenance of genomic stability, may be affected by reductions in cellular zinc. PMID- 10867038 TI - Soy isoflavone aglycones are absorbed faster and in higher amounts than their glucosides in humans. AB - Isoflavones are contained in soybean or soy foods in two chemical forms, i.e., aglycones and glucosides. We investigated the difference in the absorption of soy isoflavone aglycones and glucosides in humans. After a single, low dose intake (0.11 mmol), the highest isoflavone concentrations in plasma were reached 2 and 4 h after ingestion of aglycones and glucosides, respectively; subjects were four men (41 y old) and four women (45 y old). The highest plasma concentration after aglycone intake was more than two times greater than that after glucoside ingestion. In a similar manner, we then compared the plasma isoflavone concentration profiles after intake of a single, high dose of isoflavones (1.7 mmol) in eight subjects (four men, 40 y old; four women, 47 y old) and found the highest plasma concentration after aglycone intake was more than five times higher than that after glucoside intake. In both high and low dose intake tests, the plasma concentration of genistein was significantly higher than that of daidzein despite the similar levels of intake. After long-term (4 wk) intakes (0.30 mmol/d), we also measured the plasma concentration of isoflavones (eight men, 45 y old). After 2 and 4 wk, these concentrations remained >100% higher after ingestion of aglycones than of glucosides. The isoflavone aglycones were absorbed faster and in greater amounts than their glucosides in humans. Isoflavone aglycone-rich products may be more effective than glucoside-rich products in preventing chronic disease such as coronary heart disease. PMID- 10867039 TI - Protein feeding pattern does not affect protein retention in young women. AB - This study was undertaken to determine whether a pulse protein feeding pattern was more efficient than a spread pattern to improve protein anabolism in young women as was already shown in elderly women. After a 15-d adaptive period [1.2 g protein/(kg fat-free mass. d)], 16 young women (age 26 +/- 1 y) were given a 14-d diet providing 1.7 g protein/(kg fat-free mass. d), using either a pulse pattern (protein consumed mainly in one meal, n = 8), or a spread pattern (spreading daily protein intake over four meals, n = 8). Nitrogen balance was determined at the end of both the 15-d adaptive and the 14-d experimental periods. Whole-body protein turnover was determined at the end of the 14-d experimental period using [(15)N]glycine as an oral tracer. Nitrogen balance was 17 +/- 5 mg N/(kg fat-free mass. d) during the adaptive period. It was higher during the experimental period, but not significantly different in the women fed the spread or the pulse patterns [59 +/- 12 and 36 +/- 8 mg N/(kg fat-free mass. d) respectively]. No significant effects of the protein feeding pattern were detected on either whole body protein turnover [5.5 +/- 0.2 vs. 6.1 +/- 0.3 g protein/(kg fat-free mass. d) for spread and pulse pattern, respectively] or whole-body protein synthesis and protein breakdown. Thus, in young women, these protein feeding patterns did not have significantly different effects on protein retention. PMID- 10867040 TI - High-molecular-weight hydroxypropylmethylcellulose taken with or between meals is hypocholesterolemic in adult men. AB - Hydroxypropylmethylcellulose (HPMC) is a food gum that shares certain characteristics, such as high viscosity, with soluble fibers. In this trial, the safety and cholesterol-lowering efficacy of HPMC consumed with and between meals was evaluated in free-living male volunteers with mild-to-moderate hypercholesterolemia. After a 14-d baseline period, men (n = 51) with LDL cholesterol between 3.36 and 4.91 mmol/L and triglycerides <3.95 mmol/L were randomly assigned to consume 5.0 g/d HPMC in 240 mL of orange drink, taken either with or between meals, for a 2-wk treatment period. In the Between Meals group, total cholesterol was reduced by 8.0% vs. baseline in wk 1 of treatment (P < 0.05) and 5.1% in wk 2 (P < 0.01). LDL cholesterol concentrations fell by 12.0 and 7.7% (P < 0.01). In the With Meals group, reductions were 9.5 and 8.3% for total cholesterol, and 12.5 and 12.8% for LDL cholesterol (wk 1 and 2, respectively, P < 0.01). In both groups, HDL cholesterol decreased by approximately 5% during wk 1 of treatment (P < 0.01), but the wk 2 concentrations were not significantly different from baseline. There were no significant differences between groups in lipid responses, although there was a trend for a smaller LDL cholesterol-lowering effect during wk 2 of treatment in the Between Meals group (P < 0.06). Gastrointestinal-related adverse experiences (mostly mild) were twice as common among participants who ingested HPMC with meals (P < 0.05). These results suggest that HPMC has a lipid-lowering effect, which may be more consistent when taken with meals. PMID- 10867041 TI - Olestra consumption does not predict serum concentrations of carotenoids and fat soluble vitamins in free-living humans: early results from the sentinel site of the olestra post-marketing surveillance study. AB - In 1996, the U.S. Food and Drug Administration approved olestra, a fat substitute, for use in snack foods. Previous studies had shown that olestra consumption could reduce absorption of carotenoids and fat-soluble vitamins. To determine the association between consumption of olestra-containing snack foods and serum concentrations of carotenoids and fat-soluble vitamins in a free-living population, we interviewed independent population-based cross-sectional samples of 1043 adults before olestra was available and 933 adults 9 mo after olestra snacks were introduced into the marketplace in Marion County, IN, the first major test market for olestra. A cohort composed of 403 adults from the first survey, oversampling those most frequently reporting olestra consumption during follow-up telephone interviews, completed a second survey. We assessed diet, lifestyle factors and olestra consumption, and collected blood for assays for the serum concentrations of six carotenoids, four fat-soluble vitamins and lipids. Nine months after the introduction of olestra into the marketplace, 15.5% of Marion County residents reported consuming an olestra-containing snack in the previous month, with a median frequency among consumers of 3.0 times per month. There were no significant associations or consistent trends for decreased serum carotenoids or fat-soluble vitamins associated with olestra consumption, although cohort members consuming >/=2 g/d of olestra had adjusted total serum carotenoids 15% lower compared with baseline. There were increases in serum vitamin K concentrations associated with olestra consumption (P = 0.03 in the cross section and P = 0.06 in the cohort). In summary, there was no statistically significant evidence in this free-living population of associations between olestra consumption and decreased serum concentrations of carotenoids and fat-soluble vitamins. PMID- 10867042 TI - Prolonged tomato juice consumption has no effect on cell-mediated immunity of well-nourished elderly men and women. AB - The immunomodulatory potential of carotenoids has been investigated thoroughly only for beta-carotene. Data on the immunomodulatory activity of other carotenoids such as lycopene are scarce. The objective of this study was to investigate the effects of prolonged tomato juice consumption on cell-mediated immunity of well-nourished healthy elderly persons. In an intervention study, 33 female and 20 male subjects (aged 63-86 y) consumed 330 mL/d tomato juice (47.1 mg/d lycopene) or mineral water for 8 wk. Immune status was assessed by measuring number and lytic activity of natural killer (NK) cells, secretion of cytokines [interleukin (IL)-2, IL-4, tumor necrosis factor-alpha (TNF-alpha)] by activated peripheral blood mononuclear cells (PBMC), lymphocyte proliferation, and delayed type hypersensitivity (DTH) skin responses. Tomato juice consumption resulted in significantly increased plasma lycopene and beta-carotene concentrations over time. In both treatment groups, TNF-alpha and IL-4 secretion were increased at the end of the intervention period, whereas IL-2 secretion was decreased. Tomato juice consumption had no effect on lymphocyte proliferation, DTH or the number of NK cells. Lytic activity of NK cells was increased in both groups at the end of the intervention period. In conclusion, these results show that prolonged tomato juice consumption increased plasma lycopene concentrations without significantly affecting cell-mediated immunity in well-nourished elderly subjects. PMID- 10867044 TI - Biobehavioral factors are associated with obesity in Puerto Rican children. AB - The purpose of this case-control study was to identify predictors of obesity among Puerto Rican children from Hartford, CT. The study included 53 prepubertal children, 31 girls and 22 boys, between 7 and 10 y of age. Children were classified as obese [n = 29, body mass index (BMI) >/= 85th percentile] or controls (n = 24, BMI < 85th percentile). Multivariate logistic regression analyses indicated that frequency of fruit juice consumption [odds ratio (OR), 95% confidence interval (CI); 4.02, 1.48-10.95], hours of daily TV viewing (1.86, 1.02-3.42), maternal BMI (1.39, 1.10-1.77) and lower dairy product intake (0.41, 0.19-0.93) were associated with obesity. Television viewing was correlated (P < 0.05) with lower physical activity in girls, and with higher snacking frequency and sweets consumption in boys. Obese children were more likely than controls to have higher systolic and diastolic blood pressures and to have experienced more ear infections and diarrhea during the previous year. Results provide evidence of the multifactorial nature of childhood obesity in this community. PMID- 10867043 TI - Malaria, hookworms and recent fever are related to anemia and iron status indicators in 0- to 5-y old Zanzibari children and these relationships change with age. AB - In Zanzibar and other tropical regions, iron deficiency, malaria and multiple helminth infections coexist. We addressed the following questions: 1) What are the predictors of low hemoglobin in Zanzibari preschool children? 2) Are indicators of iron status informative in this population? 3) Does malaria modify the relation of iron indicators to hemoglobin? We used multivariate regression to analyze cross-sectional data from a community-based sample of rural Zanzibari children who were not ill (n = 490; 4-71 mo of age) in whom we assessed hemoglobin, serum ferritin (SF), erythrocyte protoporphyrin (EP), serum transferrin receptor (TfR), recent fever, malaria parasitemia and helminth fecal egg counts. Of hemoglobin values, 80% were <100 g/L and 15.5% were <70 g/L. In children <18 mo of age, 40.2% of hemoglobin values were <70 g/L. Our primary findings were as follows: 1) In children <30 mo old, hemoglobin was associated with malaria but not hookworms, whereas in children >/=30 mo, hemoglobin was related to hookworms but not malaria. In the younger age group, male sex and recent fever also predicted lower hemoglobin. 2) The three iron indicators were informative in this population but did not reflect only iron status. Malaria elevated SF in younger children and TfR and EP in both age groups. Fever elevated SF in older children and EP in both age groups, but not TfR. 3) Malaria modified the relation of all three indicators to hemoglobin. The relation of SF to hemoglobin was weak overall, and absent in malaria-infected children. EP and TfR were strongly related to hemoglobin, but this relation was attenuated by malaria. PMID- 10867045 TI - Dietary protein does not affect overloaded skeletal muscle in rats. AB - We compared the effects of three levels of dietary protein, i.e., 7% (low protein; LP); 17.5% (adequate protein; CON); or 30% (high protein; HP) on growth of functionally overloaded muscle in Sprague-Dawley male rats. Growth of plantaris and soleus muscles was induced by the surgical removal of gastrocnemius muscles in one hindlimb; muscles in the other leg were used as sham-operated, intra-animal controls. After 4 wk, rats fed the 7% LP diet gained less weight ( 29%) and had lighter livers (-20%) and kidneys (-16%) than rats fed the CON diet (P < 0.05). Measurements of rats fed the 30% HP diet were not different from those of CON rats except that their kidneys were larger (+6%) (P < 0.05). The level of dietary protein did not affect the experimentally induced muscular growth in either plantaris or soleus muscles. Gains in overloaded plantaris muscles over sham-operated muscles were not different among rats fed LP, CON and HP diets for muscle mass (+42 to +45%), total protein (+42 to +46%) and myofibrillar protein (+40 to +44%). Soleus muscles also did not differ among diet groups for gains in mass (+20 to +33%), total protein (+20 to +33%) and myofibrillar protein (+21 to +33%). No dietary protein effects were found on myosin heavy chain isoform (I, IIa, IIx, IIb) expression in either plantaris or soleus muscles. We conclude that gains in plantaris and soleus muscle mass, total protein and myofibrillar protein induced by functional overload are not affected by low (7%) or high (30%) protein feeding in young male rats for 4 wk. PMID- 10867046 TI - Preferential incorporation of docosahexaenoic acid into nonphosphorus lipids and phosphatidylethanolamine protects rats from dietary DHA-stimulated lipid peroxidation. AB - In a previous study, we found that dietary docosahexaenoic acid (DHA)-stimulated tissue lipid peroxide formation was suppressed to a lesser extent than expected from the peroxidizability index of tissue total lipids. This suppression was presumed to be potentiated by mechanisms other than the lipid peroxide-scavenging system. In this study, we focused primarily on the incorporation of DHA into tissue nonphosphorus lipids and phospholipid species. DHA and different levels of dietary vitamin E (VE; 7.5, 54, 134 and 402 mg/kg of diet) were fed to rats for 32 d. In rats with poor VE status, liver chemiluminescence intensity and kidney and testis thiobarbituric acid (TBA) values correlated with the tissue's peroxidizability index. In rats with normal VE nutriture, liver lipid peroxide formation was suppressed to a level below that expected from the peroxidizability index, likely because DHA was present in nonphosphorus lipids and utilized preferentially for phosphatidylethanolamine synthesis. In the kidney, differences in the TBA values were associated with differences in the peroxidizability index of total lipids, even in the DHA groups fed VE at higher than normal levels. This may be because the levels of lipid peroxide scavengers were lower than those of liver and because DHA was utilized preferentially for phosphatidylcholine synthesis. In testis, the lipid peroxide levels were not as high as expected from the peroxidizability index, even in rats fed a high DHA diet containing the normal level of VE. This may be because the testis was composed of a high proportion of (n-6) polyunsaturated fatty acids (PUFA), which are low in unsaturation, and thus the proportion of DHA was low. In addition, in testis, VE and ascorbic acid, which act as antioxidants, were retained at higher levels in rats with particularly poor and normal VE nutriture than those of liver and kidney. These results suggest that antioxidant protection against dietary DHA stimulated lipid peroxidation below the extent expected from the peroxidizability index of tissue total lipids differed from tissue to tissue. The suppression was likely due to not only the lipid peroxide scavenging system but also preferential incorporation of DHA into nonphosphorus lipids and phosphatidylethanolamine, particularly in liver. PMID- 10867047 TI - Carbohydrate supplementation of horses during endurance exercise: comparison of fructose and glucose. AB - To delay the onset of fatigue, endurance horses are often fed at rest stops during races. The resulting increase in blood insulin may adversely inhibit lipolysis. In humans, ingestion of fructose produces a smaller insulin rise than glucose. This study compared glucose and fructose as carbohydrate supplements for endurance horses. Three Arabian geldings were given 300 g of fructose (F), glucose (G) or 50% glucose: 50% fructose (GF), in 1.5 L water, by stomach tube. In the Resting Test, carbohydrate was administered at rest. Following treatment, blood samples were taken every 30 min for 8 h, and feces were collected for 24 h. Treatment did not affect fecal weight or water content. Plasma glucose and insulin responses did not differ among treatments. Post-treatment (60 min), plasma L-lactate tended to be higher (P = 0.06) after the F and GF treatments than after the G treatment. In the Exercise Test, two treadmill exercise bouts at 0 degrees incline (Bout 1: 90 min; Bout 2: 120 min) were separated by a 1-h rest period. A total distance of 36.84 km was covered at a mean speed of 2.9 m/s. Carbohydrate was administered 45 min before Bout 2. Plasma glucose and insulin at the start of Bout 2 were higher (P = 0.02 and 0.03, respectively) with the GF treatment than with the F treatment. However, during exercise, plasma glucose concentrations did not differ among treatments. We conclude that fructose is well absorbed by horses and rapidly converted to glucose. PMID- 10867048 TI - Soy isoflavone conjugation differs in fed and food-deprived rats. AB - An experiment clarifying the influence of food deprivation on the isoflavone conjugation pattern in rats was conducted. Food-deprived and fed rats were administered daidzein and genistein at 7.9 mcmol/kg body, and changes in their plasma metabolites (i.e., free compounds, sulfates, glucuronides, sulfates/glucuronides) were measured quantitatively as a function of time. In the food-deprived group, total plasma daidzein and genistein reached maximum concentrations of 20.9 +/- 4.4 and 11.4 +/- 3.1 mcmol/L, respectively, 10 min after administration, whereas in the fed group, the maxima were 2.4 +/- 0.8 mcmol/L for daidzein after 2 h and 1. 8 +/- 0.2 mcmol/L for genistein after 4 h. In both groups, there were significantly more daidzein sulfates than genistein sulfates. Moreover, depriving rats of food before daidzein and genistein administration significantly increased plasma isoflavone sulfates with simultaneous significant decreases in plasma isoflavone glucuronides compared with fed rats. Additionally, nonconjugated daidzein and genistein appeared in plasma of food-deprived rats for 1 h after administration. Plasma concentrations of conjugates having both sulfate and glucuronide moieties were not significantly different between the groups. PMID- 10867049 TI - In vitro fermentation pattern of D-tagatose is affected by adaptation of the microbiota from the gastrointestinal tract of pigs. AB - Knowledge of the fermentation pattern of D-tagatose is important for the assessment of energy value and compliance of D-tagatose. In vitro fermentation experiments with pig intestinal contents and bacteria harvested from the gastrointestinal tract of pigs were used to investigate the degradation of D tagatose and the formation of fermentation products. Two groups of eight pigs were fed either a control diet containing 150 g/kg sucrose or a diet which had 100 g/kg of the sucrose replaced by D-tagatose. After 18 d the pigs were killed and the gastrointestinal contents collected for in vitro studies. No microbial fermentation of D-tagatose occurred in the stomach or in the small intestine, whereas the sugar was fermented in the cecum and colon. Formate, acetate, propionate, butyrate, valerate, caproate and some heptanoate were produced by the microbial fermentation of D-tagatose by gut microbiota. Hydrogen and methane were also produced. The population of D-tagatose-degrading bacteria in fecal samples and the capacity of bacteria from the hindgut to degrade D-tagatose were higher in the pigs adapted to D-tagatose compared with unadapted pigs. In unadapted pigs, the major fermentation product from D-tagatose was acetic acid. Much more butyric and valeric acids were produced from D-tagatose by bacterial slurries of tagatose-adapted pigs compared with unadapted pigs; this was especially the case for samples from the colon. We conclude that D-tagatose is not fermented in the upper gastrointestinal tract, and the ability of the large intestinal microbiota to ferment D-tagatose is dependent on adaptation. PMID- 10867050 TI - Coarse brown rice increases fecal and large bowel short-chain fatty acids and starch but lowers calcium in the large bowel of pigs. AB - Young male pigs were fed a diet formulated from human foods including either boiled white rice plus rice bran or heat-stabilized brown rice at equivalent levels of fiber for 3 wk. Stool and starch excretion were low in pigs fed white rice during the first 2 wk of the experiment. In pigs fed brown rice, their excretion was high during wk 1 but declined in wk 2 while short-chain fatty acid (SCFA) excretion was higher at both times. Large bowel digesta mass, measured during wk 3, was higher in pigs fed brown rice but only in the proximal colon. Large bowel and fecal starch concentrations were higher in pigs fed brown rice but the difference was insufficient to explain the increase in large bowel digesta mass. In pigs with a cecal cannula, digesta starch concentrations were equally higher when white or brown rice was fed compared with the corresponding rice which had been finely milled, indicating that particle size was a determinant of ileal digestibility. Concentrations and pools of total and individual SCFA were higher in all regions of the colon but not the cecum of pigs fed brown rice. Large bowel Ca(2+) concentrations were lower in pigs fed brown rice, suggesting greater absorption. The data confirm earlier findings that brown rice raises large bowel digesta mass and SCFA through greater fermentation of starch but show that starch itself makes a relatively small contribution to digesta and stool mass. Apparently, the rate of passage of digesta is a determinant of the concentrations and pools of SCFA in the distal colon and in feces. PMID- 10867051 TI - Dietary beta-carotene is taken up by blood plasma and leukocytes in dogs. AB - beta-Carotene uptake by blood plasma and leukocytes was studied in mature beagle dogs. In expt. 1, dogs were fed once orally with 0, 50, 100 or 200 mg of beta carotene and their blood was sampled at 0, 1. 5, 3, 6, 10, 18 and 24 h. Plasma beta-carotene concentrations increased dose-dependently to peak at 6 h postfeeding. Concentrations decreased rapidly thereafter, showing a half-life of 3 to 4 h. In expt. 2, dogs were given daily doses for seven consecutive days with 0, 12.5, 25, 50 or 100 mg beta-carotene. Plasma beta-carotene concentrations increased dose-dependently; concentrations after the last dose were two- to fourfold higher than after the first dose. In expt. 3, dogs were fed 0, 50 or 100 mg beta-carotene daily for 30 d. beta-Carotene was elevated in lymphocytes and neutrophils in supplemented dogs. Furthermore, beta-carotene was taken up by the cytosol, mitochondria, microsomes (lymphocytes and neutrophils) and nuclei (lymphocytes only), proving that dogs can absorb beta-carotene. beta-Carotene is taken up by subcellular organelles of blood lymphocytes and neutrophils and in the plasma and leukocytes beta-carotene may have physiological importance as it relates to immunity in dogs. Uptake kinetics indicated that dogs are not an appropriate animal model for studying beta-carotene absorption and metabolism in humans. PMID- 10867052 TI - Fructooligosaccharide consumption enhances femoral bone volume and mineral concentrations in rats. AB - We examined whether the enhanced mineral absorption resulting from fructooligosaccharide (FOS) consumption affects femoral bone structure and mineral concentrations, using histomorphometrical and X-ray microanalysis. Male Wistar rats (n = 16; 42 d old) were divided into two groups, a control group (n = 8) and a FOS group (5 g/100 g FOS in the diet, n = 8). After a 3-d adaptation period, constant amounts of calcium (95 mg/d) and magnesium (8 mg/d) were fed to the rats in each group, using a pair-feeding protocol. At age 60 d, a 3-d metabolic study was initiated. Calcium and magnesium absorptions were calculated. The rats were then killed, and the right femur was embedded in polyester resin. The distal metaphysis was sagittal-sectioned, and the middle of the diaphysis and neck were cross-sectioned. Calcium, magnesium and phosphorus concentrations in the three samples were then measured. Calcium and magnesium absorptions were significantly greater in FOS-fed rats. Trabecular bone volume at the metaphysis and bone volume at the neck of the femur in FOS-fed rats were also significantly greater than those in control rats. The mineral concentration (Ca, Mg and P) in each region of the bone surface was greater in FOS-fed rats. There was a significant relationship between absorbed calcium and calcium concentrations in bone (r = 0.722, P < 0.001), and a similar relationship was found for magnesium (r = 0.720, P < 0.001). These results suggest that the enhanced calcium and magnesium absorption due to FOS consumption might enhance femoral bone volume and mineral concentrations. PMID- 10867053 TI - Dietary arginine requirement of juvenile red drum (Sciaenops ocellatus) based on weight gain and feed efficiency. AB - Increasing aquacultural production of red drum (Sciaenops ocellatus) has prompted the determination of many of this species' nutritional requirements. However, limited information is available concerning its amino acid requirements, especially for arginine. Therefore, a feeding trial was conducted with juvenile red drum to determine their quantitative dietary requirement for arginine. Experimental diets contained 35 g crude protein/100 g from red drum muscle and crystalline amino acids. Incremental levels of arginine were added to the diets in place of a mixture of glycine and aspartic acid to maintain all diets isonitrogenous. All diets were fed in triplicate to juvenile red drum for 7 wk. Graded levels of arginine significantly (P < 0.05) affected weight gain, feed efficiency, protein efficiency ratio (PER) and plasma arginine levels of the fish. Based on least-squares regression of feed efficiency and PER data, the minimum requirement (+/- SEM) of red drum for arginine was estimated at 1.44 (+/- 0.15) and 1.48 (+/- 0.12) g/100 g diet (4.11 and 4.23 g/100 g dietary protein), respectively. The arginine requirements estimated from weight gain data were 1.75 (+/- 0.18) g/100 g diet or 5.0 g/100 g dietary protein. These values are similar to those reported for other carnivorous fish species. PMID- 10867054 TI - Depletion of alpha-tocopherol and astaxanthin in Atlantic salmon (Salmo salar) affects autoxidative defense and fatty acid metabolism. AB - Duplicate groups of Atlantic salmon post-smolts were fed four purified diets supplemented with both vitamin E and the carotenoid astaxanthin (Ax) (+E, +Ax), or supplemented with either vitamin E or Ax (-E, +Ax and +E, -Ax) or deficient in both vitamin E and Ax (-E, -Ax) for 22 wk. There were no effects of diet on growth rate, but an extensive lipoid liver degenerative lesion was observed in 15% of fish fed diets deficient in vitamin E. Tissue vitamin E concentrations varied in accordance with dietary vitamin E in liver, muscle, heart, plasma, brain and eye; levels were reduced to approximately 3% in liver but only to 40% in eye of fish fed diets deficient in vitamin E compared with those fed diets supplemented with vitamin E. An interactive sparing of Ax supplementation on tissue vitamin E concentration was observed, but only in brain. Dietary deficiency of both vitamin E and Ax significantly increased the recovery of desaturated and elongated products of both [1-(14)C] 18:3(n-3) and [1-(14)C] 20:5(n-3) in isolated hepatocytes, suggesting that conversion of fatty acids to their long-chain highly unsaturated products can be stimulated by a deficiency of lipid-soluble antioxidants. The antioxidant synergism of vitamin E and Ax was supported by their ability to reduce malondialdehyde formation in an in vitro stimulation of microsomal lipid peroxidation and to reduce plasma levels of 8 isoprostane. The results of this study suggest that both vitamin E and the carotenoid Ax have antioxidant functions in Atlantic salmon. PMID- 10867055 TI - Dietary L-carnitine improves nitrogen utilization in growing pigs fed low energy, fat-containing diets. AB - Growing pigs (n = 25; 17.8 +/- 0.1 kg) were used to study the effects of L carnitine and protein intake on nitrogen (N) balance and body composition. Fat supplemented (40 g soy oil/kg diet), corn-soybean meal basal diets containing low or high protein (136 or 180 g/diet) were formulated so that protein accretion would be limited by metabolizable energy (ME). Each basal diet was supplemented with 0 or 500 mg/kg L-carnitine and fed to pigs for 10 d in a nutrient balance trial. Final body composition was compared with weight and age-matched pigs measured on d 0 to calculate nutrient accretion rates. High protein feeding increased (P < 0.01) average daily gain (ADG) by 34%, as well as nitrogen digestibility (4.4%), retention (5.2%), urinary excretion (29%) and crude protein (CP) accretion (33%). Total-body carnitine accretion rate was 4.5 fold greater and total body carnitine concentration was almost 100% greater than in unsupplemented controls (P < 0.01). Irrespective of protein level, carnitine increased ADG (by 7.3%, P < 0.10) and CP accretion rate (9%, P < 0.10). Congruently, carnitine supplementation improved the efficiency of nitrogen retention (P < 0. 05) and reduced urinary nitrogen excretion (14%, P < 0.10). Carcass fat content also was reduced in carnitine-supplemented pigs (P < 0. 10). Collectively, these data support the hypothesis that carnitine can improve the efficiency of nitrogen utilization in 20-kg pigs fed energy-limited, fat containing diets. We conclude that endogenous carnitine biosynthesis may be adequate to maintain sufficient tissue levels during growth, but that supplemental dietary carnitine (at 500 mg/kg) may be retained sufficiently so as to alter nutrient partitioning and thus body composition of 20-kg pigs. PMID- 10867056 TI - Dietary soy protein is associated with reduced intestinal mucosal polyamine concentration in male Wistar rats. AB - Quantitation of polyamine levels has been correlated with biomarkers of proliferation in the colon mucosa where dysregulated epithelial hyperproliferation is associated with colorectal cancer risk. This study was performed to assess the response of polyamine measurements to dietary factors in an animal model. Male Wistar rats were fed purified diet or diets substituted by 20% lard fat, 20% beet fiber and 20% soy protein. After 2 wk, mucosal polyamines were measured along intestinal tracts by HPLC. In rats fed the control diet (n = 10), mucosal polyamines were found at high levels in the duodenum, jejunum and ileum but at low levels in the cecum, colon and rectum. Compared with rats fed the control diet, those fed the 20% lard diet showed greater polyamine levels in the large intestine (P < 0.05, n = 10), but those fed the 20% fiber diet exhibited lower polyamine levels in the small intestine (P < 0.05, n = 9). However, rats fed the 20% soy protein diet had lower polyamine levels in both small and large intestines (P < 0.05, n = 15). Significant linear correlations were observed between rectal polyamine levels and the dietary energy intakes in these four diet groups (r = 0.972-0.991, P < 0.001). Supplementation of 0.1% soy isoflavones to the basal diet or 0.3% DL-methionine to the 20% soy protein diet for 4 wk did not affect polyamine levels. The results indicate that soy protein reduced mucosal polyamine levels, at least in part, through reduction of energy intakes. Further studies are warranted to verify that polyamine levels in intestinal mucosa are useful as an intermediate endpoint of the dietary risk factors. PMID- 10867057 TI - Maternal protein deficiency causes hypermethylation of DNA in the livers of rat fetuses. AB - Maternal protein deficiency during pregnancy is associated with changes in glucose tolerance and hypertension in the offspring of rats. In this study the growth of rat fetuses was examined when the dams were fed diets containing 18% casein, 9% casein or 8% casein supplemented with threonine. The extra threonine was added to reverse the decrease in circulating threonine concentrations that occurs when pregnant rats are fed protein-deficient diets. The fetuses of the group fed the low protein diet supplemented with threonine were significantly smaller than those of the control group and not significantly different from those fed low protein. Homogenates prepared from the livers of dams fed the diet containing 9% casein oxidized threonine at approximately twice the rate of homogenates prepared from dams fed the diet containing 18% casein. We conclude that circulating levels of threonine fall as a consequence of an increase in the activity of the pathway that metabolizes homocysteine produced by the transulfuration of methionine. Serum homocysteine was unaffected in the dams fed low protein diets compared with controls, but was significantly greater in dams fed the low protein diet supplemented with threonine. Elevated levels of homocysteine are associated with changes in the methylation of DNA. The endogenous methylation of DNA was greater than that of controls in the livers of fetuses from dams fed the 9% protein diets and increased further when the diet was supplemented with threonine. Our results suggest that changes in methionine metabolism increase homocysteine production, which leads to changes in DNA methylation in the fetus. An increase in maternal homocysteine may compromise fetal development, leading to the onset of glucose intolerance and hypertension in adult life. PMID- 10867058 TI - Arginine enhances In vivo immune responses in young, adult and aged mice. AB - Arginine supplementation enhances in vitro lymphocyte proliferation in healthy adult humans and in rodent models. Studies examining the effect of arginine supplementation on in vivo immune responses are lacking. The purpose of this study was to determine whether arginine supplementation could enhance in vivo immune responses in adult mice and reverse known age-associated alterations in immune function of young and aged mice. Mice (1, 10 and 33 mo old) were fed a 2% arginine or an isonitrogenous diet for 2 wk. Delayed-type hypersensitivity to 2,4 dinitrofluorobenzene-challenged ears and changes in popliteal lymph node weights to injected sheep red blood cells were measured. The mean percentage of increase in ear thickness in challenged vs. unchallenged ears was 27, 35 and 24% with arginine supplementation and 7, 12 and 0% with the isonitrogenous diet in the 1-, 10- and 33-mo-old mice, respectively (P 90% OBA). It was also noted that the degree of definition became poorer towards the distal end of the arches and was worst on the mandibular molars. It is suggested that the etched enamel morphology of different tooth types could affect composite resin bond strengths. Consequently, this feature could effect the clinical survival of orthodontic brackets and contribute to the higher failure rate of brackets on posterior teeth. PMID- 10867071 TI - The development of the index of complexity, outcome and need (ICON). AB - This paper is based on the winning submission for the 1998 Chapman prize awarded by the British Orthodontic Society for an essay on a subject promoting the interests of orthodontics. The aim of the investigation is to develop a single index for assessing treatment inputs and outcomes. An international panel of 97 orthodontists gave subjective judgements on the need for treatment, treatment complexity, treatment improvement, and acceptability on a diverse sample of 240 initial and 98 treated study models. The occlusal traits in the study models were scored according to a defined numerical protocol. Five highly predictive occlusal traits were identified (IOTN Aesthetic Component, crossbite, upper arch crowding/ spacing, buccal segment antero-posterior relationships, and anterior vertical relationship) and then used to 'predict' the panelist's decisions using regression analysis. Cut-off values were determined for the dichotomous judgements by plotting specificity sensitivity and overall accuracy. Twenty percentile ranges were used to determine 5 grades of complexity and improvement. The index prediction of decisions for treatment need, had specificity 84.4 per cent, sensitivity 85.2 per cent, and overall accuracy 85 per cent. When used to predict treatment outcomes, the new index had specificity 64.8 per cent, sensitivity 70.1 per cent, and overall accuracy 68.1 per cent. The index could explain 75.6 per cent of the variance in the mean casewise complexity score and 63.5 per cent of the mean casewise improvement score. A new orthodontic index is proposed to assess treatment need, complexity, and outcome. It is based on international orthodontic opinion. PMID- 10867072 TI - Effect of wearing cervical headgear on tongue pressure. AB - The purpose of this study was to examine whether wearing cervical headgear affected tongue pressure on the lingual surface of mandibular incisors, with particular attention to suprahyoid muscle activity. Tongue pressure was recorded using a miniature pressure sensor without cervical headgear and with two cervical headgears with traction forces of 500 and 1200 g, respectively. Electromyographic activity of suprahyoid muscles and respiratory-related movement were recorded simultaneously. Wearing cervical headgear significantly affected tongue pressure and suprahyoid muscle activity in the short-term. A significant increase in tongue pressure was observed in association with an increase in traction force from 500 to 1200 g, whereas no significant difference in suprahyoid muscles activity was seen between these force levels. These results suggest that wearing cervical headgear increases tongue pressure on the lingual surface of mandibular incisors, and this increase in tongue pressure may result from changes in the electromyographic activity of suprahyoid muscles to maintain adequate pharyngeal patency. PMID- 10867073 TI - Radiographic factors affecting the management of impacted upper permanent canines. AB - The aim of the investigation was to evaluate which radiographic factors influenced the orthodontists' decision whether to expose or remove an impacted upper permanent canine and was a retrospective, cross-sectional design. The sample consisted of all radiographic records of patients referred to the Orthodontic Department at Manchester University Dental Hospital with impacted upper permanent canines between 1994-1998 (n = 44). The following canine position measurements were made from the OPG: angulation to the midline, vertical height, antero-posterior position of the root, overlap of the adjacent incisor, and presence of root resorption of adjacent incisor(s). The labio-palatal position of the impacted canine was assessed from the lateral skull radiograph. Whether the impacted canine had been exposed and orthodontically aligned or removed was also recorded. Stepwise logistic regression analysis showed that the labio-palatal position of the crown influenced the treatment decision, with palatally positioned impacted canines more likely to be surgically exposed and those in the line of the arch, or labially situated, removed (P < 0.05). Additionally, as the canine angulation to the midline increased, the canine was more likely to be removed (P < 0.05). The orthodontists' decision to expose or remove an impacted upper permanent canine, based on radiographic information, seems to be primarily guided by two factors: labio-palatal crown position and angulation to the midline. PMID- 10867074 TI - Orthodontic treatment with fixed appliances in the General Dental Service in Scotland. AB - The records of 128 subjects treated by orthodontic specialists with fixed appliances in the General Dental Service in Scotland were randomly selected from the Scottish Dental Practice Board for analysis. The results of the study revealed that: (1) Mean post-treatment Peer Assessment Rating was 7.8 +/- 4.6 PAR points. (2) Mean reduction in PAR score was 14.9 +/- 10.6 PAR points. (3) Mean percentage reduction in PAR was 59 per cent. (4) Twenty-eight per cent of the cases were 'greatly improved', 15 per cent of the cases were made 'worse or no different'. (5) Median duration of treatment was 15 months, with a range of 2-41 months. (6) Multiple regression analysis showed that 82 per cent of the variability of PAR change could be predicted by the pretreatment PAR scores and the number of arches treated. Post-treatment PAR scores and duration of treatment could not be predicted with adequate reliability. It was concluded that although about 75 per cent of the cases were treated to at least acceptable alignment and occlusion, the modest average percentage reduction in PAR score could be explained by the low average initial PAR score and the borderline need for treatment in many cases. PMID- 10867075 TI - Investigation of a hydrophilic primer for orthodontic bonding: an in vitro study. AB - A common reason for bond failure is moisture contamination. This study investigates the in vitro bond strength of brackets bonded using a new hydrophilic primer, designed to be insensitive to moisture, and compares it with a conventional primer. Using a standardized technique, the in vitro bond strength of brackets bonded with the hydrophilic primer was compared to identical brackets bonded with a conventional primer. Although designed to be moisture insensitive, the directions for use stipulate drying the teeth before bonding. Therefore, for the purposes of comparison with a conventional primer the experiment was conducted under dry conditions. The results were analysed using the Weibull distribution modelling. The median bond strength with the hydrophilic primer (6.43 MPa, 95 per cent C.I. 7.69-9.50) was significantly lower (P = 0.0001) than the conventional primer (8.71 MPa, 95 per cent C.I. 5.89-7.59). The Weibull distribution modelling showed that brackets bonded with the hydrophilic primer were 3.96 times more at risk of failure (95 per cent C.I.: 2.39-6.56; P <0.0001). The bond strength at which 5 per cent of the brackets failed was also lower for the hydrophilic primer. The bond strengths obtained with the hydrophilic primer were significantly lower than with the conventional primer. Although the median bond strength values were promising, the laboratory results for this particular hydrophilic primer were disappointing when using the Weibull analysis, where the whole distribution of bond strength is taken into account. PMID- 10867076 TI - Undergraduate orthodontic education in Europe. PMID- 10867077 TI - Evidence-based orthodontics--how do I assess the evidence? PMID- 10867078 TI - Resorbable implants (plates and screws) in orthognathic surgery. PMID- 10867079 TI - We learn the most from our failures. PMID- 10867081 TI - Orthodontic therapists--the current situation. AB - The promise of the U.K. being allowed to use auxiliary help in orthodontics is slowly gaining momentum. At long last, key factors are under discussion, such as permitted duties, length of training, etc. This article describes the present situation and highlights the disappointing rate of progress PMID- 10867082 TI - Maurice Samuel Berman. PMID- 10867083 TI - Noninvasive strategies for the estimation of cardiac risk in stable chest pain patients. The Economics of Noninvasive Diagnosis (END) Study Group. AB - Effective allocation of medical resources in stable chest pain patients requires the accurate diagnosis of coronary artery disease and the stratification of future cardiac risk. We studied the relative predictive value for cardiac death of 3 commonly applied noninvasive strategies, clinical assessment, stress electrocardiography, and myocardial perfusion tomography, in a large, multicenter population of stable angina patients. The multicenter observational series comprised 7 community and academic medical centers and 8,411 stable chest pain patients. All patients underwent pretest clinical screening followed by stress (exercise 84% or pharmacologic 16%) electrocardiography and myocardial perfusion tomography. Risk-adjusted multivariable Cox proportional hazards models were developed to predict cardiac death. Kaplan-Meier rates of time to cardiac catheterization were also computed. Cardiac mortality was 3% during the 2.5 +/- 1.5 years of follow-up. The number of infarcted vascular territories and pretest clinical risk factors were strong predictors of cardiac mortality, whereas the number of ischemic vascular territories gained increasing importance when determining post-test resource use requirements (i.e., the decision to perform cardiac catheterization). Exertional ST-segment depression in a population with a high frequency of electrocardiographic abnormalities at rest was not a significant differentiator of cardiac death risk. Stable chest pain patients are accurately identified as being at high risk for near-term cardiac events by both physicians' screening clinical evaluation and by the results of stress myocardial perfusion imaging. Disease management strategies for stable chest pain patients aimed at risk reduction should incorporate knowledge of relevant end points in treatment and guideline development. PMID- 10867084 TI - Rates of progression of coronary calcium by electron beam tomography. AB - In this study, we sought to determine the rate of progression of atherosclerosis using coronary calcium scores derived from electron beam tomography (EBT). We studied a variety of disease states (hypertension, high cholesterol, tobacco use, diabetes mellitus) followed for 1 to 6.5 years. We evaluated 299 asymptomatic persons (227 men and 72 women) who underwent 2 consecutive EBT scans at least 12 months apart. The average change in the calcium score (Agatston method) for the entire group was 33.2 +/- 9.2%/year. The treated group (receiving statins) demonstrated an average increase in calcium scores of 15 +/- 8%/year compared with 39 +/- 12%/year for untreated patients (p <0.001). Among the 60 patients on statin monotherapy, 37% had a decrease in the calcium score from baseline to follow-up scan. The relative increase in calcium scores did not vary significantly by gender or risk factors, with the exception of statin-treated hypercholesterolemic subjects. Scores of zero on the initial scan portend a low likelihood of significant calcific deposits on repeat scanning. Only 2 of 81 participants (2%) with scores of zero at baseline had scores >10 on repeat study. In this study, statin therapy induced a 61% reduction in the rate of coronary calcium progression. This study demonstrates that EBT may be a useful tool in assessing efficacy of different interventions to retard progression of atherosclerosis, noninvasively, over relatively short time periods. PMID- 10867085 TI - Transthoracic stress echocardiography with transesophageal atrial pacing for bedside evaluation of inducible myocardial ischemia in patients with new-onset chest pain. AB - To date, there are no data on the feasibility and accuracy of bedside pacing stress echocardiography in patients admitted to the hospital with new-onset chest pain or unstable angina. We evaluated the feasibility of pacing stress echocardiography and examined its correlation with myocardial perfusion stress scintigraphy (rest thallium-201/stress technetium-99m sestamibi dual-isotope myocardial perfusion single-photon emission computerized tomography) performed within 24 hours of the pacing stress echocardiography test. We studied 70 consecutive patients after acute myocardial infarction had been excluded. The bedside pacing stress echocardiography test was performed with 10Fr transesophageal pacing catheters. We found pacing stress echocardiography to be feasible and safe (3% minor adverse event rate) at the patients' bedside. Target heart rate of >85% of the age-predicted heart rate was achieved in 96% of patients, and the mean rate-pressure product was 22,644 +/- 4,520 beats/min/mm Hg. The mean duration of the bedside pacing stress echocardiography test including technical preparations and image interpretation was 41 +/- 7 minutes. Pacing stress echocardiography and myocardial perfusion stress scintigraphy correlated well for identification or exclusion of inducible myocardial ischemia in 63 of 70 patients (90%) (kappa 0.81, p <0.001). The extent of inducible myocardial ischemia by vascular territories correlated with myocardial perfusion stress scintigraphy in 52 of 70 patients (74%) (kappa 0.6, p <0.001). We conclude that bedside pacing stress echocardiography is feasible and safe, and highly correlates with myocardial perfusion stress scintigraphy for identifying inducible myocardial ischemia in patients with new onset of chest pain or unstable angina. PMID- 10867086 TI - Program participation, exercise adherence, cardiovascular outcomes, and program cost of traditional versus modified cardiac rehabilitation. AB - Common concerns with the traditional protocol (TP) for cardiac rehabilitation include suboptimal program participation, poor facilitation of independent exercise, the use of costly continuous electrocardiographic (ECG) monitoring, and lack of insurance reimbursement. To address these concerns, a reduced cost modified protocol (MP) was developed to promote independent exercise. Eighty low- to moderate-risk cardiac patients were randomized to a TP (n = 42) or a MP (n = 38) and were compared over 6 months on program participation, exercise adherence, cardiovascular outcomes, and program costs. During month 1, patients followed identical regimens, including 3 ECG-monitored exercise sessions/week, with encouragement to achieve >/=5 thirty-minute sessions/week. In week 5, the TP continued with a facility-based regimen including 3 exercise sessions/week for 6 months and used ECG monitoring the initial 3 months. The MP discontinued ECG monitoring in week 5 and were gradually weaned to an off-site exercise regimen that was complemented with educational support meetings and telephone follow-up. Compared with TP patients, MP patients had higher rates of off-site exercise over 6 months (p = 0.05), and total exercise (on site + off site) during the final 3 months (p = 0.03). Also, MP patients were less likely to drop out (p = 0.05). Both protocols promoted comparable improvements in maximal oxygen uptake (p <0.05), blood lipids (p <0.001), and hemodynamic measurements (p <0.002). The MP cost $738 less/patient than the TP and required 30% less staff (full-time equivalents). These results suggest that a reduced cost MP was as effective as an established TP in improving physiologic outcomes while demonstrating higher rates of exercise adherence and program participation. Thus, the MP or a similar protocol has applicability to hospitals with large capitated or managed care populations to provide cost-effective cardiovascular risk reduction to patients. PMID- 10867087 TI - Use of resources, quality of life, and clinical outcomes in patients with and without new Q waves after thrombolytic therapy for acute myocardial infarction (from the GUSTO-I trial). AB - Previous reports indicate that patients who do not develop Q waves after thrombolytic therapy are a different population with a better long-term survival than those who do develop Q waves. However, the use of resources, quality of life, and health status of this population have not been fully evaluated. Using data from the Economics and Quality of Life subset of the Global Utilization of Streptokinase and tPA for Occluded Arteries study, we examined 30-day and 1-year mortality, use of resources, and quality-of-life measures among 1,830 of 3,000 patients with acute myocardial infarction and ST-segment elevation treated with thrombolytic therapy. At hospital discharge, 555 patients (30.2%) had not developed Q waves. These patients had lower mortality than patients with Q waves at 30 days (1.6% vs 4.5%, p <0.01) and at 1 year (4.7% vs 6.8%, p <0.04). Recurrent chest pain and dyspnea were similar at 30 days and 1 year. Patients without Q waves had significantly more angiography and trends toward higher readmission, revascularization, and use of calcium antagonists at 30 days. Angiography, revascularization, readmission, and quality of life were equivalent from 30 days to 1 year, with no sign of late instability. Logistic regression analysis showed an association between in-hospital revascularization and better survival and quality of life at 1 year. Conversely, there was no association between in-hospital use of calcium antagonists and outcome to explain the lower mortality in non-Q-wave patients. The absence of Q waves after thrombolytic therapy is a marker of success, implying better prognosis and equivalent quality of life, use of resources, and health status than for patients with Q-wave acute myocardial infarction and no sign of long-term unstable clinical course. PMID- 10867088 TI - Angiographic and clinical characteristics associated with increased in-hospital mortality in elderly patients with acute myocardial infarction undergoing percutaneous intervention (a pooled analysis of the primary angioplasty in myocardial infarction trials). AB - Advanced age is associated with increased mortality in acute myocardial infarction (AMI) but the mechanism remains unclear. We performed a pooled analysis of 3,032 patients from the Primary Angioplasty in Myocardial Infarction (PAMI)-2, Stent-PAMI, and PAMI-No Surgery On Site trials to determine which clinical, hemodynamic, and angiographic characteristics in the elderly were associated with in-hospital death. There were 452 patients aged >/=75 years and 2,580 patients aged <75 years. Older patients had a lower number of risk factors for coronary artery disease but more comorbidities. Acute catheterization demonstrated more 3-vessel disease, higher left ventricular (LV) end-diastolic pressure, lower LV ejection fraction, and higher initial rates of Thrombolysis In Myocardial Infarction (TIMI) trial 2 or 3 flow. Elderly patients were equally likely to undergo percutaneous intervention but had a lower procedural success rate and lower rates of final TIMI 3 flow, and older patients were more likely to have post-AMI complications. In-hospital mortality was 10.2% and 1.8%, respectively (p = 0.001). Cardiac and noncardiac mortality was higher in elderly patients, and no significant differences in causes of death were identified. Multivariate analysis revealed that the strongest predictors of death were age >/=75 years, lower LV ejection fraction, lower final TIMI flow, higher Killip class, need for an intra-aortic balloon pump (IABP), and post-AMI stroke/transient ischemic attack, or significant arrhythmia. Despite avoiding thrombolysis, elderly patients remain at increased risk of bleeding, stroke, and other post-AMI complications, and death. Cardiac risk factor analysis and acute catheterization offer prognostic information but do not completely explain the mechanism of increased in-hospital mortality in the elderly. PMID- 10867089 TI - Detection of coronary restenosis by exercise electrocardiography thallium-201 perfusion imaging and coronary angiography in asymptomatic patients after percutaneous transluminal coronary angioplasty. AB - Noninvasive detection of restenosis in patients remaining asymptomatic after percutaneous transluminal coronary angioplasty (PTCA) remains a major clinical problem. The value of exercise electrocardiography (ECG) and exercise redistribution thallium-201 single-photon emission computed tomography (SPECT) in detecting restenosis in such patients remains uncertain. Discordances between these tests and coronary angiography is a common situation. We studied 179 consecutive patients remaining asymptomatic after successful PTCA (208 vessels), who underwent 6 +/- 2 months of exercise ECG, SPECT, and coronary angiography. We sought to assess the diagnostic value of the noninvasive tests compared with coronary angiography, and identify the determinants of discordances between the tests. Restenosis (diameter stenosis >50%) was detected in 39% of patients and in 37% of vessels. The overall sensitivity, specificity, and accuracy for exercise ECG and SPECT in detecting restenosis in individual vessels were, respectively, 53% versus 63% (p = 0.06), 59% versus 77% (p = 0.0001), and 57% versus 72% (p = 0. 0001). On multivariate analysis, positive exercise ECG was associated with higher heart rate response (p = 0.02), incomplete revascularization (p = 0.004), and angiographic restenosis (p = 0. 03), whereas positive SPECT was associated with incomplete revascularization (p = 0.02), infarct-related artery PTCA (p = 0.01), and angiographic restenosis (p = 0.0001). Accuracies of the 2 tests were not significantly different in patients with incomplete revascularization or PTCA of an infarct-related vessel. Overall, SPECT is more accurate than exercise ECG in detecting asymptomatic restenosis. Nevertheless, incomplete revascularization and PTCA of an infarct-related artery could cause reversible perfusion defects regardless of restenosis, reducing the diagnostic value of SPECT in such patients. PMID- 10867090 TI - Safety and efficacy of percutaneous coronary interventions performed immediately after diagnostic catheterization in northern new england and comparison with similar procedures performed later. AB - "Ad hoc" percutaneous coronary interventions (PCIs)-those performed immediately after diagnostic catheterization-have been reported in earlier studies to be safe with a suggestion of higher risk in certain subgroups. Despite increasing use of this strategy, no data are available in recent years with new device technology. We studied use of an ad hoc strategy in a large regional population to determine its use and outcomes compared with staged procedures. A database from the 6 centers performing PCIs in northern New England and 1 center in Massachusetts was analyzed. During 1997, excluding only patients requiring emergency procedures or those with a prior PCI, 4,136 PCIs were performed, 1,748 (42.3%) of these being ad hoc procedures. Patients having ad hoc procedures were less likely to have peripheral vascular disease, renal failure, prior myocardial infarction, or coronary artery bypass surgery, congestive heart failure, or poor left ventricular function, and more likely to have received preprocedural intravenous heparin or nitroglycerin or to have required an urgent procedure. Narrowings treated during ad hoc procedures were less frequently types B and C or in saphenous vein grafts. Adjusted rates of clinical success were not different between ad hoc and non-ad hoc procedures (93.7% vs 93.6%); there was no difference in the incidence of death (0.6% vs 0.5%), emergency (0. 9% vs 0.8%) or any (1.4% vs 0.8%) coronary artery bypass surgery, or myocardial infarction (2.6% vs 2.0%). As currently practiced in our region, ad hoc intervention is used selectively with outcomes similar for ad hoc and non-ad hoc procedures. PMID- 10867091 TI - Incremental reduction of serum total cholesterol and low-density lipoprotein cholesterol with the addition of plant stanol ester-containing spread to statin therapy. AB - This study compares the effect of plant stanol ester spread with a placebo spread on cholesterol in patients taking statin therapy, but who still had elevated low density lipoprotein (LDL) cholesterol. This was a randomized, double-blind, placebo-controlled clinical trial, with 67 women and 100 men with LDL cholesterol >/=130 mg/dl and triglycerides /=3 factors. A maximal exercise test performed in asymptomatic men free of cardiovascular disease does appear to be a worthwhile tool in predicting future risk of CHD death. An abnormal exercise test is a more powerful predictor of risk in those with than without conventional risk factors. PMID- 10867093 TI - Clinical characteristics of patients intolerant to VVIR pacing. AB - The incidence and clinical predictors of the development of intolerance to VVIR pacing have not been extensively studied in prospective long-term randomized trials comparing different pacing modes. The frequency and clinical factors predicting intolerance to ventricular pacing are controversial. The Pacemaker Selection in the Elderly (PASE) Trial enrolled 407 patients aged >/=65 years in a 30-month, single-blind, randomized, controlled comparison of quality of life and clinical outcomes with ventricular pacing and dual-chamber pacing in patients undergoing dual-chamber pacemaker implantation for standard clinically accepted indications. We reviewed the clinical, hemodynamic, and electrophysiologic variables at the time of pacemaker implantation in 204 patients enrolled in the PASE trial and randomized to the VVIR mode, some of whom subsequently required crossover (reprogramming) to DDDR pacing. During a median follow-up of 555 days, 53 patients (26%) crossed over from VVIR to DDDR pacing. A decrease in systolic blood pressure during ventricular pacing at the time of pacemaker implantation (p = 0.001), use of beta blockers at the time of randomization (p = 0.01), and nonischemic cardiomyopathy (p = 0.04) were the only variables that predicted crossover in the Cox multivariate regression model. After reprogramming to the dual-chamber mode, patients showed improvement in all aspects of quality of life, with significant improvements in physical and emotional role. The high incidence of crossover from VVIR to DDDR pacing along with significant improvements in quality of life after crossover to DDDR pacing strongly favors dual-chamber pacing compared with single-chamber ventricular pacing in elderly patients requiring permanent pacing. PMID- 10867094 TI - Atrial fibrillation after beating heart surgery. AB - Postoperative atrial fibrillation (AF) is a frequent adverse event after coronary artery bypass grafting (CABG) and may negatively affect the early clinical outcome. We sought to investigate the risk factors, prevalence, and prognostic implications of postoperative AF in patients submitted to CABG without cardiopulmonary bypass (off-pump). The study population comprised 969 patients, 645 men (67%) and 324 women (33%) who had off-pump CABG at the Washington Hospital Center from January 1987 to May 1999. Preoperative AF patients were excluded (n = 15). Two hundred six patients (age 69 +/- 10 years, 137 men [66%]) developed AF, whereas 763 patients (age 61 +/- 12 years, 508 men [67%]) did not. Predictors of AF included age >75 years (odds ratio [OR] 3.0, 95% confidence interval [CI] 1.9 to 4.5; p <0.001), history of stroke (OR 2.1, CI 1.2 to 3.7; p = 0. 007), postoperative pleural effusion requiring thoracentesis (OR 3.2, CI 1.0 to 9.4; p = 0.03), and postoperative pulmonary edema (OR 5.1, CI 1.2 to 21; p = 0.02). Minimally invasive direct CABG was associated with a lower incidence of AF (OR 0.4, CI 0.3 to 0.7; p <0. 001). AF was associated with a prolonged postoperative hospital stay (9 +/- 6 days AF vs 6 +/- 5 days no AF, p <0.001). In hospital mortality was significantly higher in AF patients (3% AF vs 1% no AF, p = 0.009). Patients with persistent AF had a higher postoperative in-hospital stroke rate than patients without persistent AF (9% vs 0. 6%, p <0.001). AF after beating heart surgery is associated with a higher in-hospital morbidity, mortality, and prolonged hospital stay. A minimally invasive surgical approach (minimally invasive direct CABG) is associated with a lower risk of AF. PMID- 10867096 TI - Richard John Bing, MD: a conversation with John Willis Hurst, MD. PMID- 10867095 TI - Nonobstructive aortic valve calcium as a window to atherosclerosis of the aorta. AB - Aortic valve calcium without stenosis and mitral annulus calcium (MAC) are known to correlate with atherosclerotic risk factors. Recently, it has been reported that MAC is associated with atherosclerosis of the cardiovascular system, suggesting MAC as an atherosclerotic process by itself. Hence, the aim of the present study was to determine whether a similar association between aortic valve calcium and aortic atheroma exists. Ninety-six patients (54 men and 42 women, mean age 72 +/- 12 years) with aortic valve calcium who underwent transesophageal echocardiography (TEE) formed the study group. They were compared with 92 sex- and age-matched patients without aortic valve calcium who underwent TEE for the same indications during the same period. The presence and echocardiographic features of aortic atheromas were evaluated by TEE. No differences were found between the groups in risk factors for atherosclerosis or in indications for referral for TEE. Significantly higher rates were found in the aortic valve calcium group for prevalence of aortic atheroma (86% vs 30%, p = 0.001). This significant trend was also consistent with the dimension and complexity of the atheromas. On multivariate analysis aortic valve calcium, and MAC were the only independent predictors of aortic atheroma (p = 0.0001, 0.006 respectively). We conclude that there is a significant association between the presence of aortic valve calcium and the presence and severity of aortic atheroma. Thus, aortic valve calcium may serve as a window to atherosclerosis of the aorta. PMID- 10867097 TI - Effects of simvastatin or hormone replacement therapy, or both, on fibrinogen, factor VII, and plasminogen activator inhibitor levels in postmenopausal women with proven coronary artery disease. PMID- 10867098 TI - Relation between C-reactive protein levels on admission and pattern of acute myocardial infarction onset. PMID- 10867099 TI - Effectiveness of rotational atherectomy in narrowed left internal mammary artery grafts to the left anterior descending coronary artery. PMID- 10867100 TI - Effectiveness of rotational atherectomy in aortocoronary saphenous vein grafts. PMID- 10867101 TI - Prevalence of the brugada syndrome in an apparently healthy population. PMID- 10867102 TI - Comparison of intravenous adrenomedullin with atrial natriuretic peptide in patients with congestive heart failure. PMID- 10867103 TI - The spectrum of pulmonary abnormalities on computed chest tomographic imaging in patients with advanced heart failure. PMID- 10867104 TI - Pilot study of guided imagery use in patients with severe heart failure. PMID- 10867105 TI - Analysis of agreement between dobutamine stress echocardiography and exercise nuclear angiography in severe aortic regurgitation. PMID- 10867106 TI - Dose and duration of fenfluramine-phentermine therapy impacts the risk of significant valvular heart disease. PMID- 10867107 TI - Clinical and echocardiographic features of hypereosinophilic syndromes. PMID- 10867108 TI - Usefulness of doppler echocardiography to determine the timing of surgery for supravalvar aortic stenosis. PMID- 10867109 TI - Effect of low molecular weight heparin on coronary endothelial function in acute cellular heart transplant rejection. PMID- 10867110 TI - Anatomic landmarks for use when measuring intracardiac pressure with fluid-filled catheters. PMID- 10867111 TI - Effects of the high-affinity corticotropin-releasing hormone receptor 1 antagonist R121919 in major depression: the first 20 patients treated. AB - Clinical and preclinical data suggest that unrestrained secretion of corticoctropin-releasing hormone (CRH) in the CNS produces several signs and symptoms of depression and anxiety disorders through continuous activation of CRH(1) receptors. This led to the development of drugs that selectively antagonize CRH(1) receptors suppressing anxiety-like behavior in rats and also in monkey models of anxiety. These findings led to a clinical development program exploring the antidepressive potential of R121919, a water-soluble pyrrolopyrimidine that binds with high affinity to human CRH(1) receptors and is well absorbed in humans. This compound was administered to 24 patients with a major depressive episode primarily in order to investigate whether its endocrine mode of action compromises the stress-hormone system or whether other safety and tolerability issues exist. The patients were enrolled in two dose-escalation panels: one group (n=10) where the dose range increased from 5-40 mg and another group (n=10) where the dose escalated from 40 to 80 mg within 30 days each. Four patients dropped out because of withdrawal of consent to participate (three cases) or worsening of depressive symptomatoloy in one case. We found that R121919 was safe and well tolerated by the patients during the observation period. Moreover, the data suggested that CRH(1)-receptor blockade does not impair the corticotropin and cortisol secretory activity either at baseline or following an exogenous CRH challenge. We also observed significant reductions in depression and anxiety scores using both, patient and clinician ratings. These findings, along with the observed worsening of affective symptomatology after drug discontinuation, suggests that the pharmacological principle of CRH(1) receptor antagonism has considerable therapeutic potential in the treatment and the prevention of diseases where exaggerated central CRH activity is present at baseline or following stress exposure. PMID- 10867112 TI - Fluoxetine decreases concentrations of 3 alpha, 5 alpha tetrahydrodeoxycorticosterone (THDOC) in major depression. AB - There is evidence for a differential alteration in the concentrations of 3 alpha reduced neuroactive steroids in major depression. Because it has been suggested that fluoxetine may shift the activity of the 3 alpha-hydroxysteroid oxidoreductase towards the reductive direction, treatment of major depression may be accompanied by a further increase in plasma 3 alpha, 5 alpha tetrahydrodeoxycorticosterone (THDOC) concentration. We studied eight male depressed patients before and after treatment with fluoxetine and compared them to healthy age-matched control subjects. Blood samples were quantified for 3 alpha, 5 alpha-tetrahydroprogesterone, 3 alpha,5 beta-tetrahydroprogesterone (THP) and THDOC by means of a highly sensitive combined gas chromatography/mass spectrometry analysis. Compared to control subjects, concentrations of THDOC were higher in depressed patients and decreased after fluoxetine treatment. In contrast, THP concentrations were lower in depressed patients and increased after fluoxetine treatment. Our results give further evidence for a disequilibrium of 3 alpha-reduced neuroactive steroids in major depression, which is normalized by treatment with fluoxetine. PMID- 10867113 TI - Construct validity of life chart functioning scales for use in naturalistic studies of bipolar disorder. AB - This study examined the construct validity of the functional impairment scales of the National Institute of Mental Health (NIMH) prospective life-charting methodology (LCM-p(TM)). Twelve male and 28 female bipolar participants were recruited from the community through advertisements. Diagnoses of bipolar I or II were confirmed using Structured Clinical Interview for DSM-IV Axis I Disorders (SCID-I). Patients completed life charts for three consecutive months. At the end of each month, a trained clinician administered the Hamilton Depression Rating Scale (HDRS) and the Young Mania Rating Scale YMRS by telephone, followed by ratings using the Global Asessment of Functioning (GAF), and Clinical Global Impression (CGI) for bipolar disorder. Data was analyzed separately for each month (time 1, time 2, and time 3). Complete data was available for 35 participants at time 1, 36 at time 2 and 32 at time 3. Spearman correlations demonstrated significant convergent validity at times 2 and 3 for life chart measures of mania (HIGH) and CGI-mania, and at all three times for life chart depression (LOW) and CGI-depression, with corresponding discriminant validities. The GAF was positively correlated with HIGH and negatively correlated with LOW at time 2 and 3 only. HAM-D and LOW also showed convergent and discriminant validity for all three times. For HIGH and YMRS, however, there was a strong significant correlation at time 3 only. While the use of heterogeneous methods maximizes differences between measures, it also appears that bipolar patients are less consistent in reporting functional impairment due to mania than due to depression. The construct of life chart dysfunction due to mania does not consistently measure the same construct as similar clinician ratings of mania. Life chart dysfunction due to depression shows higher construct validity. PMID- 10867114 TI - Eight-month functional outcome from mania following a first psychiatric hospitalization. AB - The aim of this study was to identify how different areas of function (role performance, interpersonal relationships, sexual activity and recreational enjoyment) differentially recover from a manic episode during the 8 months following a first psychiatric hospitalization. Fifty patients with bipolar disorder, 16-45 years of age, who met the criteria for a current manic episode were recruited during their first psychiatric hospitalization. Forty-two (84%) of these participated in follow-up. Patients were evaluated using structured and semi-structured clinical instruments and the four areas of functional outcome were assessed with the LIFE interview. Recovery of the four areas of function were compared using survival and correlational analyses. Logistic regression identified factors associated with functional outcome. The four aspects of function were not significantly intercorrelated at baseline or during follow-up. Moreover, the survival curves for the different areas of function significantly differed. Specifically, patients demonstrated better recovery of sexual activity and worse recovery of recreational enjoyment than the other areas of function. Different clinical and demographic variables predicted recovery of the different areas of function. In conclusion, following a first manic episode, recovery of psychosocial function can be divided into separate components, i.e., role function, interpersonal relationships, sexual activity and recreational enjoyment, that appear to be relatively independent. Further clarification of recovery of these different areas of function may lead to better integrated treatments that maximize functional improvement early in the course of bipolar disorder. PMID- 10867115 TI - Comparison of preferences for health outcomes in schizophrenia among stakeholder groups. AB - BACKGROUND: To determine the effectiveness of psychiatric interventions for use in cost-effectiveness analysis, we assessed the feasibility of using a multimedia computer survey to study preferences (utilities) for health outcomes among persons with schizophrenia, family members of persons with schizophrenia, health professionals, and the public. METHODS: We developed videos depicting two patterns of mental health impairment in schizophrenia, both with and without pseudo-parkinsonism side-effects. These descriptions were integrated into a computer program that measured preferences using two psychometric methods: (1) standard gamble and (2) a visual analog scale. This program was used to compare preferences among potential stakeholder groups. RESULTS: 20 persons with schizophrenia, 11 family members, 20 healthy volunteers and 14 health professionals participated in the computerized interview. All but one subject completed the survey. The correlation among ratings of various states was high (r=0.7-0.95) and ratings were internally consistent in 89% of participants. There were significant differences in values between groups for health states (p=0.024) and in values for the effects of pseudo-parkinsonism on quality of life (p<0.001). Persons with schizophrenia valued the disease states more highly and placed more significance than did other groups on the effects of pseudo parkinsonism on quality of life. CONCLUSIONS: Computer-based multimedia techniques can offer a feasible and valid approach to measure preferences for outcomes in schizophrenia. The study found significant differences in preferences among stakeholder groups for schizophrenia outcomes. Further work is needed to clarify how these differences affect clinical decision-making and policies for health resource allocation. PMID- 10867116 TI - The primary care evaluation of mental disorders (PRIME-MD), German version: a comparison with the CIDI. AB - There is a need for the development and evaluation of diagnostic instruments suitable for daily use in primary care offices that can improve recognition rates of psychopathology. The objective of this study is the comparison of the German version of the Primary Care Evaluation of Mental Disorders (PRIME-MD), a short structured diagnostic instrument, with the Composite International Diagnostic Interview (CIDI) and to gather some information on the usefulness of the PRIME MD. Seven hundred and four patients were assessed three times, once using the physician's clinical judgement, subsequently, administering the PRIME-MD, DSM-IV version and finally, with the CIDI. The CIDI was administered on a different occasion within 1 week after the PRIME-MD evaluation by independent interviewers over the telephone. At the end of the study, the participating physicians answered a few feedback questions on the usefulness of PRIME-MD. Sensitivity (0.73), specificity (0.67), overall accuracy (0.70) were good for any psychiatric disorder. According to the diagnostic categories of mood disorders, anxiety disorders, eating disorders, and alcohol-related disorders, the sensitivity of PRIME-MD was good (0.67-0.80). For somatoform disorders, sensitivity was poor. Specificity, accuracy, and negative predictive values were good-to-excellent for all diagnostic categories. Kappas range from poor (0.12 somatoform disorders) to satisfactory (0.62 alcohol related disorders). Average time for administering PRIME-MD was 11 min. The results demonstrated that the German version of the PRIME-MD may be useful in primary care settings to enhance recognition of mental disorders by primary care physicians, even without being formally trained in diagnosing mental disorders. PMID- 10867117 TI - Sociodemographic predictors of temperament and character. AB - The Unified Biosocial Theory of Personality postulates that human personality is organized around four temperaments - Novelty Seeking, Harm Avoidance, Reward Dependence, and Persistence - and three characters - Self-Directedness, Cooperativeness, and Self-Transcendence. The objective of the present study was to investigate the influence of sociodemographic factors on temperament and character without the confounding influence of mental disorders. Volunteers (n=94) did not meet criteria for any Axis I and Axis II diagnosis, had no first degree relatives with mental disorders, and were medically healthy. After giving written informed consent, volunteers completed the Temperament and Character Inventory. Analyses were conducted to determine the degree of association of each sociodemographic factor (i.e., age, gender, ethnicity, marital status, educational attainment, and occupational status) to personality dimension, while controlling for possible interactions with other sociodemographic factors. Partial correlation analysis showed a significant association between gender and Reward Dependence, and occupational status was significantly related to Reward Dependence, Cooperativeness, and Self-Transcendence. Stepwise regression analysis indicated that gender and occupational status were significant predictors of Reward Dependence. Occupational status was the only predictor of Cooperativeness and Self-Transcendence. These data suggest that sociodemographic factors should be considered in studies investigating temperaments and characters as defined by the Unified Biosocial Theory of Personality. PMID- 10867118 TI - Changes in directed attention and short-term memory in depression. AB - This study examined changes in directed attention and short-term memory in depression using a newly constructed battery of computerized measures. A repeated measures design was used with two sample groups; 25 individuals meeting DSM-IV criteria for Major Depression and a group-matched comparison sample of 27. Both groups were tested at three points in time over a 10-week period. Test-retest reliability of the measures was examined. Profile analysis demonstrated that there were differences between the depressed and comparison groups in both directed attention and short-term memory. Recommendations for specific improvements in the testing battery are discussed. The ability to detect changes in directed attention and short-term memory may have clinical utility in early detection of impending onset of depression or subtle residual symptoms of an acute episode that may impair functioning or signal a relapse. PMID- 10867119 TI - A promoter polymorphism in the monoamine oxidase A gene and its relationships to monoamine metabolite concentrations in CSF of healthy volunteers. AB - Concentrations of monoamine metabolites (MM) in lumbar cerebrospinal fluid (CSF) have been used extensively as indirect estimates of monoamine turnover in the brain. We investigated possible relationships between a putative functional promoter polymorphism in the monoamine oxidase A (MAOA) gene and CSF concentrations of homovanillic acid (HVA), 5-hydroxyindoleacetic acid (5-HIAA), and 3-methoxy-4-hydroxyphenylglycol (MHPG) in healthy volunteers (n=88). Among women (n=37), those carrying at least one copy of the alleles associated with more efficient transcription displayed higher concentrations of HVA (p=0.01) and 5-HIAA (p=0.01). In men (n=51), however, there was a tendency in the opposite direction. The results suggest that MAOA genotypes may participate differentially in the regulation of dopamine and serotonin turnover rates under presumed steady state in the central nervous system. The results should be interpreted with caution until replicated because of the limited sample size. PMID- 10867120 TI - Habituation of the auditory evoked field component N100m in male patients with schizophrenia. AB - The auditory evoked field (AEF) component N100m represents the most prominent and stable peak of the AEF, and its alterations in patients with schizophrenia are an extensive topic of neuropsychiatric research. In our current study, the degree of N100m habituation was investigated in 20 male schizophrenics and 19 healthy male controls. Participants were stimulated monaurally with 270 trials of 1000 Hz tones separated by an interstimulus interval between 800-1800 ms. The trial sample of the bilaterally recorded AEF was separated into three consecutive blocks of 90 trials and these blocks were compared with each other. The mean global field power (MGFP) of the N100m decreased on average 9.1% from the first to the third trial block, while the N100m latency was increasing. The analyses of the influence of habituation revealed a systematic change of dipole location in inferior-superior direction, mainly in the left hemisphere. This habituation effect was found to be the same for both groups. The groups also did not differ in the N100m latency increase and MGFP decrease, except for one parameter. The right-hemispheric MGFP decrease from the first to second block was found to be more pronounced in patients compared to controls. However, this difference was related to medication with clozapine. Overall, the habituation behaviour of the N100m seems to be undisturbed in schizophrenic patients. PMID- 10867121 TI - A replication study on P300 single trial analysis in schizophrenia: confirmation of a reduced number of 'true positive' P300 waves. AB - A single trial analysis of event-related potentials (auditory odd-ball paradigm) of 20 schizophrenics was performed in comparison to matched healthy controls. The purpose of the study was to test the hypothesis that in schizophrenia the well known P300 amplitude reduction of averaged event-related potentials is due to fewer elicited single trial P300 waves. The results of the present study support this finding of our previous exploratory investigation and point to the view that schizophrenics reveal basal disturbances in information processing due to inadequately elicited electrophysiological responses to target stimuli. PMID- 10867122 TI - Foreword PMID- 10867123 TI - Eicosanoids and the esophagus. AB - Eicosanoids are products of arachidonic acid metabolism. Among the products produced are the prostaglandins and leukotrienes, products which are known to play important roles in health and disease of many gastrointestinal tissues. Here, we review current knowledge about eicosanoids in the esophagus, including production in healthy and diseased tissues and potential physiologic and pathophysiologic effects in two important esophageal mucosal disorders, reflux esophagitis and esophageal cancer. PMID- 10867124 TI - Eicosanoids and the stomach. AB - Eicosanoids are arachidonic acid derivatives that include prostaglandins, thromboxanes, and leukotrienes. During the last three decades, it has become evident that these bioactive lipids play a pivotal role in gastric physiology. The goal of the present review is to describe their involvement in the normal regulation of gastric secretion and gastric motility, as well as in gastric mucosal defense. Their role in gastric mucosal mitogenesis, apoptosis, inflammation, and immune modulatory responses is also discussed. PMID- 10867125 TI - Eicosanoids and the small intestine. AB - Prostaglandins play an important role in modulation of various physiologic processes in the small intestine. In this review, the involvement of prostaglandins in various small-intestinal functions including small-intestinal secretion, mucosal protection, epithelial and endothelial barrier function, and motility are discussed. PMID- 10867126 TI - Eicosanoids and the large intestine. AB - During the past three decades, many studies have been conducted to determine the precise role of eicosanoids in colorectal physiology and pathophysiology. This research has increased our understanding of bioactive lipid signaling, and may contribute to the development of more effective therapeutic modalities for digestive diseases in the future. The purpose of this report is to provide a brief overview of the role of eicosanoids in the colon and rectum. This information has been organized according to both functional and disease-related categories. The role of eicosanoids in colonic secretion, motility, inflammatory bowel disease, and colorectal neoplasia will be discussed. PMID- 10867127 TI - Eicosanoids and the liver. PMID- 10867128 TI - The EVI-1 gene--its role in pathogenesis of human leukemias. AB - EVI-1 (ecotropic virus integration site-1) was at first identified as an integration site of the murine leukemia retrovirus in murine myeloid leukemias. It is involved in pathogenesis of mouse and human leukemias. EVI-1 expression may be activated by retroviral insertion or is caused by chromosomal translocations. EVI-1 gene is located on human chromosome 3, spans over 100 kb and contains 12 exons with ten coding exons. EVI-1 gene encodes 1051 amino acids DNA binding protein containing ten zinc finger repeats organized in two domains. The 145 kDa EVI-1 protein is localized in the nucleus. The structure of the EVI-1 protein indicates that it functions as a transcriptional factor of the zinc finger family. The role of this transcription factor in myeloid cell transformation and the target genes of EVI-1 is still unknown. Occurrence of a few EVI-1 fusion transcripts was shown. The role of this fusion proteins is still unclear. Mouse and human sequences of the gene show a high degree of homology; 91% in nucleotide sequence and 94% in amino acid sequence. PMID- 10867129 TI - Insight into the mechanism of asparaginase-induced depletion of antithrombin III in treatment of childhood acute lymphoblastic leukemia. AB - Asparaginase (ASNase) is a widely used and successful agent against childhood acute lymphoblastic leukemia (ALL). Asparaginase cleaves asparagine (Asn) to give aspartic acid and ammonia, thereby depleting free Asn in the blood. However, treatment with ASNase has been implicated in significant reduction of plasma levels of the coagulation serine protease inhibitor (serpin) antithrombin III (AT3), predisposing patients to thromboembolic complications. Our investigation was designed to delineate the biochemical mechanism of AT3 depletion that can occur in the plasma of ALL patients undergoing ASNase therapy. SDS-PAGE showed no cleavage of purified AT3 following treatment with ASNase. Furthermore, purified AT3 treated with ASNase demonstrated no decrease in inhibitory activity. Human plasma and whole blood treated with approximate therapeutic concentrations of ASNase showed no loss of AT3 activity as detected by a plasma-based factor Xa inhibition assay. Treatment of a confluent monolayer of HepG2 (hepatocarcinoma) cells with ASNase showed no gross loss in AT3 message levels detected by rtPCR. However, a decrease of cell viability was observed in cultures treated with ASNase. Interestingly, medium from HepG2 cells treated with ASNase showed a marked decrease in secretion of AT3 and another serpin, heparin cofactor II. Collectively, these data show that ASNase has no direct effect on AT3 in blood or plasma, but that ASNase may affect plasma levels of AT3 by interfering with translation and/or secretion of the protein in liver cells. PMID- 10867130 TI - A phase I study of induction chemotherapy for older patients with newly diagnosed acute myeloid leukemia (AML) using mitoxantrone, etoposide, and the MDR modulator PSC 833: a southwest oncology group study 9617. AB - Older patients with acute myelogenous leukemia (AML) have overexpression of P glycoprotein (Pgp+), and this has been shown to correlate quantitatively with therapeutic outcome. Since Pgp-mediated efflux of cytotoxic drugs can be inhibited by the cyclosporine analogue, PSC 833, we investigated the use of this agent with a 5-day mitoxantrone/etoposide regimen in patients over age 55 with newly diagnosed AML. Previous studies suggested a 33% incidence of grade IV/V non hematologic toxicity with the use of mitoxantrone 10 mg/M(2) and etoposide 100 mg/M(2), each for 5 days, in this patient population. Since PSC 833 alters the pharmacokinetic excretion of MDR-related cytotoxins, this phase I dose-finding study was performed to identify doses of mitoxantrone/etoposide associated with a similar 33% incidence of grade IV/V non-hematologic toxicity, when given with PSC 833. Mitoxantrone/etoposide (M/E) doses were escalated in fixed ratio from a starting dose of M: 4 mg/M(2) and E: 40 mg/M(2), to M: 7 mg/M(2) and E: 70 mg/M(2), in successive cohorts of eight patients each. PSC 833 was well tolerated and the MTD of this M/E regimen with PSC 833 in this population was M: 6 mg/M(2) and E: 60 mg/M(2). The complete response (CR) rate for all patients was 50% (15/30) and was considerably higher for de novo than for secondary AML. These data suggest that the addition of PSC 833 to an M/E regimen for older patients with untreated AML is well tolerated but requires a reduction in M/E dosing to avoid increased toxicity. PMID- 10867131 TI - Pattern of immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements in childhood acute lymphoblastic leukemia in India. AB - In 120 cases of acute lymphoblastic leukemia (median age 8 years), IgH chain gene was rearranged in 99% B-Cell Precursor (BCP) ALLs and 13% T-ALLs. One or the other TCR locus was rearranged not only in all T-ALLs, but also in 87% of BCP ALLs. TCR-beta rearrangement in BCP-ALL was associated with a higher mean age at presentation (8.7 vs. 6.2 years, P=0.008), lower mean platelet counts (61.2x10(9)/l vs. 103.7x10(9)/l, P=0.003) and a poorer DFS (% cummulative survival 0 vs. 88.9+/-10.5, P=0.004). TCR-gamma rearrangement in T-ALL was associated with a higher mean WBC count (186.3x10(9)/l vs. 63. 4x10(9)/l, P=0.002). Also, the pattern of rearrangement of these genes appeared to be different from the West; viz. TCR-beta rearrangement in a higher proportion of BCP-ALLs (58%, 95% confidence intervals 45-69%), invariable deletion of Cgamma1 and only monoallelic rearrangement for TCR-delta locus. This repertoire of gene rearrangement may have a bearing on the poor treatment outcome reported previously from our geographic region. PMID- 10867132 TI - Treatment of patients with advanced chronic myelogenous leukemia with interferon alpha-2b and continuous oral cytarabine ocfosfate (YNK01): a pilot study. AB - The efficacy of continuous oral cytarabine ocfosfate (YNK01) (300 mg/day) in combination with interferon alpha (IFNalpha, 5x10(6) IU/day) was evaluated in patients with advanced chronic myelogenous leukemia, who previously failed to respond to IFNalpha-based therapies. Dose escalations up to 900 mg YNK01 were allowed in patients who failed to respond. In view of our results, four patients developed a complete hematological response after YNK01 was started. Among those who initially responded to YNK01, one complete cytogenetic response was achieved 18 months later. Although the data are preliminary, this is the first study showing that continuous administration of YNK01 along with IFNalpha is effective in patients with advanced chronic myelogenous leukemia. PMID- 10867133 TI - Descriptive epidemiology of de novo acute leukaemia in the Sultanate of Oman. AB - A hospital-based epidemiological study of de novo acute leukaemia was carried out in the Sultanate of Oman, a sparsely populated Arabian Gulf country which has undergone rapid and dramatic socio-economic changes recently. A total of 65/99 Omanis (66%) were diagnosed as acute lymphoblastic leukaemia (ALL) and 34/99 (34%) as acute myeloid leukaemia (AML). Trends and peak values in age-specific incidence rates for ALL are generally in line with those of Western countries. The proportion of T-ALL cases is however higher than that in Caucasian populations but lower than in many non-white populations. AML frequency particularly in childhood is similar to that observed in many African countries. PMID- 10867134 TI - Myelodysplastic syndrome and aplastic anemia--diagnostic and conceptual uncertainties. PMID- 10867135 TI - Refractory anemia with ringed sideroblasts with a low IPSS score progressed rapidly with de novo appearance of multiple karyotypic abnormalities and into acute erythroleukemia (AML-M6A). AB - We report here a case of refractory anemia with ringed sideroblasts (RARS) with a low risk group by the International Prognostic Scoring System (IPSS) at the time of diagnosis but had a rapid disease progression. Although the patient showed a normal male karyotype at the time of RARS diagnosis, his marrow cells had del(5)(q14) and add(17)(p12) abnormalities 2 months after the diagnosis, and later the marrow cells had multiple abnormalities and the patient expired 6 months after the initial diagnosis of RARS. The patient was diagnosed as having RARS with a low risk group by the IPSS classification, however, one should keep in mind that some patients with myelodysplastic syndromes with low risks by either the French-American-British (FAB) classification or the IPSS classification may have progressive disease and subsequential cytogenetic analysis could predict the disease progression. PMID- 10867136 TI - Clonal response of K562 leukemic cells to exogenous p21WAF1. AB - The p21WAF1 protein is involved in the control of cell differentiation and proliferation. We have previously shown that p21WAF1 is upregulated in normal, proliferating hematopoietic cells undergoing differentiation. Exogenous p21WAF1 has been reported to increase colony-formation by normal hematopoietic progenitors. We examined the effects of exogenous p21WAF1 on proliferation, differentiation, gene expression and colony-formation by K562 cells using an inducible p21WAF1 expression construct. Expression of the stathmin (oncoprotein 18) gene decreased within 24 h of p21WAF1 expression; Hox B4 expression increased. Four K562 subclones were derived which differed in their response to equivalent induction of p21WAF1. All four subclones exhibited growth arrest in response to p21WAF1 in liquid culture. Three of four clones developed cytoplasmic granulation and partial morphologic differentiation after p21WAF1 induction. One clone exhibited fewer morphologic features of differentiation following p21WAF1 induction and unlike other clones, colony formation in methlycellulose was not decreased by p21WAF1 expression in this clone. This indicates that additional cell-specific factors influence cellular fate in the presence of elevated p21WAF1. PMID- 10867137 TI - Disruption of the IFN-gamma cytokine network in chronic lymphocytic leukemia contributes to resistance of leukemic B cells to apoptosis. AB - Recent evidence indicates that the slowly expanding population of CD5+ B cells that characterizes chronic lymphocytic leukemia (CLL) results primarily from defects in responses to cytokines that regulate apoptosis (e.g. I1-4, TGF-beta, IFN-alpha, IFN-gamma). We have now demonstrated not only that the enhanced anti apoptotic effect of IFN-gamma on these neoplastic B cells is apparently mediated through increased levels of IFN-gamma receptors but also that there are increased numbers of IFN-gamma-expressing CD4 and CD8 T cells in these patients. This is the strongest evidence to date that multiple alterations in the IFN-gamma cytokine network contribute to the pathogenesis of CLL. PMID- 10867138 TI - In vitro chemo-sensitivity of B-chronic lymphocytic leukaemia to ara-G. AB - In vitro sensitivity to ara-G of peripheral blood B-CLL lymphocytes from 23 patients was assessed using the flow cytometric TdT assay. All patients had typical B-CLL, according to immunophenotype and morphology, and none had received treatment within 1 year of testing. A wide range of spontaneous apoptosis was recorded. Exposure of the cells to a concentration of ara-G, comparable to plasma levels achieved in a phase I trial, produced significant increases in the rate of apoptosis in 21 out of 23 patients. Prior treatment, stage or lymphocyte doubling time did not influence the effect of ara-G. Ten patients' samples tested, in parallel, for sensitivity to fludarabine demonstrated a good correlation of response to both drugs. This in vitro evidence of activity against B-CLL suggests that the spectrum of clinically useful action may be broader than previously demonstrated. PMID- 10867139 TI - Reactive plasmacytoses in multiple myeloma during hematopoietic recovery with G- or GM-CSF. AB - We report on three cases of reactive plasmacytoses (RP) in the course of multiple myeloma (MM). The three patients achieved complete remission following high dose melphalan and peripheral blood stem cell transplantation. These transient plasmacytoses had all the characteristics of RP, i.e. expansion of highly proliferative polyclonal plasma cells (PC) with a normal phenotype and genotype and corresponding to expansion of both PC progenitors (plasmablasts) and PC precursors (early plasma cells). These cells were easily distinguished from malignant PC of the corresponding patients evaluated at diagnosis, especially by their phenotypic and genotypic features. PMID- 10867140 TI - Clinical significance of reverse BCR/ABL gene rearrangement in Ph-negative chronic myelogenous leukemia. PMID- 10867141 TI - Aneuploidy precedes and segregates with chemical carcinogenesis. AB - A century ago, Boveri proposed that cancer is caused by aneuploidy, an abnormal balance of chromosomes, because aneuploidy correlates with cancer and because experimental aneuploidy generates "pathological" phenotypes. Half a century later, when cancers were found to be nonclonal for aneuploidy, but clonal for somatic gene mutations, this hypothesis was abandoned. As a result, aneuploidy is now generally viewed as a consequence, and mutated genes as a cause of cancer. However, we have recently proposed a two-stage mechanism of carcinogenesis that resolves the discrepancy between clonal mutation and nonclonal karyotypes. The proposal is as follows: in stage 1, a carcinogen "initiates" carcinogenesis by generating a preneoplastic aneuploidy; in stage 2, aneuploidy causes asymmetric mitosis because it biases balance-sensitive spindle and chromosomal proteins and alters centrosomes both numerically and structurally (in proportion to the degree of aneuploidy). Therefore, the karyotype of an initiated cell evolves autocatalytically, generating ever-new chromosome combinations, including neoplastic ones. Accordingly, the heterogeneous karyotypes of "clonal" cancers are an inevitable consequence of the karyotypic instability of aneuploid cells. The notorious long latent periods, of months to decades, from carcinogen to carcinogenesis, would reflect the low probability of evolving by chance karyotypes that compete favorably with normal cells, in principle analagous to natural evolution. Here, we have confirmed experimentally five predictions of the aneuploidy hypothesis: (1) the carcinogens dimethylbenzanthracene and cytosine arabinoside induced aneuploidy in a fraction of treated Chinese hamster embryo cells; (2) aneuploidy preceded malignant transformation; (3) transformation of carcinogen-treated cells occurred only months after carcinogen treatment, i.e., autocatalytically; (4) preneoplastic aneuploidy segregated with malignant transformation in vitro and with 14 of 14 tumors in animals; and (5) karyotypes of tumors were heterogeneous. We conclude that, with the carcinogens studied, aneuploidy precedes cancer and is necessary for carcinogenesis. PMID- 10867142 TI - Detailed genome-wide screening for inter- and intrachromosomal abnormalities by sequential G-banding and RxFISH color banding of the same metaphase cells. AB - While the now-classic chromosome banding methods, such as G-banding, remain the techniques of choice for the initial screening for karyotypic abnormalities, sometimes chromosomal rearrangements involve segments too small or too similarly banded to be detected or described adequately by these techniques. The necessity to use a genome-wide, fluorescence in situ hybridization (FISH)-based screening technique as a complement to G-banding is especially obvious in cases where the information obtained by the latter analysis does not provide an adequate guide to the choice of probes for chromosome-specific FISH. Furthermore, the same metaphase cells should ideally be used for both G-banding and FISH analysis to overcome the scarcity of metaphases observed in many cases and to ensure the correct interpretation of chromosomal aberrations in cytogenetically unstable neoplasms with massive cell-to-cell karyotypic variability. We describe a protocol which enables cross-species color banding (RxFISH), a new FISH-based screening technique that simultaneously imparts specific color banding patterns on all chromosomes, of preparations that have been G-banded and mounted for up to several years, as well as a procedure allowing chromosome-specific painting of the same metaphase cells to resolve whatever doubts persist after the preceding G banding and RxFISH analyses. This approach makes possible a detailed, genome-wide screening for inter- and intrachromosomal abnormalities including archival cases whose karyotypic rearrangements had been incompletely identified by G-banding. PMID- 10867143 TI - 19p deletion in recurring leiomyosarcoma lesions from the same patient. AB - Ten leiomyosarcomas (LMS) affecting the same patient over a period of 3 years were cytogenetically studied to detect nonrandom chromosomal changes with a pathogenetic significance. All tumors, likely metastases of a previous LMS presentation, were classified as small, including eight that developed before chemotherapy; the diagnoses were based on standard immunohistochemistry methods for smooth muscle tumors. Scoring of 613 metaphases revealed monosomy of chromosome 22 in six LMS, monosomy of chromosome 19 in three, and deletion of chromosome 19p in all ten. Interphase fluorescence in situ hybridization (FISH) of the 22-alphoid-specific probe allowed loss of the target chromosome to be detected in four tumors at higher frequencies than those detected by cytogenetics. Double-color FISH of the 19p- and 19q-specific YAC performed on one tumor made it possible to distinguish the monosomic and 19p deleted cells, the relative frequencies of which were found to be 10% and 20%, respectively. The deletion breakpoint could be mapped at 19p13 between YAC 957d12 and YAC 947g4. The recurrence of the 19p deletion in a subset of tumor cells from all of the analyzed LMS suggests that this structural aberration is a significant change in the development of leiomyosarcomas. PMID- 10867144 TI - Balanced translocation (3;7)(p25;q34): another mechanism of tumorigenesis in follicular thyroid carcinoma? AB - Alterations of 3p are the most frequently observed changes in follicular thyroid carcinomas. Loss of 3p25-pter has been speculated to be a critical event in the malignant transformation of a subset of thyroid follicular neoplasms. The present report describes a minimally invasive follicular thyroid carcinoma (FTC) with a balanced t(3;7)(p25;q34) and dic(15;22)(p11;p11) as the only abnormalities. The alterations were present in all metaphases analyzed and were demonstrated by G banding, spectral karyotyping (SKY), and fluorescence in situ hybridization (FISH). This study represents the second case of FTC where 3p25 is involved in a balanced translocation. The findings support the existence of a gene locus in this region which is involved in the tumorigenesis of thyroid carcinoma. PMID- 10867145 TI - Interstitial deletion of the short arm of chromosome 12 during clonal evolution in myelodysplastic syndrome with t(5;12)(q13;p13) involving the ETV6 gene. AB - We report here a 65-year-old man with a myelodysplastic syndrome (MDS), refractory anemia with excess of blasts. He had received chemotherapy with tegafur for renal carcinoma. Chromosome analysis of bone marrow cells revealed complex karyotypes; del(5)(q13) was observed in all 20 metaphase spreads, and two related aberrations, add(12)(p11) and add(12)(p13), were detected in 13 and 7 cells, respectively. Fluorescence in situ hybridization (FISH) analysis with chromosome-specific DNAs revealed that these alterations originated from a reciprocal translocation (5;12)(q13;p13). Therefore, del(5)(q13), add(12)(p11), and add(12)(p13) were revised as der(5)t(5;12)(q13;p13), der(12)del(12)(p11p13)t(5;12)(q13;p13), and der(12)t(5;12)(q13;p13), respectively. Fluorescence in situ hybridization with a series of cosmid probes spanning the ETV6 gene showed that the 12p13 breakpoint on the der(12)t(5;12)(q13;p13) was located in intron 1, but the exon 1 signal was deleted. Our results suggest that a fusion gene was generated between the 5'-end of an unidentified partner at 5q13 and the 3'-end of ETV6 by t(5;12)(q13;p13), and that the interstitial deletion (12)(p11p13) occurred following t(5;12) during clonal evolution. del(12)(p11p13), including the rearranged ETV6 gene, may be implicated in the progression of MDS. PMID- 10867146 TI - Characterization of chromosomal breakpoints in an ALL patient using cross-species color banding. AB - Cross-species color-banded karyotype (Rx-FISH) results were compared with those of conventional G-banded metaphases from the same sample. Breakpoints and karyotype were confirmed as 46,XX,t(8;22)(q24;q11), der(9)t(1;9)(q21;p13) through the novel technology of cross-species color banding in an acute leukemic patient (ALL, L3); the karyotype was 46,XX,t(8;22)(q24;q11),der(9)t(1;9)(q25;p24) by conventional G-banding. PMID- 10867147 TI - Late-appearing PML/RARalpha fusion transcript with coincidental t(12;13)(p13.2;q14) in acute promyelocytic leukemia lacking the t(15;17) cytogenetic anomaly. AB - The late appearance of a cytogenetic/molecular hallmark in human leukemias is a rare event. We report on a case of acute myeloid leukemia with morphology, immunophenotype and clinical features typical of promyelocytic subtype (APL), in which the specific PML/RARalpha gene rearrangement was molecularly detected only at second relapse of disease, without cytogenetic evidence of the t(15;17). The emergence of the PML/RARalpha gene may be therapy-related or may represent the exceptional result of a clonal evolution during progression of neoplasia. At second relapse, a novel cell clone bearing a t(12;13)(p13.2;q14) was also observed and a molecular deletion and rearrangement of a locus at 13q14, distinct from retinoblastoma (Rb1) locus, was found. In this unusual case, the PML/RARalpha product seems to be not essential for the expression of the promyelocytic phenotype at diagnosis and, when detectable, it is not the sole genetic defect. PMID- 10867148 TI - Sclerosing epithelioid fibrosarcoma: a cytogenetic, immunohistochemical, and ultrastructural study of an unusual histological variant. AB - A case of sclerosing epithelioid fibrosarcoma was studied. The tumor cells expressed vimentin, focally epithelial membrane antigen and CD34, contained cisternae of rough endoplasmic reticulum, large Golgi apparatus, many pinocytotic vesicles, and were devoid of basal lamina. Their composite karyotype was 45,Y,t(X;6)(q13;q15), t(6;13)(p11.2;q13),-22?2/46,Y,t(X;6)(q13;q15),add(13)(p12), add(22)(q13)?3/44 approximately 46,der(X)t(X;6)(q13;q21),-Y, t(13;14)(q10;q10), 22,add(22)(q13)?7/46,XY?8. PMID- 10867149 TI - Comparative genomic hybridization reveals complex genetic changes in primary breast cancer tumors and their cell lines. AB - DNA copy number changes were characterized by comparative genomic hybridization (CGH) in 18 breast cancer cell lines. In 5 of these, the results were comparable with those from the primary tumors of which the cell lines were established. All of the cell lines showed extensive DNA copy number changes, with a mean of 16.3 +/- 1.1 aberrations per sample (range 7-26). All of the cell lines had a gain at 8q22-qter. Other common gains of DNA sequences occurred at 1q31-32 (89%), 20q12 q13.2 (83%), 8q13 (72%), 3q26.1-qter (67%), 17q21-qter (67%) 5p14 (61%), 6p22 (56%), and 22pter-qter (50%). High-level amplifications were observed in all cell lines; the most frequent minimal common regions were 8q24.1 (89%), 20q12 (61%), 1q41 (39%), and 20p11.2 (28%). Losses were observed less frequently than gains and the minimal common regions of the most frequent losses were Xq11-q12 (56%), Xp11.2-pter (50%), 13q21 (50%), 8p12-pter (44%), 4p13-p14 (39%), 6q15-q22 (39%), and 18q11.2-qter (33%). Although the cell lines showed more DNA copy number changes than the primary tumors, all aberrations, except one found in a primary tumor, were always present in the corresponding cell line. High-level amplifications found both in primary tumors and cell lines were at 1q, 8q, 17q, and 20q. The DNA copy number changes detected in these cell lines can be valuable in investigation of tumor progression in vitro and for a more detailed mapping and isolation of genes implicated in breast cancer. PMID- 10867150 TI - The expression frequency of common fragile sites and genetic predisposition to colon cancer. AB - The expression frequency of common fragile sites induced by aphidicolin (Apc), bromodeoxyuridine (BrdU), and caffeine was evaluated on prometaphase chromosomes obtained from the peripheral blood lymphocytes of 32 patients with colon cancer, 30 of their clinically healthy family members and 30 age-matched normal controls. The proportion of damaged cells (P < 0.001), the mean number of chromosomal aberrations and the expression frequencies of fragile sites were significantly higher in the patient and relative groups compared to the control group. Our findings show an increased genetic instability in patients with colon cancer and their first-degree relatives. In addition, common fragile sites can be used as a suitable marker for determining genetic predisposition to cancer. PMID- 10867151 TI - TP53 deletions but not trisomy 12 are adverse in B-cell lymphoproliferative disorders. AB - Abnormalities of the TP53 tumor suppressor gene at 17p13.1 are prognostically adverse in a variety of hematolymphoid malignancies. The present study utilized interphase fluorescence in situ hybridization (I-FISH) to detect TP53 deletions and trisomy 12 in 101 clinical specimens from 98 patients with B-cell lymphoproliferative disorders (B-LPDs). Twelve patients had TP53 deletions (group A), 23 had trisomy 12 (group B), and 63 had neither (group C). The groups did not significantly differ in age, duration of disease, absolute lymphocyte count, or percentage with an immunophenotype or cytology atypical for chronic lymphocytic leukemia (CLL). The clinical stage of disease and lactate dehydrogenase (LDH) level were higher in group A, with less response to therapy. After a median follow-up of 19 months, seven of the patients in group A had died of disease (another patient subsequently has had large cell transformation) compared with none in group B and nine in group C. Multivariate analysis found the stage of disease and TP53 deletions as the only parameters independently associated with shortened survival (P < 0.001). Thirty-nine patients had conventional cytogenetic analysis (CCA) which was complexly abnormal in 11 patients; 6 of whom died of disease. There was a trend for complex cytogenetics to be seen more frequently in group A, often with 17p involvement. For most laboratories, CCA may be the preferable initial study to identify prognostically different subgroups of B LPDs. However, as more probes and clinical outcome data become available, I-FISH will likely play an increasingly important ancillary role. PMID- 10867152 TI - Cytogenetic findings in a case of epithelioid sarcoma and a review of the literature. AB - Cytogenetic studies of epithelioid sarcoma, a rare malignant soft tissue neoplasm of adolescents and young adults, are few. A characteristic anomaly has not yet been identified for this sarcoma. In this study, cytogenetic studies of a primary epithelioid sarcoma of a 15-year-old male revealed the following abnormalities: t(6;8)(p25;q11.2) and add(7)(p15). PMID- 10867153 TI - Polysomy 13 with concomitant deletion of 13q13-14 involving the retinoblastoma gene and the D13S25 locus in a case of acute myeloid leukemia. AB - We herein describe a case of acute myeloblastic leukemia (AML), FAB subtype M4, with an unfavorable clinical course and a complex karyotype, including 4-9 copies of chromosome 13. Polysomy 13 was a result of clonal evolution. Fluorescence in situ hybridization (FISH) revealed a cytogenetically unrecognizable deletion within 13q13-14 that included the retinoblastoma gene (RB) and the D13S25 locus in all but one copy of chromosome 13. The only chromosome 13 that did not show a deletion affecting the q13-14 region was translocated to chromosome 7, resulting in a dic(7;13)(q21;p11). In this case, the coexistence of polysomy and a partial deletion within the same chromosome point toward a possible formation of a fusion product with oncogenic potential and its consecutive amplification as a critical alteration in this case. PMID- 10867154 TI - Submicroscopic insertion of RARalpha gene into chromosome 15 in two cases of acute promyelocytic leukemia. AB - Acute promyelocytic leukemia (APL) is characterized by a specific translocation (15;17)(q22;q21), resulting in the formation of PML/RARalpha chimeric transcripts. We report two female patients with PML/RARalpha-positive classical APL, whose leukemic cells expressed a variant translocation, t(5;15)(q13;q22) and t(15;17)(q22;p13), respectively. Both translocations were confirmed by whole chromosome painting which revealed no apparent involvement of 17q. A two-color fluorescence in situ hybridization with a 5' PML and a 3' RARalpha probe showed, in both cases, the presence of a PML-RARalpha fusion gene, on the der(15)t(5;15) long arm, and on the der(17)t(15;17) short arm, respectively. These two complex rearrangements resulted most probably from a two-step mechanism: (1) a submicroscopic insertion into 15q of a 17q segment including the 3' part of the RARalpha gene; (2) a reciprocal translocation between der(15) and a variable chromosome arm, with a breakpoint distal and proximal to RARalpha insertion in the case of t(5;15) and t(15;17), respectively. Molecular and prognosis significance of these variant translocations are discussed. PMID- 10867155 TI - Effect of two different combinations of antiretrovirals (AZT+ddI and AZT+3TC) on cytokine production and apoptosis in asymptomatic HIV infection. AB - Nineteen HIV-seropositive antiretroviral therapy-naive and asymptomatic individuals (200-500 CD4/microl) were enrolled in a prospective study aimed at analyzing the immunologic and virologic effects of two different combinations of nucleoside reverse transcriptase inhibitors (AZT+ddI and AZT+3TC), and randomly assigned to one of the treatment group. Immunologic (CD4 and CD8 counts, mitogen stimulated cytokine production, unstimulated and mitogen-stimulated apoptosis) and virologic (HIV viral load) determinations were performed pre-therapy and 15, 30, 90, 200 and 360 days after initiation of therapy. Results showed that the two combinations had comparable effects on increasing CD4 counts and the CD4/CD8 ratio and in reducing HIV viral load. In contrast, AZT+3TC was more efficient in improving interleukin-2 (IL-2) and interferon gamma (IFNgamma) production as well as the type 1/type 2 cytokine ratio and in down modulating the susceptibility of peripheral blood mononuclear cells to in vitro mitogen-stimulated apoptotic cell death. These data suggest that the combination of AZT+3TC has a stronger effect on potentially beneficial immune parameters (IL-2 production; reduction of apoptosis) than the one between AZT+ddI. The combination of AZT+3TC could be more advantageous in the therapy of HIV infection even when used in association with a protease inhibitor. PMID- 10867156 TI - A novel high throughput screening assay for HCV NS3 helicase activity. AB - A novel assay for measurement of Hepatitis C virus (HCV) NS3 helicase activity was developed using Flashplate technology. This assay involves the use of a DNA duplex substrate and recombinant HCV NS3 produced in Escherichia coli. The DNA duplex consisted of a pair of oligonucleotides, one biotinylated, the other radiolabeled at their respective 5' termini. This DNA duplex was immobilized, via the biotin molecule, on the surface of a neutravidin-coated SMP103 Flashplate (NEN Life Science Products). Helicase activity results in the release of the radiolabeled oligonucleotide, which translates in signal reduction with respect to control wells. Biochemical characterization of the HCV NS3 helicase activity was performed using this assay. We demonstrated that the NS3-mediated unwinding is proportional to both the amount of DNA substrate in the well, and to the NS3 concentration in the reaction. Most of the NS3-mediated unwinding was achieved in the initial 60 min of incubation. As expected the reactions were ATP-dependent and found to be affected by the concentration of MgCl(2), MnCl(2), KCl, EDTA, and by pH. We found this assay to be highly reproducible since only slight variation was observed when a total of 68 helicase reactions were performed on one plate. Therefore, this Flashplate helicase assay is fast, convenient and reproducible. These criteria make it suitable for high throughput screening of potential NS3 helicase inhibitors. PMID- 10867157 TI - The inhibitory effect of the imidazoquinolinamine S-28828 on the pathogenesis of a type II adenovirus in turkeys. AB - In this study we show that a type I-IFN inducing compound, S-28828, modulated the pathogenesis of an avian type II adenovirus in turkeys. By itself, S-28828 induced a strong reaction in the spleen characterized by hyperplasia of the red and white pulps as well as an increase in lymphoid cell aggregations. Oral administration of S-28828 before the time of virus inoculation suppressed significantly (P<0.05) the replication of hemorrhagic enteritis virus (HEV) in turkeys. Two doses of 5 or 50 mg of S-28828 administered at 2 days before and at the day of virus inoculation inhibited HEV-induced pathological and histopathological lesions. Virus-induced apoptosis and reduced IgM-surface expression of B cells were suppressed by low dose S-28828 treatment. These results are of interest because mammalian adenoviruses were shown to be resistant to antiviral effects of type I IFN, the major effector cytokine induced by S 28828. PMID- 10867158 TI - Loss of interferon alpha and interferon tau-induced antiviral protection in interferon regulatory factor-2 DNA-binding domain dominant negative mutants. AB - The role of the interferon regulatory factory (IRF) family of transcription factors in regulation of interferon alpha and interferon tau antiviral activity was investigated using a dominant negative mutant of IRF-2. The IRF-2 DNA binding domain (DBD), without the C-terminal regulatory region, was stably transfected into myeloid U937 cells. Expression of the IRF-2 DBD resulted in an increase in constitutive 2'5' oligoadenylate synthetase (OAS) levels, indicative of an active repressive mechanism, but was not sufficient to protect cells from challenge with vesicular stomatitis virus. Treatment of the DBD clones with interferons alpha A and tau failed to upregulate 2'5' OAS expression and did not elicit an antiviral response. While interferon alpha A was more sensitive than interferon tau to the inhibitory effects of the IRF-2 DBD, IRF-mediated gene induction is involved in successful interferon alpha and tau-induced anti-VSV activity. PMID- 10867159 TI - In vitro evaluation of the effect of temporary removal of HIV drug pressure. AB - We tried to establish whether MT-4 cells that were infected with HIV-1(HTLV III(B)) at a high multiplicity of infection (m.o.i.=1), and subsequently treated with high concentrations of anti-HIV drugs for several days, would be able to resume virus production after the antivirals are washed away. The HIV inhibitors studied were the nucleoside reverse transcriptase inhibitors (NRTIs) zidovudine and lamivudine, the non-nucleoside reverse transcriptase inhibitors (NNRTIs) nevirapine, delavirdine and loviride, the acyclic nucleoside phosphonate RT inhibitor (R)-9-(2-phosphonylmethoxypropyl)adenine (tenofovir) and the protease inhibitors (PIs) saquinavir, indinavir and ritonavir. The compounds, at 50 and 500 times their 50% inhibitory concentration (IC(50), determined at a m.o.i.=0.01), were added immediately after virus adsorption and removed after an incubation period of 0 (wash control), 24, 48 or 72 h. Virus breakthrough was monitored by microscopical examination of cytopathicity, viral infectivity (yield) and p24 levels in the supernatant. The presence of HIV-1(HTLV-III(B)) proviral DNA was determined after a 72-h incubation period. None of the antiviral drugs studied was able to prevent resumption of viral growth after removal of the compound. Tenofovir, lamivudine and the NNRTIs nevirapine, delavirdine and loviride, at 500 times their respective IC(50), were able to delay viral breakthrough for approximately 2-3 days. The NRTI zidovudine and the PIs saquinavir, indinavir and ritonavir, under the same conditions, were not able to delay viral breakthrough at all. Virus recovered upon treatment proved as sensitive to the anti-HIV drugs as wild-type virus. Our results suggest that viral replication at the cellular level was not completely inhibited by drug monotherapy. Consequently, virus rebounded when drug therapy stopped. In conclusion, our findings suggest that drug holidays would result in viral breakthrough, even after virus replication has been previously suppressed by adequate drug levels. PMID- 10867160 TI - Combinations of reverse transcriptase, protease, and integrase inhibitors can be synergistic in vitro against drug-sensitive and RT inhibitor-resistant molecular clones of HIV-1. AB - Combinations of anti-HIV agents including one or two reverse transcriptase inhibitors with a protease inhibitor are potent and effective. However, toxicities, costs and the emergence of drug-resistant organisms have compromised their long-term efficacy in people. A next, likely, target for anti-HIV therapy is HIV-1 integrase. Viral integration, catalyzed by integrase, is absolutely required for HIV replication. L-chicoric acid is a potent and selective inhibitor of HIV-1 integrase that also inhibits HIV-1 replication in cell culture. As a first step in understanding the potential role for integrase inhibitors in clinical medicine, the activities of L-chicoric acid alone and in combination with 2', 3'-dideoxycytidine, zidovudine, and a protease inhibitor, nelfinavir, were tested in vitro against molecular clones of HIV-1 resistant to reverse transcriptase inhibitors. L-chicoric acid was equally effective against a wild type clone of HIV-1, HIV(NL4-3), or against HIV-1 resistant to either zidovudine or dideoxycytidine. L-chicoric acid was largely synergistic with zidovudine and synergistic with both dideoxycytidine and nelfinavir. PMID- 10867161 TI - Exploration of subjective well-being and dependence in daily activities at the beginning of the geriatric rehabilitation process: a challenge to traditional goal-setting and evaluation procedures? AB - The objectives of this study were to describe a population of elderly patients at the beginning of their rehabilitation period as regards subjective well-being and dependence in activities of daily living (ADL). In a Swedish rural county 244 patients aged 65+ who had begun rehabilitation within the last month were targeted. One part of the self-administered Goteborg Quality of Life Instrument and a revised version of the ADL Staircase were used. No correlation was found between subjective well-being and ADL dependence. However, significant correlations between ADL dependence and separate subjective well-being items were found in three out of 17, i.e. the items 'energy', 'leisure', and 'sense of significance and appreciation outside home'. Overall subjective well-being did not show any gender differences, but significant gender differences due to the distribution of scores was shown; females scored the items 'health', 'sleeping', and 'economy' as bad to a larger extent than males. Males were significantly more dependent than females in three out of nine ADL: 'going to the toilet', 'dressing', and 'cooking'. Additional knowledge of subjective well-being and ADL dependence at the beginning of the rehabilitation process challenges the traditional goal-setting and evaluation procedures of geriatric rehabilitation services. PMID- 10867162 TI - Post myocardial infarction use of beta-blockers. AB - The use of beta-blockers after myocardial infarction was investigated in 100 patients (age range 30-89 years), from January to December 1997, in Caerphilly District Miner's Hospital. Out of 54 patients eligible for beta-blockers only 27 (27/54 (50%)) patients received such therapy. We conclude that beta-blockers is underused in patients with myocardial infarction. Although the reason for this is not clear from this study, this probably indicates the anxieties clinicians may have about the side effects from the use of beta-blockers in this population. PMID- 10867163 TI - Influence of age on clinical presentation of acute pulmonary embolism. AB - The aims of this study were to compare the clinical features of patients with pulmonary embolism (PE) and patients in whom the initial suspected diagnosis was not confirmed by the complementary studies and to determine the possible clinical differences among patients with PE according to age. A retrospective review of the charts of a group of patients with PE (n, 96) and another without PE (n, 96) was carried out. The patients with PE over 65 years of age (n, 64) were compared with those under 66 years of age (n, 32). The variables related to PE were absence of known heart disease, duration of symptoms /=0.4. Conversions of both proteins in the oligomers are roughly the same. The estimates of the oligomerisation yield obtained by the gel-permeation chromatography and SDS gel electrophoresis agree well. This indicates that the formation of intermolecular disulfide bonds is necessary for the oligomerisation. Thus, the oligomerisation of pressure denatured ALA and BLG is driven by the thiol<-->disulfide exchange rendered possible by the vigorous baric denaturation and the exposure of the free thiol group of BLG, which acts as an initiator of disulfide bridges scrambling. PMID- 10867182 TI - How to develop globular proteins into adhesives. AB - To make globular proteins suitable for application in adhesives, the specific bonds and interactions which shape their structure have to broken. Only then, a layer of relatively large, flexible and interwoven polymer chains, which are firmly attached to the solid surface by adsorption, can be created. Such a network layer is essential to save the adhesive bond under an applied force, because it can distribute the concentration of stresses generated at the interface into the bulk. Unfolding and swelling of a protein can be achieved by changing the solvent quality. For the globular whey protein beta-lactoglobulin, the optimal conditions for unfolding and swelling is found with 98% formic acid as a solvent. In formic acid, beta-lactoglobulin looses its amphoteric character (it is protonated, probably for approximately 20%). In addition, formic acid is less polar than water and thus a better solvent for the apolar parts of the protein. The swelling and unfolding behaviour of beta-lactoglobulin is studied by viscosity and CD-spectroscopy measurements. For the interpretation of the results we apply the Kuhn formalism that the conformation of a protein can be described in terms of a statistical chain which consists of segments of an average persistence length P. The statistical segment length P, which varies with the experimental conditions, is directly related to the adsorption energy required for a strong adhesion between coil and surface. It determines the depletion energy kT P(-2) m(-2) which must be overcome by specific attraction between side groups of the protein chain and the surface. For beta-lactoglobulin in 98% formic acid, we find a P value of approximately 2.2 nm, pointing at a relatively flexible chain. The minimum net adsorption energy kT P(-2) is then approximately 1 mJ m(-2), a relatively small value to be exceeded. Preliminary results of destructive adhesion tests on beech wood lap-shear joints reveal promising tensile strengths of approximately 2.9+/-1.1 N mm(-2), indeed. PMID- 10867183 TI - The effect of pH on heat denaturation and gel forming properties of soy proteins. AB - This study is focussed on the influence of pH on the gel forming properties of soy protein isolate and purified glycinin in relation to denaturation and aggregation. At pH 7.6 more fine-stranded gels were formed characterised by low G' values, and a smooth, slightly turbid appearance, whereas at pH 3.8 coarse gels were obtained with a high stiffness and a granulated, white appearance. Low G' values, as found at pH 7.6, correlate with a high solubility of glycinin and soy protein isolate (ca. 50%) after heating at low protein concentration. At pH 3.8 all protein precipitated upon heating, which correlates with relatively high G' values. The role of beta-conglycinin during gelation of SPI seems to be minor at pH 7.6, which is indicated by the fact that, in contrast to pH 3.8, notable gel formation did not start upon heat denaturation of beta-conglycinin. Furthermore, the mechanism of gel formation seems to be affected by pH, because at pH 7.6, in contrast to pH 3.8, the disulphide bridge between the acidic and the basic polypeptide of glycinin is broken upon heating. PMID- 10867184 TI - Gelation by phase separation in a whey protein system: in-situ kinetics of aggregation. AB - The aggregation and gelation properties of beta-lactoglobulin (BLG), a globular protein from milk, was studied in aqueous ethanol solutions at room temperature. The phase state diagrams as a function of pH and ethanol concentration showed that a gel structure appeared after a period ranging from 1 min to 1 week, depending on the physico-chemical conditions. The in-situ kinetics of aggregation were followed by several methods in order to obtain a better understanding of the building of aggregates by the addition of ethanol. It was shown that the aggregation kinetics highly depended upon the pH, the process being fastest at pH 7. Viscoelasticity and infrared measurements indicated that alcohol-induced gelation would proceed via a two-step mechanism: small aggregates loosely connected between them were first built up; a real network took place in a second step. The coarse and irregular structures formed in aqueous ethanol gels revealed by confocal laser scanning microscopy could be analysed in terms of a phase separation. This observation was supported by a syneresis phenomenon visible in the final gel state. BLG in water-ethanol solution would undergo either an inhibition of the demixing by gelation or a binary phase separation accompanied by an irreversible gelation transition. PMID- 10867185 TI - Isinglass/collagen: denaturation and functionality. AB - Isinglass is widely used commercially to clarify alcoholic beverages by aggregation of the yeast and other insoluble particles. It is derived from swim bladders of tropical fish by solubilisation in organic acids and consists predominantly of the protein collagen. The low content of intermolecular cross links allows ready dissolution of swim bladder compared to bovine hide which is cross-linked by a high proportion of stable bonds and requires enzymic digestion to solubilise. Isinglass is no longer effective as a clarifying agent if thermally denatured hence the collagenous triple helical structure must be maintained. Thermal denaturation of isinglass occurs at 29 degrees C, compared to 40-41 degrees C for mammalian collagens, primarily due to the lower hydroxyproline content. The hydroxyproline is essential for the formation of H bonded water-bridges through the hydroxyl group and the peptide chain thereby stabilising the triple helix. Based on the lower enthalpy determined by differential scanning calorimetry we have calculated that the thermally labile domain of the isinglass molecule was 41 residues compared to 66 for mammalian collagen. The fining efficiency was unaffected by pH, chelating agents, detergents and removal of surface proteins from yeast cells. Studies on the mechanism of action of isinglass have shown that higher molecular weight aggregates that increase the length of the collagen molecules (trimers, tetramers, etc.) increase efficiency and that their surface charge are important in the clarification process. By chemical modification, we have shown that blocking positively charged groups had no effect on the fining process, whilst negative charges are clearly essential and that increasing the negative charge by succinylation increases its efficacy. Solutions of bovine hide collagen were shown to be equally effective in refining beers and standard yeast preparations. The higher thermal denaturation temperature, ready availability and reproducibility of bovine collagen preparations gives it considerable advantages over isinglass. PMID- 10867186 TI - BSA structural changes during homomolecular exchange between the adsorbed and the dissolved states. AB - The secondary structure and the thermostability of bovine serum albumin (BSA), before adsorption and after homomolecular displacement from silica and polystyrene particles, are studied by circular dichroism spectroscopy and differential scanning calorimetry. The structural perturbations induced by the hydrophilic silica surface are reversible, i.e. BSA completely regains the native structure and stability after being exchanged. On the other hand, the adsorption on, and subsequent desorption from, polystyrene particles causes irreversible changes in the stability and (secondary) structure of BSA. The exchanged proteins have a higher denaturation temperature and a lower enthalpy of denaturation than native BSA. The alpha-helix content is reduced while the beta-turn fraction is increased in the exchanged molecules. Both effects are more pronounced when the protein is displaced from less crowded sorbent surfaces. The irreversible surface induced conformational change may be related to some aggregation of BSA molecules after being exposed to a hydrophobic surface. PMID- 10867187 TI - Interpretation of DSC data on protein denaturation complicated by kinetic and irreversible effects. AB - Thermal denaturation of Kunitz soybean trypsin inhibitor (KTI) and ribulose-1,5 biphosphate carboxylase (RBPC) from tobacco leafs was studied by the method of high-sensitivity differential scanning calorimetry (HS-DSC). The dependence of the denaturation temperature on the heating rate reveals in the case of both proteins a non-equilibrium character of the denaturation transition in applied conditions. Developed kinetic approach allows the determination of an equilibrium transition temperature as well as the rate constants of denaturation and renaturation from the complex data of HS-DSC. This method was applied to the analysis of the pH-induced change of the conformational stability of KTI within pH range from 2.0 to 11.0. It allowed the determination of the pH dependencies: of the excess free energy of denaturation, of the activation enthalpy and entropy of denaturation as well as of the denaturation rate constant. Conclusions have been made suggesting the contribution of side-chain hydrogen bonds in the stabilisation of the native and activated states of KTI. PMID- 10867188 TI - Protein folding and stability investigated by fluorescence, circular dichroism (CD), and nuclear magnetic resonance (NMR) spectroscopy: the flavodoxin story. AB - In this review, the experimental results obtained on the folding and stability of Azotobacter vinelandii flavodoxin are summarised. By doing so, three main spectroscopic techniques used to investigate protein folding and stability are briefly introduced. These techniques are: circular dichroism (CD) spectroscopy, fluorescence emission spectroscopy, and nuclear magnetic resonance (NMR) spectroscopy in combination with the hydrogen exchange methodology. Results on the denaturant-induced and thermal equilibrium unfolding of apoflavodoxin from A. vinelandii, i.e. flavodoxin in the absence of the riboflavin-5'-monophosphate (FMN) cofactor, are discussed. A scheme for the equilibrium unfolding of apoflavodoxin is presented which involves a relatively stable molten globule-like intermediate. Denaturant-induced apoflavodoxin (un)folding as followed at the residue-level by NMR shows that the transition of native A. vinelandii apoflavodoxin to its molten globule state is highly co-operative. However, the unfolding of the molten globule to the unfolded state of the protein is non-co operative. A comparison of the folding of A. vinelandii flavodoxin with the folding of flavodoxin from Anaboena PCC 7119 is made. The local stabilities of apo- and holoflavodoxin from A. vinelandii as measured by NMR spectroscopy are compared. Both Che Y and cutinase, which have no sequence homology with apoflavodoxin but which share the flavodoxin-like topology, have stabilisation centres different from that of apoflavodoxin from A. vinelandii. The stable centres of structurally similar proteins can thus reside in different parts of the same protein topology. Insight in the variations in (local) unfolding processes of structurally similar proteins can be used to stabilise proteins with a flavodoxin-like fold. Finally, the importance of some recent experimental and theoretical developments for the study of flavodoxin folding is briefly discussed. PMID- 10867189 TI - Denaturation of proteins from a non-food perspective. AB - Controlled denaturation, or the prevention of denaturation, is an important aspect in the development of food applications of proteins. For the use of proteins in non-food applications such as surfactants, adhesives, coatings or plastics, it is discussed that a certain degree of denaturation must occur in order to make proteins processable, and to reach the required product properties such as strength, water resistance or adhesion. By adjusting the processing parameters (temperature, water content, chemicals) conditions can be created to allow structural changes in the protein. The effect of processing on some product properties will be discussed. PMID- 10867190 TI - Immunoprecipitation of a 50-kDa protein: a candidate receptor component for tomato spotted wilt tospovirus (Bunyaviridae) in its main vector, Frankliniella occidentalis. AB - A 50-kDa protein that binds to viral particles in solid-phase assays and that is recognized by anti-idiotypic antibodies made against anti-viral glycoproteins G1/G2 (anti-Ids) has been proposed as a receptor candidate for tomato spotted wilt tospovirus (TSWV) in its main thrips vector, Frankliniella occidentalis Pergande (Bandla et al., 1998. Phytopathology 88, 98-104). Here we show the immunoprecipitation of the 50-kDa protein by anti-Ids and by an anti-G1/G2-TSWV conjugate - a new immunoprecipitation method. In addition, we show that anti-Ids made against anti-G1 (anti-IdG1) block virus replication in an insect tissue replication assay. The results indicate that (a) the TSWV-50-kDa protein interaction occurs in solution, as it must do in vivo; (b) G1 is a viral attachment protein; and (c) the 50-kDa protein is a candidate host factor essential for TSWV entry. These results provide additional support for the role of the 50-kDa thrips protein as a viral receptor. Additionally these experiments provide the basis for testing saturable binding and represent an important step toward the first cloning and identification of a cellular receptor for a plant virus. PMID- 10867191 TI - Selection of a specific peptide from a nona-peptide library for in vitro inhibition of grass carp hemorrhage virus replication. AB - Grass carp hemorrhage virus (GCHV), a member of Reoviridae, causes severe hemorrhagic disease of grass carp (Ctenopharyngodon idellus) in China. Icosahedral virions of GCHV were used as to assay the effect of specific peptides for the inhibition of GCHV infectivity. A random nona-peptide library displayed on phage fUSE5 was constructed, and the expressed peptides were fused onto the amino terminus of the minor coat protein III. By biopanning, the fused peptides were bound to the biotinylated GCHV. Phages containing specific peptides bound to GCHV were eluted and amplified in Escherichia coli K91. Three rounds of affinity selection enriched the pool of inhibiting peptides. Sixteen clones which inhibited the replication of GCHV in a grass carp kidney cell line were selected. The TCID(50) of GCHV was decreased over 10000x. Six clones having the strongest inhibitory effect shared the same DNA sequence, with a deduced amino acid sequence of NH(2)-Leu-Trp-Val-Gly-Gly-Gly-Arg-Asn-Ala-COOH. A synthesized nona peptide of identical sequence exhibited similar inhibitory activity towards GCHV replication in vitro. PMID- 10867192 TI - A new PCR-based assay amplifies the E6-E7 genes of most mucosal human papillomaviruses (HPV). AB - We established a new assay to detect the E6-E7 DNA of mucosal human papillomaviruses (HPV) by a PCR-based method using four pairs of degenerate LCR and E7 primers (LCR-E7 PCR). This assay amplifies the full length of E6 and the N terminal part of E7. HPV typing was performed using restriction-fragment-length polymorphism (RFLP), and by analyzing the sequences of cloned PCR products. We compared this assay with the first generation hybrid captured assay (HCA-I) and the MY09/11-PCR method. LCR-E7 PCR was able to detect more than 34 mucosal HPV types and theoretically should detect two additional types. LCR-157 PCR and HCA-I detected HPV DNA in 70% (69/99) and 55% (54/99) of low-grade cervical intraepithelial lesions (LSIL), 89% (105/118) and 76% (90/118) of high-grade cervical intraepithelial lesions (HSIL), and 90% (56/62) and 79% (49/62) of invasive squamous cell carcinomas (SCC), respectively. LCR-E7 PCR was more sensitive than the HCA-1 test. Discordant results between the LCR-E7 and MY 11/09 PCR tests were observed in one of 185 (0.5%) normal samples, seven of 85 (8.2%) LSIL samples, seven of 82 (8.5%) HSIL samples, and four of 72 (5.6%) SCC samples. The discordant results were mostly observed in samples with a low-copy number of the HPV genome or with multiple HPV infection. The sensitivity of LCR-E7 PCR was equivalent to that of MY 11/09 ECR, and false positives were less frequent in LCR E7 PCR. LCR-E7 PCR may be useful for determining the biological activity of detected HPV types, since this method amplifies the entire E6 gene. PMID- 10867193 TI - Phylogenetic relationships of bluetongue viruses based on gene S7. AB - Previous phylogenetic analyses based on bluetongue virus (BTV) gene segment L3, which encodes the inner core protein, VP3, indicated a geographical distribution of different genotypes. The inner core protein, VP7, of BTV has been identified as a viral attachment protein for insect cell infection. Because the inner core proteins are involved with infectivity of insect cells, we hypothesized that certain VP7 protein sequences are preferred by the insect vector species present in specific geographic locations. We compared the gene segment S7, which encodes VP7, from 39 strains of BTV isolated from Central America, the Caribbean Basin, the United States, South Africa and Australia. For comparison, the S7 sequences from strains of the related orbiviruses, epizootic hemorrhagic disease virus (EHDV) and African horse sickness virus (AHSV) were included. The S7 gene was highly conserved among BTV strains and fairly conserved among the other orbiviruses examined. VP7 sequence alignment suggests that the BTV receptor binding site in the insect is also conserved. Phylogenetic analyses revealed that the BTV S7 nucleotide sequences do not unequivocally display geographic distribution. The BTV strains can be separated into five clades based on the deduced VP7 amino acid sequence alignment and phylogeny but evidence for preferential selection by available gnat species for a particular VP7 clade is inconclusive. Differences between clades indicate allowable variation of the VP7 binding protein. PMID- 10867194 TI - Isolation of lactate dehydrogenase-elevating viruses from wild house mice and their biological and molecular characterization. AB - Lactate dehydrogenase-elevating virus (LDV) was first identified as a contaminant of transplantable mouse tumors that were passaged in laboratory mice. It has been assumed that these LDVs originated from LDVs endemic in wild house mouse populations. In order to test this hypothesis and to explore the relationships between LDVs from wild house mice among each other and to those isolated from laboratory mice, we have isolated LDVs from wild house mice and determined their biological and molecular properties. We have screened for LDV tissues of 243 wild house mice that had been caught in various regions of North, Central and South America between 1985 and 1994. We were able to isolate LDVs from the tissues of four mice, three had been caught in Baltimore, MD and one in Montana. We demonstrate that the phenotypic properties (ability to establish a long-term viremic infection, low immunogenicity of the neutralization epitope, high resistance to antibody neutralization and lack of neuropathogenicity) of the four wild house mouse LDVs are identical to those of the primary LDVs isolated from transplantable tumors (LDV-P and LDV-vx), which are distinct from those of the neuropathogenic LDV-C. Furthermore, ORF 5 and ORF 2 and their protein products (the primary envelope glycoprotein VP-3P, and the minor envelope glycoprotein, respectively) of the wild house mouse LDVs were found to be closely related to those of LDV-P and LDV-vx. The LDVs caught in Baltimore, MD were especially closely related to each other, whereas the LDV isolated in Montana was more distantly related, indicating that it had evolved independently. The ectodomain of VP-3P of all four wild house mouse LDVs, like those of LDV-P and LDV-vx, possess the same three polylactosaminoglycan chains, two of which are lacking in the VP-3P ectodomain of LDV-C. These results further strengthen the conclusion that the three polylactosaminoglycan chains are the primary determinants of the phenotypic properties of LDV-P/vx. PMID- 10867195 TI - Rearrangement generated in double genes, NSP1 and NSP3, of viable progenies from a human rotavirus strain. AB - We generated rotavirus clones with rearrangement in vitro by serial passages of a human rotavirus strain (IGV-80-3) at high multiplicity of infection and determined nucleotide sequences of the rearranged genes from two distinct rotavirus clones, each of which possesses two rearranged genes: a common rearranged NSP1 gene and NSP3 gene with slightly different migration in polyacrylamide gel electrophoresis. Sequence analysis showed that the rearranged NSP1 and NSP3 genes had similar gene structures: concatemerization in a head to tail orientation and partial duplication of the open reading frame following the termination codon. The rearranged NSP1 gene had a direct repeat, whereas in the rearranged NSP3 gene, no such pattern was found. PMID- 10867196 TI - Hepatitis C virus NS5A protein protects against TNF-alpha mediated apoptotic cell death. AB - Hepatitis C virus (HCV) often causes a prolonged and persistent infection which may lead to hepatocellular carcinoma. We have previously reported that the nonstructural 5A (NS5A) protein of HCV promotes cell growth [Ghosh, A.K., Steele, R., Meyer, K., Ray, R., Ray, R.B., 1999. Hepatitis C virus NS5A protein modulates cell cycle regulatory genes and promotes cell growth. J. Gen. Virol. 80, 1179 1183]. In this study, we investigated the role of HCV NS5A (genotype 1a, strain H) in TNF-alpha induced apoptotic cell death. HepG2 cells expressing NS5A exhibited an inhibitory role in relation to TNF-alpha mediated apoptotic cell death. The NS5A protein blocked the activation of caspase-3 and inhibited proteolytic cleavage of the death substrate poly (ADP-ribose) polymerase in TNF alpha induced cells. Together, these results suggest that HCV NS5A protein protects against TNF-alpha mediated apoptotic cell death. PMID- 10867197 TI - Inhibition of influenza C viruses by human MxA protein. AB - Human MxA protein was analyzed for its ability to inhibit the replication of different influenza C viruses. Three laboratory derivatives of viral strain C/Ann Arbor/1/50 were investigated, namely the parental wild-type virus C/AA-wt, the persistent variant C/AA-pi and the highly cytopathogenic variant C/AA-cyt. In addition, strain C/Paris/214/91 isolated from an influenza patient was used. Multiplication of all four viruses was suppressed in MxA-expressing Vero cells, as indicated by a decrease in viral RNA synthesis, viral protein synthesis, virion production and induction of a cytopathic effect. Inhibition correlated with the level of MxA expression. Furthermore, inhibition was independent of cell clone-specific differences in expression of virus receptors, as demonstrated by receptor reconstitution experiments. Thus, human MxA protein has antiviral activity against influenza C viruses. PMID- 10867198 TI - A deletion in the gI and gE genes of equine herpesvirus type 4 reduces viral virulence in the natural host and affects virus transmission during cell-to-cell spread. AB - In order to identify the role of the equine herpesvirus type 4 (EHV-4) glycoprotein I (gI) and E (gE) genes in determining viral virulence and their affect on the infection cycle, we constructed an EHV-4 recombinant strain containing a deletion in both gI and gE genes and its revertant. The recombinant was assayed in vitro in order to compare its growth kinetics with the parent and revertant viruses. Our results indicated that a deletion in the genes encoding gI and gE affected cell-to-cell spread of the virus in vitro. In order to assess the pathogenicity and vaccine efficacy of the recombinant in a natural host, colostrum-deprived foals were inoculated intranasally with the recombinant. Clinical signs obtained in foals upon the inoculation with the recombinant were milder than that for the revertant. This suggests that intact gI and/or gE genes are important factors in the expression of virulence in EHV-4 as in seen in the case of other herpesviruses. In addition, full protection against challenge infection was observed in foals, which had undergone a previous inoculation of the recombinant. PMID- 10867199 TI - The spheroidin of an entomopoxvirus isolated from the grasshopper Anacridium aegyptium (AaEPV) shares low homology with spheroidins from lepidopteran or coleopteran EPVs. AB - Based on virion morphology, the current virus taxonomy groups entomopoxviruses (EPVs) (Poxvirus: Entomopoxvirinae) from coleopteran and dipteran hosts in separated genera, wilts it keeps viruses infecting either lepidopteran or orthopteran hosts in the same genus. In contrast to the morphological criteria, the few data available from recent studies at the genetic level have suggested that EPVs infecting different insect orders are phylogenetically distant. In order to elucidate EPVs phylogeny we have cloned and sequence the highly conserved/highly expressed spheroidin gene of Anacridium aegyptium entomopoxvirus (AaEPV). This gene and its promoter is of interest for the development of genetic engineering on EPVs. The spheroidin gene was located in the AaEPV genome by Southern blot and hybridisation with specific degenerated oligonucleotides probes synthesised after partial sequencing of the purified spheroidin protein. A total of 3489 bp were sequenced. This sequence included the coding and promoter region of 969 residues 108. 8 kDa protein identified as spheroidin. AaEPV spheroidin contains 21 cysteine residues (2.2%) and 14 N-glycosylation putative sites distributed along the sequence. The cysteine residues are particularly abundant at the C-terminal end of the protein, with 11 residues in the last 118 aa. Our results confirm that the spheroidin is highly conserved only between EPVs isolated from the same insect order. Polyclonal antibodies raised against AaEPV spherules specifically revealed spheroidin in Western Blots failing to cross react with MmEPV or AmEPV spheroidins or MmEPV fusolin. Comparison of spheroidins at the aa level demonstrate that AaEPV spheroidin shares only 22.2 and 21.9% identity with the lepidopteran AmEPV and the coleopteran MmEPV spheroidins, respectively, but 82.8% identity with the orthopteran MsEPV spheroidin. Only two highly conserved domains containing the sequence (V/Y)NADTG(C/L) and LFAR(I/A) have been identified in all known spheroidins. The phylogenetic tree constructed according to the CLUSTALX analysis program revealed that EPVs are clearly separated in three groups - lepidopteran, coleopteran and orthopteran - according to the insect order of the virus hosts. In base to our results, the split of the genus Entomopoxvirus B dissociating lepidopteran and orthopteran EPVs into two different genera is suggested. PMID- 10867200 TI - Vascular risk factors for Alzheimer's disease: an epidemiologic perspective. AB - Vascular disease and Alzheimer's disease are both common disorders, in particular among elderly subjects. Therefore, it can be expected that the joint occurrence of these two disorders is not a rare phenomenon. In recent years, evidence is increasing that the two may be more closely linked than just by chance. Epidemiological studies have suggested that risk factors for vascular disease and stroke are associated with cognitive impairment and Alzheimer's disease, and that the presence of cerebrovascular disease intensifies the presence and severity of the clinical symptoms of Alzheimer's disease. In this paper, current knowledge on the relation between vascular risk factors and risk indicators and Alzheimer's disease will be reviewed. PMID- 10867201 TI - Risk factors for cerebral hypoperfusion, mild cognitive impairment, and dementia. AB - Putative risk factors accelerating mild cognitive decline and dementia were correlated with repeated measures of cerebral atrophy, CT, densitometry, perfusions, and cognitive testing among neurologically and cognitively normative aging volunteers. A total of 224 normative subjects at increased risk for cognitive decline were admitted to the study. Mean entry age was 59.5 +/- 15.8 years. Mean follow-up is 5.8 +/- 3.3 years. At follow-up, 22 developed mild cognitive impairment (41 CCSE >/= -3), 19 became demented-8 with Vascular type (VAD), 11 with Alzheimer's type (DAT)-and 183 remain cognitively unchanged. Cerebral atrophy, tissue densities, and perfusions were measured by Xe-CT. After age 60, cerebral atrophy, ventricular enlargement, and polio- and leuko-araiosis geometrically increased as perfusions declined. Risk factors accelerating perfusional decline, cerebral atrophy, polio-araiosis, and leuko-araiosis were: transient ischemic attacks (TIAs), hypertension, smoking, hyperlipidemia, and male gender. At age 71.5 +/- 11.9, mild cognitive impairment began accelerated by TIAs, hypertension and heart disease. Leuko-araiosis began before cognitive decline. TIAs, hypertension, and hyperlipidemia correlated with VAD. Excessive cortical perfusional decrease, gray and white matter hypodensities, and cerebral atrophy correlate with cognitive decline. PMID- 10867202 TI - The epidemiology of Alzheimer's disease and vascular dementia in Japanese and African-American populations: the search for etiological clues. PMID- 10867203 TI - Soluble beta-amyloid peptides mediate vasoactivity via activation of a pro inflammatory pathway. AB - Freshly solubilized beta-amyloid (Abeta) peptides display vasoactive properties, increasing both the magnitude and the duration of endothelin-1-induced vasoconstriction. We show that Abeta vasoactivity is mediated by the stimulation of a pro-inflammatory pathway involving activation of secretory phospholipase A(2) (PLA(2)), mitogen activated protein kinase (MAPK) kinase (MEK1/2), p38 MAPK, cytosolic PLA(2), and the release of arachidonic acid. Ultimately, arachidonic acid is metabolized into proinflammatory eicosanoids via the 5-lipoxygenase and cyclooxygenase-2 (COX-2) enzymes, both of which we show to be required for A beta vasoactivity. Accordingly, p38 MAPK activity is higher in the brains of transgenic mice that overproduce A beta, and COX-2 immunoreactivity is increased in the cerebrovasculature of these transgenic animals. Taken together, our data show that freshly solubilized A beta peptides can trigger a pro-inflammatory reaction in the vasculature that can be blocked by inhibiting specific target molecules, providing the basis for novel therapeutic intervention. PMID- 10867205 TI - Cleavage of amyloid precursor protein elicited by chronic cerebral hypoperfusion. AB - In the present study, we sought to determine whether low-grade, chronic vascular insufficiency induced in a rodent model of chronic cerebrohypoperfusion is sufficient, in and of itself, to trigger cleavage of the amyloid precursor protein (APP) into beta A-sized fragments. We report that chronic two vessel occlusion (2VO) results in progressive accumulation of beta A peptides detected by Western analysis in aged rats correlating with a shift in the immunohistochemical localization of APP from neurons to extracellular deposits in brain parenchyma. These data indicate that the 2VO paradigm reproduces features of beta A biogenesis characteristic of sporadic Alzheimer's disease. PMID- 10867204 TI - A damaged microcirculation contributes to neuronal cell death in Alzheimer's disease. AB - Alzheimer's disease (AD) involves multiple etiologic factors and a complex pathogenesis. Vascular factors are increasingly implicated in the pathogenesis of AD. In this paper we review evidence that AD brain microvessels are biochemically altered and contribute to neuronal injury and death by release of factors directly injurious to neurons. Our data show that when brain microvessels are "injured" by anoxia they produce high levels of reactive oxygen species. Comparisons of isolated brain microvessels from AD and age-matched controls show specific abnormalities in alpha(1) and beta receptors and in protein kinase C and protein kinase A signaling pathways. In AD but not in controls, the cerebral microcirculation expresses the inflammatory mediator CAP37 and over produces nitric oxide. Finally, we demonstrate that AD microvessels secrete toxic factors that cause neuronal cell death in vitro. These latter experiments showing that AD brain microvessels, in co-culture or vessel-conditioned media, cause lethal injury to neurons in culture, establish a direct link between endothelial cell products and neuronal cell death in this disease. PMID- 10867206 TI - Are cerebrovascular factors involved in Alzheimer's disease? AB - Recent epidemiological studies have shown that vascular risk factors may be involved in Alzheimer's disease (AD) as well as dementia in general. To investigate the relation between a vascular disorder and AD pathology, current criteria are defective because most depend on exclusion of a cerebrovascular disorder. Epidemiological studies have indicated the possibilities that arteriosclerosis, abnormal blood pressure, diabetes mellitus and smoking may be related to the pathogenesis of AD. As for the mechanism that vascular disorders influence AD, it is presumed that amyloid deposition may be caused by a vascular disorder. Alternatively, a vascular event may cause progression of subclinical AD to a clinical stage. Insulin resistance and apolipoprotein E may also be involved in these mechanisms. Our studies show that ischemia-induced the Alzheimer associated gene presenilin 1 (PS1) and endoplasmic reticulum-stress, generated from a vascular disorder, may unmask clinical AD symptoms caused by presenilin mutation, suggesting that a vascular factor might be involved in the onset of familial AD. PMID- 10867207 TI - Chronic cerebrovascular ischemia in aged rats: effects on brain metabolic capacity and behavior. AB - The objective of this study was to model one of the risk factors for the development of late-onset Alzheimer's disease, decreased cerebral blood flow. Aging rats were tested for visuospatial behavioral deficits after permanent surgical occlusion of both carotid arteries. This was followed after 4 weeks by quantitative cytochrome oxidase histochemical mapping of metabolic capacity throughout the brain. The brain regions affected were related to observed deficits in spatial memory (CA1 and posterior parietal cortex), visually guided movements (superior colliculus and secondary visual cortex), motor coordination (red nucleus), and escape behavior (central gray). The results suggest that deficits in visuospatial learning are not exclusively the result of hippocampal dysfunction, but may be directly correlated with altered oxidative energy metabolism in other integrative visuomotor regions identified in this study. It was concluded that chronic cerebrovascular ischemia in this aged rat model produces neurometabolic and behavioral alterations that may be relevant for an increased risk for the development of Alzheimer's disease. PMID- 10867208 TI - Are Alzheimer's disease, hypertension, and cerebrocapillary damage related?. AB - Alzheimer's disease (AD) patients are often subject to vascular dysfunction besides their specific CNS pathology, which warrants further examination of the interaction between vascular factors and the development of dementia. The association of decreased cerebral blood flow (CBF) or hypertension with AD has been a target of growing interest. Parallel with physiological changes, the cerebral capillaries in AD are also prone to degenerative processes. The microvascular abnormalities that are the result of such degeneration may be the morphological correlates of the vascular pathophysiology pointing to a compromised nutrient transport through the capillaries. Animal models have been developed to study the consequences of hypertension and reduced CBF. Spontaneously hypertensive rats are widely used in hypertension research whereas ligation of the carotid arteries has become a method to produce cerebral hypoperfusion. Based on these models, we propose a relationship between hypertension, cerebral hypoperfusion, cerebral capillary malformation and cognitive decline as it occurs in AD. We suggest that the above conditions are functionally related and can contribute to the progression of AD. PMID- 10867209 TI - The role of apolipoprotein E in Alzheimer's disease, acute brain injury and cerebrovascular disease: evidence of common mechanisms and utility of animal models. AB - The epsilon 4 allele of apolipoprotein E (APOE denotes gene; apoE denotes protein) is a major risk factor for Alzheimer's disease (AD). More recent evidence indicates an association with a poor outcome after acute brain injury including that due to head trauma and intracerebral hemorrhage. APOE gene polymorphism also influences the risk of hemorrhage in cerebral amyloid angiopathy. These diverse brain disorders seem to have some mechanisms in common. The multiplicity of the roles of apoE within the central nervous system is currently being unraveled. For example, apoE can interact with amyloid beta protein and tau, proteins central to the pathogenesis of AD. In addition to these effects, it is proposed that one of the major functions of apoE is to mediate neuronal protection, repair and remodeling. In all of the different roles proposed, there are marked apoE-isoform specific differences. Although it remains to be clarified which is the most important mechanism(s) in each disorder in which apoE is involved, these isoform specific differences seem to underly a genetically determined susceptibility to outcome from acute brain injury and to AD with APOE epsilon 4 conferring relative vulnerability. This review focuses on apoE research, from clinical studies to animal models, in AD, acute brain injury and cerebrovascular disease and explores the common mechanisms that may explain some of the complex underlying neurobiology. PMID- 10867211 TI - Hemorheological changes and overproduction of cytokines from immune cells in mild to moderate dementia of the Alzheimer's type: adverse effects on cerebromicrovascular system. AB - An association between hemorheological alterations (i.e., whole-blood and plasma hyperviscosity, reduced erythrocyte deformability, increased red cell aggregation, hyperfibrinogenemia and increased acute-phase protein levels) and the mild stage of senile dementia of the Alzheimer's type (DAT) was suggested in the present study. In particular, hyperfibrinogenemia and the increase of erytrhocyte aggregation were correlated with the increased generation and release of TNF-alpha and IFN-gamma (spontaneous release and IL-2-modulated release) from natural killer (NK) lymphocytes (CD16+, CD56+, CD3- cells) of patients with DAT; whereas a normal cytokine release from NK cells was found in healthy old subjects and in patients with vascular dementia (VaD). The in vitro and in vivo administration of the hemorheologic drug pentoxifylline (PTX) significantly reduced spontaneous and IL-2-modulated cytokine overproduction from NK cells (in vitro effects with 500 U/ml and 1000 U/ml/NK cells) and improved all the hemorheological parameters. Taken together, these data suggest that disturbances of cerebrovascular flow and of hemorheology could be considered a negative component related to the pathogenesis and progression of DAT neurodegeneration. The association between hemorheological changes and alterations of TNF-alpha and IFN-gamma release from NK may indicate a potential immunorheologic mechanism associated with cerebrovascular damage in DAT and could suggest the use of vascular protective drugs as support of the main pharmacological and non pharmacological therapy of AD. PMID- 10867210 TI - Hemodynamic changes during aging associated with cerebral blood flow and impaired cognitive function. AB - This study investigates the age associated changes in hemorheological properties and cerebral blood flow. Partial correlations indicate that part of the age dependent decrease in flow velocities can be attributed to a hemorheological decrement resulting in part from enhanced oxidative stress in the aged. A possible link with Alzheimer's pathology is suggested by the augmented hemorheological impairment resulting from in vitro incubation of red cells with amyloids. These results suggest that in aging, oxidative stress as well as amyloids may influence the fluid properties of blood, resulting in a potential decrement in blood flow and oxygen delivery to the brain. Animal intervention studies further demonstrate that altered hemorheological properties of blood can actually influence cognitive function. The relationships shown to exist between hemorheology, blood flow, amyloids, oxidative stress, and cognitive function suggest that these factors may be one of the mechanisms operating in the complex etiology of Alzheimer's disease. PMID- 10867212 TI - A promising approach to the treatment of multi-infarct dementia. AB - Cerebrovascular multi-infarct dementia (MID) is associated with high fibrinogen levels and lipid fractions leading to an increase of both plasma and whole blood viscosity as well as raised aggregability of blood cells. One important goal in the treatment of MID therefore should be to reduce fibrinogen and lipoproteins and thereby to improve the Hemorheological State. The effect of heparin-induced extracorporeal LDL/fibrinogen precipitation (HELP), a method for safe and immediate reduction of parameters relevant to hemorheology, such as plasma fibrinogen and the lipoproteins, was investigated in 141 patients with MID. All the patients underwent two HELP applications within 8 days. The impact of HELP on MID was studied by changes of laboratory data and by evaluation of clinical symptoms before and after treatment. Each HELP session caused an immediate, safe and significant reduction of important rheological parameters such as fibrinogen, whole blood viscosity at high and low shear rate, plasma viscosity and red cell transit time. Also total cholesterol and low-density lipoprotein, lipoprotein(a) and the triglycerides had been reduced significantly. The results in laboratory measurement were followed by a statistically significant improved neurologic recovery, represented in the values of the Mathew Scale, the Mini Mental State Examination and the Activities-of-Daily-Living-Test. These results can indicate the importance and influence of hemorheology on clinical symptoms in MID. PMID- 10867213 TI - Perfusion abnormalities in prodromal AD. AB - Cerebral perfusion abnormalities in patients with established Alzheimer's disease (AD) are most commonly seen in the temporoparietal cortex. As the disease progresses, this perfusion pattern is increasingly prevalent. Recently, investigators have begun to examine the patterns of perfusion among individuals at risk for AD. To date, such studies have been conducted either in individuals who have a progressive memory difficulty but do not yet meet clinical criteria for AD, or in individuals with a genetic risk factor or family history of AD, either with or without a memory problem. These latter studies suggest that a set of brain regions show decreased perfusion during the prodromal phase of AD, and that a brain network or networks with multiple nodes is affected early in the course of AD. These perfusion abnormalities may also shed light on how AD progresses during the prodromal phase of disease and may ultimately lead to improved diagnosis or methods of monitoring response to treatment. PMID- 10867214 TI - How does the apolipoprotein E genotype modulate the brain in aging and in Alzheimer's disease? A review of neuroimaging studies. AB - The epsilon 4 allele of apolipoprotein E is a risk factor for Alzheimer's disease, but also a modulator of its clinical picture. In this paper, recent research in neuroimaging of aging and Alzheimer's disease in relation to apolipoprotein E is reviewed, emphasizing the advances but also the controversies. Further, the possible clinicopathological implications of these findings are discussed. PMID- 10867215 TI - Vascular and metabolic reserve in Alzheimer's disease. AB - Vascular and metabolic reserve were analyzed in probable Alzheimer's disease (AD) and vascular dementia (VaD). Cerebral blood flow (CBF), cerebral blood volume (CBV), cerebral metabolic rate of oxygen (CMRO(2)), and oxygen extraction fraction (OEF) were measured quantitatively with positron emission tomography (PET). Vascular reactivity (VR) was also calculated by comparing the CBF during 5% CO(2) inhalation with the CBF during normal breathing. Vascular transit time (VTT) that was calculated as a ratio of CBV/CBF and VR reflect vasodilating capacity of the small resistance vessels, whereas OEF designates metabolic (oxygen-extraction) reserve in threatening brain ischemia. Significant increase in OEF was seen in the parieto-temporal cortex and both VTT and VR were preserved in AD patients. By constrast, there was no significant increase in OEF whereas VTT was prolonged and VR was markedly depressed in VaD patients. The increase of OEF and preserved VTT and VR seen in AD patients indicate the possible participation of vascular factors in the pathogenesis of AD perhaps at the capillary level. PMID- 10867216 TI - Aberrant expression of nitric oxide synthase III in Alzheimer's disease: relevance to cerebral vasculopathy and neurodegeneration. AB - Alzheimer's disease (AD) has heterogeneous pathology, in part due to the large subset of cases (AD+CVD) with superimposed vascular lesions that are sufficient in number and distribution to accelerate the clinical course of dementia. Brains with AD+CVD have lower densities of neurofibrillary tangles and A beta-amyloid diffuse plaques, and increased numbers of cerebral vessels exhibiting p53 associated apoptosis relative to brains with uncomplicated AD. AD and AD+CVD both exhibit altered expression of the nitric oxide synthase 3 (NOS-III) gene; however, in AD+CVD, reduced NOS-III expression in cerebral vessels is associated with an increased frequency of vascular lesions, vascular smooth muscle cell apoptosis, and A beta-amyloid plaques. In contrast, experimental and spontaneous focal acute and subacute cerebral infarcts are associated with increased NOS-III expression in perifocal neurons, glial cells, cerebrovascular smooth muscle and endothelial cells, and diffuse A beta-amyloid plaque formation. This suggests that ischemic injury and oxidative stress can precipitate NOS-III-mediated cell loss and neurodegeneration. A role for aging-associated impaired mitochondrial function as a contributing factor in AD and CVD is suggested by the reduced levels of mitochondrial protein observed in AD and AD+CVD cortical neurons and vascular smooth muscle and endothelial cells. The aggregate findings suggest that cell loss and neurodegeneration may be mediated by somewhat distinct but overlapping mechanisms in AD and AD+CVD. PMID- 10867217 TI - The role of cerebral ischemia in Alzheimer's disease. AB - The Alzheimer type of dementia and stroke are known to increase at comparable rates with age. Recent advances suggest that vascular risk factors linked to cerebrovascular disease and stroke in the elderly significantly increase the risk of developing Alzheimer's disease (AD). These include atherosclerosis, atrial fibrillation, coronary artery disease, hypertension, and diabetes mellitus. Moreover, review of various autopsy series shows that 60-90% of AD cases exhibit variable cerebrovascular pathology. Although some vascular lesions such as cerebral amyloid angiopathy, endothelial degeneration, and periventricular white matter lesions are evident in most cases of AD, a third will exhibit cerebral infarction. Despite the interpretation of pathological evidence, longitudinal clinical studies suggest that the co-existence of stroke and AD occurs more than by chance alone. Strokes known to occur in patients with Alzheimer syndrome and most frequently in the oldest old substantially worsen cognitive decline and outcome, implicating some interaction between the disorders. Nevertheless, the nature of a true relationship between the two disorders seems little explored. What predisposes to strokes in underlying cognitive decline or AD? Is it possible that cerebral ischemia is a causal factor for AD? I examined several vascular factors and the vascular pathophysiology implicated in stroke and AD, and propose that cerebral ischemia or oligemia may promote Alzheimer type of changes in the aging brain. Irrespective of the ultimate pathogenetic mechanism, these approaches implicate that management of peripheral vascular disease is important in the treatment or prevention of Alzheimer's disease or mixed dementia. PMID- 10867218 TI - Critically attained threshold of cerebral hypoperfusion: the CATCH hypothesis of Alzheimer's pathogenesis. AB - This review discusses the experimental and clinical data which indicate that chronic cerebral hypoperfusion can affect metabolic, anatomic, and cognitive function adversely. In aged but not young animals, chronic brain hypoperfusion results in regional pre- and post-synaptic changes, protein synthesis abnormalities, energy metabolic dysregulation, reduced glucose utilization, cholinergic receptor loss, and visuo-spatial memory deficits. Additionally, aging animals that are kept for prolonged periods of time after chronic brain hypoperfusion, also develop brain capillary degeneration in CA1 hippocampus and neuronal damage extending from the hippocampal region to the temporo-parietal cortex where neurodegenerative tissue atrophy eventually forms. All these pathologic events occur in rodents in the absence of senile plaques and neurofibrillary tangles. Alzheimer brains reveal similar biochemical and structural changes as those experimentally induced in aging animals. Moreover, regional cerebral hypoperfusion is one of the earlier (if not the earliest) clinical manifestations in both the sporadic and familial forms of Alzheimer's disease. In addition, therapy that improves or increases cerebral perfusion is generally of some benefit to Alzheimer patients. Conversely, a variety of disorders with different etiologies that impair or diminish cerebral perfusion are reported to be risk factors for this dementia. These findings have prompted us to propose the concept that advanced aging in the presence of a vascular risk factor can converge to create a critically attained threshold of cerebral hypoperfusion (CATCH) that triggers regional brain microcirculatory disturbances and impairs optimal delivery of energy substrates needed for normal brain cell function. The outcome of this defect generates a chain of events leading to the progressive evolution of brain metabolic, cognitive and tissue pathology that characterize Alzheimer's disease. The possible role of CATCH in familial and early onset Alzheimer's disease is briefly discussed from a theoretical vantagepoint. The growing and most recent evidence in support of the CATCH concept is the focus of this review. PMID- 10867219 TI - Monoamine oxidase-B inhibitors in the treatment of Alzheimer's disease. AB - The monoamine oxidase-B (MAO-B) inhibitor L-deprenyl (Selegiline) is effective in treating Parkinson's disease and possibly Alzheimer's disease, with a concomitant extension of life span. It has been suggested that the therapeutic efficacy of L deprenyl may involve actions other than the inhibition of the enzyme MAO-B. This article reviews some novel actions of L-deprenyl and suggests that stimulation of nitric oxide (NO) production could be central to the action of the drug. L Deprenyl induced rapid increases in NO production in brain tissue and cerebral blood vessels. In vitro or in vivo application of L-deprenyl produced vasodilatation. The drug also protected the vascular endothelium from the toxic effects of amyloid-beta peptide. Because NO modulates activities including cerebral blood flow and memory, and reduced NO production has been observed in AD brain, stimulation of NO production by L-deprenyl could contribute to the enhancement of cognitive function in AD. MAO-B inhibitors are unique in that they exert protective effects on both vascular and neuronal tissue and thus warrant further consideration in the treatment of vascular and neurodegenerative diseases associated with aging. PMID- 10867220 TI - Agrin and microvascular damage in Alzheimer's disease. AB - Heparan sulfate proteoglycans (HSPGs) are ubiquitously present within the perivascular basement membrane, and have been shown to be altered in patients with Alzheimer's Disease (AD). Although the HSPG agrin clearly orchestrates the differentiation of the neuromuscular junction, its role in the brain remains unclear. Growing evidence suggests that agrin may be an important vascular basement membrane (VBM)-associated HSPG. In previous studies, we demonstrated that agrin is present throughout the brain microvasculature, as well as in neuronal cell bodies. AD brains exhibited fragmentation of VBM-associated agrin. Agrin immunoreactivity was also seen within senile plaques and neurofibrillary tangles. These changes were accompanied by the appearance of an additional pool of insoluble agrin. In the present study, we provide further evidence for microvascular damage in AD, by examining the distribution of agrin and laminin within the VBM, and by measuring the agrin concentration within hippocampus and prefrontal cortex. Furthermore, we assessed blood-brain-barrier (BBB) leakage by examining the perivascular distribution of prothrombin immunoreactivity. Soluble agrin levels were increased approximately 30% in Braak stage III-VI AD patients relative to age-matched controls. Furthermore, agrin and laminin exhibited identical patterns of VBM fragmentation in AD and colocalized with beta-amyloid in senile plaques. Microvascular changes were associated with the appearance of perivascular prothrombin immunoreactivity. Our data suggest that agrin is an important VBM-associated HSPG in the brain and that agrin levels are altered in association with microvascular damage in AD. PMID- 10867221 TI - Vascular abnormalities: the insidious pathogenesis of Alzheimer's disease. AB - Alzheimer's disease (AD) and cerebrovascular dementia (CVD) are two major causes of senile dementia in elderly individuals. Mounting evidence from epidemiological, clinical, and neuropathological studies suggests that there is considerable overlap between AD and CVD with respect to risk factors, prevalence, and pathological changes. Although our lack of understanding on the important contribution of vascular disturbance to pathogenesis of AD has further hindered our understanding of AD, data on the roles of cerebrovascular diseases and systemic vascular diseases in AD need to be carefully analyzed to avoid misinterpretation. Here, we review studies on the cerebral vasculature, cardiac vasculature, and apoE that lead us to contend that vascular abnormalities are likely an important mechanism underlying dementia. Because early and aggressive intervention is available to prevent and treat a number of vascular diseases, therapies that attenuate vascular risk factors could be valuable in preventing and treating AD. PMID- 10867222 TI - Neuropathology of mitral valve prolapse in man and cardiopulmonary bypass (CPB) surgery in adolescent Yorkshire pigs. AB - We investigated the brains of non-demented individuals with mitral valve prolapse (MVP) and found evidence of Alzheimer-like lesions. This neuropathology consisted of premature presence of beta-amyloid-containing senile plaques (SP) without increased prevalence of neurofibrillary tangles. Low levels of SP occurred in 20 to 45- year-old subjects with MVP, and much greater densities were observed in subjects between 45 and 62 years of age. We also investigated the brains of adolescent Yorkshire pigs undergoing cardiopulmonary bypass surgery and likewise found evidence of Alzheimer-like neuropathology. This took the form of intraneuronal accumulation of beta-amyloid immunoreactivity and increasing numbers of Alz-50 immunoreactive neurons with reduced recovery of cardiac efficiency after the surgery. Based on prevailing concepts in Alzheimer's disease, it is feasible to hypothesize that cognitive dysfunction occurring after cardiopulmonary bypass surgery with coronary artery grafting or valve repair/replacement is a functional sequela of AD-like neuropathology. This postulate is based on the premise that an individual seeking such surgery would have pre-existing, elevated AD-like neuropathology to start with. It is further coupled with the probability that these forms of cardiovascular surgery exacerbate the extent and progression of AD-like neuropathology. PMID- 10867223 TI - Longitudinal effects of estrogen replacement therapy on PET cerebral blood flow and cognition. AB - Observational studies suggest that estrogen replacement therapy (ERT) may protect against age-related memory decline and lower the risk of Alzheimer's disease (AD). This study aimed to characterize the neural substrates of those effects by comparing 2-year longitudinal changes in regional cerebral blood flow (rCBF) in 12 ERT users and 16 nonusers. Positron emission tomography (PET) measurements of rCBF were obtained under three conditions: rest, and verbal and figural recognition memory tasks. Groups showed different patterns of change in rCBF over time in a number of brain areas. These group differences, for the most part, reflected regions of increased rCBF over time in users compared to nonusers. The greatest differences between ERT users and nonusers were in the hippocampus, parahippocampal gyrus, and temporal lobe, regions that form a memory circuit and that are sensitive to preclinical AD. Across a battery of standardized neuropsychological tests of memory, users obtained higher scores than did nonusers of comparable intellect. Group differences in longitudinal change in rCBF patterns may reflect one way through which hormones modulate brain activity and contribute to enhanced memory performance among ERT users. PMID- 10867224 TI - Preface PMID- 10867225 TI - Bacterial protein toxins targeting rho GTPases. AB - Several bacterial protein toxins target eukaryotic cells by modulating the functions of Rho GTPases that are involved in various signal processes and in the regulation of the actin cytoskeleton. The toxins inhibit Rho functions by ADP ribosylation or glucosylation and activate them by deamidation and transglutamination. New findings indicate that the GTPases are also targeted by various 'injected' toxins which are introduced into the eukaryotic cells by the type-III secretion system. The injected toxins do not covalently modify Rho GTPases, but manipulate their regulatory GTPase cycle by acting as GTPase activating proteins or guanine nucleotide exchange factors. PMID- 10867226 TI - Phenotypic and genotypic comparisons of 23 strains from the Bacillus cereus complex for a selection of known and putative B. thuringiensis virulence factors. AB - Sixteen Bacillus thuringiensis, four Bacillus cereus and three Bacillus anthracis isolates were screened for a selection of known and putative B. thuringiensis virulence factors. PCR primers were designed to detect genes for phosphatidylcholine specific phospholipase C, phosphatidylinositol specific phospholipase C, immune inhibitor A, vegetative insecticidal protein 3A, a protein proposed to be involved in capsule synthesis, a newly identified Ser/Thr kinase homologue and enterotoxin entS. Motility, the presence of flagella, haemolysis, chitinase and lecithinase production were also evaluated. The widely varying profiles of the 23 strains from the complex provide a pool of different genotypes that can help to identify factors involved in pathogenicity. PMID- 10867227 TI - Role of the Salmonella abortusovis virulence plasmid in the infection of BALB/c mice. AB - Following oral inoculation of BALB/c mice, Salmonella abortusovis strain SS44 was recovered in lower numbers from the Peyer's patches and mesenteric lymph nodes compared with S. typhimurium strain SL1344, whereas splenic infections were equivalent between the two serovars. SS44 was cured of its virulence plasmid or subjected to mutagenesis of the spv genes, and the Spv(-) derivatives were tested for virulence in mice. Plasmid-cured S. abortusovis SU40 retained virulence in BALB/c mice when inoculated intraperitoneally. On the other hand, mice infected orally with SU40 had greatly reduced splenic infection compared to those infected with wild-type SS44. Similar results were obtained after Tn5 insertion mutagenesis of the spvR gene or deletion of the spvABCD locus. These results suggest that in the gut-associated lymphoid tissues S. abortusovis may replicate less than S. typhimurium and that the S. abortusovis virulence plasmid primarily affects systemic infection after oral inoculation but not after intraperitoneal administration in the mouse model. PMID- 10867228 TI - Regulation of 36-kDa beta-1,3-glucanase synthesis in Trichoderma harzianum. AB - The effect of carbon sources on the level of beta-1,3-glucanases in the culture filtrates of Trichoderma harzianum (Tc) was investigated. Enzyme activity was detected in all carbon sources, but highest levels were found when laminarin and purified cell walls were used. Three isoforms of beta-1,3-glucanase were produced during growth of the fungus on purified cell walls. Two isoforms were produced on chitin, chitosan, N-acetylglucosamine and laminarin, while only one was detected when the fungus was grown on cellulose and glucose. A 36-kDa beta-1,3-glucanase (GLU36) was secreted from T. harzianum (Tc) grown on all carbon sources tested as demonstrated by Western blot analysis. We found that a significant increase in the level of GLU36 in the culture filtrate follows glucose exhaustion, suggesting that this enzyme is controlled by carbon catabolite repression. PMID- 10867229 TI - An AraC-like transcriptional activator is required for induction of genes needed for alpha-galactoside utilization in Sinorhizobium meliloti. AB - The nodulating bacterium Sinorhizobium meliloti can utilize alpha-galactosides like melibiose and raffinose as sole sources of carbon and energy. We show that this utilization requires an AraC-like transcriptional activator, AgpT. When agpT was inactivated, Rhizobium meliloti could not utilize alpha-galactosides or induce genes required for transport and catabolism of these sugars. The agpT gene was not essential for the establishment of an effective nitrogen-fixing symbiosis. PMID- 10867230 TI - Use of a lux reporter system for monitoring rapid changes in alpha-toxin gene expression in Clostridium perfringens during growth. AB - To determine whether the luxA-luxB reporter system is suitable as a sensitive reporter for rapid real-time measurements of alpha-toxin gene (cpa) expression in Clostridium perfringens, and to widen the range of alpha-toxin-producing C. perfringens strains examined with respect to cpa expression during growth, the reporter plasmid pPS14 (possessing the alpha-toxin promoter region plus 0.7 kb of upstream region linked to the luxA-luxB genes), was used in batch growth experiments of C. perfringens P90.2.2, an alpha-toxin-producing strain with no known association with disease. Levels of in vivo bioluminescence obtained during growth were broadly in agreement with previous mRNA and reporter studies of cpa expression (Bullifent et al., FEMS Microbiol. Lett. 131 (1995) 99-105), confirming the suitability of lux as an accurate reporter system in this organism, but the sensitive nature of the lux reporter permitted the in vivo detection of a very rapid reduction in expression during late-exponential phase that was not attributable to loss in cell viability or limiting bioluminescence assay substrates. There was also a small peak in cpa expression in early- to mid exponential phase cells, that was not detected in previous studies with other reporters. This may be indicative of the exquisite sensitivity of the lux reporter, or this may be a difference in cpa expression that occurs specifically in this C. perfringens strain. Whichever is the case, these results confirm the complexity of alpha-toxin gene expression in different strains of this pathogenic bacterium. PMID- 10867231 TI - Identification of a fmtA-like gene that has similarity to other PBPs and beta lactamases in Staphylococcus aureus. AB - We identified a gene from Staphylococcus aureus, flp (fmtA-like protein), encoding a protein of 489 amino acid residues with a molecular mass of 56.4 kDa. The deduced amino acid sequence shows similarity to previously characterized penicillin binding proteins (PBPs) and FmtA of S. aureus (one of the factors which affect methicillin resistance). FLP protein has three motifs, which are conserved in PBPs and beta-lactamases, suggesting that it might be associated with cell wall synthesis. Recombinant FLP protein, however, lacks penicillin binding activity, and the inactivation of flp in two methicillin-resistant S. aureus strains did not cause a reduction in the methicillin resistance. PMID- 10867232 TI - Isolation and characterization of an Achromobacter xylosoxidans strain B3 and other bacteria capable to degrade the synthetic chelating agent iminodisuccinate. AB - Three bacterial strains were isolated, which used the synthetic chelating agent iminodisuccinate (IDS) as sole carbon source for growth in mineral salts media (MSM). Taxonomic analysis and 16S rDNA sequence analysis identified one of these isolates (B3), which was isolated from sewage sludge, as a strain of Achromobacter xylosoxidans subsp. xylosoxidans. It exhibited a doubling time of approximately 3 h in liquid MSM supplemented with IDS and grew even in the presence of 1.0% (w/v) IDS. Since photometric and high performance liquid chromatography analysis showed that IDS, which came onto the market only recently as an alternative for ethylenediaminetetraacetate, was completely degraded by axenic cultures of bacteria; it will probably be readily degraded in the environment. PMID- 10867233 TI - A high yielding mutant of mycobacteriophage L1 and its application as a diagnostic tool. AB - L1 is a lysogenic phage of mycobacteria, which along with L5 and D29 constitute a closely linked family of homoimmune mycobacteriophages. These phages can be potentially used for genetic engineering of mycobacteria and diagnosis of mycobacterial infection. The effectiveness of such phage based systems depends on the efficiency with which they infect and grow within target cells. While working with phage L1c1ts which is a temperature sensitive mutant of phage L1, we observed that high yielding phage stocks were generated by repeated passage through the host, Mycobacterium smegmatis. A plaque purified mutant L1-P2, obtained from one such high yielding stock, when analyzed further was found to infect host cells with increased efficiency. The DNA obtained from L1-P2 was examined by restriction digestion, and it was observed that spontaneous loss of DNA fragment from the right arm, which encodes early regulatory factors, had occurred. It has been further demonstrated that the high yielding property of the mutant phage could be utilized to increase the sensitivity of mycobacteriophage based detection systems. PMID- 10867234 TI - Cloning, sequencing, and expression in Escherichia coli of a cytochrome P450 gene from Cunninghamella elegans. AB - A polyclonal antibody against microsomes of a fungus, Cunninghamella elegans, was used to screen a C. elegans cDNA library. A cDNA clone, containing an open reading frame (ORF) encoding a protein of 389 amino acids (aa), was obtained. GenBank comparison (BLAST) showed that the protein was closely related to P450 because a heme-binding region, which is highly conserved in all P450 sequences, was found in the ORF protein. Using an oligo probe designed from this C. elegans heme-binding region to rescreen the cDNA library, we obtained three new clones. Sequence comparison showed that the three clones, with different length cDNA inserts, were from the same mRNA of the C. elegans P450 gene. One clone had the full C. elegans P450 gene, encoding 473 aa with a molecular mass of 54958.60, whereas the 389 was a part of the 473 aa without the N-terminal. The entire C. elegans P450 gene was successfully subcloned and overexpressed in a plasmid Escherichia coli system (pQE30). Immunostaining with three antibodies (CYP1A1, CYP2E1, and CYP3A1) against mammalian P450 enzymes and benzidine staining for hemoproteins showed positive results for the recombinant protein expressed in E. coli. A phylogenetic tree was constructed by comparison of other fungal P450s to the C. elegans sequence. The C. elegans P450 clustered close to the cyp51 family and was named cyp509A1 by the International Committee on the Nomenclature for Cytochrome P450 Enzymes. PMID- 10867235 TI - Positioning of medial actin rings affected by eccentrically located nuclei in a fission yeast mutant having large vacuoles. AB - In Schizosaccharomyces pombe wild-type cells, the nucleus positions in the middle of the cell where the cortical actin ring is assembled prior to septum formation. ste12 mutants contain a few large vacuoles. In a considerable fraction of ste12 cells, the nuclei and septa were eccentrically positioned. Both extension of spindle microtubules in the anaphase and post-mitotic migration of sister nuclei to the cell poles were partially disrupted, probably due to enlarged vacuoles. In spite of the eccentric positioning of nuclei, the cortical actin ring overlays the displaced pre-mitotic nucleus. This observation supports the notion that the nucleus dictates the division site in fission yeast. PMID- 10867236 TI - Metabolism of ricinoleic acid into gamma-decalactone: beta-oxidation and long chain acyl intermediates of ricinoleic acid in the genus Sporidiobolus sp. AB - In order to study differences in gamma-decalactone production in yeast, four species of Sporidiobolus were cultivated with 5% of methyl ricinoleate as the lactone substrate. In vivo studies showed different time courses of intermediates of ricinoleic acid breakdown between the four species. In vitro studies of the beta-oxidation system were conducted with crude cell extracts of Sporidiobolus spp. and with ricinoleyl-CoA (RCoA) as substrate. The beta-oxidation was detected by measuring acyl-CoA oxidase, 3-hydroxyacyl-CoA dehydrogenase activities, and acetyl-CoA production. The time courses of the CoA esters resulting from RCoA breakdown by crude extract of Sporidiobolus spp. permit the proposal of different metabolic models in the yeast. These models explained the differences observed during in vivo studies. PMID- 10867238 TI - The fib locus in Streptococcus pneumoniae is required for peptidoglycan crosslinking and PBP-mediated beta-lactam resistance. AB - Penicillin resistance in pneumococci is mediated by modified penicillin-binding proteins (PBPs) that have decreased affinity to beta-lactams. In high-level penicillin-resistant transformants of the laboratory strain Streptococcus pneumoniae R6 containing various combinations of low-affinity PBPs, disruption of the fib locus results in a collapse of PBP-mediated resistance. In addition, crosslinked muropeptides are highly reduced. The fib operon consists of two genes, fibA and fibB, homologous to Staphylococcus aureus femA/B which are also required for expression of methicillin resistance in this organism. FibA and FibB belong to a family of proteins of Gram-positive bacteria involved in the formation of interpeptide bridges, thus representing interesting new targets for antimicrobial compounds for this group of pathogens. PMID- 10867237 TI - Three-dimensional modelling of the catalytic domain of Streptococcus mutans glucosyltransferase GtfB. AB - Glucosyltransferases (GtfB/C/D) of Streptococcus mutans, a pathogen for human dental caries, synthesize water-insoluble glucan through the hydrolysis of sucrose. Genetic and biochemical approaches have identified several active sites of these enzymes, but no three-dimensional (3D) structural evidence is yet available to elucidate the subdomain arrangement and molecular mechanism of catalysis. Based on a combined sequence and secondary structure alignment against known crystal structures of segments from closely related proteins, we propose here the 3D model of an N-terminal domain essential for the sucrose binding and splitting in GtfB. A Tim-barrel of (alpha/beta)(8) structural characteristics is revealed and the structural correlation for two peptides is described. PMID- 10867239 TI - Detection of antitumor and antimicrobial activities in marine organism associated actinomycetes isolated from the Taiwan Strait, China. AB - The purpose of this work was to screen the actinomycetes having antitumor or antimicrobial activity, which were isolated from the surface, epidermis and intestines of sea plants and animals collected from the Taiwan Strait, China. Antitumor activity was studied by the MTT assay and DNA target activity was studied by the biochemical induction assay while antimicrobial activity was determined by observing bacterial and fungal growth inhibition. 20. 6% of marine actinomycete cultures displayed cytotoxic activity on P388 cells at dilutions at and below 1:320 and 18.6% on KB cells. 2. 96% of marine actinomycete cultures displayed inducing activity. Among all marine actinomycetes isolated, the genus Micromonospora has the highest positive rate of inducing activity. However, most antimicrobial activity was found in the genus Streptomyces. These results indicate that marine organism associated actinomycetes could be a promising source for antitumor and antimicrobial bioactive agents. PMID- 10867240 TI - Decolourisation of synthetic textile dyes by Phlebia tremellosa. AB - Phlebia tremellosa decolourised eight synthetic textile dyes (200 mg l(-1)) by greater than 96% within 14 days under stationary incubation conditions. High performance liquid chromatography analysis of culture supernatants indicated that Remazol Black B was degraded by the fungus, however, complete mineralisation did not occur as a colourless organic breakdown product accumulated. Laccase activity was detectable in culture supernatants after 5 days when the fungus was grown in the presence of an artificial textile effluent, with activity reaching a maximum of 15 U l(-1) on day 14. PMID- 10867241 TI - Interaction of omega-conotoxin and the membrane calcium transport system of Escherichia coli. AB - omega-Conotoxin, a calcium channel blocker, inhibits chemotaxis by Escherichia coli. To test whether omega-conotoxin acts at the cytoplasmic membrane, the kinetics of 125I-omega-conotoxin binding was investigated. 125I-omega-Conotoxin bound to Tris-EDTA-permeabilized cells or right-side-out membrane vesicles with saturation kinetics. Binding of 125I-omega-conotoxin to membrane vesicles was inhibited by Ca(2+) ions, but not by Mg(2+) ions. The calA mutant, defective in calcium transport, was more resistant to omega-conotoxin inhibition of chemotaxis than the parental wild-type. 125I-omega-Conotoxin binding to membrane vesicles indicated that both the wild-type and the calA mutant had similar K(D)s for omega conotoxin binding. However, the saturation level was higher with the calA mutant, indicating that there are more binding sites in the calA mutant. Thus, calA does not directly affect the affinity of the omega-conotoxin binding site. Chemical cross-linking experiments identified two proteins as potential omega-conotoxin receptors. PMID- 10867242 TI - The role of the bacitracin ABC transporter in bacitracin resistance and collateral detergent sensitivity. AB - The bacitracin resistance of Bacillus licheniformis, a producer of bacitracin, is mediated by the ABC transporter Bcr. Bacillus subtilis cells carrying bcr genes on high-copy number plasmids developed collateral detergent sensitivity, as did human cells with overexpressed multidrug resistance P-glycoprotein. Resistance against bacitracin and sensitivity of resistant cells to detergents were shown to be inseparable phenomena associated with the membrane part of Bcr transporter, namely protein BcrC. A fused protein, consisting of ATP-binding protein BcrA and membrane component BcrC was constructed. It resembled a half molecule of P glycoprotein and was capable of providing a significant degree of antibiotic resistance and detergent sensitivity. PMID- 10867245 TI - Studies on the cyclosporin A loaded stearic acid nanoparticles. AB - Stearic acid nanoparticles were prepared in this study by melt-homogenization to investigate the possibility of them as a new kind of drug carrier system. Some physicochemical properties of stearic acid nanoparticles were studied and morphology examined by transmission electron microscope. Cyclosporin A as a model drug was then encapsulated into stearic acid nanoparticles. Following the establishment of high performance liquid chromatography assay for cyclosporin A analysis in stearic acid nanoparticles or blood samples, the encapsulation ratio of cyclosporin A to stearic acid nanoparticles was estimated and pharmacokinetics as well as bioavailability of cyclosporin A stearic acid nanoparticles after oral administration to Wistar rats were studied, using the Sandimmun Neoral(R) (an available microemulsion system of cyclosporin A) as a reference. The mean diameter of cyclosporin A stearic acid nanoparticles was 316.1 nm, while the encapsulation ratio of cyclosporin A to stearic acid nanoparticles reached to 88.36%. It was demonstrated by IR spectra and differential scanning calorimetry that there was no chemical reaction occurred between the cyclosporin A and stearic acid. The relative bioavailability of cyclosporin A stearic acid nanoparticles over reference was nearly 80%, and the time to reach maximum concentration (T(max)) of cyclosporin A after oral administration of cyclosporin A stearic acid nanoparticles was delayed significantly than the reference, suggesting an obvious sustained release effect. The stearic acid nanoparticles might be a very potential drug carrier. PMID- 10867246 TI - Structure and hydration properties of hydroxypropyl methylcellulose matrices containing naproxen and naproxen sodium. AB - The present study was conducted to obtain a deeper insight into the mechanism of drug release from HPMC matrices. The microstructure, mobility, internal pH and the state of water within the gel layer of hydrated HPMC matrices (having different molecular weights) containing naproxen sodium (NS) and naproxen (N) were studied using Electron Paramagnetic Resonance (EPR), Nuclear Magnetic Resonance (NMR) and Differential Scanning Calorimetry (DSC) techniques. The study show that matrices composed of various viscosity grades of HPMC are characterized by similar microviscosity values in spite of the difference in their molecular weight. The NMR and DSC results led to the conclusion that higher molecular weights of HPMC are characterized by higher water absorption capacity and higher swelling. Analysis of non-freezable water in HPMC(K4M)-NS system revealed that addition of NS to solution increased the fraction of water bound to K4M+NS compared with the equivalent solutions without NS. The results suggest that the drug is participating in the crystallization of water and leads to the formation of a three dimensional network structure that decreases the freedom of water in K4M+NS samples. Calculation of the number of hydration shells showed that up to 2.2 layers are involved in HPMC-NS hydration compared to 1.5 layers for HPMC gel without NS. This was explained based on the different water ordering in the gel induced by NS as results of its absorption to polymer surface. Microviscosity values measured by EPR for K4M/N and K4M/NS hydrated matrices were found to be higher for K4M/N matrices, especially at initial stage of hydration. Mobile compartment calculations showed lower values for K4M/N compared with K4M/NS matrices. pH measurements by EPR revealed that incorporation of N to HPMC matrix led to lower internal pH value inside the hydrated tablet compared with NS. This behavior led to lower solubility of N which dictates its surface erosion mechanism, compared with NS matrix that was characterized by higher internal pH value and higher drug solubility. These properties of HPMC/NS increased chain hydration and stability, and led to drug release by the diffusion mechanism. PMID- 10867247 TI - Stability of some novel thymidine, 5-bromo-2'-deoxyuridine and 3'-azido-2'-3' dideoxythymidine analogues. AB - In the search of new prodrugs effective against herpes simplex virus series of thymidine, 5-bromo-2'-deoxyuridine esters with amino acid and peptide chains and 3'-azido-2',3'-dideoxythymidine derivatives have been synthesized and evaluated for antiviral activity. The chemical stability of some of them containing different residues was studied at pH 1 and 7.4 and temperature of 37 degrees C. An HPLC method was developed for quantification of the unchanged ester concentration. It was proved that esters with simple aliphatic straight side chain (containing alanyl-, glycyl-, or glycyl-glycyl-glycyl-residues) are relatively stable both at acidic and neutral media, 37 degrees C. Some of them undergo negligible hydrolysis with half lifes ranging between 6 and 23 h. In contrast, more complex esters with branched side chain (valyl-), with phenyl residue (phenylalanyl-), as well as containing thiazol ring are rather unstable especially at acidic conditions and undergo rapid hydrolysis resulting in the respective chemical precursor. The stability of the former group esters outlines them as suitable candidates for prodrugs: with higher lipophilicity facilitating po absorption, satisfying chemical stability and possibility to release the active moiety following enzymatic hydrolysis. PMID- 10867248 TI - Stability of thioTEPA and its metabolites, TEPA, monochloroTEPA and thioTEPA mercapturate, in plasma and urine. AB - The degradation of N,N',N"-triethylenethiophosphoramide (thioTEPA) and its metabolites N,N',N"-triethylenephosphoramide (TEPA), N, N'-diethylene,N"-2 chloroethylphosphoramide (monochloroTEPA) and thioTEPA-mercapturate in plasma and urine has been investigated. ThioTEPA, TEPA and monochloroTEPA were analyzed using a gas chromatographic (GC) system with selective nitrogen/phosphorous detection; thioTEPA-mercapturate was analyzed on a liquid chromatography-mass spectrometric (LC-MS) system. The influences of pH and temperature on the stability of thioTEPA and its metabolites were studied. An increase in degradation rate was observed with decreasing pH as measured for all studied metabolites. In urine the rate of degradation at 37 degrees C was approximately 2.5+/-1 times higher than at 22 degrees C. At 37 degrees C thioTEPA and TEPA were more stable in plasma than in urine, with half lives ranging from 9-20 h for urine and 13-34 h for plasma at pH 6. Mono- and dichloro derivatives of thioTEPA were formed in urine and the monochloro derivative was found in plasma. Degradation of TEPA in plasma and urine resulted in the formation of monochloroTEPA. During the degradation of TEPA in plasma also the methoxy derivative of TEPA was formed as a consequence of the applied procedure. The monochloro derivative of thioTEPA-mercapturate was formed in urine, whereas for monochloroTEPA no degradation products could be detected. PMID- 10867249 TI - An in vivo investigation of the rabbit skin responses to transdermal iontophoresis. AB - To optimize the benefits of transdermal iontophoresis, it is necessary to develop a suitable animal model that would allow for extensive assessments of the biological effects associated with electro-transport. Rabbit skin responses to iontophoresis treatments were evaluated by visual scoring and by non-invasive bioengineering parameters and compared with available human data. In the current density range 0.1-1.0 mA/cm(2) applied for 1 h using 0.9% w/v NaCl and 0.5 mA/cm(2) for up to 4 h, no significant irritation was observed. 2 mA/cm(2) applied through an area of 1 cm(2) for 1 h resulted in slight erythema at both active electrode sites but without significant changes in transepidermal water loss (TEWL) and laser Doppler velocimetry (LDV). A value of 4 mA/cm(2) under similar conditions caused moderate erythema at the anode and cathode with TEWL and LDV being significantly elevated at both sites; 1 mA/cm(2) current applied for 4 h, caused moderate erythema at both anode and cathode; and 1 mA/cm(2) applied for 1 h caused no irritation when the area of exposure was increased from 1 to 4.5 cm(2). When significant irritation and barrier impairment occurred, the erythema was resolved within 24 h with barrier recovery complete 3-5 days post treatment. Rabbit skin thus shows promise as an acceptable model for iontophoresis experiments. PMID- 10867250 TI - Characterization of a diltiazem-lambda carrageenan complex. AB - In the present paper the interaction between lambda carrageenan, a natural sulphated polysaccharide, and diltiazem HCl, a Ca channel blocking agent, was studied. Dialysis equilibria were performed to quantify the binding capacity of lambda carrageenan for diltiazem. The relevance of the interaction to hydrophilic matrix systems was confirmed: a relationship was found between the binding capacity and the release profiles of matrix tablets containing a fixed amount of drug and different percentages of lambda carrageenan. Dialysis equilibria in buffered media showed that the interaction is quite insensitive to the pH of the medium (in the range 1.8-6.8), while it is reduced by increasing ionic strength; this behaviour is in line with the importance of ionic bonds in diltiazem carrageenan interaction. On the basis of the calculated binding capacity, the complex was prepared, dried and milled. A preliminary characterization of the diltiazem-carrageenan complex in the solid state was effected by means of X-ray and DSC analysis. The amount of drug going into solution from the complex was not significantly affected by the pH of the medium (in the range 1.8-6.8), while it was increased by increasing ionic strength. PMID- 10867251 TI - Relationship between polysorbate 80 solubilization descriptors and octanol-water partition coefficients of drugs. AB - The molar solubilization capacities (kappa) and the molar micelle-water partition coefficients (K(M)(N)) in Polysorbate 80 of several drugs (including barbiturates, steroids, and benzoic acid derivatives) are related to their log octanol-water partition coefficients (logP). Both kappa and K(M)(N) values were calculated from solubility versus Polysorbate 80 concentration profiles, which were either experimentally determined or obtained from the literature. There is a linear relationship between logP of the tested compounds and the logarithm of the molar micelle-water partition coefficient (logK(M)(N)). On the other hand molar solubilization capacities are nearly independent of logP. It is shown that the ability of Polysorbate 80 to solubilize a drug can be predicted from its logP value. PMID- 10867252 TI - In vitro evaluation of sol-gel processed spray dried silica gel microspheres as carrier in controlled drug delivery. AB - The objective of this study was to evaluate sol-gel-derived spray dried silica gel microspheres as carrier material for dexmedetomidine HCl and toremifene citrate. The drug was dissolved in sol-gel processed silica sol before spray drying with Buchi laboratory scale equipment. Microspheres with a low specific surface area were spherical by shape with a smooth surface without pores on the external surface. The particle size distribution was quite narrow. The in vitro release of toremifene citrate and dexmedetomidine HCl showed a dose-dependent burst followed by a slower release phase, that was proportional to the drug concentration in the concentration range between 3.9 and 15.4 wt.%. The release period for toremifene citrate was approximately 10 days and for dexmedetomidine HCl between 7 and 50 days depending on drug concentration. Spray drying is a promising way to produce spherical silica gel particles with a narrow particle size range for controlled delivery of toremifene citrate and dexmedetomidine HCl. PMID- 10867253 TI - Clonazepam release from core-shell type nanoparticles of poly(epsilon caprolactone)/poly(ethylene glycol)/poly(epsilon-caprolactone) triblock copolymers. AB - The triblock copolymer based on poly(epsilon-caprolactone) (PCL) as hydrophobic part and poly(ethylene glycol) (PEG) as hydrophilic one was synthesized and characterized. Core-shell type nanoparticles of poly(epsilon caprolactone)/poly(ethylene glycol)/poly(epsilon-caprolactone) (CEC) block copolymer were prepared by a dialysis technique. According to the amphiphilic characters, CEC block copolymer can self-associate at certain concentration and their critical association concentration (CAC) was determined by fluorescence probe technique. CAC value of the CEC-2 block copolymer was evaluated as 0.0030 g/l. CAC values of CEC block copolymer decreased with the increase of PCL chain length, i.e. the shorter the PCL chain length, the higher the CAC values. From the observation of transmission electron microscopy (TEM), the morphologies of CEC-2 core-shell type nanoparticles were spherical shapes. Particle size of CEC-2 nanoparticles was 32.3+/-17.3 nm as a monomodal and narrow distribution. Particle size, drug loading, and drug release rate of CEC-2 nanoparticles were changed by the initial solvents and the molecular weight of CEC. The degradation behavior of CEC-2 nanoparticles was observed by 1H NMR spectroscopy. It was suggested that clonazepam (CNZ) release kinetics were dominantly governed by diffusion mechanism. PMID- 10867254 TI - pH, pK(a) and dermal delivery. AB - The effect of pH on the permeation of ibuprofen and lignocaine through human skin has been modelled using a modification to the equation derived by Potts and Guy, which is normally applied to unionized entities. The results show that permeation is related to the distribution coefficient. The physicochemical properties have been predicted ab initio using commercially available software and compared to literature values. The approach is successful and shows that there is significant permeation of the ionized drugs through a lipophilic pathway, possibly as a result of ion pairing. Since the aqueous solubility of the ionized material is significantly higher than the unionized, the maximum flux through the skin may occur at a pH where ionization is high. Optimum topical or transdermal formulations may not therefore be for the free acid or free base. PMID- 10867255 TI - Inhibition of Ostwald ripening in local anesthetic emulsions by using hydrophobic excipients in the disperse phase. AB - The stability of submicron emulsions of different local anesthetic/analgesic substances was investigated in the presence and absence of different hydrophobic excipients (ripening inhibitors). Ostwald ripening was believed to be the underlying mechanism for the instability of these emulsions. In the absence of ripening inhibitors, the mean droplet size of the emulsions increased from 100 nm to about 4-5 microm within an hour of manufacture. The addition of a small amount of a second component of lower solubility to the disperse phase decreased the rate of Ostwald ripening, producing good stability of the emulsions. The efficiency of the ripening inhibitors was directly proportional to their solubility in the disperse phase, i.e. the water. The lower the solubility, the more effective the stabilization of the emulsions. The experimentally observed rates of increase in droplet size in the emulsions were closely correlated with those predicted according to the Liftshitz-Slezov-Wagner (LSW) theory. PMID- 10867256 TI - TR146 cells grown on filters as a model of human buccal epithelium: V. Enzyme activity of the TR146 cell culture model, human buccal epithelium and porcine buccal epithelium, and permeability of leu-enkephalin. AB - The objective of the present study was to characterise the TR146 cell culture model as an in vitro model of human buccal mucosa with respect to the enzyme activity in the tissues. For this purpose, the contents of aminopeptidase, carboxypeptidase and esterase in homogenate supernatants of the TR146 cell culture model, and human and porcine buccal epithelium were compared. The esterase activity in the intact cell culture model and in the porcine buccal mucosa was compared. Further, the TR146 cell culture model was used to study the permeability rate and metabolism of leu-enkephalin. The activity of the three enzymes in the TR146 homogenate supernatants was in the same range as the activity in homogenate supernatants of human buccal epithelium. In the TR146 cell culture model, the activity of aminopeptidase (13.70+/-2.10 nmol/min per mg protein) was approx. four times the activity of carboxypeptidase (3.73+/-0.53 nmol/min per mg protein), whereas the level of esterase activity was significantly higher (223.39+/-69.82 nmol/min per mg protein). In the TR146 cell culture model, the apical esterase activity was found significantly higher than the basal activity, and found comparable to the porcine buccal mucosa. However, the esterase activity on the serosal side of the porcine buccal mucosa was higher than in the TR146 cell culture model. Approx. 1.5% of leu-enkephalin permeated the TR146 cell layers within 5 h (P(app) 7.38+/-0.83x10(-7) cm/s) and approx. 77% of intact peptide was still present in the donor phase after 5 h. The present study suggests that the TR146 cell culture model is a valuable in vitro model for permeability and metabolism studies with enzymatically labile drugs, such as leu enkephalin, intended for buccal drug delivery. PMID- 10867257 TI - An atomic force microscopy investigation of bioadhesive polymer adsorption onto human buccal cells. AB - Atomic force microscopy (AFM) was used to examine the buccal cell surface in order to image the presence of adsorbed bioadhesive polymers identified from previous work. Isotonic saline solution (5 ml) containing either polycarbophil (pH 7.6), chitosan (pH 4.5) or hydroxypropyl methylcellulose (pH 7.6) (0.5% w/v) was exposed to freshly collected buccal cells (ca. 48x10(4) cells/test) for 15 min at 30 degrees C. The cells were then rinsed with a small volume of double distilled water, allowed to air-dry on a freshy cleaved mica surface and imaged using contact mode AFM. Untreated cells showed relatively smooth surface characteristics, with many small 'crater-like' pits and indentations spread over cell surfaces. Cells that had been treated with all the investigated polymers appeared to have lost the crater and indentation characteristic and gained a higher surface roughness. These results suggest that polymer chains had adsorbed onto the cell surfaces. Quantitative image analysis of cell topography showed significant increases (P<0.05) in arithmetic roughness average (R(a)) for all the investigated polymer treated cells surfaces with respect to untreated control specimens. The changes in surface topography indicate the presence of adsorbed polymer, confirming previous work. This study demonstrates the suitability of AFM as a powerful and sensitive technique for detecting and imaging bioadhesive polymers present on mucosal cell surfaces. PMID- 10867258 TI - The physico-chemical properties of salmeterol and fluticasone propionate in different solvent environments. AB - The physico-chemical properties of two anti-asthmatic drugs, salmeterol xinafoate and fluticasone propionate, have been studied in both aqueous and non-aqueous solvent environments. Ultraviolet-visible (UV-Vis) spectroscopy, fluorescence spectroscopy and electrospray ionisation mass spectrometry (ESI-MS) have been used to characterise the interaction of the drugs in 70:30 (v/v) methanol/water solutions. First derivative UV-Vis spectra measurements indicate that an interaction takes place between the two drugs in a binary solvent system. Fluorescence studies indicate that an increase in the concentration of fluticasone propionate results in a decrease in the fluorescence signal of the salmeterol for mixed solutions of the drugs. Analysis of a mixture of the two drug solutions using mass spectrometry also shows evidence of salmeterol fluticasone propionate interaction and dimer formation with respect to both the salmeterol and the fluticasone propionate. Model metered dose inhalers (MDI) of both individual samples and mixtures of the drugs were formulated as suspensions in solvent CFC-113. The extent of deposition onto different inhaler components, such as the aluminium alloy canister, Teflon coated canister and the metering valve was evaluated by high-performance liquid chromatography (HPLC) of the methanol/water washings of the deposited drug(s). Changing the nature of the surface properties of the container resulted in a significant difference in the extent of deposition. The deposition of the individual drugs was found to increase as the dispersion concentration of the drug increases. However, the formulation based on a combination of the two drugs was found to show different deposition behaviour compared to the individual drug formulations. The deposition of the drugs, onto the aluminium alloy canister and the metering valve, decreases as the combined dispersion concentration of the two drug increases. PMID- 10867259 TI - Enthalpies of solution of ampicillin, amoxycillin and their binary mixtures at 310.15 K. AB - Enthalpies of solutions of ampicillin, amoxycillin and their binary mixtures have been determined at pH 2, 5, and 7 using C-80 calorimeter. The systems showed endothermic behaviour; molar enthalpies of solutions of ampicillin were determined to be 13.32, 15.89 and 23.21 kJ mol(-1) and amoxycillin were 16.32, 18.45 and 26. 25 kJ mol(-1) at pH 2, 5, and 7, respectively. The excess molar enthalpies of solution have also been calculated to find any interaction between these two drugs. PMID- 10867260 TI - The effect of processing variables on the physical characteristics of non-ionic surfactant vesicles (niosomes) formed from a hexadecyl diglycerol ether. AB - Niosomes are vesicles formed by self-assembly of non-ionic surfactants. In this investigation, the effects of processing variables, particularly temperature and sonication, on the physical characteristics and phase transitional behaviour of two niosomal systems based on a hexadecyl diglycerol ether (C(16)G(2)) have been studied. Systems containing C(16)G(2), cholesterol and poly-24-oxyethylene cholesteryl ether (Solulan C24) in the molar ratios 91:0:9 and 49:49:2 were prepared by aqueous dispersion of films, followed by examination of 5(6) carboxyfluorescein entrapment, particle size and morphology. The thermal behaviour was examined using high sensitivity differential scanning calorimetry (HSDSC) and hot stage microscopy, while the effects of sonication were studied in terms of size and morphology, both immediately after preparation and on storing for 1 h at room temperature and 60 degrees C. Polyhedral niosomes were formed from systems containing C(16)G(2) and Solulan C24 alone, while cholesterol containing systems formed spherical vesicles mixed with tubular structures; the polyhedral systems were found to have a larger particle size and higher CF entrapment efficiency. HSDSC studies showed the polyhedral systems to exhibit an endotherm at 45.4 degrees C and a corresponding exotherm at 39.1 degrees C on cooling which were ascribed to a membrane phase transition; no equivalent transition was observed for the cholesterol containing systems. Hot stage microscopy showed the polyhedral vesicles to convert to spherical structures at approximately 48 degrees C, while on cooling the spherical vesicles split into smaller structures and reverted to the polyhedral shape at approximately 49 degrees C. Sonication resulted in the polyhedral vesicles forming spherical structures which underwent a particle size increase on storage at room temperature but not at 60 degrees C. The study suggests that the polyhedral vesicles undergo a reversible transition to spherical vesicles on heating or sonication and that this morphological change may be associated with a membrane phase transition. PMID- 10867261 TI - Ex vivo and in situ PLGA microspheres uptake by pig ileal Peyer's patch segment. AB - We investigated the ability of pig ileal Peyer's patch segments to transport intestinal poly (D,L-lactide-co-glycolide) microspheres (PLGA MS) from intestinal lumen across the mucosae using in situ and ex vivo segments with confocal laser scanning microscopy (CLSM) and transmission electronic microscopy (TEM). From a global aspect, CLSM suggested that PLGA MS were translocated by M cells labelled with a FITC-conjugated anti-cytokeratin peptide 18, and transported through the follicle-associated epithelium (FAE) in the dome area in both types of experiments. At the ultrastructural level, TEM showed the traffic of PLGA MS throughout M cells, their transport into the basolateral invaginations of the M cells and their subsequent migration into the dome area and the follicular area in contact with macrophages and lymphatic vessels. Although in situ experiments allowed following the migration of PLGA MS until mesenteric lymph nodes, an ex vivo model could be used as a useful tool to study the targeting ability of PLGA MS formulations to the gut-associated lymphoid tissue (GALT). PMID- 10867262 TI - The effect of pore formers on the controlled release of cefadroxil from a polyurethane matrix. AB - The effect of various pore formers on the controlled release of an antibacterial agent from a polymeric device was examined in order to develop a novel biomaterial that prevents bacterial adhesion and growth on its surface. Cefadroxil was chosen as the model antibiotic and was incorporated into a polyurethane matrix by the solvent-casting method. Polyethylene glycol (PEG) 1450, D-mannitol, or bovine serum albumin (BSA) was used as a pore former. The amount of cefadroxil released from various formulations at 37 degrees C was measured by HPLC. The morphological change of matrices before and after release studies was investigated by scanning electron microscopy (SEM). The duration of antimicrobial activities of matrices against Escherichia coli and Bacillus subtilis was evaluated by measuring the diameters of the inhibition zone. Changing the weight fraction and particle size of the pore formers/drug mixtures could control the release of cefadroxil from the matrix. The release rate of cefadroxil increased as the loading dose of the pore former increased (15<20<25%). Cefadroxil released from these devices exhibited antibacterial activity for 5-6 days. These results imply that an antibiotic-loaded polymeric device that could prevent bacterial infection on its surface can be formulated using appropriate pore formers. PMID- 10867263 TI - Spectroscopic investigation of the molecular state of nystatin encapsulated in liposomes. AB - The stability and spectral properties of nystatin-encapsulating liposomes, composed of various combinations of dipalmitoyl phosphatidylcholine (DPPC), cholesterol (CH) and distearoyl-N-(monomethoxy poly(ethylene glycol)succinyl) phosphatidylethanolamine (DSPE-PEG), were studied in order to elucidate the molecular state and localization of nystatin encapsulated in liposomes. Localization of nystatin at the surface region of the liposomal membrane was investigated by PEG/dextran two-phase partition and measurement of the fluorescence quenching of nystatin by p-xylene-bis-pyridinium bromide (DPX). In DPPC/DSPE-PEG liposomes and DPPC/CH/DSPE-PEG liposomes, containing 151 and 160 mcg nystatin per mg lipid, respectively, nystatin appeared to be present at the surface region of the liposomal membranes. Self-quenching of nystatin fluorescence was observed in DPPC/CH and DPPC/CH/DSPE-PEG liposomes even at low encapsulated amounts, suggesting the localization of nystatin in CH-incorporating membranes. In CH-free liposomes, nystatin molecules were at first delocalized in the membranes and then self-associated at a higher level of encapsulation. Absorption and circular dichroism (CD) spectra were also measured to examine the monomeric and aggregated states of nystatin in liposomes. High encapsulation efficacy was observed in DPPC and DPPC/DSPE-PEG liposomes, but the highest stability and retention of nystatin in liposomes were observed in DPPC/CH/DSPE PEG liposomes, evaluated in terms of the nystatin and calcein release from nystatin-encapsulating liposomes in vitro. From the results, possible encapsulation mechanisms of nystatin in liposomes narrowed down to the following three points; interaction with lipid membrane, adsorption on the liposomal surface and complex formation with DSPE-PEG. PMID- 10867264 TI - A novel approach to prepare tripolyphosphate/chitosan complex beads for controlled release drug delivery. AB - A novel approach was developed to improve the mechanical strength of tripolyphosphate (TPP)/chitosan beads prepared under coagulation condition at 4 degrees C in the presence of gelatin. Cross-sectional analysis indicated that the beads had a homogeneous crosslinked structure, as a result the beads were strengthened greatly (the mechanical strength increased more than ten times). Furthermore sodium alginate (a polyanion) can interact with cationic chitosan on the surface of these TPP/chitosan beads to form polyelectrolyte complex film for the improvement of the drug sustained release performances. The loading efficiency of model drugs (brilliant blue and FITC-dextran) in these beads was very high (more than 90%). Crosslinking time, TPP solution pH and other preparation factors had an effect on the drug release performance of beads. The release period of brilliant blue (a poor water soluble dye) was more than 2 months at a fairly constant rate in 0.9% NaCl, 10 mM PBS pH 7.4. However, for FITC-dextran (a water soluble polysaccharide) only 1-2 days in the same conditions. It seems that TPP/chitosan bead prepared by the novel method is a promising formulation for drug delivery. PMID- 10867265 TI - Pharmacokinetics and biodisposition of fluorescein-labeled arabinogalactan in rats. AB - Fluorescein-labeled arabinogalactan (FA) was prepared by the reaction with FITC in methyl sulphoxide according to the method of deBelder and Granath. A systemic kinetic analysis of FA in rats was carried out by using a specific high performance size-exclusion chromatography. Intravenously administered FA was rapidly eliminated from the blood circulation followed by an appreciable distribution to the liver and kidney. FA was accumulated in these organs over a long period whereas negligible levels of FA were detected in the other organs. A marked dose-dependency was seen in the hepatic uptake of FA which was markedly reduced by coinjected asialofetuin whereas the renal uptake of FA was not altered. Measurement of the hepatocellular localization demonstrated the overwhelming distribution of FA in the parenchymal liver cell fraction. Furthermore, the microscopic examination revealed FA that was effectively endocytosed by the parenchymal liver cells. These results suggested that FA which is bound to the asialoglycoprotein receptor with a high affinity is subsequently internalized to the hepatocyte via receptor-mediated endocytosis. FA was partially activated by periodate oxidation in order to acquire aldehyde groups to which guest molecules can be bound. A 12.5% oxidized arabinogalactan keeping a hepatocellular targetability showed a good conjugating reactivity to guest molecules via Schiff-base formation or by reductive amination. It was suggested that arabinogalactan can serve as a potential carrier for the delivery of enzymes and drugs to the parenchymal liver cells via the asialoglycoprotein receptor. PMID- 10867266 TI - Coadsorption of the sodium salts of two steroid molecules at a silica/interface as induced by the adsorption of a cationic surfactant. AB - The incorporation of two ionic steroids, namely the sodium salts of hydrocortisone 21-hemisuccinate (HNa) and prednisolone 21-succinate (PNa), at a silica/water interface in the presence of adsorbed cetyltrimethylammonium bromide has been investigated first at a constant pH value. It is shown that this coadsorption effect is qualitatively similar to the adsolubilization effect which is described for neutral molecules. The adsorption of the cationic surfactant induces the coadsorption of the anionic drug molecules although the silica surface is negatively charged. At surfactant equilibrium concentration above the critical micelle concentration the drug molecules are distributed between the adsorbed aggregates and the free micelles. At larger surfactant concentration, the drugs may be completely depleted from the silica/water interface. Based upon Langmuir-type isotherms, the equilibrium constants of the drug molecules with the adsorbed aggregates and the free micelles are calculated. The constants are about three times larger for the former than for the latter aggregates. The signification of such results is discussed. The coadsorption of HNa at low surfactant surface coverage was also investigated in the pH interval between 3 and 9. HNa is strongly coadsorbed at lower pH onto the silica surface. The coadsorption goes through a maximum at a pH value which may be considered as equal to the apparent pK of the drug and decreases to zero at higher pH values. A pK value equal to 4.2 is proposed for HNa. This behaviour is interpreted as the result of the interplay of the drug dissociation and that of the surface silanol groups upon the change of pH. PMID- 10867267 TI - Relations between crystallisation conditions and micromeritic properties of ibuprofen. AB - The effects of solvent, cooling rate and type of methacrylic polymer (Eudragit(R)) on the micromeritic properties (size, elongation ratio, roundness and fullness ratio), the temperature change in the crystallisation liquid, the crystal yield and the extent of agglomeration of ibuprofen crystals have been compared. Twenty batches of crystals were prepared and Latin square experimental design was applied with four levels for each factor. It was found that crystal yield (Y) is related to the extrapolated point of maximum rate of temperature deviation (T(d)) with a logarithmic-type equation [Y=34.45lnT(d)-28.00] and to the area under the curve of temperature-deviation versus time (AUC) with a polynomial equation including cooling rate [Y=19.95AUC-1.57AUC/CR+63.00]. Crystal size is affected by the cooling rate and analysis of variance (ANOVA) showed that elongation ratio and fullness ratio of single crystals (P=0.05 and 0.05), as well as roundness and fullness ratio of agglomerates (P=0.05 and 0.1), are affected by the solvent. Post hoc statistical analysis of the solvent effects on the shape of crystals and agglomerates (Tukey's HSD multiple pairwise comparison test of means) indicated that their significance lies in the different polarity and may be attributed to interactions of solvent (acetone) with the growing crystal faces. Extent of crystal agglomeration was found to be inversely proportional to the ratio of elongation ratio/circle equivalent diameter of the single crystals. PMID- 10867268 TI - Transition to CFC-free metered dose inhalers--into the new millennium. AB - Metered dose inhalers (MDIs) are the most popular vehicle for drug delivery into the lungs and some 500 million are manufactured each year. All MDIs marketed prior to 1995 contained chlorofluorocarbons (CFC) as a propellant. These are implicated in the depletion of stratospheric ozone and, except for specific exemptions, their production has been banned since 1996 under the terms of the Montreal Protocol. Hydrofluoroalkanes have been identified as suitable alternatives for MDI propellants but their physico-chemical properties differ significantly from CFCs and an extensive redevelopment and testing programme has been required to demonstrate the safety, quality and efficacy of HFA containing MDIs. Hydrofluoroalkanes contribute to global warming but the benefit to human health through continued MDI availability currently outweighs the environmental concern. Several HFA-MDIs have reached the market and the transition to replace existing CFC-MDIs is now underway. PMID- 10867269 TI - Gamma-scintigraphic study of the gastrointestinal transit and in vivo dissolution of a controlled release diclofenac sodium formulation in xanthan gum matrices. AB - A xanthan gum matrix controlled release tablet formulation containing diclofenac sodium was evaluated in vitro and was found to release the drug at a uniform rate. The gastrointestinal transit behaviour of the formulation as determined by gamma scintigraphy, using healthy male volunteers under fasted and fed conditions, indicated that gastric emptying was delayed with food intake. In contrast, the small intestinal transit remained practically unchanged under both food statuses. Therefore, the delay in caecal arrival observed in the fed state can be attributed to the delay in gastric emptying. Rate of diclofenac sodium absorption was generally higher in the fed state compared to the fasted state, however the total amount absorbed under both food statuses remained practically the same. The rate of in vivo dissolution of the drug in the fed state was faster compared to that in the fasted state. Thus, at the time of caecal arrival, in vivo dissolution was complete in the fed state, unlike in the fasted state, where almost 60% of the drug was delivered to the colon. PMID- 10867270 TI - On iontophoretic delivery enhancement: ionization and transport properties of lidocaine hydrochloride in aqueous propylene glycol. AB - The ionization and transport properties of lidocaine hydrochloride (LidHCl) in aqueous propylene glycol (PG) containing 20% PG by weight was studied by means of electrical precision conductometry. For drug concentrations exceeding about 1.7 mM a slight formation of LidH(+)Cl(-) ion-pairs is indicated; ion-pair association constant, K(p)=1.73 (molar scale). A two variable analysis of the experimental data yielded K(a)=1.5x10(-8) for the acid dissociation constant of LidH(+), i.e. pK(a)=7.82, and the limiting ionic conductivity, lambda(o)(LidH(+))=21.73 cm(2) S mol(-1). To enable evaluation of single ion conductivities the proton transport number of HCl in the present solvent mixture was determined using the moving boundary method. PMID- 10867271 TI - A review of evidence in support of a role for 5-HT in the perception of tinnitus. AB - Tinnitus is a debilitating condition from which some 0.5-1% of the population of the Western world suffer sufficiently badly as to interfere with their normal working and leisure life. There is no satisfactory treatment at the present time and the uncertainty surrounding the mechanism of its generation makes it difficult to devise an effective cure. After much debate, the consensus of opinion amongst researchers regarding its site of origin is that it is primarily a central nervous system pathology although there certainly exists a class of patients whose tinnitus is peripherally based. In this paper, we provide some speculative ideas on how an initial auditory insult can be translated into central neurological substrates that represent tinnitus. Plastic changes arising from sensory deprivation trigger a change in synaptology and neurotransmission with a consequent change in receptor configuration. From neuroanatomical considerations and analogies with other clinical conditions, we postulate the involvement of serotonin (5-HT) in these plastic changes and consider the evidence available from the use of serotonergic drugs in different conditions. A possible relationship between 5-HT and lidocaine is also discussed. PMID- 10867272 TI - Kinocilia heights on utricular hair cells. AB - Kinocilium height is a critical determinant of any hair cell's response to head movement, but accurate measurements of kinocilia heights have been difficult to achieve. We have developed a method for measuring kinocilia heights that combines immunochemical staining with three-dimensional morphometry, and we have used this method to measure kinocilia in the utricle of a turtle, Pseudemys scripta. Our results suggest that kinocilium height varies with position on the utricular epithelium and that kinocilia in the striola are significantly shorter than kinocilia in other regions of the utricle. PMID- 10867273 TI - The effects of maturation and stimulus parameters on the optimal f(2)/f(1) ratio of the 2f(1)-f(2) distortion product otoacoustic emission in neonates(1). AB - Distortion product otoacoustic emission (DPOAE) measurements are becoming popular in the clinical realm because they have been shown to reflect cochlear function. The primary tones used to evoke the DPOAE are important in determining the amplitude of the emission recorded in the ear canal. This study examined the ratio of the primaries necessary to determine the maximum amplitude emission as a function of development, stimulus level and frequency. Optimum f(2)/f(1) ratios were measured utilizing the f(1)-sweep technique from 105 neonates between 30-42 weeks conceptional age (CA) and 40 adults. No significant difference for optimum ratio was shown between the neonatal and the adult groups. Primary tone frequency had a significant effect on optimum ratio for both neonates and adults. Low f(2) frequencies (<4 kHz) were associated with higher optimum ratios than high f(2) frequencies (>4 kHz). The adult group was used to investigate the effect of stimulus level on the optimum f(2)/f(1) ratio for f(2) frequencies from 1.7 to 10 kHz. Regression analysis showed significant differences across levels of the primaries at all frequencies except for f(2)=3.4 and 7.0 kHz. These differences in f(2)/f(1) ratio across stimulus frequency and level may be attributed to the change in the shape of the excitation profiles along the basilar membrane. PMID- 10867274 TI - The importance of phase data and model dimensionality to cochlear mechanics. AB - The vulnerability of the mammalian cochlear amplifier to surgical trauma hinders observations of its behaviour in vivo. This produces a greater need for realistic models to aid the interpretation of the experimental observations. The emphasis in most modelling studies has been to simulate the gain of the response of the basilar membrane. This paper argues that matching the phase behaviour of the response should be given at least equal importance. When it is, many of the models used to justify hypotheses regarding the operation of the cochlear amplifier cannot simulate the response even of the dead cochlea. This discrepancy is due to oversimplification of the mechanics of the cochlear fluids. It is argued that three-dimensional fluid behaviour should be regarded as a bare minimum in any quantitative description of cochlear mechanics. Furthermore, it is shown that a three-dimensional model is consistent with experimental data from a healthy cochlea only when the main effect of the cochlear amplifier is to inject mechanical energy into the basilar membrane. The injection of mechanical energy is fundamentally different to modifying the stiffness of the basilar membrane. This means that existing models which possess cochlear amplifiers that effect large changes on the stiffness of the basilar membrane may not be accurate representations of the real organ. PMID- 10867275 TI - Cochleotopic fos immunoreactivity in cochlea and cochlear nuclei evoked by bipolar cochlear electrical stimulation. AB - Fos-like immunoreactivity evoked by basal, second or apical turn bipolar intracochlear electrical stimulation was evaluated in the spiral ganglion and cochlear nuclei. At stimulation levels of six times the electrically evoked auditory brain stem response thresholds, immunoreactive neurons were observed at appropriate discrete cochleotopic regions relative to stimulation site. The number of neurons increased with stimulus level and closely correlated to wave I amplitude. At 10 times thresholds, some spread in fos-like immunoreactivity to adjacent cochlear turns was found. However, fos-like immunoreactivity at this high level of stimulation still clearly showed a differential distribution in density of expression. These results indicated that the restricted topographic distribution of activity evoked by high levels of electrical stimulation is initiated at first order primary neurons of the system. For the profoundly deaf with cochlear implants, this indicates that place (channel) information can be maintained in the spiral ganglion and central nervous system even at very high levels of electrical stimulation. Together with our previous studies, these results indicate that fos provides a marker which can be used for evaluation of extent and pattern of cellular activation throughout the central auditory pathways, including activation of auditory nerve cells. PMID- 10867276 TI - Frequency and temporal analysis of contralateral acoustic stimulation on evoked otoacoustic emissions in humans. AB - Previous studies have shown that the effect of contralateral acoustic stimulation (CAS) on ipsilateral evoked otoacoustic emissions (EOAE) depends somewhat upon the spectrum of the eliciting stimulus. The latency of the EOAE, however, is itself frequency-dependent. Consequently, two general ways of analyzing the effects of CAS may be considered: by frequency band or by temporal segment. In this study, we analyzed the effects of CAS both ways in the same subjects, essentially simultaneously. The frequency analysis of the EOAE derived from the wavelet transform (WT). The WT is known to provide a robust approach to the analysis of non-stationary signals and was anticipated to avoid possible time frequency confounds of the cochlear mechanical system. For comparison, a more basic analysis - using a temporal moving window - was employed. The results largely support earlier findings and confirm that in humans the greatest suppression of EOAEs by CAS is obtained for lower frequency and/or longer latency EOAE components. Despite expectations for the WT analysis, the more basic, temporal, analysis tended to yield the clearer results. PMID- 10867277 TI - Effect of calmodulin antagonists on the compound action potential of the cochlea. AB - This study aimed to evaluate the effect of calmodulin antagonists on the threshold of the compound action potential (CAP) and the functional recovery of the cochlea after transient ischemia. When trifluoperazine and W-7 were administered to albino guinea pigs with perilymphatic perfusion, these drugs did not significantly affect the CAP thresholds. Transient cochlear ischemia of 30 min duration was obtained via a skull base approach. Although trifluoperazine significantly ameliorated the post-ischemic CAP threshold shifts 4 h after the onset of reperfusion, 1 to 50 microM W-7 did not affect the CAP threshold shifts. These results suggest that the action antagonizing calmodulin has no effect on the CAP threshold, while the role that calmodulin plays in cochlear ischemia reperfusion injury still remains unclear. PMID- 10867278 TI - Lateral interactions account for the pattern of the hair cell array in the chick basilar papilla. AB - It has been suggested that lateral interactions set up the array of hair cells and supporting cells in the chick basilar papilla. The presence of a hair cell would inhibit adjacent cells from becoming hair cells, and promote the formation of supporting cells. Models of cell specification were tested, starting with a closely packed array of multipotent progenitor cells. Lateral interactions, in which emerging hair cells promoted a supporting cell phenotype in adjacent cells, and in which emerging supporting cells promoted a hair cell phenotype in adjacent cells, produced an array of cells similar to that observed experimentally in the distal and central parts of the basilar papilla. In these areas, the ratio of supporting cells to hair cells is very close to 2:1, each hair cell on average being surrounded by six supporting cells, and each supporting cell being surrounded by three hair cells and three supporting cells. Identical patterns of hair and supporting cells could be produced by models in which either of the lateral inhibitory factors was replaced by a diffusive factor, i.e. a factor which acts on all cells in the model irrespective of position. The agreement of the model with observed cell ratios supports the view that the fate of both hair cells and supporting cells in the chick basilar papilla is a product of cell interactions within the papilla. It is possible that one factor, that provides contact lateral inhibition and promotes the formation of supporting cells, is the Notch/Delta system. It is possible that the other factor is retinoic acid, a diffusive factor that promotes the formation of hair cells. PMID- 10867279 TI - Development and regression of cochlear blood vessels in fetal and newborn mice. AB - Various strains of mice have been used for hearing research, but there have been few reports regarding the cochlear vasculature in mice. In this study, the development of the cochlear vasculature was investigated in C57BL/6 mice from day 15 of gestation to day 15 after birth, and mature vessels were also observed in 3 month-old mice. Both India ink injection and the resin casting method were used. On gestational day 17, spiral vessels of the basilar membrane were developing and were elaborating communicating branches that ran toward the external wall and the spiral lamina. On day 18, the spiral vessels showed the largest diameter of all vessels in the cochlea, but these vessels subsequently regressed and finally disappeared by day 14 after birth. The external wall vessels formed a single layer capillary network at birth and subsequently divided into two layers, which became the vessels of the stria vascularis and the spiral ligament vessels. This process occurred progressively from the basal turn toward the apical turn between days 5 and 8 after birth. A general tendency for the cochlear vasculature to mature from the basal turn towards the apex was observed. PMID- 10867280 TI - Measurement of the mechanical compliance of the endolymphatic compartments in the guinea pig. AB - During injection of artificial endolymph into scala media of the guinea pig, fluid pressure was simultaneously measured in endolymph and perilymph with micropipettes. Pressure differences in the order of 10 Pa could reproducibly be measured upon injection of 2-4 microl of artificial endolymph with a rate of 50 nl/s. Injection of larger volumes damaged the endolymphatic system. From the results, values were derived for the compliances of the membranes surrounding scala media and the vestibular part of the endolymphatic system. The shape of the pressure-time curve during and between repetitive injections of fluid could well be described with a two-component model for the endolymphatic system, consisting of two compartments with compliant walls, connected through a flow resistance. With this model, a larger compliance was found for the second compartment (vestibular part of endolymphatic system) than for the first compartment, into which fluid was injected (scala media). PMID- 10867281 TI - Succinate dehydrogenase (SDH) activity in hair cells: a correlate for permanent threshold elevations. AB - Hair cell loss is often used as a histological correlate of hearing loss. However, the histological and the physiological data are not always well correlated. This paper investigates the use of succinate dehydrogenase (SDH) activity in the hair cells as a marker of cellular dysfunction and so the loss of auditory sensitivity. In our previous studies, potentiation of noise-induced auditory threshold elevation by carbon monoxide (CO) was observed [Chen and Fechter, 1999; Chen et al., 1999]. However, its histological basis is still unclear. In this study, rats were exposed to 100-dB octave-band noise (center frequency=13.6 kHz, 2 h) or to the combination of the noise and CO (1200 ppm). Threshold elevation of compound action potential (CAP) and cochlear histological changes were assessed 4 weeks after exposure. The noise alone caused CAP threshold elevations with little if any or without hair cell loss. However, the SDH activity in the hair cells decreased after the exposure. The SDH reduction, especially in the inner hair cells, was well related to the loss of auditory sensitivity. The combined exposure to noise and CO caused more severe CAP threshold elevation and SDH activity reduction than did the noise alone and it also caused significant outer hair cell loss. However, across all the test frequencies, neither the hair cell loss nor the SDH reduction alone had good correlation to the reduction of the auditory sensitivity. Under this situation, CAP threshold elevation seemed to follow OHC loss at high frequencies and to follow SDH reductions in the IHCs at low frequencies, where no hair cell loss occurred. PMID- 10867282 TI - Immunolocalization of calbindin D28k and calretinin in the dog cochlea during postnatal development. AB - The calbindin (CB) and the calretinin (CR) immunoreactivities were studied in the dog cochlea during its postnatal maturation from birth to the 33rd postnatal day. At birth, CB was expressed in the Kolliker's organ, in the immature inner (IHC) and outer hair cells (OHC), in neurons of the spiral ganglion, and in nerve fibers running in the basilar membrane of the apical turn. During the cochlear maturation, non-sensorineuronal structures, such as the Kolliker's organ, the rods of Corti, and the inner sulcus cells, displayed a transient CB-staining. In the adult-like dog cochlea, CB was found in the cytoplasm, the cuticular plate, and the stereocilia of the IHC and OHC. All the neurons of the spiral ganglion and some nerves fibers in the modulius were CB-positive. At birth, CR exhibited a neuronal distribution: about 75% of the spiral ganglion neurons, some nerve fibers in the modulius and nerve fibers running in the basilar membrane were CR labeled. During the postnatal maturation, a CR-immunostaining appeared around the IHC body and CR was expressed transiently in the OHC. In the adult-like dog cochlea, a CR-positive network surrounded the unlabeled IHC. The neuronal CR labeling remained unchanged from birth. PMID- 10867283 TI - Noise damage in the C57BL/CBA mouse cochlea. AB - The present study was designed to determine the response to noise of the auditory system of a genetically well-defined laboratory mouse in preparation for examining the effect of noise on mice with specific genetic mutations. The mice were C57BL/CBA F1 hybrids. Eight mice served as non-noise-exposed controls and 39 mice were exposed for 1-24 h to an octave band of noise with a center frequency of 2, 4 or 8 kHz and a sound pressure level of 100-120 dB. Auditory brainstem response thresholds were measured pre-exposure and several times post-exposure (i.e., 0-27 days) to determine the magnitude of the temporary threshold shift (TTS) and permanent threshold shift (PTS). After fixation by cardiac perfusion, the cochleas from each mouse were embedded in plastic, dissected into quarter turns of the cochlear duct and analyzed quantitatively. Immediately post exposure, all mice had sizable TTSs at the tested frequencies (i.e., 3-50 kHz). At this time, two mice were killed. Thresholds of the other 37 mice recovered somewhat in the first 4 days post-exposure. One mouse fully recovered from its TTS; 10 mice were left with PTSs at all frequencies; 26 mice recovered at some frequencies but not others. Most mice with PTSs for 30-50 kHz had focal losses of inner and outer hair cells in the basal 20% of the organ of Corti, often with degeneration of adjacent myelinated nerve fibers in the osseous spiral lamina. On the other hand, mice with PTSs for the lower frequencies (i.e., 3-20 kHz) had stereocilia disarray without significant hair cell losses in the second and first turns. Considerable variability was found in the magnitude of hair cell losses in those mice that received identical noise exposures, despite their genetic homogeneity. PMID- 10867284 TI - An anatomically based frequency-place map for the mouse cochlea. AB - An anatomically based frequency-place map was created for the mouse using C57BL/CBA F1 hybrids by matching noise-induced lesions in the organ of Corti with permanent hearing losses as determined by auditory brainstem response (ABR) thresholds. Twenty-six mice developed 'notched' ABR threshold shifts after exposure to an octave band of noise with a center frequency of 2 kHz at 120 dB SPL for 24 h, 4 kHz at 110 dB SPL for 4 h or 8 kHz at 100 dB SPL for 1 or 2 h. ABR thresholds were determined at several intervals post-exposure until thresholds stabilized (14-27 days). Once thresholds had stabilized, the mice were killed and their cochleas were prepared for phase-contrast microscopic examination as plastic-embedded flat preparations. Hair cell loss, stereocilia damage, and myelinated nerve fiber degeneration as a function of percentage distance from the cochlear apex were determined. Frequency-position matches could be made for 22 of the 26 mice by correlating areas of hair cell loss/stereocilia damage with permanent changes in ABR thresholds. These frequency-position data were fitted with the equation: % Distance from apex=56.6 log (f(Hz))-179.1; r(2)=0.810. This frequency-place function agrees well with Ehret's (1975) theoretical function based on critical bands and masked auditory thresholds. PMID- 10867285 TI - Inferior colliculus as candidate for pitch extraction: multiple support from statistics of bilateral spontaneous otoacoustic emissions. AB - The fibrodendritic laminae of the central nucleus of the inferior colliculus (ICC) constitute a frequency map in stacked sheets that are consistently related to the psychoacoustic critical bandwidth (CB) [Schreiner and Langner, 1997. Nature 388, 383-386]. The recently observed co-occurrence of the CB and the double CB (2CB) suggested an adaptation of the ICC frequency map to the extraction of the fundamental frequency f(0) [Braun, 1999. Hear. Res. 129, 71 82]. The present study examined a possible influence of this frequency map upon efferent signaling towards the cochlea. The f(0) distribution of 2890 monaural and 2604 binaural pairs of human spontaneous otoacoustic emissions was analyzed by three statistical methods and in each case showed non-random behavior in the CB-2CB range. Single results were (1) a bias of right ear f(0) (mode at 349 Hz) and left ear f(0) (mode at 262 Hz) towards different ranges of speech f(0) (P<0.02); (2) a bias of binaural, but not monaural, f(0) towards five of 12 semitone bins, C-G-D-A-E, representing the most frequent tones in music (P<0.003); (3) a bias of binaural, but not monaural, f(0) fine-distribution towards the exact pitch frequencies used in music, according to the international standard A4=440 Hz (P=0.03). The results support a model of lamina-based f(0) extraction in the ICC and suggest a specific colliculo-cochlear feedback for f(0) enhancement. PMID- 10867286 TI - An anatomically shaped incus prosthesis for reconstruction of the ossicular chain. AB - An anatomically shaped incus replica prosthesis has been designed to reconstruct the ossicular chain. A series of in vitro studies on human temporal bones evaluated the acoustic performance of this prosthesis and compared it with a Causse partial ossicular replacement prosthesis (PORP). Pure tones in the frequency range 0. 125-8 kHz stimulated the tympanic membrane at sound pressure levels of 80, 90 and 100 dB. Measurements of the stapes footplate velocity were made with a laser interferometer. The acoustic function of the ossicular chain reconstructed with the incus replica prosthesis was found to be within 10 dB of that of the original intact ossicular chain, when both the upper and lower joints of the implant were rigidly fixed in place. It was shown that a rigid mechanical contact between the ossicular prosthesis and ossicles is a prerequisite for effective sound transmission. The anatomically shaped incus prosthesis gave a 15 dB improvement on the PORP at frequencies below 1.5 kHz. PMID- 10867287 TI - Changes of the compound action potential (CAP) and the expression of inducible nitric oxide synthase (iNOS/NOS II) in the cochlea under the inflammatory condition(1). AB - In this study, the effect of endotoxin on the guinea pig cochlea has been examined electrophysiologically and immunohistochemically. Bacterial lipopolysaccharide (LPS, 5 mg/ml, 0.2 ml) was injected into the middle ear trans tympanically. The electrocochleograms were measured before, immediately upon, and 3, 6 and 12 h after the injection continuously with an electrode inserted into the facial canal. After each measurement, some of the animals were killed with an intracardiac perfusion of fixative, temporal bones were removed and were immunohistochemically examined for inducible nitric oxide synthase (iNOS/NOS II). On serial paraffin section, iNOS could be detected first after 3 h in the lateral wall, the supporting cells of the organ of Corti and in cells of the spiral ganglion and was observed up to 12 h. After the injection of LPS, the threshold of compound action potential became significantly worse after 12 h in the LPS group. These changes became evident first at higher frequency (8 kHz). These results suggest that iNOS-generated NO is involved in the cochlea dysfunction under inflammatory conditions. PMID- 10867288 TI - Isolation of cochlear inner hair cells. AB - In order to identify hair cell specific genes, it is essential to obtain isolated hair cells in quantity. While whole-cell recordings have been made from isolated inner hair cells (IHCs) from guinea pigs, detailed methods for obtaining a fairly large amount of isolated inner hair cells have not been published. Here we describe a protocol that can yield a fairly large amount of isolated gerbil IHCs. This technique can provide sufficient numbers of solitary IHCs for either electrophysiological studies of the cell's membrane properties or identifying genes related to IHC functions using techniques of molecular biology. PMID- 10867289 TI - The perception of coherent and non-coherent auditory objects: a signature in gamma frequency band. AB - The pertinence of gamma band activity in magnetoencephalographic and electroencephalographic recordings for the performance of a gestalt recognition process is a question at issue. We investigated the functional relevance of gamma band activity for the perception of auditory objects. An auditory experiment was performed as an analog to the Kanizsa experiment in the visual modality, comprising four different coherent and non-coherent stimuli. For the first time functional differences of evoked gamma band activity due to the perception of these stimuli were demonstrated by various methods (localization of sources, wavelet analysis and independent component analysis, ICA). Responses to coherent stimuli were found to have more features in common compared to non-coherent stimuli (e.g. closer located sources and smaller number of ICA components). The results point to the existence of a pitch processor in the auditory pathway. PMID- 10867290 TI - Scopolamine reduces sensitivity to auditory gaps in the rat, suggesting a cholinergic contribution to temporal acuity. AB - Prior research [Caine et al., 1981] suggested that scopolamine, a central cholinergic antagonist, may increase gap thresholds in young human listeners. If confirmed, an effect of scopolamine on gap detection might help to explain why both aged humans and aged laboratory animals have less sensitive temporal acuity on gap detection tests, as they may be presumed to have less effective cholinergic mechanisms. Here we measured the effect of scopolamine on gap detection in rats (n=8) using reflex modification audiometry, which depends on the fact that brief gaps in noise presented immediately prior to a loud noise inhibit the acoustic startle reflex. Scopolamine increased the gap threshold and reduced reflex inhibition produced by gaps that were presented at and beyond about 40 ms prior to the startle reflex, but not at shorter lead times. A peripheral antagonist had no effect at long lead times. These data indicate that central cholinergic mechanisms are involved in relatively high level perceptual processing of gaps. This conclusion is consistent with the hypothesis that temporal acuity may be compromised in the aged listener because of deficits in the efficacy of these central mechanisms. PMID- 10867291 TI - Organisation of binaural interactions in the primary and dorsocaudal fields of the guinea pig auditory cortex. AB - This study investigated the nature and topography of binaural interactions in the primary auditory field (AI) and dorsocaudal field (DC) of the urethane anaesthetised guinea pig auditory cortex. Single and multi-units were classified by their responses to monaural and binaural stimulation. In both AI and DC, units displayed binaural facilitation, binaural inhibition, or a level dependent mixture of facilitation and inhibition. There was a significant difference in the distribution of binaural response types between the two fields. Facilitated units predominated in DC (facilitated: 58%; inhibited: 24%; mixed: 6%; non-interacting: 12%), while inhibited units were the most common class in AI (facilitated: 15%; inhibited: 44%; mixed: 18%; non-interacting: 22%). It has previously been suggested that inhibited and facilitated units are concerned with processing different areas of space suggesting a possible separation of function between the two core fields. Topographically, the binaural response properties in AI and DC varied along isofrequency bands, with neurones displaying similar interactions aggregating in clusters. These clusters were similar in size for the two fields and often overlapped neighbouring isofrequency bands. However, their shape and position varied between different animals. This clustered organisation of binaural interactions is similar to that reported in recent studies of AI in other mammals. PMID- 10867292 TI - Characterizing non-linearity in the cochlear microphonic using the instantaneous frequency. AB - In this paper, we examine the non-linearity of mechano-electric transduction in the cochlea by computing the instantaneous frequency (IF) of the cochlear microphonic (CM) in response to sinusoidal stimuli. In contrast to a linear system which yields a constant IF when driven with a sinusoid, the IF of the CM varied during one period of oscillation. This variation was not symmetric, but differed for positive and negative slopes of the CM. Administration of tetrodotoxin to eliminate neural activity indicated that the variation of the IF was not due to neural contamination. Moreover, comparing the IF of the stimulus to that of the CM indicated that the IF was not due to non-linearity in the acoustic signal. Signal frequency, signal level and acoustic trauma altered the IF. A cochlear model of the CM was developed to determine the influence of the saturation of hair-cell receptor currents and vector summation on the IF. Results indicated that these factors could not fully account for the variation in the IF. We conclude that the variation in IF within one period of cochlear partition vibration indicates that the mechanical and/or electrical oscillations which produce the CM differ from those of a linear system. PMID- 10867293 TI - Mathematical decomposition of the round window potential. AB - We present an algorithm called the median transform which can be used to decompose the round window auditory potential into AC and DC components. The first of these is identified with the cochlear microphonic, and the second with the combined summating and compound action potentials. Elsewhere in this volume, the algorithm is employed as an intermediate step in obtaining the instantaneous frequency of the CM. Since the algorithm is easily implemented and operates entirely in the time domain, it may prove useful to clinicians as well as researchers. PMID- 10867294 TI - Motivation to change substance use among offenders of domestic violence. AB - Substance use alone leads to increased rates of violence, reduction in adherence to treatment regimes, and other negative psychiatric sequelae. Given the high rates of co-occurring substance use and family violence-related problems, substance use was assessed among offenders of domestic violence who were mandated by court to attend anger management classes. Rates of substance dependence diagnoses ranged from 33 to 50%, while rates of substance abuse diagnoses ranged from 60 to 75%. This study evaluated the effectiveness of a motivational enhancement intervention on readiness to change substance use. Two anger management groups were targeted to assess substance use, violence, and motivation to change substance use behaviors. One group was randomly chosen to partake in a motivational enhancement intervention session. The comparison group was offered standard anger management classes. Forty-one clients were evaluated for substance abuse and dependence diagnosis using criteria from the Diagnostic and Statistical Manual of Mental Disorders, 4th ed. A brief motivation to change survey, adapted from the Readiness to Change subscale of the Stages of Change Readiness and Treatment Eagerness Scale was administered pre- and postsession. Results indicate that a motivational enhancement intervention is feasible and effective in increasing readiness to change substance use among domestic violence offenders. The results illustrate the importance of assessing and treating substance use among offenders of domestic violence, as this may be an important indicator for higher dropout rates and reoffenses among this population. PMID- 10867295 TI - Services provided during methadone treatment. A gender comparison. AB - Greater improvement in posttreatment outcomes has been shown in programs that tailor frequency and type of services to unique client needs. Using a sample of 635 clients (199 females and 436 males) admitted to three community-based methadone treatment programs, this study examined gender differences in services needed and provided during the first 3 months of treatment. Results revealed that compared to males, women entered treatment with more psychological symptoms and AIDS/HIV-risky behaviors; they also presented with less criminal activity, less alcohol use, and higher motivation. Counselors addressed psychological and crisis issues more frequently with women, and counseling strategies were more often directed toward developing problem-solving and communication skills. Counselors also made more medical referrals and reported having better rapport with females. Attention to employment issues and HIV/AIDS sexual-risk behaviors did not differ by gender, even though women had more needs in these areas. PMID- 10867296 TI - Continuation of high-risk behavior by HIV-positive drug users. Treatment implications. AB - Drug users who are positive for the human immunodeficiency virus (HIV) represent a major vector of HIV transmission, yet relatively little is known about their continued drug- and sex-related HIV-risk behavior, which may impede the development of effective risk-reduction interventions. In this study, 50 HIV seropositive injection drug users entering methadone maintenance treatment completed a comprehensive risk assessment battery, including self-report of HIV risk behavior since learning HIV serostatus, and measures of risk-reduction information, motivation, and behavioral skills. We found that a disconcertingly high proportion of patients (66%) reported having engaged in HIV-risk behavior since learning their HIV-seropositive status. Level of HIV-related knowledge did not predict high-risk behavior. Drug-related risk behavior was predicted by psychiatric severity and poor behavioral skills. Sex-related risk was predicted by low levels of motivation and poor behavioral skills. Implications of these findings for treatment are discussed. PMID- 10867297 TI - Parental stress and child behavioral outcomes following substance abuse residential treatment. Follow-up at 6 and 12 months. AB - Residential treatment programs specifically designed for alcohol/drug-addicted women and their children have become a popular treatment modality across the United States. Outcome evaluation of these programs are beginning to show promising results. In this article, outcome data from a study of a residential substance abuse treatment program for women and young children in rural South Carolina will be presented. Data from 35 women and 23 children in the area of addiction severity, parenting and child emotional and behavioral development at 6 and 12 months following discharge from a substance abuse residential treatment program is examined. Results showed that women who completed treatment had better scores on addiction severity and parental stress, and their children had improved behavioral and emotional functioning at 6 and 12 months after discharge from the program. These results suggest that residential treatment has benefits for mothers and their children. This data adds to the growing body of evidence supporting intensive and inclusive care for certain groups of individuals with substance use disorders during critical periods. PMID- 10867298 TI - Posttraumatic stress disorder and short-term outcome in early methadone treatment. AB - The aim of this study was to determine treatment adherence relative to frequency of violence and posttraumatic stress disorders (PTSD) among new methadone patients. Ninety-six opiate-abusing patients were evaluated for childhood physical and sexual abuse (CPSA), adulthood exposures to violence (ADVIOL), PTSD, and treatment adherence. Overall, 43% of the subjects dropped out of treatment within 3 months of intake. Occurrence of trauma or PTSD did not predict drop-out rates. A 2 (Gender) x 2 (PTSD) analysis of covariance (ANCOVA) with severity of other drug use on admission as a covariate, however, revealed a main effect for PTSD, F(4, 71) = 7. 69, p < or =.01, such that those patients with current PTSD revealed significantly more ongoing drug use at 3 months (M = 24.3, SD = 20. 9) than those without (M = 8.9, SD = 11.8). Examination of ongoing cocaine use using a 2 (Gender) x 2 (PTSD) ANCOVA also revealed a main effect for PTSD, F(4, 17) = 8.24, p < or = .005, such that those patients with current PTSD revealed significantly more ongoing cocaine use at 3 months postadmission (M = 51.6, SD = 37.6) than those without (M = 24.3, SD = 20.9). For both genders, CPSA and ADVIOL were associated with higher rates of PTSD, which in turn predicted poorer treatment adherence as measured by ongoing co-occurring drug abuse 3 months postadmission. Results underscore the need for routine assessment and targeted treatment of trauma in methadone patients. PMID- 10867299 TI - Emergency room outreach to chronically addicted individuals. A pilot study. AB - There is a dearth of literature describing the treatment needs of substance abusing or chronically mentally ill homeless individuals who frequently utilize emergency medical services. This homeless subset represents a discrete population in the larger homeless community. We describe a pilot program, supported by local county public funds, and conducted by a local nonprofit social work agency, which was designed to provide intensive case management services to such a population. Outreach and case management activities resulted in linking clients to a broad range of entitlements and community services. Among those receiving outreach and case management services (n = 10), emergency services decreased by 58% in the year following referral compared to the year before (p <.03). Emergency services for the purpose of this study are defined as ambulance response and transport followed by emergency room admission and treatment. Those in a comparable control group (n = 8) showed no decrease in emergency service use. These results suggest that such community-based outreach programs can significantly improve patient outcome and provide substantial cost savings for local governments and hospitals. PMID- 10867300 TI - Patterns of drinking and drinking outcomes among drug misusers. 1-year follow-up results. AB - This article investigates patterns of drinking and drinking outcomes among 753 drug-misusing clients recruited to the National Treatment Outcome Research Study (NTORS). More than one third of those who were drinking at intake reported problematic or highly problematic patterns of alcohol consumption. About one third of the sample were abstinent from alcohol at intake and at follow-up. Some improvements in drinking behavior were found at 1-year follow-up, especially among the heaviest and most problematic drinkers. Improvements were specifically related to patterns of preintake drinking behavior. The majority of clients made little change to their pattern of pretreatment drinking behavior and the continued heavy drinking of many drinkers at follow-up is a disappointing finding. Drinking problems have been given insufficient attention in the treatment of illicit substance misuse problems, and efforts should be made to develop and strengthen the assessment and treatment of drinking problems among drug misusers. PMID- 10867301 TI - Occupational medical program alcohol screening. Utility of the CAGE and BMAST. AB - Alcohol consumption is a primary or secondary factor in many work-related accidents, suicides, homicides, violent crimes, and motor vehicle accidents. The absentee rate in alcoholics is 3.8 to 8. 3 times greater than that for nonalcoholic workers. The purpose of this research was to evaluate the effectiveness of two interview questionnaires-the Brief Michigan Alcoholism Screening Test (BMAST) and the CAGE (cut down, annoyed by criticism, guilty about drinking, and eye-opener drinks). The validity of the BMAST and the CAGE as screening tools for alcohol problems has been verified in a number of nonworkplace settings. If they prove to be as effective for screening workers in an occupational medical setting, follow-up definitive diagnoses could result in earlier detection of alcohol problems and allow prompt intervention. Positive outcomes could include a safer workplace, less absenteeism, improved worker productivity, and a reduction in personal and family problems caused by drinking. PMID- 10867302 TI - HIV risk behaviors in male substance abusers with and without antisocial personality disorder. AB - Antisocial personality disorder (ASP) is common in male substance abusers and may be associated with increased human immunodeficiency virus (HIV) risk behaviors. In this study, 91 male substance abusers were recruited from the community, and 42% met diagnostic criteria for ASP. Although ASP and non-ASP subjects demonstrated equivalent knowledge about HIV, subjects with ASP participated in more risky behaviors. On a lifetime measure of drug risk behaviors, ASP subjects reported higher rates of intravenous drug use (IVDU), frequency of needle sharing, and number of equipment-sharing partners and lower rates of needle cleaning. On a measure of past-month risk behaviors, ASP subjects reported higher rates of IVDU and lower rates of needle-cleaning. Subjects with ASP also reported greater participation in lifetime sexual risk behaviors, including number of sexual partners and frequency of anal sex. These findings suggest that clients entering substance abuse treatment programs should be screened for ASP, and clients identified with ASP should be provided risk-reduction interventions early in treatment. PMID- 10867303 TI - Older adult treatment outcome following elder-specific inpatient alcoholism treatment. AB - This study examined multidimensional 6-month outcomes of elder-specific inpatient alcoholism treatment for 90 participants over the age of 55. At baseline, physical health functioning was similar to that reported by seriously medically ill inpatients in other studies while psychosocial functioning was worse, and nearly one third of the sample had comorbid psychiatric disorders. Based on 6 month outcomes, participants were classified into the following groups: Abstainers, Non-Binge Drinkers, and Binge Drinkers. The groups did not differ on any baseline measures (demographics, drinking history, alcohol symptoms and age of onset, comorbidity, or length of treatment). General health improved between baseline and follow-up for all groups. Psychological distress decreased for Abstainers and Non-Binge Drinkers, but did not change for Binge Drinkers. Results suggest that a large percentage of older adults who receive elder-specific treatment attain positive outcomes across a range of outcome measures. PMID- 10867304 TI - The combination of phentermine and fenfluramine reduced cocaine withdrawal symptoms in an open trial. PMID- 10867305 TI - Abstinent-contingent housing and treatment retention among crack-cocaine dependent homeless persons. AB - This study investigated Behavioral Day Treatment attendance in relation to treatment outcome among homeless persons dependent on crack-cocaine. Participants (N = 141) were 72.3% male and 82.7% African American. Days attended, activities attended, and follow-up rates over a 12-month period were positively affected by the more attractive treatment of providing immediate, rent-free, abstinent contingent housing during a 2-month Behavioral Day Treatment program. Results replicated previous findings that abstinence is a function of treatment attendance and more treatment is associated with greater abstinence. The loss of predictive power at long-term follow-up, limitations of a retrospective design, need to identify most predictive therapeutic activity types, and potential influence of mental disorders were discussed. Analytical techniques used in this study allows for the planning, predictability, and measurement of drug abuse treatment success as a function of service utilization. PMID- 10867306 TI - "That original tension". Negotiating abstinence in clinicians' accounts of harm reduction in nonresidential treatment of heroin withdrawal. AB - The aim of this article is to examine how drug and alcohol clinicians, guided by a policy of harm reduction, approach their withdrawal work in their encounter with injecting heroin users seeking nonmethadone withdrawal treatment. The study, qualitative in design, involved detailed interviews with all seven clinicians who worked in the nonmethadone withdrawal program of a nonresidential drug and alcohol center in Melbourne, Australia. I draw attention to the difficulties that these clinicians have in their withdrawal work, especially concerning the place of abstinence in withdrawal and in harm reduction. Abstinence is a legitimate goal of harm reduction. Yet, how harm-reduction knowledge is practiced and reproduced in the clinical encounter is underpinned by dominant and taken-for granted assumptions about abstinence as 'other' to harm reduction. The ideal of abstinence in drug and alcohol treatment and its decentering within the concept of harm reduction, make introducing harm-reduction strategies in the clinical encounter precarious. The work of withdrawal is compromised with an unresolved tension brought about through the paradox of legitimating illicit drug use in one context (the medical) when it is not legitimate in another context (the sociopolitical). PMID- 10867307 TI - Characterizing and dating the onset of symptoms in psychotic illness: the Symptom Onset in Schizophrenia (SOS) inventory. AB - Prodromal symptoms, including disturbances of perceptions, beliefs, cognition, affect, and behavior, are often the first symptoms of schizophrenia. Little is understood about the initial, prodromal stage of schizophrenia, despite the compelling research and clinical need. The development and psychometric properties of a new, time-efficient instrument to characterize and date the initial symptoms of a psychotic illnesses, the Symptom Onset in Schizophrenia (SOS) scale, is described in this paper. The SOS rates the presence and dates the onset of 16 general prodromal, positive, negative, and disorganizational symptoms, as well as a clinician, family, and patient global rating of onset of illness. Inter-rater reliability for the presence of each symptom in 35 patients with schizophrenia, schizoaffective, or schizophreniform disorder was good to excellent, with kappa coefficient >0.7 for 12 items, and >0. 5 for all items. Agreement on symptom duration was good to excellent for individual items (ICC=0.7 1.0) and for global rating of duration of illness (ICC=0.97). Our data indicate that the SOS is a reliable, valid, time-efficient tool useful to retrospectively assess the onset of schizophrenia and related psychotic disorders. Further study is underway to evaluate other psychometric properties of the SOS, including test retest reliability and predictive validity. PMID- 10867308 TI - Brain and ventricle instability during psychotic episodes of the schizophrenias. AB - Recent reports from serial brain scans suggest that the rate of ventricular expansion and/or brain atrophy may be accelerated in at least some schizophrenics. The authors assessed the effect of state changes upon such findings.Within-subject 3D MRIs were assessed for ventricular and brain volumes during periods of [partial] remission and of exacerbation of psychosis. Additional scans at comparable within-subject SAPS were used to assess rates of change in volumes that were independent of SAPS changes. Correlations of changes of ventricle and brain volumes vs. change of SAPS cores between scans revealed that ventricle volumes decreased during a period of psychotic exacerbation and increased at a time of [partial] remission (r(p)=-0.666; P<0.0005); conversely, brain volumes increased during psychotic exacerbation and decreased at [partial] remission (r(p)=+0.448; P=0.032). Scans at comparable SAPS scores suggested that the majority of patients had rates of ventricular expansion comparable to controls (0.9+/-0.6 cc/year), though two patients appeared to have rates of ventricular increase of 4.5+/-2. 1 cc/year (Lilliefores P=0.036; K-means clustering F=17.75). Exacerbation of psychosis in schizophrenia is accompanied by evidence of brain swelling, especially of periventricular brain, with encroachment of brain substance upon ventricular volumes. Controlled for state changes, the majority of schizophrenics show rates of ventricular expansion or brain atrophy indistinguishable from controls. PMID- 10867309 TI - Cerebral ventricular enlargement as a generalized feature of schizophrenia: a distribution analysis on 502 subjects. AB - Enlargement of cerebral ventricles is one of the most replicated biological features, and the one quantitatively most deviant in schizophrenia. It occurs in the early phases of the disease and may have pathogenetic relevance. Whether this abnormality is limited to a specific subgroup of patients or is a common feature to most or all patients affected by schizophrenia, however, is still a matter of debate. The answer to this question would improve our comprehension of the nature of this abnormality and contribute to the debate between the competing hypotheses of biological homogeneity vs heterogeneity of schizophrenia.We performed a distribution analysis of lateral ventricular dimensions of 340 schizophrenic patients and 162 non-psychiatric controls. All subjects underwent cerebral computerized tomographic scan, and ventricular dimensions were expressed as ventricular brain ratio (VBR). After removing the effect of confounding variables (age, sex and type of scanner) on individual VBR, data were power-transformed and different distribution hypotheses were tested by means of the maximum log likelihood ratio method. Our findings indicate that, in the mixed sample of patients and controls, a mixture of two gaussian curves represents the distribution better than a single gaussian curve, but no evidence emerged leading to rejection of the normality hypothesis in the schizophrenic patients sample. Lateral ventricular enlargement in schizophrenia is not a marker of a discrete subgroup of schizophrenia, but occurs in most, if not all, schizophrenic patients. This supports the hypothesis of biological homogeneity of the disease, at least relative to its major brain morphological abnormality. PMID- 10867310 TI - Serotonin(2A) receptors are reduced in the planum temporale from subjects with schizophrenia. AB - [(3)H]ketanserin binding to 5HT(2A) receptors was measured in the left planum temporale (sensory speech cortex) from schizophrenic and non-schizophrenic (control) subjects using both particulate membranes and tissue sections. There was a significant decrease in the affinity of [(3)H]ketanserin binding to particulate membranes from schizophrenic subjects who were treated with phenothiazines up to death. Adding 2nM chlorpromazine to brain tissue from control subjects caused a similar decrease in the affinity of [(3)H]ketanserin binding to particulate membranes. This suggests that the decrease in affinity observed in the phenothiazine-treated subjects was due to residual drugs. In addition, there was a significant decrease in the density of [(3)H]ketanserin binding in both particulate membranes and tissue sections from schizophrenic subjects which did not appear to be due to residual antipsychotic drugs. Analysis of the laminar distribution of 5HT(2A) receptors showed that this decrease was greatest in cortical layer III. The decrease in the density of 5HT(2A) receptors was significant whether schizophrenic subjects were receiving phenothiazines or haloperidol at the time of death, and there was no correlation between the last recorded dose of antipsychotic drug and 5HT(2A) receptor density. These data suggest that a decrease in the density of 5HT(2A) receptors in the planum temporale may be associated with the pathology of schizophrenia. PMID- 10867311 TI - Neurocognitive and social functioning in schizophrenia: a 2.5 year follow-up study. AB - In our previous study we demonstrated that, in 80 schizophrenia subjects, verbal ability, verbal memory and executive functioning were significantly associated with social problem solving. The objective of this present study was to assess the longitudinal stability of the relationship between social and neurocognitive functioning in schizophrenia. This 2.5 year longitudinal cohort study re-assessed community functioning, social problem solving and symptoms in 65 of the 80 original subjects to determine the predictive ability of neurocognitive functioning. Neurocognition was not re-assessed at this follow-up. Positive and negative symptoms were assessed with the Positive and Negative Syndrome Scale. Social functioning was assessed using the Social Functioning Scale, the Quality of Life Scale, and the Assessment of Interpersonal Problem-Solving Skills (AIPSS). Verbal ability, verbal memory and vigilance were significant predictors of social problem solving as assessed by the AIPSS. Results suggest that the association between neurocognition and social functioning remains consistent over time. PMID- 10867312 TI - Processing of context information in schizophrenia: relation to clinical symptoms and WCST performance. AB - Failure in contextual information processing has been hypothesized as being the single function responsible for several impairments in cognitive tasks and symptoms, through an involvement of the prefrontal cortex, in patients with schizophrenia. A variant of the Continuous Performance Test (CPT) designed specifically to elicit deficits in the processing of contextual information has been administered to 20 schizophrenic patients and 20 healthy controls. The relation to Wisconsin Card Sorting Test (WCST), relatively specific to prefrontal damage and executive dysfunctioning, and clinical status by using scales for the assessment of positive, negative symptoms and outcome has been investigated. The data show that multi-episode schizophrenic patients manifest inability to use contextual information to inhibit habitual response to an ambiguous stimulus and to maintain information across delay, without a general attention deficit. We also found a relationship between contextual reasoning and WCST unique errors, hallucinations, formal thought disorders, and outcome evaluation. Our results further support the hypothesis that the deficit of contextual reasoning could account for cognitive impairments and symptoms of patients with schizophrenia. PMID- 10867313 TI - Habituation of the blink reflex in first-episode schizophrenia, psychotic depression and non-psychotic depression. AB - OBJECTIVE: Electrophysiological recording of the electrically elicited blink reflex is the most reliable method of investigating habituation of the startle reflex. The purpose of this study was to compare the habituation and the late R3 component of the blink reflex between control subjects (N=19) and first-episode patients with schizophrenia (N=17), psychotic depression (N=23), and severe non psychotic depression (N=25). METHODS: The blink reflex was evoked by electrical stimulation of the supraorbital nerve, and the deficient habituation of the R2i component was measured with a computer-assisted integral area measurement. Prefrontal executive function of the patients was assessed with the Wisconsin Card Sorting Test. Current psychiatric symptoms were assessed with the Brief Psychiatric Rating Scale, the Hamilton Depression Scale, the Positive and Negative Syndrome Scale, and the Calgary Depression Scale. RESULTS: Deficient habituation of the blink reflex and occurrence of the late R3 component were associated both with a previous diagnosis of psychotic disorder and with the presence of current psychosis. The sensitivity and specificity of the abnormal habituation of the blink reflex in detecting psychotic disorder were 0.50 and 0.80, respectively. The abnormalities of the blink reflex were not associated with psychotropic medication. In schizophrenic patients, defective habituation of the blink reflex was associated with negative and cognitive symptoms, and in depressive patients with the presence of delusions. CONCLUSIONS: The deficient habituation of the blink reflex and occurrence of the late R3 component seem to be both trait and state markers of a psychotic disorder. The results suggest that schizophrenia and psychotic depression share some common neurobiological mechanisms involved in the modulation of the startle reflex. PMID- 10867314 TI - Main risk factors for schizophrenia: increased familial loading and pre- and peri natal complications antagonize the protective effect of oestrogen in women. AB - Women fall ill with schizophrenia 3 to 4 years later than men. The neurobiological mechanism, explaining the delay of onset in women until menopause, is presumably due to a sensitivity reducing effect of oestrogen on central d(2) receptors, as we have previously shown in animal experiments and in a controlled clinical study. The gender difference in age at onset seems to disappear in familial cases with schizophrenia, but it increases to highly significant values of 5 years or more in isolated cases according to a recent study by Albus and Maier (Schizophrenia Research 18:51-57, 1995). We tried to replicate these findings and to test the hypothesis of a functional antagonism between genetic predisposition to illness and the protective effect of oestrogen in a population-based sample of 232 first illness episodes of schizophrenia. In women with at least one first-degree relative suffering from schizophrenia, age at onset defined by first psychotic symptom was significantly reduced by several years and the difference with men disappeared. In sporadic female cases (no mental disorder in first-degree relatives) the age at onset was slightly increased compared with the total sample, which was in accordance with our hypothesis. In men with familial schizophrenia, but without a protective agent like oestrogen, the age at onset was only slightly and non-significantly reduced compared with the total group and with sporadic cases. This was in line with Albus and Maier and with our hypothesis that only the protective effect of oestrogen could be antagonized by a strong genetic disposition. The second main risk factor for schizophrenia is pre- and peri-natal complications. We compared men and women from our sample of first illness episodes with a history of pre- and peri-natal complications with those without a history of obstetric complications. In women the age at first psychotic symptom was markedly reduced, but due to small case numbers not significantly, compared with women without the risk factor and with the total group. Again, schizophrenic men with a history of pre- and peri-natal complications showed only a small, non-significant reduction of age at onset compared with the total and the group without the risk factor. Therefore, we concluded that the degree of genetically determined vulnerability and, presumably to a slightly lesser extent, the degree of pre- and peri-natal brain injury antagonizes the onset delaying effect of oestrogen in schizophrenia. PMID- 10867315 TI - Reproductive characteristics of Ethiopian highland sheep. II. Genetic parameters of semen characteristics and their relationships with testicular measurements in ram lambs. AB - A study was conducted to investigate the effect of season on, and to estimate heritabilities of, and genetic correlations among, semen and spermatozoa characteristics and the relationship of these characteristics with measures of testicular or scrotal size in 6-, 9- and 12-month old ram lambs of Menz and Horro sheep breeds indigenous to Ethiopian highlands. A total of 278 ram lambs with substantial pedigree information were involved in the study. There was improvement in all semen and spermatozoa traits with age, the means at 12 months being consistently superior to values at 6 and 9 months of age. There were no significant breed differences in any of the traits studied, except semen volume at 9 months (0.67, S.E. 0.07 for Horro versus 0.39, S.E. 0.05ml for Menz) and proportion of dead spermatozoa at 12 months (0.18, S.E. 0.03 versus 0.23, S.E. 0.02, respectively). Season was significant (p<0.05) for most of the traits studied, and differences were attributed primarily to nutrition. Semen collected in the wet season had higher spermatozoa concentration while samples collected in the dry season had higher proportion of abnormalities. After correcting for differences in total spermatozoa abnormalities, the wet season (of collection) had the highest spermatozoa output (volumexconcentration) in 12-month old ram lambs. Heritability estimates varied substantially in magnitude, ranging from zero to over 0.4. Traits with non-trivial heritability estimates were mass motility at 9 months (0.32, S.E. 0.11), individual motility at 9 months (0.32, S.E. 0.12) and at 12 months (0.16, S.E. 0.12) and proportion of abnormal spermatozoa at 9 months (0.35, S.E. 0.13). Genetic correlations among semen and spermatozoa characteristics, as well as correlations with testicular measurements, were medium to high and generally favourable, but were associated with large standard errors. The genetic correlations of scrotal circumference with semen volume (0.55, S.E. 0.11), mass motility (0.62, S.E. 0.20), individual motility (0.54, S.E. 0.12), concentration (0.25, S.E. 0.04) and proportion of abnormal spermatozoa (-0.75, S.E. 0.24) in 12-month old rams indicated that selection based on this trait, which is highly heritable and easy to measure and can be measured early in life, should have appreciable favourable correlated response in semen quality and spermatozoa production. PMID- 10867316 TI - Production performance, repeatability and heritability estimates for live weight, fleece weight and fiber characteristics of alpacas in New Zealand. AB - The production performance, repeatability and heritability estimates for live weight, fleece weight and fiber characteristics of alpacas farmed in the South Island of New Zealand are reported. Male alpacas produced heavier fleeces (p<0.001) than females, but with relatively similar fiber diameter. Mean (S.E.) shearing weight, greasy fleece weight (GFW), clean fleece weight (CFW), yield, staple length (SL), resistance to compression (RtC) and fiber diameter (FD) in adults were 68.0kg (1.0), 2.16kg (0.06), 2.03kg (0.06), 93.6% (0.4), 9.9cm (0.2), 5.3kPa (0.1) and 31.9um (0.5), respectively. These means in tuis were 68.1kg (1.9), 3.02kg (0.20), 2.94kg (0.27), 92.2% (0.4), 12.2cm (0.3), 4.8kPa (0.1) and 30.5um (0.9), respectively. The corresponding measurements in crias were 40.5kg (1.1), 1.97kg (0.07), 1.84kg (0.07), 93.4% (0.3), 12.6cm (0.2), 4.6kPa (0.1) and 26.4um (0.4), respectively. The birth weight (BWT) was 8.4kg (0.1) and SS was 28.4 N/ktex (1.9) in crias. The seasonal variation of fiber growth and fiber diameter was small to moderate, with lowest values in the winter. Mid-side fleece site FD was highly correlated with other main sites sampled and shown to be appropriate as a standard sampling site. The phenotypic correlation between CFW and FD was 0.40 (p<0.001) and for fleece weight and shearing live weight was 0.47 in adult alpacas (p<0.001). Correlation coefficients for GFW and CFW with FD and SL were highly positive (GFW with FD and SL: 0.32-0.45, 0.39-0.54; CFW with FD and SL: 0.37-0.46, 0.40-0.53) in both tui and cria fleeces. The heritabilities for BWT, summer weight, spring weight, GFW and CFW, yield, SL, RtC and FD were estimated as 0.63, 0.41, 0.99, 0.63, 0.68, 0.67, 0.57, 0.16, 0.69 and 0.73. Production performance and heritability estimates for these traits were markedly higher than that previously reported in South American camelids. PMID- 10867317 TI - Genetic parameters for growth traits of the Moroccan Timahdit breed of sheep. AB - Data from 522 lambs of the Timahdit breed representing 13 sires were used to estimate genetic parameters. Traits analyzed were weights at birth (BW), 30 days (W30), 70 days (W70) and 90 days (W90), average daily gains from 10 to 30 days (ADG10-30), 30-70 days (ADG30-70) and 30-90 days (ADG30-90). REML estimates of variance and covariance components were obtained assuming animal models that included the fixed effects of sex, age of dam and the combined effect of year and types of birth or rearing, and the animal random effects for the direct and the maternal genetic effects. Genetic correlations were estimated with the same fixed effects and only additive direct genetic effects. Results showed that an important direct additive and maternal genetic effects were observed. Estimates of the direct heritability and maternal heritability were respectively 0.18, 0.59 (BW); 0.31, 0.34 (W30); 0.50, 0.24 (W90); 0.42, 0.08 (ADG10-30), and 0.31, 0.22 (ADG30-90). Estimates of additive direct-maternal correlations were important and ranged from -0.94 to -1.0 for live weights, and from -0.70 and -1.0 for daily gains. The estimates of genetic and phenotypic correlations were high and showed no genetic antagonisms among the growth traits. PMID- 10867318 TI - Oral pharmacokinetics of fenbendazole in llamas, South American Camelids. AB - Llamas, South American Camelids are increasingly popular in the United States, as a source of fiber, livestock guard, and pack animals. Gastrointestinal parasites have been identified as a major health problem in all classes of livestock including llamas. Currently, there are no approved anthelmintics available for use in llamas. In this study, the pharmacokinetics of a single, oral administration of fenbendazole paste at a minimum target dose of 5mg/kg, with an upper limit of <10mg/kg, was evaluated in llamas. Plasma fenbendazole concentration time profiles were best described by a single compartment model. After oral administration of fenbendazole, T(max) and Cp(max) were 28.39+/ 12.80h, and 0.28+/-0.17ug/ml, respectively. The T(1/2alpha) and T(1/2beta) were 16.25+/-11.67 and 36.00+/-25.00h, respectively. The apparent volume of distribution (V(d)) and the area under the curve (AUC) were 11.28+/-4.66l/kg, and 22.52+/-8.67ugh/ml, respectively. The results of this preliminary study indicate that when the paste formulation of fenbendazole is administered orally to llamas, its rate of absorption appears to be very similar to that of other ruminants including sheep, goats, and cattle as indicated by the time required to reach peak plasma concentrations. It was also found that the rate of elimination of fenbendazole was prolonged in llamas as compared to sheep, goats, and cattle. PMID- 10867319 TI - Bioserogroups of Campylobacter species isolated from sheep in Kaduna State, Nigeria. AB - Sheep Campylobacter isolates from Kaduna State were characterized into their species and bioserogrouped. A total of 1100 samples were collected from Kaduna abattoir and National Animal Production Research Institute (NAPRI), Shika. The samples were from 250 gallbladder, 250 intestinal contents, 100 fetal stomach contents all from Kaduna abattoir while 250 rectal swabs and 250 vaginal swabs were from the NAPRI Small Ruminant Programme. Of a total of 1100 samples, 39 (3.54%) yielded Campylobacter organisms. The highest isolation rate (6.8%) was from samples of intestinal contents followed by those from gall bladders (4.0%). Samples from the vaginal and fetuses had the lowest isolation rates (2.80%) and (0%), respectively. Of the 39 Campylobacter isolates from all the sources, (79%) were characterized as C. fetus subsp jejuni, C. coli (13%) and C. laridis (8.0%). while C. coli and C. laridis were isolated from gall bladder and intestinal contents only. Campylobacter fetus subsp jejuni biotype 1 accounted for 40.3% of the total isolates. C. laridis biotypes I and II were also isolated and accounted for 5% and 3% of the isolates, respectively. 5% of the isolates were not typeable. The serogroups 4 (13%), 36 (10%), 9 (10%), 84 (8%), 29 (5%) and 20 (8%) were the commonest serogroups identified in sheep at two locations surveyed. The isolation of Campylobacter organisms from rectum, vagina, gallbladder, and intestinal contents is a clear indication that sheep serves as a reservoir of this organisms in Nigeria. Similarities between documented human Campylobacter isolates in Nigeria and those in the present study raised the possibility of cross-transmission between sheep and man. It is concluded that biotyping and serotyping can be used for epidemiological study of campylobacteriosis due to Campylobacter jejuni in sheep in Kaduna State of Nigeria. PMID- 10867320 TI - Effect of breed and age on the voluntary intake and the micromineral status of non-pregnant sheep. 1. Estimation of voluntary intake. AB - Twenty-four sheep were used to test breed and age differences in voluntary intake (VI). The sheep were divided into four groups of six animals each: Suffolk mature ewes (SM), Suffolk yearlings (SY), Rambouillet mature ewes (RM), and Rambouillet yearlings (RY). The animals grazed alternatively two mixed pastures containing Trifolium repens latum, Lolium perenne and Pennisetum clandestinum from 07:00 to 16:30 hours and remained penned overnight. Voluntary intake was estimated using chromic sesquioxide (1g per sheep) as an external marker, administered in two gelatin capsules, one before and one after grazing, during 65 days. Faecal grab samples were collected directly from each animal for three consecutive days beginning on Day 7 of the experiment and at 14-day intervals until Day 65 (sampling times=5). Hand-plucked samples of forage were used to estimate in vitro dry matter digestibilities (IVDMD), which were used with estimates of fecal output to calculate intakes (per animal and per kg of metabolic body weight (kg(0.75))). The data were analyzed as a completely randomized design in a factorial arrangement with repeated measurements. Voluntary intake, expressed both per animal and per kg(0.75), showed a breedxage interaction (P<0.001). Suffolk mature ewes had the greatest daily VI (1.88 versus 1.26kgDM/animal for the other three groups) because they were the heaviest animals, but their VI/kg(0.75) (0.071kgDM/day) was not different (P>0.05) from that of RY (0.063kgDM/day), which were smaller, younger animals. Suffolk yearlings and RM had similar VI/kg(0.75) (0.057 and 0.051kgDM/day, respectively), not different from that of the RY. Intakes showed a cubic trend (P<0.001) with time, possibly because of changes in IVDMD and climatic conditions. The lower intake, yet better gains of RM and RY indicate a greater ability to thrive under conditions of average pasture quality. High forage availability did not result in greater forage consumption. The information on VI was later used to assess the micromineral status of the sheep in the experiment. PMID- 10867321 TI - Effect of breed and age on the voluntary intake and the micromineral status of non-pregnant sheep. II. Micromineral status. AB - Twenty-four non-pregnant sheep, divided into four groups of six animals each, were used to test breed (Suffolk and Rambouillet) and age (mature (six years old) and yearling (16 months old) ewes) differences in micromineral status. The animals grazed alternatively two mixed pastures containing Trifolium repens latum, Lolium perenne and Pennisetum clandestinum from 07:00 to 16:30hours, and remained penned overnight. Hand-plucked forage samples, blood and wool samples were collected on Days 1 (except forage), 7 (except wool), 21, 35, 49, and 63 of the experiment and assayed for Zn, Fe, Cu, Se, and Mo. Blood and wool data were analyzed as a completely randomized design in a factorial arrangement with repeated measurements. Zinc blood levels (2.15mgkg(-1), below normal values) showed a sampling timexbreed interaction (P<0.05) with a quadratic trend (P<0.02). Iron blood levels (337.48mgkg(-1)) showed a sampling timexage interaction (P<0.05), with mature ewes having higher and more variable levels than yearlings. A sampling time effect (P<0.01) with a quadratic trend (P<0.05) dependent on the breed (P<0.01) was found in Se blood levels. Rambouillet sheep had higher (86ugkg(-1)) initial levels than Suffolk sheep (61.14ugkg(-1)) and they declined linearly, whereas Se levels in Suffolks showed a quadratic trend. Copper blood concentration (below normal values) was higher (P<0.01) in Suffolks (0.46mgkg(-1)) than in Rambouillets (0.35mgkg(-1)). Suffolk sheep had higher Zn (P<0.01) and Cu (P<0.05; below normal values) levels in wool (146.63 and 2.16mgkg(-1), respectively) than Rambouillet sheep (141.34 and 1.72mgkg(-1), respectively). No effect was found in Fe wool levels (291.22mgkg(-1)), and Se concentration in wool (204.7ugkg(-1)) only showed a sampling time effect (P<0.05) with a decreasing linear trend (P<0.01). Coefficients of variations higher than 50% were found for Zn and Se in blood and Fe and Se in wool. Molybdenum could not be detected either in blood or in wool. Rambouillet sheep, in general, showed lower and less variable micromineral levels than Suffolk sheep. Because no signs of deficiency were observed nor was the animals' productivity affected after the study, it is considered that reported normal values for Zn and Cu could be more variable than suspected, depending on breed, location, feeding practices and physiological state. PMID- 10867322 TI - Nitrogen metabolism and renal function in the dik-dik antelope (Rhynchotragus kirkii). AB - Nitrogen metabolism and kidney function of the dik-dik antelope (Rhynchotragus kirkii) were studied under a range of controlled, experimental conditions and diets. Dik-dik antelope remained in nitrogen balance even when fed a diet low in protein and high in fibre. When fed a diet high in protein (20%) and water ad libitum, 55.3% of the urea filtered by the kidney was reabsorbed. Limiting water intake increased urea reabsorption to 77.2%. The U/P urea concentrations were maintained at similar ratios on all diets, as well as during dehydration and solute loading. Minimum endogenous nitrogen excreted was 74mg/kg(0.75)/day. Dehydration (water deprivation) and solute loading (intraruminal infusion of 0.25M NaCl) had varying effects on nitrogen metabolism. It is concluded that the metabolic nitrogen economy of the dik-dik antelope is qualitatively similar to that of other domestic and wild ruminants. PMID- 10867323 TI - Lamb production and forage quality under a forage system consisting of AU Triumph tall fescue and Tifton 44 bermudagrass. AB - The lamb production potential and forage quality of a forage system consisting of AU Triumph fescue and Tifton 44 bermudagrass was studied over a period of two consecutive years. Each spring, 60 yearling Suffolk ewes and 40 wethers were divided into two groups and allowed to graze two replicated 1ha AU Triumph paddocks continuously. At the end of the spring grazing season, both groups were transferred to two replicated 1ha Tifton 44 paddocks for summer grazing. Body weight was recorded fortnightly and composite forage samples analyzed for chemical composition, in vitro dry matter disappearance (IVDMD) and in vitro organic matter disappearance (IVOMD) at the end of the study. Mean crude protein (CP), ether extract (EE), ash, neutral-detergent fiber (NDF), acid-detergent fiber (ADF), cellulose, lignin, IVDMD and IVOMD contents of AU Triumph fescue were 13.26, 1.86, 7.38, 69.4, 36.37, 29.2, 6.81, 64.44 and 62.88%, respectively. The corresponding values for Tifton 44 bermudagrass were 7.5, 1.01, 5.2, 78.3, 41.0, 28.3, 8.8, 56.97 and 55.61%. Average daily gain (ADG) and lamb gain per hectare on AU Triumph fescue were 142.1g and 412.59kg/ha, respectively. However, lamb ADG started to decline and remained low when they were transferred from AU Triumph to Tifton 44 bermudagrass resulting in a low ADG and lamb gain per hectare of only 18.2g and 84.09kg/ha, respectively. In contrast, Tifton 44 yielded higher post grazing residual DM, suggesting that the decline in performance of the lambs grazing Tifton 44 to be related to poor forage quality rather than availability. The results indicate that low nutritive value of Tiftton 44 bermudagrass is a limiting factor to lamb production in the forage system studied. To prevent the observed decline in lamb performance, it might be necessary to fertilize Tifton 44 at a higher rate or replace it with a better quality warm season forage. PMID- 10867324 TI - Fatty acid composition of goat muscles and fat depots: a review. AB - In addition to the fat content of muscle and adipose depots, the fatty acid composition of lipids affects meat quality. Furthermore, relevant reports are difficult to use for comparisons, in that samples were collected from muscles and fat depots at various anatomical locations and experiments entailed different objectives, designs, procedures and methodologies. Nonetheless, based on currently available publications, according to a recent classification of meats by concentrations of potentially cholesterol-raising, and neutral, and cholesterol-lowering effects, average values for goat muscles appear better than for beef and lamb. Feeding dry diets seems to increase levels of unsaturated fatty acids and stearic acid in fat depots compared with milk or milk replacer. Increasing concentrate consumption can increase levels of odd-numbered and branched chain fatty acids in subcutaneous fat depots. With increasing age of unweaned kids, the level of stearic acid in fat depots decreases, and with increasing live weight of weaned kids levels of saturated fatty acids increase, and contents of monounsaturated fatty acids decrease in most fat depots. This review of a currently limited database indicates need for further experimentation to characterize interactions among factors such as breed, age and nutritional conditions in the fatty acid composition of carcass lipids of goats so as to gain a fuller understanding of goat meat quality. PMID- 10867325 TI - Effect of sectioning on the number of isolated ovine preantral follicles. AB - The aim of the present study was to test the effect of the interval of serial sections in the tissue chopper on the number of isolated ovine preantral follicles. Best results were obtained when the ovarian fragments were cut in the tissue chopper at interval of 87.5um (1592 preantral follicles per treatment). Histochemical analysis showed that the follicular morphology was preserved after mechanical isolation as demonstrated by the normality of oocytes and granulosa cells as well as by preservation of basement membrane. The percentages of isolated primordial, primary and secondary follicles were 92, 6, and 2%, respectively. The follicular sizes varied from 12.5 to 96.3um in diameter. In conclusion, a greater number of isolated preantral follicles were obtained when the method of isolation used the tissue chopper adjusted at 87.5um. Besides, this treatment does not affect the follicular integrity after the isolation procedure. PMID- 10867326 TI - Caprine sperm acrosome reaction: promotion by progesterone and homologous zona pellucida. AB - Experiments were designed to characterize the effect of progesterone and the zona pellucida (ZP) on the goat sperm acrosome reaction (AR) through a comparative study. Goat spermatozoa were incubated for 4h in Krebs-Ringer bicarbonate media (KRB) for capacitation. Progesterone and ZP stimulated exocytosis of capacitated spermatozoa in a dose-dependent manner. EGTA and La(3+), added 10min before the addition of the agonists, completely abolished the stimulatory effects. Ca(2+) influx was observed to occur through a calcium phosphate transporter. Picrotoxin and bicuculline, two GABA(A)/Cl(-) channel antagonists, also inhibited progesterone-induced AR when added 10min before steroid addition. ZP-induced AR was unaffected by these antagonists. Studies using pertussis toxin (PTX) showed that, unlike ZP, progesterone acts without the involvement of a G-protein. Progesterone-3-(O-carboxymethyl) oxime: BSA conjugate (P-BSA) also induced AR in capacitated sperm suspension. Results suggest that progesterone and ZP induce AR via specific membrane receptors through different mechanisms, both requiring an influx of Ca(2+). It is assumed that both the mechanisms reconcile at some stages of the cascade and elicits a similar physiological response. PMID- 10867327 TI - Growth performance of Malpura and crossbred lambs under intensive feeding. AB - Growth performance, feed conversion efficiency and nutrient utilization of mutton synthetic, selected and randombred Malpura populations of lambs (60days) were evaluated. The mutton synthetic lambs reached the 25kg body weight in 73days of intensive feeding while the selected and randombred Malpura populations of lambs reached the same weight in 91 and 136days, respectively (P<0.01). The fitted growth curve confirmed the mutton synthetic lambs grew rapidly and were more efficient in feed conversion (P<0.01) followed by selected and randombred Malpura lambs. Dry matter intake and nutrient utilization was similar in mutton synthetic and selected Malpura lambs and required 75g DM, 5.5g DCP and 42.1g TDN/kg W(0.75) for average daily gain of 148g. The lambs were in positive N, Ca and P balance whereas their N retention was lower (25%). It is concluded from this preliminary study that growth response and feed conversion efficiency were higher in mutton synthetic than selected Malpura lambs and lower in randombred Malpura lambs. PMID- 10867328 TI - Effect of whole and rolled corn or barley on growth and carcass quality of lambs. AB - Lambs were assigned to a 2x2 factorial design with corn or barley fed either whole or rolled. Hay was fed for ad libitum intake while concentrates were fed in similar amounts. Dry matter intake of hay, ADG, the gain:feed ratio, and warm carcass weight (WCW) were higher (P<0.05) for lambs fed corn than for those fed barley. Cereal processing had no effect (P>0.05) on hay dry matter intake, ADG, the gain:feed ratio, and WCW. Time of fattening (23-46kg) was similar (P>0.05) among treatments. Carcass characteristics generally were similar among treatments. Corn was more beneficial than barley to increase the ADG and carcass weight. PMID- 10867329 TI - Statins after cardiac transplantation: which statin, what dose, and how low should we go? PMID- 10867330 TI - Allograft coronary artery disease: clinical correlations with circulating anti HLA antibodies and the immunohistopathologic pattern of vascular rejection. PMID- 10867331 TI - Crossing the vasculopathy bridge from morphology to therapy: a single center experience. PMID- 10867332 TI - Efficacy and safety of pravastatin vs simvastatin after cardiac transplantation. AB - Prior studies of cardiac transplant recipients have shown that pravastatin reduces 12-month rejection and mortality after cardiac transplantation and simvastatin reduces 4-year mortality, low-density lipoprotein (LDL) cholesterol levels, and intimal thickening. In a 12-month observational study, cardiac transplant recipients received open-label pravastatin 40 mg (n = 42) or simvastatin 20 mg daily (n = 45) on an alternating basis from the time of transplantation. Lipid levels, safety, and post-transplant outcomes were compared. We found no significant differences in total LDL or high-density lipoprotein cholesterol, triglycerides, linearized infection or rejection rates, liver function tests, or immunosuppressant dosages between groups at 1, 3, 6, or 12 months. Rhabdomyolysis or myositis occurred only in patients on simvastatin (n = 6, 13.3%) with no episodes for patients on pravastatin (p = 0. 032). Survival at 12 months on an actual treatment basis was 97.6% for patients on pravastatin and 83.7% for those on simvastatin (p = 0.078). Immunosuppression-related deaths occurred in only 2.4% (1 patient) on pravastatin vs 15.6% (n = 7) on simvastatin (p = 0.06). Pravastatin and simvastatin resulted in comparable lipid profiles. Pravastatin use was however free from the high rates of rhabdomyolysis and myositis seen with simvastatin use. Pravastatin was additionally associated with a trend toward superior survival, attributable to fewer immunosuppression-related deaths. For safety and pharmacokinetic reasons, pravastatin should be considered the statin of choice after heart transplantation. PMID- 10867333 TI - Early alterations of myocardial blood flow reserve in heart transplant recipients with angiographically normal coronary arteries. AB - BACKGROUND: The evaluation of the coronary reserve provides valuable information on the status of coronary vessels. Therefore, we studied with positron emission tomography (PET) and 13N-ammonia the myocardial blood flow (MBF) reserve in heart transplant recipients free of allograft rejection and with angiographically normal coronary arteries early after heart transplantation (HTx). The MBF reserve was calculated as the ratio between MBF after dipyridamole injection and basal MBF normalized for the rate-pressure product. METHODS: Patients were studied within 3 months (group A, n = 12) or more than 9 months (group B, n = 12) after HTx. Five patients have been studied both during the early and late period after HTx. Results were compared to those obtained in 7 normal volunteers (NL). RESULTS: Group A recipients had a significantly lower dipyridamole MBF (in ml/min/100 gr of tissue) than that of group B recipients (142+/-34 vs 195+/-59, p<0.05). This resulted in a significant decrease in MBF reserve early after HTx (group A: 1.82+/- 0.33) and a restoration to normal values thereafter (group B: 2.52+/- 0.53 vs NL: 2.62+/-0.51, p = ns). Separate analysis of 5 patients studied twice is consistent with these results. CONCLUSION: This study shows that in heart transplant recipients free of allograft rejection and with normal coronary angiography, MBF reserve is impaired early after HTx. Restoration within one year suggests that this abnormality does not represent an early stage of cardiac allograft vasculopathy. PMID- 10867334 TI - Longitudinal study of vascular remodeling in coronary arteries after heart transplantation. AB - Cross-sectional studies by intravascular ultrasound (IVUS) in heart transplant recipients have suggested that vascular remodeling occurs in coronary arteries years after transplant. However, no reports describe vascular remodeling in the same cohort of patients studied prospectively using morphometric analysis (10 evenly spaced images obtained from a slow pullback from the left anterior descending coronary artery). Morphometric analysis better reflects total vessel anatomy compared with previously reported site (2 to 3 images) analysis. We reviewed 20 patients studied by IVUS at 2 months, 1 year, 2 years, and 3 years after heart transplant.Over time, the coronary artery luminal area decreased from baseline level of 12.0 mm(2) to a 3-year mark of 9.7 mm(2) (p = 0.02). Vessel shrinkage was seen in 16/20 patients. After an initial rise in intimal parameters (maximal intimal thickness, intimal index, and plaque area) from baseline to 1 year, we found a significant decrease in intimal parameters between Year 1 and Year 3 after transplant. For example, plaque area decreased from 2.05 mm(2) at 1 year post-transplant to 1.48 mm(2) by 3 years post-transplant (p = 0.05). In a majority of heart transplant patients, early intimal thickening in the first year post-transplant is accompanied by constrictive remodeling. Over the subsequent 2 years, further constrictive remodeling is seen despite a decrease in intimal area. PMID- 10867335 TI - Polymorphism of the transforming growth factor-beta 1 gene correlates with the development of coronary vasculopathy following cardiac transplantation. AB - BACKGROUND: Expression of transforming growth factor-beta1 (TGF-beta1) is central to vascular repair due to its effects on smooth muscle cell, monocyte/macrophage, leucocyte, and extracellular matrix accumulation and proliferation. Genetic polymorphism at position +915 of the TGF-beta1 gene determines the degree of cytokine production in response to injury. We investigated this allelic variation on the development of cardiac transplant-related coronary vasculopathy (CV). METHODS: Using sequence-specific primers to the TGF-beta1 gene region of interest, a polymerase chain reaction (PCR) and gel electrophoresis identified the genotype in 129 cardiac transplant recipients. An association was sought between the presence of a high- (GG) or low/intermediate-producing (CC/GC) genotype and the development of coronary vasculopathy diagnosed by coronary angiography. RESULTS: C allele carriers made up 10.9% of the recipient population but were significantly less likely to develop coronary vasculopathy (p = 0. 0361). Mean time to diagnosis was 1240.5 days in G homozygotes relative to 2266.5 days in C allele carriers (p = 0.002). Pre- and 1-year posttransplant clinical variables were equivalent between the 2 groups. Multivariate analysis identified the GG genotype (p = 0. 042, hazard ratio 3.01, [95% CI, 1.056-10.99]), donor age (p = 0.002, hazard ratio 1.063, [95% CI, 1.029-1.097]), and number of acute rejection episodes of grade 3 or greater in the first year (p = 0.029, hazard ratio 1.11, [95% CI, 1.05-1.26]) as significant predictors of vasculopathy. CONCLUSION: This study demonstrates a correlation between a high-producing TGF beta1 genotype and an earlier onset of cardiac-transplant coronary vasculopathy. This gives an important insight into the pathophysiology of cardiac transplant vasculopathy and suggests new treatment options. PMID- 10867336 TI - The role of percutaneous transluminal coronary angioplasty in heart transplant recipients. AB - OBJECTIVES: Review the acute and late results of percutaneous transluminal coronary angioplasty (PTCA) in heart transplant recipients and examine the factors predictive of restenosis. BACKGROUND: Coronary graft disease (CGD) is the main factor responsible for late graft loss. Medical treatment, surgical revascularization, or retransplantation gives only suboptimal results in this regard. Therefore, PTCA has been attempted in this situation. METHODS: More than 332 heart transplantations in our institution have been performed since 1992, the date of the first PTCA in our patients. We are currently in charge of 450 patients. All the characteristics, procedure-related information, and clinical outcome of patients needing PTCA were assessed by review of each patient's clinical records. All coronary angiograms were reviewed by an independent cardiologist. RESULTS: Since 1992, 53 coronary sites have been dilated in the course of 39 procedures in 29 patients. Indication for PTCA was asymptomatic angiographic coronary graft disease in 35 sites (64.8%), angina in 9 (16.6%), silent ischemia in 2 (3.7%), acute myocardial infarction in 1 (1.8%), and CHF in 7 (12.9%). Primary success (< 50% residual stenosis) was obtained in 50 (94.3%) of 53 lesions. No periprocedural death occurred. Procedural complications were 1 transient acute renal failure and 1 persistent bleeding at the puncture site. Six months restenosis rate (defined as percent stenosis > 50%) was 32.5% (14/43). Mean follow-up was 1.27 year +/- 1.2 (SD). Five deaths (17. 2%) occurred in follow-up and were all in relation to coronary graft disease. Mean time separating PTCA from death was 0.9 year +/- 1.3 (SD). We also sought to look at factors predictive of restenosis. By multivariate analysis, a positive recipient's serology for cytomegalovirus (CMV) before the graft was the only factor found protective against restenosis (odds ratio 22.4; confidence interval 1.1 to 443.4). CONCLUSION: PTCA in heart transplant recipients allows a high level of primary success with a low periprocedural-complication rate. Restenosis rate seems equivalent to restenosis rate in native coronary arteries. Mortality during follow-up is increased in this population and is the consequence of a high level of coronary events. Recipient positivity for CMV before the graft is associated with a protective effect from restenosis. PMID- 10867337 TI - Intrathymic immunomodulation in sensitized rat recipients of cardiac allografts: requirements for allorecognition pathways. AB - BACKGROUND: Intrathymic injection of alloantigen in the form of donor cells, soluble major histocompatibility complex (MHC) molecules, or MHC allopeptides induces donor-specific tolerance in a variety of acute allograft rejection models. We have previously shown that a single intrathymic injection of donor spleen cells into pre-sensitized rats abrogates accelerated (circa 24-hour) rejection and prolongs the survival of cardiac allografts to about 7 days. The present study was designed to investigate the mechanisms by which intrathymic administration of donor cells modifies the course of accelerated rejection. METHODS: Lewis RT1(1) (LEW) rats sensitized by transplantation with Wistar-Furth RT1(u) (WF) skin grafts received WF cardiac allografts 7 days later-a classic model of accelerated rejection. At the time of skin challenge, however, certain animals received intrathymic cell suspensions (either allogeneic or syngeneic) or donor-derived class I and/or class II MHC peptides. RESULTS: Control animals (sensitized by skin grafts but receiving no other treatment) rejected cardiac allografts within 24 hours. Intrathymic injection of WF splenocytes at the time of skin transplantation abrogated rejection at 24 hours and prolonged cardiac allograft survival to 6.6+/-0.6 days (p<0.001), whereas intrathymic administration of syngeneic (LEW) or allogeneic third party Brown Norway RT1(n) cells was ineffective in this regard. Intrathymic injection of gamma-irradiated donor cells marginally extended cardiac allograft survival to 3.0+/-0.9 days (p< 0.001), but the grafts were still rejected in an accelerated fashion. Intrathymic injection of donor-derived class I and/or class II MHC allopeptides at the same time period also failed to prolong cardiac allograft survival beyond 3 days. In the group receiving unmodified donor cells, elevated immunoglobulin M (IgM) and immunoglobulin G (IgG) allo-antibodies were found at the time of cardiac transplantation; this pattern was not observed with any other treatment. CONCLUSION: The superiority of non-modified donor spleen cells over gamma irradiated donor cells or donor specific allopeptides in modifying the course of accelerated cardiac rejection suggests that direct allorecognition is the dominant pathway initiating rejection in sensitized transplant recipients. Marked alterations in the antidonor IgM and IgG responses are associated with successful abrogation of accelerated rejection by thymic immunomodulatory mechanisms. PMID- 10867338 TI - Evidence that non-deletional mechanisms are responsible for inducing and maintaining unresponsiveness after intrathymic injection of non-professional antigen presenting cells. AB - INTRODUCTION: Intrathymic inoculation of donor alloantigen and concomitant immunosuppressive treatment can induce immune unresponsiveness to alloantigen. To examine the role of non-deletional mechanisms in the development of unresponsiveness, fractionated splenocytes were injected into only 1 lobe of the thymus. METHODS AND RESULTS: Untreated CBA (H2(k)) mice or controls pre-treated with anti-CD4 monoclonal antibody alone (on Day -28 and -27 relative to transplantation) acutely rejected C57BL/10 (H2(b)) cardiac allografts. Intrathymic inoculation of unfractionated splenocytes, resting B (rB) cells, or dendritic cells into both thymic lobes with the antibody resulted in indefinite survival of cardiac allografts. In contrast, when donor rB cells or dendritic cells were delivered into a single lobe of the thymus with the antibody, only rB cells induced indefinite prolongation of graft survival; unfractionated splenocytes or dendritic cells were markedly less effective. Mice that had 1 of the 2 thymic lobes removed were able to reject grafts even when treated with the antibody 27 days before transplantation. Therefore, T-cell export from 1 thymic lobe was sufficient to induce graft rejection. Finally, adoptive transfer of splenocytes from mice with long-term surviving primary grafts resulting from the intrathymic injection of rB cells significantly prolonged a graft from the same donor strain in a naive syngeneic recipient. CONCLUSION: Taken together, these data suggest that regulatory mechanisms generated by intrathymic injection of a non-professional antigen presenting cell, in this study donor rB cells, suppressed the rejection response mediated by T cells exported from the uninjected lobe. PMID- 10867339 TI - Active cell migration in retransplanted rat cardiac allografts during the course of chronic rejection. AB - BACKGROUND: Chronic allograft vasculopathy (CAV) is caused by the infiltration of host immune cells to a graft, but it has been technically difficult to monitor the movements of the cells in graft rejection. METHODS: We used a male-specific gene, SRY, as a marker to investigate the dynamics of host cells in a model of CAV in which immunosuppression was unnecessary and anti-male responses were practically negligible. Fluorescent-based real-time quantitative polymerase chain reaction (PCR) was adapted to estimate the fraction of host cells in a graft by the ratio of SRY to IL-2 gene. Using this technique, we studied the turnover and migration of host cells during the course of CAV progression by retransplanting female allografts from male to female or from female to male rats. RESULTS: We detected histologic CAV 60 days after retransplantation in allografts retransplanted to the F(1) progeny of donor x recipient on the 5th day, but not in those retransplanted on the 3rd day, regardless of the mismatches in the genders. Most of the initial infiltrating cells disappeared rapidly in both cases. The fraction of migrating cells from the second recipient, however, continuously increased in allografts developing CAV, and 60 days after retransplantation exceeded 50%, whereas it stayed at 5% to 15% in those not developing CAV. ED-1-positive macrophages/monocytes were likely candidates for the migrated cells. CONCLUSION: We have developed a simple method to measure the migration of host cells into a graft. This technique was useful, at least in certain rat strains, to investigate the cellular mechanisms of chronic cardiac allograft rejection. PMID- 10867340 TI - Monitoring pig-to-primate cardiac xenografts with live Internet images of recipients and xenograft telemetric signals: histologic and immunohistochemical correlations. AB - BACKGROUND: Monitoring pig-to-primate cardiac xenografts is often difficult in awake and uncooperative primates. We investigated the possibility of monitoring xenotransplantation through Internet broadcasting of (1) continuous video images of transplant recipients and (2) xenograft telemetric signals detected by an implanted device. The telemetric readings were later compared with histology and immunohistochemistry for signs of rejection. METHODS: Heterotopic baboon-to baboon (n = 2) and transgenic pig (human complement regulatory proteins CD59/DAF, n = 3; MCP, n = 1)-to-baboon transplants were performed with serial biopsies for hematoxylin-and-eosin staining and immunohistochemical detection of immunoglobulin M (IgM) and complement membrane attack complex (MAC) deposition. Baboon recipients were continuously monitored with a QuickCamPro digital camera, whereas grafts were monitored with a Data Science International implantable telemetric system. Video images and telemetric signals were broadcast over the Internet through a laptop computer. RESULTS: Baboon allografts remained healthy until explant on Day 14, whereas pig xenografts were rejected on Day 5, 6, 7, and 11. Telemetry of allografts and xenografts documented regular rhythm with an average heart rate of 80 to 120, but xenografts developed bradycardia and widened/dampened QRS complexes 24 to 48 hours before graft loss. Continuous video monitoring of recipient activities was vital in differentiating between graft arrhythmias and telemetric artifacts. Allograft biopsies showed little cellular infiltrate, whereas xenograft biopsies showed increasing IgM and MAC deposition, with extensive thrombi and myocardial damage 24 hours before cessation of cardiac activities. CONCLUSIONS: Combined video surveillance of recipient activities and graft telemetric signals is a useful method to continuously monitor abdominal cardiac grafts in large, uncooperative, awake primates. QRS-complex widening associated with progressive bradycardia correlated with histologic and immunohistochemical evidence of xenograft rejection. PMID- 10867341 TI - Predictors of quality of life in women with heart failure. SOLVD Investigators. Studies of Left Ventricular Dysfunction. AB - BACKGROUND: Two and one half million women have heart failure (HF). Yet little is known about quality of life (QOL) in this population and the factors influencing it. Given the importance of QOL as an outcome of care, we conducted a study to evaluate predictors of QOL in women with HF. METHODS: Using baseline QOL data collected in the Studies of Left Ventricular Dysfunction (SOLVD) trials, we studied predictors of QOL in 691 women with HF. Univariate, bivariate, and multiple regression analyses were used. Potential predictors included age, education, tobacco use, social isolation, life stresses, comorbidity index, New York Heart Association (NYHA) class, HF symptoms, etiology, and medications. We measured global QOL and QOL dimensions of physical function, emotional distress, and social and general health. RESULTS: Women were older (61+/-10.5 years), predominantly Caucasian (75%), and their mean ejection fraction was 0.27 (+/ 6.51). Variables with the strongest relationship to QOL included dyspnea, NYHA class, and life stresses. As dyspnea, life stresses, and NYHA class increased, QOL decreased. Additionally, smoking behavior and vasodilator use was associated with decreased QOL. Heart failure etiology of ischemic origin was associated with decreased social life satisfaction, and use of digitalis was predictive of increased social life satisfaction. Finally, increasing age was related to an increase in general life satisfaction. CONCLUSION: Symptom amelioration, which may improve functional ability, has the greatest potential for increasing QOL in women with HF. Programs to increase physical activity in women with HF should be developed and tested. Finally, clinicians may need to optimize HF medications in women. PMID- 10867342 TI - The impact of cocaine on the donor heart: a case report. AB - With a limited supply of donor hearts in the United States and a prevalent history of cocaine abuse among potential heart donors, the question of transplanting the hearts of cocaine users presents a dilemma to the surgeon. We report a patient who died of the acute right ventricular failure of a heart from a donor with a history of binge drinking and cocaine abuse and who had sustained traumatic brain death. The donor's serum was positive for cocaine prior to transplantation, and autopsy findings were consistent with cocaine cardiomyopathy. This case illustrates the importance of accurate donor history and toxicologic screen prior to heart transplantation and suggests that hearts of cocaine users should not be transplanted, especially in a setting of traumatic brain death. PMID- 10867343 TI - Successful management of profound thrombocytopenia after lung and heart-lung transplantation. PMID- 10867344 TI - Durable cure of mucormycosis involving allograft and native lungs. AB - Among the spectrum of fungi causing disease in lung allograft recipients, fungi in the order Mucorales represent uncommon pathogens. Lung transplant patients, however, often possess more than one risk factor for development of life threatening mucormycosis. We describe a unique case of pulmonary mucormycosis involving both the allograft and the native lungs, in a single lung transplant recipient with steroid-induced diabetes. Extended intravenous amphotericin B and oral fluconazole therapy, reduction of immunosuppression, and blood glucose control achieved a durable cure without the need for surgical intervention. Early diagnosis with prompt initiation of multiagent antifungal therapy, prolonged continuation of antifungal therapies, and amelioration of contributing conditions are important elements of the treatment strategy that led to successful resolution of the infection. PMID- 10867345 TI - Resilience and protective factors in the lives of adolescents. PMID- 10867346 TI - Preliminary study examining relationship between family environment and resting mean arterial pressure in African-American youth. AB - This investigation examined the relationship between family environment and mean arterial pressure in a sample of African-American youth. Completed data were collected from 46 adolescents attending an inner-city junior high school. Blood pressure measurements were assessed in a seated position with a Dinamap 1846 Vital Signs Monitor. To assess the additive effects associated with the family environment, a composite risk score (cohesion, conflict, control, and organization subscales of the Family Environment Scale) was computed. Regression analyses indicated that the cumulative risk associated with the family environment was predictive of resting mean arterial pressure. PMID- 10867347 TI - Utilization of electronic communication (E-mail) with patients at university and college health centers. AB - PURPOSE: To examine the utilization and potential uses and problems of electronic communication with patients. METHODS: University and college health centers were surveyed about the type of utilization and policies of electronic communication with patients. The survey group consisted of 99 health centers predominantly serving students representing small-, medium-, and large-sized public and private colleges and universities. Eighty-nine health centers completed the survey. RESULTS: Of the responding health centers, 63.6% use some form of electronic communication with patients. Twenty-seven percent of the health centers give out some form of medical advice via E-mail or the Internet; 14.7% give out some laboratory results via E-mail; 3.4% make appointments via E-mail; and 63.6% give out administrative advice by E-mail. While there was consistent concern expressed about confidentiality and security, only five health centers had a policy about electronic communication. Uses were most common in nonclinical areas but did include health education, Web sites, medical advice, laboratory results, appointment-making or confirmation, and contacting hard-to-reach patients including those studying abroad. CONCLUSIONS: While electronic communication with patients was common, provision of direct medical advice was less common. Issues receiving little attention include determining the types of electronic communication that is acceptable to staff and students, determining the level of security of their current information system, educating staff about confidentiality and security issues, and establishing a comprehensive policy regarding electronic communication with patients. PMID- 10867348 TI - Contextual influences on school provision of health services. AB - PURPOSE: To examine contextual factors that may facilitate or impede the provision of school health services. METHODS: Using a composite database derived primarily from the National Longitudinal Study of Adolescent Health, we used logistic regression to examine how selected characteristics of communities, schools, and state-level policies are related to the provision of specific health services by high schools. RESULTS: Schools whose students experienced more health risks were generally more likely to provide related services than schools whose students experienced fewer risks. State policies and requirements for health related programs and services were associated with greater school-based provision of services. Availability of health care services within the community was associated with a reduced likelihood that schools provided similar services on site; however, for some health services, the reverse was true. In general, more affluent communities were more likely to provide school health services than less affluent communities. Public schools were more likely to offer health services than private schools. CONCLUSIONS: Certain characteristics of communities, schools, and state-level policies are associated with the provision of school health services. These contextual factors appear to operate by creating a demand for services and by creating the opportunity for schools to provide health services. PMID- 10867349 TI - Family planning clinic patients: their usual health care providers, insurance status, and implications for managed care. AB - PURPOSE: To understand the extent to which family planning clinic patients have health insurance or access to other health care providers, as well as their preferences for clinic versus private reproductive medical care. METHOD: An anonymous self-report questionnaire was administered at three Planned Parenthood clinics in Los Angeles County to 780 female patients aged 12-49 years. Dependent variables included insurance status, usual source of care, and a battery of questions regarding the importance of confidentiality. RESULTS: A total of 356 adolescents (aged 12-19 years) and 424 adults (aged 20-49 years) completed the survey in 1994. Fifty-nine percent of adolescents and 53% of adults had a usual source of care other than the clinic. The majority of each group reported some degree of continuity of care in their usual provider setting. Nearly half (49%) of all adolescents had health insurance compared with 27% of adults. Adolescents cited not wanting to involve family members as the primary reason for not using their usual providers, whereas adults were more likely to cite being uninsured. The majority of both adult and adolescent patients indicate they would prefer the clinic over private health care if guaranteed health care that was free, confidential, or both. CONCLUSION: Despite many patients' having health insurance and other sources of health care, family planning clinics were primarily chosen because of cost and confidentiality. Their reasons for preferring clinics may continue despite changes in access to insurance or efforts to incorporate similar reproductive services into mainstream health care provider systems. Making public or private health care funds available to family planning clinics through contracts or other mechanisms may facilitate patients' access to essential services and reduce potential service duplication. PMID- 10867350 TI - Cumulative effect of family environment on psychiatric symptomatology among multiethnic adolescents. AB - PURPOSE: To examine the influence of family adversity and support on levels of psychiatric symptomatology in Hawaiian and non-Hawaiian adolescents. METHOD: More than 4000 students from five high schools in Hawaii completed a survey during the 1992-1993 school year about their family environment and mental health. The response rate was approximately 60%. Logistic regression analyses were performed and responses for Hawaiians and non-Hawaiians were compared. RESULTS: Hawaiian adolescents experienced significantly more adversity than their non-Hawaiian counterparts. The cumulative effect of family adversity had a greater effect on psychiatric symptomatology than any single indicator. Family support reduced the risk for internalizing symptoms, particularly for Hawaiian adolescents. The influence of family support was less clear for externalizing symptoms, increasing the risk for some adolescents and decreasing the risk for others. CONCLUSION: We identified strong associations between family adversity and levels of psychiatric symptomatology. We found that Hawaiian adolescents are at increased risk for psychiatric symptomatology, given the presence of family adversity and the effect of reduced family support. However, risk was also substantial for non-Hawaiians. Clinicians need to assess the family environment routinely and implement family oriented interventions. PMID- 10867351 TI - Perceived parental monitoring and health risk behaviors among urban low-income African-American children and adolescents. AB - PURPOSE: To examine gender and age differences among urban, low-income, African American children and adolescents in perceived monitoring by their parents, and the association of perceived parental monitoring with family characteristics, health risk behaviors, and risk perceptions. METHODS: Three cross-sectional surveys were conducted in 1992 (n = 455), 1994 (n = 355), and 1996 (n = 349). Respondents aged 9-17 years were recruited from low-income urban areas including public housing communities and associated recreation centers. Both multivariate analysis of variance and correlation analysis were performed. RESULTS: Low levels of perceived parental monitoring were associated with participation in several health risk behaviors, including sexual behavior, substance/drug use, drug trafficking, school truancy, and violent behaviors. Females perceived themselves to be more monitored than did males. In general, the perceived parental monitoring tended to decrease with advancing age of the youth. CONCLUSIONS: The strong inverse correlation between perceived parental monitoring and adolescent risk behavior suggests that parental monitoring initiatives may be an effective intervention tool. Longitudinal studies are needed to determine the long-term relationship between perceived parental monitoring and adolescent risk involvements. PMID- 10867352 TI - Impact of perceived parental monitoring on adolescent risk behavior over 4 years. AB - PURPOSE: To determine the stability of perceived parental monitoring over time and its long-term effect on health risk behaviors among low-income, urban African American children and adolescents. DESIGN: Prospective, longitudinal follow-up (4 years). SUBJECTS: A total of 383 African-American youth aged 9-15 years at baseline recruited from nine recreation centers serving three public housing communities in an Eastern city. OUTCOME MEASURES: A six-item measure assessing perceived parental monitoring and an 11-item self-reported measure assessing unprotected sex, drug use, and drug trafficking were administered at baseline and at regular intervals over the subsequent 4 years. ANALYSIS: Concordance was assessed by Pearson correlation coefficients at the level of scale and by kappa scores at the level of items. The association between the monitoring score and risk involvement was determined by stepwise multiple regression analysis including parental monitoring, age, gender, intervention status, and two-way interactions between parental monitoring and age, gender, intervention status as independent variables. RESULTS: The perception of being monitored demonstrated consistency over time. Parental monitoring was inversely correlated with all three targeted risk behaviors cross-sectionally and prospectively. CONCLUSION: These data provide evidence for an inverse relationship between perceived parental monitoring and risk involvement cross-sectionally and longitudinally. These data support the long-term effect of perceived parental monitoring on risk behaviors among urban, low-income African-American children and adolescents. Coupled with some evidence suggesting that directed interventions might be able to increase parental monitoring, this study provides a solid platform for reinforcing the importance of parental monitoring and directing intervention efforts at strengthening parental monitoring to reduce adolescent risk behaviors. PMID- 10867353 TI - Three years of a clinical practice guideline for uncomplicated pelvic inflammatory disease in adolescents. AB - PURPOSE: To study the effect of continued use of a clinical practice guideline (CPG) on the course of admissions for uncomplicated pelvic inflammatory disease (PID) over 3 consecutive fiscal years (FY). METHODS: Medical charts, computerized laboratory records, and hospital charge data were reviewed for 165 consecutive inpatient admissions of adolescents meeting clinical criteria for PID during FY 1994, 1995, and 1996. Data were analyzed to compare demographics, clinical variables, length of stay (LOS), and hospital charges (total, nursing, and pharmacy) across the three FYs. RESULTS: Of admissions for clinical PID, 65% had a discharge diagnosis of PID. Of those, 90% were uncomplicated PID. Among admissions with a discharge diagnosis of uncomplicated PID, reductions were seen in mean LOS (3.75 days in FY 1994 vs. 3.24 days in FY 1995 vs. 3.08 days in FY 1996; p =.047), proportion of admissions lasting longer than 3 days (48% vs. 24% vs. 20%; p < or =.022), and mean pharmacy charge ($946 vs. $806 vs. $731; p =.002). For all admissions to CPG, mean LOS, proportion of prolonged admissions, and mean total and pharmacy charges also decreased over the first 2 years but increased in FY 1996. More patients in FY 1996 met the three major clinical criteria plus at least one additional criterion (76% in FY 1996 vs. 26% in FY 1994 and 53% in FY 1995; p <.0005) and had pelvic ultrasounds (80% in FY 1996 vs. 56% in FY 1994 and 45% in FY 1995; p < or =.001) than in other FYs. CONCLUSIONS: Continued use of a CPG can reduce hospital LOS, charges, and prolonged admissions of adolescents with uncomplicated PID. Over 3 years, variations in clinical practice such as admitting sicker patients may attenuate the effect of the CPG. PMID- 10867354 TI - The relationship between anger-coping styles and lifestyle behaviors in teenagers. AB - PURPOSE: To investigate the relationship between anger-coping styles (anger expression and anger suppression) and lifestyle behaviors (physical activity and consumption of alcohol, cigarettes, and caffeine) in adolescents. METHODS: A sample of 411 adolescents (198 males: 101 white, 97 black; 213 females: 101 white, 112 black) aged 13-20 years (mean age 15.6 years) completed the Anger Expression Scale and brief self-report questionnaires assessing physical activity (weekly amount, comparison with peers) and consumption of alcohol (frequency and amount over the past 2 weeks), cigarettes (amount over past 2 weeks), and caffeine (from coffee and soda over past week). RESULTS: Correlational and Chi square analyses showed teenagers high in anger suppression reported consuming alcohol more frequently, spending fewer hours per week in aerobic activity, and being less physically active than their peers. Teenagers high in anger expression reported consuming more caffeinated soda and coffee. CONCLUSIONS: Results suggest that excessive anger suppression or expression may be associated with an imprudent lifestyle relatively early in life. PMID- 10867356 TI - Health care for incarcerated youth. Position paper of the Society for Adolescent Medicine. PMID- 10867355 TI - Case report: treatment of catatonia in an adolescent male. PMID- 10867358 TI - Biphasic survival response to amlodipine after myocardial infarction in rats: Association with cardiac vascular remodeling. AB - In clinical studies, calcium channel blockers have been found to cause adverse cardiovascular reactions after myocardial infarction; however, such effects appear limited to short-acting agents. Thus, our aim was to evaluate the response to a long-acting calcium channel blocker, amlodipine, in terms of both survival and, cardiac and vascular remodeling after infarction. One week after permanent coronary occlusion, rats were randomized to no treatment or daily amlodipine (5 mg/kg) continued for up to 9 months. Amlodipine resulted in improved survival at 200 days (65% versus 26%; p < 0.05), but no difference at 9 months. However, rats with large infarcts died earlier than untreated animals, while those with smaller infarcts died later or survived for nine months. Amlodipine produced no difference in collagen content in non-infarcted tissue or myocyte cross-sectional area versus untreated hearts; however, scar length was increased. In addition, amlodipine was associated with vascular remodeling; muscle:lumen ratio increased in non-infarcted myocardium as did perivascular fibrosis. Vessels within the scar had reduced lumen area because of smooth muscle proliferation. We also examined infarcted hearts subjected to one week of intravenous amlodipine (1 mg/kg) initiated before occlusion and examined three weeks later. In this study, amlodipine exacerbated muscle proliferation in infarct vessels and was associated with less scar collagen. The vascular remodeling associated with amlodipine treatment is considered unfavorable and so the adverse survival for rats with large infarcts was no surprise. However, the prolonged survival associated with smaller infarcts raises the possibility that these vascular changes, under certain circumstances, are beneficial. PMID- 10867357 TI - Impact of delayed reperfusion of myocardial hibernation on myocardial ultrastructure and function and their recoveries after reperfusion in a pig model of myocardial hibernation. AB - This study examined the effect of delayed reperfusion of myocardial hibernation from 24 hours to 7 days on myocardial ultrastructural and functional changes and their recoveries after reperfusion. BACKGROUND: We have previously shown in pigs that after reperfusion the functional and structural alterations in short-term myocardial hibernation which was reperfused in 24 hours can recover in 7 days. The effect of delayed reperfusion of hibernating myocardium on the extent and severity of cellular and extracellular structural changes of hibernating myocardium, and their recoveries after reperfusion is not known. METHODS AND RESULTS: A severe LAD stenosis was created in 27 pigs, reducing resting flow by 30-40% immediately after placement of the stenosis and producing acute ischemia as evidenced by regional lactate production, a decrease in regional coronary venous pH, reduced regional wall thickening (from 38.5 +/- 5.1% to 10.4 +/- 8.0%) and a 33% reduction of regional oxygen consumption. The stenosis was maintained either for 24 hours in 9 pigs (group 1) with LAD flow of 0.65 +/- 0.13 ml/min/g (38% reduction), or for 7 days in 17 pigs (group 2) with LAD flow of 0.67 +/- 0.14 ml/min/g (36% reduction). There were no differences (p = NS) in the reduction of wall thickening, rate-pressure product, lactate production, or regional oxygen consumption between group 1 and group 2. Quantitative morphometric evaluation of the ultrastructure on electromicrographs revealed a greater decrease in sarcomere volume and a higher incidence of myocytes with reduced sarcomere volume in 7-day than in 24-hour hibernating regions (53 +/- 19% versus 33 +/- 14%, p < 0.05). Patchy myocardial necrosis with replacement fibrosis was common, but 6 of the 18 pigs had no myocardial necrosis or replacement fibrosis in the 7-day hibernating group, and 4 of 9 pigs had no patchy myocyte necrosis in the 24 hour hibernating group. In 6 pigs in group 1 in which the stenosis was then released and hibernating myocardium reperfused in 24 hours, regional wall thickening recovered to 30 +/- 6% (p = NS compared to baseline) after one week of reperfusion. In 12 pigs in group 2 in which the stenosis was released and hibernating myocardium reperfused in 7 days, regional wall thickening recovered slowly, from 10.1 +/- 7.2% to 18.1 +/- 8.3% at one week (n = 5) and to 28.0 +/- 3.6% at 3-4 weeks of reperfusion (n = 7, p < 0.05 compared to baseline). Similarly, the sarcomere volume or myofilament recovered significantly (p < 0.01) and was not different compared to the normal region (p = NS) in the 24-hour hibernating region of group 1, but the recovery was much slower and was incomplete at 4 weeks (p < 0.01) compared to baseline in the 7-day hibernating region of group 2. Recovery of regional wall thickening correlated with ultrstructural recovery (p < 0.01). By multivariate stepwise regression analysis, the degree of LAD flow reduction, the extent of fibrosis, and myofilament loss were independent predictors of the extent of functional recovery. CONCLUSIONS: In a porcine model of myocardial hibernation with myocardial hypoperfusion, systolic dysfunction, and metabolic adaptations, a longer period of myocardial hibernation with delayed reperfusion was associated with more severe abnormalities of myocytes. an increasing interstitial fibrosis, and more protracted myofibrillar and functional recoveries after reperfusion. The extent of functional recovery is related to the degree of coronary flow reduction, the severity of the ultrastructural changes, and the extent of interstitial fibrosis. PMID- 10867359 TI - Structural remodeling of the left atrial appendage in patients with chronic non valvular atrial fibrillation: Implications for thrombus formation, systemic embolism, and assessment by transesophageal echocardiography. AB - Left atrial appendage (LAA) is frequently the site of thrombus formation in patients with chronic atrial fibrillation (AF). Transesophageal echocardiography and hematologic studies have identified blood flow stasis (spontaneous echogenic contrast) and abnormal coagulation (increased serum fibrinogen) as important predisposing factors to formation of LAA thrombi. However, the third component of the Virchow's triad, i.e., endothelial abnormalities, has not been adequately studied. Accordingly, we studied, at necropsy, the LAA morphology in 46 hearts of patients with (n = 22) and without (n = 24) chronic AF. Compared to patients without AF, those with AF had significantly larger LAA volumes (1.7% 1.1 vs. 5. 4% 3.7 mL, p = 0.0002), and larger luminal surface area of the bisected LAA (4.4% 1.8 vs. 7.1% 4.5 cm(2), p = 0.01). However, both the absolute and relative surface area of the transected pectinate muscles were reduced in patients with AF (2.6% 1.1 vs. 1.8% 1.0 cm(2), p = 0.02 and 38% 15 vs. 21% 14%, p = 0.0003). In addition, in most patients (73%) with chronic AF, the LAA showed significant endocardial thickening with fibrous and elastic tissue (endocardial fibroelastosis) compared to those without AF (13%, p < 0.0001). Endocardial fibroelastosis resulted in a smooth LAA luminal surface and encased the pectinate muscles. These findings suggest that LAA remodeling (dilation, stretching, and reduction in pectinate muscle volume, as well as endocardial fibroelastosis) occurs frequently in chronic AF and may contribute to the increased risk of thrombus formation and systemic embolism. Additionally, the information may have relevance in interpreting transesophageal echocardiographic images of the LAA in patients with chronic AF. PMID- 10867360 TI - Backscattered electron imaging: A new method for the study of cardiomyocyte architecture using scanning electron microscopy. AB - Scanning electron microscopy (SEM) with secondary electron emissions is useful for the study of cardiomyocyte architecture, however, the information is limited from the cell surface. Whereas backscattered electron (BSE) emission can give a high-resolution image of the specimen's intracellular structure after heavy metal staining. In this study, we applied BSE imaging analysis to the study of the arrangement of cardiomyocytes in the myocardium. The tissue specimens from a normal fresh monkey heart, normal human heart obtained at autopsy, and surgically resected tissue from a patient with old myocardial infarction in the left ventricular aneurysmectomy were used. The tissue specimens were fixed in neutral formalin, treated with NaOH and then stained with Gomori's silver methenamine reagent followed by tannic acid and osmium tetroxide. After dehydration and drying, the specimens were coated with carbon and examined by SEM with a BSE detector. In the tissue preparations, the A bands of sarcomeres were selectively stained with silver so that the arrangements of subsarcolemmal myofibrils and the intercalated discs were clearly seen in the BSE images. In the left ventricular aneurysmal walls of old myocardial infarction, atrophied cardiomyocytes with disarray of subsarcolemmal myofibrils were observed. The results strongly suggest that BSE images are further applicable to the study of the architecture of cardiac myocytes and their branches, and the arrangement of intracellular myofibrils in various diseased myocardium. PMID- 10867361 TI - Upregulation of adhesion molecules and class I HLA in the myocardium of chronic chagasic cardiomyopathy and heart allograft rejection, but not in dilated cardiomyopathy. AB - The immunohistochemical expression of adhesion molecules and class I HLA in chronic chagasic cardiomyopathy were compared with heart allograft rejection and dilated cardiomyopathy, to obtain new knowledge on the occurrence of autoimmunity and inflammation in the pathogenesis of chronic chagasic cardiomyopathy. Semiquantitative immunohistochemistry was performed for CD8+ T cells, ICAM-1, VCAM-1, LFA-1, and class I HLA in frozen sections of myocardial biopsies from patients presenting chronic chagasic cardiomyopathy (group I, n = 12), heart allograft rejection (group II, n = 9) or dilated cardiomyopathy (group III, n = 9). A high mean number of CD8+ T cells/mm(2) was present in group I (18.26) and group II (28.60), but not in group III (0.83). The frequency of high expression for ICAM-1 and VCAM-1 on the endothelial and interstitial cells, and for class I HLA on the cardiomyocytes was greater in group I (100%, 33.3%, and 83.3%, respectively) and group II (100%, 66.7%, and 77.8%, respectively), compared to group III (66.7%, 0%, and 0%, respectively). ICAM-1 and VCAM-1 probably participate in the development of the lymphocytic inflammatory infiltrate present in chronic chagasic cardiomyopathy, as seen in heart allograft rejection. The overexpression of adhesion molecules and the induction of class I HLA on the cardiomyocytes are probably related to the high cytokine levels at the inflammatory sites in chronic chagasic cardiomyopathy. Although the induction of class I HLA on the cardiomyocytes is consistent with an autoimmune reaction, it should not be considered as irrefutable evidence for autoimmunity in chronic chagasic cardiomyopathy. The differential expression of adhesion molecules and class I HLA in dilated cardiomyopathy compared to chronic chagasic cardiomyopathy suggests differences in the pathogenesis of these cardiomyopathies. PMID- 10867362 TI - Molecular screening by polymerase chain reaction detects panleukopenia virus DNA in formalin-fixed hearts from cats with idiopathic cardiomyopathy and myocarditis. AB - Viral myocarditis has been suggested as an etiology for cardiomyopathy in several mammalian species. Myocarditis and idiopathic cardiomyopathy have been reported in the domestic cat, although a viral etiology has not been demonstrated. Because of the continuing interest in the potential relationship between viral myocarditis and cardiomyopathy, we evaluated hearts from cats with spontaneous, idiopathic cardiomyopathy for viral genomic material within myocytes by polymerase chain reaction, and for the presence of myocarditis by light microscopy. Thirty-one (31) formalin-fixed hearts from domestic cats who died of idiopathic cardiomyopathy were randomly selected from pathology archives. Seventeen (17) formalin-fixed hearts from healthy cats were similarly selected as normal controls. The polymerase chain reaction (PCR) was used to evaluate myocardial tissue for the presence of viral genome from feline panleukopenia virus, herpes virus, calici virus, and corona virus. Hearts were examined using light microscopy for histologic evidence of myocarditis according to the Dallas criteria. Panleukopenia virus was identified by PCR in 10 of 31 cats with cardiomyopathy but in none of the controls. Neither cardiomyopathic or control cats tested positive by PCR for herpes virus, calici virus, and corona virus. Myocarditis was detected by histologic examination in 18 of 31 cardiomyopathic cats and in none of 17 control cats. Myocarditis and or feline panleukopenia virus genome was detected in felines with idiopathic hypertrophic, dilated, and restrictive cardiomyopathy, suggesting a possible role of viral infection and inflammation in the pathogenesis of cardiomyopathy in this species. PMID- 10867364 TI - Editorial. PMID- 10867363 TI - Situs inversus totalis and single coronary ostium: A coincidence or a pattern? AB - Situs inversus totalis and single coronary ostium are rare congenital anomalies, and no ontogenic connection has been described between them. Only three cases of association of single coronary ostium and situs inversus have been reported in the literature, all found on angiography. Here we present the first case of this association discovered at autopsy. Based on the apparently higher than expected frequency of this finding, an underlying pathologic connection between these conditions is proposed. PMID- 10867365 TI - Effect of quinalphos and cypermethrin exposure on developing blood-brain barrier: role of nitric oxide. AB - Effect of low-level exposure of quinalphos (QP) and cypermethrin (CP) on the blood-brain barrier (BBB) permeability to macromolecular tracers, Evans blue (EB) and horseradish peroxidase (HRP) was studied in developing rat pups. Ten-day-old rat pups were daily exposed to QP and CP at a dose of approximately 1/50th of adult LD50 through oral intubation, upto postnatal day 17 (PND). Functional integrity of the BBB was assessed by measuring the brain uptake index (BUI) of HRP and by visually grading the brains of control and treated rat pups for the staining of EB. Our results have demonstrated a significant increase in the BUI for HRP (204 and 254%) and have also shown a significant amount of EB staining in QP and CP exposed brains, respectively, as compared to the age-matched controls. Studies carried out with the nitric oxide synthase (NOS) inhibitor L-NAME (30 mg/kg, i.p., on alternate days from PND 10-17) have provided significant protection against the QP-induced increase in the BBB permeability, suggesting the possible involvement of NO in the barrier disruption. Microvessel acetylcholinesterase activity was also inhibited (53%, P<0.001) in QP-exposed rat pups only, with no change observed in CP-exposed microvessels. However, membrane fluidity was found to be decreased in both QP (18%, P<0.05) and CP (15%, P<0.05) exposed microvessels compared to controls. It is evident from the study that QP and CP exposure during early postnatal period causes significant impairment in the development and maturation of the BBB that may have adverse consequences on the normal brain functioning with long-term neurotoxic effects. PMID- 10867366 TI - Cadmium-induced changes in cation-osmotic haemolysis in rats. AB - Erythrocyte microrheology changes were measured using cation-osmotic haemolysis (COH) in healthy rats and rats after 6 and 12 months of cadmium ingestion. Using COH properties of two membrane constituents, spectrine membrane skeleton and membrane bilayer, were studied. COH in rats after 6 months of cadmium ingestion was significantly lower only in the area with lower ionic strength. However, longer intake of cadmium (12 months) induced decrease of COH in both areas, with lower and also with higher ionic strength. The relation between cation-osmotic haemolysis and erythrocyte deformability is being discussed. PMID- 10867367 TI - Dinoflagellates from marine algal blooms produce neurotoxic compounds: effects on free calcium levels in neuronal cells and synaptosomes. AB - In this report, evidence is presented that the marine unicellular eukaryotic dinoflagellates can cause neurotoxicity very likely by an increase in intracellular free calcium ions ([Ca(2+)](i)). Determinations of the effects of culture supernatants from different clones of the dinoflagellate Alexandrium sp. isolated from algal blooms on the viability of rat primary neuronal cells revealed that all clones tested were toxic for these cells. In addition, all Alexandrium clones tested, except for A. ostenfeldii BAH ME-141, were found to be toxic for rat pheochromocytoma PC12 cells. No toxicity was observed for culture supernatants from Gonyaulax and Coolia monotis. Calcium ions are important in the process of apoptotic cell death; our studies revealed that the dinoflagellate supernatants from A. lusitanicum K2, A. lusitanicum BAH ME-091, and A. tamarense 1M caused an increase in [Ca(2+)](i) levels in both PC12 cells and primary neuronal cells. These dinoflagellate supernatants, as well as the A. tamarense ccmp 115 supernatant, were found to cause also an increase in free calcium concentration in isolated synaptosomes. Our results suggest that the neurotoxic effects of certain dinoflagellate supernatants may be associated with disturbances in [Ca(2+)](i) levels. PMID- 10867368 TI - Cyanobacteria as a biosorbent of heavy metals in sewage water. AB - The effect of sewage water on some physiological activities of cyanobacteria was studied. Metal-tolerant cyanobacterium (Nostoc linckia) and metal-sensitive (Nostoc rivularis) were grown at three levels of sewage water (25, 50 and 75%). The growth rate showed significant stimulation in low and moderate levels (50% for N. linckia and 25% for N. rivularis). Not only the number of cells was elevated but also, the time required to reach the exponential and the stationary phases was reduced. Also, low levels of sewage water increased chl.a content, photosynthetic O(2)-evolution, respiration and protein content. Similarly, heterocyst frequency as well as nitrogenase activity were increased in cyanobacteria grown at low and moderate levels (25 and 50% sewage). On the other hand, the high level of waste (75%) reduced growth and metabolic activities of the two species. N. linckia accumulated about 30-fold of Zn and ten-fold of Cd than those of growth medium (50% sewage water). Also, N. rivularis accumulated about ten-fold of Zn and two-fold of Cd. The distribution of Cd and Zn in cells were investigated. About 65-60% of Cd or Zn were found in pellets (sediment) as insoluble form in the two species. The soluble form (cytosolic fraction) after being fractionated on sephadex G-(75-100) revealed two peaks with molecular weights of 70-75 and 40-45 kDa. These peaks were in coincidence with Cd and Zn maxima. Nostoc rivulais showed more sensitivity to heavy metals than N. linckia, and accumulated less amount of metal-binding proteins. Nostoc linckia seems to be tolerant to heavy metals (Zn and Cd) and is able to accumulate this metal by adsorption on the pellets (cell surface) and/or through sequestration via metal binding protein. Therefore it can be recommended it to be employed in the purification of waste contaminated with these heavy metals. PMID- 10867369 TI - Comparative characterization of CHCl(3) metabolism and toxicokinetics in rodent strains differently susceptible to chloroform-induced carcinogenicity. AB - A comparative kinetic study in B6C3F1 mice, Osborne-Mendel (OM) and Sprague Dawley (SD) rats has been undertaken with the major aim to determine the extent of covalent binding of chloroform reactive metabolites produced in vivo through oxidative and/or reductive metabolism in the target organs of chloroform carcinogenicity. Some additional kinetic observations of chloroform biotransformation were also collected comparatively. Expiration of [14C]-CO(2) showed that chloroform metabolism went to saturation in all tested rodent strains. In the B6C3F1 mouse maximal rates of approximately 135 umol [14C] CO(2)/kg b.w./h were reached at a dose of approximately 150 mg/kg, while in the two rat strains saturation occurred at a dose of approximately 60 mg/kg, with a maximal rate of approximately 40 umol [14C]-CO(2)/kg b.w./h. At doses of 150-180 mg/kg b.w., limited differences were found in the distribution and elimination of [14C]-chloroform in the liver and kidney. Species differences have been found in the kinetics of alkali-extractable radioactivity in the blood. The levels of adducts of electrophilic intermediates with the polar heads (PH) of phospholipids (PL) showed a limited variability accross the rodents tested and did not correlate with the species and organ susceptibility to chloroform carcinogenicity. The levels of adducts of radical intermediates with the fatty acyl chains (FC) of PL were much lower than the PH adducts in all the samples analyzed; at the carcinogenicity bioassay doses, statistically significant levels of hepatic FC adducts were present only in the B6C3F1 mouse, where chloroform is hepatocarcinogenic. The observations in the rat kidney were suggestive of the formation of electrophilic reactive metabolites, presumably different from phosgene and associated with an initial chloroform reduction. PMID- 10867370 TI - Assessment of multipathway exposure of small children to PAH. AB - The aim of study was to assess the uptake of polycyclic aromatic hydrocarbons (PAH) by children living in a city and its effect on 1-hydroxypyrene (1-OHP) excretion. Two groups of children (n=11 and 13; 3-6 years old) were chosen: (1) a group from a kindergarten situated near a road with a high traffic density ('polluted' area); (2) a group from a kindergarten situated in a green zone ('non polluted' area). Food consumption was recorded in all children and PAH uptake from foodstuffs was estimated. Ambient air samples were collected on the playground and indoor of kindergartens during 3 days in summer 1997. Soil samples were collected on the playground. Urine samples were collected in the morning and in the evening. Mean outdoor total PAH concentration (sum of 12 individual PAH) in 'polluted' area was 12 times higher than that in 'non polluted' area (22.9 vs. 1.9 ng/m(3)). However, indoor concentrations were similar (3.0 vs. 2.1 ng/m(3)). The same trend was observed for pyrene concentrations. The contribution to the total pyrene absorbed dose from food consumption (estimated daily absorbed dose of 167 and 186 ng, respectively, in 'polluted' and 'non polluted' area) was much more important than that from inhalation (8.4 and 5.4 ng, respectively) in both areas. The estimated daily absorbed doses of pyrene from the soil were 0.061 and 0.104 ng in 'polluted' and 'non polluted' kindergarten, respectively, which correspond to 0.032 and 0.059% of the total absorbed dose. Higher urinary concentrations of 1-OHP were found in children from 'polluted' kindergarten. In conclusion, the food seems to be a main source of the total pyrene and total PAH uptake in small children, even under a relative high PAH air exposure in the city. Pyrene concentration in soil had a negligible contribution to the total pyrene absorbed dose. Usefulness of the urinary 1-OHP as an indicator of the environmental exposure to PAH needs further research. PMID- 10867371 TI - Using the metabolism of PAHs in a human cell line to characterize environmental samples. AB - P450 reporter gene system (RGS) is an in vitro assay to detect compounds that activate the Ah receptor and induce cytochrome P450 (CYP1A1). This system utilizes a human cell (101L) stably transfected with a luciferase reporter downstream of human CYP1A1 promoter sequences. When CYP1A1-inducing compounds are present, luciferase is produced as well as endogenous CYP1A1 enzymes. Polycyclic aromatic hydrocarbons (PAHs) are more readily degraded than chlorinated compounds including dioxins, furans, and coplanar polychlorinated biphenyls (PCBs). PAH induced luciferase production begins to decrease between 6 and 16 h, while chlorinated compounds produce a more sustained response. Individual and mixtures of CYP1A1-inducing compounds were tested at both 6 and 16 h. Extracts of soils containing both PAHs and dioxins were also tested, before and after cleanup to remove PAHs. Results indicate that RGS testing at 6 and 16 h is a promising tool to differentiate between PAHs and chlorinated hydrocarbons often co-occurring in environmental samples. PMID- 10867372 TI - Effects of oxidised alpha-lipoic acid and alpha-tocopherol on xenobiotic-mediated methaemoglobin formation in diabetic and non-diabetic human erythrocytes in vitro. AB - The effects of oxidised alpha-lipoic acid and alpha-tocopherol were investigated on a human erythrocytic in vitro model of diabetic metabolic stress. Preincubation of non-diabetic and diabetic erythrocytes with oxidised alpha lipoic acid or alpha-tocopherol resulted in marked increases in nitrite-mediated methaemoglobin formation. In contrast, oxidised alpha-lipoic acid resulted in considerable reductions in 4-aminophenol-mediated methaemoglobin formation in both diabetic and non-diabetic cells. alpha-Tocopherol showed an increase only in diabetic cells, at one time point. Monoacetyl dapsone hydroxylamine (MADDS-NHOH) mediated methaemoglobin formation was reduced by oxidised alpha-lipoic acid in non-diabetic and diabetic cells at all three time points, although alpha tocopherol had no effect with MADDS-NHOH. In diabetic cells only, alpha tocopherol incubation caused a reduction in GSH levels compared with non-diabetic cells. As the agents showed pro- as well as anti-oxidant effects in this study, further studies are required to demonstrate potential diabetic benefit from alpha lipoic acid adminstration. PMID- 10867373 TI - Percutaneous penetration studies for risk assessment. AB - During the last few years the general interest in the percutaneous absorption of chemicals has increased. It is generally accepted that there is very few reliable quantitative and qualitative data on dermal exposure to chemicals in the general population and in occupationally exposed workers. In order to predict the systemic risk of dermally absorbed chemicals and to enable agencies to set safety standards, data is needed on the rates of percutaneous penetration of important chemicals. Standardization of in vitro tests and comparison of their results with the in vivo data could produce internationally accepted penetration rates and/or absorption percentages very useful for regulatory toxicology. The work of the Percutaneous Penetration Subgroup of EC Dermal Exposure Network has been focussed on the standardization and validation of in vitro experiments, necessary to obtain internationally accepted penetration rates for regulatory purposes. The members of the Subgroup analyzed the guidelines on percutaneous penetration in vitro studies presented by various organizations and suggested a standardization of in vitro models for percutaneous penetration taking into account their individual experiences, literature data and guidelines already in existence. During the meetings of Percutaneous Penetration Subgroup they presented a number of short papers of up to date information on the key issues. The objective was to focus the existing knowledge and the gaps in the knowledge in the field of percutaneous penetration. This paper is an outcome of the meetings of the Percutaneous Penetration Subgroup and reports the presentations on the key issues identified throughout the 3-year duration of the Dermal Exposure Network (1997 1999). PMID- 10867374 TI - Sodium thiosulfate protects against acrylonitrile-induced elevation of glial fibrillary acidic protein levels by replenishing glutathione. AB - Acrylonitrile (ACN) like many organic solvents produce neurotoxicity by elevating brain glial fibrillary acidic protein (GFAP), a putative biomarker of astrogliosis. In this study we tested the hypothesis that sodium thiosulfate (STS) protective action against ACN-induced astrogliosis is glutathione (GSH) mediated. Male Sprague-Dawley rats were administered for 2 weeks intraperitoneal doses (50 mg/kg body weight) of the ACN, with or without STS as outline in the Methods section. Specific brain regions were tested for GFAP, GSH and glutathione S-transferase (GST) enzyme activity. In the brain regions tested STS significantly (P0.1). This study suggests a new way of comparing reproducibility in terms of kappa values between diagnostic methods in clinical studies where accuracy cannot be evaluated. The subtraction method may be useful in the assessment of caries lesion behaviour in the clinic. PMID- 10867420 TI - Digital subtraction radiography for monitoring dental demineralization. An in vitro study. AB - The aim of the present study was to evaluate the assessment of progression of demineralization by digital subtraction radiography. In each of 14 extracted human teeth, 2 approximal enamel demineralization lesions were induced in vitro to simulate dental caries. A modified tunnel preparation with glass ionomer fillings was performed on one lesion of every tooth as a model of caries inhibition. Every week, radiographs were obtained under standardized conditions over a period of 42 days. The radiographs were digitized and calibrated for grayscales. Reference landmarks were chosen and aligned for the different pictures by computer-assisted imaging to adjust the images for projective distortions. The images of the 7th, 14th, 21st, 28th, 35th and 42nd day were subtracted from the baseline radiograph. The mean value of gray values of the subtraction images was calculated and ANOVA tests for repeated measurements and paired t tests were used for statistical analysis. The results of the present study indicate that statistically significant gray level changes due to progression of demineralization could be detected in the radiographic images by subtraction analysis. Differences between glass ionomer-filled and nonfilled lesions failed to reach significance. The introduced method may have the potential to detect and document minute caries progression. PMID- 10867421 TI - Detection of natural white spot caries lesions by an ultrasonic system. AB - Ultrasound has been used in industrial business as one of the nondestructive measurement methods. It was hypothesized that nondestructiveness of the ultrasonics could be useful in determination of demineralization of noncavitated carious lesions on human enamel. This investigation was designed to determine the presence of natural carious lesions on proximal surfaces of human molar teeth using an ultrasonic system compared to radiography and histology as the gold standards. Measurements were made directly from proximal surfaces of 20 mandible molar teeth with white spot carious lesion by 2 examiners independently with the ultrasonic system. Ultrasonic evaluation of each natural white spot lesion had a sensitivity of 88%, specificity of 86%, positive predictive value of 88% and a negative predictive value of 86%, and the chance-corrected agreement was also satisfactory (kappa=0.74) compared to histology. The radiograph demonstrated chance-corrected agreement of 0.41:0.38 for the first and second examiners, respectively. Duncan test analysis of the numerical values was significantly different for the intact and the noncavitated carious surfaces (p<0.05). The results indicated that the ultrasonic evaluation is a sensitive method for the detection of the natural white spot carious lesions and can differentiate the changes in elastic properties of enamel numerically. PMID- 10867422 TI - A Raman spectroscopic investigation of dentin and enamel structures modified by lactic acid. AB - Confocal Raman microspectrometry allows a thorough molecular analysis of mineralised dental tissues. The output information is provided in the form of curves representing the intensity of the signal according to the frequency, and its mathematical exploitation permits all sorts of comparative and quantitative analyses. By this process, we investigated the in vitro action of lactic acid on enamel and dentin from human permanent teeth. Modifications due to the acidic attack essentially concern phosphate grouping PO(4)(3-), which represents the mineral phase in enamel and dentin (hydroxyapatite): on Raman spectra, changes in intensity of the PO(4)(3-) band are linked to the type of dentin, to its anatomical location, and to the age of the subject. The variability of the dentinal chemical structure was confirmed by a quantitative statistical analysis, revealing a significant spectral difference between coronal and root dentin. PMID- 10867423 TI - Comparative reduction of enamel demineralization by calcium and phosphate in vitro. AB - In theory, calcium and phosphate in the plaque fluid exert a large influence on the demineralization of enamel surface. In order to know the effect of increasing the concentration of either of these factors, the following in vitro experiment was conducted. Three thin sections, about 150 Im thick, were cut out from each of 13 human premolars. All surfaces of the sections, except for the original enamel surface, were coated with nail varnish. These sections were immersed into one of two sets of demineralizing solutions for 1 week at 25 degrees C. Each set, the 'calcium set' and the 'phosphate set', contained three solutions. The composition of these solutions differed mainly in calcium or phosphate concentrations. After 1 week, the degree of demineralization was determined by image analysis of contact microradiograms from each section. The subsurface demineralization in enamel was reduced by 95% by increasing the calcium concentration of the demineralizing solution from 7 to 21 mmol/l. A similar reduction (87%) was observed by increasing the phosphate concentration. However, the amount of phosphate needed was approximately 20 times more than that of calcium. The larger inhibitory effect that calcium has on enamel demineralization was related to the larger effect it has on the degree of saturation of the solution. Even though no statistically significant difference was found between the effect of calcium and phosphate on the demineralization of enamel (when the solutions had the same degree of saturation), the difference in the standard deviation of demineralization suggests the existence of some other factors which have an influence on the demineralization reaction. PMID- 10867424 TI - In vitro fluoride dose-response study of sterilized enamel lesions. AB - Currently our intra-oral model uses enamel specimens that have been disinfected by soaking in buffered formalin (pH 6.8). However, because of increasing emphasis on infection control, it is important to identify a way to sterilize these specimens. The aim of this study was to determine if autoclaved, or gas sterilized, lesioned enamel responds to fluoride (F) in the same way alcohol disinfected enamel lesions do. Seventy-two formalin-disinfected, human enamel specimens (3 mm) were lesioned in demineralizing solution for 96 h and were then divided into three groups. One group was autoclaved; one group was gas sterilized (ethylene oxide), and the remaining 24 specimens were further disinfected in 70% ethanol for 10 min. Specimens in each group were then treated 4 times/day for 4 weeks with 0, 250 or 1,100 ppm F dentifrice slurries in an in vitro cycling, remin/demin model. Following treatment, fluoride uptake was analyzed by microdrill biopsy, and lesion depth and mineral content changes (DeltaM) were determined by transverse microradiography. Data were analyzed by one-way ANOVA analysis. In all three groups of specimens there were significant (p<0.05) differences in fluoride uptake in response to different fluoride treatments. Autoclaved lesions failed to provide dose- response data with regard to changes in lesion mineral content. Because formalin and 70% alcohol are only disinfectants, and autoclaving altered the responsiveness of enamel lesions, results from this study suggest that, of the methods tested, gas sterilization is the preferred method for sterilizing enamel specimens that will be used in intra oral studies. PMID- 10867425 TI - Fluoride profiles in dental plaque in vivo formed on fluoride pre-treated human enamel. AB - Using a novel device capable of generating plaque in vivo on a natural enamel substrate, it has been possible to determine fluoride profiles from the saliva plaque interface towards the enamel surface. Fluoride profiles in dental plaques tended to fall from the saliva-plaque interface towards the enamel. The device also offered the possibility of examining fluoride distributions after pre treatment of the enamel with fluoride in vitro. Fluoride profiles were determined in plaque generated in vivo on enamel surfaces, which had been previously treated with a 900-ppm fluoride solution. The results showed the previously reported fall from the plaque surface, but in addition, a further rise towards the enamel surface was seen. The data imply that enamel loaded with fluoride can release some of this fluoride back into the plaque and may act as a fluoride reservoir. PMID- 10867426 TI - The anticariogenic effect of amine fluorides on Streptococcus sobrinus and glucosyltransferase in biofilms. AB - Dental caries is a chronic infectious disease caused by the accumulation of bacterial plaque (biofilm) on tooth surfaces. Antibacterial agents, in addition to other preventive measures, can control dental plaque accumulation. Amine fluorides (AmF) are known anticaries agents for over 30 years. The purpose of our study was to assess the adsorption and desorption of AmF to experimental dental biofilm and to evaluate the effect of AmF on Streptococcus sobrinus 6715 and glucosyltransferase (GTF) activity in experimental dental biofilms. The experimental plaque model used in this study consists of hydroxyapatite beads coated with human saliva (sHA), followed by adsorption of S. sobrinus and synthesis of in situ polysaccharides. Our results show that the viability of S. sobrinus in biofilm decreased as the concentration of AmF and chlorhexidine (CHX) increased. The concentration of AmF and CHX required to kill S. sobrinus adherent to sHA is about 100 times greater than the concentration required to kill the same amount of planktonic bacteria. Adsorption of AmF to surfaces was more than 90% and the desorption of AmF from our experimental model was limited. Pre adsorption of AmF on the surface increased adhesion of S. sobrinus but also resulted in surface killing of the adsorbed bacteria. At low concentrations AmF increased GTF activity in solution by about 10%, but at concentrations above 0.1 mM it inhibited GTF activity. Inhibition of GTF on the surface required about 100 times more AmF than in solution. Our results show that AmF retains its anticariogenic effects in solution and in biofilm systems. PMID- 10867427 TI - Effects of dental treatment and single application of a 40% chlorhexidine varnish on mutans Streptococci in young children under intravenous anaesthesia. AB - Clinical studies suggest that application of a highly concentrated chlorhexidine varnish results in a decrease in the number of mutans streptococci and thereby a decrease in the caries risks. The aim of this study was to determine the effect of dental treatment on the levels of mutans streptococci (MS) and lactobacilli (LB) and the additional effect of a single application of 40% chlorhexidine varnish (EC40) on the level of MS. Twenty-three children under the age of 5 years scheduled for full dental treatment were selected. Of these 23 children (mean dmf s 27.1, SD 19.3), 11 children had nursing bottle caries. The mean sugar exposure was 6.4. Subjects were randomly distributed into two groups of approximately equal size. One group received an EC40 application after dental treatment, while the other group received only full dental treatment, both with intravenous anaesthesia with propofol as a single drug. An unstimulated saliva sample and a plaque sample were taken prior to dental treatment. The saliva and plaque sampling of the subjects was repeated after 6 weeks. MS and LB were isolated and counted. The number of children harbouring more than 10(6) MS in a pooled plaque sample decreased significantly from 8 to 2 children 6 weeks after dental treatment. No additional effect of EC40 was found. The number of salivary MS did not change significantly between the groups or before and after treatment. The figures for LB remained at a high level of more than 10(4)/ml saliva before (21 children) and 6 weeks after treatment (17 children). The results of this study indicate that dental treatment results in a significant suppression of plaque MS, while a single application of EC40 showed no significant additional suppression after 6 weeks. PMID- 10867428 TI - Evaluation of Carisolv for the chemo-mechanical removal of primary root caries in vivo. AB - The objective of this study was to evaluate a new chemo-mechanical method (Carisolv) for the removal of primary root caries in vivo in terms of efficiency, treatment time and patient perception. Thirty-eight patients participated in an open, randomised and controlled study. Of the 60 root carious lesions included, 34 were randomised for chemo-mechanical treatment and 26 for drilling. A within subject comparison was used whenever the patient had two cavities, which was the case for 22 subjects. All the Carisolv-treated cavities became caries free, as did all but one of those treated with drilling, as judged by an independent examiner. Only 4 of 34 patients asked for anaesthesia in the Carisolv group compared with 6 of 26 patients in the drilling group. Of those who did not use anaesthesia, 12 individuals in the drilling group experienced some pain compared with none in the chemo-mechanical group (p<0.001). The mean treatment time for the Carisolv method was 5.9+/-2.2 min, compared with 4.5+/-2.0 min for drilling; time for anaesthesia excluded (p<0.05). No negative reactions or adverse effects were recorded during the study. All 55 teeth examined in the 1-year follow-up were found to be sensitive using an electric pulp tester, and there was no difference regarding the condition of fillings between the lesions treated with Carisolv and drilling. It can be concluded that root caries can be effectively removed using the Carisolv method. The longer treatment time was compensated by less need for anaesthesia. PMID- 10867430 TI - Ultrasonication as a method to study enamel demineralisation during acid erosion. AB - The aim of this study was to use ultrasonication as a method to measure subsurface demineralisation of enamel. Polished human enamel samples with surface profiles within +/-0.3 microm were divided into 6 groups of 10 specimens. The groups of specimens were exposed to 0. 3% citric acid (pH 3.2) for 30 min, 1, 2, 3 or 4 h. The depths of the resulting lesions were measured using a profilometer. A control group was stored in water for 4 h. Ultrasonication in water was performed on the specimen groups for 5, 30, 120, 240 and 480 s with profilometric measurements at each time point. The depth of the erosion increased linearly with the exposure time. Most of the additional loss of enamel occurred with the 5 second ultrasonication. The 30-min and 1-hour erosion lesions were further deepened by approximately 1 microm with 5 s of ultrasonication. The 2-, 3- and 4 hour lesions were deepened by 2-4 microm with 5 s of ultrasonication. There were no changes in the control group. It is concluded that ultrasonication removed softened enamel from the surface of the eroded enamel. Ultrasonication together with accurate measurement of lesion depth by profilometry offers a useful method for studying the depth of enamel softening associated with erosion. PMID- 10867429 TI - Effect of tooth-bound fluoride on enamel demineralization/ remineralization in vitro. AB - The effect of tooth-bound fluoride (F) on enamel caries formation was investigated under the condition that loosely bound F was essentially absent. Eighteen thin enamel sections, prepared from the lingual or buccal surfaces of extracted human molars, were embedded in acrylic resin with the enamel surfaces exposed. The sections were placed in a pH 7 remineralizing solution (RS; 1.2 mmol/l Ca, 0.72 mmol/l P, 30 mmol/l KCl, 50 mmol/l HEPES) for 5 days, and were randomly divided into 3 groups: (1) control group that received no treatment, (2) acidulated phosphate fluoride (APF) group that received 5 cycles of a 4 min treatment with APF gel followed by immersion in the RS for 2 days (RS changed daily) and (3) dicalcium phosphate dihydrate (DCPD) - APF group that received 5 cycles of a 4-min pH 2.1 DCPD-forming solution followed by 4 min APF gel and then placed in the RS for 2 days. After the treatment cycles, the sections were washed in a constant composition F titration system to remove loosely bound F. An in vitro model, which consisted of cycles of de- (6 h) and remineralization (18 h) each day for 5 days, was used to produce caries-like lesions in the specimens. The DeltaZ (mineral loss) values, measured by quantitative microradiography, of the lesions formed in the three groups were (mean +/- standard deviation; n = 6) 91.2+/-12.3 microm for the control group, 41.3+/-10.1 microm for the APF group and 21.2+/-4.8 microm for the DCPD-APF group. The same system produced lesions in untreated shark enamel with a mean DeltaZ of 4.4+/-0.3 microm (n = 12). One-way fixed-effects ANOVA indicated that mineral loss was significantly different among the different groups (p<0.05). The results showed that enamel resistance to lesion formation increased with increasing tooth-bound F content. Shark enamel was much more resistant to demineralization than human enamel. PMID- 10867431 TI - Studies concerning the glucosyltransferase of Streptococcus sanguis. AB - We have shown in previous studies that the glucosyltransferase (Gtf) enzymes of Streptococcus mutans have distinct properties when adsorbed to a surface. In the present study, we compared the activity of Gtf from Streptococcus sanguis, designated GtfSs, in solution and on the surface of saliva-coated hydroxyapatite (sHA) beads, and determined the ability of its product glucan to support the adherence of oral microorganisms. Gtf from S. sanguis 804 NCTC 10904 was purified from culture supernatant fluids by means of hydroxyapatite chromatography. Enzyme and the substrate were prepared in buffers at pH values from 3.5 to 7.5. Maximum activity of GtfSs occurred between pH 5.5 and pH 6.5, whether in solution or adsorbed onto a surface. The solubilized and insolubilized enzymes showed highest activity at 40 degrees C; activity was reduced by 50(+/-2)% at 20 and 30 degrees C. The enzyme did not form glucans in either phase at 10 or 60 degrees C. The K(m), determined from Lineweaver-Burk plots, for the enzyme in solution was 4.3(+/-0.4) mmol/l sucrose, and the K(m) for the enzyme on sHA beads was 5.0(+/ 1.0) mmol/l sucrose. The ability of the GtfSs glucan synthesized on the surface of sHA beads to support the adherence of oral bacteria was investigated. (3)H thymidine-labeled bacteria (S. mutans GS-5, S. sobrinus 6715, S. sobrinus 6716, S. sanguis 10904, Actinomyces viscosus OMZ105E, A. viscosus 2085, and A. viscosus 2086) were incubated with sHA beads coated with GtfSs glucan. S. mutans GS-5 displayed the highest level of binding numerically. These results show that the GtfSs of S. sanguis is active on sHA beads, that the pH optimum for activity on a surface differs slightly from that in solution, and that its product glucan can support the adherence of oral microorganisms. PMID- 10867432 TI - Changes in dental caries prevalence in 12-year-old students in the State of Mexico after 9 years of salt fluoridation. AB - The purpose of this study was to describe the dental caries experience in 12-year old students in the State of Mexico and to detect changes after 9 years of salt fluoridation (1988-1997). The 1987 WHO dental caries criteria were used in both surveys. The population studied encompassed 2,275 12-year-old students in 1988, and 1,138 in 1997. The proportion of 'caries-free' children was 10. 3% in the first survey and 27.7% in the second one. The mean DMFT index was 4.39 (SD 2.9) in 1988 and 2.47 (SD 2.4) in 1997, the confidence interval of the differences between DMFT means was [95% CI 1.73, 2.11]; the caries reduction detected was 43.7%. The Unmet Restorative Index was 82.3% in 1988 and 72.8% in 1997. The results of the present study indicate that the oral health status of State of Mexico students has improved during the last decade; however, there is still a need for further caries reduction and an increment in access to dental treatment. PMID- 10867433 TI - Muscle spindle reinnervation following phenol block. AB - Infusion of phenol into peripheral nerves is used clinically to manage spasticity. It produces relief of symptoms by chemical denervation. We simulated the clinical procedure by bathing the lateral plantar nerve of rats in 7% phenol solution for 20 min. We studied the innervation of muscle spindles in the plantar lumbrical muscles of untreated rats and in rats 4 and 6 weeks after a single phenol block. Spindles were identified by the immunoreactivity of nuclear bag(1) fibers to slow tonic myosin (antibody ALD 19). The integrity of the sensory and motor reinnervation of spindles was evaluated using a monoclonal antibody specific for a high molecular weight neurofilament protein. Four weeks after phenol block, muscle spindles were difficult to find, as their immunoreactivity to antibody ALD 19 was reduced. In those spindles studied, most (>80%) were completely denervated. The remainder of which were innervated by afferents only. None received efferent (gamma) innervation. After 6 weeks, spindles were readily identified and nearly all (>90%) received recognizable afferent innervation. A much smaller number (38%) received gamma innervation. Phenol block thus results in a complete denervation of muscle spindles, followed by a fairly rapid sensory reinnervation. Reinnervation by gamma motor neurons is either incomplete or significantly delayed. PMID- 10867434 TI - Morphological characteristics of neuromuscular junctions of the opossum (Didelphis albiventris) extraocular muscles: a scanning-electron-microscopic study. AB - Three types of neuromuscular junctions were described in the extraocular muscles of the opossum. The present study demonstrates the three-dimensional characteristics of these neuromuscular junctions after HCl connective tissue digestion. Adult opossum of both sexes were used and the neuromuscular junctions of the extraocular muscles were examined after removal of the intramuscular connective tissue and basal layer. This material was examined with a scanning electron microscope. Two types of 'en plaque' neuromuscular junction were described: the continuous type revealed elongated and branched primary synaptic grooves separated from each other by sarcolemma protuberances with different sizes, and the discontinuous or punctiform type which presents very shallow and discontinuous grooves when compared with the former. The multiple neuromuscular junctions were observed as two or three junctions associated with the same muscular fiber. The multiple junctions were present in thin fibers (around 11 microm caliber); the en plaque junctions were associated with large diameter fibers (around 21 microm). This study confirms and reveals the detailed morphological characteristics of the three neuromuscular junction types previously described by transmission electron microscope in the extraocular muscles of opossum. PMID- 10867435 TI - Ciliogenesis in the uterine tube of control and superovulated heifers. AB - Ciliogenesis was investigated in the uterine tube epithelium of control and superovulated heifers during day 1 to day 7 of the oestrous cycle. Seventy-two heifers were used, consisting of four control groups and four superovulated groups. All heifers received cloprostenol (PG) to induce oestrus (day 0). Superovulated heifers received 24 mg pFSH at doses of 4.5, 3.5, 2.5 and 1.5 mg given twice daily. Control and superovulated heifers were slaughtered on days 1, 3, 5 and 7 of the oestrous cycle. Samples from the ampulla, preisthmus and isthmus of the uterine tube were collected and processed for light and electron microscopy, following standard procedures. Both centriolar and acentriolar pathways were involved in the development of basal bodies. Formation of basal bodies and protrusion of ciliary shafts mostly occurred during days 1 and 3 of the oestrous cycle. The centriolar pathways, in which procentrioles generate with the aid of preexisting centrioles, played a minor role in the heifer uterine tube. In the acentriolar pathways, fibrous granules were the first structures which appeared in the course of ciliogenesis and they initially occurred in association with free ribosomes. Subsequently, deuterosomes arose in the aggregates of fibrous granules, and then procentrioles containing microtubules originated around deuterosomes or apart from deuterosomes. Newly formed centrioles migrated to the apical cytoplasm, and ciliary shafts extended first at the periphery of the luminal surface of ciliogenic cells. Deuterosomes as well as rootlet formation were considered to be related to the fibrous granules. Quantitative examinations by light microscopy showed that the number of ciliated cells in the ampullar, preisthmic and isthmic epithelium of the superovulated heifers was significantly higher than in the control heifers during day 1 and day 3 of the oestrous cycle. PMID- 10867436 TI - Vascularization of developing human olfactory neuroepithelium - a morphometric study. AB - The present study reveals intraepithelial capillaries in the olfactory neuroepithelium of human fetuses aged between 12 and 24 weeks of gestation, which disappear at birth. The area occupied by the intraepithelial capillaries increases significantly with fetal age (0.047 +/- 0.014 microm(2)/microm(2) at 12 weeks and 0.101 +/- 0. 025 microm(2)/microm(2) at 24 weeks) and with the thickness of the epithelium (45.00 +/- 6.74 microm at 8 weeks and 64.10 +/- 8.51 microm at 24 weeks). The vascularization of the developing neuroepithelium may suggest increased metabolic demand during development and maturation of the olfactory neuroepithelium, and postnatal retreat of capillaries to the underlying lamina propria may suggest diffusion of nutrients and gases from blood vessels into the lamina propria and direct gaseous exchange from the atmosphere. PMID- 10867437 TI - The sinus hair follicle of the cat. Hair growth and hair cycle. AB - The cat's sinus hair follicle is described histologically with special regard to structural characteristics and functional mechanisms of sinus hair growth and sinus hair cycle. Special features of both cornification of the inner epithelial root sheath and hair fixation, respectively, result in a loss of traction that is required for hair growth and hair expulsion. Instead, there is the outgrowing new anagen sinus hair that pushes the preceeding sinus hair upwards, and - secondly - there is a long-lasting cell pro- liferation. Cell proliferation - immunocytochemically detected with anti-proliferating cell nuclear antigen - continues in the preceeding sinus hair while the formation of a new, succeeding sinus hair follicle begins. No distinct, 'typical' catagen state of sinus hair follicle has been found even in a large number of collected specimens. These findings stress that the well known features of hair growth and hair cycle in the pelage hair follicle cannot be generalized and adopted in all details to the sinus hair follicle. PMID- 10867438 TI - Developmental morphological and histological studies on structures of the human fetal elbow joint. AB - In the present work, morphological changes in the developing human elbow joint were studied at different prenatal ages (8, 12, 16, 20, 29 and 40 weeks) and were compared with the same structures in the adult joint. The elbow joint had gone through its most important developmental changes during the 20th week of prenatal life, probably due to the direct dynamic effect of the newly developed fetal movement. During later prenatal development, the articular surfaces of the lower end of humerus and the upper ends of radius and ulna developed their characteristic congruencies, so that the highly curved convexities always articulate with the highly curved concavities. That process progressed postnatally and even till adult age. In full-term infants it was found that the lower end of humerus had acquired its adult shape, while the shape of the upper ends of radius and ulna were still not fully developed. They continued development in postnatal life even till adult age. In the present work, histological prenatal studies were done on longitudinal sections from the back of the capsule and synovial tissue, early (8 weeks) and late in full term, and the results were also compared with the same structures in adults. It was found that at all ages, the capsules were formed of cellular and fibrous elements, but at early prenatal age (8 weeks), this cellular condensation was more massive and prominent while in full-term infants, it became generally more fibrous, but was still different compared to adults. Basic cellular structures of the synovial tissue changed very little during the late prenatal developmental stage, as it did not become more fibrous than cellular during these periods, but differences in vascularity became more obvious. The cartilaginous content of the articular surface at 8 weeks was highly cellular with very little intercellular matrix. In contrast to that of full term, this cartilage became fully chondrogenous with a notable decrease in cellular density and massive increase in matrix content. PMID- 10867439 TI - The lumbosacral ligament. An autopsy study of young black and white people. AB - The lumbosacral ligament (LSL), situated between the L5-vertebra and the sacrum, was studied in autopsy material. Twenty-eight cadaveric specimens from 12 black and 16 white persons aged 17-30 years were studied during routine forensic autopsies. The ligaments were measured and determined in situ. Thereafter, the ligaments were removed for histologic preparation. The ligament in the black subjects was thicker compared with the white (7.5 +/- 1.4 vs. 5.7 +/- 1.2 mm), and wider (11.7 +/- 1.6 mm in the black vs. 9.2 +/- 0.5 mm in the white), yielding a greater cross-sectional area in the black group (70.7 +/- 22.8 vs. 34.5 +/- 11.4 mm(2), p < 0.001). However, no histological differences were noted. The previously described fibro-osseus tunnel could not be detected in any of the subjects. In all instances, the ligament was situated medial to the L5 nerve. Compression of the L5 nerve under the previously presented fibro-osseus tunnel could not be confirmed. PMID- 10867440 TI - Anatomy of the intrahepatic ramification of the portal vein in the right hemiliver. AB - The ramification of the portal vein in the right hemiliver was studied by anatomic dissection in 36 formalin-fixed human livers. In 28/36 (77.8%) cases, the portal vein bifurcated into a right branch and a left branch and the right branch bifurcated into anterior and posterior segmental branches. The anterior segmental branch terminated in the anterosuperior subsegment (S(8)) in two types: bifurcated when it divided into anterior P(8) and posterior P(8 )branches towards the respective regions of S(8) (24/28 cases) and monopodal when it had a single pedicle (4/28 cases). The maximum anteroinferior subsegmental branch (P(5 )maximum) originated either from the anterior segmental branch (16/28 cases) or from the anterior P(8) branch (12/28 cases). The posterior segmental branch vascularized the posteroinferior (S(6)) and the posterosuperior (S(7)) subsegments, and was terminated in three types: fan-shaped (16/28), bifurcated (9/28) and tripodal (3/28). In 4/36 (11.1%) cases the portal vein bifurcated into a right branch and a left branch but the posterior segmental branch was not present. In 4/36 (11.1%) the right branch of the portal vein was not present. These anatomical variations are explained separately and finally all cases are considered as a whole. PMID- 10867441 TI - The human anterior tympanic artery. A nutrient artery of the middle ear with highly variable origin. AB - The variability of the origin of the anterior tympanic artery was investigated in 104 individuals of both sexes. A surprising laterality was found: thus, while the left anterior tympanic artery originated as a singular vessel from either the maxillary or the superficial temporal artery with almost equal frequencies (44.7 and 45.9%, respectively), the right anterior tympanic artery predominantly branched off from the maxillary artery (77.8% of cases). Besides the origin from either the maxillary artery or the superficial temporal artery, also anterior tympanic arteries branching off from the external carotid artery were found to occur (4% on the left and 1% on the right side). Although in the majority of individuals, a singular anterior tympanic artery occurred within the infratemporal fossa, duplications of the anterior tympanic artery were found to be present: in one case on the right and in 8 cases on their left side. In 1 female individual, a triplet of left anterior tympanic arteries was found to supply the tympanic cavity. Also in these cases, the anterior tympanic artery arose from either the external carotid, the superficial temporal or the maxillary artery. In singular cases, even several other branches of the maxillary artery, viz. the deep auricular, middle, and accessory meningeal, as well as the posterior deep temporal, inferior alveolar and masseteric arteries were found to form common trunks with the anterior tympanic artery. PMID- 10867443 TI - Beta-amyloid deposition in the temporal lobe of patients with dementia with Lewy bodies: comparison with non-demented cases and Alzheimer's disease. AB - beta-Amyloid (Abeta) deposition in regions of the temporal lobe in patients with dementia with Lewy bodies (DLB) was compared with elderly, non-demented (ND) cases and with Alzheimer's disease (AD). The distribution, density and clustering patterns of diffuse, primitive and classic Abeta deposits were similar in 'pure' DLB and ND cases. The distribution of Abeta deposits and the densities of the diffuse and primitive deposits were similar in 'mixed' DLB/AD cases compared with AD. However, the density of the classic deposits was significantly lower in DLB/AD compared with AD. In addition, the primitive Abeta deposits occurred more often in small, regularly spaced clusters in the tissue and less often in a single large cluster in DLB/AD compared with 'pure' AD. These results suggest that pure DLB and AD are distinct disorders which can coexist in some patients. However, the Abeta pathology of DLB/AD cases is not identical to that observed in patients with AD alone. PMID- 10867442 TI - Cholesterol modulates the membrane-disordering effects of beta-amyloid peptides in the hippocampus: specific changes in Alzheimer's disease. AB - Cholesterol represents an important determinant of the physical state of biological membranes. In Alzheimer's disease (AD) brains, specific changes in the distribution of cholesterol and its membrane-ordering effects take place. In the present study, membrane fluidity was investigated at the level of the hydrocarbon core and of the heads of the phospholipid bilayers using two different fluorescent probes. Hippocampal membranes of AD brains showed a reduced fluidity in the hydrocarbon core region only. Fluidity was correlated with the cholesterol content in AD and control membranes. Aggregated beta-amyloid peptides (Abeta) disrupted brain membrane structure in AD patients and controls in the same fashion. However, this effect was correlated with the cholesterol content in AD membranes only. It is suggested that in AD the brain becomes specifically sensitive for the modulation by membrane-bound cholesterol of the membrane disturbing and ultimately neurotoxic properties of Abeta. PMID- 10867444 TI - The validity of the MMSE and SMQ as screening tests for dementia in the elderly general population-- a study of one rural community in Japan. AB - OBJECTIVE: To compare the validity of the Mini Mental State Examination (MMSE) and the Short-Memory Questionnaire (SMQ) as screening tests to detect dementia in the elderly general population. SUBJECTS: Six hundred and sixty-two subjects and their informants from the elderly general population sample who had completed these tests. SETTING: One rural community survey in Japan. METHOD: We used receiver-operating characteristic analysis to compare the performance of the MMSE and the SMQ with the clinical diagnosis of dementia according to DSM-III-R. RESULTS: The areas under the receiver-operating characteristic curve of the MMSE and the SMQ were 0.980 (SE = 0.006) and 0.982 (SE = 0.008), respectively. This differed from chance to a highly significant degree for both the MMSE and the SMQ, but the difference between the two scales was not statistically significant. CONCLUSION: As screening tests to detect dementia in the elderly general population, the SMQ which is assessed by informants demonstrates a statistically significant discriminating ability as well as the MMSE. PMID- 10867445 TI - No evidence of endothelial dysfunction in patients with Alzheimer's disease. AB - In view of accumulating evidence of vascular pathology in Alzheimer's disease (AD), we tested the hypothesis that AD patients have impaired endothelial function. This was assessed using the technique of strain-gauge venous occlusion plethysmography, which measures forearm blood flow (FBF). Intra-arterial (brachial) infusion of acetylcholine (ACh) and sodium nitroprusside (SNP) was used to assess local endothelial dependent and independent responses, respectively. There was no difference in the basal FBF of patients and controls. ACh and SNP caused dose-related increases in FBF from baseline, but no difference was recorded between the AD and control group. This study provides no evidence of endothelial dysfunction in the systemic circulation of patients with AD. PMID- 10867446 TI - Metrifonate therapy in Alzheimer's disease: a pooled analysis of four randomized, double-blind, placebo-controlled trials. AB - This retrospective analysis assessed the efficacy of metrifonate in the treatment of mild to moderate Alzheimer's disease (AD). Those four studies meeting Food and Drug Administration guidelines for establishing the efficacy of an AD therapeutic agent were pooled for further analysis. Data were included from all patients valid for the intent-to-treat analyses (last observation carried forward). Patients received once daily placebo (n = 550), metrifonate 30-60 mg (by weight, n = 769) or 60/80 mg (by weight, n = 197). Metrifonate 60/80 mg significantly improved the cognitive abilities [AD Assessment Scale - Cognitive Subscale (ADAS Cog), p = 0.0001; Mini Mental State Examination (MMSE), p = 0.0001], psychiatric and behavioral disturbances (Neuropsychiatric Inventory, p = 0.039; ADAS - Noncognitive Subscale, p = 0.0001), performance of instrumental and basic activities of daily living (Disability Assessment for Dementia, p = 0.0002) and global status (Clinician's Interview-Based Impression of Change with Caregiver Input, p = 0.0001) of AD patients when compared with placebo. Metrifonate effects across these domains were dose related. Metrifonate 60/80 mg significantly improved the cognitive performance relative to both placebo and to baseline as evaluated by both the ADAS-Cog and the MMSE. Metrifonate is the first cholinesterase inhibitor consistently shown under prospective, placebo-controlled conditions to improve significantly behavior in addition to cognition, function in activities of daily living and global functional status of patients with mild to moderate AD. PMID- 10867447 TI - Predictors of cognitive decline in the early stage of probable Alzheimer's disease. AB - Several papers have attempted to find neurological and neuropsychological predictors of progression in Alzheimer's disease (AD) till now. Despite this quite large amount of works, different and not univocal conclusions have been reported in this field. Different study samples, different end-points and differences in statistical methods can explain much of the inconsistency in the results obtained. In our study, AD patients were examined in a very early stage of the disease to avoid any possible risk to examine subjects at different times of evolution. All the patients underwent an extensive neuropsychological test battery twice (baseline and follow-up) spaced out over about 1 year and were divided into two groups of fast decliners (FD) and slow decliners (SD) on the basis of their rate of decay at the MMSE score. Verbal memory tests, mental control abilities and attention-demanding tasks seem to play a pivotal role in distinguishing the two groups of subjects in the early stage of the disease. Moreover, FD patients show a worse performance than SD at the baseline in most of the cognitive domains explored. In conclusion, different subtypes of AD do exist and an important predictor of progression is represented by the severity of the cognitive impairment at the onset. PMID- 10867448 TI - One year follow-up of parkinsonism in dementia with Lewy bodies. AB - The progression of parkinsonism over 1 year was evaluated in a prospective cohort of patients (n = 338), suffering from dementia with Lewy bodies (DLB), Alzheimer's disease (AD) or vascular dementia (VaD). Parkinsonism was assessed using the modified Unified Parkinson's Disease Rating Scale. Significant parkinsonism was significantly commoner in DLB sufferers (71%) than amongst patients with AD (7%) or VaD (10%). DLB patients with established parkinsonism had an annual increase in severity of 9%, but progression was more rapid (49% in 1 year) in patients with early parkinsonism. Parkinsonism was frequent at all severities in DLB patients, but usually only present in other dementias when MMSE <10. PMID- 10867449 TI - Detection of dementia in primary care: the Linkoping study. AB - We examined to what extent dementia and cognitive impairment are detected in a primary health care centre. A systematic sample of patients aged 70 years and above, who attended a primary health care centre for a doctor's consultation (n = 350) were examined with a neuropsychiatric examination and an interview with a close informant. Dementia was diagnosed according to DSM-III-R. Medical records from the health centre were examined for entries on cognitive decline or dementia, other diagnoses and prescribed drugs. The prevalence of dementia was 16.3% and a further 3.1% had questionable dementia. Cognitive disturbances or dementia were noted in case records in 15 out of 57 (26%) demented cases, and in 1 out of 11 (9%) questionable dementias. Compared to non-demented patients, the demented had more diagnoses and a higher number of prescribed drugs. Severity and duration of dementia were associated with an increased detection. PMID- 10867450 TI - A 26-week analysis of a double-blind, placebo-controlled trial of the ginkgo biloba extract EGb 761 in dementia. AB - This intent-to-treat (ITT) analysis was performed to provide a realistic image of the efficacy that could be expected after 26 weeks treatment with a 120-mg dose (40 mg t.i.d.) of EGb 761 (EGb). The data were collected during a 52-week, double blind, placebo-controlled, fixed dose, parallel-group, multicenter study. Patients were mildly to severely impaired and diagnosed with uncomplicated Alzheimer's disease or multi-infarct dementia according to ICD-10 and DSM-III-R criteria. The primary outcome measures included the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-Cog), Geriatric Evaluation by Relative's Rating Instrument (GERRI) and Clinical Global Impression of Change. From 309 patients included in the ITT analysis, 244 patients (76% for placebo and 73% for EGb) actually reached the 26th week visit. In comparison to the baseline values, the placebo group showed a statistically significant worsening in all domains of assessment, while the group receiving EGb was considered slightly improved on the cognitive assessment and the daily living and social behavior. Mean treatment differences favored EGb with 1.3 and 0.12 points, respectively, on the ADAS-Cog (p = 0.04) and the GERRI (p = 0.007). In the group receiving EGb, 26% of the patients achieved at least a 4-point improvement on the ADAS-Cog, compared to 17% with placebo (p = 0.04). On the GERRI, 30% of the EGb group improved and 17% worsened, while the placebo group showed an opposite trend with 37% of patients worsening for 25% improved (p = 0.006). Regarding safety, no differences between EGb and placebo were observed. PMID- 10867451 TI - What's new with tumor markers for colorectal cancer? AB - BACKGROUND/AIMS: Many tumor markers have been utilized in the follow-up care of colorectal cancer patients. No marker, however, has proven reliably accurate in detecting recurrent disease. METHODS: The strengths and weaknesses of currently available tumor markers are reviewed, with attention to related cost and efficacy. RESULTS: Tumor antigens, enzymes, and genetic markers have been used as tumor markers. CEA and CA 19.9 are the most widely utilized; however, genetic markers are the most promising for the future. CONCLUSIONS: Currently available markers have significant limitations. Development of genetic markers may greatly enhance our ability to predict prognosis and the need for adjuvant therapy. Marker-guided therapy may play an increasing role in this disease. PMID- 10867452 TI - 'Free omental plug': a nostalgic look at an old and dependable technique for giant peptic perforations. AB - Peptic perforation is a serious complication of peptic ulcer disease. The defect in the intestinal wall usually does not present a difficult technical problem of surgical management, in most cases perforation can be closed primarily. On rare occasions an extremely large defect (giant peptic perforation - defined as any perforation greater than 2.5 cm in size) cannot be closed by these simple techniques. Modalities of treatment advocated for such an ulcer over the years are: free omental plug in the form of a mushroom; serosal patch technique; jejunal pedicle graft, partial gastrectomy, and finally the possible addition of proximal gastrojejunostomy. The omental plug is a simple procedure which does not require expertise and can even be performed in a very short time by a trainee general surgeon in a seriously ill patient in emergency. We review 7 cases of giant peptic perforations closed by a free omental plug. PMID- 10867453 TI - Accuracy of endoscopic ultrasonography in preoperative staging of esophageal carcinoma. AB - BACKGROUND: With the advent of stage-adapted multimodal regimens for many gastrointestinal malignancies, accurate staging has become of utmost importance. In esophageal cancer, endoscopic ultrasonography (EUS) emerged as standard to determine T and N stage. OBJECTIVE: Since growth patterns of squamous carcinoma (SC) differs from adenocarcinoma (AC) and lymph nodes are located at various distances from the esophagus in a horizontal plane, we studied the accuracy of esophageal EUS as a function of tumor type and localization of the tumor within the esophagus. RESULTS: Overall staging accuracy was 79% for T and for N staging. Staging was more accurate for T3/4, when compared to T1/2 tumors, and for SC when compared to AC. Histological T1/2 stages were overstaged by EUS in 8/17 patients, mostly in patients with AC (6/10). The sensitivity of retrosternal pain and of dysphagia for extramural disease was 57 and 92% respectively, the specificity of pain for extramural disease was 73%, and of dysphagia 36%. Preoperative weight loss in this series correlated linearly with tumor stages. CONCLUSIONS: The accurate preoperative staging of T2 esophageal endodermal malignancies is crucial for treatment stratification but difficult to achieve by visual analysis of endosonography alone. Postacquisition processing of echoendosonographic images might further increase the accuracy of echoendosonography and aid in the critical differentiation of T2 versus T3 esophageal malignancies. Preoperative weight loss and retrosternal pain are good clinical indicators of extramural disease. PMID- 10867454 TI - Simple closure or vagotomy and pyloroplasty for the treatment of a perforated duodenal ulcer: comparison of results. AB - BACKGROUND/AIMS: Treatment of the perforated duodenal ulcer continues to be a controversial subject. The purpose of our study was to compare the results of simple closure of perforated duodenal ulcer versus treatment by truncal vagotomy and pyloroplasty. METHODS: We present a prospective and randomized study of 207 patients who underwent surgical treatment due to perforated duodenal ulcer. In 117 patients the surgical treatment was simple closure and postsurgery medical treatment with proton pump inhibitors for 1 month, and in 90 patients vagotomy and pyloroplasty with no additional medical treatment. RESULTS: We applied the Visick scale in order to compare postsurgery results. The postoperative morbidity and mortality rates were the same with both techniques. Statistically, in both cases, no significant differences were found in postsurgery symptomatology. The different rates of recurrent ulcers and the reinterventions due to recurrent ulcers presented no significant statistical values. CONCLUSION: We conclude that simple closure remains the selected treatment in the majority of patients who present with a perforated duodenal ulcer. The operation is a simple and safe procedure. PMID- 10867455 TI - Preoperative ERCP approach to common bile duct stones: results of a selective policy. AB - BACKGROUND: In a previous study, we made a plea for more selective indications for preoperative ERCP in patients with gallstones based on the results obtained from a liberal policy. Following 3.5 years of implementing this selective policy, a report on the results are presented here. This study was performed in a referral academic hospital. METHODS: Between June 1994 and December 1997, 328 patients underwent cholecystectomy because of symptomatic cholelithiasis. Absolute indications for preoperative ERCP were: acute cholangitis (4 patients); obstructive jaundice (22 patients); gallstone pancreatitis (within the first 24 h in 14 patients), and wide common bile duct (CBD, >8 mm) with suspicion of stones in the biliary tree (2 patients). RESULTS: In 42 patients (12.8%) a preoperative ERCP was performed for these indications. Stones were found in the CBD in 30 patients and edema in the papilla in 2 patients (total 76.2%). The stones could be extracted by endoscopic sphincterotomy in 24 of the 30 patients (80%). Complications were seen in 7 patients (16.7%). All these complications (bleeding of the papilla in 4 and mild pancreatitis in 3 patients) could be treated conservatively. During a mean follow-up of 2.5 years, CBD stones could be demonstrated postoperatively in 3 patients (0.3%). No mortality was observed in this series. CONCLUSIONS: The results of this selective policy included the expected outcome of a significant reduction in the number of ERCPs performed from 29 to 12.8% (p < 0.001, chi(2) test) and a better yield of stones, from 29 to 76.2% of the patients. The mortality of the procedure decreased from 2 to 0% whereas morbidity remained the same. This selective policy seems adequate for the preoperative assessment of CBD stones. PMID- 10867456 TI - Prostaglandin E(1) continuous hepatic arterial infusion in the treatment of postoperative acute liver failure: basic study on hepatic hemodynamics and clinical application. AB - AIM: In the treatment of severe liver damage, it is of greater advantage to administer prostaglandin E(1) (PGE(1)) directly to the liver compared with systemic intravenous infusion, because of its high inactivation rate in the lungs. In comparison with intraportal infusion, hepatic arterial infusion is more advantageous because of its easier and safer accessibility. This study was designed to prove the superiority of hepatic arterial infusion to intravenous infusion. METHODS: Changes in hepatic hemodynamics and oxygen delivery accompanying PGE(1) infusion using both methods were investigated in pigs. In addition, continuous hepatic arterial infusion was applied in 3 cases of postoperative acute liver failure, for patients in whom other conventional treatments like plasma exchange failed to improve the functioning of the liver. RESULTS: Hepatic arterial flow increased significantly accompanying hepatic arterial infusion of PGE(1) at a rate of 0.1 microg/kg/min compared with intravenous infusion at the same rate in pigs. Such an increase resulted in elevation of total hepatic blood flow and oxygen delivery to the liver. Correspondingly, bile flow significantly increased accompanying hepatic arterial infusion of PGE(1). Continuous hepatic arterial infusion was applied in 3 cases of postoperative acute liver failure. The infusion was continued for 7-9 days at a rate of 0.01 microg/kg/min without any complications through heparin-coated catheters inserted via the femoral artery. Significant increase in bile flow was observed in 2 cases in whom bile was collected, serum total bilirubin began to decrease in all these 3 cases, and the patients recovered from acute liver failure. CONCLUSION: Hepatic arterial infusion of PGE(1) is very useful and effective in the treatment of acute liver failure. PMID- 10867457 TI - Proximal and distal segments of the possum sphincter of Oddi respond differently to neural and cholecystokinin octapeptide stimulation in vitro. AB - BACKGROUND/AIMS: Previous studies have demonstrated separate pancreatic duct (PD) and bile duct (BD) components of the sphincter of Oddi (SO) and suggested distinct proximal and distal functional segments. This study was designed to determine if proximal and distal segments of the BD component of the SO (BD-SO) and PD component of the SO (PD-SO) responded equally to (1) activation of SO duodenal neural pathways, and (2) exogenous cholecystokinin octapeptide (CCK-8). METHODS: Intact SO-duodenum preparations from Australian brush-tailed possums (n = 6) were mounted in organ baths. SO activity was recorded from the proximal and distal segments of BD-SO and PD-SO +/- electrical activation of duodenal nerves at two separate sites. Full thickness muscle strips from the proximal and distal segments of the BD-SO and PD-SO were prepared (n = 8), mounted in organ baths, and exposed to CCK-8 (10(-9)- 10(-6) M), +/- tetrodotoxin. RESULTS: Activation of duodenal nerves evoked different responses in some segments of the BD-SO and PD SO, depending on the site of duodenal electrical stimulation. CCK-8 induced a concentration-dependent, tetrodotoxin-insensitive decrease in the contraction amplitude of SO muscle strips from the proximal but not the distal SO. BD-SO and PD-SO strips were not different. CONCLUSIONS: The SO is composed of BD and PD components each of which contains proximal and distal segments that can respond independently to appropriate stimuli. PMID- 10867458 TI - Influence of gastrointestinal hormones on tumor microcirculation of experimental pancreatic cancer in the rat. AB - BACKGROUND/AIMS: Gastrointestinal hormones influence the microcirculation in the normal pancreas. In the present study, we studied the effect of cerulein and somatostatin on pancreatic cancer microcirculation after orthotopic and nonorthotopic tumor implantation. METHODS: In 36 male Lewis rats (150-180 g) induction of a ductlike pancreatic cancer was achieved by intrapancreatic or intraperitoneal tumor fragment interposition between two inert transparent polymethyl methacrylate plates. After 4 weeks, intravital microscopy of the tumor microcirculation was performed in a temperature-controlled immersion chamber. The animals received 5 microg/kg cerulein or 3 mg/kg somatostatin for 1 h intravenously. The erythrocyte velocity in normal pancreatic capillaries or in tumor vessels was measured. RESULTS: The erythrocyte velocity in the capillaries of the normal pancreas was 1.01 +/- 0.11 mm/s at baseline and increased to 1.64 +/- 0.09 mm/s after cerulein stimulation (p = 0.007). Pancreatic cancer vessels demonstrated no increase in erythrocyte velocity after orthotopic (baseline 0.95 +/- 0.14 mm/s, after 1 h 0.86 +/- 0.13 mm/s; n.s.) and nonorthotopic tumor implantation (baseline 0.91 +/- 0.12 mm/s, after 1 h 0.95 +/- 0.14 mm/s; n.s.) after cerulein stimulation. Somatostatin decreased the erythrocyte velocity both in normal pancreas (baseline 0.87 +/- 0.02 mm/s, after 1 h 0.60 +/- 0.07 mm/s; p = 0.01) and in pancreatic cancer (baseline 0.85 +/- 0.20 mm/s, after 1 h 0.63 +/- 0.18 mm/s; p = 0.02) after orthotopic tumor implantation. There was no effect of somatostatin after nonorthotopic tumor implantation (baseline 0.90 +/- 0.10 mm/s, after 1 h 0.88 +/- 0.14 mm/s; n.s.). CONCLUSION: These data suggest that pancreatic cancer microcirculation lacks physiological blood flow control by stimulatory hormones, in contrast to the normal pancreas. PMID- 10867459 TI - No effect of bolus glutamine supplementation on the postresectional adaptation of small bowel mucosa in rats receiving chow ad libitum. AB - OBJECTIVE: Early postoperative enteral feeding has been reported to stimulate intestinal mucosa proliferation. Dietary components influence the intestinal adaptive response after resection and glutamine is a preferential nutrient to enterocytes. The purpose of this study was to evaluate the effects of bolus glutamine supplementation on intestinal adaptation. METHODS: Male Wistar rats underwent a 65% small bowel resection. The rats were divided into three groups receiving glutamine 2 g/kg/day, isonitrogenous glycine or saline by gavage for 10 days. All the rats were provided with ordinary rat chow ad libitum. Sampling was done 10 days after resection. Animals fed ordinary rat chow without surgery or specific treatment served as control. RESULTS: Mucosal wet weight, DNA, RNA, protein contents and sucrose activity, as well as villus height increased in the ileal remnant. No significant differences in any of these parameters or body weight could be found between the three groups. CONCLUSION: Postoperative enteral bolus glutamine supplementation at a dose of 2 g/kg b.w. did not enhance the adaptation of the residual intestine 10 days after massive intestinal resection in the rat. PMID- 10867460 TI - Early perioperative results and surgical recurrence after strictureplasty and miniresection for complicated Crohn's disease. AB - BACKGROUND/AIMS: Strictureplasty (SP) or miniresective 'bowel-sparing' techniques (MR) can prevent the risk of intestinal stomia and short bowel syndrome in patients affected by Crohn's disease (CD). The aim of this study was to analyze the perioperative morbidity and mortality in 104 of 138 consecutive patients treated for CD complications using bowel-sparing techniques. We also considered the factors that may be related to the risk of perioperative complications and the long-term outcome. METHODS: One hundred and four patients were treated with SP and/or MR and then included in a prospectively maintained database. The factors claimed to influence perioperative complications were analyzed using Fisher's exact test for categorical observations and the Mann-Whitney U test for continuous variables. A multivariate analysis, using logistic regression, and a long-term time-to-event analysis using the Kaplan-Meier function, were also performed. RESULTS: Perioperative mortality was nil. In relation to the 6 postoperative complications (5.8%), 4 patients underwent minimal bowel resection (MR), 1 a MR with SP, and 1 SP alone. Three of these patients (2.9%) needed reoperation for septic complications, and 3 (2.9%) were treated as outpatients for enterocutaneous fistulas. A correlation (p < 0.05) was found between low serum hemoglobin levels and postoperative complications at univariate and multivariate analyses. The 5-year surgical recurrence-free rate was 75% overall, 73% for patients treated with SP, 78% with MR, and 77% with MR + SP. CONCLUSIONS: Postoperative complications are not related to conservative or miniresective surgery even when active disease is present at the resection margins or the site of SP. The higher risk reported for patients with low serum hemoglobin and hematocrit levels suggests that surgeons should consider using preoperative iron and vitamin support, parenteral nutrition and erythropoietin therapy, when necessary, in those cases. Our postoperative morbidity, mortality and long-term surgical recurrence rate results support the efficacy and safety of SP and MR surgery in the treatment of complicated CD. PMID- 10867461 TI - Sacral magnetic stimulation in paradoxical puborectalis syndrome. AB - BACKGROUND/PURPOSE: Our earlier studies have demonstrated that sacral magnetic stimulation (MS) in the canine model, in healthy volunteers and in constipated subjects brought about a rise in rectal pressure and a decline in rectal neck (anal canal) pressure as well as rectal evacuation. Based on these results, we studied the effect of sacral MS on defecation in patients with paradoxical puborectalis syndrome (PPS). METHODS: Eleven subjects (8 women, 3 men; age 36-53 years) with PPS were enrolled in the study. The magnetic coil was placed on the back with its center located between L4 and L5. Stimulation parameters were set at 70% of maximum intensity, 40 Hz frequency and 2 s burst length with 2 s off. During MS, the rectal neck and gastric (intra-abdominal) pressures were measured. The procedure was performed in the empty and the full rectum using the balloon expulsion test in the latter. RESULTS: MS of the empty and balloon-filled rectum brought about a rise in the rectal pressure (p < 0.001), decline in rectal neck pressure (p < 0. 001) and no significant change in intragastric pressure (p > 0.05). The balloon was expelled by all patients. CONCLUSIONS: Sacral MS succeeded in expelling the water-filled rectal balloon. The method is simple, easy, noninvasive, nonradiologic and can be performed on an outpatient basis in the treatment of PPS. PMID- 10867462 TI - The initial approach to anorectal abscesses: fistulotomy is safe and reduces the chance of recurrences. AB - BACKGROUND/AIMS: Anorectal abscesses are most frequently based on a coexistent fistula in ano. Whether these should be searched for and excised initially or not remains controversial. Our aim was to determine which approach has less recurrences and carries a lower risk of continence disorders. METHODS: 158 patients with an anorectal abscess or anal fistula were identified in our institution over a period of 75 consecutive months. The records and follow-up questionnaires of 131 patients were evaluable. The mean follow-up period was 40 (range 3-78) months. RESULTS: When fistulotomy was performed at the time of draining the abscess, the recurrence rate could be reduced, in comparison to incision and drainage alone, from 34 to 4% (p = 0.007). In the group of patients undergoing surgery for a recurrence, the recurrence rate could even be reduced from 67 to 0% (p = 0.03) by simultaneous fistulotomy. A total of 4 of the 131 patients (3%) developed incontinence of liquid stool and flatus, but no incontinence of solid stool occurred. Incontinence did only occur after recurrent disease. CONCLUSIONS: The number of recurrences requiring surgery can be significantly reduced by initial fistulotomy. The risk to develop incontinence increases with recurrent anorectal disease, not with careful fistulotomy. PMID- 10867463 TI - Superior mesenteric and portal vein thrombosis following laparoscopic nissen fundoplication. AB - This case report describes superior mesenteric and portal vein thrombosis after laparoscopic Nissen fundoplication. As a thromboembolic prophylaxis, 2,500 IU of dalteparin was given preoperatively. After postoperative day 19, the patient experienced gradually increasing abdominal pain, mostly related to meals. Physical examination and laboratory tests were normal. CT scan revealed a portal and superior mesenteric vein thrombosis. Dalteparin and warfarin treatment was started, and symptoms relieved rapidly. In a control Doppler ultrasound 1 month after the onset of the treatment, a good flow in the portal and superior mesenteric vein was seen. Possible mechanisms are discussed. PMID- 10867464 TI - Carcinoid tumour in a caecal duplication cyst. AB - Intestinal duplication cysts are rare congenital anomalies that may occasionally undergo neoplastic change. We report the case of a 30-year-old woman who was diagnosed to have a caecal duplication cyst. The cyst was excised and histology revealed the presence of a 10 mm diameter carcinoid tumour within the cyst wall. There was no evidence of metastatic spread and the patient remains well after 2 years follow-up. The 3 previously reported cases of carcinoid tumour arising within duplication cysts are reviewed. PMID- 10867465 TI - Rare case of left-sided ureteroduodenal fistula. AB - BACKGROUND/AIMS: Ureteroduodenal fistulas are rare and only 11 cases have been reported in the literature since 1918. Diagnosis requires careful observation of symptoms. METHODS: The case presented demonstrates a 68-year-old female with left sided ureteroduodenal fistula confirmed by CT scan. A duodenal fistula was localized and an atrophic left kidney was identified and repaired. RESULTS: Nephroureterectomy was performed and an omental patch was used for the repair. No complications were encountered during the postoperative course. CONCLUSIONS: Recurrent chronic urinary tract infection, pyuria and hematuria can indicate this rare disease. Early testing and detection can improve the chances of renal preservation. PMID- 10867466 TI - Agenesis of the gallbladder found at laparoscopy in an adult patient with cardiac congenital malformation. AB - We report a case of gallbladder agenesis in a 30-year-old woman affected by a cardiac congenital malformation who had been operated on at the age of 12. The patient was sent for laparoscopic cholecystectomy due to a preoperative diagnosis of cholelithiasis using clinical and instrumental examinations such as ultrasonography and cholangiography. During laparoscopy, the gallbladder was not found, and laparotomy with intraoperative cholangiography and ultrasonography was performed which also resulted negative. The preoperative possibility of a diagnosis of gallbladder agenesis, the association with other malformations and the steps to be taken to discover agenesis of the gallbladder are discussed. PMID- 10867467 TI - Recurrent idiopathic perforations of the small intestine. AB - BACKGROUND/AIMS: Idiopathic perforation of the small intestine is extremely rare. The definition depends on the absence of any detectable pathology that could be responsible for the perforation. This study was undertaken to outline the criteria for determination of the condition. METHODS: Case report and review of the literature. RESULTS: A 50-year-old male patient underwent surgery including laparostomy and planned reexplorations for multiple recurrent small intestinal perforations of unknown origin; the patient made an uneventful recovery. Extensive investigations were carried out to search for any pathology that could explain the occurrence of perforations, but yielded no results. CONCLUSIONS: Further studies will possibly give more information concerning idiopathic perforation of the small intestine, provided that judicious criteria for its determination are applied. PMID- 10867468 TI - Intraluminal duodenal diverticulum causing acute pancreatitis: CT scan diagnosis and review of the literature. AB - BACKGROUND: Intraluminal duodenal diverticulum is a rare congenital anomaly. First described by Boyd in 1845, no more than 100 cases have been reported up to now: only 17 are associated with acute pancreatitis. METHODS: A new case of intraluminal duodenal diverticulum with acute pancreatitis is reported and the literature about this association reviewed. RESULTS: The diagnosis was made by helical CT scan. The pathogenesis of pancreatitis was possibly due to a pure duodenal content reflux through the papilla of Vater. The patient was successfully treated by surgery. CONCLUSIONS: Intraluminal duodenal diverticulum is a rare but curable cause of pancreatitis, usually affecting young people. We describe, for the first time, its unusual helical CT imaging with two-dimensional reformations. PMID- 10867469 TI - Intermittent activity-induced hemobilia caused by liver hemangioma. AB - BACKGROUND: Intestinal bleeding of unknown origin can lead to a difficult workup. Abdominal colic, melena/hematemesis, and jaundice represent the pathognomonic triad for hemobilia, but clinical presentation and etiology of this entity are varying. Seldom all of these symptoms are present, and rarely does hemobilia cause melena or hematemesis. Often the correct diagnosis is missed. Patients frequently have a long history of complaints and inadequate therapy. CASE REPORT: We report on a patient who complained of repeated, severe epigastric pain and massive melena induced by exercise activity. After 2 years of complaints and an unnecessary operation, ultrasound detected a liver hemangioma. It was supposed that the hemangioma was causing hemobilia during strenuous physical activity. The patient underwent a partial liver resection to eliminate the hemangioma. All complaints resolved, and the patient remained asymptomatic postoperatively. CONCLUSIONS: Physicians should be aware of hemobilia as a rare cause of upper gastrointestinal bleeding, especially if esophagogastroduodenoscopy cannot demonstrate any bleeding source. Ultrasound is able to visualize many diseases leading to hemobilia and should be integrated into the early workup of unclear intestinal bleedings. PMID- 10867470 TI - Electrocoagulation of perianal warts: a word of caution. AB - BACKGROUND: Perianal warts are common, and may be extensive. Electrocoagulation is a recognised management option. METHOD: A 20-year-old male underwent electrocautery of extensive perianal warts. He presented 3 months postoperatively with constipation and inability to defecate. Examination revealed severe perianal stricture, which necessitated a defunctioning colostomy. RESULT: A gradual and spontaneous resolution of the stricture was observed over the following 18 months. Closure of the colostomy was followed by satisfactory anal function. CONCLUSION: Electrocautery of extensive perianal warts should be used with caution. Preservation of healthy skin bridges between lesions is essential if perianal stricture is to be avoided, and may best be achieved by sharp scissors dissection. PMID- 10867471 TI - Intrahepatic biliary cystadenoma causing luminal common bile duct obstruction. AB - BACKGROUND: Biliary cystadenomas are rare cystic tumours that arise in the liver or less frequently in the extrahepatic biliary system. They are commoner in middle-aged women, their most favoured site is the right hepatic lobe. METHODS: Case report and review of the literature. RESULTS: We present only the second case of an intrahepatic cystadenoma causing luminal obstruction of the common bile duct. Clinical presentation is often non-specific and can prove to be a diagnostic challenge. CONCLUSION: Wide local excision of biliary cystadenomas is recommended, with regular radiological follow-up. PMID- 10867472 TI - Granular cell tumor at the hepatic duct confluence mimicking Klatskin tumor. A report of two cases and a review of the literature. AB - BACKGROUND: Granular cell tumors are rare tumors most often located in the oral cavity, skin or subcutaneous tissue. The occurrence of this tumor in the biliary tree is extremely rare. METHODS: Two patients are described presenting with biliary obstruction due to a tumor at the hepatic duct confluence. One patient is a 38-year-old white male with concomitant cutaneous granular cell tumors, and the other a 50-year-old white female. RESULTS: Hilar excision was performed in both patients. Histopathology of the tumors revealed a proliferation of cells with granular cytoplasm, diagnosed as granular cell tumor. CONCLUSION: At preoperative examination, hilar granular cell tumors are difficult to differentiate from cholangiocarcinoma, sclerosing cholangitis or more common benign biliary tumors. Treatment consists of surgical excision after which prognosis is favorable. PMID- 10867473 TI - In-continuity fundic bypass for palliation in unresectable carcinoma of the oesophagus: a previously unreported procedure. AB - Surgical bypass for the palliation of dysphagia in patients with unresectable oesophageal carcinoma continues to be an option in developing countries, as the cost of a good quality endo-prosthesis is well beyond the means of most patients. One such case is presented in which an in-continuity fundic bypass (without resection of the lesser curvature and cardia, thereby not disconnecting the oesophago-gastric junction) was made with gratifying results with regard to quality of life. Awareness of this previously unreported procedure is important because it adds to the armamentarium of surgeons wanting to provide palliation for dysphagia and aspiration in patients with unresectable carcinoma of the oesophagus. PMID- 10867474 TI - Esophageal leiomyomatosis in a woman with a history of vulvar leiomyoma and Barrett's esophagus: a case report and review of the literature. AB - BACKGROUND: The diagnosis and treatment of esophageal pathology remains a challenge despite advances in preoperative endoscopy, radiographic staging, and perioperative care. CASE REPORT: In this article, we present an interesting case of esophageal leiomyomatosis in a woman with a history of vulvar leiomyoma and Barrett's esophagus. This paper represents the first reported simultaneous occurrence of these three pathologic entities in the English literature. CONCLUSIONS: The clinical presentation and characteristic pathologic findings in patients with esophageal leiomyomatosis are reviewed. Diagnostic and therapeutic approaches to esophageal masses are discussed including the indications for esophageal resection. PMID- 10867475 TI - Lymphoepithelial cyst in the pancreas: a case report and review of the literature. AB - BACKGROUND: Lymphoepithelial cysts of the pancreas constitute a rare clinicopathologic entity. CASE REPORT: We report a case of lymphoepithelial cyst of the pancreas and review the world literature. RESULTS: Lymphoepithelial cysts are true pancreatic cysts lined by squamous epithelium and surrounded by mature lymphoid tissue. The cyst arises typically in middle aged men, and is usually asymptomatic or causes nonspecific abdominal complaints. There is no specific serologic marker for this entity. None of its radiologic characteristics can help differentiate it from other cystic lesions of the pancreas. Fine-needle aspiration cytology may be able to suggest its benign nature and identify it as a true cyst of the pancreas. The outcome after surgical excision is uniformly good with good symptom control and no recurrences. RECOMMENDATIONS: In the symptomatic patient or the asymptomatic patient with acceptable surgical risk a simple cyst excision should be performed after verification of the diagnosis with frozen section. In the asymptomatic patient with a high surgical risk, in whom fine needle aspiration suggests the diagnosis of a lymphoepithelial cyst, observation of the lesion is recommended. When simple cyst excision is technically not possible, extensive resections/reconstructions should be avoided and drainage/bypass procedures may be considered. PMID- 10867476 TI - Editorial Note: Modern Giants in Surgery. PMID- 10867477 TI - D.J. du Plessis: a surgical giant in southern Africa. PMID- 10867478 TI - Fetal impact of cholestasis of pregnancy: experience at Tenon Hospital and literature review. AB - Cholestasis of pregnancy is a liver disorder that occurs during the second half of pregnancy, causing pruritus and elevated serum bile acid levels. Its etiology remains unknown but probably involves vascular and humoral immune responses, mediated by bile acids. This disorder is associated with substantially increased fetal morbidity and mortality. The most satisfactory treatment consists in delivering the fetus as soon as pulmonary maturation has occurred. PMID- 10867479 TI - Neonatal periventricular leukomalacia preceded by fetal periventricular echodensity. AB - OBJECTIVE: The purpose of this prospective study is to verify whether fetal periventricular echodensity (PVE) precedes neonatal periventricular leukomalacia (PVL). METHODS: Fetal brains were studied with transvaginal scan in 63 high-risk fetuses from 17 to 32 weeks of pregnancy, PVE echogenicity was quantified with ultrasonic histogram, and neonatal brains and clinical courses were studied after birth. RESULTS: No fetal cystic PVL was found, instead, fetal PVE was detected in 42 fetuses. The quantified echogenicity value was higher in PVE than in normal brain. Four cases developed neonatal PVL among 28 preterm and 1 among 14 term births. Neonatal PVL developed in the 23 cases of persistent fetal PVE, whereas no neonatal PVL was found when fetal PVE was negative or disappeared. Cord compression signs were common in PVL cases. CONCLUSION: Neonatal PVL was preceded by antepartum persistent fetal PVE in the present study. PMID- 10867480 TI - Mid-trimester thoracoamniotic shunting for the treatment of fetal primary pleural effusions in a twin pregnancy. a case report. AB - Thoracoamniotic shunting has been described as having a beneficial role in the antenatal management of primary pleural effusions in singleton pregnancies. We report a case of a twin pregnancy in which progressive pleural effusions and hydrops were diagnosed in one of the fetuses at 16 weeks of gestation. An initial evaluation ruled out underlying genetic and anatomic abnormalities in both twins. At 19 weeks gestation, the first procedure of bilateral thoracoamniotic shunting was performed in the affected fetus, subsequent to which the lungs re-expanded and the hydrops resolved. Three additional shunt replacements and one therapeutic amniocentesis were required on follow-up. At 35 weeks, labor was induced. The first fetus (healthy) was delivered vaginally and the second fetus (affected) was delivered by cesarean section. Both neonates are healthy at one year follow-up. PMID- 10867481 TI - Discrepancy between cytogenetic and FISH results on an amniotic fluid sample of 45,X/46,X,idic(Y)(p11). AB - The presence of abnormal ultrasound markers showing a thick nuchal fold with short middle phalanx of the fifth finger in an otherwise normal-appearing female fetus led to the sampling of amniotic fluid at 16 weeks gestation. Cytogenetic analysis with routine G-banding showed a 45,X karyotype in all 20 cells analysed from two flasks. However, fluorescent in situ hybridization on uncultured cells showed presence of a Y signal in 9 cells, 11 cells showing a single signal for the X. A cytogenetic analysis of the fetal blood at 23 weeks confirmed the presence of two cell lines, 45,X and 46,X, idic(Y)(p11). The couple opted to have the pregnancy terminated. However, the fetus was not available to carry out confirmatory tests. PMID- 10867482 TI - Intracardiac echogenic focus: no apparent association with structural cardiac abnormality. AB - OBJECTIVE: The aim of our study was to evaluate whether intracardiac echogenic foci (ICEFs) may be associated with increased risk for structural cardiac anomalies in the low-risk population. METHODS: During a 24-month period, 3,744 low-risk patients were prospectively screened for ICEFs by prenatal sonography. The study group was composed of 138 fetuses (3.7%) with ICEF. The control group was composed of 167 fetuses without ICEF. In all fetuses a complete echocardiographic evaluation was performed. RESULTS: Among the 138 fetuses in the study group, 108 (78%) ICEFs were found in the left ventricle, 25 (18%) were found in the right ventricle, and 5 (4%) were found to be bilateral. No statistically significant difference was found between the study and the control group regarding the presence of cardiac anomalies. Only 1 case (0.7%) of pulmonic stenosis was found in the study group, compared to 1 case (0.6%) of bicuspid aortic valve in the control group. CONCLUSIONS: We conclude that ICEFs found in low-risk patients are not associated with a significant increase in the risk of cardiac anomalies. PMID- 10867483 TI - Contribution of transvaginal ultrasonography and fetal cerebral MRI in a case of congenital cytomegalovirus infection. AB - Cytomegalovirus is the most common cause of congenital viral infection. In utero this infection is usually suspected on the basis of ultrasound findings. We present a case in which routine ultrasound examination demonstrated a decrease in fetal cephalic dimensions at 32 weeks' gestation in an asymptomatic patient. Transvaginal ultrasound revealed echogenic vessels in the thalami and lesions in the subependymal region. Suspected diagnosis of fetal cytomegalovirus infection was confirmed by positive titers of anti-cytomegalovirus-IgM antibodies in fetal blood and amniotic-fluid PCR studies. Fetal cerebral MRI demonstrated parenchymal atrophy and polymicrogyria. The parents decided to terminate the pregnancy, and necropsy confirmed the diagnosis. Suspicion of CMV fetal infection should prompt transvaginal ultrasound and fetal brain MRI. PMID- 10867484 TI - Influence of amnioinfusion in a model of in utero created gastroschisis in the pregnant ewe. AB - OBJECTIVE: Recent studies on the management of human fetal gastroschisis have produced two major findings: (1) there is an inflammatory response in the amniotic fluid of these fetuses, and (2) amniotic fluid exchange designed to disrupt the inflammatory loop seems to have a favorable impact on the immediate and late outcome of these early operated neonates. To test this hypothesis, we used serial amniotic fluid exchanges in a model of gastroschisis developed in the ewe. METHODS: Gastroschisis was created at midgestation in 21 lamb fetuses by an in utero technique. Saline was amnioinfused in some fetuses every 10 days to term. Fetuses were sacrificed on day 145 by cesarean section. Extra-abdominal bowels with fibrous peel were processed for histologic examination. Comparisons were done between fetuses without gastroschisis (controls), fetuses with gastroschisis and amnioinfusion, and fetuses with gastroschisis without amnioinfusion. RESULTS: Of 21 fetuses operated, 8 died in utero or were stillborn; 5 were not amnioinfused, and 8 underwent amnioinfusion. Thickness of bowel muscularis (micrometer) was 92.6 +/- 20.2 for controls, 126.2 +/- 21 for the amnioinfused fetuses, and 182.8 +/- 58.3 for the nonamnioinfused fetuses (p = 0.001). The same significant results were obtained for thickness of serous fibrosis (p = 0.02) and plasma cell infiltration (p = 0.015). CONCLUSIONS: We have created a model of gastroschisis suitable for experimentation in the fetal sheep. Our amnioinfusion data in this model indicate a clear improvement of the deleterious process. This finding correlates well with recent data on amnioinfusion as a therapeutic approach to human gastroschisis. PMID- 10867485 TI - Usefulness of fluorescence in situ hybridization for the diagnosis of Turner mosaic fetuses with small ring X chromosomes. AB - OBJECTIVE: To emphasize the usefulness of fluorescence in situ hybridization (FISH) techniques on uncultured amniocytes for the diagnosis of abnormal mosaic karyotypes. METHODS: In the course of three prenatal diagnoses, specific fluorescent probes, coding, respectively, for chromosomes X, Y, 18, 13, and 21, were applied on amniocyte preparations directly after amniocentesis. At least 50 nuclei were counted in each case. Parallel to the FISH procedure, cell cultures were set up in order to obtain karyotypes. FISH and cytogenetic results were then compared. RESULTS: In each case, FISH showed an abnormal mosaic chromosomal constitution, 45,X/46,XX, which was related to the existence of tiny ring X chromosomes in karyotypes. CONCLUSION: Because very small ring X chromosomes can escape identification when standard cytogenetic techniques are used alone, we show that misdiagnosis can be avoided when FISH is performed beforehand. PMID- 10867486 TI - Parvovirus as a differential diagnosis of hydrops fetalis in the first trimester. AB - We report a case of fetal parvovirus B19 infection which appears to have resulted in hydrops in the first trimester. An ultrasound scan performed at the booking visit of a woman in the first trimester showed generalised oedema. Karyotyping to exclude a fetal abnormality was normal. Maternal radioimmunoassay serological investigations showed parvovirus B19 IgG and IgM. Post-mortem analysis revealed intranuclear inclusion bodies typical of parvovirus in the erythroid cells in the liver and in the myocardium. PMID- 10867487 TI - The development of a genetic profile of placental gene expression during the first trimester of pregnancy: a potential tool for identifying novel secreted markers. AB - OBJECTIVES: Many of the maternal serum markers used in prenatal screening have been developed or evolved based on serendipity. This study determines the feasibility of developing a genetic profile of placental gene expression during early pregnancy; such a gene repertoire may serve as a tool for identifying novel secreted markers. METHODS: RNA fingerprinting was used to produce a differential expression map in normal aborted placentae of weeks 9 and 13. RESULTS: Out of 212 gene expression differences, 115 were up-regulated at week 9 and the remaining 97 up-regulated at week 13. Ninety-four were found to be previously characterised genes and 118 were expressed sequence tags or novel genes. Seven of the known genes were found to be secreted proteins and included well-characterised pregnancy markers. mRNA levels of these secreted proteins were found to correlate with levels found in maternal serum. CONCLUSION: This approach enabled the identification of known secreted markers leaving open the possibility of finding new markers. With the human genome project nearing completion, it will be vital to use a systematic approach to understand the actual pattern of gene expression during pregnancy. This will also allow a more ordered and precise identification of novel prenatal diagnostic markers. PMID- 10867488 TI - Congenital cystic adenomatoid malformation of the lung: antenatal ultrasound findings and fetal-neonatal outcome. Fifteen years of experience. AB - Seventeen cases of congenital cystic adenomatoid malformation of the lung (CCAM) are reported. They were followed up over a period of 1 month to 15 years. Diagnosis was made by prenatal ultrasound. Our purpose was to evaluate the fetal neonatal outcome and the prognostic elements observable through ultrasound techniques, and to compare all types of CCAM. The outcome observed ranged from total prenatal resolution to postnatal spontaneous regression of the lesion, to complications due to the presence of nonimmune fetal hydrops (NIFH), intrauterine death and the necessity of surgical intervention. In our experience only hydrops represented a negative predictor of outcome since death occurred in all cases with this pathology. In the absence of NIFH, counselling should stress the prevalence of a positive outcome, even in cases of surgical intervention. PMID- 10867489 TI - New classification for analysing uterine Doppler waveforms during pregnancy: is it useful? PMID- 10867490 TI - [Developments in minimally invasive breast surgery - overview and our own experience: new diagnostic and therapeutic challenges in breast cancer]. AB - New diagnostic and therapeutic modalities in breast cancer are necessary because nowadays an increased number of suspicious breast lesions are referred to breast clinics for a histological diagnosis. In the future more than one quarter of patients might present with a preinvasive lesion and more than half of the patients will have tumors less than 2 cm. Mammography screening helps to find more preinvasive lesions. Therefore we need new tools for exact stereotactic breast biopsies in the outpatient setting, such as the advanced breast biopsy instrumentation (ABBI((R))) or the Mammotome((R)) system. For axillary clearance we need methods that lead to less morbidity. The detection of the sentinel lymph node is one of these new techniques. Endoscopic axillary clearance after liposuction also helps to reduce morbidity. Due to better visualization of the anatomic structures with axilloscopy less traumatic surgery is possible. We also combined these two new methods and described the endoscopic clearance of the sentinel lymph node in the axilla with the additional help of isosulfan blue. However, this combined method is not only time-consuming but also more expensive and shows no obvious advantage compared to the open sentinel technique. Therefore we stopped using the endoscopic sentinel technique and we now promote open sentinel lymph node biopsy without full axillary clearance when frozen section shows sentinel node-negative lymph nodes. PMID- 10867491 TI - [Sexuality in the second half of the life span]. AB - Compared to other research areas of sexology, the state of knowledge regarding sexual behavior and experiences in middle and older age is rather poor. This is due to not only the continuing taboo placed on questions regarding sexuality of elderly people, but also to limited understanding of factors influencing the sexual realm. The present review presents findings on sexuality in the second half of the life span from a number of differing disciplinary perspectives. Focus is on gender differences in sexual life and the influence of critical life events on sexuality in women in the second half of the life span. Apart from restrictive social norms and individual personality factors, physiological changes during the menopause can also have an effect on sexuality. The special situation of elderly women without partners and changes in partnership sexuality during this phase of the life span are discussed. Some recommendations for the field of gynecology are derived from a multidisciplinary approach. PMID- 10867492 TI - [The cardiotocographic score of Hammacher in computer analysis]. AB - OBJECTIVE: After the quantitative, fully computerized evaluation of cardiotocography (CTG) after G.S. Dawes had yielded reliable results, an investigation was to be carried out to see whether a semiquantitative CTG analysis together with a CTG score (K. Hammacher) produced similarly useful results, too. METHODS: The last 120 directly recorded intrapartum CTG minutes (HP 8040A, 80300A) of 393 vaginally delivered fetuses were evaluated by simple means (magnifying glass, dividers, Plexiglas sliding caliper). The number of turning points, the bandwidth and the mean frequency level were determined for each individual CTG minute. Further, all decelerations were registered numerically (start, end and depth of decelerations), both qualitatively (28 key positions) and quantitatively. The CTG score of Hammacher was programmed in Fortran IV and integrated as a subroutine into a larger software system. RESULTS: Score 4 was noted most frequently (35.4%) during the last 30 min ante partum. The score values 10, 11, 12, 14, 15, 16, 17 and 18 were not seen at all. On the basis of the 'percent distribution of points' of the three score components, it could be shown that the decelerations were numerically overrepresented in comparison with the two other factors. The rank correlation between the score value and the relevant blood pH of the umbilical artery was highly significant (p << 0.001). The scoring of the CTG rises perceptibly only above pH 7.15. The sensitivity of score < or =4 is 87.2% for pH 7. 10 and 100% for pH 7.00. CONCLUSIONS: The interpretation of the cardiotocogram by means of the Hammacher score, which is based on a semiquantitative evaluation, yields clinically useful results. These may further be improved by changing the score structure. Beside the fully computerized analysis, the semiquantitative CTG analysis is able to finally determine the reliability of this monitoring method. PMID- 10867493 TI - [Lichen sclerosus - present-day hormonal therapy]. AB - Several treatment modalities of lichen sclerosus have been described. Topical testosterone (2%) has been the therapy of choice with improvement of symptoms in 67-100% of the patients. Newer placebo-controlled studies report success rates for testosterone of 20% which is not much higher than for placebo (10%). Treatment with the very potent steroid-like clobetasol 0.05% has been shown to be much more effective with success rates of up to 75%, while corticosteroid-related side effects like skin atrophy have not been a major problem. PMID- 10867494 TI - [Laser conization and laser ablation of the transformation zone]. AB - OBJECTIVE: To evaluate postoperative courses and histological results after laser therapy of the transformation zone. METHOD: From 1993 to 1997, 136 laser interventions were performed (110 conizations and 26 vaporizations); 103 (76%) of these interventions were performed as outpatient procedures and 122 (89%) under local anesthesia. RESULTS: 94 (85%) of the 110 conizations showed tumor-free margins, whereas 12 (11%) had dysplastic changes in one or both margins. Follow up of these 12 patients revealed 5 normal cytologic controls, 4 cases of hysterectomy without residual disease, 1 case of CIN III in the hysterectomy specimen, 1 case of CIN I in a radical hysterectomy (cervical cancer stage Ib in the histology of the conus), and 1 patient was lost to follow-up. In 4 cases (4%), there was only a suspicion of marginal involvement. These patients had either a normal cytologic control (n = 3) or no residual disease in the hysterectomy specimens (n = 1). There were eight bleedings that had to be treated with bicoagulation. CONCLUSIONS: Laser therapy offers the possibility of a very precise circumcision at the ectocervical site under colposcopic control. It can mostly be performed as an outpatient procedure under local anesthesia, and the frequency of postoperative complications compares to the other available methods. If the margins show dysplastic changes, follow-up rarely reveals recurrent disease. Expectative management with close cytologic follow-up is, therefore, justified in such cases, if fertility should be maintained. PMID- 10867495 TI - [Ultrasound-guided biopsy of non-palpable breast lesions: correlation with the results of fine-needle aspiration biopsy]. AB - PURPOSE: The purpose of this study was the clinical evaluation of ultrasound guided biopsy in comparison with ultrasound-guided fine-needle aspiration biopsy of identical, non-palpable breast lesions. MATERIALS AND METHODS: From August 1997 until July 1998, 73 ultrasound-guided biopsies were performed in 66 patients with non-palpable lesions of the breast. In 18 patients (age 33-77 years) with 20 non-palpable lesions, fine-needle aspiration biopsy (20-G needle) and biopsy (18 G biopsy needle) were performed on a single occasion. This was the patient selection of our retrospective study. RESULTS: One malignant neoplasm was found among the 20 biopsied lesions, while the remaining 19 lesions were of a benign nature. In 20% of the cases, the material obtained by fine-needle biopsy was not sufficient for a cytologic diagnosis, while biopsy allowed a diagnosis in 19/20 cases. No complications were observed. CONCLUSIONS: Ultrasound-guided biopsy using an 18-G needle is a suitable method for the evaluation of non-palpable lesions that are only visible on ultrasound. It represents an attractive alternative to fine-needle aspiration in the absence of experienced cytologic diagnosticians. PMID- 10867496 TI - [Neurofibromatosis of the breast in a patient with Morbus von Recklinghausen]. AB - Neurofibromatosis is a rare autosomal dominant disorder with several subtypes. Common is the appearance of specific skin alterations. Neurofibromas occurring in the breast are very rare, and in such cases they are most common in the areolar area. A case of a 46-year-old woman with von Recklinghausen's disease of the breast is reported, and the literature will be discussed. PMID- 10867497 TI - [The future of the German language in science]. AB - The German language served the scientific community for decades during the 20th century. The shift to the English language was considered by some as a loss of cultural identity. A view difficult to support, since not only Germany but also countries like Switzerland and Austria are part of the cultural heritage. The development of the impact factor, which is derived predominantly from Anglo American journals, has also contributed to this situation. This may be of concern, since it influences the scientific future of young scientists from German-speaking countries. At present, no exclusively German-language journal in obstetrics and gynecology is listed in the Index Medicus anymore. If this means that the German-language scientific journals are not needed for the transmission of scientific data, they may evolve as organs for postgraduation studies. Interest in postgraduate papers only recently became evident in academic circles. In American CVs a rubric of postgraduate studies is separately provided. These are highly esteemed. The need for such postgraduate journals will be established when postgraduate studies become mandatory also in German-speaking countries. A lack of postgraduate papers may be overcome by an impact factor for postgraduate studies to encourage young academics to write such papers. Until German-language postgraduate papers are submitted in larger numbers, the editors of German language journals are referred to the Anglo-American literature where such articles are available. Since a scientific journal, even when there are large subscription numbers, requires advertisements of the pharmaceutical industry, there is a potential for a conflict of interest between physicians and advertisers. In general, it can be expected that the number of German-language journals will drastically decline. PMID- 10867498 TI - Localization of sialyl glycoconjugates in eosinophil-specific granules after degranulation stimuli. AB - Sialyl glycoconjugates, a group of the acidic glycoconjugates, are expressed in murine eosinophil-specific granules through their maturational stages. To clarify whether the sialyl glycoconjugates work as functional molecules in the process of eosinophil degranulation, we examined the localization of sialyl residues after degranulation stimulated with calcium ionophore A23187 or interleukin-5. Although sialyl residues were localized in the specific granules before degranulation, they were dissociated from the granules after the stimuli. These results suggest that sialyl glycoconjugates participate extensively in the process of eosinophil degranulation. PMID- 10867499 TI - Ecalectin/galectin-9, a novel eosinophil chemoattractant: its function and production. AB - We have recently succeeded in purifying and cloning a novel eosinophil chemoattractant, ecalectin/galectin-9, which belong to a rapidly growing galectin family. The eosinophil chemoattractant activity of ecalectin is potent and selective for eosinophils. The cellular source of ecalectin has been thought to be antigen-stimulated T cells. Herein, we will review the significance of its divalent galactoside-binding activity in ecalectin-induced eosinophil chemoattraction and the regulation of ecalectin expression in immune cells. PMID- 10867500 TI - A rapid and simple photometric assay for quantification of eosinophil chemotaxis. AB - Various methods have been devised to assay eosinophil chemotaxis. In this review, we introduced a rapid and sensitive eosinophil chemotactic assay system based on eosinophil peroxidase (EPO) determination. Eosinophils were semipurified by dextran sedimentation followed by Percoll gradient centrifugation. Eosinophil migration was measured by the Boyden blind-well chamber technique using a 96-well multiwell chamber and polycarbonate membrane filter. The number of migrated eosinophils was assessed by determination of the EPO activity of drop-off cells, and photometric measurement was performed with a microtiter plate reader. The contribution of neutrophil peroxidase to the assay is negligible, and the assay is sensitive enough to detect 200 eosinophils. The method is both less tedious and less laborious than conventional manual methods, and it allows a large number of samples to be analyzed within 4 h. When eosinophils were exposed to 15 chemokines and their migration was assayed using this method, we found that only CC chemokine receptor 3 ligands, such as eotaxin, induced significant eosinophil migration. PMID- 10867501 TI - Eosinophil-adhesion-inducing activity produced by antigen-stimulated mononuclear cells involves GM-CSF. AB - BACKGROUND: The initial step of eosinophil accumulation in allergic inflammation is adhesion of circulating eosinophils to vascular endothelial cells (EC). There is evidence that the adhesive property of circulating eosinophils is upregulated following antigen exposure. Although the exact mechanism remains to be established, cytokine(s) produced by antigen-stimulated mononuclear cells is (are) likely key factor(s). OBJECTIVE: The objective of this study was to examine the factor(s) responsible for eosinophil adhesion and migration induced by the antigen-stimulated mononuclear cells obtained from atopic asthmatics. METHODS: Peripheral blood mononuclear cells (PBMC) isolated from house-dust-mite-sensitive bronchial asthmatics were cultured for 96 h in the presence or absence of 1 microg/ml Dermatophagoides farinae (Df) antigen. Eosinophils were isolated from peripheral blood of healthy subjects. Eosinophil-adhesion-inducing activity in the culture supernatants of PBMC was examined by the ability to modify the adhesion of eosinophils to human pulmonary microvascular endothelial cells (HPMEC) in the presence or absence of anti-cytokine/chemokine antibodies. Eosinophil migration induced by the supernatants was also examined. RESULTS: Eosinophil adhesion to HPMEC was significantly augmented by the supernatants of Df-stimulated PBMC, which was significantly inhibited by anti-GM-CSF, but not by anti-IL-5, anti-RANTES, or isotype-matched controls. On the other hand, eosinophil migration induced by the supernatants was inhibited by anti-GM-CSF and partly by anti-RANTES. CONCLUSION: Both eosinophil adhesion and migration induced by the antigen-stimulated PBMC involve GM-CSF. In contrast, RANTES is involved only in the eosinophil migration. These molecules may participate in the development of eosinophil accumulation at the allergic inflammation sites. PMID- 10867502 TI - Systemic T helper 2 cell activation is not sufficient for antigen-induced eosinophil recruitment into the airways. AB - BACKGROUND: Airway eosinophilic inflammation is a characteristic feature of asthma. We have previously shown that antigen-induced eosinophil recruitment into the airways of sensitized BALB/c mice is mediated by CD4+ T cells and IL-5. However, the basis for the eosinophil recruitment into the airway are still largely unknown. METHODS: To determine the regulatory mechanisms that control the magnitude of antigen-induced eosinophilic inflammation in the airways, we analyzed antigen-induced eosinophil and T cell infiltration into the trachea in several strains of mice, including BALB/c (H-2(d)), BALB/b (H-2(b)), C57BL/6 (H 2(b)), C57BL/10 (H-2(b)), and B10.D2 (H-2(d)) mice. In addition, cytokine production from antigen-stimulated splenocytes and the titer of antigen-specific IgE antibody in serum were assessed. RESULTS: Antigen-induced eosinophil recruitment into the trachea in BALB/c mice was more than 20 times as abundant as that in C57BL/6 mice, the latter of which was also mediated by CD4+ T cells. In contrast, systemic Th2 responses, which were evaluated by the titers of antigen specific IgE antibody in serum and antigen-induced IL-4 and IL-5 production from splenocytes, were similarly observed in BALB/c mice and C57BL/6 mice. In addition, the antigen-induced eosinophil recruitment into the trachea was high in BALB/b mice, like BALB/c mice, but low in B10.D2 mice. CONCLUSIONS: Systemic Th2 responses are not sufficient for causing antigen-induced eosinophil recruitment into the airways and an undefined determinant(s) that exists in the BALB background mice rather than the H-2 haplotype is vital for the regulation of antigen-induced eosinophil recruitment into the airways. PMID- 10867503 TI - Role of secretory IgA, secretory component, and eosinophils in mucosal inflammation. AB - Eosinophils and their products are important in the pathophysiology of allergic inflammation in mucosal tissues. Secretory component (SC) bound to IgA mediates transepithelial transport of IgA. As another biological activity of SC, we have reported that secretory IgA (sIgA) and SC preferentially activate human eosinophils. When eosinophils were stimulated with immobilized sIgA, degranulation and superoxide production were greater than when stimulated with serum IgA. In contrast, neutrophils responded similarly to sIgA and serum IgA. Superoxide production by eosinophils stimulated with cytokines was enhanced synergistically by immobilized SC, while SC showed no effect on neutrophil activation. Eosinophil superoxide production stimulated with sIgA was abolished by anti-CD18 mAb, suggesting that beta2 integrins might be crucial for this reaction. There are several reports that SC and sIgA may play important roles in regulating eosinophil functions in vivo in diseases associated with mucosal eosinophilia and in various allergic diseases. It is speculated that eosinophils in the mucosa are activated by SC or sIgA, and that subsequent degranulation and superoxide production are induced. PMID- 10867504 TI - Regulation of CD69 expression on eosinophil precursors by interferon-gamma. AB - The purpose of this study was to determine whether interferon-gamma (IFN-gamma) induced CD69 expression by eosinophil precursors. Eosinophil precursors were induced from CD34+ cord blood cells using recombinant human interleukin-3 (IL-3) and interleukin-5 (IL-5). On day 14 of culture, cells constitutively expressed CD69 and the IFN-gamma receptor (IFN-gammaR). Stimulation with IFN-gamma for 24 h did not affect IFN-gammaR expression by the cells. On the other hand, IFN-gamma significantly upregulated CD69 expression by the precursors after 24 h of incubation. A specific JAK2 inhibitor (AG-490) caused a concentration-dependent suppression of IFN-gamma-induced CD69 expression by the precursors. In conclusion, these results indicate that IFN-gamma induces CD69 expression by eosinophil precursors via the activation of JAK2. PMID- 10867505 TI - Stimulation of the beta(2) integrin, alpha(M)beta(2), triggers tyrosine phosphorylation and cellular degranulation on human eosinophils. AB - Activation of human eosinophils by specific extracellular stimuli triggers the cellular degranulation response. Because cellular adhesion is critical for this eosinophil degranulation, we have tested the hypothesis that ligation of the beta(2) integrin, alpha(M)beta(2) (Mac-1, CD11b/CD18), leads to intracellular signaling events that contribute to the eosinophil activation response. Recently, we found that engagement of beta(2) integrin using two different approaches, such as cell adhesion induced by IL-5 or direct ligation of alpha(M)beta(2), triggered tyrosine phosphorylation of Cbl, the product of the c-cbl proto-oncogene, paxillin, a cytoskeletal protein, an unidentified 115-kD protein, and subsequent cellular degranulation. The results of this study indicate that engagement of alpha(M)beta(2) on eosinophils triggers an intracellular signaling cascade leading to cellular degranulation. Tyrosine phosphorylation of Cbl, paxillin, and a 115-kD protein may play important roles in adhesion-dependent cellular functions of eosinophils. PMID- 10867506 TI - Analysis of the prolongation of rat eosinophil survival induced by recombinant rat interleukin-5. AB - Rat eosinophil survival was prolonged by recombinant rat IL-5 prepared by the baculovirus expression system. The IL-5-induced prolongation of eosinophil survival was dose-dependently inhibited by the protein synthesis inhibitor cycloheximide, the DNA-dependent RNA synthesis inhibitor actinomycin D, and the tyrosine kinase inhibitor herbimycin A. The MEK-1 inhibitor PD98059 inhibited IL 5-induced phosphorylation of both p44 and p42 MAP kinases, but the IL-5-induced prolongation of eosinophil survival was not inhibited. In contrast, the JAK2 inhibitor AG490 inhibited the IL-5-induced prolongation of eosinophil survival. Treatment of eosinophils with IL-5 resulted in phosphorylation of STAT5 but not STAT1, and the IL-5-induced phosphorylation of STAT5 was inhibited by AG490. These findings suggest that recombinant rat IL-5 activates JAK2 tyrosine kinase, which phosphorylates STAT5, and induces protein synthesis required for the prolongation of rat eosinophil survival. PMID- 10867507 TI - Induction of apoptosis in bronchial eosinophils: beneficial or harmful? AB - BACKGROUND: Prominent eosinophil infiltration takes place in asthmatic bronchi, and damages bronchial epithelial cells. AIM: This study was designed to investigate whether induction of apoptosis in infiltrated cells in the airways is beneficial or harmful. METHODS: A/J mice, which are genetically predisposed to be hyperresponsive to acetylcholine, were immunized with ovalbumin (OA) and alum. Thereafter, they were subjected to a 2-week regimen of OA inhalation, during which they were also administered either hamster anti-mouse Fas monoclonal antibody or hamster IgG (sham control) intranasally. Pulmonary function was then analyzed using whole-body plethysmography. RESULTS: Inhalation of OA increased both airway responsiveness to acetylcholine and infiltration of eosinophils. Administration of anti-Fas antibody induced apoptosis in the infiltrating eosinophils and abolished the increase in airway responsiveness to acetylcholine. CONCLUSION: Induction of apoptosis in eosinophils infiltrating asthmatic bronchi has a beneficial effect on airway hyperresponsiveness. PMID- 10867508 TI - Expression of eotaxin by normal airway epithelial cells after influenza virus A infection. AB - BACKGROUND: Viral infection is known to cause lung inflammatory disease, including bronchial asthma. The mechanisms of inflammatory cell accumulation into the airways after viral infection are not well understood. Eotaxin is a CC chemokine which is a potent and specific agonist for CC chemokine receptor 3 (CCR3). CCR3 is expressed on eosinophils, basophils and T lymphocytes. These cells are known to be key cells in the pathogenesis of asthma. Although it has recently been demonstrated that airway epithelial cells express eotaxin in vivo and in vitro, there are few data about its epxression in viral infection. We hypothesized that eotaxin may play an important role in attracting inflammatory cells to the airways after viral infection, and analyzed whether viral infection attracts eotaxin in bronchial epithelial cells in vitro. METHODS: Human airway epithelial cells obtained from bronchial tissue at lobectomy for lung cancer were infected with influenza virus A (subtype H3N2). The cells and cultured media were collected 8, 24, and 48 h after infection. Eotaxin mRNA was analyzed with reverse transcriptase-polymerase chain reaction. Eotaxin protein levels in the culture media were analyzed by enzyme-linked immunosorbent assay. We also studied a blocking assay to analyze the intervention of proinflammatory cytokines in its production induced by influenza virus. RESULTS: Eotaxin mRNA appeared to be expressed constitutively in uninfected cells but was expressed more clearly in infected cells. Eotaxin protein release into culture media significantly increased after infection. Anti-TNF-alpha and anti-IL-1beta antibodies did not alter the eotaxin protein levels after viral infection. CONCLUSIONS: These results suggest that influenza virus A infection in airway epithelial cells activates the expression of eotaxin and that eotaxin may participate in the pathogenesis of airway inflammatory disease caused by viral infection, such as infectious type asthma. PMID- 10867509 TI - Autocrine regulation of eotaxin in normal human bronchial epithelial cells. AB - Normal human bronchial epithelial cells (NHBECs) spontaneously synthesize and store chemokines including eotaxin, RANTES, or MCP-3. We also observed the constitutive expression of mRNA specific for CC chemokine receptor-3 (CCR3) in these cells. The stimulation of these cells by eotaxin (1-100 ng/ml) dose dependently increased eotaxin production. In addition, RANTES or eotaxin-2 but not MCP-3 at the higher concentrations (100 ng/ml), also induced slight eotaxin production. Eotaxin elicited the expression of eotaxin mRNA at 6 h after stimulation. Antibody against eotaxin or CCR3, and actinomycin D inhibited these effects of eotaxin. These antibodies also significantly inhibited the ability of eotaxin to produce further eotaxin. These findings strongly suggest that eotaxin which originates from inflammatory cells or cells resident in the airway of bronchial asthmatics can induce further production of eotaxin and participate in expansion of allergic inflammation. PMID- 10867511 TI - An optimal condition of bronchial cell proliferation stimulated by insulin-like growth factor-I. AB - BACKGROUND: It is known that growth hormones such as insulin-like growth factor-I (IGF-I) and several kinds of cytokines are involved in the regeneration process of injured epithelial cells in chronic respiratory inflammatory diseases such as bronchial asthma. Repetitive degeneration/regeneration processes of the airway epithelial layer is supposed to be responsible for the remodeling and irreversible organic changes of the airway in bronchial asthma. The purpose of this study is to establish a simple and reliable in vitro method for studying airway epithelial cell growth and proliferation using IGF-I. METHOD: By altering the number of cultured epithelial cells (strain NCI-H(292)), culture duration before stimulation with IGF-I, concentration of IGF-I, and duration of IGF-I stimulation, the optimum conditions for epithelial cell growth was determined. The epithelial cell growth was evaluated using [methyl-(3)H]thymidine uptake. RESULT: Among various culture conditions, the epithelial cells cultured at 1 x 10(3) cells/well for 24 h followed by 24 h of stimulation by 10(-8) M of IGF-I showed the highest growth. CONCLUSION: The method for the evaluation of epithelial cell growth established in this study requires a small number of cells and has no complicated procedure. This simple model enables us to investigate the effect of various substances on bronchial epithelial cell growth in the presence of IGF-I. PMID- 10867510 TI - Regulation of human eotaxin generation by Th1-/Th2-derived cytokines. AB - BACKGROUND: Accumulating evidence indicates that eotaxin is the primary mediator of tissue eosinophilia. In the present study, we analyzed the mechanisms of eotaxin generation by Th1-/Th2-derived cytokines in vitro. METHODS: Human dermal fibroblasts, human umbilical vein endothelial cells and A549 human bronchial epithelial cell line cells were stimulated with TNF-alpha, IL-4, IFN-gamma or TNF alpha in combination with IL-4 or IFN-gamma and the amount of eotaxin production was analyzed. RESULTS: Fibroblasts produced nanogram/milliliter quantities of eotaxin. Proinflammatory cytokine TNF-alpha and Th2-type cytokine IL-4 each induced eotaxin production, and combination of them caused a marked synergistic increase in that production. On the other hand, Th1-type cytokine IFN-gamma inhibited eotaxin generation at the protein/mRNA levels. CONCLUSION: The Th2 derived cytokine upregulated while the Th1-derived cytokine inhibited eotaxin production by fibroblasts. In view of the Th1/Th2 paradigm, these results indicate that (1) eotaxin regulates eosinophil accumulation in the Th2-dominant state such as allergic disease, and (2) direct suppression of eotaxin production by IFN-gamma is one of the major mechanisms by which IFN-gamma suppresses eosinophilic inflammation. PMID- 10867512 TI - IL-5 synthesis by T cells of allergic subjects is regulated at the transcriptional level. AB - BACKGROUND: IL-5 plays a central role in allergic diseases associated with eosinophilic inflammation. We have previously reported that IL-5 production by peripheral blood mononuclear cells (PBMC) is greatly enhanced in both atopic and nonatopic asthmatics compared to control subjects. METHOD: Concanavalin A (Con A) blast lymphocytes were derived from PBMC of allergic and control subjects. Transcriptional regulation of the IL-5 gene was investigated by transient transfection assay. RESULTS: A significant amount of IL-5 was produced by Con A blast lymphocytes derived from allergic subjects upon stimulation with phorbol ester and Ca(2+) ionophore, whereas the cells derived from control subjects did not produce a detectable amount of IL-5. Production of IL-2 and IL-4 was not significantly different between the two groups. A luciferase reporter plasmid containing the human IL-5 promoter/enhancer region was transcribed by Con A blast lymphocytes derived from allergic subjects, but not by the cells from control subjects, upon activation. CONCLUSION: IL-5 synthesis by nontransformed T cells of allergic subjects is enhanced at the level of gene transcription. Elucidation of the molecular mechanism of IL-5 gene transcription by allergic T cells may delineate the pathogenesis of allergic disease for the future therapeutic intervention. PMID- 10867513 TI - Tamm-Horsfall protein updated. PMID- 10867514 TI - Oxidative processes in glomerulonephritides: additional facts. PMID- 10867515 TI - Fas-Fas ligand system in the peripheral blood of patients with renal diseases. AB - To clarify the role of the Fas-Fas ligand (FasL) system in the peripheral blood from patients with various renal diseases, the Fas and FasL expression on mononuclear cells (MNCs) and serum levels of soluble Fas (sFas) and soluble FasL were investigated. Patients were selected from those with various types of glomerular diseases showing various degrees of renal function. Fas expression on MNCs was analyzed by a FACScan, sFas and soluble FasL were measured with an ELISA kit, and FasL expression on MNCs was counted using a FACScan after a bioassay. Fas-positive MNCs and sFas increased with statistical significance concomitantly with deterioration in renal function. Moreover, there was a significant correlation between them. sFas- and FasL-positive MNCs were significantly correlated with proteinuria. However, the Fas expression percentage on MNCs and/or serum levels of sFas did not correlate with the number of TUNEL-positive cells in the glomeruli. Also, there was no disease specificity in the activation of Fas. These results indicate that Fas expression on MNCs is activated in accordance with the deterioration in renal function without disease specificity, corresponding to the elevation of serum sFas levels to protect against Fas mediated apoptosis. PMID- 10867516 TI - Responses of the skin microcirculation to acetylcholine in patients with essential hypertension and in normotensive patients with chronic renal failure. AB - AIMS: To assess the endothelial function of the skin microcirculation in chronic renal failure (CRF) independent of hypertension, we investigated the changes of the cutaneous blood flow induced by iontophoretic delivery of acetylcholine (ACh) and of sodium nitroprusside (SNP) in CRF patients free from arterial hypertension and in patients with essential hypertension. METHODS: The study included 20 patients affected by CRF (mean creatinine clearance 12+/-2 ml/min) without arterial hypertension (mean blood pressure 96+/-1 mm Hg), 15 patients affected by essential hypertension (mean blood pressure 124 +/-1 mm Hg), and 20 normal controls. The changes of skin blood flow following iontophoretic delivery of ACh and of SNP were measured by laser Doppler flowmetry. RESULTS: Following maximal ACh or SNP delivery, the change of blood flow from the baseline was similar both in normals (683+/-92 vs. 684 +/- 87%) and in CRF patients (778+/-108 vs. 803+/ 124%), whereas in the hypertensives the response to ACh was lower than to SNP (434+/-48 vs. 702 +/- 98%, p<0.01). Since the third ACh delivery dose, the skin blood flow increments were significantly lower in the hypertensive than in the CRF or in the normal control groups, whereas no difference was observed between uremics and controls. CONCLUSIONS: The endothelium-dependent hyperemia following ACh iontophoretic delivery is impaired in the skin microcirculation of essential hypertensive patients, but this is not the case in CRF patients with no history of arterial hypertension. This suggests that CRF per se, independent of arterial hypertension, is not associated with endothelial dysfunction of skin microcirculation. PMID- 10867517 TI - Urinary excretion of glutathione S transferases alpha and pi in patients with proteinuria: reflection of the site of tubular injury. AB - In patients with renal diseases, proteinuria is a major determinant of progressive renal failure, probably by causing tubular cell injury. Little is known on extent and site of tubular cell injury in patients with proteinuria. Glutathione S transferases (GST) are cytosolic enzymes. The alpha isoform is present only in proximal tubular cells, whereas the pi isoform is confined to distal tubular cells. We have studied the urinary excretion of both isoenzymes in 56 (38 male and 18 female) patients with glomerular diseases and proteinuria. The mean age was 45 +/- (SD) 16 years, the median creatinine clearance was 80 (range 27-159) ml/min, and the median albuminuria was 4.2 (range 0.7-16.9) g/10 mmol creatinine. The excretions of both GST alpha (median 35.9 ng/10 mmol creatinine) and GST pi (median 24.8 ng/10 mmol creatinine) were elevated as compared with control values (upper limits 10 and 12 ng/10 mmol creatinine, respectively). The urinary excretion of GST pi, but not that of GST alpha, was inversely correlated with the creatinine clearance. The highest levels of GST alpha were found in patients with a well-preserved renal function, whereas highest levels of GST pi were found in patients with renal failure. In a small number of patients we performed immunofluorescent studies of renal tissue. An increased urinary excretion of GST alpha correlated with brush border damage and decreased staining of proximal tubules for that isoenzyme. Our data suggest that in patients with proteinuria initial injury is apparent at the proximal tubules. Measurements of GST alpha and GST pi appear useful to study longitudinal timing and site of proteinuria-induced tubular cell injury. PMID- 10867518 TI - Interleukin-18 and interleukin-12 synergize to stimulate the production of vascular permeability factor by T lymphocytes in normal subjects and in patients with minimal-change nephrotic syndrome. AB - BACKGROUND/AIM: In a previous study, we reported that interleukin (IL)-12 could upregulate the production of vascular permeability factor (VPF) derived from activated human peripheral blood mononuclear cells. Since IL-18, a novel immunoregulatory cytokine with potent interferon-gamma inducing activities, has been shown to be a strong cofactor for T helper type 1 cell development, we tested the hypothesis that IL-18 in combination with IL-12 can act synergistically to modulate the production of VPF. METHODS: For this purpose, T cells were isolated from heparinized venous blood, stimulated with concanavalin A, and incubated in the presence of IL-18 or IL-12, and the production of VPF was determined by the method of Lagrue. RESULTS: There was a significant increase in VPF production from concanavalin A-stimulated T cells following incubation with IL-18 or IL-12. More importantly, the combination of the cytokines was found to give a potent synergistic stimulation of VPF by concanavalin A-activated T cells from normal subjects. To determine the specificity of the stimulatory effect, neutralizing anti-IL-18 and anti-IL-12 antibodies were preincubated with IL- 18 and IL-12, respectively, prior to the addition of responder cells. The antibodies completely inhibited the effects of IL-18 and IL-12. Thus, these data show that IL-18 can synergize with IL-12 to selectively increase the production of VPF from T cells. The present study further demonstrates that IL-18 and IL-12 are in fact acting in synergy in patients with minimal-change nephrotic syndrome. CONCLUSIONS: Taken together, our results indicate that both IL-18 and IL-12 contribute to the VPF production in vitro and suggest that they play key roles in the complexity of cytokine regulation in the pathophysiology of VPF. PMID- 10867519 TI - Sensitivity and specificity of the thermodilution technique in detection of access recirculation. AB - BACKGROUND/AIM: Recirculation measured by thermodilution includes effects caused by access and cardiopulmonary recirculation. The aims of this study were to illustrate the accuracy of thermodilution in measurement of hemodialysis recirculation and also to identify a sensitive and specific threshold to detect access recirculation. METHODS: 110 studies were performed in 19 patients. Recirculation obtained directly by the blood temperature monitor (BTM) was compared to that calculated from access blood flow, pump blood flow, and cardiac output determined by ultrasound dilution using the hemodialysis monitor (HDM). RESULTS: A highly significant linear correlation was obtained between repeated BTM recirculation measurements (R(BTM, 2) = 0.99.R(BTM, 1) - 0.22%, r(2) = 0.99). There were no significant differences between repeated BTM recirculation measurements with correct placement (11.4+/-7.1 vs. 10.9+/-7.4%, p = NS) or reversed placement (30.0+/-15.6 vs. 30.2 +/-15.9%, p = NS) of blood lines. A strong linear relationship was obtained between the recirculation determined by thermodilution and the recirculation calculated from HDM measurements (R(calc) = 0.98. R(BTM) - 1.49%, r(2) = 0.95). The mean recirculation obtained by BTM was not significantly different from the recirculation calculated by HDM with correct placement (9.5+/- 2.2 vs. 8.6+/-2.5%, p = NS) or with reversed placement (25.4+/ 7.8 vs. 23.8+/-7.7%, p = NS) of blood lines. When a recirculation greater than 15% measured by the BTM was considered as the threshold at which true access recirculation occurred, sensitivity and specificity of the thermodilution method to detect access recirculation were 93 and 98%, respectively. CONCLUSIONS: Recirculation measurements made by the BTM are accurate and precise. Even though BTM thermodilution includes effects of cardiopulmonary recirculation, so that low levels of access recirculation might not be detected, a BTM recirculation >15% represents a highly significant access recirculation. PMID- 10867520 TI - Differences between blood flow as indicated by the hemodialysis blood roller pump and blood flow measured by an ultrasonic sensor. AB - BACKGROUND/AIM: The ultrasonic transit time is currently the best method for measuring the blood flow rate in the extracorporeal hemodialysis circuit. The purpose of this study was to analyze the differences between blood flow as indicated by the hemodialysis blood roller pump (prescribed blood flow) and by an ultrasonic flowmeter (delivered blood flow). METHODS: The ultrasonic blood flow was measured in 20 patients on chronic hemodialysis who were dialyzed through an arteriovenous fistula. During each dialysis session the ultrasonic blood flow was measured at three different blood roller pump flow rates (300, 350, and 400 ml/min). In order to analyze the influence of inflow and outflow pressures on blood flow, this study was conducted during nine consecutive dialysis sessions during which needles of different sizes were used. RESULTS: The ultrasonic flow was always lower than indicated by the blood roller pump: 265+/-12, 304+/-15, and 341+/- 19 ml/min for blood roller pump flow rates of 300, 350, and 400 ml/min, respectively (variability: -11.6, -13.1, and -14.8%, respectively). An univariate regression analysis showed that the reduction in flow recorded ultrasonically correlated with both venous blood line pressure (r = -0.2679, p<0.001) and negative arterial blood line pressure (r = 0. 6773, p<0.001). By multivariate analysis, only the arterial blood line pressure has a predictive value. When all measurements were grouped by arterial blood line pressure ranges, the variability between ultrasonic blood flow and blood roller pump flow was found to be similar in those groups with the same arterial blood line pressure, regardless of the blood roller pump flow rate. CONCLUSIONS: The blood flow indicated by the dialysis blood roller pump is always greater than the delivered blood flow, and this difference is in turn conditioned by the negative pressure induced by the blood roller pump in the arterial blood line. PMID- 10867521 TI - Expression of heat shock proteins 72/73 in human peritoneal mesothelial cells in vivo and in vitro. AB - Leukocyte accumulation during peritonitis leads to an injurious microenvironment which is involved in the host defense reaction but is also thought to cause peritoneal damage. We tested the hypothesis that mesothelial cells (MC) respond to the injurious microenvironment during peritonitis by an increased expression of heat shock proteins (HSP 72/73), a basic way by which cells are protected against injury. Comparison of resting MC and activated MC during peritonitis in vivo by means of immunohistochemistry revealed an increased expression of HSP 72/73. As assessed by Western immunoblotting, incubation of MC in vitro with tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) caused a time-dependent induction of HSP 72/73 expression, which was maximal 6 h after stimulation. We suggest that the increased HSP 72/73 expression of MC during peritonitis is in part induced by TNF-alpha and IL-1beta and may exert a cell protective function, lessening MC damage during peritonitis. PMID- 10867522 TI - Expression of adhesion molecules in kidney with experimental chronic obstructive uropathy: the pathogenic role of ICAM-1 and VCAM-1. AB - BACKGROUND: Chronic obstructive uropathy induced by maintained unilateral ureter ligation in the rat is characterized morphologically by interstitial inflammation, interstitial fibrosis, and tubular atrophy. Infiltrating mononuclear inflammatory cells, particularly T lymphocytes and macrophages, may contribute to the progression of this lesion by mediating tubular injury and by the activation of interstitial fibroblasts, with resultant tubular atrophy and interstitial fibrosis, respectively. Altered expression and activation of adhesion molecules by leukocytes, vascular endothelial cells, and parenchymal cells likely contributes both to the infiltration of inflammatory cells into the tubulointerstitial compartment and to the interaction of activated inflammatory cells with parenchymal cells. METHODS: In the current study, we examined changes in the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in a 90-day model of maintained unilateral ureter ligation in male Sprague-Dawley rats. RESULTS: Rat kidneys showed constitutive expression of ICAM-1 mRNA and constitutive immunostaining for ICAM-1 in peritubular capillaries, glomeruli, and a small percentage of cortical tubules. Ureter ligation resulted in a rapid increase in ICAM-1 mRNA, which was almost 2 fold greater than those of the contralateral and control kidneys as early as 3 h and which was maintained at a 4- to 6-fold higher level in the ligated vs. contralateral kidneys throughout the entire 90-day time course. There was a marked increase in ICAM-1 immunostaining within the tubular epithelium, with up to 80% of both cortical and medullary tubular cross-sections showing strong apical immunostaining from day 6 to 25, with a subsequent decrease throughout the remainder of the experiment. ICAM-1 immunostaining in the expanding interstitium in the ligated kidneys showed a gradual increase throughout the duration of the experiment. In contrast, glomerular immunostaining for ICAM-1 was decreased in the ligated compared to the contralateral kidneys throughout the entire experiment. There was a later but prominent increase in VCAM-1 mRNA in ligated kidneys, which was first evident at 2 days and which was maintained 2- to 10-fold greater than the contralateral kidneys throughout the entire time course. VCAM-1 immunostaining increased in the expanding interstitium, but decreased in glomeruli in obstructed vs. contralateral kidneys. Tubular staining for VCAM-1 did not change after ureter ligation. CONCLUSION: Increased ICAM-1 and VCAM-1 may contribute to the prominent inflammatory cell infiltration in the chronic tubulointerstitial nephritis accompanying maintained unilateral ligation. Tubule expression of ICAM-1, which occurs during a similar time course as previously documented for tubular cell proliferation and especially tubular cell apoptosis in this model, may contribute to injurious interactions of activated inflammatory cells with tubular epithelium. PMID- 10867523 TI - Melatonin, a pineal hormone with antioxidant property, protects against gentamicin-induced nephrotoxicity in rats. AB - The present study investigated the effects of melatonin, an antioxidant, on gentamicin-induced nephrotoxicity in rats. Melatonin (5 mg/kg p.o.) was used 3 days before and 8 days simultaneously with gentamicin (80 mg/kg i.p.) Saline treated animals served as controls. Determinations of urinary creatinine, N acetyl-beta-D-glucosaminidase, glucose, protein, blood urea, serum creatinine, plasma and kidney tissue malondialdehyde (MDA), and antioxidant enzyme levels in kidney tissue were done after 8 days of gentamicin treatment. The kidneys were also examined for morphological changes using histological techniques. Gentamicin caused nephrotoxicity as evidenced by marked elevation in blood urea and serum creatinine. Mean blood urea and serum creatinine levels were 289+/-50, and 2.5+/ 0.5 mg/dl, respectively, in rats treated with gentamicin. Melatonin significantly protected the rats from gentamicin-induced nephrotoxicity; blood urea and serum creatinine levels were 23+/-2.7 and 0.88+/-0.19 mg/dl, respectively. The creatinine clearance was decreased with gentamicin treatment (0.048+/- 0.007 ml/min) as compared with controls (0.41+/-0.08 ml/h/kg). In rats treated with melatonin plus gentamicin, the creatinine clearance was similar to controls (0.41+/-0.08 ml/h/kg). The product of lipid peroxidation (MDA) was markedly increased in plasma (2.10+/-0.15 nmol) and kidney tissue (8.87+/-3.2 nmol/mg protein) with gentamicin treatment. Melatonin prevented the gentamicin-induced rise in plasma MDA (1.03+/-0.27 nmol) and kidney tissue MDA (2.57+/-0.87 nmol/mg protein). An increased excretion of urinary N-acetyl-beta-D-glucosaminidase, glucose, and protein by gentamicin was also prevented by melatonin. Kidneys from gentamicin-treated rats showed tubular epithelial loss with intense granular degeneration involving more than 50% of renal cortex, while there were findings comparable to controls in melatonin plus gentamicin treated rats. The present study indicates that melatonin significantly protects against gentamicin-induced renal toxicity in Wistar rats. PMID- 10867524 TI - Chronic unilateral ureteral obstruction represented as renin-dependent hypertension. AB - A 50-year-old woman developed renin-dependent hypertension immediately after accidental unilateral ureteral ligation during hysterectomy, and the hypertension lasted for 5 months. Surgical release of the obstruction was carried out 157 days after the ligation. Then, her blood pressure was normalized. However, the obstructed kidney showed intensive tubulointerstitial fibrosis and functional recovery was not obtained. This case suggests that the renin-angiotensin system may be upregulated in human kidney during unilateral ureteral obstruction for a long duration. PMID- 10867525 TI - HIV-Associated nephropathy: is it going to disappear? PMID- 10867526 TI - Hepatitis C virus RNA as an early indicator of hepatitis C virus associated glomerulonephritis. PMID- 10867527 TI - Peritubular capillary flow and tubular function in idiopathic nephrotic syndrome. PMID- 10867528 TI - Renoprotective properties of angiotensin-converting enzyme inhibitors: proofs and limits? PMID- 10867529 TI - The unmasking of hyperreninemic hypovolemia by captopril test in a hypertensive HD patient unaccompanied by autonomic neuropathy. PMID- 10867530 TI - Urinary podocytes in patients with chronic renal failure. PMID- 10867531 TI - Successfully treated renal artery stenosis using an oral PGI(2) derivative, beraprost, in cadaveric kidney transplantation. PMID- 10867532 TI - Effect of acarbose on blood glucose and proteinuria in patients with diabetic nephropathy. PMID- 10867533 TI - Vascular hyperpermeability in nephrotic edema. AB - BACKGROUND/AIM: There is increasing evidence that hypoalbuminemia and the inability of the renal distal tubule to excrete salt are not the only factors responsible for nephrotic edema. We tested the possibility that vascular hyperpermeability also plays a role in the pathophysiology of nephrotic edema in human primary glomerulonephritis. METHODS: We investigated the capillary permeability by means of a standardized test using albumin labelled with technetium (99mTc-albumin) in 20 healthy adults and in 101 nephrotic adult patients comprising 60 patients with idiopathic nephrotic syndrome (INS; minimal change nephropathy and segmental glomerulosclerosis), 32 with idiopathic membranous nephropathy (IMN) and 9 patients with idiopathic type I membranoproliferative glomerulonephritis (MPGN). The patients and healthy controls were also compared with a group of women (n = 25) with idiopathic cyclic edema, a disease in which an increase in capillary permeability plays a pivotal role. The capillary permeability measured by the Landis isotope test is normal in edematous patients with cardiac and renal impairment unrelated to glomerular disease, cirrhosis, hypothyroidism, lymphatic obstruction and diuretic abuse. As values were not normally distributed, nonparametric analysis of variance was used (Kruskal-Wallis test), and patient groups were compared with healthy controls and with women with idiopathic cyclic edema by means of the two-tailed nonparametric Mann-Whitney test. The effects of high-dose steroids and Ginkgo biloba extract (Tanakan); an agent able to improve capillary permeability) were analyzed by means of the two-tailed nonparametric Wilcoxon test. RESULTS: The capillary permeability was significantly increased (Mann-Whitney test) in each glomerular disease group compared with the healthy controls. 99mTc-albumin extravasation values (%; median and range in parentheses were the following: healthy controls 0 (0-8.4); idiopathic cyclic edema patients 11.5 (8-24), p < 0.001; INS 20 (0-50), p < 0.0001; IMN 12.5 (0-40), p < 0.001, and MPGN 10 (0-40), p < 0.05. An increase in capillary permeability exceeding the upper limit of control values (>8% of 99mTc-albumin extravasation) was observed in 88% of INS, in 53% of IMN and in 44% of MPGN patients. The increase in capillary permeability (%) was greater in idiopathic nephrotic patients than in idiopathic cyclic edema patients (p < 0. 005, Mann-Whitney test) and was markedly reduced in nephrotic patients receiving high-dose steroids (n = 8) [before 25 (8-40); after 0 (0-25), p < 0.005 (Wilcoxon's test)] and high doses of G. biloba extract (n = 16) [before 30 (8 50); after 2.5 (0-20, p < 0. 0005 (Wilcoxon's test)]. CONCLUSIONS: We conclude that capillary permeability is severely altered in most of the nephrotic patients with primary glomerulonephritis. These results strongly suggest that the capillary hyperpermeability plays a role in the pathophysiology of nephrotic edema in primary glomerular disease, especially in INS. We postulate that this widespread abnormality in capillary permeability is related to the release of vascular permeability factor and other cytokines by immune cells. PMID- 10867534 TI - Adrenomedullin gene transcription is decreased in peripheral blood mononuclear cells of patients with IgA nephropathy. AB - We measured mRNA levels of adrenomedullin (AM), C-type natriuretic peptide (CNP), vascular endothelial growth factor (VEGF), interleukin 1beta (IL-1beta) and interleukin 6 (IL-6) in peripheral blood mononuclear cells (PBMC) of patients with IgA nephropathy. To evaluate these mRNA levels, we employed a real-time quantitative PCR method which was performed using a hybridization probe labeled with two fluorescence dyes. This strategy was found to afford the standard curves with a high correlation, suggesting that this method is useful for evaluations of mRNA levels. By this method, levels of AM, CNP, VEGF, IL-1beta and IL-6 mRNA in PBMC of 49 IgA nephropathy patients and 35 healthy volunteers were evaluated. Among the mRNAs examined, AM mRNA levels were significantly lower in severe-grade than in mild-grade IgA nephropathy patients. Furthermore, AM mRNA levels correlated with CNP mRNA levels in PBMC of patients with IgA nephropathy, and each peptide generated from these mRNAs has antiproliferative effects on mesangial cells. These data indicate that gene expression of AM in PBMC is regulated according to the pathophysiological states of IgA nephropathy and that decreased AM production may contribute to the progression of IgA nephropathy. PMID- 10867535 TI - Increased excretions of beta2-microglobulin, IL-6, and IL-8 and decreased excretion of Tamm-Horsfall glycoprotein in urine of patients with active lupus nephritis. AB - Tubulointerstitial nephritis is a less frequently recognized but important complication of systemic lupus erythematosus. We have investigated the cytokine beta2-microglobulin (beta2M) and Tamm-Horsfall glycoprotein (THG) excretions in the urine of systemic lupus erythematosus patients to identify indices for evaluation of tubulointerstitial inflammation in lupus nephritis (LN). Daily urine was collected from 15 patients with active LN, from 12 patients with inactive LN, and from 17 normal subjects. The amounts of soluble interleukin (IL) 2 receptor, IL-6, IL-8, beta2M, and THG in urine were measured. Beta2M and THG were regarded as indicators of proximal and distal renal tubule function, respectively. The urinary excretions of IL-6 and IL-8 were significantly higher in patients with active LN than in those with inactive LN and in normal individuals. The excretion of soluble IL-2 receptor in all three groups of subjects was not significantly different. On the other hand, the excretion of beta2M in patients with LN was significantly higher than that in normal individuals. The excretion of beta2M in patients with active or inactive LN was not significantly different. The THG excretion was lower in patients with active LN and tubulointerstitial inflammation as compared with patients with inactive LN or normal individuals. Six patients underwent pulse cyclophosphamide therapy during the course of experiments. Five of them showed a decrease in IL-8 and IL-6 excretions in urine after the treatment. The excretions of beta2M and THG in urine, in addition to IL-6 and IL-8, can reflect the renal inflammatory activity in patients with lupus tubulointerstitial nephritis as well as in those having lupus glomerulonephritis. PMID- 10867536 TI - A controlled trial of the effect of folate supplements on homocysteine, lipids and hemorheology in end-stage renal disease. AB - Elevated plasma homocysteine (Hcy), dyslipidemia and hemorheological abnormalities all occur commonly in end-stage renal disease (ESRD) and are recognized risk factors for arteriosclerosis. To study the effect of folate supplementation on these factors we conducted a randomized controlled trial. Thirteen hemodialysis (HD) and 8 continuous ambulatory peritoneal dialysis (CAPD) patients received either 5 mg folic acid daily or placebo for 3 months. After 1 and 3 months, fasting blood samples were taken for Hcy, lipid profile, blood and plasma viscosity, red blood cell (RBC) osmotic fragility, plasma fibrinogen concentration and in vivo platelet aggregability. At baseline, the CAPD patients had a higher mean plasma fibrinogen concentration than the HD patients and they also tended to have higher mean plasma viscosity. Folate-treated patients showed marked increases in RBC folate and an average decrease in plasma Hcy concentration of 33%. Mean total cholesterol, LDL cholesterol and triglyceride concentrations decreased significantly in the CAPD patients who took folate. Folate had no significant effect on hemorheology. In conclusion, folate supplements in ESRD reduce plasma Hcy concentrations and may improve lipid profiles. In our patients, hemorheological abnormalities were more marked in patients on CAPD than in those on HD and were not improved by folate supplementation. PMID- 10867537 TI - Reduction of neutrophil activation by vitamin E modified dialyzer membranes. AB - BACKGROUND/AIM: Transient leukopenia during hemodialysis due to neutrophil activation is attributed to bioincompatibility of the dialysis membrane, but the mechanism remains unclear. We studied the mechanism of neutrophilic activation by comparing a vitamin E modified membrane (CLEE) and a regular cellulose membrane (CLSS). METHODS: (1) CLSS and CLEE membranes were used in a crossover clinical study in 7 chronic hemodialysis patients. Neutropenia, CD11b expression, and plasma C3a and myeloperoxidase concentrations were compared between the two dialyzer membranes. (2) Normal blood was circulated through CLEE and CLSS minimodels, and the same parameters were compared. (3) Blood samples with modified complement activities (EDTA: both classical and alternative pathways inactivated; EGTA+Mg: classical pathway inactivated; heating: alternative pathway inactivated; control: no modification) were incubated in the CLSS minimodel, and the neutrophilic activation was compared. RESULTS: In clinical hemodialysis, neutropenia, CD11b expression, and C3a and myeloperoxidase levels were significantly lower when CLEE membranes were used. The same tendency was observed in minimodels. However, the degrees of inhibition in clinical dialysis, especially at the venous line, were significantly higher than in minimodels. As compared with controls, CD11b expression and myeloperoxidase level were significantly lower when both classical and alternative pathways were inactivated or when the classical pathway alone was inactivated, but were not significantly different when the alternative pathway alone was inactivated. CONCLUSIONS: Vitamin E modification of the dialyzer reduces some reactions of neutrophilic activation, such as CD11b expression and myeloperoxidase release, more effectively in the clinical situation than in ex vivo models, suggesting a possible effect of vitamin E in inhibiting bioreactions due to pyrogen in the dialysate. The classical complement pathway is required in membrane-induced neutrophilic activation, at least during the initial stage. PMID- 10867538 TI - An intron 4 gene polymorphism in endothelial cell nitric oxide synthase might modulate volume-dependent hypertension in patients on hemodialysis. AB - Nitric oxide (NO) has some relevance to the pathophysiology of hypertension. We examined the distribution of endothelial nitric oxide synthase (ecNOS4) gene polymorphism in patients with essential hypertension (n = 107) in comparison with healthy subjects (n = 362), and then studied the possibility that ecNOS4 gene polymorphism affects changes in blood pressure (BP) due to water load in hemodialysis patients. Though the NO metabolite (NOx) level in subjects with the a allele (31.2 +/- 1.76 micromol/l) was significantly lower than in those without the a allele (35.6 +/- 0.90 micromol/l) (p = 0.0263), we found no association between this gene polymorphism and essential hypertension. However, there still remains the possibility that the NO response to elevation of BP might be suppressed in patients with hypertension. In order to examine the association between ecNOS4 gene polymorphism and volume-dependent hypertension, 181 hemodialysis patients with complete anuria were recruited. Depending on the increase in mean BP (mm Hg) divided by percent increase in body weight (deltaBP/deltaBW), patients were divided into high responders (High-R: deltaBP/deltaBW >2.0 mm Hg; n = 90) and low responders (Low-R: deltaBP/deltaBW <2.0 mm Hg; n = 91). In the High-R group, the frequencies of a/a, b/a and b/b genotypes were 1.1, 32.1 and 66.8%, respectively, and in the Low-R group, these frequencies were 1.1, 9.4 and 89.5%, respectively. The relative risk for those in the High-R group conferred by the ecNOS4 a allele (b/a + a/a) was 3.64 (95% confidence intervals: 1.60-8.24, p = 0.0089). This study did not show a strong involvement of ecNOS4 gene polymorphism, at least in the basal NO production in patients with essential hypertension, however, it indicated that ecNOS4 gene polymorphism might modulate changes in BP due to water load in patients on hemodialysis, thus indicating that these polymorphisms may be involved in the pathophysiology of volume-dependent hypertension. PMID- 10867539 TI - G proteins regulate calcium channels in the luminal membranes of the rabbit nephron. AB - The filtered calcium (Ca2+) is reabsorbed by the luminal membrane of the proximal and distal nephron. Ca2+ enters cells across apical plasma membranes along a steep electrochemical gradient, through Ca2+ channels. Regulation by various hormones implies several steps, including binding of these hormones to the basolateral membrane, interaction with G proteins, liberation of messengers, activation of kinases and finally opening of the channels at the opposite pole of the cells. In the present study, we examined whether the Ca2+ entry through the luminal membranes of proximal and distal tubules is also regulated by G proteins, by a membrane-limited process. Luminal membranes were purified from rabbit proximal and distal tubule suspensions, and their vesicles were loaded with GTPgammas or the carrier. Then, the 45Ca2+ uptake by these membrane vesicles was measured in the presence and absence of 100 mM NaCl. In the absence of Na+, intravesicular GTPgammas significantly enhanced 0.5 mM Ca2+ uptake by the proximal membrane vesicles from 0.53 +/- 0.06 to 0.72 +/- 0.06 pmol/microg/10 s (p < 0.05). In the presence of Na+, however, this effect disappeared. In the distal tubules, intravesicular GTPgammas increased 0.5 mM Ca2+ uptake in the absence (from 0.57 +/- 0.02 to 0.79 +/- 0.02 pmol/microg/10 s, p < 0.02) and in the presence (from 0.36 +/- 0.03 to 0.55 +/- 0.03 pmol/microg/10 s, p < 0.02) of Na+. The action of GTPgammas, when present, was dose dependent with a half maximal effect at 20 microM. The distal luminal membrane is the site of two Ca2+ channels with different kinetics parameters. GTPgammas increased the Vmax value of the low-affinity component exclusively, in the presence as in the absence of Na+. Finally, Ca2+ uptake by the membranes of the two segments was differently influenced by toxins: cholera toxin slightly stimulated transport by the proximal membrane, but had no influence on the distal membrane, whereas pertussis toxin decreased the cation uptake by the distal tubule membrane exclusively. We conclude that the nature of Ca2+ channels differs in the proximal and distal luminal membranes: Ca2+ channels present in the proximal tubule and the low affinity Ca2+ channels present in the distal tubule membranes are directly regulated by Gs and Gi proteins respectively, whereas the high-affinity Ca2+ channel in the distal tubule membrane is insensitive to any of them. PMID- 10867540 TI - Proliferative responses of mesangial cells to growth factors during compensatory versus dietary hypertrophy. AB - While the bulk of renal hypertrophy induced by contralateral nephrectomy or a high-protein diet consists of tubular cell growth, there is some evidence suggesting that mesangial cells play a role in this phenomenon. Previous data suggest that this role of mesangial cells is associated with their proliferation. We, therefore, undertook this investigation to assess the proliferative responses of mesangial cells, originating from single remaining kidneys or from kidneys of rats fed a high-protein diet, to epinephrine, endothelin, arginine vasopressin, neo-synephrine, or epidermal growth factor (EGF). All agents significantly enhanced the proliferation of normal mesangial cells, though the responses to neo synephrine and EGF were significantly lower as compared with the other growth promoters. The mitogenic effects of the first three agents on single kidney mesangial cells were significant, but blunted as compared with control cells. This blunting was not evident in the case of the latter two mitogens. A significant enhancement of proliferation of mesangial cells originating from protein-fed rats was produced by epinephrine, neo-synephrine, and EGF. These effects were statistically not different from those observed in normal mesangial cells. The proliferative response to each of the mitogens used in the study proved highly specific for each mitogen, since it was abolished by respective specific inhibitors. Mesangial cells may play a role in the activation and later in progressive inhibition of renal hypertrophy in vivo. PMID- 10867541 TI - Transient and sequential expression of chemokine mRNA in glomeruli in puromycin aminonucleoside nephrosis. AB - Chemokines are a large family of low-molecular-weight proinflammatory cytokines that stimulate recruitment of leukocytes. We previously reported that among six chemokines, the expression of mRNAs for MCP-1, MCP-3, TCA3, and MIP-1alpha, but not for MIP-1beta and RANTES, was markedly elevated in the renal cortex of rats with puromycin aminonucleoside induced nephrosis. In this study we have determined the glomerular expression of the chemokine mRNAs in this model using quantitative competitive reverse-transcriptase polymerase chain reaction. After an injection of puromycin aminonucleoside, the number of monocytes/macrophages and CD4+ and CD8+ cells markedly increased by day 5 and increased thereafter until day 10. The levels of mRNAs for MCP-1, MCP-3, and lymphotactin increased on day 5 and returned to their normal levels by day 7. The level of TCA3 mRNA increased on day 3, and that of MIP-1alpha mRNA increased on day 7, but both returned to their normal levels within 2 days. No increase in the mRNAs of MIP 1beta or RANTES was observed until day 10. These results indicate that the expression pattern of the chemokine mRNAs in glomeruli resembles that in renal cortex, but is more transient and sequential. PMID- 10867542 TI - Expression pattern in a modified equalized kidney cDNA library of hypertensive rat. AB - BACKGROUND: Genes with important functions and rarely expressed would probably more easily be cloned from a modified equalized kidney cDNA library for further investigation. METHODS: A kidney cDNA library of a spontaneously hypertensive rat was synthesized by a modified equalization method. Inserts of random clones were amplified by PCR and sequenced. Sequences were compared against a nonredundant database in GenBank. The cDNA profile was compared with an expression profile of a mouse renal proximal tubule cDNA library. Seven clones were analyzed by Northern blot analysis. The cDNA ends of two novel genes were amplified by PCR, sequenced and analyzed. RESULTS: 336 cDNA clones were analyzed and grouped into 323 species of transcript with 77 species similar to previously reported genes. Northern blot analysis identified one kidney-specific, one rarely expressed and lung-specific, and another relatively testis-specific gene. Two novel genes were cloned. One was 4.1 kb in length and encoded a 390-amino acid zinc-finger protein. Another was 2.5 kb and encoded a 474-amino acid protein of unknown function. Compared with the expression profile of a mouse renal proximal tubule cDNA library, this kidney library had a lower proportion of ribosomal genes and had a greater proportion of genes for signal transduction and DNA or RNA binding. CONCLUSIONS: Rare or novel genes could be more easily isolated from this library for molecular study of hypertension and renal pathophysiology. PMID- 10867544 TI - L-Carnitine supplementation in a hemodialysis patient with a mutation in the mitochondrial tRNA(Leu(UUR)) gene. PMID- 10867543 TI - Hemodialysis-induced degranulation of polymorphonuclear cells: no correlation between membrane markers and degranulation products. AB - BACKGROUND/AIMS: Degranulation of polymorphonuclear leukocytes (PMN) during hemodialysis (HD) is usually assessed by measuring degranulation products. However, this process might also be estimated by the assessment of cell surface markers. In this study, the relationship between the expression of PMN degranulation markers (CD63 and CD66b) and the release of degranulation products [myeloperoxidase (MPO) and lactoferrin (LF)] was investigated during clinical HD in order to evaluate cell surface markers as a useful index of PMN degranulation. METHODS: The expression of CD63 and CD66b on PMN and the release of MPO and LF were investigated in 10 chronic HD patients, during both heparin (HDhep) and trisodium citrate anticoagulation (HDcit), in a randomized order. Samples were drawn from both the efferent and afferent lines of the dialyzer at 0, 7.5, and 180 min. RESULTS: During HDhep at first passage, a major increase in MPO (from 158 +/- 32 to 448 +/- 177 microg/l, p = 0.001) and LF (from 134 +/- 52 to 260 +/- 120 microg/l, p = 0.01) was found across the dialyzer, whereas marked changes were not observed during HDcit. The expression of CD63 and CD66b increased across the dialyzer during both anticoagulation modalities, but was only significant in the case of HDhep (CD63: mean fluorescence intensity from 247 +/- 61 to 331 +/- 118, p < 0.01; CD66b: mean fluorescence intensity from 340 +/- 76 to 434 +/- 103, p = 0.01). During HDhep a correlation was noted between the degranulation products and markers of both azurophilic and specific granules (MPO and CD63: r = 0.35; p < 0.01; LF and CD66b: r = 0.39, p < 0.01). Significant differences in the expression of CD63 and CD66b between HDhep and HDcit were not observed. When analyzing the combined data for both HDhep and HDcit, no correlation was observed between degranulation products and markers. CONCLUSION: Our data suggest that the measurements of cell surface markers may not be a reliable indicator of the degree of HD-induced PMN degranulation. PMID- 10867545 TI - Evaluation of changes in amino acid metabolism in pre-dialysis patients treated with erythropoietin. PMID- 10867546 TI - Spontaneous perirenal hematoma as the initial manifestation of polyarteritis nodosa and long-term follow-up without treatment. PMID- 10867547 TI - Effect of dilazep dihydrochloride on plasma P-selectin concentrations in patients with IgA nephropathy. PMID- 10867548 TI - Pasteurella multocida as a rare cause of peritonitis in peritoneal dialysis. PMID- 10867549 TI - Congenital hypohaptoglobinemia in childhood hemolytic uremic syndrome? PMID- 10867550 TI - (+/-)3,4-Methylenedioxymethamphetamine ('Ecstasy')-induced serotonin neurotoxicity: studies in animals. AB - The popular recreational drug, (+/-)3, 4-methylenedioxymethamphetamine (MDMA; 'Ecstasy') is a potent and selective brain serotonin (5-HT) neurotoxin in animals. MDMA-induced 5-HT neurotoxicity can be demonstrated using a variety of neurochemical, neuroanatomical and, more recently, functional measures of 5-HT neurons. Although the neurotoxic effects of MDMA in animals are widely accepted, the relevance of the animal data to human MDMA users has been questioned, largely because dosages of drugs used in animals are perceived as being much higher than those used by humans. In the present paper, we review the extensive body of data demonstrating that MDMA produced toxic effects on brain 5-HT neurons in animals and present new data indicating that levels of the type 2 vesicular monoamine transporter are reduced in MDMA-treated animals, providing further indication of MDMA's 5-HT neurotoxic potential. Further, we demonstrate, using principles of interspecies scaling, that dosages of MDMA known to be neurotoxic in animals fall squarely in the range of dosages used typically by recreational MDMA users. PMID- 10867551 TI - (+/-)3,4-Methylenedioxymethamphetamine ('Ecstasy')-induced serotonin neurotoxicity: clinical studies. AB - (+/-)3,4-Methylenedioxymethamphetamine (MDMA, 'Ecstasy') is a brain serotonergic neurotoxin in experimental animals, including nonhuman primates. It is also an increasingly popular recreational drug of abuse, and doses of MDMA that are used recreationally overlap with those that produce serotonin (5-HT) neurotoxicity in animals. Studies in human MDMA users probing for evidence of brain serotonergic neurotoxicity indicate that some MDMA users may incur MDMA-related 5-HT neural injury and, possibly, functional sequelae. In particular, MDMA users have selective decrements in cerebrospinal fluid 5-hydroxyindoleacetic acid and brain 5-HT transporters, similar to nonhuman primates with documented MDMA-induced neurotoxicity. Functional abnormalities seen in MDMA users that may be related to 5- HT injury include cognitive deficits, altered sleep architecture, altered neuroendocrine function, altered behavioral responses to 5-HT selective drugs, and increased impulsivity. Additional studies in animals, as well as longitudinal and epidemiological studies in MDMA users, are required to confirm and extend the present data, and to determine whether MDMA users are at increased risk for developing neuropsychiatric illness as they age. PMID- 10867552 TI - Human research on MDMA (3,4-methylene- dioxymethamphetamine) neurotoxicity: cognitive and behavioural indices of change. AB - Laboratory animals can develop serotonergic neurotoxicity after repeated doses of 3,4-methylenedioxymethamphetamine (MDMA) or 'Ecstasy'. If similar neural damage occurs in humans, this may be evident in cognitive or behavioural impairments. In a review of the behavioural skills shown by drug-free recreational Ecstasy users, three aspects of cognitive performance are often affected: reduced memory for new information (Rivermead Behavioral Memory, supraspan word recall), impaired higher executive processing (Wisconsin Card Sort, Tower of London), and heightened impulsivity (Impulsiveness, Venturesomeness and Empathy Questionnaire, Matching Familiar Figures test). Performance on other more basic cognitive functions is generally unimpaired (simple reaction time, choice reaction time, number vigilance, Stroop, trail making). Some Ecstasy users also complain of poor memories and/or concentration difficulties, which they attribute to MDMA use. There are many methodological problems and uncertainties with research in this field: non-random allocation of subjects to drug conditions, the deleterious effects of other psychoactive drugs, and the possibility that these adverse profiles reflect pre-existing personality characteristics in Ecstasy users. However, this particular pattern of cognitive decrements in humans, is consistent with the animal data on those brain areas showing serotonergic damage following MDMA: the frontal cortex (impulsivity and higher cognitive impairments), and hippocampus (memory deficits). Finally, this profile of cognitive deficits is also consistent with a hypothetical integrative construct: namely reduced cortical inhibition. PMID- 10867553 TI - Potential human neurotoxicity of MDMA ('Ecstasy'): subjective self-reports, evidence from an Italian drug addiction centre and clinical case studies. AB - The present paper attempts to give an updated overview of the magnitude of the phenomenon of ecstasy abuse in Italy and other European countries. It gives an account of some clinical case studies and of a larger-scale report on polydrug (including MDMA) consumers attending our Public Health Addiction Treatment Unit in recent years, with a view to clarifying the characteristics and psychopathological consequences (mainly depression, psychotic disorders, cognitive disturbances, bulimic episodes, impulse control disorders, panic disorders, social phobia) of MDMA consumption. Longer-term, larger-dose (acute or cumulative) MDMA consumers were found to be at high risk of developing these psychopathological disturbances. A tentative description of certain personological dimensions of ecstasy consumers is also given (the novelty-seeking dimension was characteristic of those who occasionally experimented with the drug) while those who ingested larger quantities revealed low harm avoidance scores). Results are discussed in the light of the complex and different methodological issues arising from this kind of study, in which MDMA is far from being the only drug of abuse. PMID- 10867554 TI - Is MDMA ('Ecstasy') neurotoxic in humans? An overview of evidence and of methodological problems in research. AB - Evidence from research with a range of animal species, from rodents to non-human primates, has shown that MDMA (+/-3, 4-methylenedioxymethamphetamine) is neurotoxic. This article explores the evidence that MDMA may be neurotoxic in humans by briefly overviewing three types of research: (1) neurobiological, (2) psychological/somatic and (3) psychiatric. The first type of evidence derives from neuropharmacological and neuroendocrine studies, the second type focuses on psychological function and somatic symptoms in MDMA users, and the third involves studies of psychiatric cases in people who have taken MDMA. Evidence from these types of studies is indirect and differs in the degree to which any causative links are implied between observed effects, MDMA use and human neurotoxicity. These issues are critically discussed within the context of the wide-ranging methodological problems in human research with MDMA. PMID- 10867555 TI - 'Is MDMA a human neurotoxin?': diverse views from the discussants. AB - Every discussant at the Novartis symposium was invited to submit a 250-word abstract, giving their views upon the question: 'Is MDMA a human neurotoxin?'. These abstracts are presented here. They illustrate a wide range of viewpoints and opinions, as might be expected from experts in such diverse fields: animal neuroscience, human cognitive testing, police pathology laboratory, psychotherapeutic institute and psychiatric hospital. Some abstracts emphasized the methodological weaknesses of the human empirical data: the uncertain nature of 'Ecstasy' tablets, the reliance on self-report data, and the contributory factors of heat, dancing/exertion, poor diet and other illicit drugs. These factors may lead to psychobiological changes, which could be misinterpreted as neural damage. The absence of gliosis in animal models was also noted, which led to suggestions that there might be alternative interpretations for the neural changes which have been observed in rats and monkeys. Others noted the absence of neural/behavioural change following a single Ecstasy tablet, or commented upon the therapeutic benefits of MDMA in a quiet supportive environment. Nevertheless, novel studies from England, Germany, Italy, the Netherlands, Scotland and Wales confirmed and extended the range of cognitive, behavioural, EEG and neurological deficits, displayed by drug-free Ecstasy users. Moreover, these deficits often remained when other illicit drug use was statistically controlled. IN CONCLUSION: If MDMA neurotoxicity in humans is a myth, then it is a myth with a heavy serotonergic component. PMID- 10867556 TI - Nonselective partial dorsal rhizotomy: a clinical experience with 1-year follow Up. AB - The ability to perform a 'selective' dorsal rhizotomy has been challenged. EMG responses are inconsistent and often do not represent reflex responses. We perform nonselective partial dorsal rhizotomy (NSPDR) when reflex response is not evident. Ten children undergoing primarily NSPDR were evaluated preoperatively and postoperatively with the Modified Ashworth Scale, gait lab analyses gross motor function measure and the NSPDR was performed by nonselectively sectioning 50-75% of the dorsal roots not demonstrating a reflex response. Standard selective rhizotomy was performed in the remainder. Only 17 of 106 (16%) dorsal roots demonstrated reflex responses. The results reported in this study demonstrate a benefit to patients undergoing primarily NSPDR which is similar to that reported for patients in whom a selective procedure was intended. PMID- 10867557 TI - Epidemiology of infantile hydrocephalus in Saudi Arabia: birth prevalence and associated factors. AB - INTRODUCTION: Hydrocephalus is a common central nervous system disorder in children. In spite of its importance, it has not been subjected to sufficient epidemiological studies, particularly in the developing countries. The aim of this study was to provide information from a representative series of newly diagnosed cases of infantile hydrocephalus on the birth prevalence, associated factors and mortality. METHODOLOGY: A prospective study was conducted over a 1 year period from April 1996 to March 1997 in the city of Al-Madinah Al-Munawarah, Saudi Arabia. Except for neural tube defects and brain tumors, all cases of hydrocephalus diagnosed within the first 28 days of life were included. A control group of 104 subjects was studied for comparison. Information about antenatal, natal and early postnatal periods were obtained by interviewing the mothers of the newborns. RESULTS: During the study period, 26 cases of infantile hydrocephalus were detected. The birth prevalence was 1. 6 per 1,000 live births. There was no sex preponderance as the male to female ratio was 1.2:1. Multiple pregnancies were detected in 21 (81%) cases. Nineteen (73%) cases were the product of consanguineous parent and 4 patients had a positive family history of hydrocephalus. The number of preterm infants was 16 (62%). The number of low birth weights (less than 2,500 g) was 18 (69%). An Apgar score of less than 8 occurred in 18 (69%) cases. The mode of delivery was vaginal in 15 (58%) women. The 6 months mortality rate was 23% (6 infants). CONCLUSION: The birth prevalence of infantile hydrocephalus in this study was significantly higher than in the developed countries. A positive family history of hydrocephalus, low birth weight, low Apgar score and abdominal delivery were found to be associated factors. The mortality rate in the first 6 months of life was significantly higher in hydrocephalus infants than in controls. PMID- 10867558 TI - Fetal MRI in the evaluation of intrauterine myelomeningocele. AB - BACKGROUND: Accurate fetal imaging is essential to the practice of maternal-fetal medicine. While ultrasonography has been the traditional mainstay of fetal imaging, its ability to resolve critical features of central nervous system (CNS) anatomy remains limited. As interest in intrauterine therapy for myelomeningocele has increased, so has the need for more accurate, noninvasive imaging of the CNS. Fetal magnetic resonance imaging (MRI) promises to fill the gap left by ultrasound. METHODS: Thirty-seven MRI scans of fetuses previously diagnosed with myelomeningocele were reviewed by 2 neuroradiologists. The ability of fetal MRI to resolve the commonest CNS stigmata of spina bifida, and the incidence and extent of interobserver error, was assessed. In 4 cases, postnatal MRIs were also available. These were compared to the corresponding fetal studies. RESULTS: The imaging quality with the technique used in this study was excellent, even without the use of maternal or fetal sedation. There were no complications, and the imaging times were minimal. Interobserver error was minimal with respect to the evaluation of ventricular dilatation and hindbrain herniation, but moderate in the description and location of the spinal lesion. As had previously been documented with ultrasonography, a reduction was seen in hindbrain herniation when comparing pre- and postnatal MRIs. CONCLUSION: It is concluded that fetal MRI is an effective, noninvasive means of assessing fetal CNS anatomy. Its ability to resolve posterior fossa anatomy is superior to ultrasonography while, with respect to the evaluation of hydrocephalus and the level and nature of the spinal lesion, it may be equivalent to inferior. Inclusion of the fetal MRI into the standard diagnostic armamentarium will probably await the next major advance in speed and resolution. It is conceivable that, with further advances, MRI might supplant ultrasonography as the diagnostic tool of choice for evaluation of fetal anomalies including myelomeningocele. PMID- 10867559 TI - Spontaneous regression of low-grade astrocytomas in childhood. AB - An 8-year-old boy with neurofibromatosis type 1 (NF1) and a biopsy-proven juvenile pilocytic astrocytoma of the hypothalamic/chiasmatic region was followed with serial MRIs over 4 years. Spontaneous tumor regression was followed by progression and biopsy; 6 months later, the tumor regressed again. This bimodal regression is rare, but highlights the variable natural history of low-grade gliomas in children with NF1 and the difficulty in evaluating response of such tumors to therapy. PMID- 10867560 TI - Arachnoid cyst rupture with concurrent subdural hygroma. AB - Arachnoid cysts (ACs) are relatively common intracranial mass lesions, which occur most often in the middle cranial fossa. While these lesions can present as a mass lesion, many are asymptomatic. Rarely, posttraumatic or spontaneous rupture of ACs can result in intracystic hemorrhage, subdural hematoma or subdural hygroma. We have encountered two cases of ruptured arachnoid cysts that resulted in subdural hygromas. Both patients harbored middle cranial fossa cysts and suffered mild closed head injuries. The presentation, radiographic findings and surgical management of these patients as well as the association between ACs and subdural hygromas are described. PMID- 10867561 TI - Recurrent craniopharyngioma with nasopharyngeal extension. AB - We report the case of a 10-year-old boy having a recurrent craniopharyngioma with nasopharyngeal extension during a course of growth hormone therapy, in whom the nasopharyngeal craniopharyngioma was totally resected despite its extensive growth by using a transbasal approach. There has been no evidence of recurrence during 6 years of follow-up. A literature review was made with respect to nasopharyngeal extension of craniopharyngiomas, and the efficacy of the transbasal approach for those tumors is discussed. PMID- 10867562 TI - Spinal atypical teratoid/rhabdoid tumor in an infant. AB - Atypical teratoid/rhabdoid tumor of the central nervous system in infancy and childhood was established as an entity based on histological, immunohistochemical, and cytogenetic studies. We report the case of a 7-month-old girl who presented with progressive paraplegia and hypesthesia of her legs. Imaging studies revealed a spinal cord mass occupying the entire spinal canal below the T(7) level. Through a T(12)-L(3) laminectomy, the intramedullary tumor was partially debulked. Histologically, the tumor specimen had rhabdoid cells, and immunostaining showed vimentin and cytokeratin positivity. No abnormality of chromosome 22q was detected with the fluorescence in situ hybridization method. PMID- 10867563 TI - Thrombosis of the internal carotid artery secondary to soft palate injury in children and childhood. Report of two cases. AB - Trauma to the soft palate is a uncommon event during childhood. Stroke following intraoral trauma is also rare, but has been well documented by the current literature as a potentially serious complication. In this article, we report 2 cases of posttraumatic internal carotid artery thrombosis depicted by imaging studies. We discuss pathogenesis, and the literature is reviewed. PMID- 10867564 TI - Initial management of children with hypothalamic and thalamic tumors and the modifying role of neurofibromatosis-1. AB - Diencephalic gliomas may be grouped into 2 clinical categories. Optic pathway/hypothalamus gliomas (OPG) arise primarily from a slower-growing juvenile pilocytic astrocytoma, and thalamic gliomas arise primarily from a fibrillary astrocytoma which can become clinically and histologically more aggressive. Children with OPG have an excellent long-term prognosis with a 10-year survival of over 85%. The major therapeutic challenge for these patients is to maximize their quality of life by preserving visual and endocrine function while minimizing treatment-related morbidity. Treatment is often initiated at diagnosis in infants and toddlers who have a major visual impairment or the diencephalic syndrome. The judicious application of chemotherapy may serve to forestall the need for radiotherapy or surgery. Children with neurofibromatosis-1 (NF-1) usually have a more indolent course. Tumors may grow more slowly or occasionally regress spontaneously. However, over 90% of children with OPG without NF-1 will require some form of therapy. Patients with thalamic gliomas present with a shorter history, often with hydrocephalus. Surgical intervention is often required to relieve intracranial pressure and establish the histologic identity of the tumor. Over 75% of these tumors will become locally aggressive. Current multimodality therapy is relatively ineffective. The bithalamic variant behaves similarly to a pontine glioma. PMID- 10867565 TI - Images in Pediatric Neurosurgery. Lipoma of the nervous system. PMID- 10867566 TI - Images in Pediatric Neurosurgery. Chiari malformations. PMID- 10867567 TI - Concerning the article by Sharma et. al., Pediatr Neurosurg 1999; 31:150-154. Calcified subdural hematoma. PMID- 10867568 TI - Articulatory vowel lengthening and coordination at phrasal junctures. AB - Recent work has demonstrated that progressively stronger prosodic boundaries result in increasing amounts of lengthening for consonant gestures, both preceding and following the boundary. The present experiment extends these earlier findings by considering kinematic data collected with a magnetometer to determine (1) if vocalic gestures demonstrate lengthening patterns comparable to those of consonantal gestures due to adjacent prosodic boundaries, and (2) if the relative timing of consonant and vowel gestures is affected by adjacent boundaries. A magnetometer system was used to track articulator movements of 3 speakers producing sentences in which a /... C(1)V(1)(#)C(2)V(2).../ sequence was embedded with varying medial boundaries. All subjects demonstrated lengthening of the vocalic articulation before stronger boundaries. Additionally, effects on the relative timing of words spanning a boundary and the relative timing of segments in boundary-adjacent syllables were observed. PMID- 10867569 TI - Searching for an explanation for diphthong perception: dynamic tones and dynamic spectral profiles. AB - The aim was to find a psychophysical explanation for the perception, by naive listeners, of diphthongs as single vowels, even though they are essentially formant movements. Subjects were asked to match sinusoidal tone and resonance glides around 1,000 Hz with two connected steady-state tones or resonances whose frequencies could be controlled independently. The expectation was that short glides (below 120 ms) would give rise to single perceptual events without any movement in a particular direction, so that the two matching steady-state patterns would not show any frequency direction either; long resonance glides (above 120 ms), on the other hand, were expected to be perceived as rising or falling and matched accordingly. The results showed an effect of duration, although it interacted with glide width. At durations shorter than about 120 ms, subjects placed the two steady profiles with which they had to match the dynamic profile closer together than with durations over 120 ms; however, this only occurred if a glide covered more than 500 Hz, and is therefore irrelevant to diphthong perception. PMID- 10867571 TI - Obituary PMID- 10867570 TI - The question of 'Stress' in west greenlandic. An acoustic investigation of rhythmicization, intonation, and syllable weight. AB - The main purpose is to investigate whether stress is a relevant category in Greenlandic word prosody. I focus on tonal and durational parameters. The data material consists of word lists read aloud seven times by two subjects. The main findings are: (1) The prosodic characteristics of words can be explained in either tonal or durational terms. (2) The four different syllable types (of different 'weight') are distinguished in durational terms; further, there appears to be only a tripartite system of short, long and overlong. (3) There are intra syllabic as well as inter-syllabic rhythmical adjustments. It is concluded that Greenlandic prosody does not include an autonomous stress category, either tonal or durational parameters alone will do. And although Greenlandic has distinctive quantity, there is room for considerable durational variation of segments. PMID- 10867572 TI - The doublets of anger. PMID- 10867573 TI - The concept of anger: universal or culture specific? AB - I will suggest that the English word 'anger' and its counterparts in diverse languages of the world are based on concepts of anger that have a great deal of complexity. This conceptual complexity derives from several sources: (1) the metaphors and metonymies that apply to the concepts in various languages; (2) the prototypes of anger that people share in these cultures, and (3) the many different senses that the word anger and its counterparts have in different languages. We can ask: Are there any universal aspects of the concept(s) of anger? On the basis of linguistic evidence from English, Chinese, Japanese, Hungarian, Zulu and Wolof, I will suggest that there are, but I will also claim that some of the aspects are culture specific. This raises the further important question of why there is both universality and culture specificity in the conceptualization of this emotion. At stake is the issue of which of the following two contradictory claims is valid: (1) that anger is conceptualized in the same way universally, or (2) that anger is a social construction and thus varies considerably from culture to culture. I will propose a compromise view, which can be called 'body-based social constructionism', that enables us to see anger and its counterparts as both universal and culture specific. PMID- 10867574 TI - Fury: heroic, bestial and collective. AB - In his furies and frenzies, man's humanity is pushed to its outermost limits; his human potential is inordinately stretched. In poetic, heroic frenzy, man reaches the gods and divinity. Yet this poetic frenzy is not an oblivion, but a memory; his frenzy is not born from forgetting himself, but from a strengthening of his own ego. Bestial fury is the tale of the self-destructive consequences of attempting to use ferocity and fury against one's own enemies, and the endeavors to keep these forces in check and bend them towards one's own ends. However, the fact that man displays anger and wrath is not the sign of a descent towards the feral, animal realm, but, on the contrary, of an ascent. Man has risen up towards civilization. With this have come collective furies - the common angers of a human multitude, where the single personality is nullified. Might it be that even heroic frenzy, or perhaps above all heroic frenzy, has been liberally used in our times to unleash bestial and collective fury? PMID- 10867575 TI - Dante's inferno: phenomenology of a strange passion. AB - Dante's Inferno presents us with a spiritual adventure where the experience of wrath plays a crucial role. The infernal pilgrim's 'passage through wrath' is their only access to the lower part of Hell. Wrath - and the place where it is punished as a vice - occupy a very central position in the moral topography of Dante's Inferno: the Stygian Marsh, which defends the heart of Hell, its stronghold, and makes it inaccessible. Wrathfulness is situated at the borderline between 'carnal' and 'spiritual' sins, higher and lower Hell, weakness and wickedness of the will. What is the meaning of this situation? How is the sin of wrathfulness related to wrath as an expression of just indignation? How can the same passion have an angelical and an infernal face? What is anger's relation to reason? Does the trial gone through by the two pilgrims teach us something on the nature of this passion? Here are some questions which this paper tries to answer, thereby outlining a phenomenological theory of those passions which might be called 'dynamic'. PMID- 10867576 TI - When to get mad: adaptive significance of rage in animals. AB - The wide distribution of rage in animals suggests that rage should have an adaptive significance. In the present work, the function of rage is explored under an evolutionary perspective. I try to assess the selective advantage conferred to the individual presenting rage compared to one that does not. In this work, I considered animals under the 'strategist' perspective rather than the 'stimulus-reactor' one. I suggest that rage has a highly adaptive significance both as: (1) an emotion to prepare antagonistic actions and (2) as a communicative act. I suggest therefore that, as a communicative act, rage can be explored through the theory of games. In three crucial scenarios, I investigate, using the theory of games framework, when, and how, there is a selective advantage for individuals expressing, bluffing and simulating rage. PMID- 10867577 TI - Dysphoria, vulnerability and identity. An eulogy for anger. AB - Compared with the bulk of psychiatric literature dedicated to sadness or euphoria, dysphoric states have received relatively little attention. Perhaps we find ourselves facing a removal of anger from the horizon of contemporary psychopathology. Even less attention is given to the 'doublets' of anger. Anger marks off a region of the psyche within which the game of identity is played: as indignation, it defends the limits of that which is tolerable, the border upon which to keep watch, the trench from which to fight. But as fury, in the excess of the absolute affirmation of one's own existence, it incarnates the wreckage and the bloody fall of identity - i.e. pathology. The topic then collects the interweaving of the subjectivising character of rage on the one side, and on the other the explosive one, which breaks up the unifying and conciliatory direction which rationality enforces upon the subject - 'anger of life' and 'anger of death'. Both dimensions are explored with special concern to the reason for empathising with anger. PMID- 10867578 TI - Paranoia and dysphoria: historical developments, current concepts. AB - Regarding diagnostic criteria for paranoia (delusional disorder) today psychiatry usually refers to Kraepelin, suggesting that this diagnostic category has existed more or less unchanged since the end of the 19th century. But, reviewing German literature on this topic, one can find a lively discussion with regard to definition, development and course. In this context, the position of affective symptomatology - especially dysphoria with its various meanings - is of major interest. This paper aims at clarifying some definitory questions and at recalling the 'old' paranoia dispute which is worth rediscovering as a stimulation for modern classification and clinical practice. PMID- 10867579 TI - Dysphoria from a transnosological perspective. AB - The objective of our psychopathological analyses is to shed light on the position of irritable mood (dysphoria) in psychiatric diagnostics and nosology. In today's most commonly applied classification systems, the ICD-10 and the DSM-IV, dysphoria is mentioned mostly in the context of diagnostic criteria of personality and affective disorders. Other authors have emphasized the importance of dysphoric states in organic psychoses and delusional disorders. Summarizing the various publications on the nosological position, dysphoria is a nosological nonspecific syndrome which may occur in the course of all psychiatric disorders and illnesses. According to the results of our psychopathological analyses, the pathogenesis of dysphoria has to be considered as a multidimensional circular process in which various mental, physical and social factors act as predisposing, triggering and disorder-maintaining factors. Stressors induced by particular experiences and perceptions and by impaired health may lead to a dysphoric state if adequate coping mechanisms are missing. Dysphoria itself usually leads to a deterioration in the mental and physical state of the patient, and shows a clear impact on the patient's social network. The reactions of people close to the patient combined with the impaired mental and physical conditions of the patient cause the circle to restart. As contemporary diagnostic entities do not refer to pathogenesis, classical categorical diagnostics cannot provide the basis for effective pathogenesis-oriented therapy. A change of paradigm in diagnostics from a categorical to a dimensional approach thus becomes necessary. Following a dimensional diagnostic approach based on a dynamic model of vulnerability, a precise differential diagnosis of the complex constellation of conditions and their interactions becomes necessary in order to develop effective treatment strategies. Disorder-maintaining factors determine the treatment of the acute symptomatology, whereas predisposing and triggering factors serve as the basis for the prophylactic treatment. PMID- 10867580 TI - Dysphoria: a key for 'understanding' delusion? AB - The 'nonunderstandability' that traditional psychopathology attributes to 'true' delusion does not have a clear demarcation line, but, rather, it is a continuum of various delusional experiences. The attention paid to emotional situations and, specifically, to dysphoria, often contributes to making the delusional phenomenon, and, above all, its persistence, more understandable. A positive correlation between productive psychotic symptoms and the dysphoric mood often prevails in delusions with unfavorable prognoses. PMID- 10867581 TI - Dysphoria and aloneness in borderline personality disorder. AB - A close examination of dysphoria, anger and aloneness (three main characteristics of the borderline syndrome) provides a theoretical model of reference for the therapist. Dysphoria results from the cyclical emotional oscillation between hope for stability and disappointment in its inattainability; a dependent-anaclitic depression arises from the mixture of anger, aloneness and inner emptiness which is so characteristic of the borderline syndrome. The tendency to be immersed in the here-and-now, an intra-festum mentality, exacerbates the sense of isolation, causing more irritation, mute frustration and, consequently, anger. The effects and ramifications of anger, and the resultant precarious cohesion of the self, are explored in the borderline syndrome; they are especially illuminated by the application of Kernberg's pain-anger-hate-vengefulness cycle concept. Meanings of solitude, in its forms of aloneness and loneliness, are explored in their pertinence. Aloneness - the constant needy search for, but condemnation to never finding, objects to fill an inner sense of emptiness - is especially germane. Suggestions for assisting subjects with borderline personality disorder to overcome aloneness and the lack of historical progression are made. PMID- 10867582 TI - Anger and narcissism: between the void of being and the hunger for having. AB - The borderline syndrome, a typical marginal structure, is certainly a specific, autonomous pathology, with its own distinctive characteristics: among them, acting out, cyclical repetition of events without historical progression and anaclitic depression. Kohut's concept of 'narcissistic hunger' is particularly pertinent to the borderline condition: the borderline patient hungers to have that which is missing in his being. Through the application of the related notions of 'tragic man', 'self objects', 'grandiose self', and 'damaged self', the authors further develop their theory that the borderline syndrome has much in common with paranoid personalities. PMID- 10867583 TI - Biopsychosocial challenges of the new millennium. PMID- 10867584 TI - Psychosomatic medicine: why fix it if it ain't broken? PMID- 10867585 TI - Consultation liaison psychiatry and psychosomatics: strange bedfellows. PMID- 10867586 TI - Psychosomatic medicine: emerging trends and perspectives. AB - Developments have occurred in all aspects of psychosomatic medicine. Among factors affecting individual vulnerability to all types of disease, the following have been highlighted by recent research: recent and early life events, chronic stress and allostatic load, personality, psychological well-being, health attitudes and behavior. As to the interaction between psychological and biological factors in the course and outcome of disease, the presence of psychiatric (DSM-IV) as well as subclinical (Diagnostic Criteria for Psychosomatic Research) symptoms, illness behavior and the impact on quality of life all need to be assessed. The prevention, treatment and rehabilitation of physical illness include the consideration for psychosomatic prevention, the treatment of psychiatric morbidity and abnormal illness behavior and the use of psychotropic drugs in the medically ill. In the past 60 years, psychosomatic medicine has addressed some fundamental questions, contributing to the growth of other related disciplines, such as psychoneuroendocrinology, psychoimmunology, consultation-liaison psychiatry, behavioral medicine, health psychology and quality of life research. Psychosomatic medicine may also provide a comprehensive frame of reference for several current issues of clinical medicine (the phenomenon of somatization, the increasing occurrence of mysterious symptoms, the demand for well-being and quality of life), including its new dialogue with mind body and alternative medicine. PMID- 10867588 TI - Cognitive-behavioral therapy for somatization and symptom syndromes: a critical review of controlled clinical trials. AB - OBJECTIVE: Few treatments for somatization have been proven effective. In the past decade, however, clinical trials of cognitive-behavioral therapy (CBT) have been promising. Our aim was to critically review and synthesize the evidence from these trials. METHODS: A search of the Medline database from 1966 through July 1999 was conducted to identify controlled trials designed to evaluate the efficacy of CBT in patients with somatization or symptom syndromes. RESULTS: A total of 31 controlled trials (29 randomized and 2 nonrandomized) were identified. Twenty-five studies targeted a specific syndrome (e.g. chronic fatigue, irritable bowel, pain) while 6 focused on more general somatization or hypochondriasis. Primary outcome assessment included physical symptoms, psychological distress and functional status in 28, 26 and 19 studies, respectively. Physical symptoms appeared the most responsive: CBT-treated patients improved more than control subjects in 71% of the studies and showed possibly greater improvement (i.e., a trend) in another 11% of the studies. A definite or possible advantage of CBT for reducing psychological distress was demonstrated in only 38 and 8% of studies, and for improving functional status in 47 and 26%. Group therapy and interventions as brief as 5 sessions proved efficacious. Benefits were sustained for up to 12 months. CONCLUSION: CBT can be an effective treatment for patients with somatization or symptom syndromes. Benefits can occur whether or not psychological distress is ameliorated. Since chronic symptoms are exceptionally common and most studies were conducted in referral populations, the optimal sequencing of CBT in treating primary care patients and the identification of those most likely to accept and respond to therapy should be further evaluated. PMID- 10867587 TI - Assessing somatization in functional gastrointestinal disorders: integration of different criteria. AB - BACKGROUND: Psychiatric assessment of somatization (the tendency to experience and communicate psychological distress in the form of physical symptoms and to seek medical help for them) currently rests on DSM criteria. An alternative diagnostic and conceptual framework has been proposed by an international group of psychosomatic investigators. The aim of this study was to compare these new criteria (Diagnostic Criteria for Psychosomatic Research, DCPR) with DSM-IV in a population where a high prevalence of psychosocial problems is expected (functional gastrointestinal disorders, FGID). METHOD: One hundred and ninety consecutive patients with FGID in a tertiary care center were assessed according to DSM-IV and DCPR criteria. RESULTS: The number of the 12 DCPR diagnoses was almost double that of DSM diagnoses. Only 9% of the patients were not identified by DCPR criteria, whereas this occurred in 25% of patients using DSM criteria. While patients who were given a DSM diagnosis frequently had additional DCPR diagnoses, many patients with DCPR syndromes did not fulfill any DSM criteria. Four DCPR syndromes appared to be particularly frequent and accounted for almost three quarters of the total diagnoses (alexithymia, persistent somatization, functional symptoms secondary to a psychiatric disorder, demoralization). CONCLUSIONS: The joint use of DSM and DCPR criteria was found to improve the identification of psychological factors in FGID. The results may pave the way for changes in DSM classification of somatoform disorders. PMID- 10867589 TI - A clinical index for rating severity in Cushing's syndrome. AB - BACKGROUND: In assessing the clinical response to medical and/or surgical treatments in patients with established Cushing's syndrome, the need for an instrument that could measure the magnitude of changes in the most common symptoms was felt. We therefore tested the usefulness of a simple clinical index, the Cushing's syndrome severity index (CSI), based on clinimetric principles. Eight clinical features were selected. Each one was graded on an ordinal 3-point scale (0-2) with specification of anchor points based on severity. The total score ranged from 0 to 16. METHODS: Interrater agreement, construct validity and concurrent validity of the index were evaluated. Fourteen patients with Cushing's syndrome were evaluated independently by 2 endocrinologists before and after successful treatment. The CSI and two global scales of illness severity were administered. Urinary cortisol measurements were also performed. RESULTS: The intraclass correlation coefficient of the CSI was 0.95 before treatment and 0.87 after treatment. The CSI significantly discriminated (p<0.001) the effects of treatment and paralleled the changes in urinary cortisol. There were significant correlations between CSI and global scales of illness severity and change after treatment. CONCLUSION: The results indicate that the CSI is a valid and reliable clinimetric method to evaluate severity in Cushing's syndrome. It may provide a tool for better assessing the complex array of signs and symptoms in this condition. The index is suitable for descriptive studies, outcome investigations and treatment trials. PMID- 10867590 TI - Teaching and training in the psychiatric-psychosomatic consultation-liaison setting. AB - BACKGROUND: The consultation-liaison (C-L) psychiatrist is in an opportune position to undertake the tasks of education, training and assessment of performance as regards future physicians, psychiatrists, specialists in other branches and nurses. This paper describes the education and training programme in the Psychiatric-Psychosomatic C-L Service of Modena University Hospital. DESCRIPTION OF THE PROGRAMME: This programme consists of the following main activities: (1) daily group-case supervision, performed by the full-time psychiatrist together with the psychiatry residents of the C-L staff; (2) a bimonthly quality management meeting, which is part of a European project of measurement and improvement of quality of service; (3) weekly lectures on selected topics; (4) monthly tutorials in research techniques; (5) bimonthly presentations of literature reviews; (6) weekly clinical case conferences, which are the nucleus of the curriculum and which focus on the following main topics: 'the patients', 'the intervention' and 'the group', and (7) liaison meetings requested by non-psychiatric departments. CONCLUSIONS: A common denominator seen throughout the teaching and training activities of such a programme is the attitude of openness and effort toward integration which should be the C-L psychiatrist's distinguishing mark, in the context of the general hospital. PMID- 10867591 TI - Diagnosis and treatment of cystic fibrosis. AB - This review discusses some diagnostic aspects of cystic fibrosis (CF) including direct mutational analysis. Treatment of major disease manifestations is discussed in more detail with an emphasis on lung disease, in particular chronic infection with Pseudomonas aeruginosa which is responsible for the majority of excess morbidity and mortality. Centralised care and aggressive antimicrobial treatment have led to increased life expectancy and this may be even further increased by the demonstration that chronic P. aeruginosa infection may be prevented, or at least postponed for many years in a majority of patients. Adjunct treatment such as the use of local and systemic anti-inflammatory agents and inhalation of human recombinant DNase are also briefly touched upon. It is emphasised that important questions concerning the link(s) between the mutated gene and lung disease are still missing but that current research raises hope of a more causal treatment in the near future. PMID- 10867592 TI - Geriatric men at altitude: hypoxic ventilatory sensitivity and blood dopamine changes. AB - BACKGROUND: Short-term exposure to high-altitude hypoxia increases hypoxic ventilatory sensitivity (HVS) in healthy humans. Dopamine (DA) is the implicated neurotransmitter in carotid body (CB) chemoreceptor response, and the microenvironmental conditions in CB tissue are comparable to blood. Continuous DA infusion affected ventilation in animals and humans. Age-related oscillations in blood DA levels may influence peripheral chemoreflexes. OBJECTIVE: Hypoxic ventilatory responses (HVR) relative to blood DA concentration and its precursor, dihydroxyphenylalanine (DOPA) was measured in young and elderly men during short term altitude adaptation. METHODS: Nine elderly climbers (group 1:61+/-1.4 years) and 7 young healthy subjects (group 2: 23+/-2 years) were tested at sea level on day 0, on day 3 after passive transport to 2,200 m, and on day 14 after climbing to 4,200 and 5,642 m. RESULTS: Sea level HVR in group 1 was 47% lower than in group 2, accompanied by higher blood DOPA (300%) and DA (37%) content. Initial DA and DOPA concentrations showed a negative correlation with initial HVR but a positive correlation with age. Passive transport to middle altitude (2,200 m) increased HVS, doubling HVR slopes in groups 1 and 2 and producing increased maximum expired minute ventilation during isocapnic rebreathing (29 and 28%, respectively). Day 3 2,200-meter blood DOPA content decreased by 22% in group 1 and increased by 300% in group 2. DA increased in both groups. CONCLUSION: The relationship between HVR and the reciprocal DA and DOPA values seen in both groups is associated with age, producing decreased DA receptor sensitivity and enhanced DA reuptake during adaptation to high altitude. PMID- 10867593 TI - Decline in FEV(1) in community-based older volunteers with higher levels of neutrophil elastase in bronchoalveolar lavage fluid. AB - BACKGROUND: Neutrophil elastase (NE) is thought to be one of the key proteinases in the development of chronic obstructive pulmonary disease (COPD). Previously, we have shown that the NE-alpha1-proteinase inhibitor (NE-alpha1PI) complex in bronchoalveolar lavage (BAL) fluid was markedly elevated in asymptomatic smokers who had subclinical emphysema on CT scans. We proposed that excessive NE-alpha1PI complex in BAL fluid was a factor which might differentiate smokers who were developing emphysema from others. OBJECTIVE: In this study, we addressed the question of whether elevated levels of the NE-alpha1PI complex in BAL fluid are linked to the accelerated decline in pulmonary functions in those subjects. METHODS: We conducted a follow-up study to analyze the decline in FEV(1) for 4.3 years on average for 26 community-based volunteers who had received pulmonary function tests, CT scans and BAL. The levels of the NE-alpha1PI complex in BAL fluid and in plasma was measured. RESULTS: Neither pulmonary function measurements nor the presence of emphysema on CT scans could predict the decline in FEV(1). The number of inflammatory cells in BAL fluid was also not an indicator of progression. By contrast, subjects with higher levels of the NE alpha1PI complex in BAL fluid had a significantly accelerated decline in FEV(1) compared to those with lower levels. CONCLUSION: These data seem to support the hypothesis that NE in the lung is related to the onset and/or progression of COPD. PMID- 10867594 TI - Sleep apnea syndrome in patients with cardiac pacemaker. AB - BACKGROUND: Heart rhythm disturbances are cardiac side effects in patients with sleep-disordered breathing (SDB), which in itself is considered to be a risk factor for bradycardic rhythm disturbances. OBJECTIVE: We analyzed the prevalence and degree of SDB in patients who received a cardiac pacemaker due to bradycardic rhythm disturbances and investigated the relationship between the severity of an underlying SDB and the type of heart rhythm disturbance. METHODS AND RESULTS: 192 patients (100 males, 92 females, mean age 62.2 +/-12.2 years) were studied using the portable screening device MESAM IV. The respiratory disturbance index (RDI) was calculated visually. The mean RDI in all patients was 9.13+/-11. 09/h, 11.7+/ 13.15/h in males and 6.33+/-7.42/h in females. The prevalence ratio of SDB between men and women was 1.7:1, with significant differences in the respective severity (p<0.05). The screening showed a prevalence of SDB (RDI >10/h) of 32.3%. The highest prevalence was found in the group of patients with atrial fibrillation and bradycardia. However, there were no significant differences compared to other types of rhythm disturbances. The RDI in the population studied depended on age and body mass index, but not on the existence or type of rhythm disturbance and not on concomitant diseases. CONCLUSION: The prevalence of SDB in cardiac pacemaker patients is similar to that in patients of comparable age without a pacemaker. A heart rhythm disturbance does not seem to be an independent risk factor for development of SDB. Nevertheless, the differential diagnosis of bradycardic rhythm disturbances in this age group should include a screening for sleep apnea. PMID- 10867595 TI - Self-adjusting continuous positive airway pressure therapy based on the measurement of impedance. A comparison of free pressure variation and individually fixed higher minimum pressure. AB - BACKGROUND: Measurement of impedance using forced oscillation technique is a sensitive means of detecting airway obstructions, including the obstructive sleep apnea syndrome (OSAS). OBJECTIVE: The present study was conducted to determine whether treatment with an automated impedance-controlled continuous positive airway pressure (CPAP) device (APAP(FOT)) is possible in patients with OSAS, and which is the best range of pressure variation in automatical CPAP treatment. We investigated two modes of APAP(FOT) with different pressure ranges: (1) the widest technically possible pressure range and (2) a range with individually defined minimum pressure. METHODS: Ten patients [9 men, age 56.6+/-10.5 years, BMI 32.0+/-4.5 kg/m(2), apnea/hypopnea index (AHI) 18.2+/-13.3 /h] had a diagnostic polysomnography (baseline). After manual titration of positive airway pressure they were submitted, in randomized order, to two modes of the APAP(FOT) device, namely pressure range of 4.0- 15.5 mbar (mode 1 free range) and an individually fixed higher minimum pressure with a maximum pressure of 15.5 mbar (mode 2). RESULTS: While the manually titrated pressure was 8.0+/-1.3 mbar, in mode 1 it was 5.6+/-2.1 mbar (p<0.01); in mode 2 7.3+/-1.6 mbar (p< 0.05). Both of these modes suppressed abnormal respiratory events (baseline AHI 18.2+/ 13.3/h; mode 1: 2.5+/-1.9; mode 2: 1.8 +/-0.7, p<0.01 in each case), and increased slow wave sleep (baseline: 10.6+/-8.0%, mode 1: 20.2+/-10.4%, p<0.05; mode 2: 22.3+/-9.3%, p<0.01). In mode 1, the pressure was lower than that titrated manually in 73.2% of total sleep time, in mode 2 in 48.6%, while pressures higher than those derived manually were observed in 13.0% in mode 1 and in 19.1% in mode 2. CONCLUSIONS: The data indicate that impedance-controlled CPAP (APAP(FOT)) allows adequate treatment of OSAS patients at significantly lower pressures as compared with manually titrated pressure. Differences between the two modes are only minor. PMID- 10867596 TI - Interleukin-10 secretion by alveolar macrophages and monocytes in sarcoidosis. AB - BACKGROUND: Alveolitis and the production of proinflammatory cytokines are known features of sarcoidosis. Because of the usually spontaneous resolution of alveolitis despite local secretion of mediators causing inflammation and granuloma formation, we hypothesized that downmodulating mechanisms such as anti inflammatory cytokines might be involved in this process. OBJECTIVE: Investigation of the secretion of the macrophage deactivating cytokines interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) by alveolar macrophages in untreated sarcoidosis of the lung. METHODS: Fourteen consecutive and untreated patients with pulmonary sarcoidosis and 18 volunteers underwent bronchoscopy. Alveolar macrophages (AM) were obtained by bronchoalveolar lavage and the secretion of IL-10 and TGF-beta was studied. RESULTS: Spontaneous IL-10 production by AM was found in 6 of 14 patients and in 2 of 18 controls. The IL-10 level of lipopolysaccharide-stimulated AM was significantly higher in patients. Monocytes secreted significantly more IL-10 than AM, but there was no difference between sarcoid and control monocytes. No difference was found in the secretion of TGF-beta between patients and controls. CONCLUSION: Increased local secretion of IL-10 - but not TGF-beta - may represent a downmodulating mechanism involved in the spontaneous resolution of alveolitis in sarcoidosis. PMID- 10867597 TI - Influence of oxygen concentration on argon plasma coagulation-induced tissue damage in isolated pig tracheas. AB - BACKGROUND: Argon plasma coagulation (APC) is used during bronchoscopy to restore passage of central airways. Concern has been raised about the possibility that elevated oxygen concentration may increase the tissue damage by APC. OBJECTIVE: to examine the extent of tissue damage at different concentrations of oxygen in isolated pig tracheas. METHODS: The APC-induced tissue damage was investigated macroscopically and microscopically in isolated and artificially ventilated pig tracheas at oxygen concentrations of 21, 50, 75 and 100%. RESULTS: There was deep coagulation of all tissue layers reaching the adventitia independent of the oxygen concentration up to 75% O(2). With 100% O(2), only tissue damage increased and combustion occurred. CONCLUSIONS: APC-induced tissue damage in ex vivo pig tracheas was independent of the oxygen concentration relevant for clinical practice. PMID- 10867598 TI - Soot-exposed mononuclear cells increase inflammatory cytokine mRNA expression and protein secretion in cocultured bronchial epithelial cells. AB - BACKGROUND: Soot particles are air pollutants capable of inducing airway and lung parenchymal injury. Mononuclear and bronchial epithelial cells are central to the maintenance of homeostasis and inflammation in the airways. OBJECTIVES: The aim of this study was to evaluate the contribution of mononuclear cells to the release of inflammatory mediators by bronchial epithelial cells. METHODS: To model the in vivo situation, an in vitro system of cocultured blood monocytes and BEAS-2B cells was established in a transwell system. Blood monocytes were exposed to soot particles (FR 101) at concentrations of up to 100 microg/10(6) cells. Inflammatory cytokine mRNA and protein concentrations were quantified in BEAS-2B mono- and BEAS-2B-BM cocultures by RT-PCR and ELISA following exposure to soot for 1 and 8 h. RESULTS: No inflammatory cytokine mRNA expression was observed in unstimulated BEAS-2B cells. IL-6 and IL-8 mRNA and protein levels showed a dose dependent elevation in FR 101-exposed blood monocytes. In addition, both IL-6 and IL-8 mRNA expression was upregulated in cocultured BEAS-2B cells while cytokine concentrations in the blood monocyte-BEAS-2B coculture medium were significantly increased. This upregulation was likely due to a synergism of two cell populations. CONCLUSIONS: Exposure to soot particles induces an autocrine stimulation of inflammatory cytokine release by blood monocytes and BEAS-2B cells. Since IL-6 and IL-8 play a major role in the pathogenesis and persistence of bronchial inflammation, these findings may serve as a partial explanation for the aggravation of asthmatic and bronchitic symptoms after exposure to soot. PMID- 10867599 TI - Comparison of the effect on bronchial hyperresponsiveness of beclomethasone dipropionate administered via a novel multidose dry-powder inhaler or a conventional pressurised metered dose inhaler. AB - BACKGROUND: Treatment with inhaled corticosteroids improves symptoms and reduces bronchial hyperresponsiveness (BHR) associated with asthma. Delivery of drugs into the lung is dependent on the inhaler device. Furthermore, environmental concerns regarding the use of chlorofluorocarbon propellants in pressurised inhalers and patient acceptability of inhaler devices both influence the extent of use of different delivery systems. OBJECTIVES: To compare the efficacy of beclomethasone dipropionate (BDP) administered via a novel multidose dry-powder inhaler (DPI) and a conventional pressurised metered-dose inhaler (pMDI) with spacer in patients with BHR. METHODS: A randomised, double-blind, crossover study was carried out in a group of 27 patients (aged 19-55 years) with a clinical diagnosis of reversible airway disease, who demonstrated BHR to methacholine (PD(20) < or =6.4 mg). Each patient received BDP (< or =2 mg/day) via the DPI or pMDI, for periods of 4 weeks. The randomised treatment periods were preceded by 3 week washout periods when no corticosteroid was used. Five clinic visits marked the start and end of each study phase. The primary efficacy endpoint was BHR as defined by the pharmacodynamic parameter, PD(20), which was determined at the start and end of each treatment period. Clinical endpoints including lung function, symptoms and adverse events were also evaluated. RESULTS: Both treatments caused a significant decrease in BHR (p<0.05 vs. pre-treatment values). Mean +/- SD changes in log PD(20) were: DPI 0.59+/-1.29; pMDI 0.59+/ 0.94 mg. There was no statistically significant difference between treatments and no evidence of a carry-over effect between treatments on BHR. Clinical efficacy and safety parameters also demonstrated no statistically significant treatment differences, and patients found the DPI easier to use. CONCLUSION: Efficacy of BDP in reducing BHR is comparable via the DPI and pMDI plus spacer. PMID- 10867600 TI - Respimat (a new soft mist inhaler) delivering fenoterol plus ipratropium bromide provides equivalent bronchodilation at half the cumulative dose compared with a conventional metered dose inhaler in asthmatic patients. AB - BACKGROUND: Respimat, a possible alternative to the conventional metered dose inhaler (MDI), is a novel, reusable, propellant-free, multidose soft mist inhaler. Respimat slowly releases a metered dose of active substance as a soft mist with a high proportion of the dose in the fine particle fraction, leading to improved lung deposition following inhalation when compared with the conventional MDI. OBJECTIVES AND METHODS: The equipotent bronchodilating efficacy and safety of a combination of fenoterol hydrobromide and ipratropium bromide (F/I) in cumulative doses delivered by either Respimat or pressurised MDI was assessed in a randomised, controlled, double-blind (within device) 4-way crossover study. Forty-three patients with stable asthma (mean FEV(1) 62% predicted) responsive to F/I inhaled cumulatively 16 puffs on each of 4 test days (1 + 1 + 2 + 4 + 8 puffs at 50-min intervals) via Respimat delivering 50/20, 25/20 or 25/10 microg F/I per puff or via MDI delivering 50/20 microg F/I per puff. RESULTS: Cumulative doses of 400/160 and 400/320 microg F/I via Respimat produced bronchodilation (evaluated by average increase in FEV(1) 45-245 min after first inhalation) equivalent to that achieved with a cumulative 800/320 microg F/I via MDI (mean increase in FEV(1) above baseline 0.76, 0.73 and 0.71 litres, respectively). The tolerability of the F/I combination via Respimat was also comparable to that of twice the dose delivered via MDI. CONCLUSION: Therefore, a fenoterol hydrobromide/ipratropium bromide combination delivered by Respimat is as safe and effective as the MDI at half the cumulative dose, on acute administration to patients with asthma. PMID- 10867601 TI - Severe pancoast tumour. PMID- 10867602 TI - Bronchiolitis obliterans organizing pneumonia: a distinct pulmonary complication in cystic fibrosis. AB - Organizing pneumonia in cystic fibrosis has hitherto been considered a nonspecific reparative process. We report on an adult patient with cystic fibrosis and histologically proven bronchiolitis obliterans organizing pneumonia, who experienced sustained clinical improvement under corticosteroid therapy. This case suggests that bronchiolitis obliterans organizing pneumonia may be a distinct pulmonary complication in cystic fibrosis and improve with specific therapy. PMID- 10867603 TI - Tracheobronchial involvement in relapsing polychondritis. AB - Relapsing polychondritis (RPC) is a multisystem disorder of chondromalacia involving any cartilage. Respiratory tract involvement is the greatest threat to life. We report a patient with stenosis of the subglottic trachea and left main bronchus who suddenly ceased breathing. As this patient did not have any other clinical features of RPC, the diagnosis was difficult. CT showed circumferential worm-eaten-like thickening suggesting a deformity and edema of the tracheal mucosa. Biopsy of the tracheal and thyroid cartilage revealed mild cartilage degeneration and infiltration with inflammatory cells. Therefore, the patient was diagnosed as having RPC. She is currently well 24 months after Montgomery T tube intubation with systemic steroids. Narrowing of the left main bronchus has not worsened. PMID- 10867604 TI - Small cell lung carcinoma presenting as acute cardiovascular collapse due to tumour cell embolisation. AB - We present a case of small cell lung carcinoma causing acute cardiovascular collapse due to pulmonary tumour emboli. Although pulmonary tumour emboli may complicate a number of malignancies, this is rarely seen in cases of carcinoma of the bronchus. Patients suffering with pulmonary tumour emboli often have previous symptoms, and show progressive dyspnoea. To our knowledge there have been no reports of tumour emboli presenting acutely without any previous history of symptoms. PMID- 10867605 TI - Cystic schwannoma presenting as massive hemoptysis in an adult. AB - A 62-year-old man who presented with the chief complaint of hemoptysis is reported. A chest radiograph obtained on admission showed a huge cystic mass located at the posterior mediastinum. Prior to surgery, transarterial embolization was done because of continuous massive hemoptysis. An uneventful removal of the tumor was performed, and the pathological diagnosis was schwannoma. The hemoptysis was thought to have been caused by changes in the cystic formation in combination with inflammation which extended to the lung. PMID- 10867606 TI - Unilateral hilar lymphadenopathy of sarcoidosis or sarcoid reaction compressing the trunk of the right pulmonary artery. AB - A case of sarcoidosis or sarcoid reaction with a rare manifestation of unilateral lymphadenopathy compressing the trunk of the right pulmonary artery is presented. A 71-year-old woman was admitted for evaluation of a left hilar mass. Chest CT scans showed a mass invading the right pulmonary artery. A frozen section obtained following open lung biopsy showed lymph node tissue largely replaced by noncaseous granulomas indicating sarcoidosis or sarcoid reaction. Old uveitis was compatible with sarcoidosis, and no malignancy was evident. These findings suggested sarcoidosis, however, other evidence of sarcoidosis was not obtained. PMID- 10867607 TI - Malignant transient pleural transudate: a sign of early lymphatic tumoral obstruction. AB - In the absence of a responsible comorbid condition, the transudative character of a pleural effusion in patients with malignancy does not imply a favorable outcome. We report a case of colon carcinoma metastatic to lung and pleura presenting as a bilateral transudative pleural effusion. Tumoral diffuse lymphatic permeation was identified as the cause of lymphatic obstruction on pleural and transbronchial biopsies. The transudative character of the pleural effusion was transient denoting its obstructive origin. PMID- 10867608 TI - Fatigue associated with obstructive sleep apnea in a patient with sarcoidosis. AB - Many patients with sarcoidosis suffer from persistent constitutional symptoms such as fatigue and general weakness, even though physiological measures of disease activity returned within normal limits. The following case report demonstrates a sarcoidosis patient with recurring fatigue caused by an obstructive sleep apnea syndrome developed during the course of the disease. PMID- 10867609 TI - Hypolucent lung in a woman with recurrent haemoptysis. PMID- 10867610 TI - Malignant peripheral nerve sheath tumor of the mediastinum. PMID- 10867611 TI - Fluticasone propionate reduces serum interleukin-8 levels in asthmatic patients. PMID- 10867612 TI - Hepatocyte-derived soluble factors regulate proliferation and autocrine growth factor expression in colon cancer cell lines of varying liver-colonizing capability. AB - We investigated the role of hepatocyte-derived soluble growth factors on cell proliferation and expression of growth factors and their receptors in four colon cancer cell lines of varying liver-colonizing ability. Cocultures of hepatocytes and colon cells and cultures of colon cells with hepatocyte-conditioned medium resulted in growth inhibition of both weakly and strongly metastatic cell lines. Growth inhibition was accompanied by a reduced expression of erb-B2 in the colon cells after 4 days in the presence of hepatocytes. In LS174T and LiM6 cells, there was a dramatic reduction in heregulin-alpha levels in the presence of hepatocytes. Interestingly, after 2 days in culture, hepatocyte-derived soluble factors increased the mRNA levels for the EGF family members amphiregulin and cripto. These studies show an inhibitory effect of hepatocyte-derived soluble factors on the proliferation of colon cell lines, mediated in part by changes in the expression of autocrine growth factors and receptors of the EGF and heregulin family. PMID- 10867613 TI - Characterization of a new MUC1 monoclonal antibody (VU-2-G7) directed to the glycosylated PDTR sequence of MUC1. AB - A monoclonal antibody (MAb), VU-2-G7, was generated against a synthetic 60-mer MUC1 triple tandem repeat peptide with N-acetyl-galactosamine (GalNAc) O-linked to the threonine in the PDTR region of each repeat (3M GalNAc). VU-2-G7 and 8 MUC1 MAbs (VU-3-C6, VU-4-H5, 139H2, A76-A/C7, VU-12-E1, BCP9, MF11 and BW835) were tested against various glycosylated and nonglycosylated MUC1 tandem repeat peptides. VU-2-G7 showed strong reactivity with its immunogen, 3M GalNAc, and much lower reactivity with the nonglycosylated 60-mer MUC1 triple tandem repeat peptide. VU-2-G7 showed no reactivity with a 60-mer MUC1 triple tandem repeat peptide modified at the PDTR region or with a 60-mer MUC1 triple tandem repeat peptide with 3 GalNAc per repeat outside the PDTR region (9M GalNAc). In ELISA and flow cytometry, VU-2-G7 ubiquitously reacted with 4 MUC1-expressing breast cancer and 2 ovarian cancer cell lines and with a MUC1-gene-transfected Chinese hamster ovary cell line. The reactivity of VU-2-G7 was always higher than that of VU-4-H5, raised against a nonglycosylated 60-mer MUC1 triple tandem repeat peptide. Immunohistochemical staining of paraffin sections of breast and ovarian tumor tissues showed strong binding of VU-2-G7 predominantly at the cell membrane. The dominant epitope of VU-2-G7 is in the glycosylated PDTR motif of the MUC1 tandem repeat, and this epitope is abundantly present on the surface of tumor cell lines and breast and ovarian tumor tissues. Given the ubiquitously aberrant glycosylation of MUC1 in malignant cells, the production of MAbs against highly purified glycosylated MUC1 tandem repeat peptides may yield MAbs better suited for the immunotherapy of carcinomas than those available at the moment. PMID- 10867614 TI - Effects of differentiation inducers on cell phenotypes of cultured nontransformed and immortalized mammary epithelial cells: a comparative immunocytochemical analysis. AB - We examined the effects of the differentiation-inducing agents 1, 25 dihydroxyvitamin D(3) (calcitriol), phorbol myristate acetate (PMA), the cytokine interferon-gamma (IFN-gamma), and the tumor necrosis factor-alpha (TNF-alpha) alone and in combination with calcitriol on cell growth and differentiation parameters of cultured nontransformed human mammary epithelial cell (HMEC) lines, the chemically transformed HMEC line H184 A1N4, and the human mammary carcinoma cell line CAL51. Cell differentiation was phenotyped by semiquantitative immunocytochemistry using a panel of 15 monoclonal antibodies against marker molecules representing epithelial cell differentiation, cell-cell adhesion processes and malignancy. Cell proliferation of HMEC and H184 A1N4, but not of CAL51 cells was reduced by the agents. Cell phenotypes were analyzed by examining the expression of cytokeratins (pan CK and CK19), the epithelial mucin (MUC1), isoactin, and the blood group-related H type 2 carbohydrate antigen. HMEC and H184 A1N4 cells showed characteristics of basal cells, whereas CAL51 cells resembled a lumenal phenotype. The cell-cell adhesion molecules E-cadherin, intracellular adhesion molecule (ICAM-1), and the tumor markers carcinoembryonic antigen (CEA) and CD44v6 were expressed on all 3 mammary cell lines, with moderate differences. With respect to effects on cell phenotypes, HMEC were sensitive to PMA and IFN-gamma resulting in an increased expression of MUC1, CEA and ICAM-1 molecules. H184 A1N4 cells responded to TNF-alpha in combination with calcitriol by increased expression of pan CK, MUC1, and decreased H type 2 antigen expression, suggesting a transition towards a lumenal phenotype. Furthermore, CEA, ICAM-1 and CD44v6 were increased by TNF-alpha plus calcitriol. In contrast, CAL51 cells were overall less sensitive to differentiation induction attempts; only TNF-alpha stimulated MUC1, isoactin and epithelial cell adhesion molecule (Ep-CAM) expression. PMID- 10867615 TI - Specific detection and quantitation of SCC antigen 1 and SCC antigen 2 mRNAs by fluorescence-based asymmetric semi-nested reverse transcription PCR. AB - Squamous cell carcinoma antigen (SCCA) is expressed in normal squamous epithelia and malignant squamous cell tissues. The serum level of SCCA has been used to evaluate treatment efficacy, clinical course of disease, and recurrence. SCCA is produced by at least two genes (SCCA1 and SCCA2); both of them have been located on chromosome 18q21.3. It has been difficult to examine the expression levels of SCCA1 and SCCA2 mRNAs separately because of their high homology at nucleotide level. In the present study, asymmetric semi-nested reverse transcription PCR, based on the principle of fluorescence energy transfer, enabled to quantitate the copy numbers of both SCCA1 and SCCA2 mRNAs. Using this method, the expression levels of these mRNAs were evaluated in normal and malignant squamous tissues. The copy number of SCCA2 mRNA was higher in malignant tissues than in normal tissues, while those of SCCA1 mRNA did not significantly differ between normal and malignant tissues. These data indicate that specific quantitation of the expression level of SCCA2 mRNA may be useful for the diagnosis and management of patients with squamous cell carcinoma. PMID- 10867616 TI - Postoperative follow-up of breast cancer patients: overview and progress in the use of tumor markers. AB - The role of serial measurement of serum tumor-associated antigens (TAA) in the postoperative follow-up of breast cancer patients is not considered by most authors. The authors of this article review the literature and original data showing benefits from the use of TAA in the postoperative follow-up of breast cancer patients. An increase in the lead time from the first pathological finding to the definite evidence of distant metastases has been observed in follow-up studies which have used TAA compared to those which did not use them. The sensitivity and specificity of TAA in the diagnosis of distant metastases have been markedly improved by selecting an appropriate combination of TAA and by identifying specific conditions associated with breast cancer responsible for false-positive results as well as by adopting a 'dynamic' evaluation of multiple successive determinations of TAA. TAA accurately predict patients with bone metastases and can be used to guide imaging techniques. The anticipation of distant metastasis by TAA can be used to initiate a relatively early treatment which has been shown to prolong overall survival in a previous study. It is hoped that these data will stimulate further trials including TAA in the follow-up of breast cancer patients. PMID- 10867617 TI - Alterations of intestinal motor responses to various stimuli after Nippostrongylus brasiliensis infection in rats: role of mast cells. AB - Nippostrongylus brasiliensis infection induces jejunal mastocytosis associated with enteric nerve remodelling in rats. The aim of this study was to evaluate the intestinal motility responses to meals and to neurotransmitters involved in the control of gut motility (acetylcholine (carbachol), substance P and neurokinin A) in both control and N. brasiliensis-infected rats 30 days post-infection. All rats were equipped with NiCr electrodes in the jejunum to record myoelectrical activity. The duration of disruption of the jejunal migrating myoelectrical complexes (MMC) induced by the different stimuli was determined. Meal ingestion and substance P administration disrupted the MMC pattern for similar durations in the two groups. Carbachol and neurokinin A induced a significantly longer MMC disruption in post-infected rats than in controls (125 +/- 8.3 vs. 70 +/- 6 min for carbachol 100 microg kg-1 and 51 +/- 4 vs. 40 +/- 2 for neurokinin A 50 microg kg-1). The enhanced motor response in postinfected rats was reduced by previous mast cell stabilization with ketotifen or mast cell degranulation with compound BrX 537 A. In conclusion, the increased intestinal motor reactivity to carbachol and neurokinin A in post-N. brasiliensis-infected rats depends upon intestinal mast cell hyperplasia and degranulation. PMID- 10867618 TI - Effect of endotoxin on opossum oesophageal motor function. AB - Endotoxin induces nitric oxide (NO*) synthase and alters gastrointestinal functions. We explored the effect of lipopolysaccharide (LPS) on oesophageal motor function at 6, 12, 24, and 48 h. The effects of inhibiting inducible NO* synthase (iNOS) were studied 12 h after administration of LPS with/without aminoguanidine (AG). Oesophageal manometry was performed and tissue bath studies were performed with muscle strips from the oesophagus and lower oesophageal sphincter (LOS). Plasma nitrite/nitrate concentrations were determined. The amplitudes of peristaltic pressure waves, resting LOS pressure and the percentage LOS relaxations were diminished by LPS. AG attenuated the decrease in amplitude of oesophageal pressure waves, LOS pressure, and percentage relaxation of LOS brought about by LPS. LPS decreased electrical field stimulation (EFS)-induced relaxation of LOS muscle. AG attenuated this decrease in LOS relaxation. The off response of transverse oesophageal muscle strips was decreased, and AG antagonized this effect. Plasma concentrations of nitrite/nitrate were increased. The increase in plasma nitrite/nitrate was attenuated by AG. These studies support the hypothesis that endotoxin modulates oesophageal motor function by increasing NO production and suggest that this results from the induction of iNOS. PMID- 10867619 TI - Gastric electrical activity and gastric emptying in term and preterm newborns. AB - The aims of this study were to evaluate the gastric electrical activity and gastric emptying in preterm and term newborns and to assess the development of gastric motility by comparing newborns of different gestational ages. The cutaneous electrogastrography and the ultrasonographic study of the gastric emptying were performed before and after milk formula in three groups of infants: 12 preterm newborns with a gestational age of 28-32 weeks, 11 preterm newborns with a gestational age of 32-36 weeks, and 10 full-term newborns with a gestational age of 36-40 weeks. All recording sessions were performed 1 week after infants had reached full enteral feeding. The percentage of normal slow waves was similar in the three groups but it was not predominant compared to tachygastria in the earliest premature infants (59.3% (12.7-92.3) vs. 29.6% (3.7 78.8); P < 0.05). In addition, a progressive increase in the normal slow wave percentage (59.3% (17.4-87.4), 60.9% (38.1-89.7), 77.8% (66.4-84.8); P < 0.05) was observed as gestation advanced. As regards gastric emptying parameters, the antral area was greater and T(1/2) was longer in the preterm newborns of 28-32 weeks than preterm newborns of 32-36 weeks and full-term newborns (fasting antral area: 0.96 cm2 (0.6-1.5), 0.63 cm2 (0.4-1.2), 0.55 cm2 (0.1-0.9) respectively, P < 0.05; T(1/2): 83.4 min (76.0-108.5), 70 min (57.5-89.5) and 71.8 min (54.9 81.2), respectively P < 0.05). The comparisons of gastric emptying curves made among the three groups showed a reduced antral dilatation in preterm newborns of 28-32 weeks compared to full-term newborns at 30 and 60 min after a meal. In conclusion, although enteral feeding is important for the development process of gastrointestinal motility, gastric electrical activity and gastric emptying show an intrinsic maturation depending on the gestational age. PMID- 10867620 TI - Twenty-four hour ambulatory antroduodenal manometry in normal subjects (co operative study). AB - Circadian antroduodenal motor activity was studied in 40 normal subjects by means of a portable recording system consisting of a computerized data logger and a probe with microtransducers. The quantitative and qualitative characteristics of contraction events during the interdigestive and digestive periods, as well as during the awake and asleep periods, were analysed. The composition and timing of meals and night recumbence were standardized. In spite of the high interindividual variability in motor parameters, significant differences in the characteristics of interdigestive and digestive periods between waking and sleep states were found. This paper confirms the existence of a circadian variation in antroduodenal motor activity and provides reference values from a large series of normal subjects that can be used for statistical comparisons with those obtained from patients recorded with the same method. PMID- 10867621 TI - The effect of fentanyl, DNQX and MK-801 on dorsal horn neurones responsive to colorectal distension in the anaesthetized rat. AB - Certain dorsal horn neurones respond in a graded manner to noxious colorectal distension (CRD). Morphine inhibits these responses in the spinalized rat, but the role of excitatory amino acids in baseline visceral nociceptive transmission is less clear. This study examines the effect of the mu-opiate receptor agonist fentanyl, and the non-NMDA and NMDA antagonists DNQX and MK-801, respectively, on such responses to CRD in the sodium pentobarbitone-anaesthetized rat. Male rats were prepared for extracellular recording from the lumbosacral spinal cord. 90 neurones responsive to CRD, located throughout the dorsal horn, were classified according to their response duration and latency to 60 mmHg distension, as SL-A (short latency-abrupt; 59%), SL-S (short latency-sustained; 23%), L-L (long latency; 10%) and Inhib (inhibited; 8%). Convergent cutaneous receptive fields were mapped for 79/90 neurones and classified as LT (low threshold), WDR (wide dynamic range) or HT (high threshold). CRD (20-100 mm Hg) elicited graded responses in most neurones. In 6/6 SL-S neurones, fentanyl (1-8 microg kg-1) dose dependently inhibited the response to 60 mm Hg CRD, in a naloxone-sensitive manner, with an ID50 value (+/-95% confidence limits) of 2.48 (1.7-3. 7) microg kg-1. In 6/6 SL-A neurones, fentanyl had no significant effect on the response to CRD. DNQX (0.03-3 mg kg-1) produced a dose-dependent inhibition of the response to CRD in 5/5 SL-A neurones, with an ID50 value of 0.32 (0.01-41.1) mg kg-1. MK 801 (0. 03-0.3 mg kg-1) had no significant effect on responses to CRD in 6/6 SL-A neurones. The differential inhibitory effects of fentanyl on two neuronal subtypes may indicate functional differences. In SL-A neurones AMPA/kainate, but not NMDA receptors are involved in mediating baseline nociceptive neurotransmission. PMID- 10867622 TI - Effect of alpha-2 adrenoceptor antagonists on colonic function in rats. AB - We studied the effect of alpha-2 adrenoceptor antagonists on colonic function stimulated by water-avoidance stress, 5-hydroxytryptamine (5-HT), bethanechol and castor oil by comparison with the effects of atropine and a 5-hydroxytryptamine 3 (5-HT3) receptor antagonist, ondansetron. Yohimbine, idazoxan and atropine, but not ondansetron, significantly inhibited water-avoidance stress-stimulated faecal excretion. Yohimbine and idazoxan inhibited neither 5-HT- nor bethanechol stimulated faecal excretion. In contrast, atropine inhibited both 5-HT- and bethanechol-stimulated faecal excretion and ondansetron inhibited 5-HT-stimulated faecal excretion. Yohimbine did not inhibit the incidence of diarrhoea induced by castor oil, but idazoxan significantly inhibited diarrhoea observed during a 1-h period after the administration of castor oil. Both atropine and ondansetron inhibited diarrhoea during a 2-h period after the administration of castor oil. These findings suggest that alpha-2 adrenoceptor antagonists specifically inhibit colonic motor function stimulated by stress in rats. PMID- 10867623 TI - Glutamate receptors in the enteric nervous system: ionotropic or metabotropic? AB - Intracellular recording methods were used to investigate actions of glutamate on morphologically identified neurones in the myenteric and submucous plexuses of guinea-pig small intestine. Glutamate evoked a tetrodotoxin-resistant, slowly activating depolarizing response in most of the submucous neurones (86 of 125, 69%) and a smaller number of myenteric neurones (6 of 60, 10%). The depolarizing responses were restricted to S-type neurones with uniaxonal morphology. The group I metabotropic glutamate receptor (mGluRs) agonists quisqualate, 1S, 3R-ACPD and DHPG mimicked the depolarizing action of glutamate. A group I mGluRs antagonist, S-4-carboxyphenylglycine (S-4CPG), suppressed the glutamate responses with an IC50 of 357 microM at 30 microM glutamate. Group II or III mGluRs agonists did not produce depolarizing responses and group II or III mGluRs antagonists did not alter glutamate-evoked depolarization. The ionotropic glutamate receptor (iGluRs) agonists NMDA, AMPA, or kainate did not evoke depolarizing responses and glutamate-evoked depolarization was unaffected by the iGluRs antagonists D-APV, MK-801, or DNQX. No rapidly activating fast depolarizing responses reminiscent of fast excitatory postsynaptic potentials (EPSPs) were ever observed during application of glutamate or AMPA and stimulus-evoked fast EPSPs were unaffected by DNQX. The results suggest that the excitatory action of glutamate on enteric neurones is mediated by group I metabotropic glutamate receptors and that ionotropic glutamate receptors are not involved. The results also suggest that glutamate-mediated fast EPSPs may not be present in myenteric and submucous neurones in guinea-pig small bowel. PMID- 10867624 TI - Anatomy: A new look. PMID- 10867626 TI - The 2000 Henry Gray Award. PMID- 10867625 TI - AAA award winners. PMID- 10867627 TI - A.J. Ladman AAA/Wiley Exemplary Service Award. PMID- 10867628 TI - Giuseppe Sterzi (1876-1919) of the University of Cagliari: a brilliant neuroanatomist and medical historian. PMID- 10867629 TI - Chordate evolution and the origin of craniates: an old brain in a new head. AB - The earliest craniates achieved a unique condition among bilaterally symmetrical animals: they possessed enlarged, elaborated brains with paired sense organs and unique derivatives of neural crest and placodal tissues, including peripheral sensory ganglia, visceral arches, and head skeleton. The craniate sister taxon, cephalochordates, has rostral portions of the neuraxis that are homologous to some of the major divisions of craniate brains. Moreover, recent data indicate that many genes involved in patterning the nervous system are common to all bilaterally symmetrical animals and have been inherited from a common ancestor. Craniates, thus, have an "old" brain in a new head, due to re-expression of these anciently acquired genes. The transition to the craniate brain from a cephalochordate-like ancestral form may have involved a mediolateral shift in expression of the genes that specify nervous system development from various parts of the ectoderm. It is suggested here that the transition was sequential. The first step involved the presence of paired, lateral eyes, elaboration of the alar plate, and enhancement of the descending visual pathway to brainstem motor centers. Subsequently, this central visual pathway served as a template for the additional sensory systems that were elaborated and/or augmented with the "bloom" of migratory neural crest and placodes. This model accounts for the marked uniformity of pattern across central sensory pathways and for the lack of any neural crest-placode cranial nerve for either the diencephalon or mesencephalon. Anat Rec (New Anat) 261:111-125, 2000. PMID- 10867631 TI - Forthcoming topics PMID- 10867630 TI - Angiogenesis: new insights and therapeutic potential. AB - Angiogenesis, the formation of vessels from pre-existing vessels, is of critical importance not only during normal growth, but also in pathological situations. In the latter, some diseases are enhanced by excessive vascular growth (e.g., tumors), whereas in others inadequate vascular growth contributes to morbidity and mortality (e. g., ischemic heart disease). Our current state of knowledge makes it clear that the cascade of angiogenic events depends on complex processes that include cell-cell interactions, various intracellular signaling pathways, and the appropriate extracellular microenvironment. The literature regarding angiogenesis has increased exponentially during the last decade. Progress in this area is largely a consequence of advances in our understanding of angiogenic growth factor and cytokine function, in part due to the determination of their complete amino acid sequences and cloning of their genes. Other factors also play key roles in angiogenesis, including the extracellular matrix, adhesion molecules and their inhibitors, and metabolic and mechanical factors. The potential for developing therapeutic protocols has been enhanced by data from both in vitro and in vivo studies and has provided the rationale for clinic trials. Angiogenic therapy strategies include inhibition of aberrant angiogenesis, as seen in tumors or diabetes, as well as stimulation of angiogenesis in conditions of ischemia, such as ischemic heart or peripheral vascular disease. Anat Rec (New Anat) 261:126-135, 2000. PMID- 10867632 TI - Comparison of the conformation and dynamics of a polysaccharide and of its isolated heptasaccharide repeating unit on the basis of nuclear Overhauser effect, long-range C-C and C-H coupling constants, and NMR relaxation data. AB - A comparison of the conformation and dynamics of the cell wall polysaccharide of S. mitis J22 and the heptasaccharide repeating unit made from this polysaccharide was performed on the basis on nmr data. We have previously reported a model for this highly flexible polysaccharide in which four residues of the antigenic epitope adopt a defined conformation as do the two residues of the lectin-binding epitope. These domains are connected by a 6-substituted galactofuranoside residue that acts as a flexible hinge and the repeating subunits are joined by phosphodiester linkages that provide further flexibility. Homonuclear nuclear Overhauser effect (NOE) and long-range C-C and C-H scalar coupling constants measured in uniform (13)C-labeled samples of the polysaccharide and heptasaccharide were very similar, indicating a similar conformational average in solution. Significant differences in the solution dynamics were found from the heteronuclear relaxation data, T(1), T(1 rho), and NOE, which reflect the faster molecular tumbling of the heptasaccharide. Internal motions occurring on a picosecond time scale are relatively uniform along the polymer while dynamics on the time scale longer than a few nanoseconds is characteristic of hinge motion. PMID- 10867633 TI - Design, synthesis, and crystal structure of self-assembling norbornene (NBE) supported two-helix bundles: a unique example of Janus helicity in the solid state structure of NBE(Aib(5))(2). AB - Norbornene-supported bis-helical peptides with the general structure NBE(Aib(n) )(2) (NBE: 2,3-trans-norbornene dicarbonyl unit; Aib: alpha,alpha'-dimethyl glycine unit; n = 4,5) have been synthesized and examined for self-assembly preferences in the solid state. An x-ray study has revealed a phenomenon of Janus helicity in the solid state structure of NBE(Aib(5))(2). The lower homologue NBE(Aib(4))(2), however, shows an identical screw sense for both the helical arms. The difference in the handedness of left and right arms is reflected in the self-assembly patterns. Thus, while the NBE(Aib(4))(2) molecule self-assembles to form an infinite hydrogen-bonded superhelical ladder, the Janus molecule NBE(Aib(5))(2) crystallizes as individual units surrounded by water molecules. The structures of Z-Aib(4)-OMe and Z-Aib(5)-OMe are also presented to compare their conformations with the helical arms of the title compound and also to the already known structures of other X-Aib(n) -Y compounds. The helices in all the molecules are the 3(10)-type. PMID- 10867634 TI - The effect of sequence patterns and local conformational preferences on the structure of collapsed polypeptide. AB - We studied the structure of a polypeptide model by means of the Monte Carlo method. The model chain consisting of two kinds of residues (hydrophobic and hydrophilic) was confined on the (310) hybrid lattice. The residues interacted with the long-range contact potential. The short-range potential was also used by introducing the preferences of conformations corresponding to the helical structure. Simulations of the coil-to-globule collapse were done by an annealing process starting from high-temperature structures and then gradual cooling of the system. The parameters describing the behavior of the system were monitored. It has been found that in a case of a helical pattern -HHPPHPP- the collapsed chains consisted of helical fragments. The resulting structures were formed in such way that the polar residues were located in the outer shell of the molecule since the hydrophobic residues filled the inner part of molecule. The results showed that the proper balance between the magnitude of the potentials used in a model is important and its influence on final structures of molecules was discussed. PMID- 10867636 TI - Differential scanning calorimetry study of the hydrophobic hydration of the elastin-based polypentapeptide, poly(VPGVG), from deficiency to excess of water. AB - The polypentapeptide of elastin, poly(VPGVG), has become an interesting model polypeptide in understanding the mechanism of protein folding and assembly. Due to its simple amino acid composition and the predominance of apolar side chains, this polymer shows strong hydrophobic-hydration phenomena. This paper explores, by calorimetric methods, the nature and structure of the clathrate-like arrangements that take place, surrounding the apolar side chains of the polymer. The performance of these methods, especially differential scanning calorimetry, has a well-gained reputation. In this work, the development of the clathrate-like structures around this model polymer has been followed from water deficiency to water-excess states. Two main conclusions have been obtained from the data obtained. First, there is an upper limit of about 170 water molecules per pentamer as the number of water molecules required to form all the possible clathrate-like structures. Second, these structures exist as an inhomogeneous population with energies spreading in a significantly broad range, which is likely related to differences in geometrical parameters (bond lengths and angles) of the clathrate structure. PMID- 10867635 TI - High interaction alginate-hyaluronate associations by hyaluronate deacetylation for the preparation of efficient biomaterials. AB - The paper presents fundamental investigations of alginate-hyaluronate association with significant polymer interactions for preparation of efficient biomaterials. For this purpose, acetamide functions of hyaluronate were partly cleaved by hydrazine at high temperature, yielding amino groups accessible to carboxylic functions of the alginate chain. Alginate-hyaluronate association was studied both in dissolved state by rheological measurements and CD, and in the form of gel slabs prepared after calcium diffusion. Appreciable interaction between carboxylic groups of alginate and the released amino groups of hyaluronate was put into evidence by enhanced values of the viscosity of mixed solutions, and by assessment of the properties of the gel formed: moderate deacetylation allowed gels of improved hardness and viscosity. Nevertheless, high deacetylation was observed to hinder the gel formation by Ca(2+) complexation of alginate, by the significant competition of COOH-NH(2) association. Interaction between alginate and modified hyaluronate results in regular gel structure, with small cavities. PMID- 10867637 TI - Design and synthesis of AB(3)-type (A = 1,3,5-benzenetricarbonyl unit; B = Glu diOME or Glu(7) octa OMe) peptide dendrimers: crystal structure of the first generation. AB - The first generation molecule of glutamic acid-based dendrons on a 1, 3,5 benzenetricarbonyl core leads to a cylindrical assembly as demonstrated by single crystal x-ray diffraction. The benzene pi-pi stack (A) is stabilized by vertical NH...O===C hydrogen bonding with each subunit participating in three intermolecular hydrogen bonds related by three-fold rotation symmetry. PMID- 10867638 TI - Initiation of eukaryotic DNA replication: conservative or liberal? AB - The mechanism for initiation of eukaryotic DNA replication is highly conserved: the proteins required to initiate replication, the sequence of events leading to initiation, and the regulation of initiation are remarkably similar throughout the eukaryotic kingdom. Nevertheless, there is a liberal attitude when it comes to selecting initiation sites. Differences appear to exist in the composition of replication origins and in the way proteins recognize these origins. In fact, some multicellular eukaryotes (the metazoans) can change the number and locations of initiation sites during animal development, revealing that selection of initiation sites depends on epigenetic as well as genetic parameters. Here we have attempted to summarize our understanding of this process, to identify the similarities and differences between single cell and multicellular eukaryotes, and to examine the extent to which origin recognition proteins and replication origins have been conserved among eukaryotes. Published 2000 Wiley-Liss, Inc. PMID- 10867639 TI - The subapical compartment and its role in intracellular trafficking and cell polarity. AB - In polarized epithelial cells and hepatocytes, apical and basolateral plasma membrane surfaces are maintained, each displaying a distinct molecular composition. In recent years, it has become apparent that a subapical compartment, referred to as SAC, plays a prominent if not crucial role in the domain-specific sorting and targeting of proteins and lipids that are in dynamic transit between these plasma membrane domains. Although the molecular identity of the traffic-regulating devices is still obscure, the organization of SAC in distinct subcompartments and/or subdomains may well be instrumental to such functions. In this review, we will focus on the potential subcompartmentalization of the SAC in terms of regulation of membrane traffic, on how SAC relates to the endosomal system, and on how this compartment may operate in the context of other intracellular sorting organelles such as the Golgi complex, in generating and maintaining cell polarity. PMID- 10867640 TI - Copper induces apoptosis in BA/F3beta cells: Bax, reactive oxygen species, and NFkappaB are involved. AB - Copper, an essential trace element, can be toxic to some cells when present in excess. But thorough investigations into the cytotoxicity of copper and subsequent molecular mechanisms are rare, although the cytotoxicity of copper has been applied to cancer chemotherapy. The present study demonstrates that Cu(2+) inhibits [(3)H] thymidine incorporation in mouse pro-B cell line BA/F3beta and induces apoptosis. Apoptosis was mainly judged by morphology of cells, quantification of subdiploid DNA contents by flow cytometry, and detection of DNA fragmentation by gel electrophoresis. The apoptotic effect is dose and time dependent. Western blotting shows Bax is upregulated by Cu(2+). Bcl-2 overexpression can partially inhibit this apoptosis. Moreover, Cu(2+) increases the production of reactive oxygen species (ROS) in a dose-dependent manner. The antioxidant N-acetylcysteine (NAC) not only significantly inhibited copper induced apoptosis but also totally blocked generation of ROS, while Bcl-2 overexpression has no effect on the generation of ROS. Furthermore, our results show that NFkappaB is downregulated by Cu(2+). Bcl-2 overexpression or NAC can sustain the activity of NFkappaB. These data indicate that Cu(2+) might induce apoptosis in BA/F3beta cells via upregulation of Bax and ROS and subsequent inactivation of NFkappaB. PMID- 10867641 TI - The mitogenic activity of fibroblast growth factor-1 correlates with its internalization and limited proteolytic processing. AB - The fibroblast growth factor-1 (FGF-1) mitogenic signal transduction pathway is not well characterized, and evidence indicates that FGF-1 binding to and activation of cell-surface receptors is not solely sufficient for a full mitogenic response. Although initiation of the phosphorylation signaling cascades are likely important in FGF-1-induced mitogenic signaling, there appear to be additional signaling requirements. In this study, we demonstrate that FGF-1 internalization and subsequent processing correlates with the mitogenic potential of the growth factor on NIH 3T3 cells. Using site-directed mutants of FGF-1 and inhibitors of the endocytic and degradative pathways, we provide evidence for growth factor internalization and exposure to an acidic environment as necessary components of FGF-1-induced mitogenesis. In addition, a protease-sensitive event(s) appears critical for a complete mitogenic response to FGF-1, whereas, this protease sensitivity was not detected under the same conditions for serum stimulated mitogenesis. Therefore, proteolytic modification of internalized FGF-1 may result in the activation of additional, intracellular signaling events. PMID- 10867642 TI - Activation of heat shock factor 1 by hyperosmotic or hypo-osmotic stress is drastically attenuated in normal human fibroblasts during senescence. AB - We have previously reported that osmotic stress prominently induces the DNA binding activity of the heat shock transcription factor 1 (HSF1). In the present study, we examined the effects of medium osmolarity on both the activation of HSF1 and the programmed cell death in normal human fibroblasts during cellular senescence. The activation of HSF1 occurred rapidly in presenescent (early passage) IMR-90 cells when exposed to either hypo-osmotic or hyperosmotic stress. In contrast, the activation of HSF1 was significantly attenuated in senescent cells. Western blot analysis indicated that equal amounts of HSF1 were present as monomers in the cytoplasm of both presenescent and senescent cells in normal growth medium. Under either hypo-osmotic or hyperosmotic stress, trimerization and nuclear localization of HSF1 occurred in presenescent cells but not in senescent cells. More than 80% of HSF1 in senescent cells remained as monomers in the cytoplasm under osmotic stress, suggesting a defect in the signal transduction pathways that lead to HSF1 trimerization or a dysfunction in the HSF1 protein itself. Possible involvement of mitogen-activated protein kinase (MAPK) signal transduction pathways in the activation HSF1 was investigated by monitoring the activation of the three MAPKs, ERK1/2, JNK1/2, and p38, in cells exposed to hypo-osmotic or hyperosmotic stress. All three MAPKs were activated by hyperosmotic stress but not hypo-osmotic stress, suggesting that the MAPK signal transduction pathways may not be directly linked to the osmotic stress-induced activation of HSF1. In contrast to the rapid heat shock transcription factor (HSF) activation, apoptosis occurred only after long-term exposure to hypo osmotic or hyperosmotic stress. Despite the prominent induction of HSF1 activation, the presenescent cells were more sensitive than the senescent cells to the osmotic stress-induced apoptosis. PMID- 10867643 TI - The expression of Met/hepatocyte growth factor receptor gene in giant cell tumors of bone and other benign musculoskeletal tumors. AB - Overexpression of the hepatocyte growth factor receptor (Met/HGF receptor), a transmembrane tyrosine kinase encoded by the MET proto-oncogene, is involved in transformation and invasive behavior of human carcinomas and sarcomas. We have previously found that bone sarcomas express high levels of Met/HGF receptor while in some cases the ligand HGF is co-expressed with the receptor, activating an autocrine loop. In this study, we analyzed 40 biopsy samples of a collection of giant cell tumors and other rare benign tumors of bone for expression of the MET proto-oncogene. These included nonossifying fibromas, osteoblastomas, desmoplastic fibromas of bone, chondroblastomas, and giant cell tumors of bone. Snap frozen samples were tested for the MET and HGF gene expression by immuno histochemistry and Western blotting with anti-MET antibodies and RT-PCR. Over 50% of all cases scored positive for MET expression being constantly positive in recurrent or locally aggressive lesions. Sporadic co-expression of the Met/HGF receptor and ligand is also demonstrated. Met/HGF receptor expression in benign bone neoplasms suggests its early involvement in sarcomagenesis. PMID- 10867644 TI - Effects of CTGF/Hcs24, a hypertrophic chondrocyte-specific gene product, on the proliferation and differentiation of osteoblastic cells in vitro. AB - Connective tissue growth factor/hypertrophic chondrocyte-specific gene product Hcs24 (CTGF/Hcs24) promotes the proliferation and differentiation of chondrocytes and endothelial cells which are involved in endochondral ossification (Shimo et al., 1998, J Biochem 124:130-140; Shimo et al., 1999, J Biochem 126:137-145; Nakanishi et al., 2000, Endocrinology 141:264-273). To further clarify the role of CTGF/Hcs24 in endochondral ossification, here we investigated the effects of CTGF/Hcs24 on the proliferation and differentiation of osteoblastic cell lines in vitro. A binding study using (125)I-labeled recombinant CTGF/Hcs24 (rCTGF/Hcs24) disclosed two classes of specific binding sites on a human osteosarcoma cell line, Saos-2. The apparent dissociation constant (Kd) value of each binding site was 17.2 and 391 nM, respectively. A cross-linking study revealed the formation of (125)I-rCTGF/Hcs24-receptor complex with an apparent molecular weight of 280 kDa. The intensity of (125)I-rCTGF/Hcs24-receptor complex decreased on the addition of increasing concentrations of unlabeled rCTGF/Hcs24, but not platelet derived growth factor-BB homodimer or basic fibroblast growth factor. These findings suggest that osteoblastic cells have specific receptor molecules for CTGF/Hcs24. rCTGF/Hcs24 promoted the proliferation of Saos-2 cells and a mouse osteoblast cell line MC3T3-E1 in a dose- and time-dependent manner. rCTGF/Hcs24 also increased mRNA expression of type I collagen, alkaline phosphatase, osteopontin, and osteocalcin in both Saos-2 cells and MC3T3-E1 cells. Moreover, rCTGF/Hcs24 increased alkaline phosphatase activity in both cells. It also stimulated collagen synthesis in MC3T3-E1 cells. Furthermore, rCTGF/Hcs24 stimulated the matrix mineralization on MC3T3-E1 cells and its stimulatory effect was comparable to that of bone morphogenetic protein-2. These findings indicate that CTGF/Hcs24 is a novel, potent stimulator for the proliferation and differentiation of osteoblasts in addition to chondrocytes and endothelial cells. Because of these functions, we are re-defining CTGF/Hcs24 as a major factor to promote endochondral ossification to be called "ecogenin: endochondral ossification genetic factor." PMID- 10867645 TI - Type I collagen-induced osteoblastic differentiation of bone-marrow cells mediated by collagen-alpha2beta1 integrin interaction. AB - Bone marrow cells are multipotent cells. When bone marrow cells were cultured with type I collagen matrix gels, they showed high alkaline phosphatase activity, collagen synthesis, and formed mineralized tissues. Furthermore, cells expressed osteocalcin and bone sialoprotein genes, which are osteoblast-specific genes. These findings indicate that type I collagen matrix gels induce osteoblastic differentiation of bone marrow cells. Type I collagen interacts with the alpha 2 beta 1 integrin receptor on the cell membrane and mediates extracellular signals into cells. DGEA peptide is a cell-binding domain of type I collagen molecule. When collagen-integrin interaction was interrupted by the addition of Asp-Gly-Glu Ala (DGEA) peptide to the culture, the expression of osteoblastic phenotypes of bone marrow cells was inhibited. Furthermore, anti-alpha 2 integrin antibody, which interacts with alpha subunit of integrin and blocks the binding of integrin with collagen, suppressed the expression of osteoblastic phenotypes. These findings imply that collagen-alpha 2 beta 1 integrin interaction is an important signal for the osteoblastic differentiation of bone marrow cells. PMID- 10867646 TI - Integrin involvement in glioblastoma multiforme: possible regulation by NF kappaB. AB - Glioblastoma multiforme (GBM) is the most malignant astroglial-derived tumors which has the propensity to aggressively infiltrate normal regions of the brain surrounding the tumor. The interaction of tumor cells with the extracellular matrix (ECM) is an integral step in the process of tumorigenesis and may play a role in the local invasion of the GBM cells. Our study investigated the role of the nuclear transcription factor NF-kappaB on GBM integrin expression and cell attachment. Our results show that treatment of GBM cell lines, SNB-19 and T98G with PMA, an inducer of NF-kappaB, increased the expression of fibronectin and vitronectin genes. Accordingly, ectopic over-expression of NFkappaB subunits in GBM cells elevated the levels of fibronectin gene expression, providing direct evidence for a regulatory role for NF-kappaB in ECM protein production. Cell attachment to the ECM proteins including fibronectin, vitronectin and laminin was increased in GBM and normal astrocytic cells. Interestingly, treatment of cells with PMA augmented attachment of SNB-19 and T98G cells to fibronectin and vitronectin, however it had no effect on attachment of normal astrocytes. Addition of the tripeptide arginine-glycine-asparatic acid (RGD), the recognition site for many integrins, significantly inhibited SNB-19 and T98G cell attachment to fibronectin and vitronectin. Finally, activation of NFkappaB upon treatment of SNB cells with PMA led to an increase in the levels of mRNA for the beta3 and the alphav integrin subunits. Collectively, these data demonstrate a possible role for NF-kappaB in glioma cell attachment. PMID- 10867647 TI - Attenuation and loss of hormonal modulation of KGF (FGF-7)/KGF receptor expression and mitogenesis during mammary tumor progression. AB - Keratinocyte growth factor (KGF), alone and in synergism with progesterone (P) and prolactin (PRL), is mitogenic for normal mammary epithelium (ME) in vitro. In addition, P can upregulate ME sensitivity to KGF by slowing KGF receptor (KGFR) mRNA turnover in vitro. These hormonal interactions with KGF in vitro raise the possibility that alterations in these interactions can play a role in hormone dependent mammary tumor growth and progression. The effect of hormones on KGF mitogenesis and the regulation of KGFR expression was examined in pregnancy dependent (PDT) and ovarian-independent (OIT) mouse mammary tumors. In serum free, collagen gel cell culture, dose/response (2-20 ng/ml) and time course studies showed that KGF stimulated the proliferation of PDT (not OIT) cells but synergism with P or PRL was not observed. The level of KGFR mRNA in PDT cells was not significantly different from normal ME but in OIT it was reduced more than 90%. P did not affect KGFR mRNA turnover in cultured PDT cells. However, KGFR mRNA was more stable in PDT cells compared to normal ME; after 6 days culture in basal medium, KGFR mRNA levels declined 40% vs. 85% previously shown for normal ME. Determination of KGF mRNA levels in tissues showed that it was lower in PDT compared to normal mammary gland and not detectable in OIT. These data show that in PDT both KGF-stimulated mitogenesis and the regulation of KGFR expression are independent of hormones. OIT has progressed to independence from any KGF influence. Thus, a subset of hormonally regulated pathways related to epithelial/stromal cell interactions can be lost in hormone-dependent mammary tumors during tumor progression. PMID- 10867649 TI - Actin depolymerization and polymerization are required during apoptosis in endothelial cells. AB - In order to understand the role of actin microfilaments in the apoptotic process, we followed their evolution during tumor necrosis factor-alpha (TNF)-induced apoptosis in bovine aortic endothelial (BAE) cells. Using Western blotting analysis and immunofluorescence microscopy, we observed that the actin microfilaments network was disrupted in apoptotic cells. Depolymerization of F actin was concomitant with internucleosomal DNA degradation and with the morphological changes associated with apoptotic cell death. However, using the actin microfilament disrupting agent, cytochalasin, we present evidence that the formation of blebs leading to apoptotic cell fragmentation requires neopolymerization of actin. Indeed, in the presence of cyochalasin, induction of apoptosis (internucleosomal DNA degradation) in BAE cells by TNF and cycloheximide was not associated with these classical morphological markers of apoptosis. Moreover, when added to BAE cells showing incipient apoptotic fragmentation, cytochalasin E reversed this process. We also observed an accumulation of actin at the basis of the apoptotic bodies in formation in these cells. Together, these results suggest that the actin network of flattened cells is disrupted concomitantly to the morphological modifications associated to the apoptotic cell death, and that the cytochalasin-sensitive reorganisation of actin is required to the formation of apoptotic blebs. PMID- 10867648 TI - Activation of surfactant protein-B transcription: signaling through the SP-A receptor utilizing the PI3 kinase pathway. AB - This study describes receptor-activated signaling initiated by surfactant protein A (SP-A), and the means by which it activates transcription of surfactant protein B. Pulmonary surfactant is a mixture of lipids and associated proteins produced by type II pneumocytes. Interaction of SP-A with its cognate receptor (SPAR) on type II cells is involved in regulating surfactant secretion. This interaction also increases transcription of surfactant proteins and several other genes. To study SP-A cytokine activity, we used as a model surfactant-protein (SP-B) transcription, the activators of which have been characterized. HNF-3 and TTF-1 transcription factors are known to stimulate SP-B transcription. SP-A caused increased phosphorylation and nuclear localization of both. Corresponding increases in protein binding to the SP-B promoter were demonstrated by gel shift analysis. SP-A increased protein binding to HNF-3 and TTF-1 consensus recognition elements. Footprinting analysis indicated that SP-A-induced protein binding to SP B promoter was greater in amount, but not different in location, from that seen in control cells, which normally transcribe SP-B. SP-A caused transient increases in PI3 kinase localization at the plasma membrane, and SP-A signaling to elicit increased SP-B transcription was blocked by LY294002, an inhibitor of PI3 kinase. Therefore, SP-A signals through PI3 kinase to increase SP-B transcription in type II pneumocytes by enhancing TTF-1 and HNF-3 activation of the SP-B promoter. SP-A activation of this signaling pathway, which affects many cellular functions and has not previously been implicated in type II cell transcriptional activity, has profound import for understanding type II cell biology. PMID- 10867650 TI - Expression of functional tyrosine kinases on immortalized Kaposi's sarcoma cells. AB - Kaposi's sarcoma (KS) is the most frequent malignant lesion in patients with AIDS and is characterized by spindle cell proliferation, inflammatory cell infiltration, angiogenesis, edema, and invasiveness. KS origin is still debated. The complex aspect of this disease is probably supported by multiple concomitant pathogenetic factors, among which growth factors and their cognate tyrosine kinase receptors are deeply involved. Here we have investigated the expression status and functional integrity of KDR and Met receptors, as well as of their ligands, in an immortalized KS cell line (KS-IMM). The MET and KDR genes encode the tyrosine kinase receptors for Hepatocyte Growth Factor (HGF) and Vascular Endothelial Growth Factor (VEGF) respectively. Both factors are pleiotropic cytokines controlling growth, survival, motility, invasive migration and differentiation of endothelial cells. We have found that KS-IMM cells, which retain most of the features of the parental tumor and can induce KS-like sarcomas when injected subcutaneously in nude mice, express the Met receptor, but not its ligand. The receptor, which is basally inactive, is functional, being tyrosine phosphorylated in response to ligand stimulation and mediating the expected HGF dependent biological responses of motility, invasion and proliferation. Moreover, we report that KS-IMM cells coexpress VEGF and KDR and that KDR is constitutively tyrosine phosphorylated, possibly as a consequence of the establishment of an autocrine loop. The receptor, however, maintains responsiveness to exogenously added ligand, by increasing the level of tyrosine phosphorylation and by responding in biological assays of motility, invasion and proliferation. Finally, we have found that the two growth factors synergize in a motility assay. These data show that HGF and VEGF are growth factors active on KS-IMM cells. PMID- 10867651 TI - Angiogenesis inhibition by transdominant mutant Ets-1. AB - The expression of transcription factor Ets-1 is induced in endothelial cells (ECs) by angiogenic factor; and in turn Ets-1 converts ECs to angiogenic invasive phenotype. In order to control angiogenesis, we constructed a transdominant mutant Ets-1 (TMEts-1) which acts as a dominant negative molecule. This molecule inhibited the DNA binding and the transactivation activity of the wild-type Ets 1. Stable transfection of murine endothelial cell line MSS31 cells with the TMets 1 gene impaired angiogenic activities including proliferation, migration, invasion, and tube formation in type-1 collagen gel. Finally, we incorporated the TMets-1 gene into a non-proliferative adenovirus vector, designated as AdTMets-1. AdTMets-1 significantly inhibited angiogenesis in the Matrigel plugs injected into the subcutaneous tissue of C57BL mice. These results indicate that TMets-1 would be a tool for angiogenic inhibition. PMID- 10867652 TI - Lysosomal accumulation of drugs in drug-sensitive MES-SA but not multidrug resistant MES-SA/Dx5 uterine sarcoma cells. AB - Sequestration of drugs in intracellular vesicles has been associated with multidrug-resistance (MDR), but it is not clear why vesicular drug accumulation, which depends upon intracellular pH gradients, should be associated with MDR. Using a human uterine sarcoma cell line (MES-SA) and a doxorubicin (DOX) resistant variant cell line (Dx-5), which expresses p-glycoprotein (PGP), we have addressed the relationship between multidrug resistance, vesicular acidification, and vesicular drug accumulation. Consistent with a pH-dependent mechanism of vesicular drug accumulation, studies of living cells vitally labeled with multiple probes indicate that DOX and daunorubicin (DNR) predominately accumulate in lysosomes, whose lumenal pH was measured at < 4.5, but are not detected in endosomes, whose pH was measured at 5.9. However, vesicular DOX accumulation is more pronounced in the drug-sensitive MES-SA cells and minimal in Dx5 cells even when cellular levels of DOX are increased by verapamil treatment. While lysosomal accumulation of DOX correlated well with pharmacologically induced differences in lysosome pH in MES-SA cells, lysosomal accumulation was minimal in Dx5 cells regardless of lysosomal pH. We found no differences in the pH of either endosomes or lysosomes between MES-SA and Dx5 cells, suggesting that, in contrast to other MDR cell systems, the drug-resistant Dx5 cells are refractory to pH-dependent vesicular drug accumulation. These studies demonstrate that altered endomembrane pH regulation is not a necessary consequence of cell transformation, and that vesicular sequestration of drugs is not a necessary characteristic of MDR. PMID- 10867653 TI - Macaque red nucleus: origins of spinal and olivary projections and terminations of cortical inputs. AB - The cerebellar, spinal, bulbar, and cortical connections of the mammalian red nucleus imply a motor role. However, what information the red nucleus receives, processes, and distributes is poorly understood, partly because the rubral microcircuitry, especially in primates, remains incompletely defined. Multiple retrogradely transported fluorescent tracers were injected into the spinal cord and inferior olive of the macaque to label rubrospinal and rubroolivary neuron populations, respectively. Anterograde dextran amines were used to label the terminals of corticorubral neurons. These data provided the topographic framework for examining the morphology of rubral neurons in the accompanying paper (Burman et al. [2000]). Soma profiles of rubrospinal and rubro-olivary neurons were respectively segregated in the magnocellular and parvocellular nuclei. A subpopulation of neurons (DL-spinal cells) with their somas immediately dorsolateral to the rostral magnocellular nucleus and its capsule, also projected to the spinal cord, as did clusters of neurons in the periaqueductal grey matter. Terminals of corticorubral axons originating from ipsilateral primary motor area 4 (the densest projection), the supplementary motor area, cingulate area 24, area 8, and posterior parietal area 5, were each mapped in the parvocellular red nucleus. Only area 4 projected to the magnocellular red nucleus, and this projection as small. DL-spinal neurons had no cortical input. The somatotopic organization of rubral connections was examined only in (a) the corticorubral input from motor area 4, and (b) the rubrospinal and DL-spinal projections. These connections and their somatotopic alignment, were mapped in a 3-dimensional reconstruction of the red nucleus. PMID- 10867654 TI - Geometry of rubrospinal, rubroolivary, and local circuit neurons in the macaque red nucleus. AB - The primate red nucleus consists of three main neuron subpopulations, namely, rubrospinal neurons in the magnocellular nucleus, rubroolivary cells in the parvocellular nucleus, and local circuit neurons in both subnuclei: Each subpopulation has unique cerebellar and neocortical inputs. The structural framework for the interactions of these rubral subpopulations remains poorly defined and was the focus of this study in six macaques. Somata of rubrospinal neurons, dorsolateral-spinal (DL-spinal) neurons, as defined in the accompanying paper (Burman et al. [2000] J. Comp. Neurol., this issue), and rubroolivary neurons were labeled retrogradely first with Fast Blue injected either into the cervical spinal cord or the inferior olive. The soma/dendrite profiles of selected cells (53 rubrospinal, 19 DL-spinal, and 17 rubroolivary cells) were visualized by the intracellular injection of Lucifer Yellow/biocytin in fixed slices (400 microm thick) of midbrain. The descriptive statistics of the somata and the dendritic arborization of each rubral neuron type were established. Projection neuron subpopulations had similar but differentiable soma/dendrite profiles, with four to six slender, spine-bearing dendritic trees radiating out approximately 400 microm from the soma. Twelve presumed interneurons, all in the parvocellular nucleus, differed from projection neurons in that they had smaller somata and many slender, spine-bearing segments that constituted the multibranching dendrite profile that radiated out approximately 250 microm from the soma. A tentative model of the macaque rubral microcircuitry was developed, and its functional implications were explored. It incorporated 1) the known topography of the nucleus and its connections, 2) our data specifying the soma/dendrite morphology of the three main rubral neuron types, and 3) the ultrastructure reported by other laboratories of intrarubral synaptic connections. PMID- 10867655 TI - Projection pattern and target selection of Clione limacina motoneurons sprouting within an intact environment. AB - In the pteropod mollusc Clione limacina, two groups of locomotor motoneurons, located in the pedal ganglion, innervate the dorsal and ventral muscle layers of the ipsilateral wing through the wing nerve. Separate branches of this nerve go either only to the dorsal muscle layer or only to the ventral one. In the present study, growth of novel neurites of the wing motoneurons was induced by cutting the wing nerve. In addition, all other peripheral nerves and connectives of the pedal ganglion were cut, except for the pedal commissure to the contralateral pedal ganglion. Thus, the neurites were allowed to grow only towards the contralateral pedal ganglion. We have found that the novel neurites, entering the contralateral pedal ganglion, were capable of growing everywhere inside the central nervous system (CNS) and into any peripheral nerve. However, they preferred the wing nerve. This finding suggests that the preference is caused by the guiding cues in the wing nerve or the attractive influence of the wing muscles. Because the contralateral pedal ganglion and nerves were left intact, the growth direction of the new neurites could be determined only by factors permanently existing in the CNS, rather than induced by nerve injury or muscle denervation. Within the wing nerve, the neurites could not discriminate between the nerve branches going to the dorsal and ventral muscle layers. They formed synapses on muscles of both layers, despite the fact that the muscles were innervated by their own motoneurons. PMID- 10867656 TI - Cone mosaic development in the goldfish retina is independent of rod neurogenesis and differentiation. AB - The goldfish retina displays a characteristic arrangement of cone photoreceptors that develop in a stereotyped sequence according to spectral phenotype. It has been suggested that the earliest differentiating photoreceptor in the teleost, the rod photoreceptor, might play an instructive role in development of the cone mosaic. This hypothesis was tested, first by examining the expression pattern of a cone subtype-specific marker with respect to that of rod opsin, and then by killing the cells that generate rods and examining the cone mosaic that formed in the absence of new rods. We find that, although there is potential for interactions between developing cones and immediately postmitotic rods, a role for such interactions in cone mosaic pattern formation is not likely. PMID- 10867657 TI - Bottlebrush dendritic endings and large dendritic fields: motion-detecting neurons in the mammalian tectum. AB - The widefield vertical neurons of the lower stratum griseum superficiale (SGS3) and upper stratum opticum (SO) of the superior colliculus provide an extrageniculate route for visual information to reach the pulvinar. Previous physiological studies indicate that SGS3/SO neurons have large receptive fields and respond to small moving stimuli. We sought to better characterize the dendritic morphology of SGS3/SO neurons with intracellular filling in slice preparations of the ground squirrel superior colliculus. We found that dendrites of widefield vertical cells end in monostratified arrays of spiny terminal specializations called "bottlebrush" dendritic endings. Two major subtypes of neurons are described. Type I neurons have somata restricted to the SGS3 and bottlebrush endings in the most superficial sublayer of the SGS. Type II neurons are found at the base of the SGS and in the upper SO, and have bottlebrush endings arrayed within the middle sublayers of the SGS. Bottlebrush endings may sample and integrate laminated afferents to the superior colliculus, and cellular subtypes may underlie multiple information streams within the tectopulvinar pathway. A similar dendritic morphology and projection pattern can be found in cells of the avian optic tectum that project upon the nucleus rotundus, a thalamic nucleus homologous to the mammalian caudal/inferior pulvinar. Because motion processing is a dominant feature of the avian tectorotundal pathway, the current results suggest that both dendritic morphology and motion processing are conserved features of widefield vertical cells in the tectopulvinar pathway of vertebrates. PMID- 10867658 TI - Chemical characterization of leptin-activated neurons in the rat brain. AB - Leptin has profound effects on food intake, body weight, and neuroendocrine status. The lack of leptin results in hormonal and metabolic alterations and a dramatic increase in body weight. Leptin acts in the brain, especially in the hypothalamus; however, the central nervous system sites that respond to leptin have not been examined comprehensively. In this study, we explored systematically the distribution of leptin-activated neurons throughout the rat brain. Furthermore, we investigated the chemical identity of subsets of these leptin activated cells. Fos-like immunoreactivity (Fos-IR) was investigated in the rat brain after two different doses of leptin (1.0 mg/kg and 5.0 mg/kg) at 2 hours and 6 hours after injections. The induction of Fos-IR was observed in hypothalamic nuclei, including the paraventricular nucleus (PVH), the retrochiasmatic area (RCA), the ventromedial nucleus (VMH), the dorsomedial nucleus (DMH), the arcuate nucleus (Arc), and the ventral premammillary nucleus (PMV). In addition, leptin-induced Fos-IR was found in several nuclei of the brainstem, including the superior lateral and external lateral subdivisions of the parabrachial nucleus (slPB and elPB, respectively), the supragenual nucleus, and the nucleus of the solitary tract (NTS). By using double-labeling immunohistochemistry or immunohistochemistry coupled with in situ hybridization, leptin-activated neurons were found that contained cocaine- and amphetamine regulated transcript mRNA in several hypothalamic nuclei, including the RCA, Arc, DMH, and PMV. In the Arc and DMH, leptin-induced Fos-IR was observed in neurons that expressed neurotensin mRNA. Dynorphin neurons in the VMH and in the Arc also expressed Fos-IR. In the brainstem, we found that cholecystokinin neurons in the slPB and glucagon-like peptide-1 neurons in the NTS were activated by leptin. We also investigated the coexpression of Fos-IR and the long form of the leptin receptor (OBRb) mRNA. We found double-labeled neurons surrounding the median eminence and in the RCA, Arc, VMH, DMH, and PMV. However, in brainstem sites, very little OBRb mRNA was found; thus, there were very few double-labeled cells. These results suggest that leptin stimulates brain pathways containing neuropeptides that are involved in the regulation of energy balance, autonomic homeostasis, and neuroendocrine status. PMID- 10867659 TI - Connections between the medial temporal cortex and the CA1 subfield of the hippocampal formation in the Japanese monkey (Macaca fuscata). AB - The connections between the medial temporal cortical areas and CA1 of the hippocampus were examined in the Japanese monkey (Macaca fuscata) by means of retrograde and anterograde tract-tracing methods with wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) and fluorescent dyes (Fast Blue and Diamidino Yellow). The posterior parahippocampal (areas TF1, TF2, and TH), perirhinal (areas 35 and 36), and ventral inferotemporal areas (areas TEav and TEpv) were reciprocally connected with CA1. Projection fibers from CA1 to the medial temporal cortical areas originated in the pyramidal cell layer, whereas those from the medial temporal cortical areas to CA1 terminated in the molecular layer. Each of these cortical areas was reciprocally connected with the entire rostrocaudal extent of CA1. However, the intensity of the connections varied along the rostrocaudal axis of CA1: areas TH and TF2 were connected most markedly with the anterior and middle parts of CA1, respectively. Areas TF, 35, 36, TEav, and TEpv were connected predominantly with the posterior part of CA1. In the coronal plane of CA1, labeled cells were located in proximal CA1 (i. e., near the prosubiculum), but not in distal CA1 (i.e., near CA2). The medial temporal cortical areas in direct reciprocal connection with CA1 were presumed to be involved in the memory system, especially in the system for declarative memory. PMID- 10867660 TI - Organization of the human paraventricular hypothalamic nucleus. AB - The cyto- and chemoarchitecture of the human paraventricular hypothalamic nucleus (Pa) was studied with the aid of three-dimensional computer reconstruction. The adult human Pa is a vertically elongated structure that abuts the wall of the third ventricle (3V) medially and is indented dorsolaterally by the descending fornix. Chemoarchitecture revealed the following five subnuclei in the human Pa. The most prominent of these is the magnocellular subnucleus (PaM) occupying the ventrolateral quadrant of the Pa and comprised of a concentration of large arginin-vasopressin (AVP)- and acetylcholinesterase (AChE)-positive cells, and small calbindin (Cb)-positive neurons. Rostrally, the PaM is succeeded by the small anterior parvicellular subnucleus (PaAP), which contains small AChE-, AVP- and tyrosin hydroxylase (TH)-positive cells. Dorsal to the PaM is found the dorsal subnucleus (PaD), containing large spindle-shaped TH-, oxytocin (OXY)-, and AChE-positive cells, as well as a population of small Cb-positive neurons. Abutting the wall of the 3V and medial to PaM and PaD is the parvicellular subnucleus (PaP). The PaP contains small cells immunoreactive for corticotropin releasing factor (CRF), neuromedin K receptor (NK3), and nonphosphorylated neurofilament protein (SMI32). The posterior subnucleus (PaPo) is situated posterior to the descending column of the fornix; it replaces all above-mentioned subdivisions caudally, and is a chemoarchitectonic amalgam that includes dispersed large AChE-, OXY-, AVP- and TH-positive cells, as well as small NK3-, CRF-, SMI32- and Cb-immunoreactive neurons. The present findings suggest that the human PaM and PaD are homologues to the magnocellular subnuclei of the rat Pa, whereas the human PaP and PaPo correspond to the rat medial parvicellular and posterior subnuclei, respectively. PMID- 10867661 TI - Autoradiographic localization of (125)I[Tyr(14)]orphanin FQ/nociceptin and (125)I[Tyr(10)]orphanin FQ/nociceptin(1-11) binding sites in rat brain. AB - The endogenous ligand for the orphan opioid receptor, orphanin FQ/nociceptin (OFQ), has recently been characterized. The OFQ peptide sequence contains paired basic amino acids, suggesting the possibility of posttranslational processing to a peptide containing the first 11 amino acids of the OFQ peptide. This peptide has been reported in the brain and it has a unique pharmacology. In the present study, we compared the autoradiographic distribution of (125)I[Tyr(14)]OFQ and (125)I[Tyr(10)]OFQ(1-11) in coronal rat brain sections. Nonspecific binding was defined with unlabeled OFQ or OFQ(1-11), respectively. Both radioligands demonstrated high levels of specific binding (>95% of total binding), with no appreciable binding in white matter areas with either ligand. (125)I[Tyr(14)]OFQ binding was widely distributed throughout the rat brain. In contrast, (125)I[Tyr(10)]OFQ(1-11) binding was more restricted. The highest (125)I[Tyr(14)]OFQ binding levels measured in this study were found in the locus coeruleus, an area which contained very low (125)I[Tyr(10)]OFQ(1-11) binding. Both ligands labeled the cortex, hippocampus and amygdala. In the thalamus, (125)I[Tyr(14)]OFQ binding was prominent in most nuclei, whereas (125)I[Tyr(10)]OFQ(1-11) binding was restricted to the midline thalamus. (125)I[Tyr(14)]OFQ binding was heavy in the suprachiasmatic hypothalamus, and moderate in other hypothalamic nuclei. (125)I[Tyr(10)]OFQ(1-11) binding in the hypothalamus, however, was present mainly in the ventromedial hypothalamic nucleus. Lower binding levels of both ligands were found in the caudate putamen. The distinct autoradiographic patterns of these two ligands are consistent with different binding sites, which might help explain their different functional activities. PMID- 10867662 TI - Presynaptic NMDA receptor subunit immunoreactivity in GABAergic terminals in rat brain. AB - N-methyl-D-aspartate (NMDA) receptors are commonly found post-synaptically; they mediate fast excitatory neurotransmission in the central nervous system. In this study, we provide immunocytochemical data supporting the existence of presynaptic NMDA receptors in GABAergic terminals using polyclonal antisera raised against the C-terminus of the NMDAR1 subunit. At the light microscope level, rich plexuses of NMDAR1-positive varicose fibers were found in various nuclei in the basal forebrain (bed nucleus of stria terminalis, septum, parastrial nucleus, vascular organ of the lamina terminalis), thalamus (paraventricular nucleus, midline nuclei), and hypothalamus (parvocellular paraventricular nucleus, arcuate nucleus, preoptic nucleus, suprachiasmatic nucleus). In the brainstem, labeled fibers were much less abundant and were confined to the ventral tegmental area, periaqueductal gray, parabrachial nucleus, and locus coeruleus. At the electron microscope level, NMDAR1-immunoreactive terminals examined in the bed nucleus of stria terminalis, parvocellular paraventricular hypothalamic nucleus, and arcuate nucleus formed symmetric synapses, contained darkly stained large dense-core vesicles, and displayed gamma-aminobutyric acid (GABA) immunoreactivity. Terminals with similar ultrastructural features were found in the paraventricular thalamic nucleus. These findings demonstrate the existence of NMDAR1 subunit immunoreactivity in subsets of GABAergic terminals, which raises questions about the potential roles and mechanisms of activation of presynaptic NMDA heteroreceptors in the rat central nervous system. The pattern of distribution and ultrastructural features of these boutons suggest that they may arise from local GABAergic projections interconnecting a group of brain structures mediating stress responses and/or other endocrine, autonomic, and limbic functions. PMID- 10867663 TI - Expression of "suppressor of cytokine signalling" (SOCS) genes in the developing and adult mouse nervous system. AB - Growth factor and cytokine signalling in the developing nervous system has multiple effects, ranging from cell differentiation and cell survival to modulation of cell phenotype. Molecules that can regulate growth factor signalling pathways will therefore be of importance in determining the cellular response to factor stimulation. Members of a recently described gene family, the suppressor of cytokine signalling (SOCS) family, can regulate signalling events downstream of predominantly cytokine stimulation and may have important roles in the nervous system. We have examined the temporal and spatial expression of SOCS 1, SOCS-2, and SOCS-3 in the developing and adult nervous system by use of Northern analysis and in situ hybridisation. All three genes were expressed in the brain, with maximal expression from embryonic day 14 to postnatal day 8 and declining thereafter, with SOCS-2 being the most highly expressed. In situ hybridisation analysis showed that SOCS-1 and SOCS-3 had a low and widespread pattern of expression, whereas SOCS-2 expression was higher and tightly regulated. Its expression pattern indicated that SOCS-2 was expressed exclusively in neurons and that it was switched on developmentally at the time of neuronal differentiation. PMID- 10867664 TI - Color Doppler sonographic appearance of renal perforating vessels in subjects with normal and impaired renal function. AB - PURPOSE: The purpose of this study was to assess the prevalence and color Doppler sonographic characteristics of perforating vessels-small arteries and veins connecting the intrarenal vasculature with the capsular plexus-in healthy subjects, in hypertensive patients, and in patients with renal failure due to hypertensive nephroangiosclerosis or ischemic nephropathy. METHODS: Fifteen healthy subjects 24-34 years old, 15 healthy subjects 68-80 years old, 25 hypertensive patients, 25 patients with hypertension and chronic renal failure (15 mild, 10 severe), and 12 patients with hypertension and chronic renal failure and acute renal insufficiency due to ischemic nephropathy underwent color Doppler sonography of both kidneys. RESULTS: The few perforating arteries in healthy and hypertensive patients had various resistance indices and flow toward the capsule. Perforating veins in these patients were much more common than perforating arteries. Perforating arteries with a lower mean resistance index than the mean interlobar resistance index and flow toward the capsule were detected in 76% of kidneys in the patients with mild chronic renal failure and in 20% of those in patients with severe chronic renal failure. Only a few perforating veins were seen in patients with chronic renal failure. In 64% of the kidneys with renal artery stenosis detected in the patients with chronic renal failure complicated by acute renal insufficiency, there were perforating arteries with flow toward the kidney and a mean resistance index higher than the mean interlobar resistance index. CONCLUSIONS: Perforating vessels are recognizable using color Doppler sonography both in healthy subjects and in patients with renal failure. The prevalence and flow characteristics of perforating vessels differ between healthy subjects, patients with mild and with severe chronic renal failure, and patients with chronic renal failure complicated by acute renal insufficiency caused by renal artery stenosis. PMID- 10867665 TI - Abdominal sonographic study of autosomal dominant polycystic kidney disease. AB - PURPOSE: The purpose of this study was to determine whether kidney size in patients who have autosomal dominant polycystic kidney disease (ADPKD) is related to renal function, hypertension, or extrarenal manifestations of the disease and to sonographically evaluate the abdominal manifestations of ADPKD. METHODS: Between 1994 and 1998, 400 individuals from 85 families with a history of ADPKD were examined. There were 213 persons with ADPKD and 187 unaffected family members; there were 182 males and 218 females, 1-82 years old (mean, 39.3 years). We obtained a complete medical history, performed a physical examination, measured the arterial blood pressure and serum creatinine levels, and performed abdominal sonography on each subject. The sonographic features that were studied were renal length and the presence and number of cysts on the kidneys, liver, and pancreas. RESULTS: There was a relationship between kidney size and age (p < 0.05), kidney size and renal function (p < 0.001), and kidney size and hypertension (p < 0.001). The overall prevalence of hepatic cysts in patients with ADPKD was 67%, and the prevalence increased with age. The presence of hepatic cysts was related to the severity of renal disease. Females had more severe polycystic liver disease, and massive polycystic liver disease (ie, hepatomegaly with innumerable cysts) was seen only in females. The prevalence of pancreatic cysts in the 187 persons in whom the pancreas was well evaluated sonographically was 5%. CONCLUSIONS: Kidney size in patients with ADPKD is related to renal function, hypertension, and extrarenal involvement and can be used to predict the outcome of the disease. Hepatic cysts are very common in patients with ADPKD and are related to age and renal function; pancreatic cysts are infrequent in these patients. PMID- 10867666 TI - Sonographically guided core-needle biopsy in the diagnosis of superficial lymphadenopathy. AB - PURPOSE: This study was designed to evaluate the efficacy of ultrasound-guided percutaneous core-needle biopsy for establishing histopathologic diagnoses of palpable enlarged lymph nodes. METHODS: Thirty patients without a history of malignancy or recent infection underwent ultrasound-guided core-needle lymph node biopsies. Only patients with enlarged lymph nodes ranging from 1.5 to 3.5 cm in greatest diameter that had been present for more than 2 months were included in this study. Two cores from each lymph node were obtained by a freehand core needle biopsy technique using a 7-MHz ultrasound transducer and an automatic spring-loaded biopsy gun with an 18-gauge cutting needle. RESULTS: The histologic diagnoses were conclusive in 24 cases (80%) and inconclusive in 6 cases (20%). In the 24 conclusive cases, the lymph node enlargement was due to benign causes in 12 cases and malignancy in 12 cases. All specimens in the conclusive cases were sufficient for histologic diagnosis, but there was insufficient material for a diagnosis in 3 of the inconclusive cases. No complications were seen in this series. CONCLUSIONS: Ultrasound-guided core-needle biopsy of enlarged lymph nodes is a safe, minimally invasive alternative to surgical biopsy, enabling a histologic diagnosis for treatment planning in most cases. PMID- 10867667 TI - Duplex Doppler sonography of the flaccid penis: potential role in the evaluation of impotence. AB - PURPOSE: Duplex Doppler sonography of the cavernosal arteries of the penis with intracavernous injection (ICI) of vasoactive agents has been widely used to evaluate arterial insufficiency in impotence. Our goal was to assess the potential value of peak systolic velocity (PSV) measurements on the flaccid penis in the diagnosis of arteriogenic impotence. METHODS: Forty-four men underwent duplex Doppler sonography with PSV measurements before and after ICI of prostaglandin E(1). Three different cutoff values for lowest normal PSV before injection-5 cm/second, 10 cm/second, and 15 cm/second-were tested. RESULTS: Thirteen patients had arteriogenic insufficiency based on post-ICI duplex sonography and clinical response. Results for our different cutoff PSV values of 5 cm/second, 10 cm/second, and 15 cm/second in diagnosing arteriogenic impotence were, respectively: sensitivity 29%, 96%, and 100%; specificity 100%, 92%, and 23%; negative predictive value 80%, 92%, and 100%; positive predictive value 100%, 81%, and 41%; and overall accuracy 79%, 93%, and 44%. In the flaccid state, there was a significant difference in mean PSV between the "normal" group (12.6 +/- 0.9 cm/second) and the arteriogenic impotence group (7.7 +/- 1.1 cm/second). Twenty-nine patients with a bilateral PSV of 10 cm/second or less before ICI had a normal clinical response. CONCLUSIONS: A cutoff PSV value of 10 cm/second in the flaccid state had the best accuracy in predicting arterial insufficiency. Duplex Doppler sonography is proposed as the initial test to evaluate the penile arterial supply and to determine whether patients are good candidates for therapy with ICI. PMID- 10867668 TI - Assessment of condoms as probe covers for transvaginal sonography. AB - PURPOSE: This prospective study assessed the incidence of transvaginal probe contamination and breakage of condoms used to cover those probes during transvaginal sonography. METHODS: Over a 9-month period, 214 women underwent transvaginal sonography with probes that had been coated with gel and then covered with a latex condom. Condom defects were detected after the scans by inspection, by adding hydrogen peroxide, and by filling the condoms with 500 ml of water. After the condoms were removed, the probe was either wiped with a dry tissue (during the first 18 weeks of the study) or wiped first with a dry tissue and then with a 70% isopropyl alcohol wipe. Probe head contamination was assessed by periodic swab sampling and culturing for bacteria and herpes simplex virus. Samples of the sonographic gel also were tested for bacterial contamination at approximately weekly intervals. RESULTS: A total of 217 condoms were used, 3 of which broke and were discarded while being applied to the probe. Two of the 214 condoms used (0.9%) were found upon visual inspection to have perforations. None of the other 212 condoms leaked upon being filled with water; none of the 204 condoms tested with hydrogen peroxide showed bubbles. Only 1 of the 46 probe swab samples was positive for bacteria (Acinetobacter species); none of the 26 probe swab samples cultured for viruses or the 25 gel samples cultured for bacteria were positive. CONCLUSIONS: Condoms used to cover transvaginal probes showed a low rate of perforation. Disinfection of the probe with isopropyl alcohol wipes further reduced the risk of contamination. PMID- 10867669 TI - Sonographic findings in masseter-muscle metastases. AB - Symptomatic metastases in skeletal muscles are rare, and involvement of the masseter muscle is extremely rare, with only 3 reported cases. We report 2 additional cases of masseter-muscle metastases. Sonographically, the masseter muscle metastases appeared as ill-defined, hypoechoic, heterogeneous lesions aligned with the long axis of the muscle. Fine-needle aspiration was confirmatory. The combination of sonography and fine-needle aspiration is the ideal method for diagnosing masseteric metastases. PMID- 10867670 TI - Retroperitoneal duodenal perforation during endoscopic sphincterotomy: sonographic findings. AB - We present the sonographic findings in 5 cases of retroperitoneal duodenal perforation during endoscopic sphincterotomy. In each case, sonography showed hyperechoic areas associated with shadowing and ring-down artifacts between the liver and the right kidney correlating with the retroperitoneal air seen on plain x-ray films. This brightly echogenic area with shadowing surrounded and obscured the kidney. In 4 patients, resolution was documented on follow-up sonographic examinations in agreement with other radiologic findings; the other patient died. PMID- 10867671 TI - Prenatal sonographic detection of congenital hepatic cyst in third trimester after normal second-trimester sonographic examination. AB - A solitary unilocular hepatic cyst (SUHC) is a rare prenatal or neonatal finding. There are few reports of the prenatal detection of SUHC, and the progression of SUHC in utero is unknown. We present a proven case of SUHC in a fetus detected on a 34-week ultrasound examination following a normal 19-week examination. The cyst was inseparable from the liver and caused some flattening of the liver edge. Prenatal detection of an SUHC inseparable from the liver and appearing in the late second or third trimester should suggest a congenital hepatic cyst. PMID- 10867672 TI - Sonographic appearance of a retained surgical sponge in the neck. AB - We report the sonographic appearance and clinical course of a retained surgical sponge in the neck beginning 6 months after a partial thyroidectomy. Sonograms showed a subcutaneous curvilinear hyperechoic interface with marked acoustic shadowing obscuring the left side of the neck. Three months later, a fistulous tract could be seen. Surgical exploration revealed a florid foreign tissue reaction due to a retained surgical sponge. Early diagnosis of retained sponges is important to enable expeditious removal before complications develop. PMID- 10867673 TI - Meckel's diverticulum mimicking infantile colic: sonographic detection. AB - We report a case of Meckel's diverticulum in a 6-month-old girl who presented with a 5-month history of chronic screaming but no symptoms or signs of intestinal obstruction. Infantile colic was the presumptive diagnosis. Abdominal sonography at 6 months of age demonstrated an abdominal mass with an anechoic center and a double-layered wall, surrounded by bowel loops. Abdominal CT and barium enema x-ray studies demonstrated nonspecific findings of a cystic mass with compression of adjacent bowel loops. Histologic examination of the resected mass revealed a Meckel's diverticulum with a perforation sealed off by the neighboring bowel and mesentery to form an inflammatory mass. PMID- 10867674 TI - A strategy for modelling the effect of a continuous covariate in medicine and epidemiology. AB - Low-dimensional parametric models are well understood, straightforward to communicate to other workers, have very smooth curves and may easily be checked for consistency with background scientific knowledge or understanding. They should therefore be ideal tools with which to represent smooth relationships between a continuous predictor and an outcome variable in medicine and epidemiology. Unfortunately, a seriously restricted set of such models is used routinely in practical data analysis - typically, linear, quadratic or occasionally cubic polynomials, or sometimes a power or logarithmic transformation of a covariate. Since their flexibility is limited, it is not surprising that the fit of such models is often poor. Royston and Altman's recent work on fractional polynomials has extended the range of available functions. It is clearly crucial that the chosen final model fits the data well. Achieving a good fit with minimal restriction on the functional form has been the motivation behind the major recent research effort on non-parametric curve-fitting techniques. Here I propose that one such model, a (possibly over-fitted) cubic smoothing spline, may be used to define a suitable reference curve against which the fit of a parametric model may be checked. I suggest a significance test for the purpose and examine its type I error and power in a small simulation study. Several families of parametric models, including some with sigmoid curves, are considered. Their suitability in fitting regression relationships found in several real data sets is investigated. With all the example data sets, a simple parametric model can be found which fits the data approximately as well as a cubic smoothing spline, but without the latter's tendency towards artefacts in the fitted curve. PMID- 10867675 TI - Estimating treatment effects in randomized clinical trials in the presence of non compliance. AB - In clinical trials where patients are randomized between two treatment arms, not all patients comply with the treatment they were randomly assigned to. The reasons for (non)compliance may be associated with the outcome variable and thereby act as confounders. The standard way of analysing such trials is by the 'intention-to-treat' principle, which allows the use of permutation tests. Conclusions drawn from such tests do not depend on untested assumptions such as absence of confounding. However, this approach may yield biased estimators for the causal effects of treatments. We consider the estimation of such effects for clinical trials where non-compliers can be considered to have switched to the other trial arm. The most important example of this is the placebo-controlled clinical trial where no substantial placebo effects are anticipated. We consider the situation where the relationship between compliance, and thus treatment received, and outcome is influenced by unobserved confounders. The residual of the regression of the actual treatment indicator variable on the randomization arm indicator variable is shown to 'intercept' the effect of such confounders. Inclusion of this residual in a multivariate analysis, in conjunction with the treatment indicator variable, should thus adjust for confounding. Examples are given. In those examples, the results are similar to those obtained by more complex methods. PMID- 10867676 TI - Modelling the cumulative risk for a false-positive under repeated screening events. AB - Screening examinations are widely utilized in detecting the presence of medical disorders, for instance, screening mammograms and clinical breast examinations for detection of breast cancer. Such procedures are invaluable in enabling early treatment but produce the possibilities of false-positive and false-negative diagnoses. Focusing on false-positive results, with increasing number of screening events, it is clear that the risk of a false-positive increases. The objective of this paper is to quantify the cumulative risk associated with repeated screening. We provide a very general framework within which to investigate this risk, both at the population and the individual level. The latter allows incorporation of evolving patient medical history to permit individualized assessment of risk. We model cumulative risk in terms of the number of screening events until first false-positive. We develop models which are essentially familiar actuarial models for life table data adding a Cox regression to enable individual level modelling. Because it offers several advantages, we employ a Bayesian inference framework and apply our modelling to the analysis of 9773 screening mammograms collected from 2227 women at an HMO serving nearly 300000 adults in and around Boston, MA. PMID- 10867677 TI - Rater agreement and the generalized Rudas-Clogg-Lindsay index of fit. AB - Rudas, Clogg and Lindsay proposed a new index of fit for contingency table analysis. Using the two-component mixture, where the first component with weight (1-w) represents the model to be tested and the second component with weight w is unstructured, the RCL index of lack of fit was defined to be the smallest mixing weight w(*) being compatible with the two-component mixture to be saturated. This index of fit, which is not sensitive to sample size, is applied to the problem of assessing agreement between two raters whereby three hypotheses (pure agreement, quasi-independence, independence) are considered. As quasi-independence comprises the two other hypotheses, a natural generalization of the RCL index of fit results from assuming the model itself to be composed of two submodels (pure agreement, independence) while the third component remains unstructured. Two examples demonstrate the application of this generalized RCL index of fit, with the first 3x3 contingency table having no empty cells, and the second 4x4 table having five of them. In both cases estimating the parameters and determining w(*) was possible without problems within the linear logistic framework for latent class analysis. A further analysis of the 4x4 table assumes the model to be the two-class latent class model that certainly does not belong to the family of standard models for contingency table analysis. Thus, in contrast to the recommendations given originally, the following conclusions seem to be justified: (i) the number of components representing the model may exceed one (generalized RCL index); (ii) the application of the original RCL index of fit may be extended to more complex models; and (iii) empty cells bear no problems so that this approach may be recommended in the case of both large and small sample sizes. PMID- 10867678 TI - Using an antedependence test to analyse post-operative pain measurements. AB - Post-operative pain is frequently measured by a visual analogue scale (VAS). The most common methods for analysing such data are the so-called time-by-time method and repeated measures analysis of variance. In the first, due to correlation between time points, successive tests are not independent and cannot measure the treatment effects over time. In the second, a rather unlikely covariance structure has to be assumed. In a simulation study we have shown that the power of the antedependence test is nearly the same as that of repeated measures of analysis of variance and it does not suffer from some of the latter's shortcomings. We also present an analysis of real data using the antedependence test. PMID- 10867679 TI - The generalization of the odds ratio, risk ratio and risk difference to r x k tables. AB - Familiar measures of association for 2 x 2 tables are the odds ratio, the risk ratio and the risk difference. Analagous measures of outcome-exposure association are desirable when there are several degrees of severity of both exposure and disease outcome. One such measure (alpha), which we label the general odds ratio (OR(G)), was proposed by Agresti. Convenient methods are given for calculation of both standard error and 95 per cent confidence intervals for OR(G). Other approaches to generalizing the odds ratio entail fitting statistical models which might not fit the data, and cannot handle some zero frequencies. We propose a generalization of the risk ratio (RR(G)) following the statistical approaches of Agresti, Goodman and Kruskal. A method of calculating the standard error and 95 per cent confidence interval for RR(G) is provided. A known statistic, Somers' d, fulfils the characteristics necessary for a generalized risk difference (RD(G)). These measures have straightforward interpretations, are easily computed, are at least as precise as other methods and do not require fitting statistical models to the data. We also examine the pooling of such measures as in, for example, meta-analysis. PMID- 10867680 TI - Comparing the performance of two indices for spatial model selection: application to two mortality data. AB - The statistical analysis of spatially correlated data has become an important scientific research topic lately. The analysis of the mortality or morbidity rates observed at different areas may help to decide if people living in certain locations are considered at higher risk than others. Once the statistical model for the data of interest has been chosen, further effort can be devoted to identifying the areas under higher risks. Many scientists, including statisticians, have tried the conditional autoregressive (CAR) model to describe the spatial autocorrelation among the observed data. This model has greater smoothing effect than the exchangeable models, such as the Poisson gamma model for spatial data. This paper focuses on comparing the two types of models using the index LG, the ratio of local to global variability. Two applications, Taiwan asthma mortality and Scotland lip cancer, are considered and the use of LG is illustrated. The estimated values for both data sets are small, implying a Poisson gamma model may be favoured over the CAR model. We discuss the implications for the two applications respectively. To evaluate the performance of the index LG, we also compute the Bayes factor, a Bayesian model selection criterion, to see which model is preferred for the two applications and simulation data. To derive the value of LG, we estimate its posterior mode based on samples derived from the BUGS program, while for Bayes factor we use the double Laplace-Metropolis method, Schwarz criterion, and a modified harmonic mean for approximations. The results of LG and Bayes factor are consistent. We conclude that LG is fairly accurate as an index for selection between Poisson gamma and CAR model. When easy and fast computation is of concern, we recommend using LG as the first and less costly index. PMID- 10867681 TI - Application of log-linear models to malaria patients in Thailand. AB - Malaria is a common infectious disease in many tropical countries, including Thailand. The country is located geographically in a tropical zone and the transmission of malaria is particularly common in some regions, for instance in Tak province. The objective of this study is to identify risk factors causing malaria in Tak province in the rainy season by using log-linear models. Tests of independence are used (chi-square and Cramer's V-value tests) to find out the relationships between any two variables. In addition two- and three-dimensional log-linear models are used to obtain estimated parameters and expected frequencies for these models. Amongst the models fitted, the best are chosen based on the analysis of deviance. The results of this study show that most observed variables are significantly related with p-values<0.05. Causes of migration and reasons for staying overnight are highly related to personal variables. Thus, it can be concluded that two of the risk factors for malaria are causes of migration and reasons for staying overnight. Knowledge of prevention is also related to personal variables. Therefore, knowledge of prevention was concluded to be a risk factor affecting prevalence of malaria. For each set of three variables, the best model shows interaction terms of variables that have a relationship but there are no interactions of three effects in these best models. PMID- 10867682 TI - Mass spectrometry applied to the analysis of estrogens in the environment. AB - Environmental analytical chemistry has recently changed focus from analysis of non-polar, persistent contaminants (e.g. polychlorinated biphenyls and dioxins) to more polar and labile compounds that interfere with biological processes. For example, natural and synthetic estrogens and their metabolites have been detected in sewage treatment plant effluents at nanogram/liter concentrations that are similar to those at which both total sex reversal and intersex (containing both testes and ova) is induced in fish exposed to these compounds in laboratory experiments. The development of techniques for the analysis of natural and synthetic estrogens in biological fluids (i.e. serum and urine) has been a priority in the biomedical field. However, the recent recognition that estrogen hormones are contaminants in the environment that may contribute to endocrine disruption has focused attention on the need for highly sensitive and specific techniques that are applicable for trace analysis in complex environmental matrices. Three optimized mass spectrometric protocols have been developed for the determination and quantitation of steroid hormones in environmental matrices using gas chromatography/tandem mass spectrometry (GC/MS/MS), liquid chromatography/mass spectrometry selected ion monitoring, (LC/MS - SIM) and liquid chromatography/tandem mass spectrometry (LC/MS/MS). The advantages and disadvantages of each method are presented. PMID- 10867683 TI - Characterization of intermolecular beta-sheet peptides by mass spectrometry and hydrogen isotope exchange. AB - The self-assembly of beta-sheet peptide domains resulting in the formation of fibrillar aggregates (amyloids) is a feature of various neurodegenerative disorders. In order to evaluate mass spectrometric methods for the characterization of intermolecular beta-sheet structures the hydrogen/deuterium exchange behaviour of model peptides DPKGDPKG-(VT)(n)-GKGDPKPD-amide (n = 3,4,5,6,7,8), (VT)(n)-peptides, composed of a central beta-sheet-forming domain and N- and C-terminal nonstructured octapeptide sequences, was measured by electrospray ionization mass spectrometry (ESI-MS) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The kinetic analysis of the hydrogen/deuterium exchange (HX) shows that intermolecular beta-sheet structures contain slowly exchanging protons (k 100% of recommended daily intake) in patients with cystic fibrosis related diabetes mellitus. The importance of calories from fat is emphasized, with no restriction on total carbohydrate intake. Insulin intake mirrors carbohydrate intake. Routine dietary therapy is straightforward, but challenges occur due to both complications of cystic fibrosis and advancing disease. If a patient with cystic fibrosis related diabetes mellitus is malnourished, overnight enteral tube feeding is often used, with an adjusted insulin regimen. There is a great need for both physiological and outcomes research to provide sound scientific evidence for the dietary treatment of cystic fibrosis related diabetes mellitus. PMID- 10867727 TI - Nutritional status of mechanically ventilated critically ill patients: comparison of different types of nutritional support. AB - Malnutrition is a common problem in hospitalized patients. Early assessment of nutritional status may help in identifying patients for whom nutritional interventions are needed. The purpose of this study was to assess and compare the nutritional status of mechanically ventilated critically ill patients who were receiving nutritional support. Forty-nine patients were divided into either enteral nutrition, total parenteral nutrition or combined (enteral plus total parenteral nutrition) groups. Anthropometric and biochemical measurements, and medical status (APACHE II score) were assessed at the 1st day and 14th day of admission in the intensive care unit (ICU) of Taichung Veteran General Hospital. The length of ventilator dependency was significantly positively correlated with calorie and carbohydrate intake in the pooled group. Patients receiving enteral and combined nutrition showed significantly lower anthropometric measurements at the 14th day after admission. Patients in all groups had abnormal mean biochemical values at the 1st day of admission. Subjects in the combined group showed a significant increase in prealbumin and the Maastricht Index levels after 14 days. Patients in all three groups were malnourished when admitted to the ICU. Patients showed a slightly improvement of nutritional status after receiving nutritional support for 14 days. PMID- 10867728 TI - Assessment of intracellular water by whole body bioelectrical impedance and total body potassium in HIV-positive patients. AB - OBJECTIVE: Bioelectrical impedance analysis (BIA) is widely used as bedside assessment of body composition. Body cell mass (BCM) and intracellular water (ICW) are clinically important body compartments. Estimates of ICW obtained from BIA by different calculation approaches were compared to a reference method in male HIV-infected patients. PATIENTS: Representative subsample of clinically stable HIV-infected outpatients, consisting of 42 men with a body mass index of 22.4 +/- 3.8 kg/m(2)(range, 13-l31 kg/m(2)). METHODS: Total body potassium was assessed in a whole body counter, and compared to 50 kHz monofrequency BIA and multifrequency bioelectrical impedance spectroscopy. Six different prediction equations for ICW from BIA data were applied. Methods were compared by the Bland Altman method. RESULTS: BIA-derived ICW estimates explained 58% to 73% of the observed variance in ICW (TBK), but limits of confidence were wide (-16.6 to +18.2% for the best method). BIA overestimated low ICW (TBK) and underestimated high ICW (TBK) when normalized for weight or height. Mono- and multifrequency BIA were not different in precision but population-specific equations tended to narrower confidence limits. CONCLUSION: BIA is an unreliable method to estimate ICW in this population, in contrast to the better established estimation of total body water and extracellular water. Potassium depletion in severe malnutrition may contribute to this finding but a major part of the residual between methods remains unexplained. PMID- 10867729 TI - Effects of parenteral infusion with medium-chain triglycerides and safflower oil emulsions on hepatic lipids, plasma amino acids and inflammatory mediators in septic rats. AB - This study was designed to investigate the effects of preinfusion with total parenteral nutrition (TPN) using medium-chain triglycerides (MCT) versus safflower oil (SO) emulsion as fat sources on hepatic lipids, plasma amino acid profiles, and inflammatory-related mediators in septic rats. Normal rats, with internal jugular catheters, were divided into two groups and received TPN. TPN provided 300kcal/kg/day with 40% of the non-protein energy provided as fat. All TPN solutions were isonitrogenous and identical in nutrient composition except for the fat emulsion, which was made of SO or a mixture of MCT and soybean oil (9:1) (MO). After receiving TPN for 6 days, each group of rats was further divided into control and sepsis subgroups. Sepsis was induced by cecal ligation and puncture, whereas control rats received sham operation. All rats were classified into four groups as follows: MCT control group (MOC, n= 8), MCT sepsis group (MOS, n= 8), safflower oil control group (SOC, n= 8), and safflower oil sepsis group (SOS, n= 11). The results of the study demonstrated that the MOS group had lower hepatic lipids than did the SOS group. Plasma leucine and isoleucine levels were significantly lower in the SOS than in the SOC group, but no differences in these two amino acids were observed between the MOC and MOS groups. Plasma arginine levels were significantly lower in septic groups than in those without sepsis despite whether MCT or safflower oil was infused. Plasma glutamine and alanine levels, however, did not differ between septic and non septic groups either in the SO or MO groups. No differences in interleukin-1b, interleukin-6, tumor necrosis factor-alpha, and leukotriene B(4)concentrations in peritoneal lavage fluid were observed between the two septic groups. These results suggest that catabolic reaction is septic rats preinfused MCT is not as obvious as those preinfused safflower oil. Compared with safflower oil, TPN with MCT administration has better effects on reducing sepsis-induced liver fat deposition. Preinfusion with MCT before sepsis, however, had no effect on inflammatory-related cytokines or leukotriene in peritoneal lavage fluid. In addition, plasma arginine appears to be a more sensitive indicator than glutamine for septic insult. PMID- 10867730 TI - Body mass index as a predictor of 1 year mortality in geriatric patients. AB - BACKGROUND AND AIMS: The predictive value of body mass and functional capacity for 1 year mortality was examined retrospectively in 552 consecutive geriatric patients categorized in 14 diagnosis groups. METHODS: Data on body mass index (BMI, kg/m(2)) was retrievable from 337 subjects. In 532 patients, Katz indexes of activities of daily life (ADL, A-G; A=independent, G total dependence) were registered. The mean age (+/- SD) was 81 +/- 8 years, two-thirds were women and 75% lived alone. Mortality data was obtained from the Swedish population records. RESULTS: Thirty-six per cent of the patients had BMI values < or =43% had BMI 21 25 and 21% > 25. Less than 2% were diagnosed as malnourished. The 1 year mortalities of those with BMI < or = 20, BMI 21-25 and BMI > 25 were 48%, 29% and 18% respectively (P< 0.001). Katz ADL index was significantly worse in those with BMI > 20 as compared with those having BMI < or = 20 (Katz D and C (P< 0.01) respectively). Patients with chronic obstructive lung disease displayed the lowest BMI values, i.e. 20 +/- 4. A logistic regression analysis indicated that BMI, gender and Katz ADL index, but not age, diagnosis or marital status, were independent predictors of 1 year mortality. CONCLUSION: Depletion may still be an overlooked problem in geriatric patients, in whom low body mass index appears to be independently associated with imminent death. PMID- 10867731 TI - Provision of rhIGF-I/IGFBP-3 complex attenuated development of cancer cachexia in an experimental tumor model. AB - Tumor growth is associated with development of cachexia which includes progressive wasting and anorexia. Our previous studies have indicated that insulin like growth factor-I (rhIGF-I) in complex with its binding protein 3 (IGFBP 3), but not free IGF-I, was a potent stimulator of muscle protein synthesis in rats with chronic undernutrition. The aim of the present study was to evaluate the effect of rhIGF-I/IGFBP-3 on the development of cancer cachexia, and to assess safety data on net tumor growth and progression during treatment. METHODS: A methylcholantrene induced sarcoma was implanted s.c. in C 57 bl mice. The animals were provided with rhIGF-I/rhIGFBP-3 (5 microg/g bw) i.v. twice daily (n= 18). Controls were provided with saline (n= 20). Body weight and food intake were registered daily. Net tumor growth was measured over 10 days. Protein synthesis in liver and muscle, as well as plasma concentrations of glucose, insulin, IGF-I and amino acids were measured at the end of the study. RESULTS: tumor size did not differ between control mice and rhIGF-I/rhIGFBP-3 treated mice (1.5 +/- 0.1 g wet tumor weight vs 1.6 +/- 0.2 g respectively). Saline treated tumor bearing controls lost 9.1 +/- 1.3 % body weight over 10 days due to rapid tumor growth while rhIGF-I/rhIGFBP-3 provision attenuated weight loss to 5.6 +/- 1.3% of body weight in study mice (P< 0.05). Food intake was improved and blood glucose concentration was reduced from 7.1 +/- 0.5 to 5.8 +/- 0.2 (P< 0.05) in response to treatment. CONCLUSION: Our results demonstrate that rhIGF-I/rhIGFBP-3 complex did not affect net tumor growth. Moreover rhIGF-I/rhIGFBP-3 complex improved tumor-host nutritional state by improving food intake, attenuating weight loss and improving glucose metabolism. PMID- 10867732 TI - The 'cut and push' method of percutaneous endoscopic gastrostomy tube removal. AB - BACKGROUND AND AIMS: The standard method of removing percutaneous endoscopic gastrostomy tubes is by gastroscopy. This has implications for endoscopy time and resources, and we believe is not always necessary. Depending on the type of percutaneous endoscopic gastrostomy tube used we often used the 'cut and push' method. This involves cutting the catheter at skin level and allowing the tube and internal bumper to spontaneously pass. The cut and push method also represents a considerable resource saving compared to the endoscopic method that we think warrants further discussion. METHOD: We reviewed all the files of the percutaneous endoscopic gastrostomy tubes removed in our unit over the last 4 years. RESULTS: During the period of July 1995 to July 1999, we have inserted 384 percutaneous endoscopic gastrostomy tubes. Seven tubes have been removed endoscopically and 73 tubes have been removed with the cut and push method. Only two possible complications have been recorded (2.7%). CONCLUSIONS: We believe that we have provided further evidence that percutaneous endoscopic gastrostomy tubes can be removed safely using the cut and push method. Patients who are often frail and who have multiple medical problems are saved an often-long journey to the endoscopy unit as well as the hazards of an endoscopy. The saving in resources in what is already an overworked system by not performing endoscopies is also considerable. PMID- 10867733 TI - Prolonged exclusive breast-feeding from vegan mother causing an acute onset of isolated methylmalonic aciduria due to a mild mutase deficiency. AB - We describe a case of methylmalonic aciduria (MMA) occurred in a 22-month-old boy whose diet was exclusively based upon breast-feeding from a mother following a long-lasting strict vegetarian diet. Clinical picture showed a dramatic onset, with a profound drowsiness associated with a severe metabolic acidosis, hyperammonemia, macrocytic anemia, ketonuria, and massive methylmalonic aciduria without homocystinuria. Both symptoms and biochemical findings quickly improved thanks to prompt vitamin B(12)parenteral therapy. Biochemical and enzymatic findings allowed a diagnosis of mild mutase deficiency, which only and inadequate dietary B(12)contribution might have revealed. Our case highlights the risk of a prolonged strictly vegetarian diet of lactating mother for providing inadequate amounts of some nutrients to the breast-fed baby. Moreover, such a dietary behaviour could dramatically unmask otherwise clinically unapparent metabolic defects in the baby. PMID- 10867734 TI - The possible effect of seasonal mood changes on the seasonal distribution of myocardial infarction. AB - The incidence of acute myocardial infarction is increased in the winter months. Seasonality of mood and behavior is common throughout the general population. Increased cardiovascular morbidity and mortality in patients with depressive disorders have been well documented. The author suggests that seasonal mood changes may contribute to the increased incidence of acute myocardial infarction in the winter. The author further suggests that patients with cardiovascular disorders who suffer from seasonal mood changes should be identified and receive the appropriate treatment for seasonal depressive symptoms that may improve the quality of life for these individuals and reduce their incidence of acute myocardial infarction. PMID- 10867735 TI - Sex, kings and serial killers and other group-selected human traits. AB - (Note: This unorthodox paper contains the first argument for heart disease being a programmed age change and promoted by the dramatic, post age-40 increases in the hormones FSH and hCG seen in some individuals.) A recent issue of Science suggests that the evolutionary purpose of sex is unknown. PARADOX: Surviving to adulthood implies a valuable gene combination which is destroyed by sexual recombination. This should be detrimental to offspring. PROPOSED: Sex is group selected in prey to allow coalescence of beneficial, and disposal of detrimental, mutations in single individuals enabling rapid adaptation to novel predation. Group selection is a universal force driven by local inter-species (not intra species) competition. Aging, metabolism, litter size, and fixed body size are directly linked. Sexual recombination and chromosomes destroy gene linkage and exist because mutations are usually detrimental, rarely positive, and occur in linked groups. In unevolving environments, sex is selected against and asexuality emerges. Periodic evolution of novel predators, like man, can explain the 'punctuated equilibria' fossil record. Genes inhibited by methylation or chromatin condensation, expressed at older ages in predation-minimized environments, allow for group selection. Stress increases mutation rates and beneficial mutation likelihood. Females select bigger, brighter, louder, or stronger males that can survive predator attention. Size approximates age and thus predator encounters; male traits represent predation-survival potential. Human male traits include, balding, acne, beard-length, wrinkling, graying, nose/ear growth. Progeria accelerates development of most male traits. Domination of groups by single males allows rapid predation-defense evolution: adolescent males are expelled, brave the wild, and expel another group's male to mate. If expelled and dominant males are culled by predation, males reaching puberty first will reproduce. Hormonal acceleration of puberty accelerates aging/population turnover, induces smaller bodies, larger litters. With a fixed group biomass, more, smaller, stressed individuals with faster aging/turnover, increase beneficial mutation likelihood. 'Kin selection', where dominant families are supported by celibate relatives, allow the best group genes to survive famine. Dominant families gorge while others starve. Equal food sharing results in group extinction leading to group-evolved human traits of social hierarchy, greed, king/queen/God worship. Menstrual hormone cycling parallels aging. FSH and DHT promote ovarian, hair, acne, dental, and arterial follicle development causing ovulation, hair growth, pimples, dental caries, and atherosclerotic soft plaques. Soft plaques contain macrophages and LDL plug; upper plaque layers thin and rupture, releasing LDL plug, causing thrombosis. FSH withdrawal or LH/hCG increases trigger ovulation and thrombosis. Artery narrowing atherosclerotic hard plaques are stress-induced through cortisol-promoted necrotic calcification. LH/hCG-induced apoptosis promotes ovulation and aging-related somatic atrophy. Long-term estradiol stimulates, while progesterone suppresses, gonadotropin levels. Estradiol protects by inhibiting gonadotropin bioactivity and has extracellular antioxidant, but intranuclear free radical, effects. Female X linked gene mosaicism conserves evolved aging systems. Maternal age factors for chromosomal trisomy suggest menopause prevents human parthenogenesis. Homosexuality and serial killing inhibit genetic contribution by individuals evolutionarily perceived as stressed. Smoking during pregnancy may induce homosexual offspring. Nitric oxide, a free radical, stimulates cGMP, but not cAMP. cGMP likely first evolved as an antioxidant defense to free radicals. Human aging syndromes might reflect human evolution progression. AS#4 affects tissues evolved from plant ancestors, AS#5a - from predators, AS#5b-immune system, and AS#6-sex tissues. (ABSTRACT TRUNCA PMID- 10867736 TI - Better use of Darwinian concepts might change the way we look at some diseases. AB - Medical science seems generally to be fixed in a paradigm within which individual organisms are viewed as harmonious units with all their component parts functioning towards a common good. This paper shows how a more revealing research methodology might be generated if a cellular, as opposed to organismic, perspective were used when thinking about some pathologies. From the cellular perspective, cells in multicellular organisms are seen to abandon intracellular cooperation and 'go solo' in some situations. This change in cellular conduct is facilitated by vestigial or introduced genetic instruction. When cells in multicellular organisms stop cooperating we call it disease. PMID- 10867737 TI - Tubaric pregnancy - an excellent idea? AB - Until now, the treatment in cases of tubal pregnancy has been focused on the patient and has not been concerned with saving the embryo. We consider a hypothetical operation: the transfer of the tube, the seat of developing pregnancy, with its vascularization, to the inside of the uterus cavity by an incision in the myometrium, and binding of the breach. In order to support this hypothesis, a review has been made of the literature on surgical treatments of transplantation of the ovary into the uterus at the beginning of the century. PMID- 10867738 TI - Pulmonary and other examples of muscular malfunction, especially muro-sphincteric conflict in contractile chambers...commentaries. AB - The mammalian body is composed of similar organs, the contractile chambers, which are therefore subject to similar inherent disorders. The basic unit is a muscular compartment which according to its function may or may not have sphincters. Muscles come in pairs, which normally cooperate. A universal form and cause of disease is failure of the essential reciprocity between mated muscles and allied groups. Clinicians encounter them as the inherent disorders of the hollow muscular organs/contractile chambers. PMID- 10867739 TI - Possible role of gangliosides in salivary gland complications of diabetes. AB - Gangliosides have previously been considered to be possible antigenic sites in Type 1 diabetes. Lymphocytic infiltration of Langerhans islands is the pathologic hallmark of autoimmune diabetes and may also be observed in salivary glands in experimental diabetes. Diabetic complications of parotid and submaxillary glands may therefore be related with an autoimmune process against sialoglycoconjugates of salivary gland tissue. PMID- 10867740 TI - Shifts in information processing level: the speed theory of intelligence revisited. AB - A hypothesis is proposed here to reconcile the inconsistencies observed in the IQ P3 latency relation. The hypothesis stems from the observation that task-induced increase in P3 latency correlates positively with IQ scores. It is hypothesised that: (a) there are several parallel information processing pathways of varying complexity which are associated with the generation of P3 waves of varying latencies; (b) with increasing workload, there is a shift in the 'information processing level' through progressive recruitment of more complex polysynaptic pathways with greater processing power and inhibition of the oligosynaptic pathways; (c) high-IQ subjects have a greater reserve of higher level processing pathways; (d) a given 'task-load' imposes a greater 'mental workload' in subjects with lower IQ than in those with higher IQ. According to this hypothesis, a meaningful comparison of the P3 correlates of IQ is possible only when the information processing level is pushed to its limits. PMID- 10867741 TI - Specific toxins destabilize virus inhibitors (e.g. AIDS viruses). AB - There are two approaches to the study of viral infections - the A-Z hypothesis of Sprietsma involving thiol-zinc inhibition of protease activation in the virus coat, and the activity of toxins in depleting the cell of thiols, or the activity of toxins activating (not detoxifying) the mixed function oxidase (MFO) system. Both of these systems generate free radicals and utilize thiols in the process. Zinc forms stable mercaptides with thiols, inhibits cyclic reduction-autoxidation of thiols and superoxide generation. When the MFO system acts as an activator and not as a detoxifier in that intermediate products are more toxic than the original compound, zinc inhibits the oxidation. An example of increased toxicity with increased MFO activity is the toxin 3-methylindole (3-MI), a toxic product of intestinal bacterial putrefaction which reactivates the infectious bovine rhinotracheitis virus (IBR). Zinc reduces MFO activity and in this regard it functions synergistically with antioxidants in protecting cell membranes. It is hypothesized that stable zinc complexes inhibit activity of proteases in the virus nucleocapside (NC) proteins in the virus coat, both directly and indirectly because zinc also inactivates some toxins that are thiol depleters or virus reactivators. PMID- 10867742 TI - A possible role of soluble receptors to cell growth factors in body aging. AB - It is assumed that production by differentiated cells of soluble receptors to cell growth factors may mediate a feedback mechanism controlling cell growth and differentiation in the body. Based on this assumption, it is hypothesized that with age a concentration of such soluble receptors in the body fluids gradually augments as a consequence of increasing a proportion of the differentiated cell pool. In the old body, when present in the cellular microenvironment at relatively high concentrations these receptors might markedly diminish ligand binding to the membrane-bound counterparts in a competitive manner and, thereby, significantly reduce cell regeneration activity. Under such conditions, the niches forming because of cell death could be being filled by connective fibers rather than newly generated cells. PMID- 10867743 TI - The beneficial effect of donor-specific transfusions: a review of existing explanations and a new hypothesis based on a relatively unapplied theory of T cell immunoregulation. A regulatory hypothesis in progress... AB - The mechanism by which donor specific transfusions protect a graft from the recipient's immune system is unknown. It is likely that this beneficial mechanism is a subset or distinct exhibition of the general rules governing the regulation of the immune system. This phenomenon provides a strong framework for investigation of immune regulation, considering its potential consanguinity to immune regulation, that it is a paradox representing a manifestation of regulatory rules, and that it provides a wealth of clinical experience and experimentation from which to make inferences. Vital in any exploration of immune regulation, is the promise held in reducing the immune system to its chief elemental regulatory mechanisms and interactions. Strangely, the majority of this consequential work may have already been accomplished by Gershon, Green and colleagues with their elegant demarcation of T cell regulation into suppressor and contrasuppressor pathways. The practical and theoretical implications of this discovery seem to be, for the most part, ignored by mainstream immunology. It is doubtful, based on the quality and quantity of their work, or confirming work by other laboratories that they were inaccurate in their findings. It remains a horrible waste that their discoveries are not in immunology's pantheon of hallowed discoveries and are little used. With all this kept in mind, a comprehensive hypothesis of regulation was put together based mainly on Gershon's portrait of the suppressor and contrasuppressor pathways' contributions to immune regulation and experimentation surrounding the unsolved paradox of donor specific transfusions. PMID- 10867744 TI - Correlation between predicted theoretical mechanistic biochemistry (TMB) data and therapeutic effects in the management of vascular disorders with calcium channel blockers. AB - Theoretical mechanistic biochemistry (TMB) analysis was used to predict the therapeutic effects of calcium channel blockers in the drug management of hypertension, cerebrovascular disorders (CVD) and coronary artery disease (CAD). This analysis was extended to acetylsalicylic acid (aspirin) a non-calcium channel blocker which is nevertheless commonly used in the management of the same disorders. TMB data have suggested nisoldipine, nicardipine and nimodipine as agents of choice in the management of cerebrovascular disease, e.g. in transient ischemic attacks (TIAs). The same agents were found preferable in the management of coronary artery disease. It is noteworthy that atherosclerosis and vascular spasm are common pathogenic events in both conditions. For lowering blood pressure, without compromising cerebral and coronary blood flows TMB data suggested nisoldipine, nicardipine, nimodipine and nifedipine in that preferential order. For tissue selectivity, TMB data have identified nisoldipine, nicardipine, nifedipine, nimodipine and nitrendipine for vascular tissue and that verapamil, diltiazem and aspirin have little or no tissue selectivity. TMB data have gone further to suggest a combination of nicardipine, nisoldipine or nimodipine with beta-blockers in order to reduce the frequently uncomfortable reflex tachycardia often induced by some calcium channel blockers. By and large, TMB predicted data have been found to correlate reasonably well with clinically observed and reported therapeutic effects of calcium channel blockers. Their consistency in the management of hypertension, cerebrovascular disease and coronary artery disease is apparent in this study. PMID- 10867745 TI - Water clusters in life. AB - The role of water in chemical, biochemical and cellular events has only been recognized as a universal solvent. The conventional wisdom holds that water is a passive agent in biological interaction. However, more and more researchers regard water as an active component in biochemical reactions and hence occupy a crucial role in life. We propose that the active component of water is due to the existence of stable water clusters in aqueous solutions. Our research demonstrated that stable water clusters could be produced in very dilute inorganic and organic water solutions, and also isolated from biological fluids such as bovine serum. Stable water clusters may play an important role in physiological and pathological processes of life. PMID- 10867746 TI - Evolutionary hypothesis of long-term memory. PMID- 10867747 TI - Origin of the immunological pregnancy test. AB - Promising innovations which enhance accuracy of laboratory testing and shorten turnaround time in the laboratory should receive timely recognition. Reporting a finding is more important than the apprehension that the finding might later prove to be an erroneous 'fluke'. Delay in reporting a finding until it is supported by more exacting and extensive testing may cause a clinically significant test to go undiscovered, whatever the reason. This article credits those who assisted the author in developing the world's first nonbiological pregnancy test. By serially injecting rabbits with human chorionic gonadotropin (HCG) mixed with Freund's adjuvant, the author induced antibodies to HCG. By our use of the latex particle fixation slide test, these antibodies could be detected and measured to a precision of within 10 IU. PMID- 10867748 TI - Five hourly measurements of serum cholesterol levels: a new methodology to assess and evaluate stress, good health and disease. AB - Research has developed a concrete link between psychological/emotional stress and life-threatening diseases such as heart disease and cancer. Here we present a technique to assess the magnitude of stress from cholesterol variation number (CVN). This number is the difference between the highest and the lowest cholesterol concentrations that relates to the five hourly cholesterol measurements performed over a five hour span. Since cholesterol in serum arises from the liver, the CVN is equated with the fluctuations in hepatic biosynthesis. This relationship is explained on the basis of the rhythmic hormonal secretions associated with cholesterol biosynthesis. Whenever stress-induced aberrations in timing of hormonal secretions occur, CVN changes. Individuals with lower CVN would have overall better health than persons with higher CVN. Thus by utilizing CVN, physicians may be able to differentiate cardiovascular health of individuals with the same or very similar serum cholesterol concentrations. PMID- 10867749 TI - Transplantation psychoneuroimmunology: building hypotheses. AB - The research findings of psychoneuroimmunology have not yet been fully applied to the field of transplantation psychiatry. Though much study has been devoted to the impact of psychiatric disease on the immunosuppressed state and disease progression in HIV-related illness, little has yet been written on the immunology implications of psychiatric disturbances in the immunosuppressed post-transplant patient. Utilizing Medline literature searches to review relevant research data in psychoneuroimmunology and transplantation immunology, the author formulates and examines four transplantation psychoneuroimmunology hypotheses involving the potential impact of depression on post-transplant organ rejection, cancer, coronary artery disease, and infections. The author concludes that though major questions remain, it appears reasonable to include the impact of depression, and possibly other psychological states, among factors that may affect the net state of immunosuppression in transplant patients. PMID- 10867750 TI - Is autism a G-alpha protein defect reversible with natural vitamin A? AB - Autism may be a disorder linked to the disruption of the G-alpha protein, affecting retinoid receptors in the brain. A study of 60 autistic children suggests that autism may be caused by inserting a G-alpha protein defect, the pertussis toxin found in the DPT vaccine, into genetically at-risk children. This toxin separates the G-alpha protein from retinoid receptors. Those most at risk report a family history of at least one parent with a pre-existing G-alpha protein defect, including night blindness, pseudohypoparathyroidism or adenoma of the thyroid or pituitary gland. Natural vitamin A may reconnect the retinoid receptors critical for vision, sensory perception, language processing and attention. Autism spectrum disorders have increased from 1 in 10 000 in 1978 to 1 in 300 in some US communities in 1999. Recent evidence indicates that autism is a disorder of the nervous system and the immune system, affecting multiple metabolic pathways. PMID- 10867751 TI - The 'consciousness primitive': a competence model of mind and will. AB - Questions about mind and will are usually raised as though mind and will are either present or absent. Here, a view is presented to suggest that they are only a particular instantiation of a 'consciousness primitive', present in lower animals as well as in primitive portions of the human brain. Physiological variables, drugs and transmitters alter it in order to produce discontinuous products, such as learning, motivation, emotions etc., which are usually studied by performance models as though they are distinctive functions of the brain. PMID- 10867752 TI - Sudden infant death syndrome: a preconditioning approach to acute arterial hypoxemia. AB - The blood hemoglobin F (HbF) concentration increases in response to chronic arterial hypoxemia and is abnormally elevated in sudden infant death syndrome (SIDS) post-mortem indicating a need for greater oxygen affinity of hemoglobin (Hb) or diminished oxygen usage by tissues or both. Modifying Hb oxygen affinity in rats revealed that increased, rather than decreased, hemoglobin-oxygen affinity permitted survival at greatly reduced environmental oxygen pressures equivalent to high altitude. Decreased Hb-oxygen affinity resulted in bradycardia 5-10 minutes before death. Cardiorespiratory recordings from infants dying suddenly and unexpectedly at home demonstrated cardiovascular failure with hypotension and bradycardia, rather than a cessation of breathing.A fall in blood pressure and acidosis due to hypoxemia in combination with reduced arterial oxygen saturation leads to circulatory failure, heart failure and death. It is speculated that the final mechanism of SIDS mimics failure to survive at high altitudes and very low environmental oxygen pressures when low arterial oxygen pressures combine with decreased Hb-oxygen affinity lead to severe hypoxemia and death. PMID- 10867753 TI - The role of antidepressants in the treatment of abdominal obesity. AB - The pathophysiology of abdominal obesity is unclear and controversial. Recent evidence now suggests that inadequate cortisol secretion is associated with abnormalities in glucose, insulin and lipid metabolism, including hypertension, bringing the importance of the hypothalamic-pituitary-adrenal (HPA) axis in the pathogenesis of abdominal obesity to the forefront. In addition, abnormal gonadal steroid concentrations and impaired plasma growth hormone levels accompany the abdominally obese state. Since the reproductive and growth axes are inhibited at many levels by various components of the HPA axis, increasing cortisol levels results in further depression of testosterone and growth hormone concentrations. Over the last decade, antidepressant (serotoninergic) drugs have proved useful as equalizers of HPA axis hyperactivity. Such therapy may interrupt the vicious circle of a hyperactive HPA axis leading to increasing abdominal obesity and endocrine perturbations that, in turn, leads to progressive accumulation of abdominal fat. Additionally, preliminary results indicate that serotoninergic agents decrease abdominal fat mass with improvements in related risk factors. PMID- 10867754 TI - A role of network arrangements of microcirculation vessels in cardiovascular function. AB - The objective of the work was to explain the mechanisms for the appearance or creation of the so-called preferential channels for blood flow in tissues. We postulated that the addressed preferential blood flow occurs at the level of network structure of vascular system and results directly from its heterogeneity. The studies have been done in regular, two-dimensional network model of microvessels. Due to poorly defined dimensions, shapes and tortuousity of real vessels, the values of network elements were random. We examined how the heterogeneity of network elements influences flow redistribution within the network and the global network resistance to flow. The results indicated that the higher segmental resistance scatters the greater probability of the appearance within the network preferential flow paths passing through the whole network or being its local singularity. These channels significantly reduce the effect the global network resistance increases. PMID- 10867755 TI - Widespread countervailing genomic responses induced by chemotherapy or radiation as a cause of therapeutic failure. AB - If chemotherapy or ionizing radiation induce widespread genomic responses tending to circumvent or antagonize their ability to kill malignant cells, an additional cause for therapeutic failure would be suggested. There is evidence that some agents evoke extensive countervailing genomic activity, the nature and extent of which can be assessed with the use of 'gene chips'. PMID- 10867756 TI - Plague as HIV vaccine adjuvant. AB - Individuals homozygous for a 32-basepair deletion mutation (null mutation) in CCR5, a chemokine receptor, are nearly immune to developing HIV infection. Recently based on molecular dating evidence it has been proposed that the bacterium Yersinia pestis, cause of plague, may have been the selective pressure which selected for a high prevalence of the 32-basepair null mutation. An association of a robust chemokine response with HIV vaccine efficacy has recently been shown. I suggest that Y. pestis may be a useful adjuvant in an HIV vaccine protocol by stimulating a vigorous chemokine response. PMID- 10867757 TI - Health effects of respirator use at low airborne concentrations. AB - Respirators are considered a personal protective mechanism for preventing occupational disease because of inhaling toxic and hazardous substances. Engineering and work practice controls, in today's industrial environment, often maintain airborne contaminants below occupational hazardous levels. However, even at low concentrations, workers are often required to use respirators. Inappropriate use of respirators, during low exposure concentrations, may result in increased incidence and prevalence of disease from respirator use caused by physiological and psychological stress. This paper examines the hypothetical aspect of increased disease from using respirators at low exposure concentrations, with specific reference to the asbestos abatement industry. PMID- 10867758 TI - Sampling irregularities, or why the present estimate for risk of HIV casual transmission is probably an underestimate. AB - Updating the list of casual transmission studies yielded a minor revision of the recently reported estimate of the risk of HIV casual household transmission, estimating it as 0.3% per year of contact. A meta-analysis of the studies used to calculate this risk indicated that the estimated risk is probably an underestimate for four main sampling irregularities: (a) The studies employed samples which were based on selective non-participation of subjects including refusals and unlocatable subjects at average rates of 36% and 14%, respectively; (b) only 17% of the studies reported full numerical details of the population studied; (c) the studies were based on unjustified exclusions of subjects by the investigators at an average rate of 31%; (d) the sampling of studies cited in the transmission studies is consistent with the hypothesis that the studies were biased against reporting cases of casual transmission. In the 13% of the studies that reported full details of exclusions, the average rate of all exclusions combined was 84%. Since it is likely that cases of casual transmission were mostly included among the exclusions, this may have resulted in an unrepresentatively low frequency of casual infection among the 16% that were left to be studied. PMID- 10867759 TI - Some new thoughts on evolution: the role of the germ layers. AB - At an early stage of embryonic development, the animal body divides itself into the three germ layers - the ectoderm, mesoderm and entoderm. It is proposed that this division repeats a fundamental functional differentiation of cells in phylogeny. The tasks of life have been divided into three groups, each germ layer specializing in one of them. The task of the entoderm is metabolism and homeostasis, that of the mesoderm is the structural organization of the body, that of the ectoderm interactions with the environment. In effect, the entoderm is the chemist of the body, the mesoderm its architect and engineer, the ectoderm deals with its external affairs. It is proposed that cognitive and reasoning faculties are inherent in every germ layer and when they separate, they all take an equal share. Each performs its complex and multitudinous tasks by virtue of its faculty of conceptual thought, except that the entoderm and mesoderm began to develop that faculty millions of years before the ectoderm. PMID- 10867761 TI - Introduction PMID- 10867760 TI - Volume transmission as a mechanism of time-dependent sensitization to antidepressant therapy. AB - Administration of certain drugs such as antidepressants have been shown to produce a phenomenon known as time-dependent sensitization, a process in which the acute effects of the drugs sensitize and grow over time. While both preclinical and clinical studies have demonstrated time-dependent sensitization, it is still uncertain how it occurs. It is possible that the mechanism by which time-dependent sensitization occurs may be via a type of transmission in the brain known as volume transmission. Signals released via this transmission have properties that may explain the process of time-dependent sensitization. PMID- 10867762 TI - Laser tissue interaction in direct myocardial revascularization. AB - This investigation examines the various laser choices used for transmyocardial laser revascularization (TMLR) with emphasis on the laser-tissue interaction. A series of in vivo (porcine model, n=27) and in vitro experiments were performed to study the effects of CO(2), holmium:YAG, and XeCl excimer lasers on the histological outcome of TMR channels. Computerized histopathological analysis has revealed that the CO(2) and holmium:YAG lasers produce substantial unpredictable thermal damage and differ predominantly in the amount of the mechanical injury or tissue shredding. In comparison, the excimer laser appears to produce the most uniform tissue ablation with the least thermal and shockwave damage. PMID- 10867763 TI - An examination of potential mechanisms underlying transmyocardial laser revascularization: channels, angiogenesis and neuronal effects. PMID- 10867764 TI - Transmyocardial laser revascularization: surgical experience overview. AB - Transmyocardial revascularization (TMR) using holmium:yttrium-aluminium-garnet (YAG) and carbon dioxide lasers has been approved by the United States Food and Drug Administration for the treatment of medically refractory angina in patients without conventional options. In prospective, randomized trials, patients who received TMR experienced improved angina, better-event free survival, and reduction in cardiac-related rehospitalizations when compared to patients remaining on medical therapy alone. In addition, TMR as an adjunct to coronary artery bypass grafting (CABG) has resulted in improved clinical status for patients who would not be completely revascularized by CABG alone. PMID- 10867765 TI - Percutaneous direct myocardial revascularization: an overview of systems. PMID- 10867766 TI - Percutaneous myocardial revascularization: European trials overview. PMID- 10867767 TI - Percutaneous transmyocardial laser revascularization: overview of US clinical trials. AB - Data from four surgical trials of laser transmyocardial revascularization (TMR) show pain reductions of at least two classes occurred among 25-76% of patients, significantly greater than with optimal medical therapy. The safety and feasibility of percutaneous TMR (PTMR) was proved in multiple uncontrolled registry studies. With PTMR in two randomized trials enrolling about 550 patients, improvements were found among 45% and 66% of patients as compared with 13% for best medical therapy. Other encouraging observations included significant improvements in exercise time (85 and 100 seconds) as compared with minimal or negative results for medical therapy, consistent overall exercise time improvements for PTMR, and overall patient perceptions of anginal stability and global health. Peri-procedural mortality is low and ranged from 0% to 0.6%. Some skepticism remains about the procedures, arising from failure in most cases to show improvements with SPECT imaging and lack of conclusive insights into the mechanism of action. A randomized 'blinded' study is underway to establish a definitive therapeutic value for catheter-based TMR. Nonetheless, preliminary clinical data seem promising for catheter-based TMR for obtaining symptomatic improvement in patients with chronic refractory ischemic coronary syndromes. PMID- 10867769 TI - Special Selection: An Ulcerated Umbilical Nodule. PMID- 10867768 TI - Acute changes of global and regional left ventricular function immediately after direct myocardial revascularization. AB - Direct myocardial revascularization (DMR) has been proposed to treat patients with severe coronary artery disease who are not amenable for classical revascularization techniques such as percutaneous coronary intervention (PCI) or bypass surgery (CABG). Although recent reports suggest its benefit in alleviating patients' complaints in the long term, there is still a paucity of data on the immediate impact on regional and global myocardial functioning following this treatment. In this overview we discuss our own experience and provide a summary of other data currently available. PMID- 10867770 TI - Patient Commentary. PMID- 10867771 TI - Patient Commentary. PMID- 10867772 TI - Finding the beginning or predicting the future? PMID- 10867773 TI - Inflammation in neurodegenerative disease: good, bad, or irrelevant? PMID- 10867774 TI - Autotoxicity and Alzheimer disease. AB - I mmunological responses are considered to be either humoral, resulting from cloning of B lymphocytes, or cell mediated, resulting from cloning of T lymphocytes. Autoimmune diseases occur when the cloned products attack host tissue. Inflammation is considered a nonspecific response to injury, characterized by exudation of serum into damaged tissue, and identified by the cardinal signs of rubor, calor, dolor, and tumor. However, these classic mechanisms do not fit pathological observations of Alzheimer disease (AD) affected brain tissue. Although many of the components prominently associated with peripheral immunological and inflammatory states are present in AD lesions, there are no identifiable B lymphocytes or antibodies, and T cells are sparse. Furthermore, the blood-brain barrier is intact, excluding exudation of exogenous serum proteins. Although "neuroinflammation" is the term commonly used to describe the pathological changes, it fails to define adequately the process that is taking place. The reaction is neither a nonspecific response to injury, as classically implied for inflammatory reactions, nor an autoimmune reaction, despite the directed attack against plaques and extracellular tangles. It is most appropriately defined as an innate immunoreaction. The fact that such a reaction can be mounted by brain, an organ frequently described as being immunologically privileged, suggests that a reevaluation is required of the dimensions of the innate immune system, including how it operates at the tissue level. The innate immune system is primitive, while the adaptive immune system, which is directed by peripheral immune organs, is an invention of vertebrates. Even in vertebrates, however, the innate immune system is the first line of defense. Much more needs to be learned about the operation of the innate immune system in health and disease. Arch Neurol. 2000. PMID- 10867775 TI - Mechanisms of neurodegenerative disorders: Part 1: protein aggregates. PMID- 10867776 TI - Mechanisms of neurodegenerative disorders: Part 2: control of cell death. PMID- 10867777 TI - The preclinical phase of alzheimer disease: A 22-year prospective study of the Framingham Cohort. AB - OBJECTIVES: To relate performance on tests of cognitive ability to the subsequent development of probable Alzheimer disease (pAD) and to identify the pattern of earliest changes in cognitive functioning associated with a diagnosis of pAD. DESIGN: From May 1975 to November 1979, a screening neuropsychological battery was administered to Framingham Study participants. They were followed up prospectively for 22 years and examined at least every 2 years for the development of pAD. SETTING: A community-based center for epidemiological research. PARTICIPANTS: Subjects were 1076 participants of the Framingham Study aged 65 to 94 years who were free of dementia and stroke at baseline (initial) neuropsychological testing. MAIN OUTCOME MEASURE: Presence or absence of pAD during a 22-year surveillance period was related to test performance at initial neuropsychological testing. RESULTS: Lower scores for measures of new learning, recall, retention, and abstract reasoning obtained during a dementia-free period were associated with the development of pAD. Lower scores for measures of abstract reasoning and retention predicted pAD after a dementia-free period of 10 years. CONCLUSIONS: The "preclinical phase" of detectable lowering of cognitive functioning precedes the appearance of pAD by many years. Measures of retention of information and abstract reasoning are among the strongest predictors of pAD when the interval between initial assessment and the development of pAD is long. Arch Neurol. 2000. PMID- 10867778 TI - Cortical inflammation in Alzheimer disease but not dementia with Lewy bodies. AB - BACKGROUND: There have been no previous studies on the role of inflammation in the brain for the second most common dementing disorder, dementia with Lewy bodies. OBJECTIVE: To investigate the degree of cortical inflammation in dementia with Lewy bodies (DLB) compared with Alzheimer disease (AD) and control brains. DESIGN AND MAIN OUTCOME MEASURES: Post-mortem tissue collection from a brain donor program using standardized diagnostic criteria. Brains collected from January 1, 1993, through December 31, 1996, were screened and selected only for the presence or absence of tau neuritic plaques. Results of immunohistochemistry for HLA-DR were quantified using area fraction counts. Counts were performed by investigators who were unaware of the diagnosis. Results were compared across groups using analysis of variance and posthoc testing. SETTING: A medical research institute in Sydney, Australia. PATIENTS: Eight brains with DLB and without the tau neuritic plaques typical of AD, 10 brains with AD and no Lewy bodies, and 11 nondemented controls without significant neuropathological features were selected from a consecutive sample. RESULTS: Compared with AD, DLB demonstrated significantly less inflammation in the form of HLA-DR-reactive microglia in all cortical regions (P<.001, posthoc). The level of inflammation in DLB was comparable to that seen in controls (P=.54, post hoc). CONCLUSIONS: Inflammation appears related to the tau neuritic plaques of AD. Despite similar clinical presentations, therapeutic anti-inflammatory strategies are not likely to be effective for pure DLB. Arch Neurol. 2000. PMID- 10867779 TI - Apolipoprotein E polymorphism and Alzheimer disease: The Indo-US Cross-National Dementia Study. AB - BACKGROUND: The APOE*E4 allele of the gene for apolipoprotein E (APOE) has been reported as a risk factor for Alzheimer disease (AD) to varying degrees in different ethnic groups. OBJECTIVE: To compare APOE*E4-AD epidemiological associations in India and the United States in a cross-national epidemiological study. DESIGN: Case-control design within 2 cohort studies, using standardized cognitive screening and clinical evaluation to identify AD and other dementias and polymerase chain reaction to identify APOE genotyping. PARTICIPANTS: Rural community samples, aged 55 years or older (n=4450) in Ballabgarh, India, and 70 years or older (n=886) in the Monongahela Valley region of southwestern Pennsylvania. MAIN OUTCOME MEASURES: Criteria of the National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association for probable and possible AD and Clinical Dementia Rating (CDR) scale for dementia staging. RESULTS: Frequency of APOE*E4 was significantly lower (P<.001) in Ballabgarh vs the Monongahela Valley (0.07 vs 0.11). Frequency of probable or possible AD, with CDR of at least 1.0, in the Indian vs US samples, was as follows: aged 55 to 69 years, 0.1% (Indian sample only); aged 70 to 79 years, 0.7% vs 3.1%; aged 80 years or older, 4.0% vs 15.7%. Among those aged 70 years or older, adjusted odds ratios (95% confidence interval) for AD among carriers of APOE*E4 vs noncarriers were 3.4 (1.2-9.3) and 2.3 (1.3-4.0) in the Indian and US samples, respectively, and not significantly different between cohorts (P=. 20). CONCLUSION: This first report of APOE*E4 and AD from the Indian subcontinent shows very low prevalence of AD in Ballabgarh, India, but association of APOE*E4 with AD at similar strength in Indian and US samples. Arch Neurol. 2000. PMID- 10867780 TI - Effect of anti-inflammatory medications on neuropathological findings in Alzheimer disease. AB - BACKGROUND: There has been no analysis of brain tissue from longitudinally observed, cognitively tested patients to validate whether anti-inflammatory medications protect against the pathological changes of Alzheimer disease. OBJECTIVE: To investigate the role of anti-inflammatory medications in alleviating the pathological features of Alzheimer disease. DESIGN AND MAIN OUTCOME MEASURES: A 5-year postmortem tissue collection was performed after a case-control study of Alzheimer disease (approximately 90 [30%] of patients died during follow-up, of whom consent for autopsy was obtained in 44 [50%]). Cases were selected on the basis of (1) adequate clinical histories of nonsteroidal anti-inflammatory drug usage, (2) no neuropathological findings other than Alzheimer disease, and (3) no generalized sepsis at death. Variables analyzed included neuropsychological test scores and amount of tissue inflammation and Alzheimer-type pathological changes. Two-way analysis of variance was used to determine whether drug usage significantly affected these variables. SETTING: The Centre for Education and Research on Ageing and the Prince of Wales Medical Research Institute, Sydney, Australia. PATIENTS: Twelve patients with Alzheimer disease (5 taking anti-inflammatory drugs) and 10 nondemented controls (3 taking anti-inflammatory drugs) were selected (50% of available sample). RESULTS: Of the patients with Alzheimer disease, anti-inflammatory drug users performed better on neuropsychological test scores than did nonusers. However, there were no significant differences in the amount of inflammatory glia, plaques, or tangles in either diagnostic group. CONCLUSION: Long-term anti-inflammatory medications in patients with Alzheimer disease enhanced cognitive performance but did not alleviate the progression of the pathological changes. Arch Neurol. 2000. PMID- 10867781 TI - The course of cognitive impairment in preclinical Alzheimer disease: three- and 6 year follow-up of a population-based sample. AB - OBJECTIVES: To examine the ability of the total score and individual items from the Mini-Mental State Examination in predicting the development of Alzheimer disease (AD) across a 3- and 6-year period in a population-based sample, and to describe the longitudinal changes in these measures across the same follow-up periods. DESIGN: Prospective follow-up of a community-based cohort, with 3 times of testing across a 6-year period. At each time of measurement, participants were clinically examined by physicians to identify demented and nondemented participants according to Diagnostic and Statistical Manual of Mental Disorders, Revised Third Edition, criteria. PARTICIPANTS: The study population consisted of all participants who were nondemented at the first follow-up and participated in the second follow-up examination. Among those, 459 remained nondemented and 73 developed AD during the second follow-up period. RESULTS: Baseline differences in the total Mini-Mental State Examination score and the delayed memory item were seen 6 years before eventual dementia diagnosis (P<.01). Analysis of the longitudinal changes showed no differences in the rate of decline for the incident AD or nondemented group between time 1 and time 2 (P>.10). However, the incident AD group exhibited precipitous declines in 8 of the 10 subscales between time 2 and time 3, the point at which they were clinically diagnosed (P<.01). Logistic regression analyses showed that only the delayed memory item was a significant predictor of who would develop AD, independent of age, sex, and years of education, at both of the first 2 times of measurement (P<.001). CONCLUSIONS: The diagnosis of AD is preceded by a long preclinical phase in which deficits in memory performance are most common. These deficits remain relatively stable up until the time that a dementia diagnosis can be rendered. Arch Neurol. 2000. PMID- 10867782 TI - Region-specific neurotrophin imbalances in Alzheimer disease: decreased levels of brain-derived neurotrophic factor and increased levels of nerve growth factor in hippocampus and cortical areas. AB - BACKGROUND: Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), and neurotrophin 4/5 (NT-4/5) are members of the neurotrophin gene family that support the survival of specific neuronal populations, including those that are affected by neurodegeneration in Alzheimer disease (AD). OBJECTIVE: To determine whether neurotrophin protein levels are altered in the AD-affected brain compared with control brains. METHODS: We quantitated protein levels of NGF, BDNF, NT-3, and NT-4/5, and calculated neurotrophin/NT-3 ratios in AD-affected postmortem hippocampus, frontal and parietal cortex, and cerebellum, and compared them with age-matched control tissue (patients with AD/controls: hippocampus, 9/9 cases; frontal cortex, 19/9; parietal cortex, 8/5; and cerebellum, 5/7, respectively). We applied highly sensitive and specific enzyme-linked immunosorbent assays in rapid-autopsy derived brain tissue (mean+/-SD postmortem interval, 2. 57+/-1.75 h, n=71) to minimize postmortem proteolytic activity. RESULTS: Levels of BDNF were significantly reduced in hippocampus and parietal cortex (P<.001, and P<.01) as well as BDNF/NT-3 ratios in frontal and parietal cortices (P<.05, and P<.01) in the group with AD compared with the control group. Levels of NGF and NGF/NT-3 ratio were significantly elevated in the group with AD compared with the control group in the hippocampus and frontal cortex (P<.001). Levels of NT-4/5 and the NT 4/NT-3 ratio were slightly reduced in hippocampus and cerebellum in the group with AD compared with the control group (P<.05). In contrast, the levels of NT-3 were unchanged in all brain regions investigated. CONCLUSION: Decreased levels of BDNF may constitute a lack of trophic support and, thus, may contribute to the degeneration of specific neuronal populations in the AD-affected brain, including the basal forebrain cholinergic system. Arch Neurol. 2000. PMID- 10867783 TI - Progression of parkinsonism and loss of cognitive function in Alzheimer disease. AB - OBJECTIVE: To assess the relation between parkinsonism and cognitive function in Alzheimer disease from cross-sectional and longitudinal perspectives. DESIGN: Prospective cohort study with annual clinical evaluations during a 4-year period. SETTING: Alzheimer disease clinic in an urban medical center. PARTICIPANTS: Four hundred ten persons with clinically diagnosed Alzheimer disease. MAIN OUTCOME MEASURES: Global and specific measures of cognitive function and parkinsonism. RESULTS: Higher levels of parkinsonism at baseline were reliably associated with lower levels of cognitive function at baseline and with more rapid cognitive decline during the 4-year study period. However, the associations were small, with baseline parkinsonism accounting for less than 10% of the variation either in baseline cognitive function or in the rate of cognitive decline. By contrast, rates of change in parkinsonism and cognitive function were strongly correlated, with 70% or more shared variance in the rates of change in many models. The association was observed with diverse measures of cognition and parkinsonism and was not explained by demographic variables or use of neuroleptic medications. CONCLUSION: In Alzheimer disease, progressive worsening of parkinsonism is more strongly associated with cognitive decline than previously recognized. Arch Neurol. 2000. PMID- 10867784 TI - Left frontotemporal hypoperfusion is associated with aggression in patients with dementia. AB - BACKGROUND: Aggressive behavior is common in patients with dementia. Temporolimbic and prefrontal cortical lesions can produce pathological aggression; however, involvement of these structures has not been established in aggressive patients with dementia. OBJECTIVE: To study the relation between regional brain perfusion and aggressive behavior in patients with dementia. METHODS: We compared the pattern of regional cerebral perfusion determined with technetium Tc 99m-labeled hexamethylpropelene amineoxime single photon emission computed tomography in 2 groups of 10 patients with dementia with and without aggression, that were comparable for demographic factors, severity of cognitive impairments, and other behavioral symptoms as measured by the Neuropsychiatric Inventory. RESULTS: Patients with aggression revealed significant (P<.001) hypoperfusion in the left anterior temporal cortex; additional bilateral dorsofrontal and right parietal cortex were also found to be significantly hypoperfused. CONCLUSION: These results indicated an association between aggression and decreased perfusion in the left anterior temporal cortex. Arch Neurol. 2000. PMID- 10867785 TI - Association between severe cerebral amyloid angiopathy and cerebrovascular lesions in Alzheimer disease is not a spurious one attributable to apolipoprotein E4. AB - BACKGROUND: We have previously reported an association between severe cerebral amyloid angiopathy (CAA) and cerebrovascular lesions in Alzheimer disease (AD), which is particularly strong for microinfarcts, hemorrhages, and multiple lesion types. Cerebral amyloid angiopathy has also been associated with the apolipoprotein E4 (APOE4) genotype, which is in turn associated with premature coronary artery disease and atherosclerosis. OBJECTIVE: To test whether severe CAA would be more strongly associated with cerebrovascular lesions than would APOE4 genotype. METHODS: We reviewed 306 cases of autopsy-confirmed AD (from the University of California, San Diego, brain autopsy series) to assess whether APOE genotype and other clinical risk factors were predictive of vascular lesions (VLs) in AD. Cerebral amyloid angiopathy severity was assessed using a semiquantitative scale in 4 brain regions (ie, hippocampus, midfrontal cortex, inferior parietal cortex, and superior temporal cortex) and an average score was computed for each case. RESULTS: We found that severe CAA was associated with an increased frequency of VLs (33% of the cases of severe CAA had VLs vs 19% of the cases of mild or absent CAA; P=.02). While the APOE4/4 genotype was associated with an increased severity of CAA, there was no significant relationship between APOE genotype and frequency of VLs. Logistic regression models showed that severe CAA, advanced age, atherosclerosis, and Hachinski Ischemia Scale score of 7 or more were all significantly associated with VLs, but the number of APOE4 alleles, history of hypertension, coronary artery disease, sex, and serum cholesterol levels had nonsignificant effects. Within strata of APOE genotype, the presence of severe CAA was associated with increased frequency of VLs (eg, within APOE4/4 homozygotes, VLs were present within 47% of the cases of severe CAA vs 9.5% of the cases of mild or absent CAA; P=.01). CONCLUSIONS: Severe CAA confers a greater risk of VLs in AD, even within strata of APOE genotype. Therefore, the association between severe CAA and VLs in AD is not a spurious one owing to APOE4. Overall, our cases of AD with APOE4 do not seem to be a more "vasculopathic" subtype of AD. The mechanisms by which CAA produces VLs of various types need to be further elucidated, as these are probably important in producing the common entity of "mixed" AD/vascular dementia. Arch Neurol. 2000. PMID- 10867786 TI - Assessing financial capacity in patients with Alzheimer disease: A conceptual model and prototype instrument. AB - OBJECTIVE: To investigate financial capacity in patients with Alzheimer disease (AD) using a new theoretical model and prototype psychometric instrument. DESIGN: Cross-sectional comparisons of older control subjects (n=23) and patients with mild (n=30) and moderate AD (n=20). MAIN OUTCOME MEASURES: Financial capacity was measured using the Financial Capacity Instrument, a prototype psychometric instrument that tests financial capacity using 14 tasks of financial ability comprising 6 clinically relevant domains of financial activity: basic monetary skills, financial conceptual knowledge, cash transactions, checkbook management, bank statement management, and financial judgment. RESULTS: The Financial Capacity Instrument tasks and domains showed adequate to excellent internal, interrater, and test-retest reliabilities. At the task level, patients with mild AD performed equivalently with controls on simple tasks such as counting coins/currency and conducting a 1-item grocery purchase, but significantly below controls on more complex tasks such as using a checkbook/register and understanding and using a bank statement. At the domain level, patients with mild AD performed significantly below controls on all domains except basic monetary skills. Patients with moderate AD performed significantly below controls and patients with mild AD on all tasks and domains. Regarding capacity status outcomes (capable, marginally capable, incapable) on domains, patients with mild AD had high proportions of marginally capable or incapable outcomes (range, 47% 87%), particularly on difficult domains like bank statement management (domain 5) and financial judgment (domain 6), but variability in individual outcomes. Patients with moderate AD had almost exclusively incapable outcomes across the 6 domains (range, 90%-100%). CONCLUSIONS: Financial capacity is already significantly impaired in mild AD. Patients with mild AD demonstrate deficits in more complex financial abilities and impairment in most financial activities. Patients with moderate AD demonstrate severe impairment of all financial abilities and activities. The Financial Capacity Instrument has promise as an instrument for assessing domain-level financial activities and task-specific financial abilities in patients with dementia. Arch Neurol. 2000. PMID- 10867787 TI - Early-onset Alzheimer disease caused by a new mutation (V717L) in the amyloid precursor protein gene. AB - CONTEXT: Alzheimer disease is the most common form of dementia. Mutations in the genes amyloid precursor protein (APP), presenilin 1(PS1) and presenilin 2(PS2) have been found in early-onset familial forms of Alzheimer disease OBJECTIVE: To determine the cause of dementia in a family with early-onset illness. DESIGN, SETTING, AND PARTICIPANTS: A family with a history of dementia was referred to the Indiana Alzheimer Disease Center, Indianapolis. All the research in this study was done in a university or university hospital. The proband and her 4 siblings took part in the study. The proband, who is still alive, showed symptoms of Alzheimer disease at 38 years of age. Genomic DNA was obtained from blood samples of 5 family members. The APPandPS1genes of the proband were screened for mutations by amplification followed by direct sequencing. RESULTS: Sequence of exon 17 of the APPgene revealed a single nucleotide (guanine to cytosine) substitution in 1 allele, resulting in an amino acid change at codon 717 (valine to leucine). Each of the proband's siblings were tested for this mutation by direct sequencing. Two of the 4 were found to have the mutation; one of whom was recently clinically diagnosed at the age of 36 years. CONCLUSIONS: A novel mutation in the APPgene (V717L) has been found in a family with a history of dementia, beginning in the mid to late 30s. The age of onset in this family is earlier than most of the other families with Alzheimer disease who also have APPmutations. Arch Neurol. 2000. PMID- 10867788 TI - Active intracerebral hemorrhage from the lateral posterior choroidal artery. PMID- 10867789 TI - History of neurology: seminal citation. Aphasia. PMID- 10867791 TI - Topography, histology, and seminology in dementia PMID- 10867790 TI - Topography, histology, and seminology in dementia. PMID- 10867792 TI - In vivo glioma model enabling regulated gene expression. AB - Experimental investigation of glioma biology and therapy requires a representative model and a convenient technique for regulating gene expression. We have established an in vivo model in which genetically modified rat C6 glioma cells (C6TL cells) are transplanted into nude mice brain, followed by specific transcriptional control of a transgene. Histologically, the tumors exhibit an astrocytic phenotype and closely resemble human malignant gliomas including diffuse brain invasion. Due to a stably integrated lacZ gene, individual tumor cells can be unequivocally identified in tissue sections by histochemistry for beta-galactosidase. Since C6TL cells carry the tet transactivator (tTA) gene, any additional gene under control of a tetracycline/tTA-responsive promoter can be transcriptionally regulated by the concentration of tetracycline. C6TL cells stably transfected with a tetracycline/tTA-responsive luciferase reporter gene showed 23-fold regulation of luciferase activity in vitro. After intracerebral transplantation a regulation of 4.5- to 8.3-fold was obtained, dependent on the concentration and the type of tetracycline in the drinking water. This model should be useful for studying the functional role of candidate genes in tumor biology as well as for experimental gene therapy studies. PMID- 10867793 TI - Selective accumulation of cyclooxygenase-1-expressing microglial cells/macrophages in lesions of human focal cerebral ischemia. AB - Cyclooxygenases (COX; prostaglandin endoperoxide H synthases) are key enzymes in the conversion of arachidonic acid into prostanoids which mediate inflammation, immunomodulation, mitogenesis, ovulation, fewer, apoptosis and blood flow. Here, we report COX-1 expression following focal cerebral infarctions (FCI). In healthy control brains, COX-1 was localized by immunohistochemistry to a few endothelial cells, single neurons and rare, evenly distributed brain microglia/macrophages. In infarctioned brains, COX-1+ cells accumulated highly significantly (P < 0.0001) in peri-infarctional areas and in the developing necrotic core early after infarction. Here, cell numbers remained persistently elevated up to several months post infarction. Further, clusters of COX-1+ cells were located in perivascular regions related to the Virchow-Robin space. Double-labeling experiments confirmed co-expression of COX-1 by CD68+ microglia/macrophages. Co expression of the activation antigens HLA-DR, -DP, -DQ (MHC class II) or the macrophage inhibitor factor-related protein MRP-8 (S100A8) by most COX-1+ microglia/macrophages was only seen early after infarction. Thus, COX-1 appeared to be expressed in microglial cells regardless of their activation state. However, the prolonged accumulation of COX-1+ microglia/macrophages restricted to peri-infarctional areas enduring the acute post-ischemic inflammatory response points to a role of COX-1 in tissue remodeling or in the pathophysiology of secondary injury. We have identified localized, accumulated COX-1 expression as a potential pharmacological target following FCI. Therefore we suggest that therapeutic approaches based on selective COX-2 blocking might not be sufficient for suppressing the local synthesis of prostanoids. PMID- 10867794 TI - Ubiquitinated filamentous inclusions in cerebellar dentate nucleus neurons in dentatorubral-pallidoluysian atrophy contain expanded polyglutamine stretches. AB - We have recently reported that, in addition to the widespread occurrence of ubiquitinated neuronal intranuclear inclusions (NIIs), the restricted occurrence of ubiquitinated intracytoplasmic filamentous inclusions in the neurons of the cerebellar dentate nucleus (CDN) is a characteristic feature of dentatorubral pallidoluysian atrophy (DRPLA). Interestingly, these neuronal intracytoplasmic filamentous inclusions (NIFIs) were morphologically indistinguishable from the skein-like inclusions (SLIs) described previously in the spinal anterior horn cells in amyotrophic lateral sclerosis (ALS). In the present study, we examined immunohistochemically the CDN in ten patients with clinicopathologically and genetically confirmed DRPLA and the spinal anterior horns in five patients with sporadic ALS, using a monoclonal antibody (1C2) directed against long polyglutamine stretches. In all of the patients with DRPLA, both the NIFIs and the NIIs were visualized clearly with 1C2. Conversely, in the patients with ALS all structures, including the SLIs, were completely negative. These findings indicate that in DRPLA, the NIFIs in the CDN are an alteration that is directly related to the causative gene abnormality (an expanded CAG repeat encoding polyglutamine) and that, from the molecular point of view, they are distinct from the SLIs in ALS. PMID- 10867795 TI - Expression of MyoD and myogenin in dystrophic mice, mdx and dy, during regeneration. AB - Expression of two myogenic regulatory factors, MyoD and myogenin, was studied in regenerating muscles of dystrophic mice and compared to a chemically induced regeneration process. First, the distribution of the two proteins was determined immunohistochemically at various time points after single administrations of a local anaesthetic, bupivacaine hydrochloride, which causes myonecrosis followed by regeneration. Detectable levels of MyoD appeared at 18 h and the expression reached their maximum levels at 48 h after the injection, which coincide with the stage when satellite cells are activated and start to proliferate. Myogenin became detectable in 24 h and its expression reached its highest level at 72 h after injection when newly formed myotubes appeared. The two genes were also expressed in the dystrophic muscles from dy and mdx mice which exhibit dystrophic pathological features but are associated with different phenotypes. In mdx mice the two genes were expressed at reasonably high levels in parallel with the active regenerating process, whereas in dy mice MyoD and myogenin expressions decreased as fibrosis progressed. However, MyoD was relatively more strongly expressed in the larger mature myotubes of dy mice than in those of mdx mice, suggesting prolonged regenerative activity. In dy and mdx mice, MyoD and myogenin were expressed in different quantities, indicating that these animals have distinct regenerating activities. Our findings confirm that expression of both MyoD and myogenin genes is necessary in the regenerative process for the proliferation of satellite cells (myoblasts) and for the development of early regenerating fibers (myotubes) even in dystrophic muscles. PMID- 10867796 TI - Constant involvement of the Betz cells and pyramidal tract in multiple system atrophy: a clinicopathological study of seven autopsy cases. AB - We investigated clinicopathologically the pyramidal signs, including spasticity, hyperreflexia, and Babinski's sign, and the involvement of the pyramidal tract and primary motor cortex, in seven Japanese autopsy cases of multiple system atrophy (MSA). Pyramidal signs were observed in six (86%) of the seven autopsy cases. Hyperreflexia and Babinski's sign were each evident in five patients, but spasticity was observed in only one patient. Loss of Betz cells and presence of glial cytoplasmic inclusions in the primary motor cortex were noticed in all seven cases. Astrocytosis in the fifth layer of the primary motor cortex was noticed in five cases, but its presence was not related to the duration of the disease. Involvement of the pyramidal tract in the spinal cord, particularly of the small myelinated fibers, was observed in all seven cases, but no involvement of the pyramidal tract in the midbrain was evident in any of the six cases in which this structure was examined. In MSA, pyramidal signs were shown to be present more frequently than believed before, and the clinicopathological correlation between pyramidal signs and involvement of the pyramidal tract was obvious. Constant involvement of Betz cells in MSA has not been reported. Our clinicopathological findings may also make a contribution to the understanding of the clinicopathological hallmarks of MSA. PMID- 10867797 TI - Electron microsocopy of brain amyloid plaques from a patient with new variant Creutzfeldt-Jakob disease. AB - Cerebral cortex biopsy from a patient with new variant Creutzfeldt-Jakob disease (nvCJD) has been examined at the electron microscope level. Spongiform changes corresponded mostly to distended neurites scattered in the neuropil or surrounding amyloid plaques. These latter exhibited heterogeneous submicroscopic morphology including variable amount of loosely interwoven amyloid fibrils admixed in a cellular-rich environment constituted essentially by abnormal neuronal processes. By immunoelectron microscopy, fibrils and some membrane structures reacted with anti-prion protein (PrP) antibodies. One striking aspect was the presence of many small dystrophic neurites without paired helical filaments. Moreover, amyloid fibrils showed unexpected intimate association with abnormal membranes, suggesting a relationship between PrP fibrillogenesis and membrane alteration. These ultrastructural findings provide an additional criterion to distinguish nvCJD-from sporadic CJD-type plaques and reinforce the hypothesis that nvCJD brain is infected by a distinctive strain of the transmissible agent encephalopathy. PMID- 10867798 TI - AIDS does not alter the total number of neurons in the hippocampal formation but induces cell atrophy: a stereological study. AB - Although cognitive dysfunction is a common finding in patients with acquired immunodeficiency syndrome (AIDS) its pathogenesis remains controversial. Given the involvement of the hippocampal formation in the processing of cognitive information and the scarcity of quantitative studies in this brain region, we have examined, using stereological methods, the hippocampal formations of AIDS patients. The study was performed in ten AIDS patients and ten age-matched controls. All cases were male. The Principle of Cavalieri was applied to estimate the volume of the layers of the dentate gyrus and of the CA3 and CA1 hippocampal fields. The fractionator and the nucleator were used as estimators of the total number, and mean somatic and nuclear volumes of the neurons in the cell containing layers of all hippocampal subdivisions. No cell death was detected in AIDS patients but the global volume of their hippocampal formations was significantly decreased due to the reduced volume of its layers, mainly the cell containing layers. Furthermore, the somatic and nuclear volumes of the neurons in the hippocampal formation were significantly decreased in AIDS patients. No correlation was found between the estimates obtained and the presence or absence of neurological involvement. Our results show that neurons in the hippocampal formation of AIDS patients display marked morphological changes, despite the maintenance of their total number. These alterations are likely to lead to dysfunction of the hippocampal circuitries and, thus, might contribute to explaining the dementia features which occur in this condition. PMID- 10867799 TI - Aciculin and its relation to dystrophin: immunocytochemical studies in human normal and Duchenne dystrophy quadriceps muscles. AB - Aciculin is a novel adherens junction antigen extracted from human uterine smooth muscle that is reported to associate biochemically with dystrophin. We attempted to determine (i) the immunostainability of anti-aciculin antibody for the 6 histochemically normal human muscles and seven muscles from boys with Duchenne muscular dystrophy (DMD) and 11 disease control muscles, (ii) the ultrastructural localization of aciculin in normal skeletal myofibers, (iii) aciculin's spacial relationship with dystrophin and beta-spectrin, and (iv) if the aciculin is ultrastructurally colocalized with dystrophin, the distance from the aciculin epitope to the epitope of the dystrophin N- or C-terminal domain. For this, rabbit anti-aciculin antibody was generated against the synthetic peptide of aciculin fragment D [4]. Immunohistochemical staining showed that the immunostainability of DMD muscles for anti-aciculin antibody was markedly decreased as compared with normal and disease control muscles. Single and double immunogold labeling electron microscopy of 6 histochemically normal human quadriceps femoris muscles revealed that aciculin was present along the inner surface of muscle plasma membrane and that aciculin formed doublets more frequently with dystrophin (23.5 +/- 1.8%; group mean +/- SE) than with beta spectrin (12.8 +/- 1.1%; P < 0.01 two tailed t test). Rabbit anti-aciculin antibody frequently formed doublets with monoclonal antibodies against the N- or C-terminal domain of dystrophin at the muscle cell surface. These results suggest that aciculin is associated with dystrophin and may interact with both the N- and C-terminal domains of dystrophin. PMID- 10867800 TI - Distinctive neuropathology revealed by alpha-synuclein antibodies in hereditary parkinsonism and dementia linked to chromosome 4p. AB - The identification of the alpha-synuclein gene on chromosome 4q as a locus for familial Lewy-body parkinsonism and of alpha-synuclein as a component of Lewy bodies has heralded a new era in the study of Parkinson's disease. We have identified a large family with Lewy body parkinsonism linked to a novel locus on chromosome 4p15 that does not have a mutation in the alpha-synuclein gene. Here we report the clinical and neuropathological findings in an individual from this family and describe unusual high molecular weight alpha-synuclein-immunoreactive proteins in brain homogenates from brain regions with the most marked neuropathology. Distinctive histopathology was revealed with alpha-synuclein immunostaining, including pleomorphic Lewy bodies, synuclein-positive glial inclusions and widespread, severe neuritic dystrophy. We also discuss the relationship of this familial disorder to a Lewy body disease clinical spectrum, ranging from Parkinson's disease to dementia with psychosis. PMID- 10867801 TI - Involvement of intercellular adhesion molecule-1 in myelin recognition by macrophages. AB - Macrophages play a crucial role in myelin removal during nerve degeneration and demyelination. The exact mechanisms of myelin recognition and uptake are not yet defined. The present experiments aimed at defining the role of intercellular adhesion molecule-1 (ICAM-1) in this process. Myelin phagocytosis was studied in an established in vitro model of cultured macrophages and sciatic nerves. Cocultures of wild-type C57BL macrophages with sciatic nerves resulted in a massive invasion of the nerves by macrophages with subsequent removal of myelin. In contrast, when macrophages of ICAM-1-deficient animals were cocultured with wild-type nerves, myelin phagocytosis was significantly retarded, whereas cell invasion was completely undisturbed. These data indicate that the ICAM-1 molecule acts as a costimulatory signal in myelin recognition and uptake by macrophages. PMID- 10867802 TI - Cyclosporine A-associated fatal central nervous system angiopathy in a bone marrow transplant recipient: an autopsy case. AB - We report here the case of a 32-year-old woman who suffered from a unique angiopathy in the central nervous system (CNS). She died of multiple infarcts in the brain stem and cerebellum during treatment with cyclosporine A after bone marrow transplantation for refractory anemia with excess of blasts. The autopsy findings showed segmental narrowing of the basilar artery, in which circumferential dissection of the internal elastic lamina had occurred. The distal portion of the basilar artery was obstructed by upward dislocation of the dissected intima. Similar angiopathy was also observed at multiple sites along the basilar artery branches. These findings suggest endothelial damage, including vasoconstriction and dissection of the CNS arteries. PMID- 10867803 TI - Diencephalic neuronal hamartoma associated with congenital obstructive hydrocephalus, anophthalmia, cleft lip and palate and severe mental retardation: a possible new syndrome. AB - A male infant was born with severe hydrocephalus, bilateral cleft lip/palate, left anophthalmos and right microphthalmos, and an equino-varus foot deformity. Imaging studies showed enlarged lateral ventricles, apparent absence of the corpus callosum and a midline density in the third ventricular region. He had a normal male karyotype. He was severely mentally retarded and died suddenly at 7 years of age. Neuropathological examination of the brain revealed enlarged and polygyric cerebral hemispheres, due to congenital obstructive hydrocephalus, and secondary thinning of the corpus callosum. An unusually large neuronal hamartoma filled the interpeduncular fossa and third ventricle. It was continuous posteriorly with the left thalamus and so was classified as diencephalic rather than as hypothalamic. The right optic nerve merged with the hamartoma, whereas the left nerve was absent. Microscopically the hamartoma consisted of mature grey matter interspersed with narrow bands of white matter. No immature or non-neural elements were identified. This combination of diencephalic neuronal hamartoma, hydrocephalus, ocular and craniofacial abnormalities has not, to our knowledge, previously been described. PMID- 10867804 TI - Intracytoplasmic hyaline inclusions in the hippocampal neurons of two cattle. AB - This report deals with the large intracytoplasmic hyaline inclusions observed in hippocampal large pyramidal cells of two Holstein-Friesian cattle. These inclusions were round to elongated polyhedral in shape with consistently homogeneous glassy appearance; they varied in size and were positive for the periodic acid-Schiff reaction and silver impregnation. Electron microscopic examination revealed that the inclusions consisted of granular materials showing moderate electron density and were bounded by a unit membrane. On the external surface of the unit membrane, there were direct connections to cellular organelles, including ribosome, rough endoplasmic reticulum, and mitochondria. These findings suggest that the inclusions might be derived from neuronal endoplasmic reticulum. PMID- 10867805 TI - Glial intranuclear inclusion bodies in a patient with Alzheimer's disease. AB - We report a case of dementia in an elderly woman with the pathological findings of Alzheimer's disease and numerous intranuclear inclusions in astrocytes and occasionally in neurons. These inclusions were seen in the cerebral cortex, limbic areas, basal ganglia, thalamus, brain stem and cerebellum. They expressed ubiquitin and were ultrastructurally composed of haphazardly arranged straight filaments. The presence of similar intranuclear inclusions in previous cases of adult-onset dementia without other neuropathological changes suggests an important link between these kind of inclusions and dementia. To our knowledge, this type of intranuclear inclusions has not been previously described in association with Alzheimer pathology. PMID- 10867806 TI - Suprasellar chordoid glioma. AB - Brat et al. (J Neuropathol Exp Neurol 57:288-290, 1998) reported eight cases of a new clinico-pathological entity, which occurs mainly in the third ventricle of middle-aged females, which they described as chordoid glioma of the third ventricle. We report a new case of a 41-year-old woman with a suprasellar chordoid glioma with histological, immunohistochemical and ultrastructural studies. We discuss the differential diagnosis between chordoma, chordoid meningioma, germinoma and pituitary adenoma. Histologically, the tumour showed cords and lobules of isomorphic epithelioid cells in a vacuolated matrix with prominent multifocal lymphoplasmacytic infiltrates in which some histiocytes and isolated Touton-type giant cells were seen; cells were immunoreactive for glial fibrillary acidic protein but negative for epithelial membrane antigen. Ultrastructural examination revealed abundant intermediate filament but no desmosomes, microvilli nor cilia were seen. PMID- 10867807 TI - The first case of new variant Creutzfeldt-Jakob disease in France: clinical data and neuropathological findings. AB - Clinical data and autopsy findings in a case of new variant Creutzfeldt-Jakob disease (vCJD) are reported. This case, the first histologically confirmed case described outside the United Kingdom, very much resembles the cases described by Will et al. [(1996) Lancet 347:921-925] and Zeidler et al. [(1997) Lancet 350:903 908, 908-910]. Neuropathological studies failed to reveal any conspicuous clues that could be relevant for understanding the pathophysiology of the disease. For epidemiological surveillance, neuropathologists should scrutinize suspected cases keeping in mind the possibility of vCJD. PMID- 10867808 TI - Central nervous system Hodgkin's lymphoma without systemic manifestation: case report and review of the literature. AB - A 66-year-old woman treated for ocular myasthenia gravis with azathioprine for 12 years presented with a left fronto-parietal mass. Histology revealed primary Hodgkin's lymphoma of the central nervous system with CD30, Epstein-Barr virus (EBV) latent membrane protein and CD20-positive, CD45 (LCA)-negative Reed Sternberg cells surrounded by T cells. Moreover, EBV-encoded RNA-1 (EBER-1) sequences and a monoclonal rearrangement of the immunoglobulin heavy chain CDR2 locus were detected. PMID- 10867809 TI - Development of a system for detection of circulating antibodies against hemidesmosomal proteins in patients with bullous pemphigoid. AB - Specific antibodies directed against special hemidesmosomal proteins are involved in the pathogenesis of bullous pemphigoid (BP), and detection of these antibodies is crucial for a correct diagnosis. As the BP autoantigen primary structures are known, the question was addressed as to whether it is possible to demonstrate circulating antibodies against BP autoantigens (BPAG1 and BPAG2) by means of an ELISA system, using antigenic epitopes. With the help of the programs Peptidestructure and Plotstructure, antigenic epitopes of BP antigens were predicted, chemically synthesized and screened using serum from 43 proven BP patients. The coding sequences of the best antigenic epitopes were then chemically synthesized and inserted as monomer and homo- or hetero-oligomer forms into fusion-expression plasmids (PGEX-4T, Pharmacia) in-frame to the C-terminus of glutathione-S-transferase. Fusion products were expressed and purified from Escherichia coli cells by affinity chromatography. The recombinant proteins were used for the detection of antibodies in the serum of 43 BP patients and of 60 controls (including 30 healthy persons, 22 patients with pemphigus vulgaris and 8 patients with other bullous dermatoses). Use of the homo- and hetero-oligomers of the recombinant fusion peptides increased the sensitivity of the disease-specific antibody detection. When a mixture of the best recombinant fusion proteins was used, the sensitivity of the ELISA assays in the case of the BP patients' serum was 0.90. This system could form the basis of a rapid and simple system for the diagnosis of BP. PMID- 10867810 TI - Antiapoptotic bcl-2 and bcl-xL in advanced malignant melanoma. AB - Apoptosis is an important cofactor in the pathogenesis of a plethora of malignancies. However, little is known about modulation of the expression of bcl gene family in melanocytic tumors. To determine the role of bcl-2, bcl-x and bax in melanocytic tumors we investigated the differential expression of these genes via RT-PCR in tissue samples from human benign nevi, primary melanomas and melanoma metastases in comparison with normal skin. Bcl-2 was strongly expressed in 14/16 metastases (87.5%), whereas only 7/13 primary melanomas (53%), 7/15 nevi (46%) and 7/16 normal tissue samples (43%) showed expression of bcl-2 (P < 0.05). There was a strong indication of a correlation between tumor thickness and bcl-2 expression in nodular malignant melanomas. Expression of bcl-x was found in 16/16 melanoma metastases (100%), 11/13 primary melanomas (84%), 12/15 nevi (80%) and 10/16 normal tissue samples (62%) (P < 0.05). Bcl-xL expression increased from primary melanoma to melanoma metastases, whereas bcl-xS showed a decreasing expression level during melanoma progression. No differences in bax expression were seen between melanoma metastases, primary melanoma, nevi and normal tissue. Immunohistochemical investigations of another 53 tissue samples showed similar results. Our results strongly indicate that bcl-2 and bcl-xL gene expression increases with progression of malignant melanoma. Bcl-2 and bcl-xL expression could reflect an increased malignant potential caused by an inhibition of apoptosis and growth advantage for metastatic melanoma cells. PMID- 10867811 TI - Behavioral differences between donor site-matched adult and neonatal melanocytes in culture. AB - Little is known about the biologic behaviors of cultured melanocytes in relation to donor age. To investigate age-dependent differences, neonatal and adult melanocytes were isolated from the same anatomical site, the foreskin, and cultured in the same growth medium supplemented with cAMP inducers (choleratoxin and 3-isobutyl-methylxanthine). The morphology, melanin content, pattern of melanosome distribution, and growth rate were then compared. Neonatal melanocytes were bipolar in appearance, whereas adult melanocytes were highly dendritic in appearance. Image analysis showed that adult melanocytes were larger and longer, and had a greater number of dendrites than neonatal melanocytes. When the growth medium was replaced by a medium without cAMP inducers, adult melanocytes showed a change in their morphology from dendritic to spindle-shaped, while the morphology of neonatal melanocytes remained unchanged. Melanosomes of adult melanocytes were distributed singly along the dendrites, and extracellular secretion of melanosomes was also found. In contrast, melanosomes of neonatal melanocytes were aggregated near the nuclei. No age-dependent differences in melanin content and growth rate were noted in the donor site-matched cultured melanocytes. These results suggest that donor age is one of the factors involved in determining melanocyte dendricity and melanosome distribution, and that increased dendricity of adult melanocytes is due to increased sensitivity to cAMP inducers. In addition, the adult melanocytes established in our culture system, which resembled dendritic melanocytes in vivo, could be considered a desirable model for studying the mechanisms of adult-onset hyperpigmentary disorders and melanogenesis. PMID- 10867812 TI - Interleukin-1 alpha enhances mast cell growth by a fibroblast-dependent mechanism. AB - Mast cell hyperplasia is observed in various inflammatory skin diseases. Although the pathogenesis of these conditions remains largely uninvestigated, it has been speculated that lesional mediators provide a favorable microenvironment for mast cell growth. We investigated the effect of an inflammatory cytokine, IL-1 alpha, on mast cell growth in a mast cell/fibroblast coculture system. When mouse bone marrow-derived cultured mast cells (BMMC) were cultured on a NIH/3T3 fibroblast monolayer, IL-1 alpha stimulated mast cell proliferation. However, IL-1 alpha did not stimulate 3H-thymidine incorporation in BMMC in the absence of fibroblasts. Separation of BMMC from fibroblasts by a permeable micropore membrane reduced the effect of IL-1 alpha. When BMMC were prepared from W/Wv mice, which lack a functional c-kit, or when NIH/3T3 fibroblasts were substituted with Sl/Sld derived fibroblasts, which lack membrane-bound stem cell factor (SCF), a lower, but significant, effect of IL-1 alpha was observed. Flow cytometric analysis revealed no enhancement of SCF expression on fibroblasts following stimulation with IL-1 alpha. Neutralizing antibodies against IL-3, IL-4, IL-10, and nerve growth factor (NGF) showed no inhibition. On the other hand, indomethacin inhibited the effect of IL-1 alpha, and prostaglandin E2 induced mast cell growth in the co-cultures. These results indicate that IL-1 alpha stimulates mast cell growth by a fibroblast-dependent mechanism, in which SCF/c-kit interaction may participate in a major way. The mast cell growth activity induced by this cytokine can, at least in part, be attributed to prostaglandins. Inflammatory cytokines may thus contribute to mast cell hyperplasia in skin diseases. PMID- 10867813 TI - Simultaneous analysis of [3H]-thymidine uptake and alpha 1(I) procollagen mRNA expression in systemic sclerosis skin fibroblasts--an in situ hybridization study. AB - Heterogeneity of DNA synthesis and collagen synthesis has been reported in skin fibroblasts from systemic sclerosis (SSc) patients. The uptake of [3H]-thymidine and expression of alpha 1(I) procollagen mRNA by cultured skin fibroblasts from four normal controls and four SSc patients was analyzed simultaneously. The grains overlying the cytoplasm representing alpha 1(I) procollagen mRNA and overlying the nucleus representing [3H]-thymidine uptake were counted using computer-aided image analysis. The results were analyzed statistically. Procollagen mRNA expression by SSc fibroblasts was significantly greater than by control fibroblasts (P < 0.01). The distribution curve of [3H]-thymidine uptake showed two peaks representing low- and high-uptake cells. Significantly more SSc fibroblasts than control fibroblasts showed high [3H]-thymidine uptake (P < 0.05). The number of SSc fibroblasts expressing low amounts of alpha 1(I) procollagen mRNA was significantly lower than the number of control fibroblasts (P < 0.05). [3H]-thymidine uptake by SSc fibroblasts expressing high amounts of alpha 1(I) procollagen mRNA was significantly lower than by those expressing low amounts (P < 0.05). These results indicate that elevated DNA synthesis and elevated collagen mRNA synthesis in SSc skin fibroblasts are due to different clones with high DNA-synthesizing ability and high collagen-producing ability. PMID- 10867814 TI - Acute keratinocyte damage stimulates platelet-activating factor production. AB - Recent evidence suggests that the phosphocholine-derived lipid mediator platelet activating factor (PAF) is involved in keratinocyte function and cutaneous inflammation. PAF is found in various inflammatory skin diseases, and intradermal injection of PAF directly results in cutaneous inflammation. Keratinocytes also synthesize PAF and related 1-acyl species in response to ionophores, cytokines and growth factors, and in response to activation of the epidermal PAF receptor. Since keratinocytes are routinely exposed to potential damage by thermal or oxidative stressors with resultant induction of cutaneous inflammation, the objective of these studies was to assess whether exogenous thermal or oxidative damage can induce the production of PAF and related 1-acyl species. Cells of the immortalized human keratinocyte cell line HaCaT were subjected to acute heat or cold, or treatment with the pro-oxidant lipid tertiary butyl hydroperoxide, and PAF and 1-palmitoyl-2-acetyl-GPC were measured by gas chromatography/mass spectrometry. We report that these diverse toxic stimuli resulted in the accumulation of these biologically active lipids. These studies suggest that the PAF system is involved in the inflammatory response seen following acute epidermal damage. PMID- 10867815 TI - Generation of reactive oxygen species by Candida albicans in relation to morphogenesis. AB - Candida albicans is able to generate significant amounts of reactive oxygen species (ROS). In this study, ROS generation by yeast and hyphal forms of the strain 3153 A was analyzed to determine whether ROS generation could be a major factor in the invasive behavior of germinative cells. Furthermore, the virulent strain CA6 and its avirulent and agerminative mutant VIR3 were compared. ROS were measured by lucigenin-enhanced chemiluminescence and a cytochrome c assay. During the blastoconidial phase of all strains moderate amounts of ROS were found at cell concentrations > 1 x 10(5)/ml. However, ROS generation appeared to be specifically inhibited at cell concentrations > 1 x 10(8)/ml, and this was found in both assays. As shown in comparative experiments, the medium used for measurement markedly affected the total amount of ROS. Hyphae of strain 3153 A generated a significantly higher amount of ROS than yeast cells and cells with germ tubes (P < 0.001). The strain CA6 showed significantly higher ROS generation than the VIR3 strain for both blastoconidiae and after 30 min of induction of hypha formation (P < 0.05). In conclusion, hypha formation, usually acknowledged as a major factor in Candida pathogenicity, was associated with markedly increased ROS formation. ROS generation was not closely linked to the ability to form hyphae, but was highest in germinative cells. PMID- 10867817 TI - Anterior endoderm and head induction in early vertebrate embryos. AB - Early work on the formation of the vertebrate body axis indicated the existence of separate head- and trunk-inducing regions in Spemann's organizer of the amphibian gastrula. In mammals some head-organizing activity may be located in anterior visceral (extraembryonic) endoderm (AVE). By analogy, the equivalent structure in the Xenopus laevis gastrula, the anterior endoderm, has been proposed to be the amphibian head organizer. Here we review recent data that challenge this notion and indicate that the involvement of AVE in head induction seems to be an exclusively mammalian characteristic. In X. laevis and chick, it is the prechordal endomesoderm that is the dominant source of head-inducing signals during early gastrulation. Furthermore, head induction in mammals needs a combination of signals from anterior primitive endoderm, prechordal plate, and anterior ectoderm. Thus, despite the homology of vertebrate anterior primitive endoderm, a role in head induction seems not to be conserved. PMID- 10867816 TI - Adrenoceptors in normal and malignant human melanocytes. PMID- 10867819 TI - Ontogenetic expression and sex differences of aromatase and estrogen receptor alpha/beta mRNA in the mouse hippocampus. AB - Estrogen plays an important role during brain development interfering with the maturation of distinct neural systems and, in particular, with the sexual differentiation of brain structures and function. Similar to other brain regions, estrogen is known to influence neuronal differentiation and plasticity in the hippocampus. The present study is concerned with the developmental expression of mRNAs for the estrogen-synthesizing enzyme aromatase and the two known nuclear estrogen receptors (alpha/beta) in the male and female mouse hippocampus. Using semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR) analysis, we found that aromatase as well as estrogen receptors (alpha/beta) are already expressed prenatally in the hippocampus of both sexes. Aromatase expression increased during the first two postnatal weeks and decreased, thereafter, to lower levels in adults. Sex differences in aromatase expression were observed postnatally with higher levels in males. Estrogen receptor alpha/beta mRNAs did not fluctuate obviously throughout pre- and postnatal development but revealed a distinct sex-specific pattern at the end of the first postnatal week. Again, higher expression was detected in males. These findings clearly demonstrate the capacity of estrogen formation and the presence of both estrogen receptor subtypes in the developing hippocampus. Sex differences in aromatase mRNA levels paralleled the sex-specific pattern of estrogen receptor expression. Thus, our data support the idea that the developing hippocampus is a target for estrogen action and estrogen receptor-mediated sexual differentiation. PMID- 10867818 TI - Expression of pituitary adenylate cyclase-activating polypeptide (PACAP) and the PACAP-selective receptor in cultured rat astrocytes, human brain tumors, and in response to acute intracranial injury. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) is a bioactive peptide with diverse activities in the nervous system. In addition to its more classic role as a neurotransmitter, PACAP functions as a neurotrophic factor. PACAP exerts these activities by binding to PACAP-selective (PAC1) or nonselective (VPAC1, VPAC2) receptors (-R). Glial cells also exhibit PACAP binding, which is associated with the increased proliferation of astrocytes. The present report demonstrates a distinct spatiotemporal regulation of PACAP, PAC1-R, VPAC1-R, and VPAC2-R expression in primary cultured rat astrocytes. To determine the role of PACAP and PAC1-R expression on glial proliferation, two in vivo models were examined--human brain tumors of glial origin and the reactive gliosis induced by a penetrating stab wound to the mature rat brain. Relative to normal human brain, PAC1-R expression is significantly upregulated in glioma, particularly oligodendrogliomas. While similar polymerase chain reaction (PCR) analysis does not detect PACAP expression, in situ hybridization studies reveal PACAP expression in a limited number of cells within the tumor. In sharp contrast, neither PACAP nor PAC1-R expression are upregulated consequent to injury. These results suggest a distinct role for PACAP and PAC1-R in glioma development and nervous system response to injury. PMID- 10867820 TI - Localization of Na/K-ATPase in developing and adult Drosophila melanogaster photoreceptors. AB - Drosophila melanogaster photoreceptors are highly polarized cells and their plasma membrane is organized into distinct domains. Zonula adherens junctions separate a smooth peripheral surface, the equivalent of the basolateral surface in other epithelial cells, from the central surface (approximately equal to apical surface). The latter consists of the microvillar rhabdomere and the juxtarhabdomeric domain, a nonmicrovillar area between the rhabdomere and the zonulae adherens. The distribution of Na/K-ATPase over these domains was examined by immunocytochemical, developmental, and genetic approaches. Immunofluorescence and immunogold labeling of adult compound eyes reveal that the distribution of Na/K-ATPase is concentrated at the peripheral surface in the photoreceptors R1 R6, but extends over the juxtarhabdomeric domain to the rhabdomere in the photoreceptors R7/R8. Developmental analysis demonstrates further that Na/K ATPase is localized over the entire plasma membrane in all photoreceptors in early pupal eyes. Redistribution of Na/K-ATPase in R1-R6 occurs at about 78% of pupal life, coinciding with the onset of Rh1-rhodopsin expression on the central surface of these cells. Despite the essential role of Rh1 in structural development and intracellular trafficking, Rh1 mutations do not affect the distribution of Na/K-ATPase. These results suggest that Na/K-ATPase and rhodopsin are involved in distinct intracellular localization mechanisms, which are maintained independent of each other. PMID- 10867821 TI - Apolipoprotein E gene expression correlates with endogenous lipid nutrition and yolk syncytial layer lipoprotein synthesis during fish development. AB - During embryogenesis of teleost fish, the formation of a yolk syncytial layer (YSL) enables the resorption of the yolk reserves and development up to the larval stage. We have examined the changes of the yolk cell structure in relation to yolk and oil-globule lipid utilization during development of the turbot (Scophthalmus maximus). After encapsulation by the YSL, resorption of the single, large oil globule occurred predominantly after yolk resorption and was slower in fasting larvae. The YSL was in contact with an enlarged perisyncytial space, but no vascular network or red blood cells were present within the walls of the yolk sac. Intrasyncytial channels infiltrated by pigmented lining cells were observed in the YSL surrounding the oil globule. Apolipoprotein E (apoE) has a prominent role in lipid metabolism because of its ability to interact with lipoprotein receptors. We performed molecular cloning of the putative low-density lipoprotein receptor binding domain of turbot apoE. In situ hybridization analysis revealed a very high level of apoE transcripts in the YSL, while no expression could be detected in the intestine. YSL apoE expression was correlated with the synthesis of very low density lipoprotein (VLDL) particles. An extraordinarily high number of VLDL particles were poured into the perisyncytial space, and intrasyncytial channels enabled the transfer of yolk- and oil globule-derived lipids to the developing embryo or larva. The pattern of apoE mRNA distribution in relation to YSL lipoprotein synthesis indicates that apoE expression is a suitable molecular marker for monitoring endogenous lipid nutrition during the endoexotrophic period of teleost fish development. PMID- 10867822 TI - Angiogenesis in the hypertensive lung: response to ambient oxygen tension. AB - The present study further analyzes the growth and reorganization of the vessels adjacent to capillaries in the hyperoxia-adapted lung in response to a lower ambient oxygen tension. The aim of the study was to determine the source of the new smooth muscle cells known to develop in these segments on return to breathing air. To accomplish this we determined the reorganization of vessel walls by quantitative light-microscopy techniques, and vascular cell phenotype(s) by high resolution microscopy, in the lungs of rats that breathed a high oxygen tension (87% O2 for 4 weeks), followed by weaning to a lower oxygen tension (87-20% O2 over 1 week) and return to breathing air (for 1, 2 or 4 weeks). Return to breathing air initially triggered wall growth in a subset of vessels and wall thinning in others before wall thinning predominated throughout the vessel population. Interstitial fibroblasts were identified as the source of new perivascular cells. The recruitment of these cells was accompanied by loss of elastic laminae from vessel walls. Subsequently, most perivascular cells expressed a smooth muscle phenotype and elastic laminae were restored. Arteriography demonstrated an increase in the number of patent vessels on return to air, and light- and high-resolution microscopy restitution of the capillary network. We propose that in the hyperoxia-adapted lung return to breathing air represents a relative hypoxia that triggers differential patterns of vessel and capillary growth to meet new functional demands set by the lower ambient oxygen tension. PMID- 10867823 TI - Expression of vascular endothelial growth factor by granulated metrial gland cells in pregnant murine uteri. AB - Granulated metrial gland (GMG) cells are a characteristic uterine component belonging to a natural killer cell lineage. This study is aimed at revealing their kinetic and spatial relationship with vascular growth during pregnancy and the expression of vascular endothelial growth factor (VEGF). GMG cells and blood vessels were identified by periodic-acid-Schiff-reagent (PAS)-stained granules and positive staining for factor-VIII-related antigen, respectively. GMG cells were widely distributed in the decidua and metrial gland and showed a numerical increase with a peak at day 13 in parallel with the increase of vascular density. Preceding the maximal vascular development at day 13, microvessels with a narrow lumen representative of neovascularization prevailed at days 7-9, and the VEGF content in the decidua/metrial gland was significantly elevated at days 7-13 concurrently with mRNA expression. By immunolight microscopy combined with PAS staining, GMG cells with PAS-stained granules were positive for VEGF. Immunoelectron microscopy demonstrated that immunoreactions were diffuse in the cytoplasm but not localized in the granules. In contrast, fibroblast-like stromal cells were negative. These data indicate that GMG cells express VEGF and may play inducing roles in uterine neovascularization during pregnancy. PMID- 10867824 TI - P2 receptors in the thymus: expression of P2X and P2Y receptors in adult rats, an immunohistochemical and in situ hybridisation study. AB - The expression of the seven P2X receptor subtypes and of two P2Y receptors was examined immunohistochemically and by in situ hybridisation in thymi of adult male rats. P2X4, P2Y2 and 4 receptor mRNA colocalisation studies combining in situ hybridisation and immunohistochemistry were also carried out. P2X and P2Y receptors were found on thymocytes. P2X receptors were also abundant in cells of the thymic microenvironment, involved in control of T-cell maturation in vivo. We are the first to describe the expression of P2X4 receptors on thymocytes and confirm the finding of P2X1 and P2Y2 receptors on subpopulations of lymphocytes. P2X1,2,3,4 and 5 receptors were present in blood vessels of the thymus. P2X1,2 and 4 receptors were detected in vascular smooth muscle, while P2X3 receptors appeared to be associated with endothelial cells; some small arteries were positive for P2X5, possibly labelling vascular smooth muscle or fibroblasts in the adventitia. P2X2,3,6 and 7 receptors were found on thymic epithelial cells. P2X2 and 3 receptors were abundant on medullary epithelial cells, whilst P2X6 receptors were prominent in Hassall's corpuscles. P2X2 receptors were found on subcapsular and perivascular epithelial cells. P2X2,6 and 7 receptors were detected in epithelial cells along the thymic septa. Expression of P2X receptors was also investigated by Western blotting of crude thymic tissue extracts under reducing conditions. All seven P2X receptor subtypes were found to be dimers of approximately 70 kDa and 140 kDa molecular weight. ATP-mediated apoptosis and cell proliferation of thymocytes are discussed. PMID- 10867825 TI - Cell-cycle distribution of pancreatic cells from rats with acute pancreatitis induced by bile-pancreatic obstruction. AB - The aim of this study was to analyze, using electron microscopy, the morphological alterations that progressively appear in the pancreas of rats with acute pancreatitis induced by bile-pancreatic obstruction over 48 h. In addition, in order to ascertain the capability of pancreas regeneration at different stages of pancreatitis, the distribution of pancreatic cells throughout the different phases of the cell cycle was also analyzed by flow cytometry using propidium iodide staining. Interstitial edema, macrophage infiltration, vacuolization, and dilatation of endoplasmic reticulum were observed from 1.5 h after obstruction onward. Interestingly, cell cycle studies showed an increased proportion of S phase cells at early stages of pancreatitis (1.5 h after obstruction), which leads to a significant increase in cells in G2/M phase 12 h after pancreatic obstruction. Histological studies revealed severe alterations in pancreas of rats with obstruction maintained over 48 h which affects the nuclear structure. Intracellular disorganization, apoptosis, and focal necrosis were observed at this stage. Furthermore, flow-cytometric analysis of cell DNA contents showed a significant decrease in the proportion of S and G2/M cells and a significant increase in G0/G1 cells, suggesting an arrest of almost all cells in quiescent states. These results suggest that rat pancreas cells are able to recover during the first 12 h after pancreatic obstruction. However, the gland would lose its ability to regenerate if the obstruction was maintained for longer periods. PMID- 10867826 TI - Transforming growth factors alpha and beta-1 are co-expressed in the uterine epithelium during early pregnancy. AB - Semi-quantitative immunoperoxidase light microscopy on unfixed frozen sections of rat uterus was used to examine the expression of transforming growth factors (TGF) alpha and beta before, during and after implantation. TGF beta-1 labelling was present in the lateral plasma membranes and basement membrane from day 3, steadily increased until the time of implantation on day 6 when it formed a dense apical band, but had disappeared by day 7. TGF alpha appeared in the apical epithelium, 3 days prior to TGF beta-1. Ultrastructural immunogold labelling showed the intense TGF beta-1 signal observed at the time of implantation to be present on the apical epithelial surface and microvilli and in the luminal secretions. We suggest that TGF alpha stimulates the expression of TGF beta-1 in rats, as in implanting transgenic mice, and that it is involved in extracellular matrix remodelling in the lateral plasma membranes and basal lamina to inhibit invasion and implantation and to stimulate the production of basement membrane. PMID- 10867827 TI - P2X receptor immunoreactivity in the male genital organs of the rat. AB - The distribution of ATP ionotropic P2X receptors in the genital organs of the male rat has been investigated with immunohistochemical techniques using specific antibodies to P2X1-7 receptors. In the excretory ducts of the testis (ductus epididymidis, vas deferens and its associated seminal vesicles), the major signals were seen with antibodies to P2X1 and P2X2 in the membranes of the smooth muscle layer, suggesting that these receptors are involved in the process of sperm transport and ejaculation. In the penis body, strong P2X1 and weaker P2X2 immunoreactivity was seen in the smooth muscle of blood vessels and the corpus cavernosum, suggesting a participation in the detumescence process. P2X5 immunoreactivity, a marker for differentiating cells in stratified squamous epithelia, was observed in the epithelia of the terminal urethra, the "horny spur" (spine-studded epithelium of the glans) and the inner surface of the prepuce. Antibodies to P2X3 reacted with nerve fibres in the adventitia of vas deferens, and the P2X6 receptor was localised in the basal lamina of the epithelium. In the prostate, there was immunostaining of the smooth muscle between the tubules with antibody for P2X1, but not with P2X2; P2X3 immunostaining of nerves and strong P2X7 immunostaining of the glandular epithelium of the prostate were also present. PMID- 10867828 TI - Factors affecting morphogenesis of rabbit gallbladder epithelial cells cultured in collagen gels. AB - Although peptide growth factors play an important role in the morphogenesis of gallbladder, little is known about how they effect the morphogenesis of gallbladder epithelial cells. Rabbit gallbladder epithelial cells (RGEC) were isolated and cultured in monolayer or collagen gels. Epidermal growth factor (EGF), hepatocyte growth factor (HGF), epimorphin, transforming growth factor beta 1 (TGF-beta 1), and fibroblast-conditioned medium (FCM) were added to the cultured cells to clarify the effects of these peptides and FCM on morphogenesis of RGEC. RGEC suspended in collagen gels form spherical cysts with morphologic polarity. EGF, HGF, epimorphin, and FCM promoted cyst maturation by accelerating the proliferation and aggregation of clear, polarized vesicles. In contrast, TGF beta 1 markedly inhibited DNA synthesis in both monolayer and collagen gel cultures and promoted formation of branching structures in collagen gels. Furthermore, in the presence of EGF, TGF-beta 1 induced a drastic change in morphogenesis, with the formation of branching networks that showed cell-cell contact only at sites where branches touched. RGEC-forming multicellular cysts did not express vimentin but expressed significant amounts of cytokeratin and regained junctional complexes. In contrast, TGF-beta 1-treated cells strongly expressed vimentin along with branching structures and showed decreases in cytokeratin expression and junctional complexes. Thus, TGF-beta 1 induces a mesenchyme-like cell shape accompanied by cytoskeletal molecular changes, with loss of both epithelial polarization and junctional complexes. These results suggest that the morphogenetic program of RGEC is likely to be determined by the interaction of these peptides and the timing of their presence. PMID- 10867829 TI - Neurons that express the AMPA receptor GluR2/3 subunits in suprachiasmatic nuclei of Syrian hamsters colocalize either vasoactive intestinal peptide, peptide histidine isoleucine or gastrin-releasing peptide. AB - In mammals, many circadian rhythms are driven by a clock located inside the suprachiasmatic nucleus of the hypothalamus. They are synchronized to environmental light-dark cycles by information coming directly from the retina via glutamatergic afferents. In rodents, retinal fibres make direct synaptic contacts with neurons synthesizing vasoactive intestinal peptide and gastrin releasing peptide. These two neuropeptides, administered alone or combined with the peptide histidine isoleucine, phase-shift the clock in the same way that light does. Using ICC and light and electron microscopy, our study demonstrates that subunits 2 and 3 of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid type glutamatergic receptors are colocalized in neurons expressing one or other of these three neuropeptides. Double-labelled neurons were located in the ventral and lateral ventral parts and near the symmetrical plane of the intermediate and caudal thirds of the nucleus. In light microscopy, brown and granular blue stainings of chromogens revealing both antigens were easily identifiable and spatially separated in perikarya. In electron microscopy, almost all the cells observed in these zones expressed the receptor subunits. A few labelled dendritic profiles, some of them post-synaptic, were observed; axon terminals were always unlabelled. Colocalization with vasoactive intestinal peptide and gastrin releasing peptide was confirmed by the immunogold technique in perikarya and some dendrites. The present study suggests that peptidergic neurons expressing the AMPA receptors are involved in photic entrainment of the clock by the retina without excluding some glutamatergic information coming from other hypothalamic nuclei. PMID- 10867831 TI - A new model for investigating hair cell degeneration in the guinea pig following damage of the stria vascularis using a photochemical reaction. AB - We have established a new model for investigating the relationship between cochlear lateral wall damage and sensory cell degeneration in guinea pigs by using a photochemical reaction between the systemic injection of Rose Bengal (RB) and controlled green light irradiation to the cochlea. The photochemical reaction produced a reactive oxygen species, which then damaged the endothelium. This triggered platelet adhesion and aggregation at the site of endothelial injury to produce thrombi and affect microcirculation in the lateral wall at the site of irradiation. Changes were studied under a scanning electron microscope (SEM), and compound action potentials (CAP) were measured. SEM observations after tangential illumination of the cochlear wall revealed degeneration of the stria vascularis (SV). Specific morphological findings at 24 h included delayed degeneration of the outer hair cells concurrent with a significant increase in the CAP. Based on these findings, we suggest that degeneration of the SV was a direct result of the photochemical reaction, but CAP changes and sensory hair cell damage were secondarily caused by SV degeneration. PMID- 10867830 TI - Effects of kynurenic acid as a glutamate receptor antagonist in the guinea pig. AB - Glutamate excitotoxicity is implicated in both the genesis of neural injury and noise-induced hearing loss (NIHL). Acoustic overstimulation may result in excessive synaptic glutamate, resulting in excessive binding to post-synaptic receptors and the initiation of a destructive cascade of cellular events, thus leading to neuronal degeneration and NIHL. The purpose of this study was to determine whether this apparent excitotoxicity can be attenuated by kynurenic acid (KYNA), a broad-spectrum glutamate receptor antagonist, and protect against noise-induced temporary threshold shifts (TTS). Guinea pigs were randomly assigned to three separate groups. Base-line compound action potentials (CAP) thresholds and cochlear microphonics (CM) were recorded. Group I was treated with physiologic saline as a vehicle control applied to the round window membrane that was followed by 110 dB SPL wide-band noise for 90 min. Group II received 5 mM KYNA followed by noise exposure, and group III received 5 mM KYNA alone without noise exposure. Post-drug and noise levels of CAP thresholds and CM were then obtained. Noise exposure in the control group caused a significant temporary threshold shift (TTS) of 30-40 dB across the frequencies tested (from 3 kHz to 18 kHz). Animals that received 5 mM KYNA prior to noise exposure (group II) showed statistically significant protection against noise-induced damage and demonstrated a minimal TTS ranging between 5 and 10 dB at the same frequencies. Animals in group III receiving KYNA without noise exposure showed no change in thresholds. Additionally, cochlear microphonics showed no considerable difference in threshold shifts when controls were compared to KYNA-treated animals. These results show that antagonizing glutamate receptors can attenuate noise-induced TTS, suggesting that glutamate excitotoxicity may play a role in acoustic trauma. PMID- 10867832 TI - Involvement of nitric oxide synthase in the physiology and pathophysiology of facial nerve function and dysfunction. AB - To date few reports have discussed the presence and function of nitric oxide (NO) in structures of the facial nerve. We performed nicotinamide adenine dinucleotide phosphate (NADPH-d)-diaphorase-histochemistry and immunohistochemistry on the intratemporal portion of the facial nerve, including the geniculate ganglion, of guinea pigs using specific antibodies to the three known isoforms of NO synthase and soluble guanylyl-cyclase (sGC). Normal facial nerves were compared to those treated intratympanically with bacterial lipopolysaccharides (LPS) and tumor necrosis factor-alpha (TNF-alpha). Both constitutive NOS isoforms and sGC could be detected in the bipolar ganglion cells of normal animals, while the inducible isoform (iNOS or NOS II) was not found. Endothelial NOS (NOS III) and sGC were present in blood vessels and were predominantly found in the perineurial sheath and less in the endoneurium. sGC could be detected in all fibers in a cross section of the facial nerve. LPS and TNF treatment led to the detection of iNOS in the perikaryia of the geniculate ganglion and the perineural sheath. These findings imply that NO may be involved in neurotransmission at least in the visceroafferent system. NO regulates vascular tone of nutrient blood vessels in the perineural sheath and endoneurium. The presence of sGC indicates that NO acts via its second messenger cGMP. NOS II expression may be a contributing factor to facial nerve palsy via two different mechanisms: NOS II-generated NO may lead to an overstimulation of the visceroefferent nerve fibers and motor fibers of the facial nerve. Dysregulation in facial nerve blood vessels could lead to edema and elevated pressure on the nerve within its osseous canal. PMID- 10867833 TI - Dynamic visual acuity during linear acceleration along the inter-aural axis. AB - We investigated visual-vestibular interactions during linear acceleration along the inter-aural axis. Eighteen healthy volunteers and two patients with central neurological diseases were subjected to transaural linear acceleration in the direction of gravity force (frequency: 0.5-1.5 Hz; amplitude: 5 cm). During linear acceleration, eye movements were recorded under three test conditions: eyes closed (EC), while staring at an imaginary target (IT) and during the testing of dynamic visual acuity (DVA). As parameters of evaluation we used the amplitude of horizontal eye movements, phase shift and the decrease of DVA threshold (DVAT). Under all test conditions, eye amplitude increased with rising stimulus frequency and exceeded, especially in the higher frequency range, a hypothetically calculated eye amplitude for smooth pursuit. The combination of a visual and vestibular input (DVA and IT) led to a better compensation (lower phase shift) than under vestibular stimulation alone (EC). Eye movements during low-frequency stimulation depended more on the visual system while responses in the higher frequency range were mainly triggered by the otolith organ. At 1.5 Hz the compensatory function of the visual-vestibular system was limited (rising phase shift) and DVAT decreased even in a significant number of healthy subjects. Patients with diseases of the central nervous system showed a higher phase shift and thus a stronger decrease of DVAT (two levels) already at a stimulus frequency of 1.25 Hz. PMID- 10867834 TI - Effects of macrolides on interleukin-8 secretion from human nasal epithelial cells. AB - Low-dose, long-term macrolide treatment has recently been reported to be very effective in patients with chronic airway diseases. We examined the in vivo and in vitro effects of 14-membered macrolide antibiotics erythromycin (EM) and clarithromycin (CAM) on interleukin (IL)-8 secretion from human nasal epithelial cells. Fifteen patients with chronic sinusitis received macrolide treatment (CAM 400 mg/day) for 1 to 3 months. The number of infiltrated neutrophils and IL-8 concentrations in the nasal discharges of these patients decreased significantly at 1 to 2 months after the treatment. In vitro effects of EM and CAM on IL-8 secretion were examined in nasal epithelial cells cultured at the air-liquid interface. After 14-day culture in the air-liquid interface, macrolide antibiotics were added in medium for 24 h. EM and CAM at concentrations of 10(-4) M did not affect spontaneous secretions or IL-1 beta-induced secretions of IL-8 either apically or basolaterally. When cells were preincubated with 10(-4) M CAM for 7 days, the IL-1 beta-induced secretion of IL-8 decreased significantly. However, no difference was observed between the effects of 10(-4) M CAM and 10( 4) M josamycin, a 16-membered macrolide. These results suggest that macrolide treatment inhibits neutrophil infiltration and IL-8 secretion in nasal epithelium in vivo and that these clinical effects depend on a mechanism other than the direct action of macrolide on nasal epithelial cells. PMID- 10867835 TI - Multicenter investigation of 1,036 subjects using a standardized method for the assessment of olfactory function combining tests of odor identification, odor discrimination, and olfactory thresholds. AB - "Sniffin' Sticks" is a test of nasal chemosensory performance that is based on penlike odor-dispensing devices. It is comprised of three tests of olfactory function: tests for odor threshold, discrimination and identification. Previous work has already established its test-retest reliability and validity in comparison to established measures of olfactory sensitivity. The results of this test are presented as a composite TDI score--i.e., the sum of results obtained for threshold, discrimination and identification measures. The present multicenter investigation aimed at providing normative values in relation to different age groups. To this end, 966 patients were investigated in 11 centers. An additional study tried to establish values for the identification of anosmic patients, with 70 anosmics investigated in five specialized centers where the presence of anosmia was confirmed by means of olfactory evoked potentials. For healthy subjects, the TDI score at the 10th percentile was 24.5 in subjects younger than 15 years, 30.3 for ages from 16 to 35 years, 28.8 for ages from 36 to 55 years and 27.5 for subjects older than 55 years. While these data can be used to estimate individual olfactory abilities in relation to a subject's age, hyposmia was defined as the 10th percentile score of 16- to 35-year-old subjects. Our latter study revealed that none of 70 anosmics reached a TDI score higher than 15. This score of 15 is regarded as the cut-off value for functional anosmia. These results provide the basis for the routine clinical evaluation of patients with olfactory disorders using "Sniffin' Sticks." PMID- 10867836 TI - A new method for reconstruction of the larynx after vertical partial resections. AB - The indications and problems of organ-preserving vertical partial laryngectomy (VPL) in cases of T1b glottic or T2 glottic and subglottic cancers are well known. The first and imperative requirement for the surgeon is adequate resection of tumor while the second prerequisite is the safe and successful correction of the excised portion of the anterolateral wall of the larynx. Since reconstruction of the defect can cause significant challenges for surgeons, the main requirements are an adequate lumen for breathing, a smooth surface for epithelialization, voice restoration and good deglution. Krajina's method for reconstruction of the larynx utilizes pedicled sternohyoid fascia, which is thin, elastic, well adaptable to defects, and resistent to infection or saliva. By providing a large surface for covering defects, granulations and synechiae can be prevented. We now use the superficial fascia colli as a new method for reconstruction of laryngeal defects after frontolateral partial resections. The technique was first refined experimentally in dogs. A Leroux-Robert partial laryngectomy was carried aut on five animals and laterally pedicled fascia was sutured to the edge of the defect created. At 2-week intervals through 8 weeks after the operation fixation, vascularization and epithelialization were examined histologically. To date, clinical reconstruction with the fascial flap has been used in 29 cases. Because the flap has a very low metabolism, no necrosis was seen. Functional results of respiration, phonation and swallowing have been good. These findings show that laterally pedicled fascia with the bipedicled sternohyoid muscles can play an important role in laryngeal reconstruction. PMID- 10867837 TI - Clinical experience with the treatment of T1b glottic cancer by means of horizontal glottectomy. AB - This study describes the oncological and functional results of horizontal glottectomy performed in a series of 37 similar patients with T1b glottic cancers. The 5-year overall and disease-free survival rates were, respectively, 85.4% and 91.0%. Decannulation was always possible within a mean period of 16.2 days, and no patient developed laryngeal stenosis. A bypass naso-gastric tube was removed a mean 4.9 days after surgery, and adequate swallowing was soon obtained. The mean duration of post-operative hospitalization was 16 days and no major post operative complications were observed. Satisfactory vocal function was obtained in all cases. On the basis of these results, horizontal glottectomy was found to be a reliable and safe procedure for the management of T1b glottic cancer. PMID- 10867838 TI - Normal configuration of the anterior commissure of the glottis on magnetic resonance imaging. AB - The purpose of this study was to evaluate the normal width of the anterior commissure of the glottis by measuring its dimensions in patients who had no laryngeal disease on magnetic resonance imaging (MRI), but had unrelated cervical pathology. In all, 27 patients were studied. Axial images through the arytenoid commissure at the level of the vocalis muscle and/or vocal process of the arytenoid cartilage were magnified on MRI. The anteroposterior width of the anterior commissure was measured by using an electronic ruler having 1-mm marks. The average width of the anterior commissure was 1.59 +/- 0.6 mm, varying from a minimum width of 1 mm to a maximum width of 3 mm. The width was less than or equal to 2 mm in 25 patients (92.5%). Our results showed that all of the patients studied had measurable mucosal thicknesses at the level of anterior commissure > or = 1.0 mm. PMID- 10867839 TI - Transoral laser surgery for early glottic carcinoma. AB - This prospective study evaluates the oncological results of transoral laser surgery (TLS) for glottic carcinoma categorized Tis, T1 and T2 in a large, unselected group of 285 consecutive patients from a university-based referral center that uses transoral laser surgery as the standard approach to these tumors. Patients were treated between 1 January 1987 and 31 December 1996. Thirty three patients had Tis disease, 174 T1 tumors and 113 T2. Main outcome measures were local control with initial therapy, ultimate local control, regional control, organ preservation, overall survival and cause-specific survival. The 5 year uncorrected actuarial survival for all 285 patients was 71.1%, and cause specific actuarial survival was 98.7%. Local control with initial treatment was 85.9%, ultimate local control with salvage for local treatment failures 98.5%, and regional control 98.4%. In all, 94.3% had their larynges preserved after 5 years. Although favorable oncological results for early laryngeal carcinoma treated with laser surgery are supported this study, no definitive recommendations can be given for the best single treatment. Partial laryngectomies lead to the highest local control rates reported so far, radiotherapy is believed to preserve voice best and laser surgery is associated with time- and cost-effectiveness, low morbidity, fair local control rates and excellent re-treatment options in case of local failure. All specialists dealing with the treatment of early glottic carcinoma should be able to offer these different treatment modalities to their patients and to deal specifically with each patient's individual needs and preferences. PMID- 10867840 TI - Endoscopic cordectomy. A proposal for a classification by the Working Committee, European Laryngological Society. AB - The European Laryngological Society is proposing a classification of different laryngeal endoscopic cordectomies in order to ensure better definitions of post operative results. We chose to keep the word "cordectomy" even for partial resections because it is the term most often used in the surgical literature. The classification comprises eight types of cordectomies: a subepithelial cordectomy (type I), which is resection of the epithelium; a subligamental cordectomy (type II), which is a resection of the epithelium, Reinke's space and vocal ligament; transmuscular cordectomy (type III), which proceeds through the vocalis muscle; total cordectomy (type IV); extended cordectomy, which encompasses the contralateral vocal fold and the anterior commissure (type Va); extended cordectomy, which includes the arytenoid (type Vb); extended cordectomy, which encompasses the subglottis (type Vc); and extended cordectomy, which includes the ventricle (type Vd). Indications for performing those cordectomies may vary from surgeon to surgeon. The operations are classified according to the surgical approach used and the degree of resection in order to facilitate use of the classification in daily practice. Each surgical procedure ensures that a specimen is available for histopathological examination. PMID- 10867841 TI - Morphometric analysis of the uvula in patients with sleep-related breathing disorders. AB - The upper-airway mucosa in obstructive sleep apnea (OSA) patients and snorers is often described as edematous and hyperplastic. The morphologic aspects of the pharyngeal mucosa, and in particular the mucosa of the uvula and soft palate, in OSA patients are, however, not well described. The aim of the present retrospective study therefore was to perform histologic examination of the pharyngeal mucosa obtained from patients with various forms of sleep-related breathing disorders, including primary snoring. A midsagittal section of uvulas obtained by uvulopalatopharyngoplasty (UPPP) was investigated in 34 OSA patients and 9 non-apneic snorers. Control tissues were taken by autopsy in 19 patients not known to have OSA or snoring. A morphometric point counting technique was used to determine the tissue composition. The data showed that OSA patients and non-apneic snorers had a significantly greater percentage of intercellular space than controls (65.7% vs 54.0%; P = 0.006). Control uvulas contained more muscle than OSA and snorers (14.0% vs 7.8%; P = 0.006). Moreover, the covering epithelium was significantly thicker in OSA and snorers than in controls (variance ratio = 7.64; P = 0.008). When taking body mass index (BMI) into account, no correlation was found between fat deposition and BMI. Findings showed that an increased clinical severity of OSA did not affect the tissue composition. Indeed, uvula morphology was similar in OSA patients with respect to non-apneic snorers. Since the increased amount of intercellular space is the expression of edema, we hypothesize that these mucosal changes together with hyperplasia of the covering epithelium are secondary effects to snoring. They presumably play a minor role in the etiopathogenesis of OSA, but may increase the severity of OSA by further narrowing the pharyngeal lumen. PMID- 10867842 TI - Therapy for respiratory tract infections caused by respiratory syncytial virus. AB - Respiratory syncytial virus (RSV) is the most common viral cause of lower respiratory tract infection (LRTI) in infancy and young children. No effective treatment for RSV lower respiratory tract infection (RSV-LRTI) exists. Ribavirin initially proved to be an effective anti-viral drug for RSV-LTRI. However, subsequently performed trials could not reproduce these positive results and, based on the current available evidence, there is no place for ribavirin in the routine treatment of RSV-LTRI. The use of nebulised bronchodilator therapy in RSV LTRI has been subject of many trials, with conflicting results. Although the individual patient may have some short-term benefit from nebulised bronchodilators, there does not seem to be a sufficient scientific basis for the standard use of bronchodilator therapy in infants and children with RSV-LTRI. There is increasing evidence that RSV-LTRI is an immune-mediated disease and therefore corticosteroids may be an effective treatment. The results from efficacy trials have demonstrated that corticosteroids are not effective for patients with mild RSV infection. In contrast there are indications that it may be beneficial in patients with more severe RSV-LTRI. It has been demonstrated that in children with RSV infection the vitamin A concentration is inversely related to disease severity. The use of vitamin A in the treatment of patients with RSV-LTRI, however, proved not to be effective. Immunoprophylaxis with hyperimmune immunoglobulins and monoclonal antibody against the viral F-protein have been shown to be effective in the prevention of RSV-LRTI. From the results of the therapeutic efficacy trials, however, it can be discerned that immunoglobulins have no place in the treatment of RSV-LRTI. CONCLUSION: Although respiratory syncytial virus infections each year have a considerable socioeconomic impact, attempts to find an effective therapy have so far been quite unsuccessful. Anti-viral therapy with ribavirin has not been proven to be effective. Symptomatic therapy with bronchodilators may give only short-term relief of symptoms in some individual patients, but has no effect on hospitalisation rates, or duration of hospitalisation. The beneficial effect of corticosteroids in patients with mild respiratory syncytial virus infection is very disappointing, however, there are indications that there might be an effect in patients with more severe infection. So far no beneficial therapeutic effect has been demonstrated with immune globulins. PMID- 10867843 TI - Treatment and prevention of respiratory syncytial virus infection. AB - This review discusses the current knowledge on treatment and prevention of respiratory syncytial virus (RSV) infections in children. Unfortunately, an effective therapy is not yet available. The efficacy of corticosteroids and bronchodilators has not yet been adequately documented and the use of ribavirin is only indicated in a highly selected group of high risk patients with T-cell immunodeficiency. The results of studies on the efficacy of vitamin A, interferon and antibiotics showed disappointing results. Vaccination research has produced candidate vaccines such as the recombinant vaccine BBG2Na, a subunit vaccine PFP 2 and cold-passaged-temperature sensitive vaccines. However, phase III efficacy trials in infants, young children and the elderly are still lacking. Passive protection against infections by RSV can be conferred by the use of RSV hyperimmune globulin or by the administration of palivizumab, a monoclonal antibody. However, large costs are involved. In addition, major differences have been reported in the prevalence of RSV lower respiratory tract infections in different countries, regions and even within well-known high-risk populations. CONCLUSION: We suggest the development of local and regional guidelines based on hospitalisation rates in high-risk infants and cost-benefit analysis studies. PMID- 10867844 TI - Malignant osteopetrosis obscured by maternal vitamin D deficiency in a neonate. AB - A neonate presented with clinical, biochemical, endocrine and radiographic features consistent with vitamin D deficiency rickets of maternal origin. Persistent hypocalcemia and subsequent development of pancytopenia, hemolysis and hepatosplenomegaly prompted further studies that led to the diagnosis of infantile osteopetrosis. CONCLUSION: Osteopetrosis is an important differential diagnosis of neonatal rickets and is not excluded by low vitamin D levels. PMID- 10867845 TI - Widening of the clinical spectrum of Bartonella henselae infection as recognized through serodiagnostics. AB - The recently improved diagnostics have widened, in children, the spectrum of clinical manifestations recognisable as Bartonella henselae infection. We report here the clinical features of 20 (14 males) consecutive children with serologically proved B. henselae infection observed within 12 months in the Paediatric Department of the University of Pisa. The patients had a mean age of 7 years 4 months (range 1.1-14.1 years). All children but one had a history of contact with kittens. Clinical manifestations included regional lymphadenopathy in 14 patients, representing in five the only clinical manifestation at onset, infectious mononucleosis-like syndrome in six, erythema nodosum in three, and Parinaud oculoglandular syndrome in one. In five patients a severe disorder was first suspected: fever of unknown origin in two with multiple hepatosplenic granulomatosis in one; osteolytic lesion suggesting bone neoplasm, marked inguinal lymph-node enlargement, suggesting Burkitt lymphoma, and an acute encephalopathy in one each. Bartonella henselae IgG antibody was positive in all patients with a titre ranging from 1:128 to 1:8590. IgM antibody was present in all except one child with an IgG titre of 1:2048. All patients recovered, some spontaneously. CONCLUSION: Bartonella henselae infection is frequent in Tuscany and probably underdiagnosed due to the high frequency of atypical onset of the clinical manifestations. An accurate clinical history and a reasonably wide use of the serological test may allow a rapid and accurate diagnosis, reassuring the family of the patient and avoiding invasive and expensive diagnostic procedures. PMID- 10867846 TI - Cord blood interleukin-10 levels are increased in preterm newborns. AB - Cell-mediated immunosuppression due to interleukin (IL)-10 may contribute to normal pregnancy. By contrast, delivery is associated with a predominance of T helper-1 (Th1) cytokines (IL-12, interferon-gamma) and might be regarded as a graft rejection process. The aim of the study was to assess IL-10 and IL-12 levels in cord blood samples from newborns and their normal mothers in relation to the gestational age and type of delivery. Cord blood and serum samples were obtained from 31 term newborns (gestational age 38-42 weeks) and 40 preterm newborns (mean gestational age 32 weeks). Serum samples were obtained from 26 mothers of term newborns at birth. There were 18 term and preterm infants born by caesarean section. Measurements of IL-10 and IL-12 levels by ELISA were repeated in mothers 15 days after delivery and in 11 preterm infants (median 14 days of age). Cord blood IL-10 levels were significantly higher in preterm than in term newborns (median 17.0 versus 3.2 pg/ml, P = 0.0001), but were similar to term newborns and paired mothers (2.2 versus 1.0 pg/ml). Term and preterm newborns also showed similar cord blood IL-12 levels (median 349 versus 320 pg/ml), and these levels were significantly higher when compared to their paired mothers (median 14.5 pg/ml, P = 0.0003). Cord blood IL-10 levels showed a significant inverse correlation with gestational age (P = 0.0001). When preterm infants, at several weeks post-delivery, were compared to gestational age matched newborns, their IL-10 levels were similar (median 8.3 pg/ml) whereas IL-12 levels were clearly lower (147 pg/ml; P = 0.0007). The type of delivery (vaginal versus caesarean) did not influence cord blood IL-10 and IL-12 results. CONCLUSION: Cord blood IL-10 levels are increased in preterm newborns and may be due to the immunosuppression occurring during pregnancy and to fetal immaturity because these levels are inversely correlated with gestational age. PMID- 10867847 TI - Serum vascular endothelial growth factor: a new predictive indicator for the occurrence of coronary artery lesions in Kawasaki disease. AB - We investigated serum vascular endothelial growth factor (SVEGF) levels in Kawasaki disease and determined whether these levels had any association with the development of coronary artery lesions. We measured SVEGF levels in 66 patients with Kawasaki disease, 18 patients with active infections and 18 afebrile controls. SVEGF levels of patients in the acute phase of Kawasaki disease (0.0 2003.6 pg/ml, median 59.87 pg/ml) were significantly higher than those of patients with active infections (0.0-45.2 pg/ml, median 8.10 pg/ml; P < 0.05) or afebrile controls (0.0-49.8 pg/ml, median 7.75 pg/ml; P < 0.05) and decreased to undetectable or low levels in the recovery phase (n = 31, acute phase: 0.0-2003.6 pg/ml, median 62.50 pg/ml versus recovery phase: 0.0-146.5 pg/ml, median 26.90 pg/ml; P = 0.0007) of the disease. There existed a positive correlation between SVEGF levels and serum C-reactive protein concentrations in the acute phase of Kawasaki disease (rs = 0.347, P = 0.0051). In addition, SVEGF level and duration of fever were found to be major risk factors for the occurrence of coronary artery lesions by univariate (P = 0.012 and P = 0.003, respectively) and multivariate (P = 0.037, OR 6.16 and P = 0.0059, OR 7.59, respectively) analyses. CONCLUSION: Serum vascular endothelial growth factor level, in combination with persistence of fever, could be a powerful predictor for the development of coronary aneurysms. PMID- 10867848 TI - Enalapril in paediatric patients with Alport syndrome: 2 years' experience. AB - Enalapril, a long-acting inhibitor of angiotensin-converting enzyme, was given for 2 years to seven children with Alport syndrome. Five patients had a classical X-linked form of the disease; two siblings had the autosomal recessive variant. Their age was between 5.15 and 13.75 years when enalapril was started. All patients had haematuria and proteinuria, creatinine clearance was > 80 ml/min per 1.73 m2 in all, and only one patient was hypertensive. The starting dose of enalapril (0.1 mg/kg body weight per day) was increased progressively according to individual clinical tolerance. The median doses were 0.13, 0.12, 0.21 and 0.29 mg/kg at 6, 12, 18 and 24 months, respectively. Median values of mean blood pressure were 95 mmHg at the start and 84 mmHg after 24 months. Median daily proteinuria decreased from 52 mg/kg to 18 mg/kg at 6 months, 21 mg/kg at 12 months, 12 mg/kg at 18 months and 30 mg/kg at 24 months. Serum creatinine increased over time from a median of 0.64 mg/dl at baseline to 0.77 mg/dl at 24 months. Concomitantly, there was a decrease in GFR from 104 to 83 ml/min per 1.73 m2 at 18 months and an increase again to 95 ml/min per 1.73 m2 at 24 months. Analysis of the individual data showed three patterns: no response (n = 2), temporary response (n = 2) and sustained response (n = 3). CONCLUSION: When given enalapril at the dosages mentioned, Alport patients as a group display a marked reduction in urinary protein excretion with a nadir of 23% of the baseline figure at 18 months, a decrease that cannot be accounted for by the slight decrease in glomerular filtration rate. Although these are preliminary data, it is recommended to try an angiotensin-converting enzyme inhibitor in every paediatric Alport patient with proteinuria. PMID- 10867849 TI - Elective use of nasal continuous positive airways pressure following extubation of preterm infants. AB - The aim of this study was to determine whether elective use of nasal continuous positive airways pressure (CPAP) following extubation of preterm infants was well tolerated and improved short- and long-term outcomes. A randomized comparison of nasal CPAP to headbox oxygen was undertaken and a meta-analysis performed including similar randomized trials involving premature infants less than 28 days of age. A total of 150 infants (median gestational age 30 weeks, range 24-34 weeks) were randomized in two centres. Fifteen nasal CPAP infants and 25 headbox infants required increased respiratory support post-extubation and 15 nasal CPAP infants and nine headbox infants required reintubation (non significant). Eight infants became intolerant of CPAP and were changed to headbox oxygen within 48 h of extubation; 19 headbox infants developed apnoeas and respiratory acidosis requiring rescue nasal CPAP, 3 ultimately were re-intubated. Seven other trials were identified, giving a total number of 569 infants. Overall, nasal CPAP significantly reduced the need for increased respiratory support (relative risk, 0.57, 95% CI 0.43-0.73), but not for re-intubation (relative risk 0.89, 95% CI 0.68-1.17). Nasal CPAP neither influenced significantly the intraventricular haemorrhage rate reported in four studies (relative risk 1.0, 95% CI 0.55, 1.82) nor that of oxygen dependency at 28 days reported in six studies (relative risk 1.0, 95% CI 0.8, 1.25). In two studies nasal CPAP had to be discontinued in 10% of infants either because of intolerance or hyperoxia. CONCLUSION: Elective use of nasal continuous positive airways pressure post-extubation is not universally tolerated, but does reduce the need for additional support. PMID- 10867850 TI - Bunk beds--a still underestimated risk for accidents in childhood? AB - A retrospective analysis of 218 bunk-bed accidents and a random sample survey with 991 family interviews were performed in order to establish guidelines for bunk-bed accident prevention. Falls from the top bed during sleep (35.1%) or while playing (34.4%) and falling off the ladder (23.2%) are the leading causes of bunk-bed accidents. Of the 218 children, 91 (41.7%) had sustained major injuries, including 3 polytrauma, 7 skull fractures, 44 cerebral concussions, 33 long bone fractures, 2 Lisfranc injuries, and 2 lacerations of the spleen. Of these accidents, 58.3% resulted in minor injuries with 18 fractures in other locations than the long bones or cranial vault, 89 contusions and sprains, 18 skin lacerations and 2 tooth fractures. A total of 23.8% of the accidents occurred in children under 3 years of age. The random sample survey demonstrated that in relation to age groups of children 30.8% (0%-45.8%) of families interviewed had been using bunk beds, with peaks at 3 years (29.8%), 7 years (36.5%) and 11 years (45.8%) of age. Of these bunk beds, 75.3% were equipped with side-rails, 57.3% had placed carpets alongside the bunk bed and 43.0% had used night lights. CONCLUSION: There is only one recommendation: no bunk beds!!! PMID- 10867851 TI - Apparent life-threatening events, facial dysmorphia and sleep-disordered breathing. AB - A standardized clinical evaluation and questionnaire was, beginning in 1985, applied to infants referred for an apparent life-threatening event (ALTE). All children who underwent this "core evaluation protocol" during a 10-year period were reviewed. Documentation of clinical complaints, symptoms and signs of sleep disordered breathing, sleep/wake evaluation, systematic evaluation of the face and naso-oro-pharynx, nocturnal polygraphic recording, and systematic follow-up was conducted. A total of 346 infants had complete data sets, with a smaller group of 46 age-matched healthy infants as controls. A scorer blind to the clinical data analyzed the polygraphic investigation and divided the 346 referred into two groups. Group A, 42.6% of the population, included infants with no abnormal findings based on nocturnal polygraphic recording. These infants were no different from controls at initial evaluation and during follow-up. Group B, 57.4% of the population, included infants who had obstructive breathing during sleep which became more obvious over time. Two-thirds of these infants not only had clinical symptoms of sleep-disordered breathing but also had mild facial dysmorphia that could be seen clearly at 6 months of age. CONCLUSION: A subgroup of infants with apparent life-threatening events present an indication of a sleep disordered breathing syndrome which is associated with a mild dysmorphia. This mild facial dysmorphia needs to be recognized early to distinguish these infants from other infants with apparent life-threatening events and to initiate appropriate treatment. PMID- 10867852 TI - Severe neonatal group A streptococcal disease. AB - Since the mid-1980s, an increase in incidence of invasive disease caused by group A streptococci has been noted amongst adults and children; however, neonatal disease is still rare. Between 1979 and 1998, seven neonates with severe group A streptococcal disease were admitted to our neonatal intensive care unit. The clinical presentation, treatment and outcome are described. In three cases of early-onset disease vertical transmission was documented. CONCLUSION: Because the incidence of group A streptococcal disease in the general population seems to have increased over the last two decades, we should be aware of the possibility and particularly the severity of group A streptococcal disease in the neonatal period. PMID- 10867853 TI - Disseminated cytomegalovirus infection as initial manifestation of hyper-IgM syndrome in a 15-month-old boy. AB - We report on the clinical, laboratory, and molecular genetic findings in a 15 month-old boy who initially presented with disseminated cytomegalovirus and concomitant para-influenza virus infection. Hyper-IgM syndrome, suspected on clinical grounds, was confirmed by immunological investigations. In addition, a previously unreported potentially disease-causing mutation in the CD40 ligand gene was detected in this patient. CONCLUSION: The present case illustrates that disseminated cytomegalovirus infection with atypical clinical features should be included in the spectrum of the hyper-IgM syndrome. PMID- 10867854 TI - Prevalence of thyroid hemiagenesis: ultrasound screening in normal children. AB - While performing a systematic ultrasound study of the thyroid gland volume for the evaluation of iodine deficiency in 2845 normal Belgian school children, we found an absence of the left lobe in 6 children (4 girls and 2 boys). There was no association with other thyroid malformations or dysfunction. CONCLUSION: This first systematic ultrasound evaluation of thyroid hemiagenesis in normal children established a prevalence of thyroid hemiagenesis of 0.2% and confirmed the female predominance and higher incidence of agenesis of the left lobe. PMID- 10867855 TI - Prediction of permanent hearing loss in high-risk preterm infants at term age. AB - The aim of this series was to assess hearing screenings; auditory brainstem responses (ABR), transient evoked otoacoustic emissions (TEOAE) and free field auditory responses (FF) for the prediction of permanent bilateral hearing loss in high-risk preterm infants at term post-conceptional age. A total of 51 preterm infants (gestational age < 34 weeks, birth weight < 1500 g) underwent examinations at term and hearing, speech and neurological development were followed up until a corrected age of 18 months. Significant hearing defects were verified by broader ABR examinations under sedation and by clinical ward observation including responsiveness to sounds and enhancement of hearing using an amplification device. Seven bilateral fails in ABR were found, together with nine bilateral fails in TEOAE and four fails in FF screening at term age. Six preterm infants were later confirmed to have a significant permanent bilateral hearing loss, four of whom had also cerebral palsy. Bilateral failure in ABR screening predicted hearing loss with a sensitivity of 100% and a specificity of 98%, TEOAE with a sensitivity of 50% and a specificity of 84% and in the FF examination at the levels of 50% and 98%, respectively. CONCLUSION: Transient evoked otoacoustic emissions alone seem not to be so applicable to the neonatal screening of hearing in high-risk preterm infants as shown earlier in full-term infants, possibly because a hearing defect may be due to retrocochlear damage. Consequently, auditory brainstem response screening seems to be more suitable for very low birth weight preterm infants. PMID- 10867856 TI - Serum inhibin B in normal term-born male and female neonates during the first week of life. AB - Inhibin B, a gonadal peptide regulating follicle stimulating hormone (FSH) secretion in adults, has been found during gestation in amniotic fluid, but at birth only in term cord blood of male babies. Since no data are available on the evolution of serum inhibin B during the 1st week of life, we studied changes in inhibin B using a specific and sensitive immunoassay in male and female neonates during the 1st week of life in relation to FSH and to evaluate the possible effect of perinatal factors on inhibin B production. Inhibin B was measured by a specific monoclonal enzyme-linked immunosorbent assay. Inhibin B was detectable in cord blood of all eight longitudinally studied male newborns, correlated negatively with the ponderal index and increased significantly on day 5 (from 54.2 +/- 18.5 to 100.4 +/- 34.8 ng/l, P < 0.005). Cord blood inhibin B was detected in only 1 out of 13 screened female neonates. In 48 at term-born females in whom inhibin B was measured on the 5th day of life, only 20 cases had a detectable level (between 8 and 68.6 ng/l). Inhibin B concentrations in cord blood and on day 5 were independent of duration of pregnancy, type of delivery, Apgar score and FSH concentration. CONCLUSION: A sexual difference in serum inhibin B is already present at the end of gestation and changes in inhibin B during the 1st week of life are independent of follicle stimulating hormone changes and perinatal factors in both sexes. Our data suggest that neonatal inhibin B could be used to study whether the newborn has functional testes, i.e. in babies with ambiguous genitalia and/or bilateral cryptorchidism. PMID- 10867857 TI - Linkage disequilibrium of the common mutations 677C > T and 1298A > C of the human methylenetetrahydrofolate reductase gene as proven by the novel polymorphisms 129C > T, 1068C > T. PMID- 10867858 TI - [A life devoted to science (Mykola Mykolaiovych Horiev)]. PMID- 10867859 TI - [The role of endothelium-dependent factors in realizing cardiogenic reflexes under normal and pathological conditions]. AB - We have investigated the role of NO-dependent mechanisms in the realization of cardiogenic reflexes (pressor and depressor) during the action of veratrin and serotonin on cardiac receptors, adrenergic changes of the myocardial contractile activity or the short-term period of myocardial ischemia on anaesthetized dogs cats and rats. NO take place in the development of cardiogenic depressor reflexes and decreases pressor reflexes from the cardiac receptors in dogs. Specific characteristics of these influences were asserted: the depressor reflexes decrease after systemic inhibition of NOS by L-NNA in dog but they are increased or are not altered in rats. The depressor reaction in rats diminished after inhibition of nNOS performing by 7-nitroindazole. We affirm that NO-dependent influences have a compensatory effect in the course of acute alterations of the coronary blood flow and our statement concerning the advantageous role of NO dependent mechanisms in the realization of self-regulatory reactions of circulation that connected with vagus nerve. PMID- 10867860 TI - [The role of nitric oxide in changes in the oxygen consumption and the oxygen cost of the work of the cardiac muscle]. AB - In experiments on guinea-pig isolated hearts, perfused under Langendorff preparation, it has been shown that heart volume load and inotropic stimulation were accompanied with an increase of nitric oxide synthesis (by 46 and 51% accordingly) and with a decrease in an oxygen cost of myocardial work. L-arginine or sodium nitroprusside administration resulted in a reduce of the oxygen consumption by 29% and a decrease of oxygen cost of myocardial work. Inhibition of NO-synthase activity lead to an opposite effect--an increase in both--O2 consumption and an oxygen cost of the heart work by 15 and 19%. The data obtained evidence that NO is a regulator of the oxygen consumption by tissues and it determines the oxygen cost of the myocardial work. Its lack can play an important role in the developing of pathological states of the cardiovascular system, which are followed with its insufficient synthesis. PMID- 10867861 TI - [The hypothalamic and brain stem mechanisms in the development of arterial hypertension during aging]. AB - Effects of stimulation of hypothalamic and brain stem nuclei on arterial blood pressure were studied in adult and old rabbits and rats. In aging, thresholds of pressor reactions due to stimulation of anterior and posterior hypothalamic areas were decreased, while thresholds of depressor reactions didn't change. Long-term stimulation of nuclei of anterior and posterior hypothalamic areas induced stable hypertension, more pronounced in old animals. In old rats, as compared to adult ones: pressor responses due to stimulation of N. tr. solitarius developed more often and thresholds of these reactions were lower; amplitude of pressor responses due to stimulation of locus coeruleus was higher; changes in ventromedial hypothalamic nuclei electrogram were induced by lower electric stimulation of locus coeruleus; pressor reactions due to intracerebroventricular norepinephrine injections developed more often; ouabain microinjections into cerebral ventricles and ventromedial hypothalamic nuclei induced higher pressor reactions. Age-related changes in hypothalamic and brain stem mechanisms of blood pressure regulation may be of importance in development of arterial hypertension in old age. PMID- 10867862 TI - [The hormonal factors regulating water-electrolyte exchange in the pathogenesis of the hemodynamic disorders in pre-eclampsia]. AB - Study of indices of different endocrine links of female organisms with gestosis of the 2nd half of pregnancy with the search of degree of taking part of this hormones in the forming of basic syndromes of hestosis and guiding line to fundamental approach in medicinal practice with gestosis. Decrease of natrium content in CBV at the same time with decrease of intravascular volume of it and decrease of excretion of it with urine in the dynamics of gestosis confirms hestosis development and retention of natrium in the expectorant women's organism with primary delay of it in tissues. Energization of renin-angiotensin aldosterone-tromboxane system, changes of prostaglandin-tromboxane link in blood contributed to forming of arterial hypertension and edema. With taking into account revealed changes of hormonal levels at the background of progressive hypovolemia, decrease of speed of excretion of natrium with urine and speed of glomerular filtration, differentiate approach in prescription for hypotensive and diuretic agents is recommended for the avoidance of possible hardening of state of expectorant woman with gestosis. PMID- 10867863 TI - [The nitric oxide system in a chronic deficiency of mesostriatal dopamine: the action of nitroglycerin]. AB - Unilateral chronic deficiency in dopamine was induced by injection of the neurotoxin 6-hydroxydopamine into the mesostriatum of male Wistar rats (n = 15). The content of the nitric oxide metabolite--NO2- was three time less than in controls in the neostriatum on the operated side only, whereas, in the heart, aorta and plasma it was reduced by 1.9, 1.7 and 3.2 times, respectively. Four hours after subcutaneous injection of nitroglycerin (10 mg/kg, n = 4), a significant 6 fold increase in NO2- content in the neostriatum in the operated hemisphere and 2 fold increase on the in the intact side was observed. In heart, aorta and plasma NO2- content increased 2, 9, 3 times, respectively. The same dose of nitroglycerin given to intact rats (n = 4) had less effect on NO2- content: a 2 fold increase in heart, aorta and plasma only. Activity of NO synthase (NOS) in animals with dopamine deficiency was reduced 2.7 fold in the neostriatum of the operated hemisphere and by 1.8 time on the intact side, in heart and aorta--by 1.8 and 3 time, respectively. Injection of nitroglycerin increased activity of NOS in the neostriatum of the operated hemisphere 2 fold and on the intact side 3 time; in heart (11 time) and aorta (1.3 time) after 4 hours. Similarly subcutaneous nitroglycerin injection restored the reduced endothelium-dependent vasodilator responses characteristic of the rats with mesostriatal dopamine deficiency. Thus the amplitude of acetylcholine-induced smooth muscles relaxation of the aorta increased by more than 70%. In these animals the injection of nitroglycerin also exerted a normalizing effect on the characteristic apomorphine-induced behaviour asymmetry (rotatory movements). On week after the injection of nitroglycerin the rotatory behaviour was reduced by 20.7% of control rotations. We show that nitroglycerin, a NO-donor, is a useful drug. It correct the abnormalities in the metabolic pathways on NO synthesis in a variety of tissues, endothelium-derived vascular responses and also the behaviour abnormalities induced by unilateral mesostriatal lesions. Our experimental animals are thought to be a model of human hemi-Parkinsonism. Thus nitroglycerin might be of use in Parkinson's disease. PMID- 10867864 TI - [The current concepts of the pathogenesis of Monckeberg-type arteriosclerosis]. AB - On the basis of experimental researches the circuit of pathogenesis of Monckeberg's arteriosclerosis--one of the most widespread types of arteriosclerosis is offered. The most important mechanisms realized during development of this pathological process, following: 1) actions of the injuring factor cause necrobiotic change of vascular wall cells; 2) activation of proteolytic enzymes, disturbance of balance between proteinases and their natural inhibitors results in destruction of extracellular matrix fibers and excessive stimulation by products limited proteolysis of vascular wall cell; 3) infringements of intracellular ion homeostasis at the expense of the mentioned below factors exhaust energy systems (last in case of metabolic poison action can be primary are damaged); 4) infringements in system of blood, in particular, in calcium homeostasis, blood coagulation, lipid spectrum promote additional damage of vascular wall cells; 5) dystrophic and metastatic calcification of media; 6) development "unspecific mesenchymal reaction", including fibrosis and sclerosis, in damage zone of vascular wall. PMID- 10867865 TI - [Modified lipoproteins--their types and role in atherogenesis]. AB - In the performed clinical and experimental investigation there were determined the significance of lipoprotein modification in atherogenesis and the mechanisms of blood atherogenicity development. The pronounced changes of total blood cholesterol and the spectrum of lipoproteins in patients with coronary atherosclerosis were not regular and were noted only in 40-55% cases. The most regular was the increasing of blood atherogenicity, reflecting the appearance of the great amount of modified lipoproteins. One of the most important modifying factors was oxidative stress induced by the activation of blood inflammatory cells. These data were confirmed in investigations on children with acute respiratory inflammatory diseases and on rabbits with the models of acute inflammation. The other important mechanism of blood atherogenicity was the increasing of blood lipoproteins enriched with triglycerides; that led to secondary monocyte activation and oxidative stress development. The usage of lipid-lowering drug lipantil in coronary patients not only decreased the amount of cholesterol and tryglicerides in blood but also lowered blood atherogenicity and suppressed the oxidative stress. Thus the results show the existence of at least two ways of atherogenic lipoprotein formation--in the course of oxidative modification and under the enrichment with tryglicerides as a result of the lipolysis disturbances with subsequent slowing of the formation and elimination of these lipoprotein remnant forms from blood. PMID- 10867866 TI - [The correction of arterial hypertension by oxygen deprivation]. AB - Curative effect of oxygen deprivation has been examined in 147 patients with initial studies of hypertonic disease. It was used the intermittent model of the oxygen deprivation. The oxygen partial pressure in the breathing gaseous mixtures was 90-110 mm Hg. The positive effect was achieved in 79% of patients. Arterial pressure decreasing at the expense of different hemodynamic mechanisms depending on the type of blood circulation. It is proposing to use the oxygen deprivation as the correction method for the initial studies of the arterial hypertension. PMID- 10867867 TI - [The role of voltage gated K(+) channels in the modulation of resting membrane potential of myocytes isolated from rat resistance arteries]. AB - K+ current which take part in the controlling of membrane potential in myocytes isolated from rat resistance mesenteric arteries have been investigated using conventional patch clamp method. The mean resting potential of myocytes was--37 mV. Charybdotoxin (200 nM)--selective blocker of large conductance Ca(2+) activated K+ (KCa) channels--inhibited transmembrane outward K+ current by 60%. 1 mM of tetraethylammonium inhibited outward K+ current same as 200 nM of charybdotoxin, also it inhibited spontaneous spike-like hyperpolarizations and did not affect the membrane potential. Transmembrane current had a 4 aminopyridine (4-AP) sensitive component of delayed rectifier current (KV). Addition of 5 mM of 4-AP evoked membrane depolarization with mean significance of 12.0 +/- 1.5 mV in 5 from 7 single myocytes which had resting potential in the range of -50 ... -35 mV. The obtained results suggest that large conductance KCa channels do not determine the resting potential, but may serve as a negative feedback mechanism at the considerable membrane depolarization. In contrast, 4-AP sensitive KV current take part in the controlling of the resting membrane potential of single myocytes from rat resistance mesenteric arteries. PMID- 10867869 TI - [The mechanisms of the GABA-ergic regulation of beta-endorphin levels in the hypothalamic structures of prenatally stressed male rats]. AB - The effect of GABA receptors agonists on the stress-induced beta-endorphin levels in the preoptic area and mediobasal hypothalamus of the intact and prenatally stressed male albino rats was studied. It has been found out that stimulation of GABAa-receptor complex by means of the muscimol leads to increasing of beta endorphin levels in the preoptic area and mediobasal hypothalamus of the control animals. GABAb receptor activation by means of the baclofen decreases opioids level in the mediobasal hypothalamus. Prenatal stress eliminates stimulant effect of the muscimol on beta-endorphin levels in the investigated brain structures and leads to the opioid level decreasing after baclofen influence in preoptic area. PMID- 10867868 TI - [The protective effect of omega-3 polyunsaturated fatty acids on the activity of the isolated rat heart during myocardial ischemia-reperfusion]. AB - It was produced plant-derived product, an omega-3 acid-enriched substrate (64%). In our study we tested the influence of this preparation, which is supposed a membrane-modifying agent, on the processes of damage to the isolated heart under conditions of ischemia-reperfusion. Animals took this substrate as nutrient addition to usually everyday diet. We assumed disorders in cardiodynamics and contractile functions of the myocardium (we measured a perfusion pressure in coronary vessel, left ventricular pressure and dp/dt) and in structure of cardiomyocytes. All mentioned parameters was much better after ischemia reperfusion in hearts from animals which took an omega-3 acid-enriched substrate in course of 4 week before experiments than in hearts from control animals. Conclusions. Omega-3 polyunsaturated acids exert protective effect on functioning and structure of the isolated rat heart during ischemia-reperfusion. PMID- 10867870 TI - [The effect of medikhronal on the psychophysiological and metabolic processes in alcoholism]. AB - Pathopsychological examination, biochemical analysis of blood and urine as well as electroencephalogram (REG) were made in patients of the second (II) stage of alcoholism. The state of higher nervous activity (HNA) before and after treatment with antialcoholic drug medichronal was also analyzed. It has been shown that the course of treatment with medichronal applied to patients with alcoholism results in the removal of the inclination to alcohol, to improvement of functional state of the central nervous system, as well as ethanol (E), acetaldehyde (AcA) and catecholamines (CA) exchange, the disturbances in their metabolism (and first of all AcA) apparently being the basis of pathogenetic mechanisms of alcoholism. PMID- 10867871 TI - [The role of modified low-density lipoproteins in physiology and pathology]. AB - Various kinds of low-density lipoprotein modification, its physiological and pathological role are characterized. Special attention has been given to the modified lipoproteins by peroxidation and their impact in the formation of atherosclerotic lesions. The mechanisms of its development are considered at cellular-molecular level. PMID- 10867872 TI - [Age and the vascular reaction to injury]. AB - Our own results and the literature dates analysis showed that spontaneous endothelial cells injury in aging, and the hypertension, hypercholesterinemia, immune disorders, that was induced in experiment aggravating the process of the development of atherosclerosis. PMID- 10867873 TI - [The perinatal stress modification of the reactivity of the hypothalamo hypophyseal-adrenal system]. AB - The literature data and the results of authors research work concerning long-term effects of prenatal stress and of early postnatal handling on the hypothalamic pituitary-adrenal stress response are reviewed in the article. Functional state of that system, and the role of hormones, biogenic monoamines and glucocorticoid receptors of the brain in pathogenesis of its alteration are discussed. Special reference is made to gender- and age-related effects of pre- and postnatal stress. The body of evidences supports basic states of the brain hormone neurotransmitter imprinting during early ontogenesis. PMID- 10867874 TI - [Multilocus sequencing--a new method of genotyping bacteria and first results of its use]. AB - Comparative characterization (molecular typing) of isolates within a bacterial species is one of the major problems in microbiology and epidemiology. However, it is rather difficult to correlate data obtained in various laboratories, because traditional, including molecular, methods employed in typing pathogenic microorganisms cannot be standardized. In 1998, Maiden et al. proposed multilocus sequence typing (MLST); through which alleles of several housekeeping genes are directly assessed by nucleotide sequencing, each unique allele combination determining a sequence type of a strain. The advantages of this approach are that the culturing of pathogenic microorganisms is avoided, as their gene fragments are amplified directly from biological samples, and that the sequencing data are unambiguous, easy to standardize, and electronically portable. The latter makes it possible to generate an expandable global database for each species at an Internet site, in order to use it for the purposes of genotyping pathogenic bacteria (and other infectious agents). MLST protocols have been elaborated for Neisseria meningitidis, Streptococcus pneumoniae, and Helicobacter pylori; those for Streptococcus pyogenes, Staphylococcus aureus, and Haemophilus influenzae are now being developed. Basic principles and the first results of MLST have been reviewed, including data on the distribution and microevolution of N. meningitidis clones causing epidemic meningococcal infection, the relative recombination and mutation rates in the N. meningitidis genome, the identification of antibiotic-resistant S. pneumoniae clones causing severe generalized infection, the grouping of H. pylori isolates from various geographic regions, etc. PMID- 10867875 TI - [Expression in vivo of the lacZ gene, delivered to mouse lungs using synthetic microspheres]. AB - A capacity of MF-2 synthetic microspheres to serve as the vehicle for transfer of the marker LacZ gene to mouse lung epithelial cells was studied after a single intranasal administration of the MF-2/gene complex. Two types of plasmids carrying marker gene LacZ were used in the experiments: with cytoplasmic (pCMV LacZ) and nuclear (pCMV-nlsLacZ) localization of the gene product (beta galactosidase). As early as 7 days after the complexes MF-2/pCMV-LacZ and MF 2/pCMV-nlsLacZ were administered, specific staining for beta-galactosidase revealed this enzyme activity in the epithelial cells of bronchi, bronchioli, and alveoli. The maximum in vivo of the marker gene in the MF-2/pCMV-LacZ complex was observed at day 14 to 21 after administration and the corresponding gene product was detected during the following two months. The MF-2-mediated gene transfer led to a twofold increase in beta-galactosidase activity relative to the case when the "unbound" pCMV-LacZ plasmid was administered. These results suggest that the synthetic microsphere-mediated transfer of alien genes to the lung of experimental animals is promising. Microspheres may be used in gene therapy for pulmonary affections, in particular cystic fibrosis. PMID- 10867876 TI - [Cloning of a new murine gene coding for a protein immunologically related to RecA protein from Escherichia coli]. AB - Using the testis-specific murine cDNA library immunoscreening with the affine purified polyclonal antibodies to the E. coli RecA protein, the 1730-bp fragment of the novel mouse gene was cloned. Northern hybridization of total RNA samples from mouse somatic and meiotic tissues showed that this gene was specifically expressed in mouse testis, producing a transcript of about 10 kb in size. Analysis of the cloned cDNA sequence revealed the presence of the-AATAAA polyadenylation site at its end, indicating that the cloned fragment represented the end part of the corresponding gene. Comparative sequence analysis revealed that the plus-chain of the cloned fragment contained motifs of the mouse RAD50 gene; the 378-bp fragment of the minus-chain showed 96% homology with the Homo sapiens Hd741-f cDNA fragment; and the whole minus-chain was by 50% homologous to the prokaryotic FTSZ/A genes. The cloned fragment contained two reading frames located in the plus- and minus-chains, respectively. The presumptive 44-kDa protein encoded by the plus-chain (sense) open reading frame contained serine-, proline-, and threonine-rich regions and was 25 to 30% homologous to a number of nuclear DNA-binding proteins of eukaryotes. Although analysis of the similarity between the 44-kDa protein and the E. coli RecA protein did not show any significant homology between them, it revealed their identity by five amino-acid residues involved in the formation of the epitope that recognized the paratope of the RecA protein antibody for subsequent epitope-paratope binding of these proteins. PMID- 10867877 TI - [TAR-cloning of the short arm of human chromosome 7 in yeast and search for terminal sequences]. AB - A partial clone library of the short arm of human chromosome 7 was created in yeast artificial chromosomes (YAC) using TAR-cloning. The DNA of monochromosome somatic hybrid cells (mouse/human) RuRag 14-4-7-44 containing short arm human chromosome 7 was used for cloning. The clone library was screened for YACs with the human DNA; the mitotic stability of these YACs, the sizes of cloned fragments, and an independent clonal distribution in the chromosome were determined. Human YACs were tested for the presence of chromosome 7p telomeric sequences. PMID- 10867878 TI - [Insertion of transposon P(lacW) with gradual expression of reporter genes]. AB - In Drosophila, insertions of the P?lacW? transposon led to uncommon expression patterns of the reporter genes white and lacZ: gene activity was gradually reduced in the neighboring cells along the dorsal-ventral or proximal-distal axes. Using in situ hybridization, the gradual gene expression was shown to be caused by P?lacW? insertions in regions 40A (lines G1 and G2) and 69C (line G3) of the Drosophila genome. The expression patterns of genes lacZ and white were similar in eye cells of each line. In salivary gland polytene cells of G3 flies, the lacZ expression was also gradually reduced, which indicates the existence of a mechanism maintaining the gene activity gradient during endomitosis. PMID- 10867879 TI - [Functional characteristics of the promoter region of the tag7/PGRP gene in KSML0, KSML100 murine mammary adenocarcinoma cell ines and VMR liver]. AB - A study was carried out on the transcription regulation of the tag7/PGRP gene, whose product is similar to cytokines and is involved in recognizing peptidoglycane in mouse mammary adenocarcinoma cell lines KSML0, KSML100, and VMR liver. The 3250-bp fragment of the promoter region was sequenced and tested for DNase I-hypersensitive sites (DHSs). In the KSML0 cells line, DHSs were found in the vicinity of the TATA box and approximately at position -3000 relative to the transcription start of the gene. Only a DHS cluster near the TATA box was found in the VMR-liver cell line. Regions involved in transcription regulation of the gene in the three cell lines were identified with the use of reporter constructions carrying the CAT gene under the control of deletion derivatives of the tag7/PGRP promoter region. The minimal promoter including only the TATA box with the nearest neighboring sequences was inactive in all cell lines. The elements (enhancers) positively regulating gene transcription in KSML0 and VMR liver were mapped to region -192, -48. An element accounting for the transcriptional inactivity of the gene in KSML100 cells was assigned to region 315, -3250. PMID- 10867880 TI - [Interaction of ATP17 gene with SUP45 and SUP35 genes in Saccharomyces cerevisiae yeast]. AB - Genes SUP35 and SUP45 have been identified in the saccharomycete yeast as genes controlling termination of translation in cytoplasmic ribosomes. However, many facts indicate that the control of translation termination is not the only function of these genes. This work is devoted to studying one of the pleiotropic effects of sup35 and sup45 mutations, a respiratory deficiency. The compensation for this deficiency in mutants for either gene can occur due to a mutation in the ATP17 gene encoding the f-subunit of mitochondrial F1F0 ATP synthase. It is assumed that the observed interaction can be related to the system of co translational protein import into mitochondria. PMID- 10867881 TI - [Assessment of genetic polymorphism in Colorado potato beetle Leptinotarsa decemlineata 8Say) by RAPD markers]. AB - Four decanucleotide primers were chosen to detect polymorphism for randomly amplified polymorphic DNA (RAPD) in the populations of Colorado potato beetle. The variability of RAPD patterns was studied in three oblasts (Kiev, Moscow and Kurgan) that are geographically distant and considerably differ in ecological and climatic conditions affecting the development of this beetle: Kiev, Moscow, and Kurgan oblasts. A high level of intrapopulation polymorphism exceeding interpopulation differences was revealed. RAPD-marker subsets differing in the frequency and distribution throughout the studied populations were revealed. PMID- 10867882 TI - [Taste sensitivity in homeotic mutants of the drosophila leg-arista-wing complex]. AB - It is well established that taste chemoreceptors in Drosophila are located on the tarsi of the first leg pair. In order to investigate the influence of the novel homeotic arista-tarsus transformation on behavior, an analysis of taste perception in the lawc-mutants, characterized by the transformation of the arista into tarsus elements, was carried out. The data were subjected to thorough statistical treatment. It was shown that elements of an additional leg that appeared as a result of homeotic transformation of the arista were sensitive to gustatory stimuli. Analysis of the innervation of the homeotic organs by means of cobalt staining of afferent projections showed that the afferents starting from the homeotic leg reached the thoracic ganglion. PMID- 10867883 TI - [Genetic diversity of reaction of common wheat (Triticum aestivum L.) cultivars to light intensity]. AB - The effect of low light intensity (LI) on the period from sprouting to earing was studied in 12 cultivars of the spring common wheat under controlled conditions. Differences between cultivars with respect to their responses to LI (RLIs) were found both for those that were photoperiod-sensitive and those that were almost photoperiod-neutral. Specifically, a prolonged photoperiod and a low LI differently increased the period from sprouting to earling in different cultivars. Genetic analysis of the RLI demonstrated, for the first time, that the weak response was incompletely dominant in F1. The results of genetic analysis agree with the hypothesis that the cultivars Pitic 62 and Novosibirskaya 22 differ in alleles of two loci controlling the RLI in wheat. PMID- 10867885 TI - [Genetic determination of stem-rust resistance in rye]. AB - The harmful effect of stem rust on the crops of short-stem diploid winter rye was studied. If stem rust affected the plants by 70-100%, this decreased the mass of 1000 grains by about 35.8%. The genes that control the stemrust resistance of rye might originate from the following cultivars and forms: Ilmen, Orlovskii Gibrid, Kharkovskaya 55, Kharkovskaya 60, Kustovka, Kombaininyai, Kazanskaya, Krupnozernaya, Novozybkovskaya 4, Alfa, Derzhavinskaya 29, Chulpan, and Rossul, as well as wild populations of the perennial rye Secale montanum. This study was first to demonstrate that the resistance of the Kharkovskaya 55 and Rossul rye cultivars to the population of stem rust was controlled by a single dominant gene, which was designated Sr1. PMID- 10867884 TI - [Chromosome maps of the plant family Trilliaceae: nucleotide composition of heterochromatin an localization of 18S-26S rRNA-genes of four-leafed paris (Paris quadrifolia L.)]. AB - Using nucleotide-specific agents Hoechst 33258, actinomycin D, chromomycin A3, and distamycin A, the Paris quadrifolia L. karyotype, and the location and nucleotide composition of heterochromatic bands were studied. The chromosome ideogram of H33258/AMD and CMA/DA heterochromatic bands was created by an image analysis system with the Videotest-Kario software. By fluorescence in situ hybridization, the 18S and 26S rRNA genes were mapped. PMID- 10867886 TI - [RAPD-analysis of genetic polymorphism of ducks: differences in breeds]. AB - Using randomly amplified polymorphic DNA (RAPD) obtained by polymerase chain reaction (PCR), the genetic differentiation of breeds of Peking and Musk ducks maintained at the Blagovarskii State Farm for Pedigree Poultry was studied. A comparison of genetic distances estimated by two different methods, as well as similarity dendrograms based on these estimates, was performed. The similarity dendrograms obtained using different primers and methods of construction were found to be similar. The pattern of breed clustering on these dendrograms adequately reflects their actual state known from the history and genealogy of breeds. PMID- 10867887 TI - [Interaction of polymorphic systems in a population of Semirechenskaya swine breed]. AB - Associations between the alleles of nonlinked polymorphic genetic systems were studied in the population of the Semirechenskaya breed of pigs. The study of marker pairs of 21 genetic systems revealed that 7% of associations are statistically significant. Most of the associations detected are assumed to arise from breeding by commercial productivity traits and from the formation of an integral genotype adapted to conditions of artificial selection in the population. PMID- 10867888 TI - [Variability of GATA-microsatellite DNA in populations of the parthenogenetic species of lizard Lacerta unisexualis Darevsky]. AB - Multilocus DNA fingerprinting was used to analyze the genome variation of mini- and microsatellite DNA regions in parthenogenetic Caucasian rock lizard Lacerta unisexualis. The DNA fingerprints obtained with probe M13 were nearly identical in all populations examined (the average similarity index S = 0.992). The fingerprints obtained with probe (GATA)4 varied (S = 0.862). Polymorphic fragments were assumed to correspond to allelic variants of genetically unstable GATA loci. Comparison of the fingerprints of animals from four geographically isolated populations revealed several population-specific GATA microsatellite markers. Based on their distribution among the populations, the corresponding alleles were assumed to originate from a common ancestral allele. PMID- 10867889 TI - [Molecular genetic study of the factor VIII gene in families from Bashkir with hemophilia A]. AB - The distribution of hemophilia A was studied in Bashkortostan. The factor VIII gene of the blood coagulation system was analyzed in 34 patients with hemophilia A and 48 of their close relatives. Inversion of intron 22 of the factor VIII gene was revealed in nine cases, which comprised 30% of the total sample analyzed. The type II and type III of this mutation occurred at a relatively high frequency, which may be explained by the founder effect and genetic drift. The allelic frequencies of the polymorphic locus HindIII at intron 19 were similar; a substantial allelic heterogeneity of both microsatellite (CA)-repeats at intron 13 and the DXS52 locus were found on normal and mutant X chromosomes. The molecular genetic analysis of (CA)-repeats and the loci HindIII and DXS52 in families with hemophilia A makes it possible to reveal up to 89% of the informative families. PMID- 10867890 TI - [Molecular genetic analysis of polymorphism of the HindII and (CA)-repeat of the factor VIII gene loci in people of the Volgo-Ural region]. AB - Molecular genetic analysis of polymorphism of the factor VIII gene at loci HindIII (intron 19) and (CA)-repeats (intron 13) was performed in seven ethnic groups of the Volga-Ural region (Bashkirs, Tatars, Chuvashes, Maris, Mordovians, Udmurts, and Komis). The allelic frequency of the HindIII locus was shown to be similar in these ethnic groups. The ethnic groups were more heterogeneous with regard to the genotype and allele frequencies of (CA)-repeats, which indicates that this polymorphic system can be used to characterize the genetic structure of the Volga-Ural human populations. PMID- 10867891 TI - [Study of the most often encountered allele of the HLA-B27 gene in Tuvinian and Russian inhabitants of Western Siberia]. AB - HLA-B27 gene frequencies and allelic polymorphism were studied in two Siberian ethnic groups: Russians from Novosibirsk (western Siberia) and Tuvinians from Kyzyl (southern Siberia). The HLA-B27 frequencies were determined by means of serologic typing of HLA antigens in 198 Tuvinians and 288 Russians. Molecular typing was performed via hybridization of oligonucleotide probes with amplified DNAs obtained from 30 HLA-B27-positive Russians and 11 HLA-B27-positive Tuvinians. The HLA-B27 gene frequencies in Tuvinians and Russians were 5.5 and 10.4%, respectively. Molecular variants of the HLA-B27 gene were studied in Tuvinians for the first time. The proportions of the HLA-B2705 and HLA-B2704 alleles were found to be 64 and 36%, respectively, in the population studied. The presence of the HLA-B2704 allele indicates a Mongoloid origin of Tuvinians. In the Russian population of Novosibirsk, the HLA-B2704 allele was not found, whereas the proportions of the HLA-B2705 and HLA-B2702 alleles were 76.2 and 23.8%, respectively, which is characteristic of Caucasoid populations. PMID- 10867892 TI - [Chromosomal instability in offspring of voles in unfavorable radiation zones]. AB - A high frequency of cells having chromosomal aberrations was shown to be preserved in the first generation of laboratory offspring of common voles, whose ancestors were captured near the East Ural radioactive track (EURT). In F2 and F3, a decrease in frequency was observed. Most likely, the cytogenetic effects observed were caused by environmental contamination with plutonium in the locality of capture. An increased chromosomal instability observed in the first laboratory generation of nonirradiated common voles was caused by reversible modifications in the expression of mutator genes inherited from parents exposed to alpha-irradiation rather than by classic mutations occurred in specific loci. PMID- 10867893 TI - [Analysis of the 32 bp deletion in the CCR5 chemokine receptor gene in people from Moscow, infected with HIV-1]. AB - Chemokine receptors have recently been shown to mediate HIV-1 entry into cells. The chemokine receptor CCR5 plays a key role in this process. A 32-bp deletion within the coding region of the CCR5 gene generates a truncated nonfunctional receptor. In HIV-1-infected individuals homozygous for this mutation, disease progression is inhibited. We analyzed the frequencies of the deletion in HIV-1 infected seropositive individuals. No significant differences in allelic frequencies of the CCR5 gene between the control and general HIV-1-infected cohorts and within the latter group between the infected individuals and patients with AIDS symptoms were revealed. PMID- 10867894 TI - [Internal medicine and peripheral neurology]. PMID- 10867895 TI - [Vascular peripheral neuropathy from neurological perspective]. PMID- 10867896 TI - [Vascular peripheral neuropathy from the internist's perspective]. PMID- 10867897 TI - [Peripheral neuropathy in diabetes mellitus from neurological perspective]. PMID- 10867898 TI - [Peripheral neuropathy in diabetes mellitus from internist's perspective]. PMID- 10867899 TI - [Inflammatory myopathies from neurologist's perspective]. PMID- 10867900 TI - [Inflammatory myopathies from internist's perspective]. PMID- 10867901 TI - [Stiff-man syndrome from neurological perspective]. PMID- 10867902 TI - [Stiff-man syndrome from the perspective of internal medicine and immunology]. PMID- 10867903 TI - [Interstitial lung diseases]. PMID- 10867904 TI - [28-year old woman requiring reanimation for heart arrest after hospitalization for treatment of deep vein thrombophlebitis]. PMID- 10867905 TI - [Splenomegaly in a 55-year old patient]. PMID- 10867906 TI - [Hypoxemia-induced symptomatology of sleep apnea]. PMID- 10867907 TI - [Metformin for prevention of diabetes?]. PMID- 10867908 TI - [Bowen's disease; association with tumors of internal organs?]. PMID- 10867909 TI - [The role of antibiotic therapy in infectious enteritis]. PMID- 10867910 TI - [Simple repeating sequences and gene expression]. PMID- 10867911 TI - [Detection of interspecies (interlineage) polymorphism in the structure of internal transcribed spacers of Zea mays corn ribosomal DNA]. PMID- 10867912 TI - [Analysis of nucleotide sequences in a region of S100b protein gene in AKR/J, DBA/dJ and BALB/cLac mice]. PMID- 10867913 TI - [Analysis of high molecular weight nuclear DNA, detected in mammalian cells by pulse-electrophoresis]. PMID- 10867914 TI - [Development of a colorimetric test system for detecting point mutations by ligating a tandem of short oligonucleotides on a solid-phase carrier]. PMID- 10867915 TI - [Allelic polymorphism at the TAQIB locus of the dopamine D2 receptor gene in groups of patients with endogenous psychoses]. PMID- 10867916 TI - [Determination and analysis of the nucleotide sequence of a segment of a Mycoplasma gallisepticum strain A5969 chromosome, containing operons S10 and rrn23-5]. PMID- 10867917 TI - [Deletion of the Alu-Vpa/MycL1 (1P34.3) locus--a sign of unfavorable prognosis in human colon cancer]. PMID- 10867918 TI - [Prediction of the effectiveness of gene expression by their nucleotide composition]. PMID- 10867919 TI - [Effect of inducers on the interaction of positive and negative cis-active elements of the CYP2B2 gene with nuclear proteins in rat liver]. PMID- 10867920 TI - [Interaction of a complex of human DNA and topoisomerase I with oligo-1,3 thiazolecarboxamides and their conjugates with oligonucleotides]. PMID- 10867921 TI - [Class 1 translation termination factors are structurally and functionally similar to suppressor of tRNA and are related to various structuro-functional families (prokaryotes and mitochondria--eukaryotes and archaebacteria)]. PMID- 10867923 TI - [Interaction of DNA with actinocin amides, containing radicals with cationoid centers in the amide groups]. PMID- 10867922 TI - [Multiplicity of site-specific DNA-methyltransferases of the BstF5I restriction modification system from Bacillus stearothermophilus F5]. PMID- 10867924 TI - [Synthetic construction, functionally imitating ribonuclease A]. PMID- 10867925 TI - [Molecular design based on polyribonucleotides, fixed in the structure of liquid crystalline dispersion and "cross-linked" via polymeric chelate bridges]. PMID- 10867926 TI - [p73 gene: deletion and expression in non-small cell lung cancer cells]. PMID- 10867927 TI - [Identification of CpG islands, hypermethylated in cancerous and transformed cells, by methyl-sensitive polymerase reaction with statistical primers]. PMID- 10867928 TI - [Design of virus-like particles, exposing HIV-1 epitopes]. PMID- 10867929 TI - [Simultaneous determination of mutations 1691 G-A of the blood coagulation system factor V gene and 20210 G-A gene, coding for prothrombin, by means of complex PCR from whole blood using one restriction enzyme (HindIII)]. PMID- 10867930 TI - [Molecular biology in Russia during the Soviet period (a short history)]. PMID- 10867931 TI - [The effect of ecoestrogens on the development of the mammalian reproductive system]. AB - Recently, it has been shown that the environment contains a large variety of chemical substances possessing estrogenic activity (ecoestrogens/environmental estrogens). In this connection, the problem of estrogenic effects exerted by the environment on animal ontogenesis is very important. Here, we review that available data concerning the effects of ecoestrogens on the development of the reproductive system in mammals. We discuss issues of reproductive toxicology related to the effects of these substances during prenatal and postnatal development, with special attention to the effect of ecoestrogens and modern reproductive technologies on preimplantation embryonic development. PMID- 10867932 TI - [The regulatory mechanisms of early embryogenesis in the mouse]. AB - The mechanisms involved in the regulation of preimplantation mammalian development have been considered using the example of mouse embryos. The role of four factors affecting the program of early embryogenesis is discussed: nucleocytoplasmic interactions, "maternal" control of development, cell-to-cell interactions, and genomic imprinting. The current concepts of the spatial and temporal regulation of developmental processes have been reviewed, as well as the perspectives of some trends in the experimental embryology of mammals. PMID- 10867933 TI - [The growth and form development of the limb buds in vertebrate animals]. AB - The development of the fin and limb buds involves a balance of centrifugal (active) and centripetal (passive) mechanical forces, the first of which acts to move the walls of these structures away from each other and the second holds them together. When the volume of the mesodermal core increases, the generated force meets with the resistance of the basal membrane, and as a result, the limb bud has a tendency to acquire cylindrical shape. Collagen fibers, individual mesenchymal cells, and their groups hold together the dorsal and the ventral wall of the limb bud, prevent the movement of these walls away from each other, and in this way direct bud growth along the proximodistal and the anteroposterior axes. The balance of the forces, which stretch the ectodermal layer, and those, which constrain it, have also been observed in the development of other body parts. PMID- 10867934 TI - [The absence of uniparental X-chromosome inheritance in spontaneous abortuses with a 46,XX karyotype]. AB - The problem of the presence of imprinted regions on the X-chromosome and the possible influence of the imprinted expression of X-linked genes on the embryonic development in man remains largely unsolved. A comparison of the uniparental inheritance of chromosomes or of their regions having different phenotypic manifestations provides an instrument with which to study the phenomenon of genomic imprinting at the chromosomal level. Assuming that the imprinted inactivation of X-chromosomes is functionally significant for embryonic development, we have studied several polymorphic micro- and minisatellite loci of X-chromosomes in 52 fetuses with karyotype 46,XX, which were spontaneously aborted during the first trimester of pregnancy. The purpose was to determine the contribution of uniparental disomy for the X-chromosome in any disturbances of the embryonic development. We found that inheritance of X-chromosomes was biparental in the studied embryos, suggesting the absence of any significant contribution of the parental origin of the X-chromosome to embryonic mortality occurring between 4 and 12 weeks of development. PMID- 10867935 TI - [Disordered differentiation of mouse embryonic lungs in organ cultures under the action of nitrosomethylurea]. AB - We have studied the effect of nitrosomethylurea (NMU) on differentiation of early rudiments of mouse embryonic lungs (12th day of embryogenesis) explanted into an organ culture. We have demonstrated that nontoxic doses of NMU are capable of accelerating normal lung differentiation both at the early (increase in the number of epithelial buds) and at the late (increase in the number of explants with regions of well-developed alveoli) stages of cultivation. However, NMU induces disturbances of differentiation, which appear as polycystic structures and hyperplastic nodules generally absent in the control. PMID- 10867936 TI - [Ontogenetic divergence in the phytoparasitic nematodes Macroposthonia nainitalensis and M. tenuiannulata (Criconematidae, Tylenchia)]. AB - Promorphogenesis, egg cleavage, and gastrulation proceed in Macroposthonia nainitalensis similarly to other phytonematodes of the subclass Tylenchia. Before the onset of gastrulation, the greatest similarity was observed in a descendant of the micromere 2b, which separates macromeres of the third division. Deviations from the general line of development started in this species during embryonal organogenesis with the formation of an external epithelium with a special rough surface layer, which was absent in the embryos of other phytonematodes. In addition, during this period of acceleration, the digestive system developed, while peristaltic changes in body shape appeared at the tadpole stage. Later, somatic muscle development was enhanced in the first instar larvae of M. nainitalensis, while in the second instar larvae of Macroposthonia spp. somatic muscle cells were located at an acute angle to the longitudinal body axis. In addition, denticles--microchaetae appeared in the second to fifth instar larvae of these nematodes. All these features, which have not been observed in other phytonematodes of the subclass Tylenchia, help the peristaltic crawling of Macroposthonia spp. In contrast to other Tylenchia, the musculocutaneous sac did not develop in Macroposthonia spp. However, there are similarities in the development of the digestive and genital systems. PMID- 10867937 TI - [The effect of prenatal stress on steroidogenesis in the gonads of Arctic foxes (Alopex lagopus)]. AB - Handling is a source of stress for farm-bred blue foxes. The influence of handling during the late gestational period was investigated in 10-day old male and female blue foxes. Testosterone and estradiol were measured by RIA in the plasma, gonadal homogenates and in vitro incubates from blue foxes of both sexes. The gonads were incubated in vitro without or with human chorionic gonadotropin. In cubs of both sexes, the gonad weights and ovarian estradiol production were decreased by stress. The testicular testosterone and ovarian estradiol contents were increased in prenatally stressed cubs as compared to the controls. The testicular content and baseline in vitro production of testosterone were not affected by prenatal stress, but the testicular response to human chorionic gonadotropin was higher in the stressed group. This study suggests that prenatal stress induced by handling pregnant vixens may influence gonadal steroidogenesis and this effect was more pronounced in female cubs. PMID- 10867938 TI - [A common link in the mechanism of the self-maintenance of malignant growth: the syndrome of the nonhealing wound]. AB - A general principle of the maintenance of malignant growth in all types of tumors has been formulated. According to this principle, stochastic but continuous death of some tumor cells due to the inherent genetic instability of their genome (fragility of chromosomes) is the main event stimulating tumor growth. The dead cells trigger a complex multicomponent process of wound healing expressed as further proliferation of living tumor cells, angiogenesis, stimulation of cell migration, and other events. Stimulation of the proliferation of living cells leads to further death of cells and, as a result, to further stimulation of the system of wound healing, etc. Thus, the tumor sacrifices a small amount of dying cells to stimulate proliferation of all its other cells. It is proposed that the nature of the genetic instability of malignant cells is related to the appearance of an uninemic structure in some regions of chromosomes, in whole chromosomes, or in whole genomes. The author bases his statements on the binemic structure of chromosomes, which has already been experimentally and theoretically substantiated. Uninemic regions have an exceedingly high frequency of spontaneous chromosome aberrations due to blockage of the mechanism of underlying repair of the DNA double break in the absence of a second DNA copy. Possible approaches to a search for more efficient methods of therapy are discussed. PMID- 10867939 TI - [Genomics. Proteomics. Bioinformatics. What else?]. AB - The biology of the past century was, undoubtedly, reductionist. Its general trend was expressed in the appearance and burst of scientific disciplines, such as biochemistry, biophysics, molecular biology, molecular genetics, virology, cell biology, and bioorganic chemistry. All these disciplines are united by an aspiration to explain the phenomena of life through the description of the properties of molecules, first of all, biopolymers, which constitute the living organisms. In current science, all these directions are often united under the general heading "physicochemical biology", while classical biological disciplines, such as zoology, botany, embryology, etc., are referred to as "general biology". PMID- 10867940 TI - [Fermentation waste--a biomodifier of concrete]. AB - The composition and properties of molasses-containing yeast fermentation waste used as a plasticizer and structural biological additive to concrete mixtures are reviewed. The basic principles of the effects of organic and inorganic components of yeast fermentation waste on the properties of the bonding system are analyzed. PMID- 10867941 TI - [Quantitative analysis of calibration dependence of biosensors]. AB - Three-parameter Hill's equation, which is used in enzyme kinetics, was shown to applicable to calibration curves of both potentiometric (glucose, pesticides, urea, etc.) and amperometric (surfactants, biphenyl, etc.) biosensors. Possible causes of errors of analyte concentration measurements are discussed. PMID- 10867942 TI - [Comparative kinetic characteristics of catalase of Penicillium species molds]. AB - Extracellular catalases produced by fungi of the genus Penicillium: P. piceum, P. varians and P. kapuscinskii were purified by consecutive filtration of culture liquids. The maximum reaction rate of H2O2 decomposition, the Michaelis constants and specific catalytic activities of isolated catalases were determined. The operational stability was characterized by effective rate of catalase inactivation during enzymatic reaction (kin at 30 degrees C). The thermal stability was determined by the rate of enzyme thermal inactivation at 45 degrees C (k*[symbol: see text]H, s-1). Catalase from P. piceum displayed the maximum activity, which was higher than the activity of catalase from bovine liver. The operational stability of catalase from P. piceum was twofold to threefold higher than the stability of catalase from bovine liver. The physicochemical characteristics of catalases of fungi are better than the characteristics of catalase from bovine liver and intracellular catalase of yeast C. boidinii. PMID- 10867943 TI - [Oxidation of organic compounds by Rhodococcus erythropolis 3/89 propanomonooxygenase]. AB - The ability of propane monooxygenase of Rhodococcus erythropolis 3/89 to catalyze oxidation of higher liquid alkenes and aromatic hydrocarbons was studied. Optimal conditions of tetradecene epoxidation and benzene hydroxylation were found. Under these conditions, oxidation was shown to be accompanied by a 100% conversion of benzene to phenol. PMID- 10867944 TI - [Certain biochemical and physicochemical properties of the inducible form of extracellular laccase from basidiomycetes Coriolus hirsutus]. AB - An inducible form of extracellular laccase (EC 1.14.18.1) was isolated from the basidiomycete Coriolus hirsutus. The induction was performed with 0.11 microM syringaldazine, a substrate of laccase. The inducible form of the enzyme consisted of two isoforms, laccase I1 and laccase I2, whose molecular weights were 69 +/- 2 and 67 +/- 2 kDa, respectively. The isoelectric points of these isoenzymes were found to be 3.5 and 4.2, respectively. The optimum pH range for both laccases was 4.4-4.6, and the optimum temperature was 50 degrees C. The thermal stability of these isoenzymes was examined, and KM values for the substrates syringaldazine and pyrocatechol were determined. Our biochemical and physicochemical studies demonstrated that inducible laccase isoforms differed from constitutive forms in molecular weight, IP, KM, and thermal stability. However, their optimum pH ranges and temperatures were identical. PMID- 10867945 TI - [Enzyme regeneration during hydrolysis of steam-pretreated willow and requirement for cellulase complex composition]. AB - In order to reduce the total enzyme consumption in high-solids static hydrolysis of nonwashed steam-exploded willow Salix caprea by mixed cellulase of Trichoderma reesei + Aspergillus foetidus, two different approaches were proposed. In the first case, the enzyme activity adsorbed on residual solids after extended hydrolysis was used for hydrolysis of the newly added substrate. The initial mixing of fresh and hydrolyzed substrates was sufficient for the adsorbed enzyme redistribution and conversion of the new substrate portion, and permanent mechanical stirring was not required. Feeding of two additional portions of the exploded hardwood adjusted to pH 4 with dry caustic into the reactor with simultaneous replacement of accumulated sugars with fresh buffer (pH 4.5) resulted, on average, in a 90% conversion of cellulose at the final enzyme loading 8 IFPU per g ODM substrate, an average sugar concentration of 12%, and a glucose/xylose ratio of 5:1. In the second approach, weakly adsorbed cellulase fractions were used for static high-solids hydrolysis followed by their ultrafiltration recovery from the resultant sugar syrup. In contrast to the initial cellulase mixture whose residual activity in a syrup did not exceed 5-10% at the end of hydrolysis (48 h), up to 60% of weakly adsorbed enzyme fraction could be separated from sugar syrups by ultrafiltration and then reused. Weakly adsorbed enzymes displayed a hydrolysis efficiency of not less than 80% per IFPU enzyme consumed in extended hydrolysis of pretreated willow as compared to the original enzyme mixture. An electrophoretic study of the weakly adsorbed enzyme fraction identified T. reesei cellobiohydrolase II as the predominant component, whereas clear domination of T. reesei cellobiohydrolase I was found by electrophoresis of proteins tightly bound to hydrolysis residual solids. PMID- 10867946 TI - [Change in composition of structural components of leaf cellulose sulfate during fermentation bleaching by xylanases]. AB - The possibility of the use of xylanase preparations for hydrolysing hemicelluloses in a non-bleached kraft pulp in order to facilitate its bleaching was studied. The effects of enzymatic preparations of the fungal and bacterial origins were examined, and the optimal conditions for xylanase activity were determined. UV spectroscopy demonstrated that the treatment of kraft pulp with the enzymatic preparations containing xylanase facilitated the subsequent removal of lignin and increased the brightness by 5%. The effect of enzymatic treatment was retained in the case of peroxide bleaching. The enzymatic preparations studied are promising for the development of chlorine-free pulp bleaching technologies. PMID- 10867947 TI - [Change in composition of hydrolyzed lignin during composting]. AB - Microbial assemblages were composed for composting hydrolysed lignin. Data on bioconversion of aromatic compounds with various types of substitution in the ring were used for this purpose. Composting of hydrolysed lignin reduced the contents of lignin, low-hydrolyzable polysaccharides, resins, and low-molecular weight phenols and resulted in accumulation of humic acids. The resulting compost showed no phytotoxicity. PMID- 10867948 TI - [Comparative characteristics of proteolytic enzyme preparations by degree of hydrolysis of a microbial protein]. AB - Proteolytic enzymatic preparations obtained from fungi and bacteria were compared by their ability to hydrolyze yeast protein. Fungal preparations were more effective. There was a more than twofold increase in the level of amine nitrogen in cell biomass hydrolysates in of cell biomass comparison to that induced by bacterial preparations. The amino acid composition of these hydrolysates was studied. Amyloprotooryzin, a preparation from Aspergillus oryzae 387, displayed the highest potency in profound protein hydrolysis: the concentration of free amino acids released was 34.7% in comparison to 20.6% induced by amyloryzin and 10.5% by protosubtilin. PMID- 10867949 TI - [Hydrolase from Xanthomonas rubrilineans for synthesis of cefalexin]. AB - The use of peptide hydrolase (EC 3.4.13.1) from Xanthomonas rubrilineans for synthesis of the antibiotic cephalexin from 7-aminodesacetoxycephalosporanic acid was studied. The optimum conditions for production of cephalexin were determined, and the yield exceeded 80%. A method for monitoring the synthesis of this antibiotic synthesis by means of a conventional amino acid analyzer is proposed. PMID- 10867950 TI - [Transposon mutagenesis in preparing mutants of a strain--a producer of a new antibiotic batumin]. AB - Various plasmids carrying transposon Tn5 were used to generate insertion mutants synthesizing batumin, a unique antibiotic with a selective antistaphylococcal effect. One of the plasmids used provided a sufficient yield of the clones in question. An analysis of over 7000 clones allowed us to select the mutant clones with increased and decreased levels of batumin synthesis and the mutants that lost the ability to synthesize this antibiotic. PMID- 10867951 TI - [Development of a new method for preparing biologically active compounds based on the typed strain Streptomyces werraensis]. AB - The regulatory function of a DNA fragment responsible for actinomycin resistance in the typed strain Streptomyces werraensis ATCC 1365, which produces a macrotetrolide antibiotic, was studied. Metabolic changes made this strain capable of producing an antibiotic complex, which comprises four biologically active compounds absent from the parent culture. PMID- 10867952 TI - [Diketopiperazine alkaloids from the fungus Penicillium piscarium Westling]. AB - Fungi of the species Penicillium piscarium produced diketopiperazine alkaloids (isorugulosuvine, puberuline, verrucosine, prolyltryptophanyldiketopiperazine, 12,13-dehydroprolyltryptophanyldiketopiperazine, fellutanine A, phenylalanylphenylalanyldiketopiperazine, as well as roquefortine and 3,12 dihydroroquefortine whose precursors are tryptophan, phenylalanine, leucine, proline, and histidine. PMID- 10867953 TI - [Effect of the mutation process on formation of alkaloids in Penicillium roquefortii BKM F-141 and P. fellutanum BKM F-1073]. AB - Mutant strains of Penicillium roquefortii VKM F-141 and P. Fellutanum VKM F-1073 were obtained by mutagenesis induced by ultraviolet irradiation, N-methyl-N nitronitrosoguanidine and bromouracil. By the rates of alkaloid production, the mutant strains can be divided into three groups: 1) unable to synthesize alkaloids; 2) with a high rate of biosynthesis; 3) with changed alkaloid composition. Compounds not characteristic of wild-type cultures were found in alkaloid fractions of some mutant strains. PMID- 10867954 TI - [Preparation of conjugated antigens based on zearalenone carboxymethyloxime and their use in immunoenzyme analysis]. AB - Zearalenone-6'-carboxymethyloxime was synthesized, and its conjugates with albumins and gelatin were prepared. Polyclonal rabbit antibodies against the conjugate with bovine serum albumin were shown to be highly specific to zearalenone and to have a lower cross-reactivity toward its structural analogues (alpha-zearalenol--28%, beta-zearalenol--6%, zearalanone--12%, and alpha zearalanol--5%). The sensitivity of enzyme immunoassay using gelatin-based immobilized conjugates for determination of zearalenone in solutions was 1 ng/ml, and this allowed us to determine this substance in feed at a threshold concentration of 200 micrograms/kg. PMID- 10867955 TI - [Phenol compounds from brown algae and their antioxidant activity]. AB - The composition and content of phenolic substances were studied in 14 species of marine brown algae of the Canary Islands littoral (Spain). The highest content of phenolic substances was found in Cystoseira compressia and Sargassum furcatum. A high antioxidant activity was found in florotannin isolated from Cystoseira sp. PMID- 10867956 TI - [Intensification of the initial stage of amino acid metabolism in germinating cereal seeds by exogenous glutamine and proline]. AB - The initial stage of amino acid metabolism was intensified in germinating wheat seeds with exogenous glutamine and proline. Exogenous glutamine and proline accumulated over 2 h at 4 degrees C in swelling seeds were spent at different rates over the following 2 h at 20 degrees C, thus compensating for insufficiency of these amino acids during the initial stage of development. Creation of an additional store of glutamine and proline during the initial stage of amino acid metabolism had positive effects on the seed germination and vital activity of the plants. PMID- 10867957 TI - [Oxyanthraquinones and flavonoids from garland chrysanthenum]. AB - The stems of the vegetable plant garland chrysanthemum (Chrysanthemum coronarium L.) were shown to contain emodin (in its aglycon and glycoside forms) and chrysophanol. Chrysophanol and chrysazin were isolated from the roots of the plant. Because the pigments identified are derivatives of 1,8 dihydroxyanthroquinone, garland chrysanthemum may be a medicinal plant and have utility as a component of laxative species. The leaves of C. coronarium were shown to be rich in quercetin and its glycosides, rutin and isoquercetin. Taken together, this observation and the known high content of ascorbic acid and carotenoids in the plant suggest that C. coronarium may be useful in preventing cardiac and vascular diseases. PMID- 10867958 TI - [A cell for bioelectrochemical studies with parallel spectrophotometric monitoring of the studied substance]. AB - An electrochemical cell for simultaneous potentiometric and spectrophotometric measurements of solutions of various substances is described. A schematic diagram of the cell, its advantages, method of use, and possible areas of application are discussed. The cell is suggested to be used for electrochemical potentiometric redox titration of leghemoglobin and electrochemical reduction of organic reductants commonly used in biochemical research. PMID- 10867959 TI - [A new method for immobilizing microbial cells by crosslinking]. AB - A method for immobilization of microbial cells was developed designed. The method uses based on generation of reactive aldehyde groups on the cell wall surface under conditions of periodate oxidation. The linking of aldehyde groups by various bifunctional aromatic diamines and then by glutaraldehyde produced immobilized cells, which are promising for the use in biocatalysis with high molecular-weight substrates of high molecular weight. PMID- 10867960 TI - Neuropharmacological treatments for alcoholism: scientific basis and clinical findings. AB - Preclinical studies have exploded our knowledge about the behavioral and biological underpinnings of alcoholism. These studies suggest that certain neurotransmitters, particularly those interacting with the opioid, N-methyl-D aspartate, and monoamine systems, may play a critical role in the expression of alcohol-drinking and other behaviors associated with its abuse liability. Built upon this foundation, important advances have been made in the development of therapeutic medications for the treatment of alcoholism. Of the medications reviewed, acamprosate's potential appears to be the most widely established. In the USA, naltrexone was approved by the Food and Drug Administration in 1995 for the treatment of alcoholism; however, the results of some studies have been less encouraging. Naltrexone's reliance on high compliance rates for efficacy may, eventually, limit its potential in clinical settings offering generic treatment for alcoholism. The relative paucity of dose-response studies on naltrexone's effects in treating alcoholics is an important gap in the literature. Recent data from a large clinical trial suggests that ondansetron, a serotonin3 antagonist, offers new hope for the treatment of early onset alcoholics; a type of alcoholism most difficult to manage with psychosocial measures alone. Different subtypes of alcoholic may, therefore, have varying treatment responses to serotonergic agents. Matching subtypes of alcoholic to effective treatment medications based upon their different biologies remains an important therapeutic goal. Combinations of effective pharmacological agents need exploration as they may prove to be synergistic, and could shepherd in a new era of treatments aimed at multiple neurotransmitter targets associated with the alcoholism disease. The coming decade promises more powerful tools for characterizing drug effects on alcohol drinking, thereby closing the gap between animal models of addiction and the human condition. PMID- 10867961 TI - Effects of the NMDA receptor channel blockers memantine and MRZ 2/579 on morphine withdrawal-facilitated aggression in mice. AB - RATIONALE: Opioid withdrawal is known to facilitate aggressive behavior in laboratory rodents. Aggression develops as the somatic signs disappear and thus may reflect protracted withdrawal-related behavioral alterations. Antagonists acting at the NMDA receptor are known to attenuate the expression of morphine withdrawal syndrome in laboratory animals. OBJECTIVE: The present study aimed to evaluate the effects of low-affinity NMDA receptor channel blockers (memantine and MRZ 2/579) on aggression facilitated by morphine withdrawal in mice. METHODS: Significant increases in aggressive behavior were observed 48 h after repeated morphine administration (8 days, b.i.d., 10-80 mg/kg, s.c.) was discontinued. Separate groups of mice were treated intraperitoneally with vehicles or different doses of memantine (1, 3, 10 or 30 mg/kg) or MRZ 2/579 (1, 3 or 10 mg/kg) 48 h after the last morphine injection. RESULTS: Both compounds dose-dependently reduced the expression of aggressive behavior while having no significant effect upon the intensity of non-aggressive social contacts. Memantine significantly diminished the occurrence of all recorded components of aggressive behavior (attacks/bites, threats, tail rattling) while MRZ 2/579 affected mainly the appetitive events of aggressive bursts (threats, tail rattling). For both compounds, anti-aggressive effects occurred at dose levels that did not produce motor impairment in the Rotarod test. CONCLUSIONS: Taken together with the evidence on the lack of selective anti-aggressive effects of these drugs in morphine-naive mice, attenuation of the aggression observed in the present study may be due to specific interaction with morphine withdrawal-triggered processes. PMID- 10867962 TI - Comparison of the neurophysiological effects of allopregnanolone and ethanol in rats. AB - RATIONALE: The central nervous system actions of allopregnanolone (3 alpha hydroxy-5 alpha-pregnan-20-one) and ethanol are at least partially mediated by modulation of gamma-aminobutyric acid (GABA)-A receptors. Although ethanol and allopregnanolone have similar behavioral effects, their macro electrophysiological profiles have not been directly compared. OBJECTIVE: The purpose of this study was to compare the effects of allopregnanolone and ethanol on the electroencephalogram (EEG) and event-related potentials (ERPs). METHODS: Male Wistar rats were implanted with cortical and amygdalar electrodes. The rats were then administered allopregnanolone (0.0-10 mg/kg), ethanol (0.0-1.0 g/kg), or a combination of the two before recording. RESULTS: Allopregnanolone and ethanol had similar effects on ERPs. When administered alone, both decreased cortical P1-N1 ERP amplitude by 25-50% and N1 amplitude in the amygdala by 75 80%. Combined administration of ethanol (0.50 g/kg) and allopregnanolone (5.0 mg/kg), doses which were ineffective alone, decreased N1 amplitude in the amygdala by 60%. Allopregnanolone and ethanol had dissimilar EEG effects. Allopregnanolone increased high frequency power in the cortex and amygdala by 25 30%. Ethanol decreased cortical and amygdalar power in the same high frequency bands by 25-45%. Allopregnanolone, but not ethanol, also shifted cortical frequency in the 32- to 50-Hz band. Combined administration of allopregnanolone and ethanol had no effect on EEG power but enhanced allopregnanolone's effect on cortical frequency. CONCLUSIONS: These data suggest that allopregnanolone's macro electrophysiological profile resembles barbiturates and benzodiazepines more than ethanol. Further, the interactions of allopregnanolone and ethanol appear complex, with multiple effects observed (enhancement or reversal) depending on the neurophysiological variable assessed. PMID- 10867963 TI - Sleep changes during long-term treatment of depression with fluvoxamine--a home based study. AB - RATIONALE: The effects of antidepressants on sleep in depression have been extensively investigated, although to date there have been relatively few studies of newer drug classes such as specific serotonin reuptake inhibitors (SSRIs). All reported studies on SSRIs have been conducted in patients admitted to sleep laboratories and very few longitudinal studies have continued to measure sleep beyond 5 weeks of treatment. The growing trend towards outpatient and community care has highlighted the need for studies of sleep in depression in a more naturalistic setting, and during longer periods of treatment in line with recommended clinical practice. OBJECTIVES: To establish if the changes in sleep architecture and continuity described during early treatment with SSRIs persist after 3 months, to relate these changes to clinical state, and to establish whether home recordings would yield similar results to previous laboratory studies. METHODS: We have recorded objective sleep parameters in 12 depressed patients before and during 12-week treatment with an SSRI, fluvoxamine. All the sleep recordings were performed in the patients' own homes, using the Oxford Medilog system. RESULTS: At 12 weeks, 7/12 patients had responded (HAM-D decreased by > 50%). REM latency showed the expected increase early in treatment; this change was less obvious at weeks 3 and 12. Amount of REM sleep was decreased at day 2 and week 3, but returned to baseline by week 12. Slow wave sleep was slightly increased at day 2 and decreased at week 12. Of the sleep continuity measures, the only significant change was in sleep onset latency, which was increased at week 3; the other measures showed non-significant worsening at night 2 and week 3, but most were better than baseline by 12 weeks. Subjective sleep (the three sleep items on the HAM-D) showed a progressive improvement over time, especially in the responders. CONCLUSIONS: The effects of the SSRI fluvoxamine on objective sleep measures are in the direction predicted by its pharmacological actions and some persist for at least 12 weeks. In addition subjective appraisal of sleep is strongly affected by mood state. All patients found the home recording procedure acceptable and only minimally disruptive. PMID- 10867964 TI - Prefrontal dopamine is directly involved in the anxiogenic interoceptive cue of pentylenetetrazol but not in the interoceptive cue of chlordiazepoxide in the rat. AB - RATIONALE: The prefrontal cortical (PFC) dopamine (DA) system has been implicated in anxiety-related behavioral changes, but direct, unequivocal support for this idea is sparse. OBJECTIVES: The present aim was to study the functional significance of prefrontal DA using the pentylenetetrazol (PTZ) discrimination model of anxiety. A comparison was made with its role in the cue of the anxiolytic drug chlordiazepoxide (CDP). METHODS: Two groups of rats were trained to discriminate either PTZ (20 mg/kg, s.c.) or CDP (10 mg/kg, i.p.) from saline using an operant drug discrimination procedure. After prolonged training, half of each group was used to assess biochemical changes induced by both drugs in different sub areas of the PFC. For the remaining rats, discrimination training continued and generalization tests with PTZ and CDP were performed. Rats were then provided with bilateral guide cannulae aimed at the ventromedial (vm) PFC, and the effects of local infusions of DAergic drugs on discriminative performance were evaluated. RESULTS: CDP did not affect PFC DA activity, but PTZ increased the DOPAC/DA ratio in the vmPFC selectively. Generalization tests showed that the cues of PTZ and CDP were dose dependent. In PTZ-trained rats, infusions of the DA receptor antagonist cis-flupenthixol into the vmPFC blocked the PTZ cue dose dependently, whereas the agonist apomorphine partially generalized to this cue. In CDP-trained rats, neither drug antagonized or generalized to the CDP cue, showing that PFC DA is not critically involved in the CDP cue and that local pharmacological manipulations of PFC DA do not affect discriminative abilities per se. CONCLUSIONS: The DAergic innervation of the PFC is directly involved in the behavioral effects of PTZ suggesting a role for it in anxiety. PMID- 10867965 TI - In vivo estimates of efficacy at 5-HT1A receptors: effects of EEDQ on the ability of agonists to produce lower-lip retraction in rats. AB - RATIONALE: Maximal responses are often used as a measure of intrinsic activity or efficacy, but cannot be directly equated to efficacy. Using irreversible antagonists, estimates of efficacy can be obtained that may be less dependent on specific conditions. OBJECTIVES: To characterize the intrinsic activity of serotonin (5-HT)1A agonists by examining the effects of an irreversible antagonist on their ability to produce 5-HT1A receptor-mediated responses. METHODS: The effects of N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) on the ability of 5-HT1A agonists to produce lower-lip retraction (LLR) in rats were studied. RESULTS: In the absence of EEDQ, each 5-HT1A agonist produced full effects, the rank order of potency being: S 14506 > 8-OH-DPAT > buspirone > ipsapirone. EEDQ decreased the number of 5-HT1A binding sites and shifted the dose-response curves (DRCs) of each agonist either to the right or, at higher EEDQ doses, to the right and downward. The manner in which these shifts occurred, however, differed among the compounds. For each agonist, all DRCs obtained after different doses of EEDQ were fitted to models proposed by Furchgott and Black and Leff, and the results indicated the following rank order of efficacy: ipsapirone < buspirone approximately 8-OH-DPAT < S 14506. 5-HT1A agonist-induced LLR appears to be mediated by 5-HT1A receptors, because the 5-HT1A antagonist, WAY 100635, shifted the agonist DRCs to the right in a parallel and dose-related manner, with pA2 values ranging from 7.8 to 8.1. Moreover, pretreatment with WAY 100635 protected against the antagonist activity of EEDQ. CONCLUSIONS: The results suggest that the effects of EEDQ on the ability of 5-HT1A agonists to produce LLR in rats may be useful to obtain estimates of their apparent efficacy at 5-HT1A receptors. PMID- 10867966 TI - Effects of methyllycaconitine (MLA), an alpha 7 nicotinic receptor antagonist, on nicotine- and cocaine-induced potentiation of brain stimulation reward. AB - It has been shown that nicotine facilitates intracranial self-stimulation (ICSS) reward and that nicotinic acetylcholine receptors (nAChRs) in the ventral tegmental area (VTA) are of primary importance for its reinforcing and dependence producing actions. Recently, we have shown that alpha 7 nicotinic receptors in the VTA contribute to both the acute effects of nicotine on the mesolimbic dopamine system, as well as to nicotine withdrawal reactions. However, it is not yet known whether the same receptor conformation is directly involved in the reinforcing actions of nicotine. Here, using the curve-shift method we studied the effects of methyllycaconitine (MLA), a selective alpha 7 receptor antagonist, microinjected (graded doses: 1, 3, 9 micrograms/microliter per side) into the VTA on the rewarding efficacy of lateral hypothalamic self-stimulation and on the systemic nicotine-induced potentiation of brain stimulation reward. MLA did not affect baseline self-stimulation. Nicotine produced a significant reduction in ICSS threshold, without altering maximal rates of responding, while MLA attenuated the effect of nicotine at the two lower doses. Given the reported interaction between nicotine and cocaine at both the neuronal and the behavioral level, we also examined whether alpha 7 receptor antagonism within the VTA can affect the reinforcing action of cocaine, as measured with ICSS. Interestingly, MLA attenuated the reinforcing effect of cocaine in all doses tested, without altering the maximal rate of responding, i.e. the performance of the animals. These results suggest that alpha 7 nAChRs in the VTA are involved in mediating the reinforcing actions of drugs of abuse, such as nicotine and cocaine, and provide evidence that alpha 7 nAChR antagonists may be clinically useful in attenuating the rewarding effects of addictive drugs. PMID- 10867967 TI - Socio-psychological stress-induced antinociception in diabetic mice. AB - RATIONALE: Although it is well established that different forms of stress produce a pronounced antinociception, the effect of diabetes on psychological stress induced antinociception is not yet clear. OBJECTIVES: The effect of diabetes on psychological stress-induced antinociceptive effect was assessed in mice. METHODS: Animals were rendered diabetic by an injection of streptozotocin (200 mg/kg, i.v.). Mice were exposed to psychological stress in the compartment of a communication box. The antinociceptive response was evaluated by the tail-flick test, using radiant heat as a stimulus, which was performed before stress (pre stress latency) and 0, 30 and 60 min after stress. RESULTS: Exposure to socio psychological stress for 5, 10 and 15 min produced duration-dependent antinociception in diabetic mice. However, in non-diabetic mice, no appreciable antinociception was found even in the case of socio-psychological stress for 15 min. Pretreatment with diazepam (0.3 mg/kg, i.p.) significantly attenuated socio psychological stress-induced antinociception in diabetic mice (vehicle: 62.9 +/- 5.5%, n = 10; diazepam: 22 +/- 1%, n = 10). Furthermore, pretreatment with flumazenil (1 mg/kg, i.v.), a benzodiazepine receptor antagonist, also significantly reduced socio-psychological stress-induced antinociception in diabetic mice (vehicle: 77.9 +/- 5.0%, n = 10; flumazenil: 5.8 +/- 1.2%, n = 10). In contrast, pretreatment with methyl beta-carboline-3-carboxylate (beta-CCM, 2 mg/kg, i.v.), a benzodiazepine receptor inverse agonist, significantly enhanced socio-psychological stress-induced antinociception in non-diabetic mice (vehicle: 4.9 +/- 0.6%, n = 10; beta-CCM: 61.5 +/- 5.9%, n = 10), but not in diabetic mice (vehicle: 50.7 +/- 4.5%, n = 10; beta-CCM: 64.4 +/- 7.2%, n = 10). CONCLUSIONS: These results indicate that emotional stress can readily induce antinociception in diabetic mice. Furthermore, this enhanced emotional stress-induced antinociception might be attributable to an increase in the production and/or release of endogenous ligands for benzodiazepine receptors, such as diazepam binding inhibitor, which act as inverse benzodiazepine receptor agonists. PMID- 10867968 TI - Acquisition of oral phencyclidine self-administration in rhesus monkeys: effect of sex. AB - RATIONALE: There are increasing reports of sex differences in the etiology of drug abuse in humans. A nonhuman primate model is useful for examining sex as a variable in drug abuse. OBJECTIVES: To determine whether there are sex differences in the acquisition of oral phencyclidine (PCP) self-administration and to compare the effect of altered feeding conditions on drug self administration in male and female monkeys. METHODS: Acquisition of orally delivered PCP was studied using 7 female and 11 male adult rhesus monkeys. Initially, the monkeys were not food restricted, and they were given access to water under concurrent fixed-ratio (FR) 1 schedules during daily 3-h sessions. Each lip-contact response on a drinking spout resulted in a 0.3 ml liquid delivery. After baseline levels of water intake were obtained for 5 days, water was replaced with PCP (0.125 mg/ml) at both drinking spouts. Body weights were then reduced to 85% of free-feeding weights, and the monkeys were fed 30 min before the session began. The FR value was increased from 1 to 2, 4, and 8, at both drinking spouts. As a final step in the procedure, water and PCP were concurrently available at the two spouts under FR 8 schedules. Acquisition of PCP reinforced behavior was considered to have occurred if PCP intake was consistently greater than water intake. RESULTS: Lip-contact responses and liquid deliveries were not significantly different between the females and males throughout the acquisition period, but there was a significant increase in responding and decrease in liquid intake as FR increased, and a significant increase in PCP consumption due to food restriction that did not differ in males and females. On a milligram per kilogram basis, female monkeys consumed nearly twice as much PCP as the males; however, this effect was not significant. The females showed significantly higher PCP than water intake while the males consumed approximately equal amounts of PCP and water. Of the seven females, 100% met the acquisition criterion of significantly greater PCP than water intake, while only 36.4% of the males met the criterion. CONCLUSION: These results concur with previous rat studies and indicate that female monkeys are more likely than males to acquire drug-reinforced behavior. PMID- 10867969 TI - The P300 brain potential is reduced in smokers. AB - RATIONALE: Tobacco smoking is the most prevalent type of substance abuse, yet its biobehavioral etiology is little understood. Identification of differences between smokers and non-smokers on basic characteristics of neurocognitive functioning may help to elucidate the mechanisms of tobacco dependence. OBJECTIVES: This study assessed the relationship between smoking status and the P300 component of event-related potential (ERP) while controlling for potential confounders such as alcoholism, drug abuse, and psychopathology. METHODS: The ERP responses elicited by a visual oddball task were measured at the mid-parietal site in 905 current smokers, 463 ex-smokers, and 979 never smokers. RESULTS: P300 amplitude was significantly lower in current cigarette smokers compared to never smokers. Ex-smokers did not differ significantly from never-smokers. P300 reduction was also associated with alcoholism, drug dependence, and family density of alcoholism. However, after controlling for smoking, only family density of alcoholism remained a significant predictor of P300 amplitude. CONCLUSIONS: The results indicate a significant effect of smoking status on P300 amplitude which is additive to family history of alcoholism and suggest that either (1) long-term tobacco smoking may produce a reversible change in brain function, or (2) reduced P300 may be a marker of risk for nicotine dependence. PMID- 10867970 TI - Mecamylamine prevents tolerance but enhances whole brain [3H]epibatidine binding in response to repeated nicotine administration in rats. AB - RATIONALE: Chronic administration of nicotine in rats results in upregulation of neuronal nicotinic receptors. Upregulation has been proposed to reflect receptor desensitization, which may underlie functional tolerance to nicotine's effects. However, evidence indicates that tolerance and upregulation do not always parallel each other, suggesting that either upregulation does not always reflect desensitization, or mechanisms other than receptor desensitization account for tolerance to nicotine. OBJECTIVES: The present studies examined tolerance to nicotine-induced antinociception and changes in receptor binding after two regimens of intermittent nicotine injections in rats. The role of receptor activation in upregulation and tolerance was also examined by co-administering nicotine with the non-competitive antagonist, mecamylamine. METHODS: Sprague Dawley rats were administered a short (once-daily, s.c. for 6 days (0.35 mg/kg)) or long (twice-daily for 11 days (0.66 mg/kg)) series of injections and tolerance to nicotine-induced antinociception and [3H]epibatidine binding in whole brain were measured. RESULTS: The short series of injections resulted in tolerance to nicotine-induced antinociception, but failed to increase [3H]epibatidine binding. In contrast, the long series of injections resulted in both tolerance and increased receptor binding. Once-daily pairings of mecamylamine (1 mg/kg, s.c.) with nicotine (0.35 mg/kg) for 6 days blocked the development of tolerance, indicating receptor activation is necessary for tolerance to occur. Pairing mecamylamine with nicotine (0.66 mg/kg) twice daily for 11 days blocked tolerance but produced a greater increase in [3H]epibatidine binding than nicotine alone. CONCLUSIONS: A dissociation of tolerance from receptor upregulation was observed in the present study. The finding that receptor activation may be necessary for tolerance but not upregulation is discussed within the context of possible mechanisms controlling tolerance to nicotine. PMID- 10867971 TI - Risperidone counteracts lethality in an animal model of the serotonin syndrome. AB - RATIONALE: The serotonin (5-HT) syndrome is the most serious side effect of antidepressants, and pharmacologic treatment should be offered in severe cases. OBJECTIVE: In the present study, the effects of risperidone, ketanserin, and haloperidol on an animal model of the serotonin (5-HT) syndrome were evaluated. METHODS: Intraperitoneal administration of 100 mg/kg 5-hydroxy-L-tryptophan (5 HTP) (a precursor of 5-HT) and 2 mg/kg clorgyline (a monoamine oxidase type-A inhibiting antidepressant) induced the 5-HT syndrome in rats. The rectal temperature of the rats was measured, and the microdialysis method was used to measure noradrenaline (NA) levels in the anterior hypothalamus. RESULTS: In the group pre-treated with saline, the NA concentration increased to 13 times the pre administration level, rectal temperature increased to more than 40 degrees C, and all of the animals died 75 min later. In the group pre-treated with risperidone (0.5 mg/kg), the 5-HT syndrome was completely inhibited, and the NA level increased to 6.5 times the pre-administration level. Ketanserin, a selective 5 HT2A antagonist (5 mg/kg) also inhibited the 5-HT syndrome. In contrast, all of the rats in the group pre-treated with haloperidol (0.5 mg/kg) died earlier than in the saline group. CONCLUSIONS: These results suggest that strong 5-HT2A antagonists such as risperidone, but not dopamine D2 antagonists, counteract lethality due to 5-HT syndrome, and that not only does enhancement of 5-HT activity occur in the 5-HT syndrome, but NA activity also increases. PMID- 10867972 TI - Carbamazepine and valproate monotherapy: feasibility, relative safety and efficacy, and therapeutic drug monitoring in manic disorder. AB - RATIONALE: Search for alternatives to lithium therapy for mood disorders commenced with anticonvulsants, carbamazepine (CBZ) and valproic acid (VPA), in the late 1970s. The comparative safety and efficacy data of CBZ and VPA monotherapy in patients with bipolar disorder remain to be established. OBJECTIVES: The main objectives of the study were to assess the relative antimanic efficacy and safety of CBZ and VPA; to study the feasibility of using either, as a first line anti-manic agent; to investigate and generate clinically relevant parameters involving therapeutic drug monitoring of the two drugs. METHODS: After a 2-day screening period, suitable patients (n = 30) were randomly assigned to treatment with CBZ or VPA. Both the drugs were started with an average dose of approximately 20 mg/kg body weight per day. Further increment in dose was carried out at weekly intervals, guided by clinical improvement, serum levels and treatment emergent adverse events. The primary efficacy measure in the protocol was defined as a change from baseline to endpoint in total score on the Young Mania Rating Scale. A favourable clinical response was defined a priori as a decrease of more than 50% from baseline in Young Mania Rating Scale total score. RESULTS: Both CBZ and VPA were found to be efficacious as single first line anti-manic agents, however VPA proved to be better. Using the intent-to treat analysis, the VPA group showed a significant fall in YMRS total scores after week 1 while the CBZ group showed a significant fall after week 2. In the primary efficacy analysis, valproate group experienced significantly greater mean improvement in Young Mania Rating Scale total score than the CBZ group. Of the VPA treated patients, 73% showed a favourable clinical response while 53% of the patients on CBZ responded favourably. In the CBZ group, significantly more patients received rescue medication during the week 2 and the requirement was quantitatively more as compared to the VPA group. The steady state serum concentration (Css) of CBZ ranged from 3 to 9 micrograms/ml; however, it did not appear to correlate with the dose or clinical response. The Css of VPA ranged from 50 to 100 micrograms/ml; a linear correlation was found between the dose and serum levels of VPA as well as between weekly rise in serum levels and clinical response. Weekly dose escalations of VPA also correlated positively with corresponding rise in serum levels. Significantly more patients in the CBZ group reported adverse events, including nausea, vomiting and dizziness, than VPA. CONCLUSIONS: The findings from this study suggest that both CBZ and VPA monotherapy is feasible for treatment of acute mania; however, VPA is more efficacious in terms of its early onset of action, lesser requirement for rescue medication and better tolerability. Further work needs to be undertaken to characterise the manic patients in terms of their differential psychopharmacologic response profile. PMID- 10867973 TI - Anatomical and pharmacological specificity of the rewarding effect elicited by microinjections of morphine into the nucleus accumbens of mice. AB - RATIONALE: The involvement of nucleus accumbens (NAc) in initiating opiate induced reward has been difficult to demonstrate in rats, and has not been studied in mice. OBJECTIVES: To determine whether a reward-sensitive strain of mice (BALB/c) would self-administer morphine directly into the NAc or sub-regions of the dorsal striatum. METHODS: BALB/c mice were unilaterally implanted with a guide-cannula above either the NAc, the anterior caudate putamen, or the posterior caudate putamen. On each experimental day, a stainless-steel injection cannula was inserted into the guide cannula to test the capacity for morphine self-administration (6.5 pmol or 65 pmol/50 nl) using a spatial discrimination task in a Y maze. RESULTS: Only the ventro-medial NAc group discriminated between the arm enabling a microinjection of morphine and the neutral arm. Once self administration had been acquired, the effects of a pretreatment with two doses of the opiate antagonist naloxone (0.4 mg/kg or 4 mg/kg) were tested. Both doses slightly disrupted self-administration on the first 2 days. Only subjects receiving the 4-mg/kg dose exhibited an extinction of self-administration, related to an increasing number of jump attempts; none of the other opiate withdrawal-associated signs were observed. Self-administration was reinstated when naloxone was replaced with saline. CONCLUSIONS: (1) Medio-ventral NAc is involved in acute rewarding effects of opiates in mice. (2) Neither anterior nor posterior dorsal striatum seem to participate in these effects. (3) NAc is involved in jumping caused by naloxone-induced extinction, a behavior presumably revealing an aversive state associated with the unexpected suppression of reward. PMID- 10867974 TI - Dissociable effects of the kappa-opioid receptor agonists bremazocine, U69593, and U50488H on locomotor activity and long-term behavioral sensitization induced by amphetamine and cocaine. AB - RATIONALE: Mesolimbic dopaminergic neurotransmission plays a critical role in the locomotor effects of psychostimulant drugs, but a general involvement in the induction of long-term psychostimulant sensitization is questionable. By influencing dopaminergic neurotransmission, opioid drugs can alter the behavioral effects of psychostimulants. OBJECTIVES: The effects of the kappa-opioid receptor agonists bremazocine, U69593, and U50488H on the locomotor stimulant and the long term sensitizing effects of amphetamine and cocaine were investigated in rats. Unlike U69593 and U50488H, bremazocine is also an antagonist at mu- and delta opioid receptors, as well as an agonist at a subtype of delta-opioid receptors inhibiting dopamine D1 receptor-stimulated adenylate cyclase. METHODS: Bremazocine, U69593, and U50488H were administered prior to amphetamine and cocaine, and locomotor activity was measured. In separate studies, the opioids were co-administered with amphetamine and cocaine for 5 days, and locomotor sensitization was assessed 3 weeks post-treatment. RESULTS: Bremazocine and U69593 attenuated the psychomotor stimulant effects of amphetamine and cocaine. U50488H attenuated the locomotor effect of cocaine and biphasically affected amphetamine-induced locomotion, i.e., suppression followed by stimulation. Bremazocine prevented the development of amphetamine-induced but not cocaine induced long-term sensitization. Neither U69593 nor U50448H affected the induction of long-term amphetamine or cocaine sensitization. CONCLUSIONS: In agreement with previous studies, the present data suggest that differential mechanisms underlie the acute stimulant versus the long-term sensitizing effects of psychostimulants, and the induction of long-term sensitization by amphetamine versus cocaine. Stimulation of kappa-opioid receptors does not seem to block the induction of long-term psychostimulant sensitization. Thus, bremazocine is likely to block the induction of amphetamine sensitization through a non-kappa-opioid receptor mechanism. We suggest that this effect of bremazocine is the result of its unique agonist action at a subtype of delta-opioid receptors, thereby acting as a functional dopamine D1 receptor antagonist. This would be consistent with the literature showing that the induction of long-term amphetamine sensitization depends on the activation of dopamine D1 receptors. In addition, the present data are in keeping with studies showing that dopamine neurotransmission is not critical for the induction of long-term cocaine sensitization. PMID- 10867975 TI - Acute administration of amitriptyline and mianserin increases dopamine release in the rat nucleus accumbens: possible involvement of serotonin2C receptors. AB - Previous studies of conventional tricyclic and non-tricyclic antidepressants have suggested that a number of these drugs display considerable pharmacological activity at 5-HT2C receptors in the brain. There is evidence that 5-HT2C receptors are involved in the control of the activity of the central dopaminergic system. Therefore, the effects of amitriptyline (5 mg/kg and 10 mg/kg i.p.) and of the atypical antidepressant mianserin (2.5 mg/kg and 5 mg/kg i.p.) were studied on the extracellular concentration of dopamine (DA) in the nucleus accumbens of chloral hydrate-anesthetized rats, using intracerebral microdialysis. Amitriptyline and mianserin significantly increased DA release (+31.1 +/- 7.9% and +33.6 +/- 4.3%, respectively) at the higher doses. In addition, lower doses of mianserin (2.5 mg/kg i.p.) and amitriptyline (5 mg/kg i.p.) blocked the inhibitory action of RO 60-0175 (1 mg/kg i.p.), a selective 5 HT2C receptor agonist, on DA release. The effect of RO 60-0175 (1 mg/kg i.p.) was completely blocked by SB 242084 (2.5 mg/kg i.p.), a selective and powerful 5-HT2C receptor antagonist. Taken together, these data indicate that amitriptyline and mianserin increase DA release in the nucleus accumbens by blocking 5-HT2C receptors. PMID- 10867976 TI - Antidepressants preferentially enhance habituation to novelty in the olfactory bulbectomized rat. AB - RATIONALE: The mechanisms whereby antidepressant drugs exert their therapeutic effects remain unknown. Responses to stressful stimuli are currently thought to contribute to the onset and course of affective disorders. It has been postulated that antidepressants might act by ameliorating response patterns to challenging life events, such as processes of reactivity and/or habituation. OBJECTIVE: Using the olfactory bulbectomy (OBX) rat model, this study examined the effects of various antidepressants on measures of reactivity and habituation in behavioral tests assessing responses to novel stimuli. METHODS: Sham-operated and OBX rats received 21 daily injections of fluoxetine (10 mg/kg), amitriptyline (10 mg/kg), desipramine (10 mg/kg), buspirone (3 mg/kg), or vehicle. Forty-eight hours after the last injection, animals were tested in the open field, elevated plus maze, and startle apparatus. For each test, time series data were collected and fit with exponential random effects models, in which estimated parameters assessed behavioral reactivity and habituation. RESULTS: Relative to sham controls, OBX rats displayed increased total locomotor activity in the open field and exhibited increased open arm behavior in the elevated plus maze. Through comparison with zinc sulfate-treated anosmic controls, these OBX-induced increases were attributed to both an augmentation of initial reactivity due to anosmia and an attenuation of the average rate of habituation. Chronic antidepressant treatment did not reduce the anosmia-related initial reactivity levels of OBX rats to that of sham controls. Rather, the antidepressants evoked their restorative effects by increasing the rate of habituation. CONCLUSIONS: These findings suggest that antidepressants restore normal responding by permitting more effective adaptation to novel stimuli. PMID- 10867977 TI - Self-administration of remifentanil, an ultra-short acting opioid, under continuous and progressive-ratio schedules of reinforcement in rats. AB - RATIONALE: Remifentanil is a mu-opioid agonist with an exceptionally short duration of action. Evaluating remifentanil's effects within the self administration model of drug abuse may provide insight into the relationship between a drug's duration of action and its effectiveness as a reinforcer. OBJECTIVES: This study was conducted to establish a dose-effect function for intravenous remifentanil self-administration in rats and to assess the drug's ability to maintain responding under intermittent schedules of reinforcement. METHODS: Inter-infusion intervals were recorded under two continuous reinforcement schedules of remifentanil self-administration. In the fixed-dose schedule, the unit dose (0.25-32 micrograms/kg) was held constant within sessions but varied across sessions. In the variable-dose schedule, four different doses were self-administered in random order within each session. For comparison, heroin (6.25-125 micrograms/kg) was studied with the variable-dose schedule. Remifentanil and heroin were also compared under a progressive-ratio schedule of reinforcement in which the response requirements increased exponentially with each successive infusion until responding ceased within each session. RESULTS: Under the continuous-reinforcement schedules, inter-infusion intervals for both drugs increased monotonically as a function of dose, with the remifentanil curve being considerably flatter. Under the progressive-ratio schedule, breaking points varied as an inverted-U shaped function, and the highest breaking points maintained by remifentanil and heroin were similar. At the doses that maintained the highest breaking points under the progressive-ratio schedule, post-infusion pauses under the continuous-reinforcement schedule were about three times shorter with remifentanil than with heroin. CONCLUSIONS: Although rates of self administration are clearly influenced by a drug's duration of action, the ability to maintain responding under intermittent schedules of reinforcement may be independent of duration of action. PMID- 10867978 TI - Role of dopaminergic system in reactivity to spatial and non-spatial changes in mice. AB - RATIONALE: While experimental evidence shows that dopamine (DA) systems have an important role in locomotor function and in motivation, their role in the reactivity to spatial and non-spatial novelty is less well established. OBJECTIVE: In this study, we investigated the effects of dopaminergic pharmacological manipulation on the capability of CD1 mice to encode spatial and non-spatial information in an open field with objects. METHODS: The effects of systemic administration of: (1) selective D1 and D2 antagonists (SCH23390, 0.015 mg/kg or 0.020 mg/kg; sulpiride, 10 mg/kg or 20 mg/kg); (2) direct and indirect DA agonists (apomorphine, 1 mg/kg or 2 mg/kg; amphetamine, 1 mg/kg or 2 mg/kg) were studied. RESULTS: On the one hand, systemic administration of either D1 or D2 antagonists induced a selective impairment in the detection of spatial change but did not affect reaction to non-spatial novelty. On the other hand, amphetamine induced a selective decrease in exploratory activity in the first three sessions. This decrease did not affect the ability of mice to react to spatial change, but a dose-dependent decrease was observed in reactivity to non spatial novelty. Such an effect does not seem to be due to amphetamine-induced hyperactivity or to non-DA mechanisms, since apomorphine induced a similar neophobic profile, without affecting locomotion. CONCLUSIONS: Taken together, these results demonstrate that manipulations of DA transmission affect reactivity to spatial and non-spatial novelty. In particular, we suggest that these two behavioral responses are modulated in opposite ways by the DA system. PMID- 10867979 TI - Pharmacological demonstration of the differential involvement of protein kinase C isoforms in short- and long-term memory formation and retrieval of one-trial avoidance in rats. AB - RATIONALE: The hippocampal protein kinase C (PKC) family is involved in the early events of consolidation of long-term potentiation (LTP) and long-term memory (LTM). Results so far are indecisive about which PKC isoform is involved and as to whether any of them plays a role in short-term memory (STM) processes, which have recently been shown to be separate from those of LTM in the hippocampus dependent one-trial step-down inhibitory avoidance task. OBJECTIVES: To measure the effect of two PKC inhibitors, one (Go 6976) selective to the calcium dependent isoforms alpha and beta I, and the other (Go 7874) unspecific as to PKC isoforms on the formation and retrieval of STM and LTM of one-trial inhibitory avoidance. METHODS: Rats bilaterally implanted with cannulae in the CA1 region of the dorsal hippocampus were trained in one-trial step-down inhibitory avoidance. The effect of these two drugs on STM and LTM formation was investigated as follows. Animals were infused 10 min before or 50, 110, or 170 min after inhibitory avoidance training with a vehicle (2% dimethylsulfoxide in saline), or with Go 6976 (0.92 nM or 4.6 nM) or Go 7874 (1.96 nM or 8 nM) dissolved in the vehicle. Infusion volume was 0.5 microliter in all cases. Animals were tested 1.5 h and 3 h after training for STM and at 24 h for LTM. In order to study the effects of these compounds on retrieval, they were infused into the hippocampus 10 min prior to STM testing at 3 h (see above) or 10 min before LTM testing at 24 h. In addition, the effect of Go 6976 and Go 7874 was studied on general activity measured in an open field, and on performance in an elevated plus maze. RESULTS: STM was suppressed by 4.6 nM Go 6976 given 10 min before or 50 min after training. LTM was cancelled by the higher dose of the two compounds given 10 min before, or 50 min or 110 min after training. None of the two compounds infused 170 min post-training affected the retrieval of STM measured 10 min later. However, both compounds given 10 min before testing inhibited the retrieval of LTM measured at 24 h. This effect cannot be attributed to influences on locomotor activity or anxiety levels, since the drugs had no effect on performance in the open field but were mildly "anxiogenic" (pro-conflict) and reduced the number of entries into open and closed arms and rearings. CONCLUSIONS: LTM consolidation requires in part alpha- and/or beta 1-PKC and in part other PKC isoforms. STM formation requires instead only alpha and/or beta I-PKC and during a more limited period of time. In addition, PKC appears not to be necessary for the retrieval of STM, but is crucial for the retrieval of LTM. These findings further point to a biochemical separation of STM and LTM, as ascertained in numerous previous studies. PMID- 10867980 TI - Comparison of the effects of clonidine and yohimbine on spontaneous pupillary fluctuations in healthy human volunteers. AB - RATIONALE: Spontaneous fluctuations in the size of the pupil in darkness are a recognized index of sleepiness, and these fluctuations can be quantitatively measured using the pupillographic sleepiness test (PST). The central noradrenergic system is believed to play a role in the maintenance of alertness, and there is evidence that alpha 2-adrenoceptor agonists (e.g. clonidine) decrease the activity of central noradrenergic neurones, whereas alpha 2 adrenoceptor antagonists (e.g. yohimbine) have the opposite effect. OBJECTIVE: To evaluate the effects of single oral doses of clonidine and yohimbine on spontaneous pupillary fluctuations in healthy volunteers. METHODS: Sixteen healthy male volunteers (18-25 years) participated in four weekly sessions, each associated with one oral drug condition (clonidine hydrochloride 0.2 mg, yohimbine hydrochloride 22 mg, clonidine hydrochloride 0.2 mg + yohimbine hydrochloride 22 mg), according to a balanced double-blind design. Pupil diameter was recorded continuously over 11 min in darkness using a dedicated monocular video pupillometer. Average pupil diameter, power of pupil diameter fluctuations (obtained by fast Fourier transformation), and the pupillary unrest index (PUI), a measure of cumulative changes in pupil size, were computed. Autonomic functions known to be sensitive to centrally acting noradrenergic drugs (blood pressure, heart rate and salivary output) were recorded. Subjective "alertness", "anxiety" and "contentedness" were rated using visual analogue scales. Measurements were carried out 2 h after drug ingestion. Data were analyzed by ANOVA followed by Dunnett's corrected t-test (criterion of significance P < 0.05). RESULTS: Clonidine decreased systolic and diastolic blood pressure, salivation and subjectively rated alertness, and tended to decrease pupil diameter, and to increase the power of pupillary fluctuations and PUI. On the other hand, yohimbine increased systolic and diastolic blood pressure, salivation, pupil diameter and decreased PUI. When the two drugs were given in combination they reduced each other's effects. CONCLUSIONS: These results confirm the alerting effect of the centrally acting noradrenergic activating drug yohimbine and the opposite effect of clonidine, and the suitability of the PST to detect these effects. PMID- 10867981 TI - Effects of dopaminergic drugs on delayed reward as a measure of impulsive behavior in rats. AB - RATIONALE: Impulsive individuals exhibit an exaggerated preference for immediate rewards over delayed but larger rewards, perhaps because they value the delayed rewards less. Dopamine (DA) has been postulated to mediate the incentive value of rewards, thus it may also mediate the exaggerated preference for immediate rewards in impulsive individuals. In this paper, we investigate the effects of DA agonists and antagonists on the value of delayed versus immediate rewards. OBJECTIVES: The study had three main objectives: (1) to determine the effects of D1 and D2 type DA antagonists on the value of delayed rewards, (2) to determine the effect of the indirect DA agonist d-amphetamine on the value of delayed rewards, (3) to determine the sensitivity of the adjusting amount (AdjAmt) procedure to acute one-day changes in delay to reward, amount of reward, deprivation level and starting amount. METHODS: In the AdjAmt procedure, rats choose between an adjusting amount of water given immediately (adjusting alternative) and a constant 150 microliters water delayed by 4 s (standard alternative). The immediate amount of water is adjusted across trials until the rat chooses both alternatives with equal frequency (indifference point). The final adjusted amount is an indicator of the subjective value of the standard alternative. RESULTS: The D1/D2 antagonist flupenthixol (25, 50, and 100 micrograms/kg) and the D2 antagonist raclopride (40, 80, and 120 micrograms/kg), decreased the indifference points, whereas the D1 antagonist SCH 23390 (5, 10, and 20 micrograms/kg) did not affect the indifference point. All three DA antagonists slowed responding. The indirect DA agonist amphetamine (0.5, and 1.0 mg/kg) had effects opposite to those of the DA antagonists, it decreased choice latency, increased the number of trials completed and increased the indifference point. Decreasing the water deprivation level (6, 24, and 48 h) had no effect on the indifference points but slowed responding. Increasing the delay to reward (2, 4, and 8 s) and decreasing the amount of water available for choosing the standard alternative (300, 150, 75 microliters) decreased the indifference point. CONCLUSIONS: The results indicate that amphetamine increased the value of delayed rewards (decreased impulsivity) and that D2 but not D1 receptor antagonists decreased the value of delayed rewards (increased impulsivity). The procedure was sensitive to acute 1-day changes in delay and magnitude of reward. PMID- 10867982 TI - Effects of AMPA/kainate receptor blockade on responses to dopamine receptor agonists in the core and shell of the rat nucleus accumbens. AB - The present experiments were conducted to investigate effects of alpha-amino-3 hydroxy-5-methyl-4-isoxazoleprionic acid (AMPA)/kainate receptor blockade (CNQX, NBQX) on locomotor responses to D2/3 (7-OH-DPAT) and D1 [(+)-SKF 38393] dopamine receptor agonists in the nucleus accumbens (NAS) core and shell. CNQX (0.25-0.5 microgram) microinjected into the NAS core or shell did not affect baseline locomotor activity. 7-OH-DPAT (2.5-5 micrograms) decreased locomotor activity. Co administration of CNQX (0.5 microgram) increased the effects of 7-OH-DPAT (5 micrograms) in the NAS core and shell. A similar increase was observed with NBQX (0.5 microgram) in the NAS shell. (+)-SKF 38393 (5 micrograms) into the NAS core and shell increased locomotor activity after 30 min; this effect was not altered by CNQX (0.5 microgram). As the D2/3 dopamine agonist (-)-quinpirole (2 micrograms) increased effects of (+)-SKF 38393 (5 micrograms) in NAS shell but not core, lack of site-selective effects of (+)-SKF-38393 and of 7-OH-DPAT within NAS is not attributable to drug diffusion. The previous observation that glutamate effects on locomotor activity depend on the relative involvement of D1 or D2/3 dopamine receptors in the NAS was based on the dopamine-depletion model. The present results demonstrate differential interactions of AMPA receptor blockade with dopamine agonists in "dopamine-intact" animals. PMID- 10867983 TI - Cholinergic neurotransmission influences covert orientation of visuospatial attention in the rat. AB - RATIONALE: Both monkey and human studies have suggested that attentional orienting may be mediated by the cholinergic neurotransmitter system. OBJECTIVES: The purpose of the present study was to examine whether the cholinergic agonist (nicotine) and/or antagonist (scopolamine) influence covert orientation in the rat. METHODS: Rats carried out a visual reaction time task to measure covert orienting of attention following systemic administration of nicotine or scopolamine. RESULTS: Nicotine reduced reaction times, abolishing the validity effect by differentially speeding the reaction times for invalidly cued targets. Conversely, scopolamine increased the validity effect by disproportionately lengthening reaction times to invalidly cued targets. CONCLUSIONS: Taken together, these data indicate that cholinergic transmission represents an important neurochemical substrate of visuospatial attention, specifically influencing disengagement or movement of attention. PMID- 10867984 TI - Effects of the nicotinic antagonist mecamylamine on inspection time. AB - RATIONALE: Several lines of evidence suggest that nicotinic acetylcholine receptors (nAchRs) are involved in speed of information processing, and inspection time appears to be particularly sensitive to nicotinic manipulation. OBJECTIVE: The present study sought to examine the effects of the nAchR antagonist mecamylamine on inspection time. Furthermore, the extent to which the anticholinesterase donepezil would reverse the effects of mecamylamine on inspection time was also examined. METHODS: A double-blind, repeated measures design was employed. Subjects (n = 6) received placebo, mecamylamine (20 mg PO) or mecamylamine (20 mg PO) and donepezil (5 mg PO). Inspection time and physiological measures were then assessed. RESULTS: The mecamylamine condition and the mecamylamine and donepezil condition were associated with an increase in heart rate, when compared to the placebo condition. There was a significant slowing of inspection time in the mecamylamine condition; compared to placebo, which was partly reversed by donepezil. CONCLUSIONS: The slowing of inspection time following mecamylamine is consistent with the role of nAchRs in speed of information processing, and add to the evidence that IT may in part index nAchR system integrity. PMID- 10867985 TI - Serotonin transporter (5-HTT) promoter genotype may influence the prolactin response to clomipramine. AB - RATIONALE: A 44-base-pair insertion/deletion polymorphism in the promoter region of the human serotonin (5-HT) transporter (5-HTT) gene gives rise to a bi-allelic polymorphism designated long (l) and short (s). The s variant is associated with a lower expression of 5-HTT sites and a reduced efficiency of 5-HT reuptake. OBJECTIVE: The aim of the present study was to determine whether the increase in brain 5-HT function produced by acute 5-HT reuptake blockade is influenced by the 5-HTT promoter l/s polymorphism. METHODS: We measured the increase in plasma prolactin that follows acute administration of the tricyclic antidepressant clomipramine as an index of 5-HT neurotransmission in 14 healthy female subjects (7 with ss genotype and 7 with ll genotype) using a placebo-controlled crossover design. RESULTS: Clomipramine-induced prolactin release was significantly greater in subjects with the ll genotype. CONCLUSION: Our findings suggest that acute 5 HT reuptake blockade produces a greater increase in 5-HT neurotransmission in subjects with the ll genotype than in those with an ss genotype. These results are consistent with clinical data indicating that subjects with an ss genotype may have a poorer therapeutic response to selective serotonin reuptake inhibitor (SSRI) monotherapy. PMID- 10867986 TI - [Omsk hemorrhagic fever--current status of the problem]. AB - The author sums up the data on the history of discovery and research of Omsk hemorrhagic fever (OHF), comparative characteristics of its natural foci, and clinical features of epizootic and epidemic processes at the end of the 1940s and at present. Presents the modern concept of OHF virus ecology and discusses differences of its biological properties in the strains isolated from different sources. Based on analysis of many-year findings and major publications, the author offers a long-term epizootological and epidemic prognosis for OHF in endemic territories. PMID- 10867987 TI - [Isolation of the West Nile Fever virus from human patients during an epidemic outbreak in the Volgograd and Astrakhan regions]. AB - Two strains of West Nile virus LEIV 27889 Vig and Ast 986 were isolated from the brain of a dead subject and from the blood of a patient, respectively, during an outbreak of serous meningitis and meningoencephalitis in July-September, 1999, in the Volgograd region, Krasnodar territory, and Astrakhan region. These strains reacted with convalescent sera in hemagglutination inhibition test, which proves their etiological role in this outbreak. PMID- 10867988 TI - [Isolation and characteristics of low molecular weight recombinant peptides, representing antigenic domains in the NP protein of lymphocytic choriomeningitis virus]. AB - High- and low-molecular recombinant peptides of LCM virus nucleoprotein, representing individual immunodominant antigenic sites, were obtained in pJC40 expressing vector and studied in solid-phase enzyme immunoassay. 2-7% proteins resultant from total cellular synthesis are recombinant peptides. Comparative analysis of antigenic properties of recombinant peptides and native viral protein showed that recombinant peptides are virtually not inferior to native viral protein in antigenic properties. PMID- 10867989 TI - [Variants of influenza H1N1 virus, adapted to mouse lungs and inhibitors of mouse serum inhibitors differ by hemagglutinin characteristics and structure]. AB - Influenza virus A/USSR/90/77 variants adapted to mouse blood serum (USSR/90-MS) and lungs (USSR/90-ML) resistant to beta-inhibitors of mouse serum differ by biological properties and hemagglutinin (HA) structure. One of glycosylation site (GS) located at the tip of HA spike near the receptor binding site is lost because of mutations in both variants: GS 158 (Asn158Asp substitution) in USSR/90 MS and GS131 (Asp131Asp substitution) in USSR/90-ML. Probably adaptation to mouse lungs and serum represents adaptation to different types of receptor molecules. From these data we conclude that mouse lungs and/or bronchi and serum contain different beta-like inhibitors. Presumably the inhibitors in the lungs contain some additional factors in comparison with the serum, adaptation to which allows a wider spectrum of virus resistance to homologous and to many other normal animal sera. PMID- 10867990 TI - [Cellular sensitivity to the effect of interferon during different forms of viral pathology]. AB - A new approach to evaluation of interferon (IFN) status of patients with acute respiratory viral infections, herpes, and urogenital infections has been developed. It consists in measurement of IFN-alpha and IFN-gamma production by leukocytes primed with IFN-alpha and IFN-gamma, respectively, and subsequent identification of groups of patients whose lymphocytes differ by their in vitro response to IFN priming: sensitive to IFN-alpha and IFN-gamma; sensitive to IFN alpha but not to IFN-gamma; sensitive to IFN-gamma but not to IFN-alpha; not sensitive to IFN-alpha and IFN-gamma. Such analysis helps understand the mechanisms of IFN system deficiency in infectious diseases and promotes more effective IFN therapy due to selection of patients whose cells retain in vitro sensitivity to a certain IFN type. PMID- 10867991 TI - [Clinical characteristics of tick-borne encephalitis complicated by Lyme borreliosis]. AB - Clinical characteristics of mixed tick borne encephalitis (TBE) + Lyme borrelliosis (LB) infection and monoinfections are compared. Eighty-five patients with TBE + LB mixed infection serologically verified by EIA and 87 with isolated TBE, who fell ill in 1996, were examined. Among patients with mixed infection, cases with blurred TBE predominated; severe forms (meningeal and focal) were almost two times less incident than in TBE monoinfection. Typical clinical symptoms of LB were observed in 63.5% patients, while in the rest 36.5% LB manifested only by circulation of antibodies to B. burgdorferi. Comparative analysis of patients with mixed TBE + LB infection and TBE monoinfection confirmed a more benign course of TBE during the acute period in patients with mixed infection. Mixed infection should be ruled out or confirmed by thorough clinical examinations with obligatory detection of antibodies to agents of both diseases. PMID- 10867992 TI - [Variability of hemagglutinin from strains of influenza virus A (H3N2), isolated in Russian from 1989 to 1999]. AB - Analysis of 154 strains isolated in Russia and CIS countries in 1989-1999 showed that influenza virus A(H3N2) caused epidemics and epidemic rises 8 times, circulating together with A(H1N1) and B viruses. Antigenic drift was revealed using polyclonal and monoclonal antibodies. Analysis of antigenic properties of the viruses in the population showed that strains isolated during the same year were usually variants of one or rarely two reference strains. A drop of isolation rate of A(H3N2) strains on chick embryos in recent years was associated with increase in these strains' sensitivity to MDCK culture. Differences in amino acid sequences of epidemic and reference strain hemagglutinins were detected. The number of positions in which the changes were detected varied from 6 to 16 in all antigenic sites: A, B, C, D. and E. PMID- 10867993 TI - [Acute hepatitis B in anti-HCV-positive patients]. AB - Seventy-six anti-HCV-positive patients with acute hepatitis B were observed. HBsAg, anti-HBc IgM, and anti-HCV were detected in the sera of all patients. During the acute phase of illness, HBV DNA was present in the sera of 90.8% patients, but not HCV RNA. The clinical picture of acute hepatitis B in anti-HCV positive patients corresponded to HBV monoinfection with prolonged intoxication and a more benign biochemical course. Out of 10 patients observed during 2 years chronic mixed HBV/HCV hepatitis was diagnosed in 1 patient, in 8 patients only HCV infection was diagnosed, and in 6 cases HCV RNA was detected. Virus interference may be responsible for successive alternative dominant replication of HBV and HCV. PMID- 10867994 TI - [Adenovirus KR95, isolated from chickens during an outbreak of hydropericarditis, is the pathogen of this disease]. AB - Virus agent KR95 was isolated from the liver of dieoff chickens during an outbreak of hydropericarditis syndrome at a poultry farm in Russia. Electron microscopic examination of the virus morphology, comparative restriction cleavage map construction, DNA-DNA hybridization, and analysis of structural proteins from purified and disrupted virions showed that the agent is to be classified as type 1 avian adenovirus. PMID- 10867995 TI - [Monoclonal antibodies to Ebola virus: isolation, characteristics, and study of cross reactivity with Marburg virus]. AB - Thirteen hybridoma strains producing monoclonal antibodies (Mabs) to Ebola virus were prepared by fusion of NS-O mouse myeloma cells with splenocytes of BALB/c mice immunized with purified and inactivated Ebola virus (Mayinga strain). Mabs directed against viral proteins were selected and tested by ELISA. Protein specificity of 13 Mabs was determined by immunoblotting with SDS-PAGE proteins of Ebola virus. Of these, 11 hybridoma Mabs reacted with 116 kDa protein (NP) and 2 with Ebola virus VP35. Antigenic cross-reactivity between Ebola and Marburg viruses was examined in ELISA and immunoblotting with polyclonal and monoclonal antibodies. In ELISA, polyclonal antibodies of immune sera to Ebola or Marburg viruses reacted with heterologous filoviruses, and two anti-NP Ebola antibodies (Mabs 7E1 and 6G8) cross-reacted with both viruses. Target proteins for cross reactivity, Ebola NP (116 kDa) and Marburg NP (96 kDa), and VP35 of both filoviruses were detected by immunoblotting with polyclonal and monoclonal antibodies (6G8) to Ebola virus. PMID- 10867996 TI - [Development of immunoenzyme methods for detecting antibodies to Aujeszky's disease virus gB glycoprotein in swine serum]. AB - Four ELISA methods have been developed for detecting antibodies to Aujeszky's disease virus (ADV) glycoprotein gB. Indirect ELISA is based on affinity-purified gB (affi-gB-ELISA); three blocking ELISAs: indirect blocking ELISA (lbgB-ELISA), direct blocking ELISA (db-gB-ELISA), and two-site "sandwich" ELISA (sb-gB-ELISA) are based on monoclonal antibodies to conservative immunodominant epitopes of gB. The specificities and sensitivities of ELISAs were compared with each other and with indirect ELISA based on purified ADV virions (vir-ELISA). Affi-gB ELISA, db gB-ELISA, and sb-gB-ELISA possess 100% sensitivity, ib-gB-ELISA 98% sensitivity, and vir-ELISA 93% sensitivity. Affi-gB ELISA, ib-gB-ELISA, db-gB-ELISA, and sb-gB ELISA possess 100% specificity and vir-ELISA 92% specificity. The efficiency of detection of ADV-specific antibodies by affi-gB ELISA, db-gB-ELISA, and sb-gB ELISA was comparable to that of analogous commercial test. Since db-gB-ELISA is easier to perform than affi-gB-ELISA or sb-gB-ELISA, it is concluded to be the most appropriate test for detecting pigs infected with ADV among non-vaccinated animals. PMID- 10867997 TI - [Myocardial hibernation: another view]. AB - In the following, three newer concepts are brought together: myocardial hibernation, heterogeneity in myocardial blood flow and oxidative metabolism, and effects of hibernating animal serum on non-hibernators. Myocardial hibernation is viewed as a protective mechanism that helps to maintain myocardial integrity and viability by down-regulating contractile function as an adaptation to reduced blood flow. Myocardial flow is considerably heterogeneous. Consequently, oxygen supply to the myocardium is also heterogeneous. Many lines of evidence show a close correlation between regional flow and regional metabolism. In low-flow/low metabolism areas, myocardial function must be reduced, since the myocardium would otherwise undergo necrosis. Thus, others and we hypothesize that function must be down-regulated to induce hibernation in low-flow areas. Because no regional histologic differences exist (the mitochondria are uniformly distributed within the myocardium), the pattern of heterogeneity seems to shift over time. Hence, we hypothesize that such very regional hibernation presents an evolutionary, protective mechanism, permitting subsequent myocardial areas to rest within the ceaselessly working heart. We also hypothesize that this mechanism ensures the down-regulation of function following myocardial ischemia in order to induce myocardial hibernation on a broader level. Surprisingly, a substance (opioid in nature) contained in hibernator serum both induced hibernation-like state in non hibernators and suppressed myocardial oxygen consumption. Thus, we lastly hypothesize that myocardial hibernation is a remnant of the early stages of evolution and is closer to physiologic hibernation than traditionally viewed. PMID- 10867998 TI - [The renin-angiotensin system as the basic principle for hypertension and coronary heart diseases--role of genetic factors]. AB - Since the discovery of renin by Tigerstedt and Bergmann, the renin angiotensin system (RAS) has been recognized as an important modulator of blood pressure and volume homeostasis. Based on these functions a pathophysiological role of the RAS in the pathogenesis of hypertension and other cardiovascular disorders has been postulated. The therapeutic benefit of RAS inhibition by angiotensin converting enzyme (ACE) inhibitors and angiotensin II (ANG II) antagonists in these conditions has been shown. It remains unclear, however, whether the changes in RAS activity associated with cardiovascular disease are primary or secondary factors. It is well known that hypertension and hypertensive end-organ disease is influenced by genetic factors. Gene polymorphisms for virtually all components of the RAS have been described and investigated in clinical studies. It remains to be determined, however, how relevant these findings are for disease etiology. This review, therefore, will attempt to discuss the causal implications of these genetic studies for cardiovascular disease. The role of angiotensinogen and ACE for hypertension, coronary artery disease and other cardiovascular disorders is discussed in this context in an exemplary fashion. PMID- 10867999 TI - [Ventricular fibrillation in intra-atrial cardioversion of atrial fibrillation]. AB - Recently intra-atrial defibrillation has become an interesting alternative to external defibrillation and drug therapy for the treatment of atrial fibrillation. Low-energy intra-atrial defibrillation can be used to restore sinus rhythm f.ex. after a failed external cardioversion or during an electrophysiologic study when the administration of antiarrhythmic drugs should be avoided. Additionally this new technique has led to the development of implantable atrial defibrillators for the treatment of selected patients suffering from chronic atrial fibrillation. Intra-atrial defibrillation seems to be a highly effective and safe method, but little experience exists concerning the outcome so far. Especially the potential risk of inducing ventricular pro arrhythmia is subject of current controversy. We report the case of a 79-year-old patient suffering from WPW syndrome with a concealed bypass tract who was subject to an intra-atrial defibrillation during an electrophysiologic study. At the beginning of the study atrial fibrillation could be converted to sinus rhythm by a single low-energy atrial defibrillation (3 J.). After a short period of time a second intra-atrial defibrillation had to be performed in the same way because of recurrent atrial fibrillation. By this atrial shock ventricular fibrillation was induced, so that high energy external defibrillation became necessary. Analyzing the ECG a correct R-wave synchronization was found, but a rather short preceding RR interval (252 ms). In conclusion, low energy atrial defibrillation is gaining importance as a highly effective new technique to restore sinus rhythm in patients suffering from atrial fibrillation resistant to conventional therapies. Nevertheless potential risks have to be considered such as the induction of ventricular pro-arrhythmia. Therefore, a correct R-wave synchronization is obligatory and shock delivery should be withheld after short RR intervals. Future prospective randomized studies will have to show whether this new technique is really safe enough and superior to the conventional methods for restoring sinus rhythm in patients suffering from atrial fibrillation. PMID- 10868000 TI - [Variable late potentials in long-term ECG of the post-infarct patient at risk for ventricular fibrillation]. AB - BACKGROUND: Ventricular late potentials are found more readily in post-infarction patients who had sustained ventricular tachycardia than in those who survived ventricular fibrillation. Hypothetically, a daytime variability of late potentials might be responsible for this finding. METHOD: Therefore a conventional late potential analysis only performed once a day was compared to a late potential analysis in time and frequency domain repeatedly performed every hour in the Holter-ECG of 160 post-infarction patients (50 patients (= VT-group) with documented, sustained ventricular tachycardia (cycle-length > 230 ms), 50 patients (= VF-group) who survived, documented ventricular fibrillation and 60 patient, without ventricular arrhythmias (= O VT/VF-group)). RESULTS: The conventional analysis showed late potentials in time domain in 72% of the patients in the VT-group, in 40% of patients in the VF-group and in 20% of the patients in the O VT/VF-group. The Holter-ECG showed late potentials to be permanently present in frequency domain in 66% of the patients in the VT-group, in only 6% in the patients in the VF-group and in no patient in the O VT/VF group. However, in at least one analysis we detected late potentials in 84% of patients of the VF-group, in 90% of patients in the VT-group and in 18% of patients in the O VT/VF-group. Transiently detectable late potentials in patients of the VF-group were predominantly seen at heart rate accelerations in the morning hours, ST-segment shifts or transitory decreased heart rate variability. CONCLUSIONS: Post-infarction patients with sustained ventricular tachycardia predominantly have constantly detectable late potentials over 24 hours. In these patients conventional late potential is successful for post-infarction risk stratification at any time of the day. However, in post-infarction patients who survived ventricular fibrillation, late potentials are found to be transitory and only detectable by Holter-ECG. Thus, late potential analysis performed in the Holter-ECG might improve post-infarction risk stratification in patients prone to sudden cardiac death. PMID- 10868001 TI - [Background and evaluation plan of a study on self-management of anticoagulation in patients with non-valvular atrial fibrillation (SMAAF Study)]. AB - The objective of this open, randomized, multicenter study is to investigate the benefits and economic efficiency of self-management of oral anticoagulation in patients with atrial fibrillation (SMAAF study) in comparison with a group of patients given conventional care by a general practitioner or specialist. Two thousand patients suitable for self-management will be assigned at random to either the self-management group or the control group. The numbers of thromboembolic and hemorrhagic complications requiring treatment during the 2 year follow-up period will be recorded as the primary end point. The secondary endpoint variables will be maintenance of the INR value in the individual target range, INR variance, the course of complications over time, and the cost efficiency of self-management compared with the routine procedures. The last of these parameters will include the diagnostic and/or therapeutic measures carried out, the duration of inpatient hospital treatment, and the social consequences (subsequent rehabilitation treatment, inability to work, forced retirement). The estimate of the required number of patients was based on the assumption that during long-term anticoagulant therapy within the framework of primary and secondary prevention 4% of patients with chronic non-valvular atrial fibrillation would have severe thromboembolic of hemorrhagic complications each year. Since this rate can be halved by self-management, a one-tailed chi 2-test of 80% power and a 5% significance threshold would require n = 997 patients per group. The results of the SMAAF study will establish the socioeconomic benefits of self management in patients with non-valvular atrial fibrillation. PMID- 10868002 TI - [Where should the implantable ECG event recorder be placed?]. AB - Aim of our study was the comparison of bipolar ECG quality (with a 4 cm lead distance) at different sites within the anterior thorax to find the preferable implantation site for an ECG event recorder (ECG-ER). In 70 patients a bipolar ECG with a short electrode distance and in the vertical position was registered at the following sites: left and right subclavicular, left and right parasternal (4th-5th ICR), left and right anterior axilla (4th-5th ICR), at the heart apex and subxiphoidal. Then it was compared to the standard lead II. In 34 patients, an additional comparison between vertical and horizontal ECG registration was performed at the above mentioned sites. During implantation of an ECG-ER in 5 patients, ECG signals were compared with electrodes placed towards the skin or towards the muscle. The best ECG quality (greatest QRS amplitude, visible P-wave and pacemaker spike, measurable QT period and bundle-branch block) and the best agreement with standard lead II was found in 67% left parasternal, significant less often (p < 0.001) right parasternal (14.3%), left subclavicular (7.1%), apical (5.7%), and subxiphoidal (4.3%). In a vertical electrode position a significantly higher QRS amplitude and a more often visible P wave was found in comparison to a horizontal electrode position. In all cases, there was good agreement between bipolar surface ECG at the implantation site and ECG-ER stored signals. When the ECG-ER is positioned with electrodes towards the muscle, significant noise-signal occurred in all 5 patients. Only in 3 patients was a P wave visible, but with a slightly greater QRS amplitude than in ECG-ERs positioned with electrodes towards the skin. From these results, it is recommended to implant ECG-ERs vertically with electrodes towards the skin and in the parasternal position. PMID- 10868003 TI - [Morphological changes in correlation with so-called idiopathic ventricular tachycardias from the right ventricular outflow tract]. AB - "Idiopathic" ventricular tachycardia is an exclusion diagnosis. The underlying reasons and mechanisms of "idiopathic" ventricular tachycardias are still not completely understood. Recent investigations showed a high prevalence of morphological abnormalities in the right ventricle of patients with "idiopathic" ventricular tachycardia out of the right ventricular outflow tract, which could often be correlated with the origin of the ventricular tachycardia. These described abnormalities were not uniform. Here we report about a patient suffering from drug-refractory "idiopathic" ventricular tachycardia for 10 years. This is the first report in which the origin of an "idiopathic" ventricular tachycardia could be localized by right-ventricular angiography, magnetic resonance tomography and electrophysiological study in the area of an interventricular septal thickening of the right ventricular outflow tract and cured by radiofrequency catheter ablation. PMID- 10868004 TI - Impact of intravascular ultrasound guidance on directional coronary atherectomy. AB - In contrast to the luminogram of coronary angiography, intravascular ultrasound (IVUS) has proven to accurately assess both coronary lumen and vessel morphology due to its 360 degrees imaging capacity. Directional coronary atherectomy (DCA) improves the coronary lumen by removing plaque mass rather than stretching the vessel and compressing the plaque as with conventional percutaneous transluminal coronary angioplasty. In an attempt to optimize the procedural result of DCA we prospectively investigated the impact of IVUS guidance in a head to head comparison to on-line quantitative coronary angiography (QCA) on the result of DCA. In 16 consecutive patients IVUS demonstrated significant residual plaque mass after DCA irrespective of a satisfactory angiographic result. After a mean of 9 +/- 2 cuts luminal improvement was obtained with an area stenosis by angiography of 39 +/- 17% and by IVUS of 50 +/- 10% (p < 0.05), a diameter stenosis by angiography of 23 +/- 10% and IVUS of 35 +/- 14% (p < 0.05) and finally a minimal lumen diameter (MLD) by angiography of 2.9 +/- 0.5 mm and by IVUS of 2.3 +/- 0.5 mm (p < 0.005). After both on-line QCA and IVUS measurements a second series of 7 +/- 2 cuts were initiated to debulk more atheroma and improve stenosis dimensions. After additional cuts IVUS revealed further luminal improvement with an area stenosis by angiography of 25 +/- 16% and IVUS of 21 +/- 18% (n.s.), a diameter stenosis by angiography of 16 +/- 11% and by IVUS of 13 +/ 19% (n.s.) and finally a MLD by angiography of 3.1 +/- 0.5 mm and by IVUS of 2.8 +/- 0.3 mm (p < 0.05). Intraprocedural use of IVUS is superior to on-line QCA to assess the immediate result of DCA. IVUS-guided DCA results in more effective atheroma debulking than luminographic evaluation. Results of larger follow-up studies are needed to substantiate the intraprocedural advantage of IVUS with DCA. PMID- 10868005 TI - [Fractional flow reserve as a deciding criterion for intervention in patients with 50% coronary stenoses and impaired myocardial perfusion]. AB - BACKGROUND: A fractional flow reserve (FFRmyo) < 0.75 is a well validated parameter for significance of coronary stenoses in cases of normal myocardial function. We used the FFRmyo limit in patients with impaired myocardial perfusion by myocardial infarction and/or hypertension for intermediate stenoses of the LAD for decision to PTCA and checked the indication by clinical follow-up. METHODS: In 20 pts (5 women) with chest pain and visual 50 D% LAD stenoses, the FFRmyo was obtained by using a RADI-Pressure-Wire, the CFR by a densitometric technique (HODGSON), and the geometry of stenosis (minimal lumen diameter and diameter stenosis) by quantitative coronary angiography (QCA). EF and the kinetics of the anterolateral wall (expressed as radial shortening fraction) were measured by laevography. RESULTS: The mean age of our 20 pts. was 59.4 years: 13 of the pts. (65%) had a history of hypertension, 9 (45%) pts. a history of myocardial infarction. The mean diameter stenosis was 50.8%. The mean value of CFR was 2.9. The FFRmyo ranged from 0.66 to 0.90, the mean value was 0.78. The 12 pts. with FFRmyo > or = 0.75 (60%, group A) were treated with the usual anti-anginal medications. A PTCA was performed only in patients with FFRmyo < 0.75 (N = 8 (40%), group B). Except for one pt. with instent restenosis, in the 7 pts. with denovo stenoses stent implantation was performed. Significant differences between the groups A and B were seen only for the total number of myocardial infarctions (8/12 vs. 1/8) and diameter stenosis (48.5% vs. 54.3%). All lesions of group B had a diameter stenosis of 50% or higher. CFR correlated significantly with the radial shortening fraction (r = 0.75), minimal lumen diameter (r = -0.51) and diameter stenosis (r = -0.46). FFRmyo correlated with diameter stenosis (r = 0.47) only. All pts. treated with PTCA were primarily free of pain or reduced angina at least 1 CCS stage; only one developed an angina due to a restenosis (74 D%) 2 months after PTCA and stent implantation. The pts. of group A did not get worse, nor were they readmitted within 6 to 13 months after catheterization. CONCLUSIONS: Pts. with 50 D% stenoses, impaired myocardial perfusion and FFRmyo < 0.75 had a good long-term benefit concerning clinical and angiographic result. No pts. with FFRmyo < 0.75 had a D% lower than 50; therefore, the PTCA of intermediate stenoses without quantification must be avoided. CFR is not helpful for a decision to PTCA in such cases, because a normal value of CFR is relevant only. PMID- 10868006 TI - [Clinical experiences with the PercuSurge GuardWire--a new system for prevention of peripheral embolisms in catheter interventions on degenerated coronary artery venous bypasses]. AB - Peripheral embolization is a typical complication of catheter interventions in degenerated aortocoronary saphenous vein grafts. Alternative techniques, such as transluminal extraction catheter (TEC), directional coronary athrectomy (DCA) and eximer laser angioplasty (ELCA) were not able to reduce the risk of peripheral vessel occlusion or "no-reflow". For the protection from peripheral embolisation we used the PercuSurge GuardWire Temporary Occlusion & Aspiration System (PercuSurge Inc., Sunnyvale, USA) in 55 patients with 58 stenosis in degenerated saphenous vein grafts. The primary endpoint of this consecutive series was to access the efficacy of the system by post-interventional TIMI flow and determination of serial CK/CK-MB. The intervention was primarily successful in 55 of 58 cases (94.8%), with a reduction in stenosis from 87.6% (range 75-100%) to 4.5% (range 0-40%). TIMI flow improved from a mean of 2.1 +/- 1.4 to 2.9 +/- 0.4 after intervention. CK/CK-MB levels were 24/5, 27/7 and 26/6 U/l (mean values) before, 8 hours, 16 hours and 24 hours after intervention, ruling out myocardial ischemia. Three (5.2%) non-q wave infarctions occurred in this series, twice caused by a peripheral embolization during the crossing of the lesion with the wire und once because of "no-reflow". In the latter case the occlusion balloon had to be deflated before the aspiration could be performed. In 80 of 92 aspirates (86.9%) macroscopic embolic debris was visible. The PercuSurge GuardWire Temporary Occlusion & Aspiration System is a safe and effective device for the protection of distal embolization during interventions in degenerated aorto-coronary saphenous vein grafts. PMID- 10868007 TI - [Safety and feasibility of intracoronary brachytherapy with the Novoste system within the scope of international multicenter studies]. AB - Clinical trials are increasingly investigating the effects of intracoronary radiation for the treatment of de-novo lesions, restenosis (without stents), and in-stent restenosis. As the first group in Germany, we had the opportunity to use the Novoste system within the international multicenter studies BETA-CATH, START and BRIE and report our preliminary experience regarding safety and feasibility of intracoronary brachytherapy with this afterloader. A total of 95 patients were enrolled. The Novoste system was used in 92 patients (104 lesions). Ischemic complications were not observed; therefore, radiation was performed as planned. The mean applied dose was 16 +/- 2 Gy (14-20 Gy, at 2 mm distance) and mean exposure time was 202 +/- 27 s (165-261 s). The addition of brachytherapy increased the total duration of the intervention for 17 +/- 8 min. At the body surface of the patients, the following dose rates were measured: left chest wall: 99 +/- 52 microSv/h; groin 3 +/- 3 microSv/h. All patients received ASS 300 mg/d o.d. Patients with stent implantation in the same session received 250 mg b.i.d. Ticlopidin or 75 mg Clopidogrel o.d. for at least three months. Total mortality and infarct rate was 0. There was no acute, subacute or late stent thrombosis. CONCLUSION: Our first experience with the Novoste Beta-Cath system showed that intracoronary brachytherapy can be safely and simply performed in the cath lab. There were no acute complications. To avoid the possible risk of late stent thrombosis, Ticlopidin or Clopidogrel must be administered for at least three months. PMID- 10868008 TI - [Intracoronary stent implantation and left ventricular function in primary PTCA in acute myocardial infarct]. AB - In acute myocardial infarction intracoronary stenting is superior to PTCA regarding interventional success and occurrence of cardiac events. It is, however, uncertain whether myocardial function also improves with stenting. We, therefore, assessed angiographic parameters of myocardial function in patients with acute myocardial infarction who were treated with primary PTCA and received additional stenting in case of an unsatisfactory angiographic result (provisional stenting). Nineteen patients with acute myocardial infarction, in whom a "stent like" angiographic result was achieved by PTCA alone, were compared with an equal number of patients receiving provisional stenting. The groups were exactly matched with respect to severity of coronary heart disease, segment of coronary occlusion, Killip class, and TIMI flow after intervention. We only included patients without inhospital cardiac events, in whom repeat angiography after ten days revealed a patent target vessel. There were no differences between both groups regarding age, gender, enzymatic infarction size, duration of ischemia (< or = 12 h), and cardiac risk factors. Myocardial function was assessed by ventriculography and was analyzed quantitatively by the center-line method. The group treated by intracoronary stenting showed a significantly improved ejection fraction (60.3 +/- 2.1% vs. 52.6 +/- 2.9%). All parameters of regional wall motion also indicated significantly less functional disturbance in the stented group compared to PTCA alone (circumferential extend of hypokinesia: 7.4 +/- 2.4% vs. 16.1 +/- 3.4% chords, maximum hypokinesia in the central infarct region: 0.98 +/- 0.20 vs. -1.52 +/- 0.15 SD, severity of regional hypokinesia: 7.3 +/- 2.6 vs. 21.9 +/- 5.4 area). In summary, these results in patients undergoing primary PTCA in acute myocardial infarction indicate that intracoronary stenting is superior to PTCA alone with respect to myocardial recovery, even if an angiographically "stent-like" result can be achieved. Probably, stenting results in a more efficient reperfusion. PMID- 10868009 TI - [New developments in diagnosis of coronary heart disease--3D fusion image]. AB - The interpretation of three-dimensional (3D) structures of the coronary tree and the myocardium by a clinician demands a subjective visual integration of two dimensional (2D) images of cardiac diagnostic procedures like coronary angiography and myocardial scintigraphy. Although in the conventional analysis of 2D display scintigraphic myocardial perfusion segments are arbitrarily assigned to three major coronary artery systems, the standard myocardial perfusion distribution territories correspond with the individual pathologic-anatomic coronary tree in only 50-60% of the patients. Hence, the mental integration of both 2D images of coronary angiography and myocardial scintigraphy does not necessarily allow an accurate assignment of particular myocardial perfusion regions to the corresponding vessels. For an objective assignment of each vessel segment of the coronary tree to the corresponding myocardial regions, we have developed a 3D "fusion image" technique and applied it to patients with coronary artery disease. Cause-and-effect relationships may be more obvious with 3D data fusion and may enable an easier comparison of anatomy and physiology. Preliminary results demonstrate that our newly developed 3D fusion image is useful for accurate assignment of coronary vessel segments to the corresponding myocardial perfusion regions and suggest that it may allow the clinician a comprehensive and accurate assessment of the patient's myocardial status. PMID- 10868010 TI - [Mitral valve prolapse]. AB - Mitral valve prolapse has previously been found to be associated with severe cardiovascular complications such as embolic insults, infectious endocarditis, and sudden cardiac death. However, at the same time, in particular after adopting M-mode and 2D echo for diagnosis, prevalence of the disease was found to be very high, especially in the young. The dilemma of a disease which is frequent and mostly asymptomatic, but in some cases has catastrophic complications, has been solved by implementation of more restrictive diagnostic criteria based on an appreciation of the spatial morphology of the mitral annulus. These criteria call for diagnosis exclusively based on long axis views and a prolapse of > 2 mm beyond a line connecting the leaflet insertion points. "Classic prolapse" additionally requires diastolic thickness of the mitral leaflets of at least 5 mm. Two recent studies, a population-based study of mitral valve prolapse prevalence, and a case-control study of juvenile stroke patients compared to a group of young patients without a history of stroke, shed further light on this disease. The authors found that prevalence of mitral valve prolapse in an average population is 2-3% (1.3% for classic prolapse), without age or sex preponderance; the rate of cerebrovascular insults, congestive heart failure, and atrial fibrillation of patients with prolapse does not exceed that of the rest of the population; however, mitral insufficiency is more frequent; young patients with a history of cerebrovascular insult do not have higher mitral valve prolapse rates than young patients without previous insult. PMID- 10868011 TI - ["Cherchez la femme"]. PMID- 10868012 TI - [Obituary for a friend. Dr. med. Karl-Ludwig Neuhaus]. PMID- 10868013 TI - [Corticoids in therapy of rheumatic diseases yesterday--today--tomorrow]. AB - The discovery of the suprarenal glands and their function is discussed. The complicated history of cortisone from the synthesis until its use as an antirheumatic is told. Pharmacotherapy with this physiologic hormone also caused serious side effects. Changing the molecular structure did not essentially solve this problem. Only after finding novel ways of application could these risks be reduced. The indications for therapy were further effectively reduced after understanding the mechanisms of action. Modern standards for corticoid therapy of rheumatoid arthritis are discussed. The important problem of corticoid osteoporosis and the myth of cortisone ulcers are also explained. The problems of coping with patient fears to cortisone are considered. Concurring drugs of cortisone and their special benefits are compared. PMID- 10868014 TI - [A high rheumatoid factor titer caused by vaccination?]. AB - A forty year old healthy adult man was admitted to our outpatient department because of a highly elevated rheumatoid factor. Since the person was planning an extended stay in Africa, we performed an extensive history, clinical and laboratory examination to rule out oligosymptomatic inflammatory diseases. We further analyzed if the person presented genetic risk factors to develop rheumatic diseases in the future or if the presence of the rheumatoid factor could be a reaction to multiple immunizations. Possible explanations for an elevated rheumatoid factor including the genetic risk factors are discussed. PMID- 10868015 TI - [Individual isokinetic strength training in patients with gonarthrosis]. AB - PROBLEM: To date, therapy of osteoarthritis of the knee is aimed at relieving pain and changing behavior patterns, which usually leads to reduced activity. The weakening of the quadricep's musculature leads to an increase in both joint instability and arthritis. Walking time is prolonged and the pain-induced reaction of knee angle velocity is onset by increased stress on other joints. The progressive muscle atrophy correlates to the degree of pain. The aim of this study was to demonstrate an improvement in strength and pain based on 4-week isokinetic strength training in gonarthritis patients. METHOD: During a conservative hospitalization period, isokinetic strength training was performed by 19 randomized patients with gonarthritis in addition to regular physiotherapy. Another 19 patients functioned as a control group. The work was examined at 60 degrees/s and 180 degrees/s and rated using a pain questionnaire at the start and end of the investigation. RESULTS: In addition to the expected increase of strength and strength endurance in the test group, the degree of pain could also be statistically significantly decreased compared to the control group. Activities of daily living, such as climbing stairs and standing-up, were also performed more easily. CONCLUSIONS: The therapeutic strategy for patients with osteoarthritis of the knee should be reconsidered to include less expensive therapeutic sport measures. Anglo-american and Scandinavian studies support this statement. Overuse and pain can be avoided by precise and low-dose strength training. Objective and reproducible measurements in the patients are essential to make individual training possible. PMID- 10868016 TI - [Long-term outcome of metatarsal head resection in rheumatoid arthritis]. AB - Between January 1983 and December 1987, metatarsal head-resections were performed on 203 patients, comprising a total of 370 feet, using the Hueter/Mayo and Hoffmann procedure. Seventy-two patients, comprising a total of 126 feet, were available for post-operative review after an average of 11.4 years from the date of the original operations. The information obtained from standardized questionnaires was compared to the information found in each patient's file. In addition, every available pre- and post-operative x-ray taken from 1983 to 1987 was analyzed. Thus, with an average follow-up period of 5.6 years, the changes found in the pre- and post-operative x-rays from a total of 183 feet could be compared. Before the operations, nearly 100% of the examined feet suffered from painful synovial hypertrophy and erosion of the metatarsophalangeal joints with dislocation and subluxation, causing approximately 70% of all patients to have great difficulties in walking. After the operations, however, 90.2% of the patients reported that this condition had noticeably improved or had completely disappeared. In fact, 87.5% of all patients reported a lasting improvement in their ability to walk longer distances. As the main criteria in determining the success of an operation (namely, the noticeable reduction of pain and increased mobility) were achieved in 87.5% of the patients, we consider the metatarsal head resection a reliable method of correcting forefoot deformities in rheumatoid arthritis. PMID- 10868017 TI - [Mucocutaneous candidosis in a patient with Sjogren syndrome]. AB - Oral candidiasis is common among patients with Sjogren's syndrome. For the first time we report on a 60-year-old female patient, who developed a non-familiar chronic mucocutaneous candidiasis of later life. She presented with the following symptoms: oral candidiasis and glossitis, angulus infectious, vulvovaginitis, lichen planus-like onychodystrophy of the finger-nails and chronic nail bed inflammation of the nails of the big toes. The knowledge of chronic mucocutaneous candidiasis is of prognostic and therapeutic relevance, since topical treatment often fails. PMID- 10868018 TI - [Prospective multicenter observational study of early rheumatoid arthritis- prognostic factors and predictors of disease course]. AB - The aim of a new prospective, multicenter trial on early rheumatoid arthritis is to evaluate parameters of potential relevance for the prediction of an erosive disease course. The integration of general practitioners, rheumatologists, arthritis care units and experimental research groups in a network structure is intended to improve the cooperation between basic and clinical research as well as the treatment of the patients. The final goal is to identify new parameters that may improve an early stratification of patients into the appropriate treatment areas. PMID- 10868019 TI - [Inception cohorts for spondyloarthropathies]. AB - The spondyloarthropathies (SpA) are important inflammatory rheumatic diseases- not only because patients of young age are frequently involved. Ankylosing spondylitis, reactive arthritis, and undifferentiated spondyloarthropathy are prototypes of SpA which share several clinical features and the strong association with HLA B27. Despite its high prevalence there are many shortcomings in early diagnosis and therapy of SpA. The same is true for the knowledge on course, prognostic factors, and the socioeconomic consequences of these diseases. Furthermore, our knowledge about pathogenesis and therapy is still limited. Due to the known deficiencies in rheumatologic education, there are clear-cut deficiencies in the health care provided by general practitioners. The aim of the project presented here is to built up a cohort of early SpA patients to learn about course and outcome of SpA. In this regard, special attention is paid to the inclusion of patients with juvenile SpA since AS starts frequently in the 2nd decade of life and because the clinical picture with more frequent peripheral joint involvement differs from AS of adult onset. Moreover, these cohorts play an essential role for other projects of the MedNet subdivision of infect-associated arthritides/spondyloarthropathies in which studies on diagnosis, therapy, immunology, and genetics of the SpA performed. PMID- 10868020 TI - [Prospective study of JCR-Oligo I of the Child and Adolescence Rheumatology Working Group]. AB - Intra-articular injection of corticosteroids is a powerful anti-inflammatory treatment with relatively few side effects. To date no prospective RCT exists on its use in children with oligoarthritis type I. Thus, in January 2000 the Arbeitsgemeinschaft started a multicenter RCT to compare in children with this diagnosis the initial treatment recommended in the textbooks, which is NSAID plus physiotherapy, with the early use of intra-articular injection of Triamcinolonhexacetonide. In the synovial fluid the presence of cytokines and the specific reactivity of T cells will be explored, and a highly sensitive screening for infectious agents will be performed. PMID- 10868022 TI - [Clinical day care treatment of patients with rheumatoid arthritis: concept of a new outcome-oriented evaluation project]. AB - Patients with rheumatic diseases are commonly treated as an outpatient in their local environment or are referred to specialized centers. Two recently founded day-patient clinics in Berlin and Frankfurt/Rhein-Main (Germany) for patients suffering from rheumatic diseases will be evaluated for their medical outcome and cost-effectiveness of comprehensive treatment of RA patients in stages of high inflammatory activity and progressive disability. The study design will be prospective, controlled, and randomized to compare outpatient and day-patient treatment. The case-control study design with matched pairs within the network of collaborative arthritis centers will be used to compare day-patient and inpatient treatment. The paper contains a review of studies published in English or German language dealing with day-patient treatment of RA patients. PMID- 10868021 TI - [Cyclooxygenase-2 (Cox-2) selective inhibitors: socioeconomic and pharmaco epidemiologic aspects]. AB - Health authorities of several European countries recently introduced guidelines for socioeconomic evaluations. Additionally the activities of OMERACT (Outcome Measures in Rheumatoid Arthritis Clinical Trials) indicate an increasing awareness for the need of socioeconomic studies in rheumatology. The planned 2000 OMERACT meeting in Toulouse will address transfer of socioeconomic standards into rheumatology. In terms of cost effectiveness of selective Cox-2 inhibitors, a reference has to be made to the preceding discussion of cost effectiveness of Misoprostol. In addition, there are two models examining the cost effectiveness of Cox-2 inhibitors: a Canadian model comparing Nabumetone and Naproxen and an unpublished model assessing the cost effectiveness of Celecoxib (ACCES: Arthritis Cost Consequence Evaluation System). German data indicate that gastrointestinal bleedings account for 32.9% of all adverse drug events leading to a hospital admission. Further data assessing the morbidity due to adverse effects of nonsteroidal anti-inflammatory drugs are needed. Such data would allow the quantification of possible savings related to the usage of Cox-2 inhibitors in Germany. PMID- 10868023 TI - [Long-term prognosis and outcome of patients with primary systemic vasculitis: initial results in Germany]. AB - Some outcome parameters have been shown to represent valuable measures for the long-term assessment of primary systemic vasculitides (PSV). These parameters include disease extent, disease activity, damage, and the health-related quality of life. We describe a concept for the establishment of a detailed documentation system for PSV with a duration of at least 5 years to obtain a reliable characterization of these patients. This database will provide the basis for a PSV cohort and the development of an interdisciplinary patient education concept followed by an evaluation with a multicenter, trial. PMID- 10868024 TI - [Quality assurance--small group work. Diagnosis/Therapy Group]. PMID- 10868025 TI - [Surgical diagnosis: a good teaching tool]. PMID- 10868026 TI - [Current treatment of malignant epithelial tumors of the ovary]. AB - Surgery is the essential element of staging and treatment of malignant ovarian tumours. Regardless of the stage, it must include peritoneal cytology, hysterectomy with bilateral adnexectomy, omentectomy, pelvic and lumbo-aortic lymphadenectomy, appendicectomy and multiple peritoneal biopsies. In stage I tumours, in young women desiring a subsequent pregnancy, preservation of the uterus and contralateral ovary can be proposed. In stages II, III and IV, the therapeutic strategy consists of primary surgery and systematic chemotherapy (6 cycles). Radical surgery is essential in these cases, as the size of the residual tumour at the end of operation constitutes the major prognostic factor. To optimize the quality of tumour debulking, the maximum of visible carcinomatous nodules must be resected with, if necessary, gastrointestinal resections. The value of second-look surgery, after 6 cycles of chemotherapy, is currently controversial: it is only indicated in the context of randomized trials. Borderline malignant ovarian tumours have a good prognosis regardless of their stage. Surgery can very often be conservative, particularly in young women. Adjuvant chemotherapy has been shown to be effective in these tumours. Many studies are underway to define the value of new cytostatic molecules and "interval" surgery (intercalated between several courses of chemotherapy). PMID- 10868027 TI - [Initial surgery in advanced epithelial cancers of the ovary]. AB - Epithelial ovarian cancer is usually diagnosed at an advanced stage with a bulky tumor in the pelvis and upper abdomen. The most common therapeutic strategy begins by a surgical operation that allows histologic diagnosis, accurate staging and maximal debulking. Since the papers by Griffiths at the end of the seventies, the volume of the residual tumor after surgery appears to be one of the most important prognostic factors in all series. Indeed, patients whose tumor is completely or optimally debulked have greater chances of prolonged survival of about 50% at 5 years. Surgeons experienced in this field can achieve optimal debulking in about 75 to 80% of cases. But, in order to reach this objective, they must often perform an ultra-radical operation with extensive peritonectomies, lymphadenectomies and intestinal resections. Moreover, since 1983, Hacker has shown that the initial tumor bulk was still a poor prognostic factor even after debulking. Today it can be demonstrated that the greater the tumor bulk the more aggressive must be the surgical procedure in order to be optimal and the final benefit will nevertheless be proportionally lower with a higher morbidity rate. This paradigm leads the surgeons to currently try to more accurately assess the initial tumor bulk in order to determine wether the tumor would be optimally debulked by means of a well-standardised operation. If not, the alternative strategy would be 3 chemotherapy courses as front-line treatment before debulking surgery, which hopefully would be easier. Trials are needed in order to validate this strategy despite the fact that some patients will unfortunately have their prognosis jeopardized by the chemoresistance of their tumor. PMID- 10868028 TI - [Early debulking surgery after chemotherapy in advanced cancer of the ovary]. AB - The main prognostic factor in advanced ovarian cancer is the volume of residual disease after the initial laparotomy. Early debulking surgery after several cycles of chemotherapy, before the emergence of resistant cell lines, could improve the prognosis of patients with bulky residual disease. This study concerns patients with advanced ovarian cancer entered into three prospective trials including IV cisplatin and anthracycline-based chemotherapy, early debulking surgery after three cycles of chemotherapy in case of initial residual disease superior 2 cm and intraperitoneal consolidation chemotherapy. Among 160 patients with stage III or IV, 80 (50%) had at least a residual tumor of more than 2 cm in diameter. Early debulking surgery was effectively performed in 54 patients (67.5%), leaving 39 patients with no residue over 2 cm. Twenty-one patients had no macroscopic residual disease. The median survival of all patients with initial residual disease over 2 cm was 23 months. Patients with no macroscopic residual disease at early debulking surgery had a median survival of 44 months. Early debulking surgery appears useful in advanced ovarian cancer with bulky residual disease. The objective of this operation is to achieve no macroscopic residual lesion. PMID- 10868029 TI - [Endosonography in the preoperative evaluation of cancers of the esophagus]. AB - OBJECTIVE: To assess the diagnostic accuracy of endoscopic ultrasonography (EUS) for the local and regional staging of esophageal cancer, and its possible alteration resulting from the performance of preoperative chemoradiation. METHODS: Prospective study of 85 consecutive patients with esophageal cancer evaluated by EUS and operated on between January 1992 and December 1995. 28 of these patients had received previous induction therapy. In all cases, EUS examination was performed by the same physician not informed about the results of previous morphological explorations. Histopathological analysis of all operative specimens was performed by the same pathologist, not informed about the results of EUS. Data were collected by another independent observer. RESULTS: EUS examination resulted in incomplete staging in 8 patients (9.5%) with severe stenosis precluding endoscope passage. The accuracy, specificity and sensitivity of EUS in detecting the depth of esophageal involvement (T0-2 vs. T3-4) were 82.3%, 78%, and 86% respectively, and 72%, 70%, and 73% respectively for lymph node metastasis. The overall accuracy of EUS in identifying the preoperative stage was 67%, with a clear-cut alteration when patients had received induction therapy (61% vs 72%). On the other hand, 7 (64%) of the 11 patients thought to have a complete response at endosonography had no residual tumor. CONCLUSION: EUS provides precise information for the preoperative identification of locally advanced esophageal tumor, even after induction therapy. The latter alters the diagnostic accuracy of EUS, although complete responders could be identified in two-thirds of cases. PMID- 10868030 TI - [Laparoscopic fundoplication for gastroesophageal reflux. Apropos of our first 123 patients]. AB - From July 1991 to March 1997, 123 patients underwent laparoscopic fundoplication. Surgical indications were as follows: either failure of medical therapy, or early recurrence of symptoms after interruption of medical treatment in young patients or large hiatal hernia associated with symptoms of reflux and/or symptoms of mediastinal compression. The type of the wrap was tailored to the preoperative manometry: circumferential fundoplication was achieved in patients with normal esophageal motility, and partial wrap in patients with altered motility. Short gastric vessels were not routinely divided. One hundred and eleven circumferential fundoplications were performed: 52 with division of short gastric vessels and 49 without, whereas there were 22 partial wraps. In 4 cases, it was necessary to switch to open surgery (conversion rate: 3.2%): 2 enlarged left liver lobes, one esophageal tear and one splenic injury. Six postoperative complications were observed (morbidity rate: 4.8%), one of whom was severe and led to the patient's death due to necrosis of the fundus. After a mean follow-up of 1.7 +/- 1.4 years, 4 patients have transient recurrent reflux, 3 patients have had annoying dysphagia requiring balloon dilatation in one case and reoperation in two cases. Four patients experienced a late thoracic migration: in one case after a violent physical effort, requiring urgent reoperation; in the other three cases, the migration remained asymptomatic. The pH- and manometric study performed in 41 consecutive patients before and after surgery allows objective evaluation of the results. PMID- 10868031 TI - [Laparoscopic treatment of small bowel obstruction arising on adhesions]. AB - OBJECTIVES: Determine the feasibility of laparoscopy, with the objective as well as the subjective benefits offered to patients, and the possible contra indications of this type of surgery in the treatment of mechanical intestinal ileus. PATIENTS AND MATERIAL: This concerned 20 patients who came to the hospital with a picture of intestinal obstruction. All had a history of at least one abdominal operation. The diagnosis of mechanical occlusion due to band adhesions was presumed on the basis of the clinical and complementary examinations, particularly x-rays, and confirmed in the course of the operation. All patients were operated by laparoscopy after failure of non-invasive treatment. RESULTS: Results obtained are encouraging since 60% of the patients were successfully treated by laparoscopy alone. The patients' comfort and length of hospital stay are improved by laparoscopy. There is no relation between previous surgical history and the results of the technique. CONCLUSIONS: Most failures were due to the time interval between onset of symptoms and the operation. Early operation therefore seems to increase the chances of success. PMID- 10868032 TI - [Splenectomy for splenomegaly of more than 1000 grams. A retrospective study of 36 patients]. AB - Splenectomy for massive splenomegaly is frequently performed for hematologic disorders for diagnostic and therapeutic indications. The role of splenectomy is complex and controversial. The aims of our retrospective study were to focus on postoperative complications and advantages of splenectomy for massive splenomegaly. Thirty six patients with splenomegaly weighing 1000 g or more, underwent splenectomy at Centre Hospitalier Universitaire Lyon Sud, from January 1st, 1982, to December 31, 1995. Thirty-one (85%) of these patients had hematologic malignancy and more than half of them were older than sixty years. The main indications for splenectomy were hypersplenism (18 patients) and diagnosis (14). Preliminary ligation of the splenic artery was performed in 25 patients (42%). All patients had drainage. The mortality and morbidity rates were 5.5% and 20%, respectively. No major septic or thromboembolic complications occurred. There was only one major bleeding complication. The advantages of splenectomy included histopathological diagnosis in 13 of 14 patients with splenomegaly of unknown origin, permanent pain relief in all cases, and immediate correction of hematological cytopenia in 27 cases (75%). We conclude that the large weight of the spleen does not constitute a contraindication to splenectomy, but indications must be carefully selected, and the operative and perioperative management, must be appropriate. PMID- 10868033 TI - [Diagnosis and treatment of rectal and sigmoid endometriosis]. AB - From October 1989 to September 1994 six resections of the bowel were performed for colorectal endometriosis. Five of, the patients, with a mean age of 32 years, presented clinical features. In all cases, colonoscopy showed a normal mucosa. All patients treated by hormonetherapy relapsed. The resection was segmental with immediate end-to-end anastomosis in 5 cases and partial in 1 case. In three cases, endometriosis of the genital tract was associated and treated during the initial laparotomy. One low rectosigmoid anastomosis fistulised. Rectosigmoid endometriosis accounts for 70% of bowel localisations and genital endometriosis is associated in 80% of cases. Deep and clinical rectosigmoid endometriosis does not respond to hormonetherapy and requires bowel resection. The pelvis should be explored and genital tract endometriosis treated. Postoperative hormonetherapy should be considered after initial surgery. PMID- 10868034 TI - [MR cholangiopancreatography]. AB - MR cholangiopancreatography (MRCP) is a new technique allowing noninvasive investigation of the bile ducts and pancreatic duct. Due to the extremely intense signal of water on T2-weighting, MR sequences can be obtained only demonstrating liquids. The bile and pancreatic ducts can therefore be studied spontaneously even in the case of major cholestasis. The contraindications of MR cholangiography are exclusively those of MR. MRCP can visualize the level of a bile duct obstruction and often the nature of this obstruction (stone, tumour). Complementary axial T1- and T2-weighted sequences can also visualize the parenchyma around the ducts. MR cholangiography therefore appears to be a technique of the future for noninvasive investigation of the bile ducts. PMID- 10868035 TI - [Isolated cerebral metastases disclosing rectal adenocarcinoma]. AB - Colorectal cancer is usually revealed by modifications of bowel habit and/or signs of haemorrhage. Hepatic and lung metastases are the common sites of metastatic involvement of this cancer. Brain metastasis are rare, especially when they are isolated. We report the case of a 37-year-old patient presenting with isolated brain metastasis revealing a rectal cancer. The patient was initially treated by surgical excision of the symptomatic brain metastasis, followed several days later by anterior resection of the rectum and whole-brain radiotherapy. This unusual strategy, due to the lack of preoperative diagnosis, did not improve the poor prognosis of brain metastasis. PMID- 10868036 TI - [Laparoscopy and viral risk]. PMID- 10868037 TI - [Anastomoses and sutures in cervical laryngotracheal and pharyngoesophageal surgery]. AB - Head and neck anastomosis techniques present common aspects with any type of anastomosis but their localisation at the junction of the respiratory and digestive tracts makes them often delicate to perform. In the present article, we first review the different surgical indications and the laryngeal, tracheal, pharyngeal and oesophageal anastomosis techniques. The main types of flaps also used for this purpose are highlighted. We then review the cases observed during the 5 last years in our department, type and technique of surgery used. In this general review, we illustrate our stand point although aware of the multiple variants favoured by different schools. PMID- 10868038 TI - [Epidemiology of breast cancers in the Somme department (1990-1993)]. AB - Breast cancer is the commonest female cancer in the Somme region and represents the leading cause of cancer mortality in women. It therefore constitutes an important public health problem. From 1990 to 1993, 1,106 new cases of breast cancer were recorded by the Somme Cancer Registry. The incidence continued to increase over this period in the Somme region. The mortality rate for this period was 35.1/100,000 women, while the standardized mortality rate for the world population was 20.9/100,000 women. The most frequent histological types were invasive ductal carcinoma (64.3%) and invasive lobular carcinoma (5%). Carcinoma in situ represented 2.9% cases; 4% of patients presented metastases at the time of diagnosis. For the period 1990-1993, 44.3% cases were classified as T1, 37.9% as T2 and 11.5% as T3-T4. The lymph node extension rate was less than 15% for tumours less than 10 mm (on the resection specimen). The 5-year survival rate was 73%. It is important to increase the rate of early diagnosis in order to improve the overall survival of this disease. PMID- 10868039 TI - [Preoperative colonic lavage and one-stage excision-anastomosis in obstruction of the left colon]. AB - In a retrospective series of 95 patients requiring emergency surgery for distal colonic obstruction, primary bowel resection followed by immediate anastomosis after intraoperative colonic irrigation was performed. Carcinoma was the cause of obstruction in 81 cases (85%); 13 patients had diverticulitis, and 1 had sigmoid volvulus. The technique of on-table lavage was similar to that described by Dudley in 1980: a caecostomy tube was used in 86 patients (90%) and was removed on the tenth postoperative day. 4 patients died, none from complications of anastomotic leakage. There were three anastomotic leakages (3.1%) and 10 radiologic leaks were observed. 3 patients were reoperated. The mean hospital stay was 23 days. The results of this study suggest that intraoperative colonic irrigation is an effective method, enabling the surgeon to perform primary anastomosis with reasonable safety after emergency resection of selected distal colonic lesions. PMID- 10868040 TI - [Laparoscopy-assisted colonic polypectomy or how to be helped by laparoscopy to prevent colectomy in benign colonic polyps considered to be unresectable by colonoscopy]. AB - The aim of this study was to prospectively evaluate the role of laparoscopic surgery in all patients presenting with colonic polyps. From April 1994 to April 1996, 16 consecutive patients were treated. The laparoscopy starts the procedure, then a colonoscopy easily and rapidly locates the colonic lesion. Under laparoscopic supervision a new snare polypectomy is often possible. If polypectomy remains impossible, a mini-laparotomy is performed above the polyp and allows polypectomy by extra-peritoneal colotomy. In one case, the polyp was not found on colonoscopy. Colonoscopic polypectomy was possible in 6 cases (40%), and 9 mini-laparotomies were necessary (60%). No colectomy was performed. The final histologic result showed two Dukes A carcinomas justifying secondary wide colectomy. Laparoscopy-assisted polypectomy is a safe and efficient procedure, allows complete excision of polyps and may avoid a colonic resection. PMID- 10868041 TI - [Complicated appendicitis in children: laparoscopy or Mac Burney incision?]. AB - PURPOSE: The aim of this retrospective study was to compare open (Mc Burney incision group MB n = 92) and laparoscopic (group LAP n = 58) appendectomy for complicated appendicitis (abscess or peritonitis). MATERIAL AND METHOD: 150 children, with a mean age of 8.5 years, were operated in our department from January 1990 to April 1996. Only children with complicated appendicitis and positive bacteriology of peritoneal fluid were included in this study. All children received parenteral antibiotics for an average of 5.4 days. RESULTS: The mean operating time was significantly longer in the LAP group (63 vs 43 min p < 0.0001). The conversion rate in the LAP group was 12%. The mean hospital stay was 8.4 days (3-29) without any difference between the 2 groups. There was no significant difference for the complication rate in the 2 groups, except for wound infections which were more frequent in the MB group (p = 0.008). Late postoperative complications occurred in 3 cases in the MB group (none in the LAP group) (NS). There were 2 small bowel obstructions and a wound dehiscence. CONCLUSION: Laparoscopic appendectomy is a safe procedure for complicated appendicitis in children, but the greatest short-term benefit is cosmetic. Long term results have to be evaluated, particularly with regards to the long-term complication rate. PMID- 10868042 TI - [Cystic pneumatosis of the colon. Report of 3 cases and review of the literature]. AB - Pneumatosis cystoides intestinalis (PCI) is a rare entity in which gas is found in cysts located in the intestinal wall. PCI is a sign, not a disease; therefore, its relevance should be interpreted within the whole clinical context. PCI has been found in several distinctive clinical settings, which the authors review in the light of their 3 new cases. Morphologic and endoscopic examinations showed "polypoid" intraluminal formations corresponding to pneumocysts which should not be misdiagnosed with a diffuse polyposis. The most important tasks of the physician include: 1) recognition of the entity of PCI so that patients are not misdiagnosed and mismanaged as having malignancy or polyposis; and 2) differentiation of the benign noncomplicated cases from life-threatening forms (bowel necrosis, perforation, and infections), in which immediate surgery is necessary, and which are associated with high mortality. PMID- 10868043 TI - [Cancer of the upper third of the bile duct]. AB - Carcinoma of the upper third of the extrahepatic bile ducts (Klatskin's tumors) are difficult to manage and there is still no consensus on the best means of diagnosis and therapy. The objective of this general review was to determine the state of the art about the pathology and the available diagnostic and therapeutic options. PMID- 10868044 TI - [Treatment of acute postoperative pain]. AB - Postoperative pain is a subjective experience involving sensations and perceptions, which may be the result of tissue damage after surgery. Various analgesic drugs and techniques can be used to relief postoperative pain. They must be adapted to the surgery and to the patient. Moreover, adequate management of postoperative pain need to be organized. This include medical attitudes, clinical orientations, disciplinary involvements, consultative protocols and program education. PMID- 10868046 TI - [Ileocaecal tuberculosis. Apropos of a case]. AB - The authors report a case of ileocaecal tuberculosis in a 27-year-old man with no particular risk factor for this disease. The initial diagnosis was terminal ileitis discovered at appendicectomy. The diagnosis of ileal tuberculosis was suspected in the presence of giant cell follicles on ileal biopsies, and was confirmed by the presence of AFB in the gastric intubation fluid. This patient presented known atypical pulmonary images for several years, which had never been investigated in more detail. A favourable course was observed in response to triple-agent, then double-agent antibiotic therapy. This case illustrates the fact that ileal tuberculosis still exists today, and that it does not exclusively affect "high-risk" patients. PMID- 10868045 TI - [Retroperitoneal malignant fibrous histiocytoma]. AB - Malignant fibrous histiocytoma is the commonest soft tissue sarcoma of adults. The soft tissue of the extremities is the commonest primary site of malignant fibrous histiocytoma. It is much less common in the female retroperitoneum, leading to diagnostic errors. The clinical, radiographic and CT signs are non specific. This tumor can only be diagnosed by histology. An initial complete resection is essential for successful treatment of the primary tumor. Radiation therapy is limited and chemotherapy has only been successful in a limited number of cases. This tumor has a poor prognosis. These lesions are relatively rare and consequently difficult to study. The authors report three cases and review the literature. PMID- 10868047 TI - [Pneumatosis cystoides intestinalis. Diagnostic elements and therapeutic approach]. AB - Pneumatosis cystoides intestinalis is an uncommon condition characterised by multiple gas-filled cysts within the small intestine or colonic wall. Clinical manifestations are unspecific and often found in many other abdominal diseases. To avoid unnecessary laparotomy, radiologic and endoscopic findings are essential to be known. The present case associates symptoms highly suspect of neoplasia like weight loss, rectal mass, bloody stools and tenesmus. Treatment of choice is medical. In the absence of an acute abdomen, surgery is only reserved when it is not responsive to medical treatment. PMID- 10868049 TI - [Two-stage laparoscopic treatment of perforated sigmoid diverticulitis: a case]. PMID- 10868048 TI - [Inflammatory pseudotumor of the liver. Apropos of a case]. AB - The authors report a case of inflammatory pseudotumor of the liver, a rare lesion of unknown pathogenesis. Conservative management could be justified due to the good prognosis of this disease, when it is diagnosed preoperatively. However, when in doubt, surgical resection is usually recommended to obtain histological diagnosis. PMID- 10868050 TI - [Thermodynamic theory of biological evolution and aging. Experimental verification of the theory]. AB - Experimental data confirming original thermodynamic theory of biological evolution and aging are presented. Biological evolution (phylogenesis) and ontogenesis can be easily described within the frames of equilibrium hierarchical thermodynamics on the basis of temporal hierarchies law and the second principle of thermodynamics. The theory explains many facts and suggests new practical proposals in medical and biological science, particularly, in dietology, gerontology, and geriatrics. Application of the temporal hierarchies model to studying living nature offers horizonless possibilities for its understanding. PMID- 10868051 TI - [Ultrastructure of the neuromuscular junctions in the rat soleus muscle under varying gravity conditions]. AB - Changes in the ultrastructure of neuromuscular junctions have been considered as an index to adaptation of Wistar rats (whose pre- and postnatal ontogenesis proceeded on a centrifuge under constant rotation until the age of two months) to the hypergravity conditions (2G) and, then, to earth gravity (1G): on the 2nd and 15th days after centrifuge stoppage. The dynamic of synaptic vesicles was shown: their number increased at 2G and gradually decreased at 1G. Local damage of muscle fibers, partial separation of the motor axonal terminal from intrafusal fiber, and membrane twisting were noted at the increased gravity-dependent static load (2G). Neuromuscular junctions with signs of remodeling occurred more frequently in the experimental rats than in the control ones. It was proposed that adaptation of rats to 1G gravity after a prolonged sojourn under the hypergravity conditions (2G) was not completed within the studied period. PMID- 10868052 TI - [Expression of glial, neurospecific, and extracellular antigens during retinal regeneration in adult newts]. AB - Spatial localization and time of expression of glial, neurospecific, and extracellular antigens were studied during retina regeneration in adult newts using antibodies against the glial acidic fibrillar protein (GFAP), neurospecific molecules (N-CAM and PSA-N-CAM), and tenascin (Tn). At the early stages of regeneration (5th day after the operation), slightly differentiated cells in the retina growth area, which are not cellular sources of regeneration, were brightly stained by the antibodies. The one-two-layered retina rudiment formed from the pigment epithelium layer a week later was also intensely stained by the antibodies to GFAP. The retina rudiment cells belong to slightly differentiated precursors of the regenerating retina. It was first shown that the antibodies to GFAP were not only markers of the glial cells, but also markers of slightly differentiated precursors of the regenerating retina. Expression of neurospecific antigens was found in depigmented cells of the retina rudiment. It appeared to have been initiated by cell interactions in the regenerate. An expression of tenascin was found in the cells migrating in the eye cavity and contacting with the retina rudiment cells. The role of tenascin in interaction with the retina rudiment is unknown. PMID- 10868053 TI - [Cytotoxic and cytogenetic effects of radiation are modulated by introduction of RNA preparations obtained from various body tissues]. AB - We studied the influence of various gamma-ray doses on the death pattern of Wistar rats and the changes in lethality after introduction of RNA isolated from various tissues in the organism. All studied RNA introduced into the irradiated animals proved to clearly stimulate repair of the cells damaged by radiation and to decrease both cytotoxic and cytogenetic effects of gamma-rays. The most pronounced effect on the cells was specific for RNA isolated from the matching tissue. PMID- 10868054 TI - [Comparative analysis of the monopodial and sympodial models of bulb branching in Galanthus L]. AB - We have examined sympodial and monopodial models of bulb branching in Galanthus. The issue of the position of the reduced prophyll is discussed. We proposed a method of formal interpretation: parts of the plant were positioned on diagrams; several variants of axial schemes were matched to each diagram; the schemes were divided into two classes, monopodial and sympodial ones, and stability of each class was estimated. In order to decide about the model of Galanthus bulb branching, we have examined plants with additional inflorescences and plants with additional leaf series. We have shown that the sympodial model predicts the presence of the reduced prophyll at the base of the innovation bud in all studied cases. Consecutive stages of prophyll reduction (prophyll of the innovation bud) can be followed in Amaryllidaceae in the following sequence: Zephyranthes, a well developed large prophyll with green lamina; Vallota, a developed prophyll with reduced green lamina; Haemanthus, a thin chaffy short-living prophyll. At the end of this sequence is Galanthus with completely reduced prophyll at the innovation bud. PMID- 10868055 TI - [Adaptive types of prolamins--specialized proteins from cereal seeds (Poaceae Banrh.)]. AB - Amino acid composition of prolamins and the whole seed in representatives of 52 cereal genera and 22 tribes, as well as amino acid composition of seed protein fractions in representatives of 9 cereal genera are presented. In terms of proposed directed evolution of the seed proteins and adaptive role of prolamins in evolution and distribution of cereals, generalized data on their content in seed protein complex, electrophoretic, immunochemical, and amino acid composition of the seeds, we develop the notion of prolamins biochemical specialization and their polyphyletic origin during evolution of cereal seed protein complex. Seven adaptive types of prolamins are recognized: Sasa, Molinia, Chloris, Zingeria, Poa, Triticum, and Panicum types. Adaptive types of prolamins were formed in response to an ecological request of the environment where taxon ancestors existed and a taxon appeared and in relation to phylogenetic history of a taxon and its assignment to particular tribe and subfamily. PMID- 10868056 TI - [Effect of salt stress on respiration metabolism in higher plants]. AB - We studied the activity of NADP-dependent isocitrate dehydrogenase, malate dehydrogenase, succinate dehydrogenase, catalase, and peroxidase as well as the rate of 14CO2 release after introduction of labeled substrates for glycolysis and citrate acid cycle within 24 h after salt stress (1% NaCl) in 10-14 days old germinants of wheat (Triticum aestivum L.) and maize (Zea mays L.) as well as thallus of small duckweed (Wolffia arrhiza (L.) Hork ex Wimmer). Oscillations in the enzymes activity with 4-6 h period have been revealed under stress conditions. Activity of glycolysis decreased in wheat and maize and increased in duckweed under the influence of stress stimulus. Six hours after NaCl action decarboxylation of exogenous citrate and succinate was enhanced in all three plants while the rate of exogenous malate decarboxylation was decreased. We conclude that adaptation of higher plans to salinization is accompanied by rearrangements in oxidative metabolism reflected by oscillations in activity of the enzymes involved in oxidative metabolism. PMID- 10868057 TI - [Nitrogen fixation--new fenomenon in rodent feeding]. AB - We have conducted a series of experiments in order to test hypothesis about the possible use of nitrogen fixation by small phytophagous mammals as a way to supplement their nitrogen balance. Two groups of red voles Clethrionomys rutilis were kept on rations with different protein nitrogen content. Three days later, the rate of nitrogen fixation was determined in different regions of the digestive tract. Nitrogen-fixing activity has been detected in the GI tract of all studied voles, specifically in those regions, where symbiocenoses were present; it was particularly high in animals of the experimental group under the conditions of dietary nitrogen limitation. A mixed culture of bacteria capable of nitrogen fixation has been isolated from the digestive tract of red voles. PMID- 10868058 TI - [Dynamics of rodent community and populations in Kalmyk semideserts under decreasing pasture load and increasing humidity]. AB - Today, pastural ecosystems of the Kalmyk Republic demonstrate progressive regrowth dynamics, which is explained by a sharp decrease in grazing load and the onset of a new humidification cycle in the Caspian Lowland. By the mid-1990s, the sparse and poor desert vegetation with a significant proportion of ephemerals, characteristic of these ecosystems in the 1980s, had been substituted by highly productive tall-grass communities typical of steppes. Under such conditions, corresponding changes in the distribution and abundance of rodents could be expected. These parameters were assessed in different types of habitats in Chernye Zemli (the Kalmyk Black Lands) from 1980 to 1983 and from 1993 to 1997. Rodents were captured in live traps distributed over permanent test plots and in crush traps arranged in lines. The results showed that the population of midday gerbils did not change significantly during this period, although their favorite open habitats considerably decreased in area. The abundance and range of tamarisk gerbils noticeably increased, whereas those of little sousliks continued to decrease, contrary to our expectations. Thus, a rapid progressive succession in plant communities did not cause the corresponding change (of similar rate and extent) in the structure of rodent community. The difference between the observed and expected results provides evidence that rodent populations are somewhat "inert" in their response to changes in environmental conditions, and their development is relatively independent of these changes, but is contingent upon the state of populations in the past (i.e., the history of colonies). PMID- 10868059 TI - [Comparative study of anticoagulants obtained from various extracts of Paeonia anomala]. AB - We have established that extracts prepared from various parts of the root of Paeonia anomala (bark and wood) have anticoagulant activity as indicated by the criterion of longer recalcification time of the normal rat blood plasma. This anticoagulant activity is due to the presence of heparin-like fragments in the preparations, which follows from the results of biochemical analysis and results of studies on the functional state of the mast cell population. Preparations from the peony root bark retain their anticoagulant properties in the circulatory system for 1.5 h after intravenous administration, similarly to heparin of the animal origin. PMID- 10868060 TI - [Morphophysiological adaptation to feeding in Aplysia dactylomela (Rang, 1828)]. AB - Hemoglobin content in radular muscle and relative weight of the main viscera were determined in Aplysia dactylomela. We propose that relatively low concentration of tissue hemoglobin as well as low heart and ctenidium indices are due to the mode of the mollusk nutrition. Nutrition by biting algae fragments requires less muscular effort than, for instance, scrapping off overgrowth from substrate and, hence, is accompanied by lower indices. PMID- 10868061 TI - [Characteristics of wintering in ground beetles (Coleoptera, Carabidae) in forest ecosystems of the East European Plain]. AB - Specific features of wintering of the ground beetles in three habitats (litter, soil, and bark of fallen trees and stumps) were comparatively studied in the forests of forest-steppe (Voronezh District) and subzone of broad-leaved-spruce forests (Moscow District). The main mass of ground beetles is concentrated in the upper 10-cm soil layer, irrespective of the type of watering (automorphous or hydromorphous soils). Wintering under the bark is a facultative feature of the most species occurring in these biocoenoses. PMID- 10868062 TI - [Protist plankton from the Khanka lake]. AB - We present data on studies of protists in Lake Khanka in late August-early September. A total of 99 species of protists have been found, of them 14 phytoflagellates, 14 zooflagellates, five sarcodins, two heliozoans, 62 infusoria, and two suctorians. Small infusoria from the class of Kinetophragminophora, order Oligotrichida, as well as suborder Tetrahymenina were most numerous. The density of many protist species in Lake Khanka exceeds markedly that in several other high-productivity water reservoirs. Most of the protists belong to bacteriovores (bacteriophages). Many of the found species have high density. This is evidence of the production of bacterial plankton in the lake. In view of very high water turbidity in the lake, we assume that the total production of phytoplankton is not that high. In this connection, an abundance of protists (protist plankton) indicates is an indication for the existence of an additional source of organic matter in Lake Khanka. PMID- 10868063 TI - [Interrelations of forest and steppe elements in the fauna of herpetobionts of the middle and lower Don river steppe zone]. AB - Herpetobionts of the forest and steppe biocoenoses were studied in the region of their contact under the conditions of the Middle and Lower Don flow limiting for the forest existence. The species composition of coleopterans of the ravine forests comprises 96 species. The dominant complex of herpetobionts of the steppe and ravine forest biocoenoses includes 13 and 9 species, respectively. Forest species predominate in both forest and steppe habitats of the northern regions, while the meadow species, predominate in the southern regions. The forest species Carabus excellens was found in the steppe zone (oak forest of the Derkul' skii region) for the first time. PMID- 10868064 TI - [Comparative analysis of the life span in HA and BA Drosophila melanogaster]. AB - We studied lifespan (LS) of Drosophila melanogaster strains HA and BA. The first one was generated by long-term selection for low male sexual activity and features decreased LS as well as the changes in a relatively constant index- coefficient of LS variation. The BA strain was derived from the HA strain by reverse selection and has the mean LS and coefficient of LS variation similar to those of the normal strain D-32. Analysis of the death curves of the HA strain insects does not exclude existence of a subpopulation with the normal LS within these strain. The obtained data are discussed in the context of possible involvement of mobile element transposition in animal LS control. PMID- 10868065 TI - [Development of Tilletia caries (D.C.) Tul. in callus and suspension wheat cultures]. AB - All phases of ontogenesis of Tilletia caries were found in combined suspension and callus culture of the wheat and causative agent of common bunt of wheat. Newly formed spores were observed on calluses of the susceptible species Triticum aestivum and resistant species T. timopheevi within 90 and 120 days after inoculation, respectively, which can be used for their production in the laboratory. On the other hand, susceptibility of the callus culture of the resistant species T. timopheevi suggests different responses of the callus cultures and vegetating plants to the pathogen. The capacity of Tilletia caries of growing on callus culture of T. timopheevi indicates to the possibility of overcoming a resistance barrier of the "timopheevi" type, which would allow a study of this phenomenon for selection purposes. PMID- 10868066 TI - [Effect of various organic compounds on the growth and hydrocarbon production by sulfur-reducing bacteria]. AB - The effects of lactate, pyruvate and ethanol on growth and formation of extracellular hydrocarbons by Desulfovibrio desulfuricans 1799 cultivated in H2 + CO2 atmosphere were studied. It was shown that sulfate reducing bacteria grow and produce hydrocarbons on all studied carbon and energy sources. Substitution of lactate in the medium for pyruvate or ethanol decreased only insignificantly the amount of synthesized hydrocarbons with concomitant increase in the content of isoform. PMID- 10868067 TI - [Ultrastructural study of local lesions and surrounding tissues in Gomphrena globosa L. leaves infected by tobacco mosaic virus]. AB - The cells in the local chlorotic areas of Gomphrena globosa L. leaves infected with tobacco mosaic virus (TMV) accumulate a significant amount of viral particles. The cells in the surrounding tissue have a relatively low number of virions, which are often degraded. The cells surrounding the forming local lesion have pronounced signs of activated autolysis. We believe that lysis activation provides for the viral particles destruction and is an important factor limiting TMV accumulation in the cells outside the chlorotic areas. PMID- 10868068 TI - [Effect of antioxidant Mannich base acetate on the growth, cyclic AMP level, and Ca(2+) uptake by yeast cells in Saccharomyces cerevisiae]. AB - We have studied the effect of Mannich base acetate (MBA), an antioxidant, on growth of the yeast, as well as on the level of cAMP and Ca2+ uptake by them. We demonstrate that the antioxidant induced an increase in the level of cAMP in yeast cells and a decrease in Ca2+ uptake by the cells. The effect of the antioxidant depends on the level of Ca2+ in the growth medium. PMID- 10868069 TI - [XV Meeting on "Preservation of genetic resources"]. PMID- 10868070 TI - Exploring the use of qualitative methods in published health services and management research. AB - There is interest in promoting greater use of qualitative methods in health care research. However, little is known about the volume or characteristics of published studies that use qualitative methods. This article explores these issues through a systematic review of 3 years (1995-1997) of articles classified as research in nine core health services research and management journals. The findings show that only about one in seven published research articles used qualitative methods. Two of the nine journals reviewed contributed 45 percent of the total number of articles using qualitative methods. Four journals contributed a combined 2 percent of this total number. The primary purposes in using these methods are description and articulating stakeholder perspectives. There is no standard number of pages devoted by journals to these studies or evidence that they require more journal space on average than quantitative studies. Most of the studies reviewed presented little or no information on methodology. These findings clarify future areas of emphasis for both editors and researchers wishing to promote the use of qualitative methodology in published health care research. PMID- 10868071 TI - Expenditures for physician services under alternative models of managed care. AB - This study compares expenditures for physician services in a closed panel gatekeeper health maintenance organization (HMO) and an open panel point of service HMO that share the same physician network. The study uses administrative files of the two study HMOs for 1994-1995 to assess differences in spending for primary care physicians' (PCPs') services, specialists' services, and total physician services. When the copayments for PCP visits and PCP-referred specialist visits were $0, total physician expenditures were 4 percent higher in the gatekeeper HMO than in the point of service plan (p < .05). When the copayments for PCP visits and PCP-referred specialist visits were $10, total physician expenditures ranged from equal in both HMOs to 7 percent higher in the gatekeeper HMO (p < .01), depending on the copayment for self-referred visits. Expenditures for specialists' services were not higher in the point of service plan. The authors conclude that direct patient access to specialists does not necessarily result in higher physician or specialist expenditures in HMOs. PMID- 10868072 TI - The impact of prior authorization on outpatient utilization in managed behavioral health plans. AB - This study examines how preauthorization affects outpatient behavioral health utilization under managed care by comparing plans with similar benefits, but differing in the number of visits authorized. The authors compare plans primarily authorizing in increments of 5 visits to plans authorizing in increments of 10 visits. They analyze the likelihood of terminating outpatient service between the two groups using conditional logistic regression. Results suggest that patients whose treatment is authorized in increments of 5 sessions are nearly 3 times more likely to terminate treatment at exactly the fifth visit than if their treatment is authorized in increments of 10 sessions conditional on being in treatment until the 5th visit. The likelihood of termination peaks in both the 5- and 10 session authorization at the 10th visit, but the difference is not statistically significant. The authorization effect differs by provider type and is weaker among psychiatrists than among nonphysician providers. PMID- 10868073 TI - Access to medical care reported by Asians and Pacific Islanders in a West Coast physician group association. AB - This study examines access to medical care for Asians and Pacific Islanders in the United States, using a survey of patients receiving care provided by a physician group practice association concentrated on the West Coast. Asians and Pacific Islanders who had used their health plan in the past year had worse access to health care than whites, blacks, Hispanics, and Native American or other ethnicities. The odds that Asians reported that they had adequate access ranged from about one quarter to three quarters that of whites, depending on the measure. Cultural differences and associated communication problems may explain the access problems experienced by Asians. Interventions need to be developed to address the problems with access to services, and better translation services may play an important role in improving access to care for Asians. Future studies need to clarify why Asians were more vulnerable to the access problems examined than other ethnic groups that might experience similar barriers. PMID- 10868074 TI - Impact of Oregon's priority list on Medicaid beneficiaries. AB - Since Oregon's controversial priority list was implemented in 1994, there has been only anecdotal evidence available on its possible impact on Medicaid beneficiaries. The authors surveyed over 1,400 beneficiaries to determine how often a service was denied because it was below the line, what kinds of services these represent, and any resulting health impact. About one third of all respondents reported that they had needed a service that Medicaid would not cover; in 38 percent of these cases, the reason was that the service was below the line. Frequently mentioned services included hernia repair, chiropractic treatment, dental splints, and newborn circumcision. About half of the respondents received the service anyway, often by paying for it themselves. Of those unsuccessful in getting the service, many reported that their health had worsened as a result. However, there was no evidence that getting (or not getting) the service had a causal impact on health status. PMID- 10868075 TI - Predicting the fate and effects of tributyltin in marine systems. AB - The available data indicate that sediment-water partitioning, bioaccumulation, and the toxicity responses for tributyltin (TBT) are predictable when using some of the assumptions and tenets of the equilibrium partitioning method, toxicokinetic modeling (1CFOK), and critical body residue (CBR) approach. Because TBT is ionizable, its speciation is strongly affected by pH, which appears to cause large variations in the octanol-water partition coefficient. In marine systems, and in freshwater systems with high pH, TBT occurs predominantly in the hydroxide form, which may explain the hydrophobic properties and its EqP behavior. Organic carbon in sediment (> 0.2%) appears to be the major controlling factor for sediment-water partitioning. The equilibrium organic carbon-normalized sediment-water partition coefficient (Koc) in marine environments is approximately 32,000 (log10 Koc approximately 4.5), which was determined from direct measurement and confirmed by the relationship between the lipid-normalized bioconcentration factor (BCF) in porewater and the biota-sediment accumulation factor (BSAF). The conclusion that sediment-water partitioning of TBT in marine systems follows EqP is supported by the similarity between its Kow and Koc and the correlation between the sediment-water partition coefficient (Kp) and sediment TOC, which results from the influence of organic carbon on pore-water concentrations. Even though the rates of uptake and elimination control tissue residues and lipid content appears to have no bearing on the amount of TBT that is accumulated, the species specific BSAF is useful for examining bioaccumulation, sediment-water partitioning, and the toxicity response. Although TBT is hydrophobic and appears to have a propensity to accumulate in lipid, the rates of uptake and elimination, not thermodynamics, appear to control whole-body tissue concentrations. Support for a toxicokinetic approach for predicting tissue residues is found in BCF and BSAF values for several species that are far in excess of that predicted by simple thermodynamic partitioning and in the comparisons of observed and predicted bioaccumulation values based on toxicokinetic coefficients. This observation is counter to the assumption of EqP that the route of uptake is of no consequence under equilibrium conditions. For TBT, it appears that kinetics determine tissue residues and that body lipid is important only for regulating the toxic response, not the amount bioaccumulated. Unlike those for neutral hydrophobic organic compounds, the toxicokinetics for this one toxicant are highly variable in diverse species but relatively accurate in predicting the amount bioaccumulated and the resulting toxicity response. For the CBR approach to be useful, a relatively constant tissue residue for a given biological response is necessary. Several studies support the CBR approach because certain biological effects, such as mortality and growth inhibition, occur at a relatively constant TBT tissue concentration. For TBT, the lethal whole-body tissue concentration affecting 50% of individuals (LR50) exhibits little variation, occurring at approximately 48 micrograms/g (166 nmol/g) dry weight in a range of species. Direct evidence and correlation of the LC50 and the bioconcentration factor (BCF) support this observation. Impaired growth, a sublethal response, also appears to be associated with a relatively constant tissue concentration, which has also been demonstrated by direct measurement and indirectly by regression of the BCF and LOEC. The lowest-observed-effect tissue residue (LOER) associated with impaired growth for several species was approximately 3 micrograms/g (10.4 nmol/g) dry wt. Because of the small number of studies linking growth impairment and tissue concentrations, additional studies are needed to confirm these values. (ABSTRACT TRUNCATED) PMID- 10868076 TI - Fate and effects of diazinon. AB - Diazinon use has significantly increased since its introduction more than four decades ago. Thus, today we are faced with environmental and health consequences that are largely inseparable from the insecticide's benefits. Fortunately, the research to date is of immeasurable value in making sound scientific and policy decisions regarding diazinon use. Overall, research shows that diazinon is globally widespread, having distributed to all environmental media. Residential uses, and its ubiquity under many farming practices, contribute to extensive non point-source pollution. In general, diazinon is degraded fairly rapidly in natural settings, although results have been variable and some degradation products are at least as toxic as the parent compound. Diazinon exhibits high acute toxicity to a wide variety of animals, leading to a wide range of sublethal biochemical effects, damage to specific target organs and tissues, cytotoxic and genotoxic effects, reproductive damage, and adverse ecological impacts. Its biological fate is complex, mediated largely by diverse metabolic mechanisms. Further research and monitoring are needed in a number of areas. For instance, it is important to develop a better understanding of the mechanism of diazinon's highly lethal effects on birds. Use restrictions at golf courses and sod farms are a welcome step, but there are still widespread avian exposures from orchards and lawns. Continued diazinon use at current rates also poses a clear threat to aquatic ecosystems and to important species such as salmon and bluegill sunfish. Although the research presented here does not indicate threats to humans from the pesticide, Wright (1990) suggests that people may be at substantial risk in unregulated settings. Further research is also needed to resolve the matter of the potential carcinogenicity of diazinon. As with all pesticides, diazinon use can result in the so-called pesticide treadmill wherein pesticide use necessitates further use as insects develop resistance and natural predators are eliminated (Gliessman 1998). It is critical that all pesticides be used with great care to minimize this consequence to avoid a repeat the occurrence in 1965 in the Culiacan Valley of Mexico. There, excessive pesticide use resulted in cotton pests that were resistant to all available insecticides, forcing growers to entirely abandon production (Wright 1990). However, used carefully, diazinon represents a powerful agricultural tool available to assist in the continued production of foodstuffs for a rapidly growing world population. PMID- 10868077 TI - Trace element contamination in Antarctic ecosystems. AB - Data concerning trace element concentrations in both abiotic and biotic components of the Antarctic ecosystems are summarized here to be used as a first background database for pollution detection. Antarctic ancient ice and snow cores have been used to assess past and present-day changes in global atmospheric levels of certain trace elements. Concentrations of Pb, Cd, and Hg in the Antarctic tropospheric cell have varied according to glacial and interglacial periods before humans began to contaminate the atmosphere with these metals. Based on data of Pb concentrations in the Antarctic ancient ice cores and samples of recent snow, most of the Pb in Antarctica is anthropogenic. Moreover, this metal has decreased in recent years as a consequence of the reduced use of leaded gasoline in countries of the Southern Hemisphere. Reliable experimental and field data have indicated, however, that human activity in Antarctica contributes significantly to increasing atmospheric Pb levels in this continent, whereas the environmental impact of other metals such as Cd, Zn, or Hg is restricted to the area a few hundreds of meters about the anthropogenic source. Trace element concentrations in Antarctic abiotic matrixes are generally at ultratrace levels, challenging analytical detection limits and increasing the risk of unwanted contamination. Pb and Hg concentrations in Antarctic snow, surficial soil, air, and marine sediment have been considered as the lowest concentrations ever reported. Conversely, concentrations in Antarctic biota, are comparable to those from polar and temperate areas of the Northern Hemisphere, in particular Cd and Hg. Environmental and biological factors favoring a greater metal accumulation by Antarctic biota are discussed. Growth rate of the organisms and detoxification mechanisms (e.g., metallothioneins, molting cycles) are largely affected by the extreme environmental conditions in Antarctica (water supply, temperature regimen, light availability) that probably interfere with uptake, storage, and excretion of trace elements by organisms. On the other hand, environmental factors such as the upwelling of Cd-rich waters and local volcanism undoubtedly increase the bioavailability of metals in the Antarctic environment. In this context, several Antarctic organisms such as fish, mollusks, lichens, and mosses have been proposed as suitable biomonitors, and their trace element concentrations have been suggested as baselines. Structure and dynamics of Antarctic ecosystems as well as quantifying metal point sources and long-range atmospheric transport require in-depth studies to improve the assessment of human impact in Antarctica. PMID- 10868078 TI - Trace metals in Antarctica related to climate change and increasing human impact. AB - Metals are natural constituents of the abiotic and biotic components of all ecosystems, and under natural conditions they are cycled within and between the geochemical spheres--the atmosphere, lithosphere, hydrosphere, and biosphere--at quite steady fluxes. In the second half of the twentieth century, the huge increase in energy and mineral consumption determined anthropogenic emissions of several metals exceeding those from natural sources, e.g., volcanoes and windborne soil particles. In the Northern Hemisphere, the biogeochemical cycles of Pb, Cd, Zn, Cu, and other metals were significantly altered, even in Arctic regions. On the contrary, available data on trace metal concentrations in abiotic matrices from continental Antarctica, summarized in this review, suggest that the biogeochemical cycle of Pb is probably the only one that has been significantly altered by anthropogenic emissions in Antarctica and elsewhere in the Southern Hemisphere, especially in the period 1950-1975. Environmental contamination by other metals from anthropogenic sources in Antarctica itself can generally only be detected in snow samples taken within a range of a few kilometers or several hundred meters from scientific stations. Local metal pollution from human activities in Antarctica may compromise studies aimed at assessing the biogeochemical cycle of trace elements and the effects of global climate change. Thus, this review focuses on concentrations of metals in atmospheric particulate, snow, surface soils, and freshwater from the Antarctic continent and surface sediments and seawater from the Southern Ocean, which can plausibly be regarded as global background values of trace elements. These baselines are also necessary in view of the construction of new stations, the expansion of existing facilities to support research, and the growth of tourism and fisheries. Despite difficulties in making comparisons with data from other remote areas of the world, concentrations of trace metals in most samples of atmospheric particulates, snow, ice, soils, and marine sediments from Antarctica can be taken as global background levels. Comparison between the results of trace element surveys in marine waters of the Southern Ocean and in other seas is practically impossible. The upwelling or subduction of water masses, the seasonality in ice cover and in phytoplankton biomass, the low fallout of atmospheric dust, and many other peculiar characteristics of the Southern Ocean make concentrations of trace metals in surface waters quite variable in space and time. The depletion of nutrients in surface waters, which is a regular feature of many marine environments, rarely occurs in the Southern Ocean. Waters in some regions are characterized by very low concentrations of Fe and Mn, whereas in others the content of Cd is relatively high at the beginning of summer and may decrease about one order of magnitude during the phytoplankton bloom. Although in most Antarctic coastal ecosystems the input of metals from geochemical and anthropogenic sources and from long-range transport is negligible, concentrations of Cd in the waters and biota may be higher than in waters and related species of organisms from polluted coastal areas. Like the Southern Ocean, Antarctic lakes have many peculiar characteristics. They are often perennially ice covered and without outlet, and their water, which is gained only from short-term melting of snow and glaciers in summer, is lost mainly by sublimation of surface ice. Several lakes are distinctly stratified: the water under the ice may be cool, rich in oxygen, and among the cleanest and clearest of natural waters, whereas water near the bottom becomes anoxic, tepid, and richer in major and trace elements. Considering the specificity of Antarctic environments, to evaluate the extent and consequences of global changes and increasing human activities in Antarctica itself, research on the biogeochemistry of trace metals and monitoring programs PMID- 10868079 TI - Mental health service use by Americans with severe mental illnesses. AB - BACKGROUND: The aim of this study was to determine the patterns and determinants of service use in severely mentally ill persons drawn from the National Institute of Mental Health Epidemiological Catchment Area (ECA) program, a community-based epidemiologic survey. This information provides a baseline against which to track ongoing changes in the US mental health service system. METHODS: Severe mental illness (SMI) was defined according to US Senate Appropriations Committee guidelines. Comparisons were made with persons who had a mental disorder that did not meet these criteria (non-SMI). Sociodemographic factors, and 1-year volume and intensity of mental or addictive services use were determined. Differences between those who used services and those who did not were examined using logistic regression. RESULTS: Persons with SMI differed from persons with non-SMI in most sociodemographic characteristics. A higher proportion of persons with SMI used ambulatory services, but the mean number of visits per person did not differ from the non-SMI population. Persons with SMI comprised the bulk of hospital inpatients admitted during a 1-year period. Several significant sociodemographic determinants of service use were found, with different patterns for general medical and specialty service use, pointing out potential barriers to care. CONCLUSIONS: As health care reform measures continue to be debated, attention to the service needs of the severely mentally ill is of crucial importance. Pre managed care (pre-1993) baseline service use benchmarks will be essential to assess the impact of managed care on access to care, particularly for the severely mentally ill. Periodic collection of epidemiologic data on prevalence and service use would thus greatly facilitate service planning and addressing barriers to receiving mental health services in this population. PMID- 10868080 TI - Methods used for parasuicide: results of the WHO/EURO Multicentre Study on Parasuicide. AB - BACKGROUND: National suicide statistics show remarkable differences in the frequencies of various methods used for completed suicide. The WHO/EURO Multicentre Study on Parasuicide makes possible for the first time an international comparison of the frequencies of methods used in attempted suicide, because the data are based on geographical catchment areas of medical institutions. METHOD: Ongoing standardized monitoring of attempted suicide in all medical institutions serving the catchment areas was performed in 14 centres in 12 European countries. The data analysis is based on 20,649 events involving 15,530 persons, recorded between 1989 and 1993. RESULTS: The comparison of rates per 100,000 shows striking differences between the centres. The highest rates for drug overdoses were found for female attempters in Oxford (347/100,000), Helsinki (238/100,000) and Stockholm (221/100,000). Guipuzcoa had the lowest rates (61/100,000). The differences were most prominent in the age group 15-24, with outstanding rates for women in Oxford (653/100,000), which was mainly due to the frequent use of analgesics. Szeged had outstandingly high rates for pesticides and solvents. In some centres the use of multiple methods was frequent. CONCLUSIONS: There is a need, especially for areas with high frequencies for certain methods, to understand the factors involved and to develop new and specific prevention projects and to monitor their effects. The WHO/EURO Multicentre Study on Parasuicide has proved to be a useful and reliable instrument for continuous monitoring of trends in parasuicide. PMID- 10868081 TI - The geographical mobility of severely mentally ill residents in London. AB - BACKGROUND: There is currently great concern over the demands on psychiatric services in metropolitan areas in most developed countries, and this is exemplified in capital cities. These greater demands were not anticipated by those planning psychiatric services and the consequences have led to insufficient beds in many areas. We investigated the geographical mobility (the number of changes of address in the past 2 years) of patients presenting to services in greater London, to determine whether this might be a possible factor in the increased demand. METHOD: The geographical mobility of the severely mentally ill was determined by taking a random sample of all psychiatric admissions to hospitals serving residents in the London area over the calendar year of 1994 (n = 156) and an equivalent sample of patients in an established community mental health team (n = 74) in one area (Paddington). The extent of geographical movement was determined for the 2 years prior to interview. RESULTS: Greater geographical movement in the in-patient group was found for those living in inner London compared with outer London and for patients admitted to hospitals outside their catchment area. Twenty-eight percent of the in-patient sample had changed address in the year before admission (including 13% more than once) and 39% had changed address in the 2 years prior to admission. By contrast, the patients seen by the community psychiatric team were less than half as likely to have changed address over the previous year as the in-patients, and none of the community team's patients had changed address more than once over the previous year. The geographically mobile patients had significantly longer periods in hospital than geographically stable patients. CONCLUSION: Geographical mobility of psychiatric patients in London is high and is particularly marked for those presenting for in patient treatment. These findings suggest that greater mobility could be one of the most important reasons for the higher than expected demands on psychiatric services and the difficulties in maintaining contact with patients in London in general and inner London in particular. More attention should be paid to geographical mobility as a predictor of psychiatric service use, and it is recommended that it is recorded regularly in information systems. PMID- 10868082 TI - Evaluation and validation of a measure profiling needs and problems of psychiatric patients in the community: a Malaysian study. AB - BACKGROUND: The Profile of Community Psychiatry Clients (PCPC) was developed in a Sydney-based sample of those with a mental illness as a 35-item measure of likely need for service recognition, review and possible assistance. METHODS: This study has three principal objectives. Firstly, to test the utility of the PCPC measure in a very different region and culture. Secondly, to review the factor structure in an independent sample. Thirdly, to pursue the extent to which the PCPC might serve as a measure of likely need, by obtaining three differing reference viewpoints of need (i.e. clients, their carers, and case managers) and examining responses against PCPC scores. The PCPC was given to a sample of 333 Malaysian clients living in the community, together with two other measures of morbidity and disability. In addition, case managers, family members and clients were requested to directly rate the level of need for service assistance. RESULTS: A principal components analysis favoured a six-factor solution, with PCPC factor scores and total scores intercorrelated with subscale and total scores on the Life Skills Profile (LSP) and Health of the Nation Outcome Scales (HoNOS). The correlation coefficients supported the concurrent validity of the derived PCPC scales. Family members rated the clients' needs as greater than did case managers who, in turn, rated severity of needs greater than the clients themselves. Most importantly, PCPC scores correlated more highly than did LSP and HoNOS scores with need estimates derived by all three rating groups, providing strong support for the PCPC meeting its objective as a measure of putative need. In addition, a refined 23-item version of the PCPC was derived, which retained the capacity of the PCPC to correlate strongly with needs estimates. CONCLUSIONS: This Malaysian study supports the use of the PCPC in a culture where service provision and family support for those with a mental illness vary considerably from Western regions, while its validation as a measure of need for service is supported. PMID- 10868084 TI - The Experience of Caregiving Inventory: further evidence. AB - BACKGROUND: The aim of this study was to reexamine the construct validity of the Experience of Caregiving Inventory (ECI) using new, independent data from a population of patients and their carers. This involved re-testing the ECI within the stress-coping model, but adding new variables which included independently rated (rather than carer-rated) assessments of the patient's symptoms and disabilities, a rating of social support (this time for the patient rather than the carer) and a measure of a range of service inputs. METHOD: Data were available on 69 patients and their carers from the PRiSM Psychosis Study. Two regression analyses were performed; the first to establish the extent to which the ECI predicted GHQ scores and the second to examine predictors of the ECI selected from the wider dataset on the basis of their likely relationship to carer appraisal. The second regression analysis was performed in two stages, allowing the effect of service factors to be assessed after controlling for the impact of patient personal characteristics such as illness-related or environmental factors. RESULTS: ECI scores accounted for 27% of the variance of GHQ scores. Over one-third of the ECI negative appraisal can be explained by a combination of patient disability (the Social Behaviour Score Total), extent of patients' social network (Social Network Schedule: Number of Friends) and involvement of a Community Psychiatric Nurse (CPN). At the same level of patient morbidity and informal social network, CPN contact reduced ECI scores. CONCLUSION: As hypothesised, ECI scores correlated significantly with the other measures in the directions predicted by the stress-coping model; that is, negative caregiving as measured by the ECI predicted carer morbidity, while it in turn was predicted by a combination of stressor variables (patient symptoms and disability) acting to increase it, and mediating variables (social support, service inputs) acting to reduce it. Implications for services are discussed. PMID- 10868083 TI - Factors associated with problem behaviors in Turkish immigrant children in The Netherlands. AB - BACKGROUND: The objective of this study was to examine the relationship between child, parent, family/support, and stress variables and problem behaviors in Turkish immigrant children in the Netherlands. METHODS: Parents of 833 children were interviewed and administered a Turkish version of the Child Behavior Checklist for ages 4 through 18 and a Turkish immigrant assessment questionnaire. RESULTS: Increased integration (i.e., children belonging to a second generation of immigrants, older children) generally reduced the risk for problem behaviors, while frequent arguments, divorce, psychological problems, and convictions/incarcerations increased the risk for problem behaviors. CONCLUSIONS: Results indicate that problem behaviors are associated with the high level of separation faced by Turkish immigrant families and that more integration leads to lower levels of problem behavior. Additionally, migration history alone does not contribute to problem behavior. Factors in the family (e.g., quarrels, divorce, conviction) associated with problem behaviors in Turkish immigrants are similar to those found in Dutch and American populations. Future studies could examine predictors of problem behaviors in other culturally separated immigrant groups in the Netherlands, and in Turkish immigrant groups in other countries. PMID- 10868085 TI - [Unemployment and mental disorders]. PMID- 10868086 TI - [Multiple sclerosis]. PMID- 10868087 TI - [Hydrocephalus in children]. PMID- 10868088 TI - [Deliveries during transportation and shortly after admission to hospital]. AB - BACKGROUND: The community midwife services in Oppland county are under review. Many pregnant women live far from hospital. This is a study of the occurrence of deliveries during transport to hospital. MATERIAL AND METHODS: We used hospital records to identify women who had given birth during transport in Oppland county in the period 1989-97. We mailed a questionnaire to these and to 100 women who had given birth shortly after arrival at Lillehammer hospital in 1996-97. RESULTS: 133 women gave birth to 133 children during transport in Oppland 1989 97. This amounts to 0.69% of all births in the county. One child died during birth. A midwife assisted 71% of the women. However, only one third of these midwives had been on call at the time. Most women gave birth at night, and had their second child. INTERPRETATION: Deliveries during transport are inevitable and mostly uncomplicated. Midwives on call may provide qualified assistance during transport when the delivery is imminent. PMID- 10868089 TI - [Polyneuropathy is an early finding in primary systemic amyloidosis]. AB - BACKGROUND: Polyneuropathy is associated with several pathological conditions. Amyloidosis is a less common cause of polyneuropathy. Amyloidosis is caused by the accumulation of insoluble protein fibrils (amyloid) in the extracellular matrix. Primary systemic amyloidosis is caused by B-lymphocyte dyscrasia. Polyneuropathy is the first sign of primary systemic amyloidosis in about 20% of the cases, and is characterised by relentless progression, painfulness, and prominent symptoms of autonomic neuropathy. MATERIAL AND METHODS: We describe three cases of primary systemic amyloidosis that started with polyneuropathy. RESULTS: The diagnosis of primary systemic amyloidosis is often delayed more than two years after the onset of polyneuropathy. Biopsy of rectum, fat tissue, bone marrow, or peripheral nerve are diagnostic tools. INTERPRETATION: The survival of non-treated patients is about 18 months after the diagnosis. With chemotherapy the survival is prolonged to approximately 38 months. PMID- 10868090 TI - [Neurosurgical shunt treatment of children with hydrocephalus]. AB - BACKGROUND: The aims of this study were to determine the ten-year outcome of hydrocephalic children, both in terms of academic results, social skills, physical functioning and surgical morbidity and mortality. MATERIAL AND METHODS: A cohort of 128 children shunted for hydrocephalus at the National Hospital in Oslo between 1985 and 1988 were retrospectively analysed by registering their medical records and in a questionnaire survey. RESULTS: We found that 23 (18%) children died during the ten-year observation period. Six patients died from shunt system failure, ten died from their malignant tumour, six died of their complex cerebral malformations and one died of unknown cause. Of the 105 (82%) children still alive, 104 returned our questionnaire. 92 (88%) reported to be attending a normal school, while 77 (74%) are in the same grade as children of similar age. Of those children at same grade level, 39 (51%) needed extra tutoring, while 35 (45%) reported speech and writing difficulties. Among children not in their adequate grade level, 25 (93%) (p < 0.01) needed extra tutoring, and 23 (85%) (p < 0.01) reported speech and writing difficulties. 50 (48%) participate in physical exercise along with the other children, while 53 (51%) report having normal social relations with children of the same age. 34 (33%) has suffered from epileptogenic seizures, while 24 (23%) daily use prescribed antiepileptic drugs. During the ten-year follow-up period, 108 (84%) patients had a total of 342 shunt revisions. We found no correlation between high incidence of revisions and negative long-term outlook. 99 out of the 105 children alive are considered to be life-long dependent of their shunt system. INTERPRETATION: The vast majority of the children shunted for hydrocephalus that are still alive, have a good or satisfactory level of functioning ten years after their first shunt insertion. This view seems to be shared by the parents of these children, of whom 50 (48%) state that the development of their child has been "very good" compared to what they initially feared. PMID- 10868091 TI - [Intracranial aneurysms associated with cerebral arteriovenous malformations]. AB - BACKGROUND: Patients known to harbour a cerebral arteriovenous malformation that is inaccessible to therapy may have a second bleeding into the subarachnoid space, but from another source. MATERIAL AND METHODS: Two patients had previously bled from an intracranial arteriovenous malformation; both were considered inaccessible to surgical or endovascular repair. The patients were then admitted to hospital with symptoms and signs suggesting a second subarachnoid haemorrhage. RESULTS: Diagnostic work-up demonstrated an aneurysm as the probable source of haemorrhage in both patients. INTERPRETATION: Patients harbouring an intracranial arteriovenous malformation are much more likely to develop an associated intracranial aneurysm than patients without such malformations, and a second bleeding in these patients will more often arise from the associated aneurysm. The cause of the frequent association of an aneurysm is probably haemodynamic stress due to the increased blood flow through the feeding artery. These patients suffer subarachnoid haemorrhage more often than patients with either an aneurysm or a malformation alone. The therapeutic strategy should be carefully individualized and the aneurysm should more often have priority. PMID- 10868092 TI - [Health and quality of life among long-term unemployed]. AB - BACKGROUND: Many studies have shown an association between unemployment and poor health. MATERIAL AND METHODS: This cross-sectional survey from 1993-94 using structured interviews and questionnaires describes health and quality of life among 148 long-term unemployed in Lillesand, Norway. RESULTS: Compared with the general population, somatoform conditions and anxiety symptoms were twice as common among the unemployed, and depression three times as common. There was a higher frequency of depression among unemployed men than among unemployed women. The unemployed had a lower quality of life score than the general population; the difference was more than twice as large among men as among women. Unemployed aged 30-39 reported the strongest symptoms of anxiety and depression. INTERPRETATION: The reason why unemployed men are more often stricken by depression and report lower quality of life than women, may be that men experience the job-loss more existentially threatening than do women. PMID- 10868093 TI - [Mastocytosis--a review illustrated by two case reports]. AB - BACKGROUND: Mastocytosis includes a range of disorders characterised by accumulation of tissue mast cells. These are derived from pluripotent haematopoietic stem cells. Recent research has improved the understanding of the mastocytosis pathogenesis. Organ manifestations and symptoms are highly variable. MATERIAL AND METHODS: Two cases of systemic mast cell disease are presented. RESULTS: One patient had urticaria pigmentosa and systemic mast cell disease; the other had systemic mast cell disease and myelodysplastic changes in the bone marrow. INTERPRETATION: The two cases illustrate different manifestations and different prognosis for various types of mastocytosis. PMID- 10868095 TI - [Subluxation of the radial head]. AB - It is our impression that subluxation of the caput radii is a little known injury, especially in general practice. This article presents a review of the literature. Subluxation of the caput radii is a common injury among toddlers. The yearly incidence is approximately 1% among children under five years of age, with peak incidence around two years. It affects the left elbow more frequently than the right and girls more frequently than boys. Pull of the affected arm as mechanism of injury, decreased arm movement and absence of local swelling is typical for this condition. Radiographic abnormalities are very subtle or non existent. Reduction can usually be obtained by supination of the affected arm. After successful reduction, normal function of the arm is regained within 15 minutes in most instances. In cases with typical mechanism of injury and typical clinical findings, reduction can be performed without X-rays. However, it should be checked that spontaneous and normal use of the affected arm is obtained after reduction. PMID- 10868094 TI - [HLA-B27 in Bechterew's disease]. AB - BACKGROUND: The role of HLA-B27 in rheumatic disease is reviewed. MATERIAL AND METHODS: The PubMed database was searched for the code words HLA-B27, ankylosing spondylitis, or their combinations. The diagnostic value of HLA-B27 for ankylosing spondylitis was calculated using Bayes' theorem. RESULTS: It is still not clear whether HLA-B27 is a contributing causal factor for ankylosing spondylitis and reactive arthritis. Knowledge of a patient's HLA-B27 status for establishing prognostic information is of uncertain clinical value. INTERPRETATION: Knowledge of HLA-B27 status has limited diagnostic value. HLA-B27 status is clinically most informative when the test result is negative, as ankylosing spondylitis can be ruled out. PMID- 10868096 TI - [Rheumatological manifestations in HIV infections]. AB - BACKGROUND: The recognised clinical spectrum of disease associated with HIV infection is rapidly expanding and now includes a variety of rheumatological manifestations. MATERIAL AND METHODS: In this review of the literature of the last 15 years, we present the most common rheumatic manifestations described in association with HIV infection. RESULTS: Manifestations include a wide array of articular syndromes and autoimmune manifestations such as Reiter's syndrome, psoriatic arthritis, HIV associated arthritis and septic arthritis. Autoimmune diseases associated with HIV infection include a Sjogren-like syndrome, myopathies and systemic vasculitis. INTERPRETATION: Rheumatological manifestations of HIV infection may present earlier than clinical signs of the infection itself. Steroid and cytostatic treatment of rheumatic diseases may worsen the HIV disease. PMID- 10868097 TI - [Neurologic follow up of premature infants and other neonatal at-risk patients in Norway]. AB - BACKGROUND: Infants with very low birthweight (< 1,500 grams) are at increased risk of neurological disabilities and impairments later in childhood. We wanted to study whether the paediatric departments in Norway had organised routine neurological follow-up programmes for these patients, and in particular whether the child habilitation departments participated in the follow-up. MATERIAL AND METHODS: A questionnaire was returned from 19 of 23 pediatric departments. The questionnaire also included other neonatal risk groups. The results showed that one senior paediatrician was in charge of the follow-up at most departments. The neonatologists worked in collaboration with the child neurologists, and most often the child habilitation department was consulted in the neonatal period. RESULTS: Only seven departments used a standardised follow-up programme. Main high risk groups were infants with birthweight < 1,500 grams and infants with birth asphyxia. There was a lack of consensus with regard to other inclusion criteria, time for follow-up and type of examinations. Only a few of the departments had performed follow-up studies concerning neurological sequelae and quality of life for this group of patients. INTERPRETATION: Registration of neurological disease in premature children should be mandatory for every department with neonatal intensive medicine. PMID- 10868098 TI - [Magnetic resonance tomography of the heart]. AB - Magnetic resonance imaging is a rarely performed procedure in cardiac disease, mainly due to easily available alternative techniques, especially echocardiography. MR imaging may, however, provide unique anatomical and functional information and represents a valuable supplement to the alternative invasive and non-invasive modalities. Clinical indications for MR imaging of the heart include congenital heart disease, acute and chronic ischaemic heart disease, valvular heart disease, cardiomyopathy, right ventricular dysplasia, pericardial disease, and cardiac tumours. In the near future, MR imaging may provide a complete "one-stop" evaluation of cardiac disease. PMID- 10868099 TI - [Neurologic magnetic resonance tomography--indications and practical use]. AB - Magnetic resonance tomography is considered the most important technique in modern neuroradiology. This article demonstrates the importance of MR imaging in the diagnosis of diseases of the central nervous system. Modern neuroradiology is completely dependent upon this modality, and sections and departments of neuroradiology should have great responsibility for both diagnostic MR imaging and the day-to-day operation MR laboratories. PMID- 10868100 TI - [Alendronate also for women without established osteoporosis?]. PMID- 10868101 TI - [Celiac disease and neurologic disease]. PMID- 10868103 TI - [Prophylaxis of venous thromboembolism]. PMID- 10868104 TI - [Acute porphyria]. PMID- 10868105 TI - [Child abuse and osteogenesis imperfecta. How do we distinguish?]. AB - Osteogenesis imperfecta is a hereditary connective tissue disorder. Typical manifestations are fragile bones with multiple bone fractures and bone deformities. A history of minimal or no trauma and recurrent fractures is a feature of OI, but is also typical of non-accidental injury (NAI). OI and NAI are relevant differential diagnoses when a child presents with unexplained fractures. The differential diagnostic problems are reviewed, all of which are important for the child both in terms of treatment and for prognosis, socially and medicolegally. We conclude that comprehensive clinical evaluation is adequate for differential diagnosis and that both OI and NAI can be diagnosed by positive anamnestic and objective signs. Mild OI IV without other signs than fracture(s) is very rare and the new entity temporary brittle bone disease is hypothetical; the diagnosis of these two clinical pictures is unacceptable in small children. Routine analysis of collagens should not be performed. PMID- 10868106 TI - [Child and adolescents with psychotic disorders. A literature review of psychology and course]. AB - The article is a literature review of psychosis in children and adolescents. Earlier studies point to psychosis in children and adolescence being of the same types as the ones seen in adults. Schizophrenia has an early onset in boys, there are more premorbid dysfunctions, a gradual onset of disease, a number of unspecific symptoms and a poorer prognosis than for adults. For the children with bipolar affective disorder the prognosis is similar to that of adults, and it is better than for the schizophrenics. The relationships between developmental disorders and schizophrenia, and between hyperactivity syndromes and manic depressive disorders need to be further clarified, as does the validity of the follow-up investigations. PMID- 10868107 TI - [Amebiasis]. AB - Amoebiasis caused by the enteric protozoan Entamoeba histolytica is a widespread parasitic disease, which causes 40,000 to 100,000 deaths per year. Cases diagnosed in Denmark are always imported. Infection results from ingestion of amoebic cysts, which after excystation form trophozoits in the small intestine, colonize the bowel lumen and invade the intestinal epithelium resulting in amoebic colitis. Spread to the liver and formation of amoebic liver abscesses occurs in one third of the cases, whereas other extraintestinal manifestations are rare. Amoebic colitis and liver abscess have a good prognosis when treated with metronidazole, whereas complications such as necrotizing colitis, peritonitis and pericarditis have a high mortality. An early diagnosis and treatment is therefore important. Intestinal amoebiasis is diagnosed by demonstration of E. histolytica cysts or amoebae in the stools, whereas serology is of value in diagnosing extraintestinal amoebiasis in nonendemic regions. PMID- 10868108 TI - [Genital prolapse]. AB - The prevalence of genital prolapse in women is unknown. The development of prolapse is dependent on the pelvic floor muscles and connective tissue. Risk factors are vaginal birth, obstipation, high abdominal pressure and surgical procedures. Preventive measures are discussed. The classification of prolapse is somewhat difficult. Conservative treatment with pessaries and pelvic floor muscle exercises and various surgical procedures are discussed. PMID- 10868109 TI - [Medical graduates from the Odense University 1972-1998. Recruitment and employment]. AB - The aim was to create and maintain a registration of all medical graduates from Odense University from the first graduates in 1972 and to describe the graduates' background and subsequent professional employment. All graduates from 1972 to 1998 are included. About 55% of the graduates came from secondary schools in the country og Funen, 20% from southern Jutland and 15% from Copenhagen and Jutland north of Vejle. About 45% became general practitioners after 5-22 years (median 11 years), a third of them in the county of Funen. Twenty-five percent became consultants after 11-17 years (median 14 years), twenty-five percent of these in the county of Funen. The cumulated death rate was 5% for the ages 30-55 years. At 17 years after graduation 83% had become specialists, at ten years the number was 55%. In conclusion, Odense University recruits 75% of its medical students from southern Denmark. Of the graduates that become general practitioners, about two thirds are employed in southern Denmark, for consultants the corresponding figure is 50%. PMID- 10868110 TI - [The Danish hernia database--the first year]. AB - The Danish Hernia Data Base started registration of inguinal herniorrhaphies 1. Jan. 1998. The purpose of the data base is, on the basis of well-defines goals, to estimate and improve the different aspects of inguinal hernia surgery. The data base has participation from 81 hospital departments and outpatient clinics. During this first year 9380 herniorrhaphies have been included, corresponding to 95% of herniorraphies performed in Denmark. The results from the data base show, that 17% of operations are for a recurrence (with a rate of reoperation within one year of at least 1.6%), that a large number of different types of operations are used, that mesh is used in approx. 70% of herniorrhaphies, that 14% of operations are performed under infiltration anaesthesia and that 55% of the operations are performed on an outpatient basis. The data base collaboration, in combination with frequent meetings where results from the data base are discussed, seems to be a useful tool in documenting and developing this type of operation. PMID- 10868111 TI - [Tension-free herniotomy using the Lichtenstein's method. Results of five years' experience]. AB - The results after 230 initial open tension-free hernioplasties a.m. Lichtenstein for groin hernias are evaluated 24 months after operation. Of the 167 primary operations 57% were performed under local analgesia, and 68% of the patients were discharged on the day of operation after primary herniotomy. Infection occurred after 2.7% of the operations, but in no case did the mesh have to be removed. Complications occurred after 7.4% of the operations, necessitating reoperation after 4.3% of the operations. After 24 months 96% of live patients were controlled for recurrences. After 154 primary herniotomies 2.6% recurred. No indirect hernia recurred, 3.6% of direct hernias recurred. After 57 operations for secondary hernias 7.0% recurred. Thirty-eight different surgeons operated 1 19 hernias (median 4). It is concluded that recurrences after open tension-free hernia-repairs are rare, complications few, the operation is simple to perform and can be done under local analgesia in a same day regime. PMID- 10868112 TI - [Endoscopic treatment of colorectal obstruction with self-expandable metal endoprosthesis]. AB - Relief of colo-rectal obstruction by means of self-expandable metal mesh stents (SEMS) has been suggested for palliation and acute decompression followed by optimization of the patients' general condition prior to definitive surgery. Twelve patients with high operative risk and/or metastatic disease were selected for stenting with a dedicated colorectal partly covered SEMS (Choo Colo-Rectal Stent, Solco Intermed Co., Seoul, Korea). Stent deployment was successful in nine, two of whom had total obstruction. In one a guidewire perforation was treated conservatively. In two patients (one benign stricture, and one rectal cancer) the stents migrated within three weeks. One re-obstructed. In the remaining six patients colonic decompression was achieved, and the stents have been patent until death (33-175 days, four patients) or are still patent (follow up 35-80 days). These results are promising, but data from several centres should be compiled prospectively in a standardized fashion in order to allow for assessment of the method's safety and success rates before randomized trials can be initiated. PMID- 10868114 TI - [Picture of the month. Toxic epidermal necrolysis]. PMID- 10868113 TI - [The relationship of Helicobacter pylori to iron status--serum ferritin and hemoglobin. A seroepidemiologic survey of 2794 Danes]. AB - We evaluated the influence of Helicobacter pylori (H. pylori) infection assessed by the levels of H. pylori serum IgG-antibodies, on iron status (serum ferritin and haemoglobin) in 2794 Danes (1425 men), aged 30-60 years. The seroprevalence of H. pylori antibodies increased with age (p < 0.01). Median serum ferritin levels were significantly lower in seropositive than in seronegative men and postmenopausal women (men 114 micrograms/L vs. 120 micrograms/L, p = 0.01; premenopausal women 37 micrograms/L vs. 40 micrograms/L, p = 0.13; postmenopausal women 63 micrograms/L vs. 77 micrograms/L, p = 0.02). Seropositive subjects had a higher prevalence of iron deficiency (serum ferritin < 15 micrograms/L) than seronegative subjects. H. pylori infection has a negative influence on iron status. We hypothesize that this may be caused by increased blood losses from the gastric mucosa. PMID- 10868115 TI - [Electromagnetic fields and pediatric cancer]. PMID- 10868116 TI - [H:S accreditation]. PMID- 10868117 TI - [Increased knowledge about our genes required more counseling]. PMID- 10868118 TI - [Molecular biology and patient counseling in the year 2000]. PMID- 10868119 TI - [Molecular biology and genetic counseling]. PMID- 10868120 TI - ["Genetification" in the year of 2000]. PMID- 10868121 TI - [Investigation of patients with syncope: normal orthostatic tolerance]. PMID- 10868122 TI - [Prevention of deformation (disability) in rheumatoid arthritis]. PMID- 10868123 TI - [Ruptured abdominal aortic aneurysms]. PMID- 10868124 TI - [Sensitivity and specificity in the diagnosis of whiplash injury. What is the list of answers?]. PMID- 10868125 TI - [Screening for Down syndrome and congenital abnormalities in the first versus the second trimester]. PMID- 10868126 TI - Back surgery. Coming back from pain. PMID- 10868127 TI - Health tips. Getting the most from juice. PMID- 10868129 TI - Extra oxygen during colon surgery may cut infection risk. PMID- 10868128 TI - Thalidomide--a treatment for cancer? PMID- 10868130 TI - Clinical trials. Removing the mystery. PMID- 10868131 TI - Botulinum toxin. Can a poison help? PMID- 10868132 TI - Home blood pressure monitors. Tools you can use. PMID- 10868133 TI - I've heard that injections of human growth hormone can slow aging. Is this true? PMID- 10868134 TI - Can allergy shots provide relief from hay fever even after the shots are discontinued? PMID- 10868135 TI - Colon cancer. Regular screenings could save your life. PMID- 10868136 TI - Respiratory viral infections in immunocompetent and immunocompromised persons. AB - The acute respiratory illnesses are the most common type of acute illness in the United States today. The respiratory viruses--which include influenza viruses, parainfluenza viruses, respiratory syncytial virus (RSV), rhinoviruses, coronaviruses, and adenoviruses--cause the majority of these illnesses. Some of these viruses cause illness throughout the year, whereas others are most common in winter. All population groups experience these infections and illnesses. As the number of elderly persons and those with underlying disease increases, awareness is growing that these common infections can have serious consequences. This has recently been emphasized for immunocompromised persons. At the M.D. Anderson Cancer Center (MDACC), infection surveillance of mostly hospitalized adults with leukemia or a recent bone marrow transplant yielded a respiratory virus from 181 of 668 (27.1%) respiratory illness episodes. In descending order of frequency, infections with RSV, rhinoviruses, influenza viruses, parainfluenza viruses, and adenoviruses were detected in each of three surveillance years. High frequencies of nosocomial acquisition occurred, as has been noted in prior reports. Similarly, persistence of infection and high frequencies of pneumonia and death among infected patients occurred, which have also been noted earlier. At MDACC, pneumonia occurred in 58-78% of infected patients, and 22-44% died. The role of the virus infection in many cases of pneumonia is uncertain, but death from pure viral pneumonia is well documented. A number of immune deficiencies in this patient population and options for control of these infections have been described that can, respectively, account for the medical problem and provide ways to approach prevention and treatment. PMID- 10868137 TI - Community respiratory virus infections in immunocompromised patients with cancer. AB - Community respiratory viruses, such as respiratory syncytial virus (RSV), influenza viruses, parainfluenza viruses, adenoviruses, and picornaviruses, are an important cause of respiratory disease in the immunocompromised adult with cancer. Recent studies have demonstrated that a minimum of 31% of adult bone marrow transplant (BMT) recipients and 18% of adults with leukemia who are hospitalized with an acute respiratory illness have a community respiratory virus infection. The temporal occurrence of these infections in immunocompromised patients tends to mirror their occurrence in the community. The clinical illnesses range from self-limited upper respiratory illnesses to fatal pneumonias, depending on the type of virus and the type and degree of immunosuppression. The pneumonias may be viral, bacterial/fungal, or mixed. The highest frequency of progression to fatal viral pneumonia has been reported for RSV infections in recently transplanted BMT recipients and myelosuppressed patients with leukemia. Studies have suggested that early therapy for RSV pneumonia with a combination of aerosolized ribavirin and intravenous immunoglobulin may be of benefit. Defining effective prophylactic and therapeutic strategies will be a challenge, given the diversity of viruses, the wide spectrum of immunocompromised patients with varying vulnerability to serious community respiratory virus disease, and the frequent presence of other opportunistic infections and medical problems. A combination of antiviral drugs and immunotherapy may need to be considered for their potential additive effect as well as to prevent the emergence of resistant virus, as occurs during monotherapy for influenza with amantadine or rimantadine. The optimal therapies need to be defined in controlled trials; however, it appears that a favorable response will hinge on the initiation of therapy at an early stage of the respiratory illness. PMID- 10868138 TI - Community respiratory viruses in individuals with human immunodeficiency virus infection. AB - Respiratory viruses, particularly influenza viruses, respiratory syncytial virus (RSV), parainfluenza viruses, and adenoviruses, are ubiquitous pathogens among humans, especially among young children. However, relatively little is known about the impact of these common infections on individuals with the human immunodeficiency virus (HIV). A review of the literature identifies three key areas that need further exploration. First, moderate-to-severe and even fatal lower respiratory viral illnesses in HIV-infected individuals have been reported. In general, the clinical presentation of these respiratory viral infections in persons with HIV infection is similar to their presentation in individuals without HIV infection. The major exception is the occurrence of fulminant, and often fatal, disseminated adenovirus infection in adults and children with HIV disease. Despite these reports, no information is available regarding the frequency of moderate-to-severe respiratory viral illnesses in individuals with HIV infection. Epidemiologic studies of respiratory viral illnesses in cohorts of HIV-infected adults and children are needed. Second, prolonged shedding of respiratory viruses for weeks and even months has been documented in HIV-infected adults and children. The frequency of prolonged shedding in this population has not been well defined, but data from a small newborn cohort study suggest that, at least for RSV, prolonged shedding is common. Prolonged respiratory viral shedding has implications for infection control in medical facilities where HIV infected individuals are treated and in nursing homes, child care centers, and group foster homes that provide care for HIV-infected individuals. Therapies to help eliminate these chronic viral infections should be explored. Finally, indirect evidence suggests that respiratory viral infection may result in changes in HIV replication and, theoretically, HIV disease progression. Increased HIV-1 replication has been demonstrated in vitro in T lymphoma cells exposed to genetic material from adenovirus. Increased HIV replication in peripheral blood from adults following inactivated influenza vaccination has been reported. The impact of natural respiratory viral infection (and perhaps vaccination against these pathogens) on HIV replication and disease progression will be an important area of study. PMID- 10868139 TI - Respiratory virus infections after marrow transplant: the Fred Hutchinson Cancer Research Center experience. AB - Respiratory virus infections are becoming increasingly appreciated causes of morbidity and mortality in bone marrow transplant recipients. Fred Hutchinson Cancer Research Center (FHCRC) has had considerable experience with respiratory syncytial virus (RSV), parainfluenza, influenza, and rhinovirus infections in these patients over the past decade. Overall, RSV accounted for the majority of community-acquired respiratory virus infections (35%), followed by parainfluenza virus (30%), rhinovirus (25%), and influenza virus (11%). Pneumonia occurred frequently among those infected with RSV (49%) or parainfluenza (22%) but infrequently among those infected with influenza virus (< 10%) or rhinovirus (3%). In one study conducted at FHCRC, intravenous ribavirin was not effective against established RSV pneumonia. In another, ongoing study, however, short course aerosolized ribavirin appears to reduce progression of upper respiratory tract infection to pneumonia, but further study is needed. Neither strategy had an obvious impact on viral shedding, although persistence of viral shedding did not correlate with either the development or the outcome of pneumonia. PMID- 10868140 TI - Community respiratory viruses: organ transplant recipients. AB - Respiratory infections are common after solid organ transplantation, but the significance of community respiratory viral infections in this patient population has not been determined. Review of the literature indicates that infection of organ transplant recipients by community respiratory viruses can result in significant morbidity with some associated mortality. These viruses include respiratory syncytial virus (RSV), parainfluenza virus (PIV), influenza virus, and adenovirus. As in normal hosts, infection of organ transplant recipients by these viruses can result in limited upper respiratory tract symptoms, such as rhinorrhea, cough, and fever. Immunocompromised patients can also have lower respiratory tract infection, resulting in bronchiolitis, pneumonitis, respiratory failure, and death. The highest incidence of infection with these viruses is reported in lung transplant recipients, with an incidence up to 21%. In addition to the effects of the usual immunosuppressant regimen, lung transplant recipients have altered lung immunity due to impaired ciliary clearance, poor cough reflex, and abnormal lymphatic drainage, predisposing these patients to lower respiratory tract infections. Of additional importance to organ transplant recipients is the correlation of organ rejection to recent viral infections with these agents. Influenza A and B, PIV, and adenovirus have been reported to be associated with acute rejection in renal transplant recipients. Diagnosis of these infections is often made by positive respiratory cultures, often with a delay between symptom onset and diagnosis. Clinical trials of antiviral agents in this patient population have not been carried out, and treatment has often been limited to severe, life-threatening cases. PMID- 10868141 TI - A comparison of cytomegalovirus and community respiratory viruses in immunocompromised patients. AB - Infection of organ transplant recipients with herpesviruses, especially cytomegalovirus (CMV), has been an important barrier to successful transplantation. Measures to prevent CMV infection have evolved based on the biology and epidemiology of the virus. Recently, community-acquired respiratory viruses have been recognized as an important cause of morbidity and mortality in immunocompromised hosts. It is unlikely that the strategies used to prevent and treat CMV disease will be applicable to the community respiratory viruses, owing to their different biology and epidemiology. Basic epidemiologic questions that focus on defining risk factors for disease and death from community-acquired respiratory viruses in the immunocompromised host have not been answered. The lack of established risk factors and a rapid "gold standard" diagnostic test for community-acquired respiratory viruses in the immunocompromised host has had a negative impact on the diagnosis and treatment of these infections. The study of CMV disease in organ transplant patients may serve as a blueprint for studies of community respiratory virus infections. PMID- 10868142 TI - Respiratory virus infections in bone marrow transplant recipients: the European perspective. AB - The role of respiratory viruses in severe complications following bone marrow transplantation was examined in a survey of selected transplant centers in Europe belonging to the European Group for Blood and Marrow Transplantation. Information was collected on 57 cases of infections with respiratory viruses, including adenoviruses. In two centers that had conducted prospective studies of bone marrow recipients with respiratory symptoms, the frequency of these infections was 7.1% and 4%, respectively. The severity of these infections varied. Pneumonias caused by respiratory syncytial virus and adenovirus were associated with mortality rates of 60% and 75%, respectively, whereas pneumonia caused by parainfluenza had a mortality rate of 20%. Influenza infections progressed to fatal pneumonia in 17% of cases. Results of antiviral therapy varied. As the results of our survey showed, respiratory virus infections are not infrequent after bone marrow transplantation and are associated with significant morbidity and mortality. PMID- 10868143 TI - Infection control of nosocomial respiratory viral disease in the immunocompromised host. AB - Among immunocompromised adults, such as bone marrow transplant recipients, more than half of respiratory viral infections are complicated by pneumonia, with an associated mortality rate > 50%. Nosocomial transmission of respiratory viral pathogens, such as respiratory syncytial virus (RSV) and influenza, in the immunocompromised patient has been reported frequently and usually occurs during a community outbreak. In view of the poor outcome in this subset of patients, intensive efforts should be directed at instituting prevention measures that would interrupt nosocomial transmission. At M.D. Anderson Cancer Center, a multifaceted infection control strategy resulted in a significant decrease in and almost complete interruption of the nosocomial transmission of RSV infections in immunocompromised patients over a 3-year period (1994-1996). For influenza virus, special emphasis should be given to vaccination of hospital personnel before the influenza season to prevent and control nosocomial transmission. In highly immunocompromised patients, prophylactic use of antiviral agents should be considered during an outbreak or when the frequency of nosocomial transmission is high. An aggressive multifaceted infection control strategy appears to be effective in reducing the frequency of nosocomial transmission of respiratory viral infections in immunocompromised patients. Universal and timely influenza vaccination of hospital personnel who care for immunocompromised patients is necessary. PMID- 10868144 TI - Prevention and treatment of influenza in immunocompromised patients. AB - Influenza A and B viruses cause serious, sometimes fatal, disease in immunocompromised patients, particularly bone marrow and solid organ transplant recipients. Protracted disease has also been recognized in certain oncology and HIV-infected patients. Currently available inactivated vaccines are variably immunogenic in such groups. Poor humoral immune responses are seen within 2 years of bone marrow transplantation, often following solid organ transplantation, and commonly in patients with advanced HIV infection. Oral amantadine and rimantadine are useful for prophylaxis and treatment of influenza A virus infections, but their efficacy, particularly in treatment of severe disease, has not been rigorously established in immunocompromised hosts. Case reports document the emergence of drug-resistant variants and prolonged viral shedding in some patients. Aerosol and intravenous ribavirin has been used to treat severe influenza in small numbers of immunosuppressed patients, but the efficacy of ribavirin by either route has not been established in such patients. The neuraminidase inhibitor GG167 is active in experimental influenza but requires topical application to the respiratory tract and has had limited clinical study in natural influenza. More effective interventions for serious influenza infections will likely require combinations of antiviral drugs. PMID- 10868145 TI - Prevention and treatment of respiratory syncytial virus and parainfluenza viruses in immunocompromised patients. AB - Immunocompromised patients are vulnerable to severe infections due to respiratory syncytial virus (RSV) and parainfluenza viruses (PIV), and therefore prevention and treatment strategies must be considered. The prevention of RSV disease with high-titer RSV-specific immune globulin has been documented in very young children but has not been systematically studied in high-risk adults. Vaccines against RSV and PIV are under development, but their use in immunocompromised patients is problematic. Ribavirin aerosol therapy is licensed for the treatment of RSV in pediatric patients and has also been used to treat RSV disease in adults and PIV disease in severely immunocompromised children and adults. Uncontrolled trials show that early therapy with ribavirin aerosol may be beneficial, but treatment of pneumonia in patients with respiratory failure is rarely successful. Other potential treatments for RSV or PIV disease include high dose, short-duration ribavirin therapy; combined immunoglobulin and ribavirin therapy; polyclonal and monoclonal antibodies; and, potentially, immunomodulators. PMID- 10868146 TI - Adenovirus infections in immunocompromised patients. AB - Adenovirus infections have been reported in as many as one-fifth of bone marrow transplant (BMT) recipients and patients with acquired immunodeficiency syndrome (AIDS), and in a lesser, though still prominent, proportion of organ transplant recipients. The relative contributions of primary infections versus reactivations from latency in immunocompromised patients remain unclear. Compared with adult BMT recipients, pediatric BMT recipients appear to be infected by adenovirus more frequently and earlier in the post-transplant period. The diagnosis of adenovirus infection is complicated by the existence of > 40 viral serotypes, although certain subgroups are more likely to be involved in certain patient populations. Adenoviruses are responsible for a broad range of clinical diseases that may be associated with high mortality, including pneumonia, hepatitis, encephalitis, hemorrhagic cystitis, and gastroenteritis. The clinical and histopathologic features of adenovirus disease may resemble those of cytomegalovirus disease, potentially complicating the diagnosis. Risk factors for clinical adenovirus disease include the number of sites from which the virus is cultured and, in BMT recipients, the presence of moderate to severe acute graft-versus-host disease. PMID- 10868147 TI - Community respiratory viral infections in the immunocompromised host: past, present, and future directions. PMID- 10868149 TI - The Health Care 250. PMID- 10868150 TI - Circle of life. PMID- 10868148 TI - The slow road to executive diversity. PMID- 10868151 TI - Advancing healthcare practices: linking quality with culture. PMID- 10868152 TI - Happy Y2K. New and old challenges for the nurse administrator. PMID- 10868153 TI - On the definition and measurement of nursing care. PMID- 10868154 TI - Documentation's effect on reimbursement for rapid treatment status patients. PMID- 10868155 TI - Advancing careers through membership in the American Academy of Nursing. PMID- 10868156 TI - Subacute care. A competitive response to providing geriatric care. PMID- 10868157 TI - Australia's nursing workforce in perspective. PMID- 10868158 TI - Intensive care unit cross training: saving dollars while retaining staff. PMID- 10868159 TI - A lesson from the last nursing shortage: the influence of work values on career satisfaction with nursing. AB - With an emerging national nursing shortage and evidence of dissatisfied registered nurses, it is useful to revisit the issue of satisfaction by analyzing data from the prior nursing shortage in 1989. The authors explore the relationships that work values have on the satisfaction that nurses experience with a career in nursing. The relevance of the findings are discussed within the context of the nursing shortage. PMID- 10868160 TI - Creating a hospital-based nursing research fellowship program for staff nurses. AB - Staff nurses are expected to participate in nursing research and to use study findings. Insufficient institutional support and uncertainties about how to participate in the research process can prevent staff nurses from meeting these expectations. We describe a newly developed nursing research fellowship program designed for staff nurses who practice in an acute care setting. An overview of the entire curriculum and the outcomes of the first year of the program are described. PMID- 10868161 TI - Moving toward restraint-free patient care. AB - After reviewing the literature and regulatory requirements, a multidisciplinary team developed a comprehensive restraint reduction program that has reduced the use of restraints by more than 60% in the acute care setting. The authors discuss the research-based restraint education program and the implementation of a restraint consultant role. PMID- 10868162 TI - The relationship of nursing practice models and job satisfaction outcomes. AB - The concept of nursing practice models--shared governance--has attracted the attention of nursing administrators in the last decade in response to maintaining nursing job satisfaction, quality care, and fiscal viability. This article contains a critique of existing research that tests the effects of nursing practice models on nursing outcomes, predominantly job contentment and autonomy. The results of the studies reviewed indicate that implementation of nursing practice models enhances job satisfaction, increases personal power and accountability, and improves climatic change. In addition, implementation of nursing practice models is highly dependent on manager skill in leading and maintaining the change process. PMID- 10868163 TI - Diagnosis and treatment of attention deficit hyperactivity disorder (ADHD). AB - OBJECTIVE: The objective of this NIH Consensus Statement is to inform the biomedical research and clinical practice communities of the results of the NIH Consensus Development Conference on Diagnosis and Treatment of Attention Deficit Hyperactivity Disorder (ADHD). The statement provides state-of-the-art information regarding effective treatments for ADHD and presents the conclusions and recommendations of the consensus panel regarding these issues. In addition, the statement identifies those areas of study that deserve further investigation. Upon completion of this educational activity, the reader should possess a clear working clinical knowledge of the state of the art regarding this topic. The target audience of clinicians for this statement includes, but is not limited to, psychiatrists, family practitioners, pediatricians, internists, neurologists psychologists, and behavioral medicine specialists. PARTICIPANTS: Participants were a non-Federal, nonadvocate, 13-member panel representing the fields of psychology, psychiatry, neurology, pediatrics, epidemiology, biostatistics, education and the public. In addition, 31 experts from these same fields presented data to the panel and a conference audience of 1215. EVIDENCE: The literature was searched through Medline and an extensive bibliography of references was provided to the panel and the conference audience. Experts prepared abstracts with relevant citations from the literature. Scientific evidence was given precedence over clinical anecdotal experience. CONSENSUS PROCESS: The panel, answering predefined questions, developed their conclusions based on the scientific evidence presented in open forum and the scientific literature. The panel composed a draft statement that was read in its entirety and circulated to the experts and the audience for comment. Thereafter, the panel resolved conflicting recommendations and released a revised statement at the end of the conference. The panel finalized the revisions within a few weeks after the conference. The draft statement was made available on the World Wide Web immediately following its release at the conference and was updated with the panel's final revisions. CONCLUSIONS: Attention deficit hyperactivity disorder or ADHD is a commonly diagnosed behavioral disorder of childhood that represents a costly major public health problem. Children with ADHD have pronounced impairments and can experience long-term adverse effects on academic performance, vocational success, and social-emotional development which have a profound impact on individuals, families, schools, and society. Despite progress in the assessment, diagnosis, and treatment of ADHD, this disorder and its treatment have remained controversial, especially the use of psychostimulants for both short and long-term treatment. Although an independent diagnostic test for ADHD does not exist, there is evidence supporting the validity of the disorder. Further research is needed on the dimensional aspects of ADHD, as well as the comorbid (coexisting) conditions present in both childhood and adult forms. Studies, (primarily short term, approximately three months) including randomized clinical trials, have established the efficacy of stimulants and psychosocial treatments for alleviating the symptoms of ADHD and associated aggressiveness and have indicated that stimulants are more effective than psychosocial therapies in treating these symptoms. Because of the lack of consistent improvement beyond the core symptoms and the paucity of long-term studies (beyond 14 months), there is a need for longer term studies with drugs and behavioral modalities and their combination. Although trials are underway, conclusive recommendations concerning treatment for the long term cannot be made presently. There are wide variations in the use of psychostimulants across communities and physicians, suggesting no consensus regarding which ADHD patients should be treated with psychostimulants. (ABSTRACT PMID- 10868164 TI - ["Medicine on the Intenet"-- model course for physicians in the use of the Internet]. AB - The Internet is efficient in coping with the ever-increasing volume of medical knowledge. This and other functions have lead to its integration into daily medical practice. It is therefore imperative to disseminate its use among physicians, as some, unfamiliar with the personal computer (PC), may be reluctant to learn to use it. We therefore developed a course for training physicians in the basics of Internet use, consisting of 6 sessions of 2 hours each. It includes training in PC operation as well as in the main Internet functions. The topics are medicine-oriented, including Medline search, general info search, exploring medical sites and reading medical journals on-line. 41 physicians (8 groups of 4 6 each) with various specialties attended, of whom 38 completed a feedback questionnaire 4 weeks later. 79% reported using the Internet either at home or at their working place. The function used mostly was Medline Search (68%). Although 83% had not been familiar with the PC before, 96% stated that the course was clear and understandable and 99% expressed satisfaction with the quality of the teaching. Based on these results we recommend this course as a model for teaching physicians the use of the Internet. PMID- 10868165 TI - [Does incidence of hepatitis A increase during shmitah (the Sabbatical year)?]. AB - In Israel the biblical injunction of the sabbatical year (shmitah) prevails, whereby all Jewish-owned land should lie fallow during every seventh year. Consequently, it is customary for members of the orthodox Jewish community to eat only produce grown by non-Jews (Arabs). Many Arab farmers use sewage water for irrigation and since such water could be infected with hepatitis A virus (HAV), there is concern about the possibility of HAV epidemics during the sabbatical year. We therefore we examined the data of the Israeli Center for Disease Control (ICDC). We found no obvious increase in incidence of viral hepatitis during, nor in the year immediately after, all sabbatical years since 1951. However, the data was not comprehensive as it included only partial information on morbidity from HAV in our Jewish inhabitants. Also, there was no data specific for the orthodox Jewish community, which is especially at risk for HAV from sewage-irrigated vegetables. Irrespective of shmitah, there should be constant effort to prevent HAV infection in Israel. PMID- 10868166 TI - [Transmission of hepatitis C in hemodialysis patients--does one-time exposure result in chronic hepatitis?]. AB - Acquired infection with hepatitis C virus (HCV) in hemodialysis patients has been described lately. In dialysis units in Italy and France, the prevalence and incidence of HCV are 20-60% and 1-2%, respectively. Most infected patients develop chronic hepatitis. The clinical presentation of acute HCV in hemodialysis patients is very mild and therefore the diagnosis is often made only by laboratory tests. Acute infection is usually followed by mild elevation of liver enzymes and the presence of HCV-RNA and anti-HCV in serum. We report a 48-year old man on hemodialysis who developed acute hepatitis C. The diagnosis was made by finding mild elevation of liver enzymes and the presence of HCV-RNA in his serum. A few months later, he developed severe hepatitis which was followed by rapid deterioration in liver function. However, the virus was eradicated and liver function tests became normal. Surprisingly, serum anti-HCV antibodies were detected 5 months later. PMID- 10868167 TI - [Spontaneous bilateral ectopic pregnancy--an rare and dangerous occurrence]. AB - Bilateral, spontaneous ectopic pregnancy is rare (1 in 125-1580 ectopic pregnancies). We describe a 30-year-old, unmarried woman with no predisposing factors for ectopic pregnancy who presented in hypovolemic shock, in the 7th week of gestation, complaining of abdominal pain. On immediate laparoscopy there were found blood and clots in the abdominal cavity, a left ampullar pregnancy (5 cm in diameter), and there was active bleeding from the fimbria of the right tube. Bilateral salpingectomy was performed and she received 3 units of packed red cells. She was discharged in good condition 3 days later. The pathologic diagnosis was pregnancy in each tube. This case emphasizes the need for thorough sonographic and laparoscopic observation in order not to miss the presence of bilateral ectopic pregnancy. PMID- 10868168 TI - [Psychogenic polydipsia leading to water intoxication]. AB - Psychogenic polydipsia and its frequent complication, water intoxication, are well-known to psychologists. There are biochemical and psychiatric theories of psychogenic polydipsia which often correlate with each other. A 48-year-old man with chronic paranoid schizophrenia developed symptoms of psychogenic polydipsia. This provoked disturbances in electrolyte balance, resulting in a rapid decrease in serum sodium concentration and eventual death. There is a paucity of information and little awareness of this problem in the professional literature. PMID- 10868169 TI - [Hemorrhoidectomy using a circular stapler]. AB - We compared traditional local excision of hemorrhoids in 40 patients (group A) with excision using the CDH-33 surgical stapler designed for bowel anastomosis, in 41 (group B). In this technique a purse-string suture is prepared 3 cm above the dentate line, and the stapler is inserted and used to form a mucosa-mucosal anastomosis. This technique is less time-consuming than the traditional technique. Data were collected from patients' medical files and from detailed questionnaires in which symptoms prior to and after operation were reported. Mean ages were 53.0 and 51.5 years and male/female ratios were 1.0:1.2 and 1.0:1.1 respectively, neither significant. The most common complaints in both groups were pain and rectal bleeding. All patients had a lower-GI investigation prior to operation to exclude other causes of rectal bleeding. Recovery averaged 2 months in both groups. Patient satisfaction was assessed by decrease or absence of symptoms on return to normal daily activities. Satisfaction tended to be greater in group B. More patients in group A complained of tightness and discomfort at the operative site, but this was not significant. We are extending our study to a larger number of patients to determine if there are statistically significant differences between the results of the 2 methods. PMID- 10868171 TI - [Vaginal vault prolapse: repair by sacrospinous ligament fixation]. AB - Sacrospinous ligament fixation is a well-known method for correction of vaginal vault prolapse. The procedure is primarily indicated after hysterectomy and as a prophylactic measure for total uterine prolapse. 8 women with post-hysterectomy vaginal vault prolapse aged 48-72 years, were referred 1-25 years following primary surgery. Sacrospinous ligament fixation was elected to enable simultaneous correction of cystocele and rectocele, and to preserve sexual function. All operations were completed without significant complications. In 6 of the 8 patients located for long-term interview, 1 reported mild bulging of the introitus, another mild urinary stress incontinence, urgency and frequency and 1 reported frequency only. Sexual function in 4 was without complaints. Defecation in all patients was normal. We conclude that this operation is safe and effective. We encourage gynecologic surgeons to consult an experienced tutor prior to performing this procedure, as this type of surgery is quite dangerous and many gynecologists are not familiar enough with it. PMID- 10868170 TI - [Corneal infection, cause and effect on vision]. AB - We conducted a retrospective 5-year survey of corneal infections treated in the ophthalmology ward of Hasharon Hospital. The most frequent type of corneal infection was corneal abscess; the most frequent cause was Staphylococcus albus, although this bacterium is not reported as a frequent cause of corneal infections. There was improvement in visual acuity in 69.2% and no change in 15.4%. Corneal infection by Pseudomonas aeruginosa was the most frequent cause of worsening of corneal acuity (23.08%). The greatest improvement of visual acuity was in those with corneal ulcers. The worst visual acuity was in those with corneal abscesses. Pseudomonas aeruginosa was the main cause of infection in contact-lens wearers. In the world medical literature, Staphylococcus albus is considered of very low virulence. This bacterium was the most frequent cause of corneal infections in our study so it may have greater virulence in Israel. PMID- 10868172 TI - [Diabetic ketoacidosis during the Ramadan fast]. AB - We report a 15-year-old Muslim boy with insulin-dependent diabetes mellitus (IDDM) who presented with diabetic ketoacidosis (DKA) during the Muslim Ramadan month of day-time fasting. DKA apparently occurred due to omitting pre-lunch insulin combined with dehydration and overeating during the permitted sunset-to sunrise meals. It is well-known that fasting accelerates development of lipolysis and ketosis and increases glucagon levels. Thus, these pathophysiological aberrations related to fasting in ketosis-prone patients, in conjunction with fasting, endanger metabolic control in IDDM. PMID- 10868173 TI - [Carcinoma of the cecum presenting as intussusception]. AB - In intussusception a proximal bowel segment (intussusceptum) invaginates into the lumen of a distal bowel segment (intussuscipiens). In adults intussusception represents a rare cause of intestinal obstruction and is usually secondary to an identifiable lesion that requires surgical treatment other than that of the intussusception itself. We present a 57-year-old woman with metastatic colonic carcinoma of the ileocecal valve and intussusception of the ileum into the cecum. Right hemicolectomy was performed. PMID- 10868174 TI - [On telomeres and telomerases: the key to cell senescence or immortality]. PMID- 10868175 TI - [Cardiac cachexia]. PMID- 10868176 TI - [Primary subacute hematogenous osteomyelitis in children]. PMID- 10868177 TI - [Neonatal testicular torsion]. PMID- 10868179 TI - [Laparoscopic management of endometrial cancer]. PMID- 10868178 TI - [Combined liver damage from primary biliary cirrhosis, primary sclerosing cholangitis, liver allograft rejection and graft-versus-host disease]. PMID- 10868180 TI - [Blast injury of the ear]. PMID- 10868181 TI - [Treatment of recurrent and chronic tonsillitis]. PMID- 10868183 TI - [Firefighting--job demands, health risks, medical surveillance and retirement]. PMID- 10868182 TI - [Prostacyclin--its function and indications for therapeutic use]. PMID- 10868184 TI - [Joint recommendations of Israel medical societies for prevention of coronary heart disease and atherosclerosis]. PMID- 10868185 TI - [Surgery in the Talmud]. PMID- 10868186 TI - A pediatrician and his mothers and infants. AB - Pediatricians are in a unique place in society by being able not only to care for the health and well-being of mothers and which, are their clinical responsibility, but also by being able to act as advocates for those patients who are often among the most vulnerable of our population. This article illustrates some of these points by referring to Australian Aboriginals from the vast desert areas of Westerns Australia. In remote areas of Western Australia, Aboriginal infants have high rates of low birth weight, failure to thrive and undernutrition. They also have high rates of respiratory, gastrointestinal and other infections. Aboriginal infant mortality has improved significantly over recent years, but Aboriginal health and mortality rates are still much worse than those of non-Aboriginal children and tend to be worst in more remote parts of the state. Overall, Aboriginal infants less than one year in age were hospitalized 9.5 times more frequently than non-Aboriginal infants for respiratory diseases (such as pneumonia, acute bronchiolitis and asthma); diarrheal diseases and skin infections were other very important causes of hospitalization for Aboriginal infants. Another poorly understood aspect of Aboriginal health is their widespread proneness to urinary tract infections. This is very important now in Australian Aboriginals in whom end-stage renal failure is becoming very prevalent. Rapid social and lifesyle changes have been very important in the poor health status of Aboriginals. They are also subject to severe socio-economic discrimination, underemployment, limited education, overcrowding, social depression and severely depressed housing conditions, relative inaccessibility to adequate and nutritious foodstuffs, and limited access to clinical services. Aboriginal people are prone to obesity, hypertension, type-2 diabetes mellitus and cardiovascular diseases. Overuse of alcohol and tobacco smoking have also become important challenges, particularly among adolescents and young adults. For the past twenty years or so, special programs have been developed to help overcome some of these problems; these include immunization programs, an extensive child health care program, special childhood screening programs, and oral rehydration therapy to reduce the high rates of mortality and morbidity associated with diarrheal diseases. These improvements have been achieved despite a set of socio-economic circumstances that face Aboriginal infants and children who live with adverse social factors. This was termed "Down and Out in 1996" in an editorial in The New Scientist (27 January 1996). A strategy that Australian Aboriginals are using now is to increase their own role through Aboriginal controlled health and medical services including child health programs. PMID- 10868187 TI - Antibiotic susceptibilities of Salmonella serogroups isolated from Turkish children. AB - This study is performed to show the serogroup distribution and in-vitro antibiotic susceptibilities of Salmonella species that cause either gastroenteritis with/without bacteremia or enteric fever at Hacettepe University Ihsan Dogramaci Children's Hospital. Of the 309 Salmonella strains evaluated, serogroup B was the most common isolate (56%) followed by serogroup D (33%). Antibiotic susceptibility tests using the disk diffusion technique revealed resistance rates of 43 percent for ampicillin, 41 percent for chloramphenicol, 29 percent for trimethoprim-sulfamethoxazole (SXT) and 32 percent for ceftriaxone among Salmonella serogroup B. The same rates were 10, eight, seven and zero percent for Salmonella serogroup D, and seven, 14, and zero percent for serogroup C, respectively. S.thypi strains susceptible to all antibiotics studied except tetracycline (33% resistant). No resistance was detected against the quinolones. The antibiotic resistance of Salmonella species isolated from children seems to be important, especially in serogroup B. Susceptibility tests should be considered in the antimicrobial therapy of Salmonella infections where indicated. PMID- 10868188 TI - Antibiotic susceptibilities of enterococci isolated from Turkish children. AB - To evaluate the antibiotic resistance rates of enterococci isolated at Hacettepe Children's Hospital, in vitro antibiotic susceptibility tests were performed in 77 enterococci (32 hospital, 45 nonhospital strains) isolated from various clinical specimens in 1994. Microbroth dilution tests against ampicillin, vancomycin, gentamicin and streptomycin were performed according to the NCCLS standards. High-level resistance to aminoglycosides was investigated. Ampicillin resistance rates were 21.9 percent and 2.2 percent for hospital and nonhospital strains, respectively (p < 0.01). The same rates were 46.9 and 13.3 percent for gentamicin (p < 0.01), and 15.6 and 13.3 percent for streptomycin (p = 0.25). No resistance was detected against vancomycin. Six strains (7.8%) showed high-level resistance to both aminoglycosides studied. Special attention should be paid to enterococci isolated from hospitalized patients. Appropriate antibiotic use in serious infections can only be achieved by choosing an appropriate regimen according to antibiotic susceptibility tests. Periodic evaluation of the antibiotic susceptibility patterns of enterococci is necessary for the empirical treatment of infections due to these microorganisms. PMID- 10868189 TI - Experience with oral rehydration therapy in moderately dehydrated children due to diarrhea. AB - Additional reports on oral rehydration therapy (ORT) in the management of moderately dehydrated patients may convince practitioners to convert from high cost intravenous treatment to low-cost and efficient ORT in moderately dehydrated patients. Therefore, the failure rate and its association with admission serum sodium, potassium and bicarbonate levels of 162 moderately dehydrated children with diarrhea treated with ORT were analyzed retrospectively. The overall failure rate was found to be 17.6 percent. This rate did not differ significantly among normonatremic, hyponatremic and hypernatremic patients (16%, 25% and 28%, respectively; p > 0.05), nor did the rate differ among normokalemic, hypokalemic and hyperkalemic patients (16%, 33% and 25%, respectively; p > 0.05). Only moderately and severely acidotic patients had higher failure rates than non acidotic and mildly acidotic cases (21%, 38%, 4% and 14% respectively; p < 0.05). Severely acidotic patients were also rehydrated over a longer time (10.4 +/- 6.6 hours) than were nonacidotic patients (6.7 +/- 2.3 hours, p < 0.001). Thus the degree of acidosis, which is closely related to the clinical severity of dehydration, was found to be much more predictive of ORT failure and the duration of rehydration than were other electrolyte disturbances. Besides correcting dehydration, ORT was safe and effective in treating various electrolyte disturbances. PMID- 10868190 TI - The angiocardiographic analysis of 73 patients with double-outlet right ventricle. AB - The purpose of this study was to determine the cardiac anatomy of patients with double-outlet right ventricle by angiocardiography. A total of 73 patients between the ages of one day and 11 years were examined. The aorta was on the right side of the pulmonary artery in 23 cases (32%), right anterior in 20 (27%) and right posterior in 17 cases (23%). Pulmonary stenosis was found in 53 patients (73%) and subaortic stenosis in six cases. Ventricular septal defect (VSD) was subaortic in 39 cases (52%), remote type in 17 (23%), doubly committed in 10 (13%), and subpulmonic in 9 (12%). Double VSD was noted in two patients. Pulmonary hypertension was more frequent in subpulmonic ventricular septal defect (78%). The most common associated anomalies were atrial septal defect (34%), anomalous coronary arteries (12%) and endocardial cushion defect (10%). Aortic root angiography was not satisfactory in half of the cases with coronary arterial anomaly. In conclusion, double-outlet right ventricle is a complex anomaly, all of whose cardiac features can be successfully demonstrated in detail by echocardiography and angiocardiography. However, in order to determine the anatomy of the coronary arteries, selective coronary angiography may be necessary in some patients. PMID- 10868191 TI - Surgical management of congenital lobar emphysema. AB - Here in nine patients with congenital lobar emphysema who had been treated surgically in the previous 10 years are reported. The ages of the patients at diagnosis ranged from 26 days to 11 months. The six patients whose symptoms started in the neonatal period had more severe dyspnea, cyanosis and respiratory distress. Tube thoracostomy was performed in two of three patients who had been misdiagnosed initially. The affected side was the left upper lobe in five patients, the right upper lobe in three, and the right middle and upper lobes in one patient. Lobectomy was performed in all cases. Dysplasia of the bronchial cartilage was found in six patients and bronchial atresia of the left upper lobe was found in another infant as the etiologic cause of the condition. Although the possibility of conservative management in congenital lobar emphysema has been reported recently, we believe that surgery is the treatment of choice in patients who have persistent or progressive, severe respiratory distress in spite of medical treatment. PMID- 10868192 TI - Transtelephonic ECG versus electrophysiologic study in children with recurrent palpitation attacks. AB - Palpitation may be a terrifying event for children and pose diagnostic problems for pediatric cardiologists. Routine methods often fail to document episodic arrhythmia because the episodes may be brief or infrequent or both. The purpose of this study was to evaluate and compare values of non-invasive and invasive techniques. Twenty-three children (mean age 11.7 +/- 2 years, range 7-16 years) with recurrent palpitation attacks (> 2 times), who had normal physical examinations, ECG, echocardiography, 24 hour ambulatory ECG monitoring and treadmill exercise tests, were included in the study. Redline Model CG 4000 TTE recorders were given to patients for 10 or 20 days. We performed an intracardiac electrophysiologic study (EPS) on 14 patients. The mean number of palpitation attacks was 11.3 +/- 7.4 (median: 5), lasting 9.6 +/- 7 (median 3) minutes. The number of transmitting records was 2 +/- 1.4 (1-20). Of the 23 patients, only 15 (65%) transmitted during the palpitation attacks. Twenty-four-hour ambulatory ECG monitoring findings were normal in all patients. One patient had a wide QRS tachycardia attack in the TTE records. We stimulated ventricular tachycardia in the same patient in the EPS. Among the other patients who transmitted TTE records during palpitation attacks, we diagnosed two cases of concealed accessory pathway and eight cases of dual AV nodal pathway in the EPS. In conclusion, TTE is a sensitive and accurate method for diagnosis and follow-up of patients with cardiac arrhythmia. It can be used before invasive studies in children with recurrent palpitation attacks. PMID- 10868193 TI - Ear, nose and throat findings in cystic fibrosis patients. AB - An ear-nose-throat survey was carried out on 30 children with cystic fibrosis (CF) between two and 17 years of age. There were three (10%) confirmed cases of secretory otitis media. One child had previous surgery for polyps. Ten children had chronic pansinusitis mildly responsive to medical therapy. Hypertrophy of the nasal conchae was detected in eight cases. The data showed that the risk of ear nose-and-throat disease was increased in CF patients, but audiologic problems are not very common in CF patients. CF should be kept in mind in children who have nasal polyps, hypertrophied turbinates or sinusitis unresponsive to medical therapy. PMID- 10868194 TI - A retrospective study on 16 collodion babies. AB - Sixteen collodion babies followed in the Neonatal Care Unit between January 1982 and December 1994 were evaluated retrospectively. The preterm/term ratio was 1.6, and complete shedding of the collodion membrane took an average of 21.9 days (range 18-46 days). Problems noted were marked temperature instability, defective barrier function, increased insensible water loss predisposing to hypernatremic dehydration, cutaneous infections and septicemia. Hypernatremia was observed in 11 (68.7%) and septic infection in seven patients (43.7%). All the infants were treated topically with vaseline containing five percent lactic acid. In the hypernatremic infants, intravenous fluid was administered for rehydration. In the septic infants, antibiotics were used according to the antibiogram. Four of the infants died due to septicemia. The mortality rate was 25 percent, and the major complications included hypernatremia, cutaneous infection and sepsis. PMID- 10868195 TI - Adenosine-sensitive right ventricular tachycardia in children. AB - Right ventricular outflow tachycardia is a distinct subgroup of idiopathic ventricular tachycardia (VT) with characteristic clinical and electrophysiologic properties. This study was planned to evaluate the effects of adenosine on idiopathic right ventricular tachycardia in children, and to assess the long-term efficacy of verapamil treatment. Diagnostic tests including electrocardiogram, chest roentgenogram, echocardiogram, exercise stress test and 24-hour ambulatory electrocardiographic monitoring were performed in each patient. Adenosine was administered in increasing amounts until an effective dose or a maximal dose of 300 micrograms/kg was reached. In adenosine-sensitive patients verapamil was given orally (3-10 mg/kg/day) for long-term suppression of arrhythmia. Seven patients with a mean age of 10.2 +/- 4.7 years were enrolled in the study. There were five male and two female patients. Six patients had VT, and one patient had frequent ventricular ectopic bests. Arrhythmia originated from the right ventricle in all patients. Adenosine was effective in terminating the arrhythmia in all patients, with a mean effective dose of 183 +/- 75 micrograms/kg. Favorable long-term results (mean follow-up period 17 +/- 8 months) were obtained with verapamil treatment at an average dose of 6.3 +/- 2.2 mg/kg. In conclusion, adenosine can be used effectively for termination of VT originating from the right ventricular outflow tract in children. In these patients verapamil may be considered as the drug of choice for long-term therapy. PMID- 10868196 TI - Gastric emptying time in children with progressive muscular dystrophy. AB - Gastric emptying was evaluated in 11 male children (mean age 8.2 +/- 3.2 years) with progressive muscular dystrophy to detect gastrointestinal smooth muscle involvement. No patient had gastrointestinal symptoms. Gastric emptying studies were performed by using 500 microCi of technetium 99 m sulfur colloid bound to a scrambled egg, and scintigraphic measurements were taken continuously for 60 to 90 minutes. The gastric emptying studies were compared with those of eight male children (mean age 8.2 +/- 2.8 years) without gastrointestinal or muscular disorders. The mean percentage of retention of gastric isotope was significantly greater in the study group than in the control group. These data suggest that dysfunction of the smooth muscle of the upper gastrointestinal tract is detectable in children with progressive muscular dystrophy, even when gastrointestinal symptoms are absent. PMID- 10868197 TI - Delta infection in children with chronic viral hepatitis B. AB - The incidence and clinical features of chronic viral hepatitis B associated with the delta infection were studied in 300 children between one and 14 years in age. Anti-delta was found in 42 (14%) of 300 children with chronic viral hepatitis B, predominantly in patients with chronic active hepatitis who had anti-HBe; exacerbations were found in 11 of 12 children with anti-delta, but in only four of 12 children without these antibodies. These findings confirm the assumption that the process of exacerbation in patients with chronic active hepatitis and anti-HBe may be related to super-infection induced by the delta virus. PMID- 10868198 TI - Clinical risk factors of Hirschsprung-associated enterocolitis. I: Preoperative enterocolitis. AB - Enterocolitis is still the main source of mortality and morbidity in Hirschsprung's disease (HD). Between 1976 and 1993, 79 (26%) of 302 Hirschsprung patients proved to have Hirschsprung-associated enterocolitis (HAEC). Mortality was 7.6 percent (6 patients). HAEC patients, those who died of HAEC and those without HAEC were analyzed for differences in 34 parameters. The length of the aganglionic segment was found not to be a risk factor for HAEC, but early diagnosis and prompt treatment were found to decrease the occurrence of preoperative HAEC. Although we defined HAEC as foul-smelling, explosive diarrhea, some other symptoms and signs, such as abdominal distention on physical examination, vomiting, dehydration, and a history of nonspecific diarrhea were encountered with significant frequency. None of the patients had Down's syndrome. Sepsis was detected in all of the patients who died of HAEC. The severity of HAEC did not increase with the number of attacks of HAEC, and mortality was greater in the first three attacks. Differences in results between some series seemed to be related to differing definitions of HAEC. PMID- 10868199 TI - Clinical risk factors of hirschsprung-associated enterocolitis. II: Postoperative enterocolitis. AB - Among 302 Hirschsprung patients diagnosed between 1976 and 1993, 213 patients who had undergone definitive operations for Hirschsprung's disease (HD) were reviewed for the occurrence of postoperative Hirschsprung-associated enterocolitis (HAEC). The role of 42 parameters were analyzed. We detected 34 patients (14.0%) with postoperative HAEC with no mortality. In nine patients (26.5%) HAEC persisted postoperatively but no effect of preoperative HAEC on postoperative HAEC was detected. The length of the aganglionic segment did not have any effect on HAEC. Down's syndrome was not present in any of our patients with HAEC. Enterocolitis appeared after Swenson, Duhamel and Boley operations in 23, 10 and one patient, respectively. The incidence of HAEC was inversely proportional to age and weight at the time of definitive operation. The presence of colostomy before or during definitive operation had no effect on postoperative HAEC. Complications of definitive operations, including stricture, did not have any effect on HAEC. PMID- 10868201 TI - Yellow nail syndrome in a 10-year-old girl. AB - A 10-year-old girl with yellow dystrophic nails, bronchiectasis, chronic sinusitis and lower-limb lymphedema is presented. The underlying mechanism remains unknown although it has been postulated to be associated with lymphatic abnormalities. To date no causative treatment exists. Our patient was treated with conservative management, including a low-fat diet supplemented with medium chain triglycerides. Moderate improvement in the lymphedema of the lower extremities was observed. To our knowledge this is the first case of yellow nail syndrome to be treated with diet. PMID- 10868200 TI - A Turkish family with 6.7 Kb deletion associated with isolated growth hormone deficiency type 1A. AB - Familial growth hormone deficiency type 1A is an autosomal recessive disease caused by homogenous deletions of both alleles of growth hormone gene 1 (hGH1) in various patterns. The hGH1 gene deletion is an event that probably occurs between the 5' and 3' flanking regions by unequal recombination, and results in deletion of the hGH1 gene in different patterns. Deletions are mostly 6.7 kb and rarely 7.0 kb, 7.6 kb and 45 kb in size. A four-year-old girl diagnosed with growth hormone deficiency syndrome was send to us for further evaluation. DNA samples of the patient, her parents and controls were amplified by polymerase chain reaction (PCR); furthermore, restriction endonuclease analysis was done with Sma I enzyme and the patterns were evaluated. Our gel electrophoresis results show that the gene deletion pattern of the patient represents a homogenous 6.7 kb deletion, while her parents had a heterogeneous 6.7 kb deletion pattern. PMID- 10868202 TI - Chloroquine in idiopathic pulmonary hemosiderosis. A case report. AB - This report describes an 11-year-old boy with idiopathic pulmonary hemosiderosis. His only presenting symptom was severe anemia due to iron deficiency. Idiopathic pulmonary hemosiderosis was diagnosed nine years after the onset of symptoms. During this period many invasive and non-contributory investigations were performed. This report describes the patient's diagnostic problems, clinical features and dramatic improvement with chloroquine (250 mg/day) after failing to respond to megadose methylprednisolone (30 mg/kg). One year later, chloroquine was discontinued. The patient has remained in remission since March 1994. Chloroquine should be used for this life-threatening condition since it is less toxic than other immunosuppressive drugs. PMID- 10868203 TI - Intestinal involvement and vasculopathy in von Recklinghausen's neurofibromatosis. AB - Involvement of the gastrointestinal system and vascular lesions are well-known features of von Recklinghausen's disease, but they are rarely detected in childhood. We report a case of von Recklinghausen's disease with intestinal involvement. The excised right hemicolectomy material of a 16-year-old girl was examined, and diffuse neurofibromatous proliferation with ganglioneuromatous features were observed. Vasculopathy was also seen in some arteries throughout the intestinal segment. PMID- 10868204 TI - Tuberculous otitis media in a four-month-old infant. AB - Tuberculosis of the middle ear is currently a rare disease but still occurs and may cause permanent hearing loss. A four-month-old infant with chronic left-ear drainage was diagnosed with tuberculous otitis media by biopsy examination and PPD positivity without BCG. He was treated successfully with antituberculous therapy. Tuberculosis should be considered in the differential diagnosis of chronic ear infection, especially in young infants. PMID- 10868205 TI - Anomalous origin of the left coronary artery from the pulmonary artery. AB - Anomalous origin of the left coronary artery from the pulmonary artery (ALCA-PA) is a rare form of congenital heart disease. In this report, three cases with this anomaly are described; two patients presented in infancy with heart failure from myocardial ischemia and infarction, while the third was asymptomatic and ALCA-PA was diagnosed during evaluation of a residual murmur after surgery for associated cardiac defects (ventricular septal defect and patent arterial duct). All three cases underwent aorto-pulmonary tunnel repair (Tukeuchi procedure), and to our knowledge two of them are the first infantile cases reported in Turkey. PMID- 10868206 TI - Non-invasive diagnosis and management of coronary arteriovenous fistula. A case report. AB - Coronary arteriovenous fistulas are rare anomalies resulting in abnormal communication between the coronary artery and any chamber of the heart. An asymptomatic patient was referred for evaluation of her murmur. Two-dimensional and color Doppler echocardiographic evaluation revealed an enlarged left main coronary artery. A retrograde, eccentric small jet was found within the right ventricular outflow tract at the pulmonary artery valvular level allowing us to detect the entrance site of the fistula. The diagnosis was confirmed by cardiac catheterization and angiocardiography. Although our case was asymptomatic, the decision to perform cardiac surgery was made because of the aneurysmatic appearance of the left coronary artery. In our opinion, visualization of coronary arteries by two-dimensional echocardiography, together with additional information obtained from the Doppler examination, provides an excellent technique for the noninvasive diagnosis of coronary artery fistula. PMID- 10868207 TI - Peripheral arterial thrombosis in systemic lupus erythematosis and nephrotic syndrome: possible association with protein S deficiency. AB - Arterial thrombosis in systemic lupus erythematosus (SLE) and nephrotic syndrome have been infrequently reported. A 16-year-old boy with SLE and longstanding nephrotic syndrome presented with peripheral arterial thrombosis when his lupus was at an inactive stage. He did not have antiphospholipid antibodies but had low serum antithrombin III and protein S levels. We suggest that the thrombotic event is not related to antiphospholipid antibodies but to nephrotic syndrome and possibly to acquired protein S deficiency. PMID- 10868208 TI - General review of medical genetics. PMID- 10868209 TI - Genetic heterogeneity of Usher syndrome. AB - Progress towards the understanding of the molecular basis of US has been substantial. Nine different loci have been found to be responsible and two have had the specific gene identified. This information is expected to lay the foundation for the eventual development of new treatment strategies. Usher syndrome is the combined loss of both of humans most important two senses and a better understanding of the genes involved should not only help the families with US but will also provide much needed basic information about the hearing and visual systems. PMID- 10868210 TI - Alport syndrome. PMID- 10868211 TI - Waardenburg syndrome. PMID- 10868212 TI - Branchio-Oto-Renal syndrome. PMID- 10868213 TI - The Jervell and Lange-Nielsen syndrome. PMID- 10868214 TI - Treacher Collins syndrome. PMID- 10868215 TI - Non-syndromic dominant DFNA1. PMID- 10868216 TI - DFNA 2, 5, 8, 12. PMID- 10868217 TI - DFNA3. PMID- 10868218 TI - Characterization of autosomal dominant non-syndromic hearing loss loci: DFNA 4, 6, 10 and 13. PMID- 10868219 TI - DFNA7. PMID- 10868220 TI - Mutations in COCH (formerly Coch5b2) cause DFNA9. PMID- 10868221 TI - Sensorineural hearing impairment non-syndromic, dominant DFNA11. PMID- 10868224 TI - Sensorineural hearing impairment: non-syndromic, recessive DFNB2. PMID- 10868223 TI - Mapping of the DFNB1 locus. PMID- 10868222 TI - DFNA15. PMID- 10868225 TI - DFNB3 families and Shaker-2 mice: mutations in an unconventional myosin, myo 15. PMID- 10868226 TI - The PDS gene, Pendred syndrome and non-syndromic deafness DFNB4. PMID- 10868227 TI - Sensorineural hearing impairment, non-syndromic: DFNB5, 6, 7. Homozygosity mapping to localize genes causing autosomal recessive non-syndromic hearing loss. PMID- 10868228 TI - Refined genetic mapping of the autosomal recessive non-syndromic deafness locus DFNB8 on human chromosome 21q22.3. PMID- 10868229 TI - DFNB9 and DFNB12. PMID- 10868230 TI - DFNB11. PMID- 10868231 TI - DFNB15: autosomal recessive non-syndromic hearing loss gene-chromosome 3q, 19p or digenic recessive inheritance? PMID- 10868232 TI - X-linked recessive deafness-dystonia syndrome (Mohr-Tranebjaerg syndrome). PMID- 10868233 TI - DFN2. PMID- 10868234 TI - The ins and outs of X-linked deafness type 3. PMID- 10868235 TI - DFN4: non-syndromic autosomal dominant X-linked sensorineural hearing impairment. PMID- 10868236 TI - X-linked non-syndromic sensorineural deafness: the DFN6 locus. PMID- 10868237 TI - Sensorineural hearing loss associated with the mitochondrial mutations. PMID- 10868238 TI - Histopathology of the inner ear in DFNA9. PMID- 10868239 TI - Histopathologic findings of the inner ears with Alport, Usher and Waardenburg syndromes. PMID- 10868240 TI - Mouse homologues for human deafness. PMID- 10868241 TI - Neurofibromatosis 2. PMID- 10868242 TI - Tumour suppressor genes and head and neck cancer. PMID- 10868243 TI - Chromosomal abnormalities in squamous cell carcinoma of the head and neck. PMID- 10868244 TI - Kallmann syndrome. PMID- 10868245 TI - Cochlear gene therapy. PMID- 10868246 TI - Gene therapy principles and strategies for head and neck cancer. AB - The ongoing preclinical and now human clinical investigation of gene therapy in head and neck cancer and cancer and cancer overall have provided a few foundation points of information. The first point is that viral and nonviral gene therapy has demonstrated efficacy in a variety of animal models and these successes support consideration and evaluation of gene therapy in human clinical trials. Regarding the retrovirus as a vehicle for gene transfer in humans, it appears to be a safe vehicle. There has been no significant short- or long-term toxicity associated with a wide application of retroviral gene therapy in human patients [12]. Regarding adenovirus as a vehicle, there are less numbers and less advanced trials, but the associated toxicities reported thus far have been both transient and relatively minor. Nonviral cationic lipid-mediated gene transfer also appears safe in human patients. These points are significant because they establish the safety foundation for delivering potentially therapeutic genes to humans. Without this safety data, gene therapy would not have a future in the treatment of cancer. At the present Phase I stage of human clinical trial investigation for head and neck cancer, the focus remains on patients with advanced or recurrent incurable cancer. Although this patient population is a standard choice for establishing the safety of novel therapies, the greatest chance of eventual success with currently available gene transfer vehicles and gene therapy strategies will most likely be in those patients with less advanced stages of disease. Another future potential for gene therapy in head and neck cancer may be in combination with surgery or radiation or chemotherapy. At some acceptable ratio of efficacy to toxicity, gene therapy may also prove effective against earlier stage cancer either as a primary or adjuvant therapy. In conjunction with the evolution of molecular diagnostics, gene therapy strategies may provide the means for preventing the malignant progression of premalignant head and neck lesions upon early diagnosis. PMID- 10868247 TI - Six- and ten-item indexes of psychological distress based on the Symptom Checklist-90. AB - Clinicians, provider organizations, and researchers need simple and valid measures to monitor mental health treatment outcomes. This article describes development of 6- and 10-item indexes of psychological distress based on the Symptom Checklist-90 (SCL-90). A review of eight factor-analytic studies identified SCL-90 items most indicative of overall distress. Convergent validity of two new indexes and the previously developed SCL-10 were compared in an archival sample of posttraumatic stress disorder patients (n = 323). One index, the SCL-6, was further validated with archival data on substance abuse patients (n = 3,014 and n = 316) and hospital staff (n = 542). The three brief indexes had similar convergent validity, correlating .87 to .97 with the SCL-90 and Brief Symptom Inventory, .49 to .76 with other symptom scales, and .46 to .73 with changes in other symptom measures over time. These results indicate the concise, easily administered indexes are valid indicators of psychological distress. PMID- 10868248 TI - Does the letter number sequencing task measure anything more than digit span? AB - A total of 102 undergraduate students performed the Letter Number Sequencing (LNS) task in addition to a series of other measures of reading, working memory, motor execution, visuo-spatial memory, and executive functions. Performance on the LNS was uniquely contributed to by reading level, digit span forward and backward, arithmetic, visual spatial learning, and by performance on the Symbol Search subtest of the WAIS-III. The results indicate that while much of the variance on the LNS task is explained by performance on the traditional measures of digit span, additional unique contributions to prediction of LNS performance are provided by measures of processing speed and visual spatial working memory. PMID- 10868249 TI - Errors in scoring objective personality tests. AB - Given the paucity of previous research, we examined the occurrence of scoring error on widely used objective personality tests and examined its possible relation to two factors: scoring procedure complexity (SPC) and commitment to accuracy (CTA). We double-checked the scoring of three tests (MMPI, Beck Depression Inventory, Spielberger State/Trait Anxiety Inventory) across three settings. Each of the tests were misscored at a surprisingly high rate in at least one setting, and some such errors altered major interpretive implications. Tests of higher SPC showed greater error rates, but high CTA greatly reduced the occurrence of error across levels of SPC. Unexpected sources of error were also uncovered, such as commercial computer scoring errors and disagreement in scoring standards among test publishers. Practical suggestions for improving scoring accuracy are offered. PMID- 10868250 TI - A shortened version of the MMPI-2. AB - A psychometrically sound method of prorating scores from a shortened version of the MMPI-2 is presented to approximate the full-scale raw scores on the basic scales. After a brief review of the history of short versions of the original MMPI and their strengths and weaknesses, justifications for developing and publishing this new version are offered. In spite of the risk of abuse by harassed and over-worked clinicians, there are cogent reasons to make this set of procedures available to practitioners and research investigators. These procedures were devised on the data from the 2,600 men and women in the original MMPI-2 restandardization sample and cross-validated on a sample of 632 test records from a psychiatric inpatient service. The dependability of estimated single raw scores as well as of the patterning of the prorated profile patterns is explored. PMID- 10868251 TI - MMPI-2 clinical scale differences between dysthymia and major depression. AB - Though dysthymia is considered less severe and more chronic than major depressive disorder, it is unclear whether the two disorders are truly different. In this study, MMPI-2 scales of 21 patients with dysthymia and 30 patients with major depressive disorder were compared. The average scores on Scales 2, 4, 6, 7, and 8 were in the clinical range for both groups. However, sizable differences between the two groups were found for Scale 1 and Scale 3. Smaller but reliable differences were found for Scale 2 and mean clinical scale T score with major depressives scoring higher on all of these measures. Results indicate that not only is major depressive disorder more severe than dysthymia, but also contains more physical/somatic symptoms than dysthymia. PMID- 10868252 TI - Reliability of the WAIS-III subtests, indexes, and IQs in individuals with substance abuse disorders. AB - Reliability of the WAIS-III for 100 male patients with substance abuse disorders was determined. Means for age and education were 46.06 were (SD = 8.81 years) and 12.70 years (SD = 1.51 years). There were 63 Caucasians and 37 African Americans. Split-half coefficients for the 11 subtests (Digit Symbol-Coding, Symbol Search, and Object Assembly were omitted) ranged from .92 for Vocabulary and Digit Span to .77 for Picture Arrangement. The median subtest reliability coefficient was .86. Composite reliabilities were excellent for the Indexes (.94 to .95) and IQs (.94 to .97), with all coefficients > or = .94. Using the Fisher z test to compare correlation coefficients from independent samples, none of the reliability estimates differed significantly from those reported for the WAIS-III standardization sample. Similar findings emerged when reliabilities were determined separately for Caucasian and African American participants. PMID- 10868253 TI - Estimation of Wechsler Adult Intelligence Scale-III index scores with the 7 subtest short form in a clinical sample. AB - A 7-subtest short form of the Wechsler Adult Intelligence Scale-III (WAIS-III) previously demonstrated good comparability in estimating Full Scale and Verbal IQ summary scores, with adequate comparability in estimating Performance IQ. In a mixed clinical sample of 295 patients, the current study assessed the equivalence of the index scores generated from the full and prorated WAIS-III. The results revealed correlations corrected for redundancy of .90, .86, .87, and .75 for the Verbal Comprehension (VCI), Perceptual Organization (POI), Working Memory (WMI), and Processing Speed (PSI) indexes, respectively. Although the 7-subtest short form of the WAIS-III was not designed to estimate index scores, adequate estimates are viable for VCI, POI, and WMI when the goal is to obtain group, rather than individual, data points. PMID- 10868254 TI - Comparisons among the Holden Psychological Screening Inventory (HPSI), the Brief Symptom Inventory (BSI), and the Balanced Inventory of Desirable Responding (BIDR). AB - Issues of reliability, item latent structure, and faking on the Holden Psychological Screening Inventory (HPSI), the Brief Symptom Inventory (BSI), and the Balanced Inventory of Desirable Responding (BIDR) were examined with a sample of 300 university undergraduates. Reliability analyses indicated that scales from all inventories had acceptable internal consistency. Confirmatory item principal component analyses supported the structures and scoring keys of the HPSI and the BIDR, but not the BSI. Although all inventories were susceptible to faking, validity indices of the HPSI and the BIDR could correctly classify over two thirds of test respondents as either responding honestly or as faking. PMID- 10868255 TI - Brief psychosocial assessment of a clinical sample: an evaluation of the Personal Problems Checklist for Adults. AB - In light of the requirements for managed health care organizations to use assessment instruments that are psychometrically sound, cost and time efficient, and theoretically useful, the present study examined the psychometric properties of one such potential instrument, the Personal Problems Checklist for Adults (PPCA). Designed to measure problems in 13 areas of everyday functioning, the PPCA along with the Brief Symptom Inventory were completed by 132 individuals in an outpatient drug rehabilitation program. Counselor ratings on the Adjective Check List were also obtained. Results clearly showed that personal problems as measured by the PPCA were related to self-reported psychological symptoms and to perceptions by their counselors. The PPCA proved to have good psychometric properties and warrants greater attention by testing psychologists given its potential to meet criteria set forth by managed health care. PMID- 10868256 TI - Factor structure of the Reitan-Indiana Neuropsychological Battery for Children. AB - The Reitan-Indiana Neuropsychological Battery (RINB) was administered to a sample of children referred for educational and behavioral problems (N = 130). Subtest scores were standardized by age at 1-year intervals (6, 7, and 8 years). A principal components analysis (PCA) with promax rotation of 18 subtest scores produced a five-factor solution. Factor 1 emphasizes tactile/spatial functions, Factor 2 emphasizes concept formation and visual/spatial abilities, Factor 3 reflects motor strength, Factor 4 emphasizes sensory perception, and Factor 5 reflects motor speed. Principal factor analysis (PFA) of these data was performed to permit comparison with the PCA solutions. The PFA and PCA solutions were similar and major conclusions about factor structure were consistent. PMID- 10868257 TI - Patient safety and clinical engineering. PMID- 10868258 TI - Ultra-fast MRI improves accuracy of prenatal ultrasound. PMID- 10868259 TI - Managing the airwaves: new FCC rules for wireless medical telemetry. PMID- 10868260 TI - Requirements engineering: the key to designing complex medical systems. AB - A variety of business systems, clinical work systems, instrumentation systems, information systems, infrastructure systems, and management systems interact to make the modern healthcare facility work. The key to designing for such a system is systems engineering, a skill often little appreciated among clinical engineers. At the heart of systems engineering is requirements engineering and management (REAM), which is defined as "the process of discovering, documenting and managing systems requirements." The principal activities of REAM include eliciting, understanding, negotiating, describing, validating, and managing system requirements. When REAM is done improperly, the resulting system will be satisfactory only if chance intervenes. Well-done REAM is likely to bring the project in on time, under budget, and at full performance. PMID- 10868261 TI - Effect of adaptive motion-artifact reduction on QRS detection. AB - Motion artifact resulting from electrode and patient movement is a significant source of noise in ECG, EEG, EMG, and impedance pneumography recording. Noise resulting from motion is particularly troublesome in ambulatory ECG recordings, such as those made during Holter monitoring or stress tests, because the bandwidth of the motion artifact overlaps with the ECG signal bandwidth. The authors investigated the effect of an adaptive motion-artifact removal algorithm on the performance of a standard QRS detector. They made four ECG recordings on each of the three subjects while manually generating artifact. Adaptive noise removal was applied to the ECG signal using a skin-stretch signal as the noise reference. Adaptive noise removal reduced the number of false QRS detections in the records from 380 to 104, for an average reduction in false detections of 72.6%. False-detection reductions for individual records ranged from 12% to 93%. PMID- 10868262 TI - Dynamic thermal responses of five commercially available infant radiant warmer systems. AB - Five commercially available infant radiant warmer systems were characterized through radiant-energy measurements at the mattress surface under identical environmental conditions. Average irradiance levels for each warmer were then calculated. In addition, all systems were thermally evaluated for warm-up time using the International IEC standard black anodized aluminum disk simulator. Warm up time and radiant-energy bed mapping tests were conducted in manual mode. To assess thermal performance under dynamic conditions, skin and core temperatures were measured using a patient simulator test-load device. Dynamic patient simulator response-time data and side-wall temperature data are provided. The radiative heat contribution from the warmer side walls on patient simulator core and skin temperatures was demonstrated. Extreme differences in warmer performances and energy to the mattress surface are documented. Differences in infant radiant warmer performances are principally due to heater-element composition, reflector design, and heater-to-mattress distance. PMID- 10868263 TI - The central equipment pool, an opportunity for improved technology management. AB - A model for a central equipment pool managed by a clinical engineering department has been presented. The advantages to patient care and to the clinical engineering department are many. The distribution of portable technology that has been traditionally managed by the materials management function is a logical match to the expanding role of clinical engineering departments in technology management. Accurate asset management tools have allowed us to provide reliable measures of infusion pump utilization, permitting us to predict future needs as programs expand. Thus we are more actively involved in strategic technology planning. The central equipment pool is an excellent opportunity for the clinical engineering department to increase its technology management activities. PMID- 10868264 TI - Improving ECG trace quality. PMID- 10868265 TI - Routing fundamentals, Part Two: Distance vector routing. PMID- 10868266 TI - Use your networking skills to connect at conferences. PMID- 10868267 TI - Detection of cellular DNA cleavage using non-proofreading thermostable DNA polymerases. PMID- 10868268 TI - Using BbvI and related restriction enzymes to cut DNA at otherwise inaccessible sites. PMID- 10868269 TI - Fluorescein PAGE analysis of microsatellite-primed PCR: a fast and efficient approach for genomic fingerprinting. PMID- 10868270 TI - Use of dialysis to prepare bacterial DNA suitable as PCR template. PMID- 10868271 TI - Single-primer PCR procedure for rapid identification of transposon insertion sites. PMID- 10868272 TI - Extraction and quantitative determination of callose from Arabidopsis leaves. PMID- 10868273 TI - Analysis of quorum sensing activity in staphylococci by RP-HPLC of staphylococcal delta-toxin. PMID- 10868274 TI - Method to decrease the titers of contaminating helper adenovirus during the production of recombinant adeno-associated virus. PMID- 10868275 TI - The sequence manipulation suite: JavaScript programs for analyzing and formatting protein and DNA sequences. PMID- 10868276 TI - PCR- and ligation-mediated synthesis of split-marker cassettes with long flanking homology regions for gene disruption in Candida albicans. AB - Because Candida albicans is a diploid organism, two consecutive steps of gene disruption are required to generate a gene knock-out. The same marker (URA3) is often used for disruption of both copies of the gene. This is possible because, after the first round of disruption, homologous recombination between direct repeats flanking the URA3 marker and the subsequent counterselection allow for the efficient recovery of Ura- revertants. Unfortunately, the URA-blaster disruption cassette cannot be used in a PCR-based disruption approach. The hisG repeats flanking the URA3 gene in the disruption cassette anneal to one another during PCR and thereby prevent amplification of the complete cassette. We explored the use of transformation based on split-marker recombination to circumvent this problem. To avoid any cloning steps and to retain the advantage of long flanking regions for disruption, we combined this with a PCR- and ligation-mediated approach for generating marker cassettes. We used this approach to disrupt the C. albicans FAL1 (ATP-dependent RNA helicase) gene. Long 5' and 3' FAL1-specific regions were amplified by PCR and individually ligated to a URA blaster cassette. The resulting ligation reactions were used separately as templates to generate two FAL1 disruption cassettes with overlapping URA3 marker regions. Simultaneous transformation with both overlapping disruption cassettes yielded efficient disruption of one FAL1 allele. PMID- 10868277 TI - Tissue factor mRNA quantitation without prior monocyte isolation. AB - Monocyte tissue factor (TF) quantitation evaluates the involvement of coagulation processes in many diseases. However, technical difficulties, such as blood sampling of cells representative of the whole intravascular pool, cell isolation, protein quantitation or activity assessment, hinder reliable evaluation of TF expression by activated monocytes. Early determination of such activation can be achieved through TF mRNA quantitation by RT-PCR and sensitive product detection, such as automated electrophoresis of fluorescently labeled products. Although it is very sensitive, this method has its limitations. It needs to be standardized using other mRNA that display two main characteristics: the absence of upregulation during inflammation and similar levels of expression when compared with the target mRNA. Widely used standardization housekeeping genes such as HLA or GAPDH genes only meet the former requirement. We demonstrate here that CD11b gene expression meets both conditions. Moreover, because of its specific expression in myelomonocytic cells, it is possible to avoid further monocyte purification from a regular mononuclear cell preparation. A rapid, sensitive, specific and accurate way to evaluate monocyte TF expression is described in this paper. PMID- 10868278 TI - Determination of transgene repeat formation and promoter methylation in transgenic plants. AB - The integration of transgenes into a plant host genome following Agrobacterium tumefaciens-mediated or direct transformation may occur as a single copy or in the form of tandem repeats. The latter has been associated with promoter methylation and silencing of transgenes. Thus, the early screening of such transgenic plants is desirable for ruling out future repeat-dependent transgene instability. We developed a simple PCR-based method in which primer pairs were specifically designed so that amplifications could only be obtained if the transgene was present in the form of multiple inserts in a transgenic line. The method was established using 35S-rolC transgenic aspen lines showing morphologically visible transgenic silencing. Later, it was possible to screen independent transgenic lines showing no visible marker gene expression. Furthermore, a method was developed in which positive PCR amplification was indicative of promoter methylation. The results were consistent and reproducible across different independent transgenic lines. The methods were quick, reliable, consistent and reproducible, and can be useful for routine screening of transgene silencing in lines derived from many different systems. PMID- 10868279 TI - Transient host range selection for genetic engineering of modified vaccinia virus Ankara. AB - Recombinant vaccinia viruses are extremely valuable tools for research in molecular biology and immunology. The extension of vaccinia vector technology to replication-deficient and safety-tested virus strains such as modified vaccinia virus Ankara (MVA) have made this versatile eukaryotic expression system even more attractive for basic and clinical research. Here, we report on easily obtaining recombinant MVA using stringent growth selection on rabbit kidney RK-13 cells. We describe the construction and use of new MVA vector plasmids that carry an expression cassette of the vaccinia virus host range gene, K1L, as a transient selectable marker. These plasmids allow either stable insertion of additional recombinant genes into the MVA genome or precisely targeted mutagenesis of MVA genomic sequences. Repetitive DNA sequences flanking the K1L gene were designed to remove the marker gene from the viral genome by homologous recombination under nonselective growth conditions. The convenience of this new selection technique is demonstrated by isolating MVA recombinants that produce green fluorescent protein and by generating MVA deletion mutants. PMID- 10868280 TI - Colorimetric detection of the tuberculosis complex using cycling probe technology and hybridization in microplates. AB - Cycling probe technology (CPT) is a simple signal amplification method for the detection of specific target DNA sequences. CPT uses a chimeric DNA-RNA-DNA probe that is cut by RNase H when bound to its complementary target sequence. In this study, a hybridization assay was developed to detect biotinylated CPT products that result from the amplification of a Mycobacterium tuberculosis complex sequence. The chimeric probe was specifically designed to avoid the formation of secondary structures. The chosen capture probe was perfectly complementary to and was the same size as OL2, one of the two CPT products. The assay was based on the observation that a long sequence, such as the initial probe, was destabilized when bound to a small capture probe as a result of steric hindrance. The capture probe preferentially bound OL2 rather than the long initial probe. We added a prehybridization step with a helper DNA to enhance this discrimination between the two sequences. Colorimetric detection was performed using a peroxidase streptavidin conjugate. After optimization, the non-isotopic hybridization assay allowed the detection of around 10 amol of target DNA. Besides being faster and easier to perform, this detection method was compared to electrophoresis separation and gave similar results. PMID- 10868281 TI - Application of DNA topoisomerase-activated adapters to riboprobe synthesis. AB - Topoisomerase-activated adapters for rapid incorporation of the T7 promoter into PCR products were made by hybridizing synthetic oligonucleotides and activating vaccinia DNA topoisomerase I. The adapters were used to incorporate the T7 promoter sequence into PCR products amplified from cDNA and genomic DNA. Modified PCR products were used as templates to synthesize digoxigenin-labeled sense and cRNA probes by in vitro transcription with phage T7 RNA polymerase. The red/green cones were labeled by the antisense probe, but no specific signal was produced by the sense probe. These results demonstrate that topoisomerase-activated adapters provide a powerful and convenient tool for the rapid modification of PCR products. PMID- 10868282 TI - Detection of proteolytic activity by fluorescent zymogram in-gel assays. AB - Proteases are involved in the regulation of many biological functions. This study describes a novel method for detecting protease activity by fluorescent zymogram in-gel protease assays, using SDS polyacrylamide gels copolymerized with a peptide-MCA (4-methyl-coumaryl-7-amide) substrate. This method allows simultaneous determination of protease cleavage specificity and molecular weight. Trypsin was electrophoresed in SDS polyacrylamide gels copolymerized with Boc-Gln Ala-Arg-MCA, the gel was then incubated in assay buffer, and trypsin cleavage of the peptide-MCA substrate generated fluorescent AMC (7-amino-4-methyl-coumarin), which was subsequently detected under UV transillumination. Chymotrypsin activity was detected in gels copolymerized with Suc-Ala-Ala-Pro-Phe-MCA substrate. Selective detection of these proteases was demonstrated by the absence of trypsin activity in gels containing the chymotrypsin substrate, and the lack of chymotrypsin activity in gels containing the trypsin substrate. Detection of proteolytic activity from secretory vesicles of adrenal medulla (chromaffin granules) was observed with the trypsin substrate, Z-Phe-Arg-MCA, but not with the chymotrypsin substrate. Overall, this sensitive fluorescent zymogram in-gel protease assay method can be used for rapid determination of protease cleavage specificity and enzyme molecular weight in biological samples. This assay should be useful for many research disciplines investigating the role of the many proteases that control cellular functions. PMID- 10868283 TI - Strategies for searching sequence databases. AB - We provide a detailed overview of the choices inherent in performing a sequence database search, including the choice of algorithm, substitution matrix and gap model. Each of these choices has implications that can be described as restrictions on the underlying model of sequence evolution, the expected degree of divergence between the query sequence and the database sequences (if one uses an evolutionary based matrix), as well as the sensitivity and selectivity of the search. We conclude with a series of recommendations for researchers performing these searches based on our experience and literature studies. PMID- 10868284 TI - DNAssist, a C++ program for editing and analysis of nucleic acid and protein sequences on PC-compatible computers running Windows 95, 98, NT4.0 or 2000. AB - The size of sequence databases, the rapid rate of generating sequence data and the need to routinely construct recombinant DNA molecules in molecular biological research necessitate the frequent handling and analysis of nucleotide and protein sequences on a computer. Although several console or Web-based utility programs are available, the application of these programs generally requires reformatting the data when proceeding from one such program to the next. The acquisition of elaborate, integrated program suites is financially prohibitive to smaller laboratories. Here, we report the development of DNAssist, a shareware program for editing and analysis of nucleic acid and protein sequences. It was developed as a multiple-document interface program--similar to a word processor--where sequences are entered, edited and analyzed in a single integrated environment. DNAssist can calculate the physicochemical properties of a sequence, convert between nucleic acid and protein sequences, translate DNA in multiple frames, identify open reading frames and locate ambiguous sequence patterns allowing gaps and mismatches. DNAssist also performs restriction enzyme and transcription factor-binding site analyses of DNA sequences, the multiple alignment of nucleic acid and protein sequences and the analysis of DNA sequences for nucleosome positioning sites. PMID- 10868285 TI - Useful Microsoft Word Macros for molecular biologists and protein chemists. AB - Biologists today make extensive use of word processing programs for the production of research reports, literature reviews and grant proposals. Frequently, such programs become the default platform for viewing and the later publication of protein and nucleic acid sequence data. Thus, researchers often switch between their word processor and more specialized programs designed to analyze protein and nucleic acid sequences. It would be more convenient to perform these simple sequence analyses using the word processor without switching to another program. The focus here is on the use of the Visual Basic programming language, which is built into all recent versions of Microsoft Word to generate surprisingly complex and useful macros that can conveniently analyze several important features of protein and nucleic acid sequences. The standard Word interface can also be easily modified to display and run these macros from a pull down menu. Several examples of this approach are provided. PMID- 10868286 TI - Experimental design optimization of filamentous phage transfection into mammalian cells by cationic lipids. AB - A previous study showed that filamentous phage could be efficiently transfected into mammalian cells in the presence of the cationic lipid Transfectam. In the present study, we used an experimental plan based on a uniform network (Doehlert) matrix to estimate optimal transfection conditions in two different cell lines, CHO and Cos-7. Using the cationic lipid RPR120535b as a model, we show that optimal conditions can be determined much more readily than with standard response curves. Under optimal conditions as analyzed by FACS, up to 60% of Cos-7 and 50% of CHO cells can be transfected. Furthermore, a comparison of different lipids (Transfectam, RPR120535b, TC1-12 and GAP-DLRIE/DOPE) suggests that lipids with multiple amine groups are more efficient for the transfection of filamentous phage. PMID- 10868287 TI - Purification and reconstitution of an integral membrane protein, the photoreaction center of Rhodobacter sphaeroides, using synthetic sugar esters. AB - Detergents are indispensable reagents for the extraction and solubilization of integral membrane proteins, but their removal from a reconstituted phospholipid protein complex is usually desirable. In this paper, we describe a novel method in which the synthetic sugar esters 6-O-octanoyl-beta-D-glucose (OG) or 6-O octanoyl-beta-D-mannose (OM) are used as detergents for both the isolation and the rapid reconstitution of the photosynthetic reaction center protein of Rhodobacter sphaeroides. Following solubilization of the reaction center with OG or OM and reconstitution of this protein in liposomes, a convenient removal of these detergents was achieved within less than two hours by hydrolytic cleavage of the sugar esters using immobilized lipases. Best results were achieved with lipase from Bacillus sp. immobilized on silica gel. PMID- 10868288 TI - Xplore mRNA assays for the quantification of IL-1 beta and TNF-alpha mRNA in lipopolysaccharide-induced mouse macrophages. AB - Because the accurate measurement of a number of cytokine mRNA transcripts provides valuable knowledge about cytokine gene regulation, we have developed the Xplore assay for the quantification of cytokine mRNA. This microplate-based assay is rapid (under four hours), quantitative over three orders of magnitude and carries no risk of false-positive values from contamination with amplified target. Here, we describe the use of Xplore assays to measure the steady-state mRNA levels of TNF-alpha and IL-1 beta produced by mouse WEHI and J774 macrophage like cell lines. PMID- 10868289 TI - New approaches to antigen delivery. AB - An improved understanding of immunologic events associated with immunization, the identification of promising new antigens, and an increased capacity to generate these antigens through chemical and biotechnology methods have led to many new vaccine opportunities. Inappropriate antigen exposure, however, can result in unwanted outcomes, such as incomplete protection, allergic reactions, autoimmunity, infection, or even tolerization. Thus, proper antigen delivery is critical for achieving the desired outcome. A number of vaccination approaches have now been described with varied degrees of success. The relative success of these approaches can be correlated with antigen delivery to specific presentation cells and stimulation of the immune system at sites where protective immunity is most appropriate. In addition, a greater understanding of mechanisms involving cells and effector molecules in the events of immunity may allow for improved possibilities for initiating, augmenting, and maintaining the response to a delivered antigen. This review provides insights into the various strategies currently being explored to optimize antigen delivery and the immune response to that antigen. PMID- 10868290 TI - Drug delivery systems for the treatment of restenosis. AB - Attempts to achieve revascularization of coronary arteries blocked by atherosclerotic plaques are hampered by restenotic hyperproliferative response of the treated vessels. The uniform failure of clinical trials using systemic therapies to prevent restenosis has prompted development of methods for arterial drug delivery systems. This review describes technologies of polymeric-based, perivascular, and intraluminal drug and gene delivery systems. The critical assessment of controversies including drug and vehicle type, dose and release rate, and preclinical validation is reviewed. PMID- 10868291 TI - Cytochemical nucleic acid research during the twentieth century. PMID- 10868292 TI - Inositide-specific phospholipase C signalling in the nucleus. PMID- 10868293 TI - Nuclear inositides. PMID- 10868294 TI - Phosphoinositide 3-kinase is associated to the nucleus of HL-60 cells and is involved in their ATRA-induced granulocytic differentiation. PMID- 10868295 TI - Nuclear envelope signaling-role of phospholipid metabolism. PMID- 10868296 TI - Intranuclear domains involved in inositol lipid signal transduction. PMID- 10868297 TI - Molecules that inhibit T-cell functions: cytochemical localization and shuttling. AB - Adaptive immune responses to antigens are mediated by specific receptors expressed on B cells (BCR's) and T cells (TCR's). Effector cells and memory cells are produced following a proliferative wave that accounts for clonal expansion. If not down-regulated, clonal expansion might lead to uncontrolled lymphoproliferation that would be harmful for the organism. Several mechanisms that account for the down-sizing of activated lymphocyte clones are briefly reviewed here. We next consider in detail one such mechanism that deals with the functional characterization and the immunocytochemical localization of two T-cell inhibitory molecules, namely the Cytotoxic T Lymphocyte Antigen-4 (CTLA-4) and the HP-F1 antigen, both present in all T lymphocytes. CTLA-4 and HP-F1 inhibit CD4+ T-helper cell proliferation and the lytic ability of CD8+ T-cytotoxic cells in non-specific and in antigen-specific cytolytic assays. Interestingly, a clonal distribution exists as for the ability of CTLA-4 and HP-F1 to inhibit T-cell functions. In resting and activated T cells, both molecules are largely confined in the endosomal compartment, as shown by immunofluorescence analyses. However, upon interaction of T cells with Antigen-Presenting Cells (APC's) or with target cells that must be killed, CTLA-4 molecules are transported to the plasma membrane, at the site of cell-to-cell contact where, following interaction with ligands, they trigger inhibitory signals. PMID- 10868298 TI - [Pharmacokinetics of panipenem in neonates and dosage recommendation]. AB - The pharmacokinetics of panipenem/batamipron (Carbenin; PAPM/BP) was studied in 17 neonates of the postconceptional age (PCA) of 25.6 to 43.1 weeks. PAPM/BP was administered at 10 mg/kg to 20 mg/kg every 12 hours over 60 minutes by intravenous infusion. Blood samples were obtained just prior to the infusion, one or two hours and six hours after the infusion. All the data for the 85 serum PAPM concentrations were analyzed by one-compartment model using a nonlinear mixed effect model (NONMEM). The pharmacokinetic parameters in these population are given below: CLPAPM; 0.239 +/- 0.206 (L/hr), VdPAPM; 0.97 +/- 0.80 (L), Half Life; 3.1 +/- 0.5 (hr). Half life in the patients with PCA < 34 (2.66 +/- 0.44 hr) were significantly lower (p < 0.001) than that with PCA > or = 34 (3.39 +/- 0.23 hr). Our results suggest that postnatal alterations in the PAPM excretion are related to maturational changes in the renal function and that we should consider the values of PCA when determining the initial PAPM/BP dosing regimen in neonates. We conclude that the dosage of 10 mg/kg to 20 mg/kg every 12 hours are adequate treatment for neonatal infectious disease. PMID- 10868299 TI - [In vitro interaction of piperacillin and imipenem/cilastatin combined with aminoglycosides against Pseudomonas aeruginosa]. AB - The in vitro interactions of piperacillin (PIPC) and imipenem/cilastatin (IPM/CS) combined with 5 kinds of aminoglycosides (gentamicin, tobramycin, amikacin (AMK), isepamicin and netilmicin) were investigated against IPM/CS-susceptible (MIC of IPM/CS was < or = 3.13 micrograms/ml) and IPM/CS-resistant (MIC of IPM/CS was > or = 12.5 micrograms/ml) Pseudomonas aeruginosa. The following results were obtained. 1. In the checkerboard dilution studies, the combinations of PIPC with aminoglycosides showed synergistic effect for more than 50% of the each 54 strains of IPM/CS-susceptible and IPM/CS-resistant P. aeruginosa. The synergistic/additive effects of PIPC with aminoglycosides were demonstrated for all tested strains. 2. In the checkerboard dilution studies, the combinations of IPM/CS with aminoglycosides showed no antagonism against any strains. The synergistic effects of IPM/CS with aminoglycosides were demonstrated for 0 to 14.8%, and these values were smaller than the combinations of PIPC with aminoglycosides. 3. Corresponding to the results of checkerboard dilution studies, the combination of PIPC with AMK was more effective than the combination of IPM/CS with AMK on the killing curve for IPM-resistant P. aeruginosa. In conclusion, PIPC showed the synergistic effects in combinations with aminoglycosides against IPM/CS-resistant P. aeruginosa. These results suggest that the combination therapies of PIPC with aminoglycosides are useful for the clinical treatment of serious infections due to P. aeruginosa. PMID- 10868300 TI - [Comparative studies on activities of antimicrobial agents against causative organisms isolated from patients with urinary tract infections (1998). I. Susceptibility distribution]. AB - The frequencies of bacterial isolation and susceptibilities to antimicrobial agents were investigated on 538 bacterial strains isolated from patients with urinary tract infections (UTIs) in 9 hospitals during the period of June 1998 to May 1999. Of the above bacterial isolates, Gram-positive bacteria accounted for 30.3% and Gram-negative bacteria accounted for 69.7%. Susceptibilities of several isolated bacteria to antimicrobial agents were as follows; against Enterococcus faecalis isolated from patients with UTIs, vancomycin (VCM), ampicillin (ABPC) and imipenem (IPM) had strong activities. Among E. faecalis strains, those with low susceptibilities to all drugs have increased in 1998, compared with those in 1997. VCM showed the highest activity against MRSA isolated from patients with UTIs. The MICs of VCM for all 34 strains were equal to or lower than 2 micrograms/ml. Arbekacin (ABK) was also active against MRSA with the MIC90s of 2 micrograms/ml. Against Escherichia coli and Klebsiella pneumoniae, all drugs except penicillins were active. Particularly, meropenem (MEPM) showed the highest activity with the MICs of 0.125 micrograms/ml or below. Almost all the drugs except minocycline (MINO) showed high activities against Proteus mirabilis. Against Pseudomonas aeruginosa, all drugs were not so active, with the MIC90s of 16 micrograms/ml or above. MEPM, IPM and gentamicin (GM) showed high activities against Serratia marcescens. Generally, it seemed that resistant strains of S. marcescens had decreased since 1996. PMID- 10868302 TI - [A study on the mechanism of seizures associated with the drug interaction between quinolones and anti-inflammatory drugs. A comparative study and the structure-activity relationship of anti-inflammatory drugs]. PMID- 10868301 TI - [Comparative studies on activities of antimicrobial agents against causative organisms isolated from patients with urinary tract infections (1998). II. Background of patients]. AB - Clinical background was investigated on 449 patients with urinary tract infections (UTIs) from whom 591 bacterial strains were isolated in 9 hospitals during the period from June, 1998 through May, 1999. About distribution of age and sex of patients and type of infections, among males, patients less than 50 years old were few, and uncomplicated UTIs without indwelling catheters was most frequent. Among females, patients less than 20 years old were few, and uncomplicated was most frequent. Escherichia coli was the most frequently isolated in uncomplicated UTIs, and the higher the ages of patients, the higher were became the isolation frequencies of Enterococcus faecalis, Proteus spp. and Klebsiella spp. In complicated UTIs with indwelling catheters and without indwelling catheters, the types of pathogens had no relation with ages. The complication of infections had decreased E. coli but that had increased Proteus spp., Pseudomonas aeruginosa and Staphylococcus aureus. Until last year, use of antibiotics had decreased pathogens isolated from patients with uncomplicated UTIs drastically in our study. But, pathogens isolated after antibiotics had increased in 1998. As for surgical procedures and types of causative organisms in UTIs, E. faecalis were more isolated when surgical procedures were experienced, and E. coli were more isolated when they were not in uncomplicated and complicated UTIs without indwelling catheters. In complicated UTIs with indwelling catheters, types of causative organisms had no relationship with surgical procedures. PMID- 10868303 TI - [Study of effects and mechanisms of macrolide as biological response modifier for lung cancer treatment]. PMID- 10868304 TI - Bioavailability and bioequivalence: average, population and/or individual. PMID- 10868305 TI - PhRMA perspective on population and individual bioequivalence. AB - The Food and Drug Administration (FDA) issued a second-draft guidance in August 1999 on the subject of in vivo bioequivalence, which is based on the concepts of individual and population bioequivalence (IBE and PBE, respectively). The intention of this guidance is to replace the 1992 guidance that requires that in vivo bioequivalence be demonstrated by average bioequivalence (ABE). Although the concepts of population and individual bioequivalence are intuitively reasonable, a detailed review of the literature has not uncovered clinical evidence to justify the additional burden to the innovator and generic companies as well as the consumer that the new guidelines would impose. The criteria for bioequivalence described in the draft guidance employ aggregate statistics that combine information about differences in bioavailability between formulation means and differences in bioavailability variation of formulations between and within subjects. The purely technical aspects of the statistical approach are reasonably sound. However, PhRMA believes that important operational issues remain that need to be resolved before any changes to current practice are implemented. PhRMA believes that the ideals of prescribability and switchability are intuitively reasonable, but it is uncertain of the extent to which the proposed guidance can achieve these goals. It is not clear whether the attainment of such goals is necessary in the evaluation of bioequivalence given the role this plays in drug development, and the lack of clinical evidence argues against a pressing need to change current practice. PhRMA is concerned that the trade-off offered by the aggregate criteria may ultimately represent more harm than good to the public interest. PhRMA recommends more rigorous evaluation of methods based on two-way crossover designs before moving to methods that require more complex designs. One such method is identified herein and contains procedures for estimating prescribability and switchability. The possibility of a phase-in or trial period to collect replicate crossover data to further evaluate IBE and PBE and possibly allow market access based on these criteria as they are being evaluated has been proposed. PhRMA believes this is unprecedented and will offer little additional information beyond that which can be obtained by simulation or has already been collected by the FDA. Simulation studies have the advantage of allowing evaluation of the sensitivity of various procedures to represent the data patterns as created within the simulation. Operating characteristics by which proposed criteria can be adequately judged have not yet been defined. The limitations of ABE for highly variable drugs and narrow therapeutic drugs are well appreciated and may be addressed by means other than a wholesale change in the current criteria. PMID- 10868306 TI - Update to the PhRMA perspective on population and individual bioequivalence. PMID- 10868307 TI - Drug labeling and risk perceptions of teratogenicity: a survey of pregnant Canadian women and their health professionals. AB - There is a general perception that medicinal drugs are not safe in pregnancy despite the fact that fewer than 30 drugs have been shown to cause major malformations in humans. A large number of women need medications in pregnancy to treat pregnancy-induced conditions, acute illnesses, and chronic diseases. The objectives of this study were the following: (1) to characterize the perception of teratogenic risk by pregnant women and their partners and by health professionals and (2) to examine the most reassuring way to present data on a drug for nausea and vomiting of pregnancy that has been proven to be safe to the fetus. A convenience sample of pregnant Canadian women and their partners, pharmacists, nurses, physicians, and hospital workers were asked to choose the "safest" among four drugs by statements describing their safety. Although the text of all four was similar, the title and narrative were modified to be more or less "reassuring" by the use of more or less terms such as malformations and abnormalities. Health professionals rated the teratogenic risk significantly lower than the parents, but even they rated the drugs as not safe, despite a scientifically reassuring text. Sixty percent of the 240 participants, regardless of their perception of teratogenic risk, believed the four drugs were of similar risks. However, in the other 40%, the less "reassuring" text led to higher teratogenic perception, and the more reassuring options tended to decrease the false perception of teratogenic risk. It was concluded that in general, four different versions of reassuring text describing a scientifically proven safe drug in pregnancy did not lead expecting parents to believe they were safe. Among those who did not rank the four drugs as having equal safety/risk, the less "reassuring" text led to a higher perception of teratogenic risk. Even health professionals reading the labels describing safe drugs rated them as unsafe. Presently, the perception of teratogenic risk is strong even for safe drugs and is difficult to change even with evidence-based facts. PMID- 10868308 TI - Pediatric pharmacodynamics of midazolam oral syrup. Pediatric Pharmacology Research Unit Network. AB - In this study, the authors evaluate the pharmacodynamics, safety, and acceptability of a new cherry-flavored oral syrup formulation of midazolam. This randomized, double-blind, parallel-group, dose-ranging clinical trial of oral midazolam was conducted at seven U.S. health care institutions focused on pediatric clinical pharmacology research (i.e., the PPRU Network). Pediatric patients (n = 85, ages 6 months through 15 years) underwent invasive procedures and were randomized to a single oral dose of midazolam syrup (0.25, 0.5, or 1.0 mg/kg). Patient taste acceptability of midazolam syrup was evaluated at the time of oral administration. Pharmacodynamic measurements included (1) sedation score using a 5-point scale at baseline and 10-, 20-, and 30-minute postdose intervals and (2) anxiety score using a 4-point scale at the time of separation from caretakers and, when applicable, at the time of mask anesthetic induction. Midazolam and alpha-hydroxymidazolam plasma concentrations were measured at all pharmacodynamic measurement time points. Adverse events were monitored continuously during the study. Most patients (99%) accepted the syrup without difficulty. Satisfactory sedation was achieved within 30 minutes by 81% of patients. The anxiety score at the time of caretaker separation and mask anesthetic induction was satisfactory for 87% and 91% of patients, respectively. A significant linear relationship between plasma drug concentration and maximal sedation score, but not anxiety score, was observed. The occurrence of adverse events was consistent with the known safety profile of midazolam. The most commonly reported adverse events were hiccoughing, hypoxemia, nausea, and emesis. It was concluded that a new oral syrup formulation of midazolam, 0.25 to 1.0 mg/kg, effectively induced rapid-onset, dose-related, adequate, and safe sedation and anxiolysis in pediatric patients who underwent operative procedures. Sedative effects were related to plasma concentrations of both midazolam and the primary metabolite, alpha-hydroxymidazolam. Oral midazolam, 1.0 mg/kg, administered within 30 minutes of the expected procedure or anesthetic induction should provide safe and effective sedation to a majority of children ages 6 months to 16 years. PMID- 10868309 TI - Absence of pharmacokinetic drug interaction of levetiracetam with phenytoin in patients with epilepsy determined by new technique. AB - Levetiracetam has recently been approved as an adjunctive medication for partial seizures and frequently will be added to phenytoin. The objective of this study was to determine the presence or absence of a pharmacokinetic drug interaction of levetiracetam with phenytoin. A stable isotope tracer technique using deuterium labeled (D10) phenytoin and high-performance liquid chromatography with ultraviolet detection (rather than mass spectrometric detection) was employed. Tracer doses of D10-phenytoin were administered i.v. before and 12 weeks after adding levetiracetam to the regimen of 6 subjects on phenytoin monotherapy for epilepsy. Blood was collected for 96 hours after each infusion. The following pharmacokinetic parameters were determined for phenytoin: Cmax, Cmin, Cavo, AUC, CL, t 1/2, VD, and free (nonprotein bound) fraction. The ratio and the 90% confidence interval of the ratio of log-transformed mean values for phenytoin pharmacokinetic parameters before (denominator) and after (numerator) adding levetiracetam all fell within the range of 0.85 to 1.17 (two one-sided test). The authors conclude that the addition of levetiracetam did not bring about clinically important changes in phenytoin pharmacokinetic parameters and that it is not necessary to change the phenytoin dosing rate when levetiracetam is added to phenytoin. PMID- 10868310 TI - Sex differences in the pharmacokinetics of dehydroepiandrosterone (DHEA) after single- and multiple-dose administration in healthy older adults. AB - The pharmacokinetics of exogenously administered DHEA have not been well characterized despite its increasing use in therapeutic and research investigations. The purpose of this study was to evaluate the pharmacokinetics of DHEA and its sulfated metabolite (DHEA-S) after single- and multiple-dose oral administration of DHEA 200 mg. Healthy older adult volunteers (7 women, 6 men) ages 65 to 79 years were studied on five visits separated by 1 week. Subjects received daily administration of placebo (days 1 to 7), DHEA 200 mg (days 8 to 22), and placebo (days 23 to 29). Blood samples were collected over 24 hours on days 1, 8, 15, 22, and 29 for DHEA and DHEA-S determinations by RIA. Pharmacokinetic parameter estimates were calculated by noncompartmental methods. Administration of DHEA 200 mg resulted in higher DHEA Cmax, AUC, and overall concentrations in women than in men (p < 0.03); DHEA-S parameter estimates were similar between men and women. Following a single dose of DHEA 200 mg, DHEA concentrations increased 5- to 6-fold in both men and women, and DHEA-S concentrations increased 5-fold in men and 21-fold in women relative to endogenous concentrations. The results of this study indicate that the pharmacokinetics of DHEA differ between older men and women. PMID- 10868311 TI - Extravascular administration of interferon alfa-N3 increases serum exposure and 2 5(A) synthetase activity. AB - The objective of this study was to evaluate the pharmacokinetics, pharmacodynamic response, and safety of single intravenous (i.v.), intramuscular (i.m.), and subcutaneous (SQ) doses of interferon alfa-n3. Six healthy adults received 10 million units of i.v., i.m., and SQ interferon alfa-n3 in a randomized three period crossover fashion. Serum interferon alfa-n3 concentrations and 2'-5' oligoadenylate synthetase (2-5[A] synthetase) activity in peripheral blood mononuclear cells were determined after each dose. Extravascular administration significantly increased mean serum interferon alfa-n3 AUC values (1152 +/- 214, 944 +/- 209, and 576 +/- 188 U.h/mL, p < 0.001, with SQ, i.m., and i.v. administration, respectively) and 2-5(A) synthetase activity at 36 and 48 hours after dosing. Mild to moderate flu-like symptoms were reported by all 6 subjects, with no route-related difference in type or incidence. Interferon alfa-n3 is generally well tolerated by the i.v., i.m., and SQ routes, with i.m. and SQ administration maximizing serum exposure and 2-5(A) synthetase activity. PMID- 10868312 TI - Pharmacodynamics and pharmacokinetic-pharmacodynamic relationships of atorvastatin, an HMG-CoA reductase inhibitor. AB - The objective of this study is to determine the relationships between plasma atorvastatin concentrations, LDL (low-density lipoprotein) cholesterol reduction, and atorvastatin dose; the earliest time at which lipid levels change when atorvastatin treatment is initiated or discontinued; and alterations in LDL particle composition. Twenty-four subjects with elevated LDL-cholesterol were treated with escalating daily doses of 5, 20, and 80 mg atorvastatin for 6 weeks each. Serial plasma lipid and lipoprotein analyses were performed during the initiation and discontinuation of atorvastatin therapy, as well as at steady state. LDL-apolipoprotein B and LDL-cholesterol were measured directly after ultracentrifugation, and LDL-cholesterol also was estimated by the method of Friedewald. Steady-state atorvastatin pharmacokinetic parameters were estimated on the last day of each dosing period. LDL-cholesterol (Friedewald) reductions of 34%, 43%, and 57% were produced by atorvastatin doses of 5, 20, and 80 mg, respectively. The mean dose-response relationship was log linear, and almost all individual dose-response curves paralleled the mean curve. LDL-apolipoprotein B reductions were slightly less than those of LDL-cholesterol. Atorvastatin area under the curve (AUC(0-24) values increased proportionally with dose, while values of Cmax (maximum concentration) increased more than proportionally, and Cmin (minimum concentration) increased less than proportionally. Following initiation of dosing, statistically significant decreases in total cholesterol, LDL-cholesterol (beta quant), and LDL-apolipoprotein B were observed within 24 hours and in LDL-C (Friedewald) within 72 hours. Following discontinuation of drug dosing, statistically significant increases were observed in total cholesterol and LDL-cholesterol (Friedewald) within 48 hours and in LDL cholesterol (beta quant) and LDL-apolipoprotein B within 72 hours. At each dose, an individual's LDL-cholesterol response was not correlated with AUC(0-24). In conclusion, atorvastatin produces marked LDL-cholesterol reductions, the mean dose-response relationship is log linear, almost all individual dose-response curves parallel the mean dose-response curve, onset and cessation of action are rapid, the estimated and measured LDL-cholesterol are the same, LDL-cholesterol and LDL-Apo B reductions are similar, and plasma concentrations are not correlated with LDL-cholesterol reduction at a given dose. PMID- 10868313 TI - Mycophenolic acid area under the curve values in African American and Caucasian renal transplant patients are comparable. AB - The possibility of an effect of ethnicity on the pharmacokinetics of mycophenolic acid, the immunosuppressive metabolite of the prodrug mycophenolate mofetil, was studied over 90 days following renal transplantation in African American (n = 13) and Caucasian patients (n = 20). Since renal dysfunction and time after transplant surgery are two factors known to alter mycophenolic acid pharmacokinetics, two-way analysis of variance of the data at each time point with ethnicity and renal function status as covariates was used to evaluate the possibility of an ethnicity effect on the pharmacokinetic parameters. No statistically significant difference based on ethnicity was detected for the primary pharmacokinetic parameters, abbreviated mycophenolic acid area under the concentration-time curve (MPA AUC), or the predose trough concentration on study days 4, 7, 14, 28, or 90. A statistically significant decrease in MPA AUC and increase in oral apparent clearance were observed in renally impaired patients regardless of ethnicity on days 4, and 4 and 7, respectively. The suggested mechanism for these differences is uremia-induced increased MPA free fraction, leading to a temporary increased clearance for this restrictively cleared drug. PMID- 10868314 TI - Single-dose pharmacokinetics of nateglinide in subjects with hepatic cirrhosis. AB - This single-dose, open-label, parallel-group study compared the pharmacokinetics and tolerability of 120 mg doses of nateglinide, a physiologic mealtime glucose regulator for type 2 diabetes, in 8 subjects with cirrhosis and 8 matched healthy subjects. In both groups, plasma concentration peaked in a median of 0.5 hours, and mean terminal elimination half-lives were comparable. Mean +/- SD pharmacokinetic parameters in cirrhotic versus healthy subjects were slightly different (Cmax, 7.7 +/- 4.9 vs. 5.6 +/- 1.3 micrograms/ml; AUC(0-t), 18.5 +/- 7.5 vs. 14.2 +/- 2.1 micrograms.h/ml, respectively). Mean apparent total clearance and mean renal clearance in both groups were comparable. Mean protein bound fractions were equivalent; binding appeared unaltered by metabolites. One cirrhotic and 2 healthy subjects each reported one adverse event. No statistically significant or clinically relevant alteration in pharmacokinetic parameters of nateglinide resulted from hepatic dysfunction, and it was well tolerated; therefore, adjustment of nateglinide dosage is not required in subjects with mild to moderate cirrhosis. PMID- 10868315 TI - Linear pharmacokinetic behavior of ropinirole during multiple dosing in patients with Parkinson's disease. AB - The objectives of this study were to measure the pharmacokinetics of ropinirole at steady state when the drug is used as an adjunct to L-dopa and evaluate the long-term tolerability of ropinirole in this indication. Twenty-four patients who were taking L-dopa for Parkinson's disease and experiencing a lack of symptomatic control were recruited. Patients received open-label adjunctive treatment with ropinirole for up to 2 years. The starting dose was 0.5 mg bid, which could be titrated to a maximum of 6.0 mg tid. Ropinirole demonstrated approximately dose linear pharmacokinetics at steady state; corresponding values were higher during tid than bid dosing. A reduction in mean L-dopa dose was maintained throughout the trial. The combination of L-dopa and ropinirole was generally well tolerated, with only 1 patient withdrawing from treatment because of adverse events. Thus, ropinirole shows approximately linear steady-state pharmacokinetics and a good safety profile when administered with L-dopa. PMID- 10868316 TI - The effect of 3-month ingestion of Ginkgo biloba extract on pancreatic beta-cell function in response to glucose loading in normal glucose tolerant individuals. AB - Ginkgo biloba extract is ingested on a chronic basis for enhancing mental focus and improving cognitive function in the elderly. This study was undertaken to determine the effect of Ginkgo biloba extract on glucose-stimulated pancreatic beta-cell function in normal glucose-tolerant individuals. Twenty individuals (14 females and 6 males, ages 21-57 years) underwent a 2-hour 75 g oral glucose tolerance test (OGTT) before and after ingestion of Ginkgo biloba extract (120 mg/day at bedtime) for 3 months. Ginkgo biloba extract ingestion caused a decrease in systolic blood pressure from 125 +/- 15 to 118 +/- 12 mmHg (p < 0.05) and in diastolic blood pressure from 86 +/- 10 to 68 +/- 10 (p < 0.01). Fasting plasma insulin and C-peptide were increased, and the insulin and C-peptide areas under the curve during the OGTT changed from 136 +/- 55 to 162 +/- 94 microU/ml/h (p = 0.1232) and 9.67 +/- 5.34 to 16.88 +/- 5.20 ng/ml/h (p < 0.001), respectively. The observed dissimilar insulin/C-peptide response curves may be due to Ginkgo biloba-induced increase of the rate of insulin metabolic clearance. PMID- 10868317 TI - Celecoxib does not significantly alter the pharmacokinetics or hypoprothrombinemic effect of warfarin in healthy subjects. AB - The objective of this study was to determine the effects of celecoxib, an anti inflammatory/analgesic agent that primarily inhibits COX-2 and not COX-1 at therapeutic doses, on the steady-state pharmacokinetic profile and hypoprothrombinemic effect of racemic warfarin in healthy volunteers. Twenty-four healthy adult volunteers on maintenance doses of racemic warfarin (2-5 mg daily), stabilized to prothrombin times (PT) 1.2 to 1.7 times pretreatment PT values for 3 consecutive days, were randomized to receive concomitant celecoxib (200 mg bid) or placebo for 7 days in an open-label, multiple-dose, randomized, placebo controlled, parallel-group study of warfarin pharmacokinetics and PT. Steady state exposure of S- and R-warfarin (area under the curve [AUC]) and maximum plasma concentration (Cmax) in subjects receiving celecoxib were within 2% to 8% of the warfarin AUC and Cmax in subjects receiving placebo during the concomitant treatment period. In addition, PT values were not significantly different in subjects receiving warfarin and celecoxib concomitantly compared with subjects receiving warfarin and placebo. In conclusion, concomitant administration of celecoxib has no significant effect on PT or steady-state pharmacokinetics of S- or R-warfarin in healthy volunteers. PMID- 10868318 TI - Prospective allometric scaling: does the emperor have clothes? AB - We are not suggesting that an allometric relationship between pharmacokinetic parameters in animals and humans does not exist; we believe that it does. We are suggesting that using prospective AS to select doses in an FTIM study may lead to a false sense of security given the large publication bias in the literature. There are a number of unrecognized pitfalls to this approach, including (1) prediction intervals so wide as to be of limited use, (2) prediction error is often no better than arbitrarily chosen constants, and (3) it is not possible to determine which drugs will fail a priori. We encourage journals to publish studies in which prospective AS has failed so as scientists we may begin to see what makes these compounds different, with a goal toward better prediction of human pharmacokinetics. PMID- 10868319 TI - Memory and ECT: from polarization to reconciliation. PMID- 10868320 TI - Balancing speed of response to ECT in major depression and adverse cognitive effects: role of treatment schedule. AB - Schedule of administration (number of ECT per week and total number of treatments in the course) is one of a number of factors that may significantly influence the degree of cognitive impairment induced by ECT. We examined the effect of twice (ECT x 2) versus three times weekly (ECT x 3) bilateral ECT on cognitive function, particularly memory, in patients with major depression. Two studies were conducted, both double blind and controlled by the administration of simulated ECT (anesthesia and muscle relaxant only with no electrical stimulation). The results of these studies showed that the antidepressant effect of the two schedules, when assessed at the end of the ECT course, was equal. Speed of response was significantly greater with ECT x 3 but this schedule induced more severe memory impairment, even when the number of ECT in the series was not significantly different between the two groups. These findings are in general accordance with other studies that were similar in design although not as rigorously controlled. They support the conclusion that ECT x 2 is the more appropriate schedule for regular clinical practice unless speed of response is an overriding concern. In an era when patients administered ECT tend to be older and are more likely to manifest cognitive impairment for other reasons, choice of schedule is of particular relevance along with other factors such as electrode placement and stimulus intensity that influence ECT-induced cognitive impairment. PMID- 10868321 TI - Electrophysiological correlates of the adverse cognitive effects of electroconvulsive therapy. AB - Resting state, eyes closed, 19-lead EEG recordings were obtained at pre-ECT baseline and just prior to penultimate treatment and during the week following the ECT course in 59 patients with major depression. Patients had been randomized to ECT conditions that varied in electrode placement and stimulus intensity. The EEG data were submitted to power spectral analysis, and global and topographic effects were characterized for the delta and theta frequency bands. Relations between the EEG changes and scores on three cognitive measures were examined. The period of disorientation immediately following RUL ECT was associated with an accentuation of delta power in anterior frontal and temporal regions. Across the electrode placements, increased theta activity in left frontotemporal regions was associated with longer recovery of orientation. Post-ECT decrements in global cognitive status, as assessed by the modified Mini-Mental State exam, were associated with a greater increase in delta relative to theta power, globally across the cortex. The magnitude of retrograde amnesia for autobiographical events correlated with increased theta activity in left frontotemporal regions. The findings suggest that distinct neurophysiological changes subserve the therapeutic and adverse cognitive effects of ECT. Postictal disorientation and post-ECT retrograde amnesia appear to share a common physiological substrate. PMID- 10868322 TI - Subjective memory complaints: a review of patient self-assessment of memory after electroconvulsive therapy. AB - Interest in patients' subjective complaints about the adverse cognitive effects of electroconvulsive therapy (ECT) spans several decades. This article reviews the major areas that have been examined in relation to patients' subjective assessment of memory function: 1) technical aspects in the administration of ECT; 2) objective tests of cognitive function; and 3) clinical state. For the most part, subjective assessments of memory following ECT have relied on a single instrument, the Squire Subjective Memory Questionnaire (SSMQ). While older reports of the impact of the technical aspects of ECT on subjective memory assessment following ECT suggest a detectable negative influence with certain forms of treatment, most recent studies indicate that subjective memory improves following ECT. This shift in findings may be due to the change in practice from sine wave to brief-pulse ECT. While the impact of ECT on objective tests of memory is clear and reproducible, the relationship of objective findings to subjective memory assessment appears to be weak. Instead, subjective reports of cognitive function are strongly influenced by mood state. Current batteries of objective tests of memory may not include components that are most affected in reports about subjective memory. In addition, the literature mainly reports group effects, and sample sizes have been small. We lack data on the number of individuals who believe ECT has had a markedly negative effect on memory functioning, and on the characteristics of memory function in this subgroup of patients who complain of severe impairment. Furthermore, there is a paucity of information relating patient characteristics to subjective memory outcomes with ECT. PMID- 10868323 TI - Electroconvulsive therapy and memory loss: a personal journey. AB - The cause for the significant gap between research and anecdotal evidence regarding the extent of some memory loss after electroconvulsive therapy (ECT) has never been adequately explained. A patient's development of awareness and self-education about her severe side effects from ECT raises questions regarding many current assumptions about memory loss. ECT-specific studies, which conclude that side effects are short term and narrow in scope, have serious limitations, including the fact that they do not take into account broader scientific knowledge about memory function. Because of the potential for devastating and permanent memory loss with ECT, informed consent needs significant enhancement until advancing research on both improved techniques and on better predictive knowledge regarding memory loss progresses to making a greater impact on clinical applications. Follow-up care and education in coping skills need to be a regular part of ECT practice when patients do experience severe effects. PMID- 10868324 TI - Herbal treatments for ECS-induced memory deficits: a review of research and a discussion on animal models. AB - During the last decade the use of herbal medicinal substances in the attenuation of anterograde and retrograde amnesia induced by electroconvulsive shock (ECS) has been studied using animal research. We will discuss the background of herbal medicine in India, review the research findings on herbal medicines for ECS induced amnestic deficits, and examine the applications and limitations of animal models in this context. We will focus on our own research and insights, with particular emphasis on practical issues. PMID- 10868325 TI - Anesthesia for electroconvulsive therapy: a review. AB - Anesthetic techniques have evolved to improve the comfort and safety of modern electroconvulsive therapy (ECT). The authors review the literature and discuss the selection, preparation, and management from an anesthetic perspective. Specifically, the management of medications preprocedure and coexisting diseases is discussed. A review of induction agents, muscle relaxants, and other medications utilized in ECT is included. PMID- 10868326 TI - A preliminary study of the effects of electroconvulsive therapy on regional brain glucose metabolism in patients with major depression. AB - Animal studies have shown that a course of electroconvulsive shock (ECS) leads to a significant reduction in glucose metabolism in rat brains 1 day after the last ECS. In humans, of the two positron emission tomography (PET) studies that assessed the effects of a course of electroconvulsive therapy (ECT) on brain glucose metabolism in depressed patients, one reported no change while the other found a trend for reduction in glucose metabolism in frontal cortical region 24 hours after last ECT. The changes in glucose metabolism detected 24 hours after the last ECS/ECT treatment might simply be due to subacute effects of a seizure. We hypothesized that the changes in brain metabolism that persist 1 week after a course of ECT are more likely to underlie the therapeutic effects of ECT. We, therefore, investigated the effects of a course of ECT on brain glucose metabolism 1 week after last ECT by using PET and [18F]fluorodeoxyglucose (FDG). Six patients who met DSM-IV criteria for a diagnosis of major depressive disorder (unipolar), and were referred for ECT as the clinically indicated treatment were recruited. They underwent two PET scans, one prior to first ECT and the second a week after last ECT. The number of ECT treatments subjects received ranged from 8 to 12 with a mean of 11. Five out of six patients responded to the ECT treatment. Cerebral metabolic rates for glucose were slightly lower in most regions post treatment compared with pretreatment but the differences were not statistically significant. Similarly, there was no significant correlation between changes in regional cerebral metabolic rates for glucose (rCMRglc) and changes in Hamilton Depression Rating Scale (HAM-D 21-item) scores. Our results might suggest that rCMRglc rates are not altered 1 week after a therapeutic course of ECT in depressed patients. Further studies using new generation PET scanners, which have a higher resolution, in larger numbers of depressed patients, are clearly needed before firm conclusions can be drawn. PMID- 10868327 TI - Cardiovascular response during ECT: a cross-over study across stimulus conditions. AB - This study examined the effect of stimulus conditions on rate pressure product (RPP) during ECT. Seizure duration as well as baseline, ictal, and postictal RPP were recorded in 28 patients who received ECT under four stimulus conditions (unilateral threshold, unilateral suprathreshold, bilateral threshold, and bilateral suprathreshold). Seizure duration did not differ between the stimulus conditions. RPP significantly rose from baseline under every stimulus condition. Both ictal and postictal RPP values were different between stimulus conditions (one-way repeated-measure analysis of variance). Pair-wise comparisons showed that unilateral threshold ECT produced significantly lower RPP than unilateral suprathreshold and bilateral suprathreshold ECT conditions. The selective differences in RPP during ECT as well as their correspondence with the known therapeutic potency of ECT under these stimulus conditions suggest that RPP may be a potential index of ECT's therapeutic potency. PMID- 10868328 TI - ECT in mixed affective states: a case series. AB - There is limited information regarding the effectiveness of electroconvulsive therapy (ECT) as a treatment for patients in a mixed affective state. The authors report their experience using this treatment in medication-resistant patients meeting Research Diagnostic Criteria (RDC) for both mania and major depression. Clinical and response characteristics of these patients are described. Forty-one consecutively admitted patients meeting the RDC for mania received pharmacotherapy. Eight patients failing to respond to pharmacotherapy were referred for ECT, and seven consented. All met RDC for both mania and major depressive disorder. All patients receiving ECT remitted. The patient who did not accept ECT did not improve and ultimately needed transfer to a state hospital for longer term care. Mixed manic-depressive states are responsive to ECT, even in medication-refractory patients. PMID- 10868330 TI - ECT and intracranial vascular masses. AB - In 1990, the APA Task Force on ECT cited no "absolute" contraindications to ECT but "Substantial Risk" to be associated with ECT for patients with space occupying or other cerebral lesions with increased intracranial pressure and with bleeding or otherwise unstable vascular aneurysm or malformation. These findings indicate that patients with intracranial vascular masses are at increased risk for serious morbidity and mortality. Several authors have reported performing ECT in patients with intracranial vascular masses without adverse events by monitoring blood pressure both with and without pharmacologic intervention. Given the relatively recent change in practice of considering ECT for patients with intracranial vascular masses and the few number of cases thus far reported, we present a review of the existing literature and two additional cases of ECT performed with good result and no adverse events. With the cases we have presented, the literature now contains eight cases of ECT performed in patients with intracranial vascular masses, none of which had adverse outcomes. While such numbers do not establish unequivocal safety in this population, and the individual ECT practitioner must continue to make a risk/benefit analysis on a case-by-case basis, this report adds to the growing literature on the safety and efficacy of ECT for such patients. PMID- 10868329 TI - ECT in the presence of brain tumor and increased intracranial pressure: evaluation and reduction of risk. AB - The presence of brain tumor and increased intracranial pressure has long been considered an absolute contraindication to electroconvulsive therapy. Recently, however, the American Psychiatric Association Task Force Report questioned the absolute nature of this contraindication and recommended a detailed evaluation of the risk-benefit ratio and measures to decrease the risks involved in treatment of affected persons. After a careful review, electroconvulsive therapy was administered to a 61-year-old patient who had severe medication-resistant major depression and a left temporal anaplastic astrocytoma with brain edema. Special attention was given to reduce intracranial pressure and minimize neurologic side effects. A course of eight nondominant unilateral electroconvulsive therapy treatments improved the depression significantly, without serious complications at the 4-month follow-up examination. With appropriate modifications, electroconvulsive therapy may be considered a treatment option even in the presence of clinical evidence of increased intracranial pressure. Further studies are needed to assess and minimize risks of electroconvulsive therapy in association with brain tumor. PMID- 10868331 TI - Combined ECT and clozapine in treatment-resistant mania. AB - A treatment-resistant manic patient failed to respond to conventional treatment, adjunctive olanzapine (20 mg/day), clozapine (600 mg/day), or ECT alone, but did respond to ECT combined with low dose clozapine (200 mg/day). Maintenance ECT combined with clozapine resulted in a remission over the 18-month period. PMID- 10868332 TI - Divided attention and memory: evidence of substantial interference effects at retrieval and encoding. AB - In 5 divided attention (DA) experiments, students (24 in each experiment) performed visual distracting tasks (e.g., recognition of words, word and digit monitoring) while either simultaneously encoding an auditory word list or engaging in oral free recall of the target word list. DA during retrieval, using either of the word-based distracting tasks, produced relatively larger interference effects than the digit-monitoring task. DA during encoding produced uniformly large interference effects, regardless of the type of distracting task. Results suggest that when attention is divided at retrieval, interference is created only when the memory and concurrent task compete for access to word specific representational systems; no such specificity is necessary to create interference at encoding. During encoding, memory and concurrent tasks compete primarily for general resources, whereas during retrieval, they compete primarily for representational systems. PMID- 10868333 TI - The persistence of structural priming: transient activation or implicit learning? AB - Structural priming in language production is a tendency to recreate a recently uttered syntactic structure in different words. This tendency can be seen independent of specific lexical items, thematic roles, or word sequences. Two alternative proposals about the mechanism behind structural priming include (a) short-term activation from a memory representation of a priming structure and (b) longer term adaptation within the cognitive mechanisms for creating sentences, as a form of procedural learning. Two experiments evaluated these hypotheses, focusing on the persistence of structural priming. Both experiments yielded priming that endured beyond adjacent sentences, persisting over 2 intervening sentences in Experiment 1 and over 10 in Experiment 2. Although memory may have short-term consequences for some components of this kind of priming, the persisting effects are more compatible with a learning account than a transient memory account. PMID- 10868334 TI - Reasoning about geography. AB - To understand the nature and etiology of biases in geographical judgments, the authors asked people to estimate latitudes (Experiments 1 and 2) and longitudes (Experiments 3 and 4) of cities throughout the Old and New Worlds. They also examined how people's biased geographical judgments change after they receive accurate information ("seeds") about actual locations. Location profiles constructed from the pre- and postseeding location estimates conveyed detailed information about the representations underlying geography knowledge, including the subjective positioning and subregionalization of regions within continents; differential seeding effects revealed between-region dependencies. The findings implicate an important role for conceptual knowledge and plausible-reasoning processes in tasks that use subjective geographical information. PMID- 10868335 TI - Why do categories affect stimulus judgment? AB - The authors tested a model of category effects on stimulus judgment. The model holds that the goal of stimulus judgment is to achieve high accuracy. For this reason, people place inexactly represented stimuli in the context of prior information, captured in categories, combining inexact fine-grain stimulus values with prior (category) information. This process can be likened to a Bayesian statistical procedure designed to maximize the average accuracy of estimation. If people follow the proposed procedure to maximize accuracy, their estimates should be affected by the distribution of instances in a category. In the present experiments, participants reproduced one-dimensional stimuli. Different prior distributions were presented. The experiments verified that people's stimulus estimates are affected by variations in a prior distribution in such a manner as to increase the accuracy of their stimulus reproductions. PMID- 10868336 TI - Are affective events richly recollected or simply familiar? The experience and process of recognizing feelings past. AB - The author used the remember/know paradigm and the dual process recognition model of A. P. Yonelinas, N. E. A. Kroll, I. Dobbins, M. Lazzara, and R. T. Knight (1998) to study the states of awareness accompanying recognition of affective images and the processes of recollection and familiarity that may underlie them. Results from all experiments showed that (a) negative stimuli tended to be remembered, whereas positive stimuli tended to be known; (b) recollection, but not familiarity, was boosted for negative or highly arousing and, to a lesser extent, positive stimuli; and (c) across experiments, variations in depth of encoding did not influence these patterns. These data suggest that greater recollection for affective events leads them to be more richly experienced in memory, and they are consistent with the idea that the states of remembering and knowing are experientially exclusive, whereas the processes underlying them are functionally independent. PMID- 10868337 TI - The extralist-feature effect: evidence against item matching in short-term recognition memory. AB - The authors varied the similarity between negative probes and study items in a short-term item-recognition task. Current models treat similarity as a function of the number of occurrences of the probe's features in the study set, a factor that is often confounded with the number of the probe's features occurring in the study set. Unconfounded comparisons showed that performance reflected only the latter factor, with response time a linear function of the number of probe features in the study set. The effect was obtained for both stimuli with manipulated features (colored shapes) and words. Number of presented features is a global property of the study list, but existing global models calculate familiarity by averaging across item matches and cannot readily accommodate the data. The authors proposed that the probe's features are compared with a global representation of the study set's features. PMID- 10868338 TI - Toxigenic strains of Fusarium moniliforme and Fusarium proliferatum isolated from dairy cattle feed produce fumonisins, moniliformin and a new C21H38N2O6 metabolite phytotoxic to Lemna minor L. AB - Corn samples suspected of causing refusal-to-eat syndrome in dairy cattle were examined mycologically. Fusarium moniliforme (14 isolates) and F. proliferatum (12 isolates) were the predominant fungi present. These isolates were tested for mycotoxin production on rice at 25 degrees C. Each strain of F. moniliforme produced fumonisin B1 (FB1: 378-15,600 ppm) and fumonisin B2 (FB2: 2-1050 ppm). Each strain of F. proliferatum produced moniliformin (45-16,000 ppm), FB1 (27 6140 ppm), and FB2 (5-1550 ppm). In addition, a new Fusarium metabolite of molecular composition C21H38N2O6 was produced by 10 of the F. moniliforme isolates and 7 of the F. proliferatum isolates. The metabolite's 1H- and 13C-NMR, HRFAB/MS and IR spectra indicate an alpha amino acid. It is toxic to Lemna minor L. duckweed (LD50 100 micrograms/mL). PMID- 10868339 TI - Cloning and sequence analysis of a Phytophthora cinnamomi gene which encodes for cinnamomin, a toxin with implications in root rot of cranberry. AB - We used a polymerase chain reaction (PCR) based cloning strategy to isolate cinnamomin genes from Phytophthora cinnamomi 8601, a pathogen responsible for cranberry root rot. Complete DNA sequence analysis of nine recombinant clones revealed two different classes of genes, each class consisting of genes with identical DNA sequences. Both classes of genes (Cin-1 and Cin-2) contained an open reading frame encoding a protein of 122 amino acid residues. The encoded proteins, named cinnamomin-1 and cinnamomin-2 (Cin-1 and Cin-2), were highly homologous to other proteins of the elicitin family and contained a 19 amino acid residue long signal peptide sequence. Both Cin-1 and Cin-2 proteins showed higher degree of sequence homology to the alpha-elicitins than beta-elicitins; moreover, a Val residue was found at position 13 of the putative mature Cin-1 and Cin-2 proteins. Because alpha-elicitins and beta-elicitins are known to contain a Val and a Lys residue, respectively, at this position, we concluded that both Cin-1 and Cin-2 genes from P. cinnamomi 8601 encode for alpha cinnamomins, Cin-1 and Cin-2. PMID- 10868340 TI - Characterization of Staphylococcus aureus beta-toxin induced leukotoxicity. AB - One virulence determinant of Staphylococcus aureus is Beta-toxin, a 37 Kd magnesium-dependent sphingomyelinase C. This toxin lyses erythrocytes (RBCs) containing sphingomyelin in the outer lipid layer of their plasma membrane. Although membranes of both human polymorphonuclear leukocytes (PMNs) and lymphocytes (MNLs) contain small amounts of sphingomyelin, the effect of Beta toxin on these cells remains controversial. The purpose of this study was to investigate the hemolytic activity of this toxin on RBCs of various species and determine the leukotoxic nature on several types of human leukocytes. One nanogram of Beta-toxin lysed 115,000 sheep erythrocytes (sRBCs) and 82,000 human erythrocytes (hRBCs) in a 'hot-cold' assay and caused cytotoxicity to 325 PMNs and MNLs. Both hemolytic and leukotoxic activity were found to be magnesium dependent. RBC susceptibility to Beta-toxin correlated with the reported sphingomyelin content of each species. Scanning electron microscopy (SEM) demonstrated that 'hot-cold' incubation with Beta-toxin in the presence of magnesium caused significant morphological changes in the surface structure of both RBCs and PMNs. The changes included the formation of pits and membrane invaginations in the RBCs. The PMNs lost their ruffled membrane appearance and showed overall membrane disintegration. This study demonstrated that the viability of sphingomyelin-containing PMNs and MNLs was significantly decreased by the addition of Beta-toxin, indicating that this toxin does, in fact, have a leukotoxic nature. Leukocytes did not have significant membrane invaginations unlike toxin-treated RBCs; therefore, it is possible that leukotoxicity does not result from membrane lysis. PMID- 10868341 TI - Evidence for palytoxin as one of the sheep erythrocyte lytic in lytic factors in crude extracts of ciguateric and non-ciguateric reef fish tissue. AB - The occurrence of palytoxin or its congener in fish extracts has been presented in this study. The presences of hemolytic factors in fish extracts of Hawaiian reef fish and their implication in ciguatera poisoning have been shown by the sheep erythrocyte assay. By use of the anti-palytoxin inhibition assay with fish extracts and sheep red blood cell (RBC), it was shown that palytoxin was one of the major factors in the lysis of sheep erythrocytes. Ouabain, an antagonist of palytoxin for the Na+/K+ ATPase receptor on RBC, also showed inhibition of sheep RBC lysis by fish extracts. From these results, it was concluded that, in part, palytoxin and other palytoxin-related, hemolysin-like factors in fish extracts were responsible for sheep cell hemolysis. PMID- 10868342 TI - Edema factor from the venom of Trimeresurus elegans (Sakishimahabu). AB - An edema factor was isolated from the venom of Trimeresurus elegans using HW-55, CM-Cellulose, and Mono S column chromatographies. Homogeneity was demonstrated by the formation of a single band in polyacrylamide gel electrophoresis (pH 8.3). The edema factor has a molecular weight of 25,500, an isoelectric point of 7.5, and express edema, proteolytic and capillary permeability-increasing activities. Edema, proteolytic and capillary permeability-increasing activities are inhibited by ethylenediaminetetraacetic acid (EDTA), o-phenanthroline, and N bromosuccinimide. Additionally, this factor exhibits kinin-releasing activity. The edema factor possesses proteolytic activity as shown by hydrolyzing the Val(3)-Asn(4), His(5)-Leu(6), Ser(9)-His(10), Ala(14)-Leu(15), Leu(15)-Tyr(16), Tyr(16)-Leu(17), and Glu(21)-Arg(22) bonds of oxidized insulin B chain. The A alpha, B beta, and gamma chains of human fibrinogen were also hydrolyzed. The edema factor was found to contain 1 mol of zinc and 2 mols of calcium per mol of protein and the amino-terminal sequence was determined. PMID- 10868343 TI - Effects of intramuscular injection of a sublethal dose of the Egyptian cobra snake on the histological and histochemical pattern of the kidney. AB - The effects of intramuscular (i.m.) injection of a sub-lethal dose of cobra venom (0.015 microgram/gm body weight) on the histological and histochemical patterns of the kidney of rabbit were examined after 3, 6, and 12 hr. of envenomation. The histological observations after 3 hr. of envenomation showed glomerular congestion together with slight swelling of the cortical tubular epithelia. However, no changes were recorded in the medullar tubules. Serious alterations were recorded after 6 hr. of envenomation. It included thickening of the Bowman's capsules, signs of mesangiolysis, and glomerular collapse. The cortical tubular epithelia were swollen and revealed cytoplasmic granulation, coagulation, or depletion. Nuclear pyknosis and cellular damage were recorded in some areas. The medullar tubules showed cytoplasmic degeneration with no nuclear changes. By 12 hr. of envenomation a higher degree of severity was recorded. The glomerular tufts were hypertrophied or suffered from partial damage. Mesangiolysis and glomerulolysis were common and some glomerular tufts were completely transformed to clumps of hyaline casts. The cortical tubules showed hyaline coagulation, together with severe tubular damage in which the boundaries of the individual tubule cannot be identified. Numerous inflammatory cells were observed invading the damaged epithelial cells and the intertubular spaces. The medullar tubules showed swollen epithelia with cytoplasmic changes and nuclear pyknosis or karyolysis. Histochemically, the polysaccharide inclusion was increased in the glomerular tufts, the Bowman's capsules, and the basement membranes and brush borders of the renal tubules after 3 and 6 hr. of envenomation. By 12 hr. of envenomation, decreased PAS reactivity was recorded in all renal components except the glomerular tufts which exhibited intensive reactivity. Time-dependent depletion of lipid, protein, and RNA components was recorded in the renal tissues of the three envenomed groups. However, no changes in DNA reactivity were detected in renal tissues of the 3 hr. envenomed group. The nuclei of certain renal tubules revealed weak DNA reactivity after 6 hr. of envenomation, while most of the nuclei lost their contents by 12 hr. of envenomation. The results indicated serious histological and histochemical alterations induced in the renal tissues by 6 hr. of envenomation. Such alterations could indicate a disturbance in the functional activity of the kidney during envenomation. Therefore, nephrotoxicity should be considered as one of the serious consequences of such venom. PMID- 10868344 TI - Dietary psoralens induce hepatotoxicity in C57 mice. AB - The psoralens are secondary plant metabolites found in many fruits and vegetables. Synthetic forms of 5-methoxypsoralen (bergapten) and 8 methoxypsoralen (xanthotoxin) have been used in combination with UV radiation in skin photochemotherapy for decades. However, handling or ingestion of psoralen containing plants as well as medicinal use of these compounds have been shown to cause human health hazards. We evaluated the subacute toxicity of bergapten and xanthotoxin in a mammalian model by mixing individual chemicals into mouse diet at 0, 250, and 1000 ppm, and in combination at 500 ppm each. Feeding on individual dietary treatments at 1000 ppm significantly reduced total liver cytochrome P450 (CYP) levels in female mice compared with the control diet, but not in males. However, combining the two chemicals resulted in a significant induction of total CYP450 in both males and females. Both the combined diet and bergapten at 250 ppm caused a weak induction of CYP1A1. Weight gain was significantly less in males fed either the combined or 1000 ppm diets, while only the combined diet induced a significant weight reduction in females compared with the control diet. The psoralens also caused hypertrophy of centrolobular hepatocytes in livers of treated animals in a manner consistent with morphological alterations seen in rodent livers exposed to liver CYP-inducing agents. Neither bergapten nor xanthotoxin, however, induced a significant dose dependent toxicity in either male or female mice, suggesting that mice may not represent a good laboratory animal model for evaluating the toxicological effects of psoralens. PMID- 10868345 TI - GC/MS/MS detection of pyrrolic metabolites in animals poisoned with the pyrrolizidine alkaloid riddelliine. AB - Pyrrolic metabolites from pyrrolizidine alkaloids (PAs) were detected in liver and dried blood samples using a gas chromatography/tandem mass spectrometry (GC/MS/MS) selected product-ion-monitoring method. A calibration curve was constructed using a protein-metabolite conjugate spiked into dried bovine blood. These spiked samples served as a model for tissues from animals poisoned by the toxic metabolite of PAs. Tissue samples from pigs fed various amounts of the PA alkaloid riddelliine (from Senecio riddellii) were analyzed for pyrrolic metabolites, and the results were applied to the calibration curve to provide a measure of the degree of PA poisoning. Pyrrolic metabolites were detected in liver and blood samples of all poisoned animals at levels between 2 and 64 ppm. Although differences in metabolite levels could be discerned under the reported experimental conditions, the amount detected did not correlate with the dose of riddelliine given; and livers fixed with formalin gave greatly reduced recovery than those same livers either frozen or freeze dried. PMID- 10868346 TI - Effect of a radical scavenger "water soluble protein" from broad beans (Vicia faba) on antioxidative enzyme activity in cellular aging. AB - We isolated a free-radical scavenger "water soluble protein (WSP)" from broad beans. Hydrocortisone (HC) is known to inhibit superoxide generation and was used as the reference scavenger. WSP was examined for its effect on antioxidation in young (PDL 20, 25% of the maximum life span) and old (PDL 50, 62.5% of the maximum life span) human fibroblasts (TIG-1). Cells were treated with WSP or HC for 4 and 6 wk in young cells, and for 3 and 6 wk in old cells. The cytosolic superoxide dismutase activity in the cells treated with WSP or HC tended to decrease as compared with that in the non-treated cells (control) with the exception of WSP-treated young cells 4 wk after culturing. Young cells were equal in glutathione peroxidase activity to the control, but the activity level in WSP- or HC-treated young cells 6 wk after culturing was 10-50% lower than that in the control. Young and old cells treated with WSP or HC were superior to the control in catalase activity with the exception of HC-treated old cells. WSP- or HC treated cells were higher in glutathione (GSH) concentration than the control with the exception of WSP-treated young cells 4 wk after culturing and HC-treated old cells 6 wk after culturing. Such increases in catalase activity and GSH concentration by WSP treatment may be related to the delay of cellular aging dependent degeneration. PMID- 10868347 TI - Comparisons of effects of raw and gelatinized sago and tapioca starches on serum and liver lipid concentrations in rats. AB - Effects of raw and gelatinized sago and tapioca starches on serum and liver lipid concentrations were compared in rats fed a cholesterol-enriched diet. Total serum cholesterol and the atherogenic index in the rats fed raw sago starch showed a statistically significant decrease and tendency to decrease, respectively, when compared with those of rats fed raw tapioca starch. Both the serum total cholesterol and atherogenic index in the rats fed gelatinized sago starch showed a tendency to decrease when compared with those of rats fed gelatinized tapioca starch. Serum triacylglycerol concentrations in the rats fed raw sago starch were significantly lower than those in the rats fed raw tapioca starch. The apparent digestibility of raw sago starch was lower than that of tapioca starch. The oval shapes of undigested raw sago starch granules collected from feces remained almost unchanged in shape, but had some holes and scrapes on their surfaces. The shape surfaces of undigested raw tapioca starch collapsed, thereby not retaining their original granule shapes. On the other hand, the amylose contents in the raw and gelatinized sago starches were higher than those in the raw and gelatinized tapioca starches. These results suggested that a decrease or a tendency to decrease in serum total cholesterol levels and atherogenic indexes in raw or gelatinized sago starch-fed rats compared to those of raw or gelatinized tapioca starch-fed rats, and a decrease in serum triacylglycerol concentrations in raw sago starch-fed rats compared to those of raw tapioca starch-fed rats, were due to the lower digestibility and higher amylose content of sago starch. PMID- 10868348 TI - Change and 1-year maintenance of nutrient and food group intakes at a 12-week worksite dietary intervention trial for men at high risk of coronary heart disease. AB - We examined how dietary habits (i.e., intake of nutrients and food groups) were changed by intervention and how once adopted diets were maintained thereafter using the data of a 12-wk worksite dietary intervention trial for men at high risk of coronary heart disease (i.e., hypercholesterolemia, hyperglycemia, and/or overweight). Dietary habits were assessed pre- and post-intervention and 1 y follow-up points using a self-administered diet history questionnaire. The intervention method was a brief individual counseling based on the results of a pre-intervention assessment and a weekly distribution of newsletters. At the pre- and post-intervention points, a control group selected from the workers was used for comparison. The Keys score, and the changes in intake of saturated fatty acids (SFA), monounsaturated fatty acid, total fat, and cholesterol (the decrease), as well as dietary fiber, potassium, calcium, and iron (the increase) were significantly different between the intervention (n = 63) and control (n = 123) groups (p < 0.05). The changes were almost maintained with little recidivism at the 1 y follow-up point in the intervention group (i.e., for the decrease in SFA and Keys score, p < 0.001). The decrease in serum cholesterol level expected from the change in Keys score and body weight, taking possible regression to the mean into consideration, was almost identical to and slightly greater than (18%) those observed at the post-intervention and 1 y follow-up points, respectively. The results suggest that the favorable changes in dietary habits adopted during an intervention period were almost maintained for the subsequent 1 y period. PMID- 10868349 TI - Preventive effect of soybean resistant proteins against experimental tumorigenesis in rat colon. AB - The insoluble 'high-molecular-weight' fraction (HMF) centrifugally separable after digestion of soy protein isolate with a microbial protease of the exo-type, of which about a quarter is regarded as an indigestible 'resistant protein,' was examined for its preventive effect against colonic tumorigenesis in a model system with male F-344 rats. The rats were intraperitoneally injected with azoxymethane (15 mg/kg BW) once a week for 3 wk and were fed a 20.6% HMF diet (+0.4% DL-Met) or 14.7% casein diet (+0.3% DL-Met) supplemented with 0.2% sodium deoxycholate (DCA) or without supplementation. Twelve wk later, 5 rats of each group were inspected for formation of tumors but no tumors were visible to the naked eye. The DCA-fed casein group was conspicuous for a low count of aberrant crypt foci. At 39 wk, 6 rats of the DCA-fed casein group (n = 10) and 3 rats of the DCA-fed HMF group (n = 9) had a total of 18 tumors with a major axis of 4.0 +/- 0.4 mm and 3 tumors with an axis of 2.0 +/- 0.1 mm, respectively, in contrast to only a single tumor for the DCA-unfed casein group (nil for the DCA-unfed HMF group). The difference in tumor number and size was considered significant between these DCA-fed casein and HMF groups; that is to say, HMF feeding retarded tumor development despite the frequent occurrence of pre-neoplastic lesions. In addition, fecal bile acid excretion was much more elevated by HMF feeding than by casein feeding. It can be assumed from these observations that the antitumorigenicity of HMF is due to the inhibitory effect of soybean resistant proteins on reabsorption as well as the mucosal contact of bile acids in the intestine. PMID- 10868350 TI - Plasma cholesterol reduction by defatted soy ontjom (fermented with Neurospora intermedia) in rats fed a cholesterol-free diet. AB - To popularize defatted soy ontjom (DSB-ontjom, soy product fermented with Neurospora intermedia) as a new food, I examined the plasma cholesterol-reducing effects of DSB-ontjom and DSB in rats fed cholesterol-free diets and compared the efficiencies of these effects. DSB-ontjom greatly reduced the plasma cholesterol level and increased fecal steroid excretion as compared to DSB. DSB-ontjom was rich in pepsin-resistant protein having a high bile acid binding capacity and was abundant in isoflavone-aglycones, especially daizein. The dietary fiber (DF) of DSB-ontjom stimulated the production of short-chain fatty acids (SCFAs) by intestinal microflora. The effect of DSB-ontjom on plasma cholesterol reduction was attributed to the collaborative effects of pepsin-resistant-protein, isoflavone-aglycones and SCFA-producing DF in DSB-ontjom. PMID- 10868351 TI - Absorption of calcium, magnesium, phosphorus, iron and zinc in growing male rats fed diets containing either phytate-free soybean protein or soybean protein isolate or casein. AB - The effect of dietary phytate-free soybean protein (PFS) on intestinal mineral absorption and retention was examined in growing male rats using a three-day mineral balance technique. The rats were fed diets containing PFS, soybean protein isolate (SPI) or casein at a 20% level for 5 wk. Total calcium (Ca), magnesium (Mg), phosphorus (P), iron (Fe) and zinc (Zn) contents in diets were adjusted to 0.35, 0.05, 0.7, 0.0035 and 0.003%, respectively, by supplementation of the diet with their salts. Mineral absorption and retention ratios in rats fed the PFS diet were significantly higher than those in rats fed either the SPI or casein diet. These results suggest that PFS may be a promising dietary protein source for improving the mineral bioavailability in humans. PMID- 10868353 TI - Time-dependent changes of tissue erythorbic acid concentrations in guinea pigs. AB - The concentrations of erythorbic acid(ErA) and L-ascorbic acid(AsA) in the tissues of guinea pigs orally administered AsA or ErA were measured over the passage of time using high-performance liquid chromatography. Guinea pigs were each administered 5 mg AsA or 100 mg ErA, and killed at a specified time thereafter. The AsA concentrations in the tissues of AsA-administered guinea pigs and the ErA concentrations in the tissues of ErA-administered guinea pigs increased for 3 h after the respective administrations and decreased thereafter in both groups. The AsA concentration in the tissues of AsA-administered guinea pigs tended to be similar to the sum of AsA and ErA concentrations in the ErA administered guinea pigs within 3 h after administration. PMID- 10868352 TI - Improved method of determining retinol and retinyl palmitate in rat liver and serum by high-performance liquid chromatography. AB - In this study, we examined the effect of certain analytical procedures to determine the best method of recovering the ingested retinoids, specifically retinol and retinyl palmitate, from rat liver and serum. In this experiment, the best extraction solvent for retinol was n-hexane and that for retinyl palmitate was ethyl acetate. The best results were obtained using a mobile phase (n-hexane diethyl ether, 76:24, v/v) as the sample solvent in the assay for liver retinol, similarly, chloroform as the sample solvent in the assay for serum retinol, and for liver retinyl palmitate, the best sample solvent was methanol-toluene (5: 5, v/v). The assayed values of retinol and retinyl palmitate measured in ethanol (0.125% BHT added) and extraction solvent (0.025% BHT added) were significantly higher than those when no BHT was added to the ethanol and extraction solvent. The determination methods for extracting retinol and retinyl palmitate from the liver varied according to the conditions layed out above. Simultaneous determination of retinol and retinyl palmitate has been illustrated in previous papers by various authors: however, we found that the individual determination of retinol and retinyl palmitate was necessary to accurately assay each retinoid. PMID- 10868354 TI - Sex-dependent developmental change of rat liver cytosolic alcohol dehydrogenase activity. AB - Rat liver cytosolic alcohol dehydrogenase (ADH) is the principal enzyme which catalyzes the oxidation of ethanol. ADH activity is known to be significantly higher in females than in males. However, the precise mechanism of the sex difference in ADH activity is uncertain. Recently, we have shown that the inhibitory action of androgen and the slight facilitatory actions of progestin and estrogen are involved in the mechanism of sex-difference in adult rat liver ADH activity. In the present study, we studied the difference of the postnatal developmental changes of ADH activity between males and females. ADH activity increased rapidly after birth up to about 12 d of age, and was not different in the two sexes. In female rats, ADH activity peaked at about 30 d of age and then increased gradually to plateau levels. However, the ADH activity of male rats fell markedly between 30 and 45 d old, and the reduced enzyme activity was observed until 104 d old. In conclusion, the ADH activity of male rats fell between 30 and 45 d after birth. At the ages over the turning point, the ADH activity was significantly lower in male than in female rats. PMID- 10868355 TI - The fibula in congenital pseudoarthrosis of the tibia: the EPOS multicenter study. European Paediatric Orthopaedic Society (EPOS). AB - Fibular involvement in congenital pseudoarthrosis of the tibia (CPT) can be either a rare isolated pathology or is in association with the tibial changes. Out of 282 patients with CPT who had complete radiographic work-up, 62% (almost two thirds) demonstrated fibular pathology: 36% had true fibular pseudoarthrosis and the rest, i.e., 26% had fibular hypoplasia or dysplastic fibula. Neurofibromatosis was found in 62% of the patients with fibular pathology. The typical radiological features of tibial pseudoarthrosis are often missing in patients with fibular pathology. It is most probably because fibular changes precede the tibial involvement. In 250 patients with tibial surgical treatment, the highest rate of fusion occurred in patients with fibular involvement compared with those with normal fibula. PMID- 10868356 TI - Treatment approaches for congenital pseudarthrosis of tibia: results of the EPOS multicenter study. European Paediatric Orthopaedic Society (EPOS). AB - This study was designed to analyze the different therapeutic methods used by European Paediatric Orthopaedic Society members from 13 countries for congenital pseudarthrosis of tibia. The treatment data of 340 patients who underwent 1287 procedures for this condition were analyzed. The essential findings were that the method of choice needed to approach the biological problem with the aims of: (1) resecting the pseudarthrosis to provide stability, the basic requirement for bony consolidation; (2) correcting length discrepancy and axial deformity; (3) achieving fusion; and (4) solving the additional problems around the main deformity such as alignment, leg length discrepancy and ankle valgus. The Ilizarov technique emerged as being the optimal method, having the highest rate of fusion (75.5%) of pseudarthrosis and rate of success in correction of the additional deformities. There was also consensus that surgery should be avoided before the third year of life. PMID- 10868357 TI - Free vascular fibular transfer in congenital pseudoarthrosis of the tibia: results of the EPOS multicenter study. European Paediatric Orthopaedic Society (EPOS). AB - This paper presents a review of the literature, describes the principal author's (B.R.) personal experience and provides the results of the European Paediatric Orthopaedic Society (EPOS) multicenter study. The objective is to evaluate the present status and future role of free vascular fibular transfer in treating congenital pseudoarthrosis of the tibia. Variables such as the selection of cases, age at operation, technical surgical details and postoperative results will be considered. The data on the EPOS study were incomplete at the time of writing, but the considerable amount of information already amassed is a valuable contribution to this updated report. PMID- 10868358 TI - Functional results at the end of skeletal growth in 30 patients affected by congenital pseudoarthrosis of the tibia. AB - From a multicentric study on Congenital Pseudoarthrosis of the tibia (CPT) conducted on 340 patients, we evaluated the functional results on a group of thirty patients who were at the end of skeletal growth (age < or = 16 years). The prognosis of CPT is very much related to the radiologic classification. Crawford type 2 and Crawford type 4 pseudoarthrosis have a worse prognosis, with a lower percentage of fusion at the site of pseudoarthrosis. Crawford 4 patients have the worst functional results. Most of them showed a severe leg length discrepancy, needed a permanent brace, with an ankle joint function fair or poor. The presence of fibula pseudoarthrosis seems to be responsible for most of the worst functional results. The level of deformity caused by either the natural course of the disease or its treatment, is decisive in the evaluation of the functional results. PMID- 10868359 TI - Congenital pseudoarthrosis of the tibia. AB - Congenital pseudoarthrosis of the tibia remains one of the most difficult conditions to treat in orthopedic surgery. Seven cases were treated in our hospital by different methods. Three out of seven patients were healed, two of these refractured. At follow-up, the success rate was 14% (one out of seven cases). It is our recommendation that early primary amputation with an appropriate prosthesis should be considered, and that the final evaluation should not be based on obtaining bone union, but on the level of function of the lower extremity. PMID- 10868360 TI - A new methodology for the measurement of the Wiberg angle in infants under 3 months. AB - This article describes the measurement of the Wiberg center edge angle in infants under 3 months using sonographic images of their hips. In the literature review, there was no reference of the application of this angle in this particular age group. Thus, a methodology has been developed based upon the fact that, at this age, the acetabular roof constitutes a large portion of hyaline cartilage. In this way, it was concluded that it was not possible to apply the original Wiberg method since the center of the femoral head can be only estimated and not accurately determined in plain radiography. For this present study, 400 hips of 200 infants were analyzed. All these hips were classified as type 1a or 1b, according to Graf. The sonographic images were transferred to a Pentium computer through a video Spigot interface. The images were analyzed by software specially developed for this purpose. Since ultrasonography allows the exact recognition of the anatomical elements of the hip joint in young children, software provided the acetabular cartilaginous roof angle that corresponds to the center edge angle in adults. The authors believe that this method will be helpful in the early detection of morphological alterations in the hip joint. PMID- 10868361 TI - A simplified valgus osteotomy of the proximal femur in children. AB - Valgus osteotomy of the hip is required in a number of orthopedic conditions in children. We present a simplified valgus osteotomy in which a dynamic compression plate is used. This technique has the advantages of using easily available equipment, requiring only one osteotomy, and providing immediate rigid fixation while not violating the proximal femoral physis. We have used this simplified technique for valgus osteotomy in six hips in four children with excellent fixation, good maintenance of correction, and no complications. PMID- 10868362 TI - Slipped capital femoral epiphysis: is the displacement always posterior? AB - The initial direction of displacement on slipped capital femoral epiphysis is generally accepted to be posterior as a consequence of retroversion of the femoral neck. We report the case of a 15-year-old boy with slipped capital femoral epiphysis in the medial direction, confirmed by three-dimensional computerized imaging. This was associated with an elongated neck without retroversion of the femoral neck. We suggest a correlation between elongated femoral neck with increased offset of the hip and the medial direction of slip. This case also underlines the need for precise definition of deformity prior to undertaking surgical treatment. PMID- 10868363 TI - The Scottish incidence of traumatic dislocation of the hip in childhood. AB - Traumatic dislocation of the hip was reviewed over a 20-year period in Scotland. The incidence of the condition was 0.8 cases per million children annually. Posterior dislocation predominates and the presence of an associated femoral fracture must always be considered. Avascular necrosis developed in two of the 15 cases (13.3%) and is more likely if the injury is severe or if there is an appreciable delay in reduction. PMID- 10868364 TI - Intraarticular knee ganglion: a case report of an unusual cause of limping in a 4 year-old. AB - This report describes a 4-year-old boy who presented to the orthopaedic clinic with a primary complaint of limping and refusal to bear full weight on his right leg. An extensive evaluation revealed an intercruciate ganglion cyst of the knee. Diagnostic arthroscopy of the right knee was performed, and the cyst was aspirated and debrided. Magnetic resonance imaging of the knee 3 months and 1 year postoperatively showed a small remnant of the cyst adjacent to the posterior cruciate ligament. At the most recent clinical examination, 13 months postoperatively, the patient was symptom free. To the best of our knowledge, this is the youngest patient in the literature to be diagnosed with an intraarticular knee ganglion. PMID- 10868365 TI - Muscular strength after extensive operative treatment of congenital talipes equinovarus. AB - Maximum isometric voluntary contraction (MIVC) strength of muscles (extensors, plantar flexors, pronators and supinators of the foot) in children with congenital talipes equinovarus who underwent extensive operative treatment was evaluated. The first group consisted of 28 children (50 clubfeet) operated on with posteromedial-lateral release aged 7 months to 76 months (mean, 22 months), with mean follow-up period of 85 months. The second group consisted of 32 children (39 clubfeet) operated on with complete subtalar release from Cincinnati incision aged from 3 months to 50 months (mean, 11 months), with mean follow-up period of 51 months. In both groups, the muscles moving the foot in the sagittal and coronal plane showed a decreased MIVC. The greatest deficit was observed in the supinators and extensors, less in the pronators and plantar flexors. Better results were accompanied by greater MIVC, but significant relations existed in the first group between the strength of the extensors and the quality of results (better results correlated with better MIVC of the extensors). No significant differences between both patient groups were noted. For the plantar flexors, the difference of the MIVC strength between the normal and affected feet is stable; instead, for the extensors, supinators and pronators, it increases in the analyzed age interval. PMID- 10868366 TI - The radiological analysis of pes cavus deformity in Charcot Marie Tooth disease. AB - Charcot Marie Tooth (CMT) is a progressive hereditary peripheral neuropathy. The most prevalent subtype is CMT-1A, wherein patients develop a characteristic cavovarus deformity. We have reviewed a series of standing lateral foot radiographs of patients with foot deformity due to CMT, and found that the hind foot of these patients is in dorsiflexion, not equinus, and that the apparent equinus is due to plantar flexion of the forefoot on the midfoot, and actually represents a cavus deformity. PMID- 10868367 TI - Social support can fight post-heart attack depression. PMID- 10868368 TI - "Mad-cow" disease transmissable to humans. PMID- 10868369 TI - Sexual abuse of nursing home residents. AB - 1. A new subgroup of rape victims resides in nursing homes. 2. Nursing home victims can suffer both compounded and silent rape trauma. 3. Innovative therapies are needed for treating elder rape trauma. PMID- 10868370 TI - Health maintenance behaviors of individuals with severe and persistent mental illness in a state prison. AB - The purpose of this grounded theory study is to define health and health-seeking behaviors of incarcerated individuals experiencing severe and persistent mental illness (SPMI) in a state prison. The strategies used to prevent loss of control and maintain health in the prison environment were examined. Nineteen incarcerated individuals with SPMI were interviewed in a state prison. Constant comparative analysis of the data revealed that while establishing a "fit" with the core variable "loss of control" and its identified properties, differences were found in the enactment of health maintenance behaviors related to properties in the prison environment. Several of these behaviors are inconsistent with behaviors expected of individuals in a community day treatment center and reflect a need for bridging programs to facilitate entry into the "free" world. Correctional nurses and mental health providers in this system can advocate for incarcerated individuals with SPMI by developing programs that provide cost effective intermediate care and collaborating with community health systems for continuity of care. The findings related to alcohol and drug abuse among incarcerated individuals with SPMI challenges mental health providers in both correctional facilities and community-based programs to generate appropriate and effective substance abuse treatment programs for these individuals. PMID- 10868371 TI - Logs as adjuncts to therapy for adolescents in a residential psychiatric program. AB - As part of a 3-year ethnography of an adolescent residential psychiatric program, staff, patients, and occasionally family members were observed communicating using written logs. Using participant observation and opportunistic interviewing, and reading the logs, the researcher was able to determine the advantages, disadvantages, and direct value of the logs to patients and staff. Logs, when used judiciously after patient assessment, demonstrated patient responsiveness to treatment and promoted patient-centered care. Writing in logs increased patients' identification of thoughts and feelings, mirrored their developmental state, and demonstrated staff members' skills in communication. They are an invaluable adjunct to therapy for certain adolescent patients. PMID- 10868372 TI - Implementation of computer-assisted treatment planning in a prison psychiatric facility. AB - 1. An effective treatment plan must be behaviorally based and descriptive, yet concise, with clear, measurable outcomes. It should be the synergistic product of the interdisciplinary team process. 2. The use of a computer program and a firm commitment to making the team process work can generate a superior, clinically sound, practical, legible product, while liberating professionals for more clinical time. 3. One does not need to be a computer expert to use software to generate treatment plans. The program you choose must be capable of being modified by the user to respond to changing needs of patients and capabilities of staff. PMID- 10868374 TI - Speech recognition software grows up. PMID- 10868373 TI - ASHRAE 52.2-1999 helps in air filter selection. PMID- 10868375 TI - The way out of the OSHA ergonomics debacle. PMID- 10868376 TI - OHS management systems: a survey. PMID- 10868377 TI - Meetings in minutes--tailgate it! PMID- 10868378 TI - The TDL advantage. PMID- 10868379 TI - Different systems, common purpose. PMID- 10868380 TI - Noise know-how. PMID- 10868381 TI - Recognition, evaluation, and control. PMID- 10868382 TI - Military ingenuity. PMID- 10868383 TI - Safety incentive programs--effective business tools. PMID- 10868384 TI - Better methods, better results. PMID- 10868385 TI - Steering clear of disaster. PMID- 10868387 TI - Reducing pedestrian-vehicle conflicts in parking lots. PMID- 10868388 TI - ONS must set the standard for health promotion. PMID- 10868386 TI - Using circuit analyzers. PMID- 10868389 TI - Obtaining financial support for patients receiving off-label drugs. PMID- 10868390 TI - Transformation of tragedy among women surviving breast cancer. AB - PURPOSE/OBJECTIVES: To describe the process of how women with breast cancer attribute positive meaning to their illness. DESIGN: Descriptive, qualitative. SETTING: Major metropolitan area in the southwestern United States. SAMPLE: Twenty-four women diagnosed with breast cancer within the past two years. METHODS: Data collected during semistructured interviews were coded and analyzed using Grounded Theory techniques. MAIN RESEARCH VARIABLES: Process and outcomes of ascribing positive meanings to cancer. FINDINGS: The author observed a basic social-psychospiritual process of transforming personal tragedy. This process involved phases labeled as encountering darkness, converting darkness, encountering light, and reflecting light. CONCLUSIONS: Varying degrees of positive meaning can be attributed to breast cancer. Encountering the darkness and moving through the other phases are normal and adaptive. IMPLICATIONS FOR NURSING PRACTICE: Recognizing that positive meanings are independent of questions of causality but that encountering the darkness is necessary for transformation can help patients and nurses to face tragedy. Future research should seek to understand why some patients get "stuck" encountering or converting darkness. PMID- 10868391 TI - Cancer-related depression: Part II--Neurologic alterations and evolving approaches to psychopharmacology. AB - PURPOSE/OBJECTIVES: To describe evolving psychopharmacologic options for managing depression related to cancer. DATA SOURCES: Literature review and case study. DATA SYNTHESIS: Optimal recovery from depression requires early detection and selection of psychopharmacologic agents--singly or in combination--to address each depressive symptom. CONCLUSIONS: Adequate psychopharmacologic management of depression requires consistent, informed involvement of healthcare professionals, patients, and caregivers. Patients and caregivers need to be prepared to assess responses to therapy, recognize adverse effects, manage minor side effects and threats to adherence, and seek appropriate assistance for adverse events, including drug-drug interactions. IMPLICATIONS FOR NURSING PRACTICE: Oncology nurses need to take an active role in assessing, documenting, and monitoring patients' responses to antidepressants, particularly during key phases (e.g., induction, substitutions, augmentation, withdrawal). Nurses also need to take an active role in screening patients for depression, preparing patients and caregivers for psychopharmacologic management of depression, and reinforcing these competencies regularly. Oncology nurses also can be instrumental in recognizing signs and symptoms of treatment resistance and addressing these issues appropriately. Because of the rapid pace of psychopharmacologic research, oncology nurses need to update knowledge continuously to provide safe care to depressed patients with cancer. PMID- 10868392 TI - How problematic are various aspects of quality of life in patients with cancer at the end of life? AB - PURPOSE/OBJECTIVES: To identify aspects of quality of life (QOL) in patients receiving end-of-life care that are most and least problematic for patients. DESIGN: Descriptive using secondary analysis of data from an earlier QOL study. SETTING: A large not-for-profit hospice that primarily provides home care in southwest Florida. SAMPLE: 231 homecare hospice patients with cancer. METHODS: Item analysis of items on the Hospice Quality-of-Life Index. MAIN RESEARCH VARIABLES: Psychophysiologic, functional, and social/spiritual well-being. FINDINGS: Patients had the most problems in the area of functional well-being and the least problems with social/spiritual well-being. Most common physical problems included constipation and dyspnea. CONCLUSIONS: Patients with end-stage cancer are able to maintain their relationships with God and with family and friends even in the face of marked functional difficulties and troublesome physical symptoms. IMPLICATIONS FOR NURSING PRACTICE: A continued focus on the patient and family by the interdisciplinary healthcare team is warranted. PMID- 10868393 TI - Enduring suffering: a grounded theory analysis of the pain experience of elderly hospice patients with cancer. AB - PURPOSE/OBJECTIVES: To identify, describe, and generate a theoretical analysis of the pain experience of elderly hospice patients with cancer. DESIGN: Qualitative, grounded theory. SETTING: Participants' homes in rural east Texas counties. SAMPLE: 11 participants older than 65 years who were receiving services from a for-profit hospice. METHODS: Broad, unstructured, face-to-face audiotaped interviews transcribed verbatim and analyzed using constant-comparative method of analysis. MAIN RESEARCH VARIABLES: Participants' descriptions of their pain experience. FINDINGS: Suffering is the basic social problem of pain. Participants deal with suffering through the basic social process of enduring. Enduring has two subprocesses: maintaining hope and adjusting. Trusting in a higher being and finding meaning are ways of maintaining hope. Dealing with uncertainty, accepting, and minimizing pain are ways of adjusting. CONCLUSIONS: Findings provide the basis for assessment of and interventions to foster enduring. Faith, caring relationships, and strategies to decrease pain helped elderly hospice patients to endure. IMPLICATIONS FOR NURSING PRACTICE: Nurses need to recognize and value the hard work of enduring to deal with suffering. Enduring may be fostered by assisting elderly hospice patients with cancer to maintain hope and adjust. PMID- 10868394 TI - Working women identify influences and obstacles to breast health practices. AB - PURPOSE/OBJECTIVES: To identify factors contributing to participation in breast screening in working women to drive health education planning and implementation. DESIGN: Survey. SETTING: Automotive plants in southern Canada. SAMPLE: Union and nonunion women working in the plants. METHODS: Survey using "Health Care Practices: A Worksite Survey," modified for Canadian population. MAIN RESEARCH VARIABLES: Age, education, breast health practices, influences on decision to participate in breast screening, and physician gender. FINDINGS: Differences were noted among three age groups (under 30 years, 30-49 years, 50 years or older) in terms of influences and perceived barriers to the different modalities of breast screening. For clinical breast exams, women preferred an expert in breast health, regardless of whether the professional was a physician or a nurse. In all groups, the physician was noted as being very influential; however, perceptions of encouragement from the physician varied across the age groups. Perceptions of barriers to breast screening differed among the age groups and between women with male physicians and those with female physicians. Coworkers were identified as being a strong influence in the older group, whereas friends and family were identified as being more influential in the younger groups. CONCLUSIONS: Health promotion and education strategies may need to be stratified for different age groups. IMPLICATIONS FOR NURSING PRACTICE: Breast health education may need to be seen as an ongoing educational process, with the target groups being both the women and the primary healthcare professionals. The worksite has strong potential as a setting for health promotion activities. PMID- 10868395 TI - Issues in families of children with brain tumors. AB - PURPOSE/OBJECTIVES: To determine the needs of children with brain tumors and their parents/guardians and siblings during the six stages of illness: diagnosis, hospitalization, posthospitalization, adjuvant treatment, recurrence, and terminal or reported cured. DESIGN: A cross-sectional, qualitative study of families using focus group methodology. SETTING/SAMPLE: Families with a child diagnosed with a brain tumor recruited from the practices and clinics of several major teaching hospitals in the New York metropolitan area. Families traveled as far as 70 miles for the group meeting. Varying ethnic groups and family structures were represented. METHODS: Groups were separated into parents/guardians, siblings, and affected children. The children's groups were divided further into age 10 and older and younger than age 10. During the focus group, moderators followed an outline of topics identified from family interviews, the literature, and content experts that were important to families in similar situations. An assistant moderator took detailed notes, and the entire group meeting was audiotaped. Information was transcribed and analyzed using qualitative analysis techniques. FINDINGS: 11 focus groups met involving 7 affected children, 24 adults, and 19 siblings. Issues identified as important by the group members fell into four categories: Interaction With Healthcare Providers, Medical Information/Education, Healthcare Utilization and Treatment, and Psychosocial Issues. The needs of family members differed at various stages of the illness. CONCLUSIONS: The particular impact of the four major areas of concern differed by family role and stage of illness. IMPLICATIONS FOR NURSING PRACTICE: To provide optimal care, healthcare professionals must be aware of the family's composition and support systems and the impact that the illness has on individual family members at each stage of illness. PMID- 10868396 TI - [Xenograft: great hopes for an old project]. PMID- 10868397 TI - [Xenografts: expectations and perspectives]. PMID- 10868398 TI - [Utilization of xenogenic liver in human clinics. Tests and perspectives]. AB - Before the 1980s, xenogeneic livers were incorporated for extracorporeal perfusion systems to support failing liver function. Transient improvement in consciousness was seen in some patients developing stage IV encephalopathy. In the 1990s, the growing donor shortage has led to reconsidering the use of xenogeneic livers. In 1992, two baboon livers were successfully transplanted to humans; the survival times were 26 and 70 days. One auxiliary pig liver was unsuccessfully transplanted. Regeneration capacity of the liver in the xenotransplantation context in man was demonstrated using a baboon donor. At the present time physiological, microbiological and immunological problems have not been solved using xenogeneic liver. PMID- 10868399 TI - [Clinical perspectives of xenografts: encapsulated chromaffin cells and pain]. AB - Intrathecal allograft of chromaffin cells can be effectively used in replacement of more conventional therapies for treating intractable chronic pain, such as in cancer. The efficacy of this technique depends on the ability of those cells to produce analgesic opioids and on the immuno-privileged property of the central nervous system, in which rejection risks are limited. However, there are some limitations to the generalization of this new therapy, mainly due to the low number of available grafts. Thus other sources than humans have to be considered. Here we discuss the pros and cons of the xenogeneic chromaffin cells of bovine or porcine origin. Graft immuno-isolation, for example, by using cell encapsulation, seems to be unavoidable in spite of the graft site. PMID- 10868400 TI - [Clinical perspectives of xenotransplantation of islets of Langerhans]. AB - This paper describes the rationale and different approaches for developing islets of Langerhans xenotransplantation. Implementing this therapeutic strategy in the treatment of human diabetes mellitus requires careful consideration of its potential risks and benefits, taking into account the current status of the treatment of this disease. PMID- 10868401 TI - [Anti-pig xenogeneic response by human CD4+ T-lymphocytes]. AB - When considering the hypothesis of xenotransplantation, and if it becomes possible to control hyperacute and delayed vascular rejection, the recognition of porcine graft by human T CD4+ lymphocytes could still constitute a very important barrier. The direct recognition of porcine MHC class II molecules (SLA-DR and SLA DQ) by human TCR has been demonstrated in vitro. It is accompanied by a proliferative lymphocytic response, as co-stimulatory molecules are able to interact across the species barrier. In vivo, this type of recognition only applies to porcine cells with antigen-presenting functions, mainly the graft dendritic cells which emigrate into the recipient lymphoid organs. The other recognition pathway is indirect, whereby the recipient dendritic cells capture porcine xenoantigens in the graft, then process and present them to the lymphocytes in the lymphoid organs. This indirect pathway can be shown in vitro by utilizing porcine MHC class II-negative endothelial cells. In this model, human purified T CD4+ lymphocyte proliferation is tightly dependent on the presence of human antigen-presenting cells and their HLA class II molecules. As the xenogenic peptides all differ from self peptides, the indirect T-cell response will be very strong and probably difficult to control. PMID- 10868402 TI - [Xenotransplantation: physiopathology of hyperacute and differentiated rejection, and therapeutic perspectives]. AB - The mechanisms of xenograft rejection involved in closely related species of donors and recipients are not the same as in disparate combinations. When the donor and the recipient are phylogenetically close, such as the monkey and the human being, the rejection of xenografts is essentially mediated by a cellular immune response, and is clinically similar to a strong allograft rejection. When the donor and the recipient belong to widely disparate species, such as in the pig-to-human combination, the graft is destroyed by a hyperacute rejection before a cellular rejection has started. Hyperacute rejection is triggered by the fixation of the recipient's preformed antibodies on the graft endothelium, and subsequent activation of the recipient's complement. The activation of endothelial cells that ensues leads to the formation of an extensive thrombosis within the graft and to its necrosis within minutes or hours after grafting. When hyperacute rejection is overcome by means of complement inhibitors, a delayed vascular rejection occurs within 36 to 48 hours, mainly involving antibody dependent cellular cytotoxicity and contributing to the extension of the thrombosis. Both hyperacute and delayed vascular rejections can be prevented by depletion of preformed antibodies, inhibition of complement activation, or by masking the dominant xeno-epitope Gal alpha 1,3 Gal. Transgenic pigs expressing molecules that inhibit xenogeneic rejection are being produced. However, prior to any clinical use, the infectious risks, and particularly the viral risk must be evaluated. PMID- 10868403 TI - [Impact of transgenes and cloning on xenografts]. AB - Transgenesis can theoretically add a foreign gene or specifically replace an endogenous gene by another gene. Gene addition in mammals is generally achieved by DNA microinjection into one-cell embryos. Gene replacement implies homologous recombination in cultured cells which must be selected and remain capable of generating a living organism. The use of totipotent cells can, though currently in the mouse only, lead to the generation of chimeric animals transmitting the genetic modification to offsprings. The embryo cloning technique has recently allowed the use of somatic fetal cells, in which gene replacement occurred to generate living sheeps. This technique is being extended to other domestic ruminants, to pigs and rabbits. The mouse, rat and rabbit are being used as models to define the genes which should be added or inactivated to reduce rejections of xenografts. Transgenic pigs harbouring the human CD59 or the DAF genes have been obtained by several groups. Heart, kidney and isolated cells from these animals are more resistant to hyperacute rejection when grafted to experimental primates. Additional genes are to be added in the future to inhibit the other rejection mechanisms as soon as their action has been demonstrated in laboratory animals. Experiments in progress aim at inactivating the gene for Gal: 1-3-galactosyl transferase by homologous recombination in the pig genome. Transgenesis might also be used to prevent expression of endogenous pig retroviral vectors and to prepare recombinant proteins having antirejection activity from the milk of animals. PMID- 10868404 TI - [Risks of infection in xenotransplantation: what are they and how are they to be controlled?]. AB - Porcine grafts are a potential source of pathogenic agents capable of contaminating the recipient. The spread of porcine retroviruses in the patient's tissues is a major risk that must be rigorously evaluated. The control of the microbiological state of the pig donor, concerning retroviruses and other pathogens, is the necessary condition for controlling this risk. PMID- 10868405 TI - [Xenografts and the probability of viral zoonotic risk: reflections based on the sanitary control of a pig population free from specific pathogenic organisms and risk analysis]. AB - An analysis of the risk factors involved has been made and a classical approach has been adopted, consisting of at least six stages. In the context of risk assessment and identification of potential danger, the pathogenic agents in the pig have been identified and those which are potentially pathogenic to man have been indicated. The risk of spreading, and exposure to certain pathogenic agents (SEPA) varies depending on the nature of the donor organs. The risk factors should be managed by a quality-control approach, which at the sanitary level should consist of controlling all production levels in the pig population, from rearing to slaughtering, and the subsequent removal of vital organs and tissue for xenotransplantation or xenografts. The personnel involved should also undergo rigorous medical examination. PMID- 10868406 TI - [Endogenous porcine retroviruses and xenotransplantation]. AB - Vertebrate DNA contains numerous genomes closely related to retroviruses, i.e. endogenous retroviruses. While most of the retroviruses are pathogenous, endogenous retroviruses have rarely been shown as such. Endogenous retroviruses, as most of the retroviruses, are able to cross 'species barriers'. Porcine endogenous retroviruses were described in 1974. They are expressed in tissues and cells involved in transplantation, endothelial cells, for instance. They are capable of interspecific transmission, expressed in vitro in human cells; no evidence of in vivo interspecific transmission has been reported so far. As far as xenotransplantation is concerned, porcine endogenous retroviruses represent a risk of adaptation to humans of a new form of retrovirus. Such a risk mandates a close monitoring of recipients and their partners. This risk has already been taken with the use of tissues and stable blood-derived products from animals harboring in their genomes endogenous retroviruses. PMID- 10868407 TI - [Risk and risk management connected with xenograft]. AB - Transmission of an animal virus to man is probably a constant reality. Pathogenicity is is not inevitable. Some vaccines were contaminated by cell culture but remain safe. Haemorrhagic fevers, despite limited outbreaks, are often cited in the media. On the other hand, the influenza A virus has been responsible for a large mortality. The cases of human infection with simian viruses (herpes virus B, cytomegalovirus, spumavirus, immunodeficiency virus) were accidental and have always remained asymptomatic. Monkeypox virus emerges in only some outbreaks. No transmission of animal endogenous retrovirus has been described. The precautionary principle, in the face of an unquantifiable risk, supposes that preventive measures should be taken in advance to reduce the risk at each step: isolation of the animal source after hysterotomy, transport in a germ-free environment, procurement and transplantation in rigorous surgical conditions, long-term follow-up surveillance of the patient, his or her family and healthcare workers. All the phases of the xenotransplantation procedure should be taken to limit the number of places and persons coming into contact with the recipient. Surveillance means regular clinical information. Various samples should be taken from the animal source, recipient, his or her family contacts, medical staff and cryogenically preserved as 'biological memory'. A National Xenotransplantation Register should be set up to gather and share all information on incidents. PMID- 10868408 TI - [Primates: another source of grafts?]. AB - The catarrhini primates of the genus Papio, and particularly Papio anubis, may be a source of transplants whose compatibility with the human receptor doesn't induce acute rejection. The risk of zoonosis transmission is not higher than with pig transplants. PMID- 10868409 TI - [Clinical trials using cell xenografts. Their place in the treatment of fulminant hepatitis]. AB - Trials involving xenocells are mainly carried out in the context of treatment for acute liver failure. In fact, in this disease there is no accompanying chronic hepatopathy, and the liver has a significant potential for regeneration. Regarding the clinical aspect, several trials have been conducted involving patients with severe liver failure awaiting hepatic transplantation. Hepatic xenocells are the treatment of choice, as there is no normal human hepatocyte line available. Hepatic xenocells of porcine origin are used. The experimental systems adopted in man mainly consist of extracorporal systems in which the hepatocytes are present in the extracapillary space of a capillary filter. The various systems used in man have resulted in a neurological improvement. However, the risk of PERV transmission (porcine endogenous retrovirus) remains, and is a preoccupying issue. Nevertheless, no documented case of this particular risk has yet been recorded. In conclusion, clinical trials utilizing hepatic xenocells in animals and in man are ongoing, and should permit progress to be made in the field of hepatic insufficiency. PMID- 10868410 TI - [Pigs, do they have their heart in their hand? Anthropological reflections on xenografts]. AB - The therapeutic perspectives initiated by xenotransplantations are of interest since they allow a great deal of hope for all patients whose transplant delays decrease in proportion to their prognosis for survival. However, this type of therapy, while resolving the qualitative problem of transplants, could possibly lead to other dilemmas, especially the sociocultural aspects. Besides the potential risk of transmission of a pathogenic agent to the human species, xenotransplants, while creating a symbolic and actual proximity between man and animal, could abolish the ritualized and ancient distance established between the protagonists. It could be pigs or any other type of animal; the central question is one of the relationship between human and animal species. Animal husbandry and slaughtering for food are submitted to very precise cultural codes. But it would be very hasty to think that the use of an animal for therapeutic transplants would be free of all cultural connotations. PMID- 10868411 TI - ["Population and xenograft" investigation. Preliminary results]. AB - The chronic shortage of human organs is the argument for xenotransplantation. In emergencies, acceptability is closely linked to the benefit. Little information is available on attitudes towards xenotransplantation. A poll of the attitudes was carried out, based on a questionnaire with targeted questions and background information. The goal of the study was to have a better understanding of people's attitudes towards xenotransplantation and to know the eventual changes in the answers after having been given information. For 75% of the sample, xenotransplantation would be a future biotechnology. The animal sources that would be considered for xenotransplantation were the pig and monkey. A period of ten years or more is necessary for 69% of the respondents before xenografts are performed routinely; for 19%, five years or less are needed. Human organ donation should be continued, according to 90% of the sample. Roughly, 46.4% support xenotransplantation, though in the case of a life-or-death situation acceptance reaches 65.7%. This level is higher (77%) for relatives or unknown people (71%); 74% of respondents were in favour of using normal animals and a large majority (88%) support research on xenotransplantation. A good level of confidence in medical biotechnology research and practice is suggested by this study, contrary to the results of a European survey on biotechnology. PMID- 10868412 TI - [Acceptability of xenograft in type 1 diabetic patients and by the general French population]. AB - OBJECTIVE: The possibility of using pig xenografts raises the questions of their acceptability and the reasons for reluctance by patients and society, which have not been clearly investigated in Europe. RESEARCH DESIGN AND METHODS: A survey using a multiple-choice questionnaire was conducted to quantify the acceptability of pig xenografts in type 1 diabetic patients potentially concerned by xenografts (n = 377) as compared to a sample of the French population (n = 697). RESULTS: Willingness to accept a xenograft was significantly greater among diabetic patients than the general population (64% vs 54%, P < 0.001). The notion of using pig xenografts appears to be rather well accepted by the general population, and more information might improve acceptability. The acceptance of xenografts in general and pig tissues in particular was higher in diabetic patients. CONCLUSIONS: Because the general population and type 1 diabetic patients are not aware of the sanitary risks specifically related to a xenograft, the decision to use xenografts cannot be based simply on the expectations of possible recipients. The sanitary risks need to be assessed before further xenografts are performed, particularly in diabetic patients whose risk/benefit ratio is not particularly favourable. PMID- 10868413 TI - [Regulation of xenografts]. AB - The practice of xenografting, if not recent then at least still rare, explains the scarcity of countries that have legislated on this subject. In most of the industrialized countries, though, events are taking place, reflections and debates are being conducted, and attempts at a legal context, at least partial, are being developed. France is among the most committed countries in the legal framing of certain forms of xenografts. A recent legal context has just been defined by law n(o) 98-535 of July 1, 1998, concerning the reinforcement of sanitary surveillance and the control of safety of products destined for man. PMID- 10868414 TI - [Some ethical considerations, especially identity, during xenotransplantation]. AB - Provided that a pandemic due to retroviruses or prions eventually transmitted by animals is under control, in accordance with the fundamental ethical rule to serve the best interest of the patient, any organ xenotransplantation raises an important ethical dilemma: how to deliver proper and adapted information to the patient on the preservation of his identity. Xenotransplantation of any part of the brain seems to us unethical. PMID- 10868415 TI - Microbiological hazards of xenotransplantation. 1. Doubts and convictions relating to the risk of xenozoonosis. AB - The use of a xenogenic organ, tissue or cells for transplantation permits in theory the transmission of microbiological agents from one species to another. The risk of transmission of an unknown animal pathogen to man is assumed to be a public health issue. The genotype homology between human beings and non-human primates, theoretically, should increase the probability of transmission of microbiological agents. It is also assumed that pathogenicity is intensified in closely related species. Historically, most zoonoses come from species that are distant from man. Viruses are more often responsible for human disease than other animal microbial agents. Exposure of humans to animal viruses does not predicate infection. The pathogenicity of an animal virus for man may be immediate or delayed by a possible recombination of adaptive processes. The ultimate risk is the inter-human transmission of a highly pathogenic and fatal animal disease. The retrovirus possesses the enzyme which enables it to become inserted into chromosomal DNA. With an endogenous retrovirus, viral genomes are transmitted through heredity. Some of the retrovirally derived sequences in mammals are fossil viruses. It has been argued that the more closely the species are related, the less likely the retrovirus is to be transmitted, because of xenotropism. PMID- 10868416 TI - Microbiological hazards of xenotransplantation. 2. Facing the risk. AB - The high number of xenotransplantations that have been carried out around the world since the beginning of the century, combined with the quantity of animal experiments that have exposed man to animal viruses under similar conditions, should be measured against the low incidence of known pathological consequences. The pathogenic potential of animal viruses for man, then, is unpredictable; and while the risk is acknowledged, it cannot be defined. Unless no benefit can be in xenotransplantation, the application of the precautionary principle to the extent of a moratorium seems to be excessive. The precautionary principle offers possible modes of action in the face of an unquantifiable risk. In the event of xenotransplantation, precautionary measures should be taken in advance, in order to limit risk to the maximum extent. The risk is reduced and the benefit/risk ratio becomes higher, rendering the challenge acceptable. Preventive measures should be taken as the knowledge grows about possible modes of transmission. Guidelines of good practice should be implemented at each step of the xenotransplantation process: birth of animal source by hysterotomy in a 'Specific Pathogen-Free' (SPF) centre rearing in incubator then in isolated conditions (SPF centre) transportation to and rearing in the procurement site (SPF hospital special wing) surgical procurement and xenotransplantation (hospital) post surgical reanimation (intensive care unit) convalescence (isolated room- hospital) long-term recipient follow-up (home) All these steps should be taken in such a way as to limit the number of persons coming into contact with the recipient. PMID- 10868417 TI - Xenotransplantation in Sweden. PMID- 10868418 TI - Summary of principles of good practice for the production of pigs used for xenotransplantation. A report of the expert committee on xenotransplantation. Groupe d'Experts Xenogreffes. AB - The choice and selection of pigs used as donors of organs or tissues are founded on the concept of quality assurance of the animals included in the original herd. The rearing conditions in this herd have to guarantee a defined health status. All the stages of production, rearing, slaughter and collection of organs or tissues used for xenotransplantation or xenograft have to be fully controlled in regard to health. PMID- 10868419 TI - [Clinical perspectives of renal xenotransplantation]. PMID- 10868420 TI - [Treatment of neurodegenerative diseases]. PMID- 10868421 TI - [The problem of microbiological risks]. PMID- 10868422 TI - [Xenotransplantation: a program for development]. PMID- 10868423 TI - [Xenografts and public health: international political perspectives]. PMID- 10868424 TI - [Ethics and xenografts: concerning some major principlesi]. PMID- 10868425 TI - [Xenograft and Islamic bioethicss]. PMID- 10868426 TI - [Xenografts: an examination by a seniors]. PMID- 10868427 TI - Office International des epizooties, presentation. PMID- 10868428 TI - Prevention of pediatric injuries: so much to do, so little time. PMID- 10868429 TI - Diagnosis and management of common toxic ingestions and inhalations. PMID- 10868430 TI - The "fatal four" indoor air pollutants. PMID- 10868431 TI - Bites, stings, and other painful things. PMID- 10868432 TI - Drowning and near drowning: a pediatric epidemic. PMID- 10868433 TI - Pediatric burns: management of thermal, electrical, and chemical burns and burn like dermatologic conditions. PMID- 10868434 TI - The master's touch. PMID- 10868435 TI - Thoracic outlet syndrome. AB - Thoracic outlet syndrome (TOS) is an often misdiagnosed cause of neck, shoulder, and arm disability. Neurovascular compression may be seen in the interscalene triangle, costoclavicular space, or posterior to the pectoralis minor, although any cause of abnormalities of shoulder girdle alignment may cause a localized area of brachial plexus compression. Nerve compression in this way may lead to upper extremity weakness, pain, paresthesias, and numbness. A careful and detailed medical history and physical examination are essential to proper identification of thoracic outlet syndrome, which remains primarily a clinical diagnosis. Diagnostic testing may differentiate other causes of pain or neurologic symptoms of the upper extremity from TOS. Clinical management is often challenging. PMID- 10868436 TI - Visual estimation of angles by orthopedic surgeons. AB - A literature review indicates that the exactness of visual estimation of angles has not adequately been addressed. This study used a series of angles to examine how accurately and consistently practicing orthopedic surgeons were able to visually estimate angles. The data indicate that orthopedic surgeons were able to visually estimate angles to within 10 degrees 93.1% of the time and to within 5 degrees 64.6% of the time. Repeat measurements 6 weeks later were within 5 degrees of the initial responses 82.2% of the time and within 10 degrees of the initial responses 94.5% of the time. The number of years in practice or in training was irrelevant to the validity and reliability of the estimates. Acute angles of 31 degrees or less were consistently overestimated. PMID- 10868437 TI - Prognostic value of histologic tumor necrosis assessment in osteogenic sarcoma of bone. AB - To assess the prognostic value of tumor necrosis in osteogenic sarcoma of bone, we designed a retrospective study of 18 patients with classic osteogenic sarcoma (OGS) in which several factors were considered as the common criteria of inclusion. Forty percent of patients with > or = 95% necrosis related to chemotherapy of their primary tumor experienced metastatic disease and/or tumor recurrence during their follow-up, while 50% of those with < 95% necrosis had a disease-free period of > or = 5 years. Tumor necrosis related to chemotherapy in OGS does not seem to represent, as a single predictor of disease-free survival, an accurate clinical prognostic indicator. Further clinical and epidemiologic studies are needed on larger series of patients with strict criteria of inclusion to confirm our results. PMID- 10868438 TI - Anatomic evaluation of the first three sacral vertebrae and dorsal screw placement. AB - Although the anthropometric studies of the first sacral vertebra have been previously described, there are few studies that define and compare the morphological features of the S-1 to S-3 vertebrae. Twenty sacra were directly measured, and then digitized images of macroscopic cross-sectional specimens were measured by using Color Image 1.31 software. Measurements, including parameters from sacral vertebral bodies, sacral canal, and auricular articular surfaces of the first three sacral vertebrae, were obtained. The contributions by the first, second, and third sacral vertebrae to the auricular articular surface area were 50%, 35%, and 15%, respectively. The parameters related to the S-1 and S-2 medial and lateral screw placement were also determined. All measured parameters of the upper three sacral vertebral bodies and the sacral canal gradually decreased from S-1 to S-3. This study provides morphometric data of the first three sacral vertebrae that may prove helpful for better understanding the unique anatomy of the sacrum and transpedicular and lateral mass screw placement. PMID- 10868439 TI - Unilateral absence of the clavicle with rapidly progressive scoliosis in an 8 year-old. AB - We report a possible association of unilateral absence of a clavicle with rapidly progressive scoliosis. Cleidocranial dysplasia (CCD) is an autosomal dominant disorder that is characterized by defective bone formation. The clavicle, pelvis, and skull are the most commonly affected bones. A review of the literature found two cases of CCD and scoliosis. Unilateral absence of the clavicle in association with rapidly progressing scoliosis has not been previously reported. Review of the patient's charts and radiographs from age 8 to 17 years, 5 years after treatment with posterior spinal instrumentation is presented, together with a review of the literature. Our patient initially presented without any spinal deformity until age 9, when she had a 10 degree curve between C-8 and L-T. Eighteen months later, the curve progressed to 52 degrees, Risser 1. Associated anomalies include posterior-element hypoplasia of the thoracic spine and posterior fusion of C4-6. She was treated with posterior spinal instrumentation from C-8 to L-4 without complications. Correction was maintained at 5-year follow up. There may be an association between unilateral absence of the clavicle and rapid progression of scoliosis in immature children. We hypothesize that the asymmetrical influence of the unilateral absent clavicle may have played a causative role in her rapidly progressive scoliosis. PMID- 10868440 TI - Treatment of cubitus varus with osteotomy and Ilizarov external fixation. AB - A cubitus varus deformity, secondary to a supracondylar fracture, was treated with a short oblique osteotomy and the Ilizarov external fixator. An excellent anatomic and functional outcome resulted. This method may prove to be of significant value in the treatment of this difficult deformity. PMID- 10868441 TI - Severe bilateral internal tibial torsion in an adult, corrected by O'Donoghue's derotation osteotomy. AB - A case report of an adult with severe bilateral internal tibial torsion is presented. The deformities were corrected by a modification of the tibial osteotomy described by O'Donoghue in 1939. The osteotomies healed well, and the deformities were completely corrected. PMID- 10868442 TI - Cost effectiveness of deep venous thrombosis prophylaxis after hip fracture. AB - Patients undergoing hip fracture repair are at significant risk for deep vein thrombosis and pulmonary embolism in the postoperative period without appropriate prophylaxis. Agents available in the United States that have undergone clinical trials as pharmacoprophylaxis for this indication include warfarin, dalteparin, and danaparoid. Safety and efficacy data from these trials were used to determine the most cost-effective agent for routine deep vein thrombosis prophylaxis in patients with hip fractures. Incremental cost-effectiveness ratio calculations demonstrate that warfarin dosed to an international normalized ratio of 2-2.7 is currently the most cost-effective agent in these patients. PMID- 10868443 TI - A modified technique for insertion of a unit rod into the pelvis. AB - The use of a pedicle probe facilitated insertion of a unit rod into the ilium of five patients with neuromuscular scoliosis. Compared to the standard technique, the pedicle probe method greatly reduced soft-tissue dissection and also provided better perception of rod alignment and cortical perforation. PMID- 10868444 TI - Screening for bone metastases. PMID- 10868445 TI - Laparoscopic surgery for uncomplicated diverticulitis: advantages? PMID- 10868446 TI - Activation of genes for superoxide dismutase, interleukin-1beta, tumor necrosis factor-alpha, and intercellular adhesion molecule-1 during healing of ischemia reperfusion-induced gastric injury. AB - BACKGROUND: Ischemia followed by reperfusion (I/R) induces gastric lesions, probably due to excessive formation of free radicals, but the role of the scavenger of these radicals, proinflammatory cytokines such as interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), in the healing of these lesions has not been extensively studied. It is also unknown whether expression of intercellular adhesion molecule-1 (ICAM-1), which mediates neutrophil-induced injury and neutrophil infiltration, is involved in the recovery from I/R lesions. METHODS: I/R lesions were induced in Wistar rats by applying a small clamp to the celiac artery for 30 min (ischemia phase), followed by the removal of the clamp for 60 min (reperfusion phase). The influence of I/R on gastric secretion was also tested in rats equipped with a gastric fistula (GF) without or with the exposure to a standard period of I/R. Two series of rats (A and B) were used to determine the effects of exogenous and endogenous superoxide dismutase SOD (series A) and allopurinol, a xanthine oxidase inhibitor (series B), on the mucosal recovery from the gastric lesions induced by I/R. The animals were killed immediately after the exposure to I/R (0 h) and at 3 h, 24 h, or 3, 5, or 10 days after this I/R, the area of gastric lesions being measured by planimetry, and the gastric blood flow (GBF) determined by the H2 gas clearance method. Blood was withdrawn for measurement of plasma IL-1beta and TNF-alpha levels with enzyme linked immunosorbent assay, and plasma gastrin with radioimmunoassay. Biopsy samples of oxyntic mucosa were taken for the assessment of SOD, IL-1beta, TNF alpha, and ICAM-1 mRNAs by reverse-transcription polymerase chain reaction and Southern blot. RESULTS: Exposure to I/R resulted in acute gastric erosions, with the maximal increase of the area of these lesions observed 3 h after the end of I/R. This effect was accompanied by a decrease in the GBF, a significant increase in blood free radicals and plasma gastrin increments, and almost complete suppression of gastric secretion. Starting 24 h after I/R, the gastric superficial lesions progressed into deeper ulcers that healed progressively within 10 days, and this was accompanied by gradual restoration of the gastric secretion and the GBF. Treatment with SOD and allopurinol accelerated significantly the healing of I/R erosions, and this effect was accompanied by a significant increase in the GBF and the attenuation of blood free radicals. At 0, 3, and 12 h after I/R a significant decrease in SOD mRNA was observed, whereas expression of TNF-alpha, IL-1beta, and ICAM-1 showed a progressive increase starting immediately after I/R, reaching a maximum on day 3. The plasma level of TNF-alpha and IL-1beta started to increase on day 3 and peaked on day 5 after I/R, being still significantly higher at day 10 than that measured in the vehicle treated control gastric mucosa. On day 10 the gastric ulcers were almost completely healed, and a decrease in the expression for TNF-alpha, IL-1beta, and ICAM-1 mRNA and an increase in the expression of SOD mRNA were observed. CONCLUSIONS: 1) exposure to I/R produces gastric lesions mediated by the excessive formation of free radicals, resulting in suppression of both gastric microcirculation and secretory activity of the stomach; 2) SOD and allopurinol accelerate the healing of I/R lesions, probably due to suppression of oxygen free radicals and improvement of gastric microcirculation; and 3) the upregulation of SOD mRNA, with subsequent increase in the SOD production and local release of IL 1beta and TNF-alpha, may activate ICAM-1 expression and neutrophil infiltration, which appear to play an important role in the progression of I/R-induced acute gastric erosions into chronic ulcers. PMID- 10868448 TI - Positive effect of rebamipide on gastric permeability in mice after eradication of Helicobacter felis. AB - BACKGROUND: Despite Helicobacter pylori eradication, gastric inflammation persists for months or years. Preliminary results have indicated that an increase in gastric permeability could be one reason. Our aim was to evaluate the effect of a mucosal protective drug, rebamipide, on gastric permeability in a model of H. felis-infected mice. METHODS: Thirty-three C57/BL6 mice were inoculated with H. felis, and seven controls were kept non-inoculated. After 2 months 20 infected mice were treated with omeprazole, amoxicillin, and clarithromycin for 1 week and then for 4 weeks with either rebamipide (n = 9) or placebo (n = 11). The 13 remaining mice were kept untreated. After cessation of treatment, a fragment of antrum obtained from each mouse was mounted in a small Ussing chamber to study the electric resistance of the tissue (R) and antral permeability. RESULTS: No modification of the paracellular permeability (R, JNa, JMan) was observed in either group. However, the transcellular permeability to horseradish peroxidase (HRP) was significantly increased in H. felis-infected non-treated mice (1131 +/- 463, 948 +/- 339, and 182 +/- 312 ng/h x cm2) as compared with non-infected controls (469 +/- 262, 458 +/- 261, and 10 +/- 6 ng/ h x cm2, for J3H-HRP, JD, and JHRPi, respectively) (P < 0.003). Eradication of the bacteria by antibiotics without subsequent treatment with rebamipide led to a non-significant decrease in the HRP fluxes. However, when rebamipide was used after the antibiotic treatment, a significant (P < 0.01) decrease in HRP fluxes as compared with non-treated mice was observed. CONCLUSIONS: These results confirm that gastric permeability to macromolecules remains increased despite H. felis eradication and show that rebamipide can facilitate the normalization of gastric permeability to macromolecules after bacterial eradication. PMID- 10868447 TI - Gastroduodenal tolerance of 75 mg clopidogrel versus 325 mg aspirin in healthy volunteers. A gastroscopic study. AB - BACKGROUND: Clopidogrel is a new antiplatelet agent that offers increased protection over aspirin in preventing vascular ischaemic events in patients with symptomatic atherosclerosis. In a large, randomized, international study of clopidogrel and aspirin (n = 19,185 patients) clopidogrel was associated with a lower incidence of gastrointestinal adverse events, including gastrointestinal haemorrhage and hospitalizations because of gastrointestinal haemorrhage. The aim of the study was to determine whether macroscopic differences in the gastric mucosa between aspirin- and clopidogrel-treated subjects could be detected by gastroscopy after short-term treatment. METHODS: Thirty-six healthy volunteers were randomized in a double-blind, double-dummy, parallel design, to 75 mg/day of clopidogrel or 325 mg/day of aspirin for 8 days. Gastroscopy was performed at base line before administration of study drug and directly after treatment completion. Gastroduodenal effects were measured in accordance with a modified Lanza scale. RESULTS: At base line no difference between the groups was detected (median Lanza score, 0.0 in both groups). At the end of treatment the aspirin group showed a median score of 7.5, and the clopidogrel group showed an unchanged median score of 0.0 (P < 0.001). In the aspirin group 13 individuals reported 19 adverse events versus 8 individuals and 13 adverse events for clopidogrel, with approximately half of the adverse events being gastrointestinal in each group. No serious adverse events were reported. CONCLUSION: In contrast to aspirin, short term treatment with clopidogrel does not induce macroscopic changes in the gastroduodenal mucosa. The study results show that in patients without gastroduodenal disease clopidogrel, but not aspirin, does not induce any gastroscopically evident erosions during short-term treatment. PMID- 10868449 TI - Relationship between serologically diagnosed chronic atrophic gastritis, Helicobacter pylori, and environmental factors in Japanese men. AB - BACKGROUND: Whereas chronic atrophic gastritis is known to be an intermediate stage in gastric carcinogenesis, information is sparse about factors associated with this precancerous lesion except for Helicobacter pylori. METHODS: In a cross sectional study of 566 men aged 50-55 years in the Japan Self-Defense Forces, we examined the relation of H. pylori infection, smoking, alcohol use, and dietary factors to the prevalence of chronic atrophic gastritis as determined by serum pepsinogen I and pepsinogen II (I/II ratio < 3.0. and pepsinogen I < 70 ng/ml). Chronic atrophic gastritis was classified as severe when the pepsinogen I/II ratio was < 2.0, and as moderate otherwise. RESULTS: The overall prevalence of chronic atrophic gastritis was 35.7% (202 of 566). The seropositivity of H. pylori was associated with a 10-fold increase in the risk of chronic atrophic gastritis, and the association was much stronger for moderate atrophic gastritis. Neither cigarette smoking nor alcohol consumption was related to the overall risk of chronic atrophic gastritis. Consumption of vegetables and fruits was each unrelated to chronic atrophic gastritis whether examined as a whole or separately for moderate and severe atrophic gastritis. Green tea was related to decreased risk of severe atrophic gastritis, although not statistically significant, whereas garlic consumption showed no protective association. CONCLUSIONS: The findings corroborate that H. pylori infection has an important role in the development of chronic atrophic gastritis in middle-aged Japanese men. Green tea consumption may be protective against the advance of atrophic gastritis. Vegetables, fruits, or garlic had no protective effect against the development of atrophic gastritis in the study. PMID- 10868450 TI - Gastric metaplasia in the duodenal bulb shows increased mucosal interleukin-8 activity in Helicobacter pylori-positive duodenal ulcer patients. AB - BACKGROUND: Although increased levels of interleukin (IL)-8 are known to be associated with infiltration of neutrophils in the gastric mucosa with Helicobacter pylori infection, no study has investigated the relationship between local IL-8 levels and neutrophil infiltration in the duodenal mucosa of patients with duodenal ulcer (DU). METHODS: Duodenal mucosal biopsy specimens with and without gastric metaplasia (GM) were obtained from patients with DU and controls with an endoscopic methylene blue (MB) staining method. Levels of IL-8 secreted in the organ cultures of biopsy specimens were measured with an enzyme-linked immunosorbent assay. The number of myeloperoxidase-positive neutrophils infiltrating the lamina propria was determined in immunohistochemically stained tissue sections. RESULTS: Histologic assessment showed that there was a strong correlation between the absence of endoscopic MB staining and the extent of GM. The levels of IL-8 in both duodenal and antral mucosal tissues were significantly higher in patients with H. pylori infection than in those without infection. In patients with DU the duodenal mucosal tissues with GM (MB-unstained mucosa) showed significantly higher levels of IL-8 than those without GM (MB-stained mucosa) or the antral mucosa. The number of neutrophils showed similar variations among DU and control patients with a positive correlation with IL-8 activity. The levels of IL-8 and the number of neutrophils decreased after H. pylori eradication in both duodenal and antral mucosal tissues, and these changes were more remarkable in the duodenal mucosal tissues with GM. CONCLUSIONS: Increased IL-8 activity in the duodenal mucosa with GM may be important for ulcerogenesis in H. pylori-positive DU patients. PMID- 10868451 TI - Gastrointestinal motor function in patients with portal hypertension. AB - BACKGROUND: Existing data on gastric emptying and small-intestinal transit rates in portal-hypertensive patients are scarce and contradictory, and so far, the motor function of the colon has not been assessed in these patients. In this study we evaluated the propulsive effect of all main segments of the gastrointestinal tract in patients with well-characterized portal hypertension. METHODS: Eight patients with a postsinusoidal hepatic pressure gradient of at least 13 mmHg and eight age- and sex-matched healthy controls participated in the study. Gastric emptying, small-intestinal transit, and colonic transit rates were evaluated in all subjects by means of a gamma camera technique. The technique was also used to measure the frequency of antral contractions. RESULTS: No difference was observed in gastric mean emptying time or small-intestinal mean transit time of liquid and solid markers between patients and controls. After 24 h, however, the geometric center of the liquid marker had a more caudal localization in the colon of the patient group than in the controls (P = 0.04); that is, the patients had a faster colonic transit. No difference was found in the frequency of antral contractions 45 min after the test meal between patients and controls. CONCLUSIONS: These data suggest that the colonic transit is often accelerated in patients with portal hypertension, whereas the motor function of the stomach and the small intestine is unaffected. PMID- 10868452 TI - Stimulation of the small intestine by nutrients in relation to phase of the migrating motor complex. AB - BACKGROUND: The relationship between the preceding phase of the migrating motor complex (MMC) and postprandial motility in the small intestine was studied. METHODS: In eight healthy subjects small-bowel manometry was performed, and a 55 ml caloric liquid bolus (280 kJ) containing paracetamol and 14C-D-xylose was instilled into the duodenum during phase I and late phase II of the intestinal MMC, respectively, in randomized order. Blood samples were drawn at regular intervals and analysed for insulin, gastrin, glucose, paracetamol, and 14C-D xylose. RESULTS: After bolus administration during late phase II a phase-III-like activity succeeded by quiescence occurred in the duodenum in seven of eight subjects, whereas administration during phase I initiated irregular contractions in seven of eight subjects (P < 0.05). The caloric bolus induced a significant increase in serum insulin and gastrin. Areas under the curves for serum insulin, gastrin, glucose, paracetamol, and 14C-D-xylose were not modulated by the preceding phase of the MMC. CONCLUSIONS: The present study shows that a nutrient bolus instilled into the intestinal lumen induces MMC-like activity when administered during late phase II. These findings provide further evidence of interference between MMC and postprandial motility. PMID- 10868453 TI - Mucosal in vitro permeability in the intestinal tract of the pig, the rat, and man: species- and region-related differences. AB - BACKGROUND: The barrier properties of the gastrointestinal mucosa may be studied by measuring its permeability to different-sized marker molecules. Owing to difficulties in obtaining human tissue it is, however, often necessary to extrapolate findings from experimental animals to man. The aim of the present study was to compare regional intestinal mucosal permeability in man, the rat, and the pig, using the same marker molecules and in vitro technique. METHODS: Segments from jejunum, ileum, colon, and rectum were mounted in Ussing diffusion chambers, and the mucosa-to-serosa passage of 14C-mannitol, fluorescein isothiocyanate (FITC)-dextran 4,400, alpha-lactalbumin, ovalbumin, and FITC dextran 70,000 was studied. RESULTS: Irrespective of species or intestinal region an inverse relationship between the molecular weight of the markers and the permeability was seen. The mannitol permeability was higher in the small intestine than in the colon in man, whereas the rat showed a higher permeability in the ileum than in the jejunum and colon. The FITC-dextran 4,400 permeability was higher in all intestinal regions in the rat than in man and the pig. The macromolecules showed low permeability with no regional differences. CONCLUSIONS: The results showed differences between intestinal regions and between species. Permeability data from the pig correlated fairly well with those of man, whereas the rat differed, making it difficult to extrapolate from the rat to man. PMID- 10868454 TI - Determination of anti-omega-gliadin antibodies in serologic tests for coeliac disease. AB - BACKGROUND: Serologic detection of coeliac disease in the general population or in subjects belonging to risk groups implies the use of a test with high efficiency, large-scale use, and low cost. The enzyme-linked immunosorbent assay (ELISA) technique is the most appropriate assay for performing this kind of studies. Even though anti-gliadin determination has been the test most frequently used as the first step in screening procedures, many false-positive results produced a low-specificity test. In a previous work a selective recognition of omega-gliadins, mainly by IgA antibodies, was observed. Results also indicated that omega-gliadins can be useful as antigens in serologic detection of coeliac disease. We therefore wanted to analyse the anti-gliadin antibody reactivity by using purified gliadins and to evaluate the actual performance of the anti-omega gliadin antibody test. METHODS: A population consisting of 105 coeliac patients, 81 healthy controls, and 73 subjects in a disease control group was analysed. Anti-endomysium (EMA), both IgG and IgA anti-omega-gliadins, and anti-tissue transglutaminase (tTG) antibodies were determined. RESULTS: Concordant results, positive and negative, in the EMA and IgG and IgA anti-gliadin determinations were observed in 220 of 259 samples from the total population analysed. The three assays showed high efficiency, being 96.9%, 90.7%, and 91.1% for EMA and anti omega-gliadins IgG and IgA, respectively. Anti-tTG determination was performed on 103 samples (69 controls and 34 coeliac patients), finding 4 false results (2 false positive and 2 false negative), whereas anti-omega-gliadins showed 10 false results (5 false negative and 5 false positive), 3 of which were coincident with anti-tTG determination. To compare the reactivity of anti-gliadin antibodies, alpha-, beta-, gamma- and omega-gliadins were isolated under non-denaturing conditions by acid preparative electrophoresis and cation-exchange fast protein liquid chromatography (FPLC) and used in an indirect ELISA test. The composition of these fractions was analysed by means of capillary electrophoresis, showing no cross-contamination among them. CONCLUSIONS: The comparison of results using purified gliadins shows that omega-gliadins present a differential reactivity that has not previously been documented. Results using omega-gliadins isolated by either preparative electrophoresis or FPLC were similar. Tests using the purified omega-gliadin fraction present the best performance when anti-gliadin antibodies are evaluated. PMID- 10868455 TI - The effect of dexamethasone treatment on murine colitis. AB - BACKGROUND: Corticosteroids are used as anti-inflammatory drugs in the treatment of inflammatory bowel disease. We wanted to know whether dexamethasone (DEX) treatment could prevent dextran sulphate sodium (DSS)-induced colitis in mice. METHODS: Acute colitis was induced after oral administration of 10% DSS for 2 days. Controls received normal tap water. Five days before and during DSS or tap water exposure half the group was treated with 0.06 mg/day DEX, and the other half received saline. After the mice had been killed, macroscopic observation and histologic evaluation were used to determine the degree of colonic inflammation. RESULTS: The macroscopic score was significantly increased in untreated DSS mice (P < 0.001). The induction of colitis was not prevented by DEX pretreatment (5.9 +/- 0.9 versus 4.2 +/- 0.6; NS). In addition, the macroscopic scores of DEX treated controls were significantly increased (1.8 +/- 0.2 versus 0.7 +/- 0.2; P = 0.007), which suggests that DEX has a stimulating effect on colitis induction. The histology score was significantly increased in untreated DSS mice compared with controls (P = 0.016). Analogous to the macroscopic scoring results, the histology score of DEX-treated controls was significantly increased compared with untreated controls (P = 0.046). CONCLUSIONS: Pretreatment with dexamethasone did not prevent the induction of acute DSS colitis, reflected by both aggravated macroscopic and histologic inflammation scores. PMID- 10868456 TI - Deficiency of the intestinal growth factor, glucagon-like peptide 2, in the colon of SCID mice with inflammatory bowel disease induced by transplantation of CD4+ T cells. AB - BACKGROUND: Glucagon-like peptide 2 (GLP-2) is produced in endocrine L-cells of the intestinal mucosa. Recently, GLP-2 was found to stimulate intestinal mucosal growth. Our objective was to study the content of GLP-2 in the large intestine in a murine model of T-cell-induced inflammatory bowel disease. METHODS: Inflammation was induced by adoptive transfer of CD4+ blast T cells from BALB/c mice to SCID mice. The amount of GLP-2 (1-33) was measured with a specific, NH2 terminally directed radioimmunoassay in tissue extracts from the large intestine of transplanted mice developing colitis and from BALB/c and SCID control mice. RESULTS: In the middle and descending colon segments showing the most severe signs of inflammatory lesions in the CD4+ T-cell-transplanted mice, the amount of GLP-2 was significantly lower than in similar colon segments in both untransplanted SCID mice and normal BALB/c mice (P = 0.0013 and 0.0033). In the descending colon the amount of GLP-2 was 6.7 +/- 1.0 pmol/g protein in the CD4+ transplanted mice compared with 68.4 +/- 20.3 and 42.7 +/- 4.3 in the two groups of control mice. Similar findings were made with regard to the contents of the two other proglucagon-derived intestinal peptides, glicentin and GLP-1. CONCLUSION: The amount of GLP-2 is markedly reduced in the colon of mice with a T cell-induced inflammatory bowel disease histopathologically resembling both Crohn disease and ulcerative colitis. This observation may provide a pathophysiologic rationale for administration of GLP-2 as a trophic factor in inflammatory bowel disease. PMID- 10868457 TI - Melanoma antigen genes 1 and 2 are differentially expressed in human gastric and cardial carcinomas. AB - BACKGROUND: MAGE genes encode for tumor-rejection antigens and are expressed in tumors of different histologic types but not in normal tissues, with the exception of testis and placenta. The aim of this study was to evaluate the frequency of MAGE-1 and -2 expression in gastric and in cardial carcinomas; these conditions have been described as two distinct diseases, having different etiologies, epidemiologic patterns, and gene mutations. METHODS: Two groups of patients were studied: patients with distal gastric carcinoma and patients with carcinoma of the cardia. A group of patients with intestinal metaplasia in the gastric mucosa and controls were also included. All of them underwent upper GI endoscopy. Paired biopsy specimens were taken for routine histology and for RNA extraction, to study the expression of MAGE-1 and -2 genes. RESULTS: None of the intestinal metaplastic samples or controls expressed MAGE-1 and -2 at detectable levels. Whereas 40% of the gastric cancer patients expressed either MAGE-1 or -2, 26.6% transcribed both. In the cardial cancer group, 20% of the cases expressed at least one MAGE, and only 6.6% expressed both genes. These results might reinforce the concept that cancer of the cardia is a distinct neoplastic disease with regard to esophageal and gastric (distal) carcinomas. CONCLUSIONS: Here we show that MAGE gene expression occurs in advanced stages of gastric and cardial cancer and therefore appears to be a late event. This might point to a reconsideration of their potential role in cancer immunotherapy. PMID- 10868458 TI - Identification and quantification of aberrant crypt foci in the colon of Min mice -a murine model of familial adenomatous polyposis. AB - Min mice are heterozygous for a nonsense mutation in the murine adenomatous polyposis coli (APC) gene and spontaneously develop multiple intestinal neoplasms similar to the familial adenomatous polyposis (FAP) syndrome in humans. Aberrant crypt foci (ACF) are assumed to be preneoplastic lesions in both murine and human colon carcinogenesis and have been observed in FAP patients. Light microscopic examination of the colonic mucosa of 42 Min mice did not show even a single 'classical' ACF on the basis of previously defined criteria, specifying that they are elevated above the surrounding mucosa. However, in Min mice we discovered aberrant crypt foci of a different type, which we denoted ACF(Min). In contrast to the classical type, ACF(Min) were not elevated above the surrounding mucosa, their detection was totally dependent on methylene blue staining and transillumination, and they could not be identified with scanning electron microscopy. Histopathologic examination of ACF(Min) showed dysplastic crypts, similar to those found in larger lesions--that is, microadenomas in the Min mouse. The number of ACF(Min) increased up to the age of 6 weeks and then seemed to remain at a constant level of approximately four per colon. In conclusion, by transillumination of whole-mount preparations stained with methylene blue, we have identified and quantified small microscopic lesions that may be precursors of colonic adenomas in Min mice. PMID- 10868459 TI - Leukocyte interferon-alpha in the treatment of chronic hepatitis C in Finland. AB - BACKGROUND: To evaluate the efficacy of leukocyte interferon in previously untreated patients with chronic hepatitis C, 97 patients were enrolled in a prospective study in Finland with a stepped-care management protocol. METHODS: The treatment was initiated with 3 million units of interferon-alpha subcutaneously three times a week. At 3 months, if the serum alanine aminotransferase was still abnormal, the dose was doubled. If serum hepatitis C virus (HCV) RNA had turned negative at 6 months, the treatment was stopped; if it was still positive, treatment was continued for up to 12 months. All patients were followed up after treatment for 6 months. Altogether, 74 patients completed the treatment and follow-up periods. RESULTS: Of all the originally enrolled patients 36% (35 of 97) achieved sustained virologic response, defined as HCV RNA negativity 6 months after the end of treatment. The commonest HCV genotype among these patients was 3a, and as many as 52% of such patients achieved sustained virologic response. Thirty-two per cent of the patients had HCV genotype 1a, 1b, or a mixture of these; a sustained response was achieved in only 6% of such patients but in 50% of patients with a non-1 genotype. Adverse effects caused treatment cessation for 10% of the patients and IFN dose reduction for 20%. CONCLUSIONS: Monotherapy with human leukocyte interferon resulted in sustained virologic response in 36% of patients with chronic hepatitis C. In those infected with a HCV genotype other than 1, the sustained virologic response rate was 50%. PMID- 10868460 TI - Protein S-100beta: a biochemical marker for increased intracranial pressure in pigs with acute hepatic failure. AB - BACKGROUND: Acute hepatic failure (AHF) may cause encephalopathy. Intracranial pressure (ICP) is frequently monitored to guide therapy, but such monitoring may cause intracerebral haemorrhagic complications. We hypothesize that determination of serum levels of S-100beta, a protein synthesized in astroglial cells, will provide useful clinical information on the presence and extent of intracranial hypertension in AHF. METHODS: Continuous intraparenchymatous ICP monitoring and serial S-100beta measurements in serum were performed in 11 Norwegian Landrace pigs with surgically induced AHF and in 4 sham-operated controls. RESULTS: ICP increased hour by hour in the devascularized pigs in parallel with increased serum levels of protein S-100beta. In the sham-operated controls S-100beta was not detectable at any time point. CONCLUSIONS: Serum levels of S-100beta are increased early in experimental AHF. Determination of protein S-100beta may provide useful information on the presence and extent of intracranial hypertension in AHF. PMID- 10868461 TI - Long-term follow-up after the first episode of acute alcoholic pancreatitis: time course and risk factors for recurrence. AB - BACKGROUND: Owing to the current lack of long-term follow-up data on the recurrence of alcohol-induced acute pancreatitis (AP), we studied the pattern of recurrence and determined the characteristics of the disease to predict the recurrence. METHODS: Between 1972 and 1991, 2678 AP episodes were detected; 1555 were induced by alcohol, and 591 of them were the first episode. During the first alcohol-induced AP 29 patients died and were excluded from further analysis. Of the 562 included, 503 were men, and 59 women. Admission serum tests, severity index, development of complications, intensive care unit and hospital stay, and need for surgery were assessed. Case records were studied. The national database was used to detect admissions to other hospitals. RESULTS: Overall, 260 (46%) developed recurrent disease. Of the first relapses, 80% developed during 4 years. The recurrence rate has not changed with time. Age less than 45 years increased the risk (odds ratio (OR) = 2.42; 95% confidence interval (CI), 1.30-4.50). The risk factors of the first alcohol-induced AP associated with the development of multi-recurring pancreatitis are age <45 years (OR, 2.42; 95% CI, 1.59-13.0), 0-2 positive Glasgow criteria (OR, 2.45; 95% CI, 1.16-5.19), and arterial oxygen tension >60 mmHg (OR, 9.90; 95% CI, 1.32-74.3). CONCLUSIONS: Fewer than half of the patients develop recurrent alcohol-induced AP. Younger patients are at the highest risk of recurrence. Those whose first alcohol-induced pancreatitis episode was not severe are at a higher risk of developing multi-recurring pancreatitis. PMID- 10868462 TI - Heterotopic bone formation in two cases of colon carcinoma. AB - Heterotopic bone formation is rare in the gastrointestinal tract. We here present the clinical and pathologic details of a 56-year-old male patient with mucinous adenocarcinoma of the colon and a 70-year-old male patient with colon metastasis previously operated on for signet-ring-cell carcinoma of the stomach who was treated with radiotherapy postoperatively. Both of them showed diffuse bone metaplasia. Heterotopic bone formation is usually present with mucin-producing benign or malignant tumors. The pathogenesis of osseous metaplasia is not well known; however, it is speculated that the extravasation of mucin may have a stimulatory role. PMID- 10868463 TI - Acute haemorrhagic colitis possibly caused by Clostridium perfringens. PMID- 10868464 TI - Genetics and psychopathology of spectrum phenotypes. PMID- 10868465 TI - Atypical antipsychotics and weight gain--a systematic review. AB - OBJECTIVE: To review systematically data relating to weight changes with atypical antipsychotics. METHOD: We conducted a Medline search on October 29 1999 and covered the period 1980-99. All recovered papers were examined for further relevant reports. In addition, we wrote to pharmaceutical manufacturers and 10 practising clinicians to ask them to provide other relevant reports known to them. RESULTS: Eighty reports mentioning change in body weight were retrieved. Data relating to weight changes were of variable quality. Weight changes were indicated by a variety of measures. The majority of reports related to short-term changes. CONCLUSION: All atypical drugs, with the exception of ziprasidone, have been associated with weight increases. Clozapine seems to have the highest risk of weight gain, followed by olanzapine and quetiapine. There is probably a lower risk with risperidone, sertindole and zotepine and a still lower risk with amisulpride. Ziprasidone appears not to be associated with weight gain. In the absence of more compelling data, these rankings must be considered approximate and preliminary. Longer, more robust trials are needed. PMID- 10868466 TI - Finnish adoptive family study: sample selection and adoptee DSM-III-R diagnoses. AB - OBJECTIVE: To evaluate the genetic contribution to schizophrenia using an adoption design that disentangles genetic and environmental factors. METHOD: Finnish hospital diagnoses of schizophrenic/paranoid psychosis in a nationwide sample of adopting-away women are compared with DSM-III-R research diagnoses for these mothers. DSM-III-R diagnoses of their index offspring are blindly compared with adopted-away offspring of epidemiologically unscreened control mothers. RESULTS: Primary sampling diagnoses of index mothers were confirmed using DSM-III R criteria. Lifetime prevalence of typical schizophrenia in 164 index adoptees was 6.7% (age-corrected morbid risk 8.1%), significantly different from 2.0% prevalence (2.3% age-corrected morbid risk) in 197 control adoptees. When adoptees with diagnoses of schizoaffective disorder, schizophreniform disorder, schizotypal disorder and affective psychoses were added, the contrast between the index and control adoptees increased. CONCLUSION: The genetic liability to 'typical' DSM-III-R schizophrenia is decisively confirmed. Additionally, the liability also extends to a broad spectrum of other psychotic and non-psychotic disorders. PMID- 10868467 TI - Mania: differential effects of previous depressive and manic episodes on response to treatment. AB - OBJECTIVE: We compared effects of previous depressive or manic episodes on antimanic response. METHOD: In-patients in a parallel-groups, double-blind comparison of lithium, divalproex or placebo for manic episodes had comprehensive evaluations of illness history. We used non-linear curve fitting of change in Manic Syndrome Score (MSS) of the Schedule for Affective Disorders and Schizophrenia (SADS) versus previous depressive or manic episodes to investigate their relationships to MSS improvement. RESULTS: Response to lithium, but not to divalproex or placebo, worsened with increased depressive or manic episodes. More than 11 manic, or four depressive, episodes was associated with response to lithium that did not differ from placebo. Effects of previous depressive and manic episodes appeared independent, and could not be accounted for by increased rapid cycling or mixed states. CONCLUSION: At least four previous depressive or 12 previous manic episodes are associated with reduced antimanic response to lithium. PMID- 10868468 TI - The prevalence of comorbid substance misuse and its influence on suicidal ideation among in-patients with schizophrenia. AB - OBJECTIVE: To estimate the prevalence, and identify the clinical correlates of comorbid substance misuse (abuse or dependence) among readmissions with schizophrenia, particularly to establish whether comorbid substance misuse is associated with higher rates of depressive symptoms and suicidal ideation. METHOD: Over 12 months, consecutive readmissions from a catchment area psychiatric service with DSM-IV schizophrenia/schizoaffective disorder were assessed using clinical assessments of symptomatology including depression, insight and compliance. Comorbid substance misuse was diagnosed using DSM-IV criteria. RESULTS: Of 102 readmissions 40% had lifetime, while 20% had current comorbid substance misuse and were predominately young males. Comorbid substance misuse had no statistically significant impact on positive, negative or depressive symptomatology. However, those currently misusing substances reported more suicidal ideation compared with past or non-substance misusers. CONCLUSION: Readmissions with comorbid substance misuse were more likely to report suicidal ideation, and may represent a group of individuals who are at higher risk of suicide. PMID- 10868469 TI - Long-term psychiatric patients in vocational rehabilitation programmes: a naturalistic follow-up study over 3 years. AB - OBJECTIVE: This paper aims at assessing the vocational integration attained after a 3-year period by psychiatric patients who participated in different vocational rehabilitation programmes. METHOD: In the north-western German region of Westphalia-Lippe a naturalistic follow-up study was carried out on 471 patients from three different types of vocational rehabilitation programmes. The sample comprised chronically ill patients with a history of repeated and long-term hospitalization. RESULTS: After 3 years 11% of the patients were in competitive employment, 67% (still) in sheltered employment, 7% in out-patient work therapy programmes and 15% were unemployed. It is important to notice that 74% achieved their subjective rehabilitation goals expressed at baseline. CONCLUSION: Vocational rehabilitation programmes are an essential part in the treatment of people with chronic mental illness. Integration into work varies markedly while patients' satisfaction is comparably good. Competitive employment represents a realistic objective only for patients with high motivation and favourable preconditions. PMID- 10868470 TI - Service use and costs of people with dual diagnosis in South London. AB - OBJECTIVE: To compare the service use and costs of individuals who have a dual diagnosis of psychosis and substance abuse with those who have a diagnosis of psychosis but no substance abuse. METHOD: Patients with psychosis were identified and a representative sample were interviewed. Six-month service use was measured and costs calculated. Regression models were developed to predict costs from background characteristics and dual diagnosis status. RESULTS: A greater proportion of the patients with dual diagnosis used community psychiatric nurses, in-patient care and the emergency clinic. The regression analysis revealed that dual diagnosis patients had significantly higher 'core' psychiatric service costs (a difference of pound sterling 1362) and non-accommodation service costs (pound sterling 1360) than non-dual-diagnosis patients. The difference when all services were analysed was pound sterling 1046, but this was not statistically significant. CONCLUSION: Specific interventions for dual diagnosis patients should be introduced and assessed in terms of individual outcomes, service use and costs. PMID- 10868471 TI - Prevalence estimates of pathological gambling in Switzerland. AB - OBJECTIVE: The purpose of this study was to evaluate the prevalence of pathological gambling in the Swiss adult population before the introduction of new forms of gambling, and the link between pathological gambling and alcohol abuse. METHOD: 2526 telephone interviews were completed using standardized assessment instruments for identifying potential and probable pathological gamblers (SOGS) and alcohol abuse (CAGE). RESULTS: The current prevalence of probable and potential pathological gamblers were estimated to be 0.8% and 2.2%, respectively. The results also show a clear correlation between alcohol abuse and gambling behaviours. CONCLUSION: For the first time estimations are available of the Swiss prevalence rates of pathological gambling. Considering the link between gambling availability and increases in the prevalence of pathological gambling and the correlation between alcohol abuse and pathological gambling, the implications of these results for the prevention and treatment of this debilitating disorder are discussed. PMID- 10868472 TI - Visual perseveration: a new side effect of maprotiline. AB - OBJECTIVE: The aim of this paper is to describe a case of visual perseveration including palinopsia during maprotiline therapy. METHOD: A single case report. RESULTS: The patient, a 56-year-old depressive man, suffered from visual perseveration during maprotiline therapy. The visual perseveration was dose related and disappeared with reduction and cessation of the therapy. CONCLUSION: The present findings suggest that maprotiline can induce visual perseveration including palinopsia in some patients. PMID- 10868473 TI - Who's afraid of homocysteine? PMID- 10868474 TI - New insights into the role of COX 2 in inflammation. AB - Cyclo-oxygenase (COX) is responsible for the synthesis of bioactive prostanoids, the inhibition of which serves as the basis for the mode of action of clinically used nonsteroidal anti-inflammatory drugs. While there were suggestions as early as the 1970s that an inducible isoform of COX exists, it was only in the early 1990s that COX 2 was identified, cloned and sequenced. Not surprisingly, this new isoform was expressed at sites of inflammation and reported to contribute to the inflammatory response. Recently, however, evidence is emerging to suggest that COX 2 also has anti-inflammatory properties. In this review, the two faces of COX 2 are examined, with emphasis on its role in regulating inflammatory resolution, including possible mechanisms of action PMID- 10868475 TI - Regulation of renin: new evidence from cultured cells and genetically modified mice. AB - Renin, as the rate-limiting enzyme in the synthesis of the potent vasoactive peptide angiotensin II, has been studied for more than 100 years. Transgenic and knockout mice for renin and other proteins involved in renin regulation and function have recently revealed new evidence that can improve our understanding of its biological relevance. Furthermore, transgenic mice have been the source of the novel cell line As4.1. This cell line has been effective in the analysis of renin secretion and regulation because of its similarity with renin-producing juxtaglomerular (JG) cells. Renin secretion and synthesis by the JG cells of the kidney is upregulated by cAMP and downregulated by intracellular calcium. The effect of cGMP, once elevated by nitric oxide, depends on the present level of cAMP in the cells, which can be stimulatory in the presence of and inhibitory in the absence of the other cyclic nucleotides. All known effectors of renin regulation affect one of these molecules. Adenosine and ATP, released by macula densa cells in response to high salt load in the distal tubule and stretch of the JG cell by renal perfusion pressure, increase calcium. Furthermore, noradrenaline, derived from sympathetic nerve endings, and prostaglandins, generated by macula densa cells under low-salt conditions, increase cAMP. In addition to its stimulatory effect on secretion, cAMP also effectively augments renin mRNA levels by acting at the transcriptional and posttranscriptional levels. Several DNA elements in the distal and proximal promoter regions as well as in intron I have been implicated in cAMP regulation and in tissue specificity of renin gene expression. A second intracellular renin isoform, coded by the same gene but applying a different promoter located in intron I, has recently been detected. Transgenic technology will help to clarify the function of this isoform as well as some of the other unresolved aspects of renin regulation and function and may become the motor of the second century in renin research. PMID- 10868476 TI - Involvement of PTCH gene in various noninflammatory cysts. AB - Constitutional hemizygous inactivation of PTCH, the Shh signaling pathway gene that moderates the signal, manifests itself as nevoid basal cell carcinoma syndrome or Gorlin syndrome, a condition variably characterized by a number of developmental disorders and malformations, and by predisposition to some malignancies, basal cell carcinoma in particular. Loss of heterozygosity for the PTCH region was found several years ago in the epithelial lining of odontogenic keratocysts, the cyst type with highly increased incidence in nevoid basal cell carcinoma syndrome. This finding confirmed the expectations that the gene responsible for the syndrome would have a decisive role in the genesis of these cysts even when they are not syndrome related. Suggestive temporal distribution of Shh signaling, recently observed during tooth development, lead us to investigate PTCH association with dentigerous cysts, the other major noninflammatory cyst of odontogenic origin. We report here that PTCH appears to be inactivated in dentigerous cysts, suggesting that it is responsible for their genesis as well. More generally, if our similar observations of incomplete heterozygosity in this region for dermoid cysts can be interpreted as loss of heterozygosity, PTCH alterations may prove to be a necessary, and perhaps the initiating event, in formation and growth of various noninflammatory cysts. This would be consistent with our view that local PTCH inactivation can, under favorable circumstances, lead to persistent though not by itself truly aggressive cell proliferation. PMID- 10868477 TI - Determinants of plasma homocyst(e)ine in patients with nephrotic syndrome. AB - Hyperhomocyst(e)inemia is an independent risk factor for atherothrombosis in several clinical settings in which renal function is impaired, but its prevalence in the nephrotic syndrome has not been investigated in detail, even though this syndrome provides an excellent model in which to study a possible link between albuminuria, proteinuria, and hyperhomocyst(e)inemia. We obtained plasma and urine from 27 patients with biopsy-confirmed membranous glomerulonephritis presenting nephrotic syndrome and 27 matched controls and determined the concentrations of homocyst(e)ine and proteins considered putative markers of glomerular and tubular function. Hyperhomocyst(e)inemia, defined as the mean +SD of the plasma homocyst(e)ine concentration of the controls [plasma homocyst(e)ine concentration >10.8 micromol/l] was present in 26% of the patients with nephrotic syndrome but in only 7.4% of the controls. Furthermore, the degree of hyperhomocyst(e)inemia was more severe in the nephrotic patients than in the controls. The existence of renal failure, tubular damage, and, interestingly, relatively well conserved glomerular function barrier were the main predictors of increased levels of plasma homocyst(e)ine. In conclusion, hyperhomocyst(e)inemia is a frequent cardiovascular risk factor present in patients with nephrotic syndrome and renal failure, but it is not directly associated with proteinuria. PMID- 10868478 TI - Suppression of breast cancer invasion and migration by indole-3-carbinol: associated with up-regulation of BRCA1 and E-cadherin/catenin complexes. AB - Indole-3-carbinol (I3C) is a compound occurring naturally in cruciferous vegetables and has been indicated as a promising agent in preventing breast cancer development and progression. In the present study we have investigated the effect of I3C on the cell migration and invasion behavior in estrogen receptor positive MCF-7 and estrogen receptor negative MDA-MB-468 human breast cancer cell lines. Both MCF-7 and MDA-MB-468 were poorly invasive cell lines and exhibited modest invasion and migration capacity in the presence of fibronectin as the chemoattractant. I3C (50 or 100 microM) elicited a significant inhibition of in vitro cell adhesion, migration, and invasion as well as in vivo lung metastasis formation in both cell lines. I3C also suppressed the 17beta-estradiol stimulated migration and invasion in estrogen-responsive MCF-7 cells. These results indicate that anti-invasion and antimigration activities of I3C occur via estrogen independent and estrogen-dependent pathways. Moreover, I3C significantly caused a dose-dependent increase in E-cadherin, three major catenins (alpha, beta, and gamma-catenin) and BRCA1 expression. Our current finding is the first demonstration that I3C can activate the function of invasion suppressor molecules associated with the suppression of invasion and migration in breast cancer cells. Thus, clinical application of I3C may contribute to the potential benefit for suppression of breast cancer invasion and metastasis. PMID- 10868479 TI - Antitumor efficacy of regional oncolytic viral therapy for peritoneally disseminated cancer. AB - Oncolytic viral therapy is a promising new method of cancer treatment. Peritoneal dissemination of cancer is a common and fatal clinical condition seen in many malignancies, with few effective therapies available. G207, a multimutated replication-competent herpes simplex virus type-1, effectively treats disseminated peritoneal cancer. This study evaluates viral proliferation and subsequent tumoricidal effects in vitro and in vivo after regional viral delivery. In vitro studies demonstrate that G207 efficiently kills five human gastric cancer cell lines, and that permissiveness to viral replication is correlated with cytotoxicity. In a murine xenograft model of human gastric carcinomatosis, peritoneal delivery of G207 effectively kills tumor and prolongs survival. Data from quantitative PCR characterizes peritoneal clearance of virus after intraperitoneal injection, and identifies G207 replication within tumor cells in vivo, similar to in vitro proliferation. Further analysis of various organs confirms that G207 does not replicate within normal tissue after peritoneal delivery. Wild-type KOS viral replication was also demonstrated in vivo, with significant toxicity secondary to dissemination and encephalitis. In vivo viral proliferation of G207 is restricted to tumor cells, is correlated with in vitro assays, and is an important mechanism of anticancer efficacy. PMID- 10868480 TI - Hippocampal anatomy and water maze performance are affected by neonatal cryoanesthesia in rats of both sexes. AB - There is recent evidence that cryoanesthesia, commonly used during neonatal hormone manipulations (e.g., gonadectomy), has deleterious effects on the morphology of the splenium of the corpus callosum and primary visual cortex in adult rats of both sexes. (Nunez and Juraska, 1998; Nunez, Kim, and Juraska, 1998). In the present study, the effect of neonatal cryoanesthesia on the morphology of the hippocampus and dentate gyrus and on performance in the Morris water maze was investigated. Cold exposure for as brief as 30 min (5 degrees C) on Postnatal Day 1 resulted in a significant decrease in the volume of the hippocampus and in brain weight of adults. Performance on the water maze was also impaired in cold-exposed animals. This study indicates that not only morphology but also behavioral performance in adulthood are affected by neonatal cryoanesthesia. PMID- 10868481 TI - Stimulatory effects on the reproductive axis in female songbirds by conspecific and heterospecific male song. AB - Courtship vocalizations of male songbirds can profoundly enhance the reproductive physiology and behavior of conspecific females. However, no study has fully investigated the selectivity of conspecific song effects on reproductive development in birds. We studied the effects of conspecific and heterospecific song on reproductive development in domesticated (canaries) and wild songbirds (song sparrows). As expected, conspecific song enhanced follicular development. Unexpectedly, however, birds exposed to heterospecific song also underwent enhanced follicular development (compared to birds exposed to no song); conspecific and heterospecific songs were equally effective in enhancing ovarian development. In canaries exposed to 18L:6D, conspecific song induced oviposition earlier and at a greater frequency than in heterospecific and no song groups, with the fewest eggs being laid in the no song group. These results indicate that conspecific and heterospecific male song can enhance reproductive activity in female songbirds. Whether or not activation of the reproductive axis in female songbirds by heterospecific song occurs in the wild remains unclear. It is also unclear as to whether the ability of the reproductive axis to respond to heterospecific song performs a specific function, or whether it is simply a consequence of greater selection pressure acting upon behavioral responses to song. PMID- 10868482 TI - Estrus and estrogen changes in mated and unmated free-living European ground squirrels. AB - The course of behavioral and vaginal estrus and patterns of circulating estrogens were followed in free-living European ground squirrels (Spermophilus citellus) after their emergence from hibernation. Normally mating females were compared to a second group in an area where males had been removed from the population before female emergence. Both groups showed vaginal estrus, but the patterns differed. Mating shortened vaginal estrus to a 3-day period compared to 8 days in unmated females. The extent (cell number) of cell cornification during estrus and the cellular components (percentage distribution) of metestrus did not differ between the two groups. Females in the area without males had significantly higher estrogen levels during estrus and metestrus compared to those in the control area. European ground squirrels were found to be monestrous, as none of the unmated females reentered estrus after metor diestrus was detected. The prolongation of vaginal estrus in unmated females can be viewed as either a physiological inevitability or an adaptation to low mate availability. The extension is still relatively short compared to other sciurid species and perhaps a product of constraints producing a strict time frame for reproduction. PMID- 10868483 TI - Sex steroids relative to alternative mating behaviors in the simultaneous hermaphrodite Serranus subligarius (Perciformes: Serranidae). AB - This study is the first investigation of reproductive endocrinology in a simultaneously hermaphroditic teleost, the belted sandfish (Serranus subligarius). We address two questions: (1) Do steroid hormone levels vary during the spawning season or during the daily spawning cycle of sandfish? (2) Do hormone levels vary relative to an individual's phenotype-size, frequency of spawning and aggressive behaviors, and proportion of testis in the gonad? We analyzed circulating estradiol-17beta (E2), testosterone (T), 11-ketotestosterone (11KT), 17alpha,20beta,21-trihydroxy-4-pregnen-3-one (20betaS), and 17alpha,20beta-dihydroxy-4-pregnen-3-one (DHP) concentrations in a field population. Only E2 levels were significantly higher at the new and full moon, suggesting peak periods of vitellogenesis at these times. Naturally spawning sandfish were sampled every 2 h during the photophase of a 25-h period (12 pm to 1 pm the following day) and gonadosomatic index, degree of oocyte hydration and ovulation, and plasma levels of E2, T, DHP, and 20betaS were analyzed. E2 and T levels did not vary during photophase, suggesting continuous recruitment of oocytes into vitellogenesis. The 20betaS levels peaked around the time of final oocyte maturation. Since frequency of spawning behaviors changes with body size, we captured individuals of various sizes throughout the spawning season and analyzed circulating levels of hormones. 11KT and 20betaS levels increased significantly with body size. In 1992, we quantified frequency of spawning and aggressive behaviors, circulating T and 11KT levels and testicular mass relative to ovotestis mass in focal animals. 11KT levels tended to be positively correlated with frequency of courting male behavior, but were unrelated to the frequency of aggressive behavior or testis mass. Because hormone levels increased with size and frequency of each spawning behavior changes with size, we propose that sex steroids influence growth-related changes in spawning tactics of individuals. PMID- 10868484 TI - Effects of gonadotropin-releasing hormones, corticotropin-releasing hormone, and vasopressin on female sexual behavior. AB - The effects of intracerebroventricular (icv) infusion of four neuropeptides on female sexual behavior were examined in the female musk shrew (Suncus murinus). In the first experiment, (icv) infusion of 100 ng of the mammalian form of gonadotropin-releasing hormone (mGnRH) facilitated rapid display of receptivity. Gonadotropin-releasing hormone-infused females had shorter latencies to rump present and tail wag, compared with controls. In a second experiment, icv administration of the other form of GnRH present in musk shrew brain, the chicken GnRH-II form, produced no changes in female behavior relative to the control condition. In Experiment 3, icv delivery of corticotropin-releasing hormone (CRH) facilitated female sexual behavior, relative to vasopressin and controls. The females treated with CRH had shorter latencies to display rump present, tail wag, and for the receipt of the first missed intromission compared with females in the other treatment groups. Vasopressin increased female scent marking relative to that of CRH-treated females. These data indicate that neurohormones of the hypothalamic-pituitary-gonadal and the hypothalamic-pituitary-adrenal axes can facilitate reproductive behavior in S. murinus. PMID- 10868485 TI - Circulating LH levels and the response to exogenous GnRH in the common mole-rat: implications for reproductive regulation in this social, seasonal breeding species. AB - The effects of breeding season and reproductive status on male and female reproduction were investigated in the common mole-rat, Cryptomys hottentotus hottentotus, a cooperatively breeding rodent which exhibits a unique combination of seasonal breeding and a reproductive division of labor. Pituitary function was examined by measuring the luteinizing hormone (LH) responses to single doses of 2 microg exogenous gonadotrophin-releasing hormone (GnRH) and physiological saline in 69 males and 58 females from 35 wild caught colonies. Neither males nor females exhibited any apparent manifestation of season on basal LH concentrations or on pituitary sensitivity to stimulation by exogenous GnRH. The continuance of reproductive function during the nonbreeding period is essential in common mole rat males and females, as this period coincides with the period of maximal dispersal opportunity in the winter rainfall area they inhabit. Normal circulating levels of reproductive hormones in dispersing animals may aid intersexual recognition, assist pairbond formation, and thus prime animals for independent reproduction. Circulating basal concentrations of LH as well as LH levels measured in response to a single exogenous GnRH challenge were not significantly different between the reproductive and non-reproductive groups of either sex, suggest the absence of a physiologically well-defined suppression of reproduction in subordinate common mole-rats. PMID- 10868486 TI - Effects of prostaglandin F2alpha treatment on the behavior of pseudopregnant pigs in an extensive environment. AB - In seminatural environments, prepartum sows leave the herd and construct a maternal nest (a dug out hollow lined with vegetation) prior to the birth of their piglets. The endocrine drives motivating this behavior are not understood, but may involve prostaglandin (PG) F2alpha. This study examined the effect of PGF2alpha treatment on the behavior of pseudopregnant gifts housed in a large enclosure. Pseudopregnancy was induced using 5 mg/ml estradiol valerate/day im from days 11 to 15 of the estrous cycle (first day of estrus = day 0). The gifts' behavior was recorded on a control day, during which no treatment was given, and a test day (= 45.9 +/- 0.42 days of pseudopregnancy) when gilts received either 15 mg PGF2alpha (dinoprost: Lutalyse, Upjohn, Crawley, UK, n = 11) or 0.9% saline (n = 10) im at 11.00 h. PGF2alpha-treated gilts traveled further and were more frequently >10 m from the nearest pig than saline-treated animals. In the hour following injection, PGF2alpha-treated animals also showed increased frequencies of rooting and pawing the ground and stood for longer than saline-treated animals. However, gathering and carrying nest materials were not increased. These results suggest that PGF2alpha, given as a single dose to extensively housed gilts, initiated many, but not all, of the behaviors characteristic of prepartum nest building. The dose and duration of PGF2alpha treatment may have limited the observed behaviors. In addition, environmental feedback is likely to affect the degree to which some nest building behaviors are expressed. PMID- 10868488 TI - Mating season aggression and fecal testosterone levels in male ring-tailed lemurs (Lemur catta). AB - The challenge hypothesis (J. C. Wingfield, R. E. Hegner, B. G. Ball, and A. M. Duffy, 1990, Am. Nat. 136, 829-846) proposes that in birds, reptiles, and fish, "the frequency or intensity of reproductive aggression as an effect of T[estosterone] is strongest in situations of social instability, such as during the formation of dominance relationships, the establishment of territorial boundaries, or challenges by a conspecific male for a territory or access to mates" (p. 833). To determine the extension of this hypothesis to mammalian species, we tested predictions of the hypothesis in a nonpaternal, seasonal breeding, prosimian primate (ring-tailed lemurs, Lemur catta). Semi-free-ranging males were studied during periods of social stability (premating period) and instability (mating period). The annual mating season consists of several days during which males fight for access to promiscuous group females as each individually becomes sexually receptive for 1 day. Male rates of aggression were compared to fecal testosterone levels within premating and mating periods. In the premating period male rate of aggression was not significantly correlated with testosterone level. By contrast, during the mating season testosterone and aggression levels were positively and significantly correlated. However, on days just preceding estrus, male rate of aggression was not significantly correlated with testosterone, but on days of estrus, when aggressive challenges peaked sharply, testosterone and aggression were highly positively correlated. These results suggest that the challenge hypothesis applies to mammals as well as to birds, reptiles, and fish. In addition, elevations in testosterone were tightly circumscribed around days of estrus, suggesting a compromise between costs and benefits of elevated testosterone levels. PMID- 10868489 TI - Issues regarding growth hormone replacement in growth hormone deficient adults. PMID- 10868487 TI - Conversion of testosterone to estradiol may not be necessary for the expression of mating behavior in male Syrian hamsters (Mesocricetus auratus). AB - Male sexual behavior is mediated in part by androgens, but in several species, mating is also influenced by estradiol formed locally in the brain by the aromatization of testosterone. The role of testosterone aromatization in the copulatory behavior of male Syrian hamsters is unclear because prior studies are equivocal. Therefore, the present study tested whether blocking the conversion of testosterone to estradiol would inhibit male hamster sexual behavior. Chronic systemic administration of the nonsteroidal aromatase inhibitor Fadrozole (2.0 mg/kg/day) for 5 or 8 weeks did not significantly increase mount latency or reduce mount frequency, intromission frequency, ejaculation frequency, or anogenital investigation relative to levels shown by surgical controls. However, Fadrozole effectively inhibited aromatase activity, as evidenced by the suppression of estrogen-dependent progesterone receptor immunoreactivity in the male hamster brain. The JZB39 anti-progesterone receptor antibody labeled significantly more neurons in brains of sham-treated hamsters than in brains of Fadrozole-treated hamsters. These data suggest that aromatization of testosterone to estradiol is not necessary for normal mating behavior in Syrian hamsters. PMID- 10868490 TI - Advances in management of melanoma. AB - It is clear from the above that the sentinel node technique and assessment of circulating melanoma cells has provided important additional methods for staging and assessment of prognosis of patients with melanoma. Cure of the disease still depends on good quality surgery and implementation of quality control measures to ensure the adequacy of surgical removal of primary melanoma and regional lymph node metastases is fundamental to improvement in survival from the disease. Systemic medical treatments have yet to show any significant impact on the disease when given in an adjuvant setting or in treatment of metastatic disease. There is much hope that new initiatives in immunotherapy and in apoptosis research will provide more effective treatments of the disease. PMID- 10868491 TI - Risk factors for development of hepatocellular carcinoma among Australians with hepatitis C: a case-control study. AB - BACKGROUND: Older patients with cirrhosis due to hepatitis C are at risk of developing hepatocellular carcinoma (HCC), but additional risk factors may vary between countries. AIM: In the present study, we sought to identify additional risk factors for HCC among a cohort of Australian patients with chronic hepatitis C. METHODS: Case-control study of patients with advanced fibrosis stage hepatitis C who developed HCC during five-year follow up at a referral liver clinic. Cases were compared to twice the number of age-matched patients with chronic hepatitis C of similar fibrotic severity who did not develop HCC over a similar interval, using conditional logistic regression analysis (CLRA) and multivariate analysis. The main outcome measures were demographic and disease-related variables at first presentation in relation to the development of HCC. RESULTS: HCC developed in 17 cases, an annual incidence among those considered to be at risk of 2%. The duration of follow up since first assessment was comparable among the cases and 34 selected age-matched controls (4.1 and 5.2 years respectively, p=0.5). Cases were more often male (p=0.03), born in Asia (p=0.05), and had poorer liver function as indicated by serum albumin concentration (p=0.02). Anti-hepatitis B core-antibody (anti-HBc) was detected in 59% (ten/17) of cases, compared to 21% (seven/34) of the controls (p=0.01). No patient with a sustained response to interferon developed HCC during follow up. There were no significant differences in the mode of HCV transmission, HCV genotype, alcohol exposure, serum bilirubin level or prothrombin time between the cases and the controls. Although the data set was small, multivariate CLR analysis identified serum albumin < or = 35 g/L and anti-HBc positivity to be independent risk factors for development of HCC. CONCLUSIONS: Among older Australian patients (over the age of 40 years) with advanced fibrosis stage hepatitis C, the annual incidence of HCC is about 2%. Those who have low serum albumin and evidence of previous exposure to hepatitis B virus (anti-HBc positivity) appear to have the highest risk of developing HCC during follow up, but males and those born in Asia could also be at increased risk. PMID- 10868492 TI - Mini-allografts' for haematological malignancies: an alternative to conventional myeloablative marrow transplantation. AB - BACKGROUND: Considerable morbidity and mortality are consequences of the myeloablative chemoradiotherapy utilised in conventional allogeneic marrow transplantation. This has generally restricted such potentially curative treatment to patients <50-55 years with normal organ function. Recent studies suggest that purine-analogue based non-myeloablative regimens are sufficiently immunosuppressive to facilitate allogeneic donor cell engraftment. AIM: To review the outcome of HLA-compatible sibling allogeneic peripheral blood progenitor cell (PBPC) transplants using fludarabine-based conditioning in patients ineligible for a conventional transplant, with emphasis on engraftment, graft vs host disease (GVHD) and graft vs tumour effects. METHODS: Eleven patients (nine > or = 47 years age) with advanced haematological malignancies received one of three different fludarabine-based chemotherapy followed by infusion of granulocyte colony-stimulating factor mobilised PBPC. Cyclosporin-methotrexate was used as GVHD prophylaxis. RESULTS: Two patients died early; minimal non-haematological toxicity apart from mucositis occurred in the other nine. Eight of the nine evaluable patients had evidence of durable donor cell engraftment in preference to recipient cells. Acute GVHD (> or = grade 2) occurred in seven of eight patients with donor engraftment who survived beyond two months. Four patients are alive in complete remission 8-28 months post-transplant; another is alive in probable remission at four months. CONCLUSIONS: Fludarabine-based non myeloablative chemotherapy facilitates rapid engraftment of allogeneic donor cells, which produce powerful GVHD and graft vs tumour effects. This approach has the potential to extend the applicability of allogeneic stem cell transplants to patients traditionally regarded as too old or sick for conventional marrow transplants. PMID- 10868494 TI - Cerebrovascular disease as a general medicine discharge diagnosis: assessment of diagnostic agreement on case note review. AB - BACKGROUND: Accurate clerical coding of discharge diagnoses is important partly because results may be used to derive a recommended costing for hospital length of stays (LOS). Some authors think that discharge coding undertaken by clinicians will result in less diagnostic misclassification than clerical coding. This presupposes a high degree of between-observer diagnostic agreement between clinicians. AIMS: To compare discharge coding undertaken by two general physicians, for patients receiving a clerical discharge code of cerebrovascular disease. The recommended LOS was then calculated from each observer's discharge codes using the Physicians Diagnosis Related Group Working Guidebook. RESULTS: Eighty-two cases were coded as stroke by the clerical coder. Both medical coders agreed with this diagnosis in 68 (83%) of these cases. The corresponding figure for cases coded by the clerical coder as transient cerebral ischaemia was 47% (32/68) agreement between all three observers. Correcting for chance agreement between medical observers using the kappa statistic, a value of 0.64 was obtained for the combined stroke and transient cerebral ischaemia discharges, suggesting moderate diagnostic agreement. Using the clerical coder's results, the mean recommended LOS for all cases of cerebrovascular disease over the study period was calculated at 6.68 days. The corresponding values for the two medical coders were 6.68 days and 6.75 days. CONCLUSIONS: Diagnostic agreement between clinicians was moderate. Consideration of alternative diagnostic possibilities and the difficulty in determining the duration of neurological deficit were the main reasons for diagnostic disagreement. The mean recommended LOS was similar, however, when comparing results from all three observers. PMID- 10868493 TI - Influence of angiotensin converting enzyme (ACE) genotype on interpretation of diagnostic tests for serum ACE activity. AB - BACKGROUND: The discovery that an insertion/deletion (I/D) polymorphism of the angiotensin converting enzyme (ACE) gene influences the circulating concentration of ACE may have implications for the proper use of serum ACE activity measurements in screening for sarcoidosis. AIM: To determine whether the sensitivity of the serum ACE test improves if ACE genotype is taken into account. METHODS: A retrospective determination of ACE genotype and clinical diagnosis was done in 54 patients with serum ACE activity above the upper limit of the reference range for the insertion (II) genotype. ACE was measured by radioenzymatic and spectrophotometric techniques, and genotype by PCR. RESULTS: When serum ACE values determined diagnostically were related to the appropriate genotype-specific reference range, sensitivity and specificity for diagnosis of sarcoidosis were 65-70% and 58% respectively, compared to 47-57% and 77% with a reference range unsegregated for genotype. CONCLUSION: ACE genotyping may be helpful in determining the diagnostic significance of mildly elevated serum ACE activity in patients with the II and ID genotypes. PMID- 10868495 TI - Opinion of New Zealand physicians on management of acute ischaemic stroke: results of a national survey. AB - BACKGROUND: Randomised trials have evaluated various treatments for acute ischaemic stroke, but it is unclear how the results of these studies are used in everyday practice. AIMS: To obtain the opinions of physicians on the management of acute ischaemic stroke. METHODS: A questionnaire was sent to 368 New Zealand Fellows of the Royal Australasian College of Physicians. The survey included questions about the availability of hospital services for stroke patients, management of acute ischaemic stroke and opinion on the efficacy of treatments used in acute ischaemic stroke. RESULTS: Of the 293 physicians who responded to the questionnaire, 171 managed patients in the first week after stroke. Forty seven per cent of these physicians were general physicians. Ninety-five per cent usually managed these patients in a general medical ward. Only five physicians admitted patients to an acute stroke unit and only 57% considered acute stroke units were beneficial. Aspirin was usually or sometimes used for patients with acute ischaemic stroke by 92% of physicians, intravenous heparin by 43%, low-dose subcutaneous heparin by 41%, low-molecular-weight heparin by 25% and tissue plasminogen activator (t-PA) by 3%. Two thirds considered that aspirin was definitely beneficial, but most were uncertain about the efficacy of intravenous heparin, low-dose subcutaneous heparin, low-molecular-weight heparin and t-PA. Sixty-two per cent were prepared to begin aspirin and 21% subcutaneous heparin before computerised tomography (CT). Twenty-three per cent used anti-hypertensive treatment in the first few hours after an ischaemic stroke. CONCLUSIONS: Several common deficiencies in the management of acute ischaemic stroke were identified. The widespread lack of stroke units, use of aspirin and heparin before CT, and lowering of blood pressure after an acute ischaemic stroke differed from accepted guidelines. Many physicians used heparin despite lack of evidence from randomised trials that it is beneficial. The development of stroke units and the appointment of physicians with a special interest in the management of stroke may improve the management of patients with acute stroke. PMID- 10868497 TI - Growth hormone deficiency in adults: to treat or not to treat. PMID- 10868496 TI - Comparison of the long-term efficacy of implantable defibrillators and sotalol for documented spontaneous sustained ventricular tachyarrhythmias secondary to coronary artery disease. AB - BACKGROUND: The relative efficacy of antitachycardia pacing implantable cardioverter defibrillators (ATPICD) and sotalol in the treatment of ventricular tachyarrhythmias is controversial. AIM: To compare the mortality in patients treated with ATPICD and sotalol for documented spontaneous sustained ventricular tachyarrhythmias occurring late after previous myocardial infarction. METHODS: In this non-randomised retrospective study of 139 consecutive patients all patients had inducible ventricular tachycardia at baseline electrophysiological studies. Before the availability of ATPICD, 22 patients were treated with sotalol as part of a randomised study comparing the efficacy of sotalol to amiodarone. After ATPICD became available sotalol was used in 49 patients in whom intravenous testing predicted sotalol to be effective and ATPICD were implanted in 68 patients in whom sotalol was predicted to be ineffective at electrophysiological testing. Thus, 68 patients were treated with an ATPICD and 71 with sotalol. RESULTS: The two groups were well-matched for age, type of presenting arrhythmia, severity of coronary artery disease and ventricular function. At 36 months Kaplan Meier estimates of mortality from ventricular tachyarrhythmia were 0% with ATPICD and 15% with sotalol (p=0.03). Kaplan-Meier estimates of total mortality at 36 months were 12% with ATPICD and 25% with sotalol (p=0.09). Multivariate analysis showed hazard ratio of 7.9 (p=0.06) for death from ventricular tachyarrhythmia in patients treated with sotalol compared to ATPICD. CONCLUSIONS: While no difference in total mortality was demonstrated, treatment with ATPICD is probably superior to sotalol for preventing deaths due to ventricular tachyarrhythmia. PMID- 10868498 TI - Problem-based learning: process and practice. PMID- 10868499 TI - The insulin-like growth factor system: basic and clinical aspects. PMID- 10868500 TI - A review of isoniazid-related hepatotoxicity during chemoprophylaxis. PMID- 10868501 TI - Sudden cardiac death in familial hypertrophic cardiomyopathy: an Australian experience. PMID- 10868502 TI - Olanzapine: extrapyramidal side effects in the elderly. PMID- 10868503 TI - Cardiac toxicity associated with ciguatera poisoning. PMID- 10868504 TI - Vegebilia'--a late complication of endoscopic sphincterotomy. PMID- 10868505 TI - Focal necrotising vasculitis with secondary myositis following fluoxetine administration. PMID- 10868506 TI - Delayed diagnosis of recurrent visceral angio-oedema secondary to ACE inhibitor therapy. PMID- 10868507 TI - Hyperemesis gravidarum, transient hyperthyroxinaemia and primary hyperparathyroidism in a Chinese woman. PMID- 10868508 TI - Herpes simplex encephalitis in multiple myeloma. PMID- 10868509 TI - Haemolytic uraemic syndrome and prostatic cancer. PMID- 10868510 TI - Arsenic trioxide treatment of relapsed acute promyelocytic leukaemia: initial Australian experience. PMID- 10868511 TI - Large animal models of heart failure. AB - Congestive heart failure (HF) is a major focus of medical research. Its incidence has greatly increased in recent decades because of an aging population base and the increasingly successful treatment of other forms of chronic cardiac disease. Relevant large animal models of HF should reflect the complex interactions of cardiac dysfunction, neurohumoral dynamics and peripheral vascular abnormalities found in human HF. A number of large animal models have been developed, especially in dogs, sheep and swine, using naturally occurring HF, or single or combinations of interventions, as instruments to trigger the development of HF. Naturally occurring HF models are not commonly used because of ethical or perceived ethical grounds, however, King Charles Cavalier Spaniel and Yucatan Mini Pig models have been described. Tachycardia induced HF is the most commonly used HF model. Ventricular pacing at 220-240 bpm results in profound low output, biventricular, oedematous failure in two to three weeks. Lower pacing rates result in a more stable, sustainable, lesser degree of failure. Positive features of this model include 'acceptance', aetiological relevance to patient tachycardia induced HF, neurohumoral and functional profile similar to most human HF, relatively low cost simple preparation, ability to manipulate the degree of failure with pacing rate, reversibility, reliability and a large amount of published multi species data. Limitations to the use of the model are the rapid onset, the fact that reversibility is only relevant to the tachycardia induced patient HF, the absence of hypertrophy in failure, the diminished plasma atrial natriuretic peptide (ANP) levels, absence of ANP of ventricular origin, and the interference between rapid pacing and therapeutic interventions. Myocardial damage models of HF include those models induced by ischaemia, eg due to coronary occlusion (ligation or aneroid) or intracoronary microembolism, transmyocardial DC shock, toxic cardiomyopathy from adriamycin, doxorubicin or catecholamines. Overload models of HF may be induced by high pressure from aortic constriction, aortic regurgitation, renal artery constriction, pulmonary stenosis or aortocaval shunts, or by induction of mitral regurgitation from chordae or leaflet damage. No single, all-encompassing, large animal model of HF exists to date. Selection of the type of model to be used should be based primarily on the hypotheses to be tested and secondarily on the available resources and facilities. PMID- 10868512 TI - Animal models of heart failure. AB - Animal models of heart failure present homogenous groups of animals all with heart failure produced by a well defined lesion at a particular stage of evolution, in contrast to humans, who present with heart failure of uncertain duration from a wide variety of causes and with marked variation in age and pre morbid health and fitness. Animal models of heart failure provide diseased groups of animals in which experimental procedures, not possible in humans, can be evaluated and in which new treatments can be tested before their safety is established in humans. An ideal model should have a common human counterpart and should closely mimic heart failure in humans. Thus the haemodynamic changes should include increased cardiac filling pressures and low cardiac output. There should be evidence of activation of the sympathetic nervous system and increased secretion of hormones such as renin, angiotensin, aldosterone, vasopressin, atrial natriuretic factor and endothelin. The clinical features of the human syndrome such as cardiomegaly, lung and peripheral oedema and decreased exercise tolerance should be present. Lastly, the model should be inexpensive and technically simple to produce and study. This paper reviews some commonly used models of heart failure in relation to the criteria listed above. There is no perfect animal model of heart failure and in practice one should match the model to the purpose of the study. PMID- 10868513 TI - Mechanical circulatory support devices in the treatment of heart failure. AB - With an increasingly aging population, heart failure is a major health issue, affecting more than 10% of the population over 65 years of age, and costing hundreds of millions of dollars per year for ongoing care. Even with maximal medical therapy, annual mortality rates of in excess of 25% are commonly reported. Over the last three decades, various surgical approaches have been examined in the hope of improving the outcome of congestive cardiac failure. These procedures range from simple coronary revascularisation to left ventricular reduction surgery and cardiac transplantation. Although of value in selected situations, no surgical approach, beyond transplantation, has had significant impact on the outcome of heart failure. In the last decade, development in the area of mechanical support for the failing heart has continued to expand at a rapid rate. Strong evidence now exists to show that in many patients with advanced heart failure, prolonged mechanical support results in significant myocardial recovery. There are currently several mechanical support devices available for clinical use, although most are considered experimental in this country. These devices are expensive and are not without significant complications, but early results of their use as either a bridge to transplantation or as a stand alone treatment, have been very encouraging. Currently available mechanical assist devices are described, with discussion of indications for implantation, complications and results of their use. PMID- 10868514 TI - Sympathetic activation and the role of beta-blockers in chronic heart failure. AB - Chronic heart failure (CHF) is associated with activation of the sympathetic nervous system. This activation provides short term haemodynamic support to the failing myocardium, but may be deleterious over longer periods as chronic catecholamine excess appears to contribute to disease progression and increased mortality in this condition. Therefore, blockade of sympathetic activation represents a logical, if somewhat counter-intuitive, approach to the management of the patient with systolic CHF. Pharmacological approaches to blockade of this system include inhibition of central sympathetic outflow (using central sympatholytics, e.g. rilmenidine, moxonidine), blockade of the catecholamine biosynthetic pathway (dopamine beta hydroxylase antagonists) and blockade of the cardiac effects of sympathetic activation (beta-adrenoceptor blocking agents). Beta-blockers have now been extensively studied in patients with symptomatic CHF of the New York Heart Association (NYHA) Class II and III severity. Provided beta blocker therapy is carefully up-titrated from sub-therapeutic doses and given for at least three to four months, these agents have been associated with favourable long term haemodynamic, functional and mortality outcomes. Furthermore, beta blockers delay disease progression in CHF by reversing the pathological myocardial remodelling process that accompanies the disease. Non-selective beta adrenoceptor blocking drugs appear to be of particular benefit in CHF therapy. Myocardial beta2 receptors are down-regulated to a lesser extent than beta1 receptors in CHF, therefore blockade of the beta2 receptor sub-type may assume greater importance in inhibiting the deleterious effects of sympathetic activation on the myocardium in this disease. Newer agents that possess additional vasodilator properties (carvedilol, bucindolol, nebivolol) may be useful in overcoming the initial negative inotropy of the beta-blocking component of the drug, however, it is unlikely that vasodilation contributes greatly to long term clinical benefits. Drugs such as carvedilol are also anti-oxidant, anti proliferative and have anti-endothelin actions; the clinical significance of these properties is yet to be determined. Unanswered questions remain regarding the use of beta-blockers in heart failure. Ongoing studies are further examining mechanisms underlying the clinical benefits of these agents as well as their therapeutic potential in NYHA Class IV patients, heart failure post-myocardial infarction and patients with asymptomatic left ventricular dysfunction. PMID- 10868515 TI - The role of diastolic ventricular interaction in abnormal cardiac baroreflex function in chronic heart failure. AB - Baroreflex abnormalities have been well documented in both patients with chronic heart failure and experimental animal models of heart failure. These abnormalities are associated with increased mortality and probably contribute to neurohumoral activation. While it is likely that several mechanisms contribute to reduced baroreflex sensitivity, it has been difficult to explain why baroreflex control mechanisms during acute volume unloading in patients with severe chronic heart failure should be directionally opposite to those in normal subjects. Volume unloading normally causes a reduction in baroreceptor activity, and hence an increase in sympathetic outflow; however, patients with chronic heart failure develop attenuated increases or paradoxical reductions in forearm vascular resistance, muscle sympathetic nerve activity, and noradrenaline spillover. It has been suggested that this probably represents paradoxical activation of left ventricular (LV) mechanoreceptors, but why LV receptors should behave in such a fashion has not been determined. In the setting of diastolic ventricular interaction, the filling of the left ventricle is constrained by the surrounding pericardium and right ventricle. In these patients, the reduction in right ventricular (RV) volume that normally occurs during acute volume unloading allows for an increase in LV end-diastolic volume (as opposed to the reduction in LV volume that normally occurs). We have demonstrated this to be important in some patients with chronic heart failure, and observed that baroreflex control of forearm vascular resistance was markedly impaired in these patients. We propose that the increase in LV volume that occurred during volume unloading would increase LV mechanoreceptor activity, and could therefore explain the paradoxical reductions in sympathetic outflow. As discussed, this has important therapeutic implications. PMID- 10868516 TI - New thrombolytic drugs. AB - Thrombolytic drugs reduce mortality from myocardial infarction and research is focused on newer drugs with improved clinical efficacy. The ideal thrombolytic drug should be effective in dissolving fresh and older thrombi, be rapid in its action with complete dissolution of the thrombus, be able to be given as a bolus and be safe without hypotensive or allergic or immunogenic reactions. The types of agents being developed fall into five broad categories: * mutants or variants of single chain urokinase type plasminogen activator; * mutants or variants of tissue type plasminogen activator; * recombinant chimaeric plasminogen activators; * conjugates of plasminogen activators and anti-fibrin monoclonal antibodies; * compounds derived from haemophagous animals. Within the mutant and variant groups there may be single point mutations which increase the half life or deletion of various amino acid sequences to increase resistance to plasma protease inhibitors or cause more selective binding to fibrin. The recombinant chimaeric plasminogen activators are designer drugs where the kringle regions, the growth hormone domain region, the finger domain region and the serine protease part of the molecules are all cleaved and recombined in various ways to improve potency. The conjugates of plasminogen activators and anti-fibrin and monoclonal antibodies improve the targeting of the agent to fibrin clot. Monoclonal antibodies are commonly used against the seven aminoterminal residues of the beta chain of fibrin. These are cross linked and bound to plasminogen activators. A variety of substances from haemophageous animals are currently being studied including salivary plasminogen activator from vampire bats, venom from southern copperhead snakes and staphylokinase from bacteria. Currently available thrombolytic agents have many limitations, but the novel thrombolytic agents have yet to be tested in clinical trials. With few exceptions, they all act via the plasminogen system and it may be in the future that combinations of anti-platelet and thrombolytic agents may prove to be more efficacious than thrombolytics and aspirin alone. PMID- 10868517 TI - Troponin and other cardiac markers: role in management of acute coronary syndromes. AB - Many adjuncts to the traditional history, physical examination, and baseline ECG have been developed to improve the differential diagnosis of acute coronary syndromes in the Emergency Department (ED). These include serial ECGs, continuous ST-segment monitoring, and serum markers of myocardial necrosis. In general, the serum markers of coronary ischaemia, particularly the troponins, provide the most information toward initial and later risk stratification. Serial marker measures not only increase the sensitivity and negative predictive value for acute infarction (thus reducing hospital costs) but also can add to prognostic ability. More importantly, a positive marker result indicates an increased risk of an early event, which could allow earlier intervention and thus a better outcome. PMID- 10868518 TI - Antithrombins and the importance of good control. AB - The generation of thrombin and the formation of platelet rich intra-coronary thrombus in response to atherosclerotic plaque rupture is pathognomonic of acute coronary syndromes. An understanding of the process of thrombin generation and the unique relationship between the structure and function of thrombin is essential to developing more effective anti-thrombotic strategies than the use of standard unfractionated heparin in the acute coronary syndromes. The mechanisms of action of heparin, low molecular weight heparins (LMWHs) and the newer direct anti-thrombins, recombinant hirudin and Hirulog, are reviewed. Evidence from the currently available phase 2 and 3 trials of these drugs regarding their efficacy in the acute coronary syndromes is also reviewed. PMID- 10868519 TI - Platelet adhesion receptors: novel targets for anti-thrombotic therapy. AB - The critical role of platelets in the development of the acute coronary syndromes is now well recognised, and a great deal of effort has therefore focused on elucidating the key adhesion receptors mediating platelet-vessel wall and platelet-platelet interactions. The vascular adhesion protein von Willebrand factor (vWf) plays a key role in supporting platelet adhesion to the damaged vessel wall and binds to two adhesion receptors on the platelet surface, the glycoprotein (GP) Ib-V-IX complex and glycoprotein IIb-IIIa. The GP Ib-V-IX complex is a unique adhesion receptor which enables platelets to roll on a vWf matrix under conditions of rapid blood flow as well as transducing signals leading to the activation of GP IIb-IIIa. This latter receptor binds to a distinct site on vWf and is essential for stabilising platelet adhesion to the site of vessel wall injury. In addition to supporting platelet adhesion, GP IIb IIIa plays a key role in a number of other platelet responses including platelet spreading, aggregation, the release of procoagulant-rich microvesicles, and clot retraction. Given its central role in platelet function GP IIb-IIIa has become an attractive target for the development of novel anti-thrombotic agents. In this paper, we consider the advantages of inhibitors of GP IIb-IIIa compared with other established anti-platelet drugs including aspirin and ticlopidine, and also discuss some potential problems associated with the inhibition of GP IIb/IIIa and other platelet adhesion receptors. PMID- 10868520 TI - Reperfusion therapy for stroke. AB - Stroke is a heterogeneous disease, but about 85% of strokes are as a result of cerebral ischaemia due to arterial occlusion. It seems logical to assume that, as in myocardial infarction, treatment designed to dissolve clots should be helpful. We now have a substantial amount of data on the use of aspirin, heparin and thrombolytic drugs in the treatment of acute ischaemic stroke. Aspirin 300 mg daily has a modest effect in reducing mortality and handicap when used within 48 hours of stroke onset. The beneficial effects of low dose, medium dose subcutaneous unfractionated heparin, and various low molecular weight heparins in reducing early recurrent ischaemic stroke seem to be outweighed by haemorrhagic side effects. Streptokinase used within six hours of stroke onset results in excess mortality with some reduction in handicap in survivors, while in carefully selected patients recombinant tissue plasminogen activator (r-TPA) may be less hazardous. At the moment it is unclear which stroke patients will benefit from the use of r-TPA, and the use of criteria, as outlined by the NINDS group, means that only a very small proportion of stroke victims are suitable for thrombolytic therapy. Further research is necessary, while the concept of a 'Brain Attack' with appropriate urgency being used in the assessment of possible stroke needs development. PMID- 10868521 TI - Non-rheumatic atrial fibrillation and stroke. AB - The risk of systemic embolism and stroke in patients with non-rheumatic atrial fibrillation (NRAF) should not be underestimated. The annual embolic rate is approximately 5% and in those with left atrial enlargement and/or left ventricular (LV) dysfunction, or who have already had systemic embolism, this rate may be as high as 20%. Decisions on patient management and the prophylaxis of stroke must always be individualised. The risk of bleeding related to warfarin is almost certainly greater than that encountered in the previous randomised trials. Also, clinical and echocardiographic features can further define absolute risk in an individual patient with NRAF. Clinical markers of increased risk of embolism in patients with NRAF include older age, previous cerebral embolism, recent congestive heart failure, hypertension and diabetes mellitus. Transthoracic echocardiography improves risk stratification and should be performed in the vast majority of patients. Embolic risk is greatest in those with increasing left atrial dilation, atrial dysfunction and LV dysfunction. Transoesophageal echocardiography sharpens the risk profile in selected patients. Overall randomised trials show greater benefit with warfarin than aspirin. In general, increasing age is associated with a greater incidence of structural heart disease and probably implies greater potential benefit with warfarin. Increasing age per se may not increase the risk of warfarin-related bleeding. When the decision is made to warfarinise patients, at the present time data suggest that the target INR should be in the range of 2.0-3.0. PMID- 10868522 TI - Oestrogen and vascular disease. AB - Cardiovascular disease (CVD) is the leading cause of morbidity and mortality in postmenopausal women. Epidemiological studies consistently suggest that oestrogen administered to postmenopausal women confers an estimated 30-50% reduction in risk of development and progression of CVD. The long term effect on the cardiovascular system of the addition of a progestogen to the replacement regimen is currently unknown. In addition, it may be argued that it remains to be proven whether the magnitude of the oestrogen-induced cardioprotective effect demonstrated in these observational studies is a real biological phenomenon. No prospective, randomised, controlled studies examining the effect of oestrogen on primary and secondary prevention of CVD have been completed. However, a large number of biologically plausible mechanisms have been identified which provide evidence to support the proposed oestrogen-induced cardioprotection. These include oestrogen mediated favourable changes in metabolic profile, in particular changes in lipid metabolism, insulin resistance and the fibrinolytic system. In addition, recent data have shown that oestrogen may affect vascular structure and function by a variety of mechanisms. It has been shown that oestrogen may induce acute and chronic coronary and cerebral vasodilation through both direct (vascular smooth muscle) and indirect (endothelium dependent) mechanisms. Oestrogen also has recently been shown to have complex anti-atherogenic and antioxidant properties. Much less is known of the vascular effects of progestogens. Progestogens currently in clinical use have androgenic properties and may attenuate the beneficial effects of oestrogen by neutralising or opposing the lipid lowering, vasodilatory and anti-atherogenic actions of oestrogen. Thus further studies are required to elucidate the effects on arterial physiology and CVD outcome of the oestrogens and progestogens of different types, doses and routes of administration which are collectively referred to as postmenopausal 'hormone replacement therapy'. PMID- 10868523 TI - Stemming the tide: reducing cardiovascular disease and renal failure in Australian Aborigines. AB - An epidemic of cardiovascular disease (CVD) and end stage renal disease (ESRD) has developed among Aborigines in the Northern Territory; CVD deaths increased over the 1980s (tripling among women!), and are now more than five times those of non-Aboriginal people, while ESRD rates are increasing more than 20-fold and are doubling every three to four years. Dialysis costs (>$75,000 per person/year) pose a crisis for health care budgets, but premature mortality is the greater human catastrophe. Health services are not meeting the challenge of timely diagnosis, prevention and containment. We screened 90% of adults (20+ years) in one community, with CVD mortality among the highest in Australia, and ESRD rates increased 60-fold. Seventy-five per cent of persons were smokers. Central obesity was common, but BMIs only modestly increased by Caucasian standards, 23% had hypertension (>140/90), 29% had diabetes or impaired glucose tolerance (IGT) (peaking at 65% of persons aged 40-49 years), high triglyceride and insulin levels were common, and 55% had albuminuria (albumin/creatinine ratio (ACR), >3.4 gm/moL). Progressive albuminuria predicted renal failure. ACR was correlated with age, BMI, blood pressure, lipid, glucose and insulin levels, heavy drinking and past and current skin infections, and, inversely with birth weight. ACR correlated strongly with a composite CV risk score, and in a two to five year follow-up, microalbuminuria (ACR 3.4-33) and overt albuminuria (ACR 34+) have both predicted increased rate of premature death from natural causes of lower ACRs. Thus albuminuria marks CV risk/disease. This implies that renal and CV disease share common risk factors, and should respond to the same interventions, and that this response might be monitored through ACR levels. Robust public health programmes could reduce all these reversible risk factors, lowering disease rates over the intermediate term, however, few such programmes are in place. Modification of disease in persons already afflicted is a parallel responsibility. To this end, in November 1995, we introduced a treatment programme with Coversyl (perindopril, Servier) for all persons in the study community with hypertension (>140/90), for all diabetics with ACR 3.4+ and for all nondiabetic, non-hypertensive persons with progressive overt albuminuria (ACR 34+). One-quarter of all adults, or 224 persons have enrolled; 162 have reached one year of treatment and 100 have passed two years. Compliance is reasonable and enthusiasm high. Average SBP has fallen 12 mmHg (24 mmHg in hypertensive persons), while average ACR and estimated glomerular filtration rate (GFR) have stabilised. This contrasts favourably with the pretreatment course (average 2.7 years) in the same persons, when SBP had increased by 3 mmHg, ACR had increased by 15% and GFR had decreased by 3.5 mL/min each year. Cautious estimates suggest a >50% fall in ESRD, and a reduction in all-cause and CV deaths, even at this early stage, although more extended observation is needed. These data predict a dramatic and rapid fall in morbidity, premature deaths and health care costs if these basic principles of medical care are extended to all Aboriginal people. A national, concerted, multi-disciplinary effort to implement a coherent, effective strategy to this end is of great urgency. PMID- 10868524 TI - Overprescribing: have we made any progress? PMID- 10868525 TI - The addition of allogeneic peripheral blood-derived progenitor cells to bone marrow for transplantation: results of a randomised clinical trial. AB - BACKGROUND: The use of cytokine-mobilised peripheral-blood-derived progenitor cells (PBPC) has resulted in a significant improvement in the safety of autologous transplantation. Collections of PBPC contain large numbers of haemopoietic progenitors and T-lymphocytes when compared with bone marrow (BM). AIMS: To assess the clinical effects and safety of filgrastim-mobilised allogeneic PBPC, in particular whether PBPC would alter the kinetics of engraftment or the incidence and severity of graft-versus-host disease (GVHD). METHODS: Twenty-seven patients undergoing allogeneic transplantation were randomised to receive BM or BM supplemented by PBPC. Patients were conditioned with busulphan 16 mg/kg and cyclophosphamide 120 mg/kg. GVHD prophylaxis was with methotrexate and cyclosporin. Ganciclovir prophylaxis was administered to all cytomegalovirus seropositive patients. No haemopoietic growth factor was used after BMT. Donors of PBPC underwent a single, seven litre apheresis after four days of filgrastim, 10 microg/kg/day by subcutaneous injection. RESULTS: Donor toxicity was mild, with the most frequently reported being bone pain in the cytokine-treated donors. The patients transplanted with BM+PBPC received an eight fold increase in CD3+ T-lymphocytes (1.65X 10(8)kg vs 0.22 x 10(8)/kg, p=0.04) and the cytokine product contained a median of 5 x 10(6) CD34+ cells/kg. BM+PBPC patients had more rapid engraftment of neutrophils to 0.5x 10(9)/L (17 days vs 19 days, p=0.02) and platelets to 50X 10(9)/L (22 days vs 35 days, p=0.04). BM megakaryocyte numbers were significantly enhanced at days 14 and 28 after BMT (both p=0.04), however platelet transfusions were not significantly different. The incidence, organ distribution, severity and time to onset of acute and chronic GVHD were not different between the treatment groups and there was no difference in overall survival or event-free survival. CONCLUSIONS: Allogeneic PBPC from HLA-identical siblings may speed engraftment of neutrophils and platelets without detrimental effects on GVHD or survival. PMID- 10868526 TI - Profile of people referred to an emergency department from residential care. AB - BACKGROUND: Elderly people in residential care are among the most infirm in society and are at high risk of developing acute medical problems. There are no Australian data on the use of acute hospital emergency services by this group. AIM: To determine patterns of use of a major public hospital's Emergency Department (ED) by elderly people living in residential care, their presenting problems and the outcome of attendance. METHODS: Prospective study of 300 consecutive referrals to a teaching hospital's ED involving people aged over 65 years and living in residential care in southern Adelaide, South Australia. Case records were examined and residential care staff were interviewed by telephone when information required clarification. This occurred in 25% of referrals. RESULTS: The 300 referrals were seen over a three month period and accounted for 2.43% of the 12,371 ED attendances during this period. During this time, at least 4.9% of people in residential care in the region were referred to the ED. The referrals involved 239 residents, 196 (82%) who were referred once only, 32 (13%) twice and 11 (5%) three or more times. Residents had a mean age of 84 years and 70% were female. A broad range of acute medical problems precipitated referral and 61% of people referred were immediately hospitalised. There was no general practitioner (GP) involvement in the management of the presenting illness in 58% of all referrals and in 45% of those where symptoms had been present for over three days. CONCLUSIONS: People living in residential care are frequently referred to an ED service, often bypassing their GP in the process. They present with a wide range of acute medical problems for which most are hospitalised. Strategies that anticipate, prevent and manage health breakdown in residential care and so minimise the need for ED referral should be trialed. PMID- 10868527 TI - Monoclonal gammopathy of undetermined significance (MGUS)-31 year follow up of a community study. AB - BACKGROUND: When unselected healthy adults in the community have their serum screened by cellulose acetate or paper electrophoresis, monoclonal gammopathy of undetermined significance (MGUS) may be found in 0.5-1%. AIM: To report upon a 31 year follow up of MGUS in a New Zealand community. METHODS: Serum from 2,192 subjects (82% of the adult population) of a New Zealand town was collected in 1967 and subsequently screened by cellulose acetate electrophoresis. Eleven of the 2,192 (0.5%) were found to have MGUS. Clinical correlation was sought in 1970 and subsequently to elucidate the underlying cause. RESULTS: Seven of the 11 patients have developed a haematological malignancy. Two have been diagnosed as having Waldenstrom's macroglobulinaemia, one malignant lymphoma and acute myelomonocytic leukaemia and four developed myeloma. Myeloma developed at one, nine, 23 and 25 years after the original screening. One myeloma patient and one patient with MGUS are currently alive and well, 31 years after discovery of their gammopathy. CONCLUSIONS: The incidence of MGUS in this community is only half that detected in a comparable study. The association with haematological malignancy in this study (64%) is considerably higher than that found in the Swedish study (6%), possibly because of the longer follow up in New Zealand. MGUS should not only be studied in depth at the time of its discovery, but needs very long term follow up as the underlying disease may not surface until the third decade. PMID- 10868528 TI - Review of Emergency Department thrombolytic therapy and changes in inpatient mortality of acute myocardial infarction on the NSW Central Coast 1986 to 1994 96. AB - AIMS: To examine changes in inpatient mortality of acute myocardial infarction (AMI) from 1986 to 1994-96 and to review the Emergency Department (ED) use of thrombolytic therapy (TT) for AMI on the NSW Central Coast. METHOD: A retrospective review of medical records of patients presenting to the EDs of Gosford and Wyong Hospitals with a discharge diagnosis of AMI (ICD9 code 410.x) from 1 January 1986 to 31 December 1986 and 1 January 1994 to 31 December 1996. Data were collected on patients' age, sex, duration of symptoms on arrival at the ED, ECG changes and presence of positive ECG criteria for thrombolysis, agent used, contraindications to TT, and inpatient mortality. The main measure of outcome was inpatient mortality. RESULTS: There were 423 admissions for AMI in 1986 and 1,220 admissions in 1994-96. The overall inpatient mortality has declined from 18.9% in 1986 to 9% in 1994-96 (p<0.0001). The mean age of patients has increased from 67.5 years to 68.1 years (p=0.35). The proportion of patients over age 75 years has increased significantly from 24.6% to 30.3% (p<0.0001). Presentation times from onset of symptoms have not changed significantly from a median time of two hours in 1986 to 2.5 hours in 1994 to 1996 (p=0.52). The overall proportion of patients with ECG criteria for TT was 53.2% in 1994-96. TT was administered to 42.9% of patients with a mean door to needle time of 67 minutes (median 45 minutes). The Australasian College for Emergency Medicine benchmark door to needle time of 60 minutes was achieved in 71.3% of patients. Streptokinase was the predominant agent given in 78%, while recombinant tissue plasminogen activator accounted for 15.7% of patients. Patients not receiving TT due to negative ECG criteria showed a decline in mortality from 18.6% to 6.7% (p<0.0001). Patients who underwent mechanical revascularisation (by bypass graft or angioplasty) increased from 8.7% to 17.4% (p<0.0001). Inpatient mortality has declined for all age groups, for both sexes, and for all sites of AMI. CONCLUSION: There have been significant declines in inpatient mortality of patients with AMI on the Central Coast. TT has had a significant impact on this decline but has an eligibility rate of less than half. Significant declines in mortality have also been seen in patients ineligible for thrombolysis. These patients have benefited from other therapies introduced or more widely used in the last decade. The results achieved on the Central Coast compare favourably with published reviews in Australia and overseas despite the lack of facilities for coronary angiography, coronary angioplasty and cardiothoracic surgery. PMID- 10868529 TI - Helicobacter pylori treatment and antibiotic susceptibility: results of a five year audit. AB - BACKGROUND: Helicobacter pylori eradication treatment has been a rapidly evolving field. Audit of treatment results provides reassurance that trial data can be translated into routine clinical practice. METHODS: Data were collected prospectively over five years. Patients were given four different treatment regimens over the audit period 'standard' triple therapy, two types of clarithromycin-based treatment or ranitidine, amoxycillin and metronidazole. Eradication was proven by a urea breath test at least four weeks after completing treatment. RESULTS: Eradication treatment for H. pylori was given to 665 patients; 89% had follow-up data. H. pylori eradication was significantly associated with treatment type (p<0.0001) and smoking (p=0.04) by univariate analysis, but was not associated with sex, age, alcohol consumption, endoscopic diagnosis, recent treatment with anti-secretory drugs or NSAIDs. By logistic regression analysis, only treatment type was significant (p=0.0001). H. pylori culture and sensitivities were available for 255 patients. Metronidazole resistance was shown for 84 isolates (32%) and clarithromycin resistance for 18 isolates (6.8%). Metronidazole resistance was significantly associated with younger age (p=0.02), ethnicity (p=0.02), female sex (p=0.02), and year of endoscopy (p=0.04), but was not associated with clarithromycin resistance. Clarithromycin resistance was not associated with age, sex, or ethnicity. Metronidazole resistance significantly affected H. pylori eradication for regimens containing metronidazole without clarithromycin. Eradication with metronidazole without clarithromycin was achieved in 90% of sensitive strains but only 55% of resistant strains (p<0.001). Metronidazole resistance was not significantly associated with treatment failure when metronidazole was combined with clarithromycin. Eradication with metronidazole and clarithromycin was achieved in 86% of sensitive strains and 78% of resistant strains (p=0.42). CONCLUSION: Treatment type and antibiotic susceptibility are the most important determinants of treatment success. PMID- 10868530 TI - Treatment of metastatic breast cancer with continuous infusional 5 fluorouracil. AB - BACKGROUND: Single agent continuous infusional 5 fluorouracil (CI-5FU) via a central venous catheter (CVC) is usually reserved for breast cancer patients who have previously failed one or more chemotherapy regimens. The patients are usually heavily pre-treated with later stage disease. Previously published studies of CI-5FU have reported response rates as high as 54%. It is considered an approach with an acceptable side effect profile in such patients. AIMS: To evaluate the efficacy and toxicity of CI-5FU in previously treated metastatic breast cancer. METHODS: A retrospective review of advanced breast cancer patients treated with CI-5FU between October 1992 and October 1996 was performed. Response to treatment, toxicity, CVC complications and patient survival were analysed. RESULTS: Twenty-four patients with metastatic breast cancer were treated with CI 5FU. All had received previous chemotherapy, including 19 patients (79%) with prior 5FU exposure and eight patients (33%) who had previous high dose chemotherapy with autologous stem cell transplantation. The median duration of CI 5FU treatment was 3.1 months. Nineteen patients had evaluable disease, three (16%) of whom demonstrated a partial response and four patients had stable disease. There were no complete responses. All responses occurred in soft tissue sites with no objective evidence of response in liver or bone metastases. The survival rate at one year was 21% (five of 24) and the median survival of all patients was 6.1 months. Five patients (21%) stopped treatment due to treatment related morbidity (two CVC complications and three CI-5FU side effects). Diarrhoea, nausea, and palmar-plantar erythrodysaesthesia were the major side effects of chemotherapy. CVC complications requiring intervention, the most notable of which were infection and thrombosis, occurred in 11 patients (46%). There were no treatment related deaths. CONCLUSIONS: Single agent CI-5FU has modest activity in women with previously treated advanced breast cancer. The efficacy is lower than in previously published series. This may reflect patient selection factors. The toxicity was mainly related to CVC complications. Important issues relating to quality of life need to be objectively measured in future studies of CI-5FU. PMID- 10868531 TI - Endothelial dysfunction, insulin resistance and diabetes: exploring the web of causality. PMID- 10868532 TI - Current trends in the management of early stage Hodgkin's disease. PMID- 10868533 TI - Faecal occult blood test screening for colorectal cancer--what are we waiting for? PMID- 10868534 TI - Treatment of peritoneal dialysis related peritonitis--an Australian and New Zealand perspective. PMID- 10868535 TI - Fludarabine and high dose cytarabine (FLA): a well tolerated salvage regimen in acute myeloid leukaemia. PMID- 10868536 TI - Suspected Ross River virus encephalitis in Papua New Guinea. PMID- 10868537 TI - Clozapine-induced intestinal obstruction. PMID- 10868538 TI - Coexistent cryptococcosis and carcinoma within a solitary pulmonary nodule. PMID- 10868539 TI - Lymphogranuloma venereum presenting with a psoas abscess. PMID- 10868540 TI - Successful management of pain syndrome due to Angiostrongylus cantonensis by implantable spinal cord stimulator. PMID- 10868541 TI - Deep venous thrombosis as the presenting feature in a patient with coeliac disease and homocysteinaemia. PMID- 10868542 TI - Atrial fibrillation--prevalence and management. PMID- 10868543 TI - Atrial fibrillation--prevalence and management. PMID- 10868544 TI - Septic portal vein thrombosis due to Fusobacterium necrophorum. PMID- 10868545 TI - Presentation of Candida glabrata spinal osteomyelitis 25 months after documented candidaemia. PMID- 10868546 TI - Chylous ascites caused by an occult pancreatic malignancy. PMID- 10868547 TI - Camel kidney ferritin: isolation and partial characterization. AB - Camel kidney ferritin was isolated from a tissue homogenate by thermal denaturation, ammonium sulphate fractionation, Sephacryl S-300 gel filtration and DEAE-blue gel affinity chromatography. The yield and the iron and neutral carbohydrate contents were 0.012 mg/g wet tissue, 4.0% and 2.7%, respectively. The phosphate:iron ratio was 0.13, twofold lower than that reported for camel liver ferritin. Native gel electrophoresis revealed the presence of a monomeric ferritin. SDS gel electrophoresis and immunoblotting showed two types of subunits, heavy and light, contrary to the extensive heterogeneity observed in camel liver ferritin. In general, the tissue ferritins shared a similar amino acid composition. However, a twofold lower glycine and an eightfold higher arginine content were recorded for camel kidney ferritin. In addition, kidney ferritin had a relatively high content of glutamic acid. Cross-reactivity studies by Ouchterlony double diffusion and noncompetitive indirect ELISA revealed a distinct cross-reactivity between buffalo ferritin antiserum and camel liver ferritin, but camel liver ferritin showed only weak cross-reactivity. PMID- 10868548 TI - Tissue culture of the enteric nervous system from equine ileum. AB - Ileal samples were harvested fresh from euthanized adult horses. The tissues were microdissected to prepare wholemount preparations for immunohistochemistry and for either explant or dissociated culture systems of the enteric nervous system. Explant culture systems were established using whole-mounts of either the submucous plexus or the muscularis externa (including the myenteric plexus). Dissociated cell cultures could only be obtained from the submucous plexus. Culture systems were maintained for up to 5 days. Immunoreactivity for a neuronal marker (Pan-N) and for glial cell markers (GFAP and S100) indicated the presence of both neurons and enteric glia in the tissue culture preparations. This is the first report of equine enteric neurons being grown in tissue culture Further refinements to the techniques will be required before this in vitro model can be used for quantitative analysis. PMID- 10868549 TI - Anticoccidial vaccination of broiler chickens in various management programmes: relationship between oocyst accumulation in litter and the development of protective immunity. AB - Paracox anticoccidial vaccine was administered to a 7-day-old flock of commercial broiler breeder stock subsequently reared to point-of-lay in the same house. For comparison, three subgroups of another flock of broiler breeders were also vaccinated with Paracox at 7 days of age, reared to 42 days and then transferred to new litter on another farm until point-of-lay. The first subgroup received no further treatment, but the second and third each received a second vaccination with Paracox, either immediately after transfer to the new litter or 42 days after transfer. Using an Eiomeria necatrix model, protective immunity was demonstrated by virulent challenge of samples of birds from all groups by the age of 37-40 days (30-33 days after the first vaccination), and was maintained to at least 122-125 days of age, whether the birds remained on the same litter or were transferred to another farm, and whether they received one or two anticoccidial vaccinations. Therefore, there is no disadvantage in transferring birds onto new litter 35 days after a single Paracox vaccination, nor is there any advantage in giving a second vaccination after such a transfer. Vaccinated birds seeded the new litter with oocysts, despite being clinically immune to coccidiosis. A supplementary laboratory experiment showed that birds vaccinated at 8 days of age passed almost no oocysts after a second vaccination at 43 days of age. This indicated that they were not only protected against clinical coccidiosis, but were almost solidly immune to a homologous infection 5 weeks after a single vaccination. Nevertheless, oocysts appeared in the litter of all four groups of commercial breeders throughout the trial, showing that wild-type heterologous infections occurred whether the birds were transferred to new litter or not, but these did not overwhelm the acquired protective immunity and cause clinical coccidiosis. PMID- 10868550 TI - Studies on the mechanism of diarrhoea induced by Escherichia coli heat-stable enterotoxin (STa) in newborn calves. AB - Enterotoxigenic Escherichia coli (ETEC) produces a heat-stable enterotoxin (STa) that binds to and activates a putative intestinal receptor, guanylate cyclase, causing an increase in the intracellular levels of cyclic guanosine monophosphate (cGMP). Using flow cytometry and 125I-STa binding assays, we studied the distribution of STa-receptors on enterocytes isolated from different segments of the newborn calf's intestinal tract. We also investigated the effect of STa on the intracellular levels of cGMP and ion transport to the intestinal lumen. More STa-receptors were found on enterocytes prepared from the ileum than on enterocytes obtained from the other segments of the intestinal tract. Guanylate cyclase activity was higher in the ileum of STa-challenged calves than in the ileum of control calves. No changes were observed in the guanylate cyclase activity of the other intestinal segments of the STa-challenged and control calves. Na+ levels, as measured by atomic absorption spectroscopy, were significantly increased in the luminal contents of the ileum of STa-challenged calves, whereas serum Cl- levels were significantly lower in the STa-challenged calves than in control calves. This study supports previous observations on the role of guanylate cyclase in the initiation of STa-induced secretory diarrhoea and suggests that Na+/Cl- coupling may be the major mechanism for the loss of ions in the diarrhoeal response that is mostly induced in the ileum of newborn calves. PMID- 10868551 TI - Pharmacokinetic behaviour of albendazole sulphoxide enantiomers in male and female sheep. AB - Benzimidazole anthelmintic drugs are widely used in veterinary practice. Albendazole sulphoxide (ABZSO) is a benzimidazole drug with two enantiomers, as a consequence of a chiral centre in the sulphoxide group. The kinetics of these enantiomers were studied in male and female sheep. Plasma samples were obtained from the animals between 0.5 and 72 h after oral administration of 7.5 mg/kg of a racemic formulation of ABZSO (total-ABZSO). After a liquid-liquid extraction, the samples were analysed by HPLC to determine the concentrations of total-ABZSO and of the sulphone metabolite (ABZSO2). During the chromatographic analysis, the ABZSO peak was collected and reanalysed by an HPLC technique using a Chiral AGP column to quantify the enantiomeric proportion therein. After kinetic analysis, the AUCs obtained for the (+)-ABZSO were 5.8 and 4.0 times higher than those for the (-)-ABZSO in male and female animals, respectively. The mean residence times were 23.4 and 16.1 h for (+)-ABZSO and 22.2 and 17.4 h for (-)-ABZSO for male and female animals, respectively. The only significant difference between the sexes (p < 0.05) was in the Tmax of the (-)-ABZSO. Comparing both enantiomers within each sex, significant differences were found in all the kinetic parameters. Finally, no kinetic differences were found between sex for total-ABZSO or ABZSO2. PMID- 10868552 TI - Pathological lesions in lambs fed raw or processed cottonseed meal. AB - Thirty male lambs of 3-4 months of age, were assigned equally to five dietary treatments in a completely randomized design and fed isonitrogenous and isocaloric concentrate mixtures containing 30% de-oiled peanut meal (DPNM) or 40%, cottonseed meal, which was raw, cooked for 45 min or treated with either 1%, calcium hydroxide or iron (1:3, free gossypol: Fe). The mixtures containing raw or variously processed CSM replaced about 50% of the nitrogen of the reference concentrate mixture. These concentrate mixtures were fed to meet 80% of the animals' crude protein requirements along with ad libitum feeding of maize (Zea mays) hay for 180 days. The free gossypol content of the raw cottonseed meal (0.27%) was reduced to 0.16% 0.20% and 0.21% by the cooking, Ca(OH)2 and iron treatments, respectively. At the end of the experiment, the tissues of various organs were fixed in 10% formol saline. embedded in paraffin and sectioned at 4-5 microm thickness, and duplicate sections were stained with either haematoxylin and eosin or Perl's Prussian blue. The lambs fed diets incorporating raw, cooked, Ca(OH)2- or iron-treated cottonseed meal consumed respectively 302, 215, 250 and 222 mg free gossypol/day. No morbidity. mortality or gross lesions were observed in any organs and the histopathological lesions due to cottonseed meal were limited to the testes and epididymis. Spermatogonial cells were absent in the majority of the seminiferous tubules of testes from lambs fed raw cottonseed meal. Most seminiferous tubules were collapsed, with a reduced wall thickness, owing to there being fewer germ cell layers and vacuolation of the basal cells. The epithelium of the epididymal ductules was degenerated, desquamated to a variable degree with hyperplastic changes, and they were devoid of spermatozoa. Most lambs fed any of the processed cottonseed meals did not show any of these lesions, and such lesions as occurred in affected lambs in these groups were relatively mild. Iron pigments were deposited around the portal areas of the liver, the tip of intestinal villi and the spleen of lambs fed the iron-treated cottonseed meal diet. Cooking or treatment with 1%, Ca(OH)2 effectively minimized the toxic effects of free gossypol. PMID- 10868553 TI - Over-the-counter analgesics and antipyretics: a critical assessment. AB - BACKGROUND: It is just 100 years since the introduction of aspirin to medicine. Since then, aspirin and its derivatives have been joined by acetaminophen, and the nonsteroidal anti-inflammatory drugs--ibuprofen, naproxen sodium, and ketoprofen--as the only over-the-counter (OTC) agents approved by the US Food and Drug Administration for the short-term treatment of pain, headache, dysmenorrhea, and fever. Recently the prescription use of aspirin has expanded to include a number of antiplatelet indications. OBJECTIVE: The purpose of this paper is to review critically the history, mechanisms of action, efficacy, and tolerability of OTC analgesic and antipyretic products. Relatively new and potential future indications for these drugs are also discussed. CONCLUSION: Although all of the OTC analgesic/antipyretic agents seem to share a common mechanism of prostaglandin inhibition, there are important differences in their pharmacology, efficacy, and side-effect profiles. Considering their often-unsupervised use, the risk-benefit ratio of this class of drugs has been extremely favorable. However, when used inappropriately, even these drugs pose significant risks to certain patient populations. PMID- 10868554 TI - Amprenavir: a new human immunodeficiency virus type 1 protease inhibitor. AB - OBJECTIVE: This paper reviews the pharmacologic properties and clinical usefulness of amprenavir, a new human immunodeficiency virus type 1 (HIV-1) protease inhibitor. BACKGROUND: Amprenavir, the most recent HIV-1 protease inhibitor to receive marketing approval from the US Food and Drug Administration, is a potent competitive inhibitor of HIV-1 protease and a relatively weak inhibitor of HIV-2 protease. Inhibition of the HIV-1 protease enzyme results in immature and noninfectious viral particles. Amprenavir is rapidly absorbed following oral administration. The time to peak concentration (Tmax) in adults is between 1 and 2 hours, the area under the plasma concentration versus time curve is roughly proportional to the dose, the half-life is approximately 8 hours, and the volume of distribution is approximately 430 L. The Tmax in children 4 to 12 years of age is between 1.1 and 1.4 hours. The bioavailability of the solution is 86% relative to the capsule formulation. It is metabolized by the cytochrome P 450 isozyme CYP3A4 and to a lesser extent by CYP2D6 and CYP2C9. METHODS: We searched MEDLINE (1966 to January 2000), AIDSLINE (1980 to January 2000), International Pharmaceutical Abstracts (1970 to January 2000), PharmaProjects (January 2000 version), and Web sites of major HIV/acquired immunodeficiency syndrome conferences for appropriate published references (1996 to February 2000). RESULTS: Data reported to date indicate that amprenavir is efficacious in the treatment of HIV disease in patients with primary HIV infection, antiretroviral-naive patients, protease inhibitor-naive patients, protease inhibitor-experienced patients, and pediatric patients. Adverse effects were usually of early onset (range, 2 to 21 days) and transient (range, 3 to 46 days), although the incidence of metabolic abnormalities such as lipodystrophy, hyperlipidemia, and diabetes mellitus has not yet been defined. Amprenavir should be avoided in patients with a known sulfonamide allergy. Concomitant use of other medications that are CYP3A4 inducers or inhibitors should be done cautiously and only if the potential benefit clearly outweighs potential risk. The dose should be reduced in patients with significant hepatic impairment (Child-Pugh score, > or = 5). Amprenavir probably should not be administered with rifabutin, rifampin, astemizole, midazolam , triazolam, bepridil, dihydroergotamine, ergotamine, or cisapride. The recommended adult dose is 1200 mg twice daily. For patients between 4 and 12 years of age or between 13 and 16 years of age who weigh < 50 kg, the recommended dosage of the capsule form is 20 mg/kg (22.5 mg/kg for oral solution) twice daily or 15 mg/kg (17 mg/kg for oral solution) 3 times a day to a maximum dose of 2400 mg (2800 mg for oral solution). Patients should not take vitamin E supplements because amprenavir is formulated with a large amount of vitamin E (109 IU/capsule and 46 IU/mL oral solution) to improve oral absorption. Amprenavir may be administered with or without food, but a high-fat meal (> 67 g fat) should be avoided. CONCLUSIONS: Published clinical data are limited, but amprenavir appears to be efficacious and generally well tolerated in patients with HIV infection. Pharmacoeconomic data are not yet available. The introduction of amprenavir appears to be important, since it provides an additional treatment option as a component of both initial and salvage combination therapies for patients with HIV. PMID- 10868555 TI - Loratadine versus cetirizine: assessment of somnolence and motivation during the workday. AB - OBJECTIVE: This parallel-group, double-blind study compared the somnolence and motivation profiles of 2 second-generation antihistamines, loratadine and cetirizine, in patients with allergic rhinitis. BACKGROUND: Second-generation antihistamines were developed to provide symptomatic relief from allergic disorders without the unwanted side effects of first-generation antihistamines, including somnolence. Recent research has indicated that not all second generation antihistamines are comparable with respect to somnolence and other cognitive processes. METHODS: Patients aged > or = 12 years and actively exhibiting symptoms of allergic rhinitis were randomized to 2 treatment groups to receive 10 mg loratadine or 10 mg cetirizine daily at 8:00 AM for 1 week. After patients took the medication, their somnolence and degree of motivation to perform activities were recorded in an electronic diary using a visual analog scale 4 times during the workday (8:00 AM, 10:00 AM, noon, and 3:00 PM). RESULTS: Sixty patients (31 men, 29 women) were randomized to treatment. Somnolence scores were similar for both groups at baseline and at the time of dosing (8:00 AM). However, there was a statistically significant difference in somnolence scores between the loratadine and cetirizine groups at 10:00 AM (P = 0.008), noon (P = 0.001), and 3:00 PM (P < 0.001), with the cetirizine group showing a greater degree of somnolence. The scores on motivation to perform activities were similar for both groups at the baseline and 8:00-AM measurements. In parallel with the somnolence scores, there were statistically significant differences in motivation scores between the loratadine and cetirizine groups at 10:00 AM (P = 0.014), noon (P = 0.001), and 3:00 PM (P < 0.001), indicating that patients taking loratadine were relatively more motivated during the workday. CONCLUSION: The results of this study demonstrate that in patients aged > or = 12 years who had allergic rhinitis, cetirizine use promoted somnolence and decreased motivation to perform activities during the workday compared with loratadine. PMID- 10868556 TI - Doses of olanzapine, risperidone, and haloperidol used in clinical practice: results of a prospective pharmacoepidemiologic study. EFESO Study Group. Estudio Farmacoepidemiologico en la Esquizofrenia con Olanzapina. AB - OBJECTIVE: The objectives of this study were to determine the doses of olanzapine (OLZ), risperidone (RIS), and haloperidol (HAL) used in clinical practice in outpatients with schizophrenia and the rates of occurrence of extrapyramidal symptoms (EPS) and other adverse events, clinical response, and use of concomitant medications. METHODS: The present study involved a subset of patients from a 6-month, open-label, prospective observational study. Data were collected by 293 psychiatrists at mental health centers and other outpatient treatment facilities in Spain. Medications and doses used, occurrence of EPS and other adverse events, and scores on the Clinical Global Impression (CGI) of Severity Scale and Global Assessment of Function (GAF) were recorded. Clinical response was defined as a decrease of > or = 2 points on the CGI, with a final CGI score < or = 4. RESULTS: A total of 2657 patients were included in the analysis. The initial and overall mean daily doses for the 3 groups were as follows: OLZ, 12.2 and 13.0 mg, respectively; RIS, 5.2 and 5.4 mg; and HAL, 13.9 and 13.6 mg. Initial and overall median daily doses were the same in each group: OLZ, 10 mg; RIS, 6 mg; and HAL, 10 mg. A significantly lower proportion of OLZ-treated patients (36.9%) experienced EPS compared with RIS-treated (49.6%) and HAL treated (76.0%) patients (P < or = 0.001). A significantly lower proportion of patients in the OLZ group (47.8%) experienced adverse events compared with patients in the RIS (57.2%) and HAL (79.8%) groups (P < or = 0.001). A significantly greater proportion of OLZ-treated patients (37.3%) were responders compared with RIS-treated patients (31.5%) (P < 0.05). In all 3 groups, patients who had an initial CGI score > or = 5 received significantly higher overall mean daily doses than did patients with an initial CGI score < 5 (P < 0.001). A significantly lower proportion of OLZ-treated patients (10.2%) were receiving concomitant anticholinergic medication at the end of the study (month 6) compared with RIS-treated (19.9%) and HAL-treated (44.0%) patients (P < 0.001). CONCLUSION: The mean daily doses recorded in this analysis based on data from a naturalistic setting are consistent with recommendations based on clinical trials. Compared with both RIS- and HAL-treated patients, OLZ-treated patients were less likely to experience EPS or other adverse events, and less likely to use concomitant anticholinergic medications. OLZ-treated patients were also more likely to respond to treatment than were RIS-treated patients. PMID- 10868557 TI - Combination hydrocodone and ibuprofen versus combination oxycodone and acetaminophen in the treatment of postoperative obstetric or gynecologic pain. AB - OBJECTIVE: The objective of this study was to compare the effectiveness of combination hydrocodone and ibuprofen with that of combination oxycodone and acetaminophen in the treatment of moderate to severe postoperative obstetric or gynecologic pain. BACKGROUND: Hydrocodone 7.5 mg with ibuprofen 200 mg is the only approved fixed-dose combination analgesic containing an opioid and ibuprofen. METHODS: This randomized, double-blind, parallel-group, single-dose, active-comparator, placebo-controlled study compared the effects of a 2-tablet dose of hydrocodone 7.5 mg and ibuprofen 200 mg with those of a 2-tablet dose of oxycodone 5 mg and acetaminophen 325 mg and placebo. Analgesia was assessed over 8 hours. RESULTS: Mean pain relief (PR) scores were similar for the hydrocodone with ibuprofen and oxycodone with acetaminophen groups (n = 61 and 59, respectively) at 0.5, 1, 1.5, 2, 2.5, 3, 4, and 7 hours and significantly greater for the hydrocodone with ibuprofen group at 5, 6, and 8 hours (P < or = 0.05). Mean pain intensity difference (PID) scores were similar for hydrocodone with ibuprofen and oxycodone with acetaminophen at 0.5, 1, 1.5, 2, 2.5, 3, and 4 hours and significantly greater for hydrocodone with ibuprofen at 5, 6, 7, and 8 hours (P < or = 0.05). Total PR scores were similar for hydrocodone with ibuprofen and oxycodone with acetaminophen for the 0- to 3- and 0- to 4-hour intervals and significantly greater for hydrocodone with ibuprofen for the 0- to 6- and 0- to 8 hour intervals (P < 0.05). The sum of the PID scores was similar for hydrocodone with ibuprofen and oxycodone with acetaminophen for the 0- to 3-, 0- to 4-, 0- to 6-, and 0- to 8-hour intervals. The median estimated time to onset of analgesia, mean peak PR score, median time to remedication, and mean global assessment score were similar for hydrocodone with ibuprofen and oxycodone with acetaminophen. Assay sensitivity was demonstrated by the presence of statistically significant differences between both active treatments and placebo (n = 60). The number of patients experiencing adverse events was similar for each of the 3 groups (11 [18.0%], hydrocodone with ibuprofen; 7 [11.9%], oxycodone with acetaminophen; and 6 [10.0%], placebo). CONCLUSIONS: In this study, a 2-tablet dose of combination hydrocodone 7.5 mg and ibuprofen 200 mg was as effective as a 2-tablet dose of combination oxycodone 5 mg and acetaminophen 325 mg in the treatment of moderate to severe postoperative obstetric or gynecologic pain. Both treatments were superior to placebo. The results of this study suggest that the combination of hydrocodone 7.5 mg and ibuprofen 200 mg may offer prescribers an additional option in combination pain therapy. PMID- 10868558 TI - The pharmacokinetics, electrocardiographic effects, and tolerability of loratadine syrup in children aged 2 to 5 years. AB - OBJECTIVE: We assessed the pharmacokinetics and tolerability of 5 mg loratadine syrup (1 mg/mL) in children aged 2 to 5 years. METHODS: Two studies were undertaken. A single-dose, open-label bioavailability study was performed to characterize the pharmacokinetic profiles of loratadine and its metabolite desloratadine. Plasma concentrations of loratadine and desloratadine were determined at 0, 1, 2, 4, 8, 12, 24, 48, and 72 hours after a single administration of 5 mg loratadine syrup to 18 healthy children (11 male, 7 female; 12 black, 5 white, 1 other; mean age +/- SD, 3.8 +/- 1.1 years; mean weight +/- SD, 17.4 +/- 4.4 kg). In addition, a randomized, double-blind, placebo controlled, parallel-group study was performed to assess the tolerability of 5 mg loratadine syrup after multiple doses. Loratadine (n = 60) or placebo (n = 61) was given once daily for 15 days to children with a history of allergic rhinitis or chronic idiopathic urticaria. In the loratadine group, 27 boys and 33 girls (52 white, 8 black) were enrolled, with a mean age +/- SD of 3.67 +/- 1.13 years and a mean weight +/- SD of 17.2 +/- 3.8 kg. In the placebo group, 27 boys and 34 girls (53 white, 7 black, 1 Asian) were enrolled, with a mean age +/- SD of 3.52 +/- 1.12 years and a mean weight +/- SD of 17.3 +/- 2.9 kg. Tolerability was assessed based on electrocardiographic results, occurrence of adverse events, changes in vital signs, and results of laboratory tests and physical examinations. RESULTS: The peak plasma concentrations of loratadine and desloratadine were 7.78 and 5.09 ng/mL, respectively, observed 1.17 and 2.33 hours after administration of loratadine; the areas under the plasma concentration-time curve to the last quantifiable time point for loratadine and desloratadine were 16.7 and 87.2 ng x h/mL, respectively. Single and multiple doses were well tolerated, with no adverse events occurring with greater frequency after multiple doses of loratadine than after placebo. Electrocardiographic parameters were not altered by loratadine compared with placebo. There were no clinically meaningful changes in other tolerability assessments. CONCLUSION: Loratadine was well tolerated in this small, selected group of children aged 2 to 5 years at a dose providing exposure similar to that with the adult dose (ie, 10 mg once daily). PMID- 10868559 TI - Effects on serum lipid profiles of continuous 17beta-estradiol, intermittent norgestimate regimens versus continuous combined 17beta-estradiol/norethisterone acetate hormone replacement therapy. AB - OBJECTIVE: To compare the effects of a daily oral 1-mg dose of continuous 17beta estradiol (E2) plus intermittent (3 days off, 3 days on) norgestimate (NGM) 90 microg (n = 221), an oral 2-mg dose of continuous E2 plus intermittent NGM 180 microg (n = 219), and an oral 2-mg dose of continuous E2 plus continuous norethisterone acetate (NETA) 1 mg (n = 217) on blood lipids and lipoproteins in postmenopausal women. BACKGROUND: The present study was undertaken because some progestins have adverse effects on lipid profiles, thereby negating the favorable effects of estrogens. METHODS: This was a multicenter, randomized, parallel-group trial that focused primarily on the 2 marketed regimens--E2 1 mg/NGM 90 microg and E2/NETA. Both subjects and investigators were blinded to the intermittent regimens; the continuous combined regimen was administered open-label. After a minimum 12-hour overnight fast, blood samples were collected at baseline and during months 7 and 12 to determine lipid and lipoprotein concentrations using validated methods. RESULTS: E2 1 mg/NGM 90 microg was associated with significant (ie, the 95% CI did not include 0) increases in high-density lipoprotein cholesterol (HDL-C) (6.8% [95% CI = 4.7%, 9.0%] and 4.8% [2.3%, 7.2%] at months 7 and 12, respectively) and high-density lipoprotein 2 cholesterol (HDL2-C) (10.8% [6.2%, 15.3%] and 24.1% [18.9%, 29.4%]) concentrations, and decreases in total cholesterol (-7.7% [-9.0%, -6.3%] and -9.2% [-10.5%, -7.9%]), low-density lipoprotein cholesterol (-14.3% [-16.3%, -12.4%] and -14.9% [-16.7%, -13.2%]), and lipoprotein(a) (-30.6% [-41.4%, -20.0%] at month 12) concentrations. A significant difference (P < 0.001 by analysis of variance) between the E2 1 mg/NGM 90-microg and NETA regimens was seen for HDL-C and HDL2-C concentrations, which were elevated in subjects receiving E2 1 mg/NGM 90 microg but reduced ( 9.1% [-11.1%, -7.1%] and -12.3% [-14.3%, -10.3%] for HDL-C at months 7 and 12, respectively; -14.2% [-18.0%, -10.4%] and -2.5% [-7.8%, +2.8%] for HDL2-C at months 7 and 12, respectively) in those receiving E2/NETA. CONCLUSIONS: In the present study, continuous E2 1 mg/NGM 90 microg was associated with beneficial effects on lipids and lipoproteins in healthy postmenopausal women, effects that were greater at least for HDL-C and HDL2-C than those observed with continuous combined E2/NETA. The applicability of the study results to women with preexisting cardiovascular disease or dyslipidemia, or those who are overweight, remains to be investigated. PMID- 10868560 TI - An outpatient prescription-medication benefit in Medicare: so much time, so little consensus. PMID- 10868561 TI - The effect of new and continuing prescription drug use on cost: a longitudinal analysis of chronic and seasonal utilization. AB - OBJECTIVE: To provide basic information about 2 factors contributing to rising prescription drug costs--utilization trends and product selection. BACKGROUND: Prescription drug costs have risen sharply in recent years, and continued growth is expected. There is little consensus about appropriate cost-management strategies, in part because quantitative data on the causes and implications of increased drug costs are lacking. METHODS: This study followed 463,820 continuously enrolled adult (> or = 18 years of age on January 1, 1996) utilizers of 15 chronic or seasonal therapeutic classes for 2.5 years (January 1996 through June 1998) using a pharmacy benefit manager's multiple-payer claims database. Outcome measures included (1) change in utilization rate, (2) relationship between new use and utilization growth, (3) stability of the treated population (ie, mostly long-term use vs high rates of turnover), and (4) product mix changes (ie, cost per dispensed day for 1996 vs 1997 and for new vs continuing users, controlling for inflation). RESULTS: Of the 463,820 utilizers, 97% were commercially insured and 3% enrolled in Medicare risk plans; 40% were enrolled in managed care and the remainder covered by indemnity insurance. Rates of growth and turnover varied substantially by class. The highest 2-year utilization rate change was 66.7% for antihyperlipidemic agents; change was < 10% in only 3 classes. Across classes, an average of 38.7% of 1997 users were new (ie, no use in 1996) and an average of 34.0% of 1996 users were dropouts (ie, no use in 1997). Utilization growth depended heavily on treatment continuation; classes with high dropout rates (eg, antirheumatic, antiasthmatic) did not have high growth rates, even with high rates of new use. In most classes, costs were not higher for new than for continuing users. In some classes, however (eg, antipsychotic, antidiabetic), both new and continuing users increased their use of newer, more expensive products. CONCLUSIONS: Because factors underlying rising prescription drug costs vary by therapeutic class, cost-containment strategies should address these differences. Further research is needed to assess the clinical and economic costs and benefits of rapid growth in the utilization of certain therapeutic classes. PMID- 10868562 TI - Spectral editing of 13C cp/MAS NMR spectra of complex systems: application to the structural characterisation of cork cell walls. AB - A mathematical method of obtaining 13C CP/MAS subspectra of single components of a complex system is presented and applied to three- and four-component systems. The method is based on previously reported work that exploits different proton relaxation properties for different domains of an heterogeneous system. However, unlike the original method that obtained subspectra through a trial-and-error approach, the method here presented solves the problem mathematically, thus avoiding the time-consuming and non-rigorous trial-and-error step. The method is applied to mixtures of three and four polymers and to a more complex system: cork cell walls. As expected, as the number of components increases, the sharing of relaxation properties between different components is increasingly probable, either due to incidental coincidence of relaxation times or to specific interactions and intimate mixing of compounds. While this hinders the calculation of the subspectra of single chemical components, it may provide useful information about inter-component interactions. This possibility was demonstrated by the application of this method to cork cell walls. Both three-component and four-component approaches showed that three domains exist in cork cell walls: carbohydrate/lignin matrix, mobile suberin close to (probably bonded to) lignin groups (about 42% w/w) and hindered suberin close to (probably bonded to) carbohydrate-OCH2O groups (about 4% w/w). PMID- 10868563 TI - Comparative study of motions in dimethylsulfone by noise excitation and solid echo spectroscopy. AB - 2H NMR spectra of dimethylsulfone were measured with noise excitation and solid echo NMR spectroscopy in the temperature range from 125 to 355 K. Besides the known fact that broad NMR spectra can be measured with both methods, in comparable times it is shown that for noise excitation, the signal loss is negligible compared to echo spectroscopy in the regime when the correlation times of the motions are of the order of magnitude of the echo pulse spacing. For simulating the dynamic NMR spectra acquired with noise excitation, only the motional process must be taken into account and relaxation can be neglected. Furthermore, the problem of restricted acquisition bandwidth in noise NMR spectroscopy is discussed. PMID- 10868564 TI - 13C high-resolution solid state NMR studies of the structure and dynamics of 1,6:3,4-dianhydro-2-O-tosyl-beta-D-galactopyranose. AB - 13C CP/MAS, dipolar dephasing MAS and theoretical GIAO calculations were employed to assign 13C resonances to the molecular structure of 1,6:3,4-dianhydro-2-O tosyl-beta-D-galactopyranose 1. From spinning sideband intensities, employing the graphical method of Herzfeld and Berger the 13C delta(ii) parameters for aromatic residue were calculated. The experimental data were compared with computed results obtained by means of the B3PW91 hybrid method and 6-311G (df, p) basis set. The X-ray geometry of 1 with the correlated position of hydrogen atoms was taken as input data for theoretical calculations. As concluded from Cambridge Crystallographic Database (CSD) search, there are two reports describing the X ray studies of 1 that show the slightly different geometry of the compound under investigation. This work shows that such discrepancies in geometry can generate differences between computed 13C delta(ii) parameters up to 6 ppm. 13C T1 and 1H T1rho relaxation times reveal that 1 is very rigid in crystal lattice. This structure is characterized by extremely long 1H T1rho, found to be in range ca. 200 ms. PMID- 10868565 TI - Moment analysis as a systematic tool for NMR powder pattern analysis. AB - The low order moments for chemical shift and second-order quadrupolar powder patterns have been calculated as functions of the anisotropy and asymmetry parameter of the governing interaction, and the expressions inverted to give these parameters as a function of the moments. Theoretical simulations and experimental experience show that moment analysis in most cases equals and in some cases exceeds the accuracy of direct inspection as a method of obtaining NMR parameters. We illustrate the efficacy of the method applied to 31P chemical shift spectra of nucleic acids, and 39K second-order patterns of series of potassium salts. PMID- 10868566 TI - Heating of samples induced by fast magic-angle spinning. AB - Intense sample heating through high-speed magic-angle spinning (MAS; up to 58 K temperature difference) is demonstrated. The role of probehead and spinner design, as well as that of the temperature of the bearing air on the heating of a rotating sample, is examined. MAS-induced heating can affect the accurate determination of the isotropic value of the chemical shift as well as the principal values, asymmetry and anisotropy parameters of the chemical shift tensor. In some cases, a very large temperature gradient (12 K) within the fast rotating sample was found, which may limit the resolution of high-speed 1H MAS nuclear magnetic resonance (NMR) spectra. PMID- 10868567 TI - Effect of hydrostatic pressure on N(CH3)4+ cation motion in ferroelectric N(CH3)4H(Cl3CCOO)2. AB - The proton spin-lattice relaxation time in ferroelectric N(CH3)4H(Cl3CCOO)2 has been studied under isobaric conditions at pressures 0.1, 200 and 400 MPa over a wide range of temperature. The data indicate that the dominant relaxation mechanism for T1 can be attributed to the classical CH3 group reorientation of N(CH3)4+ cation. The influence of pressure on methyl group reorientation of N(CH3)4+ cation was analysed. PMID- 10868568 TI - Heteronuclear 1H-13C spin echoes mediated by cross-polarization. AB - Heteronuclear spin echoes having the spin coherence transferred by means of cross polarization (CP) under Hartmann-Hahn condition are reported. The particular characteristic of heteronuclear spin echoes is that the dephasing and refocusing processes take place within different nuclear species. Originally, they were demonstrated using the heteronuclear scalar interaction to transfer the spin coherence between the two spin systems. In this work, it is shown that spin coherence transferred by CP can also lead to the formation of heteronuclear spin echoes. A semiclassical formalism was used to explain the mechanism of generation of the spin echoes. These spin echoes were observed so far in elastomeric materials. PMID- 10868569 TI - NMR determination of the Sternheimer antishielding factor of Ca2+. AB - The experimental determination of the Sternheimer antishielding factor gamma(infinity) for Ca2+ in a sold material is reported for the first time. The gamma(infinity) value was obtained by comparing the quadrupole frequency measured by 43Ca NMR with the electric field gradient calculated in a frame of the point charge approximation in the high-Tc superconducting compound, Tl0.5Pb0.5Sr2CaCu2O7. The deduced value (gamma(infinity)= -6.1+/-.9) is about one third of that obtained from the quantum mechanical calculations, whereas in the same sample, the gamma(infinity)value for O2- deduced by the same procedure (gamma(infinity)= -13.7+/-2.0) is very close to the calculated one. PMID- 10868570 TI - Characterization of 15N chemical shift tensors via 15N-13C REDOR and 1N-1H dipolar-shift CPMAS NMR spectroscopy. AB - As part of our studies on the characterization of 15N chemical shift anisotropy (CSA) via magic angle spinning (MAS) NMR spectroscopy, we have investigated via numerical simulations the sensitivity of two different REDOR experimental protocols to the angles defining the orientation of the 15N-13C' bond vector in the principal axis system of the 15N CSA tensor of the amide nitrogen in a peptide bond. Additionally, employing polycrystalline samples of 15N and 13C', 15N-labeled acetanilide, we have obtained, in a first study of this type, the orientation of the 15N CSA tensor in the molecular frame by orienting the tensor with respect to the 15N-3C' and 15N-1H dipolar vectors via 15N-13C' REDOR and 15N 1H dipolar-shift MAS experiments, respectively. PMID- 10868571 TI - Through-bond connectivity in solids by continuous-wave spin lock. AB - A simple two-dimensional correlation experiment that enables determination of through-bond connectivity in the solid state is described. The experiment is performed under fast magic angle spinning (MAS) conditions. After the initial pi/2 pulse, the magnetization develops freely under the MAS Hamiltonian. The t1 period is followed by a strong spin locking pulse used as mixing period. The dipolar coupling is averaged out by magic angle spinning, and the chemical shifts and r.f.-offsets are scaled by the applied spin locking field. Hence, for strong locking conditions, the isotropic J-coupling is the dominant interaction. The mixing Hamiltonian is thus identical to the well-known TOCSY-Hamiltonian, resulting in a net through-bond magnetization transfer. The mixing-time dependence of the exchange rates is investigated. Applications to crystalline P4S7 and MgP4O11 are shown. PMID- 10868572 TI - Spin diffusion of dipolar energy in NQR. AB - The theory of spin diffusion was extended to the case of nuclear dipolar order in solids containing paramagnetic impurities and nuclei with spin I > 1/2 having nuclear quadrupole moment. We show that spin diffusion process of dipolar order takes place in solids containing paramagnetic impurities. At the start of relaxation process, the direct relaxation regime is realized with non-exponential time dependence. Then the relaxation regime will be changed to diffusion-limited one. Using obtained expressions for the spin lattice relaxation times for these two relaxation regimes, the diffusion coefficient of the dipolar order in nuclear quadrupole resonance can be estimated from experimental data. PMID- 10868573 TI - Multiple-quantum MAS NMR of quadrupolar nuclei. Do five-, seven- and nine-quantum experiments yield higher resolution than the three-quantum experiment? AB - The question of whether or not higher-order (five-, seven- and nine-quantum) multiple-quantum magic angle spinning (MQMAS) experiments yield isotropic NMR spectra of half-integer quadrupolar nuclei with higher resolution than the basic three-quantum MAS experiment is examined. The frequency dispersion is shown theoretically to be greatly increased in higher-order MQMAS spectra, but it is argued that whether or not this translates into an increase in resolution depends upon the ratio of the homogeneous to inhomogeneous contributions to the isotropic linewidth. Experimentally, it is demonstrated using three-, five- and seven quantum 45Sc MAS NMR and three- and five-quantum 27Al MAS NMR of crystalline samples that higher-order MQMAS experiments can yield a real and useful increase in resolution but that, owing to the presence of inhomogeneous broadening in the isotropic spectra, this increase is less than the theoretically predicted value. A number of practical issues relating to resolution in MQMAS NMR are also pointed out. PMID- 10868574 TI - Determination of histamine in tomatoes by liquid chromatography/mass spectrometry. AB - The histamine level in tomato fruits and pastes was determined by 2 orthogonal techniques as a means of comparing accuracy. Close statistical agreement was found between assays for free histamine by capillary electrophoresis (with UV absorbance detection), and for the dansyl derivative by reversed-phase liquid chromatography (LC). Both techniques have adequate sensitivity for the analysis of endogenous histamine in tomatoes, but LC/electrospray tandem mass spectrometry was more sensitive by at least an order of magnitude, down to a level of 0.05 mg/kg. PMID- 10868575 TI - Validation of a method for determination of phospholipase A2 and melittin in bee venom preparations by capillary electrophoresis. AB - A method was validated for the determination of the 2 main components of bee venom, phospholipase A2 and melittin, by capillary electrophesis (CE). Optimum resolution and selectivity were attained with a running electrolyte of 150 mM phosphoric acid, pH 1.8. The repeatability and day-to-day reproducibility of the migration times were better than 0.36 and 2.8%, respectively. The repeatability and day-to-day reproducibility of the normalized peak areas were better than 1.3 and 2.6%, respectively. The response of the UV detector at 190 nm was linear over < 2 concentration decades, from 0.05 to 1.5 mg/mL, with correlation coefficients of 0.9994 for phospholipase A2 and 0.9997 for melittin. The limits of detection and quantitation were 4.5 and 15 microg/mL, respectively, for phospholipase A2 and 1.6 and 6 microg/mL, respectively, for melittin. The reproducibility of the measurements with 2 different CE instruments was satisfactory; the mean concentration and relative standard deviation (RSD) values for phospholipase A2 and melittin were 14.4% (RSD, 1.3%) and 51.4% (RSD, 1.1%), respectively, with instrument I; the corresponding values with instrument II were 14.5% (RSD, 2.8%) and 52.3% (RSD, 2.2%). The accuracy was estimated by comparison with a liquid chromatographic (LC) method. Differences between the CE and LC measurements were attributed to irreversible adsorption of the analytes on the LC column. The recoveries of phospholipase A2 and melittin with the CE method were 98.8 and 101.7%, respectively. PMID- 10868576 TI - Determination of tilmicosin residues in cow and sheep milk by liquid chromatography. AB - This method was developed and validated to detect and quantitate tilmicosin residues in cow milk over a range of 0.010-10 microg/mL, and in sheep milk over a range of 0.025-0.5 microg/mL. The procedure involves extracting the milk sample with acetonitrile and using a C18 cartridge to perform a solid-phase extraction cleanup of the extract. A reversed-phase gradient liquid chromatography method with detection at 280 nm is used to separate the tilmicosin from matrix components in a 30 min run time. The limit of quantitation of the method is 0.010 microg/mL for cow milk, and 0.025 microg/mL for sheep milk. Average percentage recoveries for milk samples ranged from 82 to 94%. Percentage relative standard deviation values ranged from 3.1 to 17.2%. PMID- 10868577 TI - Method for determination of xanthene dyes in guava fruits and its application in a field dissipation study. AB - Xanthene dyes, i.e., phloxine B and uranine or phloxine B alone, are phototoxic to tephritid fruit flies infesting guava fruits. An analytical method was developed for determination of residues of these dyes used in bait solutions for suppression of the tephritid fruit fly population in guava fruits. The procedure involved solvent extraction, anion-exchange cleanup, and determination by liquid chromatography or capillary zone electrophoresis. The dyes were extracted from 50 g guava fruit at 45 degrees degrees with 400 mL methanol-acetonitrile (1 + 1) and 5 g magnesium oxide added as an alkaline and clarifying agent. The guava extract was adjusted to pH 8.5 and subjected to an amino column cleanup. Average recoveries of xanthene dyes added to guava purees ranged from 77 to 99% for phloxine B and from 79 to 102% for uranine at spiking levels of 0.05-1.00 microg/g. The method was applied to the determination of phloxine B residues in guava fruits collected from a dye-sprayed orchard. After phloxine B was applied at a rate of 62.5 g/ha for 14 weekly sprayings, it was found on guava fruits at an average concentration of 111 +/- 18 ng/g 4 h after the llth spraying. The concentration of phloxine B was 426 +/- 94 ng/g in selected fruits with high deposits of the dye 4 h after spraying. Average concentrations of phloxine B 5 days after the 7th and 14th sprayings were 29 +/- 7 and 19 +/- 8 ng/g, respectively. PMID- 10868579 TI - Evaluation of a hypercrosslinked polystyrene, MN-200, as a sorbent for the preconcentration of volatile organic compounds in air. AB - Breakthrough volumes, average percentage recoveries, and storage stabilities were obtained for vapors of 8 volatile organic compounds (pentane, octane, undecane, isooctane, cyclohexane, toluene, methanol, and dichloromethane) on a new adsorbent material, Hypersol-Macronet, MN-200. Breakthrough volumes were estimated as half of the gas chromatographic specific retention volumes at 20 degrees C for the compounds. Recoveries of the adsorbates were determined by both solvent extraction and thermal desorption methods. The results obtained compare favorably with those for Tenax GR (values reported in the published literature and others obtained in our laboratory). Results of storage stability studies on MN-200 meet the criterion for acceptability (<10% loss). High adsorption capacity for very volatile and polar compounds, combined with ease of desorption of less volatile compounds, render MN-200 a highly promising adsorbent for sampling volatile organic compounds in indoor and outdoor air. PMID- 10868578 TI - Estimation of residues of carbofuran and its metabolites in sugarcane and soil by derivatization with 1-fluoro-2,4-dinitrobenzene and gas chromatography with nitrogen-phosphorus detection. AB - Residues of carbofuran and its metabolites were studied in sugarcane plants and soil after application at 1 and 2 kg/ha. The residues of carbofuran and its metabolites were extracted by refluxing with 0.25N HCI, partitioned into dichloromethane, and cleaned up on acidic alumina. The respective 7-phenols of carbofuran, 3-ketocarbofuran, and 3-hydroxycarbofuran were destroyed by treatment with ceric ammonium sulfate, and the residues were derivatized with 1-fluoro-2,4 dinitrobenzene. The derivatives were estimated by gas chromatography with nitrogen-phosphorus detection. The concentration of 3-hydroxycarbofuran in sugarcane plants remained higher and persisted longer than that of the parent compound. Carbofuran-derived residues were not detected in cane juice. Soil samples were found to contain only carbofuran, which declined at a very fast rate that followed a first-order kinetics rate of reaction. PMID- 10868580 TI - Methodology for studying oxidation of organic species in solution. AB - A photoreactor was developed to study products of photochemical oxidation in a wide range of organic compounds. Analysis of the products from the reactor were used to determine the extent of mineralization of the organic material, to characterize any intermediate compounds formed, and to obtain information on the decomposition mechanism. Appropriate methods for separation and characterization include LC, UV spectrophotometry, gas chromatography/mass spectrometry, total organic carbon, and total inorganic carbon. The uses of the reactor are described for the photocatalytic decomposition of phenol and of its major decomposition intermediates 1,2- and 1,4-dihydroxybenzene. PMID- 10868581 TI - Determination of hydroperoxides in liposomes by the modified IDF and the modified tiron methods. AB - Two new methods, the modified International Dairy Federation and the modified Tiron methods, were developed for the determination of hydroperoxides in liposomes. Interferences of alpha-tocoferol and egg-phosphatidylcholine (EPC) required a solid-phase extraction before the analysis to eliminate alpha tocoferol and a fluid-fluid extraction based on the solvent triangle of Bligh and Dyer to separate EPC. The developed color, using thiocyanate and Tiron, respectively, as complex-formers for the generated ferri-ions, was measured spectrophotometrically. Both techniques showed good reproducibility and high sensitivity. Peroxide contents of 0.04 and 0.07% (g/g) in EPC samples were easily determined. PMID- 10868582 TI - Evaluation of silica- and sepharose-based immunoaffinity sorbents for sample cleanup in determination of fumonisins B1 and B2 in corn products. AB - Anti-fumonisin B1 polyclonal antibodies were isolated from the serum of rabbits, immobilized onto the surface of glutaraldehyde-activated silica or Sepharose CL 4B particles, and placed into empty small plastic solid-phase extraction cartridges. The immobilized antibodies were evaluated for their ability to retain fumonisin B1 and fumonisin B2. Cartridge capacity and elution conditions were determined, and the results were compared to those obtained with a commercially available cartridge. The cartridges, which were tested for their effectiveness to isolate the fumonisins from extracts of corn flour and nacho chips, detected fumonisins down to levels of about 20 ng/g. However, additional cleanup was required for detection at lower concentrations. With the use of a strong anion exchange cartridge as a preliminary cleanup before immunoaffinity chromatography, the detection limit reached 2-5 ng/g in the products tested. The silica sorbent material exhibited strong interactions with the fumonisins, requiring acidified ethanol-water mixtures for elution and resulting in an additional degree of selectivity in isolating fumonisins from sample extracts. The silica-based immunoaffinity cartridges were successfully reused more than 10 times; the Sepharose-based cartridges were less robust. Liquid chromatography with fluorescence detection was used after prechromatographic derivatization with o phthaldialdehyde-mercaptoethanol. PMID- 10868583 TI - Effect of temperature and solvent composition on extraction of fumonisins B1 and B2 from corn products. AB - Fumonisins B1 and B2 were extracted from naturally contaminated corn products by using different extraction solvent compositions (methanol-water, acetonitrile methanol-water, ethanol-water, and 100% water) and a range of temperatures from ambient to 150 degrees C. Ground samples of several corn products and 1 rice sample were mixed with an adsorbent material (Hydromatrix), and the fumonisins were extracted in 2 sequential 5 min static extractions at various temperatures. The combined extracts were cleaned up and analyzed by reversed-phase liquid chromatography with fluorescence detection after o-phthaldialdehyde mercaptoethanol derivatization. The results showed a clear influence of temperature and solvent composition on recovery of fumonisins from some matrixes. With acetonitrile-methanol-water (1 + 1 + 2) the quantity of fumonisins extracted from naturally contaminated taco shells almost tripled in going from 23 degrees to 80 degrees C, and increased by another 30% when ethanol-water (3 + 7) was used as extraction solvent at 80 degrees C. Similar results were obtained with nacho chips. These effects were less pronounced with cornmeal, and small differences due to temperature and solvent composition were observed for corn flakes and rice. The ethanol-water extraction solvent combinations were specifically evaluated in an effort to use the cheapest, least toxic, and most environmentally friendly solvents for organic residue analysis. At 80 degrees C, ethanol-water combinations performed equally or better than methanol-water (8 + 2) or acetonitrile-methanol-water (1 + 1 + 2), combinations which are commonly used for fumonisin extractions. Even 100% water was successful for extracting fumonisins from the products, except for rice. However, increased amounts of water created technical problems and required an increased amount of Hydromatrix in the samples prior to extraction. PMID- 10868584 TI - Confirmation of patulin and 5-hydroxymethylfurfural in apple juice by gas chromatography/mass spectrometry. AB - A gas chromatographic/mass spectrometric (GC/MS) method was developed for the confirmation of patulin and 5-hydroxymethylfurfural (HMF) extracted from apple juice. The extraction is based on the official AOAC method for liquid chromatographic analysis. Juice extracts are quickly and easily derivatized with bis(trimethylsilyl)trifluoracetamide under mild conditions, and the trimethylsilyl ethers of the analytes are stable for at least several hours. The analytes are determined by GC/MS using an electron-impact source and selected ion monitoring of characteristic ions. For both analytes, the interassay differences between base-peak ratios for samples and standards were all <7.1% (absolute). The presence of patulin was confirmed at fortification levels of about 30-400 microg/L and naturally occurring levels of about 80-400 microg/L. The presence of HMF was also confirmed at levels < or = 2 mg/L. The proposed mass spectral fragmentation pathways of the analytes are presented. PMID- 10868585 TI - Characterization of varietal differences in essential oil components of hops (Humulus lupulus) by SFC-FTIR spectroscopy. AB - A supercritical fluid chromatographic method combined with Fourier-transform infrared spectroscopy detection (SFC-FTIR) was developed for determination of varietal differences in essential oil constituents in hops (Humulus lupulus). Infrared spectra (IR) of the major constituents of essential oil of hops were taken as films deposited on AgCl discs and compared with those obtained after chromatographic separation in the IR flow-cell with supercritical carbon dioxide (scCO2). Spectra from AgCl discs were comparable to those in scCO2, but in scCO2 most of the bands appeared approximately 8-10 cm-1 to higher wave numbers. Open tubular SFC-FTIR analysis of the essential oil of 4 different hop varieties was performed. The SFC-FTIR chromatograms showed differences in the location and relative intensity of the peaks depending on the variety, which was further confirmed by consideration of their FTIR spectra. PMID- 10868586 TI - Liquid chromatographic determination of tocopherols and tocotrienols in vegetable oils, formulated preparations, and biscuits. AB - A precise and selective liquid chromatographic procedure for determining tocopherol and tocotrienol isomers in vegetable oils, formulated preparations, and biscuits was developed and validated. The proposed method quantitates vitamin E in better conditions of recoverability and reproducibility than the standard saponification procedure. Tocopherols and tocotrienols were extracted in hexane from vegetable oils, passed through a silica Sep-pak, chromatographed on a mu Bondapak C18 column with a mobile phase of methanol-water (95 + 5, v/v), identified at 292 nm, and detected with fluorescence procedure (excitation 296 nm, and emission 330 nm). The correlation coefficient on the calibration curve was 0.9995 over the range of 0.1 to 100 microg/mL. Overall recovery of vitamin E isomers was 93%; coefficients of variation for intra- and interday precision, < 2.25%. The results obtained from extraction methods 1 (with saponification) and 2 (without saponification) were compared by ANOVA test. Significant differences appeared between vitamin E isomers (p < or = 0.05). PMID- 10868587 TI - Determination of the levels of isoflavonoids in soybeans and soy-derived foods and estimation of isoflavonoids in the Japanese daily intake. AB - The levels of 6 kinds of isoflavonoids found in 11 domestic and imported soybeans, and 12 kinds of soybean-based processed foods in Japan were systematically analyzed, and the Japanese daily intake of isoflavonoids from those foods was estimated. The total isoflavonoids (daidzein, glycitein, and genistein) were analyzed with acid hydrolysis and the intact isoflavonoids (daidzein, glycitein, genistein, daidzin, glycitin, and genistin) were analyzed without hydrolysis. This was followed by cleanup with an ODS cartridge column and determined by liquid chromatography with a diode array detector. The highest content of isoflavonoids was found in kinako (a roasted soybean powder) and the lowest was found in soy sauce. The contents and composition of the isoflavonoids in the 11 soybeans varied by species and country of origin. The level of isoflavonoids found in the processed foods varied by manufacturing method or ingredients. The percentage of aglycone tended to be higher in miso (fermented soybean paste) and soy sauce, which are heated and fermented during the manufacturing process. Japanese daily intake of isoflavonoids from soybeans and soybean-based processed foods was estimated as 27.80 mg per day (daidzein 12.02 mg, glycitein 2.30 mg, and genistein 13.48 mg). PMID- 10868588 TI - Coupling of size-exclusion chromatography to liquid chromatography/mass spectrometry for determination of trace levels of thifensulfuron-methyl and tribenuron-methyl in cottonseed and cotton gin trash. AB - Size-exclusion chromatography (SEC) was coupled to reversed-phase liquid chromatography/mass spectrometry for the determination of thifensulfuron-methyl and tribenuron-methyl in cottonseed and cotton gin trash. The limit of quantitation was 20 parts per billion (ppb), and the limit of detection was 6 ppb. The analytes were extracted by homogenization in a buffer solution. The extracts underwent a solvent exchange into methanol and were injected onto an SEC column. As the analytes eluted from the SEC column, the eluate was diverted onto a reversed-phase column for additional separation of the analytes and their detection via mass spectrometry. This method is unique because the samples are not cleaned up before analysis, the analytes are injected in methanol, and the entire analysis is completed in 30 min. Average recoveries and standard deviations for thifensulfuronmethyl and tribenuron-methyl in cotton gin trash were 91 +/- 6% and 88 +/- 5%, respectively. Average recoveries and standard deviations for thifensulfuron-methyl and tribenuron-methyl in cottonseed were 91 +/- 11% and 99 +/- 12%, respectively. This is an effective method for the detection and determination of thifensulfuron-methyl and tribenuron-methyl in cotton. PMID- 10868589 TI - Averaging the closest two out of three observations. AB - The average of 3 observations is the standard method of estimating the location of their distribution. The average of the closest 2 out of 3 observations, although often used, has more variability than the average of all 3 observations for a standard normal distribution. One statistic uses a threshold to decide when to use the average of all 3 observations rather than the closest 2. With the standard normal, this statistic still has more variability than the average of all 3 observations. PMID- 10868590 TI - Comparative study of epithelial and fibroblastic cell lines as an alternative cytotoxicity test to the Draize method. AB - Several methods are being used with considerable advantage as alternatives to the Draize test, although some technical difficulties still persist. This work compared the sensitivity of HeLa and NCTC L 929 cells to evaluate the cytotoxicity of shampoos used by adults and children (undiluted and diluted to 25, 5, 1, and 0.1%), and eye drops and their containers and surfactants (diluted to 30, 10, 1, and 0.1%). Nondiluted adult shampoos and their 25 and 5% dilutions were cytotoxic for both cell lines. When diluted to 1%, only one of the shampoos was noncytotoxic, whereas among those diluted to 0.1%, only one was cytotoxic. Children's shampoos were cytotoxic when not diluted or diluted to 25%. From those diluted to 5%, only one was noncytotoxic for both cell lines. The cytotoxic tests showed that the eye drops and their containers were noncytotoxic. Surfactants were cytotoxic when diluted to 30 and 10% and noncytotoxic when diluted to 1 and 0.1%. An excellent correlation (r = 0.95) was demonstrated between the sensitivity of the HeLa and NCTC L929 cells in the evaluation of cytotoxicity reactions. PMID- 10868592 TI - Determination of inulin in foods. AB - A method was developed for determining fructan inulin in various foods (yogurts, honey cakes, chocolates). Warm water was applied for extraction of samples, and mono- and dissacharides were determined by a thin-layer chromatographic densitometric method. A portion of the test solution was hydrolyzed 30 min with 1% oxalic acid in a boiling water bath. Fructose was determined in the hydrolysate. The amount of inulin in a sample was calculated as the difference between the amount of fructose in the sample before and after hydrolysis. The fructose from sucrose formed during the hydrolysis was also considered. The mean recovery from yogurt fortified with 4% inulin was 95.5 +/- 4.5% (mean +/- standard deviation); from honey cakes extract fortified with 10% inulin, 97.3 +/- 5.5%; and from chocolate extract fortified with 30% inulin, 98.6 +/- 6.6% (6 replicates in all cases). Determination of glucose is not necessary for analyzing fructans with the composition expressed shortened to GFn-1 (G, glucose; F, fructosyl) with the average degree of polymerization 8 < or = n < or = 15. PMID- 10868591 TI - Color measurement of rosemary honey in the solid state by reflectance spectroscopy with black background. AB - A colorimetric method was developed to measure the color of rosemary honeys in the solid state, without liquefaction. The color of 20 solid samples of rosemary honeys was measured by reflectance spectroscopy with white and black background in cells of 1 cm pathlength. The Kulbelka-Munk theory of turbid media was applied to calculate the spectral distribution of reflectivity, internal transmittance, and coefficients of absorbance and scattering of light. From these spectral distributions, 2 different types of honey were found. The honey colors measured from reflectivity and internal transmittance are well grouped. The colors obtained from reflectivity and those obtained from reflectance with black background show high linear correlation (r2 > 0.99). As a practical application, measurements of reflectance with black background in cells of 1 cm pathlength can be used to determine the color of these honeys in the solid state. PMID- 10868593 TI - Analysis of pesticide residues in mixed fruit and vegetable extracts by direct sample introduction/gas chromatography/tandem mass spectrometry. AB - Direct sample introduction (DSI), or "dirty sample injection," was investigated in the determination of 22 diverse pesticide residues in mixed apple, green bean, and carrot extracts by benchtop gas chromatography/tandem mass spectrometry (DSI/GC/MS-MS). The targeted pesticides, some of which were incurred in the samples, included chlorpyrifos, azinphos-methyl, parathion-methyl, diazinon, terbufos, p,p'-DDE, endosulfan sulfate, carbofuran, carbaryl, propargite, bifenthrin, dacthal, trifluralin, metalaxyl, pendimethalin, atrazine, piperonyl butoxide, diphenylamine, vinclozolin, chlorothalonil, quintozene, and tetrahydrophthalimide (the breakdown product of captan). The analytical DSI method entailed the following steps: (1) blend 30 g sample with 60 mL acetonitrile for 1 min in a centrifuge bottle; (2) add 6 g NaCl and blend 30 s; (3) centrifuge for 1-2 min; (4) add 5 mL upper layer to 1 g anhydrous MgSO4 in a vial; and (5) analyze 11 microL extract, using DSI/GC/MS-MS. Sample cleanup is not needed because GCIMS-MS is exceptionally selective for the targeted analytes, and nonvolatile coextracted matrix components do not contaminate the injector or the GC/MS-MS system. Average recoveries of the pesticides were 103 +/- 7% with relative standard deviations of 14 +/- 5% on average, and limits of detection were <2 ng/g for nearly all pesticides studied. The DSI/GC/ MS-MS approach for targeted pesticides is quantitative, confirmatory, sensitive, selective, rugged, rapid, simple, and inexpensive. PMID- 10868595 TI - Multiresidue method for the gas chromatographic determination of pesticides in honey after solid-phase extraction cleanup. AB - A new multiresidue method is described for the determination of pesticides in honey. The method involves dissolution of the honey in a methanol-water mixture, followed by solid-phase extraction cleanup and gas chromatographic determination. Twenty-six pesticides used on flowering field crops, on flowering fruit and vegetables, or as acaricides to control Varroa jacobsoni in beehives are determined by the method. Recoveries from honey, spiked at 0.02-1.6 mg/kg, ranged from 85 to 127% with a relative standard deviation (RSD) of 2-16%, except for the RSD of 27% for captan at 0.05 mg/kg. PMID- 10868594 TI - Multiresidue method for the determination of residues of 251 pesticides in fruits and vegetables by gas chromatography/mass spectrometry and liquid chromatography with fluorescence detection. AB - A method is described for the determination of 251 pesticide and degradation product residues in fruit and vegetable samples. Extraction of the sample with acetonitrile is followed by a salting-out step. Co-extractives are removed by passing a portion of the acetonitrile extract through an octadecyl (C18) solid phase extraction cleanup cartridge and then, in a second cleanup, through a carbon cartridge coupled to an amino propyl cartridge. Determination is by gas chromatography with mass-selective detection in the selected-ion monitoring mode, and by liquid chromatography with post-column reaction and fluorescence detection for N-methyl carbamates. The method has been used for analysis of various fruits and vegetables, such as apple, banana, cabbage, carrot, cucumber, lettuce, orange, pear, pepper, and pineapple. Limits of detection range between 0.02 and 1.0 mg/kg for most compounds. Over 80% of the compounds have a limit of detection of < or = 0.04 mg/kg. PMID- 10868596 TI - Selective trace determination of dithiocarbamate fungicides in fruits and vegetables by reversed-phase ion-pair liquid chromatography with ultraviolet and electrochemical detection. AB - The predominant methods for determination of dithiocarbamate fungicides (DTC) have been based on quantitation of carbon disulfide released by hot acid digestion. Because the subgroups of the DTC family differ in their chemical properties and toxicological behavior, selective determination is required. To meet the demand for a fast, simple, and sensitive procedure, a new reversed-phase ion-pair chromatographic method was developed, consisting of surface extraction followed by direct injection into a liquid chromatographic system equipped with ultraviolet and electrochemical detectors, connected in series. The procedure is applicable to residues of N-methyldithiocarbamates (metam-sodium), N,N dimethyldithiocarbamates (e.g., ziram and ferbam), ethylenebisdithiocarbamates (e.g., nabam, maneb, zineb, and mancozeb), and propylenebisdithiocarbamates (e.g., propineb) in fruits and vegetables. Limits of quantitation, calculated according to the procedure of the Deutsche Forschungsgemeinschaft, are 9, 12, 8, and 12 microg CS2/L for N-methyl-DTC, N,N-dimethyl-DTC, ethylenebis-DTC, and propylenebis-DTC, respectively, when electrochemical detection is used. PMID- 10868597 TI - Determination of glyphosate residues in plants by precolumn derivatization and coupled-column liquid chromatography with fluorescence detection. AB - A rapid method was developed for the trace-level determination of glyphosate in olives. After extraction of the glyphosate with water-dichloromethane and simultaneous removal of the olive oil, an aliquot of the aqueous extract is derivatized with 9-fluoroenylmethyl chloroformate (9-fluorenylmethoxycarbonyl chloride; FMOC-CI) to produce a highly fluorescent derivative. A 2 mL aliquot of this extract is injected directly into a coupled-column liquid chromatography system with fluorimetric detection (LC/LC-FD). The procedure was validated by recovery experiments at 3 spiking levels; recoveries ranged from 80 to 97% with relative standard deviations of 3-6%. The limits of detection and quantitation were estimated to be 0.01 and 0.05 mg/kg, respectively. The method was also applied to other plant materials, i.e., tomato plants, strawberry plants, and pear trees (branches, leaves, and fruits) suspected to be contaminated by glyphosate. In all these cases, the extraction was performed in aqueous media. The derivatization reaction was modified by increasing the FMOC-CI concentration, to ensure a quantitative reaction between analyte and reagent in the presence of high levels of coextractives, which also react with FMOC-CI. The final determination was by LC/LC-FD, yielding a rapid, selective, and sensitive method for the determination of glyphosate residues in these samples. The method was tested with real-world samples after application of glyphosate to the surrounding area of crops. PMID- 10868598 TI - Determination of residues of pirimicarb and its desmethyl and desmethylformamido metabolites in fruits and vegetables by liquid chromatography-electrospray/mass spectrometry. AB - A method was developed for the simultaneous determination of residues of pirimicarb (I) and its desmethylformamido (II) and desmethyl (III) metabolites in plums, peas, green beans, broad beans, carrots, and swedes. The compounds were extracted with ethyl acetate and determined, without cleanup, by reversed-phase liquid chromatography and electrospray mass spectrometry (MS). MS and MS/MS were used concurrently to monitor the protonated molecules and their common collision induced dissociation product. The limit of detection (signal-to-noise ratio of >3) was 1 ng/mL, corresponding to crop concentrations of <0.0015 mg/kg. All 3 compounds were determined in plums, broad beans, and green beans by MS without interference. Interferences which affected the determination of desmethylformamido-pirimicarb in peas, and to a lesser extent in carrots and swedes, were eliminated by MS/ MS. Recoveries for all 3 compounds, at 0.05 mg/kg for plums and 0.005 mg/kg for other commodities, were in the range 83-124%. No interconversion of I, II and III, occurred during extraction, and the compounds were stable in extracts for > or = 7 days under appropriate conditions. PMID- 10868599 TI - Determination of chlormequat in pears by liquid chromotagraphy/mass spectrometry. AB - A straightforward and reliable method was developed for the determination of chlormequat in pears by liquid chromatography/mass spectrometry (LC/MS). Water and methanol were compared as extraction solvents. Because no significant differences in extraction efficiency or repeatability were found, water was chosen as the extraction solvent. The extracts were analyzed without cleanup by either an ion-trap liquid chromatograph/mass spectrometer in the single MS mode or a triple-quadrupole instrument in the MS/MS mode, using electrospray ionization. Both instruments were equally suitable for quantitation and confirmation of identity. Recoveries were 76-103%, and reproducibility was < or = 12%. The lowest detection limit (0.007 mg/kg) was obtained with the triple quadrupole instrument in the MS/MS mode. PMID- 10868600 TI - Determination of imidacloprid and benzimidazole residues in fruits and vegetables by liquid chromatography-mass spectrometry after ethyl acetate multiresidue extraction. AB - A simple and sensitive method based on liquid chromatography-atmospheric pressure ionization-mass spectrometry is described for the determination of 4 benzimidazole pesticides (carbendazim, thiabendazole, benomyl, and thiophanate methyl) and imidacloprid in vegetables and fruits. Food samples were typically extracted with ethyl acetate to draw the analytes into the organic phase. No cleanup step was necessary before injection into the liquid chromatographic (LC) system with electrospray mass spectrometric detection. The analytes were separated on a reversed-phase C8LC column. Limits of detection for the compounds were in the microg/L range. Results are reported for validation studies with fortified pear and tomato samples and for residues of the target compounds found in the pesticide residue monitoring program during 1998. PMID- 10868601 TI - Use of a particle beam interface combined with mass spectrometry/negative chemical ionization to determine polar herbicide residues in soil by liquid chromatography. AB - The use of mass spectrometry/negative chemical ionization (MS/NCI), in combination with selected-ion monitoring, allows sensitive and selective determination of polar and thermally unstable herbicide residues by liquid chromatography coupled to a particle beam interface. The method has been applied to the analysis of soil samples for herbicide residues, using various procedures for their extraction and employing matrix-standard calibration to avoid quantitative errors due to transfer of matter through the interface. Data for the fragment ions found in the MS/NCI procedure and chromatograms from analyses of real samples are also presented. PMID- 10868602 TI - Rapid target analysis for pesticides in water by online coated capillary microextraction combined with liquid chromatography and tandem mass spectrometry. AB - The applicability of online trace enrichment with custom-made coated capillaries combined with tandem mass spectrometry was demonstrated for the target analysis of selected pesticides (mainly herbicides, e.g., triazines, phenylureas, and acetanilides) in water. The developed method allows rapid determination of several widely used plant protectants within a total analysis time of 11 min. Good linearity (r > or = 0.995) was obtained for the selected pesticides in the range of 0.050-50 microg/L. The relative standard deviations (RSDs) of the peak areas were < or = 3.8% for spiked Milli-Q water (5 microg/L). The RSDs obtained in analyses of spiked (1 microg/L) water samples (brook water, river water, sewage plant effluent) ranged from 2.9 to 6.8%, indicating low influence of the matrix on enrichment and detection. The detection limits, which ranged from 10 to 90 ng/L, fulfilled the requirements of the European regulations for drinking water. The polyacrylate coating of the extraction capillary showed good stability in the presence of water and acetonitrile and allowed > or = 100 extractions with 1 capillary. PMID- 10868603 TI - The thyroid-cholesterol connection: an association between varying degrees of hypothyroidism and hypercholesterolemia in women. PMID- 10868604 TI - The silent epidemic of Chlamydia trachomatis: the urgent need for detection and treatment in women. PMID- 10868605 TI - Toward optimal health: the experts discuss thyroid disease. Interview by Jodi Godfrey Meisler, M.S., R.D.. PMID- 10868606 TI - Women in midlife: a nutritional perspective. PMID- 10868607 TI - Uterine artery embolization: a minimally invasive technique for the treatment of uterine fibroids. AB - Uterine artery embolization (UAE) as a primary therapy for symptomatic fibroids was first used in France in 1991. Currently, there are at least 250 centers in the United States, as well as centers in Canada and England, with experience in this technique. Initial published results worldwide indicate that after UAE, uterine fibroids shrink at least 50% in volume on average and symptoms of refractory vaginal bleeding and chronic pelvic pain are controlled in approximately 85% of patients. Major complications are rare. Overall, this technique is minimally invasive, preserves the uterus, and requires a shorter hospitalization than hysterectomy or myomectomy. PMID- 10868608 TI - Engaging women's interest in colorectal cancer screening: a public health strategy. AB - Screening rates for colorectal cancer are unacceptably low. New guidelines, public education campaigns, and expanded coverage of screening costs by healthcare insurance are expected to increase screening rates, but interventions targeting women may accelerate this change. Most American women already participate in regular cancer screening, in the form of Papanicolaou (Pap) tests and mammography, so they may be receptive to tailored messages about the need to add regular colorectal cancer screening to their preventive health regimen. In addition, their role in promoting the health of family members may position women to influence screening behavior in family and friends. Women may be particularly valuable change agents in populations where screening rates are traditionally low, such as medically underserved populations, the elderly or low socioeconomic status groups with competing health priorities, and populations with cultural values or practices inconsistent with the adoption of a new screening behavior. To serve as agents of change in their family and social networks, women must understand that colorectal cancer is not solely a man's disease and that the benefits of colorectal screening are similar to those of Pap testing and mammography. Colorectal cancer screening should also be promoted within a framework of a lifelong strategy for health maintenance for both men and women. The message to women should emphasize the value of colorectal cancer screening rather than the disagreement among experts over preferred screening strategies and should emphasize the value of shared decision making between the patient and her healthcare provider. PMID- 10868609 TI - Lower extremity arterial disease in older women: the Rancho Bernardo Study. AB - Most studies of lower extremity arterial disease (LEAD) have not included women. To study the frequency of LEAD and its association with cardiovascular disease risk factors and estrogen use in community-dwelling postmenopausal women, we conducted a cross-sectional study of LEAD in 826 women whose average age was 74 years. Cardiovascular disease risk factors and medical history, body mass index (BMI), blood pressure, glucose tolerance, lipids and lipoproteins, and current and past medication use were determined using a standard protocol. Ankle-brachial artery index (ABI) of systolic blood pressure was measured by a trained technician using Doppler ultrasound. LEAD was defined as ABI <0.8. LEAD prevalence increased with age from nearly 5% in the 60-69-year-old group to >25% in women aged 90 and older. In age-adjusted analyses, women with LEAD had significantly lower levels of high-density lipoprotein (HDL) cholesterol, were less likely to exercise regularly, and were less likely to have ever used estrogen replacement therapy. They also had significantly higher levels of blood pressure, low-density lipoprotein (LDL) cholesterol, triglycerides, glucose, and insulin. In multivariate analyses, high HDL cholesterol, regular exercise, and estrogen use were each associated with a reduced risk of LEAD, whereas age, high blood pressure, and abnormal glucose tolerance were each associated with increased risk. Few women (6%) were smokers, but they had twice the risk of LEAD compared with nonsmokers. Estrogen was independently associated with LEAD in a model containing all covariates except LDL and HDL, and this association was no longer significant in a second model adjusting for these lipoproteins. LEAD is common in older women and associated with modifiable risk factors. The apparent protection associated with estrogen should be studied in clinical trials. PMID- 10868610 TI - Comparison of regimens containing oral micronized progesterone or medroxyprogesterone acetate on quality of life in postmenopausal women: a cross sectional survey. AB - A cross-sectional survey was conducted to examine quality of life (QOL) related to physiological, somatic, and vasomotor effects of changing progestogen treatment from medroxyprogesterone acetate (MPA) to micronized progesterone in postmenopausal women. Eligible women (n = 176) were currently using hormone replacement therapy (HRT) containing micronized progesterone for 1-6 months and had previously received HRT containing MPA. QOL was assessed via telephone interview using the Greene Climacteric Scale and the Women's Health Questionnaire. When compared with the MPA-containing regimen, women using micronized progesterone-containing HRT experienced significant improvement in vasomotor symptoms, somatic complaints, and anxiety and depressive symptoms. Women reported improved perceptions of their patterns of vaginal bleeding and control of menopausal symptoms while on the micronized progesterone-containing regimen. Approximately 80% of women reported overall satisfaction with the micronized progesterone-containing regimen. A micronized progesterone-containing HRT regimen offers the potential for improved QOL as measured by improvement of menopause-associated symptoms. PMID- 10868611 TI - Preventive healthcare in women with multiple sclerosis. AB - The health maintenance of women with diverse disabilities and chronic disabling conditions has been neglected by medical professionals. Interest in their basic health promotion has been eclipsed by the narrowed focus on their underlying conditions. We surveyed preventive medical practices of 220 women with multiple sclerosis (MS). The objectives of this study were to evaluate the adequacy of preventive healthcare delivery for women with MS and to explore the adequacy of the detection, prevention, and treatment of osteoporosis in this high-risk population. Our survey revealed that 50% of the women do not get regular medical preventive checkups. Twenty-five percent do not have regular pelvic examinations, and 11% have not had a Pap smear within 3-5 years. In women over 40 years old, 52% do not have yearly mammograms. Risk factors for premature osteoporosis in our sample included impaired mobility (53%), corticosteroid use (82%), and vitamin D deficiency as a result of avoidance of sunlight. Despite these risks, 85% have never had bone density testing, 50% are not taking calcium supplements, and 71% are not taking vitamin D. Among the postmenopausal sample, 81% have never had bone density testing, 50% are not taking calcium supplements, and 70% are not receiving hormone replacement therapy (HRT). Only 1% are taking bone resorption inhibitors. The benefits of preventive healthcare and cancer prevention screening should be stressed to women with MS. Referrals should be facilitated by neurologists for dignified, knowledgeable examinations in fully accessible facilities. Osteoporosis prevention, screening, and treatment protocols must be part of the medical plan for all women with MS. PMID- 10868612 TI - Declining levels of erythrocyte folate during the postpartum period among Hispanic women living on the Texas-Mexico border. AB - Hispanic women have higher parity and shorter interbirth intervals than women of other ethnic groups. Thus, they are more likely to become pregnant relatively soon after giving birth, which may place these women at risk of low or deficient levels of specific nutrients. Folic acid is of particular concern because recent studies suggest that maternal use of folic acid supplements may be associated with better reproductive outcomes. The purpose of this study was to assess folic acid levels in postpartum Hispanic women. Using a cross-sectional design, we measured erythrocyte folate values for 188 low-income Hispanic women 1-12 months postpartum who were receiving services at the Women, Infants, and Children (WIC) clinics in El Paso, Texas. An interview was administered to collect information on diet, vitamin use, and method of infant feeding. Mean erythrocyte folate levels decreased from >1300 ng/ml during the first 4 months postpartum to a low of 1017 ng/ml by 12 months postpartum, for an overall decrease of approximately 23% (p = 0.004). Use of postpartum vitamin supplements was significantly associated with higher folate levels. However, only 35% of mothers used vitamins beyond 1 month postpartum. Study results suggest that these mothers may be at risk of developing low or deficient levels of folic acid during the postpartum period. Educational campaigns targeting these women as well as other groups of postpartum women should encourage them to comply with the U.S. Public Health Service recommendation that women of childbearing age consume 0.4 mg of folic acid daily. PMID- 10868613 TI - Physician beliefs about victims of spouse abuse and about the physician role. AB - Our purpose was to measure the beliefs of physicians about victims of spouse abuse and to examine factors related to holding positive (e.g., supportive) and negative beliefs about providing services to victims of domestic violence. This was a total site sample of 150 physicians (76 responded; RR 51%), surveyed at one time, practicing in a large general hospital and the surrounding urban/periurban area. Four specialities are represented: emergency medicine, family practice, obstetrics-gynecology, and psychiatry. Three aspects of beliefs are measured: beliefs toward physician role in assisting victims of spouse abuse, beliefs about victims of spouse abuse, and beliefs about resources available to physicians to assist victims of spouse abuse. Almost all (97%) physicians believe it is part of their role to assist victims of domestic violence. Almost one third (30%) hold victim-blaming attitudes toward victims of spouse abuse, and the majority (70%) do not believe that they have the resources available to them to assist victims of domestic violence. Being female, younger, practicing obstetrics-gynecology, and having fewer years in practice are all significantly related to holding supportive (positive) beliefs. The majority of negative beliefs held are about resource availability. Hence, training programs may need to be developed locally for physicians and tailored to individual community characteristics. Training programs should also emphasize the importance of understanding the victims of spouse abuse and of not blaming the victims for the violence. PMID- 10868614 TI - Gender-related behavior during childhood and associations with adult abdominal obesity: a nested case-control study in women. AB - Abdominal obesity affects many aspects of women's health, and recent studies indicate that hyperandrogenicity (HA) may contribute to the excess of body fat in women. As hormone behavior research attributes male-like play patterns in childhood to the effects of androgens, the aim of the present study was to assess the potential association of such behavior with obesity in adult women. In a randomly selected sample of 40-year-old women (n = 1464), 78% volunteered to respond to a questionnaire collecting information on the effect of other variables on childhood behavior. Self-reported body weight, height, and waist and hip circumferences were used to calculate body mass index (BMI) and waist/hip ratio (WHR). Age at menarche showed an inverse association with overweight (BMI > or = 25) (odds ratio [OR] = 0.82). Reports of gender-related behavior as a child showed that playing with girls and girl toys was negatively related to both overweight and abdominal obesity (WHR > or = 0.85). Among respondents who were overweight, relationships were found for playing with boys (OR = 0.90) and fighting (OR = 1.70). The OR of playing with boy toys and fighting among respondents with abdominal obesity were increased 1.12 and 1.65, respectively. Interests in athletics as a child seemed to decrease the risk for overweight (OR = 0.89) and abdominal obesity (OR = 0.91). Furthermore, dose-response analysis between the individual exposure levels and the OR for overweight showed a negative trend for playing with girls (p = 0.002) and girl toys (p = 0.017) and a positive trend for playing with boys (p = 0.011) and fighting (p = 0.031). Among respondents with abdominal obesity, positive dose-response effects were found for playing with boys (p = 0.026) and boy toys (p = 0.036) and fighting (p = 0.008). Thus, women with an elevated WHR showed a preference to play with boys and boy toys and also fought frequently as children. This might be a sign of a relative HA in childhood ("tomboyism"). These preliminary observations suggest that HA may originate in childhood. PMID- 10868616 TI - Women's Health LiteratureWatch. PMID- 10868615 TI - Single-dose miconazole nitrate vaginal ovule in the treatment of vulvovaginal candidiasis: two single-blind, controlled studies versus miconazole nitrate 100 mg cream for 7 days. AB - To determine the efficacy and safety of a single-dose (1200 mg) soft gel insert (vaginal ovule) with miconazole nitrate (2%) topical cream compared with Monistat 7 (miconazole nitrate 2%) Vaginal Cream (Advanced Care Products, North Brunswick NJ) in treating vulvovaginal candidiasis (VVC), two randomized, single-blind, multicenter, controlled, comparative phase III studies were performed. Five hundred fifty-eight patients received either a single-dose miconazole nitrate (1200 mg) ovule or seven consecutive doses of Monistat 7. Ovule arm patients also received miconazole nitrate 2% cream for symptom relief, as needed, up to twice daily. The primary end point was a therapeutic cure. Also evaluated were time to complete symptom relief, safety, and patient preference. The ovule had overall cure rates of 71.7% (71 of 99 patients) and 61.5% (64 of 104 patients). Monistat 7 had overall cure rates of 70.1% (68 of 97 patients) and 61.1% (55 of 90 patients). A significantly greater proportion of patients experienced complete symptom relief by day 3 with the ovule (p = 0.008 and p = 0.025), and time to complete relief was significantly faster (median 4 versus 5 days and 3 versus 4 days). Overall safety results were consistent between groups in both studies. Miconazole nitrate vaginal ovule is as safe and efficacious in curing VVC as Monistat 7 while providing complete symptom relief significantly faster. Patients preferred the ovule to prior therapy. PMID- 10868617 TI - Laparoscopic live donor nephrectomy. PMID- 10868618 TI - Role of the thymus in T lymphocyte reconstitution. PMID- 10868619 TI - Is there hope to improve chronic renal allograft outcome? PMID- 10868620 TI - Tuning to the right frequency: cytotoxic T lymphocytes and cytomegalovirus. PMID- 10868621 TI - Characterization of CMVpp65-specific CD8+ T lymphocytes using MHC tetramers in kidney transplant patients and healthy participants. AB - BACKGROUND: Cytomegalovirus (CMV) is a ubiquitous herpesvirus that infects 50-90% of individuals in different populations. After primary infection, the virus persists latently in myeloid cells under the control of specific T-cells. Reactivation of CMV infection may cause lethal organ dysfunction and is frequently seen in immunosuppressed individuals. CD8+ cytotoxic T-cells (CTL) have a primary role in suppressing CMV reactivation, and the dominating CTL response is directed against pp65. METHODS: MHC tetramers, that is, complexes between HLA class I (or class II) molecules and antigenic peptides conjugated to fluorochromes allow the direct visualization of antigen-specific receptor carrying T-cells using flow cytometry. We constructed a novel MHC tetramer for identification of CMVpp65-specific CD8+ T-cells using HLA-A2 molecules folded with the immunodominant NLVPMVATV peptide. RESULTS: The A2/pp65 tetramer specifically stained CMV-directed T-cell lines, and sorted cells showed CMV specific cytotoxicity. High proportions (0.1-9%) of the CD8+ T-cells were A2/pp65 tetramer+ in healthy HLA-A2+ CMV carriers and in immunosuppressed kidney transplant patients with latent infection. Patients with reactivated CMV infection exhibited up to 15% A2/pp65 tetramer+ cells, which seemed to correlate with CMV load over time. A2/pp65 tetramer+ cells expressed T-cell activation markers. CONCLUSIONS: The construction of a novel A2/pp65 MHC tetramer enabled the design of a rapid and precise flow cytometric method allowing quantitative and qualitative analysis of CMV-specific T-cells. The number of A2/pp65 tetramer binding CTLs in blood may prove to be clinically relevant in assessing the immune response to CMV. PMID- 10868622 TI - Direct visualization and quantitation of cytomegalovirus-specific CD8+ cytotoxic T-lymphocytes in liver transplant patients. AB - BACKGROUND: CMV infection remains a significant clinical problem in the context of LT. Changes in the magnitude of the CMV-specific CTL response after LT have not previously been assessed but may be important in determining the outcome of CMV infection. METHOD: We used a fluorescent HLA-B*0702-CMV peptide tetrameric complex to directly visualize and quantitate CMV-specific CD8+ CTL both in immunosuppressed patients after LT and in immunocompetent controls. RESULTS: CMV specific CD8+ CTL, at a frequency ranging from 0.1 to 5.8% of CD8+, were detected in the peripheral blood of 22 of 25 B*0702, CMV immunoglobulin G seropositive individuals, with no difference observed between immunocompetent controls and patients >3 years after LT. In CMV seropositive LT recipients who did not have symptomatic CMV infection during the first 3 months after LT, CMV-specific CD8+ CTL magnitude initially decreased, then increased up to 5 times higher than pre LT levels within 3 months. Two CMV seronegative recipients of seropositive donors had symptomatic CMV infection in association with high viral load. In both patients, no CD8+ CTL response was detected before the onset of symptoms, and a reduction in viral load was observed during antiviral therapy. However, polymerase chain reaction negativity was achieved only when a demonstrable CMV specific CD8+ CTL response was generated. Responses were never observed in asymptomatic CMV seronegative patients. CONCLUSIONS: We suggest that the generation of CMV-specific CD8+ CTL may be driven by, and seems to coincide with the suppression of, viral reactivation. Direct monitoring of CMV-specific CD8+ CTL using an HLA-peptide tetramer may prove to be of value in the management of patients after LT. PMID- 10868623 TI - Immunosuppressive therapies for the prevention and treatment of obliterative airway disease in heterotopic rat trachea allografts. AB - BACKGROUND: Obliterative bronchiolitis remains a major long-term complication after lung transplantation. Using a reproducible model of heterotopically transplanted rat tracheas, this study examined the role of several novel immunosuppresive compounds to prevent and reverse obliterative airway disease in these animals. METHODS: Brown Norway rat trachea were transplanted into the greater omentum of Lewis (allografts) or Brown Norway (isografts) animals. Recipient animals were treated with rapamycin, cyclosporine, 15-deoxyspergulin, mycophenolate mofetil, or leflunomide from day 0, 7, or 14 until day of graft removal, either day 28 or 50. Trachea segments were evaluated for degree of lumenal occlusion, as well as percent and type of lumen epithelial cell coverage. RESULTS: All untreated allografted tracheas obliterated completely, although isografts appeared patent with normal respiratory epithelium when they were removed. Leflunomide, rapamycin, and cyclosporine effectively prevented obliteration when treatment was initiated at day 0, with rapamycin showing continued efficacy when initiated as late as day 7. 15-deoxyspergulin and mycophenolate mofetil failed to consistently inhibit obliteration with any treatment schedule. An inverse correlation was found between epithelial coverage and degree of obliteration, and was especially pronounced in grafts from cyclosporine-treated animals. CONCLUSIONS: Immunosuppressive drug therapy will inhibit airway obliteration, but efficacy sharply diminishes if initiation of treatment is delayed. Efficacy also varies among immunosuppressive compounds, and results indicate those drugs that enable epithelial regrowth most effectively inhibit airway graft obliteration. PMID- 10868624 TI - The roles of platelet-activating factor and endothelin-1 in renal damage after total hepatic ischemia and reperfusion. AB - BACKGROUND: This study was designed to verify the involvement of platelet activating factor (PAF) in renal damage associated with hepatic ischemia and reperfusion (HIR) injury through the release of endothelin (ET)-1 and to determine the modulating effect of a specific PAF receptor antagonist on these insults in rats. METHODS: Male rats pretreated with either normal saline as a vehicle (NS group) or intravenous TCV-309, a PAF receptor antagonist (TCV group), were subjected to 120 min of total hepatic ischemia under an extracorporeal portosystemic shunt. Plasma aspartate transaminase, creatinine, blood urea nitrogen, and ET-1 levels and the relative renal wet weight were determined under nonischemic conditions and at 1, 3, and 6 hr of reperfusion after hepatic ischemia. Changes in mean arterial blood pressure and renal tissue blood flow measurements in the kidney were determined throughout the experiment. RESULTS: Increased plasma aspartate transaminase, creatinine, blood urea nitrogen, and ET 1 levels and the relative renal wet weight after HIR in the NS group were significantly suppressed by TCV-309 pretreatment. Mean arterial blood pressure and renal tissue blood flow after HIR in the TCV group were significantly improved when compared with those in the NS group. These effects resulted in attenuation of structural hepatic and renal damage with the improvement of 7-day survival (62%). CONCLUSIONS: The present study demonstrates that renal damage as well as critical liver injury is produced after reperfusion following 120 min of total hepatic ischemia. A PAF receptor antagonist may be therapeutically useful to protect against these types of damage via indirect modulation of plasma ET-1 levels. PMID- 10868625 TI - Stress responses in graft and native intestine after rat heterotopic small bowel transplantation. AB - BACKGROUND: Cardiac transplantation has been shown to induce heat shock protein expression, and reactivity to these stress proteins has been implicated in acute and chronic allograft rejection. This study assessed Hsp60 and Hsp70 expression in graft and native small intestine after rat small bowel transplantation. METHODS: Heterotopic small bowel transplantation was performed between PVG donor and DA recipient rats, a subgroup of which received tacrolimus immunosuppression (1 mg x kg(-1) x day(-1)). Untransplanted and isografted (PVG-->PVG) animals served as controls. Paraffin sections of graft and native intestine on day 5 after transplantation were stained by immunohistochemistry, and heat shock protein expression was graded blindly by three observers. RESULTS: Villus epithelial cell expression of Hsp60, but not Hsp70, was increased in allografts. The induction of Hsp60 in the villus epithelium was not controlled by tacrolimus. Hsp60 and Hsp70 expression was induced in the lamina propria of isografts and allografts. This response was more pronounced in allografts and was significantly reduced, but not totally abrogated, by tacrolimus. Interestingly, heat shock protein expression was also induced in the native intestine lamina propria and epithelium of allograft recipients, suggesting the induction of stress responses at sites other than the transplanted organ. CONCLUSIONS: Small bowel transplantation induces a stress response in both the graft and native intestine. The early and prolonged expression of these proteins may influence the induction of anti-heat shock protein reactivity and have an adverse effect on graft outcome after small bowel transplantation. PMID- 10868626 TI - Adenosine deaminase inhibition attenuates reperfusion low flow and improves graft survival after rat liver transplantation. AB - BACKGROUND: Low flow or no flow is a prefinal step after reperfusion of hepatic allografts. Adenosine is an intrinsic key regulator of physiological and pathological hepatic blood flow. METHODS: In a model of rat liver transplantation, the effect of donor pretreatment with adenosine deaminase inhibitors (0, 0.1, 1, 10 micromol erythro-9-[2-hydroxy-3-nonyl]adenine) was studied on hepatic interstitial adenosine concentrations, microcirculatory flow, leukocyte adhesion, and graft survival by means of microdialysis sampling, intravital video microscopy, and laser Doppler flowmetry. RESULTS: Donor pretreatment with 1 micromol erythro-9-[2-hydroxy-3-nonyl]adenine increased interstitial adenosine concentrations 5- to 10-fold, for more than 24 hr of cold storage. In LDF studies, mean donor blood flow was increased from 420 +/- 42 perfusion units (PU) to 832 +/- 52 PU and from 475 +/- 79 to 720 +/- 81 PU after reperfusion, and in intravital video microscopy studies from 247 +/- 24 to 281 +/ 39 pl/sec. There was no difference in the number of leukocytes sticking, but a significantly lower percentage of leukocytes rolling (26.1 +/- 1.9 vs. 36.5 +/- 7.5%) along the endothelial wall in the treatment group. Transplant survival after 44 hr cold storage in UW solution was 8/10 in the treatment group and 1/13 in the control group. CONCLUSIONS: Donor pretreatment with erythro-9-[2-hydroxy-3 nonyl]adenine increases survival of critically injured liver grafts. Donor or recipient treatment rather than addition of protectants to cold storage solutions are successful strategies to overcome preservation injury and possibly adverse donor factors. PMID- 10868627 TI - A recombinant soluble chimeric complement inhibitor composed of human CD46 and CD55 reduces acute cardiac tissue injury in models of pig-to-human heart transplantation. AB - BACKGROUND: Inasmuch as complement plays a critical role in many pathological processes and in xenograft rejection, efficient complement inhibitors are of great interest. Because the membrane-associated complement inhibitors are very effective, recombinant soluble molecules have been generated. METHODS: We tested the efficacy of complement activation blocker-2 (CAB-2), a recombinant soluble chimeric protein derived from human decay accelerating factor (DAF, CD55) and membrane cofactor protein (MCP, CD46), in two models of pig-to-human xenotransplantation in which tissue injury is complement mediated. The in vitro model consisted of porcine aortic endothelial cells and human serum, and the ex vivo model consisted of a porcine heart perfused with human blood. RESULTS: In vitro, addition of CAB-2 to serum inhibited cytotoxicity and the deposition of C4b and iC3b on the endothelial cells. Ex vivo, addition of CAB-2 to human blood prolonged organ survival from 17.3 +/- 6.4 min in controls to 108 +/- 55.6 min with 910 nM (100 microg/ml) CAB-2 and 219.8 +/- 62.7 min with 1820 nM (200 microg/ml) CAB-2. CAB-2 also retarded the onset of increased coronary vascular resistance. The complement activity of the perfusate was reduced by CAB-2, as was the generation of C3a and SC5b-9. The myocardial tissues had similar deposition of IgG, IgM, and Clq; however, CAB-2 reduced the deposition of C3, C4, and C9. Hearts surviving >240 min demonstrated trace to no deposition of C9 and normal histologic architecture. CONCLUSION: These results indicate that CAB-2 can function as an inhibitor of complement activation and markedly reduce tissue injury in models of pig-to-human xenotransplantation and thus may represent a useful therapeutic agent for xenotransplantation and other complement-mediated conditions. PMID- 10868628 TI - Taurine attenuates cold ischemia-reoxygenation injury in rat liver. AB - BACKGROUND: Taurine, betaine, and inositol were recently identified as osmolytes in liver cells interfering with cell volume regulation and cell function. In this study, the effect of osmolytes on cold ischemia-reoxygenation injury was investigated in rat liver. METHODS AND RESULTS: Isolated rat livers were flushed for 15 min with Krebs-Henseleit buffer (KHB), then stored for 16 hr in KHB at 4 degrees C, and thereafter reperfused with oxygenated KHB for 180 min. When taurine, betaine, and inositol (2 mmol/L, each) were added to the preperfusion and storage buffer, lactate dehydrogenase, aspartate amino transferase, and glutathione S-transferase leakage into the effluent perfusate during the reoxygenation period were less than half compared to controls without osmolytes and bile flow was higher. The effect of taurine (2 mmol/L) was similar to a mixture of all three osmolytes, indicating that taurine is the most important constituent. When livers were stored for 24 hr in University of Wisconsin solution, osmolyte addition to the storage solution also decreased lactate dehydrogenase and aspartate aminotransferase leakage during reoxygenation. Increasing liver taurine content by a 7-day taurine supplementation of drinking water attenuated reoxygenation injury in cold and warm ischemia in rat livers, whereas taurine depletion by beta-alanine feeding had the opposite effect. CONCLUSIONS: The data show that taurine protects livers from ischemia reoxygenation. Taurine addition to perfusion and storage solutions in low millimolar concentrations or taurine supplementation of the donor may be useful to protect transplanted organs. PMID- 10868629 TI - High-dose porcine hematopoietic cell transplantation combined with CD40 ligand blockade in baboons prevents an induced anti-pig humoral response. AB - BACKGROUND: In pig-to-primate organ transplantation, hyperacute rejection can be prevented, but the organ is rejected within days by acute vascular rejection, in which induced high-affinity anti-Gal alpha1-3Gal (alphaGal) IgG and possibly antibodies directed against new porcine (non-alphaGal) antigenic determinants are considered to play a major role. We have explored the role of an anti-CD40L monoclonal antibody in modifying the humoral response to porcine hematopoietic cells in baboons pretreated with a nonmyeloablative regimen. METHODS: Porcine peripheral blood mobilized progenitor cells obtained by leukapheresis from both major histocompatibility complex-inbred miniature swine (n=7) and human decay accelerating factor pigs (n=3) were transplanted into baboons. Group 1 baboons (n=3) underwent whole body (300 cGy) and thymic (700 cGy) irradiation, T cell depletion with ATG, complement depletion with cobra venom factor, short courses of cyclosporine, mycophenolate mofetil, porcine hematopoietic growth factors, and anti-alphaGal antibody depletion by immunoadsorption before transplantation of high doses (2-4 x 10(10)/cells/kg) of peripheral blood mobilized progenitor cells. In group 2 (n=5), cyclosporine was replaced by eight doses of anti-CD40L monoclonal antibodies over 14 days. The group 3 baboons (n=2) received the group 1 regimen plus 2 doses of anti-CD40L monoclonal antibodies (on days 0 and 2). RESULTS: In group 1, sensitization to alphaGal (with increases in IgM and IgG of 3- to 6-fold and 100-fold, respectively) and the development of antibodies to new non-alphaGal porcine antigens occurred within 20 days. In group 2, no sensitization to alphaGal or non-alphaGal determinants was seen, but alphaGal reactive antibodies did return to their pre- peripheral blood mobilized progenitor cells transplant levels. In group 3, attenuated sensitization to alphaGal antigens was seen after cessation of cyclosporine and mycophenolate mofetil therapy at 30 days (IgM 4-fold, IgG 8-30-fold), but no antibodies developed against new porcine determinants. In no baboon did anti-CD40L monoclonal antibodies prevent sensitization to its own murine antigens. CONCLUSIONS: We believe these studies are the first to consistently demonstrate prevention of a secondary humoral response after cell or organ transplantation in a pig-to-primate model. The development of sensitization to the murine elements of the anti-CD40L monoclonal antibodies suggests that nonresponsiveness to cell membrane-bound antigen (e.g., alphaGal) is a specific phenomenon and not a general manifestation of immunological unresponsiveness. T cell costimulatory blockade may facilitate induction of mixed hematopoietic chimerism and, consequently, of tolerance to pig organs and tissues. PMID- 10868630 TI - A 12-day course of FK506 allows long-term acceptance of semi-identical liver allograft in inbred miniature swine. AB - BACKGROUND: Spontaneous tolerance to liver allograft has been reported previously in outbred pig models, but the lack of genetic background did not allow to analyze the impact of the major histocompatibility complex (MHC) on tolerance induction. A model of semi-identical liver allograft was therefore developed in inbred miniature swine in order to mimic the clinical situation of living related liver transplant (parent into infant) and to study a protocol for inducing tolerance to liver allograft. METHODS: SLAdd (class Id/d, class IId/d) pigs received orthotopic liver allograft from heterozygous SLAcd (class Ic/d, class IIc/d) miniature swine. Eight animals did not receive immunosuppression. Fourteen SLAdd animals had a 12-day course of FK506 and were divided in two subgroups. In subgroup FK-1, six pigs received a daily intramuscular injection of FK506 at 0.1 0.4 mg/kg, in order to reach daily trough levels between 7 and 20 ng/ml; in subgroup FK-2, eight additional animals received two daily injections of FK506 at 0.05 mg/kg regardless of the daily trough levels. Graft survival, liver biological tests, histology, cellular and humoral immune responses, as well as detection of microchimerism were assessed in all groups. RESULTS: All untreated animals rejected their allograft and died within 28.1 +/- 9.5 days. These rejector animals developed a significant anti-donor cellular and humoral immune response. No peripheral or lymphoid tissue microchimerism was detected in this group. In contrast, long-term survival was obtained in five FK-treated animals (112, 154, 406, 413, and 440 days), whereas several pigs died with a normal allograft function from either overimmunosuppression or intercurrent causes. All FK-treated pigs developed a specific anti-donor unresponsiveness in both cell mediated lymphocytotoxicity and mixed lymphocyte reaction and did not develop anti-donor alloantibodies. The study of the anti-donor immune response by mixed lymphocyte reaction, during the first postoperative week, demonstrated a specific anti-donor unresponsiveness in the peripheral blood from the first posttransplant day. Although microchimerism was detectable in the peripheral blood for several postoperative weeks (maximum 10 weeks) in FK-treated animals, donor cells or DNA were not detected during the long-term follow-up in peripheral blood or lymphoid tissues. CONCLUSIONS: Spontaneous tolerance to semi-identical orthotopic liver allograft did not occur, whereas a 12-day course of FK506 allowed long-term graft acceptance. All FK-treated animals developed in vitro signs of specific immune unresponsiveness and transient peripheral microchimerism. The specific anti-donor cellular unresponsiveness occurred on the first postoperative day after surgery and was of long-term duration. The study of the early immunological events in this model could be of major importance regarding clinical living related liver transplantation. PMID- 10868631 TI - The inhibitory effect of prostaglandin E1 on oxidative stress-induced hepatocyte injury evaluated by calpain-mu activation. AB - BACKGROUND: Prostaglandin E1 (PGE1) is known to inhibit ischemia-reperfusion injury of the liver. The calcium-dependent neutral proteinase, calpain-mu, is involved in oxidative stress-induced hepatocyte injury. We investigated the mechanisms of cytoprotection by PGE1, focusing on the elevation of intracellular calcium ([Ca2+]i), activation of calpain-mu, and calpain-mu-mediated activation of protein kinase C-alpha (PKC-alpha). METHODS: Cultured hepatocytes were treated with various amounts of PGE1 (0, 0.1, 1.0, 10, and 100 ng/ml) for 30 min and subsequently with 0.5 mM tert-butyl hydroperoxide (TBHP). Cell injury was evaluated by the release of lactate dehydrogenase. Plasma membrane bleb formation was examined by phase contrast microscopy. Activation of calpain-mu and limited degradation of PKC-alpha was evaluated by Western blotting using antibodies that specifically recognize the amino-terminal regions of calpain-mu and PKC-alpha. [Ca2+]i was measured by confocal microscopy using Fluo-3AM. RESULTS: LDH release from cells treated with 10 ng/ml PGE1 was significantly lower than from untreated cells (135 +/- 12 vs. 258 +/- 18 IU/L, respectively; P < 0.05). Morphologically, many blebs were observed in untreated cells, but very few were seen in those treated with 10 ng/ml PGE1. Western blotting revealed that the amount of activated calpain-mu and [Ca2+]i increased up to 1,300 nM at 35 min after the addition of TBHP (0.5 mmol/L) in control experiments (without PGE1). PGE1 (10 ng/ml) delayed the rise in [Ca2+]i for about 30 min, but did not suppress it completely. PKC-alpha decreased in experiments using PGE1 (10 ng/ml). CONCLUSION: PGE1 exerts its cytoprotective effect in TBHP-induced hepatocyte injury partly by inhibiting Ca2+-calpain-mu-mediated mechanisms. PMID- 10868632 TI - Laparoscopic live donor nephrectomy: the recipient. AB - BACKGROUND: Laparoscopic live donor nephrectomy offers advantages to the donor in terms of decreased pain and shorter recuperation. Heretofore no detailed analysis of the recipient of laparoscopically procured kidneys has been performed. The purpose of this study was to determine whether laparoscopic donor nephrectomy had any deleterious effect on the recipient. METHODS: A retrospective review was conducted of all live donor renal transplantations performed from January 1995 through April 1998. The control group received kidneys procured via a standard flank approach (Open). Rejection was diagnosed histologically. Creatinine clearance was calculated using the Cockroft-Gault formula. RESULTS: A total of 110 patients received kidneys from laparoscopic (Lap) and 48 from open donors. One-year recipient (100% vs. 97.0%) and graft (93.5% vs. 91.1%) survival rates were similar for the Open and Lap groups, respectively. A similar incidence of vascular thrombosis (3.4% vs. 2.1%, P=NS) and ureteral complications (9.1% vs. 6.3%, P=NS) were seen in the Lap and Open groups, respectively. The incidence of acute rejection for the first month was 30.1% for the Lap group and 31.9% for the Open group (P=NS). The rate of decline of serum creatinine level in the early posttransplantation period was initially greater in the Open group, but by postoperative day 4 no significant difference existed. No difference was observed in allograft function long-term. The median length of hospital stay was 7.0 days for both groups. CONCLUSIONS: Laparoscopic live donor nephrectomy does not adversely effect recipient outcome. The previously demonstrated benefits to the donor, and the increased willingness of individuals to undergo live kidney donation, coupled with the acceptable outcomes experienced by recipients of laparoscopically procured kidneys justifies the continued development and adoption of this operation. PMID- 10868633 TI - Isolated liver transplant and sequential small bowel transplantation for intestinal failure and related liver disease in children. AB - Liver dysfunction is a well-recognized complication of intestinal failure in children. Advances in total parenteral nutrition (TPN) have allowed these children to survive while their intestinal tract gradually adapts. Unfortunately TPN may lead to cholestatic liver disease particularly in the young children. Progression of liver disease is associated with a poor prognosis and is an indication for small bowel transplantation. We report our experience of orthotopic liver transplantation in four children with short gut and sequential liver and small bowel transplantation in one child. All children had TPN-related liver failure. Causes of intestinal failure included necrotising enterocolitis (n=2), gastroschisis (n=1), intestinal atresia (n=1), and megacystic, microcolon syndrome (n=1). At the time of liver transplantation the children's mean age was 10.9 months (2.5-24) and weight 6.7 kg (4.8-10.1). The mean serum bilirubin was 522 micromol/liter (299-823), aspartate transaminase 423 IU/liter (49-1024) and international normalized ratio 2.8 (2-3.9). There were two deaths both from respiratory failure secondary to adenovirus pneumonia including the child who received a sequential small bowel transplant. Three children with isolated liver grafts are alive and off TPN at 20 months (mean) follow up (range 6-35). Isolated orthotopic liver transplantation has a role in selected children with intestinal failure, particularly those with short but normally functioning gut and progressing with satisfactory intestinal adaptation but developing liver disease. Those children with TPN-related liver disease and unadapted gut or irreversible intestinal disease need combined liver and small bowel transplantation. Sequential small bowel transplantation is feasible after orthotopic liver transplantation and may provide an option for the child with terminal liver and small bowel failure. PMID- 10868634 TI - The impact of routine mycophenolate mofetil drug monitoring on the treatment of cardiac allograft rejection. AB - INTRODUCTION: Mycophenolate mofetil (MMF) is a unique immunosupressive agent that has been shown to be efficacious in the treatment of cardiac allograft rejection. The utility of therapeutic drug monitoring on rejection prophylaxis and treatment is inconclusive. This study was undertaken to evaluate the incidence of rejection in relation to MMF trough level following heart transplantation. METHODS: Between May 1998 and February 1999, we retrospectively analyzed the clinical outcome of 215 heart transplant patients who had routine monitoring of MMF trough level at the time of scheduled endomyocardial biopsy. Patients were divided into three groups according to the time interval post transplant, and were evaluated in relation to the MMF trough level. Group I, 104 patients within 6 months of transplant; Group II, 90 patients, 6-12 months post transplant; and Group III, 71 patients beyond one year of transplant. Fifty patients had samples in more than one group. Rejection was defined as Grade > or = 3A based on ISHLT criteria. Mean follow-up period was 179+/-52 days. RESULTS: A significantly decreased incidence of rejection was noted in the samples with MMF trough level > or = mg/l compared to those with less than 2 mg/l inpatients evaluated within the first year of transplant (Group I: 8.8% vs. 14.9%, Group II: 4.2% vs. 11.3%, both P=0.05). In the presence of therapeutic cyclosporine (CSA) or tacrolimus (FK) blood levels, the incidence of rejection decreased significantly when MMF trough level was > or = 2 mg/l compared to samples with MMF trough level <2 mg/l (3.6% vs. 14.4%, P=0.005). No significant difference was noted in the presence of subtherapeutic CSA or FK levels (15.4% vs. 13.9%, P=NS). CONCLUSIONS: Monitoring of MMF trough levels may play a role in the management of cardiac transplant recipients during the first year post transplant. PMID- 10868636 TI - Excellent outcome of renal transplantation in patients with Fabry's disease. AB - INTRODUCTION: Fabry's disease is an X-linked error of glycosphingolipid metabolism. Clinical manifestations of the disease are secondary to accumulation of glycosphingolipids in various tissues. Renal failure and vascular complications are common. There are conflicting reports regarding the outcomes of patients with Fabry's disease after renal transplantation. METHODS: We reviewed the United States Renal Data System Registry database from 1988 and 1998, and found 93 patients with Fabry's disease who had received a renal transplant. Case matched patients were identified to serve as controls. RESULTS: Patients with Fabry's disease demonstrated equivalent 5-year patient and graft survival, compared with controls (83% and 75%, respectively, for those with Fabry's disease vs. 82% and 67% for controls). CONCLUSION: Despite their high risk for cardiovascular complications, patients with Fabry's disease have excellent outcomes after renal transplantation. PMID- 10868635 TI - Chronic liver allograft rejection in a population treated primarily with tacrolimus as baseline immunosuppression: long-term follow-up and evaluation of features for histopathological staging. AB - BACKGROUND: Predisposing factors, long-term occurrence, and histopathological changes associated with recovery or progression to allograft failure from chronic rejection (CR) were studied in adult patients treated primarily with tacrolimus. METHODS: CR cases were identified using stringent criteria applied to a retrospective review of computerized clinicopathological data and slides. RESULTS: After 1973 days median follow-up, 35 (3.3%) of 1049 primary liver allograft recipients first developed CR between 16 and 2532 (median 242) days. The most significant risk factors for CR were the number (P<0.001) and histological severity (P<0.005) of acute rejection episodes and donor age >40 years (P<0.03). Other demographic and matching parameters were not associated with CR in this cohort. Ten patients died with, but not of, CR. Eight required retransplantation because of CR at a median of 268 days. Ten resolved either histologically or by normalization of liver injury tests over a median of 548 days. CR persisted for 340 to 2116 days in the remaining seven patients. More extensive bile duct loss (P<0.01), smallarterial loss (P<0.03), foam cell clusters (P<0.01) and higher total bilirubin (P<0.02) and aspartate aminotransferase (P<0.03) were associated with allograft failure from CR. CONCLUSIONS: Early chronic liver allograft rejection is potentially reversible and a combination of histological, clinical, and laboratory data can be used to stage CR. Unique immunological and regenerative properties of liver allografts, which lead to a low incidence and reversibility of early CR, can provide insights into transplantation biology. PMID- 10868637 TI - Manganese deposition in the globus pallidus in patients with biliary atresia. AB - BACKGROUND: Chronic liver diseases may alter trace element contents in the brain. Among these trace elements, manganese is a ubiquitous transition metal excreted by the liver into the bile. Blood concentrations of manganese are elevated in patients with biliary atresia who have undergone hepatic portoenterostomy. The present study investigated the effects of liver transplantation on manganese deposition in the brain in such patients. METHODS: The signal intensity of the globus pallidus was calculated as an index defined as the percentile ratio of signal intensity in the globus pallidus to the subcortical frontal white-matter in sagittal T1-weighted magnetic resonance imaging planes. RESULTS: Brain magnetic resonance imaging revealed hyperintense signals in the globus pallidus due to manganese deposition in biliary atresia patients. Few neurologic symptoms related to manganese intoxication were observed. However, one 23-year-old female with biliary atresia had depressive symptoms and dyskinesia; she improved after oral administration of the dopamine precursor, L-DOPA. Manganese deposition disappeared in two patients after living-related reduced-size hepatic transplantation. CONCLUSIONS: Manganese accumulates in the brain during cholestasis associated with biliary atresia and disappears after hepatic transplantation. Manganese deposition is likely to be subclinical and reversible but may be associated with some age-related neurologic symptoms. PMID- 10868638 TI - Quadruple tacrolimus-based induction therapy including azathioprine and ALG does not significantly improve outcome after liver transplantation when compared with standard induction with tacrolimus and steroids: results of a prospective, randomized trial. AB - BACKGROUND: Tacrolimus in combination with prednisolone has been proven to be a safe and effective immunosuppressive induction therapy in solid organ transplantation. However, it remains unclear whether a tacrolimus-based quadruple induction regimen with azathioprine and an antilymphocytic preparation could further improve the results after orthotopic liver transplantation. Therefore, we designed a prospective, randomized study to compare the immunosuppressive efficacy of dual (tacrolimus and prednisolone) and quadruple (tacrolimus, azathioprine, ALG Merieux and prednisolone) induction after liver transplantation. METHODS: After randomization, 120 consecutive patients of primary liver transplants were divided into the dual group (n=59) and the quadruple group (n=61) and followed for a minimum of 3 years. RESULTS: Patient survival at 3 years was 88.2% in the dual versus 94.9% in the quadruple group. Overall 25 patients in each group (41 and 42%, respectively) developed acute rejection. There was no difference in the number and severity of rejections. In each group only four patients required OKT3-therapy, however, although three of four patients in the quadruple group responded to OKT3 and cleared rejection, none of the four patients in the dual group were treated successfully with OKT3 (P<0.02). Rejection in these patients resolved only after additional treatment with mycophenolate mofetil. Adverse events and infections were equally distributed in both groups. Asymptomatic Cytomegalovirus infections were more common in the quadruple group (P<0.02). As of today, only one patient developed posttransplant lymphoproliferative disease (dual group). CONCLUSIONS: The data from our single-center study indicate that both tacrolimus-based dual and quadruple immunosuppressive induction regimens yield similar safety and effectiveness after liver transplantation. PMID- 10868639 TI - Dobutamine stress echocardiography in patients undergoing liver transplantation evaluation. AB - BACKGROUND: Coronary artery disease has an important impact on perioperative morbidity and mortality in patients undergoing liver transplantation. To assess the role of dobutamine stress echocardiography (DSE) in these patients, DSE was included in the preoperative evaluation. METHODS: Patients under consideration for liver transplantation underwent detailed clinical history, electrocardiography, and echocardiography. Patients with documented coronary disease or symptoms of myocardial ischemia underwent angiography. The remaining patients with cardiac risk factors, atypical chest pain, or age > or = 60 years underwent DSE. RESULTS: These 121 patients (77 men and 44 women) ranged in age from 34 to 73 years (mean 53). Among the 61 patients who underwent liver transplantation, DSE was normal in 25, nondiagnostic in 34 because of inadequate heart rate response, and abnormal in two patients. Major perioperative events occurred in eight patients, all with normal or nondiagnostic DSE studies (negative predictive value 86%). CONCLUSIONS: In patients with low to moderate risk of cardiac disease, DSE performed as part of an evaluation for liver transplantation is a poor predictor of major perioperative events. PMID- 10868640 TI - Clinical determinants of multiple acute rejection episodes in kidney transplant recipients. AB - BACKGROUND: Recipients with multiple (more than one) acute rejection (AR) episodes have significantly lower graft survival rates than those with no AR or only one treated episode. However, fewer than 50% of recipients treated for one AR episode will have another episode. METHODS: We studied recipients with at least one AR episode to determine whether any clinical features could identify risk factors for multiple AR. RESULTS: Between January 1, 1984, and June 30, 1997, a total of 1793 recipients underwent a kidney transplant at our institution. Of these, 354 were treated for one AR episode, 307 for more than one. By multivariate analysis, recipients at highest risk for multiple AR episodes were those with initial delayed or slow graft function (relative risk=1.5, P=0.05), those with initially severe AR (as judged by vascular involvement or steroid resistance), and those with an initial early AR episode (<6 months posttransplant). The remaining variables tested were not significant. Graft survival in recipients with more than one AR episode was significantly lower than in those with only one AR episode. Graft survival at 5 years posttransplant was 52.5% in recipients with more than one AR episode and 85.1% in recipients with one AR episode (P=0.0001). Chronic rejection as a cause of graft loss was significantly more common in recipients with more than one vs. only one AR episode (34.8% vs. 8.9%, P=0.001). CONCLUSION: Clinical features may be used to identify recipients at higher risk for multiple AR episodes. These recipients can then be targeted with more aggressive or novel immunosuppressive regimens in an attempt to reduce the likelihood of another AR episode. PMID- 10868641 TI - Significance of blood stream infection after lung transplantation: analysis in 176 consecutive patients. AB - BACKGROUND: Although infection is a leading cause of death after lung transplantation, very little is known about the incidence, epidemiology, and clinical significance of bloodstream infections in lung transplant recipients. METHODS: All blood cultures were reviewed in 176 consecutive lung transplant recipients over a 6-year period. Data were obtained from a prospectively collected microbiological database. RESULTS: Bloodstream infection (BSI) occurred in 25% (44/176) of all lung transplant recipients over the 6-year study period. Staphylococcus aureus, Pseudomonas aeruginosa, and Candida species were the most common bloodstream isolates after lung transplantation. The epidemiology of posttransplant BSI, however, varied considerably between early and late posttransplant time periods and also differed between cystic fibrosis (CF) and non-CF patients. BSI infection after transplantation was associated with significantly worse survival by Kaplan-Meir analysis (P value log rank test=0.0001). In a multivariable logistic regression model, posttransplant BSI was a significant predictor of posttransplant death (odds ratio 5.62, CI 2.41 13.11, P=0.001), independent of other pre- and posttransplant factors. CONCLUSIONS: Bloodstream infection represents a serious complication after lung transplantation, occurring more frequently than previously recognized, and independently contributing to posttransplant mortality. PMID- 10868642 TI - Requirement of residual thymus to restore normal T-cell subsets after human allogeneic bone marrow transplantation. AB - BACKGROUND: To determine the effect of residual thymic activity in reconstituting the T-cell system after T cell-depleting therapy, we monitored T-cell subsets of a unique thymectomized cancer patient in comparison to thymus-bearing patients after allogeneic bone marrow transplantation (BMT). METHODS: T cells and T-cell subsets previously shown in murine studies to be regulated by the thymus were analyzed by FACS from 6 to >48 months after BMT. The investigation of thymus bearing patients included 32 examinations of 9 children and 14 adults. None of the investigated cases had severe graft-versus-host disease or severe infections when examined. RESULTS: In the thymectomized host, T-cell regeneration occurred by donor cell expansion and was characterized by two prominent features: (i) a persistent failure to regenerate naive (CD45RA+) T-helper cells (14%, median), consistent with the recently developed concept of a thymus-dependency; and (ii) persistently elevated proportions of CD3+CD4-CD8- cells (double-negative cells, median 29%), which were identified in T cell receptor (TCR)gamma delta+ (22%, median of CD3+ cells, 88% double negatives) but also TCRalpha beta+ T-cell populations (78%, median of CD3+ cells, 17% double negatives). In thymus-bearing patients, 10 of 12 and 6 of 14 examinations of children and adults, respectively, performed later than 12 months after BMT showed the proportion of CD4+CD45RA+ cells appropriate for age (>52% and >28% in children and adults, respectively). Elevated double-negative cells (>10%) were found in only three patients, but none had elevated double-negative cells with a TCRalpha beta+ phenotype. CONCLUSION: Residual thymic activity might, in addition to its well-established role for regenerating naive T-helper (CD4+CD45RA+) cells, control the expansion of double negative cells. A normal T-cell subset regeneration in a proportion of thymus bearing adult hosts indicates the potential of an effective residual thymic activity even beyond childhood. PMID- 10868643 TI - A randomized prospective trial of low-dose OKT3 induction therapy to prevent rejection and minimize side effects in recipients of kidney transplants. AB - BACKGROUND: We attempted to minimize the undesired side effects and maximize the benefit of OKT3 induction therapy in renal transplantation. METHODS: One hundred and one recipients of kidney-only transplants were randomized to three groups. Each received low-dose 2.5-mg OKT3 induction for 7-14 days, but different premedication on days 0, 1, and 2. Group I was given 250 mg i.v. methylprednisolone at 1 and 6 hr, and group II received another 500 mg at 1 hr before initial OKT3. Group III received Atgam 15 mg/kg on day 0 and began OKT3 on day 1. A CD3+ T-cell cut-off of 50/mm3 was used to guide therapy. Maintenance therapy included cyclosporine and steroids for each patient. However, groups I and II were also given mycophenolate mofetil, and group III received azathioprine as a third agent. All rejections were biopsy confirmed and Banff scored. RESULTS: No differences in demographic or transplant characteristics were noted between groups I, II, and III, and mean follow-up was 25.7 (1-38) months. There was no significant difference in actuarial patient (90%, 91%, 94%) or graft survival (83%, 88%, 84%) at 3 years between the respective groups. Mean creatinine values and infectious complications were similar for each group. No patient experienced acute rejection during induction, and eight patients required dose escalation to sustain suppression of CD3 counts. The incidence of acute rejection at 6 and 12 months was significantly (P=0.004) greater in group III (38.2, 44.1%) than in either group I (15.1, 18.1%) or group II (14.7, 17.6%); relative risk 1.988 (95% CI 1.012-3.906). Formation of anti-OKT3 antibody was significantly (P=0.006) greater in group III (26.5%) than in group I (6%) or group II (2.9%). Group I recipients enjoyed significantly (P=0.001) fewer (2.17) OKT3 side effects on days 0, 1, and 2 than group II (3.03) or group III (2.49), and contained the largest number (61%) of recipients who experienced no side effects. Group I also exhibited the most suppressed profile of OKT3-induced release of tumor necrosis factor-alpha (P=0.006), interferon-gamma (P=NS), and interleukin-6 (P=0.01) on days 0 and 1. CONCLUSIONS: Low-dose 2.5-mg OKT3 with pretreatment of split-dose steroids on days 0, 1, and 2 provides the most effective method for OKT3 induction, which minimizes side effects for most patients. Subsequent maintenance therapy with cyclosporine, mycophenolate mofetil, and steroids provides effective rejection prophylaxis without increased complications for up to 3 years. Predepletion of T cells before exposure to OKT3 does not prevent cytokine release. PMID- 10868644 TI - Extension of transplantation free time by lamivudine in patients with hepatitis B induced decompensated cirrhosis. AB - BACKGROUND: Liver transplantation for hepatitis B virus (HBV)-induced cirrhosis carries a high risk of graft reinfection and poor prognosis. Active viral replication is considered a contraindication for transplantation in most centers. Lamivudine, a new nucleoside analog, is a potent inhibitor of HBV replication that has been used safely for pretransplantation suppression of HBV replication. METHODS: We report the pattern of response to lamivudine treatment in three consecutive patients with decompensated cirrhosis due to the replicative phase of chronic HBV infection. RESULTS: In addition to virological and biochemical response, impressive clinical improvement was noted in all three patients, with disappearance of the ascites and marked improvement of synthetic liver function tests. One patient converted to anti-hepatitis B surface and is free of symptoms 20 months after initiation of treatment. The other two patients experienced significant clinical improvement for 8 to 9 months and were removed from the waiting list for transplantation. However, progressive liver disease recurred in both patients--one underwent liver transplantation and the other is a candidate for the procedure. CONCLUSION: The administration of lamivudine for pretransplantation HBV suppression was associated with impressive clinical and biochemical improvement. Lamivudine may extend the transplantation free time in such patients. The mechanism of this desirable effect should be explored. PMID- 10868645 TI - Are wedge biopsies of cadaveric kidneys obtained at procurement reliable? AB - BACKGROUND: Single wedge biopsy of cadaveric kidneys from donors older than 55 is currently the standard method of evaluating their viability for transplantation. The degree of glomerulosclerosis presently determines whether a kidney can be transplanted, but most biopsies sample only the subcapsular region and may not accurately represent the true renal architecture. Our study evaluated the accuracy of transplant suitability determinations based upon the single wedge biopsy of cadaveric kidneys. METHODS: We took kidneys that were refused by UNOS centers on the basis of biopsy results, examined their histology in detail, and reviewed donor medical histories. Sections were taken from the upper, lower, and mid-portion of each kidney and stained with the periodic acid Schiff stain. Percentage and location of glomerulosclerosis and other relevant pathology were then determined in each section. We compared our findings with the results of the original wedge biopsies obtained at the time of procurement. RESULTS: Nine kidneys were obtained and examined. The wedge biopsies at the time of procurement showed glomerulosclerosis ranging from 8 to 36% (median 17%). The multiple kidney sections we analyzed showed fewer sclerosed glomeruli, ranging from 3 to 15% (median 7%, P<0.001), with most of the sclerosed glomeruli identified located in the immediate subcapsular region (P<0.001). CONCLUSIONS: Wedge biopsies of donor kidneys can overestimate the total amount of glomerulosclerosis, apparently because of a predominance of sclerosis in the kidney's subcapsular region, the area predominantly sampled by the usual wedge biopsy. These inappropriately high estimates of glomerulosclerosis can result in refusal of kidneys that might be suitable for transplantation. PMID- 10868646 TI - Fibrous intimal thickening at implantation as a risk factor for the outcome of cadaveric renal allografts. AB - BACKGROUND: During the past decade, the donor age of cadaveric renal allografts steadily increased. Because cerebrovascular injury is the main cause of death in this donor population, an increased prevalence of atherosclerotic lesions in the retrieved grafts could be anticipated. In a prospective study, we investigated the predictive value of morphologic lesions at implantation for the functional and morphologic outcome of cadaveric renal allografts at 1 1/2 years. METHODS: In 50 consecutive adult recipients of a cadaveric renal allograft, under cyclosporine-based regimen, implantation biopsies and subsequent protocol biopsies at 18 months were performed, and morphometrically analyzed for the extent of glomerulosclerosis, interstitial fibrosis, and atherosclerosis. Risk factors were assessed at implantation and during the subsequent observation period of 18 months. Endpoints for this study were: the 24-hr creatinine clearance (normalized for body surface area) and the fractional interstitial volume at 1 1/2 years. RESULTS: In multivariate analysis, fibrous intimal thickening at implantation (FIT) was the main determinant of the functional and morphologic outcome at 1 1/2 years. FIT represented a relative risk of 4.55 for interstitial fibrosis (95% CI=1.855-11.138), and 1.89 for impaired renal function (95% CI=1.185-3.007) at 1 1/2 years. FIT adversely affected fractional interstitial volume at 1 1/2 years (34.3 vs. 27.7%, P=0.004), as well as renal function (54 vs. 68 ml/min/1.73 m2, P=0.028). CONCLUSIONS: Fibrous intimal thickening at implantation is a determinant risk factor for the functional and morphologic outcome of cadaveric renal allografts at 1 1/2 years. PMID- 10868647 TI - Intrathoracic fluid volumes and pulmonary function during orthotopic liver transplantation. AB - BACKGROUND: Impaired pulmonary function is a frequent finding in patients undergoing orthotopic liver transplantation (OLT). Experimental data suggest an essential contribution of splanchnic ischemia and reperfusion as a result of intraoperative volume shifts, i.e., the accumulation of extravascular lung water (EVLW). Increases of intrathoracic blood volume (ITBV) and pulmonary blood volume (PBV) might additionally influence pulmonary capillary fluid filtration. The main objective of this study was to determine the intrathoracic volume changes during OLT and to test whether there were any relationships between intra- and extravascular volume shifts and pulmonary function, as determined by the calculation of venous admixture (QS/QT) and alveolar-arterial oxygen gradient (AaDO2). METHODS: Twenty-five patients undergoing OLT were studied. Using the transpulmonary double indicator dilution method, ITBV, PBV, and EVLW were determined from the mean transit times and exponential decay times of the indocyanine green and the thermal indicator curves recorded simultaneously with a fiberoptic catheter in the descending aorta. Recordings were made after induction of anesthesia, at the end of the anhepatic stage, immediately after reperfusion, and 1 and 4 h postoperatively. RESULTS: Significant increases in QS/QT related to changes of ITBV were observed after reperfusion. Only a minor impact on AaDO2 was perceived. EVLW remained constant during the study period. CONCLUSIONS: Postreperfusion increases of ITBV influence pulmonary function, as demonstrated by the increase in QS/QT. However, they need not be associated with greater EVLW levels, and impact on oxygenation is less severe than assumed. Hence, sufficient mechanisms protecting oxygenation and stalling increased EVLW seem to be present during uncomplicated human OLT. PMID- 10868648 TI - Prospective study of human betaherpesviruses after renal transplantation: association of human herpesvirus 7 and cytomegalovirus co-infection with cytomegalovirus disease and increased rejection. AB - BACKGROUND: Human herpesvirus 6 (HHV-6) and HHV-7 are two lymphotropic herpesviruses, which, like cytomegalovirus (CMV), have the potential to be pathogenic in immunocompromised individuals. We have conducted a prospective investigation to compare the natural history of HHV-6 and HHV-7 infection with that of CMV after renal transplantation. METHODS: Polymerase chain reaction was used to identify infections and quantify the viral load of CMV, HHV-6, and HHV-7 in peripheral blood samples from 52 renal transplant recipients. Betaherpesvirus infections were related to defined clinical criteria obtained by detailed examination of the clinical records of each patient for the immediate 120-day posttransplant period. RESULTS: CMV was the most commonly detected virus after transplant (58% of patients), followed by HHV-7 (46%) and HHV-6 (23%). Examining the time to first polymerase chain reaction positivity, HHV-7 infection was detected earlier than CMV (P=0.05). The median maximum CMV viral load was significantly higher than those for HHV-6 (P=0.01) and HHV-7 (P<0.0001) and a trend for HHV-7 viral load to be greater than HHV-6 (P=0.08). Clinicopathological analyses revealed that, in those patients with rejection, HHV-7 was associated with more episodes of rejection (P=0.02). In addition, there was a significant increase in CMV disease occurring in patients with CMV and HHV-7 co-infection compared to those with CMV infection only (P=0.04). CONCLUSIONS: HHV-7 should be further investigated as a possible co-factor in the development of CMV disease in renal transplant patients and may potentially exacerbate graft rejection. No clear pathological role was observed for HHV-6. PMID- 10868649 TI - Mycophenolate mofetil reduces late renal allograft loss independent of acute rejection. AB - BACKGROUND: Mycophenolate Mofetil (MMF) has been shown to significantly decrease the number of acute rejection episodes in renal transplant recipients during the 1st year. A beneficial effect of MMF on long-term graft survival has been more difficult to demonstrate. This beneficial effect has not been detected, despite the impact of acute rejection on the development of chronic allograft nephropathy and experimental evidence that MMF may have a salutary effect on chronic allograft nephropathy independent of that of rejection. METHODS: Data on 66,774 renal transplant recipients from the U.S. renal transplant scientific registry were analyzed. Patients who received a solitary renal transplant between October 1, 1988 and June 30, 1997 were studied. The Cox proportional hazard regression was used to estimate relevant risk factors. Kaplan-Meier analysis was performed for censored graft survival. RESULTS: MMF decreased the relative risk for development of chronic allograft failure (CAF) by 27% (risk ratio [RR] 0.73, P<0.001). This effect was independent of its outcome on acute rejection. Censored graft survival using MMF versus azathioprine was significantly improved by Kaplan Meier analysis at 4 years (85.61% v. 81.9%). The effect of an acute rejection episode on the risk of developing CAF seems to be increasing over time (RR=1.9, 1988-91; RR=2.9, 1992-94; RR=3.7, 1995-97). CONCLUSION: MMF therapy decreases the risk of developing CAF. This improvement is only partly caused by the decrease in the incidence of acute rejection observed with MMF; but, is also caused by an effect independent of acute rejection. PMID- 10868650 TI - Selection and outcome of living donors for adult to adult right lobe transplantation. AB - BACKGROUND: The shortage of cadaveric livers has sparked an interest in adult-to adult living donor transplantation. Right lobe donor hepatectomy is frequently required to obtain a graft of adequate size for adult recipients. Careful donor selection is necessary to minimize complications and assure a functional graft. METHODS: A four-step evaluation protocol was used for donor selection and satisfactory results of all tests in each step were required before proceeding to the next. Donors were selected based on a battery of laboratory studies chosen to exclude unrecognized infection, liver disease, metabolic disorders, and conditions representing undue surgical risk. Imaging studies included ultrasonography, angiography, magnetic resonance imaging, and intraoperative cholangiography and ultrasonography. The information obtained from liver biopsy was used to correct the estimated graft mass for the degree of steatosis. RESULTS: From March 1998 to August 1999, 126 candidates were evaluated for living donation. A total of 35 underwent donor right lobectomy with no significant complications. Forty percent of all donors that came to surgery were genetically unrelated to the recipient. A total of 69% of those evaluated were excluded. ABO incompatibility was the primary reason for exclusion after the first step (71%) and the presence of steatosis yielding an inadequate estimated graft mass after the second step (20%). CONCLUSIONS: Donor selection limits the application of living donor liver transplantation in the adult population. Unrelated individuals increase the size of the donor pool. Right lobe hepatectomy can be performed safely in healthy adult liver donors. Preoperative liver biopsy is an essential part of the evaluation protocol, particularly when the estimated graft mass is marginal. PMID- 10868651 TI - Bacterial translocation during graft-versus-host disease after small bowel transplantation is reduced following inhibition of inducible nitric oxide synthesis. AB - BACKGROUND: Increased nitric oxide (NO) production may contribute to intestinal barrier dysfunction and increased bacterial translocation (BT). Since BT may play a major role in graft-versus-host disease (GVHD) after small bowel transplantation (SBTx), we evaluated the role of NO production in GVHD after SBTX in the rat. METHODS: Using the standard model of semiallogeneic SBTx in the rat, we prepared three experimental groups. Recipients in group 1 received LBNF1-LBNF1 transplants and were treated with aminoguanidine (AG) (200 mg/kg), recipients in group 2 received Lewis-LBNF1 grafts and were injected with saline, and recipients in group 3 received Lewis-LBNF1 transplants and AG (200 mg/kg). Urine nitrite/nitrate levels were measured daily, and BT was determined by culturing peritoneal swabs, mesenteric lymph nodes, spleen, liver, and blood. RESULTS: In group 1 we detected indefinite survival with normal histology. In group 2 a survival of 10.5 +/- 1.1 days was reached, and the typical histological features of acute GVHD were observed. The animals in group 3 showed a mean survival of 14.8 +/- 0.6 days (P<0.02 compared with group 2) and the histological features of acute GVHD, although with a prolonged time course. Comparing NO production and BT between groups 2 and 3 we detected significantly reduced NO production on postoperative days 2-9 (P<0.03) and significantly decreased BT on postoperative days 3 and 9 (P<0.03). CONCLUSION: Inhibition of inducible NO synthesis with AG reduces NO production, decreases BT, and prolongs survival during GVHD after SBTx and therefore may be a useful addition to standard treatment protocols for GVHD. PMID- 10868652 TI - Binding of human natural antibodies to nonalphaGal xenoantigens on porcine erythrocytes. AB - BACKGROUND: Xenotransplantation is considered one of possible solutions for the serious shortage of organs and cells in transplantation. Although the alphaGal epitope (Gal alpha1,3Gal beta1,4GlcNAc-R) has been identified as being a major xenoantigen responsible for hyperacute rejection, the removal of anti-alphaGal antibody alone from human serum is insufficient to circumvent antibody-mediated immune responses. METHODS AND RESULTS: We report here the characterization of xenoreactive human natural antibodies against antigens without the alphaGal epitope (nonalphaGal xenoantigens) on porcine erythrocytes using flow cytometry and the evidence for their involvement in complement-mediated hemolysis. Furthermore, a novel protein of 45-kDa has been isolated from the porcine erythrocyte membrane as a major protein antigen recognized by human anti nonalphaGal. CONCLUSION: The data presented here will add to our knowledge of xenoantigens on porcine red cells and be important for developing strategies to produce modified red cells immunologically compatible to humans. PMID- 10868653 TI - Alloantigen-driven T cell death mediated by Fas ligand and tumor necrosis factor alpha is not essential for the induction of allograft acceptance. AB - BACKGROUND: Fas ligand (FasL)-Fas and tumor necrosis factor alpha (TNFalpha) tumor necrosis factor receptor (TNFR) interactions regulate immune responses and contribute to self-tolerance by mediating antigen-driven T cell apoptosis. It is not known whether FasL and TNFalpha, expressed by the recipient's lymphoid or nonlymphoid cells, are essential for the apoptosis of alloreactive T lymphocytes and the induction of allograft acceptance. METHODS: We compared the survival of fully allogeneic vascularized cardiac allografts between wild-type (wt) and FasL mutant (gld) recipient mice. In addition, we studied cardiac allograft survival in gld mice injected with TNFalpha-neutralizing antibody. Allograft acceptance (graft survival >100 days) was induced by treating the recipients with CTLA4Ig, a recombinant fusion protein that blocks B7-CD28 T cell costimulation. In vivo alloantigen-driven apoptosis of mature CD4+ and CD8+ T lymphocytes was analyzed in mice repeatedly stimulated with allogeneic splenocytes. RESULTS: We found that CTLA4Ig induces 100% long-term acceptance of cardiac allografts in wt and gld mice. Similarly, CTLA4Ig induced 100% allograft acceptance in gld recipients injected with TNFalpha-neutralizing antibody. In vivo alloantigen-driven apoptosis of mature CD4+ and CD8+ T cells was significantly reduced in gld mice and in wt mice treated with anti-TNFalpha antibody. However, neutralizing TNFalpha activity in gld mice failed to abrogate alloantigen-driven T cell apoptosis. CONCLUSIONS: These data indicate that: (1) FasL and TNFalpha expression are not obligatory for the induction of long-term allograft acceptance by CTLA4Ig and (2) FasL- and TNFalpha-independent death pathways contribute to alloantigen-driven T cell apoptosis. PMID- 10868654 TI - Organ donor or graft pretreatment to prolong allograft survival: lessons learned in the murine model. AB - Permanent donor-specific tolerance to allografts is the goal of transplantation research. Currently, morbid immunosuppressive therapy is used to mitigate rejection initiated in part by Ia-bearing interstitial graft dendritic or antigen presenting cells (APCs) that are thought to migrate into the host after transplantation. We hypothesized that donor or organ immune modulation directed against graft APCs might influence graft immunogenicity and promote prolonged graft acceptance in histoincompatible hosts in the absence of immunosuppressive therapy. Haplotype-specific monoclonal antibodies (mAb), mAb specific to graft APC, adhesion or costimulatory molecules and anti-LFA-1-Ricin and anti-Iak-Ricin immunoconjugates (IC) were prepared and administered in varying doses and time intervals to donor C3H/HeJ (H-2k) mice. Thereafter, their spleens and hearts were removed at varying time intervals and used either as stimulator cells in one-way mixed lymphocyte reaction or transplanted into naive Balb/c (H-2d) recipients, respectively. Explanted C3H hearts were pretreated with anti-Iak mAb on the Langendorf apparatus. Hearts were also used from major histocompatibility complex (MHC) class I, MHC class II, and MHC class I- and II-deficient "knockout" mice. Splenocytes exposed to at least 500 microg of anti-Iak mAb in vivo for more than 4 hr were able to inhibit the mixed lymphocyte reaction to almost background levels, but only after incubation with rabbit complement in vitro. Similarly pretreated cardiac allografts (both in vivo or explanted and pretreated on the Langendorf apparatus) did not experience prolonged survival in nonimmunosuppressed Balb/C recipients when compared with control solutions, regardless of the concomitant use of complement. Splenocytes from immunoconjugate pretreated donors inhibited the mixed lymphocyte reaction completely without the use of complement; however, hearts from these donors also did not experience prolonged survival nor donor hearts exposed to mAb specific for graft APC, adhesion or costimulatory molecules. Only hearts from MHC class I and class II "knockout" mice survived significantly longer than controls. We conclude that donor or graft pretreatment with haplotype-specific anti-Ia mAb, haplotype specific immunoconjugates, or mAb directed against graft APC, adhesion or costimulation molecules have little efficacy in promoting acceptance of cardiac allografts in nonimmunosuppressed recipients. The enhanced survival of hearts from MHC class I- and class II-deficient donors suggest that novel methods to effect the immunogenicity of the graft will be required if long-term allograft acceptance is to be achieved in the absence of host immunosuppression. PMID- 10868655 TI - Expression of GMP-140 (P-selectin) correlates with graft viability in cold preserved rat livers. AB - BACKGROUND: Ischemia/reperfusion injury is an inflammatory process involving cytokine release, Kupffer cell activation, and sinusoidal endothelial cell activation. GMP-140 is synthesized by endothelial cells. METHODS: We analyzed by Western blotting the expression of GMP-140 in a syngeneic rat liver transplantation model using grafts preserved for different periods of time. RESULTS: Compared with prereperfusion samples, expression did not change significantly in freshly harvested and 4-hr preserved livers. In grafts preserved for 24 hr (100% survival), GMP-140 levels increased dramatically at 1 hr, then returned to baseline at 24 hr after transplantation. Forty-eight hour preserved grafts (0% survival) showed a decreasing expression. To identify possible mediators, the effects of tumor necrosis factor-alpha and interleukin-1beta on GMP-140 expression in primary sinusoidal endothelial cells were analyzed. These cytokines increased both the percentage of stained cells as well as their mean staining fluorescence. CONCLUSIONS: The absence of increase in 48-hr grafts suggests that GMP-140 may distinguish viable from nonviable livers. PMID- 10868656 TI - Successful reuse of portal-enteric technique in pancreas retransplantation. AB - BACKGROUND: The portal venous and enteric drainage (P-E) technique was developed to avoid systemic hyperinsulinemia and bladder related complications. Pancreas retransplantation (Re-Tx) is an important option for patients who have lost their primary grafts. It is unknown whether the P-E technique can be repeated safely in patients who have lost their first pancreas transplant. METHODS: Five patients who lost their pancreas graft after simultaneous kidney-pancreas transplantation with P-E drainage underwent pancreas Re-Tx, again using the P-E technique. RESULTS: P-E Re-Tx was successful in all 5 patients without any technical difficulties or complications. CONCLUSION: The P-E technique can be reused with excellent results in pancreas Re-Tx. PMID- 10868657 TI - Lung transplantation for advanced bronchioloalveolar carcinoma confined to the lungs. AB - BACKGROUND: Bronchioloalveolar carcinoma (BAC) is a well-differentiated lung adenocarcinoma that has a tendency to spread chiefly within the confines of the lung by aerogenous and lymphatic routes and may therefore be amenable to local therapy. However, a high rate of local recurrence after lung transplantation was recently reported. We describe two patients with unresectable and recurrent extensive BAC limited to the lung parenchyma who underwent lung transplantation with curative intent. METHODS: Patients were chosen to receive lung transplants for BAC if they met the following criteria: (1) recurrent or unresectable BAC limited to the lung parenchyma without nodal involvement and (2) suitable candidate for lung transplantation. RESULTS: The first patient relapsed in the lungs at 9 months after transplantation. The pattern of disease suggested contamination of the new lungs at the time of implantation. Repeat lung transplantation was performed, with cardiopulmonary bypass and irrigation of the remaining upper airway. This patient has had no evidence of local or systemic tumor recurrence at more than 4 years since the second transplantation. The second patient underwent transplantation using the modified technique and expired 16 months after transplantation of other causes. An autopsy showed no evidence of recurrent BAC in the lungs or of metastatic lesions at any site. CONCLUSIONS: Lung transplantation may be an option for unresectable or recurrent BAC confined to the lungs. Isolation of the diseased lungs and the use of cardiopulmonary bypass during surgery may be important in this disease and should be studied further. PMID- 10868658 TI - Influenza vaccination does not promote cellular or humoral activation among heart transplant recipients. AB - BACKGROUND: The impact of influenza vaccination on in vitro parameters of cellular and humoral immunity, anti-viral titers, and clinical outcome was evaluated among cardiac transplant recipients. METHODS: Blood was collected from 29 patients before and 3-4 weeks after influenza vaccination and tested for phenotypic changes in lymphoid subpopulations and generation of antibodies against the allograft and vaccine. RESULTS: Vaccination did not change the percentage of lymphoid subpopulations and did not induce generation of anti-HLA alloantibodies. Anti-vaccine response was detected in 12 of 29 patients and did not correlate with rejection history, length of graft survival, or immunosuppressive therapy. Vaccination did not change the frequency of rejection. Flu-like symptoms were reported in one patient but not confirmed microbiologically. CONCLUSION: Despite the small number of patients in the study, influenza vaccination did not induce undesirable side effects, such as graft rejection or allo-sensitization. Generation of a positive anti-vaccine response was lower among the transplant recipients than healthy volunteers (41% vs. 80%). Clinical efficacy of the vaccine among the responders was not evaluated. PMID- 10868659 TI - Liver transplantation improves cirrhosis-associated impaired oral glucose tolerance. AB - BACKGROUND: Thirty-five percent to 80% of cirrhotic patients have impaired glucose tolerance (IGT) or diabetes mellitus (DM). Diabetic cirrhotics have higher morbidity and mortality than nondiabetics. Therefore, it would be worthwhile to determine whether liver transplantation improves glucose homeostasis in these patients. METHOD: A total of 26 patients awaiting liver transplantation were evaluated for impaired glucose homeostasis by fasting blood glucose and/or oral glucose tolerance tests (OGTT). Five patients underwent transplant surgery within 1 year of OGTT and had a repeat OGTT 3-6 months after transplantation. RESULTS: Sixty-five percent (17/26) of the patients had abnormal glucose homeostasis. Twenty-three percent (6/26) met American Diabetes Association criteria for DM, and another 42.3% (11/26) had IGT. All patients had normal HbA1C levels. After transplantation, the 2-hr blood glucose improved in four patients and the mean 2-hr glucose level was reduced (204 +/- 94 vs. 132 +/- 53 mg/dl [mean +/- SD, P=0.051]). CONCLUSION: Liver transplantation can reverse cirrhosis-associated impaired glucose tolerance. PMID- 10868660 TI - Successful treatment of a patient suffering from severe acute potassium dichromate poisoning with liver transplantation. AB - BACKGROUND: Oral ingestion of potassium dichromate produces a complex spectrum of complications. It has an extremely poor prognosis and usually leads to rapid death. METHODS: We report the case of a 16-year-old male patient who was admitted to hospital after oral ingestion of potassium dichromate with suicidal intention. RESULTS: The patient's condition deteriorated, and he became comatose within 5 days in spite of immediate attempts at detoxification. Because of irreversible liver failure, which occurred within 2 days after admission, and because of cerebral edema, the decision to perform a liver transplantation was made. On day 6 after admission, a compatible donor liver was transplanted. The course of liver transplantation and the patient's subsequent recovery were uneventful. CONCLUSION: The rationale for the delayed transplantation was to avoid damage of the new organ because of high serum chromium levels. Despite severe organ damage, the chromium content of the liver was increased. To the authors' knowledge, this is the first case report of acute toxic liver failure, caused by potassium dichromate poisoning, treated successfully by means of liver transplantation. PMID- 10868661 TI - Induction of RANTES, HLA-DR, and intercellular adhesion molecule-1 on highly purified distal tubular cells from human kidney. AB - BACKGROUND: Expression of proinflammatory molecules by tubular epithelial cells plays an important role in renal allograft rejection and inflammatory kidney diseases. Different studies from patients with acute rejection point to the involvement of distal tubular segments. At present no in vitro system for the human distal tubule is established. METHODS: Human distal tubular cells were isolated immunomagnetically. Cultured cells were stimulated with cytokines (interferon-gamma, tumor necrosis factor-alpha, interleukin-1beta, or a cytokine mix). Secretion of RANTES (regulated upon activation, normal T-cell expressed and secreted) was evaluated with an enzyme-linked immunoassay. Expression of HLA-DR and intercellular adhesion molecule (ICAM)-1 was assessed by flow cytometric analysis and immunofluorescence studies. RESULTS: Our data clearly indicate that distal tubular cells express RANTES, HLA-DR, and ICAM-1 in response to a mixture of specific cytokines. Dexamethasone inhibited the induced expression of RANTES and HLA-DR significantly, but not that of ICAM-1. CONCLUSIONS: We demonstrate an appropriate in vitro system for the human distal tubule. The present study proves the involvement of the distal tubular segment during inflammatory kidney diseases. PMID- 10868662 TI - Renal transplant for recipients over 60 years old. PMID- 10868663 TI - PSA-based screening for prostate cancer after renal transplantation. PMID- 10868664 TI - Job stress and the occupational gradient in coronary heart disease risk in women. The Stockholm Female Coronary Risk Study. AB - Recent studies of men have shown that job stress is important in understanding the occupational gradient in coronary heart disease (CHD), but these relationships have rarely been studied in women. With increasing numbers of women in the workforce it is important to have a more complete understanding of how CHD risk may be mediated by job stress as well as other biological and behavioural risk factors. The objective of this study was to examine the occupational gradient in CHD risk in relation to job stress and other traditional risk factors in currently employed women. We used data from the Stockholm Female Coronary Risk Study, a population based case-control study, comprising 292 women with CHD aged 65 years or younger and 292 age-matched healthy women (controls). An inversely graded association was observed between occupational class and CHD risk. Compared with the highest (executive/professional), women in the lowest occupational class (semi/unskilled) had a four-fold (95% CI 1.75-8.83) increased age-adjusted risk for CHD. Simultaneous adjustment for traditional risk factors and job stress attenuated this risk to 2.45 (95% CI 1.01-6.14). Neither job control nor the Karasek demand-control model of job stress substantially explained the increased CHD risk of women in the lowest occupational classes. It is likely that lower occupational class working women face multiple and sometimes interacting sources of work and non-work stress that are mediated by behavioural and biological factors that increase their CHD risk. PMID- 10868665 TI - Hospital pharmacy automation: collective mobility or collective control? AB - This study examines the relationship between new technology and pharmacy's changing role in the division of labour in health care. Does automation advance pharmacy's collective mobility project and transition to its new role as a clinical rather than a merely technical profession? Alternatively, does it threaten pharmacy's cultural authority and control of medication dispensing? Does technology serve as an artifact demarcating occupational and gender boundaries in the drug distribution labour process? These issues were examined through qualitative case studies conducted at three hospital pharmacies located in mid sized Canadian health care facilities. All three institutions had implemented automated drug distribution technology in the expectation that it would allow more pharmacist's time to be shifted from dispensing-related to clinical activities. It was found that pharmacists had attempted to maintain control of their labour process and had resisted peripheralization from drug dispensing. Technological processes did serve as artifacts reinforcing occupational demarcations as well as professional solidarity vis a vis hospital administrators although its impact on gender boundaries was less evident. PMID- 10868666 TI - The politics of herbal drugs in Korea. AB - Hanbang, the Korean medical practice with origins in classical Chinese texts, is a prominent part of the Korean health care system. Hanbang physicians, called hanuisas, are looked down on by biomedical doctors, but their practice has enjoyed increasing popularity for several decades. As the market for herbal preparations has become more lucrative, biomedical pharmacists have begun to participate in it. The Pharmaceutical Act in 1993 explicitly allowed pharmacists to prescribe and dispense herbal drugs. This provoked a bitter public conflict between hanuisas and pharmacists, involving street demonstrations and strikes. The hanuisas asserted that the pharmacists were unqualified to assume their traditional practice. They also agitated for recognition in the state-sponsored system of health care and for the state's support for developing Hanbang medicine. This paper attributes the conflicts concerning Hanbang to the expanding market for herbal preparations, Korean nationalism, and to the oversupply of biomedical pharmacists. PMID- 10868667 TI - Caesarean section delivery in Kerala, India: evidence from a National Family Health Survey. AB - Ensuring safe pregnancy and motherhood occupies a pivotal role and has been considered as one of the key issues in the framework of reproductive and child health programmes. Evidence from research studies indicate that there is a growing tendency for caesarean section deliveries especially during complications confronted at the time of pregnancy and delivery. The present study focuses on the demographic, antenatal care, spatial and socio-economic variables associated with caesarean section delivery in Kerala, India. The data from the National Family Health Survey has been utilised for this purpose. The results from logistic regression models indicate that maternal age, birth order, current age, births in health institutions and spatial differences were significantly associated with caesarean section deliveries in Kerala. The older cohorts of mothers were found at higher risk to have caesarean section when compared to their younger counterparts. When controlled for demographic variables, the odds for caesarean section was about 1.7 times more likely to occur in private health institutions. The inclusion of spatial and socio-economic variables has neither influenced the demographic and antenatal care variables nor showed any significant association with caesarean section delivery in the state. The present study calls for that a detailed investigation on behavioural aspects of both the physician and the patient with regard to type of delivery in the state. Information related to pregnancy and health related aspects needs to be monitored more accurately, both in the public and private hospitals, to understand the determinants associated with caesarean section. PMID- 10868668 TI - Defensive medicine during hospital obstetrical care: a byproduct of the technological age. AB - This paper presents an alternative perspective on defensive medicine. Defensive medicine is usually understood as arising from the effect of law on medicine through fear of litigation. Of equal significance, however, is the complementary influence of medicine on law through technological innovation, and, more importantly, the way that medicine and law develop dialectically. Each shapes the other in establishing the standards of care central to both clinical medicine and to actual or potential legal action. Excessive testing owing to fear of litigation indicates that defensive medicine is being practised in a particular setting, but it does not explain why this is so. To understand why defensive medicine occurs and why it is so troubling to clinicians requires an understanding, not only of medical and legal developments, but of a political economic system and the beliefs and values of a society. Defensive medicine is discussed in relation to hospital obstetrical scenarios commonly associated with fear of litigation: fetal oxygen deprivation ("distress"), which is detected using an electronic fetal monitor, and prolonged labor, known as "dystocia". The material presented is taken from a medical anthropological study of obstetrical care in rural British Columbia, Canada. Litigation fears are shown to result less from rare, albeit often devastating, allegations of malpractice than from doctors adopting a role as "fetal champions", together with the introduction of electronic monitoring technology. The paper concludes by asserting that, rather than being in an adversarial relationship, medical practice and associated litigation primarily work together to reinforce each other, and the social conditions in which defensive medicine occurs. PMID- 10868669 TI - Searching for socioeconomic risk factors in perinatal mortality in Kuwait: a case control study. AB - The aim of this paper was to investigate whether socioeconomic factors such as parent's education, occupation, and income constitute risk factors in perinatal mortality after controlling for biological variables such as birth weight and length of gestation, and maternal factors such as age, parity and reproductive history. A case-control study covering all perinatal deaths in Kuwait was conducted for one year from 1 October, 1997 to 30 September, 1998. Each case (perinatal death) was matched with a control (live birth). Matching criteria were: father's nationality, place, and date of birth. Information was successfully collected on 463 matched pairs, 274 Kuwaitis and 189 non-Kuwaitis. Only singleton births were included in the analysis. Bivariate analysis showed that several of the socioeconomic variables (e.g. lower education, lower income) increased the risk of a perinatal death. However, none of these variables remained significant in the multivariate analysis in which birth weight and length of gestation emerged as the two major determinants of perinatal deaths among both nationality groups. Among the Kuwaitis, primiparity and high parity, and previous history of miscarriage were also significant risk factors. Among the non-Kuwaitis, none of the socioeconomic factors, or the maternal factors, were significant predictors of perinatal mortality. For Kuwaitis, it appears that the government's policies and programs aimed at reducing social inequalities in the society have been effective in eliminating perinatal mortality differences between socioeconomic groups. Among non-Kuwaitis, the lack of differences is reflective of the fact that this group is relatively homogenous and selective of the more affluent who can bring the family to Kuwait. Both nationality groups benefit from the government's free health services. However, charges for non Kuwaitis are due to be levied soon which may increase disparities in access to health care. PMID- 10868670 TI - Frequency and timing of antenatal care in Kenya: explaining the variations between women of different communities. AB - Appropriate antenatal care is important in identifying and mitigating risk factors in pregnancy but many mothers in the developing world do not receive such care. This paper uses data from the 1993 Kenya Demographic and Health Survey to study the variations in the use of antenatal services in Kenya. The analysis is based on modelling the frequency and timing of antenatal visits using three-level linear regression models. The results show that the use of antenatal care in Kenya is associated with a range of socio-economic, cultural and reproductive factors. The availability and accessibility of health services and the desirability of a pregnancy are also important. Use of antenatal care is infrequent for unwanted and mistimed pregnancies; even women who use antenatal care frequently appear to be less consistent if a pregnancy is mistimed. The results also indicate that women are highly consistent in the use of antenatal care during pregnancies. The intra-woman correlation coefficient for the frequency of antenatal visits ranges between 50% and 80% with greater correlation for wanted pregnancies to women in urban areas. PMID- 10868671 TI - Social inequality, population health, and housing: a study of two Vancouver neighborhoods. AB - An emerging 'population health' framework for understanding inequalities in health identifies the structure of social relations as a crucial factor in shaping human health and well-being. However, there remain many unanswered questions about the mechanisms through which social relations might shape the health status of individuals and populations. Housing plays a central role in routinized, everyday life and is fundamentally bound up in one's sense of control over life circumstances. Housing and property markets are significant in the distribution of wealth and are an important arena for the exercise of power relations. Housing circumstance is crucial in the production and reproduction of social identity and social status. Yet little has been written on the influence of inequalities generated by housing and housing markets on the differential distribution of health status. This paper reports the findings of an empirical study of relationships between socioeconomic status, material and meaningful dimensions of housing and home, and health status. Our objective is to investigate ways in which material and meaningful factors related to housing, in conjunction with other dimensions of the social environment, could operate to produce systematic inequalities in health status across social strata. The data for this study were obtained through a mailed survey of residents in the Mount Pleasant (n = 322) and Sunset (n = 206) neighborhoods of Vancouver, Canada. They suggest that, in concert with commonly used measures of socioeconomic status, both material and meaningful dimensions of housing and home are associated with health status in a direction consistent with expectations following from our analytical model. PMID- 10868672 TI - How was life after treatment of a malignant brain tumour? AB - The malignant glioma is a severe disease with an unfavourable prognosis. Aside from a few case studies, the knowledge of the victimised patients' lives from diagnosis to death is mainly restricted to studies assessing functional status and rating quality of life by means of questionnaires. From a clinician's perspective this knowledge is not sufficient. By introducing the concepts 'time of everyday life' and 'time of disease', the purpose of this paper is to supplement with descriptive knowledge of clinical value. Twenty-eight patients with malignant gliomas and their spouses were followed during the course of the disease by repeated interviews. The time after treatment was then judged as representing, 'time of everyday life' or 'time of disease'. Life after treatment turned out to be quite varied. To slightly more than a third of the patients', life-continuity was lost, experiencing only 'time of disease'. Among the others who were judged to experience 'time of everyday life' and who were of working age, nearly two-thirds were able to resume work or studies on a part-time basis. In the total sample, the mean 'time of everyday life' turned out to be nearly equal to 'time of disease', 6.1 and 5.4 months, respectively. The findings are illustrated by case descriptions and the conceptualisation of time into 'everyday life' and 'disease' is proposed as meriting further study. PMID- 10868673 TI - Measuring equity in access to health care. AB - This article develops and uses methodologies to: (1) measure equity in the distribution of access to health services; and (2) measure the impact of health insurance programs on equity. The article proposes two egalitarian-based indicators for measuring equity in terms of access to health care--a concentration coefficient derived from the Gini coefficient, and the Atkinson distributional measure and also employs a weighted Utilitarian social welfare function to measure overall levels of access. The article defines access as the use of health care by individuals with a need for care; need is measured as self reported morbidity. The setting for the empirical application is the country of Ecuador. The Ecuador Social Security Institute runs a General Health Insurance (GHI) program, whose affiliates are primarily workers in the formal sector of the economy. The principal data source is the 1995 Ecuador Living Standards Measurement Survey. The study uses a microeconomic health care demand model and bivariate probit estimation techniques to measure the impact of insurance on health service use for each quintile of adjusted per-capita household expenditure. The study also predicts health care use and program impact for each quintile under a series of simulation scenarios corresponding to proposed expansion of eligibility for the GHI program. The GHI program increases overall access to health care, but has a negative impact on equity in the distribution of health services. The benefits of the program, calculated as its marginal impact on the probability of using of health care, have a strongly regressive distribution. Expanding eligibility to the self-employed makes the benefit more equitably distributed (but still inequitable), and increases overall social welfare considerably. Expanding eligibility to the dependents of the insured person has similar effects, although less important in magnitude. PMID- 10868674 TI - Health risk and inequitable distribution of liquor stores in African American neighborhood. AB - In this paper we examine whether the physical availability of alcohol is greater in predominantly African American communities compared to predominantly white communities as indicated by the presence of off premises liquor stores. We investigate the extent to which the income status of the residents of a community mediates the relationship between community racial composition and alcohol availability; and explore whether the intersection of race and class places low income African American communities at increased risk to have such stores located in their communities. Multivariate analytic techniques are used to examine the relationship between community racial composition, median income of neighborhood residents and per capita number of alcohol outlets in 194 census tracts in Baltimore, Maryland. The analysis found that liquor stores are disproportionately located in predominantly black census tracts, even after controlling for census tract socioeconomic status. Census tracts that are both low income and predominantly African American have substantially more liquor stores per capita than other census tracts. Although it is beyond the scope of the present study, our data reveal significant associations between the presence of liquor stores and the risk of health-related social problems in low income neighborhoods. More research needs to be done on the impact of alcohol on the social, psychological, and physiological health of low income urban populations. PMID- 10868675 TI - Going down to the local: incorporating social organisation and political culture into assessments of decentralised health care. AB - The social organisation and political culture of the society in which an organisation is embedded can have major effects on the way in which organisational policy is implemented and on how that organisation functions. Research on health sector reforms has paid scant attention to this aspect. If the claims made for decentralised management in the health sector are to be evaluated seriously, it is critical to develop concepts and methods to evaluate not only the formal organisation and the outputs of the health system, but also the aspects of local social organisation and political culture within which that local health system is embedded that may mediate their relationship. The paper explores three cases of district health systems in Northeast Brazil in order to identify aspects of local social organisation and political culture that appear to influence the implementation of the reforms and thereby potentially impact upon the quality of the care provided. The results of the study indicate the importance that aspects of local social organisation and political culture may exert on the operations of a decentralised health system. Key aspects identified are: the space for autonomy; the space for local voice in political life; personalized and institutionalised influences on autonomy and local voice; differences of involvement of health staff with the district; different spaces of acceptable practice and accountability. These factors are seen to moderate the intent of the health reforms at all stages in their implementation. Three possibilities are discussed for the nature of the interaction in terms of cause and effect between the formal organisation of the health system and its local context. Seeing this relationship as one of a dialogue offers some cautious optimism for the potential of the reform agenda. The paper closes with suggestions on how to take this line of research forward. PMID- 10868676 TI - Use of a ground beef model to assess the effect of the lactoperoxidase system on the growth of Escherichia coli O157:H7, Listeria monocytogenes and Staphylococcus aureus in red meat. AB - The ability to preserve food in a state that is both appetising and nutritious is a basic requirement for health. Food poisoning represents a major source of illness and loss of productivity in many developed countries. Of particular concern in recent years are outbreaks of food poisoning associated with Escherichia coli O157:H7 or Listeria monocytogenes, many of which have been associated with the consumption of ground meat. Many of the chemicals presently licensed for use as food preservatives are increasingly being questioned with regard to their effects on humans, creating pressure on food suppliers to consider the use of 'natural' alternatives to these chemical agents. The potential use of one such agent, the lactoperoxidase system (LPS), for use in ground meat preparations is examined in this study. The degree of inhibition of growth of E. coli O157:H7, L. monocytogenes L45 and S. aureus R37 by LPS was examined in a broth system at 37 degrees C and in a ground beef system at 0, 6 and 12 degrees C. The degree of inhibition by LPS of natural populations of microorganisms present in ground beef obtained from eight retail outlets and incubated at room temperature was also examined. For each of the strains examined, sensitivity from most to least sensitive followed the order L. monocytogenes L45, S. aureus R37 and E. coli O157:H7. In each case the ability of LPS to inhibit growth was highly temperature dependent and maximal at a temperature permissive but not optimal for growth of the test strain. The numbers of bacteria detected in ground beef obtained from retail outlets varied considerably between the eight samples. In all cases, numbers of bacteria increased markedly in the uninhibited control over the 4 h incubation time and, with the exception of one faecal coliform count, growth of the microbial populations was strongly inhibited by the presence of LPS. It was concluded that LPS could potentially be applied to a considerably wider range of food products than those to which it is presently restricted. PMID- 10868677 TI - Growth of Salmonella choleraesuis subspecies diarizonae serovar 61:k:1,5,(7) in broth and fresh mutton. AB - Three serovars of Salmonella choleraesuis (IIIb 61:k:1,5,(7), Enteritidis and Dublin) were grown in broths of pH 5.5 and 6.2 and incubated at 4, 6, 8 or 12 degrees C. Growth in the broth, measured by means of an increase of absorbance, was not observed below 8 degrees C. At 8 and 12 degrees C, the maximum growth rate (mu(max)), lag period and maximum absorbance level (max(abs)) varied according to serovar and pH. In general, serovar IIIb 61:k:1,5,(7) and serovar Enteritidis grew better than serovar Dublin. The effect of pH on lag period, seen for serovar IIIb 61:k:1,5,(7) and serovar Enteritidis at 8 degrees C, was absent at 12 degrees C, while the effect of pH regarding the mu(max) and the max(abs) was observed also at 12 degrees C. Furthermore, the growth serovar IIIb 61:k:1,5,(7) in normal and dark, firm and dry meat at 8 degrees C with ambient air in competition with the natural microbial flora was tested in minced meat and chops. Slow growth of serovar IIIb 61:k:1,5,(7) was observed in minced meat. The low virulence and the ordinary growth capabilities indicate that serovar IIIb 61:k:1,5,(7) will probably not represent a serious hazard to the public health. PMID- 10868678 TI - A model describing the effect of temperature history on lag time for Listeria monocytogenes. AB - A model was built to describe the influence of the temperature and the duration of pre-incubation on the lag time for regrowth of Listeria monocytogenes at low temperature. This model is consistent with the usual procedure used to calculate lag times of cultures growing under fluctuating temperatures. It also describes the effect of prolonged starvation conditions on the regrowth lag time and takes into account the influence of the physiological state of inocula in predictive models. PMID- 10868679 TI - Predictive microbiology and food safety. AB - The evaluation of risk in food safety requires knowledge of the probability that microbial population sizes will not exceed defined levels. This probability is evaluated assuming that the growth of the microbial population can be described by the Gompertz equation with the variance of growth depending on the population size. It is shown that the probability density associated with this phenomenon is skewed, so that the risk of a high microbial population is greater than that which would be estimated using a symmetrical probability distribution such as the Gaussian. Maximum likelihood estimates of the parameters of the Gompertz equation based on the derived probability density are calculated using data published by Zwietering et al. [23] for the growth of Lactobacillus plantarum under different temperatures. The probability that a microbial population of a given size will exceed an unacceptable level within a given time is calculated for growth at two temperatures, 10 and 25 degrees C. The implication of these theoretical results for the management of risk in food safety and in the design of hazard analysis critical control point procedures is discussed. PMID- 10868680 TI - Effects of atmospheric conditions, NaCl and pH on growth and interactions between moulds and yeasts related to blue cheese production. AB - The starter culture Penicillium roqueforti, undesired cultures Penicillium caseifulvum and Geotrichum candidum and the potential starter culture Debaryomyces hansenii were examined for their growth and interactions at environmental conditions similar to the Danish blue cheese Danablu. The combined effect of low oxygen (0.3%) and high level of carbon dioxide (25%) at 4% NaCl w/v (a(w) 0.97) and pH 4.5 and 6.5 on radial growth was examined on a cheese medium at 10 degrees C. P. roqueforti and G. candidum were well adapted to growth at low levels of oxygen and high levels of carbon dioxide but G. candidum was not able to grow in the presence of 4% NaCl (w/v). Growth of P. caseifulvum was strongly inhibited at atmospheric conditions comprising 25% carbon dioxide, especially in combination with 0.3% oxygen. Generally D. hansenii showed strong growth at all environmental conditions examined, except at 0.3% oxygen combined with 25% carbon dioxide and 4% NaCl (w/v). Growth and sporulation of P. roqueforti was highly affected in the presence of G. candidum at 25% carbon dioxide irrespective of levels of oxygen and NaCl in the cheese media. P. caseifulvum caused a pronounced inhibitory effect towards growth of P. roqueforti and D. hansenii at 21% oxygen. D. hansenii caused weak inhibition of P. roqueforti at 21% oxygen, while positive interactions between the two species were indicated at 25% carbon dioxide and 0.3% oxygen. PMID- 10868681 TI - Characterization of Brevibacterium linens pigmentation using spectrocolorimetry. AB - Spectrocolorimetry was used for the evaluation of Brevibacterium linens pigmentation. A total of 23 strains of Brevibacterium linens, from various sources, were plated onto solid agar media and their color coordinates were measured and expressed by the L*a*b* colorimetric system. All the strains were located along a straight line with a hue of 66 degrees (orange color). The effect of light exposure versus storage in the dark was also investigated using this approach. Three groups with distinct behaviors were demonstrated for the 23 B. linens strains. PMID- 10868682 TI - Modification of the bile salts-Irgasan-brilliant green agar for enumeration of Aeromonas species from food. AB - The present study evaluated the productivity of BIBG medium for the isolation of Aeromonas spp. from food and describes a modification of the BIBG medium (mBIBG) (increased pH (8.7), replacement of xylose by soluble starch as a carbon source, decreased concentration of bile salts) to increase its selectivity and electivity. Using the mBIBG medium, growth of the majority of the Enterobacteriaceae (9/10) was suppressed except for Citrobacter freundii. The mBIBG medium supported growth of Pseudomonas species but a clear distinction between Aeromonas and Pseudomonas colonies could be made. Interpretation of the mBIBG medium should be performed after 24 h of incubation. It was noted that three of the 27 Aeromonas strains tested did not develop on the mBIBG medium. The ability or inability to grow on a selective medium is strain-dependent. Enumeration of Aeromonas species (A. hydrophila LMG 3771, A. caviae LMG 3775, A. veronii biovar veronii LMG 9075, A. veronii biovar sobria LMG 13071) from artificially contaminated foods (shrimp, minced meat (beef/pork), precut leek, and shredded carrots) confirmed that the mBIBG medium is suitable for quantitative recovery of aeromonads (ca. 10(2)-10(7) cfu/g) in the presence of a high background flora (10(5)-10(6) cfu/g). Screening of naturally contaminated foods (vegetables, seafood, meat) for the presence of Aeromonas resulted in three out of 14 food samples showing presumptive Aeromonas colonies on mBIBG. PMID- 10868683 TI - Evaluation of a monoclonal antibody able to detect live Listeria monocytogenes and Listeria innocua. AB - A monoclonal Listeria antibody, designated B4, was evaluated. The ability of the antibody to bind to viable bacteria belonging to Listeria spp. compared to bacteria of the same species killed by heat treatment, acid or base treatment, sanitizers, and irradiation was examined. The antibody was found to react with viable L. monocytogenes and L. innocua, but not with heat-killed (72 degrees C, 5 min) strains of these organisms. When L. monocytogenes and L. innocua were killed by methods other than heat treatment, it was ambiguous whether the antibody detected the organism or not. It was concluded that the B4 antibody has potential to be used in an immuno capture step to capture live L. monocytogenes and L. innocua from foods prior to identification of L. monocytogenes by polymerase chain reaction (PCR). PMID- 10868684 TI - Evaluation of a Yersinia adhesion gene (yadA) specific PCR for the identification of enteropathogenic Yersinia enterocolitica. AB - A total of 101 Yersinia enterocolitica strains was investigated with a PCR assay [Blais and Phillipe, Food Control, 6 (1995) 211-214] targeting the Yersinia adhesin gene (yadA) responsible for autoagglutination. Compared to the autoagglutination test the PCR assay has a specificity of 100% but a sensitivity of only 70%. This failure might be caused by the sequence heterogeneity of yadA. PMID- 10868685 TI - Patient and tumor characteristics of colon cancers with microsatellite instability: a population-based study. AB - Molecular screening for microsatellite instability (MSI) in colon cancers has been proposed to identify individuals with hereditary nonpolyposis colorectal cancer. To date, most reports of MSI in colorectal cancer have been based on studies of clinical case series or high-risk families. We examined the proportion of incident colon cancers in the general population that exhibit MSI by patient and tumor characteristics. We interviewed 201 colon cancer cases ascertained by the New Mexico Tumor Registry in the metropolitan Albuquerque area for demographic information, lifestyle factors, medical history, and family cancer history. Paired normal and tumor tissue specimens were obtained for each case. Three microsatellite markers were used; instability was defined as observed alteration at two or more loci. Overall, 37 of 201 (18%) colon cancers exhibited instability. MSI was more common among cases >70 years (26%) and most common among cases >80 years (38%). MSI was significantly associated with tumors in the proximal colon and with later stage and poor differentiation among cases >70 years. MSI was not associated with a history of polyps. Family history of colorectal cancer was associated with MSI only among cases <50 years. When all factors were analyzed jointly in a regression model, proximal subsite and poor differentiation remained significantly associated with MSI. One patient, whose tumor exhibited MSI, fulfilled the Amsterdam Criteria for hereditary nonpolyposis colorectal cancer. Our study provides a population-based estimate of MSI in colon tumors and a representative estimate of the proportion of colorectal cancer patients in the general population who consent to be interviewed for family cancer history and to have biological samples analyzed. PMID- 10868686 TI - Aspirin induction of apoptosis in esophageal cancer: a potential for chemoprevention. AB - The potential use of non-steroidal anti-inflammatory drugs (NSAIDs) in the prevention of gastrointestinal cancers has been highlighted recently. However, it is not known whether NSAIDs could also be useful for preventing esophageal cancer, although regular users of these drugs appear to have a decreased incidence of esophageal cancer. Therefore, we examined the effect of aspirin on growth and apoptosis in 10 esophageal cancer cell lines as well as the expression and modulation of its target enzymes, cyclooxygenases (COXs), and their product prostaglandin E2. Growth inhibition of these cells by aspirin was dose- and time dependent and associated with the induction of apoptosis. COX-1 and COX-2 were expressed in 7 of the 10 cell lines. Bile acids could induce COX-2 expression in six of eight cell lines tested, which was correlated with prostaglandin E2 production, and aspirin could inhibit COX-2 enzymatic activity even after bile acid stimulation but was unable to change the COX-2 protein level in these cell lines. Down-regulation of bcl-2 by aspirin was found in the two cell lines tested. These results suggest that induction of apoptosis by aspirin may be a mechanism by which it can intervene in esophageal carcinogenesis and may be indicative of the potential of NSAIDs as chemopreventive agents in esophageal cancer. PMID- 10868687 TI - Impact of genetic polymorphisms in cytochrome P450 2E1 and glutathione S transferases M1, T1, and P1 on susceptibility to esophageal cancer among high risk individuals in China. AB - Esophageal cancer, which is prevalent in China, is believed to be induced by environmental carcinogens such as nitrosamines and other agents. The disproportionate geographical distribution of this cancer among individuals suggests a role for gene-environment interactions in developing the disease. We have shown in our preliminary study that a genetic polymorphism in cytochrome P450 2E1 (CYP2E1) that is known to activate nitrosamines may be a susceptibility factor involved in the early events leading to the development of esophageal cancer (Lin et al., Cancer Epidemiol. Biomark. Prev., 7: 1013-1018, 1998). This relatively larger study was conducted to compare the results with our previous findings. One hundred and fifty cases with esophageal cancer, 146 cases with esophageal dysplasia, and 150 normal controls were residents of Linxian, China, a high-risk area. Genomic DNA samples were assayed for restriction fragment length polymorphisms in the CYP2E1 and GSTP1 loci by PCR amplification followed by digestion with RsaI and Alw26I, respectively. Deletion of the GSTM1 and GSTT1 genes was detected by multiplex PCR. The distribution of CYP2E1 c1/c1 allele frequency was found to be significantly different between controls (44.0%) and cases with cancer (71.3%) or cases with dysplasia (70.6%; P < 0.0001). Individuals having the c1/c1 genotype were at a 3.1-fold [95% confidence interval (CI), 2.4-3.9] increased risk of developing dysplasia and a 3.2-fold (95% CI, 2.5 4.1) increased risk of developing squamous cell carcinoma of the esophagus. Although polymorphisms in the GSTT1 and GSTP1 were not significantly different between cases with cancer or cases with dysplasia and controls, the frequency of the GSTM1 non-null (+/+ and +/0) genotypes appeared to be overrepresented in cases with cancer compared with controls (odds ratio, 2.3; 95% CI, 1.8-3.0). Furthermore, a joint effect of the CYP2E1 c1/c1 genotype and GSTM1 non-null genotype on the cancer risk was observed, showing an odds ratio of 8.5 (95% CI, 3.7-19.9). These results demonstrate that CYP2E1 and perhaps GSTM1 are genetic determinants in the development of squamous cell carcinoma of the esophagus. PMID- 10868688 TI - Genetic polymorphisms of N-acetyltransferases 1 and 2 and gene-gene interaction in the susceptibility to childhood acute lymphoblastic leukemia. AB - Acute lymphoblastic leukemia (ALL) is the most common pediatric cancer. In utero and postnatal exposures to various carcinogens may play a role in the etiology of this disease. N-acetyltransferases, encoded by the NAT1 and NAT2 genes are involved in the biotransformation of aromatic amines present in tobacco smoke, environment, and diet. Their rapid and slow acetylation activity alleles have been shown to modify the risk to a variety of solid tumors in adults. To investigate the role of NAT1 and NAT2 variants as risk-modifying factors in leukemogenesis, we conducted a case-control study on 176 ALL patients and 306 healthy controls of French-Canadian origin. Slow NAT2 acetylation genotype was found to be a significant risk determinant of ALL (odds ratio, 1.5; 95% confidence interval, 1.0-2.2) because of overrepresentation of the alleles NAT2*5C and *7B and underrepresentation of NAT2*4. Besides a slight increase in NAT1*4 allele frequency among cases, no independent association of NAT1 acetylation genotypes and ALL risk was observed. However, the risk associated with NAT2 slow acetylators was more apparent among homozygous individuals for NAT1*4 (odds ratio, 1.9; 95% confidence interval, 1.1-3.4). When NAT2 slow acetylators were considered together with the other risk-elevating genotypes, GSTM1 null and CYP1A1*2A, the risk of ALL increased further, which showed that the combination of these genotypes is more predictive of risk then either of them independently. These findings suggest that leukemogenesis in children is associated with carcinogen metabolism and consequently related to environmental exposures. PMID- 10868689 TI - Glutathione S-transferase polymorphisms in children with myeloid leukemia: a Children's Cancer Group study. AB - GSTM1 and GSTT1 are polymorphic genes. Absence of enzyme activity is due to homozygous inherited deletion of the gene, reducing detoxification of carcinogens such as epoxides and alkylating agents and potentially increasing cancer risk. We hypothesized that GST null genotype would increase risk of acute myeloid leukemia and myelodysplasia (AML/MDS) in children. DNA was extracted from bone marrow slides of 292 AML/MDS patients. PCR amplification was used to assign GSTM1 and GSTT1 genotypes for cases and controls. Given that the frequency of the null genotype varies by ethnicity and that the majority of the cases were Caucasian, analyses were restricted to 232 white (non-Hispanic) cases and 153 Caucasian non cancer controls. The frequency of GSTM1 null was significantly increased in AML/MDS cases compared with controls [64 versus 47%; odds ratio (OR), 2.0 [95% confidence interval (CI), 1.3-3.1]; P = 0.001], whereas the frequency of GSTT1 null genotype in AML/MDS cases was not statistically different from controls. AML comprises biologically distinct subtypes, and a test for homogeneity revealed a statistically significant difference among subtypes (P = 0.04; df, 8) for GSTM1 only. In particular, there was an increased frequency of GSTM1 null genotypes in French-American-British groups M3 [82%; n = 22; OR, 5.1 (95% CI, 1.6-21.3)] and M4 [72%; n = 53; OR, 2.9 (95% CI, 1.4-6.0)]. We conclude that the GSTM1 null genotype is a significant risk factor for childhood AML, particularly French American-British groups M3 and M4. This may indicate an important role for exogenous carcinogens in the etiology of childhood AML. PMID- 10868690 TI - Glutathione S-transferases M1, T1, and P1 and breast cancer. AB - We examined associations for glutathione S-transferases M1 (GSTM1), T1 (GSTT1), and P1 (GSTP1) genotypes and breast cancer in the Carolina Breast Cancer Study, a population-based, case-control study in North Carolina. Odds ratios were close to the null value for each GST locus among African-American women (278 cases and 271 controls) and white women (410 cases and 392 controls), as well as pre- and postmenopausal women. For women with a history of breast cancer in one or more first-degree relatives, odds ratios were 2.1 (95% confidence interval, 1.0-4.2) for GSTM1 null and 1.9 (0.8-4.6) for GSTT1 null genotypes. Among women with a family history, age at diagnosis was significantly earlier for those with the GSTM1 null genotype. We did not observe strong evidence for modification of odds ratios for smoking according to GST genotypes. There was no evidence for combined effects of GSTM1, GSTT1, and GSTP1 genotypes, and there were no combined effects for GST genotypes and the catechol O-methyltransferase genotype. We conclude that GSTM1, GSTT1, and GSTP1 genotypes do not play a strong role in susceptibility to breast cancer. However, the role of GST genotypes in age at onset and risk of breast cancer among women with a family history merits further investigation. PMID- 10868691 TI - A prospective study of estradiol and breast cancer in Japanese women. AB - Few studies have prospectively examined endogenous hormone levels as risk factors for breast cancer. The present study compares prediagnostic hormone levels using stored serum from breast cancer cases and controls selected from the Life Span Study population of the Radiation Effects Research Foundation in Hiroshima and Nagasaki, Japan. Stored serum samples collected in 1968-1970 were assayed for 72 women subsequently diagnosed with breast cancer and 150 control subjects in 72 case-control sets matched on age, date of blood collection, exposure, radiation dose, and city. Serum levels were determined for sex hormone binding globulin, total estradiol (E2), bioavailable E2, dehydroepiandrosterone sulfate, and prolactin. Matched case-control comparisons of hormone levels were carried out by conditional logistic regression and were adjusted for menopausal status at the time of blood drawing. The odds ratio per unit log change in bioavailable E2 was 2.2 [95% confidence interval (CI), 1.02-5.31 for all subjects, and 2.3 (95% CI, 0.55-6.8) and 2.1 (95% CI, 0.55-9.7), respectively, based only on premenopausal or postmenopausal serum. The estimated odds ratios in each quintile of bioavailable E2 level, using the lowest quintile as referent, were 1.00, 1.89, 1.43, 3.45, and 3.37 (P for trend = 0.035). For sex hormone binding globulin, the overall odds ratio was 0.58 (95% CI, 0.14-2.26), and 1.00 (95% CI, 0.19-5.45) and 0.21 (95% CI, 0.02-1.88) based on premenopausal and postmenopausal serum, respectively. This study offers further prospective support for the hypothesis that a high level of biologically available E2 is a risk factor for the subsequent development of breast cancer. PMID- 10868692 TI - Premenopausal equol excretors show plasma hormone profiles associated with lowered risk of breast cancer. AB - Increased urinary excretion of equol, a metabolite of the isoflavone daidzein, has been associated with a reduced risk of breast cancer. This risk reduction has generally been presumed to be a consequence of increased isoflavone consumption. However, only 30-40% of the population excretes more than trace amounts of equol, regardless of isoflavone intake. Accordingly, we hypothesized that the observed apparent protective effect of equol is at least in part attributable to hormonal differences between equol excretors and non-excretors, and that these differences are largely independent of isoflavone intake. We measured plasma hormone and sex hormone binding globulin (SHBG) concentrations in 14 normally cycling premenopausal women during each of three diet periods in which they consumed differing isoflavone doses (0.15, 1.0, and 2.0 mg/kg of body weight/day) as a component of soy protein isolate. The plasma hormone and SHBG concentrations of equol excretors (n = 5) were then compared with those of the non-excretors (n = 9). Results showed that even at the lowest dose, urinary equol excretion values for excretors far exceeded those for non-excretors consuming the highest dose. At all doses, equol excretors generally had lower concentrations of estrone, estrone sulfate, testosterone, androstenedione, dehydroepiandrosterone (DHEA), DHEA sulfate, and cortisol and higher concentrations of SHBG and midluteal progesterone, a hormonal pattern overall consistent with lowered breast cancer risk. In conclusion, the association of equol excretion and lowered breast cancer risk may largely reflect the tendency of equol excretors to have more favorable hormonal profiles, as opposed to merely reflecting increased isoflavone intake. Equol may be a marker for the presence of colonic bacterial enzymatic activity that increases fecal steroid excretion. Alternatively, equol itself, even with very modest isoflavone intake, may exert beneficial effects on the regulation of endogenous hormones. PMID- 10868693 TI - The glutathione S-transferase M1 genotype in ovarian cancer. AB - Glutathione S-transferase mu-1 (GSTM1) is a polymorphic member of the mu class gene family of the glutathione S-transferases. Individuals who are GSTM1 null have increased susceptibility to lung and colon cancer. We hypothesized that: (a) GSTM1 null individuals might also be at increased risk for development of ovarian cancer; and (b) the GSTM1 genotype would influence response to chemotherapy. One hundred and forty-six individuals with invasive epithelial ovarian cancer were genotyped using a three-primer PCR reaction specific for the GSTM1 gene and an internal control glutathione S-transferase mu-4 (GSTM4). The products were analyzed on agarose gels. Healthy individuals without a family history of ovarian, breast, or colon cancer served as unmatched controls (n = 80). The results show that age at diagnosis, histological type, and stage of ovarian cancer were all independent of GSTM1 genotype. The frequency of the GSTM1 null genotype in the ovarian cancer cohort was similar to that in the control population, 51% versus 58%, P > 0.05. Likewise, median survival for individuals with advanced stage ovarian cancer was independent of GSTM1 genotype. We concluded that the GSTM1 null genotype does not increase ovarian cancer risk. These findings suggest that GSTM1 does not play a significant role in detoxifying environmental factors that influence ovarian carcinogenesis and does not play an important role in the resistance of ovarian cancer to chemotherapy. PMID- 10868694 TI - Early-life physical activity and postmenopausal breast cancer: effect of body size and weight change. AB - It is not yet known whether early-life physical activity reduces the risk of developing breast cancer. Subgroup analyses according to menopausal status and body mass may help clarify this association. Data from a population-based case control study of female residents of Wisconsin, Massachusetts, Maine, and New Hampshire were used to examine associations between body mass and breast cancer risk. Cases (n = 4614) were identified by each state's tumor registry; controls (n = 5817) were randomly selected from population lists. Frequency of participation in strenuous physical activity when 14-22 years of age, weight at age 18 and 5 years before interview, height, and other factors were ascertained through structured telephone interviews. Adjusted odds ratios (ORs) and 95% confidence intervals (CIs) were computed using logistic regression. Reductions in postmenopausal breast cancer risk associated with strenuous physical activity were greatest for women in the fourth quartile of body mass index at age 18; the OR for women with the highest activity frequency on average (> or =once/day) was 0.45 (95% CI = 0.26-0.79). Associations with frequency of activity also varied by weight change. Compared to women with no activity and little adult weight gain, frequent physical activity was associated with reduced postmenopausal breast cancer risk in women who had lost weight since age 18 (OR = 0.19, 95% CI = 0.05 0.70) or had gained little or modest amounts of weight (weight gain: first tertile, OR = 0.36, 95% CI = 0.05-0.85; second tertile, OR = 0.31, 95% CI = 0.14 0.66). Weighted MET score analyses yielded similar but less inverse results. These findings suggest that the reduced risk of postmenopausal breast cancer associated with frequent, early-life physical activity may be greatest in women who, over the adult years, either lost weight or gained only modest amounts. PMID- 10868695 TI - Breast and cervical cancer screening practices among Asian and Pacific Islander women in the United States, 1994-1997. AB - Recent studies suggest that Asian and Pacific Islander women in the United States may underuse cancer screening tests. We examined the breast and cervical cancer screening practices of 6048 Asian and Pacific Islander women in 49 states from 1994 through 1997 using data from the Behavioral Risk Factor Surveillance System. About 71.7% [95% confidence interval (CI), 66.3-77.0%] of women in this sample aged > or =50 years had a mammogram in the past 2 years, and 69.5% (95% CI, 63.9 75.1%) had a clinical breast exam in the past 2 years. About 73.7% (95% CI, 71.3 76.0%) of women aged > or =18 years who had not undergone a hysterectomy had a Papanicolaou test in the past 3 years. Women with health insurance and those who had seen a physician in the past year were more likely to have been screened. These results underscore the need for continued efforts to ensure that Asian and Pacific Islander women who are medically underserved, including those without health insurance, have access to cancer screening services. PMID- 10868696 TI - Indoor coal combustion emissions, GSTM1 and GSTT1 genotypes, and lung cancer risk: a case-control study in Xuan Wei, China. AB - The lung cancer mortality rate in Xuan Wei County, China is among the highest in the country and has been associated with exposure to indoor smoky coal emissions that contain high levels of polycyclic aromatic hydrocarbons. This risk may be modified by variation in metabolism genes, including GSTM1, which encodes an enzyme known to detoxify polycyclic aromatic hydrocarbons. To investigate the relationship between GST genotypes and lung cancer risk in Xuan Wei County, we analyzed GSTM1 and GSTT1 genotypes in a population-based case-control study. A total of 122 lung cancer patients and 122 controls, individually matched by age, sex, and home fuel type, were studied. Compared to subjects who used less than 130 tons of smoky coal during their lifetime, heavier users (> or =130 tons) had a 2.4-fold (95% confidence interval, 1.3-4.4) increased risk of lung cancer. The GSTM1-null genotype was associated with a 2.3-fold (95% confidence interval, 1.3 4.2) increased risk of lung cancer. Furthermore, there was some evidence that smoky coal use was more strongly associated with lung cancer risk among GSTM1 null versus GSTM1-positive individuals. In contrast, the GSTT1 genotype was not significantly associated with lung cancer risk. Our data suggest that the GSTM1 null genotype may enhance susceptibility to air pollution from indoor coal combustion emissions. PMID- 10868697 TI - Blood level of B and CD4+ lymphocytes measured before induction of an experimental tumor in rats predicts tumor progression and survival. AB - After an initial series of experiments indicated that early responses of B lymphocytes were important in controlling tumor metastases in two rat models of cancer (N. Quan et al., Cancer Res., 59: 1080-1089, 1999), the present study assessed whether differences in the number of B lymphocytes that are normally present in different individual rats before any tumor development could predict tumor growth, metastases, and length of survival when tumor challenge subsequently occurred. Repeated baseline measures of several circulating lymphocyte subtypes (i.e., natural killer, B, CD4+, CD8+ lymphocytes) were made in individual inbred WAG rats before any introduction of tumor cells, and stable baselines for these subtypes were found. Animals were then injected with 2 x 10(6) CC531 tumor cells (a syngeneic tumor) into the leg, and the size of the resulting primary tumor measured. Primary tumors were surgically removed 6-7 weeks after tumor-cell injection, and animals then followed until death from metastases. In two experiments, the size of the primary tumor as well as the length of time that animals survived correlated with the pretumor percentage of certain lymphocyte subtypes in peripheral blood before tumor-cell injection. Baseline percentage of B lymphocytes was significantly negatively correlated with the size of the primary tumor and was positively correlated with the duration of survival. Baseline percentage of CD4+ lymphocytes showed the opposite relationship, being positively correlated with tumor size and negatively correlated with survival time, although these correlations were lower than those for B lymphocytes. Percent B lymphocytes in circulation also declined during tumor development. In summary, a high percentage of endogenous peripheral blood B lymphocytes predicted growth of smaller primary tumors and longer survival after experimental tumor induction in a rat model, further suggesting that B lymphocytes are involved in protection against development of certain tumors. PMID- 10868698 TI - N-acetyltransferase 2 phenotype but not NAT1*10 genotype affects aminobiphenyl hemoglobin adduct levels. AB - Aminobiphenyls (ABPs) in tobacco have been implicated in bladder cancer etiology in smokers. N-Acetylation of ABPs in the liver, predominantly by the N acetyltransferase 2 (NAT2) isozyme, represents a detoxification pathway, whereas O-acetylation of N-hydroxy-ABPs in the bladder, predominantly by the N acetyltransferase 1 (NAT1) isozyme, represents a bioactivation pathway. We and others have demonstrated that NAT2 phenotype affects 3- and 4-ABP-hemoglobin adduct levels (higher levels in slow acetylators), which are considered valid biomarkers of the internal dose of ABP to the bladder. We have also shown that NAT1 genotype (NAT1*10 allele) is associated with increased DNA adduct levels in urothelial tissue and higher risk of bladder cancer among smokers. It is not known whether NAT1*10 genotype influences ABP-hemoglobin adduct levels. Therefore, we assessed 403 primarily non-Hispanic white residents of Los Angeles County for their NAT2 acetylator phenotype, NAT1*10 acetylator genotype, and 3- and 4-ABP-hemoglobin adduct levels. Eighty-two subjects were current tobacco smokers of varying intensities. Tobacco smokers had significantly higher mean 3- and 4-ABP-hemoglobin adduct levels relative to nonsmokers. The levels increased with increased amounts smoked per day (two-sided, P < 0.0001 in all cases). With adjustment for NAT1 genotype and race, the smoking-adjusted geometric mean level of 3-ABP-hemoglobin adducts in NAT2 slow acetylators was 47% higher than that in NAT2 rapid acetylators (P = 0.01). The comparable value for 4-ABP-hemoglobin adducts was 17% (P = 0.02). In contrast, no association between NAT1*10 genotype and 3- or 4 ABP-hemoglobin adduct levels was observed after adjustment for NAT2 phenotype, smoking, and race. The present study suggests that the impact of the NAT1*10 genotype on 3- and 4-ABP-hemoglobin adducts is noninformative on the possible association between NAT1 activity and bladder cancer risk. PMID- 10868699 TI - Serum ferritin concentration and recurrence of colorectal adenoma. AB - Both body iron stores and dietary iron intake have been reported to increase risk of colorectal neoplasms. We assessed whether serum ferritin concentration was associated with recurrence of colorectal adenomas among 733 individuals with baseline determinations of ferritin as part of a multicenter clinical trial of antioxidant supplements for adenoma prevention. All study participants had at least one adenoma removed within 3 months before enrollment, and 269 of them developed one or more adenomas between follow-up colonoscopies conducted 1 and 4 years after enrollment. Baseline serum ferritin concentrations were analyzed both as a log-transformed continuous variable and as a categorical variable, defined as whether iron stores were nonreplete and low (ferritin < or =30 microg/liter), nonreplete and borderline (31-70 microg/liter), replete and adequate (71-160 microg/liter), or replete and high (>160 microg/liter). Analyses were based on multiple logistic regression models, including age, sex, study center, energy, alcohol, fiber, folate, and total fat intake, number of months between colonoscopic examinations, smoking status, and aspirin use. Overall, there was no statistically significant linear association between log ferritin concentration and adenoma recurrence (P = 0.33). Risk of adenoma recurrence was modestly increased among participants with ferritin concentrations >70 microg/liter relative to those with lower ferritin (odds ratio, 1.39; 95% confidence interval, 0.96-2.02). This result seemed more pronounced among women than men. Dietary intake of iron and red meat was inversely associated with adenoma recurrence among participants with replete iron stores but not consistently associated among those with nonreplete stores. Our findings suggest that any role of iron stores and dietary iron in influencing risk of colorectal adenoma recurrence is likely complex. PMID- 10868700 TI - Genetic and immunohistochemical analyses of p53 independently predict regional metastasis of gastric cancers. AB - Either p53 gene mutation or immunohistochemical detection of p53 protein has not been consistently shown to have prognostic significance in human cancers, including gastric carcinomas. One hypothesis to explain this inconsistency is that some p53 mutations and p53 protein accumulation are not indicative of tumor progression. To test this hypothesis, we categorized p53 status in 105 gastric carcinomas according to types of mutations, numerical scores of immunohistochemical staining (IHC), or combinations thereof. The p53 status was then correlated with metastasis to liver or peritoneum. Gastric cancers with no p53 mutations were significantly less likely to metastasize than tumors with mutations. Intermediate IHC scores were inversely associated with metastasis. A substantial number of gastric cancers (31 of 105) showed positive p53 immunostaining without detectable mutations (p53-/IHC+), which suggested an accumulation of wild-type p53 protein, and also a significantly lower risk for metastasis. After adjusting for depth of invasion and lymph node involvement, the p53-/IHC+ combination predicted low metastatic risk better than either p53- or IHC+ with intermediate scores. These findings suggest that an accumulation of wild-type p53 protein occurs in gastric cancer cells and represents a stress response mechanism that lowers metastatic potential. PMID- 10868701 TI - Reviewing asthma and anxiety. AB - The main characteristic of asthma is sudden and unexpected attacks of impaired breathing. Both the attacks themselves and the prospect of attacks generate much anxiety amongst patients. Several different forms of anxiety can be identified which vary in intensity and the situations in which they appear. Anxiety disorders are more common in asthmatics and have a considerable influence on asthma management because they influence symptom perception. Excessive anxiety about asthma symptoms can affect the patient's response to an asthma attack; anxiety related to asthma triggers can reduce the patient's quality of life and anxiety related to medical treatment can influence compliance. The extent of this influence depends upon an individual's ability to cope. Behavioural therapeutic programmes for patient education offer an opportunity to reduce anxiety and to improve asthma self-management. Physicians should look carefully for anxiety when taking the patient's history, and should support the patient's participation in asthma education programs. PMID- 10868702 TI - Inhaled glucocorticosteroids decrease hydrogen peroxide level in expired air condensate in asthmatic patients. AB - H2O2 is elevated in the exhaled air condensate in several inflammatory disorders of the lung, including bronchial asthma, and thus may reflect inflammatory processes in the airways. Exhaled H2O2 may be used to guide the anti-inflammatory treatment of patients with asthma. Therefore in this study we analysed the effect of inhaled glucocorticosteroid beclomethasone for 4 weeks on H2O2 level in the exhaled air condensate. Seventeen asthmatics and 10 healthy subjects were included to the study. Eleven patients were given inhaled beclomethasone and six were given placebo (3M Health Care). In all patients pulmonary function tests were performed. H2O2 in the expired air condensate was measured spectrofluorimetically (homovanillic acid method). Inhaled beclomethasone significantly decreased H2O2 in the expired air condensate in the active treatment group, with a fall from baseline on day 1 which remained on day 43 (follow-up) (P<0.05). Exhaled H2O2 in the active-treatment group was significantly lower than that in placebo group (P<0.05). A negative correlation between H2O2 and forced expiratory volume in 1 sec (FEV1) on day 29 was observed. The decrease in exhaled H2O2 in the active-treatment group was accompanied by an improvement in pulmonary function tests results. Inhaled glucocorticoids reduce the level of H2O2 in the expired air condensate of asthmatic patients over a 4 week period and this may reflect their anti-inflammatory activity in lung diseases. PMID- 10868703 TI - Risk factors for community-acquired pneumonia diagnosed by general practitioners in the community. AB - The purpose of this study was to identify risk factors for pneumonia diagnosed in the community by general practitioners, using a case control study in 29 general practices in Nottingham, U.K. Patients with radiographically confirmed pneumonia were compared with adults randomly selected from electoral registers corresponding to the catchment areas of the general practices taking part in the study. Sixty-six cases and 489 controls participated. Significant risk factors in univariate analysis included age, chronic obstructive pulmonary disease, congestive heart failure and lifetime consumption of cigarettes. Multiple logistic regression analysis of these four variables showed that age [adjusted odds ratio = 2.69 (for 30 year increment), 95%CI = 1.66-4.35] and chronic obstructive pulmonary disease (adjusted odds ratio= 1.99, 95%CI = 1.15-3.45) were independent risk factors. Only age and chronic obstructive pulmonary disease were independent risk factors for pneumonia in this study. Since cigarette smoking is the major cause of chronic obstructive pulmonary disease, these data suggest that cigarette smoking is the main avoidable risk factor for community-acquired pneumonia in adults. PMID- 10868704 TI - Inhibition of GM-CSF secretion by topical corticosteroids and nedocromil sodium. A comparison study using nasal polyp epithelial cells. AB - Nasal epithelial cells maintain eosinophil survival by secreting granulocyte/macrophage colony-stimulating factor (GM-CSF). Corticosteroids antagonize eosinophil viability induced by GM-CSF. We investigated the effect of topical corticosteroids and nedocromil sodium on the release of GM-CSF from nasal polyp epithelial cells. Epithelial cells were obtained from 19 patients undergoing nasal polypectomy and cultured. After reaching confluence, cultured cells were stimulated with 10% foetal calf serum in the absence and presence of four topical corticosteroids and nedocromil sodium for 48 h. GM-CSF was measured by enzyme linked immunosorbent assay (ELISA). Fluticasone propionate was the most potent inhibitor of GM-CSF release (IC25 = 46 pM) closely followed by budesonide (IC25 = 4 nM), beclomethasone dipropionate (IC25 = 40 nM) and triamcinolone acetonide (IC25 = 75 nM). Nedocromil sodium had no effect on GM-CSF release. We conclude that the effect of topical steroids on reducing eosinophil infiltration in nasal polyps may be due in part to downregulation, among other cytokines, of epithelial GM-CSF production which prolongs eosinophil viability. Quantitatively, fluticasone propionate inhibited GM-CSF production more potently than budesonide, beclomethasone dipropionate and triamcinolone acetonide. PMID- 10868705 TI - Use of pleural fluid C-reactive protein in diagnosis of pleural effusions. AB - The aims of the study were to assess whether C-reactive protein (CRP) is a sensitive marker for discriminating between transudative and exudative and pleural effusions to evaluate whether it can be used to distinguish inflammatory pleural effusions from other types of effusion. Pleural fluid and serum CRP levels were obtained in 97 patients with pleural effusion, using an immunoturbidimetric method (Olympus AU-600 autoanalyser). We compared CRP levels between transudates and exudates, inflammatory effusions and other types of effusion. According to the criteria used, 16 patients were included in the transudate group and 81 patients in the exudate group. Pleural fluid CRP levels were significantly lower in the transudate group (P<0.04; 14.9 +/- 4.9 mg l(-1) and 35.5 +/- 4.9 mg l(-1) respectively). Also, the ratio of pleural fluid to serum was significantly lower in the transudate group (P<0.009; 0.8 +/- 0.5 mg l( 1) and 2.8 +/- 0.7 mg l(-1), respectively). In the exudate group, 35 patients had neoplastic effusions, 10 chronic non-specific pleurisy, 19 tuberculous pleurisy, 16 parapneumonic effusion and one Dressler Syndrome. When these sub-groups were compared, the parapneumonic effusion subgroup CRP levels (mean 89 +/- 16.3 mg l( 1)) were significantly higher than those in the other subgroups, other exudate of neoplastic effusion, tuberculous pleurisy and chronic non-specific effusion and the transudate group (P<0.0001; P<0.0001; P<0.0004 and P<0.0001, respectively). The ratio between pleural fluid and serum CRP was significantly higher in the parapneumonic effusion subgroup than in the neoplastic subgroup (P<0.0002; 6.6 +/ 2.7 mg l(-1) and 1 +/- 0.2 mg l(-1), respectively). Pleural fluid CRP levels > 30 mg l(-1) had a high sensitivity (93.7%) and specificity (76.5%) and a positive predictive value of 98.4%. In the differential diagnosis of pleural effusions, higher CRP levels may prove to be a rapid, practical and accurate method of differentiating parapneumonic effusions from other exudate types. Although the high level of CRP obtained in the exudate group may be due to the number of patients with parapneumonic effusion who were included, the pleural CRP level may also be helpful in discriminating between exudative and transudative pleural effusions. PMID- 10868706 TI - Influence of clinical history on airways bacterial colonization in subjects with chronic tracheostomy. AB - Patients with chronic tracheostomy are subject to significant bacterial colonization of the airways, a risk factor for respiratory infections. The aim of our study was to verify whether bacterial colonization and humoral immune response in the airways can be influenced by the disease which led to chronic respiratory failure and tracheostomy. Thirty-nine clinically stable outpatients with chronic tracheostomy were considered: 24 were affected by chronic obstructive pulmonary disease (COPD) (mean age 66 years, range 54-78, M/F 19/3; months since tracheostomy 23, range 3-62), 15 by restrictive lung disease (RLD) (12 thoracic wall deformities, three neuromuscular disease; age 57 years, range 41-72; M/F 3/12, months since tracheostomy 22, range 2-68). Recent antibiotic or corticosteroid treatments (< 1 month) were among exclusion criteria. Bacterial counts were assessed in tracheobronchial secretions with the method of serial dilutions. Identification of bacterial strains was performed by routine methods. Albumin, IgG, A, and M were measured in airways secretions with an immunoturbidimetric method. No significant differences were found between the two groups as regards either the quantitative bacterial cultures (RLD 81.4, 2.6-4200 x 10(4); COPD 75.9, 1.0-1530 x 10(4) colony forming units (cfu)/ml, geometric mean, range) or the prevalence of the main bacterial strains, (Pseudomonas species: 38 and 37%, Serratia marcescens: 31 and 23%, Staphylococcus aureus: 14 and 6%, Proteus species: 3 and 8%, for RLD and COPD respectively) as a percentage of total strains isolated (RLD = 26, COPD = 48). Immunoglobulin levels did not show significant differences, apart from being higher in underweight subjects. We conclude that in our series of stable outpatients with chronic tracheostomy, bacteria-host interaction in the airways was not influenced by the clinical history. PMID- 10868707 TI - Reproducibility of early and late asthmatic responses to allergen challenge in a large group of asthmatics. AB - The specific bronchial provocative test (sBPT) coupled with allergen is used to investigate asthma. Very few studies have examined the reproducibility of responses to allergen challenge. The aim of this study was to measure the reproducibility of PD20FEV1 allergen and late asthmatic response (LAR) in 53 asthmatics and to relate the reproducibility to the time interval between two allergen challenges. Fifty-three atopic asthmatics performed two allergen challenges not less than 2 and not more than 26 weeks apart. Randomly, 19 subjects were assigned to a short-interval group (14-35 days between the two tests) and 34 to a long-interval group (40-180 days). In each challenge, the PD20FEV1 was sought for and the maximum % fall in FEV1 from 3 to 7 h after the allergen challenge was evaluated as a measurement of magnitude of the LAR. High intraclass correlation coefficients (R(I)) were found for both PD20FEV1 (R(I) = 0.78) and LAR (R(I) = 0.77) in all subjects. PD20FEV1 allergen showed a high R(I) in the long-interval group (R(I) = 0.80), but a low R(I) in the short-interval group (R(I) = 0.63). In contrast LAR showed a lower R(I) in the long-interval group (R(I) = 0.68) than in the short-interval group (R(I) = 0.77). Moreover, the R(I) for PD20FEV1 was particularly low in subjects with a dual pattern to the allergen challenge and a short interval between the two allergen challenges. Our study confirmed that asthmatic responses induced by allergen challenge have a good reproducibility. Moreover, we have demonstrated that the interval between two allergen challenges can determine a change in reproducibility in asthmatic responses induced by allergen challenge. PMID- 10868708 TI - Comparison of Pulmicort pMDI plus Nebuhaler and Pulmicort Turbuhaler in asthmatic patients with dysphonia. AB - BACKGROUND: Dysphonia is a known local adverse effect of inhaled corticosteroids. This symptom was investigated by laryngoscopy and assessment in a voice laboratory. The effects of changing the treatment of patients with dysphonia, reported whilst using the pMDI, to pMDI plus Nebuhaler or Tubuhaler was also assessed. METHODS: Seventy-two patients reporting dysphonia and taking inhaled steroids from a pMDI entered a 12-week, open, parallel group study. Fifty-one completed the study per protocol; 26 in the Nebuhaler group [21 female, mean age 57 years (22-77)] and 25 in the Turbuhaler group [18 female, mean age 58 years (21-81)]. A dysphonia diary card was completed weekly. Voice laboratory assessments and laryngoscopy were performed on entry and at 12 weeks. RESULTS: There were no differences in voice laboratory data, laryngoscopic evidence of disordered glottic closure and diary data between the two groups at 12 weeks. At study entry laryngoscopic appearances were normal in almost half the patients. Vocal cord bowing was rarely seen. Glottic closure changed in nine patients during the study period, but there was no correlation with voice symptoms. The trend of symptomatic improvement of voice status in the Turbuhaler group did not correlate with voice laboratory assessments and laryngoscopic evidence of disordered glottic closure. After 4 weeks, 40% of patients using Turbuhaler and 8% in the Nebuhaler group scored their voice status as better (P < 0.02) but there was no significant difference between the two groups at 12 weeks (Turbuhaler 52%, Nebuhaler 23%, P=0.08). CONCLUSION: This study does not support the view that dysphonia in asthmatics inhaling corticosteroids is usually caused by myopathic bowing of the vocal cord muscles. PMID- 10868709 TI - Asthma from childhood to adulthood: asthma severity, allergies, sensitization, living conditions, gender influence and social consequences. AB - The course of asthma severity, clinical allergies, allergic sensitization, changes in living conditions and social outcome were studied prospectively over five follow-up visits from the mean age of 9 to 30 years in a cohort of 28 boys and 27 girls, selected randomly among asthmatic children attending a paediatric outpatient unit. Asthma severity improved from childhood to adulthood, judged by symptom and medication scores and by the number of hospital admissions, but only nine subjects (16%) had been free from symptoms and medication over the last year of follow-up. After adolescence, asthma continued to improve among the males but not among the females. This difference could not be explained by gender differences in the course of clinical allergies or sensitization (skin-prick tests and RAST) to common inhaled allergens, or by differences in environmental or social conditions. Sensitization to relevant perennial inhaled allergens correlated with asthma severity during adulthood. In general, clinical allergies and sensitization to inhaled allergens adopted during childhood persisted into adulthood. Approximately 10% of the subjects never adopted a clinical allergy or a positive allergy test. The social outcome was good. PMID- 10868710 TI - Asthma from childhood to adulthood: course and outcome of lung function. AB - Lung function (FEV1 before and after bronchodilatation) was studied prospectively over five visits in 55 asthmatic children (28 boys) from childhood to adulthood (age 30). At the last follow-up recordings were made at rest, after cold air challenge (CACh), and after bronchodilatation. Results were related to clinical asthma scoring and to sensitization to furred animals, as described in a companion paper. Lung function outcome was shown to be influenced by initial FEV1 (% predicted) and gender, but not by initial asthma severity or sensitization. FEV1 (% predicted) was higher in females than in males over the first two follow ups, but the reverse was found over the subsequent visits. It deteriorated from childhood to adulthood in the females but improved in the males. In adulthood the females (for height 170 cm) had a steeper normalized annual fall in post bronchodilator FEV1 than the males (55 +/- 38 vs. 25 +/- 36 ml; P = 0.006). The degree of bronchial hyperresponsiveness was associated significantly with asthma severity and the extent of sensitization to furred animals, but not with gender. The results indicate a better lung function outcome for asthmatic boys than for girls, confirming trends seen in clinical asthma severity. In adulthood the extent of sensitization to relevant perennial inhaled allergens significantly influences airway responsiveness and asthma severity, but not lung function. PMID- 10868711 TI - Circulating bronchoepithelial cells expressing mRNA for surfactant protein A in patients with pulmonary fibrosis. AB - There are several unsolved clinical findings in patients with idiopathic pulmonary fibrosis (IPF); (i) predominance of fibrosis in the lower lung fields, (ii) digital clubbing, and (iii) patchy distribution of pulmonary fibrosis. To explain these unsolved problems, we hypothesized that regenerated or premature bronchoepithelial cells may circulate in the blood in patients with IPF. To prove this, we performed the reverse transcriptase-polymerase chain reaction (RT-PCR) for cytokeratin 19 (CK19) and pulmonary surfactant protein A (SPA) in peripheral blood in patients with IPF and pulmonary fibrosis associated with collagen vascular disorders. In addition, 20 patients with chronic pulmonary emphysema as a disease control and 19 normal volunteers were also evaluated for the existence of circulating bronchoepithelial cells. RT-PCR analysis showed that CK19 was expressed in 12 of 38 blood samples (31.6%) of IPF and pulmonary fibrosis associated with collagen vascular disorders, seven of 20 (35.0%) blood samples of chronic pulmonary emphysema, and four of 19 (21.1%) blood samples of normal volunteers. mRNA for SPA was positive in eight of 38 (21.1%) blood samples of IPF. In contrast, SPA expressing cells were not detected in any blood samples obtained from patients with chronic pulmonary emphysema or normal volunteers. This evidence suggests that there were some circulating bronchoepithelial cells expressing mRNA for SPA in peripheral blood of patients with IPF and pulmonary fibrosis associated with collagen vascular disorders. PMID- 10868712 TI - Comparable effects of inhaled fluticasone propionate and budesonide on the HPA axis in adult asthmatic patients. AB - This randomized, double-blind, double-dummy, multicentre cross-over study compared the effects on the hypothalamic-pituitary-adrenal (HPA) axis of fluticasone propionate (750 microg twice daily given via the Diskus) and budesonide (800 microg twice daily given via the Turbuhaler). Two treatment periods of 2 weeks each were preceded by a 2-week run-in period and separated by a 2-week washout period. During run-in and washout, patients received beclomethasone dipropionate (BDP) or budesonide at a constant dose of 1500-1600 microg day(-1). Sixty patients aged 18-75 years with moderate to severe asthma not fully controlled by treatment with 1500-1600 microg day(-1) budesonide or BDP entered run-in and 45 completed the study. HPA axis suppression was assessed by morning serum cortisol (area under the curve from 08.00 to 10.30 hours) and 12-h nocturnal urinary cortisol excretion, measured at the end of run-in (baseline 1), at the end of washout (baseline 2), and at the end of each treatment period. Neither budesonide nor fluticasone produced significant suppression of either parameter compared to baselines. Only a few patients had serum-cortisol and urinary cortisol values below the normal range, before and after treatment. This shows that the patients did not have adrenal suppression before entering the study. The ratio between the AUC serum cortisol measured after fluticasone treatment and after budesonide treatment was 0.99 (95% CI 0.92-1.06), indicating equivalent effects on the HPA axis. This result was achieved after having omitted two patients' results, due to their very sensitive reaction to budesonide, but not to fluticasone. Two exacerbations of acute asthma occurred during budesonide treatment and none during fluticasone treatment. Both treatments were well tolerated. In conclusion, budesonide 1600 microg day(-1) via Turbuhaler and fluticasone propionate 1500 microg day(-1) via Diskus had no clinical effects on the HPA axis in patients with moderate to severe asthma. PMID- 10868713 TI - Improved delivery of ipratropium bromide using Respimat (a new soft mist inhaler) compared with a conventional metered dose inhaler: cumulative dose response study in patients with COPD. AB - Respimat (RMT) is a reusable, propellant-free, soft mist inhaler (SMI), a novel device for inhalation therapy. We conducted a three-period cross-over study to evaluate the safety and efficacy of cumulative doses of ipratropium bromide inhaled from RMT (Two dose levels) or from a pressurized metered dose inhaler (MDI), in 36 patients with chronic obstructive pulmonary disease (COPD). The bronchodilator effect of ipratropium bromide was greater when administered via RMT (10 or 20 microg per puff, given double-blind within device, to total doses of 160 or 320 microg) than via MDI (20 microg per puff, total dose 320 microg). The bronchodilator effects of the 160 and 320 microg doses delivered via RMT were similar. Cumulative ipratropium bromide doses of 320 microg given via MDI or RMT and 160 microg given via RMT produced similar safety profiles. Between 45 min after the first drug inhalation and 45 min after the final dose, greater bronchodilatory effect was obtained from half the cumulative dose of ipratropium (RMT 10 microg per puff) when compared with the MDI (20 microg per puff). Therefore, ipratropium bromide delivered by RMT is as safe as, and can be more effective than, the MDI on acute administration in patients with COPD. PMID- 10868714 TI - Inappropriate inhaler use: assessment of use and patient preference of seven inhalation devices. EDICI. AB - Inefficient inhaler technique is a common problem resulting in poor drug delivery, decreased disease control and increased inhaler use. The aim of this study was to assess patients' use of different inhaler devices and to ascertain whether patient preference is indicative of ease of use and whether current inhaler use has any influence on either technique or preference. We also wished to define the most appropriate method of selecting an inhaler for a patient, taking into account observed technique and device cost. One hundred patients received instruction, in randomized order, in the use of seven different inhaler devices. After instruction they were graded (using predetermined criteria) in their inhaler technique. After assessment patients were asked which three inhalers they most preferred and which, if any, they currently used. Technique was best using the breath-actuated inhalers; the Easi-Breathe and Autohaler, with 91% seen to have good technique. The pressurized metered dose inhaler (pMDI) fared poorly, in last position with only 79% of patients showing good technique, despite being the most commonly prescribed. The Easi-Breathe was by far the most popular device with the patients. The Autohaler came in second position closely followed by the Clickhaler and Accuhaler. The majority of patients (55%) currently used the pMDI but the pMDI did not score highly for preference or achieve better grades than the other devices. Only 79% of patients tested could use the pMDI effectively even after expert instruction yet it continues to be commonly prescribed. This has important repercussions for drug delivery and hence disease control. Prescribing a patient's preferred device increases cost but can improve efficiency and therefore be cost effective in the long term. Using an inexpensive device (pMDI) when technique is good and the patient's preferred inhaler device when not is one way to optimize delivery and may even reduce cost. PMID- 10868715 TI - Characterization of the inspiratory manoeuvre when asthmatics inhale through a Turbohaler pre- and post-counselling in a community pharmacy. AB - Dose emission from a Turbohaler has been shown to be dependent on the rate of inhalation, with an optimal flow of 60 l min(-1) recommended. Some patients may need counselling to achieve this fast inhalation. Inhalation rate profiles of 24 asthmatics were measured when they inhaled through a placebo Turbohaler. The setting was a community pharmacy when the asthmatics came to collect their next supply of medication. Profiles were measured before and after counselling on how to use the Turbohaler. The mean (SD) peak inhalation rate through the Turbohaler pre- and post-counselling was 48.0 (16.8) and 54.7 (17.6) l min(-1), and their inspiratory volume was 1.75 (0.68) and 1.94 (0.62) l, respectively. Their mean (SD) percent predicted FEV1 was 57.0 (18.9)%. After counselling, 12 patients achieved an inhalation rate of > 60 l min(-1) and a further four obtained > 55 l min(-1). Emphasis should be placed on counselling patients prescribed all types of inhaled devices rather than concentrating on metered dose inhalers. PMID- 10868716 TI - Community-acquired pneumonia: development of a bedside predictive model and scoring system to identify the aetiology. AB - Although initial presentation has been commonly used to select empirical therapy in patients with community-acquired pneumonia (CAP), few studies have provided a quantitative estimation of its value. The objective of this study was to analyse whether a combination of basic clinical and laboratory information performed at bedside can accurately predict the aetiology of pneumonia. A prospective study was developed among patients admitted to the Emergency Department University Hospital Arnau de Vilanova, Lleida, Spain, with CAP. Informed consent was obtained from patients in the study. At entry, basic clinical (age, comorbidity, symptoms and physical findings) and laboratory (white blood cell count) information commonly used by clinicians in the management of respiratory infections, was recorded. According to microbiological results, patients were assigned to the following categories: bacterial (Streptococcus pneumoniae and other pyogenic bacteria), virus-like (Mycoplasma pneumoniae, Chlamydia spp and virus) and unknown pneumonia. A scoring system to identify the aetiology was derived from the odds ratio (OR) assigned to independent variables, adjusted by a logistic regression model. The accuracy of the prediction rule was tested by using receiver operating characteristic curves. One hundred and three consecutive patients were classified as having virus-like (48), bacterial (37) and unknown (18) pneumonia, respectively. Independent predictors related to bacterial pneumonia were an acute onset of symptoms (OR 31; 95% CI, 6-150), age greater than 65 or comorbidity (OR 6.9; 95% CI, 2-23), and leukocytosis or leukopenia (OR 2; 95% CI, 0.6-7). The sensitivity and specificity of the scoring system to identify patients with bacterial pneumonia were 89% and 94%, respectively. The prediction rule developed from these three variables classified the aetiology of pneumonia with a ROC curve area of 0.84. Proper use of basic clinical and laboratory information is useful to identify the aetiology of CAP. The prediction rule may help clinicians to choose initial antibiotic therapy. PMID- 10868717 TI - Scottish national bronchoscopy audit: a prospective multicentre study of 3316 cases against agreed standards. Scottish Thoracic Society. AB - INTRODUCTION: Bronchoscopy guidelines address issues of patient and operator safety but do not give guidance on the expected yield of the procedure. Realistic standards for several outcome measures of bronchoscopy for investigating bronchial carcinoma have been derived by Scottish clinicians from a published national study. The present study describes the use of these agreed standards in prospective audit. METHODS: All Society members in Scotland (population 5.1 million) were invited to participate. Data were collected for 1 year and coded anonymously. STANDARDS: 1. Supervising bronchoscopist to have completed at least 100 procedures; 2. histology to be positive in 80% of cases where tumour seen; 3. 35%-55% of bronchoscopies to reveal a diagnosis; 4. 60% of patients admitted for bronchoscopy to be day cases; 5. 80% of day case patients to be in hospital for less than 6 h; 6. 90% of male patients and 80% of female patients willing to have repeat bronchoscopy. RESULTS: Three thousand, three hundred and sixteen bronchoscopies were performed by 45 senior pulmonologists at 22 centres. One centre reached all the standards and five centres met five standards. There was wide national variation in histological spectrum, incidence of small cell cancer ranged from 12% to 25% between centres. Participants found their own data helpful in identifying local areas for improvement. CONCLUSION: Bronchcoscopy standards set locally by practising pulmonologists can be used in collaborative audit to identify areas for improving practice. Variation in histology may be accounted for by case-mix or pathology techniques. PMID- 10868718 TI - A comparison of two approaches in the treatment of perceptual problems after stroke. AB - OBJECTIVE: To compare the effectiveness of the transfer of training and functional approaches in improving perceptual and functional abilities after stroke. DESIGN: Patients identified as having perceptual problems were randomly allocated to either the transfer of training approach or the functional approach for perceptual treatment. On completion of six weeks of treatment, each patient was reassessed for perceptual and functional abilities. SUBJECTS AND SETTING: Eighty inpatients on the Nottingham Stroke Unit. INTERVENTIONS: Perceptual treatment was given for 2.5 hours per week for six weeks. MAIN OUTCOME MEASURES: Rivermead Perceptual Assessment Battery, Barthel ADL Index and Edmans ADL index. RESULTS: There was no significant difference between the treatment groups on patient characteristics or impairments. The results also showed no significant difference between the treatment groups before and after treatment on perceptual ability total scores, individual perceptual subtest scores, or functional ability total scores (Mann-Whitney U 642.5-798.0, p > 0.05). Wilcoxon matched pairs signed ranks tests showed a significant improvement in both groups after treatment on perceptual and functional abilities (perceptual z = 6.02, p < 0.001, functional z = 6.72, p < 0.001). CONCLUSIONS: These results indicated that the improvement in perceptual abilities was equivalent using either of the two approaches. This could be due to spontaneous recovery or the effects of the Stroke Unit. PMID- 10868719 TI - Improvement in hand function and wrist range of motion following electrical stimulation of wrist extensor muscles in an adult with cerebral palsy. PMID- 10868720 TI - The efficacy of Le Bon Depart and Sensory Integration treatment for children with developmental coordination disorder: a randomized study with six single cases. AB - OBJECTIVE: Evaluation of the efficacy of Le Bon Depart (LBD) treatment and Sensory Integration (SI) treatment on motor performance of children with developmental coordination disorder. DESIGN: A single subject design with multiple baseline and alternating treatments. Order of treatment and length of phase were randomized. Measurements were blinded. SETTING: Department of Occupational Therapy at the Academic Hospital Vrije Universiteit Amsterdam, The Netherlands. SUBJECTS: Five boys and one girl with developmental coordination disorder (age: 6.0-8.1 years). INTERVENTIONS: Baseline condition, Le Bon Depart treatment and Sensory Integration treatment. MAIN OUTCOME MEASURES: The Movement ABC, Praxis Tests, a rhythm test and visual analogue scales. With the exception of the Praxis Tests, lower scores indicate better performance. RESULTS: During both treatments, the performance on the Movement ABC (x = 7.21) and the scores on the visual analogue scales (x = 46.64) were significantly better than in the baseline (Movement ABC(baseline): x = 17.38; visual analogue scales(baseline): x = 68.18). After treatment 2, performance on the Praxis Tests and scores on the visual analogue scales were significantly better than after treatment 1 (Praxis Tests: 113.54 versus 104.68; visual analogue scales: 34.74 versus 58.54). All six children performed better on the Movement ABC during treatment as compared to the baseline. Le Bon Depart led to significant improvement on all dependent variables, Sensory Integration on the visual analogue scales only. The improvements after Le Bon Depart were larger than the improvements after Sensory Integration treatment. On the rhythm test this difference was significant: LBD led to an improvement of 43.01 points, while the improvement after SI was 17.59 points (p < 0.05). CONCLUSION: Motor performance of children with developmental coordination disorder improved significantly on all dependent variables after the combination of treatments. Le Bon Depart led to more improvement than Sensory Integration. LBD appears to be a valuable treatment method for children with developmental coordination disorder. PMID- 10868721 TI - The Frontal Lobe Score: part I: construction of a mental status of frontal systems. AB - OBJECTIVE: To develop a bedside mental status examination to assess the behavioural effects of damage to the frontal lobes. DESIGN: A prospective clinical comparison of patients with cerebral lesions of different locations. SUBJECTS: A total of 118 subjects were examined: 27 patients with cerebral lesions confined to the frontal lobes, 25 patients with cerebral lesions without involvement of the frontal lobes, 18 patients with mixed frontal/nonfrontal lesions, and 48 normal control subjects. MEASURES: Twenty-three mental status tests, clinical examinations and rating scales that had been reported as indicative of frontal lobe function were brought together. By statistical analysis, 12 tests and a neurobehavioural rating scale were selected. These constitute the Frontal Lobe Score (FLS). RESULTS: The FLS detected pure frontal lesions with a sensitivity of 77.7%. It discriminated patients with frontal lesions from normal control subjects with a specificity of 100%. Differentiation from patients with nonfrontal lesions was obtained with a specificity of 84%. CONCLUSION: The Frontal Lobe Score is a useful screening instrument for the clinical detection of effects of frontal lobe damage. PMID- 10868722 TI - The Frontal Lobe Score: part II: evaluation of its clinical validity. AB - OBJECTIVE: To evaluate the ability of the Frontal Lobe Score (FLS) to differentiate patients with frontal lobe lesions from those with nonfrontal lesions and normal controls. DESIGN: In a prospective, blind setup, the sensitivity and specificity of the Frontal Lobe Score was compared with the Wisconsin Card Sorting Test (WCST) and the Stroop Test. PATIENTS: A sample of 108 subjects (26 patients with cerebral lesions confined to the frontal lobes, 28 patients with cerebral lesions without involvement of the frontal lobes, 31 patients with mixed frontal/nonfrontal lesions, 23 controls without cerebral lesions) was examined. MEASURES: Frontal Lobe Score, Wisconsin Card Sorting Test, Stroop Test. RESULTS: The Frontal Lobe Score detected pure frontal lesions with a sensitivity of 92.3%. It discriminated patients with frontal lesions from normal controls with a specificity of 100%; differentiation from patients with nonfrontal lesions was obtained with a specificity of 75.0%. For the WCST, sensitivity for detection of pure frontal lesions was 65.4%, while specificity was 60.9% compared with normal controls and 53.6% compared with nonfrontal lesions. The Stroop Test showed a sensitivity of 30.8%, a specificity compared with normal controls of 95.7% and compared with nonfrontal lesions of 92.9%. CONCLUSION: The Frontal Lobe Score has clinical usefulness for screening of effects of frontal lobe damage superior to that of the WCST and the Stroop Test. PMID- 10868723 TI - Change in identity and self-concept: a new theoretical approach to recovery following a stroke. AB - OBJECTIVE: To determine whether respondents reported a change in identity following stroke. DESIGN: A cross-sectional comparison study of perceived changes in the self-concept of stroke respondents and matched hospital volunteers. A questionnaire was administered to stroke respondents in their own homes and to hospital volunteers in their work setting. PARTICIPANTS: Twenty-six first-time stroke survivors who had no severe communication, cognitive or perceptual difficulties or previous physical disability, and who had returned home from hospital up to two years previously. The comparison group were 26 hospital volunteers matched for age, gender and the time from which past self-concept was considered. MAIN OUTCOME MEASURES: Included the Hospital Anxiety and Depression Scale, Frenchay Activity Index and the Head Injury Semantic Differential Scale to assess past and present self-concept. RESULTS: Overall, individuals described themselves in more negative terms than prior to their stroke. They saw themselves as less interested, capable and independent, (p < 0.001) and less in control, satisfied and active (p < 0.05). They still saw themselves as friendly, calm, caring, hopeful and talkative. The overall self-concept of the comparison group remained positive and stable over a comparable time period. CONCLUSIONS: The stroke respondents reported a negative sense of self, reduced social activity and psychological morbidity despite inpatient and outpatient rehabilitation. Individuals following stroke may settle for a restricted future because of their expectations of life with a disability. Clinicians need to be aware of the meaning of the stroke within the life of each individual. PMID- 10868724 TI - The Subjective Index of Physical and Social Outcome (SIPSO): a new measure for use with stroke patients. AB - OBJECTIVE: To develop a measure of social integration following stroke. DESIGN: Question and response scale generation from qualitative interview-based work followed by item and factor analytic methods of test construction. Analysis of the psychometric properties of final index. MEASURES: Frenchay Activities Index, Nottingham Health Profile, Wakefield Depression Inventory, Barthel Index. SETTING: Community setting, Bath, UK. SUBJECTS: Two hundred and sixty survivors of stroke, discharged at least six months previously from a general hospital. RESULTS: A 10-item Subjective Index of Physical and Social Outcome (SIPSO) was developed. Each question is scored on a scale of 0-4 with a low score indicating a poor level of integration. From the 157 completed questionnaires, total scores ranged from 0 to 40 with a median of 24 (interquartile range 15-32). Initial testing of the psychometric properties of the SIPSO suggest that it is able to provide assessment of two distinct areas of patient integration. From analysis of completed questionnaires it is suggested that questions 1-5 on the SIPSO measure a factor related to physical functioning/mobility whilst questions 6-10 measure a factor related to social/emotional functioning. Internal consistency, test-retest reliability and construct validity were established. CONCLUSIONS: The SIPSO provides a brief, valid and reliable assessment of an individual's ability to reintegrate to a 'normal' lifestyle. The SIPSO differs from other measures in that it provides assessment of both quantity and quality of activities and interaction, reflecting an individual's ability to reintegrate to his/her own satisfaction. As a 10-item self-report questionnaire the SIPSO can be administered quickly and cheaply to large numbers of patients. PMID- 10868725 TI - Functional deterioration in adults with cerebral palsy. AB - OBJECTIVE: To examine the problem of functional deterioration of patients with cerebral palsy. SUBJECTS: Adults with cerebral palsy who work at community workshops throughout Japan. SETTINGS: Interviews combined with physical examinations. In the first part of the study a survey was conducted, to which 686 patients responded. In the second part, physicians conducted physical examinations on 163 patients. In the third part, the patients who had been examined in the second part were studied. RESULTS: Functional deterioration was noted in approximately 35% of adult patients with cerebral palsy; it was higher among those with involuntary movements of the head and neck and abnormal movement patterns during locomotion. Deterioration was also noted among those who reported an inadequate work environment, such as poor posture during work and unsuitability of the desk, chairs and tools. Deterioration over a five-year period was noted in 8 of 122 patients. CONCLUSION: Although the factors intrinsic to cerebral palsy are often responsible for the functional deterioration, environmental factors are not to be taken lightly if functional deterioration is to be prevented. PMID- 10868726 TI - Factors associated with strain in co-resident spouses of patients following stroke. AB - OBJECTIVE: To identify the factors associated with carer strain following stroke. DESIGN: Co-resident spouses of stroke patients were sent questionnaire measures of their perceptions of strain, stress, mood, handicap, adjustment, social support, life satisfaction and personality, and patient's mood and independence in activities of daily living. SETTING: Stroke spouses were identified from the stroke register at City Hospital, Nottingham. RESULTS: In a sample of 222 carers, 37% had significant strain. Strain was highly correlated with negative affectivity on the Positive and Negative Affectivity Scale, carer mood on the General Health Questionnaire-12 (GHQ-12) and carer's perceptions of patient's independence in activities of daily living on the Extended Activities of Daily Living Scale (EADL). Logistic regression analysis of 96 of these carers supported the correlations and showed three factors, carer GHQ-12, patient EADL and negative affectivity, were independently associated with carer strain. CONCLUSION: The relationship between these factors and strain needs to be tested prospectively. Early identification of carers who may be at risk of strain later on will enable services to be targeted at prevention rather than cure. PMID- 10868727 TI - An observational study of the Stroke Association family support organizer service. AB - OBJECTIVE: The aim was to investigate the role of Stroke Association family support organizers, in order to decide what outcome measures would be appropriate for evaluating their service. DESIGN: Family support organizers (FSOs) were observed on home and hospital visits by an independent observer. Every two minutes recordings were made of the location, individuals present and activity taking place. SETTING: FSOs working in the North-West and Midland regions of Britain. RESULTS: The majority (71%) of visits took place in the patient's home, with both patients and carers (39%). The most frequently occurring activities were enquiry (17%), task-related conversation (14%) and support-related conversations (14%). Comparison of FSOs showed variation in the time dedicated to each activity. CONCLUSIONS: FSOs spend more time engaged in practical rather than emotional activities. The role of the FSO is multifaceted. Evaluation of the FSO service needs to measure the patient's mood and functional abilities, the carer's mood and strain, and patients' and carers' knowledge and satisfaction with information and services. PMID- 10868728 TI - Ulnar nerve lesions: functional outcome after five years. AB - OBJECTIVE: To analyse perceived impairments, disability, job restrictions and job changes in subjects with a neurapraxia and neurotmesis of the ulnar nerve five years after trauma. DESIGN: Retrospective, descriptive follow-up study. SETTING: Department of Rehabilitation of a University Hospital. SUBJECTS: Sixteen subjects with a neurotmesis (NT-group) and 20 subjects with a neurapraxia (NP-group) were compared by means of a structured interview assessing perceived impairments and change in job and a questionnaire assessing disabilities (Groningen Activity Restriction Scale: GARS). RESULTS: The NP-group perceived significantly more pain, loss of strength and sensation, loss of dexterity and perceived more job related restrictions than the NT-group. The NP-group had significantly higher scores on the GARS. There was a moderate correlation between the visual analogue score of the pain and the GARS in the NP- and NT-groups. No significant difference was found between the groups with respect to job changes. CONCLUSIONS: This study shows that subjects with a neurapraxia of the ulnar nerve perceive more impairments and disabilities compared with those subjects with a neurotmesis. PMID- 10868729 TI - Evaluation of three methods to rate impairment in patients with complex regional pain syndrome I of one upper extremity. AB - OBJECTIVE: To gain insight into the best way of obtaining an impairment rating in complex regional pain syndrome I (CRPS I) of the upper extremity. This syndrome can potentially result in permanent impairment. DESIGN: Comparison of three evaluation methods to obtain impairment scores. Each patient was seen by one tester; two testers in total participated in the research. SETTING: Outpatient clinic of a university hospital. SUBJECTS: Seventy-four patients (27 men, 47 women, mean age 52 years) with CRPS I of one upper extremity. MAIN OUTCOME MEASURES: Methods I and II were conducted according to the American Medical Association's Guides to the evaluation of permanent impairment method I according to the general guidelines, and method II according to the methodology specificially described for CRPS I. Method III was developed by the Dutch Association of Neurologists. For comparison, differences between methods were plotted against their mean ratings, with the limits of agreement. Also the paired t-statistics were calculated (alpha = 0.05/3). RESULTS: The mean difference between methods I and II was -0.7% whole body impairment, between methods II and III 8.1% and between methods I and III 7.3%. Outcomes obtained with method III differed significantly from the other outcomes. CONCLUSIONS: Method I most accurately and objectively reflected the permanent impairment level resulting from CRPS I. PMID- 10868730 TI - Outcome measurement in brain injury rehabilitation--towards a common language. PMID- 10868731 TI - Respiratory changes in vasovagal syncope. AB - INTRODUCTION: Respiratory changes accompany the cardiovascular changes during head-up, tilt test-induced vasovagal syncope. METHODS AND RESULTS: Using the 45 minute 60 degrees head-up Westminster protocol, 29 patients were studied (mean age 53.9+/-20.0 years; 19 females). Two groups resulted: tilt-induced vasovagal syncope positive and negative. The cardiorespiratory parameters blood pressure (BP), heart rate (HR), tidal volume, and minute volume were measured. Comparisons of the cardiorespiratory parameters were made within the positive group and negative group, and then between the two groups. There were 14 in the positive group and 15 in the negative group. Baseline measurements were normalized to 1.0. Comparing the late tilt periods between the positive and negative groups, there were differences in BP (P < 0.002), HR (P < 0.002), tidal volume (P < 0.05), and minute volume (P < 0.002). In the positive group comparing early with late intervals: BP 1.11+/-0.09 versus 0.49+/-0.17, P < 0.0001; HR 1.18+/-0.12 versus 0.85+/-0.35, P < 0.009; tidal volume 1.39+/-0.34 versus 2.17+/-1.00, P < 0.015; and minute volume 1.24+/-0.26 versus 3.3+/-2.03, P < 0.0025. There were no comparable cardiorespiratory changes in the negative group. CONCLUSION: There were significant differences in the respiratory and cardiovascular parameters measured between those who were positive and those who were negative for tilt induced vasovagal syncope. Within the positive group, in addition to the falls in HR and BP, there were significant increases in minute volume and tidal volume during late tilt. This suggests that there may be a role for respiratory sensors in vasovagal syncope that may permit earlier and hence possibly more effective therapy for selected patients. PMID- 10868732 TI - Respiratory changes during the evolving vasovagal faint: physiologic and clinical implications. PMID- 10868733 TI - Prevalence and significance of focal sources of atrial arrhythmia in patients undergoing cardioversion of persistent atrial fibrillation. AB - INTRODUCTION: Recent reports have high-lighted the importance of focal atrial arrhythmias as a curable cause for a group of patients with frequently recurrent paroxysmal atrial fibrillation (AF). The importance of this arrhythmia mechanism in the general population of patients with persistent AF is unknown. METHODS AND RESULTS: After successful internal cardioversion of 50 consecutive patients with persistent AF (mean age 60 years, mean duration of AF 26 months), endocardial activity in the immediate postcardioversion period was analyzed for the presence of focal atrial activity. Postcardioversion atrial arrhythmias were considered to be focal if there was evidence of a localized source of repetitive early atrial activation, either in the form of (1) self-terminating monomorphic atrial tachycardia (at least five beats) or (2) recurrences of AF with an initial atrial activation sequence (first five beats) that was both monomorphic and reproducible with repeated recurrences. Evidence for a focal atrial arrhythmia was present in 20 of the total group of 50 patients (40%). Multivariate analysis of clinical characteristics revealed the diagnosis of lone AF as the only independent predictor of a focal source of AF (P = 0.028). Thirty-nine patients were discharged from hospital in sinus rhythm. At 1-month follow-up, 25 (64%) of these 39 patients had suffered AF recurrence. The only significant predictor of AF recurrence was evidence of a focal source of atrial arrhythmia immediately after cardioversion, with a relative risk of 1.73 (range 1.1 to 2.7; P = 0.015). CONCLUSION: Focal atrial arrhythmias are common in patients presenting with "idiopathic" persistent AF, suggesting a possible causative role in the generation of this common arrhythmia. PMID- 10868734 TI - Left ventricular diastolic dysfunction in patients with so-called lone atrial fibrillation. AB - Lone atrial fibrillation (AF) is defined by the absence of identifiable causes of AF, but its hemodynamics have not been investigated. Twenty-eight patients with lone AF were compared with 14 control patients referred for Wolff-Parkinson-White ablation. Transthoracic and transesophageal echocardiography were performed to rule out structural heart disease, followed by transseptally performed complete hemodynamic evaluation of the left heart systolic and diastolic function. There was no evidence of diastolic dysfunction according to echocardiographic criteria in AF and control patients. There was no difference in echocardiographic measurements, except for a significantly higher inferosuperior left atrial dimension seen in the four-chamber apical view in AF patients (51+/-10 vs 40+/-6 mm, P = 0.03). Hemodynamic evaluation showed that end-diastolic left ventricular pressure and the nadir of the left atrial Y descent were significantly higher in lone AF patients versus controls: 13+/-5 versus 8+/-3 mmHg (P = 0.001) and 6.7+/ 3 versus 4.6+/-2.7 mmHg (P = 0.05). Our results demonstrated the presence of diastolic left heart dysfunction in patients with so-called lone AF. PMID- 10868736 TI - Regional hyperkalemia increases ventricular defibrillation energy requirements: role of electrical heterogeneity in defibrillation. AB - INTRODUCTION: Increased spatial electrical heterogeneity has been associated with impaired defibrillation efficacy. The current study investigated the relationship between electrical heterogeneity and defibrillation efficacy by manipulating spatial electrical heterogeneity. METHODS AND RESULTS: We increased spatial electrical heterogeneity by infusing potassium chloride (2 to 4 mEq/hour) or placebo in the left anterior descending artery in 13 pentobarbital anesthetized swine. Electrophysiologic measurements at five myocardial sites and defibrillation energy requirement (DER) values were determined at baseline and during regional hyperkalemia (n = 7) or placebo (n = 6). Regional potassium infusion was titrated to a 20% reduction in action potential duration in the perfused region. Regional hyperkalemia increased biphasic DER values by 87% (P = 0.02), whereas infusion of placebo did not alter defibrillation efficacy. Regional hyperkalemia decreased myocardial repolarization and refractoriness in the perfused region by 21% (P < 0.001) and 18% (P = 0.01), respectively. However, regional hyperkalemia increased ventricular fibrillation cycle length (VFCL) by 39% (P = 0.008). Consequently, dispersions of repolarization, refractoriness, and VFCL were significantly increased by 169%, 92%, and 200%, respectively. Regional hyperkalemia also increased ventricular conduction time to the perfused region by 54% (P = 0.006), indicating conduction velocity dispersion, while not affecting local pacing threshold or local voltage gradient. CONCLUSION: Regional hyperkalemia increased DER values. Regional hyperkalemia likely impairs defibrillation by increasing myocardial electrical heterogeneity, which supports the theory that electrical heterogeneity promotes nonuniform propagation of early postshock activations, thereby inhibiting defibrillation. PMID- 10868735 TI - Molecular remodeling of Kv4.3 potassium channels in human atrial fibrillation. AB - INTRODUCTION: Atrial fibrillation (AF) is associated with important alterations in cardiac ion channels that cause shortening and impaired rate adaptation of atrial repolarization. The mechanisms underlying potassium current remodeling in human AF are not clear. We investigated the effects of AF on the gene expression of the Kv4.3, Kv1.4, and Kv1.5 potassium channel subunits and correlated the findings with the transient outward (Ito) and the sustained outward (Isus or I(Kur)) potassium current. METHODS AND RESULTS: Semiquantitative reverse transcription-polymerase chain reaction was used to evaluate mRNA expression, and ion currents were studied with the patch clamp technique in right atrial appendages from patients in AF and compared with those from patients in stable sinus rhythm (SR). The presence of AF was associated with a 61% reduction in Kv4.3 mRNA expression (P < 0.001 vs SR), which was paralleled by a reduction in Ito current densities in this group of patients (i.e., at +50 mV: 7.44+/-0.76 pA/pF in SR and 1.24+/-0.28 pA/pF in AF; P < 0.001 vs SR). mRNA levels of Kv1.4 were identical in the two groups. AF did not affect either the gene expression of Kv1.5 or the current densities of Isus. CONCLUSION: Chronic AF in humans reduces Ito by transcriptional down-regulation of the Kv4.3 potassium channel. Altered gene expression is an important component of the electrical remodeling process and may contribute to repolarization abnormalities in AF. PMID- 10868737 TI - Fibrillation and defibrillation: the odd couple? PMID- 10868738 TI - Electrophysiologic deterioration after one-minute fibrillation increases relative biphasic defibrillation efficacy. AB - INTRODUCTION: The probability of survival decreases to 70% after 2 minutes of ventricular fibrillation. Biphasic shocks are more effective than monophasic shocks in terminating short-duration (<30 sec) ventricular fibrillation. We tested the hypotheses that developing ischemia changes the electrophysiologic characteristics of fibrillation and that the relative efficacy of biphasic shocks increases as electrophysiologic characteristics deteriorate. METHODS AND RESULTS: Monophasic (12 msec) and biphasic (6/6 msec) shocks (1 to 4 A) were tested in random order in isolated rabbit hearts after 1-minute ischemic fibrillation. Monophasic action potentials showed only a sporadic occurrence of electrical diastole after 5 seconds of fibrillation (24% of action potentials in the right ventricle and 18% in the left ventricle). After 60 seconds of fibrillation, diastole (17.83+/-1.14 msec in the right ventricle and 21.52+/-1.16 msec in the left ventricle) appeared after almost every action potential (P < 0.0001 compared with 5 sec), despite a lack of change in fibrillation cycle length and dominant frequency. Monophasic I50 was 2.89 A, and biphasic I50 was 1.4 A (77% reduction in energy). Normalized curve width decreased 28%. Retrospective analysis showed that shocks delivered early in the fibrillation action potential had a greater probability of succeeding (89%) than shocks delivered late (30%; P < 0.001). CONCLUSION: After 1-minute ischemic fibrillation, diastolic intervals occur during fibrillation. Therefore, defibrillation shocks have an approximately 29% probability of interacting with the fibrillation action potential during diastole. At this time, biphasic shocks produced a more deterministic defibrillation threshold and became even more efficacious (I50 B/M = 0.48) than at short fibrillation durations (I50 B/M = 0.7). PMID- 10868739 TI - Changes in left ventricular repolarization and ion channel currents following a transient rate increase superimposed on bradycardia in anesthetized dogs. AB - INTRODUCTION: We previously demonstrated in dogs that a transient rate increase superimposed on bradycardia causes prolongation of ventricular refractoriness that persists for hours after resumption of bradycardia. In this study, we examined changes in membrane currents that are associated with this phenomenon. METHODS AND RESULTS: The whole cell, patch clamp technique was used to record transmembrane voltages and currents, respectively, in single mid-myocardial left ventricular myocytes from dogs with 1 week of complete AV block; dogs either underwent 1 hour of left ventricular pacing at 120 beats/min or did not undergo pacing. Pacing significantly heightened mean phase 1 and peak plateau amplitudes by approximately 6 and approximately 3 mV, respectively (P < 0.02), and prolonged action potential duration at 90% repolarization from 235+/-8 msec to 278+/-8 msec (1 Hz; P = 0.02). Rapid pacing-induced changes in transmembrane ionic currents included (1) a more pronounced cumulative inactivation of the 4-aminopyridine sensitive transient outward K+ current, Ito, over the range of physiologic frequencies, resulting from a approximately 30% decrease in the population of quickly reactivating channels; (2) increases in peak density of L-type Ca2+ currents, I(Ca.L), by 15% to 35 % between +10 and +60 mV; and (3) increases in peak density of the Ca2+-activated chloride current, I(Cl.Ca), by 30% to 120% between +30 and +50 mV. CONCLUSION: Frequency-dependent reduction in Ito combined with enhanced I(Ca.L) causes an increase in net inward current that may be responsible for the observed changes in ventricular repolarization. This augmentation of net cation influx is partially antagonized by an increase in outward I(Ca.Cl). PMID- 10868740 TI - Contact fluorescence imaging of reentry in monolayers of cultured neonatal rat ventricular myocytes. AB - INTRODUCTION: We present a novel contact fluorescence imaging (CFI) approach to monitor transmembrane potentials in monolayers of cultured neonatal rat ventricular cells. We apply CFI to demonstrate, for the first time, long-term recordings as well as electrical induction and termination of reentrant activity in this in vitro model. METHODS AND RESULTS: CFI was performed in confluent cell monolayers stained with di-8-ANEPPS. An anatomic obstacle (6 x 0.5 mm) was created in the center of the monolayers. Reentry was induced with a premature stimulus after pacing at 2 Hz (both via field stimulation). Seven sustained (>3 min) reentrant episodes, anchored to the anatomic obstacle, were observed in three monolayers. Field stimulation (30 V/cm) was applied to successfully terminate 6 of the 7 reentries. Analysis of reentrant activity showed similarities with anatomic reentry in tissue preparations, such as reduced conduction velocity around the core, variable conduction velocity along the reentrant pathway due to wavefront curvature effects, and field-induced activation at the obstacle borders leading to reentry termination (cardioversion). CONCLUSION: This study demonstrates the feasibility of CFI for macroscopic optical mapping of transmembrane potentials in a single layer of cultured cells. Our results suggest that the monolayer cell culture model is an attractive complement to tissue models of reentry and cardioversion. PMID- 10868741 TI - Pulmonary vein stenosis complicating catheter ablation of focal atrial fibrillation. AB - INTRODUCTION: A recently described focal origin of atrial fibrillation, mainly inside pulmonary veins, is creating new perspectives for radiofrequency catheter ablation. However, pulmonary venous stenosis may occur with uncertain clinical consequences. This report describes a veno-occlusive syndrome secondary to left pulmonary vein stenosis after radiofrequency catheter ablation. METHODS AND RESULTS: A 36-year-old man who experienced daily episodes of atrial fibrillation that was refractory to antiarrhythmic medication, including amiodarone, was enrolled in our focal atrial fibrillation radiofrequency catheter ablation protocol. The left superior pulmonary vein was the earliest site mapped, and radiofrequency ablation was performed. Atrial fibrillation was interrupted and sinus rhythm restored after one radiofrequency pulse inside the left superior pulmonary vein. Atrial fibrillation recurred and a new procedure was performed in an attempt to isolate (26 radiofrequency pulses around the ostium) the left superior pulmonary vein. Ten days later, the patient developed chest pain and hemoptysis related to severe left superior and inferior pulmonary veins stenosis. Balloon angioplasty of both veins was followed by complete relief of symptoms after 2 months of recurrent pulmonary symptoms. The patient has been asymptomatic for 12 months, without antiarrhythmic drugs. CONCLUSION: Multiple radiofrequency pulses applied inside the pulmonary veins ostia can induce severe pulmonary venous stenosis and veno-occlusive pulmonary syndrome. PMID- 10868742 TI - Successful catheter ablation in a patient with polymorphic ventricular tachycardia. AB - We describe a patient with polymorphic ventricular tachycardia (PVT)/ventricular fibrillation (VF) without organic heart disease who was cured by radiofrequency catheter ablation. The patient was a 65-year-old woman with a 10-year history of recurrent syncope. There was no evidence of organic heart disease, and the QT interval during sinus rhythm was borderline normal (corrected QT interval = 0.45 sec1/2). ECG recording during syncope showed PVT. On one occasion, PVT degenerated into VF. This PVT was always induced by a premature ventricular complex (PVC) originating from the right ventricular (RV) outflow tract. Rapid pacing (220 beats/min) at the site of PVC origin reproduced polymorphic change of the QRS wave on surface ECG that was similar to PVT. This suggests that the PVT originated from a single focus in the RV outflow tract. Catheter ablation was performed at the site of PVC origin. During 18-month follow-up, PVT/VF was not documented. PMID- 10868743 TI - Prevention of atrial fibrillation by complete compartmentalization of the left atrium using a catheter technique. AB - INTRODUCTION: Right atrial compartmentalization has been demonstrated to only reduce the number of atrial fibrillation (AF) episodes; left atrial (LA) fibrillation still occurs. METHODS AND RESULTS: We report successful LA compartmentalization resulting in isolation of all four pulmonary veins in a 51 year-old woman suffering from paroxysmal AF. Deployment of a complete encircling line resulted in dissociation of electrical activation within the isolated area from the remaining LA. Despite attempts at reinduction by pacing maneuvers inside and outside the isolated area, AF was no longer inducible. During 21-week follow up, the patient remained in stable sinus rhythm with rare atrial extrasystoles. CONCLUSION: If reproducible, this ablation strategy could allow treatment of AF independent of suppression of any triggering event. PMID- 10868744 TI - Evidence for a cardiac ion channel mutation underlying drug-induced QT prolongation and life-threatening arrhythmias. AB - The aim of this study was to test the hypothesis that some cases of drug-induced arrhythmias depend on genetic predisposition. Excessive prolongation of the QT interval and life-threatening arrhythmias (torsades de pointes or ventricular fibrillation) may occur in response to a variety of cardiac and noncardiac drugs, with detrimental effects on patient safety and the investments made by the pharmaceutical industry. Moss and Schwartz hypothesized that some drug-induced arrhythmias might represent cases of "forme fruste" of the congenital long QT syndrome (LQTS). The availability of molecular screening techniques for LQTS genes allowed us to test this hypothesis. An elderly female patient with documented cardiac arrest related to cisapride, a prokynetic drug that blocks I(Kr), and transiently prolonged QT interval underwent mutational analysis of the known LQTS-related genes performed by single-strand conformational polymorphism and DNA sequencing. Double-electrode voltage clamp in Xenopus oocytes as the expression system was used to study the in vitro cellular phenotype caused by the genetic defect in coexpression with the wild-type (WT) gene. Molecular analysis revealed a heterozygous mutation leading to substitution of a highly conserved amino acid in the pore region of KvLQT1. This mutation was present not only in the patient with ventricular fibrillation but also in her two adult asymptomatic sons who have a normal QT interval. In vitro expression of the mutated KvLQT1 protein showed a severe loss of current with a dominant negative effect on the WT KvLQT1 channel. Our findings demonstrate that some cases of drug-induced QT prolongation may depend on a genetic substrate. Molecular screening may allow identification among family members of gene carriers potentially at risk if treated with I(Kr) blockers. Evolving technology may lead to rapid screening for mutations of candidate genes that cause drug-induced life-threatening arrhythmias and allow early identification of individuals at risk. PMID- 10868745 TI - Defibrillator challenges for the new millennium: the marriage of device and patient-making and maintaining a good match. AB - Although it has become clear that implantable cardioverter defibrillators (ICDs) are effective, important challenges remain for the physician. Due to the limitations of available risk stratification tools, patient selection for primary sudden death prevention remains controversial in many populations. Additionally, the proliferation of device choices has led to challenges in matching the appropriate device to the individual patient: device size is balanced against longevity; the advantages of dual chamber systems is weighed against their increased complexity; physician and patient preferences in device implant site are constrained by site-dependent effects on defibrillation effectiveness and lead failure rates; and special consideration must be given to the patient with a preexisting pacemaker. After ICD placement, determination of appropriate follow up frequency and methodology to assess device function must be considered. This article will review patient selection, device implant site selection, device device interactions, single versus dual chamber ICD selection, and follow-up. PMID- 10868746 TI - Long QT syndrome: ionic basis and arrhythmia mechanism in long QT syndrome type 1. AB - Long QT syndrome type 1 (LQT1) causes torsades de pointes arrhythmia, ventricular fibrillation, and sudden death. It usually is inherited as an autosomal dominant trait (Romano-Ward syndrome). The primary defect in LQT1 is a mutation in KVLQT1, a gene that encodes the pore-forming alpha-subunit of a K+ channel. KvLQT1 alpha subunits coassemble with minK beta-subunits to form channels that conduct the slow delayed rectifier K+ current (I(Ks)) in the heart. Recessive mutations in KVLQT1 cause Jervell and Lange-Nielsen syndrome, which is characterized by more severe arrhythmias and congenital neural deafness. Heterologous expression studies demonstrated that mutations in KVLQT1 reduce I(Ks) by causing loss of channel function, altered channel gating, and/or a dominant-negative effect. It remains to be proven that an understanding of the molecular basis of LQT1 will lead to more effective therapy. PMID- 10868747 TI - Supraventricular tachycardia with 2:1 atrioventricular block: what is the mechanism? PMID- 10868748 TI - Pulmonary vein aneurysm in association with arrhythmogenic foci in a patient with focally initiated atrial fibrillation. PMID- 10868749 TI - Various ways to make phase singularities by electric shock. PMID- 10868750 TI - Exploring the relationship between alcohol and the driver characteristics in motor vehicle accidents. AB - This study examines the differences in alcohol-related accident involvement among different driver groups in the state of Florida. The driver characteristics considered in this study are: age, gender, race, and residency of the driver of a motor vehicle involved in an accident while under the influence of alcohol, drugs, or alcohol and drugs. The main objective of this study is to test whether there are associations between the different driver characteristics and alcohol involvement in traffic accidents, and to identify the high-risk group within each driver factor. This would improve our understanding of the relationship between alcohol involvement, accidents, and the four aforementioned driver factors. It would also enable us to better design educational and awareness programs targeting specific groups in the population to reduce drinking and driving in the state. The relationship between alcohol-related accident involvement and the driver factors are investigated using general descriptive statistics, conditional probabilities and log-linear models. The results showed that the 25-34 age group experience the highest rate of alcohol/drug involvement in accidents. The rates decline with the increase in the age of the drivers. The results also indicated that there are significant relationships between the driver characteristics and alcohol/drug involvement in accidents. Male, white, and in-state drivers were also more involved in alcohol/drugs-related traffic accidents. PMID- 10868751 TI - The relationship of alcohol safety laws to drinking drivers in fatal crashes. AB - This paper presents an analysis of the relationships between the passage of key alcohol safety laws and the number of drinking drivers in fatal crashes. The study evaluated three major alcohol safety laws--administrative license revocation laws, 0.10 illegal per se, and 0.08 illegal per se laws--on the proportion of drinking drivers in fatal crashes. Drivers aged 21 and older in fatal crashes at two BAC levels--0.01-0.09 and 0.10 or greater--were considered separately. Drivers under age 21 were not included because they are affected by the Minimum Legal Drinking Age (MLDA) law. This study used data on drinking drivers in fatal crashes from the Fatality Analysis Reporting System (FARS) covering 16 years (1982-1997) for all 50 states and the District of Columbia. Also included in the study were such variables as per capita alcohol consumption and annual vehicle miles traveled (VMT), which could affect the number of alcohol related crashes. The results indicate that each of the three laws had a significant relationship to the downward trend in alcohol-related fatal crashes in the United States over that period. This paper points out that this long-term trend is not the product of a single law. Instead, it is the result of the growing impact of several laws over time plus the affect of some factors not included in the model tested (such as the increasing use of sobriety checkpoints and the media's attention to the drinking-and-driving problem). PMID- 10868752 TI - Factors associated with falling asleep at the wheel among long-distance truck drivers. AB - Data on the prevalence and hypothesized predictors of falling asleep while driving were gathered through face-to-face interviews with 593 long-distance truck drivers randomly selected at public and private rest areas and routine roadside truck safety inspections. Hypothesized predictor variables related to drivers' typical work and rest patterns, extent of daytime and night-time drowsiness, symptoms of sleep disorder, measures of driving exposure, and demographic characteristics. A sizeable proportion of long-distance truck drivers reported falling asleep at the wheel of the truck: 47.1% of the survey respondents had ever fallen asleep at the wheel of a truck, and 25.4% had fallen asleep at the wheel in the past year. Factor analysis reduced the large set of predictors to six underlying, independent factors: greater daytime sleepiness; more arduous schedules, with more hours of work and fewer hours off-duty; older, more experienced drivers; shorter, poorer sleep on road; symptoms of sleep disorder; and greater tendency to night-time drowsy driving. Based on multivariate logistic regression, all six factors were predictive of self reported falling asleep at the wheel. Falling asleep was also associated with not having been alerted by driving over shoulder rumble strips. The results suggest that countermeasures that limit drivers' work hours and enable drivers to get adequate rest and that identify drivers with sleep disorders are appropriate methods to reduce sleepiness-related driving by truck drivers. PMID- 10868753 TI - Characterological, situational, and behavioral risk factors for motor vehicle accidents: a prospective examination. AB - The occurrence of motor vehicle accidents (MVAs) was studied prospectively in a sample of 500 drivers aged 19-88. Over a 4-year interval from 1991 to 1995, 36% of these drivers had a minor accident and 9% had a serious (injury-producing) accident. Data collected in 1991 demonstrated that crashes could be predicted from a combination of pre-existing characterological, situational, and behavioral risk factors, and that these risk factors largely explained sex and age differences in accident rates. The best predictors of future MVAs were younger age, high hostility in combination with poor self-esteem, residence in a larger city, recent relocation, high job stress, prior MVAs, and self-reported tendencies to speed and disregard traffic rules. Failure to wear seat belts did not predict accidents but did significantly influence the severity of accidents that did occur; that is, those who had earlier reported using seat belts 'always' were less likely than others to be injured when accidents did occur. Financial stress increased the likelihood of involvement in more serious accidents. PMID- 10868754 TI - Evaluation of photo radar program in British Columbia. AB - This article presents the results of an evaluation of the speed and traffic safety effects of the photo radar program in British Columbia (BC) after 1 year of full operation. Traffic speed data were collected from the photo radar units and from induction loops installed across the province. Traffic collision and injury data were obtained from police investigation reports and from BC ambulance services records. The study employed a number of analytical frameworks, including simple before and after comparison, time-series cross-sectional analysis, and interrupted time series analysis. The study revealed a dramatic reduction of speed at photo radar deployment sites. A reduction of 2.4 km/h in mean speed was also observed at selected monitoring sites where enforcement was not likely to be present. The reduction of speed was accompanied by a decrease in collisions, injuries and fatalities. The analysis found a 25% reduction in daytime unsafe speed related collisions, an 11% reduction in daytime traffic collision victims carried by ambulances and a 17%, reduction in daytime traffic collision fatalities. PMID- 10868755 TI - Effect of Florida's graduated licensing program on the crash rate of teenage drivers. AB - On 1 July 1996, Florida instituted a graduated licensing program for drivers younger than age 18. For the first 3 months, holders of learner's licenses are not allowed to drive at all between 19:00 and 06:00 h; thereafter, they may drive until 22:00 h. Learner's licenses must be held for 6 months prior to eligibility for the intermediate license. Sixteen-year-old intermediate license holders are not permitted to drive unsupervised from 23:00 to 06:00 h, 17 year-olds from 01:00 to 06:00 h. All drivers younger than 18 have strict limits on the number of traffic violations they can accumulate and, effective 1 January 1997, all drivers younger than 21 are subject to a zero tolerance law for drinking and driving. Florida crash data for 1995-1997 were obtained and compared with similar data from Alabama, a state that borders Florida but does not have graduated licensing. For 15, 16, and 17 year-olds combined, there was a 9% reduction in the fatal and injury crash involvement rate in Florida during 1997, the first full year of graduated licensing, compared with 1995. On a percentage basis, crashes declined most among 15 year-olds, followed by 16 year-olds and then 17 year-olds. Reductions were not seen among Alabama teenagers nor among 18 year-olds in Florida. PMID- 10868756 TI - Fatalities to occupants of cargo areas of pickup trucks. AB - We sought to describe the fatalities to occupants of pickup truck cargo areas and to compare the mortality of cargo area occupants to passengers in the cab. From the Fatality Analysis Reporting System (FARS) files for 1987-1996, we identified occupants of pickup trucks with at least one fatality and at least one passenger in the cargo area. Outcomes of cargo area occupants and passengers in the cab were compared using estimating equations conditional on the crash and vehicle. Thirty-four percent of deaths to cargo occupants were in noncrash events without vehicle deformation. Fifty-five percent of those who died were age 15-29 years and 79% were male. The fatality risk ratio (FRR) comparing cargo area occupants to front seat occupants was 3.0 (95% Confidence Interval [CI] = 2.7-3.4). The risk was 7.9 (95% CI = 6.2-10.1) times that of restrained front seat occupants. The FRR ranged from 92 (95% CI = 47-179) in noncrash events to 1.7 (95% CI = 1.5 1.9) in crashes with severe vehicle deformation. The FRR was 1.8 (95% CI = 1.4 2.3) for occupants of enclosed cargo areas and 3.5 (95% CI = 3.1-4.0) for occupants of open cargo areas. We conclude that passengers in cargo areas of pickup trucks have a higher risk of death than front seat occupants, especially in noncrash events, and that camper shells offer only limited protection for cargo area occupants. PMID- 10868757 TI - Injury trends of passenger car drivers in frontal crashes in the USA. AB - Injuries trends of passenger car drivers in head-on collisions are identified based on crash data extracted from the National Automotive Sampling System. Annual injury incidence levels are estimated for years 1990-2007. Over that period, the number of crashes is predicted to rise by 71%. However, the number of serious injuries to drivers is expected to rise by only 41% and driver fatalities are anticipated to decrease by 9%. Meantime, the types of injuries suffered by drivers are changing. Year-to-year shifts in injury patterns result from changes in vehicle size classes within the US vehicle fleet population and increases in seat belt use and air bag availability. The effectiveness of air bags in saving lives is estimated to be 30%, and with more air bag-equipped cars on the road, the probability of sustaining a life-threatening head or a torso injury is reduced. Air bags, however, are not as effective in preventing upper and lower extremity injuries, and thus arm and leg injuries will become more prevalent in years to come. PMID- 10868758 TI - The effectiveness of the 'ride-bright' legislation for motorcycles in Singapore. AB - This paper examines the effectiveness of the 'ride-bright' legislation implemented in Singapore in November 1995. The odds ratio test is used to investigate if there is any significant difference in the number of daytime motorcycle accidents by severity before and after the implementation of the legislation. The findings indicate that although there is insignificant change in the number of slight injury accidents, the legislation is effective in reducing the number of fatal and serious injury accidents. PMID- 10868759 TI - Head injuries to bicyclists and the New Zealand bicycle helmet law. AB - The purpose of this study was to examine the effect of helmet wearing and the New Zealand helmet wearing law on serious head injury for cyclists involved in on road motor vehicle and non-motor vehicle crashes. The study population consisted of three age groups of cyclists (primary school children (ages 5-12 years), secondary school children (ages 13-18 years), and adults (19+ years)) admitted to public hospitals between 1988 and 1996. Data were disaggregated by diagnosis and analysed using negative binomial regression models. Results indicated that there was a positive effect of helmet wearing upon head injury and this effect was relatively consistent across age groups and head injury (diagnosis) types. We conclude that the helmet law has been an effective road safety intervention that has lead to a 19% (90% CI: 14, 23%) reduction in head injury to cyclists over its first 3 years. PMID- 10868760 TI - Influence of drivers' comprehension of posted signs on their safety related characteristics. AB - The paper studies the relationship between drivers' understanding of posted signs in three of the Gulf Cooperation Council (GCC) states, Bahrain, Qatar and the United Arab Emirates (UAE), and some of their safety related characteristics. These characteristics are driving experience, accident involvement, experience per accident, citations received in the last 3 years on speed limit violations, and seat belt usage. A total of 28 posted signs were investigated. These were categorized as warning and regulatory. To achieve the above goals a questionnaire, specially prepared to collect the necessary data, was distributed to over 6000 drivers in the three states. Over 2820 (47%) responded back. Comprehension of posted signs for drivers with high years of driving experience proved to be significantly better than those with lesser experience. However, the results revealed no significant influence on their accident involvements, even when the effect of age is incorporated; experience per accident ratios, or speed citations. Further, the seat belt usage is also found to increase with understanding of posted signs. PMID- 10868761 TI - Alcohol use in relation to driving records among injured bicyclists. AB - To prevent alcohol-related occupational mishaps, employers often conduct background checks on prospective employees for history of driving while intoxicated (DWI) and driving under the influence of alcohol (DUI) to screen out candidates with drinking problems. Few studies, however, have examined the pervasive nature of drinking behavior across activity domains. Based on trauma registry data, we constructed a historical cohort of 120 Maryland residents ages 18 years or older who were injured while riding a bicycle between 1990 and 1997. Driving records for the 120 bicyclists for the 3 years between May 6, 1995 and May 5, 1998 were obtained from the state motor vehicle administration and were analyzed in relation to BAC-positive status at the time of injury. Bicyclists with positive BACs at the time of admission to the trauma center were significantly more likely than those with negative BACs to have a record of license suspension/revocation (52% vs 14%, P < 0.01) and to have DWI/DUI convictions (30% vs 3%, P < 0.01). Despite the modest sample size, this study provides compelling evidence of the pervasive nature of risky drinking between bicycling and driving activities. PMID- 10868762 TI - An expert judgment model applied to estimating the safety effect of a bicycle facility. AB - This paper presents a risk index model that can be used for assessing the safety effect of countermeasures. The model estimates risk in a multiplicative way, which makes it possible to analyze the impact of different factors separately. Expert judgments are incorporated through a Bayesian error model. The variance of the risk estimate is determined by Monte-Carlo simulation. The model was applied to assess the safety effect of a new design of a bicycle crossing. The intent was to gain safety by raising the crossings to reduce vehicle speeds and by making the crossings more visible by painting them in a bright color. Before the implementations, bicyclists were riding on bicycle crossings of conventional Swedish type, i.e. similar to crosswalks but delineated by white squares rather than solid lines or zebra markings. Automobile speeds were reduced as anticipated. However, it seems as if the positive effect of this was more or less canceled out by increased bicycle speeds. The safety per bicyclist was still improved by approximately 20%. This improvement was primarily caused by an increase in bicycle flow, since the data show that more bicyclists at a given location seem to benefit their safety. The increase in bicycle flow was probably caused by the new layout of the crossings since bicyclists perceived them as safer and causing less delay. Some future development work is suggested. Pros and cons with the used methodology are discussed. The most crucial parameter to be added is probably a model describing the interaction between motorists and bicyclists, for example, how risk is influenced by the lateral position of the bicyclist in relation to the motorist. It is concluded that the interaction seems to be optimal when both groups share the roadway. PMID- 10868763 TI - Recent European research on older drivers. AB - This paper discusses European research on older drivers published since 1985. It is not intended to be an exhaustive review; rather, the focus is on those issues that have been most topical during the past fifteen years. First, the paper deals with general efforts to integrate ageing into transport policy design. The emphasis is placed on differences in the European and American perspectives and discourse. Second, some research issues that have been topical in the European research agenda are reviewed, with a few examples of each. Third, a brief outline is given of newly emerging research issues of importance. PMID- 10868764 TI - The shift to and from daylight savings time and motor vehicle crashes. AB - The objective of the study was to examine whether the shifts to and from daylight savings time in Sweden have short-term effects on the incidence of traffic crashes. A database maintained by the Swedish National Road Administration was used to examine crashes from 1984 through 1995, that occurred on state roads the Monday preceding, the Monday immediately after (index Monday), and the Monday 1 week after the change to daylight savings time in the spring and for the corresponding three Mondays in the autumn. The Mondays 1 week before and after the time changes were taken as representing the expected incidence of crashes. Crash incidence was calculated per 1000 person-years using population estimates for each year of the study. The association between 1 h of possible sleep loss and crash incidence was estimated by the incidence rate ratio from negative binomial regression. The incidence rate ratio was 1.04 (95% CI, 0.92-1.16) for a Monday on which drivers were expected to have had 1 h less sleep, compared with other Mondays. In the spring, the incidence rate ratio for crashes was 1.11 (95% CI, 0.93-1.31) for Mondays after the time change compared to other spring Mondays. The corresponding rate ratio for the fall was 0.98 (95% CI, 0.84-1.15) It was concluded that the shift to and from daylight savings time did not have measurable important immediate effects on crash incidence in Sweden. PMID- 10868765 TI - Religious involvement and mortality: a meta-analytic review. AB - A meta-analysis of data from 42 independent samples examining the association of a measure of religious involvement and all-cause mortality is reported. Religious involvement was significantly associated with lower mortality (odds ratio = 1.29; 95% confidence interval: 1.20-1.39), indicating that people high in religious involvement were more likely to be alive at follow-up than people lower in religious involvement. Although the strength of the religious involvement mortality association varied as a function of several moderator variables, the association of religious involvement and mortality was robust and on the order of magnitude that has come to be expected for psychosocial factors. Conclusions did not appear to be due to publication bias. PMID- 10868766 TI - The natural history of cigarette smoking from adolescence to adulthood in a midwestern community sample: multiple trajectories and their psychosocial correlates. AB - Previous research on the natural history of smoking has focused on overall group trajectories without considering the possibility of risk subgroup variation. To address this limitation, the authors of the present study aimed to identify subgroups with varying trajectories of smoking behavior. The authors accomplished this within a cohort-sequential study of a large community sample (N = 8,556) with measurements spanning ages 11-31. After removing 2 a priori groups (abstainers and erratics), the authors empirically identified 4 trajectory groups -early stable smokers, late stable smokers, experimenters, and quitters--and psychosocial variables from adolescence and young adulthood were significantly distinguished among them. Given recent advances in quantitative methods, it is now feasible to consider subgroups of trajectories within an overall longitudinal design. PMID- 10868767 TI - Measuring secondhand smoke exposure in babies: the reliability and validity of mother reports in a sample of low-income families. AB - The reliability and validity of mother's reports of their infants' exposure to secondhand smoke (SHS) were examined in an ethnically diverse sample of low income, low-education families (N = 141 mothers). At baseline and posttest, smoking mothers reported about their infants' SHS exposure at different locations and by different sources during the previous week. Findings show that mothers can give reliable accounts of the degree to which they contribute to their babies' SHS exposure. Mothers are able to differentiate between their own smoking behavior and the extent to which they expose their infants. Consistent with the overall exposure pattern, exposure caused by the mother and exposure occurring at home showed the strongest associations with biological and environmental measures. These findings suggest that smoking mothers can provide reliable and valid reports of the degree to which their infants are exposed to SHS. PMID- 10868768 TI - Reconciling conflicting findings regarding postcessation weight concerns and success in smoking cessation. AB - Correlates of concern about weight gain following smoking cessation and self efficacy about controlling weight gain were examined in 940 men and 1,166 women who were surveyed on 2 occasions as part of a randomized trial of work-site interventions for smoking cessation. Weight concerns were positively associated with female sex, body weight, dieting for weight control, nicotine addiction, and social encouragement to quit. Bivariate analyses replicated prior findings that elevated weight concerns are associated with a reduced likelihood of quitting smoking, at least in women. Analyses controlling for demographics, nicotine dependence, and social factors replicated prior findings that weight concerns are not negatively related to smoking cessation and that some measures of concern are positively related to cessation. These analyses suggest that conflicting findings found in this literature are due primarily to how weight concerns are defined and whether covariates like nicotine addiction are used in data analyses. PMID- 10868769 TI - Radon and cigarette smoking: perceptions of this synergistic health risk. AB - Past approaches to the measurement of the perceived risk of combined hazards have failed to demonstrate awareness of synergy (S. E. Hampson et al., 1998; D. Hermand, E. Mullet, & B. Coutelle, 1995; D. Hermand, E. Mullet, & S. Lavieville, 1997). Respondents (N = 650) were provided with information about the synergistic risk of lung cancer from the combination of smoking and radon, and their risk perceptions were assessed on two occasions. At Time 1, using Likert-type scales, there was no evidence of synergistic risk perception. At Time 2, using a scale based on the appraisal of relative risk with anchors allowing for the expression of synergy, the combined hazard of radon and smoking was rated as significantly more of a health risk than the single hazards. The findings are discussed in terms of methodological issues in assessing synergistic risk. PMID- 10868770 TI - Protection and vulnerability processes relevant for early onset of substance use: a test among African American children. AB - This research tested predictions from a self-regulation model of factors relevant for early onset of tobacco and alcohol use with a community sample of 889 African American children (mean age = 10.5 years). Criterion variables were peer substance use, willingness to use substances, and resistance efficacy (intention to refuse substance offers). Structural modeling indicated effects of temperament dimensions were mediated through self-control and risk-taking constructs, which were related to school involvement, life events, and perceived vulnerability to harmful effects of substances. Peer use was predicted by life events, poor self control, and parent-child conflict; willingness was predicted by life events, risk taking, and (inversely) parental support; and resistance efficacy was predicted by perceived vulnerability and (inversely) poor self-control. Findings are discussed with reference to theoretical models of early protection and vulnerability processes. PMID- 10868771 TI - Similar-other support for men undergoing coronary artery bypass surgery. AB - This field experiment examined effects of a support intervention on the physical and mental health of coronary artery bypass graft (CABG) surgery patients. Control participants (N = 90) received usual hospital care; experimental participants (N = 100) also received visits from a "similar other" while in the hospital. Similar others were Veterans Administration veterans who had CABG surgery previously and were trained in simple supportive techniques. Outcomes were assessed prior to surgery and at 1, 6, and 12 months afterwards. Unexpectedly, the intervention generally had no effects on participants' well being. Further analysis showed that participants who talked often with fellow cardiac patients in the hospital ("de facto similar others") experienced improvements in their physical and emotional well-being over time. PMID- 10868772 TI - Marital satisfaction in patients with cancer: does support from intimate partners benefit those who need it the most? AB - This cross-sectional study assessed 3 ways of providing spousal support. Active engagement means involving the patient in discussions and using constructive problem-solving methods; protective buffering means hiding one's concerns; and overprotection refers to underestimation of the patient's capabilities, resulting in unnecessary help and excessive praise for accomplishments. Ratings of received spousal support by 68 patients with cancer revealed findings similar to those of partners' ratings of provided support. The positive association between active engagement and the patient's marital satisfaction was stronger for patients with a rather poor psychological and physical condition than for those with a rather good condition. Furthermore, protective buffering and overprotection were negatively associated with marital satisfaction only when patients experienced relatively high levels of psychological distress or physical limitations. PMID- 10868773 TI - Using implementation intentions to increase attendance for cervical cancer screening. AB - This article evaluates an intervention based on P. M. Gollwitzer's (1993) concept of implementation intentions. Women registered at a medical practice in rural England (N = 114) completed measures of the theory of planned behavior variables before a manipulation that induced one half of the sample to form implementation intentions specifying when, where, and how they would make the appointment. Subsequent attendance was determined from medical records. Findings show that the theory of planned behavior variables and previous delay behavior provided good prediction of attendance. However, despite equivalent motivation to attend, participants who formed implementation intentions were much more likely to attend for screening compared with controls (92% vs. 69%). Evidence also suggests that implementation intentions attenuated the relationship between previous delay behavior and subsequent attendance. PMID- 10868774 TI - Alcohol, sexual arousal, and intentions to use condoms in young men: applying alcohol myopia theory to risky sexual behavior. AB - Data from 7 studies were aggregated to examine how reported sexual arousal and alcohol intoxication interact to affect attitudes and intentions toward engaging in unprotected sexual intercourse in college-age men (N = 358). When participants were in a sober or placebo condition, their self-reports of sexual arousal had no effect on their responses. When participants were intoxicated, however, those who felt sexually aroused reported more favorable attitudes, thoughts, and intentions toward having unprotected sex than did those who did not feel aroused. These findings support alcohol myopia theory (C. M. Steele & R. A. Josephs, 1990), which states that alcohol intoxication restricts attentional capacity so that people are highly influenced by the most salient cues in their environment. It is suggested that sexual arousal is a powerful internal cue that interacts with alcohol intoxication to enhance attitudes and intentions toward risky sexual behaviors. PMID- 10868775 TI - The long-term risks in the short-term benefits: perceptions of potentially addictive activities. AB - It is generally assumed that individuals who take risks with their health either underestimate the magnitude of those risks or seek some benefit. This study assessed whether risk taking might also result from underestimating the benefits. In Study 1, lower estimates of the pleasure of drug use and risk of addiction were significantly related to increased self-reported experimentation and problems with drug use. This relationship remained significant even after controlling for preexisting psychosocial factors. In Study 2, perceptions of these "risks-in-the-benefits" were shown (a) to be distinct from perceptions of immediate benefits and (b) to serve as a protective factor against future alcohol use. Results are discussed in terms of creating improved interventions. PMID- 10868776 TI - Research on adherence, behavior change, and mental health: a workshop overview. AB - The National Institute of Mental Health sponsored a research workshop in 1999 to examine the potential of various theoretical approaches for enhancing adherence and behavior change, with special attention to mental disorders as mediators and outcomes. Complementary, multiple levels of analysis--including individual, small group, community, and cultural perspectives--were considered. Working groups identified problems and gaps in knowledge that if addressed would transform adherence and behavior change from overlooked or nuisance variables to critical variables in research on mental and medical disorders. PMID- 10868777 TI - Autism associated with the mitochondrial DNA G8363A transfer RNA(Lys) mutation. AB - We report a family with a heterogeneous group of neurologic disorders associated with the mitochondrial DNA G8363A transfer ribonucleic acid (RNA)Lys mutation. The phenotype of one child in the family was consistent with autism. During his second year of life, he lost previously acquired language skills and developed marked hyperactivity with toe-walking, abnormal reciprocal social interaction, stereotyped mannerisms, restricted interests, self-injurious behavior, and seizures. Brain magnetic resonance imaging (MRI) and repeated serum lactate studies were normal. His older sister developed signs of Leigh syndrome with progressive ataxia, myoclonus, seizures, and cognitive regression. Her laboratory studies revealed increased MRI T2-weighted signal in the putamen and posterior medulla, elevated lactate in serum and cerebrospinal fluid, and absence of cytochrome c oxidase staining in muscle histochemistry. Molecular analysis in her revealed the G8363A mutation of the mitochondrial transfer RNA(Lys) gene in blood (82% mutant mitochondrial DNA) and muscle (86%). The proportions of mutant mitochondrial DNA from her brother with autism were lower (blood 60%, muscle 61%). It is likely that the origin of his autism phenotype is the pathogenic G8363A mitochondrial DNA mutation. This observation suggests that certain mitochondrial point mutations could be the basis for autism in some individuals. PMID- 10868778 TI - Roles of glutamate transporter and receptors, poly (ADPribose) polymerase, and transforming growth factor-beta1 in pontosubicular neuron necrosis. AB - The expression of neuron-type glutamate transporters (EAAC-1), AMPA glutamate receptor subunits (GluR1 and GluR2/3), polyadenosine (5'diphosphate-ribose) polymerase (PARP), and transforming growth factor-beta1 was investigated in 20 cases of neonatal pontosubicular neuron necrosis and 12 gestational-age matched controls. Developmental immunoreactivities of EAAC-1, GluR1, and GluR2/3 appeared in the neurons of the pontine nuclei at 29 to 30 weeks' gestation in controls, and then gradually increased with age. However, these activities were decreased in the pontine nucleus of patients with pontosubicular neuron necrosis. Decreases in these immunoreactivities might indicate early degeneration of neurons. Although PARP and transforming growth factor-beta1 immunoreactivity was insignificant or very weak in the pontine nuclei at any age in controls, PARP was markedly expressed in karyorrhectic neurons of the pontine nucleus in patients with pontosubicular neuron necrosis. Transforming growth factor-beta1 immunoreactivity was observed in nonkaryorrhectic neurons of the pontine nuclei. PARP could contribute to the pathogenesis of pontosubicular neuron necrosis more than EAAC-1 or GluR1 or GluR2/3. Transforming growth factor-beta1 could play a role in the protection and repair of damaged neurons. PMID- 10868779 TI - Magnetoencephalographic study of giant somatosensory evoked responses in patients with rolandic epilepsy. AB - We report five patients with rolandic epilepsy associated with giant somatosensory responses to median nerve stimulation, in whom we analyzed the pathophysiologic relationship between rolandic discharges and the somatosensory responses using magnetoencephalography. Four of the five patients showed giant P30m, the current source of which was localized in the primary somatosensory cortex, while the first cortical response, N20m, was not enhanced, except in one patient. The current source of the giant middle-latency component, N70m, was localized posterior to that of N20m, possibly in the posterior parietal cortex, in all patients. The initial positive peak and large negative peak of rolandic discharges were identical to P30m and N70m with respect to the current source localization, wave form, topographic pattern, and time relationship in the electroencephalogram and magnetoencephalogram, and somatosensory evoked magnetic field and somatosensory evoked potential records, respectively. In addition, the secondary sensory cortex was considered to be the generator of the middle-latency component in one patient. In one patient, the current intensity of the N70m was normalized along with clinical improvement and the disappearance of rolandic discharges, whereas those of other somatosensory evoked magnetic field components remained unchanged. Our data suggest that the rolandic discharge generator mechanism in these patients could be closely related to the developmental alteration of excitability in the primary somatosensory cortex, posterior parietal cortex, and secondary somatosensory cortex, which decreased with age, and it could share a common neuronal pathway, at least in part, with the giant P30m-N70m (N90m) in the somatosensory evoked magnetic field through the sequential and parallel processing of somatosensory information. PMID- 10868780 TI - The syndrome of inv dup (15): clinical, electroencephalographic, and imaging findings. AB - The clinical and laboratory data of four pediatric patients and one adult patient with inverted duplication (inv dup) (15) are reported. The most evident findings were dysmorphic features with frontal bossing; genital abnormalities, such as macropenis or hypospadias; mental retardation; autistic behavior; and seizures. Two additional adults with inv dup (15) from other institutions were also diagnosed in our laboratory. Seizures and mental retardation were the reasons for their referral. The clinical picture of inv dup (15) seems to be quite variable since the phenotype can also be normal. However, karyotyping and fluorescent in situ hybridization, focused in particular on chromosome 15, appear to be indicated in patients with dysmorphic phenotypes, such as the one present in our patients, and in subjects with early-onset seizures and psychomotor retardation with autistic features. PMID- 10868781 TI - Familial mitochondrial intestinal pseudo-obstruction and neurogenic bladder. AB - Intestinal dysmotility and neurogenic bladder have been described as part of two autosomal-recessive mitochondrial disorders assumed to be due to a defect in communication between the nuclear and mitochondrial genomes: myoneurogastrointestinal encephalopathy (MNGIE) and diabetes insipidus, diabetes mellitus, optic atrophy, and deafness (Wolfram syndrome). Partial cytochrome c oxidase deficiency has been described in both. We describe three Ashkenazi Jewish siblings with progressive intestinal dysmotility, neurogenic bladder, and autonomic manifestations but no central nervous system involvement. Cytochrome c oxidase deficiency was demonstrated in peripheral and multiple intestinal muscle biopsies. Mitochondrial DNA analysis of an intestinal biopsy of patient 1 showed heteroplasmy consisting of a normal 16.5-kb band and an approximately 28-kb band, suggestive of a duplication. Mitochondrial DNA analysis of a muscle biopsy of patient 2 showed multiple deletions, mainly 10- and 11-kb bands. We suggest that this unique combination of intestinal pseudo-obstruction and neurogenic bladder could comprise a new autosomal-recessive mitochondrial disorder. PMID- 10868782 TI - Novel mutation in the CPT II gene in a child with periodic febrile myalgia and myoglobinuria. AB - We have identified a novel missense mutation in the carnitine palmitoyltransferase II (CPT II) gene in a child with CPT II deficiency characterized clinically by episodes of myalgia and myoglobinuria induced by intercurrent febrile illnesses. The patient was heterozygous for a G-to-A substitution at codon 487, changing an encoded glutamic acid to a lysine (E489K), while the other allele carried the common S113L mutation. This case enlarges the spectrum of mutations in patients with CPT II deficiency, and confirms the association of the S113L mutation with the muscular form. PMID- 10868783 TI - Occurrence of tics in Asperger's syndrome and autistic disorder. AB - Asperger's syndrome is a condition in the autistic spectrum in which language development is normal. Patients with Asperger's syndrome frequently exhibit repetitive movements (stereotypies), and can have motor and phonic tics in addition to other behavioral abnormalities. We present 12 patients with autistic spectrum disorders who were referred to our Movement Disorders Clinic for evaluation of tics. Eight of the 12 had normal language development and therefore met criteria for Asperger's syndrome. All patients exhibited stereotypic movements; in addition, seven had tics and six of these met diagnostic criteria for Tourette syndrome. Of the six patients with clinical features of both Asperger's syndrome and Tourette syndrome, three had severe congenital sensory deficits. The autistic patients in our series were clinically heterogeneous and though tics were clearly present, other aberrant movements demonstrated by them were harder to classify. Our series confirms the wide range of clinical manifestations in Asperger's syndrome and autism, including tics and other features of Tourette syndrome. Furthermore, it suggests that sensory deprivation contributes to the development of adventitious movements in this population. PMID- 10868784 TI - Agenesis of corpus callosum: clinical description and etiology. AB - In 135 children (aged 3 months to 15 years) with structural defects of the central nervous system found on magnetic resonance imaging, agenesis of the corpus callosum was evident in 7. The etiology of agenesis of the corpus callosum has been established in four children: partial trisomy of chromosome 13, partial duplication of the long arm of chromosome 10, Aicardi's syndrome, and intracranial bleeding during the fetal period as a result of injury. Agenesis of the corpus callosum coexisted with a Dandy-Walker malformation in one other patient, which suggests a genetic etiology. In spite of these variable etiologies, dysmorphic features were identified in all seven patients, as was psychomotor retardation. Epileptic seizures had occurred in six patients, and all manifested abnormalities on neurologic examination. PMID- 10868785 TI - Heterotopic ossification in childhood and adolescence. AB - Heterotopic ossification, or myositis ossificans, denotes true bone in an abnormal place. The pathogenic mechanism is still unclear. A total of 643 patients (mean age, 9.1 years) admitted for neuropediatric rehabilitation were analyzed retrospectively with respect to the existence of neurogenic heterotopic ossification. The purpose of this study was to obtain information about incidence, etiology, clinical aspect, and consequences for diagnosis and therapy of this condition in childhood and adolescence. Heterotopic ossification was diagnosed in 32 patients (mean age, 14.8 years) with average time of onset of 4 months after traumatic brain injury, near drowning, strangulation, cerebral hemorrhage, hydrocephalus, or spinal cord injury. The sex ratio was not significant. In contrast to what has been found in adult studies, serum alkaline phosphatase was not elevated during heterotopic ossification formation. A persistent vegetative state for longer than 30 days proved to be a significant risk factor for heterotopic ossification. The incidence of neurogenic heterotopic ossification in children seems to be lower than in adults. A genetic predisposition to heterotopic ossification is suspected but not proven. As a prophylactic regimen against heterotopic ossification we use salicylates for those patients in a coma or persistent vegetative state with warm and painful swelling of a joint and consider continuous intrathecal baclofen infusion and botulinum toxin injection for those patients with severe spasticity. We prefer to wait at least 1 year after trauma before excision of heterotopic ossification. PMID- 10868786 TI - Utility of early single photon emission computed tomography (SPECT) in neonatal gelastic epilepsy associated with hypothalamic hamartoma. AB - Gelastic epilepsy, or laughing seizures, is a rare seizure manifestation often associated with hypothalamic hamartoma. This seizure type is well described in older children and adults, but has only rarely been reported in neonates, oftentimes recognized in retrospect when the children are older. We report a child diagnosed at 3 months of age with a large hypothalamic mass after evaluation for spells occurring since birth. The spells were characterized by bursts of hyperpnea, followed by repeated "cooing" respirations, giggling, and smiling. These spells were recognized soon after birth in the delivery room, and occurred at 15-20 minute intervals. They did not interrupt feeding and occurred during sleep. On referral to our center, the patient was noted to be thriving, with normal medical and neurologic examinations except for his spells. The laboratory evaluation was normal, as were endocrine and ophthalmologic evaluations. Neuroimaging was performed, with magnetic resonance imaging demonstrating a large 2.8-cm isodense, nonenhancing hypothalamic mass. Electroencephalogram was abnormal, demonstrating bi-frontal sharp and spike-wave discharges. Video-EEG did not demonstrate ictal discharges associated with the patient's spells. Single photon emission computed tomography (SPECT) demonstrated dramatic ictal uptake in the area of the tumor, with normalization during the interictal phase. Partial excision of hamartomatous tissue has minimally improved the spells. In conclusion, this patient manifested an unusual, early presentation of a rare seizure type. SPECT scanning confirmed the intrinsic epileptogenesis of the hamartoma, further justifying a surgical approach to such patients. Early surgical intervention is probably indicated in an attempt to minimize or prevent the cognitive and behavioral sequelae commonly seen with this seizure type. PMID- 10868787 TI - Risk factors for cerebral palsy in very low-birthweight infants in the 1980s and 1990s. AB - One-hundred twenty-nine very low-birthweight infants were treated in Newborn and Infant Care Department of Children's Memorial Health Institute between 1985 and 1994; 89 were taken to prospective neurodevelopmental care. The newborns were divided into two groups. Group I had 38 preterm infants born from 1985 to 1989 and followed up at 7 to 11 years of age. Group II had 51 very low-birthweight infants treated from 1990 to 1994 and followed up at 2 to 5 years of age. Complicated, multiple pregnancy, normal delivery, and extremely low birthweight were significantly more frequent in group II. Very low-birthweight infants were frequently born by cesarean section in severe asphyxia. Only four (7.8%) newborns received surfactant therapy. From 1990 to 1994, respiratory distress syndrome III and IV, and a longer respiratotherapy period were significantly more frequent. From 1985 to 1994, the frequency of sepsis, periventricular leukomalacia, and normal ultrasonography was constant. Intraventricular hemorrhage I, II, and IV were frequently present in the 1990s, and intraventricular hemorrhage III was frequent in the 1980s. Cerebral palsy was diagnosed in 11 (28.9%) children in group I and 18 (35.2%) in group II (not statistically different). Multiple and complicated pregnancy, cesarean section, severe asphyxia, and respiratory distress syndrome did not increase the risk of cerebral palsy in very low birthweight infants. Periventricular leukomalacia has a more predictive value for cerebral palsy in these infants than did intraventricular hemorrhage. PMID- 10868788 TI - Common peroneal neuropathy due to surfing. AB - Common peroneal neuropathy is uncommon in children and adolescents. In this population, it is usually caused by direct nerve injury at the fibular head level. Most commonly, the nerve is damaged during sports-related blunt trauma. Other etiologies such as hereditary neuropathies and bone tumors are much less frequent. In some cases, repetitive microtrauma to the peroneal nerve is felt to cause neuropathy. We describe the case of a teenager who developed common peroneal neuropathy in association with prolonged wave-surfing in the presence of weight loss. PMID- 10868789 TI - Recurrent acute disseminated encephalomyelitis associated with acute cytomegalovirus and Epstein-Barr virus infection. AB - We describe a 5 6/12-year-old girl with recurrent episodes of acute disseminated encephalomyelitis following an acute cytomegalovirus infection and associated reactivated Epstein-Barr virus. Complete clinical recovery was obtained with intravenous immunoglobulin. PMID- 10868790 TI - Current therapy for West syndrome in Japan. AB - We sent questionnaires concerning the current therapy for West syndrome to 208 institutions at which pediatric care members of the Japan Epilepsy Society were working. Of these, 129 (62%) institutions responded. Vitamin B6 was the preferred first-line drug, followed by the combination of vitamin B6 and valproate or monotherapy with valproate. Corticotropin was the third choice among the drugs. The dosage of corticotropin was lower than previously reported. The treatment of West syndrome is not well established at present and further research is needed to improve the therapeutic protocol. PMID- 10868791 TI - Construction and characterization of monoclonal antibodies against western equine encephalitis virus. AB - A repertoire of mouse monoclonal antibodies (MAbs) against western equine encephalitis virus (WEE) was constructed and characterized. Anti-WEE antibodies were expressed from hybridomas and purified by protein G chromatography. Each of the antibodies was functionally assessed by indirect enzyme-linked immunosorbent assays (ELISAs), Western blotting, and immunoprecipitations. All antibodies bound to WEE antigen in ELISAs, whereas only a subgroup of antibodies was found to be active in Western blotting and immunoprecipitations. A subset of antibodies was found to cross-react with other alphaviruses, such as Sindbis virus (SIN), Venezuelan equine encephalitis (VEE), and eastern equine encephalitis (EEE). Because many of the antibodies were highly reactive to WEE antigen in one or more of the assays, these antibodies are excellent candidates for immunodetection and immunotherapy studies. PMID- 10868792 TI - Breast cancer progression and expression of blood group-related tumor-associated antigens. AB - There is sufficient evidence that blood group related Lewis antigens are tumor associated molecules. We have conducted immunohistochemical analysis of the expression of Lewis antigens in breast cancer tissue as an indicator of the degree of malignancy and as a prognostic factor. The studies were performed by examining 43 female patients diagnosed with invasive ductal carcinoma of the breast. Postoperative specimens were stained immunohistochemically using a panel of monoclonal antibodies (MAbs) specific for tumor-associated antigens: sialosyl LewisA, LewisA, LewisB, Lewisx, and LewisY. The aims of the study were to compare the appearance of metastases, degree of cancer stage (pTNM), and its histologic differentiation with the expression of Lewis phenotype. The evaluation of antigen expression was performed quantitatively and independently by two pathologists. Statistical significance was defined by Mann-Whitney test. The presented analysis of Lewis antigens showed higher expression of LeB and LeA (p = 0.03) in patients in stage N2 than in stage N1. The expression of LeB and LeY was higher in patients in stage T4 than in stage T1 (p = 0.02). No differences were observed for histologic differentiation. These data suggest that the expression of sialosyl-LeA and LeB antigens in breast cancer may predict metastases to lymph nodes. PMID- 10868793 TI - A monoclonal antibody directed against an autoimmune epitope on the human beta1 adrenergic receptor recognized in idiopathic dilated cardiomyopathy. AB - A monoclonal antibody (MAb M16) was obtained by immunizing Balb/C mice with free peptide H26R, corresponding to the second extracellular loop of the human beta1 adrenergic receptor (beta1AR), against which functional autoantibodies have been detected in patients with idiopathic dilated cardiomyopathy. The MAb was found to be of IgG2b type and directed against a conformational epitope, encompassing the sequence recognized by the human autoantibodies. BIAcore measurements yielded an equilibrium constant of 6.5 X 10(7) M1 with an association rate constant (kon) of 6.5 X 10(4) M(-1) sec(-1) and a dissociation rate constant (koff) of 1.0 X 10(-3) sec(-1). It immunoprecipitated only poorly the solubilized beta1AR of Sf9 cell membranes. Functionally, the MAb was capable of not only reducing the number of the maximal binding sites to the beta1-adrenergic receptor of transfected Sf9 cell membranes, but also of displaying a positive chronotropic effect on cultured neonatal rat cardiomyocytes. These properties, which the MAb shares with the human autoantibodies, makes it an interesting tool for passive transfer studies in mice. PMID- 10868794 TI - Expression of neutralizing recombinant human antibodies against Varicella Zoster virus for use as a potential prophylactic. AB - Chickenpox is a highly infectious disease that can be life-threatening to certain groups such as the newborn of nonimmune mothers and immunocompromised patients. At present, prophylactic treatment of individuals at risk involves the use of a polyclonal antibody preparation derived from the pooled sera of hyperimmune donors. While this product is effective, there are problems associated with maintaining supply, which depends on the availability of donors, and the variation of potency between batches. An effective human monoclonal preparation would be of value by providing a well-characterized and standardized preparation available on demand. In this study recombinant human anti-varicella zoster virus (VZV) monoclonals were generated from the mRNA of unstable anti-VZV secreting heterohybridoma cell lines, and characterized according to their molecular weight, isoelectric point, glycosylation, binding to C1q, and efficacy at neutralizing VZV in vitro. In one antibody (AEVZ 5.3) the VH region was grafted from the IgG1 parent antibody onto an IgG3 backbone to determine the effect of isotype on neutralization in vitro. Antibodies were expressed from NSO cells at concentrations of 3-24 microg/mL and contained the expected heavy and light chain fragments and N-linked glycan structures. Both AEVZ 5.1 and AVEZ 4 antibodies were IgG1 and recognized the viral coat protein glycoprotein E; both showed complement-independent and complement-enhanced neutralization. Changing the isotype of AEVZ 5.1 from IgG1 to IgG3 (AEVZ 5.3) further enhanced VZV neutralization in the presence of complement, but reduced its neutralization capacity in the absence of complement. Complement enhancement was consistent with our findings that the IgG3 form could bind more molecules of C1q. The results demonstrate the successful use of recombinant methods to generate stable, functional monoclonal antibodies. Modifications of the original antibodies were made with the aim of improving functionality. The resulting cell lines could be used for large-scale production of well-characterized antibodies for therapeutic use. PMID- 10868795 TI - Construction of ELISA system to quantify human ST2 protein in sera of patients. AB - The human ST2 gene can be specifically induced by growth stimulation in fibroblastic cells, and can also be induced by antigen stimulation in Th2 cells. The gene encodes a soluble secreted protein, ST2, and a transmembrane protein, ST2L, which are closely related to the interleukin-1 receptor. To gain insight into the biological roles of the ST2 gene, three monoclonal antibodies (MAbs) against human ST2 gene products were obtained. To obtain these antibodies, immunization was carried out using two different immunogens: purified soluble human ST2 protein (hST2), and COS7 cells, which express the extracellular portion of human ST2L. 2A5 and FB9 MAbs were derived from the immunization with soluble hST2, and HB12 was derived from the COS7 cell immunization. All three antibodies were shown to detect native forms of the human ST2 gene products by immunoprecipitation, flow cytometry, and enzyme-linked immunosorbent assay (ELISA). In the competitive ELISA using biotinylated and nonlabelled MAbs, neither FB9 nor HB12 affected the binding of 2A5 to ST2 gene products. Based on this result, we constructed a sandwich ELISA system using 2A5 and FB9 to measure the concentration of soluble hST2 in sera. The ELISA, combined with the flow cytometry using these antibodies, will be a useful tool for elucidating the functions of human ST2 gene products in individuals. PMID- 10868796 TI - A panel of CAb antibodies recognize endogenous and ectopically expressed ING1 protein. AB - Nine monoclonal antibodies (MAbs) against human and rodent ING1 protein have been generated using an IL6-secreting mouse myeloma line. These antibodies are all effective in recognizing ING1 protein in ELISAs, Western blot assays, and by indirect immunofluorescence. Combining different CAb monoclonal antibodies in a Western blot assay also allows detection of the very low levels of endogenous ING1 found in fibroblast cells in culture and the identification of at least two isoforms of ING1 in normal human diploid fibroblasts and established brain cancer cell lines. PMID- 10868797 TI - Monoclonal antibody B9-2 recognizes an 8 amino acid sequence that spans an autocatalytic cleavage site between the two subunits of human procaspase 8. AB - Caspase 8 is synthesized in a zymogen form and must be proteolytically cleaved to be activated. The catalytically active enzyme is composed of two of the four cleavage products. We have determined that the monoclonal antibody B9-2 recognizes the octomer EMDLSSPQ. This sequence is shared by two of the cleavage products: a decamer linker region released after autocatalysis and the smaller subunit of the active enzyme. PMID- 10868798 TI - Characterization of monoclonal antibodies specific for 14-kDa human group V secretory phospholipase A2 (hVPLA2). AB - Secretory phospholipase A2 (PLA2) consists of several 14-kDa isoforms with extensive homology, which makes it difficult to identify a specific isoform. In this study, we have developed and characterized monoclonal antibodies (MAbs) directed specifically against human group V sPLA2 (hVPLA2) derived from cultured hybridomas. These hybridomas were produced from the fusion of BALB/c-derived myeloma s/p20-Ag14 and splenocytes from mice immunized with purified recombinant hVPLA2. Three hybridomas secreting MAbs, MCL-3G1, MCL-2A5, and MCL-1B7, were selected and subcloned on the basis of their specificity to recognize hVPLA2 using solid-phase enzyme-linked immunoadsorbent assay (ELISA). The purified MAbs demonstrated a common pattern of immunoreactivity to hVPLA2, but not to human group IIa isoform (hIIaPLA2). Isotype analysis indicates that these hybridomas are of the IgG1 type. Under reducing conditions, MCL-3G1 sensitively detected hVPLA2 and demonstrated no cross-reactivity to either hIIaPLA2 or group IV cytosolic PLA2. Although specific for hVPLA2, a relatively modest signal was recognized with MCL-1B7 and MCL-2A5. These newly developed MAbs allow for determination of tissue distribution and cell-specific functions of hVPLA2. PMID- 10868799 TI - Monoclonal antibody against human growth hormone receptor. AB - GHR shows a high degree of homology with the prolactin receptor and with the other receptors that belong to the hemopoietic receptor superfamily. This paper describes a monoclonal antibody (MAb) (2B4B6) specific for both the extracellular domain of human GHR and human growth hormone (GH) binding protein. Mice were immunized against a seven-aminoacid peptide sequence screened by FASTA (sequence similarity search served by Genome-Net) from the European Bioinformatics Institute to exclude the existence of human membrane proteins with significant sequence homology. MAbs were screened against the peptide sequence and 2B4B6 was selected for its capability to recognize the full-length hGHBP. As evaluated by both enzyme-linked immunoadsorbent assay (ELISA) and FACS analysis, this MAb seems to recognize and bind to a hGHR positive cell line, IM-9, as well as a murine cell line, BaF3 (8/6), transfected with a chimeric construct, hGHR/hG-CSFR and expressing hGHR on the cell membrane. Studies investigating the biological effects of this MAb showed that anti-hGHR mediated inhibition of cell proliferation was not due to competition with GH binding but rather to prevention of receptor dimerization. Because of its specificity, this MAb may be usefully applied in situations in which GHR and receptors with a high degree of homology, such as PRLR (prolactin receptor), are expressed simultaneously, as occurs in the immune system. PMID- 10868801 TI - Mass production of monoclonal antibody in an ICR mouse using hybridoma cell lines MAC-1 and Aq-12 yields 165 mL ascitic fluid. PMID- 10868800 TI - Mouse monoclonal antibodies to hepatitis B virus preS1 produced after immunization with recombinant preS1 peptide. AB - We have efficiently generated mouse monoclonal antibodies (MAbs), which bind specifically to amino acids 21-47 of the preS1 domain of hepatitis B virus (HBV) by immunizing mice with the preS1 peptide (amino acids, aa 1-56) conjugated to keyhole limpet hemocyanin. Hybridomas were screened by an indirect enzyme-linked immunosorbent assay (ELISA) using the purified preS1 peptide as a coated antigen. Eighteen positive hybridomas were selected and subjected to isotyping. Of these, 5 clones secreted immunoglobulin G (IgG) and 13 clones secreted IgM. Four (KR1, KR2, KR3, and KR4) of the 5 IgG MAbs bound to preS1 peptide (aa 21-47). Epitope mapping using bacterially expressed GST fusion proteins revealed that three clones (KR2, KR3, KR4) (IgG1, K) recognize aa 21-35, while KR1 (IgG2a, K) recognizes aa 35-47 of the preS1. These MAbs immunoprecipitated HBV particles, demonstrating that they bind to native HBV particles. PMID- 10868802 TI - Archaic strains of methicillin-resistant Staphylococcus aureus: molecular and microbiological properties of isolates from the 1960s in Denmark. AB - Methicillin-resistant Staphylococcus aureus (MRSA) isolated in the 1960s in Denmark are among the historically earliest samples of these bacteria. We determined microbiological and molecular properties of 46 such isolates. They showed remarkably uniform properties, which included: (i) low methicillin MIC value (6-25 microg/ml); (ii) heterogeneous expression of resistance; (iii) the presence of a single, common, mecA polymorph II; (iv) lack of the regulatory gene mecI; (v) frequent lack of Tn554; and (vi) a common pulsed-field gel electrophoresis (PFGE) type. These properties, together with the chronological dates of isolation, and recovery of the strains from 18 hospitals scattered over Denmark, suggest that they represent a lineage close in time to the evolutionary origin of European strains of MRSA. PMID- 10868803 TI - Quinolone resistance among Salmonella enterica from cattle, broilers, and swine in Denmark. AB - This study was conducted to determine the susceptibility to nalidixic acid and fluoroquinolones of Salmonella Dublin, S. Enteritidis, and S. Typhimurium isolates from cattle, broilers, and pigs over time in Denmark and to characterise the gyrA, gyrB, and parC genes in quinolone-resistant isolates. A total of 584 S. Typhimiurium and 573 S. Dublin isolates from cattle during 1984 through 1999, and 241 S. Enteritidis and 131 S. Typhimurium from broilers and 452 S. Typhimurium from pigs isolated during 1997-1999 were tested. All isolates from cattle from the period 1984 through 1992 were susceptible to quinolones. A single (1.1%) S. Typhimurium isolate from 1995 and three (5.9%) from 1998 were resistant to nalidixic acid. Six (9.0%) S. Dublin isolates from 1996, four (4.2%) from 1997, and one (1.7%) from 1998 were resistant to nalidixic acid. Resistance was not observed among isolates from cattle in 1999. All broiler isolates from 1997 except for one were susceptible to nalidixic acid, whereas seven (6.2%) S. Enteritidis and two (6.3%) of the S. Typhimurium isolates from 1998 and 9 S. Enteritidis (26.5%) from 1999 were resistant. Among isolates from pigs, four isolates from 1997, three from 1998, and one from 1999 were resistant to nalidixic acid. All the nalidixic acid-resistant isolates had reduced susceptibility to fluoroquinolones. Sequence analysis of the gyrA gene in 37 nalidixic-resistant isolates identified two different base substitutions at codon serine-83 and two at aspartate-87. The base substitutions in serine-83 were TCC (Ser)-->TAC (Tyr), and TCC (Ser)-->TTC (Phe). The base substitutions in aspartic 87 were GAC (Asp)-->AAC (Asn), and GAC (Asp)-->GGC (Gly). Sequence analysis of the gyrB and parC genes revealed no mutations in 27 selected isolates. This study showed that quinolone-resistant isolates have emerged in recent years among food producing animals, especially among S. Enteritidis from broilers in Denmark, and that the resistance mainly is associated with mutations in gyrA. PMID- 10868804 TI - Characterization of aminoglycoside resistance genes and class 1 integrons in porcine and bovine gentamicin-resistant Escherichia coli. AB - A total of 78 gentamicin-resistant Escherichia coli strains from swine (27) and cattle (51) were characterized by phenotypic resistance, presence of selected aminoglycoside resistance genes, class 1 integrons and gene cassettes, and pulsed field gel electrophoresis. Gentamicin resistance was mainly encoded by the ant(2'')-I gene that was found in 76% of all the strains investigated, whereas the aac(3)-IIa gene was found in 14%. The ant(2'')-I gene was predominant in strains from cattle, whereas the porcine strains contained both ant(2'')-I, aac(3)-IIa, and the aac(3)-IVa genes. The pulsed-field gel electrophoresis (PFGE) investigation indicated a close clonal relationship in half of the bovine strains whereas the remaining E. coli were unrelated. Among the E. coli investigated, 20% contained class 1 integrons. Genes encoding resistance to trimethoprim (dhfrI, dhfrIb, and dhfrVII), gentamicin, tobramycin, and kanamycin (ant(2'')-Ia streptomycin and spectinomycin (ant(3'')-Ia) and streptothricin (sat1) were identified as gene cassettes. The most prevalent gene cassettes were ant(3'')-Ia (11 isolates) and the dhfrI (nine isolates). PMID- 10868805 TI - Comparison of genes involved in penicillin resistance in staphylococci of bovine origin. AB - Ten penicillin-resistant and -susceptible staphylococci, isolated from bovine mastitis milk, were studied for the presence of genes that are, or may be, involved in resistance against penicillin. The repressor (blaI), antirepressor (blaR1), and structural (blaZ) genes of the beta-lactamase-operon were found to be closely linked in all penicillin-resistant strains. The beta-lactamase gene cluster was more commonly located on chromosomal rather than plasmid DNA in the strains studied. The transposase (p480) gene, which has been identified in the Staphylococcus aureus beta-lactamase transposon Tn552, was found in only one single penicillin-resistant S. aureus strain. The other penicillin-resistant S. aureus isolates contained IS1181 in close location with the beta-lactamase gene cluster. In only one S. haemolyticus isolate was the beta-lactamase gene cluster found in close association with IS257. Penicillin-resistant S. aureus strains, which were additionally resistant to tetracycline, contained IS257 in close association with the tetracycline resistance gene (tetK). Sequence analysis of blaI, blaR1, and blaZ in two penicillin-resistant S. aureus strains revealed 94 96% sequence homology with bla in staphylococci of human origin. The results indicate a predominance of class I bla transposons rather than Tn3 family class II transposons in the isolates used in this study. PMID- 10868806 TI - Quinupristin/dalfopristin-resistant enterococci of the satA (vatD) and satG (vatE) genotypes from different ecological origins in Germany. AB - The semisynthetic streptogramin combination quinupristin/dalfopristin (Synercid) is a promising alternative for treatment of infections due to multiply resistant gram-positive bacteria including vancomycin-resistant Enterococcus faecium. Resistance is mediated by acetyltransferases SatA (VatD) or SatG (VatE). Recent papers have indicated a possible link between the use of the streptogramin virginiamycin S/M as a feed additive in commercial animal husbandry and a selection of quinupristin/dalfopristin-resistant E. faecium (QDRE). We screened manure samples from two different turkey farms and from six different pig farms (using virginiamycin), samples from a sewage water treatment plant, 24 broiler carcasses, 10 pork samples, and 200 stool samples of nonhospitalized humans for QDRE. Our strain culture collection of hospital E. faecium isolates from the last 2 years was also reviewed for QDRE. All manure and sewage samples were positive for QDRE, as well as 11 from broiler carcasses (46%), 1 from pork (10%), and 28 from human stool specimens (14%). Thirty-six hospital isolates of E. faecium exhibited resistance to quinupristin/dalfopristin. In 141 QDRE of different origin satA (vatD) and satG (vatE) genes were detected (seven isolates from humans with an unknown resistance mechanism). Streptogramin resistance determinants were tansferable in filtermating experiments for 5 of 10 satA (vatD) and 9 of 22 satG (vatE) isolates. Different EcoRI patterns of satG (vatE) plasmids and corresponding hybridizations of the satG (vatE) gene indicated nonhomologous resistance plasmids in isolates of different origin. The results of this study indicate a common gene pool for streptogramin resistance in E. faecium of different ecological origin. A selection of QDRE using the streptogramin virginiamycin S/M as a feed additive and a spread of the resistance via the food chain to humans is probable. PMID- 10868807 TI - Typeability of Tn1546-like elements in vancomycin-resistant enterococci using long-range PCRs and specific analysis of polymorphic regions. AB - Molecular subtyping of the VanA-type resistance element Tn1546 in an international collection of 81 genomically diverse vancomycin-resistant enterococci (VRE) from human, animal, and environmental reservoirs was evaluated by restriction analysis of long-range PCR amplicons (PCR-RFLP), single gene PCRs, Southern blot analysis of genomic digests, and partial DNA sequencing. A dominant Tn1546-RFLP in accordance with Enterococcus faecium BM4147 was detected in 43 of the 49 long-range PCR positive strains from ecologically diverse sources in several European countries and the US. Tn1546-like elements from the 32 (40%) long-range PCR negative strains were typed into 17 different groups by single gene PCRs and Southern blot analysis of the ORF1, ORF2, vanS-vanH, vanX-vanY, and vanZ regions. All these isolates showed deletions in the ORF1 and/or vanZ primer binding regions explaining the failure of long-range PCR amplification. Enlarged vanS-vanH or vanX-vanY fragments were detected in 7 (22%) and 16 (50%) of the long-range PCR negative strains, respectively. The enlarged vanS-vanH regions of five clinical isolates from the US (n = 2), Ireland (n = 2), and Norway (n = 1) contained identical IS1251-like insertions indicating intercontinental spread of the vanA gene cluster. Intergenic vanS-vanH IS1251 insertions have so far not been reported in European studies. Structural rearrangements of Tn1546-like elements may represent single recombination events that can serve as fingerprints in the molecular examination of vanA gene cluster evolution and transmission. The optimal strategy for such analysis has yet to be determined. Two alternative long range PCRs with subsequent RFLP analysis were successfully used to type the majority of vanA gene clusters in an ecologically and geographically heterogeneous VRE strain collection, but failed to detect and type a group of variant Tn1546-like elements truncated in the left-end ORF1/ORF2 region. Further subtyping of such variants should specifically target the polymorphic vanS-vanH and vanX-vanY regions. PMID- 10868808 TI - Incomplete cross resistance against ionophores in Enterococcus faecium and Enterococcus faecalis strains from pigs and poultry. AB - Thirty-two Enterococcus faecium strains and 33 Enterococcus faecalis strains were tested for their susceptibility to the ionophore antibiotics salinomycin, narasin, monensin, and lasalocid. Enterococcal strains originated from poultry in which these products are in use as coccidiostats, and from pigs in which these products are allowed as growth promoters. Resistance against salinomycin and narasin in enterococci was frequent among poultry strains, whereas in pig strains, resistance was less common. No resistance was found against monensin and lasalocid. Full cross resistance between salinomycin and narasin was evident. There was no cross resistance between these two ionophores and monensin and lasalocid. PMID- 10868809 TI - Associations between the use of antimicrobial agents for growth promotion and the occurrence of resistance among Enterococcus faecium from broilers and pigs in Denmark, Finland, and Norway. AB - This study compares the susceptibility of Enterococcus faecium isolated from pigs and poultry in Denmark, Finland, and Norway to antimicrobial agents used for growth promotion. E. faecium was isolated from 211 broilers and 55 pigs in Denmark in 1997, from Norwegian 55 poultry farms (turkey and broiler farms) and 4 swine farms between 1995 and 1997, and Finnish poultry (52) and swine (43) in 1996 and examined for susceptibility to avilamycin, avoparcin, bacitracin, flavomycin, monensin, salinomycin, spiramycin, tylosin, and virginiamycin. Only a limited number of isolates were categorized as resistant to monensin or salinomycin. In general, an association between the usage of antimicrobial agents in the respective countries and the occurrence of associated resistance was observed. Resistance to avilamycin was frequently observed among isolates from broilers in Denmark, where avilamycin has been used, whereas all isolates from Finland and Norway, where these drugs have not been used, were susceptible. The same phenomenon could be observed for avoparcin, bacitracin, tylosin, and virginiamycin; resistance was frequently observed among isolates from where these antimicrobials have been widely used, but rarely among isolates from where the use has been limited. Also for avoparcin and bacitracin, an association between usage and occurrence of resistance was observed. All isolates categorized as avoparcin resistant contained the vanX gene; isolates from broilers had the T variant in position 8,234 and isolates from pigs the G variant. Three (1%) of the 222 isolates resistant to tylosin contained the ermA gene and 196 (88%) ermB. Sixteen (11%) of the 146 virginiamycin-resistant isolates from broilers and two (7%) of the 27 virginiamycin-resistant isolates from pigs in Denmark contained the satA gene, whereas satA was not observed among any of the virginiamycin resistant isolates from Finland. A total of 72% of the virginiamycin-resistant E. faecium from broilers in Denmark and all nine virginiamycin-resistant E. faecium from Finland contained satG. This gene was also observed among two (7%) of the virginiamycin-resistant isolates from pigs in Denmark. This study indicates that the use of antimicrobial agents for growth promotion in Denmark, Finland, and Norway have selected for resistance to most of these drugs among E. faecium in food animals. PMID- 10868810 TI - Association between the use of avilamycin for growth promotion and the occurrence of resistance among Enterococcus faecium from broilers: epidemiological study and changes over time. AB - This study describes the changes in the occurrence of resistance to avilamycin among Enterococcus faecium from broilers in Denmark and the epidemiological association between usage of avilamycin for growth promotion and the occurrence of avilamycin-resistant E. faecium on broiler farms. The consumption of avilamycin for growth promotion increased from 10 kg in 1990 to 2,740 kg 1996 and decreased in the following years to only 7 kg in 1998. Most of this has been used for broilers. As part of the nationwide monitoring program for antimicrobial resistance, a total of 473 E. faecium isolates from broilers and 290 isolates from pigs have been tested for their susceptibility to avilamycin from 1995 to 1998. A very limited number of isolates from pigs were resistant to avilamycin, whereas the occurrence of resistance among isolates from broilers increased from 63.6% at the end of 1995 to a maximum of 80.7% during the last half of 1996. Since then, the occurrence of resistance has decreased to 23.3% in the last half of 1998. The epidemiological association between consumption of avilamycin and occurrence of resistant E. faecium fecal droppings were examined on 10 poultry farms that had not used avilamycin for growth promotion during 1996 or 1997 and eight farms that had used avilamycin during 1997. We tested a total of E. faecium isolates from the exposed farms and 104 from the nonexposed farms for their susceptibility to avilamycin. Resistant isolates were found on all eight exposed farms, and on seven of 10 nonexposed farms. Sixty-four isolates (72%) from the exposed farms were resistant, compared with 24 (23%) of the isolates from nonexposed farms. The adjusted chi-square p value equaled 0.01065, and showed a significant association between use of avilamycin and occurrence of resistance. The national monitoring program showed a decrease in the occurrence of resistance following a decreased use of avilamycin in Denmark, and the epidemiological study showed a statistically significant association between the use of avilamycin for growth promotion and the occurrence of avilamycin-resistant E. faecium on broiler farms. PMID- 10868811 TI - Origins and consequences of antimicrobial-resistant nontyphoidal Salmonella: implications for the use of fluoroquinolones in food animals. AB - Human Salmonella infections are common; most infections are self-limiting, however severe disease may occur. Antimicrobial agents, while not essential for the treatment of Salmonella gastroenteritis, are essential for the treatment of thousands of patients each year with invasive infections. Fluoroquinolones and third-generation cephalosporins are the drugs-of-choice for invasive Salmonella infections in humans; alternative antimicrobial choices are limited by increasing antimicrobial resistance, limited efficacy, and less desirable pharmacodynamic properties. Antimicrobial-resistant Salmonella results from the use of antimicrobial agents in food animals, and these antimicrobial resistant Salmonella are subsequently transmitted to humans, usually through the food supply. The antimicrobial resistance patterns of isolates collected from persons with Salmonella infections show more resistance to antimicrobial agents used in agriculture than to antimicrobial agents used for the treatment of Salmonella infections in humans. Because of the adverse health consequences in humans and animals associated with the increasing prevalence of antimicrobial-resistant Salmonella, there is an urgent need to emphasize non-antimicrobial infection control strategies, such as improved sanitation and hygiene, to develop guidelines for the prudent usage of antimicrobial agents, and establishment of adequate public health safeguards to minimize the development and dissemination of antimicrobial resistance and dissemination of Salmonella resistant to these agents. PMID- 10868812 TI - PER-1 extended-spectrum beta-lactamase production in an Alcaligenes faecalis clinical isolate resistant to expanded-spectrum cephalosporins and monobactams from a hospital in Northern Italy. AB - An Alicaligenes faecalis (FL-424/98) resistant to expanded-spectrum cephalosporins and aztreonam was isolated from the urine of an inpatient at the Intensive Care Unit of the Varese Hospital (Northern Italy) after antimicrobial chemotherapy with cefazolin, vancomycin, and amikacin. Clavulanic acid restored the activity of expanded-spectrum cephalosporins, suggesting the production of an extended-spectrum beta-lactamase (ESbetaL). A crude extract of FL-424/98 showed the presence of two beta-lactamase activities focusing at pH 5.3 and 7.6, respectively. The ESbetaL activity, purified by means of three chromatographic steps, was found to correspond to the pI 5.3 enzyme. Determination of kinetic parameters confirmed that the enzyme efficiently hydrolyzed expanded-spectrum cephalosporins and aztreonam. A colony-blot hybridization revealed the presence of blaPER-related sequences in FL-424/98, and sequencing confirmed the identity of this determinant with blaPER-1, previously detected in Pseudomonas aeruginosa, Acinetobacter, and Salmonella clinical isolates from Turkey. Finding of blaPER-1 in a species that can be part of the resident human microbiota raises the possibility that it could be an efficient shuttle for spreading of this resistance gene among other opportunistic pathogens that are normally members of the resident microbiota. Kinetic parameters determined for the PER-1 enzyme with some cephalosporin substrates were somewhat different from those previously reported. PMID- 10868813 TI - Mammography in New Hampshire: characteristics of the women and the exams they receive. AB - New Hampshire (NH) is one of two states that has developed a population-based mammography registry. The purpose of this paper is to describe what we have learned about mammography use in New Hampshire. After collecting data for 20 months, the database contains almost 110,000 mammographic encounters representing 101,679 NH women, who range in age from 18 to 97 with a mean of 56.7 years (SD=10.91). Education levels are high with 92% having a high school education and 59% with some college. Forty-six percent report their primary insurance is private, 29% report HMO/PPO coverage, and 25% receive federal health care assistance. Risk factors represented in the database include (categories not mutually exclusive) advancing age (60% over age 50), hormone replacement therapy use by menopausal women (40.6%), and a family history of breast cancer (29%). Penetration of mammography relative to the NH population is higher for younger age groups (40-48% for those aged 44-64) than older age groups (34-39% for those aged 65-84). The majority of mammographic encounters are routine screening exams (86%), often interpreted as negative or normal with benign findings (88%). Use of comparison films to interpret either diagnostic or screening mammography occurred in 86% of encounters. We have matched 3,877 breast pathology records to these mammographic encounters. The distribution of pathology outcomes for diagnostic exams was very similar to that for screening exams (approximately 65% benign, 17% invasive breast cancer, and 6% noninvasive breast cancer). Overall, we have designed a system that is well accepted by the NH community. Challenges include careful monitoring of data for coding errors, and a limitation of linking variables in mammography and pathology data. Data represented in this registry are a critical resource for research in mammographic screening and breast cancer early detection. PMID- 10868814 TI - A comparison of long pediatric hospitalizations in 1985 and 1994. AB - The objective of this study was to identify pediatric conditions commonly resulting in long hospitalizations, to evaluate changes in hospital use for these conditions over a 9-year period, and to describe the characteristics of children hospitalized for long periods (longer than 7 days). To accomplish this purpose we conducted a population-based, descriptive analysis of pediatric hospitalizations for children aged 1 to 12 years in California in 1985 and 1994 using hospital discharge data. We found that hospitalizations of longer than 7 days accounted for 10.8% of pediatric hospitalizations in 1985 (58.4% of pediatric hospital days) and 11.8% of hospitalizations in 1994 (50.4% of hospital days). Rates of long pediatric hospitalization decreased from 312.1/100,000 children in 1985 to 236.4/100,000 children in 1994. Rates fell for both sexes, in all racial/ ethnic groups, and among both preschool-age and school-age children. Common reasons for long hospitalizations in both 1985 and 1994 included lower-limb fractures, pneumonia, appendicitis, and malignancies. The rate of long hospitalization for mental disorders increased by 57% between 1985 and 1994, while the rate for injuries and poisoning decreased by 38%. In summary, long pediatric hospitalizations in 1985 and 1994 accounted for under 12% of all hospitalizations of children but for more than 50% of all hospital days. Although the overall rate of long pediatric hospitalizations decreased, rates for certain conditions, notably mental disorders, increased. As states continue to implement major health care changes, further study of conditions among children that account for a large proportion of hospital days is warranted. PMID- 10868815 TI - Assessing the health attitudes, beliefs, and behaviors of African Americans attending church: a comparison from two communities. AB - Public health officials and researchers continue to be increasingly concerned about the health of populations of color, especially African Americans. A survey was administered in African American churches in two communities (Wichita, KS and Tuscaloosa, AL) to gather information concerning health behaviors and beliefs and to design interventions that might improve their health status. The study examined the homogeneity of attitudes, beliefs, and behaviors across these samples and to determine the readiness to change using the Transtheoritical Model. Individuals completed a 33-item survey: 6 demographic questions, 12 health behavior questions, 8 health belief questions, 3 church attendance questions, and 4 church-based health promotion program questions. The total sample consisted of 429 respondents. The results showed that 93% of respondents have had their blood pressure checked in the past 2 years. While only 44% indicated eating a high fiber diet during the week. Thirty percent of respondents indicated that their health was dependent on fate or destiny. The findings from this study confirm that among both samples that health attitudes, beliefs, and behaviors need to be changed to lower the risk of certain diseases and disorders. The findings also indicate that both samples have similar beliefs about health that may have important implications for disseminating information to the community. Innovative and culturally sensitive programs are needed in the African American community if disparities in health are to diminish. PMID- 10868816 TI - Assessing managed care's role in promoting preventive care. AB - The current trend of managed health care systems opens the door to more effective control of chronic diseases through preventive care. The goal of this study was to assess managed care's role in promoting preventive care. A mail survey was conducted of a national sample of 1,200 directors, associated with preventive care, in managed care organizations (MCOs) in the U.S. Data was obtained on perceived effectiveness, degree of importance, and likelihood of support for implementation of strategies recommended (case management, utilization review programs, selective contracting, and cost sharing) for ensuring appropriate utilization of preventive services. Also, information was collected on interventions perceived effective in encouraging plan members to utilize and providers to offer preventive services. Response rate was 17.3%. Case management and prospective and concurrent utilization review programs were perceived most effective, important, and likely to receive support for implementation while cost sharing (using deductibles and coinsurance) and retrospective utilization review programs ranked low on all dimensions. Plan member-directed interventions perceived effective in encouraging utilization of preventive services included telephone and mail reminders while computer-generated reminders and medical record audits with feedback were perceived effective in encouraging providers to offer such services. Results identified preferred MCO strategies and interventions for ensuring appropriate utilization of preventive services. Further research is needed to develop methods to encourage people at high risk for chronic diseases not currently utilizing preventive services to receive such services. PMID- 10868817 TI - Knowledge, skills, and behavior improvements on peer educators and low-income Hispanic participants after a stage of change-based bilingual nutrition education program. AB - A nutrition education program, entitled La Cocina Saludable, was designed according to the Stage of Change Model and implemented in ten southern Colorado counties. The objectives were to improve the nutrition related knowledge, skills, and behaviors that lead to healthy lifestyles in a low-income Hispanic population. The content of the program included nutrition information designed to help mothers of preschool children provide for their children's nutritional needs. Previous studies suggest that low-income Hispanics often demonstrate low intakes of vitamins A and C, calcium, iron, and protein, and high rates of diabetes, obesity, and infections. Additionally, this population presents many obstacles for nutrition educators including limited resources, child care, transportation, time, language, culture, literacy, health beliefs, and, in some cases, the transient nature of the population. The program attempted to overcome these barriers by incorporating a flexible program format carried out by abuela (Hispanic grandmother) educators using the processes described in the Stage of Change Model. The program was evaluated using a knowledge, skills and behavior pre-test, post-test, and six-month follow-up survey on both the abuela educators as well as the actual class participants. Results of the peer education training sessions suggest that this type of training program can be effective in increasing the knowledge, skills, and behavior of peer educators as well as reduce need for retraining for educators who continuously teach classes. Additionally, the results suggest that this type of program can be effective in changing selected nutrition related knowledge, skills, and behaviors leading to healthy lifestyles for low-income Hispanic mothers of preschool children. PMID- 10868818 TI - Colorectal cancer screening in older men and women: qualitative research findings and implications for intervention. AB - As part of the formative research for developing interventions to increase colorectal cancer screening in men and women aged 50 and older, 14 focus groups were conducted to identify (1) knowledge, attitudes, and beliefs about colorectal cancer and colorectal cancer screening, (2) barriers to screening, and (3) strategies for motivating and supporting behavior change. Participants had either private insurance or Medicare and reported different levels of experience with colorectal cancer screening. Overall, they were poorly informed about colorectal cancer and the possible benefits of screening, reporting little or no information from physicians or mass media, negative attitudes toward screening procedures, and fear of cancer. Despite references to the subject matter as embarrassing or private, both men and women, African Americans and whites, appeared to talk candidly and comfortably in the permissive context of the focus group. This study's findings suggest that public education campaigns, decision aids, and targeted interventions are urgently needed to put colorectal cancer screening on the public's "radar screen," to increase awareness of the prevention and early detection benefits of screening, and to encourage people 50 and older-and the health care providers who serve them-to make screening a high priority. PMID- 10868819 TI - Problems associated with subcutaneously implanted glucose sensors. PMID- 10868820 TI - Hypoglycemia and driving performance: a flashing yellow light? PMID- 10868821 TI - Hypoglycemia and driving performance. PMID- 10868822 TI - Isotypes of anti-islet autoantibodies. PMID- 10868823 TI - Can medication packaging improve glycemic control and blood pressure in type 2 diabetes? Results from a randomized controlled trial. AB - OBJECTIVE: To assess the impact of calendar blister pack (CBP) use on glycemic and blood pressure control. RESEARCH DESIGN AND METHODS: We conducted an 8-month randomized controlled double-blind study among diabetic patients with poor glucose control (HbA1c >9.0%) in an urban area of South Auckland, New Zealand, with a high proportion of Maori and Pacific Islands people. Subjects included 68 consecutive patients, of whom 50% were prescribed three or more medications per day RESULTS: HbA1c was reduced by 0.95+/-0.22% in the CBP group and 0.15+/-0.25% in the control group (P = 0.026). Diastolic blood pressure decreased 5.8+/-1.5 mm Hg in the CBP group and increased 0.1+/-1.9 mm Hg in the control group (P = 0.0041). Systolic blood pressure did not change significantly CONCLUSIONS: CBPs should be considered among diabetic patients with poor glycemic control receiving multiple medications. PMID- 10868824 TI - Efficacy and safety of HOE 901 versus NPH insulin in patients with type 1 diabetes. The European Study Group of HOE 901 in type 1 diabetes. AB - OBJECTIVE: HOE 901 (Hoechst Marion Roussel, Frankfurt, Germany) is a biosynthetic insulin with a prolonged action. The aim of this study was to compare the effect of the long-acting insulin analog HOE 901 with NPH insulin regarding glycemic control in patients with type 1 diabetes. RESEARCH DESIGN AND METHODS: A total of 333 type 1 diabetic patients were enrolled in this multinational parallel group trial. Subjects were randomized either to two different formulations of HOE 901 (the formulations differed only in zinc content) or to NPH insulin. The study was only partially blinded because patients can distinguish HOE 901 (a clear solution) from NPH (a cloudy suspension). In addition to premeal injections of regular insulin, patients received HOE 901 at bedtime or NPH once daily at bedtime or twice daily in the morning and at bedtime. RESULTS: Fasting plasma glucose levels were significantly lower with HOE 901 (-1.88 mmol/l. P = 0.0005) as were fasting self-monitored blood glucose levels (-0.80 mmol/l, P = 0.0020). HbA1c levels also showed a significant reduction with HOE 901 (-0.14%) versus NPH (P = 0.030). The overall frequency of hypoglycemia did not differ, but the frequency of nocturnal hypoglycemia was significantly (P = 0.0037) lower with HOE 901 (36 vs. 55%). However, this effect on nocturnal hypoglycemia was significant only versus NPH once daily not NPH twice daily. The pattern of adverse events and injection site reactions with HOE 901 was similar to that with NPH. CONCLUSIONS: This study indicates that HOE 901 achieves better control of fasting glucose and HbA1c levels over 4 weeks, and HOE 901 has a possible safety benefit in terms of nocturnal hypoglycemia. PMID- 10868825 TI - Progressive hypoglycemia's impact on driving simulation performance. Occurrence, awareness and correction. AB - OBJECTIVE: Progressive hypoglycemia leads to cognitive-motor and driving impairments. This study evaluated the blood glucose (BG) levels at which driving was impaired, impairment was detected, and corrective action was taken by subjects, along with the mechanisms underlying these three issues. RESEARCH DESIGN AND METHODS: There were 37 adults with type 1 diabetes who drove a simulator during continuous euglycemia and progressive hypoglycemia. During testing, driving performance, EEG, and corrective behaviors (drinking a soda or discontinuing driving) were continually monitored, and BG, symptom perception, and judgement concerning impairment were assessed every 5 min. Mean +/- SD euglycemia performance was used to quantify z scores for performance in three hypoglycemic ranges (4.0-3.4, 3.3-2.8, and <2.8 mmol/l). RESULTS: During all three hypoglycemic BG ranges, driving was significantly impaired, and subjects were aware of their impaired driving. However, corrective actions did not occur until BG was <2.8 mmol/l. Driving impairment was related to increased neurogenic symptoms and increased theta-wave activity. Awareness of impaired driving was associated with neuroglycopenic symptoms. increased beta-wave activity, and awareness of hypoglycemia. High beta and low theta activity and awareness of both hypoglycemia and the need to treat low BG influenced corrective behavior. CONCLUSIONS: Driving performance is significantly disrupted at relatively mild hypoglycemia, yet subjects demonstrated a hesitation to take corrective action. The longer treatment is delayed, the greater the neuroglycopenia (increased theta), which precludes corrective behaviors. Patients should treat themselves while driving as soon as low BG and/or impaired driving is suspected and should not begin driving when their BG is in the 5.0-4.0 mmol/l range without prophylactic treatment. PMID- 10868826 TI - Relationship between several surrogate estimates of insulin resistance and quantification of insulin-mediated glucose disposal in 490 healthy nondiabetic volunteers. AB - OBJECTIVE: The goal of this study was to define the relationship between a quantitative measure of the ability of physiological hyperinsulinemia to stimulate glucose disposal and several surrogate measures of insulin resistance. RESEARCH DESIGN AND METHODS: Insulin-mediated glucose disposal was quantified in 490 healthy nondiabetic volunteers by determining the steady-state plasma glucose (SSPG) concentration in response to a continuous infusion of somatostatin, insulin, and glucose. Because the steady-state plasma insulin concentration was similar in all subjects during the infusion (approximately 60 microU/ml), the SSPG concentration provided a direct estimate of insulin-mediated glucose disposal. Relationships between this specific measure of insulin resistance and several surrogate estimates of insulin resistance based on plasma glucose and insulin concentrations were then defined. RESULTS: The surrogate measure of insulin resistance most closely related to the direct determination of insulin action was the total integrated insulin response to a 75-g oral glucose challenge with correlation coefficients (r) varying from 0.67 to 0.79. Fasting plasma insulin concentration was significantly correlated (r = 0.61, P<0.001) to the specific estimate of insulin action. Two other surrogate estimates of insulin action, the ratio of fasting glucose-to-fasting insulin concentration and the homeostasis model assessment for insulin resistance, were no more closely related to SSPG than the fasting plasma insulin concentration. CONCLUSIONS: The total integrated insulin response to oral glucose is the best surrogate measure of insulin resistance, accounting for approximately two-thirds of the variability in insulin-mediated glucose disposal. Fasting insulin concentration accounted for approximately one-third of the variability in insulin-mediated glucose disposal, and the use of fasting plasma glucose and insulin concentrations to calculate more sophisticated estimates of insulin resistance appears to offer little advantage over the fasting plasma insulin concentration. Given the large number of nondiabetic individuals in this study, the results should have general application in population-based studies, providing evidence for both the utility and limitation of the use of these surrogate measures. PMID- 10868827 TI - American Diabetes Association diabetes diagnostic criteria, advancing age, and cardiovascular disease risk profiles: results from the Third National Health and Nutrition Examination Survey. AB - OBJECTIVE: To evaluate age-specific effects on diabetes prevalence estimates resulting from the American Diabetes Association (ADA) recommendation against use of the oral glucose tolerance test (OGTT), we contrasted the prevalence of two mutually exclusive groups: undiagnosed diabetes according to ADA criteria (no report of diabetes and fasting glucose [FG] > or =126 mg/dl) and isolated postchallenge hyperglycemia (IPH) (FG <126 mg/dl and OGTT > or =200 mg/dl), a group designated to have diabetes by World Health Organization (WHO) criteria but not ADA criteria. RESEARCH DESIGN AND METHODS: The weighted age-specific ratios of undiagnosed diabetes:IPH were calculated for 2,844 subjects aged 40-74 years without reported diabetes who had both FG and OGTT. A ratio > 1.0 indicated that the proportion of undiagnosed diabetes was greater than that of IPH. Mean levels of HbA1c and cardiovascular disease (CVD) risk factors were contrasted among people with undiagnosed diabetes and IPH and those without either abnormality ("nondiabetic"). RESULTS: Both undiagnosed diabetes and IPH increased with age, but age-specific undiagnosed diabetes:IPH ratios decreased from 5.49 in the 40-44 age-group to 0.77 in the 70-74 age-group. Regression analysis showed a significant (P = 0.006) negative association between age and these ratios. Mean HbA1c was 7.1% in the undiagnosed diabetes group and differed significantly from that of the IPH and nondiabetic groups (5.6 and 5.3%, respectively). Individuals with undiagnosed diabetes had less favorable triglycerides, BMI, and HDL cholesterol compared with people with IPH. CONCLUSIONS: Compared with WHO criteria, the ADA criteria underestimate glucose abnormalities more with increasing age. However, compared to those with undiagnosed diabetes, individuals with IPH had a mean HbA1c level that is considered in the nondiabetic range, and this group had significantly more favorable levels of several key CVD risk factors. These findings suggest that the ADA criteria, although underestimating the abnormalities of postchallenge hyperglycemia that occur frequently with increasing age, appear to be effective at identifying a group of individuals with both unfavorable CVD risk factor profiles and evidence of long-term exposure to hyperglycemia. PMID- 10868828 TI - Prevalence of undiagnosed diabetes in three American Indian populations. A comparison of the 1997 American Diabetes Association diagnostic criteria and the 1985 World Health Organization diagnostic criteria: the Strong Heart Study. AB - OBJECTIVE: In 1997, the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus of the American Diabetes Association (ADA) recommended three new sets of criteria for the diagnosis of diabetes that were different from those established by the World Health Organization (WHO) in 1985. One of these three methods was based on a fasting plasma glucose value only. This article compares ADA criteria with WHO criteria by applying them to three subgroups of American Indians in the Strong Heart Study who had no known diabetes. RESEARCH DESIGN AND METHODS: The Strong Heart Study is a prospective epidemiological study of vascular disease in three American Indian populations aged 45-74 years. During the baseline examination from 1988 to 1991, participants without diagnosed diabetes underwent a fasting glucose test and a 2-h oral glucose tolerance test. These values were used to compare the ADA and WHO diagnostic criteria. RESULTS: By using fasting and 2-h glucose values, prevalence rates of undiagnosed diabetes were 15.9% according to WHO criteria and 14.4% according to ADA criteria. The overall agreement rate was 65%, and the weighted kappa statistic was 0.474, which indicates moderate agreement. The age-specific analysis showed that, among participants between 45 and 54 years of age, the prevalence rates of undiagnosed diabetes were 13.4% according to WHO criteria and 12.7% according to ADA criteria. Among those aged 55-74 years, the rates were 18.7% according to WHO criteria and 16.3% according to ADA criteria. Thus, the difference in the prevalence rates when using WHO and ADA criteria, although generally small in this population, was three times higher in the older group (2.4%) than the difference in the younger group (0.7%). CONCLUSIONS: The Strong Heart Study found that prevalence rates of undiagnosed diabetes determined by ADA criteria and WHO criteria were similar in its American Indian population. The data suggest that the difference between the two criteria may increase as age increases. Longitudinal data will be needed to evaluate further the utility of the two criteria. PMID- 10868829 TI - Use of GHb (HbA1c) in screening for undiagnosed diabetes in the U.S. population. AB - OBJECTIVE: To evaluate the use of GHb as a screening test for undiagnosed diabetes (fasting plasma glucose > or =7.0 mmol/l) in a representative sample of the U.S. population. RESEARCH DESIGN AND METHODS: The Third National Health and Nutrition Examination Survey included national samples of non-Hispanic whites, non-Hispanic blacks, and Mexican Americans aged > or =20 years. Of these subjects, 7,832 participated in a morning examination session, of which 1,273 were excluded because of a previous diagnosis of diabetes, missing data, or fasting time of <8 h before examination. Venous blood was obtained to measure fasting plasma glucose and GHb in the remaining 6,559 subjects. Receiver operating characteristic curve analysis was used to examine the sensitivity and specificity of GHb for detecting diabetes at increasing GHb cutoff levels. RESULTS: GHb demonstrated high sensitivity (83.4%) and specificity (84.4%) for detecting undiagnosed diabetes at a GHb cutoff of 1 SD above the normal mean. Moderate sensitivity (63.2%) and very high specificity (97.4%) were evident at a GHb cutoff of 2 SD above the normal mean. Sensitivity at this level ranged from 58.6% in the non-Hispanic white population to 83.6% in the Mexican-American population; specificity ranged from 93.0% in the nonHispanic black population to 98.3% in the non-Hispanic white population. CONCLUSIONS: GHb is a highly specific and convenient alternative to fasting plasma glucose for diabetes screening. A GHb value of 2 SD above the normal mean could identify a high proportion of individuals with undiagnosed diabetes who are at risk for developing diabetes complications. PMID- 10868830 TI - Evaluation of the effect of performance monitoring and feedback on care process, utilization, and outcome. AB - OBJECTIVE: We evaluated a program of performance measurement and monitoring by assessing care process, utilization of services, and outcomes. RESEARCH DESIGN AND METHODS: Information on 63,264 diabetic individuals who were continuously enrolled as members of Kaiser Permanente Southern California from 1 January 1994 to 31 December 1997 was used to evaluate the program. Time trends in testing for glycemic test and control and screening for dyslipidemia, use of lipid-lowering drugs, and microalbuminuria were evaluated as measures of care process. Time trends in hospitalization, outpatient appointments, prescriptions, and laboratory tests were evaluated as measures of utilization. Outcomes were hospitalization for myocardial infarction, ischemic stroke, and lower-limb amputation. RESULTS: Between 1994 and 1997, improvements were evident in the process measures. The mean number of hospitalizations and the mean and median number of outpatients visits did not change. The mean number of laboratory tests increased from 13.2 in 1994 to 23.6 in 1997. The mean number of prescriptions for any medication increased from 19.7 to 24.3. Hospitalization rates for myocardial infarction did not change, but rates increased for ischemic stroke and lower-limb amputation. CONCLUSIONS: Our findings suggest that measurement and monitoring of clinical performance can bring about modest improvements in measures of the processes of care in the absence of financial incentives, centrally driven interventions, and specialty care for all patients. In our setting, process improvements were associated with higher utilization of laboratory services and more prescriptions without an immediate return in terms of lower hospital utilization. PMID- 10868831 TI - Major stressful life events in relation to prevalence of undetected type 2 diabetes: the Hoorn Study. AB - OBJECTIVE: To test whether chronic psychological stress is positively associated 1) with the prevalence of type 2 diabetes; 2) with visceral adiposity; and to test whether 3) the relationship between stress and diabetes is mainly mediated by visceral adiposity RESEARCH DESIGN AND METHODS: In a general Caucasian population aged 50-74 years without a history of diabetes (n = 2,262), the number of major stressful life events experienced during the past 5 years was assessed by self-report before the administration of an oral glucose tolerance test. RESULTS: Diabetes was newly diagnosed among 5% of the subjects. The number of stressful events was positively associated with the prevalence of hitherto undetected diabetes. The highest quintile had a 1.6-fold (95% CI 1.0-2.6) increased probability of undetected diabetes compared with the remaining four quintiles (P<0.05 by logistical regression analysis adjusted for age and sex). This increased probability remained significant after additional adjustment for family history of diabetes, heavy alcohol consumption, physical activity, and low level of education. The number of stressful events was weakly positively associated with waist-to-hip ratio (WHR) (men, P<0.01; women, P = 0.05 by multiple regression analysis adjusted for age). The age- and sex-adjusted association between stress and diabetes was only marginally reduced by adding the WHR into the logistical regression model (odds ratio 1.5 [0.9-2.4]; P = 0.08). CONCLUSIONS: These cross-sectional findings are partially consistent with Bjorntorp's theory that stressful life events, which indicate chronic psychological stress, are indeed associated with undetected type 2 diabetes and with visceral adiposity. However, in this white middle-aged population, visceral adiposity does not seem to be the main link between stress and diabetes. PMID- 10868832 TI - Rapid and short-acting mealtime insulin secretion with nateglinide controls both prandial and mean glycemia. AB - OBJECTIVE: The objective of the study was to assess the efficacy and safety of four fixed doses of nateglinide compared with placebo in the treatment of patients with type 2 diabetes with focus on the prandial state. RESEARCH DESIGN AND METHODS: This randomized double-blind placebo-controlled multicenter study was conducted in 289 patients who received either nateglinide at doses of 30 mg (n = 51), 60 mg (n = 58), 120 mg (n = 63), or 180 mg (n = 57) or placebo (n = 60) before three main meals for 12 weeks. Levels of HbA1c, fasting plasma glucose (FPG), fructosamine, and plasma lipids were measured at predetermined intervals, and the effects of nateglinide on prandial glucose insulin, C-peptide, and triglyceride levels were measured after a liquid standard meal (Sustacal; Mead Johnson, Evansville, IN). Adverse events and hypoglycemic episodes were recorded. RESULTS: After a liquid meal challenge, nateglinide rapidly increased mealtime insulin levels within 30 min of drug intake and reduced mealtime glucose excursions without affecting triglyceride levels. At study end point, reduction of HbA1c levels was statistically significantly greater with nateglinide at doses of 60, 120, and 180 mg than placebo (-0.45, -0.62, and -0.64%, respectively; P<0.05). The mean level of FPG was significantly reduced versus placebo in the nateglinide 120-mg group only (-1.14 mmol/l P<0.01). Overall, nateglinide was well tolerated. CONCLUSIONS: This study demonstrated that nateglinide improves mealtime and mean glycemic control in a dose-dependent manner by restoring early insulin secretion phase. Nateglinide was well tolerated and is suitable for the treatment of patients with type 2 diabetes. PMID- 10868833 TI - A subcutaneous glucose sensor with improved longevity, dynamic range, and stability of calibration. AB - OBJECTIVE: To evaluate the lifetime, response time, linearity, glucose range, and calibration stability of two different types of continuous glucose sensor implants in a dog model. RESEARCH DESIGN AND METHODS: Glucose sensors based on the enzyme electrode principle that are coupled to a radio transmitter were evaluated on the bench top, sterilized, and then implanted subcutaneously in nondiabetic mongrel dogs. A multichannel radio receiver and PC data processor were used to record the sensor glucose data. Initial early reliable sensor responsivity was recognized by a vigorous hyperglycemic excursion after an intramuscular injection of glucagon. Periodically the dogs were made temporarily diabetic by blocking pancreatic insulin secretion by subcutaneous injection of a synthetic somatostatin (octreotide). By using exogenous insulin injection followed by intravenous glucose infusion, glucose levels were manipulated through the entire clinical range of interest: 2.2-38.9 mmol/l (40-700 mg/dl). Every 5-10 min, reference blood glucose samples were obtained and run in our hospital clinical laboratory. The glucose sensor data was evaluated by linear least squares optimization and by the error grid method. RESULTS: Beginning as early as postimplant day 7, the in vivo performances of sensors were evaluated by using glucose infusion studies performed every 1-4 weeks. Bench-top and in vivo 90% response-time sensors were in the range of 4-7 min during sensor lifetime. Best performing sensors from both types are summarized as follows. The earlier-stage technology was less linear with a dynamic range of no more than 22 mmol/l glucose, had a best-case recalibration interval of 18 days, and had a maximum lifetime of 94 days. The improved later-stage technology sensors, which were constructed with the addition of bioprotective and angiogenic membranes, were linear over the full extended range of clinical interest (2.2-38.9 mmol/l [40-700 mg/dl glucose]), had a best-case recalibration interval of 20 days, and had a maximum lifetime of >160 days. CONCLUSIONS: Stable clinically useful sensor performance was demonstrated as early as 7 days after implantation and for a sensor lifetime of 3-5 months. This type of subcutaneous glucose sensor appears to be promising as a continuous and painless long-term method for monitoring blood glucose. Specifically sensors with top-layer materials that stimulate angiogenesis at the sensor/tissue interface may have better dynamic measurement range, longer lifetimes, and better calibration stability than our previously reported sensors. PMID- 10868834 TI - Impaired skin microvascular function in children, adolescents, and young adults with type 1 diabetes. AB - OBJECTIVE: Vascular disease in type 1 diabetes is a complex and multifactorial process, which probably begins in childhood in association with the onset of diabetes. To determine the possible factors involved, we measured microvascular responses to endothelium-dependent (acetylcholine) and endothelium-independent (sodium nitroprusside) vasodilators in 56 patients with type 1 diabetes (aged 9 22 years) and 22 control subjects. RESEARCH DESIGN AND METHODS: Skin perfusion was measured at the dorsum of the foot using laser Doppler flowmetry during low current iontophoresis of acetylcholine and sodium nitroprusside. Maximum vasodilator function was measured during local 44 degrees C skin heating. RESULTS: Vascular responses were significantly reduced in patients with type 1 diabetes compared with responses in control subjects: acetylcholine (P<0.01, analysis of variance [ANOVA]), sodium nitroprusside (P<0.01, ANOVA), and local heating (P<0.02. Mann-Whitney U test). Endothelium-dependent responses were related to duration of diabetes (r = -0.38, P<0.01) and to glycemic control (r = 0.37, P<0.01). Significant correlations were found in the patient group between responses to acetylcholine and sodium nitroprusside (r = 0.28, P<0.05) but not to heating, suggesting that a common factor (e.g., nitric oxide activity) may be responsible for the abnormal vascular responses to these chemicals. CONCLUSIONS: Early changes in microvascular function are present in young patients with type 1 diabetes, long before the initial clinical presentation. These abnormalities may be related to complex interactions between structural abnormalities and functional changes in the endothelium, smooth muscle, and nitric oxide activity. PMID- 10868835 TI - Reduced beta-cell compensation to the insulin resistance associated with obesity in members of caucasian familial type 2 diabetic kindreds. AB - OBJECTIVE: Both obesity and a family history of diabetes reduce insulin sensitivity, but the impact of obesity on insulin secretion among individuals predisposed to diabetes is uncertain. We used a pedigree-based approach to test the hypothesis that beta-cell compensation to the insulin resistance associated with obesity is defective among individuals predisposed to diabetes by virtue of a strong family history of type 2 diabetes before the development of diabetes or glucose intolerance. RESEARCH DESIGN AND METHODS: A total of 126 members of 26 families ascertained for at least a sib pair with type 2 diabetes with onset before age 65 years underwent a tolbutamide-modified frequently sampled intravenous glucose tolerance test (FSIGT). Family members included 26 individuals with impaired glucose tolerance and 100 individuals with normal glucose tolerance (NGT). The acute insulin response to glucose (AIRglucose) was determined and insulin sensitivity (S(I)) estimated by minimal model analysis of FSIGT data. The beta-cell compensation for insulin sensitivity was estimated from the disposition index (DI), calculated as the product of S(I) and AIRglucose. Obesity was measured by BMI. RESULTS: Among all individuals, BMI was a significant predictor of both S(I) and AIRglucose, as expected. However, BMI also significantly predicted DI (P = 0.002) after correcting for age, sex, family membership, and glucose tolerance status. The relationship of BMI and DI was confirmed in 85 individuals with NGT who were aged <45 (P = 0.002) but not in 91 unrelated control individuals without a family history of diabetes. When normoglycemic individuals aged <45 were separated into three classes by BMI (< or =27, 27-30, >30), S(I) decreased progressively and significantly with obesity whereas AIRglucose rose significantly from lean to most obese classes. In contrast to the expectation of complete beta-cell compensation with obesity D1 fell significantly (P = 0.004) among obese family members. This relationship was not observed in control subjects. CONCLUSIONS: Individuals with a genetic predisposition to diabetes show a reduced beta-cell compensatory response to the reduced insulin sensitivity associated with obesity. We propose that this impaired compensation may be one manifestation of the underlying genetic defect in susceptible individuals. This finding helps explain the multiplicative effects of family history and obesity on risk of type 2 diabetes. PMID- 10868836 TI - Antibodies to IA-2 and GAD65 in type 1 and type 2 diabetes: isotype restriction and polyclonality. AB - OBJECTIVE: To determine the isotypes and clonality of antibodies to GAD (GADA) and IA-2 (IA-2A) in patients with type 1 and type 2 diabetes. RESEARCH DESIGN AND METHODS: We studied the following consecutive series of patients who attended a diabetes center for antibodies to GADA and IA-2A: 52 newly diagnosed type 1 diabetic patients, 199 type 2 diabetic patients, 200 control patients, and a cohort of 34 nondiabetic identical twins of patients with type 1 diabetes (15 of whom developed diabetes) who were followed prospectively. RESULTS: GADA or IA-2A were detected in 37 (71%) type 1 diabetic patients compared with only 10 (5%) type 2 diabetic patients (P<0.0001). Both GAD and IA-2 antibodies, regardless of the type of diabetes, were usually subclass restricted to IgG1 and were polyclonal. IgM, IgG3, and IgE isotypes were also detected, but all isotypes of GADA and IA-2A were less prevalent than IgG1 (P<0.017 for either antibody). There was no evidence of spreading or switching of isotypes before the onset of type 1 diabetes. CONCLUSIONS: These observations suggest that the pathogenesis of antigen-specific antibodies in type 1 and type 2 diabetes is similar and probably involves a chronic nonrandom antigen-driven polyclonal B-cell activation that is consistent with a Th1-type immune response. PMID- 10868837 TI - Serum markers of oxidative stress and severity of diabetic retinopathy. AB - OBJECTIVE: To compare serum markers of oxidative stress with diabetic retinopathy severity RESEARCH DESIGN AND METHODS: This cross-sectional study compared patients with types 1 and 2 diabetes with control subjects in western New York and Pennsylvania. Retinopathy severity was graded from funduscopic fields based on the Early Treatment of Diabetic Retinopathy Study. Serum samples were analyzed for thiobarbituric acid-reacting substances (TBARS), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities, creatinine, HbA1, and triglycerides. Appropriate analysis of covariance models were performed. RESULTS: TBARS (P = 0.019), triglyceride (P = 0.004), and glucose and HbA1 (both P<0.001) levels were elevated in diabetic patients compared with those in control subjects. SOD (P = 0.003) and GSH-Px (P = 0.046) levels were lower in diabetic patients than in control subjects. No correlation existed between SOD levels and either glucose or HbA1 levels. No significant associations existed between levels of TBARS, SOD, or GSH-Px and severity of diabetic retinopathy There was a significant association between poorer visual acuity and worse retinopathy (P = 0.009), which was only partly explained by macular edema. CONCLUSIONS: Increased levels of TBARS and decreased levels of SOD and GSH-Px were found in diabetic patients compared with those in control subjects, but no significant associations were found between the levels of these substances and severity of retinopathy When duration and type of diabetes and serum HbA1 levels were taken into account, only visual acuity remained associated with more severe retinopathy. PMID- 10868838 TI - Reassessing the role of QTc in the diagnosis of autonomic failure among patients with diabetes: a meta-analysis. AB - OBJECTIVE: A 1992 consensus statement on autonomic testing portrayed Bazett's heart rate-corrected QT interval (QT) prolongation as a specific yet insensitive indicator of diabetic autonomic failure. At that time, only a few small studies had evaluated the accuracy of QTc. To date, even fewer studies have evaluated whether its accuracy is influenced by patient characteristics. RESEARCH DESIGN AND METHODS: We critically appraised 17 studies reporting the sensitivity and specificity of QTc for diabetic autonomic failure. The studies represented 4,584 patients with diabetes (mean age 34.9 years, 46% female, 92% with type 1 diabetes, mean duration of diabetes 14.5 years). We summarized the accuracy of QTc prolongation for diabetic autonomic failure as an odds ratio (OR) (95% CI) and determined whether patient and study design characteristics influenced the accuracy of QTc prolongation by comparing summary receiver operating characteristic curves. RESULTS: Autonomic failure, defined as > or =1.2+/-0.4 (mean +/- SD) abnormal of 2.0+/-1.6 administered cardiovascular reflex tests, was found in 26% (25-28) of patients. The pooled sensitivity and specificity of QTc > 441+/-8 ms for autonomic failure were 28% (26-29) and 86% (85-87), respectively. Autonomic failure was 2.26 times (1.90-2.70) more likely to be present in patients with than in patients without QTc prolongation. At 86% specificity, the sensitivity of QTc prolongation was 46 vs. 12% for men versus women (P = 0.0077), respectively, and, after adjustment for sex, 66 vs. 17% among patients aged 25 vs. 55 years (P = 0.1902) and 61 vs. 27% at thresholds of >420 vs. >460 ms, respectively (P = 0.2964). CONCLUSIONS: QTc prolongation is a specific albeit insensitive indicator of autonomic failure. Although QTc prolongation is relatively accurate for men, accuracy may be even greater for young men at low QTc thresholds. PMID- 10868839 TI - American Diabetes Association Annual Meeting, 1999: insulin action and the development of type 2 diabetes. PMID- 10868840 TI - Plasma adrenomedullin levels in patients with diabetes. PMID- 10868841 TI - Differential antioxidant status among Indo-Asians compared with caucasians with and without diabetes. Wolverhampton Diabetes Research Group. PMID- 10868842 TI - The Batista procedure significantly improved cardiac function and glycemic control in a diabetic patient with severe insulin resistance. PMID- 10868843 TI - Elevated serum content of macrophage migration inhibitory factor in patients with type 2 diabetes. PMID- 10868844 TI - Lamin A/C mutation in a woman and her two daughters with Dunnigan-type partial lipodystrophy and insulin resistance. PMID- 10868845 TI - Effect of bezafibrate on insulin sensitivity in nonobese Japanese type 2 diabetic patients. PMID- 10868846 TI - MTHFR gene polymorphism as an exacerbation factor of diabetic nephropathy in type 2 diabetes. Analysis in Japanese male hemodialysis patients. PMID- 10868847 TI - Contribution of abnormalities of thyroid hormones to type 2 diabetes. PMID- 10868848 TI - Recent progress in glycohemoglobin (HbA1c) testing. PMID- 10868849 TI - The family and disease management in Hispanic and European-American patients with type 2 diabetes. AB - OBJECTIVE: To determine the relationship between the characteristics of families involved in disease management and the self-care practices of Hispanic and European-American (EA) patients with type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 74 Hispanic patients and 113 EA patients with type 2 diabetes recruited from managed care settings were assessed on three domains of family life (family structure/organization, family world view, and family emotion management [four scales]) and five areas of disease management (biological, general health and function status, emotional tone, quality of life, and behavioral [seven scales]). Analyses assessed the independent associations of patient sex, family, and sex by family interactions with disease management. RESULTS: Both sex and the three domains of family life were related to disease management, but the results varied by ethnic group. For EA patients, sex, family world view, and family emotion management were related to disease management (scores for Family Coherence were negatively associated with HbA1c level and depression, and poor scores for Conflict Resolution were linked with high depression); for Hispanic patients, sex and family structure/organization were related to disease management (high scores for Organized Cohesiveness were associated with good diet and exercise, and high scores for Family Sex-Role Traditionalism were related to high quality of life). No significant interactions with sex occurred. CONCLUSIONS: Characteristics of the family setting in which disease management takes place are significantly linked to patient self-care behavior, and these linkages vary by patient ethnicity. A family's multiple independent dimensions provide multiple targets for intervention, and differences in family norms, structures, and emotion management should be considered to ensure that interventions are compatible with the setting of disease management. PMID- 10868850 TI - Predictors of glycemic control in insulin-using adults with type 2 diabetes. AB - OBJECTIVE: To determine the characteristics that influence glycemic control among insulin-using adults with type 2 diabetes. RESEARCH DESIGN AND METHODS: We studied all 1,333 eligible members of a large not-for-profit health maintenance organization who responded to a 1997 survey. We tested associations among demographic, treatment, and psychometric variables with mean 1997 HbA1c values. The Problem Areas in Diabetes (PAID) instrument was used to assess the emotional effect of living with diabetes, and the Short Form 12 Physical Function Scale was used to assess the effect of physical limitations on daily activities. Based on differences between and within treatment groups, we built models to predict glycemic control for subgroups of subjects who were using insulin alone and those who were using insulin in combination with an oral hypoglycemic agent. RESULTS: Younger age, lower BMI, and increased emotional distress about diabetes (according to the PAID scale) were all significant predictors (P < 0.05) of worse glycemic control. However, except among individuals with an HbA1c level of >8.0 who were receiving combination therapy, only approximately 10% of the variance in glycemic control could be predicted by demographic, treatment, or psychometric characteristics. CONCLUSIONS: Personal characteristics explain little of the variation in glycemic control in insulin-using adults with type 2 diabetes. Possible explanations are that the reduced complexity of control in type 2 diabetes makes the disease less sensitive to personal factors than control in type 1 diabetes, that health-related behavior is less driven by personal and environmental characteristics among older individuals, or that, in populations exposed to aggressive glycemic control with oral hypoglycemic agents and nurse care managers, personal differences become largely irrelevant. PMID- 10868851 TI - Cardiovascular risk profile in individuals with borderline glycemia: the effect of the 1997 American Diabetes Association diagnostic criteria and the 1998 World Health Organization Provisional Report. AB - OBJECTIVE: In 1997, the American Diabetes Association (ADA) recommended a new diagnostic category, impaired fasting glucose (IFG), to describe individuals with borderline glucose tolerance. On the other hand, the World Health Organization (WHO) suggested retaining the category of impaired glucose tolerance (IGT). We studied the prevalence of IFG and IGT in a multiethnic society and compared the cardiovascular risk profiles of subjects with IFG, IGT, or both IFG and IGT. RESEARCH DESIGN AND METHODS: A total of 3,568 subjects were examined from the 1992 National Health Survey of Singapore, which involved a combination of disproportionately stratified sampling and systematic sampling. Anthropometric, blood pressure, insulin, lipid profile, and uric acid measurements were taken, and a standard 75-g oral glucose tolerance test was performed after a 10-h overnight fast. RESULTS: The prevalence rates of IFG only, IGT only, and both IFT and IGT were 3.45, 10.2, and 3.4%, respectively. The degree of agreement (kappa) between the two diagnostic criteria (the ADA IFG and the WHO IGT) was only 0.25. A fasting glucose level of 5.5 mmol/l was the optimal cutoff for predicting a 2-h postload glucose level of > or =7.8 mmol/l. The following cardiovascular risk factors were higher in subjects with both IFG and IGT compared with those with either IFG or IGT alone: systolic blood pressure (131 +/- 20 vs. 125 +/- 21 and 125 +/- 19 mmHg, respectively; P < 0.05 and P < 0.001, respectively); diastolic blood pressure (77 +/- 12 vs. 73 +/- 12 and 74 +/- 12 mmHg, respectively; P < 0.05); BMI (26.2 +/- 4.2 vs. 24.4 +/- 4.0 and 24.6 +/- 4.4 kg/m2, respectively; P < 0.01 and P < 0.001, respectively); waist circumference (84.1 +/- 10.3 vs. 79.3 +/- 10.7 and 79.3 +/- 10.6 cm, respectively; P < 0.001); waist-to-hip ratio (0.84 +/- 0.08 vs. 0.82 +/- 0.09 and 0.81 +/- 0.08, respectively; P < 0.05 and P < 0.001, respectively); fasting insulin (12.1 +/- 9.7 vs. 9.2 +/- 5.3 and 9.9 +/- 7.7 mU/l; P < 0.01); insulin resistance (by homeostasis model assessment [HOMA]) (3.41 +/- 2.77 vs. 2.58 +/- 1.50 and 2.43 +/- 1.83, respectively; P < 0.01 and P < 0.001, respectively); total cholesterol (5.81 +/- 1.1 vs. 5.51 +/- 1.1 and 5.53 +/- 1.1 mmol/l, respectively; P < 0.05) and apolipoprotein(B) [apo(B)] (1.5 +/- 0.38 vs. 1.40 +/- 0.34 and 1.39 +/- 0.35 mmol/l, respectively; P < 0.01). The pattern of difference remained significant only for fasting insulin, insulin resistance (HOMA), and apo(B) (borderline) after adjustment for age, sex, and ethnic differences. CONCLUSIONS: Obvious discordance was evident in the classification of glycemic status when applying the criteria proposed by the ADA (IFG) or WHO (IGT) in a multiethnic society like Singapore. However, subjects with either IFG or IGT had similar cardiovascular risk profiles. Therefore, both criteria identified individuals at high risk for cardiovascular disease. Individuals with both IFG and IGT had a greater incidence of the cardiovascular dysmetabolic syndrome. PMID- 10868852 TI - Normal blood pressure and preserved diurnal variation in offspring of type 2 diabetic patients characterized by features of the metabolic syndrome: the Fredericia Study. AB - OBJECTIVE: To examine whether an elevated blood pressure (BP) level and an impaired reduction in nocturnal BP are already present in nondiabetic first degree relatives of type 2 diabetic patients. RESEARCH DESIGN AND METHODS: We examined 253 offspring of type 2 diabetic patients using ambulatory BP monitoring and compared the BP level and profile with 275 offspring of nondiabetic subjects. Anthropometric measures and cholesterol, fasting blood glucose, and insulin levels were also compared between groups. RESULTS: No significant differences in BP level (P > 0.05) or diurnal BP profile were evident between the nondiabetic glucose-tolerant offspring of type 2 diabetic subjects and the offspring of nondiabetic subjects. BMI (P < 0.05 and P < 0.01, male vs. female), waist-to-hip ratio (P < 0.05), fasting blood glucose (P < 0.01), C-peptide (P < 0.05 and P < 0.01, male vs. female), insulin resistance index (P < 0.05 and P < 0.01, male vs. female), triglycerides (P < 0.05), apolipoprotein B (apoB) (P < 0.01 and P < 0.05, male vs. female), and apoA1/apoB (P < 0.01) were significantly higher in the nondiabetic offspring of type 2 diabetic subjects than in the offspring of nondiabetic subjects. CONCLUSIONS: This study shows a preserved diurnal BP profile and a normal BP level in the nondiabetic glucose-tolerant offspring of type 2 diabetic subjects compared with the offspring of nondiabetic subjects, although the offspring of diabetic patients are characterized by features of the metabolic syndrome. PMID- 10868853 TI - Mortality in patients with childhood-onset type 1 diabetes in Finland, Estonia, and Lithuania: follow-up of nationwide cohorts. AB - OBJECTIVE: To assess mortality of population-based cohorts of childhood-onset type 1 diabetic patients from the Eastern European countries of Estonia and Lithuania and compare this information with recent data from Finland. RESEARCH DESIGN AND METHODS: Estonian (n = 518) and Finnish (n = 5,156) type 1 diabetic cohorts were diagnosed between 1980 and 1994, and the Lithuanian (n = 698) cohort was diagnosed between 1983 and 1994. The mortality of these cohorts was determined in 1995. Life-table analysis, Cox survival analysis with covariates, and standardized mortality ratios (SMRs) were used. Causes of death were analyzed. RESULTS: Survival after 10 years duration of type 1 diabetes was similar in Estonia (94.3%) and Lithuania (94.0%), but much higher in Finland (99.1%). In the Cox survival analysis with covariates, the country of origin and age at diagnosis were found to be significant predictors of mortality. The SMR for the Estonian cohort was 4.35 (95% CI 2.25-7.61), the highest for the Lithuanian cohort was 7.55 (4.89-11.15), and the lowest for the Finnish cohort was 1.62 (1.10-2.28). The most common cause of death in Estonia and Lithuania was diabetic ketoacidosis (DKA), and in Finland, it was violent causes. No deaths from late complications of diabetes have been documented so far in any of the three countries. CONCLUSIONS: Our results demonstrate a high rate of short-term deaths due to DKA and inferior survival of childhood-onset type 1 diabetic patients in Estonia and Lithuania compared with Finland. In Finland, the survival of childhood-onset type 1 diabetic patients has improved and is only slightly inferior to that of the background population. PMID- 10868854 TI - Use of the oral glucose tolerance test to assess insulin release and insulin sensitivity. AB - OBJECTIVE: The oral glucose tolerance test (OGTT) has often been used to evaluate apparent insulin release and insulin resistance in various clinical settings. However, because insulin sensitivity and insulin release are interdependent, to what extent they can be predicted from an OGTT is unclear. RESEARCH DESIGN AND METHODS: We studied insulin sensitivity using the euglycemic-hyperinsulinemic clamp and insulin release using the hyperglycemic clamp in 104 nondiabetic volunteers who had also undergone an OGTT. Demographic parameters (BMI, waist-to hip ratio, age) and plasma glucose and insulin values from the OGTT were subjected to multiple linear regression to predict the metabolic clearance rate (MCR) of glucose, the insulin sensitivity index (ISI), and first-phase (1st PH) and second-phase (2nd PH) insulin release as measured with the respective clamps. RESULTS: The equations predicting MCR and ISI contained BMI, insulin (120 min), and glucose (90 min) and were highly correlated with the measured MCR (r = 0.80, P < 0.00005) and ISI (r = 0.79, P < 0.00005). The equations predicting 1st PH and 2nd PH contained insulin (0 and 30 min) and glucose (30 min) and were also highly correlated with the measured 1st PH (r = 0.78, P < 0.00005) and 2nd PH (r = 0.79, P < 0.00005). The parameters predicted by our equations correlated better with the measured parameters than homeostasis model assessment for secretion and resistance, the delta30-min insulin/delta30-min glucose ratio for secretion and insulin (120 min) for insulin resistance taken from the OGTT. CONCLUSIONS: We thus conclude that predicting insulin sensitivity and insulin release with reasonable accuracy from simple demographic parameters and values obtained during an OGTT is possible. The derived equations should be used in various clinical settings in which the use of clamps or the minimal model would be impractical. PMID- 10868855 TI - Mutations in the genes for hepatocyte nuclear factor (HNF)-1alpha, -4alpha, 1beta, and -3beta; the dimerization cofactor of HNF-1; and insulin promoter factor 1 are not common causes of early-onset type 2 diabetes in Pima Indians. AB - OBJECTIVE: Maturity-onset diabetes of the young (MODY) is a genetically heterogeneous subtype of type 2 diabetes characterized by an early age at onset and autosomal dominant inheritance. MODY can result from heterozygous mutations in at least five genes. The purpose of this study was to determine whether alterations in known MODY genes and two MODY candidate genes contribute to the development of early-onset type 2 diabetes in Pima Indians. RESEARCH DESIGN AND METHODS: The coding regions of the known MODY genes hepatocyte nuclear factor (HNF)-1alpha, HNF-4alpha, HNF-1beta, and insulin promoter factor 1 and the coding regions of two MODY candidate genes, HNF-3beta and the dimerization cofactor of HNF-1, were sequenced in genomic DNA from Pima Indians. The primary "affected" study population consisted of 46 Pima Indians whose age at onset of type 2 diabetes was < or =20 years. DNA sequence variants identified in the affected group were then analyzed in a group of 80 "unaffected" Pima Indians who were at least 40 years old and had normal glucose tolerance. RESULTS: A total of 11 polymorphisms were detected in these genes. However, none of the polymorphisms differed in frequency among Pima Indians with an early age at onset of diabetes compared with older Pima Indians with normal glucose tolerance. CONCLUSIONS: Mutations in these known MODY or MODY candidate genes are not a common cause of early-onset diabetes in Pima Indians. PMID- 10868857 TI - The influence of treatment modality and ethnicity on attitudes in type 2 diabetes. AB - OBJECTIVE: The study examines diabetes attitude differences by treatment modality (insulin vs. no insulin), race/ethnicity, and the interaction of these two variables for people with type 2 diabetes. RESEARCH DESIGN AND METHODS: Data were collected with the Diabetes Care Profile (DCP), an instrument that assesses psychosocial factors related to diabetes. Participants (n = 672) were recruited in the metropolitan Detroit, Michigan, area from 1993 to 1996. A total of 68% of these participants were African-Americans with type 2 diabetes, and 32% were Caucasians with type 2 diabetes. Analyses of covariance were performed to examine the effects of race/ethnicity, treatment, and their interaction for each DCP scale. RESULTS: The four patient categories (two ethnicities by two treatment modalities) differed by age, years with diabetes, education, and sex distribution. Treatment modality had a significant effect on 6 of the 16 DCP scales (Control, Social and Personal Factors, Positive Attitude, Negative Attitude, Self-Care Ability, and Exercise Barriers). Ethnicity was a significant effect for three scales (Control, Support, and Support Attitudes). The interaction of race/ethnicity and treatment modality was a significant effect for two related attitude scales (Positive Attitude and Negative Attitude). CONCLUSIONS: The results suggest that attitudes toward diabetes are similar for African-American and Caucasian patients with type 2 diabetes. The results also suggest that treatment modality has a greater effect on attitudes than either race/ethnicity or the interaction effect. However, Caucasian patients using insulin differed from the other patient groups by having the least positive and the most negative attitudes regarding diabetes. PMID- 10868856 TI - Recovery of cognitive function and mood after severe hypoglycemia in adults with insulin-treated diabetes. AB - OBJECTIVE: Acute hypoglycemia in humans impairs cognitive functions and alters mood states. The time required for cognitive functions and moods to return to normal after an acute episode of severe hypoglycemia is unknown. RESEARCH DESIGN AND METHODS: Cognitive functions and moods were studied prospectively in 20 subjects with insulin-treated diabetes who had recently experienced a spontaneous episode of severe hypoglycemia ("hypo" subjects) and 20 matched control subjects with insulin-treated diabetes who had not experienced severe hypoglycemia during the preceding year. The hypo subjects had a history of a greater number of episodes of severe hypoglycemia (P = 0.000). Cognitive function tests and mood scales were administered at 1.5, 9, and 30 days after the severe hypoglycemia and at similar intervals for the control subjects. RESULTS: For most of the cognitive tests, no evidence of a "hangover" effect of the acute hypoglycemia on cognitive function was observed (P > 0.05). A trend was noted for levels of hedonic tone (P = 0.082) and energetic arousal (P = 0.053) to improve with time in the hypo subjects but not in the control subjects. However, the hypo subjects had chronically elevated levels of depression (P = 0.011) and anxiety (P = 0.049) and persistently performed more poorly in several cognitive tests, such as the Digit Symbol Test (P = 0.009) and the Stroop Task (P = 0.007). CONCLUSIONS: These results suggest that, in general, recovery from any acute cognitive decrement after severe hypoglycemia was complete by 1.5 days. The cognitive decrements and altered mood states noted in the hypo subjects may be persistent and may be a consequence of previous exposure to recurrent episodes of severe hypoglycemia. PMID- 10868858 TI - The pattern of dyslipidemia among urban African-Americans with type 2 diabetes. AB - OBJECTIVE: To analyze lipid profiles from a large sample of African-American patients with type 2 diabetes who receive care at an urban outpatient diabetes clinic. RESEARCH DESIGN AND METHODS: Fasting serum lipid profiles of 4,014 African-Americans and 328 Caucasians with type 2 diabetes were retrieved from a computerized registry. American Diabetes Association criteria were applied to classify LDL cholesterol, HDL cholesterol, and triglyceride (TG) levels into risk categories. The proportion of patients who had none, one, two, and three lipoprotein concentrations outside of recommended clinical targets was examined. Multiple logistical regression analyses were performed to determine the influence of sex and race on the probability of having a lipid level outside of the recommended target. RESULTS: The percentages of African-Americans with high-, borderline-, and low-risk LDL cholesterol concentrations were 58, 26, and 16%, respectively, and the percentages for Caucasians were 54, 29, and 16%, respectively (P = 0.51). For HDL cholesterol, 41, 33, and 26% of African Americans were in the high-, borderline-, and low-risk categories, respectively, compared with 73, 18, and 9% of Caucasians, respectively (P < 0.0001). Nearly 81% of African-Americans had TG concentrations that were in the low-risk category compared with only 50% of Caucasians. More women than men had high-risk LDL and HDL cholesterol profiles. The most common pattern of dyslipidemia was an LDL cholesterol level above target combined with an HDL cholesterol level below target, which was detected in nearly 50% of African-Americans and 42% of Caucasians. African-Americans had lower odds of having an HDL cholesterol or TG level outside of target. African-American women, compared to men, had greater probabilities of having abnormal levels of LDL and HDL, but a lower likelihood of having a TG level above goal. CONCLUSIONS: In a large sample of urban type 2 diabetic patients receiving care at a diabetes treatment program, race and sex differences in serum lipid profiles were present. Because hypertriglyceridemia was rare among African-American subjects, interventions will need to focus primarily on improving their LDL and HDL cholesterol levels. Further studies are required regarding how to best adapt these observed differences into more effective strategies to optimize lipid levels for this population of diabetic patients and to determine whether similar patterns of dyslipidemia occur in other clinical settings. PMID- 10868859 TI - Development of a health status measure for older African-American women with type 2 diabetes. AB - OBJECTIVE: To develop a health status measure in older African-American women with type 2 diabetes. RESEARCH DESIGN AND METHODS: African-American women, age > or =40 years with type 2 diabetes, were recruited from central North Carolina to participate in three sequential phases: 1) Seven focus groups were convened and transcripts evaluated to generate questions and identify plausible domains; 2) Ten one-on-one cognitive response interviews were performed to ensure clarity and cultural appropriateness of the questions; and 3) 217 women participated in psychometric evaluation to establish the internal consistency and validity of the instrument. RESULTS: Three broad categories--mental, physical, and social well being--captured important issues generated during the focus groups. "My diabetes" was added during the cognitive response interviews as a way of separating the impact of diabetes from coexisting issues that affect health status. The response option was changed from a six- to a four-point Likert scale to accommodate subject preference. Using principal components and subsequent promax rotation, we identified two hierarchical domains (mental and social well-being) and a physical symptom index. The internal consistency (Cronbach's alpha) of the mental and social well-being subscales are 0.83 and 0.93, respectively. A priori hypothesized correlations between subscales along with each subscale and glycated hemoglobin, diabetes duration, physical activity, and a perceived health competence scale helped establish the construct validity of the instrument. CONCLUSIONS: A culturally appropriate disease-specific health status measure for older African-American women with type 2 diabetes has been developed. We have established the internal consistency, construct validity, and factor analytic properties of the measure. This measure should prove useful for investigators who seek a health status instrument that addresses issues germane to African-American women with type 2 diabetes. PMID- 10868860 TI - Is "sugar" the same as diabetes? A community-based study among rural African Americans. AB - OBJECTIVE: To determine if differing beliefs about high blood glucose exist and are associated with blood glucose control among rural African-Americans. RESEARCH DESIGN AND METHODS: A community-based sample of rural African-Americans completed a survey, and a subsample underwent a subsequent screening that included glucose and GHb measurement. Participants were asked if they thought they had diabetes or sugar-diabetes on the survey; "sugar" was added to the screening along with specific questions about this condition. RESULTS: A total of 1,031 people completed the survey, and 403 the screening exam. The total prevalence of diabetes was 13.6% for men and 15.5% for women. Among those who reported having one of the three conditions, 64% said they had diabetes, 7% sugar-diabetes, and 29% sugar. There was a discrepancy between the survey and screening in that 31% of subjects who answered "yes" to whether they had sugar at the screening had answered "no" to the survey question about diabetes. Subjects who believed they had sugar felt their condition was less serious and had higher glucose levels than those who said they had diabetes. CONCLUSIONS: Diabetes was very common in this population. Over one-fourth of those with diabetes believed they had the condition "sugar." Efforts are needed to improve control of diabetes in this population and should consider these disparate health beliefs. PMID- 10868861 TI - Metabolic and immunologic features of Chinese patients with atypical diabetes mellitus. AB - OBJECTIVE: To determine whether atypical diabetes mellitus (ADM) is present in the Chinese population in Hong Kong. RESEARCH DESIGN AND METHODS: The records of Chinese patients who attended the Diabetes Clinic at Queen Mary Hospital were reviewed. We identified 11 patients who initially presented with acute diabetic ketoacidosis but subsequently displayed clinical features more typical of type 2 diabetes. Metabolic studies and HLA typing were performed to characterize this group of Chinese patients with ADM. RESULTS: C-peptide response of the patients with ADM 1 h after a standard meal was intermediate between that of type 1 diabetic patients (matched for age and duration of diabetes) and that of nondiabetic control subjects (matched for age and BMI) (analysis of variance, P = 0.02). Insulin sensitivity measured by a short insulin tolerance test was not significantly different between patients with ADM and their matched nondiabetic control subjects. HLA typing showed that none of the patients with ADM had the DR3 allele and that the frequency of DR9 was not increased. Only one patient had significantly increased levels of antibodies to GAD and islet cell antigen 512. CONCLUSIONS: ADM, which was first described in African-Americans, is seen also in Chinese subjects. These patients have significant residual C-peptide secretory capacity and should not be misdiagnosed and treated as patients with type 1 diabetes with life-long insulin therapy. PMID- 10868862 TI - Evaluation of HbA1c determination methods in patients with hemoglobinopathies. AB - OBJECTIVE: To evaluate commercially available determination methods for HbA1c in patients with hemoglobin variants. RESEARCH DESIGN AND METHODS: HbA1c values were determined with various commercially available methods, including ion-exchange high-performance liquid chromatography (HPLC), boronate affinity assay, and immunoagglutination in patients with the hemoglobin mutations Hb Graz, Hb Sherwood Forest, Hb O Padova, Hb D, and Hb S. RESULTS: The effect of hemoglobinopathies on glycohemoglobin measurements was highly method dependent. The HPLC methods for HbA1c determination lacked the resolution necessary to differentiate hemoglobin variants. They demonstrated additional peaks in the chromatograms and HbA1c results either too low or too high compared with the nondiabetic reference range. With all immunoassays, Hb Graz demonstrated falsely low values. The other hemoglobinopathies in our study caused falsely low and/or high HbA1c results in immunoagglutination methods. The boronate affinity method showed values in an acceptable range for all hemoglobin variants. CONCLUSIONS: Because of the local occurrence of Hb variants and the ethnic origin of a given population, every individual laboratory must establish and validate its own assay method. In managing diabetic patients, knowledge of hemoglobinopathies influencing HbA1c determination methods is essential because hemoglobin variants could cause mismanagement of diabetes resulting from false HbA1c determinations. PMID- 10868863 TI - A telemedical approach to the screening of diabetic retinopathy: digital fundus photography. AB - OBJECTIVE: The importance of screening for diabetic retinopathy has been established, but the best method for screening has not yet been determined. We report on a trial of assessment of digital photographs by telemedicine compared with standard retinal photographs of the same fields and clinical examination by ophthalmologists. RESEARCH DESIGN AND METHODS: A total of 129 diabetic inpatients were screened for diabetic retinopathy by slit-lamp biomicroscopy performed by an ophthalmologist and by two-field 50 degrees non-stereo digital fundus photographs assessed by six screening centers that received the images by electronic mail. Conventional 35-mm transparencies of the same fields as the digital photographs were assessed by a retinal specialist and served as the reference method for detection of diabetic retinopathy. Slit-lamp biomicroscopy was the reference method for the detection of macular edema. RESULTS: The prevalence of any form of diabetic retinopathy was 30% (n = 35); of sight-threatening retinopathy including macular edema, the prevalence was 6% (n = 7). The assessment of digital images by the six screening centers resulted in a median sensitivity of 85% and a median specificity of 90% for the detection of moderate nonproliferative or sight threatening diabetic retinopathy. Clinically significant macular edema (n = 4) was correctly identified in 15 of the 24 grading reports. An additional seven reports referred the patients for further investigation because of concurrent diabetic retinopathy. CONCLUSIONS: Telescreening for diabetic retinopathy by an assessment of two-field 50 degrees non-stereo digital images is a valid screening method. Although detection of clinically significant macular edema using biomicroscopy is superior to digital or standard non-stereo photographs, only few patients with sight-threatening diabetic retinopathy are missed. PMID- 10868864 TI - Improved control of mealtime glucose excursions with coadministration of nateglinide and metformin. AB - OBJECTIVE: Nateglinide, a new short-acting D-phenylalanine derivative for treating type 2 diabetes, reduces mealtime blood glucose excursions by physiologic regulation of insulin secretion. This study evaluated the pharmacokinetic and pharmacodynamic interactions of nateglinide and metformin in subjects with type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 12 type 2 diabetic subjects with the following baseline characteristics were enrolled: age, 56 +/- 13 years; BMI, 28.7 +/- 4.5 kg/m2; HbA1c, 8.4 +/- 1.3%; and fasting plasma glucose 13 +/- 2.8 mmol/l. All subjects had been previously treated with glyburide and were switched to metformin monotherapy for 3 weeks before study start. Subjects then randomly received, in combination with 500 mg metformin, either 120 mg nateglinide or placebo before meals for 1 day, followed by the alternate treatment 7 days later. After 1 week of washout from both drugs, subjects received 1 day of open-label nateglinide treatment. Plasma concentrations of glucose, insulin, nateglinide, and metformin were assessed frequently during inpatient periods. RESULTS: Postmeal plasma glucose levels were significantly lower in subjects treated with nateglinide plus metformin than in those treated with either drug alone (P < 0.001), especially after lunch and dinner. Coadministration of nateglinide and metformin did not affect the pharmacokinetics of either drug. All treatments were safe and well tolerated. CONCLUSIONS: Combination therapy with nateglinide and metformin was more effective than either treatment alone and did not result in any pharmacokinetic interactions. Coadministration of nateglinide and metformin appears to be an excellent option for treating patients with type 2 diabetes not controlled with monotherapy. PMID- 10868865 TI - Long-term effects of troglitazone: open-label extension studies in type 2 diabetic patients. AB - OBJECTIVE: To determine the long-term effects of troglitazone as monotherapy or in combination with sulfonylureas or insulin regarding glycemic and lipid measures. RESEARCH DESIGN AND METHODS: Patients who completed one of three double blind studies (a 6-month troglitazone monotherapy study, a 52-week study of troglitazone in combination with micronized glyburide, or a 6-month study of troglitazone in combination with insulin) were allowed to enter open-label extensions of their respective double-blind studies. Troglitazone dose titrations were allowed to a maximum of 600 mg in response to inadequate glycemic control during the open-label phases of troglitazone monotherapy or sulfonylurea combination therapy but not with insulin combination therapy. This article focuses on the effectiveness of the highest dose of troglitazone used in these studies (600 mg daily). Safety data from all patients studied at all doses are also presented. RESULTS: For patients who received a fixed dose of 600 mg troglitazone, mean changes in fasting serum glucose and HbA1c levels from baseline to the end of the open-label phase were -57 mg/dl and -0.4%, respectively (monotherapy); -49 mg/dl and -1.8%, respectively (sulfonylurea combination); and -31 mg/dl and -1.0%, respectively (insulin combination). The proportion of patients achieving an HbA1c level of < or =8% from the combined cohort of all three studies was 54% versus only 19% at baseline. The mean decrease in triglycerides from baseline to the end of the open-label phase was 18% among all patients in the three studies who received a fixed dose of 600 mg troglitazone. Troglitazone was well tolerated in these three open-label studies; a total of 758 patients completed a total exposure of 16,264 patient-months to troglitazone in these three studies with minimal adverse events. CONCLUSIONS: Long-term use of troglitazone alone or in combination with sulfonylureas or insulin is safe and effective in sustaining glycemic control and in reducing hypertriglyceridemia in type 2 diabetic patients. PMID- 10868866 TI - Effects of nicotinamide and intravenous insulin therapy in newly diagnosed type 1 diabetes. AB - OBJECTIVE: To investigate the effect of intravenous insulin therapy combined with nicotinamide in the metabolic control and beta-cell function of newly diagnosed type 1 diabetic subjects in comparison with intensive insulin therapy and nicotinamide alone. RESEARCH DESIGN AND METHODS: A total of 34 newly diagnosed type 1 diabetic patients were included. After the correction of initial metabolic disturbances, subjects were randomly assigned to the following three groups within 72 h after admission: 1) intensive insulin therapy + placebo (C) (n = 12); 2) intensive insulin therapy + nicotinamide, 700 mg three times a day (NIC) (n = 11); and 3) 72-h intravenous insulin followed by intensive insulin therapy + nicotinamide, 700 mg three times a day (NIV) (n = 11). The subjects were monitored for 12 months. GAD, tyrosine phosphatase antibodies, and insulin autoantibodies were measured. C-peptide was measured basally and after 2, 4, 6, 8, and 10 min of 1 mg intravenous glucagon. HbA1c, glucagon, and antibody measurements were determined initially and at 1, 3, 6, 9, and 12 months. RESULTS: HbA1c values declined to normal after treatment was initiated in all groups and remained not significantly different during the follow-up period. We did not find differences between experimental (NIC and NIV) and placebo (C) groups in terms of beta-cell function, considering basal or glucagon-stimulated C-peptide (maximal stimulated C-peptide and area under the curve [AUC] of C-peptide) values during the follow-up period. After pooling data from the NIC and NIV groups (both including nicotinamide) and comparing it with data from the C group, the results remained unchanged. At diagnosis, GAD positivity was observed in 10 of 12, 8 of 11, and 10 of 11 subjects (NS) in the C, NIC, and NIV groups, respectively, and IA2 positivity was observed in 3 of 12, 4 of 11, and 4 of 11 subjects (NS) in the C, NIC, and NIV groups, respectively. Antibody titers displayed a similar behavior in all groups during the follow-up period. CONCLUSIONS: Our pilot study failed to demonstrate that the addition of 72-h intravenous insulin and nicotinamide to conventional intensive insulin therapy produces any beneficial effect in newly diagnosed type 1 diabetic subjects in terms of beta-cell function and metabolic control. PMID- 10868867 TI - Percutaneous electrical nerve stimulation: a novel analgesic therapy for diabetic neuropathic pain. AB - OBJECTIVE: To evaluate the use of percutaneous electrical nerve stimulation (PENS) in the management of patients with painful diabetic peripheral neuropathy. RESEARCH DESIGN AND METHODS: A total of 50 adult patients with type 2 diabetes and peripheral neuropathic pain of >6 months duration involving the lower extremities were randomly assigned to receive active PENS (needles with electrical stimulation at an alternating frequency of 15 and 30 Hz) and sham (needles only) treatments for 3 weeks. Each series of treatments was administered for 30 min three times a week according to a standardized protocol. After a 1 week washout period, all patients were subsequently switched to the other modality. A 10-cm visual analog scale (VAS) was used to assess pain, physical activity, and quality of sleep before each session. The changes in VAS scores and daily requirements for oral analgesic medication were determined during each 3 week treatment period. Patients completed the MOS 36-Item Short-Form Health Survey (SF-36), the Beck Depression Inventory (BDI), and the Profile of Mood States (POMS) before and after completion of each treatment modality. At the end of the crossover study, a patient preference questionnaire was used to compare the effectiveness of the two modalities. RESULTS: Compared with the pain VAS scores before active (6.2 +/- 1.0) and sham (6.4 +/- 0.9) treatments, pain scores after treatment were reduced to 2.5 +/- 0.8 and 6.3 +/- 1.1, respectively. With active PENS treatment, the VAS activity and sleep scores were significantly improved from 5.2 +/- 1.0 and 5.8 +/- 1.3 to 7.9 +/- 1.0 and 8.3 +/- 0.7, respectively. The VAS scores for pain, activity, and sleep were unchanged from baseline values after the sham treatments. Patients' daily oral nonopioid analgesic requirements decreased by 49 and 14% after active and sham PENS treatments, respectively. The post-treatment physical and mental components of the SF-36, the BDI, and the POMS all showed a significantly greater improvement with active versus sham treatments. Active PENS treatment improved the neuropathic pain symptoms in all patients. CONCLUSIONS: PENS is a useful nonpharmacological therapeutic modality for treating diabetic neuropathic pain. In addition to decreasing extremity pain, PENS therapy improved physical activity, sense of well-being, and quality of sleep while reducing the need for oral nonopioid analgesic medication. PMID- 10868868 TI - Is leptin associated with diabetic retinopathy? AB - OBJECTIVE: In advanced stages of diabetic retinopathy, new blood vessels are formed based on undefined mechanisms. Recently, leptin was shown to possess an angiogenic action in vitro and to induce neovascularization in vivo. The aim of the present study was to investigate the relationship between plasma leptin levels and the severity of diabetic retinopathy. RESEARCH DESIGN AND METHODS: There were 70 patients with type 2 diabetes (age 47.9 +/- 9.7 years, BMI 26.4 +/- 3.3 kg/m2) who were seen in a retina outpatient clinic recruited and assigned to subgroups according to the stage of their diabetic retinopathy. There were 66 healthy volunteer subjects matched with the diabetic patients for age, BMI, and sex who served as control subjects (age 46.0 +/- 8.8 years, BMI 27.1 +/- 2.3 kg/m2). Fasting plasma leptin levels were measured. RESULTS: Plasma leptin level of the diabetic patients was not significantly different from the control subjects. In patients with proliferative diabetic retinopathy (n = 17), the mean plasma level of leptin (16.1 +/- 9.2 ng/ml) was significantly higher than that in patients with nonproliferative retinopathy (n = 20) (11.5 +/- 3.5 ng/ml, P = 0.039) or patients without retinopathy (n = 33) (5.8 +/- 3.7 ng/ml, P = 0.001). The mean leptin level in patients with nonproliferative diabetic retinopathy was also significantly higher than that in patients without retinopathy (P = 0.002). CONCLUSIONS: Our results show that the more advanced the diabetic retinopathy, the higher the plasma leptin levels, even after adjusting the leptin levels for BMI. The presence of such a positive correlation need not imply a causal relationship. Nevertheless, previously observed leptin-induced promotion of angiogenesis and neovascularization lends support to the possibility that leptin may play a role in the progression of human diabetic retinopathy to a proliferative phase. This possibility deserves further investigation. PMID- 10868869 TI - Enalapril prevents clinical proteinuria in diabetic patients with low ejection fraction. AB - OBJECTIVE: Clinical proteinuria is a risk factor for both end-stage renal disease and cardiovascular disease. The prevalence of clinical proteinuria, its correlates and predictive value, and the effect of ACE inhibitors in preventing clinical proteinuria in diabetic and nondiabetic patients with left ventricular (LV) dysfunction are unknown. RESEARCH DESIGN AND METHODS: The Studies of Left Ventricular Dysfunction (SOLVD) trials were analyzed to determine the baseline distribution of clinical proteinuria and related cardiovascular risk factors, the effect of baseline proteinuria on the risk of hospitalization for congestive heart failure (CHF) and mortality, and the effect of enalapril in preventing new clinical proteinuria. RESULTS: A total of 5,487 out of 6,797 SOLVD participants (81%) were assessed for proteinuria at baseline. A total of 177 patients (3.2%) had baseline proteinuria. These patients had significantly higher systolic (137 vs. 125 mmHg, P < or = 0.001) and diastolic (83 vs. 77 mmHg, P < or = 0.001) blood pressure levels, a higher prevalence of diabetes (41 vs. 18%, P < or = 0.001), a lower ejection fraction (26.2 vs. 27.3%, P < or = 0.05), and greater degree of CHF (New York Heart Association [NYHA] class III/IV in 22 vs. 10%, P < or = 0.001) than patients without baseline proteinuria. Patients with baseline proteinuria also had higher rates of hospitalization for CHF (relative risk 1.81 [95% CI 1.37-2.41], P = 0.0001) and mortality (1.73 [1.34-2.24], P = 0.0001). Enalapril prevented clinical proteinuria in diabetic patients (0.38 [0.17-0.81], P = 0.0123) but not in nondiabetic patients (1.43 [0.77-2.63], P = 0.2622) without baseline proteinuria. CONCLUSIONS: Clinical proteinuria is an independent predictor of hospitalization for CHF and mortality in diabetic and nondiabetic patients with LV dysfunction. Enalapril significantly reduces the risk of clinical proteinuria in diabetic patients with LV dysfunction. PMID- 10868870 TI - Type 2 diabetes in children and adolescents. American Diabetes Association. PMID- 10868872 TI - American Diabetes Association Annual Meeting, 1999: type 2 diabetes treatment. PMID- 10868871 TI - An economic analysis of interventions for diabetes. AB - The objective of this article is to stratify interventions for diabetes according to their economic impact. We conducted a review of the literature to select articles that performed a cost-benefit analysis for 17 widely practiced interventions for diabetes. A scale for categorizing interventions according to their economic impact was defined. The 17 interventions were classified as follows: 1) clearly cost-saving, 2) clearly cost-effective, 3) possibly cost effective, 4) non-cost-effective, or 5) unclear. Clearly cost-saving interventions included eye care and pre-conception care. Clearly cost-effective interventions included nephropathy prevention in type 1 diabetes and improved glycemic control. Possibly cost-effective interventions included nephropathy prevention in type 2 diabetes and self-management training. Non-cost-effective interventions were not identified. Interventions with unclear economic impact included case management, medical nutrition therapy, self-monitoring of blood glucose, foot care, blood pressure control, blood lipid control, smoking cessation, exercise, weight loss, HbA1c measurement, influenza vaccination, and pneumococcus vaccination. Widely practiced interventions for patients with diabetes can be clearly cost-saving and clearly cost-effective. These practices are attractive from both a medical and an economic perspective. PMID- 10868873 TI - Acceptability of pig xenografts by patients with type 1 diabetes and the general population. PMID- 10868874 TI - Can enterovirus infections explain the increasing incidence of type 1 diabetes? PMID- 10868875 TI - The insulin-plus-sugar method for controlling recently diagnosed type 2 diabetes without hypoglycemia. PMID- 10868876 TI - Increased spontaneous adherence of neutrophils from type 2 diabetic patients with overt proteinuria: possible role of the progression of diabetic nephropathy. PMID- 10868877 TI - Erythrocyte sodium/hydrogen exchange activity and albuminuria in type 1 diabetic families. PMID- 10868878 TI - Factors influencing plasma homocysteine levels in type 2 diabetes. PMID- 10868879 TI - Acute rhabdomyolysis associated with troglitazone. PMID- 10868880 TI - Long-term transdermal estrogen therapy improves lipid profile but not insulin resistance in healthy postmenopausal women. PMID- 10868881 TI - A severe clinical phenotype results from the co-inheritance of type 2 susceptibility genes and a hepatocyte nuclear factor-1alpha mutation. PMID- 10868882 TI - Gestational diabetes alters the male bias for cesarean section. PMID- 10868883 TI - Diabetic gastroparesis improved by percutaneous endoscopic jejunostomy. PMID- 10868884 TI - Severe nonproductive cough and cough-induced stress urinary incontinence in diabetic postmenopausal women treated with ACE inhibitor. PMID- 10868885 TI - Can mobile cellular phones affect functioning of implantable insulin pumps? PMID- 10868886 TI - Usual delay in sample processing can modify gestational diabetes screening. PMID- 10868887 TI - Operational errors cause inaccurate glucose results. PMID- 10868888 TI - Poor performance of American Diabetes Association criteria in women with gestational diabetes. PMID- 10868889 TI - A role for gestational diabetes in the excess maternal transmission of type 2 diabetes? PMID- 10868890 TI - Recent commencement of dialysis is a risk factor for lower-extremity amputation in a high-risk diabetic population. PMID- 10868891 TI - Response to editorial by Flegal. PMID- 10868892 TI - Controlled trials of HbA1c measurements. PMID- 10868893 TI - Cancer in a patient receiving IGF-I therapy. PMID- 10868894 TI - Cotherapy with recombinant human IGF-I and insulin improves glycemic control in type 1 diabetes. PMID- 10868895 TI - Therapy of Helicobacter pylori: current status and issues. PMID- 10868896 TI - Acid suppression: optimizing therapy for gastroduodenal ulcer healing, gastroesophageal reflux disease, and stress-related erosive syndrome. PMID- 10868897 TI - Motility-modifying agents and management of disorders of gastrointestinal motility. PMID- 10868898 TI - Treatment of gastrointestinal infections. PMID- 10868899 TI - Therapy of inflammatory bowel disease. AB - In the last decade, substantial gains have been made in the treatment of inflammatory bowel disease (IBD). Refinements in drug formulation have provided the ability to target distinct sites of delivery, enhancing the safety and efficacy of older agents. Immunosuppressive agents beyond corticosteroids have assumed a routine part in the care of patients with IBD. Moreover, as the century closes, we stand at the threshold of unprecedented advances in knowledge of the pathogenesis of ulcerative colitis and Crohn's disease. Simultaneous progress in biotechnology has fostered the development of new agents that strategically target pivotal processes in disease pathogenesis. This review covers agents currently used in the treatment of IBD and seeks to provide an overview of emerging therapies. PMID- 10868900 TI - Antiviral chemotherapy for the treatment of hepatitis B virus infections. AB - Approximately 5% of the world's human population have an increased risk for developing liver cancer and cirrhosis as a direct consequence of chronic infection with the hepatitis B virus (HBV). Antiviral chemotherapy remains the only option for controlling infection in these individuals, for whom the current licensed hepatitis B vaccines provide no benefit. Interferon (IFN)-alpha has proven benefit in a well-defined group of those with hepatitis B but has made little impact on the global burden of chronic liver disease. The development of more effective chemotherapy for treatment of chronic hepatitis B infection has proven to be extremely challenging, the result of both virus- and host-dependent factors, which will be reviewed in this article. Past attempts to treat chronic hepatitis B infection using nucleoside analogues were disappointing, but more recently, several nucleoside (or nucleotide) analogues have been identified that are potent and selective inhibitors of HBV replication. These agents fall into two broad categories: (1) nucleoside/nucleotides that have modified sugar residues in either cyclic or acyclic configurations and (2) stereoisomers of nucleosides in the "unnatural" L-configuration. Of the analogues that have been used clinically, representatives of the first category are purine derivatives, e.g., adefovir dipivoxil and famciclovir, whereas representatives of the second category are pyrimidine derivatives, such as lamivudine. PMID- 10868901 TI - Current therapy for chronic hepatitis C. AB - Chronic hepatitis C virus infection is common in the United States with an estimated prevalence of 2.7 million persons. Fortunately, the incidence of new infections has markedly declined in recent years and the natural history of chronic hepatitis usually only results in significant progression after several decades of infection. However, the majority of chronically infected patients acquired their infections more than 20 years ago; these patients with long standing chronic hepatitis are now presenting in increasing numbers with decompensated cirrhosis and the need for liver transplantation. Cirrhosis caused by chronic hepatitis C is now the most common indication for liver transplantation. Interferon monotherapy became clinically available 10 years ago but resulted in sustained improvement in liver disease and durable loss of detectable virus in fewer than 10% of treated patients. The recent use of the combination of interferon with the nucleoside analogue ribavirin for 6-12 months results in a sustained virological response in 30%-40% of previously untreated patients. The response to this combination therapy is also excellent in patients who had initially responded to interferon monotherapy and later relapsed. Furthermore, some recent studies suggest that a small proportion of patients who failed to respond to a prior course of interferon (primarily noncirrhotic patients with low levels of virus and genotypes other than 1) may also benefit from retreatment with this combination. PMID- 10868902 TI - Colorectal cancer prevention and treatment. PMID- 10868903 TI - Endoscopic practice at the start of the new millennium. PMID- 10868904 TI - Laparoscopic minimal-access surgery: where are we now? Where are we going? PMID- 10868905 TI - Lysophosphatidic acid and sphingosine 1-phosphate: two lipid villains provoking cardiovascular diseases? AB - Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (SIP) are potent bioactive lipids with specific and multiple effects on cells of the vessel wall and blood platelets. In this paper we suggest that these lipid molecules are involved in atherogenesis, pathological vasoconstriction, plaque rupture, and intravascular thrombus formation, which leads us to propose new strategies for the prevention and therapy of cardiovascular diseases. The conclusions are hypothetical, in that the studies were so far mainly carried out on isolated cells or cultured cells in vitro and the results were extrapolated to the situation in vivo. PMID- 10868906 TI - A replicator was not involved in the origin of life. AB - Many scientific theories of the origin of life suggest that life began with the spontaneous formation of a replicator (a self-copying organic polymer) within an unorganized chemical mixture, or "soup." A profound difficulty exists, however, with the idea of RNA, or any other replicator, at the start of life. Existing replicators can serve as templates for the synthesis of additional copies of themselves, but this device cannot be used for the preparation of the very first such molecule, which must arise spontaneously from an unorganized mixture. The formation of an information-bearing homopolymer through undirected chemical synthesis appears very improbable. The difficulties involved in such a synthesis are illustrated by considering the prospects for the assembly of a polypeptide of L-alpha-amino acids, based on the contents of the Murchison meteorite as an example of a mixture of abiotic origin. In that mixture, potential replicator components would be accompanied by a host of interfering substances, which include chain terminators (simple carboxylic acids and amines), branch-formers, D amino acids, and many classes of substances for which incorporation would disrupt the necessary structural regularity of the replicator. Laboratory experiments dealing with the nonenzymatic synthesis of biopolymers have not addressed the specificity problem. The possibility that formation of the first replicator took place through a very improbable event cannot be excluded, but greater attention should be given to metabolism-first theories, which avoid this difficulty. PMID- 10868907 TI - The p66shc protein: a mediator of the programmed death of an organism? AB - Recently knockout of the gene encoding an adaptor protein (p66shc) was shown both to prolong the life span of an animal and to prevent apoptosis of cells in response to added H2O2 (Migliaccio et al. [1999] Nature 402, 309-313). A hypothesis is put forward in which p66shc is assumed to be involved in phenoptosis, i.e., programmed death of an organism, mediated by the reactive oxygen species-dependent massive apoptosis in an organ of vital importance. The reactive oxygen species are suggested to oxidize phosphatidyl serine in the inner leaflet of the cell plasma membrane, resulting in appearance of this phospholipid in the outer membrane leaflet, an effect recognized by a special receptor and causing the p66shc phosphorylation at a serine residue. Serine-phosphorylated p66shc there is proposed to block mitosis and initiate apoptosis. The large-scale apoptosis leads to phenoptosis and, hence, shortens the life span of the organism. PMID- 10868908 TI - Rational and "irrational" design of proteins and their use in biotechnology. AB - A familiar refrain within industrial circles is better, faster, and cheaper. Efforts to place this mantra into practice within the biotechnology industry has brought a focus on protein engineering as one method to create new products quickly and inexpensively. Typically, protein engineering has utilized either rational design or combinatorial methods, both of which have been explored and improved in recent years. Continued advancement in these two areas and their application to an increasing list of industrially and medically important processes mean that the number of "synthetic" proteins displacing old technologies is likely to grow at an amazing rate over the next few years. We discuss some of the technologies available for protein redesign and illustrate these with examples from the biocatalysis, biosensor, and therapeutic fields. PMID- 10868909 TI - Genetic and molecular mechanisms of age regulation (homeostasis) of blood coagulation. AB - Blood coagulation plays a critical role not only in hemostasis but also in many physiological and pathological conditions. Epidemiological studies have shown that blood coagulation capacity in humans increases with age. Towards understanding the underlying mechanisms, the age regulation of factor IX, a key blood coagulation factor, was extensively studied. A series of human factor IX minigenes, consisting of various components of the human factor IX gene, were constructed and subjected to systematic analyses with HepG2 cells in culture and over the entire life span of transgenic mice. These studies identified critical gene structures that are essential for the unique age-dependent expression patterns of the human factor IX gene--one acting by stabilizing gene transcription and another increasing the amount of mRNA present, presumably by augmenting mRNA stability. These studies have set the stage for analyzing the overall age-based regulatory mechanisms of blood coagulation. PMID- 10868910 TI - FGF receptor mutations: dimerization syndromes, cell growth suppression, and animal models. AB - This review describes recent progress in the field of fibroblast growth factor receptors (FGFRs) with an emphasis on the role of FGFR mutants in skeletal malformations. This family of four receptors contains the most frequent germline mutations in humans. More than 75 mutations have been recorded, which account for more than seven skeletal syndromes. The common cause for all the mutant phenotypes is gain-of-function by receptor activation through three major mechanisms: receptor dimerization, kinase activation, and increased affinity for FGF. The severity of the disease is correlated with both the extent of receptor activation and the specific tissue in which the mutant receptor form is expressed. Paradoxically, the consequence of receptor activation is inhibition of chondrocyte cell growth through signaling pathways that are cell-type specific. The structure of the FGFR-FGF complex and its possible ternary complex with heparin explain the mechanism of receptor dimerization in the ectodomain and the possible contribution by some of the mutations to this process. Analysis of FGFR3 mutant mice produced by gene targeting as models for human disease, and studies in cell lines, have begun to delineate the novel signaling pathways of FGFR3 and to define possible targets for therapy. PMID- 10868911 TI - Cloning and characterization of a novel cDNA sequence encoding the precursor of a novel venom peptide (BmKbpp) related to a bradykinin-potentiating peptide from Chinese scorpion Buthus martensii Karsch. AB - Based on the amino acid sequence of a bradykinin-potentiating peptide (Bpp) (peptide K-12) from scorpion Buthus occitanus, a full-length cDNA sequence encoding the precursor of a novel venom peptide (named BmKbpp) related to this Bpp, has been isolated and analyzed. The cDNA encodes a precursor of 72 amino acid residues, including a signal peptide of 22 residues and an extra Arg-Arg-Arg tail at the C-terminal end of the precursor, which have to be removed in the processing step. The C-terminal region (21 residues) of the precursor is homologous (57% identical) with the sequence of peptide K-12. Thus, according to the primary structure of the BmKbpp precursor, there may be a propeptide between the signal peptide and the putative mature BmKbpp at the C-terminal region of the precursor. PMID- 10868912 TI - Mechanism of dihydrolipoate stimulation of the mitochondrial permeability transition: effect of different respiratory substrates. AB - The stimulation of the mitochondrial permeability transition (MPT) by dihydrolipoate (DHLA) was studied in rat liver mitochondria in the presence of different respiratory substrates. The Ca2+ threshold for the induction of MPT was lowest for pyruvate, followed by 2-hydroxybutyrate, 2-oxoglutarate, glutamate plus malate, and succinate plus rotenone, both in the presence and absence of DHLA. DHLA was not able to induce MPT in the absence of Ca2+, in the presence of cyclosporin A, or rotenone with pyridine nucleotide-dependent substrates. The difference in sensitivity of MPT to DHLA with various substrates was correlated with the redox state of pyridine nucleotides but not the redox state of glutathione. These findings demonstrate that DHLA induced MPT pore opening through the P-site thiol. The similarities between the effect of DHLA and that of production of reactive oxygen species found in model experiments suggest that DHLA stimulates MPT by production of reactive oxygen species that exhaust the antioxidant defence. PMID- 10868913 TI - Osteopontin is induced by nitric oxide in RAW 264.7 cells. AB - Nitric oxide (NO) produced by macrophages is thought to contribute to various pathological conditions. Osteopontin (OPN) is a phosphorylated glycoprotein produced principally by macrophages. OPN inhibits inducible nitric oxide synthase (iNOS), which generates large amounts of NO production. However, the relationship between NO and endogenous OPN in activated macrophages has not yet been elucidated. We therefore examined expression of endogenous iNOS and OPN in a murine macrophage cell line, RAW 264.7 cells, by treating the cells with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). Treatment of cells with LPS and IFN-gamma resulted in an increase of iNOS mRNA to maximum at 12 h after stimulation. In contrast, OPN mRNA was induced more slowly than iNOS mRNA. Induction of both iNOS and OPN mRNA in RAW 264.7 cells was markedly suppressed by addition of the specific iNOS inhibitor S-2-aminoethyl isothiourea dihydrobromide. The NOS inhibitor NG-methyl-L-arginine also suppressed induction of OPN mRNA but hardly affected iNOS mRNA expression. The NO-releasing agent spermine-NONOate but not peroxynitrite enhanced induction of OPN mRNA. These results suggest that NO directly up-regulates the endogenous OPN in macrophages stimulated with LPS and IFN-gamma. This up-regulation of endogenous OPN may represent a negative feedback system acting to reduce iNOS expression. PMID- 10868914 TI - Reverse transcription of a naturally occurring nonretroviral RNA produces a precise deletion in the majority of its cDNA products. AB - A precise, reproducible deletion made during in vitro reverse transcription of RNA2 from the icosahedral positive-stranded Helicoverpa armigera stunt virus (Tetraviridae) is described. The deletion, located between two hexamer repeats, is a 50-base sequence that includes one copy of the hexamer repeat. Only the Moloney murine leukemia virus reverse transcriptase and its derivative Superscript I, carrying a deletion of the carboxy-terminal RNase H region, showed this response, indicating a template-switching mechanism different from one proposed that involves a RNase H-dependent strand transfer. Superscript II, however, which carries point mutations to reduce RNase H activity, does not cause a deletion. A possible mechanism involves the enzyme pausing at the 3' side of a stem-loop structure and the 3' end of the nascent DNA strand separating from the template and reannealing to the upstream hexamer repeat. PMID- 10868916 TI - The fundamental role of clinical research. PMID- 10868915 TI - A short region containing an AP-1 binding site is essential for transforming growth factor-beta-induced c-jun gene expression in osteoblastic cells. AB - Transforming growth factor-beta (TGF-beta) is a multifunctional regulatory peptide that elicits different responses in different cell types. Much remains unknown about the pathway of intracellular TGF-beta signal transduction, but TGF beta is known to induce expression of several genes by way of the transcription factor AP-1. We studied the mechanism that mediates TGF-beta-induced gene expression of c-jun, a component of AP-1, in MC3T3-E1 osteoblastic cells. To map in detail the corresponding responsive elements in the rat c-jun promoter, we generated a series of 5' deletion promoter/luciferase reporter gene constructs. Transient cell transfection assays identified the region located between positions -79 and -59 as being critical for the TGF-beta response and for the basal activity of the promoter. Gel mobility shift assays indicated specific binding of nuclear proteins to this 21-bp region of the c-jun promoter containing an AP-1 binding site. These results show that the AP-1-dependent mechanism is involved in TGF-beta-induced increase of c-jun induction, suggesting positive autoregulation of AP-1. PMID- 10868917 TI - The future in hypertrophic cardiomyopathy: important clues and potential advances from an understanding of the genotype phenotype relationship. AB - Hypertrophic cardiomyopathy is a familial heart muscle disorder caused by sarcomeric contractile protein gene abnormalities, nine of which have been recognised to date. The condition has been defined clinically and pathologically as a syndrome of unexplained myocardial hypertrophy, which is associated with characteristic pathophysiological (diastolic dysfunction, ischaemia, altered vascular responses) and pathological (myocyte disarray, increased loose connective tissue, small vessel disease) abnormalities. Disease expression is variable. Preliminary observations suggest that the marked clinical and pathological heterogeneity, which has long caused controversy in hypertrophic cardiomyopathy, is at least, in part, a function of the disease-causing gene. The clinical syndrome of hypertrophic cardiomyopathy can then be seen as nine or more different, but related diseases. Greater understanding of these diseases will require broader application of DNA diagnostic testing, as well as careful and accurate evaluation of the clinical/pathological phenotype of hypertrophic cardiomyopathy. PMID- 10868918 TI - Rapid coronary artery disease progression and angiographic stenosis morphology. AB - It is established that the progression of coronary artery disease is neither linear nor predictable. The unpredictable and often episodic nature of coronary disease progression can be explained by rapid increase of stenosis severity due to thrombosis, which occurs as a complication of the atherogenic process. Rapid coronary stenosis progression is most often responsible for the acute clinical manifestations of coronary artery disease, i.e. sudden cardiac death, acute myocardial infarction and unstable coronary syndromes. Recently, it has been shown that stenosis progression, whether clinically "silent" or associated with acute coronary events, is a strong predictor of cardiovascular risk. Atheromatous plaques associated with rapid coronary artery disease progression have well defined anatomo-pathological characteristics and are usually termed "vulnerable" or "unstable", terms which indicate both their propensity to acute disruption and increased thrombogenicity that may lead to the development of acute coronary events. Plaques have been regarded, in the past, as being inert and staying almost unchanged for years. However, they are very active entities. The fibrous cap is in a balance between smooth muscle cells producing collagen and the macrophages degrading collagen. The thickness of the cap depends on the relative activity of those two components and there is, therefore, a danger of the fibrous cap rupturing. Although only a relatively small proportion of all coronary artery lesions in patients with angina pectoris undergo complications that lead to fibrous cap disruption and acute coronary events, these stenoses are responsible for the majority of cases of serious coronary events. Thus the identification of vulnerable plaques that may lead to increased risk of coronary events will most certainly help in the rational management of patients with coronary artery disease. Angiographic studies have indicated that "complex" lesion morphology is associated with increased risk of myocardial infarction and "ulcerated" plaques identify vulnerable lesions. We therefore reasoned that the identification of angiographically complex coronary stenoses could provide a valuable marker of cardiovascular risk in relation to rapid disease progression. Our group sought to investigate the role of angiographically complex lesions as a marker of rapid disease progression in different clinical settings. We took advantage of the fact that patients with stable angina pectoris requiring routine myocardial revascularisation in our institution are put on waiting lists. We observed that complex lesions progressed more than smooth stenoses of similar severity both in patients presenting with stable angina and in patients presenting with unstable angina. Why complex plaques should be particularly vulnerable to rapid stenosis progression is speculative. In this paper we discuss the possible mechanisms that may explain an association between complex stenosis and acute coronary events. PMID- 10868919 TI - Three-dimensional echocardiography: where we are, where we are going. AB - Three-dimensional reconstruction of the heart has been an important research goal ever since the introduction of two-dimensional echocardiography. Several directions have been followed. Most approaches towards three-dimensional echocardiography are off-line and are based on the sequential rotational scanning and acquisition of multiple cross-sectional images together with their spatial position and orientation using internal coordinate reference systems. From the reconstructed volumetric data set electronic slicing can be performed which allows any-plane and paraplane echocardiography. The availability and versatility using the volumetric data set permits the retrieval of an infinite number of cardiac cross-sections which allow more accurate and reproducible measurements of valve areas, masses and cavity volumes by obviating geometric assumptions. The application of algorithms based on light reflection to the grey scale data provides tissue-depicting information allowing for dynamic volume-rendered display in projection, up till now unavailable in cardiology. This capability decreases variability both in the quality and interpretation of complex pathology among investigators. Emerging clinical experience indicates the strong potential of three-dimensional echocardiography in qualitative and quantitative diagnostic appraisal of various cardiac problems. While the technique is ready for clinical applications, its widespread use can be facilitated by a number of improvements. Advances in computer technology can be applied to three-dimensional echocardiography offering an exciting opportunity to employ virtual reality and simulation of interventional and surgical procedures, to predict results and plan appropriate therapy. In the future new physiologic parameters will provide additional information and will allow us to address new clinical questions. PMID- 10868920 TI - Regional distribution and timing of wall motion abnormalities during echo dipyridamole stress test in patients with stable angina: the elusive link between coronary stenoses and myocardial ischemia. AB - BACKGROUND: Classic experimental studies have shown that in the presence of a flow-limiting coronary artery stenosis, myocardial ischemia during metabolic or pharmacological arteriolar vasodilation causes wall motion abnormalities, which precede electrocardiographic (ECG) changes in the myocardial regions supplied by the stenotic branch. The aim of this study was to establish whether in patients with chronic stable angina the regional distribution of wall motion changes and sequence of ischemic events are similar to that observed in experimental models, as currently believed. METHODS: The study population consisted of 20 men and 4 women (mean age 59 +/- 10 years) who were recruited on the basis of the following criteria: 1) a history of chronic stable angina without clinical and instrumental evidence of previous myocardial infarction; 2) reproducible positive exercise tests for ECG myocardial ischemia and anginal pain; 3) angiographically normal left ventricular function; 4) isolated stenosis of the left anterior descending coronary artery (LAD). Patients underwent continuous 12-lead ECG and echocardiographic monitoring during dipyridamole infusion. RESULTS: During dipyridamole infusion 3 patients (13%) did not develop echocardiographic changes, ECG changes or angina, 14 (58%) exhibited ECG changes, 18 (75%) lamented angina and 16 (67%) developed echocardiographic changes. In 5 of these 16 patients (31.5%) echocardiographic changes occurred in LAD-dependent territories only, in 5 they occurred in non-LAD-dependent territories only (31.5%) and in 6 (37%) they occurred in both LAD- and non-LAD-dependent territories. A total of 14 patients exhibited both echocardiographic and ECG changes and/or angina. In 6 of these 14 patients (43%) echocardiographic changes were the first ischemic events; in the remaining 8 patients (57%) ECG changes and/or angina were the first ischemic events. CONCLUSION: In the majority of patients during dipyridamole infusion regional wall motion changes occur in territories supplied by non-stenotic coronary artery branches; they are probably caused, therefore, by distal vessel dysfunction. Furthermore, the sequence of ischemic events is different in individual patients. These findings indicate that in stable angina the mechanisms of ischemia are multiple and that the link between coronary stenoses and myocardial ischemia is very elusive. PMID- 10868921 TI - Heparin-induced thrombocytopenia in patients treated with unfractionated heparin: prevalence of thrombosis in a 1 year follow-up. AB - BACKGROUND: Patients with unstable angina are usually treated with unfractionated heparin and aspirin, but very little is known about the prevalence of heparin induced antibodies and their relation to thrombotic complications some time after the acute phase of unstable angina. The aim of the present study was to establish the prevalence of heparin-induced thrombocytopenia and the prevalence of heparin dependent platelet-reactive antibodies in patients treated with unfractionated heparin and the occurrence of thrombosis in a 1 year follow-up. METHODS: Patient population included 124 consecutive patients with unstable angina treated with unfractionated heparin for almost 5 days. The prevalence of heparin-dependent platelet-reactive antibodies using an ELISA assay was measured before the beginning of heparin therapy and after 7 and 40 days. The platelet count was measured at the same time and the presence of thrombotic occurrences was checked. Clinical follow-up lasted 1 year. RESULTS: At baseline no one patient was positive for heparin-induced antibodies. On day 6, 38 patients (30%) produced a positive heparin-induced antibody result and 30 patients (24%) had an intermediate result. The majority of patients (74%) who developed antibodies became positive after 6 days of heparin therapy. The combined incidence of death, myocardial infarction, recurrent angina, urgent revascularization and stroke was 66% in patients with antibodies and 44% in patients without antibodies during a 1 year follow-up. The incidence of combined primary end points was statistically higher in patients positive for antibodies. The log-rank test was statistically significant (chi2 = 4.39, p < 0.01). CONCLUSIONS: No one patient developed a clinical evidence of thrombocytopenia. Nevertheless thrombotic events during follow-up were more common in patients who developed heparin-induced antibodies. These patients need a more accurate evaluation and surveillance after hospital discharge. PMID- 10868922 TI - Anti-heparin antibodies: are they innocent in the long run? PMID- 10868923 TI - The spectrum of pericardial effusion in acute myocardial infarction: an echocardiographic study. AB - BACKGROUND: The aim of this study was to assess the prevalence of pericardial effusion in acute myocardial infarction and the different prognosis associated with distinct patterns of pericardial effusion (anechoic/hypoechoic vs hyperechoic effusion). METHODS: Five hundred eighty-five consecutive patients admitted to the Coronary Care Unit for acute myocardial infarction were initially considered. Forty of them were excluded due to a technically poor acoustic window. The remaining 545 patients were studied by two-dimensional echocardiography at admission, before discharge (after an average of 9 days in the Coronary Care Unit) and whenever there was an important change in the clinical status (chest pain, lipothymia or syncope, hemodynamic deterioration with systolic blood pressure < 90 mmHg, cardiac arrest). RESULTS: Pericardial effusion was found in 51 patients (9%). Three distinct textural patterns of pericardial effusion were noted on the basis of the echogenic properties: 1) anechoic or hypoechoic pericardial effusion was frequent (30 patients), mild or moderate and generally benign; 2) hyperechoic type "A" effusion pattern was rare (2 patients) and associated with fever, leukocytosis and pericardial rubs; 3) hyperechoic type "B" was frequent (19 patients), large and always associated with major complications (all cases cardiac tamponade and/or death). CONCLUSIONS: Pericardial effusion is not an uncommon finding in serial echo evaluation of patients with acute myocardial infarction, especially when infarction is anterior, extensive and Q wave. Echocardiographically detected pericardial effusion shows different textural patterns with hypoanechoic effusion more frequent, limited and prognostically benign than hyperechoic effusion larger and often associated with adverse events. PMID- 10868924 TI - Procedural results, in-hospital course and six-month follow-up after rescue compared to primary stenting in acute myocardial infarction. AB - BACKGROUND: Although many previous reports showed a worse outcome after rescue compared to primary coronary angioplasty, a direct comparison of these two strategies in the era of stenting is lacking. METHODS: Fifty patients treated with rescue stenting were retrospectively compared to 61 patients treated with primary stenting during acute myocardial infarction over a 4-year period in our Laboratory. RESULTS: Baseline demographic and angiographic parameters were not significantly different in the two groups. Despite a significantly longer time-to reperfusion in rescue stenting (4.7 +/- 2.7 vs 2.8 +/- 2.1 hours, p < 0.0001), procedural success rate (98 vs 97%), in-hospital mortality (6 vs 11%) and target vessel revascularization at 6 months (8 vs 10%) were similar in rescue compared to primary stenting. CONCLUSIONS: These data suggest that stenting may help improve results of rescue angioplasty, and support the concept that aggressive treatment after failed thrombolysis can be pursued with satisfactory results. PMID- 10868925 TI - Prognostic role of heart rate variability in patients with idiopathic dilated cardiomyopathy. AB - BACKGROUND: The aim of this study was to investigate whether heart rate variability may predict the outcome in patients with idiopathic dilated cardiomyopathy. METHODS: Time-domain and frequency-domain heart rate variability was analyzed on 24-hour Holter recordings of 56 patients with idiopathic dilated cardiomyopathy (70% males, mean age 49 +/- 16 years; left ventricular ejection fraction 28 +/- 6%). RESULTS: There were 8 cardiac deaths (14.3%) and 11 arrhythmic events (19.6%, either sudden death or sustained ventricular tachycardia) at a follow-up of 18.5 months (range 3-50 months). Furthermore, 6 patients were included in the list for cardiac transplantation, leading to a prevalence of total cardiac events of 37.5 % (21 patients). All time-domain and most frequency-domain heart rate variability parameters did not show any significant relationship with the end points. However, a low frequency to high frequency (LF/HF) ratio < 1.2 was associated with cardiac death (relative risk-RR 6.8, p < 0.03), arrhythmic events (RR 11.0, p < 0.004), and total cardiac events (RR 4.8, p < 0.002). On the multivariate Cox analysis, no variable showed an independent association with cardiac death, but an LF/HF ratio < 1.2 was the only variable independently predictive of arrhythmic events (RR 8.2, p < 0.02), and the most powerful predictor of total cardiac events (RR 3.8, p < 0.009). CONCLUSIONS: Our data show that, in patients with idiopathic dilated cardiomyopathy, a low LF/HF ratio, as assessed on 24-hour Holter recordings, is a powerful predictor of cardiac events. PMID- 10868926 TI - Induction of interleukin-1beta and interleukin-6 gene expression in hypoperfused skeletal muscle of patients with peripheral arterial disease. AB - BACKGROUND: A growing amount of data supports the role of inflammation in the pathophysiology of atherosclerotic diseases but the cellular source of cytokines has not been clearly identified. Cytokines could be produced by inflammatory cells, activated endothelial and smooth muscle cells, and by the tissue exposed to recurrent ischemia. Accordingly, we evaluated whether hypoperfusion induces gene expression of interleukin (IL)-1beta and IL-6 in the skeletal muscle of patients with peripheral arterial disease and critical limb ischemia. METHODS: Skeletal muscle biopsies were obtained, during a femoral-distal bypass, from normoperfused (control) and hypoperfused skeletal muscles in 8 patients. Gene expression was assessed by semiquantitative reverse transcriptase-polymerase chain reaction, using glyceraldehyde-phosphate-deydrogenase mRNA levels as a normalization factor. RESULTS: In the hypoperfused biopsies, the level of IL 1beta gene expression was significantly higher in all but 2 patients (mean upregulation > 8.8 fold, p = 0.043), and the level of IL-6 gene expression was significantly higher in all but 1 patient (mean upregulation > 23.7 fold, p = 0.031). CONCLUSIONS: We report that IL-1beta and IL-6 gene expression is markedly upregulated in hypoperfused skeletal muscle of patients with critical lower limb ischemia. To our knowledge this is the first report of a local activation of the inflammatory cascade at the level of hypoperfused skeletal muscle. This activation, which could worsen symptoms and tissue viability and be involved in the pathophysiology of reperfusion injury, might be considered as a therapeutic target. It remains to be investigated whether our results may also apply to coronary artery disease. PMID- 10868927 TI - Oxidative stress in ischemia-reperfusion injury: assessment by three independent biochemical markers. AB - BACKGROUND: Oxidative stress plays a key role in ischemia-reperfusion injury, causing peroxidation of tissue lipids and proteins. However, it is debated whether brief ischemic episodes are sufficient to cause detectable oxidative stress in humans, since biochemical markers used so far in the setting of percutaneous transluminal coronary angioplasty (PTCA) gave conflicting results. METHODS: We determined lipid hydroperoxides (ROOHs), conjugated dienes (CD) and total radical-trapping antioxidant capacity (TRAP), three different independent markers of oxidative stress, in aortic and great cardiac vein blood of 5 patients undergoing PTCA before a single balloon inflation lasting 115 +/- 38 s (t0), and 1 min (t1), 5 min (t5), 15 min (t15) after balloon deflation (Group 1). ROOHs and CD were also determined in aortic and great cardiac vein blood of 5 patients with mitral valve disease (Group 2). RESULTS: In Group 1, great cardiac vein levels of ROOHs and CD at t1 increased by 219% and 79%, respectively, compared to t0 (p < 0.01); this sharp and consistent increase persisted up to t15 (+189% and +63%, respectively, compared to t0; p < 0.01). Great cardiac vein levels of TRAP were significantly lower than aortic levels at t0, and exhibited a further decrease at tl. No significant differences in aortic and great cardiac vein levels of ROOHs and CD at t0 were observed between Group 1 and Group 2. CONCLUSIONS: The three methods we used showed a remarkable sensitivity for the detection of post ischemic reperfusion injury in cardiac venous blood and may be useful for detecting small foci of ischemia-reperfusion injury in microvascular angina. PMID- 10868928 TI - Evidence of walk-through phenomenon during echocardiographic dipyridamole stress test. AB - This case report deals with induced regional wall motion abnormalities that spontaneously disappeared during an echocardiographic stress test with dipyridamole. A patient underwent this test because of atypical chest discomfort and a positive result of exercise stress test. Transient septal, apical and anterior akinesia were observed after the first dose of dipyridamole, but they were short-lasting and did not return during the continuation of the test. Coronary angiography showed a critical stenosis of the left coronary artery. A mechanism similar to that responsible for the walk-through phenomenon might explain the observed findings. Thus stress echo with dipyridamole needs careful continuous monitoring, because transient wall motion abnormalities can otherwise be missed resulting in a false negative test. PMID- 10868929 TI - Uncoupling proteins 2 and 3: potential regulators of mitochondrial energy metabolism. AB - Mitochondria use energy derived from fuel combustion to create a proton electrochemical gradient across the mitochondrial inner membrane. This intermediate form of energy is then used by ATP synthase to synthesize ATP. Uncoupling protein-1 (UCP1) is a brown fat-specific mitochondrial inner membrane protein with proton transport activity. UCP1 catalyzes a highly regulated proton leak, converting energy stored within the mitochondrial proton electrochemical potential gradient to heat. This uncouples fuel oxidation from conversion of ADP to ATP. In rodents, UCP1 activity and brown fat contribute importantly to whole body energy expenditure. Recently, two additional mitochondrial carriers with high similarity to UCP1 were molecularly cloned. In contrast to UCP1, UCP2 is expressed widely, and UCP3 is expressed preferentially in skeletal muscle. Biochemical studies indicate that UCP2 and UCP3, like UCP1, have uncoupling activity. While UCP1 is known to play an important role in regulating heat production during cold exposure, the biological functions of UCP2 and UCP3 are unknown. Possible functions include 1) control of adaptive thermogenesis in response to cold exposure and diet, 2) control of reactive oxygen species production by mitochondria, 3) regulation of ATP synthesis, and 4) regulation of fatty acid oxidation. This article will survey present knowledge regarding UCP1, UCP2, and UCP3, and review proposed functions for the two new uncoupling proteins. PMID- 10868930 TI - Insulin-secreting cells derived from embryonic stem cells normalize glycemia in streptozotocin-induced diabetic mice. AB - Embryonic stem (ES) cells display the ability to differentiate in vitro into a variety of cell lineages. Using a cell-trapping system, we have obtained an insulin-secreting cell clone from undifferentiated ES cells. The construction used allows the expression of a neomycin selection system under the control of the regulatory regions of the human insulin gene. The chimeric gene also contained a hygromycin resistance gene (pGK-hygro) to select transfected cells. A resulting clone (IB/3x-99) containing 16.5 ng/microg protein of total insulin displays regulated hormone secretion in vitro in the presence of various secretagogues. Clusters obtained from this clone were implanted (1 x 10(6) cells) in the spleen of streptozotocin-induced diabetic animals. Transplanted animals correct hyperglycemia within 1 week and restore body weight in 4 weeks. Whereas an intraperitoneal glucose tolerance test showed a slower recovery in transplanted versus control mice, blood glucose normalization after a challenge meal was similar. This approach opens new possibilities for tissue transplantation in the treatment of type 1 and type 2 diabetes and offers an alternative to gene therapy. PMID- 10868932 TI - Hypothalamic localization of the feeding effect of agouti-related peptide and alpha-melanocyte-stimulating hormone. AB - The melanocortin-4 receptor (MC4R) in the hypothalamus is thought to be important in physiological regulation of food intake. We investigated which hypothalamic areas known to express MC4R are involved in the regulation of feeding by using alpha-melanocyte-stimulating hormone (alpha-MSH), an endogenous MC4R agonist, and agouti-related peptide (Agrp), an endogenous MC4R antagonist. Cannulae were inserted into the rat hypothalamic paraventricular (PVN), arcuate (Arc), dorsomedial (DMN), and ventromedial (VMN) nuclei; the medial preoptic (MPO), anterior hypothalamic (AHA), and lateral hypothalamic (LHA) areas; and the extrahypothalamic central nucleus of the amygdala (CeA). Agrp (83-132) (0.1 nmol) and [Nle4, D-Phe7]alpha(-MSH (NDP-MSH) (0.1 nmol), a stable alpha-MSH analog, were administered to fed and fasted rats, respectively. The PVN, DMN, and MPO were the areas with the greatest response to Agrp and NDP-MSH. At 8 h postinjection, Agrp increased feeding in the PVN by 218 +/- 23% (P < 0.005), in the DMN by 268 +/- 42% (P < 0.005), and in the MPO by 236 +/- 31% (P < 0.01) compared with a saline control group for each nucleus. NDP-MSH decreased food intake in the PVN by 52 +/- 6% (P < 0.005), in the DMN by 44 +/- 6% (P < 0.0001), and in the MPO by 55 +/- 6% (P < 0.0001) at 1 h postinjection. Injection into the AHA and CeA resulted in smaller alterations in food intake. No changes in feeding were seen after the administration of Agrp into the Arc, LHA, or VMN, but NDP-MSH suppressed food intake in the Arc and LHA. This study indicates that the hypothalamic nuclei expressing MC4R vary in their sensitivity to Agrp and alpha MSH with regard to their effect on feeding. PMID- 10868931 TI - Independent development of pancreatic alpha- and beta-cells from neurogenin3 expressing precursors: a role for the notch pathway in repression of premature differentiation. AB - The nature and identity of the pancreatic beta-cell precursor has remained elusive for many years. One model envisions an early multihormonal precursor that gives rise to both alpha- and beta-cells and the other endocrine cell types. Alternatively, beta-cells have been suggested to arise late, directly from the GLUT2- and pancreatic duodenal homeobox factor-1 (PDX1)-expressing epithelium, which gives rise also to the acinar cells during this stage. In this study, we have identified a subset of the PDX1+ epithelial cells that are marked by expression of Neurogenin3 (Ngn3). Ngn3, a member of the basic helix-loop-helix (bHLH) family of transcription factors, is suggested to act upstream of NeuroD in a bHLH cascade. Detailed analysis of Ngn3/paired box factor 6 (PAX6) and NeuroD/PAX6 co-expression shows that the two bHLH factors are expressed in a largely nonoverlapping set of cells, but such analysis also suggests that the NeuroD+ cells arise from cells expressing Ngn3 transiently. NeuroD+ cells do not express Ki-67, a marker of proliferating cells, which shows that these cells are postmitotic. In contrast, Ki-67 is readily detected in Ngn3+ cells. Thus, Ngn3+ cells fulfill the criteria for an endocrine precursor cell. These expression patterns support the notion that both alpha- and beta-cells develop independently from PDX1+/Ngn3+ epithelial cells, rather than from GLU+/INS+ intermediate stages. The earliest sign of alpha-cell development appears to be Brain4 expression, which apparently precedes Islet-1 (ISL1) expression. Based on our expression analysis, we propose a temporal sequence of gene activation and inactivation for developing alpha- and beta-cells beginning with activation of NeuroD expression. Endocrine cells leave the cell cycle before NeuroD activation, but re-enter the cell cycle at perinatal stages. Dynamic expression of Notch1 in PDX+ epithelial cells suggests that Notch signaling could inhibit a Ngn-NeuroD cascade as seen in the nervous system and thus prevent premature differentiation of endocrine cells. PMID- 10868933 TI - The transferrin receptor defines two distinct contraction-responsive GLUT4 vesicle populations in skeletal muscle. AB - Insulin and contraction increase glucose transport in an additive fashion in skeletal muscle. However, it is still unclear whether they do so by inducing the recruitment of GLUT4 transporters from the same or distinct intracellular compartments to the plasma membrane and the T-tubules. Using the transferrin receptor as a recognized marker of recycling endosomes, we have examined whether insulin and/or contraction recruit GLUT4 from this pool to either the plasma membranes or T-tubules, isolated by subcellular fractionation of perfused hindlimb muscles. Either stimulus independently increased GLUT4 translocation from an intracellular fraction to both the plasma membrane and T-tubules. The combination of insulin and contraction induced a marked (approximately threefold) and almost fully additive increase in GLUT4 content, but only in the plasma membrane. Insulin did not stimulate transferrin receptor recruitment from the GLUT4-containing intracellular fraction to either the plasma membrane or the T tubules. In contrast, contraction stimulated the recruitment of the transferrin receptor from the same GLUT4-containing intracellular fraction to the plasma membrane but not to the T-tubules. Contraction-induced recruitment of the transferrin receptor was also observed from immunopurified GLUT4 vesicles. It is concluded that muscle contraction stimulates translocation of GLUT4 from two distinct intracellular compartments: 1) a population of recycling endosomes that is selectively recruited to the plasma membrane and 2) from GLUT4 storage vesicles that are also insulin-responsive and recruited to both the plasma membrane and the T-tubules. The lack of additive translocation of GLUT4 to the T tubules may be linked to the failure of GLUT4-containing recycling endosomes to be recruited to these structures. PMID- 10868934 TI - Insulin enhances the bradykinin response in L8 rat skeletal myoblasts. AB - Inhibitors of ACE/kininase II enhance insulin sensitivity, an action that is mediated in part by bradykinin (BK). We investigated whether insulin interacts with the BK receptor signaling to modulate the inositol 1,4,5-trisphosphate (IP3) response to BK in L8 rat skeletal myoblasts. Stimulation of the cultures with BK (10 nmol/l) for 15 s increased IP3 from a basal level of 75.2 +/- 7.6 to 200.2 +/ 15.7 pmol/mg protein. Treatment of the cultures with 1, 2, and 20 nmol/l of insulin for 90 min before adding BK increased IP3 formation by the same BK dose to 328.2 +/- 19, 434.5 +/- 18, and 460.8 +/-21.3 pmol/mg protein, respectively. When wortmannin was administered to inhibit phosphatidylinositol (PI) 3-kinases at lower concentration (1 nmol/l), it increased IP3 formation stimulated by BK only when insulin was present. At a higher concentration (100 nmol/l), wortmannin significantly enhanced BK-induced IP3 formation in the absence of insulin. Genistein and tyrphostin A-23, tyrosine kinase inhibitors, completely reversed the elevated IP3 formation by BK and insulin. The IP3 response to 10 nmol/l BK was 223.3 +/- 11.8 pmol/mg protein in the absence of insulin and 402.2 +/- 12.0 pmol/mg protein in the presence of 2 nmol/l insulin. However, when exposing the cultures to 1 nmol/l genistein or tyrphostin A-23, the IP3 response to BK in the presence of insulin decreased to 211.8 +/- 46.7 and 187.7 +/- 19.9 pmol/mg protein. Tyrphostin A-1, the inactive analog, was ineffective. Exposing the cells to 1 micromol/ 3,4,5-trimethoxybenzoic acid 8-[diethylamino]octyl ester, an intracellular Ca2+ antagonist, did not change the potentiation by insulin. But, exposing them to 0.1 micromol/l n-[6-aminohexyl]-5-chloro-1-naphthalene sulfonamide, a calmodulin antagonist, resulted in enhanced IP3 response to BK alone to 292.2 +/- 18.5 pmol/mg protein and to BK in the presence of 1, 2, and 20 nmol/l insulin to 488 +/- 22.2, 625.5 +/- 11.6, and 665.2 +/- 15.9 pmol/mg protein, respectively. In conclusion, insulin potentiates BK-induced IP3 production in L8 rat skeletal myoblasts, and this action of insulin involves a tyrosine kinase. Inhibition of PI 3-kinases potentiated BK-induced IP3 formation in the presence of insulin. Calmodulin blocked the action of insulin. These results support a modulatory effect of insulin on the BK signaling system via a tyrosine kinase in L8 rat skeletal myoblasts that results in increased IP3 formation. Because BK release from skeletal muscle increases during contractions, this action of insulin is likely to play a role in the modulation of the excitation-contraction coupling process of the skeletal muscle. PMID- 10868935 TI - Study of the regulatory properties of glucokinase by site-directed mutagenesis: conversion of glucokinase to an enzyme with high affinity for glucose. AB - To identify the amino acids involved in the specific regulatory properties of glucokinase, and particularly its low affinity for glucose, mutants of the human islet enzyme have been prepared, in which glucokinase-specific residues have been replaced. Two mutations increased the affinity for glucose by twofold (K296M) and sixfold (Y214A), the latter also decreasing the Hill coefficient from 1.75 to 1.2 with minimal change in the affinity for ATP. Combining these two mutations with N166R resulted in a 50-fold decrease in the half-saturating substrate concentration (S0.5) value, which became then comparable to the Km of hexokinase II. The location of N166, Y214, and K296 in the three-dimensional structure of glucokinase suggests that these mutations act by favoring closure of the catalytic cleft. As a rule, mutations changed the affinity for glucose and for the competitive inhibitor mannoheptulose (MH) in parallel, whereas they barely affected the affinity for N-acetylglucosamine (NAG). These and other results suggest that NAG and MH bind to the same site but to different conformations of glucokinase. A small reduction in the affinity for the regulatory protein was observed with mutations of residues on the smaller domain and in the hinge region, confirming the bipartite nature of the binding site for the regulatory protein. The K296M mutant was found to have a threefold decreased affinity for palmitoyl CoA; this effect was additive to that previously observed for the E279Q mutant, indicating that the binding site for long-chain acyl CoAs is located on the upper face of the larger domain. PMID- 10868936 TI - Maturation of the humoral autoimmune response to epitopes of GAD in preclinical childhood type 1 diabetes. AB - GAD is a major target of autoimmunity in preclinical type 1 diabetes. Here we examine the maturation of the humoral response to GAD epitopes sequentially from birth to diabetes onset or current follow-up in 29 GAD antibody (GADA)+ offspring of parents with diabetes from the BABYDIAB Study. Antibodies were measured against GAD65, GAD67, and GAD65/67 chimeras by radiobinding assay. In 28 of 29 offspring, the first GADAs contained reactivity against epitopes within GAD65 residues 96-444, suggesting that the middle GAD65 region is a primary target of GAD humoral autoimmunity. In 7 of these 28 offspring, initial antibody reactivity was against all epitope regions tested (middle GAD65, COOH-terminal GAD65 residues 445-585, NH2-terminal GAD65 residues 1-95, and GAD67); in 16 offspring, reactivity was to middle and COOH-terminal GAD65 epitopes, and in 5 offspring, reactivity was only to the middle GAD65 epitopes. The single offspring without middle GAD65 reactivity had antibodies to the NH2-terminal epitopes in the absence of all other islet autoimmunity. Subsequent GADA epitope spreading was frequent and seen in 10 of 15 offspring with informative follow-up samples. Spreading was mostly (eight cases) to NH2-terminal GAD65 epitopes. In two offspring, spreading to new epitopes was found when antibody titers to GAD65 and early epitopes were declining, suggesting determinant-specific regulation of the humoral response. None of the GADA reactivities nor any changes in reactivity over time were specifically associated with diabetes onset. The findings suggest that the humoral autoimmune response to GAD found in childhood is dynamic, is initially against epitopes within the middle portion of GAD65, and spreads to epitopes in other regions of GAD65 and GAD67. PMID- 10868937 TI - Inhibitory effect of IGF-I on type 2 nitric oxide synthase expression in Ins-1 cells and protection against activation-dependent apoptosis: involvement of phosphatidylinositol 3-kinase. AB - Challenge of Ins-1 cells, a rat beta-pancreatic cell line, with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma) promoted the expression of type 2 nitric oxide synthase (NOS-2) in a cooperative way. Treatment of Ins-1 cells with IGF-I significantly inhibited the expression of NOS-2, especially at subsaturating concentrations of LPS and IFN-gamma. The inhibitory effect of IGF-I on NOS-2 expression was abrogated when cells were incubated with wortmannin or LY294002, two inhibitors of phosphatidylinositol 3-kinase. Transient expression of the p110 subunit of phosphatidylinositol 3-kinase impaired the LPS and IFN gamma-dependent NOS-2 promoter activity in cells transfected with a 1-kb fragment corresponding to the 5'-flanking region of the NOS-2 gene. However, expression of a dominant negative form of p85 abolished the inhibitory action of IGF-I on the NOS-2 promoter activity. Analysis of the decreased NOS-2 promoter activity in cells incubated with IGF-I showed a lower nuclear factor KB binding as determined by electrophoretic mobility shift assays. The synthesis of NO, produced after LPS and IFN-gamma challenge, triggered an apoptotic response in these cells. IGF-I reduced apoptosis mainly through the decreased synthesis of NO. However, in activated cells treated with N-[3-(aminomethyl)benzyl]acetamidine, a specific NOS 2 inhibitor, IGF-I completely abolished the NO-independent apoptosis. This protection from apoptosis was dependent on phosphatidylinositol 3-kinase activity. These results suggest an important anti-inflammatory and anti-apoptotic role for IGF-I in beta-pancreatic cells, with both actions depending on the activation of phosphatidylinositol 3-kinase. PMID- 10868938 TI - Modulation of humoral islet autoimmunity by pancreas allotransplantation influences allograft outcome in patients with type 1 diabetes. AB - Pancreas transplantation in patients with type 1 diabetes presents allogeneic beta-cell autoantigens to the immune system long after the initial beta-cell destruction that leads to diabetes has occurred. The aims of this study were to determine whether re-exposure to beta-cell autoantigen through transplantation affect the humoral autoimmune response and whether its modulation correlates with graft outcome. Antibodies to the major autoantigens GAD (GADA) and protein tyrosine phosphatase IA-2 (IA-2A) were measured before and after transplantation in patients with type 1 diabetes who received pancreas and kidney allografts. In the 110 cases studied, pancreas graft survival was not significantly associated with the presence of GADA or IA-2A before transplantation. In the 75 patients with sequential follow-up samples up to 11.2 years after transplantation, autoantibodies were persistently undetectable in 44 cases (59%) and remained at stable detectable levels in 13 cases (17%). Substantial changes in antibody levels were found in 18 cases (24%), of which 13 cases (17%) had declining levels and 5 cases (7%) had marked increments after transplantation. Rising GADA and IA 2A levels in these five patients were predominantly of the IgG1 subclass, with progressive spreading of epitope reactivity. Pancreas graft function was lost 0.7 2.3 years after rising autoantibody levels in four of these five patients, and a significantly lower pancreas graft survival was found in patients with major rises in either GADA or IA-2A levels (P < 0.0001 vs. the remainder) and in patients having persistently high levels of IA-2A (P = 0.002 vs. stable antibody negative patients). Kidney graft survival was not associated with islet autoantibody status. In conclusion, a minority of patients receiving pancreas allografts under generalized immunosuppression show a stimulation of islet autoantibody reactivity characteristic of that found in preclinical type 1 diabetes, which is almost invariably followed by graft function failure and resumption of insulin therapy. PMID- 10868939 TI - Early pattern of differentiation in the human pancreas. AB - In the early human embryonic/fetal pancreas, we studied 1) the ontogenetic pattern of the endocrine cells and the evolution of the endocrine mass, and 2) the morphogenetic pattern of development and, more precisely, the complex relationship of the epithelial mass with the surrounding mesenchyme. We studied 15 pancreases between 7 and 11 weeks of development (WD) by double immunohistochemistry. Epithelial cells in these pancreatic anlage were detected by cytokeratin staining, and differentiated endocrine cells were detected by insulin, glucagon, somatostatin, and pancreatic polypeptide staining. Proliferation was quantified using a nuclear marker, the Ki-67 antibody. At this early stage, the pancreas is made up of an epithelial mass composed of central ducts intermingled with a loose mesenchyme and peripheral ducts surrounded by a dense peripancreatic mesenchyme. Hormone-containing cells appear in the epithelium at 8 WD. Newly differentiated endocrine cells coexpress insulin, glucagon, and somatostatin; endocrine differentiation starts within the central ducts of the epithelial mass, at a distance from the dense peripancreatic surrounding mesenchyme. The fraction of the primitive endocrine cells undergoing proliferation is low (5% of the insulin cells at 8 WD, 3% at 11 WD), which is in favor of massive differentiation as the major mechanism for increasing endocrine mass. By contrast, the nonendocrine epithelial cells have a higher rate of proliferation; the epithelial cells in contact with the dense peripancreatic surrounding mesenchyme show more proliferation activity than those within the central part of the epithelial mass (at 11 WD, labeling index: periphery 65% vs. center 15%, P < 0.001). In conclusion, the patterns of endocrine differentiation and epithelial proliferation observed within the human pancreas early in development suggest that the mesenchyme plays a role in these phenomena. PMID- 10868940 TI - Importance of cell-matrix interactions in rat islet beta-cell secretion in vitro: role of alpha6beta1 integrin. AB - It has long been recognized that islet cell function is rapidly altered in vitro, but can be maintained, at least in part, when cells are layered on defined extracellular matrices. The present work addresses the influence of short-term cell-matrix interactions on islet beta-cell function and provides first insight into the molecular basis of these interactions. When primary rat beta-cells were allowed to attach to a matrix produced by a rat carcinoma cell line (804G), there was an increased insulin secretory response to secretagogues. This change was the result of an increase in the proportion of actively secreting beta-cells and in the amount of insulin secreted per active cell, as shown using the reverse hemolytic plaque assay. In turn, the spreading or flattening of beta-cells on this matrix was enhanced by secretagogues, and flattened cells secreted more insulin than rounded cells. Using indirect immunofluorescence, it was found that 1)alpha6beta1 integrins are present at the surface of islet cells in situ, 2) alpha6beta1 expression is heterogeneous among purified beta-cells and is upregulated by insulin secretagogues, 3) alpha6beta1 expression is higher in spreading cells, and 4) anti-alpha6beta1-specific antibodies decrease spreading. These observations demonstrate that islet cell-matrix interactions can improve the sensitivity of insulin cells to glucose and are mediated, at least in part, by alpha6beta1 integrins, suggesting that outside-in signaling through alpha6beta1 integrin plays a major role in the regulation of beta-cell function. PMID- 10868941 TI - Effects of streptozotocin-induced diabetes and insulin treatment on the hypothalamic melanocortin system and muscle uncoupling protein 3 expression in rats. AB - Hypothalamic melanocortins are among several neuropeptides strongly implicated in the control of food intake. Agonists for melanocortin 4 (MC-4) receptors such as alpha-melanocyte-stimulating hormone (alpha-MSH), a product of proopiomelanocortin (POMC), reduce food intake, whereas hypothalamic agouti related protein (AgRP) is a MC-4 receptor antagonist that increases food intake. To investigate whether reduced melanocortin signaling contributes to hyperphagia induced by uncontrolled diabetes, male Sprague-Dawley rats were studied 7 days after administration of streptozotocin (STZ) or vehicle. In addition, we wished to determine the effect of diabetes on muscle uncoupling protein 3 (UCP-3), a potential regulator of muscle energy metabolism. STZ diabetic rats were markedly hyperglycemic (31.3 +/- 1.0 mmol/l; P < 0.005) compared with nondiabetic controls (9.3 +/- 0.2 mmol/l). Insulin treatment partially corrected the hyperglycemia (18.8 +/- 2.5 mol/l; P < 0.005). Plasma leptin was markedly reduced in STZ diabetic rats (0.4 +/- 0.1 ng/ml; P < 0.005) compared with controls (3.0 +/- 0.4 ng/ml), an effect that was also partially reversed by insulin treatment (1.8 +/- 0.3 ng/ml). Untreated diabetic rats were hyperphagic, consuming 40% more food (48 +/- 1 g/day; P < 0.005) than controls (34 +/- 1 g/day). Hyperphagia was prevented by insulin treatment (32 +/- 2 g/day). In untreated diabetic rats, hypothalamic POMC mRNA expression (measured by in situ hybridization) was reduced by 80% (P < 0.005), whereas AgRP mRNA levels were increased by 60% (P < 0.01), suggesting a marked decrease of hypothalamic melanocortin signaling. The change in POMC, but not in AgRP, mRNA levels was partially reversed by insulin treatment. By comparison, the effects of diabetes to increase hypothalamic neuropeptide Y (NPY) expression and to decrease corticotropin-releasing hormone (CRH) expression were normalized by insulin treatment, whereas the expression of mRNA encoding the long form of the leptin receptor in the arcuate nucleus was unaltered by diabetes or insulin treatment. UCP-3 mRNA expression in gastrocnemius muscle from diabetic rats was increased fourfold (P < 0.005), and the increase was prevented by insulin treatment. The effect of uncontrolled diabetes to decrease POMC, while increasing AgRP gene expression, suggests that reduced hypothalamic melanocortin signaling, along with increased NPY and decreased CRH signaling, could contribute to diabetic hyperphagia. These responses, in concert with increased muscle UCP-3 expression, may also contribute to the catabolic effects of uncontrolled diabetes on fuel metabolism in peripheral tissues. PMID- 10868942 TI - Beta-blockade, but not normoglycemia or hyperinsulinemia, markedly diminishes stress-induced hyperglycemia in diabetic dogs. AB - Stress-induced hyperglycemia can lead to significant deterioration in glycemic control in individuals with diabetes. Previously, we have shown in normal dogs that, after intracerebroventricular (ICV) administration of carbachol (a model of moderate stress), increases in both the metabolic clearance rate (MCR) of glucose and endogenous glucose production (GP) occur. However, in hyperglycemic diabetic dogs subjected to the same stress, the MCR of glucose does not increase and glycemia therefore markedly deteriorates because of stimulation of GP. Our aims were to determine the following: 1) whether insulin-induced acute normalization of glycemia, with or without beta-blockade, would correct glucose clearance and prevent the hyperglycemic effect of stress, and 2) whether hyperinsulinemia per se could correct these abnormalities. Stress was induced by ICV carbachol in 27 experiments in five alloxan-administered diabetic dogs subjected to the following protocols in random order: 1) basal insulin infusion (BI) to restore normoglycemia; 2) basal insulin infusion with beta-blockade (BI+block); 3) normoglycemic-hyperinsulinemic clamp with threefold elevation of insulin above basal (3x BI); and 4) normoglycemic-hyperinsulinemic clamp with fivefold elevation of insulin above basal (5 x BI). The BI+block protocol fully prevented stress-induced hyperglycemia, both by increasing MCR (deltaMCR at peak: 0.72 +/- 0.25 ml x kg(-1) x min(-1) vs. no change in BI, P < 0.05) and by diminishing the stress-induced increment in GP observed in BI (deltaGP at peak: 3.72 +/- 0.09 micromol x kg(-1) x min(-1) for BI+block vs. 14.10 +/- 0.31 micromol x kg(-1) x min(-1) for BI, P < 0.0001). In contrast, 3x BI and 5x BI treatments with normoglycemic-hyperinsulinemic clamps proportionately increased basal MCR at baseline, but paradoxically were not associated with an increase in MCR in response to stress, which induced a twofold increase in GP. Thus, in alloxan administered diabetic dogs, stress increased GP but not MCR, despite normalization of glycemia with basal or high insulin. In contrast, beta adrenergic blockade almost completely restored the metabolic response to stress to normal and prevented marked hyperglycemia, both by limiting the rise in GP and by increasing glucose MCR. We conclude that acute normalization of glycemia with basal insulin or hyperinsulinemia does not prevent hyperglycemic effects of stress unless accompanied by beta-blockade, and we speculate that short-term beta blockade may be a useful treatment modality under some stress conditions in patients with diabetes. PMID- 10868943 TI - Potential role of glycogen synthase kinase-3 in skeletal muscle insulin resistance of type 2 diabetes. AB - Glycogen synthase (GS) activity is reduced in skeletal muscle of type 2 diabetes, despite normal protein expression, consistent with altered GS regulation. Glycogen synthase kinase-3 (GSK-3) is involved in regulation (phosphorylation and deactivation) of GS. To access the potential role of GSK-3 in insulin resistance and reduced GS activity in type 2 diabetes, the expression and activity of GSK-3 were studied in biopsies of vastus lateralis from type 2 and nondiabetic subjects before and after 3-h hyperinsulinemic (300 mU x m(-2) x min(-1))-euglycemic clamps. The specific activity of GSK-3alpha did not differ between nondiabetic and diabetic muscle and was decreased similarly after 3-h insulin infusion. However, protein levels of both alpha and beta isoforms of GSK-3 were elevated (approximately 30%) in diabetic muscle compared with lean (P < 0.01) and weight matched obese nondiabetic subjects (P < 0.05) and were unchanged by insulin infusion. Thus, both basal and insulin-stimulated total GSK-3 activities were elevated by approximately twofold in diabetic muscle. GSK-3 expression was related to in vivo insulin action, as GSK-3 protein was negatively correlated with maximal insulin-stimulated glucose disposal rates. In summary, GSK-3 protein levels and total activities are 1) elevated in type 2 diabetic muscle independent of obesity and 2) inversely correlated with both GS activity and maximally insulin-stimulated glucose disposal. We conclude that increased GSK-3 expression in diabetic muscle may contribute to the impaired GS activity and skeletal muscle insulin resistance present in type 2 diabetes. PMID- 10868944 TI - Effects of type 2 diabetes on the ability of insulin and glucose to regulate splanchnic and muscle glucose metabolism: evidence for a defect in hepatic glucokinase activity. AB - Insulin-induced stimulation of muscle glucose uptake (MGU) is impaired in people with type 2 diabetes. To determine whether insulin-induced stimulation of splanchnic glucose uptake (SGU) is also impaired, we simultaneously measured leg glucose uptake (LGU) and SGU in 14 nondiabetic subjects and 16 subjects with type 2 diabetes using a combined organ catheterization-tracer infusion technique. Glucose was clamped at approximately 9.3 mmol/l, while insulin concentrations were maintained at approximately 72 pmol/l (low) and approximately 150 pmol/l (high) for 3 h each. Endogenous hormone secretion was inhibited with somatostatin. Total body glucose disappearance was lower (P < 0.01) and glucose production higher (P < 0.01) during both insulin infusions in the diabetic compared with the nondiabetic subjects, indicating insulin resistance. Splanchnic glucose production was higher (P < 0.05) in the diabetic subjects during the low but not the high insulin infusion. SGU was slightly lower in the diabetic than in the nondiabetic subjects during the low insulin infusion and 50-60% lower (P < 0.05) during the high insulin infusion. LGU (P < 0.001), but not SGU, was inversely correlated with the degree of visceral adiposity. The contribution of the indirect pathway to hepatic glycogen synthesis did not differ in the diabetic and nondiabetic subjects. In contrast, both flux through the UDP-glucose pool (P < 0.05) and the contribution of the direct pathway to glycogen synthesis (P < 0.01) were lower in the diabetic than in the nondiabetic subjects, indicating decreased uptake and/or phosphorylation of extracellular glucose. On the other hand, glycogenolysis was equally suppressed in both groups. In summary, type 2 diabetes impairs the ability of insulin to stimulate both MGU and SGU. The defect appears to reside at a proximal (e.g., glucokinase) metabolic step and is not related to the degree of visceral adiposity. These data suggest that impaired hepatic glucose uptake as well as MGU contribute to hyperglycemia in people with type 2 diabetes. PMID- 10868945 TI - Characterization of signal transduction and glucose transport in skeletal muscle from type 2 diabetic patients. AB - We characterized metabolic and mitogenic signaling pathways in isolated skeletal muscle from well-matched type 2 diabetic and control subjects. Time course studies of the insulin receptor, insulin receptor substrate (IRS)-1/2, and phosphatidylinositol (PI) 3-kinase revealed that signal transduction through this pathway was engaged between 4 and 40 min. Insulin-stimulated (0.6-60 nmol/l) tyrosine phosphorylation of the insulin receptor beta-subunit, mitogen-activated protein (MAP) kinase phosphorylation, and glycogen synthase activity were not altered in type 2 diabetic subjects. In contrast, insulin-stimulated tyrosine phosphorylation of IRS-1 and anti-phosphotyrosine-associated PI 3-kinase activity were reduced 40-55% in type 2 diabetic subjects at high insulin concentrations (2.4 and 60 nmol/l, respectively). Impaired glucose transport activity was noted at all insulin concentrations (0.6-60 nmol/l). Aberrant protein expression cannot account for these insulin-signaling defects because expression of insulin receptor, IRS-1, IRS-2, MAP kinase, or glycogen synthase was similar between type 2 diabetic and control subjects. In skeletal muscle from type 2 diabetic subjects, IRS-1 phosphorylation, PI 3-kinase activity, and glucose transport activity were impaired, whereas insulin receptor tyrosine phosphorylation, MAP kinase phosphorylation, and glycogen synthase activity were normal. Impaired insulin signal transduction in skeletal muscle from type 2 diabetic patients may partly account for reduced insulin-stimulated glucose transport; however, additional defects are likely to play a role. PMID- 10868946 TI - Leptin induces direct vasodilation through distinct endothelial mechanisms. AB - In this study, we reveal that leptin evokes an acute hypotensive effect in 6 hydroxydopamine sympathectomized rats (response to maximal leptin dose, mean blood pressure: from 92 +/- 4 to 78 +/- 2 mmHg, P < 0.01). This hemodynamic effect is related to a direct action of the hormone on vascular tone, since in aortic and mesenteric rings increasing doses of leptin evoke a dose-dependent vasorelaxation (aorta: from 3 +/- 1 to 36 +/- 3, n = 15; mesenteric: from 6 +/- 1 to 30 +/- 5, n = 10), which is impaired by endothelial denudation. In particular, leptin-evoked vasorelaxation is impaired by nitric oxide synthase inhibition in aorta (delta% of maximal response: from 36 +/- 3 to 3 +/- 1, P < 0.01) and by endothelium-derived hyperpolarizing factor (EDHF) inhibition in mesenteric arteries (delta% of maximal response: from 30 +/- 5 to 7 +/- 2, P < 0.01), suggesting that vasorelaxation evoked by leptin is heterogeneous and related to the vascular bed. Finally, the inhibition of nitric oxide synthase by NG-nitro-L arginine-methyl ester does not modify blood pressure response to leptin, suggesting a predominant role of the EDHF mechanism in the hypotensive effect of leptin. PMID- 10868947 TI - Polymorphism in the glycogen-associated regulatory subunit of type 1 protein phosphatase (PPP1R3) gene and insulin sensitivity. AB - A polymorphism (PP1ARE) in the 3'-untranslated region of the gene encoding the glycogen-associated regulatory subunit of type 1 protein phosphatase PPP1R3 is associated with insulin resistance in Pima Indians. The aim of this study was to investigate whether two common variants in the PPP1R3 gene, Asp905Tyr and PP1ARE, are associated with reduced insulin sensitivity or can predict the development of impaired glucose tolerance (IGT) or type 2 diabetes during a 20-year follow-up period in 696 50-year-old Caucasian men. The allelic frequency of Tyr905 was 0.11 (95% CI 0.09-0.13) and of PP1ARE 0.34 (0.31-0.37) and the two polymorphisms were in linkage disequilibrium (chi2 = 46, P < 0.0001, Fisher's exact test). None of the polymorphisms was associated with the development of IGT or type 2 diabetes, but the PP1ARE polymorphism was weakly correlated to whole-body insulin sensitivity (r = -0.08, P = 0.04). In conclusion, we found no evidence in Swedish men that the PP1ARE or the Asp905Tyr variants over a 20-year period predict the development of IGT or type 2 diabetes, but the PP1ARE polymorphism could have a higher penetrance in other populations. PMID- 10868948 TI - Beta-cell transcription factors and diabetes: no evidence for diabetes-associated mutations in the hepatocyte nuclear factor-3beta gene (HNF3B) in Japanese patients with maturity-onset diabetes of the young. AB - Mutations in the transcription factors hepatocyte nuclear factor (HNF)-4alpha and -1alpha, insulin promoter factor-1, and HNF-1beta are the causes of four forms of maturity-onset diabetes of the young (MODY1 and 3-5, respectively). The winged helix transcription factor HNF-3beta has been implicated in the regulation of expression of each of these MODY genes, suggesting that mutations in the HNF 3beta gene (HNF3B) may also cause MODY. We have tested this hypothesis by screening a panel of 57 unrelated Japanese subjects with a clinical diagnosis of MODY for mutations in HNF3B. This analysis revealed four frequent polymorphisms that were not associated with MODY, including one in the promoter region ( 213A/G), two silent mutations in the codons for Ala 97 (291C/T) and Gly 279 (837A/G), and one in the 3'-untranslated region (1424C/T). Two rare substitutions in the 5'-untranslated region, -156C/T and -67A/C, were found in a heterozygous state in two subjects, and two subjects were heterozygous for putative missense mutations, S109N (326G > A) and A328V (983C>T). The two missense mutations were not found in 106 normal chromosomes from nondiabetic subjects. It was not possible to test for co-segregation of these mutations with diabetes and thus, it is unclear whether or not these mutations can cause MODY. The results of our study suggest that mutations in HNF3B are not a common cause of MODY in Japanese subjects. PMID- 10868949 TI - Genetic variation in the hepatocyte nuclear factor-3beta gene (HNF3B) does not contribute to maturity-onset diabetes of the young in French Caucasians. AB - Mutations in genes encoding hepatocyte nuclear factor (HNF) are responsible for three of the five subtypes of maturity-onset diabetes of the young (MODY). This observation and molecular studies indicate that the HNF network is required for normal function of pancreatic beta-cells. This suggests that transcription factors involved in this complex network are candidates for genetic defects in MODY. Because the HNF-3beta gene is implicated in this network, we screened it for mutations in 21 probands of French ancestry with clinical diagnosis of MODY and early-onset type 2 diabetes. All of the five known MODY genes, HNF-4alpha, glucokinase, HNF-1alpha, HNF-1beta, and IPF1, were previously excluded as being the cause of diabetes in these families. By direct sequencing, we identified two transitions, an A-to-G at position -213 and a C-to-T at position -63 in the promoter and exon 1, respectively, of the HNF-3beta gene. A G-to-C transversion at position +32 in the intron 1 and three transitions, C-to-T at position 291, A to-G at position 837, and G-to-A at position 1188 in the exon 3, resulting in noncoding mutations Ala97Ala, Gly279Gly, and Gln396Gln, respectively, were also identified. The allele frequencies were not significantly different between a control group and MODY probands. Familial segregation studies and linkage analysis showed that genetic variation in the HNF-3beta gene is unlikely to be the cause of early-onset type 2 diabetes in these Caucasian families. PMID- 10868950 TI - Diverse roles of K(ATP) channels learned from Kir6.2 genetically engineered mice. AB - The regulation of insulin secretion from pancreatic beta-cells depends critically on the activities of their plasma membrane ion channels. ATP-sensitive K+ channels (K(ATP) channels) are present in many cells and regulate a variety of cellular functions by coupling cell metabolism with membrane potential. The activity of the K(ATP) channels in pancreatic beta-cells is regulated by changes in the ATP and ADP concentrations (ATP/ADP ratio) caused by glucose metabolism. Thus, the K(ATP) channels are the ATP and ADP sensors in the regulation of glucose-induced insulin secretion. K(ATP) channels are also the target of sulfonylureas, which are widely used in the treatment of type 2 diabetes. Molecular cloning of the two subunits of the pancreatic beta-cell K(ATP) channel, Kir6.2 (an inward rectifier K+ channel member) and SUR1 (a receptor for sulfonylureas), has provided great insight into its structure and function. Kir6.2 subunits form the K+ ion-permeable pore and primarily confer inhibition of the channels by ATP, while SUR1 subunits confer activation of the channels by MgADP and K+ channel openers, such as diazoxide, as well as inhibition by sulfonylureas. The SUR1 subunits also enhance the sensitivity of the channels to ATP. To determine the physiological roles of K(ATP) channels directly, we have generated two kinds of genetically engineered mice: mice expressing a dominant negative form of Kir6.2 specifically in the pancreatic beta-cells (Kir6.2G132S Tg mice) and mice lacking Kir6.2 (Kir6.2 knockout mice). Studies of these mice elucidated various roles of the K(ATP) channels in endocrine pancreatic function: 1) the K(ATP) channels are the major determinant of the resting membrane potential of pancreatic beta-cells, 2) both glucose- and sulfonylurea-induced membrane depolarization of beta-cells require closure of the K(ATP) channels, 3) both glucose- and sulfonylurea-induced rises in intracellular calcium concentration in beta-cells require closure of the K(ATP) channels, 4) both glucose- and sulfonylurea-induced insulin secretions are mediated principally by the K(ATP) channel-dependent pathway, 5) the K(ATP) channels are important for beta-cell survival and architecture of the islets, 6) the K(ATP) channels are important in the differentiation of islet cells, and 7) the K(ATP) channels in glucose-responsive cells generally participate in coupling glucose sensing with cell excitability. Interestingly, despite the severe defect in glucose-induced insulin secretion, Kir6.2 knockout mice show only a very mild impairment in glucose tolerance. However, when the knockout mice become obese with age, they develop fasting hyperglycemia and glucose intolerance, while neither fasting hyperglycemia nor glucose intolerance is evident in the aged knockout mice without obesity, suggesting that both the genetic defect in glucose-induced insulin secretion and the acquired insulin resistance due to environmental factors are necessary to develop diabetes in Kir6.2 knockout mice. Thus, Kir6.2G132S Tg mice and Kir6.2 knockout mice provide a model of type 2 diabetes and clarify the various roles of K(ATP) channels in endocrine pancreatic function. PMID- 10868951 TI - Induction of fatty acid translocase/CD36, peroxisome proliferator-activated receptor-gamma2, leptin, uncoupling proteins 2 and 3, and tumor necrosis factor alpha gene expression in human subcutaneous fat by lipid infusion. AB - Little is known about the mechanisms involved in the preferential channeling of different fuels to fat and how the target tissue participates in this process. Dietary fatty acids have been shown to act as signaling molecules that bind and activate a new class of nuclear receptors, the peroxisome proliferator-activated receptors (PPARs). PPAR-gamma is particularly interesting because it may have the potential to link particular fatty acids with a program of gene expression involved in lipid storage and metabolism. We investigated whether a nutrient sensing pathway is activated by an increased availability of lipid fuels in nine normal weight male volunteers. Using reverse transcriptase-polymerase chain reaction analysis, the mRNA expression of fatty acid translocase (FAT)/CD36, PPAR gamma2, leptin, uncoupling protein (UCP)-2 and UCP-3, and tumor necrosis factor (TNF)-alpha was investigated in gluteal subcutaneous fat biopsies before and after 5 h infusions of saline or Intralipid (Pharmacia and Upjohn, Milan, Italy) plus heparin, which does not modify insulinemia. Marked increases in FAT/CD36 (724+/-18%; P < 0.05), PPAR-gamma2 (200+/-8%; P < 0.05), leptin (110+/-13%; P < 0.05), UCP-2 (120+/-7%; P < 0.05), UCP-3 (80+/-5%; P < 0.05), and TNF-alpha mRNA (130+/-12%; P < 0.05) were observed in comparison with pretreatment levels, whereas there was no change after saline infusion. These data suggest that the in vivo gene expression of FAT/CD36, PPAR-gamma2, leptin, UCP-2, UCP-3, and TNF alpha in subcutaneous adipose tissue is regulated by circulating lipids independent of insulin and that prolonged hyperlipidemia may therefore contribute to increased fat metabolism and storage as a result of the increased expression of these proteins. PMID- 10868952 TI - Insulin signaling and insulin sensitivity after exercise in human skeletal muscle. AB - Muscle glucose uptake, glycogen synthase activity, and insulin signaling were investigated in response to a physiological hyperinsulinemic (600 pmol/l) euglycemic clamp in young healthy subjects. Four hours before the clamp, the subjects performed one-legged exercise for 1 h. In the exercised leg, insulin more rapidly activated glucose uptake (half activation time [t1/2] = 11 vs. 34 min) and glycogen synthase activity (t1/2 = 8 vs. 17 min), and the magnitude of increase was two- to fourfold higher compared with the rested leg. However, prior exercise did not result in a greater or more rapid increase in insulin-induced receptor tyrosine kinase (IRTK) activity (t1/2 = 50 min), serine phosphorylation of Akt (t1/2 = 1-2 min), or serine phosphorylation of glycogen synthase kinase-3 (GSK-3) (t1/2 = 1-2 min) or in a larger or more rapid decrease in GSK-3 activity (t1/2 = 3-8 min). Thirty minutes after cessation of insulin infusion, glucose uptake, glycogen synthase activity, and signaling events were partially reversed in both the rested and the exercised leg. We conclude the following: 1) physiological hyperinsulinemia induces sustained activation of insulin-signaling molecules in human skeletal muscle; 2) the more distal insulin-signaling components (Akt, GSK-3) are activated much more rapidly than the proximal signaling molecules (IRTK as well as insulin receptor substrate 1 and phosphatidylinositol 3-kinase [Wojtaszewski et al., Diabetes 46:1775-1781, 1997]); and 3) prior exercise increases insulin stimulation of both glucose uptake and glycogen synthase activity in the absence of an upregulation of signaling events in human skeletal muscle. PMID- 10868953 TI - Constitutively active mitogen-activated protein kinase kinase increases GLUT1 expression and recruits both GLUT1 and GLUT4 at the cell surface in 3T3-L1 adipocytes. AB - To address a role of mitogen-activated protein kinase (MAPK) in the regulation of glucose transport, we made a constitutively active mutant of MAPK kinase (MAPKK) and introduced it into 3T3-L1 preadipocytes by using a retrovirus-mediated transfection procedure. The deletion of 20 amino acids (those between and including 32 and 51) in the amino terminal region of Xenopus MAPKK and the replacement of serine residues on the 218 and 222 positions by glutamic acid (dSESE-MAPKK) let Xenopus MAPKK constitutively active. The isolated cell clones differently expressing dSESE-MAPKK (clone 219 higher expression, clone 233 lower expression) efficiently differentiated to adipocytes by a standard differentiation cocktail. Accordingly, the increased expression of dSESE-MAPKK protein during differentiation resulted in the increased basal MAPK activity in clone 219 adipocytes and, to a lesser extent, in clone 233 adipocytes. In contrast to clone 233 and parental adipocytes, basal 2-deoxyglucose uptake was enhanced fourfold in clone 219 adipocytes, in accordance with increased expression of GLUT1 mRNA and protein. Whereas GLUT4 mRNA was similarly expressed in all of the adipocytes, GLUT4 protein appeared to decrease in clone 219 adipocytes. More importantly, subcellular fractionation studies showed that the localization of both GLUT1 and GLUT4 in the plasma membranes (PMs) was markedly increased in the basal state in clone 219 adipocytes compared with that in clone 233 and parental adipocytes, in which both glucose transporters were preferentially located in intracellular compartments. Consequently, insulin induced translocation of GLUT1 was abolished in clone 219 adipocytes, although the remaining intracellular GLUT4 was still responsive to insulin stimulation, which led to the movement to the PM. As combined effects on the situation of GLUT1 and GLUT4, the foldness of insulin stimulation of glucose transport based on the basal activity was reduced in cells expressing constitutively active MAPKK. These results imply that chronic activation of MAPK could be one of the mechanisms for insulin resistance. PMID- 10868954 TI - Interleukin-1beta-induced alteration in a beta-cell phenotype can reduce cellular sensitivity to conditions that cause necrosis but not to cytokine-induced apoptosis. AB - Previous work has shown that interleukin-1beta (IL-1beta) alters protein expression in beta-cells. This alteration is associated with cell death in isolated rat islets but not in isolated rat beta-cells. We examined whether IL 1beta pretreatment of isolated beta-cells influences their sensitivity to toxic agents. After a 24-h culture with IL-1beta (30 U/ml), beta-cells exhibited a lower expression of the beta-cell-specific protein transcription factor pancreatic and duodenal homeobox gene (PDX)-1, glucose transporter GLUT2, and proinsulin convertase PC2, with a marked reduction (60-70%) in glucose-induced insulin production and selective sensitivity to the toxins alloxan (ALX) and streptozotocin (STZ). On the other hand, the cells presented an increased expression of Mn-superoxide dismutase, heat shock protein 70, inducible heme oxygenase, and inducible nitrite oxide synthase. This IL-1beta-induced alteration in beta-cell phenotype resulted in a reduced cellular sensitivity to the beta cell-specific toxins ALX and STZ; the production of nontoxic conditions of nitric oxide (NO) also rendered the cells less susceptible to radical-induced damage. Exposure to IL-1beta can thus protect beta-cells against conditions that cause necrosis; however, it did not protect against apoptosis induced by the additional presence of interferon-gamma or tumor necrosis factor-alpha. Release of IL-1beta in the endocrine pancreas is thus not necessarily the cause of massive NO dependent beta-cell destruction. On the contrary, IL-1beta may protect these cells against necrosis, though with a loss of their characteristic phenotype and homeostatic functions. PMID- 10868955 TI - Anti-inflammatory actions of 15-deoxy-delta 12,14-prostaglandin J2 and troglitazone: evidence for heat shock-dependent and -independent inhibition of cytokine-induced inducible nitric oxide synthase expression. AB - In this study, the anti-inflammatory actions of the peroxisome proliferator activated receptor (PPAR)-gamma agonists 15-deoxy-delta 12,14-prostaglandin J2 (15-d-delta 12,14-PGJ2) and troglitazone have been examined. Treatment of RAW 264.7 cells and CD-1 mouse peritoneal macrophages with lipopolysaccharide (LPS) + interferon-gamma (IFN-gamma) results in inducible nitric oxide synthase (iNOS), inducible cyclooxygenase (COX-2) and interleukin-1 (IL-1) expression, increased production of nitric oxide, and the release of IL-1. In a concentration-dependent manner, 15-d-delta 12,14-PGJ2 inhibits each of these proinflammatory actions of LPS + IFN-gamma, with half-maximal inhibition at approximately 0.5 microg/ml and complete inhibition at 1-5 microg/ml. The inhibitory actions of 15-d-delta 12,14 PGJ2 on LPS + IFN-gamma-induced inflammatory events are not associated with the inhibition of iNOS enzymatic activity or macrophage cell death, but appear to result from an inhibition of iNOS and IL-1 transcription. In addition, the anti inflammatory actions of 15-d-delta 12,14-PGJ2 are not limited to peritoneal macrophages, as 15-d-delta 12,14-PGJ2 prevents TNF-alpha + LPS-induced resident islet macrophage expression of IL-1beta and beta-cell expression of iNOS stimulated by the local release of IL-1 in rat islets. 15-d-delta 12,14-PGJ2 appears to be approximately 10-fold more effective at inhibiting resident islet macrophage activation (in response to TNF + LPS) than IL-1-induced nitrite production by beta-cells. Two mechanisms appear to be associated with the antiinflammatory actions of both 15-d-delta 12,14-PGJ2 and troglitazone: 1) the direct inhibition of cytokine- and endotoxin-stimulated iNOS and IL-1 transcription; and 2) the inhibition of IL-1 signaling, an event associated with PPAR-gamma agonist-induced activation of the heat shock response (as assayed by heat shock protein 70 expression). These findings indicate that the PPAR-gamma agonists, troglitazone and the J series of prostaglandins, are potent anti inflammatory agents that prevent cytokine- and endotoxin-stimulated activation of peripheral and resident tissue macrophages and cytokine-induced iNOS expression by beta-cells by the inhibition of transcriptional activation and induction of the heat shock response. PMID- 10868956 TI - T-cell lines reactive to an immunodominant epitope of the tyrosine phosphatase like autoantigen IA-2 in type 1 diabetes. AB - Type 1 diabetes is the result of destruction of the insulin-secreting beta-cells of the pancreas by a process in which T-cells play a central role. A tyrosine phosphatase-like protein, IA-2, is a major target for autoantibodies and T-cells in the disease. In this study, we have further characterized the T-cell response to IA-2 by the generation and characterization of T-cell lines. T-cell lines responsive to IA-2 antigen were generated from 17 of 32 patients and 3 of 10 control subjects. Antigen specificity was confirmed in lines from six diabetic patients and one control individual by demonstration of responses to synthetic IA 2 peptides and epitope mapping. Five lines from diabetic patients responded to one of two peptides representing amino acids 831-850 and 841-860 of IA-2. The overlapping portion may therefore represent an immunodominant region of the molecule. The sixth patient-derived line responded to a peptide representing amino acids 751-770 of IA-2 presented by the DR 4 (DRB1*0401) allele that confers susceptibility to type 1 diabetes. Primary T-cell responses to peptides of the immunodominant region were detected in 9 of 19 (47%) type 1 diabetic patients and 16 of 22 (73%) nondiabetic siblings, consistent with this region having immunostimulatory properties. The study reports for the first time T-cell lines reactive to IA-2 from diabetic patients and defines an immunodominant region of the molecule. PMID- 10868957 TI - Zinc as a paracrine effector in pancreatic islet cell death. AB - Because of a huge amount of Zn2+ in secretory granules of pancreatic islet beta cells, Zn2+ released in certain conditions might affect the function or survival of islet cells. We studied potential paracrine effects of endogenous Zn2+ on beta cell death. Zn2+ induced insulinoma/islet cell death in a dose-dependent manner. Chelation of released endogenous Zn2+ by CaEDTA significantly decreased streptozotocin (STZ)-induced islet cell death in an in vitro culture system simulating in vivo circumstances but not in the conventional culture system. Zn2+ chelation in vivo by continuous CaEDTA infusion significantly decreased the incidence of diabetes after STZ administration. N-(6-methoxy-quinolyl)-para toluene-sulfonamide staining revealed that Zn2+ was densely deposited in degenerating islet cells 24 h after STZ treatment, which was decreased by CaEDTA infusion. We show here that Zn2+ is not a passive element for insulin storage but an active participant in islet cell death in certain conditions, which in time might contribute to the development of diabetes in aged people. PMID- 10868958 TI - Accurate measurement of endogenous insulin secretion does not require separate assessment of C-peptide kinetics. AB - The implication of beta-cell failure as an early defect in type 2 diabetes exacerbates the need for accurate but facile assessment of islet cell secretory rate, particularly in large group studies in which individual assessment of C peptide kinetics is impractical. This study was designed to examine whether it is possible to obtain accurate secretory rates from the extended combined model, which provides insulin and C-peptide kinetics from plasma measurements of the two peptides. Equimolar intraportal infusions of insulin and C-peptide that are designed to simulate insulin secretion rates during both oral and intravenous glucose tolerance tests were used to generate plasma insulin and C-peptide data in conscious dogs that were examined under clamped glucose conditions. The plasma peptide kinetics were analyzed using the extended combined model to generate estimates of prehepatic insulin secretion that were then compared with the known intraportal infusion rates. The extended combined model was able to reproduce the known intraportal infusion profiles. The model-predicted rates were similar to those calculated with methods that require separate assessment of C-peptide kinetics. Simulation results supported lesser clearance of insulin during rapid changes of portal insulin (as measured by an intravenous glucose tolerance test) versus slow changes in portal insulin (as measured by an oral glucose tolerance test). The extended combined model accurately calculates prehepatic insulin appearance. It may be possible to apply this approach to large studies of beta cell function designed to identify changes in islet function in subjects at risk for diabetes. Such an approach could strengthen epidemiological and genetic studies of the pathogenesis of diabetes. PMID- 10868959 TI - Synaptotagmin III isoform is compartmentalized in pancreatic beta-cells and has a functional role in exocytosis. AB - Synaptotagmin is involved in Ca2+-regulated secretion and has been suggested to serve as a general Ca2+ sensor on the membrane of secretory vesicles in neuronal cells. Insulin exocytosis from the pancreatic beta-cell is an example of a Ca2+ dependent secretory process. Previous studies of pancreatic beta-cells were unable to show presence of synaptotagmin I. We now present biochemical and immunohistochemical data showing that synaptotagmin III is present in pancreatic beta-cells as well as in the insulin-secreting cell line HIT-T15 and in rat insulinoma. By subcellular fractionation, we found synaptotagmin III in high density fractions together with insulin and secretogranin I, indicating colocalization of synaptotagmin III and insulin in secretory granules. We could also show that blockade of synaptotagmin III by a specific antibody inhibited Ca2+-induced changes in beta-cell membrane capacitance, suggesting that synaptotagmin III is part of the functional protein complex regulating beta-cell exocytosis. The synaptotagmin III antibody did not affect the activity of the voltage-gated L-type Ca2+-channel. These findings are compatible with the view that synaptotagmin III, because of its distinct localization in the pancreatic beta-cell, functionally modulates insulin exocytosis. This indicates that synaptotagmin may have a general role in the regulation of exocytosis not only in neuronal cells but also in endocrine cells. PMID- 10868960 TI - Quantitative and functional characterization of muscarinic receptor subtypes in insulin-secreting cell lines and rat pancreatic islets. AB - Expression of muscarinic receptors in rat islets, RINm5F cells, and INS-1 cells was established by reverse transcriptase-polymerase chain reaction (RT-PCR) and quantified by RNase protection. Both methods indicated that m3 and m1 receptors were expressed approximately equally in the various cellular preparations and to a much greater extent than the m5 subtype. However, the cell lines, especially RINm5F cells, expressed less of a given receptor subtype than did islets. Immunohistochemistry indicated that m3 receptors were expressed throughout the islet core. Binding studies using the radiolabeled muscarinic receptor antagonist QNB demonstrated a maximal binding capacity of INS-1 cells of 23.0+/-2.9 fmol/mg protein. Functional analyses were undertaken using INS-1 cells stably transfected with either m1 or m3 receptor cDNAs. Overexpression of either receptor did not affect basal responses but markedly enhanced maximal responses to the muscarinic receptor agonist carbachol. Although maximal hydrolysis of phosphatidylinositol 4,5-bisphosphate (Ptd InsP2) was twofold greater in m1-transfectants as compared with m3-transfectants, cell lines overexpressing either receptor gave essentially equivalent secretory responses to a full range of carbachol doses. The results demonstrate that both m1 and m3 muscarinic receptors are well expressed in pancreatic beta-cells, functionally linked to signaling pathways, and capable of initiating insulin secretion with equal potencies. PMID- 10868961 TI - Prolonged elevation of plasma free fatty acids impairs pancreatic beta-cell function in obese nondiabetic humans but not in individuals with type 2 diabetes. AB - Our recent in vivo observations in healthy nonobese humans have demonstrated that prolonged elevation of plasma free fatty acids (FFAs) results in diminished glucose-stimulated insulin secretion (GSIS) when the FFA-mediated decrease in insulin sensitivity is taken into account. In the present study, we investigated whether obese individuals and patients with type 2 diabetes are more sensitive than healthy control subjects to the inhibitory effect of prolonged elevation of plasma FFAs on GSIS. In seven patients with type 2 diabetes and seven healthy nondiabetic obese individuals, we assessed GSIS with a programmed graded intravenous glucose infusion on two occasions, 6-8 weeks apart, with and without a prior 48-h infusion of heparin and Intralipid, which was designed to raise plasma FFA concentration approximately twofold over basal. The nondiabetic obese subjects had a significant 21% decrease in GSIS (P = 0.0008) with the heparin and Intralipid infusion, associated with a decrease in whole body insulin clearance. The impairment in GSIS was evident at low (<11 mmol/l) but not at higher plasma glucose concentrations. In contrast, the patients with type 2 diabetes had a slight increase in GSIS (P = 0.027) and no change in insulin clearance, although there was marked interindividual variability in response. Plasma proinsulin concentrations measured in a subset of subjects were not altered in either group by the infusion of heparin and Intralipid. In summary, 1) obese nondiabetic individuals are susceptible to a desensitization of GSIS with heparin and Intralipid infusion, and 2) patients with type 2 diabetes do not demonstrate such susceptibility when FFAs are elevated approximately twofold above basal with heparin and Intralipid. Our results suggest that FFAs could play an important role in the development of beta-cell failure in obese individuals who are at risk for developing type 2 diabetes. They do not, however, seem to further deteriorate the beta-cell function of patients who already have established type 2 diabetes and may even result in a slight increase in GSIS in this latter group. PMID- 10868962 TI - The extracellular calcium-sensing receptor on human beta-cells negatively modulates insulin secretion. AB - The presence and functional significance of the extracellular calcium-sensing receptor (CaR) on human pancreatic beta-cells were investigated. Reverse transcriptase-polymerase chain reaction with primers for the extracellular domain of the CaR expressed in human parathyroid-secreting cells identified a product of the expected size in human pancreatic mRNA. Immunocytochemistry using an antibody against the extracellular region of CaR showed extensive immunoreactivity in insulin- and glucagon-containing cells but not in somatostatin-containing cells. In perifusion experiments, elevations in extracellular Ca2+ produced initial transient increases in insulin secretion, followed by a concentration-dependent and prolonged, but reversible, inhibition of secretion. Microfluorometric measurements of intracellular Ca2+ ([Ca2+]i) in isolated human beta-cells demonstrated that elevations in extracellular Ca2+ (0.5-10 mmol/l) caused rapid elevations in [Ca2+]i. Increases in extracellular Ca2+ caused small increases in the cyclic AMP content of whole human islets. These studies demonstrated that human beta-cells express an extracellular CaR and that activation of the receptor inhibits basal and nutrient-stimulated insulin secretion. The transduction mechanism that mediates this inhibitory effect is unknown, but our results suggest that it is unlikely to be through the adenylate cyclase-cyclic AMP pathway or through the phospholipase C-IP3 pathway. This CaR-mediated inhibitory mechanism may be an important autoregulatory mechanism in the control of insulin secretion. PMID- 10868963 TI - Glucose modulation of insulin mRNA levels is dependent on transcription factor PDX-1 and occurs independently of changes in intracellular Ca2+. AB - Glucose regulates insulin production in pancreatic beta-cells in the long term by stimulating insulin gene transcription. These effects are partially mediated through the activity of a homeodomain transcription factor, PDX-1, which binds to four sites within the human insulin gene promoter. The availability of a human beta-like cell line, NES2Y, which lacks PDX-1 but expresses the insulin gene, allowed us to determine whether PDX-1 was essential for the stimulatory effect of glucose on insulin mRNA levels. In NES2Y cells, glucose had no effect on the insulin gene promoter linked to a firefly luciferase reporter or on endogenous insulin mRNA levels. However, in NES2Y cells stably transfected with PDX-1 (NES PDX-1), glucose exhibited a marked stimulatory effect on both the insulin promoter (5+/-0.2-fold, n = 6) and insulin mRNA levels (4.8+/-0.5-fold, n = 4). NES2Y cells were derived from a patient with persistent hyperinsulinemic hypoglycemia of infancy; the cells therefore lacked operational ATP-sensitive potassium channels, which results in the failure to control depolarization dependent intracellular Ca2+ signaling. Despite the loss of control of Ca2+ channel activity, NES-PDX-1 cells maintained normal glucose-responsive insulin gene regulation. These results demonstrate that glucose modulation of insulin mRNA levels is dependent on the activity of PDX-1 and that these effects are independent of changes in intracellular Ca2+ concentrations. PMID- 10868964 TI - Isolation of INS-1-derived cell lines with robust ATP-sensitive K+ channel dependent and -independent glucose-stimulated insulin secretion. AB - The biochemical mechanisms involved in regulation of insulin secretion are not completely understood. The rat INS-1 cell line has been used to gain insight in this area because it secretes insulin in response to glucose concentrations in the physiological range. However, the magnitude of the response is far less than that seen in freshly isolated rat islets. In the current study, we have stably transfected INS-1 cells with a plasmid containing the human proinsulin gene. After antibiotic selection and clonal expansion, 67% of the resultant clones were found to be poorly responsive to glucose in terms of insulin secretion (< or =2 fold stimulation by 15 mmol/l compared with 3 mmol/l glucose), 17% of the clones were moderately responsive (2- to 5-fold stimulation), and 16% were strongly responsive (5- to 13-fold stimulation). The differences in responsiveness could not be ascribed to differences in insulin content. Detailed analysis of one of the strongly responsive lines (832/13) revealed that its potent response to glucose (average of 10-fold) was stable over 66 population doublings (approximately 7.5 months of tissue culture) with half-maximal stimulation at 6 mmol/l glucose. Furthermore, in the presence of 15 mmol/l glucose, insulin secretion was potentiated significantly by 100 pmol/l isobutylmethylxanthine (320%), 1 mmol/l oleate/palmitate (77%), and 50 nmol/l glucagon-like peptide 1 (60%), whereas carbachol had no effect. Glucose-stimulated insulin secretion was also potentiated by the sulfonylurea tolbutamide (threefold at 3 mmol/l glucose and 50% at 15 mmol/l glucose) and was abolished by diazoxide, which demonstrates the operation of the ATP-sensitive K+ channel (K(ATP)) in 832/13 cells. Moreover, when the K(ATP) channel was bypassed by incubation of cells in depolarizing K+ (35 mmol/l), insulin secretion was more effectively stimulated by glucose in 832/13 cells than in parental INS-1 cells, which demonstrates the presence of a K(ATP) channel-independent pathway of glucose sensing. We conclude that clonal selection of INS-1 cells allows isolation of cell lines that exhibit markedly enhanced and stable responsiveness to glucose and several of its known potentiators. These lines may be attractive new vehicles for studies of beta-cell function. PMID- 10868965 TI - Impaired leptin responsiveness in aged rats. AB - We previously reported that adiposity and serum leptin levels increase with age in male F-344xBN rats and that when physiological levels of serum leptin are manipulated by fasting, there is a corresponding reciprocal change in hypothalamic neuropeptide Y (NPY) mRNA in young rats, but there are no changes in older rats. These findings suggest that the regulation of hypothalamic NPY mRNA by leptin may be impaired with age. To test this hypothesis, we infused saline or leptin for 7 days into ad libitum-fed rats and compared these with saline-infused rats that were pair-fed the amount of food consumed by the leptin-treated rats. We examined daily food consumption, body weight, whole-body oxygen consumption, serum leptin, and NPY mRNA in the hypothalamus. Food consumption decreased by 50% in the leptin-infused compared with the saline-infused young rats but only decreased by 20% in the aged rats. In the leptin-treated young rats, there was a 24% increase in oxygen consumption compared with the pair-fed rats, but there were no changes in oxygen consumption in the aged rats. Leptin infusion diminished hypothalamic NPY levels by nearly 50% compared with pair-fed young rats, whereas there were no changes in the hypothalamic NPY mRNA levels in senescent rats. In summary, aged rats demonstrate a reduced responsiveness to leptin, including a diminished decrease in food intake and no increase in energy expenditure. These diminished responses to leptin were associated with and may be the result of an impaired suppression of hypothalamic NPY mRNA levels. This leptin resistance may be due to either the elevated obesity and serum leptin with age or due to age itself, or both. PMID- 10868966 TI - Brain-derived neurotrophic factor regulates glucose metabolism by modulating energy balance in diabetic mice. AB - We previously reported that brain-derived neurotrophic factor (BDNF) regulates both food intake and blood glucose metabolism in rodent obese diabetic models such as C57BL/KsJ-lepr(db)/lepr(db) (db/db) mice. To elucidate the effect of BDNF on glucose metabolism, we designed a novel pellet pair-feeding apparatus to eliminate the effect of appetite alteration on glucose metabolism. The apparatus was used to synchronize food intake precisely between BDNF-treated and vehicle treated db/db mice. It was shown using this pellet pair-feeding apparatus that BDNF administered daily (20 mg x kg(-1) x day(-1)) to db/db mice significantly lowered blood glucose compared with pellet pair-fed db/db mice. To evaluate the effect of BDNF on insulin action, we used streptozotocin-induced type 1 diabetic mice. In this case, BDNF did not lower blood glucose concentration but rather enhanced the hypoglycemic action of insulin. In hyperglycemic db/db mice, pancreatic insulin content was reduced and glucagon content was increased compared with normoglycemic db/m mice. BDNF administered to db/db mice significantly restored both pancreatic insulin and glucagon content. Histological observations of aldehyde-fuchsin staining and immunostaining with anti-insulin indicated that insulin-positive pancreatic beta-cells were extensively regranulated by BDNF administration. We also studied the effect of BDNF on KK mice, normoglycemic animals with impaired glucose tolerance. In these mice, BDNF administration improved insulin resistance in the oral glucose tolerance test. To elucidate how blood glucose was metabolized in BDNF-treated animals, we investigated the effect of BDNF on the energy metabolism of db/db mice. Body temperature and oxygen consumption of the pellet pair-fed vehicle-treated mice were remarkably lower than the ad libitum-fed vehicle-treated mice. Daily BDNF administration for 3 weeks completely ameliorated both of the reductions. Finally, to clarify its action mechanism, the effect of intracerebroventricular administration of BDNF on db/db mice was examined. Here, a small dose of BDNF was found to be effective in lowering blood glucose concentration. This indicates that BDNF regulates glucose metabolism by acting directly on the brain. PMID- 10868967 TI - Type 2 diabetes and low birth weight: the role of paternal inheritance in the association of low birth weight and diabetes. AB - Lower birth weight is associated with an increased occurrence of type 2 diabetes in later life. Whether this relationship is explained by environmental or genetic factors is unknown. We have examined the potential for genetic influences by determining whether parental diabetes is associated with lower birth weight in 1,608 children of known birth weight and gestational age born between 1941 and 1993 in the Gila River Indian Community in Arizona. The previously described relationships of maternal diabetes to increased birth weight and offspring diabetes were observed. In contrast to this we have determined novel relationships between low birth weight and paternal diabetes. The offspring of diabetic fathers were, on average, 78 g lighter than the offspring of nondiabetic fathers. For fathers, lower birth weight in their offspring was associated with an increased risk of later diabetes, i.e., fathers of offspring in the lowest quintile of birth weight, who were not diabetic at the time of birth of their child, had a 1.8-fold increased risk of developing diabetes later in life (95% CI 1.2-2.7; P = 0.004). For children, lower birth weight predicted diabetes in the offspring if paternal but not maternal diabetes was present, but it was not associated with higher plasma glucose if neither parent had diabetes. We conclude that the risk of diabetes associated with low birth weight is strongly related to the development of paternal diabetes, suggesting a genetic link between lower birth weight and later diabetes. PMID- 10868968 TI - Renal glucose production compensates for the liver during the anhepatic phase of liver transplantation. AB - The extent of the renal contribution to postabsorptive endogenous glucose production (EGP) in humans is controversial. We measured EGP in the absence of the liver during the anhepatic phase (AH) of liver transplantation in five patients (aged 46.4+/-10.2 years, two women). Stable labeling of plasma glucose (PG) was achieved for a 2-h period before the AH by primed continuous infusion of di-deuterated 6,6[2H2]glucose (1.7 mg/min) and continued throughout the AH. PG was maintained above the fasting level (6.1+/-2.73 mmol/l) with 5% dextrose labeled with 6,6[2H2]glucose throughout the AH (mean level during the AH 0.98+/ 0.45 mg x kg(-1) x min(-1)). Isotopic enrichment remained stable at 0.84+/-0.21% atom percent excess throughout. EGP, calculated by use of a modified Steele equation, decreased from 2.6+/-1.24 at baseline to 0.97+/-0.9 mg x kg(-1) x min( 1) (36% baseline, P = 0.045) but recovered at approximately 30 min to reach 1.38+/-0.83 mg x kg(-1) x min(-1) (54% baseline) by 60 min. Epinephrine, lactate, free fatty acid, and glycerol levels increased significantly (0.79+/-0.74 to 3.65+/-2.1 nmol/l, P = 0.005; 1.88+/-0.43 to 3.46+/-0.9 mmol/l, P = 0.024; 543.9+/-215.5 to 705.5+/-219.2 micromol/l, P = 0.012; 75.6+/-30.2 to 139+/-96.3 micromol/l, P = 0.003, respectively). These data show that postabsorptive nonhepatic glucose production in humans may contribute to greater than one-third of overall EGP, increasing when required, and that it is associated with a stress response and increased gluconeogenic substrate availability. We conclude that extrahepatic tissues, most notably those of the kidney, make a significant contribution to EGP in humans. PMID- 10868969 TI - Dysregulation of the insulin/IGF binding protein-1 axis in transgenic mice is associated with hyperinsulinemia and glucose intolerance. AB - The insulin/IGF binding protein-1 (IGFBP-1) axis is important in coordinating insulin- and IGF-mediated regulation of glucose metabolism and glycemia. Dysregulation of the axis may play a role in the pathophysiology of disorders of insulin deficiency and resistance. We have investigated this hypothesis by generating transgenic mice that overexpress hIGFBP-1. To study the axis in its true physiological context, we used a human (h) IGFBP-1 cosmid clone so that transgene expression is responsive to normal hormonal stimuli. hIGFBP-1 mRNA is expressed in a tissue-specific fashion, and measurement of serum protein levels by specific immunoassay indicates normal physiological regulation in response to fasting/feeding and appropriate post-translational modification as indicated by the detection of phosphorylated and nonphosphorylated isoforms of the protein. The hypoglycemic response to exogenous IGF-I is attenuated in transgenic mice. Transgenic mice exhibit an enhanced insulin secretory response to a glucose challenge, although basal and stimulated blood glucose levels are similar to controls. There is a sexual dimorphism in phenotypic expression: male transgenic mice had higher stimulated glucose and insulin levels than did females. Transgenic mice exhibit fasting hyperglycemia and hyperinsulinemia and glucose intolerance in later life, indicating an age-related decline in glucocompetence. These findings demonstrate the importance of the normal inverse relationship between serum insulin and IGFBP-1 levels in glucoregulation and that sustained dysregulation of the insulin/IGF-I/IGFBP-1 axis is associated with impaired glucose tolerance and abnormalities of insulin action. PMID- 10868970 TI - Early glomerular macrophage recruitment in streptozotocin-induced diabetic rats. AB - Diabetic glomerulosclerosis is defined by increased glomerular extracellular matrix (ECM) that is mainly synthesized by mesangial cells that underwent an activation mediated by cytokines and growth factors from various cellular origins. In this study, we tested whether macrophages could infiltrate the glomeruli and influence ECM synthesis in experimental diabetes. To test our hypothesis, we initially studied the dynamics of glomerular macrophage recruitment in streptozotocin-induced diabetic rats at days 1, 2, 4, 8, 15, and 30 by using quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) on isolated glomeruli and immunohistochemistry and morphometry. We then assessed the role of macrophages on the basis of the pharmacological modulation of their recruitment by insulin or ACE inhibitor treatments and by X-irradiation-induced macrophage depletion at days 8 and 30. Macrophages were recruited within the glomeruli at the very early phase of hyperglycemia by using RT-PCR CD14 detection from day 2 and by using ED1 immunohistochemistry from day 8. This glomerular macrophage infiltration was associated with an increase in alpha1-chain type IV collagen mRNA. In parallel, the diabetic glomeruli became hypertrophic with an increase in the mesangial area. Macrophage recruitment was preceded by or associated with an increased glomerular expression of vascular cell adhesion molecule 1, intracellular adhesion molecule 1, and monocyte chemoattractant protein 1, which contributes to monocyte diapedesis. Glomerular interleukin-1beta mRNA synthesis was also enhanced as early as day 1 and could be involved in the increase in ECM and adhesion molecule gene expressions. Insulin treatment and irradiation-induced macrophage depletion completely prevented the glomerular macrophage recruitment and decreased alpha1-chain type IV collagen mRNA and mesangial area in diabetic rats, whereas ACE inhibitor treatment had an incomplete effect. It can be concluded that in the streptozotocin model, hyperglycemia is followed by an early macrophage recruitment that contributes to the molecular and structural events that could lead to glomerulosclerosis. Therefore, besides direct stimulation of mesangial cells by hyperglycemia, macrophages recruited in the glomeruli during the early phase of hyperglycemia could secondarily act on mesangial cells. PMID- 10868971 TI - Course of renal function in type 2 diabetic patients with abnormalities of albumin excretion rate. AB - Heterogeneity in renal structure has been described in type 2 diabetic patients with both microalbuminuria and proteinuria; in fact, only a subset of type 2 diabetic patients have the typical diabetic glomerulopathy. However, it is currently unknown whether abnormalities in albumin excretion rate (AER) have a different renal prognostic value depending on the underlying renal structure. Aims of this study were: 1) to study the course of renal function in type 2 diabetic patients with altered AER; 2) to evaluate the relationship between the course of glomerular filtration rate (GFR) and renal structure; and 3) to evaluate the relationship between the course of GFR and baseline AER levels, metabolic control, and blood pressure levels during a follow-up period of 4 years. A total of 108 type 2 diabetic patients, 74 with microalbuminuria (MA) and 34 with proteinuria (P), were recruited into a prospective study that encompassed: 1) a baseline kidney biopsy with morphometric measurements of glomerular parameters; 2) intensified antihypertensive treatment for an average 4 year period (blood pressure target <140/90 mmHg); and 3) determinations of GFR at baseline and every 6 months. Mean (+/- SD) GFR significantly decreased from baseline in both MA (-1.3+/-9.4 [95% CI -3.51 to +0.86], P < 0.05) and P (-3.0+/ 13.0 ml x min(-1) x 1.73 m(-2) per year [-7.71 to +1.61], P < 0.01). However, the changes in GFR were quite heterogeneous. Thus, on the basis of percent GFR change per year from baseline (delta%GFR), both MA and P patients were defined as progressors or nonprogressors when they were below or above the median, respectively. Baseline parameters of glomerular structure had a strong influence on the course of GFR. Indeed, the odds ratios of being progressors significantly increased across the quartiles of baseline glomerular basement membrane (GBM) width and mesangial fractional volume [Vv(mes/glom)], being 2.71 and 2.85 higher, respectively, in the fourth quartile than in the first quartile (P < 0.01 for both). Conversely, nonprogressors outnumbered progressors in the first quartile of GBM width (odds ratio: 2.14, P < 0.05) and in the first quartile of Vv(mes/glom) (odds ratio: 2.28, P < 0.01). Baseline albumin excretion rate (AER) did not influence delta%GFR; in fact, the number of progressors did not increase across quartiles of baseline AER among either MA or P. Similarly, mean blood pressure levels during follow-up (and intensified antihypertensive therapy) did not affect the course of GFR: the number of progressors and nonprogressors did not change across quartiles of mean blood pressure. In contrast, HbA1c during follow-up had an impact on delta%GFR: the odds ratio for being a progressor increased across quartiles of HbA1c, particularly for the highest quartile (HbA1c >9.0%). In conclusion, the course of renal function is heterogeneous in type 2 diabetic patients with microalbuminuria or proteinuria. In fact, a subset of patients has a rapid decline in GFR over a 4-year follow-up period; these patients have more advanced diabetic glomerulopathy and worse metabolic control than the remaining patients, whose GFR remains stable. These two cohorts are otherwise undistinguishable as regards the degree of AER at baseline and tight blood pressure control. Kidney biopsy has an important prognostic role in these patients. Thus, tight blood pressure control, when not associated with satisfactory glycemic control, is unable to prevent rapid GFR decline in type 2 diabetic patients with typical diabetic glomerulopathy. PMID- 10868972 TI - Increased levels of soluble vascular cell adhesion molecule 1 are associated with risk of cardiovascular mortality in type 2 diabetes: the Hoorn study. AB - Membrane-bound vascular cell adhesion molecule 1 (VCAM-1) allows the tethering and rolling of monocytes and lymphocytes as well as firm attachment and transendothelial migration of leukocytes. Soluble forms of VCAM (sVCAM-1) may serve as monitors of increased expression of membrane-bound VCAM-1 and thus may reflect progressive formation of atherosclerotic lesions. Levels of sVCAM-1 have been found to be increased among type 2 diabetic as compared with nondiabetic subjects. To study the association of plasma sVCAM-1 concentration and risk of cardiovascular and all-cause mortality among nondiabetic and diabetic subjects, we investigated an age-, sex-, and glucose-tolerance-stratified sample (n = 631) of a population-based cohort aged 50-75 years that was followed prospectively. Plasma levels of sVCAM-1 were determined in frozen -70 degrees C baseline samples. After 7.4 years (mean) of follow-up, 107 (17%) subjects had died (42 of cardiovascular causes). In the entire group, increased sVCAM-1 levels were significantly associated with increased risk of cardiovascular mortality (relative risks [RRs] per 100 ng/ml sVCAM-1 increase, 1.10 [1.05-1.15] after adjustment for age, sex, and glucose tolerance status). This RR was somewhat diminished by further adjustment for the presence of hypertension and cardiovascular disease; levels of total, HDL, and LDL cholesterol and homocysteine; the presence of microalbuminuria (a putative marker of endothelial dysfunction); levels of von Willebrand factor (a marker of endothelial dysfunction) and C-reactive protein (a marker of low-grade inflammation); and estimates of glomerular filtration rate. However, the RR remained statistically significant. The RR among type 2 diabetic subjects was 1.13 (1.07-1.20) per 100 ng/ml sVCAM-1 increase after adjustment for age and sex, which was somewhat higher but not significantly different from the RR in nondiabetic subjects (P value for interaction term, 0.12). Further adjustment for other risk factors gave similar results. In conclusion, levels of sVCAM-1 are independently associated with the risk of cardiovascular mortality in type 2 diabetic subjects and therefore might be useful for identifying subjects at increased cardiovascular risk. Increased plasma sVCAM-1 levels may reflect progressive formation of atherosclerotic lesions, or sVCAM-1 itself may have bioactive properties related to cardiovascular risk. Our data, however, argue against the hypotheses of sVCAM 1 levels simply being a marker of endothelial dysfunction, of low-grade inflammation, or of an impaired renal function. PMID- 10868973 TI - Genetic and physical mapping of a type 1 diabetes susceptibility gene (IDDM12) to a 100-kb phagemid artificial chromosome clone containing D2S72-CTLA4-D2S105 on chromosome 2q33. AB - Polymorphic markers within the CTLA4 gene on chromosome 2q33 have been shown to be associated with type 1 diabetes. Therefore, a gene responsible for the disease (IDDM12) most likely lies within a region of <1-2 cM of CTLA4. To define more precisely the IDDM12 interval, we genotyped a multiethnic (U.S. Caucasian, Mexican-American, French, Spanish, Korean, and Chinese) collection of 178 simplex and 350 multiplex families for 10 polymorphic markers within a genomic interval of approximately 300 kb, which contains the candidate genes CTLA4 and CD28. The order of these markers (D2S346, CD28, GGAA19E07, D2S307, D2S72, CTLA4, D2S105, and GATA52A04) was determined by sequence tagged site content mapping of bacterial artificial chromosome (BAC) and yeast artificial chromosome (YAC) clones. The transmission disequilibrium test (TDT) analyses of our data revealed significant association/linkage with three markers within CTLA4 and two immediate flanking markers (D2S72 and D2S105) on each side of CTLA4 but not with more distant markers including the candidate gene CD28. Tsp analyses revealed significant association only with the three polymorphic markers within the CTLA4 gene. The markers linked and associated with type 1 diabetes are contained within a phagemid artificial chromosome clone of 100 kb, suggesting that the IDDM12 locus is either CTLA4 or an unknown gene in very close proximity. PMID- 10868974 TI - Association of the nitric oxide synthase gene polymorphism with an increased risk for progression to diabetic nephropathy in type 2 diabetes. AB - A mutation of endothelial nitric oxide synthase (ecNOS)-a key enzyme of the endogenous nitrovasodilator system that is essential for the regulation of blood flow and blood pressure-may aggravate the progression to diabetic nephropathy and/or retinopathy. To investigate the association of ecNOS tandem repeat polymorphism with diabetic nephropathy, the ecNOS genotype was assessed in 82 Japanese type 2 diabetic patients without nephropathy, 94 patients with microalbuminuria, 39 patients with nephropathy, and 155 healthy control subjects. The analysis revealed that type 2 diabetic patients with nephropathy (not with microalbuminuria) were significantly different from type 2 diabetic patients without nephropathy and healthy control subjects in genotype distribution (P = 0.0423) and frequency of the ecNOS4a allele (19.2% vs. 7.3 and 7.1%, respectively; P = 0.0078). The odds ratio of progression to diabetic nephropathy in diabetic patients who carry the mutated allele is about 2.87 compared with noncarriers. The stepwise multiple regression analysis in these patients showed that hypertension (F = 9.760) and ecNOS gene polymorphism (F = 5.298) are the relevant variables for nephropathy. However, no association was found between the ecNOS4a allele and hypertension or diabetic retinopathy. These results imply that the ecNOS gene polymorphism may be associated with progression to diabetic nephropathy in Japanese type 2 diabetic patients. PMID- 10868975 TI - Vitamin D receptor allele combinations influence genetic susceptibility to type 1 diabetes in Germans. AB - Vitamin D has been shown to exert manifold immunomodulatory effects. Because type 1 diabetes is regarded to be immune-mediated and vitamin D prevents the development of diabetes in the NOD mouse, we investigated the role of the vitamin D receptor (VDR) gene as a candidate for type 1 diabetes susceptibility. A total of 152 Caucasian families with at least one affected offspring were genotyped for four VDR restriction-site polymorphisms (FokI, BsmI, ApaI, and TaqI). Whereas the BsmI, ApaI, and TaqI polymorphisms are in strong linkage disequilibrium with each other, no significant linkage disequilibrium with the FokI site was observed. Extended transmission disequilibrium testing (ETDT) was used to detect preferential transmission of allelic combinations to affected offspring. We found significant haplotype-wise ETDT results for the BsmI/ApaI/TaqI (chi2 = 18.886, df = 7, P = 0.0086), the BsmI/TaqI (chi2 = 8.373, df = 3, P = 0.0389), and theApaI/TaqI (chi2 = 17.182, df = 3, P = 0.0006) haplotypes. The "At" and "Bt" alleles confer an increased risk, whereas "AT" and "at" are protective. The combination with the strongest susceptibility was the "BAt" haplotype (64% transmitted, P = 0.0106). Analysis of the FokI site does not provide more information on susceptibility (FokI/BsmI/ApaI/TaqI [chi2 = 24.702, df = 15, P = 0.0541]). These findings suggest a linkage of VDR itself or a nearby gene with type 1 diabetes susceptibility in Germans, confirming respective observations previously made in Indian Asians. PMID- 10868976 TI - Physical and genetic mapping of IDDM8 on chromosome 6q27. AB - Genome-wide mapping studies have provided evidence of a type 1 diabetes susceptibility gene (IDDM8) that is located on chromosome 6q27. However, association studies of IDDM8 have so far been negative. The purpose of this investigation was to determine a linkage disequilibrium (LD) map in the chromosome 6q27 region and to better localize IDDM8. A physical map of nearly 1 Mb containing the chromosome 6 telomere was constructed, and polymorphic markers spanning this region were defined. Haplotypes composed of the markers in LD were tested for association with type 1 diabetes in 266 families. A microsatellite marker allele and multiple haplotypes were associated with IDDM8, which suggests localization of this type 1 diabetes susceptibility gene to the terminal 200 kb of chromosome 6. PMID- 10868977 TI - Linkage of serum insulin concentrations to chromosome 3p in Mexican Americans. AB - Hyperinsulinemia predicts the development of type 2 diabetes, and family studies suggest that insulin levels are regulated in part by genes. We conducted a genome wide scan to detect genes influencing variation in fasting serum insulin concentrations in 391 nondiabetic individuals from 10 large multigenerational families. Approximately 380 microsatellite markers with an average spacing of 10 cM were genotyped in all study subjects. Insulin concentrations measured by radioimmunoassay were transformed by their natural logarithms before analysis. In multipoint analysis, peak evidence for linkage occurred on chromosome 3p approximately 109 cM from pter in the region of 3p14.2-p14.1. The multipoint logarithm of odds (LOD) score was 3.07, occurring in the region flanked by markers D3S1600 and D3S1285 (P value by simulation <0.0001). In a two-point analysis, LOD scores ranged from 0.75 to 2.52 for the nine markers typed in the region spanning 88-143 cM from pter. The fasting insulin resistance index was highly correlated with fasting insulin concentrations in this sample and also provided strong evidence for linkage to this region (LOD = 2.99). There was no evidence in our genome-wide scan for linkage of insulin levels to any other chromosome. These results provide evidence that a gene-influencing variation in insulin concentrations exists on chromosome 3p. Possible candidate genes in this region include GBE1 and ACOX2, which encode enzymes involved in glycogen and fatty acid metabolism, respectively. PMID- 10868978 TI - Interleukin-6 gene polymorphism and insulin sensitivity. AB - Type 2 diabetes and the insulin resistance syndrome have been hypothesized to constitute manifestations of an ongoing acute-phase response. We aimed to study an interleukin-6 (IL-6) gene polymorphism in relation to insulin sensitivity (IL 6 is the main cytokine involved in an acute-phase response). Subjects homozygous for the C allele at position -174 of the IL-6 gene (SfaNI genotype), associated to lower plasma IL-6 levels, showed significantly lower integrated area under the curve of serum glucose concentrations (AUCglucose) after an oral glucose tolerance test, lower blood glycosylated hemoglobin, lower fasting insulin levels, lower total and differential white blood cell count (a putative marker of peripheral IL-6 action), and an increased insulin sensitivity index than carriers of the G allele, despite similar age and body composition. A gene dosage effect was especially remarkable for AUCglucose (6.4 vs. 9.3 vs. 9.7 mmol/l in C/C, C/G, and G/G individuals, respectively). The serum concentration of fully glycosylated cortisol binding globulin (another marker of IL-6 action), suggested by concanavalin A adsorption, was lower in C/C subjects than in G/G individuals (32.6+/-2.9 vs. 37.6+/-4.6 mg/l, P = 0.03). In summary, a polymorphism of the IL 6 gene influences the relationship among insulin sensitivity, postload glucose levels, and peripheral white blood cell count. PMID- 10868979 TI - A PC-1 amino acid variant (K121Q) is associated with faster progression of renal disease in patients with type 1 diabetes and albuminuria. AB - Insulin resistance characterizes type 1 diabetes in patients with albuminuria. A PC-1 glycoprotein amino acid variant, K121Q, is associated with insulin resistance. We examined the impact of the PC-1 K121Q variant on the rate of decline of the glomerular filtration rate (GFR) by creatinine clearance derived from the Cockroft-Gault formula in 77 type 1 diabetic patients with albuminuria who were followed for an average of 6.5 years (range 2.5-15). Patients carrying the Q allele (n = 22; 20 with KQ and 2 with QQ genotypes) had a faster GFR decline than those patients with the KK genotype (n = 55) (median 7.2 vs. 3.7 ml x min(-1) x year(-1); range 0.16 to 16.6 vs. -3.8 to 16.0 ml x min(-1) x year( 1); P < 0.001). Significantly more patients carrying the Q allele belonged to the highest tertile of GFR decline (odds ratio = 5.7, 95% CI 4.1-7.2, P = 0.02). Levels of blood pressure, HbA1c, and albuminuria were comparable in the two genotype groups. Albuminuria (P = 0.001), mean blood pressure (P = 0.046), and PC 1 genotype (P = 0.036) independently correlated with GFR decline. Because all patients were receiving antihypertensive treatment, the faster GFR decline in the patients carrying the Q allele could be the result of reduced sensitivity to the renoprotective effect of antihypertensive therapy. PC-1 genotyping identifies type 1 diabetic patients with a faster progression of diabetic nephropathy. PMID- 10868980 TI - Family distress and involvement in relatives of obsessive-compulsive disorder patients. AB - Obsessive compulsive disorder (OCD) affects the lives of patients as well as their relatives. Calvocoressi et al. (1995) suggested that accommodation (e.g., participating in the patient's rituals) by relatives of patients with OCD was related to global family dysfunction and distress. These investigators did not, however, examine the relationship between family accommodation and severity of the patients' OCD symptoms. In the present study we examined the relationship between families' reactions to the patient's illness (e.g., assistance with rituals, modification of family routine, rejection of the patient, etc.) and the patient's Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) score and Hamilton Depression scale ratings. We also examined the effects of family accommodation and rejection on treatment outcome. Results revealed that family distress, accommodation, and rejection were related to depression and anxiety in the family members, but not to severity of the patients' OCD. At posttreatment, family accommodation and modification of routine was related to the severity of patients' OCD. These results remained significant when the effect of pretreatment OCD severity was partialled out. PMID- 10868981 TI - Emotional processing in combat-related posttraumatic stress disorder: a comparison with traumatized and normal controls. AB - Emotional numbing (EN) symptoms are an important but poorly understood component of the response to trauma. To try to demonstrate EN, this laboratory study examined subjective and psychophysiological emotion responses to standardized visual stimuli in combat veterans with posttraumatic stress disorder (PTSD), combat veterans without PTSD, and nontraumatized controls. PTSD subjects showed no evidence of generalized reduction in subjective or psychophysiological emotion responses. In response to a subset of more evocative stimuli, PTSD subjects reported less experience of Positive Emotions, and more experience of Negative Emotions than controls. For controls, valence and arousal were uncorrelated, while they were negatively correlated for PTSD subjects. Verbal and nonverbal subjective emotion measures were positively correlated for all subject groups, but there was little correlation between subjective emotion measures and psychophysiological indices. Viewing time was positively correlated with Positive Emotions for PTSD subjects, and with Negative Emotions for combat controls. PMID- 10868982 TI - Neurocognitive correlates of anxiety disorders in children: a preliminary report. AB - Children with anxiety disorders have been suggested to possess a specific cognitive scheme that underscores negative information and leads to the formation of a negative view of themselves and of the world. The aim of the present study was to assess the neuropsychological processes of children and adolescents with anxiety disorders, as compared to healthy matched controls. Nineteen children (6 18 years) with anxiety disorders and 14 age-matched healthy controls participated in the study. Both groups scored within normal range on the Wechsler Intelligence Scale for Children-Revised (WISC-R). All children underwent neuropsychological assessment with the California Verbal Learning Test (CVLT) (Verbal Processing), the Rey-Osterrieth Complex Figure test (ROCF) (Nonverbal Processing), and the Wisconsin Card Sorting Test (WCST) (Executive Functions). The anxiety group scored lower than the control group on all measures of the CVLT and had a significantly greater number of errors, perseverative responses, and incorrect answers after negative feedback on the WCST. No differences were detected for the ROCF. We conclude that in children and adolescents, anxiety disorders may be associated with lowered linguistic abilities and cognitive flexibility, as measured by neuropsychological paradigms. Anxiety does not appear to be associated with nonverbal processes. PMID- 10868983 TI - Deliberate emotional expressions of socially anxious children and their mothers. AB - The aim of the present study was to investigate whether socially anxious children show deficits in their deliberate facial expression of emotions. To test for potential mother-child transmission effects, the mothers' facial expressions were also assessed. Fifty socially anxious and 25 socially nonanxious children (8-12 years) and their mothers participated in a facial expression posing task. The expressions produced were coded using Ekman and Friesen's (1978) Facial Action Coding System (FACS). In addition, naive raters rated their quality of emotion. FACS analyses indicated that socially anxious children show a reduced general facial activity, have a more restricted facial repertoire and differ qualitatively from controls in their facial expression of emotions. Similarly, the global ratings indicated that the socially anxious children's posed facial expressions are less accurate. For the mothers no differences between groups were found when global ratings were used. However, the FACS data demonstrate that the facial expressions of mothers of socially anxious children are less intense compared to controls. It is possible that the decreased intensity of the mothers' facial expressiveness makes it more difficult for the socially anxious children to learn adequate facial expressions. PMID- 10868984 TI - Frequency, comorbidity, and psychosocial impairment of anxiety disorders in German adolescents. AB - The frequency, comorbidity, and psychosocial impairment of anxiety disorders among German adolescents was estimated from a survey of 1,035 students aged 12-17 years. The adolescents were randomly selected from 36 schools in the province of Bremen, Germany. Anxiety disorders and other psychiatric disorders were coded based on criteria from the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders, using the computerized Munich version of the Composite International Diagnostic Interview. Anxiety disorders occurred frequently in our sample of adolescents, with a rate of 18.6%. When considering the subtypes of anxiety disorders, phobia was the most common. Posttraumatic stress disorder and obsessive-compulsive disorder occurred less frequently with rates below 2%. Panic disorder and generalized anxiety disorder were the least common, with rates well below 1%. Anxiety disorders were significantly higher in girls than in boys, and that the rates increased with age. Comorbidity occurs quite frequently, both within the anxiety disorders and also with other psychiatric disorders. The most common pattern of comorbidity was that of anxiety and depressive disorders. Although a high number of anxiety cases were psychosocially impaired, at least during the worst episode of their disorders, only a few of them sought treatment for their problems. We conclude by discussing some research priorities in the area of anxiety disorders in children and adolescents. PMID- 10868985 TI - Comparison and characteristics of motor vehicle accident (MVA) and non-MVA driving fears. AB - Prior research has revealed the diagnostic complexity among people who report driving fears. However, the focus on survivors of motor vehicle accidents (MVAs) and diagnostic samples may have inadvertently led to a relative neglect of the broader driving-fearful population. No studies could be located that compared MVA survivors with those who had not experienced an MVA. The aim of the present study was to address these deficits by comparing the characteristics of MVA and non-MVA driving-fearfuls and also exploring a range of characteristics associated with driving fears. One hundred and ninety media-recruited driving-fearfuls completed a questionnaire that assessed severity of anxiety and avoidance associated with a variety of driving situations. It was found that fear levels were similar to samples of driving phobics and MVA victims. There were no significant differences between MVA and non-MVA respondents on various measures of fear severity. In addition, the sample rated a high level of anxiety when driving with someone who criticizes their driving. Implications of the findings are discussed, along with suggestions for assessment and treatment of those with driving-related fears. PMID- 10868986 TI - Psychiatric diagnoses among self-referred dental injection phobics. AB - In order to determine the psychiatric characteristics of people with dental injection phobia. 118 dental injection phobics were systematically assessed using a structured clinical interview and a written questionnaire. Fifty-four percent of subjects had a current Axis I diagnosis other than dental injection phobia, mainly anxiety, mood or adjustment disorder, and 68.6% of subjects had an additional lifetime Axis I diagnosis. Subjects with additional current Axis I diagnoses reported higher dental anxiety, greater severity of injection fear cognitions, and poorer relationships with dental professionals, than did subjects without any or with past Axis I diagnoses. Further investigation is needed to explore the treatment possibilities for patients with and without additional current diagnoses. PMID- 10868987 TI - The effect of prior trauma exposure on the development of PTSD following spinal cord injury. AB - Posttraumatic stress disorder (PTSD) occurs in only a subset of individuals who sustain traumatic spinal cord injuries (SCIs). Several previous studies have examined the effects of additive trauma on the development of PTSD and found that a history of prior trauma increases the risk for later development of PTSD. The present study examines additive trauma by investigating the effects of previous combat exposure on the development of PTSD following spinal cord injury. Significant differences in prevalence rates for current PTSD were found for the comparisons of war theater (both combat and noncombat) versus non-war theater veterans but not for the comparison between combat and noncombat war theater veterans. Moreover, for all the comparisons, no significant differences were found in lifetime PTSD diagnoses. This implies that veterans with SCI who served in a war zone have increased difficulty recovering from their PTSD following a spinal cord injury than do non-war theater veterans. PMID- 10868988 TI - Leptin and puberty. PMID- 10868989 TI - Mental health must be "centre stage" in child welfare. PMID- 10868990 TI - Taking a population perspective on child health. PMID- 10868991 TI - The principles of management of congenital anomalies of the upper limb. AB - Management of congenital anomalies of the upper limb is reviewed with reference to classification and aetiology, incidence, diagnosis before birth, broad principles of treatment, timing of x rays and scans, functional aims, cosmetic appearance, counselling of parents, therapists, scars, skin grafts, growth, and timing of surgery. Notes on 11 congenital hand conditions are given. PMID- 10868992 TI - Stamps in paediatrics. Child welfare stamps. PMID- 10868994 TI - Better burns survival PMID- 10868993 TI - US adolescent food intake trends from 1965 to 1996. AB - AIM: To examine adolescent food consumption trends in the United States with important chronic disease implications. METHODS: Analysis of dietary intake data from four nationally representative United States Department of Agriculture surveys of individuals 11-18 years of age (n = 12 498). RESULTS: From 1965 to 1996, a considerable shift in the adolescent diet occurred. Total energy intake decreased as did the proportion of energy from total fat (39% to 32%) and saturated fat (15% to 12%). Concurrent increases occurred in the consumption of higher fat potatoes and mixed dishes (pizza, macaroni cheese). Lower fat milks replaced higher fat milks but total milk consumption decreased by 36%. This decrease was accompanied by an increase in consumption of soft drinks and non citrus juices. An increase in high fat potato consumption led to an increase in vegetable intake but the number of servings for fruits and vegetables is still below the recommended five per day. Iron, folate, and calcium intakes continue to be below recommendations for girls. CONCLUSIONS: These trends, far greater than for US adults, may compromise health of the future US population. PMID- 10868995 TI - Cognitive development of term small for gestational age children at five years of age. AB - AIM: To assess the relative significance for cognitive development of small for gestational age, parental demographic factors, and factors related to the child rearing environment. METHODS: IQ of a population based cohort of 338 term infants who were small for gestational age (SGA) and without major handicap, and a random control sample of 335 appropriate for gestational age (AGA) infants were compared at 5 years of age. RESULTS: The mean non-verbal IQ was four points lower, while the mean verbal IQ was three points lower for the children in the SGA group. The results were not confounded by parental demographic or child rearing factors. However, parental factors, including maternal non-verbal problem solving abilities, and child rearing style, accounted for 20% of the variance in non verbal IQ, while SGA versus AGA status accounted for only 2%. The comparable numbers for verbal IQ were 30 and 1%. Furthermore, we found no evidence that the cognitive development of SGA children was more sensitive to a non-optimal child rearing environment than that of AGA children. Maternal smoking at conception was associated with a reduction in mean IQ comparable to that found for SGA status, and this effect was the same for SGA and AGA children. The cognitive function of asymmetric SGA was comparable to that of symmetric SGA children. CONCLUSIONS: Our findings indicate that child cognitive development is strongly associated with parental factors, but only marginally associated with intrauterine growth retardation. PMID- 10868997 TI - Tertiary paediatrics needs a disability model. PMID- 10868996 TI - Recent advances in the genetics of severe childhood obesity. AB - Childhood obesity is becoming a global epidemic. Twin studies suggest a heritability of fat mass, and disorders of energy balance that arise from genetic defects have been identified. In the past three years, five single gene disorders resulting in early onset obesity have been characterised. The discovery of these genetic defects has biological and clinical implications which are greater than the rarity of the individual diseases might suggest. PMID- 10868998 TI - Clinical training experience in district general hospitals. AB - AIMS: To estimate the nature and quantity of clinical experience available for trainees in paediatrics or general practice in acute general hospitals of differing sizes in the UK. To discuss implications for training and service configuration taking account of current Royal College recommendations (a minimum of 1,800 acute contacts each year and ideally covering a population of 450,000 to 500,000 people). METHODS: Observed frequencies of diagnoses in Pinderfields Hospital, Wakefield were compared with those in five other hospitals in Yorkshire and four in the South of England, and with expected frequencies from a review of selected marker conditions using national routine and epidemiological data. Based on the Pinderfields data, we modelled expected frequencies of a wider range of diagnoses for different sized hospitals. RESULTS: Small units (1,800 or less acute referrals a year) provide adequate exposure to common conditions such as gastroenteritis (157 per annum) and asthma (171 per annum) but encounter serious or unusual disease rarely. When modelled for units serving larger populations, numbers of such disorders remain small. For example, about 0.5% of admissions require intensive care to the level of ventilatory support. Medium size units offer a wide range of experience but differ little from those serving the population of 500,000 proposed as being optimal for training. This standard is not justified by the evidence in this review. Closing or amalgamating units on the scale necessary to achieve this ideal would be impractical as only five hospitals in England have a paediatric workload equivalent to this population; it would also raise issues of access and equity. PMID- 10868999 TI - Corticosteroid therapy in nephrotic syndrome: a meta-analysis of randomised controlled trials. AB - AIMS: To determine the benefits and toxicity of different corticosteroid regimes in preventing relapse in steroid responsive nephrotic syndrome. DESIGN: Meta analysis of randomised controlled trials. SUBJECTS: Twelve trials involving 868 children aged 3 months to 18 years. MAIN OUTCOME MEASURE: Frequency of relapse. RESULTS: A meta-analysis of five trials, which compared two months of prednisone with three months or more in the first episode, showed that the longer duration significantly reduced the risk of relapse at 12-24 months (relative risk 0.73; 95% confidence interval 0.60 to 0.89) without an increase in adverse events. There was an inverse linear relation (relative risk 1.382 (SE 0.215) - 0.133 (SE 0.048) duration; r(2) = 0.66; p = 0.05) between the duration of treatment and risk of relapse. CONCLUSIONS: Children in their first episode of steroid responsive nephrotic syndrome should be treated with prednisone for at least three months, with an increase in benefit being shown for up to seven months of treatment. PMID- 10869000 TI - Megarectum in constipation. AB - BACKGROUND: Faecal impaction is frequently observed in children with chronic constipation. The term megarectum is often used to describe this finding. AIM: To evaluate rectal functioning and rectal measures in constipated children with a filled rectum, in order to define the terms faecal impaction, enlarged rectum, and megarectum. METHODS: All children underwent radiological investigation, colonic transit time study, anorectal manometry, and rectal volume and rectal wall compliance measurements. Patients with faecal impaction were compared with controls, who had an empty rectum on digital rectal examination. RESULTS: A total of 31 patients and six controls were included in the study. The mean duration of complaints was 4.2 years and all had faecal incontinence. The colonic transit times in the patients showed a distinct delay in the rectosigmoid segment. Anorectal manometry was not significantly different between patients and controls. The rectal width in patients was 0.68 and in controls 0.52 with an upper limit of 0.61. The pressure-volume curve in patients showed significant less relaxation at a distension of 50 ml. The slope of the curve (corresponding with rectal wall compliance) was comparable for patients and controls. CONCLUSIONS: We suggest that faecal impaction is a filled rectum found on digital rectal examination; an enlarged rectum is defined by a rectopelvic ratio greater than 0.61; and megarectum is defined in those with significant abnormalities found with anorectal manometry, pressure-volume curves, or rectal compliance investigation. A diminished relaxation of the rectum on rectal distension could be the first sign of megarectum in children with chronic constipation. PMID- 10869002 TI - ADHD: A new practice guideline from the American Academy of Pediatrics. Attention deficit hyperactive disorder. PMID- 10869001 TI - Dilated cardiomyopathy in dystrophic epidermolysis bullosa. AB - BACKGROUND: Dystrophic epidermolysis bullosa (DEB) is an uncommon genetic disorder of the skin and mucosae. In 1996, we reported the occurrence of lethal dilated cardiomyopathy (DCM) in two affected children. METHODS: In the past seven years we have routinely screened patients with severe DEB who have been under the care of this hospital by yearly clinical review, echocardiography, and quantification of plasma selenium and carnitine concentrations, as deficiency of these micronutrients is known to be associated with the development of DCM. RESULTS: Six of 61 children have developed DCM over the seven year period of this study, four of whom have not been previously reported, and three of whom have since died. We compared the concentrations of selenium and free and total carnitine in the children who developed DCM to concentrations in those with severe DEB who did not. The concentrations of free and total carnitine when first measured were significantly lower in the children with DCM, but the selenium concentrations were not. CONCLUSIONS: We now believe that DCM is a not infrequent complication of severe recessive DEB, and may be related in part to carnitine concentrations, though the exact mechanism remains unclear. We therefore recommend that patients with this condition should undergo regular cardiac review including echocardiography. PMID- 10869003 TI - The impact of diagnostic delay on the course of acute appendicitis. AB - BACKGROUND: The diagnosis of acute appendicitis is often delayed, which may complicate the further course of the disease. AIMS: To review appendectomy cases in order to determine the incidence of diagnostic delay, the underlying factors, and impact on the course of the disease. METHODS: Records of all children who underwent appendectomy from 1994 to 1997 were reviewed. The 129 cases were divided into group A (diagnostic period within 48 hours) and group B (diagnostic period 48 hours or more). RESULTS: In the group with diagnostic delay, significantly more children had first been referred to a paediatrician rather than to a surgeon. In almost half of the cases in this group initial diagnosis was not appendicitis but gastroenteritis. The perforation rate in group A was 24%, and in group B, 71%. Children under 5 years of age all presented in the delayed group B and had a perforation rate of 82%. The delayed group showed a higher number of postoperative complications and a longer hospitalisation period. CONCLUSIONS: Appendicitis is hard to diagnose when, because of a progressing disease process, the classical clinical picture is absent. The major factor in diagnostic delay is suspected gastroenteritis. Early surgical consultation in a child with deteriorating gastroenteritis is advised. Ultrasonographs can be of major help if abdominal signs and symptoms are non-specific for appendicitis. PMID- 10869004 TI - Primarily chronic and cerebrovascular course of Lyme neuroborreliosis: case reports and literature review. AB - As part of an ongoing study aiming to define the clinical spectrum of neuroborreliosis in childhood, we have identified four patients with unusual clinical manifestations. Two patients suffered from a primarily chronic form of neuroborreliosis and displayed only non-specific symptoms. An 11 year old boy presented with long standing symptoms of severe weight loss and chronic headache, while the other patient had pre-existing mental and motor retardation and developed seizures and failure to thrive. Two further children who presented with acute hemiparesis as a result of cerebral ischaemic infarction had a cerebrovascular course of neuroborreliosis. One was a 15 year old girl; the other, a 5 year old boy, is to our knowledge the youngest patient described with this course of illness. Following adequate antibiotic treatment, all patients showed substantial improvement of their respective symptoms. Laboratory and magnetic resonance imaging findings as well as clinical course are discussed and the relevant literature is reviewed. PMID- 10869005 TI - Hereditary fructose intolerance and alpha(1) antitrypsin deficiency. AB - A patient with coexisting hereditary fructose intolerance (HFI) and alpha(1) antitrypsin deficiency (alpha(1)ATD) is described. Protease inhibitor typing was not conclusive, presumably because of impaired N-glycosylation secondary to HFI. The case underlines the diagnostic role of molecular genetic techniques in inborn errors of metabolism. PMID- 10869007 TI - Recurrent pneumonia PMID- 10869006 TI - Early onset of Friedreich's ataxia in a compound heterozygote. AB - Friedreich's ataxia (FA) is an autosomal recessive condition caused by a GAA trinucleotide repeat expansion in the X25 gene on chromosome 9. We describe an unusual form of "pseudodominant" inheritance to illustrate how a diagnosis of FA in a parent does not preclude the diagnosis in the child. PMID- 10869008 TI - Are sleep studies worth doing? AB - AIMS: To evaluate a sleep study service for children suspected of having sleep related upper airway obstruction (SRUAO). DESIGN: Prospective survey. SETTING: Paediatric and ear, nose, and throat clinics of the Royal Free Hampstead NHS Trust. SUBJECTS: Consecutively referred children with SRUAO symptoms. MAIN OUTCOME MEASURES: Sleep study data, referring clinician's impression, and completed symptom questionnaires. RESULTS: A total of 120 children (aged 6 months to 15.5 years) were studied. Study scores showed that 24 were classified as normal, 42 as mild, 33 as moderate, and 21 as severe SRUAO. In the 106 cases with matching data between clinician's impression and study score, 71 had good agreement, 18 were underestimated by the clinician, and 17 were over estimated. No cases reported as moderate or severe sleep apnoea by the study were referred by the clinician as normal. There were no important associations between parental symptom scores and sleep study scores. CONCLUSION: In children with suspected SRUAO, sleep studies do contribute to assessing the need for operation, the likelihood of postoperative respiratory failure, or as a baseline or outcome measure in intervention studies. PMID- 10869009 TI - FETAL AND NEONATAL EDITION july 2000 issue PMID- 10869011 TI - Two or three myths about bioinformatics. PMID- 10869010 TI - PACS (picture archiving and communication systems): filmless radiology. AB - A picture archiving and communication system (PACS) is a computerised means of replacing the roles of conventional radiological film. This review describes the Hammersmith PACS, and discusses the advantages and disadvantages of PACS systems. PMID- 10869012 TI - Homology-based gene structure prediction: simplified matching algorithm using a translated codon (tron) and improved accuracy by allowing for long gaps. AB - MOTIVATION: Locating protein-coding exons (CDSs) on a eukaryotic genomic DNA sequence is the initial and an essential step in predicting the functions of the genes embedded in that part of the genome. Accurate prediction of CDSs may be achieved by directly matching the DNA sequence with a known protein sequence or profile of a homologous family member(s). RESULTS: A new convention for encoding a DNA sequence into a series of 23 possible letters (translated codon or tron code) was devised to improve this type of analysis. Using this convention, a dynamic programming algorithm was developed to align a DNA sequence and a protein sequence or profile so that the spliced and translated sequence optimally matches the reference the same as the standard protein sequence alignment allowing for long gaps. The objective function also takes account of frameshift errors, coding potentials, and translational initiation, termination and splicing signals. This method was tested on Caenorhabditis elegans genes of known structures. The accuracy of prediction measured in terms of a correlation coefficient (CC) was about 95% at the nucleotide level for the 288 genes tested, and 97. 0% for the 170 genes whose product and closest homologue share more than 30% identical amino acids. We also propose a strategy to improve the accuracy of prediction for a set of paralogous genes by means of iterative gene prediction and reconstruction of the reference profile derived from the predicted sequences. AVAILABILITY: The source codes for the program 'aln' written in ANSI-C and the test data will be available via anonymous FTP at ftp.genome.ad.jp/pub/genomenet/saitama-cc. CONTACT: gotoh@cancer-c.pref.saitama.jp PMID- 10869013 TI - Optimal spliced alignment of homologous cDNA to a genomic DNA template. AB - MOTIVATION: Supplementary cDNA or EST evidence is often decisive for discriminating between alternative gene predictions derived from computational sequence inspection by any of a number of requisite programs. Without additional experimental effort, this approach must rely on the occurrence of cognate ESTs for the gene under consideration in available, generally incomplete, EST collections for the given species. In some cases, particular exon assignments can be supported by sequence matching even if the cDNA or EST is produced from non cognate genomic DNA, including different loci of a gene family or homologous loci from different species. However, marginally significant sequence matching alone can also be misleading. We sought to develop an algorithm that would simultaneously score for predicted intrinsic splice site strength and sequence matching between the genomic DNA template and a related cDNA or EST. In this case, weakly predicted splice sites may be chosen for the optimal scoring spliced alignment on the basis of surrounding sequence matching. Strongly predicted splice sites will enter the optimal spliced alignment even without strong sequence matching. RESULTS: We designed a novel algorithm that produces the optimal spliced alignment of a genomic DNA with a cDNA or EST based on scoring for both sequence matching and intrinsic splice site strength. By example, we demonstrate that this combined approach appears to improve gene prediction accuracy compared with current methods that rely only on either search by content and signal or on sequence similarity. AVAILABILITY: The algorithm is available as a C subroutine and is implemented in the SplicePredictor and GeneSeqer programs. The source code is available via anonymous ftp from ftp. zmdb.iastate.edu. Both programs are also implemented as a Web service at http://gremlin1.zool.iastate.edu/cgi-bin/s p.cgiand http://gremlin1.zool.iastate.edu/cgi-bin/g s.cgi, respectively. CONTACT: vbrendel@iastate.edu PMID- 10869014 TI - Net nearest neighbor analysis (NNNA) summarizes non-compensated dinucleotides within gene sequences. AB - MOTIVATION: Net Nearest Neighbor Analysis (NNNA) measures a previously unexamined aspect of dinucleotide frequency-the non-compensated, non-repetitive dinucleotides in a sequence. Non-compensated dinucleotides are those in excess of their corresponding reverse dinucleotides. RESULTS: NNNA regards dinucleotides as vector quantities, making it possible to summarize any sequence as a group of circuits and tags. The results of NNNA are found to be consistent with traditional analytic methods, yet reveal additional characteristics of the sequences. The NNNA circuits and tags uniquely identify each tRNA in Escherichia coli K-12 and certain structural components of each tRNA, extract function specific characteristics for each of the sequences involved in the formation of insulin from preinsulin, and exhibit species-specific phylogenetic characterization (demonstrated with MONILINIA:). AVAILABILITY: Nearest neighbor analysis software has been available for many years and is a component of most gene analysis software packages, including the Staden Package which is available at no charge to academic users (http://www.mrc-1mb.cam.ac. uk/pubseq/). PMID- 10869015 TI - Combinatorial motif analysis and hypothesis generation on a genomic scale. AB - MOTIVATION: Computer-assisted methods are essential for the analysis of biosequences. Gene activity is regulated in part by the binding of regulatory molecules (transcription factors) to combinations of short motifs. The goal of our analysis is the development of algorithms to identify regulatory motifs and to predict the activity of combinations of those motifs. APPROACH: Our research begins with a new motif-finding method, using multiple objective functions and an improved stochastic iterative sampling strategy. Combinatorial motif analysis is accomplished by constructive induction that analyzes potential motif combinations. The hypothesis is generated by applying standard inductive learning algorithms. RESULTS: Tests using 10 previously identified regulons from budding yeast and 14 artificial families of sequences demonstrated the effectiveness of the new motif-finding method. Motif combination and classification approaches were used in the analysis of a sample DNA array data set derived from genome-wide gene expression analysis. AVAILABILITY: Programs will be available as executable files upon request. CONTACT: yhu@ics.uci.eduor yhu@cse.ttu.edu.tw PMID- 10869016 TI - Fast probabilistic analysis of sequence function using scoring matrices. AB - MOTIVATION: We present techniques for increasing the speed of sequence analysis using scoring matrices. Our techniques are based on calculating, for a given scoring matrix, the quantile function, which assigns a probability, or p, value to each segmental score. Our techniques also permit the user to specify a p threshold to indicate the desired trade-off between sensitivity and speed for a particular sequence analysis. The resulting increase in speed should allow scoring matrices to be used more widely in large-scale sequencing and annotation projects. RESULTS: We develop three techniques for increasing the speed of sequence analysis: probability filtering, lookahead scoring, and permuted lookahead scoring. In probability filtering, we compute the score threshold that corresponds to the user-specified p threshold. We use the score threshold to limit the number of segments that are retained in the search process. In lookahead scoring, we test intermediate scores to determine whether they will possibly exceed the score threshold. In permuted lookahead scoring, we score each segment in a particular order designed to maximize the likelihood of early termination. Our two lookahead scoring techniques reduce substantially the number of residues that must be examined. The fraction of residues examined ranges from 62 to 6%, depending on the p threshold chosen by the user. These techniques permit sequence analysis with scoring matrices at speeds that are several times faster than existing programs. On a database of 12 177 alignment blocks, our techniques permit sequence analysis at a speed of 225 residues/s for a p threshold of 10-6, and 541 residues/s for a p threshold of 10-20. In order to compute the quantile function, we may use either an independence assumption or a Markov assumption. We measure the effect of first- and second-order Markov assumptions and find that they tend to raise the p value of segments, when compared with the independence assumption, by average ratios of 1.30 and 1.69, respectively. We also compare our technique with the empirical 99. 5th percentile scores compiled in the BLOCKSPLUS database, and find that they correspond on average to a p value of 1.5 x 10-5. AVAILABILITY: The techniques described above are implemented in a software package called EMATRIX. This package is available from the authors for free academic use or for licensed commercial use. The EMATRIX set of programs is also available on the Internet at http://motif.stanford.edu/ematrix. PMID- 10869017 TI - Adaptive encoding neural networks for the recognition of human signal peptide cleavage sites. AB - MOTIVATION: Data representation and encoding are essential for classification of protein sequences with artificial neural networks (ANN). Biophysical properties are appropriate for low dimensional encoding of protein sequence data. However, in general there is no a priori knowledge of the relevant properties for extraction of representative features. RESULTS: An adaptive encoding artificial neural network (ACN) for recognition of sequence patterns is described. In this approach parameters for sequence encoding are optimized within the same process as the weight vectors by an evolutionary algorithm. The method is applied to the prediction of signal peptide cleavage sites in human secretory proteins and compared with an established predictor for signal peptides. CONCLUSION: Knowledge of physico-chemical properties is not necessary for training an ACN. The advantage is a low dimensional data representation leading to computational efficiency, easy evaluation of the detected features, and high prediction accuracy. AVAILABILITY: A cleavage site prediction server is located at the Humboldt University http://itb.biologie.hu-berlin.de/ approximately jo/sig cleave/ACNpredictor.cgi CONTACT: jo@itb.hu-berlin.de; berndj@zedat.fu-berlin.de PMID- 10869018 TI - Predicting the oxidation state of cysteines by multiple sequence alignment. AB - MOTIVATION: Protein sequences found in databanks usually do not report post translational covalent modifications such as the oxidation state of cystein (Cys) residues. Accurate prediction of whether a functionally or structurally important Cys occurs in the oxidized or thiol form would be helpful for molecular biology experiments and structure prediction. RESULTS: A new method is presented for predicting the oxidation state of Cys residues based on multiple sequence alignments and on the observation that Cys tends to occur in the same oxidation state within the same protein. The prediction of the redox state of Cys performs above 82%. The oxidation state of Cys correlates with the cellular location of the given protein within the cell, but the correlation is not perfect (up to 70%). We also perform a statistical analysis of the different redox states of Cys found in secondary structures and buried positions, and of the secondary structures linked by disulfide bonds. The results suggest that the natural borderline lies between the different oxidation states of Cys rather than between the half cystines and cysteins. AVAILABILITY: A web server implementing the prediction method is available at http://guitar.rockefeller.edu/approximately andras/cyspred.html CONTACT: fisera@rockefeller.edu PMID- 10869020 TI - An ontology for biological function based on molecular interactions. AB - MOTIVATIONS: A number of important bioinformatics computations involve computing with function: executing computational operations whose inputs or outputs are descriptions of the functions of biomolecules. Examples include performing functional queries to sequence and pathway databases, and determining functional equality to evaluate algorithms that predict function from sequence. A prerequisite to computing with function is the existence of an ontology that provides a structured semantic encoding of function. Functional bioinformatics is an emerging subfield of bioinformatics that is concerned with developing ontologies and algorithms for computing with biological function. RESULTS: The article explores the notion of computing with function, and explains the importance of ontologies of function to bioinformatics. The functional ontology developed for the EcoCyc database is presented. This ontology can encode a diverse array of biochemical processes, including enzymatic reactions involving small-molecule substrates and macromolecular substrates, signal-transduction processes, transport events, and mechanisms of regulation of gene expression. The ontology is validated through its use to express complex functional queries for the EcoCyc DB. CONTACT: pkarp@ai.sri.com PMID- 10869019 TI - Sequence-structure specificity of a knowledge based energy function at the secondary structure level. AB - MOTIVATION: This paper investigates the sequence-structure specificity of a representative knowledge based energy function by applying it to threading at the level of secondary structures of proteins. Assessing the strengths and weaknesses of an energy function at this fundamental level provides more detailed and insightful information than at the tertiary structure level and the results obtained can be useful in tertiary level threading. RESULTS: We threaded each of the 293 non-redundant proteins onto the secondary structures contained in its respective native protein (host template). We also used 68 pairs of proteins with similar folds and low sequence identity. For each pair, we threaded the sequence of one protein onto the secondary structures of the other protein. The discerning power of the total energy function and its one-body, pairwise, and mutation components is studied. We then applied our energy function to a recent study which demonstrated how a designed 11-amino acid sequence can replace distinct segments (one segment is an alpha-helix, the other is a beta-sheet) of a protein without changing its fold. We conducted random mutations of the designed sequence to determine the patterns for favorable mutations. We also studied the sequence structure specificity at the boundaries of a secondary structure. Finally, we demonstrated how to speed up tertiary level threading by filtering out alignments found to be energetically unfavorable during the secondary structure threading. AVAILABILITY: The program is available on request from the authors. CONTACT: xud@ornl.gov PMID- 10869021 TI - MPSA: integrated system for multiple protein sequence analysis with client/server capabilities. AB - MPSA is a stand-alone software intended to protein sequence analysis with a high integration level and Web clients/server capabilities. It provides many methods and tools, which are integrated into an interactive graphical user interface. It is available for most Unix/Linux and non-Unix systems. MPSA is able to connect to a Web server (e.g. http://pbil.ibcp.fr/NPSA) in order to perform large-scale sequence comparison on up-to-date databanks. AVAILABILITY: Free to academic http://www.ibcp.fr/mpsa/ CONTACT: c.blanchet@ibcp.fr PMID- 10869022 TI - Storing biological sequence databases in relational form. AB - SUMMARY: We have created a set of applications using Perl and Java in combination with XML technology to install biological sequence databases into an Oracle RDBMS. An easy-to-use interface using Java has been created for database query and other tools developed to integrate with our in-house bioinformatics applications. AVAILIBILITY: The database schema, DTD file, and source codes are available from the authors via email. CONTACT: guochun_ xie@merck. com PMID- 10869023 TI - PIR: a new resource for bioinformatics. AB - The Protein Information Resource (PIR) has greatly expanded its Web site and developed a set of interactive search and analysis tools to facilitate the analysis, annotation, and functional identification of proteins. New search engines have been implemented to combine sequence similarity search results with database annotation information. The new PIR search systems have proved very useful in providing enriched functional annotation of protein sequences, determining protein superfamily-domain relationships, and detecting annotation errors in genomic database archives. AVAILABILITY: http://pir.georgetown.edu/. CONTACT: mcgarvey@nbrf.georgetown.edu PMID- 10869024 TI - HELM: searching for helix motifs within protein sequences. AB - SUMMARY: HELM is a web tool designed to automate the analysis of protein sequences searching for alpha helix motifs. This analysis can be useful in protein engineering studies, aimed at the identification of regions to be modified in order to obtain more suitable features of local and/or global stability. AVAILABILITY: The tool is available to academic and commercial institutions at the URL http://crisceb.area.na.cnr.it/angelo/ PROTEIN_TOOLS/HELM/ CONTACT: angelo@crisceb.area.na.cnr.it PMID- 10869025 TI - EDIBLE: experimental design and information calculations in phylogenetics. AB - Although evolutionary inference from molecular sequences is a statistical problem, little attention has been paid to questions of experimental design. A computer program, EDIBLE, has been developed to perform likelihood calculations based on Markov process models of nucleotide substitution allied with phylogenetic trees, and from these to compute Fisher information measures under different experimental designs. These calculations can be used to answer questions of optimal experimental design in molecular phylogenetics. AVAILABILITY: Source code (ANSI C), executables and documentation for EDIBLE are available from http://ng-dec1.gen.cam. ac.uk/info/index.htmland 'downstream' Web pages. CONTACT: N.Goldman@gen.cam.ac.uk PMID- 10869026 TI - RRTree: relative-rate tests between groups of sequences on a phylogenetic tree. AB - RRTree is a user-friendly program for comparing substitution rates between lineages of protein or DNA sequences, relative to an outgroup, through relative rate tests. Genetic diversity is taken into account through use of several sequences, and phylogenetic relations are integrated by topological weighting. AVAILABILITY: The ANSI C source code of RRTree, and compiled versions for Macintosh, MS-DOS/Windows, SUN Solaris, and CGI, are freely available at http://pbil.univ-lyon1.fr/software/rrtree.html CONTACT: marc.robinson@ens-lyon.fr PMID- 10869027 TI - PRoMT: inferring demographic history from DNA sequences. AB - I describe a parallel implementation of Rogers' mismatch algorithm, a method for making inferences about demographic history from DNA sequence data. The program is distributed on clusters of workstations, providing a substantial speedup and low execution times on large numbers of nodes. AVAILABILITY: Source code and documentation are available at http://mombasa.anthro.utah.edu/wooding/ CONTACT: stephen.wooding@anthro.utah.edu PMID- 10869028 TI - GABAagent: a system for integrating data on GABA receptors. AB - MOTIVATION: Scientific data pertaining to GABA receptors, which are of medical importance, are widely scattered throughout numerous heterogeneous Internet resources. This situation has made the integrated acquisition of such data difficult and substantially time consuming even for researchers who are Internet aficionados. Thus, there exists a genuine need for the development of Internet applications, such as GABAagent, which provide efficient and timely access to concise and integrated information. RESULTS: We report here the establishment of a novel server (GABAagent) which has been written in Perl script, and which is freely accessible through the Internet. GABAagent is designed to assist researchers in retrieving focused and integrated information related to GABA receptors from various public domain databases. GABAagent relies on server-side flat-file databases that have been created through data mining from Internet sources such as the PubMed, DDBJ, SWISS-PROT and TrEMBL, in addition to the many World Wide Web (Web) sites which are accessible through Excite (E-Web). These warehouse databases are regularly updated and contain among other things, information concerning: (i) GABA receptor publications, (ii) DNA and protein sequences and (iii) the contents of related E-Web sites along with their addresses. Our system also provides hard links to the above-mentioned Web sites and E-Web sites; the feature which adds to it the character of virtual federation type of database. The current version of GABAagent provides two user-friendly services. The first is a search engine possessing intelligent query reformulation support (GABAengine), the second an elaborate email alert service was designed into the system (GABAalert). The GABAengine allows the user to search server-side databases exclusively for GABA receptor-related queries. Whereas, GABAalert allows the user, by means of subscription, to receive immediate and/or monthly updates automatically. AVAILABILITY: GABAagent is freely accessible at the following Web address http://www.ust.hk/gaba. PMID- 10869029 TI - JESAM: CORBA software components to create and publish EST alignments and clusters. AB - MOTIVATION: Expressed Sequence Tags (ESTs) are cheap, easy and quick to obtain relative to full genomic sequencing and currently sample more eukaryotic genes than any other data source. They are particularly useful for developing Sequence Tag Sites (STSs for mapping), polymorphism discovery, disease gene hunting, mass spectrometer proteomics, and most ironically for finding genes and predicting gene structure after the great effort of genomic sequencing. However, ESTs have many problems and the public EST databases contain all the errors and high redundancy intrinsic to the submitted data so it is often found that derived database views, which reduce both errors and redundancy, are more effective starting points for research than the original raw submissions. Existing derived views such as EST cluster databases and consensus databases have never published supporting evidence or intermediary results leading to difficulties trusting, correcting, and customizing the final published database. These difficulties have lead many groups to wastefully repeat the complex intermediary work of others in order to offer slightly different final views. A better approach might be to discover the most expensive common calculations used by all the approaches and then publish all intermediary results. Given a globally accessible database with a suitable component interface, like the JESAM software described in this paper, the creation of customized EST-derived databases could be achieved with minimum effort. RESULTS: Databases of EST and full-length mRNA sequences for four model organisms have been self-compared by searching for overlaps consistent with contiguity. The sequence comparisons are performed in parallel using a PVM process farm and previous results are stored to allow incremental updates with minimal effort. The overlap databases have been published with CORBA interfaces to enable flexible global access as demonstrated by example Java applet browsers. Simple cDNA supercluster databases built as alignment database clients are themselves published via CORBA interfaces browsable with prototypical applets. A comparison with UniGene Mouse and Rat databases revealed undesirable features in both and the advantages of contrasting perspectives on complex data. AVAILABILITY: The software is packaged as two Jar files available from: URL: http://corba.ebi.ac.uk/EST/jesam/jesam. html. One jar contains all the Java source code, and the other contains all the C, C++ and IDL code. Links to working examples of the alignment and cluster viewers (if remote firewall permits) can be found at http://corba.ebi.ac.uk/EST. All the Washington University mouse EST traces are available for browsing at the same URL. PMID- 10869030 TI - Automatic discovery of regulatory patterns in promoter regions based on whole cell expression data and functional annotation. AB - MOTIVATION: The whole genomes submitted to GenBank contain valuable information about the function of genes as well as the upstream sequences and whole cell expression provides valuable information on gene regulation. To utilize these large amounts of data for a biological understanding of the regulation of gene expression, new automatic methods for pattern finding are needed. RESULTS: Two word-analysis algorithms for automatic discovery of regulatory sequence elements have been developed. We show that sequence patterns correlated to whole cell expression data can be found using Kolmogorov-Smirnov tests on the raw data, thereby eliminating the need for clustering co-regulated genes. Regulatory elements have also been identified by systematic calculations of the significance of correlations between words found in the functional annotation of genes and DNA words occurring in their promoter regions. Application of these algorithms to the Saccharomyces cerevisiae genome and publicly available DNA array data sets revealed a highly conserved 9-mer occurring in the upstream regions of genes coding for proteasomal subunits. Several other putative and known regulatory elements were also found. AVAILABILITY: Upon request. PMID- 10869031 TI - The language of RNA: a formal grammar that includes pseudoknots. AB - MOTIVATION: In a previous paper, we presented a polynomial time dynamic programming algorithm for predicting optimal RNA secondary structure including pseudoknots. However, a formal grammatical representation for RNA secondary structure with pseudoknots was still lacking. RESULTS: Here we show a one-to-one correspondence between that algorithm and a formal transformational grammar. This grammar class encompasses the context-free grammars and goes beyond to generate pseudoknotted structures. The pseudoknot grammar avoids the use of general context-sensitive rules by introducing a small number of auxiliary symbols used to reorder the strings generated by an otherwise context-free grammar. This formal representation of the residue correlations in RNA structure is important because it means we can build full probabilistic models of RNA secondary structure, including pseudoknots, and use them to optimally parse sequences in polynomial time. PMID- 10869032 TI - SPLASH: structural pattern localization analysis by sequential histograms. AB - MOTIVATION: The discovery of sparse amino acid patterns that match repeatedly in a set of protein sequences is an important problem in computational biology. Statistically significant patterns, that is patterns that occur more frequently than expected, may identify regions that have been preserved by evolution and which may therefore play a key functional or structural role. Sparseness can be important because a handful of non-contiguous residues may play a key role, while others, in between, may be changed without significant loss of function or structure. Similar arguments may be applied to conserved DNA patterns. Available sparse pattern discovery algorithms are either inefficient or impose limitations on the type of patterns that can be discovered. RESULTS: This paper introduces a deterministic pattern discovery algorithm, called Splash, which can find sparse amino or nucleic acid patterns matching identically or similarly in a set of protein or DNA sequences. Sparse patterns of any length, up to the size of the input sequence, can be discovered without significant loss in performances. Splash is extremely efficient and embarrassingly parallel by nature. Large databases, such as a complete genome or the non-redundant SWISS-PROT database can be processed in a few hours on a typical workstation. Alternatively, a protein family or superfamily, with low overall homology, can be analyzed to discover common functional or structural signatures. Some examples of biologically interesting motifs discovered by Splash are reported for the histone I and for the G-Protein Coupled Receptor families. Due to its efficiency, Splash can be used to systematically and exhaustively identify conserved regions in protein family sets. These can then be used to build accurate and sensitive PSSM or HMM models for sequence analysis. AVAILABILITY: Splash is available to non-commercial research centers upon request, conditional on the signing of a test field agreement. CONTACT: acal@us.ibm.com, Splash main page http://www.research.ibm.com/splash PMID- 10869033 TI - PROF_ PAT 1.3: updated database of patterns used to detect local similarities. AB - MOTIVATION: When analysing novel protein sequences, it is now essential to extend search strategies to include a range of 'secondary' databases. Pattern databases have become vital tools for identifying distant relationships in sequences, and hence for predicting protein function and structure. The main drawback of such methods is the relatively small representation of proteins in trial samples at the time of their construction. Therefore, a negative result of an amino acid sequence comparison with such a databank forces a researcher to search for similarities in the original protein banks. We developed a database of patterns constructed for groups of related proteins with maximum representation of amino acid sequences of SWISS-PROT in the groups. RESULTS: Software tools and a new method have been designed to construct patterns of protein families. By using such method, a new version of databank of protein family patterns, PROF_ PAT 1.3, is produced. This bank is based on SWISS-PROT (r1.38) and TrEMBL (r1.11), and contains patterns of more than 13 000 groups of related proteins in a format similar to that of the PROSITE. Motifs of patterns, which had the minimum level of probability to be found in random sequences, were selected. Flexible fast search program accompanies the bank. The researcher can specify a similarity matrix (the type PAM, BLOSUM and other). Variable levels of similarity can be set (permitting search strategies ranging from exact matches to increasing levels of 'fuzziness'). AVAILABILITY: The Internet address for comparing sequences with the bank is: http://wwwmgs.bionet.nsc.ru/mgs/programs/prof_pat/. The local version of the bank and search programs (approximately 50 Mb) is available via ftp: ftp://ftp.bionet.nsc. ru/pub/biology/vector/prof_pat/, and ftp://ftp.ebi.ac. uk/pub/databases/prof_pat/. Another appropriate way for its external use is to mail amino acid sequences to bachin@vector.nsc.ru for comparison with PROF_ PAT 1.3. PMID- 10869034 TI - On the convergence of a clustering algorithm for protein-coding regions in microbial genomes. AB - MOTIVATION: As the number of fully sequenced prokaryotic genomes continues to grow rapidly, computational methods for reliably detecting protein-coding regions become even more important. Audic and Claverie (1998) Proc. Natl Acad. Sci. USA, 95, 10026-10031, have proposed a clustering algorithm for protein-coding regions in microbial genomes. The algorithm is based on three Markov models of order k associated with subsequences extracted from a given genome. The parameters of the three Markov models are recursively updated by the algorithm which, in simulations, always appear to converge to a unique stable partition of the genome. The partition corresponds to three kinds of regions: (1) coding on the direct strand, (2) coding on the complementary strand, (3) non-coding. RESULTS: Here we provide an explanation for the convergence of the algorithm by observing that it is essentially a form of the expectation maximization (EM) algorithm applied to the corresponding mixture model. We also provide a partial justification for the uniqueness of the partition based on identifiability. Other possible variations and improvements are briefly discussed. PMID- 10869035 TI - Database of structural motifs in proteins. AB - SUMMARY: The database of structural motifs in proteins (DSMP) contains data relevant to helices, beta-turns, gamma-turns, beta-hairpins, psi-loops, beta alpha-beta motifs, beta-sheets, beta-strands and disulphide bridges extracted from all proteins in the Protein Data Bank primarily using the PROMOTIF program and implemented as a web-based network service using the SRS. The data corresponding to the structural motifs includes; sequence, position in polypeptide chain, geometry, type, unique code, keywords and resolution of crystal structure. This data is available for a representative data set of 1028 protein chains and also for all 10 213 proteins in the Protein Data Bank. The three-dimensional coordinates for all structural motifs (except sheet and disulphide bridge) are also available for the representative data set. Using features in SRS, DSMP can be queried to extract information from one or more structural motifs that may be useful for sequence-structure analysis, prediction, modelling or design. AVAILABILITY: http://www. cdfd.org.in/dsmp.html PMID- 10869036 TI - Wavelet change-point prediction of transmembrane proteins. AB - MOTIVATION: A non-parametric method, based on a wavelet data-dependent threshold technique for change-point analysis, is applied to predict location and topology of helices in transmembrane proteins. A new propensity scale generated from a transmembrane helix database is proposed. RESULTS: We show that wavelet change point performs well for smoothing hydropathy and transmembrane profiles generated using different scales. We investigate which wavelet bases and threshold functions are overall most appropriate to detect transmembrane segments. Prediction accuracy is based on the analysis of two data sets used as standard benchmarks for transmembrane prediction algorithms. The analysis of a test set of 83 proteins results in accuracy per segment equal to 98.2%; the analysis of a 48 proteins blind-test set, i.e. containing proteins not used to generate the propensity scales, results in accuracy per segment equal to 97.4%. We believe that this method can also be applied to the detection of boundaries of other patterns such as G + Cisochores and dot-plots. AVAILABILITY: The transmembrane database, TMALN and source code are available upon request from the authors. PMID- 10869037 TI - TrExML: a maximum-likelihood approach for extensive tree-space exploration. AB - MOTIVATION: Maximum-likelihood analysis of nucleotide and amino acid sequences is a powerful approach for inferring phylogenetic relationships and for comparing evolutionary hypotheses. Because it is a computationally demanding and time consuming process, most algorithms explore only a minute portion of tree-space, with the emphasis on finding the most likely tree while ignoring the less likely, but not significantly worse, trees. However, when such trees exist, it is equally important to identify them to give due consideration to the phylogenetic uncertainty. Consequently, it is necessary to change the focus of these algorithms such that near optimal trees are also identified. RESULTS: This paper presents the Advanced Stepwise Addition Algorithm for exploring tree-space and two algorithms for generating all binary trees on a set of sequences. The Advanced Stepwise Addition Algorithm has been implemented in TrExML, a phylogenetic program for maximum-likelihood analysis of nucleotide sequences. TrExML is shown to be more effective at finding near optimal trees than a similar program, fastDNAml, implying that TrExML offers a better approach to account for phylogenetic uncertainty than has previously been possible. A program, TreeGen, is also described; it generates binary trees on a set of sequences allowing for extensive exploration of tree-space using other programs. AVAILABILITY: TreeGen, TrExML, and the sequence data used to test the programs are available from the following two WWW sites: http://whitetail.bemidji.msus. edu/trexml/and http://jcsmr.anu.edu.au/dmm/humgen.+ ++html. PMID- 10869038 TI - Estimating the rate of molecular evolution: incorporating non-contemporaneous sequences into maximum likelihood phylogenies. AB - MOTIVATION: TipDate is a program that will use sequences that have been isolated at different dates to estimate their rate of molecular evolution. The program provides a maximum likelihood estimate of the rate and also the associated date of the most recent common ancestor of the sequences, under a model which assumes a constant rate of substitution (molecular clock) but which accommodates the dates of isolation. Confidence intervals for these parameters are also estimated. RESULTS: The approach was applied to a sample of 17 dengue virus serotype 4 sequences, isolated at dates ranging from 1956 to 1994. The rate of substitution for this serotype was estimated to be 7.91 x 10(-4) substitutions per site per year (95% confidence intervals of 6.07 x 10(-4), 9.86 x 10(-4)). This is compatible with a date of 1922 (95% confidence intervals of 1900-1936) for the most recent common ancestor of these sequences. AVAILABILITY: TipDate can be obtained by WWW from http://evolve.zoo. ox.ac.uk/software. The package includes the source code, manual and example files. Both UNIX and Apple Macintosh versions are available from the same site. PMID- 10869039 TI - Detecting hypermutations in viral sequences with an emphasis on G --> A hypermutation. AB - SUMMARY: This program compares sequence sets to a reference sequence, tallies G - > A hypermutations, and presents the results in various tables and graphs, which include dinucleotide context, summaries of all observed nucleotide changes, and stop codons introduced by hypermutation. AVAILABILITY: www.hiv.lanl.gov/HYPERMUT/hypermut.html PMID- 10869040 TI - A browser for expression data. AB - SUMMARY: We have written a fully extensible Java application for visually browsing expression data, and clusters of genes or experimental conditions calculated from that data. The application requires a run-time environment for Java2. AVAILABILITY: http://www. sanger.ac.uk/Users/mrp/java/ExpressionBrowser PMID- 10869041 TI - The PSIPRED protein structure prediction server. AB - SUMMARY: The PSIPRED protein structure prediction server allows users to submit a protein sequence, perform a prediction of their choice and receive the results of the prediction both textually via e-mail and graphically via the web. The user may select one of three prediction methods to apply to their sequence: PSIPRED, a highly accurate secondary structure prediction method; MEMSAT 2, a new version of a widely used transmembrane topology prediction method; or GenTHREADER, a sequence profile based fold recognition method. AVAILABILITY: Freely available to non-commercial users at http://globin.bio.warwick.ac.uk/psipred/ PMID- 10869042 TI - Taxonomic markup language: applying XML to systematic data. AB - SUMMARY: An XML document type definition (DTD) is provided for the description of taxonomic relationships between organisms. Two XSL style-sheets for the graphical presentation of simple taxonomic trees are also provided. AVAILABILITY: The DTD and stylesheets along with sample files are available at http://www.albany.edu/gilmr/pubxml/. PMID- 10869044 TI - Mitochondria and the pathogenesis of ALS. PMID- 10869043 TI - Mathematica packages for simulation of experimental genetics. AB - This note describes add-on packages for the Mathematica software system (Wolfram 1996) which allow simulation and analysis of both Mendelian and complex genetic traits in experimental crosses of plants or animals. AVAILABILITY: The add-on packages are freely available at http://www.mathsource.com/cgi-bin/msitem?0209-30 4. SUPPLEMENTARY INFORMATION: A tutorial notebook file is included with the packages at the mathsource site. PMID- 10869045 TI - Cerebral specialization and interhemispheric communication: does the corpus callosum enable the human condition? AB - The surgical disconnection of the cerebral hemispheres creates an extraordinary opportunity to study basic neurological mechanisms: the organization of the sensory and motors systems, the cortical representation of the perceptual and cognitive processes, the lateralization of function, and, perhaps most importantly, how the divided brain yields clues to the nature of conscious experience. Studies of split-brain patients over the last 40 years have resulted in numerous insights into the processes of perception, attention, memory, language and reasoning abilities. When the constellation of findings is considered as a whole, one sees the cortical arena as a patchwork of specialized processes. When this is considered in the light of new studies on the lateralization of functions, it becomes reasonable to suppose that the corpus callosum has enabled the development of the many specialized systems by allowing the reworking of existing cortical areas while preserving existing functions. Thus, while language emerged in the left hemisphere at the cost of pre-existing perceptual systems, the critical features of the bilaterally present perceptual system were spared in the opposite half-brain. By having the callosum serve as the great communication link between redundant systems, a pre-existing system could be jettisoned as new functions developed in one hemisphere, while the other hemisphere could continue to perform the previous functions for both half-brains. Split-brain studies have also revealed the complex mosaic of mental processes that participate in human cognition. And yet, even though each cerebral hemisphere has its own set of capacities, with the left hemisphere specialized for language and speech and major problem-solving capacities and the right hemisphere specialized for tasks such as facial recognition and attentional monitoring, we all have the subjective experience of feeling totally integrated. Indeed, even though many of these functions have an automatic quality to them and are carried out by the brain prior to our conscious awareness of them, our subjective belief and feeling is that we are in charge of our actions. These phenomena appear to be related to our left hemisphere's interpreter, a device that allows us to construct theories about the relationship between perceived events, actions and feelings. PMID- 10869046 TI - The neuropathology of the vegetative state after an acute brain insult. AB - The vegetative state is often described clinically as loss of function of the cortex while the function of the brainstem is preserved. In an attempt to define the structural basis of the vegetative state we have undertaken a detailed neuropathological study of the brains of 49 patients who remained vegetative until death, 1 month to 8 years after an acute brain insult. Of these, 35 had sustained a blunt head injury and 14 some type of acute non-traumatic brain damage. In the traumatic cases the commonest structural abnormalities identified were grades 2 and 3 diffuse axonal injury (25 cases, 71%). The thalamus was abnormal in 28 cases (80%), and in 96% of the cases who survived for more than 3 months. Other abnormalities included ischaemic damage in the neocortex (13 cases, 37%) and intracranial haematoma (nine cases, 26%). In the non-traumatic cases there was diffuse ischaemic damage in the neocortex in nine cases (64%) and focal damage in four (29%); the thalamus was abnormal in every case. There were cases in both groups where the cerebral cortex, the cerebellum and the brainstem were of structurally normal appearance. In every case, however, there was profound damage to the subcortical white matter or to the major relay nuclei of the thalamus, or both. These lesions render any structurally intact cortex unable to function because connections between different cortical areas via the thalamic nuclei are no longer functional, and there is also extensive damage to afferent and efferent cerebral connections. PMID- 10869047 TI - Mitochondrial DNA abnormalities in skeletal muscle of patients with sporadic amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis is a neurodegenerative disease affecting the anterior horn cells of the spinal cord and cortical motor neurons. Previous findings have suggested a specific impairment of mitochondrial function in skeletal muscle of at least a limited number of patients. Applying flavoprotein/NAD(P)H autofluorescence imaging of mitochondrial function in saponin-permeabilized muscle fibres, we detected a heterogeneous distribution of the respiratory chain defect among individual fibres in muscle biopsies of patients (11 out of 17) with sporadic amyotrophic lateral sclerosis (SALS). These findings correlate with the presence of cytochrome c oxidase (COX)-negative muscle fibres detected histologically. We established the molecular basis for the decreased activities of NADH:CoQ oxidoreductase and COX in SALS muscle. In the skeletal muscle of the investigated patients, diminished levels (13 out of 17) or multiple deletions (one out of 17) of mitochondrial DNA (mtDNA) were observed. These alterations of mtDNA seem to be related to decreased levels of membrane associated mitochondrial Mn-superoxide dismutase. Our results support the viewpoint that an oxygen radical-induced impairment of mtDNA is of pathophysiological significance in the aetiology of at least a subgroup of patients with SALS. PMID- 10869048 TI - Visual object and visuospatial cognition in Huntington's disease: implications for information processing in corticostriatal circuits. AB - The primate visual system contains two major streams of visual information processing. The ventral stream is directed into the inferior temporal cortex and is concerned with visual object cognition, whereas the dorsal stream is directed into the posterior parietal cortex and is concerned with visuospatial cognition. Both of these processing streams send projections to the basal ganglia, and the ventral stream may also receive reciprocal connections from the basal ganglia. Although a role for the basal ganglia in visual object and visuospatial cognition has been suggested, little work has been carried out in this area in humans. The primary site of neuropathology in Huntington's disease is the basal ganglia, and hence Huntington's disease provides an important model for the role of the human basal ganglia in visual object and visuospatial cognition, and its breakdown in disease. We examined performance on a wide battery of tests of both visual object and visuospatial recognition memory, working memory, attention, associative learning and perception, enabling us to specify more fully the role of the basal ganglia in visual object and visuospatial cognition, and the disruption of these processes in Huntington's disease. Huntington's disease patients exhibited deficits on tests of pattern and spatial recognition memory; showed impaired simultaneous matching and delay-independent delayed matching-to-sample deficits; showed spared accuracy but impaired reaction times in visual search; were impaired in spatial but not visual object working memory; and showed impaired pattern-location associative learning. The results of our investigations suggest a particular role for the striatum in context-dependent action selection, in line with current computational theories of basal ganglia function. PMID- 10869049 TI - Embryonic ventral mesencephalic grafts improve levodopa-induced dyskinesia in a rat model of Parkinson's disease. AB - We investigated the role of dopamine neurons in the manifestation of levodopa induced dyskinesia in a rat model of Parkinson's disease. Daily treatment with a subthreshold dose of levodopa gradually induced abnormal involuntary movements (AIM) in 6-hydroxydopamine-lesioned rats, which included stereotypy and contraversive rotation. After 4 weeks of levodopa treatment, rats with mild and severe AIM were assigned to two treatment subgroups. The graft subgroup received embryonic ventral mesencephalic tissue into the striatum, whilst the sham-graft subgroup received vehicle only. Rats continued to receive levodopa treatment for 3 months post-graft. Brain sections at the level of the basal ganglia were processed for autoradiography using a ligand for dopamine transporter, and in situ hybridization histochemistry for mRNAs encoding postsynaptic markers. Levodopa-induced AIM significantly improved in grafted rats. The severity of AIM correlated inversely with the density of dopamine nerve terminals in the striatum (P < 0. 001), with almost no AIM when the density of dopamine nerve terminals was >10-20% of normal. Embryonic dopamine neuronal grafts normalized not only mRNA expression for preproenkephalin (PPE) in the indirect pathway, but also mRNA expression for prodynorphin (PDyn) in the direct pathway, which was upregulated by levodopa treatment. AIM scores correlated linearly with expression of PPE mRNA in the indirect pathway (P < 0.001) and also with PDyn mRNA in the direct pathway (P < 0.001). We conclude that embryonic dopamine neuronal grafts may improve levodopa-induced dyskinesia by restoring altered activities of postsynaptic neurons, resulting not only from dopamine denervation, but also from levodopa therapy, provided that the density of striatal dopaminergic nerve terminals is restored above a 'threshold' level. PMID- 10869050 TI - Bilateral caudate and putamen grafts of embryonic mesencephalic tissue treated with lazaroids in Parkinson's disease. AB - Five parkinsonian patients were transplanted bilaterally into the putamen and caudate nucleus with human embryonic mesencephalic tissue from between seven and nine donors. To increase graft survival, the lipid peroxidation inhibitor tirilazad mesylate was administered to the tissue before implantation and intravenously to the patients for 3 days thereafter. During the second postoperative year, the mean daily L-dopa dose was reduced by 54% and the UPDRS (Unified Parkinson's Disease Rating Scale) motor score in 'off' phase was reduced by a mean of 40%. At 10-23 months after grafting, PET showed a mean 61% increase of 6-L-[(18)F]fluorodopa uptake in the putamen, and 24% increase in the caudate nucleus, compared with preoperative values. No obvious differences in the pattern of motor recovery were observed between these and other previously studied cases with putamen grafts alone. The amount of mesencephalic tissue implanted in each putamen and caudate nucleus was 42 and 50% lower, respectively, compared with previously transplanted patients from our centre. Despite this reduction in grafted tissue, the magnitudes of symptomatic relief and graft survival were very similar. These findings suggest that tirilazad mesylate may improve survival of grafted dopamine neurons in patients, which is in agreement with observations in experimental animals. PMID- 10869051 TI - Differential recruitment of the speech processing system in healthy subjects and rehabilitated cochlear implant patients. AB - Differences in cerebral activation between control subjects and post-lingually deaf rehabilitated cochlear implant patients were identified with PET under various speech conditions of different linguistic complexity. Despite almost similar performance in patients and controls, different brain activation patterns were elicited. In patients, an attentional network including prefrontal and parietal modality-aspecific attentional regions and subcortical auditory regions was over-activated irrespective of the nature of the speech stimuli and during expectancy of speech stimuli. A left temporoparietal semantic region was responsive to meaningless stimuli (vowels). In response to meaningful stimuli (words, sentences, story), left middle and inferior temporal semantic regions and posterior superior temporal phonological regions were under-activated in patients, whereas anterior superior temporal phonological regions were over activated. These differences in the recruitment of the speech comprehension system reflect the alternative neural strategies that permit speech comprehension after cochlear implantation. PMID- 10869052 TI - Evidence for cellular damage in normal-appearing white matter correlates with injury severity in patients following traumatic brain injury: A magnetic resonance spectroscopy study. AB - Neuropsychological studies in patients who have suffered traumatic brain injury show that the eventual clinical outcome is frequently worse than might be predicted from using conventional (CT or T(1)/T(2)-weighted MRI) imaging. Furthermore, patients who have sustained an initial mild or moderate injury may show long-term disability. This implies that there may be abnormalities in areas of the brain that actually appear normal on conventional imaging. Proton magnetic resonance spectroscopy studies have shown that N-acetylaspartate and choline containing compounds can provide measures of cellular injury. We report MRI and proton magnetic resonance spectroscopy studies of 19 head-injured patients performed once the patients were clinically stable (mean 11 days after injury, range 3-38 days). Proton magnetic resonance spectra were acquired from frontal white matter that on conventional MRI appeared normal. The brain N acetylaspartate/creatine ratio was reduced [patients (mean +/- standard deviation), 1.28 +/- 0.25; controls, 1.47 +/- 0. 24; P = 0.04] and the choline/creatine ratio was increased (patients, 0.85 +/- 0.18; controls, 0.63 +/- 0.10; P < 0.001) compared with controls. When the severity of the injury was assessed using either the Glasgow coma scale or the length of post-traumatic amnesia, the increase in the choline/creatine ratio was significant even in the mildly injured group (P = 0.008 and P = 0.04, respectively). Furthermore, there was a significant correlation (P = 0.008) between the severity of head injury and the N-acetylaspartate/choline ratio. We conclude that there is an early reduction in N-acetylaspartate and an increase in choline compounds in normal-appearing white matter which correlate with head injury severity, and that this may provide a pathological basis for the long-term neurological disability that is seen in these patients. PMID- 10869053 TI - Loss of thalamic intralaminar nuclei in progressive supranuclear palsy and Parkinson's disease: clinical and therapeutic implications. AB - Whilst many reports mention neurofibrillary tangle pathology in the thalamus in progressive supranuclear palsy, there has been little detailed regional analysis of the distribution and density of thalamic pathology in this disease or in other parkinsonian syndromes. The caudal intralaminar thalamic nuclei are the major thalamic regulators of the caudate nucleus and putamen, areas known to be dysfunctional in progressive supranuclear palsy and Parkinson's disease. We investigated whether these thalamic nuclei degenerate in patients with these disorders compared with age-matched, neurologically normal controls. Neurofibrillary tangle and Lewy body pathology was assessed and unbiased optical disector methods were used to quantify total neuronal number. Despite different thalamic pathology, there was a dramatic reduction in the total neuronal number in the caudal intralaminar nuclei in both progressive supranuclear palsy and Parkinson's disease (40-55% loss). In contrast, there was no loss of volume or total neuronal number in the limbic thalamic nuclei in either disease group, indicating selective degeneration of the caudal intralaminar nuclei. In Parkinson's disease, Lewy bodies were found in these regions, while in progressive supranuclear palsy abundant intracellular neurofibrillary tangles and glial tangles concentrated in the caudal intralaminar nuclei. However, tangle formation accounted for only a small proportion of cell loss (4 days. This was >100-fold longer than that reported in intact cells (2 min) and 40-60-fold longer than from any other known intracellular membrane. Third, when probed with several cholesterol-binding proteins, the initial rate of sterol transfer was maximally increased nearly 80 fold and the organization of cholesterol in the lysosomal membrane was rapidly altered. Nearly half of the essentially nonexchangeable sterol in the lysosomal membrane was converted to rapidly (t(1/2) = 6 min; fraction = 0.06) and slowly (t(1/2) = 154 min; fraction = 0.36) exchangeable sterol domains/pools. In summary, the data revealed that spontaneous cholesterol transfer out of the lysosome and lysosomal membrane was extremely slow, inconsistent with rapid spontaneous diffusion across the lysosomal membrane. In contrast, the very slow spontaneous transfer of sterol out of the lysosome and lysosomal membrane was consistent with cholesterol leaving the lysosome earlier in the endocytic process and/or with cholesterol transfer out of the lysosome being mediated by additional process(es) extrinsic to the lysosome and lysosomal membrane. PMID- 10869173 TI - Structural and biochemical characterization of recombinant wild type and a C30A mutant of trimethylamine dehydrogenase from methylophilus methylotrophus (sp. W(3)A(1)). AB - Trimethylamine dehydrogenase (TMADH) is an iron-sulfur flavoprotein that catalyzes the oxidative demethylation of trimethylamine to form dimethylamine and formaldehyde. It contains a unique flavin, in the form of a 6-S-cysteinyl FMN, which is bent by approximately 25 degrees along the N5-N10 axis of the flavin isoalloxazine ring. This unusual conformation is thought to modulate the properties of the flavin to facilitate catalysis, and has been postulated to be the result of covalent linkage to Cys-30 at the flavin C6 atom. We report here the crystal structures of recombinant wild-type and the C30A mutant TMADH enzymes, both determined at 2.2 A resolution. Combined crystallographic and NMR studies reveal the presence of inorganic phosphate in the FMN binding site in the deflavo fraction of both recombinant wild-type and C30A proteins. The presence of tightly bound inorganic phosphate in the recombinant enzymes explains the inability to reconstitute the deflavo forms of the recombinant wild-type and C30A enzymes that are generated in vivo. The active site structure and flavin conformation in C30A TMADH are identical to those in recombinant and native TMADH, thus revealing that, contrary to expectation, the 6-S-cysteinyl FMN link is not responsible for the 25 degrees butterfly bending along the N5-N10 axis of the flavin in TMADH. Computational quantum chemistry studies strongly support the proposed role of the butterfly bend in modulating the redox properties of the flavin. Solution studies reveal major differences in the kinetic behavior of the wild-type and C30A proteins. Computational studies reveal a hitherto, unrecognized, contribution made by the S(gamma) atom of Cys-30 to substrate binding, and a role for Cys-30 in the optimal geometrical alignment of substrate with the 6-S-cysteinyl FMN in the enzyme active site. PMID- 10869174 TI - X-ray structure of a truncated form of cytochrome f from chlamydomonas reinhardtii. AB - A truncated form of cytochrome f from Chlamydomonas reinhardtii (an important eukaryotic model organism for photosynthetic electron transfer studies) has been crystallized (space group P2(1)2(1)2(1); three molecules/asymmetric unit) and its structure determined to 2.0 A resolution by molecular replacement using the coordinates of a truncated turnip cytochrome f as a model. The structure displays the same folding and detailed features as turnip cytochrome f, including (a) an unusual heme Fe ligation by the alpha-amino group of tyrosine 1, (b) a cluster of lysine residues (proposed docking site of plastocyanin), and (c) the presence of a chain of seven water molecules bound to conserved residues and extending between the heme pocket and K58 and K66 at the lysine cluster. For this array of waters, we propose a structural role. Two cytochrome f molecules are related by a noncrystallographic symmetry operator which is a distorted proper 2-fold rotation. This may represent the dimeric relation of the monomers in situ; however, the heme orientation suggested by this model is not consistent with previous EPR measurements on oriented membranes. PMID- 10869175 TI - Spectroscopic studies of inhibited alcohol dehydrogenase from Thermoanaerobacter brockii: proposed structure for the catalytic intermediate state. AB - Thermoanaerobacter brockii alcohol dehydrogenase (TbADH) catalyzes the reversible oxidation of secondary alcohols to the corresponding ketones using NADP(+) as the cofactor. The active site of the enzyme contains a zinc ion that is tetrahedrally coordinated by four protein residues. The enzymatic reaction leads to the formation of a ternary enzyme-cofactor-substrate complex; and catalytic hydride ion transfer is believed to take place directly between the substrate and cofactor at the ternary complex. Although crystallographic data of TbADH and other alcohol dehydrogenases as well as their complexes are available, their mode of action remains to be determined. It is firmly established that the zinc ion is essential for catalysis. However, there is no clear agreement about the coordination environment of the metal ion and the competent reaction intermediates during catalysis. We used a combination of X-ray absorption, circular dichroism (CD), and fluorescence spectroscopy, together with structural analysis and modeling studies, to investigate the ternary complexes of TbADH that are bound to a transition-state analogue inhibitor. Our structural and spectroscopic studies indicated that the coordination sphere of the catalytic zinc site in TbADH undergoes conformational changes when it binds the inhibitor and forms a pentacoordinated complex at the zinc ion. These studies provide the first active site structure of bacterial ADH bound to a substrate analogue. Here, we suggest the active site structure of the central intermediate complex and, more specifically, propose the substrate-binding site in TbADH. PMID- 10869176 TI - The annexin A3-membrane interaction is modulated by an N-terminal tryptophan. AB - The crystal structure of annexin A3 (human annexin III) solved recently revealed a well-ordered folding of its N-terminus with the side chain of tryptophan 5 interacting with residues at the extremity of the central pore. Since the pore of annexins has been suggested as the ion pathway involved in membrane permeabilization by these proteins, we investigated the effect of the N-terminal tryptophan on the channel activity of annexin A3 by a comparative study of the wild-type and the W5A mutant in structural and functional aspects. Calcium influx and patch-clamp recordings revealed that the mutant exhibited an enhanced membrane permeabilization activity as compared to the wild-type protein. Analysis of the phospholipid binding behavior of wild-type and mutant protein was carried out by cosedimentation with lipids and inhibition of PLA(2) activity. Both methods reveal a much stronger binding of the mutant to phospholipids. The structure is very similar for the wild-type and the mutant protein. The exchange of the tryptophan for an alanine results in a disordered N-terminal segment. Urea induced denaturation of the wild-type and mutant monitored by intrinsic fluorescence indicates a separate unfolding of the N-terminal region which occurs at lower urea concentrations than unfolding of the protein core. We therefore conclude that the N-terminal domain of annexin A3, and especially tryptophan 5, is involved in the modulation of membrane binding and permeabilization by annexin A3. PMID- 10869177 TI - Circular dichroic investigation of the native and non-native conformational states of the growth factor receptor-binding protein 2 N-terminal src homology domain 3: effect of binding to a proline-rich peptide from guanine nucleotide exchange factor. AB - SH3 (src homology domain 3) domains are small protein modules that interact with proline-rich peptides. The structure of the N-terminal SH3 domain from growth factor receptor-binding protein 2 (Grb2), an adapter protein in the intracellular signaling pathway to Ras, was investigated by circular dichroic (CD) spectroscopy. The compact native beta-barrel conformation, previously elucidated by NMR spectroscopy, was largely predominant at pH = 4.8, in the absence of salt. From the structural changes induced by varying pH, ionic strength, temperature, or hydrophobicity of the environment, evidence for the existence of distinct nonnative conformations was obtained in the far- and near-UV domains. Along the free energy scale, these appear to distribute into two conformational ensembles, depending on the extent of structural and thermodynamic differences compared to the native conformation. The first ensemble consists of non-native conformations with a nativelike secondary structure, and the second is composed of partially unfolded conformations having short alpha-helical fragments or turnlike motifs in their nonnative secondary structure. Most of the observed nonnative conformations exist in mild or nondenaturing conditions. They probably have distinct compactness of their inner structure, depending on the strength of nonlocal interactions, but only the native all-beta conformation possesses a condensed protein exterior, appropriate for the binding to the VPPPVPPRRR decapeptide from Sos. Upon binding, the native conformation undergoes a local tertiary structure change in a hydrophobic pocket at the binding site. This is accompanied by the PP II helix folding of the proline-rich peptide. Interestingly, in the near-UV domain, a significant change in the spectral contribution of an aromatic exciton was observed, thus allowing quantitative tracking of the binding process. PMID- 10869178 TI - Regulation of phospholipase C-beta(1) activity by phosphatidic acid. AB - The role of phosphatidic acid (PA) in regulating phospholipase C-beta(1) (PLC beta(1)) activity was determined. PA promoted the binding of PLC-beta(1) to sucrose-loaded unilamellar vesicles (SLUV) containing phosphatidylcholine. PA increased enzymatic activity over a range of Ca(2+) concentrations and reduced the Ca(2+) concentration required for half-maximal stimulation of activity. PA did not affect the apparent K(m) for phosphatidylinositol 4, 5-bisphosphate. Lysophosphatidic acid also enhanced the binding of PLC-beta(1) to SLUV but was less effective in stimulating enzymatic activity. Diacylglycerol, phosphatidylserine, and oleic acid had little effect on activity. Anionic and neutral detergents did not stimulate activity. PA stimulation was relatively independent of acyl chain length. Dipalmitoyl-PA (16:0) was comparable to PA from egg lecithin and dimyristoyl-PA (C14:0) in stimulating activity, while dilauroyl PA (C12:0) was slightly less effective. A 100 kDa catalytic fragment of PLC beta(1) lacking amino acid residues C-terminal to His(880) did not bind to PA and was insensitive to stimulation by 7-15 mol % PA. Stimulation of 100 kDa enzymatic activity required 30 mol % PA. PA increased receptor-G protein stimulation of PLC beta(1) activity in membranes. These results demonstrate that PA stimulates basal and receptor-G protein-regulated PLC-beta(1) activity. PA stimulation occurs through both a C-terminal-dependent and an independent mechanism. The C-terminal mediated mechanism for stimulation may constitute an important pathway for conferring specific regulation of PLC-beta(1) in response to increases in cellular PA levels. PMID- 10869179 TI - Study of the secondary structure of the C-terminal domain of the antiapoptotic protein bcl-2 and its interaction with model membranes. AB - Bcl-2 is a protein which inhibits programmed cell death. It is associated to many cell membranes such as mitochondrial outer membrane, endoplasmic reticulum, and nuclear envelope, apparently through a C-terminal hydrophobic domain. We have used infrared spectroscopy to study the secondary structure of a synthetic peptide (a 23mer) with the same sequence as this C-terminal domain (residues 217 239) of Bcl-2. The spectrum of this peptide in D(2)O buffer shows an amide I' band with a maximum at 1622 cm(-1), which clearly indicates its tendency to aggregate in aqueous solvent. However, the peptide incorporated in multilamellar phosphatidylcholine membranes shows a totally different spectrum of the amide I' band, with a maximum at 1655 cm(-)(1), indicating a predominantly alpha-helical structure. Addition of the peptide to unilamellar vesicles destabilized them and released encapsulated carboxyfluorescein. Differential scanning calorimetry of dimyristoylphosphatidylcholine multilamellar vesicles in which the peptide was incorporated revealed that increasing concentrations of the peptide progressively broadened the pretransition and the main transition, as is to be expected for a membrane integral molecule. Fluorescence polarization of 1,6-diphenyl-1,3,5 hexatriene in fluid phosphatidylcholine vesicles showed that increasing concentrations of the peptide produced increased polarization values, pointing to an increase in the apparent order of the membrane and indicating that high concentrations of the peptide considerably broaden the phase transition of dimyristoylphosphatidylcholine multilamellar vesicles. Quenching the intrinsic fluorescence of the Tyr-235 of the peptide, by KI, indicated that this aminoacyl residue is highly exposed to aqueous solvent when incorporated in phospholipid vesicles. The results are discussed in terms of their relevance to the proposed topology of insertion of Bcl-2 into biological membranes. PMID- 10869180 TI - Acetylthiocholine binds to asp74 at the peripheral site of human acetylcholinesterase as the first step in the catalytic pathway. AB - Studies of ligand binding to acetylcholinesterase (AChE) have demonstrated two sites of interaction. An acyl-enzyme intermediate is formed at the acylation site, and catalytic activity can be inhibited by ligand binding to a peripheral site. The three-dimensional structures of AChE-ligand complexes reveal a narrow and deep active site gorge and indicate that ligands specific for the acylation site at the base of the gorge must first traverse the peripheral site near the gorge entrance. In recent studies attempting to clarify the role of the peripheral site in the catalytic pathway for AChE, we showed that ligands which bind specifically to the peripheral site can slow the rates at which other ligands enter and exit the acylation site, a feature we called steric blockade [Szegletes, T., Mallender, W. D., and Rosenberry, T. L. (1998) Biochemistry 37, 4206-4216]. We also demonstrated that cationic substrates can form a low-affinity complex at the peripheral site that accelerates catalytic hydrolysis at low substrate concentrations but results in substrate inhibition at high concentrations because of steric blockade of product release [Szegletes, T., Mallender, W. D., Thomas, P. J., and Rosenberry, T. L. (1999) Biochemistry 38, 122-133]. In this report, we demonstrate that a key residue in the human AChE peripheral site with which the substrate acetylthiocholine interacts is D74. We extend our kinetic model to evaluate the substrate affinity for the peripheral site, indicated by the equilibrium dissociation constant K(S), from the dependence of the substrate hydrolysis rate on substrate concentration. For human AChE, a K(S) of 1.9+/-0.7 mM obtained by fitting this substrate inhibition curve agreed with a K(S) of 1.3+/-1.0 mM measured directly from acetylthiocholine inhibition of the binding of the neurotoxin fasciculin to the peripheral site. For Torpedo AChE, a K(S) of 0.5+/- 0.2 mM obtained from substrate inhibition agreed with a K(S) of 0.4+/- 0.2 mM measured with fasciculin. Introduction of the D72G mutation (corresponding to D74G in human AChE) increased the K(S) to 4-10 mM in the Torpedo enzyme and to about 33 mM in the human enzyme. While the turnover number k(cat) was unchanged in the human D74G mutant, the roughly 20-fold decrease in acetylthiocholine affinity for the peripheral site in D74G resulted in a corresponding decrease in k(cat)/K(app), the second-order hydrolysis rate constant, in the mutant. In addition, we show that D74 is important in conveying to the acylation site an inhibitory conformational effect induced by the binding of fasciculin to the peripheral site. This inhibitory effect, measured by the relative decrease in the first-order phosphorylation rate constant k(OP) for the neutral organophosphate 7-[(methylethoxyphosphonyl)oxy]-4-methylcoumarin (EMPC) that resulted from fasciculin binding, decreased from 0.002 in wild-type human AChE to 0.24 in the D74G mutant. PMID- 10869181 TI - Identification of catalytic nucleophile of Escherichia coli gamma glutamyltranspeptidase by gamma-monofluorophosphono derivative of glutamic acid: N-terminal thr-391 in small subunit is the nucleophile. AB - gamma-Glutamyltranspeptidase (EC 2.3.2.2) is the enzyme involved in glutathione metabolism and catalyzes the hydrolysis and transpeptidation of gamma-glutamyl compounds such as glutathione and its derivatives. The reaction is thought to proceed via a gamma-glutamyl-enzyme intermediate where a hitherto unknown catalytic nucleophile is gamma-glutamylated. Neither affinity labeling nor site directed mutagenesis of conserved amino acids has succeeded so far in identifying the catalytic nucleophile. We describe here the identification of the catalytic nucleophile of Escherichia coli gamma-glutamyltranspeptidase by a novel mechanism based affinity labeling agent, 2-amino-4-(fluorophosphono)butanoic acid (1), a gamma-phosphonic acid monofluoride derivative of glutamic acid. Compound 1 rapidly inactivated the enzyme in a time-dependent manner (k(on) = 4.83 x 10(4) M(-1) s(-1)). The inactivation rate was decreased by increasing the concentration of the substrate. The inactivated enzyme did not regain its activity after prolonged dialysis, suggesting that 1 served as an active-site-directed affinity label by phosphonylating the putative catalytic nucleophile. Ion-spray mass spectrometric analyses revealed that one molecule of 1 phosphonylated one molecule of the small subunit. LC/MS experiments of the proteolytic digests of the phosphonylated small subunit identified the N-terminal peptide Thr391-Lys399 as the phosphonylation site. Subsequent MS/MS experiments of this peptide revealed that the phosphonylated residue was Thr-391, the N-terminal residue of the small subunit. We conclude that the N-terminal Thr-391 is the catalytic nucleophile of E. coli gamma-glutamyltranspeptidase. This result strongly suggests that gamma-glutamyltranspeptidase is a new member of the N-terminal nucleophile hydrolase family. PMID- 10869182 TI - Structures of maltohexaose and maltoheptaose bound at the donor sites of cyclodextrin glycosyltransferase give insight into the mechanisms of transglycosylation activity and cyclodextrin size specificity. AB - The enzymes from the alpha-amylase family all share a similar alpha-retaining catalytic mechanism but can have different reaction and product specificities. One family member, cyclodextrin glycosyltransferase (CGTase), has an uncommonly high transglycosylation activity and is able to form cyclodextrins. We have determined the 2.0 and 2.5 A X-ray structures of E257A/D229A CGTase in complex with maltoheptaose and maltohexaose. Both sugars are bound at the donor subsites of the active site and the acceptor subsites are empty. These structures mimic a reaction stage in which a covalent enzyme-sugar intermediate awaits binding of an acceptor molecule. Comparison of these structures with CGTase-substrate and CGTase-product complexes reveals three different conformational states for the CGTase active site that are characterized by different orientations of the centrally located residue Tyr 195. In the maltoheptaose and maltohexaose complexed conformation, CGTase hinders binding of an acceptor sugar at subsite +1, which suggests an induced-fit mechanism that could explain the transglycosylation activity of CGTase. In addition, the maltoheptaose and maltohexaose complexes give insight into the cyclodextrin size specificity of CGTases, since they precede alpha-cyclodextrin (six glucoses) and beta cyclodextrin (seven glucoses) formation, respectively. Both ligands show conformational differences at specific sugar binding subsites, suggesting that these determine cyclodextrin product size specificity, which is confirmed by site directed mutagenesis experiments. PMID- 10869183 TI - Thermodynamics of sequence-specific binding of PNA to DNA. AB - For further characterization of the hybridization properties of peptide nucleic acids (PNAs), the thermodynamics of hybridization of mixed sequence PNA-DNA duplexes have been studied. We have characterized the binding of PNA to DNA in terms of binding affinity (perfectly matched duplexes) and sequence specificity of binding (singly mismatched duplexes) using mainly absorption hypochromicity melting curves and isothermal titration calorimetry. For perfectly sequence matched duplexes of varying lengths (6-20 bp), the average free energy of binding (DeltaG degrees ) was determined to be -6.5+/-0.3 kJ mol(-1) bp(-1), corresponding to a microscopic binding constant of about 14 M(-1) bp(-1). A variety of single mismatches were introduced in 9- and 12-mer PNA-DNA duplexes. Melting temperatures (T(m)) of 9- and 12-mer PNA-DNA duplexes with a single mismatch dropped typically 15-20 degrees C relative to that of the perfectly matched sequence with a corresponding free energy penalty of about 15 kJ mol(-1) bp(-1). The average cost of a single mismatch is therefore estimated to be on the order of or larger than the gain of two matched base pairs, resulting in an apparent binding constant of only 0.02 M(-1) per mismatch. The impact of a mismatch was found to be dependent on the neighboring base pairs. To a first approximation, increasing the stability of the surrounding region, i.e., the distribution of A.T and G.C base pairs, decreases the effect of the introduced mismatch. PMID- 10869184 TI - Synthesis of cysteinyl-tRNA(Cys) by a genome that lacks the normal cysteine-tRNA synthetase. AB - Synthesis of cysteinyl-tRNA(Cys) by cysteine-tRNA synthetase is required for decoding cysteine codons in all known organisms. The genome of the archaeon Methanococcus jannaschii lacks the gene for a normal cysteine-tRNA synthetase. The activity of the enzyme, however, was identified recently, and it allowed the purification of the enzyme and cloning of its gene. Sequence analysis of the gene showed that it encodes proline-tRNA synthetase and, thus, raised the possibility of dual activities in a single aminoacyl-tRNA synthetase. Assays of aminoacyl adenylate synthesis confirmed the ability of the enzyme to activate proline and cysteine and showed that both activities were independent of tRNA. Assays of tRNA aminoacylation established the specific attachment of proline to tRNA(Pro) and cysteine to tRNA(Cys). However, in contrast to a recent report of comparable activities with cysteine and proline, results here indicate that the adenylate synthesis and aminoacylation activities with cysteine are significantly lower than the respective activity with proline. In addition, there is evidence of overlapping amino acid-binding sites and tRNA-binding sites. These considerations, among others, raised the distinct possibility that the M. jannaschii proline-tRNA synthetase may recruit additional protein or RNA factors to facilitate the synthesis of cysteinyl-tRNA(Cys). PMID- 10869185 TI - Interaction of lipid-bound myelin basic protein with actin filaments and calmodulin. AB - Myelin basic protein (MBP) binds to negatively charged lipids on the cytosolic surface of oligodendrocytes (OLs) and is believed to be responsible for adhesion of these surfaces in the multilayered myelin sheath. MBP in solution has been shown by others to bind to both G- and F-actin, to bundle F-actin filaments, and to induce polymerization of G-actin. Here we show that MBP bound to acidic lipids can also bind to both G- and F-actin and cause their sedimentation together with MBP-lipid vesicles. Thus it can simultaneously utilize some of its basic residues to bind to the lipid bilayer and some to bind to actin. The amount of actin bound to the MBP-lipid vesicles decreased with increasing net negative surface charge of the lipid vesicles. It was also less for vesicles containing the lipid composition predicted for the cytosolic surface of myelin than for PC vesicles containing a similar amount of an acidic lipid. Calmodulin caused dissociation of actin from MBP and of the MBP-actin complex from the vesicles. However, it did not cause dissociation of bundles of actin filaments once these had formed as long as some MBP was still present. These results suggest that MBP could be a membrane actin-binding protein in OLs/myelin and its actin binding can be regulated by calmodulin and by the lipid composition of the membrane. Actin binding to MBP decreased the labeling of MBP by the hydrophobic photolabel 3 (trifluoromethyl)-3-(m-[(125)I]iodophenyl)diazirine (TID), indicating that it decreased the hydrophobic interactions of MBP with the bilayer. This change in interaction of MBP with the bilayer could then create a cytosol to membrane signal caused by changes in interaction of the cytoskeleton with the membrane. PMID- 10869187 TI - Vibrio harveyi NADPH-FMN oxidoreductase arg203 as a critical residue for NADPH recognition and binding. AB - Luminous bacteria contain three types of NAD(P)H-FMN oxidoreductases (flavin reductases) with different pyridine nucleotide specificities. Among them, the NADPH-specific flavin reductase from Vibrio harveyi exhibits a uniquely high preference for NADPH. In comparing the substrate specificity, crystal structure, and primary sequence of this flavin reductase with other structurally related proteins, we hypothesize that the conserved Arg203 residue of this reductase is critical to the specific recognition of NADPH. The mutation of this residue to an alanine resulted in only small changes in the binding and reduction potential of the FMN cofactor, the K(m) for the FMN substrate, and the k(cat). In contrast, the K(m) for NADPH was increased 36-fold by such a mutation. The characteristic perturbation of the FMN cofactor absorption spectrum upon NADP(+) binding by the wild-type reductase was abolished by the same mutation. While the k(cat)/K(m,NADPH) was reduced from 1990 x 10(5) to 46 x 10(5) M(-1) min(-1) by the mutation, the mutated variant showed a k(cat)/K(m,NADH) of 4 x 10(5) M(-1) min(-1), closely resembling that of the wild-type reductase. The deuterium isotope effects (D)V and (D)(V/K) for (4R)-[4-(2)H]-NADPH were 1.7 and 1.4, respectively, for the wild-type reductase but were increased to 3.8 and 4.0, respectively, for the mutated variant. Such a finding indicates that the rates of NADPH and NADP(+) dissociation in relation to the isotope-sensitive redox steps were both increased as a result of the mutation. These results all provide support to the critical role of the Arg203 in the specific recognition and binding of NADPH. PMID- 10869186 TI - Regulation of RYR1 activity by Ca(2+) and calmodulin. AB - The skeletal muscle calcium release channel (RYR1) is a Ca(2+)-binding protein that is regulated by another Ca(2+)-binding protein, calmodulin. The functional consequences of calmodulin's interaction with RYR1 are dependent on Ca(2+) concentration. At nanomolar Ca(2+) concentrations, calmodulin is an activator, but at micromolar Ca(2+) concentrations, calmodulin is an inhibitor of RYR1. This raises the question of whether the Ca(2+)-dependent effects of calmodulin on RYR1 function are due to Ca(2+) binding to calmodulin, RYR1, or both. To distinguish the effects of Ca(2+) binding to calmodulin from those of Ca(2+) binding to RYR1, a mutant calmodulin that cannot bind Ca(2+) was used to evaluate the effects of Ca(2+)-free calmodulin on Ca(2+)-bound RYR1. We demonstrate that Ca(2+)-free calmodulin enhances the affinity of RYR1 for Ca(2+) while Ca(2+) binding to calmodulin converts calmodulin from an activator to an inhibitor. Furthermore, Ca(2+) binding to RYR1 enhances its affinity for both Ca(2+)-free and Ca(2+) bound calmodulin. PMID- 10869188 TI - Involvement of conserved aspartate and glutamate residues in the catalysis and substrate binding of maize starch synthase. AB - Chemical modification of maize starch synthase IIb-2 (SSIIb-2) using 1-ethyl-3-(3 dimethylaminopropyl)carbodiimide (EDAC), which modifies acidic amino acid residues, resulted in a time- and concentration-dependent inactivation of SSIIb 2. ADPGlc was found to completely protect SSIIb-2 from inactivation by EDAC. These results suggest that glutamate or aspartate is important for SS activity. On the basis of the sequence identity of SS, conserved acidic amino acids were mutagenized to identify the specific amino acid residues important for SS activity. Three amino acids (D21, D139, and E391) were found to be important for SS activity. D21N showed 4% of the wild-type enzyme activity and a 10-fold decrease in the affinity for ADPGlc, while the conservative change from D21 to E resulted in a decrease in V(max) and no change in affinity for ADPGlc, suggesting that the negative charge is important for ADPGlc binding. When sites D139 and E391 were changed to their respective amide form, no SS activity was detected. With the conservative change, D139E showed a decrease in V(max) and no changes in apparent K(m) for substrates. E391D showed a 9-fold increase in K(m) for ADPGlc, a 12-fold increase in apparent K(m) for glycogen, and a 4-fold increase in apparent K(m) for amylopectin. The circular dichroism analysis indicates that these kinetic changes may not be due to a major conformation change in the protein. These results provide the first evidence that the conserved aspartate and glutamate residues could be involved in the catalysis or substrate binding of SS. PMID- 10869189 TI - Orientation of the tyrosyl D, pheophytin anion, and semiquinone Q(A)(*)(-) radicals in photosystem II determined by high-field electron paramagnetic resonance. AB - The radical forms of two cofactors and an amino acid in the photosystem II (PS II) reaction center were studied by using high-field EPR both in frozen solution and in oriented multilayers. Their orientation with respect to the membrane was determined by using one-dimensionally oriented samples. The ring plane of the stable tyrosyl radical, Y(D)(*), makes an angle of 64 degrees +/- 5 degrees with the membrane plane, and the C-O direction is tilted by 72 degrees +/- 5 degrees in the plane of the radical compared to the membrane plane. The semiquinone, Q(A)(*)(-), generated by chemical reduction in samples lacking the non-heme iron, has its ring plane at an angle of 72 degrees +/- 5 degrees to the membrane plane, and the O-O axis is tilted by 21 degrees +/- 5 degrees in the plane of the quinone compared to the membrane plane. This orientation is similar to that of Q(A)(*)(-) in purple bacteria reaction centers except for the tilt angle which is slightly bigger. The pheophytin anion was generated by photoaccumulation under reducing conditions. Its ring plane is almost perpendicular to the membrane with an angle of 70 degrees +/- 5 degrees with respect to the membrane plane. This is very similar to the orientation of the pheophytin in purple bacteria reaction centers. The position of the g tensor with respect to the molecule is tentatively assigned for the anion radical on the basis of this comparison. In this work, the treatment of orientation data from EPR spectroscopy applied to one-dimensionally oriented multilayers is examined in detail, and improvements over previous approaches are given. PMID- 10869190 TI - Thermodynamic, spectroscopic, and equilibrium binding studies of DNA sequence context effects in four 40 base pair deoxyoligonucleotides. AB - Effects of different end sequences on melting, circular dichroism spectra (CD), and enzyme binding properties were investigated for four 40 base pair, non-self complementary duplex DNA oligomers. The center sequences of these oligoduplexes have either of two 22 base pair modules flanked on both sides by sequences differing in AT content. Temperature-induced melting transitions monitored by differential scanning calorimetry (DSC) and ultraviolet absorbance were measured for the six duplexes in buffered 115 mM Na(+) solutions. Values of the melting transition enthalpy, DeltaH(cal), and entropy, DeltaS(cal), were obtained directly from DSC experiments. Melting transition parameters, DeltaH(vH) and DeltaS(vH), were also estimated from a van't Hoff analysis of optical melting curves collected as a function of DNA concentration, assuming that the melting transition is two-state. Melting free energies (20 degrees C) evaluated from DSC melting experiments on the four duplex DNAs ranged from -52.2 to -77.5 kcal/mol. Free energies based on the van't Hoff analysis were -37.9 to -58.8 kcal/mol. Although the values are different, trends in the melting free energies of the four duplex DNAs as a function of sequence were identical in both DSC and optical analyses. Subject to several assumptions, values for the initiation free energy were estimated for each duplex, defined as DeltaG(int) = DeltaG(cal) - DeltaG(pred), where DeltaG(cal) is the experimental free energy at 20 degrees C determined from the experimentially measured values of the transition enthalpy, DeltaH(cal), and entropy, DeltaS(cal). The predicted free energy of the sequence, DeltaG(pred)(20 degrees C), is obtained using published nearest-neighbor sequence stability values. For three of the four duplexes, values of DeltaG(int) are essentially nil. In contrast, the duplex with 81.8% GC has a considerably higher estimate of DeltaG(int) = 7.1 kcal/mol. The CD spectra for the six duplexes collected over the wavelength range from 200 to 320 nm are also sequence dependent. Factor analysis of the CD spectra by singular value decomposition revealed that the experimental CD spectra could be reconstructed from linear combinations of two minor and one major subspectra. Changes in the coefficients of the major subspectrum for different sequences reflect incremental sequence dependent variations of the CD spectra. Equilibrium binding by BamHI restriction endonuclease to the 40 base pair DNAs whose central eight base pairs contain the recognition sequence for BamHI restriction enzyme bounded by A.T base pairs, 5'-A GGATCC-A-3' was investigated. Binding assays were performed by titering BamHI against a constant concentration of each of the duplex DNA substrates, in the absence of Mg(2+), followed by analysis by gel retardation. Under the conditions employed, the enzyme binds but does not cleave the DNAs. Results of the assays revealed two binding modes with retarded gel mobilities. Binding isotherms for the fraction of bound DNA species versus enzyme concentration for each binding mode were constructed and analyzed with a simple two-step equilibrium binding model. This analysis provided semiquantitative estimates on the equilibrium binding constants for BamHI to the four DNAs. Values obtained for the binding constants varied only 7-fold and ranged from 6 x 10(-)(8) to 42 x 10(-)(8) M, with binding free energies from -8.6 to -9.7 (+/- 0.2) kcal/mol depending on the sequence that flanks the enzyme binding site. Unlike what was found earlier in binding studies of the 22 base pair duplexes that constitute the core modules of the present 40-mers [Riccelli, P. V., Vallone, P. M., Kashin, I., Faldasz, B. D., Lane, M. J., and Benight, A. S. (1999) Biochemistry 38, 11197-11208], no obvious relationship between binding and stability was found for these longer DNAs. Apparently, effects of flanking sequence stability on restriction enzyme binding may only be measurable in very short duplex deoxyoligonucl PMID- 10869191 TI - Anthraquinones and betaenone derivatives from the sponge-associated fungus Microsphaeropsis species: novel inhibitors of protein kinases. AB - An undescribed fungus of the genus Microsphaeropsis, isolated from the Mediterranean sponge Aplysina aerophoba, produces two new betaenone derivatives (1, 2) and three new 1,3,6, 8-tetrahydroxyanthraquinone congeners (5-7). The structures of the compounds were established on the basis of NMR spectroscopic and mass spectrometric data and by CD spectroscopy. This is the first report wherein the (1)H and (13)C NMR data of the betaenone congeners are fully and unambiguously assigned on the basis of two-dimensional NMR spectroscopy. Furthermore, we describe the first elucidation of the absolute configuration of 1 (2'-anthraquinonyl)ethanols such as 5 and 6, by quantum chemical calculation of their circular dichroism (CD) and comparison with experimentally measured spectra. Moreover, it was shown that compounds 1, 5, 6, and 7 are inhibitors of PKC-epsilon, CDK4, and EGF receptor tyrosine kinases. PMID- 10869192 TI - New alkaloids from Annona purpurea. AB - Three new alkaloids, promucosine (1), romucosine F (2), and romucosine G (3), along with 28 known compounds, were isolated from the MeOH extract of stems of Annona purpurea. The structures of 1-3 were determined on the basis of spectral data and chemical evidence. PMID- 10869193 TI - Galloylglucosides from berries of Pimenta dioica. AB - Three new galloylglucosides, (4S)-alpha-terpineol 8-O-beta-D-(6-O galloyl)glucopyranoside (1); (4R)-alpha-terpineol 8-O-beta-D-(6-O galloyl)glucopyranoside (2), and 3-(4-hydroxy-3-methoxyphenyl)propane-1,2-diol 2 O-beta-D-(6-O-galloyl)glucopyranoside (3), were isolated from the berries of Pimenta dioica together with three known compounds, gallic acid (4), pimentol (5), and eugenol 4-O-beta-D-(6-O-galloyl)glucopyranoside (6). The structures of 1 3 were elucidated on the basis of MS and NMR spectral data and enzymatic hydrolysis. These galloylglucosides (1-3, 5, and 6) showed radical-scavenging activity nearly equivalent to that of gallic acid (4) against 1,1-diphenyl-2 picrylhydrazyl radical. PMID- 10869194 TI - Inhibition of phospholipase cgamma1 and cancer cell proliferation by triterpene esters from Uncaria rhynchophylla. AB - Investigation of the hooks of Uncaria rhynchophylla resulted in isolation of six phospholipase Cgamma1 (PLCgamma1) inhibitors (1-6). The structures of these compounds were elucidated as pentacyclic triterpene esters by spectroscopic and chemical analysis. Three of them, namely uncarinic acids C (1), D (2), and E (3), are newly reported as natural products. All the compounds showed dose-dependent inhibitory activities against PLCgamma1 in vitro with IC(50) values of 9.5-44.6 microM and inhibited the proliferation of human cancer cells with IC(50) values of 0.5-6.5 microg/mL. PMID- 10869195 TI - Phytotoxic compounds from the new coprophilous fungus guanomyces polythrix. AB - Bioactivity-directed fractionation of the fermentation broth and mycelium of the coprophilous fungus Guanomyces polythrix led to the isolation of several phytotoxic compounds, including five new naphthopyranone derivatives (1-5). In addition, rubrofusarin B, emodin, citrinin, and 4-hydroxybenzoic acid methyl ester were obtained. The structures of the new compounds were established by spectral and chiroptical methods. The isolates caused significant inhibition of radicle growth of two weed seedlings (Amaranthus hypochondriacus and Echinochloa crusgalli) and interacted with both spinach and bovine brain calmodulins. PMID- 10869196 TI - Alkaloids of Andrachne aspera. AB - Two new 2,6-disubstituted piperidine alkaloids andrachcinine (1) and andrachcinidine (5) have been isolated from Andrachne aspera along with andrachamine and andrachcine (2). The absolute configurations of 1, 2, and 5 were established. (+)-Allosedridine and the new alkaloids (-)-8-epi-8-ethylnorlobelol I (4) and (-)-8-epihalosaline (7) were also identified as constituents of A. aspera. Structures were determined by MS and NMR techniques and by chemical conversions. PMID- 10869197 TI - Nematicidal constituents of the aerial parts of Lantana camara. AB - Two new constituents, lantanoside (1) and lantanone (2), and the known compounds linaroside (3) and camarinic acid (4) were isolated from the aerial parts of Lantana camara. Compounds 1, 3, and 4 were tested for nematicidal activity against root-knot nematode Meloidogyne incognita and showed 90, 85, and 100% mortality, respectively, at 1.0% concentration. The results were comparable to those obtained with the conventional nematicide furadan (100% mortality at 1.0% concentration). Structures of the new compounds were elucidated by spectroscopic and chemical techniques. PMID- 10869198 TI - Antifungal substances against pathogenic fungi, talaroconvolutins, from talaromyces convolutus. AB - The dichloromethane extract of Talaromyces convolutus cultivated on barley exhibited antifungal activity against Candida albicans. In the course of a search for the active compounds, four new tetramic acid derivatives, talaroconvolutins A (1), B (2), C (3), and D (4), were isolated along with ZG-1494alpha (5), and mitorubrin derivatives. The structures of talaroconvolutins A-D (1-4) were established on the basis of spectroscopic and chemical investigations and chemical correlations. The antifungal activity of the talaroconvolutins against the pathogenic fungi Aspergillus fumigates, Aspergillus niger, C. albicans, and Cryptococcus neoformans was determined. PMID- 10869199 TI - Insecticidal and mutagenic evaluation of two annonaceous acetogenins. AB - Annonaceous acetogenins represent a new class of bioactive compounds whose primary mode of action is the inhibition of NADH-ubiquinone oxidoreductase. Given the potential pesticidal use of such a class of compounds, we have further evaluated the antifeedant and insecticidal effects of squamocin and annonacin, two annonaceous acetogenins, on Spodoptera littoralis, Leptinotarsa decemlineata, and Myzus persicae. Additionally, to partially assess their environmental risk, we have also tested their mutagenicity in Salmonella typhimurium strains TA98, TA100, and TA102 in the presence and absence of a metabolic activation system. Among the test compounds, annonacin showed antifeedant effects on L. decemlineata, while squamocin was toxic to L. decemlineata and M. persicae. Neither acetogenin was mutagenic, although both were toxic in the absence of a metabolic activation system. We compared these results with those obtained with rotenone, a well-known respiratory inhibitor that was highly toxic to L. decemlineata and M. persicae and showed no mutagenicity/toxicity in the S. typhimurium strains tested up to a concentration of 1000 microg per plate. PMID- 10869200 TI - Ribofuranosyl triazolone: a natural product herbicide with activity on adenylosuccinate synthetase following phosphorylation. AB - 2,4-Dihydro-4-(beta-D-ribofuranosyl)-1,2,4(3H)-triazol-3-one (2) was identified as the principal phytotoxic component of a fermentation broth derived from an Actinomadura. The compound is a new natural product, but known by synthesis. Broad-spectrum herbicidal activity was demonstrated in greenhouse tests. Metabolite reversal studies suggested the target site was adenylosuccinate synthetase, which was confirmed by direct measurement of the activity of the 5' phosphorylated derivative on the isolated enzyme. PMID- 10869201 TI - Polyoxygenated bipyridine, pyrrolylpyridine, and bipyrrole alkaloids from Speranskia tuberculata. AB - Five novel polyoxygenated alkaloids, speranculatines A-C (3-5), speranskilatine A (6), and speranberculatine A (7), have been isolated from Speranskia tuberculata. Compounds 3-5, 6, and 7, have bipyridine, pyrrolylpyridine, and bipyrrole skeletons, respectively. This is the first time that these three alkaloid structural types have been reported. The structures of 3-7 were elucidated by spectroscopic methods, including 2D NMR techniques and X-ray crystallographic analysis. PMID- 10869202 TI - Absolute stereochemistry of gastric antisecretory compound P371A1 and its congener P371A2 from streptomyces species P371. AB - Absolute configurations of the gastric antisecretory compound P371A1 (1) and its congener P371A2 (2) from Streptomyces sp. P371 were determined on the basis of identification of the methyl glycosides 9, 10, and 11 obtained by the acid degradation of 1 and 2, as well as application of the modified Mosher method to the P371A2 C-glycoside MTPA esters 7 and 8 and observation of the excitation chiralities in the P371A2 C-glycoside benzoate derivatives 5 and 6. PMID- 10869203 TI - Antineoplastic agents 430. Isolation and structure of cribrostatins 3, 4, and 5 from the republic of maldives cribrochalina species. AB - Continued investigation of cancer-cell growth-inhibitory constituents of the blue marine sponge Cribrochalina sp. has led to discovery of cribrostatins 3 (4a), 4 (5), and 5 (4b) in 10(-5) to 10(-7) % of the wet weight. The structure of cribrostatin 3 (4a) was determined by results of high field (500 MHz) (1)H and (13)C NMR and HRMS interpretations. The same general approach to the structures of cribrostatins 4 (5) and 5 (4b) was completed by X-ray crystal structure determinations. Cribrostatins 3, 4, and 5 provided significant cancer cell line inhibitory activities. Cribrostatins 1 and 2(2) and the newly isolated cribrostatins 3-5 displayed antibacterial and/or antifungal activities. PMID- 10869204 TI - New lipids from the tunicate Cystodytes cf. dellechiajei, as PLA2 inhibitors. AB - Cystodytes cf. dellechiajei collected off Djerba furnished new lipids, sphingosines 1, as inhibitors of phospholipase A2, along with inactive homologous ceramides 2. Structures were determined by spectroscopic methods and chemical transformations. PMID- 10869205 TI - The application of HPLC with on-line coupled UV/MS-biochemical detection for isolation of an acetylcholinesterase inhibitor from narcissus 'Sir Winston Churchill'. AB - An HPLC with on-line coupled UV/MS-biochemical detection method for acetylcholinesterase (AChE) inhibitors in natural sources has been developed. The potential of this method is shown by the isolation of a new AChE inhibitor from the alcoholic extract of Narcissus 'Sir Winston Churchill'. Combining a prefractionation technique using centrifugal partition chromatography with the on line HPLC-UV/MS-biochemical detection resulted in the isolation of the active compound that was identified as ungiminorine. This alkaloid shows a mild inhibitory effect on AChE. PMID- 10869206 TI - Three novel monotetrahydrofuran annonaceous acetogenins from Annona montana. AB - Three new monotetrahydrofuran (mono-THF) acetogenins, anmontanins A-C (1-3) were isolated from the leaves of Annona montana. Anmontanin C (3) is the first stereochemically pure acetogenin reported to have a gamma-hydroxymethyl-gamma lactone moiety. Two known mono-THF acetogenins, murisolin and annonacin, were isolated from the seeds of the plant. The structures of compounds 1-3 were elucidated by spectral methods and chemical derivatization. PMID- 10869207 TI - New taxonomically significant sesquiterpenoids from Leontodon autumnalis. AB - The methanolic extract of subaerial parts of Leontodon autumnalis afforded four new and two known sesquiterpenoids of the guaiane type. The known compounds were identified by means of (1)H and (13)C NMR spectroscopy as crepidiaside A (1) and B (2). The structures of the new compounds were determined by extensive 1D and 2D NMR experiments as 15-glucopyranosyloxy-2-oxo-guaia-3,11(13)-dien-1alp ha,5alpha, 6beta,7alpha,10alphaH-12,6-olide (3); 15-glucopyranosyloxy-2-oxo-guai-3-en 1alpha,5alpha+ ++,6beta,7alpha, 10alpha,11betaH-12,6-olide (4); 15-hydroxy-2-oxo guai-3-en-1alpha, 5alpha,6beta,7alpha,10alpha,11betaH-12,6-+ ++olide (5); and 15 glucopyranosyloxy-2-oxo-guaia-3,11(13)-dien-1bet a,5alpha,6beta, 7alpha,10alphaH 12,6-olide (6), respectively. HPLC-DAD and HPLC-MS analyses of crude extracts of subaerial parts of 25 different taxa of the genus Leontodon revealed that compounds 1 and 2 occur in all investigated members of the section Oporinia (L. autumnalis, L. croceus, L. helveticus, L. montaniformis, L. montanus, L. pyrenaicus, and L. rilaensis) and in L. duboisii from the section Kalbfussia. Compounds 1-6 are detectable neither in other investigated taxa of Kalbfussia (L. cichoraceus, L. muelleri, and L. palisae) nor in any members of the subgenus Leontodon. Compounds 3-5 occur in high amounts only in L. croceus and L. pyrenaicus and in samples of L. autumnalis from northwestern Europe. In other members of the section Oporinia, in L. duboisii as well as in samples of L. autumnalis from the Pyrenees, the Alps, the Carpathians, and southern Central Europe, these substances occur only in trace amounts; in L. montanus and its closest relatives, compounds 3-5 are not detectable at all. Compound 6 is only detectable in samples of L. autumnalis, L. helveticus, L. pyrenaicus, L. rilaensis, and L. duboisii. PMID- 10869208 TI - Potential antitumor-promoting diterpenoids from the stem bark of Picea glehni. AB - A novel rearranged labdane-type diterpenoid, 19(4-->3)abeo-8alpha, 13(S) epoxylabda-4(18),14-diene (1), and two new abietane-type diterpenoids, 19-nor abieta-4(18),8,11,13-tetraen-7-one (2) and 12-hydroxydehydroabietic acid (3) were isolated from the stem bark of Picea glehni, together with seven known diterpenoids-13-epimanoyl oxide (4), dehydroabietic acid (5), (11E)-14, 15-bisnor 8alpha-hydroxy-11-labden-13-one (6), abieta-8,11, 13-trien-7-one (7), 9alpha,13alpha-epidioxyabiet-8(14)-en-18-oic acid (8), 9,10alpha-epoxy-9,10-seco abieta-8,11,13-trien-18-oic acid (9), and methyl 15-hydroxy-7-oxo-dehydroabietate (10). Compounds 5-8 and 10 showed potent inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by the tumor promoter 12-O tetradecanoylphorbol 13-acetate. PMID- 10869209 TI - Clathrins A-C: metabolites from a Southern Australian marine sponge, clathria species. AB - A Clathria sp. collected in the Great Australian Bight has yielded the novel metabolites clathrins A (6), B (7), and C (8). Structures were assigned to clathrins A-C on the basis of spectroscopic analysis. Clathrin A (6) represents a plausible biosynthetic intermediate that provides an inferred link between marine sesquiterpene/benzenoids and mixed terpene/shikimate biosynthesis. PMID- 10869210 TI - A new bioactive sesterterpene and antiplasmodial alkaloids from the marine sponge hyrtios cf. erecta. AB - From the CH(2)Cl(2) extract of the sponge Hyrtios cf. erecta, collected from Fiji, two new sesterterpenes, 1 and 2, and the known compounds isodehydroluffariellolide (3), homofascaplysin A (4), and fascaplysin (5) were isolated. The structures of 1-5 were established employing 1D and 2D NMR spectroscopy and mass spectrometry. All NMR resonances of fascaplysin (5) have been unambiguously assigned. Evaluation of the biological activity of the extracts and pure compounds toward Plasmodium falciparum, Trypanosoma brucei subsp. rhodesiense, Trypanosoma cruzi, hepatitis A virus (HAV), several other microbial targets, and HIV-1-RT and p56(lck) tyrosine kinase revealed new activities for homofascaplysin (4) and fascaplysin (5), both being potently active in vitro against P. falciparum. PMID- 10869211 TI - Cycloshermilamine D, a new pyridoacridine from the marine tunicate cystodytes violatinctus. AB - A new alkaloid, cycloshermilamine D, was isolated from the marine tunicate Cystodytes violatinctus, and its structure (1), which is closely related to shermilamine D, was established mainly on the basis of NMR spectroscopic data. PMID- 10869212 TI - Polycitone B and prepolycitrin A: two novel alkaloids from the marine ascidian Polycitor africanus. AB - Two novel compounds, polycitone B (4) and prepolycitrin A (3), were isolated from the marine ascidian Polycitor africanus. The structures of these new dibromotyrosine metabolites were established mainly on the basis of NMR spectroscopic data. Isolation of compound 3 supports the earlier suggestion that it is the bioprecursor of polycitrin A (2). PMID- 10869213 TI - Higher order and substituent chemical shift effects in the proton NMR of glycosides. AB - The full analysis of (1)H NMR spin systems and the charting of substituent chemical shifts offer great potential in the structure elucidation of glycosides. Due to the chiral nature of sugar residues, asymmetric induction causes diastereotopic shifts of aglycon resonance, while on the other hand aromatic substituent-induced shifts affect the sugar portion of the molecule. Therefore, it is possible to deduce critical structural information, such as the site of sugar linkage in bisdesmosides and oligoglycosides, through the analysis of exact chemical shifts, J patterns, and signal multiplicity. The potential of the method is demonstrated for three kaempferol bisdesmosides (1-3) by establishing the shift characteristics for 3-O-, 7-O-, and 6' '-O-glycosidation. Spectral simulation and soft-pulse 1D NMR turn out to be indispensable tools in the course of refined signal and structure assignment. PMID- 10869214 TI - NMR spectroscopy, X-ray crystallographic, and molecular modeling studies on a new pyranone from Haloxylon salicornicum. AB - A new pyranone, 5-hydroxy-3-methoxy-4H-pyran-4-one (1), was isolated from the aerial parts of the desert shrub Haloxylon salicornicum. The structure was elucidated by X-ray structural analysis, NMR spectroscopy, and mass spectrometry. The monoacetate was also prepared, and molecular modeling studies and full NMR data were recorded. PMID- 10869215 TI - 7alpha-hydroxytheonellasterol, a cytotoxic 4-methylene sterol from the Philippines sponge Theonella swinhoei. AB - A new 4-methylene sterol, 7alpha-hydroxytheonellasterol (3), was isolated from the Philippines sponge Theonella swinhoei, along with the known compounds theonellasterol (1) and bistheonellide A. The structure of 3 was elucidated by interpretation of its spectroscopic data, which were compared with those of 1. PMID- 10869216 TI - A cytotoxic lobane diterpene from the formosan soft coral Sinularia inelegans. AB - A new cytotoxic lobane diterpene, ineleganene (1), was isolated from the Formosan soft coral Sinularia inelegans. The structure of compound 1 was determined by 1D and 2D spectral analysis. PMID- 10869217 TI - A new acylated flavonol triglycoside from Carrichtera annua. AB - A new acylated flavonol triglycoside, quercetin 3-O-[(6-feruloyl-beta glucopyranosyl)-(1-->2)-beta-arabinopyran oside] -7-O-beta-glucopyranoside (1), as well as the known flavonoid quercetin, were isolated from the whole plants of Carrichtera annua. The structure of 1 was established by UV, MS, and 1D and 2D NMR spectroscopy, including DEPT, DQF-COSY, TOCSY, HSQC, HSQC-TOCSY, and HMBC experiments. PMID- 10869218 TI - New diterpenic altrosides of the fungus Acremonium striatisporum isolated from a sea cucumber. AB - Two new diterpenic glycosides, virescenosides M (1) and N (2), have been isolated from a marine strain of Acremonium striatisporum KMM 4401 associated with the holothurian Eupentacta fraudatrix. Their structures were determined on the basis of MS and NMR data as beta-D-altropyranosido-19-7-oxo-isopimara-8,15-diene-2alp ha, 3beta-diol (1) and beta-D-altropyranosido-19-isopimara-7, 15-diene 2alpha,3beta,6beta-triol (2). Three other altrosides (3-5), identified as virescenosides A, B, and C from the terrestrial strain Acremonium luzulae, were also isolated. The cytotoxic activity of the virescenosides was examined. PMID- 10869219 TI - Rubupungenosides A and B, two novel triterpenoid saponin dimers from the aerial parts of Rubus pungens. AB - Two new triterpenoid saponin dimers, rubupungenosides A (1) and B (2), were isolated in their methylated forms 1a and 2a, respectively, from an ethanol extract of the aerial parts of Rubus pungens. The structures of 1a and 2a were established on the basis of spectroscopic and chemical methods. PMID- 10869220 TI - Indicanine A, a new 3-phenylcoumarin from root bark of Erythrina indica. AB - A new 3-phenylcoumarin, indicanine A (1), has been isolated from the root bark of the African medicinal plant Erythrina indica, together with three known compounds, robustic acid (2), daidzein, and 8-prenyldaidzein. The structure of the new compound was characterized, as 4-hydroxy-5-methoxy-3-(4'-methoxyphenyl) 2" -(1-methylethenyl)dihydrofurano[4",5":6,7]coumarin by means of extensive spectroscopic analyses. The compounds were found to be devoid of in vitro antibacterial activity. PMID- 10869221 TI - Characterization of two structural forms of otonecine-type pyrrolizidine alkaloids from Ligularia hodgsonii by NMR spectroscopy. AB - Clivorine (1) and ligularine (2), two hepatotoxic otonecine-type pyrrolizidine alkaloids isolated from Ligularia hodgsonii, an antitussive traditional Chinese medicine, were investigated in CDCl(3) and D(2)O by various NMR techniques to delineate why this type of alkaloid displays uncharacteristic solubility properties by dissolving in both nonpolar organic and aqueous solutions. The results demonstrated that both alkaloids exist in a non-ionized form in CDCl(3), but in an ionized form in D(2)O, suggesting that this unique dual solubility may play a role in the intoxication resultant from consumption of water extracts of herbs, including herbal teas, containing otonecine-type pyrrolizidine alkaloids. PMID- 10869222 TI - Tupichigenin A, a new steroidal sapogenin from Tupistra chinensis. AB - From the underground parts of Tupistra chinensis, a novel polyhydroxylated spirostanol sapogenin, tupichigenin A [(20S, 22R)-spirost-25(27)-ene 1beta,2beta,3beta,5beta- tetraol] (1), was isolated and determined structurally on the basis of spectroscopic methods. Also isolated was the known steroidal sapogenin (20S, 22R)-spirost-25(27)-ene-1beta,2beta,3beta,4beta, 5beta, 7alpha hexaol-6-one (2). PMID- 10869223 TI - Stereochemistry of caracurine V. AB - The 3D structure of the Strychnos alkaloid caracurine V was determined by means of NMR spectroscopy and semiempirical calculations. The previously unknown absolute configuration in the central eight-membered ring was assigned as (16R, 16'R, 17R, and 17'R). PMID- 10869224 TI - Simple synthesis of benzofuranoid neolignans from Myristica fragrans. AB - The total synthesis of four neolignans-fragnasols A (1), B (2), and C (3) and dehydrodiisoeugenol (4)-starting from the readily available phenol derivative isoeugenol (5) was accomplished. The key step of the synthesis of these natural products is a novel type of dimerization of 5 into 4 with iodobenzene diacetate. PMID- 10869225 TI - Two novel glucodienoid alkaloids from Erythrina latissima seeds. AB - The structures of two novel glycodienoid alkaloids isolated from Erythrina latissima seeds have been established as (+)-16beta-D-glucoerysopine (1) and (+) 15beta-D-glucoerysopine (2). NOE measurements established that 1 and 2 are positional isomers. Their structures were elucidated from their 1D and 2D homonuclear and heteronuclear NMR spectroscopic data. In addition, seven known tetracyclic dienoid alkaloids were isolated and identified as (+)-erysovine, (+) erysodine, (+)-erysotrine, (+)-erythraline, (+)-beta-D-glucoerysodine, (+)-8 oxoerythraline, and (+)-erysotramidine. PMID- 10869226 TI - Calafianin, a bromotyrosine derivative from the marine sponge Aplysina gerardogreeni. AB - Calafianin (1) and two known compounds, aerothionin and (3, 5-dibromo-2-hydroxy-4 methoxyphenyl)acetic acid, were isolated from the marine sponge Aplysina gerardogreeni. The structure of 1 was determined by NMR analysis and mass spectrometry. PMID- 10869227 TI - Application of the BIRD sandwich for the rapid and accurate determination of (1)H (1)H NMR coupling constants in higher order spin systems. AB - A method is presented that allows for the convenient and reliable determination of (1)H-(1)H NMR coupling constants in higher order or symmetrically coupled spin systems. The method can be applied on any programmable FT-NMR spectrometer and is demonstrated here on micromole quantities of sample in a standard 5-mm NMR tube. PMID- 10869228 TI - Salvimultine, a new noricetexane diterpene from the roots of salvia multicaulis. AB - A new noricetexane diterpene, salvimultine, has been isolated from the polar fractions of Salvia multicaulis. Its structure was established as 1(10)-seco 2(10)-cyclo-icetexane by using 1D and 2D NMR spectral methods, including COSY, HETCOR, COLOC, and NOESY experiments as well as HRMS. PMID- 10869229 TI - Lupene-type triterpenes from Periploca aphylla. AB - Two new lupane derivatives, 3beta,6alpha-dihydroxylup-20(29)-ene (1) and 6alpha hydroxylup-20(29)-en-3beta-octadecanoate (2), have been isolated from the stems of Periploca aphylla, in addition to beta-sitosterol and lupeol. The structures of 1 and 2 were determined by spectral and chemical methods. Compound 1 showed strong inhibition of alpha-glucosidase type VI and a moderate antibacterial activity. PMID- 10869230 TI - Cytotoxic cembrenolide diterpenes from the formosan soft coral lobophytum crassum. AB - A new cytotoxic cembrenolide diterpene, lobocrassolide (1), and a known cytotoxic cembrenolide, lobohedleolide (2), were isolated from the Formosan soft coral Lobophytum crassum. The structure of compound 1 was determined by 1D and 2D spectral analysis. PMID- 10869231 TI - Coruscol A, a new metabolite from the marine-derived fungus penicillium species. AB - A new metabolite possessing a 1,3-dioxane ring, coruscol A (1), was isolated from the mycelium of the fungus Penicillium sp., which was separated from the Okinawan marine bivalve Mytilus coruscus, and the structure was elucidated by spectroscopic data. PMID- 10869232 TI - Constituents of Nepeta caesarea. AB - The acetone extract of Nepeta caesarea yielded four new nepetalic acid derivatives, 3'alpha-[beta-sitosteryl-3beta-oxy]dihydronepetalactone (1), 3'beta [5alpha-stigmast-7-ene-3beta-oxy]dihydronepetalact one (2), 3'alpha-[olean-12-ene 28-oyl-3beta-oxy]dihydronepetalactone (3), and 3'alpha-[lup-20(29)-ene-28-ol 3beta-oxy]dihydronepetalactone (4). The structures were elucidated by NMR and MS techniques. PMID- 10869233 TI - An optimized set of human telomere clones for studying telomere integrity and architecture. AB - Telomere-specific clones are a valuable resource for the characterization of chromosomal rearrangements. We previously reported a first-generation set of human telomere probes consisting of 34 genomic clones, which were a known distance from the end of the chromosome ( approximately 300 kb), and 7 clones corresponding to the most distal markers on the integrated genetic/physical map (1p, 5p, 6p, 9p, 12p, 15q, and 20q). Subsequently, this resource has been optimized and completed: the size of the genomic clones has been expanded to a target size of 100-200 kb, which is optimal for use in genome-scanning methodologies, and additional probes for the remaining seven telomeres have been identified. For each clone we give an associated mapped sequence-tagged site and provide distances from the telomere estimated using a combination of fiberFISH, interphase FISH, sequence analysis, and radiation-hybrid mapping. This updated set of telomeric clones is an invaluable resource for clinical diagnosis and represents an important contribution to genetic and physical mapping efforts aimed at telomeric regions. PMID- 10869234 TI - A single genetic origin for the G101W CDKN2A mutation in 20 melanoma-prone families. AB - Germline mutations within the coding region of CDKN2A have been observed in affected members of melanoma-prone families. G101W is the most common CDKN2A missense mutation identified to date. It has been reported in several families from around the world, with a particularly high occurrence in France and Italy. Given the frequency of this mutation, we were interested in determining whether the mutation resulted from a single origin or represented a mutational hotspot in the CDKN2A gene. In addition, given the geographical distribution of the mutation, we examined the date of origination of the mutation and its migratory spread. We examined 10 families from Italy, 4 families from the United States, and 6 families from France with the G101W mutation. The following eight markers were employed for the haplotype analysis: IFNA, D9S736, D9S1749, D9S942, D9S1748, D9S1604, D9S171, and D9S126. Our findings showed no significant evidence for mutational heterogeneity, suggesting that all studied families derived from a single ancestral haplotype on which the mutation arose. Using maximum-likelihood methods, we estimated the mutation to have arisen 97 generations ago (1-LOD-unit support interval 70-133 generations) providing some explanation for the wide geographical spread of this common mutation, particularly in southwestern Europe. The presence of a founder mutation in a defined geographic area can facilitate carrier detection and genetic counseling and can provide an opportunity to study disease penetrance and the effect of environmental factors on the background of a common genetic susceptibility. PMID- 10869235 TI - SNPing away at complex diseases: analysis of single-nucleotide polymorphisms around APOE in Alzheimer disease. AB - There has been great interest in the prospects of using single-nucleotide polymorphisms (SNPs) in the search for complex disease genes, and several initiatives devoted to the identification and mapping of SNPs throughout the human genome are currently underway. However, actual data investigating the use of SNPs for identification of complex disease genes are scarce. To begin to look at issues surrounding the use of SNPs in complex disease studies, we have initiated a collaborative SNP mapping study around APOE, the well-established susceptibility gene for late-onset Alzheimer disease (AD). Sixty SNPs in a 1.5-Mb region surrounding APOE were genotyped in samples of unrelated cases of AD, in controls, and in families with AD. Standard tests were conducted to look for association of SNP alleles with AD, in cases and controls. We also used family based association analyses, including recently developed methods to look for haplotype association. Evidence of association (P80 g/day) alcohol intake on the histological and clinical progression of HCV infection and their associated risk of hepatic cancer in a group of Japanese patients. A number of other variables were assessed to evaluate their impact on disease progression. We recruited 120 patients with HCV infection and categorized them into four groups, based on alcohol consumption pattern. All clinical and biochemical profiles were collected from recorded files. Liver biopsies were analysed for the degree of fibrosis, presence of cirrhosis and histological activity of necroinflammation. Hepatic tumours were detected by the follow-up imaging analysis. There was no difference in the age, length of exposure to HCV infection and HCV RNA serum levels in the alcohol and alcohol-free groups. The histological grading of necroinflammation, serum levels of alanine aminotransferase and HCV RNA did not have any correlation with each other in the alcohol and alcohol-free group. There was a 1.5-2. 5-fold greater risk of liver cirrhosis and hepatocellular carcinoma in the alcohol intake group compared to the alcohol-free group. Kruskal-Wallis analysis among four groups demonstrated a significant transition to fibrosis (P < 0.05) for alcoholics with HCV infection. The increased risk of liver cancer in the alcohol group is independent of size and growth of tumours. The clinical manifestations of gastro-oesophageal variceal bleeding, ascites, and encephalopathy were also higher in the alcohol intake group. Alcohol consumption is an important risk factor in the histological and clinical progression of HCV infection and has no relation with HCV replication. Chronic HCV carriers should avoid excessive alcohol intake to reduce the acceleration of liver disease and risk of liver cancer. PMID- 10869252 TI - Frequency and Markov chain analysis of the amino-acid sequence of human alcohol dehydrogenase alpha-chain. AB - The amino-acid sequences of human alcohol dehydrogenase alpha-chain (ADH1) were analysed according to two-, three- and four-amino-acid sequences. The measured frequencies and probabilities were compared with the predicted frequencies and probabilities. Of 373 two-amino-acid sequences in the ADH1, 92 (24.665%) and 32 (8.579%) sequences can be explained by the predicted frequencies and probabilities according to a purely random mechanism. Of 191 non-appearing two amino-acid sequences in the ADH1, 119 (62.304%) and 52 (27.225%) sequences can be explained by the predicted frequencies and probabilities according to a purely random mechanism. Of 373 measured first-order Markov transition probabilities for the second amino acid in two-amino-acid sequences, three (0.804%) probabilities match the predicted conditional probabilities and therefore can be explained by a purely random mechanism. No more-than-two-amino-acid sequences can be explained by a purely random mechanism. PMID- 10869251 TI - Drinking habits of subjects with hepatitis C virus-related chronic liver disease: prevalence and effect on clinical, virological and pathological aspects. AB - Alcohol changes the progression of hepatitis C virus (HCV)-related chronic liver disease and may affect the outcome of interferon therapy. The ethanol intake of 245 patients with biopsy-proven chronic hepatitis C with or without cirrhosis, its interaction with laboratory and histological parameters common to alcohol and HCV-mediated liver damage, and its effects on therapy were evaluated. The results show that 60-70% of subjects regularly consumed alcohol (median intake >40 g/day in about 30%). Less than 50% stopped drinking after being diagnosed as having liver disease. Ethanol intake affected: fibrosis, especially in women, HCV RNA levels, which were significantly lower in abstainers than in drinkers (0.6 +/- 0.3 vs 6.9 +/- 5.9 Eq/ml x10(6); P < 0.01), and response to interferon therapy. The number of responders decreased as ethanol intake increased. There were less abstainers than drinkers among non-responders (10.7% vs 63.1% respectively; P < 0.001). Data indicate that alcohol will induce and worsen liver damage and, in subjects with chronic liver disease who continue to drink, adversely affect their response to treatment. PMID- 10869253 TI - Stressors and alcohol consumption. AB - The objective of this study was to examine the relationship between negative life events and chronic stressors and drinking behaviour. Data suggested that some life events (getting divorced) and some chronic stressors (financial difficulties, unfavourable marital status, and unfavourable employment status) were positively related to abstinence among men and women. Furthermore, some life events (being a victim of a crime, decrease in financial position, divorce or reporting two or more life events) were positively associated with heavy drinking among men. Chronic stressors, such as unfavourable marital status and unfavourable employment status, were also related to heavy drinking among both men and women. Results presented here suggest that people under stressful conditions are more likely to either abstain or drink heavily rather than to drink lightly or moderately. PMID- 10869254 TI - Determination of ethyl glucuronide in hair samples. PMID- 10869255 TI - June 27, 2000 PMID- 10869256 TI - Myocardial cell protection : a challenging time for action and a challenging time for clinical research. PMID- 10869257 TI - Positional genomic analysis identifies the beta(2)-adrenergic receptor gene as a susceptibility locus for human hypertension. AB - BACKGROUND: -After genome-wide linkage analyses of blood pressure levels, we resequenced 5 positional candidate genes in a linkage region on chromosome 5 and genotyped selected variants in several family samples from Rochester, Minn. METHODS AND RESULTS: In a sample of 55 pedigrees containing >/=1 sibling-pair(s) discordant for systolic blood pressure, polymorphisms within the beta(2) adrenergic receptor gene (Arg16Gly, P=0.009) and the glutathione peroxidase 3 gene (-302G-->A, P=0.037; -623A-->C, P=0.013) were significantly related to blood pressure levels. In a second sample of 298 nuclear families (n=1283 individuals), the Arg16Gly polymorphism was significantly associated with diastolic blood pressure in family-based analyses (P=0.016) and with both diastolic (P=0.009) and mean arterial blood pressure (P=0.038) in analyses of the parental generation only. Neither polymorphism in the glutathione peroxidase 3 gene was associated with blood pressure levels in this sample. An additional 291 families (n=1240 individuals) were added to the nuclear family sample, and the Gln27Glu polymorphism in the beta(2)-adrenergic receptor gene was significantly associated with both systolic (P=0.034) and mean arterial blood pressure (P=0.035) in the parental generation of the combined 589 families. The frequencies of both the Gly16 and Glu27 alleles were higher in hypertensives than in normotensives (0.649 versus 0.604 and 0.490 versus 0.429, respectively), and the odds ratio for the occurrence of hypertension was 1.80 (95% confidence interval, 1.08 to 3.00; P=0. 023) for the Glu27 allele. CONCLUSIONS: The results of this study provide support for further detailed investigations of the mechanistic pathways by which variations in the beta(2)-adrenergic receptor gene may influence blood pressure levels. PMID- 10869258 TI - Monoclonal T-cell proliferation and plaque instability in acute coronary syndromes. AB - BACKGROUND: Unstable angina (UA) is associated with systemic inflammation and with expansion of interferon-gamma-producing T lymphocytes. The cause of T-cell activation and the precise role of activated T cells in plaque instability are not understood. METHODS AND RESULTS: Peripheral blood T cells from 34 patients with stable angina and 34 patients with UA were compared for the distribution of functional T-cell subsets by flow cytometric analysis. Clonality within the T cell compartment was identified by T-cell receptor spectrotyping and subsequent sequencing. Tissue-infiltrating T cells were examined in extracts from coronary arteries containing stable or unstable plaque. The subset of CD4(+)CD28(null) T cells was expanded in patients with UA and infrequent in patients with stable angina (median frequencies: 10.8% versus 1.5%, P<0.001). CD4(+)CD28(null) T cells included a large monoclonal population, with 59 clonotypes isolated from 20 UA patients. T-cell clonotypes from different UA patients used antigen receptors with similar sequences. T-cell receptor sequences derived from monoclonal T-cell populations were detected in the culprit but not in the nonculprit lesion of a patient with fatal myocardial infarction. CONCLUSIONS: UA is associated with the emergence of monoclonal T-cell populations, analogous to monoclonal gammopathy of unknown significance. Shared T-cell receptor sequences in clonotypes of different patients implicate chronic stimulation by a common antigen, for example, persistent infection. The unstable plaque but not the stable plaque is invaded by clonally expanded T cells, suggesting a direct involvement of these lymphocytes in plaque disruption. PMID- 10869259 TI - Antisense to LOX-1 inhibits oxidized LDL-mediated upregulation of monocyte chemoattractant protein-1 and monocyte adhesion to human coronary artery endothelial cells. AB - BACKGROUND: We have recently demonstrated a lectin-like receptor for oxidized (ox)-LDL (LOX-1) in human coronary artery endothelial cells (HCAECs). This receptor is upregulated by ox-LDL. The present study examined the significance of LOX-1 in monocyte adhesion to HCAECs and endothelial injury in response to ox LDL. METHODS AND RESULTS: HCAECs were incubated in the presence of antisense oligodeoxynucleotides to the 5'-coding sequence of the human LOX-1 gene (0.5 microm/L). Basal LOX-1 mRNA and protein were suppressed by antisense LOX-1. Ox LDL-mediated upregulation of LOX-1 was also suppressed by antisense LOX-1. Incubation of HCAECs with ox-LDL (40 microg/mL) for 24 hours markedly increased monocyte chemoattractant protein-1 (MCP-1) mRNA and protein expression as well as monocyte adhesion to HCAECs (P<0.01). After 48 hours of preincubation of HCAECs with antisense LOX-1, ox-LDL-mediated upregulation of MCP-1 and monocyte adhesion to HCAECs both were suppressed (P<0.01), whereas sense LOX-1 had no effect. Whereas antisense or sense LOX-1 alone (both 0.5 nmol/L) did not injure the cells, antisense LOX-1, but not sense LOX-1, reduced ox-LDL-mediated HCAEC injury, determined as LDH release (P<0.01). Activation of mitogen-activated protein kinase (MAPK) may play a critical role in signal transduction in ox-LDL mediated alteration in MCP-1 expression, since antisense LOX-1, but not the sense LOX-1, completely inhibited the ox-LDL-induced MAPK activation. CONCLUSIONS: These observations with the first use of a specific antisense to human LOX-1 mRNA suggest that LOX-1 is a key factor in ox-LDL-mediated monocyte adhesion to HCAECs. PMID- 10869260 TI - Physical activity prevents age-related impairment in nitric oxide availability in elderly athletes. AB - BACKGROUND: Aging is associated with increased cardiovascular risk and endothelial dysfunction. Since exercise can improve endothelium-dependent vasodilation, in the present study we tested whether long-term physical activity could prevent aging-related endothelial dysfunction. METHODS AND RESULTS: In 12 young and elderly (age 26.9+/-2.3 and 62.9+/-5.8 years, respectively) healthy sedentary subjects and 11 young and 14 elderly matched athletes (age 27.5+/-1.9 and 66.4+/-6.1 years, respectively), we studied (with strain-gauge plethysmography) forearm blood flow modifications induced by intrabrachial acetylcholine (0.15, 0.45, 1.5, 4.5, and 15 microg/100 mL per minute), an endothelium-dependent vasodilator, at baseline, during infusion of N(G) monomethyl-L-arginine (L-NMMA) (100 microg/100 mL forearm tissue per minute), a nitric oxide-synthase inhibitor, vitamin C (8 mg/100 mL forearm tissue per minute), an antioxidant, and finally under simultaneous infusion of L-NMMA and vitamin C. The response to sodium nitroprusside (1, 2, and 4 microg/100 mL forearm tissue per minute) was also evaluated. In young athletes and sedentary subgroups, vasodilation to acetylcholine was inhibited by L-NMMA and was not changed by vitamin C. In elderly subjects, vasodilation to acetylcholine was blunted as compared with young subjects in both control subjects and athletes, whereas the response to sodium nitroprusside was similar. Moreover, in elderly athletes, vitamin C did not change the vasodilation to acetylcholine. In contrast, in elderly sedentary subjects, the response to acetylcholine was resistant to L-NMMA. In this subgroup, vitamin C increased the vasodilation to acetylcholine and restored the inhibiting effect of L-NMMA. CONCLUSIONS: These results suggest that regular physical activity can at least in part prevent the age-induced endothelial dysfunction, probably the restoration of nitric oxide availability consequent to prevention of production of oxidative stress. PMID- 10869261 TI - Cardioprotective effects of the Na(+)/H(+) exchange inhibitor cariporide in patients with acute anterior myocardial infarction undergoing direct PTCA. AB - BACKGROUND: Activation of Na(+)/H(+) exchange in myocardial ischemia and/or reperfusion leads to calcium overload and myocardial injury. Experimental studies have shown that Na(+)/H(+) exchange inhibitors can attenuate Ca(2+) influx into cardiomyocytes. We therefore performed a multicenter, randomized, placebo controlled clinical trial to test the hypothesis that inhibition of Na(+)/H(+) exchange limits infarct size and improves myocardial function in patients with acute anterior myocardial infarction (MI) treated with direct PTCA. METHODS AND RESULTS: One hundred patients were randomized to receive placebo (n=51) or a 40 mg intravenous bolus of the Na(+)/H(+) exchange inhibitor cariporide (HOE 642) (n=49) before reperfusion. Global and regional left ventricular functions were analyzed by use of paired contrast left ventriculograms performed before and 21 days after PTCA and myocardial enzymes (ie, creatine kinase ?CK, CK-MB, and LDH) as markers for myocardial tissue injury were evaluated. At follow-up, the ejection fraction was higher (50% versus 40%; P<0.05) and the end-systolic volume was lower (69.0 versus 97.0 mL; P<0.05) in the cariporide group. Significant improvements in some indices of regional wall motion abnormalities were observed, such as the percentage of chords with hypokinesis < -2 SD (P=0.045) and the severity of hypokinesis in the border zone of the infarct region (P=0.052). In addition, CK, CK-MB, or LDH release was significantly reduced in the cariporide patients. CONCLUSIONS: Our findings suggest that inhibition of Na(+)/H(+) exchange by cariporide may attenuate reperfusion injury and thereby improve the recovery from left ventricular dysfunction after MI. PMID- 10869262 TI - Uncertainty principle of signal-averaged electrocardiography. AB - BACKGROUND: Signal-averaged ECG (SAECG) reproducibility is reported to have a component that is independent of residual noise. Methods and Results-In group 1, multiple paired SAECGs were obtained to noise levels of 0.3+/-0.1 and 0.5+/-0.2 microV. For the 0.5- and 0. 3-microV noise recordings, QRS duration (QRSd) was 101.2+/-11.3 and 104.6+/-9.6 ms, respectively (P<0.0001), and the differences in paired QRSd (DeltaQRSd) were normally distributed, with variances of 11.4 and 26.2 ms(2) (P<0.0001). Paired SAECGs were obtained in group 2 patients without and with late potentials; DeltaQRSd variance was 3.3 and 217.9 ms(2) (P<0.0001). In group 3, >/=10 SAECGs were acquired at noise levels of 0.2 to 0.8 microV, in 0.1-microV increments. QRSd increased as noise level decreased. The variance was greater in low-noise (0.2 to 0.4 microV) versus higher-noise (0. 5 to 0.8 microV) recordings. In group 4, SAECGs were analyzed with bidirectional and Bispec filters, with no difference in QRSd between the 2 filters and a normally distributed DeltaQRSd. A computer simulation demonstrated that alterations in the phase relationship of noise to signal results in a normal distribution of signal end points. CONCLUSIONS: Within the acceptable noise range for SAECG, lower noise results in longer QRSd and larger variance, suggesting that more accurate recordings may have less reproducibility. The random timing of noise relative to signal results in the distribution/variance of repeated measurements. Statistical strategies may be used to reduce some of this variance and may enhance the diagnostic utility of SAECG. PMID- 10869263 TI - Prolonged antibiotic prophylaxis after cardiovascular surgery and its effect on surgical site infections and antimicrobial resistance. AB - BACKGROUND: Despite evidence supporting short antibiotic prophylaxis (ABP), it is still common practice to continue ABP for more than 48 hours after coronary artery bypass graft (CABG) surgery. METHODS AND RESULTS: To compare the effect of short (<48 hours) versus prolonged (>48 hours) ABP on surgical site infections (SSIs) and acquired antimicrobial resistance, we conducted an observational 4 year cohort study at a tertiary-care center. An experienced infection control nurse performed prospective surveillance of 2641 patients undergoing CABG surgery. The main exposure was the duration of ABP, and main outcomes were the adjusted rate of SSI and the isolation of cephalosporin-resistant enterobacteriaceae and vancomycin-resistant enterococci (acquired antibiotic resistance). Adjustment for confounding was performed by multivariable modeling. A total of 231 SSIs (8.7%) occurred after a median of 16 days, including 93 chest wound infections (3.5%) and 13 deep-organ-space infections (0. 5%). After 1502 procedures using short ABP, 131 SSIs were recorded, compared with 100 SSIs after 1139 operations with prolonged ABP (crude OR, 1.0; CI, 0.8 to 1.3). After adjustment for possible confounding, prolonged ABP was not associated with a decreased risk of SSI (adjusted OR, 1.2; CI, 0.8 to 1.6) and was correlated with an increased risk of acquired antibiotic resistance (adjusted OR, 1.6; CI, 1.1 to 2.6). CONCLUSIONS: Our findings confirm that continuing ABP beyond 48 hours after CABG surgery is still widespread; however, this practice is ineffective in reducing SSI, increases antimicrobial resistance, and should therefore be avoided. PMID- 10869264 TI - Acute endothelin A receptor blockade causes selective pulmonary vasodilation in patients with chronic heart failure. AB - BACKGROUND: Elevated plasma endothelin-1 (ET-1) levels in patients with chronic heart failure correlate with pulmonary artery pressures and pulmonary vascular resistance. ET(A) receptors on vascular smooth muscle cells mediate pulmonary vascular contraction and hypertrophy. We determined the acute hemodynamic effects of sitaxsentan, a selective ET(A) receptor antagonist, in patients with chronic stable heart failure receiving conventional therapy. METHODS AND RESULTS: This multicenter, double-blind, placebo-controlled trial enrolled 48 patients with chronic New York Heart Association functional class III or IV heart failure (mean left ventricular ejection fraction 21+/-1%) treated with ACE inhibitors and diuretics. Patients with a baseline pulmonary capillary wedge pressure >/=15 mm Hg and a cardiac index 10 seconds), and after resuscitation from prolonged VF (>10 minutes). Short-term differences in percent change in left ventricular +dP/dt(max) (MP -16+/-28%, BP +9.1+/-24%; P=0.03) and left ventricular -dP/dt(max) (MP -37+/-26%, BP -18+/-20%; P=0.05) were present after rescue from brief VF, with BP animals exhibiting less countershock-induced dysfunction. After prolonged VF, the BP group had lower mean defibrillation thresholds (107+/-57 versus 172+/-88 J for MP, P=0.04) and significantly shorter resuscitation times (397+/-73.7 versus 488+/-74.3 seconds for MP, P=0.03). CONCLUSIONS: External defibrillation is more efficacious with BP countershocks than with MP countershocks. The lower defibrillation thresholds and shorter resuscitation times associated with BP waveform defibrillation may improve survival after prolonged VF arrest. PMID- 10869272 TI - Changes in sinus node function in a rabbit model of heart failure with ventricular arrhythmias and sudden death. AB - BACKGROUND: Heart failure is associated with profound changes in the balance of the autonomic nervous system, such as vagal withdrawal and increased catecholamine levels. It is not known whether the intrinsic sinus node function changes during the progression of heart failure. METHODS AND RESULTS: We implanted transmitters for Holter recording in an established rabbit model of heart failure (n=9) and observed changes in sinus cycle length and the occurrence of arrhythmias during the progression of heart failure. The in vitro sinus cycle length and the responses to acetylcholine and norepinephrine in the isolated right atria were analyzed in 12 rabbits with heart failure and in 6 control rabbits. In vivo cycle length increased in some animals and decreased in others. Sudden death occurred in 3 of 9 rabbits. These rabbits had developed a shorter cycle length than the surviving rabbits. Ventricular tachycardias developed in all but 1 rabbit. The in vitro sinus cycle length increased in heart failure. The response to acetylcholine also increased in heart failure, whereas the response to norepinephrine was unchanged. CONCLUSIONS: Changes in intrinsic sinus node function during the progression of heart failure cannot explain the observed decreases in heart rate variability and/or baroreflex sensitivity in this disease, because increased responsiveness to acetylcholine would be expected to cause the opposite. PMID- 10869273 TI - Left ventricular remodeling after myocardial infarction: pathophysiology and therapy. PMID- 10869274 TI - Images in cardiovascular medicine: pseudo-myocardial infarction. PMID- 10869275 TI - Osborn waves of hypothermia. PMID- 10869276 TI - The electrophysiological basis of QT dispersion: global or local repolarization? PMID- 10869277 TI - Sustained ventricular arrhythmias in patients receiving thrombolytic therapy. PMID- 10869278 TI - The significance of expression of proliferating cell nuclear antigen in the cardiovascular system: mitosis or DNA repair? PMID- 10869279 TI - Significance of myocytes with positive DNA in situ nick end-labeling (TUNEL) in hearts with dilated cardiomyopathy. PMID- 10869281 TI - Cardiovascular news PMID- 10869280 TI - Thrombocytopenia and glycoprotein IIb/IIIa inhibitors: causation or association? PMID- 10869282 TI - Metabolic and nutritional considerations in nonalcoholic fatty liver. PMID- 10869283 TI - Liver from bone marrow in humans. AB - It has been shown in animal models that hepatocytes and cholangiocytes can derive from bone marrow cells. We have investigated whether such a process occurs in humans. Archival autopsy and biopsy liver specimens were obtained from 2 female recipients of therapeutic bone marrow transplantations with male donors and from 4 male recipients of orthotopic liver transplantations from female donors. Immunohistochemical staining with monoclonal antibody CAM5.2, specific for cytokeratins 8, 18, and 19, gave typical strong staining of hepatocytes, cholangiocytes, and ductular reactions in all tissues, to the exclusion of all nonepithelial cells. Slides were systematically photographed and then restained by fluorescence in situ hybridization (FISH) for X and Y chromosomes. Using morphologic criteria, field-by-field comparison of the fluorescent images with the prior photomicrographs, and persistence of the diaminiobenzidene (DAB) stain through the FISH protease digestion, Y-positive hepatocytes and cholangiocytes could be identified in male control liver tissue and in all study specimens. Cell counts were adjusted based on the number of Y-positive cells in the male control liver to correct for partial sampling of nuclei in the 3-micron thin tissue sections. Adjusted Y-positive hepatocyte and cholangiocyte engraftment ranged from 4% to 43% and from 4% to 38%, respectively, in study specimens, with the peak values being found in a case of fibrosing cholestatic recurrent hepatitis C in one of the liver transplant recipients. We therefore show that in humans, hepatocytes and cholangiocytes can be derived from extrahepatic circulating stem cells, probably of bone marrow origin, and such "transdifferentiation can replenish large numbers of hepatic parenchymal cells. PMID- 10869284 TI - Gastrin inhibits cholangiocyte growth in bile duct-ligated rats by interaction with cholecystokinin-B/Gastrin receptors via D-myo-inositol 1,4,5-triphosphate-, Ca(2+)-, and protein kinase C alpha-dependent mechanisms. AB - We studied the role of gastrin in regulating cholangiocyte proliferation induced by bile duct ligation (BDL). In purified cholangiocytes, we evaluated (1) for the presence of cholecystokinin-B (CCK-B)/gastrin receptors, (2) the effect of gastrin on D-myo-Inositol 1,4,5-triphosphate (IP(3)) levels, and (3) the effect of gastrin on DNA synthesis and adenosine 3', 5'-monophosphate (cAMP) levels in the absence or presence of CCK-A (L-364,718) and CCK-B/gastrin (L-365,260) receptor inhibitors, 1, 2-bis(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid tetrakis(acetxymethyl ester) (BAPTA/AM; an intracellular Ca(2+) chelator), and 2 protein kinase C (PKC) inhibitors, 1-(5-Isoquinolinylsulfonyl)-2-methylpiperazine (H7) and staurosporin. To evaluate if gastrin effects on cholangiocyte proliferation are mediated by the isoform PKCalpha, we evaluated (1) for the presence of PKCalpha in cholangiocytes and (2) the effect of gastrin on the PKCalpha protein expression in a triton-soluble (containing cytoplasm + membrane) and a triton-insoluble (containing cytoskeleton) fraction. To evaluate the effects of gastrin in vivo, immediately following BDL, gastrin or bovine serum albumin (BSA) was infused by minipumps for 7 days to rats and we measured cholangiocyte growth and cAMP levels. We found CCK-B/gastrin receptors on cholangiocytes. Gastrin increased IP(3) levels. Gastrin inhibited DNA synthesis and cAMP synthesis in cholangiocytes. Gastrin effects on cholangiocyte functions were blocked by L-365,260, BAPTA/AM, H7, and staurosporin but not by L-364,718. Gastrin induced translocation of PKCalpha from cholangiocyte cytoskeleton to membrane. In vivo, gastrin decreased cholangiocyte growth and cAMP synthesis compared with controls. We concluded that gastrin inhibits cholangiocyte growth in BDL rats by interacting with CCK-B/gastrin receptors through a signal transduction pathway involving IP(3), Ca(2+), and PKCalpha. PMID- 10869285 TI - Growth control of human biliary epithelial cells by interleukin 6, hepatocyte growth factor, transforming growth factor beta1, and activin A: comparison of a cholangiocarcinoma cell line with primary cultures of non-neoplastic biliary epithelial cells. AB - A well characterized human cholangiocarcinoma (CC) cell line, SG231, was compared with primary cultures of normal human biliary epithelial cells (BECs) for alterations in interleukin 6 (IL-6) and hepatocyte growth factor (HGF)-mediated stimulation and transforming growth factor beta1 (TGF-beta1) and activin A mediated inhibition of growth. Results were compared with immunolabeling of the original tumor and after injection of SG231 into the liver of BALB/cByJ-scid mice. In vitro, both BECs and CCs expressed met, gp80, and gp130 messenger RNA (mRNA) and protein, but the levels of expression were higher in the CCs than in the BECs. In both the CCs and BECs, exogenous HGF or IL-6 induced phosphorylation of met or gp130, respectively, and a concentration-dependent increase in DNA synthesis. However, the CCs but not BECs, continued to grow in basal serum-free medium (SFM) and spontaneously produced both IL-6 and HGF under these conditions, which resulted in auto-phosphorylation of gp130 and met, respectively; and neutralizing anti-HGF or anti-IL-6 alone inhibited CC growth, indicative of autocrine growth control circuits. Conversely, activin A inhibits the growth of both BECs and CCs, but does not significantly increase apoptosis. Activin-A induced growth inhibition of both CCs and BECs can be reversed by 100 ng/mL exogenous IL-6, but not by 10 to 100 ng/mL HGF. TGF-beta1 inhibited the growth of BECs but had no mitoinhibitory or proapoptotic effects on CCs. Immunolabeling of the original tumor and after inoculation into scid mice showed positive staining for met, gp130, gp80, and IL-6. This study contributes to a further understanding of BEC growth control and derangements that can occur during cholangiocarcinogenesis. PMID- 10869286 TI - Short- and long-term outcome of severe alcohol-induced hepatitis treated with steroids or enteral nutrition: a multicenter randomized trial. AB - Steroids are recommended in severe alcohol-induced hepatitis, but some data suggest that artificial nutrition could also be effective. We conducted a randomized trial comparing the short- and long-term effects of total enteral nutrition or steroids in these patients. A total of 71 patients (80% cirrhotic) were randomized to receive 40 mg/d prednisolone (n = 36) or enteral tube feeding (2,000 kcal/d) for 28 days (n = 35), and were followed for 1 year or until death. Side effects of treatment occurred in 5 patients on steroids and 10 on enteral nutrition (not significant). Eight enterally fed patients were prematurely withdrawn from the trial. Mortality during treatment was similar in both groups (9 of 36 vs. 11 of 35, intention-to-treat) but occurred earlier with enteral feeding (median 7 vs. 23 days; P =.025). Mortality during follow-up was higher with steroids (10 of 27 vs. 2 of 24 intention-to-treat; P =. 04). Seven steroid patients died within the first 1.5 months of follow-up. In contrast to total enteral nutrition (TEN), infections accounted for 9 of 10 follow-up deaths in the steroid group. In conclusion, enteral feeding does not seem to be worse than steroids in the short-term treatment of severe alcohol-induced hepatitis, although death occurs earlier with enteral nutrition. However, steroid therapy is associated with a higher mortality rate in the immediate weeks after treatment, mainly because of infections. A possible synergistic effect of both treatments should be investigated. PMID- 10869287 TI - High-dose and long-term therapy with interferon-alfa inhibits tumor growth and recurrence in nude mice bearing human hepatocellular carcinoma xenografts with high metastatic potential. AB - Postoperative recurrence of human hepatocellular carcinoma (HCC) is the major issue that must be addressed to further improve prognosis. This study was undertaken to investigate the effects of interferon-alfa-1b (IFN-alpha-1b) on recurrent tumor and metastasis after curative resection in nude mice bearing an HCC xenograft with high metastatic potential. Tumor tissues from LCI-D20, a metastatic model of HCC in nude mice, were orthotopically implanted in 105 nude mice. Eleven days later, 64 mice underwent curative resection of liver tumors. IFN-alpha at different doses was administered subcutaneously to mice with or without resection. In mice without resection, when comparison was made among control, IFN 7.5 x 10(6) U/kg/day, 1.5 x 10(7) U/kg/day for treated groups, and 3 x 10(7) U/kg/day; tumor volume was 8,475 mm(3) +/- 2,636 mm(3), 7,963 mm(3) +/- 3,214 mm(3), 769 mm(3) +/- 287 mm(3), and 13 mm(3) +/- 9 mm(3); incidence of lung metastasis being 100%, 80%, 40%, and 0%; life span was 45 +/- 4 days, 53 +/- 8 days, 81 +/- 6 days, and 105 +/- 24 days, respectively. In mice with curative resection, when comparison was made among control, IFN 5 x 10(5) U/kg/day, 1 x 10(6) U/kg/day, 4 x 10(6) U/kg/day, 7.5 x 10(6) U/kg/day, 1.5 x 10(7) U/kg/day, and 3 x 10(7) U/kg/day for treated groups; incidence of recurrent tumor was 100%, 100%, 87.5%, 100%, 87.5%, 62.5%, and 12.5%; lung metastasis being 100%, 75%, 87.5%, 50%, 62.5%, 0%, and 0%, respectively. IFN-alpha inhibited neovascularization induced by LCI-D20 tumor specimens implanted into the micropocket of nude mice corneas. In conclusion, high-dose and long-term therapy with IFN-alpha dose-dependently inhibits tumor growth and recurrence after resection of HCC. The effect of IFN-alpha may be attributed to antiangiogenesis in this experiment. These results provide potential clinical implication, particularly for the prevention of recurrence after curative resection of HCC. PMID- 10869288 TI - Hepatic cytochrome P450 down-regulation during aseptic inflammation in the mouse is interleukin 6 dependent. AB - Expression of cytochromes P450 (CYP) is markedly reduced during inflammatory processes. In vitro studies with hepatocytes have shown that cytokines generated during these processes down-regulate CYP. However, it is not clear to what extent each individual cytokine contributes to the overall reduced expression of the various CYP isoenzymes in vivo. Interleukin 6 (IL-6), a major player during inflammatory processes, is recognized as the most important cytokine modulating the hepatic expression of acute-phase protein (APP) genes. For this reason, we selected the IL-6(-/-) mouse as a model to investigate the role of IL-6 in the down-regulation of hepatic CYP during experimental inflammation. Our results show that the reduction in messenger RNA (mRNA) levels of CYP1A2, CYP2A5, and CYP3A11 during turpentine-induced inflammation was abrogated in IL-6-deficient mice, confirming that IL-6 is an indispensable player for the down-regulation of hepatic CYP during aseptic inflammation. Moreover, the different CYP isoenzymes showed a variable grade of dependence on IL-6, CYP2A5 being the most sensitive one. In the case of CYP2E1, differences between IL-6(-/-) and wild-type mice were no longer maintained after 24 hours, suggesting a delayed, rather than abrogated, CYP down-regulation in the absence of IL-6. As opposed to that, hepatic CYP repression took place in IL-6-deficient mice during lipopolysaccharide (LPS) mediated inflammation. This contrasting behavior observed for CYP is surprisingly similar to the one seen for extracellular (serum amyloid A, beta-fibrinogen) and intracellular (metallothionein-1) APPs and points to the fact that, in the model of bacterial inflammation (LPS), the effects of IL-6 on CYP down-regulation are likely to be substituted by other cytokines or mediators. PMID- 10869290 TI - Mrp2 is essential for estradiol-17beta(beta-D-glucuronide)-induced cholestasis in rats. AB - The present study evaluates the roles of the multidrug resistance-1 P glycoprotein, Mdr1a/1b, the bile salt export pump (Bsep), and the multidrug resistance-associated protein-2 (Mrp2) in mediating cholestasis induced by estradiol-17beta(beta-D-glucuronide) (E(2)17G). Administration of ?(3)HE(2)17G (18 nmol/g body weight) gave a similar degree of cholestasis and biliary excretion of E(2)17G-equivalents in wild-type and Mdr1a(-/-)/1b(-/-) mice. When expressed in Sf9 cells, Bsep-mediated adenosine triphosphate (ATP)-dependent transport of taurocholate (TC, 1 micromol/L) in membrane vesicles was 110% +/- 12.5% and 108% +/- 17.3% of control in the presence of 10 and 50 micromol/L E(2)17G, respectively, whereas in rat canalicular membrane, both E(2)17G and the choleretic estradiol-3-beta-D-glucuronide (E(2)3G) inhibited ATP-dependent transport of TC to the same extent. Infusion of ?(3)HE(2)17G (24 micromol) did not induce cholestasis in Mrp2-deficient TR(-) rats whereas 2 micromol of ?(3)HE(2)17G inhibited bile flow by 51% in control Wistar rats. The maximal biliary concentration of E(2)17G was 3.5 and 2.5 mmol/L in control and TR(-) rats, respectively. However, 2.2 mmol/L of E(2)17G in bile is associated with inhibition of bile flow in control rats. These data show that (1) Mdr1a/1b are not essential for E(2)17G-mediated cholestasis, (2) direct inhibition of Bsep mediated bile acid transport is not the mechanism for E(2)17G cholestasis, and (3) accumulation of E(2)17G in bile alone is not sufficient to induce cholestasis. These data indicate that the process of Mrp2-mediated transport of high concentrations of E(2)17G is essential for its induction of cholestasis. PMID- 10869289 TI - Human placenta sphingomyelinase, an exogenous acidic pH-optimum sphingomyelinase, induces oxidative stress, glutathione depletion, and apoptosis in rat hepatocytes. AB - Ceramide has been identified as a putative lipid messenger that mediates diverse cellular processes including cell death. Since glutathione (GSH) depletion is known to sensitize cells to many cytotoxic agents and as a result of the reported regulation of neutral sphyngomyelinase (NSMase) by GSH, the present study compared the role of individual SMases in the induction of oxidative stress, regulation of cellular GSH, and apoptosis of rat hepatocytes. Exposure of cultured rat hepatocytes to exogenous Bacillus cereus sphingomyelinase (bSMase), a neutral SMase, or human placenta sphingomyelinase (hSMase), an acidic SMase (ASMase), generated similar ceramide levels in a dose-dependent manner. However, whereas bSMase increased hepatocellular GSH levels, hSMase depleted GSH stores, an effect that was prevented by monensin and mannose 6-phosphate (M-6-P), suggesting that exogenous hSMase enters hepatocytes by endocytosis and is delivered to an endosomal/lysosomal acidic compartment. Interestingly, despite the differential effect of either SMases on cell GSH levels, both bSMase and hSMase increased gamma-glutamylcysteine synthetase heavy-subunit chain (gamma-GCS HS) mRNA levels. Consistent with these findings on GSH regulation, hSMase, but not bSMase, generated reactive oxygen species (ROS), being accompanied by mitochondrial depolarization, suggesting that hSMase targeted mitochondria, leading to oxidative stress. Accordingly, hepatocytes displayed a selective sensitivity to hSMase in contrast to bSMase exposure, and depletion of GSH stores enhanced susceptibility to hSMase as a result of potentiation of ROS formation and caspase 3 activation. Thus, these findings reveal the ability of ASMase to induce oxidative stress as a result of the targeting of mitochondria, and that GSH depletion sensitizes hepatocytes to the ASMase-induced apoptosis. PMID- 10869291 TI - Reversion of hepatobiliary alterations By bone marrow transplantation in a murine model of erythropoietic protoporphyria. AB - Erythropoietic protoporphyria (EPP) is characterized clinically by cutaneous photosensitivity and biochemically by the accumulation of excessive amounts of protoporphyrin in erythrocytes, plasma, feces, and other tissues, such as the liver. The condition is inherited as an autosomal dominant or recessive trait, with a deficiency of ferrochelatase activity. A major concern in EPP patients is the development of cholestasis with accumulation of protoporphyrin in hepatobiliary structures and progressive cellular damage, which can rapidly lead to fatal hepatic failure. The availability of a mouse model for the disease, the Fech(m1Pas)/Fech(m1Pas) mutant mouse, allowed us to test a cellular therapy protocol to correct the porphyric phenotype. When Fech/Fech mice received bone marrow cells from normal animals, the accumulation of protoporphyrin in red blood cells and plasma was reduced 10-fold but still remained 2.5 times above normal levels. Interestingly, in very young animals, bone marrow transplantation can prevent hepatobiliary complications as well as hepatocyte alterations and partially reverse protoporphyrin accumulation in the liver. Bone marrow transplantation may be an option for EPP patients who are at risk of developing hepatic complications. PMID- 10869292 TI - Rifamycin SV and rifampicin exhibit differential inhibition of the hepatic rat organic anion transporting polypeptides, Oatp1 and Oatp2. AB - The antibiotics, rifamycin SV and rifampicin, are known to interfere with hepatic bile salt and organic anion uptake. The aim of this study was to explore which transport systems are affected. In short-term-cultured rat hepatocytes, low concentrations (10 micromol/L) of both compounds inhibited mainly sodium independent taurocholate uptake, whereas higher concentrations (100 micromol/L) also inhibited sodium-dependent taurocholate uptake. In Xenopus laevis oocytes expressing the Na(+)/taurocholate cotransporting polypeptide (Ntcp), high rifamycin SV and rifampicin concentrations were required for inhibition of taurocholate uptake. In contrast, sodium-independent taurocholate uptake mediated by the organic anion transporting polypeptides, Oatp1 and Oatp2, was already substantially inhibited by 10 micromol/L rifamycin SV. Rifampicin potently inhibited Oatp2-mediated taurocholate uptake, but did not interfere with Oatp1 mediated taurocholate uptake. Similar effects of rifamycin SV and rifampicin were found for Oatp1- and Oatp2-mediated estradiol-17beta-glucuronide transport. Dixon plot analysis yielded a pattern compatible with competitive inhibition of estradiol-17beta-glucuronide transport with K(i) estimates of 6.6 micromol/L and 7.3 micromol/L for rifamycin SV-induced inhibition of Oatp1 and Oatp2, respectively, and of 1.4 micromol/L for rifampicin-induced inhibition of Oatp2. These results demonstrate that rifamycin SV and rifampicin exhibit differential inhibition on Oatp1 and Oatp2, and identify rifampicin as a selective Oatp2 inhibitor. The data indicate that these inhibitors can be used to determine the in vivo relevance of Oatp1 and Oatp2 for the overall bioavailability and disposition of drugs and other Oatp1/2 substrates. PMID- 10869293 TI - Hepatitis C-related cirrhosis: a predictor of diabetes after liver transplantation. AB - Hepatitis C virus (HCV) infection has recently been suggested to be a risk factor for the development of diabetes mellitus. The aim of our study was to investigate whether the prevalence of diabetes is increased among liver transplant recipients infected with HCV. We compared the prevalence of diabetes among 278 liver transplant recipients whose original cause of liver failure was HCV infection (110 patients), hepatitis B virus infection (HBV; 53 patients), and cholestatic liver disease (CLD; 115 patients). The pretransplantation prevalence of diabetes was higher in the HCV group (29%) compared with the HBV (6%) and CLD (4%) groups (P <.001). The prevalence of diabetes remained higher in the HCV group 1 year after transplantation: 37%, 10%, and 5% in the HCV, HBV, and CLD groups, respectively (P <.001). The cumulative steroid dose during the first year of transplantation was significantly lower in the HCV group compared with the CLD group. Multivariate analysis revealed that HCV-related liver failure (P =.002), pretransplantation diabetes (P <.0001), and male sex (P =.019) were independent predictors of the presence of diabetes 1 year after transplantation. The high prevalence of diabetes persisted in the HCV group, with 41% diabetic at 5 years. The majority of patients with diabetes mellitus (89%) required insulin therapy after transplantation. Patient and graft survival rates were similar among patients with and without diabetes. In conclusion, our study shows that there is a high prevalence of diabetes among liver transplant recipients infected with HCV both before and after transplantation. PMID- 10869294 TI - Low frequency of cirrhosis in a hepatitis C (genotype 1b) single-source outbreak in germany: a 20-year multicenter study. AB - From August 1978 until March 1979, 14 batches of anti-D immune globulin contaminated with hepatitis C virus (HCV) genotype 1b (20, 000-480,000 copies/dose) from a single erythrocyte donor had been administered for prophylaxis of rhesus isoimmunization throughout East Germany. All 2,867 women involved had been recalled after January 12, 1979 for repeated screening of alanine transaminase (ALT). They were prospectively followed in regional centers. We have reexamined a cohort of 1,018 women (median age 24, range 16-38 years at infection) on follow-up for 20 years in 9 representative centers. Within 6 months after anti-D administration, 10% of these women had no evidence of disease and 90% had acute hepatitis C (n = 917) including 49% with symptomatic and 22% with icteric course. After 20 years, 85% of the 917 affected women still tested positive for HCV antibodies (among them 3% responded to interferon treatment) and 55% were positive for HCV RNA (among them 7% were nonresponders to interferon and 3% were apparent HCV carriers). Only 4 (0.4%) had overt cirrhosis. Two (0.2%) died of superinfected fulminant hepatitis B or alcoholism and cirrhosis, respectively. Histology obtained in 44% of the viremic women showed hepatitis of minimal to moderate grade in 96%, portal fibrosis in 47%, and septal fibrosis in 3% of the cases. In conclusion, formerly healthy young women, without hepatic comorbidity, may clear HCV (1b) infection in half of the cases or develop mild chronic hepatitis C with low risk of progression to cirrhosis within 20 years. PMID- 10869295 TI - Detection of hepatitis C virus replication by In situ hybridization in epithelial cells of anti-hepatitis C virus-positive patients with and without oral lichen planus. AB - Epidemiological studies have demonstrated that there is a correlation between oral lichen planus and chronic hepatitis C virus (HCV) infection. HCV RNA has been recently detected in epithelial cells from oral lichen planus lesions by reverse-transcription polymerase chain reaction (RT-PCR). However, this technique does not discriminate which types of cells are infected by the virus or if the viral RNA is present in the serum that contaminates the biopsy. Morphological evidence of viral replication in cells from these lesions is needed to establish a role for HCV in oral lichen planus. Consequently, we have analyzed the presence of positive and negative HCV-RNA strands in oral mucosa biopsies from 23 patients (14 anti-HCV-positive) diagnosed as having oral lichen planus and from 5 patients with chronic hepatitis C without oral lichen planus. Positive and negative HCV RNA strands were detected in epithelial cells of the mucosa biopsies from all anti-HCV-positive patients independently of whether or not they had oral lichen planus, but in none of the anti-HCV-negative cases. The percentage of stained cells ranged from 4.4% to 14.3%. These percentages do not correlate with the serum viremia levels or the intensity of the cellular infiltrate in patients with oral lichen planus. In conclusion, we have shown that HCV replicates in epithelial cells of patients with and without oral lichen planus. The pathological consequences of this finding remain to be elucidated. PMID- 10869296 TI - DNA-Based immunization produces Th1 immune responses to hepatitis delta virus in a mouse model. AB - Hepatitis delta virus (HDV) superinfection is one of the major causes of fulminant hepatitis in endemic areas of hepatitis B virus (HBV) infection. Currently, there is no effective treatment or vaccine against HDV superinfection. DNA-based immunization is a promising antiviral strategy to prevent or treat persistent viral infections. In this study, we investigated the immunological effects of DNA vaccines against HDV in BALB/c mice. Plasmid (pD) encoding large hepatitis D antigen (L-HDAg), or plasmid (pS/pD) coexpressing hepatitis B surface antigen (HBsAg) and L-HDAg, were injected into mice intramuscularly. The seroconversion rate, anti-HBs levels, anti-HDV titers, T-cell proliferation responses, and T-helper (Th)-release cytokine profiles were analyzed. Mice immunized with plasmids, pS/pD or pD, produced low, but significant, titers of anti-HDV antibodies. In contrast, pS/pD induced much stronger anti-HBs titers in the immunized animals. Interestingly, splenic lymphocytes derived from pS/pD inoculated mice demonstrated significant proliferation responses to recombinant HBsAg and HDAg, and resulted in a Th1-like immune response as suggested by the production of interferon gamma (INF-gamma) and interleukin-2 (IL-2), but not IL 4. The splenic lymphocyte derived from the pD-inoculated mice showed a similar Th1 response to the stimulation of HDAg, but not to HBsAg. In conclusion, our results suggest that DNA vaccines against HDV can induce significant cellular immune responses with a Th1 preference. HBV and HDV coimmunization can be performed by DNA vaccines. These results are promising for the future development of prophylactic and therapeutic HDV vaccines. PMID- 10869297 TI - Hepatitis C virus (HCV) infection in a community in the Nile Delta: population description and HCV prevalence. AB - This report describes a cross-sectional survey of the prevalence of antibodies to hepatitis C virus (anti-HCV) in a rural Egyptian community in the Nile Delta. One half of the village households were systematically selected and examined by questionnaire and testing sera for anti-HCV and HCV RNA. Blood samples were obtained from 3, 888 (75.4%) of 5,156 residents >/=5 years of age; an additional 111 samples were obtained from children younger than 5 years. Overall, 973 (24.3%) of 3,999 residents were anti-HCV-positive, and the age- and gender adjusted seroprevalence was 23.7%. Anti-HCV prevalence increased sharply with age, from 9.3% in those 20 years of age and younger to >50% in those older than 35 years. Currently or previously married individuals were more likely to be seropositive than those never married, controlling for age (Mantel-Haenszel risk ratio = 1.8; 95% CI: 1.3, 2.6). Of the 905 anti-HCV-positive samples tested, 65% were also positive for HCV RNA. Active schistosomal infection was not associated with anti-HCV status; however, history of antischistosomal injection therapy (reported by 19% of anti-HCV positives) was a risk for anti-HCV (age-adjusted risk ratio = 1.3; 95% CI: 1.2, 1.5). This study, the largest community-based survey to date, supports earlier reports of high levels of anti-HCV among adults in rural areas of Egypt, although many of those who are seropositive will not have active liver disease. The large reservoir of HCV infection in the community provides an opportunity to investigate risk factors for transmission, the natural history of infection and effectiveness of preventive methodologies, and raises concern about the prospect of an increasing incidence of chronic liver disease in the coming decades. PMID- 10869298 TI - Molecular analysis of hepatitis B virus DNA in serum and peripheral blood mononuclear cells from hepatitis B surface antigen-negative cases. AB - We have analyzed the molecular bases of the persistence of hepatitis B virus (HBV) DNA in serum and peripheral blood mononuclear cells (PBMC) in the absence of detectable hepatitis B surface antigen (HBsAg) in hemodialysis patients and dialysis-unit staff members who had suffered acute hepatitis B that resolved previously. HBV DNA was found in both compartments by polymerase chain reaction (PCR) using primers of the pre-S/S region. Viral DNA was transcriptionally active in PBMC, because the covalently closed circular (ccc) HBV DNA, the template for the viral RNA transcription, was detected in 47% of the samples. Furthermore, all PBMC had HBV RNA. HBsAg-negative cases had statistically lower levels of HBV DNA in serum and PBMC than a control group of chronic HBsAg carriers. We have also studied the presence of immune complexes and the existence of mutations in the pre-S/S gene to explain the lack of detection of HBsAg in these cases. No serum HBsAg/hepatitis B surface antigen antibody (anti-HBs) immune complexes or mutations in the "a determinant of the S gene were found. However, we have observed that all HBsAg-negative cases were infected by a mixture of the wild type virus and a deletion mutant in the pre-S1 region. This deletion (amino acids 58-118) affects the S gene promoter, and previous in vitro studies have shown that it produces a reduction of the HBsAg synthesis. In conclusion, this work shows that the lack of detection of HBsAg in the presence of low viral levels of replication may be caused by the existence of viral genomes harboring deletions in the pre-S1 region that affect the S promoter. PMID- 10869299 TI - Long-term follow-up study of core gene deletion mutants in children with chronic hepatitis B virus infection. AB - Core gene deletion mutants of hepatitis B virus (HBV) have been identified in adults. Because the acquisition of HBV occurs mainly in infancy and childhood in hyperendemic areas, this study aimed to learn the temporal profile of such mutants in children with chronic HBV infection. We have followed up 365 HBV infected children for more than 10 years and screened out HBV core gene deletion from their sera. Serial serum samples of positive cases were subjected to HBV-DNA nucleotide sequence analyses and quantification. Deletion mutants were found in 18 of the 365 patients (4.9%). Most cases (15 of 18) with deletion mutants heralded hepatitis B e antigen (HBeAg) seroconversion phase, while the other cases (3 of 18) remained in HBeAg-seropositive phase. Deletion mutants disappeared after HBeAg seroconversion except in 1 child. Decreased HBV-DNA levels accompanied deletion mutants for those who finally underwent HBeAg seroconversion, but the HBV-DNA level did not decline if there was no seroconversion. Deletion mutants were not associated with a particularly high peak liver enzyme. Core gene deletion mutants could appear as early as the age of 5. The duration of their appearance was 0.5 to 5 years. Horizontal rather than perinatal transmission of HBV was a favorable factor for these mutants to develop. Deletion fragments were located in the middle part of core gene. The emergence of the mutants was likely the result of host-viral interaction and mostly signified HBeAg seroconversion within 1 year. Core gene deletion mutants appeared preferably in children acquiring HBV by horizontal transmission. PMID- 10869300 TI - Adefovir dipivoxil for the treatment of lamivudine-resistant hepatitis B mutants. AB - Lamivudine has been shown to be an effective therapy for chronic hepatitis B, but resistance to this nucleoside agent is common after prolonged use. Five patients with chronic hepatitis B virus (HBV) infection developed resistance to lamivudine after 9 to 19 months of treatment. In 4 patients this occurred after liver transplantation and the remaining individual had stable cirrhosis. In each case, resistance was confirmed to be caused by one or more mutations in the HBV-DNA polymerase gene and was associated with active underlying liver disease. The patients were treated with adefovir dipivoxil in a dose of 5 to 30 mg daily. Two to 4 log(10) reductions in HBV-DNA levels were observed in 4 cases, and the fifth patient became negative by quantitative polymerase chain reaction (PCR) after retransplantation in conjunction with hepatitis B immunoglobulin (HBIg). Virologic improvement was associated with stable or declining serum alanine transaminase levels in 4 patients. HBV-DNA suppression has been sustained during a mean treatment period of 13 months (range 11 to 15 months), including 1 patient in whom lamivudine has been discontinued. Mild changes in renal function were observed during treatment in most cases but did not require early discontinuation of the drug. This study provides evidence that adefovir dipivoxil can be an effective treatment for lamivudine-resistant HBV mutants as well as wild-type HBV. PMID- 10869301 TI - Iron reduction as an adjuvant to interferon therapy in patients with chronic hepatitis C who have previously not responded to interferon: a multicenter, prospective, randomized, controlled trial. AB - Hepatic iron concentration has consistently been observed as being directly correlated with the response to interferon therapy in chronic hepatitis C virus (HCV). We therefore conducted a randomized, controlled trial comparing iron reduction by phlebotomy with iron reduction followed by retreatment with interferon in 96 patients with chronic hepatitis C who had previously not responded to a course of interferon. During the initial phase when all patients were undergoing phlebotomy, we found that serum alanine transaminase (ALT) activities decreased but by less than 50% from baseline in 67 patients (89%), decreased by more than 50% in 12 patients (13%) and became normal in 9 patients (9%) with no overall change in HCV-RNA levels. Subsequently no patient in either treatment group achieved a sustained virologic response. Improvements in necroinflammatory changes were noted in liver biopsy specimens in those patients receiving phlebotomy plus interferon (mean index 8.59 vs. 7.37, P <. 05). A slight but not statistically significant decrease in histologic activity index was noted in those subjects treated by phlebotomy alone (mean index 8.4 vs. 7.75, P not significant). We conclude that, although prior phlebotomy therapy does not improve the rate of sustained response to interferon retreatment, it does result in less liver injury manifested by a decrease in serum transaminase activity and a slight improvement in liver histopathology. PMID- 10869302 TI - Increased hepatocyte turnover and inhibition of woodchuck hepatitis B virus replication by adefovir in vitro do not lead to reduction of the closed circular DNA. AB - The aim of this study was to evaluate the inhibitory effect of the nucleotide analogue adefovir on woodchuck hepatitis B virus (WHV) replication and, in particular, to determine whether the pool of covalently closed circular DNA (cccDNA) could be reduced by adefovir treatment in primary cultures of woodchuck hepatocytes isolated from a chronic carrier. Strong reduction of WHV-DNA synthesis (90%) and secretion (up to 98%) was observed with all 3 doses of adefovir used (1, 10, and 100 micromol/L), whereas in the absence of the drug, high amounts of viral particles were continuously secreted in the culture medium until the end of the study (27 days). Secretion of envelope proteins and viral RNA levels remained constant both in the adefovir-treated and -untreated cultures for the entire course of the study. Intracellular core protein levels declined by approximately 50% in all the cultures, independent of adefovir treatment. There was no indication of cccDNA loss in the adefovir-treated hepatocyte cultures even when cell turnover was induced for 14 days by the addition of epidermal growth factor (EGF) to the culture medium. Our data show that adefovir has a very strong inhibitory effect on WHV-DNA synthesis in chronically infected primary hepatocyte cultures and indicate that cccDNA is a very stable molecule that appears to be efficiently transmitted to the dividing hepatocytes. PMID- 10869303 TI - Clinicopathological conference: hepatitis C in a patient with human immunodeficiency virus infection. PMID- 10869304 TI - Chromosomes and cirrhosis: all's well that ends well? PMID- 10869305 TI - Cyp2e1 and ALD. PMID- 10869306 TI - Nitric oxide as a calcium wave accelerator. PMID- 10869307 TI - Interpreting epidemiological evidence: how meta-analysis and causal inference methods are related. AB - Interpreting observational epidemiological evidence can involve both the quantitative method of meta-analysis and the qualitative criteria-based method of causal inference. The relationships between these two methods are examined in terms of the capacity of meta-analysis to contribute to causal claims, with special emphasis on the most commonly used causal criteria: consistency, strength of association, dose-response, and plausibility. Although meta-analysis alone is not sufficient for making causal claims, it can provide a reproducible weighted average of the estimate of effect that seems better than the rules-of-thumb (e.g. majority rules and all-or-none) often used to assess consistency. A finding of statistical heterogeneity, however, need not preclude a conclusion of consistency (e.g. consistently greater than 1.0). For the criteria of strength of association and dose-response, meta-analysis provides more precise estimates, but the causal relevance of these estimates remains a matter of judgement. Finally, meta analysis may be used to summarize evidence from biological, clinical, and social levels of knowledge, but combining evidence across levels is beyond its current capacity. Meta-analysis has a real but limited role in causal inference, adding to an understanding of some causal criteria. Meta-analysis may also point to sources of confounding or bias in its assessment of heterogeneity. PMID- 10869308 TI - Childhood cancers, birthplaces, incinerators and landfill sites. AB - BACKGROUND: In all, 70 municipal incinerators, 307 hospital incinerators and 460 toxic-waste landfill sites in Great Britain were examined for evidence of effluents causing childhood cancers. Municipal incinerators had previously shown significant excesses of adult cancers within 7.5 and 3.0 km. The relative risks for adults had been marginal and an analysis of childhood cancers seemed to offer a more sensitive approach. METHODS: A newly developed technique of analysis compares distances from suspect sources to the birth addresses and to the death addresses of cancer-children who had moved house. A localized hazard, effective at only one of these times, must be preferentially associated with the corresponding address. This creates an asymmetry of migrations towards or away from age-restricted effective sources. RESULTS: The child-cancer/leukaemia data showed no systematic migration-asymmetries around toxic-waste landfill sites; but showed highly significant excesses of migrations away from birthplaces close to municipal incinerators. Relative risks within 5.0 km of these sites were about 2:1. Hospital incinerators gave analogous results. The ratios greatly exceed findings around 'non-combustion' urban sites. CONCLUSIONS: Because of their locations, the specific effects of the municipal incinerators could not be separated clearly from those of adjacent industrial sources of combustion effluents. Both were probably carcinogenic. Landfill waste sites showed no such effect. PMID- 10869309 TI - The decline in the mortality rates of cervical cancer and a plausible explanation in Shandong, China. AB - BACKGROUND: The aim of the present study is to describe the temporal trends in mortality rates of cervical cancer in Shandong Province, China, and to elucidate their likely explanations. METHODS: Three retrospective surveys of all causes of death in 1970-1974, 1985-1989 and 1990-1992 in Shandong were carried out. An age period-cohort analysis based on similar survey data from Qixia, a county in Shandong, from 1970 to 1994 was performed. A correlation analysis between prevalence of syphilis and cervical cancer mortality rates three decades later was conducted. A cross-sectional survey of risk factors for cervical cancer was conducted in 1991 in one city and 12 villages in the province. RESULTS: A marked decline in cervical cancer mortality rates was observed from 1970 to 1992 and in successive birth cohorts from 1892 to 1927, and rates remained relatively constant in subsequent birth cohorts through that of 1952. The decline in these rates was strongly correlated with a decline in positive serological tests for syphilis 32 years previously. The percentages of women with selected risk factors were lower in younger women (30-54 years) than in older women (55-69 years) in both cities and rural areas. CONCLUSION: These trends are compatible with a decrease in risk of exposure to sexually transmitted factors at an early age after the founding of the People's Republic of China in 1949, and a decline in lifetime duration of exposure to possible co-factors in successive birth cohorts who reached sexual maturity before that time. PMID- 10869310 TI - Blood type and family cancer history in relation to precancerous gastric lesions. AB - BACKGROUND: The increased odds of stomach cancer among subjects with blood type A have been reported in epidemiological studies. AIM: To study the relation of family history of gastric cancer and ABO blood type with precancerous gastric lesions in a high-risk area for stomach cancer. Subjects and setting We examined 3400 adults aged 35-64 in a population-based gastric endoscopic screening in a county in China with one of the highest rates of stomach cancer in the world. METHODS: In this cross-sectional study, data on family cancer history, ABO blood type and other characteristics of the participants were obtained by interview and blood test. Responses were compared between those with the most advanced gastric lesions, dysplasia (DYS) or intestinal metaplasia (IM), versus those with chronic atrophic gastritis (CAG) or superficial gastritis (SG). RESULTS: The prevalence odds ratio (OR) for blood type A relative to other types was 1.39 (95% CI : 1.12 1.73) for DYS and 1.28 (95% CI : 1.06-1.53) for IM. The OR associated with parental history of stomach cancer was 1.88 (95% CI : 1.20-2.95) for DYS, but the numbers were too small to evaluate aggregation among siblings. The combined OR associated with blood type A and a parental of history of gastric cancer was 2.61 (95% CI : 1.59-4.30) for DYS and 1.46 (95% CI : 0.93-2.31) for IM. CONCLUSIONS: The findings suggest that genetic factors play a role in developing precancerous gastric lesions. PMID- 10869311 TI - Patterns of familial aggregation of three melanoma risk factors: great number of naevi, light phototype and high degree of sun exposure. AB - BACKGROUND: Besides melanoma susceptibility genes and shared environmental exposures, part of the familial clustering of cutaneous malignant melanoma (CMM) might be due to familial aggregation of melanoma-associated phenotypes. Our goal was to assess the patterns of familial aggregation of three melanoma risk factors: great number of naevi (GNN), light phototype (LP) and high degree of sun exposure (HDSE). METHODS: Familial aggregation of GNN, LP and HDSE was investigated in 66 French families with at least two CMM cases and was measured by the asssociation of the relatives' traits with the probands' traits, using the generalized estimating equations approach. The probands were the melanoma cases leading to ascertainment of the families, subdivided into cases (with the trait studied) and controls (without the trait). RESULTS: We found significant evidence for familial aggregation of GNN only among sibs (OR = 3.7, 95% CI : 1.4-10.5, P = 0.01), of LP among blood relatives (OR = 3.8, 95% CI : 1.8-8.0, P = 0.004) and of HDSE among blood relatives (OR = 4.5, 95% CI : 2.1-9.9, P < 0.001) and spouses (OR = 44.3, 95% CI : 5.1-382.2, P < 10(-3)). These results suggest that genetic factors might account for the clustering of GNN and LP and shared environment for the aggregation of HDSE. The GNN clustering was lower in families with increasing numbers of CMM (>/=3 cases) or presence of p16 mutations, the opposite being observed for LP and HDSE. Moreover, the familial aggregation of LP was significantly lower in families with highly sun-exposed members. CONCLUSION: Melanoma might not only result from specific genetic and environmental factors but also from those underlying melanoma-associated traits involving complex gene gene and gene-environment interactions. PMID- 10869312 TI - Cutaneous malignant melanoma in New Zealand: trends by anatomical site, 1969 1993. AB - BACKGROUND: Site-specific trend analysis is probably the most effective method available for assessing how the long-term trend in melanoma rates relates to changes in sun exposure and behaviour. New Zealand has very high incidence of and mortality from melanoma and the fraction of melanoma cases and deaths with a site specified has been comparatively high. METHODS: Trends in incidence and mortality from melanoma in New Zealand were analysed between 1969 and 1993, by sex and body site. A graphical representation of the trend by birth-cohort and age-period cohort modelling were used. RESULTS: For all sites combined, the annual increase in incidence was 6.7% (95% CI : 6.3-7.1%) in men and 3.1% (95% CI : 2.3-3.7%) in women. The increase was significantly greater at each site for males. The largest increases occurred for the upper limbs in males (7.3% a year) and the trunk in females (3.8% a year). Incidence rates slowed appreciably in the later years (currently about 26/100 000 for each sex) and no further increase in lifetime risk of melanoma was observed among post World War II generations. Mortality trends paralleled those for incidence with a 25-year gap, with a more modest rate of increase (2-3% per annum for each sex), essentially due to the increased risk among generations born up to 1919 or 1924. Age-standardized death rates have now stabilized in New Zealand at about 5.5/100 000 (men) and 3.2/100 000 (women). Trends between cohorts were the most marked for sites with a likely intermittent pattern of exposure, and were consistent overall for the trunk and the limbs. CONCLUSIONS: Results support the hypothesis that changes in lifestyle factors resulted in a pattern of carcinogenic exposures that explains both the upsurge in melanoma in the last few decades and the current levelling off in incidence. PMID- 10869313 TI - Cancer following cardiac catheterization in childhood. AB - BACKGROUND: Low-dose ionizing radiation is one of the definitive risk factors for cancer development. Nevertheless, only a few follow-up studies of children subjected to cardiac catheterization have been performed, yielding inconsistent results. METHODS: Our study group included 674 children who underwent cardiac catheterization due to congenital anomalies, between the years 1950-1970 in three major medical centres in Israel. A registered nurse conducted a review of the children's medical files in each hospital. Demographic data and vital status were ascertained from the Israeli National Registry, using a unique identity number. Subsequently, the study cohort was linked with the Israeli National Cancer Registry, in order to identify cancer cases that had been diagnosed through December 1996, the last follow-up date of the study. RESULTS: Over 75% of the study participants were native-born; 56.2% were males. Approximately 78% of the cohort subjects were alive at the end of follow-up; 28.6% of the participants underwent more than one procedure. All of the diagnosed cases occurred in males. Expected number of malignancies for all sites was 4.75, while the observed number was 11.0 (standardized incidence ratio [SIR] = 2. 3; 95% CI : 1.2-4.1). Of the 11 cancer cases, 4 lymphomas were observed (0.63 were expected, SIR = 6.3; 95% CI : 1.7-16.2). One of these was Hodgkin's Disease. There were also three cases of melanoma as opposed to 0.62 expected (SIR = 4.9; 95% CI : 1.0-14.2). CONCLUSIONS: This finding is compatible with current knowledge about the carcinogenic effect of low-dose irradiation but differs in the occurrence of an excess of lymphoma in the absence of an excess of leukaemia, which has not been reported before. The dissonance between males and females is yet to be resolved. IMPLICATIONS: Radiation doses that are used currently during cardiac catheterization are lower than in the past. Yet, the procedure is more common and frequently involves longer duration due to therapeutic interventions. The possible long-term results of such an exposure should be kept in mind. PMID- 10869314 TI - Siesta and the risk of coronary heart disease: results from a population-based, case-control study in Costa Rica. AB - BACKGROUND: The siesta (afternoon nap or rest), a common traditional behaviour in tropical areas, may increase the risk of myocardial infarction (MI) since the post siesta cardiovascular response very closely resembles the period soon after waking up in the morning when the onset of acute cardiovascular events is high. METHODS: We studied 505 MI survivors and 522 randomly selected controls, matched for age, gender, and area of residence, in a population-based case-control study in Costa Rica. Participation rates were 97% for cases and 90% for controls. All subjects completed a physical activity questionnaire that included occupational and leisure time components with specific questions on siesta. Five siesta frequency categories (<1/wk, 1-4/wk, 5-6/wk, daily [> or =1 h and <2 h], and daily [> or =2 h and <3:30 h]) were used to calculate the odds ratio (OR) by multiple logistic regression. RESULTS: Compared to controls, cases were more likely to take daily siestas (44 versus 35%, P = 0. 01), and spend more time per siesta (1:07 +/- 0:04 versus 0:54 +/- 0:04 h:min, P = 0.002). As compared to subjects with the lowest siesta frequency (<1/wk), the OR for MI among those in the highest category was 1.51 (95% CI : 1.02-2.25, P for trend = 0.006). After adjusting for risk factors, lifestyle, and health history the OR across the siesta categories were 1.0, 0.77, 1.28. 1.66, and 1.40 (P for trend = 0.02). CONCLUSIONS: Our data suggest that the practice of daily siesta is associated with increased risk of MI. PMID- 10869315 TI - Socioeconomic differences in fat intake in a middle-aged population: report from the Malmo Diet and Cancer Study. AB - BACKGROUND: The objective was to investigate whether socioeconomic differences in fat intake may explain socioeconomic differences in cardiovascular diseases. METHODS: The Malmo Diet and Cancer Study is a prospective cohort study. The baseline examinations used in the present cross-sectional study were undertaken in 1992-1994. Dietary habits were assessed using a modified diet history method consisting of a 7-day menu book and a 168-item questionnaire. A subpopulation of 11 837 individuals born 1926-1945 was investigated. This study examined high fat intake, defined as >35.9% among men and >34.8% among women (25% quartile limit) of the proportion of the non-alcohol energy intake contributed by fat. The subfractions saturated, mono-unsaturated and poly-unsaturated fatty acids and the P:S ratio (polyunsaturated/saturated fatty acids) were analysed in the same way. The uppermost quartile (75%) of total and subgroup fat intake was also studied. Socioeconomic differences before and after adjustment for low energy reporting (LER), defined as energy intake below 1.2 x Basal Metabolic Rate, were examined. RESULTS: No socioeconomic differences in fat intake were seen between the SES groups, except for self-employed men, and male and female pensioners. Approximately 20% in most SES groups were LER. The LER and body mass index were strongly related. The SES pattern of fat intake remained unchanged after adjustment for age, country of origin and LER in a logistic regression model. The results for the subfractions of fat and the P:S ratio did not principally differ from the total fat results. CONCLUSIONS: This study provides no evidence that fat intake contributes to the inverse socioeconomic differences in cardiovascular diseases. PMID- 10869316 TI - Increased acute myocardial infarction mortality following the 1995 Great Hanshin Awaji earthquake in Japan. AB - BACKGROUND: This study examined the factors affecting mortality from acute myocardial infarction (AMI) following the 1995 Great Hanshin-Awaji earthquake. METHODS: We examined the death certificates of all decedents between January 1994 and December 1996 in 16 municipalities, which covered most of the area affected by the 1995 Great Hanshin-Awaji earthquake. We analysed the extent and duration of the increased mortality from AMI. The standardized mortality ratio (SMR) of AMI was calculated weekly after the earthquake, taking the number of AMI deaths during the same period in 1994 as a reference. The main outcome measures were the number of deaths from AMI (ICD-9 410; ICD-10 I21, I22) in the study area before and after the earthquake, and the weekly SMR after the earthquake. RESULTS: A significant increase in mortality from AMI in the study area as a whole continued for about 8 weeks after the earthquake. There was wide variation amongst the regions with respect to the extent and duration of the increased mortality from AMI. The SMR of AMI showed a positive relationship with the percentage of houses which were completely destroyed, and was almost significant (r = 0.530, P = 0.062). CONCLUSIONS: The duration of increased cardiac mortality after the 1995 Great Hanshin-Awaji earthquake was longer than seen with previous earthquakes. Further analysis to identify the factors affecting cardiac mortality is needed so that we may reduce adverse health effects during the recovery stage following natural disaster. PMID- 10869317 TI - How accurately are height, weight and leg length reported by the elderly, and how closely are they related to measurements recorded in childhood? AB - BACKGROUND: This paper examines (1) the accuracy of self-reported height, leg length and weight in a group of subjects aged 56-78; (2) whether recent measurement of height and weight influences the accuracy of self-reporting and (3) associations between childhood and adult height, leg length and BMI measured in old age. METHODS: All 3182 surviving members of the Boyd Orr cohort were sent postal questionnaires in 1997-1998 and a sub-sample (294) was also clinically examined. RESULTS: Self-reported height was overestimated and body mass index (BMI), based on reported height and weight, underestimated. The mean difference between self-report and measured values were for height: 2.1 cm in males and 1.7 cm in females; for BMI the difference was -1.3 kg/m(2) in males and -1.2 kg/m(2) in females. Shorter individuals and older subjects tended to over-report their height more than others. The overweight under-reported their weight to a greater extent. Recent measurement appeared to decrease over-reporting of height but not weight. Correlations between self-report and measured height and BMI were generally over 0.90, but weaker for leg length (r = 0.70 in males and 0.71 in females). Adult height and leg length were quite closely related to their relative values in childhood (correlation coefficients ranged from 0.66 to 0.84), but associations between adult and childhood BMI were weak (r = 0.19 in males and 0.21 in females). CONCLUSIONS: Self-reported measures of height and weight may be used in studies of the elderly although systematic reporting errors may bias effect estimates. As overweight individuals tend to under-report and the short and underweight tend to over-report, studies investigating associations of disease with height and weight using self-reported measures will underestimate effects. The weak associations between childhood and adult BMI indicate that associations between childhood adiposity and adult cardiovascular disease found in this cohort may reflect the specific effect of childhood overweight, rather than its persistence into adulthood. This suggests that avoidance of adiposity may be as important in childhood as in adulthood. PMID- 10869318 TI - Decreasing mortality from pulmonary embolism in the United States, 1979-1996. AB - BACKGROUND: Mortality trends and patterns for pulmonary embolism may yield clues to its aetiology. Previous investigations had identified several mortality contrasts in pulmonary embolism mortality among US residents. These findings had been made in the context of a trend of increasing rates during 1962-1984. METHODS: Annual age-specific and age-adjusted pulmonary embolism mortality rates for US residents during 1979-1996 were compiled from the US National Center for Health Statistics web site. These data were analysed for mortality contrasts and trends. RESULTS: For all racial-gender groups, age-adjusted mortality declined throughout the period. The greatest rate of decline was found among black men, followed by (in decreasing order) black women, white men, other men, white women, and other women. In 1996, the previously observed demographic contrasts of blacks experiencing the highest pulmonary embolism mortality, followed by whites and then others, and the male rate being higher than the female one were still present despite this decline. Age-specific mortality from pulmonary embolism in the US during 1996 also mirrored that reported for the 1970s, with mortality increasing during the life span (the risk of death doubling with each decade of life). Specifically, the annual age-adjusted pulmonary embolism mortality rate in 1996 for white men was 2.4 per 100 000 persons; white women, 2.3 per 100 000 persons; black men, 6.0 per 100 000 persons; black women, 4.8 per 100 000 persons; non-black non-white men, 1.0 per 100 000 persons; and non-black non white women, 0.7 per 100 000 persons. CONCLUSIONS: Mortality from pulmonary embolism in the US declined significantly during 1979-1996. Several demographic contrasts, particularly an excess among men, continue to exist. PMID- 10869319 TI - Country of birth, acculturation status and abdominal obesity in a national sample of Mexican-American women and men. AB - BACKGROUND: Few studies have examined the influence of country of birth and acculturation status on indicators of obesity using national samples of Mexican American women and men. METHODS: We analysed data for 1387 Mexican-American women and 1404 Mexican- American men, ages 25-64, from the third National Health and Nutrition Examination Survey (1988-1994). We examined whether waist circumference and abdominal obesity varied by country of birth and acculturation status (primary language spoken), and whether among those with abdominal obesity, number of associated cardiovascular disease (CVD) risk factors varied by country of birth and acculturation status. RESULTS: Both country of birth and, to a lesser degree, acculturation status were significantly associated with waist circumference and abdominal obesity. Mexican-born women and men had the smallest waist circumference (90.4 cm, 94.0 cm respectively), US-born English-speaking women and men had intermediate waist circumference (93.6 cm, 97.3 cm), and US born Spanish-speaking women and men had the largest waist circumference (96.9 cm, 97.7 cm), after accounting for age, education, per cent of energy from dietary fat, leisure-time physical activity, and smoking. All women had high prevalences of abdominal obesity, particularly US-born Spanish-speaking women (68.7%). In addition, US-born Spanish-speaking women with abdominal obesity were significantly more likely than their counterparts to have one or more of the following CVD risk factors: high serum insulin, non-insulin dependent diabetes, high blood lipids, and/or hypertension. CONCLUSIONS: These findings illustrate the heterogeneity of the Mexican-American population and suggest that country of birth and lack of acculturation to the majority culture, as well as secondary lifestyle changes, may explain the significant clinical differences observed in abdominal obesity within Mexican-American population subgroups. PMID- 10869320 TI - Locomotor disability in a cohort of British men: the impact of lifestyle and disease. AB - BACKGROUND: Increasing life expectancy has brought public health concern about the increase in prevalence of disability in old age. Reducing the prevalence of disability in older age requires the identification of preventable or modifiable risk factors earlier in life. We have examined the relationship between lifestyle and other potential risk factors in men aged 40-59 years at screening and locomotor disability 12-14 years later to assess whether any of these factors have direct and independent roles in influencing disability in later life. METHODS: In 1978-1980, a longitudinal study of cardiovascular disease was initiated in 7735 men aged 40-59 years drawn from one general practice in each of 24 British towns. The present study concerns 5717 men, 88% of the surviving men who were available to follow-up (i.e. were registered with a GP and had an address) and who satisfactorily completed the disability section of a follow-up postal questionnaire in 1992 (Q92). The main endpoint from the questionnaire was locomotor disability based on self-reported inability in any one or more of the following: to get outdoors, walk 400 m, climb stairs, maintain balance, bend down, or straighten up. RESULTS: In the 5717 men (mean age 63 years) who provided information on disability status, 25.0% reported locomotor disability and the majority of these men recalled a doctor-diagnosed disease of which cardiovascular disease was most strongly associated with locomotor disability. Lifestyle factors at screening (smoking, physical inactivity, obesity and heavy drinking) and manual social class were strongly and independently associated with increased odds of locomotor disability 12-14 years later. By contrast, baseline blood pressure and serum total cholesterol showed little relationship with locomotor disability. Among men with diagnosed major cardiovascular disease (stroke, myocardial infarction, angina or aortic aneurysm) those with locomotor disability showed significantly higher adverse lifestyle factors at screening than those who were able. Similarly, adverse lifestyle factors were also seen more frequently among disabled men with respiratory disease and among disabled men with other non cardiovascular conditions than among their able counterparts. CONCLUSIONS: Smoking, obesity, physical inactivity and heavy drinking in middle age are strong predictors of locomotor disability in later life independent of the presence of diagnosed disease. Leading a healthy lifestyle improves survival and reduces the incidence of disease. It also reduces the risk of locomotor disability and increases the odds of being disability-free even in the event of developing major cardiovascular disease. PMID- 10869321 TI - Manual work as predictor for disability pensioning with osteoarthritis among the employed in Norway 1971-1990. AB - BACKGROUND: Manual work is reported to be a risk factor for becoming a disability pensioner due to osteoarthritis. This association may be due to covariation with other variables. We wanted to assess if manual work remained a risk factor after adjusting for number of hours worked, income, level of education, gender and marital status, and if the risk associated with manual work was equal in the 1970s and the 1980s. METHODS: In a prospective study, data on all new disability pensioners with osteoarthritis in Norway during the two follow-up periods, 1971 1980 and 1981-1990, were analysed by logistic regression. The study include data on all subjects living in Norway and registered as 50-56 years old and employed either in the census collected in 1970 or in the census of 1980. RESULTS: Manual workers have nearly twice the probability of becoming a disability pensioner with osteoarthritis compared to professionals after adjusting for part-time work, income, level of education, marital status and gender. Adjusted for other risk factors, the probability of becoming a disability pensioner with osteoarthritis was three times higher in the 1980s compared to the 1970s. CONCLUSION: The relatively strong association between manual work and disability pensioning with osteoarthritis suggests difficulties in adjusting manual work patterns for a person with osteoarthritis, which may have increased during the study period as implied by the separate effect of the 1980s. PMID- 10869322 TI - Comparing Swedish hospital discharge records with death certificates: implications for mortality statistics. AB - BACKGROUND: The quality of mortality statistics is of crucial importance to epidemiological research. Traditional editing techniques used by statistical offices capture only obvious errors in death certification. In this study we match Swedish hospital discharge data to death certificates and discuss the implications for mortality statistics. METHODS: Swedish death certificates for 1995 were linked to the national hospital discharge register. The resulting database comprised 69 818 individuals (75% of all deaths), 39 872 (43%) of whom died in hospital. The diagnostic statements were compared at Basic Tabulation List level. RESULTS: The last main diagnosis and the underlying cause of death agreed in 46% of cases. Agreement decreased rapidly after discharge. For hospital deaths, the main diagnosis was reported on 83% of the certificates, but only on 46% of certificates for non-hospital deaths. Malignant neoplasms and other dramatic conditions showed the best agreement and were often reported as underlying causes. Conditions that might follow from some other disease were often reported as contributory causes, while symptomatic and some chronic conditions were often omitted. In 13% of cases, an ill-defined main condition was replaced by a more specific cause of death. CONCLUSIONS: There is no apparent reason to question the death certificate if the main diagnosis and underlying cause agree, or if the main diagnosis is a probable complication of the stated underlying cause. However, cases in which the main diagnosis cannot be considered a complication of the reported underlying cause should be investigated, and assessments made of the feasibility and cost-effectiveness of routinely linking hospital records to death certificates. PMID- 10869323 TI - Economic development and traffic accident mortality in the industrialized world, 1962-1990. AB - BACKGROUND: We examined the association between prosperity and traffic accident mortality in the industrialized world in a long-term perspective. METHODS: We calculated traffic accident mortality, traffic mobility and the fatal injury rate of 21 industrialized countries from 1962 until 1990. We used mortality and population data of the World Health Organization (WHO), and figures on motor vehicle ownership of the International Road Federation (IRF). We examined cross sectional and longitudinal associations of these traffic-related variables with the prosperity level per country, derived from data of the Organization for Economic Cooperation and Development (OECD). RESULTS: We found a reversal from a positive relation between prosperity and traffic accident mortality in the 1960s to a negative association currently. At a certain level of prosperity, the growth rate of traffic mobility decelerates and the fatal injury rate continues to decline at a similar rate to earlier phases. CONCLUSIONS: In a long-term perspective, the relation between prosperity and traffic accident mortality appears to be non-linear: economic development first leads to a growing number of traffic-related deaths, but later becomes protective. Prosperity growth is not only associated with growing numbers of motor vehicles in the population, but also seems to stimulate adaptation mechanisms, such as improvements in the traffic infrastructure and trauma care. PMID- 10869324 TI - A multivariate method for estimating mortality rates among children under 5 years from health and social indicators in Iraq. AB - BACKGROUND: Many reports on Iraq suggest that a rise in rates of death and disease have occurred since the Gulf War of January/February 1991 and the economic sanctions that followed it. METHODS: Four preliminary models, based on unadjusted projections, were developed. A logistic regression model was then developed on the basis of six social variables in Iraq and comparable information from countries in the State of the World's Children report. Missing data were estimated for this model by a multiple imputation procedure. The final model depends on three socio-medical indicators: adult literacy, nutritional stunting of children under 5 years, and access to piped water. RESULTS: The model successfully predicted both the mortality rate in 1990, under stable conditions, and in 1991, following the Gulf War. For 1996, after 5 years of sanctions and prior to receipt of humanitarian food via the oil for food programme, this model shows mortality among children under 5 to have reached an estimated 87 per 1000, a rate last experienced more than 30 years ago. CONCLUSIONS: Accurate and timely estimates of mortality levels in developing countries are costly and require considerable methodological expertise. A rapid estimation technique like the one developed here may be a useful tool for quick and efficient estimation of mortality rates among under 5 year olds in countries where good mortality data are not routinely available. This is especially true for countries with complex humanitarian emergencies where information on mortality changes can guide interventions and the social stability to use standard demographic methods does not exist. PMID- 10869325 TI - Childhood deaths and injuries in Finland in 1971-1995. AB - BACKGROUND: This study examined the recent nationwide trends for the absolute number and the age- and sex-specific incidence rates of the fatal and serious non fatal injuries among 0-14 year old children in Finland in 1971-1995. METHODS: We selected from Official Cause-of-Death Statistics and National Hospital Discharge Register children aged 0-14 years who died or required treatment at a hospital department because of an injury in 1971-1995. The number of Finnish children was 1.1 million in 1971, and 1.0 million in 1995. RESULTS: During the entire study period injuries were the leading cause of death in children aged 1-14 years, but not in infants. However, in these years the incidence (per 100 000 people) of fatal injuries in Finnish children decreased considerably in all age groups and both sexes, in girls from 20.1 in 1971 to 4.6 in 1995, and in boys from 36.7 in 1971 to 9.3 in 1995. In 1995, 41% of all the injurious deaths among 0-14 year old Finnish children were motor vehicle accidents, 12% were drownings, and 24% intentional injuries. The overall number and incidence of serious non-fatal injuries among Finnish children showed no clear trend change in 1971-1995. The mean hospitalization time of injured children shortened between 1971 and 1995, from 7.4 days to 2.7 days. CONCLUSIONS: We conclude that the number and incidence of fatal childhood injuries have decreased dramatically in Finland between 1971 and 1995. The reasons for this positive development are multifactorial, but improved traffic safety and trauma care are probably very important. In children's serious non-fatal injuries the development has not been so encouraging and therefore children's injury prevention should receive continuous intense attention. PMID- 10869326 TI - Erectile dysfunction in type 1 and type 2 diabetics in Italy. On behalf of Gruppo Italiano Studio Deficit Erettile nei Diabetici. AB - BACKGROUND: Several studies reported data on the increased risk of erectile dysfunction (ED) in populations of diabetic men, but few presented data separately for Type 1 and Type 2 subjects. No comparison data for these diabetic subgroups are available with regard to risk factors for ED. METHODS: Eligible for the study were men aged 20-69 years with a diagnosis of insulin-dependent (Type 1) or non-insulin-dependent (Type 2) diabetes who were observed on randomly selected days in 178 diabetes centres in Italy. Erectile dysfunction was defined as a failure to achieve and maintain an erection sufficient for satisfactory sexual performance. RESULTS: The study population consisted of 1383 Type 1 and 8373 Type 2 men. The prevalence of ED increased with age for both groups. After taking into account the effect of age Type 2 men (37/100 men) tend to report ED less frequently than Type 1 men (51/100 men). A significant positive relationship was reported between ED and poor metabolic control and smoking for both Type 1 and Type 2 men, whereas high body mass index (BMI) increased only the risk of ED in Type 1 cases. CONCLUSIONS: The study offers a quantitative estimate of the prevalence of ED and its main risk factors in Type 1 and Type 2 diabetic subgroups. PMID- 10869327 TI - Effect of Sardinian heritage on risk and age at onset of type 1 diabetes: a demographic case-control study of Sardinian migrants. AB - BACKGROUND: Children of Sardinian heritage are at high risk of type 1 diabetes, whereas no data are available in young adults. Age at onset of type 1 diabetes could be associated with different relative weight of genetic susceptibility and environmental determinants in the pathogenesis of the disease. We test this hypothesis in subjects with Sardinian heritage 0-29 years of age living in the city of Turin, a highly industrialized area in Northern Italy. METHODS: In all, 202 cases with onset of type 1 diabetes aged 0-29 years during 1984-1991 and 1010 controls randomly selected from residents of the city of Turin, frequency-matched by sex and year of birth to cases, were included in this study. Name and place of birth of parents were ascertained by postal inquiry and linkage with city population and census files. Social class was based on the highest educational level of parents abstracted from 1991 and 1981 census files. RESULTS: Differential effects on risk of type 1 diabetes of Sardinian heritage and social class in the age groups 0-14 and 15-29 years were found. In children with one and both Sardinian parents the odds ratios (OR) were 2.09 (95% CI : 0.85-5.15) and 3.20 (95% CI : 0.75-13.64); in young adults 0.81 (95% CI : 0.18-3.64) and 1.95 (95% CI : 0.51-7.40), respectively. In subjects with low social class the OR were 1.16 (95% CI : 0.68-1.97) in children and 0.66 (95% CI : 0.41-1.05) in young adults. CONCLUSIONS: This study shows higher risk of type 1 diabetes in subjects of Sardinian heritage; higher risk in children than in young adults and a protective effect of low social class in young adults. These findings are consistent with the hypothesis of heterogeneity of type 1 diabetes by age at onset, with prevailing genetic effect in childhood and environmental determinants in adulthood. PMID- 10869328 TI - How many data sources are needed to determine diabetes prevalence by capture recapture? AB - BACKGROUND: Capture-recapture (CR) methods are increasingly used to estimate the size of human populations, including those with diabetes. Few studies have examined the demographic details needed to match patients on the lists used in these techniques, or to determine the optimum number of lists. METHODS: Six lists of known diabetic patients attending different medical settings during the study year were obtained. The effects on total enumeration after aggregation of these lists were examined using increasing numbers of demographic data items as patient identifiers. The CR estimates of prevalence were obtained using 15 different combinations of two lists. Estimates were obtained after log-linear modelling for interdependence between different combinations of three and four lists, and after combining the six available lists into three logical lists. RESULTS: For matching patients, adding date of birth to first name and family name as matching criteria increased the total of identified patients from 2500 to 2585 (3% increase), corresponding to a period prevalence of 1.5% (95% CI : 1.41-1.52). Addition of further identifiers, such as partial postcode, only increased the estimate by a further 15 patients (0.5%), and more detailed matching with full postcode introduced uncertainty. The use of two-list CR yielded widely varying estimates of the total diabetic population from 1379 (95% CI : 435-2273) to 9554 (95% CI : 7291-10 983). Log-linear modelling using different combinations of three and four lists produced estimates of 5074 (95% CI : 4417-5947) and 5578 (95% CI : 4918 7081), respectively, after compensating for statistical interdependence between the lists used. The appropriate condensation of six available lists into three lists for modelling yielded estimates of 5492 (95% CI : 4870-6285), corresponding to a CR-adjusted period prevalence of 3.1% (95% CI : 3.03-3.19%). CONCLUSIONS: In a Western population, the only demographic data required for matching patients on lists used for CR methods are first name, family name and date of birth, if unique identifiers such as social security numbers are not available. Two lists alone do not produce reliable data, and at least three lists are needed to allow for modelling for 'dependence' between datasets. The use of more than three lists does not substantially alter the absolute value or confidence of enumeration, and multiple lists (if available) should be condensed into three lists for use in CR calculations. PMID- 10869329 TI - Community-based prevention of perinatal deaths: lessons from nineteenth-century Sweden. AB - BACKGROUND: Perinatal deaths have been more difficult to prevent than infant deaths in low- income countries due to its close relation to poor maternal outcome. The aim of the study was to perform a comprehensive population-based analysis of perinatal mortality in a high mortality setting and to determine the impact of midwifery-assisted home deliveries. METHOD: The study design was a community-based cohort study. In all, 4876 perinatal deaths were recorded among 116 211 newborns in the districts of Sundsvall and Skelleftea in northern Sweden during the years 1831-1899. Relative risks, 95% CI, population attributable proportions and prevented fractions were calculated. RESULTS: The overall perinatal mortality rate was 42.0 per 1000 births. A previous stillbirth represented one of the most important risk factors (RR = 3.25, 95% CI : 2.97 3.56), with a population attributable proportion of 7%. Two or more previous stillbirths gave an RR of 8.50 (95% CI : 7.58-9.53) and a population attributable proportion of 4%. There was an increased risk of perinatal mortality for mothers over 35 years old, the primiparous and the unmarried, while grandparous women had a higher perinatal mortality that was accounted for completely by a poor history of previous stillbirths and infant deaths among these women. The children of crofters, farmers and workers had higher perinatal mortality, but area had no significant impact. During the years 1881-1890 and 1891-1899, the prevented fractions of midwifery were 15% and 32%, respectively. CONCLUSION: Poor reproductive history, particularly previously high perinatal mortality, is associated with high perinatal mortality. Midwifery-assisted at home deliveries successfully reduced perinatal mortality. PMID- 10869330 TI - Acute effects of low levels of ambient ozone on peak expiratory flow rate in a cohort of Australian children. AB - BACKGROUND: We enrolled a cohort of primary schoolchildren with a history of wheeze (n = 148) in an 11-month longitudinal study to examine the relationship between ambient ozone concentrations and peak expiratory flow rate. METHODS: Enrolled children recorded peak expiratory flow rates (PEFR) twice daily. We obtained air pollution, meteorological and pollen data. In all, 125 children remained in the final analysis. RESULTS: We found a significant negative association between daily mean deviation in PEFR and same-day mean daytime ozone concentration (beta-coefficient = 0.88; P = 0.04) after adjusting for co pollutants, time trend, meteorological variables, pollen count and ALTERNARIA: count. The association was stronger in a subgroup of children with bronchial hyperreactivity and a doctor diagnosis of asthma (beta-coefficient = -2.61; P = 0.001). There was no significant association between PEFR and same-day daily daytime maximum ozone concentration. We also demonstrated a dose-response relationship with mean daytime ozone concentration. CONCLUSIONS: Moderate levels of ambient ozone have an adverse health effect on children with a history of wheezing, and this effect is larger in children with bronchial hyperreactivity and a doctor diagnosis of asthma. PMID- 10869331 TI - Evaluating the impact of tuberculosis control: number of deaths prevented by short-course chemotherapy in China. AB - BACKGROUND: Tuberculosis (TB) is still amongst the most important causes of human morbidity and mortality, killing approximately two million people each year. Standard short-course chemotherapy (SSCC) can rapidly control illness and dramatically reduce the chance of death, but the impact of treatment has rarely been evaluated in these terms. METHOD: We developed a mathematical model that makes use of routinely-collected data to calculate the number of deaths directly prevented by TB treatment (i.e. excluding those due to reduced transmission). The method was applied to the world's largest TB control programme covering over 500 million people in 12 provinces of China. RESULTS: Counties which had been enrolled in the programme since 1991 were, by 1997, preventing at least 46% (37 56%) of the TB deaths that would otherwise have occurred. If replicated across the entire TB control programme area, this would amount to 30 000 (range 26 000 59 000) deaths directly prevented each year. CONCLUSIONS: Short-course chemotherapy has substantially reduced TB mortality in half of China. The analytical method described here could be applied to TB control operations in many other countries, and should help to quantify the true burden of tuberculosis alleviated by SSCC. PMID- 10869333 TI - Predictive value of the HIV paediatric classification system for the long-term course of perinatally infected children. AB - BACKGROUND: To compare the Centers for Disease Control and Prevention (CDC) paediatric classification system with the long-term course of perinatal human immunodeficiency virus type 1 (HIV-1) infection. METHODS: Prospective study on 366 perinatally infected children followed-up from birth and checked at least every 2 months. Survival, smoothed hazard, adjusted hazard ratio of death, and transition probabilities in clinical and immunological categories were outcome measures. RESULTS: Survival was 49% (95% CI : 40-58%) at 8 years. The risk of death was high before the age of 2, relatively low between ages 2 and 7, and contained thereafter. Children did not advance through the categories sequentially. Survival at 8 years was 61.7% (95% CI : 49.8-73.6%) in those children who had passed through clinical category A; the hazard ratio of death was 2.5 (95% CI : 1.7-3.8) for 175 (47.9%) children who skipped this category. Transition probability in clinical category B was 39.9% (95% CI : 32.3-45.6%) after one year, but 59.1% (95% CI : 51.4-66.8%) after 5 years. Before 2 years of age, the probability of entry into category C (40%; 95% CI : 35-45%) was higher than that of entry into immunological category 3 (28%; 95% CI : 22-34%). Conclusions The classification system stands comparison with the clinical reality, but the CD4-positive cell thresholds in infancy should be adjusted and category B indicator diseases better distributed to improve their predictive value. PMID- 10869332 TI - Secular trends in the survival of HIV-infected homosexual men in Amsterdam and Vancouver estimated from a death-included CD4-staged Markov model. AB - BACKGROUND: The purpose of this study was to investigate secular trends in waiting times in CD4-based stages of human immunodeficiency virus (HIV) disease progression in two cohorts of homosexual men, one in Vancouver and one in Amsterdam. All HIV-positive men with two or more CD4 counts in their AIDS-free period between 1 January 1985 and 1 January 1997 were included in this study. Data regarding clinical AIDS diagnoses (using the 1987 Centers for Disease Control and Prevention [CDC] AIDS case definition) and death were collected through active follow-up, review of hospital records, and municipal/national registries. The Vancouver Lymphadenopathy-AIDS Study (VLAS), was started in November 1982 and had enrollment until December 1984. Both HIV-negative and HIV positive men were followed at intervals of 3-6 months until 1986 and annually thereafter. The Amsterdam cohort study on HIV and AIDS (ACS) started in December 1984, has ongoing enrollment and follow-up of both HIV-negative and HIV-positive homosexual men. The HIV-positive men were followed at intervals of 3 months. METHODS: The CD4-based stage of an individual at each visit was determined using smoothed data. For each cohort and in each calendar time period, a CD4-based Markov model with death as the absorbing stage was fitted to the data. The parameters in this model were estimated using the method of maximum likelihood and confidence intervals were calculated using bootstrap methods. RESULTS: A total of 509 homosexual men participating in the VLAS were included in this study, providing 5356 visits. Some 292 men developed AIDS before 1 January 1997 and 239 died before this date. In all, 232 of the 239 deaths were AIDS related. Thirty-seven per cent of all visits were related to treatment. A total of 543 homosexual men participating in the ACS were included in this study, providing 10 043 visits; 277 men developed AIDS before 1 January 1997 and 250 died before this date. The date of AIDS diagnosis was known for 225 of the 250 deaths. Twenty per cent of all visits were related to treatment. We found that in both cohort studies the stage-specific waiting times were longer in the low CD4-based stages (stages 4, 5 and 6: i.e. CD4 count <500 cells per mm(3)) after March 1990 compared to waiting times before March 1990. The increase in mean waiting time in these stages with low CD4 count was 21%, 33% and 53%, respectively in the ACS and 20%, 2% and 29% in the VLAS. Because waiting times alone are not exclusive for progression in a reversible model we also calculated the stage-specific median incubation periods till death. Men spent considerably longer in these CD4-based stages after March 1990 compared to before March 1990. CONCLUSIONS: Data from these population-based cohort studies showed that HIV disease progression in the calendar period where treatment was administered was slower for individuals in stages with low CD4 counts. We found no evidence for shortening of the incubation period that may have appeared from increasing virulence of the HIV in the population. PMID- 10869334 TI - Assessment of different sources of variation in the antibody responses to specific malaria antigens in children in Papua New Guinea. AB - BACKGROUND: A potential problem for malaria vaccine development and testing is between-host variation in antibody responses to specific malaria antigens. Previous work in adults in an area highly endemic for Plasmodium falciparum in Papua New Guinea found that genetic regulation partly explained heterogeneity in responsiveness. We have now assessed the relative contributions of environmental and genetic factors in total IgG responses to specific malaria antigens in children, and quantified temporal variation within individuals of total IgG responses. METHODS: Total IgG responses against schizont extract, merozoite surface protein-1, merozoite surface protein-2, ring-infected erythrocyte surface antigen, and SPf66 were measured by ELISA. Variance component analysis was used to estimate the variation explained by genetic and environmental factors in these antibody responses. Intra- and inter-class correlations of antibody responses within relative pairs were estimated. We adjusted for age, P. falciparum density, sex and village differences either within or prior to the analysis. RESULTS: For all malaria antigens, temporal variation in the total IgG response was the predominant source of variation. There was substantial familial aggregation of all IgG responses, but it remained unclear how much this clustering was attributable to genetic factors and how much to a common environment in the household. The remaining variance, which could not be explained by either of the above, was very small for most of the antigens. CONCLUSIONS: Temporal variation and clustering of immune responses to specific malaria antigens need to be taken into account when planning, conducting and interpreting immuno-epidemiological and vaccine studies. PMID- 10869335 TI - Human cytomegalovirus seroprevalence in three socioeconomically different urban areas during the first trimester: a population-based cohort study. AB - BACKGROUND: To re-evaluate the impact of socioeconomic status and human cytomegalovirus (HCMV) seroprevalence during pregnancy, we carried out a population-based cohort study. METHODS: IgG and IgM antibodies to HCMV and IgG avidity were studied by enzyme-linked immunosorbent assay (ELISA) in three different socioeconomic areas (SEA) in the 9-12th week of pregnancy of 1088 consecutive mothers. RESULTS: The overall IgG seropositivity was 70.7%, ranging from 60.9 to 76.4% in 'upper' to 'lower' SEA (P = 0.0004). The HCMV IgM seropositivity was 4.0%, ranging from 3.8% in the 'upper' and 'intermediate' SEA to 4.6% in the 'lower' SEA. Serologically acute cases, defined by low avidity of IgG, represented 1.7% of the pregnancies in the 'upper' SEA compared with 1.0 and 1.1% in the other two areas. In the 'lower' SEA there were twice as many recurrent infections as in the others, 3.6 versus 1.7%. The low impact of age did not increase after elimination of the effects of SEA and parity. Miscarriages were associated neither with IgG nor with IgM positivity, although the percentage of >/=2 miscarriages was 8.8% in seronegative women compared with 11.2% and 13.6% in IgG- and IgM-positive women. CONCLUSIONS: Social environment seems to be the most powerful factor, predicting both IgG seroprevalence and recurrences during pregnancy. PMID- 10869336 TI - Helicobacter pylori infection and subsequent peptic duodenal disease among young adults. AB - BACKGROUND: Evidence for a causal relationship between presence of Helicobacter pylori (H. pylori) in gastric mucosa and development of peptic disease is based largely on intervention studies in which eradication of H. pylori led to healing of the lesion. The aim of this study was to assess the importance of H. pylori seropositivity for subsequent development of peptic disease in a prospective study design in young Israelis. METHODS: A nested case-control serum bank study based on a systematic sample of male and female inductees to the Israel Defense Force. Twenty-nine cases of duodenal ulcer or duodenitis of moderate or higher severity, diagnosed between 1986 and 1995, were individually matched for age, sex, ethnicity, education and year of induction, with five healthy controls each. Presence of anti-H. pylori antibodies in the frozen stored sera was determined by ELISA. RESULTS: The geometric mean titre of anti-H. pylori antibodies at baseline was significantly higher in cases (18. 3 U/ml) than controls (6.9 U/ml; P = 0.009). The matched odds ratio for peptic ulcer disease by seropositivity was 3.8 (95% CI : 1.4-10. 2). A stronger association was evident for subjects diagnosed > or =2 years after induction than those diagnosed earlier. The population attributable fraction was 56.6% (95% CI : 15.7-81.1). CONCLUSIONS: Pre-existing infection with H. pylori, as determined by seropositivity, is an important determinant of development of duodenal ulcer or duodenitis in young Israelis, supporting the generalizability of an apparent causal association to diverse populations. PMID- 10869337 TI - European stillbirth proportion and Chernobyl. PMID- 10869338 TI - Topical antibiotic use and circumcision-associated neonatal tetanus: protective factor or indicator of good wound care? PMID- 10869339 TI - Amyloid beta -peptide-binding alcohol dehydrogenase is a component of the cellular response to nutritional stress. AB - Amyloid beta-peptide-binding alcohol dehydrogenase (ABAD) is a member of the family of short chain dehydrogenase/reductases whose distinctive properties include the capacity to bind amyloid beta-peptide and enzymatic activity toward a broad array of substrates including n-isopropanol and beta-estradiol. In view of the wide substrate specificity of ABAD and its high activity on l-beta hydroxyacyl-CoA derivatives, we asked whether it might also catalyze the oxidation of the ketone body d-3-hydroxybutyrate. This was indeed the case, and oxidation proceeded with K(m) of approximately 4.5 mm and V(max) of approximately 4 nmol/min/mg protein. When placed in medium with d-beta-hydroxybutyrate as the principal energy substrate, COS cells stably transfected to overexpress wild-type ABAD (COS/wtABAD) better maintained 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide reduction, cellular energy charge, and morphologic phenotype compared with COS/vector cells. Using a severe model of metabolic perturbation, transgenic mice with targeted neuronal expression of ABAD subjected to transient middle cerebral artery occlusion showed strokes of smaller volume and lower neurologic deficit scores in parallel with increased brain ATP and decreased lactate, compared with nontransgenic controls. These data suggest that ABAD contributes to the protective response to metabolic stress, especially in the setting of ischemia. PMID- 10869340 TI - Novel alternative splicing and nuclear localization of human RGS12 gene products. AB - RGS proteins are GTPase-activating proteins for certain Galpha subunits, accelerating the shutoff mechanism of G protein signaling, and also may interact with receptors and effectors to modulate G protein signaling. Here, we report identification of 12 distinct transcripts of human RGS12 that arise by unusually complex splicing of the RGS12 gene, which spans 70 kilobase pairs of genomic DNA and contains 16 exons. These transcripts arise by both cis- and trans-splicing mechanisms, are expressed in a tissue-specific manner, and encode proteins ranging in size from 356 to 1447 amino acids. Both 5'- and 3'-splicing of two primary RGS12 transcripts occur to generate RGS12 mRNAs encoding proteins with four distinct N-terminal domains, three distinct C-terminal domains, and a common internal region where the semiconserved RGS domain is located. Confocal microscopy and subcellular fractionation of COS-7 cells expressing RGS12 proteins with three different N termini (brain (B), peripheral (P), and trans-spliced (TS)) and a shared short (S) C-terminal domain demonstrated exclusive nuclear localization of these proteins and an influence of the N-terminal region on the pattern of intranuclear distribution. Both native RGS12TS-S in HEK-293T cells and ectopically expressed RGS12TS-S localized to discrete nuclear foci (dots), a characteristic of various tumor suppressor proteins. Subnuclear localization of RGS12TS-S into nuclear dots was cell cycle-dependent. Native RGS12TS-S associated with the metaphase chromosome during mitosis, and ectopically expressed RGS12TS-S induced formation of abnormally shaped and multiple nuclei in COS-7 cells. Expression of RGS12 proteins with long and intermediate C-terminal domains was not observed in COS-7 cells, suggesting that 3'-splicing of RGS12 transcripts may influence the expression or stability of the encoded proteins. These results document extraordinary structural complexity in the RGS12 family and the role of alternative splicing and cell cycle-dependent mechanisms in expression and subnuclear targeting of RGS12 proteins. PMID- 10869341 TI - GAP1IP4BP contains a novel group I pleckstrin homology domain that directs constitutive plasma membrane association. AB - The group I family of pleckstrin homology (PH) domains are characterized by their inherent ability to specifically bind phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) and its corresponding inositol head-group inositol 1,3,4,5 tetrakisphosphate (Ins(1,3,4,5)P(4)). In vivo this interaction results in the regulated plasma membrane recruitment of cytosolic group I PH domain-containing proteins following agonist-stimulated PtdIns(3,4,5)P(3) production. Among group I PH domain-containing proteins, the Ras GTPase-activating protein GAP1(IP4BP) is unique in being constitutively associated with the plasma membrane. Here we show that, although the GAP1(IP4BP) PH domain interacts with PtdIns(3,4, 5)P(3), it also binds, with a comparable affinity, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) (K(d) values of 0.5 +/- 0.2 and 0.8 +/- 0.5 microm, respectively). Intriguingly, whereas this binding site overlaps with that for Ins(1,3,4,5)P(4), consistent with the constitutive plasma membrane association of GAP1(IP4BP) resulting from its PH domain-binding PtdIns(4,5)P(2), we show that in vivo depletion of PtdIns(4,5)P(2), but not PtdIns(3,4,5)P(3), results in dissociation of GAP1(IP4BP) from this membrane. Thus, the Ins(1,3,4,5)P(4) binding PH domain from GAP1(IP4BP) defines a novel class of group I PH domains that constitutively targets the protein to the plasma membrane and may allow GAP1(IP4BP) to be regulated in vivo by Ins(1,3,4,5)P(4) rather than PtdIns(3,4,5)P(3). PMID- 10869342 TI - Analysis of the adenosylcobinamide kinase/adenosylcobinamide-phosphate guanylyltransferase (CobU) enzyme of Salmonella typhimurium LT2. Identification of residue His-46 as the site of guanylylation. AB - CobU is a bifunctional enzyme involved in adenosylcobalamin (coenzyme B(12)) biosynthesis in Salmonella typhimurium LT2. In this bacterium, CobU is the adenosylcobinamide kinase/adenosylcobinamide-phosphate guanylyltransferase needed to convert cobinamide to adenosylcobinamide-GDP during the late steps of adenosylcobalamin biosynthesis. The guanylyltransferase reaction has been proposed to proceed via a covalently modified CobU-GMP intermediate. Here we show that CobU requires a nucleoside upper ligand on cobinamide for substrate recognition, with the nucleoside base, but not the 2'-OH group of the ribose, being important for this recognition. During the kinase reaction, both the nucleotide base and the 2'-OH group of the ribose are important for gamma phosphate donor recognition, and GTP is the only nucleotide competent for the complete nucleotidyltransferase reaction. Analysis of the ATP:adenosylcobinamide kinase reaction shows CobU becomes less active during this reaction due to the formation of a covalent CobU-AMP complex that holds CobU in an altered conformation. Characterization of the GTP:adenosylcobinamide-phosphate guanylyltransferase reaction shows the covalent CobU-GMP intermediate is on the reaction pathway for the generation of adenosylcobinamide-GDP. Identification of a modified histidine and analysis of cobU mutants indicate that histidine 46 is the site of guanylylation. PMID- 10869343 TI - Lipoxin A4 antagonizes the mitogenic effects of leukotriene D4 in human renal mesangial cells. Differential activation of MAP kinases through distinct receptors. AB - The lipoxygenase-derived eicosanoids leukotrienes and lipoxins are well defined regulators of hemeodynamics and leukocyte recruitment in inflammatory conditions. Here, we describe a novel bioaction of lipoxin A(4) (LXA(4)), namely inhibition of leukotriene D(4) (LTD(4))-induced human renal mesangial cell proliferation, and investigate the signal transduction mechanisms involved. LXA(4) blocked LTD(4)-stimulated phosphatidylinositol 3-kinase (PI 3-kinase) activity in parallel to inhibition of LTD(4)-induced mesangial cell proliferation. Screening of a human mesangial cell cDNA library revealed expression of the recently described cys-leukotriene(1)/LTD(4) receptor. LTD(4)-induced mesangial cell proliferation required both extracellular-related signal regulated kinase (erk) and PI 3-kinase activation and may involve platelet-derived growth factor receptor transactivation. LTD(4)-stimulated the MAP kinases erk and p38 via a pertussis toxin (PTX)-sensitive pathway dependent on PI 3-kinase and protein kinase C activation. On screening a cDNA library, mesangial cells were found to express the previously described LXA(4) receptor. In contrast to LTD(4), LXA(4) showed differential activation of erk and p38. LXA(4) activation of erk was insensitive to PTX and PI 3-kinase inhibition, whereas LXA(4) activation of p38 was sensitive to PTX and could be blocked by the LTD(4) receptor antagonist SKF 104353. These data suggest that LXA(4) stimulation of the MAP kinase superfamily involves two distinct receptors: one shared with LTD(4) and coupled to a PTX sensitive G protein (G(i)) and a second coupled via an alternative G protein, such as G(q) or G(12), to erk activation. These data expand on the spectrum of LXA(4) bioactions within an inflammatory milieu. PMID- 10869344 TI - A novel RalGEF-like protein, RGL3, as a candidate effector for rit and Ras. AB - The small GTPase Rit is a close relative of Ras, and constitutively active Rit can induce oncogenic transformation. Although the effector loops of Rit and Ras are highly related, Rit fails to interact with the majority of the known Ras candidate effector proteins, suggesting that novel cellular targets may be responsible for Rit transforming activity. To gain insight into the cellular function of Rit, we searched for Rit-binding proteins by yeast two-hybrid screening. We identified the C-terminal Rit/Ras interaction domain of a protein we have designated RGL3 (Ral GEF-like 3) that shares 35% sequence identity with the known Ral guanine nucleotide exchange factors (RalGEFs). RGL3, through a C terminal 99-amino acid domain, interacted in a GTP- and effector loop-dependent manner with Rit and Ras. Importantly, RGL3 exhibited guanine nucleotide exchange activity toward the small GTPase Ral that was stimulated in vivo by the expression of either activated Rit or Ras. These data suggest that RGL3 functions as an exchange factor for Ral and may serve as a downstream effector for both Rit and Ras. PMID- 10869345 TI - Neutrophil tissue inhibitor of matrix metalloproteinases-1 occurs in novel vesicles that do not fuse with the phagosome. AB - The human neutrophil granule location of precursors of matrix metalloproteinases (MMPs), MMP-8 and -9, has been established, but that of the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) has not. In this study, labeling for TIMP-1, pro-MMP-8, pro-MMP-9, and established granule marker proteins reveals that TIMP-1 is mainly located in distinct oval, electron translucent organelles, a little larger than azurophil granules. A lack of labeling for the fluid phase endocytic marker, bovine serum albumin-gold, the lysosome-associated membrane protein markers, and for glycosylphosphatidylinositol-linked proteins, which are enriched in secretory vesicles, indicates the non-endosomal, non-lysosomal, and non secretory nature of this organelle. Density gradient cofractionation with the least dense, secretory population and some pleomorphism of the organelle suggest it is a "vesicle" rather than a "granule" population. Colocalization with pro-MMP 9 or pro-MMP-8, in minor subpopulations, suggests that TIMP-1 vesicle biogenesis occurs between metamyelocytic and terminal differentiation and before secretory vesicle synthesis. Pulse-chased IgG-coated latex beads and immunolabeling show that specific and azurophil granules fuse with the phagosome whereas TIMP-1 and pro-MMP-9-containing organelles do not. This suggests that these play no role in phagosomal destruction of IgG-opsonized bacteria. Separate localization and colocalization of these proteins may, however, facilitate fine regulation of extracellular proteolysis. PMID- 10869346 TI - The long signal peptide isoform and its alternative processing direct the intracellular trafficking of interleukin-15. AB - Two isoforms of interleukin (IL)-15 exist: one with a short and another with a long signal peptide (LSP). Experiments using combinations of the LSP and mature proteins IL-2, IL-15, and green fluorescent protein revealed complex pathways of intracellular trafficking. In one pathway, the LSP was unprocessed, and IL-15 was not glycosylated, remained in the cytoplasm, and was degraded. The second trafficking pathway involved endoplasmic reticulum entry, N-linked glycosylation, and alternative partial LSP processing. The third pathway involved endoplasmic reticulum entry, followed by glycosylation, complete processing, and ultimately secretion. The complex intracellular trafficking patterns of LSP-IL-15 with its impediments to secretion as well as impediments to translation may be required due to the potency of IL-15 as an inflammatory cytokine. In terms of a more positive role, we propose that intracellular infection may relieve the burdens on translation and intracellular trafficking to yield effective IL-15 expression. PMID- 10869347 TI - Inhibition of insulin-induced glucose uptake by atypical protein kinase C isotype specific interacting protein in 3T3-L1 adipocytes. AB - Atypical protein kinase C (PKC) isotype-specific interacting protein (ASIP) specifically interacts with the atypical protein kinase C isozymes PKClambda and PKCzeta. ASIP and atypical PKC, as well as their Caenorhabditis elegans counterparts (PAR-3 and PKC-3, respectively), are thought to coordinately participate in intracellular signaling that contributes to the maintenance of cellular polarity and to the formation of junctional complexes. The potential role of ASIP in other cellular functions of atypical PKC was investigated by examining the effect of overexpression of ASIP on insulin-induced glucose uptake, previously shown to be mediated through PKClambda, in 3T3-L1 adipocytes. When overexpressed in these cells, which contain PKClambda but not PKCzeta, ASIP was co-immunoprecipitated with endogenous PKClambda but not with PKCepsilon or with Akt. The subcellular localization of PKClambda was also altered in cells overexpressing ASIP. Overexpression of ASIP inhibited insulin stimulation of both glucose uptake and translocation of the glucose transporter GLUT4 to the plasma membrane, but it did not inhibit glucose uptake induced by either growth hormone or hyperosmolarity both of which promote glucose uptake in a PKClambda independent manner. Moreover, glucose uptake stimulated by a constitutively active mutant of PKClambda, but not that induced by an active form of Akt, was inhibited by ASIP. Insulin-induced activation of PKClambda, but not that of phosphoinositide 3-kinase or Akt, was also inhibited by overexpression of ASIP. These data suggest that overexpression of ASIP inhibits insulin-induced glucose uptake by specifically interfering with signals transmitted through PKClambda. PMID- 10869348 TI - Nonsaturable binding indicates clustering of tau on the microtubule surface in a paired helical filament-like conformation. AB - Tau protein modulates microtubule dynamics and forms insoluble aggregates in Alzheimer's disease. Because there is a discrepancy between reported affinities of Tau to microtubules, we determined the interaction over a wide concentration range using a sensitive enzyme-linked immunosorbent assay. We found that the interaction is biphasic and not monophasic as assumed earlier. The first binding phase is typical for identical and noninteracting binding sites, with dissociation constants around 0.1 micrometer and stoichiometries around 0.2 Tau/tubulin dimer. Surprisingly, the second phase is nonsaturable and shows a nearly linear increase in bound Tau versus free Tau for free Tau concentrations higher than 2 micrometer. The slope is proportional to the microtubule concentration. From this we define an overloading parameter with values around 50 micrometer. The influence of Tau isoform, phosphorylation, and dimerization on both phases was investigated. Remarkably the overloading of Tau on microtubules leads to a thioflavin S fluorescence increase reminiscent of that seen with Tau aggregated into Alzheimer paired helical filaments. Because polyanions stimulate Tau aggregation and because the C-terminal domain of tubulin is polyanionic, we suggest that an early conformational change in Tau leading to paired helical filament aggregation occurs right on the microtubule surface. PMID- 10869349 TI - Persistent tumor necrosis factor signaling in normal human fibroblasts prevents the complete resynthesis of I kappa B-alpha. AB - Transcription factor NF-kappa B is normally sequestered in the cytoplasm, complexed with I kappa B inhibitory proteins. Tumor necrosis factor (TNF) and interleukin-1 induce I kappa B-alpha phosphorylation, leading to I kappa B-alpha degradation and translocation of NF-kappa B to the nucleus where it activates genes important in inflammatory and immune responses. TNF and interleukin-1 actions are typically terminated by desensitization, and I kappa B-alpha reappearance normally occurs within 30-60 min. We found that in normal human FS-4 fibroblasts maintained in the presence of TNF, I kappa B-alpha protein failed to return to base-line levels for up to 15 h. Removal of TNF at any time during the 15-h period resulted in complete I kappa B-alpha resynthesis, suggesting that I kappa B-alpha reappearance was prevented by continued TNF signaling. Long term exposure of FS-4 fibroblasts to TNF led to a persistent presence of I kappa B alpha mRNA, sustained I kappa B kinase activation, continuous proteasome-mediated degradation of I kappa B-alpha, and sustained nuclear localization of NF-kappa B. Continuous exposure of FS-4 cells to TNF did not lead to a sustained activation of p38 or ERK mitogen-activated protein kinases, suggesting that not all TNF induced signaling pathways are persistently activated. These findings challenge the notion that all cytokine-mediated signals are rapidly terminated by desensitization and illustrate the need to elucidate the process of deactivation of TNF-induced signaling. PMID- 10869350 TI - Dimerization with retinoid X receptors and phosphorylation modulate the retinoic acid-induced degradation of retinoic acid receptors alpha and gamma through the ubiquitin-proteasome pathway. AB - In eukaryotic cells, the ubiquitin-proteasome pathway is the major mechanism for targeted degradation of proteins. We show that, in F9 cells and in transfected COS-1 cells, the nuclear retinoid receptors, retinoic acid receptor gamma2 (RARgamma2), RARalpha1, and retinoid X receptor alpha1 (RXRalpha1) are degraded in a retinoic acid-dependent manner through the ubiquitin-proteasome pathway. The degradation of RARgamma2 is entirely dependent on its phosphorylation and on its heterodimerization with liganded RXRalpha1. In contrast, RARalpha1 degradation can occur in the absence of heterodimerization, whereas it is inhibited by phosphorylation, and heterodimerization reverses that inhibition. RXRalpha1 degradation is also modulated by heterodimerization. Thus, each partner of RARgamma/RXRalpha and RARalpha/RXRalpha heterodimers modulates the degradation of the other. We conclude that the ligand-dependent degradation of RARs and RXRs by the ubiquitin-proteasome pathway, which is regulated by heterodimerization and by phosphorylation, could be important for the regulation of the magnitude and duration of the effects of retinoid signals. PMID- 10869351 TI - The nuclear matrix protein CDP represses hepatic transcription of the human cholesterol-7alpha hydroxylase gene. AB - To date, the molecular mechanisms that govern hepatic-specific transcription of the human cholesterol 7alpha-hydroxylase (CYP7A1) gene are poorly understood. We recently reported that the region extending from -1888 to +46, which includes the promoter, is not capable of conferring expression to human CYP7A1 promoter lacZ transgenes in the livers of mice, but that expression is observed with transgenes containing the entire structural gene. To locate liver-specific elements in other segments of the human gene, DNase I hypersensitivity studies were performed with transcriptionally active, liver-derived HepG2 cells and with transcriptionally inactive HeLa cells. Three DNase I hypersensitivity sites were detected within the first intron of the human CYP7A1 gene, but only in HepG2 cells. Transient transfection experiments with HepG2 cells revealed a transcriptional repressor within intron 1. Five binding sites for the CAAT displacement protein (CDP) were detected within intron 1. Since CDP is a nuclear matrix protein, two methods were employed to localize nuclear matrix attachment sites within intron 1 of the human CYP7A1 gene. A matrix attachment site was found throughout the entirety of intron 1. Gel retardation experiments and cell transfection studies provided evidence for the repression mechanism. Repression is achieved by displacement by CDP of two hepatic activators, namely HNF-1alpha and C/EBPalpha, that bind to three different sites within intron 1. Additionally, CDP represses transactivation mediated by these two activators. PMID- 10869352 TI - Activation of extracellular-regulated kinase pathways in ovarian granulosa cells by the novel growth factor type 1 follicle-stimulating hormone receptor. Role in hormone signaling and cell proliferation. AB - Follicle-stimulating hormone (FSH) regulated growth and function of the ovarian follicle was previously thought to be mediated solely through activation of G(s) coupled receptors. In this study, we show for the first time that this function is predominantly mediated through the alternatively spliced and novel growth factor type 1 receptor (oFSH-R3) that is also present in the ovary. Immortalized granulosa cells lacking endogenous FSH receptors, when transfected with either oFSH-R3 cDNA (JC-R3) or the G(s)-coupled oFSH-R1 (JC-R1), expressed the corresponding glycosylated receptor. In JC-R3 or JC-R1 cells labeled with bromodeoxyuridine or [(3)H]thymidine, FSH stimulated the cells to progress through S-phase and divide. The growth promoting effect of recombinant FSH in JC R3 cells was preceded by the rapid activation of ERK1 and ERK2. This effect was hormone-specific and transient. In JC-R3 cells inhibitors like calphostin C, PD98059, Ag 18, or calcium chelators EGTA or 1,2-bis(O-aminophenoxy)ethane N,N,N',N'-tetraacetic acid/AM inhibited both mitogen-activated protein kinase activation and bromodeoxyuridine incorporation. FSH induced phosphorylation of the FSH-R3 receptor was blocked by pretreating cells with calphostin C. There was no cAMP induction by FSH in JC-R3 cells. The cAMP independent growth promoting effect of FSH is mediated by activation of Ca(2+) and mitogen-activated protein kinase-dependent pathways. Thus, alternative splicing of a G-protein coupled receptor creates the expression of a novel receptor motif that can mediate a widely recognized function of the glycoprotein hormone. PMID- 10869353 TI - Heterodimeric Pbx-Prep1 homeodomain protein binding to the glucagon gene restricting transcription in a cell type-dependent manner. AB - Homeodomain proteins specify developmental pathways and cell-specific gene transcription whereby proteins of the PBC subclass can direct target gene specificity of Hox proteins. Proteins encoded by nonclustered homeobox genes have been shown to be essential for cell lineage differentiation and gene expression in pancreatic islets. Using specific antiserum in an electrophoretic mobility shift assay and in vitro transcribed/translated proteins, the nuclear proteins binding domain B of the G3 enhancer-like element of the glucagon gene were identified in the present study as heterodimers consisting of the ubiquitously expressed homeodomain protein Prep1 and the also widely expressed PBC homeoprotein Pbx (isoform 1a, 1b, or 2). These heterodimeric complexes were found to bind also to the glucagon cAMP response element and to a newly identified element termed G5 (from -169 to -140). Whereas the expression of Prep1 or Pbx forms alone had no effect, coexpression of Pbx1a/1b-Prep1 inhibited the glucagon promoter when activated by cotransfected Pax6 or another transcription factor in non-glucagon-producing cells. In contrast, in glucagon-producing pancreatic islet cells, Pbx-Prep1 had no effect on GAL4-Pax6-induced mutant glucagon promoter activity or on Pax6-dependent wild-type glucagon promoter activity. Furthermore, 5'-deletion of G5 enhanced glucagon promoter activity in a non-glucagon-producing cell line but not in glucagon-producing islet cells. This study thus identifies a novel target and Hox-independent function of Pbx-Prep1 heterodimers that, through repression of glucagon gene transcription in non-glucagon-producing cells, may help to establish islet cell-specific expression of the glucagon gene. PMID- 10869354 TI - Regulation of prostaglandin E2 biosynthesis by inducible membrane-associated prostaglandin E2 synthase that acts in concert with cyclooxygenase-2. AB - Here we report the molecular identification of membrane-bound glutathione (GSH) dependent prostaglandin (PG) E(2) synthase (mPGES), a terminal enzyme of the cyclooxygenase (COX)-2-mediated PGE(2) biosynthetic pathway. The activity of mPGES was increased markedly in macrophages and osteoblasts following proinflammatory stimuli. cDNA for mouse and rat mPGESs encoded functional proteins that showed high homology with the human ortholog (microsomal glutathione S-transferase-like 1). mPGES expression was markedly induced by proinflammatory stimuli in various tissues and cells and was down-regulated by dexamethasone, accompanied by changes in COX-2 expression and delayed PGE(2) generation. Arg(110), a residue well conserved in the microsomal GSH S transferase family, was essential for catalytic function. mPGES was functionally coupled with COX-2 in marked preference to COX-1, particularly when the supply of arachidonic acid was limited. Increased supply of arachidonic acid by explosive activation of cytosolic phospholipase A(2) allowed mPGES to be coupled with COX 1. mPGES colocalized with both COX isozymes in the perinuclear envelope. Moreover, cells stably cotransfected with COX-2 and mPGES grew faster, were highly aggregated, and exhibited aberrant morphology. Thus, COX-2 and mPGES are essential components for delayed PGE(2) biosynthesis, which may be linked to inflammation, fever, osteogenesis, and even cancer. PMID- 10869355 TI - Probing the catalytic mechanism of the insulin receptor kinase with a tetrafluorotyrosine-containing peptide substrate. AB - The interaction of a synthetic tetrafluorotyrosyl peptide substrate with the activated tyrosine kinase domain of the insulin receptor was studied by steady state kinetics and x-ray crystallography. The pH-rate profiles indicate that the neutral phenol, rather than the chemically more reactive phenoxide ion, is required for enzyme-catalyzed phosphorylation. The pK(a) of the tetrafluorotyrosyl hydroxyl is elevated 2 pH units on the enzyme compared with solution, whereas the phenoxide anion species behaves as a weak competitive inhibitor of the tyrosine kinase. A structure of the binary enzyme-substrate complex shows the tetrafluorotyrosyl OH group at hydrogen bonding distances from the side chains of Asp(1132) and Arg(1136), consistent with elevation of the pK(a). These findings strongly support a reaction mechanism favoring a dissociative transition state. PMID- 10869356 TI - Activation of the pro-drug ethionamide is regulated in mycobacteria. AB - The anti-tuberculosis drug ethionamide (ETH), which is a structural analog of isoniazid (INH), is known to strongly inhibit mycolic acid synthesis in Mycobacterium tuberculosis. Although several targets have been identified for INH, only speculative information is available concerning ETH. Mutations within the promoter and the coding region of enoyl-acyl carrier protein reductase (InhA) were found to confer resistance to both drugs, thus leading to the impression that INH and ETH may share a common mode of action. However, a notable distinction between the two drugs lies in the lack of cross-resistance in clinical isolates. This may be attributed in part to the fact that the pro-drug INH must be activated via KatG, and no activation step for ETH has yet been described. Here we report the identification of an activator for ETH. The ETH activator (Rv3854c), which we have termed EthA, was found to be homologous to various monooxygenases and induced ETH sensitivity when overexpressed in mycobacteria. Interestingly, the neighboring open reading frame (Rv3855), which was found homologous to transcriptional repressors of the tetR family, led to ETH resistance when overexpressed. In addition, chromosomal inactivation of this gene by transposition led to ETH hypersensitivity. These data strongly suggest that Rv3855, which we have termed EthR, regulates the production of EthA, which subsequently activates the pro-drug ETH. This study opens up new avenues of research relating to ETH activation in mycobacteria, possibly leading to an improved efficacy of ETH and to the generation of new anti-mycobacterial agents. PMID- 10869357 TI - Substrate specificity in the highly heterogeneous M4 peptidase family is determined by a small subset of amino acids. AB - The members of the M4 peptidase family are involved in processes as diverse as pathogenicity and industrial applications. For the first time a number of M4 family members, also known as thermolysin-like proteases, has been characterized with an identical substrate set and a uniform set of assay conditions. Characterization with peptide substrates as well as high performance liquid chromatography analysis of beta-casein digests shows that the M4 family is a homogeneous family in terms of catalysis, even though there is a significant degree of amino acid sequence variation. The results of this study show that differences in substrate specificity within the M4 family do not correlate with overall sequence differences but depend on a small number of identifiable amino acids. Indeed, molecular modeling followed by site-directed mutagenesis of one of the substrate binding pocket residues of the thermolysin-like proteases of Bacillus stearothermophilus converted the catalytic characteristics of this variant into that of thermolysin. PMID- 10869358 TI - Sterol 27-hydroxylase acts on 7-ketocholesterol in human atherosclerotic lesions and macrophages in culture. AB - 27-Hydroxycholesterol (27OH) is the major oxysterol in human atherosclerotic lesions, followed by 7-ketocholesterol (7K). Whereas 7K probably originates nonenzymically, 27OH arises by the action of sterol 27-hydroxylase, a cytochrome P450 enzyme expressed at particularly high levels in the macrophage and proposed to represent an important pathway by which macrophages eliminate excess cholesterol. We hypothesized and here show that 27-hydroxylated 7-ketocholesterol (270H-7K) is present in human lesions, probably generated by the action of sterol 27-hydroxylase on 7K. Moreover, [(3)H]27OH-7K was produced by human monocyte derived macrophages (HMDMs) supplied with [(3)H]7K but not in HMDMs from a patient with cerebrotendinous xanthomatosis (CTX) shown to have a splice-junction mutation of sterol 27-hydroxylase. Whereas [(3)H]27OH-7K was predominantly secreted into the medium, [(3)H]-27OH formed from [(3)H]-cholesterol was mostly cell-associated. The majority of supplied [(3)H]7K was metabolized beyond 27OH-7K to aqueous-soluble products (apparently bile acids derived from the sterol 27 hydroxylase pathway). Metabolism to aqueous-soluble products was ablated by a sterol 27-hydroxylase inhibitor and absent in CTX cells. Sterol 27-hydroxylase therefore appears to represent an important pathway by which macrophages eliminate not only cholesterol but also oxysterols such as 7K. The fact that 7K (and cholesterol) still accumulates in lesions and foam cells indicates that this pathway may be perturbed in atherosclerosis and affords a new opportunity for the development of therapeutic strategies to regress atherosclerotic lesions. PMID- 10869359 TI - Negative regulation of the serine/threonine kinase B-Raf by Akt. AB - B-Raf contains multiple Akt consensus sites located within its amino-terminal regulatory domain. One site, Ser(364), is conserved with c-Raf but two additional sites, Ser(428) and Thr(439), are unique to B-Raf. We have investigated the role of both the conserved and unique phosphorylation sites in the regulation of B-Raf activity in vitro and in vivo. We show that phosphorylation of B-Raf by Akt occurs at multiple residues within its amino-terminal regulatory domain, at both the conserved and unique phosphorylation sites. The alteration of the serine residues within the Akt consensus sites to alanines results in a progressive increase in enzymatic activity in vitro and in vivo. Furthermore, expression of Akt inhibits epidermal growth factor-induced B-Raf activity and inhibition of Akt with LY294002 up-regulates B-Raf activity, suggesting that Akt negatively regulates B-Raf in vivo. Our results demonstrate that B-Raf activity can be negatively regulated by Akt through phosphorylation in the amino-terminal regulatory domain of B-Raf. This cross-talk between the B-Raf and Akt serine/threonine kinases is likely to play an important role in modulating the signaling specificity of the Ras/Raf pathway and in promoting biological outcome. PMID- 10869360 TI - Regulation of vesicle trafficking in madin-darby canine kidney cells by Rab11a and Rab25. AB - Polarized epithelial cells maintain the polarized distribution of basolateral and apical membrane proteins through a process of receptor-mediated endocytosis, sorting, and then recycling to the appropriate membrane domain. We have previously shown that the small GTP-binding proteins, Rab11a and Rab25, are associated with the apical recycling system of Madin-Darby canine kidney cells. Here we have utilized inducible expression of wild-type, dominant negative, and constitutively active mutants to directly compare the functions of Rab25 and Rab11a in postendocytic vesicular transport. We found that a Rab11a mutant deficient in GTP binding, Rab11aS25N, potently inhibited both transcytosis and apical recycling yet failed to inhibit transferrin recycling. Similarly, expression of either wild type Rab25 or the active mutant Rab25S21V inhibited both apical recycling and transcytosis of IgA by greater than 50% but had no effect on basolateral recycling of transferrin. Interestingly, the GTPase deficient mutant Rab11aS20V inhibited basolateral to apical transcytosis of IgA, but had no effect on either apical or basolateral recycling. These results indicate that neither Rab11a nor Rab25 function in the basolateral recycling of transferrin in polarized Madin-Darby canine kidney cells cells, consistent with recent morphological observations by others. Thus, transferrin receptors must be recycled to the plasma membrane prior to sorting of apically directed cargoes into Rab11a/Rab25-positive apical recycling endosomes. PMID- 10869361 TI - Characterization of the murMN operon involved in the synthesis of branched peptidoglycan peptides in Streptococcus pneumoniae. AB - The murMN operon, recently identified in the genome of Streptococcus pneumoniae, encodes for enzymes involved in the synthesis of branched structured muropeptides in the pneumococcal peptidoglycan; inactivation of murMN causes production of a peptidoglycan composed exclusively of linear muropeptides and a virtually complete loss of resistance in penicillin-resistant strains (Filipe, S. R., and Tomasz, A. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 4891-4896). The experiments described in this paper follow up these observations. Primer extension analysis was used to identify the putative promoter region of the murMN operon in penicillin-susceptible and -resistant strains. Selective inactivation of the murN gene in the penicillin-resistant strain Pen6 caused production of an unusual peptidoglycan that contained only single amino acid residues in the muropeptide branches, indicating that the product of murN was involved with the addition of the second amino acid and the product of murM was involved with the addition of the first amino acid (alanine or serine) to the peptidoglycan cross-bridge. Allelic replacement of the mosaic murM gene of strain Pen6 with murM of the penicillin-susceptible laboratory strain caused enrichment of the peptidoglycan in linear muropeptides. The findings suggest that the genetic determinant primarily controlling the synthesis of branched muropeptides in the pneumococcal peptidoglycan is murM. PMID- 10869362 TI - Flux control of cytochrome c oxidase in human skeletal muscle. AB - In the present work, by titrating cytochrome c oxidase (COX) with the specific inhibitor KCN, the flux control coefficient and the metabolic reserve capacity of COX have been determined in human saponin-permeabilized muscle fibers. In the presence of the substrates glutamate and malate, a 2.3 +/- 0.2-fold excess capacity of COX was observed in ADP-stimulated human skeletal muscle fibers. This value was found to be dependent on the mitochondrial substrate supply. In the combined presence of glutamate, malate, and succinate, which supported an approximately 1.4-fold higher rate of respiration, only a 1.4 +/- 0.2-fold excess capacity of COX was determined. In agreement with these findings, the flux control of COX increased, in the presence of the three substrates, from 0.27 +/- 0.03 to 0.36 +/- 0.08. These results indicate a tight in vivo control of respiration by COX in human skeletal muscle. This tight control may have significant implications for mitochondrial myopathies. In support of this conclusion, the analysis of skeletal muscle fibers from two patients with chronic progressive external ophthalmoplegia, which carried deletions in 11 and 49% of their mitochondrial DNA, revealed a substantially lowered reserve capacity and increased flux control coefficient of COX, indicating severe rate limitations of oxidative phosphorylation by this enzyme. PMID- 10869363 TI - Molecular basis for the omega-regiospecificity of the CYP4A2 and CYP4A3 fatty acid hydroxylases. AB - The CYP4A fatty acid omega-hydroxylases are involved in important physiological processes such as the regulation of vascular pressure. A previous study of chimeras of the rat CYP4A2 and CYP4A3 enzymes established that the regiochemistry of fatty acid hydroxylation is determined by the first 119 amino acid residues (Hoch, U., Zhang. Z. P., Kroetz, D. L., and Ortiz de Montellano, P. R. (2000) Arch. Biochem. Biophys. 373, 63-71). The role of the individual amino acid differences in this region has therefore been examined by site-specific mutagenesis to determine which residues actually control the omega- versus (omega 1)-regiospecificity. The results indicate that regiospecificity is controlled by the presence or absence of a three-residue insert (Ser-114, Gly-115, Ile-116) in CYP4A3 and by the residue at position 119 (CYP4A3 numbering). Furthermore, analysis of the absolute stereochemistry of the 11-hydroxylauric acid product indicates that this stereochemistry is not very sensitive to changes in the residues that line the substrate access channel. These results define a model of the specificity determinants of an important class of cytochrome P450 enzymes. PMID- 10869364 TI - Role of acyl-coenzyme A:cholesterol acyltransferase-1 in the control of hepatic very low density lipoprotein secretion and low density lipoprotein receptor expression in the mouse and hamster. AB - Cholesteryl esters present in nascent very low density lipoproteins are generated in a reaction catalyzed by acyl CoA:cholesterol acyltransferase (ACAT). To examine the effect of cholesteryl esters on the secretion of apoB-containing lipoproteins, we transiently overexpressed human (h) ACAT-1 in the livers of low density lipoprotein (LDL) receptor(-/-) mice using adenovirus-mediated gene transfer. Overexpression of hACAT-1 increased hepatic total and esterified cholesterol but did not reduce hepatic free cholesterol due to a compensatory increase in the rate of de novo cholesterol synthesis. Overexpression of hACAT-1 markedly increased the plasma concentration and hepatic secretion of apoB containing lipoproteins but had no effect on the clearance of very low density lipoprotein-apoB from plasma indicating that cholesteryl esters play an important role in regulating the assembly and secretion of apoB-containing lipoproteins. ACAT activity has been implicated in the regulation of the LDL receptor pathway by dietary fatty acids. It has been hypothesized that unsaturated fatty acids, by enhancing ACAT activity, reduce the amount of free cholesterol in a putative regulatory pool that feeds back on LDL receptor expression. We directly tested this hypothesis in hamsters by transiently overexpressing hACAT-1 in the liver. Enhanced cholesterol esterification in the liver resulted in a compensatory increase in de novo cholesterol synthesis but no induction of LDL receptor expression suggesting that fatty acids regulate LDL receptor expression via a mechanism independent of ACAT. PMID- 10869365 TI - Molecular organization of the alkali-insoluble fraction of Aspergillus fumigatus cell wall. AB - Physical and biological properties of the fungal cell wall are determined by the composition and arrangement of the structural polysaccharides. Cell wall polymers of fungi are classically divided into two groups depending on their solubility in hot alkali. We have analyzed the alkali-insoluble fraction of the Aspergillus fumigatus cell wall, which is the fraction believed to be responsible for fungal cell wall rigidity. Using enzymatic digestions with recombinant endo-beta-1,3 glucanase and chitinase, fractionation by gel filtration, affinity chromatography with immobilized lectins, and high performance liquid chromatography, several fractions that contained specific interpolysaccharide covalent linkages were isolated. Unique features of the A. fumigatus cell wall are (i) the absence of beta-1,6-glucan and (ii) the presence of a linear beta-1, 3/1,4-glucan, never previously described in fungi. Galactomannan, chitin, and beta-1,3-glucan were also found in the alkali-insoluble fraction. The beta-1,3-glucan is a branched polymer with 4% of beta-1,6 branch points. Chitin, galactomannan, and the linear beta-1, 3/1,4-glucan were covalently linked to the nonreducing end of beta-1, 3 glucan side chains. As in Saccharomyces cerevisiae, chitin was linked via a beta 1,4 linkage to beta-1,3-glucan. The data obtained suggested that the branching of beta-1,3-glucan is an early event in the construction of the cell wall, resulting in an increase of potential acceptor sites for chitin, galactomannan, and the linear beta-1,3/1,4-glucan. PMID- 10869366 TI - The distribution of P2X receptor clusters on individual neurons in sympathetic ganglia and their redistribution on agonist activation. AB - The distribution of P2X receptors on neurons in rat superior cervical ganglia and lability of P2X receptors on exposure to agonists were determined. Antibody labeling of each P2X subtype P2X(1)-P2X(7) showed neurons isolated into culture possessed primarily P2X(2) subunits with others occurring in order P2X(7) > P2X(6) > P2X(3) > P2X(1) > P2X(5) > P2X(4). Application of ATP and alpha,beta meATP to neurons showed they possessed a predominantly nondesensitizing P2X receptor type insensitive to alpha,beta-meATP, consistent with immunohistochemical observations. P2X(1)-green fluorescent protein (GFP) was used to study the time course of P2X(1) receptor clustering in plasma membranes of neurons and internalization of receptors following prolonged exposure to ATP. At 12-24 h after adenoviral infection, P2X(1)-GFP formed clusters about 1 microm diameter in the neuron membrane. Application of ATP and alpha,beta-meATP showed these neurons possessed a predominantly desensitizing P2X receptor type sensitive to alpha,beta-meATP. Infection converted the major functional P2X receptor type in the membrane to P2X(1). Exposure of infected neurons to alpha,beta-meATP for less than 60 s led to the disappearance of P2X(1)-GFP fluorescence from the cell surface that was blocked by monensin, indicating the chimera is normally endocytosed into these organelles on exposure to agonist. PMID- 10869367 TI - Combinatorial chemistry and contemporary pharmacology. AB - Both solid- and liquid-phase combinatorial chemistry have emerged as powerful tools for identifying pharmacologically active compounds and optimizing the biological activity of a lead compound. Complementary high-throughput in vitro assays are essential for compound evaluation. Cell-based assays that use optical endpoints permit investigation of a wide variety of functional properties of these compounds including specific intracellular biochemical pathways, protein protein interactions, and the subcellular localization of targets. Integration of combinatorial chemistry with contemporary pharmacology now represents an important factor in drug discovery and development. PMID- 10869368 TI - Mechanisms and sites of ocular action of 7-hydroxy-2-dipropylaminotetralin: a dopamine(3) receptor agonist. AB - The purpose of this study was to investigate mechanism(s) and site(s) of action involved in 7-hydroxy-2-dipropylaminotetralin (7-OH-DPAT)-induced ocular hypotension. As measured by pneumatonometry, the topical, unilateral application of 7-OH-DPAT (75 microg), a dopamine D(3)-preferring receptor agonist, decreased the intraocular pressure (IOP) bilaterally. The ocular hypotensive activity of 7 OH-DPAT was diminished in sympathetically denervated rabbits. Pretreatment with raclopride, a D(2)/D(3) receptor antagonist; UH232, a D(3) receptor antagonist; or U-99194A, a D(3) receptor antagonist antagonized 7-OH-DPAT-induced ocular hypotension. However, pretreatment with spiperone, a D(2) receptor antagonist, did not affect the 7-OH-DPAT-induced ocular hypotension. In addition, topically applied 7-OH-DPAT caused a reduction of aqueous humor flow rate. To examine sites of action, immunohistochemistry of D(3) dopamine receptors was performed. Dopamine D(3) receptors were found to be present on postganglionic sympathetic nerves in the ciliary body of normal rabbits but were virtually undetectable in the same tissue of sympathectomized rabbits. In summary, the IOP-lowering effect caused by 7-OH-DPAT was due, in part, to the suppression of aqueous humor flow. Immunohistochemical identification of D(3) receptors in the ciliary body, associated with the diminution of IOP-lowering effects by D(3) receptor agonist 7 OH-DPAT in sympathetically denervated rabbits provided evidence of neuronal site of action of 7-OH-DPAT. Suppression of 7-OH-DPAT-induced ocular hypotension by D(3) receptor antagonists (U-99194A and UH232) and sympathectomy, coupled with the immunohistochemical data, suggested that the primary site of D(3) receptor mediated action of 7-OH-DPAT is located on postganglionic sympathetic nerve endings in the ciliary body of rabbit. PMID- 10869369 TI - Expression and localization of multidrug resistant protein mrp2 in rat small intestine. AB - The expression of multidrug resistance-associated protein isoform 2 (mrp2), the ATP-dependent export pump that mediates the transport of glucuronic acid-, glutathione-, and sulfate-conjugated derivatives, was studied in rat small intestine. The small intestine was divided into nine equal segments, and mrp2 content was analyzed in homogenate and brush border membrane preparations by Western analysis. mrp2 protein was present mainly in brush border membrane of the proximal segments and gradually decreased from jejunum to the distal ileum. We also analyzed the content of mrp2 in three different populations of proximal enterocytes obtained from the upper and lower villus and the crypt regions. The export pump was mainly expressed in the villus cells and to a lesser degree in the crypt cells of the epithelium. Immunohistochemical analysis performed in duodenum, jejunum, and ileum confirmed in situ the Western blot findings. Analysis of mRNA encoding mrp2 in proximal and distal segments revealed a similar content in both regions, whereas distribution along the villus-crypt axis was similar to the protein gradient. Because conjugating enzymes are distributed similarly to mrp2, we conclude that they may act coordinately to contribute to first-pass metabolism of drugs and other xenobiotics in the proximal small intestine. PMID- 10869370 TI - Effect of ozone treatment on airway reactivity and epithelium-derived relaxing factor in guinea pigs. AB - Ozone (O(3)) is toxic to respiratory epithelium and causes airway inflammation and hyperreactivity. To evaluate the role of the epithelium in the development of hyperreactivity, we examined in guinea pigs the effects of inhaled O(3) (3 ppm for 1 h; 0-24 h after exposure) on 1) reactivity to inhaled methacholine (MCh), 2) reactivity of the isolated, perfused trachea (IPT) to MCh, 3) epithelium derived relaxing factor (EpDRF)-mediated relaxations of IPT induced by mucosal hyperosmolar solutions, 4) neurogenic contraction and relaxation responses, 5) transepithelial potential difference, and 6) microscopic analysis of nitrotyrosine immunofluorescence, substance P fiber density, and tracheal morphology. At 0 h, O(3) caused hyperreactivity to inhaled MCh and mucosally but not serosally applied MCh in IPT (only in the presence of the epithelium) and a decrease in transepithelial potential difference. Inhibition of EpDRF-induced relaxation responses occurred at 2 h. All of these changes returned to control by 12 to 18 h. O(3) had no effect on neurogenic responses. Nitrotyrosine immunofluorescence appeared in the trachea at 0 h in detached epithelial cell ghosts and in intrapulmonary airways by 6 h. Substance P fiber density was elevated in smooth muscle at 0 and 18 h but not in epithelium or lamina propria of intrapulmonary and extrapulmonary bronchi. Loss of cilia and mucosubstances in the mucosa occurred at 0 h; the epithelium became markedly attenuated over 12 to 24 h. A reversible increase in epithelial permeability and a decrease in EpDRF production may contribute to O(3)-induced hyperreactivity to MCh. PMID- 10869371 TI - A highly conserved aspartic acid (Asp-155) anchors the terminal amine moiety of tryptamines and is involved in membrane targeting of the 5-HT(2A) serotonin receptor but does not participate in activation via a "salt-bridge disruption" mechanism. AB - Discovering the molecular and atomic mechanism(s) by which G-protein-coupled receptors (GPCRs) are activated by agonists remains an elusive goal. Recently, studies examining two representative GPCRs (rhodopsin and alpha(1b)-adrenergic receptors) have suggested that the disruption of a putative "salt-bridge" between highly conserved residues in transmembrane (TM) helix III, involving aspartate or glutamate, and helix VII, involving a basic residue, results in receptor activation. We have tested whether this is a general mechanism for GPCR activation by constructing a model of the 5-hydroxytryptamine (5-HT)(2A) receptor and characterizing several mutations at the homologous residues (Asp-155 and Asn 363) of the 5-HT(2A) serotonin receptor. All of the mutants (D155A, D155N, D155E, D155Q, and S363A) resulted in receptors with reduced basal activity; in no case was evidence for constitutive activity revealed. Structure-function studies with tryptamine analogs and various Asp-155 mutants demonstrated that Asp-155 interacts with the terminal, and not indole, amine moiety of 5-HT(2A) agonists. Interestingly, the D155E mutation interfered with the membrane targeting of the 5 HT(2A) receptor, and an inverse relationship was discovered when comparing receptor activation and targeting for a series of Asp-155 mutants. This represents the first known instance in which a charged residue located in a putative TM helix alters the membrane targeting of a GPCR. Thus, for 5-HT(2A) receptors, the TMIII aspartic acid (Asp-155) is involved in anchoring the terminal amine moiety of indole agonists and in membrane targeting and not in receptor activation by salt-bridge disruption. PMID- 10869372 TI - Geometry and charge determine pharmacological effects of steroids on N-methyl-D aspartate receptor-induced Ca(2+) accumulation and cell death. AB - Modulation of N-methyl-D-aspartate (NMDA) receptor function by a series of sulfated steroids and dicarboxylic acid ester analogs of pregnenolone sulfate and pregnanolone sulfate was investigated in cultured hippocampal neurons. The "bent" steroid ring structure associated with 5beta-stereochemistry favors receptor inhibition, whereas the more planar ring structure of the pregn-5-enes and 5alpha pregnanes favors potentiation of NMDA-induced [Ca(2+)] increases and neuronal cell death. The nature of the negatively charged group attached to the steroid C3 position is important for both the neuroprotection afforded by pregnane steroids and the exacerbation of NMDA-induced neuronal death by pregn-5-enes. Dicarboxylic acid hemiesters of various lengths can substitute for the sulfate group of the positive modulator pregnenolone sulfate and the negative modulator pregnanolone sulfate. This result suggests that precise coordination with the oxygen atoms of the sulfate group is not critical for modulation and that the steroid recognition sites can accommodate bulky substituents at C3. The capacity of charged steroids to enhance or protect against NMDA-induced death of hippocampal neurons is strongly correlated with modulation of NMDA-induced Ca(2+) accumulation, indicating that direct enhancement or inhibition of NMDA receptor function is responsible for the proexcitotoxic or neuroprotective effects of these steroids. PMID- 10869373 TI - Characterization of contractile P2 receptors in human coronary arteries by use of the stable pyrimidines uridine 5'-O-thiodiphosphate and uridine 5'-O-3 thiotriphosphate. AB - The present study was designed to evaluate the relative contribution of the different contractile P2 receptors in endothelium-denuded human coronary arteries by use of extracellular nucleotides, including the stable pyrimidines uridine 5' O-3-thiotriphosphate (UTPgammaS) and uridine 5'-O-thiodiphosphate (UDPbetaS). The isometric tension of isolated vessel segments was recorded in vitro, and P2 receptor mRNA expression was examined by reverse transcription-polymerase chain reaction. alphabeta-Methylene-adenosine triphosphate (alphabeta-MeATP) elicited contractions at a low concentration (pEC(50) = 5.2), indicating the presence of contractile P2X receptors. The P2Y responses were analyzed after P2X receptor desensitization with 10 microM alphabeta-MeATP. The stable nucleotides UTPgammaS and adenosine 5'-O-3-thiotriphosphate (ATPgammaS), which are agonists of P2Y(2) or P2Y(4) receptors, were approximately 2 log units more potent than the endogenous UTP and ATP (pEC(50) = 4.6 and 3.8 for UTPgammaS and ATPgammaS). The efficacy of these responses were approximately double that of the P2X agonist alphabeta-MeATP (E(max) = 125% for UTPgammaS, 126% for ATPgammaS, and 68% for alphabeta-MeATP), suggesting a primary role for contractile P2Y(2/4) receptors. The P2Y(2) receptor agonist diadenosine tetraphosphate also stimulated contraction, whereas the selective P2Y(1) agonist adenosine 5'-O-thiodiphosphate and the selective P2Y(6) agonist UDPbetaS had no effect. Reverse transcription polymerase chain reaction analysis of mRNA from endothelium-denuded human coronary arteries demonstrated strong bands for P2Y(2) and P2X(1), although bands for P2Y(1), P2Y(4), and P2Y(6) receptor mRNA could also be detected. In conclusion, the stable pyrimidines UDPbetaS and UTPgammaS are important tools for P2 receptor subtype characterization in intact tissues with ectonucleotidase activity. Extracellular nucleotides elicit contraction of human coronary arteries primarily by activation of P2Y(2) and P2X receptors, whereas a role for P2Y(1) and P2Y(6) receptors can be excluded. Antagonists of P2Y(2) and P2X receptors may be useful in the treatment of coronary vasospastic disorders. PMID- 10869374 TI - Computational model of intracellular pharmacokinetics of paclitaxel. AB - The intracellular pharmacokinetics of paclitaxel is closely related to its pharmacodynamics. Although drug transport across the cell membrane and extracellular and intracellular drug binding have been shown to affect intracellular drug accumulation, their quantitative relationship is unknown. This study was designed to establish a mathematical model for computing the intracellular paclitaxel pharmacokinetics. As a starting point, the model assumes drug transport into and out of cells via passive diffusion. Experimental data on the intracellular pharmacokinetics of [(3)H]paclitaxel were obtained using monolayer cultures of human breast MCF7 tumor cells, which have negligible expression of the mdr1 P-glycoprotein. The results indicate that, in addition to drug binding and microtubule concentration, changes in cell number due to cell growth and drug effects also affected intracellular drug accumulation. A kinetic model was developed to describe several concomitant processes: 1) saturable drug binding to extracellular proteins, 2) saturable and nonsaturable drug binding to intracellular components, 3) time- and concentration-dependent drug depletion from culture medium, 4) cell density-dependent drug accumulation, and 5) time- and drug concentration-dependent enhancement of tubulin concentration. The model was validated by the close prediction (<7% deviation) of the effects of extracellular-to-intracellular concentration gradient and cell density on the kinetics of drug accumulation and efflux. This model was used to predict the effects of changing several parameters (number and binding affinity of intracellular binding sites, free fraction, and concentration of drug in extracellular fluid) on intracellular drug accumulation. In conclusion, the computational model of intracellular paclitaxel pharmacokinetics provides the means to predict drug concentration in cells. PMID- 10869375 TI - Cloning of rat histamine H(3) receptor reveals distinct species pharmacological profiles. AB - The recent cloning and characterization of the human histamine H(3) receptor cDNA marked a significant step toward a greater understanding of the role of this receptor in the central nervous system. We now report the cloning of the rat histamine H(3) receptor cDNA and demonstrate distinct pharmacological species differences. The rat cDNA clone encodes a putative 445-amino acid protein with 93% identity to the human H(3) receptor. Northern blot analysis revealed a major single entity of 2.7-kb in length expressed only in brain. Transfection of the rat receptor cDNA into SK-N-MC cells conferred an ability to inhibit forskolin stimulated cAMP formation in response to histamine and other H(3) agonists. N [(3)H]methylhistamine saturably bound to transfected cells with high affinity (K(d) = 0.8 nM). All agonists tested had low or subnanomolar K(i) values similar to that for the human recombinant receptor. The antagonists thioperamide and clobenpropit also bound with high affinity (K(i) = 4 and 0.4 nM, respectively). This is in contrast to the antagonist profile obtained for the human recombinant receptor that showed K(i) values of 58 and 0.6 nM for thioperamide and clobenpropit, respectively. These data suggest that the low affinity of thioperamide for the human H(3) receptor represents a species difference in pharmacology and not a unique pharmacological subtype. It also was found that chloroproxyfan behaved as a full agonist at the rat recombinant receptor. These findings highlight the significance of validating the pharmacology of experimental compounds at both the rat and human H(3) receptors. PMID- 10869376 TI - Down-regulation of vascular endothelial growth factor expression after A(2A) adenosine receptor activation in PC12 pheochromocytoma cells. AB - Vascular endothelial growth factor (VEGF) is an endothelial cell mitogen that promotes angiogenesis during embryonic development and the progression of certain pathologies. This study examined the regulation of VEGF expression by adenosine receptor (AR) activation in PC12 rat pheochromocytoma cells. Treatment of cells with the AR agonist CGS21680 reduced the VEGF mRNA level to approximately 20% of that in control cells with an EC(50) value of 0.47 nM, indicative of mediation by the A(2A)AR. Down-regulation of VEGF mRNA by CGS21680 was abolished by pretreatment of cells with the AR antagonist ZM241385. Additionally, ZM241385 alone increased VEGF mRNA by 2.8-fold above basal. RNase protection assays indicated that CGS21680 down-regulated VEGF(121), VEGF(165), and VEGF(189) transcripts. VEGF protein secretion was similarly decreased by CGS21680. Under hypoxic conditions, VEGF mRNA expression was reduced by 85.7% after pretreatment with CGS21680. The down-regulation response appears to be mediated predominately by coupling of the A(2A)AR to G(s) because cholera toxin treatment also reduced VEGF expression. The decrease in VEGF mRNA steady-state levels after A(2A)AR activation is apparently due to a decrease in the VEGF gene transcription rate and not to a decrease in mRNA stability. Thus, depending on the cell type, adenosine may have an inhibitory effect on VEGF production, which may have implications in blood vessel development. PMID- 10869377 TI - Ca(2+) signaling via sigma(1)-receptors: novel regulatory mechanism affecting intracellular Ca(2+) concentration. AB - The sigma(1)-receptor is a one-transmembrane endoplasmic reticulum protein that binds neurosteroids and dextrorotatory benzomorphans. The roles of sigma(1) receptors in regulating intracellular Ca(2+) in NG108 cells were examined in this study. sigma(1)-Ligands pregnenolone sulfate, (+)-pentazocine, and 2-(4 morpholino)ethyl-1-phenylcyclohexane-1-carboxylate hydrochloride modulate Ca(2+) signaling in NG108 cells via two modes of action. First, nanomolar concentrations of the ligands, without effect by themselves, potentiated the bradykinin-induced increase of the cytosolic free Ca(2+) concentration in a bell-shaped manner. This effect of sigma(1)-ligands was unaffected by depletion of Ca(2+) from perfusion buffer and was blocked by a 21-mer antisense oligodeoxynucleotide against the cloned sigma(1)-receptors. Second, after the cells were depleted of the endoplasmic reticulum Ca(2+) stores, the depolarization (75 mM KCl)-induced increase in cytosolic free Ca(2+) was potentiated by 2-(4-morpholino)ethyl-1 phenylcyclohexane-1-carboxylate hydrochloride, whereas it was inhibited by pregnenolone sulfate and (+)-pentazocine. These effects, albeit opposite in direction, were blocked by both the 21-mer antisense oligodeoxynucleotide and pertussis toxin. Western blotting indicates that sigma(1)-receptors are increased on the plasma membrane and the nuclear membrane in the presence of sigma(1) ligand. These results suggest that Ca(2+) signaling via sigma(1)-receptors may represent a novel mechanism that affects intracellular Ca(2+) concentrations. PMID- 10869378 TI - Role of corticotropin-releasing factor (CRF) receptors in the anorexic syndrome induced by CRF. AB - Genetic manipulations of corticotropin-releasing factor (CRF)(1) and CRF(2) receptors have resulted in data suggesting that the CRF(2) receptor could mediate the effects of CRF on appetite or satiety. We have attempted to obtain pharmacological evidence for this hypothesis by comparing the ability of a high affinity peptide, mixed CRF antagonist [cyclo 30-33,f12,L18,21E30, A32,K33]sucker fish urotensin (12-41)NH(2) [cUTSN (12-41)] with a small-molecule CRF(1) selective antagonist, NBI-27914, and a CRF(2)-selective peptide antagonist, antisauvagine-30, to attenuate the anorexic effects of CRF. We also monitored other behaviors that accompanied CRF-induced anorexia. CRF-induced anorexia was significantly correlated with a reduction in locomotor activity and an increase in freezing behavior and piloerection. cUTSN (12-41) and antisauvagine-30 significantly attenuated the effects of CRF (0.04 nmol) on food intake along with the behavioral syndrome that accompanied anorexia. In contrast, NBI-27914 did not attenuate either of the above-mentioned CRF-induced phenomena when given centrally at doses ranging from 0.13 to 10 nmol/2.5 microl or when given orally at 20 to 40 mg/kg. Although these data support the hypothesis that the CRF(2) receptor mediates the appetite suppression induced by CRF, they also suggest that the CRF(2) receptor could mediate the stress-like behaviors that accompany CRF induced appetite suppression. PMID- 10869379 TI - Cannabinoids protect cells from oxidative cell death: a receptor-independent mechanism. AB - Serum is required for the survival and growth of most animal cells. In serum-free medium, B lymphoblastoid cells and fibroblasts die after 2 days. We report that submicromolar concentrations of Delta(9)-tetrahydrocannabinol (THC), Delta(8) THC, cannabinol, or cannabidiol, but not WIN 55,212-2, prevented serum-deprived cell death. Delta(9)-THC also synergized with platelet-derived growth factor in activating resting NIH 3T3 fibroblasts. The cannabinoids' growth supportive effect did not correlate with their ability to bind to known cannabinoid receptors and showed no stereoselectivity, suggesting a nonreceptor-mediated pathway. Direct measurement of oxidative stress revealed that cannabinoids prevented serum-deprived cell death by antioxidation. The antioxidative property of cannabinoids was confirmed by their ability to antagonize oxidative stress and consequent cell death induced by the retinoid anhydroretinol. Therefore, cannabinoids act as antioxidants to modulate cell survival and growth of B lymphocytes and fibroblasts. PMID- 10869380 TI - AIT-082, a cognitive enhancer, is transported into brain by a nonsaturable influx mechanism and out of brain by a saturable efflux mechanism. AB - A fundamental feature of any drug designed to treat a disease of the central nervous system is the ability to cross the blood-brain barrier. Passage across the blood-brain barrier of AIT-082, a cognitive enhancer, was investigated in mice. [(14)C]AIT-082 crossed the blood-brain barrier in young male Swiss-Webster mice with a mean influx constant (K(i)) of 0.6 +/- 0.2 microl g(-1) min(-1). Furthermore, [(14)C]AIT-082 was transported into brain of both young and old male C57BL/6 mice with a K(i) of 0.35 +/- 0.06 and 0.33 +/- 0.02 microl g(-1) min(-1), respectively. There was no significant effect of age or strain on the movement of [(14)C]AIT-082 across the blood-brain barrier in mice. When 110- or 650-fold excess unlabeled AIT-082 was included in the injection solution, the K(i) was not significantly changed in either Swiss-Webster or C57BL/6 mice. This indicated that [(14)C]AIT-082 crossed the blood-brain barrier by a nonsaturable mechanism. The passage of AIT-082 into brain extracellular fluid was confirmed with capillary depletion and microdialysis. The efflux of [(14)C]AIT-082 from brain also was examined. After i.c.v. injection, [(14)C]AIT-082 levels in brain decreased over time with a t(1/2) of 20.0 +/- 1.0 min. Excess unlabeled AIT-082 (600-fold) increased the t(1/2) to 35.5 +/- 3.6 min. Together, these data indicate that AIT-082 moves into brain via a nonsaturable mechanism and is actively transported out of brain. PMID- 10869381 TI - Alkanols inhibit respiration of intact mitochondria and display cutoff similar to that measured in vivo. AB - Primary aliphatic alcohols from hexanol to pentadecanol were tested for their effects on the succinate-supported respiration of intact mitochondria isolated from rat liver. Alkanols were found to inhibit State 3 and uncoupled respiration. The ADP/oxygen ratios, a measure of the efficiency of oxidative phosphorylation, also were lowered, but to a lesser degree when compared on the basis of percentage of controls. Given each alkanol's nearly identical effect on State 3 and uncoupled respiration, action is not directly on ATP synthase, but earlier in the respiratory process. In agreement with many other studies of the homologous series of alkanols, potency increased with number of carbons in the chain until reaching a peak, in this case at undecanol, then tapered off to tridecanol before reaching a cutoff, at tetradecanol. If tetradecanol or longer homologs have activity, it is only after a lag phase of >15-min preincubation. All alkanols up to tridecanol also acted as uncouplers. At higher doses, hexanol inhibited State 4 rates, whereas longer chain alkanols did not, even at doses that completely eliminated respiratory control. Hexanol and decanol also were assayed against freeze-thawed (broken) mitochondria to distinguish effects on the mitochondrial substrate carrier from those on the electron transport chain. Both compounds were only weak inhibitors of respiration in broken mitochondria, suggesting that inhibition originates from interference with the dicarboxylate carrier, which must transport succinate across the mitochondrial membranes before it can be fed into complex II, rather than affecting the electron transport chain itself. PMID- 10869382 TI - Reduction of cisplatin nephrotoxicity by procainamide: does the formation of a cisplatin-procainamide complex play a role? AB - Procainamide protects mice bearing P388 leukemic cells against the toxicity of cisplatin without diminishing antitumor activity. The mechanism of action of procainamide protection was investigated both in vitro and in vivo. HPLC studies showed that procainamide forms a complex with cisplatin in vitro that has a UV spectrum similar to that of DPR, a triamine platinum complex that contains procaine as ligand. We report here the effect of the reaction product of cisplatin and procainamide on both cisplatin-induced DNA interstrand cross-links (ISCLs) and on the total DNA platination of isolated DNA. Total DNA platination in vitro of isolated DNA was increased by 113% (P <.01) and 17% (P <.05) after incubation times of 1.75 and 6 h, respectively, compared with products from the reaction of cisplatin with water. Furthermore, the reaction product of cisplatin and procainamide was bound to DNA to a significantly greater extent than was cisplatin itself. ISCLs were decreased by 41% when this drug combination was incubated with DNA for 1.75 h, but no changes were observed after incubation for 6 h. We also examined the influence of the time interval between administration of cisplatin and procainamide on normal kidney injury, the renal distribution and urinary excretion of platinum, and the formation of cisplatin-DNA adducts in renal tissue of Sprague-Dawley rats after i.p. administration of 7.5 mg/kg cisplatin either with or without procainamide. The plasma concentrations of urea and creatinine and kidney histology demonstrated that procainamide provided effective protection in vivo in the rat when administered either simultaneously or at 0.5 and 1 h before or after cisplatin. The protection was accompanied by both higher renal levels of platinum and cisplatin-DNA adducts and by an increase in the formation of ISCLs. Moreover, a dose-dependent reduction of urinary excretion and concentration of platinum was also observed. We propose that procainamide, after accumulation in the kidney, may coordinate with cisplatin to form a less toxic DPR-like complex that renders rats less susceptible to cisplatin-induced toxicity. PMID- 10869383 TI - Ochratoxin A induces JNK activation and apoptosis in MDCK-C7 cells at nanomolar concentrations. AB - Ochratoxin A (OTA) is a ubiquitous fungal metabolite with nephritogenic, carcinogenic, and teratogenic action. Epidemiological studies indicate that OTA may be involved in the pathogenesis of different forms of human nephropathies. Previously we have shown that OTA activates extracellular signal-regulated kinases 1 and 2, members of the mitogen-activated protein kinases (MAPK) family, in the C7-clone but not in the C11-clone of renal epithelial Madin-Darby canine kidney (MDCK) cells. Here we show that nanomolar concentrations of OTA lead to activation of a second member of the MAPK family, namely, c-jun amino-terminal kinase (JNK) in MDCK-C7 cells but virtually not in MDCK-C11 cells, as determined by kinase assay and Western blot. Furthermore, OTA potentiated the effect of tumor necrosis factor-alpha on JNK activation. In parallel to its effects on JNK, nanomolar OTA induced apoptosis in MDCK-C7 cells but not in MDCK-C11 cells, as determined by DNA fragmentation, DNA ladder formation, and caspase activation. In addition, OTA potentiated the proapoptotic action of tumor necrosis factor-alpha. Our data provide additional evidence that OTA interacts in a cell type-specific way with distinct members of the MAPK family at concentrations where no acute toxic effect can be observed. Induction of apoptosis via the JNK pathway can explain some of the OTA-induced changes in renal function as well as part of its teratogenic action. PMID- 10869384 TI - Effects of cholinomimetic injection into the brain stem reticular formation on halothane anesthesia and antinociception in rats. AB - The brain stem reticular formation plays an important role in determining consciousness and arousal. Modulation of cholinergic neurotransmission in this region alters the sleep-wake cycle. In the present study, we examined the effect of the direct application of cholinergic agents into the pontine reticular nucleus on anesthetic requirements and recovery and antinociception in rats. Sprague-Dawley rats were implanted with 24-gauge guide cannulas 1.0 mm above the oral portion of pontine reticular nucleus (PnO) while under pentobarbital anesthesia with the use of a stereotaxic apparatus. After recovery from surgery, animals were randomly assigned to one of the following protocols: minimum alveolar concentration (MAC), recovery time, tail-flick latency, or motor blockade. All measurements were performed after carbachol microinjection into the PnO after pretreatment with atropine or mecamylamine. Carbachol injection into the PnO significantly reduced MAC of halothane and prolonged recovery in a dose dependent manner. Pretreatment with atropine reversed MAC reduction by carbachol, and both atropine and mecamylamine shortened recovery time under carbachol. In unanesthetized rats, carbachol produced antinociceptive effects as reflected by a change in tail-flick latency response. Atropine and mecamylamine inhibited antinociceptive effects of carbachol. These results suggest that cholinomimetic injection into the PnO modulates the anesthetic state produced by halothane, suggesting participation of this area in the mechanisms in the brain that generate the anesthetic state. PMID- 10869385 TI - Selective vasopressin, angiotensin II, or dual receptor blockade with developing congestive heart failure. AB - With developing congestive heart failure (CHF), activation of the vasopressin V(1a) and angiotensin II type 1 (AT(1)) receptors can occur. In the present study, we examined the direct effects of V(1a) receptor blockade (V(1a) block), selective AT(1) receptor blockade (AT(1) block), and dual V(1a)/AT(1) receptor blockade (dual block) with respect to left ventricular (LV) function and contractility during the progression of CHF. LV and myocyte functions were examined in pigs with pacing CHF (rapid pacing, 240 beats/min, 3 weeks, n = 10), pacing CHF with concomitant V(1a) block (SR49059, 60 mg/kg, n = 8), pacing CHF with concomitant AT(1) block (irbesartan, 30 mg/kg, n = 7), or pacing CHF with dual block (n = 7). LV end-diastolic dimension and peak wall stress were reduced in all receptor blockade groups compared with CHF values. However, LV fractional shortening was increased only in the dual block group compared with CHF values (29 +/- 3 versus 21 +/- 2, P <.05). Basal LV myocyte percent shortening increased in the dual block group compared with CHF values (3.44 +/- 0.23 versus 2.88 +/- 0.11, P <. 05). Although V(1a) or AT(1) block reduced LV loading conditions, only dual block resulted in improved LV and myocyte shortening. PMID- 10869386 TI - Evaluation of a minimal experimental design for determination of enzyme kinetic parameters and inhibition mechanism. AB - The advent of combinatorial chemistry has led to a deluge of new chemical entities whose metabolic pathways need to be determined. A significant issue involves determination of the ability of new agents to inhibit the metabolism of existing drugs as well as its own susceptibility for altered metabolism. There is need to estimate the enzyme inhibition parameters and mechanism or mechanisms of inhibition with minimal experimental effort. We examined a minimal experimental design for obtaining reliable estimates of K(i) (and V(max) and K(m)). Simulations have been applied to a variety of experimental scenarios. The least experimentally demanding case involved three substrate concentrations, [S], for the control and one substrate-inhibitor pair, [S]-[I]. The control and inhibitor data (with 20% coefficient of variance random error) were simultaneously fit to the full nonlinear competitive inhibition equation [simultaneous nonlinear regression (SNLR)]. Excellent estimates of the correct kinetic parameters were obtained. This approach is clearly limited by the a prior assumption of mechanism. Further simulations determined whether SNLR would permit assessment of the inhibition mechanism (competitive or noncompetitive). The minimal design examined three [S] (control) and three [S]-[I] pairs. This design was successful in identifying the correct model for 98 of 100 data sets (20% coefficient of variance random error). SNLR analysis of metabolite formation rate versus [S] permits a dramatic reduction in experimental effort while providing reliable estimates of K(i), K(m), and V(max) along with an estimation of the mechanism of inhibition. The accuracy of the parameter estimates will be affected by the experimental variability of the system under investigation. PMID- 10869387 TI - Transporter-mediated release: a superfusion study on human embryonic kidney cells stably expressing the human serotonin transporter. AB - HEK 293 cells stably expressing the human serotonin transporter (hSERT) were grown on coverslips, preincubated with [(3)H]5-hydroxytryptamine (5-HT), and superfused. Substrates of the hSERT [e.g., p-chloroamphetamine (PCA)], increased the basal efflux of [(3)H]5-HT in a concentration-dependent manner. 5-HT reuptake blockers (e.g., imipramine, paroxetine) also raised [(3)H]5-HT efflux, reaching approximately one-third of the maximal effect of the hSERT substrates. In uptake experiments, both groups of substances inhibited [(3)H]5-HT uptake. Using the low affinity substrate [(3)H]N-methyl-4-phenylpyridinium (MPP(+)) to label the cells in superfusion experiments, reuptake inhibitors failed to enhance efflux. Similar results were obtained using human placental choriocarcinoma (JAR) cells that constitutively express the hSERT at a low level. By contrast, PCA raised [(3)H]MPP(+) efflux in both types of cells, and its effect was inhibited by paroxetine. The addition of the Na(+),K(+)-ATPase inhibitor ouabain (100 microM) to the superfusion buffer enhanced basal efflux of [(3)H]5-HT-loaded hSERT cells by approximately 2-fold; the effect of PCA (10 microM) was strongly augmented by ouabain, whereas the effect of imipramine was not. The Na(+)/H(+) ionophore monensin (10 microM) also augmented the effect of PCA on efflux of [(3)H]5-HT as well as on efflux of [(3)H]MPP(+). In [(3)H]5-HT-labeled cells, the combination of imipramine and monensin raised [(3)H]5-HT efflux to a greater extent than either of the two substances alone. In [(3)H]MPP(+)-labeled cells, imipramine had no effect on its own and fully reversed the effect of monensin. The results suggest that the [(3)H]5-HT efflux caused by uptake inhibitors is entirely due to interrupted high-affinity reuptake, which is ongoing even under superfusion conditions. PMID- 10869388 TI - Influence of ovarian hormones and estrous cycle on the behavioral response to cocaine in female rats. AB - Both humans and experimental animals demonstrate gender differences in response to cocaine. However, the mechanisms underlying these differences remain unclear. The purpose of the present study was to determine whether ovarian steroid hormones play a role in the locomotor response to cocaine in rats. Initial assessments of locomotor activity measured using photobeam monitors verified the robust gender difference in response to cocaine in our experimental paradigm. Subsequently, cocaine (5.0, 7.5, and 10.0 mg/kg) was shown to increase total horizontal activity in a dose-dependent manner in independent groups of intact females; the 5.0 mg/kg dose was selected for use in additional studies to determine the effect of estrogen (E) and progesterone (P) on the response to cocaine. Mature female rats were ovariectomized (OVX) or OVX and implanted with hormone-filled (E or P) Silastic capsules. Three to 4 weeks later, automated and observational measures of behavior were recorded after the administration of 5 mg/kg cocaine. Hormone replacement with E or E + P (but not P alone) resulted in greater cocaine-evoked hyperactivity than was observed in OVX animals. On measurement in normally cycling rats, hyperactivity induced by 5 mg/kg cocaine was greater during proestrus and estrus than during diestrus 2. The results of this series of experiments demonstrate that E significantly influences the responsiveness of female rats to cocaine. The enhanced response to cocaine was demonstrated in the presence of pharmacologically administered E as well as correlated with the normal estrous cycle. PMID- 10869389 TI - Single-cell recombinant pharmacology: bovine alpha(1a)-adrenoceptors in rat-1 fibroblasts release intracellular ca(2+), display subtype-characteristic agonism and antagonism, and exhibit an antagonist-reversible inverse concentration response phase. AB - Phe-activated Ca(2+) signals recorded from single rat-1 fibroblasts stably expressing the bovine alpha(1a)-adrenoceptor (AR) were characterized and used to analyze functional agonist-antagonist interactions. The response to Phe was initiated by the mobilization of stored Ca(2+) and subsequently sustained by receptor-regulated Ca(2+) influx. The selective alpha(1A)-AR agonist (R)-A-61603 was 141-fold more potent as an agonist than Phe. This potency ratio was consistent with the pharmacology of the native alpha(1A)-ARs. Functional responses evoked by concentrations of Phe of more than 0. 3 microM displayed fade, which could be explained by agonist-dependent depletion of Ca(2+) stores. The antagonists tested did not conform to the predictions of the Schild equation for competitive antagonism as expected from the nonequilibrium nature of the response. The antagonist potency series WB4101 > or = prazosin >> BMY7378, however, was consistent with alpha(1A)-ARs. Antagonism exhibited by WB4101 and prazosin was compatible with a model in which antagonists dissociate so slowly from the receptor that this is a major factor in their inhibition of the transient agonist-mediated response, leading to the appearance of insurmountable antagonism. A consequence of this phenomenon was that an inverse concentration response relationship at high agonist concentrations was abolished by low concentrations of antagonists. Overall, the results indicate that quantitative pharmacology can be studied successfully in single cells even though equilibrium could not be achieved in the agonist-antagonist-response relationship in this particular cell phenotype. The study also showed a form of fade that could be readily explained. PMID- 10869390 TI - Postnatal developmental delay and supersensitivity to organophosphate in gene targeted mice lacking acetylcholinesterase. AB - Acetylcholinesterase (AChE; EC 3.1.1.7) is the primary terminator of nerve impulse transmission at cholinergic synapses and is believed to play an important role in neural development. Targeted deletion of four exons of the ACHE gene reduced AChE activity by half in heterozygous mutant mice and totally eliminated AChE activity in nullizygous animals. Butyrylcholinesterase (EC 3.1.1.8) activity was normal in AChE -/- mice. Although nullizygous mice were born alive and lived up to 21 days, physical development was delayed. The neuromuscular junction of 12 day-old nullizygous animals appeared normal in structure. Nullizygous mice were highly sensitive to the toxic effects of the organophosphate diisopropylfluorophosphate and to the butyrylcholinesterase-specific inhibitor bambuterol. These findings indicate that butyrylcholinesterase and possibly other enzymes are capable of compensating for some functions of AChE and that the inhibition of targets other than AChE by organophosphorus agents results in death. PMID- 10869391 TI - Intravenous cereport (RMP-7) modifies topographic uptake profile of carboplatin within rat glioma and brain surrounding tumor, elevates platinum levels, and enhances survival. AB - Several experiments studied the effects of i.v. infusions of the bradykinin agonist, Cereport (RMP-7), on permeability of the blood-brain tumor barrier in rat gliomas. First, the ability of Cereport to increase uptake of two poorly blood-brain barrier-penetrating drugs (lypophilic paclitaxel and hydrophilic carboplatin) was directly compared to provide new information regarding the scope of delivery effects achieved with Cereport. Next, the increased uptake of platinum into tumor and brain surrounding tumor was shown to closely parallel that of radiolabeled carboplatin, confirming that delivery of a biologically active moiety is increased with Cereport. This study also demonstrated that the elevated tumor levels of platinum persisted for at least 2 h. The enhanced carboplatin uptake was then examined using a novel, high spatial resolution analysis of autoradiography. This revealed that the effects of Cereport were not uniform throughout the tumor, because it especially modified those areas normally impermeable to carboplatin. Finally, a range of i.v. Cereport doses (3.0 and 9.0 microg/kg) was tested in combination with carboplatin to determine whether increased survival might be achieved and to define the relationship between Cereport dose, plasma levels, uptake of carboplatin, and enhanced survival. Survival was enhanced only by the high dose of Cereport; the high dose also produced robust increases in carboplatin uptake and plasma concentrations of Cereport estimated to achieve the K(i), whereas the low dose did not. These data offer fundamental information regarding the effects of Cereport on delivery of chemotherapeutic agents to brain tumors and provide new insight into receptor mediated permeability of the blood-brain tumor barrier. PMID- 10869392 TI - Isoprostanes, novel eicosanoids that produce nociception and sensitize rat sensory neurons. AB - Isoprostanes are a novel class of eicosanoids primarily formed by peroxidation of arachidonic acid. Because of their potential as inflammatory and/or hyperalgesic agents whose formation is largely independent of cyclooxygenases, we examined whether 8-iso prostaglandin E(2) (8-iso PGE(2)) or 8-iso prostaglandin F(2alpha) (8-iso PGF(2alpha)) reduces mechanical and thermal withdrawal threshold in rats, and whether they sensitize rat sensory neurons. Injection of 1 microg of 8-iso PGE(2) (in 2.5 microl) into the hindpaw of rats significantly reduced mechanical and thermal withdrawal thresholds, whereas 1 microg of 8-iso PGF(2alpha) elicited a transient decrease in only the mechanical withdrawal threshold. Both isoprostanes enhanced the firing of C-nociceptors in a concentration-dependent manner when injected into peripheral receptive fields. Exposing sensory neurons grown in culture to 1 microM 8-iso PGE(2) or 8-iso PGF(2alpha) augmented the number of action potentials elicited by a ramp of depolarizing current. In contrast, 8-iso PGE(2) but not 8-iso PGF(2alpha) enhanced the release of substance P- and calcitonin gene-related peptide-like immunoreactivity from isolated sensory neurons. Ten micromolar 8-iso PGE(2) stimulated peptide release directly, whereas treatment with 1 microM 8-iso PGE(2) augmented the release evoked by either bradykinin or capsaicin. Pretreating neuronal cultures with the nonsteroidal anti-inflammatory drug ketorolac did not alter the sensitizing action of 8-iso PGE(2) on peptide release, suggesting that this action of the isoprostane was not secondary to the production of prostaglandins via the cyclooxygenase pathway. These data support the notion that isoprostanes are an important class of inflammatory mediators that augment nociception. PMID- 10869393 TI - Effects of aniracetam on bladder overactivity in rats with cerebral infarction. AB - Aniracetam has been used to improve the mental condition of patients with cerebrovascular disease. Previous studies have demonstrated that aniracetam activates the residual functions of cholinergic neurons in damaged brain areas. In this study, the effects of aniracetam on bladder overactivity after left middle cerebral artery occlusion were assessed through oral or i.c.v. administration in sham-operated and cerebral infarcted rats. Oral administration of aniracetam (100 and 300 mg/kg) resulted in a significant and dose-dependent increase in bladder capacity in cerebral infarcted rats but had no effect on bladder capacity in sham-operated rats. Intracerebroventricular administration of aniracetam (0.25 and 2.5 microg/rat) resulted in a significant and dose-dependent increase in bladder capacity in cerebral infarcted rats but not in sham-operated rats. Aniracetam had no significant effect on bladder contraction pressure or micturition threshold pressure in either sham-operated or cerebral infarcted rats. Furthermore, i.c.v. administration of atropine (1 microg/rat), a muscarinic acetylcholine receptor antagonist, completely inhibited the enhancing effects of aniracetam on bladder capacity in cerebral infarcted rats. The effects of aniracetam on bladder overactivity are thought to be mediated in part by activation of cholinergic inhibitory mechanisms in the brain. These results indicate that aniracetam may improve the neurogenic voiding dysfunction observed in patients with cerebrovascular disease. PMID- 10869394 TI - Protective effects of I(1)-antihypertensive agent moxonidine against neurogenic cardiac arrhythmias in halothane-anesthetized rabbits. AB - Numerous studies have addressed the antihypertensive properties of I(1) imidazoline receptor agonists such as moxonidine, but very few authors examined their cardiac antiarrhythmic potency. Due to the important role of the sympathetic nervous system in the genesis of neurogenic cardiac arrhythmias, we investigated the antiarrhythmic effects of moxonidine and compared them to those of propranolol in an experimental model of neurogenic arrhythmias. Chronic bipolar electrodes were implanted within the posterior hypothalamus of six halothane-anesthetized rabbits. Every 15 days, after three 10-min-interval control electrical stimulations, we compared the effects of randomized i.v. administrations of moxonidine (25 microg/kg), propranolol (0.5 mg/kg), and saline (0.9% NaCl) on mean arterial pressure (MAP), heart rate (HR), and ECG during 2.5 h with six stimulations every 20 min. We observed that: 1) in control conditions, intrahypothalamic stimulation increased MAP (DeltaMAP = 17 +/- 2 mm Hg) and HR (DeltaHR = 60 +/- 1 beats/min), and triggered extrasystoles (number of extrasystoles = 55 +/- 2) and abnormal complexes (number of abnormal ECG complexes = 37 +/- 1), which occurred with a 6.4 +/- 0.4-s delay and 33 +/- 1-s duration; 2) moxonidine and propranolol induced almost equihypotensive (DeltaMAP = -12 +/- 2 and -10 +/- 2 mm Hg) and pronounced bradycardic effects (DeltaHR = 47 +/- 10 and -78 +/- 9 beats/min, respectively). Arrhythmias were significantly reduced by moxonidine and propranolol: Deltanumber of extrasystoles = -83 and 98%; Deltanumber of abnormal ECG complexes = -33 and -79%; Deltadelay = +65 and +188%; Deltaduration = -35 and -58%, respectively. Our results show that moxonidine presents an antiarrhythmic potency comparable to that of propranolol that should be predominantly related to their central action. However, additional studies are required to determine whether these antiarrhythmic effects are of central and/or peripheral origin. PMID- 10869395 TI - Role of the beta(3)-adrenoceptor in urine storage in the rat: comparison between the selective beta(3)-adrenoceptor agonist, CL316, 243, and various smooth muscle relaxants. AB - The objective of this study was to compare the effects of a beta(3)-adrenoceptor (beta(3)-AR) agonist on bladder function and cardiovascular parameters in rats with those of several drugs that act on smooth muscle. CL316,243 (beta(3)-AR agonist), isoproterenol (nonselective beta-AR agonist), procaterol (beta(2)-AR agonist), verapamil (Ca(2+) antagonist), and papaverine (antispastic drug) each evoked a concentration-dependent relaxation of the detrusor in vitro. They also reduced bladder pressure in anesthetized rats, the beta-AR agonists apparently being more potent than the other drugs. Atropine (muscarinic antagonist) neither relaxed detrusor strips nor reduced bladder pressure. In anesthetized rats, CL316,243 and atropine each had only a slight influence on blood pressure and heart rate, but isoproterenol, procaterol, verapamil, and papaverine significantly affected cardiovascular function at the same dose range as that required to reduce bladder pressure. In cystometry experiments, CL316,243 (10 microg/kg i.v.), verapamil (1 mg/kg i.v.), and papaverine (1 mg/kg i.v.) all significantly prolonged micturition interval and increased bladder capacity, but did not change the residual urine volume after a micturition contraction. Procaterol (100 microg/kg i.v.) prolonged the micturition interval and increased both bladder capacity and residual urine volume (all significantly). Atropine (100 microg/kg i.v.) reduced micturition pressure and increased residual urine volume (both significantly). Because the human detrusor, like the rat detrusor, relaxes on beta(3)-AR stimulation, we conclude that this beta(3)-AR agonist may have potential in pollakiuria (frequent urination) as a therapeutic agent without cardiovascular side effects. PMID- 10869396 TI - Subsensitivity to opioids is receptor-specific in isolated guinea pig ileum and mouse vas deferens after obstructive cholestasis. AB - The rate and degree of subsensitivity development to morphine (mu-opioid receptor, preferred, but not selective agonist) and U50488H (highly selective kappa-opioid receptor agonist) were assessed in vitro on guinea pig ileum (GPI) of cholestatic animals 2, 5, and 7 days after bile duct ligation. In addition to this phenomenon of morphine, the effects of U50488H and SNC 80 (highly selective delta-opioid receptor agonist) were studied in vitro on mice vas deferens (MVD) of cholestatic animals 2, 5, 7, 10, and 15 days after bile duct ligation. The IC(50) for each compound was determined in these preparations. The ratio of the IC(50) in bile duct-ligated animals to sham and control animals provides a quantitative index for the degree of subsensitivity development to each agonist. For any given time, the highest degree of subsensitivity to morphine was observed in GPI of cholestatic animals, whereas in MVD obtained from the cholestatic animals, the highest degree of subsensitivity developed to inhibitory effect of SNC 80. The subsensitivity development in cholestatic animals was time dependent; in GPI the maximum subsensitivity developed after 7 days of the operation, whereas the maximum subsensitivity in MVD developed 15 days after bile duct ligation. Moreover, subsensitivity to exogenous acetylcholine and norepinephrine in GPI and MVD, respectively, did not develop in the presence of subsensitivity to opioids in cholestatic animals. Significant accumulation of endogenous opioids in plasma of cholestatic animals has been shown in several studies and this may account for a significant development of subsensitivity to inhibitory effects of opioid agonists. PMID- 10869397 TI - Catalytic antibodies that hydrolyze (-)-cocaine obtained by a high-throughput procedure. AB - Antibodies to a 2beta-carboxamido-2beta-phosphonate transition-state analog of ( )-cocaine benzoate ester hydrolysis were elicited in mice. A large number of hybridoma cell lines were propagated, and the catalytic activity of culture fluid was determined with a high-throughput photometric assay using cocaine benzoyl thioester as substrate. Binding avidity of the hybridoma supernatants to the phosphonate hapten was also determined. The initial rate constants for cocaine benzoyl thioester hydrolysis and binding avidity for a large number of hybridoma supernatants elicited to the phosphonate hapten did not always correlate. The lack of correlation of substrate hydrolysis with the binding affinity of 70 catalytic antibodies was also observed for (-)-cocaine hydrolysis using derivatization and HPLC analysis of methyl ecgonine as meta-nitrococaine. The k(cat) values for cocaine benzoyl thioester hydrolysis by monoclonal antibodies 3, 5, and 12 were 38, 4.2, and 0. 6 min(-1), respectively. For monoclonal antibody 5, the selectivity ratios (K(i) value divided by the K(m) value for the hydrolysis of cocaine benzoyl thioester) with ecgonine benzoyl ester, ecgonine methyl ester, norcocaine, and ecgonine were 101, 25, 9.4, and 4, respectively. Three active esterolytic monoclonal antibodies identified with the high throughput assay procedure were examined in detail for their ability to hydrolyze (-)-cocaine. The k(cat) values for the hydrolysis of (-)-cocaine with monoclonal antibodies 3, 5, and 12 were 6.6, 0.4, and 0.1 min(-1), respectively. Hydrolysis of (-)-cocaine by monoclonal antibody 3 approached the k(cat) value for that of human butyrylcholinesterase. Cocaine esterolytic catalytic antibodies that approach or exceed the catalytic efficiency of human butyrylcholinesterase may represent a new pharmacological intervention approach to the treatment of cocaine abuse, and the high-throughput process described here represents an advance in the effort to develop clinically useful antibodies. PMID- 10869398 TI - Dextromethorphan and its metabolite dextrorphan block alpha3beta4 neuronal nicotinic receptors. AB - Dextromethorphan (DM), a structural analog of morphine and codeine, has been widely used as a cough suppressant for more than 40 years. DM is not itself a potent analgesic, but it has been reported to enhance analgesia produced by morphine and nonsteroidal anti-inflammatory drugs. Although DM is considered to be nonaddictive, it has been reported to reduce morphine tolerance in rats and to be useful in helping addicted subjects to withdraw from heroin. Here we studied the effects of DM on neuronal nicotinic receptors stably expressed in human embryonic kidney cells. Studies were carried out to examine the effects of DM on nicotine-stimulated whole cell currents and nicotine-stimulated (86)Rb(+) efflux. We found that both DM and its metabolite dextrorphan block nicotinic receptor function in a noncompetitive but reversible manner, suggesting that both drugs block the receptor channel. Consistent with blockade of the receptor channel, neither drug competed for the nicotinic agonist binding sites labeled by [(3)H]epibatidine. Although DM is approximately 9-fold less potent than the widely used noncompetitive nicotinic antagonist mecamylamine in blocking nicotinic receptor function, the block by DM appears to reverse more slowly than that by mecamylamine. These data indicate that DM is a useful antagonist for studying nicotinic receptor function and suggest that it might prove to be a clinically useful neuronal nicotinic receptor antagonist, possibly helpful as an aid for helping people addicted to nicotine to refrain from smoking, as well as in other conditions where blockade of neuronal nicotinic receptors would be helpful. PMID- 10869399 TI - Oxidant stress in rat liver after lipopolysaccharide administration: effect of inducible nitric-oxide synthase inhibition. AB - The role of inducible nitric-oxide synthase (iNOS) in lipopolysaccharide (LPS) induced hepatic oxidant stress was evaluated using the iNOS inhibitor L iminoethyl-lysine (L-NIL). Male rats were divided into three groups. One group received LPS (Salmonella minnesota) 2 mg/kg i.v. A second group received LPS plus L-NIL (3 mg/kg i.p.) at the time of LPS administration followed by a second dose 3 h later. A third group received saline i.v. At 6 h, blood and liver tissue were collected. Serum nitrate/nitrite (metabolic products of nitric oxide) levels were increased from 5.4 +/- 1.5 nmol/ml in the saline group to 360 +/- 48 nmol/ml in the LPS group (n = 5). Values for the LPS + L-NIL group were significantly reduced to 35 +/- 7 nmol/ml. Tissue malondialdehyde levels were increased from 0.20 +/- 0.02 nmol/mg (n = 4) in the saline group to 0.41 +/- 0.03 nmol/mg (n = 4) in the LPS group. L-NIL significantly reduced the values in the LPS group to 0.29 +/- 0.02 nmol/mg (n = 4). 4-Hydroxynonenal-protein adducts levels were increased 3.6-fold by LPS treatment as compared with saline. L-NIL significantly reversed the levels to 1.6-fold (n = 4). Intracellular GSH levels were decreased from 8.49 +/- 0.64 nmol/mg (n = 4) in the saline group to 5.63 +/- 0.51 nmol/mg in the LPS group (n = 7). L-NIL significantly increased the levels in the LPS group to 7.04 +/- 0.46 nmol/mg (n = 7). These data indicate that LPS-induced nitric oxide generation can result in oxidant stress in the liver, and that inhibitors of iNOS may offer some protection in LPS-induced hepatic toxicity. PMID- 10869400 TI - Receptor-mediated inhibition of keratinocyte migration by nicotine involves modulations of calcium influx and intracellular concentration. AB - Early stages of wound healing rely on the ability of keratinocytes (KCs) to move over the denuded dermis to re-epithelialize the defect. The agarose gel keratinocyte outgrowth system (AGKOS) is an in vitro model of skin re epithelialization designed to study the migratory function of KCs. Endogenously secreted acetylcholine controls crawling locomotion of KCs in AGKOS by binding to the cholinergic receptors of both the nicotinic and the muscarinic classes that are expressed by KCs. In this study, we used AGKOS to elucidate the nicotinic pathway of cholinergic control of keratinocyte migration. Activation of the nicotinic acetylcholine receptors decreased the migration distance of KC in a dose-dependent fashion without altering cell viability. Nicotine also increased in a dose-dependent manner transmembrane influx of (45)Ca(2+), and caused a transient rise in the concentration of [Ca(2+)](i). Perfect correlation between concentration responses found in the migration and (45)Ca(2+) influx assays suggested that nicotine-induced inhibition of crawling locomotion relies on modulation of Ca(2+) metabolism in KCs. The effects of nicotine could be mediated by the alpha3- and the alpha7-containing nicotinic receptors visualized on KCs by immunostaining. Long-term incubation with nicotine up-regulated alpha7 and down regulated alpha3 expression. Thus, nicotine exerts inhibitory effects on keratinocyte migration, and Ca(2+) serves as a second messenger in the signaling pathway. These results help explain deleterious effects of nicotine on wound re epithelialization, and suggest that smoking may delay wound healing via nicotinic receptor-mediated pathway. PMID- 10869402 TI - Intrarenal effects of ecadotril during acute volume expansion in dogs with congestive heart failure. AB - Neutral endopeptidase 24.11 (NEP) inhibitors are known to have vascular, diuretic, and natriuretic effects that may be helpful in the treatment of congestive heart failure (CHF). Most NEP inhibitors may act principally through intrarenal mechanisms, which are not completely understood. The purpose of this study was to determine the principal renal effects of the NEP inhibitor ecadotril in dogs with progressive CHF induced by rapid ventricular pacing. Renal function was measured before, during, and after acute i.v. infusion of normal saline in a total of six dogs during normal cardiac function, early left ventricular dysfunction, and overt CHF. During overt CHF, each dog was treated with either ecadotril or placebo orally for 1 week. Parameters measured included glomerular filtration rate, renal blood flow, urine output, sodium clearance, sodium fractional excretion, and proximal and distal sodium reabsorption. Ecadotril treatment resulted in increased urine output, sodium clearance, and renal sodium excretion relative to placebo-treated controls. The principal intrarenal effect of ecadotril was decreased distal renal tubular sodium reabsorption. Both glomerular filtration rate and renal blood flow declined during overt CHF and were unaffected by ecadotril treatment. The results of this study are consistent with the principal action of ecadotril occurring by way of intrarenal events as opposed to changes in renal hemodynamics. The principal effect of ecadotril on distal tubular sodium reabsorption suggests that inhibition of NEP activity in the proximal renal tubules may allow increased binding of filtered atrial natriuretic peptide to natriuretic peptide receptor sites in the distal renal tubules and collecting ducts. PMID- 10869401 TI - Reactive oxygen species-induced apoptosis in PC12 cells and protective effect of bilobalide. AB - Although clinical studies have demonstrated that EGb 761, a standard extract of Ginkgo biloba, was effective in mild-to-moderate dementia of the Alzheimer's disease patients, the mechanism underlying its neuroprotective effect remains unclear. In this study, effects of bilobalide, the main constituent of the nonflavone fraction of EGb 761, on reactive oxygen species (ROS)-induced apoptosis in PC12 cells was studied. Exposure of cells to xanthine (100 microM)/xanthine oxidase (150 mU/ml) (ROS producer) resulted in a characteristic DNA fragmentation and an increase in the apoptosis rate. When p53, c-Myc, Bcl-2, Bcl-x(L), and Bax were measured by flow cytometry and the activities of caspase-1 and caspase-3-like protease determined with Ac-YVAD-AMC or Ac-DEVD-AMC as substrates, the profile of ROS-induced changes in these apoptosis regulatory and effector proteins suggests that elevation of c-Myc, p53, and Bax and activation of caspase-3 play an important role in the apoptosis. When cells were treated with ROS and bilobalide (25-100 microM) simultaneously, a dose-dependent reduction in the apoptotic rate was found. The percentage of cells with positive staining for c-Myc and p53 decreased from 27.8 and 50.1% to 16.7 and 23.2%, respectively, when bilobalide (25 microM) was present. Bilobalide also reduced ROS-induced elevation of Bax and activation of caspase-3 effectively. Our results provide the first direct evidence that bilobalide can protect neurons against oxidative stress. Bilobalide may block the apoptosis in the early stage and then attenuate the elevation of c-Myc, p53, and Bax and activation of caspase-3 in cells. PMID- 10869403 TI - Preclinical and clinical evidence for disappearance of long-circulating characteristics of polyethylene glycol liposomes at low lipid dose. AB - This study describes the effect of the lipid dose of (99m)Tc-polyethylene glycol (PEG) liposomes in the low-dose range (0. 02-1.0 micromol/kg) on the pharmacokinetics and biodistribution in rats, rabbits, and humans. The biodistribution and pharmacokinetics of (99m)Tc-PEG liposomes at various dose levels were studied in rats and rabbits with a focal Escherichia coli infection. Scintigraphic images were recorded on a gamma camera. In addition, the role of macrophages in the biodistribution of a low-dose PEG liposome injection was studied. Finally, the pharmacokinetics of (99m)Tc-PEG liposomes at two lipid dose levels was studied in four patients. At a dose level of 0.03 micromol/kg, the blood level in rats at 4 h postinjection was significantly lower than at the highest dose level (1.1 micromol/kg). The same effect was observed in rabbits where enhanced clearance was observed at a dose level of 0.02 micromol/kg. The circulatory half-life decreased from 10.4 to 3.5 h (at 1.0 and 0. 02 micromol/kg, respectively). At the lowest dose level, liposomes were mainly taken up by the liver and to a lesser extent by the spleen. Injection of a low dose of PEG liposomes in macrophage-depleted rabbits resulted in normal pharmacokinetics, suggesting involvement of macrophages in the effectuation of the rapid elimination of the liposomes from the circulation. Most importantly, the rapid clearance of low-dose PEG liposomes was also observed in humans when relatively low lipid doses were administered. This study showed that at very low lipid doses the biodistribution of PEG liposomes is dramatically altered. PMID- 10869404 TI - mu-Opioid receptor knockout mice do not self-administer alcohol. AB - Opioid peptides long have been hypothesized to play a role in ethanol reinforcement. Neuropharmacological studies have shown that opioid receptor antagonists decrease ethanol self-administration in rodents and prevent relapse in humans. However, the exact mechanism for such powerful effects has remained elusive. The availability of mu-opioid receptor knockout mice has made possible the direct examination of the role of the mu-opioid receptor in mediating ethanol self-administration. In the present experiments, both nosepoke and lever operant ethanol self-administration and several tests of two bottle-choice ethanol drinking were studied in these genetically engineered mice. In no case did knockout mice show evidence of ethanol self-administration, and, in fact, these mice showed evidence of an aversion to ethanol under several experimental conditions. These data provide new evidence for a critical role for mu-opioid receptors in ethanol self-administration assessed with a variety of behavioral paradigms and new insights into the neuropharmacological basis for ethanol reinforcement. PMID- 10869405 TI - Enantioselectivity of pregnanolone-induced gamma-aminobutyric acid(A) receptor modulation and anesthesia. AB - This study reports the actions of enantiomer pairs of anesthetic steroids 3alpha5alphaP/ent-3alpha5alphaP and 3alpha5betaP/ent-3alpha5betaP as modulators of gamma-aminobutyric acid (GABA)(A) receptors and as anesthetics. The enantiomers of structurally related 17-carbonitrile analogs also are examined. These studies were aimed at 1) determining whether the steroid recognition site could distinguish between molecules differing in shape, but not other physical properties (enantioselectivity); 2) providing further insight into the structure activity relationships of anesthetic steroids; and 3) determining whether modulation of GABA(A) receptor function correlates with anesthetic potency for anesthetic steroid enantiomers. Stereoselective actions of the compounds were evaluated in four different bioassays: 1) noncompetitive displacement of [(35)S]t butylbicyclophosphorothionate from the picrotoxin site of GABA(A) receptors present in rat brain membrane preparations; 2) modulation of GABA currents in cultured rat hippocampal neurons; 3) loss of righting reflex in tadpoles; and 4) loss of righting reflex in mice. The data indicate that 5alpha-reduced steroids, but not 5beta-reduced steroids, show a high degree of enantioselectivity/enantiospecificity in their actions as modulators of GABA(A) receptors and as anesthetics. For all compounds studied, the effects on GABA(A) receptor function closely tracked with anesthetic effects. These data show that the anesthetic steroid recognition site is capable of distinguishing enantiomers, suggesting a protein-binding site of specific dimensions and shape. The results are compatible either with a structural model of the binding site that can accommodate 3alpha5alphaP, 3alpha5betaP, and ent-3alpha5betaP, but not ent 3alpha5alphaP, or with two different binding sites for steroid anesthetics. PMID- 10869406 TI - Dissociation of cocaine-antagonist properties and motoric effects of the D1 receptor partial agonists SKF 83959 and SKF 77434. AB - Previous studies suggest that D1 receptor partial agonists may be viable candidates for development as pharmacotherapies for cocaine addiction. This study investigated the ability of the D1 receptor partial agonists SKF 83959 and SKF 77434 to modulate the behavioral effects of cocaine and compared these effects with those of the reference D1 receptor antagonist SCH 39166 and D1 receptor agonists SKF 81297 and 6-Br-APB. Squirrel monkeys were trained either to respond under a fixed-interval schedule of stimulus-shock termination or to discriminate cocaine from vehicle (procedures useful for evaluating the behavioral stimulant and subjective effects of cocaine, respectively). Additional monkeys were studied with quantitative observational techniques to evaluate the effects of the drugs on various forms of motor behavior. Like SCH 39166, but unlike SKF 81297 and 6-Br APB, the D1 receptor partial agonists attenuated the behavioral stimulant and discriminative stimulus effects of cocaine in a dose-dependent manner, although maximum antagonism produced by SKF 77434 was not always as great as that produced by SKF 83959 or SCH 39166. In observational studies, SKF 83959 and SKF 77434 produced less severe disruptions in motor behavior than did SCH 39166 and, for SKF 83959, showed a greater separation between the dose required to antagonize the behavioral effects of cocaine and the dose that induced catalepsy (>/=33 fold). These results suggest that D1 receptor partial agonists can act as functional cocaine antagonists with less severe behavioral effects than D1 receptor antagonists. The prominent cocaine-antagonist properties and the low incidence of motoric side effects of SKF 83959 may reflect its unique binding profile at D1 as well as nondopaminergic receptors. PMID- 10869407 TI - Latent overexpression of hepatic CYP2C7 in adult male and female rats neonatally exposed to phenobarbital: a developmental profile of gender-dependent P450s. AB - For more than 20 years it has been known that neonatal exposure to phenobarbital results in a delayed, but permanent overexpression of drug-metabolizing enzymes in adult male and female rats. Accordingly, to identify the specific isoform(s) of P450 responsible for the imprinted overexpression of hepatic monooxygenases, we have monitored the developmental profile of some dozen hepatic P450 isoforms in 4- to 150-day-old male and female rats neonatally treated with the barbiturate. Some of the cytochrome P450s (CYP), i. e., CYP2A1, 2A2, 2C6, 3A1, and 3A2, exhibit the typical transient response in which isoform levels (mRNA, protein, and/or specific catalytic activity) rise precipitously at the time of phenobarbital administration and rapidly decline to preinduction levels after withdrawal of the barbiturate. Other isoforms, i.e., CYP1A1, 1A2, 2C7, 2C11, 2C12, and 2C13, were neither constitutively expressed nor phenobarbital inducible in the neonate. Only one of these isoforms, female predominant (M:F, approximately 1:2) CYP2C7, exhibited a barbiturate-induced delayed, but persistent approximately 30 to 50% overexpression from puberty through adulthood. We propose that at the time of exposure, neonatally administered phenobarbital produces a "silent" programming defect resulting in a delayed, but persistent overexpression of the isoform, contributing, at least in part, to a permanent elevation of hepatic drug-metabolizing enzyme activities. PMID- 10869408 TI - Trans-stimulation effects of folic acid derivatives on methotrexate transport by rat renal organic anion transporter, OAT-K1. AB - We examined the pharmacological role of the renal organic anion transporter OAT K1, which localizes predominantly in the brush-border membranes of proximal straight tubules, in the urinary excretion of methotrexate and the possibility of its contribution to "folinic acid rescue." With Madin-Darby canine kidney (MDCK) cells stably transfected with OAT-K1 cDNA, OAT-K1-mediated methotrexate accumulation was inhibited in the presence of various folic acid derivatives. These derivatives included aminopterin, 5-methyltetrahydrofolic acid, unlabeled methotrexate, folinic acid (citrovorum factor, leucovorin), and folic acid with apparent inhibition constant values of 0.5, 1.2, 1.8, 8.2, and 14.1 microM, respectively. In contrast, 10 microM taurocholic acid and sulfobromophthalein did not inhibit OAT-K1-mediated methotrexate accumulation. In addition, methotrexate efflux was stimulated in the presence of inwardly directed gradients of aminopterin, 5-methyltetrahydrofolic acid, unlabeled methotrexate, folinic acid, and folic acid, but not of uric acid, taurocholic acid, and glutathione, indicating that OAT-K1-mediated methotrexate efflux is stimulated by a folic acid derivatives exchange. In conclusion, OAT-K1 was suggested to enhance the apical efflux of highly accumulated methotrexate in tubular epithelial cells and contribute at least in part to folinic acid rescue by exchanging intracellular methotrexate for extracellular folinic acid. PMID- 10869409 TI - Inhibitory effects of AE0047, a new dihydropyridine Ca(2+) channel blocker, on renal nerve stimulation-induced renal actions in anesthetized dogs. AB - The effects of AE0047, a newly developed dihydropyridine Ca(2+) channel blocker, and nicardipine on changes in the renal function and norepinephrine (NE) overflow induced by renal nerve stimulation (RNS) were examined in anesthetized dogs. RNS at a low frequency (0. 5-2.0 Hz) caused significant decreases in the urine flow, urinary excretion of sodium, and fractional excretion of sodium, while also inducing increases in the NE secretion rate (NESR), without affecting the renal hemodynamics. RNS at a high frequency (2.5-5.0 Hz), which diminishes the renal blood flow, glomerular filtration rate, and filtrate fraction, elicited more potent decreases in urine formation and increases in NESR than those seen for the low-frequency RNS. When AE0047 (10 and 50 ng/kg/min) was administered intrarenally, increases in the basal renal blood flow and urine formation were observed. During AE0047 (50 ng/kg/min) infusion, low-frequency RNS-induced antidiuretic action and increase in NESR were markedly attenuated. Qualitatively similar results were observed for high-frequency RNS. In addition, high-frequency RNS-induced renal vasoconstriction was significantly suppressed by AE0047 infusion at higher doses. Lower doses of AE0047 (10 ng/kg/min) tended to attenuate the low- and high-frequency RNS-induced antidiuretic actions, although neither of the RNS-induced increases in NESR were suppressed by lower doses of AE0047. Nicardipine (50 ng/kg/min) also increased the level of basal urine formation, but the RNS-induced changes in renal function and increases in NESR were not affected by this drug. These results suggest that AE0047 suppresses the RNS-induced NE overflow from renal nerve endings, which is probably involved in the inhibitory effects of the drug on the antidiuretic action elicited by RNS. PMID- 10869410 TI - S33084, a novel, potent, selective, and competitive antagonist at dopamine D(3) receptors: I. Receptorial, electrophysiological and neurochemical profile compared with GR218,231 and L741,626. AB - The benzopyranopyrrole S33084 displayed pronounced affinity (pK(i) = 9.6) for cloned human hD(3)-receptors, and >100-fold lower affinity for hD(2) and all other receptors (>30) examined. S33084 concentration dependently, potently, and competitively (pA(2) = 9.7) antagonized dopamine (DA)-induced [(35)S]guanosine-5' O-(3-thio)triphosphate (GTPgammaS) binding at hD(3)-receptors. It also concentration dependently abolished stimulation by DA of hD(3)-receptor-coupled mitogen-activated protein kinase. Administered alone, S33084 did not modify dialysate levels of DA in the frontal cortex, nucleus accumbens, or striatum of freely moving rats, nor the firing rate of ventrotegmental dopaminergic cell bodies. Furthermore, it had minimal effect on DA turnover in mesocortical, mesolimbic, and nigrostriatal projection regions. However, S33084 dose dependently blocked the suppressive influence of the preferential D(3)-agonist PD128,907 on frontocortical release of DA. Furthermore, it likewise antagonized the inhibitory influence of PD128,907 on the electrical activity of ventrotegmental dopaminergic neurons. Although less potent than S33084, GR218,231 likewise behaved as a selective hD(3)- versus hD(2)-receptor antagonist and its neurochemical and electrophysiological profiles were similar. In contrast, L741,626 was a preferential antagonist at hD(2) versus hD(3) sites. In vivo, on administration alone, L741,626 increased frontocortical, mesolimbic, and (more potently) striatal DA release, enhanced the firing rate of dopaminergic perikarya, and accelerated cerebral DA synthesis. It also blocked the actions of PD128,907. In conclusion, S33084 is a novel, potent, selective, and competitive antagonist at hD(3)-receptors. Although GR218,231 behaves similarly, L741,626 is a preferential D(2)-receptor antagonist. DA D(2)- but not D(3)-(auto) receptors tonically inhibit ascending dopaminergic pathways, although the latter may contribute to phasic suppression of DA release in frontal cortex. PMID- 10869411 TI - S33084, a novel, potent, selective, and competitive antagonist at dopamine D(3) receptors: II. Functional and behavioral profile compared with GR218,231 and L741,626. AB - The selective dopamine D(3)-receptor antagonist S33084 dose dependently attenuated induction of hypothermia by 7-hydroxy-2-dipropylaminotetralin (7-OH DPAT) and PD128,907. S33084 also dose dependently reduced 7-OH-DPAT-induced penile erections (PEs) but had little effect on 7-OH-DPAT-induced yawning and hypophagia, and it did not block contralateral rotation elicited by the preferential D(3) agonist quinpirole in unilateral substantia nigra-lesioned rats. In models of potential antipsychotic activity, S33084 had little effect on conditioned avoidance behavior and the locomotor response to amphetamine and cocaine in rats, and weakly inhibited apomorphine-induced climbing in mice. Moreover, S33084 was inactive in models of potential extrapyramidal activity in rats: induction of catalepsy and prolactin secretion and inhibition of methylphenidate-induced gnawing. Another selective D(3) antagonist, GR218,231, mimicked S33084 in inhibiting 7-OH-DPAT-induced PEs and hypothermia but neither hypophagia nor yawning behavior. Similarly, it was inactive in models of potential antipsychotic and extrapyramidal activity. In distinction to S33084 and GR218,231, the preferential D(2) antagonist L741,626 inhibited all responses elicited by 7-OH-DPAT. Furthermore, it displayed robust activity in models of antipsychotic and, at slightly higher doses, extrapyramidal activity. In summary, S33084 was inactive in models of potential antipsychotic and extrapyramidal activity and failed to modify spontaneous locomotor behavior. Furthermore, it did not affect hypophagia or yawns, but attenuated hypothermia and PEs, elicited by 7 OH-DPAT. This profile was shared by GR218,231, whereas L741,626 was effective in all models. Thus, D(2)-receptors are principally involved in these paradigms, although D(3)-receptors may contribute to induction of hypothermia and PEs. S33084 should comprise a useful tool for further exploration of the pathophysiological significance of D(3)- versus D(2)-receptors. PMID- 10869412 TI - Tectoridins modulate skeletal and cardiac muscle sarcoplasmic reticulum calcium release channels. AB - The isoflavones tectoridin (TTR) and 3'-hydroxy TTR (3'-TTR) were isolated from an Ayurvedic herbal preparation Vaca and evaluated for their affinity and effect on ryanodine receptors (RyR) using junctional sarcoplasmic reticulum vesicles (JSRVs). In [(3)H]ryanodine displacement binding affinity assays, TTR and 3'-TTR exhibited IC(50) values of 17.3 +/- 1.3 microM (K(d) = 6.7 +/- 0.4 microM) and 6.6 +/- 1.4 microM (K(d) = 2.4 +/- 0.2 microM), respectively, for fast skeletal muscle RyR (RyR1) compared with an IC(50) value for ryanodine of 6.2 +/- 0.4 nM (K(d) = 2.4 nM). TTR demonstrated a 3-fold higher affinity for cardiac RyR (RyR2) [IC(50) value of 5.2 +/- 0.6 microM (K(d) = 0.95 +/- 0.3 microM)] than for RyR1. The displacement isotherms for both TTRs paralleled that for ryanodine, consistent with the notion that all three are likely binding to similar site(s) on the receptors. Calcium efflux from and calcium influx into JSRVs were used to measure function effects of TTRs on binding to RyR. In calcium efflux assays, TTR (up to 1 mM) enhanced the release of (45)Ca(2+) from JSRVs in a concentration dependent manner (EC(50act) of 750 microM). Higher concentrations deactivated (partially closed) RyR1. 3'-TTR had similar effects, but was approximately 2-fold more potent, exhibiting an EC(50act) value of 480 microM. Using passive calcium influx assays, TTR activated and deactivated RyR1 in a time- and concentration dependent manner. The aglycone tectorigenin also was effective in displacing [(3)H]ryanodine from RyR1 but not from RyR2. These results demonstrate that TTRs are capable of interacting at ryanodine binding sites to differentially modulate fast skeletal and cardiac calcium-release channels. PMID- 10869413 TI - gamma-aminobutyric Acid(A) (GABA(A)) agonist 4,5,6, 7-tetrahydroisoxazolo[4,5 c]pyridin-3-ol persistently increases sleep maintenance and intensity during chronic administration to rats. AB - Many hypnotics, such as benzodiazepines, are agonistic modulators of gamma aminobutyric acid(A) (GABA(A)) receptors. Such compounds increase the ability to fall and stay asleep, but inhibit rapid-eye movement (REM) sleep and deep non-REM sleep. However, tolerance to their hypnotic action may develop rapidly. Previous findings in rats and humans demonstrate that the gamma-aminobutyric acid(A) agonist 4, 5,6,7-tetrahydroisoxazolo[4,5-c]pyridin-3-ol (THIP) promotes deep non REM sleep and increases non-REM sleep continuity. To investigate the effects of repeated administration, we assessed sleep in rats before, during, and after chronic dosing of THIP (3 mg/kg, once daily for 5 days; n = 9) or of placebo (n = 8). The substances were administered i.p. at the onset of darkness. The electroencephalogram (EEG) and electromyogram were recorded during the first 6 h after injection. During baseline recording, the placebo and the THIP group exhibited similar sleep patterns. After the first THIP injection, rats displayed more non-REM sleep, longer non-REM episodes, and higher levels of slow wave activity in the EEG within non-REM sleep than the placebo group rats. The effects were sustained during all treatment days. REM sleep was not affected. After drug withdrawal, the sleep patterns of the THIP and the placebo group were practically identical again. These observations suggest that THIP does not rapidly produce tolerance toward its sleep effects and abrupt drug withdrawal may not be associated with sleep disturbances. These findings confirm and extend the existing information suggesting that THIP may be promising for treatment of insomnia. PMID- 10869414 TI - Activity-dependent neurotrophic factor: intranasal administration of femtomolar acting peptides improve performance in a water maze. AB - Activity-dependent neurotrophic factor (ADNF) is a glia-derived protein that is neuroprotective at femtomolar concentrations. A nine-amino acid peptide derived from ADNF (Ser-Ala-Leu-Leu-Arg-Ser-Ile-Pro-Ala; ADNF-9) captured the activity of the parent protein and has been reported to protect cultured neurons from multiple neurotoxins. Antibodies recognizing ADNF-9 produced neuronal apoptosis, and identified an additional, structurally related, glia-derived peptide, Asn-Ala Pro-Val-Ser-Ile-Pro-Gln (NAP). Previous comparative studies have characterized s.c.-injected NAP as most efficacious in protecting against developmental retardation and learning impairments in apolipoprotein E-deficient mice. This study was designed to assess 1) neuroprotection after intranasal administration of ADNF-9 and NAP to rats treated with the cholinotoxin ethylcholine aziridium; and 2) bioavailability and pharmacokinetics after intranasal administration. Results showed significant improvements in short-term spatial memory, as assessed in a water maze, after daily intranasal administration of 1 microg of peptide (ADNF-9 or NAP) per animal. However, a 5-day pretreatment with ADNF-9 did not improve performance measured after cessation of treatment. Compared with rats treated with ADNF-9, NAP-pretreated animals exhibited a significantly better performance. Furthermore, NAP (and not ADNF-9) protected against loss of choline acetyl transferase activity. Significant amounts of (3)H-labeled NAP reached the brain, remained intact 30 min after administration, and dissipated 60 min after administration. This study revealed efficacy for ADNF-related peptides in rodent models for neurodegeneration. The small size of the molecules, the low dosage required, the noninvasive administration route, and the demonstrated activity in a relevant paradigm suggest NAP as a lead compound for future drug design. PMID- 10869415 TI - Intracisternal injection of somatostatin receptor 5-preferring agonists induces a vagal cholinergic stimulation of gastric emptying in rats. AB - We previously showed that the somatostatin receptor 5 (sst(5))-preferring agonist BIM-23052 injected intracisternally (i.c. ; 0.8 nmol/rat) stimulated gastric emptying of a non-nutrient meal in conscious rats. In this study, we investigated the neural pathways and specificity of BIM-23052 action. BIM-23052 (0.4, 0.8, and 1.2 nmol/rat i.c.) stimulated gastric transit; values of gastric emptying were 65.5 +/- 6.5, 77.4 +/- 5.3, and 77.7 +/- 1.9%, respectively, compared with 43.2 +/-3.2% in i.c. saline group. Intravenous injection of BIM-23052 (0.8 nmol/rat) had no effect. BIM-23052 (0.8 nmol/rat i.c.) action was prevented by subdiaphragmatic vagotomy or atropine. Medullary thyrotropin-releasing hormone (TRH) is known to play a physiological role in the vagal stimulation of gastric motor function. TRH receptor antisense oligodeoxynucleotides injected i.c. with a regimen that prevented TRH (0.3 nmol/rat i.c.)-induced enhanced gastric emptying did not influence BIM-23052 stimulatory action. Somatostatin-28 (0.2-1.2 nmol/rat i.c.), which possesses a higher affinity than somatostatin-14 for sst(5), and the cyclic octapeptide des-AA(1,2,4,5,12,13)[D-Trp(8)]somatostatin (0.2-1.2 nmol/rat i.c.), an oligo-somatostatin analog that shares similar brain actions as somatostatin-28, induced a dose-related stimulation of gastric emptying. Somatostatin-14 and the preferring peptide agonists for sst(1), CH-275; sst(2), DC-32-87; sst(3), BIM-23056 and L-796,778; and sst(4), L-803,087 had no significant effect on gastric emptying when injected i.c. at 0.8 nmol/rat. These results show that BIM-23056 injected i.c. acts in the brain independently from medullary TRH to induce a vagal cholinergic stimulation of gastric emptying through the sst(5) receptor subtype. PMID- 10869416 TI - Effect of novel motilide ABT-229 versus erythromycin and cisapride on gastric emptying in dogs. AB - ABT-229 (8,9-anhydro-4"-deoxy-3'-N-desmethyl-3'-N-ethylerythromycin B-6,9 hemiacetal), a synthetic derivative of erythromycin (ERY) with no antibiotic activity, has been shown to bind to motilin receptors and stimulate contractile activity of the antrum and small intestine. The objective of this study was to determine the effect of ABT-229 on canine gastric emptying (GE) and contractile activity of the antrum and duodenum in response to a solid meal. Six beagles were used to determine GE of a solid meal and contractile activity in response to either vehicle, ABT-229 (0.17, 0.83, 2.5, or 5.0 microg/kg/min), ERY (33.3 microg/kg/min), or cisapride (CIS) (10 microg/kg/min). Lag (t(lag)), half emptying (t(1/2)), and complete emptying (t(full)) times were determined. Contractile data were analyzed for motility index and gastroduodenal coordination. Compared with vehicle, ABT-229 dose dependently accelerated GE, t(lag) was decreased at the two highest doses, t(1/2) was decreased compared with vehicle at the three highest doses, and t(full) was decreased at all doses compared with vehicle. ERY also decreased t(1/2) and t(full), whereas CIS decreased all GE parameters. The slopes of the linear phase of GE curves for all drugs and doses were greater than those for vehicle. ABT-229 dose dependently increased the motility index as well as gastroduodenal coordination. ABT-229 (two highest doses) and CIS accelerated GE of a solid meal by decreasing the lag phase and increasing the rate of GE, whereas ERY only increased the rate of GE. The data suggest that ABT-229 is 7- to 40-fold more potent than ERY in accelerating GE. PMID- 10869417 TI - Inactivation of hepatic CYP2E1 by an epoxide of diallyl sulfone. AB - Diallyl sulfone (DASO(2)) inhibits hepatic CYP2E1. In this investigation, we have tested the hypothesis that an epoxide formed from DASO(2) is responsible for inactivation of hepatic CYP2E1 in mice. An epoxide of DASO(2) (1,2-epoxypropyl-3, 3'-sulfonyl-1'-propene; DASO(3)) was synthesized and conjugated to glutathione (GSH) to produce the conjugates S-(1R,S-[[ 1-hydroxymethyl-2,3'-sulfonyl]-1' propenyl]ethyl)glutathione (diastereomers) and S-(1-[[2R,S-hydroxypropyl]-3, 3' sulfonyl]-1'-propenyl)glutathione (diastereomers). Their identities were confirmed by (1)H NMR analysis, and these were used as analytical standards. HPLC analysis revealed a major peak for the GSH conjugates that eluted at 20.5 min. This peak was detected in liver microsomal incubations performed with DASO(2) in the presence of NADPH. A similar peak also was detected in incubations of CYP2E1 expressed lymphoblastoid microsomes, NADPH and DASO(2). The generation of the epoxide-derived GSH conjugates in the microsomal incubations was concentration dependent, and reached saturation at 0. 75 to 1.0 mM DASO(2). Formation of the conjugates was also time-dependent and peaked at 2.0 h after DASO(2). Levels of DASO(3) formed from DASO(2), as estimated by production of a 4-(p nitrobenzyl)pyridine derivative, were maximal at 1 mM DASO(2) at 30 min. CYP2E1 dependent p-nitrophenol hydroxylase activity was decreased in microsomes incubated with DASO(2), with alterations that were proportional to the concentration of DASO(2) (0.25-1.0 mM) used. Dose-dependent decreases in hydroxylase activity also were found in microsomes from mice treated in vivo with DASO(2) (25-200 mg/kg). These DASO(2)-induced decreases corresponded with reduced amounts of immunodetectable CYP2E1. Levels of spectrally detectable P450 and heme were both diminished by DASO(2). These results supported the contention that an epoxide formed from DASO(2) mediates the inactivation of hepatic CYP2E1. PMID- 10869418 TI - Expression of CX3CR1 chemokine receptors on neurons and their role in neuronal survival. AB - Recent in vitro and in vivo studies have shown that the chemokine fractalkine is widely expressed in the brain and localized principally to neurons. Central nervous system expression of CX(3)CR1, the only known receptor for fractalkine, has been demonstrated exclusively on microglia and astrocytes. Thus, it has been proposed that fractalkine regulates cellular communication between neurons (that produce fractalkine) and microglia (that express its receptor). Here we show, for the first time, that hippocampal neurons also express CX(3)CR1. Receptor activation by soluble fractalkine induces activation of the protein kinase Akt, a major component of prosurvival signaling pathways, and nuclear translocation of NF-kappaB, a downstream effector of Akt. Fractalkine protects hippocampal neurons from the neurotoxicity induced by the HIV-1 envelope protein gp120(IIIB), an effect blocked by anti-CX(3)CR1 antibodies. Experiments with two different inhibitors of the phosphatidylinositol 3-kinase, a key enzyme in the activation of Akt, and with a phospholipid activator of Akt demonstrate that Akt activation is responsible for the neuroprotective effects of fractalkine. These data show that neuronal CX(3)CR1 receptors mediate the neurotrophic effects of fractalkine, suggesting that fractalkine and its receptor are involved in a complex network of both paracrine and autocrine interactions between neurons and glia. PMID- 10869419 TI - Networks of interneurons with fast and slow gamma-aminobutyric acid type A (GABAA) kinetics provide substrate for mixed gamma-theta rhythm. AB - During active exploration, hippocampal neurons exhibit nested rhythmic activity at theta ( approximately 8 Hz) and gamma ( approximately 40 Hz) frequencies. Gamma rhythms may be generated locally by interactions within a class of interneurons mediating fast GABA(A) (GABA(A,fast)) inhibitory postsynaptic currents (IPSCs), whereas theta rhythms traditionally are thought to be imposed extrinsically. However, the hippocampus contains slow biophysical mechanisms that may contribute to the theta rhythm, either as a resonance activated by extrinsic input or as a purely local phenomenon. For example, region CA1 of the hippocampus contains a slower class of GABA(A) (GABA(A,slow)) synapses, believed to be generated by a distinct group of interneurons. Recent evidence indicates that these GABA(A,slow) interneurons project to the GABA(A, fast) interneurons that contribute to hippocampal gamma rhythms. Here, we use biophysically based simulations to explore the possible ramifications of interneuronal circuits containing separate classes of GABA(A,fast) and GABA(A,slow) interneurons. Simulated interneuronal networks with fast and slow synaptic kinetics can generate mixed theta-gamma rhythmicity under restricted conditions, including strong connections among each population, weaker connections between the two populations, and homogeneity of cellular properties and drive. Under a broader range of conditions, including heterogeneity, the networks can amplify and resynchronize phasic responses to weak phase-dispersed external drive at theta frequencies to either GABA(A,slow) or GABA(A,fast) cells. GABA(A, slow) synapses are necessary for this process of amplification and resynchronization. PMID- 10869420 TI - Galactolipid deficiency and abnormal chloroplast development in the Arabidopsis MGD synthase 1 mutant. AB - The lipid monogalactosyl diacylglycerol (MGD) is a major structural component of photosynthetic membranes in chloroplasts. Its formation is catalyzed by the enzyme MGD synthase. In many plants, MGD derives from two different biosynthetic pathways: the prokaryotic pathway, which operates entirely within the plastid, and the eukaryotic pathway, which involves steps in the endoplasmic reticulum. Here, we describe the identification and characterization of an Arabidopsis mutant with a defective MGD synthase gene (MGD1). The mutant was identified in a screen of T-DNA lines for individuals with defects in chloroplast biogenesis. It has a yellow-green phenotype that correlates with a approximately 50% deficiency in total chlorophyll per plant. A single T-DNA insertion is located adjacent to the transcription initiation site of the MGD1 gene, and the abundance of MGD1 mRNA is reduced by 75% compared with wild type. Correlation between steady-state MGD1 transcript levels and MGD synthase activity (also reduced by 75% in mgd1) suggests that MGD1 is the most important MGD synthase in green tissues. The amount of MGD in mutant leaves is reduced by 42% compared with wild type. MGD from the mutant contains 23% less 16:3 fatty acid and 10% more 18:3 fatty acid. Because 16:3 is a characteristic feature of MGD from the prokaryotic pathway, it is possible that MGD1 operates with some preference in the prokaryotic pathway. Finally, the MGD-deficiency of mgd1 is correlated with striking defects in chloroplast ultrastructure, strongly suggesting a unique role for MGD in the structural organization of plastidic membranes. PMID- 10869421 TI - Nonapoptotic neurodegeneration in a transgenic mouse model of Huntington's disease. AB - Huntington's disease (HD) is a fatal inherited neurodegenerative disorder characterized by personality changes, motor impairment, and subcortical dementia. HD is one of a number of diseases caused by expression of an expanded polyglutamine repeat. We have developed several lines of mice that are transgenic for exon 1 of the HD gene containing an expanded CAG sequence. These mice exhibit a defined neurological phenotype along with neuronal changes that are pathognomonic for the disease. We have previously observed the appearance of neuronal intranuclear inclusions, but did not find evidence for neurodegeneration. In this study, we report that all lines of these mice develop a late onset neurodegeneration within the anterior cingulate cortex, dorsal striatum, and of the Purkinje neurons of the cerebellum. Dying neurons characteristically exhibit neuronal intranuclear inclusions, condensation of both the cytoplasm and nucleus, and ruffling of the plasma membrane while maintaining ultrastructural preservation of cellular organelles. These cells do not develop blebbing of the nucleus or cytoplasm, apoptotic bodies, or fragmentation of DNA. Neuronal death occurs over a period of weeks not hours. We also find degenerating cells of similar appearance within these same regions in brains of patients who had died with HD. We therefore suggest that the mechanism of neuronal cell death in both HD and a transgenic mouse model of HD is neither by apoptosis nor by necrosis. PMID- 10869422 TI - A neuronal network model of macaque primary visual cortex (V1): orientation selectivity and dynamics in the input layer 4Calpha. AB - In this paper, we offer an explanation for how selectivity for orientation could be produced by a model with circuitry that is based on the anatomy of V1 cortex. It is a network model of layer 4Calpha in macaque primary visual cortex (area V1). The model consists of a large number of integrate-and-fire conductance-based point neurons, both excitatory and inhibitory, which represent dynamics in a small patch of 4Calpha-1 mm(2) in lateral area-which contains four orientation hypercolumns. The physiological properties and coupling architectures of the model are derived from experimental data for layer 4Calpha of macaque. Convergent feed-forward input from many neurons of the lateral geniculate nucleus sets up an orientation preference, in a pinwheel pattern with an orientation preference singularity in the center of the pattern. Recurrent cortical connections cause the network to sharpen its selectivity. The pattern of local lateral connections is taken as isotropic, with the spatial range of monosynaptic excitation exceeding that of inhibition. The model (i) obtains sharpening, diversity in selectivity, and dynamics of orientation selectivity, each in qualitative agreement with experiment; and (ii) predicts more sharpening near orientation preference singularities. PMID- 10869423 TI - Identification of renox, an NAD(P)H oxidase in kidney. AB - Oxygen sensing is essential for homeostasis in all aerobic organisms, but its mechanism is poorly understood. Data suggest that a phagocytic-like NAD(P)H oxidase producing reactive oxygen species serves as a primary sensor for oxygen. We have characterized a source of superoxide anions in the kidney that we refer to as a renal NAD(P)H oxidase or Renox. Renox is homologous to gp91(phox) (91-kDa subunit of the phagocyte oxidase), the electron-transporting subunit of phagocytic NADPH oxidase, and contains all of the structural motifs considered essential for binding of heme, flavin, and nucleotide. In situ RNA hybridization revealed that renox is highly expressed at the site of erythropoietin production in the renal cortex, showing the greatest accumulation of renox mRNA in proximal convoluted tubule epithelial cells. NIH 3T3 fibroblasts overexpressing transfected Renox show increased production of superoxide and develop signs of cellular senescence. Our data suggest that Renox, as a renal source of reactive oxygen species, is a likely candidate for the oxygen sensor function regulating oxygen-dependent gene expression and may also have a role in the development of inflammatory processes in the kidney. PMID- 10869424 TI - Mechanisms that direct ordered assembly of T cell receptor beta locus V, D, and J gene segments. AB - T cell receptor (TCR) beta variable region genes are assembled in progenitor T cells from germ-line Vbeta, Dbeta, and Jbeta segments via an ordered two-step process in which Dbeta to Jbeta rearrangements occur on both alleles before appendage of a Vbeta to a preexisting DJbeta complex. Direct joining of Vbeta segments to nonrearranged Dbeta or Jbeta segments, while compatible with known restrictions on the V(D)J recombination mechanism, are infrequent within the endogenous TCRbeta locus. We have analyzed mechanisms that mediate ordered Vbeta, Dbeta, and Jbeta assembly via an approach in which TCRbeta minilocus recombination substrates were introduced into embryonic stem cells and then analyzed for rearrangement in normal thymocytes by recombinase-activating gene 2 deficient blastocyst complementation. These analyses demonstrated that Vbeta segments are preferentially targeted for rearrangement to Dbeta as opposed to Jbeta segments. In addition, we further demonstrated that Vbeta segments can be appended to nonrearranged endogenous Dbeta segments in which we have eliminated the ability of Dbeta segments to join to Jbeta segments. Our findings are discussed in the context of the mechanisms that regulate the ordered assembly and utilization of V, D, and J segments. PMID- 10869425 TI - Gene genealogies reveal global phylogeographic structure and reproductive isolation among lineages of Fusarium graminearum, the fungus causing wheat scab. AB - During the past decade, the plant disease called scab or Fusarium head blight of wheat and barley has reached epidemic proportions in North America and elsewhere in the world. Scab is an economically devastating plant disease, not only because it causes significant reduction in seed yields and quality, but also because infested seeds are often contaminated with trichothecene and estrogenic mycotoxins that pose a serious threat to animal health and food safety. To test whether the primary etiological agent of scab, the fungus Fusarium graminearum, is panmictic throughout its range, allelic genealogies were constructed from six single-copy nuclear genes from strains selected to represent the global genetic diversity of this pathogen. Excluding one hybrid strain, all six genealogies recovered the same seven biogeographically structured lineages, suggesting that they represent phylogenetically distinct species among which gene flow has been very limited during their evolutionary history. Parsimony analysis of the combined data set comprising 7,120 aligned nucleotide characters resolved most relationships among the seven lineages of the F. graminearum clade and related fusaria included in the study. Phylogenetic evidence is also presented for introgressive hybridization and intragenic recombination among lineages of the F. graminearum clade in nature. PMID- 10869426 TI - pRB binds to and modulates the transrepressing activity of the E1A-regulated transcription factor p120E4F. AB - The retinoblastoma protein pRB is involved in the transcriptional control of genes essential for cell cycle progression and differentiation. pRB interacts with different transcription factors and thereby modulates their activity by sequestration, corepression, or activation. We report that pRB, but not p107 and p130, binds to and facilitates repression by p120(E4F), a ubiquitously expressed GLI-Kruppel-related protein identified as a cellular target of E1A. The interaction involves two distinct regions of p120(E4F) and the C-terminal part of pRB. In vivo pRB-p120(E4F) complexes can only be detected in growth-arrested cells, and accordingly contain the hypophosphorylated form of pRB. Repression of an E4F-responsive promoter is strongly increased by combined expression of p120(E4F) and pRB, which correlates with pRB-dependent enhancement of p120(E4F) binding activity. Elevated levels of p120(E4F) have been shown to block growth of mouse fibroblasts in G(1). We find this requires pRB, because RB(-/-) fibroblasts are significantly less sensitive to excess p120(E4F). PMID- 10869427 TI - Estrogens suppress RANK ligand-induced osteoclast differentiation via a stromal cell independent mechanism involving c-Jun repression. AB - Loss of ovarian function following menopause results in a substantial increase in bone turnover and a critical imbalance between bone formation and resorption. This imbalance leads to a progressive loss of trabecular bone mass and eventually osteoporosis, in part the result of increased osteoclastogenesis. Enhanced formation of functional osteoclasts appears to be the result of increased elaboration by support cells of osteoclastogenic cytokines such as IL-1, tumor necrosis factor, and IL-6, all of which are negatively regulated by estrogens. We show here that estrogen can suppress receptor activator of NF-kappaB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF)-induced differentiation of myelomonocytic precursors into multinucleated tartrate-resistant acid phosphatase-positive osteoclasts through an estrogen receptor-dependent mechanism that does not require mediation by stromal cells. This suppression is dose dependent, isomer-specific, and reversed by ICI 182780. Furthermore, the bone sparing analogues tamoxifen and raloxifene mimic estrogen's effects. Estrogen blocks RANKL/M-CSF-induced activator protein-1-dependent transcription, likely through direct regulation of c-Jun activity. This effect is the result of a classical nuclear activity by estrogen receptor to regulate both c-Jun expression and its phosphorylation by c-Jun N-terminal kinase. Our results suggest that estrogen modulates osteoclast formation both by down-regulating the expression of osteoclastogenic cytokines from supportive cells and by directly suppressing RANKL-induced osteoclast differentiation. PMID- 10869428 TI - Interaction of the iron-sulfur cluster assembly protein IscU with the Hsc66/Hsc20 molecular chaperone system of Escherichia coli. AB - The iscU gene in bacteria is located in a gene cluster encoding proteins implicated in iron-sulfur cluster assembly and an hsc70-type (heat shock cognate) molecular chaperone system, iscSUA-hscBA. To investigate possible interactions between these systems, we have overproduced and purified the IscU protein from Escherichia coli and have studied its interactions with the hscA and hscB gene products Hsc66 and Hsc20. IscU and its iron-sulfur complex (IscU-Fe/S) stimulated the basal steady-state ATPase activity of Hsc66 weakly in the absence of Hsc20 but, in the presence of Hsc20, increased the ATPase activity up to 480-fold. Hsc20 also decreased the apparent K(m) for IscU stimulation of Hsc66 ATPase activity, and surface plasmon resonance studies revealed that Hsc20 enhances binding of IscU to Hsc66. Surface plasmon resonance and isothermal titration calorimetry further showed that IscU and Hsc20 form a complex, and Hsc20 may thereby aid in the targeting of IscU to Hsc66. These results establish a direct and specific role for the Hsc66/Hsc20 chaperone system in functioning with isc gene components for the assembly of iron-sulfur cluster proteins. PMID- 10869430 TI - Assembly and activation of site-specific recombination complexes. AB - Site-specific recombination is responsible for a broad range of biological phenomena, including DNA inversion, resolution of transposition intermediates, and the integration and excision of bacteriophage genomes. Integration of mycobacteriophage L5 is catalyzed by a phage-encoded integrase with recombination occurring between specific attachment sites on the phage and mycobacterial chromosomes (attP and attB, respectively). Although some site-specific recombination systems simply involve binding of the recombinase to the sites of strand exchange, synapsis, and recombination, phage systems typically require the assembly of higher-order structures within which the recombinational potential of integrase is activated. The requirement for these structures derives from the necessity to regulate the directionality of recombination-either integration or excision-which must be closely coordinated with other aspects of the phage growth cycles. We show herein that there are multiple pathways available for the assembly of L5 recombination complexes, including the early synapsis of the attP and attB DNAs. This process is in contrast to the model for lambda integration and illustrates the different usage of molecular machineries to accomplish the same biological outcome. PMID- 10869429 TI - Requirement of the inducible nitric oxide synthase pathway for IL-1-induced osteoclastic bone resorption. AB - Nitric oxide has been suggested to be involved in the regulation of bone turnover, especially in pathological conditions characterized by release of bone resorbing cytokines. The cytokine IL-1 is thought to act as a mediator of periarticular bone loss and tissue damage in inflammatory diseases such as rheumatoid arthritis. IL-1 is a potent stimulator of both osteoclastic bone resorption and expression of inducible nitric oxide synthase (iNOS) in bone cells and other cell types. In this study, we investigated the role that the iNOS pathway plays in mediating the bone-resorbing effects of IL-1 by studying mice with targeted disruption of the iNOS gene. Studies in vitro and in vivo showed that iNOS-deficient mice exhibited profound defects of IL-1-induced osteoclastic bone resorption but responded normally to calciotropic hormones such as 1,25 dihydroxyvitamin D3 and parathyroid hormone. Immunohistochemical studies and electrophoretic mobility shift assays performed on bone marrow cocultures from iNOS-deficient mice showed abnormalities in IL-1-induced nuclear translocation of the p65 component of NFkappaB and in NFkappaB-DNA binding, which were reversed by treatment with the NO donor S-nitroso-acetyl penicillamine. These results show that the iNOS pathway is essential for IL-1-induced bone resorption and suggest that the effects of NO may be mediated by modulating IL-1-induced nuclear activation of NFkappaB in osteoclast precursors. PMID- 10869431 TI - In organello formaldehyde crosslinking of proteins to mtDNA: identification of bifunctional proteins. AB - The segregating unit of mtDNA is a protein-DNA complex called the nucleoid. In an effort to understand how nucleoid proteins contribute to mtDNA organization and inheritance, we have developed an in organello formaldehyde crosslinking procedure to identify proteins associated with mtDNA. Using highly purified mitochondria, we observed a time-dependent crosslinking of protein to mtDNA as determined by sedimentation through isopycnic cesium chloride gradients. We detected approximately 20 proteins crosslinked to mtDNA and identified 11, mostly by mass spectrometry. Among them is Abf2p, an abundant, high-mobility group protein that is known to function in nucleoid morphology, and in mtDNA transactions. In addition to several other proteins with known DNA binding properties or that function in mtDNA maintenance, we identified other mtDNA associated proteins that were not anticipated, such as the molecular chaperone Hsp60p and a Krebs cycle protein, Kgd2p. Genetic experiments indicate that hsp60 ts mutants have a petite-inducing phenotype at the permissive temperature and that a kgd2Delta mutation increases the petite-inducing phenotype of an abf2Delta mutation. Crosslinking and DNA gel shift experiments show that Hsp60p binds to single-stranded DNA with high specificity for the template strand of a putative origin of mtDNA replication. These data identify bifunctional proteins that participate in the stability of rho(+) mtDNA. PMID- 10869432 TI - In vivo urea cycle flux distinguishes and correlates with phenotypic severity in disorders of the urea cycle. AB - Urea cycle disorders are a group of inborn errors of hepatic metabolism that result in often life-threatening hyperammonemia and hyperglutaminemia. Clinical and laboratory diagnosis of partial deficiencies during asymptomatic periods is difficult, and correlation of phenotypic severity with either genotype and/or in vitro enzyme activity is often imprecise. We hypothesized that stable isotopically determined in vivo rates of total body urea synthesis and urea cycle specific nitrogen flux would correlate with both phenotypic severity and carrier status in patients with a variety of different enzymatic deficiencies of the urea cycle. We studied control subjects, patients, and their relatives with different enzymatic deficiencies affecting the urea cycle while consuming a low protein diet. On a separate occasion the subjects either received a higher protein intake or were treated with an alternative route medication sodium phenylacetate/benzoate (Ucephan), or oral arginine supplementation. Total urea synthesis from all nitrogen sources was determined from [(18)O]urea labeling, and the utilization of peripheral nitrogen was estimated from the relative isotopic enrichments of [(15)N]urea and [(15)N]glutamine during i.v. co-infusions of [5 (amide)(15)N]glutamine and [(18)O]urea. The ratio of the isotopic enrichments of (15)N-urea/(15)N-glutamine distinguished normal control subjects (ratio = 0.42 +/ 0.06) from urea cycle patients with late (0.17 +/- 0.03) and neonatal (0.003 +/- 0.007) presentations irrespective of enzymatic deficiency. This index of urea cycle activity also distinguished asymptomatic heterozygous carriers of argininosuccinate synthetase deficiency (0. 22 +/- 0.03), argininosuccinate lyase deficiency (0.35 +/- 0.11), and partial ornithine transcarbamylase deficiency (0.26 +/- 0.06) from normal controls. Administration of Ucephan lowered, and arginine increased, urea synthesis to the degree predicted from their respective rates of metabolism. The (15)N-urea/(15)N-glutamine ratio is a sensitive index of in vivo urea cycle activity and correlates with clinical severity. Urea synthesis is altered by alternative route medications and arginine supplementation to the degree that is to be expected from theory. This stable isotope protocol provides a sensitive tool for evaluating the efficacy of therapeutic modalities and acts as an aid to the diagnosis and management of urea cycle patients. PMID- 10869433 TI - A regulatory shortcut between the Snf1 protein kinase and RNA polymerase II holoenzyme. AB - RNA polymerase II holoenzymes respond to activators and repressors that are regulated by signaling pathways. Here we present evidence for a "shortcut" mechanism in which the Snf1 protein kinase of the glucose signaling pathway directly regulates transcription by the yeast holoenzyme. In response to glucose limitation, the Snf1 kinase stimulates transcription by holoenzyme that has been artificially recruited to a reporter by a LexA fusion to a holoenzyme component. We show that Snf1 interacts physically with the Srb/mediator proteins of the holoenzyme in both two-hybrid and coimmunoprecipitation assays. We also show that a catalytically hyperactive Snf1, when bound to a promoter as a LexA fusion protein, activates transcription in a glucose-regulated manner; moreover, this activation depends on the integrity of the Srb/mediator complex. These results suggest that direct regulatory interactions between signal transduction pathways and RNA polymerase II holoenzyme provide a mechanism for transcriptional control in response to important signals. PMID- 10869434 TI - Rapid and general profiling of protease specificity by using combinatorial fluorogenic substrate libraries. AB - A method is presented for the preparation and use of fluorogenic peptide substrates that allows for the configuration of general substrate libraries to rapidly identify the primary and extended specificity of proteases. The substrates contain the fluorogenic leaving group 7-amino-4 carbamoylmethylcoumarin (ACC). Substrates incorporating the ACC leaving group show kinetic profiles comparable to those with the traditionally used 7-amino-4 methylcoumarin (AMC) leaving group. The bifunctional nature of ACC allows for the efficient production of single substrates and substrate libraries by using 9 fluorenylmethoxycarbonyl (Fmoc)-based solid-phase synthesis techniques. The approximately 3-fold-increased quantum yield of ACC over AMC permits reduction in enzyme and substrate concentrations. As a consequence, a greater number of substrates can be tolerated in a single assay, thus enabling an increase in the diversity space of the library. Soluble positional protease substrate libraries of 137, 180 and 6,859 members, possessing amino acid diversity at the P4-P3-P2-P1 and P4-P3-P2 positions, respectively, were constructed. Employing this screening method, we profiled the substrate specificities of a diverse array of proteases, including the serine proteases thrombin, plasmin, factor Xa, urokinase-type plasminogen activator, tissue plasminogen activator, granzyme B, trypsin, chymotrypsin, human neutrophil elastase, and the cysteine proteases papain and cruzain. The resulting profiles create a pharmacophoric portrayal of the proteases to aid in the design of selective substrates and potent inhibitors. PMID- 10869435 TI - Regulation of histone deacetylase 4 and 5 and transcriptional activity by 14-3-3 dependent cellular localization. AB - Transcription is controlled in part by the dynamic acetylation and deacetylation of histone proteins. The latter process is mediated by histone deacetylases (HDACs). Previous analysis of the regulation of HDAC activity in transcription has focused primarily on the recruitment of HDAC proteins to specific promoters or chromosomal domains by association with DNA-binding proteins. To characterize the cellular function of the recently identified HDAC4 and HDAC5 proteins, complexes were isolated by immunoprecipitation. Both HDACs were found to interact with14-3-3 proteins at three phosphorylation sites. The association of 14-3-3 with HDAC4 and HDAC5 results in the sequestration of these proteins in the cytoplasm. Loss of this interaction allows HDAC4 and HDAC5 to translocate to the nucleus, interact with HDAC3, and repress gene expression. Regulation of the cellular localization of HDAC4 and HDAC5 by 14-3-3 represents a mechanism for controlling the transcriptional activity of these class II HDAC proteins. PMID- 10869436 TI - Deficient long-term memory and long-lasting long-term potentiation in mice with a targeted deletion of neurotrophin-4 gene. AB - We examined the learning and memory of neurotrophin-4 (NT4)-/- mice by using fear conditioning. In both cue and context conditioning, we found significant deficits in the NT4 mutants at 2 and 24 h after training but not at 30 min. Hippocampal slices from the mutant mice showed normal basal synaptic transmission, short-term plasticity, and decremental long-term potentiation (LTP) at the Schaffer collateral-CA1 synapses. These findings, together with the normal short-term memory, suggest that the hippocampal development of NT4-/- mice is largely unaffected. However, consistent with the long-term memory defects, the long lasting LTP at the same synapses was attenuated significantly in the mutant mice. Our results suggest that NT4 plays a physiological role essential for hippocampus and amygdala-dependent long-term memory and hippocampal long-lasting LTP and that NT4 may be useful in the therapy of acquired disorders of learning and memory. PMID- 10869437 TI - The transcriptional profile of early to middle sporulation in Bacillus subtilis. AB - Spore formation by Bacillus subtilis is governed by global changes in gene transcription. We used nylon-substrate DNA arrays representing approximately 96% of the predicted open reading frames in the B. subtilis chromosome to compare the pattern of transcripts from wild-type cells with the pattern from cells mutant for the sporulation transcription factors Spo0A or final sigma(F). We found 520 genes whose transcript levels were at least 3-fold dependent on Spo0A but not on final sigma(F), and an additional 66 genes whose transcript levels were dependent upon both regulatory proteins. Two strategies were used to help assign genes to the direct control of a particular developmental regulatory protein. In one approach, we analyzed the effects on global gene expression of artificially producing a constitutively active form of Spo0A during growth. In a second approach, Hidden Markov models were used to identify promoters likely to be activated by Spo0A, final sigma(F), or a third sporulation transcription factor, final sigma(E). In addition to detecting known sporulation genes, we identified many genes of unknown function whose patterns of expression and regulation suggest that they could be involved in sporulation. Disruption of two such newly identified genes, yabP and yabQ, blocked sporulation at a late stage. PMID- 10869438 TI - Nucleotide binding by the erythrocyte transglutaminase/Gh protein, probed with fluorescent analogs of GTP and GDP. AB - GTP is known to be a potent inhibitor of the protein crosslinking activity of transglutaminase (TG), probably the most abundant G protein in the human red cell. Nucleotide binding to TG was examined by fluorescence spectroscopy and anisotropy in mixtures of TG with methylanthraniloyl analogs of GTP and GDP. A characteristic feature was the appearance of a major energy transfer band (lambda(exc, max) = 290 nm, lambda(em) = 444 nm) from protein tryptophans to the bound nucleotides. Quenching of the bound fluorophore (lambda(exc) = 360 nm, lambda(em) = 444 nm) by acrylamide was barely different from that of free ligand. However, major changes were observed in anisotropy, which was used to demonstrate a facile exchange between bound and free nucleotides and to evaluate affinity constants for the binding of methylanthraniloyl GTP and GDP to TG. PMID- 10869439 TI - Ordered membrane insertion of an archaeal opsin in vivo. AB - The prevailing model of polytopic membrane protein insertion is based largely on the in vitro analysis of polypeptide chains trapped during insertion by arresting translation. To test this model under conditions of active translation in vivo, we have used a kinetic assay to determine the order and timing with which transmembrane segments of bacterioopsin (BO) are inserted into the membrane of the archaeon Halobacterium salinarum. BO is the apoprotein of bacteriorhodopsin, a structurally well characterized protein containing seven transmembrane alpha helices (A-G) with an N-out, C-in topology. H. salinarum strains were constructed that express mutant BO containing a C-terminal His-tag and a single cysteine in one of the four extracellular domains of the protein. Cysteine translocation during BO translation was monitored by pulse-chase radiolabeling and rapid derivatization with a membrane-impermeant, sulfhydryl-specific gel-shift reagent. The results show that the N-terminal domain, the BC loop, and the FG loop are translocated in order from the N terminus to the C terminus. Translocation of the DE loop could not be examined because cysteine mutants in this region did not yield a gel shift. The translocation order was confirmed by applying the assay to mutant proteins containing two cysteines in separate extracellular domains. Comparison of the translocation results with in vivo measurements of BO elongation indicated that the N-terminal domain and the BC loop are translocated cotranslationally, whereas the FG loop is translocated posttranslationally. Together, these results support a sequential, cotranslational model of archaeal polytopic membrane protein insertion in vivo. PMID- 10869440 TI - Mechanism of action of a pestivirus antiviral compound. AB - We report here the discovery of a small molecule inhibitor of pestivirus replication. The compound, designated VP32947, inhibits the replication of bovine viral diarrhea virus (BVDV) in cell culture at a 50% inhibitory concentration of approximately 20 nM. VP32947 inhibits both cytopathic and noncytopathic pestiviruses, including isolates of BVDV-1, BVDV-2, border disease virus, and classical swine fever virus. However, the compound shows no activity against viruses from unrelated virus groups. Time of drug addition studies indicated that VP32947 acts after virus adsorption and penetration and before virus assembly and release. Analysis of viral macromolecular synthesis showed VP32947 had no effect on viral protein synthesis or polyprotein processing but did inhibit viral RNA synthesis. To identify the molecular target of VP32947, we isolated drug resistant (DR) variants of BVDV-1 in cell culture. Sequence analysis of the complete genomic RNA of two DR variants revealed a single common amino acid change located within the coding region of the NS5B protein, the viral RNA dependent RNA polymerase. When this single amino acid change was introduced into an infectious clone of drug-sensitive wild-type (WT) BVDV-1, replication of the resulting virus was resistant to VP32947. The RNA-dependent RNA polymerase activity of the NS5B proteins derived from WT and DR viruses expressed and purified from recombinant baculovirus-infected insect cells confirmed the drug sensitivity of the WT enzyme and the drug resistance of the DR enzyme. This work formally validates NS5B as a target for antiviral drug discovery and development. The utility of VP32947 and similar compounds for the control of pestivirus diseases, and for hepatitis C virus drug discovery efforts, is discussed. PMID- 10869441 TI - The histidine kinase-related domain participates in phytochrome B function but is dispensable. AB - Phytochromes are photoreceptors that control many plant light responses. Phytochromes have two carboxyl-terminal structural domains called the PAS repeat domain and the histidine kinase-related domain. These domains are each related to bacterial histidine kinase domains, and biochemical studies suggest that phytochromes are light-regulated kinases. The PAS repeat domain is important for proper phytochrome function and can interact with putative signaling partners. We have characterized several new phytochrome B mutants in Arabidopsis that express phyB protein, three of which affect the histidine kinase-related domain. Point mutations in the histidine kinase-related domain cause phenotypes similar to those of null mutants, indicating that this domain is important for phyB signaling. However, a truncation that removes most of the histidine kinase related domain results in a phyB molecule with partial activity, suggesting that this domain is dispensable. These results suggest that phytochromes evolved in modular fashion. We discuss possible functions of the histidine kinase-related domain in phytochrome signaling. PMID- 10869442 TI - Toward a new anti-inflammatory and analgesic agent. PMID- 10869443 TI - Rapid restoration of visual pigment and function with oral retinoid in a mouse model of childhood blindness. AB - Mutations in the retinal pigment epithelium gene encoding RPE65 are a cause of the incurable early-onset recessive human retinal degenerations known as Leber congenital amaurosis. Rpe65-deficient mice, a model of Leber congenital amaurosis, have no rod photopigment and severely impaired rod physiology. We analyzed retinoid flow in this model and then intervened by using oral 9-cis retinal, attempting to bypass the biochemical block caused by the genetic abnormality. Within 48 h, there was formation of rod photopigment and dramatic improvement in rod physiology, thus demonstrating that mechanism-based pharmacological intervention has the potential to restore vision in otherwise incurable genetic retinal degenerations. PMID- 10869444 TI - Taming combinatorial explosion. PMID- 10869445 TI - Toxicological Highlight. PMID- 10869447 TI - Cardiovascular toxicity of inhaled ambient particulate matter. PMID- 10869448 TI - Mary O. Amdur. PMID- 10869449 TI - The practice of structure activity relationships (SAR) in toxicology. AB - Both qualitative and quantitative modeling methods relating chemical structure to biological activity, called structure-activity relationship analyses or SAR, are applied to the prediction and characterization of chemical toxicity. This minireview will discuss some generic issues and modeling approaches that are tailored to problems in toxicology. Different approaches to, and some facets and limitations of the practice and science of, SAR as they pertain to current toxicology analyses, and the basic elements of SAR and SAR-model development and prediction systems are discussed. Other topics include application of 3-D SAR to understanding of the propensity of chemicals to cause endocrine disruption, and the use of models to analyze biological activity of metal ions in toxicology. An example of integration of knowledge pertaining to mechanisms into an expert system for prediction of skin sensitization to chemicals is also discussed. This minireview will consider the utility of modeling approaches as one component for better integration of physicochemical and biological properties into risk assessment, and also consider the potential for both environmental and human health effects of chemicals and their interactions. PMID- 10869450 TI - Biliary and urinary excretion of inorganic arsenic: monomethylarsonous acid as a major biliary metabolite in rats. AB - In rats exposed to arsenite (AsIII) or arsenate (AsV), the biliary excretion of arsenic depends completely on availability of hepatic glutathione, suggesting that both AsIII and AsV are transported into bile in thiol-reactive trivalent forms (Gyurasics et al. [1991], Biochem. Pharmacol. 42, 465-468). To test this hypothesis, the bile and urine of bile duct-cannulated rats injected with AsIII or AsV (50 micromol/kg, iv) were collected periodically for 2 h and analyzed for arsenic metabolites by HPLC-hydride generation-atomic fluorescence spectrometry. Arsenic was excreted predominantly into bile in AsIII-injected rats, but the urine was the main route of excretion in AsV-exposed rats. Injected AsIII was excreted in urine practically unchanged, whereas both AsV and AsIII appeared in urine after administration of AsV. Irrespective of the arsenical administered, the bile contained 2 main arsenic species, namely AsIII and a hitherto unidentified metabolite. Formation of this metabolite could be prevented by pretreatment of the rats with the methylation inhibitor periodate-oxidized adenosine, indicating that it is a methylated arsenic compound. This metabolite could be converted in vitro into monomethylarsonic acid (MMAsV) by oxidation, whereas synthetic MMAsV could be converted into the unknown metabolite by reduction. Consequently, this biliary metabolite of both AsIII and AsV is monomethylarsonous acid (MMAsIII), a long-hypothesized, but never identified, intermediate in the biotransformation of AsIII and AsV. Although MMAsIII is thought to be formed from an oxidized precursor, rats injected with MMAsV did not excrete MMAsIII. In summary, the inorganic arsenicals investigated are transported into bile exclusively in trivalent forms, namely as AsIII and MMAsIII, but are excreted in urine in both tri- and pentavalent forms. Identification of MMAsIII is signified by the fact that this metabolite is more toxic than AsIII and AsV and thus formation of MMAsIII represents toxification of inorganic arsenic. PMID- 10869451 TI - Mu-class GSTs are responsible for aflatoxin B(1)-8, 9-epoxide-conjugating activity in the nonhuman primate macaca fascicularis liver. AB - Mice are resistant to the carcinogenic effects of the mycotoxin aflatoxin B(1) (AFB(1)) because they constitutively express an alpha-class glutathione S transferase (mGSTA3-3) that has high (approximately 200,000 pmol/min/mg) activity toward aflatoxin B(1)-8, 9-epoxide (AFBO). Rats do not constitutively express a GST with high AFBO-conjugating activity and are sensitive to AFB(1)-induced hepatocarcinogenesis. Constitutively expressed human hepatic alpha-class GSTs (hGSTA1-1 and hGSTA2-2) possess little or no AFBO-detoxifying activity (<2 pmol/min/mg). Recently, we found that the nonhuman primate, Macaca fascicularis (Mf), exhibits significant (approximately 300 pmol/min/mg) constitutive hepatic GST activity towards AFBO. To determine which specific GST isoenzyme(s) is (are) responsible for this activity, MF: GSTs were purified from liver tissue and characterized and, Mf mu-class GST cDNAs were cloned by reverse transcriptase coupled polymerase chain reaction (RT-PCR). Purification by glutathione agarose (GSHA) affinity chromatography yielded a protein, GSHA-GST, that exhibited relatively high AFBO-conjugating activity (239 pmol/min/mg) compared to other GST containing peaks. Western blotting and enzymatic activity analyses revealed that GSHA-GST belongs to the mu class. Two distinct mu-class GST cDNAs, mfaGSTM1 (GenBank accession # AF200709) and mfaGSTM2 (GenBank accession # AF200710), were generated by RT-PCR. CDNA-derived amino acid sequence analysis revealed that mfaGSTM1 and mfaGSTM2 share 97% and 96% homology with the human mu-class GSTs hGSTM4 and hGSTM2, respectively. In contrast to recombinant mfaGSTM1-1, which had no detectable AFBO-conjugating activity, mfaGSTM2-2 exhibited this activity at 333 pmol/min/mg. Activity profiles for the stereoisomers exo- and endo-AFBO, and of 1-chloro-2,4-dinitrobenzene of the purified protein GSHA-GST and recombinant mfaGSTM2-2, suggested that they are two distinct enzymes. Our results indicate that, in contrast to rodents, mu-class GSTs are responsible for the majority of AFBO-conjugating activity in the liver of Macaca fascicularis. PMID- 10869452 TI - Bioavailability of the genotoxic components in coal tar contaminated soils in Fischer 344 rats. AB - The effect of chemical aging on the bioavailability and subsequent genotoxicity of coal tar (CT)-contaminated soils was evaluated in a 17-day feeding study using Fischer 344 male rats. Rats consumed a control diet or diets amended with soil, 0.35% CT, or soil freshly prepared or aged for 9 months with 0.35% CT. Mild treatment-related microscopic lesions in liver tissue and elevated enzyme levels in serum were detected in all CT treatment groups. The (32)P-postlabeling assay was employed to determine DNA adduct formation in treated animals. All CT treatment groups induced DNA adducts in both the liver and lung. Adduct levels were 3-fold higher in lung DNA compared to hepatic DNA. After correcting adduct levels for total ingested polycyclic aromatic hydrocarbons (PAHs), a significant decrease (p < 0.05) in adduct levels was observed in both CT/soil treatment groups compared to CT control in liver and lung DNA. Adduct profiles of (32)P postlabeled hepatic and lung DNA displayed several nonpolar DNA adducts that comigrated with PAH-adducted calf thymus DNA standards as determined through both thin-layer chromatography (TLC) and high-pressure liquid chromatography (HPLC). These results suggest that soil, but not aging of contaminants in soil, decreases the bioavailability of genotoxic components in CT, as evidenced by DNA adduct analysis. PMID- 10869453 TI - Extrapolation of a PBPK model for dioxins across dosage regimen, gender, strain, and species. AB - A physiologically based pharmacodynamic (PBPK) model for 2,3,7, 8 tetrachlorodibenzo-p-dioxin (TCDD) was developed based on pharmacokinetic data from acute oral exposures of TCDD to female Sprague-Dawley rats (Wang et al., 1997, Toxicol Appl. Pharmacol 147, 151-168). In the present study, the utility of this model to predict the disposition of TCDD in male and female Sprague-Dawley and female Wistar rats exposed to TCDD through different dosage regimens was examined. The ability of the model to predict the disposition of 2-iodo-3,7,8 trichlorodibenzo-p-dioxin (ITrCDD) in mice (Leung, et al., 1990, Toxicol. Appl. Pharmacol. 103, 399-410) was also examined. The ability of the model to predict across routes of exposure was assessed with intravenous injection data (5.6 microg/kg bw) (Li et al., 1995, Fundam. Appl. Toxicol. 27, 70-76) in female rats. Analysis across gender extrapolations used data for male Sprague-Dawley rats exposed intravenously to 9.25 microg TCDD/kg bw (Weber et al., 1993, Fundam. Appl. Toxicol. 21, 523-534). The analysis of across-dosage regimen and stains of rats extrapolations were assessed using data from rats exposed to TCDD through a loading/maintenance dosage regimen (Krowke et al., 1989, Arch. Toxicol. 63, 356 360). The physiological differences between gender, strain, and species were taken into account when fitting the PBPK model to these data sets. The results demonstrate that the PBPK model for TCDD developed for female Sprague-Dawley rats exposed by acute oral dosing accurately predicts the disposition of TCDD, for different gender and strain of rats across varying dosage regimens, as well as in a strain of mice. Minimal changes in fitted parameters were required to provide accurate predictions of these data sets. This study provides further confirmation of the potential use of physiological modeling in understanding pharmacokinetics and pharmacodynamics. PMID- 10869454 TI - Assessment of the percutaneous absorption of trichloroethylene in rats and humans using MS/MS real-time breath analysis and physiologically based pharmacokinetic modeling. AB - The development and validation of noninvasive techniques for estimating the dermal bioavailability of solvents in contaminated soil and water can facilitate the overall understanding of human health risk. To assess the dermal bioavailability of trichloroethylene (TCE), exhaled breath was monitored in real time using an ion trap mass spectrometer (MS/MS) to track the uptake and elimination of TCE from dermal exposures in rats and humans. A physiologically based pharmacokinetic (PBPK) model was used to estimate total bioavailability. Male F344 rats were exposed to TCE in water or soil under occluded or nonoccluded conditions by applying a patch to a clipper-shaved area of the back. Rats were placed in off-gassing chambers and chamber air TCE concentration was quantified for 3-5 h postdosing using the MS/MS. Human volunteers were exposed either by whole-hand immersion or by attaching patches containing TCE in soil or water on each forearm. Volunteers were provided breathing air via a face mask to eliminate inhalation exposure, and exhaled breath was analyzed using the MS/MS. The total TCE absorbed and the dermal permeability coefficient (K(P)) were estimated for each individual by optimization of the PBPK model to the exhaled breath data and the changing media and/or dermal patch concentrations. Rat skin was significantly more permeable than human skin. Estimates for K(P) in a water matrix were 0.31 +/ 0.01 cm/h and 0.015 +/- 0.003 cm/h in rats and humans, respectively. K(P) estimates were more than three times higher from water than soil matrices in both species. K(P) values calculated using the standard Fick's Law equation were strongly affected by exposure length and volatilization of TCE. In comparison, K(P) values estimated using noninvasive real-time breath analysis coupled with the PBPK model were consistent, regardless of volatilization, exposure concentration, or duration. PMID- 10869455 TI - Effects of Di-2-ethylhexyl phthalate (DEHP) on gap-junctional intercellular communication (GJIC), DNA synthesis, and peroxisomal beta oxidation (PBOX) in rat, mouse, and hamster liver. AB - The present study evaluated the effect of di-2-ethylhexyl phthalate (DEHP) on gap junctional intercellular communication (GJIC), peroxisomal beta-oxidation (PBOX) activity, and replicative DNA synthesis in several rodent species with differing susceptibilities to peroxisome proliferator-induced hepatic tumorigenesis. A low (non-tumorigenic) and high (tumorigenic) dietary concentration of DEHP was administered to male F344 rats for 1, 2, 4, and 6 weeks. Additionally, a previously non-tumorigenic dose (1000 ppm) and tumorigenic dose of DEHP (12,000 ppm), as determined by chronic bioassay data, were examined following 2 weeks dietary administration. Male B6C3F1 mice were fed the non-tumorigenic concentration, 500 ppm, and the tumorigenic concentration, 6000 ppm, of DEHP for two and four weeks. The hepatic effects of low and high concentrations of DEHP, 1000 and 6000 ppm, were also examined in male Syrian Golden hamsters (refractory to peroxisome proliferator-induced tumorigenicity). In rat and mouse liver, a concentration-dependent increase in the relative liver weight, PBOX activity, and replicative DNA synthesis was observed at the earliest time point examined. Concurrent to these observations was an inhibition of GJIC. In hamster liver, a slight increase in the relative liver weight, PBOX activity, and replicative DNA synthesis was observed. However, these effects were not of the same magnitude or consistency as those observed in rats or mice. Furthermore, DEHP had no effect on GJIC in hamster liver at any of the time points examined (2 and 4 weeks). HPLC analysis of DEHP and its primary metabolites, mono-2-ethylhexyl phthalate (MEHP), and phthalate acid (PA), indicated a time- and concentration-dependent increase in the hepatic concentration of MEHP. At equivalent dietary concentrations and time points, the presence of MEHP, the primary metabolite responsible for the hepatic effects of DEHP, demonstrated a species-specific response. The largest increase in the hepatic concentration of MEHP was observed in mice, which was greater than the concentration observed in rats. The hepatic concentration of MEHP was lowest in hamsters. Hepatic concentrations of DEHP and phthalic acid were minimal and did not correlate with concentration and time. Collectively, these data demonstrate the inhibition of hepatic GJIC and increased replicative DNA synthesis correlated with the observed dose- and species-specific tumorigenicity of DEHP and may be predictive indicators of the nongenotoxic carcinogenic potential of phthalate esters. PMID- 10869456 TI - Nafenopin causes protein kinase C-mediated serine phosphorylation and loss of function of connexin 32 protein in rat hepatocytes without aberrant expression or localization. AB - The characteristics and mechanism of the inhibition of connexin-mediated gap junctional communication by the non-genotoxic rodent hepatocarcinogen, nafenopin, has been studied in rat hepatocytes. Nafenopin caused a time- and concentration dependent inhibition of dye coupling in hepatocytes as assessed by transfer of microinjected lucifer yellow. A half-maximum inhibitory effect of nafenopin occurred at approximately 50 microM, which was not cytotoxic. The inhibitory effect was reversible since a significant recovery of communication was observed 3 h after removal of the chemical. The protein kinase inhibitor Go6976 prevented the inhibition of dye coupling, but a tyrosine kinase inhibitor (genistein) did not. Connexin 32 and 26 protein expression, as assessed by immunoblotting, was similar in nafenopin-treated hepatocytes compared to controls, with the exception that in a 10-h culture with nafenopin, the level of connexin 26 was elevated compared to controls. Immunohistochemistry indicated that the localization of plaques containing connexin 32 was not affected in hepatocytes by nafenopin. Immunoprecipitated connexin 32 was, however, detected by an anti-phosphoserine antibody following nafenopin treatment, but not in controls. This serine phosphorylation was prevented in the presence of Go6976. The results give further support for a role of protein kinase C in the post-translational inactivation of connexin 32 function in rat hepatocytes by nafenopin. PMID- 10869457 TI - Potent competitive interactions of some brominated flame retardants and related compounds with human transthyretin in vitro. AB - Brominated flame retardants such as polybrominated diphenyl ethers (PBDEs), pentabromophenol (PBP), and tetrabromobisphenol A (TBBPA) are produced in large quantities for use in electronic equipment, plastics, and building materials. Because these compounds have some structural resemblance to the thyroid hormone thyroxine (T(4)), it was suggested that they may interfere with thyroid hormone metabolism and transport, e.g., by competition with T(4) on transthyretin (TTR). In the present study, we investigated the possible interaction of several brominated flame retardants with T(4) binding to TTR in an in vitro competitive binding assay, using human TTR and 125 I-T(4) as the displaceable radioligand. Compounds were tested in at least eight different concentrations ranging from 1.95 to 500 nM. In addition, we investigated the structural requirements of these and related ligands for competitive binding to TTR. We were able to show very potent competition binding for TBBPA and PBP (10.6- and 7.1-fold stronger than the natural ligand T(4), respectively). PBDEs were able to compete with T(4)-TTR binding only after metabolic conversion by induced rat liver microsomes, suggesting an important role for hydroxylation. Brominated bisphenols with a high degree of bromination appeared to be more efficient competitors, whereas chlorinated bisphenols were less potent compared to their brominated analogues. These results indicate that brominated flame retardants, especially the brominated phenols and tetrabromobisphenol A, are very potent competitors for T(4) binding to human transthyretin in vitro and may have effects on thyroid hormone homeostasis in vivo comparable to the thyroid-disrupting effects of PCBs. PMID- 10869458 TI - Respiratory sensitization to diphenyl-methane-4,4'-diisocyanate (MDI) in guinea pigs: impact of particle size on induction and elicitation of response. AB - The impact of particle size of aerosolized polymeric diphenylmethane-4,4' diisocyanate (MDI) for the induction and elicitation of respiratory sensitization was evaluated. Four groups of 16 female guinea pigs each received either the vehicle, repeated intradermal (id) injections (3 x 0.3% MDI), one high-level inhalation exposure of 15 min to 135 mg MDI/m(3) air using a small aerosol (MMAD approximately 1.7 microm) or large aerosol (MMAD approximately 3.8 microm). Three weeks later, animals were challenged subsequently with two ramped concentrations of MDI aerosol (average concentrations 16 and 49 mg/m(3) air, each for 15 min) and two different particle sizes, i.e., the MMAD was either approximately 1.6 microm or approximately 5.1 microm for the small- and large-size aerosol, respectively. Respiratory sensitization was assessed by two endpoints: the measurement of respiratory rate, and examination of influx of eosinophilic granulocytes into the mucosa and submucosa of the trachea, bronchi, and lung associated lymph nodes (LALN). The recruitment of eosinophilic granulocytes into bronchial tissues was subdivided as follows: muscularis mucosae, submucosa, and perivascular. From measurements of respiratory rate, it would appear that guinea pigs sensitized by id injections or by inhalation exposure with the large aerosol tended to display a higher responsiveness than naive controls when challenged with the small aerosol. The recruitment of eosinophilic granulocytes in the bronchial tissue was greater in both inhalation induction groups as compared to the vehicle control. It appears that there was a somewhat greater response in animals sensitized by id injections or by inhalation exposure with the large aerosol and challenged with the small aerosol. Topographically, this difference was apparent only at the bronchial perivascular level and lung-associated lymph nodes (LALN), whereas at the submucosal and muscularis mucosae level the impact on particle size tended to be less pronounced. In summary, this study suggests that a brief, high-level inhalation exposure of MDI aerosol caused a sensitization of bronchial tissues in guinea pigs. The higher sensitization potency of the large aerosol may possibly be related to a dosimetric phenomenon because of the greater fraction of deposition of large particles within the upper respiratory tract. Overall, challenge exposures with this type of irritant aerosol appear to evoke more consistent effects when the MMAD is in the range of approximately 2 rather than approximately 5 microm. PMID- 10869459 TI - Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) suppresses the humoral and cell-mediated immune responses to influenza A virus without affecting cytolytic activity in the lung. AB - The immune response to influenza virus is exquisitely sensitive to suppression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD); however, the cellular mechanisms underlying the suppressive effects of TCDD are unknown. Mice exposed to TCDD exhibited a dose-responsive increase in mortality following an otherwise non lethal influenza virus infection. Given that cytotoxic T lymphocytes (CTL) are generally thought to resolve primary infections in the lung, we tested the hypothesis that exposure to TCDD suppresses T-cell responsiveness, leading to decreased CTL in the lung. After infection with influenza virus, naive CD8+ lymphocytes are activated and differentiate in the mediastinal lymph node (MLN). In mice exposed to TCDD and infected with influenza virus, the number of CD8+ MLN cells was reduced 60% compared to vehicle-treated mice. Moreover, MLN cells from TCDD-treated mice failed to develop cytolytic activity, and the production of interleukin (IL)-2 and interferon (IFN)-gamma was suppressed. Exposure to TCDD also altered the production of virus-specific antibodies, decreased the recruitment of CD8+ cells to the lung, reduced the percentage and number of bronchoalveolar lavage cells bearing a CTL phenotype (CD8+CD44hiCD62L(l) degrees ), and suppressed IL-12 levels in the lung. Despite our findings that exposure to TCDD suppressed T cell-dependent functions, the cytolytic activity of lung lavage cells from TCDD and vehicle treated mice was equivalent, and IFN gamma levels in the lungs of mice treated with TCDD were enhanced 10-fold. Thus, while exposure to TCDD suppressed a number of responses associated with the development of adaptive immunity to influenza virus, a direct link between these effects and enhanced susceptibility to influenza remains unclear. PMID- 10869460 TI - Characterization of the ototoxicity of difluoromethylornithine and its enantiomers. AB - Difluoromethylornithine (DFMO) is an irreversible inhibitor of ornithine decarboxylase (ODC), the essential enzyme in mammalian polyamine biosynthesis (Pasic et al., 1997, Arch. Otolaryngol. Head Neck Surg. 123[12], 1281-1286). This cancer chemotherapeutic agent has significant ototoxic potential. Because the DFMO enantiomers differ in their ability to block ODC, the present study was designed to compare the ototoxic potential of each enantiomer with the racemic form of this drug for the rat and guinea pig. Determining differential ototoxicity of the enantiomers is one preliminary step in determining the optimal form of DFMO to use in human cancer chemotherapy. Daily intubation with D,L-DFMO does not produce any auditory dysfunction in rats with doses between 200 mg/kg/day and 1. 2 g/kg/day for up to 8 weeks, despite the fact that doses of 800 and 1200 mg/kg/day depressed body weight gain. In contrast to the data observed in rats, substantial ototoxicity was observed when guinea pigs were injected ip with doses of D,L-DFMO between 500 mg/kg/day and 1 g/kg/day. D,L-DFMO produced loss of compound action potential sensitivity, but not of cochlear microphonic amplitude. This finding correlated with histological data revealing loss of both outer and inner hair cells in the cochlea with inner more affected than outer hair cells, particularly in the basal turn. Higher exposure doses (2-3 g/kg/day) resulted in significant general toxicity including impaired growth and some mortality. When the enantiomers were evaluated in the guinea pig, it was found that 1 g/kg/day D-DFMO did not produce any significant hearing impairment, whereas 1 g/kg/day of the L-enantiomer of DFMO generated a threshold shift that surpassed that of 1 g/kg/day of the D,L-DFMO treatment. PMID- 10869461 TI - 2,4-Dichlorophenoxyacetic acid disrupts the cytoskeleton and disorganizes the Golgi apparatus of cultured neurons. AB - 2,4-Dichlorophenoxyacetic acid (2,4-D) is a potent neurotoxic herbicide widely used in agriculture. The basic mechanisms by which 2,4-D produces cell damage have not yet been determined. In this study we have examined the effects of 2,4-D in primary cultures of cerebellar granule cells in order to obtain insights into the possible mechanisms underlying the toxic effects of this herbicide. The results obtained indicate that a 24-hour exposure to 2,4-D produces a striking and dose-dependent inhibition of neurite extension. This phenomenon is paralleled by a significant reduction in the cellular content of both dynamic and stable microtubules, a disorganization of the Golgi apparatus, and an inhibition in the synthesis of complex gangliosides. Interestingly, 2,4-D inhibits the in vitro polymerization of purified tubulin. Taken together, the present observations raise the possibility that at least one basic mechanism underlying 2,4-D neurotoxicity involves an inhibition of microtubule assembly. That event may cause a decreased neurite outgrowth response, and could also explain the observed differences in the pattern of ganglioside biosynthesis and/or the disorganization of the Golgi apparatus. PMID- 10869462 TI - Gestational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin induces developmental defects in the rat vagina. AB - At puberty, female rats exposed in utero to 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exhibit a persistent thread of mesenchymal tissue surrounded by keratinized epithelium that partially occludes the vaginal opening. Our objective was to determine the earliest time during fetal development that morphological signs of this vaginal canal malformation could be detected and to obtain greater insight into mechanisms involved in this effect. Pregnant rats were administered a single dose of vehicle (control) or TCDD (1.0 microg/kg, po) on gestation day (GD) 15 and were sacrificed on GD 18, 19, 20, and 21 for histological evaluation of female. Gestational exposure to TCDD affected vaginal morphogenesis as early as GD 19, 4 days after exposure of pregnant dams. In exposed fetuses, the thickness of mesenchymal tissue between the caudal Mullerian ducts was increased, which resulted in a failure of the Mullerian ducts to fuse, a process normally completed prior to parturition. In addition, TCDD exposure appeared to inhibit the regression of Wolffian ducts. Thus, TCDD interferes with vaginal development by impairing regression of the Wolffian ducts, by increasing the size of interductal mesenchyme, and by preventing fusion of the Mullerian ducts. Taken together, these effects appear to cause the persistent vaginal thread defect observed in rats following in utero and lactational TCDD exposure. PMID- 10869463 TI - Effects of acetaminophen on preimplantation embryo glutathione concentration and development in vivo and in vitro. AB - This study investigated the effects of high doses of acetaminophen (APAP) on preimplantation embryos. Previous studies indicate that cleavage-stage embryos cannot synthesize reduced glutathione (GSH) de novo and may be sensitive to GSH depleting toxicants. Alternatively, there may be maternal mechanisms that protect the embryos from the adverse effects of these toxicants. To address these possibilities, we cultured two-cell stage embryos in 0, 375, 750, or 1500 microM APAP and evaluated GSH concentration and development. APAP depressed embryo development to the morula and blastocyst stages in vitro, but a decrease in embryo GSH concentration was not detected. Furthermore, administration of 800 or 1430 mg/kg APAP to female mice 12 h prior to embryo collection on day 2 of gestation, or administration of 800 mg APAP/kg/day from day -8 to day 1 or day 3 of gestation, did not significantly affect ovary or embryo GSH concentration or embryo development. Liver GSH, however, was significantly decreased. Moreover, no adverse effects on embryo development to term were observed after treatment of female mice with 1430 mg APAP/kg/day from day -8 to day 3 of gestation. In summary, in vitro embryos were adversely affected, in terms of development, by APAP. In vivo, large doses of APAP depleted liver GSH but did not affect development of preimplantation embryos. In conclusion, preimplantation embryos appear to be protected from GSH-depleting toxicants such as APAP in vivo. PMID- 10869464 TI - Influence of corn oil and diet on reproduction and the kidney in female Sprague Dawley rats. AB - The purpose of this study was to investigate the influence of corn oil administration on gestation, parturition, and lactation in rats, in conjunction with diets differing in composition of nutrients. Rats were divided into two groups, each fed different commercial pellets for rodents, CA-1 or CE-2, different from each other mainly in the source of protein. Female Sprague-Dawley rats in both diet groups were administered 0 (untreated control), 2, or 10 ml corn oil/kg body weight by gavage during the premating period (2 weeks), the mating period, the gestation period, and the lactation period (until day 3 of lactation). Food consumption of both the 10 ml/kg corn oil groups was significantly reduced throughout the study. Body weight gain in the 10 ml/kg corn oil group fed the CA-1 diet was significantly reduced on days 0 through 4 of lactation. Neither mating nor fertility indices were affected, and no clinical signs were observed during the gestation period in any groups. Several dams in the 10 ml/kg corn oil group fed the CA-1 diet, however, showed abnormal conditions after parturition, and three dams became moribund. Pup viability was also reduced in this group. Histopathologic examination of the kidneys of dams in the 10 ml/kg corn oil group fed the CA-1 diet revealed severe lesions in the proximal tubular epithelium, i.e., necrosis and fatty degeneration. Females in any group fed the CE-2 diet showed neither abnormal condition after parturition nor any severe lesions in the kidney. These data show that the combination of corn oil and diet with a particular constitution may cause adverse effects on the renal tubules in pregnant and/or lactating rats, suggesting that corn oil gavage as a vehicle can be a confounding factor in the reproductive toxicity studies, depending on the diet. PMID- 10869465 TI - Differential gene expression detected by suppression subtractive hybridization in the ethylene glycol monomethyl ether-induced testicular lesion. AB - The solvent ethylene glycol monomethyl ether (EGME) produces the same testicular lesions in rodents and human testis cultures, whose onset is characterized by apoptosis of pachytene spermatocytes. To identify gene changes early in the lesion and determine the possible involvement of cells other than the spermatocytes, we employed a suppression subtractive hybridization technique using whole testes from mice treated 8 h previously with 500 mg/kg EGME to generate two subtracted mouse testis cDNA libraries enriched for gene populations either up-regulated or down-regulated by EGME. A total of 70 clones were screened, and 6 of them were shown by Northern blotting to be differentially expressed in the EGME lesion. The three clones with increased expression after EGME treatment were identical to t-complex testis expressed gene 1 (tctex1), a gene encoding ribosomal protein S25, and a heretofore uncharacterized mouse testis expressed sequence tag. Three other genes suppressed by EGME were tctex2, alpha-2,6-sialyltransferase gene, and another uncharacterized mouse testis expressed sequence tag. Predicted peptide sequences of these clones contain multiple motifs for phosphorylation, glycosylation, and myristoylation. In situ hybridization with the antisense RNA probes further supported the expression changes of these six clones and localized the changes in multiple germ cell stages as well as other cell types (Sertoli, interstitial and peritubular cells). These data at the gene expression level are the first to demonstrate the early involvement in this lesion of cell types other than the dying spermatocytes. PMID- 10869466 TI - Oil fly ash-induced elevation of plasma fibrinogen levels in rats. AB - Particulate matter air pollution (PM) has been associated with morbidity and mortality from ischemic heart disease and stroke in humans. It has been hypothesized that alveolar inflammation, resulting from exposure to PM, may induce a state of blood hypercoagulability, triggering cardiovascular events in susceptible individuals. Previous studies in our laboratory have demonstrated acute lung injury with alveolar inflammation in rats following exposure to residual oil fly ash (ROFA), an emission source particulate. In addition, increased mortality has been documented following exposure to ROFA in rats with preexistent cardiopulmonary disease. ROFA's toxicity derives from its soluble metal content, which appears also to drive the toxicity of ambient PM. The present study was conducted to test the hypothesis that exposure of rats to a toxic PM, like ROFA, would adversely alter hemostatic parameters and cardiovascular risk factors thought to be involved in human epidemiologic findings. Sixty-day-old male Sprague-Dawley rats were exposed by intratracheal instillation (IT) to varying doses (0.3, 1. 7, or 8.3 mg/kg) of ROFA, 8.3 mg/kg Mt. Saint Helen's volcanic ash (MSH, control particle), or 0.3 ml saline (SAL, control). At 24 h post-IT, activated partial thromboplastin time (APTT), prothrombin time (PT), plasma fibrinogen (PF), plasma viscosity (PV), and complete blood count (CBC) were performed on venous blood samples. No differences from control were detected in APTT and PT in ROFA-exposed rats; however, ROFA exposure did result in elevated PF, at 8.3 mg/kg only. In addition, PV values were elevated in both ROFA and MSH-exposed rats relative to SAL-control rats, but not significantly. Although no changes were detected in APTT and PT, alteration of important hematologic parameters (notably fibrinogen) through PM induction of an inflammatory response may serve as biomarkers of cardiovascular risk in susceptible individuals. PMID- 10869467 TI - Effects of di-isononyl phthalate, di-2-ethylhexyl phthalate, and clofibrate in cynomolgus monkeys. AB - The effects of the peroxisome proliferators di-isononyl phthalate (DINP) and di-2 ethylhexyl phthalate (DEHP) were evaluated in young adult male cynomolgus monkeys after 14 days of treatment, with emphasis on detecting hepatic and other effects seen in rats and mice after treatment with high doses of phthalates. Groups of 4 monkeys received DINP (500 mg/kg/day), DEHP (500 mg/kg/day), or vehicle (0.5% methyl cellulose, 10 ml/kg) by intragastric intubation for 14 consecutive days. Clofibrate (250 mg/kg/day), a hypolipidemic drug used for cholesterol reduction in human patients was used as a reference substance. None of the test substances had any effect on body weight or liver weights. Histopathological examination of tissues from these animals revealed no distinctive treatment-related effects in the liver, kidney, or testes. There were also no changes in any of the hepatic markers for peroxisomal proliferation, including peroxisomal beta-oxidation (PBOX) or replicative DNA synthesis. Additionally, in situ dye transfer studies using fresh liver slices revealed that DINP, DEHP, and clofibrate had no effect on gap junctional intercellular communication (GJIC). None of the test substances produced any toxicologically important changes in urinalysis, hematology, or clinical chemistry; however, clofibrate produced some emesis, small increases in serum triglyceride, decreased calcium, and decreased weights of testes/epididymides and thyroid/parathyroid. The toxicological significance of these small changes is questionable. The absence of observable hepatic effects in monkeys at doses that produce hepatic effects in rodents suggests that DINP, DEHP, and clofibrate would also not elicit in primates other effects such as liver cancer. These data, along with results from in vitro hepatocyte studies, indicate that rodents are not good animal models for predicting the hepatic effects of phthalates in primates, including humans. PMID- 10869468 TI - Biomarkers of exposure to 1,3-butadiene as a basis for cancer risk assessment. AB - 1,3-Butadiene (BD) is carcinogenic in mice and rats, with mice being considerably more sensitive than rats. Urine metabolites are 1, 2-dihydroxybutyl mercapturic acid (DHBMA) and a mixture of monohydroxy-3-butenyl mercapturic acids (MHBMA). The reactive metabolite 1,2-epoxy-3-butene forms 1- and 2-hydroxy-3-butenyl valine adducts in hemoglobin (MHBVal). The objectives of the study were (1) to compare the suitability of MHBMA, DHBMA, and MHBVal as biomarkers for low levels of exposure to BD, and (2) to explore relative pathways of metabolism of BD in humans for comparison with mice and rats, which is important in relation to cancer risk assessment in man. Analytical methods of measuring MHBMA, DHBMA, and MHBVal were modified and applied in 2 studies to workers engaged in the manufacture and use of BD. Airborne BD concentrations were assessed by personal air monitoring. MHBMA in urine was more sensitive for monitoring recent exposures to BD when compared to DHBMA and could measure 8-h time weighted average exposures as low as 0.13 ppm. Relatively high natural background levels in urine restricted the sensitivity of DHBMA. The origin of this background is currently unknown. The measurement of MHBVal adducts in hemoglobin was a sensitive method for monitoring cumulative exposures to BD at or above 0.35 ppm. Statistically significant relationships were found between urinary MHBMA and DHBMA concentrations, between either of these variables and 8-h airborne BD levels and between MHBVal adducts and average airborne BD levels over 60 days. The data on biomarkers demonstrated a much higher rate of hydrolytic metabolism of 1,2-epoxy 3-butene in humans compared to mice and rats, which was reflected in a much higher DHBMA/(DHBMA + MHBMA) ratio and in much lower levels of MHBVal in humans. Assuming a genotoxic mechanism, the data of this study, coupled with other published data on DNA and hemoglobin binding in mice and rats, suggest that the cancer risk for man from exposure to BD is expected to be less than for the rat and much less than for the mouse. PMID- 10869469 TI - Pentoxifylline attenuates bacterial lipopolysaccharide-induced enhancement of allyl alcohol hepatotoxicity. AB - Small amounts of exogenous lipopolysaccharide (LPS) (10 ng/kg-100 microg/kg) enhance the hepatotoxicity of allyl alcohol in male Sprague-Dawley rats. This augmentation of allyl alcohol hepatotoxicity appears to be linked to Kupffer cell function, but the mechanism of Kupffer cell involvement is unknown. Since Kupffer cells produce tumor necrosis factor-alpha (TNF alpha) upon exposure to LPS, and this cytokine has been implicated in liver injury from large doses of LPS, we tested the hypothesis that TNF alpha contributes to LPS enhancement of allyl alcohol hepatotoxicity. Rats were treated with LPS (10-100 microg/kg iv) 2 h before allyl alcohol (30 mg/kg ip). Co-treatment with LPS and allyl alcohol caused liver injury as assessed by an increase in activity of alanine aminotransferase in plasma. Treatment with LPS caused an increase in plasma TNF alpha concentration, which was prevented by administration of either pentoxifylline (PTX) (100 mg/kg iv) or anti-TNF alpha serum (1 ml/rat iv) one h prior to LPS. Only PTX protected rats from LPS-induced enhancement of allyl alcohol hepatotoxicity; anti-TNF alpha serum had no effect. Exposure of cultured hepatocytes to LPS (1-10 microg/ml) or to TNF alpha (15-150 ng/ml) for 2 h did not increase the cytotoxicity of allyl alcohol (0.01-200 microM). These data suggest that neither LPS nor TNF alpha alone was sufficient to increase the sensitivity of isolated hepatocytes to allyl alcohol. Furthermore, hepatocytes isolated from rats treated 2 h earlier with LPS (i.e., hepatocytes which were exposed in vivo to TNF alpha and other inflammatory mediators) were no more sensitive to allyl alcohol-induced cytotoxicity than hepatocytes from naive rats. These data suggest that circulating TNF alpha is not involved in the mechanism by which LPS enhances hepatotoxicity of allyl alcohol and that the protective effect of PTX may be due to another of its biological effects. PMID- 10869470 TI - Metallothionein-null mice are more susceptible than wild-type mice to chronic CdCl(2)-induced bone injury. AB - Cadmium (Cd) is an environmental pollutant and is toxic to a number of organs. Chronic exposure to Cd causes loss of bone mass and increased incidence of bone fractures, as seen in Itai-itai patients and laboratory animals. Metallothionein (MT), a low-molecular weight, cysteine-rich, metal-binding protein, has been shown to play an important role in the detoxication of Cd. Thus, this study was designed to test the hypothesis that MT protects against Cd-induced bone injury. Wild-type and MT-I/II knockout (MT-null) mice were given repeated sc injections of CdCl(2) over a wide range of doses for 10 weeks, and Cd-induced bone injury was examined. Cd produced dose- and time-dependent increases in bone Cd content. However, the concentration of Cd in bone was much lower than that found in the liver and kidney (11 vs 400 and 120 microg/g, respectively) of the same mice. There was no difference in bone Cd content between wild-type and MT-null mice. Repeated Cd injections produced a dose-dependent loss of bone mass (up to 25%), as shown by analysis of the femur, tibia, and lumbar vertebrae. The loss of bone mass was more marked in MT-null mice than in wild-type mice, as shown by dry bone weight, defatted bone weight, bone ash weight, and total calcium content. X-ray photography showed decreasing bone density along the entire bone length with increasing dose and time of Cd exposure. The decrease in bone density was more marked in MT-null mice than in wild-type mice at the same dose and time points. Histopathology showed dilatation of haversian canals with increased osteoid seams, rounded osteocytes with expanded pericellular space, and expansion of hyperplastic bone marrow into metaphyseal cortical bone. Again, these lesions were more marked in MT-null mice. In conclusion, this study demonstrates that deficiency in MT renders animals more susceptible to Cd-induced bone mass loss and bone injury, and thus indicates that MT plays a protective role in Cd-induced toxicity in bone, as it does in other tissues. PMID- 10869471 TI - Clofibrate-induced in vitro hepatoprotection against acetaminophen is not due to altered glutathione homeostasis. AB - Prior induction of peroxisome proliferation protects mice against the in vivo hepatotoxicity of acetaminophen and various other bioactivation-dependent toxicants. The mechanisms underlying such chemoresistance are poorly understood, although they have been suggested to involve alterations in glutathione homeostasis. To clarify the role of glutathione in this phenomenon, we isolated hepatocytes from mice in which hepatic peroxisome proliferation had been induced with clofibrate. The cells were incubated with a range of acetaminophen concentrations and the extent of cell killing after up to 8 h was assessed by measuring lactate dehydrogenase leakage from the cells. Hepatocytes from clofibrate-pretreated mice were much less susceptible to acetaminophen than cells from vehicle-treated controls. However, the extent of glutathione depletion during exposure to acetaminophen was similar in both cell types, as were rates of excretion of the product of glutathione-mediated detoxication of acetaminophen's quinoneimine metabolite, 3-glutathionyl-acetaminophen. The glutathione replenishing ability of clofibrate-pretreated cells after a brief exposure to diethyl maleate also resembled that of control cells. More importantly, prior depletion of glutathione by diethyl maleate did not abolish the resistance of clofibrate-pretreated cells to acetaminophen. Taken together, these findings indicate that although glutathione-dependent pathways may contribute to hepatoprotection during peroxisome proliferation, the resistance phenomenon is not due exclusively to this mechanism. PMID- 10869472 TI - Gentamicin-induced apoptosis in renal cell lines and embryonic rat fibroblasts. AB - Gentamicin, an aminoglycoside antibiotic, induces apoptosis in the proximal tubule epithelium of rats treated at low, therapeutically relevant doses (El Mouedden et al., Antimicrob. Agents Chemother. 44, 665-675, 2000). Renal cell lines (LLC-PK(1) and MDCK-cells) have been used to further characterize and quantitate this process (electron microscopy; terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling of fragmented DNA [TUNEL]; and DNA size analysis [oligonucleosomal laddering]). Cells were exposed for up to 4 days to gentamicin concentrations of up to 3 mM. Apoptosis developed, almost linearly, with time and drug concentration, and was (i) preventable within the time-frame of the experiments by overexpression of the anti-apoptotic protein Bcl 2, and by co-incubation with cycloheximide (MDKC but not LLC-PK(1) cells); (ii) associated with an increased activity of caspases (MDCK cells; bcl-2 transfectants showed no increase of caspase activities and Z-VAD.fmk afforded full protection). Gentamicin-induced apoptosis also developed to a similar extent in embryonic fibroblasts cultured under the same conditions. In the 3 cell types, apoptosis (measured after 4 days) was directly correlated with cell gentamicin content (apoptotic index [approximately 10 to 18% of TUNEL (+) cells for a content of 20 microg of gentamicin/mg protein; kidney cortex of rats showing apoptosis in proximal tubule epithelium typically contains approximately 10 microg of gentamicin/mg protein). Thus, gentamicin has an intrinsic capability of inducing apoptosis in eucaryotic cells. Development of apoptosis in proximal tubules of kidney cortex in vivo after gentamicin systemic administration is therefore probably related to its capacity to concentrate in this epithelium after systemic administration. PMID- 10869473 TI - Purified fumonisin B(1) decreases cardiovascular function but does not alter pulmonary capillary permeability in swine. AB - Fumonisins are mycotoxins produced by Fusarium verticillioides, which induce acute pulmonary edema in swine. We previously reported that ingestion of fumonisin-containing culture material decreases cardiovascular function in swine (1996,a,b; Fundam. Appl. Toxicol. 31, 169-172; 33, 140-148; 1999, Am. J Vet. Res. 60, 1291-1300). The main purpose of this study was to confirm that fumonisin B(1) was responsible for the observed cardiovascular changes. Treated pigs (n = 6) were given daily intravenous injections of purified fumonisin B(1) at 1 mg/kg for 4 days, while controls (n = 6) were injected with equal volumes of saline. On day 5, pigs were anesthetized with butorphanol-chloralose and instrumented for hemodynamic studies. Terminally, bronchoalveolar lavage was performed on each pig to determine the relative permeability index of the pulmonary endothelium. Fumonisin B(1)-treated pigs had marked decreases in the maximal rate of change of left ventricular pressure (dP/dt(max)), mean aortic pressure, cardiac output, and arterial pO(2), accompanied by increases in mean pulmonary artery pressure, oxygen extraction ratio, and blood hemoglobin concentration. Plasma and left ventricular sphingosine and sphinganine concentrations were markedly increased in treated pigs at day 5; however, there was no difference in the relative permeability index between groups. Serum cholesterol concentrations and activities of hepatic-derived enzymes were increased, and hepatocyte apoptosis and mitoses were present in the livers of fumonisin-treated pigs. In the lungs of treated pigs, there was proteinaceous edema and membranous accumulations in capillary endothelial cells. These results indicate that cardiovascular function is altered by fumonisin B(1), and that fumonisin-induced pulmonary edema is caused by left-sided heart failure and not by altered endothelial permeability. Because of the potential for contamination of human foodstuffs by fumonisins, the cardiovascular toxicity of these compounds must be taken into consideration. PMID- 10869474 TI - Effects of low concentrations of paraoxon on Ca(2+) mobilization in a human parotid salivary cell-line HSY. AB - The salivary gland is a target organ of organophosphate pesticides (OPs). Inhibition of acetylcholinesterase (AChE) by OPs leads to a decrease in acetylcholine (ACh) breakdown that results in overstimulation of muscarinic cholinergic receptors (mChR). However, OPs may also directly interact with downstream elements of the phosphoinositide (PI) signalling pathway coupled with mChR. The present study examined the effects of exposure to low concentrations of the OP paraoxon on inositol 1,4,5-trisphosphate (IP(3)) formation and Ca(2+) mobilization in response to ACh or ATP in the human parotid cell-line HSY. Exposure to 0.1 and 1 nM, but not 10 nM, paraoxon for 24 hr significantly elevated the basal cytosolic free Ca(2+) ([Ca(2+)](i)). This increase was abolished by atropine. Ca(2+) release from the IP(3)-sensitive store in response to ACh or ATP, a P2Y-nucleotide agonist, was significantly increased in cells pre exposed to 0.1 nM paraoxon. However, IP(3) formation was inhibited by paraoxon but mChR expression was not altered. Although IP(3) receptor expression was not changed, Ca(2+) release elicited by IP(3) in streptolysin O toxin-permeabilized cells was significantly larger in cells pre-exposed to 0.1 nM paraoxon, suggesting that paraoxon increases the sensitivity of IP(3) receptors. Paraoxon exposure also induced a concentration-dependent reduction in the total capacity of intracellular Ca(2+) stores, whereas the capacity of the IP(3)-sensitive Ca(2+) store was not altered by paraoxon, as judged by discharging of the IP(3) sensitive Ca(2+) store with thapsigargin (TG). Ca(2+) influx stimulated by ACh or ATP was also enhanced by 0.1 nM, but not 1 and 10 nM, paraoxon. On the other hand, Ca(2+) influx activated by TG was enhanced by exposure to all concentrations of paraoxon, indicating that paraoxon modulates the Ca(2+) entry pathway. These results suggest that low concentrations of paraoxon interact with elements of the PI pathway, enhancing Ca(2+) release and influx mechanisms. PMID- 10869475 TI - Caries inhibitory activity of cacao bean husk extract in in-vitro and animal experiments. AB - Cacao bean husk extract (CBH) was examined for inhibitory effects on the caries inducing properties of mutans streptococci in vitro and on caries development in specific pathogen-free Sprague-Dawley rats infected with mutans streptococci. CBH reduced the growth rate of almost all oral streptococci examined, which resulted in the reduction of acid production. Furthermore, insoluble glucan synthesis by the glucosyltransferases from Streptococcus mutans MT8148R and Streptococcus sobrinus 6715 was significantly inhibited by CBH. Hence, the sucrose-dependent cell adherence of mutans streptococci was also depressed by CBH. The administration of CBH in drinking water resulted in significant reductions of caries development and dental plaque accumulation in rats infected with either Strep. sobrinus 6715 or Strep. mutans MT8148R, and the minimum cariostatic concentration was 1.0 mg/ml. These results indicate that CBH possesses powerful anticariogenic potential. PMID- 10869476 TI - Evidence for the involvement of bone morphogenetic protein-2 in phenytoin stimulated osteocalcin secretion in human bone cells. AB - Recent work has shown that the actions of phenytoin on bone cell proliferation and differentiation are, in part, mediated through the upregulation of transforming growth factor-beta1 (TGF-beta(1)). The present study was undertaken to examine the effect of phenytoin on bone morphogenetic proteins (BMP)-2 and -4, which are well-recognized osteoinductive proteins of the TGF-beta superfamily, in osteoblastic cells. Treatment with 5-50 microM of phenytoin increased the amount of mRNA for BMP-2 after a 0.5-24 h incubation in normal human mandible-derived bone cells (HOB-M cells), but failed to affect the mRNA for BMP-4. Phenytoin treatment for 48 h significantly increased the secretion of BMP-2 by approx. four fold, at an optimal concentration of 10 microM. While TGF-beta(1) inhibited osteocalcin secretion from HOB-M cells, both phenytoin and BMP-2 significantly stimulated it. Importantly, the stimulatory effects of phenytoin on osteocalcin release were completely blocked by the neutralizing antihuman BMP-2 monoclonal antibody. These results indicate that the stimulatory action of phenytoin on osteocalcin secretion in normal human bone cells is mediated, at least partly, through the upregulation of BMP-2, rather than that of TGF-beta(1). PMID- 10869478 TI - Effects of Streptococcus crista and human saliva on the viability of Fusobacterium nucleatum ATCC 10953. AB - Associations with facultative species might help planktonic oral anaerobes survive when they traverse saliva. This study investigated whether co-aggregation with Streptococcus crista ATCC 51110 enhanced the viability of Fusobacterium nucleatum ATCC 10953. Two tubes each of co-aggregates and Fus. nucleatum ATCC 10953 alone were prepared. One of each set was resuspended in buffer, and the other in clarified saliva from each of 20 donors. After 1 h, cells were stained for viability. The median percentage of viable Fus. nucleatum ATCC 10953 in buffer with Strep. crista (86%) was significantly higher (p=0.04) than in buffer alone (77%). A similar trend was observed for saliva with Strep. crista (81%) and saliva alone (74%), although that difference was not significant (p=0.41). The median percentage of Fus. nucleatum co-aggregated in buffer (44%) was significantly reduced after incubation in saliva (16%) (p<0.0002). No such change was seen when saliva was replaced with purified salivary proline-rich glycoprotein, which can bind both species. For co-aggregate suspensions, there was no difference in the viability of fusobacteria that were or were not in direct contact with Strep. crista. In both cases, viability was significantly reduced in saliva relative to buffer. Strep. crista may enhance the viability of planktonic Fus. nucleatum ATCC 10953, but it is not yet clear whether that requires co-aggregation. Transmission of fusobacteria through saliva could depend on the interplay between protective factors, such as the presence of streptococci, and antimicrobial factors, which kill cells or disassociate co aggregates. PMID- 10869477 TI - Upregulation of the expression of Fas antigen and Fas ligand in a human submandibular gland ductal cell line by okadaic acid. AB - Fas receptor is a member of a superfamily of receptors characterized by cysteine rich motifs in the extracellular domain of the molecule. Binding of Fas ligand to the receptor leads to receptor activation and the induction of intracellular signals that result in apoptotic cell death. In the present study, the expression of mRNA and proteins of Fas receptor and Fas ligand were examined in human submandibular gland ductal (HSG) cells treated with okadaic acid by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblot analysis. Six hundred and eighty-two bp of the PCR product of Fas receptor mRNA was detected in HSG cells and a protein with an estimated molecular weight of 58,000 was expressed in HSG cells. Treatment of HSG cells with an agonistic anti-Fas monoclonal antibody resulted in death of HSG cells, indicating that the functional Fas receptor protein is expressed in HSG cells. Fas receptor protein expression stimulated by okadaic acid was elevated in a dose- and time-dependent manner, with maximal expression at 20 nM and 48 h treatment. Fas ligand mRNA was also detected constitutively in HSG cells by RT-PCR. Okadaic acid stimulated the expression of Fas ligand protein in HSG cells in a time-dependent manner, while the expression of the ligand was low in untreated HSG cells. The molecular weight of Fas ligand was estimated as 68,000. An antagonistic anti-Fas ligand monoclonal antibody prevented okadaic acid-induced death in HSG cells in a dose-dependent fashion as determined by WST-1 assay. The results indicate that the expression of Fas receptor and ligand is regulated by protein phosphatase(s) sensitive to okadaic acid and is involved in okadaic acid-induced apoptosis in HSG cells. The results also suggest that the Fas receptor-ligand system might regulate apoptosis in HSG cells. PMID- 10869479 TI - Temporal coordination between mandibular and head-neck movements during jaw opening-closing tasks in man. AB - Previous finding of concomitant mandibular and head movements during jaw function suggest a functional relation between the human jaw and neck regions. This study examined the temporal coordination between mandibular and head-neck movements during maximal jaw opening-closing tasks, at fast and slow speed. Twenty-four healthy individuals, median age 25 years, participated in the study. They were seated with firm back support but without head-neck support. Mandibular and head movements were simultaneously monitored by a wireless optoelectronic system for three-dimensional movement recording. The timing of head movement in relation to mandibular movement was estimated at defined time-points (start, peak, end and maximum velocity of movement), and during the entire course of the jaw-opening and jaw-closing phases. The results showed that the head in general started to move simultaneously with or before the mandible, reached the peak position simultaneously with, before or after the mandible, and reached the end position after the mandible. A higher degree of temporal coordination was found for fast speed at the start and the peak positions. The head most often attained maximum velocity after the mandible, and mostly lagged behind the mandible during the entire jaw-opening and -closing phases. These findings support the notion of a functional linkage between the human temporomandibular and craniocervical regions. They suggest that "functional jaw movements" comprise concomitant mandibular and head-neck movements which involve the temporomandibular, the atlanto-occipital and the cervical spine joints, and are caused by jointly activated jaw and neck muscles. It is proposed that these jaw and neck muscle actions, particularly at fast speed, are elicited and synchronized by preprogrammed neural command(s) common to both the jaw and the neck motor systems. From the present results and previous observations of concurrent jaw and head movement during fetal yawning, we suggest that these motor programmes are innate. PMID- 10869480 TI - Investigation of sexual dimorphism in the rabbit masseter muscle showing different effects of androgen deprivation in adult and young adult animals. AB - To evaluate the role played by androgens in the development and maintenance of sex differences in the proportion of muscle fibres of different phenotypes, the effects of castration in adult (>6 months old) and in young adult (2-3 months old) male rabbits was examined. Immunohistochemical methods were used to evaluate the proportion of muscle fibres containing different myosin heavy-chain isoforms in 10 different neuromuscular compartments of the masseter. In young adult animals of both sexes, the proportion of fibres of different phenotypes in different compartments was not significantly different from that of normal adult females. In animals castrated as young adults, the development of adult male phenotype proportions was completely blocked in most compartments. In animals castrated as adults, proportions were not significantly different from those of the intact males. For most masseter compartments, androgens produced permanent changes in muscle fibre phenotype during a critical period of postnatal development. However, in the posterior deep compartment, androgen deprivation in young adults had no effect on phenotype proportions, but castration of adults resulted in a striking increase in the proportion of fibres containing the IIa myosin heavy-chain isoform. PMID- 10869481 TI - Correlations between functional and occlusal tooth-surface areas and food texture during natural chewing sequences in humans. AB - The dental-arch surfaces preferentially used in mastication were studied by measuring functional and occlusal surface areas and comparing these to the number of chews required to swallow foods of different texture properties. The functional surface of the teeth was defined as the total area of visible wear facets on post-incisal teeth, adding to it the contacting areas of restored teeth where no facets were visible. Occlusal surface area was taken as the total area of the occluding parts of post-incisal teeth. Both surfaces were measured with computer image processing on dental-stone casts of the teeth of 31 young adults. Functional surface areas (mean 168 mm(2), four quadrants) were positively correlated with occlusal surface areas (mean 739 mm(2), four quadrants). The left:right area ratios were more variable for functional than for occlusal surfaces. Functional surface-area ratios markedly different from 1.0 might reflect functional side-preponderance of masticatory activity. Correlations between tooth surface area and the number of cycles were examined with five different food samples of known texture during side-imposed mastication. Depending on the elastic moduli of the foods, significant negative correlations were found between the left:right ratios of functional or occlusal surface areas and the left:right ratios of cycle numbers. The rheological properties of the food particles chewed were assumed to be the key factor in the correlations with either the functional or anatomical occlusal surfaces. PMID- 10869483 TI - Inhibition of mitosis and induction of apoptosis in MG63 human osteosarcoma derived cells in vitro by surface proteins from Actinobacillus actinomycetemcomintans. AB - Gentle saline extraction releases a heterogeneous mixture of proteins associated with the cell surface of Actinobacillus actinomycetemcomintans, termed the surface protein fraction (SF). Some SF components are biologically active and may modulate cell behaviour in a manner of putative importance in the aetiology of periodontitis. To further characterize this activity, the ability of the SF to induce mitosis and apoptosis in MG63 cells was investigated. Cells were plated at 10(3)-10(4) cells/cm(2) and allowed to attach before culture in the serum-free medium in the presence of 25 microg/ml SF for 2-24 h. The apoptotic and mitotic figures present were counted and the results expressed as an apoptotic or mitotic index. The apoptotic and mitotic compartments were very small, but there was an inverse correlation between mitosis and apoptosis. In control experiments the mitotic was higher than the apoptotic index, whilst in the presence of SF this was reversed. These results were confirmed using in situ end-labelling. SF, therefore, may stimulate apoptotic, but inhibit mitotic, activity in MG63 cells. This raises the possibility that components of SF might induce subtle changes in the balance between apoptosis and mitosis, which, in turn, could contribute to the progression of periodontitis. PMID- 10869482 TI - Blockade of isoprenaline-induced fluid and protein secretion by the mandibular glands of the red kangaroo, Macropus rufus, with selective antagonists. AB - Selective and non-selective beta-adrenoceptor antagonists were used to block the increases in fluid and protein secretion caused by sympathomimetic stimulation of the mandibular gland of red kangaroos during intracarotid infusion of isoprenaline. Atenolol or ICI118551 at antagonist:agonist ratios up to 300:1 caused increasing but incomplete blockade of fluid secretion and protein release. Both selective antagonists had equal potency and both antagonists were more effective at blocking protein release than at blocking fluid secretion. Consequently, the mechanisms underpinning fluid secretion are more sensitive to beta-sympathomimetic stimulation than those causing protein release. Propranolol at antagonist:agonist ratios of 300:1 was more potent than the selective antagonists, almost totally blocking the increases in fluid secretion and protein release. The data are consistent with the acini of the kangaroo mandibular gland having both beta(1)- and beta(2)-adrenoceptors and with the increased fluid secretion and protein release by isoprenaline being mediated by both receptor subtypes. PMID- 10869484 TI - Crown formation times of human permanent anterior teeth. AB - One gap in knowledge of human dental-growth standards is the age at which crown fractions of anterior permanent teeth are attained. The aim of this study was to document stages of crown formation for permanent incisors and canines from a small skeletal collection of known age. The source was C18th and C19th coffin buried skeletal material from Spitalfields in London; developing teeth from 50 individuals with recorded age-at-death (range 0-5.40 years) and 56 unaged individuals were assessed. Teeth were dissected and crown height measured directly. Each developing crown was assigned to the nearest average fraction (C14, C12, C34, Cc). These fractions were calculated from the total crown height of unworn completed teeth from this sample. Median age for C12 of the permanent upper central incisor was 1.34 years (n=16) and for the canine was 2.52 years (n=16). Data on crown formation are also presented in relation to permanent lower first molar stages C12, C34 and Cc. When M(1) was at stage C34 the modal stage for I(1) was C34 and for other incisors and canines was C12. Although the sample is small, these results fill an important gap in tooth chronology and add to knowledge of growth variation in early childhood. PMID- 10869486 TI - Effect of donor age on the concentrations of histatins in human parotid and submandibular/sublingual saliva. AB - Histatins are small proteins of human glandular saliva that have antifungal properties. Recent studies show that oral candidal infections increase with age, suggesting an age-associated compromise in oral host defence. Here, the effect of age and of physiological gland stimulation on the concentration and secretion of salivary histatins was investigated. Parotid and submandibular/sublingual salivas were collected from six young adults under unstimulated, mechanical (chewing) and gustatory (0.025 M and 0.1 M citric acid) stimulation, and the concentration and secretion of histatins was measured by cationic polyacrylamide gel electrophoresis with subsequent densitometric scanning of the stained gels. With gland stimulation, parotid saliva showed no significant increase in histatin concentration (microg/ml); however, histatin secretion (microg/min) increased up to 26-fold (p<0.005; ANOVA). Stimulation of submandibular/sublingual saliva resulted in significant increases in both histatin concentration (p<0.005) and secretion (p<0.0005). Ageing effects on salivary histatins were determined in citric acid (0.1 M)-stimulated parotid and submandibular/sublingual saliva samples collected from 80 individuals (divided into four age groups having approximately equal numbers of males and females: 35-44 years; 45-54 years; 55-64 years and 65-76 years). None of the patients was taking medications or wore dentures. ANOVA showed no sex differences in histatins. Regression analysis showed significant age-associated decreases for parotid saliva histatin concentration (p<0.002) and secretion (p<0. 002) as well as for submandibular/sublingual saliva histatin concentration (p<0.0001) and secretion (p<0.0001). Both saliva types showed significant (p<0.0001) decreases in the histatin concentration per mg of total protein, suggesting a preferential decrease in salivary histatins compared to total salivary protein. These results suggest that the salivary histatin component of the oral host defence system is compromised with increasing age. PMID- 10869485 TI - Involvement of the endogenous nitric oxide signalling system in bradykinin receptor activation in rat submandibular salivary gland. AB - Biochemical signalling events coupled to the bradykinin B(2)-receptor subtype, related to nitric oxide and prostaglandin E(2) generation were studied in rat submandibular gland. Bradykinin stimulation of the B(2)-receptor triggered activation of phosphoinositide turnover, translocation of protein kinase C, stimulation of nitric oxide synthase activity, increased production of cGMP and release of prostaglandin E(2). Bradykinin stimulation of nitric oxide synthase and cGMP production was blunted by agents able to interfere with calcium/calmodulin and phospholipase C activities, while a protein kinase C inhibitor was able to stimulate bradykinin action. Moreover, a specific B(2) bradykinin antagonist of the reversible nitric oxide synthase inhibitor abrogated the bradykinin stimulation of nitric oxide synthase activity, cGMP accumulation and prostaglandin E(2) generation. Furthermore, a specific inhibitor of phospholipase A(2) blocked the bradykinin-induced prostaglandin E(2) release. These results suggest that apart, from the direct effect of bradykinin as an inducer of vasopermeability, it also appears to be a vasoactive chemical mediator that triggers, through release of prostaglandin E(2), a feedback mechanism that induces a protective adaptation of the gland, modulating the course of inflammation. PMID- 10869487 TI - Partial protection of rat parotid glands from irradiation-induced hyposalivation by manganese superoxide dismutase. AB - Head-and-neck irradiation in rats often results in reduction of water and food intake, weight loss, hyposalivation, and suppression of the white blood cell (WBC) count. Oxygen free radicals are believed to be involved in this deleterious process. Superoxide dismutase (SOD) is known to act as a first line of antioxidant defence against oxygen free radicals. Here, the protective effect of manganese SOD (MnSOD) and copper/zinc SOD (Cu/ZnSOD) against irradiation-induced injuries to the head and neck in rats was investigated. Wistar rats were irradiated with 15 Gy X-rays delivered to the head-and-neck region. MnSOD (50 mg/kg) or Cu/ZnSOD (100 mg/kg) were administered before and after irradiation. Body weight, food and water intake, WBC counts, and parotid and submandibular salivary functions were examined. Irradiation of 15 Gy resulted in a significant reduction of the parotid flow rate by 73% compared with non-irradiated controls (p<0.05). MnSOD but not Cu/ZnSOD partially reduced this effect on the parotid gland by 25% (p<0.05). Both MnSOD and Cu/ZnSOD demonstrated a protective effect against irradiation-induced WBC suppression, by 35% and 25%, respectively (p<0.05). Treatment with SOD did not protect the animals against irradiation induced reduction in oral intake and weight loss, or against submandibular hypofunction. These results suggest that SOD partially protects against head-and neck irradiation-induced injury. Both MnSOD and Cu/ZnSOD partially protect against irradiation-induced WBC loss. The parotid gland is partially protected by MnSOD but not Cu/ZnSOD, while the submandibular gland is not protected by either MnSOD or Cu/ZnSOD. PMID- 10869488 TI - Collagen crimping in the intra-articular disc and articular surfaces of the human temporomandibular joint. AB - The presence of crimping within soft fibrous connective tissues has a considerable role in determining the biomechanical properties of the tissue. However, there is little or no information on crimping of collagen in the human temporomandibular joint. To remedy this situation, the presence and nature of any crimping was studied in sections of human temporomandibular joints from individuals varying in age from between 4.5-63 years, using polarized light microscopy and differential interference contrast microscopy. The presence of crimping was looked for in collagen within the intra-articular disc and the articular surfaces of the mandibular fossa and mandibular condyle. By polarized light, crimping was seen throughout all three tissues at all ages studied. Quantification from micrographs enlarged to x250 showed that the periodicity of the banding (representing half a complete crimp wave) had a mean varying between about 15-20 microm. Crimping was also directly visualized by differential interference contrast microscopy. The presence of such a fundamental feature needs to be considered when explaining the normal function of the temporomandibular joint. PMID- 10869489 TI - The matrix metalloproteinase gelatinase A in human dentine. AB - A dentine protein extraction protocol was modified in order to identify matrix metalloproteinase gelatinolytic activities in the non-mineralized and mineralized phases of human dentine. Dentine proteins from 24 individual permanent molars from patients aged 15-73 years were sequentially extracted, first with guanidinium chloride (G1 extract), then EDTA (E extract), and after this demineralization step, again by guanidinium chloride (G2 extract) to dissociate collagen-associated proteins. Extracts were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis and the gels were processed by Western blotting and zymography to detect gelatinolytic activities. Active and latent forms of gelatinase A were identified in the non-mineralized dentine fraction (G1 extract) of 58% of the teeth. Other gelatinolytic species were also detected by zymography with apparent M(r) of 92, 54 and 30 kDa. Although gelatinase A was detected in the G1 extracts of teeth from all ages, indicating more recent synthesis and remodelling of the predentine, gelatinase A was never detected in any E extract or in the G2 extracts of patients older than 41 years. The presence of the active form of gelatinase A in mineralized human dentine implicates this enzyme in dentine mineralization. PMID- 10869490 TI - Effect of motor tasks on the cortical topography of the human masseter muscle. AB - Non-invasive magnetic stimulation of the brain was earlier used to reveal the corticomotor representation of the human masseter muscle but it is unclear how motor tasks affect this map. An experimental approach incorporating transcranial magnetic stimulation of verified locations on the scalp, surface electromyography, and controlled muscle facilitation was used to disclose the corticomotor output map of the masseter during three isometric tooth-contact tasks, viz., tooth clenching in the intercuspal position, biting on the left molar teeth, and biting on the incisors. Map area was significantly different for all tasks, and map height and volume were also different for biting on the incisor teeth (p<0.05). There was evidence of task-related modulation of corticobulbar activity that appeared to be mainly of corticomotoneuronal origin, although the role of differential, task-associated peripheral afferent input from orofacial receptors could not be discounted. PMID- 10869491 TI - Neuropeptide enzyme hydrolysis in human saliva. AB - The possible hydrolysis of neuropeptides by human saliva was studied using leucine enkephalin as a model. The data obtained indicate that in the presence of saliva this substrate is partially hydrolysed, and that its disappearance corresponds to the appearance of peptides whose composition is consistent with that of the substrate hydrolysis by-products. The formation of these peptides indicates the presence of all three classes of enzymes known to hydrolyse enkephalins in other tissues: aminopeptidases, dipeptidylaminopeptidases and dipeptidylcarboxypeptidases. The activity of these enzymes appears to be altered by the presence of low molecular-weight substances, whose inhibitory activity is apparent on all three classes of enkephalin-degrading enzymes. Substrate degradation was higher in male than female saliva; these differences appear to be caused by lower activity of the enzymes, and higher activity of the low molecular weight inhibitors, measurable in female as compared to male saliva. PMID- 10869492 TI - Elevation of histidine decarboxylase activity in the mandible of mice by Prevotella intermedia lipopolysaccharide and its augmentation by an aminobisphosphonate. AB - Lipopolysaccharide (LPS) produced by Gram-negative bacteria is an important cause of inflammation. Aminobisphosphonates are potent inhibitors of bone resorption but have inflammatory side-effects. Here, the effects of LPS from Prevotella intermedia (a prevalent Gram-negative bacterium both in periodontitis and endodontal infections) and alendronate (an aminobisphosphonate) on the activity of the histamine-forming enzyme, histidine decarboxylase (HDC), were examined in mouse mandible. Intravenous injection of P. intermedia LPS increased HDC activity in the mandible, maximal activity being induced within 3-6 h of the injection. The elevation of HDC activity was dependent on the dose of LPS, 10 microg/kg (0.25 microg/mouse) producing a significant elevation in enzyme activity. Intraperitoneal injection of alendronate (40 micromol/kg) also produced an increase in HDC activity. Moreover, the elevation of HDC activity induced by P. intermedia LPS was markedly augmented in mice given alendronate 3 days before the LPS injection. These results (i) suggest that P. intermedia LPS may stimulate the synthesis of histamine in the mandible and that the newly formed histamine may make at least some contribution to the development of inflammation (apical periodontitis and/or osteomyelitis); (ii) should encourage the clinical testing of antihistaminergic agents against inflammation; and (iii) confirm that care needs to be taken when administering aminobisphosphonates to patients. PMID- 10869493 TI - Crystallographic analysis of demineralized human enamel treated by laser irradiation or remineralization. AB - Crystallographic information on dental hard tissue is helpful in evaluating whether incipient caries that has received preventative treatments is resistant to subsequent attack. The aim here was to analyse crystallographically by means of high-resolution electron microscopy (HREM) demineralized human enamel that had been laser-irradiated or remineralized. Electron-microscopic observation identified a distinct layer at a depth of 100 nm in the demineralized and laser irradiated enamel. The crystallinity in the shallower area was inferior to that in the deeper area. Comparison of the HREM findings from the deeper area with the data provided by the Joint Committee on Powdered Diffraction Standards revealed that the mineral in the demineralized and laser-irradiated enamel was either alpha- or beta-tricalcium phosphate, while that in the remineralized enamel was thought to be tetracalcium diphosphate monoxide. PMID- 10869494 TI - Cariostatic activity of cacao mass extract. AB - Chocolate is suspected to contain some caries-inhibitory substances. The cariostatic activity of cacao mass extract (CM), the main component of chocolate, was examined in vitro and in experimental animals. CM showed no detectable effects on the cellular growth and acid production of mutans streptococci. On the other hand, the cell-surface hydrophobicity of mutans streptococci was significantly reduced by the presence of CM. Furthermore, insoluble glucan synthesis by the glucosyltransferases from either Streptococcus mutans MT8148R or Strep. sobrinus 6715 was inhibited by CM, but not significantly. Hence, the sucrose-dependent cell adherence of mutans streptococci was also depressed by CM. Finally, CM in both a 40% sucrose diet and drinking water resulted in reductions of caries development and plaque accumulation in rats infected with Strep. sobrinus 6715, but not significantly. These results indicate that cacao mass extract possesses some anticariogenic potential, but its anticaries activity is not strong enough to suppress significantly the cariogenic activity of sucrose. PMID- 10869495 TI - Neurochemical changes in brain serotonin neurons in immature and adult offspring prenatally exposed to cocaine. AB - The present study investigates the age-dependent effects of prenatal cocaine exposure on changes in the neurochemical and functional status of brain serotonin neurons. Pregnant rats were administered either saline or (-)cocaine HCl (15 mg/kg, subcutaneously), twice daily from gestational days 13 through 20. Neurochemical changes in frontal cortex, hypothalamus, hippocampus, striatum and midbrain of prepubescent and adult offspring were determined by measuring: (1) the content of serotonin (5-HT) and its major metabolite 5-hydroxyindolacetic acid (5-HIAA), and (2) the ability of the serotonin releasing drug p chloroamphetamine (PCA) to reduce brain serotonin levels. Brain catecholamine content was determined in progeny for comparative purposes. Prior to maturation, prenatal exposure to cocaine did not alter basal levels of brain 5-HT or 5-HIAA in any brain region examined. However, in adult progeny prenatally exposed to cocaine, basal 5-HT content was significantly reduced in the frontal cortex ( 32%) and hippocampus (-40%), suggesting maturation-dependent effects of prenatal cocaine exposure on brain 5-HT neurons. Consistent with the maturational onset of changes in 5-HT, striatal dopamine was significantly reduced (-10%) by prenatal exposure to cocaine only in adult offspring. Reductions in 5-HT in most brain regions, produced by pharmacological challenge with p-chloroamphetamine (PCA), were comparable in prenatal saline versus cocaine offspring. One notable exception was the markedly greater reduction (-40%) in 5-HT in the midbrain of immature offspring prenatally exposed to cocaine, suggesting alterations in midbrain 5-HT neurons prior to maturation. Overall, these data demonstrate prenatal cocaine exposure produces region-specific changes in 5-HT neurons in offspring with some deficits occurring only following maturation. PMID- 10869496 TI - Hypertolerance to morphine in G(z alpha)-deficient mice. AB - Our laboratory has generated a mouse deficient in the alpha (alpha) subunit of the G protein, G(z), (G(z alpha)) gene and we have examined the involvement of G(z alpha) in spinal and supraspinal analgesia and tolerance mechanisms. Spinal analgesia was tested by the response times to heat or cold tail flick times in a water bath at 50 degrees C or -5 degrees C and supraspinal analgesia was tested by the times for paw licking and jumping from a plate at 52 degrees C or 0.5 degrees C. Tolerance to morphine was induced in wild type and G(z alpha) deficient mice over a 5 day period and the behavioral tests were performed daily. The tail flick reaction times to both hot and cold stimuli did not differ between the wild type and G(z alpha)-deficient mice. Analysis of the reaction times from the hot and cold plate tests showed the G(z alpha)-deficient mice developed tolerance to morphine to a greater degree and at a faster rate than wild type mice. Opioid binding assays were performed on synaptic membranes prepared from naive and morphine tolerant wild type and G(z alpha)-deficient brains. No changes in the affinity of morphine for its receptor or in the density of mu and delta opioid receptors were found between the two groups of mice in the naive or morphine tolerant state. This indicates that the absence of G(z alpha) does not affect opioid receptor affinity or receptor up or down regulation. Our results suggest that the presence of G(z alpha) delays the development of morphine tolerance and represents a possible therapeutic target for improving the clinical use of morphine. PMID- 10869497 TI - Male mice lacking the gastrin-releasing peptide receptor (GRP-R) display elevated preference for conspecific odors and increased social investigatory behaviors. AB - Previously, we generated gastrin-releasing peptide receptor null mutant mice (GRP R-deficient mice), and found that these animals displayed increased non aggressive social responses in an ordinary social interaction test using a resident-intruder method. In the present study, we examined in more detail the social behaviors of GRP-R-deficient male mice. In social interaction tests, GRP-R deficient mice showed more social responses, such as sniffing and nosing, relative to wild-type mice, and similar results were obtained whether GRP-R deficient mice served as intruders or residents. In the same way, they showed more contact behaviors toward an anesthetized conspecific, and less locomotor activity than wild-type mice in a social investigation test toward an anesthetized male mouse. Since olfactory systems play important roles in the social behavior of rodents, olfactory preference tests were conducted in order to evaluate the olfactory properties of GRP-R-deficient mice. The results suggest that no differences exist between wild-type mice and GRP-R-deficient mice in the preference between a novel sawdust odor and their own odor, or that of other male mice. However, GRP-R-deficient mice preferred the odor of other male mice to their own, in contrast to wild-type mice. Furthermore, the preferences of GRP-R deficient and wild-type mice were not disrupted by intraperitoneal infusion of diazepam (1.5 mg/kg). These results indicate that neither the motion, nor the behavior of conspecifics, nor reduced anxiety lead to the increased non aggressive social responses and/or social investigatory behaviors in GRP-R deficient mice. Rather, these latter behaviors may be a consequence of altered cognition of conspecific odors in the mutant mice. PMID- 10869498 TI - Developmentally regulated gene expression of Th2 cytokines in the brain. AB - Given the critical role of cytokines in the regulation of an inflammatory response, we investigated whether certain cytokines are expressed in the brains of normal mice during maturation that could contribute to the immune-privileged nature of the CNS or potentially influence an immune-mediated illness such as experimental allergic encephalomyelitis. The gene expression of IFN gamma (Th1 cytokine) and IL-4 (Th2 cytokine) was analyzed in the brain of several strains of mice. IFN gamma was not detectable. However, IL-4 was present in the brains of neonatal mice, but not adult mice. Resident CNS cells are believed to be the source of the IL-4, because mice deficient in T cells (SCID and RAG2-/-) expressed the IL-4 gene in the CNS. Further analysis indicated that the gene expression of the Th2 cytokine transcription factor, GATA-3, correlated with IL-4 and IL-10 expression in the brain. Since GATA-3-deficient mice have an abnormal CNS, brain-derived Th2 cytokines may play an important role in CNS development, as well as potentially contribute to the immune-privileged nature of the brain. PMID- 10869499 TI - Changes in circadian period and morphology of the hypothalamic suprachiasmatic nucleus in fyn kinase-deficient mice. AB - Protein tyrosine phosphorylation is involved in intracellular signal transduction and plays important roles in various physiological events. To understand the role of Fyn, a non-receptor type tyrosine kinase of Src family kinases, in the mechanism of circadian rhythms, we analyzed the circadian locomotor behavior and morphology of the hypothalamic suprachiasmatic nucleus (SCN), a master circadian oscillator in Fyn mutant mice, because Fyn-like immunoreactive substance was observed in the SCN. Under constant dark (DD) condition the Fyn (-/-) mutant mice showed a free-running circadian rhythm, and the period of the circadian rhythm of the locomotor activity was significantly longer than that of the control mice. Fyn (-/-) mutant mice had abnormal distribution of neurons containing vasoactive intestinal polypeptide (VIP)-like immunoreactive substance in the SCN. These findings suggest that Fyn is involved in the mechanism of circadian oscillation and morphological formation of the SCN. The mechanism of the implication of Fyn discussed with the Fyn's roles in neural network formation and cellular signal transduction pathway. PMID- 10869500 TI - Sumatriptan modifies cortical free radical release during cortical spreading depression. A novel antimigraine action for sumatriptan? AB - Increases in concentration of brain NO are proposed to initiate and mediate migraine headache. Triggered by focal depolarisation, spreading depression (SD) represents a suitable mechanism for eliciting widespread release of nitric oxide. The current study examines the effect of sumatriptan, a 5-HT(1B/1D) agonist and effective antimigraine therapy, on free radical release (nitric oxide and superoxide) in SD in the simple and complex cortices of the rat and cat. Following initiation of SD, sumatriptan pretreatment (300 microg kg(-1) i.v., 15 min prior to SD) modulated all phases of nitric oxide release associated with each SD in both cats and rats. As a result, superoxide levels were observed to significantly (ANOVA, post hoc LSD) increase versus vehicle treated animals (saline 1 ml kg(-1) i.v. 15 min prior to SD) during specific phases of each SD depolarisation. Averaged over all SD depolarisations, mean peak SD nitric oxide levels per depolarisation were 0.73+/-0.23 microM (n=29) in cats, and 0.42+/-0.09 microM (n=34) in rats. Sumatriptan significantly (Students t-test, P<0.05, two tailed hypothesis, P<0.05) modulated this increase in cortical nitric oxide concentrations to 0.32+/-0.06 microM (n=25) and 0. 22+/-0.07 microM (n=37) in cats and rats. Sumatriptan appears to decrease the amplitude of nitric oxide release but enhances extracellular superoxide concentrations in both lissencephalic and gyrencephalic cortices during SD. PMID- 10869501 TI - Expression profiles of JunB and c-Fos proteins in the rat circadian system. AB - The immediate-early genes c-Fos and JunB are implicated in light signaling within the suprachiasmatic nucleus (SCN), the mammalian circadian clock. Light induces phase-dependent expression of c-Fos and JunB within the retinorecipient SCN. In the absence of light, rhythmic expression of these genes has been observed in the dorsal region of the SCN with peak expression observed near dawn. The present study examined the pattern of induction of c-Fos and JunB in the SCN and intergeniculate leaflet (IGL) of rats housed in constant conditions, under light dark cycles, or in dark-adapted light-stimulated animals. In contrast with previous studies, no evidence of expression of c-Fos and JunB was observed within the dorsal SCN, regardless of circadian time. Strain differences could account for the absence of rhythmic JunB expression, whereas the use of a monoclonal anti c-Fos antibody in the present study may account for the absence of dorsal SCN c Fos staining. The profile of light-induced c-Fos and JunB expression was consistent with previous observations. In the SCN, light-induced expression of c Fos and JunB was phase dependent, whereas in the IGL light-induced both c-Fos and JunB regardless of circadian time. PMID- 10869502 TI - Riluzole improves measures of oxidative stress following traumatic spinal cord injury. AB - Rats received a contusion injury to the spinal cord followed by treatment with riluzole (a glutamate release inhibitor, 8 mg/kg), methylprednisolone (MP 30 mg/kg) or both. At 4 h following injury, spinal cords were removed and synaptosomes prepared and examined using five measures of oxidative stress. Riluzole treatment was found to improve mitochondrial function, and enhance glutamate and glucose uptake. As expected, MP treatment was found to reduce lipid peroxidation, but also improved glutamate and glucose uptake. Interestingly, the combination treatment was found to be effective in improving all five measures of oxidative stress. The results of this study clearly demonstrate the potential beneficial effects of a combination approach in the treatment of oxidative stress events in traumatic spinal cord injury. PMID- 10869503 TI - Dopamine release from pharmacologically distinct storage pools in rat striatum following stimulation at frequency of neuronal bursting. AB - The increase of dopamine overflow in the rat caudate and nucleus accumbens following repeated stimulation of the median forebrain bundle (MFB) at short intervals and the role of alpha-methyl-p-tyrosine (AMPT) and reserpine sensitive storage pools in evoked dopamine overflow were investigated by in vivo voltammetry. In contrast to stimulation at 30-s intervals and longer, stimulation of the MFB at 5-s intervals led to increased dopamine release following each consecutive stimulation. This effect was much larger in the caudate than in the nucleus accumbens. We consider that the increase in dopamine release is due to translocation of dopamine from a reserpine-sensitive storage pool and this accounts for the increase of the rate of refilling of the readily releasable pool. The shorter the intervals between stimulation and the higher the frequency (with a plateau at 30 Hz), then the greater the fraction of dopamine which originates from the reserpine-sensitive storage pool. Exocytotic release of dopamine from the AMPT-sensitive storage pool does not seem to depend on the intervals between stimulation. We propose that the vesicles in presynaptic dopaminergic terminals near the outer membrane are more sensitive to AMPT. Distantly located vesicles have a relatively higher sensitivity to reserpine. Experiments using repeated stimulation at as low a frequency as 10-20 Hz, revealed that this phenomenon may take place under physiological conditions following bursting of dopamine neurones. PMID- 10869504 TI - 3-bromo-7-nitroindazole, a neuronal nitric oxide synthase inhibitor, impairs maternal aggression and citrulline immunoreactivity in prairie voles. AB - Lactating female rodents are aggressive against intruders when they are rearing and protecting pups. In prairie voles, Microtus ochrogaster, females exhibit a dramatic increase in citrulline immunoreactivity (citrulline-IR) in the paraventricular nucleus (PVN), but not in control regions of the brain, in association with maternal aggression. Citrulline is an indirect indicator of nitric oxide (NO) synthesis and it is possible that NO release in the PVN is an important element in the control of maternal aggression in prairie voles. In this study, we sought to examine the role of NO in maternal aggression by selectively inhibiting neuronal nitric oxide synthase (nNOS) in lactating prairie voles. Intraperitoneal injections of the nNOS inhibitor, 3-bromo-7-nitroindazole (3-Br 7NI) (20 mg/kg), three time per day over 4 days resulted in significant impairment of the expression of maternal aggression in terms of the average time in aggressive encounters, the average number of attacks, and the average latency to first attack. These behavioral deficiencies were observable beginning two days following the onset of drug treatment. The average time spent sniffing the intruder was indistinguishable between the 3-Br-7NI- and oil-treated females. In 3-Br-7NI-treated relative to oil-treated females, the number of citrulline positive cells was reduced by 70% in the PVN and by 50% in the anterior amygdaloid area, a control region of the brain. Taken together, these results indicate that 3-Br-7NI effectively inhibits maternal aggression and NO production in prairie voles and suggest that the central release of NO may play an important role in the production of maternal aggression in prairie voles. PMID- 10869505 TI - Sensitivity to glucocorticoid-mediated fast-feedback regulation of the hypothalamic-pituitary-adrenal axis is dependent upon stressor specific neurocircuitry. AB - Fos-protein immunoreactivity (Fos-IR) was used to identify neurocircuits potentially participating in the regulation of hypothalamic-pituitary-adrenal (HPA) axis sensitivity to glucocorticoid-mediated fast-feedback in rats exposed to the physical stressor, hemorrhage, or the psychological stressor, airpuff startle. Marked regional brain differences in the Fos-IR expression were observed in response to these stressors. Specifically, after hemorrhage, nuclear Fos-IR increased in the nucleus of the solitary tract and other brainstem regions known to regulate hemodynamic processes including the supraoptic nucleus, and the magnocellular division of hypothalamic paraventricular nucleus (PVN). In contrast, after airpuff startle Fos-IR increased in the dorsomedial and lateral hypothalamus as well as in the lateral septum. Thus, activation of brainstem neurocircuits predominated after hemorrhage whereas activation of forebrain neurocircuits predominated after airpuff startle. In other regions, the magnitude of stressor-induced Fos-IR expression varied in a region-specific manner. When stressor exposure was preceded by administration of corticosterone to achieve levels within the physiological range after stressors, HPA axis responses were suppressed in response to the airpuff startle but not to either a small or moderate hemorrhage. IN CONCLUSION: (1) fast-feedback mediated inhibition of HPA axis activity is critically dependent upon stressor modality; (2) this apparent selectivity is reflected by differences in the nature of the neurocircuitry mediating these stressors. It is suggested that determination of the central actions of glucocorticoids in mediating fast-feedback regulation of the HPA axis requires evaluation of the interactions between activated glucocorticoid receptors and intracellular signaling cascades evoked by convergent neuronal input. PMID- 10869506 TI - Food deprivation does not potentiate glucose taste reactivity responses of chronic decerebrate rats. AB - The chronic supracollicular decerebrate (CD) rat fails to increase meal size in response to systemic/metabolic aspects of food deprivation. Here we asked whether or not deprivation increases immediate oral motor responding to taste stimuli (taste reactivity) in CD rats, as it does in neurologically intact controls. The responses of CD rats were evaluated as functions of glucose concentration and deprivation state, with taste reactivity responses recorded myographically during 15-s intraoral infusions and during 45-s post-infusion periods. Five glucose concentrations (0, 3.2, 6. 25, 12.5, 25%) were each presented three times during each test session. The rats were tested when not-deprived (i.e. receiving their full complement of gavage feedings), deprived (23.5 h) of food and water, and deprived of food but not water. The number of oral motor responses emitted increased monotonically with stimulus concentration; during oral infusions the increase was greatest over the lower half of the concentration range, whereas responding increased linearly with concentration in the post-infusion period. This CD response profile resembled that obtained previously with neurologically intact rats tested according to the same protocols. In contrast to results obtained in intact rats, deprivation did not influence the CD's response to glucose at any concentration or for any observation period. Although the caudal brainstem may receive and process information associated with deprivation state, neural interactions between forebrain and brainstem structures appear necessary for the behavioral expression of deprivation effects on meal size or, as we can now conclude, on immediate oral motor responses to taste stimuli. PMID- 10869507 TI - Glucocorticoids exacerbate insult-induced declines in metabolism in selectively vulnerable hippocampal cell fields. AB - Glucocorticoids (GCs), the adrenal steroids released during stress, can compromise the ability of hippocampal neurons to survive necrotic neurological insults. This GC-induced endangerment has energetic facets, in that it can be attenuated with energy supplementation. In the present report, we studied the effects of GCs on the metabolic response of specific hippocampal cell fields to necrotic insults. We used silicon microphysiometry, which allows indirect measurement of metabolism in real time in tissue explants. Aglycemia caused a significant decline in metabolism in dentate gyrus explants, but not in CA1 or CA3 explants. When coupled with our prior report of cyanide disrupting metabolism only in CA1 explants, and the glutamatergic excitotoxin kainic acid disrupting metabolism only in CA3 explants, this demonstrates that microphysiometry can detect the selective regional vulnerability that characterizes the hippocampal response to these necrotic insults. We then examined the effects of GCs on the response to these insults, monitoring explants taken from rats that were adrenalectomized, intact, or treated with corticosterone (the GC of rats) that produced circulating levels equivalent to those of major stressors. Increased exposure to GCs worsened the decline in metabolism in dentate gyrus explants induced by hypoglycemia, and in CA1 explants induced by cyanide (after eliminating the effects of glial release of lactate for the support of neuronal metabolism). Thus, GCs worsen the metabolic consequences of necrotic insults in hippocampal explants. PMID- 10869508 TI - Diurnal metabolism of dopamine in the mouse retina. AB - Dopamine is an important retinal neurotransmitter and neuromodulator that regulates key diurnal cellular and physiological functions. In the present study we carried out a comprehensive analysis of dopamine metabolism during the light phase of the diurnal cycle and evaluated the presence of diurnal and circadian rhythms of dopaminergic activity in the mouse retina. Steady-state levels of dopamine did not change significantly between the dark phase (night) and the light phase (day) of the diurnal cycle, nor did they change between early and late points in the day. Dopamine synthesis and utilization, however, revealed significant alterations between the night and day and between early and late time points in the day. A spike in synthesis and utilization was measured immediately after light onset at the end of the night. Subsequently, dopamine synthesis and utilization partially declined and remained stable throughout the remainder of the day at a level that was significantly higher than that at night. The burst of dopamine synthesis and utilization at the beginning of the day is entirely light evoked and not driven by a circadian clock. Similarly, there was no circadian rhythm in dopamine synthesis and utilization in mice kept in constant darkness. This daily pattern of dopaminergic activity may impact upon a variety of temporally regulated retinal events. Moreover, these data will provide a basis for evaluating the role of dopamine in retinal pathology in mouse models of retinal degeneration where mutations affect light perception. PMID- 10869509 TI - Central muscarinic inhibition of lower urinary tract nociception. AB - Previous studies indicate cholinergic systems suppress somatic nociception. The present studies determined if cholinergic muscarinic systems suppress visceral nociception, specifically, chemical irritation of the lower urinary tract. Bladders of urethane-anesthetized rats were cannulated through the dome for continuous-infusion cystometrogram recordings. EMG electrodes recorded anal sphincter activity. Infusion of 0.5% acetic acid into the bladder to produce irritation increased bladder activity and anal sphincter activity (i.e. activation of a nociceptive vesicoanal reflex). Oxotremorine (a muscarinic agonist) and (-)butylthio[2.2.2] (a mixed muscarinic agonist/antagonist) dose dependently inhibited vesicoanal reflex activity. This inhibition was antagonized by atropine (a centrally active muscarinic antagonist) but not by scopolamine methylbromide (a peripherally restricted muscarinic antagonist). Physostigmine (a centrally active cholinesterase inhibitor) also dose-dependently inhibited vesicoanal reflex activity in an atropine-sensitive manner, while neostigmine (a peripherally restricted cholinesterase inhibitor) did not. Atropine alone (i.e. administered without prior administration of muscarinic agonist or cholinesterase inhibitor) produced robust but transient (15 min) increases in vesicoanal activity and bladder activity under conditions of acetic acid infusion into the bladder. Under conditions of saline infusion into the bladder, atropine had the opposite effect on bladder activity (i.e. inhibition). These studies indicate that an endogenous cholinergic muscarinic system can be activated by lower urinary tract irritation to suppress visceral nociception through central nervous system mechanisms. PMID- 10869510 TI - The involvement of ventral tegmental area cholinergic muscarinic receptors in classically conditioned fear expression as measured with fear-potentiated startle. AB - Accumulating evidence suggests that dopamine (DA) neurons in the ventral tegmental area (VTA) contribute to the complex amygdala-based neurocircuitry that mediates fear-motivated behaviors. Because of acetylcholine's (ACh) role in DA neuronal activation, the involvement of VTA cholinergic muscarinic receptors in Pavlovian conditioned fear responding was evaluated in the present study. Fear potentiated startle was used to assess the effects of intraVTA infused methylscopolamine on conditioned fear performance in laboratory rats. Application of this nonspecific muscarinic receptor antagonist to VTA neurons was observed to inhibit the ability of a conditioned stimulus (CS) previously paired with footshock to enhance the amplitude of the acoustic startle reflex. Doses of methylscopolamine that blocked conditioned fear expression did not alter baseline sensorimotor responding. These results identify ACh neurotransmission in the VTA as a potential excitatory mechanism underlying the fear-arousing properties of threatening environmental stimuli. PMID- 10869511 TI - Transhemispheric cortical reorganization in rat SmI and involvement of central noradrenergic system. AB - Responses of single units in the hindpaw representational area of the left primary somatosensory cortex to electrical stimulation of both hindpaws and the right forepaw were recorded under urethane anaesthesia in three groups of adult male rats: a control group and two groups in which the right hindpaw representational area had been ablated 3-4 weeks previously, immediately after intraperitoneal injection of saline vehicle or DSP4, to destroy cortical noradrenergic terminals arising from the locus coeruleus. The lesion increased the overall number of neurones responding within 500 ms after the stimulation of the contralateral hindpaw (from 64 to 91%), and the proportion exhibiting short latency response increased from 41 to 61%. Interestingly, the proportion of neurones with bilateral representation increased from 3 to 10% after the cortical lesioning. The changes were prevented by injection of DSP4 prior to lesioning and therefore depended on an intact central noradrenergic system. The increase in bilateral representation could not have been due to direct interhemispheric connections between corresponding representational areas because it occurred after lesioning of the homologous area in the contralateral hemisphere. The phenomenon was termed 'transhemispheric reorganization' and because it was somatotopically oriented (e.g. to either hindpaw); its function may be to ensure that when a sensory cortical area is damaged, its basic sensory functions are 'taken over' by the corresponding contralateral area. PMID- 10869512 TI - Effects of oil intake in the conditioned place preference test in mice. AB - We investigated the effects of corn oil intake in the conditioned place preference (CPP) test in mice. Voluntary intake of corn oil in the light box in the CPP test showed place preference, but its peroral administration 60 min before conditioning did not show either place preference or aversion. Acquisition of the place preference by corn oil intake was blocked by i.p. injection of SCH 23390 (0.03 mg/kg) and haloperidol (0.1 mg/kg), but not by that of (+/-) sulpiride (100 mg/kg), (-)-sulpiride (100 mg/kg), and L-741, 626 (1 mg/kg) 15 min before conditioning. These results suggest that stimulation of corn oil in the oral cavity, but not its postingestive effects, have positive reinforcing effects and the stimulation of corn oil is at least partly mediated via dopaminergic systems through the D(1) receptors. Moreover, the present results suggest that the CPP test is useful for the study of preferable stimulation and rewarding effects of food intake. PMID- 10869513 TI - Nefiracetam facilitates hippocampal neurotransmission by a mechanism independent of the piracetam and aniracetam action. AB - Nefiracetam, a nootropic (cognition-enhancing) agent, facilitated neurotransmission in the dentate gyrus of rat hippocampal slices in a dose dependent manner at concentrations ranged from 1 nM to 1 microM, being evident at 60-min washing-out of the drug. The facilitatory action was blocked by the nicotinic acetylcholine (ACh) receptor antagonists, alpha-bungarotoxin and mecamylamine. A similar facilitation was induced by the other nootropic agents, piracetam and aniracetam, but the facilitation was not inhibited by nicotinic ACh receptor antagonists and it did not occlude the potentiation induced by nefiracetam. In the Xenopus oocyte expression systems, nefiracetam potentiated currents through a variety of neuronal nicotinic ACh receptors (alpha 3beta 2, alpha 3beta 4, alpha 4 beta 2, alpha 4 beta 4, and alpha 7) to a different extent. In contrast, neither piracetam nor aniracetam had any potentiating action on alpha 7 receptor currents. While aniracetam delayed the decay time of currents through the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor, GluR1, -2, -3, expressed in oocytes, nefiracetam or piracetam had no effect on the currents. Nefiracetam, thus, appears to facilitate hippocampal neurotransmission by functionally targeting nicotinic ACh receptors, independently of the action of piracetam and aniracetam. PMID- 10869514 TI - Zinc-enriched (ZEN) terminals in mouse spinal cord: immunohistochemistry and autometallography. AB - The general distribution of zinc-enriched (ZEN) terminals in mouse spinal cord was investigated at light microscopic level by means of zinc transporter-3 immunohistochemistry (ZnT3(IHC)) and zinc selenium autometallography (ZnSe(AMG)). Staining for ZnT3(IHC) corresponded closely to the ZnSe(AMG) staining. Both appeared as dense grains of variable sizes and densities in the gray matter with a characteristic segmental laminar pattern. The white matter was unstained but contained rows of stained terminals radiating from the gray matter. In the dorsal horn, laminae I, III and IV were heavily stained, whereas lamina II appeared as the least stained area in the gray matter. Moderate staining was seen in laminae V and VI. In the ventral horn, large ZnT3(IHC) and ZnSe(AMG) grains, known from previous papers to represent ZEN terminals, were observed related in particular to motor neuronal somata and big dendrites. These ZEN terminals in the ventral horn were in general larger than those in the dorsal horn. This is the first description of the pattern of ZEN terminals in mouse spinal cord. PMID- 10869515 TI - Monoaminergic activities of limbic regions are elevated during aggression: influence of sympathetic social signaling. AB - A visual social signal inhibiting aggression is coincident with limiting serotonergic and noradrenergic activity in subiculum, hippocampus, nucleus accumbens, medial amygdala, but not lateral amygdala, septum, and hypothalamus. Darkening of postorbital skin in the lizard Anolis carolinensis is stimulated by sympathetic activation of beta(2)-adrenergic receptors via adrenal catecholamines, and occurs more rapidly in dominant males during social interaction. Eyespot darkening functions as a social signal limiting aggressive interaction. To assess the effect of this social signal on telencephalic activity of monoamines, males were painted postorbitally with green or black paint, and exposed to a mirror. Serotonergic and noradrenergic turnover, as estimated by ratios of catabolite to transmitter, were elevated in the subiculum, hippocampus, nucleus accumbens, and medial amygdala of animals in which the eyespots were masked by green paint. Conversely, dopaminergic activity in these brain regions was lower in males with hidden eyespots (painted green). Hiding the eyespot evoked significantly increased aggressive activity toward the mirror image. Furthermore, changes in monoaminergic turnover were coincident with altered aggressive behavior, suggesting a relationship between them. Changes of monoaminergic activity were not observed in the septum, lateral amygdala, or hypothalamus, when males with eyespots permanently marked (black) were compared with those with eyespots hidden (painted green). Stimulated (serotonergic and noradrenergic) or inhibited (dopaminergic) activity due to social signal and aggression are confined to regions of the brain similarly activated during social stress, and do not constitute a generalized activation of monoaminergic systems. PMID- 10869516 TI - The temporal and spatial expressions of neuropeptide Y induced by seizure in the hippocampal complex of gerbil. AB - Recent studies reported changes in neuropeptide Y (NPY) expression induced by seizures in the experimental epileptic models. However, there have been few reports of the alteration of NPY expression in hippocampal complexes of genetic epilepsy models. In the present study, we performed spatial and temporal analyses of NPY expression in the hippocampal complexes of the seizure-resistant (SR) and seizure-sensitive (SS) gerbils, one of the genetic models. In SR gerbils, most NPY(+) cells were located at the dentate hilus (DH) and the subiculum (SC). In the pre-seizure group of SS gerbils, neurons in the DH and SC were nearly devoid of NPY immunoreactivity. Interestingly, the acute NPY expressions were observed in these areas of the post-seizure group at 30 min, and its immunoreactivity was declined at 12 h after the onset of seizure. These findings suggest that in seizure, the deficiency of NPY in DH and SC may be one of the factors, and that the acute expression of NPY after seizure in these areas may be the compensatory response for reduction of seizure activity in this animal. PMID- 10869517 TI - Receptor-mediated endocytosis of transthyretin by ependymoma cells. AB - Transthyretin (TTR) is involved in the transport of thyroxine (T4) and retinol binding protein (RBP) in cerebrospinal fluid (CSF) and serum. TTR is secreted in the CSF by the epithelial cells of choroid plexus. The binding of [(125)I]TTR to cultured ependymoma cells which form the brain cerebrospinal barrier, was studied to determine whether these cells carry receptor(s) for TTR. TTR was bound by ependymoma cells in a time-dependent manner reaching equilibrium within 2 h. Scatchard analysis was consistent with a single class of high-affinity binding sites with a K(d) of approximately 18 nM. Saturable high-affinity binding of human TTR has previously been described in rat primary hepatocytes and human renal adenocarcinoma, neuroblastoma, hepatoma and astrocytoma cells, and also transformed lung cells. Endocytosis of fluorescent or biotinylated TTR was observed in ependymoma cells in cytoplasmic vesicles but TTR did not colocalize with clathrin in endocytic coated vesicles. Endocytosis of TTR was inhibited by high sucrose concentration (0.45 M). Finally, ligand blotting and chemical linking experiments revealed the presence of a approximately 100 kDa putative TTR receptor on the ependymoma cell membrane. Receptor binding of TTR provides a potential mechanism for the delivery of T4 within the central nervous system. PMID- 10869518 TI - Regional difference of neuronal vulnerability in the murine hippocampus after transient forebrain ischemia. AB - We have investigated the regional difference of neuronal vulnerability within the hippocampus in the C57BL/6 strain mice after forebrain ischemia. Both common carotid arteries of fifty mice were occluded for 12 min and the mouse brain was examined with cresyl violet staining. The CA4 sector was found to be the most vulnerable within the hippocampus. The CA2 and the medial CA1 sector was the 2nd and 3rd most vulnerable regions. However, The lateral part of the CA1 sector, CA3 sector and the dentate gyrus were resistant to ischemic insult. PMID- 10869519 TI - Morphine and methadone have different effects on calcium channel currents in neuroblastoma cells. AB - Using patch-clamp technique, cellular calcium channel currents were studied on nine mouse neuroblastoma N1E115 cells. Both morphine and methadone decreased the T-type calcium currents in dose-dependent fashion. These effects were partially blocked by naloxane. On L-type calcium currents, morphine showed no effect. However, methadone inhibited the L-calcium currents in dose-dependent fashion. This effect was not antagonised by naloxone. Hence, the action of methadone on L calcium channel is probably not associated with mu receptors. PMID- 10869520 TI - Immediate-early gene expression in cerebral cortex following exposure to chronic intermittent hypoxia. AB - Chronic-intermittent hypoxia (CIH) was postulated to evoke c-fos expression in cortical regions that modulate sympathetic discharge. Animals exposed to CIH for 30 days exhibited c-fos labeling in medial prefrontal, cingulate, retrosplenial, and insular cortices. Our findings strongly suggest activation of cortical circuits that adaptively regulate sympathetic and cardiovascular activities. PMID- 10869521 TI - Early and specific expression of monocyte chemoattractant protein-1 in the thalamus induced by cortical injury. AB - For many years it has been known that retrograde degeneration of thalamic neurons occurs following damage to the cerebral cortex, however, the molecular mechanisms which control this process are unknown. Recent studies have demonstrated microglial activation in thalamic nuclei well before the onset of retrograde neuronal cell death. Activated monocytes and microglia synthesize factors detrimental to neuronal survival as well as phagocytose damaged and dying neurons. Our previous studies demonstrated that monocyte chemoattractant protein 1 (MCP-1), a beta chemokine which attracts cells of monocytic origin to sites of injury, is rapidly expressed in the brain following visual cortical lesions. The present study examined the expression of MCP-1 messenger RNA and protein in the thalamus following a visual cortical lesion. Aspiration lesions of visual cortex were made in adult mice. At specific times after lesion, brains were harvested and dissected into specific regions. MCP-1 message as detected using northern analysis was absent in uninjured brain, but was elevated in the ipsilateral thalamus as rapidly as 1 h following the lesion. In situ hybridization localized MCP-1 message to subpial glial cells of the lateral geniculate nucleus (LGN) of the ipsilateral thalamus after injury. ELISA showed that MCP-1 protein levels were significantly elevated in the ipsilateral thalamus at 6 h, peaked at 12 h, and remained above baseline levels for at least 1 week post lesion. In addition, anti-GFAP staining demonstrated activated astrocytes localized to the ipsilateral LGN at 24 and 72 h after injury. The early expression and regional localization of MCP-1 mRNA and protein strongly suggest that MCP-1 is a critical molecule in the regulation of thalamic retrograde neuronal degeneration. PMID- 10869522 TI - Renin-angiotensin-aldosterone system and myocardial fibrosis. PMID- 10869523 TI - A depressing future for cyclosporine? PMID- 10869524 TI - The hibernators heart. Nature's response to arrhythmogenesis? PMID- 10869525 TI - Endothelial nitric oxide synthase and estrogen. PMID- 10869526 TI - Paced ventricular electrogram fractionation and sudden death in hypertrophic cardiomyopathy and other non-coronary heart diseases. PMID- 10869527 TI - Mechanical stress-induced cardiac hypertrophy: mechanisms and signal transduction pathways. AB - Cardiac hypertrophy is a well known response to increased hemodynamic load. Mechanical stress is considered to be the trigger inducing a growth response in the overloaded myocardium. Furthermore, mechanical stress induces the release of growth-promoting factors, such as angiotensin II, endothelin-1, and transforming growth factor-beta, which provide a second line of growth induction. In this review, we will focus on the primary effects of mechanical stress: how mechanical stress may be sensed, and which signal transduction pathways may couple mechanical stress to modulation of gene expression, and to increased protein synthesis. Mechanical stress may be coupled to intracellular signals that are responsible for the hypertrophic response via integrins and the cytoskeleton or via sarcolemmal proteins, such as phospholipases, ion channels and ion exchangers. The signal transduction pathways that may be involved belong to two groups: (1) the mitogen-activated protein kinases (MAPK) pathway; and (2) the janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway. The MAPK pathway can be subdivided into the extracellular-regulated kinase (ERK), the c-Jun N-terminal kinase (JNK), and the 38-kDa MAPK (p38 MAPK) pathway. Alternatively, the stress signal may be directly submitted to the nucleus via the cytoskeleton without the involvement of signal transduction pathways. Finally, by promoting an increase in intracellular Ca2+ concentration stretch may stimulate the calcium/calmodulin-dependent phosphatase calcineurin, a novel hypertrophic signalling pathway. PMID- 10869528 TI - Vascular function in preeclampsia. AB - Preeclampsia is a multisystem disorder peculiar to human pregnancy. It occurs in 4-5% of all pregnancies and remains a leading cause of maternal and neonatal mortality and morbidity. The pathophysiology of this syndrome is not fully understood. Two stages of vascular dysfunction seem to be involved. In the early stage suboptimal development of the placenta and a hemodynamic maladaptation to pregnancy exist. At this stage maternal constitutional factors such as genetic and immunological factors and pre-existing vascular diseases may play a role. Due to this defective placentation a factor is released from the placenta, supposedly under the influence of ischemia. This factor then results in the late vascular dysfunction characterised mainly by a generalised endothelial dysfunction, leading to the clinical syndrome of preeclampsia. This review attempts to unravel the mechanisms that may contribute to preeclampsia-associated changes in vascular function and to indicate the research needed to improve our understanding of this disease. PMID- 10869529 TI - Unstable angina activates myocardial heat shock protein 72, endothelial nitric oxide synthase, and transcription factors NFkappaB and AP-1. AB - OBJECTIVE: Unstable angina may improve the clinical outcome of acute myocardial infarction, but increases the morbidity and mortality of open heart surgery. We hypothetized that unstable angina influences the myocardium, and investigated the expression of the inducible heat shock protein 72 (HSP72), constitutive HSP73, and endothelial nitric oxide synthase (eNOS), and activation of the transcription factors NFkappaB and AP-1 in cardiac tissue. METHODS: Biopsies were taken from the right atrium of 15 patients with unstable and 15 with stable angina undergoing coronary artery bypass grafting. Immunoblotting with monoclonal antibodies against HSP72, HSP73, and eNOS were performed on protein extracts, while nuclear proteins were assessed by electromobility shift assay. RESULTS: When calculating the optical density of the bands, patients with unstable angina had more than twice as much HSP72 and eNOS as stable patients (P<0.005), while HSP73 was similar in both groups. Nuclear translocation of NFkappaB and AP-1 was found in patients with anginal pain shortly before surgery, but not in stable patients or in patients without symptoms for 4 days or more prior to surgery. CONCLUSIONS: HSP72 and eNOS, which may be associated with cardioprotection in ischemic preconditioning, are increased in atrial tissue of patients with unstable angina. Activation of NFkappaB and AP-1, which regulate a battery of inflammatory genes, was found in hearts of unstable patients. NFkappaB activation may induce a myocardial proinflammatory state, possibly making the unstable myocardium more susceptible to the inflammation induced by open heart surgery. PMID- 10869530 TI - Raised blood pressure, not renin-angiotensin systems, causes cardiac fibrosis in TGR m(Ren2)27 rats. AB - OBJECTIVES: Elevated systemic arterial blood pressure is associated with left ventricular hypertrophy and fibrosis. It has been suggested that both circulating and local myocardial renin-angiotensin systems play a role in mediating these responses. Here we describe the natural history of ventricular hypertrophy and fibrosis in the transgenic (mRen2)27 rat--a monogenetic model--which has a high tissue expression of the murine renin transgene, and suffers severe hypertension. We further explored the relative contribution of both hypertensive burden and circulating and tissue renin-angiotensin systems to the fibrotic process. METHODS: The transgenic rats were treated from 28 days old with (1) a hypotensive dose of the ACE inhibitor ramipril which inhibited both tissue and circulating ACE activity, (2) the calcium antagonist amlodipine, or (3) a non-hypotensive dose of ramipril which inhibited about 60% of tissue ACE activity with little effect on circulating ACE. Normotensive Sprague-Dawley rats were used as controls. RESULTS: The transgenics developed left ventricular hypertrophy along with perivascular and interstitial fibrosis which became progressively worse up to 24 weeks of age. Both the high dose of ramipril and amlodipine prevented the hypertrophy and fibrosis, whereas tissue ACE inhibition without lowering blood pressure had no effect, and actually led to a worsening of the fibrosis by 24 weeks. CONCLUSIONS: These results suggest that the development of left ventricular hypertrophy and fibrosis in the transgenic (mRen2)27 rat are regulated by blood pressure and not activity of the renin-angiotensin systems and that progression of fibrosis at 24 weeks involves a mechanism unrelated to local renin-angiotensin activity. PMID- 10869531 TI - Dinitrophenol, cyclosporin A, and trimetazidine modulate preconditioning in the isolated rat heart: support for a mitochondrial role in cardioprotection. AB - BACKGROUND: Recent studies have postulated that mitochondrial ATP-sensitive potassium (mitoK(ATP)) channel activation may modulate mitochondrial function with the resultant induction of a preconditioning phenotype in the heart. We hypothesized that the modulation of mitochondrial homeostasis might confer preconditioning-like cardioprotection. METHODS: We used a model of regional ischemia in Langendorff-perfused isolated rat hearts. Short-term administration of 2,4-dinitrophenol (DNP), an uncoupler of oxidative phosphorylation and cyclosporin A (CSA), an inhibitor of mitochondrial respiration, was used in an attempt to elicit preconditioning-like cardioprotection. The anti-ischemic drug trimetazidine, known to attenuate CSA-induced disruption in mitochondrial function, and the mitoK(ATP) channel blocker 5-hydroxydecanoic acid (5-HD) were used to inhibit the effects of DNP and CSA. Finally, we studied the effect of trimetazidine on adenosine-induced and ischemic preconditioning. Risk zone and infarct size were measured and expressed as a percentage of the risk zone (I/R ratio). RESULTS: DNP, CSA and adenosine pretreatment reduced infarct size (I/R ratio: DNP 9.0+/-2.4%, CSA 12.5+/-1.4%, adenosine 11.9+/-3.6%, all P<0.001 vs. control, 30.2+/-1.3%) similarly to ischemic preconditioning (9.5+/-0.6%, P<0.001 vs. control). Trimetazidine limited the effect of ischemic preconditioning (22.2+/-2.0%, P<0.001 vs. ischemic preconditioning) and completely reversed the DNP, CSA, and the adenosine-mediated reduction in infarct size. 5-HD abolished the effect of ischemic preconditioning and CSA. CONCLUSION: DNP and CSA trigger preconditioning-like cardioprotection in the isolated rat heart. Trimetazidine, a known mitochondrial 'protector', attenuated both drug-induced and ischemic preconditioning. These data support the hypothesis that modulation of mitochondrial homeostasis may be a common downstream cellular event linking different triggers of preconditioning. PMID- 10869532 TI - Ecto-5'-nucleotidase plays a role in the cardioprotective effects of heat shock protein 72 in ischemia-reperfusion injury in rat hearts. AB - OBJECTIVE: Heat shock protein 72 (HSP72) is involved in the myocardial self preservation system under several conditions such as ischemia-reperfusion injury or late preconditioning. However, its mechanism is not fully understood. Ecto-5' nucleotidase is a key enzyme for synthesizing adenosine and plays an important role in ischemic preconditioning. In this study, we tested the hypothesis that ecto-5'-nucleotidase plays a role in the cardioprotection of HSP72. METHODS: Rat hearts (H group, n=6) were transfected with HSP72 gene by an intracoronary infusion of hemagglutinating virus of Japan (HVJ)-liposome complex. Control hearts (C group, n=6) were transfected with the beta-galactosidase gene. Following 30 min of normothermic ischemia, grafts were reperfused using Langendorff apparatus. RESULTS: The activity of ecto-5'-nucleotidase was significantly higher in H group than C group both before and after ischemia reperfusion (H vs. C; 0.51+/-0.05 vs. 0.29+/-0.06, and 1.41+/-0.15 vs. 0.85+/ 0.11 nmol/mg protein/min, P<0.05). H group also showed significant better functional recoveries than C group (P<0.05), as well as less creatine phosphokinase leakage (4.4+/-2.8 vs. 14.2+/-3.4 mU/min, P<0.05) and higher adenosine release (247.5+/-35.1 vs. 54.3+/-1.7 pmol/min, P<0.05). Administration of alpha,beta-methylene adenosine diphosphate (AMP-CP), an inhibitor of ecto-5' nucleotidase, significantly diminished the tolerance to ischemia-reperfusion injury in H group (P<0.05). CONCLUSION: These results demonstrated that ecto-5' nucleotidase activated by an overexpression of HSP72 attenuated ischemia reperfusion injury in the rat myocardium. They suggest that ecto-5'-nucleotidase plays a role in the cardioprotective effects of HSP72 in rat hearts. PMID- 10869533 TI - Activation of sulphonylurea-sensitive channels and the NO-cGMP pathway decreases the heart rate response to sympathetic nerve stimulation. AB - OBJECTIVES: Activation of ATP sensitive K+ channels (K(ATP)) and the NO-cGMP pathway have both been implicated in reducing norepinephrine (NE) release from cardiac sympathetic nerves during stimulation. Our aim was to test whether these pathways could interact and modulate cardiac excitability during sympathetic nerve stimulation (SNS). METHODS: The effect of inhibitors and activators of K(ATP) channels and the NO-cGMP pathway on the heart rate (HR) response to cardiac SNS in the isolated guinea pig (Cavia porcellus) double atrial/right stellate ganglion preparation was studied (n=48). RESULTS: The K(ATP) channel activator, diazoxide (100 microM, n=6) or hypoxia (0% O2/5% CO2, n=6) significantly attenuated the HR response to 3 Hz SNS by -10+/-4% and -27+/-6% respectively; an effect that was reversed by the K(ATP) channel inhibitor, glibenclamide (30 microM). Glibenclamide (n=6) on its own enhanced the HR response to SNS by 20+/-8%. Bath applied NE (0.1-0.7 microM, n=6) did not affect the HR response to diazoxide, although an increased response to glibenclamide was observed at 0.3 and 0.5 microM NE. In the presence of 8-Br-cGMP (0.5 mM, n=7), diazoxide further decreased the HR response SNS (19+/-3%). The NO synthase inhibitor, N-omega-nitro-L-arginine (100 microM) significantly increased the HR response (13+/-3%) to SNS in the presence of diazoxide (100 microM, n=6). This effect was reversed with excess (1 mM) L-arginine. Conversely, the NO donor, sodium nitroprusside (SNP, 20-100 microM) significantly attenuated the HR response to SNS. The addition of glibenclamide (30 microM, n=10) could still enhance the HR response (42+/-15%) to SNS. Similar results were seen with the cyclic GMP analogue, 8-Br-cGMP (0.5 mM, n=12). CONCLUSIONS: Our results indicate that NO and sulphonlyurea-sensitive channels act in a complementary fashion, but appear to be independent of each other in the regulation of HR during cardiac SNS activation. PMID- 10869534 TI - Peripheral pre-synaptic pathway reduces the heart rate response to sympathetic activation following exercise training: role of NO. AB - OBJECTIVES: We tested the hypothesis that the attenuated heart rate (HR) response to sympathetic activation following swim training in the guinea pig (Cavia porcellus) results from a peripheral modulation of pacemaking by nitric oxide (NO). METHODS: Nitric oxide synthase (NOS) inhibition on the increase in heart rate with sympathetic nerve stimulation (SNS) was investigated in the isolated guinea pig double atrial/right stellate ganglion preparation from exercise trained (6-weeks swimming, n=20) and sedentary animals (n=20). Western blot analysis for neuronal nitric oxide synthase (nNOS) was performed on the stellate ganglion from both groups. RESULTS: Relative to the control group, the exercise group demonstrated typical exercise adaptations of increased ventricular weight/body weight ratio, enhanced skeletal muscle citrate synthase activity and higher concentrations of [3H]ouabain binding sites in both skeletal and cardiac tissue (P<0.05). The increase in heart rate (bpm) with SNS significantly decreased in the exercise group (n=16) compared to the sedentary group (n=16) from 30+/-5 to 17+/-3 bpm at 1 Hz; 67+/-7 to 47+/-4 bpm at 3 Hz; 85+/-9 to 63+/-4 bpm at 5 Hz and 101+/-9 to 78+/-5 bpm at 7 Hz stimulation (P<0.05). The increase in heart rate with cumulative doses (0.1-10 microM) or a single dose (0.1 microM) of bath-applied norepinephrine expressed as the effective doses at which the HR response was 50% of the maximum response (EC50) were similar in both exercise (EC50 -6.08+/-0.16 M, n=8) and sedentary groups (EC50 -6.18+/-0.07 M, n=7). Trained animals had significantly more nNOS protein in left stellate ganglion compared to the sedentary group. In the exercise group, the non-isoform selective NOS inhibitor, N-omega nitro-L-arginine (L-NA, 100 microM) caused a small but significant increase in the heart rate response to SNS. However, the positive chronotropic response to sympathetic nerve stimulation remained significantly attenuated in the exercise group compared to the sedentary group during NOS inhibition (P<0.05). CONCLUSIONS: Our results indicate that there is a significant peripheral pre-synaptic component reducing the HR response to sympathetic activation following training, although NO does not play a dominant role in this response. PMID- 10869535 TI - Influence of cyclosporine A on contractile function, calcium handling, and energetics in isolated human and rabbit myocardium. AB - OBJECTIVE: The immunosuppressive drug Cyclosporine A (CsA) is a key substance in pharmacological therapy following solid organ transplantation and has been suggested to prevent cardiac hypertrophy. We investigated the direct effects of CsA on myocardial function, because these are largely unknown. METHODS: In multicellular cardiac muscle preparations from end-stage failing and non-failing human hearts as well as from non-failing rabbit hearts we investigated the effects of CsA on contractile performance, sarcoplasmic reticulum (SR) Ca2+-load, cytosolic calcium transients, calcium sensitivity of the myofilaments, and myocardial oxygen consumption. RESULTS: In failing human muscle preparations there was a concentration dependent decrease in contractile force; the maximal effect amounted to 55.6+/-6.4% of control while EC50 was reached at 1.0+/-0.3 nM (n=6). These concentrations are at and even below the therapeutic plasma levels. CsA decreased the aequorin light signal in human failing trabeculae to 71.5+/ 5.9% (n=5), indicating decreased calcium transients. Estimation of the SR calcium load via measurement of rapid cooling contractures revealed a decrease to 84.4+/ 6.5% in failing human preparations (n=6). Measurements of both decreased SR calcium load and force development in presence of CsA were also observed in four non-failing human muscle preparations. In rabbit muscle preparations (n=8), developed force decreased to 50.2+/-7.7% (n=8, EC50: 1.9+/-0.4 nM) and rapid cooling contractures to 74.0+/-7.4% of control at 100 nmol/l CsA. No direct effects were observed on myofilament calcium sensitivity nor on maximal force development of permeabilized preparations from the rabbit (n=7). Oxygen consumption measurements showed that CsA decreased the economy of contraction to 76.4+/-7.9% in rabbit preparations (n=8). CONCLUSIONS: CsA causes a direct cardio depressive effect at clinically relevant concentrations, most likely due to altered handling of Ca2+ by the SR. PMID- 10869536 TI - Increased connexin43 gap junction protein in hamster cardiomyocytes during cold acclimatization and hibernation. AB - OBJECTIVE: The physiology of hibernation is characterized by dramatic reductions of heart rate, respiration, metabolism, blood pressure and body temperature and by resistance to ventricular fibrillation. Gap junctions in the heart provide low resistance pathways, facilitating electrical and metabolic coupling between cardiac muscle cells for coordinated action of the heart and tissue homeostasis. The conductance of these junctions, and therefore their function, is likely to be affected by the physiological changes that take place during hibernation. Our objective was to quantitate gap junction protein levels in cold acclimatization, hibernation and arousal. METHODS: We have used specific antibodies to connexins 43 and 40, in combination with confocal microscopy, to quantitatively analyze the expression of connexin protein in hamster (Mesocricetus auratus) left ventricles in four animal groups: normal controls at euthermy, cold controls (cold-exposed animals that did not undergo hibernation), hibernating animals and animals aroused from hibernation for 2 h. RESULTS: Connexin40 immunostaining was not detected in ventricular cardiomyocytes in any animal group but connexin43 was found in all groups. Connexin43 expression was significantly enhanced in hibernation and cold control ventricular cardiomyocytes. Total plaque area, numerical density and plaque size were higher in the cold controls and hibernating hamsters compared to normal controls and animals aroused from hibernation. CONCLUSION: It is possible that the increased size and number of connexin43 gap junction plaques in the cold controls may represent a compensatory response in order to maintain sufficient gap junction communication during physiological conditions that would reduce conductance. These changes may represent a mechanism by which the hamster avoids ventricular fibrillation during hibernation and arousal. PMID- 10869537 TI - Calcium handling and cell contraction in rat cardiomyocytes depleted of intracellular magnesium. AB - OBJECTIVES: Depressed levels of cardiac Mg have been found in patients with ischaemic heart disease or heart failure, but it is not known whether low intracellular free [Mg2+] ([Mg2+]i) is a causal factor in such myocardial dysfunction. The aims for the present study were to develop a method of lowering [Mg2+]i in myocytes isolated from normal rat hearts, and so to determine whether a low [Mg2+]i itself would cause abnormalities of intracellular Ca2+ ([Ca2+]i) homeostasis or myocyte contractile function in absence of any cardiac disease. METHODS: Rat ventricular myocytes were loaded with mag-indo-1/AM or indo-1/AM for determination of total [Mg2+]i and [Ca2+]i, respectively. Mitochondrial [Ca2+] was determined by selective loading of indo-1/AM into the mitochondria. Cell contraction was measured using an edge-tracking device. Myocytes were depleted of [Mg2+]i by incubation in absence of external Mg2+. This resulted in a decrease in [Mg2+]i from about 1.3 to 0.3 mM. In subsequent experiments, 1.2 mM MgCl2 was again present in the superfusate. RESULTS: Under basal conditions (low rate of stimulation, 0.2 Hz, and 1 mM external [Ca2+]), the Mg-depleted cells showed very similar changes in [Ca2+] to control cells, despite an increase in the amplitude of cell contraction. But in presence of high external [Ca2+] (4 mM) and 5 Hz stimulation rate, the Mg-depleted cells showed defects in systolic Ca2+ handling and in cell contraction; in particular, they were unable to increase systolic [Ca2+] in response to the stimulus, unlike control cells. Despite these alterations in total [Ca2+]i, mitochondrial Ca2+ uptake was unchanged in the Mg depleted cells. CONCLUSIONS: A low [Mg2+]i can itself cause significant cardiomyocyte dysfunction in absence of any contributing disease state. PMID- 10869538 TI - Injury current modulates afterdepolarizations in single human ventricular cells. AB - OBJECTIVE: Injury current (I(injury)) and afterdepolarizations are thought to play an important role in arrhythmias that occur during acute ischemia. However, little is known about the effects of I(injury) on afterdepolarizations. The present study was designed to study the effect of I(injury) on afterdepolarizations and action potentials in single human ventricular cells. METHODS: The patch-clamp technique was used to record action potentials and to apply I(injury) to human ventricular cells. In these cells, early and delayed afterdepolarizations (EADs and DADs) were induced by 1 microM norepinephrine. I(injury) was simulated by coupling cells via a variable coupling resistance to a passive resistance circuit with a potential of 0, -20, or -40 mV, mimicking a depolarized ischemic region. RESULTS: At all potentials, I(injury) induced depolarization of the resting membrane potential and action potential shortening. Flowing from 0 mV, I(injury) induced EADs by itself and aggravated the EADs and DADs that were induced by norepinephrine. Flowing from -40 mV, I(injury) abolished the noradrenaline-induced EADs and DADs. CONCLUSIONS: Our results demonstrate that I(injury) may either prevent or promote the occurrence of afterdepolarizations in human ventricle. The latter holds if conduction is slowed to such an extent that it permits flow of current from depolarized ischemic cells at plateau level to cells in phase 3 or phase 4. PMID- 10869539 TI - Beta(2)-adrenergic receptor overexpression driven by alpha-MHC promoter is downregulated in hypertrophied and failing myocardium. AB - OBJECTIVE: The alpha-myosin heavy chain (alpha-MHC) promoter is frequently used to direct cardiac specific transgene expression. We studied whether transgene expression controlled by this promoter was altered under conditions of cardiac hypertrophy and failure. METHODS: Transgenic (TG) mice overexpressing human beta(2)-adrenergic receptors (beta(2)AR) and wild type (WT) controls were subjected to thoracic aortic constriction (TAC) or sham operation and studied at 1, 3 and 8 weeks after surgery. RESULTS: Sham operated TG mice had higher heart rates and left ventricular (LV) contractility than WT (all P<0.01), demonstrating enhanced betaAR activation. TAC at 1, 3 and 8 weeks produced progressive LV hypertrophy which was similar between WT and TG mice. Evidence of heart failure was more marked in TG mice with a greater increase in weights of the right ventricle and lungs and a higher prevalence of atrial thrombus (P<0.05 in each case). In hypertrophied TG hearts, endogenous alpha-MHC mRNA transcripts in LV were maintained at 1 and 3 weeks, but were reduced by approximately 40% relative to the sham-operated group at 8 weeks after TAC. Transgene expression, measured as human beta(2)AR mRNA, was reduced by 45% at 1 and 3 weeks and by 70% at 8 weeks after TAC. beta(2)AR binding sites were reduced by 35, 47 and 65%, respectively, at 1, 3 and 8 weeks. CONCLUSION: Cardiac hypertrophy and failure cause downregulation of the endogenous alpha-MHC as well as cardiac specific overexpression of the transgene directed by an alpha-MHC promoter. PMID- 10869540 TI - Endothelin-receptor blockade improves endothelial vasomotor dysfunction in heart failure. AB - OBJECTIVES: To elucidate the effect of selective endothelin ET(A)- and mixed ET(A/B)-receptor antagonists on endothelial vasomotor dysfunction in rats with heart failure after myocardial infarction (MI). METHODS: Vasoreactivity and superoxide anion formation were determined in aortic rings from Wistar rats 12 weeks after extensive MI (>46% of left ventricle) compared to sham-operated animals. Rats were either treated with the selective ET(A)-receptor antagonist LU 135252 (30 mg/kg/day), the mixed ET(A/B)-receptor antagonist Bosentan (100 mg/kg/day) or placebo. RESULTS: In MI rats, the concentration-response curve of the endothelium-dependent, nitric oxide-mediated relaxation induced by acetylcholine was significantly shifted to the right and the maximum relaxation was attenuated. Long-term treatment with both ET antagonists significantly improved acetylcholine-induced relaxation in MI rats. LU 135252 was more effective than Bosentan. Endothelium-independent relaxations induced by sodium nitroprusside as well as endothelin- and phenylephrine-induced contractions were similar in all groups of rats. Plasma renin activity and aortic superoxide formation, which were enhanced in rats with heart failure, were normalized by LU 135252, but not by Bosentan treatment. CONCLUSIONS: Long-term treatment with ET receptor antagonists improves endothelial vasomotor dysfunction in rats with chronic MI. This mechanism may essentially contribute to the beneficial effects of ET receptor blockade in heart failure. PMID- 10869541 TI - Vascular dysfunction and myocardial contractility in the JCR:LA-corpulent rat. AB - OBJECTIVE: The JCR:LA-corpulent rat is a unique animal model of human vascular disease that exhibits a profound insulin resistance, vasculopathy, and cardiovascular dysfunction. We tested the hypothesis that the defects affect endothelial and smooth muscle function of the coronary microvasculature as well as cardiac contractility. Coronary, myocardial and aortic function were assessed in obese (homozygous for the cp gene, cp/cp) and lean (heterozygous or homozygous normal, +/?) littermates aged 7 and 18 weeks. METHODS: Coronary endothelial relaxation was examined in isolated perfused hearts by determining the effect of bradykinin (0. 1-1000 nmol l(-1)) on coronary perfusion pressure (CPP), myocardial mechanical function was evaluated in terms of left-ventricular developed pressure (LVDevP), and aortic relaxation with the endothelium-dependent agonist, A 23187 (1-1000 nmol l(-1)). RESULTS: In rats aged 7 weeks, bradykinin reduced CPP from 133+/-1 mmHg to 43+/-1 mmHg (-67%) in lean rats, but only to 64+/-3 mmHg (-52%) in corpulent rats (n=6, P<0.05). Similar differences were found in rats aged 18 weeks (n=8). Inhibition of NO synthase with N(G)-nitro-L arginine (L-NNA; 0.2 mmol l(-1)) impaired, and tetrahydrobiopterin (0.1 mmol l( 1)), a NO synthase cofactor, restored relaxation in cp/cp rats. Spermine/NO equally reduced CPP in both groups (-58%). Mechanical function was similar in lean and corpulent rats, aortic endothelial relaxation was attenuated by approximately 30% and aortic smooth muscle function was normal (7 weeks) or improved (18 weeks) in the cp/cp genotype. CONCLUSION: These results suggest that (i) there is a specific impairment of NO-mediated relaxation of the coronary resistance vessels in the JCR:LA-corpulent rat that is not associated with impaired baseline myocardial contractility, and (ii) exogenous tetrahydrobiopterin reversed the relaxation defects that are part of the vascular complications typical for the insulin resistance syndrome. PMID- 10869542 TI - Increase in plasma levels of secretory type II phospholipase A(2) in patients with coronary spastic angina. AB - OBJECTIVE: Plasma levels of sPLA(2) were increased in various chronic inflammatory diseases including coronary artery disease. Lipid products mediated through PLA(2) have been shown to induce impairment of endothelium-dependent dilation, contraction of smooth muscle and proliferation of smooth muscle cells, all of which might lead to coronary spasm. Thus, this study investigated whether plasma levels of secretory non-pancreatic type II phospholipase A(2) (sPLA(2)) may be increased in patients with coronary spastic angina, considering the possible link of sPLA(2) with pathogenesis of coronary artery spasm. METHODS: Plasma levels of sPLA(2) in peripheral circulation, in coronary sinus and in aortic root were measured in 57 patients with coronary spastic angina, 46 patients with stable effort angina and 53 control patients by radioimmunoassay. RESULTS: The peripheral plasma levels of sPLA(2) were increased in patients with coronary spastic angina compared with control patients. In multivariate statistical analysis, the increase in sPLA(2) levels was a significant risk for the presence of coronary spasm independent of other risk factors including C reactive protein levels. The coronary sinus-arterial difference of plasma sPLA(2) levels, reflecting sPLA(2) released into the coronary circulation, was increased during coronary spasm induced by the intracoronary infusion of acetylcholine in patients with coronary spastic angina, but it remained unchanged both during the acetylcholine infusion and during myocardial ischemia provoked by rapid atrial pacing in patients with stable effort angina and in control patients. CONCLUSION: The increase in peripheral plasma levels of sPLA(2) is a significant risk factor for the presence of coronary spasm and it may possibly reflect inflammatory activity in spasm coronary arteries. PMID- 10869543 TI - Endothelin receptor antagonism in patients with chronic heart failure. AB - OBJECTIVE: The relative importance of ETA and ETB receptors in mediating the constrictor effects of endogenous endothelin-1 in patients with chronic heart failure is not known. The primary purpose of this study was to compare the acute effects of selective ETA and ETB receptor antagonists in vivo in healthy subjects and patients with chronic heart failure. Our secondary aim was to examine more closely the effect of chronic heart failure on endothelin biosynthesis. METHODS: We studied the effects of BQ-123 (a selective ETA antagonist) and BQ-788 (a selective ETB antagonist) in ten healthy subjects and ten patients with chronic heart failure. Locally active doses of each antagonist were infused into the non dominant brachial artery for 90 min on separate days at least 1 week apart. Changes in forearm blood flow were measured by venous occlusion plethysmography. Venous blood samples were obtained prior to antagonist infusion for assay of total endothelin, big endothelin-1 and C-terminal fragment immunoreactivity. RESULTS: BQ-123 (100 nmol/min) increased blood flow by 54+/-10% (P<0.001) and 30+/-5% (P<0.001) in controls and heart failure patients, respectively. BQ-788 (1 nmol/min) reduced blood flow by 15+/-5% (P=0. 036) and 9+/-4% (P=0.001) in controls and heart failure patients, respectively. Total endothelin immunoreactivity was non significantly greater in heart failure patients than controls (6. 8+/-1.4 vs. 4.6+/-0.5 pM; P=0.13). Big endothelin-1 (2.6+/-0.4 vs. 1. 7+/-0.1 pM; P=0.04) and C-terminal fragment immunoreactivity (2. 1+/-0.3 vs. 0.6+/-0.1 pM; P<0.0001) were each significantly greater in heart failure patients than controls. CONCLUSIONS: Selective ETA receptor antagonism caused vasodilatation in the peripheral circulation of healthy subjects and patients with chronic heart failure while selective ETB receptor antagonism caused vasoconstriction in each group. ETB receptor antagonism may therefore cause potentially deleterious vasoconstriction in chronic heart failure. Chronic heart failure is associated with a significant increase in plasma big endothelin-1 and C-terminal fragment immunoreactivity. PMID- 10869544 TI - Local application of advanced glycation end products and intimal hyperplasia in the rabbit collared carotid artery. AB - OBJECTIVE: Accumulation of advanced glycation end products (AGEs) in the vessel wall has been implicated in atherogenesis. The aim of our study was to examine the effects of local application of glycated bovine serum albumin (AGE-BSA) on collar-induced intimal hyperplasia in a diabetes-free setting. METHODS: Intimal thickening was induced by placing a collar around the carotid artery of rabbits. Via a catheter attached to an osmotic minipump, control BSA or AGE-BSA was administered locally to the vessel wall in a dose of 1.5 or 15 microg h(-1) during 14 days. Vessels receiving phosphate buffered saline (PBS, 5 microl h(-1)) were used as controls. RESULTS: Infusion of AGE-BSA 15 microg h(-1) significantly enhanced intimal thickening as compared to control BSA or PBS. Positive remodelling, measured as an increase in the area comprised by the external elastic lamina and preservation of lumen size, was only significant after treatment with the higher dose of AGE-BSA. In all other groups, intimal thickening was accompanied by a decrease of the lumen without outward displacement. Infusion of control BSA or AGE-BSA changed the cell composition of the neointima, with a significant enhancement in the number of T-lymphocytes and macrophages and a reduction in the percentage of intimal area occupied by smooth muscle cells. These effects were however similar for control BSA as well as AGE BSA. CONCLUSIONS: It is concluded that infusion of control BSA or AGE-BSA may aggravate collar-induced intimal thickening by evoking an inflammatory response. This supports the concept that inflammation contributes to atherogenesis. Further, the significant enhancement in intimal hyperplasia by AGE-BSA suggests that glycated proteins provide an additional stimulus for the development of atherosclerotic lesions. PMID- 10869546 TI - Endothelial nitric oxide synthase activity in aorta of normocholesterolemic rabbits: regional variation and the effect of estrogen. AB - OBJECTIVE: Several animal studies suggest that nitric oxide (NO) produced by the endothelium attenuates arterial cholesterol accumulation. In the present study we have asked the following questions: (1) is the regional variation in aortic cholesterol accumulation in hypercholesterolemic rabbits preceded by a regional variation in endothelial NO synthase (eNOS) activity in normocholesterolemic rabbits, and (2) is the antiatherogenic effect of estrogen in hypercholesterolemic rabbits preceded by a higher eNOS activity in normocholesterolemic rabbits. METHODS: The eNOS activity was determined by conversion of 14C-L-arginine to 14C-L-citrulline in freshly isolated endothelial cells of aorta in normocholesterolemic rabbits. In the regional variation study, 16 male and eight female rabbits were used. In the estrogen study, ovariectomized female rabbits were subcutaneously injected three times weekly with either 17beta estradiol (n=7) or vehicle (n=7) for 18 weeks. RESULTS: In the regional variation study, the atherosclerosis prone aortic arch showed a significant lower eNOS activity than the more resistant abdominal aorta in both male (P<0.0001) and female (P<0.05) rabbits. In the estrogen study, the eNOS activity in the aortic arch and upper thoracic aorta was significantly higher in the estrogen than in the vehicle rabbits (P<0.05). In the lower thoracic aorta, however, the eNOS activity was the same. CONCLUSION: The findings suggest that a high NO production in the luminal endothelium of the arterial wall precedes a low cholesterol accumulation during a subsequent period of hypercholesterolemia. PMID- 10869547 TI - The active-site residue Cys-29 is responsible for the neutral-pH inactivation and the refolding barrier of human cathepsin B. AB - Human cathepsin B, the most abundant lysosomal cysteine protease, has been implicated in a variety of important physiological and pathological processes. It has been known for a long time that like other lysosomal cysteine proteases, cathepsin B becomes inactivated and undergoes irreversible denaturation at neutral or alkaline pH. However, the mechanism of this denaturation process remains mostly unknown up to this day. In the present work, nuclear magnetic resonance spectroscopy was used to characterize the molecular origin of the neutral-pH inactivation and the refolding barrier of human cathepsin B. Two forms of human cathepsin B, the native form with Cys-29 at the active site and a mutant with Cys-29 replaced by Ala, were shown to have well-folded structures at the active and slightly acidic condition of pH 5. Surprisingly, while the native cathepsin B irreversibly unfolds at pH 7.5, the C29A mutant was found to maintain a stable three-dimensional structure at neutral pH conditions. In addition, replacement of Cys-29 by Ala renders the process of the urea denaturation of human cathepsin B completely reversible, in contrast to the opposite behavior of the wild-type cathepsin B. These results are very surprising in that replacement of one single residue, the active-site Cys-29, can eliminate the neutral-pH denaturation and the refolding barrier. We speculate that this finding may have important implications in understanding the process of pH-triggered inactivation commonly observed for most lysosomal cysteine proteases. PMID- 10869545 TI - Male gender predisposes to development of endotoxic shock in the rat. AB - OBJECTIVE: After intravenous (i.v.) injection of lipopolysaccharide (LPS) macrophages release nitric oxide (NO) due to the expression of the inducible NO synthase (iNOS). After LPS NO is abundantly produced also in the cardiovascular system and may contribute to the development of hypotension and shock. Since the immune response, the synthesis of NO and the regulation of blood pressure (BP) differ between males and females, in the present study the effect of LPS on BP, renal function, the plasma and urinary concentration of the metabolites of NO as well as the splenic and aortic expression of the iNOS gene were compared between male and female rats. METHODS: BP and renal function were measured in anesthetized rats following the i.v. injection of LPS (E. coli, 4 mg/kg). The NO2 and NO3- (metabolites of NO=NOx) concentration was measured by the Griess reaction. The iNOS gene expression was studied by RT-PCR. RESULTS: Four hours after LPS, BP of males (n=9) was reduced by 63+/-12 mmHg versus 10+/-4 in females (n=7, P<0.005). Aminoguanidine, a selective inhibitor of iNOS, prevented the reduction of BP in males. The plasma concentration of NOx (P(NOx)), microM) was lower in hypotensive males (128+/-20) than in normotensive females (235+/-29, P<0.005). Males also exhibited lower urinary NOx excretion (U(NOx)V) after LPS (P<0.001 vs. females). Prior castration of males provided protection against hypotension (fall of BP: -4+/-4 mmHg, n=6, P<0.02 versus males) and resulted in higher P(NOx) as well as U(NOx)V (both P<0.001 versus males and not different from females). Prior ovariectomy (n=5) had no influence on the hemodynamic and NOx response to LPS. Male rats displayed enhanced aortic iNOS/beta-actin ratio relative to females after LPS (n=3 in each group, P<0.05). CONCLUSIONS: (1) Male gender may sensitize to LPS-induced shock and (2) sensitivity of males to endotoxin is associated with an attenuated, not exaggerated total rate of NO synthesis. PMID- 10869548 TI - Reduced hepatic fatty acid oxidation in fasting PPARalpha null mice is due to impaired mitochondrial hydroxymethylglutaryl-CoA synthase gene expression. AB - Glucose and fatty acid metabolism (oxidation versus esterification) has been measured in hepatocytes isolated from 24 h starved peroxisome proliferator activated receptor-alpha (PPARalpha) null and wild-type mice. In PPARalpha null mice, the development of hypoglycemia during starvation was due to a reduced capacity for hepatic gluconeogenesis secondary to a 70% lower rate of fatty acid oxidation. This was not due to inappropriate expression of the hepatic CPT I gene, which was similar in both genotypes, but to impaired mitochondrial hydroxymethylglutaryl-CoA synthase gene expression in the PPARalpha null mouse liver. We also demonstrate that hepatic steatosis of fasting PPARalpha null mice was not due to enhanced triglyceride synthesis. PMID- 10869549 TI - Newly identified repeat sequences, derived from human chromosome 21qter, are also localized in the subtelomeric region of particular chromosomes and 2q13, and are conserved in the chimpanzee genome. AB - Subtelomeric regions have been a target of structural and functional studies of human chromosomes. Markers having a defined structure are especially useful to such studies. Here, we report 93 bp tandem repeat sequences found in the subtelomeric region of human chromosome 21q. They were also detected in the telomeric region of several other chromosomes. Interestingly, the repeat was also found in the 2q13 region which is known to be a position of chromosomal fusion, a major difference between the human and chimpanzee karyotypes. To the best of our knowledge, this repetitive sequence is a new member of human subtelomeric interspersed repeats. PMID- 10869550 TI - Identification of an optimal Ncx binding sequence required for transcriptional activation. AB - The Ncx gene encodes a homeobox containing transcription factor that belongs to the Hox11 gene family. We determined specific Ncx protein binding consensus DNA sequences. Optimal Ncx binding sequences were 5'-CGGTAATTGG-3' (TAAT core) and 5' CGGTAAGTGG-3' (TAAG core), which coincided with the Hox11 binding sequence. Both Ncx and Hox11 could bind to the TAAT and the TAAG core oligonucleotide in vitro. However, they could efficiently transactivate the reporter plasmid linked to the TAAT core sequence but not to the TAAG core sequence. Thus, Ncx and Hox11 act as transcriptional activators via their target sequence, 5'-CGGTAATTGG-3'. PMID- 10869551 TI - Sorting of the vasopressin prohormone into the regulated secretory pathway. AB - The sorting of soluble proteins into the regulated secretory pathway (RSP) involves aggregation, but whether an additional sorting domain is also required is not clear. By fusing vasopressin prohormone (proVP) fragments to green fluorescent protein (eGFP) we have determined whether a sorting domain can function independently of the aggregative neurophysin domain. Although eGFP itself can be immunolocalised in the RSP of Neuro2A and AtT20 cells, most of the protein enters the constitutive pathway, and is found in the culture medium. In contrast, the N-terminal 27 residues of proVP promote residence in the RSP. Furthermore, only the processed form of this fusion was secreted when stimulated. We suggest a sorting mechanism based on the recognition of a sorting motif, the efficiency of which is enhanced by neurophysin aggregation. PMID- 10869552 TI - Secondary structure of the 5'-region of PGY1/MDR1 mRNA. AB - In order to identify the optimal target sites for antisense oligonucleotides in the human multiple drug resistance mRNA, the secondary structure of the 5' terminal part of this mRNA (nucleotides 1-678) was investigated. By using results of probing with ribonucleases T1, ONE and V1 and results of computer simulations, a model of the 5'-region of the PGY1/MDR1 mRNA was built. The molecule is formed by three major domains comprising several hairpins separated by single-stranded fragments. The predicted single-stranded regions of the PGY1/MDR1 mRNA efficiently bind complementary oligonucleotides. PMID- 10869553 TI - Heterocomplex formation between metastasis-related protein S100A4 (Mts1) and S100A1 as revealed by the yeast two-hybrid system. AB - S100A4 (Mts1) is a Ca(2+)-binding protein of the S100 family. This protein plays an important role in promoting tumor metastasis. In order to identify S100A4 interacting proteins, we have applied the yeast two-hybrid system as an in vivo approach. By screening a mouse mammary adenocarcinoma library, we have demonstrated that S100A4 forms a heterocomplex with S100A1, another member of the S100 family. The non-covalent heterodimerization was confirmed by fluorescence spectroscopy and electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. Mutational analysis revealed that replacement of Cys(76) and/or Cys(81) of S100A4 by Ser abolishes the S100A4/S100A1 heterodimerization, but does not affect the S100A4 homodimerization in vivo. PMID- 10869554 TI - PAI-1 stability: the role of histidine residues. AB - The role of the 13 histidine residues in plasminogen activator inhibitor 1 (PAI 1) for the stability of the molecule was studied by replacing these residues by threonine, using site-directed mutagenesis. The generated mutants were expressed in Escherichia coli, purified and characterized. All variants had a normal activity and formed stable complexes with tissue-type plasminogen activator. Most of these PAI-1 variants displayed a similar pH-dependency in stability as wild type PAI-1, with increased half-lives at lower pH. However, the variant His364Thr had a half-life of about 50 min at 37 degrees C and had almost completely lost its pH-dependency. Therefore, our data suggest that His(364), in the COOH terminal end of the molecule might be responsible for the pH-dependent stability of PAI-1. PMID- 10869556 TI - Bone morphogenetic protein-2 stimulates adipogenic differentiation of mesenchymal precursor cells in synergy with BRL 49653 (rosiglitazone). AB - Bone morphogenetic proteins (BMPs) were discovered as potent bone-inducing molecules. Their effect on adipogenic differentiation is not well understood, both stimulation and inhibition of the process have been described. We show here that BMP-2 strongly stimulates adipogenic differentiation of murine 3T3-L1 preadipocytes if applied together with an agonist of peroxisome proliferator activated receptor gamma (PPARgamma). On its own, BMP-2 (500 ng/ml) did not stimulate adipogenesis as quantified by flow cytometry with the lipophilic dye Nile Red. However, the protein strongly potentiated adipogenesis stimulated by the thiazolidinedione BRL 49653 as well as glycerol-3-phosphate dehydrogenase activity and induction of mRNAs for the adipogenic markers PPARgamma and adipsin. We confirmed the synergistic action of BMP-2 and BRL 49653 with primary cultures of rat bone marrow stromal cells. Our data demonstrate that BMP-2 can act as a potent adipogenic agent if presented together with activators of PPARgamma. PMID- 10869555 TI - Phosphorylation of CPI-17, an inhibitory phosphoprotein of smooth muscle myosin phosphatase, by Rho-kinase. AB - Phosphorylation of CPI-17 by Rho-associated kinase (Rho-kinase) and its effect on myosin phosphatase (MP) activity were investigated. CPI-17 was phosphorylated by Rho-kinase to 0.92 mol of P/mol of CPI-17 in vitro. The inhibitory phosphorylation site was Thr(38) (as reported previously) and was identified using a point mutant of CPI-17 and a phosphorylation state-specific antibody. Phosphorylation by Rho-kinase dramatically increased the inhibitory effect of CPI 17 on MP activity. Thus, CPI-17 as a substrate of Rho-kinase could be involved in the Ca(2+) sensitization of smooth muscle contraction as a downstream effector of Rho-kinase. PMID- 10869557 TI - The three-dimensional structure of human transaldolase. AB - The crystal structure of human transaldolase has been determined to 2.45 A resolution. The enzyme folds into an alpha/beta barrel structure and is thus similar in structure to other class I aldolases. Structure-based sequence alignment of available sequences of the transaldolase subfamily reveals that eight active site residues are invariant in the whole subfamily. Other invariant residues are mainly involved in the formation of the hydrophobic core of the enzyme. Noteworthy is a hydrophobic cluster consisting of five invariant residues. Human transaldolase has been implicated as an autoantigen in multiple sclerosis and four immunodominant peptide segments are located at the surface of the enzyme, accessible to autoantibodies. PMID- 10869558 TI - MSSP, a protein binding to an origin of replication in the c-myc gene, interacts with a catalytic subunit of DNA polymerase alpha and stimulates its polymerase activity. AB - MSSP has been identified as a protein that binds to both single- and double stranded sequences of a putative DNA replication origin sequence in the human c myc gene. MSSP possesses versatile functions, including stimulation of DNA replication, transcriptional regulation, apoptosis induction, and cell transformation coordinated by c-Myc. MSSP contains two RNP domains, RNP1-A and RNP1-B, both of which are necessary for all of the functions of MSSP. In this study, we found that MSSP binds to the N-terminal region of a catalytic subunit of a human DNA polymerase alpha via its RNP domains both in vitro and in human cells. Furthermore, MSSP was released from the putative DNA replication origin of the c-myc gene after it complexed with DNA polymerase alpha, and MSSP stimulated DNA polymerase activity in vitro. PMID- 10869559 TI - Protein phosphorylation/dephosphorylation in the inner membrane of potato tuber mitochondria. AB - Inside-out inner mitochondrial membranes free of matrix proteins were isolated from purified potato tuber (Solanum tuberosum L.) mitochondria and incubated with ?gamma-(32)PATP. Proteins were separated by SDS-PAGE and visualized by autoradiography. Phosphorylation of inner membrane proteins, including ATPase subunits, was strongly inhibited by the phosphoprotein phosphatase inhibitor NaF. We propose that an inner membrane phosphoprotein phosphatase is required for activation of the inner membrane protein kinase. When prelabelled inner membranes were incubated in the absence of ?gamma-(32)PATP, there was no phosphoprotein dephosphorylation unless a soluble matrix fraction was added. This dephosphorylation was inhibited by NaF, but not by okadaic acid. We conclude that the mitochondrial matrix contains a phosphoprotein phosphatase that is responsible for dephosphorylation of inner membrane phosphoproteins. PMID- 10869560 TI - Homogeneous longitudinal profiles and synchronous fluctuations of mitochondrial transmembrane potential. AB - This study reports for the first time (a) the longitudinal profile of the transmembrane potential (mDeltapsi) of single mitochondria using a Nernstian fluorescent probe and (b) the distribution of mDeltapsi fluctuations of mitochondria undergoing permanent depolarization. Our findings show that (1) mitochondria in different energetic conditions coexist in the same cell, (2) mDeltapsi is rather homogeneous along the entire length of single mitochondria, (3) mDeltapsi is not influenced by the surrounding cytoplasmic environment and (4) mDeltapsi fluctuations occur simultaneously in groups of mitochondria connected in a network. Taken together, these findings provide further evidence for a functional relationship between mitochondrial arrangement and energetic condition. PMID- 10869561 TI - Development and application of cytotoxic T lymphocyte-associated antigen 4 as a protein scaffold for the generation of novel binding ligands. AB - We have explored the possibilities of using human cytotoxic T lymphocyte associated antigen 4 (CTLA-4) as a single immunoglobulin fold-based scaffold for the generation of novel binding ligands. To obtain a suitable protein library selection system, the extracellular domain of CTLA-4 was first displayed on the surface of a filamentous phage as a fusion product of the phage coat protein p3. CTLA-4 was shown to be functionally intact by binding to its natural ligands B7-1 (CD80) and B7-2 (CD86) both in vitro and in situ. Secondly, the complementarity determining region 3 (CDR3) loop of the CTLA-4 extracellular domain was evaluated as a permissive site. We replaced the nine amino acid CDR3-like loop of CTLA-4 with the sequence XXX-RGD-XXX (where X represents any amino acid). Using phage display we selected several CTLA-4-based variants capable of binding to human alphavbeta3 integrin, one of which showed binding to integrins in situ. To explore the construction of bispecific molecules we also evaluated one other potential permissive site diametrically opposite the natural CDR-like loops, which was found to be tolerant of peptide insertion. Our data suggest that CTLA-4 is a suitable human scaffold for engineering single-domain molecules with one or possibly more binding specificities. PMID- 10869563 TI - Transcriptional regulation of the Saccharomyces cerevisiae DAL5 gene family and identification of the high affinity nicotinic acid permease TNA1 (YGR260w). AB - We have studied the transcript levels of YGR260w and YLR004c, two genes encoding members of the yeast Dal5p subfamily of the major facilitator family, and we show that they increase when extracellular nicotinic acid and thiamine, respectively, are absent. The deletion of YGR260w in a bna1 auxotrophic mutant for nicotinic acid prevents growth at low nicotinic acid concentration. This suggests that YGR260w is necessary for nicotinic acid import into the cell. The direct measurement of nicotinic acid uptake on whole cells demonstrates that YGR260w encodes the yeast high affinity nicotinic acid permease. Its apparent K(m) of 1.7 microM is low enough to allow the uptake of the low concentrations of nicotinic acid normally secreted by wild type cells. PMID- 10869562 TI - Identification of a novel secretory leukocyte protease inhibitor-binding protein involved in membrane phospholipid movement. AB - Previous studies have suggested that human salivary secretory leukocyte protease inhibitor (SLPI) inhibits HIV-1 by binding to a host cell surface protein of unknown identity. Using the yeast two-hybrid assay, we identified a gene sequence encoding a novel SLPI-binding protein (SLPI-BP). The 1.5-kb cDNA encodes a 318 amino acid protein with a predicted transmembrane segment near the C-terminus. Sequence analysis revealed that SLPI-BP is the human scramblase protein that is involved in the movement of membrane phospholipids. Co-expression of SLPI and SLPI-BP followed by an S-protein pulldown assay confirmed the specific interaction between these two proteins. Our data represent the first report for the identity of SLPI-BP. PMID- 10869564 TI - Functional crosstalk between phospholipase D(2) and signaling phospholipase A(2)/cyclooxygenase-2-mediated prostaglandin biosynthetic pathways. AB - We performed reconstitution analyses of functional interaction between phospholipase A(2) (PLA(2)) and phospholipase D (PLD) enzymes. Cotransfection of HEK293 cells with cytosolic (cPLA(2)) or type IIA secretory (sPLA(2)-IIA) PLA(2) and PLD(2), but not PLD(1), led to marked augmentation of stimulus-induced arachidonate release. Interleukin-1-stimulated arachidonate release was accompanied by prostaglandin E(2) production via cyclooxygenase-2, the expression of which was augmented by PLD(2). Conversely, activation of PLD(2), not PLD(1), was facilitated by cPLA(2) or sPLA(2)-IIA. Thus, our results revealed functional crosstalk between signaling PLA(2)s and PLD(2) in the regulation of various cellular responses in which these enzymes have been implicated. PMID- 10869565 TI - Regulation of UDP-glucose:ceramide glucosyltransferase-1 by ceramide. AB - We report that the expression of mRNA and the activity of UDP-glucose:ceramide (Cer) glucosyltransferase-1 (GlcT-1) of human hepatoma Huh7 and mouse melanoma B16 cells increases after treatment with bacterial sphingomyelinase or upon addition of short-chain Cer. Interestingly, however, GlcT-1 gene transcription was not increased by Cer when GlcT-1 cDNA was introduced with the CMV promoter in GlcT-1-deficient GM95 cells, suggesting that the normal promoter region of GlcT-1 gene is essential for the response. The conversion of C6-Cer to C6-GlcCer occurred much more rapidly in GlcT-1-overexpressing Huh7 cells than in mock transfectants. As a result, GlcT-1-overexpressing cells acquired a greater resistance to C6-Cer-mediated cell death. PMID- 10869566 TI - Dual regulation of the T-type Ca(2+) current by serum albumin and beta-estradiol in mammalian spermatogenic cells. AB - This study provides evidence for a novel mechanism of voltage-gated Ca(2+) channel regulation in mammalian spermatogenic cells by two agents that affect sperm capacitation and the acrosome reaction (AR). Patch-clamp experiments demonstrated that serum albumin induced an increase in Ca(2+) T current density in a concentration-dependent manner, and significant shifts in the voltage dependence of both steady-state activation and inactivation of the channels. These actions were not related to the ability of albumin to remove cholesterol from the membrane. In contrast, beta-estradiol significantly inhibited Ca(2+) channel activity in a concentration-dependent and essentially voltage-independent fashion. In mature sperm this dual regulation may influence capacitation and/or the AR. PMID- 10869567 TI - Cellular glucose-6-phosphate dehydrogenase (G6PD) status modulates the effects of nitric oxide (NO) on human foreskin fibroblasts. AB - Glucose-6-phosphate dehydrogenase (G6PD) plays an important role in cellular redox homeostasis, which is crucial for cell survival. In the present study, we found that G6PD status determines the response of cells exposed to nitric oxide (NO) donor. Treatment with NO donor, sodium nitroprusside (SNP), caused apoptosis in G6PD-deficient human foreskin fibroblasts (HFF1), whereas it was growth stimulatory in the normal counterpart (HFF3). Such effects were abolished by NO scavengers like hemoglobin. Ectopic expression of G6PD in HFF1 cells switched the cellular response to NO from apoptosis to growth stimulation. Experiments with 1H ?1,2,4oxadiazolo?4, 3-aquinoxalin-1-one and 8-bromo-cGMP showed that the effects of NO on HFF1 and HFF3 cells were independent of cGMP signalling pathway. Intriguingly, trolox prevented the SNP-induced apoptosis in HFF1 cells. These data demonstrate that G6PD plays a critical role in regulation of cell growth and survival. PMID- 10869568 TI - Cytochrome c oxidase rapidly metabolises nitric oxide to nitrite. AB - Previous studies have shown that the addition of nitric oxide to cytochrome c oxidase rapidly generates spectral changes compatible with the formation of nitrite at the binuclear haem:copper centre. Here we directly demonstrate nitrite release following nitric oxide addition to the enzyme. The nitrite complex is kinetically inactive and the off rate for nitrite was found to be slow (0.024 min(-1)). However, the presence of reductants enhances the off rate and enables cytochrome oxidase to catalyse the rapid oxidation of nitric oxide to nitrite free in solution. This may play a major role in the mitochondrial metabolism of nitric oxide. PMID- 10869569 TI - Changes in mitochondrial membrane potential during staurosporine-induced apoptosis in Jurkat cells. AB - Cytochrome c release from mitochondria is central to apoptosis, but the events leading up to it are disputed. The mitochondrial membrane potential has been reported to decrease, increase or remain unchanged during cytochrome c release. We measured mitochondrial membrane potential in Jurkat cells undergoing apoptosis by the uptake of the radiolabelled lipophilic cation TPMP, enabling small changes in potential to be determined. The ATP/ADP ratio, mitochondrial and cell volumes, plasma membrane potential and the mitochondrial membrane potential in permeabilised cells were also measured. Before cytochrome c release the mitochondrial membrane potential increased, followed by a decrease in potential associated with mitochondrial swelling and the release of cytochrome c and DDP-1, an intermembrane space house keeping protein. Mitochondrial swelling and cytochrome c release were both blocked by bongkrekic acid, an inhibitor of the permeability transition. We conclude that during apoptosis mitochondria undergo an initial priming phase associated with hyperpolarisation which leads to an effector phase, during which mitochondria swell and release cytochrome c. PMID- 10869570 TI - Rac-1 and Raf-1 kinases, components of distinct signaling pathways, activate myotonic dystrophy protein kinase. AB - Myotonic dystrophy protein kinase (DMPK) is a serine-threonine protein kinase encoded by the myotonic dystrophy (DM) locus on human chromosome 19q13.3. It is a close relative of other kinases that interact with members of the Rho family of small GTPases. We show here that the actin cytoskeleton-linked GTPase Rac-1 binds to DMPK, and coexpression of Rac-1 and DMPK activates its transphosphorylation activity in a GTP-sensitive manner. DMPK can also bind Raf-1 kinase, the Ras activated molecule of the MAP kinase pathway. Purified Raf-1 kinase phosphorylates and activates DMPK. The interaction of DMPK with these distinct signals suggests that it may play a role as a nexus for cross-talk between their respective pathways and may partially explain the remarkable pleiotropy of DM. PMID- 10869571 TI - Fourier transform infrared spectroscopic characterization of a photolabile precursor of glutamate. AB - Recently, it has been demonstrated that Fourier transform infrared spectroscopy (FTIR) detects conformational changes in the glutamate receptor ligand-binding domain that are associated with agonist binding. Combined with flash photolysis, this observation offers the prospect of following conformational changes at individual protein and agonist moieties in parallel and with high temporal resolution. Here, we demonstrate that gamma(alpha-carboxy-2-nitrobenzyl) glutamate (caged glutamate) does not interact with the protein, and that following photolysis with UV light the FTIR difference spectrum indicated changes in the protein tertiary and secondary interactions. These changes were similar to those observed for the protein upon addition of free glutamate. Thus, caged glutamate and its photolysis by-products are inert in this system, whereas the released glutamate exhibits full activity. Difference spectra of caged glutamate and of reaction analogs permitted identification of and correction for FTIR signals arising from the photolytic reaction and confirmed that its products are indeed glutamate and 2-nitrosophenyl glyoxalic acid. PMID- 10869572 TI - Protein synthesis is required for stabilization of hsp70 mRNA upon exposure to both hydrostatic pressurization and elevated temperature. AB - We have recently described that in chondrocytic cells high hydrostatic pressure (HP) causes a heat shock response via mRNA stabilization without a transcriptional activation of the hsp70 gene. In this study, we investigated whether this exceptional regulatory mechanism occurs more generally in different types of cells. Indeed, hsp70 mRNA and protein accumulated in HeLa, HaCat and MG 63 cells under 30 MPa HP, without DNA-binding of heat shock transcription factor 1 (HSF1) to the heat shock element of the hsp70 gene or formation of nuclear HSF1 granules, revealing a lack of transcriptional activation. Moreover, we observed that protein synthesis is needed for mRNA stabilization. Thus, high HP offers a model to study the mechanisms of hsp70 mRNA stabilization without HSF1-mediated induction of the heat shock gene response. PMID- 10869573 TI - Molecular cloning and functional characterization of the zebrafish ATP-gated ionotropic receptor P2X(3) subunit. AB - We describe a P2X subunit cloned from the zebrafish (Danio rerio) that is an orthologue of the mammalian P2X(3) subunit. Like the mammalian P2X(3), this receptor desensitizes rapidly in the presence of agonist. However, it differs in that alphabeta-meATP is a much less potent agonist than ATP and the antagonist TNP-ATP is not active at low nanomolar concentrations. Similar to the rat P2X(3) subunit, the zebrafish subunit forms hetero-oligomeric assemblies with the rat P2X(2) that possesses a phenotype distinct from either parent. This novel clone will provide an important basis for future experiments investigating the structure/function relationships of P2X subunit domains. PMID- 10869574 TI - Aquaporin 3 cloned from Xenopus laevis is regulated by the cystic fibrosis transmembrane conductance regulator. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) is essential for epithelial electrolyte transport and has been shown to be a regulator of epithelial Na(+), K(+), and Cl(-) channels. CFTR also enhances osmotic water permeability when activated by cAMP. This was detected initially in Xenopus oocytes and is also present in human airway epithelial cells, however, the mechanisms remain obscure. Here, we show that CFTR activates aquaporin 3 expressed endogenously and exogenously in oocytes of Xenopus laevis. The interaction requires stimulation of wild type CFTR by cAMP and an intact first nucleotide binding domain as demonstrated for other CFTR-protein interactions. PMID- 10869575 TI - Interhemispheric transfer of language in patients with left frontal cerebral arteriovenous malformation. AB - Cerebral arteriovenous malformations (AVMs) are frequently evaluated before therapeutic embolization by superselective injection of anesthetics into individual arterial branches so as to determine whether permanent occlusion would affect eloquent function. In Experiment 1, we used this adaptation of the Wada procedure to study three right-handed adult patients with left frontal cerebral AVMs by injecting vessels in Wernicke's and Broca's areas, respectively, and assessing language functions. The results showed that superselective testing in the inferior division of the left MCA in all three patients produced a dense Wernicke's aphasia. Injections into the left frontal regions, however, resulted in right paresis in all patients, but no language deficits including no loss of fluency. In Experiment 2, Patient 2 underwent fMRI activation for spontaneous word-list generation using multi-slice echo planar BOLD techniques at 1.5 Tesla. A voxel-by-voxel comparison of rest vs activation for each task was performed with a Z-score threshold of 2.5 SD for activated voxels. There was activation in the right hemisphere in the insula, frontal operculum pars opercularis, and inferior frontal gyrus, an area homologous to Broca's area in the left hemisphere. There was also activation in the left hemisphere in the Rolandic region, but language function was unaffected during Wada testing in this area. These data suggested that features of expressive language were no longer controlled by the left frontal lobe where the AVM was located, and provided new evidence for interhemispheric re-organization under conditions of chronic neurovascular disease. PMID- 10869576 TI - The contribution of recollection and familiarity to yes-no and forced-choice recognition tests in healthy subjects and amnesics. AB - Recent reports suggest that some amnesic patients perform relatively normally on forced-choice recognition memory tests. Their preserved performance may reflect the fact that the test relies more heavily on assessments of familiarity, a process that is relatively preserved in these patients, than do other recognition tests such as yes-no tests, which may rely more on recollection. The current study examined recognition memory using yes-no and forced-choice procedures in control and amnesic patients in order to determine whether the two tasks differentially relied on recollection and familiarity, and whether the extent of the recognition memory deficit observed in amnesia was dependent upon the type of recognition test used to measure performance. Results using the remember-know procedure with healthy subjects showed that there were no substantial differences in recognition accuracy or in the contribution of recollection to these two tasks. Moreover, amnesic patients were not found to perform better on a forced choice test than on a yes-no test, suggesting that familiarity contributed equally to these two types of recognition test. PMID- 10869577 TI - Attentional set shifting modulates the target P3b response in the Wisconsin card sorting test. AB - For years the Wisconsin card sorting test (WCST) has been used as a test of frontal lobe function. Recent event-related potential (ERP) research has shown large differences in the amplitude of P3b responses evoked by early and late trials within each WCST series ([8]: Barcelo F., Sanz M., Molina V., Rubia FJ. The Wisconsin Card Sorting Test and the assessment of frontal function: A validation study with event-related potentials. Neuropsychologia 1997;35:399 408). In this study, 16 normal subjects performed a WCST adaptation to investigate the role of attentional set shifting in these WCST P3b effects. Two control tasks were designed to examine whether early-late WCST P3b changes reflect category selection (attention) or category storage (memory) operations. Results suggest both a sharp P3b attenuation during shift WCST trials, followed by a gradual P3b build-up during post-shift trials. This P3b modulation could not be attributed to selection or storage of simple sensory stimulus dimensions, nor was it observed when the new rule was externally prompted by the first card in the WCST series. Instead, WCST P3b changes seem related to the endogenously generated shift in the perceptual rule used to sort the cards (i.e., the shift in set). The gradual build-up in P3b amplitude paralleled a progressive improvement in sorting efficiency over several post-shift WCST trials. A model based on formal theories of visual attention and attentional set shifting is proposed to account for these effects. The model offers firm grounds for prediction and bridges the gap between related clinical and experimental evidence. PMID- 10869578 TI - Disturbances of spatial vision and object vision correlate differently with regional cerebral glucose metabolism in Alzheimer's disease. AB - There is evidence of two neuronal systems for visual cognition which are referred to as spatial vision and object vision. We subjected 49 patients with mild-to moderate probable Alzheimer's disease (AD) to tasks associated with these two types of visual cognition. Visual counting was employed as a task to assess visuospatial recognition. Identification of overlapping figures and discrimination of visual forms were used as visuoperceptual tasks that require an ability to recognize objects. Regional cerebral glucose metabolism (rCMRglc) of the subjects was also determined by positron emission tomography. Multivariate statistics controlling for the confounding factors assessed the correlation between the task performances and the rCMRglc. The visual counting score significantly correlated with the metabolic rate of the bilateral inferior parietal lobules. On the other hand, the scores of the visuoperceptual tasks significantly correlated with the metabolic rate of the right middle temporal gyrus and the right inferior parietal lobule. The present study showed that visuospatial disturbance was related to bilateral parietal metabolism, and that visuoperceptual disturbance was related to right temporo-parietal metabolism in patients with mild-to-moderate AD. PMID- 10869579 TI - Steroid fluctuations modify functional cerebral asymmetries: the hypothesis of progesterone-mediated interhemispheric decoupling. AB - This study examines the modulation of functional cerebral asymmetries by gonadal hormones in three distinct groups. Young, normally cycling women performed a prototypical left (lexical decision) and two prototypical right-hemispheric tasks (figural comparison and face discrimination) during the low steroid menses and the high steroid midluteal phase. Saliva progesterone levels were measured with radioimmunoassay (RIA). Parallel to younger females, young men, and postmenopausal women were tested at matching time intervals. Results revealed significant interactions between cycle phase and visual half-field in the accuracy of all three tasks for the younger women; stronger lateralization patterns occurring during menses, while a more bilateral or at least less asymmetric cerebral organization predominated the midluteal phase, when highest levels of progesterone appear. Progesterone seemed to have a significant influence on lateralization in the figural comparison task, with high hormone levels enhancing the performance of the left hemisphere (for this task subdominant), thereby decreasing asymmetry. After menopause, when the levels of gonadal hormones are lower and more stable, the lateralization patterns for all three tasks were similar to those of men and normally cycling women during menses. These results make it likely that steroids and especially progesterone are able to reduce cerebral asymmetries. We hypothesize that progesterone attenuates the effect of glutamate on non-NMDA receptors. This could diminish cortico-cortical transmission which is mostly dependent on a glutamate-induced initial EPSP in pyramidal neurons which receive transcallosal input. The reduction in callosal transfer could then suppress the functional asymmetries. PMID- 10869580 TI - Comparison of overall brain volume and midsagittal corpus callosum surface area as obtained from NMR scans and direct anatomical measures: a within-subject study on autopsy brains. AB - Formalin fixed brains obtained from autopsy were studied by magnetic resonance imaging, using FLASH and MPR sequences. The overall volume and the midsagittal surface area of the corpus callosum (cc) of these brains were also directly measured by water displacement for overall volume determination and morphometry for cc surface area measures, allowing comparisons of these measures in a within subject design. Thin NMR sections (1 mm thickness for FLASH and 1.25 mm for MPR) yielded overall brain volume estimates that did not differ significantly from direct volume estimations but thicker (5 mm) sections led to a significant overestimation of brain volume for NMR measures relative to direct displacement measures. There was no overall effect of type of NMR measure: FLASH and MPR scans did not yield significantly different values for overall brain volume or callosal surface area. Direct and NMR measures of the corpus callosum surface at midsagittal level did not yield significantly different estimates. PMID- 10869581 TI - When does inter-hemispheric integration of visual events emerge in infancy? A developmental study on 19- to 28-month-old infants. AB - Simultaneous attention in the two visual fields and interhemispheric integration of visual information was studied in 19-23 and 24-28-month-old infants. The stimuli were schematic faces within which the pair of eyes was made of either two identical (two circles or two triangles) or two different eyes (triangle-circle, circle-triangle). The faces were presented either in one visual hemifield, on the right or left side of a central fixation point (unilateral presentation), or across the two visual hemifields (bilateral presentation), with one eye of the stimulus on each side of the fixation point. The task was an operant conditioning task where the children had to decide whether the shapes of the two eyes were identical or not. The results show that even the younger subjects were able to perform the task when presented in the unilateral presentation condition, whereas only children aged 24 months and older could learn the task when presented in the bilateral condition. It is concluded that simultaneous attention to the two visual fields and inter-hemispheric co-ordination of visual information emerge very late in development at about the age of 24 months. PMID- 10869582 TI - Weighing the evidence for cross over in neglect. AB - When patients with left-sided neglect are asked to bisect horizontal lines, they tend to place their marks to the right of the line's objective mid-point. However, when asked to bisect short lines they are either more accurate or paradoxically cross over and place their marks to the left of the objective mid point. Previous explanations of the cross over phenomenon have considered specific aberrations of spatial attention. However, these explanations make no predictions about judgments of non-spatial stimuli. Two patients with right brain damage were asked to judge weights placed on both hands simultaneously. They were biased in reporting weights on the right as being heavier than those on the left. This rightward bias changed with lighter pairs of weights presented in the context of equal reference weights. In one patient the directional bias was eliminated and in the other the bias was reversed so that she was more likely to report the left weight as heavier than the right. These data suggest that a phenomenon analogous to cross over in line bisections also occurs with judgments of non-spatial stimuli. Representations of stimuli appear to be influenced by features of the stimuli encountered on-line and by memory traces of similar stimuli encountered previously. With an attentional deficit, memory traces influence the magnitude of the representation derived on-line disproportionately. PMID- 10869583 TI - Impaired control of an action after supplementary motor area lesion: a case study. AB - The kinematics of the action formed by reaching-grasping an object and placing it on a second target was studied in a patient who suffered from an acute vascular left brain lesion, which affected the Supplementary Motor Area proper (SMA proper) (Matelli M, Luppino G. Thalamic input to mesial and superior area 6 in the macaque monkey. Journal of Comparative Neurology 1996;372:59-87, Matelli M, Luppino G, Fogassi L, Rizzolatti G. Thalamic input to inferior area 6 and area 4 in the macaque monkey. Journal of Comparative Neurology 1989;280:468-488), and in five healthy control subjects. The reach kinematics of the controls was affected by the positions of both the reaching-grasping and the placing targets (Gentilucci M, Negrotti A, Gangitano M. Planning an action. Experimental Brain Research 1997;115:116-28). In contrast, the reach kinematics of the patient was affected only by the position of the reaching-grasping target. By comparing these results with those previously found in Parkinson's disease patients executing the same action (Gentilucci M, Negrotti A. Planning and executing an action in Parkinson's disease patients. Movement Disorders 1999;1:69-79, Gentilucci M, Negrotti A. The control of an action in Parkinson's disease. Experimental Brain Research 1999;129:269-277), we suggest that the anatomical "motor" circuit formed by SMA-proper (see above), Basal Ganglia (BG) and Thalamus (Alexander GE, Crutcher MD. Functional architecture of basal ganglia circuits: neural substrates of parallel processing. Trends in the Neurosciences 1990;13:266-271, Hoover JE, Strick PL. Multiple output channels in the basal ganglia. Nature 1993;259:819 821) may be involved in the control of actions: SMA-proper assembles the sequence of the action, whereas BG updates its parameters and stores them. PMID- 10869584 TI - The effect of electroconvulsive therapy (ECT) on implicit memory: skill learning and perceptual priming in patients with major depression. AB - While explicit memory in amnesics is impaired, their implicit memory remains preserved. Memory impairment is one of the side effects of electroconvulsive therapy (ECT). ECT patients are expected to show impairment on explicit but not implicit tasks. The present study examined 17 normal controls and 17 patients with severe major depressive disorder who underwent right unilateral ECT. Patients were tested in three sessions: 24-48 hours prior to, 24-48 hours following the first ECT, and 24-48 hours following the eighth ECT. The controls were tested in three sessions, at time intervals that paralleled those of the patients. Implicit memory was tested by the perceptual priming task - Partial Picture-Identification (PPI). The skill learning task used entailed solving the Tower of Hanoi puzzle (TOHP). Explicit memory was tested by picture recall from the PPI task, verbal recall of information regarding the TOHP, and by the Visual Paired Association (VPA) test. Results showed that explicit questions about the implicit tasks were impaired following ECT treatment. Patients' learning ability, as measured by the VPA task, was only impaired in the first testing session, prior to ECT treatment, reflecting the effect of depression. In addition, groups only differed in the first session on the learning rate of the skill learning task. Perceptual priming was preserved in the patients' group in all sessions, indicating that it is resilient to the effect of depression and ECT. The results are interpreted in terms of the differential effect of depression and ECT on explicit and implicit memory. PMID- 10869585 TI - The effect of graded aversive stimuli on limbic and visual activation. AB - Activation studies have shown that in response to evocative visual stimuli, brain activity increases in the visual cortex and limbic areas. However, non-affective characteristics of these images, such as color composition and visual complexity, confound the interpretation of these results. To address this issue, we had subjects rate over 100 images on aversive intensity (facial mutilation, dead bodies) and semantic complexity (number of objects subjects could name). From these images, we assembled digitized image sets of non-aversive, mild and strong intensity, balanced on semantic complexity and content (human faces and figures), and adjusted for color composition. A fourth condition consisted of a fixation cross on a blank screen. Thirteen subjects underwent eight positron emission tomography scans using the [(15)O] water methodology. Measurement of skin conductance was recorded simultaneously. All picture conditions, relative to the blank screen, activated the amygdalae and bilateral orbitofrontal cortex, while we found activation trends associated with increasing aversive content in the sub lenticular region. Skin conductance increased during all picture conditions. Relative to the non-aversive pictures, aversive image content caused modulation of occipital and occipital-temporal cortex. These results demonstrated activation of the amygdala to salient, arousing stimuli, and not just aversive stimuli. In addition, they suggest that pictorial complexity, as indexed by our semantic measure, does not account for the modulation of visual cortex by aversive, emotional stimuli. PMID- 10869586 TI - Language and calculation within the parietal lobe: a combined cognitive, anatomical and fMRI study. AB - We report the case of a patient (ATH) who suffered from aphasia, deep dyslexia, and acalculia, following a lesion in her left perisylvian area. She showed a severe impairment in all tasks involving numbers in a verbal format, such as reading aloud, writing to dictation, or responding verbally to questions of numerical knowledge. In contrast, her ability to manipulate non-verbal representations of numbers, i.e., Arabic numerals and quantities, was comparatively well preserved, as evidenced for instance in number comparison or number bisection tasks. This dissociated impairment of verbal and non-verbal numerical abilities entailed a differential impairment of the four arithmetic operations. ATH performed much better with subtraction and addition, that can be solved on the basis of quantity manipulation, than with multiplication and division problems, that are commonly solved by retrieving stored verbal sequences. The brain lesion affected the classical language areas, but spared a subset of the left inferior parietal lobule that was active during calculation tasks, as demonstrated with functional MRI. Finally, the relative preservation of subtraction versus multiplication may be related to the fact that subtraction activated the intact right parietal lobe, while multiplication activated predominantly left-sided areas. PMID- 10869587 TI - Immunological stress in rats induces bodily alterations in saline-treated conspecifics. AB - This work was developed during an investigation on the neuroendocrine-immune interaction in rats immune challenged with sheep red blood cells (SRBC). The structures used for evaluating the immunological response was the direct plaque forming cells (PFC). An inbred strain of rat was used to overcome the problem of different timings in the peak humoral immune response. Normal rats were injected intraperitoneally with saline or SRBC and were killed 0, 3, 4, 5, and 6 days later. Body and gland weights were recorded, and. serum levels of corticosterone and prolactin were quantified by radioimmunoassay. The hormone levels and gland weights of the saline conspecifics and SRBC-treated rats were found to be similar. When new rats were housed in a separate room and treated with physiological saline, there were again no differences in the body and gland weights or the serum hormone levels between the two home cage control (HCC) groups of animals. Compared with saline conspecifics and SRBC-treated groups, the HCC groups had higher body weights from the third to the sixth day of treatment and had lower gland weights in absolute and relative analysis (pituitary, thyroid, and adrenals) mainly on the fourth and fifth days; thymus weights were highest on the third day. Corticosterone and prolactin levels were significantly lower on the fifth and sixth days, respectively. Because SRBC-treated rats showed a peak direct immune response on the fourth and fifth days and showed peak corticosterone levels on the fifth day after treatment, we conclude that the former animals were under stress and influenced their saline conspecifics through sound or smell. This conclusion agrees with other studies, showing that physically or emotionally stressed rats can influence conspecifics through noise and body odors. PMID- 10869588 TI - Age-related changes in adaptive macronutrient intake in swimming male and female Lou rats. AB - To evaluate the age-related changes in capacity to adjust the nutrient intake to needs, self-selecting male and female Lou/C/jall rats of 4, 6, 12, 16 and 23 months of age were submitted to a swimming exercise. They were given 6 consecutive days of moderate intensity training (3 x 15 minutes per day). Exercise and postexercise periods were compared with results from the pretraining period. During swimming, a body weight loss and a decrease in both caloric intake and fat selection were observed. This effect was more marked in older groups compared to 4 month-old groups. An increase in protein intake was observed in females, specially in older groups, whereas no effect was seen in males. The ability to increase caloric ingestion and regain weight during the postexercise period decreased with advancing age and was better in females than in males. We also showed an age-related effect on the recovery of initial nutrient intake rate that was more pronounced and more precocious for males. Moreover, males tended to decrease their protein intake, whereas females significantly increased it. The present findings suggest a decrease of capacity of adjusting feeding behavior to metabolic needs in aged rats, may be due to a deterioration of the central control of food intake. PMID- 10869589 TI - Effects of inescapable stress and treatment with pyridostigmine bromide on plasma butyrylcholinesterase and the acoustic startle response in rats. AB - Pyridostigmine bromide (PB) is a reversible, peripherally active inhibitor of acetylcholinesterase (AChE) activity, and is recommended by the military as a pretreatment against potential nerve gas exposure. Recent evidence suggests that exposure to inescapable stressors allows PB to cross the blood-brain barrier, and thereby affect central AChE activity in mice. Here, we evaluated the functional impact of a stress/PB treatment interaction on acoustic startle responding and plasma butyrylcholinesterase (BuChE) activity in male Sprague-Dawley rats. To model the treatment protocol used by the military, PB was delivered in the drinking water of rats for 7 consecutive days. The morning after the start of PB treatment, and for the next 6 days, half the rats were exposed to 1 h of supine restraint stress. We therefore employed a 2 x 2 (stress x PB treatment) between groups design. Exposure to supine stress alone induced a persistent decrease in plasma BuChE activity. Further decreases in BuChE activity were not observed in rats exposed to supine restraint and PB treatment. Exposure to stress also induced an exaggerated startle response, evident on the last day of stress and 24 h after stressor cessation. Treatment with PB alone produced an exaggerated startle response over the same time period, albeit to a lesser degree. Although treatment with PB concurrent with stress did not produce further changes in either BuChE activity or acoustic startle responding, stress-induced alterations in drinking behavior (and thereby the dose of PB ingested) may have affected these results. Persistent stress-induced reductions in BuChE activity may increase the risk of adverse reactions to cholinomimetics. PMID- 10869590 TI - Responses to basic taste qualities in rats selectively bred for high versus low saccharin intake. AB - Rats selectively bred for relatively high (HiS) and relatively low (LoS) saccharin intake were offered sweet (sucrose), bitter (quinine, sucrose octaacetate), salty (sodium chloride), starchy (Polycose((R))), and sour (citric acid) solutions at several concentrations; sucrose/quinine, and sucrose/citric acid mixtures were also tested. Compared to HiS rats, LoS rats displayed weaker preferences for and lower consumption of sweet, salty, and starchy solutions. HiS and LoS rats did not differ in responses to simple bitter or sour solutions or to adulteration of sucrose with citric acid. However, quinine adulteration reduced sucrose preference more among LoS rats. Thus, selection on a saccharin intake phenotype has yielded line differences on all hedonically positive tastants and, probably as a consequence of that difference, greater finickiness specifically towards bittersweet solutions in the low saccharin-consuming line. Additional work can clarify the psychobiological mechanisms for the phenotypic difference and, potentially, the reasons for its relationship to measures of emotionality. PMID- 10869591 TI - Bitter taste perception and severe vomiting in pregnancy. AB - Hyperemesis gravidarum or severe vomiting during pregnancy is a condition of elusive etiology that can harm both mother and fetus. This study examined the association between increased bitter-taste perception and history of hyperemesis gravidarum. Bitter-taste perception varies genetically and can be altered with conditions that damage taste-related cranial nerves. Sixty women were divided into high- (n = 21) and low-vomit (n = 39) groups based on vomiting exposure across all pregnancies and were screened for genetic variation in taste with bitterness of saturated 6-n-propylthiouracil (PROP) delivered on filter paper. Supertasters perceive PROP as intensely bitter; nontasters, as only weakly. Each reported their history of dysgeusia (persistent taste) and taste-related pathology (otitis media and head trauma). The vomit groups did not differ in the frequency of supertasters, but the high-vomit group had fewest nontasters. The high-vomit group also reported dysgeusia most frequently. A subsample (13 high vomit and 18 low-vomit women) rated the taste intensity of sodium chloride (1 mol), sucrose (1 mol), citric acid (0.0032 mol), and quinine hydrochloride (0.001 mol) applied to areas innervated by cranial nerves VII and IX. The groups only varied significantly in bitterness of quinine hydrochloride. High-vomit women tasted least bitterness on the anterior tongue (chorda tympani branch of VII) and highest bitterness on the posterior tongue (cranial nerve IX) and palate (superficial petrosal branch of VII). In high-vomit women, elevated bitterness on the posterior tongue and palate does not appear related to hydrochloric acid exposure in vomitus; it may explain the occurrence of dysgeusia. This pattern of spatial taste perception may indicate altered oral sensations that if present during pregnancy, could increase the risk of hyperemesis. PMID- 10869592 TI - Locomotor response to an open field during C57BL/6J active and inactive phases: differences dependent on conditions of illumination. AB - Time of day has proven to be a source of variability in diverse behavioral measures. Knowledge of the pattern of this temporal effect as well as its origin (exogenous or endogenous) is essential for a precise description of any behavior. This study analyzed the effect of the external light-dark cycle and the internal rest-activity rhythm on the response of C57BL/6J mice to a novel environment. In a first experiment, animals maintained in a 12:12-h light-dark cycle were tested in an open field at six different times of day. A diurnal rhythm of ambulation in the open field was observed with greater levels of activity exhibited by those groups tested at night. Long-term and short-term behavioral habituation to spatial novelty were also affected by phase of the light-dark cycle. A second experiment was designed to control for any direct effect of the light-dark cycle by keeping the animals in dim green light where entrainment was maintained by a skeleton photoperiod (two 15-min bright-light pulses separated by 12 hours of green, dim light). This second group of animals was tested at two different circadian phases under the same conditions of illumination. One group was tested during the subjective night and another group during the subjective day, i.e., 2 or 14 h after the onset of the active phase, as assessed by wheel-running behavior. No effect of circadian phase on ambulation or habituation of this response to the open field was observed in these animals. Taken together, these results suggest that spatial novelty is equally arousing regardless of circadian phase and that the conditions of illumination can dramatically alter the response to a novel environment. PMID- 10869593 TI - Phasic response of the photoperiodic clock to wavelength and intensity of light in the redheaded bunting, Emberiza bruniceps. AB - We have investigated phasic response of the photoperiodic clock to wavelength (color) and intensity of light in the male redheaded bunting (Emberiza bruniceps). Two experiments were performed. Experiment I examined whether varying the wavelength and intensity of first (entraining, E) and/or second (inducing, I) light pulse will alter the effects of a skeleton photoperiod. Birds were subjected for a period of 6 weeks to skeleton photoschedules (6L:5D:1L:12D; 6 h E light pulse, 1 h I-light pulse) containing either an E-pulse at 5 or 20 lx coupled with I-pulse at 100 lx, or an E-pulse at 100 lx, coupled with I-pulse at 5- or 20-lx intensity. Additional two groups that received both E- and I-pulses at 100 lx served as controls. All photoschedules were employed in two colors white and red (654 nm). There was the wavelength- and intensity-dependent stimulation of the testis growth and development. Long wavelengths of light (red light) induced faster and greater gonadal response, but the effects were also intensity dependent. Experiment II tested whether in the photostimulated birds held on long photoperiods the change in wavelength and intensity of light hours in the morning (entraining period) or the evening (inducing period) will influence the maintenance of the photoperiodic sensitivity. Birds were subjected initially to a long photoperiod of 14L:10D (L = approximately 500 lx; D = 0 lx) and then after 3 weeks, a 4-h light period in the morning (zeitgeber time, zt, 0 4) or in the evening (zt 10-14) of 14 L was substituted with white, green (528 nm), or red (654 nm) light at approximately 20-lx intensity. One group maintained on 14L:10D and other exposed to 10L:14D served as controls. After another 7 weeks, all birds were subjected to 16L:8D for a further 4 weeks to test for their responsivity to long-day photostimulation as a consequence of exposure to different experimental photoperiods. Testes regrew under 16L:8D only in birds that were exposed to 10L:14D or to 14L:10D with a green light pulse. However, there was no effect of the timing (morning, evening) of the light pulse. Taken together, the results from both the experiments indicate that in the redheaded bunting (1) the photoperiodic clock responds differentially to different wavelengths (colors, spectra) and intensities of light, and (2) the effects of wavelength and intensity of light on the clock are phase dependent, and such phasic effects can be seen in skeleton photoperiods in which light is applied discretely at different circadian phases. PMID- 10869594 TI - The relationship between isometric force requirement and forelimb tremor in the rat. AB - To explore the effects of isometric force of rodent forelimb contraction on forelimb tremor, rats were trained to press downward on an isometric force transducer to raise a water-filled dipper cup and maintain force to keep the dipper in the raised position while licking. Force requirements were then manipulated parametrically to measure the effects of escalating force output on forelimb tremor and other variables. In the Peak-Force greater than Hold-Force (PF > HF) manipulation, the forces required to raise the dipper were 20, 40, and 60 g (each condition for about 2 weeks), while the force required to maintain the dipper in the raised position remained 6.7 g for all three conditions. In the Peak-Force equal to the Hold-Force (PF = HF) manipulation, rats were required to maintain the "dipper-raising" force throughout the response. The forces required were 20 g, 40 g, and 60 g (each for 2 weeks). For all force requirement manipulations, data were analyzed within and across conditions. As expected, force output increased with increased force requirements. Spectral analysis of force-time records revealed that during all manipulations, high-frequency (>10 Hz) forelimb tremor increased with increased force output, an effect that is consistent with human studies, and that may reflect increases in the number of motor units firing at higher rates. Additionally, with the exception of the 60-g PF = HF condition, there were within-condition decreases in tremor and increases in task engagement, evidence suggesting increased muscle strength as a function of experience (i.e., "physical training"). Taken together, the results suggest that the rodent-based method may provide a valuable, noninvasive functional assay for animal models of disorders that affect skeletal muscle control in humans. PMID- 10869595 TI - Effects of hypnosis on diffuse noxious inhibitory controls. AB - The neurophysiological mechanisms of hypnotic analgesia are still under debate. It is known that pain occurring in one part of the body (counterstimulation) decreases pain in the rest of the body by activating the diffuse noxious inhibitory controls (DNICs). The aim of this study was to explore the effects of hypnosis on both pain perception and heterotopic nociceptive stimulation. The A forms of both the Harward Group Scale of Hypnotic Susceptibility and the Stanford Hypnotic Susceptibility Scale were administered to 50 healthy students. Twenty subjects were selected and assigned to two groups: group A, consisting of 10 subjects with high hypnotic susceptibility; and group B, consisting of 10 subjects with low hypnotic susceptibility. The subjects were then randomly assigned first to either a control session or a session of hypnotic analgesia. The nociceptive flexion reflex (RIII) was recorded from the biceps femoris muscle in response to stimulation of the sural nerve. The subjective pain threshold, the RIII reflex threshold, and the mean area with suprathreshold stimulation were determined. Heterotopic nociceptive stimulation was investigated by the cold pressor test (CPT). During and immediately after the CPT, the subjective pain threshold, pain tolerance, and mean RIII area were determined again. The same examinations were repeated during hypnosis. Hypnosis significantly reduced the subjective pain perception and the nociceptive flexion reflex. It also increased pain tolerance and reduced pain perception and the nociceptive reflex during the CPT. These effects were found only in highly susceptible subjects. However, the DNIC's activity was less evident during hypnosis than during the CPT effects without hypnosis. Both hypnosis and DNICs were able to modify the perception of pain. It seems likely that DNICs and hypnosis use the same descending inhibitory pathways for the control of pain. The susceptibility of the subject is a critical factor in hypnotically induced analgesia. PMID- 10869596 TI - Olfactory sensitivity and specificity of Arctic char, Salvelinus alpinus, to a putative male pheromone, prostaglandin f(2)alpha. AB - Actively spawning male Arctic char, Salvelinus alpinus, release immunoreactive F prostaglandins into the water that attract ovulated females and elicit spawning behaviour. To investigate a possible role played by the olfactory system, we determined the sensitivity and specificity of the char to prostaglandins by the electro-olfactogram (EOG) using a newly devised stimulatory apparatus. Of 18 prostaglandins tested, PGF(2)alpha and two synthetic analogues, 16,16-dimethyl PGF(2)alpha and U-46619, were detected at threshold concentrations of 0.5-1.0 x 10(-11) M, and at a concentration of 10(-8) M evoked an EOG response equivalent to that of 10(-5) M L-serine. Olfactory nerve twig recordings further demonstrated that the EOG responses were transduced into nerve impulses and transmitted to the brain. No difference was found in the responses between sexes and maturational stages. We conclude that PGF(2)alpha is a candidate of the male pheromone of Arctic char, supporting our previous behavioural studies. PMID- 10869597 TI - Urinary testosterone levels in the male blind mole rat (Spalax ehrenbergi) affect female preference. AB - This study investigated the sexual attraction of female blind mole rats to four groups of male mole rats: (a) intact males raised in captivity; (b) intact males trapped in the field; (c) captive males injected with testosterone; (d) captive castrated males. In the first part we measured blood testosterone, androstenedione, and dihydrotestosterone (DHT) levels, by radioimmunoassay; and urine testosterone levels, measured by GC-MS. The second part examined the relationship between urine testosterone levels in males and their attractiveness to females. Higher blood and urine testosterone levels were found in the field animals and in those injected with testosterone compared to captive intact or castrated animals: urine testosterone levels in the two other groups were not detectable. Blood androstenedione levels were also higher in the field animals and in those injected with testosterone compared to captive intact or castrated mole rats. Blood dihydrotestosterone levels were not detectable in all four experimental groups. Female mole rats chose to spend a longer period of time next to males with high blood and urine testosterone levels and high blood androstenedione levels than next to those with lower levels of these hormones. Because courtship and sexual behavior are influenced both by high levels of blood and urine testosterone and high levels of blood androstenedione, we suggest that the low levels of courtship and other sexual behavior in captive mole rats may be related to the lack of female attraction to these males, which display low levels of all three parameters. PMID- 10869598 TI - Effects of repeated restraint in Japanese quail genetically selected for contrasting adrenocortical responses. AB - Behavioral and adrenocortical responses to repeated mechanical restraint were compared in 28-day-old to 31-day-old male Japanese quail from two genetic lines divergently selected for reduced (low stress, LS) or exaggerated (high stress, HS) plasma corticosterone (C) responses to brief immobilization. Restraint in a metal crush cage for 5 min elicited immobility and silence in all the birds. Circulating C levels were considerably higher in quail of both lines following restraint than in the undisturbed controls of either line. As expected, both the behavioral and physiological effects were more pronounced in HS than in LS birds. Struggling increased with repeated restraint in HS and LS quail, thus suggesting behavioral habituation to the stressor in both lines. On the other hand, a line effect on the pattern of adrenocortical responses was revealed upon subtracting the change in plasma C concentrations from Day 1 to Day 4 in the undisturbed controls from the corresponding change in restrained birds. Thus, unlike LS quail, in which there were no detectable effects of repeated restraint, the adrenocortical responses of HS birds showed evidence of experience-dependent sensitization. Our results demonstrate the importance of the background genome in determining the patterns of the behavioral and adrenocortical responses elicited by repeated exposure to stressful stimulation. The present results and those of previous studies could be explained in one or both of two ways: that underlying fearfulness is lower in LS than HS quail or that they adopt active or passive coping strategies, respectively. Our findings may also have important implications for poultry welfare and productivity. @ 2000 Elsevier Science Inc. PMID- 10869599 TI - Cholecystokinin receptors do not mediate the suppression of food-motivated behavior by lipopolysaccharide and interleukin-1 beta in mice. AB - During the course of an infection, profound metabolic and behavioral changes are observed. The resulting decrease in food intake can be reproduced by administration of lipopolysaccharide (LPS) or the proinflammatory cytokines (e.g., interleukin-1 [IL-1] and tumor necrosis factor it induces. To test the possibility that cholecystokinin (CCK) mediates anorexia induced by IL-1 beta and LPS, mice trained to poke their noses in a hole to obtain a food reward according to a fixed ratio (1 reward per 20 actions) were pretreated with the CCK-A receptor antagonist L364,718 (at 1 mg/kg) or with the CCK-B receptor antagonist L365,260 (50 microg/kg) before being injected with LPS (100 microg/kg) or IL-1 beta (20 microg/kg). All injections were given via the intraperitoneal (i.p.) route. In spite of its ability to block the effects of exogenous CCK-8 on food motivated behavior in mice, the CCK-A receptor antagonist did not block the depressive actions of LPS and IL-1 beta on food-motivated behavior. The CCK-B receptor antagonist was not more effective at blocking. These results do not support a role for CCK in the anorexic effect of LPS and IL-1 beta. PMID- 10869600 TI - Neural transplants. effects On startle responses in neonatally MSG-treated rats. AB - Neonatal monosodium glutamate (MSG) treatment has been associated with dysfunctions in stress responses. Therefore, the present study aimed at examining the acoustic startle response (ASR) in MSG-treated rats and the effects of fetal neural transplantation. Male and female rats were given MSG (4 mg/g) or saline on alternate days from days 2-10 after birth. To determine whether fetal transplants could reverse behavioral impairments observed in MSG-treated rats, at 12 days of age MSG-treated rats received either arcuate nucleus (AN), cortical fetal grafts, or sham surgery into the third ventricle. ASR amplitude was measured at 35-40 days of age, and again in adulthood. MSG produced the expected decrease in the density of hypothalamic neuropeptide Y (NPY) and tyrosine hydroxylase (TH) in the AN area. Corticotropin-releasing factor (CRF) neurons/fibers were not affected by MSG. Pituitary atrophy was observed in all MSG rats. We report a permanent increase in the amplitude and reduction in short-term habituation of ASR in all MSG-treated rats. No effect was observed on long-term habituation in male rats. Cortical, but not AN tissue significantly reduced the magnitude of ASR in MSG animals. The results are discussed in terms of the central pathways mediating ASR, in particular hypothalamo-amygdala connections. It is considered that nonspecific factors mediate recovery produced by cortical tissue grafts, as observed in other models of neural transplantation. PMID- 10869601 TI - Effects of taste stimulation on beta-endorphin levels in rat cerebrospinal fluid and plasma. AB - Opioids are suggested to be involved in generation of palatability and facilitation of consumption of food and fluid. We measured the level of an endogenous opioid, beta-endorphin, in the cerebrospinal fluid (CSF) and plasma after free drinking of water and taste solutions in Wistar rats. When the water deprived animals were allowed to drink 10 mL of water, the level of beta endorphin increased significantly 60 and 90 min after the start of drinking in both samples. beta-Endorphin in the CSF increased most after ingestion of 0.5 M sucrose and 0.005 M saccharin followed by 0.1 M NaCl, 0.1 mM quinine and water. An intragastric infusion of 7 mL of water did not change the beta-endorphin level. Essentially the same results were obtained for plasma samples except that NaCl and quinine solutions did not increase beta-endorphin levels. Sucrose became ineffective in releasing beta-endorphin in both samples after the establishment of conditioned taste aversions to this taste stimulus. These results suggest that the release of beta-endorphin is positively correlated with the palatability of taste stimuli, and that CSF beta-endorphin also reflects the reinforcement of fluid intake in thirsty animals. PMID- 10869602 TI - Anticholinergic effects in frogs in a Morris water maze analog. AB - We determined the effect of two doses of the centrally acting anticholinergic drug, atropine sulfate (AS), on the performance of female Northern Leopard frogs (Rana pipiens) in a visual cue analog of the Morris water task. Untreated frogs learned the visually cued task, while frogs treated with 150 mg/kg AS were significantly slower than controls in learning to escape warm water by finding a visible platform, and there was a dose-dependent response, with frogs treated with 50 mg/kg AS performing midway between the higher dose and control frogs. These results suggest that the general role of the cholinergic system in learning is important in amphibians, and that this role is evolutionarily conserved across vertebrate species. PMID- 10869603 TI - Leptin-induced decrease in food intake in chickens. AB - The effect of intracerebroventricular (i.c.v.) injection of leptin was investigated using broiler and Single Comb White Leghorn (SCWL)-type chickens. These represent relatively fast- and slow-growing birds, respectively. The i.c.v. injection of leptin decreased food intake in both broilers and Leghorns in a dose dependent manner. The most efficacious dose appeared to be 10 microg in both types of chickens. Water intake was generally not affected by leptin, indicating that this effect was not due to general malaise. It appears that leptin can act within the central nervous sytstem of birds to decrease food intake. PMID- 10869604 TI - Capsaicin, acid and heat-evoked currents in rat trigeminal ganglion neurons: relationship to functional VR1 receptors. AB - Activation of primary trigeminal (TG) neurons by protons, capsaicin, or heat can evoke a variety of sensations, including tingling, stinging, warmth, and burning. Capsaicin and acid are trigeminal stimulants that are important in gustatory physiology. These stimuli can activate H(+)-gated ion channels and heterologously expressed VR1 receptors (vanilloid receptor 1). We have obtained evidence by using electrophysiological and pharmacological measurements on TG neurons that these three stimuli can activate many receptors, and we have determined the extent they behave similarly to VR1 receptors and H(+)-gated channels from the DEGenerin/ENaC superfamily. Whole-cell recordings from rat TG neurons revealed that protons evoked transient (Tp), sustained (Sp), and biphasic (TSp) currents. Tp currents had reversal potentials (Vr) of 24-45 mV, a pH(0.5) range from 5.5 to 6.5, and were inhibited by amiloride, suggesting the presence of functional H(+) gated channels. Sp currents were inhibited by the VR1 antagonist capsazepine, had Vr's approximately 0 mV, and had pH(0.5) = 6.4. Capsaicin also activated transient (Tc), sustained (Sc), and biphasic (TSc) currents. At pH 5.9, the sensitivity of the Sc currents increased by about a factor of 10, which may partially account for the synergistic responses of acid in foods containing capsaicin. Heating TG neurons evoked a thermally active, capsazepine-inhibitable current with threshold temperature of 43 degrees C and Vr = 5 mV that is also present in neurons activated by and protons (Sp) and capsaicin (Sc). These data suggest that TG neurons have functional receptors that behave similarly to VR1. Activation of such receptors should result in a burning sensation, whereas activation of the transient and biphasic currents should result in other taste descriptors. PMID- 10869606 TI - Letter from the editor PMID- 10869605 TI - Evidence for visceral hypersensitivity in high-anxiety rats. AB - In patients with irritable bowel syndrome, anxiety is often associated with visceral pain. Based on this information we hypothesized that rats genetically predisposed to anxiety have an increased visceral sensitivity. To test this hypothesis, visceromotor reflex recordings in response to colorectal distention were used to estimate the level of visceral stimulation in high; moderate-, and low-anxiety rats. We compared the effect of innocuous colorectal distension in rats with and without sensitized colons. In nonsensitized rats visceromotor responses were increased by colorectal distention with the greatest response in the high-anxiety Wistar-Kyoto strain. Sensitization of the colon significantly increased visceromotor responses to colorectal distention in all rat strains. In summary, our data suggested that a manifestation of a genetically determined anxiety level appeared to be abnormal neural responsiveness of the gastrointestinal tract leading to visceral hypersensitivity in high-anxiety animals. PMID- 10869607 TI - Intravesical electrical stimulation of the bladder: pro. PMID- 10869608 TI - Intravesical electrical stimulation of the bladder: con. PMID- 10869609 TI - Incidence and risk reduction of long-term outcomes: a comparison of benign prostatic hyperplasia with several other disease areas. PMID- 10869610 TI - Management of superficial bladder cancer with intravesical chemotherapy: an update. PMID- 10869611 TI - Laparoscopic ileal conduit: five-year follow-up. AB - OBJECTIVES: To report the techniques used for intracorporeal laparoscopic construction of an ileal conduit urinary diversion and long-term patient follow up after this procedure. METHODS: A 28-year-old man with cerebral palsy, a neurogenic bladder, and voiding dysfunction was referred for definitive management of his urinary tract after several episodes of pyelonephritis. A conduit urinary diversion was performed using a 5-port, transabdominal approach. An appropriate length of ileum was used for diversion, and ureterointestinal anastomoses were performed using a modified Bricker technique. All aspects of the procedure were performed intracorporeally, including isolation of conduit, bowel reanastomosis, ureteral mobilization, and ureterointestinal anastomosis. A 12-mm port site was enlarged and used as the stoma, which was constructed in routine fashion. RESULTS: Five years after surgery, this patient had normal and stable renal function, with a serum creatinine of 0.8 mg/dL. Serial imaging studies continued to reveal prompt and symmetric renal function and no evidence of obstruction or reflux. CONCLUSIONS: Laparoscopic ileal conduit construction is feasible and can provide durable results. Although technically challenging, ongoing technical refinements will make performance of reconstructive laparoscopy more widely applicable. Larger series with substantial follow-up will help illuminate the place of laparoscopic noncontinent urinary diversion in the surgical armamentarium. PMID- 10869612 TI - Laparoscopic radical cystoprostatectomy with ileal conduit performed completely intracorporeally: the initial 2 cases. AB - OBJECTIVES: To present the initial 2 patients who underwent laparoscopic radical cystoprostatectomy, bilateral pelvic lymphadenectomy, and ileal conduit urinary diversion, with the entire procedure performed exclusively by intracorporeal laparoscopic techniques. METHODS: Two male patients, 78 and 70 years old, with muscle-invasive, organ-confined, transitional cell carcinoma of the urinary bladder underwent the procedure. The entire procedure, including radical cystoprostatectomy, pelvic node dissection, isolation of the ileal loop, restoration of bowel continuity with stapled side-to-side ileoileal anastomosis, retroperitoneal transfer of the left ureter to the right side, and bilateral stented ileoureteral anastomoses were all performed exclusively by intracorporeal laparoscopic techniques. Free-hand laparoscopic suturing and in situ knot-tying techniques were used exclusively. RESULTS: The surgical time was 11.5 hours in the first patient and 10 hours in the second. The respective blood loss was 1200 mL and 1000 mL. In both patients, ambulation resumed on postoperative day 2, bowel sounds on day 3, and oral intake on day 4; the hospital stay was 6 days. Narcotic analgesia comprised 108.3 mg and 16.5 mg of morphine sulfate equivalent, respectively. Pathologic examination revealed pT4N0M0 (prostate) and pT2bN0M0 transitional cell carcinoma of the bladder with the surgical margins negative for cancer in both patients. No intraoperative or postoperative complications occurred in either patient. CONCLUSIONS: To our knowledge, this is the initial report of laparoscopic radical cystoprostatectomy with intracorporeal ileal conduit urinary diversion. We believe that with further experience and refinement in the operative technique, laparoscopic radical cystoprostatectomy with ileal conduit urinary diversion may become an attractive treatment option for selected candidates with localized muscle-invasive bladder cancer. PMID- 10869614 TI - Reply by the authors PMID- 10869613 TI - Editorial comment PMID- 10869615 TI - Outcome analysis in patients with primary necrotizing fasciitis of the male genitalia. AB - OBJECTIVES: To characterize patients with primary necrotizing fasciitis of the male genitalia (Fournier's gangrene) and to identify risk factors and prognostic variables of survival. METHODS: Fifty consecutive patients with primary necrotizing fasciitis of the male genitalia treated at our institution during a 15-year period between 1984 and 1998 were retrospectively analyzed. Of these patients, 44 (88.0%) were found to be eligible for analysis of the outcome parameters. Univariate survival analysis was performed using the Kaplan-Meier algorithm followed by multivariate analysis of statistically significant variables. Six patients (12.0%) who were severely immunocompromised were studied separately. RESULTS: Medical comorbidities were prevalent, with diabetes being the most common condition (50%). The overall mortality rate was 20% (10 of 50). Three statistically significant predictors of outcome were identified among the variables analyzed. These were the extent of the infection (P = 0.0262), the depth of the necrotizing infection (P = 0.0107), and treatment with hyperbaric oxygen (P = 0.0115). Multivariate regression analysis of these variables identified the extent of the infection (P = 0.0234) as the only statistically significant, independent predictor of outcome in the presence of other covariables. CONCLUSIONS: The involved body surface area appears to be the most important prognostic variable, with a significant impact on outcome. Given the high mortality of the disease entity and a trend toward the improved survival of patients receiving hyperbaric oxygen, this treatment form appears indicated in more severe cases. Immunocompromised patients, who frequently have an atypical and fulminant clinical course, appear to constitute a separate group with a dismal prognosis. PMID- 10869617 TI - Reply by the authors PMID- 10869616 TI - Editorial comment PMID- 10869618 TI - Urinary tract infection and patient satisfaction after flexible cystoscopy and urodynamic evaluation. AB - OBJECTIVES: To investigate the incidence of symptomatic and asymptomatic bacteriuria and to assess patient satisfaction after flexible cystoscopy (FC) and urodynamic (UD) evaluation in a prospective survey. The incidence of urinary tract infection after FC and UD studies and the use of prophylactic antibiotics are issues of debate. The tolerability and acceptance of FC and UD studies by patients have not been thoroughly documented. It would be helpful to be able to give such information to patients before performing these procedures. METHODS: A total of 215 nonconsecutive patients seen as outpatients for FC and UD studies to evaluate various indications were studied. A midstream urine sample was taken before and 48 hours after the procedures. Patients were given a questionnaire that inquired about the presence of lower urinary tract symptoms before and 48 hours after the procedures. The self-administered questionnaire included questions to assess patients' tolerance of the procedures and how it compared with their expectations. RESULTS: Of the 201 patients analyzed (FC 103, UD studies 98), 9 patients (4. 5%) developed significant bacteriuria within 48 hours of FC and UD studies. Only 2 patients with significant bacteriuria reported newly developed symptoms within 48 hours. In a subgroup of 25 patients who were given prophylactic antibiotics for various reasons, 6 (24%) reported new symptoms, although none developed significant bacteriuria. The association between patients who had preprocedure pyuria (n = 7) and the development of significant growth after the procedure (n = 6) was significant (P <0.01). In response to the patient satisfaction questionnaire, 166 (82.5%) reported that the procedure was not as bad as they expected, and 200 (99.5%) said that they would undergo the test again if necessary. CONCLUSIONS: FC and UD studies are safe, well-tolerated procedures. The addition of prophylactic antibiotics in these procedures is unnecessary, unless specific indications are present. PMID- 10869619 TI - Effects of dietary fat on the urinary risk factors of calcium stone disease. AB - OBJECTIVES: To assess the association between dietary fat and various urinary risk factors of calcium stone disease in a group of calcium stoneformers attending an outpatient stone clinic. METHODS: Mean daily fat intake from self recorded 4-day dietary food records and mean 24-hour urinary risk factors from two separate collections were evaluated in 476 patients selected randomly from an adult population attending an outpatient stone clinic for the first time. RESULTS: Mean daily total fat intake for men and women was significantly different at 105.6 and 78.1 g, respectively. Examination of the relationship between total fat intake and 24-hour urinary volume, pH, and excretions of magnesium, citrate, oxalate, calcium, and uric acid revealed no significant regressions in men and only a weak association between fat intake and urinary uric acid in women. CONCLUSIONS: Dietary fat does not have a significant effect on the urinary risk factors of calcium stone disease. PMID- 10869620 TI - Extraperitoneal laparoscopic repair of ureteropelvic junction obstruction: initial experience in 15 cases. AB - OBJECTIVES: To assess the feasibility and results of retroperitoneal laparoscopic pyeloplasty in the treatment of ureteropelvic junction obstruction. METHODS: From September 1996 to January 1999, 15 patients underwent extraperitoneal laparoscopic pyeloplasty for ureteropelvic junction obstruction. Aberrant vessels were noted in 4 patients. Dismembered pyeloplasty was performed in 7 patients and nondismembered Fenger plasty in 7 patients. Pyeloplasty was not possible in 1 patient. RESULTS: Fourteen of the 15 procedures were successfully completed. The procedure was not possible in 1 patient who had already undergone endopyelotomy repair. The mean operating time was 178 minutes (range 100 to 250), and the mean postoperative hospital stay was 4.8 days (range 1 to 14). Postoperative complications occurred in 3 patients (two hematomas and one urinoma). Radiographic assessment by intravenous urography 3 months after the procedure showed good results. CONCLUSIONS: Retroperitoneoscopy, by providing easy and rapid access to the retroperitoneal space, seems to be a valuable alternative treatment for ureteropelvic junction obstruction. PMID- 10869621 TI - The Whitaker test, a useful tool in renal grafts? AB - OBJECTIVES: To evaluate the Whitaker test, a pressure flow examination, for its prognostic value in dilated renal transplants because urologic complications, such as ureteral stenosis, are significant problems after kidney transplantation. METHODS: Twenty-five patients with obstruction of the renal transplant and subsequent percutaneous nephrostomy were evaluated with a urodynamic pressure flow test (Whitaker test) in combination with antegrade pyeloureterography. The results of the Whitaker test were related to the serum creatinine values. RESULTS: The Whitaker test demonstrated normal pressure flow (less than 15 cm H(2)O) in 7 patients, pressure flow between 15 and 25 cm H(2)O in 10, and pathologic results (greater than 25 cm H(2)O) in 8 patients. After percutaneous nephrostomy, the serum creatinine level decreased in 22 of 25 patients, although the urodynamic pressure flow revealed a significant obstruction (Whitaker test greater than 25 cm H(2)O) in only 8 patients. The sensitivity of the Whitaker test to indicate the relevance of post-renal transplant stenosis in comparison to the declining serum creatinine level after successful percutaneous nephrostomy was 79%; the specificity was 50%. CONCLUSIONS: The results of our study indicate that a decreasing creatinine level in correlation with radiologic results is the leading finding in dilation of transplanted kidneys without rejection. The Whitaker test, as a pressure flow examination, provided no additional information. PMID- 10869622 TI - Alternative or additional diagnoses on unenhanced helical computed tomography for suspected renal colic: experience with 1000 consecutive examinations. AB - OBJECTIVES: To determine the incidence and spectrum of significant alternative or additional diagnoses established or suggested on unenhanced helical computed tomography (CT) in a large series of patients with suspected renal colic. METHODS: One thousand consecutive unenhanced helical CT examinations were performed for suspected renal colic. All official CT reports were retrospectively reviewed, which was followed by review of all available relevant follow-up radiology reports. A selected image and chart review was also performed. RESULTS: Ureteral calculi were identified on 557 examinations, findings consistent with a recently passed stone were discovered on 67 examinations, and 275 CT examinations were unremarkable. An alternative or additional diagnosis was established or suggested on 101 examinations, including in 26 patients with concurrent ureteral calculi. There were 62 genitourinary and 39 nongenitourinary tract diagnoses. Eighty-seven of the diagnoses could be confirmed on retrospective image review combined with patient follow-up. There were two false-positive diagnoses for significant, alternative pathologic findings. CONCLUSIONS: A wide spectrum of significant, alternative, and additional genitourinary and nongenitourinary diagnoses can be reliably established or suggested on unenhanced helical CT performed for suspected renal colic. These abnormalities were identified in 10% of cases in this series of 1000 consecutive CT examinations. PMID- 10869623 TI - Editorial comment PMID- 10869624 TI - Incidental renal cell carcinoma-age and stage characterization and clinical implications: study of 1092 patients (1982-1997). AB - OBJECTIVES: To compare the epidemiologic, clinical, and pathologic characteristics of incidental and symptomatic renal cell carcinoma in a large series of patients, with emphasis on age distribution and its potential impact in defining groups of patients that may benefit from early detection programs. METHODS: Records of 1092 patients with renal tumors from 1982 to 1997 were reviewed. Age, clinical presentation, and pathologic stage and grade were analyzed. Special attention was given to the age distribution and its relationship to the incidental or symptomatic diagnosis. RESULTS: The overall mean age and proportion of patients older than 65 gradually increased (from 57 to 62.6 years and from 24.7% to 48.7%, respectively) from 1982 to 1997. The mean age in the incidental group rose steadily higher than in the symptomatic group. A progressive increase of incidental tumors from 13.0% in 1982 to 1983 to 59.2% in 1996 to 1997 was observed. A lower stage (74.3% versus 49.1%), grade (75.5% versus 56.9%), and percentage of metastases at presentation (10.4% versus 19.6%) were registered in the incidentally found neoplasms than in the symptomatic neoplasms. Eighty-two (80.4%) of 102 patients who underwent conservative surgery had incidental renal cell carcinoma. CONCLUSIONS: Our data confirm a rapid and dramatic change in the epidemiologic and clinical characteristics of renal cancer, with an increasing number of incidentally found tumors presenting with lower stage, grade, and percentage of metastases. An unexpected but significantly higher rate of renal neoplasms was observed in older patients. The stage, grade, and patient age observed in our series of incidentally found tumors raises the question of whether to leave the current diagnostic approach unaltered, thus benefiting a subgroup of patients with clinically unrecognized and possibly indolent renal cell carcinoma, or to extend early detection programs to younger patients with potentially more aggressive tumors. PMID- 10869625 TI - Retroperitoneal laparoscopic nephrectomy is safe and effective in obese patients: a comparative study of 55 procedures. AB - OBJECTIVES: To compare the results of retroperitoneal laparoscopic nephrectomy (RLN) in obese and nonobese patients, because various open surgical procedures have been reported to result in higher morbidity in obese patients. METHODS: Forty-eight consecutive patients underwent 55 RLNs in one center by one surgeon. Twenty-two patients were renal transplant recipients and underwent a total of 29 RLNs of the native kidney. Eight patients (9 procedures) were obese (body mass index 30 or more). The duration of the procedure, intraoperative and postoperative complications, and length of stay were compared between the obese and nonobese patients. RESULTS: The median operative duration was 100 and 70 minutes in the obese and nonobese patients, respectively. Three intraoperative complications occurred in nonobese patients. One postoperative complication (12. 5%) occurred in the obese patients; four (15.6%) occurred the nonobese patients. The median length of stay was 4 days for the obese and 3 days for the nonobese patients. The complication rate and postoperative length of stay were not significantly different between the two groups. CONCLUSIONS: RLN in obese patients was not associated with higher morbidity or longer hospitalization than in nonobese patients. We believe that RLN should be proposed to such patients when nephrectomy of a small nonfunctional kidney is indicated. PMID- 10869626 TI - Outcome of surgery in cystic renal cell carcinoma. AB - OBJECTIVES: To review cases of cystic renal cell carcinoma treated surgically at our institution and define their clinical and histopathologic features. METHODS: Between 1986 and 1998, 21 patients with cystic renal cell carcinoma were treated surgically. Cystic renal cell carcinoma was categorized using Hartman's classification. RESULTS: Histopathologic examination demonstrated cystic necrosis in 11 patients, multilocular cystic renal cell carcinoma in 9, and unilocular cystic renal cell carcinoma in 1 patient. Tumors were incidentally found during an evaluation of unrelated disease or a general health checkup in 14 patients (67%). The mean tumor size was 5.6 cm (range 0.5 to 12) for cystic necrosis and 5.4 cm (range 2 to 9) for multilocular cystic renal cell carcinoma. All 9 cases of multilocular cystic renal cell carcinoma were of the clear cell type and tumor grade 1. The mean follow-up period was 65 months (range 9 to 141). The 5-year disease-specific survival rates for multilocular cystic renal cell carcinoma and cystic necrosis were 100% and 80%, respectively. CONCLUSIONS: The prognosis for patients with cystic renal cell carcinoma is better than that for patients with solid tumors. In particular, the prognosis of multilocular cystic renal cell carcinoma is excellent. Multilocular cystic renal cell carcinoma represents a distinct subtype of renal cell carcinoma that can be completely cured by surgery. PMID- 10869627 TI - Cross-sectional study of nocturia in both sexes: analysis of a voluntary health screening project. AB - OBJECTIVES: To assess the prevalence of nocturia and its impact on the quality of life in both sexes by analyzing almost 2500 individuals participating in a health survey. METHODS: During a 12-month period, we included an incontinence questionnaire, which was largely based on the Bristol female lower urinary tract symptoms questionnaire, in the voluntary health examinations in the area of Vienna. In parallel, we recorded the medical history, concurrent medical therapy, physical examination findings, sociodemographic parameters, and blood laboratory study results. RESULTS: The data of 1247 women (age 49.8 +/- 13.5 years) and 1221 men (age 48.5 +/- 11.9 years) were analyzed. The percentage of individuals with nocturia of two or more times increased constantly with age: less than 30 years, 3.1% of women and 3.4% of men; 30 to 59 years, 7.2% of women and 5. 7% of men; and 60 years old or older, 26.7% of women and 32.4% of men. Age-adjusted extrapolation to the general population (older than 20 years) currently living in Austria yielded that 10.8% of men and 11.8% of women have nocturia of two or more times. Overall, 66. 9% of women and 62.2% of men reported a negative impact of nocturia on their quality of life. The correlation was close between the degree of nocturia with the quality-of-life impairment in both sexes. Several voiding symptoms correlated significantly (P <0.001) with nocturia. CONCLUSIONS: Nocturia is almost equally present in both sexes, and the incidence and severity increase constantly from early adolescence to senescence. Approximately 10% of the general population (older than 20 years) have nocturia of two or more times, which impairs the quality of life in two thirds. PMID- 10869629 TI - Editorial comment PMID- 10869628 TI - Transurethral hot-water balloon thermoablation for benign prostatic hyperplasia: patient tolerance and pathologic findings. AB - OBJECTIVES: To determine the patient tolerance and thermal ablation pattern in human prostatic tissue after treatment with a hot water, catheter-based system. METHODS: Twenty-seven men scheduled for surgery for symptomatic benign prostatic hyperplasia or adenocarcinoma of the prostate underwent water-induced thermotherapy. The patients were randomly assigned to one of four treatment groups. Lidocaine gel was the sole means of pain control. The patients and an observer recorded patient discomfort during therapy. A Foley catheter was left in place until surgery (n = 13) or successful voiding (n = 14). Prostates were subsequently enucleated or removed, whole mounted, and examined. RESULTS: Patients reported mild treatment discomfort, the level of which did not correlate with the extent of necrosis, balloon diameter, or water temperature (all P >0. 05). Distal penile burning was the most commonly reported discomfort. All 14 patients successfully voided within 12 days of treatment. Prostates were enucleated (n = 24) or removed (n = 3) at a mean of 27 days (range 4 to 120) after thermotherapy, except for a single adenectomy 17 months after therapy. Pathologic findings included periurethral hemorrhagic necrosis, with focal or extensive urothelial denudation and mild inflammation. The mean maximal depth of necrosis from the urethral lumen was 7, 9, 10.33, and 11 mm in groups 1, 2, 3, and 4, respectively. The extent of necrosis was similar in all groups (P = 0.11), regardless of the water temperature; conversely, the balloon diameter correlated with the depth of necrosis (P = 0.024). CONCLUSIONS: This system of tissue ablation appears to be well tolerated, and it produced consistent pathologic results. PMID- 10869630 TI - Evaluation of the effect of spinal cord injury on serum PSA levels. AB - OBJECTIVES: Prostatic structure and secretory activity are thought to be influenced by autonomic innervation of the prostate. Prostatic denervation is especially likely in patients with spinal cord injury (SCI) at the level of the cauda equina or the conus medullaris, where the peripheral nerve supply to the prostate may be specifically damaged. This may result in changes in serum prostate-specific antigen (PSA) levels, either directly or indirectly. Therefore, we measured serum PSA levels and also studied the influence of factors such as age, catheterization, duration of SCI, urinary tract infection, and history of cystitis on serum PSA values in men with SCI. METHODS: Serum PSA levels were determined in 79 men with SCI (age older than 40 years) using banked sera by the Abbott MEIA PSA assay. Variables such as age, catheterization, duration of SCI, urine culture results, and history of cystitis were obtained from a review of patient records. Comparisons were made with a randomly selected, non-SCI control population of 501 men, 40 to 89 years old, who underwent serum PSA determination at our institution. Statistical comparisons were performed using the Mann-Whitney U test (nonparametric), since the populations were not normally distributed. Multivariate logistic regression analysis was used to assess the correlation between the various factors and the serum PSA levels in men with SCI. RESULTS: No statistically significant differences were found in the median serum PSA values between the SCI group and the non-SCI control population. The age-specific PSA values obtained in the SCI group were also comparable to those reported for the general population at large. Age (P <0.03) and the presence of a catheter (P <0.0002) were the only two factors that were correlated with higher serum PSA values in the SCI group by regression analysis. CONCLUSIONS: Men with SCI tended to have serum PSA value distributions that were similar to those of the general population. However, those in the SCI group who had indwelling catheters were more likely to have higher PSA values at baseline, as were older men with SCI. PMID- 10869631 TI - Race is not an independent predictor of biochemical recurrence after radical prostatectomy in an equal access medical center. AB - OBJECTIVES: To compare the racial differences in clinical and pathologic features between black and white men who underwent radical prostatectomy (RP) in an equal access health care center and to determine whether race is an independent predictor of biochemical recurrence. METHODS: A retrospective survey of 273 patients (125 black, 148 white) who underwent RP at the West Los Angeles Veterans Affairs Medical Center between 1991 and 1999 was undertaken. Patients were analyzed for racial differences in age at diagnosis, clinical stage, preoperative serum prostate-specific antigen (PSA), and Gleason score of the prostate biopsy specimens. Surgical specimens were studied to determine pathologic stage, Gleason score, incidence of seminal vesicle invasion, positive surgical margins, capsular penetration, and pelvic lymph node involvement. Patients were followed for PSA recurrence (greater than 0.2 ng/mL). Multivariate analysis was used to determine the clinical and pathologic variables that were significant in predicting biochemical recurrence after RP and to determine whether race was an independent predictor of biochemical failure. RESULTS: No significant differences were found between black and white men in the preoperative factors (clinical stage, age at diagnosis, biopsy Gleason score, and serum PSA) or in the pathologic features of the RP specimens (Gleason score, pathologic stage, incidence of positive surgical margins, capsular penetration, seminal vesicle invasion, or lymph node involvement). In addition, no differences were found between black and white men in the PSA recurrence rates after RP using Kaplan-Meier survival curves (P = 0.651). Multivariate analysis revealed that serum PSA (P = 0.010), biopsy Gleason score (P = 0. 003), younger age (P = 0.010), surgical Gleason score (P = 0.005), and lymph node involvement (P = 0.022) were all independent predictors of biochemical recurrence. Race was not a significant predictor of biochemical failure in multivariate analysis (P = 0. 199). CONCLUSIONS: In an equal access medical care facility, no differences were evident between black and white men in the preoperative clinical factors or the pathologic features of the RP specimens. In addition, no differences were observed in the PSA recurrence rates after RP. Serum PSA, biopsy Gleason score, younger age, surgical Gleason score, and lymph node involvement were all independent predictors of biochemical recurrence. Race was not an independent predictor of biochemical recurrence. PMID- 10869632 TI - No difference in biochemical failure rates with or without pelvic lymph node dissection during radical prostatectomy in low-risk patients. AB - OBJECTIVES: To detect the short-term differences in biochemical relapse-free rates between patients with and without pelvic lymph node dissection (PLND). Recently, a trend has begun to omit PLND in patients undergoing radical prostatectomy considered at low risk of pelvic lymph node metastases. METHODS: The records of 1152 consecutive radical prostatectomy cases were reviewed. A total of 575 patients with favorable tumor characteristics (prostate-specific antigen [PSA] 10 ng/mL or less, Gleason score 6 or less, and clinical Stage T1 or T2) who were not receiving adjuvant or neoadjuvant therapy were divided into two groups according to whether PLND was performed (PLND group, n = 372) or omitted (no PLND group, n = 203). Proportional hazards were used to analyze the effect of age, race, family history, stage, biopsy Gleason score, initial PSA, PLND, and pathologic findings on the likelihood of biochemical failure. Biochemical failure free survival for each group was estimated by Kaplan-Meier analysis. The mean follow-up was 38 months (range 1 to 141). RESULTS: The actuarial 4-year biochemical relapse-free rate for the PLND versus no PLND groups was 91% and 97%, respectively (P = 0.16). On multivariate analysis, PLND was not an independent predictor of outcome (P = 0.24). CONCLUSIONS: The results of our study indicate that the omission of PLND in patients with favorable tumor characteristics does not adversely affect biochemical relapse rates. PMID- 10869633 TI - Risk factors for vesicourethral anastomotic stricture after radical prostatectomy. AB - OBJECTIVES: Preoperative comorbidities associated with microvascular disease may contribute to the development of bladder neck contracture (BNC) by alteration of anastomotic healing. We investigated potential risk factors for development of BNC after radical prostatectomy (RP) and reviewed management of this complication. METHODS: A retrospective review of 467 consecutive patients (mean age 63.2 years) undergoing RP between 1991 and 1999 was performed. In all cases, the bladder neck was tailored to 20 to 22F in a racket handle fashion. After mucosal eversion of the reconstructed bladder neck, a mucosa-to-mucosa vesicourethral anastomosis was created over an 18 to 22F catheter using 4 to 6 anastomotic sutures. The relationship between comorbidities identified preoperatively by patient interview and medical record review (coronary artery disease [CAD], diabetes mellitus [DM], hypertension [HTN], cerebral vascular accident, chronic obstructive pulmonary disease, and smoking history) and the incidence of BNC was determined. Risk factors including prior transurethral prostatectomy (TURP), estimated blood loss (EBL), and operative time (OR time) were also evaluated. Factors were evaluated for their ability to predict BNC using both univariate and multivariate analysis. Treatment results for BNC were also assessed. RESULTS: A total of 52 (11.1%) patients developed BNC. Current cigarette smoking resulted in a significantly higher (26%) rate of BNC (P <0.001). The BNC rate was also increased in patients with CAD (26%, P <0.001), HTN (19%, P = 0.015), and DM (21%, P = 0.030). Average OR time was longer (271 versus 249 minutes, P = 0.025) and EBL was greater (1639 versus 1092 mL, P <0.001) in patients developing a BNC. In multivariate analysis, current cigarette smoking was the strongest predictor of BNC and independent of other factors (P <0.001). BNC was not related to prior TURP, type of anastomotic suture used, size of catheter, or duration of catheterization. Patients were treated with transurethral dilation (73%) or transurethral incision (27%) and 58% responded to the initial treatment. No patient became incontinent as a result of the treatment for BNC.Conclusions. Several comorbidities associated with microvascular disease are significant risk factors for development of BNC after RP. Current cigarette smoking in particular is a strong predictor. Transurethral dilation and transurethral incision are equally effective as initial treatment of BNC. PMID- 10869634 TI - Using outcome data and patient satisfaction surveys to develop policies regarding minimum length of hospitalization after radical prostatectomy. AB - OBJECTIVES: Changes in health care economics have prompted new clinical pathways for radical prostatectomy to reduce length of hospitalization after surgery to 1 day. We evaluated satisfaction, outcomes, and short-term morbidity in 187 consecutive patients with overnight hospitalization after radical retropubic prostatectomy (RRP). METHODS: In 1995, we initiated a critical pathway for RRP that included epidural anesthesia with or without spinal anesthesia and postoperative methadone, acetaminophen, and ibuprofen for pain control. Patients were discharged when they were afebrile, tolerating a regular diet, ambulating without assistance, and using oral medications for analgesia. An 18-item satisfaction survey was mailed to each patient 3 weeks after discharge. Responses to the postoperative survey, morbidity, blood loss, and use of transfusions were recorded. RESULTS: Of 252 patients who underwent RRP, 187 (74. 2%) were discharged 1 day after surgery. The mean age of patients was 61.4 years (range 42 to 73). A pelvic lymphadenectomy was performed in addition to the RRP in 32 men (17%). Epidural anesthesia with or without spinal anesthesia was used for all but 3 patients. The mean estimated blood loss was 1166 mL, and 24 patients (12.8%) required transfusion, with a mean of 1.9 U (range 1 to 6) of packed red blood cells. The postoperative complication rate was 11. 8%, of which 2.1% (n = 4) were definitely or probably related to our protocol. These complications included clot retention (n = 2), gastrointestinal bleeding (n = 1), and spinal headache (n = 1). Three of 187 patients were readmitted to the hospital within 30 days but only one (0.5%) required admission because of our protocol. The survey response rate was 91.4%. No patient was dissatisfied with his overall care, and only 10.5% of patients would have preferred to stay in the hospital longer. CONCLUSIONS: One day hospitalization after RRP is associated with minimal postoperative morbidity and high patient satisfaction. Similar data are needed for RRP from other centers before policy decisions regarding the length of stay after this procedure are made. PMID- 10869636 TI - Reply by the authors PMID- 10869635 TI - Editorial comment PMID- 10869637 TI - Radical prostatectomy: geographic and demographic variation. AB - OBJECTIVES: Previous reports have documented a geographic variation in the use of radical prostatectomy. We examined whether this phenomenon can be explained by factors other than geography alone. METHODS: This study was based on the data from nine geographic regions of the Surveillance, Epidemiology, and End Results (SEER) program for the years 1983 through 1994. Patients with localized or regional prostate cancer were included in the analysis. Logistic regression analysis was used to investigate the influence of geographic and demographic factors on the use of radical prostatectomy. The squared multiple correlation coefficient R(2) was used to measure the proportion of variation in the selection of radical prostatectomy explained by each factor of interest. RESULTS: As previously reported, the use of radical prostatectomy was significantly associated with geographic location; the degree of geographic variation varied as a function of age and was most dramatic in the youngest (younger than 45 years) and the oldest (75 years or older) groups. Overall, however, geography explained less than 2% of the total variation in the use of radical prostatectomy. Age was the most important factor that influenced the use of radical prostatectomy. CONCLUSIONS: Geography explains only a small proportion of the variation in the use of radical prostatectomy. In fact, of the factors examined, only age appeared to meaningfully explain the variation in the use of radical prostatectomy. Overall, our ability to explain the variation in the use of radical prostatectomy remains meager, and new factors must be identified if we are to better understand how patients and physicians make clinical decisions. PMID- 10869638 TI - Complications after radical retropubic prostatectomy in the medicare population. AB - OBJECTIVES: Radical prostatectomy is the standard of care for the treatment of clinically localized prostate cancer in the appropriate patient. However, the morbidity associated with this procedure remains controversial, since complications from centers of excellence are low but nationwide surveys have reported a much higher risk of complications. This study reports the complication rates after radical retropubic prostatectomy (RRP) for men in the Medicare population. METHODS: All men in the Medicare population who underwent RRP in 1991 were identified. All inpatient, outpatient, and physician (Part B) Medicare claims for these men for 1991 to 1993 were then analyzed to determine outcomes. Procedures performed for complications resulting from RRP were recorded, as were the diagnosis codes that may have heralded a complication after RRP. RESULTS: In 1991, 25,651 men in the Medicare population underwent RRP. The mean age of these men was 70.5 years. Procedures for the relief of bladder outlet obstruction or urethral strictures after RRP occurred in 19.5% of these men. A penile prosthesis was implanted in 718 men (2.8%) after prostatectomy, and 593 men (2.3%) had an artificial urinary sphincter placed after prostatectomy. A diagnosis of urinary incontinence was reported in 5573 men (21.7%) after radical prostatectomy, but only 2025 of these men (7.9%) continued to carry this diagnosis more than 1 year after prostatectomy. A diagnosis of erectile dysfunction was reported in 5510 men (21.5%) after radical prostatectomy, but only 3276 of these men (12.8%) continued to carry this diagnosis more than 1 year after surgery. CONCLUSIONS: A review of a large, nationwide, heterogenous cohort of men revealed a morbidity rate that is slightly higher than that reported by major centers that perform large numbers of radical retropubic prostatectomies but is lower than complication rates obtained by patient surveys. The limitations of claim information in determining patient outcomes, however, must be considered when evaluating these data. PMID- 10869639 TI - Extraperitoneal laparoscopic bladder neck suspension using hernia mesh and tacker. AB - OBJECTIVES: To report our initial experience with extraperitoneal bladder neck suspension for female stress incontinence due to urethral hypermobility. METHODS: Between September 1996 and September 1999, 35 patients (mean age 49.5 years) underwent extraperitoneal bladder neck suspension at our institution. An extraperitoneal space was created by a trocar-mounted balloon device, and suspension was created using a 5-mm endoscopic hernia stapler and polypropylene mesh. RESULTS: The mean operative time was 39.5 minutes. In 2 patients, the bladder was inadvertently perforated during the bladder neck dissection. The perforation was repaired by laparoscopic suture ligation. The mean urethral catheterization and hospitalization time was 2.1 and 2.3 days, respectively. Urethral recatheterization because of temporary urinary retention was required in 11.4% of the patients. Symptoms of bladder instability were experienced by 13.5% of the patients in the early postoperative period. A total of 28 patients (80.0%) reported that they were totally dry after a mean of 23.2 months. CONCLUSIONS: Extraperitoneal bladder neck suspension using hernia mesh and a stapler seems to be an effective and safe procedure, with a shorter operative time, in selected patient groups. PMID- 10869640 TI - Cavernous and systemic testosterone levels in different phases of human penile erection. AB - OBJECTIVES: To examine changes in testosterone levels in the cavernous and peripheral blood during different phases of erection because, although the determination of systemic testosterone levels has been well established in the diagnostic workup of erectile dysfunction, the exact role of testosterone in adult male sexual function remains unclear. METHODS: Blood samples were drawn simultaneously from the corpus cavernosum and the cubital vein of 54 healthy and normally potent volunteers during four different stages of the cavernous erectile tissue (flaccidity, tumescence, rigidity, and detumescence). Penile erections were induced by audiovisual and tactile stimulation, and testosterone levels were determined by radioimmunoassay. RESULTS: The mean testosterone level in the corpus cavernosum plasma during the flaccid state was 2.9 +/- 1.2 ng/mL. During tumescence and rigidity, the testosterone levels in the cavernous blood significantly increased, to 4.3 +/- 1.3 ng/mL and 4. 4 +/- 1.4 ng/mL, respectively. During detumescence, the cavernous testosterone levels dropped to 3.5 +/- 1.4 ng/mL. The changes in the testosterone levels in the peripheral plasma were less pronounced. A significant increase was also found in the peripheral testosterone levels from flaccidity (4.1 +/- 1.1 ng/mL) to tumescence (4.4 +/- 1. 4 ng/mL). No further increase in testosterone occurred during the phase of rigidity. From rigidity to detumescence, the peripheral testosterone levels dropped to 4.1 +/- 1.2 ng/mL. CONCLUSIONS: Penile erection was found to be accompanied by a significant increase in cavernous and systemic testosterone plasma levels. The estimated difference between the systemic and cavernous testosterone levels during penile flaccidity, when blood flow through the cavernous body is minimized, might be a diagnostic tool to evaluate the amount of bioavailable testosterone and the activity of testosterone receptors in the corpus cavernosum smooth musculature. PMID- 10869641 TI - Efficacy and safety of fixed-dose and dose-optimization regimens of sublingual apomorphine versus placebo in men with erectile dysfunction. The Apomorphine Study Group. AB - OBJECTIVES: A sublingual (SL) formulation of apomorphine has been developed and found effective in penile erectile dysfunction (ED). This study assessed the efficacy and safety of several doses of apomorphine SL in a dose-optimization schedule compared with placebo. METHODS: In this 8-week, multicenter, double blind clinical trial, 569 patients were randomized to four groups: a dose optimization group in which patients began with 2 mg, increased or decreased the dosage as needed for 4 weeks, and thereafter maintained an optimal dose for 4 weeks; two fixed-dose groups of either 5 or 6 mg; and a placebo group. Efficacy was assessed by patient and partner responses to home-use questionnaires about sexual function and activity and by responses to the International Index of Erectile Function and the Brief Sexual Function Inventory. RESULTS: In all apomorphine SL groups, a significantly higher percentage of patients compared with the placebo group achieved and maintained an erection firm enough for intercourse (48% to 53% versus 35% for placebo, P < or =0.001) and a significantly higher percentage of attempts resulted in intercourse (45% to 51% versus 33%, P < or =0.001). The responses to the questionnaires completed by the patients and partners were similar. Apomorphine SL was well tolerated; nausea, the most common side effect, was dose related and diminished substantially during the second 4-week period at all doses. The dose-optimization schedule resulted in fewer adverse events without impacting efficacy. CONCLUSIONS: Apomorphine SL is an effective and safe treatment for ED, with 2 and 4 mg providing the most acceptable therapeutic index. PMID- 10869642 TI - Effect of adult microsurgical varicocelectomy on testicular volume. AB - OBJECTIVES: To re-examine the potential influence of varicocelectomy on testicular volume using scrotal ultrasonography, because it has been reported that total testicular volume (assessed by physical examination) increases after adult varicocele ligation. METHODS: A retrospective review of the testicular volume and semen parameters of 61 men who underwent microsurgical varicocelectomy between 1996 and 1998 was performed. Ultrasound-derived testicular volumes and total motile sperm counts were compared before varicocelectomy and at a mean of 7.2 months postoperatively. RESULTS: Bilateral varicocelectomy was performed in 22 men; 39 men underwent a left-sided procedure only. Overall, no significant change was found in the mean total testicular volume after varicocelectomy compared with preoperatively (24.0 versus 23.9 mL, respectively; P = 0.74). Similarly, the testicular volumes did not change significantly after left or bilateral varicocelectomy (P >0.05). Overall, the mean total motile sperm count increased significantly after varicocelectomy (17. 9 to 25.4, P = 0.05). CONCLUSIONS: This was the first study to examine the effect of adult varicocelectomy on testicular volume using ultrasound-derived measurements of volume. Unlike previous findings, our data suggest that although adult varicocelectomy improves semen quality in most infertile men, it does not result in a significant increase in testicular volume. PMID- 10869643 TI - Importance of identifying the inconspicuous penis: prevention of circumcision complications. AB - OBJECTIVES: To report the incidence of inconspicuous penis in those referred to a pediatric urologist for circumcision or revision of circumcision. METHODS: We performed a chart review of 129 boys referred to one pediatric urologist during a 3-year period for circumcision. RESULTS: Of 129 patients, 30 (23%) had evidence of an inconspicuous penis on the physical examination. Eight (9%) of 94 patients referred for the initial circumcision and 22 (63%) of 35 patients referred for a circumcision revision had evidence of an inconspicuous penis on physical examination. CONCLUSIONS: Inconspicuous penis is an umbrella term used to categorize several anatomic anomalies that occur in newborn boys. The incidence of inconspicuous penis at physical examination in children referred for circumcision revision is remarkable. Circumcision is best delayed in these newborn patients until a later time. PMID- 10869644 TI - Editorial comment PMID- 10869646 TI - A new technique for securing a foley catheter. AB - We describe a new technique to secure a urethral catheter using a horizontal drain tube stabilizer. This device is reliable, inexpensive, and more comfortable for patients than either adhesive tape or leg straps. PMID- 10869645 TI - Early orchiopexy: prepubertal intratubular germ cell neoplasia and fertility outcome. AB - OBJECTIVES: To investigate the prepubertal prevalence of intratubular germ cell neoplasia of the unclassified type (ITGCNU) and its significance as a predictor of testicular cancer and to evaluate the effect of early orchiopexy (at younger than 2 years of age) on subsequent fertility of patients with bilateral cryptorchidism. METHODS: Testicular biopsies (n = 660) from 440 prepubertal patients with cryptorchidism who underwent orchiopexy between January 1, 1970 and December 31, 1979 were evaluated for ITGCNU using placental-like alkaline phosphatase (PLAP) antibody. The clinical outcome in 15 patients with PLAP positive germ cells was evaluated in 1997. In addition, the effect of age at surgery on the fertility of patients with bilateral cryptorchidism was assessed by clinical follow-up until 1997 and was correlated with the histologic data at orchiopexy. RESULTS: PLAP-positive germ cells morphologically identical with adult ITGCNU were found in the biopsies of 22 patients (5%). After more than two decades, none of the 15 patients with successful follow-up developed testicular cancer. The fertility outcome in the patients with bilateral cryptorchidism correlated with the number of spermatogonia at orchiopexy (P = 0.018), but correlated inversely with age at orchiopexy (P = 0.021). CONCLUSIONS: PLAP positive germ cells in prepubertal testicular biopsy specimens are not necessarily precursors of testicular cancer after orchiopexy. In addition, our data support the idea that early orchiopexy may be beneficial in preventing infertility. PMID- 10869647 TI - Tube gastrostomy after radical cystectomy and urinary diversion: surgical technique and experience in 709 patients. AB - We describe our surgical technique of tube gastrostomy and report our experience with 709 patients who underwent cystectomy and urinary diversion with gastrostomy tube placement from January 1988 to December 1997. This modified Stamm technique provides an effective means of gastric decompression without the discomfort associated with nasogastric decompression, is associated with a low complication rate (0.05%), and may be considered as the procedure of choice when gastric drainage is required after radical cystectomy and lower urinary tract reconstruction. PMID- 10869648 TI - Hand-assisted laparoscopic bilateral nephrectomy. AB - Open bilateral nephrectomy in renal transplant patients may be indicated for various reasons and is associated with a significant rate of morbidity and mortality. Laparoscopic bilateral nephrectomy may favorably influence postoperative recovery but is technically difficult. This case report is the first description of hand-assisted laparoscopic bilateral nephrectomy. We believe this technique significantly shortens the operative time, increases the safety of the procedure, and assures the patient the benefits of minimal invasive surgery in terms of postoperative pain and recovery. PMID- 10869649 TI - Fournier's gangrene caused by Candida species as the primary organism. AB - Fournier's gangrene is a rare entity caused by polymicrobial aerobic and anaerobic bacteria. We report a case of Fournier's gangrene caused by Candida as the primary organism. A 65-year-old man presented with perineal soft-tissue infections. He underwent surgical debridement and suprapubic cystostomy with both antifungal and antimicrobial therapy. The histopathologic examination revealed necrotizing fasciitis with Candida species as the sole initial pathogen. The case suggests that primary fungal pathogens should be considered as a causative organism of Fournier's gangrene. PMID- 10869650 TI - Bazex syndrome: acrokeratosis paraneoplastica in association with simultaneous multiple genitourinary tumors. AB - We report a patient with Bazex (Bazex-Dupre-Christol) syndrome in association with multifocal basal cell carcinoma, epidermoid carcinoma of the lung, adenocarcinoma of the prostate, and possibly undifferentiated carcinoma of the bladder. To our knowledge, this is the first report of a patient with Bazex syndrome that consisted of four tumors, including two genitourinary tumors. PMID- 10869651 TI - Cushing syndrome as the presenting feature of metastatic Leydig cell tumor of the testis. AB - We report a patient with a history of orchiectomy for Leydig cell tumor of the testis who developed Cushing syndrome. This syndrome was due to ectopic production of cortisol and was the primary feature of tumor recurrence. PMID- 10869652 TI - Sclerotherapy of hemangioma of the glans penis. AB - Penile hemangiomas are very rare, small lesions. Possible therapies include surgical excision, electrofulguration, cryotherapy, and sclerotherapy. We describe an extremely rare case of a large hemangioma of the glans that was treated by means of repeated injections of 2% polidocanol under local anesthesia (5% lidocaine-prilocaine cream). No complications were observed after the operation, and a satisfactory aesthetic result was obtained. No relapse was noted 18 months after the treatment. A therapeutic reference standard for the treatment of penile hemangiomas is still lacking because of the rarity of the disease. Sclerotherapy proved to be an effective, low-cost, and easy-to-perform procedure. Moreover, it is repeatable in case of failure. PMID- 10869653 TI - Ureteral obstruction after kidney transplantation secondary to bone metaplasia. AB - We report a case of ureteral obstruction after kidney transplantation caused by localized bone metaplasia in the donor ureter. Surgical treatment consisted of removal of the involved ureteral segment, which was located 3 cm above the bladder and creation of a spatulated end-to-end anastomosis. Although bone metaplasia has been observed in the ureteral wall of some animal species secondary to experimental ischemia and microtrauma, it is exceedingly rare in humans and has never before been documented after kidney transplantation. PMID- 10869654 TI - Mucoepidermoid penile carcinoma: clinical, histologic, and immunohistochemical characterization of an uncommon neoplasm. AB - We report a case of a mucoepidermoid penile carcinoma. The specimen was studied by immunohistochemistry and DNA cytometry. Mucoepidermoid and adenosquamous penile carcinoma are exceedingly rare variants of penile cancer, with very little follow-up data available. To evaluate the possible prognostic significance of these differentiation patterns, it is necessary to document more cases of this unusual tumor entity. PMID- 10869655 TI - Indium-111 capromab pendetide (ProstaScint) uptake in neurofibromatosis. AB - We report on the demonstration of nonspecific accumulation of indium-111 capromab pendetide (ProstaScint) in neurofibromas in a patient with a rising prostate specific antigen level after prostatectomy. Although the immunoconjugate is targeted to a specific membrane antigen present in human prostate carcinoma, nonspecific uptake can occur in other entities. Indium-111 capromab pendetide scintigraphy is a valuable diagnostic tool that should always be interpreted within the context of the clinical history and the laboratory and radiologic findings to avoid misinterpretation. PMID- 10869656 TI - Granular cell tumor of the penis in a 5-year-old boy. AB - We evaluate a rare case of a 5-year-old boy with granular cell tumor of the penis, located on the coronal margin. We discuss the anatomic distribution of granular cell tumors, noting the marked female predominance of genital involvement and reviewing the distribution of granular cell tumors by site in a pediatric series. We also discuss clinical implications, management, and histogenesis. PMID- 10869657 TI - Unusual cause of early preeclampsia: bladder paraganglioma. AB - Ten to twenty percent of paragangliomas occur at extra-adrenal locations and less than 1% are at the urinary bladder. The most common presenting symptom of bladder paraganglioma is hypertensive attacks precipitated by micturition and hematuria. Paraganglioma of the urinary bladder occurring at pregnancy is extremely rare. We present a case of bladder paraganglioma as an unusual cause of early preeclampsia. After termination of the pregnancy, surgical resection was performed and the histopathologic diagnosis of paraganglioma confirmed. At 24 months of follow-up the patient felt well and was normotensive without any foci of paraganglioma. Although rare, paraganglioma must be considered in the differential diagnosis of early preeclampsia. PMID- 10869658 TI - Biosafety of in vivo adenovirus-p53 intravesical administration in mice. AB - OBJECTIVES: To evaluate the biosafety and in vivo biodistribution of intravesical instillation of an adenovirus that contains human p53 gene. Mutations of p53, which are found in as many as 40% of transitional cell carcinomas, are associated with a poor prognosis and resistance to chemotherapy and radiation therapy. Restoration of wild-type p53 status by means of adenoviral-mediated therapy may enhance apoptosis and improve the response to therapy, but the issues of biosafety and toxicity have not yet been addressed. METHODS: Adenovirus-p53 (1 x 10(8), 1 x 10(9), and 5 x 10(9) pfu/mL) and luciferase reporter gene (5 x 10(9)) were instilled into the bladders of anesthetized female BALB/c mice. The mice were killed on days 1, 3, 6, and 13, and representative samples of the bladder, ureter, kidney, adrenal gland, ovary, liver, heart, and lung were removed for histologic evaluation. RESULTS: No histologic signs of toxicity were found. The hematologic and biochemical profiles of the mice were normal, with the exception of a transient elevation in liver function tests on day 1 in the three treatment groups. CONCLUSIONS: Intravesical instillation of adenovirus-p53 was well tolerated; the bladder urothelium appeared to prevent systemic dissemination. The results of these experiments support the safety of intravesical gene transfer by intravesical instillation. PMID- 10869659 TI - Expression of senescence-associated beta-galactosidase in enlarged prostates from men with benign prostatic hyperplasia. AB - OBJECTIVES: Cellular senescence is a unique cellular response pathway thought to be closely associated with the aging process. The senescent phenotype is characterized by the loss of a cell's ability to respond to proliferative and apoptotic stimuli even while normal metabolic activity and vitality is maintained. Recently, a novel biomarker, senescent-associated beta-galactosidase (SA-beta-gal), was found to identify cells with the senescent phenotype. In the present study, we examined whether human prostatic epithelial cells adopt a senescence-associated phenotype after prolonged culture and analyzed a series of human benign prostatic hyperplasia (BPH) specimens to determine whether the cellular senescence process might be a factor in the development of BPH. METHODS: A primary culture of epithelial cells was established from the normal tissue of the peripheral zone of a radical prostatectomy specimen and was serially passaged until senescence. Forty-three human prostate specimens were obtained subsequent to radical prostatectomy or transrectal ultrasound-guided biopsy. The cultured cells and tissue specimens were histochemically stained to reveal the expression of SA-beta-gal, the cellular senescence biomarker. RESULTS: As has been reported for other types of cultured cells, human prostatic epithelial cells demonstrated widespread expression of the cellular senescence marker, SA-beta-gal, on prolonged culture. In our survey of hypertrophied human prostate tissues, 17 specimens (40%) of the 43 analyzed demonstrated positive staining for SA-beta gal. In these tissues, SA-beta-gal expression was noted only in the epithelial cells. No statistical correlation (P = 0.42) between the chronologic age of the patient donor and SA-beta-gal expression was found. However, a high prostate weight (greater than 55 g) was found to correlate strongly with the expression of the SA-beta-gal biomarker (P = 0. 0001). CONCLUSIONS: Cultured prostatic epithelial cells expressed SA-beta-gal on reaching replicative senescence in vitro. The survey of human BPH specimens for the senescent marker showed that prostatic epithelial cells in patients with BPH with more advanced enlargement of the prostate (greater than 55 g prostate weight) expressed SA-beta-gal, and the prostates from patients with BPH that weighed less than 55 g tended to lack senescent epithelial cells. On the basis of these results, we propose that the accumulation of senescent epithelial cells may play a role in the development of the prostatic enlargement associated with BPH. PMID- 10869660 TI - Laser-induced hyperthermia in rat prostate cancer: role of site of tumor implantation. AB - OBJECTIVES: To investigate the importance of the site of tumor implantation on the treatment response to laser-induced hyperthermia (LIH) of rat prostate cancer (PCa), because interventional manipulations of PCa have been reported to increase metastatic dissemination. METHODS: Seven to nine days after either subcutaneous or orthotopic implantation of MatLyLu cells, LIH (46.5 degrees C) was induced using pulsed irradiations of a neodymium:yttrium-aluminum-garnet laser. Both local control and distant metastases were evaluated. Plasma metalloproteinase 9 (MMP-9) was tested as a possible marker of PCa progression and LIH response. RESULTS: Twelve days after LIH treatment of subcutaneous tumors, the volumes were reduced by 64% after 8 minutes of irradiation, 73% after 10 minutes, 81% after 15 minutes, and 91.1% after 20 minutes. In the orthotopic model, the corresponding tumor reductions were 44% after 10 minutes, 61% after 20 minutes, and 65% after 30 minutes. Lung metastases were observed in only 1 animal with subcutaneous tumors. In contrast, 86% of the orthotopic tumor-bearing animals treated for 30 minutes had lung metastases compared with 23% of the untreated tumor-bearing rats. MMP-9 expression was detected in both orthotopic and subcutaneous tumor tissue and in the plasma of tumor-bearing rats. The prostate tissue of healthy rats and subcutaneous tumor-bearing rats was devoid of MMP-9. The plasma levels of MMP-9 showed a trend toward direct correlation with local tumor control but no correlation with the incidence of metastasis. CONCLUSIONS: These data emphasize the importance of the site of tumor implantation for evaluation of the efficacy of therapeutic interventions and may warrant further studies before widespread clinical application of LIH as monotherapy. PMID- 10869661 TI - Distribution of neuropeptides, histamine content, and inflammatory cells in the ureter. AB - OBJECTIVES: To determine the anatomic distribution of select neuropeptides (neurokinin A [NKA], substance P [SP], and bradykinin [BK]), of inflammatory cells (leukocytes and mast cells), and the histamine content in the normal swine ureter and compare the findings with regions of increased ureteral contractility. METHODS: Ureters from 10 pigs were obtained and cut into eight segments, proximally to distally. A portion of each ureteral segment was suspended in Krebs buffer (37 degrees C) and attached to force displacement transducers, and spontaneous contractility was measured for 30 minutes. A second portion was assayed for histamine, NKA, SP, and BK using enzyme-linked immunosorbent assay. A third portion was fixed in 10% buffered formalin, stained with hematoxylin-eosin, and evaluated histologically. RESULTS: Ureteral contractility was found to be highest in the most proximal and most distal regions of the ureter. Similarly, SP content was three times greater in the proximal ureter and two times greater in the distal ureter than in the midureter (P <0.05, n = 10). The total NKA and BK content were also higher in the proximal and distal ureter than in the midureter. Conversely, the histamine content was consistent throughout the ureter. Moreover, no significant difference in the distribution of inflammatory cells was identified throughout the ureter. CONCLUSIONS: The anatomic distribution of NKA, SP, and BK in the ureter corresponded to regions of increased spontaneous ureteral contractility, more specifically the proximal and distal ureter. Neuropeptides may play a significant role in ureteral contractility and may be a target for pharmacologic mediation during obstruction and stone passage. PMID- 10869662 TI - A chat with John Lattimer. PMID- 10869663 TI - Supercritical-fluid chromatography and supercritical-fluid extraction. PMID- 10869664 TI - Progress in packed column supercritical fluid chromatography: materials and methods. AB - This article summarizes recent developments in packed column supercritical fluid chromatography. Silica-based chemically bonded sorbents, similar to those used for HPLC, are widely used with solvent-modified fluids containing additives to suppress undesirable solute-sorbent interactions that lead to poor peak shapes. Composition programming is the most useful approach to gradient elution separations since solvent-modified fluids have low compressibility. Packed column SFC is most useful for the separation of mixtures usually separated by normal phase HPLC. Compared to normal-phase HPLC it offers faster separations, higher efficiencies, faster column re-equilibration, and a wider range of experimental variables for optimization. Packed column SFC is being increasingly selected for the analytical and preparative separation of racemic mixtures using enantiomer selective sorbents. PMID- 10869665 TI - Tuning of mobile and stationary phase polarity for the separation of polar compounds by SFC. AB - The separation of polar compounds by supercritical-fluid chromatography (SFC) is reviewed. New developments in mobile and stationary phase tuning are reviewed for packed and packed capillary SFC. In terms of mobile phase polarity adjustment, new pure and multiple component fluids are presented. The approach of tuning the polarity of the stationary phase as a way of increasing the range of polar compounds analyzed by SFC using pure CO(2) is discussed using either silica-based or new materials as stationary phase. Chiral, liquid crystal and polymer-based stationary phases coated on particles are widely covered in this review as an interesting approach to separate polar compounds avoiding the major drawbacks associated to the use of modifiers in SFC. PMID- 10869666 TI - Element selective detection for supercritical-fluid chromatography. AB - This manuscript describes the use of Supercritical-Fluid Chromatography (SFC) with plasma spectrometric detection for the analysis of organometallics. An introduction on the principles and characteristics of Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES) and Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) is included, along with a discussion about requirements for coupling SFC to plasma detection and the different approaches for interfacing SFC to ICP. The last part of this review paper provides a comprehensive description of SFC-ICP applications for the analysis of organometallics containing iron, silicon, tin, chromium, arsenic, lead, mercury and antimony. PMID- 10869668 TI - High-speed screening of combinatorial libraries by gradient packed-column supercritical fluid chromatography. AB - Packed-column supercritical fluid chromatography (pSFC) with a fast composition gradient is used as a rapid screening tool for combinatorial chemistry. The advantages of fast analysis speed and fast recovery to initial conditions are demonstrated. Retention time reproducibility is similar to isocratic analyses by pSFC and ranges from 0.37 to 0.64% relative standard deviation. A mixture of beta blockers illustrates the 'normal phase' retention mechanism. For these solutes and a range of analytes, the peak width is relatively constant. Such behavior permits the classical peak capacity equation to be reduced to a simple, straightforward form. Using this as a performance metric, pSFC is shown to be 5 10 times faster than reversed phase HPLC for library screening. PMID- 10869667 TI - Separation of drugs by packed-column supercritical fluid chromatography. AB - Packed-column supercritical fluid chromatography (pSFC) has been expected to analyze various kinds of compounds. Many researchers have expected a new chromatographic technique that overcomes the limitations of other techniques, HPLC and GC. In pharmaceutical development, chromatography plays an important role in the evaluation of safety and efficacy of a new compound. This article provides an overview of the separation of drugs by pSFC. The effects of the chromatographic parameters were studied for the separation of steroids. In chiral separation, the successful results were shown and compared with HPLC. PMID- 10869669 TI - The development of a semi-preparatory scale supercritical-fluid chromatograph for high-throughput purification of 'combi-chem' libraries. AB - A unique separator was developed which allowed automatic separation and peak collection using semi-preparative supercritical fluid chromatography (SFC). A peak detector switched the effluent between waste and special collection cassettes. Up to 50 mg of various solutes were injected onto a 21-mm I.D. Cyano column. The entire flow path was contained and no aerosols were generated. Collection efficiency was as high as 95%. Peak purity was often greater than 99. 9%. Typical run times were less than 10 min. An analytical SFC was used to screen the performance of a wide range of mobile and stationary phases for the elution of more than 60 miscellaneous small drug compounds. The best 'universal' gradient employed 0.4% isobutyl or isopropylamine dissolved in methanol, then mixed from 5 to 55% into carbon dioxide at 10%/min. Flow rate was 50 ml/min. The analytical SFC was shown to be a good predictor of the semi-prep instrumental performance. PMID- 10869670 TI - Separation of vitamins by supercritical fluid chromatography with water-modified carbon dioxide as the mobile phase. AB - Supercritical fluid chromatography (SFC) has become a technique for solving problems that are difficult to be monitored by other chromatographic methods. However, the most widely used fluid, is no more polar than hexane. Polar samples which are difficult to be analyzed with pure supercritical CO(2) because of their high polarity can be separated by adding polar modifiers to supercritical CO(2). In this paper various vitamins were well separated using water-modified supercritical CO(2) fluid. The amount of water dissolved in supercritical CO(2) was measured using an amperometric microsensor made of a thin film of perfluorosulfonate ionomer (PFSI). PMID- 10869671 TI - Development of a packed-column supercritical fluid chromatography/atmospheric pressure chemical-ionisation mass spectrometric technique for the analysis of atropine. AB - A packed-column supercritical fluid chromatography/atmospheric pressure chemical ionisation mass spectrometry (pSFC-APCI/MS) method has been developed for the determination of atropine from Atropa belladonna L extracts. The technique does not require any kind of derivatisation prior to the analysis. The optimum conditions were studied by using the pure substance in methanol (MeOH). All samples were simply dissolved in MeOH and injected into the mobile phase. Detection was achieved by using mass spectrometry (MS) with atmospheric pressure chemical ionisation (APCI). Terbutaline was used as an internal standard for the determination of the analytical reproducibility. The supercritical carbon dioxide (scCO(2)) mobile phase was modified by 15% MeOH containing 0.5% trifluoroacetic acid (TFAA) and 0.5% diethylamine (DEA) additives. Concentrations of atropine were determined with a relative standard deviation of less than 1% by the pSFC APCI/MS procedure for a sample containing atropine and terbutaline. The correlation coefficient was 0.997 and detection limit 700 pg. The absolute retention time was 9.87 min with a standard deviation of 5.2x10(-3) min and a relative standard deviation of 0.61% with respect to terbutaline. PMID- 10869672 TI - Packed-column supercritical fluid chromatography for the analysis of isosorbide-5 mononitrate and related compounds in bulk substance and tablets. AB - We describe a packed-column supercritical fluid chromatographic method that can be used for the analysis of isosorbide-5-mononitrate (5-ISMN) bulk substance and the 5-ISMN content of Imdur tablets. The method is based on methanol-modified carbon dioxide as the mobile phase and porous graphitized carbon (PGC, Hypercarb) as column support at 40 degrees C and 100 bar back pressure. The method makes it possible to simultaneously determine 5-ISMN and related compounds. In order to elute NO(3)(-) with acceptable retention time a quarternary ammonium hydrogen sulfate salt is added to the methanol modifier. An almost linear increase of the retention time with increasing carbon content of the counter ion was found. Tetramethyl ammonium hydrogen sulfate 5 mM in methanol was used in the final method as polar modifier for the simultaneous determination of possible degradation products within 12 min. The present method can separate and detect related compounds such as isosorbide-2, 5-dinitrate, isomannidemononitrate and isosorbide-2-mononitrate at the 0.1% (w/w) level as required by regulatory guidelines. Nitrate can be detected down to about 0.02% (w/w). Repeated analyses of ground tablet powder gave an assay precision for isosorbide-5-mononitrate of 1.4% (R.S.D., eight samples and two injections of each). For related substances at an area percent of 0. 1 the precision was less than 10%. PMID- 10869673 TI - Analysis of the sulfomycin component of alexomycin in animal feed by enhanced solvent extraction and supercritical fluid chromatography. AB - Enhanced solvent extraction (ESE) was used to isolate components of alexomycin from different types of animal feed samples. It was demonstrated that ESE was more economical as regards money and, for this particular matrix, twice as fast as supercritical fluid extraction (SFE) even though it was demonstrated that alexomycin can be extracted from feed quantitatively using both ESE and SFE. Supercritical fluid chromatography was used to separate alexomycin from other co extractives of feed since liquid chromatography was unable to isolate the alexomycin for quantification purposes. Our results also showed that the concentration of alexomycin in an extruded sample was approximately half of the concentration which was originally added to the feed. PMID- 10869674 TI - Supercritical fluid chromatography as a method of analysis for the determination of 4-hydroxybenzylglucosinolate degradation products. AB - In the present study analytical and preparative supercritical fluid chromatography (SFC) were used for investigation of myrosinase catalysed degradation of 4-hydroxybenzylglucosinolate (sinalbin). Sinalbin occurs as a major glucosinolate in seeds of Sinapis alba L., in various mustards and other food products. The degradation products were identified and quantified by analysis based on a developed SFC method using a bare silica column. Determinations comprised transformation products of sinalbin, produced both during degradation of isolated sinalbin, and during autolysis of meal from S. alba seeds. The conditions in the developed SFC method were used as basis for the preparative SFC procedure applied for isolation of the components prior to their identification by nuclear magnetic resonance (NMR) spectroscopy. Myrosinase catalysed sinalbin hydrolysis resulted in the reactive 4-hydroxybenzyl isothiocyanate as an initial product at pH values from 3.5 to 7.5 whereas 4 hydroxybenzyl cyanide was one of the major products at low pH values. 4 Hydroxybenzyl isothiocyanate was found to disappear from the aqueous reaction mixtures in a few hours, as it reacted easily with available nucleophilic reagents. 4-Hydroxybenzyl alcohol was found as the product from reaction with water, and with ascorbic acid, 4-hydroxybenzylascorbigen was produced. PMID- 10869675 TI - Supercritical fluid chromatography as basis for identification and quantitative determination of indol-3-ylmethyl oligomers and ascorbigens. AB - Indol-3-ylmethylglucosinolate (glucobrassicin) occurs in most plants of the Brassicaceae family together with hydroxy and methoxy derivatives of glucobrassicin. These compounds and products produced therefrom have been the subject of considerable research interest due to their potential anticarcinogenic effects, and thereby a need for techniques to work with the individual compounds. A method using normal-phase supercritical fluid chromatography (SFC) with methanol as modifier has been developed for determination and quantification of the various indol-3-ylmethyl derivatives including ascorbigens formed from the glucobrassicin degradation product, indol-3-ylmethanol, under acidic conditions (pH 2-6) with and without the presence of ascorbic acid. The SFC method had detection limits in the 10-100-pmol range. In the absence of ascorbic acid a range of oligomers were formed, whereas the presence of ascorbic acid favoured the formation of ascorbigen and products thereof. Quantitatively important indol 3-ylmethyl oligomers consisting of up to five indol rings have been purified with preparative SFC and identified from MS and 1D and 2D NMR experiments with complete assignment of chemical shifts to all of the atoms. Investigation of the autolysis products of white cabbage showed that ascorbigens were the quantitatively dominating degradation products of indol-3-ylmethylglucosinolates. PMID- 10869676 TI - Carbon-based quantitation of pyrethrins by supercritical-fluid chromatography. AB - All six insecticide active ingredients in pyrethrum extract were quantified by supercritical fluid chromatography and carbon calibration. Allethrin is a suitable reference compound for carbon calibration and pyrethrins calibrations. Carbon quantification in SFC is also applied to pyrethroids (phenothrin, permethrin, cypermethrin, fenvalerate and deltamethrin) and alkanes. Halogen substitution on pyrethroids requires halogens on the reference calibration compound. The method was applied to commercial extracts. PMID- 10869677 TI - SFC/FTIR, SFC/APCI-MS and MALDI-TOF-MS for the analysis of siloxane-ethylene oxide copolymers. AB - Polydimethylsiloxanes (PDMSs) modified by introducing ethylene oxide units with the aim of forming sufficiently water-soluble siloxane compounds were characterized using supercritical fluid chromatography (SFC) coupled with Fourier transform infrared (FTIR) spectroscopy and atmospheric pressure chemical ionization mass spectrometry (APCI-MS), and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). SFC has a domain in analyzing oligomers. Hyphenated techniques enable elucidation of the components. Remarkable is the resolution and short analysis time of MALDI-TOF-MS. SFC also allows quantification of the basic and reaction products. PMID- 10869678 TI - Enantiomer separation by complexation SFC on immobilized Chirasil-nickel and Chirasil-zinc. AB - The use of complexation SFC for enantiomer separation of Lewis base selectands on chiral nickel(II)- and zinc(II)-bis[(3-heptafluorobutanoyl)-10-methylene-(1R) camphora te] chemically bonded to poly(dimethylsiloxane) (Chirasil-nickel and Chirasil-zinc) and employed as Lewis acid selectors is described. The method is especially suited for less volatile and configurationally labile racemates. The variation of the experimental parameters temperature T, pressure p and density rho of the mobile phase carbon dioxide on the retention factor k, relative retention r and chiral separation factor alpha is studied, providing insights into the mechanisms of chiral recognition under supercritical conditions. For mecoprop methyl ester (methyl 2-(4-chloro-2-methylphenoxy)propanoate) an unusual increase of alpha at increased temperature is observed on Chirasil-nickel. Supercritical carbon dioxide does not inadvertently affect the complexation equilibria between Lewis donor selectands and the Lewis acid metal selectors during complexation SFC. PMID- 10869679 TI - Separation of ketoconazole enantiomers by chiral subcritical-fluid chromatography. AB - The separation of ketoconazole enantiomers by subcritical-fluid chromatography using an amylose-based column is described. Drastic changes in the resolution have been obtained for the different organic modifiers evaluated, with ethanol providing the best results. Other chromatographic parameters such as temperature, pressure and flow-rate have also been studied. The best results in terms of resolution and analysis time were obtained using 30% ethanol (containing 0.1% triethylamine and 0.1% trifluoroacetic acid), a pressure of 300 bar, a temperature of 35 degrees C and a flow-rate of 3 ml/min. Under these conditions the ketoconazole enantiomers are resolved in a short time (less than 7 min) and with high resolution (4.29). PMID- 10869680 TI - Determination of the 2-bromomethyl-2-(2,4-dichlorophenyl)-1, 3-dioxolan-4 yl)methyl benzoate diastereoisomers by supercritical fluid chromatography. AB - The study of the separation and determination of the two 2-bromomethyl-2-(2,4 dichlorophenyl)-1,3-dioxolan-4-yl-methyl benzoate diastereoisomers is presented in this work. The separation was achieved using packed column supercritical fluid chromatography. Three stationary phases were checked, the C(18) packing providing the best results. The influence of several parameters (nature and percentage of modifier, temperature and pressure) on the separation were also evaluated. The baseline separation (R(s)=1.78) of the compounds was obtained in only 2 min, using a C(18) column, a pressure of 100 bar, a temperature of 35 degrees C, a flow-rate of 2. 5 ml/min and 5% of 2-propanol as organic modifier. Under these conditions the unwanted diastereoisomer (trans) was eluted before the main compound (cis), so the determination of the purity was more accurate. PMID- 10869681 TI - Applications of supercritical fluid extraction and chromatography in forensic science. AB - Supercritical fluid technology is a rapidly expanding analytical technique. Here we give a brief insight into the background of supercritical fluid technology and how supercritical fluid extraction and supercritical fluid chromatography work in analysis. The applications of these two techniques in forensic science are known to be important. The main area of forensic use of supercritical fluid technology is in the sample preparation and separation of drugs of abuse particularly opiates, cannabinoids, cocaine and sedatives. Supercritical fluid technology can be used for both time-of-death-related drug analysis and for obtaining information relating to long term drug abuse. We also give a review of the use of supercritical fluids in two other major forensic areas, fingerprinting and the extraction and separation of explosives from both bombing events and gunshot residues. Overall we show that supercritical fluid technology is fast becoming a major part of forensic investigations and that it is an invaluable analysis technique. PMID- 10869683 TI - Introducing selective supercritical fluid extraction as a new tool for determining sorption/desorption behavior and bioavailability of persistent organic pollutants in sediment. AB - This review article intends to introduce the possibility of utilizing selective supercritical fluid extraction (SFE) as a tool to study sorption/desorption processes and bioavailability of persistent organic pollutants (POP) in sediment. Sorption/desorption behavior and bioavailability studies of POPs is a large research area, but still many unsolved problems exists. Therefore novel approaches to investigate mechanistic behavior of POPs in sediments are needed. Present literature on SFE points to the fact that selective SFE measurements can improve our knowledge, and recent investigations have been performed that demonstrate this. Results obtained with selective SFE can be connected to desorption of POPs in sediments under natural conditions in aquatic ecosytems. The ultimate goal is to use selective SFE as a way to determine the bioavailable fraction present within a matrix. A few preliminary results are presented here which may serve as a starting point for future studies. PMID- 10869682 TI - Separation of metal chelates and organometallic compounds by SFC and SFE/GC. AB - Supercritical fluid chromatography (SFC) combines the high diffusion coefficients of gas chromatography (GC) and the solubility properties of liquid chromatography (LC). SFC generally requires lower temperatures for chromatographic separations and thus is more suitable for analyzing thermally labile compounds including a number of metal chelates and organometallic compounds. SFC also allows interfacing between supercritical fluid extraction (SFE) and chromatographic analysis of metal-containing compounds. A large number of metal chelates and organometallic compounds can be separated by SFC. This article summarizes SFC separation of various chelates of transition metals, heavy metals, lanthanides and actinides as well as organometallic compounds of lead, mercury, and tin reported in the recent literature. This article also discusses SFC detection systems and the determination of solubility of organometallic compounds by SFC. PMID- 10869684 TI - Supercritical fluid extraction for the analysis of pesticide residues in miscellaneous samples. AB - Supercritical fluid extraction (SFE) procedures for pesticide residue analysis are reviewed and discussed. A variety of applications were classified, on matrices such as fruits, vegetables, soils, biological tissues, and other materials. Emphasis is placed on analysis of samples with a high water content containing polar pesticides, with particular attention paid to the multiresidue analyses. PMID- 10869685 TI - On-line detection for supercritical-fluid extraction. AB - An overview of the direct coupling of supercritical-fluid extraction (SFE) to distinct detection is presented. The versatile construction of this kind of hybrid analytical instrumentation is manifested through the discussion of several interface prototypes, whose characteristics satisfy the requirements of both the SF extractor and detector. The application of several experimental strategies during the whole analytical process are proposed for the successful quantification of the leaching yield. Typical examples of SFE with on-line detection in several fields of analysis are presented, considering of special interest the information provided by the molecular and atomic techniques used. Finally, the real analytical capabilities of the automation of sample handling by SFE-continuous detection are also discussed. PMID- 10869686 TI - Supercritical carbon dioxide extraction of lipids from Eucalyptus globulus wood. AB - Various typical lipid components of wood extractives have been isolated from Eucalyptus globulus wood by supercritical carbon dioxide modified with methanol. The influence of various extraction parameters on the yield and qualitative composition of the extracts have been studied. The extracts were analyzed by gas chromatography-mass spectrometry and compared with those obtained by Soxhlet extraction with acetone, the standard method for the determination of wood extractives. The qualitative and quantitative results obtained by both methods were in good agreement. The experimental planning to asses the influence of pressure, temperature and percentage of methanol and their interactions on the extraction efficiency was carried out with a factorial design, followed by multiple linear regression algorithm. PMID- 10869687 TI - Experimental design in supercritical fluid extraction of cocaine from coca leaves. AB - An optimisation procedure for the supercritical fluid extraction (SFE) of cocaine from the leaves of Erythroxylum coca var. coca was investigated by means of experimental design. After preliminary experiments where the SFE rate-controlling mechanism was determined, a central composite design was applied to evaluate interactions between selected SFE factors such as pressure, temperature, nature and percentage of the polar modifier, as well as to optimise these factors. Predicted and experimental contents of cocaine were compared and robustness of the extraction method estimated by drawing response surfaces. The analysis of cocaine in crude extracts was carried out by capillary GC equipped with a flame ionisation detector (GC-FID), as well as by capillary GC coupled with a mass spectrometer (GC-MS) for peak identification. PMID- 10869688 TI - A non-destructive method to determine the safranal content of saffron (Crocus sativus L.) by supercritical carbon dioxide extraction combined with high performance liquid chromatography and gas chromatography. AB - A supercritical carbon dioxide extraction method to obtain selectively volatile compounds of saffron without sample destruction has been developed. The influence of both pressure and temperature was studied, 20 MPa and 100 degrees C being the best conditions to extract the total safranal content. A decrease in supercritical fluid density was shown to be a critical parameter for enhancing the extraction power of carbon dioxide. For all the assay conditions, the extracts mainly contained safranal and HTCC, as demonstrated by gas chromatography and high-performance liquid chromatography analyses. Both chromatographic methods were suitable for safranal quantification and showed excellent agreement. Supercritical extracts from five different saffron types were studied by high-performance liquid chromatography and their safranal contents were determined. PMID- 10869689 TI - High-resolution gas-chromatographic analysis of the secondary metabolites obtained by subcritical-fluid extraction from Colombian rue (Ruta graveolens L.). AB - Subcritical (CO(2)) extraction, carried out in a J&W Scientific High Pressure Soxhlet Extractor, was used to isolate secondary metabolites from leaves, flowers, stems and roots of Colombian rue (Ruta graveolens L.). The various extracts were analyzed by capillary chromatography, on an HP-5 (30 m) column, using nitrogen-phosphorus, flame ionization, and mass selective detection systems. Kovats indexes and mass spectra (electron impact, 70 eV) were employed for compound identification. The extracts from the various parts of rue studied had different compositions. The number of compounds detected at concentrations above 0.01% (w/w) in the extracts from leaves, flowers, stems and roots, was 78, 45, 25 and 24, respectively. 2-Nonanone (8.9%), 2-undecanone (13.4%), chalepensin (13.0%), and geijerene (19.3%) were the main constituents found in the extracts from rue leaves, flowers, stems and roots, respectively. Furanocoumarins, furoquinolines, hydrocarbons and benzodioxol derivatives were the main compound families found in all extracts, at total concentrations between 3.7 and 33.9%, depending on the part of the plant. The extraction method used has low environmental impact and produced solvent-free extracts in good yield with no pigments, waxes, resins, or high-molecular weight compounds which may interfere with the isolation and analysis of the alkaloids responsible for rue's biological activity, which were extracted in relatively high yield. PMID- 10869690 TI - Determination of food constituents based on SFE: applications to vitamins A and E in meat and milk. AB - A method has been developed for the determination of vitamins A and E in food using supercritical fluid extraction (SFE), applying liquid or solid trapping, with an accuracy equal to conventional solvent extraction methods. Under optimal conditions, using methanol modified carbon dioxide as a supercritical fluid, Hydromatrix as a water adsorbent, and with a small amount of ascorbic acid and methanol added to the sample, the extraction time is reduced to 80 min. This time is considerably shorter than in conventional methods. Other advantages are the reduction of manual manipulations leading to lower labour costs and reduced consumption of organic solvents in the sample preparation step. PMID- 10869691 TI - Use of supercritical fluid extraction in the analysis of pesticides in soil. AB - The applicability of supercritical fluid extraction (SFE) in pesticide residue analysis in soil was investigated by analysing real soil samples from field experiments. Additionally, radiotracer batch experiments were performed to study the release of non-extractable residues. High repeatability, accuracy and high selectivity were the most important advantages of SFE in residue analysis. Extracts with low amounts of coextractants from the soil matrix were achieved, allowing extracts to be pooled and concentrated without further clean up steps. Thus, the limited volume of extraction thimbles of the SFE apparatus used could be compensated and insufficiently high limits of determination could be improved. Although the application of methanol-modified supercritical CO(2) was a time saving extraction procedure which reduced solvent usage and solvent waste, SFE efficiency proved only competitive to conventional slurry and Soxhlet extraction. No exhaustive release of non-extractable residues was achieved in radiotracer batch experiments. PMID- 10869692 TI - Supercritical-fluid extraction of synthetic pyrethroids from wool. AB - A stepwise approach was used to develop a supercritical fluid extraction (SFE) method for analysis of synthetic pyrethroids (SPs) on a wool matrix, commencing with a simple inert matrix to examine the solubility of the pyrethroids in the extraction fluid CO(2) and then extended to the real wool matrix. Chemometric approaches were used to determine the SFE optimum conditions. It was found that pyrethroids were readily extractable from an inert matrix over a wide range of pressure (170-350 atm) and at low temperature (<90 degrees C). Subambient hexane efficiently trapped the compounds from the depressurised fluid. Excessively high pressure and temperature resulted in poor trapping, isomerisation and possibly degradation of some components. With spiked wool samples method modifications focused on reducing the coextraction of grease, a bulk matrix component of raw wool. By using alumina (containing 8% moisture) and operating the extraction at 50 degrees C, 200 atm for 60 min, sufficiently clean extracts of pyrethroids suitable for gas chromatography-electron-capture detection analysis were obtained. The recoveries of all SPs were satisfactory (78-101%) over the range of 0.5-5 microg/g levels of these compounds. The precision of the entire analysis procedure was comparable to the conventional Soxhlet extraction method. Detection limits of some commonly used SPs for sheep treatment were also evaluated. Comparable results relative to those achieved by solvent extraction for incurred wool samples were obtained with a recovery of 81-85%. The results, however, suffered high uncertainties (R.S.D. approximately 19-24%) due to the small amount of wool sample taken in each extraction and the suspected inhomogeneity of the wool. Different persistences of cypermethrin isomers in wool were observed. PMID- 10869693 TI - A novel gliotic P2 receptor mediating cyclooxygenase-2 induction in rat and human astrocytes. AB - In astrocytic cultures maintained in vitro, a brief challenge with the ATP analog alpha,beta methyleneATP (alpha,betameATP) results, 3 days later, in marked elongation of astrocytic processes, an event that resembles the astrocytic hypertrophy known to occur in vivo during reactive astrogliosis. alpha,beta meATP induced effects were observed in primary astrocytes obtained from both rat striatum and cortex (a brain area highly involved in chronic neurodegenerative pathologies), as well as in human astrocytoma cells (ADF cells). Purine-induced gliosis could be reversed by the non-selective P2X/P2Y receptor antagonist pyridoxalphosphate-6-azophenyl-2', 4'-disulphonic acid (PPADS), but not by oxidized ATP (an antagonist of the P2X(7) receptor), in line with previous studies of our laboratory suggesting the involvement of a P2Y receptor subtype. Induction of reactive gliosis was preceded by increased expression of cyclooxygenase-2 (COX-2), an enzyme whose excessive activation has been implicated in both acute and chronic neurodegenerative diseases. The selective COX-2 inhibitor NS-398 prevented both purine-induced astrogliosis and the associated COX-2 induction, suggesting that inhibition of the transcription of the COX-2 gene may also contribute to the anti-inflammatory properties of this agent. Significant blockade of both alpha,beta meATP-mediated reactive gliosis and COX-2 induction was also observed with PPADS. These data suggest that COX-2 mediates P2Y receptor-induced reactive astrogliosis, and that antagonists selective for this receptor subtype may represent a novel class of anti inflammatory agents of potential interest in acute and chronic neurological disorders characterized by an inflammatory component and reactive gliosis. PMID- 10869694 TI - Imagination and reality in the search for the P2Y receptors. AB - A great body of evidence based on tissue and organ physiology and pharmacology led to the recognition, widespread by about 1990, that there must be cell membrane receptors for extracellular nucleotides to transduce their effects. This evidence was provided by the pioneering work of Geoffrey Burnstock and those who worked with him, or was developed by others starting from that information. This article will review how we could start from that foundation to clone the first known gene for such a receptor, P2Y(1). Some unusual properties of that receptor were revealed. I will consider further the P2Y receptors as a class - its definition, now that many such genes have become known. Imagination and reality have been intertwined in this saga. PMID- 10869695 TI - The effects of exercise and training on human cardiovascular reflex control. AB - During physical activity, there is a graded withdrawal of vagal cardiac tone and a graded increase in sympathetic cardiac and vasomotor tone, initiated through both central command from the somatic motor cortex and muscle chemoreceptive and mechanoreceptive inputs. In parallel, there is an upward resetting of the operating point of the arterial baroreflex, with preserved reflex sensitivity. In contrast to the traditional interpretation that blood flow through exercising muscle is independent of vasomotor neural influences because of the dominance of local dilator metabolites, recent evidence suggests that both constrictor and dilator sympathetic neural influences may be involved in determining absolute levels of perfusion. Post-exercise, there is a period of relative hypotension that is associated with decreased peripheral resistance. Some, but not all, evidence indicates a causal role for reduced sympathetic drive. Chronic exercise training appears to reduce resting sympathetic activity, with parallel changes in the gain of a variety of cardiovascular autonomic reflexes initiated from cardiovascular sites. These changes may be attributable at least partly to masking of arterial baroreflexes by the impact of elevated blood volume on low pressure baroreceptors. The reductions in sympathetic drive that follow training are more pronounced in patients with essential hypertension than in normotensive individuals and are likely to underlie the anti-hypertensive effect of exercise. PMID- 10869696 TI - NANC transmission at a varicosity: the individuality of single synapses. AB - Nerve terminals consist of several hundred varicosities or synapses, each with a single active zone. The smooth muscle membrane apposing varicosities within about 50 nm is occupied by a 1-microm diameter cluster of P2X(1) receptors together with a mixture of other P2X subtypes; the rest of the membrane possesses small (0.4 microm diameter) clusters of P2X(1) to P2X(6) subunits. The small P2X clusters appear to form large clusters during development. This is supported by the observation that chimeras of P2X(1) subunits and green fluorescent protein (P2X(1)-GFP), when packaged into adenoviruses used to infect excitable cells, initially form a diffuse distribution of small clusters of P2X(1)-GFP in the membrane; these can be later observed in real time to form large clusters. Recording the electrical signs of ATP release from single adjacent varicosities, or using antibodies to label the extent of exocytosis from them, shows that they release with quite different probabilities. There are large quantitative differences in the extent of P2X autoreceptors on the membranes of individual varicosities. These will contribute to the differences in the probability of secretion from individual varicosities. The present analysis of NANC transmission at single varicosities indicates that individual synapses possess different probabilities for the secretion of transmitter as well as different complements of autoreceptors and mixtures of postjunctional receptor subunits. PMID- 10869697 TI - Inhibition of potassium and calcium currents in neurones by molecularly-defined P2Y receptors. AB - Messenger RNAs and cDNAs for individual cloned P2Y(1), P2Y2 and P2Y(6) nucleotide receptors have been expressed by micro-injection into dissociated rat superior cervical sympathetic neurones and the effects of stimulating the expressed receptors on voltage-activated N-type Ca(2+) currents and M-type K(+) currents recorded. Both currents were reduced by stimulating all three receptors, with the following mean IC(50) values: P2Y(1) (agonist: ADP) - I(K(M)) 6.9 nM, I(Ca) 8.2 nM; P2Y(2) (agonist: UTP) - I(K(M)) 1.5 microM, I(Ca) 0.5 microM; P2Y(6) (agonist: UDP) - I(K(M)) 30 nM, I(Ca) 5.9 nM. Inhibition of I(K(M)) was voltage independent and insensitive to Pertussis toxin; inhibition of I(Ca) showed both voltage-sensitive and insensitive, and Pertussis toxin-sensitive and insensitive components. It is concluded that these P2Y receptors can couple to more than one G protein and thereby modulate more than one ion channel. It is suggested that these effects on K(M) and Ca(N) channels may induce both postsynaptic excitory and presynaptic inhibitory responses. PMID- 10869698 TI - Platelet P2 receptors: from curiosity to clinical targets. AB - Adenosine 5'-diphosphate (ADP) is a paracrine mediator that activates human blood platelets, causing them to become adhesive and thereby contributing to their role in hemostasis. The actions of ADP were initially thought to be mediated by a unique ADP receptor termed P2(T) found only on platelets and antagonized by ATP, but it appears that at least two P2Y receptor subtypes are involved, a P2Y(1) receptor linked in some way to control of intracellular-free calcium levels and another P2Y receptor linked via an inhibitory G protein to adenylate cyclase. In addition, the presence of excitatory P2X(1) receptors that mediate the influx of monovalent and divalent cations in response to both ADP and ATP has been demonstrated. The precise contribution that each of these P2 receptors make to the overall phenomena associated with platelet aggregation, adhesion and hemostasis is yet to be defined. Antithrombotic agents that interfere with the actions of ADP are marketed, and P2 receptor antagonists are entering clinical trials for acute treatments of thrombosis. This review seeks to summarize the present state of knowledge of platelet P2 receptor pharmacology and therapeutics. PMID- 10869699 TI - Alkylxanthines as research tools. AB - (1) The methylxanthine caffeine has many pharmacological effects, most of which can be linked to blockade of adenosine receptors, inhibition of phosphodiesterases, and augmentation of calcium-dependent release of calcium from intracellular stores. (2) A variety of xanthines have been developed as potent and/or selective antagonists for adenosine receptors. (3) Several xanthines have been developed that are more potent and more selective inhibitors of cyclic nucleotide phosphodiesterase than caffeine or theophylline. (4) Caffeine remains the xanthine of choice for activation of intracellular calcium-sensitive calcium release channels although millimolar concentrations are required, which can have effects on other aspects of calcium regulation. PMID- 10869700 TI - Relaxation of rat distal colon by activation of muscarinic, neuronal receptors: possible involvement of P(2y)-purinoceptors. AB - McN-A-343, which is a ligand at muscarinic receptors on myenteric ganglia, was found to concentration-dependently (1-44 microM) elicit non-adrenergic relaxation of the longitudinal muscle of rat distal colon, having been precontracted with carbachol (1 microM). This effect was partly antagonized by the muscarinic receptor antagonist, pirenzepine (0.3 microM), the nerve blocker, tetrodotoxin (1 microM), or by drugs which interfere with purinergic neurotransmission (apamin [0.5 microM], reactive blue 2 [50 microM]). Blockade of nitric oxide synthase (L NNA [100 microM]), or of the cAMP (H-89 [1 microM]), or cGMP (ODQ [10 microM]) second messenger pathways did not affect the relaxatory response to McN-A-343 (14 microM). An additional, non-neurogenic component of the relaxation to this compound on carbachol induced tone is suggested to reflect a partial antagonism of the muscarinic receptors on the gut muscle by McN-A-343. PMID- 10869701 TI - Dr. Jekyll/Mr. Hyde: the dual role of extracellular ATP. PMID- 10869702 TI - Purinergic signaling at immunological synapses. AB - The early studies and hypotheses of Geoffrey Burnstock catalyzed intensive characterization of roles for nucleotides and P2 nucleotide receptors in neurotransmission and neuromodulation. These latter analyses have focused on the mechanisms of nucleotide release and action in the microenvironments of nerve endings and synapses. However, studies of various white blood cells, such as monocytes, neutrophils, and lymphocytes, suggest that locally released nucleotides also modulate intercellular signaling at so-called 'immunological synapses'. This communication describes recent findings and speculations regarding nucleotide release and signaling in several key phases of the immune and inflammatory responses. PMID- 10869703 TI - Comparison of P2X receptors in rat mesenteric, basilar and septal (coronary) arteries. AB - alpha beta meATP-evoked concentration-dependent, PPADS-sensitive, desensitising, P2X receptor-mediated, constrictions of mesenteric, basilar and septal artery rings with EC(50) values of 1, 1 and 30 microM, respectively. In patch clamp studies on acutely dissociated artery smooth cells alpha beta meATP-evoked transient inward currents (tau approximately 100 ms) with mean current densities of approximately 340, 175 and 120 pA/pF, respectively. P2X(1) receptor immunoreactivity was expressed in mesenteric and basilar arteries and this receptor subunit appears to dominate the P2X receptor phenotype in these vessels. In contrast P2X(1) receptor immunoreactivity was not detected in septal arteries and the alpha beta meATP sensitivity of constriction was not consistent with the involvement of P2X(1) receptors. These results suggest that not all arteries share a common P2X receptor phenotype. PMID- 10869704 TI - Regulated expression of the rat recombinant P2X(3) receptor in stably transfected CHO-K1 tTA cells. AB - In this report, the regulatable expression by tetracycline of the rat recombinant P2X(3) receptor in stably transfected Chinese hamster ovary (CHO-K1) expressing the tetracycline-controlled transactivator (tTA) is described. cDNA encoding the rat P2X(3)-receptor was subcloned into pTRE (a tetracycline-repressible expression vector) which was used to transfect stably CHO-K1 tTA cells. Using whole cell patch clamp techniques, 100 microM ATP evoked inward currents of 2.9+/ 1.6 nA in transfected cells grown in the absence of tetracycline (tet-). The P2X(3) receptor protein was detectable by immunoblot as early as 24 h and protein expression levels continued to increase as much as 192 h following activation of tTA by the removal of the antibiotic. Saturation binding isotherms using [35S]ATP gamma S yielded a pK(d) of 8.2+/-0.1 and a B(max) of 31.9+/-3.5 pmol/mg protein in tet- cell membranes and a pK(d) of 8.1+/-0.1 and a B(max) of 5.8+/-0.8 pmol/mg protein in tet+ cell membranes. The agonist ligands 2MeSATP and alpha beta MeATP displaced the binding of [35S]ATP gamma S in tet- cell membranes with very high affinity, yielding pIC(50) values of 9.4+/-0.2 and 7.5+/-0. 2, respectively. In tet+ cell membrane, displacement of [35S]ATP gamma S by 2MeSATP and alpha beta MeATP was of much lower affinity (pIC(50) values of 7.8 and 6.2, respectively). ATP, ADP and UTP showed similar displacement of [35S]ATP gamma S binding in tet- and tet+ cell membranes. In other experiments, cytosolic Ca(2+) was monitored using the fluorescent indicator, fluo-3. Increases in cytosolic Ca(2+) were elicited by 100 nM alpha beta MeATP in tet- cells while no increases in cytosolic Ca(2+) were detected below 100 microM alpha beta MeATP in either tet+ cells or untransfected cells. These calcium responses to alpha beta MeATP had a pEC(50) of 6.7 and were transient, returning to baseline within 120 s. Suramin produced concentration-dependent, parallel, dextral shifts of E/[A] curves to alpha beta MeATP yielding a pK(B) of 5.6. PPADS produced non-parallel, dextral shifts of E/[A] curves to alpha beta MeATP which were insurmountable. These results show for the first time, expression of a functional, homomeric recombinant rat P2X(3) receptor which is under regulated expression in a stably transfected mammalian cell line. PMID- 10869705 TI - Nitric oxide interacts with oxygen free radicals to evoke the release of adenosine and adenine nucleotides from rat hippocampal slices. AB - The present study examined some possible mechanisms underlying the previously demonstrated release of adenosine by nitric oxide (NO) donors. Perfusion with the NO-donor S-nitroso-N-acetyl penicillamine (SNAP; 300 microM) led to a significant increase in the release of [3H]purines from both unstimulated and electrically stimulated hippocampal slices prelabeled with [3H]adenine. The NO-donor also evoked the release of endogenous ATP and ADP from unstimulated slices and, when combined with electrical stimulation, the release of ATP, AMP and adenosine. The SNAP-induced [3H]purine release was calcium-dependent, but not affected by the glutamate receptor antagonists MK-801 ((+)-5-methyl-10,11-dihydro-5H-dibenzo[a, d]-cyclohepten-5,10-imine;100 nM) and CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 10 microM). Zaprinast (5 microM), an inhibitor of the cyclic GMP-dependent phosphodiesterase and 8-Br-cyclic GMP (0.01-1 mM) failed to evoke the release of purines, whereas generation of oxygen free radicals by xanthine plus xanthine oxidase did evoke purine release. Coperfusion of SNAP with the free radical scavengers superoxide dismutase (SOD; 60 microg/ml) and catalase (50 microg/ml) reduced or eliminated the ability of the NO-donor to enhance [3H]purine release, but the poly (ADP-ribosyl) synthetase (PARS) inhibitor benzamide (500 microM) did not affect it. These data indicate that NO interacts with superoxide, likely forming peroxynitrite, which subsequently acts to release adenosine and adenine nucleotides from hippocampal tissue. PMID- 10869706 TI - Types of neurons in the enteric nervous system. AB - This paper, written for the symposium in honour of more than 40 years' contribution to autonomic research by Professor Geoffrey Burnstock, highlights the progress made in understanding the organisation of the enteric nervous system over this time. Forty years ago, the prevailing view was that the neurons within the gut wall were post-ganglionic neurons of parasympathetic pathways. This view was replaced as evidence accrued that the neurons are part of the enteric nervous system and are involved in reflex and integrative activities that can occur even in the absence of neuronal influence from extrinsic sources. Work in Burnstock's laboratory led to the discovery of intrinsic inhibitory neurons with then novel pharmacology of transmission, and precipitated investigation of neuron types in the enteric nervous system. All the types of neurons in the enteric nervous system of the small intestine of the guinea-pig have now been identified in terms of their morphologies, projections, primary neurotransmitters and physiological identification. In this region there are 14 functionally defined neuron types, each with a characteristic combination of morphological, neurochemical and biophysical properties. The nerve circuits underlying effects on motility, blood flow and secretion that are mediated through the enteric nervous system are constructed from these neurons. The circuits for simple motility reflexes are now known, and progress has been made in analysing those involved in local control of blood flow and transmucosal fluid movement in the small intestine. PMID- 10869707 TI - Multiple mechanisms of fast excitatory synaptic transmission in the enteric nervous system. AB - The enteric nervous system (ENS) can control gastrointestinal function independent of direct connections with the central nervous system. Enteric nerves perform this important function using multiple mechanisms of excitatory neurotransmission in enteric ganglia. Fast excitatory synaptic transmission in the autonomic nervous system (ANS) is largely mediated by acetylcholine (ACh) acting at nicotinic cholinergic receptors but in the ENS there are noncholinergic fast excitatory neurotransmitters. There are two broad types of neurons in the ENS: S neurons and AH neurons. S neurons are interneurons and motoneurons while AH neurons are sensory neurons. Three subsets of S neurons in the myenteric plexus can be distinguished on the basis of the neurotransmitters producing fast excitatory postsynaptic potentials (fEPSPs) in each subset. In one subset, fEPSPs are mediated solely by ACh acting at nicotinic cholinergic receptors. In a second subset of S neurons, ATP acting at P2X purine receptors and ACh contribute to the fEPSP while in a third subset, fEPSPs are mediated by 5-hydroxytryptamine (5-HT) acting at 5-HT(3) receptors and ACh. Some AH neurons also receive fast excitatory synaptic input. The fEPSPs recorded from AH neurons are mediated ACh and also by glutamate acting at alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors. Multiple mechanisms of fast excitatory synaptic transmission in the ENS are likely to contribute to its capacity to regulate complex gastrointestinal functions. PMID- 10869708 TI - Neuronal morphology and the synaptic organisation of sympathetic ganglia. AB - In this article, we provide a short review of the structure and synaptic organisation of the final motor neurons in the sympathetic ganglia of mammals. Combinations of pathway tracing, multiple-labelling immunofluorescence and intracellular dye injection have shown that neurons in different functional pathways differ not only in their patterns of neuropeptide expression, but also in the size of their cell bodies and dendritic fields. Thus, vasoconstrictor neurons consistently are smaller than any other major functional class of neurons. Serial section ultrastructural analysis of dye filled neurons, together with electron microscopic and confocal microscopic analysis of immunolabelled synaptic inputs to sympathetic final motor neurons indicate that synapses are rare and randomly distributed over the surface of the neurons. The total number of synapses is simply proportional to the total surface area of the neurons. Many terminal boutons of peptide-containing preganglionic neurons do not make conventional synapses with target neurons. Furthermore, there is a spatial mismatch in the distribution of peptide-containing terminals and neurons expressing receptors for the corresponding peptides. Together, these results suggest that there are likely to be significant differences in the ways that the final sympathetic motor neurons in distinct functional pathways integrate their synaptic inputs. In at least some pathways, heterosynaptic actions of neuropeptides probably contribute to subtle modulation of ganglionic transmission. PMID- 10869709 TI - Modulation of fast synaptic transmission by presynaptic ligand-gated cation channels. AB - There is now considerable evidence demonstrating that ligand-gated cation channels (i.e., P2X, nicotinic, kainate, NMDA, AMPA and 5-HT(3) receptors), in addition to mediating fast excitatory neurotransmission, may be located presynaptically on nerve terminals in the peripheral and central nervous systems where they function to modulate neurotransmitter release. This modulation can be facilitation, inhibition or both. In this article, we first outline the multiple mechanisms by which activation of presynaptic ligand-gated cation channels can modulate spontaneous and evoked neurotransmitter release, before reviewing in detail published electrophysiological studies of presynaptic P2X, nicotinic, kainate, NMDA, AMPA and 5-HT(3) receptors. PMID- 10869710 TI - Role of gap junctions in acetylcholine-induced vasodilation of proximal and distal arteries of the rat mesentery. AB - We have previously shown that myoendothelial gap junctions are more prevalent in distal than in proximal arteries of the rat mesentery. In the present study we have investigated the role of gap junctions in the mechanism of action of endothelium-derived hyperpolarizing factor (EDHF) in these same vessels following relaxation with acetylcholine. Arteries were pre-constricted with phenylephrine and concentration response curves to acetylcholine were constructed in the presence of N(G)-nitro-L-arginine methyl ester (L-NAME; 10(-5) M) and indomethacin (10(-5) M) to prevent effects due to the release of nitric oxide and prostacyclins. Nitric oxide was found to have only a small role in the relaxation of the proximal vessels and was not involved in the relaxations of the distal vessels. 18 alpha-Glycyrrhetinic acid (10(-5) M), a putative gap junction uncoupler, significantly reduced acetylcholine-induced relaxations by 50% in both proximal and distal vessels. Potassium channel antagonists, tetraethylammonium chloride (TEA; 10(-3) M) and barium chloride (10(-4) M), together abolished the dilatory response in the proximal mesenteric arteries, but did not completely block responses in the distal arteries. The data suggest that gap junctions contribute significantly to the acetylcholine-induced relaxation in both proximal and distal arteries of the rat mesentery. We hypothesize that the absence of a correlation between the role of gap junctions and the incidence of myoendothelial gap junctions in these same vessels is due to significant effects of the inhibitors on gap junctions located in the smooth muscle layers of the larger vessels. PMID- 10869711 TI - Effect of NGF, BDNF, bFGF, aFGF and cell density on NPY expression in cultured rat dorsal root ganglion neurones. AB - The effect of neurotrophic factors on neuropeptide Y (NPY) expression was studied in adult rat dispersed dorsal root ganglion (DRG) cultures. Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), acidic fibroblast growth factor (aFGF) or basic FGF was included in the culture medium during incubation for 72 h. In untreated cultures, around 18% of all neurones (visualized by antibodies to PGP 9.5) expressed NPY-like immunoreactivity (LI). In contrast, in vivo uninjured neurones do not contain detectable levels of NPY-LI. In the immunohistochemical analysis aFGF increased the percentage of NPY-immunoreactive (-IR) neurones 1.8 fold, while NGF, BDNF or bFGF had no significant effect on NPY expression. When the effect of these growth factors was monitored with non-radioactive in situ hybridization, both aFGF and bFGF caused a significant increase (2.25- and 1.8 fold, respectively), whereas, again, NGF and BDNF had no effect. The results also showed an effect of cell density on NPY expression, whereby fewer neurones expressed NPY in high than in low density cultures. This difference was seen in untreated as well as growth factor-treated cultures. The present results support the hypothesis that DRG neurones in culture are in an axotomized state, since they express NPY to about the same extent as axotomized DRG neurones in vivo. Surprisingly, two growth factors of the FGF family enhance NPY expression in DRG neurones, which is in apparent contrast to a published in vivo study [Ji, R.-R., Zhang, Q., Pettersson, R.F., Hokfelt, T., 1996. aFGF, bFGF and NGF differentially regulate neuropeptide expression in dorsal root ganglia after axotomy and induce autotomy. Reg. Pept. 66, 179-189.]. Finally, NPY expression was also influenced by cell density. PMID- 10869712 TI - Purinergic signalling: an experimental perspective. AB - Investigation of the multiple roles of extracellular nucleotides in the cochlea has developed from analysis of ATP-activated conductances in single sensory hair cells. Molecular probes such as radiolabelled ATP analogues and radiolabelled mRNA for ATP-gated ion channel subunits (P2X receptors) rapidly revealed the extensive nature of ATP signalling in this sensory organ. This has provided a foundation for physiological investigations which put extracellular nucleotides at the centre of homeostatic regulation of the driving force for sound transduction, modulation of mechanical tuning, control of cochlear blood flow and auditory neurotransmission. The purinergic signal transduction pathways associated with these processes have several novel features of significance to the broader field of purinergic neuroscience. In turn, these studies have benefited from the recent experimental advances in the field of purinergic signalling, a significant component of which is associated with the work of Professor Geoffrey Burnstock. PMID- 10869713 TI - P2X receptors mediate ATP-induced primary nociceptive neurone activation. AB - ATP-gated P2X ion-channel receptors are localised throughout the mammalian nervous system and have been identified on neurones which participate in conduction of nociceptive information from the periphery to, and within, the CNS. This article briefly reviews recently published research describing the role that ATP and P2X receptors may play in pain perception, highlighting the importance of the P2X(3) receptor in this process. The P2X(3) receptor subunit is almost exclusively expressed on a subset of small and medium diameter sensory neurones innervating cutaneous and visceral tissue. Activation of P2X receptors present on the peripheral terminals of primary afferents results in neuronal depolarisation and, in conscious animals, leads to the manifestation of acute nociceptive behaviour. Recent animal studies have also shown that P2X(3) receptor expression is increased in sensory ganglia following acute neuronal injury, hinting that similar plasticity in the expression of this receptor subtype could underlie the mechanisms involved in a range of conditions characterised by sensory hypersensitivity in man. It is apparent from the evidence available that functional antagonists at specific P2X receptor subtypes could represent an important class of novel analgesic agents. PMID- 10869715 TI - The discovery and development of P2 receptor subtypes. AB - Extracellular purine and pyrimidine nucleotides modulate cellular activity by acting at P2 receptors. The first receptor to be identified was the P(2) purinoceptor, which was characterised and named in 1978. In the 1980s this site was subdivided into P(2X) and P(2Y) purinoceptors on the basis of pharmacological criteria in functional studies on native receptors. Subsequently, a similar approach led to the characterisation of the P(2T), P(2Z), P(2U) and P(2D) purinoceptors. In the 1990s a molecular biological approach has led to the cloning and functional expression of at least 12 mammalian P2 receptor subtypes. The challenge now is to relate these recombinant receptors to native receptors present within a wide range of tissues. PMID- 10869714 TI - In search of selective P2 receptor ligands: interaction of dihydropyridine derivatives at recombinant rat P2X(2) receptors. AB - 1,4-Dihydropyridines are regarded as privileged structures for drug design, i.e. they tend to bind to a wide variety of receptor sites. We have shown that upon appropriate manipulation of the substituent groups on a 1,4-dihydropyridine template, high affinity and selectivity for the A(3) subtype of adenosine receptors ('P1 receptors') may be attained. In the present study we have begun to extend this approach to P2 receptors which are activated by ATP and other nucleotides. Nicardipine, a representative dihydropyridine, used otherwise as an L-type calcium channel blocker, was shown to be an antagonist at recombinant rat P2X(2) (IC(50)=25 microM) and P2X(4) (IC(50) approximately 220 microM) receptors expressed in Xenopus oocytes. Thus, this class of compounds represents a suitable lead for enhancement of affinity through chemical synthesis. In an attempt to modify the 1,4-dihydropyridine structure with a predicted P2 receptor recognition moiety, we have replaced one of the ester groups with a negatively charged phosphonate group. Several 4-phenyl-5-phosphonato-1,4-dihydropyridine derivatives, MRS 2154 (2, 6-dimethyl), MRS 2155 (6-methyl-2-phenyl), and MRS 2156 (2-methyl-6-phenyl), were synthesized through three component condensation reactions. These derivatives were not pure antagonists of the effects of ATP at P2X(2) receptors, rather were either inactive (MRS 2156) or potentiated the effects of ATP in a concentration-dependent manner (MRS 2154 in the 0.3-10 microM range and MRS 2155 at >1 microM). Antagonism of the effects of ATP at P2X(2) receptor superimposed on the potentiation was also observed at >10 microM (MRS 2154) or 0.3-1 microM (MRS 2155). Thus, while a conventional dihydropyridine, nicardipine, was found to antagonize rat P2X(2) receptors ninefold more potently than P2X(4) receptors, the effects of novel, anionic 5-phosphonate analogues at the receptor were more complex. PMID- 10869716 TI - Recombinant P2Y receptors: the UCL experience. AB - The beginning of the last decade heralded three important and sequential developments in our understanding of cell-to-cell signalling by extracellular ATP via its cell surface receptors, the P2 purinoceptors. One major development in ATP signalling culminated in a timely review in 1991, when it was established in the clearest of terms that ATP receptors exploited discrete signal transduction pathways (Dubyak, G.R., 1991. Signal transduction by P2-purinergic receptors for extracellular ATP. Am. J. Respir. Cell. Mol. Biol. 4, 295-300; and later in Dubyak, G.R., El-Moatassim, C., 1993. Signal transduction via P2-purinergic receptors for extracellular ATP and other nucleotides. Am. J. Physiol. 265, C577 C606). Henceforth, it was universally acknowledged that some P2 purinoceptors interacted with heterotrimeric G-proteins to activate intracellular signalling cascades (metabotropic ATP receptors), whereas others contained intrinsic ion channels (ionotropic ATP receptors). A second key development can be traced to 1992, from the discovery that ATP receptors were involved in excitatory neurotransmission in the CNS and PNS (Edwards, F.A., Gibb, A.J., Colquhoun, D., 1992. ATP receptor-mediated synaptic currents in the central nervous system. Nature 359, 144-147; Evans, R.J., Derkach, V., Surprenant, A., 1992. ATP mediates fast synaptic transmission in mammalian neurons. Nature 357, 503-505; Silinsky, E.M., Gerzanich, V., Vanner, S.M., 1992. ATP mediates excitatory synaptic transmission in mammalian neurones. Br. J. Pharmacol., 106, 762-763). Thereafter, it was accepted that ATP could play a neurotransmitter and/or modulatory role throughout the entire nervous system. The third key development stemmed from the isolation of a cDNA, from chick brain, encoding a metabotropic ATP receptor (Webb, T.E., Simon, J., Krishek, B.J., Bateson, A.N., Smart, T.G., King, B.F., Burnstock, G., Barnard, E.A., 1993. Cloning and functional expression of a brain G-protein-coupled ATP receptor. FEBS Lett. 324, 219-225). The cloning of a membrane protein serving as an ATP receptor ignited a widespread international interest in purinergic signalling. Investigators at University College London (UCL) - colleagues and associates of Geoffrey Burnstock - were at the forefront of this rapid phase of discovery. In this review, we highlight the UCL experience when the fields of molecular biology, physiology and cell biology converged to help advance our understanding of ATP as an extracellular signalling molecule. PMID- 10869717 TI - The novel heteromeric bivalent ligand SB9 potently antagonizes P2Y(1) receptor mediated responses. AB - Effects of 6-[(4,6,8-trisulfo-1-naphthyl)iminocarbonyl-1, 3-(4 methylphenylene)iminocarbonyl-1, 3-phenylene-azo]-pyridoxal-5'-phosphate (SB9), a heterodimeric bivalent ligand consisting of pyridoxal-5'-phosphate and the suramin monomer, were studied on contractions of the rat vas deferens elicited by alpha beta-methylene ATP (alpha beta meATP; mediated by P2X(1)-like receptors), contractions of the guinea-pig ileal longitudinal smooth muscle elicited by adenosine 5'-O-(2-thiodiphosphate) (ADP beta S mediated by P2Y(1)-like receptors), and the degradation of ATP by ecto-nucleotidases in folliculated Xenopus laevis oocytes. SB9 (0.1-10 microM) antagonized contractile responses produced by alpha beta meATP or ADP beta S in a concentration-dependent manner. Schild analysis yielded linear regression lines of unit slope, indicating competitive antagonism. From the rightward shifts of the agonist concentration response curves pA(2) values of 6.05+/-0.13 (vas deferens) and 6.98+/-0.07 (ileum) were derived. In both preparations, SB9 behaved as a slow onset, slow offset antagonist. Incubation of three oocytes in the presence of ATP produced an increase in inorganic phosphate (P(i)) over a 30-min period, which amounted to 35.1+/-1.9 microM P(i) from 100 microM ATP. SB9 (10-1000 microM) reduced this degradation (pIC(50)=4.33+/-0.10). The results illustrate that SB9 is a high affinity P2Y(1) receptor antagonist with a remarkable selectivity for P2Y(1) vs. P2X(1) receptors (about 10-fold) and ecto-nucleotidases (447-fold). These properties make it unique among the pyridoxal-5'-phosphate and suramin derivatives reported to date. PMID- 10869718 TI - Adventures in the pharmacological analysis of P2 receptors. AB - The pharmacological classification of P2 receptors owes its origin to the pioneering efforts of Geoff Burnstock and those who followed him, research that was conducted primarily in physiological experimental systems. Over recent years, the techniques of molecular biology have been increasingly applied in the study of P2 receptors while, at the same time, advances in their pharmacological analysis have been limited by a lack of potent and selective agonist or antagonist ligands. This has resulted in a classification scheme which is largely structural in nature, with relatively little contribution from pharmacology. Our endeavours in this area have been directed towards the discovery of ligands with which the pharmacological analysis and definition of P2 receptors could be advanced, the ultimate goal being the design of therapeutic agents. This article will describe some of our experiences in this challenging but rewarding area. PMID- 10869719 TI - ATP as a peripheral mediator of pain. AB - This article reviews the extent to which recent studies substantiate the hypothesis that ATP functions as a peripheral pain mediator. The discovery of the P2X family of ion channels (for which ATP is a ligand) and, in particular, the highly selective distribution of the P2X(3) receptor within the rat nociceptive system has inspired a variety of approaches to elucidate the potential role of ATP as a pain mediator. ATP elicits excitatory inward currents in small diameter sensory ganglion cells. These currents resemble those elicited by ATP on recombinantly expressed heteromeric P2X(2/3) channels as well as homomultimers consisting of P2X(2) and P2X(3). In vivo behavioural models have characterised the algogenic properties of ATP in normal conditions and in models of peripheral sensitisation. In humans, iontophoresis of ATP induces modest pain. In rats and humans the response is dependent on capsaicin sensitive neurons and is augmented in the presence of inflammatory mediators. Since ATP can be released in the vicinity of peripheral nociceptive terminals under a variety of conditions, there exists a purinergic chain of biological processes linking tissue damage to pain perception. The challenge remains to prove a physiological role for endogenous ATP in activating this chain of events. PMID- 10869720 TI - Presynaptic signalling mediated by mono- and dinucleotides in the central nervous system. AB - Synaptosomal preparations from rat midbrain exhibit specific responses to both ATP and Ap(5)A, which elicit a Ca(2+) entrance to the presynaptic terminals. Studies of isolated single terminals showed that not all the terminals contain ionotropic receptors for nucleotides, in fact only 46% of them do. Of these, 12% responded only to the dinucleotide Ap(5)A, and 20% to the mononucleotide ATP. At the presynaptic level, diinosine pentaphosphate, Ip(5)I, is a good tool to specifically block dinucleotide responses, which are inhibited at low nM concentration, versus the high microM concentrations required to block ATP responses. There is evidence for a presynaptic control of mononucleotide and dinucleotide responses, mediated by metabotropic and ionotropic receptors. Stimulation of adenosine A1 receptors increases the affinity of dinucleotide receptors by five orders of magnitude, from 30 microM to 680 pM for control and in the presence of A1 agonist, respectively. PMID- 10869721 TI - Trophic actions of extracellular ATP: gene expression profiling by DNA array analysis. AB - In addition to Professor Burnstock's work on the short-term signaling actions of extracellular nucleotides and nucleosides, Geoff has had a long-standing interest in trophic actions of purines in development and in pathophysiological conditions which has been instrumental in encouraging my work in this area. The trophic actions of extracellular ATP, alone or in combination with polypeptide growth factors, may play an important role in brain development and may contribute to the reactive gliosis that accompanies brain injury and neurodegeneration. P2Y receptors in astrocytes are coupled to the ERK/MAPK cascade, a signal transduction mechanism crucial for cellular proliferation and differentiation. The mitogenic signaling pathway from P2Y receptors to ERK involves phospholipase D and a calcium-independent PKC isoform, PKCdelta. DNA array analysis reveals a number of changes in gene expression after P2Y receptor occupancy, indicating that this methodology will be a powerful tool in understanding the mechanisms underlying the trophic actions of extracellular nucleotides and nucleosides. PMID- 10869722 TI - P2 receptors in the central and peripheral nervous systems modulating sympathetic vasomotor tone. AB - Arterial pressure depends on the level of activity of sympathetic vasoconstrictor outflow to blood vessels. This activity is generated in the central nervous system, and involves inputs from a variety of brain regions projecting to sympathetic preganglionic neurones. Of especial interest are a group of neurones in the rostral ventrolateral medulla (RVLM), as they have been demonstrated to have a fundamental role in reflex regulation of the cardiovascular system, and in generation of tonic drive to sympathetic outflow. Sympathetic outflow to blood vessels is additionally modulated at sympathetic ganglia, and at the peripheral terminals of sympathetic nerves. This review considers the role of P2 purine receptors in this neural pathway. Ionotropic P2X receptors are expressed in the RVLM, in sympathetic ganglia, and at the sympathetic neuromuscular junction, and mediate fast excitatory neurotransmission, indicating a general role for ATP as a regulator of sympathetic vasomotor tone. P2Y receptors couple to G proteins and mediate slower signalling to ATP; they have been reported to inhibit prejunctionally neurotransmission at the peripheral terminals of sympathetic nerves, but little is known about their possible role in the central nervous system and in sympathetic ganglia. PMID- 10869723 TI - Purinergic nerves and purinoceptors: early perspectives. AB - I have had the pleasure and privilege of being involved in one facet of Geoffrey Burnstock's early career. I have reviewed this work together with more recent developments in the area. In 1968, the presence of non-adrenergic, non cholinergic inhibitory nerves had been established but the identity of their neurotransmitter was unknown. Stimulation of these nerves in recycled perfused toad and guinea-pig stomachs caused release of adenosine and inosine. When ATP was added to recycled perfusates, it was broken down to adenosine and inosine. These findings together with information that AMP was released from stimulated, isolated turkey Auerbach's plexus which was known to contain the nerves, suggested that ATP could be the neurotransmitter. This was supported by observations that ATP elicited responses similar to that of nerve stimulation in a variety of tissues. Developments from the early purinergic nerve hypothesis are considered including independence of extracellular actions of ATP from its intracellular actions, identification and cloning of purinoceptors and cotransmission of ATP with other substances. PMID- 10869724 TI - Electrophysiology of autonomic neuromuscular transmission involving ATP. AB - Electrophysiological investigations of autonomic neuromuscular transmission have provided great insights into the role of ATP as a neurotransmitter. Burnstock and Holman made the first recordings of excitatory junction potentials (e.j.p.s) produced by sympathetic nerves innervating the smooth muscle of the guinea-pig vas deferens. This led to the identification of ATP as the mediator of e.j.p.s in this tissue, where ATP acts as a cotransmitter with noradrenaline. The e.j.p.s are mediated solely by ATP acting on P2X(1) receptors leading to action potentials and a rapid phasic contraction, whilst noradrenaline mediates a slower, tonic contraction which is not dependent on membrane depolarisation. Subsequent electrophysiological studies of the autonomic innervation of smooth muscles of the urogenital, gastrointestinal and cardiovascular systems have revealed a similar pattern of response, where ATP mediates a fast electrical and mechanical response, whilst another transmitter such as noradrenaline, acetylcholine, nitric oxide or a peptide mediates a slower response. The modulation of junction potentials by a variety of pre-junctional receptors and the mechanism of inactivation of ATP as a neurotransmitter will also be described. PMID- 10869725 TI - Antagonists and the purinergic nerve hypothesis: 2, 2'-pyridylisatogen tosylate (PIT), an allosteric modulator of P2Y receptors. A retrospective on a quarter century of progress. AB - 2,2'-Pyridylisatogen tosylate (PIT) is a selective antagonist of P2Y responses in smooth muscle and does not antagonise the effects of adenosine. Responses to purinergic nerve stimulation are resistant to PIT. PIT is an allosteric modulator of responses to ATP in recombinant P2Y(1) receptors expressed in Xenopus oocytes with potentiation of ATP at low concentrations (0.1-10 microM) and antagonism at higher ones (>10 microM). A radioligand binding profile showed that PIT did not interact with any other receptors, with the exception of low affinity for the adenosine A(1) receptor (pK(i), 5.3). The compound recognises purine sites and then may cause irreversible binding to sulfhydryl groups following prolonged incubation or high concentrations. PIT is a potent spin trapper. PMID- 10869726 TI - Sensing arterial CO(2) levels: a role for medullary P2X receptors. AB - ATP has been shown to act as an excitatory neurotransmitter in the central nervous system. In this review, evidence is presented to indicate that when ATP is micro-injected into the ventrolateral medulla (VLM) of the rat, changes in respiratory activity are elicited. These effects, and accompanying changes in heart rate and blood pressure are mediated by P2X purinoreceptors. Immunocytochemistry indicates a prevalence of P2X(2) and P2X(6) purinoreceptors in this region of the medulla. The P2 purinoceptor antagonists, suramin and PPADS blunt the respiratory responses to changes in arterial CO(2) levels when micro injected into the VLM. This effect is shown electrophysiologically to be mediated by purinoreceptors located primarily on respiratory neurones of the VLM including the Botzinger complex. As the effects of agonist activation of P2X(2) purinoceptors expressed in HEK293 cells and Xenopus oocytes are potentiated by lowering pH, these data imply that the central respiratory response to CO(2) depends in part on the pH sensitivity of purinoreceptors located on inspiratory neurones. The implications for respiratory activity and control are discussed. PMID- 10869727 TI - Do sympathetic nerves release noradrenaline in "quanta"? AB - The discovery of excitatory junction potentials (EJPs) in guinea-pig vas deferens by Burnstock and Holman (1960) showed for the first time that a sympathetic transmitter, now known to be ATP, is secreted in "quanta". As it was assumed at the time that EJPS are triggered by noradrenaline, this discovery led to attempts to use the fractional overflow of noradrenaline from sympathetically innervated tissues to assess, indirectly, the number of noradrenaline molecules in the average "quantum". The basic finding was that each pulse released 1/50000 of the tissue content of noradrenaline, when reuptake was blocked and prejunctional alpha(2)-adrenoceptors were intact. This provided the constraints, two extreme alternatives: (i) each pulse releases 0.2-3% of the content of a vesicle from all varicosities, or (ii) each pulse releases the whole content of a vesicle from 0.2 to 3% of the varicosities. New techniques have made it possible to address questions about the release probability in individual sites, or the "quantal" size, more directly. Results by optical (comparison of the labelling of SV2 and synaptotagmin, proteins in the membrane of transmitter vesicles), electrophysiological (excitatory junction currents, EJCs, at single visualized varicosities) and amperometric (the noradrenaline oxidation current at a carbon fibre electrode) methods reveal that transmitter exocytosis in varicosities is intermittent. The EJC and noradrenaline oxidation current responses (in rat arteries) to a train of single pulses were observed to be similar in intermittency and amplitude fluctuation. This suggests that they are caused by exocytosis of single or very few "quanta" of ATP and noradrenaline, respectively, equal to the contents of single vesicles, from a small population of release sites. These findings support, but do not conclusively prove the validity of the "intermittent" model of noradrenaline release. The question if noradrenaline is always secreted in packets of preset size ("quanta") and if the "quantum" is a subfraction or the whole content of single synaptic vesicles, still remains open. PMID- 10869728 TI - Presynaptic P2 receptors? AB - Although the emphasis in ATP research has been on postjunctional receptors, there is also evidence for presynaptic receptors regulating transmitter release in the autonomic nervous system. Recent work has attempted to identify similar mechanisms in the central nervous system. Some of the existing results can be explained by the metabolism of nucleotides to adenosine or adenosine 5' monophosphate (AMP). However, studies of presynaptic effects using sensitive electrophysiological tests such as paired-pulse interactions indicate that nucleotides can act at presynaptic sites, but that their effects may be mediated by a release of adenosine. Results are also described which indicate that, under some conditions, nucleotides can mediate phenomena such as long-term potentiation, which probably involves a significant presynaptic element. In part these effects may involve a nucleotide-induced release of adenosine and the simultaneous activation of P1 and P2 receptors. PMID- 10869729 TI - Functional properties of heteromeric P2X(1/5) receptors expressed in HEK cells and excitatory junction potentials in guinea-pig submucosal arterioles. AB - P2X receptors are ATP-gated cation channels; they form as homomers or heteromers from a family of seven related subunits. In particular, heteromeric channels comprising P2X(2) and P2X(3) subunits, or P2X(1) and P2X(5) subunits, show distinctive physiological and pharmacological properties in heterologous expression systems. There is substantial evidence that one of the native P2X receptors in sensory neurones corresponds to the P2X(2/3) heteromer, but there is no evidence for P2X(1/5) heteromers in native tissue. We recorded currents in response to activation of heteromeric P2X(1/5) receptors expressed in HEK293 cells to characterize further their functional properties. The ATP concentration response curve had a threshold concentration of 1 nM, and a Hill slope of one. TNP-ATP was a weak partial agonist, and a non-competitive antagonist which inhibited maximal ATP currents by 60%. Increasing or decreasing pH from 7.3 shifted the ATP concentration-response curves to the right by fivefold and decreased the maximum current by 40%. Calcium permeability was lower than that observed for other P2X receptors (P(Ca)/P(Na) ratio=1.1). The nanomolar sensitivity of this receptor revealed a steady release of ATP from HEK293 cells, providing an extracellular concentration which ranged from 3 to 300 nM. Noradrenaline (0.3-30 microM) increased ATP-evoked currents by 35%; this facilitation occurred within 20 ms. We also recorded excitatory junction potentials (EJPs) from guinea-pig submucosal arterioles. EJPs were inhibited by suramin and PPADS (IC(50)s of 0.2 microM and 20 microM) but TNP-ATP (0.1-10 microM) inhibited EJPs by <30%. Noradrenaline (0.3-30 microM in the presence of phentolamine and propranolol) decreased EJPs in control preparations but facilitated EJPs by 5-20% in submucosal arterioles from reserpinized guinea-pigs. These properties are discussed in relation to P2X receptors underlying EJPs at autonomic neuroeffector junctions. PMID- 10869730 TI - P2 receptors in the kidney. AB - Our understanding of the actions of extracellular ATP in controlling kidney function via stimulation of P2 receptors is still at an early stage. Recently, several groups, including our own, have begun to address this subject: in this brief review, we discuss some of these effects and speculate on likely function of extracellular nucleotides in the kidney. PMID- 10869731 TI - Say NO to ET. AB - The endothelial cells release both relaxing [nitric oxide (NO), endothelium derived hyperpolarizing factor (EDHF), prostacyclin] and contracting factors [endoperoxides, thromboxane A(2), superoxide anions, endothelin-1 (ET)]. The production of ET is inhibited by NO. The latter also strongly opposes the direct effects of the former on vascular smooth muscle. With aging and vascular disease, the production of enothelial NO declines, and thus ET can be released, act and contribute to the symptoms. PMID- 10869732 TI - Stimulation-dependent release, breakdown, and action of endogenous ATP in mouse hemidiaphragm preparation: the possible role of ATP in neuromuscular transmission. AB - In this study the in vitro mouse phrenic nerve- hemidiaphragm preparation was utilized to study the release and extracellular catabolism of endogenous ATP and its action on the postsynaptic site, i.e. on the contraction force evoked by nerve stimulation. ATP, measured by the luciferin-luciferase assay, was released stimulation-dependently from the mouse hemidiaphragm in response to electrical field stimulation at 10 Hz. Blockade of the Na(+) channel activity by tetrodotoxin inhibited the majority of the release of ATP in response to stimulation, showing that it is related to neuronal activity. The nicotinic receptor antagonists d-tubocurarine, and alpha-bungarotoxin and cooling the bath temperature to 7 degrees C also reduced stimulation-induced ATP outflow, suggesting that nicotinic receptors are responsible for the part of the release of ATP that is released from postsynaptic sites in a carrier-mediated manner. Exogenous ATP (20-500 microM) added to the bath was degraded to ADP and AMP by the action of ectoATPase and ectoATPdiphosphohydrolase; the K(m) and v(max) values of these enzymes were 185.8 microM and 55.16 nmol/min.g respectively. However, the total amount of nucleotides ([ATP+ADP+AMP]) was increased after the addition of ATP, indicating that ATP itself promoted further adenine nucleotide release. Twitch contractions of the rat hemidiaphragm preparation evoked by low frequency electrical stimulation was blocked concentration-dependently by the non depolarizing muscle relaxants d-tubocurarine and pancuronium. Suramin (100 microM 1 mM) reversed neuromuscular blockade by d-tubocurarine and pancuronium; i.e., it shifted their concentration-response curves to the right Taken together our data, that endogenous ATP is released by stimulation and subsequently catabolized in the hemidiaphragm preparation and that suramin inhibits ecto-ATPase activity could be interpreted as meaning that suramin prolongs the action of endogenous ATP to elicit twitch contraction, which points to a new, undefined role of ATP in neuromuscular transmission. The source of ATP is partly postsynaptic, released from the muscle in response to activation of nicotinic ACh receptors expressed on the muscle. PMID- 10869733 TI - Purines: from premise to promise. AB - Geoff Burnstock's remarkable insight and tenacity has established the area of purinergic research as a bona fide target for drug discovery. While efforts in P1 receptor-based medicinal chemistry and biology efforts over the past 25 years have not reached the level of success that the pharmaceutical industry investment may have anticipated, the P2 area, with knowledge of the selective localization of members of the P2X and P2Y family members and data from transgenic knockouts, has identified several potential therapeutic areas of major promise including cystic fibrosis, chronic bronchitis, male contraception and neurodegeneration. In addition, interest in the potential of purinergic therapeutics has extended outside the major pharmaceutical companies to the 'biotech industry' resulting in an environment where the inherent risks of 'first in field' in a therapeutic area may be more appropriately nurtured. PMID- 10869734 TI - ATP, P2X receptors and pain pathways. AB - A role for ATP in nociception and pain induction was proposed on the basis of human psychophysical experiments shortly after the formulation of the purinergic hypothesis. Following the pharmacological definition of distinct P2X and P2Y purinergic receptor subtypes by Burnstock and his collaborators, molecular cloning studies have identified the gene products that underlie the effects of ATP on peripheral sensory neurons. One particular receptor, P2X(3), is of particular interest in the context of pain pathways, because it is relatively selectively expressed at high levels by nociceptive sensory neurons. Evidence that this receptor may play a role in the excitation of sensory neurons has recently been complemented by studies that suggest an additional presynaptic role in the regulation of glutamate release from primary afferent neurons in the dorsal horn of the spinal cord. In this brief review, we discuss the present state of knowledge of the role of ATP in pain induction through its action on peripheral P2X receptors. PMID- 10869735 TI - A tribute to the founding editor PMID- 10869737 TI - Factors in decision making in the treatment of breast cancer PMID- 10869736 TI - FECS Clinical Award lecture 1989. Factors in decision making in the treatment of breast cancer. Federation of European Cancer Societies. PMID- 10869738 TI - The influence of patient, tumor and treatment factors on the cosmetic results after breast-conserving therapy in the EORTC 'boost vs. no boost' trial. EORTC Radiotherapy and Breast Cancer Cooperative Groups. AB - PURPOSE: To analyze the influence of different patient, tumor, and treatment parameters on the cosmetic outcome after breast-conserving therapy at 3-year follow-up. A subjective and an objective cosmetic scoring method was used and the results of both methods were compared. PATIENTS AND METHODS: In EORTC trial 22881/10882, 5569 early-stage breast cancer patients were treated with tumorectomy and axillary dissection, followed by tangential fields irradiation of the breast to a dose of 50 Gy in 5 weeks, at 2 Gy per fraction. A total of 5318 patients, having a microscopically complete tumorectomy, were randomized between no further treatment and a boost of 16 Gy to the primary tumor bed. The cosmetic result at 3-year follow-up was assessed by a panel for 731 patients, and by digitizer measurements, measuring the displacement of the nipple, for 1141 patients. Univariate and multivariate analyses were used to evaluate the correlation between various patient, tumor, and treatment factors and cosmesis. RESULTS: The factors associated with a worsened cosmesis according to the panel evaluation were: an inferior tumor location, a large excision volume, the presence of postoperative breast complications, and the radiotherapy boost. According to the digitizer measurements, a central/superior tumor location, a large excision volume, an increased pathological tumor size, an increased radiation dose inhomogeneity, and an increased bra cup size resulted in an increased asymmetry in nipple position. It appeared that the evaluation of the nipple position (whether by panel or by digitizer) is only moderately representative of the overall cosmetic outcome. CONCLUSION: To achieve a good cosmesis, it is necessary to excise the tumor with a limited margin, to avoid postoperative complications, to assess the need for a boost in the individual patient, and to give the radiation dose as homogeneously as possible. As far as the method of evaluation is concerned, the panel evaluation is the most appropriate method for giving an overall impression of the cosmetic result after breast-conserving therapy (BCT). The use of the digitizer is recommended for comparing the cosmetic outcome of two different approaches to BCT or for analyzing cosmetic changes over time. PMID- 10869739 TI - Clinical experience with intensity modulated radiation therapy (IMRT) in prostate cancer. AB - PURPOSE: To compare acute and late toxicities of high-dose radiation for prostate cancer delivered by either conventional three-dimensional conformal radiation therapy (3D-CRT) or intensity modulated radiation therapy (IMRT). MATERIALS AND METHODS: Between September 1992 and February 1998, 61 patients with clinical stage T1c- T3 prostate cancer were treated with 3D-CRT and 171 with IMRT to a prescribed dose of 81 Gy. To quantitatively evaluate the differences between conventional 3D-CRT and IMRT, 20 randomly selected patients were planned concomitantly by both techniques and the resulting treatment plans were compared. Acute and late radiation-induced morbidity was evaluated in all patients and graded according to the Radiation Therapy Oncology Group toxicity scale. RESULTS: Compared with conventional 3D-CRT, IMRT improved the coverage of the clinical target volume (CTV) by the prescription dose and reduced the volumes of the rectal and bladder walls carried to high dose levels (P<0.01), indicating improved conformality with IMRT. Acute and late urinary toxicities were not significantly different for the two methods. However, the combined rates of acute grade 1 and 2 rectal toxicities and the risk of late grade 2 rectal bleeding were significantly lower in the IMRT patients. The 2-year actuarial risk of grade 2 bleeding was 2% for IMRT and 10% for conventional 3D-CRT (P<0.001). CONCLUSIONS: The data demonstrate the feasibility and safety of high-dose IMRT for patients with localized prostate cancer and provide a proof-of-principle that this method improves dose conformality relative to tumor coverage and exposure to normal tissues. PMID- 10869740 TI - Improved management of radiotherapy departments through accurate cost data. AB - BACKGROUND AND PURPOSE: Escalating health care expenses urge governments towards cost containment. More accurate data on the precise costs of health care interventions are needed. We performed an aggregate cost calculation of radiation therapy departments and treatments and discussed the different cost components. MATERIALS AND METHODS: The costs of a radiotherapy department were estimated, based on accreditation norms for radiotherapy departments set forth in the Belgian legislation. RESULTS: The major cost components of radiotherapy are the cost of buildings and facilities, equipment, medical and non-medical staff, materials and overhead. They respectively represent around 3, 30, 50, 4 and 13% of the total costs, irrespective of the department size. The average cost per patient lowers with increasing department size and optimal utilization of resources. Radiotherapy treatment costs vary in a stepwise fashion: minor variations of patient load do not affect the cost picture significantly due to a small impact of variable costs. With larger increases in patient load however, additional equipment and/or staff will become necessary, resulting in additional semi-fixed costs and an important increase in costs. A sensitivity analysis of these two major cost inputs shows that a decrease in total costs of 12-13% can be obtained by assuming a 20% less than full time availability of personnel; that due to evolving seniority levels, the annual increase in wage costs is estimated to be more than 1%; that by changing the clinical life-time of buildings and equipment with unchanged interest rate, a 5% reduction of total costs and cost per patient can be calculated. More sophisticated equipment will not have a very large impact on the cost (+/-4000 BEF/patient), provided that the additional equipment is adapted to the size of the department. That the recommendations we used, based on the Belgian legislation, are not outrageous is shown by replacing them by the USA Blue book recommendations. Depending on the department size, costs in our model would then increase with 14-36%. CONCLUSION: We showed that cost information can be used to analyze the precise financial consequences of changes in routine clinical practice in radiotherapy. Comparing the cost data with the prevailing reimbursement may reveal inconsistencies and stimulate to develop improved financing systems. PMID- 10869741 TI - Adjuvant radiotherapy for breast cancer significantly improves overall survival: the missing link. AB - BACKGROUND AND PURPOSE: The influence of surgical adjuvant radiotherapy on overall survival of patients with operable breast cancer is still a controversial subject. The negative result of the EBCTCG meta-analysis (Early breast cancer trialists', collaborative group. Effects of radiotherapy and surgery in early breast cancer. An overview of the randomised trials. N. Engl. J. Med. 1995;333:1444-1455) of clinical randomized trials on adjuvant radiotherapy in breast cancer is in strong contrast with the Danish 82B, 82C and British Columbia trials (Overgaard M, Hanse PS, Overgaar J, et al. Postoperative radiotherapy in high-risk premenopausal women with breast cancer who receive adjuvant chemotherapy. Danish Breast Cancer Cooperative Group 82b Trial. N. Engl. J. Med. 1997;337:949-955; Overgaard M, Jensen MB, Overgaard J, et al. Postoperative radiotherapy in high-risk postmenopausal breast-cancer patients given adjuvant tamoxifen: Danish Breast Cancer Cooperative Group DBCG 82c randomized trial. Lancet 1999;353:1641-1648; Ragaz J, Jackson S, Le N, et al. Adjuvant radiotherapy and chemotherapy in node-positive premenopausal women with breast cancer. N. Engl. J. Med. 1997;337:956-962) showing an impressive survival benefit. This paper tries to fill in the gap between the conflicting results. MATERIALS AND METHODS: The 36 trials of the EBCTCG (Early breast cancer trialists', collaborative group, 1995) were prospectively screened for a number of objective parameters that are usually not analyzed in review papers. The odds of death data (and its variance) were borrowed from the original meta-analysis (Early breast cancer trialists', collaborative group, 1995) to check whether the objective features were significant predictors for overall survival benefit. RESULTS: A significant survival benefit for the radiotherapy arm was found for recent trials (2P<0.05), large trials (2P<0.03), trials that used standard fractionation (2P<0.02), and trials with a favourable crude survival (2P<0.03). For these four parameters clear parameter-effect relations were found. In recent and large trials the odds reduction was 12.4% (2P=0.004). CONCLUSIONS: Surgical adjuvant radiotherapy significantly improves overall survival of breast cancer patients provided that current techniques are used and treatment is given with standard fractionation. For the best subgroups we observed an odds of death reduction of more than 20%. The results of this study stress the importance of reducing cardiovascular and other late toxicity in adjuvant radiotherapy for breast cancer. PMID- 10869742 TI - The ESTRO-QUALity assurance network (EQUAL). AB - BACKGROUND AND PURPOSE: ESTRO has set up a Quality Assurance network (EQUAL) to check the dose delivered on axis in reference and non-reference conditions for external radiotherapy. The external audits covered by the network are based on measurements made with mailed thermoluminescent dosimeters (TLD). MATERIAL AND METHODS: The TLD consist of LiF powder type DTL 937 read with a PCL 3 automatic TLD reader. The participating centres are instructed to deliver to the TLDs absorbed doses of 2 Gy calculated with the Treatment Planning System used in clinical routine. A maximum of three photon energies by participating centre have been checked with 10 on-axis points per beam. The quantities checked include the reference beam output, beam output variation with collimator opening, depth dose data and wedge transmission factor. RESULTS: During the 1998 EQUAL programme 102 centres have been checked corresponding to 235 beams (28 (60)Co beams and 207 X ray beams). About 3% of the outputs in reference conditions show deviations outside tolerance level (>+/-5%). A similar rate of deviation is noted for the percentage depth doses. A rate of deviation (6%) has been observed for the beam output variation (open and wedged beams) and the wedge transmission factor. The analysis of the results shows that for 24 out of the 102 centres, a deviation outside tolerance level is observed at least in one point, mainly for the large and rectangular field sizes and for the wedged beams. CONCLUSIONS: The results for the EQUAL programme show the importance of a quality assurance network in Radiotherapy especially for the non reference points even if they are only located on the beam axis (In order to participate in this network, please contact EQUAL secretariat or download the attached application form ESTRO web site: Dr I.H. Ferreira or Mrs Aline Mechet, EQUAL-ESTRO, Physics Department, Institut Gustave-Roussy 39 Rue Camille Desmoulins, F-94805 Villejuif Cedex, France. e mail:equal@igr.fr or http://www.estro.be/). PMID- 10869743 TI - A comparative description of three multipurpose phantoms (MPP) for external audits of photon beams in radiotherapy: the water MPP, the Umea MPP and the EC MPP. AB - AIM: To present a technical description and intercomparison of three multipurpose phantoms (MPP) developed for mailed dosimetry checks of therapeutic photon beams in reference and non-reference conditions. MATERIALS: The W-MPP is a water MPP, whereas the Umea-MPP, made of perspex (PMMA, Plexiglas), and the EC-MPP, made of polystyrene, are solid MPPs. The W-MPP uses only thermoluminescent dosimeters (TLD) for dosimetry checks, the EC MPP uses film and TLD; the Umea phantom uses film and TLD, and offers in addition the possibility for ionization chamber measurements. Either using TLD or films, the MPPs have been designed to check on axis and off-axis the following irradiation conditions: square and rectangular fields, asymmetric fields, wedged beams, oblique incidence and, for the solid MPPs, also the influence of inhomogeneities. RESULTS AND DISCUSSION: The MPPs have been compared for different aspects going from their dosimetric performance (number of dosimetric parameters that can be checked) to some practical consideration in the use of the different MPPs (set-up time, stability, instruction sheets, etc.). From a comparison between the solid multi-purpose phantoms, it turns out that the EC-MPP is capable of checking the largest number of dosimetric parameters per beam, but has the longest set-up time ( approximately 2 h) per beam according to the users. The Umea-MPP has a smaller number of set-ups (hence a smaller average time) and also includes some parameters not checked with the EC-MPP (e.g. SSD accuracy). The major drawback however of the Umea-MPP is considered to be its high density (>1.1 g/cm(3)) which increases the difficulty of the analysis with the treatment planning system. The W-MPP checks the smallest number of parameters, but is the fastest in set-up time, the easiest for mailing, and is water equivalent, which is advantageous for the TPS checks. The major drawback of this MPP is however the inability to check complete dose distribution (film) or inhomogeneities. PMID- 10869744 TI - Clinical radiation doses for spinal cord: the 1998 international questionnaire. AB - BACKGROUND AND PURPOSE: Emmanuel van der Schueren gave a keynote lecture at the 1988 ASTRO annual conference pointing out that the spinal cord 'tolerance doses' then prescribed were probably unnecessarily cautious, resulting in probable underdosing of some tumours. This lecture was supported both by an international questionnaire which he and two of the present authors had conducted, and by animal experimental data. In 1997 he initiated a 10-year follow-up questionnaire, the results of which are summarised here. MATERIALS AND METHODS: The present report analyses the change in prescriptions from 1988 to 1998 and the variation in prescriptions among various regions of the World. RESULTS AND CONCLUSIONS: The main conclusion is that prescribed dose levels have increased significantly in this period. Large geographical variations still exist. Among responders who use a formula to correct for changed dose per fraction, 90% are now using the linear quadratic model vs. 33% in 1988. The current status of clinically acceptable doses to spinal cord in 2-Gy fractions is discussed briefly. Further details from the responses to the 1998 questionnaire will be presented in another publication. PMID- 10869745 TI - Radiation tolerance of rat spinal cord to pulsed dose rate (PDR-) brachytherapy: the impact of differences in temporal dose distribution. AB - PURPOSE: To investigate the impact of a time-variable dose rate during a high dose rate (HDR-) or pulsed dose rate (PDR-) brachytherapy fraction with the HDR microSelectron and to compare this with the biological effect of a constant dose rate treatment with the same average dose rate (as in the case of (192)Ir-wires). Moreover, the kinetics of repair in rat spinal cord are investigated using a wide spectrum of temporal dose distributions. MATERIALS AND METHODS: Two parallel catheters are inserted on each side of the vertebral bodies of the rat spinal column (Th(10)-L(4)) and connected to the HDR-microSelectron. Interstitial irradiation (IRT) is performed with a stepping (192)Ir-point source, varying the activity of the point source between 0.3 and 6.5 Ci. Three different groups of experiments are defined, varying the overall treatment time and average dose rates in the range of 3-8, 28-53 and 82-182 min and 312-489 Gy/h, 32-56 Gy/h and 13-15 Gy/h, respectively. Difference in temporal dose distribution (dose rate variation) during almost the same overall treatment time is obtained by varying the number of pulses per dwell position in either one or ten runs through the implant. For reasons of comparison, previously reported results of continuous irradiation at a constant dose rate obtained with two (192)Ir-wires in a fixed position are reanalyzed. Paralysis of the hindlegs after 5-6 months and histopathological examination of the spinal cord of each animal are used as experimental endpoints. RESULTS: During one run of the (192)Ir-point source, the peak dose rate is at least 25 times higher as compared with the minimum local dose rate and almost four times higher as compared with the average dose rate. For the three different groups of varying overall treatment times and average dose rates there is a significant difference in biological effect, with an ED(50) value of 23.1-23.6 Gy (average dose rate 312-489 Gy/h), 25.4-27.9 Gy (average dose rate 312-489 Gy/h) and 29.3-33 Gy (average dose rate 13-15 Gy/h). For these range of single doses, difference in temporal dose distribution with either one or ten runs is only significant for treatment times less then 1 h. For the prolonged treatment times at lower average dose rates, the difference between one or ten run is no longer significant. However, the results with the (192)Ir-point source at an average dose rate/run of 13-15 Gy/h are significantly different from the ED(50)-value of 33 Gy using (192)Ir-wires at the same but constant dose rate. Using different types of analysis to estimate the repair parameters, the best fit of the data is obtained assuming biphasic repair kinetics and a variable dose rate (geometrically dependent) for the (192)Ir-point source. On the basis of the incomplete repair LQ model, two repair processes with an alpha/beta ratio=2.47 Gy and repair halftimes of 0.19 and 2.16 h are detected. The partition coefficient for the longer repair process is 0.98. This results in the proportion of total damage associated with the longer repair halftime being 0.495 for short sharp fractions with complete repair in between. CONCLUSIONS: Even in the range of high dose rates of 15-500 Gy/h, spinal cord radiation tolerance is significantly increased by a reduction in dose rate. For larger doses per fraction in PDR brachytherapy dose rate variation is important, especially for tissues with very short repair half times (components). In rat spinal cord the repair of sublethal damage (SLD) is governed by a biphasic repair process with repair halftimes of 0.19 and 2.16 h. PMID- 10869746 TI - The impact of (18)F-fluoro-2-deoxy-D-glucose positron emission tomography (FDG PET) lymph node staging on the radiation treatment volumes in patients with non small cell lung cancer. AB - PURPOSE: (18)F-fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) combined with computer tomography (PET-CT) is superior to CT alone in mediastinal lymph node (LN) staging in non-small cell lung cancer (NSCLC). We studied the potential impact of this non-invasive LN staging procedure on the radiation treatment plan of patients with NSCLC. PATIENTS AND METHODS: The imaging and surgical pathology data from 105 patients included in two previously published prospective LN staging protocols form the basis for the present analysis. For 73 of these patients, with positive LN's on CT and/or on PET, a theoretical study was performed in which for each patient the gross tumour volume (GTV) was defined based on CT and on PET-CT data. For each GTV, the completeness of tumour coverage was assessed, using the available surgical pathology data as gold standard. A more detailed analysis was done for the first ten consecutive patients in whom the PET-CT-GTV was smaller than the CT-GTV. Theoretical radiation treatment plans were constructed based on both CT-GTV and PET-CT-GTV. Dose-volume histograms for the planning target volume (PTV), for the total lung volume and the lung volume receiving more than 20 Gy (V(lung(20))), were calculated. RESULTS: Data from 988 assessed LN stations were available. In the subgroup of 73 patients with CT or PET positive LN's, tumour coverage improved from 75% when the CT-GTV was used to 89% with the PET-CT-GTV (P=0.005). In 45 patients (62%) the information obtained from PET would have led to a change of the treatment volumes. For the ten patients in the dosimetry study, the use of PET-CT to define the GTV, resulted in an average reduction of the PTV by 29+/-18% (+/-1 SD) (P=0.002) and of the V(lung(20)) of 27+/-18% (+/-1 SD) (P=0.001). CONCLUSION: In patients with NSCLC considered for curative radiation treatment, assessment of locoregional LN tumour extension by PET will improve tumour coverage, and in selected patients, will reduce the volume of normal tissues irradiated, and thus toxicity. This subgroup of patients could then become candidates for treatment intensification. PMID- 10869747 TI - Diffusion limited hypoxia estimated by vascular image analysis: comparison with pimonidazole staining in human tumors. AB - PURPOSE: To assess diffusion limited hypoxia in human tumors using image analysis of vasculature and to compare it with the bioreductive marker pimonidazole as an independent method. MATERIALS AND METHODS: To set up the method, nine rectal adenocarcinomas and ten squamous cell carcinomas were analyzed. To validate the method, ten squamous cell carcinomas of the cervix were analyzed from patients who were injected with pimonidazole and biopsied approximately 24 h later. Sections of the rectal and esophageal tumors were stained for vasculature, while cervix tumor sections were double stained for vasculature and pimonidazole. Tumor areas were delineated on digitized images, and the proportion of tumor tissue greater than a fixed distance from the nearest blood vessel (called diffusion limited fraction, DLF) was then calculated. The proportion of tumor area stained for pimonidazole was also measured. RESULTS: There was a wide variation between tumors in both the vascular-derived DLF and in the pimonidazole-stained fraction. Average DLFs varied between 1.5 and 92% for different tumors, with significant differences between them. The area stained by pimonidazole was significantly smaller than DLF for all tumors. The correlation between pimonidazole area and DLF was significant in three of seven tumors containing > or = 3 images. When images from all tumors (n=123) were analyzed together, the correlation was highly significant (r=0.47, P<0.0001). CONCLUSION: The vascular derived DLF correlates significantly with pimonidazole staining, but there was large scatter. Both methods may underestimate perfusion limited hypoxia. PMID- 10869748 TI - The role of low-dose total body irradiation in treatment of non-Hodgkin's lymphoma: a new look at an old method. AB - The use of low-dose total body irradiation (LTBI) in treatment of lymphomatous malignancies dates back to the 1920s. The usual practice was to give very low individual TBI fraction sizes (0.1-0. 25 Gy) several times a week to a total dose of 1.5-2 Gy. Despite this very low total dose, LTBI could induce long term remissions and was always as effective as the chemotherapy to which it was compared. In modern radiotherapy, LTBI is still a valid option in treatment of chronic lymphocytic leukaemia (CLL) and the advanced stages of indolent low-grade non-Hodgkin's lymphoma (NHL). Its use in the early stages of low-grade NHL is under investigation in a large multi-institutional trial. The efficacy of LTBI is believed to stem from three mechanisms, namely; immune-enhancement, induction of apoptosis, and the intrinsic hypersensitivity to low-radiation doses demonstrated in many cell lines and tumour systems. Thus, LTBI seems to provide 'alternative' mechanisms of action against cancer cells. This should encourage researchers to explore strategies that integrate LTBI in new and innovative experimental treatment protocols that explore the possible synergism between LTBI and chemotherapy, biological response modifiers and/or immunotherapy. The increased incidence of secondary leukaemia that occurs when LTBI is combined with alkylating agents and/or total lymphoid irradiation should be kept in mind when designing such protocols as it may limit the use of LTBI in highly curable diseases and young patients in whom long survival is expected. PMID- 10869749 TI - Radiotherapy in the management of cervical cancer in elderly patients. AB - PURPOSE: To report treatment results and complications experienced by elderly patients treated with curatively intended radiotherapy for cancer of the uterine cervix. PATIENTS AND METHODS: One hundred and fourteen elderly patients (median 75.5 years, range 70.0-85.9) consecutively referred for curative radiotherapy in the period 1987-1996 were prospectively followed with regard to tumour control and complications. The importance of age, stage (FIGO), tumour size, histology, tumour fixation, haemoglobin, concurrent disease, performance status (WHO) and type of radiotherapy were assessed using univariate and multivariate analyses. RESULTS: Treatment was completed as planned in 68%, delayed in 29% and stopped prematurely in 3%. The frequency of grade 3 late complications was 11% and the actuarial probability at 5 years was 20%. Overall 5-year survival according to FIGO was 61% (I), 34% (II) and 25% (III). Cox multivariate analysis identified tumour size as independent prognostic factor for tumour control, disease-free survival and overall survival. FIGO stage was predictive for late grade 2 complications. We were unable to identify significant factors with respect to grade 3 complications. Age was not a significant parameter for any of the investigated endpoints. CONCLUSION: Elderly patients in good performance status with advanced cancer of the uterine may tolerate radical radiotherapy with acceptable morbidity and reasonable survival. Radiotherapy may also be a good alternative in early stage disease for surgically unfit elderly patients. PMID- 10869750 TI - Post-operative high dose rate brachytherapy in patients with low to intermediate risk endometrial cancer. AB - BACKGROUND AND PURPOSE: This paper investigates the outcome using different dose/fractionation schedules in high dose rate (HDR) post-operative vaginal vault radiotherapy in patients with low to intermediate risk endometrial cancer. MATERIALS AND METHODS: The world literature was reviewed and thirteen series were analyzed representing 1800 cases. RESULTS: A total of 12 vaginal vault recurrences were identified representing an overall vaginal control rate of 99.3%. A wide range of dose fractionation schedules and techniques have been reported. In order to analyze a dose response relationship for tumor control and complications, the biologically effective doses to the tumor and late responding tissues were calculated using the linear quadratic model. A threshold was identified for complications, but not vaginal control. While dose fractionation schedules that delivered a biologically effective dose to the late responding tissues in excess of 100 Gy(3) (LQED=60 Gy) predicted for late complications, dose fractionation schedules that delivered a modest dose to the vaginal surface (50 Gy(10) or LQED=30 Gy) appeared tumoricidal with vaginal control rates of at least 98%. CONCLUSIONS: By using convenient, modest dose fractionation schedules, HDR vaginal vault - brachytherapy yields very high local control and extremely low morbidity rates. PMID- 10869751 TI - Pelvic fractures following irradiation of endometrial and vaginal cancers-a case series and review of literature. AB - PURPOSE: To review the induction of pelvic fractures as a result of radiation therapy and to assess their management. MATERIALS AND METHODS: The charts of patients with endometrial and vaginal cancers irradiated between 1991 and 1995 were reviewed. All patients were treated with megavoltage machines, energy ranging from cobalt to 25 MV photons. RESULTS: We treated 336 patients, with a median follow-up duration of 28.9 months (range 0-73.3). Sixteen patients had symptomatic pelvic fractures. The 5-year actuarial incidence of symptomatic pelvic fracture was 2.1%. All patients had pain as the first symptom. The median time of onset was 11 months (range 4-46). Imaging studies of 37.5% (6/16) were initially interpreted to be recurrent malignancy. All patients were managed conservatively and nine patients showed radiological evidence of healing over a median time of 13 months (range 2-34). Six patients had specific drug treatment including provera, premarin, calcium supplements, or pamidronate. Of these, five healed. For the ten patients who did not have any specific treatment, only four showed signs of healing at the time of last follow-up. There was a trend toward earlier healing with specific drug treatment (P=0.11). CONCLUSIONS: Fractures can easily be mistaken for metastatic lesions (37.5% in this series) which might be treated with further irradiation. Although not statistically significant, there was a trend towards early healing with drug therapy. More studies are required to generate quantitative data for dose-response relationships and to evaluate the effect of drug therapy on the healing of such fractures. PMID- 10869752 TI - Urethral carcinoma in women: results of treatment with primary radiotherapy. AB - BACKGROUND AND PURPOSE: Urethral carcinoma in women is uncommon. This study was undertaken to evaluate the role of radiotherapy in the treatment of these tumors. MATERIALS AND METHODS: The hospital records of 34 women with primary urethral carcinoma were retrospectively reviewed. There were 15 squamous cell carcinomas, 13 transitional cell carcinomas, and six adenocarcinomas. The primary tumor was >4cm in size in eight patients, involved the proximal urethra in 19 and extended to adjacent organs in 22. Inguinal or iliac lymphadenopathy was present in nine patients. There were eight TNM stage I/II tumors, 11 stage III tumors and 15 stage IV tumors. Radiotherapy was administered only to the primary tumor in 15 patients, and to the primary tumor and regional lymph nodes in the remaining 19 patients. Brachytherapy with or without external radiation was used to treat the primary tumor in 20 patients. RESULTS: Tumor recurred in 21 patients. The 7-year actuarial overall and cause-specific survivals were 41 and 45%, respectively. Large primary tumor bulk and treatment with external beam radiation alone (no brachytherapy) were independent adverse prognostic factors for local tumor recurrence. Brachytherapy reduced the risk of local recurrence by a factor of 4.2. The beneficial effect of brachytherapy was most prominently seen in patients with bulky primary disease. Large tumor size was the only independent adverse predictor of overall disease recurrence and death from cancer. CONCLUSIONS: Radiotherapy is an effective treatment for carcinoma of the female urethra and preserves normal anatomy and function. Brachytherapy improves local tumor control, possibly as a result of the higher radiation dose that can safely be delivered. PMID- 10869753 TI - Inter-observer variation in delineation of bladder and rectum contours for brachytherapy of cervical cancer. AB - BACKGROUND AND PURPOSE: In 3D treatment planning of low dose rate brachytherapy of cervical carcinoma the dose in bladder and rectum can be estimated from dose volume histograms (DVHs). In this study, the influence of inter-observer variation in delineation of bladder and rectum on DVHs and dose at specific bladder and rectum points was investigated. MATERIALS AND METHODS: Three observers delineated bladder and rectum on axial CT images of ten patients. The highest minimum dose in bladder and rectum was determined for, respectively, 2 cm(3) (D(2)) and 5 cm(3) (D(5)), as well as the dose at specific points placed on the bladder and rectum wall. RESULTS: The inter-observer variation in D(2) was 10% (1 average relative SD) in bladder and 11% (1 SD) in rectum. In D(5) the variation was 8% (1 SD) in bladder and 11% in rectum. The variation in the bladder point was 13% (1 SD) and in the rectum point 11% (1 SD). Differences in delineation among the observers were caused by unclear organ boundaries on the CT images. CONCLUSIONS: Taking the inter-observer variation caused by delineation differences into account, dose in bladder and rectum can be determined within an accuracy of about 10% (1 SD). PMID- 10869754 TI - Carcinoma of the uterine cervix: a 3D - CT analysis of dose to the internal, external and common iliac nodes in tandem and ovoid applications. AB - PURPOSE: To describe external, internal and common iliac dose rates estimated with 3D-computed tomography (CT) based dose calculations in tandem and ovoid brachytherapy. MATERIALS AND METHODS: Thirty patients with carcinoma of the uterine cervix received low dose rate brachytherapy with a CT-compatible Fletcher Suit-Deldos device. A total of 36 implants were performed with axial CT images used to identify internal iliac, external iliac, and common iliac vessels. Dose rates on the surfaces of these vessels were calculated for the purpose of estimating the dose to their associated lymph nodes. RESULTS: In 22 out of 72 comparisons, point B overestimated the maximum dose with the external iliac nodes. In 21 out of 72 comparisons, point B overestimated the maximum dose with the internal iliac nodes. In all cases, Point B overestimated the minimum dose to the internal and external iliac nodal chains. CONCLUSION: It was found that Point B dose is similar to the maximum common iliac nodal dose. Patient to patient variability, of Point B dose, warrants further study of dose distributions to the nodal chains. The minimum dose to the external iliac nodal chain at the bifurcation of the nodal chains may provide a useful measure of 'pelvic side wall dose' and deserves further study to see if it can be correlated with pelvic side wall control and complications. PMID- 10869755 TI - Navigation system for interstitial brachytherapy. AB - PURPOSE: To develop a computed tomography (CT) based electromagnetic navigation system for interstitial brachytherapy. This is especially designed for situations when needles have to be positioned adjacent to or within critical anatomical structures. In such instances interactive 3D visualisation of the needle positions is essential. METHODS AND MATERIALS: The material consisted of a Polhemus electromagnetic 3D digitizer, a Pentium 200 MHz laptop and a voice recognition for continuous speech. In addition, we developed an external reference system constructed of Perspex which could be positioned above the tumour region and attached to the patient using a non-invasive fixation method. A specially designed needle holder and patient bed were also developed. Measurements were made on a series of phantoms in order to study the efficacy and accuracy of the navigation system. RESULTS: The mean navigation accuracy of positioning the 20.0 cm length metallic needles within the phantoms was in the range 2.0-4.1 mm with a maximum of 5.4 mm. This is an improvement on the accuracy of a CT-guided technique which was in the range 6.1-11.3 mm with a maximum of 19.4 mm. The mean reconstruction accuracy of the implant geometry was 3.2 mm within a non-ferromagnetic environment. We found that although the needles were metallic this did not have a significant influence. We also found for our experimental setups that the CT table and operation table non-ferromagnetic parts had no significant influence on the navigation accuracy. CONCLUSIONS: This navigation system will be a very useful clinical tool for interstitial brachytherapy applications, particularly when critical structures have to be avoided. It also should provide a significant improvement on our existing technique. PMID- 10869756 TI - Treatment for pyothorax-associated lymphoma. AB - In eight patients with pyothorax-associated lymphoma (PAL), which resulted from artificial pneumothorax for the treatment of pulmonary tuberculosis, seven patients received radiotherapy and five showed no local recurrence. All four patients treated by primary chemotherapy had disease progression. Radiotherapy of 50 Gy with wide margins is recommended to treat PAL. PMID- 10869757 TI - Breast-conserving radiation therapy using combined electron and intensity modulated radiotherapy technique. AB - BACKGROUND AND PURPOSE: To explore the feasibility of a multi-modality breast conserving radiation therapy treatment technique to reduce high dose to the ipsilateral lung and the heart when compared with the conventional treatment technique using two tangential fields. MATERIALS AND METHODS: An electron beam with appropriate energy was combined with four intensity modulated photon beams. The direction of the electron beam was chosen to be tilted 10-20 degrees laterally from the anteroposterior direction. Two of the intensity-modulated photon beams had the same gantry angles as the conventional tangential fields, whereas the other two beams were rotated 15-25 degrees toward the anteroposterior directions from the first two photon beams. An iterative algorithm was developed which optimizes the weight of the electron beam as well as the fluence profiles of the photon beams for a given patient. Two breast cancer patients with early stage breast tumors were planned with the new technique and the results were compared with those from 3D planning using tangential fields as well as 9-field intensity-modulated radiotherapy (IMRT) techniques. RESULTS: The combined electron and IMRT plans showed better dose conformity to the target with significantly reduced dose to the ipsilateral lung and, in the case of the left breast patient, reduced dose to the heart, than the tangential field plans. In both the right-sided and left-sided breast plans, the dose to other normal structures was similar to that from conventional plans and was much smaller than that from the 9-field IMRT plans. The optimized electron beam provided between 70 to 80% of the prescribed dose at the depth of maximum dose of the electron beam. CONCLUSIONS: The combined electron and IMRT technique showed improvement over the conventional treatment technique using tangential fields with reduced dose to the ipsilateral lung and the heart. The customized beam directions of the four IMRT fields also kept the dose to other critical structures to a minimum. PMID- 10869758 TI - Target volume definition in conformal radiotherapy for prostate cancer: quality assurance in the MRC RT-01 trial. AB - BACKGROUND AND PURPOSE: Prior to randomization of patients into the UK Medical Research Council multicentre randomized trial (RT-01) of conformal radiotherapy (CFRT) in prostate cancer, clinicians at participating centres were required to complete a quality assurance (QA) clinical planning exercise to enable an investigation of inter-observer variability in gross target volume (GTV) and normal structure outlining. MATERIALS AND METHODS: Thirteen participating centres and two investigators completed the clinical planning exercise of three practice planning cases. Clinicians were asked to draw outlines of the GTV, rectum and bladder on hard-copy computerized tomography (CT) films of the pelvis, which were transferred onto the Cadplan computer planning system by a single investigator. Centre, inferior and superior CT levels of GTV, rectum and bladder were noted, and volume calculations performed. Planning target volumes (PTV) were generated using automatic volume expansion of GTVs by a 1 cm margin. Anterior, right and left lateral beam eye views (BEV) of the PTVs were generated. Using a common central point, the BEV PTVs were superimposed for each beam direction of each case. Radial PTV variation was investigated by measurement of a novel parameter, termed the radial line measurement variation (RLMV). RESULTS: GTV central slice and length were defined with reasonable consistency. The RLMV analysis showed that the main part of the prostate gland, bladder and inferior rectum were outlined with good consistency among clinicians. However, the outlining of the prostatic apex, superior aspect of the prostate projecting into the bladder, seminal vesicles, the base of seminal vesicles and superior rectum were more variable. CONCLUSION: This exercise has demonstrated adequate consistency of GTV definition. The RLMV method of analysis indicates particular regions of clinician uncertainty. Appropriate feedback has been given to all participating clinicians, and the final RT-01 trial protocol has been modified to accommodate these findings. PMID- 10869759 TI - Quality assurance by systematic in vivo dosimetry: results on a large cohort of patients. AB - BACKGROUND: In vivo dosimetry is widely considered to be an important tool for quality assurance in external radiotherapy. INTRODUCTION: In this study we report on our experience over more than 4 years in systematic in vivo dosimetry with diodes. MATERIALS AND METHODS: From November '94 an in vivo entrance dosimetry check was performed for every new patient irradiated at one of our treatment units (Linac 6/100, 6 MV X-rays). Diodes were calibrated in terms of entrance dose; appropriate correction factors had been previously assessed (taking SSDs, field width, wedge, oblique incidence and blocking tray into account) and were individually applied to in vivo diode readings. The in vivo measured entrance dose was compared with the expected one, with a 5% action level; if a larger deviation was found, all treatment parameters were verified, and the in vivo dosimetry check was repeated. During the period November '94-May '99, 2824 measurements on 1433 patients were collected. RESULTS: Nine out of 1433 (0.63%) serious systematic errors (leading to a 5% or more on the delivered dose to the PTV) were detected by in vivo dosimetry; four out of nine would produce a 10% or more error if not detected. The rate of serious systematic errors detected by an independent check of treatment chart and MU calculation was found to be 1.5%, showing that less than 1/3 of the errors escapes this check. One hundred and twelve out of 1433 (7.8%) patients had more than one check: the rate of second checks was significantly higher for breast patients (31/250, 12.4%) against non breast patients (81/1183, 6.8%, P=0.003). A number of patients demonstrated a persistent relatively large error even after two or more checks. For almost all patients the cause of the deviation was assessed; the most frequent cause was the difficulty in correctly positioning the patient and/or the diode. When analyzing the distribution of the deviations between measured and expected entrance doses (excluding first checks in the case of repetition of the in vivo dosimetry control) the mean deviation was 0.4% with a standard deviation equal to 3.0%. The rates of deviations larger than 5 and 7% were 9.9 and 2.6%, respectively. When considering the same data taking the average deviation in the case of opposed beams, the SD became 2.6% and the rates of deviations larger than 5 and 7%, respectively, 5.2 and 0.8%. When dividing the beams according to their orientation, significantly higher rates of large deviations (>5 and 7%) were found for oblique and posterior-anterior (PA) fields against lateral and anterior posterior (AP) fields (P<0.05). Similarly, higher rates of large deviations were found for wedged fields against unwedged fields (P<0.03) and for blocked fields against unblocked fields (P<0.01). When dividing the data according to the anatomical district, accuracy was worse for breast (mean deviation 0.1%, 1 SD: 3.5%) and neck AP-PA fields (mean deviation 1%, 1 SD: 3,4%). Better accuracy was found for vertebrae (0.1%, 1 SD 2. 1%) and brain patients (-0.7%, 1 SD: 2.6%). During the considered period, in vivo dosimetry was also able to promptly detect a systematic error caused by a wrong resetting of the simulator height couch indicator, with a consequent error in the estimate of patient thickness of about 4 cm. CONCLUSIONS: In our experience, systematic in vivo dosimetry demonstrated to be a valid tool for quality assurance, both in detecting systematic errors which may escape the data transfer/MU calculation check and in giving an effective way of estimating the accuracy of treatment delivery. PMID- 10869760 TI - Dosimetric effects of patient displacement and collimator and gantry angle misalignment on intensity modulated radiation therapy. AB - PURPOSE AND OBJECTIVE: The primary goal of this study was to examine systematically the dosimetric effect of small patient movements and linear accelerator angular setting misalignments in the delivery of intensity modulated radiation therapy. We will also provide a method for estimating dosimetric errors for an arbitrary combination of these uncertainties. MATERIALS AND METHODS: Sites in two patients (lumbar-vertebra and nasopharynx) were studied. Optimized intensity modulated radiation therapy treatment plans were computed for each patient using a commercially available inverse planning system (CORVUS, NOMOS Corporation, Sewickley, PA). The plans used nine coplanar beams. For each patient the dose distributions and relevant dosimetric quantities were calculated, including the maximum, minimum, and average doses in targets and sensitive structures. The corresponding dose volumetric information was recalculated by purposely varying the collimator angle or gantry angle of an incident beam while keeping other beams unchanged. Similar calculations were carried out by varying the couch indices in either horizontal or vertical directions. The intensity maps of all the beams were kept the same as those in the optimized plan. The change of a dosimetric quantity, Q, for a combination of collimator and gantry angle misalignments and patient displacements was estimated using Delta=Sigma(DeltaQ/Deltax(i))Deltax(i). Here DeltaQ is the variation of Q due to Deltax(i), which is the change of the i-th variable (collimator angle, gantry angle, or couch indices), and DeltaQ/Deltax(i) is a quantity equivalent to the partial derivative of the dosimetric quantity Q with respect to x(i). RESULTS: While the change in dosimetric quantities was case dependent, it was found that the results were much more sensitive to small changes in the couch indices than to changes in the accelerator angular setting. For instance, in the first example in the paper, a 3-mm movement of the couch in the anterior-posterior direction can cause a 38% decrease in the minimum target dose or a 41% increase in the maximum cord dose, whereas a 5 degrees change in the θ(1)=20 degrees beam only gave rise to a 1.5% decrease in the target minimum or 5.1% in the cord maximum. The effect of systematic positioning uncertainties of the machine settings was more serious than random uncertainties, which tended to smear out the errors in dose distributions. CONCLUSIONS: The dose distribution of an intensity modulated radiation therapy (IMRT) plan changes with patient displacement and angular misalignment in a complex way. A method was proposed to estimate dosimetric errors for an arbitrary combination of uncertainties in these quantities. While it is important to eliminate the angular misalignment, it was found that the couch indices (or patient positioning) played a much more important role. Accurate patient set-up and patient immobilization is crucial in order to take advantage fully of the technological advances of IMRT. In practice, a sensitivity check should be useful to foresee potential IMRT treatment complications and a warning should be given if the sensitivity exceeds an empirical value. Quality assurance action levels for a given plan can be established out of the sensitivity calculation. PMID- 10869761 TI - Quality assurance in radiotherapy by identifying standards and monitoring treatment preparation. AB - BACKGROUND AND PURPOSE: Due to the complexity of the treatment preparation in radiotherapy, a number of errors go undetected until after the first treatment session. Some of these errors could easily have been noticed before treatment if an objective filter existed in addition to human supervision. With this in mind, a conceptually novel extension to conventional quality assurance procedures was explored to create a global platform monitoring treatment preparation by comparison with the existing local standards. MATERIALS AND METHODS: The feasibility of developing such a platform was evaluated for a test case on a cohort of 202 patients having received breast irradiation. By statistical analysis of the treatment parameters, mean values and tolerance levels could be defined for most parameters based on the observed standard deviations. Useful correlations were traced providing us with a means to automatically track errors, the detection of which would otherwise solely depend upon the alertness of the supervisor. RESULTS AND CONCLUSIONS: Apart from its possibilities as a mere quality control tool, the platform, developed in the framework of EQUART (European Quality Assurance Program in Radiotherapy by Monitoring Treatment Preparation), can be incorporated in the treatment preparation chain, providing standard setup values for the simulation. A crucial achievement of EQUART lies in the fact that filtering out of errors occurs prior to treatment initiation. PMID- 10869762 TI - Prompt radiation oncology record access by patient centered digital image chart system. AB - The authors have developed and evaluated a radiation oncology digital image chart system (RODICS). With this system we could achieve paperless and filmless practice, and thus improved operational efficiency within the department. In this paper, we describe characteristics and clinical usage of RODICS. PMID- 10869763 TI - An integrated service digital network (ISDN)-based international telecommunication between Samsung Medical Center and Hokkaido University using telecommunication helped radiotherapy planning and information system (THERAPIS). AB - This study introduces the integrated service digital network (ISDN)-based international teleradiotherapy system (THERAPIS) in radiation oncology between hospitals in Seoul, South Korea and in Sapporo, Japan. THERAPIS has the following functions: (1) exchange of patient's image data, (2) real-time teleconference, and (3) communication of the treatment planning, dose calculation and distribution, and of portal verification images between the remote hospitals. Our preliminary results of applications on eight patients demonstrated that the international telecommunication using THERAPIS was clinically useful and satisfactory with sufficient bandwidth for the transfer of patient data for clinical use in radiation oncology. PMID- 10869764 TI - A simple and reliable method to simulate multileaf-plans. AB - To combine conformity of the irradiation with time effectiveness during treatment, the use of multileaf collimators has become more and more common. However, the simulation of the leaf positions is rather difficult compared with metal blocks. We developed a new method, utilizing an acrylic template in which the contour produced by the leaves is machined in the form of a 1 mm groove by a computerized numerically controlled milling machine. This template is then inserted into a mount attached to the simulator. The main advantages are the errorfree, direct communication from the therapy planning system to the milling machine via a network, the possibility to transfer the contour to the skin, and the documentation on the simulation film. The use of templates is reliable and, e.g. the costs of the materials are lower than for block simulation. PMID- 10869765 TI - A comparison of immunogenicity and in vivo distribution of Salmonella enterica serovar Typhi and Typhimurium live vector vaccines delivered by mucosal routes in the murine model. AB - We evaluated the immune responses elicited by attenuated Salmonella enterica serovar Typhi vaccine strain CVD 908-htrA and serovar Typhimurium strain SL3261 alone or as live vectors carrying a plasmid encoding fragment C of tetanus toxin (pTETnir15) in mice immunized intranasally and orogastrically, as well as the in vivo distribution of vaccine organisms following immunization. Higher serologic and proliferative responses against both vector and the foreign antigen were elicited when vaccines were delivered by intranasal route. Whereas both Salmonella strains were detected in the nasal tissue, lungs, and Peyer's patches following intranasal and orogastric immunization, larger numbers of vaccine organisms were recovered from these tissues when the vaccines were delivered intranasally. PMID- 10869766 TI - Cross-protection against a lethal influenza virus infection by DNA vaccine to neuraminidase. AB - Cross-protection against a lethal influenza virus infection was examined in BALB/c mice immunized with plasmid DNAs encoding the neuraminidase (NA) from different subtype A viruses. Each NA-DNA was administered twice, 3 weeks apart, at the dose of 1 microg per mouse by particle-mediated DNA transfer to the epidermis (gene gun) or at a dose of 30 microg per mouse by electroporation into the muscle. Three weeks after the second vaccination, the mice were challenged with lethal doses of homologous or heterologous viruses and the ability of each NA-DNA to protect the mice from influenza was evaluated by determining the lung virus titers, body weight and survival rates. The H3N2 virus NA-DNA conferred cross-protection against lethal challenge with antigenic variants within the same subtype, but failed to provide protection against infection by a different subtype virus (H1N1). The degree of cross-protection against infection was related to titers of the cross-reacting antibodies. These results suggest that NA DNA can be used as a vaccine component to provide effective protection against infection not only with homologous virus but also with drift viruses. PMID- 10869767 TI - Intranasal vaccination of mice against infection with Mycobacterium tuberculosis. AB - The intranasal (i.n.) route of immunisation, has recently been of active interest in endeavours to improve the efficacy of vaccination against a number of respiratory infections. Here, we examined the outcome of tuberculous infection in BALB/c mice. I.n. application of the BCG-Pasteur strain was found to be highly protective against challenge infection with the pathogenic H37Rv strain given after a 4-week interval, reflected by the 100-fold reduction of CFUs in both lungs and spleens. Vaccination with the recombinant PstS-1 antigen and cholera toxin significantly protected against the challenge given 10 days later, but only marginally after 12 weeks. Histological examination showed, that i.n. vaccination abrogated the confluent infiltration of lungs with inflammatory cells, which surrounds the granulomas in H37Rv challenged control mice. In conclusion, the strong protection demonstrated by BCG suggests that the i.n. route of vaccine delivery deserves further attention toward improving vaccination against tuberculosis. PMID- 10869768 TI - Protection from IBDV-induced bursal damage by a recombinant fowlpox vaccine, fpIBD1, is dependent on the titre of challenge virus and chicken genotype. AB - Expression of the VP2 capsid protein of infectious bursal disease virus (IBDV) in an vaccine strain of fowlpox has produced an experimental recombinant vaccine, fpIBD1. Successful vaccination with fpIBD1 was dependent on the titre of challenge virus for high titres of challenge virus were able to overcome protection induced by fpIBD1 whereas challenge with a low titre of virus did not. The genotype of chicken also has an important effect on the outcome of challenge possibly as a result of the major histocompatability complex and its ability to present VP2-derived peptides to the immune system. It was not possible to protect the inbred white leghorn chicken strain, line 15I, from IBDV-induced bursal damage by vaccination with fpIBD1 even at the lowest titre of challenge virus used. All other inbred white leghorn chickens examined (line 6(1), C. B4 and C.B12) and outbred Rhode Island Red chickens were protected by fpIBD1. Protection by the fpIBD1 vaccine is induced in the absence of detectable serum antibodies, suggesting the possibility of a significant role for cell-mediated immunity in protection from IBDV challenge. PMID- 10869769 TI - Superiority of intramuscular route and full length glycoprotein D for DNA vaccination against herpes simplex 2. Enhancement of protection by the co delivery of the GM-CSF gene. AB - Immunization with naked DNA has been analyzed in two critical variables: the site of injection and the cellular compartment to which the coded protein is directed. The gene for the full length of the glycoprotein D (gD) of HSV-2 under the control of the citomegalovirus (CMV) promoter was injected via the intradermal (i.d.) or the intramuscular (i.m.) routes in mice. Immunization in the quadricep muscle was superior to the intradermal immunization in the footpads. A stronger activation of IFN-gamma-secreting cells in the spleen and draining lymph nodes (DLN) was induced, resulting in a more efficient protection against an intravaginal challenge. In order to analyze the effect of the cellular localizations of the coded protein, the DNA for the truncated form of the gD (DeltagD) was injected via the i.m. route. Immunization with a vector encoding for DeltagD resulted in higher antibody levels in serum and vaginal washes than immunization with the gene for the full length gD. However, immunization with the DeltagD DNA elicited a much weaker cell-mediated immune response and was inferior to gD DNA in providing protection against a lethal intravaginal challenge with HSV. Co-injection of an expression cassette for the granulocyte-macrophage colony stimulating factor (GM-CSF) increased both the humoral and cell-mediated immune response with both gD and DeltagD. A strong activation of IL-4-secreting cells was observed in the spleen and DLN together with an increase in the number of IFN gamma-secreting cells. In addition, a reduction in the vaginal virus titers after an intravaginal challenge was observed in mice co-injected with the GM-CSF gene as compared to those immunized with pCDNAgD only. PMID- 10869770 TI - Mucosal immunization with experimental feline immunodeficiency virus (FIV) vaccines induces both antibody and T cell responses but does not protect against rectal FIV challenge. AB - Feline immunodeficiency virus (FIV) is a natural lentiviral pathogen of cats which can be experimentally transmitted via rectal and vaginal routes--the major routes of human immunodeficiency virus type 1 transmission in man. An important objective for lentiviral research is the development of vaccine strategies which generate good mucosal immune responses capable of giving protection from a mucosal virus challenge. The experimental vaccines employed in this study were based on (a) a peptide from the third variable region of the FIV envelope glycoprotein and (b) fixed whole FIV, Glasgow-8 strain. Adjuvants used were Quil A and cholera toxin for mucosal administration and incomplete Freund's adjuvant and immune stimulating complexes for subcutaneous injection. Mucosal immunization was given by rectal and intranasal routes. Both antibody and proliferative responses were elicited by mucosal immunization and cholera toxin was found to be a good mucosal adjuvant. The addition of a lipo thioester to the FIV peptide improved IgG and IgA responses upon parenteral administration. However, no protection from a rectal FIV challenge was achieved. PMID- 10869771 TI - Protection studies following bronchopulmonary and intramuscular immunisation with yersinia pestis F1 and V subunit vaccines coencapsulated in biodegradable microspheres: a comparison of efficacy. AB - We have compared the ability of intramuscularly and intratracheally administered recombinant F1 and V subunit antigens to safeguard mice from a lethal systemic challenge with plague. The combined subunits (1 microg V plus 5 microg F1) were inoculated either in the 'free' state as a solution, or entrapped within microspheres composed of a biodegradable polyester (Poly-L-lactide), on day 1 and 60 of the experiment. In comparison to the other regimens, introduction of microsphere suspensions into the respiratory tract resulted in statistically elevated levels of specific immunoglobulins in day 82 lung wash samples. A subcutaneous challenge with virulent Yersinia pestis bacteria on day 137, equivalent to more than 10(5) mouse LD(50)s, was comparatively well tolerated by all subunit treatment groups (with survival rates between 66 and 90%). In contrast, 80% of the mice injected intramuscularly with soluble F1 and V were defeated by a 10(7) MLD(50) subcutaneous challenge, whereas the group immunised intramuscularly with microparticles were significantly better protected (p<0.1) with 50% survival. Similarly, mice immunised intratracheally with microparticles were significantly better safeguarded (56% survival) compared with the group immunised with soluble subunits intramuscularly (p<0.01). Soluble sub-units delivered intratracheally afforded 33% protection against 10(7) MLD(50)s. These data indicate that bronchopulmonary administration of microsphere co-encapsulated recombinant F1 and V antigens elicits a similar level of protective immunity against systemic plague infection as that evoked by injecting co-encapsulated subunits into the muscle. Such findings corroborate the thesis that introduction of appropriately formulated F1 and V subunits into the respiratory tract may be an alternative to parenteral immunisation schedules for protecting individuals from plague. PMID- 10869772 TI - ELISA test for rabies antibody titration in orally vaccinated foxes sampled in the fields. AB - The assessment of the efficacy of rabies oral vaccination campaigns requires the titration of specific antibodies in the target species. Unfortunately, in Continental Europe, most fox serum samples are in fact "body fluids" taken from cadavers and the lack of a validated titration method for these poor quality sera made it impossible to survey and compare the efficacy of various oral vaccination protocols used by the different European teams. By using ready to use microplates sensitised with rabies virus glycoprotein purchased from a manufacturer and applying a simple and rapid ELISA technique on field fox sera, we obtained antibody quantitation highly correlated with seroneutralising antibody titres measured with a seroneutralisation test on cell culture. We obtained, with fox sera sampled in the same area, the same distribution of high, medium and low titres within all categories of serum quality (from high to very poor quality) and therefore conclude that this ELISA test allows a reliable titration even with highly contaminated body fluids. This test was shown to be equally capable of detecting rabies antibodies in serum samples taken from foxes vaccinated with an highly attenuated rabies virus (the SAG2 double mutant of the Street Alabama Dufferin strain) or with the VRG, the Vaccinia recombinant glycoprotein. Additionally, a strong correlation was demonstrated between titres given by this ELISA (or by the seroneutralisation test) and protection against challenge of foxes orally vaccinated with SAG2 vaccine baits. In view of this validation, this simple and reliable test is proposed for sero-surveying foxes following rabies oral vaccination campaigns. PMID- 10869773 TI - Antibody response to influenza immunization in two consecutive epidemic seasons in patients with renal diseases. AB - The aim of this study was to assess antihemagglutinin and antineuraminidase antibody kinetics in 26 patients with renal diseases vaccinated against influenza in two consecutive epidemic seasons. Antibody responses were measured before immunization and 1, 3 and 6 months after immunization. Antihemagglutinin (HI) antibodies were determined by the hemagglutinin inhibition test and antineuraminidase (NI) antibody levels by the neuraminidase inhibition test. After vaccination HI and NI antibody titers significantly increased when compared with the pre-vaccination levels. Three months after vaccination the protection rates ranged from 50 to 61.5% in the 1995/96 season and 100% for all antigens in the 1996/97 season. Response rates ranged from 50 to 57.7% and 93.8 to 100% respectively. Significantly higher humoral response was recorded in the 1996/97 season than in the 1995/96 season. No serious adverse reactions were observed in the vaccinated patients and no symptoms of influenza or influenza-like infection were noted. In spite of some doubts about the safety and efficacy of influenza vaccination in patients from high-risk groups, the results of this study showed that many of them are able to produce HI antibodies in titers which are sufficient to protect against the influenza infection. PMID- 10869774 TI - European sero-epidemiology network: standardisation of the results of diphtheria antitoxin assays. AB - A European Sero-Epidemiological Network (ESEN) was established with the aim to co ordinate and harmonise serological surveillance of immunity to communicable diseases in Europe. In this study the inter-laboratory standardisation of diphtheria toxin antibody measurements is reported. A standard panel of 162 sera was tested by the participating laboratories using an in vitro assay of their choice: VERO cell toxin neutralisation assay (NT), double-antigen delayed time resolved fluorescence immuno-assay (DA-DELFIA), double-antigen enzyme-linked immunosorbent assay (DAE), toxin binding inhibition test (ToBI) and an indirect enzyme-linked immunosorbent assay (ELISA). The results were standardised using regression against the NT. The variations due to inter-laboratory and inter-assay variation, which would otherwise make it difficult directly to compare the main serum bank results by the different laboratories and the various assays were successfully minimised by the standardisation. The regression equations obtained will be used to transform the respective local results of testing the main serum bank into the reference test unitages. This study also gave the opportunity to compare the various assays within and between laboratories. This demonstrated a very high correlation between DA-DELFIA, DAE, ToBI and the NT. The ELISA showed a good correlation, too, however sera below some 0.1 IU/ml were overestimated. PMID- 10869775 TI - The immune response of mice and cynomolgus monkeys to macaque mucin 1-mannan. AB - Mice immunised with human epithelial mucin MUC1 coupled to oxidised mannan produce MUC1 specific MHC Class 1 restricted CD8(+) cytotoxic T cells and are completely protected from the development of MUC1(+) tumours; such therapy may be applicable to humans. In this light we describe pre-clinical studies in cynomolgus monkeys (Macaca fascicularis), to test the efficacy of mannan-MUC1 in higher primates. Monkey MUC1 genomic clones were isolated from a macaque library, peptides and fusion protein synthesised and mice and monkeys immunised with macaque MUC1-mannan. In mice CTL responses were induced (as has been found with human MUC1 mannan conjugates), but in contrast monkeys produced a humoral response, with no T cell proliferative, cytotoxic responses or CTLp found. In spite of the presence of anti-MUC1 auto-antibodies, there was no toxicity or induction of autoimmunity. PMID- 10869776 TI - Induction of protective immunity against pathogenic simian immunodeficiency virus by a foreign receptor-dependent replication of an engineered avirulent virus. AB - In AIDS vaccine strategies, live attenuated vaccines can confer good resistance against pathogenic virus infections but have the potential risk of inducing disease, whereas safer replication-negative strategies such as DNA vaccinations have so far failed to prevent the disease onset. Here, we developed a novel DNA vaccine strategy to induce restricted replication of an avirulent virus and evaluated it in a simian immunodeficiency virus (SIV) infection model. We generated a chimeric SIV, FMSIV, by replacing SIV env with ecotropic Friend murine leukemia virus (FMLV) env to confine its replication to FMLV receptor (mCAT1)-expressing cells. In primate cells lacking mCAT1, FMSIV did not replicate unless mCAT1 was introduced exogenously. Vaccination to macaques with both the FMSIV DNA and the mCAT1-expression plasmid DNA induced SIV Gag-specific cellular immune responses and resistance against pathogenic SIV(mac239) challenge more efficiently than the replication-negative control vaccination with the FMSIV DNA alone. This strategy may be useful for development of safe and effective vaccines against various kinds of pathogenic viruses. PMID- 10869778 TI - From hepatitis B to hepatitis A and B prevention: the puglia (Italy) experience PMID- 10869777 TI - Alkyl-polyacrylate esters are strong mucosal adjuvants. AB - Synthetic polymers were examined for their potency to enhance mucosal immune responses to inactivated antigens. Aqueous solutions of polyacrylic acid with a MW of 450 kDa (p[AA]) or an butyl-ester thereof with 16% esterification (Butyl16 p[AA]) plus antigen were administered twice intranasally in mice with a 2 week interval. The frequency of IgA-antibody secreting cells (ASCs) in lung cell suspensions was determined 1 week after the second immunisation. Both polymers significantly enhanced the IgA response against inactivated Newcastle disease virus (iNDV), inactivated influenza virus strain MRC-11 (iMRC-11), haemagglutinin/neuraminidase subunits of influenza virus strain A/Texas (HA/NA) and bovine serum albumin (BSA). Butyl16-p(AA) was significantly more effective than non-derivatised p(AA), cholera toxin B subunit (CTB) or liposomes. The factor of increase in IgA-ASCs varied from <10- to >100-fold and depended on the type of antigen, the dose of antigen and the adjuvant. Extremely high responses of about 10,000 IgA-ASCs per million lung cells were detected after immunisation with 5 microg HA/NA plus 50 microg Butyl16-p(AA). Intranasal immunisation with Butyl16-p(AA) resulted in high IgA responses, not only in the lungs, but also in the spleen and in high IgG responses in these organs. We concluded that alkyl esters of polyacrylate are an interesting, novel category of mucosal adjuvants. PMID- 10869779 TI - What role(s) for TGFalpha in the central nervous system? AB - Transforming growth factor alpha (TGFalpha) is a member of the epidermal growth factor (EGF) family with which it shares the same receptor, the EGF receptor (EGFR or erbB1). Identified since 1985 in the central nervous system (CNS), its functions in this organ have started to be determined during the past decade although numerous questions remain unanswered. TGFalpha is widely distributed in the nervous system, both glial and neuronal cells contributing to its synthesis. Although astrocytes appear as its main targets, mediating in part TGFalpha effects on different neuronal populations, results from different studies have raised the possibility for a direct action of this growth factor on neurons. A large array of experimental data have thus pointed to TGFalpha as a multifunctional factor in the CNS. This review is an attempt to present, in a comprehensive manner, the very diverse works performed in vitro and in vivo which have provided evidences for (i) an intervention of TGFalpha in the control of developmental events such as neural progenitors proliferation/cell fate choice, neuronal survival/differentiation, and neuronal control of female puberty onset, (ii) its role as a potent regulator of astroglial metabolism including astrocytic reactivity, (iii) its neuroprotective potential, and (iv) its participation to neuropathological processes as exemplified by astroglial neoplasia. In addition, informations regarding the complex modes of TGFalpha action at the molecular level are provided, and its place within the large EGF family is precised with regard to the potential interactions and substitutions which may take place between TGFalpha and its kindred. PMID- 10869780 TI - The midline glia of Drosophila: a molecular genetic model for the developmental functions of glia. AB - The Midline Glia of Drosophila are required for nervous system morphogenesis and midline axon guidance during embryogenesis. In origin, gene expression and function, this lineage is analogous to the floorplate of the vertebrate neural tube. The expression or function of over 50 genes, summarised here, has been linked to the Midline Glia. Like the floorplate, the cells which generate the Midline Glia lineage, the mesectoderm, are determined by the interaction of ectoderm and mesoderm during gastrulation. Determination and differentiation of the Midline Glia involves the Drosophila EGF, Notch and segment polarity signaling pathways, as well as twelve identified transcription factors. The Midline Glia lineage has two phases of cell proliferation and of programmed cell death. During embryogenesis, the EGF receptor pathway signaling and Wrapper protein both function to suppress apoptosis only in those MG which are appropriately positioned to separate and ensheath midline axonal commissures. Apoptosis during metamorphosis is regulated by the insect steroid, Ecdysone. The Midline Glia participate in both the attraction of axonal growth cones towards the midline, as well as repulsion of growth cones from the midline. Midline axon guidance requires the Drosophila orthologs of vertebrate genes expressed in the floorplate, which perform the same function. Genetic and molecular evidence of the interaction of attractive (Netrin) and repellent (Slit) signaling is reviewed and summarised in a model. The Midline Glia participate also in the generation of extracellular matrix and in trophic interactions with axons. Genetic evidence for these functions is reviewed. PMID- 10869781 TI - The pharmacology of headache. AB - Headache is a common problem which besets most of us at some time or the other. The pharmacology of headache is complex in an overall sense but can be understood in terms of the anatomy and physiology of the pain-producing structures. Migraine can be used as a template to understand the activation of nociceptive systems in the head and thus their neurotransmitter mediation and modulation. In recent years, the role of serotonin (5-HT) in headache pharmacology has been unravelled in the context of both understanding its role in the nociceptive systems related to headache and by exploiting its 5-HT1 receptor subtypes in headache therapeutics. The pharmacology of the head pain systems, as they are known and as they might evolve, are explored in the context of both, the anatomy and physiology of trigeminovascular nociception and in the context of clinical questions, such as those of efficacy, headache recurrence and adverse events. PMID- 10869782 TI - Neurotrophic factors in the primary olfactory pathway. AB - The number of identified growth factors continues to increase rapidly with many being implicated in the development of the nervous system, although for most of them the autocrine and paracrine pathways of cellular regulation still remain to be elucidated. The primary olfactory pathway, consisting of the olfactory epithelium and olfactory bulb, is presented here as a very useful model for the analysis of growth factor function. Review of the available literature suggests that a large proportion of neuroactive growth factors and their receptors are present in the olfactory epithelium or olfactory bulb. Furthermore, the primary olfactory pathway is one of the most plastic in the nervous system with neurogenesis continuing to contribute new sensory neurones in the olfactory epithelium and new interneurones in the olfactory bulb throughout adult life. The rich diversity of growth factors and their receptors in the olfactory system indicates that it will be useful in elucidating how these molecules regulate the formation of the nervous system. The olfactory epithelium in particular is proving useful as a model for the actions of growth factors in directing the neuronal lineage from stem cell to mature neurone. PMID- 10869784 TI - The glutathione/glutathione-related enzyme system in reproduction. PMID- 10869783 TI - Is endometriosis an endometrial disease? AB - Endometriosis is characterised by the presence of abnormally located tissue resembling the endometrium with glands and stroma. Several hypotheses have attempted to explain the development of such tissue. The most often cited theory, that of implantation, proposes that the physiological phenomenon of endometrial reflux in the fallopian tubes during menstruation may, in certain conditions, overcome local defense mechanisms, implant, and proliferate. The implantation theory does not explain why endometriosis will develop only in approximately 10 15% of women, while the reflux of endometrial tissue via the fallopian tubes during menstruation is a quasi-universal phenomenon. The endometrium of women affected by endometriosis could be abnormal compared with endometrium of healthy women. The abnormal endometrium could be able to protect itself from harmful effects of immune cells by expressing specific antigens, by harbouring a different immune cell population and by synthetizing and secreting immunosuppressive factors. Several others characteristic features of endometrium have been described in women with endometriosis: (1) production of its own estrogens in too heavy amount; (2) aptitude for setting up on peritoneum; (3) tendencies to proliferate and to invade tissue; (4) aggressiveness for the peritoneum; (5) auto-protection from physiological apoptosis; (6) abnormal expression of heat shock proteins; and (7) excessive angiogenesis. PMID- 10869785 TI - Correction of hyperinsulinemia in oligoovulatory women with clomiphene-resistant polycystic ovary syndrome: a review of therapeutic rationale and reproductive outcomes. AB - Polycystic ovary syndrome (PCOS) describes a convergence of chronic multisystem endocrine derangements, including irregular menses, hirsutism, obesity, hyperlipidemia, androgenization, large and cystic-appearing ovaries, insulin resistance and subfertility. Few PCOS patients exhibit all of these features, and often only one sign or symptom is evident. The sequelae of PCOS reach beyond reproductive health, as women affected with PCOS have increased relative risks for myocardial infarction, hypertension, ischemic heart disease, thromboembolic disease and diabetes. Although the adverse health consequences associated with PCOS are substantial, unfortunately most women are not aware of these risks. Indeed, in infertility practice such concerns are secondary as most patients are referred for treatment specifically to achieve a pregnancy. Impairments in insulin metabolism appear central to the physiologic cascade of PCOS, yet clomiphene therapy fails to remedy this defect. Several investigators have described satisfactory reproductive outcomes for PCOS patients treated with oral insulin-lowering agents. In this report, we outline a diagnostic and therapeutic approach for women with PCOS refractory to clomiphene with attention to the underlying insulin imbalance associated with impaired fertility. PMID- 10869786 TI - Influence of menstrual factors and dietary habits on menstrual pain in adolescence age. AB - OBJECTIVES: The aim of this study was to determine the frequency of the primary dysmenorrhea in adolescence age and investigate correlation between menstrual factors, dietary habits and this pathology. STUDY DESIGN: The sample was constituted from 356 students that were subjected to questionnaire, abdominal ultrasound, and in some cases, hormonal dosing. RESULTS: The frequency of the primary dysmenorrhea was 85%. Early menarche was related to an increase of its prevalence and its severity. A long and heavy menstrual flow was related to an increase of its severity. As far as dietary habits, it was noted that a higher consumption of fish, eggs, fruit and a lower consumption of wine is correlated with a lower frequency. CONCLUSION: Primary dysmenorrhea is very common in young women. The risk factors for this pathology are early menarche, long and heavy menstrual flow, and lower consumption of fish, eggs, and fruit. PMID- 10869787 TI - The changing HIV epidemic in Italian pregnant women. AB - OBJECTIVE: To describe changes in the characteristics of HIV-pregnant women in Italy and the impact of strategies for prevention of HIV vertical transmission. STUDY DESIGN: Since 1985, HIV-infected women and their children are followed in 23 European centres in the European Collaborative Study (ECS), according to a standard protocol. Eight Italian Obstetric units participating in the ECS enrolled 815 patients. RESULTS: Overall use of zidovudine to reduce HIV vertical transmission has increased significantly since 1994 and between 1995 and 1997, 57% of Italian women were treated. However, 27% of babies received the infant component of the 076 regimen. Over the years, age at delivery has increased and their CD4 count at delivery decreased, most likely reflecting heterosexually infected women with a longer duration of infection. The increasing rate of elective caesarean section (42%) is not related to maternal, foetal or obstetrical indications, but its use as an intervention to reduce HIV vertical transmission. CONCLUSIONS: The identification of HIV-infected women during pregnancy or before delivery ensures the appropriate management of the woman and her child, and clinicians should be aware of the increasing number of women with heterosexual acquisition of HIV-infection who may be less easily identified. PMID- 10869788 TI - Clinical value of detecting microalbuminuria as a risk factor for pregnancy induced hypertension in insulin-treated diabetic pregnancies. AB - OBJECTIVE: We evaluated the role of clinical non-overt stage III diabetic nephropathy concerning the development of more frequent hypertensive complications during pregnancies of women requiring insulin. METHODS: 122 unselected pregnant women treated with insulin were enrolled in the study, of whom 56 were type-1 diabetic patients and 66 patients had gestational diabetes. In 24-h urine samples, excretion rates of albumin (UAE) and beta(2)-microglobulin were determined by nephelometric analysis and a radioimmunoassay, respectively. These parameters were also measured in the serum as well as HbA(1c), fructosamine and daily blood glucose profiles. RESULTS: Sixteen (15.7%) women had an elevated UAE (>30 mg/24 h), of whom three had macroalbuminuria (UAE>300 mg/24 h). Thirteen (12.7%) insulin-treated women showed microalbuminuria during pregnancy, eight with type-1 diabetes and five with gestational diabetes requiring insulin. In the entire group hypertension was observed in seven (6.9%) women of whom six had microalbuminuria. The sensitivity and specificity were 85.7% and 92.6%, respectively. The positive predictive value reached 46.2%, whereas the negative predictive value was 98.9%. CONCLUSION: Measurement of UAE in diabetic pregnancies seems to be an useful additional parameter for risk evaluation of hypertensive disorders. PMID- 10869789 TI - Intrauterine infection, magnesium sulfate exposure and cerebral palsy in infants born between 26 and 30 weeks of gestation. AB - OBJECTIVE: To identify prenatal events associated with cerebral palsy (CP) in infants born between 26 and 30 weeks of gestation. STUDY DESIGN: Case (n=22) control (n=170) study was performed using a logistic regression model. RESULTS: Significant association of intrauterine infection with increased risk of CP was found in a logistic regression model that controlled for abnormal FHR patterns, placental infection, fetal acidosis at birth (umbilical artery pH<7. 1), and low Apgar score (<7) (odds ratio (OR) 5.47, 95% confidence interval (CI) 1.46-20.4). Magnesium sulfate exposure was associated with decreased risk (OR 0.13, CI 0.03 0.66) after exclusion of premature rupture of the membranes and abruptio placentae. In the magnesium exposure group, cases were infants born less than 28 weeks of gestation (3/21 vs. 0/61, P=0.015). CONCLUSION: In this case-control study, both intrauterine infection and magnesium sulfate exposure were significant factors related to the occurrence of cerebral palsy. PMID- 10869790 TI - Metastasectomy, a feasible treatment in selected cases with gynecologic malignancy. AB - We present the course of six gynecological patients who underwent surgical intervention because of a solitary metastasis. After a considerable follow-up period five patients are alive without evidence of disease and with a good quality of life. Metastasectomy should play a role in the management of gynecologic malignancies. PMID- 10869791 TI - Primary carcinoma of the fallopian tube: study of 11 cases. AB - OBJECTIVE: Primary fallopian tube carcinoma is a rare tumor that histologically and clinically resembles primary ovarian carcinoma. The purpose of this study was to present the experience of the Soroka Medical Center (SMC), Beer-Sheva, Israel of handling this tumor. STUDY DESIGN: Data from the files of 11 patients with primary fallopian tube carcinoma who were managed at the SMC between January 1978 and December 1998 were evaluated. RESULTS: The mean age of the patients was 59.4 years. Presenting symptoms and signs included abdominal pain, postmenopausal bleeding, watery vaginal discharge and adnexal mass. In all patients, the diagnosis of primary fallopian tube carcinoma was not made preoperatively. In ten patients in whom the adnexal mass was discovered preoperatively it was thought to be an ovarian tumor and in one patient the adnexal mass was first noticed during vaginal hysterectomy. Postoperatively, multi-drug chemotherapy was given to seven patients, multi-drug chemotherapy followed by pelvic radiotherapy to one patient, pelvic radiotherapy followed by single-agent chemotherapy to two patients, and one patient received no further treatment. The actuarial 5-year survival rate was 50%. CONCLUSIONS: Fallopian tube carcinoma is rarely suspected preoperatively. The symptom complex of 'hydrops tubae profluence', said to be pathognomonic for this tumor, is rarely encountered. The treatment approach is similar to that used for ovarian carcinoma and includes primary surgery comprised of total abdominal hysterectomy, bilateral salpingo-oophorectomy and staging followed by chemotherapy. The prognosis of patients with primary fallopian tube carcinoma is similar to that of patients with primary ovarian carcinoma. PMID- 10869792 TI - ICSI outcome in patients of 40 years age and over: a retrospective analysis. AB - OBJECTIVE: To report and analyse our experience with ICSI treatment in infertile women >/=40 years of age, with the intention of contributing to current debates on the effect of aging on the reproductive potential. STUDY DESIGN: 107 infertile couples in which the female partner was aged >/=40 years and who received ICSI treatment between January 1996 and December 1998. RESULTS: A total of 107 women underwent 171 treatment cycles during this period. Of 171 cycles initiated 33 were cancelled (cancellation rate=19.3%). In this way, 17 women did not have embryo transfer at all, while 90 patients had 138 cycles with oocyte retrieval and successful embryo transfer, with a mean number of embryos per transfer 2.36. Sixteen pregnancies occurred and eight of them ended in spontaneous abortion. The implantation rate was 4.9%, the pregnancy rate per initiated cycle was 9.35% and per transfer cycle 11.59%. The miscarriage rate was 50%. Moreover, 12 patients had supernumerary embryos, that were cryopreserved and transferred in 17 thawing cycles and resulted in two pregnancies ending in abortion. All pregnancies occurred when three embryos were available, except in two cases with two available embryos. The great majority of the total pregnancies (16 of 18) resulting in women aged between 40 and 42 years. CONCLUSION: Our data show that women 40 and older with existing ovarian function may benefit from ICSI treatment, even when the indication for treatment is male factor infertility. Supernumerary embryos, that are cryopreserved and transferred in subsequent cycles can improve the overall pregnancy rates per oocyte retrieval, although these women should be aware of the very high risk of miscarriage. PMID- 10869793 TI - Clinical implications of the didelphic uterus: long-term follow-up of 49 cases. AB - OBJECTIVE: The aim of the study was to evaluate reproductive performance of women with didelphic uterus and to consider possible long-term consequences associated with this uterine anomaly. STUDY DESIGN: Forty-nine women were diagnosed as having a didelphic uterus with a longitudinal vaginal septum at Tampere University Hospital, Finland between 1962 and 1998. The presence of other anomalies, gynecologic disorders, fertility and outcome of pregnancies were reviewed. The long-term clinical implications associated with a didelphic uterus were evaluated during the mean (S.D.) follow-up period of 9.1 (6.3) years. RESULTS: An obstructed hemivagina was found in nine (18%) out of forty-nine cases; eight of these had ipsilateral renal agenesis. A longitudinal vaginal septum was excised in twenty-six (53%) cases, but metroplasty in none. Five (13%) patients had primary infertility. Thirty-four (94%) out of thirty-six women who wanted to conceive had at least one pregnancy, and they produced seventy-one pregnancies; 21% miscarried, and ectopic pregnancy occurred in 2%. The fetal survival rate was 75%, prematurity 24%, fetal growth retardation 11%, perinatal mortality 5. 3%, and cesarean section rate 84%. Pregnancy located more commonly (76%) in the right uterus than in the left. During the follow-up period endometriosis was observed in seven (16%) out of forty-five cases. Ovarian neoplasm was found in four (9%) cases, one of them had ovarian cancer. CONCLUSIONS: Fertility in women with didelphic uterus is not notably impaired. The prognosis of pregnancy is comparatively good, while prematurity and fetal growth retardation indicate meticulous prenatal care. Long-term follow-up did not reveal that didelphic uterus is associated with increased frequency of endometriosis or genital neoplasm. PMID- 10869794 TI - (A)symptomatic necrotizing arteritis of the female genital tract. AB - AIMS: The vasculitides represent a heterogenous set of disorders that differ in prognosis and response to therapy. Beside systemic vasculitides, the development of localized forms of arteritis is well known though uncommon and the etiopathogenesis is not yet definitely clear. METHODS: Patients with necrotizing arteritis of the female genital tract proven by histology are studied in a retrospective analysis. RESULTS: Three cases of necrotizing arteritis with histological features of panarteritis nodosa apparently confined to the female genital tract are presented. None of these patients had prior history of systemic vasculitis. The acute necrotizing vasculitis was confined only to the uterine cervix in two patients and involved all the internal genital organs in the third patient. The patients have been observed for up to 4 years without any therapy for these lesions and without any manifestation of systemic vasculitic progression. CONCLUSION: It is to speculate that focal arteritis of the female genital tract is a benign form of panarteritis nodosa or moreover a totally different entity with identical morphogenesis but possibly different pathogenesis. Furthermore it seems to be important to be aware of the specificity of focal arteritis in female genital tract as distinct from the generalized form to prevent unnecessary surgical or chemotherapeutical therapy for this lesion. The benign entity of local arteritis in the female genital tract is discussed in contrast to the severe prognosis of systemic panarteritis nodosa. PMID- 10869795 TI - Factor V Leiden mutation and the risk of thrombo-embolic disease in pregnancy: a case report. AB - Factor V Leiden mutation is a risk factor for the development of thrombo-embolic episodes in pregnancy. A case is presented of a pregnant woman with repeated episodes of venous thrombosis with a complicated clinical course. PMID- 10869796 TI - Mediastinal metastasis of ovarian carcinoma. AB - Mediastinal metastasis of ovarian tumors are not rare as autopsy findings. Ovarian carcinomas usually spread by transcaelomic, lymphatic or haematogenous dissemination to peritoneum, pelvic and para-aortic lymph nodes, lung and pleura. A case of mediastinal metastasis of ovarian carcinoma is reported. A retrosternal mass was identified by CT scan and resected by VTC surgery. PMID- 10869797 TI - Rectal carcinoma during pregnancy: a reminder and updated treatment protocols. AB - Rectal carcinoma is rare during pregnancy. Prognosis is usually unfavorable due to late diagnosis, and management, especially regarding the mode of delivery, is controversial. Current treatment of rectal carcinoma includes neoadjuvant chemoradiotherapy, which may influence obstetrical management. We present a case report and discuss obstetrical management in view of updated knowledge and therapeutic approaches. PMID- 10869798 TI - The eighth meeting and exchange programme of European obstetricians and gynaecologists in training (Paris, 3rd-5th December 1998). PMID- 10869802 TI - Reorganization of adult rat barrel cortex intrinsic signals following kainic acid induced central lesion. AB - A plasticity model studying the adult rat barrel cortex intrinsic signal after a central lesion was developed. Repeated optical imaging studies of the barrel cortex of five rats were performed over variable periods of time (2 days to 6 weeks) after intracortical injection of kainic acid. The signal of the elicited principal whisker corresponding to the injected barrel in the repeat studies relocated to the perimeter of the lesion. The area of the signals of this principal whisker and of surrounding whiskers were larger in the first two weeks studies than those obtained before injection (P<0.01) resulting in increase overlapping of adjacent signals (P=0.01). Even though the signal of the PW remains relocated in the later studies (>2 weeks), all the signals returned to normal size. These findings demonstrate recovery and reorganization of sensory representation in the somatosensory cortex following a central lesion. PMID- 10869801 TI - Antiamnesic effect of metoprine and of selective histamine H(1) receptor agonists in a modified mouse passive avoidance test. AB - The aim of this study was to elucidate the effect caused by the inhibition of histamine catabolism by means of metoprine and the activation of histamine H(1) receptors by selective agonists on learning and memory processes, using a modified method of the mouse passive avoidance test. The administration of scopolamine 1 mg/kg (i. p.) immediately after the training session caused statistically-significant amnesia during the retention trial performed 24 h later. Piracetam (30 mg/kg (i.p.)), used as a positive control, and administered 20 min before the training session, prevented scopolamine-induced memory impairment. The histamine-N-methyltransferase inhibitor, metoprine, (2 and 5 mg/kg (s.c.)) had effects similar to those of this nootropic drug. The highly selective H(1) receptor agonist, 2-(3-trifluoromethylphenyl)histamine (FMPH) (2.65 and 6.5 microg/mouse (i.c.v.)) and the less selective agonist, 2 thiazolylethylamine (2-TEA) (0.1 and 0.3 microg/mouse (i.c.v.)) both antagonized the scopolamine-induced amnesia significantly and in a dose-related manner. The selective H(1) receptor antagonist, pyrilamine (20 mg/kg (i.p.)), revealed no effect by itself, but significantly prevented the antiamnesic action both that of the H(1) receptor agonists, and that of endogenous histamine, released by metoprine, thus suggesting a cognitive improvement via the activation of H(1) receptors. PMID- 10869803 TI - Similar levels of long-term potentiation in amyloid precursor protein -null and wild-type mice in the CA1 region of picrotoxin treated slices. AB - Although mutations in amyloid precursor protein (APP) are known to be involved in the development of Alzheimer's disease in some individuals, the role of this protein in normal brain function is poorly understood. We have reported previously that in APP-null mice long-term potentiation (LTP) in the CA1 region of the hippocampus is present but its magnitude is reduced compared to wild-type littermate controls. In the present study, we have confirmed this deficit using a different theta burst induction protocol. Significantly, however, we find that this deficit is no longer apparent when LTP experiments are performed following blockade of gamma-aminobutyric acid(A) receptors. These results suggest that the LTP process per se is not altered by the absence of APP. The deficit may therefore be an indirect consequence of other changes in the hippocampus that occur in the APP-null animal. PMID- 10869804 TI - Classical conditioning of oxidative DNA damage in rats. AB - This study investigated whether the formation of 8-hydroxydeoxyguanosine (8-OH dG), a known oxidative DNA modification relevant to carcinogenicity, can be classically conditioned to a novel taste in order to clarify the possible role of the central nervous system (CNS) or psychological stress on cancer initiation via a classical conditioning mechanism. Male Wistar rats underwent one or two conditioned taste aversion (CTA) experiments in which ferric nitrilotriacetate (Fe-NTA), which has renal toxicity and can induce renal cell carcinoma, served as a visceral unconditioned stimulus (US), and a saccharin solution (SAC) was used as a conditioned stimulus (CS). The 8-OH-dG levels in the group conditioned with the combination of SAC and Fe-NTA significantly increased as compared to those of the uncombined groups by two repeats of the conditioning procedure (P=0.013). The rats that showed a painful response at the Fe-NTA administration had significantly higher values of 8-OH-dG than those without pain (P=0. 003). These results not only provide the first evidence regarding classical conditioning of oxidative DNA damage using the CTA procedure, but also suggest the involvement of the CNS and psychological stress in the pathogenesis of cancer via oxidative DNA damage. PMID- 10869805 TI - Effects of ambient light on body temperature regulation in resting and exercising rats. AB - We investigated the effects of environmental light and darkness on thermoregulation during both daytime and nighttime by monitoring body temperature (T(b)) and physical activity of rats using a telemetry system. Experiments were performed in both resting and exercising rats. In resting rats, lights-off during the daytime resulted in an increase in both T(b) and activity. Conversely, during the nighttime, T(b) decreased with the lights-on stimulus despite the fact that the activity was left unchanged. Treadmill exercise (10 m/min) always increased T(b) from the basal resting level. In both daytime and nighttime, exercising rats exhibited a persistent T(b)-rise when lights were on. However, in the lights-off condition at nighttime, the T(b) of exercising rats increased to a level significantly higher than that of exercising rats with the lights-on. Our results suggest that light at nighttime causes the suppression of T(b) in both resting and exercising rats. PMID- 10869806 TI - Genetic polymorphisms in the cathespin D and interleukin-6 genes and the risk of Alzheimer's disease. AB - Alzheimer's disease (AD) is a complex multi-factorial disease with the involvement of several possible genes. The apolipoprotein E*4 (APOE*4) allele of the known susceptibility gene, APOE, is neither necessary nor sufficient to cause AD. This has prompted the search for other candidate genes associated with the risk of AD. Cathepsin D (Cat D) is an intracellular aspartyl protease that has been reported to have in vitro beta and gamma-secretase activity. The presence of a C-->T (Ala-->Val) polymorphism in exon 2 of the Cat D gene has been reported to be associated with an increased risk of AD. Further, as inflammation is reported to play a prominent role is AD pathogenesis, IL-6, a known mediator of inflammation, is another candidate gene proposed to be associated with the risk of AD. Genetic variation in the IL-6 gene has been reported to be associated with the risk of AD. We performed a genetic screening of sporadic, late-onset AD cases and age-matched controls to evaluate the role of Cat D and IL-6 polymorphisms in AD. Our data indicate no significant association between these polymorphisms and the risk of AD. When the data were stratified by APOE*4 status, no significant difference was seen either between cases and controls. These data suggest that the Cat D and IL-6 polymorphisms do not significantly alter the risk of AD in our case-control cohort. PMID- 10869807 TI - Temporal profile of cerebrospinal fluid glutamate, interleukin-6, and tumor necrosis factor-alpha in relation to brain edema and contusion following controlled cortical impact injury in rats. AB - Traumatic brain injury is associated with release of the excitotoxin glutamate and production of pro-inflammatory cytokines IL-6 and tumor necrosis factor-alpha (TNF-alpha). Following controlled cortical impact injury, cerebrospinal fluid (CSF) glutamate, IL-6, and TNF-alpha concentrations were measured to investigate their relationship to evolving tissue damage. Compared to non-traumatized rats CSF glutamate, IL-6 and TNF-alpha levels were significantly increased by 8 h after trauma (P<0.005). Parallel to increasing brain swelling and contusion CSF glutamate was significantly elevated over time, reaching highest levels by 48 h (33+/-4 microM) while IL-6 and TNF-alpha showed maximum values at 24 h after trauma (42+/-7 and 4.7+/-1 pg/ml) (P<0.005). The observed different temporal profile of CSF glutamate, IL-6, and TNF-alpha following focal traumatic brain injury could be of therapeutic importance. PMID- 10869808 TI - Immunocytochemical evidence for a progesterone receptor in neurons and glial cells of the rat spinal cord. AB - Using the KC 146 monoclonal antibody recognizing the B-form of the progesterone receptor (PR) and immunocytochemical techniques, we investigated if PR immunoreactive cells are present in the rat spinal cord. Neurons from ventral horn Lamina IX, glial cells in gray and white matter and ependymal cells were PR positive. Evidence for estrogen-inducibility of PR in ovariectomized rats was not observed. There were no significant gender differences in neuronal PR immunostaining intensity in the spinal cord, measured by computerized image analysis. In pituitary and uterus from estrogenized female rats, PR showed a strict nuclear localization, whereas in neurons and glial cells of the spinal cord, PR localized in cytoplasm and/or nucleus and in some cell processes. This receptor may be implicated in some of the biological effects of progesterone described in the spinal cord. PMID- 10869809 TI - Activity of neurons in the nucleus of the solitary tract of rats: effect of osmotic and mechanical stimuli. AB - Patch-clamp recordings were used to examine the osmosensitivity and mechanosensitivity of neurons in the caudal part of the nucleus tractus solitarius in coronal slices from rat brain. Firing rates and membrane potentials were measured as slices were exposed to perfusate which varied in its osmolality and/or sodium concentration. In all cells tested, the responses to change in the sodium concentration of perfusate were duplicated by osmolality changes of sucrose or mannitol. When nucleus tractus solitarius cells were tested with changes in pressure applied via the pipette, responses to positive or negative pressure paralleled their responses to osmotic stimulation. We suggest that a mechanosensitive receptor exists on osmosensitive neurons within the nucleus tractus solitarius, and this receptor may be responsible for changes in the firing rate and membrane potential which occur in the nucleus tractus solitarius neurons. PMID- 10869811 TI - Oral administration of the anti-inflammatory substance triflusal results in the downregulation of constitutive transcription factor NF-kappaB in the postnatal rat brain. AB - In this study we have evaluated the in vivo ability of triflusal (2-acetoxy-4-tri fluoromethylbenzoic acid) to inhibit constitutive nuclear factor-kappa B (NF kappaB) activation in the brain of postnatal rats. One week old Long-Evans black hooded rat pups received three oral administrations of triflusal (30 mg/kg) and were sacrificed at 9 days of age. After fixation, brains were cut in a cryostat and processed immunocytochemically for the demonstration of NF-kappaB. In control postnatal rats, NF-kappaB is constitutively present in some neuronal populations and in glial cells of white matter tracts. In contrast, triflusal treated rats showed a drastic downregulation of neuronal and glial NF-kappaB, both in the number of labelled cells and in the intensity of staining. The inhibition of NF kappaB activation could be an important step in the modulation of inflammatory processes occurring after several pathological conditions. PMID- 10869810 TI - Induction of rat L-phosphoserine phosphatase by amyloid-beta (1-42) is inhibited by interleukin-11. AB - Alzheimer's disease (AD) is characterized by the presence of beta-amyloid (Abeta) protein deposits in the brain and increased Abeta (1-42) peptide production is thought to be one of the early events in the pathogenesis of AD that leads to progressive neurodegenerative processes and dementia. Using cDNA subtraction and reverse transcription-polymerase chain reaction, we examined the Abeta (1-42) peptide-induced gene expression in rat neuroblastoma B104 cells. In addition we hypothesized that interleukin-11 (IL-11) supports neuronal survival. We found that Abeta (1-42) activates L-phosphoserine phosphatase in neuronal cells which is inhibited by IL-11. Moreover, IL-11 inhibits Abeta (1-42)-induced neurotoxicity in a dose-dependent manner. Our study suggests that L-phosphoserine phosphatase may play a role in altered neuronal function in AD via enhancing glutamate-induced neurotoxicity by D-serine and the IL-11 receptor system may act as a neuroprotective cytokine in human brain. PMID- 10869812 TI - Decreased nuclear factor-kappaB DNA binding activity following permanent focal cerebral ischaemia in the rat. AB - Many factors implicated in the pathogenesis of cerebral ischaemia such as glutamate, tumour necrosis factor and interleukin-1 have also been shown to activate nuclear factor-kappaB (NF-kappaB). In the present study we have investigated NF-kappaB activity at various times following permanent focal cerebral ischaemia in rats using immunohistochemistry, western blotting and electrophoretic mobility shift assay (EMSA). Three hours following middle cerebral artery occlusion nuclear translocation of NF-kappaB was detected using immunohistochemical and western blotting techniques. This was reflected in a trend towards increased NF-kappaB binding activity (EMSA) in the ischaemic cortex compared to histologically normal tissue. In contrast however, from 6 to 48 h post-occlusion nuclear translocation and NF-kappaB binding activity was decreased in the ischaemic cortex. Decreased NF-kappaB binding activity detected in degenerating neurones, suggests that decreased NF-kappaB activity may exacerbate ischaemia induced neuronal cell death. PMID- 10869813 TI - Biological effects of tiagabine on primary cortical astrocyte cultures of rat. AB - Biological effects of tiagabine, a new antiepileptic drug, were analyzed on cultures of rat's cortical astrocytes. Tiagabine was added to the cultures at concentrations of 1 and 10 microg/ml, correspondent to therapeutic range; cell viability (tetrazolium salt assay and lactic dehydrogenase release), maturation and differentiation (glutamine synthetase activity) and presence of stress conditions (reactive oxygen species formation, inducible nitric oxide synthetase expression and 70 kDa heath shock protein production) were tested. Our results indicate that the addition of Tiagabine to primary astrocytes not only did not change significantly the examined metabolic activities but also seems to exert a protective action against oxidative stress. Thus, our data reinforce the idea that Tiagabine may be considered an effective promising drug in the treatment of epilepsy. PMID- 10869814 TI - Astrocytes in culture express the full-length Trk-B receptor and respond to brain derived neurotrophic factor by changing intracellular calcium levels: effect of ethanol exposure in rats. AB - Although cultured astroglial cells were reported to express exclusively the truncated non-catalytic Trk B receptor for brain-derived neurotrophic factor (BDNF), we detect here, using a sensitive ribonuclease protection assay, mRNAs for both truncated (TrkB-T) and the full length catalytic (TrkB-fl) form of BDNF receptor in developing cortical astrocytes and neurons in culture. Cortical neurons and immature astroglia, such as radial glia and proliferating astrocytes, express both the protein and mRNAs for TrkB-fl and TrkB-T, whereas the differentiation of astrocytes leads to a decrease in the trkB-fl mRNA, being the truncated TrkB the predominant receptor in differentiating and confluent astrocytes. The levels of TrkB-fl expression in proliferating and differentiating astrocytes and neurons correlates with the cell response to BDNF, monitored by the rise in intracellular [Ca(2+)](i). Foetal exposure to ethanol alters astroglial development and delays the reduction in trkB-fl mRNA levels observed with differentiation of astrocytes. These results demonstrate that immature astrocytes are able to express the catalytic Trk B receptors and to respond to BDNF with the activation of conventional signal transduction pathways. The results suggest that this signalling pathway is more activated in ethanol-exposed cells. PMID- 10869815 TI - The cAMP-dependent kinase pathway does not sensitize the cloned vanilloid receptor type 1 expressed in xenopus oocytes or Aplysia neurons. AB - Capsaicin-activated channels present in sensory neurons are ligand-gated cation channels that largely account for mediating some types of pain. The cAMP dependent protein kinase (PKA) signal pathway was suggested to mediate the prostaglandin-induced enhancement of capsaicin-evoked inward current (I(CAP)) in rat sensory neurons. It is not clear, however, whether PKA acts directly on the capsaicin-sensitive channel that is responsible for I(CAP). To address this issue, we overexpressed the cloned capsaicin receptor, VR1, in heterologous expression systems such as Xenopus oocytes or Aplysia R2 neuron and stimulated PKA pathways. As a result, activation of PKA by applying either 8-bromo-cAMP or forskolin with 3-isobutyl-1-methylxanthine or through activation of beta(2) adrenergic receptors failed to enhance I(CAP) in oocytes or R2 neurons expressing VR1. Our results raise two possibilities. (1) Direct phosphorylation of VR1 by PKA may not be responsible for the sensitization; instead, phosphorylation of regulatory proteins associated with VR1 would account for the sensitization of I(CAP) evoked by prostaglandin E(2) in dorsal root ganglion (DRG) neurons. (2) DRG neurons may have a different PKA signaling mechanism that is not replicable in Xenopus oocytes or Aplysia R2 neurons. PMID- 10869816 TI - The effect of intravenous insulin on accumulation of excitotoxic and other amino acids in the ischemic rat cerebral cortex. AB - Insulin has been reported to be neuroprotective during cerebral ischemia/reperfusion. However, it may also increase the sensitivity of cultured cortical neurons to glutamate toxicity. The experiments described here utilized a rat four-vessel occlusion model with cerebral cortical windows to determine the effects of intravenous insulin, alone (I) or combined with glucose (IG) to maintain physiologic blood glucose levels, on the extracellular accumulation of amino acids in superfusates of the cerebral cortex. Aspartate, phosphoethanolamine, taurine and gamma-aminobutyric acid were increased in the I and IG groups and glutamate was increased in the IG group compared to controls during ischemia/reperfusion. Insulin treatment attenuated the rebound in cortical superfusate glucose levels in both groups of animals during reperfusion. The increases in amino acid release during reperfusion may be due to a lack of glycolytically derived energy available for amino acid uptake systems and ionic pumps. PMID- 10869817 TI - Reduced expression of insulin-like growth factor 1 messenger RNA in the hippocampus of aged rats. AB - Adult neurones remain dependent on neurotrophic factors such as insulin-like growth factor 1 (IGF-1) to sustain neuronal viability by maintaining cell phenotype, supporting synaptic plasticity and providing neuroprotective and neuroregenerative mechanisms. A decline in the cellular expression of neurotrophic factors has been speculated to contribute to the age-related changes that occur in the brain, where the hippocampus appears particularly susceptible. Using in situ hybridisation, we have made a detailed comparison of the expression of IGF-1 mRNA in the hippocampal formation between young (6 months) and aged (23 months) rats. IGF-1 mRNA expression was measured from cell populations containing only high density radioactive labelling (>20 grains/cell) to avoid ambiguity of signal. The amount of IGF-1 mRNA signal was significantly lower in cells of the alveus (P<0.05) and stratum lacunosum moleculare (P<0.01) of aged compared to young rats. These findings challenge reports that IGF-1 mRNA is unaltered in the ageing hippocampus and provide further evidence that changes in the IGF-1 system is a significant factor in the progressive age-related deterioration of normal neuronal function. PMID- 10869818 TI - Neuroprotection of nerve growth factor-loaded microspheres on the D2 dopaminergic receptor positive-striatal neurones in quinolinic acid-lesioned rats: a quantitative autoradiographic assessment with iodobenzamide. AB - Huntington's disease (HD) results from the degeneration of striatal neurones, mainly gamma-aminobutyric acid (GABA)ergic projection neurones and lately cholinergic interneurones. The use of trophic factors as agents able to prevent such neural degeneration is a promising strategy. The aim of this study was to validate nerve growth factor-loaded (NGF-loaded) poly-D,L-lactide-co-glycolide (PLGA) microspheres for treatment of HD in a rat model with quinolinic acid lesion using autoradiographic study of D2 dopaminergic receptors (D2R). This target is expressed by about half of striatal neurones and its scintigraphic exploration has already been performed for the follow-up of this degenerative process. Ex vivo autoradiography of D2R performed with iodobenzamide, the widely used ligand for single photo emission computerized tomography, revealed slight neuroprotection. Moreover, tolerance of microspheres was demonstrated by in vitro autoradiography with the marker of gliosis, [(3)H]-PK 11195. PMID- 10869819 TI - Dual effects of dextromethorphan on cocaine-induced conditioned place preference in mice. AB - Dextromethorphan (DM) at supra-antitussive doses might produce psychotomimetic effects in humans. In order to understand the underlying mechanisms responsible for the behavior induced by DM, we examined the effects of DM on cocaine-induced conditioned place preference (CPP) and locomotor pattern in mice, and Fos-related antigen immunoreactivity (FRA-IR) in the striatal complex (nucleus accumbens and striatum) of the mouse brain. The effects of DM (20 and 40 mg/kg, i.p.) on the CPP for 2.5, 5, 10, and 20 mg cocaine/kg, i.p. were assessed. Pretreatment with DM dose-dependently decreased the CPP for 20 mg cocaine/kg. Similarly, pretreatment with DM appeared to reduce the CPP for 10 mg cocaine/kg, but increase the CPP for 5 mg cocaine/kg. This finding was more pronounced for 2.5 mg cocaine/kg; DM significantly increased the CPP for 2.5 mg cocaine/kg in a dose related manner. Furthermore, these results were correlated with alterations in the locomotor pattern (marginal activity) and FRA-IR in the striatal complex. Thus, our results suggest that DM exhibits a biphasic effect on the cocaine induced CPP and locomotor pattern. PMID- 10869820 TI - The promoter of human acetylcholinesterase is activated by a cyclic adenosine 3',5'-monophosphate-dependent pathway in cultured NG108-15 neuroblastoma cells. AB - Different transcription elements have been proposed to play a role in the regulation of acetylcholinesterase (AChE) in muscle and neuron, and cyclic adenosine 3',5'-monophosphate (cAMP)-dependent pathway is one of them. In order to test the possible role of cAMP in regulating the expression of human AChE, an approximately 2.2 kb DNA fragment of human AChE promoter was linked up stream to a luciferase reporter. The chimeric DNA was transfected into cultured NG108-15 neuroblastoma cells. Application of Bt(2)-cAMP and forskolin increased the promoter driven luciferase activity over 2-fold in the transfected NG108-15 cells; the increase was parallel to the activation of endogenous AChE protein and enzymatic activity. The intracellular cAMP level was increased in the Galpha(sQL) (constitutively active mutant of Galpha(s)) cDNA transfected NG108-15 cells. The Galpha(sQL) cDNA transfected cells showed an increase of over 10-fold in the luciferase activity. In addition, a constitutively active mutant of activating transcription factor-1 (ATF-1) was able to turn on human AChE promoter by approximately 4-fold when they were co-expressed in the neuroblastoma cells. These results support the involvement of a cAMP-dependent pathway in regulating the expression of human AChE. PMID- 10869821 TI - Are nitrite/nitrate (NOx) levels elevated by inhalation injury? AB - The relationship between airway burn and nitric oxide (NO) levels in the early burn stage was investigated by quantifying nitrite/nitrate (NOx), which are the final metabolites of NO, in 22 burn patients. Although total body surface area (TBSA) and burn index (BI) were significantly higher in patients with airway-burn than in patients without airway burn (P=0.0347 and 0.0422, respectively), no significant difference in NOx levels was observed between the two groups (P=0.6196). The NOx levels were found to correlate significantly with TBSA (r=0.4775, P=0.0246). A significant correlation was also noted between the NOx levels and BI (r=0.4391, P=0.0409). These results suggest that NO reflects the intensity of inflammation in the early burn stage, but that excessive NO formation is unlikely to be induced by stress, such as that caused by airway burn. PMID- 10869822 TI - The relationship between tumor necrosis factor (TNF)-alpha and survival following granulocyte-colony stimulating factor (G-CSF) administration in burn sepsis. AB - Blood levels of tumor necrosis factor (TNF)-alpha were determined in 78 patients with burn sepsis. Of these patients, 51 were managed with additional administration of granulocyte colony-stimulating factor (G-CSF) in addition to routine treatment procedures (group A), while 27 received only routine treatment (group B). G-CSF was administrated for at least nine and at most 14 days; doses were gradually decreased in each 3 day period. On the 1st, 4th, 7th, 10th and 15th days, blood levels of TNF-alpha were determined. We sought to determine whether TNF alpha levels had a prognostic value in the management of burn induced sepsis that was treated with G-CSF. In our study, patients with gradually decreasing TNF-alpha levels in the second 3 day period, were strong candidates for survival, because TNF-alpha levels decreased little in nonsurvivors but decreased greatly in survivors. The survival rate was 42/51 (82.3%) in group A and 9/27 (33.3%) in group B. In conclusion, G-CSF had positive effects on survival, and TNF-alpha was a predictor of prognosis in burn-induced sepsis. PMID- 10869823 TI - Increased activity of lymph node cells in experimental thermal injury: changes in accessory cells in injured area-draining lymph nodes. AB - Accessory cell content and some of their functional characteristics were determined in regional lymph nodes which drain burn injury (DLN) in rats. Increase in percentages of non-specific esterase-positive cells and NBT+ macrophages and in numbers of dendritic cells were noted in cytospin preparations of draining lymph node cells (DLC) 24 and 72 h following thermal injury. An accumulation of B cells was also noted in the DLN paracortex region at these time points. Enrichment of ED1+ (rat macrophage marker) cells was noted in the adherent DLC population. Increased activity of interleukin-1 (IL-1) in conditioned medium from adherent DLC population and the increased stimulatory capacity of whole DLC or dendritic cell enriched-DLC fraction were noted in functional assays. Enrichment in accessory cells and an increase in their functional activity could contribute to the endogenous activity of regional lymph nodes which drain burned areas. PMID- 10869824 TI - Long-term viability of cryopreserved cultured epithelial grafts. AB - Human cultured epithelial grafts are frozen for long-term preservation. To assess the viability of these stored grafts, their cell survival rate and colony-forming efficiency of grafts cryopreserved at -135 degrees C and at -80 degrees C were followed over time. Flow cytometry showed that the cell survival rate of the grafts cryopreserved at -135 degrees C for 1 month, 6 months and 1 year averaged 89.3%, 61.7% and 61.6%. Cryopreservation at -80 degrees C maintained cell survival rate as well for 1 month, but after 6 months of cryopreservation survival was reduced at -80 degrees C (35.2%) compared with that of -135 degrees C. In histological examination, the cell structure and basal layer were very well preserved after 6 months of storage at -135 degrees C, but not at -80 degrees C. Cell survival rate at -135 degrees C was also assessed by colony-forming efficiency. Colony-forming efficiency of the grafts cryopreserved for 1 month, 6 months and 1 year averaged 66.1%, 58.5% and 55.1% of control (noncryopreserved) grafts. These findings suggest that, even when cultured epithelial grafts are subjected to long-term cryopreservation, cell viability remains sufficient, reculturing is possible, and that graft banking could be used for clinical applications. PMID- 10869825 TI - Dissociation of osmoregulation from plasma arginine vasopressin levels following thermal injury in childhood. AB - Although the syndrome of inappropriate anti-diuretic hormone secretion has been recognised as a complication associated with burn and other trauma in adults, relatively little is known about its incidence in children. The objective of this study was to investigate whether it is a complication associated with burn injury in children. Plasma and urine levels of arginine vasopressin (anti-diuretic hormone), sodium and osmolality were measured in samples collected from 16 burn injured children admitted to the burns unit of the regional children's hospital. No significant correlations were found between plasma vasopressin and plasma sodium or osmolality levels, but there were significant correlations between plasma vasopressin and urine osmolality, 36 (r=0.74, p=0. 009), 60 (r=0.92, p=0.000) and 84 h (r=0.84, p=0.001) after admission, respectively. There were also significant correlations between plasma sodium and plasma osmolality, 24 (r=0.7, p=0.005), 36 (r=0.57, p=0.04) and 84 h (r=0.84, p=0.004) after admission. The data suggest dissociation between the osmolar control of vasopressin secretion and vasopressin levels after burn injury in children, but do not support the incidence of inappropriate secretion of antidiuretic hormone. PMID- 10869826 TI - Anxiety: current practices in assessment and treatment of anxiety of burn patients. AB - Anxiety is an affective response commonly experienced by persons after emotional and physical trauma, as well as associated with aversive medical treatments. The scientific information related to the conceptualization, assessment, and treatment of anxiety is limited. In order to develop a pilot protocol for anxiety management, nursing directors at 64 burn centers were surveyed. At 89% of the centers, anxiety measures were not used. Most of the teams assess informally through observation of patient (n=21), dialogue with patient (n=12), or both observation and dialogue with patient (n=15). Assessors of anxiety range in breadth from nurse only to the entire burn team, including pastoral care representatives and family. The class of medication most frequently endorsed in treating anxiety is the benzodiazipine, most often lorazepam (Ativan). A number of non-pharmacologic techniques are used to manage anxiety, e.g., muscle relaxation, breathing, imagery. Consideration should be given to assessing anxiety systematically, so knowledge can be gleaned and applied to conceptualization of symptom presentation and application of treatment resources. PMID- 10869827 TI - Utility of an intervention scoring system in documenting effects of changes in burn treatment. AB - The consequences of the introduction of a program of consistent use of topical antimicrobials and early aggressive excision of deep burn wounds by utilizing a comprehensive, computerized patient registry/therapeutic intervention scoring system, were investigated. Prospectively, the clinical course, mortality, outcome and hospital costs were compared for the year preceding (89 patients) and the 4 years following (226 patients) the introduction of the new treatment program. It was found that mortality decreased from 10.1 to 4.6% after change in therapy (P<0.001), despite an increase in mean burn extent. The length of hospital stay per % burn surface area declined from 1.2 to 1.0 days (P<0.001). The number and complexity of therapeutic interventions and the associated costs, also declined. Patients in the new treatment program had a better level of physical and psychosocial function at follow up. In conclusion, the introduction of a program of consistent use of topical antimicrobials and early, aggressive surgical excision was associated with an improved outcome at lesser cost. The combined registry-intervention scoring system permits ready analysis of results using data entered on a daily, near-real time basis. PMID- 10869828 TI - Burns in epileptics in Saudi Arabia. AB - In a prospective study, 10 cases of burns in epileptics in Saudi Arabia, occurring over a five year period, were included. All burns were sustained during seizures. All patients were adults, and eight out of the 10 patients were female. The percentage of burn varied between 1 and 30% of the total body surface area. Four cases sustained the burn injury while fasting during the Arabic month of Ramadan because they did not take their anti-epileptic medication. Eight of the 10 burn injuries were sustained in the kitchen while cooking or ironing. One male patient had his epileptic fit at noon time while crossing the street. Contact with the hot ground in this patient resulted in full thickness burns. Recommendations regarding prevention of burn injuries in epileptics in Saudi Arabia are given. PMID- 10869829 TI - Work-related burns in South Wales 1995-96. AB - The aim of this work was to evaluate the epidemiology of occupational burns referred to the Welsh Regional Burns Unit from 1st January 1995 to 31st December 1996. Three hundred and twenty-four patients were identified as having sustained occupational burns and the case notes of 319 were available for review. Data on age, sex, occupation, aetiology, percentage and site of burn, treatment, complications and length of hospital stay were recorded. Twenty percent of all burns referred to our unit occurred in the workplace, the majority of whom were male (male:female 11:1), with a mean age of 34 years. Patients presented late to our unit in 35% of cases, with an average delay of 5 days. Chemical burns predominated (23%), followed by flame (14%) and scald (14%). Small burns (< or =1% TBSA) were seen in 70% of all patients. Five patients had burns involving >15% TBSA. One hundred and seventy-five patients were admitted, of whom 79 required surgery. The length of stay ranged from 1-110 days (mean 8.5), with an average follow-up for all patients of 3.5 months. One patient died as a result of his burn injury. In conclusion, occupational burn injuries continue to account for a significant proportion of all burn injuries, affecting mainly young males in physical occupations. Despite Health and Safety guidance, chemical burns are the predominant cause and more needs to be done to educate those working with chemicals to prevent injury. PMID- 10869830 TI - Preauricular flap for post burn ear lobe reconstruction - a case report. AB - Burns of the face often affect the ear causing deformation of the pinna or ear lobe due to perichondritis or loss of cartilage and excess scarring. The reconstruction of the ear lobe may be limited due to scarring of the surrounding tissues. We describe a preauricular flap for reconstruction of the anterior and posterior surface of the ear lobe following burn injury. This two stage technique of ear lobe reconstruction by a preauricular flap is not described in the available literature. An inferiorly based preauricular flap was harvested in a young lady who had post burn loss of the right ear lobe. This was folded on itself and sutured to reconstruct both surfaces of the ear lobe. After 6 months, it was released from its attachment at the retromandibular area to define the cheek lobe angle and to shape it. The preauricular flap for ear lobe contouring is a simple and easy technique. An adequate length may be harvested without significant donor site morbidity. PMID- 10869831 TI - Medium thickness plantar skin graft for the management of digital and palmar flexion contractures. AB - The fundamental principle of reconstruction is to replace the lost tissue with similar tissue. Because of the unique similarities in characteristics, plantar skin is an excellent donor area for skin grafting for the palmar aspect of the digits and hand. It provides an ideal color and texture match and long durability with an inconspicuous donor site. However, the use of plantar skin in management of digital and palmar defects is only rarely reported in the literature. Doubts were raised on donor site healing and incidence of recurrence of contracture. There are reports of using full thickness skin graft and dermal grafting from planta in similar cases. We have used medium thickness split grafts from sole of foot in correcting 262 cases of flexion contracture of digits and palms. Thick split skin grafts, full thickness grafts, local flaps and Z plasties were used in numbers of other cases. The technique of plantar skin grafts, application and results with reference to take of graft, quality of skin, donor area healing and recurrence are discussed and compared with available methods and studies. PMID- 10869832 TI - Surgical management of post-burn skin dyspigmentation of the upper limb. AB - The technique and results of surgery in 23 patients with skin dyspigmentation of the upper limb are presented. The study population was divided into two groups. Group A (n=15 patients) had hyperpigmented skin grafts in the palm or the palmar aspect of the digits. All patients underwent excision of the hyperpigmented grafts and coverage with split-thickness plantar skin grafts. The plantar grafts gave an excellent colour and texture match and all patients were satisfied with the result. Group B (n=8 patients) had post-burn hypopigmentation on the dorsal aspect of digits, hand or forearm. These patients underwent dermabrasion and thin split-thickness grafting harvested from the upper thigh or buttock. All grafts healed well with no residual hypopigmentation. However, the graft was slightly hyperpigmented when compared to the surrounding skin. Despite this complication, all patients were satisfied and considered this slight hyperpigmentation much better than the preoperative hypopigmentation. The pathogenesis of skin dyspigmentation and other treatment techniques are discussed. PMID- 10869833 TI - Successful treatment of a case of electrical burn with visceral injury and full thickness loss of the abdominal wall. AB - A 13-year-old male received high-voltage electrical burns with a resultant large direct wound on the upper abdomen involving the full-thickness of the abdominal wall, including the peritoneum. Early debridement, exploratory laparotomy and temporary restoration of the excised abdominal wall with a fascial prosthesis were carried out at 6 h post-burn. The bilateral upper and right lower limbs were amputated on the 10th post-burn day. The patient developed a 4x4 cm duodenocutaneous fistula on the 28th post-burn day, but was free of peritonitis. After 5 months of the conservative treatment, the fistula closed, and the abdominal wall defect was reconstructed with a free latissimus dorsi musculocutaneous flap. One month later, the patient was discharged following an uneventful recovery. PMID- 10869834 TI - Post-burn hallux varus: a case report and management of a rare deformity. AB - Post-burn hallux varus is an extremely rare condition. A 22-year-old male, with a history of campfire burns in childhood, presented with secondary hallux varus of the left great toe. Surgical correction included medial soft tissue release, metatarsophalangeal joint arthrodesis, two-pin fixation of bones, metatarsophalangeal joint capsulorrhaphy, and coverage of the skin defect with a "Z" plasty of the skin and split thickness skin grafting. Follow up 20 months later showed satisfactory results. PMID- 10869835 TI - Enhanced fos expression within the primary olfactory and limbic pathways induced by an aversive conditioned odor stimulus. AB - A central question in olfactory learning is how animals become tuned to odor stimuli that gain significance through conditioning. A leading view is that tuning to conditioned odor stimuli involves functional modifications within the primary olfactory pathways, themselves.(7) Here we studied this idea further by investigating responses within the olfactory system to an odor that had previously been paired with footshock in classical fear-conditioning trials in adult rats. Using the transcription factor Fos as a marker of odor-induced neuronal activation,(1,14) we found that in rats that had received forward pairings of odor and footshock during training, presentation of the conditioned odor stimulus, alone, produced an enhanced increase in levels of Fos in the main and accessory olfactory bulbs and anterior olfactory nucleus compared with that found in animals that had received backward presentations of the stimuli or of odor alone. These results demonstrate that Fos responses to an odor within the primary olfactory pathways can be modified through aversive conditioning, and are consistent with other evidence that olfactory conditioning can lead to functional changes within these pathways.(7) PMID- 10869836 TI - The multifarious hippocampal mossy fiber pathway: a review. AB - The hippocampal mossy fiber pathway between the granule cells of the dentate gyrus and the pyramidal cells of area CA3 has been the target of numerous scientific studies. Initially, attention was focused on the mossy fiber to CA3 pyramidal cell synapse because it was suggested to be a model synapse for studying the basic properties of synaptic transmission in the CNS. However, the accumulated body of research suggests that the mossy fiber synapse is rather unique in that it has many distinct features not usually observed in cortical synapses. In this review, we have attempted to summarize the many unique features of this hippocampal pathway. We also have attempted to reconcile some discrepancies that exist in the literature concerning the pharmacology, physiology and plasticity of this pathway. In addition we also point out some of the experimental challenges that make electrophysiological study of this pathway so difficult.Finally, we suggest that understanding the functional role of the hippocampal mossy fiber pathway may lie in an appreciation of its variety of unique properties that make it a strong yet broadly modulated synaptic input to postsynaptic targets in the hilus of the dentate gyrus and area CA3 of the hippocampal formation. PMID- 10869837 TI - Zinc-rich synaptic boutons in human temporal cortex biopsies. AB - The distribution of zinc-rich synaptic boutons in biopsies of the temporal cortex from epileptic patients who had undergone surgery is described. Unfixed cryostat sections were exposed to H(2)S vapour to precipitate endogenous zinc, which was subsequently shown by silver enhancement. In the temporal cortex, the stain for zinc was arranged in bands: stain was heavy in layers II and VI, moderate-to heavy in layers I, III and V, and low in layer IV. The white matter was virtually devoid of staining. At the electron microscope level, labelling was found in synaptic boutons that made asymmetric synaptic contacts. Immunohistochemical staining for glutamate receptor subunits GluR2/3 was observed in cell bodies in layers II, III, V and VI, coincident with the layers that showed heavy staining for zinc. Immunostaining for glutamate receptor subunit GluR1 was prominent in non-pyramidal neurons in deep cortical layers. These results support findings in other mammals and indicate that the human neocortex may contain an extensive system of zinc-rich cortico-cortical connections. This system may be altered in pathological conditions. PMID- 10869838 TI - Neocortical neurons lacking the protein-tyrosine kinase B receptor display abnormal differentiation and process elongation in vitro and in vivo. AB - The spatial and temporal expression of the protein-tyrosine kinase B (TrkB) receptor and its ligands has been correlated with the development of the neocortex. Activation of the receptor has been associated with neocortical neuronal survival, differentiation, connectivity and neurotransmitter release. Although such findings suggest an important role for TrkB signaling in corticogenesis, conclusive evidence from targeted gene deletion ("knockout"; TrkB -/-) mice has been limited, due in part to the neonatal lethality of most of these mutant mice and the confounding variables associated with the poor health of those few surviving slightly longer postnatally. In the present study, the effects of TrkB signaling on the survival, differentiation and integration of neocortical neurons was directly investigated in vitro and in vivo. First, we conducted a neuron-specific immunocytochemical analysis of TrkB -/- mice to determine whether early cortical structure and patterns of histogenesis were normal or perturbed. We then employed in vitro and in vivo approaches to extend the life of TrkB -/- neocortical neurons beyond the period possible in TrkB -/- mutant mice themselves: (i) dissociated cell culture to directly compare the developmental potential of TrkB -/-, +/- and +/+ neurons; and (ii) neural transplantation into homochronic wild-type recipients to investigate the cell autonomous effects of the receptor knockout on the differentiation, growth and integration of neocortical neurons. These latter experiments allowed, for the first time, study of the survival and differentiation potential of TrkB -/- neocortical neurons beyond the initial stages of corticogenesis. Direct comparison of brains of TrkB -/-, +/- and +/+ littermates immunocytochemically labeled with antibodies to microtubule-associated protein-2, neurofilament and beta-tubulin III revealed subtle anatomical anomalies in the mutant mice. These anomalies include abnormally diffuse microtubule-associated protein-2 positive neurons just dorsal to the corpus callosum, and heterotopic aggregations of postmitotic neurons in the subventricular zones of the ganglionic eminences, both suggesting delayed neuronal migration and differentiation. Cell culture experiments revealed substantially reduced survival by TrkB -/- neocortical neurons, and a significant reduction in neurite outgrowth by surviving TrkB -/- neurons. In experiments where prelabeled embryonic or neonatal TrkB -/- neocortical neurons were transplanted into the cerebral cortices of neonatal wild type recipients, a similar quantitatively significant defect in the formation of dendrites, as well as reduced integration of TrkB -/- neocortical neurons, was also evident. These findings demonstrate cell-autonomous abnormalities in the development of neocortical neurons from TrkB -/- mice, and the subtle, but potentially critical, role of protein-tyrosine kinase B signaling in neocortical neuronal survival, differentiation and connectivity. PMID- 10869839 TI - Cerebral cortical muscarinic cholinergic and N-methyl-D-aspartate receptors mediate increase in cortical blood flow elicited by chemical stimulation of periaqueductal gray matter. AB - The periaqueductal gray matter is implicated in the central processing of defensive reactions. We found previously that, when stimulated by N-methyl-D aspartate, the caudal third of its lateral subdivision elicited an increase in blood flow over widespread cerebral neocortical areas and that a major proportion of the flow increase was inhibited by topical cortical application of scopolamine, an antagonist of muscarinic receptors. The present study was undertaken to elucidate the roles of cortical nicotinic and excitatory amino acid receptors in the mediation of the flow increase in 66 anaesthetized, cervically cordotomized, artificially ventilated rats with open cranial windows. We found that the flow increase (laser-Doppler flowmetry) was resistant to antagonists of non-N-methyl-D-aspartate receptors and of nicotinic receptors. The response was, however, attenuated to a substantial extent by topical and intravenous N-methyl-D aspartate receptor antagonists, MK-801 and topical D(-)-2-amino-5 phosphonopentanoic acid. Combined topical application of the latter antagonist with scopolamine attenuated the flow increase to a further extent beyond that achieved with either of the antagonists alone. Topical applications of acetylcholine and N-methyl-D-aspartate individually increased the cortical blood flow. A modest synergism was observed between the actions of these two agonists.Overall, we suggest that in the face of stimuli which provoke defensive reactions, the periaqueductal gray matter may elicit an increase in cortical blood flow by utilizing the cortical acetylcholine-muscarinic receptor system and the cortical excitatory amino acid-N-methyl-D-aspartate receptor system. The vasomotor actions of these two transmitter-receptor systems may operate independently of each other as well as in harmony with each other. PMID- 10869840 TI - Priming pattern determines the correlation between hippocampal theta activity and locomotor stepping elicited by stimulation in anesthetized rats. AB - The after-effects of locomotor stimulation are a transient facilitation of locomotor initiation (the priming effect), and a transient increase in hippocampal rhythmic slow activity in the 3-6 Hz band of the theta range. The similar time course of the two effects suggests that hippocampal 3-6 Hz activity may be linked to the excitability of locomotor initiation. This study tested the hypothesis that power in the 3-6 Hz band that is present prior to stimulation would predict the magnitude of elicited stepping. Stimulation electrodes were implanted in 15 locomotor sites of 10 anesthetized rats (urethane, 800 mg/kg). Hindlimb stepping was elicited by a single control train of electrical stimulation presented once every 62 s. On test trials, a test train at the same intensity followed the control train at varying control/test intervals (15-36 s) to assess the priming effect on stepping. The priming pattern determined whether hippocampal 3-6 Hz power predicted the amount of stepping to be elicited by a stimulation train. Positive correlation (0.47>r>0.22) was found for seven out of eight sites showing positive priming effects. Correlation was absent for three other sites that showed non-significant priming effects and were mixed for four sites that showed negative effects. Sites with positive priming patterns, compared to sites with inconsistent or negative priming patterns, had similar trends in post-stimulation 3-6 Hz power, smaller increases in 6-8 Hz power during the control train and lower 1-3 Hz power during the periods immediately before the control stimulation. For six of 15 sites, regardless of the priming pattern, 1-3 Hz power was inversely related to subsequent stepping, and in three cases provided an independent predictor of stepping. Stimulation at two sites produced discrete episodes of post-stimulation stepping. In one of these cases, a 0.5-Hz increase in peak frequency of hippocampal activity preceded stepping. The results show that the association between hippocampal 3-6 Hz activity and the excitability of locomotor initiation is sufficiently specific to allow prediction of the magnitude of stepping by the prior levels of 3-6 Hz power. However, the occurrence of negative priming effects during prominent 3-6 Hz activity indicates that other factors determine the actual stepping and they can suppress the correlation between theta activity and subsequent locomotion. PMID- 10869841 TI - Benzodiazepines protect hippocampal neurons from degeneration after transient cerebral ischemia: an ultrastructural study. AB - The ability of full and partial benzodiazepine receptor agonists to prevent DNA fragmentation and neuronal death after transient cerebral ischemia was investigated in the Mongolian gerbil. Diazepam (10mg/kg, i.p.) or the partial agonist imidazenil (3mg/kg, i.p.) was administered 30 and 90min after transient forebrain ischemia produced by occlusion of the carotid arteries for 5min. Treatment with diazepam completely protected CA1b hippocampal pyramidal neurons in 94% of the animals and partially protected pyramidal neurons in 6% of the animals, as assessed with a standard Nissl stain three and four days after ischemia. DNA fragmentation was examined by the terminal dUTP nick-end labeling (TUNEL) reaction. Prior to cell death, there were no TUNEL-positive neurons in area CA1b. By three days after ischemia, when neuronal degeneration was nearly complete, 14 out of 16 gerbils exhibited a positive TUNEL reaction throughout area CA1b stratum pyramidale. In 13 out of 14 gerbils treated with diazepam, no TUNEL-positive neurons were observed in this region. Imidazenil was less effective than diazepam with respect to both neuroprotection and prevention of DNA fragmentation. Three days after ischemia, six out of eight gerbils treated with imidazenil showed partial to complete neuroprotection. Imidazenil completely prevented DNA fragmentation in only one of the animals; varying degrees of TUNEL reaction persisted in the remainder. To determine whether the neurons protected by diazepam had a normal ultrastructure, gerbils were killed two to 30 days after ischemia and the hippocampal neurons in area CA1b were examined by electron microscopy. Within the first 48h after ischemia, early cytoplasmic changes of varying degrees (e.g., vacuolation, rough endoplasmic reticulum stacking, swollen mitochondria) and electron-dense dendrites were observed in gerbils not treated with diazepam. Degeneration was nearly complete by three days after ischemia. In contrast, pyramidal neuron ultrastructure appeared normal in gerbils that exhibited complete area CA1b neuroprotection (defined at the light microscope level) by diazepam when studied two, seven or 30 days after ischemia. In gerbils with partial protection of area CA1b, most of the remaining neurons exhibited varying degrees of necrosis when studied 30 days after ischemia. No apoptotic bodies were observed. We conclude that: (i) diazepam can fully protect CA1 pyramidal cells from the toxic effects of transient cerebral ischemia; (ii) when diazepam affords only partial neuroprotection, the residual CA1 pyramidal cells exhibit ultrastructural abnormalities consistent with necrotic damage; and (iii) diazepam is a more efficacious neuroprotectant than the partial benzodiazepine receptor agonist, imidazenil. PMID- 10869842 TI - Comparative effects of neurotensin, neurotensin(8-13) and [D-Tyr(11)]neurotensin applied into the ventral tegmental area on extracellular dopamine in the rat prefrontal cortex and nucleus accumbens. AB - Ejections of 10(-5)-10(-3)M neurotensin into the ventral tegmental area increased dopamine efflux measured by electrochemical approaches in the prefrontal cortex of anaesthetized rats. In the same conditions, the effects evoked on dopamine efflux by 10(-5)M neurotensin(8-13) and [D-Tyr(11)]neurotensin were different from each other and depended on the explored area: the prefrontal cortex and the caudal and rostral nucleus accumbens. In the prefrontal cortex, neurotensin(8-13) was as potent as neurotensin, whereas [D-Tyr(11)]neurotensin was ineffective. In the caudal nucleus accumbens, when considering the initial intensity of the effect, neurotensin(8-13) and neurotensin appeared more potent than [D Tyr(11)]neurotensin. In contrast, in the rostral nucleus accumbens, neurotensin(8 13) was less potent than [D-Tyr(11)]neurotensin and neurotensin. These results support the differential involvement of two pharmacologically distinct neurotensin receptor entities on ventral tegmental area neurons in the modulation of mesolimbic and mesocortical dopaminergic activity. PMID- 10869843 TI - Changes in expression of N-methyl-D-aspartate receptor subunits in the rat neostriatum after a single dose of antisense oligonucleotide specific for N methyl-D-aspartate receptor 1 subunit. AB - In order to investigate the process of gene expression of N-methyl-D-aspartate glutamate receptor (NMDAR) subunits in the rat neostriatum and how this relates to motor behaviors, a single dose of antisense phosphodiester oligodeoxynucleotide specific for NMDAR1 was unilaterally applied in the neostriatum in a stereotaxic apparatus. After one day of antisense treatment, ipsilateral rotation behaviors that were induced by apomorphine were found in the treated animals. Reductions in the levels of expression of NMDAR1 and NMDAR2A messenger RNAs (NMDAR1: 20.6%; NMDAR2A: 19.7%) were found in the antisense treated striatal tissues by reverse transcriptase-polymerase chain reaction. There was no change in the levels of NMDAR2B, NMDAR2C and NMDAR2D messenger RNAs. After two days, western blotting experiments showed that there were decreases in the levels of expression of NMDAR1 (decreased 27.6%) and NMDAR2A (decreased 19.2%) proteins in the NMDAR1 antisense-treated striatal tissues. In addition, NMDAR1 immunoreactivity was found to decrease in intensity in the NMDAR1 antisense-treated neostriatum. At the cellular level, the intensity of NMDAR1 immunoreactivity in perikarya of presumed medium spiny neurons was found to decrease. These results indicate that a single dose of NMDAR1 antisense modifies the expression of NMDAR1 messenger RNA and protein in neurons in the neostriatum. The modification in the expression of NMDAR1 has differential effects in the expression of NMDAR2 subunits. Gene expression of the native NMDAR subunits is likely to be a dynamic process. The change in gene expression of the NMDAR subunits in the neostriatum may have a profound effect on the motor behaviors of rats. PMID- 10869844 TI - Striatal cells containing aromatic L-amino acid decarboxylase: an immunohistochemical comparison with other classes of striatal neurons. AB - In a previous study, we described a population of striatal cells in the rat brain containing aromatic L-amino acid decarboxylase, the enzyme involved in the conversion of L-DOPA into dopamine. We have also presented evidence that these cells produce dopamine in the presence of exogenous L-DOPA. In this paper, we further characterize these striatal aromatic L-amino acid decarboxylase containing cells in order to determine whether they form a subclass of one of the known categories of striatal neurons or if they represent a novel cell type. Using immunohistochemical methods, we compared the morphology and distribution of the aromatic L-amino acid decarboxylase-immunolabeled cells with those of other classes of striatal neurons. Our results show that both the morphology and distribution of aromatic L-amino acid decarboxylase-immunolabeled cells are very distinctive and do not resemble those of cells labeled for other striatal neuronal markers. Double-labeling procedures revealed that aromatic L-amino acid decarboxylase cells do not co-localize somatostatin or parvalbumin, and only a very small percentage of them co-localize calretinin. However, the population of aromatic L-amino acid decarboxylase cells label intensely for GABA.Overall, our results suggest that these aromatic L-amino acid decarboxylase-containing cells represent a class of striatal GABAergic neurons not described previously. PMID- 10869845 TI - Somatostatin inhibits GABAergic transmission in the sensory thalamus via presynaptic receptors. AB - The action of somatostatin on GABA-mediated transmission was investigated in cat and rat thalamocortical neurons of the dorsal lateral geniculate nucleus and ventrobasal thalamus in vitro. In the cat thalamus, somatostatin (10 microM) had no effect on the passive membrane properties of thalamocortical neurons and on the postsynaptic response elicited in these cells by bath or iontophoretic application of (+/-)baclofen (5-10 microM) or GABA, respectively. However, somatostatin (1-10 microM) decreased by a similar amount (45-55%) the amplitude of electrically evoked GABA(A) and GABA(B) inhibitory postsynaptic potentials in 71 and 50% of neurons in the lateral geniculate and ventrobasal nucleus, respectively. In addition, the neuropeptide abolished spontaneous bursts of GABA(A) inhibitory postsynaptic potentials in 85% of kitten lateral geniculate neurons, and decreased (40%) the amplitude of single spontaneous GABA(A) inhibitory postsynaptic potentials in 87% of neurons in the cat lateral geniculate nucleus. Similar results were obtained in the rat thalamus. Somatostatin (10 microM) had no effect on the passive membrane properties of thalamocortical neurons in this species, or on the outward current elicited by puff-application of (+/-)baclofen (5-10 microM). However, in 57 and 22% of neurons in the rat lateral geniculate and ventrobasal nuclei, respectively, somatostatin (1 microM) reduced the frequency, but not the amplitude, of miniature GABA(A) inhibitory postsynaptic currents by 31 and 37%, respectively. In addition, the neuropeptide (1 microM) decreased the amplitude of evoked GABA(A) inhibitory postsynaptic currents in 20 and 55% of rat ventrobasal neurons recorded in normal conditions and during enhanced excitability, respectively: this effect was stronger on bursts of inhibitory postsynaptic currents(100% decrease) than on single inhibitory postsynaptic currents (41% decrease). These results demonstrate that in the sensory thalamus somatostatin inhibits GABA(A)- and GABA(B)-mediated transmission via a presynaptic mechanism, and its action is more prominent on bursts of GABAergic synaptic currents/potentials. PMID- 10869846 TI - Immunohistochemical localization of the somatostatin sst(4) receptor in rat brain. AB - The biological actions of the neuromodulator somatostatin are mediated through a family of G-protein-coupled receptors, of which five members, sst(1-5), have been identified. Although the messenger RNA distribution of the sst(4) receptor has been reported, no information about the distribution of the receptor protein in the central nervous system is available. We have therefore raised a polyclonal peptide antibody against a rat carboxy-terminal sst(4) peptide. The selectivity of the affinity-purified antibody was demonstrated by western blotting of membrane proteins isolated from Chinese hamster ovary-K1 cells expressing the recombinant sst(4) receptor and from the rat hippocampus. This resulted in both cases in the identification of a single band of approximately 42,000 mol. wt. Furthermore, the sst(4) receptor antibody selectively labelled Chinese hamster ovary-K1 cells expressing the recombinant sst(4) receptor in immunocytochemistry. No cross-reactivity was observed with other recombinant somatostatin receptors. Immunohistochemistry on adult rat brain sections showed the sst(4) receptor to have a widespread distribution. This included labelling of cell bodies as well as processes in the cerebral cortex, hippocampus and several nuclei in the brainstem. All signals were absent following antibody preabsorption with the synthetic sst(4) peptide. This study provides the first detailed analysis of the distribution of sst(4) receptor protein in the rat brain. PMID- 10869847 TI - Differential regulation of fos family genes in the ventrolateral and dorsomedial subdivisions of the rat suprachiasmatic nucleus. AB - Extensive studies have established that light regulates c-fos gene expression in the suprachiasmatic nucleus, the site of an endogenous circadian clock, but relatively little is known about the expression of genes structurally related to c-fos, including fra-1, fra-2 and fosB. We analysed the photic and temporal regulation of these genes at the messenger RNA and immunoreactive protein levels in rat suprachiasmatic nucleus, and we found different expression patterns after photic stimulation and depending on location in the ventrolateral or dorsomedial subdivisions. In the ventrolateral suprachiasmatic nucleus, c-fos, fra-2 and fosB expression was stimulated after a subjective-night (but not subjective-day) light pulse. Expression of the fra-2 gene was prolonged following photic stimulation, with elevated messenger RNA and protein levels that appeared unchanged for at least a few hours beyond the c-fos peak. Unlike c-fos and fra-2, the fosB gene appeared to be expressed constitutively in the ventrolateral suprachiasmatic nucleus throughout the circadian cycle; immunohistochemical analysis suggested that delta FosB was the protein product accounting for this constitutive expression, while FosB was induced by the subjective-night light pulse. In the dorsomedial suprachiasmatic nucleus, c-fos and fra-2 expression exhibited an endogenous circadian rhythm, with higher levels during the early subjective day, although the relative abundance was much lower than that measured after light pulses in the ventrolateral suprachiasmatic nucleus. Double-label immunohistochemistry suggested that some of the dorsomedial cells responsible for the circadian expression of c-Fos also synthesized arginine vasopressin. No evidence of suprachiasmatic nucleus fra-1 expression was found. In summary, fos family genes exhibit differences in their specific expression patterns in the suprachiasmatic nucleus, including their photic and circadian regulation in separate cell populations in the ventrolateral and dorsomedial subdivisions. The data, in combination with our previous results [Takeuchi J. et al. (1993) Neuron 11, 825-836], suggest that activator protein-1 binding sites on ventrolateral suprachiasmatic nucleus target genes are constitutively occupied by DeltaFosB/JunD complexes, and that c-Fos, Fra-2, FosB and JunB compete for binding after photic stimulation. The differential regulation of fos family genes in the ventrolateral and dorsomedial suprachiasmatic nucleus suggests that their circadian function(s) and downstream target(s) are likely to be cell specific. PMID- 10869848 TI - Changes in sleep-wakefulness after 6-hydroxydopamine lesion of the preoptic area. AB - This study was undertaken to assess the role of catecholamine fibers, terminating in the preoptic area, in regulating sleep-wakefulness in rats. Sleep-wakefulness was assessed on the basis of 24h electroencephalogram, electromyogram and electro oculogram recordings before and after destruction of catecholaminergic terminals at the medial preoptic area by bilateral intracerebral injection of 6 hydroxydopamine (8 microg in 0.2 microl). There was a mild reduction in sleep and increase in wakefulness after the lesion. The increase in active wakefulness observed after eight days of lesion persisted even on the 12th day. In spite of the reduction in sleep, the day-night sleep ratio was not affected by 6 hydroxydopamine lesion of the preoptic area. The results indicate that the noradrenergic fibers at the preoptic area have a hypnogenic role. PMID- 10869849 TI - Beta-endorphin-, adrenocorticotrophic hormone- and neuropeptide y-containing projection fibers from the arcuate hypothalamic nucleus make synaptic contacts on to nucleus preopticus medianus neurons projecting to the paraventricular hypothalamic nucleus in the rat. AB - The nucleus preopticus medianus is known to be situated in a key site in pathways regulating the paraventricular hypothalamic nucleus. To investigate the innervation pattern to nucleus preopticus medianus neurons by afferent fibers containing beta-endorphin, adrenocorticotrophic hormone and neuropeptide Y, a retrograde tracing method was combined with immunohistochemistry for these peptides in the rat. In the first experiment with injection of a retrograde tracer in the nucleus preopticus medianus, retrogradely labeled neurons were found in many regions throughout the brain. Among these, the arcuate hypothalamic nucleus contained a number of retrogradely labeled neurons showing immunoreactivity to the neuropeptides examined. About 20%, 20% and 40% of retrogradely labeled arcuate hypothalamic nucleus neurons showed beta-endorphin, adrenocorticotrophic hormone and neuropeptide Y immunoreactivity, respectively. About 18% and 57% of retrogradely labeled neurons in the nucleus tractus solitarius and ventrolateral medulla, respectively, were immunoreactive to neuropeptide Y. There were many more neuropeptide Y-immunoreactive projections to the nucleus preopticus medianus from the arcuate hypothalamic nucleus than those from the medulla. None of the retrogradely labeled neurons in the medulla showed immunoreactivity to beta-endorphin or adrenocorticotrophic hormone. In the second experiment with injection of a retrograde tracer in the paraventricular hypothalamic nucleus, electron microscopic observation revealed that retrogradely labeled neurons in the nucleus preopticus medianus were in synaptic contact with beta-endorphin-, adrenocorticotrophic hormone- and neuropeptide Y-immunoreactive axon terminals. The present finding indicates that nucleus preopticus medianus neurons projecting to the paraventricular hypothalamic nucleus are innervated by beta-endorphin-, adrenocorticotrophic hormone- and neuropeptide Y-containing arcuate hypothalamic nucleus neurons in addition to being innervated by neuropeptide Y-containing catecholaminergic medullary neurons which have been reported in our previous study. PMID- 10869850 TI - Development of an antibody against a 40,000 mol. wt brain injury-derived neurotrophic peptide-binding protein and identification of a 40,000 mol. wt brain injury-derived neurotrophic peptide-binding protein in hippocampal neurons. AB - Brain injury-derived neurotrophic peptide is a 13-amino acid peptide derived from a 15,000 mol. wt neurotrophic factor released from sites of mechanical injury in neonatal rat brain. This peptide promotes survival of septal cholinergic neurons and mesencephalic dopaminergic neurons, and protects hippocampal neurons from glutamate-induced neurotoxicity. In this study, we have developed a monoclonal antibody against a brain injury-derived neurotrophic peptide-binding protein by immunizing mice with septal synaptosomes from five-week-old rat brain. Monoclonal antibodies were screened for inhibition of the binding of a 125I-labeled analogue of brain injury-derived neurotrophic peptide to rat brain synaptosomes. The monoclonal antibody 6A22 suppressed the biological activity of brain injury derived neurotrophic peptide and abolished the protective effect of the neurotrophic peptide against glutamate-induced neurotoxicity. This monoclonal antibody recognized a 40,000 mol. wt brain injury-derived neurotrophic peptide binding protein, which was also identified by cross-linking experiments. Immunohistochemical studies showed that the 6A22 antibody bound to the cell surfaces of a subpopulation (about 60%) of hippocampal neurons in culture. These results are consistent with the possibility that the 40,000 mol. wt protein belongs to brain injury-derived neurotrophic peptide receptors. PMID- 10869851 TI - In vivo study of motoneuron death induced by nerve injury in mice deficient in the caspase 1/ interleukin-1 beta-converting enzyme. AB - The apoptotic cell death program is orchestrated by members of the caspase family. Among these caspases, several in vitro and in vivo reports indicate that the interleukin-1 beta-converting enzyme (or caspase 1) may be involved in neurodegenerative processes. In view of these findings, and in order to characterize the role of the interleukin-1beta-converting enzyme in mediating or modulating cell death processes in vivo, we have investigated the effects of its deletion on motoneuron survival after a facial nerve transection in newborn and adult interleukin-1 beta-converting enzyme knock-out mice. During the postnatal period of development, when facial motoneurons are highly vulnerable to axotomy, we did not observe any significant effect of the interleukin-1 beta-converting enzyme-deletion on the percentage of cell death in the lesioned nuclei. In addition, the spontaneous cell death characteristic of the postnatal period was not altered in knock-out mice. In contrast, in adult knock-out mice, a significant reduction (16%) in the number of surviving facial motoneurons was observed six weeks after axotomy. We therefore conclude that the interleukin-1 beta-converting enzyme does not appear to be critical for cell death during the postnatal period but may favor motoneuron survival during adulthood. Given the key role of caspase 3 in neuronal apoptosis during embryonic development of the central nervous system, we also investigated the role of this caspase in cell death following axotomy. Combined immunofluorescence revealed that, at least during the postnatal period, axotomized motoneurons that have apoptotic nuclear morphologies were immunopositive for the active form of caspase 3. Double-stained cells could be also observed on the unlesioned side. These results strongly suggest that caspase 3 may be involved in both the postnatal spontaneous- and axotomy-induced facial motoneuron death processes. Similar results were obtained in interleukin-1 beta-converting enzyme-deficient and wild-type mice, indicating that the interleukin-1 beta-converting enzyme may not be required for caspase 3 activation. PMID- 10869852 TI - Murine models of inflammatory, neuropathic and cancer pain each generates a unique set of neurochemical changes in the spinal cord and sensory neurons. AB - The aim of this investigation was to determine whether murine models of inflammatory, neuropathic and cancer pain are each characterized by a unique set of neurochemical changes in the spinal cord and sensory neurons. All models were generated in C3H/HeJ mice and hyperalgesia and allodynia behaviorally characterized. A variety of neurochemical markers that have been implicated in the generation and maintenance of chronic pain were then examined in spinal cord and primary afferent neurons.Three days after injection of complete Freund's adjuvant into the hindpaw (a model of persistent inflammatory pain) increases in substance P, calcitonin gene-related peptide, protein kinase C gamma, and substance P receptor were observed in the spinal cord. Following sciatic nerve transection or L5 spinal nerve ligation (a model of persistent neuropathic pain) significant decreases in substance P and calcitonin gene-related peptide and increases in galanin and neuropeptide Y were observed in both primary afferent neurons and the spinal cord. In contrast, in a model of cancer pain induced by injection of osteolytic sarcoma cells into the femur, there were no detectable changes in any of these markers in either primary afferent neurons or the spinal cord. However, in this cancer-pain model, changes including massive astrocyte hypertrophy without neuronal loss, increase in the neuronal expression of c-Fos, and increase in the number of dynorphin-immunoreactive neurons were observed in the spinal cord, ipsilateral to the limb with cancer. These results indicate that a unique set of neurochemical changes occur with inflammatory, neuropathic and cancer pain in C3H/HeJ mice and further suggest that cancer induces a unique persistent pain state. Determining whether these neurochemical changes are involved in the generation and maintenance of each type of persistent pain may provide insight into the mechanisms that underlie each of these pain states. PMID- 10869853 TI - Alpha(1)-adrenoceptor-mediated excitation of substantia nigra pars reticulata neurons. AB - The effect of noradrenaline was studied in principal neurons of the substantia nigra pars reticulata in rat brain slices using patch clamp recordings. Perfusion of noradrenaline or the alpha(1)-adrenoceptor agonist phenylephrine increased the spontaneous firing activity of reticulata cells. The alpha(1)-adrenoceptor antagonist prazosin counteracted the effects of noradrenaline. In contrast, the beta-adrenoceptor agonist isoproterenol did not affect the activity of reticulata cells and the beta-adrenoceptor antagonist pindolol did not prevent noradrenaline's effect. In whole-cell recordings, at -60 mV holding potential, noradrenaline caused a tetrodotoxin-resistant inward current with a time-course similar to the increase in firing activity. Analysis of the reversal potential of this current did not give homogeneous results. The net noradrenaline current could be associated with a conductance decrease or increase, or in some cases it did not reverse over a range from -120 to -30 mV. It is suggested that noradrenaline increases the excitability of substantia nigra reticulata cells through alpha(1)-adrenoceptors. Both a reduction and an increase in membrane conductance may mediate this effect. The increase in the tonic firing of principal reticulata cells caused by noradrenaline may have significant consequences in regulating the final output of the basal ganglia and consequently in motor-related behaviours. PMID- 10869854 TI - The F-actin cytoskeleton modulates slow secretory components rather than readily releasable vesicle pools in bovine chromaffin cells. AB - Adrenal chromaffin cells were used to test the role of the peripheral cytoskeleton of F-actin in controlling different vesicle pools. Phorbol 12 myristate 13-acetate and calyculin A, two substances affecting phosphorylation dephosphorylation cycles, produced different degrees of F-actin reorganization, inducing the partial and the almost total disassembly of this structure, respectively, as visualized using rhodamine-phalloidin staining. Consequently, electron microscopy studies revealed the higher efficiency of calyculin-A over phorbol 12-myristate 13-acetate in promoting vesicle access to the plasmalemma boundary. Surprisingly, only the phorbol ester enhanced fast kinetics and the population of rapidly releasable vesicle pools as studied by single-cell amperometry, whereas both agents, as well as the F-actin severing compound, Latrunculin A, promoted an increase in the population of vesicles recruited in response to prolonged or repetitive stimulations. Taken together, our data support the notion that the F-actin peripheral barrier controls primary granule recruitment from reserve vesicle pools, whereas the phorbol ester effect on the rapidly releasable pools might be related to the alteration of late secretory stage through protein kinase C-dependent phosphorylation of an unidentified target. PMID- 10869855 TI - Ovine follicular fluid inhibits aromatase activity. AB - "Within follicle" regulations may be important for the fine tuning of gonadotrophin action in ovarian follicles. While numerous growth factors, steroids or proteins which are present in follicular fluid have been shown to have the ability of positively or negatively affecting follicle function, the net effect of follicular fluid of the dominant follicle on its function is unclear.A bioassay measuring aromatase activity of follicular walls was used (1) to check whether follicular fluid from dominant follicles can alter aromatase activity (2), to check how follicle size, atresia and specific gonadotrophins alter the effects of follicular fluid (3), to identify the nature (steroid or protein) of the active compound(s), and (4) to check whether the inhibition is specific of aromatase. Dominant follicular fluid had the ability to reduce aromatase activity. This effect was dose dependent and was obvious whether or not a protease inhibitor was added to the incubation medium. There was no difference in the magnitude of the inhibitory effect of follicular fluid when FSH (2 ng/ml) or no FSH was added to the incubation medium. LH, however, could potentialise the inhibitory effects of follicular fluid. Dominant follicular fluid was more potent to inhibit aromatase than follicular fluid from atretic follicles. Medium conditioned by granulosa cells, but not by theca cells could inhibit aromatase activity when added to the incubation medium. Charcoal treatment of dominant follicular fluid did not remove its inhibitory potential. Fractionation of dominant follicular fluid by a desalting column demonstrated that the inhibition was related to a compound(s) > 10 kDa. Finally, the effect of dominant follicular fluid on aromatase appears specific of this enzyme as follicular fluid does not affect androgen output by thecal shells or progesterone output by luteal cells. Further research is required to check whether the activity observed in dominant follicular fluid is related to compounds known to affect aromatase activity (inhibin, mullerian inhibiting substance, heat shock protein 90, superoxyde dismutase) or to another peptide/protein. PMID- 10869857 TI - cDNA cloning of canine common alpha gene and its co-expression with canine thyrotropin beta gene in baculovirus expression system. AB - The common alpha gene of the canine glycoprotein hormones was cloned, sequenced and co-expressed with the canine thyrotropin beta (TSH beta) gene in the baculovirus expression system, and a bioactive recombinant canine TSH was purified. The canine common alpha gene was cloned from the total RNA extracted from the canine pituitary gland by the reverse transcription polymerase chain reaction (RT-PCR) using primers that were designed based on the consensus sequences from other species. The resulting 476 bp PCR product is consisted of the full coding sequence for the 96 amino acid mature alpha subunit, and a sequence encoding a 24 amino acid signal peptide. Homology analysis with other species revealed that the canine common alpha subunit potentially contains five disulfide bonds and two oligosaccharide chains N-linked to Asn residues located at positions 56 and 82. For expression in the baculovirus expression system, the common alpha gene was cloned downstream of the p10 promoter of the pAcUW51 transfer vector, and the previously cloned canine TSH beta gene was inserted under the polyhedrin promoter of the same vector. The recombinant virus containing both alpha and beta genes was generated and propagated before being used to transfect the Sf9 insect cells for expression. The medium from the Sf9 cultures, presumably containing canine TSH alpha and beta in native heterodimer confirmation, exhibited TSH bioactivity as indicated in the cAMP stimulation assay in FRTL-5 cells. The expressed recombinant protein was purified from the culture medium with an affinity column that was coupled with IgG purified from the polyclonal antibodies generated against the partially purified native canine TSH. PMID- 10869856 TI - Canine thyrotropin beta-subunit gene: cloning and expression in Escherichia coli, generation of monoclonal antibodies, and transient expression in the Chinese hamster ovary cells. AB - The gene encoding the mature beta subunit of canine thyroid stimulating hormone (cTSH beta) was cloned, sequenced and expressed in Escherichia coli and in Chinese hamster ovary (CHO) cells, and monoclonal antibodies against the recombinant cTSH beta purified from E. coli were generated. The gene fragment that encodes mature TSH beta was cloned from the canine genomic DNA by direct polymerase chain reaction (PCR) using primers that were designed based on the consensus sequences from other species. The resulting 891 basepairs (bp) of genomic DNA consisted of two coding exons of the canine TSH beta gene and an intron of 450 bp. The two exons, which encode the mature cTSH beta subunit, was joined together by an overlap PCR and was expressed in E. coli as 6xHis-tagged protein. The purified recombinant cTSH beta with a molecular weight of about 15 kDa was recognized by the polyclonal antibodies prepared against the native canine TSH in Western blot. Monoclonal antibodies were raised against the purified cTSH beta and subsequently characterized. For transient expression in CHO cells that are permanently transfected with the bovine common alpha gene, a 60-oligonucleotide signal peptide coding sequence was added to the 5' end of the cTSH beta gene before it was cloned into the mammalian expression vector pRSV and used to transfect CHO cells. The medium from these transfected cells, presumably containing the bovine alpha and canine TSH beta in heterodimeric confirmation, exhibited TSH bioactivity as indicated by the stimulation of cAMP production in the cultured FRTL-5 thyrocytes. PMID- 10869858 TI - Evidence for a seasonal variation in the ability of exogenous melatonin to suppress prolactin secretion in the mare. AB - In seasonally breeding species photoperiodic information is thought to be conveyed to the reproductive and prolactin axis via changes in circulating concentrations of melatonin. For some species, a constant melatonin stimulus is perceived as a short day, whereas in others no photoperiodic information is provided. In the mare, a preliminary study demonstrated that constant administration of melatonin did not modify prolactin secretion, suggesting that this treatment regimen failed to provide photoperiodic information. To further investigate this proposal and to investigate an alternative explanation, namely a seasonal variation in response to melatonin, 4 experiments were performed. In experiments 1-3, the effects of constant administration of melatonin on prolactin secretion were investigated. In each study the time of treatment initiation varied beginning before the summer solstice, (May 9; Exp. 1), at the autumnal equinox (Sept. 21; Exp. 2) or the winter solstice (Dec. 21; Exp. 3). In Experiment 4, melatonin was administered as a timed daily injection (5 PM) for 6 months, beginning at the summer solstice (June 21). Constantly elevated physiological concentrations of melatonin (expts. 1-3) and an extended nighttime elevation of melatonin (exp. 4) suppressed prolactin concentrations only during the spring and early summer months (April-August). At other times during the year prolactin concentrations were similar to untreated mares. In the presence of a continuous melatonin implant the circannual rhythm of prolactin secretion was not disturbed. The results suggest that the prolactin axis of the mare is sensitive to an inhibitory melatonin signal during a restricted period of time and that at other times is refractory to this signal. PMID- 10869859 TI - Metabolic responses to mid-pregnancy shearing that are associated with a selective increase in the birth weight of twin lambs. AB - Previous studies have shown that shearing pregnant ewes at mid- or late-pregnancy is associated with an increase in lamb birth weight. The present study was designed to investigate metabolic responses that may underlie this response. Single- and twin-bearing ewes were either unshorn or shorn at mid-pregnancy (Day 69 of pregnancy; P69), and insulin, glucose and epinephrine challenges were conducted on P109-111 and P132-134. Shearing increased the birth weight of twin lambs by over 1 kg (P < 0.001) without having any effect on singleton birth weight. This response was associated with a 10-20% reduction in the insulin response to a glucose challenge (P < 0.05) without a change in glucose clearance following either glucose or insulin challenges. The lipolytic response to epinephrine challenge increased as pregnancy progressed, but was not associated with the increased birth weight of twin lambs born to shorn ewes. By late pregnancy, a 25% reduction in maternal IGF-I concentration and a two- to threefold increase in maternal IGFBP-1 concentration (P < 0. 05) associated with shearing were observed. The increase in lamb birth weight associated with mid pregnancy shearing may have been associated with an increase in the non-insulin dependent uptake of glucose by the placental-fetal unit. PMID- 10869862 TI - Gamma-hydroxybutyric acid efficacy, potential abuse, and dependence in the treatment of alcohol addiction. AB - The main objective in alcoholism therapy is to achieve and maintain abstinence and to prevent relapse. Pharmacotherapy may be necessary in treating persons who are not helped by group or psychosocial support alone. Among the substances experimented with in the past few years, gamma-hydroxybutyric acid has been effective in preventing alcohol withdrawal syndrome and in inducing a reduction in craving and an increase in the abstinence rate in treated alcoholics, in view of the alcohol-mimicking effects of the drug on the central nervous system. However, a possible development of craving for the drug and the risk of abuse and physical dependence have been reported in subjects who used gamma-hydroxybutyric acid for different reasons, including alcoholism therapy. The present review updates the existing differences in drug abuse behavior, side effects, and poisoning in the use of gamma-hydroxybutyric acid in a treatment alcoholism program and in self nonclinical illicit use. PMID- 10869863 TI - Safety and tolerability of gamma-hydroxybutyric acid in the treatment of alcohol dependent patients. AB - Gamma-hydroxybutyric acid (GHB) has been in clinical use in Italy since 1991 for treatment of alcohol dependence. Results of phase III and phase IV studies have shown that the drug is effective and well tolerated in the treatment of alcohol withdrawal syndrome and in reducing alcohol consumption and alcohol craving. Pharmacosurveillance indicates that abuse of gamma-hydroxybutyric acid is a limited phenomenon in clinical settings when the drug is dispensed under strict medical surveillance and entrusted to a referring familiar member of the patient. PMID- 10869864 TI - Design and structure-activity relationship analysis of ligands of gamma hydroxybutyric acid receptors. AB - With the use of [3H]gamma-hydroxybutyric acid, binding experiments allowed the screening of new compounds as ligands of gamma-hydroxybutyric acid receptors. Starting from the acid-alcohol gamma-hydroxybutyric acid structure, structure activity relation analysis and lead optimization highlighted gamma-hydroxybutyric acid derivatives with significantly increased affinities, when compared with the affinity of gamma-hydroxybutyric acid. Further pharmacological studies with the use of gamma-hydroxybutyric acid derivatives allowed the characterization of the first competitive antagonist acting at gamma-hydroxybutyric acid receptors (NCS 382). PMID- 10869865 TI - Characterization of the discriminative stimulus effects of gamma-hydroxybutyric acid as a means for unraveling the neurochemical basis of gamma-hydroxybutyric acid actions and its similarities to those of ethanol. AB - The present paper reviews the drug discrimination studies, both from the literature and from this laboratory, conducted to investigate the pharmacological profile of the discriminative stimulus effects of gamma-hydroxybutyric acid. Collectively, the results of these studies suggest that: (1) the discriminative stimulus effects of gamma-hydroxybutyric acid are composed of different cues, each one being the effect of gamma-hydroxybutyric acid on a specific receptor system; (2) the proportion of each component cue varies as the training dose of gamma-hydroxybutyric acid is increased; (3) the gamma-aminobutyric acid B mediated cue is a major ingredient of the mixed stimulus of gamma-hydroxybutyric acid, but it is more prominent at high training doses than at low training doses of gamma-hydroxybutyric acid; and (4) positive modulation of the gamma aminobutyric acid A receptor is a relevant part of the discriminative stimulus effects of low gamma-hydroxybutyric acid doses. Finally, data indicating symmetrical generalization between the discriminative stimulus effects of a specific range of doses of gamma-hydroxybutyric acid and those of ethanol are discussed in regard to their further support of the hypothesis that gamma hydroxybutyric acid may exert its antialcohol effects through a substitution mechanism. PMID- 10869866 TI - Gamma-hydroxybutyric acid: an evaluation of its rewarding properties in rats and mice. AB - Gamma-hydroxybutyric acid, an endogenous compound present in mammalian brain and supposed to be a neurotransmitter or neuromodulator, has been shown to affect several aspects of dependence from some drugs of abuse. It has been successfully used in clinical practice to alleviate both alcohol and opiate withdrawal symptoms. The aim of this study was to investigate whether gamma-hydroxybutyric acid possesses rewarding properties by means of conditioned place preference and intravenous self-administration paradigms. In the present study, gamma hydroxybutyric acid induced conditioned place preference in rats, was intravenously self-administered by drug-naive mice, and altered cocaine intravenous self-administration in rats. Although to date the physiological role of this compound still remains unclear, there is no doubt that gamma hydroxybutyric acid, in addition to its proved effect on alcohol and opiate dependence, possesses reinforcing properties of its own and may interfere with the neurochemical events in the rewarding effects produced by psychostimulant drugs. Our investigation points out the abuse liability of this drug, suggesting the use of particular precaution in handling gamma-hydroxybutyric acid as a clinically useful drug. PMID- 10869867 TI - Gamma-hydroxybutyric acid in the treatment of alcohol and heroin dependence. AB - We briefly review two double-blind, placebo-controlled surveys conducted in this laboratory with the aim of evaluating the efficacy of gamma-hydroxybutyric acid in the treatment of alcohol withdrawal syndrome as well as alcohol craving and consumption in alcoholics. In the first study, acute administration of 50 mg/kg gamma-hydroxybutyric acid, a nonhypnotic dose in alcoholic patients, resulted in a rapid and significant reduction of the severity score of alcohol withdrawal signs and symptoms that lasted as long as 7 hours. In the second study, treatment with 50 mg/kg/day gamma-hydroxybutyric acid for 3 consecutive months (1) reduced the number of daily drinks by approximately 50%, (2) increased the days of abstinence approximately threefold, and (3) reduced the alcohol craving score by up to 60%. These results feature gamma-hydroxybutyric acid as an effective agent for the treatment of alcohol dependence. Data on the effect of gamma hydroxybutyric acid on opiate withdrawal syndrome also are reviewed. Administration of 25 mg/kg induced a marked reduction of opiate withdrawal score in both heroin- and methadone-dependent subjects. Finally, we report the cases of adverse reactions to and abuse of gamma-hydroxybutyric acid revealed in a retrospective analysis of patients recruited in this laboratory over a 10-year period. PMID- 10869868 TI - Abuse and therapeutic potential of gamma-hydroxybutyric acid. AB - Gamma-hydroxbutyric acid is a compound found in mammalian brain that is structurally related to the neurotransmitters gamma-aminobutyric acid and glutamic acid. Gamma-hydroxybutyric acid effects dopaminergic systems in the brain and may be a neurotransmitter. Gamma-hydroxybutyric acid was first reported as a drug of abuse in 1990 and continues to be abused by bodybuilders, participants of "rave" dance parties, and polydrug abusers. Physical dependence can develop after prolonged, high-dose use, and overdoses have been widely reported. Its use in sexual assaults as a "date rape" drug and availability on the internet have recently emerged. Gamma-hydroxybutyric acid has established efficacy as an anesthetic agent, and preliminary evidence supports its utility in the treatment of alcohol dependence, opiate dependence, and narcolepsy. PMID- 10869869 TI - Mechanism of the antialcohol effect of gamma-hydroxybutyric acid. AB - Treatment with gamma-hydroxybutyric acid has been reported to effectively decrease alcohol craving and consumption as well as alcohol withdrawal symptoms in alcoholics. We describe the results of animal studies demonstrating the ability of gamma-hydroxybutyric acid to reduce (1) the severity of ethanol withdrawal signs in rats rendered physically dependent on ethanol and (2) voluntary ethanol intake in selectively bred Sardinian alcohol-preferring rats. Furthermore, we review experimental data suggesting that gamma-hydroxybutyric acid and ethanol have several pharmacological effects in common. Relevant similarities are: (1) stimulation of firing rate of dopaminergic neurons and dopamine release in specific rat brain areas; (2) development of cross-tolerance to the motor-impairing effects after repeated administration in rats; 3) abuse potential, as indicated by self-administration of pharmacologically relevant doses of gamma-hydroxybutyric acid in rats and mice; (4) induction of anxiolytic effects in rats; and (5) induction of similar discriminative stimulus effects, as evidenced by symmetrical generalization in a drug discrimination study in rats. These lines of evidence are discussed in relation to gamma-hydroxybutyric acid exerting its antialcohol effects by a substitution mechanism. PMID- 10869870 TI - Gamma-hydroxybutyric acid as a signaling molecule in brain. AB - Gamma-hydroxybutyric acid was synthesized 35 years ago to obtain a GABAergic substance that penetrates the brain freely. Since then, gamma-hydroxybutyric acid has been used in human beings for its sedative and anesthetic properties when administered at high doses, and most of the studies on gamma-hydroxybutyric acid have focused on its pharmacological effects. However, gamma-hydroxybutyric acid is also an endogenous substance, which is synthesized and released in the brain by specific neuronal pathways, implicated in the control of the GABAergic, dopaminergic, and opioid systems. This control is mediated by specific gamma hydroxybutyric acid receptors with a unique distribution in brain and a specific ontogenesis and pharmacology. Stimulation of these receptors induces specific cellular responses. Taken together, these results suggest that gamma hydroxybutyric acid possesses most of the properties required of a neurotransmitter/neuromodulator in the brain. PMID- 10869871 TI - Gamma-hydroxybutyric acid and alcohol-related syndromes. AB - We report on the effectiveness and safety of gamma-hydroxybutyric acid in the therapy of overt alcohol withdrawal syndromes, their prevention, and the prevention of relapses in formerly detoxified alcoholics. We studied 321 patients (236 men, 85 women), divided into two open-study groups for the treatment and prevention of alcohol withdrawal syndromes and one double-blind study group to evaluate the effects of gamma-hydroxybutyric acid versus placebo on alcoholic craving and relapses in detoxified patients. Gamma-hydroxybutyric acid treatment promptly reduced withdrawal symptoms in all patients and prevented alcohol withdrawal syndromes in 55% of cases. The attenuation of craving in detoxified patients was significantly greater in the gamma-hydroxybutyric acid-treated group in comparison with the placebo-treated group. The therapeutic use of gamma hydroxybutyric acid was not accompanied by serious side effects. Gamma hydroxybutyric acid diversion was poorly represented: gamma-hydroxybutyric acid induced abuse was reported in 4 (1.1%) of 345 treated patients, and only 9 cases of gamma-hydroxybutyric acid acute poisoning were reported in the years 1992 1995. Our results suggest that gamma-hydroxybutyric acid, with a favorable risk/benefit ratio, is a clinically useful drug in the treatment of alcohol dependence. PMID- 10869872 TI - Gamma-hydroxybutyric acid and growth hormone secretion studies in rats and dogs. AB - Gamma-hydroxybutyric acid, a gamma-aminobutyric acid metabolite, and baclofen, a gamma-aminobutyric acid B agonist, are endowed with a small growth hormone releasing activity in human beings. In this study, we have investigated the reciprocal interactions of gamma-hydroxybutyric acid and the gamma-aminobutyric acid B system by evaluating the growth hormone-releasing activity of the two compounds and their respective antagonists in in vivo and in vitro experiments performed in rats and dogs. In in vivo experiments, neither gamma-hydroxybutyric acid (25, 100, 150, and 300 mg/kg, SC) nor baclofen (0.25, 1, 2, 4, and 8 mg/kg, SC) significantly modified growth hormone secretion in 9-day-old rat pups. Similarly, no growth hormone and prolactin release was observed in adult anesthetized rats after administration of gamma-hydroxybutyric acid (100 mg/kg, IP) or baclofen (10 mg/kg IP). Equally ineffective on the somatotropic response was the administration of gamma-hydroxybutyric acid (200 mg/kg, IP) alone or associated with its specific receptor antagonist NCS-382 (150 mg/kg, IP) given to adult anesthetized rats. In addition, a toxicological dose of gamma hydroxybutyric acid (1500 mg/kg, IP) did not alter baseline growth hormone levels in adult conscious rats. gamma-Hydroxybutyric acid (50 mg/kg, IP) given for 10 days to adult conscious rats did not alter the growth hormone response to the same gamma-hydroxybutyric acid dose given acutely. In conscious dogs, gamma hydroxybutyric acid (20 and 50 mg/kg, IV) and baclofen (0.15, 0.30 mg/kg, IV) also were ineffective in stimulating growth hormone secretion. In this species, growth hormone response to hexarelin (31.25 microg/kg, IV), a potent growth hormone-releasing peptide, was not modified by coadministration of gamma hydroxybutyric acid (50 mg/kg, IV). In in vitro experiments, increasing doses of gamma-hydroxybutyric acid (10(-7), 10(-5), and 10(-3) M) did not alter growth hormone concentrations in media of rat pituitary cell cultures. In contrast, growth hormone-releasing hormone (10(-7) M) induced a significant growth hormone release into the media. In conclusion (1) gamma-hydroxybutyric acid is not an effective growth hormone secretagogue; (2) the reciprocal functional interactions between gamma-hydroxybutyric acid and the gamma-aminobutyric acid B system could not be investigated, due to the ineffectiveness of gamma-hydroxybutyric acid and baclofen to stimulate growth hormone release; and (3) short-term administration of gamma-hydroxybutyric acid does not induce adverse effects amenable to activation of the somatotropic function. PMID- 10869873 TI - Oncology staff recognition of depressive symptoms on videotaped interviews of depressed cancer patients: implications for designing a training program. AB - We examined oncologists' and nurses' ability to recognize depressive symptoms in two cancer patients who were interviewed on videotape. The study was conducted in a rural community, hospital-based outreach network. Staff were given a one-hour in-service on the use of the Mini International Neuropsychiatric Interview (MINI) a brief diagnostic interview-to provide a differential diagnosis (no psychiatric diagnosis, major depressive disorder, or adjustment disorder with depressed mood). Next, the staff viewed a videotape of an investigator (S.P.) utilizing the MINI to interview two depressed breast cancer patients. Staff subsequently rated depressive symptoms on the MINI and made a diagnosis. Findings indicated a high concordance among staff regarding symptom ratings on a straightforward example of major depressive disorder. Concordance on diagnosis, severity level, and specific symptoms declined slightly on a more difficult case involving primarily cognitive symptoms and a diagnosis of adjustment disorder. Following brief didactic training on depressive disorders, oncologists and nurses were able to identify depressive symptoms in cancer patients on videotape. Learning to use a semistructured interview can increase oncologists' awareness of depressive symptoms and may be a good training model. PMID- 10869874 TI - Pain and pain treatment in AIDS patients: a longitudinal study. AB - To determine the prevalence, incidence, and characteristics of pain connected with AIDS, 95 AIDS patients were enrolled in a prospective longitudinal study and interviewed every six months during a 2-year period or until death. The overall incidence of pain was 88%, and 69% of the patients suffered from constant pain interfering with daily living to a degree described as moderate or severe. The most common pain localizations were: extremities (32%), head (24%), upper gastrointestinal tract (23%) and lower gastrointestinal tract (22%). Pain conditions were connected to various opportunistic infections, Kaposi's sarcoma, or lymphoma. Pain in the extremities was predominantly of neuropathic origin (21%). The number of pain localizations increased significantly as death approached (0.8 +/- 1. 0 vs. 1.4 +/- 0.8, p = 0.03). The survival rate for patients without pain at entry was significantly higher than the survival rate of patients in pain, probably related to differences in the duration of AIDS at the time of inclusion. Sustained-release morphine preparations were prescribed in 29% of the patients. Of 39 patients (41%) who died in the department, 7 patients were prescribed continuous intravenous morphine infusion for pain treatment in the terminal phase and 20 patients received short-acting opioids. According to the Pain Management Index (PMI), the patients were insufficiently treated at the beginning of the study. Although the PMI improved significantly during the observation period, the patients felt that pain was not taken seriously by the physicians. However, the patients were convinced that treatment was optimal and, therefore, only 9% of the patients were dissatisfied. Patients were reluctant to take analgesics, primarily because of fear of addiction. PMID- 10869875 TI - A pharmacokinetic and tolerability evaluation of two continuous subcutaneous infusion systems compared to an oral controlled-release morphine. AB - The pharmacokinetic profiles, safety, and tolerability of continuous subcutaneous infusion with a novel drug deliver system (the MEDIPAD system) was compared to a standard infusion pump (the CADD-Micro) and to controlled-release tablets (MS Contin) for the administration of morphine sulfate. This was a single-center, open-label, three-treatment study conducted in 24 male and female healthy volunteers. The mean age was 40.6 yr (SD = +/- 12.27). A three treatment design was chosen to compare differences between modes of administration within each subject to minimize the impact of intersubject variability: Treatment A was a continuous 48-hr subcutaneous infusion of morphine sulfate (165.6 mg at a rate of 3. 45 mg/hr) with the MEDIPAD system attached to the chest, Treatment B was a series of four oral doses of morphine sulfate (120 mg each) at 12-hr intervals, and Treatment C was a continuous 48-hr subcutaneous infusion of morphine sulfate (163.2 mg at a rate of 3.40 mg/hr) with the CADD-Micro device attached to the chest. Subjects began treatment after eligibility was established and informed consent was obtained. The primary pharmacokinetic parameters (AUC, C(max)) for the two devices were similar; 90% confidence intervals showed that the MEDIPAD system was bioequivalent to the CADD-Micro in terms of both rate and extent of morphine absorption. The mean morphine plasma concentration versus time plot suggested that plasma concentrations rise more rapidly with the MEDIPAD device than with the CADD-Micro or oral administrations. The MEDIPAD system showed mild application and injection site reactions; there were no site reactions for the CADD-Micro or oral doses. As expected nausea, somnolence, and abdominal pain occurred more frequently in the oral treatment than the two infusion devices. These data suggest that the MEDIPAD system, which is currently undergoing clinical evaluation, is an acceptable alternative to the traditional oral treatment of morphine sulfate for delivery of analgesics as it allows rapid absorption of morphine; is small, easy to use, and disposable; and achieves plasma levels that are essentially equal to other standard infusion pumps. PMID- 10869876 TI - The frequency and correlates of dyspnea in patients with advanced cancer. AB - Dyspnea is a devastating symptom in patients with advanced cancer. Unfortunately, very limited research has been done on the frequency and correlates of dyspnea in this particular patient population. The purpose of this prospective study was to assess the frequency of moderate to severe dyspnea and the correlates of dyspnea in a population of ambulatory terminally ill cancer patients. One hundred thirty five consecutive patients attending a multidisciplinary pain clinic were tested for respiratory function (vital capacity, peak flow, maximal inspiratory pressure, and oxygen saturation). All patients gave their rating of dyspnea, anxiety, and fatigue/tiredness using visual analogue scales (VAS). Lung involvement by the tumor (primary or metastatic) was determined from the patient's chart. Moderate dyspnea occurred in 74/135 (55%) patients with terminal cancer. Lung involvement (r = 0.285, P = 0. 0009), anxiety (r = 0.306, P = 0.0003), fatigue/tiredness (r = 0.211, P = 0.0146), and vital capacity (r = 0.189, P = 0.0444) were significantly correlated with the intensity of dyspnea. Multivariate analysis demonstrated that lung involvement (P = 0.0016) and anxiety (P = 0.0027) were independently correlated with the intensity of dyspnea. In the subgroup of patients with moderate to severe dyspnea, multivariate analysis found anxiety (P = 0.0318) and maximal inspiratory pressure (P = 0.0187) to be independent correlates of the intensity of dyspnea. Dyspnea is a frequent symptom in patients with advanced cancer. The presence of cancer in the lungs, anxiety, and maximal inspiratory pressure are correlates of the intensity of dyspnea in this patient population. Possible treatments addressing low maximal inspiratory pressure and anxiety are needed, as well as further research in finding new correlates of dyspnea in advanced cancer patients. PMID- 10869877 TI - The measurement of symptoms in children with cancer. AB - The purpose of this study was to determine symptom prevalence, characteristics, and distress in children with cancer. The Memorial Symptom Assessment Scale (MSAS) 10-18, a 30-item patient-rated instrument adapted from a previously validated adult version, provided multidimensional information about the symptoms experienced by children with cancer. This instrument was administered to 160 children with cancer aged 10-18 (45 inpatients, 115 outpatients). To confirm the instrument's reliability and validity, additional data about symptoms were collected from both the parents and the medical charts, and retesting was performed on a subgroup of inpatients. Patients could easily complete the scale in a mean of 11 minutes. The analyses supported the reliability and validity of the MSAS 10-18 subscale scores as measures of physical, psychological, and global symptom distress, respectively. Symptom prevalence ranged from 49.7% for lack of energy to 6.3% for problems with urination. The mean (+/- SD) number of symptoms per inpatient was 12.7 +/- 4.9 (range, 4-26), significantly more than the mean 6.5 +/- 5.7 (range, 0-28) symptoms per outpatient. Patients who had recently received chemotherapy had significantly more symptoms than patients who had not received chemotherapy for more than 4 months (11.6 +/- 6.0 vs. 5. 2 +/- 5.1), and those patients with solid tumors had significantly more symptoms than patients with either leukemia, lymphoma, or central nervous system malignancies (9.9 +/- 7.0 vs. 6.8 +/- 5.5 vs. 6.8 +/- 5.0 vs. 8.0 +/- 6.1). The most common symptoms (prevalence > 35%) were lack of energy, pain, drowsiness, nausea, cough, lack of appetite, and psychological symptoms (feeling sad, feeling nervous, worrying, feeling irritable). Of the symptoms with prevalence rates > 35%, those that caused high distress in more than one-third of patients were feeling sad, pain, nausea, lack of appetite, and feeling irritable. Subscale scores demonstrated large variability in symptom distress and could identify subgroups with high distress. The prevalence, characteristics, and distress associated with physical and psychological symptoms could be quantified in older children with cancer. The data confirm a high prevalence of symptoms overall and the existence of subgroups with high distress associated with one or multiple symptoms. Symptom distress is relatively higher among inpatients, children with solid tumors, and children who are undergoing antineoplastic treatment. Systematic symptom assessment may be useful in future epidemiological studies of symptoms and in clinical chemotherapeutic trials. Symptom epidemiology may also provide a focus for future clinical trials related to symptom management in children with cancer. PMID- 10869878 TI - Management of dyspnea in severe chronic obstructive pulmonary disease. AB - Progression of chronic obstructive pulmonary disease (COPD) is frequently associated with increasing dyspnea; indeed, patients with severe COPD constitute the largest group of patients with chronic respiratory insufficiency. The sensation of dyspnea in these patients is mostly related to increased work of breathing, a consequence of an increased resistive load, of hyperinflation, and of the deleterious effect of intrinsic positive end-expiratory pressure (PEEP(i)). Once optimal medical treatment has been provided, pharmacological treatments of dyspnea exist (beta2-agonists, methylxanthines, opiates) but seldom suffice. Nonpharmacological complementary treatments must be envisioned. Patients with severe hyperinflation should be screened as possible candidates for lung reduction surgery. Pulmonary rehabilitation-including chest therapy, patient education, exercise training-has been established as effective on quality of life (QoL) and dyspnea. Noninvasive positive pressure devices may be effective for symptomatic treatment of severe dyspnea: continuous positive airway pressure (CPAP) counteracts the deleterious effect of PEEP(i) in patients with severe hyperinflation; intermittent positive pressure breathing (IPPB) may decrease dyspnea and discomfort during nebulized therapy; finally noninvasive positive pressure ventilation (NIPPV) has been shown to be effective on the sensation of dyspnea and QoL in COPD with severe hypercapnia. PMID- 10869879 TI - Prolonged oral morphine therapy for severe angina pectoris. AB - Patients with intractable angina pectoris despite optimal drug therapy, who are not candidates for revascularization procedures, pose a very difficult problem. We evaluated the role of chronic opioid therapy in four such patients. The patients (mean age 79.5 years) were treated by low doses (mean 40 mg/day) of controlled-release oral morphine (CRM) for 1 to 5 years. The treatment was followed by a marked decline in the rate of admissions and hospitalization periods. The number of admissions decreased from a mean of 6 during the year prior to CRM therapy to 1.5 the following year. The duration of hospitalization for angina pectoris during these periods decreased from a mean of 42 +/- 35 days to 6 +/- 10 days (p < 0.05). Side effects were negligible and consisted mainly of lactulose-responsive constipation. We suggest that prolonged oral morphine therapy may be administered with good efficacy and no significant side effects in selected patients with intractable angina pectoris. PMID- 10869880 TI - Lamotrigine in the treatment of chronic refractory neuropathic pain. AB - Many patients suffer from chronic, intractable neuropathic pain. Despite similar diagnoses and presumed pathophysiologies, symptoms and response to treatment can differ. Monotherapy is only occasionally successful. In this prospective survey, 20 patients with chronic, neuropathic pain not responding to interventional therapy received lamotrigine, sometimes as monotherapy and sometimes combined with oral morphine. The latter occurred in patients who lost pain relief from morphine after time. Ten patients did not respond to the drug; 4 were temporary responders and 6 patients obtained sustained pain relief. It is interesting that 5 patients regained opioid responsiveness and that the drug combination produced excellent pain relief for more than 5 months. We hypothesize an additive effect between morphine and lamotrigine. PMID- 10869881 TI - The genes for major psychosis: aberrant sequence or regulation? AB - A number of recent clinical and molecular observations in major psychosis indicate that epigenetic factors may be operational in the origin of major mental illness. This article further develops the idea that epigenetic factors may play an etiopathogenic role in schizophrenia and bipolar affective disorder. The putative role of epigenetic factors is shown by the epigenetic interpretation of genetic association studies of the genes for serotonin 2A (HTR2A) and the dopamine D3 (DRD3) receptors in schizophrenia. The idea of epigenetic polymorphism of genetic alleles is introduced, and it is argued that epigenetic variation may explain a number of controversial and unclear findings in allelic and genotypic association studies of HTR2A and DRD3. In linkage analyses of multiplex families with bipolar affective disorder (BPAD), different loci on chromosome 18 indicated co-segregation of alleles of one parental sex with the disease phenotype, and this finding implies that the epigenetic mechanism of genomic imprinting may be involved. Evidence for genomic imprinting provides the background for epigenetic cloning of BPAD risk factors by searching for differentially modified genes on chromosome 18. Finally, epigenetic studies could be relevant to the better understanding of the molecular action of antipsychotic medications. In addition to this, if epimutations are detected in major psychosis, epigenetic treatment directed at correction of epigenetic status of a specific brain gene may eventually be developed. PMID- 10869882 TI - Effects of antidepressants on weight and on the plasma levels of leptin, TNF alpha and soluble TNF receptors: A longitudinal study in patients treated with amitriptyline or paroxetine. AB - Leptin, tumor necrosis factor-alpha (TNF-alpha), and soluble TNF receptors are involved in weight regulation. Antipsychotic agents, such as clozapine, induce weight gain and increase circulating levels of these cytokines. To assess whether obesity-inducing antidepressants have a similar effect, we measured plasma cytokine levels in depressive inpatients during the first six weeks of treatment with tricyclic agents (amitriptyline or nortriptyline, n = 12), with paroxetine (n = 10), or without medication (n = 14). There was an increase in the body mass index at week 6 of treatment with the tricyclics, which was preceded by a significant increase in soluble TNF receptor p75 plasma levels. Circulating levels of leptin were not affected. Paroxetine and drug-free treatment did not affect any of these parameters. We conclude that weight gain induced by psychotropic agents may occur without increased circulating levels of leptin. However, activation of the TNF-alpha system might be an early and sensitive marker of ensuing weight gain. PMID- 10869883 TI - Somatodendritic 5-HT(1A) autoreceptors mediate the anti-aggressive actions of 5 HT(1A) receptor agonists in rats: an ethopharmacological study with S-15535, alnespirone, and WAY-100635. AB - To elucidate the relative contribution of somatodendritic 5-HT(1A) autoreceptors and postsynaptic 5-HT(1A) receptors in the specific anti-aggressive properties of 5-HT(1A) receptor agonists, the influence of the novel benzodioxopiperazine compound S-15535, which behaves in vivo as a competitive antagonist at postsynaptic 5-HT(1A) receptors and as an agonist at 5-HT(1A) autoreceptors, upon offensive and defensive aggression was investigated in wild-type rats using a resident-intruder paradigm. S-15535 exerted a potent dose-dependent decrease in offensive, but not defensive, aggressive behavior (inhibitory dose (ID)(50) = 1.11 mg/kg). This anti-aggressive profile was roughly similar to that of the potent pre- and postsynaptic 5-HT(1A) full agonist alnespirone (ID(50) = 1. 24). The drug's profound anti-aggressive actions were not accompanied by sedative side effects or signs of the "5-HT(1A) receptor-mediated behavioral syndrome," which are characteristically induced by prototypical 5-HT(1A) receptor agonists like 8 OH-DPAT and buspirone. The selective pre- and postsynaptic 5-HT(1A) antagonist WAY-100635, which was inactive given alone, abolished the anti-aggressive effects of S-15535 and alnespirone, thereby confirming the involvement of 5-HT(1A) receptors. Furthermore, combined administration of S-15535 and alnespirone elicited an additive anti-aggressive effect, providing further support for somatodendritic 5-HT(1A) receptor involvement. Finally, the postsynaptic 5-HT(1A) antagonistic properties of S-15535 were confirmed by showing blockade of the alnespirone-induced hypothermia, a postsynaptic 5-HT(1A) mediated response in the rat. These data provide extensive evidence that the anti-aggressive effects of 5 HT(1A) receptor agonists are expressed via their action on somatodendritic 5 HT(1A) autoreceptors, thereby most likely attenuating intruder-activated serotonergic neurotransmission. PMID- 10869884 TI - Chronic oral administration of CG-3703, a thyrotropin releasing hormone analog, increases wake and decreases cataplexy in canine narcolepsy. AB - The effects on cataplexy and daytime sleep of acute and chronic oral administration of CG-3703, a potent TRH analog were assessed in canine narcolepsy. CG-3703 was found to be orally active and to reduce cataplexy (0.25 to 16 mg/kg) and sleep (8 and 16 mg/kg) in a dose-dependent manner. Two-week oral administration of CG-3703 (16 mg/kg) significantly reduced cataplexy and daytime sleep. The anticataplectic effects of CG-3703 were not associated with changes in general behavior, heart rate, blood pressure, rectal temperature, blood chemistry and thyroid function. Although drug tolerance for the effects on cataplexy and sleep were observed during the second week of chronic drug administration, therapeutic efficacy on cataplexy was improved with individual dose adjustment (final dose range: 16 to 28 mg/kg, p.o.). These results suggest that TRH analogs could be a promising new form of treatment for human narcolepsy. PMID- 10869885 TI - Further evidence that behavioral tests and neuropeptide mRNA and tissue level alterations can differentiate between typical and atypical antipsychotic drugs. AB - This study was designed to compare some behavioral and biochemical effects of chronic treatment with a range of antipsychotic drugs. Gene expression of enkephalin, chromogranin A, chromogranin B, and secretogranin II and their respective peptide products were studied with in situ hybridization and radioimmunoassays after daily oral administration of haloperidol, clozapine, risperidone, or zotepine for 21 days. In behavioral tests, significant catalepsy was induced by haloperidol only. All four antipsychotic drugs increased hind paw retraction time but only haloperidol also increased forelimb retraction time. In the caudate putamen, haloperidol increased both enkephalin mRNA expression and enkephalin tissue levels. Neither of these parameters was altered by the other three drugs. In the prefrontal cortex, antipsychotic drugs generated a distinct pattern of gene expression in two regards. First, the dopamine D(2) receptor antagonist, haloperidol, did not significantly alter synaptic protein levels or their encoding mRNAs. Secondly, there was a differential change in tissue levels and mRNA expression since secretogranin II was not affected by any tested antipsychotic drug. This study shows that different types of antipsychotic drug induce distinct behavioural effects as well as differential changes in the biosynthesis of synaptic proteins and their encoding mRNAs. The data reinforce the notion that haloperidol can be classed as a typical antipsychotic drug whilst clozapine, zotepine, and risperidone reflect their atypical classification. PMID- 10869886 TI - In vivo olanzapine occupancy of muscarinic acetylcholine receptors in patients with schizophrenia. AB - Olanzapine is an atypical antipsychotic with potent antimuscarinic properties in vitro (K(i) = 2-25 nM). We studied in vivo muscarinic receptor occupancy by olanzapine at both low dose (5 mg/dy) and high dose (20 mg/dy) in several regions of cortex, striatum, thalamus and pons by analyzing [I-123]IQNB SPECT images of seven schizophrenia patients. Both low-dose and high-dose olanzapine studies revealed significantly lower [I-123]IQNB binding than that of drug-free schizophrenia patients (N = 12) in all regions except striatum. [I-123]IQNB binding was significantly lower at high-dose than low-dose in the same regions. Muscarinic occupancy by olanzapine ranged from 13% to 57% at 5 mg/dy and 26% to 79% at 20 mg/dy with an anatomical pattern indicating M(2) subtype selectivity. The [I-123]IQNB data indicate that olanzapine is a potent and subtype-selective muscarinic antagonist in vivo, perhaps explaining its low extrapyramidal side effect profile and low incidence of anticholinergic side effects. PMID- 10869887 TI - Modulation by estrogen-receptor directed drugs of 5-hydroxytryptamine-2A receptors in rat brain. AB - Hormonal specificity of modulation of brain 5-HT(2A) receptors was investigated by comparing activity of compounds with varying effects on estrogen response in breast, bone, and uterus. A two-week estradiol treatment stimulated the decreased uterine weight of ovariectomized rats to intact rat values whereas an increase of 29% with tamoxifen and 16% with raloxifene was observed compared to vehicle treated ovariectomized rats. In 18 assayed brain regions, ovariectomy decreased 5 HT(2A) receptor binding and mRNA levels in anterior cingulate and frontal cortices, striatum, and nucleus accumbens; estradiol restored this decrease to intact rat values. Dehydroepiandrosterone (DHEA) increased ovariectomized rats 5 HT(2A) receptor expression only in striatum and cortical amygdala. Tamoxifen increased 5-HT(2A) receptor density only in striatum. Raloxifene, an uterine estrogen receptor (ER) antagonist, increased, like estradiol, 5-HT(2A) receptor density and expression in cingulate and frontal cortices, striatum, and nucleus accumbens. Brain regional specificity of estradiol, DHEA, tamoxifen, and raloxifene on 5-HT(2A) receptors was observed which can be dissociated from peripheral activity. PMID- 10869888 TI - Effects of repeated nicotine administration and footshock stress on rat mesoprefrontal dopamine systems: Evidence for opioid mechanisms. AB - We have examined the effects of nicotine pre-treatment on mesoprefrontal dopamine (DA) function in the presence and absence of acute stress, and the involvement of endogenous opiate peptide systems (EOPS). Acute electrical footshock stress preferentially increases DA utilization in medial prefrontal cortex (mPFC) compared to nucleus accumbens (NAS) and striatal terminal fields, and this is correlated with profound locomotor immobility. Our recent studies have demonstrated that repeated, but not acute, nicotine pre-treatment significantly reduced mPFC DA utilization and footshock stress-induced immobility responses. There is increasing evidence that the biochemical and behavioral effects of nicotine are mediated by EOPS, and we hypothesized that the stress-reducing effects of repeated nicotine administration in these studies were mediated by EOPS. Accordingly, rats pre-treated subcutaneously with repeated nicotine were given a single dose of the opiate receptor antagonist naloxone (0.1-10.0 mg/kg, i.p.) or saline as a co-treatment with nicotine or saline 10 min prior to acute footshock stress. Naloxone had no effects on non-stressed or acute footshock stress-induced mPFC DA utilization, but dose-dependently antagonized repeated nicotine's attenuation of stress-induced mesoprefrontal DA utilization and immobility responses. Furthermore, naloxone dose-dependently blocked repeated nicotine's augmentation of accumbal DA utilization. These results suggest that EOPS may be involved in mediating repeated nicotine administration effects on mesoprefrontal dopaminergic and immobility responses to acute footshock stress. PMID- 10869889 TI - Effects of serotonin and serotonergic agonists and antagonists on the production of interferon-gamma and interleukin-10. AB - Serotonin (5-HT) is a neurotransmitter and an immune modulator. In vitro, antidepressants with a serotonergic mode of action have, at concentrations within the therapeutical range, negative immunoregulatory effects, i.e., they increase the production rate of interleukin-10 (IL-10), a negative immunoregulatory cytokine. We have hypothesized that part of these effects may be explained by the serotonergic activities of antidepressants on immunocytes. This study was carried out to examine the effects of 5-HT, p-chlorophenylalanine (PCPA), a 5-HT depleting agent, flesinoxan (a 5-HT1A agonist), m-chlorophenylpiperazine (mCPP; a 5-HT2A/2C agonist), and ritanserin (a 5-HT2A/2C antagonist) on the production rate of interferon-gamma (IFNgamma), a proinflammatory cytokine, and IL-10 by whole blood stimulated with polyclonal activators. The IFNgamma/IL-10 production ratio was computed, since this ratio reflects the pro- versus anti-inflammatory capacity of cultured whole blood. We found that: 1) 5-HT, 150 ng/mL, 1.5 microg/mL, and 15 microg/mL significantly decreased the IFNgamma/IL-10 ratio; 2) PCPA (5 microM) significantly suppressed the production of IFNgamma and IL-10; 3) flesinoxan (15 ng/mL; 1.5 microg/mL) had no significant effects on the production of the above cytokines; and 4) mCPP (2.7 microg/mL) and ritanserin (5.0 microg/mL) suppressed the IFNgamma/IL-10 ratio. It is concluded that intracellular 5-HT may be necessary for an optimal synthesis of IFNgamma and IL 10, and that extracellular 5-HT concentrations at or above serum values may suppress the production of the proinflammatory cytokine IFNgamma. The negative immunoregulatory effects of antidepressive drugs are probably not related to their serotonergic activities. PMID- 10869890 TI - The long-term effects of maternal deprivation depend on the genetic background. AB - The neurodevelopmental hypothesis of schizophrenia has led to a series of new animal models in which the long term consequences of early manipulations are investigated. We have recently shown that a single 24-hr period of maternal deprivation (at postnatal day (pnd) 9) increases apomorphine susceptibility and decreases prepulse inhibition in Wistar rats, viz. phenomena also seen in schizophrenic patients. In the present paper, we investigated whether the effects of maternal deprivation were dependent on a specific genetic background, by using different rat strains (Fischer 344 and Lewis) that differ in the Hypothalamus Pituitary-Adrenal axis and in dopaminergic sensitivity. The data show that in Wistar rats, basal startle amplitude was not affected by maternal deprivation, but prepulse inhibition was reduced, and apomorphine susceptibility enhanced. In Fischer 344 rats on the other hand, neither basal startle amplitude, nor prepulse inhibition were affected, but apomorphine susceptibility was reduced. In Lewis rats, maternal deprivation significantly reduced basal startle amplitude, but did not affect prepulse inhibition or apomorphine susceptibility. The differential response to maternal deprivation can best be explained by differences in baseline dopamine sensitivity between the rat strains. Since a reduced prepulse inhibition and an enhanced susceptibility to apomorphine is also seen in schizophrenic patients, the data indicate that maternally deprived Wistar rats may represent an interesting developmental model for (aspects of) schizophrenia. PMID- 10869892 TI - Computer assisted surgery-joined forces in medicine for the next millennium. PMID- 10869893 TI - Effects of medium-chain and long-chain triacylglycerols in pediatric surgical patients. AB - Medium-chain triacylglycerols (MCTs) have been shown to provide better nutritional support than long-chain triacylglycerols (LCTs). This study compares the efficacy of MCT combined with LCT with LCT alone in pediatric patients with surgical stress. Two groups of patients (n = 19 in each) received equivalent amounts of glucose (12 g. kg. d) and amino acids (2 g. kg. d), but one group received 10% Lipofundin MCT/LCT and the other received 10% LCT (1.5 g. kg. d) in a randomized study. Total parenteral nutrition (TPN) was given for 14 d. Blood and urine samples were collected before and after TPN administration for determination of various biochemical parameters. Indirect calorimetry was also performed to determine respiratory quotients and fuel utilization. After 14 d of TPN in the MCT/LCT group, there was a significantly higher blood lymphocyte percentage, a decreasing tendency of serum asparate aminotransferase and of total and direct bilirubin (P < 0.05). These changes were not observed in the LCT group. A significantly better nitrogen balance and a higher ketogenesis from day 3 were observed in the MCT/LCT group. The MCT/LCT group showed a more marked increased utilization of fat than the LCT group, whereas carbohydrate oxidation was less in the MCT/LCT group than in the LCT group (P < 0.05). In children after surgery, MCT/LCT is more protein sparing and induces a better immune response when compared with LCT-containing lipid emulsion. A TPN regimen containing MCT/LCT is likely to result in rapid oxidation of fats for energy without compromising the respiratory system. PMID- 10869894 TI - Diabetes mellitus and Helicobacter pylori infection. AB - Alterations of glucose metabolism in diabetes have been suggested as promoting Helicobacter pylori colonization. We performed a cross-sectional sero-prevalence study of diabetic patients (insulin-dependent, or type 1, and non-insulin dependent, or type 2, diabetes mellitus) with H. pylori and compared them with a control group. Consecutive diabetic outpatients aged 12 to 75 y and with disease duration of greater than 1 y were enrolled. Helicobacter pylori status was evaluated by using an enzyme-linked immunosorbent assay for anti-H. pylori immunoglobulin G. Demographic data were obtained from each individual, and socioeconomic class was assessed by occupation and education level. A total of 891 individuals participated (240 with type-2 diabetes, 145 with type-1 diabetes, and 506 control subjects). After controlling for age, there was no significant difference in the prevalence of H. pylori infection in any age group. In fact, the prevalence of H. pylori was numerically higher among children in the control group than among children with type-1 diabetes (25% versus 9%, respectively; P = 0.1). Previous associations of H. pylori and diabetes may have arisen from failure to consider socioeconomic status or age. Because childhood is the most common period for acquisition of H. pylori infection, the higher prevalence of infection among the normal children as opposed to those with type-1 diabetes confirms the lack of an association. PMID- 10869895 TI - Correlates of total plasma homocysteine: folic acid, copper, and cervical dysplasia. AB - We examined correlates of total plasma homocysteine (tHcy) in 294 subjects with cervical intraepithelial neoplasia and 170 control subjects. Associations of tHcy with risk factors for cervical intraepithelial neoplasia and 24-h intakes and biochemical indices of nutrients were examined. Plasma and red blood cell folate and plasma B(12) were strong inverse correlates of tHcy (r = -0.35, -0. 31, and 0.27, respectively). Plasma copper and severity of dysplasia were positively correlated with tHcy (r = 0.14 and 0.21, respectively). A stepwise regression model that included red blood cell folate, plasma copper, grade of dysplasia, ethnicity, intake of polyunsaturated fatty acids, plasma vitamin B(12), intake of fat, and oral contraceptive use explained 29% of the variation in tHcy. Two hundred thirty-five subjects with cervical intraepithelial neoplasia were randomized to receive folic acid (10 mg/d) or placebo for 6 mo. After 2, 4, and 6 mo, mean tHcy in the folate-supplemented group (7.2 +/- 1.8, 7.0 +/- 1.9, and 7.0 +/- 2.3 micromol/L, respectively) was significantly lower than baseline and the placebo group at 2, 4, and 6 mo (8.9 +/- 3.1, 8.4 +/- 3.0, and 8.9 +/- 3.1 micromol/L, respectively). Supplementation lowered tHcy even in subjects in the highest quintile of baseline folate. Folate, vitamin B(12), copper, and severity of dysplasia are associated with tHcy. Folate supplementation significantly lowers tHcy even in folate-replete subjects. PMID- 10869896 TI - Normal values of the bioelectrical impedance vector in childhood and puberty. AB - The purpose of this study was to determine the reference, bivariate, and tolerance intervals of the whole-body impedance vector in Italian children. This was a cross-sectional, multicenter study, and participants were chosen from the general school population. The impedance vector (standard, tetrapolar analysis at 50-kHz frequency) was measured in 3110 subjects, ages 2 to 15 y, and 2044 healthy children (1014 male and 1030 female) with weight and height within the 95th percentile were selected for the analysis (resistance-reactance graph method). The age-specific 95% confidence intervals of mean vectors and the 95%, 75%, and 50% tolerance intervals for individual vector measurements were plotted using resistance and reactance components standardized by the subject's height. Mean vectors from both sexes with separate 95% confidence ellipses were considered as representative of eight different age groups, from 2 to 13 y. There was a statistically significant sex effect on vector distribution from boys and girls in the age group of 14 to 15 y. The impedance vector distribution of children was also compared with healthy adult subjects (354 male and 372 female, age 15 to 85 y). There was a progressive, statistically significant vector shortening from age 2 to 15 y toward the adults' vector position. In conclusion, we established the trajectory followed by the mean impedance vector in children over ages 2 to 15 y and also obtained the reference, bivariate, and 95%, 75%, and 50% tolerance intervals of the impedance vector by age for healthy children, with which the vectors from children with altered body composition can be tested. PMID- 10869898 TI - Serum leptin levels in patients with hyperlipidemias. AB - Leptin is a protein hormone produced by adipocytes that reflects the body fat content. The aim of our study was to compare serum leptin levels in randomly selected untreated males and females with hypercholesterolemia and combined hyperlipidemia and in healthy control subjects matched for age and body mass index and to study the relations between leptin and serum lipids and lipoproteins. No statistically significant differences in serum leptin levels were found between the male control group (5.26 +/- 2.81 ng/mL(-1)) and the male group with hypercholesterolemia (8.16 +/- 3.85 ng/mL(-1)) or combined hyperlipidemia (7.51 +/- 4.83 ng/mL(-1)) and between the female control group (13.0 +/- 8.12 ng/mL(-1)) and the female group with hypercholesterolemia (15.36 +/- 8.89 ng/mL(-1)) or combined hyperlipidemia (18.63 +/- 10.15 ng/mL(-1)). Leptin concentration in male group with hypercholesterolemia did not differ significantly from the female control group; in the other male groups, leptin levels were significantly lower than those of the other female groups. Serum leptin levels in all studied groups except for the male group with hypercholesterolemia positively correlated with body mass index. Serum leptin levels correlated negatively with high-density lipoprotein cholesterol in the female group with hypercholesterolemia (r = -0.67, P < 0.01) and the male group with combined hyperlipidemia (r = -0.56, P < 0.01). A positive correlation between serum leptin and high-density lipoprotein cholesterol (r = 0.67, P < 0.01) and between leptin and lipoprotein (a) (r = 0.71, P < 005) was found in female group with combined hyperlipidemia. No other significant relationships between leptin and serum lipids or lipoproteins were found. We conclude that serum leptin levels in patients with hyperlipidemias do not significantly differ from those healthy control subjects matched by age and body mass index. PMID- 10869897 TI - Changes in micronutrient concentrations following anti-inflammatory treatment in patients with gastrointestinal cancer. AB - Circulating concentrations of vitamin antioxidants (retinol, alpha-tocopherol, lutein, lycopene, alpha- and beta-carotene) and trace elements (zinc, copper, iron and selenium) plus carrier proteins (albumin, transferrin, caeruloplasmin) in gastrointestinal cancer patients (n = 12) with an inflammatory response (as demonstrated by an elevated C-reactive protein concentration) were compared with a control group (n = 12). Further, the effect of moderating the inflammatory response, using the anti-inflammatory agent ibuprofen, on these measurements was examined in the cancer group. The control and cancer groups were similar in terms of age, sex, and body mass index. However, the cancer group had significantly higher C-reactive protein concentrations (P < 0.001). Concentrations of vitamin antioxidants and trace elements (and carrier proteins) were significantly lower (P < 0.001), except copper (ceruloplasmin) which was significantly higher (P < 0.05). After anti-inflammatory treatment, there were small but significant increases in lutein, lycopene, and beta-carotene (P < 0.05) and in iron and selenium (P < 0.05), whereas ceruloplasmin decreased (P < 0. 05). The micronutrient concentrations in the cancer patients remained different from those in the control subjects. These results support the concept that the magnitude of inflammation plays an important role in the regulation of circulating concentrations of vitamin antioxidants and trace elements in patients with gastrointestinal cancer. PMID- 10869900 TI - Monosaccharide-enriched diets cause hyperleptinemia without hypophagia. AB - To determine the effect of monosaccharide-enriched diets on plasma leptin and food consumption, body weight, food intake, and serum glucose, insulin, and leptin concentrations were measured in rats maintained on a 10-d course of 60% glucose or 60% fructose diet. The serum leptin concentration in rats fed a high glucose diet (7.60 +/- 0.6 ng/mL) or a high-fructose diet (5.12 +/- 0.8 ng/mL) was significantly increased compared with that in control rats (2.45 +/- 0.10 ng/mL; P < 0.001). To ascertain that the observed effect was related to hyperinsulinemia, a group of rats were infused with exogenous insulin or rendered insulin resistent with a high-fat diet. When hyperinsulinemia was induced with exogenous infusion, the serum leptin was increased (5.56 +/- 0.23 ng/mL; P < 0.001). High-fat feeding was associated with increased serum leptin (12.1 +/- 1.4 ng/mL) and insulin levels. The increased serum leptin concentration was not associated with decreased food intake. We conclude that monosaccharide-enriched diets, probably through hyperinsulinemia or relative or absolute insulin resistance, cause hyperleptinemia, which does not appear to change feeding behavior. PMID- 10869899 TI - Low serum folate but normal homocysteine levels in patients with atherosclerotic vascular disease and matched healthy controls. AB - Mild hyperhomocysteinemia has been considered a cardiovascular risk factor. However, recent prospective studies have not demonstrated that hyperhomocysteinemia or the underlying genetic defect on methylentetrahydrofolate reductase is associated with a higher risk of coronary or peripheral artery disease. We compared serum homocysteine, folate, and vitamin B(12) levels of patients with coronary and peripheral vascular disease with those of age- and sex matched healthy individuals. Subjects taking multivitamins, with diabetes mellitus, or serum creatinine levels over 1.5 mg/dL were excluded from the study. Homocysteine was measured by fluorimetric high-performance liquid chromatography. Serum folate and vitamin B(12) levels were measured by an ion-capture method. We studied 32 patients with peripheral vascular disease (10 female), aged 69.6 +/- 11 y, 24 age- and sex-matched control subjects, 52 patients with coronary artery disease (7 female), aged 59.5 +/- 10.4 y, and 42 age- and sex-matched control subjects. Serum homocysteine levels were 11.7 +/- 7.4 and 9.3 +/- 4.5 micromol/L in vascular patients and in the control counterparts, respectively (not significant). The levels for coronary patients and the control counterparts were 9.0 +/- 3.9 and 8.6 +/- 3.6 micromol/L, respectively (not significant). Folate levels were 4.48 +/- 2.42 and 7.14 +/- 4.04 ng/mL in vascular patients and control subjects, respectively (P < 0.02); the levels in coronary patients and control counterparts were 5.15 +/- 1.9 and 6.59 +/- 2.49 ng/mL, respectively (P < 0.01). No differences in vitamin B(12) or tocopherol levels were observed between patients and control subjects. There were no differences in homocysteine levels, but lower serum folate levels were observed when comparing patients with atherosclerotic vascular disease and healthy control subjects. PMID- 10869901 TI - Dietary nucleotides modulate antigen-specific type 1 and type 2 t-cell responses in young c57bl/6 mice. AB - Mice fed a nucleotide-free (NF) diet have impaired antibody (Ab) responses. The mechanisms responsible for this effect are not understood but may be related to specific changes in T-cell functions. The objective of this study was to examine the effects of dietary nucleotides on serum immunoglobulin-G (IgG) subclass Ab levels and T-cell cytokine production by cells from the lymph nodes draining the site of antigen challenge. C57BL/6 (B6) mice were fed an NF diet or the same diet supplemented with nucleotides (NS diet; 4.74 g nucleotides/kg). Keyhole limpet hemocyanin (KLH; 25 microg/dose), a T-dependent protein neoantigen, was given with incomplete Freund's adjuvant. We administered KLH at 3, 6, and 9 wk to determine primary and secondary responses. Anti-KLH IgG subclass Ab levels were measured 3 wk after the first KLH challenge and 2 wk after the last KLH challenge. T-cell responses in lymph nodes draining the site of KLH challenge were assessed 5 d after the primary and 14 d after the final KLH challenge. We measured mRNA expression and production of interferon-gamma and interleukin-5, type-1 and type-2 T-cell cytokines, respectively. Anti-KLH IgG2a and IgG2b Ab levels were higher in the NS diet group than in the NF diet group after the last KLH challenge. The NS diet group had higher interferon-gamma production and mRNA expression than did the NF diet group after the first KLH challenge. Because increased levels of interferon-gamma and IgG2a/IgG2b Ab reflect a shift toward type-1 responses to antigen stimuli, our results show that dietary nucleotides preferentially enhance type-1 responses to KLH given with incomplete Freund's adjuvant. PMID- 10869902 TI - Effects of protein-energy malnutrition and human immunodeficiency virus-1 infection on essential fatty acid metabolism in children. AB - This report summarizes data on the availability of essential fatty acids (EFAs) and their long-chain polyunsaturated fatty acid (LCPUFA) metabolites in protein energy malnutrition (PEM), in human immunodeficiency virus-1 (HIV-1) infection for which less information is available, and the combination of both PEM and HIV 1. The contribution of different EFAs and LCPUFAs to the fatty-acid composition of plasma and erythrocyte membrane lipids was found to be reduced in children with PEM in comparison with well-nourished children. In addition to limited dietary EFA supply, reduced bioconversion of EFAs to their respective LCPUFA metabolites and/or peroxidative degradation of LCPUFAs may contribute to the reduction of LCPUFA status in malnourished children. Restoration of normal energy, protein, and EFA intakes does not appear to readily correct abnormalities of plasma and erythrocyte membrane LCPUFA values. Enhanced dietary supply of LCPUFAs and/or improved supply of antioxidant vitamins may represent novel therapeutic modalities in severe PEM. With and without PEM, HIV infection was related to altered availability of various EFAs and LCPUFAs in HIV-seropositive children. The plasma total lipid fatty-acid profiles seen in well-nourished children with HIV infection were compatible with an HIV infection-related enhancement of the metabolic activity of the conversion of EFAs to their respective LCPUFA metabolites. However, the plasma phospholipid EFA and LCPUFA profiles seen in severely malnourished children with HIV infection more closely resembled those seen in children with PEM but without HIV infection than in those in children with HIV infection but no PEM. Metabolic studies using stable isotope labeled fatty acids may contribute to better understanding of the HIV-related changes in EFA metabolism and clearly are needed before therapeutic conclusions can be drawn. PMID- 10869903 TI - Beef allergy in children. AB - Beef allergy was poorly known before the '90s. Since then, a number of papers appeared elucidating the nature, epidemiology, and symptoms of beef allergy in children allergic to cow's milk and children suffering from atopic dermatitis. It is now clear that beef allergy is not an infrequent occurrence, with an incidence between 3. 28% and 6.52% among children with atopic dermatitis, its incidence may be as much as 0.3% in the general population. A diagnosis of beef allergy must be supported by skin prick tests, RASTs, and challenges. The specificity and sensitivity according to type of test and the type of extract, however, remains to be evaluated. Despite the fact that other allergens can be sensitizing, the major beef allergen is bovine serum albumin (BSA). Beef-sensitive children are also sensitized to ovine serum albumin, as well as to other serum albumins; therefore, the use of alternative meats in beef-allergic children must be carefully evaluated on an individual basis. Because industrial heat processing is more efficient than domestic cooking in reducing reactivity in beef-sensitive children, freeze-drying and homogenization may support the introduction of processed beef into the diet of beef-allergic children. PMID- 10869904 TI - Glutamine regulation of human immune cell function. PMID- 10869905 TI - Soy protein and obesity. PMID- 10869906 TI - Use of medium-chain triacylglycerols in parenteral nutrition of children. PMID- 10869907 TI - Some aspects of interpretation of the odds ratio. PMID- 10869908 TI - The great protein fiasco revisited. PMID- 10869909 TI - Is there a pharmacist in the house? PMID- 10869910 TI - The nutraceutical benefit, part iii: honey. PMID- 10869911 TI - Antioxidant supplementation in critical illness: what do we know? PMID- 10869913 TI - Pharmacodynamics and intubating conditions of cisatracurium in children during halothane and opioid anesthesia. AB - STUDY OBJECTIVES: To determine the pharmacodynamics and intubating conditions of cisatracurium 0.2 mg/kg in children aged 2 to 12 years. DESIGN: Open-label, randomized study. SETTING: Operating room of a university-affiliated hospital. PATIENTS: 42 ASA physical status I and II patients, 24 to 155 months of age. INTERVENTIONS: Patients were assigned to one of two groups: halothane anesthesia (G1) and opioid anesthesia (G2). Subsequently, each group was divided into two age subgroups: 24-59 months and 60-155 months. All patients were premedicated with midazolam intranasal 0.1 to 0.2 mg/kg. In G1, anesthesia was induced with halothane up to 3% and N(2)O/O(2) (60-70/30-40%). Halothane was reduced to 0.05). CONCLUSIONS: Cisatracurium 0.2 mg/kg offered acceptable intubating conditions at 90 seconds in 98% of pediatric patients, regardless of the anesthesia-based technique. Longer clinical duration in the halothane group in younger children may be due to age-related potentiation or to the small number of patients enrolled in the younger subgroup. PMID- 10869914 TI - Determinants of core temperature at the time of admission to intensive care following cardiac surgery. AB - OBJECTIVE: To determine the predictors of core temperature on arrival in the intensive care unit (ICU) after cardiac surgery. DESIGN: Prospective, randomized trial. SETTING: Tertiary care medical center, operating rooms (ORs), and ICU. PATIENTS: 72 patients presenting for coronary artery bypass surgery. INTERVENTIONS: Randomized assignment for ambient OR temperature (16-18 degrees C vs. 21-23 degrees C) and rewarming endpoint on cardiopulmonary bypass (CPB; nasopharyngeal and urinary bladder temperatures >/=36.5 degrees C and 34.0 degrees C, respectively, vs. nasopharyngeal and urinary bladder temperatures >/=37.5 degrees C and 36.0 degrees C, respectively) at the time of separation from bypass. MEASUREMENTS AND MAIN RESULTS: The best (and only significant) predictor of core temperature on arrival in the ICU was rewarming endpoint at the time of separation from CPB (p = 0.004). Patient weight, height, body habitus, and nitroprusside administration did not significantly predict core temperature. Ambient temperature affected only body temperature when the duration of time in the OR after separation from bypass was prolonged (>90 min). A weighted average body temperature was a better predictor of complete rewarming than was any single monitoring site. CONCLUSIONS: To reduce the incidence of hypothermia after cardiac surgery, the most important variable is rewarming endpoint achieved before separation from bypass. A warm ambient temperature (>21 degrees C) may be beneficial if the duration of time in the OR after bypass is prolonged (>90 min). PMID- 10869915 TI - Rapid induction of anesthesia with high concentrations of halothane or sevoflurane in children. AB - STUDY OBJECTIVE: To compare the characteristics of the rapid induction of anesthesia in pediatric patients with high concentrations of sevoflurane or halothane, and to determine the ability of anesthesiologists to correctly identify the anesthetic drug when administered in this fashion. DESIGN: Randomized, prospective, open-label study. SETTING: Academic university hospital. PATIENTS: 78 ASA physical status I and II healthy children scheduled for brief surgical procedures with general anesthesia and medicated with midazolam. INTERVENTIONS: Assessments were made by 5 pediatric anesthesiologists and 18 anesthesiology residents. Sevoflurane or halothane was randomly selected for anesthetic induction. The anesthetic circuit was primed with the drug (8% sevoflurane or 4% halothane) in 50% nitrous oxide and oxygen. The anesthesiologists were blinded as to the anesthetics being administered. After completion of anesthetic induction, the anesthesiologists were asked to identify the anesthetic and to assess the quality and speed of induction. MEASUREMENT AND MAIN RESULTS: The pediatric anesthesiologists correctly identified the anesthetic in 55 of 78 assessments (70.5%). This figure is statistically better than what could be achieved by random guessing (p < 0.001). The residents correctly identified the anesthetic in only 46 of 78 assessments (60.0%). Statistically, this figure is no better than what could be achieved by random guessing (p = 0.06). Speed of induction was subjectively felt to be faster with sevoflurane than halothane but there were no differences in actual induction time (sevoflurane group, 3.7 +/- 2.7 min; halothane group, 3.7 +/- 2.6 min). There were no differences in the quality of induction or the incidence of airway complications. The perceived incidence of tachycardia was significantly higher with sevoflurane than halothane(sevoflurane group, 74%; halothane group 20%). CONCLUSION: The induction of anesthesia with high concentrations of either halothane or sevoflurane can be safely accomplished. Pediatric anesthesiologists can differentiate between halothane and sevoflurane when either drug is given in high initial concentrations. The presence of tachycardia may have served as the primary clue in determining which drug was being used. PMID- 10869916 TI - Propofol dosage achieving spontaneous breathing during balanced regional anesthesia with the laryngeal mask airway. AB - STUDY OBJECTIVE: To assess an anesthetic technique achieving spontaneous breathing through the laryngeal mask airway (LMA) during combined epidural block and propofol anesthesia. DESIGN: Prospective, consecutive case series study. SETTING: Operating room at a general hospital. PATIENTS: 112 ASA physical status I and II adult surgery patients; 32 patients for lower extremity surgery are enrolled into study 1, and 30 patients for lower extremity surgery and 50 patients for lower abdominal gynecology surgery are enrolled into study 2. INTERVENTIONS: In study 1, patients were given 1.5 to 2.0 mg/kg followed by a 3 mg/kg/h of infusion of propofol, after epidural block, and they were fitted with the LMA. Thirty minutes after induction, the dose of propofol was increased to 4, 5, 6, and 7 mg/kg/h every 15 minutes. In study 2, the patients were given 1.5 to 2.0 mg/kg and 5 mg/kg/h of propofol and the LMA insertion, after epidural block. MEASUREMENTS AND MAIN RESULTS: PaO(2)/FIO(2), PaCO(2), tidal volume or respiratory rate, blood pressure, heart rate, and eye opening and motor response scales in conformity with Glasgow coma scale were recorded. Study 1 suggested an induction dose of 1.5 to 2.0 mg/kg and an infusion of 5 mg/kg/h as an appropriate dose to preserve spontaneous breathing with the LMA and to maintain reasonable depth of anesthesia. Study 2 showed that respiratory and circulatory conditions, depth of anesthesia, and other data related to anesthesia were clinically acceptable. CONCLUSIONS: The best infusion dose of propofol to achieve spontaneous breathing with the LMA seems to be 5 mg/kg/h, and the present balanced regional anesthesia with the LMA, using propofol infusion at 1.5 to 2.0 mg/kg and 5 mg/kg/h combined with epidural block, may be useful in clinical practice for lower extremity and lower abdominal gynecologic operations. PMID- 10869917 TI - Effect of xenon on autonomic cardiovascular control--comparison with isoflurane and nitrous oxide. AB - STUDY OBJECTIVES: To clarify the effect of xenon on the autonomic nervous system by comparing similar effects of isoflurane and nitrous oxide. DESIGN: Prospective, randomized study. SETTING: Operating room at a university hospital. PATIENTS: 39 ASA physical status I and II patients scheduled for general anesthesia. INTERVENTIONS: Patients were randomly allocated into one of three groups and received one of the following inhalational anesthetics: 56% of xenon (Group X), 0.94% of isoflurane (Group I), or 70% of nitrous oxide and 0.15% of isoflurane (Group N). Phenylephrine (pressor test) and nicardipine (depressor test) were given to assess baroreflex sensitivity. MEASUREMENTS AND MAIN RESULTS: Continuous blood pressure (BP) and electrocardiogram (ECG) were recorded before and during anesthesia to analyze heart rate (HR) variability and baroreflex sensitivity. Power spectrum of HR variability was calculated by fast Fourier transformation and power spectrum densities at low frequency (LF: 0.04-0.15Hz) and high frequency (HF: 0.15-0.40 Hz) were compared. Baroreflex sensitivity was calculated from the slope of regression for BP changes versus associated changes in R-R intervals. For HR variability, Group X showed lower power spectrum densities (ms(2).Hz(-1)) in LF and HF than did Group I (LF: 0.09 +/- 0.06 vs. 0.35 +/- 0.53; p < 0.05; HF: 0.40 +/- 0.34 vs. 0.98 +/- 0.68, p < 0.01). Group X had the lowest baroreflex sensitivity (ms.mmHg(-1)) via pressor test of the three study groups (Group X: 2.00 +/- 0.87, Group I: 3.53 +/- 2.14, Group N: 3.78 +/- 2. 17, p < 0.05). CONCLUSIONS: Xenon depressed both sympathetic and parasympathetic transmission more than isoflurane at 0.8 MAC. Xenon was also suggested to be relatively vagotonic. PMID- 10869918 TI - Moderate controlled hypotension with sodium nitroprusside does not improve surgical conditions or decrease blood loss in endoscopic sinus surgery. AB - STUDY OBJECTIVES To determine if moderate controlled hypotension can improve the dryness of the surgical field in endoscopic sinus surgery. STUDY DESIGN: Randomized, prospective study. SETTING: University-affiliated hospital. PATIENTS: 32 ASA physical status I and II adult patients undergoing endoscopic sinus surgery. INTERVENTIONS: All patients were premedicated orally with chlorazepate. Patients in Group H received 12.5 mg captopril orally prior to surgery. Anesthesia was provided using an intravenous (IV) technique supplemented with nitrous oxide (N(2)O); anesthesia was maintained with boluses of 2 mcg/kg fentanyl and a propofol infusion at rates between 3 and 9 mg/kg/h at the discretion of the anesthetist. In Group H, sodium nitroprusside was infused at a rate of 1 to 2.5 mcg/kg/min to maintain moderate controlled hypotension with mean blood pressure of 65 to 75 mm Hg. MEASUREMENTS AND MAIN RESULTS: Arterial blood pressure was assessed via the radial artery. Readings were recorded prior to intubation, immediately after intubation, at the start of surgery, then at 5, 15, 30, 45, and 60 minutes intraoperatively, and at the end of surgery. Intraoperative blood loss, dryness of the surgical field, adrenocorticotropic (ACTH) hormone, arginin-vasopressin (AVP), cortisol, and the preoperative and postoperative psychomotoric function were examined. At the start of surgery and thereafter, MAP increased in Group N but not in Group H. Throughout surgery, MAP was significantly lower in Group H than in Group N. Blood loss, dryness of the surgical field, ACTH, AVP, and cortisol levels, and psychomotoric function were not significantly different between the groups. CONCLUSION: Intravenous anesthesia supplemented with N(2) is as effective as moderate controlled hypotension when blood loss, visibility in the surgical field, ACTH, AVP, and cortisol are examined. PMID- 10869919 TI - Perioperative myocardial cell injury: the relationship between troponin T and cortisol. AB - STUDY OBJECTIVE: To investigate whether there is an association between Troponin T (TnT), reflecting myocardial cell injury, and cortisol, reflecting the degree of surgical trauma and associated stress, in light of our recent evaluation of TnT as a marker of perioperative myocardial cell injury. DESIGN: Prospective, cohort study. PATIENTS: 70 patients (67.4 +/- 8.7 yrs) with definite or at-risk coronary artery disease (CAD) undergoing elective noncardiac surgery (vascular n = 38, abdominal n = 21, orthopedic n = 8) with general (n = 63) or regional (n = 4) anesthesia with postoperative on-demand analgesia. MEASUREMENTS AND MAIN RESULTS: Morning blood samples for TnT (upper limit of normal: <0.2 ng/mL), CK-MB (reference range .9 by the Kruskal-Wallace test). CONCLUSIONS: The isolation of C. trachomatis from the cervix of sexually active adolescent females at a single point in time does not impact on the prevalence of significant cervical smear abnormalities. PMID- 10869966 TI - Congenital perineal lipoma presenting as "ambiguous genitalia": a case report. AB - A case is presented in which an unusual, phallic-shaped perineal lipoma raised the question of ambiguous genitalia following the delivery of an otherwise healthy female infant. The management of the case is described, and the critical features of the physical examination that contradicted that diagnosis are highlighted. PMID- 10869967 TI - The vulvar mass in the prepubertal child. AB - The differential diagnosis of the vulvar mass in the prepubertal patient is extensive and reported cases of a vulvar hamartoma in the literature are limited. The case presented in this work and review of the literature demonstrate that when considering the differential diagnosis of vulvar masses in the prepubertal female, hamartoma should be included. This review outlines the differential diagnosis of the vulvar mass and gives a comprehensive review of benign masses arising from embryonic remnants, those of mesenchymal origin as well as sarcoma botryoides-embryonal rhabdomyosarcoma and the embryonal sinus tumor. PMID- 10869968 TI - Recognizing teen dating violence. PMID- 10869969 TI - Ovarian cysts in premenarchal children. PMID- 10869970 TI - Longitudinal risk of std acquisition in adolescent girls using a generalized estimating equations model AB - Background: To date, few studies have used a longitudinal approach in examining risk factors for STD acquisition in teenage girls. In the present study, we examined risk of STD acquisition in adolescent girls over a three-year period using longitudinal analyses.Methods: One hundred and seventy-four girls between the ages of 12 and 16 (mean age = 14.5) participated in a longitudinal study of adolescent romantic relationships. The racial composition of the subject pool was 80% African-American and 20% Caucasian. These girls were followed for three years at six-month intervals. By the end of the study, 127 of the girls were sexually experienced, and thus, were included in the analyses regarding risk for STDs. The risk for history of STD was evaluated using a generalized estimating equations (GEE) model, which allows for the inclusion of data from subjects with missing visits. The following independent variables were entered into the model: age, race, screening IQ score, qualitative cognitive functioning, perceptions of STD prevalence among female friends, and whether a condom was used at last intercourse. Additional independent variables included number of lifetime partners (4 or more lifetime partners, 2 to 3 lifetime partners, only one partner) and age of sexual debut (less than 14 years, 14-16 years, and 17 years and older), five factors from the Family Environment Scale, and three types of parental monitoring (direct, direct when with peers, and indirect).Results: The results of the longitudinal (GEE) model indicated that having a history of an STD was significantly related to a younger age of sexual debut (p <.01), having more partners (p <.0001), being African-American (p =. 01), and having a lower screening IQ score (p <.01).Conclusions: These results suggest that risk of STD is associated with both modifiable and non-modifiable variables. Race and intellectual functioning independently contributed to STD risk, stressing the importance of utilizing prevention programs that are culturally and developmentally specific. For example, less intelligent girls may need to have information presented in a more concrete manner, with a focus on the present rather than the future. The targets of these prevention programs should continue to emphasize delaying initiation, as well as partner selection. Finally, the fact that variables for STD acquisition in this longitudinal model are comparable to previous cross-sectional studies of STD acquisition suggests that these are potent predictors of risk. PMID- 10869971 TI - Early medical abortion with methotrexate. Outcome and satisfaction among women aged 15-20 years AB - Background: We investigated the outcomes of women under age 21 who participated in a multi-center case series of early medical abortion using methotrexate and misoprostol.Methods: We enrolled 1973 women in a case series for medical abortion in the first seven weeks of pregnancy, using a standardized protocol and consent for methotrexate (50 mg/m(2)) and misoprostol (800 mg vaginally, repeated as needed).For this study, women who presented for abortion prior to their 21(st) birthday were compared to older women. Outcomes of abortion were classified as complete medical abortion (CMA) and suction curettage (SC). Secondary outcomes included symptoms. We did an exit interview assessing patient satisfaction during the first year of the study, and have exit interview data for approximately half of the women enrolled.We assessed the relationship of age and outcomes first by bivariate analysis using SAS (SAS Institute, Inc., Cary, NC). A multiple logistic regression model was constructed using age, gestational age, and measures of parity.Results: There were 219 women who were under 21 (18 under the age of 18). Compared to older women, adolescents presented for abortion at the same gestational ages. Compared to older women, younger women were less likely to have finished college (2.8% vs. 38. 7%), and were less likely to have had previous pregnancies (45.4% vs 76.6%) and live births (20.2% vs. 51.3%)The distribution of symptoms during abortion, such as bleeding and cramping, was the same across age groups; the only symptom which was more frequently found in younger women was headache.Overall younger women had a slightly higher rate of CMA as older women (90.2% vs. 86.5%). However, the multiple regression (MR) model did not show an effect of the woman's age on CMA. MR demonstrated an adverse effect of prior live birth, and advancing gestational age on rates of CMA; younger women were less likely to have had prior live births.Younger women were just as likely to report overall satisfaction (86.7% vs. 84.6%) with the abortion procedure and other questions of satisfaction, but were less likely to agree that the bleeding and cramping were acceptable. Conclusions: Younger women having medical abortion with methotrexate and misoprostol have similar outcomes and satisfaction levels as do older women. There is no reason to discourage adolescents who seek abortion from using a medical technique. PMID- 10869972 TI - Signs of genital trauma in adolescent rape victims examined acutely AB - Background: Adolescent females are the most frequent victims of sexual assault, but studies to document the presence of genital findings in patients examined within 72 hours, using magnification and dye, have not been published. This study was designed to document the frequency and types of genital injuries in adolescent women following acute sexual assault, using chart and photograph review.Methods: A retrospective chart review was done of examination records of all female patients age 14 to 19 years of age who were evaluated at a Sexual Assault Response Team program over a 5 year period. Data was abstracted from charts by the nurse examiners, and photographs were evaluated by the physician reviewer. Analysis was done to determine the frequency, location, and severity of genital and anal injuries, and any historical factors correlating with injury, using Pearson correlation and two-tailed t tests.Results: Charts of 214 female subjects (mean age 16.3 years) were reviewed. The most common findings recorded by the nurse examiner were posterior fourchette tear (36%), erythema of the labia minora, hymen, cervix or posterior fourchette (18% to 32%), and swelling of the hymen (19%). Uptake of Toluidine dye was noted in 66% of patients in whom it was applied. Overall, 21% of patients were found to have no findings, and 40% had tears of the posterior fourchette or fossa. Time to examination was highly correlated with the degree of injury noted (p =.000). The incidence of hymenal tears in self-described virgins was higher than in non-virgins (19% vs. 3%, p =.008), however the total number or severity of other injuries was not significantly higher in virgins. Reported anal penetration was associated with a high frequency of anal bruising, abrasions or tears (14/23, 61%), while only 2/150 victims who denied anal penetration had tears (1%, p =.000). Victims who reported multiple physical symptoms such as pain, nausea, or vomiting were significantly more likely to be older (p =.034) and to have an increased number of non-genital injuries such as bruising, abrasions, and bite marks (p =.001). A higher number of non-genital injuries was also correlated with a higher number of total genital injuries (p =.003).Conclusions: Adolescent victims of sexual assault who were examined within 72 hours, using a magnification and dye were found to have tears of the posterior fourchette or fossa in 40% of cases. Hymenal tears were rare, even in self-described virginal girls. Timely examination of adolescent victims is important to document injuries, however, many victims will still have non-specific examination findings. PMID- 10869973 TI - Prevalence and predictors of low sexual assertiveness AB - Background: Personal beliefs about sexual attitudes and rights likely affect reproductive health behaviors. We examined the prevalence and predictors of low sexual assertiveness (SA) among sexually active females across three age groups (14-17 yo, 18-21 yo, and 22-26 yo).Methods: 904 white, black, and Mexican American females presenting at community-based family planning clinics in Southeast Texas between April 1997 and February 1998 completed an anonymous self report questionnaire that assessed demographic and reproductive characteristics, dating behaviors, mental health measures and occurrence of physical and sexual assault. Subjects also completed a sexual assertiveness scale that assessed one's right to make various reproductive health decisions and sexual communications to partners. A total SA score was obtained by summing the 13 items; scores were then dichotomized so those scoring in the lowest quartile could be compared to the others. Data were analyzed using chi-square and logistic regression stratified by age group (244 aged 14-17; 392 aged 18-21; and 268 aged 22-26).Results: 27% of subjects aged 14-17, 22% aged 18-21, and 17% aged 22-26 were identified as having low SA. For 14-17 years olds, logistic regression analyses revealed that those with low SA as compared to adequate SA were significantly more likely to be Mexican American (OR = 4.8) or black (OR = 3.1) than white, to have hand /= 1 (OR = 1.8). Among 22 26 years olds, low SA was associated with gravidity >/= 1 (OR = 2.8), being a high school dropout (OR = 2.1), and inconsistent birth control use in the last 12 months (OR = 1.9).Conclusions: These data suggest that 1 in 4 sexually active women report low SA and for two age groups these attitudes are associated with inconsistent contraception. Among 18-21 years olds, low SA is also associated with forced sexual contact and depressive symptoms. Thus, efforts to prevent unintended pregnancy and unwanted sexual contact may be more effective if individual levels of SA are taken into account. PMID- 10869974 TI - The independent and combined effects of physical and sexual abuse on health. Results Of a national survey AB - Background: Although physical and sexual abuse have been linked to health risk behaviors as well as mental health problems, it is unclear whether those young women who have experienced both physical and sexual abuse are at greatest risk. To examine the independent associations between physical, sexual, and/or both types of abuse and health status, mental health, and health risk behaviors among a national school-aged sample of girls. We hypothesized that the magnitude of risk would be highest for those reporting both types of abuse compared to those reporting neither or one type of abuse. Methods: In 1997, 3,015 girls in grades 5 through 12 participated in the Commonwealth Fund Adolescent Health Survey and responded to both questions inquiring about physical and sexual abuse. This sample was derives from a nationally representative cross-section of 265 public, private, and parochial schools with an oversampling of 32 urban schools to obtain ethnic diversity. Data were analyzed using chi-square and binary or multinomial logistic regression stratified by type of abuse (none, physical abuse, sexual abuse, or both). Results: About 8% (n = 246) of girls reported a past history of only physical abuse, 5% (n = 140) reported only sexual abuse, and 5% (n = 160) reported experiencing both physical and sexual abuse. Logistic regression controlling for demographic characteristics (grade level, ethnicity, family structure, and socioeconomic status) found those who reported both types of abuse as compared to those who did not report any abuse were significantly more likely to experience moderate-to-severe depressive symptoms (OR = 5.1), moderate to high life stress (OR = 3.3), history of bingeing and purging behavior (OR = 4.4), regular smoking (OR = 5.9) regular drinking (3.8), illicit drug use in the past 30 days (RR = 3.5) and fair to poor health status (OR = 1.9). In contrast, lowered adjusted odds ratios (1.8-2. 5) were seen for those reporting one type of abuse as compared to no abuse across most health outcomes.Conclusions: Those experiencing any type of abuse are at risk; however, those adolescent females who report both physical and sexual victimization are at much greater risk. PMID- 10869975 TI - Parents' knowledge of adolescent girls' boyfriends AB - Background: Previous research has demonstrated that parents play an important role in adolescents' reduction of risk of STD and unintended pregnancy through knowledge of their daughters whereabouts and by knowing their friends. Little is known, however, about parents; relationships with their daughters' boyfriends. Therefore, we sought to describe the daughters' perceptions of their parents' knowledge of their boyfriends.Methods: Questions regarding relationships with boyfriends and parents were assessed in the last wave of a longitudinal study of adolescent girls' romantic relationships. Of the 137 girls interviewed, 109 had had a boyfriend in the past 6 months, 78 had a current boyfriend. The results of this study were based on this subsample of 109.Results: The 109 adolescent girls had a mean age of 17.6 (range 14.9 to 19.3 years), and 80% were African-American and 20% Caucasian. Of those girls, 92 were sexually experienced (which 88% of their parents knew about), and 80 had engaged in sexual intercourse with this identified boyfriend. Nine of the girls reported that their parents had had no contact with their boyfriend in person or on the phone, and ten reported that they had neither met or talked on the phone to their boyfriend's parents. Three girls reported that neither they nor their boyfriends had had contact with the others' parents. Thirteen girls reported that their parents did not know their boyfriends at all well, 40 reported that their parents knew their boyfriends somewhat, and 56 reported that their parents knew their boyfriends well. Girls who had been involved in the relationship for a longer amount of time (F(2, 105) = 7.90, p <.001) and girls who perceived that their relationship would last a year or longer were more likely to think their parents knew their boyfriends well (chi sq = 14.1, p <.001). The results of the analysis between satisfaction with their romantic relationship and how well their parents knew their boyfriends approached significance (chi square = 5.5, p =.06). Of the six girls with a current boyfriend whose parents did not know their boyfriends, none were "very satisfied" with their relationship. Neither age nor race were related to how well the parents knew the boyfriend.Conclusions: These results indicate that most adolescent girls' parents have some contact with their boyfriends. Contact appeared to be related to length of relationship. Health care professionals should be sensitized to the needs of girls who are beginning new relationships, or are not satisfied with their relationship and who do not have access to parental support. Future research should examine familial/parental factors related to knowledge of the boyfriend and whether the lack of a developmental trend is the result of a restricted age range or of other family issues. PMID- 10869976 TI - Musculoskeletal pelvic pain in a pediatric and adolescent gynecology practice AB - Background: Pelvic pain is a common complaint encountered in pediatric and adolescent gynecology. Etiologies are similar to those found in adult women, but the incidence and presentations vary with age. The purpose of this study is to review musculoskeletal (MS) pelvic pain in a pediatric and adolescent gynecology setting. Methods: A retrospective review of charts of 63 patients presenting to a private practice pediatric and adolescent gynecologist between 7/1/97 and 6/30/99 was performed. To be included in analysis, patients had a diagnosis of pelvic pain which could not be explained by standard gynecologic history, physical exam, laboratory, and ultrasound evaluation or did not respond to standard treatments for known endometriosis. A history of laparoscopy was not required, but when it was performed it could be used to exclude patients from analysis if a reason for the pain was identified. All patients who fulfilled these criteria had been screened for MS etiologies of pelvic pain using the leg lift (Carnett test) and/or head lift. Results: Sixty-three patients aged 9-23 (mean 15.21, SD 2.71) fulfilled the criteria for evaluation. Diagnoses included irritable bowel syndrome (N = 4, 6.35%), interstitial cystitis (N = 1, 1.56%), unexplained (N = 7, 11.11%), endometriosis not responding to ablation & GnRH agonists (N = 2, 3.17%), endometriosis not responding to ablation & GnRH agonists & MS pain (N = 7, 11.11%), and MS pain (N = 42, 66.67%). Mean age of those with MS pain was 15. 27 (SD 2.94), and mean duration of symptoms prior to diagnosis was 17.97 mo (SD 20.90, range 1 week-7 yr). On physical exam, trigger points were identified as causative factors in 5 (10.20%), and 40 (81.63%) had a + Carnett test. Of those with a final diagnosis of MS pain, only 5/31 (16.31%) responded to nonsteroidal anti-inflammatory agents, 6/30 (20.0%) responded to OCPs, and 3/11 (27.27%) responded to DMPA-2/3 also had a diagnosis of endometriosis. Nineteen (38.78%) had been surgically explored for the pain in the past, 1 by laparotomy & 18 by laparoscopy. Only 3 (15.79%) had symptomatic improvement after surgery. Physical therapy resulted in resolution of symptoms in 20/21 (95.24%) who completed treatment. Four of 5 (80%) who underwent trigger point injections responded.Conclusions: MS etiologies of pelvic pain are common in the adolescent age group and respond well to physical therapy. Physical therapy might be employed as an early intervention prior to surgery in adolescent girls with unexplained pelvic pain. PMID- 10869977 TI - Monitoring std prevalence and reproductive health care among high-risk adolescent women AB - Background: Urban adolescent women, particularly those in socially disadvantaged situations are at high risk for infections with Chlamydia trachomatis (CT), and Neisseria gonorrhoeae (GC) and their sequelae. Non-invasive screening technologies, including ligase chain reaction (LCR) and polymerase chain reaction (PCR) urine tests for CT and GC allow for provision of screening services in settings frequented by high-risk adolescents where such services have not been traditionally provided. In 1999, the CDC lead a multi-site project monitoring STD prevalence and reproductive health service, collecting a standard set of variables in women <20 yrs in juvenile justice system, drug treatment centers, and school-based clinics. This report provides preliminary data from Alabama and Colorado. Methods: Adolescent women who present for intake/services at selected facilities are enrolled in the project. In collaboration with CDC, sites developed a set of standard variables to be collected, including demographics, risk behaviors, symptoms, test performed, and test results. Dependent on services already performed, sites added CT and/or GC urine PCR/LCR and pregnancy testing to existing screening protocols.Results: In first 3-months, 533 adolescent women were enrolled in this on-going study; 379 in juvenile institutions, 38 in drug treatment centers, and 116 in school-based clinics. Mean age was 16 yrs, over 90% were sexually experienced, nearly 50% did not use condom with the last sex, less than 40% reported "current" birth control use, over 2/3 had sexual intercourse in past 3 months, many with multiple partners. Prevalence rates for CT and GC are summarized in the table. Conclusions: Non-invasive (urine) LCR/PCR tests allow for easy monitoring of selected STDs among adolescent women in high-risk settings. These findings may contribute to a sentinel surveillance system among high-risk populations and, in turn, may provide the rationale for the development of health care/intervention priorities at policy level. PMID- 10869978 TI - Screening females for chlamydia trachomatis (CT) In a large managed care organization (Mco). A new hedis measure AB - Background: Since CT testing for females is a new Health Plan Employer Data and Information Set (HEDIS) measure to assess MCO quality of care, we determined the proportion of females enrolled in a large closed panel vertically integrated MCO serving a demographically diverse population who were tested for CT at least once during 1998 and resulting CT prevalences.Methods: A data base with MCO members >/= 12 yrs old tested with CT DNA probes (GenProbe) from 1/1/98-12/30/98 was examined. Only females 12-24 yrs old and enrolled at least 11 continuous months in 1998 were included in the analysis.Results: In 1998, 5425 (13.1%) of 41,566 females 12-24 yrs were CT tested and 849 (2.0%) tested positive at least once. Proportion of females tested and proportion of females testing positive at least once varied by age and clinic location. Among 11, 562 12-14 yr females, 277 (2.4%) were CT tested and 48 (0.4%) tested positive; among 18,155 15-19 yr females, 2,424 (13.4%) were tested and 572 (3.2%) tested positive; and among 11,849 20-24 yr females, 2, 724 (23.0%) were tested and 229 (1.9%) tested positive. The proportion 12-24 yr females tested was 579/3,945 (14.7%) with 92/3, 945 (2.3%) testing positive in the Baltimore area; in Washington, DC/Maryland area 2,999/21,085 (14.2%) were tested with 601/21,085 (2. 9%) testing positive; in Northern Virginia 1,847/16,536 (11.2%) were tested with 156/16,536 (0.9%) testing positive.Proportion of females with at least one positive test among those females who were tested for CT also varied by age and clinic location. Among 277 of 12-14 yr females tested 48 (17.3%) tested positive at least once; among 2,424 of 15-19 yr females tested, 572 (23.6%) tested positive at least once; and among 2,724 of 20-24 yr females tested, 229 (8.4%) tested positive at least once. The proportion of females tested with at least one positive test among 12-24 yr females was 92/579 (15.9%) in the Baltimore area, 601/2,999 (20.0%) in Washington DC/Maryland area, and 156/1,847 (8.4%) in Northern Virginia.Conclusion: CT testing of adolescent females 12-19 yrs revealed a high proportion of positive tests. The lower proportion of positive tests in young adults 20-24 yrs may be due to differences in prevalence and increased number of tests performed. CT testing practices and rates varied by location. Therefore, in a large MCO, a uniform system wide approach in identifying and screening sexually active adolescent females may identify a large reservoir or asymptomatic infection. PMID- 10869979 TI - Do they, don't they, or why haven't they?. Contraceptive use patterns among inner city sexually active female adolescents AB - Background: Female adolescents frequently practice unprotected vaginal intercourse leading to nearly 1 million unintended pregnancies and 3 million sexually transmitted diseases (STDs) a year. Qualitative reasons for three contraceptive use patterns were explored. Contraceptive use patterns were quantified and correlated with gynecologic history.Methods: A total of 146 predominantly non-white (81%) sexually active female adolescents with a mean age of 17.2 +/- 1.9 years (13-21 years), completed a 45-minute self-administered, computerized questionnaire as part of a larger project. The questionnaire assessed sexual, contraceptive, gynecologic and obstetric history. Nearly a third (32%) had ever been pregnant and 32% had ever had a STD. Subjects were asked to describe their contraceptive use patterns for condoms, pills, and Depo-Provera as either "never used," "have used, but stopped," or "used before and am still using." Adolescents typed in their qualitative reasons for never using, stopping, or still using each of the three methods of contraception.Results:None of the three condom use patterns were significantly correlated with ever having a STD or being pregnant. Of those who had ever been pregnant, 70% had stopped using Depo Provera as compared to 38% who were still using Depo-Provera, p < 0.02; 47% had stopped using pills as compared to 25% who were still using pills, p = 0.04; 47% had stopped using pills as compared to 22% who had never used pills, p < 0.01. "Using another contraceptive" was the reason why one-third of subjects never used condoms. The most common reasons for never using pills were "on Depo-Provera," "would forget," and not liking to take pills. Reasons for never using Depo Provera included using another contraceptive, not liking shots and "might make me fat." The top four reasons for discontinuing condom use were being in a trusting, monogamous relationship, using another form of contraception, abstinence, and "don't like the way condoms feel." Two of the main reasons for stopping pills and Depo-Provera were weight gain and altered menses. However, those subjects who stopped taking the pill reported "kept forgetting" and "they made me sick" as other reasons for stopping. "Prevent pregnancy" was by far the most common reason for still using pills and Depo-Provera, while for condoms "prevent STDs" was cited with equal frequency to "prevent pregnancy." Having regular or no periods were frequent reasons for ongoing use of pills and Depo-Provera, respectively. Depo-Provera users also reported "don't have to worry about missing a pill" as a major reason for continuing use.Conclusions: Understanding the reasons why adolescents never use, stop using, or continue to use a particular method of contraception may provide a focus for more effective counseling. PMID- 10869980 TI - Vaginal douching in adolescents attending a family planning clinic AB - Background: One of the variables most consistently associated with vaginal douching is race, with African-American women douching more regularly. Sparse data exists in the medical literature about the practice of vaginal douching among adolescents. The purpose of this study was to assess the prevalence, knowledge, attitude, and practices of vaginal douching among adolescent females attending a public family planning clinic, and determine whether African-American (AA) females douche to a greater degree than Caucasian females.Methods: In this cross-sectional study, a one-page questionnaire was administered to all adolescent females (/= 18 months) and the least (< 6 months) time were most likely to be correct.Conclusions: This study demonstrates that many (40%) adolescents do not know when their injection is due, and in fact, the correctness of adolescents' knowledge is not consistent over time. Age and race were not significantly related to knowledge of timing. Therefore, ALL adolescents should be provided with reminders about the timing of their next injection. Finally, the curvilinear relationship needs to be examined further. This finding that adolescents on Depo Provera for 6-12 months were the least knowledgeable is consistent with research on oral contraceptive adherence indicating this is a vulnerable time period for contraceptive discontinuation. PMID- 10869982 TI - Growth hormone deficiency as the only identifiable cause for primary amenorrhea AB - Background: There is much evidence that growth hormone plays an important role in the development and function of the reproductive system of both males and females. Growth hormone exerts its effects on the ovarian follicular cycle directly or by local production of insulin-like growth factor 1 (IGF-1). It is known that growth hormone deficiency during childhood may delay pubertal development, but there is limited data about primary amenorrhea in GH-deficient girls with sufficient stimulated gonadotropin levels.Methods: Case series.Results: In the evaluation of primary amenorrhea and delayed puberty, 3 cases of adolescent females aged 17-19 years were identified as isolated GH deficiency. Among the 3 patients, 2 had history of intracranial surgery due to hydrocephalus (shunt operation) or prolactin-secreting pituitary macro-adenoma (transphenoidal surgery, one year before). 17-year-old patient with shunted hydrocephalus and 19-year-old patient with primary amenorrhea showed short statue (< 5%) and delayed bone maturation. The patient undertaken transphenoidal surgery for prolactinoma showed normal height and bone maturation. There was no familial history of delayed puberty. On physical examination, 3 patients showed variable degree of breast development from Tanner stage II to IV without sex-steroid replacement. In sella MRI, small pituitary gland were identified in 2 patients with short statue and delayed bone maturation. All of the 3 patients underwent combined pituitary function test. After insulin-induced hypoglycemia, peak growth hormone levels of the 3 patients were 0.08, 1.4 and 1.4 ng/ml and were compatible with growth hormone deficiency. Peak LH after intravenous gonadrelin (FACTREL) were 19.0 to 56.1 mIU/ml and LH % responses were 217 to 1100% and were hence defined as not being gonadotropin deficiency. Other anterior pituitary functions were normal in all of the 3 patients.Conclusions: We found isolated growth hormone deficiency as the only identifiable cause for primary amenorrhea in three patients with sufficient gonadotropins secretion. These findings suggest a complementary role of GH to gonadotropins in the occurrence of menarche. PMID- 10869983 TI - 47,Xxx in an adolescent with premature ovarian failure and autoimmune disease AB - Background: Premature ovarian failure (POF) is often associated with autoimmune disorders. The 47,XXX karyotype has also been associated with POF and other genitourinary abnormalities. Following is a case of a 17 year old with immune thrombocytopenic purpura (ITP), POF, 47, XXX and a positive antinuclear antibody (ANA).Case Report: A 17 year old Caucasian female was referred to the Adolescent Health Clinic for evaluation of oligomenorrhea with secondary amenorrhea. Thelarche occurred at 12 years, and menarche at 13 years of age. Since then she had a total of five menstrual periods, spaced 1-15 months apart and lasting 3-5 days. Her last menstrual period was six months prior to presentation. Past medical history was significant for chronic ITP diagnosed seven months prior to presentation, when she developed easy bruising. She was treated with IV gamma globulin and had a moderate response, but relapsed several weeks later. She was started on oral prednisone and had a good response, but continued to relapse whenever steroids were tapered. She was therefore maintained on prednisone 10 mg QOD. There was no family history of irregular menses or autoimmune disease. Physical exam revealed a well-appearing, slightly Cushingoid 17 year old. Physical and cognitive development were age-appropriate. There were no stigmata of Turner Syndrome. The thyroid was normal. Breasts were Tanner 5; public hair was Tanner 3. The external genitalia were normal and appeared well-estrogenized. The remainder of the exam was unremarkable. Pelvic ultrasound demonstrated a normal uterus and ovaries. Laboratory evaluation was significant for elevated gonadotropins and nondetectable estradiol. ANA was positive at 1:320 with a speckled pattern. Blood counts, serologies, complement levels, and coagulation studies were otherwise normal. Cytogenetic studies revealed a 47,XXX karyotype. The patient was placed on an estrogen/norethindrone hormone replacement patch for premature ovarian failure. To date, she has developed no further symptoms, and does not meet criteria for a diagnosis of systemic lupus erythematosis.Conclusions: A 47,XXX karyotype was found in a 17 year old with POF and ITP with a positive ANA. The presence of known autoimmune disease in a woman with POF should not dissuade the physician from evaluating for a potential genetic cause. PMID- 10869984 TI - Isolated tubal torsion at menarche- a case report AB - Background: Adnexal torsion is a well-recognized cause of acute pelvic pain. Isolated tubal torsion with ovarian sparing has certainly been documented, but is uncommon. Although risk factors for the latter include a menstrual period, menarche in particular is not known to predispose a patient to this event. Severe unilateral pelvic pain with first menses is more likely to herald a congenital mullerian anomaly and cryptomenorrhea, particularly when accompanied by a pelvic mass. We present a case of tubal torsion where a coincidental, yet misleading temporal relation to menarche led to a delay in laparoscopy and ultimate diagnosis.Case: KG, an eleven-year-old female, experienced severe right-sided dysmenorrhea with her first and second menses in August and September 1999 respectively. Between episodes, pain, although still present, was more tolerable and the patient never required hospitalization. Ultrasound revealed a lobulated inhomogeneous mass posterior to the uterus and extending from one normal ovary to the other (Figures). MRI further described the mass as pseudoencapsulated with inhomogeneous areas of high attenuation on T1 and T2 images (Figures). Findings were consistent with an endometrioma, but admittedly could have represented a hemorrhagic cystic mass. No definite mullerian anomaly was seen to explain advanced endometriosis, but two focal areas within the endometrial canal raised the possibility of a uterine septum. Examination of the patient (one week after presentation) was not very helpful although she was pubertal, did have a hymenal septum and was mildly tender on bimanual examination in the Pouch of Douglas. The patient had been started on continuous oral contraceptives while undergoing investigations. Pain only recurred during an episode of break-through bleeding. Ultimately she came to laparoscopy and hysteroscopy where chronic right tubal torsion and necrosis was identified with an inflammatory/hemorrhagic reaction in the pelvis (Photos). There were no identifiable fimbria of the right tube which was densely adherent distally to perirectal fat (Photo). No obvious precipitant was found. Laparoscopic lysis of adhesions and right distal salpingectomy was performed (Photo). Her uterine cavity was in fact normal (Photo)Conclusion: Whether or not this patient's right tube was originally normal will never be known. Congenital abnormalities of fallopian tubes do occur and can predispose to torsion. Nonetheless, adnexal torsion must always be kept in mind whenever a woman presents with unilateral pelvic pain. Early diagnosis is paramount in children and women of reproductive age in order to improve the likelihood of adnexal salvage and future fertility. A "gold-standard" radiological investigative tool continues to elude us. Laparoscopy, albeit more invasive, remains an invaluable procedure in this context with relatively low morbidity as compared to the consequences of delayed diagnosis. PMID- 10869985 TI - Labial adhesion as a complication of primary genital herpes in young women AB - Background: Genital herpes is a common sexually transmitted disease in adolescents. It may be associated with significant morbidity if not diagnosed on time or not properly treated. The objective of this study was to determine the incidence of labial adhesion secondary to primary herpes in young women and the possible predisposing factors for this complication.Methods: Analysis of the clinical data regarding primary genital herpes in young women in the adolescent clinic at a university hospital in outpatient clinic setting. Cases of primary genital herpes seen between December 1(st) 1998 and November 30(th) 1999 were included.Results: A total of 34 female adolescents with age range 12-19 years were diagnosed with primary genital herpes during this time period. 7 patients (20.6%) were found to have severe labial adhesion at time of diagnosis. All 7 patients were seen by other providers prior to their visit to the adolescent clinic and 4 were correctly diagnosed. All 7 patients were given antiviral therapy, but none was given local treatment. At time of diagnosis all 7 patients had anuria for more than 24 hours and severe pain and discomfort. 3 patients had Diabetes Mellitus (one of these was also pregnant) and one patient had asthma. The age range for these 7 patients was 13-17. Treatment with local anesthetics helped resolve the adhesion in 5 patients and surgical treatment was needed in the remaining two patients.Conclusion: Labial adhesion is a common, severe complication of primary genital herpes in young women. Very young age, chronic medical conditions, incorrect diagnosis and lack of topical treatment may predispose to the development of this complication. Use of topical therapy should be an integral part of the comprehensive treatment for primary genital herpes in female adolescents to alleviate discomfort and prevent urinary retention and labial adhesion. PMID- 10869986 TI - Urban girls, internet use, and accessing health information AB - Background: With the burgeoning growth of the Internet, many have raised concerns over whether and how underserved populations might be included in the Information Revolution. This study examines how urban adolescent girls access the Internet and whether it is a viable source for health information.Methods: During Fall 1999 in New York City, 176 girls completed an anonymous survey where half were from a private high school (N = 86) and the other half were patients at a health clinic serving ethnically diverse and disadvantaged youth (N = 90). Data were analyzed to provide information on how urban girls use the Internet.Results: Internet use was widespread; 99% of the high school and 83% of the clinic girls said they used this medium (p <.001). On frequency, 32% of high school girls compared to 13% of the clinic girls used the Internet 6-7 days/wk (p <.001). Among the high school girls, 30% indicated that they used the Internet less than 30 min/wk and another 18% reported using the Internet more than 5 hrs/wk. Among clinic girls, 51% used the Internet less than 30 min/wk while 9% reported more than 5 hrs/wk. Comfort levels differed with 20% of the high school compared to 10% of the clinic girls reporting "extreme" comfort using the Internet (p <.001).Urban girls accessed the Web from multiple locales. Almost all (98%) of the high school girls compared to 45% of the clinic girls reported that they used the Internet from home (p <.001). Most high school girls (82%) used the Internet at school compared to 63% of the clinic girls currently in school (p <.05). More clinic girls used the Internet at another family member's house (28% vs. 14%, p <.05). Similar percentages of high school and clinic girls used the Internet at a friend's house (46%), the library (31%), or a community center (3%).With similar rates among high school and clinic girls, 44% reported that they had tried to get health information from the Web. Of these (N = 74), 50% said that they got information on different diseases, 43% on diet/nutrition, 34% on fitness/exercise, 26% on sex, 24% on alcohol/drug abuse, 20% on mental health, 14% on medicines, 12% on violence among peers, 10% on parenting, 8% on violence among dating partners, 7% on tobacco and smoking, 7% on emotional or physical abuse, 3% on sexual abuse, and 1% on illness support groups. Clinic girls were more likely to have tried to get information on sex (p <. 001) and parenting (p <.001) from the Web. Many girls (59%) indicated that it was "very" or "extremely" worthwhile to have general health information on the Web, with no significant differences between high school and clinic girls. As well, 71% and 62% said it was "very" or "extremely" worthwhile to have access to contraceptive and diet/nutrition information, respectively. Conclusions. High school girls, as anticipated, were technologically savvy. Somewhat surprising, however, were the clinic girls' high levels of Internet use and access. Significant differences revealed that the high school girls were more on-line than clinic girls, but clinic girls were still using the Web frequently. A considerable number or urban girls had tried to get a range of health information from the Internet. This study offers promising statistics on how urban adolescent girls are accessing and using the Internet, especially for health information. PMID- 10869987 TI - Population-level intervention to promote chlamydia screening. Moving toward implementation of chlamydia hedis 2000 measure AB - Background: HEDIS 2000 measure includes chlamydia screening in women which is designed to assess the percentage of sexually active women 15 to 25 years who have received at least one screening test for chlamydia during the reporting year. This study is being undertaken to determine feasibility of implementing a population-level intervention within HMOs to promote chlamydia screening. This abstract presents preliminary findings from the Birmingham project of this multicenter study.Methods: In partnerships with two HMOs, series of outreach methods were used in a stepwise fashion to determine potential barriers and enabling factors for the implementation of chlamydia HEDIS measure in a conservative social environment. Mail outreach was sequentially combined with newspaper, TV, radio advertisements and poster displays. Both qualitative and quantitative impact of the outreach efforts were measured across the timeline. The measures included reporting for chlamydia screening (urine LCR) and infection rate, monitoring chlamydia hotline and staffed phoneline use, and assessment of untoward effects and cost-analysis of the chlamydia outreach campaign.Results: The key findings are: the benefit of chlamydia screening is not understood by general public, letters send by Health Plans to their members are not read by many subscribers, and there are wide gaps between adolescents and their parents, in knowledge, attitudes, beliefs in regard to obtaining information and accessing the screening services (teens prefer hotline, brochure in an envelop addressed to teens, incentives for reporting to the clinic for screening, vs. parents prefer staffed phone consults, "exposed" brochure addressed to parents, and no incentives). A month of sustained and repeat multi-media campaign resulted in 330 hotline calls, 83 phone calls and only 17 subjects being tested (3 were positive) though many more intended to come. Cumulative effects and cost of various outreach efforts are being monitored. Informational chlamydia brochures and free test cards mailed to the homes generated no negative response from the subscribers, contrary to the concerns of the HMOs. Conclusion: To be effective, investment in public education campaign and social marketing strategies must be integrated in population-level intervention for the implementation of the chlamydia HEDIS 2000 measure. PMID- 10869988 TI - Ovarian cortex cryopreservation in pediatric and adolescent medicine AB - Background: Increased pediatric/adolescent cancer survivor rates have enhanced awareness of long-term effects of therapy, specifically gonadal failure. Ovarian cortex cryopreservation may hold the promise of fertility for those at risk for ovarian failure due to medical therapy. The object of this study was to determine if an ovarian cryopreservation program is feasible and to define suitable candidates.Method: A MEDLINE search supplemented by bibliographies. The review was limited to English articles on ovarian failure rates following radiation and/or chemotherapy and on ovarian cryopreservation. Investigators in the field were consulted to identify other sources.Results: Approximately one third of postpubertal females exposed to chemotherapy or radiotherapy develop ovarian failure. The risk is mostly significant for patients exposed to pelvic radiotherapy (up to 32% decrease in fertility) and alkylating agent based chemotherapy (infertility in 22%). A ninefold increase in premature ovarian failure results from exposure to combined pelvic radiotherapy and alkylator based chemotherapy. Practically all patients exposed to multiple agent chemotherapy combined with pelvic radiotherapy at doses used in preparation for bone marrow transplant will undergo irreversible loss of ovarian function. Currently human ovarian cortex can be cryopreserved, thawed and stimulated with gonadotrophins to produce follicles when transplanted into immunosuppressed mice, however there has yet to be any human pregnancies. The immunosupressed mouse model could also serve as a test to determine whether the tissue carries metastatic risk prior to reimplantation into the donor.Conclusion: Based on the literature we propose ovarian cortex cryopreservation and banking for postpubertal females prior to chemotherapy and/or radiation therapy that holds a high risk of ovarian failure. In the future this may provide oocytes for reproductive purposes. A protocol is currently under approval by the Hospital for Sick Children's ethics committee. PMID- 10869989 TI - What do adolescent females know about breast cancer and prevention? AB - Background: Many lifelong habits begin during adolescence. These habits can have profound, long-term ramifications on health. An important health habit is self breast examination (SBE). The purpose of this study is to assess adolescent females' knowledge of breast cancer and breast cancer prevention. A survey was developed with questions that focused on female adolescents' knowledge of these topics. This information will assist health professionals in educating teens and forming public policy.Methods: Anonymous surveys consisting of ten questions were distributed to 280 females aged 13-17. These questions pertained to breast cancer with an emphasis on SBE, mammography, and risk factors. Questions were analyzed by chi square calculations. An additional section of the survey contained demographic characteristics of the respondents. Parental consent was obtained prior to completion of the surveys in accordance with the Institutional Review Board at St. Luke's Hospital. Surveys were administered during health class in a local high school.Results: One hundred and fifteen surveys were returned. Ages of participants were 13 to 17. The ethnic background of all participants were similar in that the majority were Caucasian. The overall percentage of correct answers was 65%. The majority of students knew what a mammogram is (92%) and how often screening should occur (65%), however, only 25% knew at what age screening should begin. It was encouraging that 80% of the students knew how often to perform SBE although only about half (53%) knew the time of the month this should be done. It was also encouraging that 83% knew that breast cancer is the second leading cause of cancer death, but the knowledge regarding risk factors that could possibly affect them was poor (36%). A statistically significant findings was that in the twenty percent of the students who had been taught SBE, 10 (43.5%) actually perform them. This is in relation to 2.2% of students who perform exams without any prior instruction. There was no statistically significant difference in the final score between students who had been taught how to perform exams and students who had not.Conclusions: Developing sound health habits as an adolescent should transcend to good health maintenance practices as an adult. Our study showed that adolescent females significantly lack knowledge relating to breast cancer. Adolescent females need to be better educated on the basic facts, including risk factors, screening procedures and SBE. With the incidence of breast cancer so high, knowledge of breast cancer and its prevention may result in earlier diagnosis and subsequently better long term outcomes. PMID- 10869990 TI - Obstetric outcome of adolescent pregnancies AB - Background: To determine if adolescent pregnancies are at increased risk of poor obstetrical outcome compared with a general obstetrical population.Methods: A five-year retrospective review of the Toronto Hospital for Sick Children's Teenage Pregnancy Unit was carried out. Information was available on 209 patients < 19 years age between January 1994 and December 1998. This was compared to information available from a database of all women delivering at the same hospital, The Toronto Hospital General Division, during the same time period (n = 13,557). The Chi-square test of independence was used to compare the data and is reported as adolescent group vs. hospital group.Results: Labour was induced in 25.5% vs. 21.8% (p = 0. 20). Epidural anaesthesia was received by 63.5% vs. 53% (p < 0.05). The incidence of preterm delivery (<37 wks) was 13.5% vs. 8.1% (p < 0.05), low-birth-weight babies (< 2500 g) 13.4% vs. 8.6% (p < 0.05) and small-for gestational-age babies (<2 SD) 1.9%. The incidence of post-term delivery (>41 wks) was 12.5% vs. 4.3% (p < 0.001), macrosomia (>4000 g) 1.9% vs. 9.2% (p < 0.001) and large-for-gestational-age babies (>2 SD) 0.5%. Operative delivery (forceps or vacuum) occurred in 19.7% vs. 19.9% (p = 0.94) and caesarian section in 6.2% vs. 20.1% (p < 0.001). APGARs <7 at five minutes were found in 2.4% vs. 3.1% (p = 0.60). 12.0% of infants were admitted to the neonatal nursery. There were no stillbirths. Conclusions: Both preterm deliveries and low-birth-weight babies were more frequent in the adolescent group although the incidence of SGA babies was low. The low caesarian section rate also likely reflects these findings. Postterm delivery was also more common, yet macrosomia occurred less frequently. PMID- 10869991 TI - Non-classic 21-hydroxylase deficiency in an 18-year-old female athlete. A case report AB - Background: In non-classic 21-hydroxylase deficiency, age at presentation and genital findings are variable. Late diagnosis with dramatic signs of virilization precludes early treatment and thus prevention of anatomic and psychosocial consequences. The following case illustrates the complexity of late diagnosis.Case: An 18-year-old West Indian female was seen for evaluation of clitoromegaly and hirsutism discovered in the emergency department when she presented after sexual assault. She had allegedly been drugged and raped in her dorm room. She was a college student with an athletic scholarship and had a striking masculinized, broad-shouldered appearance. She denied any use of anabolic steroids or other drugs. Menarche was at age 16 with infrequent menses. She was sexually active with 4 life-time partners, all male. On physical exam, her height was 152 cm, weight 50 kg, blood pressure 110/70 mm Hg. Breasts were hypoplastic with hyperpigmented nipples. She was hirsute with a Ferriman-Gallwey score of 14. Genitalia were abnormal with clitoris measuring 5.5 x 1.5 cm and posterior labial fusion. Initial non-fasting serum 17-hydroxyprogesterone level was 2890 ng/dL, testosterone was 274 ng/dL, and cortisol was 9 &mgr;g/dL. Chromosome analysis was 46, XX and ACTH stimulation test confirmed the diagnosis of 21-hydroxylase deficiency. The patient was initially reluctant to begin glucocorticoid treatment because of concern that it would decrease her muscle mass and negatively impact on her athletic performance and scholarship support. One year after diagnosis and 10 months after beginning treatment, she elected surgical correction of her clitoromegaly because of extreme embarrassment over having erections during sex. She underwent excision of most of the corpus cavernosum with repositioning of the glans. The neurovascular elements were preserved. The patient is pleased with the cosmetic result and reports no change in achieving orgasm. She has not notices any change in muscle mass or athletic performance since beginning glucocorticoid therapy.Conclusion: This case illustrates the somatic and genital abnormalities as well as the psychosocial impact of a delayed diagnosis of 21-hydroxylase deficiency in this young female athlete. PMID- 10869992 TI - Teen compliance with contraception. Analysis Of the ohio medicaid claims data AB - Background: Teen pregnancy is an important and costly issue for society and often commits the mother to a life of poverty. Physicians play an important role in preventing teen pregnancy by identifying teens at risk for pregnancy and prescribing contraception. This study examines for the first time, whether pharmacy data can be used to evaluate teens' compliance with contraception. Such a method may be useful to physician groups, health plans or state agencies to identify teens at risk for unwanted pregnancy and target interventions.Methods: Secondary analysis of the Ohio Medicaid Claims Data from 1998 was done identifying continuously enrolled young women ages 12-19 years, at high risk for pregnancy defined by ICD 9 codes for an STD, an abnormal pap smear, contraception, or a CPT code for a pregnancy test.Results: Between 1/1/98-3/31/98 we identified 4009 females, ages 12-19 years at high risk for pregnancy. During the study interval, 1084 (27%) became pregnant. 1732 (43%) used no prescription contraception and 1190 (30%) used a method at some time. Depo only (551) and OCP only (552) use was equal. Eighty-three combined the use of Depo and OCP. Compliance was poor. Only 20% of the contracepting group had coverage for the full year and approximately 30% used a method for 3 months or less. There was little difference in age or concurrent chronic physical or mental illness between the contracepting and non-contracepting groups. General practice physicians provided 40% of the OCP prescriptions to teens compared with 36% Ob/Gyn, 10% Pediatrics, and 6% Internal Medicine.Conclusions: Analysis of pharmacy data to evaluate the contraceptive compliance of teens at high risk for pregnancy demonstrated poor compliance. This may be a useful tool for identifying teens at risk for unwanted pregnancy and targeting interventions. PMID- 10869993 TI - Self-induced carving and scarification of the forearms as a manifestation of sexual abuse in a 14-YEAR-old adolescent girl AB - Background: Numerous cutaneous abnormalities have been described in adolescent girls who have been sexually abused. These include bruising, bite marks, cuts, scratches, abrasions, edema, hematomas or other evidence of struggle. Victims frequently shower or bathe excessively in an effort to cleanse their skin following such an unwanted encounter. However, there is a paucity of information in the literature regarding the association of sexual abuse and removal of the superficial layers of the skin as a more desperate attempt by teenagers to rid themselves of the perpetrator. The purpose of this paper is to heighten awareness among practitioners that self-induced cutting and carving of the forearms with scarification may occur as a manifestation of sexual abuse in young women.Methods: A 14-year-old girl was seen in an adolescent medicine consultation setting during the spring of 1999 for evaluation of an anxiety disorder. During the interview the girl related that she had been under considerable stress and that she was having difficulty sleeping. She also had worsening of facial tics that had been previously noted in association with obsessive compulsive behaviors. She had been receiving psychotherapy and was being treated with fluoxetine, but the symptoms were becoming more severe. On examination she appeared very anxious and demonstrated numerous involuntary, repetitive facial grimaces. Similar twitching movements of the neck were also noted. In addition, she had several well healed scars over both forearms. The lesions were linear with a range of one half to one inch in width and three to four inches in length. The remainder of the general physical examination was entirely unremarkable.Results: The etiology of the scars was initially unknown. Upon further questioning the patient was asked directly about what had caused these marks. At that point she broke down and cried as she related that had been sexually assaulted several months earlier. She stated that she carved out tatoos on her arms to get rid of the skin that the perpetrator had touched when he forcibly held her down and raped her. She believed that the scars were "clean" as they were covered with newly regenerated skin. Her gynecologic exam was normal. Further psychiatric intervention was then obtained as it became very apparent that the patient had numerous unresolved emotional conflicts stemming from the attack. Conclusions: This paper documents that carving and scarification of the forearms may be due to self-induced injury in adolescent girls who have been the victims of a sexual assault. Clinicians who care for teens should be aware of this finding as it appears to be an important dermatologic manifestation of sexual abuse. Early recognition of this sign and prompt treatment of the underlying psychiatric issues are essential to an optimal outcome so that any further complications following such a traumatic event can be minimized. PMID- 10869994 TI - Previous clinical diagnosis of chlamydia helps patients predict outcome of new chlamydia clinical test AB - Background: As part of an ongoing clinical trial evaluating the effectiveness of an intervention aimed at decreasing STDs in adolescent females, a host of different background measures were collected at baseline. These measures include self-reported behaviors, risk and probability estimates concerning STDs, self reported prior medical diagnoses, and a Chlamydia trachomatis (Ct) infection assay, among other measures. This analysis examines participants' accuracy in estimating their own chance of chlamydia infection.Methods: Participants were 131 sexually active adolescent females, 79% black, 13% white, mean age 16 (range 13 19), who had been recruited from an adolescent health service. At the time of this study, participants were not primarily being seen for a clinical visit. Participants answered two questions that are examined here: The first asked, "What is the percent chance that you have chlamydia right now," accompanied by a visual scale ranging from 0% (no chance) to 100% (certainly). The second question asked, "In your life, have you ever been told by a doctor or nurse that you had chlamydia?" Following all questions a vaginal swab was collected by the participant, which was analyzed by polymerase chain reaction (PCR) for Ct. A multiple regression was performed, predicting the outcome of the chlamydia swab test using participants' estimates that they had chlamydia, their self-reported prior diagnosis with chlamydia (dichotomous), and the interaction between these two variables.Results: The regression F(3, 127) = 10.95, p <.01, revealed that participants' estimates of the chance that they had chlamydia significantly predicted outcome of the clinical test, t(127) = 3.34, p <.01. Previous diagnosis with chlamydia alone did not predict outcome of the clinical test, t(127) = 0.84, ns, but the interaction between the two variables significantly predicted clinical outcome, t(127) = -2.07, p <.05. A previous chlamydia diagnosis made participants' own estimates highly predictive of the clinical outcome; the estimates of those who had never been previously diagnosed with chlamydia had very little predictive power. Conclusions: Those participants who had prior experience with a chlamydia diagnosis were much more able to interpret their own risks and symptoms, and arrive at a very accurate estimate of the chance that they had a chlamydia infection. It is important to note that these participants were not seeking care at the time of this study, but that they had enough information to predict clinical outcome of a chlamydia test, especially those who had been diagnosed with chlamydia in the past. Arguably, these are they very patients who are most at risk for long-term sequelae of chlamydia infection, and asking (or having the patient ask) the relevant questions may increase the likelihood that they are identified and treated promptly, and early in the course of infection. PMID- 10869995 TI - Comparison of microscopic examination and human papilloma virus dna subtyping in vulvar lesions of premenarchal girls AB - Background: In premenarchal children the diagnosis of vulvar condyloma is often based on the clinical or microscopic appearance of the lesions. Other techniques for diagnosing condyloma such as DNA subtyping are not always used by providers. The purpose of this study is to compare the microscopic examination and Human Papilloma Virus (HPV) DNA subtyping of vulvar specimens from premenarchal girls to determine whether DNA subtyping aids in the diagnosis process.Methods: Eleven premenarchal girls were taken to the operating room for the treatment of clinically diagnosed condyloma between 1993 and 1999 at the University of Michigan Medical Center. In all patients, tissue was sent for pathologic evaluation and in 10 patients the specimens also underwent DNA subtyping. One patient had prior DNA subtyping. All the other lesions were surgically ablated. Results: The average age of our patients at the time of surgery was 2.3 years, range 1-6 years. Four patients had prior surgical treatment and two patients had undergone prior medical treatment. The microscopic diagnosis was condyloma in 8 patients, squamous papilloma with focal koilocytosis not totally diagnostic for condyloma in 1 patient, chronic inflammatory infiltrate in 1 patient, and 1 patient had papillary squamous hyperplasia with no koilocytosis.All 11 specimens tested positive for HPV DNA, 10 specimens contained at least one of the low risk subtypes (6, 11, 42, 43, 44) and 1 tests positive for low risk as well as high risk HPV type (16, 18, 31, 33, 35, 45, 51, 52, 56).Conclusion: Although all our patients with a clinical diagnosis of condyloma tested positive for HPV DNA, only 8 of 11 were definitely diagnosed with condyloma by microscopic examination. We therefore suggest that in premenarchal patients with verrucous lesions in the anogenital area HPV DNA subtyping be considered to avoid confusion with the diagnosis. PMID- 10869996 TI - What is the influence of self-image and perceived parenting role expectations on adolescent fathers' perceived role performance? AB - Background: "Adolescent pregnancy is one of the most pressing, persistent, and poignant problems facing society" (Yoos, 1987, p. 247). Manitoba's teen pregnancy rates are among the highest in Canada. Yet, little is known about adolescent fathers and their parenting involvement. The purpose of this descriptive correlational study was to explore variables which may influence teen fathers' participation in parenting.Methods: A convenience sample of 30 adolescent fathers, whose partners were attending an Adolescent Prenatal Clinic, completed two questionnaires: Offer Self-Image-Revised, and Perceived Parenting Role Performance. Guided by family role theory, four hypotheses were examined utilizing a quantitative research method.Results: Data analysis revealed that 30% of these respondents had a low to very low self-image. Pearson's correlation coefficient, which facilitated hypotheses testing, failed to validate a relationship between teen fathers' perceived role performance and self-image, and perceived parenting role expectations. Nevertheless, a moderate negative correlation was noted between teen fathers' self-image and their perceived parenting role expectations (r = -.35, p <.05).Conclusions: These findings contradict the philosophy of family role theory. Results of this study indicated that teen fathers want to be involved in parenting. However, unrealistic expectations and the inability to combine the developmental tasks of adolescence with the responsibilities of fatherhood increase their vulnerability to parenting failure. PMID- 10869997 TI - Unusual presentation of lichen sclerosuis in an adolescent AB - Background: The presentation of lichen sclerosus has been described in detail in the adult literature. Typically present with symptoms of itching and soreness in the vulvar area at which time a vulvar evaluation reveals a specific appearance. The presentation is believed to be similar in prepubertal children and adolescents. In this case report we encountered an unusual initial presentation of this disease.Methods: Case presentation.Results: An 18-year-old female presented for the first time to her gynecologist with complaint of difficulty with complete emptying of bladder and dribbling. She had noted the onset of these symptoms two months prior to presentation. She denied any long-term history of vulvar itching or irritation. Her menses were normal with no complaints of dysmenorrhea. Onset of menarche and pubertal development were also normal. She denied any pre-pubertal history of labial adhesions or lichen sclerosis. The patient was not sexually active. She was diagnosed with labial adhesions and her first course of treatment included topical estrogen therapy for 8 weeks. Her second course of therapy included topical testosterone for 6 weeks without any improvement or side effects. On evaluation at our institution the posterior aspect of the labia minora could not be seen and the area of the vaginal introitus was completely obstructed (see picture). The clitoral hood could not be retracted and the surrounding vulva appeared atrophic and white. The degree of obstruction was such that the urethra could not be seen. In the operating room the labia minora were manually separated. The patient applied clobetasol.05% ointment for the next two weeks to the vulva and then switched to a less potent steroid. Follow-up evaluation 2 and 4 weeks after the procedure did not show any adhesions. Punch biopsy was consistent with diagnosis of lichen sclerosis.Conclusion: The presentation of lichen sclerosis may be variable in adolescents; thus, a high index of suspicion must be maintained to make this diagnosis. PMID- 10869998 TI - A new technique for the creation of a neovagina AB - Background: There are many described reconstructive techniques for vaginal agenesis including vaginal dilators, skin covered molds, sigmoid grafts, vulval and large muscle flaps all of which aim to produce a vagina of normal axis, secretory capacity and length. We report the laparoscopic approach to Davydov's operation which utilizes peritoneum to line the newly dissected vesicorectal space. Methods: A case report detailing preoperative evaluation, surgical technique and outcome.Results: There were no intraoperative or immediate postoperative complications. The patient was discharged from hospital within 23 hours of surgery. Six month follow up revealed a vagina 7-8 cm in length, lined with squamous epithelium. The patient reports satisfactory sexual intercourse.Conclusion: This technique provides a satisfactory option for the surgical management of vaginal agenesis. PMID- 10869999 TI - Present status of spontaneous pneumothorax in Japan. AB - Today, spontaneous pneumothorax (SPT) is a common disease in Japan. It is easy to diagnose but difficult to estimate how to manage it. The curative treatment of SPT is resection of the ruptured bulla. In Japan, almost all surgical cases of SPT are operated by video-assisted thoracic surgery (VATS). The recurrence rate after VATS is only a few percent in our center. The cause of recurrence is usually attributable to overlooking bullae and newly developed bullae. Newly developed devices in Japan which help to reduce the recurrence rate are presented, and the Japan Association for Pneumothorax (JASP) and the Pneumothorax Center are introduced. PMID- 10870000 TI - Bronchogenic carcinoma in patients age 30 and younger. AB - BACKGROUND: Lung cancer is rare in patients 30 years of age or younger. There is very little published data on lung cancer in this group of patients. METHODS: A retrospective review of patients 30 years of age and younger with bronchogenic carcinoma treated at Roswell Park Cancer Institute between 1973 and 1994 was done. RESULTS: There were 20 patients (11 female and 9 male). Mean age was 27 years (range, 19-30). The predominant histologic types were adenocarcinoma in 11 patients (55%), and undifferentiated large-cell carcinoma in 5 patients (25%). All patients presented with either stage III (8 patients) or IV disease (12 patients). Eight patients (40%) underwent surgical resection (2 lobectomies, 6 pneumonectomies). Other treatments included chemotherapy in 15 patients (75%) and radiation therapy in 7 (35%). Median survival was only 5.5 months, and there were no 5-year survivors. Univariate analysis identified stage (p = 0.05), resection (p = 0.0005), and treatment with chemotherapy (p = 0.001) as predictors of survival. On multivariate analysis, resection (p = 0.0001) and chemotherapy (p = 0.001) remained as independent predictors of survival. CONCLUSIONS: Young patients with lung cancer present with advanced-stage disease and their cancers appear to be biologically aggressive. Although curative treatment is rarely possible, aggressive multimodality therapy is warranted. PMID- 10870001 TI - Intramyocardial electrogram recordings (IMEG) for diagnosis of cellular and humoral mediated cardiac allograft rejection. AB - OBJECTIVE: The purpose of this study was to prove the reliability of intramyocardial electrogram (IMEG) recordings for diagnosis and treatment monitoring of (1) cellular and (2) humoral mediated allograft rejection after heart transplantation. MATERIAL AND METHODS: Fifteen beagle dogs underwent heterotopic neck-heart transplantation. Eight of them were previously sensitized through several skin transplantations. IMEG recordings were performed daily. Donor-specific antibodies (IgG, IgM) were determinated in serum daily. Transmyocardial biopsies were performed every two days. RESULTS: In the sensitized group (group I) accelerated rejection occurred under triple drug immunosuppression with cyclosporine A, azathioprine, and cortisone on the fifth postoperative day (range: 4th-5th). All episodes were detected through IMEG diagnosis. In each case rejection could be treated successfully. In the cellular mediated group (group II), the average sensitivity for rejection diagnosis of a single lead was 24% for the unipolar and 42% for the bipolar leads. When the voltages of different leads were summed up the sensitivity rose to 36% (3 unipolar), 81% (3 bipolar) and 100% (all leads). During rejection therapy the IMEG recovered within 24-48 hours. CONCLUSION: The IMEG detects cellular and humoral mediated rejection early and with high reliability. The rejection-related changes of grade 2/3a rejection in IMEG seem to follow a Ofocal patternO similar to the histology. Therefore the recording of several, preferably bipolar, electrode configurations appears to enhance diagnostic reliability. PMID- 10870002 TI - Experiential assessment of endoventricular circular patch plasty in patients with depressed left ventricular function due to ischemia. AB - A concept of volume reduction surgery for patients with end-stage dilated cardiomyopathy was introduced to the surgical aspect. We started consequently to employ this operation for patients with ischemic cadiomyopathy. Between October 1997 and April 1999, 14 patients (mean age 64 years; 12 men, 2 women) underwent endoventricular circular patch plasty (EVCPP) in our institute. TECHNIQUE: The left ventricle (LV) is opened by placing an incision parallel to the left anterior descending (LAD) coronary artery. An endocardial purse-string suture is placed circumferentially at the root of the posterior papillary muscle. The remaining opening of the cavity is closed with a circular bovine pericardial patch. The excluded edges of LV free wall and septum are directly closed with felt strips. RESULTS: There were no significant hemodynamic differences in heart rate, cardiac index and pulmonary capillary wedge pressure before and after the operation. Either LV end-diastolic volume index (from 134.8 +/- 30.7 to 77.1 +/- 16.3 ml/m2) or LV end-systolic volume index (from 85.0 +/- 21.8 to 40.0 +/- 12.5 ml/m2) definitely reduced postoperatively, the LV ejection fraction (EF), consequently, significantly improved from 0.31 +/- 0. 09 to 0.48 +/- 0.10 (p=0.0007). In all patients the New York Heart Association (NYHA) functional class improved from 2.8 +/- 0.7 to 1.3 +/- 0.5 after operation. We will aggressively employ this procedure for patients with ischemic cardiomyopathy in consideration of the limitations of the efficacy and the indication of this operation. PMID- 10870003 TI - Cardiac operations in patients with severe pulmonary impairment. AB - Many reviews concerning pulmonary complications after cardiac surgical procedures in patients with serious pulmonary disease have been published. However, no strict pulmonary function guidelines were proposed to help the clinician identify the patients at greater risk. We considered whether a low pulmonary function became a risk factor of cardiac operations. We conducted a retrospective analysis of records of 32 patients with severely impaired preoperative pulmonary function who had undergone cardiac operations between July 1988 and March 1999. There was 1 hospital death. The over-all mortality rate was 3.1% (1 of 32). However, this death could not be directly attributed to postoperative pulmonary complications. Postoperative pulmonary complications were seen in 2 patients (6.3%) who required tracheostomy due to atelectasis and pneumonia. No late deaths due to pulmonary complications were observed during the follow-up period. The actual survival rate is 68% at 7 years. A low pulmonary function did not, by itself, become a risk factor of cardiac operations, although a pulmonary function test can be used to alert the clinician to possible postoperative complications, including the requirement of tracheostomy. Especially strict control of postoperative respiration is necessary in patients with forced expiratory volume (FEV) of 1.0 <= 800 ml and/or FEV1.0/BSA <= 600 ml/m2. PMID- 10870005 TI - Off-pump coronary artery bypass: early results. AB - BACKGROUND: Coronary artery bypass grafting (CABG) on a beating-heart has gained the attention of cardiac surgeons and shown favorable initial results. However, only a few follow-up results have been reported. We report herein our one-year experiences of off-pump CABG performed at Shin-Tokyo Hospital. METHODS: Retrospective chart review was performed for patients who underwent off-pump CABG and conventional isolated CABG between 01/01/98 and 12/31/98. Preoperative, perioperative, and follow-up data were collected. RESULTS: Among 315 cases of isolated CABG, 94 cases were off-pump CABG (male/female 69/25, mean age 67.7). Mean number of distal anastomoses performed by off-pump CABG was 1.7 +/- 0.7 (42 cases of single-vessel revascularization and 52 cases of more than double- vessel revascularization). In off-pump CABG, there were no hospital deaths and 6 major complications including 2 incidences of perioperative myocardial infarction. Postoperative angiography before hospital discharge was performed in 56 patients (59.6%, 98 anastomosis) and revealed 5 occlusions, giving a graft patency rate of 94.9%. During the follow-up (11.4 +/- 4.1 months), there was 1 late non-cardiac death and 11 cardiac events. The event-free rate at 18 months was 94.0% in off pump CABG, showing no significant difference from the event-free rate after conventional CABG (94.0% at 18 months, p = 0.135). Follow-up angiography was performed in 21 patients (33 anastomoses) at a mean interval of 3.6 months and showed 4 graft occlusions, giving a patency rate of 92.7%. CONCLUSION: Both hospital and early results of off-pump CABG were acceptable. Off-pump CABG can be safely performed in selected patients. PMID- 10870004 TI - Effects of docarpamine on hemodynamics after open heart surgery in children. AB - Docarpamine is a dopamine prodrug which has been selected from a large number of dopamine derivatives in order to develop an orally effective dopamine. The pharmacokinetics after oral administration of docarpamine have not yet been studied in children undergoing open heart surgery. This study examined the effects of docarpamine on hemodynamics and evaluated its safety in 11 children undergoing open heart surgery for congenital heart disease. This study began when the patientOs postoperative condition was stabilized by continuous dopamine infusion into the vein at a rate of 5 micro g/kg/min. The patients were administered 40 mg/kg of docarpamine every 8 hours, and hemodynamics were measured every 4 hours for 16 hours after the initial docarpamine administration. Immediately after the initial docarpamine administration, the dose of dopamine was reduced to 3 micro g/kg/min. Infusion of dopamine was stopped 8 hours after the initial docarpamine administration. Systemic systolic and diastolic blood pressure and heart rate showed no significant changes. Mean right atrial pressure decreased 4 hours after docarpamine administration. Mixed venous oxygen saturation and mean velocity of circumferential fiber shortening increased significantly after docarpamine administration. No significant changes were observed in urine volume. All patients could be weaned from dopamine within 8 hours. No changes were observed in ECG, and no arrhythmia-inducing action was noted. Our study indicates that 40 mg/kg oral doses of docarpamine produce plasma dopamine concentration equivalent to those of a 3 to 5 micro g/kg/min dopamine infusion. Our data suggest that docarpamine is a safe and effective drug for children who have undergone open heart surgery. PMID- 10870006 TI - Intrapulmonary sequestration with arterial supply from the left internal thoracic artery: a case report. AB - Pulmonary sequestration is uncommon in the upper lobe. Its arterial supply from the internal thoracic artery is very rare. Reported here is a case of a 20-year old male whose presenting symptom was recurrent pneumonia. Helical computed tomography (CT) and three-dimensional reconstruction images showed that aberrant arteries arising from the left internal thoracic artery were supplying the area of sequestration and draining into the pulmonary vein. Selective intra-arterial digital substraction angiogram also showed left internal thoracic artery supplying the area of the sequestration. Helical three-dimensional CT is noninvasive and provides as accurate three-dimensional information of the aberrant vascular supply in intrapulmonary sequestration as the angiography. PMID- 10870007 TI - A rare case of lung cancer associated with renal cell cancer and benign histiocytoma of bone. AB - Lung cancer often metastasizes to organs outside the thorax, and consequently radiological evaluation of distant metastasis has become standard procedure prior to surgery. Although positive radiological findings generally suggest distant metastasis, the possibility of the co-existence of a benign tumor and primary malignancies must be considered. Herein we report a case of surgical resection of histologically confirmed lung cancer associated with renal cell cancer and benign histiocytoma of the humerus. PMID- 10870008 TI - Double simultaneous cysts of the mediastinum. AB - We describe a rare case of double mediastinal tumors in a 60-year-old male with spinocerebellar degeneration. Magnetic resonance imaging (MRI) accidentally revealed double cystic tumors in the anterior and posterior mediastinum. Surgical management by video-assisted thoracic surgery (VATS) was successfully performed. The histological diagnoses were confirmed as a thymic cyst in the anterior and a thoracic duct cyst in the posterior mediastinum, respectively. PMID- 10870009 TI - Pathological findings of tissue reactivity of gelatin resorcin formalin glue: an autopsy case report of the repair of ventricular septal perforation. AB - A seventy-three-year-old man was treated for ventricular septal perforation with Gelatin Resorcin Formalin (GRF) glue. The patient died of multiple organ failure 36 days after the surgery. In autopsy, macroscopically, the inferior wall was reconstructed successfully by the GRF glue. Furthermore, microscopic study revealed the excellent growth of collagen and elastic fiber where the GRF was glued. No infiltration of inflammatory cells was evident. There have been no reports that the safety and efficacy of GRF glue was pathologically proven in an autopsy case. PMID- 10870011 TI - Minimally invasive direct coronary artery bypass grafting using the saphenous vein in redo CABG. AB - We describe a patient who underwent minimally invasive direct coronary artery bypass (MIDCAB), who had previously undergone coronary artery bypass grafting (CABG) through a median sternotomy with a left internal mammary artery (LIMA) graft to the left anterior descending artery (LAD) and a right gastroepiploic artery (GEA) graft to the posterior descending artery. MIDCAB was less invasive and was an effective alternative procedure for the second operation. Because the patient had no LIMA or GEA available for a graft because of prior use, we used a saphenous vein graft (SVG) for bypassing from the left subclavian artery to the coronary artery by MIDCAB via a left minithoracotomy. The left subclavian artery was selected as the proximal anastomotic site because this artery was less diseased and was easier to reach. The SVG-to-coronary artery anastomosis was facilitated by firm adhesion between the epicardium and the pericardium, which reduced the motion of the epicardium itself. These results suggest that the procedure is safe and promising in selected cases of redo CABG. PMID- 10870010 TI - A case report of surgical treatment of quadricuspid aortic valve associated with regurgitation. AB - A case of a 65-year-old woman who had a quadricuspid aortic valve associated with aortic regurgitation is reported. The patient had severe aortic regurgitation and four equally divided aortic cusps. The valve abnormality was detected by a transesophageal echo and an aortography. The incomplete aortic valve was excised and replaced by a St. Jude Medical prosthesis. Although this case had no coronary abnormality, a coronary displacement is often reported in quadricuspid aortic valve cases. In order to perform an operation safely, accurate information which is obtained by a non-invasive examination of the transesoph-ageal echo is quite valuable as it can indicate the need for further preoperative examinations of the coronary arteries. PMID- 10870012 TI - Self-expandable vascular stent covered with polyurethane membrane: an experimental preliminary study of a placement in the thoracic descending aorta of a rabbit using endovascular intervention. AB - The effectiveness of self-expandable vascular endoprosthesis covered with polyurethane membrane for arterial substitution was examined in the descending thoracic aorta of a rabbit, followed by an observation period of 937 days. In this model there was no evidence of thrombus, aneurysmal formation, and/or infection. The self-expandable vascular stent covered with a polyurethane membrane showed long-term patency as well as excellent function, and the histological evaluation revealed endothelial cells covering all of the surface of the endoprosthesis, as was expected. Minimal intimal hyperplasia and no calcifica tion were demonstrated in any portions. This study suggests that our newly designed self-expandable vascular stent covered with a polyurethane membrane could serve as a satisfactory vascular endoprosthesis with a good long-term patency for substitution. Furthermore, stenting using our model is a safe, simple technique, and an effective treatment for vascular remodeling. PMID- 10870013 TI - Who is being served? PMID- 10870014 TI - Knowledge-work: building and modeling PMID- 10870015 TI - What is happening to advocacy? PMID- 10870016 TI - How the ordinary becomes extraordinary. PMID- 10870017 TI - Nurse practitioner education in the new millennium: challenges and opportunities. PMID- 10870018 TI - Nursing's perspective on improving communication about nursing home residents' preferences for medical treatments at end of life. PMID- 10870019 TI - Nursing perspectives on practitioner-assisted suicide. PMID- 10870020 TI - Minority nurses in leadership positions: a call for action. PMID- 10870021 TI - Cultural influence on nursing scholarship and education. PMID- 10870022 TI - Utility of structured care approaches in education and clinical practice. PMID- 10870023 TI - Comments on the Secretary's Advisory Committee on Genetic Testing document: a public consultation on oversight of genetic tests, January 27, 2000, Baltimore, Maryland. PMID- 10870024 TI - Notes from the executive director PMID- 10870025 TI - Novel insights into the neuroendocrinology of critical illness. AB - An unexplained hallmark of prolonged critical illness is the fact that food does not prevent or reverse protein wasting, while fat is paradoxically accrued. This 'wasting syndrome' often persists after the underlying disease has been resolved and thus perpetuates intensive care dependency. Although the crucial role of an intact hypothalamus-pituitary axis for homeostasis during stress is well recognized, the differences between the neuroendocrine changes observed in acute and prolonged critical illness were only recently described. Novel insights in this area are reviewed here. The initial endocrine stress response consists primarily of a peripheral inactivation of anabolic pathways while pituitary activity is essentially amplified or maintained. These responses presumably provide the metabolic substrates and host defense required for survival and to delay anabolism, and thus should be considered as adaptive and beneficial. Persistence of this acute stress response throughout the course of critical illness was hitherto assumed. This assumption has now been invalidated, since a uniformly reduced pulsatile secretion of ACTH, TSH, LH, prolactin (PRL) and GH has been observed in protracted critical illness, causing diminished stimulation of several target organs. Impaired pulsatile secretion of anterior pituitary hormones in the chronic phase of critical illness seems to have a hypothalamic rather than a pituitary origin, as administration of relevant releasing factors evoked immediate and pronounced pituitary hormone release. A reduced availability of TRH, one of the endogenous ligands of the GH-releasing peptide (GHRP) receptor (such as the recently discovered ghrelin) and, in very long-stay critically ill men, also of GHRH, appear to be involved. This hypothesis was further explored by investigating the effects of continuous i.v. infusion of GHRH, GHRP, TRH and their combinations for several days. Pulsatile secretion of GH, TSH and PRL was re-amplified by relevant combinations of releasing factors which also substantially increased circulating levels of IGF-I, GH-dependent binding proteins, thyroxine and tri-iodothyronine (T3) while avoiding a rise in reverse T3. Active feedback-inhibition loops prevented overstimulation of target organs and metabolic improvement was noted with the combined infusion of GHRP and TRH. Whether this novel endocrine strategy will also enhance clinical recovery from critical illness remains to be explored. PMID- 10870026 TI - Innovative strategies for the treatment of thyroid cancer. AB - Normally, thyroid cancer is a disease with a good prognosis, but about 30% of the tumours dedifferentiate and may finally develop into highly malignant anaplastic thyroid carcinomas with a mean survival time of less than 8 months. Due to the loss of thyroid-specific functions associated with dedifferentiation, these tumours are inaccessible to standard therapeutic procedures such as radioiodide therapy and thyroxine-mediated thyrotrophin suppression. Medullary thyroid carcinomas are also highly aggressive. Here, therapy is limited to surgery, and no alternative is left if patients do not respond to this standard procedure. Obviously, new approaches would be desirable. Several novel approaches are currently being tested for the treatment of thyroid cancer. Many of them utilise methods of gene therapy, but follow different strategies: (1) reintroduction of the tumour suppressor p53 into a background lacking functional p53; (2) suicide gene therapy with ganciclovir and a transduced gene for herpes simplex virus thymidine kinase controlled by the thyroglobulin promoter; (3) strengthening of the antitumour immune response by expression of an adenovirus-delivered interleukin-2 (IL-2) gene; (4) induction of an immune response by DNA vaccination against the tumour marker calcitonin; (5) transduction of the thyroid sodium/iodide transporter gene to make tissues that do not accumulate iodide treatable by radioiodide therapy; (6) blocking of the expression of the oncogene c-myc by antisense oligonucleotides. While these approaches are still tested in vitro or in animal models, first results from pilot studies concerning other novel treatment modalities are available: (7) radioimmunotherapy exploits the carcinoembryonic antigen expressed on medullary thyroid carcinomas to target a radiolabelled antibody to the tumour; and (8) retinoic acid is used for a redifferentiation therapy in the case of thyroid cancer. Hopefully, one or the other of these novel strategies may probably extend after some time the current therapeutic repertoire for thyroid cancers and provide a perspective for otherwise untreatable patients. PMID- 10870027 TI - The human thyrotropin receptor is highly mutable: a review of gain-of-function mutations. AB - OBJECTIVES: To find whether germline and somatic gain-of-function mutations of the thyrotropin receptor (TSHR) differ in location and/or mutational mechanisms, as well as to explore the degree to which these mutations are specific to TSHR compared with pituitary glycoprotein hormone receptors. METHODS: We examined the data on the TSHR website (www.unvi-leipzeig approximately innerre/TSH) supplemented with recent literature. Comparisons were also made with gain-of function mutations of lutropin/choriogonadotropin (LH/CGR) and follicle stimulating hormone receptors (FSHR). RESULTS: Some mutations (at residues 183, 505, 509 and 597) are exclusively germline, whereas mutations at 630 and 633 are characteristic of somatic mutations. Several residues located mainly in a mutation cluster region (619-639) are shared by both. Germline mutations are more likely to be transitions than transversions compared with somatic mutations. The lack of mutations involving deamination of CpG dinucleotides, a common mechanism for C-->T transitions, reflects the low CG prevalence in the mutable regions of TSHR. Comparison of the mutation sites with the equivalent positions in LH/CGR showed a significant difference (P<0.0001), whereas those in the mutation cluster region comprising the sixth transmembrane helix (TM6) and the adjoining third intracellular loop were concordant (P>0.90). We suggest that there is specific clustering of mutations in the juxtacytoplasmic end of TM6 in LH/CGR, a hydrophobic patch that is tightly packed with a face on TM5 whose sequences diverge from those of TSHR. CONCLUSIONS: TSHR exhibited frequent mutations outside the mutation cluster region. A role for a mutagenic environment created by the thyroid for other TSHR-specific codons cannot be discounted, nor can genetic factors, when accounting for the variation in the prevalence of TSHR activating mutations worldwide. PMID- 10870028 TI - Consistent production of a higher TH1:TH2 cytokine ratio by stimulated T cells in men compared with women. AB - OBJECTIVE: To evaluate the T helper 1 (T(H)1)/T helper 2 (T(H)2) lymphocyte cytokine profiles in women and men and to study the in vitro effects of sex hormones on lymphocyte secretion of cytokines. METHODS: Analysis of serum concentration and lymphocyte synthesis of T(H)1 (gamma interferon (INF-gamma) and interleukin 2 (IL-2)) and T(H)2 (interleukin 4 (IL-4) and interleukin 10 (IL-10)) cytokines was performed in 20 women and 15 men. Analysis of modifications in cytokine secretion induced by supplementation of lymphocyte culture with increasing concentrations of sex hormones was carried out. RESULTS: Higher levels of INF-gamma and IL-2 and lower levels of IL-4 and IL-10 were detected in the phytohemagglutinin-stimulated lymphocyte culture supernatants of men compared with women; the INF-gamma:IL-4 ratio was significantly higher in men. In women, similar concentrations of all the cytokines were detected in culture supernatants obtained during the follicular and the luteal phases. The addition of sex hormones did not modify the concentration of cytokines in supernatants of phytohemagglutinin-stimulated T-cell cultures. CONCLUSIONS: Women present a predominant T(H)2 cytokine profile, which could be involved in immune responses characterized principally by the secretion of antibodies. This could be a factor implicated in the higher concentration of immunoglobulins or the increased prevalence of autoimmune diseases detected in females. PMID- 10870029 TI - Thyroid ultrasound is the best prevalence indicator for assessment of iodine deficiency disorders: a study in rural/tribal schoolchildren from Gujarat (Western India). AB - OBJECTIVES: (i) To assess the severity of iodine deficiency disorders (IDD), (ii) to determine the aetiology of IDD in Gujarat, (iii) to identify the best prevalence indicator of IDD, and (iv) to compare thyroid volume (TV) results with the WHO International reference. METHODS: Five hundred and thirty schoolchildren (6-15 years) were studied from two districts (Baroda and Dang) and data were collected on dietary habits and parameters such as height, weight, thyroid size by palpation and ultrasonography, urinary iodine (UI), and blood thyroid stimulating hormone (TSH). Drinking water was analyzed for iodine content and food articles for goitrogens. RESULTS: In Gujarat children median UI (interquartile range)=56 (30-96)microg/l, mean TSH=1.71 +/- 2.10mU/l, goiter by palpatio n = 30%, and median TV = 27.8 (23-35)ml. Females had lower median UI (48 (27-82) microg/l) and higher mean TSH levels (2.0 +/- 2.5mU/l) than males. Applying the WHO ultrasonography reference to Gujarat children resulted in an enlarged TV-for-body surface area in almost 100% of subjects. Ninety-nine percent of females and 95% of males had enlarged TV-for-age. Three to eight times larger TV were seen in all subjects as compared with European children. Dang children were severely malnourished. Flavonoids like vitexin, glucosyl vitexin and apigenin were detected in pearl millet. Apigenin was never identified in pearl millet. Dang district water was lacking in iodine content. CONCLUSIONS: IDD is a severe public health problem in Gujarat. Baroda district is a new pocket of IDD. High amounts of dietary flavonoids in Baroda and Dang districts, and lack of iodine in Dang water, account for IDD. TV measurement by ultrasound is the best prevalence indicator of IDD. PMID- 10870030 TI - Frequency of somatic MEN1 gene mutations in monoclonal parathyroid tumours of patients with primary hyperparathyroidism. AB - OBJECTIVES: Investigation of small numbers of parathyroid tumours by X-chromosome inactivation analysis suggests that the majority of them are monoclonal lesions most likely caused by a somatic mutation. Somatic mutations in the MEN1 gene located on chromosome 11q13 have recently been identified in 12-17% of solitary parathyroid tumours in patients with sporadic primary hyperparathyroidism, and they may be the precipitating genetic defect leading to monoclonal cell proliferation in these tumours. DESIGN: To determine the prevalence of MEN1 gene mutations in monoclonal parathyroid neoplasias we investigated 33 parathyroid tumours of patients with primary hyperparathyroidism for clonality and mutations in the MEN1 gene. METHODS: X-chromosome inactivation analysis was used to assess the clonal status of the tumours, direct sequencing of the complete coding region was applied to identify mutations in the MEN1 gene. RESULTS: Twenty-eight female patients (26 patients with solitary adenoma, 2 patients with hyperplasia) were informative for the polymorphism of the androgen receptor on the X-chromosome and could be tested for inactivation pattern. Nineteen of twenty-six (73%) solitary adenomas were monoclonal. Somatic mutations in the MEN1 gene were identified in nine cases. Six of them were found in the relatively large second exon of the MEN1 gene (A49D, 193del36, 402delC, 482del22, 547delT, W126X). One was found in exon 5 (904del9), one in exon 7 (Y327X) and one in exon 9 (R415X). Of the monoclonal tumours, 5 out of 19 (26%) harboured a somatic MEN1 gene mutation. CONCLUSIONS: In summary, 73% of the solitary parathyroid adenomas were monoclonal. In 26% of the monoclonal tumours a somatic MEN1 gene mutation has been identified. However, for 74% of monoclonal tumours of the parathyroids the underlying genetic defects are still not known. PMID- 10870031 TI - The influence of menopause and body mass index on serum leptin concentrations. AB - OBJECTIVE: The aim of this study was to evaluate the influence of menopausal status, serum estradiol and body mass index (BMI) on serum leptin concentration in a large sample of pre- and postmenopausal women. DESIGN: 434 healthy women (mean age +/-s.d., 52.2 +/- 10.3 years) were recruited at the University of Marburg on the occasion of a routine gynecological visit. Two hundred and eighteen (50.2%) women were premenopausal (mean age, 36.5 +/- 10.4 years) and not on oral contraceptives or hormone replacement therapy (HRT) and 216 (49.8%) women were postmenopausal (mean age 61.8 +/- 8.9 years) not on HRT. To evaluate the influence of menopausal status, estradiol level and BMI on serum leptin concentrations, women were allocated to one of the four groups: (a) premenopausal women BMI <25 kg/m(2) (n=137), (b) premenopausal women BMI >25 kg/m(2) (n=81), (c) postmenopausal women BMI <25 kg/m(2) (n=94) and (d) postmenopausal women BMI >25 kg/m(2) (n=122). RESULTS: Irrespective of the menopausal status, women with a BMI >25 kg/m(2) had significantly higher leptin concentrations in all age groups compared with women with a BMI <25 kg/m(2) (P<0.001). The multiple linear regression analyses showed that BMI was the only statistically significant independent predictor for leptin. In comparison to postmenopausal women, premenopausal women showed a significantly lower mean age, weight, BMI and FSH concentration (P<0. 001), a higher mean height and serum estradiol (P<0.01 and P<0.001 respectively) but significantly lower serum leptin concentration (P<0.01). The multiple linear regression model showed no significant influence of menopausal status or serum estradiol on serum leptin concentration, even after controlling for BMI. CONCLUSIONS: Serum leptin concentrations are significantly higher in pre- and postmenopausal obese women, compared with normal weight controls. Serum leptin concentrations are not influenced by menopausal status or serum estradiol level. PMID- 10870032 TI - Effect of GH and/or testosterone deficiency on the prostate: an ultrasonographic and endocrine study in GH-deficient adult patients. AB - BACKGROUND: The role of IGF-I in prostate development is currently under thorough investigation since it has been claimed that IGF-I is a positive predictor of prostate cancer. OBJECTIVE: To investigate the effect of chronic GH and IGF-I deficiency alone or associated with testosterone deficiency on prostate pathophysiology in a series of patients with hypopituitarism. DESIGN: Pituitary, androgen and prostate hormonal assessments and transrectal prostate ultrasonography (TRUS) were performed in 30 men with adulthood onset GH deficiency (GHD) and 30 age-matched healthy controls, free from previous or concomitant prostate disorders. RESULTS: Plasma IGF-I levels were significantly lower in GHD patients than in controls (Pearson's coefficient P<0.0001). At study entry, 6 of the 13 hypogonadal patients and 7 of the 17 eugonadal patients had plasma IGF-I below the age-adjusted normal range. At study entry, testosterone levels were low in 13 patients (mean +/-s.e.m., 3.8+/-1.0 nmol/l) while they were normal in the remaining 17 (19.4+/-1.4 nmol/l). No difference in prostate specific antigen (PSA), and PSA density was found between GHD patients (either hypo- or eugonadal) and controls, while free PSA levels were significantly higher in eugonadal GHD than in controls (0.4+/-0.04 vs 0.2+/-0.03 microg/l; P<0.01). No difference in antero-posterior prostate diameter and transitional zone volume (TZV) was observed among groups, while both transverse and cranio-caudal diameters were significantly lower in hypogonadal (P<0.01) and eugonadal GHD patients (P<0.05) than in controls. Prostate volume (PV) was significantly lower in hypogonadal GHD patients (18.2+/-3.0 ml) and eugonadal GHD patients (22.3+/ 1.6 ml), than in controls (25.7+/-1.4, P<0.05). The prevalence of prostate hyperplasia (PV>30 ml) was significantly lower in hypogonadal and eugonadal GHD patients, without any difference between them (15.3% and 5.8%), than in controls (43.3%) (chi(2)=6.90, P=0.005). No difference was found in PV between patients with normal or deficient IGF-I levels both in the hypogonadal group (19. 9+/-4.7 vs 17.3+/-4.0 ml) and in the eugonadal group (22.6+/-2.3 vs 21.8+/-2.5 ml). When controls and patients were divided according to age (<60 years and >60 years), PV was significantly lower in hypogonadal GHD patients aged below 60 years than in age-matched controls (P<0.01) or eugonadal GHD patients (P<0.01), without any difference between controls and eugonadal GHD patients. Controls aged above 60 years had significantly higher PV than both hypogonadal and eugonadal GHD patients (P<0.01). Calcifications, cysts or nodules were found in 56.7% of patients and in 50% of controls (chi(2)=0.067, P=0.79). In controls, but not in GHD patients, PV and TZV were correlated with age (r=0.82, r=0.46, P<0. 0001 and P<0.01 respectively). PV was also correlated with GH (r=-0. 52, P=0.0026), IGF-I (r=-0.62, P=0.0002) and IGF-binding protein 3 (IGFBP-3) levels (r=-0.39, P=0.032) but neither with testosterone or dihydrotestosterone (DHT) levels. In GHD patients TZV but not PV was correlated with age (r=0.58, P=0.0007) and neither TZV nor PV were correlated with GH, IGF-I or IGFBP-3 levels. CONCLUSIONS: Chronic GH deficiency in adulthood causes a decrease in prostate size, mostly in patients with concomitant androgen deficiency and age below 60 years, without significant changes in the prevalence of structural prostate abnormalities. PMID- 10870033 TI - Characterisation of novel missense mutations in the GH receptor gene causing severe growth retardation. AB - Two Swedish brothers, 2.5 and 4 years of age, were found to fulfil all the clinical and laboratory characteristics of Laron's syndrome. They were shown to have unique missense mutations in the GH receptor gene. Both of their parents were of normal height, but they both separately carried one of the identified gene alterations. A molecular model of the first receptor alteration suggests that a collapse in three-dimensional receptor structure most likely contributed to the GH insensitivity in these patients. PMID- 10870034 TI - Inverse correlation between baseline inhibin B and FSH after stimulation with GnRH: a study of serum levels of inhibin A and B, pro alpha-C and activin A in women with ovulatory disturbances before and after stimulation with GnRH. AB - OBJECTIVE: Interest has focused recently on the influences of the polypeptide factors inhibin and activin on the selective regulation of the pituitary secretion of gonadotropins. DESIGN: Measurement of the concentrations of inhibin related proteins in relation to the changes in pituitary gonadotropin (FSH, LH) parameters, after GnRH stimulation with a bolus injection of 100 microg gonadorelin, in 19 women with ovulatory disturbances. METHODS: Serum levels of inhibin A and B, activin A, and pro alpha-C were measured using sensitive ELISA kits. RESULTS: Within 60 min after GnRH stimulation, FSH values doubled from 5 to 10 mU/ml (P < 0.001). LH increased 12-fold from 2 to 24 mU/ml (P < 0.001). Activin A showed a significant decrease from 0.47 to 0.36 ng/ml (P < 0.001), whereas pro alpha-C increased from 127 to 156 pg/ml (P = 0.039). The median inhibin A concentration did not show a significant change between baseline and the 60 min value, whereas inhibin B was characterized by a minor, but not significant, increase in the median from 168 to 179 pg/ml (P = 0.408). A significant inverse correlation (P = 0.014) with a mean coefficient of correlation of 0.5516 was found, demonstrating a strong relationship between high inhibin B baseline levels and a small increase of FSH after 60 min. CONCLUSION: Our results show an interesting correlation between the baseline inhibin B and the change in FSH before and after GnRH stimulation. A high baseline inhibin B implies only a minor increase of FSH after 60 min. PMID- 10870035 TI - Sex hormone-binding globulin as a marker for hyperinsulinemia and/or insulin resistance in obese children. AB - OBJECTIVE: A relationship between hyperinsulinemia and decreased serum sex hormone-binding globulin (SHBG) has been described in adults. We evaluated the usefulness of SHBG as an index of hyperinsulinemia and/or insulin resistance in obese children (aged 6-9 years) of both sexes and its possible influence on the androgenic status. DESIGN: We carried out a cross-sectional study of cases and controls. We studied 61 obese children (22 males, 39 females) with body mass index (BMI) superior to the 90(th) percentile and a control group of age- and sex matched non-obese children. We measured serum glucose, insulin, TSH, free thyroxine, 17beta-estradiol, testosterone and SHBG. Also, we correlated these parameters with anthropometric measures. RESULTS: The obese group presented significantly elevated levels of insulin (P=0.001) and insulin/glucose ratio (P=0.0012) compared with the control group. SHBG (P=0.0001) and testosterone (P=0.0169) levels were significantly lower than those in the non-obese group. We did not find any difference in the free androgen index (FAI). Fasting insulin (r= 0.4512; P<0.001), BMI (r=-0.3185; P<0.05) and testosterone (r=-0.3705; P<0.01) were inversely correlated with SHBG concentration. According to multivariate analyses, insulin was the only independent predictor factor for serum SHBG concentration in the obese group (r partial=0.1280; P=0.0171). CONCLUSIONS: In summary, at this age there is a strong relationship between insulin and SHBG. The changes in SHBG levels of the obese group did not affect FAI and, therefore, they did not cause changes in the androgenic status. Our data support the role of insulin in the regulation of serum SHBG levels. PMID- 10870036 TI - Adrenocortical function in patients with macrometastases of the adrenal gland. AB - OBJECTIVE: Metastases of the adrenal gland are a frequent finding in patients with malignant tumors like bronchogenic carcinoma or breast cancer. Only limited and conflicting data on adrenocortical function in these patients are available. DESIGN: Cross-sectional study. METHODS: We investigated the impact of adrenal macrometastases on adrenocortical function in a series of 28 tumor patients using the ACTH(1-24) stimulation test and dexamethasone suppression test. Seven normal controls (Con), eleven patients without adrenal metastases (No Met), eight patients with unilateral (Uni Met) and nine patients with bilateral adrenal metastases (Bil Met) were investigated. RESULTS: The prevalence of adrenal insufficiency was low in our study population, with only two of nine patients with bilateral metastases having subclinical adrenocortical insufficiency. In the remaining patients with uni- or bilateral metastases, baseline and stimulated cortisol concentrations were higher than in controls and cancer patients without metastases (baseline cortisol (in nmol/l): Con: 307+/-33.2 vs Uni Met: 440+/ 53.5, and Bil Met: 637.6+/-92.1, P=0.04 by ANOVA; cortisol 60 min after ACTH(1 24): Con: 794.6+/-41.2 vs Uni Met: 990.8+/-92.9, and Bil Met: 1151.4+/-155.5, P=0.03 by ANOVA). Simultaneously, baseline and stimulated serum aldosterone concentrations were significantly blunted in the tumor groups. CONCLUSIONS: Adrenal insufficiency is infrequent and develops only in patients with bilateral metastases. However, the majority of patients have activation of the hypothalamic pituitary-adrenal axis despite adrenal metastases with strongly elevated cortisol concentrations. PMID- 10870037 TI - Interaction between glucagon and human corticotropin-releasing hormone or vasopressin on ACTH and cortisol secretion in humans. AB - OBJECTIVE: It is known that glucagon administration elicits ACTH and cortisol responses in humans, although this effect takes place after intramuscular or subcutaneous but not after the intravenous route of administration. The mechanisms underlying this stimulatory effect on corticotroph secretion are unknown but they are unrelated to glucose variations and stress-mediated actions. DESIGN AND METHODS: To throw further light on the stimulatory effect of i.m. glucagon on the pituitary-adrenal axis, using six normal young female volunteers (26-32 years, body mass index 19.7-22.5 kg/m(2)) we studied the interaction between glucagon (GLU; 0.017 mg/kg i.m.) and human corticotropin-releasing hormone (hCRH; 2.0 microg/kg i.v.) or vasopressin (AVP; 0.17 U/kg i.m.). The interactions between hCRH and AVP on the hypothalamo-pituitary-adrenal (HPA) axis and the GH response to GLU alone or combined with hCRH or AVP were also studied. RESULTS: GLU i.m. administration elicited a clear increase in ACTH (peak vs baseline, means+/-s.e.m.: 11.6+/-3.3 vs 4.2+/-0.3 pmol/l, P<0.05), cortisol (613.5+/-65.6 vs 436.9+/-19.3 nmol/l, P<0.05) and GH levels (11.6+/-3.4 vs 3.3+/ 0.7 microg/l, P<0.05). The ACTH response to GLU (area under the curve: 426.4+/ 80.9 pmol/l per 120 min) was higher than that to AVP (206.3+/-38.8 pmol/l per 120 min, P<0.02) and that to hCRH (299.8+/-39.8 pmol/l per 120 min) although this latter difference did not attain statistical significance. The GLU-induced cortisol response (28336.9+/-2430.7 nmol/l per 120 min) was similar to those after hCRH (24099.2+/-2075.2 nmol/l per 120 min) and AVP (21808.7+/-1948.2 nmol/l per 120 min). GLU and hCRH had an additive effect on ACTH (964.9+/-106.6 pmol/l per 120 min, P<0.02) and a less than additive effect on cortisol levels (35542.5+/-2720. 2 nmol/l per 120 min). Similarly, GLU and AVP had an additive effect on ACTH (825.6+/-139.6 pmol/l per 120 min, P<0.02) and an effect less than additive on cortisol levels (33059.2+/-1965.3 nmol/l per 120 min). The effects of GLU co-administered with hCRH or AVP were similar to those of the combined administration of hCRH and AVP on ACTH (906. 0+/-152.7 pmol/l per 120 min) and cortisol (34383.5+/-1669.2 nmol/l per 120min) levels. The GH response to GLU was not modified by hCRH or AVP. CONCLUSIONS: These results show that i.m. glucagon administration is a provocative stimulus of ACTH and cortisol secretion, at least as potent as hCRH and AVP. The ACTH-releasing effect of i.m. glucagon is not mediated by selective CRH or AVP stimulation but the possibility that both neurohormones play a role could be hypothesized. PMID- 10870038 TI - Combined clonidine-short-ACTH test for the simultaneous assessment of growth hormone reserve and hypothalamic-pituitary-adrenal axis integrity in children. AB - OBJECTIVE: To determine the feasibility of using the combined oral clonidine and the short-ACTH test instead of the sometimes dangerous insulin-induced hypoglycemia test as a screening procedure, for the simultaneous assessment of growth hormone reserve and hypothalamic-pituitary-adrenal axis integrity in children with growth retardation. DESIGN: Evaluative study. METHOD: Seventy-three children (52 males) aged 11+/-3 years with attenuated growth (group 1) were tested by combined clonidine (150 microg/m(2)) and short-ACTH test (either the standard 250 microg or the low-dose 1 microg/1. 73 m(2)). Thirty-one children received no pretreatment (nonprimed) (subgroup 1NP), and 42 were primed with ethynylestradiol 40 microg/m(2)/day two days before testing (subgroup 1P). The control group for the short-ACTH test (group 2) consisted of 42 children and adolescents (13 males) aged 12+/-3 years with early or accelerated puberty or premature closure of epiphyses, who received ACTH only (21 standard, 21 low-dose) with no evidence of adrenal or pituitary pathology. The peak GH response was compared between the primed and the nonprimed group 1 subjects, and the cortisol levels were compared between the combined test subgroups and the controls. The peak pass level for growth hormone was 10 ng/ml; the peak pass level for cortisol was 520 nmol/l. RESULTS: Sixty-four of the 73 children in group 1 (87.7%) showed a growth hormone level of >/=10 ng/ml on the first stimulation test, including 26/31 (84%) nonprimed and 38/42 (90.5%) primed. Of the 9 patients who failed the first clonidine test, 4 also failed the second, primed test, including 1/5 nonprimed patients (20%) and 3/4 primed patients (75%). This yielded a GH deficiency/insufficiency rate of 5.5% and a rather low false-positive rate of 13.3% (4/30) for the nonprimed subjects and 2. 6% (1/39) for the primed subjects. Peak 30-min cortisol in response to ACTH stimulation was similar in the patients who underwent the 250 microg or the 1 microg test within each group (subgroup 1NP, subgroup 1P and group 2); therefore, the results for the two tests were considered together. Compared with group 2, subgroup 1NP patients had a similar 30-min cortisol response (P=NS), and subgroup 1P patients had a much higher response (P<0.05) (group 2=690+/-145 nmol/l, subgroup 1NP=772+/-195 nmol/l, subgroup 1P=934+/-209 nmol/l). However, there was no significant difference in the increment in cortisol response between the three groups. CONCLUSIONS: Our results suggest that the combined clonidine-short-ACTH test is a reliable and safe tool for the simultaneous assessment of growth hormone reserve and hypothalamic-pituitary-adrenal axis integrity in children. PMID- 10870039 TI - Outcome in patients with adrenal incidentaloma selected for surgery: an analysis of 88 cases investigated in a single clinical center. AB - OBJECTIVE: The study was designed to evaluate the clinical, endocrinological and radiological parameters used to investigate adrenal incidentalomas and select patients for surgery. DESIGN AND METHODS: An analysis of 88 consecutive patients with adrenal incidentaloma selected for surgery and investigated in a single clinical center was performed. RESULTS: Mean (+/-s.d.) age of the patients was 53+/-14 years. Fourteen (16%) of the adrenal incidentalomas were malignant tumors (2 adrenocortical carcinomas, 3 metastases, 4 adenocarcinomas, 4 sarcomas and 1 mesenchymoma), 10 (11%) were pheochromocytomas, 32 (36%) were non-secretory benign adrenal adenomas and the remaining were benign adrenal (n = 8; 9%) or extra-adrenal (n = 24; 27%) masses. Endocrinological investigations revealed 1 Conn adenoma, 4 tumors responsible for Cushing's syndrome or silent hypercortisolism and 1 androgen secreting tumor. Abnormalities of endocrine evaluations were observed in the 2 malignant adrenocortical carcinomas. Elevated 24-h urinary metanephrine levels were observed in the 9 pheochromocytomas tested. Complications of surgery occurred in 14% of the cases. Regardless of the endocrine status, parameters associated with malignant tumors were: older age (mean age of patients harboring malignant tumors vs patients with benign incidentalomas: 62+/-17 years vs 52+/-13 years, P = 0.005), weight loss (39% vs 7%, P = 0. 005), and mass diameter greater than 60mm (69% vs 15%, P < 0.001). By multiple logistic regression analysis malignant tumors were associated with increased age, diameter greater than 60mm and bilateral masses. CONCLUSION: This study points to a high rate of pheochromocytomas and malignant tumors in patients selected for surgery. This high rate differs from some previous reports and might be explained by the criteria used to select patients for surgery. Among these two groups of tumors, careful systematic endocrinological investigations allow the detection of altered secretion in the vast majority - if not all - malignant tumors of adrenal origin and pheochromocytomas. Only 5% of the incidentalomas below 30 mm selected for surgery in this study were malignant, in keeping with the use of this criteria as an important parameter to select patients with normal hormonal investigations for careful follow-up. PMID- 10870040 TI - Effect of tryptophan on the early tri-iodothyronine uptake in mouse thymocytes. AB - OBJECTIVE: We have studied the effect of tryptophan on cellular [(125)I]tri iodothyronine (T3) uptake by mouse thymocytes. MATERIALS AND METHODS: Mouse thymocytes (20 x 10(6 )cells/ml) were suspended in Krebs-Ringer solution buffered by Tris-HCl and incubation (23 degrees C at pH7.45+/-0.6), in the presence or absence of 1mM tryptophan, was started by adding 25 pM [(125)I]T3. At the end of incubation, samples were cooled in ice, centrifuged over a 30% sucrose cushion and the cell-associated radioactivity was measured in the pellet. RESULTS: Tryptophan reduced both the total and the saturable fraction of [(125)I]T3 uptake by 44% (P=0.0009) and 60% (P=0.0006) respectively, following 1 min of incubation. This effect was specific and dose-dependent, being maximal at 5mM concentration ( 82%). In contrast, the pre-exposure of cells to tryptophan for up to 2h had no effect on the subsequent uptake of [(125)I]T3, in the absence of tryptophan. The effect of D-tryptophan on saturable T3 uptake was not different from that obtained using the L-stereoisomer. Tryptophan reduced the V(max) of the initial rate of saturable [(125)I]T3 uptake by two-thirds without affecting the apparent K(m) (2.2 nM) of the process, thus indicating the non-competitive nature of the inhibition. In sodium-free medium the saturable [(125)I]T3 uptake was reduced by 43%. The inhibitory effect of tryptophan on [(125)I]T3 uptake was exerted in both the presence and the absence of sodium. In fact, the inhibitory effect of tryptophan on T3 transport was greater and significantly different (P=0.0046) from that obtained by sodium depletion alone. CONCLUSIONS: Tryptophan interferes with both the sodium-dependent and -independent components of [(125)I]T3 uptake by a dose-dependent, non-competitive mechanism which operates in cis-modality at the plasma membrane level of mouse thymocytes. PMID- 10870041 TI - Functional characteristics of insulin receptors with a Thr-->Ser1200 mutation overexpressed in Chinese hamster ovary cells. AB - OBJECTIVE: To investigate the functional properties of insulin receptors with a Thr-->Ser(1200)-mutation that is associated with severe insulin resistance in humans. DESIGN AND METHODS: The effect of in situ insulin-stimulation on insulin receptor kinase activity was studied in Chinese hamster ovary cells with overexpressed human Ser(1200)-mutated, non-mutated, and ATP-binding site-mutated (Lys-->Arg(1030)) receptors using a microwell-based assay that only detects human (and not hamster) insulin receptors. Moreover, the fraction of anti phosphotyrosine antibody-binding receptors following in situ stimulation was separated, and autophosphorylation and kinase activity resulting from in situ and/or in vitro activation evaluated in this fraction. RESULTS: Although insulin stimulated kinase activity of human-specific anti-insulin receptor antibody binding receptors in cells with Ser(1200)-mutated insulin receptors represented only 3.3% of that reached in cells with non-mutated receptors, a clear insulin induced increase in kinase activity was observed (3.4-fold; P<0.05). This increase was associated with a 2.3+/-0.6% (P<0.05) increase in anti phosphotyrosine-binding receptors with a kinase activity representing 43+/-8% of that found in activated non-mutated receptors. In vitro autophosphorylation and kinase activation proceeded much more slowly in Ser(1200)-mutated receptors (t(1/2)): 100 min) compared with non-mutated receptors (t(1/2)): 1 min) and were inhibitable by lower alkaline phosphatase concentrations (EC(50): 3 U/ml and 70 U/ml respectively). No activation of insulin receptor kinase was observed with Arg(1030)-mutated receptors. CONCLUSIONS: Overexpressed Ser(1200)-mutated human insulin receptors possess insulin-stimulated kinase activity and can be activated in situ and in vitro. They are characterized by a markedly slower autophosphorylation reaction, which, in a phosphatase-containing environment, results in a small fraction of phosphorylated and activated receptors. PMID- 10870042 TI - Human GnRH-secreting cultured neurons express activin betaA subunit mRNA and secrete dimeric activin A. AB - OBJECTIVE: To evaluate the expression of activin betaA-subunit mRNA and the secretion of activin A in/from cultured GnRH-secreting neuronal cells cloned from human olfactory epithelium (FNC-B4), which showed biochemical and antigenic properties of GnRH-secreting neurons. DESIGN: FNC-B4 cells were cultured in basal and conditioned media. METHODS: Reverse transcription-polymerase chain reaction (RTR-PCR) evaluated the expression of activin betaA-subunit mRNA. By using a specific ELISA, dimeric activin A concentrations were measured in culture media, in the absence or presence of carvone or forskolin and with different doses of progesterone, GnRH, and estradiol. RESULTS: RT-PCR experiments performed on total RNA isolated from FNC-B4 cells, using specific primers for the activin betaA gene, showed a 787bp DNA band corresponding to the betaA gene. FNC-B4 cells secreted activin A, and the highest accumulation in conditioned medium was achieved after 3h culture: the addition of forskolin, but not of carvone, was able to stimulate the release of activin A from cultured neuronal cells (P<0.01). When progesterone or GnRH was added, a significant accumulation of activin A was observed (P<0.01), while estradiol administration did not significantly affect activin A secretion. CONCLUSION: To date, this is the only study, in an in vitro human model reporting, that GnRH-secreting neuronal cells expressed activin betaA subunit mRNA, and released dimeric activin A in culture medium. The expression and secretion of activin suggests that in these cells activin A might exert its action by autocrine/paracrine mechanisms. PMID- 10870043 TI - Preventive effects of a herbal medicine on bone loss in rats treated with a GnRH agonist. AB - The study was designed to evaluate the effects of a traditional Chinese herbal medicine Hochu-ekki-to (Bu-zong-yi-qi-tang), which was composed of 10 herbal medicines and had been used for the treatment of oligospermia and as a postoperative medication in Japan, on bone loss in rats treated with a gonadotropin-releasing hormone (GnRH) agonist. Female rats at 40 weeks of age were divided into 4 groups of 8 rats each. In the three experimental groups, each animal received subcutaneous injections of the long-acting GnRH agonist, buserelin acetate, once every four weeks throughout the experiment. Beginning at 48 weeks of age, the experimental groups were given diets containing conjugated estrogens or Hochu-ekki-to for 8 weeks. The administration of the GnRH agonist reduced the bone mineral density in the whole femur to 91.0% of that in the control group. However, administration of conjugated estrogens and Hochu-ekki-to increased the serum concentrations of estradiol 16.8- and 5.3-fold respectively compared with concentrations in the GnRH agonist-treated group, resulting in the augmentation of the bone mineral density to 110.3% and 106.2% respectively. These findings indicate that Hochu-ekki-to enhances the reduced bone mineral density and causes a slight elevation of the serum estradiol levels in the chemically castrated rats. PMID- 10870044 TI - Cyclin D and cyclin E expression in normal and adenomatous pituitary. AB - OBJECTIVES: Cyclins play an important role in the regulation of cell progression through the cell cycle. Over-expression of the cyclins has been shown in many different tumour types. Pituitary adenomas are a common form of endocrine neoplasia in the human, but have been little studied in terms of the expression of the principal cyclins regulating checkpoint exit, cyclin D1 and cyclin E. METHODS: We therefore investigated the expression of cyclin D1 and cyclin E in a range of benign and metastatic pituitary tumours. We studied a total of 95 pituitaries, including normal pituitary (n=20), Cushing's disease (n=19), somatotroph tumours (n=19), non-functioning adenomas (n=18), prolactinomas (n=7), aggressive tumours (n=9) and pituitary carcinoma (n=3). All tumours and normal tissue were immunostained for cyclin D1 and cyclin E using a standard technique, and were then subjected to blinded analysis by a single observer and the extent of staining quantified on the basis of 500 cell counts per tissue. The distribution of positive staining between different tissues was analysed by non parametric test procedures. RESULTS: There was no cytoplasmic staining for cyclin D1 in any tissue. Nuclear staining was generally sparse, but was statistically more frequent in non-functioning and aggressive tumours compared with other tumour types or normal pituitary. Cyclin E was also sparsely expressed, but was specifically increased in corticotroph tumours from patients with Cushing's disease. CONCLUSIONS: We report cyclin D1 over-expression in aggressive and non functioning pituitary tumours, and that cyclin E expression is more frequently seen in Cushing's disease. The high level of cyclin E expression in Cushing's disease may relate to the low level of p27 protein expression previously reported in corticotroph tumours. PMID- 10870045 TI - Arthur I. Holleb, M.D PMID- 10870046 TI - Artificial neural networks for prostate carcinoma risk assessment: An overview. PMID- 10870047 TI - Allelic loss is heterogeneous throughout the tumor in colorectal carcinoma. AB - BACKGROUND: Loss of heterozygosity (LOH) at 17p and 18q in colorectal carcinoma has been depicted as a potential prognostic marker for the disease. However, conclusions vary among reports, and evidence of clinically useful genetic prognostic markers is still lacking. As a rule, single biopsies are used. In this study, the authors hypothesized that an important cause of earlier contradictory results was the heterogeneity of colorectal neoplasms. METHODS: In this study, DNA originating in each quadrant of tumors from 64 patients with colorectal carcinoma was analyzed. Microsatellite markers for chromosome 18q and 17p were amplified by polymerase chain reaction and automatically analyzed. RESULTS: The authors found that, regardless of stage, LOH and non-LOH in both 17p and 18q varied among biopsies within the tumors in a random fashion. LOH in 18q was detected in all 4 quadrants in 22% and in 1 of 4, 2 of 4, or 3 of 4 quadrants in 56% of the tumors, whereas 22% of the tumors were homogeneously without LOH in 18q. LOH 17p was distributed similarly throughout the tumors and was present in 1 of 4, 2 of 4, or 3 of 4 of the quadrants in 44%. The authors also reexamined a subset of tumors by subdividing one biopsy from each into four. Analysis of the microsatellite markers then yielded identical results. No correlation between the degree of LOH status and patient survival was observed. CONCLUSIONS: LOH status within a colorectal tumor is extensively heterogeneous. However, it is more homologous on a lower macroscopic level. For relevant genetic analysis, multiple biopsies and DNA sampling preceded by careful morphologic examination must be standard in the preparation of DNA. PMID- 10870048 TI - Is heterozygous alpha-1-antitrypsin deficiency type PIZ a risk factor for primary liver carcinoma? AB - BACKGROUND: It is well known that homozygotes with alpha-1-antitrypsin deficiency type PiZ are associated with an increased risk of chronic liver disease and liver carcinoma. The aim of this study was to determine whether heterozygous PiZ status is a risk factor for liver carcinoma development. METHODS: Three hundred seventeen consecutive primary liver carcinomas and the tumor-bearing liver tissue (tumor series) from adult patients were screened immunohistochemically for hepatocellular PiZ deposits. Liver specimens from 1663 consecutive adult patients (biopsy series) and liver tissue from 1030 consecutive adult autopsies (autopsy series) served as controls. The zygosity status of alpha-1-antitrypsin was verified by analysis of single strand conformational polymorphism and by sequencing DNA extracted from paraffin embedded tissue. RESULTS: The PiZ frequency in the tumor series (5.99%) was significantly higher than in the biopsy series (3.43%) or the autopsy series (1.84%). Cholangiocarcinomas and/or combined hepatocholangiocarcinomas were seen significantly more frequently in PiZ associated liver carcinomas (57.9%) than in non-PiZ-associated carcinomas (27.2%). Cirrhosis was found in only 3 of the 19 PiZ-associated carcinomas. The remaining 16 livers showed varying stages of fibrosis or normal tissue. All nine cases with PiZ-associated liver carcinoma suitable for genetic analysis showed heterozygous PiZ mutations. CONCLUSIONS: Heterozygotes of type PiZ are associated with an increased risk of primary liver carcinoma. PiZ-associated carcinoma may develop in noncirrhotic liver tissue and without concurrent liver disease, and is frequently characterized by cholangiocellular differentiation. The site specific antibody ATZ11 is a reliable morphologic tool for detecting PiZ individuals. PMID- 10870049 TI - Vinorelbine in elderly patients with inoperable nonsmall cell lung carcinoma: a phase II study. AB - BACKGROUND: Cancer in the elderly is becoming a complex and frequent issue. At least 30% of lung carcinomas are expected to arise each year in elderly patients, who often have significant comorbidity. The most appropriate treatment for this large portion of cancer patients remains unknown. The purpose of this Phase II trial was to make a comprehensive evaluation of the activity, toxicity, and tolerability of single-agent vinorelbine in elderly and relatively poorly performing patients with inoperable nonsmall cell lung carcinoma (NSCLC). METHODS: Patients age 70 years or older were eligible to participate in this trial if they had a pathologic diagnosis, a performance status lower than 4 (Eastern Cooperative Oncology Group [ECOG] scale), and gave informed consent. Vinorelbine was given intravenously (i.v.) at a dose of 25 mg/m(2) every week until progression, persistent toxicity, or refusal. RESULTS: Forty-six patients entered the study; their median age was 75 years (range, 70-83 years). Five patients never started on vinorelbine; 27 others had early treatment suspensions. The median number of weekly infusions was 5 (range, 0-28); the median dose intensity was 70% of projected. Toxicity was generally mild, mainly hematologic, and never life-threatening. ECOG performance status, body weight, and almost all the scores from the quality-of-life questionnaires remained constant during the first 6 weeks of treatment. Two patients obtained partial response, 10 patients had some tumor regression, and 26 had tumor stabilization. The estimated median time to progression was 19 weeks (quartile range, 11-23 weeks), and the median survival 34 weeks (quartile range, 16-63 weeks). CONCLUSIONS: In our group of patients who had poor prognoses, vinorelbine was well tolerated, moderately active, and capable of ensuring relatively long survival. It may represent a valuable therapeutic option for the treatment of nonresectable NSCLC in elderly patients. PMID- 10870050 TI - Mast cell density is associated with angiogenesis and poor prognosis in pulmonary adenocarcinoma. AB - BACKGROUND: Angiogenesis in individuals with various solid tumors has provided useful information on tumor progression and prognosis, and many examples of mast cell (MC) accumulation coupled with angiogenesis can be found in the literature. METHODS: Utilizing monoclonal antibodies for tryptase that are specific markers for MC and for the endothelial surface marker CD34, the authors quantified MC infiltration in 180 patients with pulmonary adenocarcinoma who underwent curative tumor resection, to investigate the relation between mast cell density (MCD) and microvessel density (MVD), clinicopathologic factors, and prognosis. RESULTS: A significant association was found between MCD and MVD (P < 0.0001). The MC count was significantly related to tumor progression, involving N classification and stage (P = 0.0002 for N classification and P = 0. 0015 for stage). A significant difference in the rate of patient survival was detected between patients whose tumors had an MCD defined as high and those whose tumors had an MCD defined as low (P < 0.0001). Multivariate analysis also showed that MCD was significantly related to survival (P = 0.0378). CONCLUSIONS: These data indicate that MC infiltration may contribute to tumor angiogenesis and tumor progression, and that MCD is a useful prognostic marker in pulmonary adenocarcinoma. PMID- 10870051 TI - Analysis of therapeutic and immunologic effects of R(24) anti-GD3 monoclonal antibody in 37 patients with metastatic melanoma. AB - BACKGROUND: Antitumor effects of antibodies against ganglioside antigens of melanoma have been reported, but neither optimal doses nor mechanisms have been established. METHODS: This Phase IB trial of the murine immunoglobulin IgG(3) monoclonal antibody R(24) against disialoganglioside GD3 was conducted with 37 patients to define better the dose-response relation and mechanism of action of R(24) in patients with metastatic melanoma. RESULTS: Dose-limiting toxicity consisted of a pulmonary capillary leak syndrome in 3 of 5 patients in the 80 mg/M(2)/day dosage tier. Serial blood and tumor biopsy samples were obtained prior to therapy and on Days 5, 9, and 22 following R(24) infusion. Tumor biopsy infiltrating lymphocytes were enumerated in peritumoral, endotumoral, and perivascular compartments: endotumoral CD4(+) and CD8(+) T cells and HLA-DR(+) T cells increased over time on R(24) antibody. Endotumoral CD4 lymphoid infiltrate activation (DR expression) and antibody-dependent cytotoxicity were the greatest in the one patient who achieved a complete response. CONCLUSIONS: Clinical response was associated with depression in natural killer (CD56(+) and CD56(+)DR(+)) blood cells (P = 0.03) and was associated with R(24) dosage (P = 0.01). A complete response that lasted 2 years and a partial response that lasted 2 months occurred at a dose of 1 mg/M(2)/day. The limited number of clinical responses observed in this trial hampered the correlation of antitumor and immune parameters but provided a rational foundation for the future evaluation of antiganglioside antibodies. PMID- 10870052 TI - Pigmentary traits, modalities of sun reaction, history of sunburns, and melanocytic nevi as risk factors for cutaneous malignant melanoma in the Italian population: results of a collaborative case-control study. AB - BACKGROUND: To the authors' knowledge, limited data are available from Mediterranean populations concerning risk factors for malignant melanoma. A few Italian case-control studies have produced conflicting results regarding the association between malignant melanoma and pigmentary traits, sunburns, and melanocytic nevi. METHODS: A case-control study was conducted within the framework of the Italian Group for Epidemiologic Research in Dermatology (GISED). Twenty-seven centers in the north and south of Italy participated. A total of 542 cases and 538 controls were entered onto the study. A standardized questionnaire was administered to cases and controls. Cases and controls also were examined by trained dermatologists who were required to count the number of melanocytic nevi (those measuring > or = 2 mm and > 6 mm in greatest dimension, separately) and to make judgments regarding pigmentary traits. RESULTS: In the multivariate analysis, eye and skin color, propensity to sunburn, history of sunburns before age 15 years, and solar lentigines all were associated with malignant melanoma. In addition, the risk of melanoma increased with the number of melanocytic nevi > or = 2 mm. Nevi > 6 mm in greatest dimension had effects on risk that appeared to be independent from the effects of smaller nevi (2-6 mm). CONCLUSIONS: The results of the current study largely are similar to those obtained in northern European countries, the U.S., and Australia and provide further evidence of the importance of selected pigmentary traits, sun exposure, and the number of melanocytic nevi in the risk of cutaneous malignant melanoma. PMID- 10870053 TI - Dermatofibrosarcoma protuberans: A clinicopathologic analysis of patients treated and followed at a single institution. AB - BACKGROUND: Despite optimal surgical therapy for patients with dermatofibrosarcoma protuberans (DFSP), some patients still continue to develop local recurrence. The authors' objective was to identify and analyze clinicopathologic factors for disease free survival in a large group of patients who were followed prospectively at a single institution. METHODS: Prospectively collected data and pathology slides were available for review from 159 patients with primary or recurrent DFSP who underwent treatment between July 1950 and July 1998. The study group was comprised of patients with either the "classic" form of DFSP or the fibrosarcomatous "high grade" variant of DFSP (FS-DFSP). Patient, tumor, pathologic, and treatment factors were analyzed using the log rank test for univariate influence and Cox regression analysis for multivariate influence. Local recurrence free survival was determined by the Kaplan-Meier actuarial method. RESULTS: Of the 159 patients who comprised the current study group, 134 (84%) had the classic form of DFSP. The FS-DFSP variant was found in the remaining 25 patients (16%). The overall 5-year local recurrence free survival rate was 75%, with a median follow-up of 4. 75 years. The 5-year recurrence free survival rate for each group was 81% and 28%, respectively. On univariate analysis, age > 50 years, very close (< 1 mm) to positive microscopic margins, FS DFSP variant, high mitotic rate, and increased cellularity were unfavorable prognostic factors. Multivariate analysis determined very close (< 1 mm) to positive microscopic margins and FS-DFSP variant to be independent adverse prognostic factors. For the 34 patients who developed a recurrence after surgical resection (21%), the median time to local recurrence was 32 months. Of the patients in this group, two died from metastatic disease. CONCLUSIONS: The prognosis after surgical resection with negative and sometimes positive microscopic margins for patients with DFSP is very good. However, increased age, high mitotic index, and increased cellularity are predictors of poor clinical outcome. The FS-DFSP variant represents a much more aggressive tumor with metastatic potential. Patients who are treated with curative intent for FS-DFSP should undergo aggressive attempts at complete surgical resection. Patients with recurrent classic DFSP without evidence of adverse prognostic features may benefit from conservative management, especially in the setting of potentially unresectable disease. PMID- 10870054 TI - A comparison of staging systems for localized extremity soft tissue sarcoma. AB - BACKGROUND: Staging systems for soft tissue sarcoma (STS) are important to identify patients with similar systemic risk who might benefit from specific treatments. This study compared four commonly used staging systems for predicting systemic outcomes of patients with localized extremity STS, as proposed by the fourth and fifth editions of the American Joint Committee on Cancer/International Union Against Cancer (AJCC/UICC) staging system, the Memorial Sloan-Kettering Cancer Center (MSK) system, and the Surgical Staging System (SSS) of the Musculoskeletal Tumor Society. METHODS: Three hundred consecutive adult patients with newly diagnosed nonmetastatic STS of the lower extremity were treated at Memorial Sloan-Kettering Cancer Center between 1982 and 1989. Metastasis free survival was the end point of the study. The prognostic value of the four staging systems and their components were examined in univariate and multivariate analyses. The Akaike information criterion (AIC) was used to identify the system that best predicted the risk of systemic recurrence. RESULTS: Compartment status, depth, grade, and size were all independent predictors of outcome within their respective staging systems. However, when compared with one another, only depth, grade, and size retained their prognostic significance. Of the four models, the AIC predicted that the MSK was the best predictor of systemic relapse, followed by the fifth edition of the AJCC/UICC staging system. CONCLUSIONS: Staging systems such as the MSK system or the fifth edition of the AJCC/UICC system, which include tumor depth, grade, and size as prognostic factors, are the most predictive of systemic relapse in patients presenting with localized extremity STS. Both of these systems identify the same group of patients at the highest risk who would be the most suitable for adjuvant chemotherapy trials. PMID- 10870055 TI - Vinorelbine combined with paclitaxel infused over 96 hours (VI-TA-96) for patients with metastatic breast carcinoma. AB - BACKGROUND: Vinorelbine (VI) and paclitaxel (TA) are among the most active single agents in the treatment of patients with breast carcinoma, and both have microtubules as their cytotoxic target. This Phase I-II study combined these 2 agents and used a 96-hour intravenous (i.v.) infusion of paclitaxel to maximize their cytotoxic activities. METHODS: Patients with metastatic breast carcinoma who were previously treated with chemotherapy were administered increasing doses of a 96-hour paclitaxel i.v. infusion from Days 1 to 5, with a first fixed dose of vinorelbine (12.5 mg/m(2) on Days 1 and 5) every 3 weeks. The dose of paclitaxel was then decreased starting from the previously established tolerated dose, and a second fixed dose of vinorelbine (15 mg/m(2) on Days 1 and 5) was given. This identified 2 acceptable doses of paclitaxel (110 mg/m(2) with VI 12.5 mg/m(2) and 90 mg/m(2) with VI 15 mg/m(2)). The latter was used in the subsequent Phase II study. RESULTS: For the 50 patients treated with any dose, the complete response (CR) and the CR plus partial response (PR) rates were, respectively, 14% and 48% (95% confidence interval [CI], 34-67%). When only the 27 patients treated with the Phase II dose were considered, the figures were, respectively, 11% and 52% (95% CI, 42-62%). The median time to progression was 26 weeks, and the median survival 51 weeks. The dose-limiting toxicity was febrile neutropenia. CONCLUSIONS: At the dose schedule identified for the Phase II study, the VI-TA-96 combination has considerable antitumor activity; pharmacoeconomic interest (it requires about half the doses of the agents administered singly); no major toxicity, except G4 neutropenia; and no need for premedication. This combination may be recommended as one of the most effective therapeutic options for patients with metastatic breast carcinoma who were pretreated mainly with anthracycline containing chemotherapy. PMID- 10870056 TI - Bilateral breast carcinoma: risk factors and outcomes for patients with synchronous and metachronous disease. AB - BACKGROUND: The aim of this study was to compare the outcomes of bilateral breast carcinoma (BBC) patients with those of patients who had unilateral disease. METHODS: From 1960 to 1995, 1465 Stage 0-III patients with primary breast carcinoma were treated with either mastectomy or breast conservation therapy at the Kimmel Cancer Center of Jefferson Medical College and Thomas Jefferson University Hospital. There were 1315 (89.9%) unilateral, 103 (7.1%) metachronous, and 47 (3.0%) synchronous breast carcinoma patients. Patients with synchronous breast carcinoma were defined as having a contralateral cancer diagnosed within 1 year of initial diagnosis. The percentage of patients with Stage 0-I disease at initial diagnosis was 49.4%, whereas 68% had Stage 0-I disease at subsequent diagnosis. For patients with metachronous breast carcinomas, the median interval between the first and second diagnosis was 44 months (range, 13-287 months). The median follow-up time was 58 months (range, 12-229 months) for patients with synchronous cancers and 87 months (range, 0.25-414 months) for those with metachronous cancers. Rates of overall survival and survival with no evidence of disease (NED survival), local control, and distant metastasis from the time of the second diagnosis were calculated for patients with synchronous and metachronous disease. These figures were then compared with each other and also with those for unilateral breast carcinoma patients. RESULTS: Patients with synchronous and metachronous breast carcinoma had worse 5- and 8-year NED survival rates compared with unilateral breast carcinoma patients, as well as an increased risk of distant metastasis. In multivariate analysis, differences in local control and overall survival were not statistically significant for patients who had bilateral disease compared with those who had unilateral disease. There was no difference when patients with metachronous and unilateral breast carcinoma were compared with respect to local control and overall survival. CONCLUSIONS: Patients with bilateral breast carcinoma who present with synchronous disease are at greater risk for distant metastasis than women with unilateral or metachronous breast tumors. There was a trend toward decreased overall survival and local control for patients with synchronous bilateral breast carcinoma compared with patients who had either metachronous or unilateral disease. PMID- 10870057 TI - Android obesity at diagnosis and breast carcinoma survival: Evaluation of the effects of anthropometric variables at diagnosis, including body composition and body fat distribution and weight gain during life span,and survival from breast carcinoma. AB - BACKGROUND: Although a large body of research exists concerning pathologic prognostic indicators of the rate of incidence and survival from breast carcinoma, to the authors' knowledge very few studies have examined the effects of anthropometric variables such as height, obesity, weight gain in adulthood, timing of weight gain, and body composition to survival, although these variables are related to the incidence rate. METHODS: The survival status of 166 patients diagnosed with primary breast carcinoma and followed for at least 10 years was obtained from the Cancer Center's registry, and significant anthropometric and other known prognostic indicators regarding survival after diagnosis were determined by Cox proportional hazards analysis. RESULTS: Eighty-three of 166 breast carcinoma patients (50%) with up to 10 years of follow-up died of disease. Android body fat distribution, as indicated by a higher suprailiac:thigh ratio, was a statistically significant (P < 0.0001) prognostic indicator for survival after controlling for stage of disease, with a hazards ratio of 2.6 (95% confidence interval [95% CI], 1.63-4.17). Adult weight gain, as indicated specifically by weight at age 30 years, was a statistically significant (P < 0.05) prognostic indicator for survival with a hazards ratio of 1.15 (95% CI, 1.0 1.28). In addition, the authors observed the Quatelet Index, a negatively significant (P < 0.01) prognostic indicator for survival with a hazards ratio of 0.92 (95% CI, 0.87-0.98). Other markers of general obesity such as weight at diagnosis, percent body fat, and body surface area were not significant markers influencing survival. Similarly, height; triceps, biceps; subscapular, suprailiac, abdominal, and thigh skinfolds; waist and hip circumferences; family history; and reproductive and hormonal variables at the time of diagnosis showed no apparent significant relation to survival. CONCLUSIONS: The results of the current study provide some evidence that android body fat distribution at diagnosis and increased weight at age 30 years increases a woman's risk of dying of breast carcinoma. PMID- 10870058 TI - Negative immunomagnetic purging of peripheral blood stem cell harvests from breast carcinoma patients reduces tumor cell contamination while not affecting hematopoietic recovery. AB - BACKGROUND: Because tumor contamination of hematopoietic stem cell grafts may influence the outcome in breast carcinoma (BC) patients undergoing high dose chemotherapy (HDC), several ex vivo procedures for the purging of autologous harvests have been investigated. The authors studied the presence of epithelial tumor cells and the growth of hematopoietic progenitors in peripheral blood stem cell (PBSC) collections from patients with metastatic breast carcinoma before and after a purging procedure performed by a negative immunomagnetic BC cell separation. METHODS: Eighteen patients entered the study. Tumor contamination was assessed by conventional immunocytochemistry (ICC) and by a liquid culture assay developed in the study laboratory. Committed and more primitive hematopoietic progenitors were quantitated before and after the negative selection. Ten patients received HDC with purged PBSC support. RESULTS: Before purging, 4 of 18 PBSC collections were found to be contaminated by liquid culture; among these samples, only 1 was positive by ICC. Three of the four positive collections, including the ICC positive sample, became negative after immunomagnetic selection whereas BC cells still were present after the procedure in one harvest. A high recovery of both primitive and mature hematopoietic progenitors was found after the purging procedure. Patients receiving purged PBSC after myeloablation had a prompt and complete hematopoietic reconstitution, and no graft failure was observed at a median follow-up of 1 year. CONCLUSIONS: The preliminary results of the current study suggest that negative selection of BC cells is able to purge PBSC effectively while having no apparent affect on hematopoietic progenitor recovery in vitro and in vivo. PMID- 10870059 TI - Expression of p27 and p53 in cervical squamous cell carcinoma patients treated with radiotherapy alone: radiotherapeutic effect and prognosis. AB - BACKGROUND: The p27/Kip1 gene inhibits a variety of cyclin-cyclin-dependent kinase complexes and regulates cell growth. The p53 gene acts as a tumor suppressor gene, controlling entry into the S-phase of the cell cycle. METHODS: A total of 202 biopsy specimens obtained from 77 patients with squamous cell carcinoma of the cervix before and during radiotherapy (RT) was investigated for expression of p27 and p53 in conventionally fixed and processed histologic specimens using an immunohistochemical method. DNA samples exhibiting high p53 overexpression were analyzed for detection of the wild-type or mutant-type of p53 by polymerase chain reaction-single strand conformation polymorphism analysis. RESULTS: Carcinoma cells and degenerated or swollen carcinoma cells after RT that were positive for p27 and p53 showed intranuclear reactivity. Degenerated or swollen carcinoma cells after RT with 27 Gy showed stronger p53 positivity than carcinoma cells before RT. The mean p27 labeling index was decreased significantly after 27 Gy; conversely, the mean p53 labeling index was increased significantly after 27 Gy of RT. A high p27 labeling index before RT was associated significantly with good disease free and metastasis free survival. A high p53 labeling index before RT was associated with poor overall survival. Both samples examined before RT showed no mutations of p53 (exons 5-8). Four of 5 samples showed mutations in exon 5 or 7 of the p53 gene after 27 Gy of RT. CONCLUSIONS: The high p27 expression and low p53 expression in carcinoma cells before RT are regarded as predictive factors for good prognosis of patients with cervical squamous cell carcinoma treated with RT alone. The mean p27 and p53 indices change in an inverse fashion during the period between the initiation of RT and the period after 27 Gy of RT. RT induces the mutant-type p53 oncogene after 27 Gy. PMID- 10870060 TI - Immunohistochemical staining of GLUT1 in benign, hyperplastic, and malignant endometrial epithelia. AB - BACKGROUND: Aberrant expression of the facilitative glucose transporter, GLUT1, is found in a wide spectrum of epithelial malignancies. The current study describes an immunohistochemical study of GLUT1 expression in benign, hyperplastic, and malignant endometrial epithelia. METHODS: One hundred sixteen formalin fixed, paraffin embedded sections of benign, hyperplastic, and malignant endometrial epithelium were immunostained with rabbit anti-GLUT1 antibody using the streptavidin-biotin method. RESULTS: Proliferative, secretory, and atrophic endometrium were not stained with GLUT1 antiserum. Rare, minute foci of tubal metaplasia stained positively. Simple and complex endometrial hyperplasias were consistently GLUT1 negative. All specimens of atypical hyperplasia had foci that were positive for GLUT1. All endometrial adenocarcinomas were GLUT1 positive. CONCLUSIONS: The results of the current study appear to indicate that 1) aberrant overexpression of GLUT1 is a consistent feature of endometrioid adenocarcinoma; 2) GLUT1 immunostaining may be useful in distinguishing benign hyperplasias from hyperplasias that are associated strongly with malignancy; and 3) some or all cases of atypical hyperplasia may be neoplastic rather than hyperplastic, given the existence of a molecular defect common to this hyperplastic subtype and endometrioid adenocarcinoma. PMID- 10870061 TI - Pathologic variables and adjuvant therapy as predictors of recurrence and survival for patients with surgically evaluated carcinosarcoma of the uterus. AB - BACKGROUND: The purpose of this study was to determine clinicopathologic variables associated with extrauterine disease, recurrence, and survival in patients with carcinosarcoma (CS) of the uterus. METHODS: Patients believed to have disease confined to the uterine corpus who underwent primary surgical assessment were identified and data retrospectively reviewed. RESULTS: Occult metastases were found in 38 (61%) of 62 patients. At last follow-up, 31 (50%) had had recurrence, with an extrapelvic component in 43%, and 53% had died. Depth of myometrial invasion and lymph-vascular space invasion (LVSI) were associated with extrauterine disease. Five-year survival for patients with disease confined to the corpus (74%) was significantly greater than for those with more advanced disease (24%, P = 0.0013). Factors associated with recurrence and survival included depth of myometrial invasion, LVSI, adnexal and serosal involvement, positive cytology, and lymph node metastases. Of 24 patients with uterine disease only, 11 received no adjuvant therapy, yet 8 (73%) were free of disease at last follow-up. Neither adjuvant radiotherapy nor chemotherapy was identified as an independent prognostic variable for recurrence or survival. CONCLUSIONS: More than half of patients with CS clinically confined to the uterine corpus harbor occult metastases in a pattern similar to that found with endometrial carcinoma. Survival is significantly diminished for this group. Although the benefit of adjuvant therapy cannot be demonstrated by this study, a number of early stage patients survive without adjuvant therapy. This argues for extending the International Federation of Gynecology and Obstetrics endometrial carcinoma surgical staging system to include CS, and also for conducting prospective trials to examine the benefits of adjuvant therapy for patients with early stage disease. PMID- 10870062 TI - Identification and characterization of circulating prostate carcinoma cells. AB - BACKGROUND: Analysis of prostate carcinoma cells isolated from the peripheral blood suggested a classification based on three categories. METHODS: Centrifugation density gradients and magnetic cell sorting were used to isolate circulating prostate carcinoma cells from peripheral blood. Immunocytochemistry staining and fluorescent in situ hybridization allowed characterization of isolated cancer cells. RESULTS: Terminal cells can be divided into 3 classes: 1) large, buoyant, fragile cells with a large nucleus that were captured in a 1.068 g/mL gradient; 2) enucleate cells (4, 6-diamidino-2-phenylindole [DAPI] negative) that were positive for cytokeratin and PSMA antibodies; and 3) cellular debris exhibiting cytokeratin and PSMA positive staining as well as nuclear debris identified by DAPI staining, which included cytoplasmic debris. Growing cells also exhibited three morphologic characteristics: those possessing stem cell-like morphology and characteristics such as small size, high density, developed cytokeratin systems, PSMA expression, and aneuploidy; those in M phase; and cell clusters. The majority of isolated cells exhibited intermediate characteristics and thus comprised the third group of circulating cancer cells. CONCLUSIONS: Although the significance of the cluster remains undetermined, observation suggests that the cluster has the ability to circulate as a microtumor and subsequently arrest in the small veins and capillaries. It is hypothesized that the clusters could escape certain facets of immune surveillance and possibly gain a selective growth advantage over single cells in a distant site. Further hypothesis proposes that arrested cells recruit growth-promoting nutrients, which would result in the invasion of local blood vessels and vascularization. PMID- 10870063 TI - Brachytherapy: Results of two different therapy strategies for patients with primary glioblastoma multiforme. AB - BACKGROUND: In the current study, the authors describe and compare two different strategies of brachytherapy for the treatment of patients with primary glioblastoma multiforme (GBM). METHODS: The study was comprised of 84 patients. Forty-five patients were implanted with permanent or temporary low activity iodine-125 ((125)I) seeds in Cologne and 21 patients were implanted with temporary iridium-192 ((192)Ir) wires in Amsterdam. Both groups received external beam radiation therapy (EBRT); the (125)I group received 10-30 grays (Gy) with the implant in situ and the (192)Ir group received 60 Gy before implantation. In Cologne, implantation was performed after a diagnostic stereotactic biopsy whereas in Amsterdam implantation took place after cytoreductive diagnostic surgery. In addition, 18 patients in Amsterdam served as a control group. This group received only EBRT after cytoreductive surgery. RESULTS: In both groups the mean age of the patients was between 50-55 years, with 80% of the patients age > 45 years. The mean implantation volume encompassed by the referenced isodose was 23 cm(3) for (125)I and 48 cm(3) for (192)Ir. Initial dose rates were 2. 5-2.9 centigrays (cGy)/hour for permanent (125)I, 4.6 cGy/hour for temporary (125)I, and 44-100 cGy/hour (mean, 61 cGy) for (192)Ir. A total dose of 50-60 Gy, 60-80 Gy, and 40 Gy, respectively, was administered at the outer margins of the tumor. The median survival was approximately 16 months for both the (125)I group and the (192)Ir group. This was 6 months longer than the median survival in the control group. Reoperations were performed in 4 patients in the (125)I group (9%) versus 7 patients in the (192)Ir group (33%). No complications or late reactions were reported in the (125)I group, whereas one case of hemorrhage and three cases of delayed stroke were observed in the (192)Ir group. CONCLUSIONS: The equal median survival times in these two brachytherapy groups with such different dose rate radiation schedules support the hypothesis that dose rate does not play a major role in the survival of patients with primary GBM. PMID- 10870064 TI - Progression of eIF4e gene amplification and overexpression in benign and malignant tumors of the head and neck. AB - BACKGROUND: The overexpression of eukaryotic initiation factor 4E (eIF4E) results in the up-regulation of gene products of mRNAs with long 5' untranslated regions (5' UTRs). The degree of gene amplification increases from the tumor free zone to the tumor core. This led the authors to hypothesize that the degree of eIF4E gene amplification and oncoprotein overexpression is progressive in the cells from normal head and neck tissue, to benign tumors, and eventually to invasive carcinomas (HNCA). METHODS: Using competitive polymerase chain reaction (PCR) and Western blot analysis, benign specimens from similar sites of 10 noncancer patients, 8 pleomorphic adenoma specimens, and 18 HNCA specimens were studied. DNA and protein extracts from each specimen were quantified for eIF4E gene copy number and level of eIF4E protein expression. RESULTS: There was no detectable eIF4E gene amplification and oncoprotein overexpression in benign tissue from noncancer patients (1.1 +/- 0.5 gene copy number [mean +/- standard deviation] and 0.9 +/- 0.5-fold protein elevation, respectively). Four of the eight pleomorphic adenomas analyzed showed eIF4E gene amplification of at least twofold, but none demonstrated protein elevation of any significance. In contrast, all HNCA specimens had detectable eIF4E gene amplification and protein overexpression. Furthermore, the mean degree of eIF4E gene amplification and overexpression was found to increase as cells from benign head and neck tissues (1.1 +/- 0.5 and 0.9 +/- 0.5), benign tumors (2.2 +/- 1.3 and 1.02 +/- 0.19), and HNCA (4.3 +/- 1.2 and 15.5 +/- 9.3) were compared. CONCLUSIONS: Progressive eIF4E gene amplification and overexpression were detected when normal tissues, benign tumors, and HNCA were compared. The degree of gene amplification and overexpression is variable within each tissue category. However, progression to malignant phenotype appears to be associated with an increasing degree of eIF4E gene amplification and overexpression. PMID- 10870065 TI - Estimation of growth rate in patients with head and neck paragangliomas influences the treatment proposal. AB - BACKGROUND: Extraadrenal paragangliomas of the head and neck are tumors with variable clinical behavior. Because tumor growth as well as surgery can cause disabling loss of function, knowledge of the natural history of paragangliomas is important for the development of treatment strategies. METHODS: The tumor volume, growth rate, and tumor doubling time of 48 paragangliomas at different anatomic locations in the head and neck region were estimated retrospectively using sequential radiologic imaging. RESULTS: During a mean follow-up period of 4.2 years, a volume increase of > 20% was observed in 60% of the paragangliomas. In these cases the median growth rate was 1.0 mm/year and the median tumor doubling time was 4. 2 years. More growing tumors were observed in intermediate size tumors than in very small or large paragangliomas, suggesting a biphasic growth pattern. CONCLUSIONS: The majority of head and neck paragangliomas have a very low growth rate. Although management of paragangliomas also is determined by other parameters, preoperative estimation of the tumor doubling time may influence the treatment proposal. A "wait and scan" policy must be considered in all cases. PMID- 10870066 TI - Prevalence, predictive factors, and screening for psychologic distress in patients with newly diagnosed head and neck cancer. AB - BACKGROUND: High levels of distress are a concern regarding patients with head and neck cancer. Early detection of and intervention for such distress are needed to predict patients' adaptation to treatment or rehabilitation, but few studies have investigated the detection of their distress in a patient population of significant size. METHODS: The authors examined 107 consecutive patients with head and neck cancer to assess their psychologic distress (adjustment disorders or major depression) or other psychiatric problems by structured psychiatric interview before the initial cancer treatment. They also evaluated predictive factors for psychologic distress and assessed the ability of a self-rating questionnaire (Hospital Anxiety and Depression Scale, HADS) to screen for distress. RESULTS: Of 107 subjects, 18 (16.8%) had an adjustment disorder or major depression. Thirty-six (33.6%), 7 (6. 5%), and 35 (32.7%) met criteria for alcohol dependence, alcohol abuse, and nicotine dependence, respectively. Logistic regression analysis revealed that having advanced stage cancer (odds ratio, 5. 77; 95% confidence interval [CI], 1.41-39.7; P = 0.03) and living alone (odds ratio, 4.83; 95% CI, 1.04-22.2; P = 0.04) were significantly associated with having psychologic distress. The optimal cutoff point for the HADS screening for psychologic distress was 15. This cutoff point gave 72.2% sensitivity and 81.4% specificity. CONCLUSIONS: Head and neck cancer patients who have advanced disease or live alone should be assessed so that psychologic distress can be detected and intervention made. HADS is a useful clinical instrument to screen for their distress. PMID- 10870067 TI - Organizational systems used by California capitated medical groups and independent practice associations to increase cancer screening. AB - BACKGROUND: Patients in health maintenance organizations (HMOs) appear to have higher utilization of cancer screening tests than patients with fee-for-service insurance. METHODS: The authors surveyed the medical directors of 174 physician organizations in a California network model HMO to obtain information regarding their organizational structure, implementation of guidelines, and use of systems to increase cancer screening. RESULTS: The majority of independent practice associations (IPAs) and medical groups (IMGs) in this California HMO had guidelines and office systems aimed at improving cancer screening. These activities were reported more frequently for mammography and Papanicolaou (Pap) smears than for colorectal carcinoma screening. IMGs reported using flow sheets more often than IPAs. Chart audits were performed more frequently for mammography (48% for IMGs and 40% for IPAs) and Pap smears (45% and 40%, respectively) than for colorectal carcinoma screening (38% and 30%, respectively). Approximately 50% of IPAs and IMGs reported mailing reminders to patients for mammography and Pap smears, but only a few did so for colorectal carcinoma screening. Annual fecal occult blood testing was believed by most medical directors to be a reasonable strategy for managed care patients (86% of IPAs and 96% of IMGs); however, fewer believed that screening sigmoidoscopy for patients ages 50-70 years was a reasonable expectation (71% and 78%, respectively). CONCLUSIONS: The majority of IPAs and IMGs in this California HMO reported using both guidelines and office systems to improve cancer screening rates. Further research is needed to understand the effect of these systems, as well their complex interactions with competing incentives, on cancer screening in managed care patients. PMID- 10870068 TI - A randomized, controlled, parallel-group clinical trial comparing multilayer bandaging followed by hosiery versus hosiery alone in the treatment of patients with lymphedema of the limb. AB - BACKGROUND: Multilayered, low stretch bandages (MLB) combined with exercises, skin care, and manual lymphatic drainage therapy are recommended as an intensive phase of treatment for lymphedema patients. The relative efficacy of each of the components of this comprehensive treatment program have not been determined. This study aimed to compare the effect of multilayer bandaging as an initial phase of lymphedema treatment followed by elastic hosiery versus hosiery alone. METHODS: A randomized, controlled, parallel-group trial was undertaken in the setting of the Lymphedema Clinic, The Royal Marsden Hospital, London. Ninety women with unilateral lymphedema (of the upper or lower limbs) were enrolled in the study. The interventions consisted of 18 days of multilayer bandaging followed by elastic hosiery or hosiery alone, each for a total period of 24 weeks. The main outcome measure was the percentage reduction in excess limb volume. RESULTS: The reduction in limb volume by MLB followed by hosiery was approximately double that from hosiery alone and was sustained over the 24-week period. The mean overall percentage reduction at 24 weeks was 31% (n = 32) for MLB versus 15.8% (n = 46) for hosiery alone, for a mean difference of 15. 2% (95% confidence interval, 6.2 24.2) (P = 0.001). CONCLUSIONS: Multilayer bandaging as an initial phase of treatment for lymphedema patients, followed by hosiery, achieves greater and more sustained limb volume reduction than hosiery alone. PMID- 10870069 TI - A phase I dose escalation of combination chemotherapy with granulocyte-macrophage colony stimulating factor in patients with neuroblastoma. AB - BACKGROUND: Dose intensity is important in the response to chemotherapy in patients with advanced neuroblastoma. The aim of the current study was to determine the maximum tolerated dose of a combination chemotherapy regimen in the treatment of patients with recurrent neuroblastoma and peripheral neuroepithelioma (primitive neuroectodermal tumor [PNET]) and whether the use of growth factor would allow increased dose intensity. METHODS: Twenty-nine patients diagnosed with recurrent neuroblastoma or PNET were treated with a combination chemotherapy regimen of cisplatin, 160 mg/m(2)/96 hours; doxorubicin, 40 mg/m(2)/96 hours; and escalated doses of etoposide and ifosfamide. Granulocyte macrophage-colony stimulating factor (GM-CSF) was administered beginning 24 hours after the completion of the chemotherapy. Courses were repeated at 28-day intervals. Once the maximum tolerated dose (MTD) was defined the interval between courses was shortened by administering the next course as soon as the patient's neutrophil and platelet counts had recovered to > 1500/microL and > 75,000/microL, respectively. RESULTS: Sixteen patients were treated at 3 dose levels. The MTD was defined as 10 g/m(2)/96 hours of ifosfamide and 800 mg/m(2)/96 hours of etoposide. Thirteen additional patients then were treated at 1 level below the MTD to try and decrease the interval between courses. A total of 12 of 29 patients developed a dose-limiting toxicity (DLT) after the first course of therapy. The most common DLT was gastrointestinal toxicity followed by hematologic toxicity. Twenty-seven patients developed standard National Cancer Institute criteria Grade 3 or 4 toxicity after the first course of treatment and 7 patients achieved a complete or partial response to the first course. The use of GM-CSF did not allow further dose intensification. CONCLUSIONS: This chemotherapy combination achieved a 31% overall response rate. A further increase in the dose intensity of this regimen may require supportive measures other than GM-CSF to decrease toxicity. PMID- 10870070 TI - The risk for cancer among children of women who underwent in vitro fertilization. AB - BACKGROUND: The possible association between ovarian carcinoma and ovulation inducing drugs has led to renewed interest in the potential carcinogenic risks of these drugs. In vitro fertilization (IVF) has been linked to multiple pregnancies and possibly congenital malformations. However, to the authors' knowledge the association between IVF and pediatric cancer has been described only in sporadic case reports. The aim of this study was to assess the incidence rate of pediatric cancer among a cohort of children born after IVF. METHODS: A historic prospective study based on a cohort of 332 children from 1254 women who underwent IVF treatment between 1981-1994 was performed. Medical files were reviewed and names were linked to the National Population and Cancer Registries. Pediatric cancer incidence rates were compared with the expected age-adjusted rates of the general population during the respective time period. RESULTS: No cancer cases were observed among the study cohort with respect to 1.7 cases that were expected. CONCLUSIONS: Because the small cohort analysis in the current study lacked the necessary power to reach definite conclusions, larger prospective studies are needed to assess the potential carcinogenic effect on children born after ovulation induction and IVF. PMID- 10870071 TI - Oral administration of cefixime to lower risk febrile neutropenic children with cancer. AB - BACKGROUND: Febrile neutropenia is a heterogeneous condition. Recently, several risk factors have been defined, permitting the definition of a lower risk group of patients who may benefit form less aggressive therapy. The use of an oral antibiotic approach was tested in the current trial. METHODS: From May 1997 to March 1998, 154 episodes of lower risk febrile neutropenia in 128 children with a mean age of 62 (range, 8-200) months were enrolled in this randomized, single institution trial. Inclusion criteria were fever (> 38 degrees C), neutropenia (absolute neutrophil count < 500/mm(3)), lower risk features (i.e., absence of severe comorbidity factors, good clinical condition, negative blood cultures, control of local infection, no fever during the last 24 hours), and compliance of parents. After 3 days of ceftriaxone (100 mg/kg/day administered intravenously [i.v.]) every 12 hours plus amikacin (15 mg/kg/day i.v.) every 24 hours for 3 days, all patients were discharged and randomized to be allocated to 2 treatment arms. Group A (n = 74) received ceftriaxone cefixime (8 mg/kg/day administered orally) every 24 hours for 4 days, whereas Group B (n = 80) was treated with ceftriaxone plus amikacin for 7 days. Failure was defined as the need for second hospitalization during the same episode of neutropenia, or fever during the 7 days after discharge. RESULTS: Most of the patients (49% in Group A and 55% in Group B) had acute leukemia. Fifty-four (72%) children in Group A and 46 (56%) in Group B had fever of unknown origin (P = not significant [NS]). No significant differences were found in the sites of initial infection between the two groups. Overall results were outstanding, with a favorable outcome in 73 of 78 cases (98.6%) in Group A and 78 of 80 cases (97.5%) in Group B (P = NS). Three patients needed a second hospitalization due to failure of the initial therapy: one in Group A and two in Group B. All three did well with secondary treatment. CONCLUSIONS: In lower risk febrile neutropenic children receiving anticancer therapy, the efficacy of oral cefixime, given for 4 days after 72 hours of intravenous ceftriaxone plus amikacin, was similar to that of 7 days of parenteral ceftriaxone plus amikacin. The oral outpatient therapy approach to the treatment of lower risk febrile neutropenia after chemotherapy is safe and may be cost-saving. This strategy might be adopted as standard therapy in the future. PMID- 10870072 TI - Pleuropulmonary blastoma: A case report documenting transition from type I (cystic) to type III (solid). AB - BACKGROUND: Pleuropulmonary blastoma is a rare, aggressive neoplasm that typically occurs in young children. It has been classified as type I, II, or III on the basis of the cystic versus solid nature of the lesion as well as the histologic appearance. Although it has been speculated that type I lesions may have a tendency to progress into type III lesions, no such case has been reported to date. METHODS: A case of type I pleuropulmonary blastoma in a girl age 2 years 9 months was found in our departmental archive. This case, originally diagnosed as a hamartoma over 20 years ago, predated the description of this pathologic entity. Over a 3-year period, the patient underwent excisions of a primary tumor and 3 subsequent recurrences, thus allowing us to follow the natural history of this neoplasm. RESULTS: The primary tumor was a large, multicystic mass (roughly 90% cystic by volume) with benign histologic appearance except for occasional foci with bland, embryonal rhabdomyosarcomatous features. In subsequent recurrences, the resected specimens became increasingly solid and had an anaplastic, multiphasic mesenchymal pattern. CONCLUSIONS: The course of the patient described here represents the first case in which transition from type I (cystic) to type III (solid) was documented. PMID- 10870073 TI - Clinical utility, factor analysis, and further validation of the memorial delirium assessment scale in patients with advanced cancer: Assessing delirium in advanced cancer. AB - BACKGROUND: Delirium is a common neuropsychiatric complication in patients with advanced cancer. The Memorial Delirium Assessment Scale (MDAS) is a recently developed 10-item severity rating instrument. The purpose of the current prospective study was to further assess the clinical utility, factor structure, and validity of the MDAS in a relatively homogeneous population of patients with advanced cancer. METHODS: Study entry of 104 patients occurred on their consecutive admission to a tertiary-level, acute palliative care unit in a university-affiliated teaching hospital. Patients underwent regular cognitive screening using the Mini-Mental State Examination, and serial monitoring of delirium using standardized semistructured interviews and MDAS ratings, up to the study endpoints of either patient discharge or death. RESULTS: Seventy-one patients met Diagnostic and Statistical Manual (of Mental Disorders)-IV criteria for a first episode of delirium. In 15 of 71 (21%) patients with a first episode of delirium, the first MDAS ratings were prorated because of dyspnea, fatigue, or profound delirium. In the remaining 56 patients (79%), the first MDAS ratings were rated fully and therefore evaluable. Correlations among the scale items ranged from moderate to low (correlation coefficient [r] = 0.68-0.02). Analysis of the pattern of factor loadings identified two primary correlated factors: global cognitive (Factor I) and neurobehavioral (Factor II) (r = 0.33). Cronbach alpha coefficients for Factors I and II were 0.8 and 0.66, respectively, indicating a relatively high level of correlation for items within each. The Cronbach alpha coefficient for all 10 items was 0.78, suggesting a general underlying factor. In a larger sample of complete MDAS ratings (n = 330) a cutoff total MDAS score of 7 of 30 yielded the highest sensitivity (98%) and specificity (96%) for delirium diagnosis. The MDAS was correlated moderately with the Mini Mental State Examination (r = 0.55). CONCLUSIONS: The authors concluded that the MDAS structure is representative of the many features of delirium, broadly grouped as global cognitive and neurobehavioral dimensions. Prorating item scores is necessary in approximately 20% of advanced cancer patients with delirium. This poses potential limitations on the applicability of the MDAS in research. Conversely, the ability to prorate item scores confers a clinical advantage to the instrument when assessing delirium in a patient population with advanced cancer. PMID- 10870074 TI - The schedule of attitudes toward hastened death: Measuring desire for death in terminally ill cancer patients. AB - BACKGROUND: The authors examined the reliability and validity of the Schedule of Attitudes toward Hastened Death (SAHD), a self-report measure of desire for death previously validated in a population of individuals with the acquired immunodeficiency syndrome (AIDS), among terminally ill patients with cancer. METHODS: The authors interviewed 92 terminally ill cancer patients, all with a life expectancy of < 6 months, after admission to a palliative care hospital. Patients were administered the SAHD, a clinician-rated measure of desire for death (the Desire for Death Rating Scale [DDRS]), and several measures of physical and psychosocial well-being. RESULTS: The average number of SAHD items endorsed was 4. 76 (standard deviation, 4.3); 15 patients (16.3%) endorsed > or = 10 items, indicating a high desire for death. Internal consistency was strong (coefficient alpha = 0.88, median item-total correlation = 0. 49), as were indices of convergent validity. Total SAHD scores were correlated significantly (correlation coefficient [r] = 0.67) with the DDRS, and somewhat less so with measures of depression (r = 0. 49) and hopelessness (r = 0.55). Lower, but substantial, correlations were observed between the SAHD and measures of spiritual well-being (r = -0.42), quality of life (r = -0.36), physical symptoms (r = 0.38), and symptom distress (r = 0.38). No significant correlation was observed between SAHD scores and social support (r = -0.06) or pain intensity (r = 0.16); however, pain-related functional interference and overall physical functioning were correlated significantly with SAHD scores (r = 0.31 and r = 0.23, respectively). CONCLUSIONS: The SAHD appears to be a reliable and valid measure of desire for death among terminally ill cancer patients. Coupled with previous research in patients with AIDS, these results support the utility of the SAHD for research addressing interest in hastened death in patients with a life threatening medical illness. PMID- 10870075 TI - Purchasing oncology services. Kerr L. White Institute/American Cancer Society Task Force on Purchasing Oncology Services. AB - BACKGROUND: A multidisciplinary panel representing various stakeholders in the health care delivery and oncology services marketplace was convened to develop specific criteria for healthcare purchasers to consider when evaluating the structures and processes of health plans. These rank ordered criteria also can be used by oncologic service providers and health plan designers as a yardstick for the services they offer. METHODS: A multidisciplinary 31-member Task Force was assembled by the Kerr L. White Institute and the American Cancer Society in March 1997. Task Force members were selected for their ability to offer expert insight as purchasers, suppliers, policymakers, consumers, or stakeholders in the health care marketplace. A preference-weighted majority voting rule was used to identify the three most important recommendations of the 10 that were generated through a modified Delphi technique. To test the practicality of the top three recommendations, leaders of large managed care organizations (MCOs) were surveyed; the results of this survey then were compared with the results of the Task Force survey. RESULTS: The three most important recommendations from the Task Force were that health plans provide access to: 1) comprehensive cancer care, 2) preventive and screening services, and 3) second opinions and treatment options supported by scientific evidence. The difference between the responses of the Task Force and the MCOs was that MCOs placed the highest importance on evidence-based decision-making, with their next three rankings coinciding with those identified by the Task Force. CONCLUSIONS: The value of these summary recommendations will be realized through their use by both purchasers and suppliers to influence the structure and content of the delivery of oncologic services. PMID- 10870076 TI - World Health Organization classification of tumors. PMID- 10870078 TI - Localization of epidermal growth factor receptors and putative neuroblasts in human subependymal zone. AB - Studies in rodents and monkeys suggest that neuronal precursor cells continue to exist and differentiate well into adulthood in these species. These results challenge the long held assumption that neurogenesis does not occur in the postnatal human brain. We examined the rostral subependymal zone (SEZ) of postnatal human brain for expression of cell phenotypic markers that have been associated with neuronal precursors and neuroblasts in rodent brain. We found epidermal growth factor receptor (EGF-R) mRNA and protein to be expressed in infant, teen, young adult, and adult human SEZ. Some SEZ cells expressed the polysialic acid form of neural cell adhesion molecule (PSA-NCAM), characteristic of migrating neuroblasts, as well as class III beta-tubulin and Hu protein, characteristic of neuroblasts and early neurons. These neuroblast-like cells were negative for glial fibrillary acidic protein (GFAP), 2;,3;-cyclic nucleotide 3; phosphohydrolase (CNPase), and vimentin, suggesting that they were not differentiating as glia. Our results show that neuroblast-like cells exist in the human SEZ and support the theory that SEZ of postnatal human brain has neurogenic potential. PMID- 10870079 TI - Fetuin in the developing neocortex of the rat: distribution and origin. AB - Immunocytochemical distribution of the fetal protein fetuin in the neocortex of developing rat brain and the presence of its mRNA, as detected by using reverse transcriptase-polymerase chain reaction analysis, was studied in fetuses at embryonic day 15 (E15) through E22, in neonates at postnatal day 0 (P0) through P20, and in adults. Quantitative estimates of fetuin in cerebrospinal fluid (CSF) and plasma were obtained over the same period. Exogenous (bovine) fetuin injected intraperitoneally into fetal and postnatal rats was used to study the uptake of fetuin into CSF and brain and its distribution compared with endogenous fetuin; bovine albumin was used as a control. Fetuin was identified immunocytochemically in the cortical plate and subplate cells of the developing neocortex. In the rat fetus, fetuin first was apparent at E17, mainly in cell processes, but a few subplate cells also were positive. By E18, there was strong staining in subplate neurons and in inner cells of the cortical plate. At E21, these inner cells of the cortical plate were beginning to differentiate into layer VI neurons, many of which were positive for fetuin. By P0-P1, more layer VI neurons and some layer V neurons had become positive for fetuin. Fetuin immunoreactivity generally was weaker at P1, and, by P2-P3, it had disappeared from all of the layers of the developing neocortex. Bovine fetuin (but not albumin), probably taken up through CSF over the neocortical dorsal surface, had a cytoplasmic distribution; endogenous rat fetuin was both cytoplasmic and membrane bound. Thus, much of this fetuin can be accounted for by uptake, although the presence of fetuin mRNA indicates that in situ synthesis may also contribute. PMID- 10870080 TI - Immunocytochemical evidence that collision sensing neurons in the locust visual system contain acetylcholine. AB - The lobula giant movement detector (LGMD1 and -2) neurons in the locust visual system are parts of motion-sensitive pathways that detect objects approaching on a collision course. The dendritic processes of the LGMD1 and -2 in the lobula are localised to discrete regions, allowing the dendrites of each neuron to be distinguished uniquely. As was described previously for the LGMD1, the afferent processes onto the LGMD2 synapse directly with each other, and these synapses are immediately adjacent to their outputs onto the LGMD2. Here we present immunocytochemical evidence, using antibodies against choline-protein conjugates and a polyclonal antiserum against choline acetyltransferase (ChAT; Chemicon Ab 143), that the LGMD1 and -2 and the retinotopic units presynaptic to them contain acetylcholine (ACh). It is proposed that these retinotopic units excite the LGMD1 or -2 but inhibit each other. It is well established that ACh has both excitatory and inhibitory effects and may provide the substrate for a critical race in the LGMD1 or -2, between excitation caused by edges moving out over successive photoreceptors, and inhibition spreading laterally resulting in the selective response to objects approaching on a collision course. In the optic lobe, ACh was also found to be localised in discrete layers of the medulla and in the outer chiasm between the lamina and medulla. In the brain, the antennal lobes contained neurons that reacted positively for ACh. Silver- or haematoxylin and eosin stained sections through the optic lobe confirmed the identities of the positively immunostained neurons. PMID- 10870081 TI - Localization of mGluR6 to dendrites of ON bipolar cells in primate retina. AB - We prepared antibodies selective for the C-terminus of the human mGluR6 receptor and used confocal and electron microscopy to study the patterns of immunostaining in retina of monkey, cat, and rabbit. In all three species punctate stain was restricted to the outer plexiform layer. In monkey, stain was always observed in the central element of the postsynaptic "triad" of rod and cone terminals. In monkey peripheral retina, stain was seen only in central elements, but in the fovea, stain was also observed in some dendrites contacting the base of the cone terminal. S-cone terminals, identified by staining for S opsin, showed staining of postsynaptic dendrites. These were identified as dendrites of the ON S-cone bipolar cell by immunostaining for the marker cholecystokinin precursor. The staining pattern suggests that all types of ON bipolar cells, despite their marked differences in function, express a single isoform of mGluR6. Ultrastructurally, mGluR6 was located not on the tip of the central element, near the site of vesicle release, but on its base at the mouth of the invagination, 400-800 nm from the release site. Thus, the mGluR6 receptors of ON bipolar cells lie at about the same distance from sites of vesicle release as the iGluR receptors of OFF bipolar cells at the basal contacts. PMID- 10870082 TI - Neural pathways for bilateral vocal control in songbirds. AB - Ipsilateral and contralateral projections of nucleus robustus archistriatalis (RA), a telencephalic vocal premotor nucleus, to respiratory-vocal nuclei in the brainstem were defined in adult male Wasserschlager canaries, grey catbirds, and zebra finches, three songbird species that appear to differ in the degree of lateralized syringeal dominance. In all three species, ipsilateral projections of RA to the medulla included the tracheosyringeal part of the hypoglossal nucleus (XIIts), that innervates the syrinx, the bird's vocal organ, the suprahypoglossal area (SH), and two respiratory-related nuclei, retroambigualis (RAm) and parambigualis (PAm; Reinke and Wild [1998] J Comp Neurol 391:147-163). Projections of RA to the contralateral XIIts, SH and RAm, were substantial in canaries, which use the left side of the syrinx predominantly during singing; less pronounced in catbirds, which have no lateral dominance for song control; and least pronounced in zebra finches, in which there is a right-sided dominance for song control. There were no obvious differences in the number of crossed projections in birds injected in the left or right RA. Local sources of inputs to XIIts and RAm were defined anatomically in zebra finches and canaries. RAm, including neurons in close proximity to XIIts, was found to project to XIIts and the suprahypoglossal area bilaterally but predominantly ipsilaterally. RAm also had reciprocal connections with its contralateral homologue. These results suggest a pattern of connections between premotor and motor respiratory-vocal nuclei that may be involved in bilateral control of vocal output at medullary levels, a control that involves a high degree of coordination between vocal and respiratory structures on both sides of the body. PMID- 10870083 TI - Parvocells: a novel interneuron type in the pacemaker nucleus of a weakly electric fish. AB - Gymnotiform weakly electric fish produce electric organ discharges (EODs) that function in electrolocation and communication. The command signal for the EOD is produced by the medullary pacemaker nucleus, which contains two well characterized neuron types: pacemaker cells and relay cells. In this study, we characterized a third neuron type in the pacemaker nucleus. These neurons, which we have named parvocells, were smaller (7-15 microm in diameter) than relay and pacemaker cells. The parvocells were labeled with an antibody against the neuronal calcium-binding protein, parvalbumin, and were not labeled with several glial-specific antibodies. Parvocells had one to three fine processes that often terminated at the periphery of relay and pacemaker cell bodies. The parvalbumin positive terminals of the parvocells colocalized with immunoreactivity for SV-2, suggesting that the parvocells form chemical synapses on the relay and pacemaker cells. Parvalbumin-positive neurons are frequently gamma-aminobutyric acid (GABA)ergic or glycinergic, and the cytoplasm of the parvocell somata was immunoreactive with a glycine antibody. Antibodies against glycine receptors and gephyrin, however, did not label any cells in the pacemaker nucleus, suggesting that the pacemaker nucleus does not contain glycine or GABA((A)) receptors. Electron microscopy revealed gap junctions between the membranes of parvocells and adjacent terminal-like structures. Furthermore, neurobiotin injected into individual pacemaker or relay cells labeled parvocells as well as other pacemaker and relay cells, demonstrating that the parvocells are dye-coupled to the other neuron types in the pacemaker nucleus. These findings indicate that the parvocells are histochemically distinct from relay and pacemaker cells and that they receive electrotonic inputs from and make chemical synapses back onto pacemaker and relay cells. Further study is needed to investigate the function of these neurons in regulating the EOD. PMID- 10870084 TI - Immunocytochemical localization of the glutamate transporter GLT-1 in goldfish (Carassius auratus) retina. AB - Glutamate is the major excitatory neurotransmitter in the retina of vertebrates. Electrophysiological experiments in goldfish and salamander have shown that neuronal glutamate transporters play an important role in the clearance of glutamate from cone synaptic clefts. In this study, the localization of the glutamate transporter GLT-1 has been investigated immunocytochemically at the light and electron microscopical levels in the goldfish retina using a GLT-1 specific antibody. GLT immunoreactivity (IR) was observed at the light microscopical level in Muller cells, bipolar cells, the outer plexiform layer (OPL), and the inner plexiform layer (IPL). At the electron microscopical level, membrane-bound and cytoplasmic GLT-IR in the OPL was located in finger-like protrusions of the cone terminal located near the invaginating postsynaptic processes of bipolar and horizontal cells. GLT-IR was not observed in the vicinity of synaptic ribbons. This location of GLT-1 allows modulation of the glutamate concentration in the synaptic cleft, thereby shaping the dynamics of synaptic transmission between cones and second-order neurons. In the inner IPL, GLT-IR was observed in the cytoplasm and was membrane bound in mixed rod/cone bipolar cell terminals and cone bipolar cell terminals. The membrane-bound GLT-1 was generally observed at some distance from the synaptic ribbon. The morphology of the bipolar cell terminal together with the localization of GLT-1 suggests that at least these glutamate transporters are not primarily involved in rapid uptake of glutamate release by the bipolar cells. The GLT-IR in the cytoplasm of Muller cells was located throughout the entire goldfish retina from the outer limiting membrane to the inner limiting membrane. The location of GLT-1 in Muller cells is consistent with the role of Muller cells in converting glutamate to glutamine. PMID- 10870085 TI - Nitric oxide/cyclic GMP pathway attenuates ATP-evoked intracellular calcium increase in supporting cells of the guinea pig cochlea. AB - We demonstrate here that nitric oxide (NO) attenuates ATP-evoked calcium transients in Deiters' and Hensen's cells, "supporting" (nonsensory) cells of the guinea pig cochlea, by means of activation of soluble guanylyl cyclase and protein kinase G. The enzymatic activities associated with the nitric oxide/cGMP/protein kinase G pathway had previously been demonstrated to be present in Deiters' and Hensen's cells. We now isolate these cells and measure changes in intracellular free calcium by using the calcium indicator fluo-3. In Deiters' cells, calcium increased rapidly in response to the application of ATP. The increase was attenuated when the pathway was stimulated by NO donors (diethylamine NONOate or sodium nitroprusside) or the cyclic GMP analog, 8-bromo cyclic GMP. When the activation of the pathway was blocked by the additional presence of inhibitors of soluble guanylyl cyclase (LY83583) or protein kinase G (Rp-8-bromo-cyclic GMP or KT5823), the response to ATP was restored. The reactions also occurred in calcium-free media. Hensen's cells responded similarly. These results provide evidence that intracellular calcium is regulated by the NO/cGMP/protein kinase G pathway in the inner ear. PMID- 10870086 TI - Differential expression of neurotrophin and neurotrophin receptor mRNAs in and adjacent to fetal midbrain grafts implanted into the dopamine-denervated striatum. AB - This study examined the expression of neurotrophins and neurotrophin receptors in the lesion/transplanted striatum at four different time points after transplantation. The ventral mesencephalic region was dissected from a single rat fetus at embryonic day 14 (E14) and implanted into the denervated striatum of rats with unilateral 6-hydroxydopamine lesions. Transplanted rats were killed at 1, 2, 3, or 4 weeks after transplantation surgery and the brains subsequently prepared for semiquantitative in situ hybridization analysis of neurotrophin and neurotrophin trk receptors. Hybridization of cRNA probes for trkB or trkC showed a time-dependent reduction within the transplant during the first 4 weeks after transplantation; hybridization of brain-derived neurotrophic factor or tyrosine hydroxylase mRNA probes within the transplant did not change significantly during the same posttransplantation period. Hybridization of the trkB mRNA probe in host striatum adjacent to the transplant was significantly higher than probe hybridization in the corresponding region of the intact striatum during the first 2 weeks after transplantation, but by the 3rd and 4th week, probe hybridization in the denervated/transplanted and intact striatum were the same. Lesioned animals without transplants maintained higher trkB mRNA probe hybridization in the denervated striatum than in the intact striatum at the same postlesion time points suggesting that lesioned/transplanted animals show a normalization of trkB mRNA probe hybridization. Hybridization of the trkC mRNA probe in the lesioned/transplanted striatum was significantly lower than that observed in the intact striatum 4 weeks after transplantation; however, at this same time point we observed a similar reduction of trkC probed hybridization in lesioned animals without transplants. The results of the study show dynamic neurotrophic activity occurring within the transplant and host tissue during the first month of transplant development. PMID- 10870087 TI - Connections of some auditory-responsive posterior thalamic nuclei putatively involved in activation of the hypothalamo-pituitary-adrenocortical axis in response to audiogenic stress in rats: an anterograde and retrograde tract tracing study combined with Fos expression. AB - Prior studies in our laboratory demonstrated that part of the thalamus is necessary for activating the hypothalamo-pituitary-adrenocortical (HPA) axis in response to audiogenic stress in rats. The present studies were designed to determine how the auditory-responsive thalamic nuclei might activate the HPA axis. Both retrograde [Fluoro-Gold (FG)] and anterograde [Phasoleus vulgaris leucoagglutinin (PHA-L) and biotinylated dextran amines (BDA)] tracers were employed to study the putative connectivity between the thalamus and the medial parvocellular region of the hypothalamic paraventricular nucleus (PAmp). In addition, rats receiving FG in the PAmp were subjected to audiogenic stress, and the distribution of both FG and the protein product of the immediate-early gene c fos, Fos, were determined by double immunohistochemistry, to help assess putative functional links between the auditory-responsive thalamic nuclei and PAmp. The results of PAmp FG placement indicated retrogradely labeled cells in several areas, including the bed nucleus of the stria terminalis, hypothalamic regions, the supramammillary nucleus, some thalamic regions, and importantly, a few multisensory nuclei of the thalamus, including the parvicellular division of the subparafascicular and posterior intralaminar nuclei. Injections of the tracers PHA-L or BDA into these auditory-responsive posterior thalamic nuclei provided further evidence of projections to the PAmp. In addition, several forebrain areas were observed to receive moderate to heavy innervation. These areas included most of the regions described above, which, in turn, project to the PAmp. Because cells in the multisensory thalamic nuclei, hypothalamic, and forebrain areas were double labeled with FG and Fos, the results suggest that either direct projections from the thalamus to PAmp neurons, or indirect projections from the thalamus to stress-responsive forebrain areas projecting to the PAmp, might mediate activation of the HPA axis by audiogenic stress. PMID- 10870088 TI - Peripheral, but not central, axotomy induces neuropilin-1 mRNA expression in adult large diameter primary sensory neurons. AB - Neuropilin-1 (NP-1) is a component of the receptor for semaphorin3a (Sema3a), a member of a large family of molecules with widespread expression and demonstrable influence (via their ability to repel growing axons) on nervous system development. Recent studies have shown that some types of adult mammalian neurons retain the capacity to respond to Sema3a, particularly in relation to neuronal injury and regeneration. Although variations in expression of Sema3a mRNA have been revealed in neurons in both the central and peripheral nervous systems in this context, relatively little is known about NP-1 expression patterns. In this study we investigated the expression of NP-1 mRNA in adult dorsal root ganglion (DRG) neurons in intact and lesioned animals. We compared the effect of unilateral lesioning of the sciatic nerve or unilateral dorsal rhizotomy at lumbar levels L4/5, and bilateral dorsal funiculus lesioning at thoracic levels T10/11 on NP-1 mRNA expression in the cell bodies of lumbar DRGs. A significantly increased level of NP-1 mRNA expression was detected only following sciatic nerve lesioning (P < 0.001), but not after rhizotomy or dorsal funiculus lesioning. Furthermore, this upregulation was mainly confined to large diameter neurons of DRGs at lumbar levels L4/5, which provide the main sensory contribution to the sciatic nerve. These results suggest a role for NP-1 in the axonal response to peripheral nerve injury, which may be specific to a particular subset of primary sensory neurons. PMID- 10870089 TI - Neurokinin 1 receptor distribution in cholinergic neurons and targets of substance P terminals in the rat nucleus accumbens. AB - Substance P (SP) is the major endogenous ligand for neurokinin 1 (NK1) receptors and, together with acetylcholine, has an important role in motivated behaviors involving the limbic shell and motor core of the nucleus accumbens (NAc). To determine the functional sites for SP activation of NK-1 receptors and potential interactions with cholinergic neurons in these regions, the authors examined the electron microscopic immunocytochemical localization either of antisera against the NK1 receptor or of the NK1 receptor and either 1) SP or 2) the vesicular acetylcholine transporter (VAchT) in rat NAc. In both the NAc shell and core, NK1 receptor labeling was localized mainly to somatic and dendritic plasma membranes and nearby endosomal organelles in aspiny neurons. In sections through the ventromedial shell that were processed for NK1/SP labeling, 46% of the NK1 immunoreactive dendrites (n = 603 dendrites) showed symmetric or appositional contacts with SP-containing terminals. These terminals and several others that formed symmetric synapses also occasionally were immunoreactive for NK1 receptors. Analysis of the shell region for NK1/VAchT labeling showed that 61% of the total immunoreactive dendrites (n = 534 dendrites) contained NK1 receptors without VAchT, 29% contained both products, and 10% contained VAchT only. Many of the labeled somata and dendrites also received synaptic contact from VAchT containing terminals. These findings suggest that, in the NAc, NK1 receptors are recycled through endosomal compartments and play a role in modulating mainly the postsynaptic responses, but also the presynaptic release, of SP and/or inhibitory neurotransmitters onto aspiny interneurons, some of which are cholinergic. PMID- 10870090 TI - Xenopus laevis peripherin (XIF3) is expressed in radial glia and proliferating neural epithelial cells as well as in neurons. AB - Neuronal intermediate filament (nIF) proteins form the most abundant component of the axonal cytoskeleton. Thus, understanding their function and the regulation of their expression is essential for comprehending how axonal structure is regulated. Although most vertebrate nIF proteins are classified as type IV intermediate filament (IF) proteins, additional nIF proteins exist in frogs (Xenopus laevis), cyprinid fishes, and mammals (called XIF3, plasticin, and peripherin, respectively) that are classified as type III. Expression of a type III nIF protein is correlated strongly with the earliest phases of axonal outgrowth in fishes but less so in mammals. To understand better how the correlation between type III nIF protein expression and early phases of axonal outgrowth has changed during evolution, the authors examined XIF3 expression in Xenopus laevis. In Xenopus, the association between XIF3 expression and early axonal outgrowth was especially strong. For example, during early axonal development, XIF3 expression preceded and was more abundant and widespread than that of any of the type IV nIF proteins. As axons matured, neuronal expression of XIF3 gradually became more restricted while that of type IV nIF proteins increased. These results support the idea that type III nIF proteins play a special role during early phases of axonal outgrowth. In addition to finding XIF3 in neurons, the authors also unexpectedly found it in regions of the central nervous system that contain proliferating cells and radial glia. As a framework for interpreting variations in nIF expression in different vertebrate species, the authors built phylogenetic trees to clarify relationships among vertebrate nIF proteins. These trees supported the classification of XIF3, plasticin, and peripherin as orthologs (products of the same genetic locus, evolving separately only since the species lineages diverged). Thus, XIF3, plasticin, and peripherin probably should be referred to as Xenopus, fish, and mammalian peripherin, respectively. This finding argues that differences in expression of these three proteins in frogs, fishes, and mammals are the result of regulatory changes to the peripherin ancestral gene along each lineage. The expression of a peripherin ortholog in Xenopus glia may represent either an adaptation that arose since the divergence of Xenopus from mammals or, alternatively, a feature retained from an ancestral IF protein that was expressed originally both in neurons and in glia. PMID- 10870091 TI - Postnatal development of efferent synapses in the rat cochlea. AB - The development of olivocochlear efferent axons and their contacts in the postnatal cochlea was studied after DiI applications to the olivocochlear bundle in the ipsilateral brainstem of rats from 0 to 10 days of age (P0-10). Light microscopic analyses showed that labeled axons reached the vicinity of inner hair cells by P0 and outer hair cells by P2. Electron microscopic analyses demonstrated that labeled immature efferent axons are present among supporting cells of the greater epithelial ridge as well as inner hair cells at P0. The first efferent contacts that contacted inner hair cells contained a few irregularly sized vesicles and, occasionally, mitochondria. Postsynaptic specializations within inner hair cells apposed to labeled efferent axons included subsynaptic cisterns, irregularly sized vesicles, and synaptic bodies. Similar features were present in unlabeled profiles, presumed to be afferents, indicating that immature efferent axons could not be reliably distinguished from afferents without positive labeling. Efferent axons synapsed with outer hair cells by P4 and had synapse-like contacts at the bases of Deiters' cells at P4 and P6. Contacts between afferents and efferents were observed frequently in the inner spiral bundle from P6. As they matured, efferent axon terminals contacting hair cells contained increasing numbers of synaptic vesicles and were typically apposed by well-defined postsynaptic cisterns, thus acquiring distinctive profiles. PMID- 10870092 TI - Binding and transport of methotrexate and its derivative, MX-68, across the brush border membrane in rat kidney. AB - Binding and transport properties of methotrexate (MTX) and its novel derivative, MX-68, were examined in brush-border membrane vesicles (BBMVs) isolated from rat kidneys. The uptake of MTX, MX-68 and folic acid by BBMVs was stimulated by an inwardly-directed H(+) gradient. Such H(+)-dependent uptake of folic acid is compatible with a previous report (Bhandari et al., Biochim Biophys Acta 1988; 937: 211). The MTX uptake exhibits saturation with a K(m) of 0.834 microM. Although the uptake of these three compounds at optimal pH depended on the osmolarity of the medium, a substantial portion of the uptake was osmolarity insensitive. By changing the medium osmolarity, the uptake by BBMVs could be separately discriminated as osmolarity-sensitive and insensitive portions, representing transport into the intravesicular space and binding to the surface of BBMVs, respectively. For all three compounds, the binding increased in a time dependent manner, while the amount transported reached a maximum after a relatively short incubation period. The transport of folic acid, but not its binding, exhibited an overshoot phenomenon under conditions of an inward H(+) gradient. The present results suggest that reabsorption of MTX and MX-68 in the kidney is governed by both their binding and transport mechanisms, with a similar kinetic profile to that of folic acid. The involvement of a transport system seems to make a relatively small contribution to the reabsorption of MTX assessed in BBMVs compared with MX-68 and folic acid. PMID- 10870093 TI - Pharmacokinetics and tissue distribution of olanzapine in rats. AB - The single dose pharmacokinetics of olanzapine in rats, following an oral dose and its distribution in the brain and other tissues after repeated oral and intra peritoneal (i.p.) administration, were studied. Olanzapine in plasma, brain, liver, lung, kidney, spleen and fat was assayed at predose, 0.25, 0.5, 1, 2, 5, 12, 24, 36, 48 h postoral dose of 6 mg/kg and after daily oral and i.p. doses of 0.25, 1, 3, and 6 mg/kg/day of olanzapine for 15 consecutive days by a sensitive and specific HPLC method with electrochemical detection. Olanzapine was readily absorbed and distributed in plasma and tissues as the peak concentrations were reached within approximately 45 min after the oral dose. The terminal half-life of olanzapine in plasma was 2.5 h and in tissues it ranged from 3 to 5.2 h. The area under the concentration-time curve (AUC(last)) was lowest in plasma and largest in liver and lung. The AUC(last) of olanzapine was eight times larger in brain and three to 32 times larger in other tissues than that in plasma. After repeated oral doses, the plasma and tissue concentrations of olanzapine were generally higher than those after repeated i.p. doses. The liver and spleen had the highest concentrations after oral and i.p doses, respectively. In both cases, the tissue concentrations were four- to 46-fold higher than that in plasma and correlated with administered doses. Likewise, plasma concentrations strongly correlated with the simultaneous brain and tissue concentrations (r(2)>0.908, p<0.0001). On average, the brain levels were 6.3-13.1 and 5.4-17.6 times higher than the corresponding plasma level after oral and i.p. doses, respectively. The tissue to plasma level ratio of olanzapine was higher in other tissues. The data indicated that olanzapine is rapidly absorbed and widely distributed in the tissues of rats after oral and i.p. administration. The plasma concentration appears to predict the simultaneous concentration in brain and other tissues. There was no marked localized accumulation of olanzapine in any of the regions of the rat brain. PMID- 10870094 TI - Pharmacokinetics and tissue distribution of betulinic acid in CD-1 mice. AB - Betulinic acid is a naturally occurring pentacyclic triterpenoid. Betulinic acid has recently been selected by the National Cancer Institute for addition into the RAID (Rapid Access to Intervention in Development) programme. The agent exhibits potential anti-tumour activity and functions in this regard via apoptosis. The objective of the present study was to determine the pharmacokinetics of betulinic acid in CD-1 mice. Serum samples were obtained at designed times after a single 250 or 500 mg/kg intraperitoneal (IP) dose of betulinic acid. Tissue samples (skin, heart, liver, spleen, kidney, lung, brain, colon, caecum, ovary, uterus, thymus, lymph node, bladder, perirenal fat, mammary gland and small intestine) were collected after betulinic acid administration (500 mg/kg). Betulinic acid was extracted with methylene chloride and quantitatively analysed by HPLC/MS. Oleanolic acid and madecassic acid were used as internal standards. Pharmacokinetic parameters were calculated using the WinNonlin pharmacokinetic software package. A two-compartment, first-order model was selected for pharmacokinetic modelling. The results showed that after IP 250 and 500 mg/kg betulinic acid, the serum concentrations reached peaks at 0.15 and 0.23 h, respectively. The 250 and 500 mg/kg above betulinic acid IP doses were found to have elimination half-lives of 11.5 and 11.8 h and total clearances of 13.6 and 13.5 L/kg/h, respectively. The pharmacokinetic parameters observed for IP betulinic acid 500 mg/kg in the skin of mice were as follows: k(a) (h(-1)) 0.257, k(10) (h(-1)) 0.234, t(1/2(alpha)) (h) 2.63, t(1/2(beta)) (h) 20.2, V (L/kg) 0.61, AUC (microg/h/mL) 3504, T(max) (h) 3.90 and C(max) (microg/mL) 300.9. The distribution of betulinic acid in tissues at 24 h post-IP administration in a descending order was as follows: perirenal fat, ovary, spleen, mammary gland, uterus, bladder, lymph node, liver, small intestine, caecum, lung, thymus, colon, kidney, skin, heart and brain. PMID- 10870095 TI - Inhibition of human cytochrome P450 isoforms in vitro by zafirlukast. AB - Zafirlukast is a cysteinyl leukotriene antagonist used to treat allergic and exercise-induced asthma. This in vitro study used human liver microsomes to evaluate the inhibitory activity of zafirlukast versus six human cytochrome P450 (CYP) isoforms. Zafirlukast (0-250 microM) was co-incubated with fixed concentrations of index substrates. Zafirlukast inhibited the hydroxylation of tolbutamide (CYP2C9; mean IC(50)=7.0 microM), triazolam (CYP3A; IC(50)=20.9 microM) and S-mephenytoin (CYP2C19; IC(50)=32.7 microM), and was a less potent inhibitor of phenacetin O-deethylation (CYP1A2; IC(50)=56 microM) and dextromethorphan O-demethylation (CYP2D6; IC(50)=116 microM). Zafirlukast produced negligible inhibition of CYP2E1. In vitro inhibition of CYP2C9 by zafirlukast is consistent with clinical studies showing impaired clearance of S warfarin and enhanced anti-thrombotic effects, although the in vitro IC(50) value is higher than the usual range of clinically relevant plasma concentrations. Zafirlukast deserves further clinical study as an inhibitor of other CYP2C9 substrates such as nonsteroidal anti-inflammatory agents, tolbutamide, phenytoin and mestranol. Clinically important inhibition by zafirlukast of other CYP isoforms is not established. PMID- 10870097 TI - Nonlinear pharmacokinetics of L-N(G)-methyl-arginine in rats: characterization by an improved HPLC assay. AB - L-N(G)-methyl-arginine (L-NMMA) is an inhibitor of nitric oxide synthase (NOS) enzymes. We have characterized the pharmacokinetics of L-NMMA in rats using HPLC. The HPLC assay requires pre-column derivatization, gradient elution and ultraviolet detection. The limit of sensitivity in plasma was 3.0 microM (0.75 microg mL(-1)). Using this assay, the pharmacokinetics of L-NMMA were characterized following iv bolus doses of 25, 50 and 100 mg kg(-1). Compartmental and noncompartmental data analysis suggest that L-NMMA pharmacokinetics are nonlinear at these doses. From the nonlinear compartmental analysis, we estimated the K(m) and V(max) parameters of L-NMMA elimination to be 70.2 microM and 4.59 microM min(-1), respectively. This estimated K(m) value of L-NMMA elimination is consistent with its nonlinear elimination characteristics in humans and its saturable metabolism by the N(G), N(G)-dimethylarginine dimethylamino-hydrolase enzyme in isolated rat tissue. PMID- 10870096 TI - Additive effect of indomethacin and methotrexate on suppression of growth in rats. AB - The purpose of this study was to investigate the medium-term effects of methotrexate (MTX) and indomethacin on the growth of young rats. Four equal groups of Sprague-Dawley male rats (four animals in each group; mean+/-S.D. body weight, 183+/-13 g, in their rapid growth phase) were subjected to the following drug treatment: one group was given MTX (0.2 mg kg(-1) body weight) subcutaneously on every fourth day, another received indomethacin (2.5 mg kg(-1) body weight) subcutaneously daily and the third group was given both of these drugs (MTX on every fourth day and indomethacin daily). The fourth group was injected subcutaneously with physiological saline every day to serve as a control group. Total body weight, food and water consumption by animals in each group were monitored every second day for a period of 10 weeks. After this period, liver, spleen and kidneys were excised, weighed and analysed for MTX and dihydrofolate reductase activity. Compared with the groups, which received MTX alone, indomethacin alone, or physiological saline, mean increase (17+/-11 g) in body weight of rats was minimal in the group receiving both MTX and indomethacin. The difference was statistically significant (p=0.001) when the values of mean increase in body weight of rats in different treatment groups after a 10-week treatment were compared. The mean weights of liver and spleen in this group receiving both MTX and indomethacin were also found to be significantly less than the weights of these organs in the control group (p<0.01). There also appears to be a decline in food consumption in this group (p<0.05). This negative effect on growth of animals in this group appears to be not only due to decreased food consumption but also due to increased inhibition of de novo pathway of DNA synthesis. This is supported by increased accumulation of MTX and decreased dihydrofolate reductase activity in this group receiving both MTX and indomethacin, as compared with the group receiving MTX alone. The data indicate an additive effect of MTX and indomethacin on the suppression of growth in young rats, alluding to the notion that patients suffering from juvenile rheumatoid arthritis or acute lymphoblastic leukaemia receiving these two drugs concomitantly over a long period of time might be at a risk of experiencing short term suppression of growth. PMID- 10870098 TI - Epinephrine absorption after different routes of administration in an animal model. AB - BACKGROUND: The administration of epinephrine is the most important initial treatment in systemic anaphylaxis. PURPOSE: We wanted to determine the relationship between the route of epinephrine administration and the rate and extent of epinephrine absorption. METHODS: In a prospective, randomized, five-way crossover study in rabbits, we measured plasma epinephrine concentrations before, and at intervals up to 180 min after epinephrine administration by intramuscular or subcutaneous injection, or by inhalation, with intravenous epinephrine and intramuscular saline as the positive and negative controls, respectively. RESULTS: Maximum plasma epinephrine concentrations were higher, and occurred more rapidly, after intramuscular injection than after subcutaneous injection or inhalation, and were 7719+/-3943 (S.E.M.) pg/mL at 32.5+/-6.6 min, 2692+/-863 pg/mL at 111.7+/-30.8 min and 1196+/-369 pg/mL at 45. 8+/-19.2 min, respectively. Intravenous injection of epinephrine resulted in a plasma concentration of 3544+/ 422 pg/mL at 5 min, and an elimination half-life (t(1/2)) of 11.0+/-2.5 min. In the saline control study, the endogenous epinephrine concentration peaked at 518+/-142 pg/mL. CONCLUSION: In this model, absorption of epinephrine was significantly faster after intramuscular injection than after subcutaneous injection or inhalation. The extent of absorption was satisfactory after both intramuscular and subcutaneous injections. Neither the rate nor the extent of absorption was satisfactory after administration by inhalation. PMID- 10870099 TI - SSU1 mediates sulphite efflux in Saccharomyces cerevisiae. AB - Ssu1p, a plasma membrane protein involved in sulphite metabolism in Saccharomyces cerevisiae, was found to be required for efficient sulphite efflux. An SSU1 null mutant accumulated significantly more sulphite than wild-type, whereas cells expressing multicopy SSU1 accumulated significantly less. Cells expressing FZF1 4, a dominant allele of a transcriptional activator of SSU1 that confers sulphite resistance, also accumulated less sulphite. beta-galactosidase activity in the FZF1-4 strain carrying an SSU1::lacZ fusion was found to be 8.5-fold higher than in a strain carrying wild-type FZF1, confirming that the heightened resistance was correlated with hyperactivation of SSU1. Multicopy SSU1 was also found to increase the sulphite resistance of a number of unrelated sulphite-sensitive strains by a factor of 3- to 8-fold. Rates of efflux of free sulphite from cells expressing multicopy SSU1 or FZF1-4 were significantly greater than that from wild-type or from a SSU1 null mutant. Rates of efflux of bound sulphite from wild type, a SSU1 null mutant, a FZF1-4 mutant, or cells expressing multicopy SSU1 were not significantly different, suggesting that Ssu1p specifically mediates efflux of the free form of sulphite. PMID- 10870100 TI - Cryopreservation of competent intact yeast cells for efficient electroporation. AB - We have developed a simple method for cryopreserving Schizosaccharomyces pombe and Saccharomyces cerevisiae competent intact cells that permits high transformation efficiency and long-term storage for electroporation. Transformation efficiency is significantly decreased if intact cells are frozen in common permeating cryoprotectants such as glycerol or dimethyl sulphoxide. On the other hand, we found that a high transformation efficiency could be maintained if the cells were frozen in a non-permeating cryoprotectant such as sorbitol. The optimum concentration of sorbitol was found in a hypertonic solution of around 2 M. It was also very important to use S. pombe cells grown in minimal medium and S. cerevisiae cells grown in nutrient medium in the exponential growth phase. A slow freezing rate of 10 degrees C/min and a rapid thawing rate of 200 degrees C/min resulted in the highest transformation efficiency. We also found it necessary to wash the thawed cells with 1.0 M of non electrolyte sorbitol, since the intracellular electrolytes had leaked as a result of cryoinjury. The frozen competent cells stored at -80 degrees C could be used for more than 9 months without any loss of transformation efficiency. This cryopreservation method for electroporation is simple and useful for routine transformations of intact cells. Frozen competent cells offer the advantages of long-term storage with high efficiency and freedom from the preparation of fresh competent cells for each transformation. PMID- 10870101 TI - Polymorphism of Saccharomyces cerevisiae aquaporins. AB - Aquaporin water channels facilitate the transmembrane diffusion of water and higher organisms possess a large number of isoforms. The genome of the yeast Saccharomyces cerevisiae contains two highly similar aquaporin genes, AQY1 and AQY2. AQY1 has been shown to encode a functional water channel but only in certain laboratory strains. Here we show that the AQY2 gene is interrupted by an 11 bp deletion in 23 of the 27 laboratory strains tested, with the exception of strains from the sigma 1278b background, which also exhibit a functional Aqy1p. However, although the AQY2 gene from sigma 1278b is highly homologous to functional aquaporins, we did not observe Aqy2p-mediated water transport in Xenopus oocytes. A survey of 52 yeast strains revealed that all industrial and wild yeasts carry the allele encoding a functional Aqy1p, while none of these strains appear to have a functional Aqy2p. We conclude that natural and industrial conditions provide selective pressure to maintain AQY1 but apparently not AQY2. PMID- 10870102 TI - The 'SUN' family: yeast SUN4/SCW3 is involved in cell septation. AB - SUN4 is the fourth member of the SUN gene family from S. cerevisiae, whose products display high homology in their 258 amino acid C-terminal domain. SIM1, UTH1, NCA3 (the founding members) are involved in different cellular processes (DNA replication, ageing, mitochondrial biogenesis) and it is shown herein that SUN4 plays a role in the cell septation process. sun4 delta cells are larger than wild-type and begin a new cell cycle before they have separated from their mother cell. This phenotype is more pronounced in sun4Delta cells also deleted for UTH1. FACS analysis shows apparent polyploidy which disappears when the cell cycle is arrested by mating factor or nocodazole, indicating that cell septation is delayed without modification of the doubling time. Elutriated sun4 delta uth1 delta daughter cells are born larger, and therefore enter S phase sooner than their wild-type counterpart. S phase duration, as well as timing of Clb2 degradation, is normal, but cell septation is delayed. Sun4p/Scw3p was recently described as a cell wall protein (Cappellaro et al., 1998) and, consistent with this notion, electron micrographs of sun4 delta cells show defects in the final steps of cell wall septation. Our data suggest that Sun4p and Uth1p might contribute to the regulated process of cell wall morphogenesis and septation. PMID- 10870103 TI - Possible regulatory function of the Saccharomyces cerevisiae Ty1 retrotransposon core protein. AB - The yeast Ty1 retrotransposon encodes proteins and RNA that assemble into virus like particles (VLPs) as part of the life cycle of the retro-element. The Tya protein, which is equivalent to the retroviral Gag, is the major structural component of these particles. In this work, we demonstrate that Tya proteins fulfil other functions apart from their structural role. We show that Tya interacts in vitro with the Ty1 RNA domain required for RNA packaging, suggesting that this RNA-protein interaction may direct the packaging process. Furthermore, the overexpression of both Tya proteins, i.e. p1, the primary translation product, and p2, the mature form, increases endogenous Ty1 RNA levels in trans without increasing translation significantly. These observations suggest that Tya may exert a regulatory function during transposition. Interestingly, however, only p2, the mature form of Tya, trans-activates transposition of a marked genomic Ty element. This confirms that processing is required for transposition. PMID- 10870104 TI - A phosphatidylinositol 3-kinase of Candida albicans: molecular cloning and characterization. AB - A phosphatidylinositol (PI) 3-kinase gene (CaVPS34) of the human pathogenic yeast Candida albicans was cloned by a PCR-based homology approach. The open reading frame encodes a 1020 amino acid protein with a calculated molecular weight of 118 kDa and a relative isoelectric point of 6.9. It shares 47% sequence identity with Saccharomyces cerevisiae Vps34p. Southern pattern indicated that CaVPS34 is probably present as a single copy gene per haploid genome in C. albicans. We localized the CaVPS34 gene on chromosome 1. Under all conditions tested a major CaVPS34 transcript of approximately 3. 5 kb could be detected. CaVPS34 mRNA levels increased during exponential growth up to 12-fold followed by a decline upon entry into stationary phase. The size of a 6xHis tag-CaVps34p fusion protein purified from Escherichia coli is in agreement with the calculated molecular mass of CaVps34p. It exhibits in vitro PI 3-kinase activity and produces only phosphatidylinositol 3-phosphate. The CaVPS34 gene under the control of its own promoter were not able to complement the temperature-sensitive growth of S. cerevisiae vps34. However, overexpression of CaVPS34 was sufficient to rescue the temperature-sensitive vps34 phenotype, suggesting a functional conservation in C. albicans. PMID- 10870105 TI - Isolation and nucleotide sequence of a gene encoding tRNA nucleotidyltransferase from Kluyveromyces lactis. AB - A gene (KlCCA1) encoding ATP(CTP):tRNA specific tRNA nucleotidyltransferase (EC 2.7.7.25) was isolated from Kluyveromyces lactis by complementation of the Saccharomyces cerevisiae cca1-1 mutation. Sequencing of a 2665 bp EcoRI-SpeI restriction fragment revealed an open reading frame potentially encoding a protein of 489 amino acids with 57% sequence similarity to its S. cerevisiae homologue. Southern hybridization revealed a single copy of KlCCA1 in the K. lactis genome. KlCCA1 was able to complement both the mitochondrial and cytosolic defects in the cca1-1 mutant, suggesting that, as in S. cerevisiae, the K. lactis gene encodes a sorting isozyme that is targeted to mitochondria and the nucleus and/or cytosol. An altered KlCCA1 gene encoding a tRNA nucleotidyltransferase that lacked its first 35 amino acids was able to complement the nuclear/cytosolic but not the mitochondrial defect in the S. cerevisiae cca1-1 mutant, suggesting that the 35 amino-terminal amino acids are necessary for targeting to mitochondria but are not required for enzyme activity. Our results suggest that the mechanisms for production and distribution of mitochondrial and nuclear/cytosolic tRNA nucleotidyltransferase in K. lactis differ from those seen in S. cerevisiae. PMID- 10870106 TI - Cloning and sequence analysis of the Candida boidinii ADE2 gene. AB - Candida boidinii ADE2 gene (phosphoribosyl-5-aminoimidazole carboxylase; AIRC, EC 4. 1. 1. 21) has been cloned by homology to the Saccharomyces cerevisiae ADE2 gene. The cloned C. boidinii ADE2 gene complemented the ade2 mutation of S. cerevisiae. Sequence analysis showed a single open reading frame of 1719 nucleotides coding for a polypeptide of 573 residues. Comparison of the deduced amino acid sequence with those of AIRC enzymes from other yeasts showed marked homology among yeast AIRCs. PMID- 10870107 TI - Functional cloning of the adenylate cyclase gene of Candida albicans in Saccharomyces cerevisiae within a genomic fragment containing five other genes, including homologues of CHS6 and SAP185. AB - We have cloned a genomic fragment of Candida albicans by complementation of a Saccharomyces cerevisiae cyr1 mutant. This fragment contains the two-thirds C terminal part of the adenylate cyclase CaCYR1. The complete gene has been sequenced from PCR fragments amplified from genomic DNA, and contains an ORF of 1690 amino acids closely related to other fungal adenylate cyclases. Adjacent to the adenylate cyclase gene, we have sequenced six other putative genes. CaCHS6, CaYNL191 and CaYJL098 are named on the basis of their close similarity with S. cerevisiae genes. ORFs CaYJL097a and CaYJL097b represent two repeated homologues of the S. cerevisiae YJL097w, which probably arose from an ancient duplication. The last one is a hypothetical ORF, CaYKR049, which presents only a very weak similarity with YKR049. The S. cerevisiae homologues of three of these genes are co-localized on chromosome X but with a different order and orientation. PMID- 10870108 TI - Identifying tagged transposon insertion sites in yeast by direct genomic sequencing. AB - Tagged transposons are powerful tools for large-scale studies of gene expression, protein localization, and gene disruption in Saccharomyces cerevisiae. The current techniques used to identify transposon insertion sites in the yeast genome require a DNA amplification step that can be time-consuming and problematic. We show that the DNA amplification step can be bypassed. Insertion sites can be identified rapidly and reliably by direct genomic sequencing using a transposon-specific primer, BigDye-labelled terminators, and an automated sequencer. Direct genomic sequencing can also save time on the genetic analysis phase of transposon-based projects. PMID- 10870109 TI - Current awareness on yeast. PMID- 10870110 TI - A global strategy for prevention and detection of blood doping with erythropoietin and related drugs. PMID- 10870111 TI - A novel method utilising markers of altered erythropoiesis for the detection of recombinant human erythropoietin abuse in athletes. AB - BACKGROUND AND OBJECTIVE: The use of recombinant human erythropoietin (r-HuEPO) to enhance athletic performance is prohibited. Existing tests cannot readily differentiate between exogenous and endogenous EPO. Therefore the aim of our study was to investigate possible indirect detection of r-HuEPO use via blood markers of altered erythropoiesis. DESIGN AND METHODS: Twenty-seven recreational athletes were assigned to three groups prior to a 25 day drug administration phase, with the following protocols: EPO+IM group (n = 10), 50 Ukg(-1) r-HuEPO at a frequency of 3wk(-1), 100 mg intramuscular (IM) iron 1wk(-1) and a sham iron tablet daily; EPO+OR group (n = 8), 50 U.kg(-1) r-HuEPO 3wk(-1), sham iron injection 1wk(-1) and 105 mg of oral elemental iron daily; placebo group (n = 9), sham r-HuEPO injections 3wk(-1), sham iron injections 1wk(-1) and sham iron tablets daily. Each group was monitored during and for 4 weeks after drug administration. RESULTS: Models incorporating combinations of the variables reticulocyte hematocrit (RetHct), serum EPO, soluble transferrin receptor, hematocrit (Hct) and % macrocytes were analyzed by logistic regression. One model (ON-model) repeatedly identified 94-100% of r-HuEPO group members during the final 2 wk of the r-HuEPO administration phase. One false positive was recorded from a possible 189. Another model (OFF-model) incorporating RetHct, EPO and Hct was applied during the wash-out phase and, during the period of 12-21 days after the last r-HuEPO injection, it repeatedly identified 67-72% of recent users with no false positives. INTERPRETATION AND CONCLUSIONS: Multiple indirect hematologic and biochemical markers used simultaneously are potentially effective for identifying current or recent users of r-HuEPO. PMID- 10870112 TI - Glutathione S-transferase enzyme expression in hematopoietic cell lines implies a differential protective role for T1 and A1 isoenzymes in erythroid and for M1 in lymphoid lineages. AB - BACKGROUND AND OBJECTIVES: Glutathione S-transferases (GSTs) are phase II metabolizing enzymes which catalyze the conjugation of glutathione (GSH) to electrophilic substrates and possess selenium-independent glutathione peroxidase activity. The GST enzyme family includes the cytosolic isoforms GST-alpha, mu (GSTM), pi (GSTP), theta (GSTT) and sigma (GSTS). GSTT1, P1 and M1 are polymorphic and altered polymorphic frequency of genes encoding these proteins has been suggested as a potential risk factor for the development of hematopoietic malignancies. Overexpression of GSTs has also been implicated in chemotherapeutic drug resistance. This study was undertaken to elucidate the potential functional relevance of these genetic polymorphisms in hematopoiesis. DESIGN AND METHODS: GST genotype of 14 hematopoietic cell lines was determined by polymerase- chain-reaction (PCR). Gene expression of GSTs in a cell line was detected by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) on TaqMan 7700 and by semi-quantitative RT-PCR. Cytosolic GST protein expression was detected by Western blot. GST conjugation activity was assayed using 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. RESULTS: GSTP1 expression was higher than other GSTs in 13/14 cell lines and paralleled CDNB conjugation activity. GSTP1 and GSTM1 predominated in lymphoid lines whilst T1 expression was relatively greatest in erythroid lines but was absent in 7/12 non null lines. GSTT2 was expressed in only 3/4 lines. The 3 cell lines which expressed GSTA1 were all erythroid. INTERPRETATION AND CONCLUSIONS: Glutathione S transerases showed differential lineage expression in hematopoietic cell lines. This implies a greater cytoprotective role for GSTT1 and GSTA1 in erythroid cells and GSTM1 in lymphoid cells. We postulate that inherited gene deletion of GSTT1 and M1 may produce increased genotoxic susceptibility for erythroid and lymphoid cell respectively, following exposure to xenobiotics that are substrates for these enzymes. PMID- 10870113 TI - Age-related expression of the cellular prion protein in human peripheral blood leukocytes. AB - BACKGROUND AND OBJECTIVE: Creutzfeldt-Jakob disease typically affects older patients, yet victims of new-variant Creutzfeldt-Jakob disease (nvCJD) are unusually young. Because the cellular prion protein PrP(C) is required for disease development, we investigated age-dependent variability in cell surface PrP(C) expression on various subclasses of human peripheral blood leukocytes (PBL) as a possible susceptibility factor. DESIGN AND METHODS: Three age groups of healthy individuals (mean ages: 6, 33 and 68) were studied by two-color FACS analysis of PBL with fluorescent monoclonal antibodies to PrP(C) and to the lineage markers CD3, CD19, CD4, and CD8. For each subclass marker, surface PrP(C) levels were expressed as mean fluorescence intensity ratios (MFIR) by dividing the geometric mean of the fluorescence of each test antibody by the geometric mean of its isotype-matched control antibody. PrP(C) expression levels in each age and lineage group were compared using appropriate non-parametric tests. RESULTS: We found significant age-related differences in PrP(C) expression on lymphocytes (p=0. 0004). The elderly expressed significantly higher levels than children (p=0.0006) and adults (p=0.0009). PrP(C) expression was also significantly higher in CD3(+) compared to CD19(+ )(p=0.0004) and in CD8(+) compared to CD4(+ )lymphocytes (p=0.0044). INTERPRETATION AND CONCLUSIONS: If PrP(C) expression on PBL were a significant susceptibility factor for nvCJD, young persons would display higher levels. Instead, the elderly expressed the highest amounts of PrP(C) on PBL. This argues against the hypothesis that variability in cell surface expression of PrP(C) in PBL contributes to the exquisite susceptibility of the young to nvCJD. PMID- 10870114 TI - The interaction of gemcitabine and cytarabine on murine leukemias L1210 or P388 and on human normal and leukemic cell growth in vitro. AB - BACKGROUND AND OBJECTIVE: Gemcitabine (dFdC) is a new nucleoside antimetabolite of deoxycytidine that resembles cytarabine (Ara-C) in both its structure and metabolism. Little is known about dFdC efficacy in hematologic malignancies, either as a single drug or in combination with other drugs. In this study we have tried to determine whether the cytotoxic effect of Ara-C can be increased by using it in combined therapy with dFdC. DESIGN AND METHODS: In the in vivo part of our study, mice bearing L1210 or P388 leukemia were treated with dFdC and Ara C. The drugs were administered alone and in combination according to the following schedules: Ara-C and dFdC at the same time, dFdC before Ara-C, and Ara C before dFdC. The efficacy of the therapy against leukemia (defined as the increase in lifespan, ILS) was assessed as the percentage of the median survival time (MST) of the treated group (T) in relationship to that of the control group (C): ILS=[(MST(C)/MST(T)) -1]x100. In the in vitro part of our study, normal granulocyte-macrophage colony-forming unit (CFU-GM) cells as well as CFU-GM cells obtained from patients with chronic myeloid leukemia (CML) were incubated either with dFdC or Ara-C alone or with adequate concentrations of a combination of these drugs. RESULTS: The in vivo experiment revealed that in both leukemias tested, combined therapy with dFdC given before Ara-C and dFdC given at the same time with Ara-C were more effective than monotherapy with either dFdC or Ara-C. The other treatment schedule (Ara-C before dFdC) did not significantly prolong the survival time of the treated mice bearing L1210 or P388 leukemia as compared with the treatment with dFdC alone. The in vitro experiments showed that dFdC used together with Ara-C acted additively on normal as well as CML CFU-GM cells. Furthermore, the drugs used jointly inhibited the growth of colonies formed by CML CFU-GM cells to a significantly higher degree than normal CFU-GM and the differences were statistically significant in the case of the combination of highest concentrations. INTERPRETATION AND CONCLUSIONS: Gemcitabine increased the activity of Ara-C. As these agents incorporate into DNA blocking chain elongation, and moreover, dFdC influences the cytotoxicity of Ara-C, our results could be explained by the drugs acting at these levels. dFdC used jointly with Ara-C may have an important clinical implication in the treatment of CML and other hematologic malignancies in future. PMID- 10870115 TI - Danazol treatment of idiopathic myelofibrosis with severe anemia. AB - BACKGROUND AND OBJECTIVE: Severe anemia is an important problem in patients with idiopathic myelofibrosis (IM). When other therapeutic measures are unsuccessful or not applicable, 40-50% favorable responses are obtained with androgen therapy. Oxymetholone is the drug usually employed, but good results have also been reported with danazol, although the experience is limited to a few patients. The aim of the present study was to evaluate the effect of danazol on the anemia of IM. DESIGN AND METHODS: Seven out of 22 consecutive IM patients were eligible for danazol treatment because of severe anemia not treatable with (four cases) or refractory to (three cases) other therapies. Danazol (600-800 mg/day) was given orally for six months and thereafter progressively tapered to the minimum effective dose in responding patients or discontinued in non-responders. Complete response was considered cessation of transfusion requirements with normalization of hemoglobin (Hb) values; partial response was defined as a > 30% reduction in transfusional needs or an increase > 10 g/L in the Hb. The effect on platelet counts was also analyzed. RESULTS: One patient splenectomized three years earlier achieved a complete response and three a partial response, giving an overall response rate of 57 %. A significant increase in platelet counts was also observed in three responders. The responses were first seen between three and six months after the start of treatment, which was usually well tolerated. INTERPRETATION AND CONCLUSIONS: Danazol, given at an appropriate dosage for a sufficient time, is an effective treatment for a substantial proportion of IM patients with severe anemia without marked splenomegaly or who have been previously splenectomized. PMID- 10870116 TI - Interleukin-1 beta, interleukin-1 receptor antagonist and interleukin-6 plasma levels and cytokine gene polymorphisms in chronic lymphocytic leukemia: correlation with prognostic parameters. AB - BACKGROUND AND OBJECTIVE: The growth of B-cell chronic lymphocytic leukemia (B CLL) cells has been shown to be dependent on exogenous growth factors in vitro. We wanted to evaluate the clinical relevance of interleukin (IL)-6, IL-1 beta and interleukin-1 receptor antagonist (IL-1Ra) in B-CLL. As the plasma levels of IL 6, IL-1 beta and IL-1Ra have been suggested to be partly dependent on gene polymorphism, the previously described polymorphisms of the IL-1 complex genes and the IL-6 gene were also studied. DESIGN AND METHODS: The plasma levels of these cytokines were measured in a cohort of 36 patients with B-CLL and in 400 healthy subjects. The previously described polymorphisms of the IL-1 complex genes and the IL-6 gene were studied using PCR and RFLP. These data was correlated with other parameters associated with severity and prognosis of B-CLL and a number of clinical and laboratory findings. RESULTS: The plasma concentrations of IL-1 beta and IL-1Ra were lower in B-CLL patients than in normal controls (p < 0.001). The IL-1 beta plasma levels were dependent on the cell immunophenotype score and state of progression of the disease. Moreover, plasma concentrations of IL-6 were elevated in B-CLL patients compared with healthy subjects (p < 0.005) and correlated with disease stage, hemoglobin levels, anemia and erythrocyte sedimentation rate in the patients. The allele frequencies of the analyzed genes were similar in patients and controls. INTERPRETATION AND CONCLUSIONS: Our data demonstrate that in B-CLL, plasma levels of IL-1 beta, IL-1Ra and IL-6 differ from normal, and mechanisms other than allelic imbalance of their genes account for the distinct cytokine profiles observed in this disease. PMID- 10870117 TI - Application of cross-species color banding (RxFISH) in the study of T prolymphocytic leukemia. AB - BACKGROUND AND OBJECTIVE: Cross-species color banding (RxFISH) is a new FISH technology based on the use of differentially labeled gibbon chromosome probes to obtain a specific color banding pattern for each human chromosome. The aim of the study was to test the RxFISH technique for better characterization of complex karyotypes in patients with T-prolymphocytic leukemia (T-PLL). DESIGN AND METHODS: The study evaluated the cross-species color banding technique in four patients affected with T-PLL previously studied by conventional cytogenetics. RESULTS: All patients showed an abnormal karyotype and three of them had a complex karyotype. The involvement of 14q11 in all four cases, the gain of 8q in three cases and a loss of chromosome 10, 15 and 17 and a gain of chromosome 21 in two cases were noted. The RxFISH technique identified from 2 to 7 not previously recognized aberrations per case and confirmed the inv(14)(q11q32). INTERPRETATION AND CONCLUSIONS: To our knowledge, this is the first application of RxFISH to characterize chromosomal rearrangements in T-cell neoplasms. RxFISH gave rapid and easy identification of chromosome rearrangements that were difficult to recognize by conventional cytogenetics. Using this new technology we identified 15 rearrangements not detected by conventional cytogenetics. PMID- 10870118 TI - Persistent tumor 18F-FDG uptake after a few cycles of polychemotherapy is predictive of treatment failure in non-Hodgkin's lymphoma. AB - BACKGROUND AND OBJECTIVE: Early recognition of the ineffectiveness of chemotherapy could result in lower cumulative drug toxicity and tumor burden at the start of salvage therapy, which might improve clinical outcome. Therefore, we studied the value of (18)F-FDG PET for early evaluation of response in patients with non-Hodgkin's lymphoma (NHL). DESIGN AND METHODS: We studied 28 patients by (18)F-FDG PET after a median of 3 cycles of polychemotherapy. The presence or absence of abnormal (18)F-FDG uptake was correlated to clinical outcome (median follow-up: 17.5 months, range 4-47 months). RESULTS: Five of 28 patients still had increased (18)F-FDG uptake in one or more sites previously shown to be involved by lymphoma at baseline evaluation. Only one of these five patients entered complete remission (CR), whereas among the 23 patients with negative (18)F-FDG PET studies, two died of toxicity during chemotherapy and all the others entered clinical CR (p<0.00001). All five patients with and 7/21 patients without residual abnormal (18)F-FDG uptake relapsed or reprogressed (positive predictive value for relapse: 100%, negative predictive value: 67%). By Kaplan Meier analysis, progression-free survival (PFS) at 1 and 2 years was respectively 20+/-18% and 0% for (18)F-FDG PET positive patients and 81+/-9% and 62+/-12% for (18)F-FDG PET negative patients (p=0.0001). Overall survival (OS) at 1 and 2 years was respectively 20+/-18% and 0% for (18)F-FDG PET positive and 87+/-7% and 68+/-11% for (18)F-FDG PET negative patients (p<0.0001). INTERPRETATION AND CONCLUSIONS: Persistent tumoral (18)F-FDG uptake after a few cycles of polychemotherapy is predictive of CR, PFS and OS in NHL. Further studies are warranted to determine whether (18)F-FDG PET has a predictive value independent from conventional prognostic factors. However, the sensitivity of qualitative (18)F-FDG PET imaging in identifying patients with a poor outcome was insufficient. Earlier evaluation after only one cycle of chemotherapy and quantitative analysis might increase the sensitivity of 18F-FDG PET is predicting treatment failure. PMID- 10870119 TI - Platelet ultrastructural morphometry for diagnosis of partial delta-storage pool disease in patients with mild platelet dysfunction and/or thrombocytopenia of unknown origin. A study of 24 cases. AB - BACKGROUND AND OBJECTIVE: Exact diagnosis is sometimes difficult in patients presenting with a slight bleeding diathesis, prolonged bleeding times, non specific aggregometric abnormalities, and/or mild thrombocytopenia. The objective of this study was to evaluate the use of platelet ultrastructural morphometry in detecting a partial d-storage pool disease in such patients. DESIGN AND METHODS: Platelets from 52 patients and 15 controls were fixed immediately in glutaraldehyde in White's saline without anticoagulant and processed for transmission electron microscopy. Using computer-assisted morphometry, the size and shape of the platelets were measured, as were the size and number per platelet of the dense- and a-granules. Ultrastructural morphology of the above and other intraplatelet structures was observed. RESULTS: Twenty-four cases were diagnosed as having a partial d-storage pool disease. Mean platelet area (2.28 microm(2)) and maximum diameter (2.58 microm) were significantly greater in patients than in control subjects (1.64 microm(2) and 2. 25 microm, respectively) but discoid shape was preserved. Mean dense-granule number was decreased, both per platelet and per microm(2) of platelet area (patients 0.22 and 0.09; controls 0.42 and 0.24). Seven patients also had a marked decrease in a-granules, resulting in a significantly lower mean number of granules per microm(2 )(patients 2.43; controls 3.15). Additionally, the patients' platelets had significant increases in both lipid droplets and surface-connected canalicular system. INTERPRETATION AND CONCLUSIONS: A partial dense-granule deficiency, sometimes associated with partial a-granule deficiency, should be borne in mind faced with patients who have a slight bleeding diathesis, non-specific platelet dysfunction tests and/or mild thrombocytopenia of unknown origin. Platelet ultrastructural morphometry is useful in diagnosing this condition. PMID- 10870121 TI - Prevalence and clinical significance of antiprothrombin antibodies in patients with systemic lupus erythematosus or with primary antiphospholipid syndrome. AB - BACKGROUND AND OBJECTIVE: Antibodies to prothrombin (aII) have been identified in patients with antiphospholipid antibodies, but their clinical significance is not well known. The aim of our study was to investigate their prevalence and association with clinical manifestations of the antiphospholipid syndrome (APS) in patients with primary APS or with systemic lupus erythematosus (SLE). DESIGN AND METHODS: A series of 177 patients with autoimmune diseases was studied: 70 with primary APS and 107 with systemic lupus erythematosus. A control group of 87 healthy volunteers were included in the study. aII were investigated in sera by an ELISA, using human prothrombin as antigen fixed in irradiated polystyrene plates. RESULTS: aII prevalence in patients with autoimmune disease was 47% (57% and 40% in patients with primary APS or with SLE, respectively) significantly higher than in controls (5%) (p<0.0001). In the whole series, thrombotic events were more prevalent in patients with aII (45% vs 28%; p=0.02). Moreover, aII was found to be an independent risk factor for arterial thrombosis (OR=2.4; p=0. 04). Similarly, in patients with SLE, aII were associated with both arterial and venous thrombosis (35% vs 14%; p=0.01), although only IgG-aII (OR=3.7; p=0.01) had an independent value as risk factor for thrombosis. However, a relationship between aII and thrombosis was not found in primary APS. aII were associated with thrombocytopenia only in patients with primary APS (OR=6.7; p=0.007). INTERPRETATION AND CONCLUSIONS: aII seem to be a serological marker of thrombosis in autoimmune diseases, mainly in SLE patients and/or in the arterial territory. PMID- 10870122 TI - Hematopoietic stem cell transplantation in childhood: report from the bone marrow transplantation group of the Associazione Italiana Ematologia Oncologia Pediatrica (AIEOP). AB - BACKGROUND AND OBJECTIVE: Transplantation of hematopoietic stem cells from different sources is being increasingly used to treat a variety of diseases in children. Transplant procedures and indications have changed considerably during recent years. Monitoring of information about these changes is useful for interpretation of nationwide collected data. DESIGN AND METHODS: Since 1985, Centers belonging to the AIEOP (Associazione Italiana Ematologia Oncologia Pediatrica), performing hematopoietic stem cell transplants (HSCT) in children, and members of the AIEOP-Bone Marrow Transplant (BMT) Group annually report data on their transplant activity to the AIEOP-BMT Registry employing specially prepared patient-oriented forms. RESULTS: From January 1985 to December 1998, a total of 2,474 bone marrow (BM), peripheral blood (PB) or umbilical cord blood (CB) transplants were reported: 1,296 (52%) were allogeneic (Allo) and 1,178 (48%) autologous (Auto) transplants. These transplants were performed in 19 Italian Centers on 2,249 patients aged less than 17 years. Among Allo transplants, 1,198 (92%) were performed using BM progenitor cells, whereas 49 (4%) CB, 42 (3%) were PB, 4 BM plus PB, and 3 BM plus CB allografts; they were performed using HLA-identical sibling donors in 867 cases (67%) and alternative donors (i.e. partially-matched relatives or unrelated donors) in the remaining 429 (33%) cases. Allogeneic transplants were performed on 786 (67%) patients with malignancy and on 395 (33%) patients with non-malignant disorders. In the last 6 years, the number of Allo-transplants per year exceeded that of Auto-transplants. Of the Auto-transplants, 775 (66%) were performed using BM, and 403 (34%) using PB alone or combined with BM hematopoietic stem cells. Indications for Auto-BMT were myelo-lymphoproliferative disorders in 524 (49%) cases, solid tumor in 533 (50%) cases and non-malignant disease in 11 (1%) cases. In the last 5 years, the use of PB for autografts has increased from 7% to 70%. INTERPRETATION AND CONCLUSIONS: These data reflect the development and present status of HSCT in Italy and provide a basis for patient counseling and health care planning. PMID- 10870123 TI - Collection of Ph-negative progenitor cells from interferon responsive patients with chronic myeloid leukemia: effect of granulocyte-colony-stimulating factor mobilization. AB - BACKGROUND AND OBJECTIVE: The observation that patients with chronic myeloid leukemia (CML) may relapse following stem cell transplantation because of Philadelphia positive cells contaminating the graft have led to a variety of strategies to reduce this contamination. This study investigate the feasibility of collective, Ph-re cells from patients with CML in chronic phase. DESIGN AND METHODS: A total of 18 patients with chronic myeloid leukemia in chronic phase who had responded to varying degrees to treatment with interferon-a (IFN) were subjected to mobilization with granulocyte colony-stimulating factors and peripheral blood progenitor cell collection. Nine patients were in complete cytogenetic remission (CCR) and nine were partial responders. IFN was stopped 2 to 4 weeks before the procedure. G-CSF was given by subcutaneous injection once daily at a dose of 10 microg/kg. RESULTS: Five patients underwent one collection procedure only, 10 underwent two procedures and 3 patients had three collections. The median number of nucleated cells (NC) per patient collected was 10.2 x 10(8)/kg (4.4-19.7) and the median number of CD34(+) cells was 2.5 x 10(6)/kg (0.4-9.4). Analyzable cytogenetic data were available for 26/34 (76%) leukapheresis procedures. The median percentage of Ph- negative metaphases for patients in CCR was 100% (73-100). Patients not in CCR had a higher level of Ph positive cells in their collections (median 23%, range 0-79%, p=0.01). Of the nine patients in CCR, 8 had at least one apheresis from which progenitor cells were 100% Ph-negative; conversely, patients not in CCR had detectable Ph-positive cells in every collection. Four patients have undergone autologous stem cell transplantation. INTERPRETATION AND CONCLUSIONS: It was possible to collect sufficient Ph negative progenitor cells from patients in CCR but collections from other patients contained significant numbers of Ph-positive cells. PMID- 10870124 TI - Clinical value of quantitative long-term assessment of bcr-abl chimeric transcript in chronic myelogenous leukemia patients after allogeneic bone marrow transplantation. AB - BACKGROUND AND OBJECTIVE: For purposes of therapeutic decision making, we used quantitative polymerase chain reaction (PCR) for molecular follow-up of 55 patients with chronic myeloid leukemia (CML) in complete remission (CR) after allogeneic bone marrow transplantation (BMT) from HLA compatible donors. DESIGN AND METHODS: A total of 402 bone marrow samples from 40 patients transplanted in chronic phase (group 1) and 15 in accelerated/blastic phase (group 2) were analyzed by qualitative and quantitative PCR. RESULTS: Regarding clinical outcome, 34/40 (85%) group 1 vs. 8/15 (54%) group 2 patients are alive. Only 1/40 (2.5%) group 1 patient relapsed, as against 6/15 (40%) in group 2 (p = 0. 0002). At qualitative PCR, 8/40 (19%) group 1 vs. 9/15 (60%) group 2 patients were positive, with a significantly greater total number of positive samples in group 2 (33/129, 27% vs. 16/273, 5%; p<0.001). The probability of qualitative PCR positivity >1 year after BMT was significantly lower in group 1 patients (4/40 pts, 10% vs. 9/15 pts, 60%; p = 0.01). At quantitative PCR, 4/8 (50%) group 1 patients were positive only once (< 400 transcripts/microg RNA). In group 2, 9/15 (60%) patients had 3 or more positive samples (always with >4,000 copies/mg RNA); therapeutic interventions (cyclosporin A discontinuation, temporary a-interferon or donor lymphocyte infusion) restored molecular remission in 4/9 (44%) cases. INTERPRETATION AND CONCLUSIONS: This study indicates that quantitative PCR could provide practical indications capable of directing therapeutic interventions for transplanted CML patients, especially those transplanted in accelerated/blastic phase, for whom intensive monitoring is required. PMID- 10870125 TI - Tacrolimus (FK 506) induced thrombotic thrombocytopenic purpura after ABO mismatched second liver transplantation: salvage with plasmapheresis and prostacyclin. AB - We report the course of thrombotic thrombocytopenic purpura (TTP) in a patient receiving tacrolimus (FK506) immunosuppression for an ABO mismatched second liver graft. A Chinese woman with fulminant hepatitis-B reactivation failed a living related orthotopic liver transplantation (OLT) due to portal vein thrombosis. An ABO mismatched cadaveric OLT (group A to O) was performed, with peri-operative plasmapheresis to reduce anti-A hemagglutinin titers. On day 30, she developed fever, hemolysis, thrombocytopenia and neurologic dulling. Prominent microangiopathic features in peripheral blood film, and characteristic brain lesions on magnetic resonance imaging confirmed TTP. She responded initially to intensive plasmapheresis with cryosupernatant replacement, and withdrawal of FK506. An attempted reintroduction of FK506 for threatened rejection led to TTP exacerbation. This was controlled with prolonged plasmapheresis and a ten-day infusion of prostacyclin. Immunosuppression was changed to mycophenolate mofetil. By day 53, the peripheral film and lactate dehydrogenase level had returned to baseline and plasmapheresis was stopped. PMID- 10870126 TI - Mitosis in peripheral blood in prematurity. PMID- 10870127 TI - Investigation of minimal residual disease in adult Ph1 positive acute lymphoblastic leukemia by a combination of cell sorting and fluorescence in situ hybridization: a preliminary study on 6 cases. PMID- 10870129 TI - Primary non-Hodgkin's lymphoma of the vagina. PMID- 10870128 TI - Transformation of severe aplastic anemia into myelodysplastic syndrome with monosomy 7: monoclonal origin detected by HUMARA gene analysis during the aplastic anemia phase. PMID- 10870130 TI - Homozygous Constant Spring: the first case described in the west. PMID- 10870131 TI - Liver nodular regenerative hyperplasia after bone marrow transplant. PMID- 10870132 TI - Incidence of Factor V Leiden and prothrombin G20210A in patients submitted to stem cell transplantation. PMID- 10870133 TI - Acute rhabdomyolysis after high dose chemotherapy and circulating progenitor cell autografting for breast cancer. PMID- 10870134 TI - Vulnerability of the sympathetic trunk during the anterior approach to the lower cervical spine. AB - STUDY DESIGN: Anatomic dissection and measurements of the cervical sympathetic trunk relative to the medial border of the longus colli muscle and lateral angulation of the sympathetic trunk relative to the midline on both sides were performed. OBJECTIVE: To determine the course and location of the sympathetic trunk quantitatively and relate this to the vulnerability of the sympathetic trunk during the anterior approach to the lower cervical spine. SUMMARY OF BACKGROUND DATA: The sympathetic trunk is sometimes damaged during the anterior approach to lower cervical spine, resulting in Horner's syndrome with its associated ptosis, meiosis, and anhydrosis. No quantitative regional anatomy describing the course and location of the sympathetic trunk and its relation to the longus colli muscle is available in the literature. METHODS: In this study, 28 adult cadavers were used for dissection and measurements of the sympathetic trunk. The distance between the sympathetic trunk and the medial borders of the longus colli muscle at C6 and the angle of the sympathetic trunk with respect to the midline were determined bilaterally. The distance between the medial borders of the longus colli muscle from C3 to C6 and the angle between the medial borders of the longus colli muscle also were measured. RESULTS: The sympathetic trunk runs in a superior and lateral direction, with an average angle of 10.4 +/- 3.8 degrees relative to the midline. The average distance between the sympathetic trunk and the medial border of the longus colli muscle is 10.6 +/- 2.6 mm. The average diameter of the sympathetic trunk at C6 is 2.7 +/- 0.6 mm. The length and width of the middle cervical ganglion were 9.7 +/- 2.1 mm and 5.2 +/- 1.3 mm, respectively. The distance between the medial borders of the longus colli muscle was 7.9 +/- 2.2 mm at C3, 10.1 +/- 3.1 mm at C4, 12.3 +/- 3.1 mm at C5, and 13.8 +/- 2.2 mm at C6, and the angle between the medial borders of the longus colli muscle was 12.5 +/- 4. 7 degrees. CONCLUSIONS: The sympathetic trunk may be more vulnerable to damage during anterior lower cervical spine procedures because it is situated closer to the medial border of the the longus colli muscle at C6 than at C3. The longus colli muscles diverge laterally, whereas the sympathetic trunks converge medially at C6. As the transverse foramen or uncovertebral joint is exposed with dissection or transverse severance of the longus colli muscle at the lower cervical levels, the sympathetic trunk should be identified and protected. PMID- 10870135 TI - Biomechanical rationale for the pathology of rheumatoid arthritis in the craniovertebral junction. AB - STUDY DESIGN: A finite-element model of the craniovertebral junction was developed and used to determine whether a biomechanical mechanism, in addition to inflammatory synovitis, is involved in the progression of rheumatoid arthritis in this region of the spine. OBJECTIVES: To determine specific structure involvement during the progression of rheumatoid arthritis and to evaluate these structures in terms of their effect on clinically observed erosive changes associated with the disease by assessing changes in loading patterns and degree of anterior atlantoaxial subluxation. SUMMARY OF BACKGROUND DATA: Rheumatoid arthritis involvement of the occipito-atlantoaxial (C0-C1-C2) complex is commonly seen. However, the biomechanical contribution to the development and progression of the disease is neither well understood nor quantified. Although previous cadaver studies have elucidated information on kinematic motion and fusion techniques, the modeling of progressive disease states is not easily accomplished using these methods. The finite-element method is well suited for studying progressive disease states caused by the gradual changes in material properties that can be modeled. METHODS: A ligamentous, nonlinear, sliding-contact, three-dimensional finite-element model of the C0-C1-C2 complex was generated from 0.5 mm thick serial computed tomography scans. Validation of the model was accomplished by comparing baseline kinematic predictions with experimental data. Transverse, alar, and capsular ligament stiffness were reduced sequentially by 50%, 75%, and 100% (removal) of their intact values. All models were subjected to flexion moments replicating the clinical diagnosis of rheumatoid arthritis using full flexion lateral plane radiographs. Stress profiles at the transverse ligament odontoid process junction were monitored. Changes in loading profiles through the C0-C1 and C1-C2 lateral articulations and their associated capsular ligaments were calculated. Anterior and posterior atlantodental interval values were calculated to correlate ligamentous destruction with advancement of atlantoaxial subluxation. RESULTS: Model predictions (at 0.3 Nm) fell within one standard deviation of experimental means, and range of motion data agreed with published in vitro and in vivo values. The model predicted that stresses at the posterior base of the odontoid process were greatly reduced with transverse ligament compromise beyond 75%. Decreases through the lateral C0-C1 and C1-C2 articulations were compensated by their capsular ligaments. Anterior and posterior atlantodental interval values indicate that the transverse ligament stiffness decreases beyond 75% had the greatest effect on atlantoaxial subluxation during the early stages of the disease (no alar and capsular ligament damage). Subsequent involvement of the alar and capsular ligaments produced advanced atlantoaxial subluxation, for which surgical intervention may be warranted. CONCLUSIONS: To the best of the authors' knowledge, this is the first report of a validated, three-dimensional model of the C0-C1-C2 complex with application to rheumatoid arthritis. The data indicate that there may be a mechanical component (in addition to enzymatic degradation) associated with the osseous resorption observed during rheumatoid arthritis. Specifically, erosion of the odontoid base may involve Wolff's law of unloading considerations. Changes through the lateral aspects of the atlas suggest that this same mechanism may be partially responsible for the erosive changes seen during progressive rheumatoid arthritis. Anterior and posterior atlantodental interval values indicate that complete destruction of the transverse ligament coupled with alar and/or capsular ligament compromise is requisite if advanced levels of atlantoaxial subluxation are present. PMID- 10870136 TI - The biomechanical effect of postoperative hypolordosis in instrumented lumbar fusion on instrumented and adjacent spinal segments. AB - STUDY DESIGN: Change in lumbar lordosis was measured in patients that had undergone posterolateral lumbar fusions using transpedicular instrumentation. The biomechanical effects of postoperative lumbar malalignment were measured in cadaveric specimens. OBJECTIVES: To determine the extent of postoperative lumbar sagittal malalignment caused by an intraoperative kneeling position with 90 degrees of hip and knee flexion, and to assess its effect on the mechanical loading of the instrumented and adjacent segments. SUMMARY OF BACKGROUND DATA: The importance of maintaining the baseline lumbar lordosis after surgery has been stressed in the literature. However, there are few objective data to evaluate whether postoperative hypolordosis in the instrumented segments can increase the likelihood of junctional breakdown. METHODS: Segmental lordosis was measured on preoperative standing, intraoperative prone, and postoperative standing radiographs. In human cadaveric spines, a lordosis loss of up to 8 degrees was created across L4-S1 using calibrated transpedicular devices. Specimens were tested in extension and under axial loading in the upright posture. RESULTS: In patients who underwent L4-S1 fusions, the lordosis within the fusion decreased by 10 degrees intraoperatively and after surgery. Postoperative lordosis in the proximal (L2-L3 and L3-L4) segments increased by 2 degrees each, as compared with the preoperative measures. Hypolordosis in the instrumented segments increased the load across the posterior transpedicular devices, the posterior shear force, and the lamina strain at the adjacent level. CONCLUSIONS: Hypolordosis in the instrumented segments caused increased loading of the posterior column of the adjacent segments. These biomechanical effects may explain the degenerative changes at the junctional level that have been observed as long-term consequences of lumbar fusion. PMID- 10870137 TI - Mechanical initiation of intervertebral disc degeneration. AB - STUDY DESIGN: Mechanical testing of cadaveric lumbar motion segments. OBJECTIVES: To test the hypothesis that minor damage to a vertebral body can lead to progressive disruption of the adjacent intervertebral disc. SUMMARY OF BACKGROUND DATA: Disc degeneration involves gross structural disruption as well as cell mediated changes in matrix composition, but there is little evidence concerning which comes first. Comparatively minor damage to a vertebral body is known to decompress the adjacent discs, and this may adversely affect both structure and cell function in the disc. METHODS: In this study, 38 cadaveric lumbar motion segments (mean age, 51 years) were subjected to complex mechanical loading to simulate typical activities in vivo while the distribution of compressive stress in the disc matrix was measured using a pressure transducer mounted in a needle 1.3 mm in diameter. "Stress profiles" were repeated after a controlled compressive overload injury had reduced motion segment height by approximately 1%. Moderate repetitive loading, appropriate for the simulation of light manual labor, then was applied to the damaged specimens for approximately 4 hours, and stress profilometry was repeated a third time. Discs then were sectioned and photographed. RESULTS: Endplate damage reduced pressure in the adjacent nucleus pulposus by 25% +/- 27% and generated peaks of compressive stress in the anulus, usually posteriorly to the nucleus. Discs 50 to 70 years of age were affected the most. Repetitive loading further decompressed the nucleus and intensified stress concentrations in the anulus, especially in simulated lordotic postures. Sagittal plane sections of 15 of the discs showed an inwardly collapsing anulus in 9 discs, extreme outward bulging of the anulus in 11 discs, and complete radial fissures in 2 discs, 1 of which allowed posterior migration of nucleus pulposus. Comparisons with the results from tissue culture experiments indicated that the observed changes in matrix compressive stress would inhibit disc cell metabolism throughout the disc, and could lead to progressive deterioration of the matrix. CONCLUSIONS: Minor damage to a vertebral body endplate leads to progressive structural changes in the adjacent intervertebral discs. PMID- 10870138 TI - The effects of pedicle screw adjustments on neural spaces in burst fracture surgery. AB - STUDY DESIGN: An in vitro biomechanical study of experimental burst fractures and a pedicle screw device. OBJECTIVES: To investigate the effects that adjustments of a pedicle screw device have on the neural spaces of the burst fracture. SUMMARY OF BACKGROUND DATA: Decompression of the neural spaces is important for the recovery of neural function after a burst fracture. No studies, experimental or clinical, are available that have attempted to relate the pedicle screw device adjustments to the decompression of the neural spaces. METHODS: Burst fractures were produced at L1 vertebra in nine human lumbar spine specimens. Pedicle screw devices were attached to T12 and L2. Eight device adjustments, consisting of pure translations (distraction or compression), pure extension, and combinations of translation and extension were applied. The dimensional changes in the canal and the superior and inferior intervertebral foramens were measured. Functions of restoration and improvement were determined for the adjustments to evaluate the effects of each adjustment and to determine the optimal adjustment. Analysis of variance was used to find statistically significant differences, with significance set at P values less than 0.05. RESULTS: Significant differences were observed in the results of the eight adjustments. The most effective adjustments were the combination of 5-mm distraction with 6 degrees extension or 10-mm distraction alone. The worst adjustment was 5 mm of compression. CONCLUSIONS: Restoring compromised neural spaces in a patient with burst fracture is necessary. The choice of a device adjustment was found to be an important factor in the decompression of the neural spaces after the burst fracture. Combined distraction with extension and distraction alone were found to decompress the canal andintervertebral foramens maximally in a burst fracture. PMID- 10870139 TI - The effect of kyphosis on the mechanical strength of a long-segment posterior construct using a synthetic model. AB - STUDY DESIGN: This experimental study used synthetic spine models to compare the effect of the angle of kyphosis, rod diameter, and hook number on the biomechanical stiffness of a long-segment posterior spinal construct. OBJECTIVE: To examine the biomechanical effects of incremental kyphosis on variously instrumented long-segment posterior spinal constructs. SUMMARY OF BACKGROUND DATA: Euler's formula for loading of curved long columns would suggest that kyphosis has a profound impact on the biomechanical behavior of long-segment posterior spinal constructs. The effects of sagittal contour on the mechanical properties of long-segment posterior spinal constructs have not been well documented. METHODS: Kyphotic and straight synthetic spine models were used to test long-segment posterior instrumentation constructs biomechanically while varying rod diameter and the number of hook sites. The synthetic spines, composed of polypropylene vertebral blocks and isoprene elastomer intervertebral spacers, were fabricated with either 0 degrees, 27 degrees, or 53 degrees of sagittal contour. The models were instrumented with 5.5- or 6.35-mm titanium rods, and with either 8 or 12 hooks. The models were loaded from 0 to 300 N in a cyclical ramp fashion using an MTS 858 Bionix testing device testing device. Construct stiffness (force and displacement) during axial compression was determined. RESULTS: Straight model: Changing the hook number from 8 to 12 caused a 32% increase in construct stiffness with the 5.5-mm rod. Changing the rod diameter from 5.5 to 6.35 mm caused a 36% increase in construct stiffness with the 8-hook pattern. Changing both the rods and hooks caused the stiffness to increase 44%. 27 degrees MODEL: Changing the hook number from 8 to 12 caused a 20% increase in construct stiffness with the 6.5-mm rod. Changing the rod diameter from 5.5 to 6.35 mm caused a 29% increase in construct stiffness with the 12-hook pattern. Changing both the rods and hooks caused the construct stiffness to increase 26%. 53 degrees MODEL: Changing the hook number from 8 to 12 caused a 14% increase in construct stiffness with the 6.35-mm rod. Changing the rod diameter from 5.5 to 6.35 mm caused a 17% (P<0.0005) increase in construct stiffness with the 12 hookpattern. Changing both rods and hooks caused the stiffness to increase 21%. Summary data on angular kyphosis: Using the same rod diameter and the same number of hooks, and progressing from a straight alignment to 27 degrees of sagittal contour decreased construct stiffness 32%. Going from straight alignment to 53 degrees decreased the stiffness 59.6%. All reported values were statistically significant (P < 0.0005). CONCLUSIONS: The biomechanical stiffness of the straight spine was sensitive to both an increase in hook fixation sites and an increase in rod diameter. The kyphotic spines, however, were more sensitive to variations in rod diameter. Although with increasing kyphosis, the optimum instrumentation strategy will maximize both rod diameter and the number of hook sites, instrumented kyphotic spines remain biomechanically "disadvantaged" as compared with nonkyphotic instrumented spines. PMID- 10870140 TI - The role of bone graft force in stabilizing the multilevel anterior cervical spine plate system. AB - STUDY DESIGN: The role of bone graft force in stabilizing an instrumented cervical spine was evaluated for one-level and three-level corpectomy models using in vitro experiments. OBJECTIVES: To investigate the role of bone graft force in enhancing stability of anterior cervical plate, and to study effects of fatigue loading. SUMMARY OF BACKGROUND DATA: The anterior cervical plate system is used widely in stabilizing the cervical spine after spinal corpectomy and grafting. Many factors such as applied screw torque, screw pullout force, plate strength, plate geometry, and type of bone graft have been studied. However, the role of bone graft in stabilizing the anterior plate system has not been explored. METHODS: Two models (one-level and three-level) incorporating corpectomy, strut graft, and anterior plate were constructed from eight human spine specimens (C2-T1). The flexibility of an intact specimen and two constructs with graft forces of 0 N and 100 N was determined. A flexibility test, simulating physiologic loads, consisted of pure moments of flexion, extension, lateral bending, and axial torques up to 1 Nm. For each moment, range of motion and neutral zone were determined. The stability potential index was defined as the decrease in motion caused by instrumentation, as compared with intact motion. A larger stability potential index indicates a more stable spinal construct. Repeated measures analysis of variance was used to determine the significant changes. RESULTS: In both models, bone graft force increased during extension, decreased during flexion, and showed minor changes during axial torsion and lateral bending. Higher bone graft force increased stability potential index neutral zone and stability potential index-range of motion in the three-level model in all directions, but only in flexion-extension in the one-level model. Fatigue loading decreased bone graft force to a greater extent in the three-level model. CONCLUSIONS: In the corpectomy-graft-anterior-plate model, graft force decreased in flexion and increased in extension. Higher graft force increased and fatigue decreased stability of the spinal construct in the three-level model. PMID- 10870141 TI - Placement of pedicle screws in the human cadaveric cervical spine: comparative accuracy of three techniques. AB - STUDY DESIGN: This investigation was conducted in two parts. In the first part, a morphometric analysis of critical cervical pedicle dimensions were measured to create guidelines for cervical pedicle screw fixation based on posterior cervical topography. In the second part of the study, a human cadaver model was used to assess the accuracy and safety of transpedicular screw placement in the subaxial spine using three different surgical techniques: 1) using surface landmarks established in the first part of the study, 2) using supplemental visual and tactile cues provided by performing laminoforaminotomies, and 3) using a computer assisted surgical guidance system. OBJECTIVE: To assess the accuracy of transpedicular screw placement in the cervical spine using three surgical techniques. SUMMARY OF BACKGROUND DATA: A three-column fixation device implanted to secure an unstable cervical spine can be a valuable tool with a biomechanical advantage in the spine surgeon's armamentarium. Despite this advantage, concerns over surgical neurovascular complications have surfaced. Cadaver-based morphometric measurements used to guide the surgeon in the placement of a pedicle screw show significant variability, raising legitimate concerns as to whether transpedicular fixation can be applied safely. METHODS: Precise measurements of 14 human cadaveric cervical spines were made by two independent examiners of pedicle dimensions, angulation, and offset relative to the lateral mass boundaries. On the basis of this analysis, guidelines for pedicle screw placement relative to posterior cervical topography were derived. In the second part of the study, 12 human cadaveric cervical spines were instrumented with 3.5-mm screws placed in the pedicles C3-C7 according to one of three techniques. Cortical integrity and neurovascular injury were then assessed by obtaining postoperative computed tomography scans (1-mm cuts) of each specimen. Cortical breaches were classified into critical or noncritical breaches. RESULTS: Linear measurements of pedicle dimensions had a wide range of values with only fair interobservercorrelation. Angular measurements showed similarangulation in the transverse plane (40 degrees ) at each level. With respect to the sagittal plane, both C3 and C4 pedicles were oriented superiorly relative to the axis of the lateral mass, whereas the C6 and C7 pedicles were oriented inferiorly. The dorsal entry point of the pedicle on the lateral mass defined by transverse and sagittal offset had similar mean values with wide ranges, although there often was excellent correlation between observers. There were no significant interlevel, right/left, or male/female differences noted with respect to offset. Using one of three techniques, 120 pedicles were instrumented. In group 1 (morphometric data): 12.5% of the screws were placed entirely within the pedicle; 21.9% had a noncritical breach; and 65. 5% had a critical breach. In group 2 (laminoforaminotomy), 45% of the screws were within the pedicle; 15.4% had a noncritical breach; and 39.6% had a critical breach. In group 3 (computer assisted surgical guidance system), 76% of the screws were entirely within the pedicle; 13.4% had a noncritical breach; and 10.6% had a critical breach. Regardless of the technique used, the vertebral artery was the structure most likely to be injured. CONCLUSIONS: On the basis of the morphometric data, guidelines for cervical spine pedicle screw placement at each subaxial level were derived. Although a statistical analysis of cadaveric morphometric data obtained from the cervical spine could provide guidelines for transpedicular screw placement based on topographic landmarks, sufficient variation exists to preclude safe instrumentation without additional anatomic data. Insufficient correlation between different surgeons' assessments of surface landmarks attests to the inadequacy of screw insertion techniques in the cervical spine based on such specific topographic guide PMID- 10870142 TI - Hematogenous pyogenic spinal infections and their surgical management. AB - STUDY DESIGN: Mainly a retrospective study of 101 cases of pyogenic spinal infection, excluding postoperative infections. Data were obtained through medical record review, imaging examination, and patient follow-up evaluation. SUMMARY OF BACKGROUND DATA: Hematogenous pyogenic spinal infection has been described variously as spondylodiscitis, discitis, vertebral osteomyelitis, and epidural abscess. Recommended treatment options have included conservative methods (antibiotics and bracing) and surgical intervention. However, a comprehensive classification that would aid in diagnosis, treatment planning, and prognosis has not yet been devised. OBJECTIVES: To analyze the bacteriology, pathologic entities, complications, and results of treatment options for pyogenic spinal infection. METHOD: All patients received plain radiographs, gadolinium-enhanced magnetic resonance imaging scans, and bone/gallium radionuclide studies. All patients had tissue biopsies. Bacteriology, hematology, and predisposing factors were analyzed. All patients received intravenous and oral antibiotics. A total of 58 patients underwent surgery. Patient outcomes were correlated with clinical status, with treatment method and, where applicable, with location and nature of epidural compression. Statistical analyses were performed. RESULTS: Spondylodiscitis occurred most commonly with primary epidural abscess, spondylitis, discitis, and pyogenic facet arthropathy, all occurring rarely. Staphylococcus aureus was the main organism. Infection elsewhere was the most common predisposing factor. Leukocyte counts were elevated in 42.6% of spondylodiscitis cases. The erythrocyte sedimentation rate was elevated in all cases of epidural abscess. There were 35 cases of epidural abscess (frank abscess, 29; granulation tissue, 6). Epidural abscess complicating spondylodiscitis occurred most often in the cervical spine, followed by thoracic and lumbar areas. The rate of paraplegia or paraparesis also was highest in cervical and thoracic regions. There were no cases of quadriplegia. All patients with either epidural granulation tissue or paraparesis recovered completely after surgical decompression. Only 18% of patients with frank epidural abscess and 23% of patients with paralysis recovered completely after surgical decompression. Patients with spondylodiscitis who were treated nonsurgically reported residual back pain more often (64%) than patients treated surgically (26.3%). CONCLUSIONS: Pyogenic spinal infection can be thought of as a spectrum of disease comprising spondylitis, discitis, spondylodiscitis, pyogenic facet arthropathy, and epidural abscess. Spondylodiscitis is more prone to develop epidural abscesses in the cervical spine (90%) than the thoracic (33.3%) or lumbar (23.6%) areas. Thecal sac neurocompression has a greater chance of causing neurologic deficit in the thoracic spine (81.8%). Treatment of neurologic deficit caused by epidural abscess is prompt surgical decompression, with or without fusion. Patients with frank abscess had less favorable outcomes than those with granulation tissue, and paraplegia responded to treatment more poorly than paraparesis. Surgery was preferable to nonsurgical treatment for improving back pain. PMID- 10870143 TI - A comparison of single-rod instrumentation with double-rod instrumentation in adolescent idiopathic scoliosis. AB - STUDY DESIGN: A consecutive series of patients with idiopathic scoliosis treated with single-rod instrumentation was followed prospectively. Outcomes were compared with results obtained from a retrospective review of a consecutive series of patients treated with double-rod instrumentation. OBJECTIVE: To compare single-rod instrumentation with segmental fixation with double-rod instrumentation for the treatment of adolescent idiopathic scoliosis. SUMMARY OF BACKGROUND DATA: Mechanical testing of single-rod instrumentation with segmental fixation at every level showed it to be as resistant to torsion as a double-rod construct. A clinical trial was initiated to document the clinical outcome in single-rod patients. METHODS: A total of 43 of 51 consecutive patients underwent spinal fusion with a single rod. Outcome was evaluated at a minimum of 2 years after surgery. The control group comprised 103 patients who had standard double rod instrumentation at the same institution. RESULTS: The single- and double-rod groups were similar with respect to age, sex, curve type, length of follow-up, curve magnitude, and best bend. For King III-V curves undergoing posterior spinal fusion, there was significantly less blood loss in the single-rod group (703 mL vs 1011 mL), less cell saver collection (189 mL vs 367 mL), and less operating time (220 minutes vs 260 minutes). Blood loss and operating time were not different for patients with King I and King II curves. There were eight patients (19%) requiring reoperation because of hardware-related problems in the single rod group compared with four (4%) in the double-rod group. There were nine patients (21%) with broken rods in the single-rod group, six of whom were symptomatic and five of whom required reoperation. Two patients required multiple operations because of pseudarthrosis in the single-rod group. There were no broken rods in the double-rod group. The single-rod group had 2 early postoperative infections and no late infections compared with 10 late infections in the double-rod group. There was a statistically significant relationship between hardware problems and fusion below L1 in the single-rod group. CONCLUSION: Because of rod failure, single-rod instrumentation should be considered only in curves that can be instrumented to L1 and higher. PMID- 10870144 TI - A noninvasive anthropometric technique for measuring kyphosis and lordosis: an application for idiopathic scoliosis. AB - STUDY DESIGN: Cross-sectional measurement of the sagittal geometry of adolescent idiopathic scoliosis patients. OBJECTIVES: To evaluate the accuracy of a noninvasive anthropometric approach for the measurement of kyphosis and lordosis. SUMMARY OF BACKGROUND DATA: Noninvasive approaches were developed to estimate the sagittal curvatures of the spine. However, the magnitude of the estimation error could be high for an important proportion of patients, which leads to a difficult clinical application. METHODS: The group was composed of 124 female patients with a mean age of 13.5 years (SD 2. 7 years) with Cobb angles ranging from 4 degrees to 66 degrees. Kyphosis and lordosis were measured on the lateral radiograph. The spine sagittal curvature of the same patients was also estimated using the spatial localization of skin markers placed overlying the spinous processes. These coordinates served as input into a simple trigonometric model. Data were collected by means of a stereovideographic technique (Motion Analysis Corp., Santa Rosa, CA). RESULTS: The intraclass correlation coefficient between both approaches was 0.94 for kyphosis and 0.91 for lordosis; the mean absolute differences were 5 degrees (SD 4 degrees ) and 6 degrees (SD 6 degrees ), respectively. The difference was less than 10 degrees in 91% of the patients for kyphosis, and in 79% for lordosis. CONCLUSIONS: The proposed technique appears to give more representative results than those presented in the literature. It has the advantage of being part of a global noninvasive postural evaluation. Using this approach in a systematic manner could help reduce radiograph exposure while keeping track of the spine sagittal curvatures. PMID- 10870145 TI - Magnetic resonance anatomic study of iliocava junction and left iliac vein positions related to L5-S1 disc. AB - STUDY DESIGN: An in vivo anatomic study analyzing the venous anatomy in the lumbosacral area was performed. OBJECTIVES: To obtain in vivo data concerning iliocava junction and left common iliac vein positions at L5-S1. SUMMARY OF BACKGROUND DATA: The left common iliac vein and the iliocava junction are at risk during L5-S1 anterior lumbar interbody fusion. Anatomic studies have demonstrated great interindividual variability in this vascular anatomy. METHODS: Magnetic resonance angiography was used to study 134 patients. Image processing was carried out with maximum intensity projection algorithm and the maximum intensity projection and addition algorithm. Iliocava junction position was measured in the maximum intensity projection and addition image. Four groups of junction position were established: very high, high, low, and very low. The left common iliac vein position was measured in axial magnetic resonance images, and three groups were established: lateral, intermediate, and medial. To describe the operative window delimited by the venous structures at L5-S1, the study population was classified into 12 configurations by combining junction position and vein position values. RESULTS: Very high lateral included 3.76% of the patients, high lateral 48.12%, high intermediate 10.53%, high medial 0.75%, low lateral 15.04%, low intermediate 4.51%, low medial 6.77%, very low lateral 0.75%, very low intermediate 2.26%, and very low medial 7.52%. Medial vein position was significantly more frequent in men. CONCLUSIONS: In 18.05% of the study population, the venous structures overlapped the center of the L5-S1 disc, reducing the operative window. PMID- 10870146 TI - Is there increased intervertebral mobility in isthmic adult spondylolisthesis? A matched comparative study using roentgen stereophotogrammetry. AB - STUDY DESIGN: By roentgen stereophotogrammetric technique, the intervertebral mobility of the spondylolytic segment in eight patients was measured and compared with the mobility of eight nonspondylolytic patients matched according to sex, afflicted segment, and grade of disc degeneration. OBJECTIVES: To compare the intervertebral mobility of a spondylolytic segment with the mobility of a segment without spondylolysis in adult patients with back pain. SUMMARY OF BACKGROUND DATA: Evidenced by the resulting olisthetic deformity and supported by the outcome from prior investigations, spondylolysis is assumed to induce spinal segmental instability/hypermobility. METHODS: After percutaneous application of tantalum indicators for roentgen stereophotogrammetric technique, the intervertebral translations of the spondylolytic fifth lumbar vertebra were measured in eight adult patients with low back pain and low-grade olisthesis. Eight other patients without spondylolysis but with low back pain presumably on degenerative basis were chosen for comparison and had an identical measuring procedure using roentgen stereophotogrammetric technique. The two groups were matched in pairs according to sex, afflicted segment, and grade of disc degeneration. RESULTS: No significant difference was registered considering the intervertebral mobility for matched pairs in the two groups neither along the sagittal nor the vertical axis. The transverse translations were mostly negligible in both groups. CONCLUSION: The spondylolytic defect in pars interarticularis does not cause permanent instability/hypermobility detectable in the adult patient with low back pain and low-grade olisthesis. PMID- 10870147 TI - Aerobic fitness testing in patients with chronic low back pain: which test is best? AB - STUDY DESIGN: This is a randomized comparison of three exercise tests in a sample of 30 patients with chronic low back pain. OBJECTIVES: To determine, by comparing three exercise tests, which test yields the highest peak and predicted oxygen consumption in a sample of patients with chronic low back pain. SUMMARY OF BACKGROUND DATA: Little is known about the level of aerobic fitness in patients with chronic low back pain, although many rehabilitation programs emphasize aerobic exercise as an important part of their therapy. Measurement of aerobic fitness levels in these patients remains a problem. In healthy individuals, the highest oxygen consumption values come from exercise tests that use the largest muscle groups. For a number of reasons, this may not be true in patients with chronic low back pain. METHODS: In this study, 30 participants with chronic low back pain performed three symptom-limited maximal exercise tests: a treadmill, an upper extremity ergometer, and a bicycle ergometer. The tests were administered in randomized order. Heart rate was continuously monitored and oxygen consumption in terms of mL/kg/minute was measured by indirect calorimetry each 30 seconds. RESULTS: The statistical difference among the tests was highly significant (P < 0.0001). The treadmill test yielded the highest peak and predicted oxygen consumption followed by the bicycle and the upper extremity ergometer test, respectively. CONCLUSIONS: The treadmill test is the best test for measuring aerobic fitness levels in patients with chronic low back pain. It yielded the highest peak oxygen consumption compared with the other tests, coming closest to measuring maximal oxygen consumption. PMID- 10870148 TI - Surgery versus conservative management in adult isthmic spondylolisthesis--a prospective randomized study: part 1. AB - STUDY DESIGN: A prospective randomized study was performed. OBJECTIVE: To determine whether posterolateral fusion in patients with adult isthmic spondylolisthesis results in an improved outcome compared with an exercise program. SUMMARY OF BACKGROUND DATA: In spondylolisthesis, satisfactory results have been reported with both surgical and conservative management. The evidence for treatment efficacy, however, is weak because prospective randomized studies are lacking. METHODS: In this study, 111 patients were randomly allocated to an exercise program (n = 34) or posterolateral fusion with or without transpedicular fixation (n = 77). The inclusion criteria were lumbar isthmic spondylolisthesis of any grade, at least 1 year of low back pain or sciatica, and a severely restricted functional ability in individuals 18 to 55 years of age. Pain and functional disability were quantified before treatment and at 1- and 2-year follow-up assessments by visual analog scales (VAS). RESULTS: The 2-year follow up rate was 93%. The functional outcome, as assessed by the Disability Rating Index and the pain reduction, was better in the surgically treated group than in the exercise group at both the 1- and 2-year follow-up assessments (P < 0.01). In the longitudinal analysis, the mean Disability Rating Index and pain improved in the surgical group (P < 0.0001). In the exercise group, the Disability Rating Index did not change at all, whereas the pain decreased slightly (P < 0.02). CONCLUSIONS: Surgical management of adult isthmic spondylolisthesis improves function and relieves pain more efficiently than an exercise program. PMID- 10870149 TI - Instrumented and noninstrumented posterolateral fusion in adult spondylolisthesis -a prospective randomized study: part 2. AB - STUDY DESIGN: A prospective randomized study was performed. OBJECTIVE: To determine whether transpedicular fixation improves the outcome of posterolateral fusion in patients with adult isthmic spondylolisthesis. SUMMARY OF BACKGROUND DATA: The use of transpedicular fixation remains controversial. Both a positive effect and no effect from additional transpedicular fixation have been reported. METHODS: In this study, 77 patients randomly underwent posterolateral fusion with (n = 37) or without (n = 40) transpedicular fixation. The inclusion criteria were lumbar isthmic spondylolisthesis of any grade, at least 1 year of low back pain or sciatica, and severely restricted functional ability in individuals 18 to 55 years of age. RESULTS: The follow-up rate was 94%. At a 2-year follow-up assessment, the level of pain and functional disability were strikingly similar in the two groups, and there was no significant difference in fusion rate. CONCLUSIONS: Lumbar posterolateral fusion performed in situ for adult isthmic spondylolisthesis relieves pain and improves function. The use of supplementary transpedicular instrumentation does not add to the fusion rate or improve the clinical outcome. PMID- 10870150 TI - Comparison between the outer table and intracortical methods of obtaining autogenous bone graft from the iliac crest. AB - STUDY DESIGN: A prospective study in two groups of patients selected randomly. OBJECTIVES: To determine whether keeping the outer and inner cortices of the ilium intact, while obtaining bone graft, would result in reduced postoperative bleeding and less postoperative pain. SUMMARY OF BACKGROUND DATA: Donor site complications after harvesting bone from the iliac crest are frequent. They comprise pain and bleeding related to the large bone exposed, injuries to the cluneal nerve, and sacroiliac instability. METHOD: Sixty patients who were admitted for elective fusion of lumbar segments were included in the study. In half of them, the iliac bone graft was taken in the outer Table method (group A), which included the outer cortex and the cancellous bone beneath, and in the remaining 30 patients only the cancellous bone from between the cortices was collected (group B). The amount of bone harvested, and the time taken to obtain it, were measured, as was the blood volume in the drains. At fixed intervals after surgery and up to 2 years thereafter, the patients were asked to grade the severity of pain in their back and at the donor site. RESULTS: Two years after surgery, 22% of the patients in group A and 17% of the patients in group B reported to have significant pain at the donor site. This difference was not found to be statistically significant, nor was the postoperative bleeding. The average amount of bone harvested in group A was 36 grams compared with 25.7 grams in group B, taking 14 minutes and 20.3 minutes, respectively, to harvest it. These differences were found to be statistically significant. CONCLUSIONS: Preserving the iliac cortices, while obtaining bone graft, does not reduce the postoperative bleeding or the severity of pain at the donor site. In the intraosseous method, less bone is harvested and longer duration of surgery is required, compared with that of the outer Table method. PMID- 10870151 TI - Cervical disc herniation in a patient with congenital insensitivity to pain: a case report. AB - STUDY DESIGN: A case report of a patient with a known diagnosis of congenital insensitivity to pain who developed a herniated cervical disc. OBJECTIVES: To study the clinical manifestations of cervical radiculopathy in a patient with congenital insensitivity to pain and the long-term outcome after surgical treatment. SUMMARY OF BACKGROUND DATA: There have been no reports in the English literature documenting such a patient. METHODS: Retrospective case report and long-term clinical and radiographic follow-up. RESULTS: This patient with a known diagnosis of congenital insensitivity to pain had neurologic motor weakness with "neck and shoulder pain." Clear radicular pattern could not be elicited. The patient underwent a successful anterior discectomy and fusion with long-term clinical and radiographic results. CONCLUSION: Patients with congenital insensitivity to pain who develop a cervical disc herniation may present with atypical symptoms not manifesting in the classic radicular pattern. Higher index of suspicion by the clinician must be practiced to make the appropriate diagnosis. Successful surgical outcome may be achieved in these patients. PMID- 10870152 TI - Postoperative synergistic gangrene after spinal fusion. AB - STUDY DESIGN: A case of synergistic necrotizing gangrenous fasciitis after spinal surgery is reported. OBJECTIVES: To describe this unusual complication, explain the rationale of treatment, and increase awareness of this serious postoperative complication. SUMMARY OF BACKGROUND DATA: Although several cases of postoperative synergistic necrotizing fasciitis have been reported, there are no previously reported cases of this condition after spinal surgery. METHODS: A rapidly progressive necrotizing spinal wound infection after fusion for degenerative disc disease was treated in a 39-year-old man. RESULTS: The infection was successfully treated with serial debridements, appropriate antibiotics, and hyperbaric wound oxygenation. CONCLUSIONS: The authors suggest adherence to the fundamental principles of treatment including radical surgical debridement and appropriate antibiosis for necrotizing gangrene after spinal surgery. In evaluation of aggressive spinal wound infections, diagnosis of synergistic necrotizing fasciitis should be kept in mind. Although hyperbaric wound oxygenation was implemented as an adjunct and appeared to aid in controlling the infection, its effect on outcome is not clear. PMID- 10870153 TI - Liposarcoma in the epidural space. AB - STUDY DESIGN: A patient with myxoid liposarcoma in the lumbar epidural space is reported. OBJECTIVE: The subject was treated with marginal resection and posterior instrumentation. SUMMARY OF THE BACKGROUND DATA: Liposarcoma is a malignant tumor of the soft tissues. It is commonly seen in the thigh. Lumbar extradural space is an unusual localization. Two cases have been reported with this localization in the literature. METHODS: The authors have treated a female patient with myxoid liposarcoma in lumbar extradural space with marginal resection and posterior instrumentation. RESULTS: The histopathologic examination showed myxoid liposarcoma. Two cases have been reported in the literature. CONCLUSION: Myxoid liposarcoma is a malignant tumor of the soft tissues. The extradural tumor was probably originated from the epidural fat tissue. Although wide resection is advised in the thigh localization, extradural localization of the tumor can be treated with marginal resection if there is no invasion to the surrounding tissue. PMID- 10870154 TI - Septic arthritis of a lumbar facet joint caused by Staphylococcus aureus. AB - STUDY DESIGN: Case report of a 35-year-old woman with septic arthritis of a lumbar facet joint. OBJECTIVES: To report a rare case of severe low back pain and the specific differential diagnostic problems. SUMMARY OF BACKGROUND DATA: Differential diagnosis between spondylodiscitis and facet joint septic arthritis on a clinical basis is very difficult. The lesions of the joint appear on a plain film only approximately 1.5 months after onset of the symptoms. Although the radionuclide bone scan is sensitive and shows a more laterally and vertically localized uptake than in spondylodiscitis, this technique is not very specific. Computed tomography scan and magnetic resonance imaging are the most reliable investigations even at the very early stages of the disease. Confirmation of the diagnosis has to be obtained by blood cultures or, in exceptional cases, by direct puncture of the joint. Appropriate antibiotic treatment is in most cases sufficient to heal this lesion. METHODS: The etiology, clinical presentation, technical examinations, and treatment are reviewed. RESULTS: Computed tomography scan and magnetic resonance imaging complemented by positive blood cultures led to the very early diagnosis of septic arthritis of the lumbar facet joint in this relatively young patient. CONCLUSIONS: With our case report we confirm the very small number of data reported in the literature, indicating that infections of the facet joint can be detected at a very early stage using magnetic resonance imaging and computed tomography scan. PMID- 10870156 TI - Imagery. PMID- 10870155 TI - Re: Does spinal kyphoitc deformity influence the biomechanical characteristics of the adjacent motion segment? An in vivo animal model. PMID- 10870157 TI - Whither the Hong Kong Medical Journal? PMID- 10870158 TI - Temporal lobe resection for intractable epilepsy: review of 11 cases. AB - OBJECTIVE: To review the management of medically intractable epilepsy by performing temporal lobe resection. DESIGN: Retrospective study. SETTING: University teaching hospital, Hong Kong. PATIENTS: Eleven patients: seven women and four men (mean age, 28 years; range, 19-49 years) who underwent temporal lobe resection for intractable epilepsy from 1994 through 1998. MAIN OUTCOME MEASURES: Preoperative and operative aspects of treatment, postoperative complications, mortality, and seizure control before and after surgery. RESULTS: All but one patient had long-standing medically intractable temporal lobe epilepsy; the duration between the onset of seizure and surgery ranged from 12 to 27 years (mean, 17.2 years). A total of 12 resections were performed without any mortalities or major postoperative complications. After surgery, two patients became seizure-free without the need for antiepileptic medication; six patients were seizure-free but required medication; and two patients showed >90% of improvement in seizure control, whereas one patient showed between 50% and 90% of improvement. Nine (81%) of the 11 patients reported significant improvement in their social life and performance of daily activities. Two (18%) patients, including one with improved seizure control, reported no improvement in their performance of daily functions. CONCLUSIONS: Temporal lobe resection can produce significant improvements in patients who have medically intractable epilepsy. The risks of surgery are relatively small and justifiable. PMID- 10870159 TI - The shaken baby syndrome: review of 10 cases. AB - OBJECTIVE: To study the characteristics of the shaken baby syndrome from 10 cases in Hong Kong. DESIGN: Retrospective study. SETTING: Regional public hospital, Hong Kong. PATIENTS: Six boys and four girls (mean age, 0.54 years; range, 0.18 1.42 years) in whom the shaken baby syndrome was diagnosed between January 1994 and June 1998. MAIN OUTCOME MEASURES: Clinical features at presentation, radiological findings, management, and outcome. RESULTS: All 10 patients presented with coma: the mean score on the Glasgow coma scale was 4.8 (range, 3 10). In all 10 cases, the history provided by the carers were incompatible with the patient's presentation. Nine patients presented with seizures. Retinal haemorrhages were detected in all patients, but peripheral signs of bruising were observed in only three. Acute subdural haematoma was found in eight patients; one of the remaining two children had subarachnoid and subdural haemorrhages, whereas the other had subarachnoid and intracerebral haemorrhages. Skeletal fractures were detected in two patients. The suspected abusers included either or both parents (n=3), childminders (n=3), and maids (n=2); the identity of the abusers were unknown in two cases. Prosecution by the police was initiated in three cases and two abusers were found to be guilty. Three children died of the abuse; the seven survivors had significant neurological handicaps. CONCLUSION: Medical practitioners should be alert to the occurrence of abusive head injury in children. Peripheral signs are uncommon and a high degree of suspicion is needed for the management to be successful. PMID- 10870160 TI - Genetic linkage study of family members of a patient with adult polycystic kidney disease. AB - OBJECTIVE: To study the feasibility of making an early diagnosis of adult polycystic kidney disease by using genetic linkage analysis in Hong Kong. DESIGN: Genetic linkage study. SETTING: University teaching hospital, Hong Kong. PARTICIPANTS: Six members of a Chinese family with a history of adult polycystic kidney disease. MAIN OUTCOME MEASURES: The inheritance pattern of adult polycystic kidney disease, as detected by polyacrylamide gel electrophoresis of polymerase chain reaction products using radioactively labelled primers specific to six microsatellite DNA markers that are closely linked to the PKD1 gene on chromosome 16. RESULTS: Four of the six members of the family studied were shown to be positive for disease-linked markers, and the inheritance of adult polycystic kidney disease could be traced in this family with a higher degree of precision (93.7%) using genetic linkage analysis, than could be predicted otherwise. CONCLUSION: The success of genetic linkage analysis in providing an early diagnosis of adult polycystic kidney disease is dependent on having a sufficient number of family members whose disease status has been established by imaging methods to allow the disease-linked marker haplotype to be determined. The establishment of a genetic data bank for families with adult polycystic kidney disease should be considered to maximise the effectiveness of this diagnostic approach. PMID- 10870161 TI - Women's knowledge of and attitudes towards emergency contraception in Hong Kong: questionnaire survey. AB - OBJECTIVE: To study the level of knowledge of and attitude towards emergency contraception in a group of women requesting the termination of pregnancy. DESIGN: Structured questionnaire survey. SETTING: Family Planning Association and university teaching hospital, Hong Kong. PARTICIPANTS: Two hundred women who requested the termination of an unplanned pregnancy between May 1997 and March 1998. MAIN OUTCOME MEASURES: Demographic data, basic knowledge of contraception, reasons for terminating the pregnancy, and knowledge and usage of emergency contraception. RESULTS: A substantial proportion (33.0%) of women was ignorant of the existence of emergency contraception. Only 10.0% of women had used emergency contraception before and only 2.5% had used it in an attempt to prevent this pregnancy. Of the 134 women who knew about emergency contraception, the main reason (41.8%) for not using it was risk-taking behaviour. More nulliparous women (88.5% versus 57.6%; P<0.001) and women younger than 20 years (84.0% versus 61.3%; P<0.01) had heard of emergency contraception. Women who were educated beyond secondary school level (71.0% versus 37.5%; P<0.01) and unmarried women compared with married, cohabiting, or divorced women (87.1% versus 49.5%; P<0.001) were also more likely to have heard of emergency contraception. Women younger than 20 years were more likely to have used this form of birth control in the past (18.0% versus 7.3%; P<0.05). CONCLUSION: There is a need to improve women's education about emergency contraception in Hong Kong. PMID- 10870162 TI - Hepatic resection for primary liver cancer at a private community hospital: retrospective study of 61 patients. AB - OBJECTIVE: To review the outcome after surgical resection for primary liver cancer. DESIGN: Retrospective study. SETTING: Private community hospital, Hong Kong. PATIENTS: Sixty-one consecutive patients who underwent liver resection for primary cancer from 1992 through 1997. MAIN OUTCOME MEASURES: Clinicopathological features, type of resection, duration of hospital stay, and actuarial overall and disease-free 5-year survival rates. RESULTS: Cirrhosis was present in 46 (75%) of the patients, and 42 (69%) were positive for hepatitis B surface antigen. The median tumour diameter was 8 cm (range, 1-16 cm). Liver resections consisted of hemihepatectomy (n=37), trisegmentectomy (n=4), segmentectomy (n=11), and wedge resection (n=9). Postoperative death and major morbidity occurred in 0% and 36% of patients, respectively; ascites was the most common complication. The median hospital stay was 11 days. The actuarial overall and disease-free 5-year survival rates were 36.0% and 22.8%, respectively. These results are similar to or better than those recently reported from local or overseas centres. Multivariate analysis showed that the Child-Pugh class significantly influenced the development of complications and the length of hospital stay, whereas a well circumscribed tumour margin, the tumour-node-metastasis stage of the tumour, and the Child-Pugh class were independent predictors of survival. CONCLUSION: Surgical resection for primary liver cancer can be performed with acceptable safety and efficacy in a suitably staffed and equipped private community hospital. PMID- 10870163 TI - Intoeing gait in children. AB - OBJECTIVE: To review the aetiology and management of intoeing. DATA SOURCES: Medline and non-Medline literature search, and personal experience. STUDY SELECTION: Studies that provided evidence-based information about the aetiology and management of paediatric intoeing gait were selected. DATA EXTRACTION: Data were extracted and reviewed independently by both authors. DATA SYNTHESIS: An intoeing gait affects many children and, as with flexible flatfoot, bowleg, and knock-knee, it falls into the category of physiological problems that occur in normal children. The usual causes are excessive femoral anteversion, internal tibial torsion, and metatarsus adductus. Management is based on understanding the causes and the natural course of the condition and the effectiveness of various treatment modalities. Unfortunately, due to poor understanding of the condition, intoeing is commonly overtreated with braces or special footwear. CONCLUSIONS: Intoeing is one of the most common conditions encountered in paediatric orthopaedic practice. It is important to make an early diagnosis of pathological causes of intoeing such as cerebral palsy and developmental dysplasia of the hips so that treatment can be commenced as soon as possible. PMID- 10870164 TI - Changes in chemotherapy for pancreatic cancer. AB - OBJECTIVE: To review the systemic chemotherapy regimens for pancreatic cancer. DATA SOURCES: Medline and non-Medline literature search (1966-1999). STUDY SELECTION: The following key words were used: pancreatic carcinoma; chemotherapy; antineoplastic agent; fluorouracil; gemcitabine. DATA EXTRACTION: Reports of phase II studies, randomised controlled studies, and preclinical studies were reviewed. DATA SYNTHESIS: Less than 20% of patients are suitable candidates for surgery; for the remainder, palliative chemotherapy is of only marginal benefit. Combining fluorouracil with folinic acid or interferon has not led to any significant improvement in tumour response or the patient survival rate. The early encouraging results with combination chemotherapy have not been confirmed in subsequent controlled studies. New approaches include immunotherapy and novel cytotoxic drugs. In vitro studies of monoclonal antibodies have shown promise but have failed to show clinical efficacy. Recently, gemcitabine has been shown to be more effective than fluorouracil in delivering pain relief and reducing disease related symptoms. CONCLUSION: Systemic chemotherapy is generally ineffective in increasing the survival time of patients with pancreatic cancer. Future clinical investigations concerning treatment should focus on gemcitabine-based combination chemotherapy or combined modality treatment with radiotherapy. PMID- 10870165 TI - Primary immunoglobulin A nephropathy through the 'retrospectroscope' AB - OBJECTIVE: To review pathological approaches to the diagnosis and prognosis of primary immunoglobulin A nephropathy. DATA SOURCES: Medline and non-Medline literature search (1966-1999) and personal experience. STUDY SELECTION: The following key words were used: IgA nephropathy, pathology, grading. DATA EXTRACTION: Data were extracted and analysed independently by the authors. DATA SYNTHESIS: Primary immunoglobulin A nephropathy is the most common glomerular disease worldwide and this also holds true in Hong Kong, where it represents the most common condition encountered in diagnostic renal biopsy examinations. This type of chronic nephropathy currently has no effective curable therapy, and in a significant proportion of patients, it progress to end-stage renal disease. Supportive treatment is very important, because it may alter the natural course and slow down the progression of this nephritis. CONCLUSION: Pathological grading of immunoglobulin A nephropathy currently represents the most useful method to appraise the renal outcome. PMID- 10870166 TI - Management of chronic subdural haematoma: burr hole drainage, replacement with Hartmann's solution, and closed-system drainage. AB - Although the treatment of chronic subdural haematoma by burr hole drainage has been performed in the past with or without using a closed drainage system, the problem of intracranial air entrapment still persists and can cause a deterioration in the level of consciousness or seizures in the postoperative period. Cerebral infarction may also develop a few days after surgery because of the intracranial hypotension that occurs during the drainage procedure. In an attempt to minimise these complications and to prevent cerebral infarction and its attendant morbidity, we have developed a technique of treating chronic subdural haematoma-namely, performing burr hole drainage, irrigation and replacement of the haematoma with Hartmann's solution, and closed-system drainage of the subdural space with a silicone catheter. The blood pressure is closely monitored and maintained by the infusion of fluids throughout the procedure. An illustrative case using this technique is presented in this paper. PMID- 10870167 TI - Carotid endarterectomy for carotid stenosis: audit at a tertiary referral centre. AB - Prospective randomised controlled trials performed in North America and Europe have demonstrated that the risk of future stroke or death is substantially reduced by performing carotid endarterectomy in symptomatic patients who have severe carotid stenosis. An audit was conducted to analyse the results of carotid endarterectomy performed during a 3-year period, at a tertiary referral vascular centre in Hong Kong. A total of 35 patients who had significant carotid stenosis underwent 36 carotid endarterectomies from October 1994 to September 1997. All patients recovered uneventfully without neurological complications or mortality. The audit showed that the current results of carotid endarterectomy at this institution fulfilled the criteria advised by the Stroke Council of the American Heart Association in 1995. Therefore, carotid endarterectomy is a validated therapeutic option for carotid stenosis in Hong Kong. PMID- 10870168 TI - Carotid endarterectomy for non-hemispheric cerebrovascular symptoms: an unusual indication. AB - We report on an 80-year-old man who presented with non-hemispheric cerebrovascular symptoms in the form of daily multiple syncope. The left common carotid artery, its two main divisions, and the right vertebral artery were completely occluded. There was high-grade stenosis in the right carotid artery (82%) and left vertebral artery (60%). After excluding other causes of syncope such as postural hypotension, hypoglycaemia, cardiac arrhythymia, and epileptic seizure, a diagnosis of global ischaemia was made. The patient subsequently underwent carotid endarterectomy and the symptoms were relieved. This case represents an unusual indication for carotid endarterectomy. PMID- 10870169 TI - Antithyroid drug-induced agranulocytosis. AB - Thyrotoxicosis is a common endocrine disorder. Antithyroid drug therapy is the standard treatment for this disease, especially in young women of reproductive age. A serious side effect of antithyroid drug use, however, is agranulocytosis. We report on two patients with antithyroid drug-induced agranulocytosis. Both patients presented with fever and severe neutropenia. The administration of granulocyte colony-stimulating factor resulted in a dramatic improvement in the white blood cell count and symptoms. Antithyroid drug-induced agranulocytosis is a potentially lethal condition but is completely reversible when recognised early and when prompt treatment is offered. PMID- 10870170 TI - Vertebral artery dissection: a treatable cause of ischaemic stroke. AB - Vertebral artery dissection used to be an uncommon diagnosis but it is now being diagnosed more frequently owing to the use of magnetic resonance imaging. We report on three patients with vertebral artery dissection to illustrate the importance of establishing this diagnosis by using magnetic resonance imaging in patients who present with cerebrovascular accident that involves the posterior territory. Treatment with heparin can help prevent recurrent embolic events and should be given in the absence of subarachnoid haemorrhage or other contra indications. PMID- 10870171 TI - Familial form of arrhythmogenic right ventricular dysplasia presenting with recurrent ventricular tachycardia. AB - We report on a 48-year-old man who presented with recurrent sustained monomorphic ventricular tachycardia, which resulted in syncope on one occasion. Subsequent investigation confirmed the diagnosis of arrhythmogenic right ventricular dysplasia. Familial screening for the disease was conducted using 12-lead electrocardiography, signal-averaged electrocardiography, and magnetic resonance imaging. Two of the four relatives who were screened showed evidence of the disease, thus confirming the familial form of arrhythmogenic right ventricular dysplasia in the patient. He underwent a subpectoral implantation of an implantable cardioverter defibrillator in view of the malignant nature of the ventricular tachycardia and his intolerance to drug treatment. PMID- 10870172 TI - Emergence of Streptococcus pneumoniae with high-level resistance to cefotaxime in Hong Kong. AB - We report on two cases of pneumococcal infection caused by strains demonstrating high-level cefotaxime resistance (minimal inhibitory concentration, 4 ug/mL). One patient had acute community-acquired meningitis with bacteraemia and the other had bacteraemia probably as a result of nosocomial pneumonia. Both patients died despite treatment with third generation cephalosporins. This is the first report from Hong Kong of infection with Streptococcus pneumoniae with high-level cefotaxime resistance that resulted in death. The emergence of high-level resistance to third-generation cephalosporins will result in treatment failure when these agents or penicillin are used alone, especially in cases of severe infection, such as meningitis, in which drug penetration of the blood-brain barrier is critical. The treatment of severe infections due to these isolates is problematic. Indiscriminate use of life-saving third-generation cephalosporins as out-patient treatment of minor infections or as first-line therapy for uncomplicated community-acquired infections in the hospital and in the community should be discouraged. PMID- 10870173 TI - Multiple skeletal muscle metastases. PMID- 10870174 TI - What did we learn from the Shanghai hepatitis A epidemic? AB - A major outbreak of hepatitis A (HAV), associated with consumption of raw clams, occurred in Shanghai, China in 1988. Over 300 000 cases were reported, of which 47 (0.015%) were fatal. An elevated mortality rate was observed in hepatitis B surface antigen (HBsAg)-positive patients (0.05%). The majority of these patients were also hepatitis B e antigen (HBeAg)-positive, indicating active liver disease and high viral replication rates. The increased mortality in hepatitis B virus (HBV)/HAV coinfected individuals is hypothesized to be the result of T-cell mediated destruction of HBV-infected hepatocytes, enhanced by acute HAV infection. Following recovery from HAV there is an increase in HBV expression and activated cytotoxic cells and subsequent cytolysis. Patients with chronic HBV infection are clearly at considerable risk of severe disease and increased mortality in the event of HAV infection. The period of greatest risk is during the immunoeliminative phase of HBV infection, which generally occurs in early adulthood. With the prevalence of HBV approaching 10% in this group, there is a clear opportunity for benefit from vaccination. PMID- 10870175 TI - Hepatitis A in patients with chronic liver disease - severity of illness and prevention with vaccination. AB - Acute hepatitis A in patients with pre-existing chronic liver disease (CLD) may theoretically predispose such patients to a more severe outcome of infection. The results of reports covering epidemics and case series suggest but do not universally support this possibility. An analysis of the larger case series support the suggestion that acute hepatitis A superimposed on chronic hepatitis B infection is associated with higher peak laboratory abnormalities, more severe disease and a higher fatality rate. The safety and immunogenicity of an inactivated hepatitis A vaccination was evaluated in patients with CLD and found to be satisfactory for the protection of this group of patients. PMID- 10870176 TI - Hepatitis C and the British Columbia experience with hepatitis A vaccination. AB - The prevalence of hepatitis C virus (HCV) infection in British Columbia is approximately 1.5%. Experience in this province has revealed that groups at high risk of HCV infection are also prone to outbreaks of hepatitis A virus (HAV). As hepatitis A infection can be fatal in HCV-positive individuals, numerous strategies have been implemented to help minimize its spread. The importance of vaccinating individuals at high risk for hepatitis C infection against HAV has been recognized. PMID- 10870177 TI - Is lycopene beneficial to human health? AB - Since humans cannot synthesise carotenoids de novo, we depend upon the diet exclusively for the source of these micronutrients. Although the necessity for beta-carotene, as the precursor of vitamin A has been recognised for many years, it is lycopene that has attracted substantial interest more recently. Lycopene is the red-coloured carotenoid predominantly found in tomato fruit, but in few other fruits or vegetables. It has claimed that it may alleviate chronic diseases such as cancers and coronary heart disease. This possibility has been studied extensively, by epidemiological studies and biochemical investigations of its properties and its bioavailability from tomato-based diets. This article summarises the current state of knowledge of the properties of lycopene, its possible role in human health and areas for future research. PMID- 10870178 TI - Application of mass spectrometry for identification and structural studies of flavonoid glycosides. AB - Mass spectrometry is an important tool for the identification and structural determination of flavonoid glycosides. The advantages of mass spectrometry are high sensitivity and possibilities of hyphenation with liquid chromatographic methods for the analysis of mixtures of compounds. Different desorption ionization methods allow the analysis of underivatized glycosides. A review of mass spectrometric techniques applied to the identification and structural studies of flavonoid glycosides is presented. PMID- 10870179 TI - Monoterpene synthase activities in leaves of Picea abies (L.) Karst. and Quercus ilex L. AB - In addition to direct ecological functions in the interaction of plants with the environment, the emission of monoterpenes, especially from the foliage of evergreen trees, is of great importance for the production of ozone and photochemical oxidants in the troposphere. In the present work, we established a reproducible non-radioactive standard enzyme assay and characterized monoterpene synthase activities in needles of Norway spruce (Picea abies (L.) Karst.) and in leaves of holm oak (Quercus ilex L.). In Norway spruce, the dominant monoterpenes formed were alpha-pinene, camphene, and to a lesser extent beta-pinene and limonene. In holm oak, alpha-pinene, sabinene, and beta-pinene were the main products, while limonene was a minor component. Under optimum conditions, in both Norway spruce and holm oak, monoterpene formation remained constant up to 180 min and 90 min, respectively, and varied with the buffer and Mg2+ and Mn2+ concentrations used. Optimum temperature for monoterpene synthase activity was 40 degrees C in both species; optimal pH ranged between 6.5 and 7.5 in both species. Apparent Michaelis-constants for the substrate GDP were ca. 17.9 +/- 5.1 microM for Norway spruce and ca. 69.4 +/- 22.1 microM for holm oak. Molecular weight determination by FPLC indicated that the monoterpene synthases in Norway spruce and holm oak have native molecular weights of ca. 59 and 50 kDa, respectively. PMID- 10870180 TI - Taxonomic distribution of plant glutathione S-transferases acting on xenobiotics. AB - Soluble and microsomal glutathione S-transferase activities for five model xenobiotics (nitrobenzene derivatives), two pesticidal xenobiotics (atrazine and fluorodifen), and a natural substrate (cinnamic acid), were determined in 59 different plant species and four plant cell suspension cultures. These enzyme activities were widely distributed over the plant kingdom with certain species showing particularly high activities. Marine macroalgae had a remarkably broad substrate range that included the substrates atrazine and fluorodifen. It is concluded that the evolutionary 'green liver' concept derived for xenobiotic metabolism in higher plant species is also valid for the constitutive soluble and microsomal glutathione S-transferases of lower plant species. PMID- 10870181 TI - Phototropic stimulation induces the conversion of glucosinolate to phototropism regulating substances of radish hypocotyls. AB - The distribution of natural growth inhibitors, the raphanusanins (isomers of 3 (methylthio)methylene-2-pyrrolidinethione) and their precursors (4-methylthio-3 butenyl glucosinolate (MTBG) and 4-methylthio-3-butenyl isothiocyanate (MTBI), between illuminated and shaded halves of radish hypocotyls during phototropic curvature was analyzed using a physicochemical assay. Phototropic stimulation rapidly decreased MTBG content, and abruptly increased contents of MTBI and raphanusanins in the illuminated halves of radish hypocotyls within 30 min after the onset of unilateral illumination. Content in the shaded halves was similar to that in dark controls. When MTBG, MTBI, and raphanusanins at endogenous levels were applied unilaterally to etiolated hypocotyls, MTBI and raphanusanins caused hypocotyls to bend but MTBG showed no activity. Blue illumination promoted myrosinase (thioglucosidase) activity, which releases MTBI from MTBG, in hypocotyls after 10 min, although enzyme activity in dark controls did not change. These results suggest that phototropic stimulation promotes myrosinase activity in the illuminated side of radish hypocotyls, releasing bioactive MTBI from inactive MTBG and simultaneously producing bioactive raphanusanins. PMID- 10870182 TI - Two antifeedant lignans from the freshwater macrophyte Saururus cernuus. AB - Two diarylbutane derivatives of dihydroguaiaretic acid have been isolated from emergent portions of the southeastern United States freshwater angiosperm Saururus cernuus L. (Saururaceae). Bioassay-guided fractionation of organic extracts of S. cernuus led to the compounds, sauriols A and B, in addition to five previously known lignoids. These metabolites deter feeding by the omnivorous crayfish Procambarus clarkii. The two lignans were identified by analysis of nuclear magnetic resonance and mass spectral data, and by comparison with spectral data of dihydroguaiaretic acid. PMID- 10870183 TI - Lipid profile: a useful chemotaxonomic marker for classification of a new cyanobacterium in Spirulina genus. AB - The morphological, physiological and genetic characteristics of an isolate cyanobacterium from hard sand of the lake Venere in the Pantelleria island (Italy) were described. The isolate with a small-size coiled helix shape, growing optimally at pH 9.2-9.5 at 30 degrees C under continuous illumination and aeration, possessed a 61.5 mol% of Guanine + Cytosine content of DNA. The lipid profile showed the presence of mono-, di-glycosyl, sulphoquinovolosyl and phosphatidyl (MGDG, DGDG, SQDG and PG). The fatty acid profile was also studied, characterized by the absence of gamma-linolenic acid and the presence of saturated and monounsaturated C16 and C18. The latter was also present as a dienoic component. The fatty acid composition was affected by growth temperature by increasing the degree of desaturation at a lower temperature and the biosynthesis of shorter acyl chains. The effects of growth conditions other than temperature, physical, nutritional and chemical on lipid composition were also studied. The overall features of the cyanobacterium isolated from Pantelleria clustered it into Spirulina genus. PMID- 10870184 TI - Fatty acids and tocochromanols in seeds of Orobanche. AB - The evaluation of tocochromanols (tocopherols and tocotrienols) in 49 accessions from 21 Orobanche species revealed three well separated groups. The first one, characterized by high gamma-tocotrienol content, included all the accessions of sect. Orobanche. The second one, exhibiting high gamma-tocopherol content, comprised the accessions of O. arenaria Borkh. and O. purpurea Jacq. (sect. Trionychon Wallr.). All the other accessions of this section presented high delta tocopherol content. Differences for tocochromanol derivatives within sect. Trionychon were paralleled by differences in the fatty acid profile, with the high delta-tocopherol class having also a higher oleic to linoleic acid ratio. PMID- 10870185 TI - Secondary metabolites characteristic of Penicillium citrinum, Penicillium steckii and related species. AB - Two new carboxylic acids, tanzawaic acid E (1) and F (2) in addition to the unknown benzopyran 3,7-dimethyl-1,8-dihydroxy-6-methoxy-isochroman (3), and the known mycotoxin 3,7-dimethyl-8-hydroxy-6-methoxyisochroman (4) were produced by a marine-derived strain of Penicillium steckii isolated from an unidentified tunicate. The carboxylic acids and the benzopyran were identified on the basis of mass spectrometry, and one and two dimensional NMR spectroscopic techniques. The structures 1 and 2 resemble tanzawaic acid A-D, previously isolated from Penicillium citrinum. Screening of isolates of species related to P. citrinum and P. steckii showed that P. citrinum (25 isolates) consistently produced citrinin and tanzawaic acid A, P. steckii (18 isolates) produced isochroman toxins (except 2) and tanzawaic acid E, P. sizovae consistently produced tanzawaic acid A, P. corylophilum (10 isolates) produced citreoisocoumarinol and P. sumatrense (15 isolates) always produced curvularin. PMID- 10870186 TI - 3-acetylaltholactone and related styryl-lactones, mitochondrial respiratory chain inhibitors. AB - A novel furano-pyrone, 3-acetylaltholactone, and two other known styryl-lactones, altholactone and 5-acetoxyisogoniothalamin oxide, have been isolated from Goniothalamus arvensis (Annonaceae) stem bark. We report here the isolation and structural elucidation of these compounds with furane-pyrone and styryl-pyrone skeletons, postulating also for the first time their mechanism of cytotoxicity based on inhibition on mammalian mitochondrial respiratory chain. PMID- 10870187 TI - Covalent anthocyanin-flavonol complexes from flowers of chive, Allium schoenoprasum. AB - The structures of eight anthocyanins have been determined in acidified methanolic extract of pale-purple flowers of chive, Allium schoenoprasum. Four of them have been identified as the anthocyanin-flavonol complexes (cyanidin 3-O-beta glucosideAII) (kaempferol 3-O-(2-O-beta-glucosylFIII-beta-glucosideFII)-7-O-beta gl ucosiduronic acidFIV) malonateAIII (AII-6-->AIII-1, FIV-2-->AIII-3), 1, (cyanidin 3-O-(3-O-acetyl-beta-glucosideAII) (kaempferol 3-O-(2-O-beta glucosylFIII-beta-glucosideFII)-7-O-beta-gl ucosiduronic acidFIV) malonateAIII (AII-6-->AIII-1, FIV-2-->AIII-3), 2, and their 7-O-(methyl-O-beta glucosiduronateFIV) analogous, 3 and 4. Pigments 1 and 2 are the first final identification of covalent complexes between an anthocyanin and a flavonol, while 3 and 4 are formed during the isolation process. The other four anthocyanins (5 8) were found to be the 3-acetylglucoside, 3-glucoside, 3-(6-malonylglucoside) and 3-(3,6-dimalonylglucoside) of cyanidin. The three latter pigments have earlier been identified as the major anthocyanins of the chive stem. The covalent anthocyanin-flavonol complexes show intramolecular association between the anthocyanidin (cyanidin) and flavonol (kaempferol) units, which influence the colour. PMID- 10870188 TI - Composition of lipids from sunflower pollen (Helianthus annuus). AB - The contents of the pollen lipids of the sunflower Helianthus annuus are described. The major component is the seco-triterpene helianyl octanoate, followed by new beta-diketones as second major group of compounds. They exhibit a shorter chain length and often other positions of the functional group compared to already known beta-diketones. Of particular note are the 1-phenyl-beta diketones, not previously reported from nature. Further lipid classes present are related hydroxyketones and diols. Interestingly, new beta-dioxoalkanoic acids are present in the extracts, which most likely are biogenetic precursors of the diketones. Additionally, we investigated the composition of the pollen coat which resembles the total extract, but lacks the dioxoalkanoic acids and certain estolides. PMID- 10870189 TI - 8-O-methyldioncophyllinol B and revised structures of other 7,6'-coupled naphthylisoquinoline alkaloids from Triphyophyllum peltatum (Dioncophyllaceae). AB - The isolation and structural elucidation of a new naphthylisoquinoline alkaloid, 8-O-methyldioncophyllinol B, from Triphyophyllum peltatum (Hutch. et Dalz.) Airy Shaw (Dioncophyllaceae) is described, together with the revised structures of other 'B-type' compounds previously misidentified as dioncophylline D, dioncophyllinol D, and 8-O-methyldioncophylline D. All of the presently described structures are 7,6'-coupled and thus have to be addressed as 'B-type' naphthylisoquinoline alkaloids. This is in contrast to the initially defined 'D type' structures, which are 7,8'-coupled as confirmed by a total synthesis of dioncophylline D. Some of these natural and synthetic naphthylisoquinolines were found to display good in vitro antiplasmodial activities. PMID- 10870190 TI - [Thymectomy in myasthenia gravis: long-term evolution and predictive factors]. AB - INTRODUCTION: Thymectomy is a surgical procedure which is generally accepted for treatment of myasthenia gravis. OBJECTIVE: To describe the long-term evolution of 217 myasthenic patients who had had thymectomies in the Hospital Clinico Quirurgico Hermanos Ameijeiras in La Habana, Cuba. PATIENTS AND METHODS: We determined the stage of evolution of 217 patients, followed-up periodically, with an average observation time of 83.4 months and an interval of between 5 and 155 months. We also studied the frequency of remission and the influence of a group of variables on this, as well as the mortality and its causes. RESULTS: The clinical state of the patients was: remission 77 (35.4%); pharmacological remission 45 (20.7%); significant improvement 70 (32.2%); the same or worse 5 (2.3%); deaths 11 (5%), and unknown 9 (4.1%). The cases with thymomas had a more unsatisfactory course than the other patients, with fewer remissions and greater mortality. Over the first five years of the evolution of the disease, there was a 30% rate of remission; after five years this rose to 35-40% and after ten years reached 47%. The age of the patient, duration of symptoms, histology of the thymus, age of onset of the disease and duration of follow-up did not have any significant effect (p < 0.05) on the long-term evolution of the thymectomy, in contrast to the gravity of the condition according to Osserman's scale. CONCLUSION: The results of thymectomy, in our study were good and similar to the results reported in the international literature. PMID- 10870191 TI - [Study of higher brain function in first grade students and its relationship with reading and writing acquisition]. AB - INTRODUCTION: The study of higher cortical functions gives valuable information and new insights in the understanding of the complex functioning of the central nervous system. Developmental neurological examination is an important semiologic tool in the evaluation of cortical functions. OBJECTIVE: The main of this study was to evaluate, using the neurological developmental exam, cortical functions and their association with learning. PATIENTS AND METHODS: This was an observational, analytic and transversal study with a random and proportional sample (484 children) of first grade students of Porto Alegre, Brazil. Chi-square and ANOVA tests were used, with a significant p value < 0.05. RESULTS AND CONCLUSION: Results show that altered neurologic performance disturbs reading and writing acquisition. This study, showed an association between altered neurological developmental examination and WISC subtests (numbers, figure completion and code), with learning disorders. PMID- 10870192 TI - [Early prognosis in chronic subdural hematomas. Multivariate analysis of 137 cases]. AB - INTRODUCTION: In the literature there is evidence relating different factors such as age and preoperative clinical condition with prognosis in patients treated surgically for chronic subdural haematoma. OBJECTIVE: To clarify and quantify the magnitude of the factors which determine early prognosis (during hospital admission) of these patients. PATIENTS AND METHODS: We made a prospective study of 137 patients who had been operated on in our centre and found the relationship between different clinical and therapeutic variables with the clinical course and morbidity-mortality by means of multivariate and survival analysis. RESULTS: A high Markwalder functional score (3-4) is an independent factor of poor prognosis (OR = 13.15; CI 95% 6.1-28.4; p = 0.01), as is the presence of a coexisting coagulopathy (OR = 27.2; CI 95% 9.3-79.5; p = 0.01). Advanced age tended to increase the risk (OR = 1.104) but did not reach statistical significance (p = 0.0654). A multivariate logistic model, which included the functional score and presence of coagulopathy, correctly classified 94.7% of the cases studied. Analysis of survival showed two groups with different early mortality as a function of the Markwalder score (high: 3-4 and low: 0-1-2), which could be differentiated statistically (Log-Rank chi squared test: 3.95; p = 0.0468). CONCLUSIONS: The preoperative clinical state classified by functional scores and the presence of underlying coagulopathy are the main prognostic factors in chronic subdural haematoma during hospital admission. Advanced age is probably not in itself an independent factor for bad prognosis. PMID- 10870193 TI - [Implementation of an indirect method for the measurement of memory in the elderly]. AB - INTRODUCTION: There is interference between emotional factors and intellectual output in the elderly, in whom there is a high prevalence of depression. Low output in conventional psychological tests may lead to false positive diagnoses of dementia. PATIENTS AND METHODS: We evaluated 48 patients aged between 50 and 84 years with regard to their emotional and intellectual state by means of the SCAN system. We also used a test designed by ourselves to implicitly examine memory. This consisted of a task involving naming objects. Six stimuli were repeated three times, seeking an effect of perceptual representation of 'priming' which facilitated later recall. RESULTS: The results showed that patients with dementia recalled 0 to 2 of the repeated stimuli, as a result of weakened priming. Meanwhile the non-demented persons, in spite of having other neurological disorders answered between 4 and 6 times. Multivariate analysis of the variance showed that measurement of memory, both implicit and explicit, allowed discrimination between patients with and without dementia. However, the explicit results were not independent of the effects of depression. CONCLUSIONS: Implicit measurements of memory were superior to direct or explicit measurements in the diagnosis of dementia. Patients with other degenerative conditions, such as Parkinsonism and multisystemic atrophy, had above average results. This corroborates the view that the neurological basis of this type of memory is not at subcortical level. PMID- 10870194 TI - [Chronic tension-type headache and depression]. AB - INTRODUCTION: Tension-type headache (TTH) is the commonest head pain to be seen in medical practice and is erroneously thought to have a psychogenic origin. OBJECTIVE: To evaluate the presence of depression in a cohort of persons with TTH and compare them with patients with migraine and with a control group. The clinical characteristics of patients with TTH were also noted. PATIENTS AND METHODS: We studied 89 patients with chronic TTH. Their level of depression (on the Hamilton scale) was compared with 31 patients with migraine with typical aura and a control group of 34 asymptomatic volunteers, matched for age, marital and job status. RESULTS: No difference was observed in the index of depression of the group of patients with chronic TTH as compared with the control group with migraine: p < 0.9412 and OR 1.07. Compared with the asymptomatic control group: p < 0.0272 and OR 3.81, which are significant figures. After 11 months of prophylactic treatment with imipramine (50 mg/day), levels of depression persisted in 20.02% of the patients, p < 0.06319 and OR 2.01 even when treatment was optimum in 98.8%. CONCLUSIONS: Patients with chronic TTH showed indices of depression in 33.7%. This figure was similar in those with migraine, 32.2%. Also response to treatment was independent of this figure. In 48% of the patients there was fear of having cerebral vascular disease. PMID- 10870195 TI - [Multiple sclerosis and epileptic seizures]. AB - INTRODUCTION: Although epileptic seizures are uncommon in multiple sclerosis they are more prevalent than in the general population, which supports an aetiological relationship. Similarly in a considerable proportion of patients with multiple sclerosis and epileptic seizures, alterations in magnetic resonance and electroencephalogram studies which could be correlated with the clinical features of epilepsy were observed. Nevertheless, it is difficult to establish definite clinical characteristics in these patients since the underlying pathogenic mechanisms are poorly understood and there is great variability with regard to the type of seizure, point at which this occurs during the course of the disease, degree of recurrence and other aspects. CLINICAL CASE: We report the clinical, electroencephalographical and neuroimaging findings of seven patients with multiple sclerosis who had epileptic seizures and those in whom there was no evidence of other potentially epileptogenic pathology. In two patients the epileptic seizures formed part of the first episode of their illness. One patient presented more than one type of epileptic seizure. These seizures were generalized in two cases, partial sensory and/or motor with secondary generalization in three, simple partial motor in one and partial complex in two. The epileptic seizures coincided with other clinical features of episodes in three cases and the electroencephalogram showed anomalies in five cases. CONCLUSIONS: The findings observed were of a wide variety, as was found in other reported series. We point out certain correlations between the clinical data, magnetic resonance and electroencephalogram which may help to orientate the management of these patients. PMID- 10870196 TI - [Lhermitte-Duclos disease associated with tuberous sclerosis. A case report and review of literature]. AB - INTRODUCTION: Lhermitte-Duclos disease is a rare disorder of the cerebellum of unknown origin in which dysplasic thickening of the cerebellar convolutions is seen. It usually occurs in young adults. Currently it is included in the phacomatosis group of disorders. CLINICAL CASE: A 19 year old woman attended the Emergency Department complaining of progressive orthostatic headache for the previous three months. On examination there were striking facial micronodular lesions suggestive of angiofibromas, a hypo-pigmented macula in the inframammary region and a hyperpigmented 'cafe-au-lait' macula in the right hypochondrium. On computerized tomography there was tetraventricular hydrocephalia. Cerebral magnetic resonance showed significant descent of the tonsils, hypertensive hydrocephalia and a lesion in the left cerebellum, apparently laminar hyperintensity in DP and T2, with thickening of some folia, not enhanced by intravenous contrast and suggestive of a dysplasic gangliocytoma. Laboratory investigations showed subclinical hypothyroidism. Other investigations were normal. The patient was treated by implanting a ventriculo-peritoneal shunt which has relieved the symptoms to date. CONCLUSIONS: Lhermitte-Duclos disease is probably not a single anatomo-clinical condition, assuming that it may be a cerebellar hamartoma associated with a phacomatosis with few clinical signs, whether it be Cowden's disease, tuberous sclerosis as in this case or an 'overlapping' syndrome. The magnetic resonance findings are necessary and sufficient for the diagnosis of Lhermitte-Duclos disease. PMID- 10870197 TI - [Occipital leptomeningeal angiomatosis without facial angioma. Could it be considered a variant of Sturge-Weber syndrome?]. AB - INTRODUCTION: The association of cerebral leptomeningeal angioma and facial nevus flameus in the territory of the first branch of the trigeminal nerve ipsilateral to the angioma is known as the Sturge-Weber syndrome. The cases with absence of a facial angioma are usually considered to be variants of the syndrome. OBJECTIVE: To present four cases with occipital leptomeningeal angioma without facial angioma and describe the characteristics which differentiate them from or permit their inclusion within the group of Sturge-Weber syndrome, and also to establish the differences between this and the Gobbi syndrome (occipital cerebral calcifications, epilepsy and coeliac disease. CLINICAL CASES: We selected four cases in whom cranial magnetic resonance was done with intravenous gadolinium and three cases studied to rule out coeliac disease. The cerebral calcifications, unilateral in all four cases, were similar to those observed in the Sturge-Weber syndrome. All cases had leptomeningeal angiomas at the level of the cerebral calcification shown by the uptake of contrast material on magnetic resonance. Three patients had epilepsy but none had facial angiomas, hemiparesis or glaucoma. Coeliac disease was also ruled out, both on laboratory investigations and on intestinal biopsy. CONCLUSIONS: The cases described coincide with the Sturge-Weber syndrome in all having cerebral leptomeningeal angiomas. This differentiated them from the Gobbi syndrome which does not include meningeal angiomata. Another characteristic of the Sturge-Weber syndrome is the occurrence of epilepsy and mental deficiency. Whilst awaiting molecular genetic studies, our cases may be included semantically as a variant of the Sturge-Weber syndrome without the characteristic facial angioma, although they may possibly correspond to genetically different conditions. PMID- 10870198 TI - [Leptomeningeal carcinomatosis as presenting symptom of a gallbladder carcinoma]. AB - INTRODUCTION: Meningeal carcinomatosis is rare accounting for 4-5% in autopsy of patients with solid tumors, and even less frequent, 1%, in its pure form without brain metastases. We report a case of psychosis symptomatic of a meningeal carcinomatosis as presenting manifestation of a gallbladder carcinoma. This clinicopathological combination has not been described previously. CASE REPORT: 74 years-old man. His past medical history included Parkinson's disease treated with L-dopa 50 mg/8 hour and selegiline; duodenal ulceration and hypertiroidism. He started with delirium and visual hallucinations that do not responded to a reduction of L-dopa and suppression of selegiline. The examination of CSF was diagnostic, malignant cells were identified in the initial examination. The patient dead and his autopsy diagnostic was gallbladder carcinoma with meningeal carcinomatosis. CONCLUSIONS: Leptomeningeal carcinomatosis in its pure form is the infiltration of the leptomeninges without brain metastases. It is less than 1% of meningeal metastases from solid tumors. The most frequent primary tumors are: lung, breast, stomach-esophagus, melanoma, colo-rectal, genital and urinary; and the most frequent histological type is adenocarcinoma. The commonest presenting symptoms are focal brain, medullar or radicular symptoms. Psychiatric as isolated symptom are exceptional. Diagnosis is confirmed by the examination of CSF. Malignant cells appear in the first examination in 45-50% of cases. Leptomeningeal carcinomatosis in gallbladder carcinoma is rare; only four cases has been described previously none of them presenting as isolated psychiatric clinical picture. PMID- 10870199 TI - [How does the nucleus accumbens function?]. AB - INTRODUCTION: The nucleus accumbens is considered as the neural interface between motivation and action, playing a key role on feeding, sexual, reward, stress related, drug self-administration behaviors, etc. DEVELOPMENT: The nucleus accumbens possesses two territories, the core and shell, whose connectivity wiring gives a good picture of its motor and limbic aspects. The shell seems to behave as a 'coincidence detector', which can be activated during behavioral situations of adaptive value, thanks to its connections with prefrontal cortex, amygdala and hippocampus. The activation of the shell leads to the reinforcing of goal-directed motor sequences mediated by the core and prefrontal cortex, areas which are, linked to pyramidal and extrapyramidal motor systems. Dopamine secreted within the nucleus accumbens would acts as a 'neurostabilizer' of such processes. CONCLUSION: The nucleus accumbens is made up of an 'electrophysiological coincidence detector' or shell serially connected to a 'motor sequencer' or core, both supporting the role of the nucleus accumbens as a limbic-motor interface. PMID- 10870201 TI - [A critical review of the specificity of the Wisconsin card sorting test for the assessment of prefrontal function]. AB - INTRODUCTION AND OBJECTIVE: Clinical and experimental research with the Wisconsin Card Sorting Test (WCST) has shown inconsistencies which bring into question the specificity of the test as a marker of frontal dysfunction. The aim of the present review is to evaluate the causes and the consequences of those criticisms for the assessment of both prefrontal function and the executive system of attention. DEVELOPMENT: Clinical evidence confirms that, in its present form, the WCST can not discriminate between lesions in frontal and non frontal brain regions. Moreover, functional neuroimaging studies have shown rapid and widespread activation of frontal and non frontal brain regions during WCST performance. On the one hand, these studies strongly suggest that the concept of anatomically 'pure' tasks is deceptive, but they also provide us with evidence that inconsistencies in WCST research might be motivated by problems with the internal validity and reliability of the original test as a measure of attentional set shifting ability. In contrast, recent studies have successfully employed WCST analogues to link precise cognitive processes with anatomically and functionally well defined prefrontal areas. CONCLUSIONS: It is deemed necessary to apply the new technical and methodological developments to generate more valid and reliable neuropsychological tests, that yield a better correspondence between anatomy and function. This will make possible future progress in the clinical assessment of higher brain functions. PMID- 10870200 TI - [Agitation in head injury. I. Definition and treatment with anxiolytic neuroleptics and antiepileptic drugs]. AB - OBJECTIVE: To carry out a bibliographic review of articles indexed in MEDLINE over the past 20 years concerning the pharmacological treatment of agitation in head injury. DEVELOPMENT: Head injury may cause different behaviour changes, of which agitation is the most dramatic. The incidence of agitation after severe head injury varies from 11% to 50% depending on the study involved. This incidence is high enough to warrant specific management. Drug treatment has variable results. When there is imminent danger of harm to the patient himself or to others, or when aggressive behaviour makes medical management difficult, the benzodiazepines have been found useful. Antipsychotic drugs are only indicated in head injury when the agitation causes a clinical emergency, and in such a case the more potent drugs such as haloperidol are best, since they have less sedative effect. They are also effective when the clinical features are similar to those of classical schizophrenia. Antiepileptic drugs have been used successfully for treating agitation-aggressiveness, specially in paroxystic behaviour disorders. We also consider other treatments used for posttraumatic agitation. CONCLUSION: There is no general agreement amongst doctors as to the best treatment for posttraumatic agitation in head injury. However, with regard to certain characteristics of agitation different drugs may be recommended for treatment. PMID- 10870202 TI - [Antiepileptic drugs in the elderly , pregnant women, children and in systemic disorders]. AB - OBJECTIVE: To review the known pharmacokinetic and pharmacodynamic bases of current antiepileptic drugs in the elderly, pregnant and child populations, and in patients with systemic disorders so as to offer recommendations as to their use. DEVELOPMENT: In the elderly it should be pointed out that there is a slower rate of metabolism of drugs due to poorer liver function and also slower renal clearance leading to slower elimination from the body. In children there is slower metabolism in the new-born, but metabolism increases as the child grows and becomes normal when he reaches adolescence. In childhood the adverse effects of antiepileptic drugs may have an important effect on the proper growth of the body. In pregnant women there are several reasons why the levels of antiepileptic drugs may drop, and following child-birth the risk of toxicity increase considerably. The terratogenic effect of antiepileptic drugs is well known and seems to be related to the level of antiepileptic drugs and of polytherapy. Early administration of folic acid may help to avoid some of the more serious malformations. Patients with chronic liver or kidney disease should be treated with drugs eliminated via the more healthy organ. CONCLUSIONS: Because of their specific pharmacological properties, the new antiepileptic drugs would seem to be the treatment of choice in the groups at risk which have been considered, although there is still insufficient experience to recommend their use in pregnant women. PMID- 10870203 TI - [Advances in the treatment of epilepsy: status epilepticus]. AB - OBJECTIVE: The complexity of the treatment of status epilepticus (SE) is due to the wide variety of forms of clinical presentation. In this review we wish to emphasize that satisfactory management of SE requires a system which takes account of the successive phases of the gravity of the electroclinical course of SE and the different types of SE according to the electroclinical semiology of the seizure. DEVELOPMENT: The concepts and classifications which, in current epileptology, are used in SE are mainly based on criteria developed at three international symposia: in Marseilles in 1962 and Santa Monica, California, in 1979 and 1997. Current knowledge permits distinction of different therapeutic periods depending on the chronology of each SE and to classify the SE according to the type of seizures, age of the patient and underlying pathology. CONCLUSIONS: The classifications described permit the standardization of treatment: preventive measure in high-risk patients; immediate and in situ treatment during the prodromal phase; three parallel lines of action- differential diagnosis, general measures and antiepileptic treatment--during the initial phase; measures in hospital emergencies and in the Intensive Care Unit when the SE is at a fully established phase; special measures, including induction of anaesthesia in the refractory phase; transition to long-term treatment, recovery of autonomy by the patient and long-term management in the phases following remission of the SE. The standardization proposed may perhaps serve as a basis for the future development of guidelines. PMID- 10870204 TI - [Evaluation of drug-resistant epilepsy]. AB - OBJECTIVE: Drug-resistant epilepsy makes up between 10 and 30% of all epilepsies, and is an important cause of morbidity and mortality. In this article we review the guidelines and techniques used when assessing a patient with drug-resistant epilepsy. DEVELOPMENT: Evaluation of a patient with drug-resistant epilepsy requires three steps: first a careful, detailed clinical history to establish the semiological data of the seizure, determine the factors predisposing to drug resistance (partial epilepsy, history of severe head injury etc.) and to evaluate the antiepileptic treatment given previously and whether the patient actually took the drugs prescribed. Second, a prolonged video-EEG recording is made to determine the type and site of the seizure and decide whether surgical treatment would be suitable. Thirdly, whether structural imaging investigations (magnetic resonance) or functional imaging (sole photon emission computerized tomography or positron emission tomography) should be done. It is also useful to make neuropsychological and psychiatric studies to detect possible associated defects and preexisting psychiatric pathology--which is very common in this population- in order to establish a rehabilitation programme according to the needs of each patient. CONCLUSION: Careful evaluation of each patient with drug-resistant crises leads to the correct diagnosis of epilepsy with the possibility of its correct classification and localization and may permit improvement or change of the treatment received. PMID- 10870205 TI - [Rational anti-epileptic polytherapy. Drug interactions and choice of treatment]. AB - INTRODUCTION: Following twenty years of antiepileptic biotherapy, monotherapy superseded this when it was shown to be equally effective, have fewer problems of drug interactions and the patients were more likely to actually take their drugs. However, the development of the so-called new antiepileptic drugs gives rational biotherapy a new chance. OBJECTIVE: In this article we discuss the basis of reasoned polytherapy and some of the possible interactions between antiepileptic and other drugs. DEVELOPMENT AND CONCLUSIONS: When polytherapy is considered, antiepileptic drugs with different modes of action and profiles of adverse effects should be used. Some epileptic patients may attain control of their seizures with rational polytherapy. The drug reactions between the antiepileptic drugs themselves and with other drugs are frequent but not usually of clinical importance. We discuss some of the more relevant interactions. Choice of the initial antiepileptic drug is based on various factors, basically the type of seizure, age of the patient, epileptic syndrome, possibility of pregnancy, family and job considerations. PMID- 10870206 TI - [Dissertations about leprosy in the Seville at the end of the Enlightenment Age]. AB - INTRODUCTION: Leprosy is a well-known disease from ancient history. Society reacts violently due to the fear of infection, and the fact that it causes appalling physical mutilation. It is produced by Mycobacterium leprae, which only affects the nervous system of human beings. DEVELOPMENT: The norms and examinations that for many years were practiced upon those suspected of being infected by the leprosy organism are based almost always in a series of requirements that were in keeping with cases of verification, thus named the 'declaration of leprosy'. Doctors in the 18th Century, conscious of the consequences of the disease, established a diagnostic procedure for leprosy. But as a result of the medical limitations of the time, and the innate risk of examination of the sufferer in the early phases and their changing symptoms, they adopted a cautious stance and on occasion were overly prudent. These problems remained established in different dissertations presented in the Royal Society of Medicine and other Sciences of Seville during the last third of the 18th Century. A total of eight dissertations related to this disease are analyzed. Two presented by Doctor Bonifacio Ximenez de Lorite in 1765 and 1788 are noteworthy due to the contents and quality. PMID- 10870207 TI - [Acute disseminated postinfectious encephalomyelitis]. PMID- 10870208 TI - [Mortality due to cerebrovascular diseases in Guanabacoa, Cuba]. PMID- 10870209 TI - [Rabdomyolysis due to influenza A]. PMID- 10870210 TI - [Cyclically alternating recordings and alternate trace]. PMID- 10870211 TI - [Multiple infarcts in vertebro-basilar territory and cardio-embolism]. PMID- 10870212 TI - [Coordination of the activity of monoaminergic pedal neurons in fresh water snails]. AB - In fresh water pulmonate snails, contact with the water surface as well as a tactile stimulation of the mantle wall induces a co-ordinated activity in symmetrical pedal neurones RPeD1 and LPeD1. The GDC and GSC being not primary sensory neurones, their activity is co-ordinated by polysynaptic mechanisms. In both species, a depolarising current applied to the GDC and GSC caused a discharge in one of the columellar nerves. The data obtained suggest that the co ordination of activity in the GDC and GSC may have a functional significance for controlling the left and right halves of the columellar muscle in respiration. PMID- 10870213 TI - [Intracellular cAMP involvement in the synchronized activity of noradrenaline in response to evoked release of the transmitter quanta in the frog synapses]. AB - An analogue of cyclic AMP (db-cAMP) penetrating into the frog neuromuscular junction's cell, as well as the adenylyl cyclase activator forskolin, and inhibitor of nucleotide-depending phosphodiesterase isobutilmethylxantine alter the kinetics of the quanta secretion resulting in synchronizing of the process of the transmitter release. Following a db-cAMP preliminary action, no such synchronizing of the transmitter release occurred. Action of noradrenaline on the time course of the secretion seems to be realised through activation of presynaptic beta-adrenoreceptors, augmentation of the adenylyl cyclase activity, and the rise of the intracellular cAMP. PMID- 10870214 TI - [Functional interrelations between suprachiasmatic nuclei and the hippocampus during elaboration and recovery of time conditioning in rats]. AB - In hippocampectomised rats or in rats subjected to hippocampectomy combined with destruction of the hypothalamic suprachiasmatic nuclei, time conditioning could not be elaborated. The lesion of the suprachiasmatic nucleus alone, however, accelerated elaboration of the conditioned reflex. In denucleated rats, the hippocampectomy did not affect the time of a previous conditioning recovery. PMID- 10870215 TI - [The role of the thalamic ventrolateral nucleus in the cortical effect on the activity of vestibular neurons]. AB - Electrocoagulation of lateral vestibular nucleus (NVL) reduces inhibitory effect of the motor and somatosensory areas and enhances the inhibitory effect of limbic, vestibular, and orbital cortical areas. Facilitating effect was enhanced by electrostimulation of the motor area and reduced by the stimulation of other cortical areas. Following the coagulation of the NVL, the ascending afferent flow to the cortex seems to be reduced. This results in diminishing of the cortical neurones tone and readjusts the descending influences upon the NVL neurones activity. PMID- 10870216 TI - [Local cerebral blood flow, cortical impedance, cerebral intravascular kinin production during forebrain ischemia and reperfusion in rats]. AB - In the rat model of forebrain ischemia with subsequent reperfusion, an obvious formation of intravascular kinin (IVKF) occurred. The IVKF preceded and coincided in time with the maximum hyperemia and a vasogenic oedema. Local cerebral blood flow (LCBF) increased up to about 187%, at that. In three groups of experimental rats, a correlation was revealed among the ischemia obviousness, IVKF, and development of the brain oedema. PMID- 10870217 TI - [Functional activity of blood platelets during cerebral ischemia/reperfusion in rats]. AB - In rats with an incomplete brain ischemia and subsequent reperfusion, the platelets sensitivity to the ADP decreased more obviously during the postischemic reperfusion. The platelets seem to be transformed into a refraction state after their activation in brain ischemia. The platelets refraction state might depend on a secondary activation of the nitric oxide-synthase in the platelets. PMID- 10870218 TI - [Relations between number and volume of erythrocytes and leukocytes in the human blood]. AB - Total concentration of erythrocytes and leukocytes, a relative contents of neutrophils, lymphocytes, monocytes, average volume of these types of cells, were determined in humans. Changes in the cell indices seem to be correlated and sex dependent. Thus, erythrocytes and leukocytes are correlated in their contents and volume characteristics. A control mechanism seems to exist in humans, involving microcirculation in capillaries. PMID- 10870219 TI - [Structural and functional characteristics of hemoglobin molecules from the preserved donor blood during long-term storage]. AB - The human blood haemoglobin molecules keep their quaternary structure for 25 days at a long-term storage of the blood stabilised with glugicyr. The molecules' electronic characteristics did not change during 5 days, and their oxygen-binding ability remained unaltered just during 2 days after taking the blood. PMID- 10870220 TI - [The role of glycolysis in the protective effect of amtizol during acute hypoxia]. AB - In rats, protective effect of amtizol was found to depend on the rate of carbohydrate substrate in the organism as well as on the glycolysis activity. Amtizol was able to activate utilising of carbohydrates in a glycolytic way in satisfied rats with a blockade of glycolysis by monoiodacetate, whereas glyconeogenesis was activated in starving rats. PMID- 10870221 TI - [Nitrogen oxide storage as a factor of adaptive defence]. AB - Adaptation to environmental factors possesses multiple NO-dependent protective effects and stimulates the NO storage. An adaptation to a mild stress was shown to reduce the death rate in rats from 57% to 8% and to prevent a heat shock induced hypotension and endothelial overactivation. Treatment of the rats with the NO-synthase inhibitor L-NNA interfered with the NO storage and formation of protective effects, while the NO donor dinitrosyl iron complex facilitated the NO storage and simulated the adaptive defence. The data obtained suggest an important role of the NO storage in adaptive defence of the organism. PMID- 10870222 TI - [Effect of testosterone and progesterone on carboxypeptidase H activity in the hypothalamo-hypophyseal-gonadal axis and the adrenal gland in mice]. AB - Testosterone propionate decreased activity of carboxypeptidase H in the male mongrel mice pituitary gland, activity of the CP H in testicles, and increased these parameters in 4 hours after treatment. Progesterone decreased the CP H activity in pituitary gland and increased it in testicles. Progesterone increased the CP H activity in female pituitary gland and decreased it in adrenal gland and hypothalamus. The role of the CP H in the effects of the sex steroids on the function of peptidergic systems is discussed. PMID- 10870223 TI - [Blood physiological resources and their role in physiological adaptation of the body to blood loss (based on the experimental data and calculations)]. AB - In white rats, loss of about 66 +/- 3% of the blood volume results in an abrupt drop of the BP and arrest of the lung respiration. Introduction of a plasma substitute in the volume equal to the lost blood resulted in an increase of the BP and restoration of rhythmical respiration in 9 out of 14 experimental rats. Haematocrit after the blood loss and subsequent administration of the substitute was about 15%. Previously obtained data show that the Ht of about 15% decreases the oxygen transport to tissues. However, an increase in the lung ventilation, a shift of the haemoglobin dissociation curve to the right, an increase of oxygen extraction from the blood, and an increase in the minute blood volume by 50-100%, allow a sufficient oxygen transport to the organism tissues to be ensured at the Ht of about 15%. Thus, the reserve of the blood respiration function of the blood remaining after a life-threatening blood loss plays an important physiological role in replenishing the volume of the lost blood with the plasma substitute and in survival of the organism. PMID- 10870224 TI - [Variational chronoreflexometric evaluation of the central nervous system functions in elementary school children with mental retardation]. AB - Parameters of variational chronoreflexometry variation curve of distribution were shown to correlate with the level of mental development in mentally retarded children. The borderline values of the CNS function condition's criteria were determined for both normal children and those mentally retarded. A feasibility of assessing the interhemisphere functional asymmetry by means o this particular method, was shown. PMID- 10870225 TI - [Effect of the protein kinase C inhibitor on changes in Ca(2+) level in pig granulosa cells induced by prolactin]. AB - The effect of the PKC inhibitor on Ca2+ responses to prolactin in the pig granulosa cells was studied using fluorescent dye and chlortetracycline. The effect was shown to be connected with activation of the PKC. The Ro 31-8220 increased penetration of extracellular calcium and exit of calcium from intracellular stores. The data obtained suggest an involvement of the PKC in changes of calcium contents in the pig granulosa cells activated by prolactin. PMID- 10870226 TI - [Expectant management in localized prostatic cancer]. PMID- 10870227 TI - [Renal colic and lithiasis in HIV(+)-patients treated with protease inhibitors]. AB - INTRODUCTION: Protease inhibitors, mainly Indinavir, are widely used drugs for the treatment of patients infected by the human immunodeficiency virus (HIV) and are related to renal colic and urinary obstruction. These conditions are the result of urine excretion of these drugs which favours the formation of small calculi (crystalluria and lithiasis). MATERIAL AND METHODS: Five PI treated HIV(+) patients; four males, one female, have recently been seen for renal colic at the Lithiasis Unit, Fundacion Jimenez Diaz (FJD). All five patients had renal colic, one bilateral and one renal obstruction and fever. Small lithiasic concretions of null or minor radiological calcium density were identified by urinary X-ray and UIV. The patients had haematuria, crystalluria and urinary pH 5.0-6.0. Treatment was symptomatic, pharmacologic, emergency in situ extracorporeal shock-wave lithotrity (ESWL), or ureteral catheterisation, as appropriate. RESULTS: Patients had been treated with these antiviral agents for several months. They all required urologic care: pharmacologic, ureteral catheterisation, or ESWL, with good results. No stones were obtained for mineralogic analysis, but crystalluria was identified as being due to Indinavir and calcium oxalate. CONCLUSIONS: Renal excretion and urinary elimination of PIs (or their metabolites) results in asymptomatic crystalluria in HIV(+) patients treated with this class of drugs. Other cases present genuine calcium oxalate calculi with sings of renal colic and urinary obstruction requiring urologic care. PMID- 10870229 TI - [Self-administered questionnaire for assessing the Raz technique in stress urinary incontinence: analysis of results]. AB - OBJECTIVES: Knowing our results in the treatment of the stress incontinence by means of the use of the Raz technique evaluated by sending a questionnaire. MATERIAL AND METHODS: From July 1995 until the 31st of December 1998, 137 patients were operated by the Raz technique. The 102 people who were monitored for over 12 months were sent a questionnaire asking them about the postoperative evolution and the degree of the satisfaction obtained. From the 78 (76.47%) people who answered, 75 were included in this study with an average age of 53.96 and an average follow-up of 27.12 months. RESULTS: Thirty patients (40%) showed the disappearance of their stress incontinence, seventeen patients (22.7%), an important improvement and ten (13.3%) a slight improvement, but only 58.20% of the patients were very or quite pleased with the results. None of the preoperatives variables analysed showed any meaningful influence in the outcome, while the longest time of persistence of postmiccional residue was shown as a factor of prediction in a better result. CONCLUSIONS: The mediocre results and the low rate of satisfaction got with this technique justifies the research of other therapeutic and more effective options. PMID- 10870228 TI - [Absorptive hypercalciuria type III or caused by renal loss of phosphates]. AB - Contribution of a retrospective series of 16 patients with type II absorptive hypercalciuria over a total of 1.041 patients undergoing metabolic study due to relapsing renal lithiasis. Clinical history of lithiasis, biochemistry prior to treatment and instituted therapy were examined in all cases. Stones composition, radiologic appearance of lithiasis and evolution of biochemical parameters after medical treatment with a mixture of phosphates were also studied. PMID- 10870230 TI - [Patient response to and assessment of intracavernous drug injection and vacuum]. AB - One of the various therapeutic options currently used in erectile dysfunction (ED), vacuum is a procedure that should not be ruled out as it can benefit certain types of patient. To understand each patient's response and usage readiness, a vacuum test is routinely performed by the andrology service as part of the diagnostic study. The present study conducted on 200 patients with ED of different etiologies, included the vacuum test and the intracavernous injection of vasoactive drugs. The assessment included the response to each methodology; which of the two systems offered better response; and initial usage readiness to each system. Positive response was 93% in the vacuum test, and 40% in the intracavernous injection. Improved erection was reported by 73% with vacuum and only 13% with the intracavernous injection. With regard to usage readiness 60.5% favoured vacuum and only 14.5% the intracavernous injection. It was apparent that vacuum is a highly effective methodology with very high initial usage readiness, although ultimate use may be limited for different reasons. PMID- 10870231 TI - [Urethral substitution with synthetic material]. AB - In spite of the numerous surgical techniques described, management of urethral stenosis continues to be an unresolved problem. Free graft urethroplasty is indicated in selected cases and several organic and synthetic materials have been described for this use. Our group reviews the synthetic alloplasts used for partial or total replacement of the male urethra. The search for an appropriate alloplast for urethral replacement has been rather frustrating. Complication rates are still too high to allow routine usage. The most encouraging results are those with new absorbable materials which cause minimal inflammatory reactions of a foreign body type. These are readily available and allow urethral replacement using tissues regenerated from removed ends. PMID- 10870232 TI - [Anterograde insertion of ureteral catheter]. AB - We report twenty-four patients with urinary obstruction, in which twenty-seven antegrade ureteral stent (double J) insertions were attempted (in six patients the obstruction was bilateral and in three other patients we failed). In all of them access to the urinary tract was through a nephrostomy catheter, in seventeen cases we proceeded to insert the antegrade catheter immediately after percutaneous nephrostomy and in ten remaining cases we achieved in a second try after carrying nephrostomy and failing a conventional retrograde approach to ureteral stent insertion. We got a 90-per cent success rate. A case of perirrenal hematoma occurred after applying a nephrostomy. It was the only relevant complication. In conclusion we consider that the antegrade ureteral stent insertion is a good alternative when, under several circumstances, the conventional retrograde insertion fails. PMID- 10870233 TI - [Anatomic and clinical evidence of intrapelvic pudendal nerve and its relation with striated sphincter of the urethra]. AB - OBJECTIVE: To demonstrate that somatic innervation of the urethrae striate sphincter is intrapelvic and not through an internal pudendal nerve, an extrapelvic nerve in its entire path. To study the relationship of the pelvic plexus with genito-urinary organs and its surgical implications. METHODS: 6 embryos and 2 fetuses, sliced and stained with techniques suitable for nervous structures were studied. Sequential observation of the pelvic plexus structures and the internal pudendal nerve was carried out using light microscopy. Three dimensional reconstruction of the two fetuses was performed to study the relationships of these structures with the genito-urinary organs. RESULTS: A nervous branch was identified in the 19, 25 and 30 mm long specimens that started at the internal pudendal nerve and joined the hypogastric ganglion close to the site of pelvic nerves binding. Hypogastric ganglion efferent branches penetrating the striate sphincter after a short descendent run were seen in the 30 and 39 mm embryos. CONCLUSIONS: Our findings confirm that the urethrae striate sphincter received the autonomous and somatic innervation from the pelvic plexus and, therefore, is susceptible to damage during cancer related surgery of the pelvic organs. An improved knowledge of these structures and the use of nerve preservation surgical techniques can reduce the incidence of post-operative incontinence in this type of surgery. PMID- 10870234 TI - [Mullerian-type papillary serous tumor: exceptional pathology of the testis. Report of a case and discussion]. AB - Within the exceptional tumoral pathology of the testis and paratesticular region are the common epithelial type tumors. Although, its histogenesis is under discussion, it has been interpreted as arising from the remnants of the mullerian duct, or from the mesothelium of the tunica vaginalis differentiating in a mullerian direction. Similarities with homologous ovarian tumors are well recognized and in generally we accept a good prognostic due to its low malignant potential or borderline type, more experience with these uncommon lesions is necessary to evaluate their biologic potential. PMID- 10870235 TI - [Lymphangioma of the spermatic cord]. AB - We report a new case of spermatic cord lymphangioma in a infant 2 years old. The initial diagnosis was funicular hydrocele. The treatment was the local excision of tumor and the diagnostic was histological. Postoperative course was excellent. Must be explored the transillumination of the mass which would have led us to think other the diagnosis different from that of the cord hydrocele before the operation since it would have given negative. During the operation, must the assured that the cystic anomaly is limited to spermatic cord, to evite recurrences in the postoperative course. PMID- 10870236 TI - [Penile incarceration with metal rings]. AB - Contribution of a case report of penial incarceration following placement of three thick metal rings and its resolution. Literature review. PMID- 10870237 TI - [Tuberous angioma of the glans penis: treatment with laser and local anesthesia]. AB - A case of angioma tuberous of the glans penis treated with Neodymiun:Yag laser under local anesthesia in a eleven-years-old outpatient is reported. These lesions are extremely rare. Local excision of this tumor was accepted treatment, but we think that laser treatment (Neodymiun:Yag) is far superior to surgical excision and we think this treatment of choice. The particularity of the case we report, is aside the rarity, the possibility of treatment in an outpatient child, with local anesthesia and excellent tolerance. PMID- 10870238 TI - [Inverted papilloma of the prostatic urethra]. AB - The inverted papilloma is a rare urothelial tumor, and its localization at the prosthatic urethra is also exceptional. We present a case of inverted papilloma of the prostatic urethra in a 72 years-old male, with symptoms of urinary flow obstruction. The diagnose is obtained after urethrocistoscopy and transurethral resection at the same time. We discuss about the etiology, clinical presentation, diagnose and treatment of this rare tumor, making special attention to its malignancy ability. PMID- 10870239 TI - [Penile fracture: report of a case]. AB - Traumatic rupture of the corpus cavernosum of the penis is rare, and has been reported infrequently. We present a case with this type of trauma, with immediate surgical management, with good morphologic and functional results. PMID- 10870240 TI - [Bladder hyporeflexia caused by vinca alkaloids]. AB - Presentation of one case of hyporreflexic bladder like a first step of neurotoxicity due to Vinca alkaloids. These drugs produces peripheral neuropathies as usual, but in some rare occasions they may affect to the autonomic nervous system with its effects in the bladder producing hyporreflexic. This disease reverts spontaneously after suppressing drugs. PMID- 10870241 TI - Automatic control of a robot camera for broadcasting based on cameramen's techniques and subjective evaluation and analysis of reproduced images. AB - With the goal of achieving an intelligent robot camera system that can take dynamic images automatically through humanlike, natural camera work, we analyzed how images were shot, subjectively evaluated reproduced images, and examined effects of camerawork, using camera control technique as a parameter. It was found that (1) A high evaluation is obtained when human-based data are used for the position adjusting velocity curve of the target; (2) Evaluation scores are relatively high for images taken with feedback-feedforward camera control method for target movement in one direction; (3) Keeping the target within the image area using the control method that imitates human camera handling becomes increasingly difficult when the target changes both direction and velocity and becomes bigger and faster, and (4) The mechanical feedback method can cope with rapid changes in the target's direction and velocity, constantly keeping the target within the image area, though the viewer finds the image rather mechanical as opposed to humanlike. PMID- 10870242 TI - Deflection in center of gravity at the simulated operations of cabinet in the aged. AB - The study projected a scenario where the aged handled home furniture in their daily lives. With the furniture designated at 5 different heights for task performance, different aspects of the center of gravity(CG) deflection in the young and the aged were investigated. The following points which should be considered were suggested; (1) The findings indicated that remarkable increases in CG deflection were detected in performing tasks on cabinet as age advanced. The age factor appeared to yield the major effect in all indices measured, whether the drawer was being drawn open or closed. (2) The major effects with age accompanied by significant mutual interactions between age and operation height were established when the drawer was drawn open. (3) In cases where the aged drew open the drawer, the LR deflections with reference to the CG were pronounced. When performing the task especially at greater heights, the deflections in the aged were enlarged, a tendency that was in contrast to the extended LR deflections (cf. at lower heights) in the young, contributing therefore to the increases in CG-DD and CG-DA, accordingly. PMID- 10870243 TI - Change in knowledge and reported use of sport science by elite New Zealand olympic class sailors. AB - The objective of this study was to examine the change in knowledge and use of sport science in 46 elite New Zealand Olympic class dinghy sailors' one year after the adoption of a sport science support (SSS) programme by Yachting New Zealand. Twenty eight (22 males, six females) sailors responded to a questionnaire which was administered during a training camp in April 1994 and 28 (also 22 males and six females) responded to the same questionnaire at a training camp in April 1995. Ten of the sailors responded in both 1994 and 1995. The questionnaire asked whether or not the sailors used a training race diary and inquired about their knowledge and use of sport science in the areas of nutrition, psychology and physical conditioning. In 1995, additional questions enquired about sailors' perception of sport science and its affect on their racing performance. In the intervening year, six of the sailors received sport science support (SSS) in nutrition; eleven received SSS in sport psychology; eleven received SSS in physical conditioning. In 1995, the sailors reported a greater amount of fluid taken and drunk on a four-hour sail and a greater proportion of sailors ate a high carbohydrate meal after a race. They also reported feeling less anxiety before a race. Increases were also observed in the sailors' volume and intensity of physical training and in their chosen type of aerobic training. Most sailors believed that their knowledge and use of sport science had increased and that this had led to improvements in racing performance. It is concluded that elite New Zealand sailors' reported use of sport science improved in the areas of physical conditioning and nutrition between 1994 and 1995. Improvement in the use of sports psychology was less clear and the eleven sailors who received psychology SSS reported feeling more anxious before a race. There was little evidence to suggest that the sport science programme was responsible for the improvements. This study indicates that sailors are beginning to understand the importance of sport science support, but there is still much room for improvement in their use of sport science. PMID- 10870244 TI - Total activation change of visual and motor area due to various disturbances. AB - Brain activation is affected by gradient acoustic noise and various disturbances as well as by other primary tasks. Therefore, we have studied the effects of various disturbances of two different levels of difficulty, that is, weak and strong difficulty levels for primary visual and motor tasks. In the case of visual task with motor and mental disturbances, we found it decreased as motor and mental disturbance difficulty-level increased, compared with the case of without motor and mental disturbances. To the contrary, in the case of motor activity, the total activation of motor cortex with weak and with strong mental disturbance increased as mental disturbance difficulty-levels increased. Therefore, one can conclude that when mental disturbance is added, the visual cortex and motor cortex have an opposite result and when the difficulty-level of the disturbance is increased, the primary tasks are affected more significantly. Although the current observation is preliminary and requires more careful experimental study, it appears that various disturbance effects on brain functions (such as motor and visual cortical responses) produce significant differences in data observations. PMID- 10870245 TI - Research on nitrogen-oxygen saturation diving with repetitive excursions. AB - For some tasks of underwater operation the need for longer dive duration and more working divers necessitates the use of saturation diving techniques with excursions. Saturation diving with excursion has high working efficiency. A collaborative experiment with Chinese Underwater Technology Institute, American National Office of Research Undersea Program and Hamilton Research Ltd. was conducted at our Institute in Shanghai. The main experiment objectives were to assess the longer, deeper repetitive excursions during nitrogen-oxygen saturation situation, oxygen exposure management, nitrox saturation decompression after excursions and performance aspects. Four Chinese professional experienced divers were saturated at 25 msw for 5 days at the hyperbaric facility, where they did 15 air excursions to depths between 50 and 75 msw, for duration up to 240 min. Decompression from excursions to the storage were mostly no-stops, but 5 required stops for 3 to 116 min. Saturation decompression began with the "precursory" ascent following a brief return to 25 msw. Doppler bubble detection showed some bubbles of Spencer Grade II and occasionally III, following excursions and during saturation decompression, especially after muscle flexing. No symptoms of decompression sickness were reported: one diver was more of fatigued on one occasion than other times. Oxygen exposure reached its peak of 3103 Oxygen Toxicity Units on Day 6. The only subjective symptom of oxygen toxicity was mild and transient numb fingertips. No significant change was seen in vital capacity. PMID- 10870246 TI - Effects of skin pressure applied by cuffs on resting salivary secretion. AB - The effects of pressure applied by cuffs to the abdomen, thighs and legs on resting salivary flow rate and digestive function of saliva were investigated in 9 healthy female students, aged 18 to 33 yrs (Experiment I) in a climatic chamber (Ta: 28 degrees C, RH: 50%). Each participant changed from street clothing into loose-fitting experimental garments so as to avoid any skin pressure on the body, and sat calmly in a reclining chair throughout the experimental period (195 min). After 90 min (FREE period), the physiological effects of skin pressure applied by their own clothing disappeared, and skin pressure was applied for the next 60 min to the abdomen (40 mmHg) and thighs (40 mmHg) then to the legs (60 mmHg) by the use of air-inflated cuffs (PRESSURE period). During the next 45 min, the skin pressure was again removed by letting the air of the cuffs out (FREE' period). The resting salivary flow rate was significantly suppressed while the skin pressure was applied by the cuffs. The digestive time for starch investigated in terms of the iodine starch reaction was longer with the skin pressure than without. The concentration of amylase measured in 20 female participants aged 21 to 23 yrs, decreased with skin pressure applied by the usage of the rubber tape (Experiment II). These results suggest that the pressure applied to the body can influence the digestive response by decreasing the amount of saliva via the autonomic nervous system. PMID- 10870247 TI - A significant Surgeon General's report. PMID- 10870248 TI - Bringing psychiatric nursing into the twenty-first century. AB - Psychiatric nursing stands at a crossroads, in danger of losing both its identity and standing within the larger body of nursing. Enrollment in graduate programs is at an all time low and many traditional employment opportunities for psychiatric nurses are disappearing. Many explanations have been proposed to account for the crisis in psychiatric nursing practice. Although many of the identified external forces have had real impact on psychiatric nursing, these factors have impacted all of nursing. Yet our Nurse Practitioner (NP) peers are thriving, with proliferation of NP programs with unprecedented enrollment for this nationally accepted and understood role. The psychiatric nursing crisis may be most directly related to the reality that we as a professional group have thus far failed to adequately respond to external realities that have dramatically altered the environment in which psychiatric nursing occurs. This article argues for reframing the discipline of psychiatric nursing, accomplished as a national consensus building process, and including 4 critical components: (1) reconceptualization of what constitutes the core of psychiatric nursing content and represents the epistemological heart of the profession; (2) identification of the critical clinical competencies that reflect the core content, represent the role and scope of psychiatric nursing, and that match current and anticipated practice realities; (3) identification and standardization of measurable clinical outcomes, predicated on both content and competencies, which will allow psychiatric nurses to measure, in meaningful ways, the impact of their practice on patients' health; and (4) establishment of a research agenda that will allow psychiatric nursing to expand its unique knowledge base. PMID- 10870249 TI - A consumer-oriented practice model for psychiatric mental health nursing. AB - Psychiatric mental health (PMH) nurses are systematically examining the concepts of health, wellness, and culture as they relate to recovery processes in mental health consumers. This article describes the development of a holistic, consumer oriented model, the Recovery and Monitoring Model, which integrates concepts involved in the recovery process and the manifestation and management of psychiatric symptoms. Information regarding a test of the model is presented using data from a cross-sectional study of 199 community-based, adult, public mental health consumers. The discussion focuses on issues surrounding the model's applicability for PMH nursing education, practice, and research. PMID- 10870250 TI - Psychiatric home care and family therapy: a window of opportunity for the psychiatric clinical nurse specialist. AB - This article discusses use of the Developmental-Interactional Model of family therapy by a Psychiatric Clinical Nurse Specialist (CNS) for selected patients receiving psychiatric home care services. This form of family therapy is an integrative approach to working with individuals, couples, and families that combines elements of structural-strategic family therapy with life cycle and intergenerational approaches. Applied to patients and families in a home care setting, this model permits the CNS to assess relational dynamics over time, determining how these transitions relate to a family's problem-solving capability. Case studies are provided to show the application of this model for desired outcomes. PMID- 10870251 TI - Alienation from self and others: the psychosocial problem of rural alcoholic women. AB - The purpose of this study was to describe women's perspectives in becoming and being alcohol dependent. Using grounded theory techniques, 14 adult Black and White women receiving treatment for alcohol addiction at rural substance abuse centers participated in an intensive interview. Data analysis focused on the identification of the basic psychosocial problem and the process of becoming alcohol dependent. The results are presented in 2 parts. Part 1, "Alienation From Self and Others," focuses on the basic psychosocial problem faced by women in becoming alcohol dependent. Part 2, "Running Away to Nowhere," on page 142 of this issue of Archives, describes the process used by women to resolve this problem. PMID- 10870252 TI - Running away to nowhere: rural women's experiences of becoming alcohol dependent. AB - The purpose of this study was to describe women's perspectives in becoming and being alcohol dependent. Part 1, "Alienation From Self and Others," on page 134 in this issue of Archives, describes the study methods, the sample, and the basic psychosocial problem faced by rural, alcoholic women. Part 2, "Running Away to Nowhere," focuses on the basic psychosocial process that women used to resolve the pain caused by their "Alienation From Self and Others." The article concludes with suggestions for nursing intervention. PMID- 10870253 TI - Codependency and related health variables. AB - Codependency is a controversial concept especially for feminist scholars who are concerned about pathologizing traditional female roles. This study's purpose was to determine: (1) the prevalence of codependency in a sample of older women who because of age may ascribe to traditional roles; (2) how the Hughes Hammer/Martsolf theoretical model of codependency relates to other health variables; and (3) whether previous findings about the relationship between codependency and depression replicate. Survey design was used with a sample of 238 women (ages 65 to 91) attending a flu shot clinic. Subjects completed the Codependency Assessment Tool, Beck Depression Inventory, Quality of Life Scale, Perceived Health Report, Measurement of Patient Functional Abilities, and Illness Prevention Screening Behaviors Checklist. Of these women, 99% had low codependency scores. Statistically significant correlations existed between codependency and perceived health (p < .01), and functional ability (p < .01). Codependency was not significantly correlated with illness prevention behaviors and quality of life. Codependency and depression, as in previous studies, were significantly correlated (r = .446, p = .0001). Using analysis of variance, 3 codependency subscales had significant positive effect on depression: Low Self Worth, Medical Problems, and Hiding Self. Further studies should examine the degree of ascribing to traditional female roles in women dealing with codependency issues. PMID- 10870254 TI - Plasma-levels of endothelin-1 and angiotensin-II and reactivity of arterial blood pressure to exogenous sympathomimetics and vasoactive peptides in rat model of malignant renal hypertension. AB - BACKGROUND: There is still considerable uncertainty regarding sensitivity of arterial blood pressure to endogenous peptides in renal hypertension. Many pathological processes including hypertension have been shown to be associated with release of endothelin-1 (ET-1). However the role of ET-1 in regulation of arterial blood pressure in hypertension is still controversial. OBJECTIVES: The role of endothelin-1 (ET-1) and angiotensin-II (AT-II) in malignant phase of renovascular hypertension has been assessed on the basis of arterial blood pressure increase and ETA receptor density measurements in Glodblatt-hypertensive rats (RVH). RESULTS: The arterial blood pressure response to sympatomimetic amines, vasopressors, the plasma ET-1 and AT-II levels as well as renal subtype ETA receptor density were significantly increased in RVH rats with malignant hypertension. The dominance of vasopressor ETA receptors in RVH rats suggest the contribution of endothelin peptides to malignant renovascular hypertension. (Tab. 1, Fig. 7, Ref. 25.) PMID- 10870255 TI - [Indirect diagnosis of type I neurofibromatosis using RFLP in Slovak families]. AB - Neurofibromatosis type I clinical diagnosis confirmation as well as antenatal diagnostics of the disease are recently provided by molecular genetics. The authors analyze 17 Slovak families with multiple NFI incidence, in whom the detection of mutated gene transfer was performed using indirect diagnostics-bound with of restrictive fragments length polymorphism RFLP. With the help of PCR 7 polymorphic sequencies were amplified and subsequently broken with restrictive endonucleases localized close to the neurofibrin gene. The system informative capacity was comparable with the results of other Caucasian population studies. Although direct detection of mutation is the perspective of the diagnostics, binding analysis in informative families with multiple incidence of the disease provides reliable and cheaper possibility of NFI diagnostic on the level of DNA analysis. (Tab. 1, Fig. 3, Ref. 26.) PMID- 10870256 TI - [Comparison of the results of HLA typing using serologic and molecular genetics methods]. AB - A comparison of the HLA class I typing in 50 unrelated individuals by means of serological and molecular genetic (PCR-SSP) methods was carried out. DNA-typing is more fast and reliable method in comparison with serology. It is necessary to introduce molecular genetic methods for the detection of HLA class I alleles. On the other hand there are alleles, which are not expressed on cell surface. In our laboratory both methods are established and the results of both were compared. It may be useful for determining the selection strategy of HLA-identical donor recipient pair suitable for bone marrow transplantation. The results demonstrated 9% misassignments of HLA-A antigens by serology, 11% of HLA-B and 39% of HLA-C. The serological discrepancies found were of three categories: false negatives, false positives, and an incomplete typing. The vast majority of the discrepancies were due to a combination of relatively low expression of HLA antigens, lack of serological reagents and misclassification of antigens within cross-reactive groups. These results indicate that nowadays the serological typing is insufficient for clinical histocompatibility testing. (Tab. 3, Ref. 16.) PMID- 10870257 TI - [The role of adhesion molecules in the immune system]. AB - Adhesion molecules play a major role in many biological functions. They are crucial in the development of embryo into formed organism; later they mediate many physiological and pathological functions. From the immunological point of view they are involved in virtually every process of cell interactions, involving thymic selection and antigen priming, antigen recognition and cell activation, cytotoxicity and lymphocyte recirculation. This review focuses mainly on the role of adhesion molecules in close contact between the cells, crucial for the inflammatory and immune responses. (Tab. 3, Fig. 1, Ref. 29.) PMID- 10870258 TI - [Adhesion molecules as the strategic goal of immunotherapy]. AB - Adhesion molecules are cell surface glycoproteins involved in cell-cell and cell matrix interactions. They play a central role in leukocyte adhesion to endothelium, transmigration, binding to target cells and cytotoxicity. Many pathological processes underlie abnormal cell-cell interactions in the organism. The adhesion molecules are critical for the communication mechanism between cells in inflammatory and immunologic response. The control of cell-cell adhesive interactions, i.e. modulation of adhesion molecules, is an important strategy for anti-inflammatory therapy. (Ref. 65.) PMID- 10870259 TI - [Effect of vitamin C and E on nonenzymatic glycation and physico- chemical properties of isolated erythrocyte membranes in diabetic patients]. AB - Non-enzymatic glycation, accompanied by the formation of free radicals, represents a serious problem in diabetes mellitus. It is supposed to be the cause of the development of long-term diabetic complications. The aim of this work was to estimate the effect of treatment with vitamin C (1 g per day) and E (600 mg per day) on selected biochemical parameters as well as to determine the physicochemical state of erythrocyte membranes in diabetics. The paper also compares the physicochemical state of diabetic and control erythrocyte membranes. The changes in the values of glycaemia, glycated haemoglobin, and fructosamine were insignificant after three months of treatment. This points out that the doses used could be low or that the patient compliance was poor. An anionic fluorescent probe merocyanine 540 (MC540) was used to monitor possible changes in the physicochemical properties of isolated diabetic erythrocyte membranes. Significantly higher affinity of MC540 monomers to the membrane in diabetics treated with vitamin E was observed, which can be the result of the antioxidative effect of the vitamin (p < 0.02). A comparison of absorption spectra of MC540 in diabetic and control membranes revealed significant changes in the position of the bands and in their absorbances (p < 0.01 and less). They result from substantial alterations in the structure, surface charge, and the fluidity of erythrocyte membranes in diabetes mellitus. (Tab. 2, Fig. 3, Ref. 22.) PMID- 10870260 TI - [New findings on the ecology and epidemiology of murine herpes virus isolated in Slovakia]. AB - Rodents are important reservoir animals of many human microbial pathogens. Of small rodents, trapped on Slovakia territory, were several strains of murine herpes virus (MHV) isolated. Our purpose was to complete the existing knowledge about circulation of MHV in rodents and to find out, whether also other animal species including man are MHV sensitive or not. The presence of antibodies against MHV in serum of the tested animals and men was followed by virus neutralization test (VNT) and ELISA. Pathological changes in differential white blood cell count of the trapped rodents, were also observed because it is known, that MHV induces them in laboratory mice. A total of 627 small terrestrial mammals of nine species were collected in four localities of western and eastern Slovakia during 1984-1988. Neutralizing antibodies to MHV were detected in five species of rodents in 130 cases (20.7%). Antibodies were most frequently detected in Apodemus flavicollis (34.9%). Pathological changes in differential white blood cell count of trapped rodents were detected in 37% (34/92). Neutralizing antibodies were found also in serum of fallow deers (Dama dama), wild boars (Sus scrofa), deers (Cervus elaphus) and sheep but not in serum of pheasants (Phasianus colchicus) and muflons (Ovis musimon). ELISA and VNT tests were used to investigate 20 serums of employees of the Institute of Virology, Slovak Academy of Sciences and Faculty of Natural Sciences of Comenius University. There were eight samples positive (40%). The titers of antibodies were 4-32 in VNT and 1000 in ELISA. PMID- 10870261 TI - [Epidemiologic and microbiological aspects of mycobacteriosis in Slovakia- Mycobacterium szulgai]. AB - The first case of mycobacteriosis caused by M. szulgai in the territory of Czechoslovakia was discovered in the year 1979 in southern Slovakia and was published in our and foreign literature in the year 1981. The purpose of this investigation is to describe the epidemiological situation of the diseases caused by M. szulgai in Slovakia and to compared it with the experiences in chosen developed countries, especially focused on the localization of the disease, factors of transmission, mechanism of transmission and other epidemiological characteristics. The methodology of this paper is based upon surveillance of tuberculosis applied on mycobacterioses. During the period of last 20 years two cases of the disease without mutual epidemiological connection were discovered. In contradiction to other mycobacterioses, diseases caused by M. szulgai, have not the tendency to endemic occurrence. The most often transmission factors are contaminated water and soil. Both cases of M. szulgai in Slovakia suffered from pulmonary diseases. Other localization reported in other countries, for example: olecranon bursitis, skin infections, cervical adenitis, osteomyelitis and renal disease were not reported in our country up to now. (Ref. 23.) PMID- 10870262 TI - [Lyme borreliosis: open questions and problems at the turn of the millennium]. AB - At the beginning of new millennium, Lyme borreliosis is still the subject of intensive research, polemic discussions and open questions. The authors present minute analysis of the issues associated with Lyme borreliosis, concentrating on biological aspects and taxonomic classification of the agent, co-transmission and co-infection, diagnostic criteria and their validity, laboratory diagnostics an therapy of the disease including perspectives of active immunisation in the future. Special attention is paid to open questions in clinical and laboratory diagnostics of the disease and to the prospects for the near future. The authors also discuss the importance of international and interdisciplinary cooperation in the process of articulation of diagnostic criteria and recommended procedures for laboratory diagnostics. (Ref. 17.) PMID- 10870263 TI - [Determinants of the effectiveness of electric stimulation of the auditory nerve with cochlear implants: II. Configuration of the stimulating electrodes]. AB - The influence of the electrode configuration on the efficacy of the electrical stimulation were investigated in the auditory nerve. For this purpose normal hearing cats were deafened by intrascalar application of neomycin and stimulated by a NUCLEUS-22 implant. Stimulation was performed in monopolar, bipolar and tripolar mode with all electrodes of the implant. The smallest thresholds were determined for monopolar stimulation, followed by tripolar and bipolar stimulation. The most focused stimulation was achieved by tripolar stimulation, followed by bipolar and monopolar stimulation. Tripolar stimulation is therefore suitable for beam forming in electrical stimulation of the auditory nerve. PMID- 10870264 TI - [Selection of donor-recipient pairs in families for allogenic bone marrow transplantation]. AB - In the years 1985-1999 452 were families investigated with the aim to find up an HLA-identical sibling for a patient suffering from a disease, the treatment of which requires bone marrow transplantation. Total number of investigated siblings (including patients) was 1334. HLA typing was done by serological methods, mixed lymphocyte culture test (MLC), and in the last three years also by DNA-typing (PCR-SSP). 188 HLA identical donor-recipient sib-pairs were found. At the same time the number shows that a potential donor could be found in 41.5% of investigated families. PMID- 10870265 TI - The phosphoinositide signaling pathway in the carotid body mechanism. PMID- 10870266 TI - Interpretation of clinical laboratory results. PMID- 10870267 TI - Metabolic X-syndrome, atherogenic risk factors of premature coronary heart disease. A retrospective study in a Hungarian patient-group. PMID- 10870268 TI - Metabolic syndrome X and fibrinolytic disturbances. PMID- 10870269 TI - Assessment of insertion/deletion polymorphism of angiotensin-converting enzyme gene and its clinical importance. PMID- 10870270 TI - [Epidemiology of osteoporosis]. PMID- 10870271 TI - [Early diagnosis of postmenopausal osteoporosis]. AB - Osteoporosis can be diagnosed by history, physical examination, laboratory test and bone mass measurement. A low bone mass is the most important risk factor for osteoporotic fractures. In the assessment of risk is very important the rate of bone loss too. Combining bone density and bone turnover with age and other associated risk factors can improve the assessment of the risk of osteoporosis especially at the postmenopausal women. As in many medical conditions early diagnosis is the key to success in prevention and treatment in osteoporosis. PMID- 10870272 TI - [Hormone replacement therapy and osteoporosis]. AB - Estrogen therapy has clearly been shown to maintain bone mass and to reduce postmenopausal bone loss, thereby lowering the risk of osteoporotic fractures. Current estrogen use is better than past use for maintenance of bone density and current estrogen therapy begun 10 or more years after the menopause might be as beneficial as continuous treatment begun at the time of menopause. PMID- 10870273 TI - [Prevention and non-hormone therapy of osteoporosis]. AB - Osteoporosis is an important age-related disease, constituting a major risk of fracture in the elderly, and in postmenopausal women in particular. The main aim of any therapy for osteoporosis is to prevent fractures. Primary prevention attempts to increase peak bone mass by diet and exercise. Secondary prevention aims to prevent bone loss after the menopause by the use of drugs (HRT, calcitonin, bisphosphonates, etc) and is more efficient strategy. PMID- 10870274 TI - Is need enough? PMID- 10870275 TI - Careers of academic dentists. PMID- 10870276 TI - Careers of academic dentists. PMID- 10870277 TI - Careers of academic dentists. PMID- 10870278 TI - Registration: stage I--creating and outlining the form of the upper denture. PMID- 10870279 TI - Maintaining good dental practice: the East Lancashire approach to dentists whose performance gives cause for concern. AB - The Consultant in Dental Public Health and the Local Dental Committee in East Lancashire have developed a local system, similar to that being set up for general medical practitioners, to help general dental practitioners whose performance is causing concern. GDPs approved it at an open meeting of practitioners. A Dental Performance and Assessment Group has been set up and one problem has already been resolved to everyone's satisfaction. The model is recommended to dentists in other areas. PMID- 10870280 TI - Pleomorphic salivary adenoma in an adolescent. AB - This case study illustrates an unusual condition in childhood, pleomorphic salivary adenoma in a 12-year-old. The management, pathology and treatment are discussed with emphasis on the practitioner to be vigilant for rare conditions. A review of the literature on childhood salivary tumours illustrates the rare nature of this pathology. PMID- 10870281 TI - A study of therapeutic antibiotic prescribing in National Health Service general dental practice in England. AB - OBJECTIVE: To study the therapeutic prescribing of antibiotics by general dental practitioners. DESIGN: A postal questionnaire of National Health Service general dental practitioners in ten English Health Authorities. SUBJECTS: General dental practitioners (1,544) contracted to provide NHS treatment in the Health Authorities of Liverpool, Wirral, Oxfordshire, Buckinghamshire, Nottingham, North Nottinghamshire, Sheffield, Newcastle, Northumberland and North Tyneside. MAIN OUTCOME MEASURES: The questionnaires were analysed and the responses to each question expressed as absolute frequencies. RESULTS: Responses to the questionnaire were received from 929 (60.1%) practitioners. More than 95% of practitioners recognised the need for prescribing antibiotics where there was evidence of spreading infection. Some practitioners (12.5%) prescribed antibiotics for acute pulpitis and (3.3%) for chronic marginal gingivitis. Antibiotics were prescribed by practitioners before drainage of acute abscesses (69%) and by 23% after drainage. Practitioners were generally not influenced by patient's expectations of receiving antibiotics (92%), but would prescribe when under pressure of time (30.3%), if they were unable to make a definitive diagnosis (47.3%), or if treatment had to be delayed (72.5%). Amoxicillin was the most frequently prescribed antibiotic used for most clinical conditions apart from pericoronitis, acute ulcerative gingivitis and dry sockets where metronidazole was the drug of choice. There was a wide variety of dosage, frequency and duration for all the antibiotics used in the treatment of acute dental infections. CONCLUSIONS: The results obtained from this questionnaire support the conclusion that the therapeutic prescribing of antibiotics in general dental practice varies widely and is suboptimal. There is a clear need for the development of prescribing guidelines and educational initiatives to encourage the rational and appropriate use of the antibiotics in National Health Service general dental practice. PMID- 10870282 TI - Socioeconomic and geographical influences on primary dental care preferences in a population of young children. AB - OBJECTIVE: To compare the socioeconomic profiles of children registered in the GDS, with those using the CDS services and unregistered children. Secondly to examine the effects of socioeconomic status on travelling to access primary dental care, and finally to map out the location of unregistered children in relation to primary dental care services. SETTING: The study was carried out in 1998 in Ellesmere Port in the North West of England. SUBJECTS AND MATERIALS: The study population was all children younger than 6 years who used primary dental care services in, or were residents of, Ellesmere Port. The study population was categorized into those registered with a GDS dentist, those using CDS services and those unregistered by matching GDS and CDS data to the HA population register. Socioeconomic status was measured using the Super Profiles geodemographic classification. The relationship between service preferences and travelling to access primary dental care with socioeconomic status were compared using cross-tabulations and chi square tests. RESULTS: There was a significant socioeconomic trend evident in the use of dental services. Two thirds of those using CDS services came from the most deprived area types. Of those who were unregistered half lived in the most deprived area types compared with one third of those registered with the GDS. Those who travelled into Ellesmere Port to access primary dental care were significantly more likely to live in an affluent area. Unregistered patients were homogeneously spread across the town. CONCLUSIONS: The ability to match GDS and CDS data to the HA population register is essential to understand how dental services are used by the local population. Children from deprived areas are more likely to use the CDS and a service local to their homes, therefore primary dental care services for deprived communities have to be provided locally. PMID- 10870283 TI - An evaluation of general professional training for dentistry in Scotland. AB - The first General Professional Training (GPT) scheme in Scotland ran from August 1996 to July 1998 with sixteen trainees completing one year in general practice and two six-month blocks in the hospital and community services. An independent evaluation of the scheme focussed on the advantages and disadvantages of GPT and the level of support for the scheme as well as providing stakeholders with an opportunity to contribute to its development. This paper reports the findings of the evaluation. PMID- 10870284 TI - Dentistry 2000--two sides of the same coin. PMID- 10870285 TI - Routine immunisation for influenza to include those aged 65 years and over and health care workers. PMID- 10870286 TI - Microbiological investigation into wound botulism. PMID- 10870287 TI - AIDS and HIV infection in the United Kingdom: monthly report. PMID- 10870288 TI - Women's and men's personal goals during the transition to parenthood. AB - To investigate how women's and men's personal goals change during the transition to parenthood, the authors studied 348 women (152 primiparous and 196 multiparous) and 277 of their partners at 3 times: early in pregnancy, 1 month before the birth, and 3 months afterward. At each measurement, participants completed the Personal Project Analysis questionnaire (B. R. Little, 1983). The results showed that during pregnancy women became more interested in goals related to childbirth, the child's health, and motherhood and less interested in achievement-related goals. After the birth women were more interested in family- and health-related issues. These changes were more substantial among the primiparous than among the multiparous mothers. Although the men's personal goals changed during the transition to parenthood, these changes were less substantial than those found among the women. PMID- 10870289 TI - Parents' attitudes about children: associations with parental life histories and child-rearing quality. AB - This study examined relations among parents' perceptions of their childhood, attitudes about life, expectations for child behavior, attitudes about their child's behavior, and the child-rearing environment parents provide. Eighty mothers of 1- to 5-year-olds were interviewed about perceptions of receiving harsh parenting as children, current attitudes about life, developmental expectations, and views of intentionality and severity of their child's misbehavior. The home environment was measured using the Home Observation for Measurement of the Environment (R. H. Bradley & B. Caldwell, 1979) scale. Mothers who reported harsh parenting as children, negative attitudes about life, and unrealistic developmental expectations had negative attitudes about their own child. These attitudes were related to provision of lower quality home environments. Results support a constructivist approach to understanding parental social cognitions and behavior. PMID- 10870290 TI - Factors associated with fathers' caregiving activities and sensitivity with young children. NICHD Early Child Care Research Network. AB - A multifactorial model was used to identify child, sociodemographic, paternal, and maternal characteristics associated with 2 aspects of fathers' parenting. Fathers were interviewed about their caregiving responsibilities at 6, 15, 24, and 36 months, and a subset was videotaped during father-child play at 6 and 36 months. Caregiving activities and sensitivity during play interactions were predicted by different factors. Fathers were more involved in caregiving when fathers worked fewer hours and mothers worked more hours, when fathers and mothers were younger, when fathers had more positive personalities, when mothers reported greater marital intimacy, and when children were boys. Fathers who had less traditional child-rearing beliefs, were older, and reported more marital intimacy were more sensitive during play. These findings are consistent with a multifactorial and multidimensional view of fathering. PMID- 10870291 TI - Parental reports of coparenting and observed coparenting behavior during the toddler period. AB - Fifty-two married partners played with their 30-month-olds in both dyadic (parent child) and whole family contexts and reported on their own coparenting activities (family integrity-promoting behavior, conflict, disparagement, and reprimand). Coparenting behavior observed in the whole family context was evaluated for antagonism, warmth and cooperation, child-adult centeredness, balance of positive involvement, and management of toddler behavior. Parallel balance and management scores were also formed using dyadic session data. Men's reported family integrity-promoting activities and women's reported conflict and reprimand activities were reliable correlates of family group process in both bivariate and discriminant analyses, with links enduring even after controlling for marital quality. Whole family- and dyad-based estimates of coparenting were altogether unrelated, and reported coparenting was tied only to behavior in family context, not to family measures created from dyad-based data. PMID- 10870292 TI - Rule compliance and peer sociability: a study of family process, school-focused parent-child interactions, and children's classroom behavior. AB - This study examined the associations among family processes (cohesion, control, and conflict), school-focused parent-child interactions (support and pressure about achievement), and the child's own characteristics (assertiveness, frustration tolerance, intellectual effectiveness, and self-esteem) as correlates of rule compliance and peer sociability in the classroom. The sample consisted of 161 Grade 4 and 151 Grade 7 children. Family processes and parent-child interactions about school issues were associated with children's personal characteristics, which, in turn, predicted children's rule compliance and peer sociability. Some differences were found between the 4th- and 7th-grade samples; however, many variables consistently predicted the same outcomes across grades. PMID- 10870293 TI - Quality of couples' relationship and adjustment to metastatic breast cancer. AB - This study examined mood disturbance among women with metastatic breast cancer in relationship to partnership status, relationship quality, and partner's coping and mood disturbance. These associations were examined within a total sample of 125 metastatic breast cancer patients and a subsample of 48 of these patients and their partners. Partnered and single women were indistinguishable in mood disturbance when household income was statistically controlled. Results also showed that patients were less distressed when they rated the relationship higher in Cohesion--Expression and in Conflict and when their partners reported lower mood disturbance. One possible implication of these results is that in relationships in which a woman has metastatic cancer, she may benefit from open engagement of difficulties and conflict. Furthermore, alleviating her distress may be better achieved by focus on the couple relationship rather than her individual coping. PMID- 10870294 TI - The longitudinal association between attributions and marital satisfaction: direction of effects and role of efficacy expectations. AB - This study investigated the direction of possible causal effects between attributions for negative partner behavior and marital satisfaction and tested whether any effects are mediated by efficacy expectations regarding marital conflict. Couples married for 15-20 months completed measures of attribution and satisfaction at Time 1 and at Time 3 (18 months later). At Time 2 (6 months after Time 1) they completed a measure of efficacy expectations. For both husbands and wives, a cross-lagged effects model showed that the paths from causal attributions to later satisfaction and from satisfaction to later causal attributions were significant. Efficacy expectations mediated the temporal relation between attributions and satisfaction. These findings support the assumption that there is a reciprocal causal influence between attributions and satisfaction but suggest important modifications to models of close relationships and marital therapy. PMID- 10870295 TI - Interaction structure of husbands' and wives' disclosure of marital conflict to their respective best friend. AB - Husbands' and wives' conversations with their respective best friend (N = 88) were coded to assess spouses' and friends' mutual influence in regulating support and interference with regard to spouses' marriage and to assess the impact of spouses' sex and marital satisfaction on the conversation processes. Dissatisfied husbands and wives expressed fewer positive and more negative views of marriage than satisfied husbands and wives and the friends in the 2 groups. There were no group and no sex differences in interference sequences. There were group and sex differences in support sequences. Friends of satisfied wives and those of dissatisfied husbands were more likely than satisfied wives and dissatisfied husbands to get support for their positive views of marriage. The findings are discussed with reference to the specific effects of outsiders' support and interference on satisfied and dissatisfied spouses. PMID- 10870296 TI - Effects of parental separation and divorce on very young children. AB - Data from the National Institute of Child Health and Human Development Study of Early Child Care were analyzed to explore effects of marital separation on children in the first 3 years of life. The sample included 73 never-married mothers and 97 separated mothers; a comparison group of 170 was conditionally randomly selected from the 2-parent families. Children in 2-parent families performed better than children in 1-parent families on assessments of cognitive and social abilities, problem behavior, attachment security, and behavior with mother. However, controlling for mothers' education and family income reduced these differences, and associations with separated-intact marital status were nonsignificant (the effect size was .01). Thus, children's psychological development was not affected by parental separation per se; it was related to mothers' income, education, ethnicity, child-rearing beliefs, depressive symptoms, and behavior. PMID- 10870297 TI - Bartter's syndrome in pregnancy: a case report and review. AB - Bartter's syndrome is a rare renal tubular disorder, involving juxtaglomerular cells hyperplasia, characterized by normotensive hyper-reninism and secondary hyperaldosteronism, marked renal loss of potassium and profound hypokalaemia. Both clinical and biochemical features are heterogeneous, ranging from the incidental finding in an asymptomatic patient to marked clinical features of hypokalaemia. Inheritance is likely to be an autosomal recessive. We present a case of Bartter's syndrome complicating pregnancy in a Chinese woman. We documented an increasing demand for potassium supplement during pregnancy which stabilized by mid-trimester. The absence of pregnancy complications such as polyhydramnios indicated that the fetus was unlikely to be affected by the condition. PMID- 10870298 TI - Changes in the serum levels of human chorionic gonadotropin and the pulsatility index of uterine arteries during conservative management of retained adherent placenta. AB - OBJECTIVE: Our purpose was to assess the natural course of retained adherent placenta at term. METHODS: Five cases of retained adherent placenta, clinically diagnosed as placenta accreta, were managed conservatively without methotrexate. To assess the biochemical and circulatory changes in the placentas, the serum levels of human chorionic gonadotropin (hCG) and the pulsatility index (PI) of the uterine arteries were examined. RESULTS: Serum hCG levels decreased spontaneously; the half-life of serum hCG was calculated to be 5.2 +/- 0.26 days (mean +/- SEM). The PI of the uterine arteries remained at the level of pregnant women at term, but became elevated within a few days after the removal of the placentas. All the placentas were successfully removed transvaginally within 6 weeks postpartum. CONCLUSIONS: The changes in serum hCG observed in this study indicated the spontaneous degeneration of the placenta. Such changes might be similar to those reported to occur during treatment with methotrexate. In contrast, the PI of the uterine arteries did not reflect degeneration of the placenta. PMID- 10870299 TI - A retrospective survey of clinical, pathologic, and prognostic features of adnexal masses operated on during pregnancy. AB - OBJECTIVES: To evaluate retrospective data concerning patients with adnexal masses that were managed surgically during pregnancy and their effect on fetal outcome. METHODS: Data were reviewed concerning pregnant women who required surgery at our hospital between 1980 and 1997 for an adnexal mass. RESULTS: In the past 19 years at our hospital a total of 69 Japanese women aged 28.5 +/- 3.4 years (including 2 women with twin pregnancies) were diagnosed with adnexal masses that required surgery. The masses (10.2 +/- 4.5 cm in the largest diameter) were removed at 13.9 +/- 3.7 weeks of gestation. The pathologic features of the 69 lesions were as follows: 33 mature cystic teratomas, 13 functional cysts, 8 mucinous cystadenomas, 6 endometriotic cysts, 4 paraovarian cysts, 3 serous cystadenomas, and 2 malignant neoplasms. Of the 60 patients for whom the outcome of pregnancy was available, 7 (12%) gave birth before 37 weeks of gestation, while 2 (3.3%) experienced spontaneous abortions. There were 3 perinatal deaths among the 60 infants. Two of these 3 infants died due to major anomalies. CONCLUSIONS: Although larger studies are required for confirmation, our results suggest that an adnexal mass might be associated with an adverse fetal outcome. Surgical intervention at < 24 weeks of gestation per se might not have been related to the adverse outcomes. We emphasize that surgical intervention during pregnancy can be avoided in patients who have ultrasonographically pathognomonic features of benign cystic teratomas, which are the most common neoplasms operated on during pregnancy. PMID- 10870300 TI - Relationship between endometrial estrogen and progesterone receptors, and sonographic endometrial appearance in the preovulatory phase. AB - OBJECTIVES: The purpose of the present study is to evaluate the relationship between endometrial concentrations of estrogen receptor (ER) and progesterone receptor (PR), and sonographic endometrial findings in the preovulatory phase of menstrual cycle. STUDY DESIGN: In 45 cycles of 45 infertile women with tubal factor only, transvaginal sonographic assessments and biopsy for immunohistochemical staining of the endometrium were made in the preovulatory phase of unstimulated, normal menstrual cycle. Immunohistochemical localization of ER and PR was scored according to intensity of staining and proportion of cells specifically stained in glandular epithelium and stroma, and the results were analysed according to the sonographic endometrial thickness (< 6 mm, 6-10 mm, or > 10 mm) and patterns. Endometrial patterns were classified as A, centrally hyperechogenic triple-line pattern or non-A, not triple-line. RESULTS: There were no significant differences in the endometrial thickness, serum estradiol level and serum progesterone level between A and non-A groups. The receptor scores of epithelial and stromal ER and epithelial PR were comparable in A and non-A groups. However, the receptor score of stromal PR was significantly higher in A group, with 4.8 +/- 1.4 compared with 2.7 +/- 1.7 in non-A group (p < 0.001). There were no differences in the receptor scores of epithelial ER, epithelial PR, stromal ER and stromal PR among the 3 groups according to the endometrial thickness. CONCLUSIONS: This study suggests that high PR expression in endometrial stroma could be related to the sonographic triple-line or multilayered pattern of endometrium in the preovulatory period. PMID- 10870301 TI - Anemia as a risk factor of hemorrhagic tendency during surgery. AB - OBJECTIVES: Although it is well-known that a hemorrhagic tendency in patients with massive bleeding during surgery, or with anemia before surgery, is difficult to control, its mechanism is still obscure. STUDY DESIGN: Using a damped oscillation rheometer, we measured the initiation time (Ti) of whole blood with several concentrations of red blood cells (RBCs) and the Ti of platelet-free plasma (PFP) with various concentrations of RBCs. RESULTS: The Ti of the whole blood became prolonged according to the dilution of the RBC concentration. RBCs could coagulate the PFP, and the Ti of PFP also was prolonged according to the dilution of the RBC concentration. CONCLUSION: These results suggest that RBCs might have an important role as a phospholipid supplier in the intrinsic pathway of the blood-coagulation mechanism. Anemia could thus represent a risk factor for hemorrhagic tendency during surgery due to lower RBC concentrations. PMID- 10870302 TI - Antepartum evaluation of monochorionic diamniotic (MD) twins; MD-twin score: a new scoring method for perinatal outcome. AB - OBJECTIVE: Our purpose was to establish a new scoring method to survey monochorionic diamniotic (MD) twins during antepartum periods. STUDY DESIGN: A retrospective study was performed regarding MD twins delivered between January 1992 and July 1996. Maternal and neonatal records were assessed for the following 5 perinatal variables; birth-weight discordance, amniotic-fluid discordance, hydrops fetalis, umbilical-cord insertion, and fetal-heart-rate monitoring. Each variable was coded as normal or abnormal and then assigned an arbitrary weight of 0 if normal and 1 if abnormal, yielding a range of scores from 0 (all normal) to 5 (all abnormal). The relationships between individual variables and their combinations and the outcome of pregnancy was determined. A poor pregnancy outcome consisted of intrauterine death, neonatal death, or neurological sequelae of at least one twin. The 5-variable combination was termed as the MD-twin score. A chi-square test and logistic regression analysis were used to determine statistical significance. RESULTS: There were 59 MD pregnancies, of which 13 pregnancies resulted in a poor outcome. The single variable that most likely contributed to a poor outcome was amniotic-fluid discordance. All 35 pregnancies with an MD-twin score of < or = 2 had a good outcome. There were 14 pregnancies with a score of 3, and 21% of them had a poor outcome. All of the pregnancies with a score of > or = 4 had a poor outcome. When we chose the MD-twin score of 3 as the critical point for a poor outcome, the likelihood ratio statistics became the highest of any single variable or any combination of variables. CONCLUSION: The MD-twin score predicted poor outcomes better than did any single variable or combination of variables. PMID- 10870303 TI - Spinal cord compression: a rareness in pregnant thalassemic woman. AB - Thalassemia is a common hematological disease in Southeast Asia. Extramedullary hematopoiesis is common sequelae in thalassemic patients but extramedullary hematopoiesis in the spinal epidural space that leads to paraparesis in pregnancy is very rare. We managed a thalassemic patient with extramedullary hematopoiesis and spinal cord compression during pregnancy. The diagnosis was made on clinical features and magnetic resonance imaging (MRI) showing a paravertebral mass infiltrating the epidural space. She was treated successfully with repeated blood transfusions until delivery. Fetal growth restriction was found but otherwise the fetus was clinically normal. She had an uneventful recovery when she was seen 6 weeks after delivery. PMID- 10870304 TI - Vaginal misoprostol in termination of second trimester pregnancy. AB - OBJECTIVE: To study the effectiveness and complications of 600 micrograms of intravaginal misoprostol for terminating second trimester pregnancies. STUDY DESIGN: One hundred and seventy-two patients undergoing termination of pregnancy between March 1997 and April 1999 were studied. Each patient received 600 micrograms of intravaginal misoprostol every 12 hours until abortion occurred. RESULTS: The mean induction to abortion time was 24.1 +/- 21.6 hours. The percentage of women aborting within 24 and 48 hours was 68.6 and 89.5 respectively. There was no significant difference in the mean induction to abortion time and the percentage of women aborted within 48 hours between nulliparous and multiparous women. The mean amount of misoprostol used was 1405.5 +/- 1084.6 micrograms. Incomplete abortion occurred in 23.3% of women. The most common complication was temperature of more than 38 degrees C occurred in 41% followed by diarrhoea (20%), nausea and vomiting (15%). CONCLUSION: Six hundred micrograms of vaginal misoprostol is effective, but whether the 48 hours abortion rate can be improved with a large dose or shortened the time interval between doses, requires further study. PMID- 10870305 TI - Profile of maternal smokers and their pregnancy outcomes in south western Sydney. AB - OBJECTIVE: The prevalence of maternal smoking remains high in New South Wales. In order to better understand the profile of maternal smokers, a study has been conducted to examine the social and demographic characteristics and pregnancy outcomes of women who smoked during their pregnancy in south western Sydney. METHODS: Women and babies of a retrospective cohort of 7,191 singleton births between March 1996 and December 1998, at Liverpool Hospital were analyzed. RESULTS: The prevalence of maternal smoking for the study population was 18.8%. The study found that the sociodemographic factors, such as marital status, ethnic origin, English speaking background, working status during pregnancy, and private health insurance status were independent risk factors for maternal smoking, but maternal age was not. Women who smoked during their pregnancy had higher rates of abruptio placenta, threatened premature labour, and premature labour. The adverse neonatal outcomes due to maternal smoking were low birth weight and increased neonatal morbidity. Smoking appears to have a protective effect on pregnancy induced hypertension. CONCLUSION: The findings of this study suggests that future smoking cessation programs should pay more attention to addressing sociodemographic and cultural factors that influence the behaviour of maternal smokers. PMID- 10870306 TI - A case of stage-IVb cervical adenocarcinoma successfully treated by combination chemotherapy: case report. AB - A 54-year-old patient with a Stage IVb adenocarcinoma of the cervix was treated with combination chemotherapy. This regimen consisted of intravenous cisplatin (70 mg/m2) and aclacinomycin A (30 mg/m2) on Day 1, followed by mitomycin C (5 mg/m2) on Day 2 and 3. A pathologically complete response was achieved by this regimen. The patient is well and has been free of symptoms for 66 months. PMID- 10870307 TI - A survey of the current practice of obstetric anaesthesia and analgesia in Malaysia [correction of Malaysis]. AB - OBJECTIVE: A survey covering 30% of the deliveries in Malaysia was done to determine the practice of obstetric anaesthesia and analgesia for 1996. RESULTS: From the survey, it was found that the regional anaesthesia rate for caesarean section was 46% in the government hospitals compared to 29.2% in the private hospitals, with spinal anaesthesia being the most common regional anaesthetic technique used in both types of hospitals. The epidural rate for labour analgesia was only 1.5% overall for the country. Epidural analgesia services were available in all private hospitals whereas 17.6% of government hospitals surveyed did not offer this service at all. CONCLUSIONS: Although the use of epidural analgesia for labour was low in Malaysia, the overall rate of regional anaesthesia for caesarean section (41.9%) is very much in keeping with the standards of safe practice recommended by the United Kingdom. PMID- 10870308 TI - Peritoneal and peripheral B-1-cell populations in patients with endometriosis. AB - OBJECTIVE: The purpose of this study was to investigate the frequency of B-1 cells in the peritoneal cavity and peripheral blood of patients with endometriosis. MATERIALS AND METHODS: We examined 31 patients with endometriosis and 14 normal nonpregnant women. Peripheral blood cells and peritoneal exudate cells (PECs) were stained with FITC or PE-labeled anti-CD5/CD19 monoclonal antibodies. Immunofluorescence analysis was performed using a flow cytometer. The significance of differences between the patient and control groups was determined by the non-parametric Mann-Whitney test. RESULTS: There was no significant difference in the percentages of B-1 cells in the peripheral blood of women with and without endometriosis (median, 22.7%; range, 4.7-92.3% vs median, 20.05%; range, 11.1-12.6%, respectively). Endometriosis patients with antinuclear antibodies (ANAs) demonstrated significantly elevated B-1 cells compared to both endometriosis patients without ANAs and normal controls (p < 0.005 and p < 0.05, respectively). Endometriosis patients demonstrated significantly higher B-1 cell populations (B-1 cells/total B-cell ratio) in PECs than did non-endometriosis patients (p < 0.05). CONCLUSIONS: The peripheral B-1-cell population in patients with endometriosis is related to ANA production. B-1 cells might play important roles in the development of endometriosis through autoantibody production. PMID- 10870309 TI - Prophylactic cervical cerclage for the prevention of early premature delivery in nulliparous women with twin pregnancies. PMID- 10870310 TI - Cytogenetic pattern of childhood leukemia in Taiwan. AB - BACKGROUND: Cytogenetic analyses of leukemic cells can be used to define specific subgroups of leukemia with different prognoses and, thereby, indicate appropriate treatment for individual cases. In this study, we investigated the cytogenetic pattern of childhood leukemia in Taiwanese patients. METHODS: A modified trypsin method of Seabright was used for G-banding of metaphase cells. RESULTS: From October 1996 to January 1999, 111 children with a diagnosis of leukemia were enrolled in the study. Of these, 73 patients had a diagnosis of acute lymphoblastic leukemia (ALL) and 63 of these patients had successful karyotyping of their leukemic cells. Among them, 20 (30.3%) had a normal karyotype, five had hypodiploidy (all had 45 chromosomes), five had low hyperdiploidy (47-50 chromosomes), 16 (24.2%) had high hyperdiploidy (> 50 chromosomes), and 20 had pseudodiploidy. Chromosomal translocation was identified in 24 (36.4%) of the ALL patients, 17 of whom had recurrent translocations including 10 with CD10+ B precursor ALL [4 with t(9;22), 5 with t(1;19), and 1 infant with t(8;14)(q24;q11)], one neonate with CD10- early pre-B ALL with t(4;11), three B cell cases with t(8;14), and three T-cell cases [2 with t(11;19)(q23;p13), and 1 (11;14)(p13;q11)]. One B-precursor patient had dic (9;12). Karyotypes of the 30 patients with acute myeloid leukemia (AML) included eight with t(8;21); seven with the French-American-British-M2 subtype (FAB-M2) and one with M1. All four of the patients with M3 had t(15;17), one patient with M4 had inv(16) and 7q-, one with M4Eo (M4 with eosinophilia) had t(7;16)(q21;q22), one with M0 had t(4;11)(q21;q23), and the remaining 11 had a normal karyotype. Three of the five adult-type chronic myeloid leukemia patients had standard Philadelphia chromosome, and the other two had a variant-form of Philadelphia chromosome. Both of the patients with juvenile myelomonocytic leukemia and one patient with myelodysplastic syndrome had a normal karyotype. CONCLUSIONS: Most findings were similar to previous reports. Although the high proportion of FAB-M2 patients (7/8) with t(8;21) and the consequently higher frequency (26.7%) of this translocation in the 30 AML cases in this study might have significance, a larger series of cases is needed to establish this finding. PMID- 10870311 TI - Strong association of antiepithelial cell antibodies with HLA-DR3 or DR7 phenotype in patients with recurrent oral ulcers. AB - BACKGROUND: Previous studies showed that antiepithelial cell antibodies (anti ECA) were present in 71% (15/21) of patients with recurrent oral ulcers (ROU) and that there was a strong association of human leukocyte antigen (HLA)-DRw9 with ROU in Chinese patients. In this study, we assessed anti-ECA in a larger group of Chinese patients with ROU (n = 88) in order to further investigate the association of anti-ECA with HLA-DR and -DQ antigens. METHODS: The anti-ECA in the sera of ROU patients were detected by an indirect immunofluorescence technique with rat esophagus as the substrate, and the HLA-DR and -DQ antigens in ROU patients were typed by a standard microcytotoxicity assay using Terasaki's oriental tray. RESULTS: The rate of anti-ECA positivity was significantly higher (p < 0.0001) in ROU patients (68%) than in healthy control subjects (0%). Furthermore, the rate of anti-ECA positivity in patients with major or minor oral ulcers (72%) was significantly higher (29%) than that in patients with herpetiform ulcers (p < 0.05). There was a significant increase in the frequency of DR3 or DR7 antigen expression (p < 0.0001, pc [p corrected] < 0.001, relative risk [RR] = 4.3, etiologic fraction = 0.41) in anti-ECA-positive ROU patients compared with the corresponding frequencies in healthy control subjects. There was also a significant increase in the frequency of DR7 or DRw9 antigen expression (p < 0.005, pc < 0.05, RR = 4.7, etiologic fraction = 0.45) compared to healthy controls. CONCLUSIONS: Because only DR3 or DR7 antigen occurred more frequently in anti-ECA-positive than in anti-ECA-negative ROU patients (p < 0.0007, pc < 0.05, RR = 19.6, etiologic fraction = 0.51), we concluded that the gene coding for DR3 or DR7 antigen may contribute to the presence of anti-ECA in Chinese patients with ROU. PMID- 10870312 TI - Metabolic disorders mimicking Reye's syndrome. AB - BACKGROUND: Several metabolic disorders such as encephalopathy and hepatic dysfunction have been described as Reye's-like syndrome because they present with similar clinical manifestations that mimic Reye's syndrome. We performed a retrospective study to explore the underlying metabolic etiologies of Reye's-like syndrome in patients treated at National Taiwan University Hospital. METHODS: From January 1991 to June 1998, 19 children with a syndrome fitting the Reye's like syndrome description were identified for study. Urine organic acid analysis, plasma amino acid analysis, liver pathology, and skin fibroblast enzyme assays were studied during the acute stage of illness. RESULTS: The etiologies of patients' syndromes included urea cycle disorders (n = 7), glycogen storage disease type Ia (4), primary carnitine deficiency (2), hereditary fructose intolerance (1), methylmalonic acidemia (2), and 3-hydroxy-3-methylglutaric acidemia (1). Fatty acid oxidation defects were suspected in the remaining two cases. CONCLUSIONS: A significant number of patients who present with Reye's-like syndrome have an underlying inherited metabolic disorder. In patients with Reye's like syndrome, an accurate diagnosis is essential to ensure normal growth and development and to prevent recurrence of the condition. PMID- 10870313 TI - Microbiologic features of adult community-acquired bacterial meningitis in Taiwan. AB - BACKGROUND AND PURPOSE: Community-acquired bacterial meningitis (CABM) is a life threatening disease that requires prompt initiation of appropriate antibiotic therapy. The purpose of this study was to determine the causative microorganisms of CABM and their antimicrobial susceptibility patterns at a major teaching hospital in Taipei from 1993 to 1998. METHODS: A review of medical records and microbiologic data was used to identify cases of CABM and causative pathogens. Antimicrobial susceptibility testing for bacterial isolates was performed by the disk diffusion method. RESULTS: Among the 48 adult patients with a diagnosis of CABM during the study period, the causative pathogens were identified in 36 cases. Unlike reports from other countries, Klebsiella pneumoniae was the leading causative pathogen (33%), followed by Streptococcus pneumoniae (28%), Listeria monocytogenes (11%), Neisseria meningitidis (6%), Staphylococcus aureus (6%), streptococci (6%), and Pseudomonas aeruginosa (6%). The incidence of CABM due to K. pneumoniae increased during the study period (p = 0.012, Poisson regression), while the incidence of CABM due to other pathogens remained stable. All of the CABM-associated K. pneumoniae isolates were susceptible to cefotaxime but 25% of the CABM-associated S. pneumoniae strains were not susceptible to penicillin G. CONCLUSIONS: Penicillin G alone was not an appropriate empiric therapy for adult CABM because a high percentage of cases were due to K. pneumoniae or penicillin nonsusceptible S. pneumoniae. While the recommendations for the initial empiric regimen for CABM due to S. pneumoniae in Taiwan remain to be developed, third generation cephalosporins appear to be an appropriate initial empiric regimen for the treatment of CABM due to K. pneumoniae. PMID- 10870314 TI - Characterization of a highly glycopeptide-resistant Enterococcus gallinarum isolate. AB - BACKGROUND AND PURPOSE: Adequate treatment of emergency infection involving antibiotic-resistant bacteria such as vancomycin-resistant Enterococcus requires a convergence of clinical and bacteriologic techniques. An isolate of Enterococcus gallinarum, designated as TSGH63, is known to be uncommonly vancomycin-resistant. This study investigated the genetic determinant for this unique characteristic. METHODS: After completing the conventional identification and sensitivity tests, the genomic content of E. gallinarum TSGH63 was extracted and analyzed by pulse-field electrophoresis. A set of specific primers for vanA, vanB, vanC1, and vanC2/C3 genes was then applied in a multiplex polymerase chain reaction (PCR) to differentiate its genetic content. To locate the determinant for high vancomycin resistance, the electrophoresis profile was further analyzed by Southern blot using the digoxigenin (DIG)-labeled vanA gene probe. Finally, interspecies transfer of the vancomycin-resistance determinant of E. gallinarum TSGH63 was tested by a conjugation experiment in vitro. RESULTS: A 50-kb plasmid was identified in the analysis of the genomic extract of E. gallinarum TSGH63 by pulse field electrophoresis. Using multiplex PCR, we demonstrated that E. gallinarum TSGH63 harbors a vanA gene in addition to a vanC1 gene. The DIG labeled vanA gene-specific probe bound to the plasmid exclusively on the Southern blot. The plasmid-carried vanA gene, but not the vanC1 gene, was found to be transferable from TSGH63 to E. faecalis JH2-2 by conjugation in vitro. CONCLUSIONS: This is the first report of isolation of E. gallinarum with a high level of resistance to glycopeptides in Taiwan. The demonstrated interspecies transfer of the vancomycin-resistance gene highlights the importance of stringent control of the use of vancomycin. PMID- 10870315 TI - Verapamil modulation of multidrug resistance in renal cell carcinoma. AB - BACKGROUND AND PURPOSE: Renal cell carcinoma (RCC) is well known for its chemoresistance. The membranous p-glycoprotein (gp-170) is believed to be highly correlated with multidrug resistance (MDR) of cancer cells with energy-dependent pumping efflux of anticancer drugs. Verapamil, a calcium antagonist, inhibits the efflux function of gp-170 and cytoskeletal transportation. The aim of this study was to determine the effect of verapamil on gp-170 expression and intracellular drug accumulation in RCC tumor cells and the modulation of cytotoxicity of various chemotherapeutic drugs on native RCC cell lines and acquired MDR sublines by verapamil. METHODS: Using cultured cell lines of RCC and their MDR sublines as target cells, the effect of verapamil on gp-170 expression was analyzed by immunofluorescence flow cytometry. The influence of verapamil on intracellular drug accumulation in RCC tumor cells was measured by autofluorescence flow cytometry. The modulation of verapamil on cytotoxicity of various chemotherapeutic drugs on native RCC cell lines and acquired MDR sublines was analyzed by the methyl tetrazolium method. RESULTS: From flow cytometric measurement, the expression of gp-170 was significantly decreased in A704 and Caki-1 tumor cells after verapamil treatment. The uptake of adriamycin and maintenance of intracellular drugs were also significantly increased following verapamil treatment in RCC8701 tumor cells. These effects were sustained for as long as 8 hours after verapamil withdrawal. The cytotoxicity of adriamycin and epirubicin on RCC8701 and its MDR subline tumor cells was markedly intensified by verapamil. The verapamil modulation of cytotoxicity was in an immediate-reaction pattern and was dose-dependent, with synergistic effects. Long-term treatment was more effective than short-term treatment in RCC MDR sublines. The cytotoxicity of vinca alkaloid (vinblastine) and alkylators (carboplatin) was also enhanced by verapamil. CONCLUSIONS: These results suggest that verapamil plays an important role in the circumvention of native and acquired chemoresistance of RRC because it suppresses membranous gp-170 expression and cytoplasmic drug transportation. PMID- 10870316 TI - Cutaneous sarcoidosis among Taiwanese. AB - BACKGROUND AND PURPOSE: Sarcoidosis is a multisystem disorder characterized by noncaseating, naked, epithelioid granulomas, and commonly involving the hilar lymph nodes, lungs, skin, and eyes. Once thought to be rare in Chinese, it is being encountered with increasing frequency in Taiwan. In this study, we examined the clinicopathologic findings of sarcoidosis in Chinese patients in Taiwan and investigated the role of Mycobacterium tuberculosis in the disease. METHODS: The clinicopathologic features of all patients treated for sarcoidosis at a tertiary care hospital in Taiwan during a 10-year period were retrospectively examined. Nested polymerase chain reaction (PCR), using the IS6110 sequence as the amplification target, was used to detect M. tuberculosis complex DNA in formalin fixed, paraffin-embedded skin specimens. RESULTS: Sarcoidosis was diagnosed in a total of 12 patients (7 men, 5 women; mean age, 53 yr) during the study period. All patients presented with skin lesions varying from erythematous papules or nodules mainly located on the face (8 patients), to widespread papules, large plaques, and tumors on the extremities, trunk, or scalp (4 patients). Seven of the 12 patients had extracutaneous involvement. One developed cutaneous T cell lymphoma after 14 years of severe disease. Histopathologic examination of skin lesions revealed sarcoid or mixed sarcoid/tuberculoid granulomatous infiltration without evidence of mycobacterial, deep fungal infection, or foreign body material. Nested PCR failed to detect M. tuberculosis complex DNA in skin specimens. CONCLUSIONS: Our data suggest that sarcoidosis manifests with a wide range of cutaneous lesions in Taiwanese patients and that extracutaneous involvement is not rare. Dermatologists and general practitioners should be made aware that sarcoidosis may first manifest with skin lesions, so that the correct diagnosis can be made promptly. M. tuberculosis did not appear to be associated with sarcoidosis in this study group. PMID- 10870317 TI - Cartilage repair by free periosteal grafts in the knees of pigs: a histologic study. AB - BACKGROUND AND PURPOSE: Periosteal grafts may result in cartilage formation and, therefore, have the potential to repair cartilage defects. We evaluated the histologic results of free periosteal grafts for the repair of full-thickness cartilage defects in pigs. METHODS: A free autogenous periosteal flap from the proximal tibia was grafted to a size-matched, full-thickness articular defect on the lateral femoral condyle of the knees of 12 pigs. The same defect on the medial femoral condyle was used as a control lesion. Biopsies were performed at 4, 8, and 12 weeks after grafting. RESULTS: The control lesions showed dense fibrous tissue with no evidence of cartilage-like tissues. The predominant tissues after grafting were mixtures of fibrous tissue, fibrocartilage, mesenchyme tissue, and occasional bone islands, but no cartilage tissue was identified. The tissue distribution did not change in the same knee from the week 4 to the week 8 biopsy; nonetheless, there were interindividual variations in tissue distribution. CONCLUSIONS: The results of this study do not support the use of free periosteal transplantation for full-thickness cartilage defects of the knee. PMID- 10870318 TI - Long-term effects of azathioprine therapy for juvenile rheumatoid arthritis. AB - BACKGROUND AND PURPOSE: Juvenile rheumatoid arthritis (JRA) can result in disability, growth disturbance, and systemic complications. This study investigated the efficacy and adverse effects of azathioprine (AZA) therapy in children with JRA. METHODS: Data from the medical records of 24 children with JRA treated with oral AZA during the period from 1988 to 1998 were retrospectively analyzed. All 24 patients had received two or more nonsteroidal anti-inflammatory drugs (NSAIDs) and 12 had received disease-modifying antirheumatic drugs (DMARDs) prior to the start of AZA. Of the 24 patients, 21 were corticosteroid-dependent prior to the onset of AZA therapy. The indication for AZA therapy was lack of efficacy of the current treatment regimen. The initial and maximal doses of AZA averaged 1.7 mg.kg-1.d-1 (range, 1-3 mg. kg-1.d-1) and 1.9 mg.kg-1.d-1 (range, 1 6 mg.kg-1.d-1), respectively. The mean duration of treatment was 13 months (range, 4-37 mo). The mean duration of follow-up was 45 months (range, 7-137 mo) from the start of AZA therapy. RESULTS: Fifteen children (62.5%) showed clinical improvement, while the other nine (37.5%) achieved clinical remission. AZA treatment resulted in a more than 50% reduction in the required corticosteroid dose in seven children and complete discontinuation of corticosteroid administration in eight children. None of the patients treated with AZA doses of 1 to 3 mg.kg-1.d-1 developed AZA-related side-effects. Two patients suffered from AZA-related adverse effects due to AZA overdose (6 mg.kg-1.d-1). Both experienced pancytopenia and disseminated infection, which resolved following reduction of the AZA dose to 3 mg.kg-1.d-1. CONCLUSIONS: AZA is an effective and well tolerated steroid-sparing agent for JRA refractory to NSAIDs or DMARDs. PMID- 10870319 TI - HeartMate left ventricular assist device for long-term circulatory support. AB - We describe three successful cases of HeartMate left ventricular assist device (LVAD; Thermo Cardiosystems, Woburn, MA, USA) implantation in patients with end stage heart failure for long-term circulatory support. Patient 1 was a 34-year old woman with postpartum cardiomyopathy. Patients 2 and 3 were both males with dilated cardiomyopathy, 50 years and 21 years of age, respectively. They all presented in cardiogenic shock with decreased sensorium and anuria. Temporary mechanical support with an intra-aortic balloon pump or extracorporeal membrane oxygenation (ECMO) was needed for life support. Because bleeding and right ventricular failure often occur after HeartMate LVAD implantation, we used a Vascutek tube (Vascutek Ltd, Inchinnan, Scotland) graft to wrap inflow and outflow valve conduits and ECMO as a bridge to HeartMate LVAD implantation. Following surgery, cardiac output increased from 2.70, 2.06 and 2.53 L/min to 4.50, 5.80 and 5.00 L/min in the three patients. HeartMate LVAD can provide safe and stable long-term circulatory support without the need for anticoagulation. One of the patients remained on HeartMate for 287 days before undergoing successful heart transplantation. Patients with HeartMate LVAD are ambulatory and may be discharged while awaiting heart transplantation. Heart function may recover after long-term ventricular unloading with HeartMate LVAD. PMID- 10870320 TI - Pregnancy complicated by concurrent primary hyperparathyroidism and arrhythmia. AB - Primary hyperparathyroidism during pregnancy results in a high rate of fetal complications and maternal morbidity. Maternal hypercalcemia in pregnancy results in fetal hypercalcemia, which leads to suppression of fetal parathyroid function. Spontaneous abortion and stillbirth can occur, and the loss of maternal calcium after birth leads to neonatal hypocalcemia. It is essential to detect primary hyperparathyroidism during pregnancy because early diagnosis and management can decrease the rate of fetal and maternal complications. We present the case of a 27-year-old gravida 1, para 0 woman whose pregnancy was complicated by hyperparathyroidism and arrhythmia. The patient complained of dyspnea and palpitations in the seventh and 15th weeks of gestation. Electrocardiography showed ventricular premature contraction bigeminy and trigeminy in association with hypercalcemia (3.3 mmol/L). A parathyroidectomy in the second trimester revealed parathyroid adenoma. Hypercalcemia and arrhythmia resolved completely and the patient delivered a term baby without any maternal or fetal complications. The simultaneous occurrence of arrhythmia with ventricular premature contractions and hyperparathyroidism in pregnancy is rarely reported. Palpitations and dyspnea due to arrhythmia may be associated with primary hyperparathyroidism in pregnancy and should be considered in the differential diagnosis. In the management of symptomatic primary hyperparathyroidism during pregnancy, surgical intervention is preferable in the second trimester when organogenesis is completed and the risk of spontaneous abortion is low. PMID- 10870321 TI - Laparoscopic cystectomy of a twisted, benign, ovarian teratoma in the first trimester of pregnancy. AB - Adnexal torsion is an unusual, but serious complication in pregnancy. The treatment is surgical, but this may increase the risk of pregnancy loss in the first trimester. The use of laparoscopic surgery, which is less invasive than traditional laparotomy, has been limited by diagnostic and technical difficulties including determination of ovarian tumor nature and spillage of cyst contents intraoperatively. A 25-year-old woman in her 11th week of pregnancy had acute severe left lower-abdominal pain, which was diagnosed as left ovarian teratoma with torsion. She underwent emergency laparoscopic surgery with unwinding of the twisted fallopian tube and ovary and cystectomy of the teratoma. The patient subsequently delivered a full-term baby, without complications. Accurate ultrasound and cytologic diagnoses along with copious intraoperative warm, normal saline irrigation were likely contributing factors to the successful outcome of this case. PMID- 10870322 TI - Orthotopic bladder substitution in women using the ileal neobladder. AB - Before 1990, orthotopic bladder substitution was limited to men and urethrectomy was routinely performed in women--making orthotopic reconstruction impossible. Following studies on pelvic exenteration for transitional cell bladder cancer patients, and after anatomic studies of the female urethra, the first report on orthotopic bladder substitution was published in 1994. Here, we present our ileal neobladder technique, which was applied for the treatment of bladder cancer in three women, and sigmoid colon cancer with vesical metastasis in one woman. This surgical procedure preserves the urethral sphincter, the endopelvic fascia, and the pubourethral ligaments. The innervation of the sphincter is also maintained. A 45-cm segment is isolated from the distal ileum and arranged in a W-shaped configuration. An anastomosis between the ileal bladder and the urethra is made and an antireflux ureteroileal implantation is created. Postoperatively, all patients reported daytime continence. The average urinary flow rate was 13.5 +/- 2.1 mL/sec 3 months after surgery. The reservoir had low pressure and high capacity. There was no reflux or deterioration of renal function. In properly selected female patients undergoing radical cystectomy, an ileal neobladder is now an option. PMID- 10870323 TI - Neonatal intussusception due to a cecal duplication cyst. AB - Intussusception in the first month of life is rare; however, it should be considered a distinct clinical and pathologic entity. Cecal duplication as the cause of intussusception in a neonate is extremely rare. We report a case of a newborn with ileocecocolic intussusception. She presented with vomiting and bloody stools. Abdominal sonography revealed a target lesion with a cystic component. We performed a laparotomy and found an ileocecocolic-type intussusception which was caused by a cecal duplication cyst. Right hemicolectomy with ileocolostomy was performed. She remained well at one year follow-up. PMID- 10870324 TI - Indolent cutaneous mucormycosis with pulmonary dissemination in an asthmatic patient: survival after local debridement and amphotericin B therapy. AB - We describe a 68-year-old asthmatic female patient with multiple pulmonary cavities. A preexisting ecthyma on the left lower leg became erythematous and swollen during exacerbation of her asthma which was under treatment with high dose steroids. Nonseptate broad hyphae were found in her sputum, pus from the wound, and debrided skin tissue. Hematogenous spread of septic emboli from indolent cutaneous mucormycosis to both lungs was the suspected mechanism of dissemination. High-dose steroid therapy may have been the major contributory factor. The patient was successfully treated with local surgical debridement of the wound and intravenous amphotericin B. PMID- 10870325 TI - DNA polymorphisms at the apolipoprotein A1-CIII loci in Taiwanese: correlation of plasma APOCIII with triglyceride level and body mass index. AB - BACKGROUND AND PURPOSE: Apolipoprotein (APO) A1-CIII genes are linked within a 2.6-kb region on human chromosome 11. ApoA1 is the main component of high-density lipoprotein (HDL), and apoCIII inhibits lipoprotein lipase activity. Genetic variations in APOA1-CIII may affect the function of apoA1/apoCIII and plasma lipid/lipoprotein levels, and thus, the risk of developing atherosclerosis. This study compared the frequency distributions of genetic variations in APOA1-CIII genes and their influence on plasma lipid concentrations in Taiwanese patients with coronary artery disease (CAD) and in healthy controls. METHODS: Six restriction site variations (RSVs) of the APOA1-CIII gene complex were investigated by DNA amplification using polymerase chain reaction and restriction enzyme digestion in 229 control subjects and 131 CAD patients during the period from 1992 through 1996. The blood lipid profiles of these subjects were also determined. RESULTS: Thirty-seven distinct six-RSV genotypes were observed. Separate comparisons of the frequency distributions of the six genetic variations showed no significant differences between CAD patients and controls subjects, but the combined six-RSV-genotypes showed different frequency distributions between these two groups. Nine of the 37 six-RSV genotypes were found only in the CAD patients and higher frequencies of two of these types were observed in the CAD patients than in healthy controls. The effects of these genetic variations were on high-density lipoprotein cholesterol in women (for MspIB, PstI, SstI and PvuII RSV) and total cholesterol (for PvuII RSV), low-density lipoprotein cholesterol (for XmnI RSV), and apolipoprotein B (for MspI and SstI RSV) levels in men in the control group. Elevated plasma apoCIII concentration was significantly associated with an increased plasma triglyceride level and body mass index in the control group (P < 0.0001). CONCLUSIONS: Analysis of the frequency distribution of six RSVs of the APOA1-CIII gene complex in Taiwanese CAD patients and control subjects showed that the effect of genotype on plasma lipid levels was gender specific and that the apoCIII level was closely associated with plasma triglyceride level and body mass index. PMID- 10870327 TI - Modified technique for introducer-type percutaneous endoscopic gastrostomy and jejunostomy: assisted by a novel trocar and fasteners. AB - BACKGROUND AND PURPOSE: Commercially available kits for percutaneous endoscopic gastrostomy (PEG) and jejunostomy comprise a substantial proportion of the cost of patients for this procedure. A modified introducer-type technique and new instrumentation for PEG and jejunostomy that substantially reduces the cost of the kit were tested for efficacy and safety. METHODS: This technique was tested on 10 pigs as a pilot study, and then applied to seven consecutive patients undergoing gastrostomy and three patients undergoing gastrostomy with jejunostomy. The endoscopy and site selection for gastrostomy were the same as in standard PEG. Two novel fasteners for fixing the gastric wall to the abdominal wall and a guide-wire in the selected site were inserted separately into the stomach through 15-gauge needles. A stainless steel trocar with a detachable sheath was introduced into the stomach over the guide-wire. After the inner stylet was removed, a 24-French Foley catheter was inserted as a feeding tube. Gastrostomy was completed after balloon inflation and external fixation. If jejunostomy was indicated, a 12-French nasogastric tube was inserted through the gastrostomy. The procedure time, complications, and costs were compared with those for another 15 consecutive patients who underwent the conventional pull through method of PEG and jejunostomy using commercially available kits. RESULTS: No significant difference was found in procedure time between patients who underwent the modified or conventional gastrostomy procedures (mean +/- standard deviation, 15.4 +/- 5.6 min). There was a similar incidence of short-term complications in the two treatment groups. The feeding catheters required replacement more quickly than did those in the commercial kits (80 +/- 58 vs 217 +/- 140 d). The cost to patients was much less with the new method than with conventional PEG. CONCLUSIONS: The new gastrostomy method achieves the same medical quality at far less cost for patients. PMID- 10870326 TI - Comparison of different diagnostic methods for lupus pleuritis and pericarditis: a prospective three-year study. AB - BACKGROUND AND PURPOSE: Pleural or pericardial effusions, or both, are commonly encountered, but the differential diagnosis is sometimes difficult. We evaluated the diagnostic value of effusion immunofluorescent antinuclear antibody (ANA) titer, systemic lupus erythematosus (SLE) latex agglutination slide test, and cytologic LE cell examination in patients with pleural and/or pericardial effusions of various etiologies. METHODS: A total of 153 pleural and/or pericardial effusion specimens were collected by aspiration from 152 patients (14 SLE and 138 non-SLE patients). All specimens were sent for routine biochemistry testing, determination of ANA titer, SLE latex agglutination slide test, and LE cell examination. RESULTS: Ten of the 14 SLE patients had lupus serositis and all of them had high ANA titers (> or = 1:160) in their effusions. SLE latex and LE cell tests were positive in seven and eight patients with lupus serositis, respectively. The remaining four SLE patients with effusion of etiologies other than lupus serositis had low or negative effusion ANA titers. Among the non-SLE patients, 29 of 112 patients (26%) with pleural effusion and six of 26 patients (23%) with pericardial effusion had positive ANA tests (> or = 1:40). None of them had a positive SLE latex or LE cell test result. Thirteen of the 138 non-SLE patients (11%) had high effusion ANA titers (> or = 1:160). Effusion in 11 of 13 non-SLE patients (85%) was due to malignancy. CONCLUSIONS: Effusion ANA titer detection is a very sensitive but nonspecific test for the diagnosis of lupus serositis. SLE latex and cytologic LE cell tests can aid in the differential diagnosis as complementary tools. The specificity, positive and negative predictive values of these two tests are excellent for the diagnosis of lupus serositis. PMID- 10870328 TI - Videourodynamic study for diagnosis of bladder outlet obstruction in women. AB - BACKGROUND AND PURPOSE: In order to characterize the etiology of bladder outlet obstruction (BOO) in Taiwanese women, the results of videourodynamic studies performed in women with lower urinary tract symptoms (LUTS) were analyzed. The treatment results were compared with the underlying pathophysiology among the three etiologies of urethral stricture, spastic urethral sphincter, and cystocele. METHODS: From October 1997 through February 1999, 364 female patients underwent videourodynamic study to investigate the underlying etiology of LUTS. BOO was defined as a voiding detrusor pressure of 50 cm H2O or greater and a narrow urethra on the voiding cystourethrography. Cystoscopy, urethral sounding, and external sphincter electromyography were used in conjunction with the pressure/flow findings for differential diagnosis of the etiology of BOO. RESULTS: Among the 364 patients with LUTS who underwent videourodynamic study, 35 (9.6%) had BOO. Detailed investigation revealed that 15 of these women had urethral stricture (43%), 14 had spastic urethral sphincter (40%), and six had cystocele (17%). No significant difference was found in the urodynamic parameters among these three groups of patients. The incidence of bladder trabeculation and urge incontinence was significantly higher in patients with urethral stricture. Improvement of urine flow and voiding symptoms was achieved in 87% of the patients with urethral stricture, 50% female patients with spastic urethral sphincter, and 100% of those with cystocele. CONCLUSIONS: The results suggest that BOO is not uncommon in women with LUTS, and that a correct diagnosis of BOO should be based on comprehensive urodynamic study. PMID- 10870329 TI - Comparison of two-dimensional and three-dimensional techniques for determination of facial motion--absolute movement in a local face frame. AB - BACKGROUND AND PURPOSE: Few studies have used motion analysis in the study of facial animation. A facial animation model using an expert vision motion analysis system was developed in this study to quantitatively evaluate absolute movement during five facial animations. METHODS: Skin markers were adhered to the face of each subject at 16 anatomic landmarks selected to represent the functional movement of the facial muscles. Three of the 16 skin markers were used to establish a local face frame with the origin in the mid-point of the face. All of the coordinates measured in the laboratory frame were rotated and translated to the local face frame for analysis. The trajectory of the facial markers was evaluated from the local face frame when comparing two-dimensional (2-D) displacement of skin markers (frontal plane) with three-dimensional (3-D) values by paired Student's t-test. RESULTS: Although the correlation of 2-D and 3-D displacements of skin markers was high (r > 0.69), the differences between 2-D and 3-D motion were significant (p < 0.001). The 2-D displacement of skin markers underestimated the 3-D facial animation in each marker and animation. There were no significant differences in the movement of both mouth angles or of the eyes. Thirty repeated measurements of a subject revealed good concentration in 3-D displacement, velocity, and angle of movement in smiling. The measurement error was less than 0.06 mm. The normative displacement of individual anatomic landmarks was evaluated to avoid size differences of individual faces. CONCLUSIONS: We suggest that the expert vision motion analysis technique is feasible for quantitative evaluation of absolute facial movement and would be useful for further clinical evaluation of patients with facial palsy. PMID- 10870330 TI - Molecular diagnosis of fragile X syndrome and distribution of CGG repeats in the FMR-1 gene in Taiwanese. AB - BACKGROUND AND PURPOSE: Fragile X syndrome, the most frequent form of inherited mental retardation, is caused by abnormal expansion of the CGG trinucleotide repeats in the 5' untranslated region of the FMR-1 gene. In this study, we describe the prenatal diagnosis of fragile X syndrome and the distribution of CGG repeat numbers in the FMR-1 gene, which has not been previously reported in Taiwanese. METHODS: Using polymerase chain reaction (PCR), we determined the range of the CGG repeats in the FMR-1 gene in 316 normal individuals (350 X chromosomes) and 349 mentally retarded patients (429 X chromosomes). For prenatal diagnosis of fragile X syndrome, DNA extracted from amniotic fluid cells was used for PCR determination of CGG repeats. RESULTS: Because there were no significant differences between the distribution of the (CGG)n alleles between the mentally retarded and normal subjects, the data were pooled. Among the 779 X chromosomes studied, 24 different alleles were identified with a low of 16 and a high of 45 CGG repeats. The 29 repeat allele was the most common, followed by the 30 and the 28 repeat alleles. We effectively amplified slightly expanded premutation alleles of up to about 90 CGG repeats. In the prenatally diagnosed fetus, a normal 29 repeat allele was found. CONCLUSIONS: Determination of the distribution of the CGG repeats in the FMR-1 gene in Taiwanese is useful in genetic counseling regarding fragile X syndrome. Prenatal molecular diagnosis of the syndrome can be successfully performed using amniotic fluid cells. PMID- 10870331 TI - In vitro activity of rifabutin and rifampin against clinical isolates of Mycobacterium tuberculosis in Taiwan. AB - BACKGROUND AND PURPOSE: To determine the in vitro activity of rifabutin against Mycobacterium tuberculosis (MTB) and the cross-resistance rate between rifampin and rifabutin. METHODS: A total of 56 clinical isolates of MTB, including 23 multidrug-resistant (MDR) isolates and 33 susceptible isolates, were tested for susceptibility to rifampin and rifabutin using the absolute concentration method. The concentrations of drugs tested were 2.5 and 5 mg/mL for rifampin and 0.1, 0.5, 1, 2.5, 5, and 10 mg/mL for rifabutin. RESULTS: All 33 MTB isolates that were susceptible to rifampin were also susceptible to rifabutin. None of the 23 MDR-MTB isolates were inhibited by rifabutin at a concentration of 0.1 mg/mL. Among these 23 MDR isolates, three were susceptible to rifabutin at concentrations > or = 0.5 mg/mL, six were susceptible to rifabutin at concentrations > or = 5 mg/mL, 18 were susceptible to rifabutin at concentrations > or = 10 mg/mL and five were not inhibited at any of the concentrations tested. The cross-resistance rate between rifampin and rifabutin was 87%. CONCLUSIONS: Our results indicate that the in vitro activity of rifabutin against drug susceptible MTB isolates is greater than that of rifampin. For MDR-MTB isolates, the cross-resistance is high between rifampin and rifabutin. PMID- 10870332 TI - Effects of norepinephrine on apoptosis in rat neonatal cardiomyocytes. AB - BACKGROUND AND PURPOSE: Norepinephrine (NE) is elevated in heart failure and can induce apoptosis in adult cardiac myocytes. However, it is not known whether NE can induce apoptosis in neonatal cardiac myocytes. This study examined the ability of NE to stimulate apoptosis in rat neonatal cardiac myocytes in vitro. METHODS: Neonatal rat cardiac myocytes were exposed to NE alone, NE + propranolol, or NE + prazosin for 24 hours. Apoptosis was assayed by DNA laddering with agarose gel electrophoresis and immunofluorescent terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. Reverse transcription polymerase chain reaction was used to evaluate the expression of Mcl-1. Creatine kinase activity in the cultured medium was used as a measure of the toxicity of NE on myocytes. RESULTS: NE increased DNA laddering on agarose gel electrophoresis and increased the number of apoptotic cells in a dose-dependent manner. No increase in apoptosis was found in response to NE doses between 1 and 50 mumol/L. NE at concentrations of 100 to 400 mumol/L increased apoptosis from 10% to 31% of cells. The ability of NE to stimulate apoptosis in rat neonatal cardiac myocytes was completely blocked by propranolol, but not prazosin. NE treatment at high concentrations sharply reduced the level of Mcl-1 mRNA, coincident with the increase in the number of apoptotic cells. Creatine kinase activity in the cultured medium was similar among the controls and NE treated myocytes. CONCLUSIONS: Our results showed that NE at high concentrations stimulated apoptosis in rat neonatal cardiac myocytes in vitro. Apoptosis induced by NE was associated with down-regulation of Mcl-1. However, NE at the same concentration was not toxic to rat neonatal cardiac myocytes. PMID- 10870333 TI - First successful ventricular septation of double inlet left ventricle in Taiwan. AB - Patients with a double inlet ventricle may undergo surgery using a modified Fontan procedure, in which the pulmonary ventricle is not utilized, or a procedure in which a pulmonary ventricle is created through ventricular septation. Ventricular septation is preferred to the Fontan procedure because there is better cardiorespiratory response to exercise after surgery. A 4-year old girl with Holmes heart underwent ventricular septation on 12 May 1998. Pulmonary artery banding had been performed at 3 months of age and rebanding 16 days later. She was well and continued to grow. Ultrafast computed tomography and cardiac catheterization prior to surgery showed a double inlet left ventricle (LV) connected to a right posterior aorta with a right-sided rudimentary right ventricle that drained to the left anterior pulmonary trunk. Left ventricular end diastolic volume was 218% of normal and the ejection fraction was 79%. After debanding and enlargement of the bulboventricular foramen, a 3 x 4-cm composite patch of equine pericardium and Dacron velour was used to septate the ventricle, with transmural stitching at the apical portion. The patient survived the operation with complete atrioventricular block, and was extubated 6 days later. A permanent pacemaker was implanted 1 month later. One year after surgery, she was doing well. Echocardiography revealed paradoxical septal motion with good ventricular function. This is the first report of successful ventricular septation of a double inlet left ventricle performed in Taiwan. PMID- 10870334 TI - Well-differentiated fetal adenocarcinoma of the lung. AB - We describe the case of a 43-year-old woman with a tumor shadow in the upper lobe of the left lung. The tumor was initially suspected to be a carcinoid tumor, following percutaneous needle biopsy. Subsequently, a left upper lobectomy was performed, and a well-differentiated fetal adenocarcinoma was diagnosed histologically. Unlike the biphasic epithelial and stromal features of pulmonary blastoma, it was composed solely of malignant glands of embryonal appearance. PMID- 10870335 TI - Hemangiopericytoma of the pleura causing massive hemothorax. AB - Hemangiopericytoma is an unusual soft tissue tumor. A 54-year-old man presented with sudden onset of chest pain and dyspnea for 1 day. The initial chest x-ray showed a massive left pleural effusion. A contrast-enhanced computed tomographic scan of the chest showed a homogenously enhanced mass in the intrathoracic extrapulmonary space. A tube thoracostomy was performed and hemothorax was confirmed. A posterolateral thoracotomy was performed and a tumor in the parietal pleura of the left chest wall was resected. Grossly, the resected tumor arose from the parietal pleura, and the cut surface was elastic, soft, and pale yellow. There were several cystic formations and hemorrhages. Based on histologic findings, hemangiopericytoma with lower grade malignancy was diagnosed. The patient was alive and free from tumor recurrence 1 year after surgery. Intrathoracic extrapulmonary hemangiopericytoma is extremely rare, and surgical excision is the treatment of choice. Adjuvant chemotherapy or radiotherapy is indicated because of the high risk of recurrence and potential malignancy. PMID- 10870336 TI - Fatal outcome of Erysipelothrix rhusiopathiae bacteremia in a patient with oropharyngeal cancer. AB - Bacteremia due to Erysipelothrix rhusiopathiae is rare; the most common presentation reported in the literature is endocarditis. We report a 32-year-old man with oropharyngeal cancer who developed aspiration pneumonia and E. rhusiopathiae bacteremia, and presented with fever, chills, dyspnea, and productive cough with purulent sputum. Despite treatment with amoxicillin/clavulanate and nutritional support for 9 days, he died of respiratory failure. He had no clinical evidence of endocarditis. He had no history of animal or occupational exposure, and might have been colonized with E. rhusiopathiae in the oral cavity, followed by aspiration pneumonia and bacteremia. A fatal outcome in a patient with bacteremia due to E. rhusiopathiae without endocarditis is rare. PMID- 10870337 TI - Pediatric endogenous endophthalmitis. AB - Pediatric endogenous endophthalmitis is a rare disease that can cause serious ophthalmic damage. We describe two cases of pediatric endogenous endophthalmitis. The first occurred in an 8-month-old boy and the second in a 7-day-old girl. These two patients had developed pneumonia due to Pseudomonas aeruginosa infection prior to the onset of ocular symptoms. The interval between the onset of pneumonia and ocular symptoms was 1 week, but endophthalmitis was diagnosed 9 days after the onset of ocular symptoms in the first case and 3 days after the onset of ocular symptoms in the second case. The ocular manifestations included eyelid swelling, purulent discharge, redness, corneal edema, hypopyon, and poor red reflex. Despite treatment with aggressive antimicrobial therapy, both patients became totally blind with eyeball atrophy. PMID- 10870338 TI - Metoclopramide-induced methemoglobinemia in an adult. AB - Methemoglobinemia is a condition characterized by increased level of methemoglobin in the erythrocytes and brownish cyanosis. Acquired methemoglobinemia is diagnosed by elevated methemoglobin with normal hemoglobin electrophoresis and normal NADH cytochrome b5 reductase. We report a patient who developed lethargy, confusion, and cyanosis during post-operative period. He had arterial methemoglobin level of 40.6% and oxygen saturation of 59%. No other cause could be found for his methemoglobinemia other than metoclopramide, even though it is rarely reported to cause methemoglobinemia in adults compared to infants. He had an excellent clinical response to treatment with methylene blue with which his clinical symptoms improved and the methemoglobin level returned to normal within 24 hours. Here we discuss the clinical features, diagnosis, and treatment of acquired methemoglobinemia induced by metoclopramide. PMID- 10870339 TI - Reduction of cholesterol in Kentucky--a state at high risk. AB - Coronary artery disease (CAD) or coronary atherosclerosis mortality rates vary markedly in the different regions of the United States. It has been observed that the states bordering the Ohio and Mississippi Rivers have a dis-proportionately high coronary heart disease death rate. Kentucky is one of the most severely afflicted states, especially the eastern portion that includes the Appalachian region. The high CAD mortality rates in Kentucky are mainly due to a high prevalence of major traditional risk factors for coronary artery disease: elevated cholesterol, heavy cigarette smoking, and hypertension. Physical inactivity is also widespread. Attempts to reverse these risk factors have often failed for multiple reasons. Primary prevention is often not given priority by individuals because they have not, by definition, experienced any cardiac symptoms. Advice by physicians regarding treatments that can promote atherosclerosis regression and secondary prevention is suboptimal. A major campaign to educate patients and physicians about the importance of lowering cholesterol, dietary counseling, exercising, smoking cessation, and other interventions is essential. The ultimate methods of coronary atherosclerosis prevention will be through interventions at the cellular and subcellular levels. While such interventions are not yet available, major risk factor control and marked cholesterol reduction can be achieved and thereby significantly decrease the personal and economic suffering caused by coronary heart disease. PMID- 10870340 TI - Privacy on the technology highways. PMID- 10870341 TI - [Dynamics in the health of women occupied in agricultural sector of the Ukraine]. AB - Work conditions of women engaged into crop-raising and animal husbandry are characterized by variable complex of occupational physical, chemical and biologic hazards caused by the production specificity. The latter results in increased prevalence of chronic somatic diseases and reproductive disorders (menstrual dysfunction, higher incidence of miscarriage, etc.). Further dynamic research based on multifactorial approach are required to study health state of rural population. PMID- 10870342 TI - [Reproductive function in women of Tajikistan (in crisis)]. AB - During the civil war the state of midwifery in Tagikistan worsened dramatically with consequent lower birth rate, higher maternal and perinatal mortality, increased neonatal morbidity. After the crisis resolution the stated parameters improved considerably. PMID- 10870343 TI - [The evaluation of hazardous cinder wastes in Ulan Ude thermal power stations]. AB - Considering possibility of waste materials use in building materials production, the authors evaluated toxicity of ash and clinker waste of electric power stations in Ulan-Ude city. The ash dust and its tincture in water, when injected even in maximal amounts, induced no intoxication symptoms and death in experimental animals, therefore bear no toxicity. Using toxicity indexes to compare ash and clinker waste dust with nontoxic dust proved the studied waste to be nontoxic and acceptable for use in building industry. PMID- 10870345 TI - [Toxic vinyl chloride related melanodermia]. AB - The case of toxic melanoderma in female patient 32 years aged is described. The erythematous and pigmented lesions are present on the face, neck and upper part of breast. The disease developed after 4 months exposure to vinilchloride in workplace. The diagnosis was made in 2 years after the beginning of the disease. PMID- 10870344 TI - [Social and medical problems of occupational health of railway workers]. AB - The article "Social and medical problems of healthcare in railway transport" presents principal factors influencing railway workers' health. The factors are those of social importance and influencing occupational suitability, general morbidity and morbidity with transitory disablement, disability in railway transport and its causes. The article shows therapeutic, sanitary and epidemiologic, social measures of prophylaxis for better work conditions and preservation of railway workers' health. PMID- 10870346 TI - [Mechanisms of homeostatic disorders induced by stress vibration injury (literature review)]. PMID- 10870347 TI - [Occupational disease services in Chuvash Republic]. AB - In 1984 an Occupational Pathology Center for 30 patients opened in Tchuvash Republic; since 1989 the Center was given a right to certify primary diagnosis of an occupational disease. Occupational morbidity structure in Tchuvash Republic corresponds to that in Russia, but the morbidity level in Tchuvash Republic is slightly lower than that in Russia. The Occupational Pathology Center in Tchuvash Republic served as an object of experiments in elaboration of occupational regulation parameters. PMID- 10870348 TI - [Parameters of blood coagulation in workers of Astrakhan gas processing plant]. AB - The workers demonstrated longer prothrombin and thrombin time showing activity of extrinsic coagulation mechanism, shorter activated partial thromboplastin time characterizing activity of intrinsic coagulation mechanism. There was no definite dependence of hemostasis parameters on age and length of service. Coagulation parameters with biochemical and hematologic studies could be used to detect hazardous influence of occupational factors on workers. PMID- 10870349 TI - [Human biosybstrates in ecologic analytical monitoring of heavy metals]. AB - The authors revealed direct correlation between higher level of some heavy metals in environmental objects, foods and the metals level in human biologic materials. Human biologic materials appeared to reflect total metals intake and to serve objective criteria of environmental pollution. PMID- 10870350 TI - [Comparative physiologic cost of work performed by pregnant vs non-pregnant women]. PMID- 10870351 TI - [Mortality among wood-processing industry workers in West Siberia]. PMID- 10870352 TI - [Occupational risk in the polyurethane ware production]. PMID- 10870353 TI - [Aspects of sanitary and hygienic work conditions of operators servicing purification installations of the printing enterprise]. PMID- 10870354 TI - [Scientific fundamentals of occupational regulation in illumination]. PMID- 10870355 TI - Competency to waive Miranda rights: clinical and legal issues. PMID- 10870356 TI - Aeromedical evacuation: remembering the past, bridging to the future. AB - Operations Desert Shield/Desert Storm saw the largest mobilization of aeromedical evacuation (AE) assets since the Vietnam War. Ultimately, more than 1,950 AE personnel were deployed to support the medical airlift of personnel. With aircrews based at 17 locations in the region, at its peak the system could move up to 3,600 intratheater and 2,500 intertheater casualties per day. Fortunately, the demand for AE fell far short of predictions. During the period from August 12, 1990, to March 31, 1991, more than 12,500 patients were successfully airlifted using converted cargo aircraft, a concept originally validated in World War II. The authors describe the Operations Desert Shield/Desert Storm AE system and identify the efforts underway to construct a new aeromedical evacuation system capable of meeting the needs of the battlefield of the 21st century. PMID- 10870357 TI - Traumatic stress disorders: a classification with implications for prevention and management. AB - The management and prevention of acute and post-traumatic stress disorders are current themes of great importance to the defense health services of many nations. Currently, between 2% and 8% of service members deployed on combat operations, United Nations peacekeeping tasks, and humanitarian and disaster relief operations present with one or more stress disorders within 3 years of deployment. The management of acute stress disorders and the prevention and management of post-traumatic stress disorders necessitate an understanding of the nosology of this group of illnesses. Research into some preventive options--such as critical incident stress debriefing--also necessitates the selection of syndrome-specific subjects during case finding if controversies about the efficacy of such interventions are to be resolved. Diagnostic features, a summary of the nosological evolution, and key points of differential treatment options are presented for 5 acute operational stress disorders (acute combat stress disorder, conversion reactions, the counter-disaster syndrome, peacekeeper's acute stress syndrome, and the Stockholm syndrome) and for 11 post-traumatic disorders, including classic post-traumatic stress disorder, chronic fatigue syndrome, Gulf War syndrome, peacekeeper's stress syndrome, survivor's guilt syndrome, and the syndrome of lifestyle and cultural change. PMID- 10870358 TI - The pediatric critical care experience at Naval Hospital Guam: suggestions for critical care training during residency. AB - OBJECTIVES: To determine the critical care experience encountered by three recently graduated military pediatricians at an overseas military hospital and present one model of maximizing allowable critical care training time during residency. METHOD: Retrospective reviews of all admissions to the special care nursery and intensive care unit at U.S. Naval Hospital Guam were performed for a 3-year and a 2-year period, respectively. Age, diagnosis, birth weight (if applicable), level of nursery care, invasive procedures performed in the nursery (endotracheal tube, umbilical artery, and umbilical venous catheter placement), patient outcome, and the need for medical transport were recorded. RESULTS: During a 3-year period, there were 122 admissions to the special care nursery (7.1% of all deliveries). In addition, pediatricians performed a total of 53 invasive procedures on these patients, and 29 infants required medical transport to an off-island neonatal intensive care unit for additional care. During a 2 year period, 70 pediatric patients were admitted to the adult intensive care unit, representing 10.2% of all intensive care unit admissions during this period. Fourteen of these patients required medical transport to an off-island referral hospital. CONCLUSION: Graduating military pediatric residents may be faced with caring for a wide range of critically ill neonatal and pediatric patients depending on their assignment. Residency training programs, with the recent increased emphasis on primary pediatric care, will need to streamline instruction in pediatric critical care to provide maximal benefit to the resident while maintaining compliance with Residency Review Committee guidelines. PMID- 10870360 TI - Treatment of acute asthma in a field environment using albuterol and a large volume spacer. PMID- 10870359 TI - Intensive care medicine in the German Field Hospital during the implementation force mission in Trogir, Croatia. AB - To provide medical care for multinational personnel deployed during the Implementation Force mission, the German Army provided a mobile surgical hospital with sophisticated equipment to perform advanced resuscitation and lifesaving surgical procedures, as well as preoperative and postoperative intensive care, under conditions similar to those in rear hospitals. During an observation period of 4 months (December 1995 to April 1996), nearly 10,000 patients were treated at this facility. In addition to the presentation of statistical data from a unique critical care facility during the Implementation Force mission, this article discusses the usefulness of neurosurgical services as part of a forward field hospital. PMID- 10870361 TI - Ketamine and oxycodone in the management of postoperative pain. AB - Relief of pain, whether post-traumatic or postoperative, is a prerequisite for the prevention of its deleterious effects on the whole organism. Unalleviated pain also increases the victim's or patient's anxiety and apprehension, which in turn increase the intensity of the pain. In the management of pain, opiates have maintained their position as the most common form of analgesic therapy despite the many side effects associated with their use. This double-blind study compared the analgesic effects of low doses of racemic ketamine and the morphine derivative oxycodone on postoperative pain after elective tonsillectomy. Also, the suitability of oxycodone for field use was evaluated with respect to ketamine. Plethysmographic pulse-wave amplitude changes were compared with the pain visual analogue scale scores as measures of postoperative pain. The results of this study did not reveal any significant differences between the analgesic potencies of the studied drugs and clearly demonstrate that even suboptimal doses of both ketamine and oxycodone can provide appreciable relief of pain. PMID- 10870362 TI - Uvulopalatopharyngoplasty versus sequential uvulopalatoplasty for surgical treatment of snoring. AB - Military personnel serving on active duty suffering from loud, bothersome snoring often require surgical treatment. This elective treatment should not disrupt the command and should have minimal impact on military readiness. Major drawbacks of the standard procedure, uvulopalatopharyngoplasty, include postoperative pain requiring convalescent leave, postoperative bleeding, and velopharyngeal incompetence. In addition, the surgery consumes limited operating room time for what many consider elective surgery. Sequential uvulopalatoplasty was developed as an alternative. This is performed with a carbon dioxide laser under local anesthesia in a clinic setting, but it requires cumbersome laser precautions and expensive laser equipment. Standard electrocautery can be used instead of a laser for this procedure. This avoids additional expense, special precautions, and equipment but is equally safe and effective. We compare time lost from work, duration and level of pain experienced, number of days until regular diet resumed, and effectiveness between uvulopalatopharyngoplasty and sequential uvulopalatoplasty. We found sequential uvulopalatoplasty to have less impact on military readiness while being as effective as uvulopalatopharyngoplasty for snoring. This makes it an ideal treatment modality for patients desiring surgical correction of snoring. PMID- 10870363 TI - The psychologist's role in the Garrison mission of combat stress control units. AB - Active duty psychologists frequently are called upon to provide services that extend beyond the model of direct patient care. Army psychologists in combat stress control teams or division mental health services, Navy psychologists deployed to surgical companies, and Air Force psychologists deployed with air transportable hospitals or mental health rapid response teams may find themselves acting as organizational consultants as well as clinicians. Psychologists assigned to hospitals and clinics also have opportunities to make contact with their units for purposes of consultation and education. Organizational consultations that offer interventions for improving unit readiness and/or increasing combat effectiveness are often welcomed by commanders and provide a mechanism for the application of training and experience directly to military populations. Transferring skills from patient care to performance enhancement may not be a clear progression for many clinicians. This article describes the strategies and materials developed as part of a combat stress control garrison mission at Fort Lewis, Washington, as an example of one approach to working with combat units. The article also calls for the development of a formal mechanism to train psychologists for such roles and for the maintenance and dissemination of research materials to support organizational interventions. PMID- 10870364 TI - A review of orthopedic injuries in three recent U.S. military conflicts. AB - We conducted a retrospective review of all patients with orthopedic injuries evacuated to a single medical center to evaluate the treatment and outcome of these injuries in three recent U.S. military conflicts: Operation Urgent Fury (Grenada), Operation Desert Shield/Storm (southwest Asia), and Operation Restore Hope (Somalia). Sixteen orthopedic casualties were originally treated at the medical detachment in Grenada before evacuation to the medical center. Most of these injuries were gunshot wounds to the extremities (11), with three known open fractures. Two patients (three extremities) sustained traumatic amputation (19% amputation rate). One hundred eighty-one patients with orthopedic injuries were medically evacuated from southeast Asia to the medical center for definitive treatment. Of these injuries, there were 143 fractures in 69 patients. One hundred of these fractures were open fractures, and 60% of these injuries were blast injuries. Furthermore, there were 26 amputations (14%). Twenty-two patients with orthopedic injuries were treated in Somalia and evacuated to the medical center. Thirteen of the 22 patients (59%) sustained gunshot wounds, and 2 (9%) sustained blast injuries. There were eight open fractures (36%) and three amputations in two patients (14%). Three of the 22 patients underwent successful limb salvage when ablation was the only other surgical alternative. It appears that a large percentage of medical center evacuations from military conflicts are for orthopedic injuries. Many of these injuries are the result of high-velocity weapons or blast injuries. Regardless of the size and/or purpose of the intervention, similar injury patterns and severity can be expected, because 51% of orthopedic patients had open fractures. Similarly, the rate of amputation associated with extremity trauma has not varied significantly since the Vietnam War. PMID- 10870365 TI - Low cholesterol and noncardiovascular mortality. AB - Recent clinical trial data have suggested that low cholesterol might cause increased mortality from noncardiovascular conditions. Several randomized trials have suggested an increase in noncoronary deaths at low levels of total cholesterol. When subcategories for causes of death were investigated, it was noted that hemorrhagic stroke risk was inversely related to total cholesterol, whereas nonhemorrhagic stroke risk was positively related to total cholesterol. Certain cancers were shown to be more common at low total cholesterol levels, namely lung, liver, lymphatic, and hematopoietic cancer. Another issue to consider is whether increased mortality rates are seen in individuals with "naturally" occurring low cholesterol or whether they are seen in individuals whose cholesterol has been deliberately lowered through dietary or drug intervention, or in both. If an association between low cholesterol and noncardiovascular mortality is present, there is continuing uncertainty regarding the mechanism by which it occurs. PMID- 10870366 TI - Sexually transmitted disease screening and reporting practices in a military medical center. AB - Sexually transmitted diseases (STDs) and their sequelae are responsible for significant human and economic costs. Military personnel are one of many core populations at increased risk for acquiring STDs. This study was designed to assess primary care physician/practitioner compliance with secondary screening recommendations and reporting practices of STDs in a military setting. Data from approximately 27,000 covered lives from the Naval Hospital and the Naval Air Station Branch Medical Clinic in Jacksonville, Florida, were used in this analysis. Because chlamydia is the most prevalent STD, laboratory results indicative of infection with chlamydia from July 1 to December 31, 1996, were used as a marker of a patient population requiring additional (secondary) STD screening. Patients with laboratory-confirmed chlamydia infection were identified using the Composite Health Care System. The medical records of these index cases were then analyzed for the presence of laboratory test results of human immunodeficiency virus (HIV), rapid plasma reagin, and hepatitis B virus (HBV) within 6 months of a positive chlamydia test. To assess compliance with mandated reporting of particular STDs, total laboratory-confirmed cases of chlamydia, syphilis, and HBV were compared with total cases reported to the Office of Preventive Medicine at the Bureau of Medicine and Surgery, U.S. Navy, during a 1 year period from July 1, 1996, to June 30, 1997. In 32% of chlamydia cases, no additional laboratory tests for HIV, syphilis, or HBV were obtained within 6 months. Fourteen percent of chlamydia cases were reported to the Office of preventive Medicine. Compliance with screening for multiple STDs after the identification of a single STD should be improved. In addition, better methods for reporting cases of STDs should be implemented. PMID- 10870367 TI - Increased regional cerebral perfusion by 99mTc hexamethyl propylene amine oxime single photon emission computed tomography in post-traumatic stress disorder. AB - OBJECTIVE: Because of the treatment resistance and chronic affective lability of many post-traumatic stress disorder (PTSD) patients and the hypothesized association of these behaviors with temporal and limbic structures, a study was conducted to determine whether these patients would exhibit alterations in regional cerebral perfusion in the temporal and limbic regions compared with age matched normal volunteers at rest. METHOD: We studied 17 patients using 99mTc hexamethyl propylene amine oxime single photon emission computed tomography. Seven of the patients were on a selective serotonin reuptake inhibitor, five were on a tricyclic antidepressant, and five were on no medication at the time of the study. Patients were compared with eight age-matched normal controls. RESULTS: All PTSD patients showed a relative increase in regional cerebral perfusion in the anterior and posterior cingulate regions bilaterally, the right temporal and parietal regions, the right caudate/putamen region, and the left orbital and hippocampal regions compared with the control group. When the group of PTSD patients who were free of medication were compared with the control group, increased regional cerebral perfusion was found in the right and left caudate/putamen regions and the right orbital and anterior cingulate cortex bilaterally. CONCLUSIONS: PTSD is associated with increased regional blood flow in limbic areas and the right temporal and parietal cortex compared with age matched normal volunteers. PMID- 10870368 TI - Adaptation of immigrant psychiatrists from the former Soviet Union in the Department of Mental Health of the Israel Defense Forces. AB - Psychiatrists from the former Soviet Union serve in the Department of Mental Health of the Israel Defense Forces. The new immigrant psychiatrists confront a wide range of difficulties during the process of integration to the military system and adaptation to the specifically military aspects of psychiatry. These include unfamiliarity with the military system, cultural clashes with the different groups of soldiers representing the various subgroups of the absorbing society, the psychopathology of soldiers, which is different from that seen in civil psychiatry, and the change in focus in the military mental health service, which emphasizes the importance of evaluating ego strength. Arbitrarily, one can describe four stages of adaptation that the immigrant psychiatrist has to pass through before recruitment and during service until adaptation and integration in the new role take place. Individual and group supervision are the main means by which the assimilation process is eased. The military service smooth the acculturation process and has an important role in helping the immigrant's adaptation to Israeli society and in building his or her professional identity. PMID- 10870369 TI - Development of a medical supply set for corpsmen in the field. AB - Fleet Marine Force corpsmen are the first medical responders to treat casualties in the field. They carry an outdated bag of supplies called the surgical instrument and supply set. The purpose of this investigation is to develop an updated supply set for field corpsmen by linking each supply item to specific medical tasks conducted in the field, which then creates an audit trail. The review of medical supplies generated an updated list of supplies to be carried by corpsmen in a new medical module and a list of items that corpsmen can pull from the battalion aid station authorized medical allowance lists as needed. Items without a clinical requirement were not included. This improved set of supplies for corpsmen will greatly enhance treatment capability in the field. As technology and needs change, replacements, additions, and deletions of the items can easily be made. PMID- 10870370 TI - Antiphospholipid antibodies, ischemic stroke in young adults, and calcium supplementation: a hypothesis. AB - The proper treatment of younger patients who have suffered ischemic stroke and who have no stroke risk factors other than antiphospholipid antibodies is unsettled. We propose a rationale to support adding dietary supplementation with calcium and vitamin D to the present standard regimen of anticoagulant therapy for these patients. We expect that the benefits from this additional therapy will prove additive. Proving this hypothesis will require large numbers of patients unlikely to present to any one center. The military health care system is well suited to such a study. PMID- 10870371 TI - Bladder entrapment after external fixation of traumatic pubic diastasis: importance of follow-up computed tomography in establishing prompt diagnosis. AB - A 30-year-old male was an unrestrained driver in a high-speed motor vehicle crash. On presentation, the patient was profoundly hypotensive with multiple injuries, including a 20-cm-deep perineal laceration with avulsion of the rectum, a diffusely tender abdomen, an unstable open-book pelvic fracture, and multiple rib fractures. Blood noted at the urethral meatus prompted a retrograde urethrogram and cystogram, which were within normal limits. A Foley catheter was placed with the return of clear urine. Closed reduction and external fixation of the pelvic fracture were performed emergently without difficulty. Postoperative computed tomography of the abdomen and pelvis revealed a retrovesical pelvic hematoma and entrapment of the bladder in the reduced pubic symphysis diastasis. Lower abdominal exploration revealed an intact bladder without evidence of gross bladder wall injury. On release of the external fixator, the bladder was easily reduced into the normal retropubic location. Definitive internal fixation of the pubic diastasis was performed. No urologic sequelae were noted postoperatively. PMID- 10870372 TI - Central pontine myelinolysis: association with parenteral magnesium administration. AB - A 29-year-old woman with diabetes mellitus and nephrotic syndrome was given 30 g of magnesium sulfate over 14 hours after a cesarian section. Her serum magnesium level increased to 7.4 mg/dl. Five days later, she became quadriplegic with inability to speak or swallow. Cranial magnetic resonance imaging demonstrated central pontine myelinolysis (CPM). Initial serum sodium was not measured. Although CPM is usually associated with a rapid increase in serum osmolality, most patients who experience a rapid increase in serum osmolality do not develop the clinical syndrome of CPM. Consequently, additional factors may also be important in the pathogenesis of CPM. Parenteral magnesium administration may be a potential contributing factor in the pathogenesis of some cases of CPM. PMID- 10870373 TI - Global improvement in intellectual and neuropsychological functioning after removal of a suprasellar cystic craniopharyngioma. AB - This is a case report of a patient who presented with cognitive and behavioral decline and underwent surgery for removal of a suprasellar craniopharyngioma. Previous studies have reported memory impairments in the presence of craniopharyngioma, but information is lacking regarding the impact of craniopharyngioma on other brain functions as well as on intellectual abilities. In this study, comprehensive neuropsychological testing, including intellectual testing, was conducted 9 to 14 days before surgery and 16 to 22 days after surgery. Results before surgery demonstrated average intellectual abilities, which were decreased from premorbid estimates, and diffuse neuropsychological impairments. Postoperatively, significant improvement in both intellectual abilities and neuropsychological test scores was observed. These results suggest that craniopharyngiomas can be associated with impairments in intellectual abilities and in brain functions aside from memory. The results are contrasted with those of previous reports, and implications for future research are discussed. PMID- 10870374 TI - Neuropsychological sequelae of heat stroke: report of three cases and discussion. AB - Heat stroke includes neurologic impairment as a person's body temperature reaches 40.5 degrees C (105 degrees F) as a result of a failure of thermoregulation. The physiologic complications of heat stroke are well described in the literature. The domains of cognitive functioning affected and the severity of impairment resulting from heat stroke can vary from mild deficits in attention and memory to severe global dementia. There can also be changes in affect and personality that are equally debilitating. This article presents the neuropsychological test results of three active duty soldiers who suffered heat stroke and were tested within 2.5 months of injury. PMID- 10870375 TI - Sinus bradycardia secondary to mitoxantrone. PMID- 10870376 TI - [Treatment errors and sequelae. The physician's image]. PMID- 10870377 TI - [Public health portals for laity. Gold diggers or public health educators]. PMID- 10870378 TI - [Neurolysis per enzyme probe. Placebo for the intervertebral disk?. Interview by Bodo Dorra]. PMID- 10870379 TI - [Heart failure in the elderly. Characteristics of diagnosis and therapy]. AB - The incidence and prevalence of cardiac insufficiency increases dramatically in the elderly. The reason for this is the more frequent cardiac disease in this age group, the functional effects of which compound the sequence of the physiological aging and exogenous noxae. The clinical presentation of heart failure in the elderly may be atypical, echocardiography should always be performed to confirm the diagnosis and obtain a better evaluation of the underlying condition, before therapeutic measures are initiated. For, in the elderly in particular, treatment must be planned on an individual basis. Primarily, reversible cardiac or extracardiac disorders, potential triggers of cardiac insufficiency, must be identified and treated. All the data currently available indicate that in the elderly, too, all the established options for specific treatment should be utilized. While the reservations often applied to the use of drugs to treat chronic cardiac insufficiency--in particular the treatment of acute myocardiac infarction by thrombolysis or acute intervention--and based on a fear of iatrogenic complications, are understandable, but to date do not justify such a stance. Although contraindications in the elderly are more common and complications not always avoidable, old patients can also benefit from these therapeutic measures. PMID- 10870380 TI - [Therapy of heart failure. General practice references for individual treatment]. AB - Cardiac insufficiency is today the most common reason for hospitalizing patients older than 65 years, and is associated with considerable costs for the health care system. The economic consequences (drug costs, hospitalization and costs of early retirement) necessitate close cooperation between family doctor and cardiologist if a rational (evidence based medicine) and cost-optimized treatment is to be achieved. The implementation of the current therapeutic concepts and guidelines issued by the specialist societies has so far been inadequate, in particular due to a misguided approach to budgeting, and the pressure of rising costs. Few diseases are as accessible to a differentiated drug treatment as chronic heart failure. Numerous studies have shown that an optimally adapted and adequately dosed treatment with ACE inhibitors, beta blockers, diuretics, aldosterone antagonists and digitalis is capable, not only of lowering the mortality rate, but also of achieving a significant reduction in morbidity (improved quality of life, shorter hospitalization). The treatment of the chronic forms of cardiac insufficiency is a domain of ambulatory patient care that, with an eye to the multimorbidity of the elderly, requires individually adapted pharmacotherapy. PMID- 10870381 TI - [Therapy study of acute exacerbated chronic bronchitis. Quinolone rapidly improves cough and chest pain. BRONCHIMOX Study Group]. PMID- 10870382 TI - [Gastritis with or without Helicobacter pylori. Proper therapy disclosed by the pathologist]. PMID- 10870383 TI - [Chronic headache. Differential diagnosis and therapy]. PMID- 10870384 TI - [Acupuncture--indications and risks. Complementary medicine treatment, 2]. PMID- 10870385 TI - [Travel pharmaceuticals for the physician. 3: Additional drugs for diving and trekking]. PMID- 10870386 TI - [Epigastric pain with ECG changes. Unstable angina pectoris with intramural anterior wall infarct]. PMID- 10870387 TI - [Endothelium research. ACE inhibitor with high vascular affinity]. PMID- 10870388 TI - [Chronic inflammatory bowel diseases. Do antihistaminics reduce the need for steroids?]. PMID- 10870389 TI - [Lowering blood pressure, smoking cessation, thrombocyte inhibition. What is new in stroke prevention?]. PMID- 10870390 TI - [Fast prophylaxis for last-minute travelers. Which measures are still possible 1 week before traveling?]. AB - Vaccinations or booster injections against tetanus, diphtheria and polio shortly before leaving on a journey are both possible and to be recommended. Active hepatitis-A-vaccination can also be applied immediately prior to the journey, and offers better protection than gamma globulins. As a rule, vaccinations against hepatitis B, yellow fever and typhoid must be given one to four weeks before the journey. Effective malaria prophylaxis for last-minute travellers is always possible. In addition to mandatory "exposure prevention", effective chemoprophylaxis is also recommended for travellers to tropical Africa. The dose of the first week should, whenever possible, be taken prior to the start of the journey. PMID- 10870391 TI - [When travel joy winds up in the pants... Prevention, differential diagnosis and therapy of travel diarrhea]. AB - Diarrhea is the most common symptom experienced during and after a visit to a tropical or subtropical region. It is estimated that one-third of all travellers abroad contract diarrhea. The most frequent causes are infectious pathogens that, under the circumstance of poor sanitation, are picked up via the fecal-oral route. As a preventive measure, therefore, recommendations regarding food hygiene should be strictly observed. The most important therapeutic measure is replacement of lost fluid and electrolytes. In the event of bloody stools and fever, suitable antimicrobial treatment should be instituted. PMID- 10870392 TI - [Poisonous animals at bathing beaches]. AB - Tourists and native inhabitants of tropical and subtropical regions differ significantly with regard to the risk and nature of incidents involving venomous and poisonous animals. While the indigenous population encounters such risks daily during work and other activities, tourists are usually endangered while swimming or diving, or by ingesting toxin-containing fish and/or other seafood. Whether abroad or at home, allergic reactions to the stings of bees, wasps and hornets are probably the most common manifestations of an encounter with a "poisonous animal". Travellers should be well acquainted with the dangers entailed in encountering or ingesting a venomous or poisonous animal--prevention is the most important measure. PMID- 10870393 TI - [Prevention of malaria. Drugs, side effects and practical tips for use]. AB - The malaria risk for travellers to West or East Africa is roughly 2-4% per month. In Germany, the mortality rate, in comparison with other countries, is a relatively high 2.4%. This means that in the absence of prophylactic measures, 4 8 out of 10,000 travellers will die. The risk of contracting malaria in Asia and South America is lower. Travellers to regions in which malaria is endemic, should take advantage of the effective prophylactic measures available. The two most important preventive strategies are: avoidance of mosquito bites (exposure prophylaxis) and the eradication of plasmodia that have already invaded the body. Currently, no antimalarial vaccination is available. PMID- 10870394 TI - [Risk for mother and child. Hypertension during pregnancy]. PMID- 10870395 TI - [Cardiovascular risk is correlated with serum magnesium. Recommendations for diagnosis of magnesium deficiency]. PMID- 10870396 TI - [Aids for self care. Diagnostic series: Self determination of blood coagulation values]. PMID- 10870397 TI - [Collapsing polyps in the large intestine. Pneumatosis cystoides intestinalis]. PMID- 10870399 TI - [Atopic dermatitis in children. Prevention can reduce the incidence of asthma]. PMID- 10870398 TI - [Crime in public health. White coat and dirty hands]. PMID- 10870400 TI - [Androgenetic alopecia. Denser hair growth on two-thirds of balding scalps]. PMID- 10870402 TI - Births, marriages, divorces, and deaths: provisional data for June 1999. PMID- 10870401 TI - [Type 2 diabetic patients. Aggressive management of increased blood lipids]. PMID- 10870403 TI - State society testifies on market dominance. PMID- 10870404 TI - Campbell Bill's advance is victory for patients and physicians. PMID- 10870405 TI - What's new on the hill. PMID- 10870406 TI - Statewide patient education campaign on generic drugs. PMID- 10870407 TI - HIV confidentiality law has serious flaws. PMID- 10870408 TI - Bemused by medical error controversy. PMID- 10870409 TI - Don't crumple the paperless office. PMID- 10870410 TI - A potpourri of drug news. PMID- 10870411 TI - Perspectives on health care quality. PMID- 10870412 TI - [Parathyroid cancer]. AB - Parathyroid cancer is a rare disease, causing 0.5% to 5.2% of primary hyperparathyroidism cases. Hyperparathyroidism accompanying a cancer is usually more severe with richer clinical symptomatology then hyperparathyroidism in the course of benign lesions. Every palpable neck tumour with high blood plasma calcium level should suggest the presence of parathyroid cancer. Development of cancer is slow but life prognosis depends on the extent of the first surgery. The prognosis is most favourable in case of 'en bloc' resection of tumour during the first surgical procedure. Therefore, diagnosis of cancer before operation is very important. If the surgery fails, the treatment should be aimed at lowering hypercalcaemia, which is the most common cause of the fatal sequel. PMID- 10870413 TI - [The lateral extraperitoneal access in surgery of the adrenal glands]. AB - This study aimed at evaluating lateral extraperitoneal access to adrenals in 233 patients (103 patients with pheochromocytoma, 101 with Conn syndrome, 11 with Cushing syndrome and 18 patients with hormonally inactive tumours) who underwent surgery in our department within 1981-1998. The age of patients ranged from 10 to 84 years. Biochemical tests have been performed and hormones assayed in all of them prior to surgery, and the localization of tumours has been established by means of the abdominal cavity ultrasound examination and CT scans. Two hundred thirty five tumours have been excised. The mean duration of surgery was 91 minutes and the mean time of postoperative hospital stay--5 days. Postoperative complications have been noted in 4.7% of the operated patients but in the group with pheochromocytoma their incidence increased to 8.7%. In 13.7% of cases an accidental opening of pleural cavity was noted. The majority of cases has had drain left in pleural cavity whereas the remaining patients have had pleurae sewn. There were two hospital deaths in the group of pheochromocytoma, i.e. in 0.85% of all patients. These results and complications were analysed in a discussion. It seems that this kind of surgical access to adrenals can be successfully used in most unilateral adrenal pathologies and in cases of extra adrenal tumours localized along abdominal aorta. Patients with pheochromocytoma are still in increased surgical risk. PMID- 10870414 TI - [Diagnostic significance of cancer procoagulant activity in colorectal cancer]. AB - This study aimed at evaluating diagnostic significance of cancer procoagulant (CP) activity in the homogenates of colon cancer tissues and in blood serum of patients with this neoplasm. Procoagulant activity, depending of specific cancer procoagulant, has been found in all examined tissues as well as in blood serum of cancer patients. CP activity in homogenates of colon cancer tissues as well as in blood serum of the examined cancer patients has been markedly higher than in the normal subjects. These data indicate that CP activity in the neoplastic tissue homogenates and in blood serum may be of value in the diagnosis of cancer. PMID- 10870415 TI - [Diagnostic value of some cytokines levels in chronic myelogenous leukemia]. AB - Some cytokines play an important role in aetiology and pathogenesis of leukaemia. Majority of data available in the literature relates to a single cytokine in the cell culture in vitro. In patients with chronic myelogenous leukaemia several different cytokines are probably active. Therefore, this study aimed to determine the concentrations of interleukin-1 beta, G-CSF, L-selectin in leukaemia cell culture supernatant, broken granulocytes and plasma in the course of disease exacerbation and remission. Cytokines have been assayed with available immunoenzymatic kits of ELISA type. IL-1 beta, G-CSF and L-selectin levels have been increased in cell supernatant but IL-1 beta level has been decreased in non stimulated broken granulocytes. G-CSF levels have been low both in stimulated and non-stimulated granulocytes. In comparison to control the lowered levels of G-CSF and L-selectin have been observed in blood plasma of patients with CML. Different levels of assayed cytokines and adhesion molecule may suggest their contribution to leukaemia cells proliferation regulation. PMID- 10870416 TI - [Ultrasonographic assessment in differential diagnosis of non-palpable breast tumors]. AB - In this study we analyse the value of high frequency ultrasound as diagnostic tool in non-palpable breast nodules (NPBN). We selected 121 women with NPBN revealed by ultrasound (US). The size of the nodules varied from 3 mm to 32 mm. Women with NPBN detected by US underwent sonographically-guided fine-needle aspiration biopsy (FNAB) and/or histological examination. Based on the US examination findings, the disease was diagnosed as malignant in 6 cases, as benign in 90, and in 28 cases it was unclassified. 15 patients underwent surgery. In 8, the disease was histopathologically confirmed as carcinoma (US 2 unclassified), 7 were benign lesions. US with FNAB as tool in diagnosing NPBN is an accurate technique for distinguishing between benign and malignant NPBN observable sonographically. PMID- 10870417 TI - [Primary management of facial skeleton injuries in patients treated at the maxillofacial surgery ward]. AB - This study aimed at discussing 278 patients with facial skull fractures and injuries, being managed at the Maxillofacial Surgery Ward of the Voivodeship Specialist Hospital in Rzeszow within 1991-1998. There have been 201 male, 65 female and 12 paediatric patients, aged between 6 and 67 years. The majority of injures in 161 (57.9%) patients required simultaneous treatment of fractures and soft tissue lesions. 68 (24.6%) patients have had closed injuries and face wounds. In 49 (17.6%) patients reposition and immobilisation of fractures have been necessary. In most cases injures have been dressed at emergency room. Fractures of facial skull, coexisting with face injures and wounds, often required specialist treatment provided by the maxillofacial surgeon, laryngologist and ophthalmologist, combining reposition and immobilisation of fracture with the wire, Nichrominox plates and Champy miniplates, followed by wound dressing, according to the commonly approved rules of the primary medical care. PMID- 10870418 TI - [Multiple hepatic hemangiomas]. AB - Hepatic hemangiomas are the most common benign tumours of the liver and the second most common of all hepatic tumours. Most often they are asymptomatic, although sometimes are the cause of severe complications. In this report we present female patient with multiple hepatic hemangiomas who underwent surgery for hemobilia. A several years long asymptomatic postoperative period is remarkable. PMID- 10870420 TI - [Two cases of the giant ovarian cysts treated surgically]. AB - The author presents two cases of the giant ovarian cysts, treated surgically. Diagnosis was based on the clinical examination and ultrasonography. Very large size suggested a malignant character but histological examination has shown that both cysts were benign. PMID- 10870419 TI - [Obstruction of the small intestine caused by malignant lymphoma]. AB - Primary malignant tumors of the small intestine are a rare finding, not thought to be a likely case of intestinal obstruction. A case is reported of a patient presenting with chronic abdominal pain, apparently due to adhesions and a postoperative hernia (already after its repair). On laparotomy two tumors of the small, intestine were found obstructing its lumen, diagnosed histopathologically as primary malignant lymphoma. PMID- 10870421 TI - [Isolated pulmonary eosinophilic granuloma]. AB - One clinical case of pulmonary eosinophilic granuloma were followed-up during 10 months of chemotherapy. Clinical and radiological symptoms of disease were described. Series of 4 HRCT examinations revealed improvement of pulmonary changes. PMID- 10870422 TI - [Hepatic hemangioma: review of diagnostic methods]. AB - Hepatic hemangiomas are the most common benign tumours of the liver. In most cases they are asymptomatic and are being detected accidentally. Then, differential diagnosis excluding other hepatic focal lesions and follow-up are required. In this paper we survey different methods of diagnosis and assessment of the hepatic hemangiomas. PMID- 10870423 TI - [Monitoring of bone metastases]. AB - Current knowledge about an incidence of bone metastases, use of bisphosphonates and assessment of response to the treatment are surveyed. The bone metastases are quite frequent in patients with breast and prostate cancers. High doses of intravenous pamidronate are particularly useful in the treatment of these patients. Prior to therapy with bisphosphonate special score elaborated by RE Coleman should be calculated. The new biochemical bone resorption markers especially Ntx i Crosslaps seems to be the most efficient for evaluation metastases response. PMID- 10870424 TI - [Contemporary opinions on the role and properties of L-ascorbic acid]. AB - There is growing interest in the role of vitamin C physiological role. In this paper L-ascorbic acid is discussed in detail, including its physical properties, chemical activity and biological properties. The primary goal of this review is to present the chemical characteristics of this compound and to discuss its relation to various biological functions of vitamin C, mainly as a free radical scavenger. PMID- 10870425 TI - [The achievement of Warsowian physicians in otolaryngology after introduction of laryngoscopy]. AB - The professional and scientific activities of Konstanty Karwowski (1834-1918), one of the pioneers of the laryngoscopy in Warsaw are described. The widespread contacts of Ignacy Baranowski (1833-1919) with the Viennese scientific medical world are stressed. The beginning of otological lectures by Wiktor Szokalski (1811-1897) in 1864, the famous Polish ophtalmologist is mentioned. The achievements of eminent Polish surgeons: Aleksander A. Le Brun (1803-1868), Hipolit Korzeniowski (1827-1897), Polikarp Girsztowt (1827-1877) and Julian Kosinski (1833-1914) from surgical clinics in Warsaw are presented. The first total laryngectomy in Poland, performed by Kosinski in 1877 is presented in detail. PMID- 10870426 TI - Sex differences in strategy and performance on computerized neuropsychological tests as related to gender identity and age at puberty. AB - Neuropsychological sex differences have since long been under debate. Support for the relation between behavioral differences and biological variables like hormone influence is, however, emerging. Sixteen men and sixteen women, all university students, were tested with computerized neuropsychological tests (APT), the Bem Sexual Role Inventory, and asked about pubertal age. The results were in line with earlier findings of sex differences in neuropsychological tests, men being faster and women more cautious. The assumption that women tend to use left hemispheric, verbal/serial strategies also in spatial tasks was also partly supported. In women, late onset of puberty was related to better spatial performance, and there were also more intercorrelations between verbal and spatial tests in the female than in the male group, indicating that women use less specific strategies (more g-factor intelligence) in problem solving, or that aptitudes are less compartmentalized in women than in men. PMID- 10870427 TI - More than one way to change: a study of course heterogeneity during and after short-term psychiatric in-patient treatment. AB - This study describes differences in course and outcome, defined by GSI (SCL-90) at admission, discharge, and one-year follow-up, in 458 patients receiving in patient treatment for long-standing symptom and/or personality disorders. A K mean cluster analysis identified seven subgroups of patients, representing four clinical distinct, meaningful patterns of change: early improvement, late improvement, relapsing after discharge, and a severe chronic course. MAIN FINDINGS: the subgroups had unique correlates among socio-demographic, diagnostic, and treatment-related characteristics. One of the relapsing groups had a high rate of Cluster C personality disorders, whereas the other had low participation in the anxiety programme. The group with severe chronic course showed occupational maladjustment and high number of both Axis I and II disorders. IMPLICATIONS: anxiety patients should participate in anxiety-treatment programmes, Cluster C patients should be followed and monitored for relapse, and severe chronic patients should be offered specialised treatment for their co existing substance abuse and/or eating disorders. PMID- 10870428 TI - Parental divorce: long-term effects on mental health, family relations and adult sexual behavior. AB - Specific long term effects of parental divorce were examined in a sample of 179 Icelanders, 20 to 30 years of age. The participants answered the Borromean Family Index, the Affect Balance Scale and a number of questions on sexual behavior and attitudes towards marriage and divorce. Results showed that compared to adults whose parents remained married, those of divorced parents reported more negative emotional experiences at the time of the study and had looser family ties. They also had greater number of short love affairs, had their first love affair at a younger age, had a greater number of sexual partners, and were younger at the time of their first sexual intercourse than adults whose parents remained married. PMID- 10870429 TI - The association between implicit theories of personality and the attributional process. AB - The influence of implicit theories of personality (entity vs. incremental theorists; see Dweck, Chiu and Hong, 1995) on the stages of the Sequential Operations Model of attribution (Gilbert, Pelham, and Krull, 1988) was investigated. Two hundred eighty Norwegian participants were given a Norwegian translation of the implicit personality theories measure. Participants then read two essays, one advocating the pro-life position and the other advocating the pro choice position on the abortion issue. The essay positions were ostensibly assigned rather than freely chosen by the author. After each essay, participants were asked to rate the essay position and the true attitude of the author. Entity and incremental theorists showed no differences in their ratings of the essay position; however, entity theorists made significantly stronger correspondent inferences about the author's attitude than did incremental theorists. These results support the contention that entity theorists engage in less attributional correction than incremental theorists. PMID- 10870430 TI - Comprehensive driving assessment: neuropsychological testing and on-road evaluation of brain injured patients. AB - The study investigates the correspondence between neuropsychological test results and on-road driving performance among 55 patients with a CT-verified brain damage or documented neurological disorder (cerebrovascular accident: 43, traumatic brain injury: 5, multiple sclerosis: 4, other: 3). 5 patients showed unimpaired test profiles and passed the on-road evaluation. 18 patients showed severe neuropsychological deficits contrary to driving and were not recommended for on road evaluation. Of the remaining 32 patients with some neuropsychological deficits, all 100% in the minor impaired group (n = 8) passed the driving evaluation, compared to 69% in the mildly impaired (n = 16) and 38% in the moderately impaired group (n = 8). Measures of reduced visuoconstructive ability, reaction time, visual attention, and awareness of cognitive impairments, were found to discriminate between groups. It is concluded that neuropsychological assessment of targeted functions provide an ecological valid prediction of driving skill after brain damage, but that on-road evaluation is needed as supplement in cases with ambiguous test findings. PMID- 10870431 TI - Cross-modality priming for individual words in memory for coherent texts. AB - This study explores implicit memory within the domain of text processing. Three experiments were designed to study cross-modality priming in a word-stem completion test following presentation of target words in the context of a coherent text. Four main results emerged. First, we found a significant priming effect for words previously studied in a text, this priming is higher with low frequency words than with high-frequency words. Second, subjects demonstrated more repetition priming when study and test modalities matched than when they were different. Third, the magnitude of the priming effect in the visual condition varied with the perceptual processing of the text read. Fourth, priming effects did not depend on subjects' remembering of the words of text read as measured by a yes/no recognition test since no modality effect was found in this latter memory test. These results challenge Levy's (1993) view and are discussed in the framework of the transfer-appropriate processing view proposed by Roediger, Weldon and Challis (1989). PMID- 10870432 TI - Communicative development in Swedish children 16-28 months old: the Swedish early communicative development inventory--words and sentences. AB - To describe the development of words and sentences in Swedish children 16-28 months old, 900 parental reports on 336 children were analyzed. Subjects were randomly selected from the national birth register, and there was a response rate of 88%. The assessments were made using the Swedish Early Communicative Development Inventory--words and sentences (SECDI--w&s). Age-based norms for productive vocabulary, pragmatic skills, grammar skills, and maximum length of utterance (MaxLU) were determined. We describe the development of feedback morphemes, semantic categories, and single words and tasks. Correlation across measures was significant, and especially strong between vocabulary size and grammar skills. Optimized positive predictive values were high for 25 to 28 month predictions (71%-88%), and vocabulary scores were found to be of particular predictive importance. No significant gender differences were detected. The clinical relevance of the instrument is discussed. PMID- 10870433 TI - Individual postdecision processes in group settings. AB - In two experiments Differentiation and Consolidation Theory (Diff Con) (Svenson, 1992) was used to investigate individual postdecision making processes in three member groups. It was predicted that in groups in which the subjects preferred different alternatives (conflict groups), subjects would consolidate their own preferred alternative, and not the group's final decision. A second hypothesis was that no consolidation would be indicated in groups in which all members preferred the same alternative (non-conflict groups). The results showed that in conflict groups, the members who gave up their preferred alternative (minority members) consolidated their own preference, thereby significantly regretting the group decision. In contrast, members who got their own will through in the majority decision (majority members) showed no consolidation of the group decision. The corresponding pattern of results was replicated in a second experiment, using a different decision situation. The results indicated that perceptions of social support, agreement in a group and decreasing responsibility for a group's decision, could all partly substitute consolidation by attractiveness restructuring. PMID- 10870434 TI - Postdecision consolidation in a trend prediction task. AB - To be able to learn from experience it is necessary to correctly apprehend experienced feedback and the situation in which it is provided. The results indicate how post-decision consolidation in complex domains may affect learning. The problem may be particularly pertinent in recurrent decision making where considerable risk is involved. The study explores the changes in aspect (signal) importance from pre- to postdiction as a function of outcome information. By postdiction we mean the remembering of an earlier prediction (cf. Hawkins & Hastie, 1990). Subjects were asked to decide on which of four alternative future price developments would follow a historical price trajectory for different commodities, and to rate the importance of each of the chosen alternative's corresponding aspects. The subjects revealed a bias in their support ratings of aspects--seeing support in aspects that traditionally (by themselves and in many contexts) would be seen as neutral or even counter-indicative of the alternative chosen. After an intermission, the subjects were also given information about what was indicated to be the actual development of the market. One group was told that their decisions were correct (irrespective of what the decisions were), another group that they were incorrect but close, a third group that they were incorrect by far, while a fourth group served as a control. Following this information the subjects were again asked to judge the importance of the aspects for their own prior decision on the most likely future development. The results indicated that outcome feed-back had an effect on post decision restructuring of facts. Subjects in the correct condition showed an average consolidation that increased the support, while the wrong conditions lead to negative consolidation (in retrospect indicating that they never found as much support for their decision in the past as they actually did). Thus, in a choice between consolidating their own initial prediction and the price trajectory they would have to live with, the decision makers consolidated the outcome. Therefore, the results of the study were related to the hindsight bias phenomenon (Fischhoff, 1975) and to Kahneman and Miller's (1986) mutability concept. PMID- 10870435 TI - Specifying factors in radiation risk perception. AB - This is a study of risk perception and a test of a model of perceived risk. A scale measuring fear was factor analyzed and the resulting five fear factors were related to a large number of risk dimensions, both personal and general. Fear was only rather weakly related to perceived risk. Furthermore, perceived risk of two ionizing radiation hazards (nuclear power, X-rays) were investigated in more detail. These risk ratings were modelled on the basis of attitude, risk sensitivity, specific perceived risk of radiation, trust and an extended version of the traditional Psychometric Model, enhanced by the introduction of a factor of Tampering with Nature. It was found that risk perception could be well explained with this approach and the importance of Tampering with Nature, as well as specific perceived risk, were stressed. PMID- 10870436 TI - A gender perspective on Person-Manager fit and managerial advancement. AB - This article presents two studies examining (1) the relationship between Person Manager (P-M) fit and managerial advancement of women and men with, and without managerial aspirations and (2) the P-M fit as related to managerial and non managerial women. The P-M fit was assessed by computing the congruence between participants' self-rated personality profile and the perceived personality profile of a manager. Sex (men show a higher P-M fit than women), gender (the higher the individual's masculine gender-role, the higher the P-M fit) and group (managers and managerial aspirants show a higher P-M fit than non-managerial aspirants and non-managers) hypotheses were tested. There was no support for the sex difference hypothesis. However, the group and gender hypotheses were confirmed showing that managers and managerial aspirants had a higher P-M fit than non-managers and non-aspirants. Further, analyses revealed that the higher the participants' masculinity scores, the higher the P-M fit. Implications of these findings are discussed in relation to the gendered image of the managerial role and adaptation theory. PMID- 10870437 TI - [Experience in organizing the surgical work of a garrison hospital in an armed conflict]. AB - The authors have summarized organizational experience of surgical work of garrison military hospital strengthened with specialized brigades during the period of armed conflict in Republic of Dagestan (August-September, 1999). From the start of active actions in order to render assistance specialized surgical teams from district military hospital equipped with special kits (at the rate of 7 operations/day during a week) were sent to garrison hospital. In this armed conflict there are features characterising both mine-and-explosive war in Afghanistan and sniper war in Chechen Republic resulting in increase in the number of seriously wounded (up to 46.7%) casualties during Botlikhskii operation constituted 1:4, Novolakskii (Kadarskii)--1:5. Bullet injuries were fatal in 49.4% of the cases, fragmentation (including MET)--50.6%. During 1.5 month of hospital work there were performed 303 surgical interventions. 22.7% of slightly wounded from local garrisons were treated in garrison hospitals. Treatment results--postoperative lethality in gunshot trauma at the given stage constituted 1.1%. PMID- 10870438 TI - [The trends and outlook in the development of mobile military field medical unit formations]. PMID- 10870439 TI - [Specialized diagnosis at the polyclinic stage]. PMID- 10870440 TI - [From experience in delivering sociomedical care to war veterans and to the Armed Forces]. PMID- 10870441 TI - [The training of dermatovenereologists in a medical staff internship]. PMID- 10870442 TI - [The ethical-psychological and hygienic aspects in the cleanup of the aftermath of catastrophes]. PMID- 10870443 TI - [Food additives in the prevention of iron-deficiency anemia]. PMID- 10870444 TI - [Surgical care for the wounded in an armed conflict: the problems and the ways for their improvement (4)]. PMID- 10870445 TI - [Experience with the use of the phacoemulsification method in traumatic and senile cataracts]. PMID- 10870446 TI - [Mucociliary transport in patients with lower respiratory tract infections]. PMID- 10870447 TI - [Primarily detected lymphogranulomatosis]. PMID- 10870448 TI - [The efficacy of reflexotherapy methods in the rehabilitation of servicemen with the sequelae of closed craniocerebral trauma]. PMID- 10870449 TI - [The medical rehabilitation of servicemen following aortocoronary bypass]. AB - The results of aortocoronary shunting (ACSh) were studied in 243 patients after myocardial infarction at different stages of rehabilitation--in rehabilitative hospital and military sanatorium. Result assessment at the final stage of therapy in sanatorium confirmed ACSh efficiency in overwhelming majority of patients. At the same time principal peculiarities were revealed showing the necessity of differential approach to determination of volume and place for conduction of restorative therapy. Process of successful adaptation in most patients is achieved during the periods up to 8 weeks that corresponds to two-staged rehabilitative system. More long periods of therapy are required for patients having complications not arrested at hospital stage, low exercise tolerance, poor psychological readaptation. Optimization of rehabilitative system in order to determine the volume and place of rehabilitation will contribute to further improvement of staged therapy in such patient group. PMID- 10870450 TI - [The new possibilities of automated diagnosis and millimeter-wave therapy]. PMID- 10870451 TI - [An algorithm for the reconstruction of irradiation doses based on computational and instrumental methods]. PMID- 10870452 TI - [The health of the command officer staff doing their service in Polar regions]. AB - Initial referral (not considering dental diseases) for the last 4 years was analyzed in servicemen from the Northern Fleet, group of navy specialists serving in Headquarters was investigated. Anamnesis and central hemodynamic parameters were studied in casual sample (388 senior officers), special questionnaires were used to determine intensity of service activity. According to the data obtained during the last 4 years level of morbidity, hospitalization and disability (without dental diseases) in servicemen constituted 857.01%, in Headquarters senior officers it was 488.77%, hospitalization level was 40.14% and 18.05% respectively; disability level constituted 259.77% and 127.06%. Distribution of persons investigated in the groups according to category of working intensity was the following: the 1st (low working intensity)--0%, the 2nd (middle working intensity)--35.6%, the 3rd (intensive work)--61.3%, the 4th (high working intensity)--3.1%. Good health level was established in the group of senior officer staff. The results obtained allow to conclude that in conditions of Kol'sky Zapolarie morbidity can be decreased approximately in 1.75 times. PMID- 10870453 TI - [The optimization of antibacterial therapy in medical and prophylactic institutions]. PMID- 10870454 TI - [The rights of citizens while being delivered psychiatric care]. PMID- 10870455 TI - [The date of the creation of the main administrative organ of military medical affairs in Russia]. PMID- 10870456 TI - [The fall of 1941, Kalinin]. PMID- 10870457 TI - [The 115th anniversary of the Krasnoyarsk Military Garrison Hospital]. PMID- 10870459 TI - Health tips. New drug labels. PMID- 10870458 TI - Anesthesia. Safer than ever before. PMID- 10870460 TI - Study finds that centenarians tolerate surgery well. PMID- 10870461 TI - New treatment for macular degeneration being tested. PMID- 10870462 TI - Flexibility. Stretching to stay limber. PMID- 10870463 TI - Alcohol and older people. A hidden epidemic. PMID- 10870464 TI - Seafood safety. Steps to safer eating. PMID- 10870465 TI - I know there are antibacterial soaps. Are there "antiviral" soaps? PMID- 10870466 TI - I have tinnitus--a ringing or buzzing in my ears. My doctor says it's related to my hearing loss. Is there any way to cure tinnitus? PMID- 10870467 TI - Chronic obstructive pulmonary disease. Stopping smoking can make a huge difference for people with this lung condition. PMID- 10870468 TI - Human serum antibody responses to oral microorganisms. IV. Correlation with homologous infection. AB - Recent microbiological studies of periodontal disease in humans have supported the concept of a specific bacterial etiology. While individual agents have not been unequivocally identified, numerous Gram-negative members of the subgingival microflora have been implicated. In addition, elevations in systemic antibody responses have been consistent with certain oral microorganisms being involved in an infectious process associated with the disease. This report delineates the relationship between elevated systemic antibody levels and oral colonization with the homologous microorganism at active disease sites. Thirty-four patients with various types of periodontal disease were examined. Using ELISA, each patient was shown to have an elevated antibody response to at least one organism from a battery of 18 oral microorganisms that were tested. Subsequently, subgingival plaque was cultured from disease-active and -inactive sites of each subject. The results demonstrated that the same microorganism to which the individual exhibited elevated serum antibody responses was detected in nearly 55% of the disease-active sites, while only 18% of the inactive sites showed the microorganism. Certain microorganisms including Actinobacillus actinomycetemcomitans, Bacteroides gingivalis, Eikenella corrodens and Wolinella recta were primarily or exclusively correlated with active disease lesions. These findings support the hypothesis that elevated systemic antibodies to periodontopathic bacteria are reflective of subgingival colonization and exist as a response to a bacterial infection at disease-active sites. PMID- 10870469 TI - Suspected periodontopathic microorganisms and their oral habitats in young children. AB - Samples of subgingival plaque from 67 children, 5-7 years of age, were examined for the presence of certain suspected periodontal pathogenic species using the conventional technique of anaerobic sonification, dilution and spiral plating. When this technique was compared with a direct plating procedure which involved no preliminary dispersion and dilution of plaque specimens, it was found that the direct method resulted in double the frequency of children in whom black pigmented Bacteroides (BPB) were detected and a 10-times increase in the number of subjects harbouring Actinobacillus actinomycetemcomitans. Samples from the tongue, tonsils and saliva were also plated using the direct technique. BPB were detected less commonly in the plaque specimens (61.3% of children) than in saliva (89.5%), or on the tongue (86.6%) and tonsils (97.1%). Expressed as percentages of a pooled sample of the total BPB population, the most frequently detected species in plaque were Bacteroides intermedius (44.4%) and Bacteroides melaninogenicus (48.0%). The most prevalent isolate in all other oral sites was B. melaninogenicus. Expressed as percentages of children in whom BPB were detected, the most frequently isolated species from plaque using the conventional dilution technique was B. intermedius (21.3%), whereas other BPB species were present in fewer than 5% of children. Fusobacterium nucleatum and Capnocytophaga species were isolated most frequently from plaque but were also commonly detected in the various other oral sites. PMID- 10870470 TI - Prevalence and microbiology of localized prepubertal periodontitis. AB - Loss of crestal alveolar bone at primary teeth was ascertained radiographically in a dental school clinical population of 2264 children. 19 patients (0.84%) demonstrated distinct periodontal bone destruction around one or more primary teeth; in only 2 of these patients had periodontal disease been identified in previous clinical examinations. A microbiological study of 35 subgingival samples from 9 available patients revealed a high prevalence of black-pigmented Bacteroides spp., mainly Bacteroides intermedius. Actinobacillus actinomycetemcomitans and Capnocytophaga spp. were predominant organisms in some samples. The present data indicate that localized prepubertal periodontitis is more common than previously realized and is associated with bacteria generally regarded as major periodontal pathogens. PMID- 10870471 TI - Microbiology of subgingival plaque from children with localized prepubertal periodontitis. AB - Localized prepubertal periodontitis has been described as a host-defect mediated form of bacterially induced periodontitis, with an early onset and rapid progression around a few teeth in children prior to puberty. To further our understanding of the etiology of this disease, we have examined the microbiological components of subgingival dental plaque in 9 children with localized prepubertal periodontitis to determine if patterns of putative pathogens existed, and have compared these results with those obtained from 4 children with no periodontitis. Subgingival plaque samples were plated onto a selective medium for Actinobacillus actinomycetemcomitans and onto a non selective medium for anaerobes, and the predominant cultivable microbiota of 2 sites per child was determined. The subgingival microbiota of children with localized prepubertal periodontitis clearly differs from non-diseased children in the detection of high levels of several suspected pathogens, including A. actinomycetemcomitans, Bacteroides intermedius, Eikenella corrodens, and Capnocytophaga sputigena. These putative pathogens were found in various combinations. These findings suggest that localized prepubertal periodontitis is associated with specific subgingival bacteria which are generally not found in children without periodontitis. PMID- 10870472 TI - Purification and characterization of a thiol-protease from Bacteroides gingivalis strain 381. AB - A protease from the culture supernatant of Bacteroides gingivalis 381, which hydrolyzes the chromogenic substrate Na-benzoyl-DL-arginine-p-nitroanilide (BAPNA), was partially purified by ammonium sulfate precipitation, gel filtration, and anion exchange chromatography. The molecular weight of this protease was determined by SDS-PAGE to be ca. 49,000. The optimum pH was around 7.6. The protease was stable at neutral pH and up to 40 degrees C. The isoelectric point was 4.9. The enzyme activity was enhanced by dithiothreitol, L cysteine, and 2-mercaptoethanol and inhibited by p-chloromercuribenzoic acid, N ethylmaleimide, and iodoacetic acid. PMID- 10870473 TI - Taxonomic study of spirochetes isolated from human periodontal lesions. AB - Nineteen strains of spirochetes obtained from subgingival plaque of 19 patients with advanced periodontitis were studied morphologically, biochemically, and genetically and compared with type strains of Treponema denticola and Treponema socranskii. The results showed that 16 strains which biochemically resembled T. denticola could be divided into two groups based on G + C content and DNA homology. Two isolates appeared to belong to the T. socranskii species. One intermediate size isolate did not fit any known Treponema species. PMID- 10870474 TI - Characterization and cross-reactivity of rabbit antisera to plaque toxins. AB - The effects of heat labile, high molecular weight water-soluble toxins from bacterial plaque on HL60 promyelocytic cells were examined. On gel filtration, four inhibitors of HL60 cell growth and two inhibitors of HeLa cell growth (PT1, PT2) were detected. The first and third HL60 cell inhibitors corresponded to the two HeLa cell inhibitors. The last eluted HL60 cell inhibitor (plaque leukotoxin, PL) did not inhibit HeLa cell growth. Anti-PT2 antibodies reduced the activity of enriched PT2 by 20-50%, but all other antisera tested exhibited no effect. Anti PL antibodies detected antigens from Actinobacillus actinomycetemcomitans, although anti-A. actinomycetemcomitans and anti-Capnocytophaga sputigena antibodies did not react with plaque extract. These findings suggest that the plaque toxins examined in this study were probably not derived from these two bacteria. PMID- 10870475 TI - Effect of dental plaque on the oxidative metabolism of normal neutrophils. AB - The purpose of this study was to investigate the interaction between dental plaque and the oxidative metabolism of blood (PB), crevicular (CR) and salivary (SAL) PMN. Data indicated that supragingival plaque induced an in vitro production of chemiluminescence by both PB and CR--PMN in a dose-dependent manner with a maximum activity after 30 min incubation. Comparison between PB, CR and SAL-PMNs indicated that 1) both CR and SAL-PMNs spontaneously produced large quantities of oxygen radicals, 2) CR and SAL-PMNs further produced oxygen radicals upon phorbol myristate acetate or opsonized-zymosan stimulation, and 3) SAL-PMN could not be further activated by supragingival plaque. PMID- 10870476 TI - Current issues in the management of acute promyelocytic leukemia. AB - Acute promyelocytic leukemia (APL) is caused by one of four genetic lesions that disrupt the alpha receptor for retinoic acid, RAR alpha. The fusion protein responsible for greater than 99% of APL cases, PML-RAR alpha, inhibits PML dependent apoptotic pathways in a dominant negative fashion and blocks myeloid differentiation by direct transcriptional inhibition of retinoic acid target genes. This transcriptional inhibition is mediated by recruitment of co-repressor proteins and resultant deacetylation of histones in the promoter regions of genes (yet to be identified) that control promyelocyte development. In the presence of high levels of all-trans retinoic acid (ATRA), both PML-dependent apoptotic mechanisms and myeloid-specific gene expression programs are reactivated. In the clinic, the combination of anthracycline-based chemotherapy plus ATRA cures approximately 80% of APL patients, and a high percentage of relapsed patients can achieve second remissions with arsenic trioxide. With the publication of results from the European APL 93 trial, the 'standard-of-care' for induction treatment of APL now includes ATRA plus concurrent anthracycline-based chemotherapy. The amount and type of consolidation therapy necessary for an individual APL patient remains somewhat of an open question, but at present should include at least two cycles of chemotherapy. Based on recent trials that demonstrate a benefit of maintenance ATRA (+/- low-dose chemotherapy), all APL patients should probably receive some type of maintenance therapy. While the above approach currently cures the majority of APL patients, future improvements in the treatment of this disease will require risk-adapted protocols that incorporate real-time molecular monitoring and appropriate introduction of novel therapeutic agents. PMID- 10870477 TI - Prevalence of prothrombin 20210A allele and methylenetetrahydrofolate reductase C677T genetic mutations in the Chinese population. AB - From July 1997 to June 1998, a total of 1323 subjects, including 1180 controls, 94 patients with diabetes mellitus, and 49 patients with deep-vein thrombosis, varying in age and gender, were consecutively entered into our study. Their mean (+/- SD) age was 50.0 +/- 18.0 years, range 1-99 years; 930 were male and 393 were female. None of the subjects was found to have abnormal prothrombin 20210A allele mutation. In total, 150 subjects (11.3%) were found to have a homozygous 677 C-->T mutation of the methylenetetrahydrofolate reductase gene, in which 125 were controls (10.6%), 17 were diabetics (18.1%) and 8 were patients with deep vein thrombosis (16.3%). However, 524 subjects (39.6%) were found to have a heterozygous methylenetetrahydrofolate reductase 677 C-->T mutation. We suggested that the Chinese race dose not have the prothrombin 20210A allele, but can carry the 677 C-->T mutation of the methylenetetrahydrofolate reductase gene. PMID- 10870478 TI - Effect of interleukin-3, stem cell factor and granulocyte-macrophage colony stimulating factor on committed stem cells, long-term culture initiating cells and bone marrow stroma in a one-step long-term bone marrow culture. AB - Long-term bone marrow culture (LTBMC) supports both differentiation and conservation of hematopoietic progenitor cells. During the culture period, the frequency and proliferative potential of early repopulating stem cells that give rise to progenitors detectable after 5 weeks in LTBMC--so-called long-term culture-initiating cells (LTC-IC)--can be investigated. The adherent stroma cell layer was modified by interleukin-3 (IL-3) + granulocyte-macrophage colony stimulating factor (GM-CSF)+ stem-cell factor (SCF) with an increased cellularity and higher percentage of differentiated myeloid cells and a reduced percentage of stroma cells. We have studied the effects of stimulative cytokines, such as GM CSF, SCF, and IL-3, on proliferation of committed colony-forming unit cells (CFU C), LTC-IC, and stroma cell formation in LTBMC of unseparated bone-marrow cells. IL-3, GM-CSF, and SCF significantly stimulated the cumulative proliferation of nucleated cells and committed CFU-C. Weekly stimulation of LTBMC during the culture period did not exhaust the proliferative capacity of stem cells as seen in maintenance of LTC-IC after 5 weeks in LTBMC. In contrast, no LTC-IC were seen in limiting dilution analyses from LTBMC cultured 5 weeks without cytokine supplementation. Our data indicate that an increased proliferation of committed stem cells can be achieved by the addition of stimulatory growth factors, and maintenance of LTC-IC is possible over a period of 5 weeks. A net expansion of LTC-IC if compared with the starting bone-marrow suspension could not be obtained after a 5-week LTBMC period. PMID- 10870479 TI - Critical role of nitric oxide for proliferation and apoptosis of bone-marrow cells under septic conditions. AB - Sepsis is a state of high turnover of bone-marrow cells. Nitric oxide (NO) is reported to be involved in cell proliferation and demise. Murine bone-marrow cells were incubated with lipopolysaccharide together with tumor necrosis factor alpha, interferon gamma and interleukin-1 beta for 48 h. The basal proliferation rate of the cells remained unchanged, but granulocyte-macrophage colony stimulating factor-induced proliferation was suppressed and the percentage of apoptotic cells significantly raised. Levels of nitrite in the culture supernatants were inversely correlated with the suppression of proliferation, but directly correlated with apoptosis. The NO synthesis inhibitor N-methyl-arginine inhibited the suppression of proliferation as well as the induction of apoptosis and NO synthesis. Our results indicate that NO is a negative feedback regulator of cell turnover in sepsis, which limits growth-factor-induced proliferation and induces apoptosis of bone marrow cells. PMID- 10870480 TI - Effect of treatment with amifostine used as a single agent in patients with refractory anemia on clinical outcome and serum tumor necrosis factor alpha levels. AB - Amifostine increases in vitro burst-forming unit-erythroid and colony-forming unit-granulocyte/granulcoyte-macrophage cultured from bone-marrow cells from patients with myelodysplastic syndrome (MDS). Several small clinical studies give divergent informations about the potential of amifostine as single agent to improve hematopoiesis in MDS patients. In these studies, patients with refractory anemia (RA), RA with excess of blasts (RAEB), and RAEB in transformation (RAEB-T) were analyzed together, resulting in response rates varying from 8% to 30%. The present multi-center study evaluated whether treatment with amifostine is of clinical benefit in patients with RA who are transfusion dependent. The effect on transfusion frequency as well as on platelets and absolute neutrophil count (ANC) was examined in 14 patients with RA [median age 67 years (55-72 years), male:female 9:5]. Four treatment cycles were planned, each consisting of intravenous amifostine at 200 mg/m2/day three times per week followed by a 2-week interval. Since tumor necrosis factor (TNF) alpha is a main suppressive cytokine for hematopoiesis in RA patients, serum samples for analyzing endogenous levels of TNF alpha were collected prior to the study and after four treatment cycles. In three patients (21%), reduced transfusion requirement with prolongation of the transfusion interval from 4 weeks to 8 weeks (two patients) and 4 weeks to 6 weeks was seen. An increase in ANC from 400/microliter to 2600/microliter and 200/microliter to 3400/microliter was observed in two patients. Platelets increased from 129,000/microliter to 277,000/microliter in an additional patient. In one patient, disease progression from RA to RAEB was observed. Serum TNF alpha levels were increased in MDS patients compared with normal controls (18.8 pg/ml vs 9.1 pg/ml), and there was no change during the treatment with amifostine (17.5 pg/ml). In conclusion, treatment with amifostine as a single agent was of limited benefit in patients with RA. The serum TNF alpha levels were unchanged during treatment with amifostine in RA patients. PMID- 10870481 TI - The value of interphase fluorescence in situ hybridization for the detection of translocation t(12;21) in childhood acute lymphoblastic leukemia. AB - Translocation t(12;21)(p13;q22) is the most frequent cytogenetic abnormality in childhood acute lymphoblastic leukemia (ALL) and is generally associated with favorable prognosis. In this report, we assessed the value of dual-color interphase fluorescence in situ hybridization (FISH) for the detection of t(12;21). Fifty-three patients were screened for ETV6/CBFA2 fusion by means of FISH, using two cosmid probes mapped on ETV6 and on CBFA2, respectively. The cut off value (mean + three standard deviations) for positivity established on control patients was 9.3%. A comparison between FISH and molecular methods [reverse-transcriptase polymerase chain reaction/Southern blot (RT-PCR/SB)] was possible in 52 patients: 34 of 52 (65.4%) showed negative results with both approaches, and 13 of 52 (25%) were positive; 5 of 52 (9.6%) showed discrepancies: four patients who were positive using RT-PCR/SB were negative using FISH. Conversely, one patient negative when using RT-PCR/SB was positive with FISH. Further investigations on this patients, cytogenetically characterized by add(12p), showed an atypical breakpoint on ETV6, located 5' to the common breakpoint. Compared with RT-PCR and SB, dual-color interphase FISH with the cosmid probe set proved to be highly specific but showed limited sensitivity. PMID- 10870482 TI - Recurrent idiopathic iridocyclitis after autologous peripheral blood stem-cell transplantation followed by G-CSF administration for acute lymphoblastic leukemia. AB - We describe a patient who experienced a recurrence of idiopathic iridocyclitis on day 12 after autologous peripheral blood stem-cell transplantation (auto-PBSCT) followed by G-CSF administration for acute lymphoblastic leukemia (ALL). Autologous SCT has been reported to be effective and safe in achieving dose intensification of chemotherapeutic drugs for the treatment of hematopoietic malignancies, but its therapeutic effect on autoimmune diseases is not definite. The findings from the present case suggest that auto-PBSCT followed by G-CSF administration for patients with a history of some kind of autoimmune disorders may induce exacerbation or recurrence of its symptoms after hematopoietic recovery. PMID- 10870483 TI - Granular acute lymphoblastic leukemia with hypereosinophilic syndrome. AB - A four-year-old boy presented with marked peripheral blood eosinophilia (absolute eosinophil count of 54 x 10(9)/1), features of hypereosinophilic syndrome, and acute lymphoblastic leukemia (ALL-L2), the latter characterized by the presence of granular blasts. Blasts were negative for myeloperoxidase, non-specific esterase, acid phosphatase, periodic-acid Schiff stain, and toluidine blue. They exhibited an early pre-B immunophenotype (TdT, CD19, CD10, CD20 and CD22 positive) and stained negative for T (CD7, CD2, CD5 and CD3) and myeloid markers (MPO, CD33 and CD13). Chromosomal analysis revealed a normal karyotype. To the best of our knowledge, this case represents the first report of the coexistence of granular ALL and hypereosinophilic syndrome. PMID- 10870485 TI - Staphylococcal pyomyositis in a patient with non-Hodgkin's lymphoma. AB - Pyomyositis is a rare disease, encountered mainly in tropical climates. The diagnosis of this entity is difficult, if not misdiagnosed, because of its rarity and its subacute presentation. We report of a 42-year-old man, in whom pyomyositis developed while he was receiving the standard chemotherapy for T-cell non-Hodgkin's lymphoma (NHL). Three months following splenectomy, multiple abscesses occurred in the muscles of both thighs while the patient was receiving the third course of the CHOP regimen. A purulent exudate was aspirated from the abscesses under computed tomographic guidance. Coagulase-positive Staphylococcus aureus was cultured in the aspirate. Pyomyositis was completely resolved following the surgical drainage and the antistaphylococcal antibiotic treatment. This patient has shown that immunosuppression due to splenectomy, NHL, and chemotherapy, especially when using steroids, could be risk factors for pyomyositis in nontropical or semitropical countries. PMID- 10870484 TI - Acute myelofibrosis: multifocal bone marrow infiltration detected by scintigraphy and magnetic resonance imaging. AB - Acute myelofibrosis is a rare, malignant hematological disorder of unknown etiology with an inevitably fatal outcome. Here we present the study of a 63-year old Caucasian man with acute onset of pancytopenia. Repeated bone marrow biopsies showed dense fibrosis and hypoplastic hematopoiesis raising various differential diagnoses of malignant and nonmalignant conditions. Bone marrow scintigraphy and magnetic resonance imaging (MRI) showed areas suggesting neoplastic infiltration, mainly in both femurs and tibias. Histological examination of a surgical biopsy of the left tibia revealed acute megakaryoblastic leukemia. As the patient refused polychemotherapy, therapy with interferon gamma was initiated but discontinued prematurely because of intolerable side effects. The presented case therefore suggests that the combination of bone marrow scintigraphy and MRI is a valuable diagnostic tool in patients presenting with myelofibrosis of unknown origin. PMID- 10870486 TI - Palliative care. PMID- 10870487 TI - The management of cancer pain. AB - Any therapeutic strategy developed for patients experiencing cancer pain depends on the goals of care, which can be broadly categorized as prolonging survival, optimizing comfort, and optimizing function. The relative priority of these goals for any individual should direct therapeutic decision-making. By combining primary treatments, systemic analgesic agents, and other techniques, most cancer patients can achieve satisfactory relief of pain. In cases where pain appears refractory to these interventions, invasive anesthetic or neurosurgical maneuvers may be necessary, and sedation may be offered to those with unrelieved pain at the end of life. The principles of analgesic therapy are presented, as well as the practical issues involved in drug administration, ranging from calculating dosage to adverse effects, and, when necessary, how to switch and/or combine therapies. Adjuvant analgesics, which are drugs indicated for purposes other than relief of pain but which may have analgesic effects, are also listed and discussed in some detail. Surgical and neurodestructive techniques, such as rhizotomy or cordotomy, although not frequently required or performed, represent yet other options for patients with unremitting pain and diminished hope of relief. Although cancer pain can be a complex medical problem arising from multiple sources, patients should be assured that suffering is not inevitable and that relief is attainable. PMID- 10870488 TI - Completing the continuum of cancer care: integrating life-prolongation and palliation. AB - Cancer care extends from diagnosis through the late stages of advanced illness as patients confront dying and their families cope with caregiving and grief. Palliative care is a rapidly developing area of clinical focus that offers valuable services to patients in terms of symptom management and adjustment to illness, including issues of life completion and life closure. It is often appropriate to offer certain elements of palliative care early in the course of illness. As disease progresses, physical comfort and enhancing quality of life increasingly become primary goals of cancer care. Specialized palliative care programs, epitomized by hospice, are invaluable resources for patients with far advanced illness and their families. Current regulations and prevailing payment structures limit access to and the scope of hospice services and highlight the need for innovative models of delivering and financing palliative care. PMID- 10870489 TI - The temperate doctor. PMID- 10870491 TI - Electronic wonderland. PMID- 10870490 TI - Training aboriginal health care professionals in Manitoba. PMID- 10870492 TI - Electronic wonderland. PMID- 10870493 TI - Do the right thing. PMID- 10870494 TI - The relative risks and etiologic fractions of different causes of death and disease attributable to alcohol, tobacco and illicit drug use in Canada. AB - BACKGROUND: In 1996 the number of deaths and admissions to hospital in Canada that could be attributed to the use of alcohol, tobacco and illicit drugs were estimated from 1992 data. In this paper we update these estimates to the year 1995. METHODS: On the basis of pooled estimates of relative risk, etiologic fractions were calculated by age, sex and province for 90 causes of disease or death attributable to alcohol, tobacco or illicit drugs; the etiologic fractions were then applied to national mortality and morbidity data for 1995 to estimate the number of deaths and admissions to hospital attributable to substance abuse. RESULTS: In 1995, 6507 deaths and 82,014 admissions to hospital were attributed to alcohol, 34,728 deaths and 194,072 admissions to hospital were attributed to tobacco, and 805 deaths and 6940 admissions to hospital were due to illicit drugs. INTERPRETATION: The use and misuse of alcohol, tobacco and illicit drugs accounted for 20.0% of deaths, 22.2% of years of potential life lost and 9.4% of admissions to hospital in Canada in 1995. PMID- 10870495 TI - Nonmedical drug use among adolescent students: highlights from the 1999 Ontario Student Drug Use Survey. AB - BACKGROUND: During the 1990s, rates of nonmedical drug use among adolescents escalated. We assessed data from 5 cycles of the Ontario Student Drug Use Survey for overall trends in the proportion of students reporting illegal drug use between 1991 and 1999. METHODS: The survey is a repeated, cross-sectional, 2 stage cluster-design survey of students enrolled in grades 7, 9, 11 and 13. Outcome measures were prevalence of use of 17 drugs, including alcohol and tobacco, over the 12 months preceding the survey. RESULTS: The rates of drug use increased between 1993 and 1999. The 95% confidence intervals (CIs) for the differences in proportions between 1997 and 1999 indicated significant increases in the overall use of 6 drugs: alcohol (95% CIdiff 6.1, 1.9-10.3), cannabis (95% CIdiff 46.3, 0.2-8.4), glue (95% CIdiff 2.3, 1.3-3.3), other solvents (95% CIdiff 5.0, 3.1-6.3), barbiturates (95% CIdiff 1.9, 0.4-3.4) and hallucinogens such as mescaline and psilocybin (95% CIdiff 3.5, 0.8-6.9). Fewer grade 7 students in 1999 than in earlier cohorts reported using alcohol or cigarettes by age 9. INTERPRETATION: The public health implications of the findings are mixed. On the positive side, there is no evidence of increases in early onset of drug use. On the negative side, the overall proportion of students reporting illegal drug use has continued to rise. PMID- 10870496 TI - Self-reported medical use of marijuana: a survey of the general population. PMID- 10870497 TI - Substance abuse: tempering the debate. PMID- 10870498 TI - Stemming needless deaths: "medicalizing" the problem of injection drug use. PMID- 10870499 TI - The cannabis policy debate: finding a way forward. PMID- 10870500 TI - Substance use: time for drug law reform. PMID- 10870501 TI - Interpreting the relation between injection drug use and harm: a cautionary note. PMID- 10870502 TI - Substance abuse and developments in harm reduction. AB - A drug is a substance that produces a psychoactive, chemical or medicinal effect on the user. The psychoactive effect of mood-altering drugs is modulated by the user's perception of the risks of drug use, his or her ability to control drug use and the demographic, socioeconomic and cultural context. The ability to control drug use may vary along a continuum from compulsive use at one end to controlled use at the other. The "drug problem" has been socially constructed, and the presence of a moral panic has led to public support for the prohibitionist approach. The legalization approach has severely attacked the dominant prohibitionist approach but has failed to gain much support in society because of its extreme libertarian views. The harm reduction approach, which is based on public health principles, avoids the extremes of value-loaded judgements on drug use and focuses on the reduction of drug-related harm through pragmatic and low-threshold programs. This approach is likely to be important in tackling the drug problem in the 21st century. PMID- 10870503 TI - Identifying and treating patients with alcohol-related problems. AB - Problem drinking is a serious health issue, but often patients whose alcohol consumption places them at risk are not diagnosed by physicians. Case finding is an essential component of "best practice." In many cases if given the appropriate advice, counselling and behavioural interventions, problem drinkers can be helped to reduce their use of alcohol and improve functioning in other areas of their lives. Some patients may benefit from more comprehensive therapy including the prescription of disulfiram, calcium carbimide or naltrexone. For those with serious problems with alcohol, referral to specialized addiction treatment programs and other community resource centres may also be appropriate. PMID- 10870504 TI - Injection drug use and preventive measures: a comparison of Canadian and western European jurisdictions over time. PMID- 10870505 TI - Prevention must be health focus in northern Ontario. PMID- 10870506 TI - Screening program lets Alberta test newborns' hearing. PMID- 10870507 TI - Transplant queues grow as donor numbers wane. PMID- 10870509 TI - Benefits of genetic research must be shared, international genome organization warns. PMID- 10870508 TI - Substance abuse among physicians. PMID- 10870510 TI - Are MDs at risk when they don't offer patients new medical technologies? PMID- 10870512 TI - Effect of fluoxetine on intraocular pressure in the rabbit. AB - The effects of fluoxetine, which is a selective inhibitor of serotonin reuptake (SSRI) have been studied on the intraocular pressure (IOP) in the rabbit. IOP was measured using a Perkins tonometer. Intravenous administration of fluoxetine (6.0 mg kg-1) significantly increased IOP by 7.2 +/- 1.3 mmHg (P < 0.001). Fluoxetine administration also reduced the amount of urine excreted during the 24 hr, and increased the urine concentration of 5-hydroxyindole acetic acid (5-HIAA). Topical preadministration of ketanserin prevented a rise in IOP, without there being any effects on the other parameters examined. These findings indicate that intravenous administration of fluoxetine is able to raise IOP, through increased plasma levels of serotonin, which is confirmed by elevated 5-HIAA urine excretion and reduction in diuresis. Ketanserin, a specific 5-HT2A antagonist, counteracts the IOP increase brought about by fluoxetine, thus emphasizing the role of serotonin in the regulation of IOP and stressing the importance of including ophthalmological examination in the protocol of depressed patients undergoing SSRI therapy. PMID- 10870513 TI - Tear secretion by lacrimal glands in transgenic mice lacking water channels AQP1, AQP3, AQP4 and AQP5. AB - The expression of three aquaporin (AQP)-type water channels has been reported in the lacrimal gland: AQP5 in the apical membranes of acinar and duct cells, AQP4 in the basolateral membranes of acinar cells, and AQP1 in microvascular endothelia. Recent experiments indicate that water movement through AQP5 in the salivary gland is important in saliva secretion. To investigate the role of aquaporins in lacrimal gland function, basal and pilocarpine-stimulated tear secretion was compared in wildtype mice and knockout mice lacking AQP1, AQP4 and AQP5, as well as AQP3, which was found here to be expressed in the basolateral membrane of acinar cells. Tear fluid was collected in anesthetized mice using microcapillary tubes before and at 4 min intervals after pilocarpine administration. Tear fluid volumes were (in microliter per 4 min, S.E.): 0.69 +/- 0.06 (wildtype mice), 0.70 +/- 0.07 (AQP1 -/-), 0.81 +/- 0.13 (AQP3 -/-), 0.62 +/ 0.14 (AQP4 -/-), and 0.78 +/- 0.09 (AQP5 -/-) (differences not significant). Chloride concentrations (average 155 +/- 13 mM) measured by a fluorescence assay were also not different in tear fluid collected from wildtype vs aquaporin null mice. These findings provide direct evidence against an essential role for aquaporins in lacrimal gland fluid secretion. The requirement for aquaporins in salivary but not lacrimal gland secretion, may involve the substantially slower fluid secretion rate across lacrimal gland acinar cells. PMID- 10870511 TI - An evidence-based approach to the management of uninvestigated dyspepsia in the era of Helicobacter pylori. Canadian Dyspepsia Working Group. AB - OBJECTIVES: To provide Canadian primary care physicians with an evidence-based clinical management tool, including diagnostic and treatment recommendations, for patients who present with uninvestigated dyspepsia. RECOMMENDATIONS: The management tool has 5 key decision steps addressing the following: (1) evidence that symptoms originate in the upper gastrointestinal tract, (2) presence of alarm features, (3) use of nonsteroidal anti-inflammatory drugs (NSAIDs), (4) dominant reflux symptoms and (5) evidence of Helicobacter pylori infection. All patients over 50 years of age who present with new-onset dyspepsia and patients who present with alarm features should receive prompt investigation, preferably by endoscopy. The management options for patients with uninvestigated dyspepsia who use NSAIDs regularly are: (1) to stop NSAID therapy and assess symptomatic response, (2) to treat with NSAID prophylaxis if NSAID therapy cannot be stopped or (3) to refer for investigation. Gastroesophageal reflux disease can be diagnosed clinically if the patient's dominant symptoms are heartburn or acid regurgitation, or both; these patients should be treated with acid suppressive therapy. The remaining patients should be tested for H. pylori infection, and those with a positive result should be treated with H. pylori-eradication therapy. Those with a negative result should have their symptoms treated with optimal antisecretory therapy or a prokinetic agent. VALIDATION AND EVIDENCE: Evidence for resolution of the dyspepsia symptoms was the main outcome measure. Supporting evidence for the 5 steps in the management tool and the recommendations for treatment were graded according to the strength of the evidence and were endorsed by consensus of committee members. If no randomized controlled clinical trials were available, the recommendations were based on the best available evidence. LITERATURE REVIEW: Evidence was obtained from MEDLINE searches for pertinent articles published from 1966 to October 1999. The searches focused on dyspepsia, diagnosis and treatment. Additional articles were retrieved through a manual search of bibliographies and abstracts from international gastroenterology conferences. PMID- 10870514 TI - Effects of latanoprost on tyrosinase activity and mitotic index of cultured melanoma lines. AB - The intraocular pressure-lowering drug latanoprost, a phenyl-substituted analogue of prostaglandin F2 alpha (PGF2 alpha), increases iris pigmentation in a small number of patients. In theory, this could be due to increased melanogenesis or melanocyte proliferation. To distinguish these two possibilities, the present study examined the effects of latanoprost on tyrosinase activity (the rate limiting step for melanin synthesis) and mitotic index of cultured melanoma lines. Murine cutaneous melanoma lines (S91 and B16), and human uveal (OCM1, OCM3, and OM431) and cutaneous (SK-MEL5 and M21) melanoma lines were cultured with PGE1, PGE2, PGF2 alpha, latanoprost, or the adenylate cyclase stimulating agent forskolin. After treatment, tyrosinase was assayed with respect to its dopa oxidase activity using a colorimetric assay. PGE1, PGE2, PGF2 alpha, and latanoprost greatly increased tyrosinase activity in murine melanoma lines and caused small increases in tyrosinase activity in human uveal and cutaneous melanoma lines. Similar results were obtained with the cAMP-elevating compound forskolin. Cyclic AMP content, as determined by an enzyme-linked immunoassay, was similarly increased by all treatments, with forskolin being the most potent stimulator. Since the species difference in tyrosinase activity was observed without an apparent difference in induction of cAMP, latanoprost would appear to induce tyrosinase activity through a non-cAMP-dependent pathway. Finally, latanoprost and PGF2 alpha did not enhance the mitotic index of human uveal or cutaneous melanoma lines, measured by [6-3H] thymidine uptake, although they increased the mitotic index of one murine cutaneous line. Given that latanoprost induced tyrosinase activity, but did not increase the mitotic index in any of the human melanoma lines studied, this suggests that the in vivo iris pigmentation side effect of latanoprost may not result from increased cell division, but from elevated tyrosinase activity. PMID- 10870515 TI - Intrascleral concentration vs depth profile of mitomycin-C after episcleral application: impact of applied concentration and volume of mitomycin-C solution. AB - The purpose of this study was to investigate the impact of different concentrations and volumes of Mitomycin-C (MMC) on the intrascleral concentration vs depth profile of MMC in an experimental model. The episcleral sides of scleral quadrants of human donor eyes were exposed for 1 min to sponges (corneal light shield, Merocel Corp.) soaked with MMC. After irrigation with 40 ml saline a central 8 mm diameter scleral disk was horizontally dissected with a cryotome. MMC concentrations of six layers of 140 microns thickness were analysed by means of high-performance liquid chromatography. In Experiment 1 (11 eyes) the sponges were soaked with 50 microliters of 10, 100 and 200 micrograms ml-1 MMC solutions. In Experiment 2 (12 eyes) the sponges were soaked with 10, 30, 50 and 80 microliters of a 200 micrograms ml-1 isotonic MMC solution. In Experiment 1 the MMC concentrations (microgram g-1) of layer 1 were 0.35 (+/- 0.20; 10 micrograms ml-1 group) and 9.22 (+/- 2.92; 200 micrograms ml-1 group). In Experiment 2 the MMC concentrations were 2.57 (+/- 1.17; 10 microliters group), 7.35 (+/- 2.49; 30 microliters group) and 11.67 (+/- 3.25; 80 microliters group). The scleral MMC concentrations were significantly influenced by the applied concentrations (layers 1-5) and by the applied volumes (all layers) of MMC solution. The intrascleral MMC concentration increased linearly with increasing concentration and not linearly with increasing volume of the applied MMC solution. To achieve more predictable scleral concentrations of MMC after trabeculectomy with MMC it seems advisable to control both the concentration and the volume of the MMC solution used to soak the sponge. PMID- 10870516 TI - Adenosine A2A receptors regulate the extracellular accumulation of excitatory amino acids upon metabolic dysfunction in chick cultured retinal cells. AB - The role of endogenous extracellular adenosine as a tonic modulator of the extracellular accumulation of excitatory amino acids (glutamate and aspartate) caused by metabolic inhibition was investigated in cultured retinal cells. The selective adenosine A2A receptor antagonist, 4-[2-[7-amino-2-(2-furyl)(1,2,4) triazin-5-ylamino]-ethyl]ph enol (ZM241385) (50 nM), increased the release of glutamate (three- to four-fold) and of aspartate (nearly two-fold) upon iodoacetic acid-induced glycolysis inhibition, in the presence or in the absence of Ca2+. Blockade of tonic activation of A2A receptors by ZM241385 also increased (nearly two-fold) the ischemia-induced release of glutamate and aspartate. Furthermore, another selective A2A receptor antagonist, 5-amino-7-(2-phenylethyl) 2-(2-furyl)pyrazolo[4,3-e]-1,2,4-triazolo[1,5- c] pyrimidine (SCH58261), also increased the release of aspartate and glutamate by about two-fold in cells submitted to glycolysis inhibition. In contrast, the selective adenosine A1 receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) (100 nM), did not significantly modify the extracellular accumulation of either glutamate or aspartate caused by inducers of chemical ischemia or glycolytic inhibitors. Inhibition of glycolysis also increased (about three-fold) the extracellular accumulation of GABA, which was virtually unchanged by ZM241385. Furthermore, the GABAA receptor antagonist, bicuculline (10 microM), only increased (nearly two fold) the iodoacetic acid-induced Ca(2+)-dependent release of glutamate, whereas the GABAB receptor antagonist, 3-aminopropyl(diethoxymethyl) phosphinic acid, CGP35348 (100 microM), was devoid of effects on the extracellular accumulation of glutamate and aspartate. These results show that endogenous extracellular adenosine, which rises under conditions of inhibited glycolysis, tonically inhibits the extracellular accumulation of excitatory amino acid through the activation of A2A, but not A1, adenosine receptors, and this effect is independent of GABAA and GABAB functions in the cultured retinal cells. PMID- 10870517 TI - Synthesis of elastic microfibrillar components fibrillin-1 and fibrillin-2 by human optic nerve head astrocytes in situ and in vitro. AB - The purpose of this study was to identify elastic microfibrillar components fibrillin-1 and fibrillin-2 in optic nerve heads of adult normal and glaucomatous subjects, in cultured optic nerve head astrocytes (type 1B astrocytes), as well as fibrillin-1 in fetal optic nerve heads. To characterize synthesis and gene expression of microfibrillar proteins in human optic nerve heads and cultured type 1B astrocytes, light microscopy immunohistochemistry, in situ hybridization, and RT-PCR or Northern blots were performed. Our results demonstrated that fibrillin-1 was associated with blood vessels, astrocytes in the glial columns and cribriform plates, and with astrocyte processes in the nerve bundles in all samples. In glaucomatous optic nerves there was enhanced fibrillin-1 immunoreactivity, especially surrounding blood vessels. Fibrillin-2 was localized primarily to blood vessels in all samples, without qualitative differences between normal and glaucomatous samples. In fetal optic nerve heads fibrillin-1 mRNA was localized to glial cells and to the blood vessel walls. In adult optic nerve heads, there was little fibrillin-1 mRNA as detectable by in situ hybridization and RT-PCR. There was no detectable upregulation of fibrillin-1 mRNA in glaucoma. In cultured type 1B astrocytes, fibrillin-1 staining was mostly pericellular. There was little fibrillin-2 immunoreactivity. In conclusion, astrocytes from the optic nerve head deposit elastic microfibrillar components in situ and in vitro, with a predominance of fibrillin-1. Upregulation of fibrillin 1 mRNA was not observed in glaucoma, suggesting that increased transcription may occur early in the disease process. Cultures of type 1B astrocytes from the optic nerve head provides a useful model to study mechanisms regulating the interactions of elastin and the microfibrils in optic nerve head astrocytes. PMID- 10870518 TI - Low doses of pilocarpine do not significantly increase outflow facility in the cynomolgus monkey. AB - Low doses (10(-9)-10(-6) M) of pilocarpine reportedly increase outflow facility in the organ-cultured human eye, suggesting a direct action on the trabecular meshwork. M3 muscarinic receptors have been found in both cultured human trabecular meshwork cells and tissue. We determined whether low pilo doses would increase outflow facility in the living monkey. The anterior chambers of both eyes of 17 pentobarbital anesthetized cynomolgus monkeys were cannulated and outflow facility measured bilaterally by 2-level constant pressure perfusion after an initial 2 ml exchange with Barany's perfusand containing 24.5 microM phenylephrine (PE). Two subsequent exchanges were performed with one eye receiving Barany's + PE + 10(-10)-10(-4) M pilocarpine and the contralateral eye receiving only Barany's + PE. Outflow facility was measured for 35-40 min following each exchange. Accommodation and pupil diameter were measured before each exchange and approximately every 10 min during facility measurements. Outflow facility was significantly increased by 154 and 313% in eyes treated with 10(-5) M and 10(-4) M pilocarpine, respectively, related to contralateral controls. Accommodation and miosis also were induced only at 10(-5) M (accommodation, 3.3 +/- 1.6 diopters, NS; miosis, -4.1 +/- 0.5 mm, P < or = 0.001) and 10(-4) M (accommodation, 10.6 +/- 0.0 diopters, P < or = 0.02; miosis, -3.4 +/- 1.0 mm, P < or = 0.025) pilocarpine. We conclude that low anterior chamber doses of pilocarpine do not increase outflow facility in the living monkey as reported in the organ-cultured human eye, nor do they induce miosis or accommodation. All three parameters respond to pilocarpine at similar doses, and there is no functional evidence of a meaningful outflow facility-relevant pilocarpine effect on the trabecular meshwork at doses lower than those which affect the ciliary muscle. PMID- 10870519 TI - Suppressive actions of betaxolol on ionic currents in retinal ganglion cells may explain its neuroprotective effects. AB - Betaxolol, a beta 1-selective adrenoceptor antagonist, is widely used in the treatment of glaucoma. In addition to its ocular hypotensive effects, betaxolol has been suggested to act as a retinal neuroprotective agent (Osborne et al., 1997). To investigate possible mechanisms underlying the neuroprotective effects, we tested the actions of betaxolol on ion channels and calcium signaling in isolated retinal ganglion cells. Betaxolol (50 microM) reduced by about 20% the high-voltage-activated (HVA) Ca channel currents in ganglion cells isolated from tiger salamander retina. In contrast, the beta 1-adrenoceptor antagonists propranolol (10 microM) and timolol (50 microM) had no inhibitory actions on HVA Ca channel currents. The L-type Ca channel antagonist, nisoldipine, blocked the HVA Ca channel current partially and the remaining current was not inhibited by betaxolol. Outward current was inhibited in the presence of betaxolol. Both iberiotoxin (IBTX; 10 nM), a selective inhibitor of large-conductance Ca activated K channels, and Cd2+ (100 microM), which suppresses Ca-activated K channels subsequent to its block of Ca channels, reduced outward current and the remaining current was not blocked significantly with betaxolol. In the presence of betaxolol, Na channel currents were reduced by about 20%, as were currents evoked by glutamate (10 mM) and GABA (1 mM). Current clamp recordings from isolated ganglion cells showed that betaxolol had several effects on excitability: spike height decreased, repetitive spike activity was suppressed, spike width increased and hyperpolarization following spikes was reduced. Calcium imaging in isolated rat retinal ganglion cells revealed that betaxolol inhibited glutamate-induced increases in [Ca2+]i. These results suggest that betaxolol has a diversity of suppressive actions on ganglion cell ion channels and that, as a consequence, one of the net actions of the drug is to reduce Ca2+ influx. The subsequent reduction in [Ca2+]i may contribute to the apparent neuroprotective actions of betaxolol in promoting ganglion cell survival following ischemic insult, as may occur in glaucoma and retinal disease. PMID- 10870520 TI - Localization of prostaglandin E receptor subtypes in the ciliary body of mouse eye. AB - Prostaglandin E2 (PGE2) markedly reduces intraocular pressure (IOP) when applied topically and induces strong relaxation of pre-contracted isolated ciliary muscle through PGE2 receptor. Because the ciliary muscle relaxation reduces IOP by enhancing uveoscleral aqueous outflow, the ciliary muscle where the existence of PGE2 receptors has been demonstrated is thought to be one of the target tissues for PGE2-induced IOP reduction. To investigate the subtypes of PGE2 receptors in the ciliary muscle, the regional distribution of four PGE2 receptor subtypes (EP1, EP2, EP3 and EP4) in the mouse ciliary body was investigated by in situ hybridization using specific probes. Consistent messenger RNA signals for EP1 and EP4 receptors were expressed in the ciliary muscle, although signal levels for these subtypes were less potent as compared with the kidney, which was used as a reference organ. EP2 and EP3 signals were not detected. Stimulation of the EP4 receptor activates adenylate cyclase, which should induce ciliary muscle relaxation. Therefore, the IOP reduction induced by PGE2 analogs may be mediated by the EP4 receptor. In contrast, stimulation of the EP1 receptor is believed to promote intracellular Ca2+ mobilization, and hence should cause ciliary muscle contraction. Thus, the coexistence of EP1 and EP4 receptors in the ciliary muscle suggests that the regulation of ciliary muscle tone by PGE2 is based on a complex mechanism involving multiple receptor subtypes. PMID- 10870521 TI - Choroidal vascular permeability in visually regulated eye growth. AB - The choroidal thickness fluctuates both diurnally and in response to changes in visual input. The fluctuations may represent a physiologic means of aligning the retinal photoreceptors with the focal position of distant images during the emmetropization process. To evaluate the basis for choroidal thickness changes, we studied the sources of the extravascular fluid in the chick choroid in two visually-regulated ocular growth conditions: accelerated ocular growth in goggle induced form-deprivation myopia and ocular growth retardation in the recovery from myopia after goggle removal. Two week old chicks, controls, myopic and those recovering from myopia, received fluorescein dextran (MW = 140,000) as a tracer. It was given by intravenous injection to identify a potential vascular pathway and by intracameral injection to identify a potential pathway from the anterior chamber to the suprachoroidal space. Using a microscopically positioned needle, clear fluid was aspirated from the suprachoroidal space of the enucleated chick eye; this fluid presumably corresponds to the contents of the lacunae, prominent lymphatic-like structures of the chick choroid. Plasma, aqueous humor and suprachoroidal fluid were sampled 1 hr after injection and assayed for both protein content and the tracer dye. Sulfated glycosaminoglycans were assayed in the suprachoroidal fluid, choroid and sclera under each experimental condition. In control chicks, aqueous humor and suprachoroidal fluid protein concentrations were about 0.8 and 9% of plasma levels respectively. Aqueous humor protein concentration was unaltered in myopic or recovering eyes. Suprachoroidal fluid protein concentration in myopic eyes fell dramatically to 1.5% of plasma levels (P < 0.001). In contrast, recovery from myopia led to a marked increase in suprachoroidal fluid protein level to 30% of that in plasma (P < 0.001). None of the procedures affected suprachoroidal fluid protein in the contralateral control eyes. In all three groups of chicks, fluorescein dextran distribution in the suprachoroidal fluid at 1 hr after intravenous injection tracked protein levels, with reduced levels in myopic eyes and elevated levels in recovering eyes. After intracameral injection, suprachoroidal fluid dextran levels were higher in injected eyes of control chicks (P < 0.01) and in recovering eyes (P < 0.001) but lower in myopic eyes (P < 0.01), compared to the levels in the respective contralateral non-injected eyes in each group. Sulfated glycosaminoglycan levels were at the limits of detection in the suprachoroidal fluid under all conditions and, on a whole choroid basis, were unaltered in the choroid in either myopia or recovery. Suprachoroidal fluid is lymph-like in nature and largely derives from plasma. Sulfated glycosaminoglycan levels do not seem to regulate the fluid content of the choroid in either myopia or recovery. Instead, the changes in protein and marker dye levels in myopic and recovering eyes suggest markedly altered choroidal circulatory dynamics and capillary permeability in both conditions. PMID- 10870522 TI - Proliferation of lacrimal gland acinar cells in primary culture. Stimulation by extracellular matrix, EGF, and DHT. AB - The study of lacrimal dysfunction and insufficiency, a major cause of dry eye, has been hampered by the inability to induce the proliferation of primary lacrimal acinar cells in vitro. Particularly in light of observations that androgens are able to support the overall size and functional status of the lacrimal glands as well as certain specific lacrimal functions, an in vitro culture system that is permissive for cell proliferation would be most beneficial to study the molecular basis for these processes. Here, we report on the successful establishment of such a system. Using a culture system containing Hepato Stim Medium and Matrigel, we were able to induce the efficient proliferation of primary rabbit lacrimal gland acinar cells with epidermal growth factor (EGF) and dihydrotestosterone (DHT). The generation of this in vitro cell culture system should greatly facilitate study of the regulation of acinar cell function at the molecular and cellular levels. PMID- 10870523 TI - Quantitation of rat lacrimal secretion: a novel sandwich ELISA with high sensitivity. AB - Modulation of lacrimal acinar cell tear secretion may involve multiple factors acting both in subtle and pronounced ways. Functional screens of recombinant protein products arising from gene array technologies, or protein fractions derived from lacrimal conditioned media or extracellular matrix, will require a highly sensitive assay capable of monitoring tear protein secretion by small replicate cultures. To improve significantly on current methods, a rat- and mouse specific sandwich ELISA was developed. For this purpose, chickens and rabbits were immunized with serum-free secretion media from carbachol and VIP-stimulated rat lacrimal acinar cell cultures. Immune sera were characterized by ELISA, Western blotting and immunohistochemistry, and subsequently optimized for use in a sandwich ELISA. Both antisera detected a wide range of different rat tear proteins, and immunostained only the secretory granule-rich juxtalumenal region in sections of rat lacrimal gland. Chicken, but not rabbit, antiserum cross reacted with rabbit and human tears. In sandwich ELISA, capture with purified chicken immunoglobulin fraction and detection with rabbit antiserum detected as little as 1 ng ml-1 tear protein in 10,000-fold diluted rat secretion medium--a level of sensitivity 8000 times greater than the rat tear peroxidase assay. Such specificity and sensitivity greatly reduce the quantity of media needed for assay, and makes feasible functional screens for scarce factors that may influence lacrimal secretory processes, and in turn possibly play a role in human lacrimal insufficiency syndromes. PMID- 10870524 TI - Timolol lowers intraocular pressure but does not inhibit the development of experimental myopia in chick. AB - Reports of intraocular pressure (IOP) being higher in myopes than emmetropes and of myopes being over-represented in glaucoma statistics, are consistent with a role of IOP in the excessive eye growth typically associated with myopia. We tested the hypothesis, based on these observations, that ocular hypotensive drugs would slow myopia progression using the chick as an animal model and timolol as an example of such a drug. To induce myopia, chicks (n = 56) were fitted with either monocular translucent diffusers or -15 D spectacle lenses from day 8. The drug treatment protocol comprised topical applications of 0.4% benoxinate, a local anaesthetic (to improve drug absorption), followed either by 0.5% timolol or distilled water (control), either daily (1000 hr) or twice daily (1000, 1600 hr). Refractive errors and ocular dimensions were measured on days 12 and 17. We also verified the ocular hypotensive effect of timolol in both normal (n = 8) and myopic (n = 12 diffusers; n = 12-15 D lenses) chicks. Here, we took baseline IOP measurements, instilled timolol and then monitored IOP over a further 5-9 hr. We found no difference in the amount of myopia produced in the timolol and control groups at either measurement time point (e.g. day 17, once per day application, diffusers: -26.9 +/- 3.3 D vs -22.7 +/- 9.1 D; lenses: -14.9 +/- 3.8 D vs -14.9 +/- 3.6 D). This was in spite of the fact that timolol did lower IOP in both normal and myopic chicks (27 and 18% reduction, respectively) While timolol does have an ocular hypotensive effect in the chick, it does not inhibit the development of myopia in this animal model. PMID- 10870525 TI - DC electric fields induce rapid directional migration in cultured human corneal epithelial cells. AB - After an epithelium is wounded, multiple soluble and extracellular matrix associated signals induce a repair response. An often-overlooked signal is the endogenous electrical field established in the vicinity of the wound immediately upon disruption of epithelial integrity. Previous studies have detected lateral electric fields of approximately 42 mV mm-1 near bovine corneal wounds. In addition, electric fields on the order of 100-200 mV mm-1 have been measured lateral to wounds in mammalian epidermis. Here we report the migratory response of human corneal epithelial cells to DC electric fields of similar, physiologic magnitude. Our findings demonstrate that in a 100 mV mm-1 DC field, corneal epithelial cells demonstrate directed migration towards the cathode. The migratory speed and distances traversed by cultured human corneal epithelial cells is remarkably similar to those of cultured skin-derived keratinocytes under similar conditions; however, corneal epithelial cells demonstrate a more rapid directional response to the field than keratinocytes. These findings suggest that endogenous, wound-induced electric fields present in the cornea play an important role in human corneal wound healing, by orienting the directional response of migratory cells so that they efficiently re-epithelialize the wounded area. PMID- 10870526 TI - Modulation of ocular hydrodynamics and iris function by bremazocine, a kappa opioid receptor agonist. AB - This study was designed to determine the activity of bremazocine (BRE), a relatively selective kappa opioid receptor agonist, on intraocular pressure (IOP), aqueous humor formation and pupil diameter (PD) in conscious, normal, dark adapted New Zealand white (NZW) rabbits. IOP was measured in normal and unilaterally sympathectomized rabbits using a calibrated pneumatonometer and the aqueous flow rate was determined by the use of a Fluorotron Master. A masked design study was conducted in which the rabbits' eyes were treated with BRE topically and unilaterally; the fellow eyes received vehicle. IOP and PD measurements were taken at 0.5 hr and 0 time before BRE and 0.5, 1, 2, 3, 4 and 5 hr post-treatment. Fluorophotometry recordings were taken at 1 hr before and 0.5, 1.5, 2.5 and 3.5 hr after topical application of the drug or vehicle. The effect of the relatively selective kappa opioid receptor antagonist, nor-binaltorphimine (nor-BNI), on bremazocine-induced changes in IOP, PD and aqueous flow was also determined. BRE (10 and 100 micrograms 25 microliters-1 vehicle) produced dose related, bilateral reductions in IOP, PD and aqueous humor flow. A large increase in IOP (14 mmHg) was observed when BRE (100 micrograms) was applied to sympathectomized eyes. This ocular hypertensive effect was antagonized when the sympathectomized eyes were pretreated with naloxone (200 micrograms), a non selective opioid receptor antagonist. BRE (10 and 100 micrograms) decreased the aqueous humor flow rate bilaterally by approximately 48 and 60%, respectively, at 0.5 hr after administration to the ipsilateral eye. Nor-BNI (100 micrograms) antagonized the effect of BRE (10 micrograms) on IOP and aqueous flow rates more effectively than on PD. These data indicate that bremazocine causes reductions in IOP by suppressing aqueous flow, but the ocular hypotensive effects are dependent on the presence of intact sympathetic nerves. Antagonism of BRE's effects on aqueous humor dynamics by nor-BNI suggests that the mechanism of IOP and aqueous flow reduction may involve, in part, an action on kappa receptors. Further experiments are necessary to fully define the opioid receptor populations in the ciliary body. PMID- 10870527 TI - Normal mouse and rat strains as models for age-related cataract and the effect of caloric restriction on its development. AB - The purpose of this study was to determine: (1) which of the commonly used strains of laboratory rats and mice provide good models for human age-related cataract, and (2) whether long term caloric restriction, a regimen that prolongs both median and maximum life span in rodents, would also delay the time of appearance of this age-related pathology. Three strains of mice and two rat strains commonly used in laboratory work and maintained on either ad libitum (AL) or calorically restricted (CR) diets in the National Institutes of Aging and Diet Restriction colony were examined by slit lamp for age-related cataracts at four or more time points during their life spans. These strains were Brown Norway and Fischer 344 rats, and C57BL/6, (C57BL6 x DBA/2)F1 and (C57BL/6 x C3H)F1 mice. None of these strains develop congenital cataracts. Various stages of cataract were found in the great majority of these animals in old age. In both rat strains and one mouse strain the cataracts occurred after mid-life, were most advanced late in life, and were similar in locations and appearance to those in humans. In the two mouse strains in which some cataracts appeared as early as 10-14 months of age, previously identified genetic defects affecting the eye were probably involved in the early appearances. CR extended life spain in all five rat and mouse strains and also delayed both the time of first appearances and the subsequent increase in cataract severity over time in the four dark-eyed strains. CR did not delay cataract formation in the single albino rat strain studied. In summation: (1) commonly used strains of laboratory rats and mice that are free of congenital or early appearing cataracts due to genetic defects would appear to serve as appropriate models for human age-related cataract, (2) caloric restriction (CR) provides a protective effect, delaying development of cataracts in the dark-eyed mouse and rat strains, while also extending their life spans, (3) CR did not delay the development of lens damage in the nonpigmented eye of the single albino strain studied, although it extended life span. PMID- 10870528 TI - Quantitative relationship of the scotopic and photopic ERG to photoreceptor cell loss in light damaged rats. AB - In order to use the ERG to track the effects of potential photoreceptor rescue treatments, we have compared retinal histology to the ERG in light damage. Male albino CD rats (40) were purchased at 7 weeks of age and reared in 50 lx cyclic light until 8 week old. They were exposed to a range of light intensities using white fluorescent light (1000, 1500, 2000, 2500 or 3000 lx) for 24 or 48 hr (n = 5 per group). Controls remained in dim cyclic light. Seven days after exposure, dark and light adapted ERGs were recorded from threshold up to 200 cd m-2 using 50 ms Ganzfeld white light stimuli. The STR, and scotopic and photopic b-wave thresholds and amplitudes were measured. After recording the ERG, the eyes were removed from the animals in each of the five 48 hr light exposed groups and control group for histological measurements. These included: (1) outer nuclear layer width in rod photoreceptor cell number (cell count) and micrometers, and (2) outer + inner segment layer width along the vertical meridian in the inferior retina. The product of cell count and outer + inner segment length was calculated. All histological measures showed a statistically significant linear relationship to light exposure intensity (P < 0.0001): r2 = 0.94 (cell count), 0.90 (outer nuclear layer width), 0.77 (outer + inner segment length). The log of the scotopic b-wave threshold and log amplitude showed a significant linear correlation to all histological parameters (P < 0.0001) and there was no significant difference between b-wave threshold and amplitude for any one of the histology measures used. However, overall, log b-wave threshold was significantly better correlated to histology P < 0.02. Only log b-wave amplitude showed a significant increase in variability in light damaged retinas (P < 0.02). The b wave threshold intensity increased 0.33 log cd m-2 and the maximum amplitude decreased 0.23 log microV with each 10% decrease in cell number in the outer nuclear layer. The sensitivity of the scotopic threshold response, which originates from third order neurons, changed much more slowly with cell loss, than did the b-wave (P < 0.0005) and was well fit by a linear relationship to cell loss. The increase in photopic b-wave threshold was not significant for a cell loss of less than 70-80%. Neither the photopic or scotopic b-wave could be reliably recorded with more than 80% cell loss, but the scotopic threshold response remained. Both the scotopic and photopic ERG showed similar waveform changes near the threshold, including loss of the positive going b-wave and the predominance of a negative going response. Outer nuclear layer cell counts in this study showed the same relationship to log b-wave threshold elevation, as has been previously shown for whole retinal rhodopsin content in light damage, indicating that regional histology measurements can be good indicators of overall cell survival. Both the b-wave threshold and amplitude can be reliably used to track photoreceptor cell loss due to the damaging effects of constant light, but the scotopic threshold response may be more useful in severe damage. PMID- 10870529 TI - Migraine treatment and mistreatment: primum non nocere. PMID- 10870530 TI - Government policymaking, private health insurance and hospital-efficiency issues. PMID- 10870531 TI - Clinical pathways and fractured neck of femur. PMID- 10870532 TI - Virtually viewing the large bowel: the future of colorectal cancer screening? PMID- 10870533 TI - Minimally invasive parathyroidectomy: 50 consecutive cases. AB - OBJECTIVE: To determine the effectiveness and outcomes of minimally invasive parathyroidectomy. DESIGN: Prospective, non-randomised, non-blinded trial. SETTING: Affiliated university teaching hospitals of the Northern Clinical School, University of Sydney, New South Wales, May 1998 to October 1999. PATIENTS: 50 consecutive patients who underwent minimally invasive parathyroidectomy for primary hyperparathyroidism, and 150 consecutive patients undergoing open parathyroidectomy over the same period. RESULTS: Minimally invasive parathyroidectomy was successfully completed and resulted in cure (normocalcaemia) in 42 of 50 patients (84%). Seven patients (14%) required conversion to an open procedure, all of which also resulted in normocalcaemia, giving an overall cure rate of 98%. One patient had persistent hyperparathyroidism after minimally invasive parathyroidectomy which was cured at subsequent open reoperation. Three patients had a temporary recurrent laryngeal nerve palsy. Open parathyroidectomy was successful in 147 of 150 patients (98%) at initial operation; one patient had a temporary recurrent laryngeal nerve palsy. Intraoperative measurement of parathyroid hormone levels by a quick technique in 23 of the patients (13 having minimally invasive and 10 open procedures) correctly identified the presence of multiple-gland disease. CONCLUSION: Minimally invasive parathyroidectomy is a feasible procedure, although there are concerns about the complication rate. PMID- 10870534 TI - Clinical pathway for fractured neck of femur: a prospective, controlled study. AB - OBJECTIVE: To assess outcomes of using a clinical pathway for managing patients with fractured neck of femur. DESIGN: Prospective, pseudorandomised, controlled trial. SETTING: St Vincent's Hospital, Melbourne, Victoria (a tertiary referral, university teaching hospital), 1 October 1997 to 30 November 1998. PARTICIPANTS: 111 patients (80 women and 31 men; mean age, 81 years) admitted via the emergency department with a primary diagnosis of fractured neck of femur. INTERVENTIONS: Management guided by a clinical pathway (55 patients) or established standard of care (control group, 56 patients). MAIN OUTCOME MEASURES: Timing of referrals and discharge planning; total length of stay; and complication and readmission rates within 28 days of discharge. RESULTS: Patients managed according to the clinical pathway had a shorter total stay (6.6 versus 8.0 days; P = 0.03), even if assessment for placement by the Aged Care Assessment Service was required (9.5 versus 13.6 days; P = 0.03). There were no significant differences in complication and readmission rates between pathway and control patients (complication rates, 24% versus 36%; P = 0.40; readmission rates, 4% versus 11%; P = 0.28). CONCLUSION: Coordinated multidisciplinary care of patients with fractured neck of femur reduces length of stay without increasing complications. PMID- 10870535 TI - Distribution of colorectal adenomas: implications for bowel cancer screening. AB - OBJECTIVE: To determine the distribution of colorectal adenomas relative to the splenic flexure in an asymptomatic population undergoing colonoscopy, as an indicator of the number of patients with adenomas who would be missed by screening with flexible sigmoidoscopy. DESIGN: Retrospective survey of medical records. SETTING: Private endoscopy centres in Melbourne, Victoria. SUBJECTS: All 1131 asymptomatic individuals who underwent full colonoscopy between 1 January 1995 and 31 December 1997 after referral from a bowel cancer prevention program organised by the endoscopy centres. People referred were aged either 40 years or over with a first-degree relative with bowel cancer, or 50 years or over with marked anxiety about bowel cancer. MAIN OUTCOME MEASURES: Presence and distribution of colorectal adenomas. RESULTS: Polyps were found in 270 individuals (24%) and were confirmed to be adenomas in 138 (12%). These 138 comprised 106 men and 32 women, with mean age 54 years (range, 40-78 years). Most (86%) had a single adenoma. Position of adenomas in relation to the splenic flexure was: distal only in 85 of the 138 people (62%), proximal only in 34 (25%), and both distal and proximal in 19 (14%). CONCLUSIONS: In 25% of asymptomatic people found to have adenomas by this bowel cancer prevention program, the adenomas were found only in the proximal colon, well beyond the reach of the flexible sigmoidoscope. This distribution of adenomas suggests that screening programs cannot rely solely on flexible sigmoidoscopy. PMID- 10870536 TI - Management of a patient with a complete rupture of a main bronchus in a community hospital. AB - We describe the successful use of a portable extracorporeal membrane oxygenation machine for a patient with complete rupture of the left main bronchus after a road crash. The machine was used before and during left main bronchus reanastomosis at a community hospital 30 km from Melbourne, and then during acute interhospital transfer. PMID- 10870537 TI - Socioeconomic status and health in Australia. AB - Consistent with international evidence, the findings of Australian research show that socioeconomically disadvantaged groups experience significantly higher mortality and morbidity rates. Despite marked improvements in the health of all segments of the Australian population in recent decades, during this same period there has also been an increase in socioeconomically related mortality inequalities for some conditions. Socioeconomically disadvantaged groups are more likely to engage in health-damaging behaviours, experience poorer psychosocial health, make less use of the healthcare system for preventive purposes, and have a more adverse risk factor profile. These are the main contributing factors to the poorer physiological health of low socioeconomic groups. At present, our knowledge of how socioeconomic status and health are related is limited. A necessary step in improving our understanding of this issue is to draw together all the empirical evidence and use it as the basis for developing a theory of socioeconomic health inequalities. We present a conceptual framework to facilitate this process. PMID- 10870538 TI - The new health insurance rebate: an inefficient way of assisting public hospitals. AB - Private health insurance subsidy is now estimated to cost $2.19 billion; government support for private health care includes a further $1.2 billion of Medicare benefits expenditure in hospitals. The subsidy cannot be justified on efficiency grounds, as, on the basis of available evidence and taking casemix into account, public hospitals are more efficient than private hospitals. The original stated objective of the subsidy was to "take pressure off public hospitals". If the insurance subsidy and the Medicare Benefit Schedule rebate expenditure were applied to purchasing public hospital treatment at full average cost, 58% of current private sector demand could be accommodated. If 10% of the demand were met at marginal cost, this would increase to 65%. The objective of "taking pressure off public hospitals" could be more efficiently achieved by direct funding of public hospitals rather than through subsidies for private health insurance. PMID- 10870539 TI - Organisation of healthcare: challenging the "ego systems". PMID- 10870540 TI - Heparin-induced thrombocytopenia presenting after the cessation of low molecular weight heparin prophylaxis with enoxaparin. PMID- 10870541 TI - Headache and face pain. AB - A systematic case history is the primary diagnostic tool for headache problems. An acute ("thunderclap") headache or progressive subacute headache may have sinister implications and warrants urgent investigation. Common recurrent headaches (tension-type, migraine and cluster headaches) are not life-threatening but can destroy the enjoyment of life. Specific medications are available for acute treatment and prophylaxis for most types of recurrent headache. General practitioners should be well informed about management, as they are the most appropriate people to deal with most headache patients. PMID- 10870542 TI - Vancomycin-resistant enterococci and use of avoparcin in animal feed: is there a link? PMID- 10870543 TI - Training for rural general practice in north Queensland. PMID- 10870545 TI - Suicide trends: adolescence and beyond. PMID- 10870544 TI - Evaluation of the role of positron emission tomography in oncology. PMID- 10870546 TI - Rates of food poisoning in Australia. PMID- 10870547 TI - kConFab: a research resource of Australasian breast cancer families. Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer. PMID- 10870548 TI - Global environmental change. PMID- 10870549 TI - Diversity and correlation of specific aromatic hydrocarbon biodegradation capabilities. AB - This work investigated the biodegradation capabilities of indigenous microorganisms exposed to different combinations of aromatic hydrocarbons. Considerable diversity was found in the catabolic specificity of 55 strains. Toluene was the most commonly degraded compound, followed by p-xylene, m-xylene and ethylbenzene. Strains capable of degrading o-xylene and benzene, which were the least-frequently-degraded compounds, exhibited broader biodegradation capabilities. Kappa statistics showed a significant correlation between the abilities to degrade toluene and ethylbenzene, p-xylene and m-xylene, and p xylene and o-xylene. The ability to degrade naphthalene was correlated to the ability to degrade other alkylbenzenes, but not benzene. In addition, the inability to degrade benzene was correlated to the inability to degrade o-xylene. Factorial analysis of variance showed that biodegradation capabilities were generally broader when aromatic hydrocarbons were fed as mixtures than when fed separately. Beneficial substrate interactions included enhanced degradation of benzene, p-xylene, and naphthalene when toluene was present, and enhanced degradation of naphthalene by ethylbenzene. Such heuristic relationships may be useful to predict biodegradation patterns when bacteria are exposed to different aromatic hydrocarbon mixtures. PMID- 10870550 TI - Biodegradation of gasoline and BTEX in a microaerophilic biobarrier. AB - Continuous bioremediation of gasoline-contaminated water in a packed-bed biobarrier system under oxygen-limited conditions is discussed. This study was part of an extensive effort to develop an alternative technology for the in situ bioremediation of hydrocarbons where there is a limited supply of oxygen. Protruded stainless steel pieces and granulated peat moss were used as packing material to support microbial growth in two biobarriers. The inoculum was an enrichment culture of an indigenous microbial population from a soil sample. The biobarriers' inlet gasoline concentrations and the linear liquid velocities were similar to those commonly found at in situ conditions. Gasoline removal efficiencies ranged from 94% to 99.9% in the stainless steel-packed biobarrier, and from 86.6% to 99.6% in the peat moss-packed biobarrier. Effluent gasoline concentrations below 0.03 mg/l were obtained at gasoline loading rates less than 27.5 mg/l.d in the stainless steel-packed biobarrier. The remaining fraction of gasoline in the effluent consisted mainly of three aliphatic compounds and not the aromatic compounds. Both biobarrier packings supported near complete removal of the most soluble aromatic hydrocarbons of gasoline (BTEX) under all the conditions examined. The consumption of sulfate and the presence of sulfate reducing microorganisms suggested the presence of anaerobic metabolism during the degradation of gasoline. Up to 92% gasoline was removed during the first 3 cm of the biobarriers' length. PMID- 10870551 TI - Degradation of mono-chlorophenols by a mixed microbial community via a meta- cleavage pathway. AB - A mixed microbial community, specially designed to degrade a wide range of substituted aromatic compounds, was examined for its ability to degrade mono chlorophenols as sole carbon source in aerobic batch cultures. The mixed culture degraded 2-, 3-, and 4-chlorophenol (1.56 mM) via a meta- cleavage pathway. During the degradation of 2- and 3-chlorophenol by the mixed culture, 3 chlorocatechol production was observed. Further metabolism was toxic to cells as it led to inactivation of the catechol 2,3-dioxygenase enzyme upon meta- cleavage of 3-chlorocatechol resulting in incomplete degradation. Inactivation of the meta cleavage enzyme led to an accumulation of brown coloured polymers, which interfered with the measurement of cell growth using optical density. Degradation of 4-chlorophenol by the mixed culture led to an accumulation of 5-chloro-2 hydroxymuconic semialdehyde, the meta- cleavage product of 4-chlorocatechol. The accumulation of this compound did not interfere with the measurement of cell growth using optical density. 5-chloro-2-hydroxymuconic semialdehyde was further metabolized by the mixed culture with a stoichiometric release of chloride, indicating complete degradation of 4-chlorophenol by the mixed culture via a meta cleavage pathway. PMID- 10870553 TI - Biotransformation of lantadene A (22 beta-angeloyloxy-3-oxoolean-12-en-28-oic acid), the pentacyclic triterpenoid, by Alcaligenes faecalis. AB - A bacterial strain capable of biotransformation of lantadene A (22 beta angeloyloxy-3-oxo-olean-12-en-28-oic acid), the pentacyclic hepatotoxin of lantana (Lantana camara var. aculeata) has been isolated from soil using lantadene A as the sole carbon source. The organism is Gram negative, rod shaped, motile, catalase positive and has been identified as Alcaligenes faecalis. The isolate has been found to be specific for lantadene A and did not utilize lantadene B. In studies using sucrose as an additional carbon source, A. faecalis elicited biotransformation of lantadene A to its trans isomer 22 beta-tigloyloxy 3-oxoolean-12-en-28-oic acid, designated as lantadene X and two other minor metabolites which could not be isolated in pure state. PMID- 10870552 TI - Degradation of anaerobic reductive dechlorination products of Aroclor 1242 by four aerobic bacteria. AB - We studied the aerobic degradation of eight PCB congeners which comprise from 70 to 85% of the anaerobic dechlorination products from Aroclor 1242, including 2-, 4-, 2,4-, 2,6-, 2,2'-, 2,4'-, 2,2', 4-, and 2,4,4'-chlorobiphenyl (CB), and the biodegradation of their mixtures designed to simulate anaerobic dechlorination profiles M and C. Strains Comamonas testosteroni VP44 and Rhodococcus erythreus NY05 preferentially oxidized a para-substituted ring, while Rhodococcus sp. RHA1, similar to well known strain Burkholderia sp. LB400, preferably attacked an ortho chlorinated ring. Strains with ortho-directed attack extensively degraded 2,4'- and 2,4,4'-CB into 4-chlorobenzoate, while bacteria with para-directed attack transformed these congeners mostly into potentially problematic meta-cleavage products. The strains that preferentially oxidized an ortho-substituted ring readily degraded seven of the eight congeners supplied individually; only 2,6-CB was poorly degraded. Degradation of 2,2'- and 2,4,4'-CB was reduced when present in mixtures M and C. Higher efficiencies of degradation of the individual congeners and defined PCB mixtures M and C and greater production of chlorobenzoates were observed with bacteria that preferentially attack an ortho substituted ring. PCB congeners 2,4'-, 2,2',4-, and 2,4,4'-CB can be used to easily identify bacteria with ortho-directed attack which are advantageous for use in the aerobic stage of the two-phase (anaerobic/aerobic) PCB bioremediation scheme. PMID- 10870554 TI - A new species, Toxocara lyncis, in the caracal (Lynx caracal). AB - Toxocara lyncis, sp. n. is described from Lynx caracal in Somalia. It most closely resembles T. cati, the only species of Toxocara reported from L. caracal. It differs from T. cati in the comparative length of the spicules and the esophagus, and in the shape of the cervical alae. Cervical alae have a nearly uniform width along their length in T. lyncis, while they are narrow anteriorly and broad posteriorly forming an arrow head shaped cephalic end in T. cati. PMID- 10870555 TI - The parasites of the badger (Meles meles) in the north of Mugello (Florence, Italy). AB - During the period January 1993-June 1994, a parasitological survey was carried out on 19 badgers (Meles meles) road killed in Northern Mugello (Florence). The following helminths (together with their prevalence) were isolated and classified: Uncinaria criniformis (84.2%); Capillaria sp. (31.6%); Molineus patens (21.1%); Mesocestoides melesi (21.1%); Aelurostrongylus falciformis (52.6%); Crenosoma melesi (21.1%). According to results, only sex related differences in prevalence were studied. The parasite biocenosis is composed exclusively by dominant and codominant species. Mesocestoides melesi represents the first record for Italy. All the species found fitted a negative binomial distribution. PMID- 10870556 TI - [Four species of mosquito as possible vectors for Dirofilaria immitis piedmont rice-fields]. AB - Wild mosquitoes were allowed to feed, during two nights, on a heartworm-infected dog with a very high microfilaraemia (72,000 and 78,000 microfilariae/ml just prior to be allocated in the live trap). A heartworm-free dog was used as control bait in the second night. Engorged mosquitoes (Aedes caspius, Anopheles maculipennis s.l., Culex modestus, and Cx pipiens) were kept under laboratory conditions, artificially fed until day 10 post-infection (PI) and then dissected for the presence of Dirofilaria immitis larvae. Mortality of Ae. caspius and Cx modestus was significantly higher than controls on day 3 PI (89.4 and 80.3%, respectively), but survival rates were similar in the following period. Third stage larvae were observed from day 12 to 17 PI in the four mosquito species studied. However, vector efficiency was significantly higher in Ae. caspius which produced 102 (73.9%) of the 138 infective larvae found. Although less efficient vectors, the other 3 species may contribute to D. immitis transmission in the study area due to their abundance (Cx modestus) or higher resistance to the negative effects of infection (An. maculipennis s.l., Cx pipiens). As far as Cx pipiens is concerned, this mosquito, which luckily fed the least frequently on the dog, confirmed to act as powerful vector in iperendemic areas. The risks for veterinary and medical health, related to the zoo-antropophylic host-feeding pattern of the studied species, are discussed. PMID- 10870557 TI - [Report in Fruili-Venezia Giulia about Aedes cataphylla, Ae. communis, and Ae. pullatus, three rare mosquitoes in Italy]. AB - Data are provided on the ecology and morphology of Aedes cataphylla, Ae. communis e Ae. pullatus (Diptera, Culicidae), rare species for Italy, collected in the Friuli-Venezia Giulia Region (North-eastern Italy). PMID- 10870558 TI - Occurrence of Thelazia lacrymalis (Nematoda, Spirurida, Thelaziidae) in native horses in Italy. AB - A survey on the prevalence of Thelazia spp. in the province of Bari (Apulia region, Italy) in slaughtered native horses was conducted from June 20, 1995 to April 3, 1996. Both eyes from 409 ten-month- to 4-year-old native animals were examined. Sixty horses (14.7%) were found parasitized by Thelazia lacrymalis. Three hundred-sixty one parasite specimens (220 females, 99 males and 42 larvae) were collected with a mean count burden of 6.0-5.1 (range 1 to 20) per head. T. lacrymalis specimens were found free in the conjunctiva and behind the nictitancte, in the excretory ducts of the Harderian gland and in the ducts of the lacrimal glands. Thelazia specimens were significantly more common in horses of 3-4 years. Gross examination of all infected animals showed a follicular conjunctivitis. This is the first report of T. lacrymalis horse infection in Italy. PMID- 10870559 TI - [Spread of hemoparasites in reptiles and amphibians in Italy]. AB - Hemoparasites were harvested from 993 individuals belonging to 15 reptilian and 1 amphibian species, from various Italian localities. Hemogregarins were found in 10 reptilian species while a flagellate and microfilariae were found only in Tarentola mauritanica from Lampedusa. For each host species and place of origin the frequencies of hemogregarins are reported and discussed. Longitudinal studies with periodical thin smears were carried out on 5 Tarentola mauritanica, 4 Lacerta viridis, 26 Podarcis filfolensis, 10 Podarcis muralis, 38 Podarcis sicula, 8 Chalcides ocellatus. This material, whose study has not yet been completed, is made available by the author who strongly encourages further investigations on this subject. PMID- 10870560 TI - Tick species parasitizing people in an area endemic for tick-borne diseases in north-western Italy. AB - In 1995 and 1996, 318 ticks were recovered from 240 people in Liguria (Province of Savona, Italy). Most of the ticks (284; 89.3%) were sheep ticks, Ixodes ricinus. Rhipicephalus sanguineus (31; 9.8%) and Dermacentor marginatus (3; 0.9%) were also recorded. All three life stages of I. ricinus were found on humans while only nymphs and adults of R. sanguineus and adults of D. marginatus were collected. Human tickbites were most frequent in the municipalities of the Province where roe deer density was highest. The number of tickbites in this area was about 500 per 100,000 residents over the period of observation. Most cases of tickbite were observed in children (11.2%), students (25.6%), workers (22.4%) and retired people (24%). Although ticks were recovered from people throughout the year, the highest frequencies of I. ricinus bites were in May, June and July. In the study area natural hosts were also studied. Six tick species were identified (I. ricinus, I. hexagonus, D. marginatus, R. bursa, R. sanguineus, R. turanicus). I. ricinus and R. sanguineus were the most abundant species. I. ricinus was recorded more frequently from ruminants particularly roe deer, while R. sanguineus was found to be associated with the dog. PMID- 10870561 TI - Intensity of nematode infections in cyclic and non-cyclic rock partridge (Alectoris graeca saxatilis) populations. AB - Populations of rock partridge (Alectoris graeca saxatilis), in the Trentino province of Italy, exhibit cyclic fluctuations in abundance associated with relatively dry habitat. One of the hypothesis to explain these cycles is that survival of some free living parasitic stages and rates of infection are greater in these areas leading to higher parasite burden. This hypothesis was examined by investigating the intensity of parasite infection in cyclic and non cyclic rock partridge populations. Analyses of 87 intestine samples collected from shot rock partridges during 1994 and 1995 identified 8 species of helminths parasites: Ascaridia compar (P = 33.33%; I = 9.28 +/- 1.78), Heterakis tenuicauda (P = 19.54%; I = 10.29 +/- 4.58), Heterakis gallinarum (P = 1.15%; I = 1.0 +/- 0.0), Heterakis altaica (P = 1.15%; I = 17 +/- 0.0), Aonchoteca caudinflata (P = 6.89; I = 2.17 +/- 0.65), Postharmostomum commutatum (P = 5.75; I = 7.0 +/- 3.48), Brachylaema fuscata (P = 1.15; I = 7.0 +/- 0.0), Platynosomum alectoris (P = 2.29; I = 5.5 +/- 1.5). Cestoda, recorded with a prevalence of 5.75, were not identified to species level. A. compar and H. tenuicauda were prevalent in the rock partridge populations and there was no positive association between these species. Intensity of infection in both species was not influenced by host age, sex or year of study but levels of infection with A. compar burdens were significantly greater in cyclic populations than in non cyclic populations and there was a tendency for H. tenuicauda to be greater in cyclic populations. There was no negative relationship between intensity of infection with A. compar or H. tenuicauda and host body mass. These data provide some support for the hypothesis that these parasites may play a role in generating rock partridge population cycles. PMID- 10870562 TI - Effect of seasonality on abomasal helminth community in alpine ibex (Capra ibex ibex). AB - Abomasal helminths of 81 adult female ibex (Capra ibex ibex) from Piz Albris colony (CH) culled monthly from December 1989 to May 1991 were analyzed. Subjects were divided into six quarterly groups (I: December 1989-February 1990; II: March May 1990; III: June-August 1990; IV: September-November 1990; V: December 1990 February 1991; VI: March-May 1991) in order to investigate the seasonal fluctuation of helminth community parameters (abundance, richness, sex-ratio, Berger-Parker dominance index, importance index, overall association between species by Schluter's variance test). All parameters showed considerable seasonality. Abundance was highest in group III, while richness was highest and Berger-Parker index lowest in group IV. Overall associations between species were positive in winter-spring groups and negative in summer-autumn. The importance index of the two dominant species (Teladorsagia circumcincta and Marshallagia marshalli) showed a symmetrical and opposite seasonal development. A significant negative correlation (Spearman r = -.30; p < .01) has been demonstrated between the Berger-Parker dominance index and factor scores descriptive of age and weight of the subjects. These results suggest that seasonal factors determine the community composition while host control can act as a filter maintaining a stable community structure. PMID- 10870563 TI - Pentastomes (Pentastomida, Armillifer armillatus Wyman, 1848) in snakes from Zambia. AB - Twenty-three snakes, belonging to eight different species, were collected from rural areas of Zambia and inspected for the presence of pentastomes. Pentastomid parasites were found in four snakes: one African rock python (Python sebae), one puff adder (Bitis arietans) and two Mozambique spitting cobras (Naja mossambica) were infested with a small number of Armillifer armillatus, respectively five, two and one adult parasites. As humans can be incidental/intermediate hosts for reptilian pentastomes, the zoonotic potential of these parasites, especially in tropical countries, is discussed. PMID- 10870564 TI - Preliminary results on Digenea found in fishes in the coastal waters of Terra Nova Bay, Antarctica. AB - A list of fish parasites (Digenea) from the coastal waters of Terra Nova Bay (Ross Sea, Antarctica) concerns 10 identified species, namely Macvicaria pennelli, Neolebouria terranovaensis, Helicometra pisanoae, Lepidapedon balgueriasi, L. garradi, Genolinea bowersi, Derogenes johnstoni, Gonocerca phycidis, Elytrophalloides oatesi, Lecithaster macrocotyle, 1 genus Aporocotyle sp. and one specimen undetermined. The total number of species known from the Ross Sea increased from 9 to 12. This number is smaller than those of other Antarctic areas, such as Western Antarctic and Weddell Sea where more extensive investigations were carried out to date. PMID- 10870566 TI - A case of myiasis in man due to Wohlfahrtia magnifica (Schiner) recorded near Rome. AB - In this report the case of a 14-year-old Italian boy is described in whom scratching wounds of the scalp were invaded by maggots of the dipterous Wohlfahrtia magnifica (Schiner). PMID- 10870565 TI - [Research on antifungal activity of flowers and leaves of Inula viscosa (Asteraceae)]. AB - The authors carried out a preliminary screening about the in vitro antifungal activity of some extracts of flowers and leaves of Inula viscosa obtained with different solvents. All extracts showed antifungal activity against dermatophytes and Candida species. The best results were obtained with Inula viscosa flowers extracts. These results may be ascribed to the different flavonoids and different flavonoid concentrations in our samples. PMID- 10870567 TI - Antifungal activity of Apulia region propolis. AB - A study was carried out to assess the in vitro antifungal activity of some natural Apulian propolis extracts of different origin. Their antifungal activity was compared to the antifungal activity of conifers and commercial propolis extracts. All extracts revealed antifungal activity against dermatophytes and Candida species. The antifungal activity differences found depended on the origin of the propolis and the solvent used for extraction. The best antifungal activity was given by the 'Orimini' propolis. The antifungal activity may have been influenced by the presence of different cinnamic and flavonoid components and their different concentration in the extracts. Further investigations are needed to validate this hypothesis. PMID- 10870568 TI - Diagnosis of a case of gastric anisakidosis by PCR-based restriction fragment length polymorphism analysis. AB - A set of genetic markers, based on PCR-RFLPs of three diagnostic restriction enzymes (Hhal, Hinfl and Taql), which proved to be suitable for the identification of the species of the genus Anisakis, was used for the first molecular identification of a larva obtained by endoscopy in a case of gastric anisakidosis, in a 51 year old woman from Southern Italy. The analysis of the restriction profiles obtained allowed the larva to be identified as Anisakis pegreffii, one of the three sibling species of the A. simplex complex. PCR-RFLP proved to be a cost-effective and reliable tool for the exact identification of Anisakis larvae recovered from infected humans. PMID- 10870569 TI - Chromosomal and bionomic heterogeneities suggest incipient speciation in Anopheles funestus from Burkina Faso. AB - Sampling of day-resting Anopheles funestus was carried out in September-November 1991, October-December 1992, and November 1994 at two sites near Ouagadougou, Burkina Faso: the small village of Noungou where humans outnumber cattle, and the nearby Fulani settlement of Loumbila where cattle outnumber humans. Collections made inside human dwellings were supplemented in 1992 by outdoor-resting samples from artificial pit-shelters. Indoor-resting An. funestus were also collected in November 1992 and November 1994 in four villages of the Banfora area (southern Burkina Faso) and in a sudanese-sahelian village in northern Burkina Faso (Tougouri). Half-gravid female sub-samples were preserved in carnoy's fixative and processed for polytene chromosome analysis. The material from the two villages near Ouagadougou was analysed by ELISA to know (i) the human/animal origin of the blood meal; (ii) the infectivity for Plasmodium falciparum malaria; and (iii) the possible correlation of these parameters with chromosomal variants. A total of 1416 An. funestus could be scored for the whole polytenic complement, while the origin of the blood meal and circumsporozoite protein (CSP) positivity were asserted from 1076 and 1154 specimens, respectively. With a few exceptions, four polymorphic paracentric inversions (3a, 3b, 2a and 5a in decreasing order of frequency) were observed in all populations. Inversion 2s, whose breaking points include those of inversion 2a, was found only as the heterokaryotype 2s/+ floating at an overall frequency of 3.7% in two villages of the Banfora area and in the two sites near Ouagadougou. Two heterokaryotypes 2a/t out of 186 scored specimens were observed in different years from one village of the Banfora area. Wide variations in inversion frequencies were observed among the samples without consistent geographical or temporal clines. Highly significant departures from Hardy-Weinberg equilibrium were recorded for inversions 3a and 3b in most samples, with the alternative homokaryotypes (standard and inverted) significantly more frequent than expected. Conversely, inversion 5a was in Hardy Weinberg equilibrium in most samples, whereas the 2a-s inversion system was intermediate between these extremes. However, a deficit of heterokaryotypes was apparent practically in all samples. Significantly higher frequencies of the standard homokaryotypes were recorded (i) in the exophilic samples collected in Loumbila for arrangement 3a; (ii) in the animal-fed sub-sample collected outdoors in Noungou vs. the parallel human-fed sub-sample for arrangements 2a-s, 3a, and 3b, or vs. the samples obtained from indoor catches in both Loumbila and Noungou in the case of inversion 3a; (iii) in the December CSP-negative sub-sample from Loumbila vs. the parallel CSP-positive sub-sample for arrangement 2a. A plausible working hypothesis is that An. funestus in Burkina Faso includes two taxonomic units, one of which is mainly monomorphic standard with most inverted arrangements floating at very low frequencies, and probably uniquely characterised by arrangement 2s, while the other taxon is nearly fixed for arrangement 3ab and polymorphic for all the other inversions at intermediate to high frequencies. The latter would be characterised by a higher vectorial capacity and would probably correspond to An. funestus s.s. from East Africa. About the former hypothetical taxon, its endophily and anthropophily appear less marked and its relationship with other members of the An. funestus subgroup will require specific investigations. PMID- 10870570 TI - Preliminary observations on entomopathogens of phlebotomine sandflies in Pakistan. PMID- 10870571 TI - Surgery of aortic aneurysms and brain protection. PMID- 10870572 TI - Advances in the development of immunosuppressive agents in organ transplantation. PMID- 10870573 TI - Gastric carcinoids: a statistical evaluation of 1,094 cases collected from the literature. AB - Gastric carcinoids are a rare gut endocrinoma, and only a few series dealing with limited aspects have been published. This study evaluates the present status and characteristics of gastric carcinoids in a statistically reliable series of 1,094 cases that were carefully evaluated, computerized, and analyzed by the "Gut Pancreatic Endocrinoma Analyzing System." Routine statistical analysis was carried out on 1,011 patients, excluding 83 with atypical carcinoids, focusing on clinical manifestations, location, depth, and size of the lesions in relation to metastases, immunohistochemistry, carcinoid syndrome, serotonin activity, electron microscopy, multicarcinoid complex with type A gastritis, and postoperative outcome. A tumor size of 20 mm or less comprised 60.8% of the series, with a metastasis rate of 15.1%, and depth of invasion to the submucosa occupied 53.8%, with a metastasis rate of 13.2%. Carcinoid syndrome was encountered in 4.0% of the patients. Elevated serotonin activity was detected in 22.3% overall and in 67.7% of the patients with carcinoid syndrome (P < 0.01). Multicarcinoid complex with type A gastritis was detected in 140 of 347 patients with multicarcinoid complex, and 97.1% had associated lesions in the nonantral regions of the stomach. A comparative evaluation between patients with and without type A gastritis indicated a number of significant differences including male to female ratio, age distribution, location, tumor size, depth of invasion, metastasis, and prognosis. PMID- 10870574 TI - The impact of lymph node metastases on the survival of breast cancer patients with ten or more positive lymph nodes. AB - To investigate the impact of the number of involved lymph nodes on survival, we retrospectively reviewed the data for 37 patients with breast cancer and metastases of ten or more lymph nodes who underwent treatment between 1987 and 1995. Based on the number of positive lymph nodes, the patients were allocated to one of three groups. The 5-year disease-free and overall survival rates for all patients were both 53.0%. The 7 patients with 26 or more positive nodes had significantly poorer survival than either the 19 patients with 10-15 nodes, or the 11 with 16-25 nodes, although there were no differences in survival related to the extent of node involvement as defined using the Japanese staging system. Patients with 50%-75% frequency of metastasis, defined as the positive nodes/total resected nodes, had significantly better survival than those with < 50% or > 75% frequency. These results indicate that the number of involved lymph nodes is related to survival and that 25 positive nodes is a cutoff point in breast cancer patients with ten or more positive lymph nodes. PMID- 10870575 TI - Acute acalculous cholecystitis after cardiovascular surgery. AB - The development of acute acalculous cholecystitis (AAC) after cardiovascular surgery is an infrequent but devastating complication, the etiology and management of which remains controversial. To evaluate the etiology, treatment, and outcome of patients with AAC, the cases of six patients encountered within an 8-year period who developed AAC after cardiovascular surgery requiring cardiopulmonary bypass (CPB) were reviewed. Atherosclerotic risk factors including diabetes, hyperlipidemia, and smoking were evident in five patients, three of whom had a history of stroke or arteriosclerosis obliterans, while low cardiac output was recognized in three. Percutaneous transhepatic cholecystostomy was performed in five patients, and another required cholecystectomy for peritonitis due to gangrene of the gallbladder. Two patients died of respiratory failure and sepsis after 15 and 82 days of percutaneous drainage, respectively; however, the four survivors had an excellent outcome without any biliary tract disease during a mean follow-up period of 5.3 years. In conclusion, AAC after cardiovascular surgery may result from hypoperfusion of the gallbladder due to various factors including CPB, visceral atherosclerosis, and low cardiac output. We advocate early percutaneous cholecystostomy for patients without peritonitis, while early cholecystectomy is indicated for those with peritonitis. PMID- 10870576 TI - The management of patients with dissection of the descending thoracic aorta: a comparison between closing and nonclosing dissections. AB - This study was designed to clarify and compare the clinical characteristics and prognoses of patients with closing and nonclosing dissection of the descending thoracic aorta. Between January 1991 and December 1994, 19 patients with closing dissection (Group A) and 20 with nonclosing dissection (Group B) underwent surgical repair or medical treatment at our institution. There were 29 men and 10 women, aged between 37 and 74 years, with a mean age of 62 years. There was a significant difference in age between the two groups, being 67 +/- 7 and 58 +/- 12 years for Groups A and B, respectively (P = 0.009). The presence of a concurrent abdominal aortic aneurysm was confirmed in 32% and 10% of Groups A and B, respectively (P = 0.095). A total of 15 patients experienced a variety of complications related to the dissection, but there were no significant differences in the morbidity rate between the two groups. Visceral ischemic disorders such as renal failure, leg ischemia, and ileus were the most common complications. The overall survival rate 4 years after the development of dissection was 80%, with no significant difference between the two groups. These findings led to the establishment of our policy to place all patients with dissection of the descending thoracic aorta on careful antihypertensive therapy and frequent follow-up imaging studies to assess the aorta, regardless of the condition of the false lumen. PMID- 10870577 TI - Basic studies on the application of an artificial esophagus using cultured epidermal cells. AB - In making an artificial esophagus, the transplantation of the epithelialized granulation tube fabricated by organized synthetic material was studied mainly from the viewpoint of preventing anastomotic leakage and stricture formation. The possibility of epithelialization of the inner surface of a granulation tube using cultured epidermal cells was studied in rats and dogs. A stainless steel mesh tube coated with silicon served as the granulation tube. Epithelialization on the inner surface of a granulation tube was evaluated by seeding cultured epidermal cells. A skin sample was treated with dispase and trypsin to collect epidermal cells, which were cultured in a keratinocyte growth medium. Once confluence was achieved, the epidermal cell suspension was harvested using the following methods: trypsin treatment (n = 15), mechanical separation with a cell scraper (n = 6), and dispase treatment (n = 9). The cultured epidermal cell suspension was then seeded into the lumen of the granulation tubes. The attachment of cultured epidermal cells was attained in 2 of 15 cases by trypsin treatment, and in 5 of 9 cases by dispase treatment. No attachment occurred using the cell scraper method. All attached epidermal cells exhibited a cobblestone appearance on the granulation tissue with a tendency toward stratification. These findings show that the inner surface of a steel mesh granulation tube was epithelialized by cultured epidermal cells. PMID- 10870578 TI - High submucosal blood flow and low anastomotic tension prevent anastomotic leakage in rabbits. AB - In many cases of long-gap congenital esophageal atresia, direct anastomosis is difficult. In these cases, the esophagus is first lengthened by myotomy before anastomosis. We determined the degree of submucosal blood flow and/or approximation force at the site of anastomosis in rabbits after (1) separation of the esophagus from the outer membrane, (2) 1 cm and 2 cm resection of the esophagus, and (3) circular or spiral myotomy of the esophagus after 2 cm resection. In the first experimental group, submucosal blood flow volume < 115.2 ml/min/100 g resulted in anastomotic leakage. In the second experimental group, a 1 cm resected esophagus with an approximation force of 33.3 +/- 8.2 g did not result in leakage, while a 2 cm resected esophagus with an approximation force of 111.7 +/- 13.3 g resulted in leakage. It was found that leakage occurred when the approximation force was higher than 49.1 g even if submucosal blood flow volume was greater than 131.8 ml/min/100 g. In the third experiment, both circular and spiral myotomy reduced the approximation force. Although there was no difference in the changes in submucosal blood flow volume between the two types myotomy, circular myotomy was superior to spiral myotomy in the reduction of the approximation force at the site of anastomosis. We conclude that both approximation force and submucosal blood flow are important factors in preventing anastomotic leakage. PMID- 10870579 TI - A new hydroxyl radical scavenger "EPC" on cadaver heart transplantation in a canine model. AB - This study was performed to determine if an "arrested" heart, resuscitated with cardiopulmonary bypass (CPB) after the cessation of beating, can be successfully transplanted, and whether a hydroxyl radical scavenger EPC can reduce ischemic and reperfusion injury during resuscitation of the arrested heart and following orthotopic heart transplantation. A total of 16 pairs of canines were divided into a control group of eight pairs and an EPC-treated group of eight pairs. Cardiac arrest of the donor heart was induced by the discontinuation of respiratory support after the induction of brain death. The cadaver heart was then resuscitated and core-cooled to myocardial temperature of 15 degrees C using CPB. The donor heart was harvested using cold cardioplegia and orthotopically transplanted. All of the transplanted hearts in the EPC group were weaned from CPB without any inotropic support after 60 min of bypass support, whereas all the animals in the control group required 5 micrograms/kg/min dopamine (P = 0.001). Moreover, cardiac function (Emax) 1 h after orthotopic heart transplantation was better preserved in the EPC group than in the control group, at 110 +/- 36% vs. 70 +/- 21% of the post brain death values (P = 0.02) These findings demonstrate that EPC reduces posttransplant reperfusion injury, and thus it may prove to be a valuable adjunct in this challenging model. PMID- 10870580 TI - The release of nitroglycerin absorbed into the central venous catheter. AB - This study was conducted to evaluate the release of nitroglycerin (NG) that has been absorbed into the central venous catheter. A 0.05% NG solution was infused through a central venous catheter and the flow rates were set at 1, 5, or 10 ml/h, given over 12, 24, or 48 h. The catheter was flushed with lactate Ringer solution after completion of the NG infusion. The elution of the lactate Ringer solution from the tip of the catheter was then collected and assayed for its NG concentration by high performance liquid chromatography (HPLC). A higher concentration of NG was released with a faster flow rate and a longer infusion. The high level of NG release continued during the first 20 min, and ranged from a minimum of 0.07 mg/ml to a maximum that exceeded 0.15 mg/ml. Subsequently, the NG concentration gradually declined, but low concentrations of 0.006-0.02 mg/ml were still maintained 360 min later. Thus, it is suggested that if a catheter such as the Swan-Ganz continues to be used after the completion of a NG infusion, certain pharmacological effects due to the absorption of NG into the catheter body should be expected for at least 60 min. PMID- 10870581 TI - The effects of proteoglycan on GALT in rats treated with TPN. AB - It is well known that total parenteral nutrition (TPN) causes atrophy of the intestinal mucosa, resulting in degeneration and atrophy of the gut-associated lymphoid tissues (GALT). This study was conducted to examine the suppressive effect of TPN on GALT in rats. Rats that received TPN alone for 2 weeks, i.e., the TPN group, showed a decreased number of Peyer's patches and thoracic duct lymphocytes (TDL), as well as atrophy. Conversely, those treated with TPN in combination with polysaccharide K (PSK) at a daily dose of 1000 mg/kg for 2 weeks, i.e., the PSK group, showed increases in the number of Peyer's patches and TDL and improvement in the TDL subsets compared with the TPN group. Immunohistological examination of the changes in immunocytes in GALT using monoclonal antibodies revealed increases in the production of the major histocompatibility complex (MHC)-I and (MHC)-II, helper T cells, and interleukin 2 (IL-2). These findings indicate that PSK improves GALT suppression induced by TPN. PMID- 10870582 TI - The beneficial effects of recombinant human insulin-like growth factor-I (IGF-I) on wound healing in severely wounded senescent mice. AB - The effects of recombinant insulin-like growth factor I (rIGF-I) on wound healing were tested using senescent and young BDF-1 mice, aged 108 weeks and 10 weeks, respectively. After inflicting a full thickness dermal burn encompassing 15% of the body surface, a skin incision, 2 cm in length, was made in the back. A silicone tube containing a piece of polyvinyl sponge was then implanted into a subcutaneous pocket in the flank to collect body fluid. An osmotic pump was buried in the abdominal subcutaneous tissue for the continuous infusion of rIGF I, the control being treated with the solvent of IGF-I, physiological saline, only. The administration of IGF-I produced favorable effects on wound healing in the senescent mice, shown by enhanced tensile strength and an elevated concentration in the hydroxyproline of the polyvinyl sponge content. The IGF-I treated severely wounded senescent mice healed better than their counterparts and their skeletal muscles contained more glutamine. Furthermore, they showed more enhanced cutaneous hypersensitivity towards dinitrofluorobenzene than the controls, suggesting an enhanced grade of cellular immunity. There were no conspicuous differences between the two groups of young mice. These data may suggest the beneficial effects of rIGF-I on wound healing, especially in geriatric surgery. PMID- 10870583 TI - The effect of oral sodium taurocholate on endotoxemia and intestinal anastomotic wound healing in rats with obstructive jaundice. AB - The effect of sodium taurocholate (ST) on endotoxemia and intestinal anastomotic wound healing in obstructive jaundice was evaluated in a rat model. A total of 108 Wistar rats were divided into three main groups. Thus, 36 animals were given ileal anastomosis (IA) alone (IA group), 36 were given IA with bile duct ligation (BDL) (IA + BDL group), and 36 were given IA with BDL and oral sodium taurocholate (ST) (IA + BDL + ST group). These three main groups were then divided into three equal subgroups, A, B, and C, which were killed on postoperative days (POD) 3, 5, and 9, respectively. In the IA + BDL + ST group, ST was administrated perioperatively and ceased from POD 5 onwards. The anastomotic hydroxyproline level and bursting pressure were significantly lower in the IA + BDL animals compared with the others on POD 3, 5, and 9 (P < 0.008). Endotoxemia was prominent in the IA + BDL group from POD 3 (P = 0.011). After ST was stopped, 42% of the AI + BDL + ST animals developed endotoxemia by POD 9 (P = 0.008). Anastomotic wound healing was better in the IA + BDL + ST group (P < 0.01). These findings suggest that endotoxemia and its adverse effects on wound healing in obstructive jaundice can be prevented by the oral administration of ST. PMID- 10870584 TI - Development of black gallstones after the nonsurgical management of splenic injury: report of a case. AB - A 22-year-old man was admitted to our Emergency Department after suffering splenic injury in a traffic accident. His intraabdominal bleeding was treated nonsurgically by the administration of total parenteral nutrition (TPN) and blood transfusions of packed red cells. He presented again 2 months after his discharge, being 3 months after the injury, for right hypochondralgia, at which time a gallstone was demonstrated on ultrasound (US) and computed tomography (CT). After endoscopic laparoscopic cholecystectomy, his symptoms disappeared and he has remained well since. The clinical course of this patient indicates that hemolytic hyperbilirubinemia can cause black gallstones as a late complication of the nonsurgical management of abdominal blunt trauma. PMID- 10870585 TI - Thyroid cancer in children: report of three cases and a review of the Japanese literature. AB - We experienced three cases of thyroid cancer in children less than 15 years of age between 1982 and 1995. We herein present these three cases with a review of 141 reported cases of childhood thyroid cancer in Japan. Our patients were 6, 13, and 14 years old. The patients, all girls, were diagnosed as having thyroid cancer based on diagnostic imaging. One of them was also diagnosed by a fine needle aspiration biopsy (FNAB). One of them underwent subtotal thyroidectomy, and the other two underwent lobectomy. Modified neck dissections were performed on all three. Pathologically, the tumors were all papillary carcinomas. Multiple lymph node metastases were present in all patients. However, the postoperative courses have been good, and there have been no signs of recurrence, 10, 8, and 2 years after their respective operations. In 144 reported cases of childhood thyroid cancer in Japan including ours, the youngest patient was a 2-year-old boy, and the female to male ratio was 2.1:1. FNAB was performed in 25 cases, and 23 (92%) of the tumors were diagnosed as malignant. Histologically, 76% were papillary carcinoma and 20% follicular carcinoma. At operation, lymph node metastases were found in 80% of the cases, and lung metastases in 17%. For treatment, 88% of the patients received a more extensive operation than a lobectomy. Of the 144 patients, 8 died. PMID- 10870586 TI - Coronary artery fistula into a persistent left superior vena cava: report of a case. AB - We herein report the rare case of a patient with coronary artery fistula (CAF) between the left circumflex coronary artery and persistent left superior vena cava (PLSVC) with a complete absence of the right superior vena cava (SVC). PMID- 10870587 TI - Iatrogenic paraplegia caused by surgicel used for hemostasis during a thoracotomy: report of a case. AB - A 46-year-old Japanese woman underwent a right lower lobectomy through a posterolateral incision made in the fifth intercostal space under general and epidural anesthesia on January 23, 1995. During the procedure, oxidized regenerated cellulose (Surgicel) was used to prevent postoperative rebleeding from the dorsal branch of the fifth intercostal artery. The following day it became evident that complete paraplegia had developed below the Th5 level, the cause of which was revealed by an emergency laminectomy, performed within 20 h after the thoracotomy, to be the Surgical treatment. By 50 days after the operation the patient had begun to show improvement, and was able to move her lower extremities against gravity. Her condition is continuing to improve. PMID- 10870588 TI - Mediastinal extraadrenal myelolipoma: report of a case. AB - We herein report a case of surgically resected mediastinal extraadrenal myelolipoma. Myelolipoma is an uncommon tumor composed of adipose tissue and normal hematopoietic elements, and is most often found in the adrenal glands. We could find only five such cases of mediastinal myelolipoma in the English literature. PMID- 10870589 TI - Clinical significance of the DNA index in hematogenous dissemination of aneuploid colorectal cancer. AB - The DNA index (DI), defined as the ratio of the G0/G1 channel number of tumor cells to the G0/G1 channel number of stromal cells, was analyzed in 121 aneuploid colorectal cancers. In patients with aneuploid tumors, a high DI significantly correlated with tumor penetration beyond the proper muscle layer, positive vascular invasion, and the presence of liver metastasis. This observation led us to conclude that a high DI is likely to be associated with hematogenous dissemination in aneuploid colorectal cancer. PMID- 10870590 TI - Fissurectomy with posterior midline sphincterotomy and anoplasty (FPSA) in the management of chronic anal fissures. AB - Although lateral internal sphincterotomy is widely accepted as the treatment of choice for anal fissures, we report our experience of successfully treating 195 consecutive patients with posterior chronic anal fissures by performing fissurectomy with midline sphincterotomy and anoplasty (FPSA). The surgical technique is described and its indications and results are briefly discussed. PMID- 10870591 TI - AIDS and lipodystrophy: complications of HIV and HIV treatments. PMID- 10870592 TI - Short-term human growth hormone therapy for HIV-associated wasting. PMID- 10870593 TI - Altered body shape in HIV disease: a side effect of therapy? AB - An interactive teleconference on HIV-associated lipodystrophies and altered body habitus was held on January 20, 1999, part of a series known as HIV Treatment LIVE!. It was moderated by Ronald Baker, Ph.D., and sponsored by the AIDS Wasting Foundation (AWF). AWF, located at 15 East 26th Street, New York, NY 10010, was founded in 1997 as an education and advocacy organization, under the direction of Carola Burroughs, Executive Director. This tele-conference brought together three AIDS experts to address important issues concerning the etiology, diagnosis, and treatment of this novel and enigmatic syndrome. The resulting report is an edited, selected transcript of that discussion. PMID- 10870594 TI - Willingness to treat HIV-positive patients at different stages of medical education and experience. AB - The willingness of physicians to provide care to HIV-positive patients has been linked to a number of attitudinal factors, but little is known concerning the impact of premedical, medical, and residency training on these factors. The purpose of this study is to elicit responses to the same series of questions concerning HIV and its treatment from respondents at different stages of training, to detect trends in attitudes and to measure the impact of those attitudes on willingness to provide care for HIV/AIDS patients. Study data come from a cross-sectional survey (n = 249) of respondents across the training continuum, from premedical students to faculty physicians, using a self administered questionnaire at a single medical school. The response rate was 59.6%. The study showed significant decreases in personal fear and misgivings concerning HIV, coupled with a substantial decrease in the perceived need for testing of non-high-risk individuals, as respondents gained additional education and training. Overall, the intent to treat HIV did not change significantly by training level, but multivariate analyses showed that while the initially strong influence of attitudes toward AIDS and its attendant risks diminishes, comfort relative to being around homosexuals per se continues to exert an impact on the intent to treat. Appropriate use of protective measures when providing care becomes far more common once individuals enter their clinical training years. The impact of medical education through its entire continuum therefore shows a positive impact on attitudes toward HIV, despite the absence of a significant trend in respondents' stated intent to treat. However, negative attitudes toward homosexuals continue to exert a negative influence on intent to treat that endures into the clinical training years. PMID- 10870595 TI - Pathogenesis of non-AIDS-defining cancers: a review. AB - As the AIDS epidemic advances, the number of HIV-infected subjects developing AIDS-related neoplasms is rapidly increasing, and the spectrum of malignancies encountered is expanding. Several non-AIDS-defining cancers are being reported at an increasing incidence in HIV-infected individuals, including anal, skin, oral mucosa, head and neck and lung carcinomas, testicular tumors, and pediatric soft tissue sarcoma. There appears to be an emerging role for various concurrent viral infections in the HIV-infected host that are likely implicated in the pathogenesis of some nondefining-AIDS neoplasms. Our recent findings in HIV associated lung cancers and in the precursor lesions of cervical carcinoma suggest that wide-spread genomic instability, as manifested by the development of increased numbers of microsatellite alterations (MAs), may occur frequently in HIV-associated tumors and they may play an important role in the pathogenesis of those neoplasms. Although the mechanism underlying the development of increased MAs is unknown, it may play a crucial role in the development of many HIV associated tumors. It will be important to track the epidemiological and biological features of non-AIDS-defining cancers in HIV-infected patients, and compare them to those tumors in the general population. It is likely that further clues about malignant transformation and oncogenesis unraveled in the HIV setting will have broad clinical implications. PMID- 10870596 TI - Perception of quality of life of persons with HIV/AIDS and maintenance of nutritional parameters while on protease inhibitors. AB - This descriptive study investigated quality of life issues, biochemical indices, and nutritional parameters of individuals with HIV/AIDS before the initiation of protease inhibitors (PI) and after PI therapy. Telephone interviews were conducted with 45 men and women who were diagnosed with HIV/AIDS. A 26-item subjective questionnaire was used to determine intake of liquid nutritional supplements, use of micronutrient and herb supplements, adherence to modified diets, gastrointestinal symptoms, employment status, sociability, and ability to conduct activities of daily living. A medical chart review was conducted to collect data on biochemical indices, weight, medication regimens, and incidence of opportunistic infections. The results of the study suggest that HIV/AIDS individuals gain weight, improve CD4 counts, and decrease HIV RNA viral load while on PI-based drug combination therapy. Opportunistic infections decreased, quality of life was improved, and blood albumin was elevated. Hyperlipidemia, that is, elevations in total cholesterol and triglycerides, was observed in study participants (44% and 40% of patients, respectively) after PI therapy. These findings support the need for future investigations to examine the long-term influence of PI-based combination drug therapies on nutrient intake, body composition, and quality of life of persons with HIV/AIDS. PMID- 10870597 TI - HIV/AIDS case histories: diagnostic problems. So-called "bone marrow pattern of AIDS". PMID- 10870598 TI - Soft-gel Norvir. PMID- 10870599 TI - Viagra reacts with ritonavir. PMID- 10870600 TI - Monoclonal antibody has efficacy against HIV. PMID- 10870601 TI - Amprenavir approved. PMID- 10870602 TI - 48-week data released for combination treatment with efavirenz. PMID- 10870604 TI - Vaccine protects against chlamydia. PMID- 10870603 TI - Home hepatitis C test approved. PMID- 10870606 TI - Herpes attitudes studied. PMID- 10870605 TI - Contraceptive sponge returns. PMID- 10870607 TI - Injectable antifungal approved. PMID- 10870608 TI - Substances may help women. PMID- 10870610 TI - Patient education CD-ROM available. PMID- 10870609 TI - Herpes cream developed. PMID- 10870611 TI - Local production of condoms encouraged. PMID- 10870612 TI - HIV and STD prevention update: perception of risk varies dramatically. PMID- 10870613 TI - Actual problems in medical informatics. AB - Medical informatics has been the first science that connects all traditional medical disciplines, thanking to common information needs and requirements of these disciplines. Its goal is incorporating of information technology into medical praxis, getting medical professionals in touch with capabilities of computer technology and preparing the professionals for the future that belongs to more and more powerful computer technologies. Information as a "crucial" component of medicine is in the focus of each investigation. There are many problems that have to be solved through fundamental investigation: knowledge databases; knowledge structuring problems; joining the different kinds of knowledge; knowledge and representation of decision making process; medical concepts; integration and information and knowledge exchange; education of medical staff; structure and organisation. What kinds of management are the best for application in the field of medical informatics? What are the changes in organisation and structure when computer are used? How to connect education processes in clinical centres, libraries, clinical functions and investigations? In this article author discuss about the theoretical and practical part of the education process on biomedical faculties in Bosnia and Herzegovina, according to new curriculum. PMID- 10870614 TI - Contribution of EFMI to development of medical informatics. European Federation for Medical Informatics. AB - The European Federation for Medical Informatics has been established in 1976. At the MIE 96 it has celebrated the 20th anniversary of its existence. During these 20 years the number of number of national societies who became a member has been increased from 10 to 26 and nowadays is 29 not mentioning 2-3 applicants. The objectives of EFMI are: a) to advance international co-operation and dissemination of information, b) to promote research and development, c) to promote high standards in the application, d) to encourage high standards in education in this field. The achieve these goals EFMI organizes yearly European Congresses the MIE-s. Through its working groups contributes very well to the scientific development of medical informatics. The Working Group Chairmen regularly organizes tutorials workshops and many of them participate in teaching medical informatics in their homeland as well as on international courses. EFMI publishes scientific papers from its congresses in the Medical Informatics and in the International Journal of Medical Informatics. The most important meeting, however is the regular Council Meeting--twice a year--where council members can exchange opinions, have opportunity to discuss problems of medical informatics and last but not lease recently medical Inform-ETHICS. This Council is operating as a HUMAN-NET counterbalancing the dysinformations coming from the Internet. PMID- 10870615 TI - [Y2K in medical practice]. AB - The data in computing programs could make two types of problems, functioning of medical equipment and analyses of medical datas. The first one is not use limiting problem. The troubles could be performed in calculation of pregnancy dates but the final calculation in up to medical practitioners. The former situation, analyses of medical datas, could be limited without complete and correct datas, mainly in analyses of life expectancy tables, and related fields. PMID- 10870616 TI - Education of medical informatics in the medical curriculum on biomedical faculties in Sarajevo. AB - In this paper author discussed about experiences of implementing curriculum of education of Medical informatics on biomedical faculties in Sarajevo. Theoretical and practical part of education process, according to new curriculum, during 6 years period of studying at Faculty of medicine in Sarajevo, is hold within the second semester and consists of 30 hours. At Faculty of Dental Medicine in Sarajevo, education is hold in the fourth semester, and consists of 45 hours. At Higher Medical School in Sarajevo, education is also hold in the fourth semester, and consists of 30 hours of theoretical and 30 hours of practical hours as well. Curriculum of Medical Informatics is identical at all these three institutions. Faculty of Dental Medicine education process points out dental informatics, and Higher Medical School devotes more time in education on nursing informatics. PMID- 10870617 TI - [Information technology in medical education]. AB - The role of information technology in educational models of under-graduate and post-graduate medical education is growing in 1980's influenced by PC's break-in in medical practice and creating relevant data basis, and, particularly, in 1990's by integration of information technology on international level, development of international network, Internet, Telemedicin, etc. The development of new educational information technology is evident, proving that information in transfer of medical knowledge, medical informatics and communication systems represent the base of medical practice, medical education and research in medical sciences. In relation to the traditional approaches in concept, contents and techniques of medical education, new models of education in training of health professionals, using new information technology, offer a number of benefits, such as: decentralization and access to relevant data sources, collecting and updating of data, multidisciplinary approach in solving problems and effective decision making, and affirmation of team work within medical and non-medical disciplines. Without regard to the dynamics of change and progressive reform orientation within health sector, the development of modern medical education is inevitable for all systems a in which information technology and available data basis, as a base of effective and scientifically based medical education of health care providers, give guarantees for efficient health care and improvement of health of population. PMID- 10870618 TI - [Telemedicine and telematics in medical education]. AB - Health telematics is a composite term for health-related activities, services and systems carried out over a distance by means of information and communications technologies, for the purposes of global health promotion, disease control and health care, as well as education, management, and research for health. The concept of health telematics encompasses the following functional areas: --tele education;--telemedicine;--telematics for health research;--telematics for health services management. Communications technologies are rapidly revolutionizing health care. For example, electronic communications support diagnosis and treatment of disease. TeleMedicine is an umbrella term for growing disciplines such as TeleRadiology. TelePathology. TeleCardiology, TelePsychiatry and TeleEducation. TeleMedicine is a component of TeleHealth, which includes the use of telecommunications technology and services for the surveillance and control of diseases and education. In this article authors describes the role of telemedicine and telematics in medical education and medical praxis. PMID- 10870619 TI - [Expert systems in medicine]. AB - Expert systems are software systems developed using different techniques of artificial intelligence that can act parallel to the "human" experts. The main role is consultative These are intelligent information systems that use more then 2000 different rules and that are capable to explain their decisions. Databases of such systems can contain huge number of data about different diseases and therapy modalities. In development of Medical Expert systems the rule of human experts is crucial. The teams of such experts are developing expert system considering the changes in medicine. Several modes of work are available. Consultation mode is used in cases when the diagnosis and treatment is uncertain. The human enter data about symptoms and signs of some medical disorder and computer creates a list of possible diagnosis and additional diagnostic test. Therapy for condition is also suggested. Simulation mode can simulate virtual patient and allows students and doctors to learn mode about some medical conditions. Some expert system as HEPAT can make "Decision Tree" for new-born jaundice. Similar expert system will be available in future for other fields in medicine. Some of expert systems are described in article. PMID- 10870620 TI - [Use of actual medical information in daily practice]. AB - Use of medical information in everyday practice has been described in this paper. Importance of systematic collection, analysis and use, including correct "data management" is noticed. Information system is formed of every noted information, not only in computing form. Use of databases allows us connection to many information, but rational use should allow us to select only these in number an quality which are necessary for decision making. PMID- 10870621 TI - [Experience in the development of information systems at the canton level]. AB - In the aim of need and necessity of stronger orientation and more efficient aim realization 35 Strategy SZO (health information system) country members should have the health information system able to provide information support to their National strategy "Health for All until 2005". Such a health information system should provide the information support for the planning process, monitoring and health evaluation, health care and work in public-health services sector. The experiences in the work of information system organization in Tuzla Canton Public Health Institute should be seen from two aspects: Aspect of the information system significance in the framework of the social-medical, epidemiological, hygiene-ecological and other activities of the Institute; Aspect of the information system development within the Public Health facilities of Canton in relation to the automatic data processing important for registration, tracking and recognition of success and population health care needs. From the aspect of establishing the information system in the area of Tuzla Canton and the Public Health Institute role, there is a need for adequate high technology technical equipment, with the task to enable practical realization of defined projects and in such a way, varied acceleration of work giving as a result usable information. The canton Public Health Institute needs as follows: necessary resources for free information system work; to provide free computer access to all; to ensure connection with other PHI through communications. In this way the canton Public Health Institute information system would be connected with other information systems in the field of health. In would be a base for development of integrated health information system. The information system is desirable aim for the work of Public Health Institute, method of selection and leading content of modernization. Analysing the Bill of Law on Health Care, a defectiveness has been noticed, since the information system is not regulated. Therefore, the need to be issued by Lay is of great importance. A statement that all projects on local information systems, started before war or during the war have been stooped, joins to the aforesaid, and also a series of independent information system in different health segments, which are not connected into one, unique information system. In any case, this disparaged the quality of medical decision making and therefore, providing of high-quality services at all levels of health work. PMID- 10870622 TI - [Rapid development of a uniform computerized health information system in the Zenica-Doboj Canton]. AB - It is very expensive and long-termed process to establish a complex information system, and it requires to include large number of people and processing of large number of information. As I believe that health economics and management are the key factors for maintaining health system, I think that it is now essential to create information system which will, above all, ensure accurate and prompt information to the health managers. Therefore, I suggest building of a cheaper variant of the information system which could be upgrated, and which will involve, on this stage, small number of health workers. Computerized part of this information system would be built in several levels. At each of these levels data for the needs of health statistics, economic-financial activities, and management would be processed. Communication within information system, on this stage, would be enabled through telephone lines and modems. The system would work "off line". The change of data would be carried out periodically. PMID- 10870623 TI - [Local information systems at the Pediatric Clinic at the University Clinical Center in Sarajevo--experience and perspectives]. AB - Computer were first introduced at Pediatric hospital in Sarajevo in 1989 and since 1990 first programs for managing data have been started. They were used for administration of patients and history taking, as well as for collecting clinical data of them. In the beginning, introduction was slow because lot of doctors and nurses were reluctant in using new techniques. But, in a year most of them realized all the advantages PC offers. At that time all the PCs were separated, that has limited their full facilitated data gathering especially in the periods when we lacked all other office materials (paper, typing machines ... even pencils). Thanks to them we have preserved all medical data about patients in 4 years war period. After the end of the War we started project of making clinical network and program that should run most of the work that is performed at Pediatric Hospital in Sarajevo. Everything that is done at hospital and could be helped by the use of the computers was recorded and algorithms were made. The network consists of 15 PC units. Program was developed through several phases from the admittance of the patients and administration regarding it to the discharge letter. Outpatient work was incorporated, as well as gathering all the medical findings of the patients at one place. First experiences are extremely positive. We have speeded up "paper work" and freed much time that medical stuff can spend with patients. The main problems that we encounter are need for permanent education in working with system, lack of more powerful server that can handle more data and introducing of the pictures in the medical records. We conclude that clinical network with the use of good program for managing all the data gathered in the hospital is essential for today's work. PMID- 10870624 TI - [Problems in health statistics reporting in Bosnia-Herzegovina]. AB - Health-statistic information system in Bosnia and Herzegovina is presented, its content, problems and reform objectives in accordance with new European trends. Adequate and prompt health information plays a key role in the policy of planning and decision-making at all health levels. Therefore, data collecting must be conducted in accordance with the country's needs aimed to the health promotion and health of the population improvement. Scope and structure of health information system varies from country to country depending on health status of population, health policy and, of course, economic capacity. Public Health Institute of Federation BH is authorized by the Law to conduct statistical research in the health sector, and, therefore, the Proposal of statistical research in health sector is made, according to these regulations. This Proposal offered 17 statistical researches for 1998. Unfortunately, only 7 researches have been carried out. The shortcomings of the existing health-statistic information system have been found out on the local, cantonal, entity and state level. PMID- 10870625 TI - [Evidence law in health care, development of a health care information system and use of automatic processing of information]. AB - The aim of work is to analyse the state of health-statistic and information system within the Federation of Bosnia and Herzegovina, analyse the Program on health statistic research important for the entire country, analyse the Law on records in the field of health and present how deficient it is, as well as to propose the set of other data, based on our own research, that could fill noticed defects in the practice, provide the use of computer technology in data tracking from the Law on issued records in the field of health within the activity as an example for others--health statistic sheet, form 3-21-61/A, and to present further possibilities and recommendations for development of health information system. The used method of work is descriptive analysis of methodological approach to the health statistic information system of Bosnia and Herzegovina (taken from the former Yugoslavia). We have also applied the data from the statistic records (retrospective analysis) of the registered cases from the neurological ward of the Neurological Clinic Tuzla for December 1997. In such a way we formed the data base and presented the statistic data processing ideological project including modern computer technologies. PMID- 10870626 TI - [Health status indicators and the importance of their use in daily practice]. AB - Health information undoubtedly has great social, political and economic importance. To have relevant information, it means to have basis for timely reporting, basis for action, and prerequisites for success. The purpose is to produce and use adequately appropriate information about health status, health risks and health outcomes in order to establish priorities. Health information should be useful and easy accessible to all potential users in every moment, from health care providers to the general public. Health indicators, as aggregated health statistical data, present measure instruments in this process, and because of that definition, development and implementation of basic set of health indicators at all levels (public health and clinic) must be the priority activity in the current moment of health sector reform in Bosnia and Herzegovina. Basic set of relevant and measurable indicators should be founded on different aspects of health and health care (health promotion, diseases prevention, clinical treatment, rehabilitation), monitoring and evaluation as a part of everyday's routine in health sector. PMID- 10870627 TI - [New electronic data carriers in Bosnia-Herzegovina]. AB - Bosnia and Herzegovina has been developing new Health Care System based on Electronic Registration Card. Developing countries proceeded from the manual and semiautomatic method of medical data processing to the new method of entering, storage, transfer, searching and protection of data using electronic equipment. Currently, many European countries have developed a Medical Card Based Electronic Information System. Both technologies offer the advantages and disadvantages. Three types of electronic card are currently in use: Hybrid Card, Smart Card and Laser Card. Hybrid Card offers characteristics of both Smart Card and Laser Card. The differences among these cards, such as a capacity, total price, price per byte, security system are discussed here. The dilemma is, which card should be used as a data carrier. The Electronic Family Registration Card is a question of strategic interest for B&H, but also a big investment. We should avoid the errors of other countries that have been developing card-based system. In this article we present all mentioned cards and compare advantages and disadvantages of different technologies. PMID- 10870628 TI - [Anatomic-therapeutic-chemical classification of drugs]. AB - European Research Association for Pharmaceutical Market and an international group have developed the ADC Drug Classification system, which is recommended by the World Health Organization (WHO). It has been in use since 1987. According to this classification, drugs are grouped into fourteen basic groups according to the organic system of the organism where they work. These fourteen anatomical groups represent the first anatomical level and is labeled with one capital letter, as follows: A Alimentary tract and metabolism B Blood and blood forming organs C Cardiovascular system D Dermatologicals G Genito urinary system and sex hormones H Systemic hormonal preparations, excl. sex hormones J General antiinfectives for systemic use L Antineoplastic and immunomodulating agents M Muscle-skeletal system N Nervous system P Antiparasitic products, insecticides and repellents R Respiratory system S Sensory organs V Various. Drugs are further divided into therapeutic groups and subgroups (2nd and 3rd level), the 4th level is the chemical-therapeutic subgroup. The 5th level is the generic drug name. Each drug is represented by 7 numeric-character bytes code. These seven bytes are determining the group (1st anatomical classification level marked by a capital letter) with the corresponding therapeutic subgroups on the 2nd and 3rd classification level, 4th level labels the chemical-therapeutic subgroup, while the 5th level is signified by the individual chemical compounds (generic name) and is marked by an Arabic number. The importance of the ATC classification is the possibility of the international comparability, monitoring of the use and consumption from various aspects. The standardized monitoring methodology incorporates too the daily doze determining methodology (DDD). Defined Daily Dose (DDD) is the drug amount used for the most common indication, and is, therefore, the basic statistical unit of drug use monitoring. It represents not only the recommended doze, but is also the only means of acquiring the number of patients receiving that particular drug (DDD per 1000 inhabitants). This makes the basis for the comparability of drug use in various places (country, region, institution, etc.). PMID- 10870629 TI - [Quality of medical records as a basis for DRG classification in the health care system in Bosnia-Herzegovina]. AB - Medical Record contains data about use of health care services of every member of certain population, no matter if they come from preventive or curative health care. Quality of Medical Records is very often in direct as well as indirect connection with health care quality. Better health care systems usually have better Medical Records. These are computer processed and stored on some of contemporary computer media, like: disks, microfilms, magnetic tapes and so called "smart cards". Good Medical Record should have at least next characteristics: 1. Time dimension, which should contain chronological: past, present and future data, relevant to health care consumers. 2. It should be complete in a sense that every Medical Record must contain adequate medical and other relevant data for health care planning, organization and control. From the aspects of our research described here, it is very important that Medical Records contain all financial data. These usually are consequences of consumption of health care services from various resources of different levels. The results of our research about the application of DRG classification in B&H health system, that have been done in Tuzla University-Clinical Center, show that hospital Medical Records have many inadequacies, both in respect of medical as well as non medical part. Physicians use very little ICD coding system. There is no uniform anamnesis, no detailed evidences about all hospital services, no adequately defined severity of disease and patient status at discharge. Due to manual based information system, there is no evidence about spending of resources per patient with financial data. The development and application of computer based health information system must go towards direction of solving these problems. PMID- 10870630 TI - [Dilemmas in the classification of foot injuries caused by land mines]. AB - In international classification of injuries, very often there are some specific conditions that are not clearly defined. War initiated that about specific conditions before all injuries a discussion in term of more correct presentation through the code is being made. Very often, some conditions in international classification are being led as INSUFFICIENTLY DEFINED or UNMARKED PART. When we have present large destruction due to wounding with projectiles of high kinetic energy (initial velocity), and especially stepping on land-mines, we have the situations that can not be correctly defined. The amount of these injuries which are about to follow in the future period authors will declare in few examples which have been surgically treated in Traumatological Clinic in Sarajevo. All injuries were result of stepping on land-mines and in all cases it was about foot. The stated injuries we were not able to code as amputations because the distal part of the injury was vascularized, later on we found out that there are possibilities of surgical reconstruction in meaning of functional foot, since we do not have open (explosive) fracture here. May-be, as a clinical formulation or for initial observation a more appropriate name is (term analogue in surgery of other regions beside limbs) INTERCALARY amputation of limbs. The same one could be moved from deficiency to a complete lack which surely must be linked to its causality. PMID- 10870631 TI - [The importance of Internet in medical education]. AB - Internet is more and more involved in medical education in many countries including Bosnia and Herzegovina. Not only medical student but also physicians are using Internet to find out the latest information in specific field of medicine. Some sites are specially designed to be used for medical education. Information about some programs or courses of medical education can be found here. Improvements of network resources and multimedia technologies have made it possible to satisfy needs for medical Education. Multimedia approach offer possibility to show text, picture, sound or movie considering specific need. All of that is available on Internet. Many search engine are available in the world and student can use all of them when they have access to Internet. The more precise search can be done on specific sites that include information about medical conditions and medical education. The most important is MEDLINE. MEDLINE is bibliographic database of National Library Of Medicine in USA. This database can be explored from several sites. All relevant information about article can be find here including abstract and service to obtain full text of specific article. Database can be searched using specific keywords that can be find in text or in MESH thesaurus. Data about authors, their addresses and title of article can be found, too. The possibility of using Internet in medical education are considered in this article. Some of Internet sites are described, too. PMID- 10870632 TI - [Telemedicine with respect to telepathology in Bosnia-Herzegovina]. AB - Telemedicine is rather new activity covering various approaches in order to fulfill the main task which can be expressed in more comprehensive words such as "remote medicine" or "medicine on distance". One of definitions of the telemedicine given by the European Commission for Telemedicine says that "telemedicine is fast access to distributed medical expert knowledge using telemedicine and information technologies regardless of actual location of a patient or relevant information". The first connection has been established with Zagreb, Croatia in September by means of PHAROS. In October 1996 the Institute of Pathology with the Radiology and Ophthalmology Clinic of University hospital of Sarajevo has joined the experimental project "SHARED" (a telemedicine initiative to support remote health care structure, proposed by the San Raffaele International Biomedical Science Park with Stato Maggiore della diffesa and European Space Agency). Due to problems in infrastructure and lack of experts, small country like, Bosnia and Herzegovina (B&H) sees telemedicine as a future in its health care organizing. PMID- 10870633 TI - [Telemedicine in vitreoretinal surgery]. AB - The authors present one-year experience of the multimedia role in ophthalmology with particularly review on example in vitreoretinal surgery. It has been shown goal of the project, methodology, evaluation of one-year work results and possibility of project's expansion. PMID- 10870634 TI - [Clinical algorithms in the treatment of status epilepticus in children]. AB - The clinical algorithm is a text format that is specially suited for presenting a sequence of clinical decisions, for teaching clinical decision making, and for guiding patient care. Clinical algorithms are compared as to their clinical usefulness with decision analysis. We have tried to make clinical algorithm for managing status epilepticus in children that can be applicable to our conditions. Most of the algorithms that are made on this subject include drugs and procedures that are not available at our hospital. We identified performance requirement, defined the set of problems to be solved as well as who would solve them, developed drafts in several versions and put them in the discussion with experts in this field. Algorithm was tested and revised and graphical acceptability was achieved. In the algorithm we tried to clearly define how the clinician should make the decision and to be provided with appropriate feedback. In one year period of experience in working we found this algorithm very useful in managing status epilepticus in children, as well as in teaching young doctors the specifities of algorithms and this specific issue. Their feedback is that they find that it provides the framework for facilitating thinking about clinical problems. Sometimes we hear objection that algorithms may not apply to a specific patient. This objection is based on misunderstanding how algorithms are used and should be corrected by a proper explanation of their use. We conclude that methods should be sought for writing clinical algorithms that represent expert consensus. A clinical algorithm can then be written for many areas of medical decision making that can be standardized. Medical practice would then be presented to students more effectively, accurately and understood better. PMID- 10870635 TI - [An addition to the clinical algorithm for treatment of patients with community acquired pneumonia in Bosnia-Herzegovina: a multicenter prospective study in Sarajevo]. AB - The Patient Pneumonia Outcomes Research Team (PORT) has developed a scoring system that allows the severity of illness to be quantitatively measured. Using an algorithm, the patient can be classified as to risk. The total number of points assigned to the patient increases risk of poor outcome. Patients in risk classes I, II and III have an expected mortality (< 5%) and should be managed as outpatients. Patients with risk class IV and V have mortality rates of 8-29% and should be admitted to the hospital. Patients with risk class V will usually require intensive care. AIM: We started a multicenter prospective study in Sarajevo, to investigate some differences in demographic factors, comorbid diseases, physical examination findings, laboratory findings, the duration, complications and mortality in our population with CAP, than in recommended algorithm. MATERIAL AND METHODS: We recorded the cases of 163 patients (100 male, 63 female) with mean age 58.6 years (17-91). The sample was statistical elaborated with tests of distribution frequency and hi square test. RESULTS: Between demographic factors, as significant, beside age, there was nicotine abuse in 53% (for our CAP population). In comorbid diseases as significant disease beside preexisting cancer (10.5%), chronic liver disease (4%), renal disease (13%), congestive hearth failure (27%), cerebrovascular disease (5%), there was COPD (31%) and diabetes mellitus (9%). Some differences were in physical examination findings, and laboratory findings. The duration of illness, complications and mortality in our CAP population was like in other studies. CONCLUSIONS: This work speaks us that our CAP population has some differences in observed characteristics of scoring system. But, without regard to all this, patients with following characteristics should also be admitted to an intensive care unit and would be classified as patients with severe pneumonia: hypotensia (systolic blood pressure < 90 mm Hg), impending respiratory failure that may require mechanical ventilation, hypoxaemia (pO2 < 60 mm Hg), hemodynamic instability, heart failure, diabetes mellitus (tip I), COPD, poor dental hygiene (anaerobes--necrotizing pneumonia, empyema, abscess). PMID- 10870636 TI - [Algorithms in ultrasonic diagnosis of diseases of the hepatobiliary tract]. AB - The hallmark in clinical diagnostic in last two decade is ultrasound. Today, the diagnosis of numerous diseases detected was using US diagnostic. US the first diagnostic procedure in medical practice of all specialties, especially in abdominal disease. Liver and spleen diffuse or focal lesions, and other abdominal disease very easy will be detected using by US. There are some diagnostic algorithms that solved diagnostic dilemmas, for instance in the case of jaundice, focal and diffuse liver and spleen lesions, and other gallbladder, biliary ducts, pancreas disease and abdominal solid or liquid collections. The algorithms in diagnostic of abdominal disease are the choice for successful diagnostic procedure, very often used in medical practice. PMID- 10870637 TI - [Criteria for evaluation in family medicine]. AB - For successful evaluation and the follow up of the family medicine team work it is necessary to develop evaluation criteria. Monitoring and evaluation criteria are numerous and different among countries depending on the health care development and the economic power of the country. In the present circumstances, taking in consideration health care reform through Primary Health Care (PHC) through family medicine team, the following criteria are proposed: equity, availability, cost-effectiveness, efficiency, efficacy, quality and satisfaction of user and health services providers. Indicators for the above mentioned criteria are also defined to estimate criteria achieved development. First indicator group relates on the medical-social indicator (for example number of families, structure of families and structures of concerning population) and the others social-economic indicators (GNP, expenditure in the health and social sector per capita, etc.). Second indicator group follows up the content of family medicine team work as follows: indicator of active and passive health care (for example number of doctor services, number of doctor and nurses home visits, consultation services, diagnostic procedures etc.). Third indicator group relates on the economic aspect, meaning the cost of implementation of such program, according indicator on content of work. Suggested criteria with indicators present the basic minimum databases, which would give the possibilities to have continuous overview in family medicine team functioning as well in its quality of work. PMID- 10870638 TI - [Computerized surveillance of communicable diseases as a part of public health surveillance]. AB - Conducting surveillance of communicable diseases and/or public health surveillance in general, in developing countries such is B&H has some common specific things-health care system is an integral part of organized government services thus, fewer impediments to implementing any part of a surveillance system are recorded. Limited health care providers and laboratories reduce the number of data source and can facilitate quality assurance. An ideal surveillance system is discussed in this paper (what kind of surveillance system an epidemiologist would like to have in future), as well as barriers to the ideal surveillance system and technology of the future. Some elements of computerised public health surveillance system and surveillance of communicable diseases, particularly, are given (e.g. hardware, software, data entry, editing the data, analysis of data, ...) Computerising a surveillance system will results in increasing speed of processing providing graphic capability, enhancing analytic capabilities, improving quality of data of reports and improving quantity of data. The most important step in beginning to conduct computerising a surveillance system is identifying the public health objective such is, for example, surveillance of communicable diseases. PMID- 10870639 TI - Short exposure to millimolar concentrations of ethanol induces apoptotic cell death in multicellular HepG2 spheroids. AB - PURPOSE: We have shown previously that 1 mM ethanol reduces cell proliferation and increases apoptosis in monolayers of human hepatocellular carcinoma (HepG2) cells. However, in vivo liver tumors are usually three-dimensional and multicellular. The purpose of this study was therefore to determine the effect of ethanol in multicellular tumor spheroids (MCTS) as a model system in vitro. METHODS: After the application of 1 mM ethanol for 24 h and 48 h, viable, apoptotic and necrotic cells within MCTS were stained with specific fluorescent dyes, and their amount and distribution within the MCTS were assessed by confocal laser scanning microscopy. To evaluate the effect on HepG2 cell migration and cell proliferation, the outgrowth potential after 1 week in culture was evaluated. RESULTS: As assessed by YO-PRO-1 staining, ethanol increased the number of apoptotic cells from 21.5 units (U) in control spheroids to 364 U and 482.2 U after 24 h and 48 h in ethanol-treated spheroids, respectively (P < 0.001). Merocyanine staining fluorescence increased from 10.7 U in the control to 122 U after 24 h and 293.2 U after 48 h (P < 0.001). Cell viability, as determined by staining with the acetoxymethyl ester of calcein, decreased from 578.5 U in the control to 236 U and 73.4 U after 24 h and 48 h of ethanol exposure respectively (P < 0.001). Necrosis showed an increase from 2 U in control to 24.9 after 24 h and 54 U after 48 h. MCTS treated with ethanol showed almost complete inhibition of outgrowth potential after 1 week in culture, compared to controls (P < 0.005). CONCLUSIONS: Small concentrations of ethanol (1 mM) induced apoptosis in HepG2 MCTS with a concomitant inhibition on outgrowth potential, accompanied with a low degree of necrosis. These findings suggest that low concentrations of ethanol may already be sufficient for the treatment of hepatocellular carcinoma. PMID- 10870640 TI - Cytotoxicity, cell-cycle perturbations and apoptosis in human tumor cells by lipophilic N4-alkyl-1-beta-D-arabinofuranosylcytosine derivatives and the new heteronucleoside phosphate dimer arabinocytidylyl-(5'-->5')-N4-octadecyl-1-beta-D arabinofuranosylcytosi ne. AB - The arabinofuranosylcytosine (AraC) derivative N4-octadecyl-1-beta-D arabinofuranosylcytosine (NOAC) and its (5'-->5')-heterodinucleoside phosphate analog NOAC-AraC were compared with AraC for cytotoxicity, cell-cycle dependence, phosphorylation by deoxycytidine (dC) kinase and apoptosis induction in native, AraC- or NOAC-resistant HL-60 cells. NOAC was cytotoxic in all cells with three to seven-fold lower IC50 concentrations than those of NOAC-AraC or AraC. In contrast to NOAC-AraC, the lipophilic monomer NOAC overcame AraC resistance, inducing apoptosis in more than 80% of native and AraC-resistant HL-60 cells. This suggests that NOAC-AraC may be cleaved intracellularly only at very slow rates to AraC and NOAC or to the 5'-monophosphates, whereas NOAC exerts different mechanisms of action from AraC. In vitro the dimer was cleaved by phosphodiesterase or human serum to NOAC, AraC and AraC monophosphate. In contrast to AraC, N4-alkylated AraC derivatives with alkyl chains ranging from 6 18 C atoms were not substrates for dC kinase. Furthermore, treatment of the multidrug-resistant cell lines KB-ChR-8-5 and KB-V1 with the N4-hexadecyl-AraC derivative NHAC did not induce P-170 glycoprotein expression, suggesting that the N4-alkyl-AraC derivatives are able to circumvent MDR1 multidrug resistance. The in vivo activity of liposomal NOAC in a human acute lymphatic leukemia xenograft model confirmed the antitumor activity of this representative of the N4-alkyl arabinofuranosylcytosines. PMID- 10870641 TI - Adhesion proteins, cellular morphology and fibrous components around the cell/extracellular-matrix interface in myxoid liposarcomas. AB - We examined the cell/extracellular-matrix interface in 13 myxoid liposarcomas by determining the distribution of collagen and reticular fibers in the myxoid matrix, the presence of adhesion proteins and the morphological features of the cytoplasmic border. Adhesion proteins (fibronectin, integrin alpha3) and the cytoplasmic border were examined by immunofluorescence and a differential interference-contrast image analysis respectively. A network of reticular fibers and collagen fibers was present in the myxoid matrix of 11 cases (85%) and 3 cases (23%) respectively. Tumor cells with dendritic cytoplasmic processes were observed in 8 cases (62%). Adhesion proteins were sparsely present in tumor cells and there was no correlation between those proteins and collagen fibers, reticular fibers or cytoplasmic processes. Collagen fibers, dense reticular fibers or well-developed cytoplasmic processes were more frequently observed in the cases of long-term-surviving patients than those with recently developed tumors or patients who died. All 3 cases positive for collagen fibers also contained both dense reticular fibers and cytoplasmic processes. Our findings suggest that the fibrous components in the myxoid matrix and the well-developed cytoplasmic processes may limit the invasiveness of malignant cells. This peculiar architecture may also explain the slowly progressive nature of myxoid liposarcomas. PMID- 10870642 TI - Value of 31P NMR spectroscopy in predicting the response of a xenografted human hypopharynx carcinoma to irradiation. AB - PURPOSE: An early indicator of tumor sensitivity to irradiation could provide useful information on the effectiveness of therapy and may facilitate more individual designs of treatment protocols. The aim of the present study was to evaluate the potential of in vivo 31P nuclear magnetic resonance spectroscopy in predicting the response of a xenografted human hypopharynx carcinoma to radiotherapy. METHODS: The tumor had been serially heterotransplanted to athymic mice. 31P NMR spectra were collected before and at four intervals (24, 48, 72, and 120 h) after irradiation with 15 Gy or 30 Gy. Alterations of phosphorus metabolism were compared with the growth delays, the histological appearance, and the mitotic activity of the treated tumors. RESULTS: Radiation with 30 Gy induced increases of the phosphodiester level (P < 0.001) as well as of the tumor pH (P < 0.05) and decreases of the phosphomonoester level (P < 0.001) within 48 h. The changes clearly preceded measurable tumor responses and were accompanied by severe histological destruction and marked depression of mitotic indices. However, none of these spectral alterations was significantly correlated with individual delays of tumor growth. The only parameters allowing a prediction of radiation-induced tumor responses were the pre-treatment levels of phosphomonoesters and -diesters. The 31P NMR spectroscopic changes observed after therapy with 15 Gy were either unsystematic or insignificant. CONCLUSIONS: Pretreatment levels of tumor phospholipids were indicative of radiosensitivity in the xenografted human hypopharynx carcinoma investigated here. However, since phosphorus metabolism varies considerably among different tumor lines, it seems unlikely that there exists a uniform 31P NMR spectroscopic parameter predicting tumor response to radiation therapy. PMID- 10870644 TI - Salvage of the upper extremity in cases of tumorous destruction of the proximal humerus. AB - Malignant bone tumours or metastasis of the upper humerus may cause significant loss of function especially in those patients with resectional arthroplasty of the shoulder. One method for achieving functional reconstruction of the humerus concerned is replacement with a modular endoprosthesis. Little is known about clinical and radiological results in these rare circumstances. Between 1993 and 1997 we treated 21 patients (22 shoulders) with enlarged osteolytic destructions of the proximal humerus caused by metastatic spread or primary malignant tumours. Patients with additional involvement of the glenoid were excluded from this study. The average follow-up was 3.9 years. Every 3 months all patients were followed-up clinically and radiographically. Prior to surgery, diagnosis was established by incisional biopsy and the outcome determined the therapeutic algorithm (radiotherapy, chemotherapy, surgery). In most cases of metastatic lesions, surgery was the first treatment. According to the regional spread of the tumour, various amount of bone and soft tissues had to be removed. The distal stem of the prosthesis was inserted in a cementless way and secured to bone with two interlocking screws. The length of the diaphyseal part depended on the site of osteotomy. Soft-tissue coverage of the large implant was achieved in all patients. Early complications were lymphogenic oedema and superficial wound dehiscence. One patient developed a deep infection, which had to be managed surgically. According to the functional rating system of the Musculoskeletal Tumour Society for the upper extremity the overall results were inversely proportional to the extent of resection. None of our patients achieved unrestricted motion of the shoulder concerned. The most important finding was a proximal migration of the prosthesis causing a painful subacromial impingement, mainly a consequence of the resection of the deltoid muscle and the rotator cuff. In summary, a modular endoprosthesis cannot be recommended generally as the method of choice. If the muscular balance of the shoulder is too weak to act as a joint centralizer the endoprosthesis has no advantage over a simple diaphyseal spacer. PMID- 10870643 TI - Serum CYFRA 21-1 in cervical cancer patients treated with radiation therapy. AB - BACKGROUND: A fragment of cytokeratin 19, referred to as CYFRA 21-1, is abundant in the serum of many patients with malignant tumors and is recognized as one of the established tumor markers, especially for non-small-cell lung cancer. In this study, the clinical usefulness of CYFRA 21-1 was investigated in cervical cancer patients treated with radiation therapy with reference to squamous-cell-carcinoma related antigen (SCC-Ag), a common tumor marker of cervical squamous cell carcinoma. MATERIALS AND METHODS: The serum levels of CYFRA 21-1 and SCC-Ag of 50 patients with squamous cell carcinoma of the uterine cervix were measured before and after radiation therapy. RESULTS: CYFRA 21-1 was positive in 52% of the patients. The incidence increased with the stage of the cancer, and post treatment increases were a sign of disease progression. During radiation, serum levels of CYFRA 21-1 decreased significantly and reflected the radiation effect well. In addition, CYFRA 21-1 was negative in all patients without distant metastasis at the end of radiation therapy. Compared with SCC-Ag, patients were less often positive for CYFRA 21-1, but there was a statistically positive correlation between the two markers (correlation matrix=0.69). CONCLUSIONS: CYFRA 21-1 can be used in monitoring the outcome of patients with squamous cell carcinoma of the uterine cervix. It may be particularly useful for patients without SCC-Ag. PMID- 10870645 TI - Serum concentration of E-selectin in patients with chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. AB - OBJECTIVES: High levels of soluble E-selectin have been reported in acute and chronic inflammatory disorders. Moreover, in some types of tumor elevated values have been found while in other types reduced levels have been reported. Our aims were to determine whether soluble E-selectin levels might be useful in monitoring the progression of chronic liver disease, including hepatocellular carcinoma. METHODS: Circulating soluble E-selectin was measured by an enzyme-linked immunosorbent assay in the sera of 18 patients with chronic hepatitis, 44 with liver cirrhosis, and 38 with hepatocellular-carcinoma-associated liver cirrhosis. Immunohistochemical localization of E-selectin was also performed on liver tissue specimens of patients with chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. RESULTS: Serum levels of soluble E-selectin were higher in the chronic hepatitis and liver cirrhosis patients than in the hepatocellular carcinoma patients and healthy controls. Levels in the hepatocellular carcinoma patients and controls were not significantly different. In the liver cirrhosis group, divided according to the Child-Pugh classification, soluble E-selectin decreased with disease severity. Similarly, in patients with liver cirrhosis who developed hepatocellular carcinoma, soluble E-selectin decreased as the disease progressed. Immunohistochemical localization showed strong membrane staining on endothelial cells in areas rich in inflammatory cells in severe chronic hepatitis. In some hepatocellular carcinoma tissues a marked E-selectin staining was observed on endothelial cells of tumor-associated small vessels. CONCLUSIONS: The results obtained suggest that high serum levels of soluble E-selectin are associated with chronic hepatitis and liver cirrhosis, and that levels decrease in liver cirrhosis patients as the disease progresses. Patients with hepatocellular carcinoma have different types of soluble E-selectin behaviour the significance of which requires further investigation. PMID- 10870646 TI - Further evidence for oxidant-induced vascular endothelial growth factor up regulation in the bronchoalveolar lavage fluid of lung cancer patients undergoing radio-chemotherapy. AB - BACKGROUND: Vascular endothelial growth factor (VEGF) is a potent inducer of physiological and neoplastic blood vessel growth. Moreover, in vitro studies have demonstrated that VEGF can be up-regulated by conditions associated with the generation of free radicals and reactive oxygen species. In a previous study we reported on strongly increased VEGF concentrations in the bronchoalveolar lavage fluid (BALF) of patients with lung cancer under therapy. In this study we aimed to reveal whether this increase was due to the therapy-associated intrapulmonary oxidative burden. PATIENTS AND METHODS: A total of 103 BALF samples from 94 patients with lung cancer (82 patients with non-small-cell lung cancer, 12 patients with small-cell lung cancer) were studied at different times before, during or after cancer treatment. VEGF levels in the lavage fluid and ratios of oxidised methionine in proteins of epithelial lining fluid (ELF) were determined. RESULTS: As reported previously, strongly increased VEGF levels in the ELF were observed in patients undergoing chemotherapy when radiotherapy had been administered before. Increased levels of oxidised methionine indicated that these patients suffered from severe pulmonary oxidative stress that was significantly less in patients undergoing only chemotherapy. Similarly, VEGF concentrations in the ELF were significantly elevated in cancer patients at the time of diagnosis, but the oxidised methionine levels did not reveal significant oxidant/antioxidant imbalances in these patients. CONCLUSION: Systemic chemotherapy is associated with oxidative stress in vivo, which is more pronounced if patients are additionally treated with radiation. VEGF levels in the ELF are increased by this condition as well as by the activity of the tumour itself. PMID- 10870647 TI - Pediatric tumors and secondary cancer. The Ninth International Symposium of the Hiroshima Cancer Seminar, September 1999, Hiroshima, Japan. PMID- 10870648 TI - Osteoporotic fractures, ageing, and the bone density T-score. PMID- 10870649 TI - The fallacy of BMD: a critical review of the diagnostic use of dual X-ray absorptiometry. AB - The diagnostic use of BMD should be cautious as BMD is not an ideal measure of true bone density; it is not an ideal measure of bone strength; it does not predict fractures well; and it has inherent problems of accuracy and linearity. The limitations of BMD, based on the physical deficiencies of DXA, are further obscured by the introduction of T-scores. It is suggested that BMD and BMC, when used diagnostically and for fracture risk classification, be used after correction for body size and/or bone size, age and sex, and that measured values be evaluated in the light of established mean fracture incidence data. BMD is not a parameter of sufficient validity to be the sole indicator of present and future fracture risk. A low BMD should be regarded one of several fracture risk factors. It seems that there is a need to redefine the T-score based definition of osteoporosis. PMID- 10870650 TI - Gynaecologic history in systemic sclerosis. AB - The aim of the study was to analyse the gynaecologic history of 150 Brazilian patients with systemic sclerosis (SSc) by comparing the outcome of the pregnancies before and after disease onset and in the two clinical variants of SSc, as well as to assess the effects of the pregnancy on the progress of the disease. A retrospective analysis was carried out of 150 female SSc patients, more than 18 years old, who attended the outpatient clinic of the Unit of Rheumatology of the State University of Campinas. The patients were questioned about the number of pregnancies, deliveries (full-term infants, premature births and twins) and fetal deaths (spontaneous abortions and perinatal deaths). These data were subdivided into pregnancies before and after SSc onset. In those gestations started after disease onset the patients were questioned about the evolution of SSc during the pregnancy. The patients were also asked about dyspareunia and the age at menopause. Thirty-two patients (21 %) had never been pregnant, and only five of them were considered infertile. One hundred and eighteen patients (79%) had a total of 406 pregnancies, with an average of 3.4 per patient; there were 364 pregnancies before and 42 after SSc onset. There were 58 fetal deaths (14% of the pregnancies), 50 of these occurring before and eight after disease onset; 55 were spontaneous abortions and the other three were perinatal deaths. The fertility rate was higher in the limited SSc (3.6) than in the diffuse SSc patients (3.1), although the percentage of fetal deaths and the evolution of SSc during the pregnancy were similar in the two clinical variants. In the pregnancies that occurred after the onset of SSc, the clinical course remained stable in 72% of the cases, worsened in 14% and improved in 14%. Dyspareunia was mentioned by 49 patients (37% of those with an active sexual life). Menopause was reported by 72 patients, predominantly with limited SSc (61 patients). The fertility rate in the postmenopausal SSc patients was 3.9, similar to that observed in general postmenopausal population in Brazil. The analysis of the gynaecologic history in this series of SSc patients showed no increased risk in infertility or spontaneous abortions. The fertility rate in the two SSc clinical variants was higher than that observed in the local global population. Most of the patients who became pregnant after the onset of SSc showed no signs of worsening during the course of the disease. PMID- 10870651 TI - Study of peripheral bone mineral density in patients with diffuse idiopathic skeletal hyperostosis. AB - Diffuse idiopathic skeletal hyperostosis (DISH) is an ossifying systemic enthesopathy which involves not only the spine but which may also appear in other sites. Degenerative, inflammatory and metabolic factors have been reported for a possible pathogenic role in the new bone growth that characterises DISH. In the present study peripheral bone mineral density (BMD) has been measured in patients affected by DISH and the results compared to those of a control group. Forty-two patients (33 females and 9 males) affected by DISH and 84 controls (66 females and 18 males) were examined. All subjects underwent radiological study of the lumbar and dorsal spine and the pelvis. BMD was evaluated using dual-energy X-ray absorptiometry and the examination was performed in the distal radius. In DISH patients the mean value of BMD was significantly higher than in controls (P<0.002), even when it was referred to sex subgroups. Statistical analysis showed significant differences between both the two male groups (P<0.002) and the two female groups (P<0.01). In the two female subgroups (DISH patients and controls) BMD was significantly inversely related to age and to the duration of the postmenopausal period. The present study showed higher BMD in DISH patients than in the control group. PMID- 10870652 TI - Questionnaire on NSAID gastropathy among Dutch rheumatologists. AB - Dutch rheumatologists were given a questionnaire on NSAID gastropathy, in which older age (>60 years) and previous ulcers were mentioned by them as being the most important risk factors. Dutch rheumatologists follow different strategies for the prevention of NSAID gastropathy, with a slight preference for proton pump inhibitors and the use of COX-2 selective NSAIDs. PMID- 10870653 TI - How does the short form 36 health questionnaire (SF-36) in rheumatoid arthritis (RA) relate to RA outcome measures and SF-36 population values? A cross-sectional study. AB - The aim of the study was to show that the SF-36 is a practical tool for use on outpatients with RA, to examine the relationship between the SF-36 and indices of outcome in RA, and to compare the results with population norms and other disease states. Eighty-six consecutive RA patients attending the Haywood Hospital in Stoke-on-Trent and starting or changing second-line therapy were enrolled. Disease outcome was assessed using the American College of Rheumatology core set and all subjects completed the SF-36 health questionnaire. The cohort had moderately active disease (median ESR 46) and appreciable disability (median HAQ 1.875). Impairment of health status was moderate to marked by the SF-36, with significant differences from population norms and chronic disease states such as low back pain. Good correlations were observed between HAQ and physical function (r>0.75, p<10(-6)) and HAQ and social function (r>0.61, p<10(-6)). In contrast, SF-36 scales for physical and emotional role showed no association with activity measures. We concluded that, SF-36 is a practical tool for use in patients with RA. HAQ is associated with its physical and social function scales. Other SF-36 scales, such as physical and emotional role, are not associated with activity core set measures; this suggests different information is involved. RA has a considerable impact on health status compared to other diseases. PMID- 10870654 TI - Antinuclear antibody (ANA) and ANA profile tests in children with autoimmune disorders: a retrospective study. AB - The study objective was to determine the clinical value of positive antinuclear antibody (ANA) and ANA profile tests in children with autoimmune disorders. A retrospective chart review was carried out of all patients under 18 years of age with a positive ANA test (HEp-2 cell substrate, titre > or =1:40) and ANA profile (ELISA) referred to the paediatric rheumatology service at the authors' institution between 1992 and 1996. Of 245 children with a positive ANA test, 134 (55%) had an autoimmune disease, including juvenile rheumatoid arthritis (n = 49), systemic lupus erythematosus (SLE) (n = 40) and others (n = 45). The remaining 111 patients did not have identifiable autoimmune diseases. Patients with autoimmune disorders had significantly higher ANA titres of > or = 1:160 (chi2 = 16, P<0.0001). In addition, of the 245 patients with a positive ANA test, 86 had an ANA profile performed; this was positive in 32 and negative in 54. All 32 patients with a positive ANA profile (100%) had an autoimmune disorder, compared to 22 (41%) of 54 with a negative ANA profile who had autoimmune disorders. Of 22 SLE patients with a positive ANA profile, 16 (73%) had positive anti-dsDNA and 15 (68%) had positive anti-Sm and positive anti-RNP. A positive ANA profile correlated strongly with an ANA titre > or = 1:640 (chi2 = 5.7 , P<0.02). The study demonstrated that only 55% of children with a positive ANA test had a definitive diagnosis of autoimmune disorder. These children tend to have higher ANA titres of > or =1:160. However, a positive ANA profile was strongly correlated with an ANA titre > or =1:640 and highly indicative of an autoimmune disorder (100%). We suggest that in order to reduce cost, an ANA profile should not be performed on all patients with positive ANA, but reserved for those with an ANA titre of > or =1:640 and/or those with a high clinical index of suspicion for autoimmune disorder, especially SLE. PMID- 10870655 TI - Intra-articular treatment with hyaluronic acid. Comparative study of Hyalgan and Adant. AB - Forty-nine patients diagnosed as having gonarthrosis were given intra-articular treatment with hyaluronic acid (Adant or Hyalgan) in a blind randomised study. We concluded that the efficacy with Adant at 3 months after treatment was greater than with Hyalgan (50% versus 21.1%). The maximum improvement with hyaluronic acid was seen at 5 weeks in 75.4% and the adverse effects consisted of pain in the infiltration side which was almost twice as great with Adant (16.3%). PMID- 10870656 TI - Skin score decrease in systemic sclerosis patients treated with intravenous immunoglobulin--a preliminary report. AB - The aim of the study was to determine for the first time the response of systemic sclerosis (SSc) patients to treatment with intravenous immunoglobulin (IVIg). Three patients with progressive and rapidly deteriorating disease (mainly affecting the skin) were planned to receive six monthly courses of high-dose IVIg (2 g/kg). All had a thorough physical examination, clinical evaluation by the modified Rodnan total skin thickness score, and measurement of the titres of PM Scl antibodies before and after the treatment, and before and after each treatment course. Two of the three patients received six IVIg courses as planned and no adverse effects or disease progression occurred during the therapy. The third patient received three courses, after which he developed renal failure and later died of sepsis. All three patients had a large decrease in their skin score after the treatment compared to that before the treatment. No modification of PM Scl antibody titres was noted in any patient. Intravenous immunoglobulin (IVIg) may have a role in the treatment of SSc patients with rapidly deteriorating skin disease. The specific indications, as well as the safety of this treatment, should be further researched. PMID- 10870657 TI - Erythema nodosum: the underlying conditions. AB - Erythema nodosum (EN) is a cutaneous reaction consisting of inflammatory, tender nodular lesions and is associated with a wide variety of disease processes. The aim of our study was to investigate the frequency of different aetiologies of EN. One hundred and thirty-two EN patients were investigated in a prospective study during the period 1984-1990. The evaluation of all patients began with a medical and family history and completed with a thorough physical examination and detailed laboratory and immunological work-up. In addition, various diagnostic procedures were performed where and when indicated. One hundred and ten patients (83%) were women. Their mean age was 41.0+/-14.0 years, range 18-79 years. In 35% the cause of EN was not found. Sarcoidosis was revealed in 28% of the patients, infections in 17.3% and tuberculosis in 1.5%. Other aetiologic factors were Adamantiadis-Behcet's syndrome (3.8%), pregnancy (6%), oral contraceptives (3.8%) and other drugs (3.8%). The aetiology of EN was not found in 35% of the patients. Sarcoidosis and infections were frequent causes of EN, whereas autoimmune rheumatic diseases rarely cause EN. PMID- 10870658 TI - Prevalence of pulmonary disorders in patients with newly diagnosed rheumatoid arthritis. AB - Our objective was to determine the prevalence of airway hyperreactivity (AHR) in patients with newly diagnosed rheumatoid arthritis (RA) who had received no disease-modifying antirheumatic drugs (DMARDs) and to characterise the spectrum of lung diseases identifiable in these patients at the time of presentation. Eighteen consecutive patients with newly diagnosed RA referred to our medical centre's rheumatology clinic over 2 years underwent pulmonary evaluation with arterial blood gas analysis, chest radiographs, spirometry before and after bronchodilator medication, and body plethysmography. They returned on subsequent days in random order for methacholine inhalation challenge (MIC) and eucapnic voluntary hyperventilation (EVH) as bronchoprovocation techniques. One patient had severe obstructive disease at presentation and therefore did not undergo bronchoprovocation. We found a wide variety of pulmonary abnormalities, including two patients with hypoxia (12%), two with obstruction (12%), three with restriction (18%) and four with AHR (23%). The data also suggest a strong association between pulmonary diseases in RA and cigarette smoking. Although no single characteristic lung disease such as AHR was identified in patients presenting with RA, the association between lung disease and cigarette smoking is striking and underscores the need to emphasise smoking cessation in this patient population. PMID- 10870659 TI - Acute hepatitis in adult Still's disease apparently resulting from oral iron substitution--a case report. AB - The authors report a case of a young woman with adult-onset Still's disease (AOSD) with massive hyperferritinaemia who developed acute florid hepatitis with intraparenchymatous histiocytic infiltration following oral iron substitution for presumed iron deficiency, which settled on withdrawal of the iron. This suggests that the iron exacerbated the macrophage hyperactivity which is presumed to be present in AOSD. Oral iron substitution in the acute phase of this disease may be inadvisable. PMID- 10870660 TI - Takayasu's arteritis-associated aneurysm formation without steno-occlusive lesions. AB - The authors report a case of a 20-year-old woman with Takayasu's arteritis (TA) presenting with recurrent erythema nodosum-like lesions, elevated acute-phase proteins and aortographic findings of multiple aneurysmal dilatations of the aorta without the coexistence of steno-occlusive lesions. This finding indicates that aneurysms could be an early manifestation of TA and not necessarily a change secondary to stenotic lesions. PMID- 10870661 TI - Synovitis of the wrist joint caused by an intra-articular perforation of an osteoid osteoma of the radial styloid. AB - An osteoid osteoma in the radial styloid with perforation into the radiocarpal joint was observed. Resection cured the patient. PMID- 10870662 TI - Neuro-Behcet's syndrome in a patient not fulfilling criteria for Behcet's disease: clinical features and value of brain imaging. AB - Central nervous system involvement is rarely an initial presenting manifestation of Behcet's disease (BD). We report the case of a 33-year-old man with recurrent attacks of fever, oral mucosal ulcers, deep venous thrombosis, diplopia, vertigo and headache. Sequential brain magnetic resonance imaging (MRI) scans showed fluctuating lesions of the brain stem, mesencephalon and thalamus. F-18-fluoro-2 deoxy-D-glucose positron emission tomography (FDG-PET) revealed hypometabolism at the parieto-occipital cortex at both sides and the brain stem. Treatment with prednisone and cyclosporine A led to a complete remission and normalisation of MRI and FDG-PET lesions. The present case illustrates the difficulty in the differential diagnosis of early neuro-BD. PMID- 10870663 TI - Inflammatory demyelinating polyradiculoneuropathy associated with interstitial lung disease. AB - A 58-year-old woman presented with inflammatory demyelinating polyradiculoneuropathy accompanied by sensory and motor disturbance and interstitial lung disease. Corticosteroid therapy led to a marked amelioration of both the neuropathy and the lung disease. We suggest that a demyelinating neuropathy is associated with an interstitial lung disease. PMID- 10870664 TI - Spondyloepiphyseal dysplasia tarda with progressive arthropathy. AB - Spondyloepiphyseal dysplasia tarda with progressive arthropathy, described by Wynne-Davies et al., is a rare autosomal recessive disorder. It is characterised by generalised platyspondyly and epiphyseal involvement, with enlargement of both ends of the short tubular bones of the hands. Clinical features include onset in childhood, a disproportionately short stature and premature osteoarthritis. We describe the clinical and radiographic findings of a young woman suffering from spondyloepiphyseal dysplasia tarda with progressive arthropathy. PMID- 10870666 TI - Hypertrichosis in systemic lupus erythematosus (SLE). AB - We describe a 14-year-old female with systemic lupus erythematosus (SLE) involving the skin, joints and central nervous system who developed hypertrichosis of the upper eyelashes. This clinical finding has been observed in immunocompromised patients with acquired immune deficiency syndrome (AIDS), malnutrition, cancer or kala-azar. Although the pathogenic mechanism for this type of hypertrichosis is unknown, we believe the immunological defects seen in SLE may be responsible for such manifestation in our patient. PMID- 10870665 TI - Naproxen-induced leukocytoclastic vasculitis. AB - Cutaneous reactions to non-steroidal anti-inflammatory drugs (NSAIDs) are rare in spite of their wide use. Only a few cases of hypersensitivity angiitis related to naproxen have been described. We report the case of a 62-year-old woman in whom leukocytoclastic skin vasculitis, peripheral neuropathy and nephritis developed after a short naproxen treatment, and gradually regressed after discontinuation of the drug and under glucocorticoid therapy. In the light of the relevant literature, the clinical and laboratory features of this reversible condition are described. PMID- 10870667 TI - Remitting seronegative symmetrical synovitis with pitting oedema associated with chronic lymphocytic leukaemia. AB - We describe a case of remitting seronegative symmetrical synovitis with pitting oedema (RS3PE syndrome) in a 67-year-old man. Immunophenotyping studies and histology of biopsy specimens revealed chronic lymphocytic leukaemia (CLL). Polyarthritis and oedema were revealed by small doses of corticosteroids. The association of RS3PE syndrome with rheumatological and malign disorders are discussed. PMID- 10870668 TI - Evaluation of pathergy test in North Jordan. AB - The positivity of pathergy's test in Behcet disease varies throughout the world. We tried to evaluate its diagnostic significance in known Behcet patients in the region of North Jordan. Twenty-two patients were tested, readings were taken after 48 hours. The patients were not tested previously and a comparison between the subcutaneous injection of 0.1 ml normal saline and skin puncture was recorded. The development of a skin eruption (papule, nodule, pustule) were observed and described. The percentage of positivity was 20% in the studied population. The patients tested who were not taking medication at the time of the procedure are more likely to illustrate a positive Pathergy reaction. PMID- 10870669 TI - Study of immunological and virological parameters during thalidomide treatment of SIV-infected cynomolgus monkeys. AB - The potential therapeutic utility of thalidomide (Thd), an effective inhibitor of tumor necrosis factor (TNF)-alpha in vitro, was investigated in cynomolgus monkeys (Macaca fascicularis) at 10 months after infection with simian immunodeficiency virus (SIV). Thd-treated macaques (n = 8) received an oral dose (10 mg) daily for 7 days, followed by a wash-out period of 5 weeks. A 2nd cycle of treatment was performed on the same animals at higher doses (20 mg Thd/day) for 14 days. The control monkeys (n = 7) received a placebo for the same period of time. In the present study, we show that Thd, in addition to inhibiting TNF alpha production after in vitro mitogen stimulation of peripheral blood mononuclear cells (PBMCs), was able to restore the proliferative responses to SIV peptides in monkeys that were infected with SIV. Interestingly, we found that such effects are associated with an increased expression of CD28 cell surface receptors on CD4+ T-cells paralleled by a decrease on CD8+ T-cells. At the same time, significant reduction in either cell-associated viral load or plasma viral RNA was not observed among the SIV-infected monkeys during the two treatment cycles, when compared with the placebo group. PMID- 10870670 TI - Age and gender differences in body composition, energy expenditure, and glucoregulation of adult rhesus monkeys. AB - The purpose of this study was to examine the relationship of age to body composition, glucoregulation, activity, and energy expenditure in male and female rhesus monkeys. The animals were studied in three groups, young adults (YA, 7-9 years), middle-aged adults (MA, 13-17 years), and older adults (OA, > 23 years) adults. OA had a lower (P < 0.05) lean body mass than the YA and MA. OA also had the lowest values (P < 0.06) for energy expenditure (kJ/minute). Age-related differences (P < 0.05) were observed in time spent resting and moving. The OA spent the most time resting and the least time in vertical movement. There was a trend towards an age-related decrease in acute insulin response to glucose, while other glucoregulatory parameters were not changed with age. These results are similar to findings in humans, providing further evidence that the rhesus monkey is an appropriate model of human aging. PMID- 10870671 TI - Dietary content may prevent secondary hyperparathyroidism in female rhesus monkeys (Macaca mulatta). AB - Macaque laboratory chows provide relatively more calcium (Ca) and vitamin D (D) than human diets; this may influence skeletal aging. To evaluate this possibility, parameters of skeletal relevance in premenopausal and naturally postmenopausal rhesus monkeys were measured in a cross-sectional study. Serum osteocalcin (Oc) was elevated in the postmenopausal group (P < 0.01), but levels of parathyroid hormone (PTH) and 25-hydroxyvitamin D (25OHD) were not different. Subsequently, in premenopausal animals, dietary Ca and/or D intake was reduced to optimal human levels for 8 weeks prior to the evaluation of the skeletal parameters. Serum 25OHD concentration was reduced (P < 0.01) and a trend (P=0.10) towards increased PTH was observed in both low D groups. In addition, serum alkaline phosphatase (ALP) levels were increased in the low Ca group (P < 0.01). In conclusion, skeletal turnover, as measured by serum Oc, was increased in naturally postmenopausal rhesus monkeys in the absence of hyperparathyroidism. Dietary D reduction causes a decline in serum 25OHD and an upward trend in PTH. PMID- 10870672 TI - Changes of hormonal function of the adrenal and gonadal glands in baboons of different age groups. AB - The purpose of this study was to characterize the changes of hormonal function of the adrenals and gonads during aging in male baboons (Papio hamadryas). Basal levels of plasma dehydroepiandrosterone, dehydroepiandrosterone sulfate, pregnenolone, and 17-hydroxypregnenolone progressively decrease with age from 10 15 years when analyzed by specific radioimmunoassay. However, no significant changes were found in cortisol and 11-desoxycortisol concentrations. The levels of sexual hormones did not differ in young and mature groups. In the 20-26-year old animals, the concentration of testicular androgens showed a tendency to decrease, while the concentration of biologically active luteinizing hormone (LH) showed a tendency to increase. The old animals exhibited a decrease of plasticity of the pituitary testicular system, which was manifested in the deceleration of the decrease of LH and T concentrations after the peak values had been reached in response to luteinizing hormone-releasing hormone (LHRH) administration. The oldest male also developed some refractoriness of the pituitary gonadal system to the prolonged administration of LHRH agonist. The hormonal imbalance which develops with age may play an important role in the age-related involutional process. PMID- 10870673 TI - Vaginal myeloperoxidase and flora in the pig-tailed macaque. AB - Myeloperoxidase (MPO) is an enzyme in neutrophils and monocytes which reacts with H2O2 and chloride to kill microbes after phagocytosis. Instillation of MPO into the vagina may augment vaginal defenses against sexually transmitted diseases, since the normal vaginal flora is characterized by the presence of H2O2-producing lactobacilli. We assessed the menstrual cycle stage, vaginal flora, pH, macroscopic appearance, and endogenous MPO in the adult female pig-tailed macaque (Macaca nemestrina) at baseline (n = 26; 60 observations) and at 0, 4, and 24 hours in untreated animals (n = 6) or in animals treated with intravaginal MPO gel at time 0 (n = 5). Baseline MPO levels were highly variable, and there was no detectable effect of cycle stage. In untreated animals, there was no significant effect of vaginal swab collection on vaginal flora or MPO levels. MPO treatment did not reduce vaginal H2O2-producing organisms, and vaginal MPO levels tended to increase at 4 hours in treated animals. Vaginal/cervical colposcopic changes were not detected in either group. PMID- 10870674 TI - Dilative cardiomyopathy leading to congestive heart failure in a male squirrel monkey (Saimiri sciureus). AB - A 17-year-old, 1-kg, colony-housed, male squirrel monkey (Samiri sciureus) developed clinical signs of congestive heart failure. The monkey presented with lethargy, increased heart and respiratory rates, and mild abdominal distention. The clinical history, laboratory analysis, and radiographic findings were consistent with heart failure due to dilative cardiomyopathy. Gross and microscopic examination of the heart confirmed a dilative cardiomyopathy. This is the first report describing congestive heart failure caused by dilative cardiomyopathy in a squirrel monkey. Spontaneous dilative cardiomyopathy may be infrequently observed in the squirrel monkeys because they are not routinely housed in the research environment during their advancing years. PMID- 10870675 TI - Our animal model of coronary spasm--my personal view. AB - We developed an animal model of coronary spasm in swine, similar to coronary spasm in patients with variant angina based on the angiographic findings. In this animal model, an impairment of endothelium dependent dilatation appeared to play a minor role while the hypercontraction of the medial muscle cells by histamine and serotonin at the spastic site played a major role in the induction of coronary spasm. In Gottingen male miniature swine receiving focal endothelial denudation, moderate hypercholesterolemia and X ray irradiation, the abrupt, severe and prolonged coronary spasm resulted in a sudden progression of organic coronary stenosis mainly due to intraplaque hemorrhage and also in acute myocardial infarction. PMID- 10870676 TI - QT dispersion is increased in diabetic patients with foot ulcer. AB - QT dispersion, a measure of inhomogenous ventricular repolarization, was measured in diabetic patients with foot ulcer. We recruited 75 patients with non insulin dependent diabetes mellitus: patients with neuropathic ulcer (n=15, NU group), with ischemic ulcer (n=20, IU group), with previous myocardial infarction (n=20, MI group) and without any diabetic microangiopathies (n=20, DC group). We also studied normal control subjects (n=15, NC group). The interlead variability of rate corrected QT interval (QTc dispersion) was calculated. QTc interval in the MI group was significantly higher than that in the NC or DC but showed no difference in the NU and IU groups. QTc dispersion in the IU (54+/-15 msec) as well as MI (60+/-21 msec) group were significantly higher than the NC (36+/-18 msec) or DC group (39+/-14 msec). This may be due to complicated coronary artery disease in the IU group. Furthermore, QTc dispersion was also increased (49+/-14 msec) in the NU group in which cardiac autonomic nervous dysfunction was suggested. Patients with both types of diabetic ulcer demonstrated increased QT dispersion due to atherosclerosis or neurological disorder. PMID- 10870677 TI - Use of a temporary caval filter in a young man with pulmonary embolism to prevent migration of massive caval thrombus during an attempt of caval thrombolysis. AB - A 33 year old male with no known risk factors for hypercoagulability developed a massive thrombi in the inferior vena cava (IVC). The patient had a history of both pulmonary embolism and embolism related syncope. The thrombus which extended proximally to the level of the renal vein and distally to the left superficial femoral vein did not respond to anticoagulant therapy or thrombolysis. Thirteen days after admission, we decided to use a temporary caval filter to provide protection from migration of the thrombus while attempting invasive thrombolytic therapy, which was performed using a tissue type plasminogen activator through a coaxial catheter of the temporary filter. This resulted in a marked decrease in the size of the thrombus, and multiple thrombi were found to be trapped in the temporary filter. Although the temporary caval filter was effective in capturing emboli, resulting in a decrease in the thrombus size, the thrombus was not completely dissolved within two weeks, which is the maximal implantation time. A permanent filter was eventually used to prevent pulmonary embolism, which could arise from the remaining thrombus. We have found placement of a temporary caval filter to be a safe and effective adjunct, in select cases, when attempting thrombolysis of massive thrombi in the IVC. Since we inserted the temporary filter 13 days after admission, use of a temporary filter during thrombolysis may have been more effective if conducted earlier in our patient's clinical course. PMID- 10870678 TI - Apolipoprotein E accelerates the efflux of cholesterol from macrophages: mechanism of xanthoma formation in apolipoprotein E deficiency. AB - A patient with congenitally deficient apolipoprotein (apo) E showed numerous tuberoerutive, tendon xanthomas and severe atherosclerosis, despite a low LDL concentration. In order to study the mechanism of xanthoma formation observed in apo E deficient patients, we evaluated the effect of VLDL and HDL from the patient on cholesterol ester (CE) accumulation in macrophages. The results showed that there was no difference in CE formation in macrophages among normal VLDL, the patient's VLDL and apo E containing VLDL, which was prepared by incubation of the patient's VLDL with recombinant apo E. On the other hand, apo E containing HDL, which was prepared by incubation of the patient's HDL with recombinant apo E, accelerated cholesterol efflux more effectively than did the patient's HDL and decreased intracellular CE content. Moreover, free apo E accelerated cholesterol efflux from lipid loaded macrophages. These results suggest that macrophages are prevented from transforming into foam cells by their secretion of apo E. This may also explain the marked atherosclerosis and xanthomatosis observed in the patient with apo E-deficiency. PMID- 10870679 TI - Association between basal serum and leptin levels and changes in abdominal fat distribution during weight loss. AB - The aim of this study was to evaluate the relationship between changes in abdominal fat areas and the baseline serum leptin levels of Japanese obese women during weight reduction. The study was performed on 100 obese female Japanese volunteers. We measured the BMI and abdominal fat areas (visceral, subcutaneous and total) by magnetic resonance imaging and determined the fasting serum leptin levels before and after a 3 month weight reduction program. We examined whether or not a relationship exists between the baseline leptin levels and the subsequent changes in the abdominal fat areas after a weight reduction program. Multiple linear regression analysis was performed to examine the relationship between the baseline leptin levels and changes in abdominal visceral, subcutaneous, and total fat areas, and demonstrated that the baseline leptin level was a significant predictive factor for changes in the abdominal visceral fat area in both pre and postmenopausal Japanese obese women. We thus concluded the relatively higher baseline leptin levels in Japanese obese women to be associated with a relatively smaller reduction in the amount of abdominal visceral fat after undergoing a weight reduction program. PMID- 10870680 TI - Tamoxifen in cancer therapy: minireview. AB - Tamoxifen belongs among relatively new drugs. As it has already been shown, it undoubtedly brings a benefit to oncology patients. However, there are still questions regarding its broader use in therapy or cancer prevention. This review puts together some data available at present time with the aim of elucidating the most important aspects of its use in medical oncology. PMID- 10870681 TI - Immunohistochemical analysis of expression of a 65 kDa oncofetal protein (p65), epidermal growth factor receptor (EGFR), oncogene c-erb B2 and tumor suppressor gene p53 protein products in breast cancer patients. AB - Paraffin-embedded tissue slides from 88 infiltrating ductal breast carcinoma were examined by immunohistochemistry technique with the use of monoclonal antibody against human p65 antigen and polyclonal antibody against p65-like protein present in fetal bovine serum. Immunohistochemical analysis of expression of growth factor receptors (EGFR), protein product of oncogene c-erb B2 as well as protein product of mutated anti-oncogene p53 was also done. It was established that there is no correlation between p65 and c-erbB2, EGFR or p53 expression. In low differentiated tumors (grade III) high p53 index and high EGFR and c-erbB2 expression was connected with low p65 expression. The lack of c-erbB2 and EGFR and low p53 expression was combined usually with high p65 oncoprotein levels. PMID- 10870682 TI - Influence of estrogen, antiestrogen and UV-light on the balance between proliferation and apoptosis in MCF-7 breast adenocarcinoma cells culture. AB - Studies of the mechanism of actions of estrogen, antiestrogen and physical factors may provide clues to an understanding of breast cancer growth and/or regression regulation and thus identify novel targets for therapeutic intervention. Defective control of apoptosis appears to play a central role in the pathogenesis of neoplasia. Conversely, cancer therapy and ionizing radiation can induce cancer cell death by apoptosis and/or necrosis. bcl-2 gene and p-53 gene products have been both linked to programmed cell death pathways. We have analyzed the effect of estradiol, tamoxifen and UV exposure on the induction of apoptosis, expression of p53 and bcl-2 gene products as well as the proliferative activity (expressed as [3H]thymidine incorporation and PCNA and MPM2 antigens involvement) in MCF7. It has been found that estradiol increases the speed of cell cycle in MCF7 and acts as antiapoptotic factor. Tamoxifen has multiple influence on the rate of growth of cancer cells: depends on estrogen receptor (ER), conducts reduction of proliferation rate; depends on ER and other mechanisms conducts to suppressions of Bcl-2 protein expression and induction of cell death through apoptotic pathway. Estradiol prevents the apoptotic influence of tamoxifen probably by enhancement of Bcl-2 protein expression and does not prevent the inhibition of proliferation rate. The irradiation with UV induces apoptosis by over-expression of p53 and down-regulation of bcl-2 gene. PMID- 10870683 TI - Radiation-induced apoptosis and cell cycle alterations in human carcinoma cell lines with different radiosensitivities. AB - Radiosensitivity of examined human neoplastic cell lines was assessed with the aid of MTT assay. Differences between radiosensitive and radioresistant human neoplastic cell lines were as follow: a) radiation-induced apoptosis detected by flow cytometry was apparent in the most radiosensitive (i.e. CH-1 ovarian carcinoma cell line), but not in the radioresistant (i.e. SKOV-3 ovarian carcinoma) cell lines, b) radiation-induced G2/M arrest appeared early after irradiation (6 hours) in both the radioresistant SKOV-3 cells and in the radiosensitive CH-1 human ovarian carcinoma cell line, but a different pattern was observed 24 hours after irradiation with 2 Gy dose with G2/M arrest only in radiosensitive cell line. The radiosensitivity and resistance to radiation induced apoptosis in the radioresistant human breast carcinoma MDA-MB-231 cell line were similar to those observed in SKOV-3 cells. These data suggest that radiation-induced apoptosis and cell cycle alterations can predict radiosensitivity at least in some examined human malignant cells in vitro. PMID- 10870684 TI - Effect of epinephrine and combination treatment of epinephrine and heat on melanoma cell lines. Study of cytotoxicity and kinetics of 72-kD stress protein. AB - To elucidate the effect of epinephrine and the combination with heat on malignant cells, using two melanoma cell lines, HM6KO and G361, we have examined the cytotoxicity and induction of 72-kD stress protein (HSP72) after the treatments. After epinephrine treatment, in both cell lines, cell survival rates decreased gradually in a concentration-dependent manner. After the combination treatment, cell survival rates decreased more than those when two treatments were done separately. The cytotoxicity of epinephrine was more enhanced in G361 than in HM6KO by heat. After epinephrine treatment, in both cell lines, the level of HSP72 did not elevate. After combination treatment, in HM6KO, the level of HSP72 were higher than those of heat alone. In G361, the kinetics of HSP72 level was similar to that of heat alone. These results suggest that epinephrine has a cytotoxicity to melanoma cells and the cytotoxicity is enhanced by the combination. In addition, it is probable that epinephrine does not have HSP72 inducibility in HM6KO and G361, and the different kinetics of HSP72 between the cell lines in the combination treatment may play an important role to determine the degree of enhancement. PMID- 10870685 TI - Blood levels of natural antioxidants in gastric and colorectal precancerous lesions and cancers in Slovakia. AB - A long-term sufficient intake of fruits and vegetables reduces significantly the risk of gastric and colorectal carcinoma. It is anticipated that natural antioxidants are involved in this effect in addition to other substances. The aim of this study was to determine levels of vitamins A, C and E, as well as beta carotene, selenium, zinc and copper in blood of 249 patients with precancerous lesions (atrophic gastritis, gastric hyperplastic polyp, gastric, colonic and rectal adenoma, chronic ulcerative colitis) and in 96 individuals with gastric, colonic or rectal carcinoma and to compare these levels with the values of a control group of 130 healthy individuals. We have found that the frequency of average values of analyzed micronutrients in precancerous groups was decreasing in the order vit C > vit E/vit A > Se > beta-car. The average levels of vitamins and beta-carotene were significantly reduced in all carcinoma groups, while selenium level showed a decrease only in the gastric carcinoma group. Copper level was elevated in the ulcerative colitis group and in all groups with carcinoma. The results indicate a frequent insufficient saturation of organism by natural antioxidants in groups with precancerous lesions and carcinomas of stomach and colorectum. Therefore, it is necessary to increase the general consumption of fruits and vegetables in Slovakia as a part of primary prevention of malignant diseases in these organs. Chemoprevention may be recommended in individuals with precancerous lesions. PMID- 10870686 TI - Do de novo acute myeloid leukemias with normal cytogenetics involve two main prognostic categories distinguished by the presence of erythroblastic and/or megakaryocytic dysplasia? AB - De novo acute myeloid leukemias (AML) patients with normal cytogenetics represent a standard risk cytogenetic group. Erythroblastic and/or megakaryocytic dysplasia (EMD) in diagnostic bone marrow smears of 28 consecutive AML patients with a normal karyotype was studied. Twelve patients 21-85 (median 48) years old were categorized without EMD, 14 patients 34-90 (median 58) years old with EMD, and 2 patients were not evaluable for EMD. One cycle of induction therapy 4 + 7, with 4 doses of daunorubicin 45 mg/m2/d and standard doses of cytosine arabinoside for 7 days induced 10 complete and 2 partial remissions in 12 cases without EMD but lead to only one complete remission, 6 non-responses and 3 induction deaths in 10 cases with EMD (p = 0.002). However, high doses of cytosine arabinoside plus daunorubicin induced complete remission in 6 of 7 patients with EMD. In patients under 66 years treated by intensive consolidations the estimate of median survival was 50.6 months in 10 cases without EMD, significantly higher than 8.0 months in 11 cases with EMD (p = 0.043). De novo AML with normal cytogenetics might be divided into two biological categories, the first favorable-risk category without EMD and the second poor-risk category with EMD. PMID- 10870687 TI - Chromosomal changes in somatic cells in seminoma patients after treatment with ionizing radiation or cytostatics. AB - Seminomas are sensitive to both ionizing radiation and cytostatic drugs. The study's objective was to find out the effects of cytostatics or ionizing radiation by comparing the results of genome testing before treatment and immediately afterwards. Repeat cytogenetic testing six months after completion of treatment was used to find out changes resulting from reparatory processes after various types of treatment and the degree of elimination of defective lymphocytes from circulation. Three cytogenetic tests were used in our study to find out structural changes in chromosomes (percentage of aberrations), sister chromatid exchanges (SCE) and the number of micronuclei in binuclear lymphocytes (MN). In patients treated with ionizing radiation, strong inhibition of the mitotic activity of lymphocytes occurred after irradiation of para-aortal and ipsilateral inguinal lymph nodes. However, it is difficult to make a connection between the mitotic activity of lymphocytes and their total number, which was found to be within a normal range throughout the study. There is, therefore, another possibility, i.e. that this process actually involves impairment of intracellular enzymes and blockage of the synthesis of macromolecules in lymphocytes which have suffered a large degree of genome damage. Six months after a completed course of irradiation, mitotic activity was found to be mostly normal; however, there was still a very high percentage of aberrations compared with group II (patients treated with a cytostatic, paraplatin) or with respect to the control group, in which the average percentage of aberrations was 1.42 (excluding dicentrics and rings, which are found in all irradiated patients). From the cytological mutagenetic point of view, chemotherapy proved to be less aggressive to patients. The results of recovery were visible earlier and the elimination of damaged cells was quicker. PMID- 10870688 TI - HDR intraluminal brachytherapy in the treatment of malignant bronchial obstructions. AB - Symptomatology of malignant intrabronchial obstructions has a serious negative effect on the quality of patients' life. Intrabronchial brachytherapy can play an important role in the palliation of these symptoms. Between December 1996 and September 1998 48 patients suffering from malignant intrabronchial obstructions were treated with intraluminal brachytherapy in the Dept. of Radiation Oncology at the University Maternity Hospital in Brno. Gammamed HDR automatical afterloading equipment was used to treat all patients. The first group (23 patients) was treated with a combination of intraluminal brachytherapy and external radiotherapy. The second group (18 patients who had relapsed after previous external radiotherapy) was given intraluminal radiotherapy only. A third group (7 patients) underwent intraluminal brachytherapy only. In the first group 17 patients (77%) showed symptomatic relief with tumor regression on X-ray in 16 patients and with bronchoscopic regression in 19 patients. Seven patients died before October 1998 having survived 1-6 months after the first brachytherapy application. Sixteen patients are still alive (1-14 months). In the second group, 10 patients (56%) reported significant improvement of symptoms, with endoscopic regression in 12 patients. Twelve patients died before October 1998 surviving 1-6 months after the first brachytherapy session, 6 patients are still alive 1-5 months after the first brachytherapy fraction. In the third group, bronchoscopy confirmed a complete disappearance of intrabronchial lesion in two cases with early intrabronchial tumor. Five patients reported symptomatic improvement with endoscopic regression of the tumor. There was only one complication recorded: bronchospasm in one patient. The short follow up and limited number of patients does not allow comment on the late effects and survival, yet. In conclusion, intraluminal brachytherapy is an effective and safe approach for palliation of malignant bronchial obstructions. PMID- 10870690 TI - Recent trends in cancer mortality in the Slovak Republic and in Europe. AB - During the last 30 years the trends in cancer mortality in Europe significantly changed. In 1970 the male and female cancer mortality (all ages) was higher in the Western Europe than in the "socialist" Central and Eastern Europe. After 1970 in most Western countries decrease or no change of male premature (0-64 years) cancer mortality was observed. The decrease was deepest in Finland and in United Kingdom. In the most of the former communist countries an increase of both total and premature male cancer mortality was observed, especially in Hungary, Poland, Roumania, Bulgaria and in some regions of the former Soviet Union. Present male premature cancer mortality in Hungary is two-times higher than the average of European Union. Male cancer mortality in the Slovak Republic is at least two times higher than in United Kingdom, Switzerland or Sweden. These differences are partially explainable by the higher prevalence of smoking in the East. The further risk factor could be oxidative stress, caused by low intake of antioxidants and high intake of spirits. In female populations, the differences between East and West are not so dramatic, with the exception of extremely high mortality in Hungarian females. Parallel increase of female lung cancer mortality both in the West and East is caused probably by the continually increasing smoking prevalence in females almost in the whole Europe. Further local risk factors in Eastern Europe (e.g. pollution) need to be identified with more specificity for preventive programs in Eastern Europe. This region is a prospective area for the research on lesser known cancer risk factors, e.g., chronic deficiency of antioxidants, natural anticarcinogens and psychosocial disorders. PMID- 10870689 TI - Synergistic cell killing by combination therapy of retinoic acid and hyperthermia in human epidermoid laryngeal carcinoma cells in culture. AB - In vitro monolayer culture and clonogenic assay were used to investigate the individual and combined effect of temperature and retinoic acid (RA) on cellular morphology and colony forming ability of human epidermoid laryngeal carcinoma (HEp-2) cells. 20 micromol. RA alone inhibited multilayer formation and induced cell flattening. Hyperthermia (42 degrees C) individually caused formation of cytoplasmic processes and irregularities in cellular shape and size. Combined effect of hyperthermia (42 degrees C) and 20 micromol. RA treatment caused bleb formation on cell surfaces and lysis of cytoplasmic and nuclear membrane. RA treatment also caused dose-dependent reduction of colony growth. Heat-induced cell killing was only observed at lethal temperatures of 43 degrees C and above. RA in combination with heat synergistically inhibited colony formation even at non lethal temperatures of 41 and 42 degrees C. These results indicate that RA in combination with hyperthermia may facilitate the therapy of human epidermoid larynx carcinoma. PMID- 10870691 TI - Determination of rice herbicides, their transformation products and clofibric acid using on-line solid-phase extraction followed by liquid chromatography with diode array and atmospheric pressure chemical ionization mass spectrometric detection. AB - A simultaneous method for the trace determination of acidic, neutral herbicides and their transformation products in estuarine waters has been developed through an on-line solid-phase extraction method followed by liquid chromatography with diode array and mass spectrometric detection. An atmospheric pressure chemical ionization (APCI) interface was used in the negative ionization mode after optimization of the main APCI parameters. Limits of detection ranged from 0.1 to 0.02 ng/ml for 50 ml of acidified estuarine waters preconcentrated into polymeric precolumns and using time-scheduled selected ion monitoring mode. Two degradation products of the acidic herbicides (4-chloro-2-methylphenol and 2,4 dichlorophenol) did not show good signal response using APCI-MS at the concentration studied due to the higher fragmentor voltage needed for their determination. For molinate and the major degradation product of propanil, 3,4 dichloroaniline, positive ion mode was needed for APCI-MS detection. The proposed method was applied to the determination of herbicides in drainage waters from rice fields of the Delta del Ebro (Spain). During the 3-month monitoring of the herbicides, 8-hydroxybentazone and 4-chloro-2-methylphenoxyacetic acid were successively found in those samples. PMID- 10870692 TI - Monitoring of priority pesticides and other organic pollutants in river water from portugal by gas chromatography-mass spectrometry and liquid chromatography atmospheric pressure chemical ionization mass spectrometry. AB - Gas chromatography-mass spectrometry (GC-MS) and liquid chromatography atmospheric pressure chemical ionization mass spectrometry (LC-APCI-MS) were optimized and applied for the trace-level determination of 42 priority pesticides and 33 priority organic pollutants from European Union Directive EC 76/464. First, off-line solid-phase extraction of 200 ml of river water using an OASIS solid-phase extraction cartridge, followed by GC-MS was used. Next, selected samples that were positive to GC-MS were analyzed by LC-APCI-MS in order to detect further polar byproducts or to improve the determination of previously detected polar analytes. The transformation products of triazine pesticides like deethylatrazine (DEA) and deisopropylatrazine (DIA) and compounds such as diuron and several chlorophenols were positively identified by LC-APCI-MS. The present methodology has also been used for searching for new analytes not included in the EC 76/464 list, like Irgarol, DEA and DIA. In addition it was applied to target pollutants in 43 river water samples from Portugal during a pilot survey from April to July 1999. Atrazine followed by simazine and 2,4,6-trichlorophenol were the most ubiquitous compounds detected in this area. The levels detected of the different compounds were in the range of: 0.01-2.73 microg/l, 0.05-0.74 microg/l, 0.02-1.65 microg/l, 0.02-5.43 microg/l, 0.01-0.40 microg/l, 0.01-0.26 microg/l, 0.02-0.61 microg/l, 0.01-3.90 microg/l, 0.01-1.24 microg/l, 0.02-2.3 microg/l, 0.01-0.13 microg/l and 0.01-0.5 microg/l for atrazine, simazine, terbuthylazine, alachlor, metolachlor, Irgarol, propanil; tributhylphosphate, diuron, 2,4,6 trichlorophenol, deisopropylatrazine and deethylatrazine, respectively. PMID- 10870693 TI - Part-per-trillion level determination of antifouling pesticides and their byproducts in seawater samples by off-line solid-phase extraction followed by high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry. AB - A new method for the simultaneous determination of antifouling pesticides and some of their byproducts such as dichlofluanid, diuron and its byproducts [demethyldiuron and 1-(3,4-dichlorophenyl)urea], (2-thiocyanomethylthio)ben: zothiazole, chlorothalonil, Sea-nine 211, Irgarol 1051 and one of its byproducts (2-methylthio-4-tert.-butylamino-s-triazine) in seawater was developed. The extraction of these compounds from the filtered seawater samples was performed off-line with different solid-phase extraction sorbents using (I) a 500 mg graphitized carbon black cartridge (ENVI-Carb) and (II) 200 mg polymeric cartridges (LiChrolut EN and Isolute ENV+) and passing 500 ml of the sample through these cartridges. The detection was carried out by reversed-phase high performance liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometry both in the negative and positive ion modes. The recovery ranged from 76 to 96% for the whole antifouling group with the ENVI-Carb cartridges and the detection limit was at the part-per-trillion level except for TCMTB. The method utilizing the polymeric cartridge proved to be very useful, time saving and with good recoveries when only Irgarol and its byproduct, Sea nine 211 and diuron and its byproducts, have to be analyzed. The different cartridges were applied to the analysis of these pesticides in different marinas of the Catalan coast; diuron, dichlofluanid, Sea-nine 211, Irgarol as well as demethyldiuron and the Irgarol byproduct being the must ubiquitous pollutants. Maximum concentration levels were 2-3.5 microg/l of diuron and Sea-nine 211, respectively. PMID- 10870694 TI - Comparison of various liquid chromatographic methods involving UV and atmospheric pressure chemical ionization mass spectrometric detection for the efficient trace analysis of phenylurea herbicides in various types of water samples. AB - The performance of mass spectrometric (MS) detection and UV detection in combination with reversed-phase liquid chromatography without and with the use of coupled column RPLC (LC-LC) has been compared for the trace analysis of phenylurea herbicides in environmental waters. The selected samples of this comparative study originated from an inter-laboratory study. For both detection modes, a 50 mm x 4.6 mm I.D. column and a 100 mm x 4.6 mm I.D. column packed with 3 microm C18 were used as the first (C-1) and second (C-2) column, respectively. Atmospheric pressure chemical ionization mass spectrometry was performed on a magnetic sector instrument. The LC-LC-MS analysis was carried out on-line by means of direct large volume (11.7 ml) injection (LVI). The performance of both on-line (LVI, 4 ml of sample) and off-line LC-LC-UV (244 nm) analysis was investigated. The latter procedure consisted of a solid-phase extraction (SPE) of 250 ml of water sample on a 500 mg C18 cartridge. The comparative study showed that LC-LC-MS is more selective then LC-LC-UV and, in most cases, more sensitive. The LVI-LC-LC-MS approach combines direct quantification and confirmation of most of the analytes down to a level of 0.01 microg/l in water samples in less then 30 min. As regards LC-LC-UV, the off-line method appeared to be a more viable approach in comparison with the on-line procedure. This method allows the screening of phenylurea's in various types of water samples down to a level of at least 0.05 microg/l. On-line analysis with LVI provided marginal sensitivity (limits of detection of about 0.1 microg/l) and selectivity was sometimes less in case of surface water samples. Both the on-line LVI-LC-LC-MS method and the LC-LC UV method using off-line SPE were validated by analysing a series of real-life reference samples. These samples were part of an inter-laboratory test and contained residues of herbicides ranging from 0.02 to 0.8 microg/l. Beside good correlation between the methods the data agreed very well with the true values of the samples. PMID- 10870695 TI - Application of liquid chromatography with mass spectrometry combined with photodiode array detection and tandem mass spectrometry for monitoring pesticides in surface waters. AB - Liquid chromatography with photodiode array detection (LC-DAD) and liquid chromatography with mass spectrometry (LC-MS) are two techniques that have been widely used in monitoring pesticides and their degradation products in the environment. However, the application of liquid chromatography with tandem mass spectrometry (LC-MS-MS) for such purposes, once considered too costly, is now gaining considerable ground. In this study, we compare these methods for the multi-residue analysis of pesticides in surface waters collected from the central and southeastern regions of France, and from the St. Lawrence River in Canada. Forty-eight pesticides belonging to eight different classes (triazine, amide, phenylurea, triazole, triazinone, benzimidazole, morpholine, phenoxyalkanoic), along with some of their degradation products, were monitored on a regular basis in the surface waters. For LC-MS, we used the electrospray ionization (ESI) interface in the negative ionization mode on acidic pesticides (phenoxyalkanoic, sulfonylurea), and the atmospheric pressure chemical ionization (APCI) interface in the positive ionization mode on the remaining chemicals. Different extraction techniques were employed, including liquid-liquid extraction with dichloromethane, and solid-phase extraction using C18-bonded silica and graphitized carbon black cartridges. Eleven of the target chemicals (desethylatrazine, desisopropylatrazine, atrazine, simazine, terbuthylazine, metolachlor, carbendazime, bentazone, penconazole, diuron and isoproturon) were detected by LC-MS at concentrations ranging from 20 to 900 ng/l in the surface waters from France, and six pesticides (atrazine, desethylatrazine, desisopropylatrazine, cyanazine, simazine and metolachlor) were detected by LC-MS and LC-MS-MS at concentrations ranging from 3 to 52 ng/l in the samples drawn from the St. Lawrence River. There was good correlation between the LC-DAD and LC MS techniques for 60 samples. The slope of the curves expressing the relationship between the results obtained with LC-DAD versus those obtained by LC-MS was near 1, with a correlation coefficient (r) of over 0.93. The identification potential of the LC-MS technique, however, was greater than that of the LC-DAD; its mass spectra, mainly reflecting the pseudomolecular ion resulting from a protonation or a deprotonation of the molecule, was rich in information. The LC-MS-MS technique with ion trap detectors, tested against the LC-MS on 10 surface water samples, gave results that correlated well with the LC-MS results, albeit generating mass spectra that yielded far more information about the structure of unknown substances. The sensitivity of the LC-MS-MS was equivalent to the selected ion monitoring (SIM) acquisition mode in LC-MS. The detection limits of the target pesticides ranged from 20 to 100 ng/l for the LC-MS technique (under full scan acquisition), and from 2 to 6 ng/l for LC-MS-MS. These limits were improved by a factor of almost 10 by increasing the sample volume to 10 l. PMID- 10870696 TI - Stability of organophosphorus insecticides on graphitized carbon black extraction cartridges used for large volumes of surface water. AB - The stability of nine organophosphorus insecticides (azinphos-ethyl, azinphos methyl, diazinon, EPN, ethion, fonofos, malathion, phosmet and parathion-methyl) was evaluated under a variety of storage conditions. Large volumes of surface water (4 l) were extracted using large-particle-size graphitized carbon black cartridges (Carbopack B 60-80 mesh). The effects of temperature, matrix type and drying of cartridges on the recovery of these contaminants, after different storage periods, were studied and compared to the conservation of surface water in bottles. After a 2-month period, all the chemicals stored on cartridges and kept at -20 degrees C remained stable, with recoveries ranging from 70 to 134%. By contrast, phosmet and EPN could no longer be recovered from the bottled surface water. Cartridges kept at -20 degrees C fared better than did those stored at 4 degrees C and 20 degrees C. The type of matrix water selected appears to have kept the target pesticides stored on cartridges from degrading, compared to the Milli-Q water, in which malathion and phosmet were unstable. The effect of the cartridges being either wet or dry made no difference in terms of improving the recovery of chemicals. After immediate surface water extraction, the most practical storage condition for the target insecticides was found to be storage on cartridges in the dark at -20 degrees C, with no drying or solvent washing of the Carbopack B material. PMID- 10870697 TI - Considerations involved with the use of semipermeable membrane devices for monitoring environmental contaminants. AB - Semipermeable membrane devices (SPMDs) are used with increasing frequency, and throughout the world as samplers of organic contaminants. The devices can be used to detect a variety of lipophilic chemicals in water, sediment/soil, and air. SPMDs are designed to sample nonpolar, hydrophobic chemicals. The maximum concentration factor achievable for a particular chemical is proportional to its octanol-water partition coefficient. Techniques used for cleanup of SPMD extracts for targeted analytes and for general screening by full-scan mass spectrometry do not differ greatly from techniques used for extracts of other matrices. However, SPMD extracts contain potential interferences that are specific to the membrane lipid matrix. Procedures have been developed or modified to alleviate these potential interferences. The SPMD approach has been demonstrated to be applicable to sequestering and analyzing a wide array of environmental contaminants including organochlorine pesticides, polychlorinated biphenyls, polycyclic aromatic hydrocarbons, polychlorinated dioxins and dibenzofurans, selected organophosphate pesticides and pyrethroid insecticides, and other nonpolar organic chemicals. We present herein an overview of effective procedural steps for analyzing exposed SPMDs for trace to ultra-trace levels of contaminants sequestered from environmental matrices. PMID- 10870698 TI - Determination of endocrine disruptors in water after derivatization with N-methyl N-(tert.-butyldimethyltrifluoroacetamide) using gas chromatography with mass spectrometric detection. AB - The combined gas chromatographic determination of a number of hydroxyl-group containing endocrine disruptors, including 4-octylphenol, 4-nonylphenol, 2,4 dichlorophenol, pentachlorophenol, 4-tert.-butylbenzoic acid, bisphenol-A, 17beta estradiol and 17alpha-ethynylestradiol, was investigated. Derivatization, required for sensitive determination of these compounds, was carried out using N methyl-N-(tert.-butyldimethyltrifluoroacetamide). A number of parameters affecting the derivatization reaction, like temperature, time, matrix, solvent, and amount of reagent were studied in detail. Quantitative yields were obtained for real-life extracts after optimization, but the hormones were only mono substituted. Both solid-phase extraction (SPE) and liquid-liquid extraction were studied as extraction methods, with emphasis on SPE material and effect of pH. Recoveries and RSD for analysis of surface water samples were 58-106 and 6-16% (n=4), respectively, when using SPE, and 109-117 and 6-14% (n=6) when using liquid-liquid extraction. The method developed allows routine analysis of surface water for traces of endocrine disruptors. The limits of detection of were 4-6 ng/l but higher for the hormones. PMID- 10870699 TI - Tissue motion and elasticity imaging. PMID- 10870700 TI - Elastography--the movement begins. AB - The advent of real-time ultrasound in the 1970s, together with a growing interest in tissue characterization, led to a number of investigators using the nature of tissue motion to distinguish healthy from diseased tissue. Our group at the (then) Ultrasonics Institute demonstrated the use of phase methods for detecting very small tissue motions, using natural stimuli. The method could also be applied in the lag (autocorrelation) domain to directly measure the amount of deformation to high accuracy. This method was also applied to measuring the amount of dilatation of blood vessels using both conventional and intravascular ultrasound. A basic limitation of these techniques was the poor spatial resolution, and quasistatic methods soon replaced this method of measuring tissue deformation. However, a new way of assessing the health of tissues had been established. PMID- 10870701 TI - New real-time strain imaging concepts using diagnostic ultrasound. AB - Two real-time strain imaging concepts and systems are presented. Both systems are based on a conventional ultrasound scanner that is connected to a PC with an A/D converter card for real-time data acquisition of rf data. Differential strain between successively acquired rf frames are estimated using phase root seeking. The first concept uses a special real-time implementation of manual elastography. In the second concept, denoted 'vibrography', the static compression is replaced by low-frequency axial vibration of the probe, still operating in quasistatic acquisition mode. The properties of both concepts are discussed with regard to noise and motion artefacts, and it is shown, using simulations and phantom experiments, that both imaging concepts yield the same kind of strain images. Vibrography has the advantage that no manual compression has to be applied, total compression can be very low and some motion artefacts are better suppressed. PMID- 10870702 TI - A method of imaging viscoelastic parameters with acoustic radiation force. AB - Acoustic radiation force has been proposed as a method of interrogating the mechanical properties of tissue. One simple approach applies a series of focused ultrasonic pulses to generate an acoustic radiation force, then processes the echoes returned from these pulses to estimate the radiation-force-induced displacement as a function of time. This process can be repeated at a number of locations to acquire data for image formation. In previous work we have formed images of tissue stiffness by depicting the maximum displacement induced at each tissue location after a finite period of insonification. While these maximum displacement images are able to differentiate materials of disparate mechanical properties, they exploit only a fraction of the information available. In this paper we show that the time-displacement curves acquired from tissue mimicking phantoms exhibit a viscoelastic response which is accurately described by the Voigt model. We describe how the viscous and elastic parameters of this model may be determined from experimental data. Finally, we show phantom images that depict not only the maximum local displacement, but also the viscous and elastic model parameters. These images offer complementary information about the target. PMID- 10870703 TI - Probing the dynamics of tissue at low frequencies with the radiation force of ultrasound. AB - Over the past few years there has been an increasing interest in using the radiation force of ultrasound for evaluating, characterizing and imaging biological tissues. Of particular interest are those methods that measure the dynamic properties of tissue at low frequencies. In this paper we present dynamic radiation force methods for probing tissue as a new field, discuss the interrelationship of several methods within this field and compare their features. The techniques in this field can be categorized into three groups: transient methods, shear-wave measurement methods and a recently developed method called vibro-acoustography. The last method is the focus of this paper. After briefly describing the key concepts of the first two methods, we will present a detailed description of vibro-acoustography. Finally, we will compare the capabilities and limitations of these methods. PMID- 10870704 TI - Characterization of plaque components and vulnerability with intravascular ultrasound elastography. AB - Intravascular ultrasound elastography is a method for measuring the local elastic properties using intravascular ultrasound (IVUS). The elastic properties of the different tissues within the atherosclerotic plaque are measured through the strain. Knowledge of these elastic properties is useful for guiding interventional procedures (balloon dilatation, ablation) and detection of the vulnerable plaque. In the last decade, several groups have applied elastography intravascularly with various levels of success. In this paper, the approaches of the different research groups will be discussed. The focus will be on our approach to the application of intravascular elastography. Elastograms were acquired in vitro and in vivo using the relative local displacements between IVUS images acquired at two levels of intravascular pressure with a 30 MHz mechanical or a 20 MHz array echo catheter. These displacements were estimated from the time shift between gated radiofrequency echo signals using cross-correlation algorithms with interpolation around the peak. Experiments on gel-based phantoms mimicking atherosclerotic vessels demonstrated the capability of elastography to identify soft and hard tissues independently of the echogenicity contrast. In vitro experiments on human arteries have demonstrated the potential of intravascular elastography to identify different plaque types based on their mechanical properties. These plaques could not be identified using the IVUS image alone. In vivo experiments revealed that reproducible elastograms could be obtained near end-diastole. Partial validation using the echogram was performed. Intravascular elastography provides information that is frequently unavailable or inconclusive from the IVUS image and which may therefore assist in the diagnosis and treatment of atherosclerotic disease. PMID- 10870705 TI - Three-dimensional sonoelastography: principles and practices. AB - Sonoelastography is an ultrasound imaging technique where low amplitude, low frequency shear waves (less than 0.1 mm displacement and less than 1 kHz frequency) are propagated through internal organs, while real-time Doppler techniques are used to image the resulting vibration pattern. When a discrete hard inhomogeneity, such as a tumour, is present within a region of soft tissue, a decrease in the vibration amplitude will occur at its location. This forms the basis for tumour detection using sonoelastography. For three-dimensional (3D) imaging the acquisition of sequential tomographic slices using this technique, combined with image segmentation, enables the reconstruction, quantification and visualization of tumour volumes. Sonoelastography and magnetic resonance images (MRI) of a tissue phantom containing a hard isoechoic inclusion are compared to evaluate the accuracy of this method. The tumour delineation from sonoelastography was found to have good agreement with the tumour from MRI except for a bleeding at one of its ends. Although sonoelastography is still in an experimental phase, the principles behind this imaging modality are explained and some practical aspects of acquiring sonoelastography images are described. Results from a 3D sonoelastography reconstruction of a tissue mimicking phantom and an ex vivo whole prostate specimen are presented. PMID- 10870706 TI - Non-invasive quantitative reconstruction of tissue elasticity using an iterative forward approach. AB - A novel iterative approach is presented to estimate Young's modulus in homogeneous soft tissues using vibration sonoelastography. A low-frequency (below 100 Hz) external vibration is applied and three or more consecutive frames of B scan image data are recorded. The internal vibrational motion of the soft tissue structures is calculated from 2D displacements between pairs of consecutive frames, which are estimated using a mesh-based speckle tracking method. An iterative forward finite element approach has been developed to reconstruct Young's modulus from the measured vibrational motion. This is accomplished by subdividing the 2D image domain into sample blocks in which Young's modulus is assumed to be constant. Because the finite element equations are internally consistent, boundary values other than displacement are not required. The sensitivity of the results to Poisson's ratio and the damping coefficient (viscosity) is investigated. The approach is verified using simulated displacement data and using data from tissue-mimicking phantoms. PMID- 10870707 TI - An effective ultrasonic strain measurement-based shear modulus reconstruction technique for superficial tissues--demonstration on in vitro pork ribs and in vivo human breast tissues. AB - An effective shear modulus reconstruction technique is described which uses ultrasonic strain measurements for diagnosis of superficial tissues, i.e. our previously developed ultrasonic strain measurement and shear modulus reconstruction methods are combined and enhanced. The technique realizes very low computational load, yet yields fairly high quantitativeness, high stability and spatial resolution, and large dynamic range. The suitability of the method is demonstrated on in vitro pork ribs and in vivo human breast tissues (fibroadenoma and scirrhous carcinoma). PMID- 10870708 TI - Evaluation of an iterative reconstruction method for quantitative elastography. AB - This paper describes an inverse reconstruction technique based on a modified Newton Raphson iterative scheme and the finite element method, which has been developed for computing the spatial distribution of Young's modulus from within soft tissues. Computer simulations were conducted to determine the relative merits of reconstructing tissue elasticity using knowledge of (a) known displacement boundary conditions (DBC), and (b) known stress boundary conditions (SBC). The results demonstrated that computing Young's modulus using knowledge of SBC allows accurate quantification of Young's modulus. However, the quality of the images produced using this reconstruction approach was dependent on the Young's modulus distribution assumed at the start of the reconstruction procedure. Computing Young's modulus from known DBC provided relative estimates of tissue elasticity which, despite the disadvantage of not being able to accurately quantify Young's modulus, formed images that were generally superior in quality to those produced using the known SBC, and were not affected by the trial solution. The results of preliminary experiments on phantoms demonstrated that this reconstruction technique is capable in practice of improving the fidelity of tissue elasticity images, reducing the artefacts otherwise present in strain images, and recovering Young's modulus images that possess excellent spatial and contrast resolution. PMID- 10870709 TI - A novel interpolation strategy for estimating subsample speckle motion. AB - Multidimensional, high-resolution ultrasonic imaging of rapidly moving tissue is primarily limited by sparse sampling in the lateral dimension. In order to achieve acceptable spatial resolution and velocity quantization, interpolation of laterally sampled data is necessary. We present a novel method for estimating lateral subsample speckle motion and compare it with traditional interpolation methods. This method, called grid slopes, requires no a priori knowledge and can be applied to data with as few as two samples in the lateral dimension. Computer simulations were performed to compare grid slopes with two conventional interpolation schemes, parabolic fit and cubic spline. Results of computer simulations show that parabolic fit and cubic spline performed poorly at translations greater than 0.5 samples, and translations less than 0.5 samples were subject to an estimation bias. Grid slopes accurately estimated translations between 0 and 1 samples without estimation bias at high signal-to-noise ratios. Given that the grid slopes interpolation technique performs well at high signal to-noise ratios, one pertinent clinical application might be tissue motion tracking. PMID- 10870710 TI - Precision estimation and imaging of normal and shear components of the 3D strain tensor in elastography. AB - In elastography we have previously developed a tracking and correction method that estimates the axial and lateral strain components along and perpendicular to the compressor/scanning axis following an externally applied compression. However, the resulting motion is a three-dimensional problem. Therefore, in order to fully describe this motion we need to consider a 3D model and estimate all three principal strain components, i.e. axial, lateral and elevational (out-of plane), for a full 3D tensor description. Since motion is coupled in all three dimensions, the three motion components have to be decoupled prior to their estimation. In this paper, we describe a method that estimates and corrects motion in three dimensions, which is an extension of the 2D motion tracking and correction method discussed before. In a similar way as in the 2D motion estimation, and by assuming that ultrasonic frames are available in more than one parallel elevational plane, we used methods of interpolation and cross correlation between elevationally displaced RF echo segments to estimate the elevational displacement and strain. In addition, the axial, lateral and elevational displacements were used to estimate all three shear strain components that, together with the normal strain estimates, fully describe the full 3D normal strain tensor resulting from the uniform compression. Results of this method from three-dimensional finite-element simulations are shown. PMID- 10870711 TI - Dynamics of errors in 3D motion estimation and implications for strain-tensor imaging in acoustic elastography. AB - For the purpose of quantifying the noise in acoustic elastography, a displacement covariance matrix is derived analytically for the cross-correlation based 3D motion estimator. Static deformation induced in tissue from an external mechanical source is represented by a second-order strain tensor. A generalized 3D model is introduced for the ultrasonic echo signals. The components of the covariance matrix are related to the variances of the displacement errors and the errors made in estimating the elements of the strain tensor. The results are combined to investigate the dependences of these errors on the experimental and signal-processing parameters as well as to determine the effects of one strain component on the estimation of the other. The expressions are evaluated for special cases of axial strain estimation in the presence of axial, axial-shear and lateral-shear type deformations in 2D. The signals are shown to decorrelate with any of these deformations, with strengths depending on the reorganization and interaction of tissue scatterers with the ultrasonic point spread function following the deformation. Conditions that favour the improvements in motion estimation performance are discussed, and advantages gained by signal companding and pulse compression are illustrated. PMID- 10870712 TI - Tissue characterization using magnetic resonance elastography: preliminary results. AB - The well-documented effectiveness of palpation as a diagnostic technique for detecting cancer and other diseases has provided motivation for developing imaging techniques for noninvasively evaluating the mechanical properties of tissue. A recently described approach for elasticity imaging, using propagating acoustic shear waves and phase-contrast MRI, has been called magnetic resonance elastography (MRE). The purpose of this work was to conduct preliminary studies to define methods for using MRE as a tool for addressing the paucity of quantitative tissue mechanical property data in the literature. Fresh animal liver and kidney tissue specimens were evaluated with MRE at multiple shear wave frequencies. The influence of specimen temperature and orientation on measurements of stiffness was studied in skeletal muscle. The results demonstrated that all of the materials tested (liver, kidney, muscle and tissue simulating gel) exhibit systematic dependence of shear stiffness on shear rate. These data are consistent with a viscoelastic model of tissue mechanical properties, allowing calculation of two independent tissue properties from multiple-frequency MRE data: shear modulus and shear viscosity. The shear stiffness of tissue can be substantially affected by specimen temperature. The results also demonstrated evidence of shear anisotropy in skeletal muscle but not liver tissue. The measured shear stiffness in skeletal muscle was found to depend on both the direction of propagation and polarization of the shear waves. PMID- 10870713 TI - Visualization and quantification of breast cancer biomechanical properties with magnetic resonance elastography. AB - A quasistatic magnetic resonance elastography (MRE) method for the evaluation of breast cancer is proposed. Using a phase contrast, stimulated echo MRI approach, strain imaging in phantoms and volunteers is presented. First-order assessment of tissue biomechanical properties based on inverse strain mapping is outlined and demonstrated. The accuracy of inverse strain imaging is studied through simulations in a two-dimensional model and in an anthropomorphic, three dimensional finite-element model of the breast. To improve the accuracy of modulus assessment by elastography, inverse methods are discussed as an extension to strain imaging, and simulations quantify MRE in terms of displacement signal/noise required for robust inversion. A direct inversion strategy providing information on tissue modulus and pressure distribution is described along with a novel iterative method utilizing a priori knowledge of tissue geometry. It is shown that through the judicious choice of information from previous contrast enhanced MRI breast images, MRE data acquisition requirements can be significantly reduced while maintaining robust modulus reconstruction in the presence of strain noise. An experimental apparatus for clinical breast MRE and preliminary images of a normal volunteer are presented. PMID- 10870714 TI - Left ventricular motion reconstruction from planar tagged MR images: a comparison. AB - Through recent development of MR techniques, it is now possible to assess regional myocardial wall function in a non-invasive way. Using MR tagging, space is marked with planes which deform with the tissue, providing markers for tracking the local motion of the myocardium. Numerous methods to reconstruct the three-dimensional displacement field have been developed. The aim of this article is to provide a framework to quantitatively compare the performance of four methods the authors have developed. Five sets of experiments are described, and their results are reported. Instructions are also provided to perform similar tests on any method using the same data. The experiments show that some characteristic properties of the methods, such as sensitivity to noise or spatial resolution, can be quantitatively classified. Cross-comparison of performances show what range values for these properties can be considered acceptable. PMID- 10870715 TI - Three-dimensional static displacement, stimulated echo NMR elasticity imaging. AB - This article presents a method for measuring three-dimensional mechanical displacement and strain fields using stimulated echo MRI. Additional gradient pulses encode internal displacements in response to an externally applied deformation. By limiting the mechanical transition to the stimulated echo mixing time, a more accurate static displacement measurement is obtained. A three dimensional elasticity reconstruction within a region of interest having a uniform shear modulus along its boundary is performed by numerically solving discretized elasticity equilibrium equations. Data acquisition, strain measurements and reconstruction were performed using a silicone gel phantom containing an inclusion of known elastic properties. A comparison between two dimensional and three-dimensional reconstructions from simulated and experimental displacement data shows higher accuracy from the three-dimensional reconstruction. The long-term objective of this work is to provide a method for remotely palpating and elastically quantitating manually inaccessible tissues. PMID- 10870716 TI - High-resolution tensor MR elastography for breast tumour detection. AB - MR elastography is a novel imaging technique for the visualization of elastic properties of tissue. It is expected that this method will have diagnostic value for the clarification of suspicious breast lesions. Low-frequency mechanical waves are coupled into the tissue and visualized via an MR sequence which is phase-locked to the mechanical excitation. Commonly, elasticity is assumed to be isotropic and reconstruction is performed in only two dimensions. The technique is extended to three dimensions such that the entire symmetric elasticity tensor is assessed. This is achieved by measuring different phases of the mechanical wave during one oscillatory cycle. Thereby it is possible to provide information about the anisotropy of the elasticity tensor. Finite-element simulations as well as phantom experiments are performed to demonstrate the feasibility of the method. Initial clinical results of a breast carcinoma are presented. The analysis of the eigenvalues of the elasticity tensor support the hypothesis that breast carcinoma might exhibit an anisotropic elasticity distribution. The surrounding benign tissue appears isotropic. Thereby new and additional diagnostic information is provided which might help in distinguishing between benign and malignant breast diseases. PMID- 10870717 TI - Visualizing myocardial function using HARP MRI. AB - Harmonic phase magnetic resonance imaging (HARP) is a new technique for measuring the motion of the left ventricle of the heart. HARP uses magnetic resonance tagging, Fourier filtering and special processing algorithms to calculate key indices of myocardial motion including Eulerian and Lagrangian strain. This paper presents several new methods for visualizing myocardial motion based on HARP. Quantities that are computed and visualized include motion grids, velocity fields, strain rates, pathlines, tracked Eulerian strain, and contraction angle. The computations are fast and fully automated and have the potential for clinical application. PMID- 10870718 TI - Four-dimensional B-spline based motion analysis of tagged MR images: introduction and in vivo validation. AB - In MRI tagging, magnetic tags-spatially encoded magnetic saturation planes-are created within tissues acting as temporary markers. Their deformation pattern provides useful qualitative and quantitative information about the functional properties of underlying tissue and allows non-invasive analysis of mechanical function. The measured displacement at a given tag point contains only unidirectional information; in order to track the full 3D motion, these data have to be combined with information from other orthogonal tag sets over all time frames. Here, we provide a method to describe the motion of the heart using a four-dimensional tensor product of B-splines. In vivo validation of this tracking algorithm is performed using different tagging sets on the same heart. Using the validation results, the appropriate control point density was determined for normal cardiac motion tracking. Since our motion fields are parametric and based on an image plane based Cartesian coordinate system, trajectories or other derived values (velocity, acceleration, strains ...) can be calculated for any desired point within the volume spanned by the control points. This method does not rely on specific chamber geometry, so the motion of any tagged structure can be tracked. Examples of displacement and strain analysis for both ventricles are also presented. PMID- 10870719 TI - Estimation of aortic compliance using magnetic resonance pulse wave velocity measurement. AB - A method for compliance estimation employing magnetic resonance pulse wave velocity measurement is presented. Time-resolved flow waves are recorded at several positions along the vessel using a phase contrast sequence, and pulse wave velocity is calculated from the delay of the wave onsets. Using retrospective cardiac gating in combination with an optically decoupled electrocardiogram acquisition, a high temporal resolution of 3 ms can be achieved. A phantom set-up for the simulation of pulsatile flow in a compliant vessel is described. In the phantom, relative errors of pulse wave velocity estimation were found to be about 15%, whereas in a volunteer, larger errors were found that might be caused by vessel branches. Results of pulse wave velocity estimation agree with direct aortic distension measurements which rely on a peripheral estimate of aortic pressure and are therefore less accurate. Studies in 12 volunteers show values of pulse wave velocity consistent with the literature; in particular the well-known increase in pulse wave velocity with age was observed. Preliminary results show that the method can be applied to aortic aneurysms. PMID- 10870720 TI - "Everyone has won and all must have prizes". PMID- 10870721 TI - Plasma norepinephrine levels, vagal tone index, and flexor reflex threshold in premature neonates receiving intravenous morphine during the postoperative period: a pilot study. AB - OBJECTIVE: The goal of this study was to evaluate the effects of a single dose of intravenous morphine on postoperative pain in extremely premature neonates after thoracotomy. DESIGN: Descriptive correlational study. PATIENTS: Twenty-four critically ill mechanically ventilated premature neonates with a mean gestational age of 26.1 +/- 2.1 (SD) weeks and a postnatal age of 13.8 +/- 8.1 (SD) days. OUTCOME MEASURES: Plasma norepinephrine (NE) levels, vagal tone index (VTI), and flexor reflex threshold were measured preoperatively, immediately before, and 20 and 60 minutes after the administration of the first postoperative dose of morphine (0.1 mg/kg). RESULTS: One-way repeated-measures ANOVA revealed no significant change in plasma NE levels from baseline levels (df[2,32] = 2.40, p = 0.11). Pre- and postmorphine VTI values were significantly lower than preoperative values (df[3,60] = 6.04, p = 0.0012), but no significant differences were found between pre- and postmorphine VTI values. Neonates (n = 10) who had a flexor reflex response during the postoperative period demonstrated no significant differences in the force required to elicit a flexor reflex across the four measurements (df[3,27] = 0.76, p = 0.53); however, the flexor reflex responses were significantly less vigorous during the postoperative period than at baseline. CONCLUSIONS: Findings from this pilot study suggest that the dose of morphine commonly used to treat postoperative pain in premature neonates does not affect NE, VTI, and flexor reflex threshold values within 1 hour of administration. PMID- 10870722 TI - Pain relief after arthroscopic knee surgery: intravenous morphine, epidural morphine, and intra-articular morphine. AB - OBJECTIVE: The aim of this study was to compare the analgesic efficacy and side effects of intravenous (IV), epidural, and intra-articular (IA) morphine after arthroscopic knee surgery. DESIGN: Prospective, randomized, double-blind clinical investigation. SETTING: Medical center, university teaching hospital. PATIENTS: Inpatients with an American Society of Anesthesiologists physical status of I or II who were scheduled for elective arthroscopic knee surgery. INTERVENTIONS AND OUTCOME MEASURES: A total of 75 patients scheduled for arthroscopic knee surgery under epidural anesthesia were randomly divided into three groups (n = 25 in each group). At the end of surgery, patients in group 1 received 3 mg of IV morphine, patients in group 2 received 3 mg of epidural morphine, and patients in group 3 received 3 mg of IA morphine. Patients were then observed for 24 hours. During the observation period, the proportion of patients requiring rescue analgesia with intramuscular diclofenac in each group was calculated and the occurrence of morphine-related side effects was recorded. RESULTS: We found that patients who received IV morphine requested more rescue analgesia than those who received either epidural or IA morphine. The proportions of patients requiring rescue analgesia in the IV, epidural, and IA groups were 65%, 13%, and 9%, respectively (p < 0.01 in group 1 vs. group 2 and in group 1 vs. group 3). Epidural morphine was associated with higher incidences of nausea and vomiting, pruritus, and urinary retention than IA morphine (range, p < 0.05-0.01 in group 2 vs. group 3). CONCLUSIONS: Patients who received IA morphine consumed less rescue analgesia than those who received IV morphine. They also reported fewer side effects than those patients who received epidural morphine. Intra-articular morphine may be the method of choice for pain relief after arthroscopic knee surgery. PMID- 10870723 TI - Negative affect, self-report of depressive symptoms, and clinical depression: relation to the experience of chronic pain. AB - OBJECTIVE: The goal of this study was to examine the relative importance of global affective distress, self-report of depressive symptoms, and presence or absence of major depression to the experience of chronic pain. SETTING: A multidisciplinary pain program at a university medical center was the setting for this study. PATIENTS: Subjects in this study were 211 consecutive patients with chronic pain. OUTCOME MEASURES: Pain duration, compensation, and litigation status were controlled for in the statistical analyses, as each correlated significantly with at least one of the measures of affect. Global affective distress was assessed using the Global Severity Index (GSI) from the Brief Symptom Inventory. The Beck Depression Inventory and the Center for Epidemiological Studies Depression Scale were used as measures of self-report of depressive symptoms. Presence or absence of major depression was based on DSM-IV criteria. RESULTS AND CONCLUSIONS: The GSI, Beck Depression Inventory, and Center for Epidemiological Studies Depression Scale were significantly correlated with each measure of the experience of pain, although clinical depression was only significantly related to self-reported disability and negative thoughts about pain. The self-report measures of depression maintained their relation to the dependent measures when the somatic items from the scales were removed, suggesting that the relations were not spuriously due to the influence of pain symptoms on the scales. When examining the unique contribution of each variable to the experience of pain (by simultaneously controlling for the other measures of affect), the GSI was uniquely related to the sensory and affective components of pain. Self-report of depressive symptoms was more highly related to a measure of the evaluative component of pain and uniquely related to self-reported disability and negative thoughts about pain. The results are discussed within the context of theoretical models of the relation between pain and affect, and suggestions for future research are presented. PMID- 10870724 TI - Clinical pain perception and hormone replacement therapy in postmenopausal women experiencing orofacial pain. AB - OBJECTIVE: The purpose of this study was to examine the magnitude of the relation between a postmenopausal woman's hormonal replacement status and clinical pain report in a sample of women experiencing orofacial pain. DESIGN: To accomplish this, pain ratings were collected during a routine chronic pain evaluation at an orofacial pain clinic from a sample of 87 postmenopausal women. RESULTS: Results of ANCOVA (controlling for pain duration) demonstrated that postmenopausal women receiving hormone replacement therapy (HRT) reported higher levels of pain than postmenopausal women not taking HRT. Numeric pain rating scales revealed large effect sizes for worst pain report (0.62), moderate differences for average (0.48) and current (0.39) pain levels, and trivial differences for least pain (0.04). Effect sizes for the McGill Pain Questionnaire indicated somewhat smaller differences (0.35-0.24) between the two groups. CONCLUSIONS: This study is among the first to examine the relation between a woman's hormonal status and clinical pain perception and is the first to investigate the role of HRT in a postmenopausal woman's orofacial pain report in a clinical treatment setting. This area of inquiry is particularly salient given the high percentage of women who choose to initiate HRT either after hysterectomy or with the onset of menopause. PMID- 10870725 TI - Prevalence and impact of posttraumatic stress disorder-like symptoms on patients with fibromyalgia syndrome. AB - OBJECTIVE: Traumatic events can result in a set of symptoms including nightmares, recurrent and intrusive recollections, avoidance of thoughts or activities associated with the traumatic event, and symptoms of increased arousal such as insomnia and hypervigilance. These posttraumatic stress disorder (PTSD)-like symptoms are frequently observed in persons with chronic pain syndromes. Little is known about how these two phenomena interact with one another. The present study evaluated PTSD-like symptoms in patients with fibromyalgia syndrome (FMS) and examined the relation between PTSD-like symptoms and problems associated with FMS. DESIGN: Ninety-three consecutive patients underwent a comprehensive FMS evaluation and completed self-report questionnaires measuring PTSD-like symptoms, disability, and psychosocial responses to their pain condition. Subjects were divided in two groups based on level of self-reported PTSD-like symptoms. RESULTS: Approximately 56% of the sample reported clinically significant levels of PTSD-like symptoms (PTSD+). The PTSD+ patients reported significantly greater levels of pain (p < 0.01), emotional distress (p < 0.01), life interference (p < 0.01), and disability (p < 0.01) than did the patients without clinically significant levels of PTSD-like symptoms (PTSD-). Over 85% of the PTSD+ patients compared with 50% of the PTSD- patients demonstrated significant disability. Based on response to the Multidimensional Pain Inventory, a significantly smaller percentage of PTSD+ patients were classified as adaptive copers (15%) compared with the PTSD- group (48.2%). CONCLUSIONS: Results suggest that PTSD-like symptoms are prevalent in FMS patients and may influence adaptation to this chronic illness. Clinicians should assess the presence of these symptoms, as the failure to attend to them in treatment may impede successful outcomes. PMID- 10870726 TI - Painful pricks and prickle pains: is there a relation between children's ratings of venipuncture pain and parental assessments of usual reaction to other pains? AB - OBJECTIVE: The objective of this study was to examine whether parental assessment of a child's usual behavioral reaction to common painful events predicts the child's ratings of needle pain intensity from routine venipuncture. DESIGN: Children aged 3 to 12 years (n = 88) used the Faces Pain Scale to rate how much venipuncture hurt and also indicated whether the pain was more, less, or the same as expected. The child's parent (mother) used the same scale to predict how much the needle would hurt the child as well as to rate the child's pain as observed at the time of venipuncture. Parents also estimated their child's usual reaction to six common painful events. An independent observer used a behavioral checklist to rate the child's pain response at the time of venipuncture as well as to assign a global pain rating on the Faces Pain Scale. OUTCOME MEASURES: The Faces Pain Scale and a behavioral checklist (scoring facial, vocal, motor, and verbal reactions) were used in this study. RESULTS AND CONCLUSIONS: Those children who reported venipuncture as hurting more than expected also gave the highest mean needle pain ratings and tended to have their pain underpredicted by their parents before venipuncture. For these children, parental estimates of reactions to other painful events proved to be a useful predictor of self-reported needle pain. Parent and child ratings of pain agreed more closely for those parents who indicated having relied on what their child "did" rather than "said." Additionally, and consistent with previous studies, independent observation of children's facial responses was the most useful indicator of needle pain severity. Preparation of children for venipuncture may be enhanced by asking a parent beforehand how the child usually responds to everyday pain. Specifically, reaction to other sharp time-limited pains (e.g., finger pinch, stepping on a prickle) may provide a useful guide to identifying which children will report experiencing greater pain than expected from venipuncture. PMID- 10870728 TI - Pressure pain thresholds and electromyographically defined muscular fatigue induced by a muscular endurance test in normal women. AB - OBJECTIVE: This study was undertaken to examine the relation between muscular tenderness measured as pressure pain thresholds (PPTs) and electromyographic (EMG) signs of fatigue before and after a local standardized static muscle contraction. DESIGN: Pressure pain thresholds were measured in the shoulder region before, immediately after, and 10 minutes after a standardized static endurance test while monitoring the EMG signs of local muscular fatigue and its recovery. The study did not address local biochemical issues. SETTING: The study was conducted at the Department of Rehabilitation, Lund University Hospital, Lund, Sweden. SUBJECTS: Twenty-five healthy female volunteers without musculoskeletal problems participated in this study. INTERVENTION: A static endurance test was performed, which consisted of a submaximal unilateral activation of the right trapezius and deltoid muscles for as long as possible. OUTCOME MEASURES: Bilateral PPTs over the trapezius and deltoid muscles were measured with an electronic pressure algometer. Established surface EMG parameters of local muscular fatigue were assessed. The Borg Rating of Perceived Exertion scale was used. RESULTS: The average endurance time was 330 seconds. Immediately after the test, significant bilateral increases in the normalized PPTs over both muscles were found, although the increase was more pronounced on the test side: over the right trapezius muscle by 13% (p <0.001), over the right deltoid muscle by 23% (p <0.001), and over the left trapezius and deltoid muscles by 6% (p = 0.04) and (p = 0.009), respectively. These increases persisted 10 minutes after the end of the test. The subjects developed significant signs of fatigue as defined by EMG criteria in both muscles on the right side during the test. The recovery from fatigue was approximately half complete 15 seconds after the end of the test and complete or almost complete 10 minutes thereafter. CONCLUSIONS: Pressure pain thresholds over shoulder muscles remained elevated up to 10 minutes after a unilateral static endurance test. This time course was completely different from that of EMG-defined muscle fatigue, which showed a fast recovery. These findings indicate that the mechanisms of recovery from fatigue and nociception are independent of each other. The bilateral PPT increases might be explained by central antinociceptive mechanisms activated by static muscle work. PMID- 10870727 TI - Automatic and strategic processing of threat cues in patients with chronic pain: a modified stroop evaluation. AB - OBJECTIVE: The goal of this study was to clarify whether patients with chronic pain selectively attend to syndrome-specific (i.e., pain-related) information and, if so, to determine whether this occurs at the conscious (i.e., strategic) or unconscious (i.e., automatic) level. SETTING: This study was conducted at a tertiary care rehabilitation center. PATIENTS: Thirty-three patients with chronic back and/or neck pain and 33 healthy volunteers matched for age, sex, and education participated in this study. OUTCOME MEASURES: A computerized version of a modified Stroop color-naming task, with unmasked and masked conditions, was used to assess strategic and automatic information processing of words related to sensory pain, affect pain, physical threat, social threat, and neutral themes. RESULTS: A repeated-measures ANOVA indicated that patients with chronic pain but not healthy volunteers had delayed color-naming latencies to both sensory and affect pain words in the unmasked condition. Color-naming latency differences were not evident for other word types in the unmasked condition or for any word types in the masked condition. Correlational and regression analyses indicated that the delayed color-naming latencies to pain words in the unmasked condition observed for the chronic pain patients were, in part, associated with high pain specific cognitive anxiety and interference and lower levels of anxiety sensitivity. CONCLUSIONS: Individuals with chronic pain selectively process pain related cues at the strategic level but not at the automatic level. Implications of the findings and future research directions are discussed. PMID- 10870729 TI - Radiofrequency lesions of the stellate ganglion in chronic pain syndromes: retrospective analysis of clinical efficacy in 86 patients. AB - OBJECTIVE: Stellate ganglion (SG) blockade is used for the treatment of chronic pain syndromes in which the sympathetic nervous system is hypothesized to be involved. A possible treatment modality to achieve long-term pain reduction is blockade of the SG by means of a radiofrequency lesion (RF-SG). To evaluate the outcome of RF-SG as a therapy for different chronic pain syndromes, we reviewed 86 RF-SG procedures. DESIGN: Medical records containing treatment information were reviewed systematically. A systematic MEDLINE literature review search on SG blockade was also performed. RESULTS: In our clinic, 39.5% of 221 patients who received a prognostic SG block subsequently underwent RF-SG. Of these patients, 40.7% noted a more than 50% reduction of pain, 54.7% reported no effect on pain, and 4.7% showed worsening of pain. The mean follow-up interval was 52 weeks. The computer-assisted literature search resulted in 31 studies: 12 about complications and 19 about the efficacy of SG blockade. A review of these studies showed partial pain relief in 41.3% of patients, complete pain relief in 37.8%, and no pain relief in 20.9%. CONCLUSIONS: The efficacy of RF-SG blockade seems to be in line with that of other SG blockade procedures reported in the literature. Our retrospective study shows that an RF-SG block is most likely to be of benefit for patients suffering from complex regional pain syndrome type 2, ischemic pain, cervicobrachialgia, or postthoracotomy pain. Clinical efficacy remains to be proven in a randomized controlled trial, however. PMID- 10870730 TI - Perceived treatment helpfulness and cost in chronic pain rehabilitation. AB - OBJECTIVE: This article examines the perceived helpfulness of treatment components in comprehensive interdisciplinary pain management programs as they relate to cost. DESIGN: Patient satisfaction results assessed by the Treatment Helpfulness Questionnaire (THQ) and treatment costs were compared for 309 subjects at three comprehensive interdisciplinary chronic pain management centers. All subjects completed the THQ immediately after treatment, and follow up data were gathered 3 to 6 months after the end of treatment at two of the three centers. RESULTS: Ratings of treatment helpfulness were not found to be related to either demographic or medical variables. Mean THQ ratings for many treatment modalities did differ significantly between centers, but subjects at all centers generally gave higher THQ ratings to psychological and educational therapies than to physical therapy and medical modalities both at posttreatment and at follow-up evaluations. More costly treatments generally did not receive higher ratings than less costly ones. THQ ratings tended to decline modestly from posttreatment to follow-up evaluations. CONCLUSIONS: For the selected population of patients undergoing comprehensive interdisciplinary pain management, educational and psychological approaches received high ratings of helpfulness at a relatively low cost. Further research is needed to address whether comparative patient satisfaction data can be used at pain centers to produce improved outcomes at reduced costs. PMID- 10870731 TI - Allergic reaction to spinal cord stimulator. AB - OBJECTIVE: The objective was to report on the possibility of allergic reaction to the components of a spinal cord stimulator. DESIGN: We describe a severe allergic reaction after the insertion of a spinal cord stimulator in a patient with complex regional pain syndrome type 1. SETTING: The patient was being followed in an office-based pain management practice. PATIENT: The patient is a 41-year-old woman with complex regional pain syndrome type 1, posttrauma. INTERVENTION: Insertion of a cervical and lumbar spinal cord stimulator. OUTCOME MEASURES: The outcome measures were a numerical scale of pain intensity and the ability to perform the activities of daily living. RESULTS: Adequate pain control complicated by allergic reaction. CONCLUSIONS: There exists a possibility that a patient may experience an allergic reaction to spinal cord stimulator components. Recognition of such contact sensitivity is important for physicians implanting such devices. Patients may be misdiagnosed as having infections, which can delay appropriate management; definitive diagnosis can be confirmed with a patch test. Treatment consists of removal of such devices. PMID- 10870732 TI - Fibromyalgia and related matters. PMID- 10870733 TI - Pain and suffering. AB - It is suffering, not pain, that brings patients into doctor's offices in hopes of finding relief. Astounding developments in our understanding of the mechanisms of nociception should not cause us to lose sight of our patients' goals. Chronic pain is far more than a sensory process. We must maintain the biopsychosocial model of chronic pain if we are to provide effective health care to our patients. Understanding the components of pain facilitates this goal. Suffering is an emergent property of the human brain and is dependent upon consciousness. It too is worthy of study by scientists and of concern to clinicians. PMID- 10870734 TI - Neurotrophic factors and pain. AB - Neurotrophic factors have been shown to play significant roles in the transmission of physiologic and pathologic pain. Nerve growth factor appears to be particularly important. It is crucial for the development of sympathetic and small fiber sensory neurons that serve as nociceptors. It stimulates the expression and release of neuropeptides involved in pain transmission, and interacts with cellular and molecular mediators of inflammation. Blockade with nerve growth factor antiserum demonstrates the critical role of the growth factor in mediating inflammatory hyperalgesia. Administration of nerve growth factor to rodents results in the rapid onset of hyperalgesia. Although the exact mechanism is unknown, several possibilities have been proposed. In clinical trials for the treatment of Alzheimer disease and peripheral neuropathy, induction of pain has been the major adverse event. When administered intracebrebroventricularly, a dull constant back pain resulted. Subcutaneous injection of nerve growth factor induces injection site hyperalgesia, as well as generalized myalgias and arthralgias. Whether the mechanisms underlying these adverse events are identical to those associated with the hyperalgesia in rodents is unknown. In addition to nerve growth factor, other growth factors, such as brain-derived neurotrophic factor and glial cell-derived neurotrophic factor, may be involved in pain pathways. Their precise roles are still being defined, but evidence suggests that they may have particular relevance to neuropathic pain. Understanding the role all these factors play may change the way we approach the treatment of pain in general, and neuropathic pain in particular. PMID- 10870735 TI - Peripheral neuropathic pain: from mechanisms to symptoms. AB - Several independent pathophysiological mechanisms in both the peripheral and central nervous system are responsible for sensory symptoms as well as spontaneous and evoked pains in peripheral neuropathies: (1) Pathologic active or sensitized nociceptors can induce secondary changes in central processing, leading to spinal cord hyperexcitability that causes input from mechanoreceptive Abeta-fibers (light touching) to be perceived as pain. These patients characteristically have spontaneous pain, heat hyperalgesia, static mechanical allodynia, and and severe dynamic mechanical allodynia. (2) Nociceptor function may be selectively impaired within the allodynic skin. Pain and temperature sensation are profoundly impaired but light moving mechanical stimuli can often produce severe pain (dynamic mechanical allodynia). Anatomic reorganization in the dorsal horn resulting from C-fiber degeneration may lead to Abeta-fiber mediated allodynia. (3) Persistent inflammatory reactions of the nerve trunk can induce ectopic activity in primary afferent nociceptors and thus is a potential cause of spontaneous pain and allodynia. This effect is mediated by the cytokine tumor necrosis factor-alpha produced by activated macrophages. (4) After nerve lesion the sympathetic nervous system might interact with afferent neurons. Activity in sympathetic fibers can induce further activity in sensitized nociceptors and, therefore, enhance pain and allodynia (sympathetically maintained pain). This pathologic interaction acts via noradrenaline released from sympathetic terminals and newly expressed receptors on the afferent neuron membrane. These mechanisms can operate in concert in a single disease entity (e.g., postherpetic neuralgia) and also in a single patient. Distinct pathophysiological mechanisms lead to specific sensory symptoms (e.g., dynamic mechanical allodynia, cold hyperalgesia). It is also possible that the pain generating mechanism and the symptoms change during the course of the disease. A thorough analysis of sensory symptoms may, reveal the underlying mechanisms that are mainly active in a particular patient. The treatment of neuropathic pain is currently unsatisfactory. In the future, drugs will be developed that address specifically the relevant combination of mechanisms. PMID- 10870736 TI - Mechanisms of central pain. AB - Central pain is common in patients with stroke, multiple sclerosis, syringomyelia, and spinal cord injury. It frequently develops after a delay of weeks or months, is associated with sensory change involving the spinothalamic pathways, and has a poor prognosis for spontaneous remission. Hypotheses to explain the varied clinical manifestations can be divided in two categories: those stressing aberrant neural activity in the deafferented circuits and those focusing on the postlesion imbalance between facilitatory and inhibitory neural pathways. All models inherently assume a degree of specialization of cerebral structures in pain processing, which has not been proved conclusively. PMID- 10870737 TI - A clinical approach to complex regional pain syndrome. AB - The clinical approach to complex regional pain syndrome (CRPS) is complicated by a lack of precision diagnostically, and a lack of evidence-based information for treatment. The vagaries of diagnosis were somewhat improved by the Orlando Conference (1993), where a consensus panel of experts developed a new taxonomy and criteria. Unfortunately the criteria can be based entirely on subjective grounds (patient history), and as such provides a very sensitive but not very specific device. There is some effort in the research community to amend these criteria to make them more specific. We encourage the practicing physician to include as much objective data along with the quasi-objective and subjective information currently used in formulating the diagnosis. This imprecision in diagnostic issues has significantly hampered treatment because it has not led to solid, generalizable, randomized controlled trials. To date there are no substantial scientific trials of any particular therapy or medication in the specific diagnosis of CRPS. Much can be inferred from the work with peripheral neuropathy and central pain. However, it is unlikely that this will be a perfect concordance with best therapy for CRPS. It remains our responsibility to diagnose each patient as best we can, supported by the best possible objective signs and testing. Once the diagnosis is made it is necessary to proceed in a pragmatic empirical way, following the best guidelines available. The guidelines should be considered a "rough sketch" and the key to clinical success will be flexibility, a vast fund of the available knowledge, patience, and compassion. To allow the deficiencies in the science to paralyze the clinical process is therapeutic nihilism, and not acceptable. PMID- 10870738 TI - Complex regional pain syndrome (type I, RSD; type II, causalgia): controversies. AB - Introduction of the term complex regional pain syndromes (CRPS) as a replacement of the older terminology, reflex sympathetic dystrophy (RSD) and causalgia, has achieved two goals: it has focused attention on the diagnosis and treatment, and sent basic scientists back to their laboratories. The relation of sympathetically maintained pain and sympatholysis is examined, particularly as a neuropathic process that is found in many conditions, including CRPS. This review also focuses on recent observations proposing a pathologic basis in support of diagnosis and treatment of these disorders. PMID- 10870739 TI - Neuropathic pain in cancer and AIDS. AB - Neuropathic pain is highly prevalent in patients with cancer and patients with AIDS, has profound effects on ability to function and quality of life, and is undertreated. Multiple obstacles to the adequate treatment of pain in patients with cancer and AIDS have been identified. Specific factors relevant to neuropathic pain, as well as the prevalence of substance abuse disorders in the AIDS population, contribute heavily to the undertreatment of pain in these patients. The differential diagnosis of neuropathic pain in these settings is broad, and a methodical diagnostic approach is required to achieve the primary objective of effective primary therapy. The parallel objective of providing optimal analgesic treatment also requires an aggressive and systematic approach. The presence of comorbid substance abuse issues requires special considerations that ordinarily do not compromise analgesic approaches. This review summarized the neuropathic pain syndromes seen in cancer and in AIDS, presents principles of pain assessment, highlights treatment options, and addresses the issue of pain and chemical dependency. PMID- 10870740 TI - The treatment of neuropathic pain: antidepressants and opioids. AB - OBJECTIVE: The objective of this article was to review the positive scientific data on antidepressants and opioids, which are largely confined to randomized controlled trials in two neuropathic pain conditions that have proved to be good models for clinical investigation. These two disorders are postherpetic neuralgia and painful diabetic neuropathy. DESIGN: This is a review of the literature using MEDLINE, CINAHL, and the Cochrane Database. RESULTS: There is extensive literature supporting the use of the older antidepressants such as amitriptyline in neuropathic pain. Newer randomized controlled trials support the use of opioids. CONCLUSIONS: First-line therapy for neuropathic pain may be either an older generation antidepressant such as amitriptyline or nortriptyline or the anticonvulsant gabapentin. For refractory cases, chronic opioid therapy may be the only avenue of relief, and evidence is accumulating that this approach is safe if proper guidelines are observed. PMID- 10870741 TI - Neural blockade for neuropathic pain. AB - OBJECTIVE: This study reviews the available literature regarding the use of nerve blocks for the management of peripheral neuropathy. DESIGN: A MEDLINE literature review was carried out, examining the existing literature about the use of nerve blocks for the treatment of mononeuropathies and radiculopathy. INTERVENTIONS: The study encompassed the use of local anesthetic peripheral and sympathetic nerve blocks, perineural steroid injections, and neurolytic blocks. RESULTS: Local anesthetic peripheral and sympathetic blocks provide useful diagnostic information but tend to afford only temporary therapeutic benefits in patients with peripheral neuropathy. Perineural steroids provide lasting relief for some patients with radiculopathy and peripheral neuropathy, but in most studies, it is a minority of patients who experience long-term relief. Peripheral neurolytic blocks are helpful for some patients with peripheral mononeuropathy, particularly in the setting of cancer pain. Although there are the risks of exacerbating pain and serious complications, certain patients experience long-term relief. CONCLUSIONS: Most discussions on the management of peripheral neuropathy do not include the use of nerve blocks. Nevertheless, the nerve block procedures discussed here can play an important role in the management of these conditions. PMID- 10870742 TI - New analgesics for neuropathic pain: the lidocaine patch. AB - Despite the availability of different pharmacologic agents for the treatment of various chronic neuropathic pain syndromes, complete symptom reduction and/or complete functional restoration is rarely achieved. New, safe, and effective treatments for chronic neuropathic pain, therefore, must be developed. One such agent, the lidocaine patch (Lidoderm, Endo Pharmaceuticals, Inc., Chadds Ford, PA), has been approved recently by the US Food and Drug Administration for the treatment of postherpetic neuralgia. Like other local anesthetics, the lidocaine patch results in sodium channel blockade, dampening, both peripheral nociceptor sensitization and, ultimately, central nervous system hyperexcitability. The Lidoderm patch is a topical agent and, consequently, insignificant serum levels are achieved even with chronic use. This fact enhances its safety. Recent studies have suggested that the lidocaine patch may be effective for chronic neuropathic pain conditions other than postherpetic neuralgia as well. PMID- 10870743 TI - Anticonvulsants (antineuropathics) for neuropathic pain syndromes. AB - Our knowledge about the pathogenesis of neuropathic pain has grown significantly during last two decades. Basic research with animal models of neuropathic pain and human clinical trials with neuropathic pain have accumulated solid evidence that a number of pathophysiologic and biochemical changes take place in the nervous system at a peripheral or central level as a result of the insult or disease. Many similarities between the pathophysiologic phenomena observed in some epilepsy models and neuropathic pain models justify the rationale for the use of anticonvulsant drugs in the symptomatic management of neuropathic pain disorders. Carbamazepine (CBZ) was the first representative from this class of drugs to be studied in clinical trials. It has been used for the treatment of neuropathic pain syndromes, in particular, trigeminal neuralgia (TN), for the longest time of any of the drugs in this class. Results from clinical trials support the use of CBZ in the treatment of TN, painful diabetic neuropathy, and postherpetic neuralgia. The use of CBZ was not studied for complex regional pain syndrome, phantom limb pain, and other neuropathic conditions, however. Phenytoin was the first anticonvulsant to be used as an antinociceptive agent, but based on clinical trials, there is no evidence for its efficacy in relieving neuropathic pain. Newer anticonvulsants have marked a new era in the treatment of neuropathic pain, with clinical trials of higher quality standards. Gabapentin (GBP) has most clearly demonstrated an analgesic effect for the treatment of neuropathic pain, specifically for the treatment of painful diabetic neuropathy and postherpetic neuralgia. Gabapentin has a favorable side effects profile, and based on the results of these studies, it should be considered a first-line treatment for neuropathic pain. Gabapentin mechanisms of action are still not thoroughly defined, but GBP is effective in relieving indexes of allodynia and hyperalgesia in animal models. It still remains to be seen whether GBP is as effective in other painful disorders. One small clinical trial with lamotrigine demonstrated improved pain control in TN. Evidence in support of the efficacy of anticonvulsant drugs in the treatment of neuropathic pain continues to evolve, and benefits have been clearly demonstrated in the case of GBP and CBZ. More advances in our understanding of the mechanisms underlying neuropathic pain syndromes should further our opportunities to establish the role of anticonvulsants in the treatment of neuropathic pain. PMID- 10870744 TI - The use of NMDA-receptor antagonists in the treatment of chronic pain. AB - Chronic pain can be maintained by a state of sensitization within the central nervous system that is mediated in part by the excitatory amino acids glutamate and aspartate binding to the N-methyl-D-aspartate (NMDA) receptor. A number of antagonists to the NMDA receptor are antinociceptive in animal models but are associated with significant dose-limiting side effects. Commercially available NMDA-receptor antagonists include ketamine, dextromethorphan, memantine, and amantadine. The opioids methadone, dextropropoxyphene, and ketobemidone are also antagonists at the NMDA receptor. The NMDA-receptor antagonists have a significant impact on the development of tolerance to opioid analgesics. Consequently, NMDA-receptor antagonists may represent a new class of analgesics and may have potential as coanalgesics when used in combination with opioids. PMID- 10870745 TI - Calcium and sodium channel antagonists for the treatment of pain. AB - Several lines of evidence developed in preclinical models suggest that both spontaneous and evoked pain are mediated in part by voltage-sensitive sodium and calcium channels, which are in abundance in both the peripheral and the central nervous system. There is an abundance of research, both preclinical and clinical, on the effects of the sodium and calcium channel antagonists on nociceptive processing. Clinical studies on the efficacy of the sodium channel antagonists in the treatment of acute and chronic pain have had mixed results. Preliminary studies of the N-type calcium channel antagonists for the treatment of acute and chronic pain are promising but too early to enable researchers to make firm conclusions. This review summarizes the current literature on the effects of the sodium and calcium channel antagonists on acute nociceptive processing, facilitated pain, and neuropathic pain. PMID- 10870746 TI - Clinical applications of capsaicinoids. AB - Capsaicinoids are molecules derived from chili peppers. They are best known for their pungency, producing mild to intense spice upon ingestion. It has long been known that peppers possess unusual qualities. Historically, they were burned in war rituals, and used to stimulate appetite. In 1997, a gene that encoded for a receptor specific for capsaicinoids was identified. Called VR1, this fatty acid receptor is present only on C fibers, and when activated produces desensitization or degeneration of the sensory afferent. Today, capsaicinoids are being studied as effective treatments for a number of sensory nerve fiber disorders, including pain associated with arthritis, cystitis, human immunodeficiency virus, and diabetic neuropathy. PMID- 10870747 TI - Prospects for the prevention of postherpetic neuralgia in herpes zoster patients. AB - OBJECTIVE: Herpes zoster is a common and painful disease that is caused by reactivation of the varicella-zoster virus. Herpes zoster pain that persists after healing of the acute infection is termed postherpetic neuralgia (PHN), a chronic pain syndrome that is often refractory to all treatment. The prevalence of PHN is expected to increase substantially in the coming decades, because the incidence of herpes zoster and the risk of PHN will both increase as the population ages. Although the results of recent studies provide a basis for improved treatment of patients with PHN, as many as half of all PHN patients do not obtain relief of their pain. Research on the development of improved treatments is continuing, but it has not been generally recognized that an equally important goal should be the design of interventions to prevent PHN. The prevention of PHN would lead to major reductions in disability, suffering, and the use of health care resources. DESIGN: The results of recent studies that have identified risk factors for the development of PHN and have implicated several peripheral and central mechanisms in its pathophysiology are reviewed. OUTCOME MEASURES: These risk factors and mechanisms of PHN provide a basis for hypothesizing that combining antiviral therapy with analgesic treatment beginning as soon as possible after the onset of herpes zoster would reduce the risk of PHN beyond that achieved by antiviral therapy alone. CONCLUSIONS: This treatment approach would be expected to reduce the risk of PHN in herpes zoster patients by attenuating acute pain and thereby preventing the initiation of central mechanisms of chronic pain. PMID- 10870748 TI - Psychological aspects of neuropathic pain. AB - Studies on the psychosocial impact of neuropathic pain conditions, including postherpetic neuralgia, diabetic neuropathy, complex regional pain syndrome, post spinal cord injury, postamputation, and AIDS-related neuropathy, are reviewed. Although limited, data are consistent with the larger literature on chronic pain and indicate that neuropathic pain reduces quality of life, including mood and physical and social functioning. Depression and pain coping strategies such as catastrophizing and social support predict pain severity, and a single diary study demonstrates a prospective relation between depressed mood and increased pain. Clinical trials of psychological interventions have not been reported, although some case series of successful treatment of neuropathic pain are reported, primarily in the area of biofeedback. Given the evidence indicating the broad impact of neuropathic pain on many areas of function, it is surprising that so few studies have investigated the impact of psychological interventions in these populations. PMID- 10870749 TI - What is clinically meaningful: outcome measures in pain clinical trials. AB - OBJECTIVE: The goal of this analysis is a better understanding of the issues involved in establishing the amount of change in pain that must be reported by subjects, participating in clinical trials and using standard pain scales, to indicate a clinically important difference. DESIGN: A review of the literature and a discussion of relevant concepts are presented. The focus is on outcome measures of pain commonly used in the studies described, including pain intensity, pain relief, global assessment of the medication effect, and requirement for an extra dose of rescue medication to treat a pain episode. The standard analysis statistics used to summarize the data are the central tendency of the groups being compared (i.e., mean, median, or mode), and the proportion of subjects that achieve one or more specific levels of benefit. RESULTS: The analysis of the proportion of responders in the groups being compared allows for a more easily understandable clinical importance of the results. CONCLUSIONS: An analysis of the proportion of responders is a clinically relevant analysis for many pain clinical trials and should be presented for one or more levels of response as appropriate. This will allow the readers to more easily interpret the results and apply them to clinical practice. PMID- 10870750 TI - Maxillofacial fractures following airbag deployment. AB - Maxillofacial fractures and associated lesions following airbag deployment were studied in six patients who suffered frontal or fronto-lateral car crashes. Installation of airbags in motor vehicles has reduced the morbidity and the mortality following motor vehicle accidents, but the appearance of new types of trauma directly related to airbag deployment raise questions about the potential danger of these devices when used improperly. The results of this limited study suggest that airbag injuries can be aggravated if: (1) seat belts are not worn; and (2) if the driver's chest is too close to the steering wheel as can easily happen with small people (in our study, two women). There needs to be a way of disconnecting the system. PMID- 10870751 TI - The role of thorax imaging in staging head and neck squamous cell carcinoma. AB - The overall survival rate for patients with head neck squamous cell carcinoma remains disappointingly static despite improved locoregional control. This has been attributed to the development of distant metastases and second primary malignancies in these patients, a large proportion of which occur in the thorax. We retrospectively analysed the incidence of thoracic malignancies in 138 patients presenting with newly diagnosed (n = 107) or recurrent (n = 31) cancer of the head and neck over a 4-year period. All 138 patients had undergone both computerised tomography of the thorax (CT) and conventional chest radiography within one month of presenting with biopsy proven squamous cell carcinoma. Seventeen percent of these were found to have simultaneous thoracic malignancies. CT thorax was more sensitive in detecting simultaneous thoracic malignancies compared with standard chest X-ray (24/138 versus 9/138, odds ratio of 3:1 in favour of CT). All thoracic malignancies detected by chest X-ray were also detected by CT thorax. Patients presenting with recurrent tumors were significantly more likely to have simultaneous thoracic malignancies than those with newly diagnosed cancer (11/31 versus 13/107, chi2 test with Yates correction, chi2 = 4.66, p = 0.03). The primary site (laryngeal, oral or pharyngeal) or presence of nodal disease did not have an effect on the incidence of simultaneous thoracic malignancies. The presence of distant metastases and second primary malignancies has major implications in the management and prognosis of patients presenting with head and neck squamous cell carcinoma, with a large proportion of such patients succumbing to their disease within one year of diagnosis. As CT scanning of the thorax was a more effective screening investigation than standard chest X-ray in the detection of simultaneous thoracic malignancy, we recommend it for use in the staging of patients presenting with cancer of the head and neck. PMID- 10870752 TI - Computer aided three-dimensional analysis of nostril forms: application in normal and operated cleft lip patients. AB - The appearance of the nostril in cleft lip patients is very important in the subjective assessment of naso-labial forms and patient satisfaction. To improve the outcome of plastic surgery, a computer aided diagnostic system was developed. Facial forms were measured with a three-dimensional optical scanner (Ogis Range Finder RFX-IV) XYZ coordinates (256x240) and RGB (red, green, blue) image (512x480) data sets were then obtained with the apparatus. The nostril area was determined by discriminant analysis of the RGB data, and the landmarks of the nostril were extracted under geometric conditions. To assess the reliability of this technique with head inclination, five volunteers were measured in seven postures. Landmark stability was within approximately 1 mm when the Frankfort plane was 45-60 degrees. Subsequently, this system was applied to two cleft lip patients who had undergone a secondary nasal correction. For control data, 37 healthy adults (22 males and 15 females) were measured in the same manner. Nasal asymmetry in the unilateral case and wide and flat nostrils in the bilateral case were greatly improved after surgery. Conversely, the volume of the nasal tip decreased. This system was a great help in the diagnosis of nostril abnormalities. PMID- 10870753 TI - Dental implants in alveolar cleft patients: a retrospective study. AB - Since 1994, 15 cleft patients have received dental implants after treatment of the alveolar clefts by osteoplastic measures. Altogether 23 dental implants were undertaken. Twelve implants were inserted after tertiary osteoplasty, 7 with simultaneous repeat-osteoplasty, and another 4 implants following two-stage re osteoplasty. In 11 patients with a total of 17 implants clinical and radiological follow-ups were performed. An individual crown was constructed on each of the implants. The radiological evaluation revealed that 70% of the implants were fully embedded in bone. All implants except one remained in situ and stable. Thus, the success rate was 96% retention. It is believed that a dental implant in the alveolar cleft may be lost because of its unfavourable location in terms of stability and function or because it was inserted too late. Scars resulting from previous surgery can also play a role in the failure of an implant. Therefore it is proposed that dental implants should not be inserted later than 6 to 8 weeks following osteoplasty of the cleft alveolus. PMID- 10870755 TI - Finite-element-analysis of different screw-diameters in the sagittal split osteotomy of the mandible. AB - A three dimensional finite element model of the mandible was developed to simulate and study the biomechanical loads of osteosynthesis screws in bilateral sagittal osteotomy. Using the finite-element method clinical conditions were simulated. Different bicortical screw configurations and diameters were evaluated. When bite forces were applied, the most stable configuration was found to be a triangular one. This confirms the results found in the literature. A mini screw of 2.0 mm diameter can provide sufficient stability at the osteotomy site after ramus split osteotomy. Even screws with a diameter of 1.5 mm would withstand forces up to 89.5 N, which would not normally be reached by patients after ramus split osteotomy in the early period of healing. Forces exerted by patients after bilateral ramus split osteotomy do not exceed these values. The finite-element analysis appears to be an adequate method to evaluate this clinical question of interest. It might well replace mechanical models and the results are comparable with those reported in the International literature. PMID- 10870754 TI - A cephalometric study of the stability of the base of the pharyngeal flap following a modified 'unified velopharyngoplasty procedure'. AB - A cephalometric study was conducted on 12 patients with repaired cleft palate to evaluate the stability in level and length of the base attachment of the velopharyngeal complex following pharyngeal flap surgery by a modified velopharyngoplasty. Complete velopharyngeal closure and normal articulation with a speech appliance were confirmed in all patients prior to pharyngeal flap surgery, which was performed on patients 10 years of age and above. Cephalometric radiographs were taken immediately, 1 year, 2 years and 3 years postoperatively. Cephalometric analysis revealed that although the level and length of the base of the velopharyngeal complex showed changes during the first postoperative year, they remained stable when compared with the palatal plane during the last two years. This indicated therefore that the base of a velopharyngoplasty should be attached at the same level of the palatal plane, namely the level of velopharyngeal closure, and that the procedure appeared useful in producing a stable velopharyngeal complex. PMID- 10870756 TI - Condylar changes after contralateral mandibular osteotomy in the rabbit. AB - Following orthognathic surgery undesirable changes in the temporomandibular joints (TMJs) are noted sometimes. Altered stress on the condyle and surrounding tissue frequently causes morphological changes in the TMJ. The purpose of the present study was to assess these morphologic changes in a rabbit following mandibular osteotomy and the resulting mandibular body rotation. Adult male Japanese white rabbits (3 kg, 12-16 weeks old) were applied in this study. An experimental mandibular osteotomy was performed on the right side of the body of the mandible. The surgery includes vertical mandibular body osteotomy 5 mm in width after masseter muscle reflection. Then the anterior and posterior osseous segments were fixed with wire at previously made holes. As a result, the condylar medio-lateral width on the contralateral (left) side was significantly larger than that of the ipsilateral side 2 weeks postoperatively (p < 0.05). There were significant differences in the angle between the right and the left condylar heads 4 weeks postoperatively (p < 0.05). The present experimental study demonstrated morphological changes in the contralateral TMJ caused by unilateral mandibular osteotomy. Postoperative morphological changes implied that there is a process of biological adaptation in the rabbit TMJ. PMID- 10870757 TI - Proximal segment position after distraction with the MD-DOS device. AB - We investigated cephalometrically the movement of the proximal segment in the sagittal plane in patients treated with distraction (MD-DOS device) for mandibular lengthening. The proximal segment was anteriorly rotated, whilst the distal segment was posteriorly rotated after the lengthening procedure. Thus the angle of the jaw was advanced half the distance of the advancement of the distal segment. One possible reason for the anterior rotation of 3.3 degrees on average is the repositioning of the proximal segment during application of the anterior fixation unit in the cases where mobilization was complete. Another more plausible reason is the anterior pull by the masticatory muscles and elastic bands being greater than the reactive distraction vector component in concert with a flexible telescopic distraction module and a single posterior screw anchorage. The positional movements of both distal and proximal segments were similar to those observed after mandibular advancement with bilateral sagittal split osteotomies. PMID- 10870758 TI - Report of the 'Distraction in Bruges' meeting, held under the auspices of the EACMFS. European Association for Cranio-Maxillofacial Surgery. PMID- 10870759 TI - Arthroscopic eminoplasty for habitual dislocation of the temporomandibular joint: preliminary study. AB - A unique surgical technique, arthroscopic eminoplasty was undertaken in 16 joints of 11 patients with habitual dislocation of the temporomandibular joint (TMJ). There were 10 joints with subluxation and 6 joints with complete dislocation in 4 male and 7 female patients with a mean age of 33 +/- 20 years. The procedure consisted of conventional diagnostic arthroscopy, followed by shaving of approximately 3 to 5 mm in height of the articular eminence with an electric motorized shaver with bone files, depending on the bone thickness as detected by preoperative imaging. The arthroscopic eminoplasty was accomplished without any peri- or postoperative complication. During the postoperative follow-up period of 19 months on average (6-36 months), all patients were free of dislocation of the TMJ, except for one joint. The patients could open their mouth 42 +/- 6 mm without arthralgia 2 weeks after surgery, and finally 47 +/- 7 mm without any subjective symptom but small joint noises (clicking or crepitus) in 10 joints. On postoperative radiographs only minor changes of the mandibular condyle were apparent in four joints. Arthroscopic eminoplasty might become a significant procedure for habitual dislocation of the TMJ and seems to produce results comparable to open arthrotomy. Further study will be required to assess this method as an acceptable modality in the future. PMID- 10870760 TI - Distraction osteogenesis of a fibula free flap used for mandibular reconstruction: preliminary report Sergio Siciliano, Benoit Lengere, Henre Reychler. PMID- 10870761 TI - Measurement of plasma oestradiol after an injection of a gonadotrophin as a test for neutered bitches. AB - Twenty-eight bitches with unknown reproductive histories were injected intravenously with either human chorionic gonadotrophin (hCG) or equine chorionic gonadotrophin (eCG) (pregnant mare's serum gonadotrophin) and their oestradiol responses were measured at the time of the injection and 90 minutes later. They were at various stages of the oestrous cycle as determined by histology and a progesterone assay for luteal function. Twenty-six of them were considered to be entire because they showed either an increase in plasma oestradiol over preinjection values or steady high values. The ovaries were removed from 25 of these animals and the other probably had a remnant of ovary because it came into oestrus some weeks later. In two remaining bitches no oestradiol could be detected either before or after the injection of gonadotrophin and they were predicted to have been neutered, which was confirmed at laparotomy. In the entire bitches, the highest plasma oestradiol concentration was measured during metoestrus and the lowest during anoestrus. PMID- 10870762 TI - Newcastle disease: outbreak losses and control policy costs. AB - The costs of controlling and eradicating the epidemic of Newcastle disease in Northern Ireland in 1973 are presented. The parameters of the 1973 epidemic have been adjusted to simulate the effect of the same epidemic in 1997, taking into account the relative changes of input and output prices, and the changes in the structure of the poultry industry. The costs and their distribution between producers, government and the industry, in 1973 and 1997 are compared, and the costs of an alternative vaccination strategy are compared with the eradication policy in both years. PMID- 10870763 TI - Frictional properties of cattle hooves and their conformation after trimming. AB - The conformation of the hooves of nine heifers obtained from an abattoir was measured and their coefficient of static friction (mu) was determined for movement in forward, backward and sideways directions. The hooves were trimmed by the Dutch method and their conformation and p were measured again. Trimming increased mu, and decreased the length of the digit and the angle between it and the floor. The value of mu was considerably greater for movement in a forward or backwards direction than sideways. The relationship between the value of mu for sideways movement and hoof conformation was examined by using 52 lower limbs from a variety of cattle breeds. Although mu was greater for Belgian blue than other cattle breeds, there were no relationships between mu and hoof conformation, including a measure of the roughness of the sole. PMID- 10870764 TI - Unidentified parasitic cysts in cattle. AB - Between February 18, 1995, and July 1, 1996, 38 cysts derived from New Zealand cattle were subjected to a diagnostic polymerase chain reaction (PCR) designed to identify genomic Taenia saginata DNA. No Tsaginata DNA was identified but an amplification product of 1078 bp was obtained consistently from several of the cysts. In Switzerland, suspect Tsaginata cysts are commonly positive for Tsaginata by PCR, but recently three cysts have also given a PCR fragment of 1078 bp, originating from a putatively unknown Taenia species. PMID- 10870765 TI - Application of the Vettest 8008 system for the biochemical analysis of vervet monkey plasma. AB - A clinical biochemistry analyser designed specifically for veterinary use was used to analyse plasma samples from 24 vervet monkeys (Cercopithecus aethiops). Two millilitres of heparinised blood was collected from each of the 24 monkeys on four occasions at intervals of one week. Plasma was separated and analysed for the concentrations of triglycerides, cholesterol, total proteins, albumin, globulins, creatinine and blood urea nitrogen (BUN) and the activities of alkaline phosphatase (AP), lactate dehydrogenase (LDH), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and creatine kinase (CK). The tests were easy to perform, used small volumes of plasma, and yielded consistent results for most of the analytes. The activities of CK and AP, but not AST, appeared to be influenced by haemolysis, and there were significant individual variations in the activity of LDH. PMID- 10870766 TI - Intestinal intussusception in five postparturient queens. PMID- 10870767 TI - MRI of the equine digit with a dedicated low-field magnet. PMID- 10870768 TI - Dispensing under review. PMID- 10870769 TI - Use of lignocaine in food animals. PMID- 10870770 TI - Possible treatment options for PMWS/PDNS. Porcine multisystemic wasting syndrome/porcine dermatitis and neuropathy syndrome. PMID- 10870771 TI - Cardiac disease in guinea pigs. PMID- 10870772 TI - A wish-list for improving Australian women's reproductive health in the new millennium. PMID- 10870773 TI - A template for defining a causal relationship between acute intrapartum events and cerebral palsy: international consensus statement. International Cerebral Palsy Task Force. PMID- 10870774 TI - Term breech birth in New South Wales, 1990-1997. AB - Data on 636,708 women delivering a singleton infant of gestational age > or =37 weeks in NSW from 1 January 1990 to 31 December 1997 were used to examine trends in breech births at term and the mode of delivery. From 1990 to 1997, although the crude rate of breech births at term remained stable at 3.4%, the adjusted odds ratio for breech birth compared with cephalic birth decreased over time. Among live breech births, the crude rate of vaginal breech birth declined from 29.4% to 19.7%, with an attendant increase in elective Caesarean sections from 49.1% to 58.4%. Most of this increase was at 38 and 39 weeks gestation. There was no change in the perinatal mortality rate among breech births during the study period. Despite increasing maternal age, the adjusted odds of a breech birth at term decreased over time. This could be due to offsetting factors, such as increased use of external cephalic version. If the decrease in vaginal breech birth continues, it may lead to the skills for this procedure being lost. PMID- 10870775 TI - Risk factors related to premature rupture of membranes in term pregnant women: a case-control study. AB - A case-control study of premature rupture of membranes (PROM) of full term pregnant women was undertaken between 1 November 1996 and 30 July 1997 at Rajavithi Hospital to determine the risk factors related to PROM. Two hundred and twenty pregnant women with PROM and 220 pregnant women without PROM were recruited by a simple random sampling. The diagnosis of rupture of membranes was made from history and from positive microscopic ferning and pH tests performed during a speculum examination. The demographic data was not significantly different between the two groups. The risk factors, such as a history of PROM in a previous pregnancy, a history of abortion in a previous pregnancy, and body mass index (BMI) < 20 were significantly different between the PROM group and the control group. However, on using multiple logistic regression analysis, we found that the residual significant risk factors were a history of PROM in a previous pregnancy and BMI < 20. PMID- 10870776 TI - Management of essential thrombocythaemia during pregnancy. AB - Essential thrombocythaemia is a rare myeloproliferative disorder that often presents with haemorrhagic or thrombotic complications. It may be detected incidentally in an asymptomatic younger adult and there are only a few case reports of essential thrombocythaemia in pregnant women. The risks posed by essential thrombocythaemia during pregnancy and its optimal management are uncertain. To determine if there is increased incidence of obstetric complications seen in women who have essential thrombocythaemia, we collected a large case series from a number of tertiary obstetric units in Australia and New Zealand. There were 30 pregnancies in 12 women who had essential thrombocythaemia. There were 17 live births (57%), 7 stillbirths (23%), 5 miscarriages (17%) and 1 ectopic (3%). Five pregnancies were complicated by placental abruption. When the outcomes of those women who received treatment with aspirin or interferon were compared to those that did not receive any treatment, there was a trend towards a higher livebirth rate (79% v. 38%, p = 0.06). Seven women were treated with aspirin and 5 had successful outcomes with no fetal complications. Four women were treated with alpha-interferon which reduced their platelet counts and all had successful outcomes with no fetal complication. These findings suggest that there is a high incidence of miscarriage, stillbirth and abruption in women with essential thrombocythaemia. Their pregnancies should be carefully monitored. Treatment with low dose aspirin and/or the use of alpha interferon may be associated with an improved pregnancy outcome. PMID- 10870777 TI - Outcome of pregnancies complicated by pre-gestational diabetes mellitus. AB - Pregestational diabetes mellitus (DM) is associated with adverse fetal and maternal outcomes. Studies suggest that optimal control of diabetes before and during pregnancy minimises these risks. There are few recent reviews of outcomes of pregnancies complicated by DM in Australia. Ninety-three pregnancies in women with DM at our hospital since 1989 were identified. We collected data for maternal age, type of diabetes, duration of therapy, complications of diabetes, maternal complications of pregnancy and fetal outcomes including malformations. The rate of pregnancy planning with optimal glycaemic control at conception was low in our population, particularly in patients with Type 1 diabetes. Women who smoked had worse glycaemic control, and a higher rate of miscarriage. There was a high rate of Caesarean section, particularly in those women with Type 1 diabetes (77.4%). The rate of Caesarean section was lower in planned pregnancies. There were no perinatal deaths. The number of neonates with major congenital anomalies was high (13%) in the Type 1 population. It is important to increase the rates of prepregnancy planning and to optimise glycaemic control before pregnancy. In many cases there has been a long interval between diagnosis and pregnancy, so all women with diabetes should receive counselling at frequent intervals about pregnancy and the importance of planning. Women who planned their pregnancies had improved outcomes, with decreased rate of Caesarean section, better glycaemic control and better neonatal Apgar scores. Women with diabetes should not smoke during pregnancy because of the increased risk of miscarriage and poorer glycaemic control. PMID- 10870778 TI - Single dose oral azithromycin versus seven day doxycycline in the treatment of non-gonococcal mucopurulent endocervicitis. AB - The aim of this study was to compare single dose oral azithromycin versus seven day doxycycline in the treatment of non-gonococcal mucopurulent cervicitis (MPC). One hundred and thirty-one women with non-gonococcal MPC were enrolled in a prospective-randomised study to compare the efficacy and safety of a single oral dose of 1 g azithromycin and a seven-day course of 100 mg doxycycline twice daily. Clinical examination and culture samples for Chlamydia trachomatis and other microorganisms were performed before and approximately 14 days after starting the treatment. Of the 131 women recruited (67 in the azithromycin group and 64 in the doxycycline group), Ureaplasma urealyticum was isolated from 21 (16%); Chlamydia trachomatis from 15 (11.5%); and Mycoplasma hominis from 3 (2.3%) of the patients at the initial examination. The eradication rate of baseline culture-positive cases at the follow-up visit in the azithromycin group was 71.4%, and 77.3% in the doxycycline group. There was no statistically significant difference in efficacy between the single dose azithromycin and seven day course of doxycycline in the treatment of culture-positive cases. Azithromycin 1 g appears to be an effective and safe alternative to doxycycline for the treatment of non-gonococcal MPC. PMID- 10870779 TI - The relationship of physical trauma and surgical stress to menstrual dysfunction. AB - To evaluate the incidence and pattern of menstrual dysfunction in reproductive age group women suffering acute musculoskeletal trauma, 198 women between 15 and 50 years of age admitted consecutively into an acute orthopaedic unit were recruited over a 6-month period. The patients were then followed up for 6 months with menstrual diaries to compare their menstrual pattern with their preadmission status. Excluding those with significant menstrual problems before admission, the menstrual pattern remained normal in 135 (68%) (EM), while 12 (6%) developed polymenorrhoea (PM), and 51 (25%) had oligomenorrhoea or amenorrhoea (OAM) within the 6-month observation. The three groups did not differ in their mean age, body mass index, parity or age of menarche, but previous cycle lengths were shortest in the PM group (25.4 days, SD 7.64) (p<0.05) and history of amenorrhoea in the previous one year was most common in the OAM group (p<0.025). Univariate analysis showed the incidence of moderate to major trauma,operative treatment, longer operative time, general anaesthesia, blood transfusion and immobilisation were significantly higher in the PM and OAM groups compared to the unchanged group (p<0.05). A logistic regression model showed that general anaesthesia and longer surgical operations remained significantly related to the development of menstrual dysfunction. We conclude that the pattern of menstrual dysfunction after acute orthopaedic trauma appeared to be dictated by the woman's pre existing menstrual characteristics and the stress of surgical treatment. PMID- 10870780 TI - Prevalence of gestational diabetes mellitus in polycystic ovarian syndrome (PCOS) patients pregnant after ovulation induction with gonadotrophins. AB - Our aims were: 1. To investigate if women with PCOS who become pregnant using gonadotrophins have a higher incidence of gestational diabetes mellitus (GDM) compared to spontaneously pregnant matched control women, 2. To compare the prevalence of GDM in PCOS women with that in women with hypo/eugonadotrophic hypogonadism and in unexplained infertility and 3. To investigate differences in pregnancy outcomes between the groups. This was a retrospective case-control study. Women with PCOS were matched with a control by age, BMI, and ethnicity. There were 60 women with PCOS, 11 with hypogonadotrophic hypogonadism, 6 with eugonadotrophic hypogonadism, and 12 with unexplained infertility. Control women were those who attended a major public hospital for antenatal care and delivery We found no difference in the prevalence of GDM between the PCOS (22%) and the controls (17%) or between the PCOS and other groups. Women with GDM (diet or insulin controlled) had a significantly higher BMI than women without GDM (p = 0.019). There was no difference in pregnancy outcomes between the groups. There was a significant dependence of babies' birthweight on mother's BMI (p<0.001). PMID- 10870781 TI - Incisionless cystourethropexy--the Intac device--an initial experience. AB - Our aim was to evaluate a new minimally invasive device for the treatment of genuine stress incontinence in women by both objective and subjective measures of outcome. Fifteen women with Type I or Type II urinary stress incontinence were treated with a new per vaginal bone anchor device designed to fix periurethral tissues to the pubic bone. No patients had significant operative morbidity. After 6-13 months a follow-up questionnaire and repeat urodynamic testing was performed for all the women. Subjectively 73% of patients had improvement in symptoms and on urodynamic testing 53% of patients were cured. PMID- 10870782 TI - Referral patterns for gynaecological radiotherapy in Victoria. AB - A retrospective chart review was undertaken on all patients in Victoria who were referred for radiotherapy for a gynaecological cancer from February 1997 to January, 1998. Three hundred and ten patients were identified which represents less than one-third of all gynaecological cancers diagnosed in Victoria each year. Ninety-two of the 310 patients (30%) referred for radiotherapy were managed without the prior involvement of a certified gynaecological oncologist. The 310 patients included 95 patients with cervical cancer, 33 patients with ovarian cancer and 142 patients with endometrial cancer. The initial management strategies employed for patients with the major gynaecological cancers varied depending on the source of referral. This difference was most marked in endometrial cancer due mainly to differing indications for full surgical staging and subsequent referral for radiotherapy both between types of specialists and also between gynaecological oncology units. The development of evidence based guidelines in the major gynaecological cancers should lead to a more uniform approach to the care of women with gynaecological malignancies. PMID- 10870783 TI - Surgery and post-operative radiotherapy for early stage cervical cancer. AB - The use of post-operative radiotherapy in the treatment of cervical cancer is controversial. The aim of this study was to document the results and toxicity of adjuvant irradiation in patients with Stage 1B and 2A cervical cancer. We performed a retrospective review of all patients treated with post-operative radiotherapy at Royal Prince Alfred Hospital between 1986 and 1993. Patient, tumour and treatment factors and late toxicity were recorded. Relapse-free and overall survival were calculated. Eighty-one patients form the study population. The median follow-up was 6.1 years. Fifty-eight patients (72%) had stage 1B cervical cancer and 23 (28%) stage 2A. The 5 year relapse-free and overall survival were 78% and 80% respectively. Six patients (7%) had late toxicity requiring inpatient medical treatment and 6 patients (7%) required surgery. The survival was comparable to other series reported in the literature. There was an incidence of 14% late toxicity requiring medical or surgical intervention which is greater than with hysterectomy or pelvic irradiation alone. Clinical prognostic factors should be used to select patients for either surgery or radiotherapy alone to minimise the increased toxicities associated with a combination of surgery and radiotherapy. PMID- 10870784 TI - Antenatal exercise and birthweight. AB - Does strenuous antenatal exercise reduce birthweight? Does reducing maternal exercise increase birthweight? What to advise about exercise during pregnancy? We recruited 117 women who intended to exercise 5 or more times weekly during pregnancy to a study of whether reducing the amount of maternal exercise during pregnancy is associated with an increase in birthweight. Only 61/117 (52%) of women agreed to be randomised to either continue or to reduce (to 3 or fewer sessions of exercise weekly) their intended pregnancy exercise program. Most women who refused randomisation did not want to risk being asked to reduce their exercise during pregnancy. Within the randomised trial, there was no statistically significant difference between the mean birthweight of babies born to women who continued and those who reduced their intended exercise program. The high rate of refusal of randomisation limits the power of the study to find a difference in birthweight, limits the generalisability of the results and shows that many women intending to exercise at this level during pregnancy have an uncompromising attitude to exercise. PMID- 10870785 TI - Validation of women's self-reported smoking status in a Sydney colposcopy clinic. AB - This study validated a self-reported smoking prevalence questionnaire against urinary cotinine levels among women referred to a Sydney based public hospital colposcopy clinic over the period November 1997 to June 1998. Of 213 eligible women, 160 (75%) agreed to participate. Of these, 151 (94%) completed the smoking prevalence item and 130 (86%) also provided a urine specimen. 40% (95% CI: 33% 49%) of respondents self-reported as current smokers. Observed agreement between self-reported and biochemically validated smoking status was 94% compared with 52% chance agreement (kappa = 0.87, p<0.001). This high level of agreement was also robust to sensitivity analyses using a more conservative cutoff for the urinary cotinine levels. We conclude that self-reported smoking status provides valid estimates of actual rates of smoking among women referred with abnormal cervical smears. PMID- 10870786 TI - Plasma HIV-1 RNA viral load and serum vitamin A and E levels in HIV-1 infected pregnant women. AB - The aim of this study was to assess the correlation between plasma HIV-1 RNA viral load and serum vitamin A and E concentrations and vitamin E/cholesterol ratio in HIV-1 infected pregnant women not receiving antiretroviral therapy There were no significant correlations between plasma HIV-1 RNA viral load and serum vitamin A and E concentrations and vitamin E/cholesterol ratio. However, the presence of underlying vitamin A deficiency in these pregnant women was common. PMID- 10870787 TI - A RACOG sponsored pilot study of a quality assurance program regarding management of labour by provincial Fellows. Royal Australian College of Obstetricians and Gynaecologists. AB - OBJECTIVE: To develop an effective and practical self-administered obstetric audit program for use by clinicians within their own practice. SETTING: The private and public practices of specialists in provincial practice. SAMPLE: Two periods of 3 months in each Fellow's practice, separated by a period of 3 months to allow for data review, resulting in the review of management of 6708 singleton births. METHODS: All provincial Fellows in active practice in Australia in early 1998 were invited to take part in a voluntary 'quality cycle' obstetric practice audit. The data from the first 3 month period was fed back to participating Fellows for review before a second 3-month audit period was undertaken. RESULTS: One hundred and twenty provincial Fellows were invited to take part; 62 registered for the study, 58 commenced the project, and 52 completed the entire cycle. 60.1% of the 6708 women studied laboured spontaneously, 25.8% had labour induced, and 14.1% had elective Caesarean sections. 87.8% of the 5759 women who laboured gave birth vaginally. There was little change in the incidence of intervention in labour between the first and second study periods. CONCLUSIONS: It is possible to design a worthwhile self-administered clinical audit in obstetric practice with which specialists in full-time practice can cope and which provides useful personalised feedback for the specialist. PMID- 10870788 TI - Vaginal birth after Caesarean section: an Australian multicentre study. VBAC Study Group. AB - Retrospective analysis of medical records and individual case review was undertaken at 11 major obstetric hospitals for a 5 year period from July 1992 to June 1997 to investigate rates of vaginal birth after Caesarean section (VBAC), the occurrences of uterine rupture, and the outcomes for mother and infant following rupture. Total deliveries were 234,015, of which 21,452 or 9.2% were associated with one or more previous Caesarean sections. Within this scar group, 5419 patients or 25.3% were delivered vaginally. There were 62 cases of significant uterine rupture with no maternal deaths. Perinatal mortality with rupture was 25% and serious maternal complications (usually hysterectomy) occurred in 25% of those with uterine rupture. In women attempting vaginal delivery after a previous lower segment Caesarean section, the uterine rupture rate was estimated at 0.3%, with 0.05% experiencing a perinatal death and 0.05% requiring a hysterectomy. Although VBAC rates in Australia remain lower than many overseas reported series, rates are increasing. While rupture continues to be associated with serious adverse outcomes, the incidence of rupture during trial of labour is low and appears to be associated with a better outcome than rupture of an unscarred uterus. PMID- 10870789 TI - Rate of vasectomy rises with increasing income. PMID- 10870791 TI - Sickle cell disease in pregnancy. AB - Homozygous sickle cell anaemia (Hb S) is the most common major haemoglobinopathy in the United States, occurring in approximately 1 in 626 African Americans. While haemoglobinopathies involving Hb S occur commonly in blacks of African descent, they are also found in people of Middle Eastern, East Indian and Mediterranean origin. It is an uncommon disease, especially in homozygous form, in Australia. We present the case of a woman in her third pregnancy, originally from Ghana, with HB F, and discuss the current issues in the management of sickle cell disease in pregnancy. PMID- 10870790 TI - More on asphyxia, cerebral palsy and litigation. PMID- 10870792 TI - An unusual case of choriocarcinoma following live term pregnancy. AB - Post-term choriocarcinoma is an infrequent event with poor prognosis. The diagnosis is usually delayed due to failure to recognise the mode of presentation of this disease. Being a rare occurrence, limited data is available regarding its clinical features. The choriocarcinoma in our patient presented as an isolated huge pedunculated growth over the uterine serosa without intrauterine involvement and distant metastasis. PMID- 10870793 TI - Incisional hernia in a 5 mm laparoscopic port site incision. AB - Herniation of omentum or bowel through laparoscopic incision sites is uncommon. Herniations through 5 mm ports sites are very rare, with less than a handful described in the literature. Undoubtedly however, with the increasing interest. PMID- 10870794 TI - Pneumomediastinum following vaginal delivery. PMID- 10870795 TI - Re: Home within 24 hours of laparoscopic hysterectomy. PMID- 10870796 TI - Vulvar vestibulitis syndrome (VVS) is not primarily a psychosexual disorder. PMID- 10870797 TI - Re: An unusual case of antepartrum haemorrhage. PMID- 10870798 TI - Re: The integrity of surgical gloves during gynaecological operations. PMID- 10870800 TI - Lipoprotein (a), cholesterol and triglycerides in women with venous thromboembolism. AB - Plasma concentrations of lipoprotein (a), total cholesterol, high-density lipoprotein (HDL)-cholesterol, low-density lipoprotein (LDL)-cholesterol and triglyceride were measured in 62 women who had suffered an episode of objectively confirmed venous thromboembolism (VTE) at < or = 50 years of age, and in 98 age matched female controls. The mean body mass index (BMI) of cases was significantly (P < 0.001) higher than that of controls. Plasma triglyceride was significantly higher, and total cholesterol/LDL- and HDL-cholesterol significantly lower, in cases compared with controls. After adjustment for BMI, the plasma total cholesterol and LDL-cholesterol remained significantly lower in cases. No significant differences in mean plasma lipoprotein (a) levels were identified between cases and controls. Lipoprotein (a) does not appear to be significantly associated with the development of VTE in young women. The increased risk of VTE in obese subjects may be mediated, at least in part, via hypertriglyceridaemia, which has previously been demonstrated to have effects on levels of coagulation factors, natural anticoagulants, and plasminogen activator inhibitor type 1. PMID- 10870799 TI - Tissue factor antigen and activity in serum of postoperatively shed blood used for autologous transfusion. AB - Orthopaedic surgery involves extensive dissection of connective and richly vascularised tissues rich in tissue factor (TF). It was, therefore, of interest to quantify the amount of TF antigen and activity in postoperatively drained, unwashed wound blood collected for the purpose of autologous transfusion. In nine young patients subjected to surgery for idiopathic thoracic scoliosis, samples were drawn postoperatively from collected shed blood, a pulmonary artery catheter and a radial arterial cannula prior to, during and after reinfusion of shed blood (10 ml/kg body weight), and analysed for TF antigen and activity. Preoperative arterial control samples contained 128 pg/ml TF antigen compared with 40 pg/ml postoperatively. During reinfusion of drained blood, arterial TF concentration rose to 96 pg/ml and dropped to 64 pg/ml after infusion. Arterial and mixed venous blood did not differ significantly in TF levels. Serum from drained blood contained high concentrations of TF antigen (773 pg/ml), but no TF activity was detected. It is concluded that the high concentrations of TF antigen in serum from postoperatively drained blood collected for autologous transfusion are devoid of procoagulant activity. The TF antigen in plasma of drained blood is suggested to be a soluble, proteolysed TF-apoprotein or a TF complex inactivated by the TF pathway inhibitor (TFPI). PMID- 10870801 TI - A low dose of aspirin (75 mg/day) lowers thrombin generation to a similar extent as a high dose of aspirin (300 mg/day). AB - This randomized, double-blind, parallel-group study was performed to assess the effect of 1-week treatment with 75 and 300 mg aspirin on thrombin generation. Eighteen healthy men, aged 20-25 years, entered the study. After 1 week of aspirin treatment with a daily dose of 75 mg, bleeding time became prolonged by 102 s (P = 0.02), and it was prolonged by 165 s with 300 mg (P = 0.0005). None of the doses of aspirin affected peripheral blood concentrations of prothrombin fragment 1+2 and fibrinopeptide A. At the site of microvascular injury, 75 mg aspirin led to a marked, about 60%, reduction in the total amount of thrombin generated (P = 0.04). A similar decrease was observed after 7-day treatment with 300 mg aspirin (P = 0.009). We conclude that the thrombin-lowering action of aspirin in the range between 75 and 300 mg daily given for 7 days is not dose dependent. PMID- 10870802 TI - Validation of an enzyme immunoassay for the determination of total homocysteine in plasma. AB - In recent years, the determination of homocysteine (Hcy) has become increasingly important, since high levels of Hcy in plasma or serum represent an independent risk factor for occlusive vascular diseases. Nowadays, clinical laboratories use several analytical techniques to measure Hcy, of which high-performance liquid chromatography (HPLC) is the most popular. Recently, assays for Hcy quantification based on enzyme immunoassays (EIA) have become commercially available. Our group carried out the validation of the Axis method and compared results with those obtained by an established HPLC assay. Intra- and inter-assay coefficients of variation were < or = 8.5%. Compared with HPLC, linear regression analysis showed r=0.984, slope=0.952, intercept = 1.24 /mol/l; Bland-Altman procedure, the mean of the difference EIA-HPLC results = 0.5 micromol/l. Our results suggest that Hcy determinations by both methods are equivalent, and that the Axis assay provides reproducible and reliable data. PMID- 10870803 TI - Decline of factor VIII and factor IX inhibitors during long-term treatment with NovoSeven. AB - Recombinant factor VIIa (rFVIIa) (NovoSeveng) is used to treat bleeding episodes in hemophilia A and B patients with inhibitor antibodies against factor VIII (FVIII) and factor IX. rFVIIIa has been studied in home treatment of mild-to moderate joint, muscle, and mucocutaneous bleeds to assess safety and efficacy. Treatment with other factor concentrates was allowed according to treating physician's judgment. Blood samples were drawn before study start and after 6 and 12 months. It has thus been possible to follow the inhibitor titres during this period. Analyses of 53 patients (49 hemophilia A, four hemophilia B) showed inhibitor levels up to 1,208 BU/ml before study start. Based on the first analysis, hemophilia A patients were divided into high responders (> 5 BU/ml; 28 patients), low responders (> 1 and < 5 BU/ml; 15 patients) and very low responders (< or = 1 BU/ml; six patients). In high responders receiving rFVIIa as only treatment, FVIII inhibitor titre decreased to one-third of the initial level. For high responders receiving other factor treatments such as FVIII or prothrombin complex concentrates, inhibitor titre remained unchanged. Titres for low responders and very low responders remained unchanged independent of treatment. Thus, when rFVIIa is used as the only coagulation factor to treat hemophilia A/B high-responder inhibitor patients, inhibitor level declines significantly. PMID- 10870804 TI - Fechtner syndrome: physiologic analysis of macrothrombocytopenia. AB - Fechtner syndrome is a rare autosomal dominant disorder consisting of macrothrombocytopenia and leukocyte inclusions, associated with Alport's syndrome (hereditary nephropathy, sensorineural hearing loss, and ocular anomalies). We describe a 71-year-old Caucasian male with a history of hearing loss and asymptomatic macrothrombocytopenia incidentally noted in 1985. Several challenges to hemostasis were uneventful, including total hip arthroplasty. He subsequently developed progressive renal failure, with 'nil lesions' by light and electron microscopy, which was responsive to corticosteroid therapy. Eight family members are affected variably by either thrombocytopenia or renal failure. Laboratory testing gave the following results: hemoglobin, 10.2 g/dl; leukocytes, 5.0 x 109/l; platelets, 64 x 109/l (mean platelet volume, 13.3 fl; normal platelet volume, 7.6-10.8 fl). Peripheral blood smear revealed thrombocytopenia and leukocytes with inclusions. Electron microscopy of the buffy coat confirmed Fechtner inclusions within the patient's leukocytes. Whole mount and thin section electron microscopy revealed a population of large, although not giant, platelets. Despite thrombocytopenia, platelet aggregation was normal. Flow cytometry of dilute platelets revealed normal glycoprotein alphaII beta beta3 activation and alpha-granule p-selectin secretory response to 10 nmol/l human alpha-thrombin. Dense granule adenosine triphosphate secretory response to thrombin was likewise normal. This case illustrates that 'giant' platelets are not universally present in Fechtner syndrome cases, although the platelets are enlarged. Finally, renal pathology other than Alport's nephropathy may be associated with this syndrome. PMID- 10870805 TI - Markers of hemostatic system activation in pulmonary embolism. Changes during and after cessation of anticoagulant treatment. AB - Plasma levels of prothrombin fragment 1+2 (Fl+2), thrombin-antithrombin complexes (TAT) and D-dimers were measured in 15 patients with pulmonary embolism during heparin therapy, oral anticoagulation, and after cessation of warfarin therapy. Each patient had a favorable outcome during anticoagulant therapy (3 months), but late venous thromboembolism occurred in six cases. The mean levels of the three markers were significantly increased on day 4 after the thrombotic event, and normalized during warfarin therapy. Nine months after the initial pulmonary embolism, mean levels of the three markers, as compared with a control population, were significantly higher in the patients with late recurrences, whereas only TAT were slightly higher in patients without recurrences as compared with controls. Only TAT levels were significantly higher in the patients with late recurrences than in those without late recurrences. Thus, the levels of the three markers 9 months after pulmonary embolism seem to be interesting to identify patients with high risk of recurrence and who might require longer anticoagulant treatment. PMID- 10870807 TI - Structural model of human alpha1-microglobulin: proposed scheme for the interaction with the Gla domain of anticoagulant protein C. AB - Alpha1-microglobulin (alpha1m) is a small glycoprotein with immunomodulatory properties. It is a member of the lipocalin family, a group of proteins that exhibit a well-conserved three-dimensional structure despite low sequence identity and that are known to bind small hydrophobic ligands. The types of ligands carried by alpha1m are still unknown, but it is known that this protein has yellow-brown chromophores attached to three lysines at position 92, 118 and 130. Alpha1m has one unpaired cysteine residue (Cys 34) that can form a disulphide bond with other proteins that also possess an exposed free unpaired cysteine. For instance, alpha1m interacts with the protein C (PC) Gla domain containing the Arg9Cys or Ser12Cys substitution. In order to gain insights about the alpha1m molecule and analyze the intriguing alpha1m-Gla domain interaction, it was decided to use bioinformatics. The three-dimensional structures of alpha1m and PC Gla domain were predicted. Alpha1m Cys 34 is solvent exposed and located near the entrance of the ligand-binding pocket. The chromophore-carrying lysines are found buried into the pocket, and the area around the entrance of this cavity displays about 10 positively charged residues. This electropositive region in alpha1m complements the essentially electronegative Gla domain and may play a role during intermolecular interactions. In addition, a few hydrophobic residues surround alpha1m Cys 34 and could be of importance during its interaction with macromolecular ligands. PMID- 10870806 TI - Human coagulation factor FVIIa (recombinant) in the management of limb threatening bleeds unresponsive to alternative therapies: results from the NovoSeven emergency-use programme in patients with severe haemophilia or with acquired inhibitors. AB - This open-label, emergency-use study evaluated the efficacy and safety of activated human coagulation factor VIIa (recombinant) (rFVIIa) (NovoSeven; Novo Nordisk Pharmaceuticals, Inc., New Jersey, USA) in treating limb-threatening joint or muscle bleeds in 17 patients with haemophilia A or B and six patients with acquired inhibitors to factor VIII or factor IX. All patients had previously failed on one or more alternative therapies. rFVIIa administration was effective or partially effective in controlling joint or muscle bleeds in 34 out of 35 (97%) bleeding episodes; in 23 patients, 14 of 17 (82%) muscle bleeds and 16 of 18 (89%) joint bleeds were effectively controlled. These findings suggest that rFVIIa is an effective and well-tolerated therapeutic option in the management of joint or muscle haemorrhage in patients with haemophilia and in patients with acquired inhibitors. PMID- 10870808 TI - Activated factor XII levels are dependent on factor XII 46C/T genotypes and factor XII zymogen levels, and are associated with vascular risk factors in patients and healthy subjects. AB - Coagulation factor XII (FXII) is activated on contact with various biologic surfaces, including subendothelial tissues and lipoprotein particles. Thus, the plasma level of activated FXII (XIIa) might represent vascular lesions or be a marker of abnormal lipid metabolism. A 46C/T polymorphism was recently described in the FXII gene close to the ATG translation initiation codon, which was associated with inter-individual variation of plasma FXII zymogen levels. The present paper reports the association of the 46C/T polymorphism with plasma XIIa levels in apparently healthy subjects, and in patients with ischemic cerebrovascular disease (CVD) and arteriosclerosis obliterans (ASO). XIIa levels were not significantly different between patients and controls, but were strongly dependent on XII 46C/T genotypes (2.07 +/- 0.81, 1.65 +/- 0.63, and 0.93 +/- 0.41 ng/ml for C/C, C/T, and T/T genotypes, respectively; P < 0.0001). This association was evident for each group studied (P < 0.0001 for CVD and controls; P= 0.0007 for ASO). There were positive correlations between plasma FXII clotting activity and XIIa levels. In a univariate analysis, XIIa correlated with total cholesterol, triglycerides, plasminogen activator inhibitor-1, and C-reactive protein (CRP), although the presence of conventional cardiovascular risk factors (male sex, smoking, hypertension, hypercholesterolemia, diabetes) did not significantly increase XIIa. Stepwise regression analyses revealed that the XII clotting activity had the strongest association with XIIa. In conclusion, XIIa levels depended on XII 46C/T genotype and correlated with some cardiovascular risk factors. Thus, the FXII genotype should be taken into consideration for interpretation of plasma XIIa levels. PMID- 10870809 TI - Anticoagulant versus amidolytic activity of tissue factor pathway inhibitor in coronary artery disease. AB - In view of raised levels of endothelial markers in coronary artery disease (CAD), the aim of the present study was to investigate the status of tissue factor pathway inhibitor (TFPI), another endothelium-associated glycoprotein and coagulation protease inhibitor, in CAD. The intravascular pool of TFPI is heterogeneous with respect to assay-dependent activity. While the standard amidolytic assay works well with both full-length and truncated (lipoprotein associated) TFPI, the anticoagulant assay works better with the former. The anticoagulant activity of TFPI can be estimated using dilute tissue factor (TF) to trigger clotting of plasma. In the present study, recombinant TF diluted 2,000 fold was used to initiate coagulation. A dose-dependent shortening of clotting time of normal plasma pools with polyclonal antibody against the C-terminal but not the N-terminal peptide of TFPI demonstrated the importance of the C-terminal region, and hence that of full-length TFPI, in conferring its anticoagulant activity, corroborating current opinion. As a further confirmation, the C terminal peptide itself prolonged dilute TF clotting time of normal pooled plasma in a concentration-dependent manner. The amidolytic and anticoagulant activities of TFPI were determined in 20 patients with clinically and angiographically assessed CAD and in 68 asymptomatic controls. The mean +/- SD ages of patients and controls were 54.9 +/- 10.3 and 48.8 +/- 11.6 years, respectively, the difference being statistically significant (P = 0.04). The mean TFPI activity measured by amidolytic assay was comparable for patients and controls (1.2 +/- 0.3 and 1.3 +/- 0.5 U/ml, respectively). However, the dilute TF clotting time was 115 +/- 26 s in patients, against 99 +/- 10 s in controls (P < 0.0001, irrespective of age adjustment). Since none of the patients had received heparin or had coagulation factor deficiency that may interfere with the assay, prolongation of clotting time may be attributed to the presence of TFPI, particularly the full-length form. To verify this inference, 33 extra aliquots left over from 88 samples (62.5%), 21 from controls and 12 from patients, were incubated with 1:10 diluted antibody against the C-terminal peptide of TFPI prior to dilute TF assay. The mean clotting time of both patients and controls decreased, and the between-group difference leveled (90 +/- 10 versus 88 +/- 20 s for controls and patients, respectively; P = 0.841). The mean drop in clotting time was 9% for the controls and 24% for the patients. This illustrates the specificity of dilute TF assay for full-length TFPI and supports the conclusion that relative to lipoprotein-associated TFPI, the proportion of the full-length form was possibly greater in patients with CAD. Contribution of lipoprotein associated TFPI to the overall anticoagulant activity by its activated factor X dependent inhibition of activated factor VII-TF complex seems less important considering the similar between-group mean amidolytic activities. PMID- 10870810 TI - Coexisting dysfibrinogenemia (gammaR275C) and factor V Leiden deficiency associated with thromboembolic disease (fibrinogen Cedar Rapids). AB - Fibrinogen Cedar Rapids is a heterozygous dysfibrinogenemia (gammaR275C) that was associated with thromboembolism during and following pregnancy in three second generation family members who also were heterozygotic for factor V Leiden (V R506Q). Like other dysfibrinogenemias with substitutions at position 275 of the gamma-chain, fibrinogen Cedar Rapids is characterized by defective end-to-end intermolecular fibrinogen and fibrin 'D : D' associations, a fibrin network structure that is composed of thicker and more highly branched fibers, normal fibrin 'D: E' associations, and normal factor XIII-mediated crosslinking of fibrinogen and fibrin. In addition, Cedar Rapids fibrinogen and fibrin displayed delayed plasmin lysis rates. Compared with normal fibrinogen, platelet aggregation or platelet fibrinogen receptor clustering was defective in the presence of fibrinogen Cedar Rapids. Most subjects with gammaR275 mutations do not experience clinical thrombotic disorders, suggesting that the combination of a factor V Leiden defect and a gammaR275C dysfibrinogenemia predisposes to thromboembolic disease. PMID- 10870811 TI - Inability of serial fibrin monomer measurements to predict or exclude deep venous thrombosis in asymptomatic patients undergoing total knee arthroplasty. AB - Fibrin monomer (FM) is a highly sensitive marker of venous thromboembolism and can be used to rule out deep venous thrombosis (DVT) and/or pulmonary embolism in symptomatic outpatients. The aim of the study was to investigate the usefulness of serial fibrin monomer determinations to predict or rule out DVT after total knee arthroplasty in asymptomatic patients. One hundred and thirty consecutive patients underwent total knee replacement. Blood samples were obtained in 104 of them the day before, at days 1, 3, 6 after surgery and at the day of phlebography. Phlebography was performed in all these patients between days 8 and 12 after surgery. There were 44 DVT (44/104, 42%). As compared with the patients without DVT, FM mean levels were 2 and 1.5 times higher in the DVT group at day 3 (P < 0.001) and day 6 (P < 0.01), respectively. However, no useful cut-off values for DVT prediction or exclusion could be determined due to the scattering of the values. Therefore, despite differences between patients with or without DVT, serial FM determinations are of no value for predicting or ruling out DVT in individual patients undergoing total knee arthroplasty. PMID- 10870812 TI - Severe acquired vitamin K deficiency: a hypothesis for rapid response to therapy. AB - The potential mechanism underlying the rapid response to vitamin K replacement in acquired deficiency states is incompletely understood. To examine vitamin K metabolism, a 10-year-old boy with autoimmune enteropathy on oral vitamin K supplementation, who presented with profuse nosebleeds and calf tenderness, was evaluated. Laboratory analyses were consistent with severe vitamin K deficiency: vitamin K dependent protein (VKDP) levels < 5%, normal vitamin K epoxide level and depressed total prothrombin antigen (carboxylated and undercarboxyated forms). Intramuscular vitamin K (10 mg) was administered. Nine hours following therapy, VKDP levels corrected completely. Total prothrombin antigen increased indicating new prothrombin synthesis. However, the increase in the prothrombin clotting assay far exceeded the increase in total prothrombin, supporting storage of undercarboxylated prothrombin in vitamin K deficiency states, with carboxylation and secretion after vitamin K replacement. Although this mechanism is known to occur in rodents, it has not been reported in humans. Our findings suggest a new potential mechanism of prothrombin metabolism in humans. PMID- 10870813 TI - June 2000: microgravity effects in space. PMID- 10870814 TI - Design and current status of the longitudinal study of astronaut health. AB - BACKGROUND: Information has been collected regarding the immediate physiological effects of spaceflight on humans. However, little is yet known regarding long term effects. The purpose of this paper is to describe the Longitudinal Study of Astronaut Health (LSAH) and report current mortality data. METHODS: All astronauts selected for the United States Space Program are followed from selection throughout their lifetime or until the end of the study. Comparisons are ground-based Johnson Space Center (JSC) employees matched to the astronauts at a 3:1 ratio by sex-specific age and body mass index. They are followed in the same manner as astronauts. Morbidity and mortality data are obtained from medical records supplemented with study questionnaires. Checks for death certificates are made to ascertain death of participants who miss routine examinations. RESULTS: Current cause-specific mortality rates for astronauts selected from 1959 through 1991 are not statistically different from rates for comparison participants for cardiovascular (p = 0.8112), cancer (p = 0.2382), or other disease (p = 0.5040) mortality. Astronauts have a significantly higher mortality rate due to accidents and injuries (p < 0.0001). CONCLUSIONS: Astronauts have a similar risk of death due to chronic diseases as ground-based participants, but are at greater risk for occupational-related accidental death. PMID- 10870815 TI - A tactile display for international space station (ISS) extravehicular activity (EVA). AB - BACKGROUND: A tactile display to increase an astronaut's situational awareness during an extravehicular activity (EVA) has been developed and ground tested. The Tactor Locator System (TLS) is a non-intrusive, intuitive display capable of conveying position and velocity information via a vibrotactile stimulus applied to the subject's neck and torso. In the Earth's 1 G environment, perception of position and velocity is determined by the body's individual sensory systems. Under normal sensory conditions, redundant information from these sensory systems provides humans with an accurate sense of their position and motion. However, altered environments, including exposure to weightlessness, can lead to conflicting visual and vestibular cues, resulting in decreased situational awareness. The TLS was designed to provide somatosensory cues to complement the visual system during EVA operations. METHODS: An EVA task was simulated on a computer graphics workstation with a display of the International Space Station (ISS) and a target astronaut at an unknown location. Subjects were required to move about the ISS and acquire the target astronaut using either an auditory cue at the outset, or the TLS. Subjects used a 6 degree of freedom input device to command translational and rotational motion. The TLS was configured to act as a position aid, providing target direction information to the subject through a localized stimulus. RESULTS: Results show that the TLS decreases reaction time (p = 0.001) and movement time (p = 0.001) for simulated subject (astronaut) motion around the ISS. CONCLUSION: The TLS is a useful aid in increasing an astronaut's situational awareness, and warrants further testing to explore other uses, tasks and configurations. PMID- 10870816 TI - Thermoregulation and heat exchange in a nonuniform thermal environment during simulated extended EVA. Extravehicular activities. AB - BACKGROUND: Nonuniform heating and cooling of the body, a possibility during extended duration extravehicular activities (EVA), was studied by means of a specially designed water circulating garment that independently heated or cooled the right and left sides of the body. The purpose was to assess whether there was a generalized reaction on the finger in extreme contradictory temperatures on the body surface, as a potential heat status controller. METHOD: Eight subjects, six men and two women, were studied while wearing a sagittally divided experimental garment with hands exposed in the following conditions: Stage 1 baseline--total body garment inlet water temperature at 33 degrees C; Stage 2--left side inlet water temperature heated to 45 degrees C; right side cooled to 8 degrees C; Stage 3--left side inlet water temperature cooled to 8 degrees C, right side heated to 45 degrees C. RESULTS: Temperatures on each side of the body surface as well as ear canal temperature (Tec) showed statistically significant Stage x Side interactions, demonstrating responsiveness to the thermal manipulations. Right and left finger temperatures (Tfing) were not significantly different across stages; their dynamic across time was similar. Rectal temperature (Tre) was not reactive to prevailing cold on the body surface, and therefore not informative. Subjective perception of heat and cold on the left and right sides of the body was consistent with actual temperature manipulations. CONCLUSIONS: Tec and Tre estimates of internal temperature do not provide accurate data for evaluating overall thermal status in nonuniform thermal conditions on the body surface. The use of Tfing has significant potential in providing more accurate information on thermal status and as a feedback method for more precise thermal regulation of the astronaut within the EVA space suit. PMID- 10870817 TI - Bone loss during simulated weightlessness: a biomechanical and mineralization study in the rat model. AB - BACKGROUND: Astronauts exposed to weightlessness for extended periods experience significant decreases in bone mineral density. The clinical implications of this demineralization are not entirely clear, and the biomechanics involved are not completely understood. HYPOTHESIS: Local (rather than global) measurements of geometry and calcium concentration effectively predict femur strength in adult rats exposed to a hind-limb suspension model of weightlessness. METHODS: Female Fischer rats (6-mo-old) were divided into groups of control and hind-limb suspended animals. Animals were sacrificed after 2 or 4 wk of hind-limb suspension, and both femurs removed from each animal. The 3-point bending strength and total bone mineralization were determined for one femur from each animal, and the mid-shaft cross-sectional geometrical properties and distribution of calcium were determined for the contralateral femur. RESULTS: Although suspension led to significant decreases in total bone mineralization, the concentration of calcium at the anterior periosteal surface was unaffected. Total bone percent mineralization was not well correlated with structural properties, but bone geometrical properties (particularly cross-sectional moment of inertia and length) correlated strongly with ultimate bending strength (r2 = 0.81). Differences in bone geometry due to suspension were consistent with a distribution of bone material closer to the axis of the femur. CONCLUSIONS: Structural properties of bone are predicted well by bone geometry and poorly by total bone percent mineralization. Decreased bone mechanical strength in this model of weightlessness is primarily due to a distribution of bone material nearer the axis of the bone. PMID- 10870818 TI - Aerobic exercise as a countermeasure for microgravity-induced bone loss and muscle atrophy in a rat hindlimb suspension model. AB - BACKGROUND: Loss of bone and skeletal muscle atrophy resulting from non-weight bearing are major concerns associated with microgravity environment and spaceflight deconditioning. The objective of this research was to address the fundamental issue of whether bone loss and muscle atrophy could be attenuated using weight-bearing aerobic exercise on a treadmill as a countermeasure in rats subjected to simulated weightlessness by hindlimb suspension. METHOD: Bone and muscle from control and hindlimb-suspended groups with and without exercise were evaluated by bone mineral density (BMD), mechanical tests, bone histomorphometry and muscle mass. RESULTS: Femoral BMD of hindlimb-suspended (HS) rats subjected to treadmill exercise was significantly greater than femoral BMD of HS rats without exercise and also was equivalent to that of weight-bearing controls. Muscle mass from HS rats exercised on a treadmill was significantly greater than muscle mass from HS rats that did not exercise. Exercise did not result in muscle mass equal to that of controls, however. In addition, histomorphometric analysis of the metaphysis of the proximal tibia revealed that HS rats that exercised did not maintain bone formation equivalent to controls. No other bone parameters were found to vary significantly between groups. CONCLUSIONS: It was concluded that moderate aerobic exercise on a treadmill did attenuate bone loss and muscle atrophy due to simulated weightlessness by hindlimb suspension, however its effectiveness differed by tissue, anatomical site and parameter investigated. PMID- 10870819 TI - Rat growth, body composition, and renal function during 30 days increased ambient CO2 exposure. AB - BACKGROUND: In experiments using rodents onboard orbiting spacecraft, specimens may be exposed to an increase in ambient CO2. HYPOTHESIS: Many of the physiological changes reported in rats (and humans) for spaceflight are similar to those observed with increased CO2, raising the question whether the observed changes are due to spaceflight or more specifically, the elevated ambient CO2. METHODS: To evaluate the effects of increased CO2, at levels similar to those experienced during spaceflight, three groups of adult male rats (n = 10) were exposed to ambient CO2 concentrations of 0.3, 0.7 and 2.0% for 30 d. Control rats were exposed to atmospheric conditions (0.03% CO2) for each group. RESULTS: There were alterations in water turnover, food intake, and renal function with increased CO2. Blood pH, total CO2, and plasma concentrations of Na+, Ca2+, and corticosterone were significantly elevated at the 2.0% exposure, while plasma PO4(3-) was reduced. At the 0.3% and 0.7% CO2 exposures, many of these changes were not significant. Animals exposed to 0.3% CO2 showed a significant increase in total body Na+. Urinary Ca2+, K+, creatinine, corticosterone, and total CO2 excretion were higher at 2.0%, but only Ca2+ and CO2 excretion were significantly elevated at 0.7%, and there was no significant alteration in renal function at 0.3%. CONCLUSION: Chronic increased ambient CO2 levels, similar to those observed on the Space Shuttle and proposed for the International Space Station, elicit compensatory responses in rats which may affect interpretation of experiments designed to evaluate the effects of exposure to microgravity. PMID- 10870820 TI - Effects of hindlimb suspension and elevated ambient CO2 on rat growth and renal function. AB - BACKGROUND: Previous studies show that an ambient CO2 concentration of 2.0% may complicate interpretation of animal experiments conducted in spaceflight, while 0.7% CO2 exposure produces minimal effects. HYPOTHESIS: With additional spaceflight factors, such as microgravity, effects from the 0.7% CO2 exposure may be amplified. METHODS: To investigate the combined effects from microgravity and elevated CO2 on growth and renal function, two groups of rats were hindlimb suspended for 37 d, and a third group served as an ambulatory vivarium control (AMB). One suspension group was exposed to 0.7% CO2 for 30 d (HLS + 0.7% CO2), while the other group (HLS) served as a suspended control. Both the AMB and HLS groups breathed room air at 0.03% CO2. RESULTS: The HLS group showed responses consistent with past hindlimb suspension studies when compared with AMB, indicating similar reductions in organ and tissue weights and body weight gain. When comparing HLS + 0.7% CO2 animals to HLS controls, exposure to CO2 revealed lowered food consumption and increased urine volume, NH3 and CO2 excretion, with no differences in any of the other measured parameters, such as body weight gain, pH values, or electrolyte handling. CONCLUSION: This study shows that chronic exposure to both 0.7% ambient CO2 and hindlimb suspension together have little additional effect on rat growth and renal function. PMID- 10870821 TI - Predictors of behavior and performance in extreme environments: the Antarctic space analogue program. AB - BACKGROUND: To determine which, if any, characteristics should be incorporated into a select-in approach to screening personnel for long-duration spaceflight, we examined the influence of crewmember social/ demographic characteristics, personality traits, interpersonal needs, and characteristics of station physical environments on performance measures in 657 American men who spent an austral winter in Antarctica between 1963 and 1974. METHODS: During screening, subjects completed a Personal History Questionnaire which obtained information on social and demographic characteristics, the Deep Freeze Opinion Survey which assessed 5 different personality traits, and the Fundamental Interpersonal Relations Orientation-Behavior (FIRO-B) Scale which measured 6 dimensions of interpersonal needs. Station environment included measures of crew size and severity of physical environment. Performance was assessed on the basis of combined peer supervisor evaluations of overall performance, peer nominations of fellow crew members who made ideal winter-over candidates, and self-reported depressive symptoms. RESULTS: Social/demographic characteristics, personality traits, interpersonal needs, and characteristics of station environments collectively accounted for 9-17% of the variance in performance measures. The following characteristics were significant independent predictors of more than one performance measure: military service, low levels of neuroticism, extraversion and conscientiousness, and a low desire for affection from others. CONCLUSIONS: These results represent an important first step in the development of select-in criteria for personnel on long-duration missions in space and other extreme environments. These criteria must take into consideration the characteristics of the environment and the limitations they place on meeting needs for interpersonal relations and task performance, as well as the characteristics of the individuals and groups who live and work in these environments. PMID- 10870822 TI - Effects of gender on the autonomic modulation of the cardiovascular responses to lower body negative pressure. AB - BACKGROUND: The cardiovascular responses to submaximal lower body negative pressure (LBNP) appear to differ between genders, but the underlying mechanisms are uncertain. HYPOTHESIS: These differences are due to differences in the autonomic modulation of the cardiovascular system. METHODS: There were 14 women and 13 men who underwent LBNP to -50 mmHg in 10 mmHg increments of 6 min each. Heart rate (HR), stroke volume (SV), BP, forearm blood flow and R-R interval data were acquired. Spectral analysis of the R-R interval data was used to assess autonomic modulation with the low frequency component (LF) set at 0.04 to 0.15 Hz and the high frequency component (HF) at 0.15 to 0.4 Hz. RESULTS: The responses to LBNP to -40 mmHg did not differ between groups. LBNP of -50 mmHg evoked greater HR increases in the women than the men (7.2 +/- 1.0 vs. 3.8 +/- 1.1 bpm; p < 0.05), while SV, cardiac output and total peripheral conductance decreased more (-15 +/- 2 vs. -8 +/- 2 ml x beat(-1); -0.668 +/- 0.131 vs. -0.1778 +/- 0.124 L x min(-1); -0.009 +/- 0.002 vs. -0.004 +/- 0.001 units; p < 0.05). Normalized HF, an indicator of the vagal influence on HR variability, declined below rest at -40 mmHg while the LF/HF ratio, an indicator of sympathetic neural modulation of HR variability, increased above rest at -40 mmHg. These responses did not differ significantly between groups. CONCLUSIONS: These results suggest that gender differences in the cardiovascular responses to LBNP are not due to gross differences in modulation of the autonomic nervous systems. PMID- 10870823 TI - Continuous monitoring of change in hemodilution during water immersion in humans: effect of water temperature. AB - BACKGROUND: The present study was designed to examine whether water temperature during head-out immersion (HOI) modifies hemodilution dynamics. METHODS: We made continuous measurements of blood density (rho(b)) during HOI at 3 different water temperatures; the lower critical (32 degrees C), neutral (34.5 degrees C), and upper critical (36 degrees C) temperatures in 6 healthy male volunteers. Blood was withdrawn continuously from the antecubital vein for measurement of rho(b) during 60 min of water immersion with a 10-min control period before the immersion. The density was measured with the mechanical oscillator technique. Hematocrit (Hct), plasma density (rho(p)), and osmolality were measured at 5-min intervals. Erythrocyte density (rho(e)) was calculated from rho(b), rho(p) and Hct. Cardiac output and BP were measured to calculate total peripheral resistance. RESULTS: Hct, rho(b), and rho(p) decreased rapidly in the first 20-25 min of immersion and were maintained at a reduced level during immersion. Plasma volume calculated from rho(p) and Hct increased with the rho(b) reduction. These immersion-induced changes were independent of these water temperatures. Plasma osmolality and rho(e) remained constant throughout the experimental period in the three temperature conditions, indicating that the increase in plasma volume and hence hemodilution was induced by an isotonic fluid shift from extravascular space. The total peripheral resistance increased inversely in proportion to the water temperature during HOI. CONCLUSION: In the present condition, water temperature did not modify the net transcapillary fluid transfer during HOI in the presence of the temperature dependent increase in vascular tone. PMID- 10870824 TI - Medical imaging in microgravity. AB - Microgravity environment induces significant pathophysiological changes in humans. As long-term space habitation and interplanetary missions become commonplace in the next century, researchers and imaging specialists must embrace the unique challenges of microgravity research in developing innovative uses of imaging technology. Microgravity results in cephalad fluid shift, loss of electrolytes, loss of muscle and bone mass, anemia, reduced immune response, variability in gastric emptying and hepatic metabolism, increased bowel transit, and development of space motion sickness. Medical imaging is uniquely capable of assessing all of these physiologic adaptations to weightlessness. The imaging information will also aid in identifying those changes which are potentially beneficial and those which are harmful to humans living in space. Interventions may then be employed to capitalize on the benefits and prevent the harmful effects. This article reviews the physiological adaptations to weightlessness and outlines ways in which medical imaging techniques may provide insight into these adaptations. PMID- 10870825 TI - Galactic cosmic radiation exposure of pregnant flight crewmembers. AB - BACKGROUND: In recommending the occupational dose limit of ionizing radiation for pregnant women, the International Commission on Radiological Protection apparently assumes that the dose to the conceptus from ionizing radiation exposure is about half the dose at the surface of the mother's abdomen. METHODS: To test this assumption with respect to galactic cosmic radiation, calculations were made using FAA computer program CARI-LF2, which calculates equivalent doses from galactic cosmic rays at selected depths in soft tissue at any specified location in the atmosphere or on user-entered flight profiles. RESULTS: The calculations showed that the equivalent dose of galactic radiation was almost the same at all depths. CONCLUSIONS: Thus the assumption of considerable shielding of the conceptus being provided by the woman's body is not correct with respect to galactic cosmic radiation, the principal type of radiation to which aircrews are exposed. The effective dose as calculated with FAA computer program CARI-5E, which calculates effective dose in an anthropomorphic phantom at any specified location in the atmosphere or on user-entered flight profiles, was found to be a good estimate of the equivalent dose at the depth of the conceptus. PMID- 10870826 TI - The medical implications of space tourism. AB - Commercial space travel may soon be a reality. If so, microgravity, high acceleration, and radiation exposure, all known hazards, will be accessible to the general public. Therefore, space tourism has medical implications. Even though the first flights will feature space exposure times of only a few minutes, the potential may someday exist for exposure times long enough to warrant careful consideration of the potential hazards to the space-faring public. The effects of acceleration and microgravity exposure are well known on the corps of astronauts and cosmonauts. The effects of space radiation are partially known on astronauts, but much remains to be discovered. However, there are problems using astronaut data to make inferences about the general public. Astronauts are not necessarily representative of the general public, since they are highly fit, highly screened individuals. Astronaut data can tell us very little about the potential hazards of microgravity in pediatric, obstetric and geriatric populations, all of whom are potential space tourists. Key issues in standard setting will be determining acceptable limits of pre-existing disease and inferring medical standards from mission profiles. It will not be a trivial task drafting minimal medical standards for commercial space travel. It will require the collaboration of space medicine physicians, making the best guesses possible, based on limited amounts of data, with limited applicability. A helpful departure point may be the USAF Class 3 medical standard, applicable to NASA payload specialists. It is time to begin preliminary discussions toward defining those standards. KEYWORDS: acceleration, aerospace medicine, medical standards, microgravity, radiation, space, space tourism, environmental hazards, environmental medicine. PMID- 10870827 TI - Chest-mounted left ventricular nuclear monitoring technologies. PMID- 10870828 TI - Development and application of surface plasmon resonance-based biosensors for the detection of cell-ligand interactions. AB - Surface plasmon resonance (SPR)-based biosensors were investigated with a view to providing a portable, inexpensive alternative to existing technologies for "real time" biomolecular interaction analysis of whole cell-ligand interactions. A fiber optic SPR-based (FOSPR) biosensor, employing wavelength-dependent SPR, was constructed to enable continuous real-time data acquisition. In addition, a commercially available integrated angle-dependent SPR-based refractometer (ISPR) was modified to facilitate biosensing applications. Solid-phase detection of whole red blood cells (RBCs) using affinity-captured blood group specific antibodies was demonstrated using the BIACORE 1000, BIACORE Probe, FOSPR, and ISPR sensors. Nonspecific binding of RBCs to the hydrogel-based biointerface was negligible. However, the background noise level of the FOSPR-based biosensor was approximately 25-fold higher than that of the widely used BIACORE 1000 system while that of the ISPR-based biosensor was over 100-fold higher. Nevertheless, the FOSPR biosensor was suitable for the analysis of macromolecular analytes contained in crude matrices. PMID- 10870829 TI - Quantitative determination of zwitterionic detergents using salt-induced phase separation of Triton X-100. AB - Zwitterionic detergents interfere with the salt-induced phase separation for nonionic detergents in a concentration-dependent manner by shifting the normal cloud point of nonionic detergents to a higher ionic strength at room temperature. This phenomenon was used to determine the concentration of the zwitterionic detergents CHAPS, CHAPSO, and sulfobetaine SB-12 in solution by titration with ammonium sulfate in the presence of Triton X-100. Among the ionic detergents tested, the method was only applicable to sodium cholate. The assay can be used to control the removal of zwitterionic detergents during the reconstitution of membrane proteins in liposomes. However, it cannot be used to determine the specific binding of zwitterionic detergents to highly diluted, pure membrane proteins because of the limited sensitivity. Neither proteins nor phospholipids interfered with this method at concentrations up to 20 mg/ml of test solution (human serum albumin) or 10 mg/ml (phospholipids), respectively. Since the assay is based on the competition between salts and nonionic detergents for water molecules, it is important to equalize the ionic strength of samples and calibration standards. PMID- 10870830 TI - Analysis of carbohydrates using liquid chromatography--surface plasmon resonance immunosensing systems. AB - An immunosensing system based on surface plasmon resonance (SPR) was used for on line detection and characterization of carbohydrate molecules separated by high performance liquid chromatography. These analytes, with or without serum, were continuously separated and analyzed in the combined liquid chromatography-surface plasmon resonance (LC-SPR) system. By using weak and readily reversible monoclonal antibodies, the SPR system allowed specific on-line monitoring of the substances. To increase the specificity of the immunosensor, nonrelevant antibodies were used as reference in a serial flow cell. The sensitivity of the LC-SPR system was dependent on molecular weight of the carbohydrate, affinity of binding, and design of the sensor. PMID- 10870831 TI - Fluorescence lifetime microscopy of the Na+ indicator Sodium Green in HeLa cells. AB - This study investigates the usefulness of lifetime measurements of Sodium Green for evaluating intracellular Na+ concentration ([Na+]i) in HeLa cells. Frequency domain lifetime measurements are performed in HeLa cells and in different buffer solutions (with and without K+ and bovine serum albumin). In all cases, the fluorescence decays of Sodium Green are multiexponential, with decay times independent of [Na+]. Three relaxation times are found in the various buffer solutions. Binding of the indicator to albumin results in an increase in the long and intermediate decay times. For Sodium Green inside HeLa cells, the intensity decay can be approximated by a biexponential. The ratio of the fractional intensity of the long decay time (tau2 = 2.4 +/- 0.2 ns) to that of the short component (tau1 = 0.4 +/- 0.1 ns) increases with [Na+]i. The changes in fluorescence decay with [Na+] are significantly less pronounced in cells as compared with the buffer solutions. Similar values for the resting [Na+]i were estimated from lifetime measurements of Sodium Green and from ratiometric measurements using SBFI. Alternatively, [Na+]i can be monitored by measuring only the phase angle at the modulation frequency of 160 MHz. The usefulness of this latter approach is demonstrated by following the changes in [Na+]i induced by reversible inhibition of the Na+/K+ pump. PMID- 10870832 TI - Determination of L-carnitine by flow injection analysis with NADH fluorescence detection. AB - A flow injection analysis method for determining L-carnitine is reported. The system uses the enzyme L-carnitine dehydrogenase covalently immobilized to Eupergit C. The NADH produced by the action of the enzyme, which is proportional to the L-carnitine concentration, is quantified using fluorescence detection. The system response was rapid and had a wide range of linearity. At a flow rate of 0.2 ml/min, a detection limit of 1 microM (20 pmol) was obtained for L-carnitine, peak areas were linear up to 100 microM, and samples could be injected every 4 min. The method performed well as a routine assay, showing high sensitivity (54,000 AU/microM), a precision of 0.96%, and the ability to carry out 144 consecutive assays with an RSD of 1.47% (good stability). Comparisons were made with other known methods for L-carnitine determination. Presence of D-carnitine had no effect on L-carnitine assay. The analysis was valid for determining L carnitine concentrations in commercial pharmaceutical preparations. PMID- 10870833 TI - A high-throughput homogeneous assay for reverse transcriptase using generic reagents and time-resolved fluorescence detection. AB - A homogeneous time-resolved fluorescence (HTRF) assay has been developed for determining the activity of HIV reverse transcriptase (HIV-RT). By using a sequential capping strategy, the assay has been configured to utilize only generic reagents such as biotinylated dUTP, streptavidin-allophycocyanin, and streptavidin-europium. The assay was optimized for a HIV-RT high-throughput screen. Under optimized conditions, a signal-to-background ratio of approximately 10:1 and a Z' factor of 0.8 were obtained. The titration curves of several known HIV-RT inhibitors have been evaluated with this assay format. This HTRF format can be used for high-throughput assays with other nucleotide polymerases. PMID- 10870834 TI - Use of a dual firefly and Renilla luciferase reporter gene assay to simultaneously determine drug selectivity at human corticotrophin releasing hormone 1 and 2 receptors. AB - The aim of this study was to investigate and validate the use of a dual glow signal luciferase reporter gene assay to simultaneously evaluate drug activity at two different seven-transmembrane receptor subtypes. Stable cell lines (CHO) transfected with either human corticotrophin releasing hormone 1 (hCRH1) receptors and a firefly luciferase reporter gene or hCRH2 and a Renilla luciferase reporter gene were created to provide different luciferase readouts for CRH1 and CRH2 receptors, respectively. Cells were combined for stimulation and measurement of luciferase luminescence in a 96-well plate format assay. The nonselective CRH agonists rat/human CRH and sauvagine caused concentration dependent increases in luminescence via activation of CRH1 (firefly luciferase; pEC50 = 8.40 +/- 0.06 and 8.39 +/- 0.08, respectively, n = 8) and CRH2 (Renilla luciferase; pEC50 = 8.89 +/- 0.14 and 8.92 +/- 0.13, respectively, n = 8) receptors. The nonselective CRH antagonist astressin blocked these agonist induced increases in luciferase at both CRH1 and CRH2 receptors. The selective CRH1 antagonist CP154,526 blocked r/hCRH- and sauvagine-induced increases in luciferase at CRH1 receptors only. These data report the expected pharmacology for CRH1 and CRH2 receptors. This assay enabled two receptor subtypes to be studied simultaneously in the same 96-well plate and generated robust data with low variability. It has the potential advantage of significant time and cost savings, with application to both basic research and compound screening. PMID- 10870835 TI - Determination of N-acetyl- and N-glycolylneuraminic acids in gangliosides by combination of neuraminidase hydrolysis and fluorometric high-performance liquid chromatography using a GM3 derivative as an internal standard. AB - A highly sensitive method for quantification of sialic acids in gangliosides was developed. The sialic acids, released by hydrolysis of gangliosides, were converted to fluorescent derivatives with 1,2-diamino-4,5-(methylenedioxy)benzene (DMB) and separated on a reversed-phase C18 column with an isocratic elution. As little as 0.1-1.0 nmol of sialic acid in ganglioside was quantified. The use of acetate buffer instead of water in the mobile phase could prevent damage on the column and reduce background peaks derived from the reagents. When gangliosides were subjected to acid hydrolysis, the velocity of hydrolysis varied depending on their structures and a part of the sialic acid liberated decomposed with prolonged heating time. Therefore gangliosides were hydrolyzed by Arthrobacter ureafaciens neuraminidase in the presence of sodium cholate after addition of an internal standard. For the internal standard, GM3 with N-propionylneuraminic acid (GM3(NeuPr)) was synthesized from GM3(NeuAc) by N-deacylation followed by N propionylation. Folch partition was used to decrease lipophilic materials included in the sample, and the sialic acids released were recovered from the upper phase. The present method has a satisfactory sensitivity in the simultaneous quantification of NeuAc and NeuGc in purified gangliosides as well as in crude lipid fractions containing a variety of gangliosides. PMID- 10870836 TI - Binding of avian ovomucoid to shiga-like toxin type 1 and its utilization for receptor analog affinity chromatography. AB - Development of a simple and efficient purification procedure for Shiga-like toxin I (Stx1) was attempted. Since it has been suggested that pigeon egg white ovomucoid carries a P1 antigenic determinant, we examined its ability to bind Stx1. The ovomucoid glycoprotein fraction (GPro) was prepared from pigeon egg white by acetone precipitation, and a portion of the GPro was treated with pronase to obtain the glycopeptide fraction (GPep). When both GPro and GPep were coupled to CNBr-activated Sepharose 4B and subjected to affinity chromatography, Stx1 specifically bound to both columns. The Stx1 eluted with a buffer containing 4.5 M MgCl2 was shown to be highly purified to homogeneity by polyacrylamide gel electrophoresis under denatured condition; only two protein bands with molecular weights of 32,000 and 8000, which correspond to the A and the B subunits of Stx1, respectively, were recognized. The purified toxin showed cytotoxicity on Vero cells with a specific activity of approximately 6 x 10(8) CD50/mg protein; almost 100% of the activity was recovered from Escherichia coli cell lysate. We propose that the utilization of avian ovmucoid for the affinity chromatography provides a potentially simple, convenient, and widely available method to purify Shiga-like toxins. PMID- 10870837 TI - Occurrence and determination of a natriuretic hormone, 2,7,8-trimethyl-2-(beta carboxyethyl)-6-hydroxy chroman, in rat plasma, urine, and bile. AB - The occurrence of a new natriuretic hormone, 2,7,8-trimethyl-2-(beta carboxyethyl)-6-hydroxy chroman (LLU-alpha, gamma-CEHC) in rat plasma was demonstrated and its concentration was determined using a coupled-column HPLC with a fluorometric derivatization with 4-N,N-dimethylaminosulfonyl-7-piperazino 2,1,3-benzoxadiazol e (DBD-PZ) followed by O-acetylation. The concentration of LLU-alpha was 328 +/- 113 nM in rat plasma (N = 5). LLU-alpha was found in not only urine, but also bile, suggesting an enterohepatic circulation in body. We also assigned the configuration at C-2 of LLU-alpha in these biological fluids as (S)-form by an HPLC with a chiral column. The LLU-alpha concentration decreased significantly by fasting for 3 days (P < 0.01). These results support the possibility that LLU-alpha is produced from gamma-tocopherol in diet via oxidative metabolism without racemization. PMID- 10870838 TI - Real-time fluorescence assay for O6-alkylguanine-DNA alkyltransferase. AB - O6-alkylguanine-DNA alkyltransferase (AGT) is a DNA-repair protein that reverses the effects of alkylating agents by removing DNA adducts from the O6-position of guanine. We developed a real-time AGT assay that utilizes a fluorescent guanosine analog (3-methylisoxantopterin, 3-MI). 3-MI fluorescence is quenched in DNA and fluorescence intensity increases substantially with digestion of the oligonucleotide and release of 3-MI. The substrate is a doubled-stranded oligonucleotide with 3'-overhangs on each end and a PvuII recognition site. PvuII is inhibited by O6-methylguanine, positioned within the restriction site. 3-MI is incorporated in the opposite strand just outside of the PvuII restriction site. AGT repairs O6-methylguanine; PvuII cleaves at its restriction site, yielding a blunt-ended double strand, which is then digested by exonuclease III. This releases 3-MI from the oligonucleotide, resulting in an increase in fluorescence intensity. All reaction components (100-microL volume) are monitored in a single microcuvette. Rate of increase in fluorescence intensity is related to the amount of AGT in the reaction mixture. We measured AGT levels in extracts from a leukemia cell line, from leukemic lymphoblasts from patients, and from peripheral blood mononuclear cells from normal controls. This method may prove useful for mechanistic studies of AGT. PMID- 10870839 TI - A spectrophotometric method for the direct detection and quantitation of nitric oxide, nitrite, and nitrate in cell culture media. AB - A method for the spectrophotometric determination of nitric oxide, nitrite, and nitrate in tissue culture media is presented. The method is based on the nitric oxide-mediated nitrosative modification of sulfanilic acid that reacts with N-(1 naphthyl)ethylenediamine dihydrochloride forming an orange-colored product absorbing at 496 nm. Nitric oxide levels were determined in culture media from this absorbance measurement using chemiluminescence standardization. Extinction coefficients of 5400 and 6600 M(-1) cm(-1) were determined for the nitric oxide product in assay solutions containing 0.1 or 100 mM KPO4 buffer (pH 7.4), respectively, with a limit of detection of 1 microM. Acidification of these reactions (pH 2.4) generated a pink-colored product absorbing at 540 nm allowing for quantitation of total nitric oxide/nitrite levels using extinction coefficients of 38,000 and 36,900 M(-1) cm(-1), for the assay solutions described. The limit of detection of this assay was approximately 300 nM. Using the 100 mM KPO4 buffer system, nitrate levels were determined following reduction to nitrite using a copper-coated cadmium reagent with an extinction coefficient of 29,500 M(-1) cm(-1) and a detection limit of 0.5 microM. The utility of these assays was demonstrated in the standardization of nitric oxide-saturated cell culture media, and the release of nitric oxide by the NONOate compound DEA/NO. PMID- 10870840 TI - Immunopolymerase chain reaction using real-time polymerase chain reaction for detection. PMID- 10870841 TI - High-yield expression and purification of recombinant proteins in bacteria: a versatile vector for glutathione S-transferase fusion proteins containing two protease cleavage sites. PMID- 10870842 TI - Zymogram of pancreatic lipases. PMID- 10870843 TI - Two fibrin zymography methods for analysis of plasminogen activators on gels. PMID- 10870845 TI - Enzyme replacement therapy in mucopolysaccharidosis I: altered distribution and targeting of alpha-L-iduronidase in immunized rats. AB - Enzyme replacement therapy (ERT) has been developed and trialed for the treatment of human lysosomal storage disorder patients. The viability of ERT for the treatment of these severe multiple pathology disorders has subsequently been established. However, in both animal model studies and human clinical trials, some individuals have been shown to develop an immune response to the replacement protein. This potential complication for treatment has been investigated by the infusion of recombinant human alpha-L-iduronidase (rh-alpha-L-iduronidase) into nonimmune and immunized rats to simulate mucopolysaccharidosis type I ERT in the presence of different levels of antibody. In rats with high antibody titers to rh alpha-L-iduronidase (titer 1,024,000) there was evidence of altered organ distribution and subcellular targeting when compared to either lower titer immunized rats (titers less than 64,000) or nonimmune rats (titers 512-1024). In addition, hypersensitivity reactions were observed for high titer rats (titer 1,024,000) during rh-alpha-L-iduronidase infusion, but not for the other two treatment groups. A rat with an antibody titer of 64,000 had only minor changes in subcellular targeting and organ distribution when infused with rh-alpha-L iduronidase. This implied that a high level of antibody was required to effect changes in alpha-L-iduronidase enzyme targeting and distribution. Notably, in the high titer rats, the antibody produced appeared to increase the tissue and subcellular level of rh-alpha-L-iduronidase specific activity. This suggested that antibody production may not always result in an adverse effect on ERT. PMID- 10870844 TI - Adenovirus-mediated in utero gene transfer in mice and guinea pigs: tissue distribution of recombinant adenovirus determined by quantitative TaqMan polymerase chain reaction assay. AB - Fetal somatic cell gene therapy could become an attractive solution for some congenital genetic diseases or the disorders which manifest themselves during the fetal period. We performed adenovirus-mediated gene transfer to mice and guinea pig fetuses in utero and evaluated the efficiency of gene transfer by histochemical analysis and a quantitative TaqMan-polymerase chain reaction (TaqMan-PCR) assay. We first injected a replication-deficient recombinant adenovirus containing the Escherichia coli LacZ gene driven by a CAG promoter (AxCALacZ) into pregnant mice through the amniotic space, placenta, or intraperitoneal space of the fetus. Histochemical analysis showed limited transgene expression in fetal tissues. We then administered AxCALacZ to guinea pig fetuses in the late stage of pregnancy through the umbilical vein. The highest beta-galactosidase expression was observed in liver followed by moderate expression in heart, spleen, and adrenal gland. The transgene expression was also present in kidney, intestine, and placenta to a lesser degree. No positively stained cells were observed in lung, muscle, or pancreas except in the vascular endothelium of these organs. Quantitative measurement of recombinant adenoviral DNA by the TaqMan-PCR assay showed that the vast majority of the injected viruses was present in liver. The current study indicated that adenovirus-mediated gene transfer into guinea pig fetus through the umbilical vein is feasible and results in efficient transgene expression in fetal tissues. The experimental procedures using pregnant guinea pigs might serve as a good experimental model for in utero gene transfer. Since our TaqMan-PCR assay detects the LacZ gene, one of the most widely used reporter genes, it may be generally applicable to adenovirus quantification in various gene transfer experiments. PMID- 10870846 TI - "Hypotyrosinemia" in phenylketonuria. AB - It has been postulated that the significant incidence of learning disabilities in well-treated patients with phenylketonuria (PKU) may be due, in part, to reduced production of neurotransmitters as a result of deficient tyrosine transport across the neuronal cell membrane. Hypotyrosinemia has been reported in treated and untreated PKU but virtually no data are available. We decided to examine this in our patient population and to compare it with the published norms, patient data from our hospital clinical biochemical laboratory database, and a group of normal children and adolescents in a private pediatric practice. We found that the mean nonfasting plasma tyrosine in 99 classical PKU patients was 41.1 micromol/L, in 26 mild (atypical) PKU patients 53.3 micromol/L, and in 35 non-PKU mild hyperphenylalaninemia patients 66.6 micromol/L. This compared to nonfasting plasma tyrosine levels in 102 non-PKU subjects of 64.0 micromol/L in our hospital biochemistry database, 69.1 micromol/L in 58 volunteers in the private office practice, and 64-78.8 micromol/L in infants, children, and adolescents in the literature review. Our data support the previously undocumented statements in the literature that plasma tyrosine levels are low in PKU. PMID- 10870847 TI - Relation between HFE mutations and mild iron-overload expression. AB - The identification of the HFE gene involved in hemochromatosis allows genetic tests based on mutation analysis to be performed. However, discrepancies in the correlation between HFE genotypes and iron-loading status have arisen. We investigated 708 patients with various signs or symptoms suggesting a putative iron overload that, nevertheless, did not reach the current criteria for hemochromatosis diagnosis. Most of the patients (91.4%) included in our study displayed one of three classical iron marker values above the threshold defined for iron overloading. HFE mutation analysis allowed us to identify 45.7% of carrier chromosomes in the studied group of patients that showed higher frequencies of HFE mutations compared with controls. In addition, the frequencies of compound C282Y/H63D heterozygous, H63D/H63D homozygous, and C282Y heterozygous genotypes were higher than those in HH probands and controls; they accounted for 16, 5.6, and 22.5% of the patients, respectively. All genotypic groups had a significantly higher value of serum ferritin concentration compared to the normal value; only the C282Y homozygotes and compound heterozygotes with H63D had a transferrin saturation significantly higher than the normal value. On the whole the H63D homozygous and compound heterozygous patients constitute an intermediate phenotypic group between HH and controls. Some of them may reach the critical overloading defined for HH diagnosis along with a potential risk of developing complications, whereas others only show a partial phenotypic expression. PMID- 10870848 TI - Evaluation of urinary acylglycines by electrospray tandem mass spectrometry in mitochondrial energy metabolism defects and organic acidurias. AB - We analyzed the urinary acylglycine excretion in 26 patients with mitochondrial energy metabolism disorders and in 55 patients with organic acidurias by electrospray tandem mass spectrometry (ESI-MS/MS), monitoring precursor ions of m/z 90. Urinary concentrations of the different acylglycines were quantified using deuterated internal standards. Normal values for the most important acylglycines were established. In MCAD and MAD (neonatal form) deficiencies, typical excretion patterns of urinary acylglycines were found in all the samples. In isovaleric aciduria, propionic aciduria, and 3-methylcrotonylglycinuria typical glycine conjugates were always found. Methylmalonic aciduria (mutase deficiency), multiple carboxylase deficiency, and 3-hydroxy-3-methylglutaric aciduria revealed pathological acylglycine profiles, even if not specific for the disease. In all these diseases acylglycine excretion seems to be less influenced by the clinical status than organic acid excretion. This method is a useful diagnostic tool for these metabolic disorders, complementary to organic acids and acylcarnitine profiles. PMID- 10870849 TI - Lovastatin therapy for X-linked adrenoleukodystrophy: clinical and biochemical observations on 12 patients. AB - X-linked adrenoleukodystrophy (X-ALD) is a progressive demyelinating disorder whose neurological signs and symptoms can manifest in childhood as cerebral ALD or in adulthood in the form of a progressive myelopathy (AMN). The consistent metabolic abnormality in all forms of X-ALD is an inherited defect in the peroxisomal beta-oxidation of very long chain (VLC) fatty acids (>C(22:0)) which may in turn lead to a neuroinflammatory process associated with demyelination of the cerebral white matter. The current treatment for X-ALD with Lorenzo's oil aims to lower the excessive quantities of VLC fatty acids that accumulate in the patients' plasma and tissues, but does not directly address the inflammatory process in X-ALD. We have previously demonstrated that lovastatin and other 3-HMG CoA reductase inhibitors are capable of normalizing VLC fatty acid levels in primary skin fibroblasts derived from X-ALD patients. Lovastatin can block the induction of inducible nitric oxide synthase and proinflammatory cytokines in astrocytes, microglia, and macrophages in vitro. In a preliminary report, we demonstrated that lovastatin therapy can normalize VLC fatty acids in the plasma of patients with X-ALD. Here we report our clinical and biochemical observations on 12 patients with X-ALD who were treated with lovastatin for up to 12 months. Our results show that the high plasma levels of hexacosanoic acid (C(26:0)) showed a decline from pretreatment values within 1 to 3 months of starting therapy with 40 mg of lovastatin per day and stabilized at various levels during a period of observation up to 12 months. The percentage decline from pretreatment values varied and did not correlate with the type of ALD gene mutation (point mutation versus gene deletion). In 6 patients, in whom red cell membrane fatty acid composition was studied, a mean correction of 50% of the excess C(26:0) was observed after 6 months of therapy suggesting sustained benefit. In a few patients who discontinued lovastatin therapy plasma C(26:0) levels reverted to pretreatment values suggesting a cause and effect relationship between these events. Two patients dropped out of the study claiming no clinical benefit, 1 was withdrawn due to adverse effects, and an adult patient with cerebral involvement died during the study. A 10-year-old boy with severe cerebral involvement showed worsening of his neurological status. All patients with AMN remained neurologically stable or showed modest subjective improvement. All patients who did not have Addison's disease at the time of enrollment maintained normal adrenal function throughout the study. The implications of our findings for developing an effective therapy for X-ALD are discussed. PMID- 10870850 TI - Homozygous variegate porphyria in South Africa: genotypic analysis in two cases. AB - Variegate porphyria is an autosomal dominant disorder of heme metabolism which results from decreased activity of the enzyme protoporphyrinogen oxidase. Clinically, the disease manifests postpubertally and is characterized by photocutaneous sensitivity and/or acute neurovisceral crises. However, in homozygous variegate porphyria, onset of the disease usually occurs in infancy with severe skin manifestations. The molecular basis of variegate porphyria in two severely affected probands in two South African families is described. Mutation detection included combined SSCP-heteroduplex analysis followed by direct sequencing. The unrelated probands both had the common R59W mutation while the other lesion was Y348C or R138P (both novel mutations), causing homozygous variegate porphyria. PMID- 10870851 TI - Studies on human porin XXII: cell membrane integrated human porin channels are involved in regulatory volume decrease (RVD) of HeLa cells. AB - Cell volume regulation receives increasing attention not only as the basis of regulatory volume increase or regulatory volume decrease (RVD) of cells in surroundings of changing osmolarity, but also appears to be relevant in cell proliferation, differentiation, and apoptosis. A central event in RVD is the opening of a volume-sensitive chloride/anion channel(s), and blocking this pathway would abolish RVD. This is shown here with monoclonal mouse anti-human type-1 porin antibodies, proving that porin is involved in this process. HeLa cells preincubated with these antibodies dramatically increase their volume within about 1 min after a hypotonic stimulus by 70 mM NaCl Ringer solution, but do not move back toward their starting volume, thus indicating abolished RVD. Corresponding effects are induced by the established anion channel inhibitor DIDS. Video camera monitoring of cell size over time was used as a direct and noninvasive approach. We had already accumulated evidence that plasmalemma integrated eukaryotic porin channels form chloride/anion channels in this cell compartment and that they are involved in cell volume regulation. Finally, the present data again demonstrate the suitability of our anti-porin antibodies in physiological studies. PMID- 10870852 TI - Fructose-1,6-diphosphatase deficiency and glyceroluria: one possible etiology for GIS. AB - Fructose-1,6-diphosphatase (FDPase) deficiency is characterized by episodes of lactic acidemia, hypoglycemia, and ketonuria. Liver biopsy and subsequent enzyme analysis most reliably make the diagnosis. Review of the literature reveals 85 cases. Glycerol intolerance syndrome (GIS) is less well defined. There are only a handful of cases reported. We describe a patient with FDPase deficiency and significant glyceroluria and propose that GIS may be caused by partial deficiency of FDPase. PMID- 10870853 TI - Impact of a 4-week treatment with prostaglandin E1 on health-related quality of life of patients with intermittent claudication. AB - Intermittent claudication impairs functional status and quality of life in many patients by limiting walking capacity. The aim of this study was to evaluate the effects of a 4-week treatment with prostaglandin E1 (PGE1), a drug inducing vasodilation and inhibiting platelet aggregation, on improving functional status and health-related quality of life in patients with disabling intermittent claudication. Forty-two untrained outpatients (37 men and five women, mean age 64 +/- 8 years) with intermittent claudication,and maximum walking distance (MWD) of at least 50 and no more than 200 m on treadmill test (5% slope, 3 km/hr) were randomized to 4 weeks of double-blind treatment either with 60 mcg PGE1 daily given IV in 250 mL saline over a period of 2 hours (21 patients) or placebo (250 mL saline, 21 patients). Treatment-free follow-up was completed 8 weeks after the final infusion. Pain free walking distance (PFWD), MWD, and questionnaire evaluation were determined at baseline, after the 4-week treatment period, and at the end of the 8 weeks of the treatment-free follow-up period. After 4 weeks of treatment with PGE1 PFWD and MWD increased from 72 +/- 16 m to 135 +/- 33 m (+87%, p<0.001)and from 140 +/- 30 m to 266 +/- 62 m (+90%, p<0.001), respectively. Analysis of the Walking Impairment Questionnaire responses in the PGE1 group at 4 weeks demonstrated significant improvements in the walking impairment score (+19 percentage points, p<0.001), in the distance score (+25 percentage points, p<0.001), in the speed score (+24 percentage points, p<0.001), in the stair climbing score (+20 percentage points, p<0.001). The RAND survey responses showed improvements in physical function and bodily pain scores (+14 percentage points, p<0.001, and +15 percentage points, p<0.01, respectively). After the treatment-free follow-up period of 8 weeks, increases in PFWD and MWD were maintained (113 +/- 26 m, +57%, p<0.001, and 229 +/- 55 m, +63%, p<0.001, respectively). Similarly, at the end of the treatment-free follow-up, the walking impairment score (+16 percentage points, p<0.001), the distance score (+23 percentage points, p<0.001), the speed score (+22 percentage points, p<0.001), the stair climbing score (+18 percentage points, p<0.001) as well as the RAND physical function and bodily pain scores (+10 percentage points, p<0.001, and +13 percentage points, p<0.01, respectively) were still increased compared with baseline. No change from baseline was found in all the target parameters in the placebo group after 4 weeks of treatment and at the end of the treatment-free follow-up period. These data show that a 4-week treatment with PGE1 improves functional status and quality of life as well as treadmill performance in patients with disabling intermittent claudication as compared with placebo treated patients. The improvements are also maintained for a period of 8 weeks beyond the end of the treatment. Additional studies are needed to determine the duration of functional benefits after the end of treatment. PMID- 10870854 TI - Isolated nonfilling of contrast in deep leg vein segments seen on phlebography, and a comparison with color Doppler ultrasound, to assess the incidence of deep leg vein thrombosis. AB - Nonfilling of contrast in deep veins on phlebography is claimed to be an indirect sign of deep vein thrombosis (DVT) by some authors but rejected by others. The aim of this study was to prospectively assess, with color Doppler ultrasound (CDU), the occurrence and distribution of DVT in isolated, nonfilling, deep vein segments seen on a phlebogram. One hundred consecutive patients with clinical signs of acute DVT, in whom phlebography displayed nonfilling of the posterior tibial veins and/or the deep calf muscle veins, were examined with CDU on the same occasion. Ultrasound confirmed a DVT in 31 (31%) patients; in another 38 (38%) patients other pathology, without concomitantly detected DVT, such as edema, bleedings, ligament and muscle ruptures, Baker cysts, or superficial thrombophlebitis were found instead; and in the remaining 31 (31%) patients no pathology that could explain the nonfilling was identified. Isolated, nonfilling of the posterior tibial and/or deep muscle veins of the calf found by phlebography may be an indirect sign of DVT but is equally commonly caused by other pathological conditions or arises without any detectable explanation. When the thrombotic burden is to be scored, and to facilitate the establishment of the correct diagnosis, additional CDU is recommended when isolated nonfilling is present. PMID- 10870855 TI - Feasibility and reliability of ankle/arm blood pressure index in preventive medicine. AB - Despite its potential usefulness for assessing preclinical atherosclerosis and cardiovascular risk, the ankle/arm blood pressure index (AAI) has not yet been the matter of study evaluating its feasibility and reliability by nonspecialist doctors in a general population. This study was planned for two steps. In step 1, the measurement of AAI, (ratio between Doppler systolic pressure at the ankle for each lower limb and the highest value of Doppler systolic pressure of the two upper limbs), should be performed by 50 general practitioners (GPs), 50 social security center physicians, and 50 occupational health physicians in 3,000 male smokers, 40 to 59 years, without clinical cardiovascular disease. In step 2, AAI measurement, coupled with echography-Doppler of iliofemoral arteries, should be repeated by a specialist in all subjects with decreased AAI (<0.90) and the first two subjects with normal AAI recruited in step 1 by each nonspecialist. The number of physicians and subjects participating in step 1 was lower than planned (80 physicians and 962 subjects) with the greatest defect for GPs (six physicians and 35 subjects) and the prevalence of decreased AAI was low (28 subjects). AAI measurement was repeated in step 2 in only 12 subjects with decreased AAI in step 1 and in 124 subjects with normal AAI in step 1. Five of the six subjects with decreased AAI in step 2 also had decreased AAI in step 1 and 123 of the 130 subjects with normal AAI in step 2 also had normal AAI in step 1. As regards echographic stenosis, decreased AAI had a sensitivity of 44% and a specificity of 98%. AAI seems more feasible for occupational health physicians and social security center physicians and AAI is also reliable for nonspecialists previously trained, but its predictive value as regards echographic stenosis is poor in asymptomatic subjects, which may limit its usefulness for detecting preclinical atherosclerosis. PMID- 10870856 TI - Prevalence of anticardiolipin antibodies in peripheral arterial thrombosis. AB - The aim of this study was to assess prospectively the prevalence of anticardiolipin antibodies in patients with peripheral arterial thrombosis. Fifty nine patients with arterial thrombosis (38 men, 21 women), ages ranging from 24 to 80 years, mean age, 45.5 years, were studied. The control group included 100 volunteer blood donors. After confirmation of the thrombotic event, anticardiolipin antibodies were dosed by the immunoassay enzyme method and by dilution. In Group I these antibodies were present in 62.7% of the patients, of which 33.9% were borderline and 28.8% were positive. In the control group 6% of the patients were borderline and 1% was positive. The statistical analysis showed that the prevalence found was statistically significant (p < 0.001), even when the borderline titers were not considered positive. Patients with peripheral arterial thrombosis have a high prevalence of anticardiolipin antibodies. PMID- 10870857 TI - Paravalvular regurgitation: a rare complication following valve replacement surgery. AB - Paravalvular regurgitation is an uncommon but important complication, usually following valve replacement surgery. Early recognition and management are important for reoperations are associated with high morbidity and mortality rates. Presently, little data are available on this topic. The authors review the subject. PMID- 10870858 TI - Homocysteine is related to neopterin and endothelin-1 in plasma of subjects with disturbed glucose metabolism and reference subjects. AB - Hyperhomocysteinemia is an independent risk factor for vascular disease. In order to evaluate relations between hyperhomocysteinemia and endothelial and leukocyte function, the investigators related homocysteine to indices of endothelial function (plasma endothelin-1 [p-ET-1] and intraplatelet levels of the nitric oxide [NO] and prostacyclin mediators 3'-5' guanosine monophosphate [cGMP] and cyclic 3'-5' adenosine monophosphate [cAMP]) and the monocyte-derived inflammatory mediator neopterin in 168 men (mean age 69, range 49-72 years) with disturbed glucose metabolism and a reference group of 52 male subjects (mean age 70, range 61-79 years). Among the 168 patients with disturbed glucose metabolism plasma (p)-homocysteine correlated significantly with age (r=0.20; p<0.01), glycosylated hemoglobin (HbA1c) (r=0.17; p<0.05), triglycerides (r=0.20; p<0.05), intraplatelet GMP (r=0.16; p<0.05), p-ET-1 (r=0.21; p<0.05), and p-neopterin (r=0.31; p<0.001). The correlation between p-homocysteine and p-ET-1 persisted (p<0.01) in multiple regression analysis. Among the 52 reference subjects p homocysteine correlated significantly with p-ET-1 (r=0.32; p<0.05) and p neopterin (r=0.37; p<0.01). The correlation between p-homocysteine and p neopterin persisted (p<0.05) in multiple regression analysis. In conclusion, homocysteine is related to neopterin and endothelin-1 in plasma of subjects with disturbed glucose metabolism and in reference subjects, suggesting that homocysteine exerts its deleterious effects on vascular function through interference with endothelial and leukocyte function. PMID- 10870859 TI - Increased QT interval dispersion after hemodialysis: role of peridialytic electrolyte gradients. AB - Chronic renal failure patients on maintenance hemodialysis (HD) have a number of ECG abnormalities and cardiac arrhythmias. Clinical and experimental data have shown that increased QT dispersion is associated with severe ventricular arrhythmias and sudden cardiac death. Therefore, the aim of this study was to investigate whether the uremic patients receiving long-term HD have increased QTc interval and/or QTc dispersion compared to normal subjects and to evaluate the effect of electrolyte changes between the predialysis and postdialysis phases on these parameters. Forty patients with end-stage renal failure on long-term HD (22 men, 18 women, mean age 44 years) were included in this study. Serum concentrations of K+, Na+, Ca++, Mg++, Cl-, phosphate, urea, creatinine, HCO3-, and arterial blood gases (PO2, PCO2), together with blood pH, were monitored and QTc intervals and QTc dispersion were measured from 12-lead ECG in predialysis and postdialysis phases. The hemodialyzed patients had an increased predialysis QTc maximum interval and QTc dispersion compared to normal subjects (480 +/- 51 vs 310 +/- 38 msec, p < 0.001 and 61 +/- 17 vs 42 +/- 14 msec, p < 0.001, respectively). Both QTc maximum interval and QTc dispersion increased significantly at the end of the HD (480 +/- 51 vs 505 +/- 49 msec p < 0.001 and 61 +/- 17 vs 86 +/- 18 msec, p < 0.001, respectively). The serum K+ (5.3 +/- 0.56 vs 3.36 +/- 0.41 mEq/L, p < 0.001), phosphate (7.19 +/- 1.62 vs 3.81 +/- 1.02 mg/dL, p < 0.001), magnesium (0.87 +/- 18 vs 0.75 +/- 0.14 mg/dL) and urea concentrations (174 +/- 22 vs 74 +/- 14 mg/dL, p < 0.001) significantly decreased, whereas the Ca++ (2.21 +/- 0.18 vs 2.47 +/- 0.24 mg/dL, p < 0.001), HCO3- (15.5 +/- 3.2 vs 20.1 +/- 3.4 mmol/L, p<0.001) concentrations and pH (7.27 +/- 1.1 vs 7.43 +/- 1.2, p < 0.001) significantly increased after HD compared to predialysis values. There was significant correlation between the QT dispersion increase and serum electrolyte changes (K+, Ca++, and pH levels) (p < 0.05). The association between serum electrolyte changes, acid-base status and QT measurements might provide new insights into the evaluation of the ionic bases involved in inhomogeneous ventricular repolarization. PMID- 10870860 TI - Intravenous leiomyomatosis extending into the right ventricular cavity: one-stage radical operation using cardiopulmonary bypass--a case report. AB - The authors describe a 47-year-old woman with intravenous leiomyomatosis (IVL) extending into the right ventricular cavity. This rare entity is a neoplasm originating from smooth muscle of the uterus, with vermiform extensions into the inferior vena cava. The patient underwent a one-stage operation under simultaneous sternotomy and laparotomy, and radical excision of the tumor was successfully achieved with use of normothermic cardiopulmonary bypass. Although this tumor is histologically benign, it sometimes extends into the cardiac cavity and causes sudden death due to incarceration into the atrioventricular orifice. Moreover, recurrence or lung metastasis of IVL has been reported. The authors recommend a one stage-radical resection of the tumor or a two-staged operation within a short interval. In the literature, 24 surgical cases of the intravenous leiomyomatosis with intracardiac extension have been reported. The diagnosis and surgical treatment of this tumor are reviewed and discussed. PMID- 10870861 TI - Transjugular approach to treat portal vein stenosis after liver transplantation- a case report. AB - The authors present a case of portal vein stenosis that developed shortly after liver transplantation in a 48-year-old woman treated successfully with portal stent implantation by use of a transjugular approach to achieve portal vein access. To their knowledge, there are no other reports in the literature on the use of a transjugular approach with vein stent placement in the treatment of portal stenosis complicating orthotopic liver transplantation. PMID- 10870862 TI - Acute occlusion of an abdominal aortic aneurysm--case report and review of the literature. AB - Acute thrombosis of an abdominal aortic aneurysm (AAA) is a surgical emergency. Only 44 cases have been reported in the literature. The mechanism of the thrombosis has not been delineated. The proposed etiologies include propagation of thrombus from distal artery occlusion, cardiac thromboembolism, and dislodgment of a mural thrombus. Patients often present bilateral lower extremity ischemia, mimicking a saddle embolism. Systemic heparinization immediately after diagnosis and prompt surgical revascularization can reduce the mortality rate. The authors present a patient with sudden thrombosis of an AAA who was successfully treated with an axillobifemoral bypass graft. All published cases of thrombosed AAAs are analyzed. PMID- 10870863 TI - Middle aortic syndrome: an atypical case--a case report. AB - This case concerns a 61-year-old woman presenting with middle aortic syndrome treated by an aortoaortic thoracoabdominal polytetrafluoroethylene bypass and a right renal Dacron bypass. The case was atypical because of the patient's advanced age at the time of clinical presentation. PMID- 10870865 TI - Perspectives in the utilisation of Fourier-transform infrared spectroscopy of serum in sports medicine: health monitoring of athletes and prevention of doping. AB - Doping prevention is mainly directed to providing information on the dangers of doping to young athletes and to every profession concerned with athletic performance. Unfortunately, repression is also necessary in the fight against doping. Measurement of performance-enhancing drugs is complex, partly because of the large number of prohibited substances. A number of sophisticated analytical techniques are increasingly being used to provide the maximum detection time window. However, the effectiveness of methods to separate exogenous from endogenous biological molecules and the cost of antidoping analyses makes controls invalid or impossible. Moreover, most athletes, because of the metabolic and psychological stresses caused, legitimately refuse blood testing. It is becoming crucial to introduce new methods in the form of longitudinal health monitoring, since this is probably the most effective tool to prevent the use of doping agents when athletes become overtrained and/or overstressed. This paper describes new methods using Fourier-transform infrared spectroscopy to analyse serum from 50 microl samples of capillary blood. This technique has been shown to allow determination of the concentration of a wide range of biological molecules in a single microsample with clinically useful accuracy, and to provide a 'discriminatory biomolecular profile' to differentiate individuals on the basis of their physiological status. A specific application of this methodology is to perform longitudinal health monitoring in athletes, allowing prevention of overtraining. It is proposed to apply such methods in longitudinal studies for health monitoring and prevention of doping. PMID- 10870866 TI - Risk factors associated with anabolic-androgenic steroid use among adolescents. AB - To identify risk factors associated with anabolic-androgenic steroid (AAS) use among adolescents, computerised and manual literature searches were performed and the resultant local, state, national and international reports of illicit AAS use by adolescents that referenced risk factors were reviewed. Results indicate that adolescent AAS users are significantly more likely to be males and to use other illicit drugs, alcohol and tobacco. Student athletes are also more likely than non-athletes to use AAS, and football players, wrestlers, weightlifters and bodybuilders have significantly higher prevalence rates than students not engaged in these activities. Currently, only a partial profile can be created to characterise the adolescent AAS user. Further research will be needed before associations can be made with a reasonable degree of confidence regarding risk factors such as athletic participation, ethnicity, socioeconomic status and educational level. More importantly, to improve prevention and intervention strategies, a better understanding of the process involved in initiating AAS use is needed, including vulnerability factors, age of initiation and the use of other illicit drugs. PMID- 10870864 TI - The effect of endurance training on parameters of aerobic fitness. AB - Endurance exercise training results in profound adaptations of the cardiorespiratory and neuromuscular systems that enhance the delivery of oxygen from the atmosphere to the mitochondria and enable a tighter regulation of muscle metabolism. These adaptations effect an improvement in endurance performance that is manifest as a rightward shift in the 'velocity-time curve'. This shift enables athletes to exercise for longer at a given absolute exercise intensity, or to exercise at a higher exercise intensity for a given duration. There are 4 key parameters of aerobic fitness that affect the nature of the velocity-time curve that can be measured in the human athlete. These are the maximal oxygen uptake (VO2max), exercise economy, the lactate/ventilatory threshold and oxygen uptake kinetics. Other parameters that may help determine endurance performance, and that are related to the other 4 parameters, are the velocity at VO2max (V-VO2max) and the maximal lactate steady state or critical power. This review considers the effect of endurance training on the key parameters of aerobic (endurance) fitness and attempts to relate these changes to the adaptations seen in the body's physiological systems with training. The importance of improvements in the aerobic fitness parameters to the enhancement of endurance performance is highlighted, as are the training methods that may be considered optimal for facilitating such improvements. PMID- 10870867 TI - Oxidation of carbohydrate feedings during prolonged exercise: current thoughts, guidelines and directions for future research. AB - Although it is known that carbohydrate (CHO) feedings during exercise improve endurance performance, the effects of different feeding strategies are less clear. Studies using (stable) isotope methodology have shown that not all carbohydrates are oxidised at similar rates and hence they may not be equally effective. Glucose, sucrose, maltose, maltodextrins and amylopectin are oxidised at high rates. Fructose, galactose and amylose have been shown to be oxidised at 25 to 50% lower rates. Combinations of multiple transportable CHO may increase the total CHO absorption and total exogenous CHO oxidation. Increasing the CHO intake up to 1.0 to 1.5 g/min will increase the oxidation up to about 1.0 to 1.1 g/min. However, a further increase of the intake will not further increase the oxidation rates. Training status does not affect exogenous CHO oxidation. The effects of fasting and muscle glycogen depletion are less clear. The most remarkable conclusion is probably that exogenous CHO oxidation rates do not exceed 1.0 to 1.1 g/min. There is convincing evidence that this limitation is not at the muscular level but most likely located in the intestine or the liver. Intestinal perfusion studies seem to suggest that the capacity to absorb glucose is only slightly in excess of the observed entrance of glucose into the blood and the rate of absorption may thus be a factor contributing to the limitation. However, the liver may play an additional important role, in that it provides glucose to the bloodstream at a rate of about 1 g/min by balancing the glucose from the gut and from glycogenolysis/gluconeogenesis. It is possible that when large amounts of glucose are ingested absorption is a limiting factor, and the liver will retain some glucose and thus act as a second limiting factor to exogenous CHO oxidation. PMID- 10870869 TI - Tinea gladiatorum: wrestling's emerging foe. AB - Tinea corporis, or ringworm, has become a common nuisance in competitive wrestling. Although it is a fairly benign infectious skin disease, it has significant effects on the ability of a wrestler to compete because of infection control issues. Very little has been published in the medical literature describing this problem. The majority of the literature has described outbreaks in an isolated group of wrestlers. One must examine ringworm infections in wrestlers as an entity distinct from tinea corporis infections typically seen in the paediatric population, thus the term 'tinea gladiatorum'. Tinea gladiatorum outbreaks have been caused by the dermatophyte, Trichophyton tonsurans. The epidemiology and microbiology point to person-to-person contact as the main source of transmission in wrestlers. The clinical features of tinea gladiatorum may or may not be consistent with those found in the general population. Ancillary tests, including potassium hydroxide preparations and fungal cultures may have to be done to confirm the diagnosis. Treatment guidelines for tinea corporis have failed to produce the desired goals in this particular population. More research studying different treatment regimens in the wrestling environment is needed to define the optimal treatment to return wrestlers to competition quickly without putting other wrestlers at risk for infection. Intuitive hygiene practices have been suggested to prevent spread of the infection, but they have not been substantiated. Anecdotal reports suggest that hygiene practices fall short of producing adequate primary or secondary prevention. Pharmaceutical prophylaxis has been effective, but universal drug prophylaxis carries risks including drug adverse effects and potential drug resistance. The role of potential asymptomatic carriers of dermatophytes has yet to be elucidated in the origin and/or perpetuation of tinea gladiatorum outbreaks. There are many unanswered questions about tinea gladiatorum. Sports medicine professionals must work to define this entity more completely before making recommendations about treatment, prevention and infection control. The ultimate goal is the eradication of tinea infections from the wrestling world. Energy should be focused on primary and secondary prevention, as well as treatment. Without a thorough knowledge of tinea gladiatorum as a distinct disease entity, wrestling has been losing its battle with this formidable opponent. PMID- 10870868 TI - Biomechanical analysis of the effect of orthotic shoe inserts: a review of the literature. AB - Physical activity is increasingly recognised as an important component of primary disease prevention. Overuse injuries are common sequelae of exercise and sporting activities in general, and of running in particular, frequently resulting in cessation of activity. It has been proposed that there is a link between foot shape, foot function and the occurrence of injury. As a means of treatment and prevention of further injury, orthoses and shoe inserts are widely prescribed in the belief that they can alter the pattern of lower extremity joints' alignment and movement. Although this is an assumption widely made in the treatment of many joint conditions, the manner through which this treatment could be effective is not clear. This article aims to examine the literature to gain an improved understanding of the present state of knowledge regarding the effect of foot shape and orthotic use on foot kinematic and plantar pressure characteristics. The effects of foot type on the occurrence of lower limb injury during sporting activities and different aspects of biomechanics are reviewed, and the effects of applying orthoses on injury treatment and prevention and on various aspects of biomechanics of the lower limb joints are discussed. Further research is required, firstly to establish the casual effect of foot type and function on the risk of lower extremity overuse injury, and secondly to document the specific effect of orthotic therapy on injury treatment and prevention. Specifically, more prospective studies are necessary to investigate the long term effect of orthotic intervention. PMID- 10870870 TI - A comparison of the effects of quetiapine ('seroquel') and haloperidol in schizophrenic patients with a history of and a demonstrated, partial response to conventional antipsychotic treatment. PRIZE Study Group. AB - Quetiapine ('Seroquel') is a well-tolerated, novel, atypical antipsychotic with consistent efficacy in the treatment of schizophrenia. To date, no clinical studies have evaluated the effect of quetiapine in patients who only partially respond to conventional antipsychotics, yet this type of patient is most frequently seen by psychiatrists. Therefore, this international, multicentre, double-blind study was conducted to compare the efficacy and tolerability of 8 weeks' treatment of quetiapine 600 mg/day with haloperidol 20 mg/day in 288 patients who had a history of partial response to conventional antipsychotics and displayed a partial or no response to 1 month of fluphenazine (20 mg/day) treatment. Patients on quetiapine tended to have greater improvement than those on haloperidol in the primary efficacy measure, mean Positive and Negative Symptom Scale (PANSS) score, after 4 weeks' treatment (-9.05, -5.82, respectively, P = 0.061) and at study end (-11.50, -8.87, respectively, P = 0.234). Similarly, there was a trend towards patients on quetiapine demonstrating greater improvements in the secondary efficacy measures (Clinical Global Impression, PANSS subscale and Brief Psychiatric Rating Scale scores) [week 4 (baseline) to week 12 (end)], but the difference between treatments did not reach significance. Significantly more patients on quetiapine than on haloperidol showed a clinical response-patient response rates, defined as > 20% reduction in PANSS total score between weeks 4 and 12, were 52.2% for quetiapine and 38.0% for haloperidol (P = 0.043). Patients receiving quetiapine required less anticholinergic medication (P < 0.011), had greater reduction in extrapyramidal symptoms (EPS) (P = 0.005) and fewer treatment-emergent EPS-related adverse events compared to those on haloperidol (P < 0.001). Serum prolactin concentrations were elevated at the end of fluphenazine treatment in 73% of patients. Between weeks 4 and 12, elevated serum prolactin concentrations significantly decreased in quetiapine-treated patients compared to those receiving haloperidol (P < 0.001). At the end of quetiapine treatment, 83% of patients had normal prolactin levels while only 21% of patients receiving haloperidol were within the normal range. These results suggest that quetiapine may make a valuable contribution to the management of patients with a history of partial response to conventional antipsychotics. PMID- 10870871 TI - Milnacipran efficacy in the prevention of recurrent depression: a 12-month placebo-controlled study. Milnacipran recurrence prevention study group. AB - To compare the efficacy and assess the tolerability of milnacipran 50mg p.o. b.i.d. to placebo in the prevention of recurrence in depressed patients who had responded an acute treatment and had remained in remission during a 4-month continuation phase. Remission criteria were: a Hamilton Depression Rating Scale (HDRS) (21-item) < or = 8, improvement or disappearance of the initial symptoms, and an assessment of 'very much improved' or 'much improved' on the Clinical Global Impression (CGI) Subscale: Global Improvement. Recurrence was defined by a major depressive episode according to DSM-III-R criteria and a minimum score of 18 on the HDRS, with the need to treat the recurrence. The primary analysis was the rate of recurrence as a function of time in the intent-to-treat population. Groups were compared using the Cox model. Absolute recurrence rates were 16.3% (17/104) in milnacipran-treated patients and 23.6% (26/110) in placebo-treated patients, with a significant difference in the reduction of recurrence as a function of time (Kaplan Meier Survival Analysis analysis, P < 0.05). There was no difference in tolerability between groups. This study demonstrates that milnacipran is effective with good tolerability in preventing recurrence in major depressive disorder over 1 year in patients with recurrent depression who responded to acute treatment with milnacipran and continued their response for 18 weeks. PMID- 10870872 TI - Zaleplon improves sleep without producing rebound effects in outpatients with insomnia. Zaleplon Clinical Study Group. AB - The efficacy and safety of three doses of zaleplon, a novel non-benzodiazepine hypnotic, were compared with those of placebo in outpatients with insomnia in this 4-week study, using zolpidem 10 mg as active comparator. Postsleep questionnaires were used to determine treatment effects on the patient's perception of sleep, as well as any development of pharmacological tolerance during therapy or rebound insomnia or withdrawal symptoms upon discontinuation of therapy. During week 1, sleep latency was significantly shorter with zaleplon 5, 10, and 20 mg compared to placebo. The significant decrease in sleep latency persisted through week 4 with zaleplon 20 mg, and was again evident with zaleplon 10 mg at week 3. Zaleplon 20 mg also had significant effects on sleep duration, number of awakenings, and sleep quality compared to placebo. No pharmacological tolerance developed during treatment with zaleplon and there were no indications of rebound insomnia or withdrawal symptoms after treatment discontinuation. Zolpidem 10 mg had significant effects on sleep latency, sleep duration, and sleep quality compared to placebo. However, a significantly greater incidence of withdrawal symptoms and a suggestion of sleep difficulty after treatment discontinuation (rebound insomnia) for all sleep measures was seen with zolpidem compared to placebo. There was no significant difference in the frequency of adverse events with active treatment compared to placebo. These results show that zaleplon provides effective treatment of insomnia with a favourable safety profile. PMID- 10870873 TI - Increased psychological responses and divergent neuroendocrine responses to m-CPP and ipsapirone in patients with panic disorder. AB - In patients with panic disorder and /or agoraphobia (PDA) an increased sensitivity of central 5-HT2C receptors and a decreased responsiveness of 5-HT1A receptors has been postulated. In the present study, neuroendocrine challenges were performed using oral doses of the non-selective 5-HT2C agonist m chlorophenylpiperazine (m-CPP) (0.4 mg/kg), the selective 5-HT1A antagonist ipsapirone (0.3 mg/kg), and placebo in 40 patients with PDA and 12 healthy controls in order to compare 5-HT2C and 5-HT1A-specific psychobehavioural and neuroendocrine response patterns. At baseline, all psychobehavioural variables and the plasma concentration of noradrenaline (NE) were significantly increased in the patient group compared to the controls. The administration of m-CPP or ipsapirone was followed by comparable psychological symptoms and, in 55% of all patients, panic attacks. In comparison to the control subjects, patients were characterized by significantly higher psychological reactions to both challenge agents and a significantly higher NE response to m-CPP. In the patient group, there was also a trend towards an increased cortisol response after administration of m-CPP and a decreased cortisol and hypothermia response after administration of ipsapirone compared to the control group. The neuroendocrine findings of our study support earlier reports of opposite changes in the responsiveness of 5-HT2C and 5-HT1A-related receptors in PDA patients. The behavioural hypersensitivity to both, m-CPP and ipsapiron, shows that the provocation of anxiety and other psychological symptoms might be influenced by PMID- 10870874 TI - Parental obsessive-compulsive disorder as a prognostic factor in a year long fluvoxamine treatment in childhood and adolescent obsessive-compulsive disorder. AB - Interest in the treatment of pediatric obsessive-compulsive disorder (OCD) has increased as our knowledge of adult OCD has expanded. Although adults are still the majority of patients, children and adolescents with OCD are being identified and treated more frequently. As this population is better identified, prognostic factors need to be addressed to improve treatment outcome. The purpose of this study was to determine the role of family psychiatric pathology in fluvoxamine treatment outcome. Eleven children and adolescents with OCD and one of their parents participated in the study. Four parents were diagnosed with OCD, six had an Axis I diagnosis other than OCD, and one had no mental disorder [Structured Clinical Interview for the DSM-Non-Patient edition (SCID-NP)]. Each patient received fluvoxamine for 58 weeks. Dependent measures included the Children Yale Brown Obsessive Compulsive Scale (CY-BOCS), the NIMH-Global Obsessive Compulsive Scale (NIMH-GOCS) and the Clinical Global Impression (CGI). Based on CY-BOCS, CGI and NIMH-GOCS scores, patients with parents who have OCD showed a clinically and statistically significant reduction in symptoms from pre- to post-treatment. Patients whose parents did not have OCD also improved. However, the improvement was statistically but not clinically significant. The presence of OCD in one parent seems to modify a child's response to medication. The results suggest that family psychopathology, specifically presence of OCD, may contribute to treatment efficacy. Further research is suggested in this area. PMID- 10870875 TI - Beneficial effect of olanzapine in schizophrenic patients with obsessive compulsive symptoms. AB - Some studies suggest that obsessive-compulsive symptoms may be common (7.8-46%) in schizophrenic patients and seem to be poorly responsive to drug therapy. Conventional neuroleptics are of limited value, but adjunctive anti-obsessive agents (clomipramine, fluvoxamine) may be an option. Although novel atypical antipsychotics (clozapine, risperidone) reportedly aggravate the obsessive compulsive symptoms, a recent trial has shown that olanzapine did not induce new onset obsessive-compulsive symptoms in schizophrenic patients. We report our experience with three schizophrenic patients with obsessive-compulsive symptoms who were unsuccessfully treated with various conventional neuroleptics in combination with anti-obsessive agents and subsequently showed resistance or intolerance to clozapine. All of them were switched to olanzapine (10-20 mg/ day). All patients demonstrated a significant improvement in both schizophrenic and obsessive-compulsive symptoms as measured by the Brief Psychiatric Rating Scale (BPRS) and the Yale-Brown Obsessive Compulsive Scale (Y-BOCS). Within 5-8 weeks of initiation of olanzapine, the BPRS scores of the three patient decreased by 53%, 51% and 48%, and the Y-BOCS scores by 68%, 73% and 85%. Olanzapine was well tolerated. These preliminary results suggest that olanzapine may be a therapeutic option in schizophrenic patients with obsessive-compulsive symptoms. PMID- 10870876 TI - Effect of risperidone on plasma free 3-methoxy-4-hydroxyphenylglycol (pMHPG) levels in schizophrenic patients: relationship among plasma concentrations of risperidone and 9-hydroxyrisperidone, pMHPG levels, and clinical improvement. AB - We examined the relationships among the clinical efficacies of risperidone, plasma concentrations of risperidone and its active metabolite, 9 hydroxyrisperidone, and changes in plasma free MHPG (pMHPG) in 14 schizophrenic patients. Clinical improvement in negative symptoms of schizophrenia treated with risperidone has been associated with increased pMHPG and, in the present study, there were positive correlations between plasma 9-hydroxyrisperidone concentrations and increased pMHPG levels. These results suggest that risperidone might improve negative symptoms in schizophrenia by influencing noradrenergic neurons. PMID- 10870877 TI - Dependence on zolpidem in high dose. AB - There are some indications that zolpidem is open to potential abuse. We present four cases of former drug or alcohol abusers with personality disorders, who all developed dependence on zolpidem in high dose. PMID- 10870878 TI - Diverse genetic mechanisms operate to generate atypical betaS haplotypes in the population of Guadeloupe. AB - In a survey of the chromosomal background associated with the sickle cell gene in Guadeloupe, a French Caribbean island, we identified 37 unrelated patients with sickle cell disease (27 SS, nine SC, and one S-beta-thalassemia) of 477 unrelated sickle cell patients where the beta3 gene was linked to 20 different atypical haplotypes. These atypical chromosomes account for about 5% of the overall betaS chromosomes in this population. To investigate the origin of these atypical betaS haplotypes, we performed extensive typing of betaS and betaA chromosomes. Twenty two different 5' subhaplotypes were identified among the betaS chromosomes. Fifteen of 20 different atypical haplotypes are likely to be the product of recombination by a single crossover around the <> 5' to the beta-globin gene, or between a major betaS haplotype and one of the betaS haplotypes present in the population. The remaining cases require genetic mechanisms (gene conversions, additional substitutions in a given haplotype) other than crossovers to generate these atypical haplotypes. PMID- 10870879 TI - Geographic and ethnic distribution of beta-thalassemia mutations in Uttar Pradesh, India. AB - We have studied the geographic and ethnic distribution of mutations in 376 subjects who were carriers of beta-thalassemia, and identified the mutations in 365 chromosomes. The majority of the beta-thalassemia carriers were of Uttar Pradesh (India) origin. Their pattern of mutations differed from the other states of India and from those families who had migrated from Pakistan. The frequency of the IVS-I-5 (G-->C) and 619 bp deletion mutations were 64.3 and 2.5%, respectively, among families originating from Uttar Pradesh, compared to a prevalence of 37.5 and 27.5%, respectively in the population of Pakistani immigrants. Of the 10 common Asian Indian mutations, only eight were observed in subjects studied from different parts of India. By use of the amplification refractory mutation system along with DNA sequencing techniques, the mutations were successfully identified in 97.1% of subjects, while 11 cases (2.9%) still remain to be characterized by single strand conformation polymorphism and sequencing analyses. The application of this knowledge has facilitated the successful implementation of the program of genetic counseling and prenatal diagnosis of beta-thalassemia, thus helping to avoid the birth of an affected child in India. PMID- 10870880 TI - Molecular analysis of beta-thalassemia in Vietnam. AB - The molecular basis of the thalassemias has been studied in many of the world's populations. Here we report the results of the first screening for mutations in Vietnam. Twenty-three unrelated patients, of which 17 have Hb E/beta-thalassemia, were diagnosed and beta-globin mutations were detected in all 46 chromosomes. Four previously reported South Asian mutations were found. The most common mutations were the nonsense in codon 17 (A-->T) and the frameshift at codons 41/42 (-TTCT) (30 and 22%, respectively). The rare frameshift mutation at codon 95 (+A) was present in 9% of the 46 chromosomes studied, suggesting that it is indigenous to Vietnam. These results will serve as an initial database for DNA based prenatal diagnosis of thalassemia in Vietnam. PMID- 10870881 TI - Relative quantitation of mRNA in beta-thalassemia/Hb E using real-time polymerase chain reaction. AB - beta-Thalassemia and Hb E patients, with seemingly identical genotypes, have a remarkable variability in severity. Reduction in red cell survival in beta thalassemia is correlated with the amount of intracellular unmatched alpha-globin chains. However, it was only recently realized that mRNA, whose translation is prematurely terminated, is also unstable. No systematic attempts have been made to investigate mRNA stability in beta-thalassemia arising from nonsense mutations located upstream from the normal termination codon. In this study, one-step real time polymerase chain reaction has been employed to compare the levels of alpha- and beta-globin mRNA in reticulocytes from beta-thalassemia/Hb E subjects. The results showed the highest alpha/beta-globin mRNA ratio (median = 5.70, n = 13) in frameshift codons 41/42 (-TTCT)/Hb E individuals compared to normal subjects (median = 1.02, n = 6), or those with Hb E trait (median = 2.15, n = 8). In addition, there was a concomitant increase in the alpha/beta-globin mRNA ratio with decrease in hemoglobin level, i.e., increase in severity. The difference in the ratio among beta-thalassemia/Hb E patients with the same genotype may be attributed to individual variations of efficiency in betaE-globin mRNA splicing and in the destruction of prematurely terminated mRNA. PMID- 10870882 TI - Hb Tsukumi [beta117(G19)His-->Tyr]: a new hemoglobin variant found in a Japanese male. AB - We accidentally observed an abnormal elution pattern on high performance liquid chromatogram when we examined the Hb A1c level in a 65-year-old male patient who suffered from pneumoconiosis and alcoholic liver injury. The value of the glycated fraction was within the normal range but the elution patterns on high performance liquid chromatography varied with the glycohemoglobin analyzers. Isoelectrofocusing and urea-cellulose column chromatography showed an anomalous fast-moving beta chain estimated at approximately 47%. The instability test of the hemolysate was slightly positive. Structural analysis demonstrated that the mutant was consisted by a substitution of His-Tyr at beta117. This new variant was named Hb Tsukumi for the place of residence of the patient. Additionally, the nucleotide sequence showed a change of C-->T [CAC (His)-->TAC (Tyr)] at the first base in the 117th codon of the beta gene. PMID- 10870883 TI - Two new variants with the same substitution at position beta122: Hb Bushey [beta122(GH5)Phe-->Leu] and Hb Casablanca [beta65(E9)lys-->Met; beta122(GH5)Phe- >Leu]. AB - Hb Bushey, found in a Chinese baby and his father, is a new variant with a point mutation leading to the substitution Phe-->Leu at position beta122. Hb Casablanca, found in a family from Morocco, is a further example of a hemoglobin variant that carries two abnormalities in the same chain; the first is identical to that of Hb Bushey and the second to that of Hb J-Antakya [beta65 (E9)Lys- >Met]. Structural abnormalities of both Hbs were determined by protein chemistry methods including electrospray and tandem mass spectrometry. Their stability and oxygen binding properties were found to be identical to those of Hb A. Various mechanisms that may lead to two point mutations in the same chain are reviewed briefly. PMID- 10870884 TI - Hb Madrid [beta115(G17)Ala-->Pro] in a Korean family with chronic hemolytic anemia. AB - Hb Madrid, in which the alanine residue at beta115 (G17) is replaced by proline, results in a moderately severe hemolytic anemia due to the disruption of an alpha helical region and the weakening of an alpha1 beta1 contact (1,2). It was first discovered in a single Spanish patient, by protein structural analysis, whose parents did not carry the abnormality (1). The second observation of Hb Madrid was in an American Black teenager by DNA analysis, who had the family history of chronic hemolytic anemia, but none of family members were available for evaluation (3). PMID- 10870885 TI - A second case of Hb Bologna-St. Orsola [beta146(HC3)His-->Tyr] in an unrelated family of Anglo-Celtic origin. PMID- 10870886 TI - Hb Campinas [alpha26(B7)Ala -->Val]: a novel, electrophoretically silent, variant. PMID- 10870887 TI - Hb Saint Etienne or Hb Istanbul [beta92(F8)His-->Gln] found in an Argentinean family. PMID- 10870889 TI - Hb Rambam [beta69(E13)Gly-->Asp]/beta0-thalassemia [codon 5 (-CT)] in a family from Argentina. PMID- 10870888 TI - Hb I-high Wycombe [beta59(E3)Lys-->Glu]: the first instance in Japan. PMID- 10870890 TI - Two unrelated cases of Hb Old Dominion/Burton-upon-Trent [beta143(H21)His-->Tyr]: a rare variant causing spuriously elevated Hb A1c values. PMID- 10870891 TI - Identification of the nucleotide change (CAC-->CGC) responsible for Hb P Galveston [BETA117(G19)His-->Arg]. PMID- 10870892 TI - Dynamic mechanical conditioning of collagen-gel blood vessel constructs induces remodeling in vitro. AB - Dynamic mechanical conditioning is investigated as a means of improving the mechanical properties of tissue-engineered blood vessel constructs composed of living cells embedded in a collagen-gel scaffold. This approach attempts to elicit a unique response from the embedded cells so as to reorganize their surrounding matrix, thus improving the overall mechanical stability of the constructs. Mechanical conditioning, in the form of cyclic strain, was applied to the tubular constructs at a frequency of 1 Hz for 4 and 8 days. The response to conditioning thus evinced involved increased contraction and mechanical strength, as compared to statically cultured controls. Significant increases in ultimate stress and material modulus were seen over an 8 day culture period. Accompanying morphological changes showed increased circumferential orientation in response to the cyclic stimulus. We conclude that dynamic mechanical conditioning during tissue culture leads to an improvement in the properties of tissue-engineered blood vessel constructs in terms of mechanical strength and histological organization. This concept, in conjunction with a proper biochemical environment, could present a better model for engineering vascular constructs. PMID- 10870893 TI - In vitro cell shearing device to investigate the dynamic response of cells in a controlled hydrodynamic environment. AB - Mechanical stresses and strains play important roles in the normal growth and development of biological tissues, yet the cellular mechanisms of mechanotransduction have not been identified. A variety of in vitro systems for applying mechanical loads to cell populations have been developed to gain insight into these mechanisms. However, limitations in the ability to control precisely relevant aspects of the mechanical stimuli have obscured the physical relationships between mechanical loading and the biochemical signals that mediate the cellular response. We present a novel in vitro cell shearing device based on the principles of a cone and plate viscometer that utilizes microstepper motor technology to control independently the dynamic and steady components of a hydrodynamic shear-stress environment. Physical measurements of the cone velocity demonstrated faithful reproduction of user-defined input wave forms. Computational modeling of the fluid environment for the unsteady startup confirmed small inertial contributions and negligible secondary flows. Finally, we present experimental results demonstrating the onset rate dependence of functional and structural responses of endothelial cell cultures to dynamically applied shear stress. The controlled cell shearing device is a novel tool for elucidating mechanisms by which mechanical forces give rise to the biological signals that modulate cellular morphology and metabolism. PMID- 10870894 TI - Shear augmented dispersion of a solute in a casson fluid flowing in a conduit. AB - The unsteady dispersion of a solute in a Casson fluid flowing in a conduit (pipe/channel) is studied using the generalized dispersion model of Gill and Sankarasubramanian. With this approach, the entire dispersion process is described appropriately in terms of a simple diffusion process with the effective diffusion coefficient as a function of time, in addition to its dependence on the yield stress of the fluid. The results are accurate up to a first approximation for small times, but verified with Sharp to be exact for large times. The model brings out mainly the effect of yield stress, or equivalently, the plug flow region on the overall dispersion process. It is found that the rate of dispersion is reduced (i.e., the effective diffusivity decreases) due to the yield stress of the fluid, or equivalently, the plug flow region in the conduit. Also, the effective diffusivity increases with time, but eventually attains its steady state value below a critical time [0.48(a2/Dm) for dispersion in a pipe and 0.55(a2/Dm) for dispersion in a channel-the critical transient time for a Newtonian fluid-where "a" is the radius of the pipe and Dm is the molecular diffusivity]. At steady state, for dispersion in a pipe with the plug flow radius one tenth of the radius of the pipe, the effective diffusivity is reduced to about 0.78 times of the corresponding value for a Newtonian fluid at equivalent flow rates; for dispersion in a channel, the reduction factor is about 0.73 confirming the earlier result of Sharp. Further, the location of the center of mass of a passive species over a cross section is found to remain unperturbed during the course of dispersion and for different values of the plug flow parameter (i.e., the yield stress of the fluid). The study can be used as a starting first approximate solution for studying the dispersion in the cardiovascular system or blood oxygenators. PMID- 10870895 TI - Experimental and computational flow evaluation of coronary stents. AB - Local flow alterations created by a metallic stent in a simulated coronary artery were studied to compare the hemodynamic effects of two different stent geometries. Dye injection flow visualization and computational fluid dynamics were used. Resting and exercise conditions were studied. Flow visualization using the dye injection method provided a qualitative picture of stent hemodynamics while the computational approach provided detailed quantitative information on the flow next to the vessel wall near the intersections of stent wires. Dye injection visualization revealed that more dye became entrapped between the wires where the wire spacing was smallest. The dye washout times were shorter under exercise conditions for both wire spacings tested. The computational results showed that stagnation zones were continuous from one wire to the next when the wire spacing was small. Results from greater wire spacing (more than six wire diameters) showed that the stagnation zones were separate for at least part of the cardiac cycle. The sizes of the stagnation zones were larger under exercise conditions, and the largest stagnation zones were observed distal to the stent. These studies demonstrate that stent geometry has a significant effect on local hemodynamics. The observation that fluid stagnation is continuous in stents with wire spacings of less than six wire diameters may provide a criterion for future stent design. PMID- 10870896 TI - Shear rate dependence of ultrasound backscattering from blood samples characterized by different levels of erythrocyte aggregation. AB - The objectives were (1) to determine the effect of the erythrocyte aggregation level (wide range of aggregation) and shear rate (which also affects aggregation) on the ultrasound backscattered power, and (2) to evaluate the reproducibility of the ultrasound method. Experiments were performed under steady flow (100-1250 ml/min) in a 12.7 mm diameter vertical tube. Doppler ultrasound at 10 MHz was used to measure simultaneously the velocity and the backscattered power across the tube. For each radial position, the shear rate was computed from the derivative of the velocity profile. The backscattered power decayed exponentially as a function of the shear rate, and for a given shear rate, the power increased monotonically with the level of aggregation measured by laser reflectometry. Using blood samples simulating hypo-, normal, and hyperaggregating erythrocytes, the power of the ultrasound signal varied respectively by -7.8, -13.2, and -16.1 dB as a function of the shear rate (from 0.4 to 50 s-1). The reproducibility of the backscattered power was 5.5 dB, which is less than the variations observed as a function of the shear rate. In conclusion, ultrasound backscattering is sensitive to the level of erythrocyte aggregation. At a first glance, ultrasound seems less accurate when compared to laser reflectometry but it is suggested that this is because ultrasound backscattering may be sensitive to structural aggregate changes that are not detected by the laser method. PMID- 10870897 TI - Short term in vivo precision of proximal femoral finite element modeling. AB - As more therapies are introduced to treat osteoporosis, precise in vivo methods are needed to monitor response to therapy and to estimate the gains in bone strength that result from treatment. A method for evaluating the strength of the proximal femur was developed and its short term reproducibility, or precision, was determined in vivo. Ten volunteer subjects aged 51-62 years (mean 55.6 years), eight women and two men, were examined using a quantitative computed tomography (QCT) protocol. They were positioned, scanned, repositioned and re scanned. The QCT images were registered in three-dimensional space, and finite element (FE) models were generated and processed to simulate a stance phase load configuration. Stiffness was computed from each FE model, and strength was computed using a regression equation between FE stiffness and fracture load for a small set (n = 6) of experimental specimens. The coefficients of variation (COV) and repeatability (COR= 2.23* 42*COV) were determined. The COV for the FE fracture load computed was 1.85%, and the detectable limit (coefficient of repeatability) for serial measurements was 5.85%. That is, if a change of 5.85% or more in computed FE fracture load is observed, it will be too large to be consistent with measurement variation, but instead can be interpreted as a real change in the strength of the bone. The detectable limit of this method makes it suitable for serial research studies on changes in femoral bone strength in vivo. PMID- 10870898 TI - Flow patterns at the stenosed carotid bifurcation: effect of concentric versus eccentric stenosis. AB - Carotid stenosis severity is a commonly used indicator for assessing risk of stroke. However, the majority of individuals with severe carotid artery disease never suffer a stroke, and strokes can occur even with only mild or moderate stenosis. This suggests local factors (other than stenosis severity) at or near the carotid artery bifurcation may be important in determining stroke risk. In this paper we investigate the effect of stenosis geometry on flow patterns in the stenosed carotid bifurcation, using concentrically and eccentrically stenosed anthropomorphic carotid bifurcation models having identical stenosis severity. Computational simulations and experimental flow visualizations both demonstrate marked differences in flow patterns of concentric and eccentric stenosis models for moderately and severely stenosed cases, respectively. In particular, we identify post-stenotic recirculation zone size and location, and spatial extent of elevated wall shear stress as key factors differing between the two geometries. As these are also rotid plaque more vulnerable to cerebral embolus prokey biophysical factors promoting thrombogenesis, we propose that the stenosed carotid bifurcation geometry--or the induced flow patterns themselves--may provide more specific indicators for those plaques that are vulnerable to enhanced thromboembolic potential, and hence, increased risk of ischemic stroke. PMID- 10870899 TI - Remodeling of the bifurcation asymmetry of right coronary ventricular branches in hypertrophy. AB - To document the remodeling of the asymmetric branching pattern of the coronary right ventricular branches (RVBs) in right ventricular hypertrophy (RVH), we computed an asymmetry ratio, S, for the diameters and lengths of all vessels, defined as the ratio of the daughter diameters and lengths, respectively. We have previously induced RVH in pigs by pulmonary stenosis for five weeks. At autopsy, silicone elastomer casts of the right coronary arteries were made and the morphometric data on the branching pattern and vascular geometry of the RVB were collected. Data on smaller vessels were obtained from histological specimens while data on larger vessels were obtained from vascular casts. The results show that the diameter asymmetry ratio was significantly decreased in RVH hearts. The asymmetry ratios of diameters and lengths were used to compute the asymmetry ratios for vascular resistance and flow of the various daughter vessels. It was found that the degree of asymmetry of the resistance and flow were decreased, which implies that the flow heterogeneity at a bifurcation is decreased in the RVH hearts. PMID- 10870900 TI - Flow visualization in mechanical heart valves: occluder rebound and cavitation potential. AB - High density particle image velocimetry, with spatial resolution of O(1 mm), was used to measure the effect of occluder rebound on the flow field near a Bjork Shiley Monostrut tilting-disk mitral valve. The ability to measure two velocity components over an entire plane simultaneously provides a very different insight into the flow compared to the more traditional point to point techniques (like Laser Doppler Velocimetry) that were utilized in previous investigations of the regurgitant flow. A picture of the effects of occluder rebound on the fluid flow in the atrial chamber is presented. Specifically, fluid velocities in excess of 1.5 m/s traveling away from the atrial side were detected 3 mm away from the valve seat in the local low pressure region created by the occluder rebound on the major orifice side where cavitation has been observed. This analysis is the first spatially detailed flow description of the effects of occluder rebound on the flow field past a tilting-disk mechanical heart valve and further reinforces the hypothesis that the rebound effect plays a significant role in the formation of cavitation, which has been implicated in the hemolysis and wear associated with tilting-disk valves in vivo. PMID- 10870901 TI - Scaling approach to study the changes through the gestation of human fetal cardiac and circulatory behaviors. AB - During human gestation, fetal body size increases considerably and important transformations occur to hemodynamics of the cardiovascular system of the fetus. Vascular compliances and resistances as well as the cardiac function show important changes. In order to investigate these modifications, a mathematical approach based on scaling techniques was developed. Vascular and cardiac parameters of the human fetus were related by allometric equations to the anatomical dimensions of vessels that, in turn, depend on the fetal body weight and the gestational age. A scaling factor (b) was identified for each parameter under study: vascular resistances and flow inertances decrease with gestational age (b= -0.33 for flow inertances) whereas vascular compliances remarkably increase (b= 1.33). Scaling factors were also adopted for the fetal cardiac parameters, according to experimental data on the development of fetal myocardium. Parameter values calculated for each week of the last trimester of the fetal gestation, were tested using a mathematical lumped parameter model, previously developed for a human fetus near the term of the gestation. The validation of the scaling method adopted for the parameters was performed by comparing the results of the simulations with a group of data obtained by Doppler velocimetry at different stages of fetal normal gestation. The adopted allometric equations were appropriate in describing the development of the human fetal circulatory system. The ductus venosus, the ductus arteriosus, and the foramen ovale, that conclude their function at the birth moment, as well as the lungs and the brain, do not follow the general growth rate and require different scaling factors. PMID- 10870902 TI - Diffusion-convection equation solved in parallel regions of the lung. AB - The single path model of airway gas transport was incorporated into each of Cruz (Cruz, J. C. Respir. Physiol. 86:1-14, 1991). Thus, the effect of time on the predicted gas fractions in and out of the lung could be evaluated. Two experimental maneuvers were simulated: (1) fast inhalation of an argon-oxygen mixture from a functional residual capacity and fast exhalation to residual volume, including inspiratory breath holdings of 5-20 s, and (2) the standard single-breath nitrogen washout test. Expired argon and nitrogen are predicted within a +3% error of the experimental data with no breath holding. Breath holding predictions were at variance with experimental results because the solution of the diffusion-convection equation produced even mixing in the alveoli at the end of inspiration. The minimum square of the difference between the experimental data (standard single-breath nitrogen washout test) and those provided by the model was 0.0016. This model is capable of generating a nitrogen expirogram with four phases when a vital capacity of oxygen is inhaled. However, the model failed to produce a sharp distinction between phase 3 and phase 4. Thus, we conclude that uneven emptying of parallel regions generates any expirogram (a fast or slow expiratory maneuver). The alveolar gas stratification that is created during inspiration disappears at the end of the inspiratory maneuver. As a result, breath holding maneuvers cannot be predicted in the anatomical model used. PMID- 10870903 TI - Fatigue-related loading imbalance on the shank in running: a possible factor in stress fractures. AB - In previous reports we have shown that in long distance running the impact acceleration on the shank increases with progressing fatigue. The aim of the present study was to test whether, in parallel to this increase, an imbalance in the activities between the ankle plantar and dorsi flexor muscles develops. The tests were made on fourteen subjects during 30 min treadmill running above their anaerobic thresholds. Respiratory data were collected to determine the anaerobic threshold speed and to indicate the progressively developing metabolic fatigue. Surface electromyogram (EMG) was monitored to indicate the changing activity of the shank muscles. In the tibialis anterior the average integrated EMG (iEMG) and the mean power frequency (MPF) significantly decreased from the beginning to the end of running. In the gastrocnemius iEMG did not change, while MPF increased during the course of running. The impact acceleration, measured by means of an accelerometer attached to the tibial tuberosity, significantly increased during the course of running. It was concluded that, with developing fatigue, an imbalance in the contraction of the shank muscles develops in parallel to an increase in shank shock acceleration. The combination of these two changes may hamper the loading balance on the tibia since the bone becomes exposed to excessive bending stresses and to higher risk of stress injury. PMID- 10870904 TI - A simulation study for the design of a control system for the blood concentration process in autotransfusion. AB - Autotransfusion is the process in which a patient serves as his or her own blood donor; its most important application is the intraoperative blood salvage, in which the blood collected during a surgical operation is filtered, concentrated, washed, and then reinfused. In an automatic autotransfusion device, such as the DIDECO Compact Advanced, red blood cells (RBCs) are separated from the other unwanted components and concentrated by using a rotating bowl and the effect of centrifugal force. An important characteristic of concentrated RBCs is their hematocrit (Hct), i.e., percent RBC volume divided by total blood volume. The aim of this study is to assess the feasibility of a controller, based on the artificial neural network approach, which is able to provide a closed loop control of the hematocrit of the blood in the bowl at the end of the concentration phase. A simulation approach was adopted both for training the network and for assessing its performances. The results obtained are quite satisfactory, since the target Hct was typically reached within a 3% error, and always within 6% in highly challenging situations. PMID- 10870905 TI - Mood and heuristics: the influence of happy and sad states on sensitivity and bias in stereotyping. AB - The influence of mood states on the propensity to use heuristics as expressed in stereotypes was examined using signal detection statistics. Participants experienced happy, neutral, or sad moods and "remembered" whether names connoting race (African American, European American) belonged to social categories (criminal, politician, basketball player). Positive mood increased reliance on heuristics, indexed by higher false identification of members of stereotyped groups. Positive mood lowered sensitivity (d'), even among relative experts, and shifted bias (beta) or criterion to be more lenient for stereotypical names. In contrast, sad mood did not disrupt sensitivity and, in fact, revealed the use of a stricter criterion compared with baseline mood. Results support theories that characterize happy mood as a mental state that predisposes reliance on heuristics and sad mood as dampening such reliance. PMID- 10870906 TI - Automatic vigilance: the attention-grabbing power of approach- and avoidance related social information. AB - The automatic processing of information was investigated, varying valence (positive vs. negative) and relevance (other-relevant traits [ORT] vs. possessor relevant traits [PRT]; G. Peeters, 1983) of stimuli. ORTs denote unconditionally positive or negative consequences for persons in the social environment of the holder of the trait (e.g., honest, brutal) whereas PRTs denote unconditionally positive or negative consequences for the trait holder (e.g., happy, depressive). In 2 experiments using the Stroop paradigm, larger interference effects were found for ORTs than PRTs. This is due to the behavior-relatedness of ORTs. In a go/no-go lexical decision task (Experiment 3), participants either had to withdraw their finger from a pressed key (i.e., "avoid") or had to press a key (i.e., "approach") if a word was presented. Responses to negative ORTs were relatively faster in the withdraw condition, whereas positive ORTs were relatively faster in the press condition. PMID- 10870907 TI - The use of category and exemplar knowledge in the solution of anchoring tasks. AB - Five studies examine the role that category and exemplar knowledge play in the mediation of anchoring effects--the assimilation of an absolute estimate to a previously considered standard. Studies 1 through 3 demonstrate that comparing the target object with a plausible anchor (i.e., a standard that constitutes a possible value for the target) leads to a selective increase in the accessibility of anchor-consistent exemplar knowledge about the target. This easily accessible knowledge is then used to generate the absolute estimate, which leads to its assimilation to the standard. Studies 4 and 5 demonstrate that comparing the target with an implausible anchor, however, involves the activation of knowledge about the general category of the target, rather than exemplar knowledge about the target itself. PMID- 10870908 TI - A safe haven: an attachment theory perspective on support seeking and caregiving in intimate relationships. AB - This study used an attachment theoretical framework to investigate support seeking and caregiving processes in intimate relationships. Dating couples (N = 93) were videotaped while one member of the couple (support seeker) disclosed a personal problem to his or her partner (caregiver). Results indicated that when support seekers rated their problem as more stressful, they engaged in more direct support-seeking behavior, which led their partners to respond with more helpful forms of caregiving. Responsive caregiving then led seekers to feel cared for and to experience improved mood. Evidence for individual differences was also obtained: Avoidant attachment predicted ineffective support seeking, and anxious attachment predicted poor caregiving. Finally, couples in better functioning relationships engaged in more supportive interactions, and participants' perceptions of their interaction were biased by relationship quality and attachment style. PMID- 10870909 TI - Violence is a curvilinear function of temperature in Dallas: a replication. AB - Data on weather and aggravated assaults were obtained to determine whether the curvilinear relationship between temperature and violence previously observed in Minneapolis, Minnesota (E. G. Cohn & J. Rotton, 1997), could be replicated. The data consisted of calls for services received by police in Dallas between January 1, 1994, and December 31, 1995. Controlling for holidays, school closings, time of day, day of the week, season of the year, and their interactions, moderator variable autoregression analyses indicated that assaults were an inverted U shaped function of temperature. Replicating past research, the curvilinear relationship was dominant during daylight hours and spring months, whereas linear relationships were observed during nighttime hours and other seasons. The results are interpreted in terms of routine activity theory and the negative affect escape model of aggression. PMID- 10870910 TI - Psychophysiological responses to imagined infidelity: the specific innate modular view of jealousy reconsidered. AB - Three studies measured psychophysiological reactivity (heart rate, blood pressure, and electrodermal activity) while participants imagined a mate's infidelity. The specific innate modular theory of gender differences in jealousy hypothesizes that men are upset by sexual infidelity and women are upset by emotional infidelity, because of having faced different adaptive challenges (cuckoldry and loss of a mate's resources, respectively). This view was not supported. In men, sexual-infidelity imagery elicited greater reactivity than emotional-infidelity imagery. But, sexual imagery elicited greater reactivity even when infidelity was not involved, suggesting that the differential reactivity may not specifically index greater jealousy. In two studies with reasonable power, women did not respond more strongly to emotional infidelity. Moreover, women with committed sexual relationship experience showed reactivity patterns similar to those of men. Hypothetical infidelity self-reports were unrelated to reactivity. PMID- 10870911 TI - The influence of cognitive load on self-presentation: can cognitive busyness help as well as harm social performance? AB - Extra cognitive loads can hinder challenging self-presentations by usurping needed cognitive resources but also may sometimes improve them by shifting attention away from negative self-preoccupation. In Study 1, extraverts and introverts participated in an interview in which they presented themselves as either extraverted or introverted. Congruent self-presentations, which should be cognitively nondemanding, were unaffected by a cognitive busyness manipulation (rehearsing an 8-digit number). However, incongruent self-presentations were affected by busyness. Busyness decreased the effectiveness of extraverts who tried to appear introverted but increased the effectiveness of introverts who tried to appear extraverted. Study 2 found that introverts, who also tend to be socially anxious, reported less public self-consciousness and fewer negative self focused thoughts when they were busy than when they were not busy. PMID- 10870912 TI - Taxometric analyses of sexual orientation and gender identity. AB - Taxa are nonarbitrary classes whose existence is an empirical question and not a matter of mere semantic convenience. Taxometric procedures detect whether numerical relations between purported indicators of conjectured taxa bear the hallmarks of true taxa. On the basis of theoretical considerations, the current study tested whether taxa underlie sexual orientation and related measures of gender identity. Two taxometric procedures, maximum covariance, making hits maximum (MAXCOV) and mean above minus below a cut (MAMBAC), were applied to Kinsey Scales and measures of childhood gender nonconformity and adult gender identity in a sample of nearly 5,000 members of the Australian Twin Registry. Results suggest that latent taxa underlie these measures. About 12-15% of men and 5-10% of women belong to latent taxa associated with homosexual preference. These percentages are greater than those of individuals who report homosexual preference, however, and hence it appears that an appreciable proportion of individuals in these taxa have heterosexual preference. An understanding of the origins of these latent taxa may be important to understanding the development of sexual orientation and gender identity. PMID- 10870913 TI - The pleasures and pains of distinct self-construals: the role of interdependence in regulatory focus. AB - Regulatory focus theory distinguishes between self-regulatory processes that focus on promotion and prevention strategies for goal pursuit. Five studies provide support for the hypothesis that these strategies differ for individuals with distinct self-construals. Specifically, individuals with a dominant independent self-construal were predicted to place more emphasis on promotion focused information, and those with a dominant interdependent self-construal on prevention-focused information. Support for this hypothesis was obtained for participants who scored high versus low on the Self-Construal Scale, participants who were presented with an independent versus interdependent situation, and participants from a Western versus Eastern culture. The influence of interdependence on regulatory focus was observed in both importance ratings of information and affective responses consistent with promotion or prevention focus. PMID- 10870914 TI - Behavioral activation and inhibition in everyday life. AB - Joint effects of daily events and dispositional sensitivities to cues of reward and punishment on daily positive affect (PA) and negative affect (NA) were examined in 3 diary studies. Study 1 showed that positive events were strongly related to PA but not NA, whereas negative events were strongly related to NA but not PA. Studies 2 and 3 examined how the dispositional sensitivities of independent appetitive and aversive motivational systems, the Behavioral Activation System (BAS) and the Behavioral Inhibition System (BIS), moderated these relationships. Participants in Study 2 with higher BAS sensitivity reported more PA on average; those with more sensitive BIS reported more NA. Also, BIS moderated reactions to negative events, such that higher BIS sensitivity magnified reactions to negative events. Study 3 replicated these findings and showed that BAS predisposed people to experience more positive events. Results demonstrate the value of distinguishing within-person and between-person effects to clarify the functionally independent processes by which dispositional sensitivities influence affect. PMID- 10870916 TI - Thrombocytosis in temporal arteritis rising platelet counts: a red flag for giant cell arteritis. AB - OBJECTIVES: To determine whether early recognition and detection of thrombocytosis in patients with giant cell arteritis can help secure an earlier diagnosis, and whether it can help differentiate cases of arteritic optic neuropathy from other forms of optic neuropathy. METHODS: Medical and ophthalmologic records from 1993 to 1998 of the authors' patients with biopsy proven temporal arteritis versus the authors' patients with nonarteritic anterior ischemic optic neuropathy and idiopathic demyelinating optic neuritis were prospectively collected. Past and present blood analyses were collected, and platelet counts were compared between patients with giant cell arteritis and control populations. This was done to determine whether thrombocytosis could help with the diagnosis and differentiation of these different disease states. RESULTS: There was a significant difference in the frequency of thrombocytosis in patients with giant cell arteritis (13 out of 19 patients), with or without arteritic ischemic optic neuropathy, as compared with those with nonarteritic anterior ischemic optic neuropathy (zero out of 30 patients), idiopathic optic neuritis (zero out of 26 patients), and healthy age-matched controls (one out of 22 control subjects). This difference was especially helpful in patients whose sedimentation rates were within the normal range (adjusting for age). Also noted was the finding that the rise in the platelet counts was not acute, but rather it was a slow gradual increase for approximately 12 months before the onset of significant systemic or visual symptoms. CONCLUSION: Thrombocytosis should be considered an important marker in patients being referred for evaluation of ischemic optic neuropathy, diplopia, amaurosis fugax, headache, or even generalized malaise. Westegren sedimentation rates <50 mm/hr are often erroneously viewed as nondiagnostic or equivocal in the elderly and just followed. An over-the-phone review of patients' sedimentation rates, complete blood counts, and platelet counts can lead to expedited evaluation and treatment of patients who may be at high risk of visual loss from temporal arteritis. Thrombocytosis should lower a physician's threshold to acutely treat patients for possible arteritic ischemic optic neuropathy until the disease is definitely ruled out. PMID- 10870915 TI - Coping through emotional approach: scale construction and validation. AB - Four studies demonstrate the psychometric adequacy and validity of scales designed to assess coping through emotional approach. In separate undergraduate samples, exploratory and confirmatory factor analyses of dispositional (Study 1) and situational (Study 3) coping item sets yielded 2 distinct emotional approach coping factors: emotional processing (i.e., active attempts to acknowledge and understand emotions) and emotional expression. The 2 scales yielded high internal consistency and test-retest reliability, as well as convergent and discriminant validity. A study (Study 2) of young adults and their parents established the scales' interjudge reliabilities. Longitudinal (Study 3) and experimental (Study 4) research supported the predictive validity of the emotional approach coping scales with regard to adjustment to stressful encounters. Findings highlight the utility of functionalist theories of emotion as applied to coping theory. PMID- 10870917 TI - Ocular motility review for 1997-1998: part I. AB - Each year brings new scientific knowledge that builds on itself in a geometric fashion. Ocular motility basic and clinical neurosciences continue to advance with this accelerating pace. The years 1997 through 1998 brought new knowledge to the motility world. This review focuses on the clinical advances within this realm, presented in supranuclear to myopathic organization. Part II of this review will appear in the September 2000 (20:3) issue. PMID- 10870918 TI - Postpartum optic neuritis: etiologic and pathophysiologic considerations. AB - The clinical course of four patients with visual loss in the postpartum period due to acute optic neuritis is described. Factors that disclosed the underlying etiology and expression of disease are discussed. The clinical records of four women examined and managed for visual loss after uncomplicated pregnancies and term deliveries were reviewed. Neurodiagnostic examination, treatment modalities, and outcomes were assessed. These four women with varied and confounding medical histories, all with optic neuropathy, eventually were demonstrated to harbor demyelinating disease. Although visual loss in the postpartum period evokes differential diagnostic considerations, the authors' experience suggests that puerperal immune-mediated changes are responsible for activation of optic neuritis associated with relapsing multiple sclerosis. PMID- 10870920 TI - Homonymous visual field defects in patients without corresponding structural lesions on neuroimaging. AB - Homonymous visual field defects usually occur with structural processes affecting retrochiasmal visual pathways. The responsible lesion is usually evident on magnetic resonance imaging or on other neuroimaging studies. When results of neuroimaging are normal, functional illness is often suspected. The authors report four patients with homonymous visual field defects who presented with no evident corresponding lesion on magnetic resonance or computed tomography imaging. Etiologies for the visual field defects included the Heidenhain variant of Creutzfeldt-Jacob disease, degenerative dementia, subtle occipital ischemia demonstrated only on positron-emission tomography scanning, and nonketotic hyperglycemia. Clinicians should be aware of the alternative etiologies of organic homonymous visual field loss in patients with normal neuroimaging. PMID- 10870921 TI - Monocular central dazzle after thalamic infarcts. AB - The authors observed a patient after he had ischemic strokes in both paramedian thalamic regions, which were more marked on the left side. Symptoms included dysphasia, vertical binocular diplopia, right-sided hemianopia, and a right-sided sensory and motor deficit, sparing the face. However, the most disturbing phenomenon was a painless, left monocular dazzle, which was the presenting symptom and also the only persisting symptom. This report shows that a thalamic lesion may be at the origin of central dazzle, and to the authors' knowledge, it is the first clinical observation of its monocular occurrence. It is conceivable that this dazzle was due to optic-trigeminal summation. PMID- 10870919 TI - The representation of the horizontal meridian in the primary visual cortex. AB - The authors report the findings of two patients that confirm the location of the horizontal meridian in the human visual cortex. The first patient had an inferior quadrant defect with a band of horizontal meridian sparing. Magnetic resonance imaging showed a lesion concentrated along the medial striate cortex. The second patient had a homonymous horizontal defect that resulted from removal of an arteriovenous malformation located in the lateral striate cortex. The findings of these two patients demonstrate that the horizontal meridian is represented at the calcarine fissure base in the primary visual cortex. PMID- 10870922 TI - Anisocoria associated with the medical treatment of irritable bowel syndrome. AB - A case of anisocoria associated with oral pharmacologic treatment of irritable bowel syndrome is reported. A 26-year-old woman developed sudden onset of anisocoria and compromised accommodation that lasted 2 days after the use of oral scopolamine methylbromide for treatment of irritable bowel syndrome. The anisocoria and compromised accommodation occurred after contamination of the ocular surface after administration of scopolamine methylbromide and resolved within 1 week without further contamination. Oral preparations used for the pharmacologic treatment of irritable bowel syndrome can cause anisocoria due to anticholinergic pharmacologic blockade of the iris sphincter muscle. PMID- 10870923 TI - Myasthenia gravis with a paraneoplastic marker. AB - Ocular manifestations of myasthenia gravis are very common. Myasthenia gravis may be associated with lung carcinoma. Lambert-Eaton syndrome is also commonly associated with lung carcinoma and can have ocular manifestations. Overlap of these two entities has been described. The case of a patient with fatigable diplopia and ptosis 3 years after removal of a large-cell lung carcinoma is presented. Tests results for acetylcholine receptor binding and modulating antibodies were positive for myasthenia gravis. Test results for presynaptic voltage-gated calcium channel antibodies of the N-type were also positive. However, test results for the P/Q-type voltage-gated calcium channel antibodies, which are consistent with Lambert-Eaton syndrome, were negative. Autoantibodies can be used to serologically distinguish paraneoplastic myasthenia gravis from Lambert-Eaton syndrome. PMID- 10870924 TI - Neuro-ophthalmologic manifestations of neuroendocrine carcinoma. AB - The neuro-ophthalmologic findings of parasellar neuroendocrine carcinoma are reported. Two patients with parasellar neuroendocrine carcinoma had headache, ptosis, and ophthalmoplegia. In both patients, neuroimaging revealed a parasellar mass with extension into the cavernous sinus. The tumors initially were believed to be pituitary adenomas, but histopathology confirmed neuroendocrine carcinoma. Clinicians should be aware of neuroendocrine carcinoma in the differential diagnosis of sellar/parasellar lesions causing ophthalmoplegia. PMID- 10870925 TI - Vision loss as the presenting sign in juvenile neuronal ceroid lipofuscinosis. AB - OBJECTIVE: To review cases of juvenile neuronal ceroid lipofuscinosis (JNCL) and highlight salient clinical and diagnostic features, thereby enhancing recognition of this disease among ophthalmologists. MATERIALS AND METHODS: Twelve cases of JNCL seen from 1982 to 1999 were reviewed. Diagnosis was based on characteristic clinical history, ophthalmoscopic findings, electroretinography, neuroimaging, histopathology, and molecular analysis. RESULTS: Vision loss was the first subjective symptom of the disease in all 12 cases. Among these cases, nine of 12 patients (75%) developed neurologic deficits an average of 3 years after the onset of visual deterioration. CONCLUSION: Because visual symptoms usually precede neurologic dysfunction, JNCL should be considered in the differential diagnosis when an apparently healthy child presents with unexplained bilateral vision loss. PMID- 10870926 TI - Selective loss of the electroretinogram B-wave in a patient with Creutzfeldt Jakob disease. AB - Creutzfeldt-Jakob disease (CJD) is a neurodegenerative disease characterized by movement abnormalities and dementia that inevitably progress to death. Familial, infectious, and sporadic forms of the disease are recognized. The worldwide incidence of CJD is estimated at 1:1,000,000 per year, and it affects middle-aged men and women in roughly equal proportions. The disease is caused by a unique infectious vector, the prion, which is a mutant form of a normally occurring cell surface protein found predominantly in the central nervous system. A significant proportion of patients with CJD will have visual disturbances at some point in their illness and may therefore consult a neuro-ophthalmologist. The case of a woman in whom the diagnosis of CJD was not known until autopsy is reported. Early in the course of her disease, she sought ophthalmic consultation because of vision problems. PMID- 10870927 TI - Posterior fixation suture augmentation of full-tendon vertical rectus muscle transposition for abducens palsy. AB - OBJECTIVE: To evaluate the effect of augmenting full-tendon vertical rectus transpositions with posterior fixation sutures in patients with complete or near complete lateral rectus palsy. METHODS: Transposition of the vertical recti to the lateral rectus muscle was performed in seven patients with unilateral lateral rectus palsy (the mean angle of preoperative horizontal deviation in primary gaze was 36.7 prism diopters (delta); range, 25-62delta of esotropia). A posterior fixation suture of 5.0 Mersilene (Ethicon, Somerville, NJ) was placed in sclera (14-16 mm posterior to the limbus) adjacent to the lateral rectus and incorporated 1/3 belly width of each transposed vertical rectus muscle. RESULTS: The mean angle of postoperative horizontal deviation in primary gaze was 7.1delta (range, 0-20delta). The mean change in primary-position horizontal deviation postoperatively was 41.2delta (range, 37-72delta). Four patients were able to fuse without prism in primary gaze; three patients were orthophoric and one patient had a consecutive intermittent exotropia. The remaining three patients required prism correction to neutralize the postoperative gaze deviation. All patients had improvement in abduction. Mild limitation of adduction was noted in three patients (range, -0.5 to -2.0). CONCLUSIONS: Augmenting full vertical rectus muscle transpositions with posterior fixation sutures improves the abducting effect of surgery without significant limitation of adduction. PMID- 10870928 TI - Internuclear ophthalmoplegia after coronary artery catheterization and percutaneous transluminal coronary balloon angioplasty. AB - A retrospective chart review was performed for identification of patients with isolated internuclear ophthalmoplegia (INO) postcardiac catheterization from two neuro-ophthalmology units. Of the 110 patients with a diagnosis of INO who were evaluated during the observation period, five patients (4.5%) demonstrated relatively isolated INO occurring in the perioperative period of a cardiac endovascular procedure. These five patients underwent diagnostic catheterization alone (three patients), balloon angioplasty (one patient), or stent placement (one patient). All patients improved, with resolution of diplopia in primary position after a mean period of 82 days. The occurrence of INO in the postcardiac catheterization setting is not uncommon, and it appears to be related to dorsal pontine ischemia. The pontomesencephalic medial longitudinal fasciculus is supplied by small-caliber perforating end-arteries from the basilar trunk, which increases selective vulnerability of this area. Cardiac catheterization may precipitate microemboli involving these vessels, leading to internuclear ophthalmoplegia. PMID- 10870929 TI - Unilateral thalamic infarction and vertical gaze palsy: cause or coincidence? AB - Although vertical gaze palsy (VGP) is commonly associated with lesions of the rostral mesencephalon, there is some evidence that VGP may also be caused by a unilateral thalamic lesion. The case of a 68-year-old man with persistent upward gaze palsy after a unilateral thalamic infarction, demonstrated on computed tomography and magnetic resonance imaging scans, is presented. Subsequent high resolution magnetic resonance scanning, however, showed involvement of the rostral mesencephalon as well. The authors suggest that in previous patients with VGP ascribed to a unilateral thalamic infarction, a coexisting mesencephalic involvement may have been missed because of inappropriate imaging techniques. Strong evidence of unilateral thalamic infarction as a cause of VGP is still lacking. PMID- 10870930 TI - Bilateral ptosis with pupil sparing because of a discrete midbrain lesion: magnetic resonance imaging evidence of topographic arrangement within the oculomotor nerve. AB - The topographic arrangement within the midbrain oculomotor nerve is not adequately elucidated in humans. Two patients with a partial oculomotor palsy because of a localized infarction or hematoma were treated. Both patients had bilateral ptosis, impaired adduction, and supraduction. One patient had impaired infraduction and pupillary involvement on one side. Results of computed tomography and magnetic resonance imaging revealed discrete lesions at the dorsal midbrain tegmentum that spared the rostral midbrain. The authors' cases elucidate that pupillary components take the most rostral course. This report provides indirect magnetic resonance imaging evidence to prove the course of pupillary fibers. Based on the different neuro-ophthalmologic findings in the authors' cases (sparing or affecting pupillary component and infraduction), the nerves of the inferior rectus and inferior oblique for infraduction pass more rostrally than those of medial rectus, superior rectus, and levator palpebrae. The nuclear and fascicular arrangement within the midbrain oculomotor nerve is speculated to be pupillary, extraocular, and eyelid elevation in the rostro-caudal order, based on the neuro-ophthalmologic impairment and magnetic resonance imaging findings in the authors' patients and in previous animal experiments. Knowing the fascicular and nuclear arrangement within the midbrain in detail will offer diagnostic clues for differentiation of causes for partial oculomotor palsy. PMID- 10870931 TI - 4.0 Tesla magnetic resonance imaging of brainstem lesions with ocular motility deficits. AB - The authors studied six patients with brainstem ocular motility deficits with 4.0 Tesla (T) magnetic resonance imaging to investigate whether a higher field strength would produce superior images compared with 1.5T. In four patients whose lesions were evident on 1.5T, the increased signal-to-noise achieved with 4.0T allowed for better resolution at 1-mm slice thickness than was achieved at the standard 5-mm slice thickness with 1.5T. In the two patients with unremarkable 1.5T scan results, 4.0T also failed to demonstrate a lesion. Therefore, 4.0T imaging has superior resolution to 1.5T imaging and can provide more detailed images of lesions identified by 1.5T. PMID- 10870932 TI - Insights into the three-dimensional structure of the oculomotor nuclear complex and fascicles. AB - The authors report the case of a patient with an ischemic lesion in the left midbrain. The patient presented with paresis of left inferior rectus, pupil, right superior rectus, convergence and transiently, of the left medial rectus. A lesion in the left dorsal midbrain close to the oculomotor nuclear complex, selectively involving the fascicles innervating the above muscles, is proposed. Fine magnetic resonance sections showed a consistent lesion in the left paramedian dorsal midbrain. A detailed, three-dimensional, schematic computer model of the oculomotor nucleus and fascicles was constructed. Using this model, the authors topographically validate the putative site of the lesion. The medial rectus subnucleus is divided into three subgroups, A, B, and C. Subgroup C is thought to be the site of the majority of neurons controlling convergence. In the above model, the putative lesion is closer to subgroup A than to C; this suggests that subgroup A, rather than subgroup C, may have a higher concentration of neurons involved in convergence. PMID- 10870933 TI - Hepatitis B in the family. AB - During a 3-year period (1992-1995), 239 index cases of hepatitis B virus (HBV) infection and 459 members of their households from the Osijek-Baranja county were examined. The aim of the study was to determine the spread of HBV infection in the families with a member verified as a virus carrier, and to identify the family members with the highest risk of infection according to kinship degrees. The retrospective and prospective methods were used in the study. The probable route of infection was assessed by the use of an epidemiologic questionnaire, and the serologic status of the study subjects concerning infection with HBV was determined by enzyme immunoassays (HBsAg, anti-HBs, anti-HBe and anti-HBc). The first member of a family identified as a virus carrier was considered an index case. HBV infection was demonstrated in 334 (47.85%) out of a total of 698 subjects. Only 21 (6.28%) of the 334 subjects with verified HBV infection developed the clinical picture of acute hepatitis B. The ratio of clinically manifest vs inapparent infection was 1:16. Serologic traces of infection were detected in 95 of the 459 family members of the index cases, yielding a mean rate of the infection among the virus carrier family members of 20.70%. PMID- 10870934 TI - Soluble tumor necrosis factor alpha receptors (sTNF-Rs) in HIV-1-infected intravenous drug users: change in circulating sTNF-R type II level and survival for AIDS patients. AB - This study in intravenous drug users (IVDUs) investigated differences in serum soluble tumor necrosis factor types I and II (sTNFR-I and II) concentrations in HIV-1-infected IVDUs and controls. This study also investigated whether changes of sTNFRs concentration affect the risk of death among patients with AIDS. A cross-sectional study of 54 subjects with AIDS, 47 HIV-seropositive IVDUs, 47 HIV seronegative IVDUs, and 21 healthy subjects showed that sTNFRs concentration increases from healthy controls to AIDS patients through HIV-seronegative and HIV seropositive subjects (p < 0.01). sTNFR-I concentration, however, was shown to be similar in HIV-seronegative IVDUs and healthy controls. In the longitudinal study, serum concentration of sTNFRs was determined near AIDS diagnosis in 21 IVDUs and 1 year later (start for the survival study). Cox proportional hazards regression was performed to assess the prognostic value of percent change of sTNFR level alone and in combination with T lymphocyte subsets, HIV-p24 antigenemia and opportunistic infections for death within 240 days. Uni- and multivariate Cox modelling for dichotomised variables according to its median showed an increase of sTNFR-II by at least 30% to be single significant predictor of death: crude relative risk 3.69, p = 0.03; adjusted relative risk 5.67, p = 0.02. Mean survival was 126 days in 11 patients whose sTNFR-II level increased by at least 30%, and 176 days in 10 patients with less change in sTNFR-II (p = 0.02). CONCLUSIONS: sTNFRs concentration is higher in IVDUs than in healthy controls and is highest in AIDS patients. Survival of patients with AIDS is associated with variation in the concentration of sTNFR-II. PMID- 10870935 TI - Survey on influenza laboratory diagnostic and surveillance methods in Europe. European Scientific Working Group on Influenza. AB - The survey was undertaken by ESWI in order to investigate the comparability of the laboratory diagnostic methods and the influenza surveillance systems used in 24 European countries. The results indicate considerable consensus in the general approaches to collection and use of clinical specimens, rapid diagnostic techniques, virus isolation techniques in eggs or/and MDCK cell lines, virus identification and use of inhibition of hemagglutination (IHA) and complement fixation (CF) tests for serological diagnostics. However, the details of the techniques used are somewhat heterogeneous: antigen detection methods (immunofluorescence versus immuno adsorbent assay), isolation methods (eggs versus tissue culture), reagents (locally produced, WHO, commercial) are not always equivalent and results are therefore not really comparable. Some of these discrepancies are due to a lack of resources or a lack of priority for influenza in the country. The greatest differences between individual countries exist in the epidemiological part of surveillance programmes. The mode of collection of influenza related mortality and absentism from work varies considerably in different countries. These findings indicate the need to harmonize viral procedures and surveillance systems in European countries in order to improve validity and comparability of results and as a prerequisite for early information on influenza etiology and spread. PMID- 10870937 TI - Assessment of aluminium in human deciduous teeth. AB - The possible role of environmental aluminium exposure in the pathogenesis of various diseases has highlighted the need for methods by which the long-term exposure to aluminium can be assessed. Therefore, we have further developed a method to determine aluminium in human deciduous teeth and applied this method for studying populations in Sweden, Crete and Iceland. PMID- 10870936 TI - Risk factors for low bone mineral density among a large group of Norwegian women with fractures. AB - The objectives of this study were to determine factors related to fractures and bone mineral density (BMD) in a large group of Norwegian women. In a cross sectional study, 3803 women aged 50-75, all with a history of fractures, were included in the study. BMD was measured with Dual energy X-ray absorptiometry at both hip (neck) and spine (L1-L4), while information on other factors thought to influence BMD were obtained through a questionnaire. In multivariate analysis, the strongest positive predictor of both hip and spine BMD was current body weight, while weight loss since the age of 25 and number of years since menopause were the strongest inverse predictors. In addition, use of cortisone and maternal history of fractures were associated with lower BMD, as was loss of height since the age of 25. Physical activity was positively correlated with BMD. These results show the complexity of factors involved in the etiology of osteoporosis, with several factors acting in synergism. This points to the need for multifactorial prevention strategies, which most effectively need to be instituted at an early age, before peak bone mass is achieved. PMID- 10870938 TI - Comparison of different procedures to identify probable cases of myocardial infarction and stroke in two Swedish prospective cohort studies using local and national routine registers. AB - In prospective cohort studies, person-time time is calculated from baseline until the first definite event occurs or until censoring. A way to correctly identify and date definite events when only routine registers are available is to retrieve all hospital discharge notes and death certificates with a diagnosis of probable event and perform a consecutive revision. It is important to detect all possible hospital stays as they may contain useful information for the revision study. Furthermore, loss to follow-up can be avoided by extending the retrieval outside the specific geographical area where the cohort was defined. The aims of this study were (i) to describe a comprehensive retrieval of probable myocardial infarctions (diagnosis with International Classification of Diseases 8th and 9th revisions codes 410-414) or stroke (codes 430-438), (ii) to quantify the relative efficiency of different local and national routine registers or their combination compared with the use of all available registers together, and (iii) to audit local and national registers by comparing their outcome at the county level. The study was performed in two prospective cohorts studies i.e., 'Men-born-1914' (n = 500) from Skane (period 1982-1993), and Skara-1 (n = 683) from Skaraborg (period 1988-1994.). All available routine registers were linked to the cohorts. The use of all available routine registers improved retrieval of both individual and hospital stays with a discharge diagnosis of probable event and gave an enhanced basis for a future validation study. Local registers were not completely covered by the national register, but the accessible combination of national inpatient and mortality registers was an efficient alternative. PMID- 10870939 TI - Increase of specific symptoms after long-term use of chlorophenol polluted drinking water in a community. AB - Chlorophenols contaminated the drinking water system and a local lake in the village of Jarvela in southern Finland. Period prevalence rates of symptoms, signs and diseases among the residents 15 years or older who responded (69%) to a survey in the contaminated area (1773 subjects) were compared with the rates of three uncontaminated areas (2018 subjects). Gastrointestinal and skin symptoms, in particular, were significantly (p < 0.05) more common in the contaminated area than in each control area. Nausea, general malaise, headache, anorexia, exceptional tiredness, and respiratory infections were significantly increased compared to the control areas combined. A dose-response was also observed: higher consumption of drinking water and contaminated fish further significantly increased (p < 0.05) reported symptoms. In conclusion, long-term use of chlorophenol polluted household water and fish can cause symptoms already familiar in connection with occupational chlorophenol exposures. PMID- 10870940 TI - Evaluation of a transit first-aid station providing emergency care to former Yugoslavian war victims evacuated in Ancona, Italy. AB - BACKGROUND: A first-aid station was implemented in Falconara Marittima airport (Ancona, Italy). It provided medical emergency care to war victims evacuated from former Yugoslavia in transit for further treatment. MATERIALS AND METHODS: A descriptive analysis of the displaced population arriving at the first-aid station was performed using three independent datasets for administrative information, of which one included medical information. The implemented resources were also evaluated. RESULTS: From August 1993 to March 1995, 2272 displaced persons were registered at the first-aid station, out of which 54.2% were accompanying family members. Among those needing medical intervention (45.8% of total), most frequent diagnoses were traumatisms and burns (59.8%), neoplasms (15.6%), and congenital malformations (13.2%). The medical care provided at the first-aid station was most often basic: a medical examination alone was performed on 77.0% of the patients, and a minor dressing on 17.3%. Median length of stay was 1 day. Patients were sent to 30 different countries and 8% were forwarded to the local regional hospital. Deployed logistical resources exceeded by far actual needs but a lack of psychological assistance was observed, mainly for children. The agencies involved did not coordinate data sharing and follow-up information. CONCLUSIONS: The medical assistance to the war victims was efficient regarding provided care and timeliness. Effectiveness of such a programme could be improved by a better coordination between partners, allowing more adequate logistics according to appropriate epidemiological information. PMID- 10870941 TI - Comparison of serotyping, ribotyping and pulsed-field gel electrophoresis for distinguishing group A Streptococcus strains isolated in Albania. AB - Conventional serotyping for T antigens, rRNA gene restriction fragment length polymorphism analysis (ribotyping) and pulsed-field gel electrophoresis were compared for distinguishing among group A streptococci isolated in Albania between 1980-1982 and in 1995. A total of twelve serotypes were identified among seventy GAS strains. Ribotyping revealed eight and eleven distinct patterns after digestion with HindIII and PvuII, respectively. Twenty-three strains of serotype T12 were subdivided in 10 ribotypes and 11 strains of T2 serotype were differentiated in 5 ribotypes. By comparison, PFGE generated 37 patterns after SmaI digestion. The index of discrimination, using the Hunter-Gaston formula, was applied to assess the value of these methods for interpretation of the epidemiological data. For serotyping the value of index was 0.85. The ribotyping system revealed an ID of 0.83 when the combination HindIII and PvuII was used. This index reached 0.97 for PFGE. The methods used were useful to subtype the isolates of GAS. PMID- 10870942 TI - Prevalence, awareness, treatment and control of hypertension in a working Bulgarian population. AB - Arterial hypertension is a major risk factor for coronary heart disease and stroke mortality. Few data exist on prevalence, awareness, and management of hypertension in Bulgaria, precluding development of potentially beneficial risk reduction initiatives. Between September 1996 and July 1997, an age-sex stratified sample of 847 male and 771 female employees (age 18-64 y) of the national transport industry resident in Sofia was recruited during their annual physical examination. A structured interview was conducted and resting blood pressure (BP) measured. Prevalence: Elevated BP (mean of two consecutive readings SBP > or = 140 mmHg and/or DBP > or = 90 mmHg) was observed among 24% of women and 58% of men (p < 0.001). Prevalence increased with age in both men and women. Awareness: Among 722 employees with elevated BP, 49% of women and 33% of men (p < 0.001) reported history of hypertension. Awareness increased with age. MANAGEMENT: Among 345 employees with history of hypertension, 37% of women and 36% of men (p > 0.05) reported taking antihypertensive treatment. The proportion under management increased with age. CONTROL: Normal BP was measured in only 6% of men and 7% of women taking antihypertensive medication (p > 0.05; no consistent trends by age). Elevated BP is widespread and hypertension is underdiagnosed and poorly controlled in this urban working-age Bulgarian population, especially among those under 40 y. This may contribute to the high rates of coronary heart disease and stroke incidence and mortality in Bulgaria. PMID- 10870943 TI - Tularemia in Bursa, Turkey: 205 cases in ten years. AB - Tularemia is a zoonotic disease caused by the coccobacillus F. tularensis. Small epidemics and sporadic cases were seen around Bursa since November 1988. In this study, a total of 205 cases of tularemia were observed. All the cases were diagnosed on clinical, bacteriological and serological grounds. The epidemics were thought to be waterborne. The majority of the patients were young and female. In most of the cases the disease presented itself in oropharyngeal form (83%). Analysing sera from the patients with microagglutination method demonstrated that titers were > or = 1:160 in approximately 85% of the cases, including the ones in subclinical form. Five of ten patients from who the bacteria was isolated were seronegative. Streptomycin was given to the most of the patients by combining with tetracycline, doxycycline or chloramphenicol. The early administration of these antibiotics (before the third week of disease) was found to be much more effective to resolve the infection. As a result, the main mode of transmission of F. tularensis is waterborne in our region. In our region, tularemia should be considered in differential diagnosis for the cases with fever, tonsillopharyngitis and cervical lymphadenopathy to make an early diagnosis and to design relevant treatment. PMID- 10870944 TI - Haemorrhagic papular rash associated to Flavimonas oryzihabitans bacteraemia in a child. AB - Flavimonas oryzihabitans is a gram-negative rod that has rarely been implicated in human infections. The involvement of this organism has been documented in serious infections, the majority of which were cases of bacteraemia or peritonitis. We report the first isolation of the organism in Greece, from a case of bacteraemia, associated with haemorrhagic papular rash, in a paediatric patient and describe the phenotypic characteristics of the strain. PMID- 10870945 TI - Sleep position, bedding and heating practices in high- and low-risk ethnic groups for unexpected death in infancy (UDI). AB - In a previous study, a significant increased risk for unexpected death in infancy (UDI) among Arab infants as compared to Jews (RR: 5.2) was found. The incidence has significantly decreased in both groups during the 'back to sleep' campaign. The objective of this study was to compare the prevalence of three risk factors, i.e. positioning, night dressing/covering and heating practices in these ethnic groups. A community sampling procedure was employed, resulting in the participation of 264 Jewish and 146 Arab mothers of infants between 1 and 4 months, born at term and with no chronic illness at the time of the study. A questionnaire in Arabic and Hebrew was designed, pertaining to sleep positioning at the time of the study and of the previous infant, prior to the SIDS prevention campaign, as well as clothing and heating practices. Significantly, more Arab infants were put to sleep in a supine or side position as compared to Jewish infants both during the study (p = 0.002) as well as prior to the SIDS campaign (p = 0.001). No ethnic difference was related to clothing practices. Open heating, however, was significantly more common in the Arab sector (p = 0.001). A logistic procedure for each of the practices indicated that ethnicity is related significantly to both sleep position (p = 0.002) and beating practices (p = 0.001). Prone sleep positioning was still prevalent (32.2%) more so among Jews (35.2%) than Arabs (27%). CONCLUSION: Sleep positioning and overdressing do not appear to be the major attributable risk factors for UDI among Arab infants as compared to Jews. The compliance with positioning recommendations is lower than expected in both ethnic groups. PMID- 10870947 TI - Acute myeloid leukaemia and exposure to organic solvents--a case-control study. AB - The objective was to study the relation between exposure to organic solvents and the risk of developing acute myeloid leukaemia (AML) in Caucasians aged 18 years and more. Ninety-eight cases of AML were diagnosed from September 1986 to March 1990 in Novi Sad, Yugoslavia, and in two London hospitals from September 1988 and May 1994 and from July 1992 and July 1994, respectively. Two controls were matched to each case by hospital, year of admission, gender and 5-year age group. Information on solvent exposure was collected by interview. Odds ratios (ORs) were derived with conditional logistic regression. The degree of solvent exposure was determined by three experts blind to the status of the subject with good agreement between them (the kappa coefficient ranged between 0.52 and 0.86). The response rate for cases was 80%. Exposure to solvents was associated with the increased risk of AML (OR: 2.52; 95% CI: 1.45-4.39; p = 0.001) and those with probable exposure to solvents were found to have an odds ratio of AML over three times greater than non-exposed. We also found an elevated OR for exposure to oils (OR: 1.56) but this was not statistically significant. There is no clear pattern of increasing risks with increasing duration of employment but a significant risk was found for exposures of 10 years or less. An induction period of less than 10 years or more than 30 years was associated with a significantly raised OR. There was a significant excess of machinery mechanics and fitters among the cases. PMID- 10870946 TI - Prevalence of enteroparasites in a residence for children in the Cordoba Province, Argentina. AB - A study of the prevalence of enteroparasites in a population belonging to a substitute home that gives shelter to orphaned and homeless children was done using conventional methods of analysis. This home is located in Cordoba Province, Argentina, and has the following characteristics: It has nine houses located inside the main plot of ground, that shelter 139 individuals, and 25 houses outside this plot distributed randomly in Unquillo city and that shelter 257 individuals. The overall parasitic infection, pathogen and commensal organisms included, yielded 84.8% and the prevalence of the most important parasites was: Enterobius vermicularis (43.4%), Giardia lamblia (23.0%), Ascaris lumbricoides (13.1%), Entamoeba coli (45.5%), Blastocystis hominis (44.4%) and Endolimax nana (34.6%). We also analyzed the population dividing it according to the residence place (inside or outside the plot), age and sex of the individuals. In reference to the location of the patients, A. lumbricoides and E. coli showed significant prevalence in the individuals living inside the plot (p < 0.001 and p < 0.005, respectively) and of B. hominis in those living outside the main plot (p < 0.005). Results indicated a greater parasitism level in the outside residents (61.5%, p < 0.001). When the individuals were studied according to sex, no significant difference was observed, except for E. vermicularis that showed greater prevalence in the male sex (p < 0.04). When the individuals were grouped according to age ranges, a greater prevalence in individuals from 5 to 14 years was noticed (p < 0.01). In this study is also included an analysis of the multiparasitism level that comprises the whole population. PMID- 10870948 TI - Incidence of E. coli O157:H7 and other enteropathogens in a Spanish hospital. AB - From January to December 1997, stools submitted for routine culture were also screened for E. coli O157:H7 to investigate its incidence in our area. Bloody and non-bloody stools were studied. E. coli O157:H7 was not recovered from any of the samples tested. We believe, therefore, that we are in a low rate of infection area, and accordingly do not recommend routine screening of this pathogen. PMID- 10870949 TI - Trends in mortality from major diseases in Europe during 1980 to 1993. PMID- 10870950 TI - Simplification of complex peptide mixtures for proteomic analysis: reversible biotinylation of cysteinyl peptides. AB - A rapid means of identifying many components in an enriched mixture of proteins is enzymatic digestion of the entire protein fraction. This complex peptide mixture is then subjected to reversed-phase high performance liquid chromatography (HPLC) coupled on-line with a mass spectrometer capable of data dependent ion selection for fragmentation (LC-tandem mass spectrometry; MS/MS). Thus, as many peptides as possible in the sample are fragmented to produce MS/MS spectra, which can then be searched against sequence databases. Ideally, one peptide from each protein in the mixture would be fragmented and identified. To this end, we employed an affinity selection method to capture cysteinyl peptides and thereby simplify the mixture. Both the captured cysteinyl and the noncysteinyl peptides are analyzed by LC-MS/MS, to increase the number of proteins identified. The method was tested on a limited set of standard proteins and applied to the analysis of a protein fraction obtained from isolated mitochondria treated with atractyloside. To further increase the number of different precursor ions selected for fragmentation, dynamic exclusion and ion selection from multiple narrow mass ranges of consecutive runs were employed. PMID- 10870951 TI - Fluorescent modification for peptide sequencing by postsource decay-matrix assisted laser desorption/ionization-mass spectrometry. AB - The sequential analysis of a peptide of CDYEGRLI, relating to the nucleic proteins in influenza virus, was performed by the postsource decay (PSD) fragmentation method using matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The sequence of the peptide was difficult to analyze by MALDI-MS since the PSD fragment ions of the peptide were almost never observed and were not amenable to complete sequence interpretation. The peptide was modified by 4(5)-(iodoacetamide) fluorescent reagent to improve the sensitivity of the MALDI-PSD fragment spectrum. In the spectrum of the fluorescent modified peptide, almost all sequential b-series fragment ions were observed clearly, which was sufficient for complete sequence interpretation. The results indicate the advantage of fluorescent modification for the total sequencing of the peptides by MALDI-MS. PMID- 10870952 TI - Preparative two-dimensional gel electrophoresis with agarose gels in the first dimension for high molecular mass proteins. AB - A two-dimensional gel electrophoresis (2-DE) method that uses an agarose isoelectric focusing (IEF) gel in the first dimension (agarose 2-DE) was compared with an immobilized pH gradient 2-DE method (IPG-Dalt). The former method was shown to produce significant improvements in the 2-D electrophoretic separation of high molecular mass proteins larger than 150 kDa, up to 500 kDa, and to have a higher loading capacity, as much as 1.5 mg proteins in total for micropreparative runs. The extraction medium found best in this study for agarose 2-DE of mammal tissues was 6 M urea, 1 M thiourea, 0.5% 2-mercaptoethanol, protease inhibitor cocktail (Complete Mini EDTA-free), 1% Triton X-100 and 3% 3-[(3 cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS). Trichloroacetic acid (TCA) treatment of the agarose gel after IEF is to be carefully weighed beforehand, because some high molecular mass proteins were less likely to enter the second-dimensional polyacrylamide gel after TCA fixation, and proteins such as mouse skeletal muscle actin gave pseudospots in the agarose 2-DE patterns without TCA fixation. As a good compromise we suggest fixation of proteins in the agarose gel with TCA for one hour or less. The first-dimensional agarose IEF gel containing Pharmalyte as a carrier ampholyte was 180 mm in length and 2.5-4.8 mm in diameter. The gel diameter was shown to determine the loading capacity of the agarose 2-DE, and 1.5 mg liver proteins in total were successfully separated by the use of a 4.8 mm diameter agarose gel. PMID- 10870953 TI - Sequence information of peptides and proteins with in-source decay in matrix assisted laser desorption/ionization-time of flight-mass spectrometry. AB - In-source decay coupled with matrix assisted laser desorption/ionization-mass spectrometry, which is a mass spectrometric degradation method for the sequencing of peptides and proteins, has been applied to several different polypeptides and proteins. The influence of the nature of the constituent amino acids on positively charged product ions is described. Relatively small molecular mass peptides produced c-, b-, and/or a-series ions usable for C-terminal sequencing as well as y- and/or z-series ions usable for N-terminal sequencing. The formation of the C-terminal sequencing ions (c, b and a) and the N-terminal sequencing ions (y and z) was strongly dependent on the location(s) of basic arginine and lysine residues. The presence of the arginine and/or lysine residues at the N-terminal region was one-sided in the formation of c-, b-, and/or a series ions, while the presence of those at the C-terminal region was favorable for the formation of y- and z-series ions. In-source decay experiments of intact proteins, apomyoglobin and two viral coat proteins, led to large amounts of c series ions and small amounts of y-series ions, which reflected internal sequences. PMID- 10870954 TI - A simplified device for protein identification by microcapillary gradient liquid chromatography-tandem mass spectrometry. AB - A simplified device and procedure have been developed for microcapillary gradient liquid chromatography-tandem mass spectrometry (LC-MS/MS). This procedure has proved useful in identifying low level quantities of proteins from sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel bands. Microelectrospray needles are packed with reversed-phase resin and function both as a high performance liquid chromatography (HPLC) column and a nanospray mass spectrometer tip when interfaced between an HPLC and ion trap mass spectrometer. Variable submicroliter flow rates are generated by flow splitting between the microelectrospray capillary and an HPLC system. A manual injector is used to inject a protein digest mixture that binds to the column and is then washed at a high flow rate (2 microL/min post split). Gradient elution of bound peptides was initiated by the injection of a filled loop of 70% v/v methanol (5 microL) concomitant with a reduction of flow rate (0.1 microL/min post split). This forms a diffusion-dependent gradient of variable length (typically 15-30 min in length) depending upon the final flow rate. Chromatographic separations of a standard solution digest demonstrate that this diffusion-dependent gradient provides reasonable separations such that multiple peptide identifications by MS/MS can be obtained. Application of this methodology to the analysis of several in-gel digested gel-separated proteins is presented to demonstrate its utility. PMID- 10870955 TI - Cardiac sarcoplasmic reticulum and sarcolemmal proteins separated by two dimensional electrophoresis: surfactant effects on membrane solubilization. AB - We evaluated the use of the alkyaryl amidosulfobetaine zwitterionic detergent, designated as C8psi, to facilitate the solubilization of cardiac subcellular, membrane-associated proteins. Hearts from 7-week-old male Sprague-Dawley rats were isolated, and the left ventricles dissected and subsequently homogenized. The sarcolemma (SL) and the sarcoplasmic reticulum (SR) were isolated from different homogenate preparations originating from rat hearts by ultracentrifugation methods. The isolated membrane preparations were solubilized and the proteins precipitated. After resuspension, protein separation was achieved in first-dimensional IEF using an immobilized (pH 4-7) gradient and in the second dimension using 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Gels were then stained, images analyzed, and protein spots excised for subsequent identification. Protein identification from both SR and SL samples did not identify any of the known major membrane-associated proteins. Solubilization of whole tissue lysates with C8psi resulted in no increase in the total number of proteins detected relative to samples solubilized in the presence of 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulforate (CHAPS). The data suggest the utility of newer surfactants such as C8psi to improve both the resolution of (2-D) protein profiles and increase the number of proteins extracted from subcellular organelle fractions. PMID- 10870956 TI - Automated interpretation of low-energy collision-induced dissociation spectra by SeqMS, a software aid for de novo sequencing by tandem mass spectrometry. AB - SeqMS, a software aid for de novo sequencing by tandem mass spectrometry (MS/MS), which was initially developed for the automated interpretation of high-energy collision-induced dissociation (CID) MS/MS spectra of peptides, has been applied to the interpretation of low-energy CID and post-source decay (PSD) spectra of peptides. Based on peptide backbone fragmented ions and their related ions, which are the dominant ions observed in the latter two techniques, the types of ions and their propensities to be observed have been optimized for efficient interpretation of the spectra. In a typical example, the modified SeqMS allowed the complete sequencing of a 31-amino acid synthetic peptide, except for the isobaric amino acids (Leu or Ile, and Lys or Gln), based on only the low-energy CID-MS/MS spectrum. PMID- 10870957 TI - Blocks-based methods for detecting protein homology. AB - The most highly conserved regions of proteins can be represented as blocks of aligned sequence segments, typically with multiple blocks for a given protein family. The Blocks Database World Wide Web (http://blocks.fhcrc.org) and e-mail (blocks@blocks. fhcrc.org) servers provide tools to search DNA and protein queries against the Blocks+ Database of multiple alignments. We describe features for detection of distant relationships using blocks. Blocks+ includes protein families from the PROSITE, Prints, Pfam-A, ProDom and Domo databases. Other features include searching Blocks+ with the BLIMPS and NCBI's IMPALA programs, sequence logos, phylogenetic trees, three-dimensional display of blocks on PDB structures, and a polymerase chain reaction (PCR) primer design strategy based on blocks. PMID- 10870958 TI - Proteomic analysis of the human colon carcinoma cell line (LIM 1215): development of a membrane protein database. AB - The proteomic definition of plasma membrane proteins is an important initial step in searching for novel tumor marker proteins expressed during the different stages of cancer progression. However, due to the charge heterogeneity and poor solubility of membrane-associated proteins this subsection of the cell's proteome is often refractory to two-dimensional electrophoresis (2-DE), the current paradigm technology for studying protein expression profiles. Here, we describe a non-2-DE method for identifying membrane proteins. Proteins from an enriched membrane preparation of the human colorectal carcinoma cell line LIM1215 were initially fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE, 4-20%). The unstained gel was cut into 16 x 3 mm slices, and peptide mixtures resulting from in-gel tryptic digestion of each slice were individually subjected to capillary-column reversed phase-high performance liquid chromatography (RP-HPLC) coupled with electrospray ionization ion trap-mass spectrometry (ESI-IT-MS). Interrogation of genomic databases with the resulting collision-induced dissociation (CID) generated peptide ion fragment data was used to identify the proteins in each gel slice. Over 284 proteins (including 92 membrane proteins) were identified, including many integral membrane proteins not previously identified by 2-DE, many proteins seen at the genomic level only, as well as several proteins identified by expressed sequence tags (ESTs) only. Additionally, a number of peptides, identified by de novo MS sequence analysis, have not been described in the databases. Further, a "targeted" ion approach was used to unambiguously identify known low-abundance plasma membrane proteins, using the membrane-associated A33 antigen, a gastrointestinal-specific epithelial cell protein, as an example. Following localization of the A33 antigen in the gel by immunoblotting, ions corresponding to the theoretical A33 antigen tryptic peptide masses were selected using an "inclusion" mass list for automated sequence analysis. Six peptides corresponding to the A33 antigen, present at levels well below those accessible using the standard automated "nontargeted" approach, were identified. The membrane protein database may be accessed via the World Wide Web (WWW) at http://www.ludwig. edu.au/jpsl/jpslhome.html. PMID- 10870959 TI - Characterization of native and recombinant peptidyl prolyl cis-trans isomerases derived from Methanococcus thermolithotrophicus based on cDNA sequence. AB - It is important to establish whether a recombinant protein is an authentic copy of the predicted cDNA sequence. In this study, recombinant protein for native peptidyl prolyl cis-trans isomerase (N-PPIase) and double-labeled (13C- and 15N-) protein (DL-PPIase) appeared on the sodium dodecyl sulfate (SDS) electropherograms as two bands for N-PPIase and four bands for DL-PPIase. Since the N-terminal amino acid residues of all bands were the same, we characterized these bands using the peptide mapping method and amino acid composition analysis. Peptide mapping of the proteins seemed to be almost identical but they could not reflect the whole amino acid sequences of the protein. The bands on the polyvinylidene difluoride (PVDF) membrane, electroblotted after SDS polyacrylamide gel electrophoresis (SDS-PAGE), were hydrolyzed and their amino acid composition was analyzed using a highly sensitive 6-aminoquinolyl-N hydroxysuccinimidyl carbamate (AQC) amino acid analysis and compared with the cDNA sequences for proteins. The matching score (sigma(T%-E%)2) for similarity of proteins was calculated by summation of the square difference between the theoretical (T%) and the experimental (E%) amino acid composition of the recombinant protein. The amino acid composition of all bands of both proteins showed more than 93% of the theoretical values. The major molecular weights of both proteins were 16812 and 17694 by electrospray ionization (ESI)-mass spectrometry. However, the purified proteins also contained minor compounds with Mr of 3721 for N-PPIase and 5285 for DL-PPIase. These compounds were considered to be nonpeptidyl products that comigrated with the protein. Similarities of the amino acid composition of the four bands were more than 98%. Our results indicate that AQC amino acid analysis is the most suitable method for characterization of a recombinant protein. PMID- 10870960 TI - A proteomic analysis of secreted proteins from xylan-induced Bacillus sp. strain K-1. AB - The expression level of extracellular proteins in an alkaliphilic bacterium, Bacillus sp. strain K-1, grown in a xylan-containing medium, is significantly increased when compared with that grown in the nonxylan culture medium. A proteomic approach has been efficiently applied to separate and characterize these differentially expressed secretory proteins. Eight prominent protein spots were identified and subjected to N-terminal amino acid sequencing. The results show that three spots share considerable similarity with the xylanolytic enzymes and that two spots share considerable similarity with the GltC regulatory protein and 3-dehydroquinate dehydratase, respectively. In addition, the three other proteins show little similarity with the known proteins in the database. In conclusion, our results demonstrate that the proteomic approach is a highly efficient method to rapidly study the differential expression of the secreted proteins by Bacillus sp. strain K-1 grown under xylan-induced condition. PMID- 10870961 TI - Proteomic study of the peripheral proteins from thylakoid membranes of the cyanobacterium Synechocystis sp. PCC 6803. AB - Thylakoid membranes of cyanobacteria and plants contain enzymes that function in diverse metabolic reactions. Many of these enzymes and regulatory proteins are associated with the membranes as peripheral proteins. To identify these proteins, we separated and identified the peripheral proteins of thylakoid membranes of the cyanobacterium Synechocystis sp. PCC 6803. Trichloroacetic acid (TCA)-acetone extraction was used to enrich samples with peripheral proteins and to remove integral membrane proteins. The proteins were separated by two-dimensional electrophoresis (2-DE) and identified by peptide mass fingerprinting. More than 200 proteins were detected on the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel that was stained with colloidal Coomassie blue. We analyzed 116 spots by peptide mass fingerprinting and identified 78 spots that were derived from 51 genes. Some proteins were found in multiple spots, indicating differential modifications resulting in charge differences. Therefore, a significant fraction of the peripheral proteins in thylakoid membranes is modified post-translationally. In our analysis, products of 17 hypothetical genes could be identified in the peripheral protein fraction. Therefore, proteomic analysis is a powerful tool to identify location of the products of hypothetical genes and to characterize complexity in gene expression due to post-translational modifications. PMID- 10870962 TI - Profiling of Caenorhabditis elegans proteins using two-dimensional gel electrophoresis and matrix assisted laser desorption/ionization-time of flight mass spectrometry. AB - The nematode Caenorhabditis elegans (C. elegans) is the first animal whose whole 97 Mb genome sequence, encoding ca. 19000 open reading frames (ORF's), has been essentially determined. We tried to establish a 2-DE map of the nematode proteome by means of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). A soluble protein fraction of mixed stages of the worm, wild-type strain N2, was applied to 2-D PAGE. After Coomassie Brilliant Blue (CBB) staining, 1200 spots were detected and 140 major spots were excised from the gel and subjected to in gel digestion with Achromobacter protease I (lysyl endopeptidase). Resulting peptides were analyzed by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) followed by peptide mass fingerprinting for protein identification. With this approach we have obtained a two-dimensional electrophoresis (2-DE) protein map in which 69 spots were localized as landmarks for comparison of expression profiles to elucidate the basis of various biological events. PMID- 10870963 TI - Effect of heat stress on tomato fruit protein expression. AB - We delayed the ripening of tomato fruit for several days (average 5 days) by a 1 day heat treatment at 37-42 degrees C. We analyzed the tomato fruit pericarp proteins, which were altered by the heat stress, using two-dimensional electrophoresis. Heat stress caused about 23.7% of the proteins in the pericarp to disappear and about 1.1% of new proteins to appear. We determined their apparent molecular mass, isoelectric point, and N-terminal amino acid sequence. Identified proteins included antioxidant enzymes, heat shock proteins, cell-wall related proteins, etc. PMID- 10870964 TI - Proteomic analysis of colonic crypts from normal, multiple intestinal neoplasia and p53-null mice: a comparison with colonic polyps. AB - In order to observe cellular changes caused by mutation of the tumor suppressors, APC and p53, we have generated protein expression profiles of mouse colon epithelial cells using two-dimensional electrophoresis (2-DE). Crypts, polyps and stroma were isolated from normal, multiple intestinal neoplasia (MIN) and p53 null mice, each with a C57Black/6J background, and subjected to 2-DE in two separate pH ranges (pH 3-10 and pH 6-11). No significant differences in protein expression patterns were observed between the normal, MIN and p53-null colon epithelial crypts. However, 64 proteins from the MIN polyps showed a 2-fold or greater difference in intensity that was statistically significant as assessed by the Wilcoxon rank-sum test (p < or = 0.05). Of these, calreticulin, carbonic anhydrase I and a new member of the glutathione-S-transferase theta family of proteins have so far been identified using an in-gel digestion protocol coupled with reversed-phase high performance liquid chromatography (RP-HPLC) ion-trap mass spectrometry. In addition, 38 marker proteins have been identified in a continuing effort to generate a comprehensive 2-DE database of proteins expressed by mouse colon epithelial cells (these databases are available at http://www.ludwig.edu.au/jpsl/jpslhome. html). PMID- 10870965 TI - Proteomic analysis of the small intestine and colon epithelia of adenomatous polyposis coli gene-mutant mice by two-dimensional gel electrophoresis. AB - Mutations of the adenomatous polyposis coli gene (APC) have been implicated in the occurrence of sporadic colon cancer. Various APC mutant strains of mice have been created to better understand the function of this gene. Previously, we had mice express a mutant form of mRNA of the APC protein that encoded 474 amino acids instead of the 2845 amino acids due to exon duplication. These APC mutant mice (APC delta 474) developed intestinal and mammary tumors, as have other APC mutant mice previously reported (Sasai, H., et al. Carcinogenesis, in press). To elucidate the mechanism of the tumor development, we prepared protein samples from both normal and tumor tissues from APC delta 474 mutant mice, as well as tissues from normal mice, and used them for proteomic analysis. After two dimensional electrophoresis, the gels were silver stained and the protein spots were analyzed. We analyzed about 1000 protein spots per sample and found several protein spots that are specific for normal or tumor samples from APC delta 474 mutant mice, as well as proteins with altered expression levels. Among the identified protein spots, truncated beta-tubulins were specific to APC delta 474 mutant mice polyp samples. The apparent molecular mass of these proteins suggested that these beta-tubulins may be truncated very close to the binding site of the anti-tumor drug taxol. PMID- 10870966 TI - Two-dimensional electrophoresis map of the human hepatocellular carcinoma cell line, HCC-M, and identification of the separated proteins by mass spectrometry. AB - Currently, one of the most popular applications of proteomics is in the area of cancer research. In Africa, Southeast Asia, and China, hepatocellular carcinoma is one of the most common cancers, occurring as one of the top five cancers in frequency. This project was initiated with the purpose of separating and identifying the proteins of a human hepatocellular carcinoma cell line, HCC-M. After two-dimensional gel electrophoresis separation, silver staining, matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analyses, tryptic peptide masses were searched for matches in the SWISS-PROT and NCBI nonredundant databases. Approximately 400 spots were analyzed using this approach. Among the proteins identified were housekeeping proteins such as alcohol dehydrogenase, alpha-enolase, asparagine synthetase, isocitrate dehydrogenase, and glucose-6-phosphate 1-dehydrogenase. In addition, we also identified proteins with expression patterns that have been postulated to be related to the process of carcinogenesis. These include 14-3-3 protein, annexin, prohibitin, and thioredoxin peroxidase. This study of the HCC-M proteome, coupled with similar proteome analyses of normal liver tissues, tumors, and other hepatocellular carcinoma cell lines, represents the first step towards the establishment of protein databases, which are valuable resources in studies on the differential protein expressions of human hepatocellular carcinoma. PMID- 10870967 TI - Proteomic analysis of Epstein-Barr virus-transformed human B-lymphoblastoid cell lines before and after immortalization. AB - Infection of human B lymphocytes with Epstein-Barr virus (EBV) induces proliferative B-lymphoblastoid cell lines (LCLs). However, the majority of EBV transformed LCLs are mortal and unable to avoid cellular senescence. In our previous experiment, three immortalized LCLs were established by passages of EBV transformed LCLs for nearly five years accompanied by strong telomerase activity. In the present study, proteomic profiles of these three LCLs were analyzed comparatively at the early and the late passages of cell culture, and a protein spot was found which most significantly decreased with the immortalization in two LCLs. The expression of the protein in the third LCL was suppressed at 17 population doubling level (PDL), already suggesting that part of the immortalization process had been initiated before 17 PDL. The protein was assigned to ssp7001 (16.3 kDa, pI 6.0) by referring to our TMIG-2DPAGE proteome database. The protein was transferred onto a polyvinylidene difluoride (PVDF) membrane and digested with lysilendopeptidase to perform peptide mass fingerprinting by nanoelectrospray ionization mass spectrometry (nano-ESI-MS). Subsequent MS-Fit database search indicated that ssp7001 is a phosphoprotein stathmin. This speculation was confirmed by the tandem MS (MS/MS) analysis in a Q Tof system and by Edman degradation microsequencing. PMID- 10870969 TI - Analysis of blood plasma proteins in patients with Alzheimer's disease by two dimensional electrophoresis, sequence homology and immunodetection. AB - Blood plasma proteins of patients with Alzheimer's disease (AD; senile dementia) and non-AD-type dementia were resolved by two-dimensional electrophoresis and identified by migration position in the electrophoresis pattern, sequence homology, and immunodetection by using antibodies. For the control experiments, blood plasma proteins of a healthy young individual and non-dementia patients were examined in a manner similar to that of the plasma samples of AD patients. In the plasma sample of the healthy young individual, more than 350 spots of silver-stained proteins were observed and among these spots, 73 spots were identified. Blood plasma proteins of the AD and non-AD-type dementia patients were compared with those of the control and non-dementia patients. In the blood plasma samples of five AD patients, three patients had apolipoprotein E4, and another patient showed apolipoprotein L and complement factor H. For the AD related proteins apolipoprotein E, tau-1, and presenilin 2, proteins were examined by immunostaining with antibodies, in both AD and non-AD patients. Among the three samples of non-AD-type dementia patients, one was distinguishable by amyloid A proteins, and the other by haptoglobin isoforms. PMID- 10870968 TI - Comprehensive analyses of prostate gene expression: convergence of expressed sequence tag databases, transcript profiling and proteomics. AB - Several methods have been developed for the comprehensive analysis of gene expression in complex biological systems. Generally these procedures assess either a portion of the cellular transcriptome or a portion of the cellular proteome. Each approach has distinct conceptual and methodological advantages and disadvantages. We have investigated the application of both methods to characterize the gene expression pathway mediated by androgens and the androgen receptor in prostate cancer cells. This pathway is of critical importance for the development and progression of prostate cancer. Of clinical importance, modulation of androgens remains the mainstay of treatment for patients with advanced disease. To facilitate global gene expression studies we have first sought to define the prostate transcriptome by assembling and annotating prostate derived expressed sequence tags (ESTs). A total of 55000 prostate ESTs were assembled into a set of 15953 clusters putatively representing 15953 distinct transcripts. These clusters were used to construct cDNA microarrays suitable for examining the androgen-response pathway at the level of transcription. The expression of 20 genes was found to be induced by androgens. This cohort included known androgen-regulated genes such as prostate-specific antigen (PSA) and several novel complementary DNAs (cDNAs). Protein expression profiles of androgen stimulated prostate cancer cells were generated by two-dimensional electrophoresis (2-DE). Mass spectrometric analysis of androgen-regulated proteins in these cells identified the metastasis-suppressor gene NDKA/nm23, a finding that may explain a marked reduction in metastatic potential when these cells express a functional androgen receptor pathway. PMID- 10870970 TI - Alteration of protein composition in mouse thymocytes by signals through T-cell receptor. AB - To avoid destructive autoimmunity, T-cell precursors (thymocytes) expressing autoreactive T-cell receptor are deleted in the thymus via an apoptotic process by the signals from the T-cell receptor-CD3 complexes. In order to analyze the apoptotic mechanism, we established a cell-free system using the lysates from mouse thymocytes treated in vivo with anti-CD3 monoclonal antibody (mAb). The soluble cytosolic high molecular mass protein fraction from the anti-CD3-treated thymocytes revealed an activity that directly induces nuclear apoptotic morphological changes and DNA fragmentation. This fragmentation activity was not observed in the fraction from the thymocytes without anti-CD3 treatment. Proteins in both fractions were separated by two-dimensional electrophoresis. The silver stained gels revealed differences in protein spots. These protein spots were identified by database searching of mass spectrometric (MS) and tandem mass spectrometric (MS/MS) data obtained from in-gel tryptic digests of the spots, using an integrated system of liquid chromatography/electrospray ionization/ion trap mass spectrometry. In this study, the high mobility group protein HMG2 was identified as one of the cytosolic proteins that is increased by the signals from the T-cell receptor, and heterogeneous nuclear ribonucleoprotein A2/B1 and glyceraldehyde 3-phosphate dehydrogenase were found to be decreased by the signals. PMID- 10870971 TI - Proteome analysis of mouse brain: two-dimensional electrophoresis profiles of tissue proteins during the course of aging. AB - Mouse brain proteins were isolated from five regions (cerebellum, cerebral cortex, hippocampus, striatum, and cervical spinal cord) at five ages from the 10th week to the 24th month, and separated by two-dimensional gel electrophoresis (2-DE). 2-DE was carried out with an immobilized pH gradient bar in the first dimension, and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the second dimension. Over one thousand protein spots were visualized by silver staining and quantified by image processing. In the analyses, 58 protein spots were distinguishable among the above five brain regions, and 17 proteins were shown to be varied in quantity in the course of aging. Partial amino-terminal sequences and/or internal sequences for a total of 301 protein spots were analyzed. One hundred and eighty proteins appeared to have blocked N-termini and 122 proteins were identified. Twenty-seven new proteins were identified by sequence homology search. A mouse brain proteome database was constructed, which consists of the 2-DE map images and the respective spot data files with 15 related references. PMID- 10870972 TI - Protein profiling of rat cerebella during development. AB - Protein profiles of developing rat cerebella were analyzed by means of two dimensional gel electrophoresis (2-DE) and mass spectrometry (MS). The analysis of adult rat cerebellum gave rise to a protein map comprising approximately 3000 spots detectable by silver staining following high resolution 2-DE with a pH range of 3-10 and a mass range of 8-100 kDa. To obtain landmarks for comparison of developmental profiles of cerebellar proteins, 100 spots were subjected to peptide mass fingerprinting using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), and 67 spots were assigned on the map. Analysis of profiles of the developing cerebella revealed significant changes in the expression of proteins during development. In most cases the expression levels of proteins increased as the cerebellum matured, while the expression of 42 spots appeared specific or remarkably abundant in the immature cerebellum. Peptide mass fingerprinting of these spots allowed us to identify 29 proteins, which include, in addition to proteins of unknown function, many proteins known to have roles in the development of the central nervous system. These results suggest that the proteomic approach is valuable for mass identification of proteins involved in cerebellar morphogenesis. PMID- 10870973 TI - Comparative analysis of brain proteins from p53-deficient mice by two-dimensional electrophoresis. AB - p53 is a tumor suppressor protein that regulates many cellular processes including the cell cycle, DNA repair, and apoptosis. It also serves as a critical regulator of neuronal apoptosis in the central nervous system (CNS). To elucidate the role of p53 in the CNS, brain proteins of p53 knock-out mice (p53-/-) were analyzed by two-dimensional gel electrophoresis (2-DE) and compared with those from p53 wild type (p53+/+) mice. Six types of brain tissue (temporal cortex, cerebellum, hippocampus, striatum, olfactory bulb, and cervical spinal cord) and other control tissues (lung and blood) from 18-week-old non-stress-induced mice were analyzed. The morphology of brains from p53-/- mice appeared to be normal and identical to that of p53+/+ mice, although lungs showed diffuse tumors that may have been caused by p53 deficiency. Comparative 2-D gel analysis showed that, on average, 7 of 886 spots from brain tissue were p53-/- specific, whereas 12 of 1008 spots from lung tissue were p53-/- specific. N-terminal amino acid sequence was determined for p53-/- specific proteins. In all brain tissues from p53-/- mice, a newly identified mouse mitochondrial NADH-ubiquinone oxidoreductase 24 kDa subunit showed decreased expression, and apolipoprotein A1 acidic forms showed increased expression. In addition, brain-type creatine kinase B chain and tubulin beta-5 N-terminal fragment were increased in the p53-/- cerebellum, and a new protein in mouse, hydroxyacylglutathione hydrolase (glyoxalase II) was decreased in the temporal cortex of p53-/- mice. The alterations in protein expression identified in this study may imply a p53-related brain function. This is the first proteomic analysis on the p53-/- mouse brain, and further information based on this study will provide new insights into the p53 function in the CNS. PMID- 10870974 TI - Matrix assisted laser desorption/ionization-time of flight-mass spectrometry analysis of proteins detected by anti-phosphotyrosine antibody on two-dimensional gels of fibrolast cell lysates after tumor necrosis factor-alpha stimulation. AB - We describe efficient methods for using functional proteomics analysis to study signal transduction pathways in murine fibroblast L929 cells following stimulation with tumor necrosis factor (TNF)-alpha. After stimulation with TNF alpha, cellular proteins of L929 cells were extracted with a lysis buffer containing 0.3% sodium dodecyl sulfate (SDS) for 10-30 min time intervals, and were separated by two-dimensional (2-D) electrophoresis followed by immunoblot analysis with anti-phosphotyrosine antibody and alkaline phosphatase-anti IgG antibody conjugate. To improve detection sensitivity by immunoblot analysis we used a chemifluorescent substrate for alkaline phosphatase. One hundred protein spots were detected in the TNF-alpha stimulated L929 cell extract by immunoblot analysis. The use of chemifluorescence allowed us to quantitate immunoblotted spots with fluoroscanner so that we were able to detect time-dependent changes of a number of immunoblotted spots. Protein spots on a silver-stained 2-D gel corresponding to those detected by immunoblot analysis were subjected to in-gel trypsin digestion- matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF)-mass spectrometry analysis, respectively. Twenty-one proteins detected by immunoblot analysis were identified by MS-Fit database search analysis. Among them, the proteins that show time-dependent changes in staining intensity include vimentin, tubulin beta-chain, eukaryotic translation initiation factor 1A, chromatin assembly factor 1 (P48 subunit), probable protein disulfide isomerase P5, and several other proteins. Vimentin and tubulin beta-chain have been reported to be phosphorylated at tyrosine residues and involved in the signal transduction pathway induced by TNF-alpha. However, the other proteins have no previously known function in the signal transduction pathway. Thus, the methods used in this study seem to be suitable for the identification of time dependent changes in many proteins that are involved in signal transduction. Usefulness of the method for comprehensive analysis of the proteins involved in signal transduction pathway and the limitations of the method are discussed. PMID- 10870975 TI - Application of proteomics for determining protein markers for wool quality traits. AB - The technique of two-dimensional electrophoresis (2-DE) has been under investigation for its usefulness in identifying protein markers for wool quality traits in sheep. However, before this could be achieved, unique problems relating to the detection and quantitation of wool proteins needed to be overcome so that 2-DE protein maps could be examined using computational programs like Melanie II. Four protein staining regimes were examined. Colloidal Coomassie Blue G-250 was found to be superior to Coomassie Blue R-250 and gave satisfactory staining of all protein classes. Silver staining detects minor strings of keratinous proteins, but unfortunately it negatively stains intermediate filament proteins, the major high sulphur proteins (HSPs) and the high glycine tyrosine proteins and the latter two classes can only be seen by overstaining the background of the gel. In contrast, labeling reduced keratins with [14C]iodoacetamide, followed by autoradiography detection, results in a protein map with low background and all protein spots stained positively. 2-DE has been used to obtain wool protein maps of Lincoln/Merino chimeric sheep to examine wool originating from two genotypes grown with different crimp frequencies within the same fleece. Between fleece, variations have also been examined. Work to date suggests that several major HSPs may be associated with the fibre curvature trait known as crimp frequency. From matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectral mapping, one of these proteins has been identified as being from the B2A family from the HSP class. PMID- 10870976 TI - Proteome analysis of Oncorhynchus species during embryogenesis. AB - To understand the molecular mechanisms underlying normal and abnormal development of two salmonids, masu salmon (Oncorhynchus masou) and rainbow trout (O. mykiss), we used two-dimensional (2-D) electrophoresis to construct a series of 2-D maps during the embryonic period. We identified all visible protein spots on the 2-D map by assigning numbers for masu salmon and rainbow trout, and we determined N terminal sequences of proteins for one hundred of the spots, that appear at very high concentrations in the whole embryos of masu salmon and rainbow trout. We also characterized embryonic stages according to the periods of appearance of spots. Most of the N-terminal sequences were identical or at least highly similar to partial sequences reported for vitellogenin (Vtg) of O. mykiss. A potential proteolytic processing of Vtg for rainbow trout is discussed in relation to the time of appearance and relative position of Vtg fragments within the complete protein sequence. PMID- 10870977 TI - Natural and biotech-derived therapeutic proteins: what is the future? AB - A myriad of novel proteins and ligands of unknown function will be generated by the Human Genomic Project. Due to differences in post-translational processing, proteins produced by recombinant DNA technology may not possess proper biological activity. One way to find their function is to search for their natural counterparts. Proteins are produced in the tissues, and many of them are secreted into plasma and excreted into urine. There is a virtually "unlimited" array of human proteins in our plasma and urine, many of them in a fully active form. They include small molecules like steroids, peptides, and large glycoproteins like human menopausal gonadotropin. A library of plasma and urinary proteins could be developed to serve as a reference for the novel proteins generated by the functional genomic projects. PMID- 10870978 TI - Subject-by-formulation interaction in bioequivalence: conceptual and statistical issues. FDA Population/Individual Bioequivalence Working Group. Food and Drug Administration. AB - PURPOSE: The FDA has proposed replacing the current average bioequivalence criterion with population and individual bioequivalence criteria that consider variances in addition to the difference of averages. One of these variances in the individual bioequivalence criterion measures subject-by-formulation interaction, the extent to which the test-reference difference varies from person to person. This paper discusses conceptual and statistical issues raised in various publications and presentations with respect to the presence and estimation of such an interaction. METHODS: We focus on the importance of subject by-formulation interaction, an understanding of what is a large interaction, and the assessment of the magnitude of this interaction in bioequivalence studies. Simulation studies, examples from the literature, and data from FDA files are used to demonstrate the magnitude of the interaction and its distribution under various conditions. RESULTS: The concept of a large interaction is tied to the concept of a large mean difference. We suggest that an interaction greater than 0.15 is a conservative criterion for a large interaction. Magnitudes of estimated interaction are affected by variability, sample size, and the selection of data sets that pass average bioequivalence. CONCLUSIONS: Examples of substantial interactions are beginning to appear. More data is needed before reaching definitive conclusions regarding the frequency and importance of observed interactions. PMID- 10870979 TI - Bioequivalence of methylphenidate immediate-release tablets using a replicated study design to characterize intrasubject variability. AB - PURPOSE: To determine the relative bioavailability of two marketed, immediate release methylphenidate tablets. The study used a replicated study design to characterize intrasubject variability, and determine bioequivalence using both average and individual bioequivalence criteria. METHODS: A replicated crossover design was employed using 20 subjects. Each subject received a single 20 mg dose of the reference tablet on two occasions and two doses of the test tablet on two occasions. Blood samples were obtained for 10 hr after dosing, and plasma was assayed for methylphenidate by GC/MS. RESULTS: The test product was more rapidly dissolved in vitro and more rapidly absorbed in vivo than the reference product. The mean Cmax and AUC(0-infinity) differed by 11% and 9%, respectively. Using an average bioequivalence criterion, the 90% confidence limits for the Ln transformed Cmax and AUC(0-infinity), comparing the two replicates of the test to the reference product, fell within the acceptable range of 80-125%. Using an individual bioequivalence criterion the test product failed to demonstrate equivalence in Cmax to the reference product. CONCLUSIONS: The test and reference tablets were bioequivalent using an average bioequivalence criterion. The intrasubject variability of the generic product was greater and the subject-by formulation interaction variance was borderline high. For these reasons, the test tablets were not individually bioequivalent to the reference tablets. PMID- 10870980 TI - Pharmacokinetics of a hematoregulatory peptide (SK&F107647) in healthy male volunteers and in patients with colorectal or pancreatic adenocarcinoma not amenable to standard therapy. AB - PURPOSE: To describe the pharmacokinetics of SK&F 107647, a synthetic hematoregulatory peptide, in healthy volunteers and in patients with adenocarcinoma. METHODS: SK&F 107647 pharmacokinetics were evaluated in 2 dose escalation studies. Volunteers received SK&F 107647 as single 15-minute iv infusion doses of 1, 10, 100, 500, and 1,000 microg/kg. Cancer patients received 2-hour iv infusions of 0.001, 0.01, 0.1 and 1 microg/kg once daily for 10 days. Drug concentrations were quantified in plasma and urine of healthy volunteers and on days 1 and 10 in plasma of cancer patients receiving the two top dose levels. RESULTS: In volunteers, mean clearance (CL) ranged from 76.7 to 101 ml/hour/kg; mean volume of distribution at steady-state (Vss) ranged from 175 to 268 ml/kg. Most of the administered dose was renally excreted as intact peptide within 24 hours postinfusion. In patients, mean CL was 57.6 ml/hour/kg, mean Vss ranged from 128 to 150 ml/kg and terminal half-life from 2.1 to 3.4 hours. There was little accumulation of drug. In both studies, linear pharmacokinetics was observed. Clearance approached normal glomerular filtration rate (GFR) in volunteers and correlated with creatinine clearance in cancer patients. CONCLUSIONS: SK&F 107647 exhibits linear pharmacokinetics, a small Vss, and clearance, primarily renal, approaching normal GFR. PMID- 10870981 TI - Stability of human serum albumin during bioprocessing: denaturation and aggregation during processing of albumin paste. AB - PURPOSE: To assess the impact of various bioprocessing steps on the stability of freshly precipitated human serum albumin (HSA) obtained from pooled human plasma. METHODS: After initial precipitation of HSA from plasma, the resultant paste is either (a) lyophilized or (b) washed with acetone and then air-dried in order to obtain a dry powder. The structure of HSA was examined using Fourier transform infrared (IR) spectroscopy. The extent of aggregation of redissolved HSA was measured using both dynamic light scattering and SDS-polyacrylamide gel electrophoresis (SDS-PAGE). RESULTS: Both lyophilization and air-drying perturb the secondary structural composition of HSA, as detected by infrared (IR) spectroscopy. Upon dissolution of dried paste, most of the protein refolds to a native-like conformation. However, a small fraction of the protein molecules form soluble aggregates that can be detected by both dynamic light scattering and SDS PAGE. The level of aggregation is so low that it could not be detected in the bulk by either circular dichroism or IR spectroscopy. The lyophilized protein, which appears to be more unfolded in the solid state than the acetone washed/air dried material, exhibits a higher level of aggregation upon dissolution. CONCLUSIONS: There is a direct correlation between the extent of unfolding in the solid state and the amount of soluble aggregate present after dissolution. Moreover, the presence of the aggregates persists throughout the remainder of the purification process, which includes dissolution, chromatography, sterile filtration and viral inactivation steps. Analytical methods used to monitor the stability of biopharmaceuticals in the final product can be used to assess damage inflicted during processing of protein pharmaceuticals. PMID- 10870982 TI - What is the true solubility advantage for amorphous pharmaceuticals? AB - PURPOSE: To evaluate the magnitude of the solubility advantage for amorphous pharmaceutical materials when compared to their crystalline counterparts. METHODS: The thermal properties of several drugs in their amorphous and crystalline states were determined using differential scanning calorimetry. From these properties the solubility advantage for the amorphous form was predicted as a function of temperature using a simple thermodynamic analysis. These predictions were compared to the results of experimental measurements of the aqueous solubilities of the amorphous and crystalline forms of the drugs at several temperatures. RESULTS: By treating each amorphous drug as either an equilibrium supercooled liquid or a pseudo-equilibrium glass, the solubility advantage compared to the most stable crystalline form was predicted to be between 10 and 1,600 fold. The measured solubility advantage was usually considerably less than this, and for one compound studied in detail its temperature dependence was also less than predicted. It was calculated that even for partially amorphous materials the apparent solubility enhancement (theoretical or measured) is likely to influence in-vitro and in-vivo dissolution behavior. CONCLUSIONS: Amorphous pharmaceuticals are markedly more soluble than their crystalline counterparts, however, their experimental solubility advantage is typically less than that predicted from simple thermodynamic considerations. This appears to be the result of difficulties in determining the solubility of amorphous materials under true equilibrium conditions. Simple thermodynamic predictions can provide a useful indication of the theoretical maximum solubility advantage for amorphous pharmaceuticals, which directly reflects the driving force for their initial dissolution. PMID- 10870983 TI - Photomechanical drug delivery into bacterial biofilms. AB - PURPOSE: To investigate whether photomechanical waves generated by lasers can increase the permeability of a biofilm of the oral pathogen Actinomyces viscosus. METHODS: Biofilms of Actinomyces viscosus were formed on bovine enamel surfaces. The photomechanical wave was generated by ablation of a target with a Q-switched ruby laser and launched into the biofilm in the presence of 50 microg/ml methylene blue. The penetration depth of methylene blue was measured by confocal scanning laser microscopy. Also, the exposed biofilms were irradiated with light at 666 nm. After illumination, adherent bacteria were scraped and spread over the surfaces of blood agar plates. Survival fractions were calculated by counting bacterial colonies. RESULTS: Confocal scanning laser microscopy revealed that a single photomechanical wave was sufficient to induce a 75% increase in the penetration depth of methylene blue into the biofilm. This significantly increased the concentration of methylene blue in the biofilm enabling its photodestruction. CONCLUSIONS: Photomechanical waves provide a potentially powerful tool for drug delivery that might be utilized for treatment of microbial infections. PMID- 10870984 TI - Allometric pharmacokinetic scaling: towards the prediction of human oral pharmacokinetics. AB - PURPOSE: To evaluate (1) allometric scaling of systemic clearance (CL) using unbound drug concentration, (2) the potential usage of brain weight (BRW) correction in allometric scaling of both CL and oral clearance (CL/F). METHODS: Human clearance was predicted allometrically (CLu = a x W(biv)) using unbound plasma concentration for eight Parke-Davis compounds and 29 drugs from literature sources. When the exponent b(iv) was higher than 0.85, BRW was incorporated into the allometric relationship (CLu*BRW = a x W(biv)). This approach was also applied to the prediction of CLu/F for 10 Parke-Davis compounds. Human oral t1/2, Cmax, AUC, and bioavailability were estimated based on allometrically predicted pharmacokinetic (PK) parameters. RESULTS: Human CL and CL/F were more accurately estimated using unbound drug concentration and the prediction was further improved when BRW was incorporated into the allometric relationship. For Parke Davis compounds, the predicted human CL and CL/F were within 50-200% and 50-220% of the actual values, respectively. The estimated human oral t1/2, Cmax, and AUC were within 82-220%, 56-240%, and 73-190% of the actual values for all 7 compounds, suggesting that human oral PK parameters of those drugs could be reasonably predicted from animal data. CONCLUSIONS: Results from the retrospective analysis indicate that allometric scaling of free concentration could be applied to orally administered drugs to gain knowledge of drug disposition in man, and to help decision-making at early stages of drug development. PMID- 10870985 TI - Prediction of in vivo interaction between triazolam and erythromycin based on in vitro studies using human liver microsomes and recombinant human CYP3A4. AB - PURPOSE: To quantitatively predict the in vivo interaction between triazolam and erythromycin, which involves mechanism-based inhibition of CYP3A4, from in vitro studies using human liver microsomes (HLM) and recombinant human CYP3A4 (REC). METHODS: HLM or REC was preincubated with erythromycin in the presence of NADPH and then triazolam was added. alpha- and 4-hydroxy (OH) triazolam were quantified after a 3 min incubation and the kinetic parameters for enzyme inactivation (k(inact) and K('app)) were obtained. Drug-drug interaction in vivo was predicted based on a physiologically-based pharmacokinetic (PBPK) model, using triazolam and erythromycin pharmacokinetic parameters obtained from the literature and kinetic parameters for the enzyme inactivation obtained in the in vitro studies. RESULTS: Whichever enzyme was used, triazolam metabolism was not inhibited without preincubation, even if the erythromycin concentration was increased. The degree of inhibition depended on preincubation time and erythromycin concentration. The values obtained for k(inact) and K('app) were 0.062 min(-1) and 15.9 microM (alpha-OH, HLM), 0.055 min(-1) and 17.4 microM (4-OH, HLM), 0.173 min(-1) and 19.1 microM (alpha-OH, REC), and 0.097 min(-1) and 18.9 microM (4-OH, REC). Based on the kinetic parameters obtained using HLM and REC, the AUCpo of triazolam was predicted to increase 2.0- and 2.6-fold, respectively, following oral administration of erythromycin (333 mg t.i.d. for 3 days), which agreed well with the reported data. CONCLUSIONS: In vivo interaction between triazolam and erythromycin was successfully predicted from in vitro data based on a PBPK model involving a mechanism-based inhibition of CYP3A4. PMID- 10870986 TI - The stereoselective distribution of halofantrine enantiomers within human, dog, and rat plasma lipoproteins. AB - PURPOSE: To study the in vitro distribution of the enantiomers of the antimalarial drug halofantrine in human, dog and rat plasma lipoprotein fractions. METHODS: Plasma was spiked with racemic halofantrine (1,000 ng/ml) and incubated for 1 h at 37 degree C. The fractions (high and low density lipoproteins, triglyceride-rich lipoproteins and lipoprotein deficient plasma) were separated using density gradient ultracentrifugation. Fractions were assayed for halofantrine enantiomer using stereospecific high performance liquid chromatography. RESULTS: The (-) enantiomer of halofantrine displayed higher affinity for the lipoprotein-deficient fraction than the (+) enantiomer in all three species. The (+) enantiomer was predominately located in the lipoprotein rich fractions of dog and human plasma (the (+):(-) ratio ranging from 1.2-9.6). In contrast, the (+):(-) ratio was consistently < 1 in lipoprotein-deficient fractions. Dog displayed a large magnitude of stereoselectivity in halofantrine distribution to the plasma fractions tested. There were substantial interspecies differences in the pattern of distribution of halofantrine enantiomers within the different fractions. A significant positive relationship was observed between halofantrine uptake into lipoprotein-rich fractions and the percent of apolar core lipid in those fractions. There was also a strong negative correlation between total protein concentration and the enantiomeric ratio in the lipoprotein deficient plasma fraction. CONCLUSION: Distribution of halofantrine enantiomer to plasma lipoprotein-fractions is stereoselective and species specific. This differential binding of halofantrine enantiomers to lipoproteins may need to be considered in viewing pharmacokinetic and pharmacodynamic data involving the drug. PMID- 10870987 TI - Pharmacokinetic study of the hepatobiliary transport of indomethacin. AB - PURPOSE: The biliary excreted amount of indomethacin and its glucuronide is related to the intestinal toxicity of this drug. In the present study, we investigated the hepatobiliary transport of indomethacin. METHODS: The uptake of indomethacin into primary cultured rat hepatocytes and COS-7 cells transfected with cDNA encoding sodium taurocholate co-transporting polypeptide or organic anion transporting polypeptide 1 was examined. Moreover, we compared the biliary excretion of indomethacin and its glucuronide between Sprague-Dawley (SD) rats and Eisai hyperbilirubinemic rats (EHBR) whose canalicular multispecific organic anion transporter/multidrug resistance associated protein 2 (cMOAT/MRP2) function is hereditarily defective. RESULTS: The uptake of indomethacin into rat hepatocytes was mediated by Na+-dependent and independent active transport systems. Neither transfectant stimulated the uptake of indomethacin. After intravenous infusion of indomethacin to SD rats, the biliary excretion of indomethacin glucuronide exceeded that of indomethacin. The indomethacin transport clearance across the bile canalicular membrane was comparable between SD rats and EHBR, whereas the corresponding value for indomethacin glucuronide in EHBR was approximately 50% that in SD rats. CONCLUSIONS: These results indicate that another transporter(s) is involved in the hepatic uptake of indomethacin and the canalicular transport of indomethacin glucuronide is mediated by cMOAT/MRP2 whereas that of indomethacin is not mediated by cMOAT/MRP2. PMID- 10870988 TI - Dissolution testing as a prognostic tool for oral drug absorption: dissolution behavior of glibenclamide. AB - PURPOSE: The dissolution behavior of two commercially available glibenclamide formulations was tested in various media. The aim of the study was to investigate whether the use of biorelevant dissolution media (BDM) would be advantageous over the use of standard media for predicting the in vivo performance of the two formulations. METHODS: The dissolution tests were performed using USP 23 apparatus 2. Conventional buffers and USP media were compared with two BDM containing different amounts of lecithin and sodium taurocholate. RESULTS: The dissolution of two drug powders was highly dependent on wetting, particle size, pH, and the composition of the medium used. In addition, the dissolution behavior of the two glibenclamide formulations showed differences in all media tested. The dissolution results of the two formulations were compared with those from an in vivo bioequivalence study undertaken by the central quality control laboratory of the German pharmacists (ZL). The bioequivalence criterion set by the ZL requires more than 80% drug release within 10 minutes. Results in FaSSIF, one of the BDMs, met the ZL criterion and this medium was also able to discriminate between the two formulations. This was not the case for the other media tested. CONCLUSIONS: The study indicates that BDM are better able to discriminate between glibenclamide formulations than standard dissolution media. PMID- 10870989 TI - Effect of the concurrent LHRH antagonist administration with a LHRH superagonist in rats. AB - PURPOSE: The purpose of this study was to investigate the effect of a novel LHRH antagonist, Orntide acetate, on the initial testosterone elevation in rats during treatment with a LHRH superagonist, Leuprolide acetate. METHODS: Thirteen groups of a rat animal model were administered either liquid Orntide or Orntide PLGA microspheres before or simultaneously with Leuprolide injections. Serum levels of testosterone were monitored during the time course of the study using a radioimmunoassay method. RESULTS: Administration of a single daily dose of liquid Orntide resulted in testosterone suppression within 6 h to levels below 0.5 ng/ml (castration level). However, combined administration of liquid Orntide and liquid Leuprolide did not have a significant effect on the initial testosterone elevation in studied rats. Similarly, there was no effect when liquid Orntide was co-administered with Leuprolide microspheres. Administration of Orntide microspheres 48 h before Leuprolide microspheres suppressed testosterone levels below the castration level within 24 h, however, did not prevent a rise in testosterone serum concentration upon administration of Leuprolide microspheres. Also, a second testosterone peak was observed between days 3 and 15 in the animals which were simultaneously treated with Orntide microspheres and Leuprolide microspheres. CONCLUSIONS: Orntide acetate was found to be an effective LHRH antagonist with a rapid onset of pharmacological action and a short biological half-life. Administration of a single dose of liquid Orntide or Orntide microspheres, resulted in rapid testosterone suppression without an initial elevation, as seen with LHRH superagonists. However, combined administration of Orntide and Leuprolide did not have an effect on the initial testosterone elevation in rats. PMID- 10870990 TI - Synthetic glycopeptide-based delivery systems for systemic gene targeting to hepatocytes. AB - PURPOSE: To design, synthesize, and test synthetic glycopeptide-based delivery systems for gene targeting to hepatocytes by systemic administration. METHODS: All peptides were synthesized by the solid phase method developed using Fmoc chemistry on a peptide synthesizer. The binding of galactosylated peptides to HepG2 cells and accessibility of the galactose residues on particle surface was demonstrated by a competition assay using 125I-labeled asialoorosomucoid and RCA lectin agglutination assay, respectively. DNA plasmid encoding chloramphenicol acetyl transferase (CAT) gene was complexed with a tri-galactosylated peptide (GM245.3) or tri-galactosylated lipopeptide (GM246.3) in the presence of an endosomolytic peptide (GM225.1) or endosomolytic lipopeptide (GM227.3) to obtain DNA particles of 100-150 nm in size. The plasmid/peptide complexes were added to HepG2 cell cultures or intravenously administered by tail vein injection into normal mice or rats. Plasmid uptake and expression was quantified by qPCR and ELISA, respectively. RESULTS: Multiple antennary glycopeptides that have the ability to condense and deliver DNA plasmid to hepatocytes were synthesized and complexed with DNA plasmid to obtain colloidally stable DNA/peptide complexes. Addition of DNA/GM245.3/GM225.1 peptide complexes (1:3:1 (-/+/-)) to HepG2 cell cultures yielded CAT expression in transfected cells. The transfection efficiency was significantly reduced in the absence of galactose ligand or removal of endosomolytic peptide. Intravenous administration of DNA/GM245.3 peptide complexes (1:0.5 (-/+)) into the tail vein of normal rats yielded DNA uptake in the liver. Substitution of GM245.3 by galactosylated lipopeptide GM246.3 resulted in more stable DNA particles, and a 10-fold enhancement in liver plasmid uptake. CAT expression was detectable in liver following intravenous administration of DNA/GM246.3 complexes. Addition of endosomolytic lipopeptide GM227.3 into the complexes (DNA/ GM246.3/GM227.3 (1:0.5:1 (-/+/-))) yielded a 5-fold increase in CAT expression. Liver expression was 8-fold and 40-fold higher than lung and spleen, respectively, and localized in the hepatocytes only. The transfection efficiency in liver was enhanced by increasing DNA dose and injection volume. The plasmid uptake and expression in liver using DNA/GM246.3/GM227.3 complexes was 100-200-fold higher than DNA formulated in glucose. Tissue examination and serum biochemistry did not show any adverse effect of the DNA/GM246.3/ GM227.3 (1:0.5:1 (-/+/-)) complexes after intravenous delivery. CONCLUSIONS: Gene targeting to hepatocytes was achieved by systemic administration of a well-tolerated synthetic glycopeptide-based delivery system. The transfection efficiency of this glycopeptide delivery system was dependent on peptide structure, endosomolytic activity, colloidal particle stability, and injection volume. PMID- 10870991 TI - Formation of multilayers in the caco-2 cell culture model: a confocal laser scanning microscopy study. AB - PURPOSE: To introduce confocal laser scanning microscopy (CLSM) combined with digital image restoration to characterise Caco-2 cells under different culture conditions, and thus to define additional valid criteria for the optimisation of culture models. METHODS: Growth curves were established and transepithelial electrical resistance (TEER) measured for cells grown in EMEM or DMEM medium on Cyclopore membranes. Cytoskeleton, cell nuclei and tight junctions (TJ) were investigated by CLSM. RESULTS: Cultures reached a plateau of approximately 4.5 x 10(5) cells/cm2 after approximately 10 days. At the same time TEER reached 750 omega cm2. An irregular, fairly complete network of TJ was present at confluence (approximately 2 d). Between 15 and 30 days a regular TJ network was established. Cells formed mixed mono- and multilayers under most conditions with two exceptions: flat monolayers were observed on polycarbonate filters with EMEM and with the Biocoat intestinal epithelium differentiation environment system. In multilayers TJ were found in the upper as well as in the lower cell layers although the regular vertical polarity was disturbed. CONCLUSIONS: CLSM represents an important tool to investigate the cytoarchitecture of Caco-2 cells. 3D-analysis of confocal data gives important clues on the characteristics of cell layers and thus helps to validate optimisation strategies. PMID- 10870992 TI - Transdermal extraction of analytes using low-frequency ultrasound. AB - PURPOSE: Transdermal extraction of clinically relevant analytes offers a potentially non-invasive method of diagnostics. However, development of such a method is limited by the low skin permeability. In this paper, we report a potential method for non-invasive diagnostics based on ultrasonic skin permeabilization and subsequent extraction of interstitial fluid (ISF) across the skin. METHODS: In vivo experiments were performed using Sprague Dawley rats to assess ultrasound-induced skin permeabilization and subsequent extraction of various analytes. Serum and ISF concentrations of various analytes were measured. RESULTS: Application of low-frequency ultrasound rapidly increased skin permeability. Skin remained in a state of high permeability for at least three hours. During this period, application of vacuum extracted ISF across rat skin in vivo at a rate of 25.7 microl/cm2/hr. We measured concentrations of various analytes including glucose, albumin, calcium, urea, triglycerides, lactate, and dextran in transdermally extracted fluid. The composition of the fluid extracted transdermally is similar to that of ISF. CONCLUSIONS: Application of low frequency ultrasound allows skin permeabilization and extraction of ISF across the skin. PMID- 10870993 TI - Visualization and analysis of electroosmotic flow in hairless mouse skin. AB - PURPOSE: To identify the physiological structures in hairless mouse skin responsible for the generation of electroosmotic flow during iontophoresis. Also, to determine the effects of changing the pH of the contacting solution on the magnitude of electroosmotic flow in these structures. METHODS: Localized diffusive and iontophoretic fluxes of a neutral molecule, hydroquinone (HQ), across hairless mouse skin were quantified using scanning electrochemical microscopy (SECM). The iontophoretic flux was determined as a function of the direction of the applied current and pH of the contacting solution. RESULTS: SECM images of HQ transport recorded during iontophoresis at moderate current densities (+/-0.1 mA/cm2) demonstrate that electroosmotic flow is localized to hair follicles. The direction of flow is from anode to cathode at pH > 3.5 and from cathode to anode at pH <3.5. CONCLUSIONS: Electroosmotic flow through hair follicles is an efficient and controllable means of transporting small, electrically neutral molecules across hairless mouse skin. Transport through the appendages is sensitive to the pH of the solution in contact with the skin. The isoelectric point of hair follicles, pI, is estimated to be 3.5 from the dependence of electroosmotic flow on the solution pH. PMID- 10870994 TI - Iontophoresis-enhanced absorptive flux of polar molecules across intestinal tissue in vitro. PMID- 10870995 TI - Role of P-glycoprotein in ocular clearance of rhodamine 123 in rabbits. PMID- 10870996 TI - In vivo absorption properties of orally administered recombinant human interleukin-11. PMID- 10870997 TI - Solid state NMR investigations on nanosized carrier systems. PMID- 10870998 TI - Effect of the storage conditions on the tensile strength of tablets in relation to the enthalpy relaxation of the binder. PMID- 10870999 TI - Noninvasive visualization of serotonergically mediated pathophysiology. PMID- 10871000 TI - MR imaging of hypertrophic olivary degeneration: is there a role for metaanalysis? PMID- 10871001 TI - Etiology of congenital growth hormone deficiency. PMID- 10871002 TI - MDMA ("Ecstasy") and its association with cerebrovascular accidents: preliminary findings. AB - BACKGROUND AND PURPOSE: Abuse of the popular recreational drug "Ecstasy" (MDMA) has been linked to the occurrence of cerebrovascular accidents. It is known that MDMA alters brain serotonin (5-HT) concentrations and that brain postsynaptic 5 HT(2) receptors play a role in the regulation of brain microvasculature. Therefore, we used brain imaging to find out whether MDMA use predisposes one to cerebrovascular accidents by altering brain 5-HT neurotransmission. METHODS: The effects of MDMA use on brain cortical 5-HT(2A) receptor densities were studied using [(123)I]R91150 single-photon emission CT in 10 abstinent recent MDMA users, five former MDMA users, and 10 healthy control subjects. Furthermore, to examine whether changes in brain 5-HT(2A) receptor densities are associated with alterations in blood vessel volumes, we calculated relative cerebral blood volume maps from dynamic MR imaging sets in five MDMA users and six healthy control subjects. RESULTS: An analysis of variance revealed that mean cortical [(123)I]R91150 binding ratios were significantly lower in recent MDMA users than in former MDMA users and control subjects. This finding suggests down-regulation of 5-HT(2) receptors caused by MDMA-induced 5-HT release. Furthermore, in MDMA users, low cortical 5-HT(2) receptor densities were significantly associated with low cerebral blood vessel volumes (implicating vasoconstriction) and high cortical 5-HT(2) receptor densities with high cerebral blood vessel volumes (implicating vasodilatation) in specific brain regions. CONCLUSION: These findings suggest a relationship between the serotonergic system and an altered regulation of 5-HT(2) receptors in human MDMA users. MDMA users may therefore be at risk for cerebrovascular accidents resulting from alterations in the 5-HT neurotransmission system. PMID- 10871003 TI - Moyamoya-like vasculopathy from cocaine dependency. AB - We herein describe two cases of moyamoya vasculopathy occurring in two men who used alkaloidal cocaine for years. One patient presented with aneurysmal subarachnoid hemorrhage and one with infarction in both lobes. Particularly impressive was a significant degree of collateral development with lenticulostriate networks. PMID- 10871004 TI - CT angiography for the detection of cerebral vasospasm in patients with acute subarachnoid hemorrhage. AB - BACKGROUND AND PURPOSE: Digital subtraction angiography (DSA) is the standard of reference for detecting cerebral vasospasm after subarachnoid hemorrhage (SAH). CT angiography (CTA) is a relatively recent method for depicting the intracranial arterial vasculature. The purpose of this study was to compare CTA and DSA in the detection and quantification of cerebral vasospasm. METHODS: Seventeen patients with SAH underwent initial CTA with or without DSA and follow-up CTA and DSA. The follow-up CTA and DSA studies were performed within 24 hours of each other and 5 to 10 days after SAH. Maximum intensity projection images were produced for each CTA. Six arterial locations were examined for spasm: the suprasellar internal carotid artery (ICA), the M1 and M2 segments of the middle cerebral artery, the A1 and A2 segments of the anterior cerebral artery, and the basilar artery. Vasospasm was categorized as none, mild (<30% luminal reduction), moderate (30% to 50% reduction), or severe (>50% reduction). RESULTS: The overall correlation between CTA and DSA was 0.757, but was better for proximal than distal locations (0.88-1.00 versus 0.152-0.446). Agreement between CTA and DSA was greater for no spasm (92%) and severe spasm (100%) than for mild (57%) or moderate (64%) spasm. CTA was highly accurate for no spasm or severe spasm in proximal locations (96%, and 100%, respectively); it was less accurate (90% and 95%, respectively) for mild or moderate spasm in these locations. For distal locations, the accuracy for absent, mild, moderate, or severe spasm was 78%, 81%, 94%, and 100%, respectively. CONCLUSION: CTA is highly sensitive, specific, and accurate in detecting no spasm or severe cerebral vasospasm in proximal arterial locations; it is less accurate for detecting mild and moderate spasm in distal locations. PMID- 10871005 TI - Angiographic and clinical characteristics of patients with cerebral arteriovenous malformations associated with hereditary hemorrhagic telangiectasia. AB - BACKGROUND AND PURPOSE: Cerebral arteriovenous malformations (AVMs) are occasionally associated with hereditary hemorrhagic telangiectasia (HHT), which is characterized by the presence of multiple mucocutaneous telangiectasia, epistaxis, and familial inheritance. We analyzed the angiographic and clinical characteristics of patients with cerebral AVMs related to HHT. METHODS: Among 638 patients with cerebral AVMs, we identified 14 patients with HHT. The AVMs were classified as those with nidi of 1 cm or less (micro AVMs), those with nidi between 1 and 3 cm (small AVMs), and those of the fistulous type (arteriovenous fistulas [AVFs]). RESULTS: A total of 28 AVMs were found; seven of 14 patients had multiple AVMs. The 28 AVMs were categorized as 12 micro AVMs, eight small AVMs, and eight AVFs. All except one micro AVM were asymptomatic, whereas all small AVMs were symptomatic. Three of eight AVFs were asymptomatic. All 28 AVMs were located on the cortex. All micro AVMs and AVFs had single feeders and single draining veins, whereas the small AVMs had multiple feeders in all lesions and single draining veins in six of eight lesions. CONCLUSION: Multiple, cortical, micro AVMs or AVFs harboring single feeding arteries and single draining veins should raise clinical suspicion of HHT-related AVMs. PMID- 10871007 TI - Contrast-enhanced MR angiography: the effects of k-space truncation on luminal representation in a carotid artery phantom model. AB - Using carotid bifurcation phantom models with different degrees of stenoses, we evaluated the accuracy of vessel lumen representation on MR images obtained from the inverse Fourier transform of different k-space percentages. Our results show that the lower thresholds of truncated k-space sampling are dictated by the severity of luminal narrowing. The defined thresholds may help improve efficiency of 3D MR imaging of the carotid arteries while maintaining adequate luminal representation. PMID- 10871006 TI - Contrast-enhanced MR angiography of supraaortic vessels: the effect of voxel size on image quality. AB - BACKGROUND AND PURPOSE: Several acquisition strategies may be used for imaging supraaortic vessels by using contrast-enhanced MR angiography. The purpose of this study was to assess the effect of voxel size on image quality of MR angiograms. METHODS: Fourty-four patients underwent 3D MR angiography in the coronal plane. Patients were randomly assigned into two groups according to the voxel size of MR angiograms: group 1 referred to a 1.3 x 1.29 x 1.25-mm voxel size and group 2 to a 0.95 x 0.76 x 0.82 mm voxel size. Signal-to-noise ratios (SNRs) were measured and image artifacts were analyzed by consensus between observers. The delineation of the arterial lumen was independently ranked on a four-point scale (1 = not assessable; 2 = poor delineation; 3 = fair delineation; 4 = optimal delineation). RESULTS: The overall interobserver agreement for the delineation of the arterial lumen was good (K = .84, P < .0001), with a rank significantly higher in group 2 (68% of arteries graded as 4) compared with group 1 (76% graded as 3). SNRs were significantly higher by using the conventional resolution technique, with a negative correlation between SNRs and artery delineation (P < .0001). Image artifacts, however, were more frequent with the high-resolution technique, including wrap-around artifacts and signal fall-off at the origin of the great vessels. CONCLUSION: MR angiograms with a decreased voxel size improve the delineation of cervical carotid and vertebral arteries, despite reduced SNRs and additional artifacts. PMID- 10871008 TI - Scalp vein detected using internal carotid angiography that did not result in venous sinus compromise. AB - We present an unusual case of a scalp vein detected by using angiography of the internal carotid artery. The vein arose from the superior sagittal sinus and drained into the deep posterior cervical vein via the parietal emissary vein. This scalp vein may be a collateral pathway for venous sinuses; however, the patient had no evidence of venous sinus occlusive disease or intracranial hypertension. PMID- 10871009 TI - Prognostic value of MR and magnetization transfer imaging findings in patients with clinically isolated syndromes suggestive of multiple sclerosis at presentation. AB - BACKGROUND AND PURPOSE: The extent of abnormalities on T2-weighted MR images of the brain of patients with clinically isolated syndromes (CIS) suggestive of multiple sclerosis (MS) at presentation is associated with an increased risk of developing clinically definite MS (CDMS). We evaluated whether subtle changes outside T2-visible lesions are present in the brain of these patients and whether their extent increases the risk of subsequent development of CDMS. METHODS: Dual echo, T1-weighted, and magnetization transfer (MT) images of the brain were obtained from 24 patients with CIS at presentation. These patients were followed up for a mean period of 33 months (range, 25-42 months). Twenty age- and sex matched healthy volunteers served as control subjects. To create MT histograms of the normal-appearing brain tissue (NABT), macroscopic lesions were segmented from dual-echo images, were superimposed automatically, and were nulled out from the coregistered and scalp-stripped MT ratio (MTR) maps. The following MTR histogram derived measures were considered: average MTR, MTR(25), MTR(50), MTR(75), peak height, and peak position. T2 and T1 lesion loads, average lesion MTR, and brain volume were also measured. RESULTS: Patients with CIS had lower average NABT-MTR (P < .0001) and peak position (P = .002) than did control volunteers, but patient brain size was similar to that of volunteers. At follow-up, 10 (41%) patients developed CDMS. Patients who developed CDMS during the follow-up period had higher T2 lesion volume (P = .003) and lower average NABT-MTR (P = .005) and peak position (P = .006) than did those who did not develop CDMS. T2 lesion volume (odd ratio, 3.54; P = .0005) and average NABT-MTR (odd ratio, 0.81; P = .01) were independent predictors of the subsequent development of CDMS. CONCLUSION: Subtle changes occur outside lesions visible on conventional MR images among patients with CIS suggestive of MS at presentation. The greater the extent of such abnormalities is, the higher is the risk of subsequent development of CDMS. PMID- 10871010 TI - MR venography of multiple sclerosis. AB - BACKGROUND AND PURPOSE: The distribution of multiple sclerosis (MS) lesions in the brain follows a specific pattern, with most lesions in the periventricular regions and in the deep white matter; histopathologic studies have shown a perivenous distribution. The aim of this study was to illustrate these distribution patterns in vivo using high-resolution MR venography. METHODS: Seventeen MS patients underwent MR imaging at 1.5 T. Venographic studies were obtained with a 3D gradient-echo technique. MS lesions were identified on T2 weighted images, and their shape, orientation, and location were compared with the venous anatomy on the venograms. RESULTS: The use of contrast material facilitated the visualization of small veins and increased the number of veins seen. A total of 95 MS lesions could be identified on both the T2-weighted series and the venograms; a central vein was visible in all 43 periventricular lesions and in all but one of the 52 focal deep white matter lesions. The typical ovoid shape and orientation of the long axis of the MS lesions correlated well with the course of these veins. CONCLUSION: With MR venography, the perivenous distribution of MS lesions in the brain can be visualized in vivo. The venous anatomy defines the typical form and orientation of these lesions. PMID- 10871011 TI - Magnetization transfer histogram analysis of monosymptomatic episodes of neurologic dysfunction: preliminary findings. AB - BACKGROUND AND PURPOSE: Patients presenting with a monosymptomatic episode of neurologic dysfunction (MEND) have a high probability of developing multiple sclerosis (MS). Our study was designed to determine whether magnetization transfer (MT) histogram analysis could predict the development of MS for a cohort of patients presenting with a MEND. METHODS: Eleven patients with a MEND and 21 age-matched control volunteers underwent MR imaging. Six patients underwent serial MR examinations. MT ratio histogram peak height (MTRHPH) and the location of the MT ratio histogram peak (LOC MTRHP) were determined for patients and control volunteers. T2 lesion volume was also calculated. Patients were clinically followed up for 587 +/- 308 days to determine or rule out the development of MS. RESULTS: Three patients went on to develop MS. There was no statistically significant difference in the MTRHPH (P = .65) and the LOC MTRHP (P = .71) between patients and control volunteers. For those patients who underwent multiple examinations, no statistically significant differences in the MTRHPH (P = .64), LOC MTRHP (P =.58), and T2 lesion volume (P = .47) were seen. There were no statistically significant correlations between any of the parameters studied. CONCLUSION: We found no difference in MT histogram parameters among control volunteers, patients with a MEND without MS, and patients with a MEND who went on to a diagnosis of MS. Our preliminary findings suggest that there may not be a substrate of disease in the normal-appearing white matter that is predictive of the development of MS. PMID- 10871012 TI - Cerebral activation during multiplication: a functional MR imaging study of number processing. AB - BACKGROUND AND PURPOSE: Current models of brain function propose that number processing involves the interaction of different neuronal networks. Our purpose was to use functional MR (fMR) imaging to elucidate the brain regions engaged by multiplication. METHODS: Eighteen adults underwent fMR imaging while performing matching, multiplication, and control tasks. For each task, three or four single digit or low-value double-digit numbers were presented serially followed by a 12 second delay. A target stimulus then appeared and subjects made a judgement by pressing a button box that recorded responses. During the matching task, subjects judged whether the target stimulus matched one of the previous numbers. During the multiplication task, subjects judged whether the target stimulus was the product of the previous numbers. For the control task, the numbers were always zeros, and the subjects responded to a target stimulus that was always four zeros. Composite statistical parametric maps of the time course of activation comparing the control task with the matching and multiplication tasks, respectively, were generated and the significance of signal changes was estimated by randomization of statistical parametric maps. RESULTS: The matching and multiplication tasks resulted in activation (P < .005) in the medial superior frontal gyrus; the anterior cingulate gyrus; the intraparietal sulci, bilaterally; the right superior frontal sulcus bilaterally; the middle, inferior and precentral frontal gyri (left greater than right); the left basal ganglia; and the right lateral and inferior occipital gyri. There was a larger area of early activation in the right middle frontal gyrus during the matching task compared with the multiplication task, and there was a longer interval of activation in the left middle frontal gyrus during the multiplication task (10 seconds) than in the matching task (6 seconds). CONCLUSION: Multiplication and memory of numbers share an integrated network of brain regions. The left frontal lobe, an area also involved in memory and language processes, appears to play an important role in multiplication. PMID- 10871013 TI - The influence of gliomas and nonglial space-occupying lesions on blood-oxygen level-dependent contrast enhancement. AB - BACKGROUND AND PURPOSE: Functional MR (fMR) imaging with blood-oxygen-level dependent (BOLD) contrast enhancement is increasingly used as a noninvasive tool for presurgical mapping in patients with intracranial tumors. Most physiologic studies of task-related BOLD contrast enhancement have involved healthy volunteers. Therefore, it is not known whether BOLD contrast is evoked in the same way in or adjacent to tumor tissue. The purpose of this study was to study the influence of different intracranial tumors on BOLD contrast enhancement. METHODS: fMR mapping of the sensorimotor cortex was successfully performed in 15 of 21 patients with intracranial space-occupying lesions by using a bimanual motor task. Tumors were located either within the sensorimotor area itself or in adjacent brain areas, inducing changes of signal intensity on T2-weighted images along the pre- or postcentral gyrus. Space-occupying lesions were divided into a group comprising gliomas (seven cases) and a group comprising nonglial space occupying lesions (three metastases, two cavernomas, one abscess, one arteriovenous malformation, one meningioma). A hemispheric activation index was calculated using the volume of activation on the affected and on the contralateral hemisphere. Hemispheric activation indices of gliomas and nonglial lesions were compared statistically. RESULTS: The activated volume in the hemispheres ipsilateral to the nonglial lesions was 14% larger than in the contralateral hemisphere, whereas in the hemispheres ipsilateral to gliomas, the activated volume decreased by 36% in comparison with the contralateral hemisphere. The difference between nonglial lesions and gliomas was significant (P < .05). CONCLUSION: The generation of BOLD contrast enhancement is reduced near gliomas but is not affected by nonglial tumors. PMID- 10871014 TI - Patterns of disease spread in metastatic breast carcinoma: influence of estrogen and progesterone receptor status. AB - BACKGROUND AND PURPOSE: It is widely recognized that tumor hormone receptor status correlates with overall survival in metastatic breast carcinoma; however, the influence of hormone receptors on the pattern of disease spread is not well known. PURPOSE: We set out to determine the common distributions of metastatic disease spread in metastatic breast carcinoma, and to evaluate tumor hormone receptor status as predictor of disease spread. METHODS: Thirty-six patients being imaged for possible metastatic breast carcinoma between 1995 and 1998, in whom the presence or absence of tumor estrogen and progesterone receptors (ER+ or ER- / PR+ or PR-) was known, who underwent both contrast-enhanced MR of the brain and total body skeletal scintigraphy, were studied retrospectively. RESULTS: Of twelve patients with skeletal metastases but no brain metastases, 83% were ER+/PR+. Ten patients had brain metastases but no skeletal involvement, 80% of which were ER-/PR-. Seven patients had no brain or osseous metastases, but had metastatic disease in the chest or abdomen. Eighty-six percent of patients in this group were ER-/PR-. The tumor receptor status was statistically different between these three distribution groups (P = .01). A final group, consisting of seven patients, showed widespread disease, with diffuse metastases to the brain, viscera, and skeleton. In this group, no patients were ER+/PR+. CONCLUSION: There are two major patterns of disease spread in metastatic breast carcinoma, excluding patients with extensive diffuse metastases. Patients with ER+/PR+ tumors tend to develop osseous but not brain metastases. Patients with ER-/PR- tumors tend to develop brain but not osseous metastases. Appreciation of these distributions can aid the radiologist in detecting metastatic lesions, and will help the clinician to estimate the likelihood of metastases to various organ systems, as well as to potentially target therapy. PMID- 10871015 TI - Bilateral pituitary adenomas occurring with multiple endocrine neoplasia type one. AB - We report a case of synchronous bilateral pituitary adenomas in a patient with multiple endocrine neoplasia type one (MEN1). The patient was previously known to have a pancreatic gastrinoma and had first-degree relatives with MEN1. Both adenomas were concurrently revealed by high-resolution MR imaging of the pituitary gland as part of the investigation of the patient's severe, persistent headaches and elevated serum prolactin level. PMID- 10871016 TI - Increased conspicuity of intraventricular lesions revealed by three-dimensional constructive interference in steady state sequences. AB - We describe our preliminary experience with the three-dimensional constructive interference in steady state (3D-CISS) sequence for the evaluation of intraventricular lesions. Cyst walls, extent and margins of tumors, and intratumoral cystic structures were clearly depicted on 3D-CISS images. The 3D CISS sequence can offer additional information to conventional MR studies to define intraventricular lesions better. PMID- 10871017 TI - Hypertrophic olivary degeneration: metaanalysis of the temporal evolution of MR findings. AB - BACKGROUND AND PURPOSE: Hypertrophic olivary degeneration (HOD) is usually caused by a lesion in the triangle of Guillain and Mollaret and presents clinically as palatal tremor. Although the imaging features have been well described, the temporal course of hypertrophy and T2 signal increase in the inferior olivary nucleus (ION) has not been fully characterized. Our purpose was to evaluate the time course of MR imaging features of HOD caused by a lesion within the triangle of Guillain and Mollaret. METHODS: The temporal progression of HOD in 45 patients with symptomatic palatal tremor was obtained by extrapolation of combined MR imaging data from six patients treated at our institution and 39 patients reported in the literature. The MR examinations and reports were reviewed for presence of hyperintense signal in the ION on T2-weighted images, hypertrophy of the ION, and an inciting lesion in the triangle of Guillain and Mollaret. The interval between the MR examination and the inciting lesion was determined. RESULTS: Increased olivary signal on T2-weighted images first appeared 1 month after the inciting lesion and persisted for at least 3 to 4 years. Olivary hypertrophy initially developed 6 months after the acute event and resolved by 3 to 4 years. CONCLUSION: Visible changes on MR images in the ION in patients with a lesion in the triangle of Guillain and Mollaret correlate well with the described sequential histopathologic findings. PMID- 10871018 TI - Diffusion changes in the aging human brain. AB - BACKGROUND AND PURPOSE: Quantifying changes in the human brain that occur as part of normal aging may help in the diagnosis of diseases that affect the elderly and that cause structural changes in the brain. We sought to assess diffusion changes that are inherently related to brain structure during aging. METHODS: MR scans were obtained from 11 healthy volunteers and 27 patients (ages 26 to 86 years [53.4 +/- 17.0 years]). Images acquired from the patients either showed no abnormalities, contained minimal periventricular white matter changes, or revealed focal lesions. Maps of the average diffusion constant (D(av)) were calculated for each subject. Changes in D(av) were determined with distribution analysis (histogram) for the entire brain and compared with region-of-interest measurements from the periventricular white matter and thalamus. RESULTS: Mean D(av) of the entire brain (0.74 +/- 0.02 x 10(-5) cm2/s) showed weaker age dependency compared with the periventricular white matter D(av)(0.76 +/- 0.04 x 10(-5) cm2/s). The D(av) of the thalamus D(av) (0.75 +/- 0.03 x 10(-5) cm2/s) had no age dependency. The age-dependent changes of entire brain D(av) may be significant for subjects older than 60 years compared with younger subjects. CONCLUSION: In this study, we observed minimal changes in the D(av) of the entire brain with aging. The mean D(av) of the human brain is nearly constant throughout most of adulthood. PMID- 10871019 TI - Effects of contrast material on single-volume proton MR spectroscopy. AB - BACKGROUND AND PURPOSE: Administration of contrast material before proton MR spectroscopy may allow more accurate placement of the volume of interest, particularly in tumors; yet, some data have suggested that contrast material may alter the results of MR spectroscopy. To determine the validity of this contention, we performed pre- and postcontrast MR spectroscopy in patients with brain tumors and compared the results with those obtained from a phantom. METHODS: Ten patients with astrocytomas were examined with single-volume MR spectroscopy before and after administration of contrast material. Voxel placement was identical for all studies. Peak area, peak height, and width at half maximum were measured for N-acetyl aspartate (NAA), creatine (Cr), and choline (Cho) in all studies. A phantom containing a 10 mmol concentration of NAA, Cr, and Cho was prepared in phosphate-buffered saline and mixed with contrast concentrations varying from 0.1 to 1.0 mmol. The phantom was studied by MR spectroscopy with the same parameters as used for the clinical studies. RESULTS: No significant differences were found between the pre- and postcontrast MR spectroscopy studies for the three parameters measured. In phantom studies, there was a significant decline in the Cho peak area and height and an increase in the width at half maximum as the concentration of contrast material increased from 0.1 to 1.0 mmol. NAA and Cr peaks showed no significant changes in peak height or area. CONCLUSION: Contrast material may be administered before clinical MR spectroscopy without affecting its interpretation. PMID- 10871020 TI - MR findings of eosinophilic meningoencephalitis attributed to Angiostrongylus cantonensis. AB - Eosinophilic meningoencephalitis is prevalent and widely distributed in Thailand, especially in the northeastern and central parts of the country. Angiostrongylus cantonensis is one of the causative agents of fatal eosinophilic meningoencephalitis. The nematodes produce extensive tissue damage by moving through the brain and inducing an inflammatory reaction. We report the clinical features and the findings revealed by MR imaging and MR spectroscopy in six patients with eosinophilic meningoencephalitis. The clinical presentation included severe headache, clouded consciousness, and meningeal irritation. Abnormal findings on MR images included prominence of the Virchow-Robin spaces, subcortical enhancing lesions, and abnormal high T2 signal lesions in the periventricular regions. Proton brain MR spectroscopy was performed in three patients and was abnormal in one severe case, showing decreased choline in a lesion. Small hemorrhagic tracts were found in one case. Lesions thought to be due to microcavities and migratory tracts were found in only one case. We believe the MR imaging and MR spectroscopy findings are of diagnostic value and helpful in understanding the pathogenetic mechanisms of the disease. PMID- 10871021 TI - Applicability and advantages of flow artifact-insensitive fluid-attenuated inversion-recovery MR sequences for imaging the posterior fossa. AB - We describe a new sequence, flow artifact-insensitive fluid-attenuated inversion recovery (FAIS-FLAIR), that capitalizes on the advantages of fluid-attenuated inversion recovery (FLAIR) while minimizing FLAIR-related artifacts such as those often encountered in the posterior fossa. Twenty-eight patients with posterior fossa disease underwent FAIS-FLAIR, conventional FLAIR, and spin-echo MR studies, and the findings yielded by the three techniques were compared. In this patient population, postcontrast FAIS-FLAIR imaging was obtained in 20 patients and compared with postcontrast T1-weighted images. The images were assessed for lesion conspicuity by three radiologists. FAIS-FLAIR markedly reduces the inflow artifacts from noninverted CSF on FLAIR images. It does so with and without contrast agent administration, and produces higher lesion conspicuity compared with T1- and T2-weighted spin-echo sequences and conventional FLAIR images of the posterior fossa. PMID- 10871022 TI - Concepts of myelin and myelination in neuroradiology. AB - Until the advent of MR imaging, knowledge of the structure of myelin and the process of myelination were of little importance to the neuroradiologist. Other than some mild changes in the attenuation of white matter, myelination resulted in no significant alterations of CT (1) or sonographic studies. MR studies, on the other hand, have been increasingly used for pediatric brain imaging. MR imaging's greater sensitivity to small changes in the water content of brain tissue, to changes in the binding of free water (revealed by magnetization transfer), and to the extent and anisotropy of water diffusion (revealed by diffusion imaging) has cast new light on this very complex and important molecule. Assessing myelination has become a key component of evaluating the child with delayed development. Moreover, better understanding of the nature of myelin and the effect of its different components on MR imaging parameters may help us to understand and diagnose inborn errors of metabolism better. In this review, I discuss what is known regarding the function and structure of CNS myelin and the effects of the various components of myelin on the signal imparted to the MR image. Summary Abnormalities of myelin can cause a wide variety of disorders of the nervous system. MR imaging is a powerful tool for the study of myelin and its disorders. However, only by understanding the physiologic properties and structure of myelin can we use MR imaging to its fullest capacity for studying patients with myelin disorders. In this review, I have discussed the structure of myelin as it relates to MR imaging of normal myelination and to neurologic disorders resulting from abnormalities of myelin. Thinking of myelin and its disorders in this manner will be critical to using MR imaging techniques optimally to diagnose and study these disorders further. PMID- 10871023 TI - Cranial MR imaging of osteopetrosis. AB - BACKGROUND AND PURPOSE: The purpose of this study was to describe the cranial MR imaging manifestations of osteopetrosis. These features have not previously been reported in the literature. METHODS: Cranial MR studies, obtained with a uniform imaging protocol, were reviewed in 47 patients with osteopetrosis. Thirty-four patients had autosomal recessive (malignant) osteopetrosis (AROP), seven had intermediate osteopetrosis (IOP), and six had either type I or type II autosomal dominant osteopetrosis (ADOP I or II). The prevalence of abnormalities was tabulated and compared with the specific osteopetrosis variants. RESULTS: All patients with osteopetrosis had thickening and sclerosis of the calvaria. Ventriculomegaly, tonsillar herniation, proptosis, and dural venous sinus stenosis were observed in the majority of patients with AROP and ADOP I. Optic nerve sheath dilatation occurred in many of the patients with AROP and in all patients with ADOP I. Acquired cephaloceles were also observed only in these two groups. Optic nerve atrophy and optic canal stenosis were observed in a majority of patients with AROP, IOP, and ADOP II. Middle ear fluid was prevalent in AROP and IOP, present in over half the patients in each group. Features seen most prevalently, or exclusively, in AROP included stenosis of the internal carotid and vertebral arteries and extramedullary hematopoiesis. CONCLUSION: The cranial MR imaging features of osteopetrosis are both shared and unique among the various subtypes of the disease. The specific cranial and intracranial manifestations reflect the predominant calvarial or skull base patterns of bone thickening. The unique features seen in patients with AROP probably reflect the early age of onset and the greater severity of this form of the disease. PMID- 10871024 TI - Posterior pituitary ectopia: another hint toward a genetic etiology. AB - We report the MR and clinical findings of two patients with growth hormone deficiency and posterior pituitary ectopia (PPE). Possible causes of PPE are discussed. PMID- 10871025 TI - A heterotopic cerebellum presenting as a suprasellar mass with associated nasopharyngeal teratoma. AB - We present a case of nasopharyngeal teratoma that was discovered in association with a suprasellar heterotopic cerebellum in a newborn. Well-differentiated, heterotopic, cerebellar masses have been reported in the orbits, spine, and frontal encephalocele but not, to our knowledge, in the suprasellar region. In this report, we describe the imaging findings and discuss the possible origins of the two masses discovered in this case. PMID- 10871026 TI - Diffuse leptomeningeal oligodendrogliomatosis: radiologic/pathologic correlation. AB - We present the radiologic and pathologic findings in a boy who presented with diffuse leptomeningeal enhancement and whose clinical status deteriorated over the course of 5 years. During this period, MR images showed progression of the enhancement in the subarachnoid spaces, formation of intraaxial cysts, and hydrocephalus. Autopsy findings revealed diffuse oligodendroglioma throughout the leptomeninges of the brain and spine, with no definite intraaxial focus. The radiologic and pathologic features of diffuse leptomeningeal oligodendrogliomatosis are reviewed. PMID- 10871027 TI - Sonography for the detection of cervical lymph node metastases among patients with tongue cancer: criteria for early detection and assessment of follow-up examination intervals. AB - BACKGROUND AND PURPOSE: Because the presence of cervical metastasis is one of the factors influencing the outcome of patients with carcinoma of the head and neck, its early detection is potentially very important. The purpose of this study was to evaluate the characteristic changes of cervical metastasis revealed by sonography during follow-up and to assess an adequate interval for follow-up sonography of the neck among patients with tongue cancer. METHODS: Forty-three of 44 consecutive patients with squamous cell carcinoma of the tongue, who had undergone interstitial brachytherapy, were examined with sonography of the neck during the posttherapeutic follow-up period. RESULTS: Seventeen cervical lymph node metastases were found in 12 of 43 patients during follow-up. Of these 17 cervical metastases, 16 (94.1%) were accurately diagnosed and one (5.9%) was misdiagnosed as nonmetastatic based on sonographic findings. Sonography of the neck performed in seven patients at an interval of less than 1 month since the last follow-up imaging showed 9 (90.0%) of 10 metastases increased by up to 2 mm in short-axis diameter. Five patients who were followed up at an interval of more than 1 month since the last follow-up imaging had seven metastases increase by 3 to 8 mm in short-axis diameter or a change of echogenicity in the internal structure of lymph nodes or both. Pathologic examinations showed extranodal spread in 3 (42.9%) of these 7 metastases. CONCLUSION: Changes both in size and internal echogenicity can occur as quickly as 2 to 4 weeks between sonographic examinations. Therefore, in high-risk patients, or in those with suspicious sonographic findings, short-interval follow-up sonographic examinations are recommended at least during the first posttherapeutic year. Our findings suggest that follow-up sonography of the neck should be performed monthly, at least during the first posttherapeutic year. PMID- 10871028 TI - The role of neural networks in improving the accuracy of MR spectroscopy for the diagnosis of head and neck squamous cell carcinoma. AB - BACKGROUND AND PURPOSE: MR Spectroscopy (MRS) has the unique ability to analyze tissue at the molecular level noninvasively. The purpose of this study was to determine if peak heights revealed by proton MRS ((1)H-MRS) signals showed that neural networks (NN) provided better accuracy than linear discriminant analysis (LDA) in differentiating head and neck squamous cell carcinoma (SCCA) from muscle METHODS: In vitro 11-T (1)H-MR spectra were obtained on SCCA tissue samples (n = 16) and muscle (n = 12). The peak heights at seven metabolite resonances were measured: olefinic acids at 5.3 ppm, inositol at 3.5 ppm, taurine at 3.4 ppm, choline (Cho) at 3.2 ppm, creatine (Cr) at 3.0 ppm, sialic acid at 2.2 ppm, and methyl at 0.9 ppm. Using leave-one-out experimental design and receiver operating characteristic curve analysis, the ability of NN and LDA classifiers to distinguish SCCA from muscle were compared (given equal weighting of false negative and false-positive errors). These classifiers were also compared with an existing method that forms a diagnosis by using LDA of the Cho/Cr peak area ratio. RESULTS: NN classifiers, which were identified using height data, achieved better sensitivity and specificity rates in distinguishing SCAA from muscle than did LDA using height or area data. Sensitivity/specificity for the NN analysis of the seven metabolite peak heights were 87.5 % and 83.3%, respectively, for a one hidden-node network and 81.2% and 91.7%, respectively, for a two-hidden-node network. Additional nodes did not improve accuracy. The sensitivity and specificity were 81.2% and 50%, respectively, for LDA of the seven peak heights, and 68% and 83%, respectively, for LDA of the Cho/Cr peak area ratio. CONCLUSION: NN classifiers with peak height data were superior to LDA of the peak heights and LDA of the Cho/Cr peak area ratio for differentiating SCCA from normal muscle. These results show neural network analysis can improve the diagnostic accuracy of (1)H-MRS in differentiating muscle from malignant tissue. Further studies are necessary to confirm our initial findings. PMID- 10871029 TI - Imaging findings in schwannomas of the jugular foramen. AB - BACKGROUND AND PURPOSE: Tumors of the cranial nerve sheath constitute 5% to 10% of all intracranial neoplasms, yet few articles have described their CT and MR characteristics. We report the imaging findings in a relatively large series of schwannomas of the jugular foramen, contrasting them with other disease entities, especially vestibular schwannomas and tumors of the glomus jugulare. METHODS: CT and/or MR studies of eight patients who underwent surgery for histologically proved schwannomas were reviewed retrospectively. One additional patient with an assumed schwannoma of the jugular foramen, who did not have surgery, was also included. RESULTS: Surgical findings showed schwannomas of the glossopharyngeal nerve in seven patients and tumor involvement of both the glossopharyngeal and vagal nerves in one patient. All tumors were partially located within the jugular foramen. Growth extending within the temporal bone was typical. Tumor extended into the posterior cranial fossa in all nine patients and produced mass effect on the brain stem and/or cerebellum in seven patients; in five patients, tumor extended below the skull base. On unenhanced CT scans, tumors were isodense with brain in six patients and hypodense in two. In seven patients, CT scans with bone algorithm showed an enlarged jugular foramen with sharply rounded bone borders and a sclerotic rim. On MR images, T1 signal from tumor was low and T2 signal was high relative to white matter in all patients. Contrast enhancement on CT and/or MR studies was strong in eight patients and moderate in one. CONCLUSION: Schwannoma of the jugular foramen is characteristically a sharply demarcated, contrast-enhancing tumor, typically centered on or based in an enlarged jugular foramen with sharply rounded bone borders and a sclerotic rim. Intraosseous extension may be marked. PMID- 10871030 TI - MR dacryocystography: comparison with dacryocystography and CT dacryocystography. AB - BACKGROUND AND PURPOSE: Several techniques have been used to image the nasolacrimal system, providing functional (dacryoscintigraphy) or morphologic (dacryocystography, CT dacryocystography [CTD]) information. Using gadopentetate dimeglumine-diluted solution injected into the lacrimal canaliculus or instilled into the conjunctival sac, we compared the sensitivity of MR dacryocystography (MRD) with that of CTD. METHODS: Eleven healthy volunteers and 25 patients affected by primary epiphora (21 patients) or postsurgical recurrent epiphora (four patients) underwent MRD after the topical administration of contrast media or cannulation of the lacrimal canaliculus. The MR imaging findings were compared with irrigation and CTD data. All patients underwent surgical treatment (dacryocystorhinostomy), which served as a standard of reference for confirming the MRD findings. RESULTS: The topical administration of contrast-enhanced saline solution and the injection of contrast-enhanced saline solution after cannulation were always well tolerated. In healthy volunteers, outflow of contrast media was always revealed by MRD. Eight (32%) of 25 patients with epiphora had stenosis proximal to the lacrimal sac revealed by MRD, whereas 17 (68%) of 25 showed a dilated lacrimal sac and nasolacrimal duct stenosis, as confirmed by surgical findings. The findings of MRD after the topical administration of contrast medium and MRD after cannulation of the lacrimal canaliculus were comparable with irrigation or CTD data for all patients except one. CONCLUSION: In patients with epiphora, MR imaging performed after the topical administration of diluted contrast material can reveal stenosis of the lacrimal apparatus and can be added to the standard orbital imaging protocol when lacrimal system involvement is suspected. PMID- 10871031 TI - Evolution of a new multidisciplinary subspecialty: interventional neuroradiology/neuroendovascular surgery. PMID- 10871032 TI - Program requirements for residency/fellowship education in neuroendovascular surgery/interventional neuroradiology: a special report on graduate medical education. AB - BACKGROUND AND PURPOSE: Neuroendovascular surgery/interventional neuroradiology is a relatively new subspecialty that has been evolving since the mid-1970s. During the past 2 decades, significant advances have been made in this field of minimally invasive therapy for the treatment of intracranial cerebral aneurysms; acute stroke therapy intervention; cerebral arteriovenous malformations; carotid cavernous sinus fistulas; head, neck, and spinal cord vascular lesions; and other complex cerebrovascular diseases. Advanced postresidency fellowship programs have now been established in North America, Europe, and Japan, specifically for training in this new subspecialty. METHODS: From 1986 to the present, an ad hoc committee of senior executive committee members from the American Society of Interventional and Therapeutic Neuroradiology, the Joint Section of Cerebrovascular Neurosurgery, and the American Society of Neuroradiology met to establish, by consensus, general guidelines for training physicians in this field. RESULTS: In April 1999, the Executive Committee of the Joint Section of Cerebrovascular Neurosurgery voted unanimously to endorse these training standard guidelines. In May 1999, the Executive Committee of the American Society of Interventional and Therapeutic Neuroradiology and the American Society of Neuroradiology also unanimously voted to endorse these guidelines. In June 1999, the Executive Council of the American Association of Neurological Surgeons and the Congress of Neurological Surgeons unanimously voted to endorse these guidelines. CONCLUSION: The following guidelines for residency/fellowship education have now been endorsed by the parent organization of both the interventional and diagnostic neuroradiology community, as well as both senior organizations representing neurosurgery in North America. These guidelines for training should be used as a reference and guide to any institution establishing a training program in neuroendovascular surgery/interventional neuroradiology. PMID- 10871033 TI - The utility of the microcrystalline cellulose sphere as a particulate embolic agent: an experimental study. AB - BACKGROUND AND PURPOSE: Although various particulate materials have been developed as embolization agents, their biocompatibility remains unclear. We used an animal model to examine the possibility of using FDA-approved microcrystalline cellulose spheres (CELPHERE) as solid embolic material for the permanent occlusion of blood vessels. METHODS: Angiographic and histologic studies in 12 canine renal arterial systems were conducted to evaluate the performance of CELPHERE beads at 1 hour, and at 4 and 12 weeks after embolization. RESULTS: The CELPHERE beads traveled to vessels with diameters approximating their own. Larger vessels were occluded by aggregations of beads. There was no disruption of vessel walls and no evidence of perivascular hemorrhage or inflammatory changes. CONCLUSION: Because CELPHERE beads are easy to handle, highly biocompatible, and have few adverse effects, they are suitable for intravascular applications. PMID- 10871034 TI - The utility of diffusion-weighted imaging with navigator-echo technique for the diagnosis of spinal epidermoid cysts. AB - We report a case of a spinal epidermoid cyst in which diffusion-weighted imaging with a navigator-echo technique was useful for the differential diagnosis from other cystic tumors. Motion artifacts are inherent on diffusion-weighted images of the spinal region; however, the navigator-echo technique compensated for this problem and provided high-quality images. Diffusion-weighted imaging with navigator echoes is considered to be a potentially useful tool in the differential diagnosis of spinal cystic tumors. PMID- 10871035 TI - Partial aplasia of the posterior arch of the atlas with an isolated posterior arch remnant: findings in three cases. AB - We report the imaging findings in three symptomatic cases of partial aplasia of the posterior arch of the atlas with an isolated posterior remnant of the arch. These cases are instructive in illustrating the mechanism of cord impingement produced by the posterior arch remnant during extension of the cervical spine. Additionally, focal increased T2 signal was observed within the cord at the level of the anomaly in two of the patients. PMID- 10871036 TI - Diffusion-weighted imaging of cerebral abscess and subdural empyema. PMID- 10871037 TI - In Euglena gracilis, a heat-shock protein related to hsc73 is constitutive and stress inducible. AB - Using monoclonal antibodies directed against different cytoplasmic isoforms of hsp70 proteins, namely, the constitutive hsc73 and the inducible hsp72 isoforms, we found that one isoform related to hsc73 was present in Euglena gracilis. This hsc73-like protein is expressed with a higher rate of synthesis in cells growing under heat shock than in control cells. Moreover, in cadmium-resistant cells, cultured at normal growth temperature, the rate of synthesis of this protein is constitutively increased. These results indicate that a heat-shock protein related to hsc73 is present in an ancestral eukaryote, Euglena gracilis, and that this protein may be constitutive and stress inducible as well. PMID- 10871038 TI - Primary structure and functional characterization of bilitoxin-1, a novel dimeric P-II snake venom metalloproteinase from Agkistrodon bilineatus venom. AB - The amino acid sequence of the hemorrhagic toxin, bilitoxin-1, isolated from the venom of Agkistrodon bilineatus was determined by the Edman sequencing procedure of peptides derived from digests utilizing cyanogen bromide, clostripain, lysyl endopeptidase, and Staphylococcus aureus V8 protease. A molecular mass of 80,000 Da was observed in the nonreduced state and 48,000 Da was observed in the reduced state, as demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Each subunit consists of 291 amino acid residues and has a calculated molecular mass of 32,276 Da. The toxin contains fucose, galactosamine, glucosamine, galactose, mannose, and N-acetylneuraminic acid and three N-linked glycosylation consensus sites. Hydrazinolysis and ESI mass spectrometry revealed that asparagine was the carboxyl-terminal amino acid. The disintegrin-like domain of bilitoxin-1 lacks the RGD cell-binding sequence, which is substituted by the MGD sequence. Under certain conditions, the disintegrin domain is autoproteolytically processed from the native protein. Studies with the bilitoxin disintegrin demonstrated that it lacks platelet aggregation inhibitory activity, probably reflecting the substitution of RGD by MGD. The hemorrhagic activity of the asialobilitoxin-1 was only 25% of bilitoxin-1, while proteolytic activity was unaffected. The three-dimensional structure of this toxin was modeled and was shown to likely possess a structure similar to that of adamalysin II (Gomis-Ruth et al., EMBO J. 12, 151-157 (1993)) and the disintegrin kistrin (Adler et al., Biochemistry 32, 282-289 (1993)). In summary, here we report the first primary structure of a dimeric, P-II snake venom metalloproteinase and the biological role of bilitoxin-1 glycosylation and the disintegrin domain. PMID- 10871039 TI - Tropomodulin-binding site mapped to residues 7-14 at the N-terminal heptad repeats of tropomyosin isoform 5. AB - Tropomodulin is a globular protein that caps the pointed end of actin filaments by complexing with the N-terminus of a tropomyosin (TM) molecule. TM consists of coiled coils except for the N-terminus, which may be globular. Here we report that human TM isoform 5 (hTM5) lacking the N-terminal 18 residues lost its binding activity toward tropomodulin. We further characterized the tropomodulin binding site by creating a series of deletion and missense mutations within this region, followed by a solid-phase binding assay. I7, V10, and I14, hydrophobic residues located at the a and d positions of N-terminal heptad repeats involving intertwine, are essential for tropomodulin binding. R12, a positively charged residue at the f position, is also involved in recognition. In contrast, A2R and G3Y mutations, each creating a bulky N-terminus, did not alter the binding. In addition, rat TM5b, which differs from hTM5 in residues 4-6, exhibits a similar binding affinity. The tropomodulin-binding site, therefore, is mapped to residues 7-14 at the beginning of the long heptad repeats. Column chromatography revealed that hTM5 mutants remained capable of dimerization. Results also suggest tropomodulin has a groove-type, rather than a cavity-type, binding site for hTM5. We also mapped the epitope of monoclonal antibody LC1 to residues 4-10 of hTM5 and showed the competition between mAb LC1 and tropomodulin in hTM5 binding. Since the N-terminal residues need to overlap with the C-terminus of TM in their head-to-tail association, this investigation elucidates the mechanisms by which the tropomodulin-hTM5 complex is formed and functions in regulating the actin filaments. PMID- 10871040 TI - Effect of D-amino acid substitution in a mucin 2 epitope on mucin-specific monoclonal antibody recognition. AB - We have studied the influence of D-amino acid substitution in the flanking region on the antibody recognition of the 19TGTQ22 epitope core in the tandem repeat of mucin 2 (MUC2) glycoprotein. Analogue peptides corresponding to the optimal epitope sequence (16PTPTGTQ22) have been prepared by the replacement of single or multiple L-amino acid residues at the N-terminal part of the molecule. According to previous studies, this portion of the all-L 16PTPTGTQ22 peptide possesses a beta-turn secondary structure important for efficient monoclonal antibody interaction. The binding properties of sequentially modified peptides (pTPTGTQ, ptPTGTQ, ptpTGTQ, and ptptGTQ) have been analyzed by a MUC2 glycoprotein specific monoclonal antibody (MAb 996) using RIA inhibition assay and characterized by IC50 values. At the same time, we have investigated the secondary structure of the compounds by circular dichroism and Fourier transform infrared spectroscopy in solution. Our data showed that the presence of D-amino acid residue(s) at position(s) 16P, 16PT17, or 16PTP18 resulted in gradually decreasing antibody binding, but the replacement of the L-Thr at position 19 almost abolished activity. Parallel with this reduction, changes in the conformer population have been detected. The propensity of the pTPTGTQ peptide to adopt folded, most probably beta-turn, structure in water can be in correlation with its essentially preserved antibody recognition. After further substitution, the peptide still contained beta- and/or gamma-turn folded secondary structural elements. The conformation of peptide ptptGTQ could be characterized mostly by semiextended (polyproline II) and probably classic gamma-turn conformers built up from D residues. PMID- 10871041 TI - Two-dimensional peptide mapping of cross-linked type IX collagen in human cartilage. AB - Type IX collagen is a quantitatively minor component of hyaline cartilage that is essential for the normal structural integrity of the tissue. Purification and analysis are difficult because the mature protein is insoluble as a cross-linked integral component of the fibrillar matrix. In order to view a peptide map of the total pool of type IX collagen in a cartilage sample, a selective method based on Western blot analysis was developed for displaying collagen IX peptides in a cyanogen bromide digest of tissue. Digests were partially resolved by reverse phase HPLC, individual fractions were run on SDS-PAGE and then transblotted to membrane, and the collagen IX fragments were revealed using an anti-collagen IX rabbit antiserum. All major CB-peptides from alpha1(IX), alpha2(IX), and alpha3(IX) chains in the resulting two-dimensional display were identified by amino-terminal sequence analysis. Cross-linked peptides originating from sites of covalent interaction between collagen types IX and II and between IX and IX were also defined. By comparison with an analysis of soluble type IX collagen from chondrocyte culture medium, the results showed that the pool of type IX collagen molecules in fetal and adult human cartilage is extensively cross-linked intermolecularly at sites previously revealed by other methods using purified protein. This sensitive, direct method has the potential to screen for abnormalities in the content and properties of type IX collagen in tissue samples, for example, in the study of heritable chondrodysplasia syndromes and the pathogenesis of cartilage destruction in osteoarthritis. PMID- 10871042 TI - Complex formation between Chromatium vinosum ferric cytochrome c' and bromophenol blue. AB - An unusual complex has been observed between the common electrophoresis tracer bromophenol blue (BPB) and the cytochrome c' from Chromatium vinosum during polyacrylamide gel electrophoresis. Complex formation results in a shift and increase in the intensity of the visible absorption band of BPB. Differential spectrophotometric titration of BPB with cytochrome c' indicates that one BPB binds to each of the two subunits of cytochrome c' with a binding constant of 4.2(0.5) x 10(5). The absence of a significant effect of ionic strength on the binding constant and the effect of Triton X-100 on the spectrum of BPB suggest that hydrophobic interactions are important to binding. An analysis of the structure of C. vinosum cytochrome c' shows the presence of a surface hydrophobic patch which may participate in the binding interaction. Many of the hydrophobic amino acids in the patch are well conserved by type among all known sequences of cytochrome c' and are found in loop elements of the 3D structure, suggesting a functional basis for conservation. It is proposed that the binding of BPB may mimic a relevant interaction involving the cytochrome c' biological function. PMID- 10871043 TI - Regulation of the binding of myristoylated alanine-rich C kinase substrate (MARCKS) related protein to lipid bilayer membranes by calmodulin. AB - The effector domain (ED) of MARCKS proteins can associate with calmodulin (CaM) as well as with phospholipids. It is not clear, however, whether a complex between MARCKS proteins and CaM can form at the surface of phospholipid membranes or whether CaM and membranes compete for ED binding. Using two-mode waveguide spectroscopy, we have investigated how CaM regulates the association of MARCKS related protein (MRP) with planar supported phospholipid bilayer membranes. Bringing a solution containing CaM into contact with membranes on which MRP had previously been deposited results in low-affinity binding of CaM to MRP. A preformed, high-affinity CaM MRP complex in the aqueous phase binds much more slowly than pure MRP to membranes. Similar observations were made when a peptide corresponding to the ED of MRP was used instead of MRP. Hence CaM cannot form a stable complex with MRP once the latter is bound at the membrane surface. CaM can, however, strongly retard the association of MRP with lipid membranes. The most likely interpretation of these results is that CaM and the phospholipid membrane share the same binding region at the ED and that the ED is forced by membrane binding to adopt a conformation unfavorable for CaM binding. PMID- 10871044 TI - Cloning, chromosomal sublocalization of the human soluble aminopeptidase P gene (XPNPEP1) to 10q25.3 and conservation of the putative proton shuttle and metal ligand binding sites with XPNPEP2. AB - Human soluble ("cytosolic") aminopeptidase P (hsAmP) is an aminoacylprolyl hydrolase (EC 3.4.11.9) present in all tissues yet examined. hsAmP is related in terms of catalytic specificity to an ectoenzyme, membrane aminopeptidase P (hmAmP), which is largely limited in distribution to endothelia and brush border epithelia. Although both enzymes can degrade oligopeptides having N-terminal Xaa Pro- moieties, hsAmP and hmAmP are of relatively low sequence homology. Recently, it has been shown that the two enzymes are not products of splice variants of the same gene. How hsAmP relates to hmAmP has clinical significance in that both can inactivate bradykinin, and AmP deficiency states have been described. The hmAmP gene (XPNPEP2) is disposed at chromosome Xq25, a disposition with clear meaning in terms of inheritance of hmAmP deficiencies. To further explore similarities and differences between hsAmP and hmAmP, the present study was begun to determine the chromosomal disposition of the hsAmP gene. Here we show that the gene is sublocalized on chromosome 10q25.3. We also show that hsAmP and hmAmP contain homologous blocks of sequence common to members of the "pita bread-fold" protein family, of which Escherichia coli methionine aminopeptidase is the prototype. The prototype is known to contain a proton shuttle and five divalent metal ligands, counterparts of which we identify in the homologous blocks of sequence in both hsAmP and hmAmP and compare to E. coli aminopeptidase. PMID- 10871046 TI - Oxidation of linoleyl alcohol by potato tuber lipoxygenase: kinetics and positional, stereo, and geometrical (cis, trans) specificity of the reaction. AB - The dioxygenation of linoleyl alcohol (LAL) by potato tuber lipoxygenase leads to formation of two positional isomeric products--9- and 13-hydroperoxyoctadecadien 1-ols (Butovich, I. A., Luk'yanova, S. M., and Reddy, C. C. (1998) Biochem. Biophys. Res. Commun. 249, 344-349). In the present study, we examined the stereospecificity and double-bond conformation of primary dioxygenation products of LAL catalyzed by potato lipoxygenase. In contrast to the product profiles of linoleic acid oxidation by potato lipoxygenase, oxidation of LAL led to all possible positional (9- and 13-), stereo, and geometrical (cis,trans and all trans) isomers in equimolar mixtures at 25 degrees C. The reaction appears to proceed through an enzyme-catalyzed formation of a pentadiene carbon-centered radical followed by resonance stabilization of the radical and molecular oxygen insertion in an enzyme-dependent as well as an enzyme-independent pathway. A strict positional, stereo, and geometrical specificity of the dioxygenation products of LAL oxidation appears to be maintained when the reaction occurs at the active site of the enzyme. However, when the pentadiene carbon-centered radical of LAL is dissociated from the active site of the enzyme, it appears to be nonenzymatically transformed into a mixture of all possible positional and geometrical stereoisomers of primary dioxygenation products. The latter pathway was effectively blocked by the free radical scavenger 4-hydroxy-TEMPO, which substantially reduced the production of all-trans hydroperoxyoctadecadienols. In the presence of the scavenger, 9(S)-hydroperoxy-10E,12Z-octadecadien-1-ol was the predominant LAL oxidation product, representing approximately 80% of the total conjugated dienes, with 13(S)-hydroxy-9Z,11E-octadecadien-1-ol the expected product of reverse orientation of the substrate at the active site, accounting for approximately 10%. A similar pattern in oxidation of LAL was observed when the reactions were carried out at 0 degrees C. PMID- 10871045 TI - CCAAT/enhancer binding protein beta mediates the activation of the murine alpha1(I) collagen promoter by acetaldehyde. AB - Acetaldehyde was previously shown to activate the alpha1(I) and alpha2(I) collagen promoters and to increase collagen production in activated stellate cells. Also, CCAAT/enhancer binding protein beta (C/EBPbeta) binds and activates the mouse alpha1(I) collagen promoter. This study investigates the role of C/EBPbeta in mediating the activation of the alpha1(I) collagen promoter by acetaldehyde. Nuclear extracts isolated from cultured activated rat hepatic stellate cells formed four protein-DNA complexes on electrophoretic mobility shift assay with an oligonucleotide including the C/EBP binding site between -365 and -335 in the alpha1(I) collagen promoter. The four complexes were identified to represent C/EBPbeta binding to the oligonucleotide by supershift with C/EBPbeta antibody. The principal C/EBP isoform found in the nuclear extracts from stellate cells was C/EBPbeta, with very low amounts of C/EBPalpha detected. Acetaldehyde (200 microM) increased C/EBPbeta protein in stellate nuclear extracts, increased its binding to the promoter, and activated the alpha1(I) collagen promoter in transfected stellate cells. Mutation of the C/EBPbeta binding site markedly decreased nuclear protein binding. A transfected promoter, mutated at the C/EBP binding site, had decreased basal activity, was not activated by acetaldehyde, and was not activated when cotransfected with a C/EBPbeta expression vector. This study shows that C/EBPbeta is the predominant C/EBP isoform found in activated stellate cells and that increased C/EBPbeta protein and C/EBPbeta binding to a proximal C/EBP binding site in the promoter mediates the activating effect of acetaldehyde. PMID- 10871047 TI - Effects of lipid-derivatized glycosaminoglycans (GAGs), a novel probe for functional analyses of GAGs, on cell-to-substratum adhesion and neurite elongation in primary cultures of fetal rat hippocampal neurons. AB - The effects of glycosaminoglycans (GAG) on cell-to-substratum adhesion and neurite elongation were examined in primary cultures of fetal rat hippocampal neurons using tissue culture dishes coated with GAGs coupled to dipalmitoylphosphatidylethanolamine (PE), a novel probe for biological functions of GAGs. Both chondroitin sulfate conjugate to PE (CS-PE) and hyaluronic acid conjugate to PE (HA-PE) promoted neurite elongation from neurons in a dose dependent manner when immobilized onto polylysine-coated dishes at various concentrations up to 1.0 microg/ml. The coating of CS-PE or HA-PE at a concentration higher than 1.0 microg/ml resulted in failure of neurite extension and adhesion of neurons to the substrata. In contrast, heparin conjugate to PE (HP-PE) did not exert any effects on neurite elongation or on cell attachment at these concentrations. These findings suggest that GAGs serve as a modulator for neurite elongation during neuronal network formation in the developing central nervous system. PMID- 10871048 TI - Catalytic properties of dihydroorotate dehydrogenase from Saccharomyces cerevisiae: studies on pH, alternate substrates, and inhibitors. AB - Yeast dihydroorotate dehydrogenase (DHOD) was purified 2800-fold to homogeneity from its natural source. Its sequence is 70% identical to that of the Lactococcus lactis DHOD (family IA) and the two active sites are nearly the same. Incubations of the yeast DHOD with dideuterodihydroorotate (deuterated in the positions eliminated in the dehydrogenation) as the donor and [14C]orotate as the acceptor revealed that the C5 deuteron exchanged with H2O solvent at a rate equal to the 14C exchange rate, whereas the C6 deuteron was infrequently exchanged with H2O solvent, thus indicating that the C6 deuteron of the dihydroorotate is sticky on the flavin cofactor. The pH dependencies of the steady-state parameters (k(cat) and k(cat)/Km) are similar, indicating that k(cat)/Km reports the productive binding of substrate, and the parameters are dependent on the donor-acceptor pair. The lower pKa values for k(cat) and k(cat)/Km observed for substrate dihydroorotate (around 6) in comparison to the values determined for dihydrooxonate (around 8) suggest that the C5 pro S hydrogen atom of dihydroorotate (but not the analogous hydrogen of dihydrooxonate), which is removed in the dehydrogenation, assists in lowering the pKa of the active site base (Cys133). The pH dependencies of the kinetic isotope effects on steady-state parameters observed for the dideuterated dihydroorotate are consistent with the dehydrogenation of substrate being rate limiting at low pH values, with a pKa value approximating that assigned to Cys133. Electron acceptors with dihydroorotate as donor were preferred in the following order: ferricyanide (1), DCPIP (0.54), Qo (0.28), fumarate (0.15), and O2 (0.035). Orotate inhibition profiles versus varied concentrations of dihydroorotate with ferricyanide or O2 as acceptors suggest that both orotate and dihydroorotate have significant affinities for the reduced and oxidized forms of the enzyme. PMID- 10871049 TI - Interaction of the peptide antibiotic alamethicin with bilayer- and non-bilayer forming lipids: influence of increasing alamethicin concentration on the lipids supramolecular structures. AB - Incorporation of the helical antimicrobial peptide alamethicin from aqueous phase into hydrated phases of dioleoylphosphatidylethanolamine (DOPE) and dioleoylphosphatidylcholine (DOPC) was investigated within a range of peptide concentrations and temperatures by time-resolved synchrotron X-ray diffraction. It was found that alamethicin influences the organizations of the non-bilayer forming (DOPE) and the bilayer-forming (DOPC) lipids in different ways. In DOPC, only the bilayer thickness was affected, while in DOPE new phases were induced. At low peptide concentrations (<1.10(-4) M), an inverted hexagonal (H(II)) phase was observed as with DOPE dispersions in pure buffer solution. A coexistence of two cubic structures was found at the critical peptide concentration for induction of new lipid/peptide phases. The first one Q224 (space group Pn3m) was identified within the entire temperature region studied (from 1 to 45 degrees C) and was found in coexistence with H(II)-phase domains. The second lipid/peptide cubic structure was present only at temperatures below 16 degrees C and its X-ray reflections were better fitted by a Q212 (P4(3)32) space group, rather than by the expected Q229 (Im3m) space group. At alamethicin concentrations of 1 mM and higher, a nonlamellar phase transition from a Q224 cubic phase into an H(II) phase was observed. Within the investigated range of peptide concentrations, lamellar structures of two different bilayer periods were established with the bilayer-forming lipid DOPC. They correspond to lipid domains of associated and nonassociated helical peptide. The obtained X-ray results suggest that the amphiphilic alamethicin molecules adsorb from the aqueous phase at the lipid head group/water interface of the DOPE and DOPC membranes. At sufficiently high (>1.10(-4) M) solution concentrations, the peptide is probably accommodated in the head group region of the lipids thus inducing structural features of mixed lipid/peptide phases. PMID- 10871050 TI - Purification and characterization of chorion peroxidase from Aedes aegypti eggs. AB - Previous study has shown that a peroxidase is present in the mature eggs of Aedes aegypti mosquitoes, and the enzyme is involved in the formation of a rigid and insoluble chorion by catalyzing chorion protein crosslinking through dityrosine formation. In this study, chorion peroxidase was solubilized from egg chorion by 1% SDS and 2 M urea and purified by various chromatographic techniques. The enzyme has a relative molecular mass of 63,000 as estimated by SDS-PAGE. Spectral analysis of the enzyme revealed the presence of the Soret band with a lambda(max) at 415 nm, indicating that chorion peroxidase is a hemoprotein. Treatment of the native enzyme with H2O2 in excess in the absence of reducing agents shifted the Soret band from 415 to 422 nm, and reduction of the native enzyme with sodium hydrosulfite under anaerobic conditions changed the Soret band from 415 to 446 nm. These results show that the chorion peroxidase behaves similarly to other peroxidases under oxidative and reductive conditions, respectively. Compared to other peroxidases, the chorion peroxidase, however, is extremely resistant to denaturing agents, such as SDS and organic solvents. For example, chorion peroxidase remained active for several weeks in 1% SDS, while horseradish peroxidase irreversibly lost all its activity in 2 h under the same conditions. Comparative analysis between mosquito chorion peroxidase and horseradish peroxidase showed that the specific activity of chorion peroxidase to tyrosine was at least 100 times greater than that of horseradish peroxidase to tyrosine. Chorion peroxidase is also capable of catalyzing polypeptide and chorion protein crosslinking through dityrosine formation during in vitro assays. Our data suggest that the characteristics of the chorion peroxidase in mosquitoes closely reflect its functions in chorion formation and hardening. PMID- 10871051 TI - An N-terminal peptide from link protein can stimulate biosynthesis of collagen by human articular cartilage. AB - Previous studies have shown that a peptide identical in sequence to the N terminal of link protein can function as a growth factor and up-regulate proteoglycan synthesis by human articular cartilage in explant culture (L. A. McKenna et al., Arthritis Rheum. 41, 157-162, 1998). The present study has extended these investigations to determine the effects of this peptide on the synthesis of collagen, another essential component of normal cartilage matrix. Explants from normal adult knee cartilage were maintained for periods of up to 8 days in medium with or without serum. Peptides were added during each day of culture. Synthesis of collagen was determined by the incorporation of [3H]proline into hydroxyproline and proteoglycans by incorporation of [35S]sulfate. The type of newly synthesized collagen was measured by SDS-polyacrylamide gel electrophoresis, fluorography, and immunoblotting. The link protein peptide stimulated synthesis of type II collagen in cartilage from a number of different subjects. Maximum up-regulation of synthesis was attained at a concentration of 100 ng/ml, similar to that observed previously for up-regulation of proteoglycan. Synthesis was up-regulated in both the presence and the absence of serum, although the overall rate of synthesis was greater when serum was added. The findings that this link peptide growth factor stimulated synthesis of proteins, including collagen, in a manner analogous to that shown previously for proteoglycans support the hypothesis that this peptide may have an important role in the feedback control of cartilage matrix synthesis. PMID- 10871052 TI - Redox capacity of cells affects inactivation of glutathione reductase by nitrosative stress. AB - Glutathione reductase (GR) plays a pivotal role in maintaining glutathione (GSH) in its reduced form. We have isolated a cDNA for rat GR and constructed a baculovirus system to produce recombinant GR on a large scale. This protein was purified by simple, two-step chromatographic procedure using DE52 and 2',5'-ADP Sepharose. Tissue distributions of GR were examined by Northern and Western blotting with a rabbit antibody to purified GR. GR was expressed in the order of reactivity; kidney, colon, liver, stomach, etc. Western blot analysis showed that both the cytosolic and the mitochondrial fractions of liver homogenate gave immunoreactive bands of similar size. This indicates that the same gene products exist in these fractions. Since nitric oxide (NO) produced under inflammatory conditions causes nitrosative stress and affects the redox states of surrounding tissues, we investigated the effects of NO donors on the enzymatic activities of purified GR. S-nitrosoglutathione (GSNO), 3-morpholinosydnonimine N ethylcarbamide (SIN-1), and S-nitroso-N-acetyl-D,L-penicillamine (SNAP) at 1 mM gave 39, 15, and 12% inhibitions, respectively. In RAW 264.7 cells the GR activity was reported to be inhibited by GSNO. In A549 cells, however, no such change in the activity, protein levels and mRNA of GR was noted. Since these cells have a much higher redox capacity than RAW 264.7 cells as judged by GR activity and thioredoxin reductase activity it wound minimize cellular damage, including inactivation of GR caused by nitrosative stress. PMID- 10871053 TI - Isolation of a proteinase with plasminogen-activating activity from Lachesis muta muta (bushmaster) snake venom. AB - A plasminogen activator enzyme (LV-PA) from Lachesis muta muta venom was purified to homogeneity using gel filtration and anion exchange chromatography. SDS-PAGE under reducing conditions showed a single protein band with an Mr of 33,000 Da. It is an acidic glycoprotein which activates plasminogen to plasmin indirectly, functioning via prior formation of a molecular complex, known as plasminogen activator. The purified preparation catalyzes the hydrolysis of several p nitroanilide peptide substrates containing Lys at the scissile bond. In contrast, no hydrolysis was detected on the synthetic substrates TAME and BAPNA, which contain arginine. By the use of the plasmin-specific chromogenic substrate Tos Gly-Pro-Lys-pNA, the preparation had a plasmin-like activity of 0.68 U/mg, which was 35.8-fold higher than that of the crude venom from which it was prepared. In vitro, fibrin hydrolysis using LV-PA as plasminogen activator displayed more similarity with the effect produced by streptokinase (SK). SDS-PAGE (10%) analysis showed a 115-kDa complex formation after incubation of plasminogen with either LV-PA or SK. At a molar ratio of 50:1 (fibrinogen:enzyme), the preparation exhibited weakly fibrinogenolytic activity. However, LV-PA is distinguished from thrombin in that it does not clot fibrinogen. After incubation of LV-PA with platelet-rich plasma, the enzyme (2 microM) showed no effect on platelet aggregation induced by ADP, epinephrine, or collagen. Comparison of the N terminal sequence of LV-PA with other snake venom plasminogen activators revealed that LV-PA exhibits a high degree of sequence identity with the TsVPA from Trimeresurus stejnegeri (90%) and with the Haly-PA from Agkistrodon halys (85%). LV-PA also has homology with other snake venom serine proteinases such as the thrombin-like/gyroxin analogue (38%) from bushmaster venom and with other coagulation serine proteases. The proteinase was readily inhibited by treatment with p-nitrophenyl p-guanidinebenzoate, p-aminobenzamidine, and phenylmethanesulfonyl fluoride but was not affected by metal chelators. PMID- 10871054 TI - Comparative study of the N-glycans of human monoclonal immunoglobulins M produced by hybridoma and parental cells. AB - Cell-cell hybridization is one method of establishing cell lines capable of producing an abundance of antibodies. In order to clearly characterize antibodies produced by hybridomas, the influence of cell-cell hybridization on the glycosylation of produced antibodies should be studied. In this report, we describe structural changes of the N-glycans in immunoglobulin M (IgM) produced by a hybridoma cell line termed 3-4, which was established through hybridization of an IgM-producing Epstein-Barr virus transformed human B-cell line termed No. 12, and a human myeloma cell line termed P109. We analyzed the structures of sugar chains on the constant region of the mu-chain of IgMs produced by parental No. 12 cells and hybridoma 3-4 cells. In both parental cells and hybridoma cells, the predominant structures at Asn171, Asn332, and N395 were fully galactosylated biantennary complex types, with or without core fucose and/or bisecting GlcNAc. However, the amount of bisecting GlcNAc was markedly decreased in the hybridoma cells. Therefore, the activity of UDP-N-acetylglucosamine:beta-D-mannoside beta 1,4-N-acetylglucosaminyltransferase (GnT-III) responsible for the formation of bisecting GlcNAc was measured in parental cells and hybridoma cells. No. 12 cells showed some GnT-III activity, whereas P109 cells showed no such activity. The corresponding level of activity observed in hybridoma 3-4 cells was much lower than that in No. 12 cells. The above results demonstrated a reduction in the intracellular activity of GnT-III in the hybridoma cells, which was largely due to the influence of P109 cells. Moreover, the sugar chain structures of IgMs produced by the cells reflected the level of GnT-III activity. PMID- 10871055 TI - Affinity purification of pancreastatin receptor-Gq/11 protein complex from rat liver membranes. AB - Pancreastatin, a chromogranin A derived peptide, exerts a glycogenolytic effect on the hepatocyte. This effect is initiated by binding to membrane receptors which are coupled to pertussis toxin insensitive G proteins belonging to the Gq/11 family. We have recently solubilized active pancreastatin receptors from rat liver membranes still functionally coupled to G proteins. Here, we have purified pancreastatin receptors by a two-step procedure. First, pancreastatin receptors with their associated Gq/11 regulatory proteins were purified from liver membranes by lectin absorption chromatography on wheat germ agglutinin immobilized on agarose. A biotinylated rat pancreastatin analog was tested for binding to liver membranes before using it for affinity purification. Unlabeled biotinylated rat pancreastatin competed for 125I-labeled [Tyr0]PST binding to solubilized receptors with a Kd = 0.27 nM, comparable to that of native pancreastatin. The biotinylated analog was immobilized on streptavidin-coated Sepharose beads and used to further affinity purify wheat germ agglutinin eluted receptor material. Specific elution at low pH showed that the receptor protein was purified as an 80-kDa protein in association with a G protein of the q/11 family, as demonstrated by specific immunoblot analysis. The specificity of the receptor band was assessed by chemical cross-linking of the purified material followed by SDS-PAGE and autoradiography. In conclusion, we have purified pancreastatin receptor as a glycoprotein of 80 kDa physically associated with a Gq/11 protein. PMID- 10871056 TI - Mechanistic studies of cytochrome P450 2B1 inactivation by xanthates. AB - Xanthates have previously been shown to inactivate the phenobarbital-inducible rat cytochrome P450 2B1 as well as its human homologue P450 2B6. The inactivation was mechanism-based and the loss in enzymatic activity was due to covalent binding of a reactive xanthate intermediate to the P450 2B1 apoprotein. In this report, we investigated various mechanistic events to elucidate the individual step(s) in the P450 catalytic cycle that are compromised due to the inactivation by xanthates. Different xanthates displayed typical type I binding spectra and the spectral binding constants were in the low-millimolar range. A dramatic loss in 7-ethoxy-4-(trifluoromethyl)coumarin activity was observed when P450 2B1 was incubated with five different xanthates in the presence of NADPH. With the exception of the C14 xanthate, virtually no loss of absorbance at 418 or 450 nm in the reduced-CO complex was observed. Long-chain xanthates were able to affect the rate of the first electron transfer in the P450 catalytic cycle by stabilizing the heme in its low-spin state. n-Octyl xanthate (C8) metabolism led to very little observable oxy-ferro intermediate complex formation. The alternate oxidant tert-butyl hydroperoxide was able to support the inactivation reaction of C8 in the absence of reductase or NADPH. The rates of reduction of native, C8 exposed, and C8-inactivated P450 2B1 were measured. The C8-inactivated P450 had a 62% lower rate of reduction in the absence or presence of benzphetamine compared to the native enzyme. Product formation of the three enzyme preparations was quantified with benzphetamine as the substrate. The C8-inactivated P450 2B1 exhibited a much lower rate of NADPH consumption and formation of formaldehyde. However, the ratio of H2O2 to formaldehyde production increased from 1:1 for the native enzyme to 2.8:1 for the inactivated P450. Together these observations indicate that the covalent modification of P450 2B1 by a reactive intermediate of xanthates reduces the rate of the first electron transfer by the reductase and also leads to uncoupling of electron transfer from product formation by diverting a greater proportion of the electrons to H2O2 formation. PMID- 10871057 TI - Identification of active-site residues in Bradyrhizobium japonicum malonyl coenzyme A synthetase. AB - Malonyl-CoA synthetase (MCS) has been previously purified and characterized from Bradyrhizobium japonicum USDA 110. The gene encoding this enzyme is now cloned, sequenced, and expressed in Escherichia coli. The enzyme contains 509 amino acid residues, with a calculated molecular mass of 55,239 Da. The recombinant enzyme was also purified from the transformed E. coli. The enzyme was essentially indistinguishable from the MCS of B. japonicum by the criteria of polyacrylamide gel electrophoresis and biochemical properties. Based on inhibitor studies of Rhizobium trifolii MCS reported previously and database analysis, Arg173, Lys175, His211, and Glu308 were selected for site-directed mutagenesis in order to identify amino acid residues essential for substrate binding and/or catalysis. Five different mutant enzymes (R173G, K175M, H211L, K175M/H211L, and E308Q) were prepared and then subjected to steady-state kinetic studies. The kinetic data measured for the mutants suggest that Lys175 and His211 participate in the formation of malonyl-AMP, whereas Glu308 may play a role in malonate binding. PMID- 10871058 TI - Characterization and functional analysis of two common human cytochrome P450 1B1 variants. AB - Cytochrome P450 1B1 (CYP1B1) is a human extrahepatic P450 that activates procarinogens, metabolizes 17beta-estradiol, and may well have a role in the pathogenesis of some forms of cancer. Besides rare deleterious mutations reported for the CYP1B1 gene, six single-nucleotide polymorphisms have been reported, of which four cause amino acid exchanges. We have expressed two of the common CYP1B1 alleles in yeast cells and mammalian COS-1 cells in order to functionally characterize the alleles with respect to kinetic properties and protein stability. The CYP1B1.2 variant contains the two linked amino acid substitutions R48G and A119S compared to CYP1B1.1. The kinetic parameters of two structurally unrelated CYP1B1 substrates for the two variants were examined. No kinetic differences were seen of 17beta-estradiol hydroxylation activities between the two CYP1B1 variants and an only minor increase in the apparent Km for ethoxyresorufin was observed for CYP1B1.2. It therefore appears that they have very similar catalytic properties and the substitutions do not appear to alter CYP1B1 catalytic function. The two CYP1B1 variants were similarly stable when expressed in mammalian COS-1 cells, indicating that the substitutions have no effect on protein folding or stability. The combined results indicate that these two CYP1B1 variants show very similar properties with respect to catalytic activities and protein stability and do not alter CYP1B1 function. PMID- 10871060 TI - Stability determines formation rate of four-helix-bundle protein. PMID- 10871059 TI - T cell receptor assembly and expression in the absence of calnexin. AB - Most subunits of the alphabeta deltaepsilon gammaepsilon zetazeta T cell antigen receptor (TCR) complex associate with the molecular chaperone calnexin shortly after their synthesis in the endoplasmic reticulum, including clonotypic TCRalpha,beta molecules and invariant CD3gamma,delta,epsilon chains. While calnexin interaction is suggested to be important for the stability of newly synthesized TCRalpha subunits, the role of calnexin in the survival and assembly of remaining TCR components is unknown. Here we evaluated the expression of TCR proteins in CEM T cells and the calnexin-deficient CEM variant CEM.NK(R). We found that CEM and CEM.NK(R) cells constitutively synthesized all TCR subunits except for TCRalpha and that CD3gamma,delta,epsilon components and CD3-beta complexes were effectively assembled together in both cell types. The stability and folding of core CD3epsilon chains were similar in CEM and CEM.NK(R) cells. Interestingly, TCRalpha synthesis was differentially induced by phorbol myristate acetate treatment in CEM and CEM.NK(R) cells and TCRalpha proteins synthesized in CEM.NK(R) cells showed reduced survival compared to those made in CEM cells. Importantly, these data show that TCR complexes were inducibly expressed on CEM.NK(R) cells in the absence of calnexin synthesis. These results demonstrate that TCR complexes can be expressed in the absence of calnexin and suggest that the role of calnexin in the quality control of TCR assembly is primarily restricted to the stabilization of newly synthesized TCRalpha proteins. PMID- 10871061 TI - Use of sulfhydryl-directed inhibitors in vitro to distinguish activities of the mitochondrial and plastidic forms of pyruvate dehydrogenase. PMID- 10871062 TI - Extranodal marginal zone B-cell lymphoma of the skin: a morphologic and immunophenotypic study of 11 cases. AB - Extranodal marginal zone B-cell lymphoma (MZBL) is a recently recognized low grade lymphoma that has been well described in other organs such as the stomach and salivary gland. It has only recently been described in skin, where it may be difficult to distinguish from reactive processes and other types of B-cell lymphoma such as follicle center lymphoma. These cases may have been classified as pseudolymphomas in the past. Extranodal MZBL was referred to as mucosa associated lymphoid tissue (MALT) lymphoma before the Revised European-American Classification of Lymphoid Neoplasms was published in 1994. Important histologic features that aid in the diagnosis of MALT lymphoma are atypical lymphocytes (centrocyte-like and monocytoid B cells) often admixed with plasmacytoid lymphocytes, a prominent plasma cell component, lymphoepithelial lesions, intranuclear inclusions (Dutcher bodies), and reactive germinal centers that may be colonized by neoplastic cells. Immunophenotypic studies demonstrating a B-cell phenotype, light chain restriction, coexpression of CD43, and staining of atypical lymphocytes with bcl-2 support a diagnosis of MALT lymphoma. We studied 11 cases of extranodal MZBL of the skin from the Armed Forces Institute of Pathology files. There were six women and five men ranging in age from 30 to 69 years (median, 54 years). The anatomical sites included the trunk, head and neck areas, and upper extremities. There were no other sites of disease besides the skin in any of the cases. The follow-up period ranged from 5 months to 8 years (median, 24 months). Histologic results included an atypical lymphoid infiltrate with B-cell phenotype, reactive germinal centers, and a variable plasma cell component in all cases. No Dutcher bodies or lymphoepithelial lesions were noted. Extranodal MZBL of skin is a diagnostic challenge because of a heterogeneous cellular infiltrate that may be interpreted as a reactive process. The most significant neoplasm with which it is confused is follicular lymphoma. It is important to recognize the characteristic histologic and immunophenotypic features of extranodal MZBL so that the appropriate therapeutic approach may be applied. PMID- 10871063 TI - Mycosis fungoides with CD30-positive cells in the epidermis. AB - Large numbers of CD30-positive tumor cells are not typically observed in mycosis fungoides (MF), but CD30 expression may occur on large cells of MF that have transformed into high-grade large cell lymphoma. Of 202 patch/plaque phase MF cases studied by immunohistochemistry, we encountered 4 patients with patch-phase MF at stage Ia or Ib, whose lesions contained a high proportion (greater than 50%) of CD30-positive tumor cells within the epidermis. The morphologic and immunohistochemical features of these four cases were otherwise similar to those of other patch-phase MF cases, and were different from those of pagetoid reticulosis. More importantly, the clinical behavior of these cases did not differ from that of other cases of early MF without CD30 expression. The mechanism underlying the high levels of CD30 expression by epidermotropic tumor cells is unknown. With increased use of the CD30 immunohistochemical stain in the diagnosis of cutaneous lymphomas, similar cases are likely to be encountered, and perhaps will be evaluated for their clinical behavior. PMID- 10871064 TI - The relationship between melanocytes and peripheral nerve sheath cells (Part I): melanocytic nevus (excluding so-called "blue nevus") with peripheral nerve sheath differentiation. AB - Among thousands of specimens of melanocytic nevi, not including giant congenital melanocytic nevus or blue nevus, 42 melanocytic nevi that showed peripheral nerve sheath differentiation were collected. The patterns of melanocytic nevi with peripheral nerve sheath differentiation may be classified into three groups: 1) "neurotized and neural nevi" with nests of "neuroid cords" and "nevic corpuscles" (the most common pattern); 2) nerve fascicle-like structures with no relation to neurotized and neural nevi; and 3) palisading melanocytes of a nevus in nests of conventional melanocytic nevi (a rare pattern). Each pattern may represent a different expression of nerve sheath differentiation in melanocytic nevi. Some melanocytic nevi with nerve fascicle-like structures show discrete structures closely resembling authentic nerve fascicles, confirming a close relationship between melanocytes and peripheral nerve sheath cells (Schwann cells and probably perineurial cells in part) and suggesting derivation of the two types of cells from common precursor cells of the neural crest and their de novo development in the dermis rather than by Abtropfung of melanocytes from the epidermis. In addition, the high prevalence of Unna, Miescher, and superficial congenital nevi in melanocytic nevi with peripheral nerve sheath differentiation suggests a different character or process for these congenital melanocytic nevi than for Clark and Spitz nevi (junctional and compound types). PMID- 10871065 TI - The relationship between melanocytes and peripheral nerve sheath cells (Part II): blue nevus with peripheral nerve sheath differentiation. AB - Peripheral nerve sheath differentiation was studied in 120 specimens of blue nevi (112 specimens of "common" blue nevi and 8 specimens of "cellular" blue nevi). In 10 of 112 common blue nevi, fascicles of pigmented dendritic melanocytes or less pigmented spindle-shaped melanocytes with S-shaped nuclei were associated with wavy delicate collagen bundles. In 4 of these 10 specimens, the melanocytes showed a perifollicular arrangement. These nerve fascicle-like structures were seen in some cellular blue nevi (5 of 8 specimens). Structures closely resembling authentic nerve fascicles were not observed in common or cellular blue nevi. The fascicles with S-shaped nuclei and fibrillary collagenous tissue observed in blue nevi (which were well detected in the cellular type but rarely found in the common type) may be peripheral nerve sheath features and perhaps evidence that dermal dendritic melanocytes in the reticular dermis may have arisen within a peripheral nerve sheath milieu from primitive fibroblast-like precursors. Some of the examples presented here may be identical to those reported for "pilar neurocristic hamartoma" or "neurocristic hamartoma." I also speculate on the pathogenesis of blue nevi based on the observations made in this study. PMID- 10871066 TI - Immunohistochemical distinction of epithelioid histiocytic proliferations from epithelioid melanocytic nevi. AB - Histiocytic tumors can be confused with melanocytic nevi and malignant melanoma and vice versa. To explore the use of immunohistochemistry for this diagnostic problem, we examined the expression of S-100 protein, gp100 (the antigen recognized by HMB-45), tyrosinase (T311), Melan-A (A103), Factor XIIIa (FXIIIa), and CD68 in 10 juvenile xanthogranulomas (JXGs), five epithelioid histiocytomas (EHs), and 15 melanocytic nevi composed of large epithelioid cells. All epithelioid melanocytic nevi were immunoreactive for Melan-A, tyrosinase, and S 100 protein in most melanocytes. Four nevi were completely negative with HMB-45. Nine nevi had only a minor HMB-45-positive component in the superficial dermis. Two nevi were diffusely HMB-45-positive. Melanocytes in all nevi were completely negative for FXIIIa. Thirteen nevi were completely negative for CD68. Two nevi contained rare cells with weak staining for CD68. All 15 histiocytic proliferations were completely negative for Melan-A, tyrosinase, and gp100. They lacked expression of S-100 protein or had at most 10% immunopositive cells. In JXGs, most cells were strongly reactive for CD68, although only a few were positive for FXIIIa. In EHs, 40% to 60% of cells were immunoreactive for FXIIIa, and only 20% to 30% were positive for CD68. Our results demonstrate that Melan-A and tyrosinase are sensitive and specific markers to distinguish epithelioid melanocytic nevi from epithelioid histiocytic tumors. PMID- 10871067 TI - Histological findings after brown recluse spider envenomation. AB - Histologic specimens from 41 rabbits were studied for changes resulting from the manual injection of brown recluse spider venom. Major findings included a mixed inflammatory cell infiltrate, coagulative tissue necrosis, and vasculitis. All specimens demonstrated a well-delineated zone of eosinophilic staining recognizable as "mummified" coagulative necrosis of the epidermis and dermis. A dense band of neutrophils bordered the zone of necrosis. Immediately adjacent to the neutrophilic band, small vessel vasculitis was a universal finding. Degranulated eosinophils and neutrophils and macrophages filled with eosinophilic granules were common. Inflammatory foci were often centered on groups of lipocytes within the dermis. Large vessel vasculitis resembling that seen in polyarteritis nodosa was present deep to 7 of the 40 eschars. Large vessel vasculitis may contribute to the large zones of necrosis seen after some brown recluse spider bites. Eosinophils may play a role in tissue damage after envenomation. PMID- 10871068 TI - Staining of melanocytic neoplasms by melanoma antigen recognized by T cells. AB - We stained benign melanocytic nevi and malignant melanoma with antibodies to melanoma antigen recognized by T cells (Mart-1) to determine if this was useful in differentiating benign from malignant melanocytic neoplasms. Forty-five primary malignant melanomas and 71 benign melanocytic nevi were stained with antibodies to Mart-1. Two cases of malignant melanoma metastatic to lymph node and three cutaneous metastases of malignant melanoma were also stained. The degree of staining was graded into diffuse positive staining, focal positive staining, and negative staining. Thirty-six of 45 primary malignant melanomas stained diffusely positive with antibodies to Mart-1. This included three of five desmoplastic malignant melanomas that showed positive staining. Four melanomas showed faint or focal positive staining. One of two metastases to lymph node showed strong positive staining and one showed no staining. All three cutaneous metastases showed diffuse positive staining. Sixty-one of 71 melanocytic nevi showed no staining or faint staining with antibodies to Mart-1. Ten of 71 melanocytic nevi showed strong positive staining. The majority of these were congenital nevi. Staining with antibodies to Mart-1 antigen was a useful marker of malignant melanoma. However, staining may also be seen in benign melanocytic neoplasms. The presence or absence of staining for Mart-1 antigen cannot be used to differentiate benign melanocytic nevi from malignant melanocytic tumors. PMID- 10871069 TI - Desmoplastic leiomyosarcoma of the skin. AB - This report focuses on two unusual cases of cutaneous leiomyosarcoma composed of sparse numbers of neoplastic cells embedded in abundant sclerotic stroma throughout the entire neoplasm. To the best of our knowledge, only one example of this rare lesion has been described previously as "sclerotic primary cutaneous leiomyosarcoma." However, the resemblance of this tumor to other desmoplastic tumors of the skin is striking and, therefore, we propose the term desmoplastic leiomyosarcoma of the skin for this neoplasm. Because of the sparse cellularity and the abundant stroma, desmoplastic leiomyosarcoma of the skin can be easily misinterpreted, especially in small biopsies. It should be included in the differential diagnosis of inflammatory skin diseases associated with sclerosis, such as radiation dermatitis, and of desmoplastic tumors of the skin, including desmoplastic melanocytic nevus, desmoplastic melanoma, and desmoplastic squamous cell carcinoma. PMID- 10871070 TI - Primary cutaneous B-cell lymphoma of the leg in a chronic lymphedematous extremity. AB - This is a case report of a woman who had chronic lymphedema on one leg and who developed a primary cutaneous large B-cell lymphoma of the leg at that site. She received radiotherapy and did not show any systemic involvement thereafter. Other neoplasms may appear in a clinical setting of chronic lymphedema, namely, lymphangiosarcoma (Stewart-Treves), melanoma, and metastatic carcinoma. There are four other reports in the English literature of cutaneous lymphoma arising in an extremity with chronic lymphedema. PMID- 10871071 TI - Apocrine poroma with sebaceous differentiation. AB - Poromas have been classified as eccrine neoplasms, but several recent reports of poroid tumors with sebaceous, follicular, and apocrine differentiation have challenged this concept. We report a case of apocrine poroma with sebaceous differentiation. A 69-year-old man presented with an asymptomatic elevated erythematous plaque. Histopathology revealed cellular nests composed of cuboidal poroid cells and sebocytes. The nests varied in size and were entirely intraepidermally arranged in a growth pattern similar to that of hidroacanthoma simplex. Given the common embryologic origin of folliculosebaceous and apocrine units, we believe that this lesion represents an apocrine poroma with sebaceous differentiation. PMID- 10871072 TI - Adult rhabdomyoma: report of two cases of rhabdomyoma of the lip and of the eyelid. AB - We describe two cases of adult rhabdomyoma. One was located in the lip of a 66 year-old woman and was removed because it was clinically suspicious for infiltrating carcinoma. The other arose in the eyelid of a 60-year-old woman with a glass eye and was initially interpreted as a reactive process due to the prosthesis. Both lesions were composed of cells with oval nuclei and deeply eosinophilic cytoplasms with occasional cross striations. Immunoreactivity for desmin and myoglobin excluded the diagnosis of other tumors with similar morphology. The unusual association of the eyelid tumor with the prosthesis suggests a role for chronic irritation in the pathogenesis of rhabdomyoma. PMID- 10871074 TI - Histopathology of psoriasis treated with zinc pyrithione. AB - Psoriasis is still a relatively poorly understood inflammatory dermatosis that is resistant to many therapies. Because the pathogenesis is poorly understood, rational treatment is elusive. Until recently, the North American public was able to achieve successful resolution with an over-the-counter topical preparation marketed for dandruff and seborrheic dermatitis called SkinCap, which has now been withdrawn from the market. The purpose of this study was to examine the histologic changes induced by this preparation in a well-developed psoriatic plaque. Serial punch biopsies were taken over a 2-week period during which time SkinCap was applied topically twice daily. The biopsies were examined histologically, and features were evaluated semiquantitatively. The classic histologic features of psoriasis resolved completely over 2 weeks, with the reversal beginning with disappearance of neutrophils and the most striking finding being prominent apoptosis at 48 hours. The mechanism of this normalization is unknown. Hypotheses include blockage of cytokine and growth factor effect at some level and induction of apoptosis. PMID- 10871073 TI - Pleomorphic angioleiomyoma. AB - We describe two cases of pleomorphic angioleiomyoma. In one case, a 46-year-old man presented with a single nodule on his scrotum of 1 year's duration, and in another, a 38-year-old woman presented with a single nodule on her right knee of 1 year's duration. In both cases, histopathologic examination showed a well circumscribed nodule composed of smooth muscle and numerous veins and capillaries. Contrary to the ordinary angioleiomyoma, marked nuclear pleomorphism was noted. Although mitoses were rare, immunohistochemistry revealed many tumor cells that were positive for proliferating cell nuclear antigen, Ki-67, and p53, indicating that the pleomorphic appearance does not simply represent a degenerative change of some tumor cells. PMID- 10871075 TI - Lamellar ichthyosis: response to etretinate with transglutaminase 1 recovery. AB - We studied a case of typical lamellar ichthyosis before and after etretinate treatment for the expression of transglutaminase 1 (TGK) and the presence or absence of the marginal band. Before the treatment, TGK was undetectable, although two other components of the marginal band, loricrin and involucrin, were detected by immunostaining in a normal pattern. The marginal band of the corneocytes was either thin and irregular or completely absent by electron microscopic study. After therapy with etretinate, 50 mg/day, the patient' s skin improved, and biopsies were taken at 4 and 8 months. Transglutaminase 1 became detectable by immunostaining. The marginal band was still absent in most corneocytes. PMID- 10871076 TI - Dermatofibroma with diffuse eosinophilic infiltrate. AB - Dermatofibroma, a common form of benign fibrous histiocytoma, is characterized by the presence of different cell types consisting of fibroblastic, histiocytic, and even multinucleated cells. Dermatofibromas are always accompanied by an inflammatory lymphoid cell reaction, including B cells and T cells at their border. To our knowledge, however, there is no report of an infiltration of eosinophils within this tumor. We describe a dermatofibroma on the pretibial region of an 8-year-old boy. Microscopic examination revealed a diffuse eosinophilic infiltration within the dermatofibroma, which was typically composed of histiocytic cells and mature collagen. The diagnosis of dermatofibroma was confirmed by demonstrating the immunoreactivity of tumor cells to an anti-factor XIIIa antibody. PMID- 10871078 TI - A critical review of apoptosis in historical perspective PMID- 10871077 TI - A vessel runs through it. PMID- 10871079 TI - Characteristics of the Birt-Hogg-Dube/Hornstein-Knickenberg syndrome. PMID- 10871080 TI - "Early" in Kaposi sarcoma: signifier and signified. PMID- 10871081 TI - Reconstruction of a parotidectomy defect using a paraumbilical perforator flap without deep inferior epigastric vessels. AB - A 37-year-old woman who underwent a parotidectomy for acinic cell carcinoma was referred for correction of the resulting defect. As an assistant principal, the patient was often in public and, because she was somewhat self-conscious about her facial deformity, she sought reconstruction. Physical examination revealed a pre-auricular soft-tissue defect that measured approximately 5 x 5 cm. After consultation with the patient, microsurgical transfer of fat from the lower abdomen based on branches of the deep inferior epigastric vessels, rather than the deep inferior epigastric artery and vein itself, was planned. A vertical skin ellipse measuring 6 x 5 cm was raised from the right lower abdomen with a pedicle consisting of only a branch of the deep inferior epigastric artery and vein. The donor vessels were then microsurgically anastomosed to the superficial temporal artery and vein on the left side of the face. Two weeks postoperatively the flap was defatted, with removal of the skin-monitoring island. The patient continues to do well with a normal contour of the face and decreased anxiety secondary to correction of her facial deformity. PMID- 10871082 TI - Vascularized metacarpal bone graft for scaphoid non-union and Kienbock's disease. AB - A new vascularized bone graft from the base of the second metacarpal was used to treat scaphoid non-union and Kienbock's disease. In two patients with scaphoid non-union, the procedure promoted healing, even when the proximal segment was poorly vascularized. In one patient with Kienbock's disease, the graft held stable and wrist pain was markedly reduced. Further clinical experience may establish this procedure as an option for carpal bone fractures or disease. PMID- 10871083 TI - It is never too late: limb salvage following a half-century of long-bone osteomyelitis. AB - The author presents a case of osteomyelitis of the long bones, of over a half century duration. An aggressive surgical approach comprising wide debridement of all compromised soft tissues and generous ostectomies, followed by myocutaneous flap reconstruction, achieved limb salvage. PMID- 10871084 TI - Recalcitrant post-surgical neuropathy of the ulnar nerve at the elbow: treatment with autogenous saphenous vein wrapping. AB - Surgical decompression or transposition is generally efficacious for cubital tunnel syndrome. However, recurrence is not rare and its management is both challenging and difficult. Four patients with refractory cubital tunnel syndrome were operated on with the vein-wrapping technique, using the autologous saphenous vein. A total of 16 operative procedures were performed on these patients prior to wrapping the ulnar nerve with a saphenous vein graft. The mean patient age was 43 years (range: 30 to 54 years) and the mean follow-up was 34 months (range: 24 to 44 months). All patients reported significant pain relief and improvement in sensation. Two-point discrimination and EMG findings also improved. This is the first study reporting long-term results of the vein-wrapping technique for the treatment of recalcitrant cubital tunnel syndrome. PMID- 10871085 TI - Lateral arm fascial flap: microarterial anatomy and potential clinical applications. AB - Previously, muscle flaps and the omentum have been used to indirectly vascularize tissues. Induction of synangiogenesis, or indirect vascularization through the formation of collateral vessels, occurs through the development of vascular connections at the interface between the donor and recipient tissues. Unfortunately, muscle and omental flaps are bulky and, when used to salvage ischemic hands and digits, may limit digital range of motion. Additionally, disadvantages to using omentum include a requirement for an intraabdominal procedure and a lack of subsequent donor tissue if the contralateral limb becomes involved at a later time. The purpose of this anatomic study was to develop a customized lateral arm fascial flap (LAFF) which may be used for flap prefabrication or synangiogenesis of non-bypassable ischemia. Detailed anatomic dissections were performed to more thoroughly define the microvascular anatomy of the LAFF. Computer analysis of the data was performed to demonstrate the potential clinical application of using the LAFF. Dissections revealed a consistent pattern of vessels branching within the lateral arm fascia and to the neighboring musculature. In order to optimize the surgical use of available tissue, computer-aided design techniques were used to model a reliable fascial free flap for inducing synangiogenesis while imparting minimal donor-site morbidity. Anatomic studies of the LAFF revealed pitfalls in flap dissection, while computer-generated models illustrated the detailed microarterial anatomy of the LAFF and potential limitations in flap design. Potential clinical applications for use of this low-profile fasciovascular conduit are noted. PMID- 10871086 TI - Vascularized plantaris tendon graft: anatomic study of the donor. AB - In order to determine new vascularized donor tendons for grafting, a detailed anatomic study of the plantaris tendon and vascular connections with the posterior tibial artery or its branches was undertaken in 20 legs of 10 fresh adult cadavers. A histologic evaluation was also undertaken. Findings demonstrated that there is a close vascular connection between the crural fascial linked section of the plantaris tendon and the posterior tibial artery. Through its rich surrounding paratenon, the blood supply of the tendon is provided by two to four transfascial branches of the posterior artery in the lower-middle portion of the leg. Out of these branches, one or two anastomosing arteries (more than 1.0 mm in diameter), together with accompanying veins, consistently emerge 5 to 8 cm from the insertion of the plantaris tendon. Histologic observations demonstrate the reliable vascularity of the paratenon and crural fascia. The authors consider this vascularized tendon donor a good option for grafting. A composite tendofascial flap with vascularized pedicle from the posterior tibial artery or an isolated vascularized tendon graft from its branches would both be alternative techniques for clinical vascularized tendon grafting. PMID- 10871087 TI - The life and work of Theodore Schwann. PMID- 10871088 TI - Effect of distant septic foci on the patency of microvascular anastomoses. AB - Despite all technical improvements, some free-tissue transfers are still subject to failure in the early postoperative period due to anastomotic occlusion. Local or distant foci of sepsis may be present in most of these cases. Using 40 Sprague Dawley adult small female rats, distant septic abscesses were formed with Pseudomonas aeruginosa (including 2 x 10(8) bacteria/ml) suspension. Microarterial and microvenous anastomoses were carried out and animals were divided into experimental and control groups. With the use of postoperative exploration, anastomotic patency was evaluated, cultures of wound, blood, and tissue were taken, and the anastomosis line and thrombi were evaluated using different staining techniques on prepared histopathologic sections. Compared to the control groups, thrombus formation in animals with distant septic foci, both with arterial and microvenous anastomosis, was found to be greater, and the difference was statistically significant. Colonies of Pseudomonas were not demonstrated in samples taken from wound infections, hemocultures, cultures of vessel wall and thrombi, and in histopathologic sections. In this experimental model, it appears that distant septic foci have an occluding effect on the microanastomosis. PMID- 10871089 TI - Long-term evaluation of rat peripheral nerve repair with end-to-side neurorrhaphy. AB - This study was designed to assess long-term reinnervation of end-to-side neurorrhaphy in the rat. The cut right peroneal nerve was repaired and sutured to the side of the intact tibial nerve. Both the extent of reinnervation and the integrity of the intact donor nerve were evaluated in 48 Sprague-Dawley rats randomly treated with fresh or delayed nerve repair with or without perineurotomy. Evaluations included nerve conduction velocity (NCV) of both the peroneal and tibial nerves, dry muscle weight, and histologic examination (neurofilament stain and morphometric assessment) at 8 and 12 months postoperatively. Although animals treated with perineurotomy tended to have better NCV and dry muscle weight recovery than those without, the difference was not statistically significant. No difference was observed between fresh and predegenerated nerve repair. The mean total (all four subgroups) NCV recovery rates were 87 percent and 94 percent for the peroneal nerve, and 93 percent and 95 percent for the tibial nerve, compared to the contralateral intact nerves, at 8 and 12 months, respectively. Tibialis anterior muscle mass measurements revealed a recovery in dry muscle weight of about 85 percent and 89 percent at 8 and 12 months, respectively, compared to the intact contralateral tibialis anterior muscles. Histologic studies with neurofilament staining revealed numerous axons at the distal end of the peroneal nerve in all groups, indicative of myelinated axonal regeneration. Morphometric analysis demonstrated that the presence of a window in the perioneurium improved the histologic picture. The mean number of myelinated fibers at 12 months postoperatively was significantly higher in animals with a perineurotomy window (compared to without) in both fresh and predegenerated nerve repair subgroups, respectively (p <0.05). These results indicated that end-to-side neurorrhaphy permits axonal regeneration from the intact donor nerve and is associated with satisfactory recovery. The effect of the procedure on the donor nerve was negligible. PMID- 10871090 TI - Ultrastructure of early axonal regeneration in an end-to-side neurorrhaphy model. AB - The ultrastructure of the early regenerative response in an end-to-side neurorrhaphy rat model was studied using transmission electron microscopy. The ipsilateral saphenous nerve was grafted to the sciatic nerve under the following conditions: Group 1, the epineurium and perineurium of the sciatic nerve remained intact; Group 2, an epineurial and perineurial window was created at the site of the lateral neurorrhaphy; Group 3, the same as in Group 2 and, in addition, the sciatic nerve sustained a partial neurectomy. Rats were perfused through the heart with fixative containing 2 percent paraformaldehyde and 2.5 percent glutaraldehyde in 0.1 M cacodylate buffer (pH 7.4) at 4, 8, 12, 24 and 48 hr after surgery. In Group 1, no regenerating axons were observed and the myelin sheath in the donor nerve did not demonstrate any degenerative changes through 48 hr. In Group 2, an increased diameter of the unmyelinated axons and growth cones was observed in the donor nerve proximal to the coaptation site after 12 hr. Degenerative changes in the myelin sheath were observed after 12 hr within the several layers under the coaptation site. In Group 3, many growth cone-like structures were observed in the area proximal to the coaptation site after 12 hr. After 24 hr, proximal regenerating axons elongated to the coaptation site and, at 48 hr, many regenerating nerves grew inside the Schwann cell basement membrane of the graft nerve. These results indicate that the perineurial window and nerve graft are the critical conditions for inciting nerve regeneration in the donor nerve. PMID- 10871091 TI - Pedicle variation of the free gracilis muscle flap in the rat. PMID- 10871092 TI - New technique for endoscopic sural nerve harvest. PMID- 10871093 TI - Sensitivity, specificity and positive predictive value of patch testing: the more you test, the more you get? ESCD Working Party on Epidemiology. AB - Pathophysiological variability affects the results of patch testing. In addition, even a minimal degree of test-imprecision due to this variability has a number of important statistical consequences for the analysis and interpretation of any patch test data set. One such statistical phenomenon that is often overlooked is the dependence of the positive predictive value (i.e., the predictive value of a positive patch test) on sensitivity and specificity, the impact of which is heavily dependent on the proportion of truly allergic subjects that are studied. A 2nd important issue is the fact that patch testing is performed in series, which means multiple tests. If we assume, for example, a patch test series of only 10 allergens, then it can be demonstrated that there is a random probability of over 40%) to find, simply by chance, for at least 1 allergen, a statistically significant difference between 2 groups of patients. Comparison of the results of series between patients calls for statistical adjustments in order to prevent erroneously positive differences and/or associations. PMID- 10871094 TI - Effect of dark test-substance pigmentation on skin perfusion assessments and effect of test technique on balsam of Peru patch-test results. AB - 13 balsam of Peru (Myroxylon Pereirae) patch-test-positive subjects are re-tested with 25% balsam of Peru in petrolatum and with serial doses printed on polyester squares. All substances are applied with tape strips for 3, 6, 24 (1 day [D]), 48 (2D), 72 (3D) and 96 h (4D) on each subject and for 96 h (4D) with plastic foils. Tests are followed visually and with perfusion assessments from 3 h to 9 days. Results show that pigment remnants following detachment of patches affect perfusion assessments. Such effect due to pigment is supported by readings of patch tests through the petrolatum test substance while applied with transparent foils. For most reactions, good agreement is observed between the assessment techniques when peak assessment values of reactions are compared. There is inter individual variation in perfusion with identical tests. With the petrolatum test substance, increased visible reactivity was observed when the application time was extended up to 24 h (1D), while extension of application time increased perfusion in most cases except for an extension from 24 (2D) to 48 h (4D) where decreased perfusion resulted in most cases. Dose and application time did not affect the timing of highest reactivity of reactions in most cases. PMID- 10871095 TI - Assessment of balsam of Peru patch tests. AB - To find an ideal test technique for as low a dose of balsam of Peru (Myroxylon Pereirae) as possible, subjects testing positive to balsam of Peru are re-tested with a 25% concentration of balsam of Peru in petrolatum. Applications are with Finn Chambers for 6 different application times, and directly by foils for 96 h (4 days (D)). The goals are to confirm which subjects are positive and which are not, and, using that information, to see if it is possible to distinguish between these 2 groups, tested concomitantly at much lower serial dose levels, in terms of perfusion or by visual assessments. 5 different serial doses are applied with strips for 3-96 h (4D) and with foils for 96 h (4D). The Finn Chamber tests allow a distinction between visually positive and negative subjects supported by perfusion assessments. With the foils, a 24x lower serial dose level than with the 25% test substance is sufficient to distinguish between positive and negative subjects in terms of perfusion values. This approach requires readings up to 9 days. With this test, the visual approach yields only 3 of 10 positive subjects. This study demonstrates that a lower test dose is possible with perfusion assessments compared to visual ones. PMID- 10871096 TI - The outcome of an additional patch-test reading on days 6 or 7. AB - In a retrospective study (period 1997-1998), the usefulness of an additional patch-test reading on day (D) 6 or 7 was investigated. In 62 out of 760 patients (8.2%), 77 late-positive reactions, those manifest after D3, were seen. 4 late reactions of allergens of which a related allergen reacted earlier (at D2 or D3) in the same patient were not incorporated in the study as late-positive. Allergens most involved in producing late-positive reactions were nickel sulfate (20 reactions), neomycin sulfate (7), tixocortol-21-pivalate (5), p.t. butylphenol formaldehyde resin (5) and Cl+ Me isothiazolinone (5). Special attention is paid to ?+ reactions at D6 or D7, because these reactions could add extra information to the outcome of patch-test readings, considering that this might be a group of allergens that has been positive between D3 and D6/7 or become positive later on. When this group is included, the total number of late positive reactions is 105 in 88 out of 760 patients (11.6%). By doing D3 readings in combination with D6 or D7 readings only, one would miss 24 positive reactions. However, we concluded that it was preferable to have a reading on D2 and D3. An extra reading on D6 or D7 is very useful as it gives additional information in 8.2% of patch-tested patients. PMID- 10871097 TI - Multicentre study for the development of an in vivo model to evaluate the influence of topical formulations on irritation. AB - Although skin protective products to prevent irritant skin reactions are in wide use, neither standardized test models to prove differences in efficacy exist, nor has the quality or the reproducibility of results been evaluated in a multicentre approach. This should be mandatory when developing or testing skin care products. Therefore, we have designed a multicentre study in an approach to find a standardized test procedure for the evaluation of skin protective products. In this irritation study, a repeated short-time occlusive irritation test (ROIT) with a standardized protocol has been evaluated in 2 phases (12 days and 5 days protocol) in 4 (n=20) respectively 6 (n=33) skilled centres. The skin reaction was induced by 2 irritants (0.5% aq. SLS and toluene, 2x a day for 30 min). Its modification by 3 different cream bases with different hydrophilicity was analyzed. The irritation was monitored by bioengineering methods (TEWL measurement, colorimetry) and by clinical scoring. The evaluation showed that significant results could already be achieved with the 5-day protocol. Furthermore, in spite of the expected inter-centre variations due to heterogeneity of the individual threshold of irritation, interpretation of clinical score, and inter-instrumental variability, the ranking of the vehicles regarding reduction of the irritant reaction was consistent in all centres. PMID- 10871098 TI - Use of the local lymph node assay for the estimation of relative contact allergenic potency. AB - The effective toxicological evaluation of skin sensitization demands that potential contact allergens are identified and that the likely risks of sensitization among exposed populations assessed. By definition, chemicals which possess the toxicological property of skin sensitization potentially are capable of causing allergic contact dermatitis (ACD) in humans. However, this hazard is not an all-or-none phenomenon; clear dose-response relationships can be discerned and thresholds identified for both the induction of sensitization and the elicitation of contact dermatitis. Commonly, these parameters are grouped under the heading of potency, determination of which is vital for risk assessment. In the present investigation, the local lymph node assay (LLNA) has been employed to determine the relative potency of a range of 20 chemicals. The parameter used is the estimated concentration required to produce a 3-fold increase in draining lymph-node cell proliferative activity, the EC3 value. These measurements have been compared with an assessment of the human sensitizing potency of the 20 selected chemicals, each being assigned to 1 of 5 classes based on their human sensitizing potency. The EC3 value, derived from LLNA work carried out in acetone/ olive oil vehicle, correlated well with the human classification, with the strongest sensitizers having low EC3 values (1000 fatalities in the United States per year. Understanding of this manner of death is likewise increasing, as noted by the growing number of cases reported in the literature. However, this form of accidental death is much less frequently seen in females (male:female ratio >50:1), and there is correspondingly less literature on female victims of autoerotic asphyxiation. The authors present the case of a 31-year-old woman who died of an autoerotic ligature strangulation and review the current literature on the subject. The forensic examiner must be able to discern this syndrome from similar forms of accidental and suicidal death, and from homicidal hanging/strangulation. PMID- 10871124 TI - Stereolithography: a potential new tool in forensic medicine. AB - Stereolithography is a computer-mediated method that can be used to quickly create anatomically correct three-dimensional epoxy and acrylic resin models from various types of medical data. Multiple imaging modalities can be exploited, including computed tomography and magnetic resonance imaging. The technology was first developed and used in 1986 to overcome limitations in previous computer aided manufacturing/milling techniques. Stereolithography is presently used to accurately reproduce both the external and internal anatomy of body structures. Current medical uses of stereolithography include preoperative planning of orthopedic and maxillofacial surgeries, the fabrication of custom prosthetic devices; and the assessment of the degree of bony and soft-tissue injury caused by trauma. We propose that there is a useful, as yet untapped, potential for this technology in forensic medicine. PMID- 10871125 TI - Forensic aspects of ocular injury. AB - A case of homicidal stabbing resulting in bilateral penetrating ocular injuries is described. The case is noteworthy in that it highlights an unusual mechanism of death in homicidal stabbing. Disturbances in heart rhythm including asystole can be ascribed to the so-called oculocardiac or trigeminocardiac reflex. Although this phenomenon is well known to ophthalmologists, neurosurgeons, and anesthetists, it is much less familiar to forensic pathologists. This is a potential mechanism of death worthy of consideration in cases of sudden unexpected death occurring in the context of facial injury. PMID- 10871126 TI - A baby, a virus, and a rat. AB - The authors present a case initially thought to be a child abuse homicide that, after complete autopsy and thorough investigation, was determined to be caused by a viral infection and complicated by postmortem animal activity. Neonatal herpes simplex infection and postmortem skin defects are discussed. PMID- 10871127 TI - Spontaneous rupture of the liver associated with a primary angiosarcoma: case report. AB - A case of fatal spontaneous rupture of the liver caused by primary angiosarcoma is described. Spontaneous rupture of the liver is a rare clinical and pathological entity associated with high morbidity and mortality rates. Possible causes include infections such as hydatid disease, infiltrating conditions such as amyloidosis, inflammatory disorders such as the vasculitides, malignant and benign tumors, and tumorlike conditions. As in the case presented, the finding of a ruptured liver raises the possibility of blunt abdominal trauma, and the circumstances and scene should be assessed. The possibility of terminal resuscitation attempts should also be considered as a possible cause. PMID- 10871128 TI - Identification of remains by sequencing of mitochondrial DNA control region. AB - The maternity of two newborns who were murdered and abandoned >5 and 10 years were analyzed by amplification and direct sequencing of mitochondrial DNA (mtDNA) control regions. Sequences of two hypervariable segments from each femur bone sample and the blood of the putative mother showed four mutations in hypervariable region I and two mutations in addition to two nucleotide insertions in hypervariable region II compared with the reference sequence, and all sequences were identical. The genotype of these individuals is found to be relatively rare in the Japanese population, and it was strongly suggested that both sets of newborn remains really were children of the putative mother. Sexes of the remains were determined to be female and male by amplifying a segment of the X-Y homologous gene, amelogenin. These results demonstrate that sequencing of mtDNA is a useful tool for genetic identification of aged and decomposed materials. PMID- 10871129 TI - An unexpected death during oxygen-ozone therapy. AB - An unexpected death is described that was caused by gas embolism that occurred during oxygen-ozone (O2/O3) therapy administered by autohemotransfusion for psoriasis. This unusual complication suggests the necessity of investigating benefits and adverse effects of medical ozone application. PMID- 10871130 TI - Asbestos body burden in decomposed human lungs. AB - The authors discuss the influence of postmortem tissue decomposition on the lung asbestos body (AB) burden, with the aim of evaluating the reliability of data obtained from autopsies performed for medicolegal purposes several months after deaths in possible connection with asbestos-related pathology. Eight autopsy cases were selected, each one with occupational exposure considered very probable on the basis of the history or pathologic findings. In each case the AB concentrations were assessed soon after death in one lung and after periods of 1 to 18 months in the others, which had been stored in sealed containers without fixation. AB concentrations consistently decreased with time in rotten lungs. The counts in some cases became negative a few months after death, even in cases with very high AB counts at first examination. It may be reasonably inferred that, in putrefied lungs from corpses exhumed after months of internment, the counts in digested tissues and the screening of histologic sections for AB may give false negative results. PMID- 10871131 TI - Necrotizing fasciitis: reports of three fatal cases simulating and resulting from assaults. AB - Necrotizing fasciitis is a progressive, potentially fatal, rapid, necrotizing infection of the subcutaneous tissues and fascia often caused by a mixture of organisms or by infection with group A Streptococcus pyogenes with or without Staphylococcus aureus. Three cases are presented that have been encountered in forensic pathologic practice. Two cases presented after assaults, and the third simulated an assault and burglary. The history, scene, and pathologic findings are presented with a brief review of the literature. PMID- 10871132 TI - Unappreciated agenesis of cerebellum in an adult: case report of a 38-year-old man. AB - An unexpected finding at autopsy of almost complete agenesis of the cerebellum in an apparently functional, mentally subnormal 38-year-old man who died as the result of an accidental electrocution is reported. The posterior fossa was normal in appearance despite nearly complete absence of the cerebellum. A number of syndromes of cerebellar atrophy or dysgenesis have been reported, but congenital agenesis is considered a very rare condition. It does not resemble most common cerebellar malformations or acquired conditions, especially in an adult, who apparently had reasonable motor and coordinative function. The relevant literature is reviewed. PMID- 10871133 TI - Identification of an alleged offender of murder by VNTR analysis: case report. AB - DNA typing was used to demonstrate that three human body pieces found disposed of in the countryside around Athens and the suspicions about the perpetrator's identity were connected. Reverse paternity testing was attempted by comparative typing of three variable number tandem repeats loci in the remains, as well as in the presumptive parents and sister of the decedent, demonstrating Mendelian inheritance of the alleles of the loci analyzed. Confronting the results of the abovementioned analysis, the suspect accepted the accusation. PMID- 10871134 TI - Correlation of the incidence of cocaine and cocaethylene in hair and postmortem biologic samples. AB - Hair samples are useful as a matrix for drug testing because drugs can be detected in hair for longer periods than in blood or urine. The authors report a prospective comparison of the detection of cocaine and cocaethylene in routine postmortem biologic specimens to the detection of cocaine and cocaethylene in hair. The authors collected hair samples from various areas of the head in 53 autopsy cases, prepared them, and analyzed them by gas chromatography/mass spectrometry (GC/MS) for cocaine and cocaethylene. The authors compared the results of hair analysis with the results of toxicologic analysis performed on routine postmortem samples by enzyme multiplied immunoassay technique and GC/MS. Cocaine was found in either biologic fluids or in hair in 16 of 53 samples tested. Nine samples were positive for cocaine in both biologic fluids and hair. Five samples contained cocaine only in biologic fluids, and two contained cocaine only in hair. Cocaethylene was present in two cases. Drug screening of hair provides additional information in some autopsy cases, but the authors have not made hair analysis a routine practice. It may prove useful to save hair samples in all cases for later analysis if warranted by additional history or autopsy findings. PMID- 10871135 TI - Analysis of eight polymorphic human genetic markers in a well-defined Greek population. AB - The use of DNA in forensic science has become a basic tool for person identification or parentage testing. The use of polymorphic markers permits the formation of a unique profile for each individual. The knowledge of allele frequencies in a given population allows the scientist to estimate the probability of a particular allele combination. For this task, allele databanks are essential. In this report, the authors estimate the frequencies of eight polymorphic markers (namely, HumFES, HumF13A1, HumTHO1, HumVWA, HumFABP2, HumLIPOL, D1S80, and D17S5) in a randomly selected sample from Crete, Greece. The allele profile of all markers, with the exception of D17S5 and HumFABP2, concurs with previous reports and international data. PMID- 10871136 TI - Dental identification using digital images via computer network. AB - Dental identification is a useful scientific method. In Japan, however, there are only a few forensic odontologists; moreover, until now, forensic dental services have only been offered by general dentists. These dentists may not be able to offer such forensic services during office time. For a quick comparison, the authors tried sending digital photos, taken with a 2-million-pixel digital camera, to dental offices via the Internet. If a dental office has Internet access, it is possible for dental charting to be sent directly to the autopsy room. Of course, digital images only provide the first outline. However, when antemortem dental records of the person in question are available at autopsy, a quick comparison can be made. PMID- 10871137 TI - A study on polymerase chain reaction-based HLA DQ alpha locus in Elazig, Turkey. AB - The polymerase chain reaction (PCR) was used for genetic characterization of 45 samples taken from the city of Elazig in Turkey. The polymorphism at the human leukocyte antigen DQalpha locus was detected. Allele and genotype frequencies were determined for unrelated individuals at this locus. Laboratory analyses were done by PCR amplification of DNA. Hybridization to allele specific oligonucleotide probes was performed using a reversed dot-blot typing method. The collected genotype and allele frequencies have been tested, and a comparison was made with other population surveys of this locus. Allele frequencies ranged from 3.3% (allele 1.3) to 36.7% (allele 4), with a discrimination power of 0.92. No deviation was seen from Hardy-Weinberg equilibrium in the findings. PMID- 10871138 TI - Child deaths in Virginia, 1996: a review of investigations of sudden, unexpected, or unnatural deaths of children less than age 13. AB - This study examines the consistency of investigative procedures used by the Office of the Chief Medical Examiner, law enforcement, and child protective services, when investigating the violent, sudden, unexpected, or unnatural deaths of children. The study also assessed the status of communication and cooperation among the investigating agencies, to determine whether improvements in the level of cooperation and communication among the systems recommended by prior legislative studies had been achieved. The subjects of this study were children from birth through age 12 who died a sudden, unexpected, or unnatural death in Virginia in 1996. The findings from this research provide both justification to celebrate the progress that has been made and the stimulus to improve the investigation into the sudden, unexpected, or unnatural deaths of children in Virginia. Data suggested that the level of cooperation and communication among child protective services workers, medical examiners, and law enforcement personnel in Virginia had increased between 1986 and 1996. The results demonstrated that some investigative procedures were consistent, especially within regional boundaries. However, the results also showed that inconsistencies exist in the way some deaths are investigated, and that room for improvement exists. PMID- 10871139 TI - Custody restraint asphyxia. PMID- 10871140 TI - Resetting of baroreflexes, changes in autonomic controls and sudden unexpected death during sleep. PMID- 10871141 TI - Shaken baby syndrome. PMID- 10871142 TI - Effect of knee and hip position on hip extension range of motion in individuals with and without low back pain. AB - STUDY DESIGN: A 2-group, nonrandomized, mixed design with 1 between-subjects factor (group) and 2 within-subjects factors (knee and hip position). OBJECTIVES: To determine the amount of passive hip extension during changes in the knee angle in the sagittal plane, and the hip angle in the frontal plane in back-healthy (BH) subjects and subjects with low back pain (LBP). BACKGROUND: Information regarding the specific contributions of hip flexor muscles to limitations in hip extension range of motion (ROM) is necessary for the prescription of appropriate treatment. METHODS AND MEASURES: Thirty-five BH subjects (24 women and 11 men, mean age = 31.37 +/- 11.36) and 10 subjects with LBP (6 women and 4 men, mean age = 33.70 +/- 9.31) participated in the study. The passive length of the one- and two-joint hip flexor muscles was tested in 4 different conditions in which the positions of the knee and the hip were varied. The knee was positioned passively in full extension or 80 degrees of flexion while the hip was positioned passively in zero abduction or full abduction. RESULTS: Subjects with LBP displayed less passive hip extension than BH subjects (LBP, -5.61 degrees +/- 4.30; BH, -2.57 degrees +/- 4.18). Both groups had less hip extension when the knee was in flexion of 80 degrees than when the knee was fully extended (flexed, -5.51 +/- 4.50; extended, -0.98 degrees +/- 4.65), and when the hip was in zero hip abduction than when the hip was fully abducted (zero, -7.55 degrees +/- 5.03; full, 1.06 degrees +/- 4.31). The contribution of the different hip flexors to a hip extension limitation differed between BH and subjects with LBP. BH subjects demonstrated an effect of knee angle on hip extension when the hip was in zero abduction (flexed, -11.43 degrees +/- 5.81; extended, -2.49 degrees +/- 5.39), but not when the hip was in full abduction (flexed, 1.74 degrees +/- 3.91; extended, 1.89 degrees +/- 3.94). Subjects with LBP demonstrated an effect of knee angle on hip extension when the hip was in zero abduction (flexed, -12.60 degrees +/- 4.91; extended, -6.65 degrees +/- 5.03) and when the hip was in full abduction (flexed, -3.10 degrees +/- 5.53; extended, -0.10 degrees +/- 5.18). CONCLUSIONS: The results of this study provide evidence that changing the knee joint angle in the sagittal plane and the hip joint angle in the frontal plane, during the hip flexor length test, can affect the amount of passive hip extension ROM. The contribution of specific hip flexor muscles to a hip extension limitation may differ depending on the individual's movement dysfunction. Modifying the hip flexor length test, as described, should provide information about the specific muscles contributing to a hip joint extension limitation. PMID- 10871143 TI - Inertial effects on moment development during isokinetic concentric knee extension testing. AB - STUDY DESIGN: Two group pre-test post-test design was used with 2 isokinetic dynamometers in a laboratory setting. OBJECTIVES: To evaluate the inertial effects on moment measurements during the initial acceleration period of concentric isokinetic knee extension. BACKGROUND: Torque acceleration energy is a controversial measure of muscle "explosive power." Moments due to acceleration during the initial period of isokinetic movements have been ignored in the majority of isokinetic studies. METHODS AND MEASURES: These inertial effects were assessed at various angular velocities measuring the work production at the initial 0.125 second on a Biodex and a Lido dynamometer. Four women (age, 23.33 +/- 2.49 years) and 5 men (age, 26.00 +/- 2.63 years) were tested on Biodex. A different group of 9 men (age, 23.4 +/- 3.41 years, height, 1.77 +/- 1.02 meters, and mass, 74 +/- 8.26 kg) was tested on the Lido. Joint moment was calculated by including moments associated with the angular acceleration of the lower limb and the lever arm. RESULTS: Torque acceleration energy was significantly greater after correction for inertial effects compared with uncorrected measures at moderate (165 and 180 deg/s) and high velocities (300 deg/s) on both dynamometers. CONCLUSION: In cases where moment development in the initial 0.125 second of an isokinetic movement is measured, it would be better to use slow velocities where inertial effects are minimum. At high angular velocities inertial correction is essential to acquire valid moment measurements and conclusions about muscle and joint function. PMID- 10871144 TI - Gilmore's groin repair in athletes. AB - Incapacitating groin pain is a frequent problem among athletes and its etiology may be multifactorial. A specific clinical syndrome relating to injury to the lower abdominal wall has been described and successfully treated by O. J. Gilmore. This paper presents our results of 100 consecutive groin repairs in 85 young athletes using a diagnostic and therapeutic strategy similar to Gilmore's. Ninety-six percent of our patients returned to competitive sport within 15 weeks; we suggest that this is an appropriate therapeutic intervention for athletes who develop chronic incapacitating groin pain. PMID- 10871145 TI - Effect of antipronation tape and temporary orthotic on vertical navicular height before and after exercise. AB - STUDY DESIGN: A randomized controlled, crossover, within-subjects study evaluating 2 antipronation treatments. OBJECTIVES: To investigate the antipronation effect of 2 treatments designed to reduce abnormal pronation, and the effect of an exercise challenge on the treatments. BACKGROUND: Control of abnormal pronation in order to ameliorate inappropriate stresses on injured soft tissues is frequently sought in the treatment of overuse injuries of the lower limb. Tape and temporary soft orthotics are used to control abnormal pronation. The effects of these treatments remain largely untested. METHODS AND MEASURES: Fourteen subjects (age = 23.8 +/- 3.5 years) who had at least a 10-mm navicular drop were studied. The dependent variable was vertical navicular height. The two independent variables were the treatment conditions (temporary felt orthotics, augmented LowDye tape, and control) and the exercise challenge (0, 10, and 20 minutes of controlled jogging). The subjects' vertical navicular height was measured before and after the application of the treatment conditions, and then after 10 and 20 minutes of jogging. RESULTS: Tape and orthotic treatments produced approximately a 19% and 14% increase in vertical navicular height, respectively, which were both significantly greater than the control condition (0%). The treatment effect, although significantly diminished following exercise challenge, remained superior to control (6.5% for orthotic and 3.5% for tape compared to -7.3% for control). CONCLUSION: Antipronation tape and temporary orthotics help to control excessive foot pronation initially after application and following exercise. These treatments may be useful in the assessment and treatment of lower limb injuries that are associated with abnormal foot pronation. PMID- 10871146 TI - Effects of ultrasound and stretch on knee ligament extensibility. AB - STUDY DESIGN: Randomized, double-blind pre-test and post-test with repeated measures. OBJECTIVE: To determine whether heating with continuous wattage ultrasound (CWUS) augmented the effects of stretching on ligament extensibility in nonimpaired human subjects. BACKGROUND: Heating with CWUS, combined with static stretching, is often used to treat ligament "tightness," but "heat and stretch" has not been studied well in vivo. METHODS AND MEASURES: Twenty-one nonimpaired women subjects (aged 31.5 +/- 11.0 years) underwent serial measurements of knee joint displacement (valgus and varus) on a Genucom arthrometer before and after valgus stretch (10 ft-lb x 2.5 minutes). Subjects received either simultaneous CWUS (3 MHz, 1.25 W/cm2) or sham CWUS applied over the medial collateral ligament. Five trials (2 before, 3 after treatment) were conducted with the right knee positioned in 20 degrees of flexion. Subjects received the alternate treatment 28 days later. For each subject, all testing was performed by the same investigator. RESULTS: Repeated measures ANOVA revealed that stretching, combined with sham CWUS, increased mean valgus displacement from 8.95 degrees (+/-1.72 degrees) to 10.00 degrees (+/-2.10 degrees). Stretching, combined with CWUS, increased mean valgus displacement from 9.24 degrees (+/-2.36 degrees) to 10.48 degrees (+/-2.54 degrees). This was a 13.4% change from the control condition. CONCLUSION: Heating with CWUS did not augment the effects of stretching. "Heat and stretch" with CWUS may not be more effective than stretching alone for increasing the extensibility of dense connective tissue. PMID- 10871147 TI - Varicella-Zoster virus: pathogenesis, immunity, and clinical management in hematopoietic cell transplant recipients. AB - New information about the mechanisms of varicella-zoster virus (VZV) pathogenesis and the host response to the virus has improved our understanding of the threat that VZV reactivation may pose after hematopoietic cell transplantation (HCT). Antiviral therapy compensates for some of the deficiencies in VZV immunity in HCT recipients, and inactivated varicella vaccine may be useful for the early reconstitution of adaptive immunity to VZV after HCT. PMID- 10871148 TI - T-cell subsets mediate graft-versus-myeloid leukemia responses via different cytotoxic mechanisms. AB - Analysis of the cytotoxic effector mechanisms by which T-cell subsets mediate graft-versus-leukemia (GVL) activity is complicated by systems that use unfractionated T cells and leukemias that express alloantigens in addition to tumor-specific antigens. In this study, we used MMB1.10, a myeloid leukemia of C57Bl/6 (B6) origin, to examine the cytolytic pathways employed by syngeneic GVL mediating, and therefore tumor antigen-specific, T-cell subsets. Wright-Giemsa staining and flow cytometric analysis indicated that MMB1.10 cells exhibited the morphology and markers most consistent with a monocytic-myeloid origin. Although reverse transcription-polymerase chain reaction analysis revealed that MMB1.10 cells expressed tumor necrosis factor (TNF) receptor types I and H, in vitro assays suggested that these cells were resistant to TNF-alpha-mediated cytotoxicity. For study of in vivo GVL responses, mice were challenged with MMBl.10 cells, lethally irradiated, and administered anti-Thy-1-treated (T-cell depleted) bone marrow (ATBM) either alone or in combination with T-cell subsets from MMB1.10-presensitized mice. In regard to CD4+ donor T cells, 4 x 10(6) MMB1.10-presensitized wild-type (wt) cells exhibited increased GVL responses and survival values relative to tumor-challenged recipients of ATBM only. CD4 T cells from either perforin-deficient (pfp0) or Fas ligand (FasL)-deficient (gld) mice exhibited a lower level of GVL activity but did not produce any long-term survivors. Recipients of 5 x 10(6) wt B6 CD8+ T cells had significantly improved survival relative to tumor-challenged mice that received ATBM only. The same dose of gld CD8+ T cells exhibited a reduced but significant level of GVL activity, whereas cells from mice that were perforin-deficient or cytotoxicity doubly deficient (cdd) (ie, lacking perforin and FasL) exhibited no discernable GVL activity. Doubling the gld CD8+ T-cell dose to 10(7) cells resulted in further improved survival of recipients. We conclude that GVL effects mediated by CD4+ T cells can depend on either perforin- or FasL-mediated mechanisms, whereas the CD8+ T-cell subset is heavily dependent on perforin-mediated cytotoxicity. PMID- 10871149 TI - Real-Time polymerase chain reaction of immunoglobulin rearrangements for quantitative evaluation of minimal residual disease in multiple myeloma. AB - The majority of patients with multiple myeloma (MM) have persistence of minimal residual disease (MRD), as determined by polymerase chain reaction (PCR) detection of clonal immunoglobulin H (IgH) gene rearrangements. As a result, PCR analysis has not provided clinically useful prognostic information in myeloma patients. Instead, quantitative PCR approaches are required to predict patient outcomes and assess response to novel treatment strategies. We adapted real-time PCR technology to quantify myeloma cells using the IgH rearrangement and then assessed the utility of this approach in 29 patients with myeloma who had undergone autologous stem cell transplantation. Because of the high cost of producing a specific reporting probe for each patient, H-chain V-region family specific consensus probes were used in association with allele-specific oligonucleotides for PCR amplification. Because of the high frequency with which somatic hypermutation at the immunoglobulin locus occurs in MM, a number of mismatches occurred between the patient sequences and the consensus probe. However, construction of a limited number of probes allowed real-time PCR with a sensitivity of 10(-4) to 10(-5). To validate this method, we extensively evaluated assay accuracy and reproducibility. Results indicate that real-time PCR using consensus probes provides a feasible, accurate, and reproducible method for evaluating MRD in M M and possibly in other differentiated B-cell malignancies, and one that is less expensive than the use of patient-specific probes. This technique is being used to assess tumor depletion after immunologic purging and changes in tumor burden in patients undergoing stem cell transplantation and novel treatment approaches. PMID- 10871150 TI - Equivalence of 2 effective graft-versus-host disease prophylaxis regimens: results of a prospective double-blind randomized trial. AB - We have previously demonstrated a decrease in the incidence of acute graft-versus host disease (GVHD) with the addition of methotrexate (MTX) to cyclosporine (CSP) and prednisone (PSE) chemotherapy in patients with leukemia. We have now completed a prospective randomized trial comparing the 3-drug regimen (CSP/MTX/PSE, including 3 doses of MTX) to the standard 2-drug regimen (CSP/MTX, including 4 doses of MTX) to investigate the benefit of PSE used up front for the prevention of acute and chronic GVHD. In the trial, 193 patients were randomized and 186 were included in the final analysis. All patients received a bone marrow graft from a fully histocompatible sibling donor. The preparatory regimen consisted of fractionated total-body irradiation (fTBI) and etoposide in all but 13 patients, who received fTBI and cyclophosphamide. The patients were randomized to receive either CSP/MTX/PSE or CSP/MTX. The 2 groups were well balanced with respect to diagnosis, disease stage, age, donor-recipient sex, and parity. In an intent-to-treat analysis, the incidence of acute GVHD was 18% (95% confidence interval [CI] 12-28) for the CSP/MTX/PSE group compared with 20% (CI 10-26) for the CSP/,MTX group (P = .60), with a median follow up of 2.2 years. Overall survival was 65% for those receiving CSP/MTX/PSE and 72% for those receiving CSP/MTX (P = .10); the relapse rate was 15% for the CSP/MTX/PSE group and 12% for the CSP/MTX group (P = .83). The incidence of chronic GVHD was similar (46% versus 52%; P = .38), with a follow-up of 0.7 to 6.0 years. Of interest, 21 patients went off study due to GVHD (5 in the CSP/MTX/PSE group and 16 in the CSP/MITX group [P = .02]), and 11 patients went off study because of alveolar hemorrhage (3 in the CSP/MTX/PSE group and 8 in the CSP/MTX group [P = .22]). The addition of PSE did not result in a higher incidence of infectious complications, bacterial (66% versus 58%), viral (77% versus 66%), or fungal (20% versus 20%), in those receiving CSP/MTX/PSE versus CSP/MTX, respectively. These data suggest that the addition of PSE was associated with a somewhat lower incidence of early posttransplantation complications but did not have a positive impact on the incidence of acute or chronic GVHD or event-free or overall survival. PMID- 10871151 TI - Transplantation of highly purified CD34+Thy-1+ hematopoietic stem cells in patients with metastatic breast cancer. AB - We report here the transplantation of extensively purified "mobilized" peripheral blood CD34Thy-1 hematopoietic stem cells from 22 patients with recurrent or metastatic breast cancer. Patients were mobilized with either high-dose granulocyte colony-stimulating factor (G-CSF) alone or cyclophosphamide plus G CSE Median purity of the stem cell product at cryopreservation was 95.3% (range, 91.1%-98.3%), and viability was 98.6% (range, 96.5%-100%). After high-dose chemotherapy with carmustine, cisplatin, and cyclophosphamide, CD34+Thy-1 cells at a median dose of 11.3 x 10(5) per kilogram (range, 4.7-163 x 10(5) per kilogram) were infused. No infusion-related toxicity was observed. Neutrophil recovery was prompt, with median absolute neutrophil count >500/microL by day 10 (range, 8-15 days) and >1000/microL by day 11 (range, 8-17 days). Median platelet recovery (>20,000/microL) was observed by day 14 (range, 9-42 days) and >50,000/microL by day 17 (range, 11-49 days). Tumor cell depletion below the limits of detection of a sensitive immunofluorescence-based assay was accomplished in all patients who had detectable tumor cells in apheresis products before processing. Although CD4+ T-cell reconstitution was slow, no unusual infections were observed. Neither early nor late graft failure was observed, and no patient required infusion of unmanipulated backup cells. At a median follow-up of approximately 1.4 years and a maximum follow-up of 2.5 years, 16 of the 22 patients remain alive, with 9 free of disease progression, and have stable blood counts. In summary, highly purified CD34+Thy-1+ cells used as the sole source of the hematopoietic graft result in rapid and sustained hematopoietic engraftment. PMID- 10871152 TI - Second allogeneic transplantation after failure of first autologous transplantation. AB - We evaluated the outcome of second allogeneic bone marrow transplantations (BMTs) in 59 patients aged 1-57 years who relapsed after initial autologous transplantation. Patients received a second transplantation for recurrent acute myeloid leukemia (AML) (n = 24), acute lymphoblastic leukemia (ALL) (n = 13), lymphoma (n = 18), multiple myeloma (n = 3), or chronic myelogenous leukemia (n = 1) from an HLA-matched related (n = 14), mismatched related (n = 25), or matched unrelated (n = 20) donor. The probabilities of nonrelapse mortality, relapse, and disease-free survival (DFS) 2 years after the second BMT were 51%, 26%, and 23%, respectively. The 2-year DFS estimates for AML, ALL, and lymphoma were 46%, 23%, and 0%. Univariate analysis demonstrated that superior DFS was associated with age < or =17 years at the time of the second transplantation, remission before the second transplantation, total-body irradiation-based preparative regimen for the second transplantation, and the diagnosis of AML. These data demonstrate that an allogeneic transplantation after a failed autologous transplantation can result in disease-free survivors, especially in the young. The outcomes after a second transplantation for patients aged >17 years and for those with lymphoma were especially grim. These data suggest that pediatric patients may be appropriate candidates for a second transplantation. In adults, however, the use of an allogeneic transplantation as salvage therapy after failure of the initial autologous transplantation is generally unsuccessful. Alternative experimental strategies, such as low-dose nonmyeloablative allogeneic minitransplantations, should be considered. PMID- 10871153 TI - CMV antigenemia following bone marrow transplantation: risk factors and outcomes. AB - Cytomegalovirus (CMV) infection remains a major problem in blood and bone marrow transplant (BMT) recipients. Recent efforts have been directed at prevention, early diagnosis, and treatment of CMV disease following BMT. Assay for CMV early antigen pp65 on circulating leukocytes has been shown to be sensitive, and specific in detecting early CMV infection. We examined the frequency, risk factors, and outcomes of a positive CMV antigen assay in 118 consecutive BMT patients. Forty-three (36%) of the 118 patients developed CMV antigenemia a median of 26 days post-BMT (range, -6 to 209 days). The incidence of antigenemia in autologous, related donor, and unrelated donor BMT recipients was 15%, 50%, and 48%, respectively (P < .01) and was lower in CMV-seronegative patients (19% versus 51% in seropositive patients; P < .01). Patients with grade II to IV acute graft-versus-host disease (GVHD) had 2.2 times the risk of antigenemia of patients with no or only limited GVHD (P = .03). Age at transplantation, underlying disease, CMV prophylaxis regimen, and GVHD prophylaxis regimen did not affect the risk of CMV antigenemia. Ten of the 43 antigenemic patients, all CMV seropositive allogeneic BMT (alloBMT) recipients, developed CMV organ disease a median of 101 days (range, 28-283 daya) post-BMT. These data suggest that CMV seropositive alloBMT patients are at highest risk for CMV antigenemia and for organ disease as well. CMV disease may occur before antigenemia is detectable in leukopenic patients and may also develop late post-BMT, even in patients still receiving antiviral prophylaxis. In high-risk groups, intensive surveillance continuing for more than 6 months after BMT may be indicated. PMID- 10871154 TI - Comparative distribution of glutamyl and aspartyl aminopeptidase activities in mouse organs. AB - To evaluate the functional role of glutamyl and aspartyl aminopeptidases, their soluble and membrane-bound activities were measured simultaneously in several tissues of normal mice using arylamide derivatives as substrates. Although the soluble aspartyl aminopeptidase activity showed its highest levels in the testicle, the rest of the activities presented their highest levels in the kidney. Different patterns of distribution were observed for glutamyl and aspartyl aminopeptidase activities and also for soluble and membrane-bound aspartyl aminopeptidase activities. However no major differences were observed between soluble and membrane-bound glutamyl aminopeptidase activities. This unequal distribution suggests that the use of arylamide derivatives as substrates is a sensitive method that distinguishes between these enzymatic activities. The results also suggest different functions for soluble and membrane-bound aspartyl aminopeptidase activities, and for glutamyl and aspartyl aminopeptidase activities. PMID- 10871155 TI - Increased expression of interleukin 6 in term compared to the first trimester human placental villi. AB - Cytokines and their specific receptors expressed at the feto-maternal interface are known to play a critical role in regulating various placental functions. Interleukin 6 (IL-6) has been shown to be produced by both decidua and the trophoblast cells of the placenta. The aim of the present study was to examine the expression profile of placental IL-6 protein and mRNA at early and late stages of gestation. Placental villi were obtained from women undergoing first trimester pregnancy termination or elective Cesarean section at term. Functionally active placental explant culture system was used to study the release of IL-6 by these tissues. IL-6 was detected in placental conditioned media of all the samples from first trimester and term group. The mean levels of IL-6 produced by term villi were found to be 5.5, 7.5 and 5-fold higher at term when compared with the first trimester at 24 h, 48 h and 72 h of culture, respectively. Expression of IL-6 mRNA was demonstrated by RT-PCR performed on total RNA isolated from these tissues. IL-6 mRNA expression was detected in both early and late gestational placental tissues. Moreover, the level of IL-6 mRNA was found to be approximately 4-fold higher at term compared with first trimester. These data are consistent with the hypothesis that levels of IL-6 production by the placenta are developmental stage-specific and suggest that expression of IL-6 in the placenta could be subjected to transcriptional regulation. PMID- 10871156 TI - Complete sequencing and mRNA expression analysis of the MEN-I gene in adrenal myelolipoma. AB - The molecular pathogenesis of adrenal myelolipoma is unclear. Endocrine activity of these tumors and association with other endocrine tumors have stimulated the hypothesis that it may belong to the group of sporadic tumors caused by defects of the gene responsible for multiple endocrine neoplasia type I (MEN-I). DNA of blood and tumoral sections from two patients with adrenal myelolipoma were analyzed by examination of variable number of tandem repeats (VNTR) loci PYGM, D11S987, D11S480, and D11S449 on chromosome 11q13 and by complete direct DNA sequencing of all coding exons and splice junctions of the MEN-I gene. Menin expression was examined by RT-PCR. RT-PCR did not detect menin expression in one adrenal myelolipoma. No loss of heterozygozity on chromosome 11q13 was identified. Intragenic heterozygozity was retained in codon 418 of the menin gene in both patients. No mutation was identified in the coding exons of the menin gene. Complete DNA sequencing yielded no hint that defects of the MEN-I gene are responsible for the formation of adrenal myelolipomas. Adrenal myelolipomas do not share the loss of heterozygozity on chromosome 11q13 observed in some benign adenomatous and many malignant adrenocortical tumors. PMID- 10871157 TI - Excess of glucocorticoids impairs whole-body antioxidant status in young rats. relation to the effect of dexamethasone in soleus muscle and spleen. AB - The action of glucocorticoids in high doses is catabolic, but not much is known about the accompanying effects on antioxidative capacity of the entire body. Animals were treated (or not) with dexamethasone (Dex) 2 mg/kg b.w. d-1 during 5 consecutive days followed by recovery, during which an additional group received 3-hydroxy-3-methylbutyrate (40 mg/kg b.w.). Animals were killed after treatment with Dex, and after 5 days of the recovery period. Dexamethasone treatment decreased appetite almost twofold (from 20 g/day to 10 g/day, P < 0.001). Feed restriction, however, seemed to have only minor impact on the effects observed since body weight loss of pair-fed rats after the 5th day of treatment was only 2% and Dex-treated rats decrease in body weight was 22% (P < 0.05). In turn, wet weight of the soleus muscle (expressed per body weight) did not significantly decrease after Dex treatment, suggesting relative resistance of oxidative type muscles to the catabolic action of dexamethasone. Spleen wet weight expressed per body weight dropped by 65% (P<0.001). Additionally, there was a 46% reduction (P<0.001) of blood glutathione (GSH/Hb), and 36% (P < 0.001) of muscle glutathione (GSH/tissue wet weight). This suggests that dexamethasone directly and/or indirectly impaired antioxidant reactions. This was further confirmed by a significant (49%) decline of SOD-1 activity in erythrocytes isolated from the group treated with dexamethasone. Another index of lipid peroxidation (TBARS) was also significantly increased. Activity of blood plasma CK increased by 73% (P<0.001) in Dex-treated rats, indicating moderate injury of muscle tissue. In conclusion, young growing rats were sensitive to the dosage of dexamethasone, but in contrast to lymphoid tissue, could easily compensate the outcomes of impaired antioxidative defence within 5 days of recovery. PMID- 10871158 TI - Tamoxifen prevents bone loss in castrated male mice. AB - The selective estrogen receptor modulator tamoxifen was administered to intact and castrated male mice, and its effects on tibial bones and circulatory calcium, phosphate and testosterone were compared with controls and castrated animals. Tamoxifen in a dose used in humans for treatment of breast cancer decreased the weight of seminal vesicles, an organ which is highly sensitive to the androgenic effect, decreased the concentration of testosterone, but did not have any negative effect on bone density or mineral content in intact mice. When castrated mice with extraordinarily low concentrations of testosterone and weights of seminal vesicles were treated with tamoxifen, the changes in bone density and bone mineral resulting from castration were not only entirely prevented, but increased above the values of intact mice. At the same time, cortical bone was lost in orchidectomized mice, and this decrease in cortical thickness of femur was completely prevented by tamoxifen treatment. Pharmacological therapy with estrogen agonist on bone, tamoxifen in androgen deficient adult male mice prevents bone loss. PMID- 10871159 TI - Clinical parameters (body mass index and age) are the best predictors for the need of insulin therapy during the first 18 months of diabetes mellitus in young adult patients. AB - To address the question whether there are simple clinical predictors of need for insulin in the first 18 months of treatment of diabetes presenting in young adult subjects, a prospective study of 24 patients with diabetes mellitus (age: 18-40 years) was designed. At diagnosis of diabetes, age, sex, body mass index (BMI), glycemia, ketonuria, C-peptide, insulin autoantibodies, islet cell antibodies and glutamic acid decarboxylase antibodies were recorded before starting any treatment. At the end of the follow-up (18 +/- 4 months), they were divided into two groups according to their need for insulin therapy: group 1 (n=15; 62%), who needed insulin therapy, and group 2 (n=9; 38%), who did not. Each marker was related to actual need for therapy necessity. Multivariate analysis showed that BMI and age were the variables with greatest predictive value regarding need for insulin. These data reveal that the need for insulin therapy in young adult diabetic patients may be supported by the clinical criteria of age and BMI, which are both easily and quickly determined. PMID- 10871160 TI - Goiter and impairment of thyroid function in acromegalic patients: basal evaluation and follow-up. AB - AIMS: We evaluated morphological, biochemical and cytological thyroid parameters in acromegalic patients, investigated before and after treatment for acromegaly. PATIENTS: 28 acromegalics were investigated before and, in 18 cases, after 2-7 years of therapy. Fourteen patients were from areas of moderate iodine deficiency in Southern Italy. One patient underwent thyroidectomy before entering this study. RESULTS: 19 patients were euthyroid (FT4: 17.7 +/- 0.8 pmol/l and FT3 4.6 +/- 0.2 pmol/l), but TSH was undetectable in 5/19. Among them, TRH-stimulated TSH increase was absent/impaired or exaggerated/delayed in 9 and one cases, respectively. Decreased FT3 and/or FT4 values with low/normal TSH values were detected in 7 cases; TRH-stimulated TSH response was absent/impaired in 2 patients and exaggerated/delayed in another two. Increased free T4 and free T3 concentrations with undetectable TSH levels were found in one. Two euthyroid patients had high TPOAb levels. Goiter was diagnosed in 21 cases and nodules were found in 14/21. 99Tc scintiscan showed "cold" areas in 13/14 cases and a "hot" nodule in the hyperthyroid patient. Acromegalics from iodine deficient areas showed a not significant increase of prevalence of goiter (86 vs. 71 %) and of mean thyroid volume (35 +/- 7 vs. 28 +/- 4 ml, NS), compared to others. Thyroid volume (TV) did not correlate with GH, IGF-1 and TSH levels, the area under the curve of insulin-increase during OGTT, the age of patients or the duration of acromegaly. Fine needle aspiration biopsy (FNAB), performed in 11/14 patients with nodular goiter, showed colloid nodules in 8 cases, hyperplastic nodules in 2 and an adenomatous nodule in one. Neurosurgery, radiotherapy or medical treatment for acromegaly induced a significant decrease of mean GH and IGF-1 levels (21.5 +/- 8.5 vs. 12.9 +/- 9.6 ng/ml, p< 0.005 and 747 +/- 94 vs. 503 +/- 88 ng/ml, p < 0.02, respectively), but both GH and IGF-1 values normalized only in 3 cases. No significant variation of mean TSH levels was found. Although TV normalized in 3 patients, ultrasound evaluation showed a not significant decrease of mean TV and no changes in the diameter and number of nodules. FNAB was unchanged. CONCLUSIONS: Our results suggest that, despite no correlation between serum GH and IGF-1 levels and thyroid volume being found, a decrease in serum GH and IGF-1 levels has favourable effects on thyroid status. PMID- 10871161 TI - Systemic levels contribute significantly to increased intraocular IGF-I, IGF-II and IGF-BP3 [correction of IFG-BP3] in proliferative diabetic retinopathy. AB - Increased intraocular levels of angiogenic growth factors such as insulin-like growth factor I (IGF-I) have been demonstrated in proliferative diabetic retinopathy (PDR). It is unclear whether increased leakage of the blood retina barrier or local synthesis primarily determine intraocular levels of IGFs in man, which is of special interest regarding possible therapeutic options with somatostatin analogues in PDR. This is the first study investigating parallelly serum and vitreous levels of IGF-I/II, IGF-BP3 and the liver-derived permeability marker albumin to determine in vivo the amount of circulation-derived intraocular IGFs. A control group without retinal proliferation and patients with PDR were compared. Levels of IGF-I/II, IGF-BP3 and albumin were determined by immunological methods. Vitreous levels of albumin were 2.2-fold elevated in patients with PDR (254.1 +/- 37.2mg/dl; n = 27; p = 0.0027) compared to controls (115.7 +/- 36.2mg/dl; n =10), whereas serum levels were slightly decreased in diabetes patients (5049 +/- 196 mg/dl vs. 4330 +/- 186 mg/dl; p = 0.0283). This was comparable to an increase of IGF-I/11 and IGF-BP3 in vitreous from PDR patients (IGF-I: 2.3 +/- 1.1 ng/ml p = 0.005. IGF-II: 37.9 +/- 4.9 ng/ml; p = 0.0003. IGF-BP3: 97.9 +/- 26.9 ng/ml; p = 0.0001; n = 34) compared to controls (IGF-I: 0.7 +/- 0.1 ng/ml. IGF-II: 21.3 +/- 4.2 ng/ml. IGF-BP3: 31.3 +/- 4.9 ng/ml: n = 19). Serum levels did not differ significantly among the groups regarding IGF-I, II and IGF-BP3. Intraocular albumin and IGF-I levels calculated as percentage of the respective serum levels correlated significantly (r = 0.42; p = 0.012). This study demonstrates that influx of IGF-I, II and IGF-BP3 in PDR quantitatively parallels influx of the liver derived serum protein albumin suggesting that leakage of the blood retina barrier and serum levels of IGF primarily determine intravitreal IGF levels rather than local synthesis. Suppression of systemic IGF levels by new, highly effective somatostatin analogues therefore provides a promising approach to prevent PDR. PMID- 10871162 TI - Early reperfusion, late reperfusion, and the open artery hypothesis: an overview. AB - Randomized clinical trials have clearly shown the beneficial effects of early reperfusion within 12 hours, and possibly up to 24 hours, after acute myocardial infarction (AMI). The data on late reperfusion beyond 24 hours are less convincing. Many studies show that an open infarct-related artery after MI, irrespective of the initial reperfusion strategy, is independently associated with improved long-term clinical outcome. However, similar analysis of the large Global Utilization of Streptokinase and tPA for Occluded Arteries (GUSTO) 1 study did not confirm this finding. It is unclear whether mechanical reperfusion of an occluded infarct-related artery late after MI (>24 hours) in asymptomatic patients will confer long-term benefits. The late open artery hypothesis, which proposes several mechanisms by which late reperfusion may offer benefit, remains to be tested in a large clinical trial. This overview focuses on the definitions of early reperfusion, late reperfusion, the relationship between timing of reperfusion and prognosis after AMI, and the late open artery hypothesis. PMID- 10871163 TI - Occluded infarct-related arteries and clinical events. AB - Late patency of the infarct-related artery has been shown to be associated with improved long-term survival rates in observational cohort studies. However, there is a dearth of randomized trials correlating the opening of persistently occluded infarct-related arteries with clinical outcomes. Recent technological advances have improved the success and safety of percutaneous revascularization, resulting in lower restenosis and reocclusion rates. A large randomized trial is needed to evaluate clinical outcomes with percutaneous revascularization versus medical management of occluded infarct-related arteries in the absence of inducible ischemia. PMID- 10871164 TI - The open artery hypothesis: potential mechanisms of action. AB - The postinfarction period is one of intense dynamic activity because the cardiovascular system undergoes a number of adaptive responses attempting to maintain cardiac output. These homeostatic responses contribute to the processes involved in postinfarction ventricular remodeling and involve acute, chronic, systemic, and local reactions. Almost immediately, neurohormones are activated that alter hemodynamic load, and, later, stimulate myocyte hypertrophy, while locally, inflammatory processes clear necrotic debris, reorganize the extracellular matrix, and orchestrate scar formation. Where the initial cardiac insult is sufficiently large (eg, necrosis of more than 40% of left ventricular mass), remodeling responses are exaggerated and may become maladaptive, contributing to the poor long-term prognosis associated with myocardial infarction. Although several studies have shown clear benefits after neurohormonal modulation and/or manipulation of ventricular loading forces in the postinfarction setting, relatively few studies have investigated the potential merits of a patent infarct-related artery during postinfarction ventricular remodeling. In particular, the salutary effects of late revascularization of previously occluded vessels remains controversial. Thus, though this issue remains speculative, our present practice must be governed by circumstantial evidence and hypothetical arguments. This article discusses the potential mechanisms whereby late reperfusion of an infarcted myocardial region may benefit long-term prognosis. PMID- 10871165 TI - Does an open infarct-related artery after myocardial infarction improve electrical stability? AB - Arrhythmic events are responsible for the majority of sudden cardiac deaths after myocardial infarction. Many clinical studies have suggested that patency of the infarct-related artery, achieved by thrombolytic therapy or revascularization procedures, is a predictor of survival rates irrespective of myocardial salvage. The open-artery hypothesis suggests that an open infarct-related artery may result in other potential mechanisms, of benefits including electrical stability. This review focuses on the various levels and types of evidence supporting this contention. PMID- 10871166 TI - The role of myocardial viability in deriving benefit from reestablishing infarct related artery flow after acute myocardial infarction. AB - Early, sustained patency of the infarct-related artery (IRA) induces myocardial salvage, which preserves left ventricular (LV) function and mediates better long term outcome. However, the time course and the mechanisms of muscle recovery after myocardial infarction are not completely understood. A large body of evidence suggests that most of the improvement occurs during the hospital phase and is related to early and sustained thrombolysis in myocardial infarction 3 flow in the IRA. Nevertheless, the relationship between IRA status and regional and global LV mechanics in the chronic phase of the disease remains controversial. Some late recovery may occur, either spontaneously or after revascularization, even in the absence of documented myocardial ischemia. The interplay between vessel patency, coronary flow grade and severity of the residual stenosis, and the presence of stunned or hibernating myocardium in the area at jeopardy may explain this delayed improvement. Although there seems to be a limited time window in which myocardium can be salvaged, timely testing for viability, particularly in patients with poor LV function, is justified even in a later phase of the disease to challenge potential cardiac recovery. PMID- 10871167 TI - Beneficial effects of an open artery on left ventricular remodeling after myocardial infarction. AB - During the last 2 decades researchers have developed a clearer understanding of the pathophysiology of acute myocardial infarction (AMI). The use of thrombolysis and coronary angioplasty early in the course of AMI salvages myocardium and preserves left ventricular function, which results in improved survival rates. Late reperfusion after AMI, however, may provide beneficial effects on long-term prognosis, especially owing to attenuation of left ventricular remodeling. This article reviews the evidence of the benefits of an open infarct-related artery on left ventricular remodeling after AMI. PMID- 10871168 TI - Rationale and intended use for the Veress needle: a translation of the original descriptive article. AB - The technical development of equipment in the last decade has resulted in a rapid expansion in the range of procedures capable of being performed safely by a laparoscopic technique. For many procedures, the first step is induction of a pneumoperitoneum. This has inherent danger, and there is disagreement on the preferred technique. The Veress needle is an instrument developed in the 1930s that has continued to be used into the 1990s. In view of the controversy about its present role, the authors reviewed the article that provided the original description of the needle. This review demonstrates that the designer had a clear intention for its use and an understanding of the hazards involved. In his hands, the complications were few. A translation of the article from German into English is provided. PMID- 10871169 TI - Ergonomic surgeon's chair for use during minimally invasive surgery. AB - The characteristic working situation in laparoscopic surgery involves elongated instruments and limited mobility of the surgeon during the operation. These circumstances require new technical solutions to enhance the surgeon's comfort. In other surgical fields with special ergonomic situations, such as microsurgery, some surgeons prefer to operate from a seated position at the operating room table. We developed a new surgeon's chair dedicated to the ergonomic and functional requirements of laparoscopic surgery. The chair allows the surgeon to maintain a semi-standing position during the operation. Foot pedals for high frequency and suction/irrigation are integrated into the base of the chair. The pedals are purposely aligned to be comparable to foot pedals in a car. The chair is driven by electromotors, controlled with a special foot switch that operates independent of assisting personnel during surgery. Initial clinical testing of the chair could prove the theory that supporting the surgeon with a cockpit type of operating room chair helps to avoid fatigue during long endoscopic procedures. Such assistance is especially important in combination with robotic devices for use during solo surgery. PMID- 10871170 TI - Technique for laparoscopic gastric surgery. AB - As technology and surgeon experience expand, laparoscopic surgery is playing a larger role in the treatment of gastric conditions. We present our technical approach to various laparoscopic gastric resections and outline our preliminary results. Contrary to the majority of publications on laparoscopic gastric resection, we believe gastric mobilization should be carried out by incising the avascular plane between the greater omentum and transverse colon. This gives easy access to the origin of the left gastric artery and permits an acceptable D1 oncologic resection. For small lesions, tumor localization and resection margins should be mapped with the aid of routine intraoperative endoscopy. Nine patients underwent formal gastric resections, six of which were done for malignancy. Median time to discharge and length of follow-up were 4.5 days (range 3-10) and 25 months (range 24-35), respectively. One patient died postoperatively, and the remaining five patients operated for malignancy are alive and well with no evidence of recurrent disease or port site metastases. PMID- 10871171 TI - Management of early dislodgment of percutaneous endoscopic gastrostomy tubes. AB - One of the most serious complications of percutaneous endoscopic gastrostomy (PEG) is premature removal of the gastrostomy tube. In an attempt to clarify the optimal therapy of this complication, the records of 197 patients undergoing PEG were reviewed. Six patients whose PEG tubes were removed 2.9 +/- 1.3 days after placement were identified; only one patient required an emergent operation. The patients managed nonoperatively were treated by immediate replacement of tubes through the tracts (two patients), observation prior to repeat PEG (two), and delayed laparoscopic gastrostomy (one). Nonoperative management is feasible in the majority of patients suffering this complication. When an operation is indicated because of suspected intraperitoneal spillage, laparoscopy allows wide visualization and irrigation of the peritoneal cavity, closure of the gastrotomy, and placement of new enteral access while avoiding the morbidity associated with laparotomy in these often-debilitated patients. PMID- 10871172 TI - "Tension-free" hiatoplasty, gastrophrenic anchorage, and 360 degrees fundoplication in the laparoscopic treatment of paraesophageal hernia. AB - In our initial experience of four cases from March to November 1994, large paraesophageal hernias were repaired by conventional primary closure of the hiatus with interrupted, nonabsorbable sutures, adding a 360 degrees fundoplication. In all four cases the hernia recurred. Subsequently, we modified the procedure. The technique and results are described. From March 1995 to May 1998, 12 patients with paraesophageal hernia (4 following a previous Nissen procedure) underwent elective laparoscopic repair. In all patients a "tension free" hiatoplasty and a floppy 360 degrees fundoplication were performed. The hiatal defect was repaired with a polypropylene mesh, fixed to the diaphragm by staples. A gastrophrenic anchorage procedure was added in the eight patients undergoing surgery for the first time, utilizing the peritoneum of the hernia sac. There were no conversions to open surgery or intraoperative complications. Two patients developed postoperative pleural effusion, which was treated medically. Mean hospital stay was 5 days. Three patients developed postoperative transient dysphagia to solid food that lasted 10 days. At a mean follow-up of 22.7 months (range 1-40), all patients are asymptomatic without dysphagia, reflux, or hernia recurrence. Laparoscopic "tension-free" hiatoplasty, 360 degrees fundoplication, and anterior gastrophrenic anchorage are effective in the treatment of large paraesophageal hernias. PMID- 10871174 TI - Anatomic rationale for arterial bleeding from the liver bed during and/or after laparoscopic cholecystectomy: a postmortem study. AB - The aim of this study was to establish an anatomic rationale for liver bed arterial bleeding during laparoscopic cholecystectomy. Fifty consecutive human cadavers were dissected. A corrosion cast method was used. Six anastomotic branches (12%) of the cystic artery to the right or left hepatic artery ran underneath the gallbladder serosa surface and entered liver parenchyma after crossing the medial or lateral edge of the liver fossa without passing through the areolar tissue of the liver bed. Their mean length was 18.3 mm (range 4-60), and the mean diameter was 0.38 mm (range 0.2-0.8). Two cystic arteries that ascended in the midline between the gallbladder and liver bed were identified in 50 (4%) casts. Their lengths were 16 and 18 mm, and their diameters were 1.9 and 2.2 mm. Five and seven branches encircling the gallbladder arose radially. These two arterial branching patterns can cause arterial bleeding from the liver bed during and/or after laparoscopic cholecystectomy. PMID- 10871173 TI - Gallbladder rupture during laparoscopic cholecystectomy: does it have an effect on postoperative morbidity? AB - Gallbladder rupture during laparoscopic cholecystectomy is a common event that may lead to increased postoperative morbidity. To evaluate this event, we reviewed 300 cases of laparoscopic cholecystectomy. Duration of surgery and hospitalization, postoperative symptoms, wound infection, and late complications were analyzed by comparing two groups of patients, one without gallbladder rupture (A) and one with rupture (B). Gallbladder rupture was found in 40 cases (13.9%). Duration of surgery averaged 81 min for group A and 96.5 min for group B. Postoperative symptoms in the first 24 hours were present in approximately 10% of patients in both groups. Within the first 24 hours, 92.3% of patients in group A were discharged compared with 85% in group B. One patient (0.4%) in group A developed wound infection compared with 2 patients (5%) in group B (p = 0.05). To date, no patients have developed late abdominal complications associated with the procedure. Although this was a retrospective and uncontrolled study, gallbladder rupture during laparoscopic cholecystectomy was found to be associated with increased wound infections. No other significant effects on postoperative morbidity were detected. PMID- 10871175 TI - Laparoscopic appendectomy for ruptured appendicitis. AB - We conducted a retrospective analysis to assess the feasibility of laparoscopic appendectomy in cases of ruptured appendicitis. Between August 1993 and April 1998, a total of 328 laparoscopic appendectomies were performed in Min-Shen General Hospital. There were 34 cases of pathology-proven ruptured appendicitis. Patients were divided into three groups according to the operative findings: group 1 (10 cases) consisted of patients with a perforated appendix with local peritonitis, group 2 (15 cases) consisted of patients with perforated appendix with diffused peritonitis, and group 3 (9 cases) consisted of patients with abscess formation around the perforated appendix. Three cases in group 3 were converted to laparotomy and were excluded from this study. Mean age (+/- SD) was 30 +/- 15 years in group 1, 39 +/- 23 years in group 2, and 37 +/- 13 years in group 3. Duration of symptoms was longer in group 3 (4.2 +/-1.2 days) than in group 1 (1.8 +/- 1.3 days) and group 2 (2.3 +/- 1.2 days). There was no difference in operation time among groups 1 (52 +/- 10 min), 2 (64 +/- 13 min), and 3 (67 +/- 16 min). The time of flatus passage after operation was similar in the three groups (group 1, 17 +/- 11 hours; group 2, 21 +/- 12 hours; group 3, 24 +/- 8 hours). Hospital stay was significantly shorter in group 1 (3.0 +/- 1.1 days) than in group 2 (5.1 +/- 2.2 days) and group 3 (4.2 +/- 1.2 days). There were no complications and no readmissions. Our results indicate that the laparoscopic approach is feasible for ruptured appendicitis with local or diffuse peritonitis and in selected cases with abscess formation. However, prospective randomized controlled trials are needed to determine which procedure is to be recommended. PMID- 10871176 TI - Prevention of neoplastic port site implants in laparoscopy: an experimental study. AB - Enthusiasm about the application of videolaparoscopy to oncologic diseases has been limited by the growing number of port site implants. Adult Wistar rats were submitted to 6-7 mm Hg carbonic gas pneumoperitoneum. Rats were randomly divided into two groups: group I rats with tumor (200,000 viable cells of Walker tumor) and group 11 rats with no tumor. The pneumoperitoneum was deflated after 30 min. Group I was further randomized into five groups: no treatment; or abdominal irrigation with saline, heparin, chemotherapy (doxorubicin), or chemotherapy associated with heparin. After a period lasting no more than 18 days, the abdominal wall and intraperitoneal organs macroscopically affected were studied histologically. Chemotherapy groups had no port site implants and were significantly different (p < 0.05) than the no treatment, saline, and heparin solution groups, which had incisional implants at frequencies of 100%, 85.7%, and 82.5%, respectively. Intraperitoneal irrigation with chemotherapy solution was effective in preventing incisional implants in this animal model. PMID- 10871177 TI - Convenient murine pneumoperitoneal model for the study of laparoscopic cancer surgery. AB - To investigate the effects of laparoscopic surgery on the progression of cancers, it is necessary to establish a reliable and economical animal model. We describe a convenient murine pneumoperitoneal model for the study of laparoscopic cancer surgery. Under anesthesia using diethyl ether, peritoneal cavity was insufflated with gas through an intravenous catheter placed in the left lower quadrant. Syringe pump was used for continuous gas insufflation, and intraperitoneal pressure was measured. Intraperitoneal pressure increased and reached 10 cm H2O when 15 mL of CO2 gas was injected, but fell to 1 cm H2O 5 min after stopping the injection. When the continuous flow was adjusted by syringe pump between 20 and 160 mL/hour, intraperitoneal pressure was easily maintained at 8 cm H2O for >60 min. We believe that this murine model for pneumoperitoneum may be useful for the study of laparoscopic cancer surgery. PMID- 10871178 TI - Thoracoscopic approach to giant lymph node hyperplasia (Castleman's disease). AB - We describe the case of a thoracoscopic approach to giant lymph node hyperplasia (Castleman's disease) located in the mediastinum. In our patient the initial diagnosis was substernal goiter, but at cervical exploration the mass was found not to be continuous with the thyroid. The mass was easily identified through a lateral thoracoscopic approach and carefully removed. The postoperative course was uneventful, and the patient was discharged home on the fifth postoperative day. Histopathology revealed the features of Castleman's disease, mixed type. The prevalence, location, and pathogenesis of giant lymph node hyperplasia are described, together with the histology and clinical signs. Our report is proposed as the first case of an endoscopic approach to giant lymph node hyperplasia, which could be the best surgical approach for a mediastinal location. PMID- 10871179 TI - Simplified laparoscopic approach to "second-look" laparotomy: a review. AB - Acute mesenteric vascular accidents are being diagnosed more commonly as a consequence of an aging population and often result in emergency bowel resection for ischemia. Because predicting postoperative intestinal viability remains difficult, second-look laparotomy has been advocated to improve outcomes. Recently, laparoscopy has emerged as an alternative to laparotomy for the diagnosis and treatment of ongoing postoperative ischemia. A review of the literature since 1994 reveals that, to date, 19 procedures have been reported to prevent 13 (68%) unnecessary laparotomies. We describe our laparoscopic second- look technique and review the literature. Second-look laparoscopy has been shown repeatedly to be a safe alternative to laparotomy. It is simple and reduces negative second-look laparotomy in critically ill patients. PMID- 10871180 TI - Laparoscopic distal pancreatectomy with preservation of the spleen. AB - We describe a case of chronic pancreatitis treated by laparoscopic distal pancreatectomy with conservation of the spleen involving the resection of the splenic vessels. A proximal ligation of the splenic artery and vein was performed, followed by transection of the body of the pancreas. Retroperitoneum was dissected to the left by mobilizing the stump of the transected pancreas. The entire distal pancreas was freed posteriorly. The distal splenic artery and vein were ligated and divided individually adjacent to the tail of the pancreas at the hilum of the spleen. The points of this operation were to ligate the splenic artery and vein at both sides of the resected pancreas and to save the spleen with the blood supply continuing through the short gastric vessels and the splenocolic ligament. This operation with splenic preservation is more suitable for a patient who is a candidate for laparoscopic distal pancreatectomy, which will minimize the operation time, preserve the useful immunologic role of the spleen, and obtain the intact resected specimen. Furthermore, this procedure is useful in chronic pancreatitis patients because it avoids the difficult dissection of the posterior pancreas off of the splenic vessels. PMID- 10871181 TI - Revision laparoscopy for incarcerated hernia at a 5-mm trocar site following pediatric laparoscopic surgery. AB - We report the case of a 6-month-old female infant who developed post-operative bowel obstruction due to an incarcerated hernia through a 5-mm laparoscopic wound. The patient underwent laparoscopic Nissen fundoplication for gastroesophageal reflux. On day 6, she showed symptoms of ileus, and the diagnosis of a trocar wound hernia was made on day 13. The herniated intestine was reduced and the defective peritoneum and fascia were closed under relaparoscopic guidance, thus avoiding full-scale laparotomy. A trocar wound hernia causing early postoperative bowel obstruction is a rare complication, especially at 5-mm trocar puncture sites. Intraoperative dislodgment and reinsertion of working trocars may create fascial defects larger than the actual size of the trocar. All laparoscopic puncture wounds, even those <10 mm in size, should be closed at the fascial level in infants. Revision laparoscopy is considered preferable to manage trocar site complications in children. PMID- 10871182 TI - Laparoscopic management of spermatic cord entrapment after laparoscopic inguinal herniorrhaphy. AB - Spermatic cord entrapment is an unusual complication of inguinal herniorrhaphy. The case of a 52-year-old man who presented with severe pain along the left spermatic cord and testicle, varicocele, and dyspareunia after a laparoscopic bilateral inguinal hernia repair performed elsewhere is reported. Medical treatment failed, and laparoscopic exploration showed the vas deferens and spermatic vessels entrapped by a mesh slit that was pulling the genital branch of the genitofemoral nerve. The vas deferens and spermatic vessels were released, neurotomy of the affected nerve branch was performed, and a new mesh was positioned in the residual space. The patient's pain disappeared completely after the surgery and the varicocele decreased progressively. The patient remains asymptomatic at 1-year follow-up. Laparoscopy might be the approach of choice to treat some of the complications of laparoscopic hernia repair, not only because it allows better observation of the anatomic structures, but also because the surgical therapy can be done with minimal tissue damage compared to the traditional approach. PMID- 10871184 TI - Laparoscopic management of operative vena cava injury. AB - Major vascular injury can result from trocar placement, but it is a rare event during an intra-abdominal laparoscopic procedure. The authors report a major vena cava injury that occurred during a right adrenalectomy. The injury was managed entirely laparoscopically. Management of a major vascular injury occurring in such circumstances requires appropriate instruments and equipment, including a vascular clamp and good suction. Conversion to an open procedure is recommended if the injury cannot be safely resolved laparoscopically. PMID- 10871183 TI - Ureteral complications after gasless laparoscopic hysterectomy. AB - Laparoscopic hysterectomy is becoming a more common operation. Gasless laparoscopy initially seems to be a better technique, reducing CO2 complications and allowing the use of conventional instruments rather than more expensive laparoscopic tools. We report our experience with 50 cases of laparoscopic hysterectomies, 5 of which were performed using a gasless technique. Of these five cases, there were two cases of ureteral stenosis. Ureteral injuries are common with hysterectomy, even in nonlaparoscopic procedures, and the literature is replete with recommendations to avoid this damage. In the gasless procedure, the ureters cannot be repositioned completely from the cervix after the hydrodissection. Extreme caution must be taken when applying bipolar or monopolar energy. The abdominal cavity shape does not allow complete avoidance of the ureters using the gasless technique. We have decided not to use a gasless technique with hysterectomy. We believe that the actual complication rate may be higher than reported, due to investigators' reluctance to report such complications. Our hope is that this report will encourage other investigators to help establish a more accurate rate of possible complications associated with this procedure. PMID- 10871185 TI - Laparoscopic resection of a retroperitoneal schwannoma. AB - Although laparoscopic surgery for retroperitoneal diseases has been widely performed, there are few reports of laparoscopic resection for retroperitoneal tumors. We present the case of a 5-cm retroperitoneal tumor compressing the right common iliac vein and inferior vena cava that was successfully resected using a laparoscopic technique. Dissection between the tumor and the large vessels was performed safely using a harmonic scalpel and an ultrasonic surgical aspirator. Histopathology of the resected tumor showed a benign schwannoma. Laparoscopic surgical techniques should be considered for treatment of selected retroperitoneal tumors. PMID- 10871186 TI - Looking back on open hepatectomies for liver cysts in the era of endosurgery. PMID- 10871187 TI - Three-port laparoscopic cholecystectomy: a plea for scientific evidence. PMID- 10871188 TI - Metabolic stress and altered glucose transport: activation of AMP-activated protein kinase as a unifying coupling mechanism. AB - 5'AMP-activated protein kinase (AMPK) can be activated in response to cellular fuel depletion and leads to switching off ATP-consuming pathways and switching on ATP-regenerating pathways in many cell types. We have hypothesized that AMPK is a central mediator of insulin-independent glucose transport, which enables fuel depleted muscle cells to take up glucose for ATP regeneration under conditions of metabolic stress. To test this hypothesis, rat epitrochlearis muscles were isolated and incubated in vitro under several conditions that evoke metabolic stress accompanied by intracellular fuel depletion. Rates of glucose transport in the isolated muscles were increased by all of these conditions, including contraction (5-fold above basal), hypoxia (8-fold), 2,4-dinotrophenol (11-fold), rotenone (7-fold), and hyperosmolarity (8-fold). All of these stimuli simultaneously increased both alpha1 and alpha2 isoform-specific AMPK activity. There was close correlation between alpha1 (r2 = 0.72) and alpha2 (r2 = 0.67) AMPK activities and the rate of glucose transport, irrespective of the metabolic stress used, all of which compromised muscle fuel status as judged by ATP, phosphocreatine, and glycogen content. 5-Aminoimidazole-4-carboxamide ribonucleoside, a pharmacological AMPK activator that is metabolized to an AMP mimetic ZMP, also increased both glucose transport and AMPK activity but did not change fuel status. Insulin stimulated glucose transport by 6.5-fold above basal but did not affect AMPK activity. These results suggest that the activation of AMPK may be a common mechanism leading to insulin-independent glucose transport in skeletal muscle under conditions of metabolic stress. PMID- 10871189 TI - Immunohistochemical and ultrastructural localization of leptin and leptin receptor in human white adipose tissue and differentiating human adipose cells in primary culture. AB - Leptin is mainly produced in white adipose tissue and acts both at distant sites and locally at the tissue from which it originates. The cellular and subcellular localization of leptin and its receptor (Ob-receptor [Ob-R]) and their relationship to various stages of fat cell maturation have not been characterized as yet. Therefore, we analyzed leptin and Ob-R by using reverse transcriptase polymerase chain reaction, immunohistochemistry, and ultrastructural immunogold labeling in human white adipose tissue and in human adipocyte cell cultures at early and late stages of differentiation. Both leptin and its receptor were present in mature unilocular fat cells. The thin cytoplasmic rim of the adipocytes exhibited the strongest expression of both leptin and Ob-R. At early stages of differentiating human adipocytes, leptin was mainly expressed in multilocular preadipocytes, whereas the Ob-R was found predominantly on fibroblast-like cells. Other cellular components of human white adipose tissue were characterized by anti-CD31 for endothelial cells, anti-CD68 for macrophages, and antibodies specifically labeling B-cells and T-cells. In addition to fat cells, endothelial cells were immunopositive for the full-length leptin receptor. On the ultrastructural level, leptin was mainly found attached to cellular membranes and in small alveolate vesicle-like structures in the cytoplasm of adipocytes. Leptin was also present on the cell membranes of endothelial cells and macrophages. We conclude that the expression of the Ob-R in human white adipose tissue is not restricted to adipocytes but is present in resident endothelial and immune cells. Ultrastructural localization studies revealed an association of leptin with cell membranes and small vesicles. The cellular and subcellular distribution of leptin and its receptor suggests an important autocrine and paracrine role for leptin in human adipose tissue. PMID- 10871190 TI - Differential activation of peroxisome proliferator-activated receptor-gamma by troglitazone and rosiglitazone. AB - The antidiabetic thiazolidinediones, which include troglitazone and rosiglitazone, are ligands for the nuclear receptor peroxisome proliferator activated receptor (PPAR)-gamma and exert their antihyperglycemic effects by regulation of PPAR-gamma-responsive genes. We report here that PPAR-gamma activation by troglitazone depends on the experimental setting. Troglitazone acts as a partial agonist for PPAR-gamma in transfected muscle (C2C12) and kidney (HEK 293T) cells, producing a submaximal transcriptional response (1.8- to 2.5-fold activation) compared with rosiglitazone (7.4- to 13-fold activation). Additionally, troglitazone antagonizes rosiglitazone-stimulated PPAR-gamma transcriptional activity. Limited protease digestion of PPAR-gamma suggests conformational differences in the receptor bound to troglitazone versus rosiglitazone. Consistent with this finding, an in vitro coactivator association assay demonstrated that troglitazone-bound PPAR-gamma recruited the transcriptional coactivators p300 and steroid receptor coactivator 1 less efficiently than rosiglitazone-bound receptor. In contrast to these observations, troglitazone behaves as a full agonist of PPAR-gamma in 3T3L1 adipocytes. Two dimensional protein gel electrophoresis demonstrated that troglitazone and rosiglitazone regulated distinct but overlapping sets of genes in several cell types. Thus, troglitazone may behave as a partial agonist under certain physiological circumstances and as a full agonist in others. These differences could be caused by variations in the amount of specific cofactors, differences in PPAR response elements, or the presence of different isoforms of PPAR-gamma. PMID- 10871191 TI - Co-expression of HLA DR3 and DQ8 results in the development of spontaneous insulitis and loss of tolerance to GAD65 in transgenic mice. AB - Specific HLA DQ and DR alleles have been associated with susceptibility to type 1 diabetes. HLA DQ8 and DQ2 have been shown to strongly predispose to disease and to be in linkage disequilibrium with at-risk DR4 and DR3 alleles, respectively. Inheritance of a mixed DR3/DR4 haplotype confers the greatest risk. A double transgenic mouse expressing both DR3 and DQ8 was generated to investigate potential major histocompatibility complex class II interactions. The DR3/DQ8 transgenic mice developed a spontaneous loss of tolerance to GAD65, in which the T-cell response to GAD65 was restricted by HLA DR. Although the mice also showed spontaneous insulitis, they did not progress to overt diabetes. Mice expressing either transgene (DQ8 or DR3) alone showed mild infiltration of their islets, which disappeared when DQ8 or DR3 was co-expressed with a resistant DR2 allele or the neutral DQ6 allele. Therefore, in a fashion analogous to human diabetes, the murine model demonstrated a requirement for a combination of at-risk DR and DQ allotypes for the initiation of spontaneous autoimmunity. PMID- 10871192 TI - Sex-determining region Y-related protein SOX13 is a diabetes autoantigen expressed in pancreatic islets. AB - The SOX (sex-determining region [SRY]-type high mobility group [HMG] box) family of transcription factors play key roles in determining cell fate during organ development. In this study, we have identified a new human SOX gene, SOX13, as encoding the type 1 diabetes autoantigen, islet cell antigen 12 (ICA12). Sequence analysis showed that SOX13 belongs to the class D subgroup of SOX transcription factors, which contain a leucine zipper motif and a region rich in glutamine. SOX13 autoantibodies occurred at a significantly higher frequency among 188 people with type 1 diabetes (18%) than among 88 with type 2 diabetes (6%) or 175 healthy control subjects (4%). Deletion mapping of the antibody epitopes showed that the autoantibodies were primarily directed against an epitope requiring the majority of the protein. SOX13 RNA was detected in most human tissues, with the highest levels in the pancreas, placenta, and kidney. Immunohistochemistry on sections of human pancreas identified SOX13 in the islets of Langerhans, where staining was mostly cytoplasmic. In mouse pancreas, Sox13 was present in the nucleus and cytoplasm of beta-cells as well as other islet cell types. Recombinant SOX13 protein bound to the SOX consensus DNA motif AACAAT, and binding was inhibited by homodimer formation. These observations-along with the known molecular interactions of the closely related protein, rainbow trout Sox23 suggest that SOX13 may be activated for nuclear import and DNA binding through heterodimer formation. In conclusion, we have identified ICA12 as the putative transcription factor SOX13 and demonstrated an increased frequency of autoantibody reactivity in sera from type 1 diabetic subjects compared with type 2 diabetic and healthy control subjects. PMID- 10871193 TI - Selection of insulinoma cell lines with resistance to interleukin-1beta- and gamma-interferon-induced cytotoxicity. AB - Engineered insulinoma cell lines may represent an alternative to isolated islets for transplantation therapy of type 1 diabetes. Success of this approach may require development of cell lines that can withstand cytokine-mediated damage. To this end, we have cultured INS-1 insulinoma cells in increasing concentrations of interleukin-1beta (IL-1beta) + gamma-interferon (IFN-gamma), with approximate weekly iterations over an 8-week period. Based on the C,N diphenyl-N'-[4,5 dimethylthiazol-2-yl]-2,5-diphenyltetrazolium+ ++ bromide (MTT) viability assay, the selected cells, termed INS-1res, were 100% viable after 5 days of treatment with 10 ng/ml of IL-1beta. These cells were also 78 +/- 1.2% viable after 5 days of exposure to the combination of 10 ng/ml IL-1beta and 100 U/ml IFN-gamma, whereas parental INS-1 cells treated in the same manner were only 0.3 +/- 0.03% viable. INS-1res cells were also resistant to treatment with supernatants from activated rat peripheral blood mononuclear cells, whereas only 20% of parental INS-1 cells survived such treatment. The resistance to IL-1beta conferred by this procedure was stable, whereas the partial resistance to IFN-gamma was transient but reinducible by culture in the presence of cytokines. Stable transfection of INS-1res cells with a plasmid containing the human insulin cDNA and expansion of the transfected colonies in the absence of cytokines produced cell lines that were on average more resistant to IL-1beta + IFN-gamma (53 +/- 11%) than similarly transfected clones derived from parental INS-1 cells (15 +/- 7%). Importantly, several INS-1res-derived clones retained the capacity to secrete insulin in response to glucose concentrations over the normal physiological range. With regard to the mechanism by which selection was conferred, we found normal levels of IFN-gamma receptor mRNA, but a 60% reduction in expression of the IL-1 receptor type I (IL-1RI) in INS-1res cells compared with parental INS-1 cells. IL-1beta signaling through p38 MAP kinase was found to be normal in INS 1res cells, suggesting that their expression of IL-1RI is sufficient to maintain cytokine action. However, normal IL-1beta-mediated translocation of NF-kappaB and induction of inducible nitric oxide synthase expression and nitric oxide production was severely impaired in the INS-1res cell lines, suggesting a mechanism for the IL-1beta resistance. In sum, this study defines a strategy for isolation of cytokine-resistant beta-cell lines and provides a new system for studying the mechanisms by which such resistance can be achieved. PMID- 10871194 TI - Adult human pancreatic duct and islet cells exhibit similarities in expression and differences in phosphorylation and complex formation of the homeodomain protein Ipf-1. AB - The homeodomain transcription factor encoded by the pancreatic and duodenal homeobox gene-1 (Ipf-1) is essential for pancreatic ontogenesis. Whether Ipf-1 is also involved in the neogenesis of beta-cells in the adult pancreas is unknown. We examined whether Ipf-1 is expressed in adult human pancreatic ducts, which are thought to generate new beta-cells. In tissue sections, virtually all duct cells were immunopositive for Ipf-1, as were the islet beta-cells but not the acinar cells. After isolation and culture, both duct and islet cell preparations contained the Ipf-1 immunoreactive proteins p42 and p45 (42 and 45 kDa, respectively) in similar proportions, but the expression levels were twofold lower in duct cells. After 4 h of labeling, the endocrine cells exhibited a sevenfold higher phosphorylation of p42 than the duct cells, whereas p45 was phosphorylated only in endocrine cells. Homeobox binding transcription factor complexes with Ipf-1 in duct cells differed from those in endocrine cells in terms of gel mobility, sequence specificity, and affinity. The observed similarities in Ipf-1 expression by adult human pancreatic duct cells and endocrine cells may reflect their common ontogenic origin, whereas the differences in Ipf-1 phosphorylation and complex formation may correlate with their divergent differentiation. PMID- 10871195 TI - Validation of methods for measurement of insulin secretion in humans in vivo. AB - To detect and understand the changes in beta-cell function in the pathogenesis of type 2 diabetes, an accurate and precise estimation of prehepatic insulin secretion rate (ISR) is essential. There are two common methods to assess ISR, the deconvolution method (by Eaton and Polonsky)-considered the "gold standard" and the combined model (by Volund et al.). The deconvolution method is a 2-day method, which generally requires separate assessment of C-peptide kinetics, whereas the combined model is a single-day method that uses insulin and C-peptide data from a single test of interest. The validity of these mathematical techniques for quantification of insulin secretion have been tested in dogs, but not in humans. In the present studies, we examined the validity of both methods to recover the known infusion rates of insulin and C-peptide mimicking ISR during an oral glucose tolerance test. ISR from both the combined model and the deconvolution method were accurate, i.e., recovery of true ISR was not significantly different from 100%. Furthermore, both maximal and total ISRs from the combined model were strongly correlated to those obtained by the deconvolution method (r = 0.89 and r = 0.82, respectively). These results indicate that both approaches provide accurate assessment of prehepatic ISRs in type 2 diabetic patients and control subjects. A simplified version of the deconvolution method based on standard kinetic parameters for C-peptide (Van Cauter et al.) was compared with the 2-day deconvolution method, and a close agreement was found for the results of an oral glucose tolerance test. We also studied whether C-peptide kinetics are influenced by somatostatin infusion. The decay curves after bolus injection of exogenous biosynthetic human C-peptide, the kinetic parameters, and the metabolic clearance rate were similar whether measured during constant peripheral somatostatin infusion or without somatostatin infusion. Assessment of C-peptide kinetics can be performed without infusion of somatostatin, because the endogenous insulin concentration remains constant. Assessment of C-peptide kinetics with and without infusion of somatostatin results in nearly identical secretion rates for insulin during an oral glucose tolerance test. PMID- 10871196 TI - Genetic modifiers of the insulin resistance phenotype in mice. AB - Insulin resistance can result from genetic interactions among susceptibility alleles. To identify genetic loci predisposing to insulin resistance, we used crosses between different strains of mice with a targeted null allele of the insulin receptor gene. On the genetic background of B6 mice, the insulin receptor gene mutation causes mild hyperinsulinemia. In contrast, on the genetic background of 129/Sv mice, the same mutation causes severe hyperinsulinemia, suggesting that the 129/Sv strain harbors alleles that interact with the insulin receptor mutation and predispose to insulin resistance. As a first step to identify these alleles, we generated an F2 intercross between insulin receptor heterozygous mutant mice on B6 and 129/Sv backgrounds (B6IR x 129IR) and performed a genome-wide scan with polymorphic markers at a 20-cM resolution. We report the identification of loci on chromosomes 2 (logarithm of odds [LOD] 5.58) and 10 (LOD 5.58) that show significant evidence for linkage to plasma insulin levels as a quantitative trait. These findings indicate that targeted mutations in knockout mice can be used to unravel the complex genetic interactions underlying insulin resistance. PMID- 10871197 TI - Upregulation of macrophage lipoprotein lipase in patients with type 2 diabetes: role of peripheral factors. AB - Atherosclerosis is the major complication of diabetes. Accumulating evidence indicates that lipoprotein lipase (LPL) produced by macrophages in the vascular wall may favor the development of atherosclerosis by promoting lipid accumulation within the lesion. We previously demonstrated that high glucose stimulates in vitro murine and human macrophage LPL production. In this study, we measured macrophage LPL mRNA expression, immunoreactive mass, and activity in normotriglyceridemic subjects with type 2 diabetes. Monocytes isolated from healthy control subjects and patients with type 2 diabetes were differentiated into macrophages in RPMI medium containing 20% autologous serum. After 5 days in culture, macrophage LPL mRNA expression, mass, and activity were determined. Macrophages of diabetic patients cultured in their own sera showed a significant increase in LPL mRNA levels, mass, and activity compared with macrophages of control subjects. Differentiation of macrophages of diabetic patients in sera obtained from control subjects significantly reduced these anomalies. Conversely, culturing macrophages of control subjects in sera of diabetic patients significantly increased LPL mass and activity in these cells. Besides LPL overproduction, macrophages of diabetic patients exhibited an increase in basal and LPL-induced tumor necrosis factor (TNF)-alpha release. TNF-alpha alterations were reduced by exposing these cells to sera of control subjects. Overall, these data demonstrate that macrophages of diabetic patients overexpress LPL and TNF alpha and that peripheral factors dysregulated in diabetes are, at least in part, responsible for these alterations. PMID- 10871198 TI - Decreased insulin receptor tyrosine kinase activity and plasma cell membrane glycoprotein-1 overexpression in skeletal muscle from obese women with gestational diabetes mellitus (GDM): evidence for increased serine/threonine phosphorylation in pregnancy and GDM. AB - The cellular mechanisms for the insulin resistance of pregnancy and gestational diabetes mellitus (GDM) are unknown. The membrane protein plasma cell membrane glycoprotein-1 (PC-1) has been identified as an inhibitor of insulin receptor tyrosine kinase (IRTK) activity. We investigated insulin receptor function and PC 1 levels in muscle from three groups of obese subjects: women with GDM, pregnant women with normal glucose tolerance, and nonpregnant control subjects. Subjects (n = 6 for each group) were similar in age and degree of obesity (body fat >30%). IRTK activity, insulin receptor tyrosine phosphorylation, and protein levels of membrane glycoprotein PC-1 were determined in rectus abdominus muscle biopsies obtained at the time of either elective cesarean section or gynecological surgery. No significant differences were evident in basal insulin receptor tyrosine phosphorylation or IRTK activity in the three groups. After maximal insulin (10(-7) mol/l) stimulation, IRTK activity measured with the artificial substrate poly(Glu,Tyr) increased in all subjects but was lower in women with GDM by 25% (P < 0.05) and 39% (P < 0.001) compared with pregnant and nonpregnant control subjects, respectively. Similarly, insulin receptor tyrosine phosphorylation was significantly decreased in subjects with GDM (P < 0.05) compared with pregnant and nonpregnant control subjects. Treatment of the insulin receptors with alkaline phosphatase to dephosphorylate serine/threonine residues increased insulin-stimulated IRTK activity significantly in pregnant control and GDM subjects (P < 0.05), but these rates were still lower compared with nonpregnant control subjects (P < 0.05). PC-1 content in muscle from GDM subjects was increased by 63% compared with pregnant control subjects (P < 0.05) and by 206% compared with nonpregnant control subjects (P < 0.001). PC-1 content was negatively correlated with insulin receptor phosphorylation (r = -0.55, P < 0.05) and IRTK activity (r = -0.66, P < 0.05). These results indicate that pregnant control and GDM subjects had increased PC-1 content and suggest excessive phosphorylation of serine/threonine residues in muscle insulin receptors and that both may contribute to decreased IRTK activity. These changes worsen in women with GDM when controlling for obesity. These postreceptor defects in insulin signaling may contribute to the pathogenesis of GDM and the increased risk for type 2 diabetes later in life. PMID- 10871199 TI - Effect of glucagon-like peptide 1(7-36) amide on glucose effectiveness and insulin action in people with type 2 diabetes. AB - Although it is well established that glucagon-like peptide 1(7-36) amide (GLP-1) is a potent stimulator of insulin secretion, its effects on insulin action and glucose effectiveness are less clear. To determine whether GLP-1 increases insulin action and glucose effectiveness, subjects with type 2 diabetes were studied on two occasions. Insulin was infused during the night on both occasions to ensure that baseline glucose concentrations were comparable. On the morning of study, either GLP-1 (1.2 pmol x kg(-1) x min(-1)) or saline were infused along with somatostatin and replacement amounts of glucagon. Glucose also was infused in a pattern mimicking that typically observed after a carbohydrate meal. Insulin concentrations were either kept constant at basal levels (n = 6) or varied so as to create a prandial insulin profile (n = 6). The increase in glucose concentration was virtually identical on the GLP-1 and saline study days during both the basal (1.21 +/- 0.15 vs. 1.32 +/- 0.19 mol/l per 6 h) and prandial (0.56 +/- 0.14 vs. 0.56 +/- 0.10 mol/l per 6 h) insulin infusions. During both the basal and prandial insulin infusions, glucose disappearance promptly increased after initiation of the glucose infusion to rates that did not differ on the GLP 1 and saline study days. Suppression of endogenous glucose production also was comparable on the GLP-1 and saline study days during both the basal (-2.7 +/- 0.3 vs. -3.1 +/- 0.2 micromol/kg) and prandial (-3.1 +/- 0.4 vs. -3.0 +/- 0.6 pmol/kg) insulin infusions. We conclude that when insulin and glucagon concentrations are matched, GLP-1 has negligible effects on either insulin action or glucose effectiveness in people with type 2 diabetes. These data strongly support the concept that GLP-1 improves glycemic control in people with type 2 diabetes by increasing insulin secretion, by inhibiting glucagon secretion, and by delaying gastric emptying rather than by altering extrapancreatic glucose metabolism. PMID- 10871200 TI - v- and t-SNARE protein expression in models of insulin resistance: normalization of glycemia by rosiglitazone treatment corrects overexpression of cellubrevin, vesicle-associated membrane protein-2, and syntaxin 4 in skeletal muscle of Zucker diabetic fatty rats. AB - Insulin stimulation of adipose and muscle cells results in the translocation of GLUT4 from an intracellular location to the plasma membrane; this translocation is defective in insulin resistance. Studies have suggested an important role for synaptobrevin and syntaxin homologues in this event, particularly the v-soluble N ethylmaleimide attachment protein receptors (SNAREs) cellubrevin and vesicle associated membrane protein-2 (VAMP-2) and the t-SNARE syntaxin 4, but the expression of these proteins has not been studied in insulin-resistant tissues. Therefore, we examined SNARE protein content in skeletal muscle from Zucker diabetic fatty (ZDF) rats compared with lean controls and determined the effect of the thiazolidinedione insulin sensitizer rosiglitazone on these proteins. GLUT4 levels in skeletal muscle from ZDF rats were similar to those in lean control animals. In contrast, cellubrevin, VAMP-2, and syntaxin 4 protein levels were elevated (2.8-fold, P = 0.02; 3.7-fold, P = 0.01; and 2.2-fold, P < 0.05, respectively) in skeletal muscle from ZDF rats compared with lean controls. Restoration of normoglycemia and normoinsulinemia in ZDF rats with rosiglitazone (30 micromol/kg) normalized cellubrevin, VAMP-2, and syntaxin 4 protein to levels approaching those observed in lean control animals. These data show that elevated v- and t-SNARE protein levels are associated with insulin resistance in skeletal muscle and that these increases may be reversed by rosiglitazone treatment concomitant with a restoration of glycemic control. Such increases in SNARE protein levels were not observed in streptozotocin-induced diabetic rats, which suggests that hyperinsulinemia rather than hyperglycemia may be more important in modulating SNARE protein expression in rodent models of insulin resistance. Consistent with this hypothesis, elevated levels of SNARE proteins were also observed in 3T3-L1 adipocytes chronically treated with insulin (500 nmol/l for 24 h). These data argue that SNARE protein levels may be altered in insulin resistant states and that the levels of these proteins are modulated by agents that increase insulin sensitivity. Moreover, these data demonstrate for the first time altered expression of proteins known to regulate GLUT4 translocation in a model of diabetes. PMID- 10871201 TI - Can clinical factors estimate insulin resistance in type 1 diabetes? AB - An insulin resistance syndrome (IRS) score was developed based on clinical risk factors in adults with childhood-onset type 1 diabetes in the Epidemiology of Diabetes Complications (EDC) Study and was validated using euglycemic hyperinsulinemic clamp studies. Hypertension, waist-to-hip ratio (WHR), triglyceride and HDL cholesterol levels, family history of type 2 diabetes, and glycemic control were risk factors used to define the score. A score of 1 (lowest likelihood IRS) to 3 (highest likelihood IRS) was assigned for each risk factor. Eligible subjects (n = 24) were recruited from the EDC cohort based on tertile of IRS score. Subjects received an overnight insulin infusion to normalize glucose levels, then underwent a 3-h euglycemic-hyperinsulinemic (60 mU x m(-2) x min( 1)) clamp. Glucose disposal rate (GDR) was determined during the last 30 min of the clamp. The GDR differed significantly by IRS group (9.65 +/- 2.99, 8.02 +/- 1.39, and 5.68 +/- 2.16 mg x kg(-1) x min(-1), P < 0.01). The GDR was inversely correlated with the IRS score (r = -0.64, P < 0.01). Using linear regression, the combination of risk factors that yielded the highest adjusted r2 value (0.57, P < 0.001) were WHR, hypertension, and HbA1. This study found that clinical risk factors can be used to identify subjects with type 1 diabetes who are insulin resistant, and it provides validation of a score based on clinical factors to determine the extent of insulin resistance in type 1 diabetes. This score will be applied to the entire EDC population in future studies to determine the effect of insulin resistance on complications. PMID- 10871202 TI - Effects of troglitazone on blood concentrations of plasminogen activator inhibitor 1 in patients with type 2 diabetes and in lean and obese normal subjects. AB - Low plasma fibrinolytic activity in association with increased plasma plasminogen activator inhibitor 1 (PAI-1) levels has been linked to an increased risk of atherosclerosis in obesity and type 2 diabetes. We tested the hypothesis that troglitazone, which improves insulin sensitivity and lowers plasma insulin levels in insulin-resistant obese subjects and patients with type 2 diabetes, would also lower circulating PAI-1 antigen concentrations and activity. We assessed insulin sensitivity (5-h, 80 mU x m(-2) x min(-1) hyperinsulinemic-euglycemic clamp) and measured plasma PAI-1 antigen and activities and tissue plasminogen activator (tPA) in 14 patients with type 2 diabetes and 20 normal control subjects (10 lean, 10 obese) before and after 3 months of treatment with troglitazone (600 mg/day). At baseline, plasma PAI-1 antigen levels after an overnight fast were significantly higher in the obese (33.5 +/- 4.7 microg/l) and type 2 diabetic subjects (54.9 +/- 6.3 microg/l) than in the lean control subjects (16.3 +/- 3.2 microg/l; P < 0.01 and P < 0.001, respectively). Troglitazone decreased plasma PAI-1 antigen concentrations in the diabetic patients (36.8 +/- 5.0 microg/l; P < 0.001 vs. baseline), but the reduction in the obese subjects did not reach statistical significance (baseline, 33.5 +/- 4.7; after troglitazone, 25.6 +/- 5.2 microg/l). Changes in plasma PAI-1 activity paralleled those of PAI-1 antigen. The extent of the reduction in plasma PAI-1 antigen concentrations in the diabetic patients after troglitazone correlated with the reductions in fasting plasma insulin (r = 0.60, P < 0.05), nonesterified fatty acid (r = 0.63, P < 0.02), and glucose concentrations (r = 0.64, P < 0.02) but not with the improvement in glucose disposal rates during the glucose clamps. Three nonresponders to troglitazone with respect to effects on insulin sensitivity and fasting glucose and insulin levels also had no reduction in circulating PAI-1. In conclusion, troglitazone enhances fibrinolytic system activity in insulin resistant type 2 diabetic patients. This effect appears to be intimately linked to its potential to lower plasma insulin levels and improve glycemic control through its peripheral tissue insulin-sensitizing effects. PMID- 10871203 TI - Increase in fat oxidation on a high-fat diet is accompanied by an increase in triglyceride-derived fatty acid oxidation. AB - The aim of this study is to investigate the mechanism behind the slow increase in fat oxidation on a high-fat diet. Therefore, we determined 24-h substrate oxidation using respiration chambers and the rate of appearance and oxidation of plasma-derived fatty acids in seven healthy nonobese men (age 23 +/- 2 years, height 1.85 +/- 0.03 m, weight 70.4 +/- 2.3 kg, % body fat 13 +/- 1). Before testing, they consumed a low-fat diet (30% fat, 55% carbohydrate) at home for 3 days. Measurements were performed after 1 day consumption of either a low-fat diet (LF), a high-fat diet (HF1, 60% fat, 25% carbohydrate), or a high-fat diet preceded by a glycogen-lowering exercise test (HF1+EX), and after 7 days on a high-fat diet (HF7). After an overnight fast, an infusion of [U-13C]palmitate (0.00806 micromol x min(-1) x kg(-1)) was started and continued for 2 h at rest followed by 1 h of exercise at 50% of maximal power output (Wmax). Whole-body fat oxidation was measured using indirect calorimetry, and plasma-derived fatty acid oxidation was evaluated by measuring breath 13CO2 enrichment and corrected with the acetate recovery factor. Twenty-four-hour fat oxidation gradually increased on the high-fat diet. Both at rest and during exercise, there was no change in rate of appearance of fatty acids and plasma-derived fatty acid oxidation. Triglyceride-derived fatty acid oxidation tended to be higher after 7 days of high-fat diet at rest (P < 0.07). This difference was significant during exercise (P < 0.05). In conclusion, the results from this study suggest that triglyceride derived fatty acid oxidation (VLDL or intramuscular triglycerides) plays a role in the increase in fat oxidation on a high-fat diet, but plasma-derived fatty acids remain the major source for fat oxidation. PMID- 10871204 TI - Use of a novel impermeable biotinylated photolabeling reagent to assess insulin- and hypoxia-stimulated cell surface GLUT4 content in skeletal muscle from type 2 diabetic patients. AB - Cell surface GLUT4 levels in skeletal muscle from nine type 2 diabetic subjects and nine healthy control subjects have been assessed by a new technique that involves the use of a biotinylated photo-affinity label. A profound impairment in GLUT4 translocation to the skeletal muscle cell surface in response to insulin was observed in type 2 diabetic patients. Levels of insulin-stimulated cell surface GLUT4 above basal in type 2 diabetic patients were only approximately 10% of those observed in healthy subjects. The magnitude of the defect in GLUT4 translocation in type 2 diabetic patients was greater than that observed for glucose transport activity, which was approximately 50% of that in healthy subjects. Reduced GLUT4 translocation is therefore a major contributor to the impaired glucose transport activity in skeletal muscle from type 2 diabetic subjects. When a marked impairment in GLUT4 translocation occurs, the contribution of other transporters to transport activity becomes apparent. In response to hypoxia, marked reductions in skeletal muscle cell surface GLUT4 levels were also observed in type 2 diabetic patients. Therefore, a defect in a common late stage in signal transduction and/or a direct impairment in the GLUT4 translocation process accounts for reduced glucose transport in type 2 diabetic patients. PMID- 10871205 TI - Mechanical stretch-induced fibronectin and transforming growth factor-beta1 production in human mesangial cells is p38 mitogen-activated protein kinase dependent. AB - Hemodynamic abnormalities are important in the pathogenesis of the excess mesangial matrix deposition of diabetic and other glomerulopathies. p38-Mitogen activated protein (MAP) kinase, an important intracellular signaling molecule, is activated in the glomeruli of diabetic rats. We studied, in human mesangial cells, the effect of stretch on p38 MAP kinase activation and the role of p38 MAP kinase in stretch-induced fibronectin and transforming growth factor-beta1 (TGF beta1) accumulation. p38 MAP kinase was activated by stretch in a rapid (11-fold increase at 30 min, P < 0.001) and sustained manner (3-fold increase at 33 h, P < 0.001); this activation was mediated by protein kinase C (PKC). Stretch-induced fibronectin and TGF-beta1 protein levels were completely abolished (100% inhibition, P < 0.001; and 92% inhibition, P < 0.01, respectively) by SB203580, a specific p38 MAP kinase inhibitor. At 33 h, TGF-beta1 blockade did not affect stretch-induced fibronectin production, but partially prevented stretch-induced p38 MAP kinase activation (59% inhibition, P < 0.05). TGF-beta1 induced fibronectin accumulation after 72 h of exposure via a p38 MAP kinase-dependent mechanism (30% increase over control, P < 0.01). In human mesangial cells, stretch activates, via a PKC-dependent mechanism, p38 MAP kinase, which independently induces TGF-beta1 and fibronectin. In turn, TGF-beta1 contributes to maintaining late p38 MAP kinase activation, which perpetuates fibronectin accumulation. PMID- 10871206 TI - Endothelin receptor blockade prevents augmented extracellular matrix component mRNA expression and capillary basement membrane thickening in the retina of diabetic and galactose-fed rats. AB - Endothelins (ETs) are a family of vasoactive peptides that have mitogenic properties and have also been associated with altered long-term nuclear signaling. We have previously shown that mRNA levels for ET-1, ET-3, and their receptors are upregulated under hyperhexosemic conditions. In this study, an endothelin antagonist was used to assess the effects of endothelin blockage on the production of two basement membrane transcripts, fibronectin and collagen alpha1 (IV). The microvascular basement membranes were analyzed using ultrastructural morphometry. Streptozotocin-induced diabetic rats, galactose-fed rats (30% galactose in diet), and nondiabetic, non-galactose-fed control rats were studied after 1-month and 6-month follow-up. Simultaneously, similar animal groups were treated with a general ET receptor blocker (bosentan, 100 mg x kg(-1) x day(-1)) and investigated. Semiquantitative reverse transcription-polymerase chain reaction for fibronectin and collagen alpha1 (IV) was conducted after 1 month of follow-up with comparison to beta-actin housekeeping gene using slot blot hybridization and densitometry. Basement membrane thickness was assessed after 6 months of follow-up in diabetic rats, using the orthogonal intercept method. After 1 month of follow-up, increased fibronectin and collagen alpha1 (IV) mRNA were present in the retina of diabetic and galactosemic animals, and the bosentan-treated groups exhibited mRNA levels similar to the control animals. After 6 months of follow-up, diabetes and galactose feeding induced basement membrane thickening, which was partially prevented by bosentan treatment. The above findings indicate that increased production of extracellular matrix proteins leading to thickening of microvascular basement membrane, secondary to hyperhexosemia, may be mediated via augmented ET production. PMID- 10871207 TI - Molecular basis and characterization of the hyperinsulinism/hyperammonemia syndrome: predominance of mutations in exons 11 and 12 of the glutamate dehydrogenase gene. HI/HA Contributing Investigators. AB - Glutamate dehydrogenase (GDH) is allosterically activated by the amino acid leucine to mediate protein stimulation of insulin secretion. Children with the hyperinsulinism/hyperammonemia (HI/HA) syndrome have symptomatic hypoglycemia plus persistent elevations of plasma ammonium. We have reported that HI/HA may be caused by dominant mutations of GDH that lie in a unique allosteric domain that is encoded within GDH exons 11 and 12. To examine the frequency of mutations in this domain, we screened genomic DNA from 48 unrelated cases with the HI/HA syndrome for exon 11 and 12 mutations in GDH. Twenty-five (52%) had mutations in these exons; 74% of the mutations were sporadic. Clinical manifestations included normal birth weight, late onset of hypoglycemia, diazoxide responsiveness, and protein-sensitive hypoglycemia. Enzymatic studies of lymphoblast GDH in seven of the mutations showed that all had reduced sensitivity to inhibition with GTP, consistent with an increase in enzyme activity. Mutations had little or no effect on enzyme responses to positive allosteric effectors, such as ADP or leucine. Based on the three-dimensional structure of GDH, the mutations may function by impairing the binding of an inhibitory GTP to a domain responsible for the allosteric and cooperativity properties of GDH. PMID- 10871208 TI - Viewpoint on ISA TR84.0.02--simplified methods and fault tree analysis. AB - ANSI/ISA-S84.01-1996 and IEC 61508 require the establishment of a safety integrity level for any safety instrumented system or safety related system used to mitigate risk. Each stage of design, operation, maintenance, and testing is judged against this safety integrity level. Quantitative techniques can be used to verify whether the safety integrity level is met. ISA-dTR84.0.02 is a technical report under development by ISA, which discusses how to apply quantitative analysis techniques to safety instrumented systems. This paper discusses two of those techniques: (1) Simplified equations and (2) Fault tree analysis. PMID- 10871209 TI - Static and dynamic stresses of practical thermowells AB - A flexible beam model is proposed for calculating the flow induced stresses and application limits of common thermowell designs. It is intended to replace traditional selection methods with one which emphasizes mechanical integrity and process containment. PMID- 10871210 TI - Simulation modeling of nuclear steam generator water level process--a case study AB - Simulation modeling of the nuclear steam generator (SG) water level process in Qinshan Nuclear Power Plant (QNPP) is described in this paper. A practical methodology was adopted so that the model is both simple and accurate for control engineering implementation. The structure of the model is in the form of a transfer function, which was determined based on first-principles analysis and expert experience. The parameters of the model were obtained by taking advantage of the recorded historical response curves under the existing closed-loop control system. The results of process dimensional data verification and experimental tests demonstrate that the simulation model depicts the main dynamic characteristics of the SG water level process and is in accordance with the field recorded response curves. The model has been successfully applied to the design and test of an advanced digital feedwater control system in QNPP. PMID- 10871211 TI - Workflow management with systems approach: anticipated and ad-hoc workflow for scientific applications. AB - While workflow management has mostly been considered in business applications so far, the focus of the Workflow Management with Systems Approach (WOMASA) is on scientific applications such as geoprocessing, molecular biology, or laboratory environments. In particular, WOMASA aims at flexible and platform-indepented workflow support, with respect to both specification and execution of workflows. It turns out that the modeling and execution of workflows in traditional and in scientific applications exhibit significant differences. In particular, the need for dynamic modifications of workflow models while workflows are running is an important feature in scientific applications. The conceptual design and functionality of a WOMASA prototype is outlined, in particular that of its core workflow engine, and it is shown how the requirements of flexibility in modeling and executing workflows, imposed by scientific applications, are met. PMID- 10871212 TI - Fiber Bragg grating temperature sensor for practical use AB - Fiber Bragg grating (FBG) is a promising measurement technology for future sensor system applications. Little attention has been given to the FBG sensor housing, however it is important that the FBG is embedded in a conventional electrical sensor housing so that it may be installed easily in instrumentation systems. We have experimentally fabricated a practical pass-through type FBG temperature sensor which is embedded in the conventional thermocouple housing. A small radius U-turn fiber spliced to the FBG is placed inside the top of a stainless sheath. Employing a high numerical aperture fiber with polyamide recoating, we have determined that the bending loss of the small radius U-turn fiber and heat resistance is less than 0.1 dB and up to 250 degrees C respectively. The wavelength shift of this sensor due to temperature change is 10.3 pm/degrees C equal to that of general FBG. Consequently the practical use of this sensor has been confirmed. PMID- 10871213 TI - Auto-calibration and -compensation of a capacitive pressure sensor using multilayer perceptrons AB - Using multilayer perceptrons (MLPs), a smart model for a capacitive pressure sensor (CPS) is proposed. When the ambient temperature changes, the nonlinear response characteristics of a CPS may vary widely. Under such conditions, calibration of the sensor and compensation of the nonlinear sensor characteristics to obtain correct readout becomes a difficult task. The proposed MLP model can provide automatic nonlinear compensation and calibration of the CPS characteristics. A microcontroller unit (MCU)-based implementation scheme for this model is also considered. Simulation results show that this model can estimate the pressure with a maximum full-scale error of +/- 1% over a variation of temperature from -50 to 150 degrees C. PMID- 10871214 TI - Controller designs for constant cutting force turning machine control AB - A simulation study of a constant cutting force metal turning process is investigated. The process is a challenging control problem due to its nonlinear and time varying dynamics. Simulated implementations of PID, adaptive and non adaptive sliding mode and model reference adaptive controllers were developed. Tests of the closed loop systems were performed for a range of cutting conditions including specific machined part contours that are presented here to verify the force tracking capability and flexibility of each control scheme. Results indicate that careful design and use of a fixed-gain sliding mode controller with output feedback can give roughly equivalent performance to that of more complex adaptive controllers. PMID- 10871215 TI - Sliding mode control: an approach to regulate nonlinear chemical processes AB - A new approach for the design of sliding mode controllers based on a first-order plus-deadtime model of the process, is developed. This approach results in a fixed structure controller with a set of tuning equations as a function of the characteristic parameters of the model. The controller performance is judged by simulations on two nonlinear chemical processes. PMID- 10871216 TI - Robust on-line relay automatic tuning of PID control systems AB - In this paper, a robust on-line relay automatic tuning method for PID control systems is developed which expand on the application domain of Astrom's renowned relay autotuning method. In the proposed configuration, a relay is applied to an inner loop of a controller-stabilised process in the usual manner. Using the induced limit cycle oscillations from the closed-loop system, the controller settings may be re-tuned non-iteratively to achieve enhanced performance without disrupting closed-loop control. Two control tuning methodologies are developed -- a direct and an indirect method based on an explicit process model. Simulation examples and a real-time experiment are provided to illustrate the practical appeal and potential advantages of the proposed method over the basic one. PMID- 10871217 TI - Simultaneous online automatic tuning of cascade control for open loop stable processes AB - In this paper, we present a new simultaneous online automatic tuning method for cascade control using a relay feedback approach. Departing from the traditional approach towards tuning of cascade control systems where the secondary and primary loops are tuned in strict sequence, the proposed approach is to carry out the entire tuning process in one experiment. For ease of practical applications, the entire procedure of controller design may be automated and carried out online. A direct controller tuning approach to tune the controllers is proposed here. Robustness analysis of the new cascade control design is further carried out in the paper by drawing on existing results for SISO feedback systems. Simulations are provided to illustrate the applicability and effectiveness of both the online autotuning approach and the new cascade control design. PMID- 10871218 TI - LQG controller design using GUI: application to antennas and radio-telescopes AB - The Linear Quadratic Gaussian (LQG) algorithm has been used to control the JPL's beam wave-guide, and 70-m antennas. This algorithm significantly improves tracking precision in a wind disturbed environment. Based on this algorithm and the implementation experience a Matlab based Graphical User Interface (GUI) was developed to design the LQG controllers applicable to antennas and radiotelescopes. The GUI is described in this paper. It consists of two parts the basic LQG design and the fine-tuning of the basic design using a constrained optimization algorithm. The presented GUI was developed to simplify the design process, to make the design process user-friendly, and to enable design of an LQG controller for one with a limited control engineering background. The user is asked to manipulate the GUI sliders and radio buttons to watch the antenna performance. Simple rules are given at the GUI display. PMID- 10871219 TI - Control valves and process variability AB - A theoretical analysis is developed to explain some experimental results for a flow loop from the literature. Process variability is shown to be dependent on the disturbance or noise parameters, the closed loop time constant, the loop dead time and the control valve dead band. The results can be extended to other process variables. PMID- 10871220 TI - Integration of advanced process control and full-scale dynamic simulation AB - In the process control industry, multivariable model predictive controller and dynamic simulation for operator training are usually available in separate packages. It is very difficult for the operators and plant engineers to find good tools for them to get trained in multivariable advanced process control. This paper presents a system, which integrates the advanced process control and full scale dynamic simulation. The advanced process control uses multivariable model predictive control techniques. The model used in the predictive control algorithms is generated from the dynamic simulated process. The advanced process controller can control the simulated plant directly, or through a DCS system to control the simulated plant. The combined system provides an excellent environment for training operators in process operation with multivariable advanced process control. The same environment is also very useful for engineers in designing and tuning the advanced process controllers, and in testing communication between the advanced process controller and the DCS systems, or the other type of process control systems. PMID- 10871221 TI - Foundation fieldbus economics comparison AB - This paper will provide a life cycle cost comparison between installations of three different control system configurations. 1. Conventional analogue instruments. 2. HART protocol instruments with a parallel asset management system. 3. Foundation fieldbus installation. Engineering, procurement, construction, commissioning and ongoing maintenance costs will be included as part of the analysis. Data for the analysis includes equipment from multiple manufacturers so a range of expected expenses will also be provided to indicate the sensitivity of the economic calculations to supplier. All calculations will be based on the median equipment values. PMID- 10871222 TI - T-plate fixation of distal radial closing wedge osteotomies for treatment of angular limb deformities in 18 dogs. AB - OBJECTIVE: To describe the surgical technique and clinical results of treatment for forelimb angular limb deformities, secondary to premature distal radial or ulnar physeal closure, by using T-plate fixation of a distal radial closing wedge osteotomy in 18 dogs. STUDY DESIGN: Retrospective clinical study. SAMPLE POPULATION: 18 client-owned dogs. METHODS: The medical records of 18 dogs that underwent a distal radial closing wedge osteotomy with T-plate fixation for correction of a forelimb angular limb deformity were reviewed. Small pins (Kirschner wires) were used to obtain the appropriate alignment of the antebrachiocarpal and elbow joints and proper limb orientation. In-hospital follow-up evaluation was obtained at the time fracture healing was observed radiographically. Further long-term follow-up was obtained by owner interview. RESULTS: Osteotomy sites were radiographically healed within 4 to 12 weeks (mean, 8 weeks) after surgery in the 14 dogs that returned for in-hospital follow-up. Limb function was graded as good or excellent in all dogs. Long-term follow-up by owner interview rated limb function and cosmetic appearance as good to excellent in all dogs. Plate removal was necessary in one dog 7 months after surgery because of osteopenia in the radius. CONCLUSION: This surgical technique was considered successful in the treatment of angular limb deformities in all dogs. A good to excellent prognosis is to be expected with this technique, with minimal complications. CLINICAL RELEVANCE: The use of a T-plate for the correction of angular limb deformities has not been previously described in the literature. This technique permits accurate correction of the angular limb deformity and minimizes postoperative complications. PMID- 10871223 TI - Comparison of bone healing by demineralized bone matrix and autogenous cancellous bone in horses. AB - OBJECTIVE: The purpose of this study was to compare bone healing induced by equine demineralized bone matrix (DBM) to autogenous cancellous bone graft (ACB) or no graft (control) in a rib-defect model in horses. STUDY DESIGN: The osteogenic properties of ACB and DBM were evaluated in bilateral 19-mm circular defects created in the outer cortex of the 6th and 8th ribs of each horse. ANIMALS OR SAMPLE POPULATION: Eight mature horses. METHODS: Three rib defects in each horse were randomly treated with each of the 3 treatment groups, and the fourth rib defect received a random treatment. Rib sections, including the defects, were harvested 56 days after implantation and examined for bone mineral density, percent ash and calcium and graded for signs of radiographic and histological healing. RESULTS: All ribs were fractured at the defect site and were classified as nonunion fractures 56 days after implantation. There were no significant differences among groups in bone mineral density and signs of radiographic or histological healing. There was an increased volume of bone in control and ACB-treated sites compared with DBM-treated sites. Rib defects treated with ACB were significantly higher in percent ash and calcium than those treated with DBM. DBM elicited no inflammatory reaction, and remodeling occurred around the periphery and within vascular channels of the decalcified particles. CONCLUSION: DBM particles remodel from the periphery, which may explain the significantly lower percent ash, calcium, and bone when compared with ACB, because 2- to 4-microL pieces of DBM may act as space-occupying masses until completely mineralized. There was no evidence of enhanced healing associated with the use of DBM in this model. CLINICAL RELEVANCE: Particles of 2 to 4 mm DBM should not be used as an aid to fracture repair because particles of this size interfere with normal mineralization. However, our model of nonunion fracture healing may be useful in future studies. PMID- 10871224 TI - Femoral medullary infarction secondary to canine total hip arthroplasty. AB - OBJECTIVE: To evaluate the prevalence of femoral intramedullary infarction after total hip arthroplasty (THA) and to determine whether any specific femoral morphology predisposes to bone infarction. STUDY DESIGN: Retrospective clinical study. SAMPLE POPULATION: All dogs from our hospital population undergoing THA between 1984 and 1997 with radiographic follow-up available at 1 year or more postoperatively. METHODS: A case control study was conducted within the THA group to determine risk factors predisposing to femoral infarction after THA. Medical records and radiographs were reviewed. Data were collected on clinical parameters, femoral morphology, prosthesis, and bone changes. Radiographic diagnosis was confirmed using histopathology in 11 femora. Radiographs of 50 age matched control dogs weighing more than 20 kg with coxofemoral degenerative joint disease were randomly chosen to determine the prevalence of bone infarction in nonoperated dogs. RESULTS: Ninety-one dogs with 110 THA were included in the study. Fifteen of the 110 femora with THA had radiographic evidence of infarction (14%). Infarction was not present in any femora in the control group. There was no significant difference in the prevalence of infarction between dogs that received cemented or uncemented prostheses. Clinical signs were not reported in any patient that developed femoral infarction. Young age (P = .03) and a distance between the greater trochanter and nutrient foramen greater than 79 mm (P = .008) predisposed dogs to femoral infarction. Over time, three infarcts decreased in size radiographically, five remained unchanged, and three expanded. An osteosarcoma developed at the site of a bone infarct in one dog. CONCLUSION: Femoral intramedullary infarction occurred in 15 of 110 THA. Young age at the time of THA and a greater distance between the greater trochanter and the nutrient foramen predisposed to infarction. CLINICAL RELEVANCE: Intramedullary infarction occurs after canine THA. These bone infarcts do not appear to cause clinical signs; however, they may present a diagnostic challenge. Malignant transformation could potentially result from medullary infarction. PMID- 10871225 TI - Fracture healing after stabilization with intramedullary xenograft cortical bone pins: a study in pigeons. AB - OBJECTIVE: To investigate the effectiveness of intramedullary xenograft cortical bone pins compared with stainless steel Kirschner wire for the repair of a standardized avian humeral fracture. STUDY DESIGN: Prospective randomized study. SAMPLE POPULATION: Thirty mature pigeons (Columba livia). METHODS: Birds were randomly assigned to 3 groups. Transverse mid-diaphyseal humeral fractures were created in 1 humerus in each bird. Fractures were stabilized with intramedullary ostrich or canine xenograft cortical bone pins or Kirschner wire. Radiographic, histological, and biomechanical assessments were used to compare fracture healing 6 weeks after fracture stabilization. The contralateral humerus of each bird was used as a control. RESULTS: All fractures healed regardless of intramedullary pin type. There were no statistically significant biomechanical differences among groups or within groups. Xenograft cortical bone pins induced a mononuclear inflammatory reaction that did not impair bone healing. Bones stabilized with intramedullary cortical bone pins had more periosteal callus and inflammation at the fracture site than bones stabilized with stainless steel Kirschner wires. CONCLUSIONS: Intramedullary xenograft cortical bone pins, derived from mammalian or avian sources, appear to represent an alternative for the repair of avian humeral fractures. CLINICAL RELEVANCE: Intramedullary xenograft cortical bone pins are biodegradable and may reduce the need for additional surgery to remove implants after fracture healing. PMID- 10871227 TI - Evaluation of samarium-153 for synovectomy in an osteochondral fragment-induced model of synovitis in horses. AB - OBJECTIVE: To determine the effects of intraarticular administration of Samarium 153 (153Sm) bound to hydroxyapatite microspheres (153SmM) on an osteochondral chip-induced synovitis. STUDY DESIGN: Sixty days after implantation of autogenous osteochondral fragments in the middle carpal and metacarpophalangeal joints, 153SmM was administered into 1 joint of each type. The contralateral joints were used as untreated controls. ANIMALS OR SAMPLE POPULATION: Fifteen horses without preexisting joint disease were randomly divided into 2 groups (7 in the carpal group, 8 in the metacarpophalangeal group). METHODS: Horses had osteochondral fragments that were harvested from the lateral ridge of the trochlea of the talus and implanted bilaterally into a middle carpal joint and a metacarpophalangeal joint; the opposite joint type served as a control. Sixty days later, 10 to 15 mCi of 153SmM (20 to 50 microm diam) was injected into the fragment-implanted joints. Three horses were treated with nonradioactive hydroxyapatite fragments. Horses were examined clinically until they were killed 14 or 30 days later. Control and treated joints were examined grossly and microscopically to determine the effects of 153SmM on synovial membrane and cartilage. RESULTS: Intraarticular 153SmM caused a transient flare with lameness, effusion, and edema for 48 to 72 hours. Implanted osteochondral chips induced a synovitis characterized by variable degrees of joint damage and synovial infiltrate. Use of 153SmM resulted in synovectomy of variable depth and extent. CONCLUSIONS: Intraarticular 153SmM may be a useful method for synovectomy of inflamed synovial membrane. CLINICAL RELEVANCE: With further testing, radioactive pharmaceuticals might become useful clinical treatments for persistent synovitis not responsive to conventional techniques. PMID- 10871226 TI - In vitro evaluation of antibiotic elution from polymethylmethacrylate (PMMA) and mechanical assessment of antibiotic-PMMA composites. AB - OBJECTIVE: To determine whether different methods of sterilization of antibiotic vials or the heat of polymerization altered the antimicrobial activity or mechanical properties of antibiotic/polymethylmethacrylate (PMMA) composites when compared to antibiotic-free PMMA. STUDY DESIGN: In vitro study. METHODS: Steam sterilized, gas-sterilized, and non-sterilized 1 gram vials of cefazolin and injectable gentamicin sulfate (high and low doses) were mixed with PMMA to prepare composites for antibiotic elution evaluation, compression, and elongation testing. Blocks of PMMA that contained antibiotic were assayed for antibacterial activity using an agar gel diffusion method or were placed in phosphate buffered saline (PBS) to assess elution of antibiotic. Phosphate buffered saline samples from steam-sterilized cefazolin and high-dose gentamicin groups were assayed on days 1, 2, 5, and 9 for cefazolin or gentamicin concentration by high-pressure liquid chromatography or fluorescent polarization immunoassay, respectively. RESULTS: PMMA blocks containing antibiotic inhibited bacterial growth of Staphylococcus aureus 25923 for an average of 9 days. Cefazolin and gentamicin concentration in PBS decreased dramatically after the first 24 hours, but remained above minimum inhibitory concentration (MIC) throughout the experiment for all groups except low-dose gentamicin. Compressive strength of plugs made from plain cement and plugs made from PMMA mixed with untreated and steam sterilized cefazolin was similar, but was significantly different from the other groups. There appeared to be an inverse relationship between compressive strength and elongation. CONCLUSION: PMMA/antibiotic composites inhibited bacterial growth for 7 to 10 days. Compressive strength was affected by different additions of antibiotic. CLINICAL RELEVANCE: Bacteria introduced during a surgical procedure may be inhibited by elution of antibiotic from PMMA at the time of contamination. PMID- 10871228 TI - Esophageal obstruction caused by a left aortic arch and an anomalous right patent ductus arteriosus in two German Shepherd littermates. AB - OBJECTIVE: To present details of surgical management of an unusual vascular ring anomaly in two German Shepherd littermates. STUDY DESIGN: Clinical case report. STUDY POPULATION: Three-month-old intact male and female German Shepherd littermates. RESULTS: In each dog, the esophagus was obstructed by a vascular ring comprised of the left aortic arch, an anomalous patent right ductus arteriosus, and the pulmonary artery ventrally. Surgical treatment consisted of dividing and oversewing the patent right ductus arteriosus. Neither dog has clinical signs of esophageal disease one year after treatment. CONCLUSION AND CLINICAL RELEVANCE: This vascular ring anomaly should be considered a possibility in any young dog with esophageal obstruction and a machinery murmur. PMID- 10871229 TI - The hemodynamic effects of intrathecal oxytocin in normal dogs. AB - OBJECTIVE: To evaluate the hemodynamic effects produced by intrathecal administration of oxytocin in healthy isoflurane-anesthetized dogs. STUDY DESIGN: Prospective single-dose trial. ANIMAL POPULATION: Six healthy purpose-bred adult dogs weighing between 7.3 and 14.5 kg. METHODS: Dogs were anesthetized with isoflurane and instrumented. Oxytocin at a dosage of 1.6 microg/kg was administered intrathecally at the cisternal space at time 0. Hemodynamic data were recorded immediately before and at 1, 5, 15, 30, and 60 minutes after oxytocin administration. Statistical analysis included an analysis of variance (ANOVA) for repeated measures over time. A P < .05 was considered significant. RESULTS: Baseline values +/- standard error of the mean for heart rate, mean arterial pressure, central venous pressure, cardiac output, systemic vascular resistance, mean pulmonary arterial pressure, pulmonary arterial occlusion pressure, and pulmonary vascular resistance were 101 +/- 11 beats/minute, 76 +/- 7 mm Hg, 4 +/- 4 mm Hg, 1.9 +/- 0.7 L/min, 3834 +/- 2556 dynes x sec/cm5, 14 +/- 3 mm Hg, 4 +/- 2 mm Hg, and 430 +/- 201 dynes x sec/cm5, respectively. Variations from the baseline values were seen in all parameters after intrathecal oxytocin administration, but no statistically significant differences were found. CONCLUSION: The intrathecal injection of 1.6 microg/kg of oxytocin is associated with minimal hemodynamic effects during isoflurane anesthesia. CLINICAL RELEVANCE: This study revealed no clinically significant deleterious effects from the intrathecal administration of oxytocin, and investigations into its use as a perioperative analgesic are therefore warranted. PMID- 10871230 TI - A panoply of anabolic and catabolic mediators. PMID- 10871231 TI - Effects of inactivity and hormonal mediators on skeletal muscle during recovery from trauma. AB - Severe injury profoundly alters two important hormonal mediators of skeletal muscle. Cortisol production, and the subsequent effect on muscle catabolism, is immediately and persistently elevated. To the contrary, testosterone, a primary anabolic stimulus of skeletal muscle, is considerably suppressed with trauma. The result is a dramatically increased catabolic/anabolic hormonal profile that is further exacerbated by the prolonged inactivity that occurs with severe injury. These factors combine to produce a conspicuous loss of lean body mass throughout hospitalization. Emerging evidence suggests that one approach to ameliorating the loss of skeletal muscle nitrogen is restoration of the anabolic influence. The safe and effective normalization of testosterone concentrations after severe injury attenuates the loss of muscle protein. The retention of lean body mass will positively affect clinical and rehabilitative outcomes. PMID- 10871232 TI - Cancer cachexia: from experimental models to patient management. AB - Cachexia is frequently associated with advanced or terminal cancer states, but it can also develop early during the course of neoplastic disease. This syndrome, which is characterized by body weight loss and negative nitrogen balance, significantly affects patient survival and quality of life. Studies on experimental models have shown that a complex interplay of different factors, such as anorexia, classical hormones, cytokines and other less well defined factors, concur in causing tissue wasting. On the basis of these results, it has been possible to prevent the onset of experimental cachexia by targeting therapeutic interventions at the underlying metabolic perturbations. Anticytokine treatments, either acting centrally or peripherally, have received particular attention, and are currently reaching the stage of clinical trials. PMID- 10871233 TI - Anabolic and catabolic mediators of intestinal protein turnover: a new experimental approach. AB - Studies on regulation of protein turnover in skeletal muscle have revealed the important contributions of protein synthesis and catabolism to tissue protein balance, and have identified a host of specific anabolic and catabolic stimuli and biochemical mechanisms that regulate these processes. This knowledge is critical to current efforts designed to promote anabolism and limit atrophy. Of the tissues with a potentially large contribution to whole-body amino acid metabolism, protein turnover of the intestine stands out as being poorly understood. The intestine is subject to complexities in regulation of its metabolism that are not apparent for other tissues. The study of intestinal protein turnover also entails some important technical challenges. We recently developed an in-situ experimental system for study of intestinal mucosal protein synthesis with the following unique features: multiple observations within an animal; controlled delivery of nutritional stimuli to the apical side, basolateral side, or both; and luminal delivery of tracer in a flooding dose for determination of protein synthesis. We have begun to use the system to test the specific roles of individual luminal nutrients in regulation of mucosal protein synthesis. We have also used protease gene expression as an index of potential regulation of catabolic pathways. PMID- 10871234 TI - Anti- and proinflammatory cytokines in the pathogenesis of tissue damage in Crohn's disease. AB - Crohn's disease is a chronic inflammatory bowel disease of unknown origin. The understanding of the pathogenesis of inflammatory bowel diseases has been greatly advanced by manipulations of the immune system in mice using targeted disruptions of genes that encode specific anti- and proinflammatory cytokines, as well as T cell subsets. The outcome of these experiments has implicated CD4+ lymphocytes and certain proinflammatory cytokines (tumour necrosis factor-alpha, interleukin 12) as playing a central role in the pathogenesis of mucosal inflammation in Crohn's disease. The present review focuses on these recent important immunological observations, and discusses several newly developed therapeutic strategies that are based either on blocking proinflammatory cytokines or on the administration of anti-inflammatory cytokines. PMID- 10871235 TI - Role of nitric oxide in wound healing. AB - Nitric oxide is a short-lived free radical, that is capable of multiple effects at the molecular, cellular, and physiologic levels. Over the past several years, nitric oxide has been proved to play an important role in the healing of various types of wounds. The present review examines some of the recently defined roles of nitric oxide in normal and pathologic healing. PMID- 10871236 TI - Anabolic and destructive mediators in osteoarthritis. AB - Osteoarthritis is a joint disease that is characterized by focal degradation of articular cartilage. In addition to the degeneration of articular cartilage, attempts at repair are found in the affected tissue. Cartilage cells (chondrocytes) play a key role, not only in the destructive process, but also in the repair response. It has become apparent that anabolic and catabolic mediators, released from chondrocytes themselves or from other joint cells, drive both destructive and repair activities in the osteoarthritic joint. PMID- 10871237 TI - Specialized feeding in hospital and home: computers, regulations and reimbursement issues. PMID- 10871238 TI - Central venous access in neonates through the peripheral route. AB - Central venous access is frequently used for monitoring and administration of drugs in the intensive care unit, and for administration of parenteral nutrition. The improving results of neonatal surgery have closely followed the evolution of neonatal intensive care and parenteral nutrition. Nutritional support by the parenteral route is required in the majority of cases for only a few weeks. Percutaneous sialastic long line introduced into a central vein through a peripheral venous puncture has emerged as one of the most popular techniques for delivery of parenteral nutrition in neonates. PMID- 10871239 TI - Technical aspects of feeding the disabled child. AB - Children with neurological impairment frequently have difficulties in consuming sufficient energy and other nutrients to maintain adequate nutritional status. Under-nutrition is a significant contributory factor to growth failure. Eating may be distressing and time-consuming for the child and carer. Aspiration of feeds is common and may predispose to chronic chest infections. Gastro oesophageal reflux is also common and may contribute to significant morbidity. This paper discusses some of the issues involved in the nutritional management of neurologically impaired children. PMID- 10871240 TI - Practicalities of nutrition support in chronic liver disease. AB - The underlying causative agent in the majority of patients with chronic liver disease is ethanol; the rest of the cases are largely viral in aetiology (hepatitis B or C viruses). Nutrition supplementation improves the quality of life and decreases morbidity in chronic liver disease, but evidence that it prolongs long-term survival is lacking. The situation is very different in chronic liver disease patients who receive a liver transplant, however, with better pretransplantation nutrition status predicting a distinctly improved survival after transplantation. PMID- 10871241 TI - Use of computers in long-term/home parenteral nutrition--a missed opportunity? AB - Desk-top microcomputers and total parenteral nutrition grew up together, and in their early days there was considerable progress in both camps. Since that time, the power of computing devices has increased dramatically, as has their ability to share information both between individual systems and worldwide through facilities such as the Internet and e-mail. Although there are some signs of continuing progress, to date there appears to be little evidence in peer-reviewed journals that this increased power is being utilized or that the vision of the pioneers in this area has been realized. PMID- 10871243 TI - Bibliography. Current world literature. Anabolic and catabolic signals. PMID- 10871242 TI - Economic aspects of paediatric home parenteral nutrition. AB - Children account for 15-20% of home parenteral nutrition programmes. Underlying irreversible intestinal diseases lead to potential indications for intestinal transplantation. Therefore, new controversial issues are timing for referring children for transplantation, and comparison between home parenteral nutrition and transplantation in terms of costs and quality of life. PMID- 10871244 TI - Bibliography. Current world literature. Technical aspects of nutritional support. PMID- 10871245 TI - Why treat early multiple sclerosis patients? AB - Class 1 clinical trials demonstrated that immunomodulatory treatments (interferon beta and glatiramer acetate) reduce the disease activity and the accumulation of disability in relapsing remitting multiple sclerosis. Moreover interferon beta-1b also had similar positive effects in secondary progressive multiple sclerosis. The magnitude of these clinical effects was modest, but the reduction of inflammatory activity, as revealed by magnetic resonance imaging, was marked. Converging evidence from new pathological studies and new magnetic resonance techniques, characterized by increased pathological specificity, has shown that already in the early phases of the disease inflammatory activity determines irreversible axonal damage. Moreover, the amount of inflammatory activity at the clinical presentation of the disease has some value in predicting long-term disability. Taken together, these data indicate that patients may benefit from early treatment; the positive results of the Controlled High Risk Subjects Avonex Multiple Sclerosis Prevention Study support this conclusion. PMID- 10871246 TI - Susceptibility to multiple sclerosis: interplay between genes and environment. AB - Multiple sclerosis is a complex trait of unknown etiology. Epidemiological data have shown that susceptibility to multiple sclerosis is determined by both genetic and environmental factors. It is unknown whether the clinical subcategories of multiple sclerosis are separate diseases with separate etiologies and causes. Recent theories of the pathogenesis of multiple sclerosis and candidate genes are discussed. Other potential nonchromosomal factors involved in multiple sclerosis susceptibility such as mitochondrial DNA and viral factors such as Chlamydia pneumoniae are reviewed. PMID- 10871247 TI - The value of new magnetic resonance techniques in multiple sclerosis. AB - Cell-specific magnetic resonance imaging, magnetization transfer imaging, proton magnetic resonance spectroscopy and diffusion-weighted imaging have all recently been applied to the study of multiple sclerosis. These techniques can give more accurate and pathologically specific estimates of the multiple sclerosis lesion burden than conventional magnetic resonance. Their extensive application should improve the understanding of the mechanisms leading to multiple sclerosis-related irreversible disability and help in the diagnostic work-out of patients with suspected multiple sclerosis. PMID- 10871248 TI - Neurophysiological investigations in multiple sclerosis. AB - The advent of magnetic resonance imaging techniques has greatly reduced the diagnostic value of neurophysiological tests, particularly evoked potentials, in multiple sclerosis patients, because of the higher sensitivity in revealing subclinical involvement of the central nervous system. Technical progress and new methods of investigating afferent and efferent nervous pathways would seem to increase the sensitivity in detecting neural dysfunction, but the 'clinical gain' is modest at best. More promising is the utilization of neurophysiological tests to quantify the severity of white matter involvement. Transversal and longitudinal studies have demonstrated good correlations between neurophysiological parameters and disability measures, indicating that a battery of neurophysiological tests could be useful in monitoring the disease evolution in single patients and as surrogate endpoints in clinical trials. Further studies are needed for a better definition of the applications of evoked potentials and other neurophysiological techniques. Finally, event-related potentials and advanced electroencephalogram techniques, such as coherence analysis, could provide useful information on the pathophysiology of cognitive dysfunction, so common in multiple sclerosis patients, and with a strong impact on the quality of life. PMID- 10871249 TI - The management of multiple sclerosis patients. AB - The management of individuals with multiple sclerosis must be considered under the following headings: administration and follow-up of adequate disease modifying treatment, symptomatic relief and neurorehabilitation. Neurorehabilitation deserves a four-step strategy: multidisciplinary assessment, identification of areas of potential functional improvement, setting of goals of short/long-term duration, and measurement of outcomes. The patient's perspective is important to evaluate through questionnaires about quality of life. Well organized disability services with multidisciplinary specialists are probably cost effective and efficient. Determining the actual economic impact of multiple sclerosis and defining the most cost-effective type of care for persons with multiple sclerosis is a need in all countries faced with the limitation of healthcare resources. In persons with multiple sclerosis the range of main symptoms includes the loss of mobility and spasticity, pain, tremor, abnormal eye movements, paroxysmal symptoms, bladder and bowel dysfunction, sexual disturbances, fatigue and depression. Current palliative treatments, which are reviewed, are partly successful depending on the type of symptoms under consideration. PMID- 10871250 TI - Migraine: imaging the aura. AB - We currently conceive of a migraine attack as originating in the brain. Triggers of an attack initiate a depolarizing neuroelectric and metabolic event likened to the spreading depression of Leao. This event activates the headache and associated features of the attack by mechanisms that remain to be determined, but appear to involve either peripheral trigeminovascular or brainstem pathways, or both. The excitability of cell membranes, perhaps partly genetically determined, is the brain's susceptibility to attacks. Factors that increase or decrease neuronal excitability constitute the threshold for triggering attacks. Using a model of visual stress-induced migraine or by studying spontaneous attacks and applying advanced imaging and neurophysiological methods, results have been obtained that support spreading neuronal inhibition as the basis of aura. This neuroelectric event is accompanied by hyperoxia of the brain, possibly associated with vasodilation. Evidence has also been obtained that the spreading cortical event can activate the subcortical centers possibly involved in nociception and associated symptoms of the migraine attack. Susceptibility to migraine attacks appears to be related to brain hyperexcitability. These newer techniques of functional neuroimaging have confirmed the primary neural basis of the migraine attack with secondary vascular changes, reconciling previous theories into a neurovascular mechanism. PMID- 10871251 TI - Chronic daily headache. AB - Primary chronic daily headache can be subclassified into disorders of short duration (<4 h/attack) including chronic cluster and disorders of long duration (> or =4 h/attack). Primary chronic daily headache disorders of long duration include chronic tension-type headache, chronic daily migraine (previously called transformed migraine), new daily persistent headache, and hemicrania continua. Four to 5% of the general population have primary chronic daily headache. Most chronic daily headache patients overuse analgesics or ergots. This article will consider recent insights into specific disorders, then psychiatric comorbidity, epidemiology, pathophysiology, and treatment. PMID- 10871252 TI - Tension-type headache: an update on mechanisms and treatment. AB - Tension-type headache represents one of the most costly diseases in modern society because of its very high prevalence. Very little research on this disease has actually been carried out, and knowledge about key pathophysiological issues such as the nature and site of the noxious stimulus is surprisingly limited. As a result of this and the lack of scientific interest from the medical field, treatment is widely non-specific, very often ineffective and consists mainly of simple analgesics. The only new strategy is the pericranial injection of botulinum toxin. If current progress in our understanding of the mechanisms of tension-type headache continues, this may lead to greater scientific interest and the development of more specific and more effective drugs in the future. PMID- 10871253 TI - Facial pain. AB - Facial pain is a debilitating disorder if left untreated. Too often patients are labelled as having psychopathology when face pain etiology is unclear. These patients are categorized as 'atypical', 'idiopathic' or 'psychogenic'. Idiopathic, when referring to a medical problem suggests that there is something unknown, and does not define the problem. The same applies to terms incorporating the word 'atypical'. It is postulated that the most commonly undiagnosed facial pain conditions include neuropathic and myofascial pains because their pathophysiologies are not well understood. Peripheral and central mechanisms associated with these disorders are used to provide an update of these frequently seen clinical conditions. PMID- 10871254 TI - Rasmussen's encephalitis: a challenge to neuroimmunology. PMID- 10871255 TI - Epstein-Barr virus and the nervous system. AB - The neurological complications of Epstein-Barr virus infection include viral meningitis, encephalitis and neuromuscular complications. The introduction of cerebrospinal fluid polymerase chain reaction for Epstein-Barr virus DNA has improved diagnosis of these conditions and of primary central nervous system lymphoma in acquired immune deficiency syndrome, and has enabled cerebrospinal fluid monitoring of therapy. Prognosis remains good for most Epstein-Barr virus related neurological complications; for primary central nervous system lymphoma in acquired immune deficiency syndrome the prognosis is still poor. PMID- 10871256 TI - Inflammatory neuropathies: update. AB - This review focuses on recent neuroimmunological findings in autoimmune inflammatory neuropathies. In Guillain-Barre syndrome and paraneoplastic neuropathy most current investigations are centred on the hypothesis of molecular mimicry. In chronic inflammatory demyelinating polyradiculoneuropathy and multifocal motor neuropathy the data on immunopathology are more fragmentary. Why and how patients with autoimmune inflammatory neuropathies raise an increased anti-self-reactivity and how this leads to disease remains a major challenge for future research. PMID- 10871257 TI - Myelitis. AB - Acute transverse myelitis (ATM) with moderate symptomatology and smaller multiple magnetic resonance imaging lesions is often caused by multiple sclerosis. Severe ATM with extensive magnetic resonance imaging lesions with or without associated meningitis often has a viral cause, particularly in the younger age groups, whereas vascular disorders may prevail among older patients. Previously, one had to rely on indirect evidence such as viral serology or viral identification in throat washings to confirm a diagnosis of myelitis. Thus, mycoplasma myelitis may occur coincident with a mycoplasma pneumonia. Viral myelitis is now often diagnosed by specific polymerase chain reaction of the cerebrospinal fluid, for echovirus, Coxsackie virus, mumps virus, herpes simplex virus or varicella-zoster virus, but an autoimmune component may still be important. An anterior horn syndrome may be produced by the tick-borne encephalomyelitis virus. Severe ATM may also be a postinfectious or postvaccinal disorder [i.e. a partial acute disseminated encephalomyelitis (ADEM)]. Neuromyelitis optica, a combination of severe myelitis and optic neuritis, is often a manifestation of ADEM or systemic lupus erythematosus. Many of these disorders are potentially treatable with specific antiviral agents or immunosuppression. 'Idiopathic' ATM is probably a consequence of inadequate examination and follow up. The differential diagnoses viral myelitis, multiple sclerosis, ADEM, neuromyelitis optica, spinal arteriovenous malformation and arteritis-should be considered and are usually identified by a rapid diagnostic work-up, leaving few ATM cases undiagnosed. PMID- 10871258 TI - Leprosy. AB - Leprosy is a unique infectious disease with a prolonged incubation period and a predilection for skin and nerves. The involvement of nerves by the primary infection as well as the immunologically mediated reversal reactions result in impairment of nerve function and severe disabilities. The introduction of the World Health Organization Multi Drug Therapy over the last two decades has produced dramatic changes in the management and control programmes of leprosy. A recent important contribution to the understanding of leprosy pathogenesis has been the elucidation of the molecular basis for the entry of Mycobacterium leprae into the Schwann cell and the peripheral nerve. Leprosy still remains the commonest cause of peripheral neuropathy in developing countries. PMID- 10871259 TI - Neurosarcoidosis. AB - Although there continues to be sporadic reporting of small numbers of patients with neurosarcoidosis, relatively little progress is being made in defining this condition in more detail, making it difficult for clinicians to be confident in their diagnoses and treatments. Although groups continue to use retrospective data to extrapolate diagnostic criteria, there is a need for a multinational collaboration to produce a prospective data set that would more adequately inform the diagnostic and therapeutic process for individual clinicians. PMID- 10871261 TI - Bibliography. Current world literature. Headache. PMID- 10871260 TI - Bibliography. Current world literature. Demyelinating diseases. PMID- 10871262 TI - Bibliography. Current world literature. Inflammatory diseases. PMID- 10871263 TI - From data to knowledge. PMID- 10871264 TI - Assessing the accuracy of prediction algorithms for classification: an overview. AB - We provide a unified overview of methods that currently are widely used to assess the accuracy of prediction algorithms, from raw percentages, quadratic error measures and other distances, and correlation coefficients, and to information theoretic measures such as relative entropy and mutual information. We briefly discuss the advantages and disadvantages of each approach. For classification tasks, we derive new learning algorithms for the design of prediction systems by directly optimising the correlation coefficient. We observe and prove several results relating sensitivity and specificity of optimal systems. While the principles are general, we illustrate the applicability on specific problems such as protein secondary structure and signal peptide prediction. PMID- 10871265 TI - Prediction of genetic structure in eukaryotic DNA using reference point logistic regression and sequence alignment. AB - MOTIVATION: Current software tools are moderately effective in predicting genetic structure (exons, introns, intergenic regions, and complete genes) from raw DNA sequence data. Improvements in accuracy and speed are needed to deal with the increasing volume of data from large scale sequencing projects. RESULTS: We present a two-stage computer program to predict genetic structure in eukaryotic DNA. The first stage makes use of a novel statistical technique, called reference point logistic (RPL) regression, to calculate scores for potential functional sites. These site scores are combined with interval content, length, and state scores, via a Generalized Hidden Markov Model, to determine a combined score for each possible parse of a given DNA sequence into exons, introns, and intergenic regions. An optimal parse is found using a dynamic programming algorithm. In the second stage, protein sequence alignment methods are applied to improve the accuracy of the initial parse. Computation in the first stage of the program is very fast (1 s on a 360 MHz CPU for a 16 kb sequence) and its predictive accuracy typically matches or exceeds the best results reported for other methods (Sensitivity = 0.93 and Specificity = 0.93 for the Burset/Guigotest set). Computation in the second stage is slower, but the final predictions are more accurate (Sn = 0.97, Sp = 0.97). The program (called GRPL) can handle partial, single, and multi-gene sequences. The program is also capable of predicting the genetic structure of vertebrate, invertebrate, and plant DNA with nearly equal accuracy. Statistical techniques have also been introduced to model the effects of varying C+G content in a continuous manner and to control overfitting of parameters for smaller training sets. AVAILABILITY: An academic implementation of GRPL, compiled for SUN workstations, is available by anonymous ftp from snipe.pharmacy. ualberta.ca/pub. The training and test sets used in this work, together with supplementary material, can be found at the same location. A commercial implementation is available as a component of GeneTool (BioTools Inc., http://biotools.com). PMID- 10871266 TI - PatSearch: a pattern matcher software that finds functional elements in nucleotide and protein sequences and assesses their statistical significance. AB - MOTIVATION: The identification of sequence patterns involved in gene regulation and expression is a major challenge in molecular biology. In this paper we describe a novel algorithm and the software for searching nucleotide and protein sequences for complex nucleotide patterns including potential secondary structure elements, also allowing for mismatches/mispairings below a user-fixed threshold, and assessing the statistical significance of their occurrence through a Markov chain simulation. RESULTS: The application of the proposed algorithm allowed the identification of some functional elements, such as the Iron Responsive Element, the Histone stem-loop structure and the Selenocysteine Insertion Sequence, located in the mRNA untranslated regions of post-transcriptionally regulated genes with the assessment of sensitivity and selectivity of the searching method. AVAILABILITY: A Web interface is available at: http://bigarea.area.ba.cnr.it:8000/EmbIT/Pats earch.html. PMID- 10871267 TI - GeneRAGE: a robust algorithm for sequence clustering and domain detection. AB - MOTIVATION: Efficient, accurate and automatic clustering of large protein sequence datasets, such as complete proteomes, into families, according to sequence similarity. Detection and correction of false positive and negative relationships with subsequent detection and resolution of multi-domain proteins. RESULTS: A new algorithm for the automatic clustering of protein sequence datasets has been developed. This algorithm represents all similarity relationships within the dataset in a binary matrix. Removal of false positives is achieved through subsequent symmetrification of the matrix using a Smith Waterman dynamic programming alignment algorithm. Detection of multi-domain protein families and further false positive relationships within the symmetrical matrix is achieved through iterative processing of matrix elements with successive rounds of Smith-Waterman dynamic programming alignments. Recursive single-linkage clustering of the corrected matrix allows efficient and accurate family representation for each protein in the dataset. Initial clusters containing multi-domain families, are split into their constituent clusters using the information obtained by the multi-domain detection step. This algorithm can hence quickly and accurately cluster large protein datasets into families. Problems due to the presence of multi-domain proteins are minimized, allowing more precise clustering information to be obtained automatically. AVAILABILITY: GeneRAGE (version 1.0) executable binaries for most platforms may be obtained from the authors on request. The system is available to academic users free of charge under license. PMID- 10871268 TI - RSDB: representative protein sequence databases have high information content. AB - MOTIVATION: Biological sequence databases are highly redundant for two main reasons: 1. various databanks keep redundant sequences with many identical and nearly identical sequences 2. natural sequences often have high sequence identities due to gene duplication. We wanted to know how many sequences can be removed before the databases start losing homology information. Can a database of sequences with mutual sequence identity of 50% or less provide us with the same amount of biological information as the original full database? RESULTS: Comparisons of nine representative sequence databases (RSDB) derived from full protein databanks showed that the information content of sequence databases is not linearly proportional to its size. An RSDB reduced to mutual sequence identity of around 50% (RSDB50) was equivalent to the original full database in terms of the effectiveness of homology searching. It was a third of the full database size which resulted in a six times faster iterative profile searching. The RSDBs are produced at different granularity for efficient homology searching. AVAILABILITY: All the RSDB files generated and the full analysis results are available through internet: ftp://ftp.ebi.ac. uk/pub/contrib/jong/RSDB/http://cyrah.e bi.ac.uk:1111/Proj/Bio/RSDB PMID- 10871269 TI - BIND--a data specification for storing and describing biomolecular interactions, molecular complexes and pathways. AB - MOTIVATION: Proteomics is gearing up towards high-throughput methods for identifying and characterizing all of the proteins, protein domains and protein interactions in a cell and will eventually create more recorded biological information than the Human Genome Project. Each protein expressed in a cell can interact with various other proteins and molecules in the course of its function. A standard data specification is required that can describe and store this information in all its detail and allow efficient cross-platform transfer of data. A complete specification must be the basis for any database or tool for managing and analysing this information. RESULTS: We have defined a complete data specification in ASN.1 that can describe information about biomolecular interactions, complexes and pathways. Our group is using this data specification in our database, the Biomolecular Interaction Network Database (BIND). An interaction record is based on the interaction between two objects. An object can be a protein, DNA, RNA, ligand, molecular complex or an interaction. Interaction description encompasses cellular location, experimental conditions used to observe the interaction, conserved sequence, molecular location, chemical action, kinetics, thermodynamics, and chemical state. Molecular complexes are defined as collections of more than two interactions that form a complex, with extra descriptive information such as complex topology. Pathways are defined as collections of more than two interactions that form a pathway, with additional descriptive information such as cell cycle stage. A request for proposal of a human readable flat-file format that mirrors the BIND data specification is also tendered for interested parties. AVAILABILITY: The ASN.1 data specification for biomolecular interaction, molecular complex and pathway data is available at ftp://bioinfo.mshri.on.ca/pub/BIND/Spec/bind.asn. An interactive browser for this document is available through our homepage at http://bioinfo.mshri.on.ca/BIND/asn browser/. PMID- 10871270 TI - A homolog of mammalian antizyme is present in fission yeast Schizosaccharomyces pombe but not detected in budding yeast Saccharomyces cerevisiae. AB - MOTIVATION: The antizymes (AZ) are proteins that regulate cellular polyamine pools in metazoa. To search for remote homologs in single-celled eukaryotes, we used computer software based on hidden Markov models. The most divergent homolog detected was that of the fission yeast Schizosaccharomyces pombe. Sequence identities between S.POMBE: AZ and known AZs are as low as 18-22% in the most conserved C-terminal regions. The authenticity of the S.POMBE: AZ is validated by the presence of a conserved nucleotide sequence that, in metazoa, promotes a +1 programmed ribosomal frameshift required for AZ expression. However, no homolog was detected in the completed genome of the budding yeast Saccharomyces cerevisiae. Procedural details and supplementary information can be found at http://itsa.ucsf.edu/ approximately czhu/AZ. PMID- 10871271 TI - GESTALT: a workbench for automatic integration and visualization of large-scale genomic sequence analyses. AB - SUMMARY: The GESTALT Workbench is a WWW-based tool for genomic sequence analysis, comparison and annotation, with strong emphasis on visualization. GESTALT integrates graphically the output of diverse sequence analysis algorithms producing an information-rich, interactive genomic map. AVAILABILITY: The GESTALT Workbench, as well as a more detailed description, are available at http://bioinfo. weizmann.ac.il/GESTALT/. PMID- 10871272 TI - Viral Genome DataBase: storing and analyzing genes and proteins from complete viral genomes. AB - SUMMARY: The Viral Genome DataBase (VGDB) contains detailed information of the genes and predicted protein sequences from 15 completely sequenced genomes of large (&100 kb) viruses (2847 genes). The data that is stored includes DNA sequence, protein sequence, GenBank and user-entered notes, molecular weight (MW), isoelectric point (pI), amino acid content, A + T%, nucleotide frequency, dinucleotide frequency and codon use. The VGDB is a mySQL database with a user friendly JAVA GUI. Results of queries can be easily sorted by any of the individual parameters. AVAILABILITY: The software and additional figures and information are available at http://athena.bioc.uvic.ca/genomes/index.html . PMID- 10871273 TI - RadCon: phylogenetic tree comparison and consensus. AB - SUMMARY: RadCon is a Macintosh program for manipulating and analysing phylogenetic trees. The program can determine the Cladistic Information Content of individual trees, the stability of leaves across a set of bootstrap trees, produce the strict basic Reduced Cladistic Consensus profile of a set of trees and convert a set of trees into its matrix representation for supertree construction. AVAILABILITY: The program is free and available at http://taxonomy.zoology.gla.ac.uk/ approximately jthorley/radcon/radcon.html. PMID- 10871274 TI - Concerning the accuracy of MAST E-values. PMID- 10871275 TI - Role of EGR1 in hippocampal synaptic enhancement induced by tetanic stimulation and amputation. AB - Hippocampal neurons fire spikes when an animal is at a particular location or performs certain behaviors in a particular place, providing a cellular basis for hippocampal involvement in spatial learning and memory. In a natural environment, spatial memory is often associated with potentially dangerous sensory experiences such as noxious or painful stimuli. The central sites for such pain-associated memory or plasticity have not been identified. Here we present evidence that excitatory glutamatergic synapses within the CA1 region of the hippocampus may play a role in storing pain-related information. Peripheral noxious stimulation induced excitatory postsynaptic potentials (EPSPs) in CA1 pyramidal cells in anesthetized animals. Tissue/nerve injury caused a rapid increase in the level of the immediate-early gene product Egr1 (also called NGFI-A, Krox24, or zif/268) in hippocampal CA1 neurons. In parallel, synaptic potentiation induced by a single tetanic stimulation (100 Hz for 1 s) was enhanced after the injury. This enhancement of synaptic potentiation was absent in mice lacking Egr1. Our data suggest that Egr1 may act as an important regulator of pain-related synaptic plasticity within the hippocampus. PMID- 10871276 TI - The plant vacuolar sorting receptor AtELP is involved in transport of NH(2) terminal propeptide-containing vacuolar proteins in Arabidopsis thaliana. AB - Many soluble plant vacuolar proteins are sorted away from secreted proteins into small vesicles at the trans-Golgi network by transmembrane cargo receptors. Cleavable vacuolar sorting signals include the NH(2)-terminal propeptide (NTPP) present in sweet potato sporamin (Spo) and the COOH-terminal propeptide (CTPP) present in barley lectin (BL). These two proteins have been found to be transported by different mechanisms to the vacuole. We examined the ability of the vacuolar cargo receptor AtELP to interact with the sorting signals of heterologous and endogenous plant vacuolar proteins in mediating vacuolar transport in Arabidopsis thaliana. AtELP extracted from microsomes was found to interact with the NTPPs of barley aleurain and Spo, but not with the CTPPs of BL or tobacco chitinase, in a pH-dependent and sequence-specific manner. In addition, EM studies revealed the colocalization of AtELP with NTPP-Spo at the Golgi apparatus, but not with BL-CTPP in roots of transgenic Arabidopsis plants. Further, we found that AtELP interacts in a similar manner with the NTPP of the endogenous vacuolar protein AtALEU (Arabidopsis thaliana Aleu), a protein highly homologous to barley aleurain. We hypothesize that AtELP functions as a vacuolar sorting receptor involved in the targeting of NTPP-, but not CTPP-containing proteins in Arabidopsis. PMID- 10871277 TI - Inhibitors of COPI and COPII do not block PEX3-mediated peroxisome synthesis. AB - In humans, defects in peroxisome biogenesis are the cause of lethal diseases typified by Zellweger syndrome. Here, we show that inactivating mutations in human PEX3 cause Zellweger syndrome, abrogate peroxisome membrane synthesis, and result in reduced abundance of peroxisomal membrane proteins (PMPs) and/or mislocalization of PMPs to the mitochondria. Previous studies have suggested that PEX3 may traffic through the ER en route to the peroxisome, that the COPI inhibitor, brefeldin A, leads to accumulation of PEX3 in the ER, and that PEX3 overexpression alters the morphology of the ER. However, we were unable to detect PEX3 in the ER at early times after expression. Furthermore, we find that inhibition of COPI function by brefeldin A has no effect on trafficking of PEX3 to peroxisomes and does not inhibit PEX3-mediated peroxisome biogenesis. We also find that inhibition of COPII-dependent membrane traffic by a dominant negative SAR1 mutant fails to block PEX3 transport to peroxisomes and PEX3-mediated peroxisome synthesis. Based on these results, we propose that PEX3 targeting to peroxisomes and PEX3-mediated peroxisome membrane synthesis may occur independently of COPI- and COPII-dependent membrane traffic. PMID- 10871278 TI - Cdc28 activates exit from mitosis in budding yeast. AB - The activity of the cyclin-dependent kinase 1 (Cdk1), Cdc28, inhibits the transition from anaphase to G1 in budding yeast. CDC28-T18V, Y19F (CDC28-VF), a mutant that lacks inhibitory phosphorylation sites, delays the exit from mitosis and is hypersensitive to perturbations that arrest cells in mitosis. Surprisingly, this behavior is not due to a lack of inhibitory phosphorylation or increased kinase activity, but reflects reduced activity of the anaphase promoting complex (APC), a defect shared with other mutants that lower Cdc28/Clb activity in mitosis. CDC28-VF has reduced Cdc20- dependent APC activity in mitosis, but normal Hct1- dependent APC activity in the G1 phase of the cell cycle. The defect in Cdc20-dependent APC activity in CDC28-VF correlates with reduced association of Cdc20 with the APC. The defects of CDC28-VF suggest that Cdc28 activity is required to induce the metaphase to anaphase transition and initiate the transition from anaphase to G1 in budding yeast. PMID- 10871279 TI - Phosphorylation by Cdc28 activates the Cdc20-dependent activity of the anaphase promoting complex. AB - Budding yeast initiates anaphase by activating the Cdc20-dependent anaphase promoting complex (APC). The mitotic activity of Cdc28 (Cdk1) is required to activate this form of the APC, and mutants that are impaired in mitotic Cdc28 function have difficulty leaving mitosis. This defect can be explained by a defect in APC phosphorylation, which depends on mitotic Cdc28 activity in vivo and can be catalyzed by purified Cdc28 in vitro. Mutating putative Cdc28 phosphorylation sites in three components of the APC, Cdc16, Cdc23, and Cdc27, makes the APC resistant to phosphorylation both in vivo and in vitro. The nonphosphorylatable APC has normal activity in G1, but its mitotic, Cdc20 dependent activity is compromised. These results show that Cdc28 activates the APC in budding yeast to trigger anaphase. Previous reports have shown that the budding yeast Cdc5 homologue, Plk, can also phosphorylate and activate the APC in vitro. We show that, like cdc28 mutants, cdc5 mutants affect APC phosphorylation in vivo. However, although Cdc5 can phosphorylate Cdc16 and Cdc27 in vitro, this in vitro phosphorylation does not occur on in vivo sites of phosphorylation. PMID- 10871280 TI - CYK-4: A Rho family gtpase activating protein (GAP) required for central spindle formation and cytokinesis. AB - During cytokinesis of animal cells, the mitotic spindle plays at least two roles. Initially, the spindle positions the contractile ring. Subsequently, the central spindle, which is composed of microtubule bundles that form during anaphase, promotes a late step in cytokinesis. How the central spindle assembles and functions in cytokinesis is poorly understood. The cyk-4 gene has been identified by genetic analysis in Caenorhabditis elegans. Embryos from cyk-4(t1689ts) mutant hermaphrodites initiate, but fail to complete, cytokinesis. These embryos also fail to assemble the central spindle. We show that the cyk-4 gene encodes a GTPase activating protein (GAP) for Rho family GTPases. CYK-4 activates GTP hydrolysis by RhoA, Rac1, and Cdc42 in vitro. RNA-mediated interference of RhoA, Rac1, and Cdc42 indicates that only RhoA is essential for cytokinesis and, thus, RhoA is the likely target of CYK-4 GAP activity for cytokinesis. CYK-4 and a CYK 4:GFP fusion protein localize to the central spindle and persist at cell division remnants. CYK-4 localization is dependent on the kinesin-like protein ZEN 4/CeMKLP1 and vice versa. These data suggest that CYK-4 and ZEN-4/CeMKLP1 cooperate in central spindle assembly. Central spindle localization of CYK-4 could accelerate GTP hydrolysis by RhoA, thereby allowing contractile ring disassembly and completion of cytokinesis. PMID- 10871281 TI - TPX2, A novel xenopus MAP involved in spindle pole organization. AB - TPX2, the targeting protein for Xenopus kinesin-like protein 2 (Xklp2), was identified as a microtubule-associated protein that mediates the binding of the COOH-terminal domain of Xklp2 to microtubules (Wittmann, T., H. Boleti, C. Antony, E. Karsenti, and I. Vernos. 1998. J. Cell Biol. 143:673-685). Here, we report the cloning and functional characterization of Xenopus TPX2. TPX2 is a novel, basic 82.4-kD protein that is phosphorylated during mitosis in a microtubule-dependent way. TPX2 is nuclear during interphase and becomes localized to spindle poles in mitosis. Spindle pole localization of TPX2 requires the activity of the dynein-dynactin complex. In late anaphase TPX2 becomes relocalized from the spindle poles to the midbody. TPX2 is highly homologous to a human protein of unknown function and thus defines a new family of vertebrate spindle pole components. We investigated the function of TPX2 using spindle assembly in Xenopus egg extracts. Immunodepletion of TPX2 from mitotic egg extracts resulted in bipolar structures with disintegrating poles and a decreased microtubule density. Addition of an excess of TPX2 to spindle assembly reactions gave rise to monopolar structures with abnormally enlarged poles. We conclude that, in addition to its function in targeting Xklp2 to microtubule minus ends during mitosis, TPX2 also participates in the organization of spindle poles. PMID- 10871282 TI - Coupling of Gab1 to c-Met, Grb2, and Shp2 mediates biological responses. AB - Gab1 is a substrate of the receptor tyrosine kinase c-Met and involved in c-Met specific branching morphogenesis. It associates directly with c-Met via the c-Met binding domain, which is not related to known phosphotyrosine-binding domains. In addition, Gab1 is engaged in a constitutive complex with the adaptor protein Grb2. We have now mapped the c-Met and Grb2 interaction sites using reverse yeast two-hybrid technology. The c-Met-binding site is localized to a 13-amino acid region unique to Gab1. Insertion of this site into the Gab1-related protein p97/Gab2 was sufficient to confer c-Met-binding activity. Association with Grb2 was mapped to two sites: a classical SH3-binding site (PXXP) and a novel Grb2 SH3 consensus-binding motif (PX(V/I)(D/N)RXXKP). To detect phosphorylation-dependent interactions of Gab1 with downstream substrates, we developed a modified yeast two-hybrid assay and identified PI(3)K, Shc, Shp2, and CRKL as interaction partners of Gab1. In a trk-met-Gab1-specific branching morphogenesis assay, association of Gab1 with Shp2, but not PI(3)K, CRKL, or Shc was essential to induce a biological response in MDCK cells. Overexpression of a Gab1 mutant deficient in Shp2 interaction could also block HGF/SF-induced activation of the MAPK pathway, suggesting that Shp2 is critical for c-Met/Gab1-specific signaling. PMID- 10871283 TI - Colocalization and redistribution of dishevelled and actin during Wnt-induced mesenchymal morphogenesis. AB - Activation of the Wnt signaling pathway is important for induction of gene expression and cell morphogenesis throughout embryonic development. We examined the subcellular localization of dishevelled, the immediate downstream component from the Wnt receptor, in the embryonic mouse kidney. Using immunofluorescence staining, confocal microscopy, and coimmunoprecipitation experiments, we show that dishevelled associates with actin fibers and focal adhesion plaques in metanephric mesenchymal cells. Stimulation of Wnt signaling leads to profound changes in metanephric mesenchymal cell morphology, including disruption of the actin cytoskeleton, increased cell spreading, and increased karyokinesis. Upon activation of Wnt signaling, dishevelled also accumulates in and around the nucleus. Casein kinase Iepsilon colocalizes with dishevelled along actin fibers and in the perinuclear region, whereas axin and GSK-3 are only present around the nucleus. These data indicate a branched Wnt signaling pathway comprising a canonical signal that targets the nucleus and gene expression, and another signal that targets the cytoskeleton and regulates cell morphogenesis. PMID- 10871284 TI - Shared and unique roles of CAP23 and GAP43 in actin regulation, neurite outgrowth, and anatomical plasticity. AB - CAP23 is a major cortical cytoskeleton-associated and calmodulin binding protein that is widely and abundantly expressed during development, maintained in selected brain structures in the adult, and reinduced during nerve regeneration. Overexpression of CAP23 in adult neurons of transgenic mice promotes nerve sprouting, but the role of this protein in process outgrowth was not clear. Here, we show that CAP23 is functionally related to GAP43, and plays a critical role to regulate nerve sprouting and the actin cytoskeleton. Knockout mice lacking CAP23 exhibited a pronounced and complex phenotype, including a defect to produce stimulus-induced nerve sprouting at the adult neuromuscular junction. This sprouting deficit was rescued by transgenic overexpression of either CAP23 or GAP43 in adult motoneurons. Knockin mice expressing GAP43 instead of CAP23 were essentially normal, indicating that, although these proteins do not share homologous sequences, GAP43 can functionally substitute for CAP23 in vivo. Cultured sensory neurons lacking CAP23 exhibited striking alterations in neurite outgrowth that were phenocopied by low doses of cytochalasin D. A detailed analysis of such cultures revealed common and unique functions of CAP23 and GAP43 on the actin cytoskeleton and neurite outgrowth. The results provide compelling experimental evidence for the notion that CAP23 and GAP43 are functionally related intrinsic determinants of anatomical plasticity, and suggest that these proteins function by locally promoting subplasmalemmal actin cytoskeleton accumulation. PMID- 10871285 TI - GAP43, MARCKS, and CAP23 modulate PI(4,5)P(2) at plasmalemmal rafts, and regulate cell cortex actin dynamics through a common mechanism. AB - The dynamic properties of the cell cortex and its actin cytoskeleton determine important aspects of cell behavior and are a major target of cell regulation. GAP43, myristoylated alanine-rich C kinase substrate (MARCKS), and CAP23 (GMC) are locally abundant, plasmalemma-associated PKC substrates that affect actin cytoskeleton. Their expression correlates with morphogenic processes and cell motility, but their role in cortex regulation has been difficult to define mechanistically. We now show that the three proteins accumulate at rafts, where they codistribute with PI(4,5)P(2), and promote its retention and clustering. Binding and modulation of PI(4, 5)P(2) depended on the basic effector domain (ED) of these proteins, and constructs lacking the ED functioned as dominant inhibitors of plasmalemmal PI(4,5)P(2) modulation. In the neuron-like cell line, PC12, NGF- and substrate-induced peripheral actin structures, and neurite outgrowth were greatly augmented by any of the three proteins, and suppressed by DeltaED mutants. Agents that globally mask PI(4,5)P(2) mimicked the effects of GMC on peripheral actin recruitment and cell spreading, but interfered with polarization and process formation. Dominant negative GAP43(DeltaED) also interfered with peripheral nerve regeneration, stimulus-induced nerve sprouting and control of anatomical plasticity at the neuromuscular junction of transgenic mice. These results suggest that GMC are functionally and mechanistically related PI(4,5)P(2) modulating proteins, upstream of actin and cell cortex dynamics regulation. PMID- 10871286 TI - Apical membrane targeting of Nedd4 is mediated by an association of its C2 domain with annexin XIIIb. AB - Nedd4 is a ubiquitin protein ligase (E3) containing a C2 domain, three or four WW domains, and a ubiquitin ligase HECT domain. We have shown previously that the C2 domain of Nedd4 is responsible for its Ca(2+)-dependent targeting to the plasma membrane, particularly the apical region of epithelial MDCK cells. To investigate this apical preference, we searched for Nedd4-C2 domain-interacting proteins that might be involved in targeting Nedd4 to the apical surface. Using immobilized Nedd4-C2 domain to trap interacting proteins from MDCK cell lysate, we isolated, in the presence of Ca(2+), a approximately 35-40-kD protein that we identified as annexin XIII using mass spectrometry. Annexin XIII has two known isoforms, a and b, that are apically localized, although XIIIa is also found in the basolateral compartment. In vitro binding and coprecipitation experiments showed that the Nedd4-C2 domain interacts with both annexin XIIIa and b in the presence of Ca(2+), and the interaction is direct and optimal at 1 microM Ca(2+). Immunofluorescence and immunogold electron microscopy revealed colocalization of Nedd4 and annexin XIIIb in apical carriers and at the apical plasma membrane. Moreover, we show that Nedd4 associates with raft lipid microdomains in a Ca(2+) dependent manner, as determined by detergent extraction and floatation assays. These results suggest that the apical membrane localization of Nedd4 is mediated by an association of its C2 domain with the apically targeted annexin XIIIb. PMID- 10871287 TI - Plasmin-sensitive dibasic sequences in the third fibronectin-like domain of L1 cell adhesion molecule (CAM) facilitate homomultimerization and concomitant integrin recruitment. AB - L1 is a multidomain transmembrane neural recognition molecule essential for neurohistogenesis. While moieties in the immunoglobulin-like domains of L1 have been implicated in both heterophilic and homophilic binding, the function of the fibronectin (FN)-like repeats remains largely unresolved. Here, we demonstrate that the third FN-like repeat of L1 (FN3) spontaneously homomultimerizes to form trimeric and higher order complexes. Remarkably, these complexes support direct RGD-independent interactions with several integrins, including alpha(v)beta(3) and alpha(5)beta(1). A pep- tide derived from the putative C-C' loop of FN3 (GSQRKHSKRHIHKDHV(852)) also forms trimeric complexes and supports alpha(v)beta(3) and alpha(5)beta(1) binding. Substitution of the dibasic RK(841) and KR(845) sequences within this peptide or the FN3 domain limited multimerization and abrogated integrin binding. Evidence is presented that the multimerization of, and integrin binding to, the FN3 domain is regulated both by conformational constraints imposed by other domains and by plasmin- mediated cleavage within the sequence RK( downward arrow)HSK( downward arrow)RH(846). The integrin alpha(9)beta(1), which also recognizes the FN3 domain, colocalizes with L1 in a manner restricted to sites of cell-cell contact. We propose that distal receptor ligation events at the cell-cell interface may induce a conformational change within the L1 ectodomain that culminates in receptor multimerization and integrin recruitment via interaction with the FN3 domain. PMID- 10871288 TI - Phosphorylation of connexin43 on serine368 by protein kinase C regulates gap junctional communication. AB - Phorbol esters (e.g., TPA) activate protein kinase C (PKC), increase connexin43 (Cx43) phosphorylation, and decrease cell-cell communication via gap junctions in many cell types. We asked whether PKC directly phosphorylates and regulates Cx43. Rat epithelial T51B cells metabolically labeled with (32)P(i) yielded two dimensional phosphotryptic maps of Cx43 with several phosphopeptides that increased in intensity upon TPA treatment. One of these peptides comigrated with the major phosphopeptide observed after PKC phosphorylation of immunoaffinity purified Cx43. Purification of this comigrating peptide and subsequent sequencing indicated that the phosphorylated serine was residue 368. To pursue the functional importance of phosphorylation at this site, fibroblasts from Cx43(-/-) mice were transfected with either wild-type (Cx43wt) or mutant Cx43 (Cx43-S368A). Intercellular dye transfer studies revealed different responses to TPA and were followed by single channel analyses. TPA stimulation of T51B cells or Cx43wt transfected fibroblasts caused a large increase in the relative frequency of approximately 50-pS channel events and a concomitant loss of approximately 100-pS channel events. This change to approximately 50-pS events was absent when cells transfected with Cx43-S368A were treated with TPA. These data strongly suggest that PKC directly phosphorylates Cx43 on S368 in vivo, which results in a change in single channel behavior that contributes to a decrease in intercellular communication. PMID- 10871290 TI - Neuroreceptors and ion channels as the basis for drug action: past, present, and future. AB - This article summarizes the development of cellular neuropharmacology and neurotoxicology, based primarily on my own research. The progress of this field depends at least in part on the theoretical and technological developments of excitable cell physiology, biophysics, and biochemistry. First, a brief historical development is described. Second, my earlier studies of the mechanism of action of insecticides on the nervous system are introduced. The most significant is the early discovery of the increase in depolarizing after potential caused by DDT and pyrethroids. This laid the foundation of subsequent analyses of sodium channel modulation as the major mechanism of action of DDT/pyrethroids. Third, my initial contributions to cellular neuropharmacology are described. The discovery of the potent and selective block of sodium channels by tetrodotoxin aroused interest not only in using this toxin and other chemicals as useful laboratory tools but also in studying receptors/channels as important targets of various drugs. Using internally perfused squid giant axons, pioneering studies of local anesthetic action led to the conclusion that these anesthetics block the sodium channel from inside the nerve membrane in the cationic form. Fourth, a few examples of my more recent studies using voltage-clamp and patch clamp techniques are described. Pyrethroid modulation of sodium channels was analyzed in great detail, including single-channel kinetics, toxicity amplification from channels to animal behaviors, temperature dependence, selective toxicity, and vitamin E antagonism. The neuroprotective drug riluzole blocked sodium channels and high-voltage-activated calcium channels, thereby preventing excess stimulation of N-methyl-D-aspartate receptors and massive influx of calcium, thereby retarding spread of infarction in the brain. Neuronal nicotinic acetylcholine receptors have received much attention recently, and I launched an extensive study of the mechanism whereby alcohols and general anesthetics modulate their activity. Ethanol potently stimulates the alpha bungarotoxin-insensitive, alpha4beta2-type acetylcholine receptors, thereby causing release of various transmitters; this leads to a cascade of multisynaptic events and behavioral changes. Inhalational general anesthetics augment the activity of gamma-aminobutyric acid(A) receptors and inhibit the activity of alpha4beta2-type acetylcholine receptors, causing a variety of clinical syndromes. Fifth, one of the possible future directions of cellular neuropharmacology and neurotoxicology is discussed. Emphasis is placed on the three-dimensional structure-activity relationship, in particular how changes in the molecular structure of drugs and receptors/channels result in kinetic changes in the function of receptors/channels. PMID- 10871289 TI - Activity-dependent neuronal control of gap-junctional communication in astrocytes. AB - A typical feature of astrocytes is their high degree of intercellular communication through gap junction channels. Using different models of astrocyte cultures and astrocyte/neuron cocultures, we have demonstrated that neurons upregulate gap-junctional communication and the expression of connexin 43 (Cx43) in astrocytes. The propagation of intercellular calcium waves triggered in astrocytes by mechanical stimulation was also increased in cocultures. This facilitation depends on the age and number of neurons, indicating that the state of neuronal differentiation and neuron density constitute two crucial factors of this interaction. The effects of neurons on astrocytic communication and Cx43 expression were reversed completely after neurotoxic treatments. Moreover, the neuronal facilitation of glial coupling was suppressed, without change in Cx43 expression, after prolonged pharmacological treatments that prevented spontaneous synaptic activity. Altogether, these results demonstrate that neurons exert multiple and differential controls on astrocytic gap-junctional communication. Since astrocytes have been shown to facilitate synaptic efficacy, our findings suggest that neuronal and astrocytic networks interact actively through mutual setting of their respective modes of communication. PMID- 10871291 TI - Endocannabinoids and vascular function. AB - Marijuana is used by humans for its psychoactive and medicinal effects. The active constituents of marijuana, the cannabinoids, exert effects via a G protein coupled receptor, CB(1). Two arachidonic acid analogs, N-arachidonylethanolamine and 2-arachidonylglycerol are hypothesized to function as endogenous ligands of the CB(1) receptor. The cannabinoids exert significant vascular effects in humans and laboratory animals. In particular, the cannabinoids produce vasodilation and hypotension. The possible mechanisms for these effects are inhibition of transmitter release from sympathetic nerve terminals, direct effects on vascular smooth muscle cells, and effects on endothelial cell function. The data regarding these effects of the cannabinoids and possible sources of endocannabinoid ligands in the vasculature are the subjects of this review. PMID- 10871292 TI - Z-338 facilitates acetylcholine release from enteric neurons due to blockade of muscarinic autoreceptors in guinea pig stomach. AB - The mechanism by which Z-338, a novel gastroprokinetic agent, stimulates gastric motility was studied in relation to muscarinic receptors in the guinea pig. Z-338 (3-30 microM) enhanced electrically stimulated contractions and the release of acetylcholine (ACh) that was tetrodotoxin sensitive and extracellular Ca(2+) dependent, in gastric strips. Membrane-binding assay revealed that Z-338 possessed binding affinity for muscarinic M(1) and M(2), but not M(3) receptors. In Xenopus oocytes expressing M(1) and M(2) muscarinic receptors, Z-338 did not produce any response, but inhibited ACh-induced outward currents, thereby indicating that Z-338 acts on the M(1) and M(2) muscarinic receptors as an antagonist. The M(1) receptor antagonist pirenzepine (0.5 microM) and M(2) receptor antagonist AF-DX 116 (1 microM) also enhanced electrically stimulated release of ACh. These results indicate that Z-338 facilitates ACh release from cholinergic nerve terminals by blocking muscarinic M(1) and M(2) autoreceptors, which regulate the release of ACh. PMID- 10871293 TI - Evidence for Y1-receptor-mediated facilitatory, modulatory cotransmission by NPY in the rat anococcygeus muscle. AB - The potential role of neuropeptide Y (NPY) as a neuromodulatory cotransmitter was investigated in rat anococcygeus muscle. The effects of NPY on contraction to norepinephrine or adrenergic nerve stimulation and on relaxation to nonadrenergic, noncholinergic nerve stimulation were analyzed. Norepinephrine induced contraction was enhanced by NPY (0.1 microM). The Y1 receptor antagonist BIBP 3226 (1 microM) completely reversed this effect. NPY (0.01 or 0.1 microM) increased contractions induced by electrical field stimulation of sympathetic nerves. This increase was reduced by BIBP 3226 (1 microM), indicating Y1 receptor involvement. NPY (13-36) a Y2 receptor agonist, at 0.1 microM but not 0.01 microM, caused an increase of the nerve-induced contraction, which was reversed by BIBP 3226 (1 microM), indicating no Y2 receptor involvement. BIBP 3226 (1-1 microM) produced a concentration-dependent attenuation of nerve-mediated but not norepinephrine-mediated contraction. The reduction in nonadrenergic, noncholinergic nerve-induced relaxation to nerve stimulation by NPY (0.1 microM) was not affected by BIBP 3226 (1 microM). It is concluded that 1) exogenous NPY increases excitatory nerve-induced contraction mainly via a Y1 receptor-mediated effect on smooth muscle with a small non-Y1 receptor component due to blocking inhibitory nitrergic nerves and 2) endogenous NPY is a modulatory cotransmitter, which facilitates the primarily noradrenergic contractile responses to sympathetic nerve stimulation via smooth muscle Y1 receptors. PMID- 10871294 TI - Effect of allelic polymorphism of the B(1) and B(2) receptor genes on the contractile responses of the human umbilical vein to kinins. AB - The human genes corresponding to the two receptor (R) subtypes for bradykinin (BK)-related peptides, the B(1)R and B(2)R, are known to be polymorphic. The human isolated umbilical vein responds by contractions to stimulation by kinins via constitutive B(2)Rs and inducible B(1)Rs. Vascular rings from 100 different umbilical cords were submitted to a standardized protocol where E(max) values were obtained at 2 and 6 h of incubation, and EC(50) values were estimated at 6 h for the B(1)R agonist Sar-?D-Phe(8)des-Arg(9)-BK; E(max) and EC(50) values were also obtained for the B(2)R agonist BK at 4 h. The genotype of each tissue donor was determined for two polymorphic sites in the B(1)R gene and three such sites in the B(2)R gene. The (-/-) genotype of a frequent insertion/deletion polymorphism of the B(2)R exon 1 was associated with increased contractile efficiency of the B(1)R agonist, Sar-?D-Phe(8)des-Arg(9)-BK, but had no effect on BK-induced contractility. A B(2)R exon 2 polymorphism (C(181) --> T) selectively influenced the potency of BK (EC(50) higher when the T allele was present). The other polymorphisms studied were not found to affect kinin-induced contractility. Although most of the frequent polymorphic alleles of the kinin receptor genes are functionally neutral or determine functional alterations that are not detectable using the method used here, two B(2)R polymorphic sites (exon 1, exon 2) modestly influence function. As the exon 1 B(2)R polymorphism predicts the response of the B(1)R agonist, it may be in linkage disequilibrium with an unknown, functionally important polymorphism of the neighboring B(1)R gene. PMID- 10871295 TI - Induction of cAMP response element modulator (CREM) and inducible cAMP early repressor (ICER) expression in rat brain by uncompetitive N-methyl-D-aspartate receptor antagonists. AB - The N-methyl-D-aspartate (NMDA) subtype of glutamate receptor mediates fast excitatory neurotransmission, and agents that attenuate this function are neuroprotective, anesthetic, and psychotropic. To determine whether cAMP regulatable transcription factors play a role in the neurochemical actions of agents acting through NMDA receptors, the effects of the acute administration of uncompetitive and competitive antagonists on the expression of cAMP response element modulator (CREM) and inducible cAMP early repressor (ICER) transcription factors were examined. In situ hybridization to rat brain sections revealed that ICER mRNA expression was significantly increased by uncompetitive NMDA receptor antagonists (MK-801, phencyclidine, ketamine, memantine) but not by the competitive antagonist CPP [(+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid] or other psychotropic agents (clozapine, haloperidol, desipramine). Major brain regions where ICER transcripts were increased were the hippocampus, parietal cortex layers IV and VI, temporal cortex, cingulate cortex, thalamus, and granule cell layer of the olfactory bulb. Northern and Western blot analyses indicated that CREM mRNA and protein, respectively, were also increased after MK 801 treatment, but ICER isoforms predominate during both basal and induced conditions. MK-801 also transiently increased the binding of proteins to cAMP response element, which was supershifted by anti-CREM antibody, indicating that increased mRNA and protein levels have functional consequences on gene transcription. These results provide evidence for the involvement of CREM and ICER family transcription factors in the pharmacologic effects of uncompetitive NMDA receptor antagonists. PMID- 10871296 TI - Amylin receptor phenotypes derived from human calcitonin receptor/RAMP coexpression exhibit pharmacological differences dependent on receptor isoform and host cell environment. AB - Receptor activity modifying proteins (RAMPs) constitute a group of three proteins, designated as RAMP1, 2, and 3, which are able to effect functional changes in some members of the G protein-coupled receptor family. Thus, RAMP1 or RAMP3 can modify the calcitonin receptor (CTR) to also function as a high affinity amylin receptor-like phenotype. To examine the RAMP/CTR interaction, individual RAMPs were coexpressed with either of the two human CTR (hCTR) isoforms, the insert negative (hCTR(I1-)) or the insert positive (hCTR(I1+)), in Chinese hamster ovary (CHO-P) or African monkey kidney (COS-7) cells. CHO-P cells provide an environment conducive to a low, but significant, level of amylin binding with either hCTR isoform alone, unlike in COS-7, where RAMP coexpression is imperative for amylin binding. Also, in CHO-P, hCTR(I1-) induced amylin binding with all three RAMPs, in contrast to COS-7, where only RAMP1 or RAMP3 generate an amylin receptor phenotype. hCTR(I1+) induced high-affinity amylin binding with any RAMP in either cell line. In COS-7 cells, hCTR(I1+)/RAMP generated receptor displayed high- and low-affinity states, in contrast with the single-state binding seen with hCTR(I1-)/RAMP-generated receptor, whereas in CHO P cells a two-affinity state receptor phenotype was evident with both hCTR isoforms. Endogenous RAMP expression is low and similar between cell lines. The results suggest that CTR/RAMP interaction in these cells is complex with other cellular factors such as the levels of different G proteins and/or receptor/RAMP stoichiometry following heterologous coexpression contributing to the ultimate receptor phenotype. PMID- 10871297 TI - Organic anion-transporting polypeptides mediate transport of opioid peptides across blood-brain barrier. AB - Organic anion-transporting polypeptides (Oatps) are a rapidly growing gene family of polyspecific membrane transporters. In rat brain, Oatp1 (gene symbol Slc21a1) and Oatp2 (Slc21a5) are localized at the apical and basolateral domains, respectively, of the choroid plexus epithelium. Furthermore, Oatp2 is strongly expressed at the rat blood-brain barrier (BBB). This study localizes the human OATP (now called OATP-A; SLC21A3) at the BBB in humans. Furthermore, with the Xenopus laevis oocyte system the delta-opioid receptor agonists [D penicillamine(2,5)]enkephalin (DPDPE) and deltorphin II were identified as new transport substrates of OATP-A. This OATP-A-mediated DPDPE and deltorphin II transport exhibited apparent K(m) values of approximately 202 and 330 microM, respectively, and OATP-A-mediated deltorphin II transport was inhibited by the mu opioid receptor agonist Tyr-D-Ala-Gly-N-methyl-Phe-glycinol, the endogenous peptide Leu-enkephalin, and the opiate antagonists naloxone and naltrindole. DPDPE also was transported by rat Oatp1 (K(m) approximately 48 microM) and Oatp2 (K(m) approximately 19 microM), whereas deltorphin II was only transported by Oatp1 (K(m) approximately 137 microM). These results demonstrate that OATP-A can mediate transport of the analgesic opioid peptides DPDPE and deltorphin II across the human BBB. Furthermore, because rat Oatp1 and Oatp2 exhibit similar but not identical transport activities as OATP-A, the results generally indicate that members of the Oatp/OATP gene family of membrane transporters play an important role in carrier-mediated transport of opioid peptides across the BBB and blood cerebrospinal fluid barrier of the mammalian brain. PMID- 10871298 TI - Phenacetin deacetylase activity in human liver microsomes: distribution, kinetics, and chemical inhibition and stimulation. AB - Microsomal and cytosolic phenacetin deacetylase activities were examined in human liver and kidneys. Kinetic properties of the activities were also studied in human liver microsomes. Phenacetin deacetylase activity was predominantly localized in the liver microsomal fraction. The specific activities of phenacetin deacetylation in liver cytosol and in kidney microsomes and cytosol were all less than 5% of that in liver microsomes. In human liver microsomes, Eadie-Hofstee plots for phenacetin deacetylation were monophasic, indicating a single-enzyme catalytic reaction. The Michaelis-Menten parameters, K(m) and V(max), for the deacetylation were 4.7 mM and 5.54 nmol/min/mg of protein, respectively. The intrinsic clearance, calculated as V(max)/K(m), was 1.18 microl/min/mg of protein. Although the organophosphate bis(4-nitrophenyl)phosphoric acid markedly inhibited the reaction in human liver microsomes, the activity has a tolerance to the treatment of phenylmethylsulfonyl fluoride, a serine hydrolase inhibitor. Prazosin, a peripheral alpha(1)-adrenergic antagonist, noncompetitively inhibited the phenacetin deacetylation with a K(i) value of 19.0 microM. Flutamide, a nonsteroidal androgen receptor antagonist, stimulated the activity by up to 349%. This increase was accompanied by a decrease in the K(m) value and no change in the V(max) value, resulting in an increase in the intrinsic clearance by up to 700% of the control. These results suggest that the phenacetin deacetylase localized in human liver microsomes has not only a catalytic site but also a negative and/or positive modulation site or sites. PMID- 10871299 TI - Inhibition of human cytochrome P450 enzymes by constituents of St. John's Wort, an herbal preparation used in the treatment of depression. AB - Commercially available St. John's wort (Hypericum perforatum) extracts, preparations that are used in the treatment of depression, were examined for the potential to inhibit human cytochrome P450 (CYP) enzyme activities, specifically CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4. Crude extracts demonstrated inhibition of each of these five enzymes, with CYP2D6, CYP2C9, and CYP3A4 being more sensitive than CYP1A2 and CYP2C19. Extracts were fractionated by HPLC, and each of the fractions was tested for inhibition of these five CYPs to identify individual constituents with inhibitory activity. Several fractions were shown to possess inhibitory activity, including the fractions containing hyperforin (the putative active antidepressant constituent), I3,II8-biapigenin, and hypericin. Hyperforin and I3,II8-biapigenin were isolated from the extract, and inhibition constants for the five CYP activities were measured. In addition, three other constituents, hypericin, quercetin, and chlorogenic acid, were tested for inhibitory activity toward the CYP enzymes. The flavonoid compound I3,II8 biapigenin was shown to be a potent, competitive inhibitor of CYP3A4, CYP2C9, and CYP1A2 activities with K(i) values of 0.038, 0.32, and 0.95 microM, respectively. Hyperforin was a potent noncompetitive inhibitor of CYP2D6 activity (K(i) = 1.5 microM) and competitive inhibitor of CYP2C9 and CYP3A4 activities (K(i) = 1.8 and 0.48 microM, respectively). Hypericin also demonstrated potent inhibition of several CYP activities. These in vitro data indicate that St. John's wort preparations contain constituents that can potently inhibit the activities of major human drug-metabolizing enzymes and suggest that these preparations should be examined for potential pharmacokinetic drug interactions in vivo. PMID- 10871300 TI - MK-886, a leukotriene biosynthesis inhibitor, as an activator of Ca(2+) mobilization in Madin-Darby canine kidney (MDCK) cells. AB - The effect of 3-?1-(p-chlorobenzyl)-5-(isopropyl)-3-tert-butylthioindol-2-yl-2, 2 dimethylpropanoic acid (MK-886), a leukotriene biosynthesis inhibitor, on Ca(2+) mobilization in Madin- Darby canine kidney cells has been examined by fluorimetry using fura-2 as a Ca(2+) indicator. MK-886 at 0.5 to 25 microM concentration dependently increased ?Ca(2+)(i). The ?Ca(2+)(i) increase comprised an immediate initial rise and a slowly decaying phase. Ca(2+) removal inhibited the Ca(2+) signals by reducing both the initial rise and the decay phase, suggesting that MK 886 activated Ca(2+) influx and Ca(2+) release. In Ca(2+)-free medium, 10 microM MK-886 still increased ?Ca(2+)(i) after pretreatment with carbonylcyanide m chlorophenylhydrazone (CCCP; 2 microM), a mitochondrial uncoupler, and thapsigargin (1 microM), an endoplasmic reticulum Ca(2+) pump inhibitor. Conversely, pretreatment with MK-886 abolished CCCP- and thapsigargin-induced Ca(2+) release. This suggests that 10 microM MK-886 released Ca(2+) from the endoplasmic reticulum, mitochondria, and other stores. The addition of 3 mM Ca(2+) increased ?Ca(2+)(i) after pretreatment with 10 microM MK-886 for 700 s in Ca(2+)-free medium, indicating that MK-886 induced capacitative Ca(2+) entry. This capacitative Ca(2+) entry was partly inhibited by SKF96365 (50 microM), by econazole (25 microM), and by inhibiting phospholipase A(2) with aristolochic acid (40 microM) but not by inhibiting phospholipase D with 0.1 mM propranolol. MK-886 (10 microM)-induced Ca(2+) release was not altered by inhibiting phospholipase C with U73122 (2 microM) but was inhibited by 50% by suppressing phospholipase D and phospholipase A(2) with propranolol (0.1 mM) and aristolochic acid (40 microM), respectively. PMID- 10871301 TI - Adrenal glucocorticoids modulate [3H]cyclic AMP binding to protein kinase A (PKA), cyclic AMP-dependent PKA activity, and protein levels of selective regulatory and catalytic subunit isoforms of PKA in rat brain. AB - Alterations in hypothalamic-pituitary-adrenal (HPA) function are associated with changes in mood and behavior. Protein kinase A (PKA), on activation, phosphorylates many important intracellular proteins and thereby plays a major role in mediating various physiological functions in brain. We systematically examined the relationship of altered HPA function with PKA modifications in rat brain after administering corticosterone to normal rats and by first adrenalectomizing rats and then simultaneously treating them with different doses of corticosterone. Rats were decapitated on day 1, 4, or 14. Subcutaneously implanted 50- or 100-mg corticosterone pellets in normal rats for 4 or 14 days significantly decreased PKA activity, B(max) of [3H]cyclic AMP binding, and protein levels of selective PKA regulatory (RIalpha, RIIbeta) and catalytic (Catbeta) subunit isoforms in cortex and hippocampus in a dose-dependent manner without any significant changes at day 1; these changes were more pronounced at day 14. However, adrenalectomy caused the opposite changes in these measures at day 4 or 14 in both cortex and hippocampus, and the magnitude of the changes was more pronounced at day 14. Simultaneous treatment with implanted corticosterone at 50- or 100-mg doses in adrenalectomized rats reversed the adrenalectomy induced increases in PKA measures in a dose-dependent manner. These results suggest that endogenous glucocorticoid modifies the expression of RIalpha, RIIalpha, and Catbeta subunit isoforms of PKA, as well as the catalytic and regulatory activities of PKA, and that these alterations in PKA may in part explain HPA axis-mediated changes in mood and behavior. PMID- 10871302 TI - Involvement of a receptor-mediated component in cellular translocation of riboflavin. AB - This study addresses the transport mechanism of riboflavin (vitamin B(2)) across intestinal epithelium in the presence and absence of pharmacologically active compounds. A polarized transport process with a 6-fold higher basolateral (BL)-to apical (AP) flux was observed in both a human intestinal cell model (Caco-2) and rat intestinal tissue. Riboflavin-specific translocation systems on both the AP and BL cell surfaces were saturable with affinity values close to most receptors (K(m): 9.72 +/- 0.85 and 4.06 +/- 0.03 nM, respectively). Pharmacological agents known to alter intracellular endocytic events were used to examine the potential involvement of receptor-mediated events. Nocodazole significantly inhibited AP uptake (58.4%), BL-to-AP riboflavin (56.7%) and fluorescein isothiocyanate labeled transferrin (FITC-Tf) (31.8%) transport without affecting mannitol or cholic acid transport, whereas AP-to-BL riboflavin (252.8%) and FITC-Tf (145.1%) transport was increased. Brefeldin A significantly enhanced AP-to-BL riboflavin (37.1%) and bidirectional FITC-Tf transport (AP-to-BL: 13-fold; BL-to-AP: 5 fold). without affecting BL-to-AP riboflavin transport. Combined, these data suggest an essential role of microtubule-dependent movement and vesicular sorting component(s) in the bidirectional transport of riboflavin. Dissociation of riboflavin from the cell surface was pH-dependent with significantly higher substrate release at acidic pH, indicating the presence of riboflavin-specific cell surface receptors. In summary, our studies provide biochemical evidence of the involvement of a receptor-mediated mechanism in the cellular translocation of riboflavin. PMID- 10871303 TI - Chronic oral administration of ATP modulates nucleoside transport and purine metabolism in rats. AB - The effect of repeated oral administration of ATP on purine transport and metabolism was investigated in rats. An increased ability of the gut to capture intraluminal purine nucleosides and to export ATP and nucleosides toward portal bloodstream was observed in rats after 30 days of treatment with 5 mg/kg/day ATP. This was accompanied in erythrocytes by an increased transport of adenosine rapidly transformed into ATP, which in turn was exported toward extracellular fluid. However, these metabolic changes were associated with a paradoxical and progressive diminution of plasma ATP level below that found in control rats and that was not strictly dependent on the ATP dose administered, whereas plasma adenosine concentration remained unchanged. This diminution likely resulted from an increased ectonucleotidase activity, suggesting that the chronic administration of ATP seems to induce a progressive adaptation of purine metabolism. This adaptive response to free purine supplementation affects both intracellular metabolism and purine exchange between intracellular and extracellular compartments. This modification of free purine turnover and delivery may affect physiological parameters under the control of P(1) and P(2) purinoceptors described in different experimental models. PMID- 10871304 TI - Iron-mediated free radical injury in ethanol-exposed mouse neural crest cells. AB - Previous studies using cell and whole embryo cultures have shown that free radicals play an important role in the ethanol-induced death of mouse neural crest cells (NCCs; a significant cell type with respect to the genesis of alcohol related birth defects). This investigation was spurred by reports of increased iron in ethanol-exposed fetuses and the knowledge that iron can initiate the production of reactive oxygen species. Initially, the ameliorative potential of two iron chelators, deferoxamine and phenanthroline, relative to ethanol-induced cell death was examined. Cotreatment of cultured NCCs with 100 mM ethanol and either 1 or 10 microM deferoxamine or 10, 50, or 250 microM phenanthroline significantly increased the percentage of viable cells as compared with exposure to 100 mM ethanol alone. These data indicate that iron is involved in the ethanol induced cytotoxicity. To support this premise, the direct toxicity of iron to NCCs was also examined. As expected, loading the cells with Fe(II)/Fe(III) using 8-hydroxyquinoline as a carrier had an adverse effect on their viability as did treatment with a neurotoxin, 6-hydroxydopamine, that releases iron from ferritin storage. Cotreatment with an antioxidant, N-acetylcysteine, significantly diminished the toxicity of ethanol alone, that resulting from iron loading, as well as from the combination of ethanol exposure and iron loading. These results confirm the role of free radical-mediated damage in ethanol-induced cytotoxicity and highlight the potential role of iron relative to the genesis of alcohol related birth defects. PMID- 10871305 TI - The effects of cocaine and nicotine on amino acid transport across the human placental cotyledon perfused in vitro. AB - The inhibitory effects of cocaine and nicotine on placental amino acid transport, as a mechanism contributing to intrauterine growth restriction, were investigated in the in vitro placental perfusion model. Amino acids that represent substrates for known placental transporters were selected: alanine (system A), glutamine (system N), phenylalanine and valine (system l), and arginine (system y(+)). Amino acid accumulation on the fetal side was measured in the absence of cocaine or nicotine (n = 7) and in the presence of 1.2 microg/ml cocaine (n = 6), 120 ng/ml nicotine (n = 6), or both (n = 6). Neither cocaine nor nicotine alone significantly inhibited alanine transport, whereas their combination did (P =.02). Significant inhibition of arginine transport was detected with nicotine (P =.007), cocaine (P =.01), and their combination (P =.01), whereas phenylalanine (P =.03, P =.04) and valine (P =.03, P =.04) transport was affected by cocaine and the combination of cocaine and nicotine, respectively. For glutamine, neither cocaine, nicotine, nor their combination had a statistically significant inhibitory effect. In conclusion, both cocaine and nicotine may contribute to fetal growth restriction by interfering with the activity of amino acids transporters that are necessary to maintain the nutrient gradients associated with normal fetal growth. PMID- 10871306 TI - Tachykinin receptor subtypes in the isolated guinea pig heart and their role in mediating responses to neurokinin A. AB - Selective tachykinin agonists were used to identify cardiac and coronary responses mediated by specific tachykinin receptor subtypes in isolated, perfused guinea pig hearts. Receptor desensitization with selective agonists and blockade with selective antagonists were used to determine the role of specific subtypes in generating responses to neurokinin A (NKA). Dose-dependent cardiac and coronary effects were evoked by bolus injections of ?Sar(9), Met(O(2))(11)substance P (?Sar(9),Met(O(2))(11)SP), GR64349, and ?MePhe(7)neurokinin B (?MePhe(7)NKB) (selective agonists for NK(1), NK(2), and NK(3) receptors, respectively). Each agonist caused bradycardia, but GR64349 was most effective (34 +/- 4% decrease in heart rate with 32 nmol, n = 8). Prominent increases in ventricular contractility and perfusion pressure also occurred with 32 nmol of GR64349 (25 +/- 6 and 33 +/- 4%, respectively). ?Sar(9), Met(O(2))(11)SP was unique in having a high potency for decreasing ventricular contractility and perfusion pressure. Bolus injections of 25 nmol of NKA decreased rate (48 +/- 2%, n = 51), increased contractility (26 +/- 2%), and had biphasic effects on perfusion pressure (24 +/- 1% decrease followed by 9.2 +/- 1.4% increase). Desensitization with GR64349 or treatment with the NK(2) antagonist SR48968 reduced the bradycardic response to NKA by greater than 75% and eliminated the positive inotropic response. The remaining bradycardia occurred through NK(3) receptors. Desensitization with ?Sar(9),Met(O(2))(11)SP or NK(1) blockade with FK888 eliminated the coronary relaxant action of NKA and enhanced the pressor response. It is concluded that three tachykinin receptor subtypes are present in the guinea pig heart and that each contributes to the overall response evoked by NKA. PMID- 10871307 TI - Effect of oral and intravenous S,N-diacetylcysteine monoethyl ester on circulating and hepatic sulfhydryls in the Rat. AB - The role of GSH in the detoxification of reactive metabolites of oxygen and xenobiotics, in gene expression, and as a source of cysteine is well established. Because decreased circulating and intracellular concentrations of GSH might be of pathogenetic relevance in several clinical conditions, there is a growing interest in pharmacological interventions to correct a deranged sulfhydryl status. In this study, the disposition and the effect of S,N-diacetylcysteine monoethyl ester (DACE) on sulfhydryls were investigated after i.v. and intraduodenal (i.d.) administrations to rats. DACE was rapidly hydrolyzed and deacetylated to N-acetylcysteine and cysteine in plasma. High concentrations of cysteine were attained in the circulation and in the liver after i.v. and i.d. administrations of 5 mmol/kg DACE, and physiological levels of GSH in the liver and in plasma increased by 30 and 300%, respectively, with i.v. and i.d. administrations. Incubation of peripheral blood mononuclear cells with 1 mM DACE resulted in higher intracellular concentrations of cysteine and GSH after 24 h than incubations with equimolar concentrations of cysteine, N-acetylcysteine, or oxothiazolidine carboxylic acid, respectively. It is concluded that DACE provides an efficient delivery system for cysteine that markedly increases intra- and extracellular cysteine and GSH after i.v. and i.d. administrations. Because its uptake into cells is probably not dependent on an active transport process, DACE results in higher intracellular concentrations of cysteine than those resulting from other prodrugs of cysteine and cysteine itself. The compound may thus have advantages over other compounds for the correction of a deranged sulfhydryl status. PMID- 10871308 TI - Hepatotoxicity of tacrine: occurrence of membrane fluidity alterations without involvement of lipid peroxidation. AB - Tacrine (THA), used in the treatment of Alzheimer's disease, is known to induce hepatotoxicity, the mechanisms of which remain to be fully established. We have previously shown that THA reduced intracellular glutathione concentration in rat hepatocytes in primary culture, thus pointing to a possible role for oxidative stress in THA toxicity. To test this, the effects of antioxidant molecules, namely, the flavonoids silibinin, silibinin dihydrogensuccinate, and silymarin, were evaluated on the toxicity of THA in cultured rat hepatocytes. This toxicity was investigated after a 24-h treatment over a concentration range from 0 to 1 mM, in the presence or absence of antioxidant (1 and 10 microM). We found that simultaneous treatment of hepatocytes with any of the antioxidants and THA remained ineffective on the lactate dehydrogenase release induced by THA. Then, the production of lipid-derived radicals (to estimate lipid peroxidation) was measured in THA (0.05-0.50 mM)-treated cells using a spin-trapping technique coupled to electron paramagnetic resonance (EPR) spectroscopy. No increase of the EPR signal was observed over the period of 30 min to 24 h. In contrast, treatment of cells with the spin label 12-doxyl stearic acid followed by EPR spectroscopy showed that THA (0.05 and 0.25 mM) rapidly increased hepatocyte membrane fluidity. Extracellular application of GM1 ganglioside (60 microM) both reversed this increase in fluidity and partially reduced lactate dehydrogenase release on THA exposure. In conclusion, this work indicates that early alterations of membrane fluidity, not resulting from lipid peroxidation, are likely to play an important role in the development of THA toxicity. PMID- 10871309 TI - Chronic l-alpha-acetylmethadol (LAAM) in rhesus monkeys: tolerance and cross tolerance to the antinociceptive, ventilatory, and rate-decreasing effects of opioids. AB - Although l-alpha-acetylmethadol (LAAM) is a maintenance treatment for opioid dependence, few studies have systematically assessed the behavioral effects of LAAM and other drugs in LAAM-treated subjects. In the current study, we assessed the ventilatory, antinociceptive, and rate-decreasing effects of drugs (s.c. except dynorphin, which was administered i.v.) in rhesus monkeys (n = 3 or 4) before and during chronic treatment with 1.0 mg/kg/12 h LAAM (s.c.). Minute volume (V(E)) was reduced to 62% of baseline during LAAM treatment and remained depressed after more than 10 months of LAAM treatment. A cumulative dose of 10.0 mg/kg morphine decreased V(E) to similar values under baseline (53%) and LAAM treated (52%) conditions; however, larger doses of morphine (up to 56.0 mg/kg) could be administered safely to LAAM-treated monkeys. LAAM treatment produced dependence as evidenced by a 220% increase in V(E) after a dose of naltrexone (0.032 mg/kg) that did not modify ventilation under baseline conditions. Compared with baseline, LAAM treatment increased the ED(50) values for the rate-decreasing effects of nalbuphine, morphine, and alfentanil by 7-, 7-, and 2-fold, respectively, in monkeys responding under a fixed ratio 10 schedule of food presentation. Similarly, LAAM treatment increased ED(50) values for the antinociceptive effects of morphine and alfentanil by 5- and 3-fold, respectively. LAAM treatment also increased the ED(50) values for the antinociceptive effects of the kappa-agonist enadoline by 5-fold and not those of U-50,488. That tolerance developed differentially to the ventilatory, rate, and antinociceptive effects of mu-agonists in LAAM-treated monkeys suggests that cross-tolerance might not be a safe therapeutic approach for the treatment of some opioid abusers. PMID- 10871310 TI - Differential inhibition of the prejunctional actions of angiotensin II in rat atria by valsartan, irbesartan, eprosartan, and losartan. AB - The effects of valsartan and other nonpeptide angiotensin II type 1 (AT(1)) receptor blockers on the prejunctional actions of angiotensin II were investigated in the isolated left atria of rat. Norepinephrine stores in rat atria were loaded with [(3)H]norepinephrine, and neuronal norepinephrine release was deduced from the radioactivity efflux. Angiotensin II (10(-9) to 10(-6) M) produced concentration-dependent enhancement of the electrical stimulation induced efflux of [(3)H]norepinephrine from the preparation. Pretreatment of tissues with valsartan, irbesartan, eprosartan, or losartan (10(-8) to 10(-6) M) produced concentration-dependent inhibitions of the stimulation-induced efflux of radioactivity observed in the presence of angiotensin II (10(-7) M). The AT(1) receptor blockers did not decrease the "basal stimulation-induced overflow of radioactivity but rather selectively inhibited the angiotensin II-mediated augmentation of the response. Regression analyses of the inhibition of the angiotensin II-mediated response by valsartan, irbesartan, eprosartan, and losartan revealed corresponding log IC(50) values (log M, with 95% confidence intervals) of -7.78 (-8.19, -7.51), -7.65 (-8.02, -7.40), -7.12 (-7. 37, -6.86), and -6.75 (-7.00, -6.40), indicating that the IC(50) values for valsartan and irbesartan are significantly lower than those for eprosartan and losartan. Thus, valsartan is a potent inhibitor of the prejunctional facilitatory effect of angiotensin II on the release of norepinephrine from peripheral sympathetic nerves. This implies that the therapeutic domain of valsartan may be extended to include pathophysiological conditions such as congestive heart failure wherein prejunctional angiotensin II receptors apparently play a significant role. Whether the high potency of valsartan translates into a significant clinical advantage relative to the other agents tested remains to be ascertained. PMID- 10871311 TI - Biochemical and neurobehavioral profile of CHF2819, a novel, orally active acetylcholinesterase inhibitor for Alzheimer's disease. AB - 1,2,3,3a,8,8a-Hexahydro-1,3a,8-trimethylpyrrolo?2,3-bindol-5-ol 2 ethylphenylcarbamate N-oxide hydrochloride (3aS-cis) (CHF2819) is a novel acetylcholinesterase inhibitor that produces central cholinergic stimulation after oral administration in rats. In vivo studies show that CHF2819 (0.5, 1.5, and 4.5 mg/kg p.o.) significantly increases acetylcholine levels in young adult rat hippocampus in a dose-dependent manner. Moreover, aged animals, which show a significant decrease in basal acetylcholine levels with respect to young adult rats, also exhibit a marked increase in the hippocampal concentrations of this neurotransmitter after the administration of CHF2819. This compound (1.5 mg/kg p.o.) significantly attenuates scopolamine-induced amnesia in a passive avoidance task. Furthermore, CHF2819 induces a significant decrease in dopamine levels and a significant elevation of extracellular concentrations of 5-hydroxytryptamine, whereas it does not modify norepinephrine and gamma-aminobutyric acid levels in the hippocampus of young adult rats. Functional observational battery screening demonstrates that CHF2819 (1.5 and 4.5 mg/kg p.o.) does not affect activity, excitability, autonomic, neuromuscular, and sensorimotor domains, as well as physiological end points (body weight and temperature). However, this compound induces involuntary motor movements (ranging from mild tremors to myoclonic jerks) in a dose-dependent manner. These findings suggest that the anti-amnestic properties of CHF2819, together with its stimulatory effect on cholinergic and serotonergic functions, might have a therapeutic potential mainly for the symptomatic treatment of Alzheimer's disease patients in which the cognitive impairment is accompanied by a depressive syndrome. PMID- 10871312 TI - A molecular model of agonist and nonpeptide antagonist binding to the human V(1) vascular vasopressin receptor. AB - The affinity of the nonpeptide antagonist OPC-21268 is greater for the rat V(1) arginine vasopressin (AVP) receptor (V(1)R) than for the human V(1)R. Site specific mutagenesis was carried out to identify the residues that determine interspecies selectivity for nonpeptide antagonist binding. The introduction of rat amino acids in position 224, 310, 324, or 337 of the human V(1)R sequence dramatically altered OPC-21268 affinity for the receptor, whereas binding of AVP, the peptide V(1)R antagonist d(CH(2))(5)Tyr(Me)AVP, and the nonpeptide V(1)R antagonist SR49059 was not altered by these mutations. Computer modeling explained the mutagenesis results. Docking of OPC-21268 onto a homology-built model of the V(1)R receptor yielded a model for the bound ligand in which the hydrophobic part is deeply embedded in the transmembrane region, whereas the polar part is located on the surface of the extracellular side. The increased affinity of the G337A mutant is due to two additional van der Waals contacts of the alanine methyl group with carbon atoms on the antagonist. The I310V mutant reduces the hydrophobicity in the vicinity of the polar oxygen atom of the antagonist. The I224V mutant relieves overcrowding in a hydrophobic binding pocket involving the aromatic residues Trp(175), Phe(179), Phe(307), and Trp(304). Finally, the E324D mutant enables the formation of a hydrogen bond of the carboxylate side chain with the amide side chain of Gln(311), which in turn forms a hydrogen bond with the N57 nitrogen atom of OPC-21268. Thus, a few residues, distinct from those involved in agonist binding, control interspecies selectivity toward OPC-21268 nonpeptide antagonist binding. PMID- 10871313 TI - CB1 cannabinoid receptor-mediated cell migration. AB - Recent studies have suggested that cell migratory responses are often mediated by G(i) protein-coupled receptors. Because it is known that CB1 cannabinoid receptors are coupled to pertussis toxin-sensitive G proteins, we proposed that CB1 may mediate cell migration. To test this hypothesis, modified Boyden chamber assays were used to investigate cell migration mediated by CB1 cannabinoid receptors. HU-210, WIN55212-2, and anandamide, three cannabinoid agonists with distinct chemical structures, induced migration of human embryonic kidney 293 cells stably transfected with human CB1 gene, but not 293 cells transfected with an empty expression vector. These migratory responses were concentration dependent. The EC(50) values for HU-210, WIN55212-2, and anandamide were 0.19 +/- 0.04, 12. 2 +/- 1.4, and 39.9 +/- 3.7 nM, respectively. The maximal migration index for HU-210, WIN55212-2, and anandamide were 8.9 +/- 1.6, 9.5 +/- 1.6, and 8.8 +/- 1.3, respectively. Pretreating cells with 100 ng/ml pertussis toxin eliminated the cannabinoid agonist-induced cell migration. SR141716A, a selective antagonist for CB1, inhibited the cannabinoid agonist-induced migratory responses in a concentration-dependent manner. Checkerboard analysis demonstrated that anandamide-induced cell migrations are due to chemotaxis as well as chemokinesis. Furthermore, anandamide-induced migratory responses were inhibited, in a concentration-dependent manner, by PD098059, an inhibitor of mitogen-activated protein kinase activation, but not by 8-bromoadenosine-3',5'-cyclic monophosphate, a cell-permeable cAMP analog. These data demonstrate that cannabinoid agonists are able to induce chemotaxis and chemokinesis, and that these migratory responses are mediated by G protein-coupled, CB1 cannabinoid receptors. In addition, these data suggest that activation of mitogen-activated protein kinase plays an important role, whereas inhibition of adenylate cyclase is probably not involved in the cell migration mediated by CB1. PMID- 10871314 TI - Depletion of natriuretic peptide C receptors eliminates inhibitory effects of C type natriuretic peptide on evoked neurotransmitter efflux. AB - Natriuretic peptides suppress evoked catecholamine efflux by a mechanism attributed to activation of the natriuretic peptide receptor (NPR)-C, but this designation relies on the absolute specificity of truncated natriuretic peptide analogs for the NPR-C. The NPR-C involvement in evoked catecholamine efflux was defined better in this study by selectively ablating the NPR-C in pheochromocytoma cells with antisense oligodeoxynucleotides. This treatment suppressed NPR-C levels by 52 +/- 4% relative to missense treatment. The reduction of NPR-C levels suppressed evoked catecholamine efflux 33 +/- 6% and eliminated the effect of C-type natriuretic peptide to suppress evoked catecholamine efflux. The native peptide, C-type natriuretic peptide, reduced evoked catecholamine efflux 39 +/- 3% in cells with a normal complement of NPR-C. The NPR-C reduction failed to alter neuromodulatory effects of N-nitro-L-arginine methyl ester or an active fragment of the NPR-C receptor administered in permeabilized cells. Furthermore, the NPR-C reduction did not prevent guanylyl cyclase activation in response to C-type natriuretic peptide. These latter experiments indicate that the antisense treatment resulted in a specific suppression of the NPR-C and did not affect alternative neuromodulatory pathways or guanylyl cyclase receptors. The novel aspects of this study include both the inhibitory effect of NPR-C reduction on basal-evoked neurotransmitter efflux and the ablation of natriuretic peptide effects on neurotransmitter efflux by NPR-C reduction. The results are consistent with the notion of a key signal-transducing role of the NPR-C in mediating inhibitory effects of natriuretic peptides on neurotransmitter efflux. PMID- 10871315 TI - Effects of N(G)-monomethyl-L-arginine on Ca(2+) current and nitric-oxide synthase in rat ventricular myocytes. AB - The effects of N(G)-monomethyl-L-arginine (L-NMMA), a nitric-oxide synthase (NOS) inhibitor, on the L-type Ca(2+) current (ICa) and NO effects on NOS were determined in rat ventricular myocytes. L-NMMA (10 and 100 microM) had no significant effect on basal ICa, but in a cAMP-stimulated condition due to forskolin (1 microM) or milrinone (10 microM), a cGMP-inhibited cAMP phosphodiesterase (PDE), L-NMMA (10 and 100 microM) concentration dependently augmented ICa. The enhancing effects of L-NMMA (10 and 100 microM) on ICa were not seen in the presence of either a nonselective inhibitor of PDE, 3-isobutyl-1 methylxanthine (20 microM), resulting in a stimulated ICa condition or a cGMP dependent protein kinase activator, 8-bromo-cGMP (200 microM). 8-Bromo-cGMP (200 microM) inhibited 100 microM L-NMMA-induced ICa increase in the simultaneous application of forskolin (1 microM). Acetylcholine (ACh; 1 and 3 microM) inhibited 1 microM forskolin-stimulated ICa in a concentration-dependent manner, but this inhibitory action of ACh was significantly attenuated by the additional application of L-NMMA (100 microM). In the continuing presence of both L-NMMA (100 microM) and forskolin (1 microM), ACh (6 microM) had no inhibitory effect on ICa. In another series of experiments with isolated ventricular myocytes, we obtained both the positive staining of NADPH-diaphorase activity and the expression of the endothelial isoform of NOS. These data suggest that the effect of L-NMMA on ICa in a cAMP-stimulated condition with or without cholinergic inhibition is due to inhibition (acute effects) of a cGMP-stimulated cAMP-PDE via inhibition of the endothelial isoform of NOS. PMID- 10871316 TI - An investigation of the uroselective properties of four novel alpha(1a) adrenergic receptor subtype-selective antagonists. AB - The development of alpha(1a)-adrenergic receptor (AR) subtype-selective antagonists is likely to result in uroselective agents that effectively treat benign prostatic hyperplasia (BPH) symptoms without causing undesirable side effects that may be due to vascular alpha(1)-AR blockade. The properties of four aryl piperazine compounds (RWJ-38063, RWJ-68141, RWJ-68157, and RWJ-69736) are described in this report and compared with the properties of tamsulosin, an alpha(1)-AR antagonist that is used in the treatment of BPH. Radioligand binding studies show that all four RWJ compounds have significantly higher affinity for the alpha(1a)-AR subtype than for the alpha(1b) or alpha(1d) subtype and display a higher level of receptor subtype selectivity than tamsulosin. The RWJ compounds were more potent in inhibiting (+/-)-norepinephrine-induced contractions of isolated rat prostate tissue than those of isolated rat aorta tissue, whereas tamsulosin had the reversed tissue selectivity. RWJ-38063 and RWJ-69736 had the highest potency in the isolated prostate tissue assays of the four RWJ compounds, with pK(B) values of 8.24 and 9.26, respectively, and were 319- and 100-fold more potent in their effects on isolated prostate tissue than aorta tissue. The in vivo uroselectivities of RWJ-38063, RWJ-69736, and tamsulosin were examined in anesthetized dogs. Both RWJ compounds suppressed the intraurethral pressure response to phenylephrine to a greater extent than the mean arterial pressure response; however, RWJ-69736 also caused a marked transient rise in heart rate. Although less potent, RWJ-38063 and RWJ-69736 were notably more uroselective than tamsulosin in this canine model. PMID- 10871317 TI - Brainstem nicotinic receptor subtypes that influence intragastric and arterial blood pressures. AB - The purpose of this study was to investigate the effect of microinjection of nicotine and nicotinic receptor antagonists into the dorsal motor nucleus of the vagus (DMV) or medial subnucleus of the tractus solitarius (mNTS) on intragastric (IGP) and arterial blood pressures (BP) in anesthetized rats. Nicotine microinjected into the DMV (10-300 pmol) produced dose-related increases in IGP (ED(50) = 89 pmol); no significant changes were noted for BP. Ipsilateral vagotomy abolished nicotine-induced increases in IGP. Nicotine microinjected into the mNTS in a dose range of 0.1 to 300 pmol produced dose-related decreases in IGP (ED(50) = 0.6 pmol) and BP (ED(50) = 5.4 pmol). Bilateral vagotomy abolished nicotine-induced decreases in IGP while having no effect on BP. In rats treated with daily s.c. injections of nicotine (0.8 mg/kg of base) for 10 days, microinjections of nicotine into the DMV produced similar increases in IGP. BP responses from the mNTS were not affected by chronic treatment. However, nicotine microinjections into the mNTS no longer produced a decrease in IGP in these chronically treated animals. alpha-Bungarotoxin (100 pmol) significantly blocked nicotine-evoked increases in IGP from the DMV while having no effect on nicotine induced responses elicited from the mNTS. Hexamethonium (10 and 100 pmol) microinjected into the mNTS dose-dependently blocked nicotine-induced effects but did not interfere with the action of nicotine at the DMV. Our data indicate that nicotine-induced changes in IGP result from nicotine acting at two sites, the DMV and mNTS; and that at least three different nicotinic receptors in the dorsal medulla oblongata can influence gastrointestinal and cardiovascular function. PMID- 10871318 TI - Effects of cocaine self-administration on plasma corticosterone and prolactin in rats. AB - The effects of i.v. cocaine self-administration under "naturalistic" conditions on plasma corticosterone (CORT) and prolactin (PRL) were investigated in male Sprague-Dawley rats. After the determination of plasma CORT and PRL levels under basal conditions before access to cocaine for self-administration, rats were allowed to self-administer cocaine (0.25, 0.5, 1.0, or 2.0 mg/kg/infusion i.v.) by pressing a response lever under a continuous schedule of cocaine reinforcement during five daily consecutive 10-h sessions. Plasma CORT was significantly increased and plasma PRL was significantly reduced after each of the five self administration sessions. The effects of cocaine on plasma CORT were intake dependent, as demonstrated by significant positive correlations between postsession plasma CORT and total cocaine intake within the preceding sessions. The effects of cocaine on PRL were also intake-dependent but only on the first day of self-administration, on which a significant negative correlation was observed between cocaine intake and postsession PRL. In contrast, significant correlations between PRL and cocaine intake were not observed during any subsequent session, apparently reflecting adaptations to cocaine-induced PRL release. Alterations in neuroendocrine homeostasis emerged over time. Reductions in presession CORT values, as well as a persistent blunting of the diurnal CORT peak, were observed. Similarly, there was a modest but significant attenuation of plasma PRL when measured 4 days after the termination of cocaine self administration. Alterations in neuroendocrine function associated with self administration may be related to the development of cocaine dependence and could contribute to relapse in abstinent users. PMID- 10871319 TI - Angiotensin-converting enzyme-independent angiotensin formation in a human model of myocardial ischemia: modulation of norepinephrine release by angiotensin type 1 and angiotensin type 2 receptors. AB - Angiotensin II (Ang II) promotes norepinephrine (NE) release from cardiac sympathetic nerve endings. We assessed in a human model in vitro whether locally formed Ang II contributes to NE release in myocardial ischemia. Surgical specimens of human right atrium were incubated in anoxic conditions. After 70 min of anoxia, NE release (carrier-mediated; caused by NE transporter reversal) was 8 fold greater than normoxic release. Angiotensin-converting enzyme inhibition with enalaprilat failed to reduce anoxic NE release. In contrast, prevention of chymase-dependent Ang II formation with chymostatin, Bowman-Birk inhibitor, or alpha(1)-antitrypsin significantly inhibited anoxic, but not exocytotic, NE release. Two mast-cell stabilizers, cromolyn and lodoxamide, markedly reduced NE release, implicating cardiac mast cells as a major source of chymase. Angiotensin type 1 receptor (AT(1)R) blockade with EXP3174 inhibited NE release, whereas angiotensin type 2 receptor (AT(2)R) blockade with PD123319 did not. Interestingly, PD123319 reversed the inhibitory effect of EXP3174. Furthermore, synergisms were uncovered between EXP3174 and an AT(2)R agonist, and between EXP3174 and a Na(+)/H(+) exchanger inhibitor. Thus, angiotensin-converting enzyme independent Ang II formation via chymase is important for carrier-mediated ischemic NE release in the human heart. Locally generated Ang II promotes NE release by acting predominantly at AT(1)Rs, which are likely coupled to the Na(+)/H(+) exchanger. Effects of Ang II at AT(2)Rs, seemingly opposite to those resulting from AT(1)R activation, are uncovered when AT(1)Rs are blocked. Because NE release is associated with coronary vasoconstriction and arrhythmias, and mast cell density and chymase content increase in the ischemic heart, the notion that chymase-generated Ang II plays a major role in carrier-mediated NE release may have important clinical implications. PMID- 10871320 TI - Microinjection of nociceptin (Orphanin FQ) into nucleus tractus solitarii elevates blood pressure and heart rate in both anesthetized and conscious rats. AB - The role of nociceptinergic transmission in the nucleus tractus solitarii (NTS) in the central modulation of cardiovascular activity was investigated in pentobarbital-anesthetized and conscious rats. Pharmacological activation of nociceptin receptors with a unilateral injection of synthetic nociceptin into the NTS, wherein injection of L-glutamate (1 nmol) caused typical depressor responses, elevated blood pressure and heart rate (HR) in most of the anesthetized rats. The elevation of blood pressure and HR by nociceptin was dose dependent (0.04, 0.2, and 1 nmol) with a threshold dose of 0.2 nmol. At 1 nmol, changes in blood pressure and HR were evident at 5 min, and remained for 45 min after injection. Pretreatment with the selective nociceptin receptor antagonist nocistatin (1 nmol) into the NTS abolished the nociceptin-induced hypertension and tachycardia. In contrast, the nonselective opioid receptor antagonist naloxone (5 nmol) did not modify the cardiovascular responses to nociceptin. Intra-NTS injection of nocistatin (0.04 and 1 nmol) and naloxone alone had no significant effect on baseline blood pressure and HR. In chronically cannulated and conscious rats, similar pressor and tachycardic responses were induced by intra-NTS injection of 1 nmol of nociceptin. However, changes in blood pressure and HR were rapid, and quickly returned to normal levels within 10 min. These data suggest that the newly discovered nociceptinergic transmission in the NTS has a powerful influence on peripheral hemodynamic activity. This influence is inhibitory and may not be tonically active under normal physiological conditions. Moreover, the cardiovascular responses to exogenous nociceptin were mediated through activation of specific nociceptin receptors rather than typical naloxone sensitive opioid receptors. PMID- 10871321 TI - Metabolism of bradykinin In vivo in humans: identification of BK1-5 as a stable plasma peptide metabolite. AB - Studies investigating the role of bradykinin in disease states such as hypertension, sepsis, and asthma have been confounded by difficulties in measuring the concentration of this short-lived peptide. The purpose of this study was to determine a stable metabolite of bradykinin in the systemic circulation of humans. Bradykinin (containing trace concentrations of [(3)H]bradykinin) was administered i.v. into three human volunteers in increasing amounts up to a maintenance rate of 200 ng/kg/min until a total dose of 1 mg was given. Metabolic products were purified and identified by HPLC and by electrospray ionization mass spectrometry. Infused bradykinin was rapidly degraded, such that no exogenous bradykinin was detected in venous plasma sampled during infusion. BK1-5 (Arg-Pro-Pro-Gly-Phe), the 1-to-5 amino acid fragment of bradykinin, was identified as a major stable plasma metabolite of bradykinin. Plasma concentrations of BK1-5 correlated with dose of bradykinin infused and concentrations at the end of bradykinin infusion were 1510 to 4600 fmol/ml of blood. BK1-5 was cleared from blood with a terminal half-life of 86 to 101 min. Thus, in humans, bradykinin is rapidly degraded in vivo to BK1-5, a stable metabolite. Measurement of this metabolite could provide a tool to assess pathophysiologic and pharmacologic alterations in systemic bradykinin generation associated with human disease. PMID- 10871322 TI - Nitric oxide from enteric nerves acts by a different mechanism from myogenic nitric oxide in canine lower esophageal sphincter. AB - In canine lower esophageal sphincter, myogenic constitutive nitric-oxide (NO) synthase (NOS) in plasma membrane limits tone by opening large conductance Ca(2+) dependent K(+) channels (BK(Ca) channels) and hyperpolarizing the membrane. We examined whether K(V) channels were involved and whether NO from enteric nerves and from NO donors used the same mechanisms. With nerves inactive, 100 nM iberiotoxin, like N-nitro-L-arginine (L-NOARG), increased tone but less. 4 Aminopyridine (4-AP) at 5 mM behaved similarly. Tetraethyl ammonium (TEA) at 20 mM equaled the effect of L-NOARG and occluded any tone increase from any combination of these agents. More than iberiotoxin or 4-AP, TEA decreased relaxations in response to sodium nitroprusside (SNP) or 3-morpholino-sydnonimine (Sin-1) by approximately 50%. In whole-cell patch-clamp recordings, TEA and 4-AP reduced outward K(+) currents additively by >90% at depolarization of +90 mV. Thus, K(+) channels in addition to BK(Ca) channels are opened by myogenic NO, and exogenous NO had relaxing effects both related and unrelated to K(+) channel openings. TEA (20 mM) increased tone but did not inhibit relaxations to electrical field stimulation (EFS) of enteric nerves. 4-AP relaxed tone, an effect that was abolished and reversed by L-NOARG. 4-AP apparently released NO and acetylcholine from nerves. The putative Cl(-) channel blocker niflumic acid (NFA; 30-100 microM) dose dependently reduced tone, but tone, restored by 10(-6) M carbachol or 20 mM TEA, was still relaxed by EFS and by SNP. 4,4' Diisothiocyanatostilbene-2, 2'-disulfonic acid (DIDS) at 500 to 1000 microM did not inhibit relaxation to EFS or SNP. The addition of TEA (20 mM) to DIDS (1000 microM) induced tonic and phasic activity and markedly inhibited relaxations to EFS. DIDS plus TEA reduced the relaxations to SNP like TEA alone. Reduction in extracellular ?Cl(-) by isethionate substitution reduced tone but did not reduce relaxations when tone was restored. The combination of reduced extracellular ?Cl( ) and TEA did not abolish relaxation to EFS until DIDS was added. Thus, multiple K(+) channels are opened by myogenic NO, and openings of these channels, as well as DIDS-sensitive, undefined mechanisms, are induced when NO is released from nerves or SNP. PMID- 10871323 TI - Effect of lipoteichoic acid on dermal vascular permeability in mice. AB - Lipoteichoic acid (LTA), the cell wall component of Gram-positive bacteria, has been shown to cause inflammatory responses comparable to lipopolysaccharide (LPS) of Gram-negative bacteria. This study examined the activity of LTA to induce dermal microvascular permeability changes in mice. Vascular permeability was assessed by extravasation of Pontamine sky blue. Subcutaneous injection of LTA (200-400 microg/site) in mice that were preinjected i.v. with the dye increased local dye leakage in the skin at 1 to 3 h. The LTA-induced dye leakage was inhibited by indomethacin, valeryl salicylate, diphenhydramine, and a platelet activating factor antagonist but not by inhibitors of nitric-oxide synthase, cyclooxygenase-2, or guanylate cyclase or by antibodies against tumor necrosis factor-alpha or interleukin-1alpha. LTA induced comparable increases in dye leakage in inducible nitric-oxide synthase-deficient mice and wild-type controls. Pretreatment of normal mice with i.v. LTA did not confer tolerance to LTA- or LPS induced dye leakage. In contrast, systemic LPS administration induced tolerance against subsequent challenge with LPS but not LTA. Serum corticosterone levels, which were suggested to induce tolerance, were not increased by LTA pretreatment but were increased by LPS. Thus, LTA increases dermal microvascular permeability in mice. Among the inflammatory mediators, eicosanoids, platelet-activating factor, and histamine mediate the effect of both LTA and LPS, whereas nitric oxide, tumor necrosis factor-alpha, and interleukin-1alpha may not play a major role in LTA-induced dye leakage. The difference between LTA and LPS to stimulate corticosterone may partially explain the failure of LTA to induce tolerance against vascular dye leakage. PMID- 10871324 TI - Mechanisms of N-methyl-D-aspartate-induced apoptosis in phencyclidine-treated cultured forebrain neurons. AB - Chronic administration of phencyclidine (PCP) to rats has been demonstrated to produce a sensitized locomotor response to PCP challenge that is associated with apoptotic cell death and an up-regulation of the N-methyl-D-aspartate (NMDA) receptor. To determine the underlying mechanisms, dissociated forebrain cultures were treated for 2 days with 3 microM PCP. After washout of PCP, NMDA was added (in the presence of Mg(2+)) for 20 h. The uptake of a vital dye and the release of lactate dehydrogenase measured cell viability. Apoptosis was assessed by an enzyme-linked immunosorbent assay that was specific for fragmented (histone associated) DNA and an in situ assay for nicked DNA, terminal dUTP nick-end labeling. These assays showed that the effect of a nontoxic concentration of NMDA (30 microM) became lethal to approximately one-third of the neurons after chronic (48-h) PCP treatment. This treatment also resulted in a 47% increase in NR1 subunit mRNA, suggesting that NMDA-induced neuronal cell death after chronic PCP is due to NMDA receptor up-regulation. Furthermore, exposure of PCP-treated cultures to NMDA led to increased expression of Bax and decreased expression of Bcl-X(L). The Bcl-X(L)/Bax ratio was markedly decreased by 30 microM NMDA in the PCP-treated, but not control, cultures. Addition of superoxide dismutase and catalase prevented the decrease in Bcl-X(L)/Bax. This study suggests that NMDA induced changes in Bax and/or Bcl-X(L) involve the formation of reactive oxygen species. By extrapolation, these data suggest that PCP-induced apoptosis in vivo may involve similar mechanisms and that cultured neurons may be a suitable model for the mechanistic study PCP toxicity in vivo. PMID- 10871325 TI - Coadministration of 5-hydroxytryptamine(1A) antagonist WAY-100635 prevents fluoxetine-induced desensitization of postsynaptic 5-hydroxytryptamine(1A) receptors in hypothalamus. AB - Treatment with selective serotonin reuptake inhibitors induces a desensitization of hypothalamic postsynaptic 5-hydroxytryptamine (5-HT)(1A) receptors in humans and rats. This study investigated whether fluoxetine-induced desensitization is due to overactivation of postsynaptic 5-HT(1A) receptors; whether blockade of somatodendritic 5-HT(1A) autoreceptors accelerates this desensitization; and whether desensitization is associated with a reduction of Gz proteins, which couple to 5-HT(1A) receptors. WAY-100635 was tested at low doses (0.03-0.3 mg/kg), which antagonize somatodendritic 5-HT(1A) autoreceptors in the raphe nuclei, and at a higher dose (1 mg/kg), which completely blocks postsynaptic 5 HT(1A) receptors. Plasma levels of oxytocin and adrenal corticotrophic hormone (corticotropin) were measured as peripheral indicators of hypothalamic 5-HT(1A) receptor function. Daily injections of fluoxetine (10 mg/kg/day i.p.) for 2 days did not desensitize 5-HT(1A) receptors but three daily injections of fluoxetine produced a partial desensitization of the hormone responses to (+/-)-8-hydroxy-2 dipropylaminoetetralin (50 microg/kg s.c.). WAY-100635 (0.03-0.3 mg/kg) did not accelerate or potentiate the fluoxetine-induced desensitization of 5-HT(1A) receptors. However, WAY-100635 at a dose that completely blocks postsynaptic 5 HT(1A) receptors (1.0 mg/kg) completely prevented the fluoxetine-induced desensitization of 5-HT(1A) receptors. These data demonstrate that at least 3 days of fluoxetine exposure is required to produce a homologous desensitization of hypothalamic 5-HT(1A) receptors. Although previous studies indicate that injections of fluoxetine for 14 days produce a reduction of Gz protein levels in the hypothalamus, the levels of Gz proteins were not affected by either fluoxetine or WAY-100635. Alternative mechanisms mediating the initial stages of 5-HT(1A) receptor desensitization could involve post-translational modifications in the 5-HT(1A) receptor-Gz protein-signaling cascade. PMID- 10871326 TI - Multiple cellular mechanisms mediate the effect of lobeline on the release of norepinephrine. AB - The complex effect of lobeline on [(3)H]norepinephrine ([(3)H]NE) release was investigated in this study. Lobeline-induced release of [(3)H]NE from the vas deferens was strictly concentration-dependent. In contrast, electrical stimulation-evoked release was characterized by diverse effects of lobeline depending on the concentration used: at lower concentration (10 microM), it increased the release and at high concentration (100 and 300 microM), the evoked release of [(3)H]NE was abolished. The effect of lobeline on the basal release was [Ca(2+)]-independent, insensitive to mecamylamine, a nicotinic acetylcholine receptor antagonist, and to desipramine, a noradrenaline uptake inhibitor. However, lobeline-induced release was temperature-dependent: at low temperature (12 degrees C), at which the membrane carrier proteins are inhibited, lobeline failed to increase the basal release. Lobeline dose dependently inhibited the uptake of [(3)H]NE into rat hippocampal synaptic vesicles and purified synaptosomes with IC(50) values of 1.19 +/- 0.11 and 6.53 +/- 1.37 microM, respectively. Lobeline also inhibited Ca(2+) influx induced by KCl depolarization in sympathetic neurons measured with the Fura-2 technique. In addition, phenylephrine, an alpha(1)-adrenoceptor agonist, contracted the smooth muscle of the vas deferens and enhanced stimulation-evoked contraction. Both effects were inhibited by lobeline. Our results can be best explained as a reversal of the monoamine uptake by lobeline that is facilitated by the increased intracellular NE level after lobeline blocks vesicular uptake. At high concentrations, lobeline acts as a nonselective Ca(2+) channel antagonist blocking pre- and postjunctional Ca(2+) channels serving as a counterbalance for the multiple transmitter releasing actions. PMID- 10871327 TI - Effect of methoxychlor administration to male rats on hepatic, microsomal iodothyronine 5'-deiodinase, form I. AB - We previously reported that methoxychlor administration inhibits the activity of the hepatic, microsomal iodothyronine 5'-deiodinase, form I (ID-I; ). Our data further suggested that the inhibition was due to the covalent binding of a methoxychlor metabolite to a 56-kDa protein identified as ID-I (; ). This protein is 98% homologous to the thiol:protein disulfide oxidoreductase, form Q5 (ERp55;; ). Although at the time there was some controversy, most studies now suggest that ID-I is actually catalyzed by a 27-kDa selenoprotein that does not form adducts with methoxychlor (;; ). Because the 27-kDa protein is considered to be ID-I instead of ERp55, we have further examined the basis for the decreased ID-I activity observed after methoxychlor administration. Male, 150- to 200-g Sprague Dawley rats were given methoxychlor (0-100 mg/kg/day) in corn oil by gavage for 14 days. ID-I was determined by a thyronine-specific immunoassay. Treated rats showed a significant 15% decline in total hepatic, microsomal protein at all doses. The ID-I-specific activity showed a linear decrease with increasing log doses of methoxychlor. The maximum decrease was 42% at 100 mg/kg/day. The 27-kDa protein specific content declined 37%. In rats given methoxychlor the ratios of the 27-kDa protein mRNA to the 18S ribosomal RNA declined from 2.2 +/- 0.27 x 10( 3) (controls) to 0.99 +/- 0.09 x 10(-3) (100 mg/kg/day). These data suggest that the decreased ID-I observed with chronic methoxychlor administration was due to decreased transcription or stability of the mRNA encoding the 27-kDa protein. PMID- 10871328 TI - Persistent expression of 3-methylcholanthrene-inducible cytochromes P4501A in rat hepatic and extrahepatic tissues. AB - We reported earlier that 3-methylcholanthrene (MC) persistently induces hepatic ethoxyresorufin O-deethylase activities (CYP1A1) in rats for up to 45 days. In this investigation, we tested the hypotheses that persistent expression of CYP1A1 activities is paralleled by sustained induction of CYP1A1/CYP1A2 apoproteins and their mRNAs and that this phenomenon is mediated by mechanisms other than retention of MC in the rat. Rats were given MC (93 micromol/kg) i.p., once daily for 4 days, and CYP1A1/1A2 parameters were measured in liver at selected time points. MC-elicited increases in CYP1A1/1A2 activities, apoprotein contents, and mRNA levels were sustained for several weeks after the last dose of MC treatment. MC also caused long-term induction of CYP1A1 in lungs and mammary glands. Rats treated with [(3)H]MC once daily for 4 days excreted 92. 3% of the administered radioactivity in feces and urine by day 15. The intrahepatic concentration of MC at the 15-day time point was 270 pmol/g. Dose-response studies showed that administration of MC (2 micromol/kg), which produced an intrahepatic concentration of 271 pmol/g after 24 h, did not induce CYP1A1/1A2 activities, strongly suggesting that the sustained induction of CYP1A1/1A2 was not due to retention of the parent MC in the body. Electrophoretic mobility shift assays revealed that persistent CYP1A1 induction by MC involved Ah receptor-independent mechanisms. In conclusion, our results support the hypothesis that persistent expression of CYP1A1/1A2 by MC is mediated by mechanisms independent of the retention of the parent carcinogen. PMID- 10871329 TI - Tissue distribution and clinical monitoring of the novel macrolide immunosuppressant SDZ-RAD and its metabolites in monkey lung transplant recipients: interaction with cyclosporine. AB - We report the tissue distribution and clinical monitoring of the novel macrolide immunosuppressant SDZ-RAD ?40-O-(2-hydroxyethyl)-rapamycin and its metabolites in monkey lung transplant recipients as well as its interaction with cyclosporine as the Neoral formulation. After left unilateral lung transplantation, cynomolgus monkeys received by oral administration either 1) 1.5 mg/kg/day SDZ-RAD (n = 4); 2) 100 mg/kg/day cyclosporine (n = 4); 3) 0.3 mg/kg/day SDZ-RAD + 100 mg/kg/day cyclosporine (n = 6); 4) 1.5 mg/kg/day SDZ-RAD + 50 mg/kg/day cyclosporine (n = 5); or 5) SDZ-RAD and cyclosporine doses adjusted according to trough blood concentration measurements (n = 6). At the end of the observation period (usually 29 days after transplantation), and 24 h after the last doses, tissue samples were collected and analyzed with HPLC/mass spectrometry. Gall bladder, pancreas, the transplant lung, cerebellum, kidneys, and spleen had the highest SDZ-RAD concentrations. Coadministration of cyclosporine increased SDZ-RAD concentrations in most tissues as well as tissue-to-blood distribution coefficients. In contrast, SDZ-RAD had only a small effect on cyclosporine blood and tissue concentrations. Rejection in lung grafts in monkeys treated with either of the cyclosporine/SDZ-RAD combinations was significantly less than in the monotherapy groups (P <.002). Histological rejection scores were inversely correlated with SDZ-RAD concentrations in blood (r = -0. 68; P <.001; n = 24), lymph nodes (P = 0.58; P <.003; n = 24), thymus (r = -0.63; P <.001; n = 23) and transplant lung tissue (r = -0.58; P <.003; n = 24). We conclude that, in addition to the synergistic pharmacodynamic interaction, a pharmacokinetic interaction resulting in higher SDZ-RAD tissue concentrations contributed to the significantly better immunosuppressive efficacy when both drugs were combined compared with monotherapy. PMID- 10871330 TI - Resveratrol modulates arachidonic acid release, prostaglandin synthesis, and 3T6 fibroblast growth. AB - Previous results suggested that the cyclooxygenase-2 pathway and prostaglandins might modulate 3T6 fibroblast growth. This study shows the effect of resveratrol on the main elements of arachidonic acid (AA) cascade and 3T6 fibroblast growth. The polyphenol reduced the reactive oxygen species production stimulated by fetal calf serum or platelet-derived growth factor, as well as phospholipase A(2) activity translocation and the subsequent [(3)H]AA release and prostaglandin E(2) synthesis induced by these growth factors. A Western blot analysis demonstrated that cyclooxygenase-2 induction stimulated by fetal calf serum or platelet derived growth factor was inhibited by resveratrol. The effects of resveratrol on AA cascade were correlated with an impairment of 3T6 fibroblast proliferation and DNA synthesis. These results suggest that reactive oxygen species and AA, and/or prostaglandins such as prostaglandin E(2) might be involved in the control of 3T6 fibroblast growth by resveratrol. PMID- 10871331 TI - Inhibitory effects of vesnarinone on cloned cardiac delayed rectifier K(+) channels expressed in a mammalian cell line. AB - Vesnarinone, a phosphodiesterase inhibitor, prolongs cardiac action potential duration by inhibiting the delayed rectifier K(+) current, I(K). We examined the effect of this agent on human ether-a-go-go related gene (HERG) and KvLQT1/minK K(+) channels heterologously expressed in human embryonic kidney 293T cells with the whole-cell patch-clamp technique. HERG channel current was inhibited by vesnarinone in a concentration-dependent manner, whereas KvLQT1/minK current was hardly affected by the drug. The inhibition of HERG current by vesnarinone became more prominent and faster as the membrane potential was more depolarized. The properties of inhibition could be described by a first order reaction between the drug and the channel that was apparently independent of HERG channel gating. Although the unbinding rate constant of the drug was constant, the apparent binding rate constant increased as the membrane was more depolarized and the drug concentration was raised. This model also could explain the fast recovery from the drug's effect at hyperpolarized potentials and its rate-dependent inhibition of HERG. Therefore, the effect of vesnarinone on the HERG-K(+) current could be adequately described by a simple kinetic model of drug-channel interaction. PMID- 10871332 TI - Quantification of pharmacodynamic interactions between dexmedetomidine and midazolam in the rat. AB - The pharmacodynamic (PD) interaction between the benzodiazepine agonist midazolam and the alpha(2)-adrenergic agonist dexmedetomidine was characterized for defined measures of anesthetic action and cardiovascular and ventilatory side effects in 33 rats. For various combinations of constant plasma concentrations of midazolam (0.1-20 microg/ml) and dexmedetomidine (0.3-19 ng/ml) obtained by target controlled infusion, the whisker reflex (WR), righting reflex (RR), startle reflex to noise (SR), tail clamp response (TC), and corneal reflex (CR) were assessed. EEG (power in 0.5-3.5-Hz frequency band), mean arterial pressure, and heart rate were recorded continuously. Blood gas values and arterial drug concentrations were determined regularly. The nature and extent of PD interaction was quantified by the model parameter synergy (SYN < 0, antagonism; SYN = 0, additivity; and SYN > 0, synergy). With increasing drug concentrations WR was lost first, followed by RR, SR, TC, and CR. These effects were accompanied by an increase of the EEG measure. The drug interaction was synergistic for all stimulus-response measures and the degree of synergy increased with deeper levels of central nervous system depression (SYN was 7.3, 145, 560, 374, and 1490 for WR, RR, SR, TC, and CR, respectively). The cardiovascular side effects of dexmedetomidine, evaluated at similar PD endpoints, were reduced in the presence of midazolam. Ventilatory side effects were minor for all drug combinations. The nature and extent of the PD interactions were not reflected in the EEG measure. PMID- 10871333 TI - Characterization of the alpha(2)-adrenoceptor subtype, which functions as alpha(2)-autoreceptor in human neocortex. AB - The pharmacological properties of the alpha(2)-adrenergic receptors regulating the release of norepinephrine were investigated in human neocortex. Slices were preincubated with [(3)H]norepinephrine, superfused under blockade of transmitter reuptake, and stimulated electrically. First, the autoinhibitory circuit of [(3)H]norepinephrine release was analyzed quantitatively by estimation of the K(d) of norepinephrine at the alpha(2)-autoreceptor (10(-7.99) M), the concentration of the endogenous transmitter causing this autoinhibition at a stimulation frequency of 3 Hz (10(-7.61) M), and the maximum inhibition obtainable through the autoreceptor (83%). Second, antagonist pK(b) values of nine antagonists were determined by using their pEC(50) values (negative logarithms of antagonist concentrations that increased the electrically evoked overflow of tritium by 50%) against the release-inhibiting effect of the endogenous transmitter. When compared with binding or functional data from the literature, the pK(b) values correlated best with the antagonist affinities at alpha(2A) binding sites. In contrast, the correlations with alpha(2B), alpha(2C), and alpha(2D) sites were not as good. It is concluded that in human neocortex prejunctional autoreceptors are alpha(2A). PMID- 10871334 TI - Nicotinic acetylcholine receptors at sites of neurotransmitter release to the guinea pig intestinal circular muscle. AB - Experiments were designed to test the hypothesis that nicotinic acetylcholine receptors (nAChRs) are present at sites of neurotransmission to the guinea pig ileum circular smooth muscle. Circular smooth muscle preparations, from which the myenteric plexus had been removed (circular muscle-axon preparation), were used for this purpose. Nicotine and dimethylphenyl piperazinium iodide (10-100 microM) induced contraction of the circular smooth muscle. Agonist-induced contraction was inhibited by 1 microM scopolamine and abolished in the combined presence of 1 microM scopolamine and 0. 3 microM CP 96,345-01, a neurokinin-1 receptor antagonist. Contractions induced by electric field stimulation (30 pulses, 0.5 ms, 70 V, 10 Hz) were abolished by 0.3 microM tetrodotoxin (TTX); in contrast, agonist-induced contraction was attenuated but not abolished by 0.3 microM TTX. Mecamylamine (3 or 30 microM), an nAChR antagonist, blocked agonist-induced contractions. Frequency-response curves for both "ON and "OFF electric field stimulation contractions were abolished by the combined presence of 1 microM scopolamine and 0.3 microM CP 96,345-01 or by 0.3 microM TTX. At stimulation frequencies greater than 2 Hz, the ON contraction was increased in the presence of 100 microM nitro-L-arginine. Mecamylamine (3 microM) was used to block the stimulatory prejunctional nAChRs located near sites of neurotransmitter release to the circular smooth muscle; however, ON and OFF contractions were not affected by mecamylamine. Although the prejunctional nAChRs are not targets for endogenously released acetylcholine under the conditions tested here, these receptors may be targets for the development of new prokinetic agents. PMID- 10871335 TI - Biospecific interaction analysis (BIA) of low-molecular weight DNA-binding drugs. AB - DNA-binding drugs have been reported to be able to interfere with the activity of transcription factors in a sequence-dependent manner, leading to alteration of transcription. This and similar effects could have important practical applications in the experimental therapy of many human pathologies, including neoplastic diseases and viral infections. The analysis of the biological activity of DNA-binding drugs by footprinting, gel retardation, polymerase chain reaction, and in vitro transcription studies does not allow a real time study of binding to DNA and dissociation of the generated drugs/DNA complexes. The recent development of biosensor technologies for biospecific interaction analysis (BIA) enables monitoring of a variety of molecular reactions in real-time by surface plasmon resonance (SPR). In this study, we demonstrate that molecular interactions between DNA-binding drugs (chromomycin, mithramycin, distamycin, and MEN 10567) and biotinylated target DNA probes immobilized on sensor chips is detectable by SPR technology using a commercially available biosensor. The target DNA sequences were synthetic oligonucleotides mimicking the Sp1, NF-kB, and TFIID binding sites of the long terminal repeat of the human immunodeficiency type 1 virus. The results obtained demonstrate that mithramycin/DNA complexes are less stable than chromomycin/DNA complexes; distamycin binds to both NF-kB and TATA box oligonucleotides, but distamycin/(NF-kB)DNA complexes are not stable; the distamycin analog MEN 10567 binds to the NF-kB mer and the generated drug/DNA complexes are stable. The experimental approach described in this study allows fast analysis of molecular interactions between DNA-binding drugs and selected target DNA sequences. Therefore, this method could be used to identify new drugs exhibiting differential binding activities to selected regions of viral and eukaryotic gene promoters. PMID- 10871336 TI - Neuroprotective efficacy and therapeutic window of the high-affinity N-methyl-D aspartate antagonist conantokin-G: in vitro (primary cerebellar neurons) and in vivo (rat model of transient focal brain ischemia) studies. AB - Conantokin-G (Con-G), a 17-amino-acid peptide derived from marine snails and a potent N-methyl-D-aspartate (NMDA) antagonist, was evaluated for its neuroprotective properties in vitro and in vivo. In primary cerebellar neurons, Con-G was shown to decrease excitotoxic calcium responses to NMDA and to exhibit differential neuroprotection potencies against hypoxia/hypoglycemia-, NMDA-, glutamate-, or veratridine-induced injury. Using the intraluminal filament method of middle cerebral artery occlusion as an in vivo rat model of transient focal brain ischemia, the neuroprotective dose-response effect of Con-G administration beginning 30 min postocclusion was evaluated after 2 h of ischemia and 22 h of reperfusion. In the core region of injury, an 89% reduction in brain infarction was measured with significant neurological and electroencephalographic recovery at the maximal dose tested (2 nmol), although mild sedation was noted. Lower doses of Con-G (0.001-0.5 nmol) were significantly neuroprotective without causing sedation. Postinjury time course experiments demonstrated a therapeutic window out to at least 4 to 8 h from the start of the injury, providing a 47% reduction in core injury. The neuroprotective effect of Con-G (0. 5 nmol) was also evaluated after 72 h of injury, where a 54% reduction in core brain infarction was measured. Critically, in both recovery models (i.e., 24 and 72 h), the reduction in brain infarction was associated with significant improvements in neurological and electroencephalographic recovery. These data provide evidence for the potent and highly efficacious effect of Con-G as a neuroprotective agent, with an excellent therapeutic window for the potential intervention against ischemic/excitotoxic brain injury. PMID- 10871337 TI - Detoxication of vinca alkaloids by human P450 CYP3A4-mediated metabolism: implications for the development of drug resistance. AB - Vinca alkaloids are important chemotherapeutic agents, and their pharmacokinetic properties display significant interindividual variations, possibly due to CYP3A4 mediated metabolism. We have evaluated the relevance of this metabolism for the chemotherapeutic and the toxicological properties of these drugs. Analysis was performed using Chinese hamster ovary cell lines that expressed either CYP2D6 or CYP3A4. The latter cells metabolized vinblastine with a turnover number of 0.4 min(-1), resulting in a decreased cytotoxicity of this compound. Whereas vincristine and vinblastine at a concentration of 100 nM killed more than 90% of the parental cells, more than 50 and 35%, respectively, of cells that coexpressed CYP3A4 and cytochrome P450 (P450) reductase survived these treatments. No additional increase in cytotoxicity was noted above 100 nM. Similarly, preincubation of vinblastine with bacterial membranes that contained recombinant CYP3A4 and P450 reductase decreased the cytotoxicity of vinblastine for parental Chinese hamster ovary cells. We also demonstrate that the presence of vinblastine in a coculture of cells that expressed beta-galactosidase together with cells that expressed CYP3A4 strongly selected for the latter cells, resulting in an increased level of CYP3A4 in the surviving cell population. Similarly, treatment of the human colon adenocarcinoma cell line LS174T with vinblastine selected for a cell population with higher levels of endogenous CYP3A4 as revealed by immunohistochemistry without simultaneous increase of multidrug resistance protein 1 (MDR1). This is the first evidence that tumor P450s have the potential to contribute to the development of drug resistance during chemotherapy. PMID- 10871338 TI - Cardiac peroxynitrite formation and left ventricular dysfunction following doxorubicin treatment in mice. AB - Selective cardiotoxicity of doxorubicin remains a significant and dose-limiting clinical problem. The mechanisms involved have not been fully defined but may involve the production of reactive oxygen species and/or alteration of cardiac energetics. Here, we tested the hypotheses that doxorubicin causes left ventricular dysfunction in mice and is associated with dysregulation of nitric oxide in cardiac tissue, leading to the accumulation of 3-nitrotyrosine (a biomarker of peroxynitrite formation). Animals were dosed with doxorubicin (20 mg/kg i.p.), and left ventricular performance was assessed in vivo using M-mode and Doppler echocardiography. Five days after doxorubicin administration, left ventricular fractional shortening, cardiac output, and stroke volume parameters were significantly reduced relative to control values (30.0 +/- 3.6 versus 46.1 +/- 1. 6%, 8.9 +/- 0.9 versus 11.5 +/- 0.6 ml/min, and 21.2 +/- 0.1 versus 29.5 +/- 0.1 microl for doxorubicin versus control, P <.05). Statistically significant (P <.05) increases in the immunoprevalence of myocardial inducible nitric oxide synthase (33 +/- 18 versus 9 +/- 2%, via quantitative image analysis) and 3 nitrotyrosine formation (56 +/- 24 versus 0.3 +/- 0.4%) were also observed after doxorubicin. Correlation analyses revealed a highly significant inverse relationship between left ventricular fractional shortening and cardiac 3 nitrotyrosine immunoprevalence (P <.01). No such relationship was observed for inducible nitric oxide synthase. Western blot analyses of cardiac myofibrillar fractions revealed extensive nitration of an abundant 40-kDa protein, shown to be the myofibrillar isoform of creatine kinase. These data demonstrate that alteration of cardiac nitric oxide control and attendant peroxynitrite formation may be an important contributor to doxorubicin-induced cardiac dysfunction. Furthermore, nitration of key myofibrillar proteins and alteration of myocyte energetics are implicated. PMID- 10871339 TI - Meeting report of the ASPET-Ray Fuller Symposium: insulin resistance in diabetes and hypertension: syndrome X and beyond. PMID- 10871340 TI - Substitution of Asp-210 in HAP1 (APE/Ref-1) eliminates endonuclease activity but stabilises substrate binding. AB - HAP1, also known as APE/Ref-1, is the major apurinic/apyrimidinic (AP) endonuclease in human cells. Previous structural studies have suggested a possible role for the Asp-210 residue of HAP1 in the enzymatic function of this enzyme. Here, we demonstrate that substitution of Asp-210 by Asn or Ala eliminates the AP endonuclease activity of HAP1, while substitution by Glu reduces specific activity approximately 500-fold. Nevertheless, these mutant proteins still bind efficiently to oligonucleotides containing either AP sites or the chemically unrelated bulky p-benzoquinone (pBQ) derivatives of dC, dA and dG, all of which are substrates for HAP1. These results indicate that Asp-210 is required for catalysis, but not substrate recognition, consistent with enzyme kinetic data indicating that the HAP1-D210E protein has a 3000-fold reduced K(cat )for AP site cleavage, but an unchanged K(m). Through analysis of the binding of Asp-210 substitution mutants to oligonucleotides containing either an AP site or a pBQ adduct, we conclude that the absence of Asp-210 allows the formation of a stable HAP1-substrate complex that exists only transiently during the catalytic cycle of wild-type HAP1 protein. We interpret these data in the context of the structure of the HAP1 active site and the recently determined co-crystal structure of HAP1 bound to DNA substrates. PMID- 10871341 TI - Isolation of an essential Schizosaccharomyces pombe gene, prp31(+), that links splicing and meiosis. AB - We carried out a screen for mutants that arrest prior to premeiotic S phase. One of the strains we isolated contains a temperature-sensitive allele mutation in the fission yeast prp31(+) gene. The prp31-E1 mutant is defective in vegetative cell growth and in meiotic progression. It is synthetically lethal with prp6 and displays a pre-mRNA splicing defect at the restrictive temperature. We cloned the wild-type gene by complementation of the temperature-sensitive mutant phenotype. Prp31p is closely related to human and budding yeast PRP31 homologs and is likely to function as a general splicing factor in both vegetative growth and sexual differentiation. PMID- 10871342 TI - Characterization of an ATP-dependent DNA ligase from the thermophilic archaeon Methanobacterium thermoautotrophicum. AB - We report the production, purification and characterization of a DNA ligase encoded by the thermophilic archaeon Methanobacterium thermoautotrophicum. The 561 amino acid MTH: ligase catalyzed strand-joining on a singly nicked DNA in the presence of a divalent cation (magnesium, manganese or cobalt) and ATP (K(m) 1.1 microM). dATP can substitute for ATP, but CTP, GTP, UTP and NAD(+) cannot. MTH: ligase activity is thermophilic in vitro, with optimal nick-joining at 60 degrees C. Mutational analysis of the conserved active site motif I (KxDG) illuminated essential roles for Lys251 and Asp253 at different steps of the ligation reaction. Mutant K251A is unable to form the covalent ligase-adenylate intermediate (step 1) and hence cannot seal a 3'-OH/5'-PO(4) nick. Yet, K251A catalyzes phosphodiester bond formation at a pre-adenylated nick (step 3). Mutant D253A is active in ligase-adenylate formation, but defective in activating the nick via formation of the DNA-adenylate intermediate (step 2). D253A is also impaired in phosphodiester bond formation at a pre-adenylated nick. A profound step 3 arrest, with accumulation of high levels of DNA-adenylate, could be elicited for the wild-type MTH: ligase by inclusion of calcium as the divalent cation cofactor. MTH: ligase sediments as a monomer in a glycerol gradient. Structure probing by limited proteolysis suggested that MTH: ligase is a tightly folded protein punctuated by a surface-accessible loop between nucleotidyl transferase motifs III and IIIa. PMID- 10871344 TI - Loss-of-function genetics in mammalian cells: the p53 tumor suppressor model. AB - Using an improved system for the functional identification of active antisense fragments, we have isolated antisense fragments which inactivate the p53 tumour suppressor gene. These antisense fragments map in two small regions between nt 350 and 700 and nt 800 and 950 of the coding sequence. These antisense fragments appear to act by inhibition of p53 mRNA translation both in vivo and in vitro. Expression of these antisense fragments overcame the p53-induced growth arrest in a cell line which expresses a thermolabile mutant of p53 and extended the in vitro lifespan of primary mouse embryonic fibroblasts. Continued expression of the p53 antisense fragment contributed to immortalisation of primary mouse fibroblasts. Subsequent elimination of the antisense fragment in these immortalised cells led to restoration of p53 expression and growth arrest, indicating that immortal cells continuously require inactivation of p53. Expression of MDM2 or SV40 large T antigen, but not E7 nor oncogenic ras, overcomes the arrest induced by restoration of p53 expression. Functional inactivation of both p21 and bax (by overexpression of Bcl2), but not either alone, allowed some bypass of p53-induced growth arrest, indicating that multiple transcriptional targets of p53 may mediate its antiproliferative action. The ability to conditionally inactivate and subsequently restore normal gene function may be extremely valuable for genetic analysis of genes for which loss-of function is involved in specific phenotypes. PMID- 10871343 TI - Two tricks in one bundle: helix-turn-helix gains enzymatic activity. AB - Many examples of enzymes that have lost their catalytic activity and perform other biological functions are known. The opposite situation is rare. A previously unnoticed structural similarity between the lambda integrase family (Int) proteins and the AraC family of transcriptional activators implies that the Int family evolved by duplication of an ancient DNA-binding homeodomain-like module, which acquired enzymatic activity. The two helix-turn-helix (HTH) motifs in Int proteins incorporate catalytic residues and participate in DNA binding. The active site of Int proteins, which include the type IB topoisomerases, is formed at the domain interface and the catalytic tyrosine residue is located in the second helix of the C-terminal HTH motif. Structural analysis of other 'tyrosine' DNA-breaking/rejoining enzymes with similar enzyme mechanisms, namely prokaryotic topoisomerase I, topoisomerase II and archaeal topoisomerase VI, reveals that the catalytic tyrosine is placed in a HTH domain as well. Surprisingly, the location of this tyrosine residue in the structure is not conserved, suggesting independent, parallel evolution leading to the same catalytic function by homologous HTH domains. The 'tyrosine' recombinases give a rare example of enzymes that evolved from ancient DNA-binding modules and present a unique case for homologous enzymatic domains with similar catalytic mechanisms but different locations of catalytic residues, which are placed at non-homologous sites. PMID- 10871345 TI - Oligodeoxynucleotide 5mers containing a 5'-CpG induce apoptosis through a mitochondrial mechanism in T lymphocytic leukaemia cells. AB - A chimeric methylphosphonodiester/phosphodiester 15mer oligodeoxynucleotide of randomly selected sequence was observed to rapidly induce apoptosis in MOLT-4 and Jurkat E6 T lymphocytic leukaemia cells following intracytoplasmic delivery. A series of further methylphosphonate substitutions and mutations and truncations of the oligodeoxynucleotide served to establish that the phosphodiester-linked sequence CGGTA present in the 15mer was responsible for this biological activity. End-protected CpG oligodeoxy-nucleotide 5mers of sequence type CGNNN exhibited a range of apoptosis-inducing potencies, with CGTTA being the most active. The latter was shown to significantly reduce the rate of RNA synthesis in MOLT-4 cells within 1 h; DNA laddering and redistribution of phosphatidylserine to the outer surface of the plasma membrane were marked by 160 min and mitochondrial transmembrane potential collapsed over roughly the same time scale. Pro-caspase 8 was reduced within 130 min and the proteolytically activated caspase 8 substrate Bid was also down by this time, implicating release of cytochrome c from mitochondria by the active 15 kDa fragment of Bid. Substantial proteolytic activation of pro-caspase 3 was relatively delayed. These findings support a mitochondrial amplification mechanism for apoptosis triggered by CpG 5mers. PMID- 10871346 TI - Topoisomerase activity of the hyperthermophilic replication initiator protein Rep75. AB - The plasmid pGT5 from the hyperthermophilic archaeon Pyrococcus abyssi replicates via the rolling circle mechanism. pGT5 encodes the replication initiator protein Rep75 that exhibits a nicking-closing (NC) activity in vitro on single-stranded oligonucleotides containing the pGT5 double-stranded origin (dso) sequence. Some mesophilic Rep proteins present site-specific DNA topo-isomerase-like activity on a negatively supercoiled plasmid harbouring the dso. We report here that Rep75 also exhibits topoisomerase activity on a negatively supercoiled DNA substrate. This DNA topoisomerase-like activity is dependent on the amino acids involved in NC activity of Rep75. However, in contrast with mesophilic Rep proteins, Rep75 topoisomerase activity is not dso dependent. Moreover, although pGT5 is known to be relaxed in vivo, Rep75 was not able to act on a relaxed plasmid in vitro, whether or not it contained the dso. PMID- 10871347 TI - CD43 gene expression is mediated by a nuclear factor which binds pyrimidine-rich single-stranded DNA. AB - CD43 is a leukocyte-specific surface molecule which plays an important role both in adhesion and signal transduction. We have identified a site spanning nucleotides +18 to +39 within the human CD43 gene promoter which in vitro is hypersensitive to cleavage by nuclease S1. Repeats of this region are sufficient to activate expression of a heterologous promoter in CD43-positive cell lines. Two nuclear factors, PyRo1 and PyRo2, interact with the hypersensitive site. PyRo1 is a single-stranded DNA-binding protein which binds the pyrimidine-rich sense strand. Mutation analysis demonstrates that the motif TCCCCT is critical for PyRo1 interaction. Replacement of this motif with the sequence CATATA abolishes PyRo1 binding and reduces expression of the CD43 promoter by 35% in Jurkat T lymphocytic cells and by 52% in the pre-erythroid/pre-megakaryocytic cell line K562. However, this same replacement failed to affect expression in U937 monocytic cells or in CEM T lymphocytic cells. PyRo1, therefore, exhibits cell-specific differences in its functional activity. Further analysis demonstrated that PyRo1 not only interacts with the CD43 gene promoter but also motifs present within the promoters of the CD11a, CD11b, CD11c and CD11d genes. These genes encode the alpha subunits of the beta2 integrin family of leukocyte adhesion receptors. Deletion of the PyRo1 binding site within the CD11c gene reduced promoter activity in T lymphocytic cells by 47%. However, consistent with our analysis of the CD43 gene, the effect of this same deletion within U937 monocytic cells was less severe. That PyRo1 binds preferentially to single stranded DNA and sequences within the CD43 and CD11 gene promoters suggests that expression of these genes is influenced by DNA secondary structure. PMID- 10871348 TI - Activation of the myc oncoprotein leads to increased turnover of thrombospondin-1 mRNA. AB - The Myc oncoprotein is implicated in transcriptional regulation of a variety of genes pertaining to cell cycle and neoplastic transformation. Examples of both positive and negative regulation have been reported that involve E-box and initiator (Inr) promoter elements, respectively. In both cases, Myc is thought to induce changes in transcription initiation. We have previously shown that overexpression of Myc causes down-regulation of the thrombospondin-1 (tsp-1) gene, an important negative modulator of tumor angiogenesis. In this study, we demonstrate that Myc in combination with Max can bind, albeit with low affinity, to an E-box-like element in the tsp-1 promoter. However, the 2.7 kb DNA segment containing both this non-canonical E-box and an Inr-like sequence does not constitute a Myc-responsive element in a transient expression system. Furthermore, Myc does not significantly affect the rate of initiation or elongation of the tsp-1 mRNA. Thus, in this instance Myc does not act as a canonical transcription factor. Instead, as demonstrated by blocking de novo RNA synthesis, down-regulation of the tsp-1 gene by Myc occurs through increased mRNA turnover. To our knowledge, this is the first example of gene regulation by Myc that involves mRNA destabilization. PMID- 10871349 TI - Escherichia coli exonuclease III enhances long PCR amplification of damaged DNA templates. AB - Recent development of the long PCR technology has provided an invaluable tool in many areas of molecular biology. However, long PCR amplification fails whenever the DNA template is imperfectly preserved. We report that Escherichia coli exonuclease III, a major repair enzyme in bacteria, strikingly improves the long PCR amplification of damaged DNA templates. Escherichia coli exonuclease III permitted or improved long PCR amplification with DNA samples submitted to different in vitro treatments known to induce DNA strand breaks and/or apurinic/apyrimidinic (AP) sites, including high temperature (99 degrees C), depurination at low pH and near-UV radiation. Exonuclease III also permitted or improved amplification with DNA samples that had been isolated several years ago by the phenol/chloroform method. Amelioration of long PCR amplification was achieved for PCR products ranging in size from 5 to 15.4 kb and with DNA target sequences located either within mitochondrial DNA or the nuclear genome. Exonuclease III increased the amplification of damaged templates using either rTth DNA polymerase alone or rTth plus Vent DNA polymerases or TAQ: plus PWO: DNA polymerases. However, exonuclease III could not improve PCR amplification from extensively damaged DNA samples. In conclusion, supplementation of long PCR mixes with E.COLI: exonuclease III may represent a major technical advance whenever DNA samples have been partly damaged during isolation or subsequent storage. PMID- 10871350 TI - Early melting of supercoiled DNA topoisomers observed by TGGE. AB - We have used temperature gradient gel electrophoresis (TGGE) to measure the progress of local denaturation in closed circular topoisomer DNA as a function of temperature and superhelicity (sigma). We describe the versatility of this method as a tool for detecting various conformational modifications of plasmid DNAs. The early melting temperature of a structural transition for any topoisomer is dependent on the value of superhelicity. Supercoiled topo-isomers represent a system of molecules that is sensitive to changes in temperature. We show that the topoisomer with the highest absolute value of superhelicity melts earlier than topoisomers with lower values. Thermal sensitivity of highly supercoiled plasmids could play a biologically important role in regulation of replication and expression in cells under thermal stress. The estimated melting temperature for plasmids with sigma < -0.05 is very significant because these temperatures for early melting are below physiological temperatures. PMID- 10871351 TI - Using molecular beacons as a sensitive fluorescence assay for enzymatic cleavage of single-stranded DNA. AB - Traditional methods to assay enzymatic cleavage of DNA are discontinuous and time consuming. In contrast, recently developed fluorescence methods are continuous and convenient. However, no fluorescence method has been developed for single stranded DNA digestion. Here we introduce a novel method, based on molecular beacons, to assay single-stranded DNA cleavage by single strand-specific nucleases. A molecular beacon, a hairpin-shaped DNA probe labeled with a fluorophore and a quencher, is used as the substrate and enzymatic cleavage leads to fluorescence enhancement in the molecular beacon. This method permits real time detection of DNA cleavage and makes it easy to characterize the activity of DNA nucleases and to study the steady-state cleavage reaction kinetics. The excellent sensitivity, reproducibility and convenience will enable molecular beacons to be widely useful for the study of single-stranded DNA cleaving reactions. PMID- 10871352 TI - Insertional mutagenesis based on illegitimate recombination in Schizosaccharomyces pombe. AB - An efficient insertional mutagenesis system has been developed for Schizosaccharomyces pombe based on linear PCR-generated cassettes containing selectable markers. It depends upon illegitimate recombination for integration into the genome. Various selectable markers of different sizes can be used to obtain sufficiently high transformation and integration frequencies. Based on Southern blotting, a single insertion is found in each strain and integration sites are broadly distributed in the genome. Sequence analysis of the insert junctions frequently reveals small regions of homology (4-10 bp) between the ends of the integrated cassette and the disrupted gene. The system has been used for simple genetic screens of various types and as a promoter trap for in-frame GFP fusions. PMID- 10871353 TI - Expression profiling across many samples via manifold-assisted mRNA processing. AB - Analysis of mRNA provides a condensed view of gene structure, and quantitative analyses can reveal induction of physiological or pathological gene expression programs. One of the main hurdles for routine mRNA analyses is the need to prepare large sets of samples in a rapid and standardized manner. We describe here a procedure for mRNA isolation and cDNA synthesis using manifold devices, consisting of a set of prongs that project into individual reaction wells. The prongs have a high binding capacity for the polyA-tails of mRNA and the captured mRNA is directly used to synthesize cDNA on the supports, followed by amplification. The convenience and reproducibility of the procedure allows profiling of gene expression over time, by comparing many different samples. Using the device mRNA was simultaneously isolated and accurately measured from up to 96 different samples of anywhere between 10 and 200 000 cells. The amounts of a leukemia-specific transcript could be measured when the malignant cells represented 5' exonuclease. AB - Werner's syndrome (WS) is an autosomal recessive disorder in humans characterized by the premature development of a partial array of age-associated pathologies. WRN, the gene defective in WS, encodes a 1432 amino acid protein (hWRN) with intrinsic 3'-->5' DNA helicase activity. We recently showed that hWRN is also a 3'-->5' exonuclease. Here, we further characterize the hWRN exonuclease. hWRN efficiently degraded the 3' recessed strands of double-stranded DNA or a DNA-RNA heteroduplex. It had little or no activity on blunt-ended DNA, DNA with a 3' protruding strand, or single-stranded DNA. The hWRN exonuclease efficiently removed a mismatched nucleotide at a 3' recessed terminus, and was capable of initiating DNA degradation from a 12-nt gap, or a nick. We further show that the mouse WRN (mWRN) is also a 3'-->5' exonuclease, with substrate specificity similar to that of hWRN. Finally, we show that hWRN forms a trimer and interacts with the proliferating cell nuclear antigen in vitro. These findings provide new data on the biochemical activities of WRN that may help elucidate its role(s) in DNA metabolism. PMID- 10871375 TI - Fine organization of Bombyx mori fibroin heavy chain gene. AB - The complete sequence of the Bombyx mori fibroin gene has been determined by means of combining a shotgun sequencing strategy with physical map-based sequencing procedures. It consists of two exons (67 and 15 750 bp, respectively) and one intron (971 bp). The fibroin coding sequence presents a spectacular organization, with a highly repetitive and G-rich (approximately 45%) core flanked by non-repetitive 5' and 3' ends. This repetitive core is composed of alternate arrays of 12 repetitive and 11 amorphous domains. The sequences of the amorphous domains are evolutionarily conserved and the repetitive domains differ from each other in length by a variety of tandem repeats of subdomains of approximately 208 bp which are reminiscent of the repetitive nucleosome organization. A typical composition of a subdomain is a cluster of repetitive units, Ua, followed by a cluster of units, Ub, (with a Ua:Ub ratio of 2:1) flanked by conserved boundary elements at the 3' end. Moreover some repeats are also perfectly conserved at the peptide level indicating that the evolutionary pressure is not identical along the sequence. A tentative model for the constitution and evolution of this unusual gene is discussed. PMID- 10871377 TI - The global intrinsic curvature of archaeal and eubacterial genomes is mostly contained in their dinucleotide composition and is probably not an adaptation. AB - Until now, the genomic DNA of all eubacteria analyzed has been hyper-curved, its global intrinsic curvature being higher than that of a random sequence. In contrast, that rule failed for archaea or eukaryotes, which could be either hypo- or hyper-curved. The existence of the rule suggested that, at least for eubacteria, global intrinsic curvature is adaptive. However, the present results from analyzing 21 eubacterial and six archaeal genomes argue against adaptation. First, there are two eubacterial exceptions to the former rule. More significantly, we found that the dinucleotide composition of the genome alone (which lacks all sequence information) is enough to determine the genome curvature. Additional evidence against adaptation came from showing that the global curvature of bacterial genomes could not have evolved under either of two complementary models of curvature selection: (i) that curvature is selected locally from unbiased variability; (ii) that curvature is established globally through the selection of a curvature-altering mutational bias. We found that the observed relationship between curvature and dinucleotide composition is incompatible with model (i). We also found that, contrary to the predictions of model (ii), the dinucleo-tide compositions of bacterial genomes were not statistically special in their curvature-related properties (when compared to stochastically generated dinucleotide compositions). PMID- 10871376 TI - Potent inhibition of werner and bloom helicases by DNA minor groove binding drugs. AB - Maintenance of genomic integrity is vital to all organisms. A number of human genetic disorders, including Werner Syndrome, Bloom Syndrome and Rothmund-Thomson Syndrome, exhibit genomic instability with some phenotypic characteristics of premature aging and cancer predisposition. Presumably the aberrant cellular and clinical phenotypes in these disorders arise from defects in important DNA metabolic pathways such as replication, recombination or repair. These syndromes are all characterized by defects in a member of the RecQ family of DNA helicases. To obtain a better understanding of how these enzymes function in DNA metabolic pathways that directly influence chromosomal integrity, we have examined the effects of non-covalent DNA modifications on the catalytic activities of purified Werner (WRN) and Bloom (BLM) DNA helicases. A panel of DNA-binding ligands displaying unique properties for interacting with double helical DNA was tested for their effects on the unwinding activity of WRN and BLM helicases on a partial duplex DNA substrate. The levels of inhibition by a number of these compounds were distinct from previously reported values for viral, prokaryotic and eukaryotic helicases. The results demonstrate that BLM and WRN proteins exhibit similar sensitivity profiles to these DNA-binding ligands and are most potently inhibited by the structurally related minor groove binders distamycin A and netropsin (K(i) 20-30%, good regression fits were obtained when values derived from densitometric scanning of an agarose gel and those derived from the SCFluo method were compared. The method represents an attractive alternative to currently established methods because it is simple, rapid and quantitative. During large-scale processing and long-term storage, enzymatic, chemical and shear degradation may substantially decrease the SC content of plasmid DNA preparations. Regulations for pharmaceutical grade products for use in gene therapy and DNA vaccination may require >90% of the plasmid to be in the SC form. In the present study the SC content of 6.9, 13 and 20 kb plasmid preparations that had been subjected to chemical and shear degradation was successfully quantified using the new method. PMID- 10871379 TI - Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. AB - Host cell factor (HCF, C1, VCAF or CFF) is a cellular protein that is required for transcription activation of herpes simplex virus (HSV) immediate-early (IE) genes by the virion protein VP16. The biological function of HCF remains unclear. Recently we identified a cellular transcription activator, Luman. As with VP16, the transactivation function of Luman is also regulated by HCF. Here we report a second human protein, Zhangfei (ZF) that interacts with HCF in a fashion similar to Luman and VP16. Although ZF shares no significant sequence homology with Luman, the two proteins have some structural similarities. These include: a basic domain-leucine zipper (bZIP) region, an acidic activation domain and a consensus HCF-binding motif. Unlike Luman, or most other bZIP proteins, ZF by itself did not appear to bind consensus bZIP-binding sites. It was also unable to activate promoters containing these response elements. Although in transient expression assays ectopically expressed ZF was unable to block transactivation by VP16 of a HSV IE promoter, ZF could prevent the expression of several HSV proteins in cells infected with the virus. The ability of ZF to block the synthesis of the HSV IE protein ICP0 relied on its binding to HCF, since a mutant of ZF that was unable to bind HCF was also unable to prevent viral IE protein expression. PMID- 10871381 TI - PCR-generated padlock probes detect single nucleotide variation in genomic DNA. AB - Circularizing oligonucleotide probes, so-called padlock probes, have properties that should prove valuable in a wide range of genetic investigations, including in situ analyses, genotyping and measurement of gene expression. However, padlock probes can be difficult to obtain by standard oligonucleotide synthesis because they are relatively long and require intact 5'- and 3'-end sequences to function. We describe a PCR-based protocol for flexible small-scale enzymatic synthesis of such probes. The protocol also offers the advantage over chemical synthesis that longer probes can be made that are densely labeled with detectable functions, resulting in an increased detection signal. The utility of probes synthesized according to this protocol is demonstrated for the analysis of single nucleotide variations in human genomic DNA both in situ and in solution. PMID- 10871382 TI - Real-time monitoring of in vitro transcriptional RNA synthesis using fluorescence resonance energy transfer. AB - We have developed a novel method for real-time monitoring of RNA synthesis in in vitro transcription reactions using fluorescence resonance energy transfer (FRET). Two 15mer DNAs, either of which was labeled with Bodipy493/503 as a donor or Cy5 as an acceptor, were prepared. When the two fluorescent DNAs hybridized to adjacent locations on Xenopus: elongation factor 1-alpha (xelf1-alpha) RNA, the distance between the two fluorophores became very close, causing FRET to occur and resulting in changes in fluorescence spectra. A high accessibility 30mer site of xelf1-alpha RNA was found and excess amounts of a pair of donor and acceptor DNA probes that were complementary to the site were added to the in vitro transcription reaction solution. Changes in fluorescence spectra were observed in response to progression of xelf1-alpha RNA synthesis that showed that the fluorescent probes hybridized to the synthesized RNA. Furthermore, when probes hybridizing to the synthesized xelf1-alpha RNA with less efficiency were used to monitor the reaction, spectral changes in response to RNA synthesis were also observed. This result suggests that the probes hybridized to synthesizing RNA molecules before they folded to form secondary structure and that there is no need to select sites on the RNA for the probes, which is required for probes hybridizing to folded RNA molecules. PMID- 10871383 TI - A miniature integrated device for automated multistep genetic assays. AB - A highly integrated monolithic device was developed that automatically carries out a complex series of molecular processes on multiple samples. The device is capable of extracting and concentrating nucleic acids from milliliter aqueous samples and performing microliter chemical amplification, serial enzymatic reactions, metering, mixing and nucleic acid hybridization. The device, which is smaller than a credit card, can manipulate over 10 reagents in more than 60 sequential operations and was tested for the detection of mutations in a 1.6 kb region of the HIV genome from serum samples containing as few as 500 copies of the RNA. The elements in this device are readily linked into complex, flexible and highly parallel analysis networks for high throughput sample preparation or, conversely, for low cost portable DNA analysis instruments in point-of-care medical diagnostics, environmental testing and defensive biological agent detection. PMID- 10871384 TI - Rapid purification of protein complexes from mammalian cells. AB - The evaluation of the protein binding partner(s) of biologically important proteins is currently an area of intense research, especially since the development of the yeast two-hybrid assay. However, not all protein-protein interactions uncovered by this assay are biologically relevant and another confirmatory assay must be performed. Ideally, this assay should be rapid, versatile and performed under conditions which mimic the 'normal' physiological state as closely as possible. Towards this goal, we have constructed two eukaryotic expression vectors that facilitate the purification of a protein of interest, along with any associated proteins, from mammalian cells. These vectors incorporate the following features: (i) a tetracycline-responsive promoter so that the level of protein production can be regulated; (ii) an N-terminal glutathione S-transferase tag or a triple repeat of the HA1 epitope, to facilitate purification of the protein either by glutathione affinity chromatography or immunoprecipitation, respectively, followed by a multiple cloning site; (iii) the gene for the enhanced green fluorescent protein (for detection of the presence of the fusion protein and subcellular localization); (iv) a puromycin marker for the selection of stable transformants; (v) a truncated EBNA protein and oriP sequence for episomal replication of the vector. These latter two features permit expansion of small cultures of transfected cells under puromycin selection, thereby increasing the amount of tagged protein that can be purified. We show that these vectors can be used to direct the doxycycline inducible expression of tagged proteins and to recover tagged CIP1-p21 protein complexes from HeLa cells. Furthermore, we show that these tagged p21-purified complexes contain both cyclin A and Cdk2, which are known to interact with p21, but not beta-actin. PMID- 10871385 TI - Improved NlaIII digestion of PAGE-purified 102 bp ditags by addition of a single purification step in both the SAGE and microSAGE protocols. AB - Despite the success of microarray technologies, serial analysis of gene expression (SAGE) still remains the only technique that allows an accurate quantitative and qualitative analysis of cell transcription in a variety of physiological and pathological conditions. Nevertheless, the efficiency of SAGE is limited by the numerous gel purification steps required and these increase the possibility of contamination and reduce or inhibit the activity of the enzymes used in the protocol. In order to eliminate this problem, we have modified the original protocol by adding a single purification step before NLA:III digestion of the ditags. This allows us to increase the yield of digested ditags without reducing the amount of DNA or affecting the subsequent concatemerization. PMID- 10871386 TI - Loop-mediated isothermal amplification of DNA. AB - We have developed a novel method, termed loop-mediated isothermal amplification (LAMP), that amplifies DNA with high specificity, efficiency and rapidity under isothermal conditions. This method employs a DNA polymerase and a set of four specially designed primers that recognize a total of six distinct sequences on the target DNA. An inner primer containing sequences of the sense and antisense strands of the target DNA initiates LAMP. The following strand displacement DNA synthesis primed by an outer primer releases a single-stranded DNA. This serves as template for DNA synthesis primed by the second inner and outer primers that hybridize to the other end of the target, which produces a stem-loop DNA structure. In subsequent LAMP cycling one inner primer hybridizes to the loop on the product and initiates displacement DNA synthesis, yielding the original stem loop DNA and a new stem-loop DNA with a stem twice as long. The cycling reaction continues with accumulation of 10(9) copies of target in less than an hour. The final products are stem-loop DNAs with several inverted repeats of the target and cauliflower-like structures with multiple loops formed by annealing between alternately inverted repeats of the target in the same strand. Because LAMP recognizes the target by six distinct sequences initially and by four distinct sequences afterwards, it is expected to amplify the target sequence with high selectivity. PMID- 10871387 TI - New chromatographic and biochemical strategies for quick preparative isolation of tRNA. AB - A combination of hydrophobic chromatography on phenyl-Sepharose and reversed phase HPLC was used to purify individual tRNAs with high specific activity. The efficiency of chromatographic separation was enhanced by biochemical manipulations of the tRNA molecule, such as aminoacylation, formylation of the aminoacyl moiety and enzymatic deacylation. Optimal combinations are presented for three different cases. (i) tRNA(Phe) from Escherichia coli. This species was isolated by a combination of low pressure phenyl-Sepharose hydrophobic chromatography with RP-HPLC. (ii) tRNA(Ile) from E. coli: Aminoacylation increases the retention time for this tRNA in RP-HPLC. The recovered acylated intermediate is deacylated by reversion of the aminoacylation reaction and submitted to a second RP-HPLC run, in which deacylated tRNA(Ile) is recovered with high specific activity. (iii) tRNA(i)(Met) from Saccharomyces cerevisiae. The aminoacylated form of this tRNA is unstable. To increase stability, the aminoacylated form was formylated using E.coli: enzymes and, after one RP-HPLC step, the formylated derivative was deacylated using peptidyl-tRNA hydrolase from E.COLI: The tRNA(i)(Met) recovered after a second RP-HPLC run exhibited electrophoretic homogeneity and high specific activity upon aminoacylation. These combinations of chromatographic separation and biochemical modification can be readily adapted to the large-scale isolation of any particular tRNA. PMID- 10871388 TI - Modification of human beta-globin locus PAC clones by homologous recombination in Escherichia coli. AB - We report here modifications of human beta-globin PAC clones by homologous recombination in Escherichia coli DH10B, utilising a plasmid temperature sensitive for replication, the recA gene and a wild-type copy of the rpsL gene which allows for an efficient selection for plasmid loss in this host. High frequencies of recombination are observed even with very small lengths of homology and the method has general utility for introducing insertions, deletions and point mutations. No rearrangements were detected with the exception of one highly repetitive genomic sequence when either the E.COLI: RecA- or the lambdoid phage encoded RecT and RecE-dependent recombination systems were used. PMID- 10871389 TI - Preparation of DNA and protein micro arrays on glass slides coated with an agarose film. AB - A thin layered agarose film on microscope slides provides a versatile support for the preparation of arrayed molecular libraries. An activation step leading to the formation of aldehyde groups in the agarose creates reactive sites that allow covalent immobilization of molecules containing amino groups. Arrays of oligonucleotides and PCR products were prepared by tip printing. After hybridization with complementary fluorescence labeled nucleic acid probes strong fluorescence signals of sequence-specific binding to the immobilized probes were detected. The intensity of the fluorescence signals was proportional to the relative amount of immobilized oligonucleotides and to the concentration of the fluorescence labeled probe. We also used the agarose film-coated slides for the preparation of protein arrays. In combination with specific fluorescence labeled antibodies these protein arrays can be used for fluorescence linked immune assays. With this approach different protein tests can be performed in parallel in a single reaction with minimal amounts of the binding reagents. PMID- 10871390 TI - A simple and efficient method for PCR amplifiable DNA extraction from ancient bones. AB - A simple and effective modified ethanol precipitation-based protocol is described for the preparation of DNA from ancient human bones. This method is fast and requires neither hazardous chemicals nor special devices. After the powdering and incubating of the bone samples Dextran Blue was added as a carrier for removing the PCR inhibitors with selective ethanol precipitation. This method could eliminate the time-consuming separate decalcification step, dialysis, application of centrifugation-driven microconcentrators and the second consecutive PCR amplification. The efficiency of this procedure was demonstrated on ten 500-1200 year-old human bones from four different Hungarian burial sites. A mitochondrial specific primer pair was used to obtain sequence information from the purified ancient DNA. The PCR amplification, after our DNA extraction protocol, was successful from each of the 10 bone samples investigated. The results demonstrate that extraction of DNA from ancient bone samples with this new approach increases the success rate of PCR amplification. PMID- 10871391 TI - A new MALDI-TOF based mini-sequencing assay for genotyping of SNPS. AB - A new MALDI-TOF based mini-sequencing assay termed VSET was developed for genotyping of SNPs. In this assay, specific fragments of genomic DNA containing the SNP site(s) are first amplified, followed by mini-sequencing in the presence of three ddNTPs and the fourth nucleotide in the deoxy form. In this way, the primer is extended by only one base from one allele, while it is typically extended by two bases from another allele. The products are then analyzed using MALDI-TOF mass spectrometry. The genotype of the SNP site is identified based on the number of nucleotides added. This assay has been examined using both synthetic and genomic DNA samples. In addition, multiplexed assays were successfully performed to genotype four SNP sites in a single tube. The main aspect of this assay is that it can overcome the key problems associated with the currently used mini-sequencing methods. First, it significantly reduces the stringent high-resolution and extensive desalting requirements that are essential to the pinpoint assay. Second, it avoids the long extension problem associated with the PROBE assay. PMID- 10871392 TI - Position and degree of mismatches and the mobility of DNA heteroduplexes. AB - Heteroduplex mobility assay (HMA) is a fast and inexpensive method for determining relatedness between DNA sequences. Rapidly evolving viruses such as HIV-1 develop marked sequence differences in their genomes over the course of the epidemic and infection in a single individual. HMA can be used to monitor both processes. Here, we systematically evaluated the influence of single base mismatches on heteroduplex mobility. The impact of mismatches at nine different positions in 559 bp double-stranded DNA molecules, within a background of overall sequence divergence ranging from 1.97 to 9.65%, was evaluated in both non denaturing and partially-denaturing acrylamide gels. We found that the electrophoretic mobility of heteroduplexes was proportional to the level of mismatch when that level exceeded 4.5%. Overall, mismatches near the center of the fragment and clustered mismatches tended to have an exaggerated influence on the mobility of heteroduplexes. Thus, the use of HMA for quantitative inference of genetic distances under the conditions we describe is of greatest utility at levels of mismatch >5%. PMID- 10871393 TI - RNA accessibility prediction: a theoretical approach is consistent with experimental studies in cell extracts. AB - The use of antisense oligodeoxyribonucleotides (ODN) or ribozymes to specifically suppress gene expression is simple in concept and relies on efficient binding of the antisense strand to the target RNA. Although the identification of target sites accessible to base pairing is gradually being overcome by different techniques, it remains a major problem in the antisense and ribozyme approaches. In this study we have investigated the potential of a recent experimental and theoretical approach to predict the local accessibility of murine DNA methyltransferase (MTase) mRNA in a comparative way. The accessibility of the native target RNA was probed with antisense ODN in cellular extracts. The results strongly correlated with the theoretically predicted target accessibility. This work suggests an effective two-step procedure for predicting RNA accessibility: first, computer-aided selection of ODN binding sites defined by an accessibility score followed by a more detailed experimental procedure to derive information about target accessibility at the single nucleotide level. PMID- 10871394 TI - In vitro selection supports the view of a kinetic control of antisense RNA mediated inhibition of gene expression in mammalian cells. AB - In principle, the steady-state concentrations of biomolecules in complex systems can be far from the thermodynamic equilibrium concentrations of individual processes. This means that, in addition to thermodynamics, reaction kinetics may play an important role. This view is not fully reflected in combinatorial studies in biochemistry that focus on the selection of stably interacting molecules reflected by high equilibrium constants. For kinetically controlled processes in vivo, forward or backward reaction rates are critical but not necessarily an equilibrium state. Here we have studied the control of antisense RNA-mediated gene suppression in human cells on a general basis and in a way that excludes individual structure-specific influences. The complete antisense sequence space against the chloramphenicol acetyltransferase gene (cat) was generated and a kinetic selection technique was established to enrich for fast annealing antisense species. Selected sub-populations showed successively faster annealing which was related to increased inhibition of cat gene expression in HeLa cells, providing strong evidence for the view that the suppression of gene expression by antisense RNA is controlled kinetically regardless of specific RNA structures. PMID- 10871395 TI - Retrotransposition of the I factor, a non-long terminal repeat retrotransposon of Drosophila, generates tandem repeats at the 3' end. AB - Non-long terminal repeat (LTR) retrotransposons or LINEs transpose by reverse transcription of an RNA intermediate and are thought to use the 3' hydroxyl of a chromosomal cleavage to initiate synthesis of the first strand of the cDNA. Many of them terminate in a poly(dA) sequence at the 3' end of the coding strand although some, like the I factor of Drosophila melanogaster, have 3' ends formed by repeats of the trinucleotide TAA. We report results showing that I factor transcripts end a few nucleotides downstream of the TAA repeats and that these extra nucleotides are not integrated into chromosomal DNA during retrotransposition. We also show that the TAA repeats are not required for transposition and that I elements containing mutations affecting the TAA sequences generate transposed copies ending with tandem repeats of various types. Our results suggest that during integration the 3' end of the I factor RNA template can pair with nucleotides at the target site and that tandem duplications are generated by the reverse transcriptase of the I factor in a manner that is reminiscent of the activity of the reverse transcriptases of telomerases. Reverse transcriptases of other non-LTR retrotransposons may function in a similar way. PMID- 10871396 TI - Complementation of defective translesion synthesis and UV light sensitivity in xeroderma pigmentosum variant cells by human and mouse DNA polymerase eta. AB - Defects in the human gene XPV result in the variant form of the genetic disease xeroderma pigmentosum (XP-V). XPV encodes DNA polymerase eta, a novel DNA polymerase that belongs to the UmuC/DinB/Rad30 superfamily. This polymerase catalyzes the efficient and accurate translesion synthesis of DNA past cis-syn cyclobutane di-thymine lesions. In this report we present the cDNA sequence and expression profiles of the mouse XPV gene and demonstrate its ability to complement defective DNA synthesis in XP-V cells. The mouse XPV protein shares 80.3% amino acid identity and 86.9% similarity with the human XPV protein. The recombinant mouse XPV protein corrected the inability of XP-V cell extracts to carry out DNA replication, by bypassing thymine dimers on template DNA. Transfection of the mouse or human XPV cDNA into human XP-V cells corrected UV sensitivity. Northern blot analysis revealed that the mouse XPV gene is expressed ubiquitously, but at a higher level in testis, liver, skin and thymus compared to other tissues. Although the mouse XPV gene was not induced by UV irradiation, its expression was elevated approximately 4-fold during cell proliferation. These results suggest that DNA polymerase eta plays a role in DNA replication, though the enzyme is not essential for viability. PMID- 10871398 TI - Quantitative characterization of the interaction between purified human estrogen receptor alpha and DNA using fluorescence anisotropy. AB - In an effort to better define the molecular mechanisms of the functional specificity of human estrogen receptor alpha, we have carried out equilibrium binding assays to study the interaction of the receptor with a palindromic estrogen response element derived from the vitellogenin ERE. These assays are based on the observation of the fluorescence anisotropy of a fluorescein moiety covalently bound to the target oligonucleotide. The low anisotropy value due to the fast tumbling of the free oligonucleotide in solution increases substantially upon binding the receptor to the labeled ERE. The quality of our data are sufficient to ascertain that the binding is clearly cooperative in nature, ruling out a simple monomer interaction and implicating a dimerization energetically coupled to DNA binding in the nanomolar range. The salt concentration dependence of the affinity reveals formation of high stoichiometry, low specificity complexes at low salt concentration. Increasing the KCl concentration above 200 mM leads to specific binding of ER dimer. We interpret the lack of temperature dependence of the apparent affinity as indicative of an entropy driven interaction. Finally, binding assays using fluorescent target EREs bearing mutations of each of the base pairs in the palindromic ERE half-site indicate that the energy of interaction between ER and its target is relatively evenly distributed throughout the site. PMID- 10871397 TI - Structure-based predictions of Rad1, Rad9, Hus1 and Rad17 participation in sliding clamp and clamp-loading complexes. AB - The repair of damaged DNA is coupled to the completion of DNA replication by several cell cycle checkpoint proteins, including, for example, in fission yeast Rad1(Sp), Hus1(Sp), Rad9(Sp) and Rad17(Sp). We have found that these four proteins are conserved with protein sequences throughout eukaryotic evolution. Using computational techniques, including fold recognition, comparative modeling and generalized sequence profiles, we have made high confidence structure predictions for the each of the Rad1, Hus1 and Rad9 protein families (Rad17(Sc), Mec3(Sc) and Ddc1(Sc) in budding yeast, respectively). Each of these families was found to share a common protein fold with that of PCNA, the sliding clamp protein that tethers DNA polymerase to its template. We used previously reported genetic and biochemical data for these proteins from yeast and human cells to predict a heterotrimeric PCNA-like ring structure for the functional Rad1/Rad9/Hus1 complex and to determine their exact order within it. In addition, for each individual protein family, contact regions with neighbors within the PCNA-like ring were identified. Based on a molecular model for Rad17(Sp), we concluded that members of this family, similar to the subunits of the RFC clamp-loading complex, are capable of coupling ATP binding with conformational changes required to load a sliding clamp onto DNA. This model substantiates previous findings regarding the behavior of Rad17 family proteins upon DNA damage and within the RFC complex of clamp-loading proteins. PMID- 10871400 TI - The mitochondrial genome of the stramenopile alga Chrysodidymus synuroideus. Complete sequence, gene content and genome organization. AB - This is the first report of a complete mitochondrial genome sequence from a photosynthetic member of the stramenopiles, the chrysophyte alga Chrysodidymus synuroideus. The circular-mapping mitochondrial DNA (mtDNA) of 34 119 bp contains 58 densely packed genes (all without introns) and five unique open reading frames (ORFs). Protein genes code for components of respiratory chain complexes, ATP synthase and the mitoribosome, as well as one product of unknown function, encoded in many other protist mtDNAs (YMF16). In addition to small and large subunit ribosomal RNAs, 23 tRNAs are mtDNA-encoded, permitting translation of all codons present in protein-coding genes except ACN (Thr) and CGN (Arg). The missing tRNAs are assumed to be imported from the cytosol. Comparison of the C.SYNUROIDEUS: mtDNA with that of other stramenopiles allowed us to draw conclusions about mitochondrial genome organization, expression and evolution. First, we provide evidence that mitochondrial ORFs code for highly derived, unrecognizable versions of ribosomal or respiratory genes otherwise 'missing' in a particular mtDNA. Secondly, the observed constraints in mitochondrial genome rearrangements suggest operon-based, co-ordinated expression of genes functioning in common biological processes. Finally, stramenopile mtDNAs reveal an unexpectedly low variability in genome size and gene complement, testifying to substantial differences in the tempo of mtDNA evolution between major eukaryotic lineages. PMID- 10871399 TI - Expression and function of the homeodomain-containing protein Hex in thyroid cells. AB - The homeodomain-containing protein Hex (also named Prh) is expressed in primitive endoderm (during the early phases of development), in some endoderm-derived tissues and in endothelial and hematopoietic precursors. Hex expression is exting uished during terminal differentiation of endothelial and hematopoietic cells as well as in adult lung. Previous investigations have demonstrated that Hex is expressed during early thyroid gland development. No information has been reported on Hex expression in adult thyroid gland or on the function of this protein in follicular thyroid cells. These issues represent the focus of the present study. We demonstrate that Hex mRNA is present in rat and human adult thyroid gland as well as in differentiated follicular thyroid cell lines. In FRTL 5 cells TSH reduces Hex expression. In thyroid cell lines transformed by several oncogenes Hex expression is completely abolished. By using co-transfection assays we demonstrate that Hex is a repressor of the thyroglobulin promoter and that it is able to abolish the activating effects of both TTF-1 and Pax8. These data would suggest that Hex may play an important role in thyroid cell differentiation. Protein-DNA interaction experiments indicate that Hex is able to bind sites of the thyroglobulin promoter containing either the core sequence 5' TAAT-3' or 5'-CAAG-3'. The DNA binding specificity of the Hex homeodomain, therefore, is more 'relaxed' than that observed in the majority of other homeo domains. PMID- 10871401 TI - Identification of high affinity Tbf1p-binding sites within the budding yeast genome. AB - The yeast TBF1 gene is essential for mitotic growth and encodes a protein that binds the human telomere repeats in vitro, although its cellular function is unknown. The sequence of the DNA-binding domain of Tbf1p is more closely related to that of the human telomeric proteins TRF1 and TRF2 than to any yeast protein sequence, yet the functional homologue of TRF1 and TRF2 is thought to be Rap1p. In this study we show that the Tbf1p DNA-binding domain can target the Gal4 transactivation domain to a (TTAGGG)(n) sequence inserted in the yeast genome, supporting the model that Tbf1p binds this sub-telomeric repeat motif in vivo. Immunofluorescence of Tbf1p shows a spotty pattern throughout the interphase nucleus and along synapsed chromosomes in meiosis, suggesting that Tbf1p binds internal chromosomal sites in addition to sub-telomeric regions. PCR-assisted binding site selection was used to define a consensus for high affinity Tbf1p binding sites. Compilation of 50 selected oligonucleotides identified the consensus TAGGGTTGG. Five potential Tbf1p-binding sites resulting from a search of the total yeast genome were tested directly in gel shift assays and shown to bind Tbf1p efficiently in vitro, thus confirming this as a valid consensus for Tbf1p recognition. PMID- 10871402 TI - Importance of discriminator base stacking interactions: molecular dynamics analysis of A73 microhelix(Ala) variants. AB - Transfer of alanine from Escherichia coli alanyl-tRNA synthetase (AlaRS) to RNA minihelices that mimic the amino acid acceptor stem of tRNA(Ala) has been shown, by analysis of variant minihelix aminoacylation activities, to involve a transition state sensitive to changes in the 'discriminator' base at position 73. Solution NMR has indicated that this single-stranded nucleotide is predominantly stacked onto G1 of the first base pair of the alanine acceptor stem helix. We report the activity of a new variant with the adenine at position 73 substituted by its non-polar isostere 4-methylindole (M). Despite lacking N7, this analog is well tolerated by AlaRS. Molecular dynamics (MD) simulations show that the M substitution improves position 73 base stacking over G1, as measured by a stacking lifetime analysis. Additional MD simulations of wild-type microhelix(Ala) and six variants reveal a positive correlation between N73 base stacking propensity over G1 and aminoacylation activity. For the two DeltaN7 variants simulated we found that the propensity to stack over G1 was similar to the analogous variants that contain N7 and we conclude that the decrease in aminoacylation efficiency observed upon deletion of N7 is likely due to loss of a direct stabilizing interaction with the synthetase. PMID- 10871403 TI - Recognition of GT mismatches by Vsr mismatch endonuclease. AB - The Vsr mismatch endonuclease recognises the sequence CTWGG (W = A or T) in which the underlined thymine is paired with guanine and nicks the DNA backbone on the 5'-side of the mispaired thymine. By using base analogues of G and T we have explored the functional groups on the mismatch pair which are recognised by the enzyme. Removal of the thymine 5-methyl group causes a 60% reduction in activity, while removing the 2-amino group of guanine reduces cleavage by 90%. Placing 2 amino-purine or nebularine opposite T generates mis-matches which are cut at a much lower rate (0.1%). When either base is removed, generating a pseudoabasic site (1', 2'-dideoxyribose), the enzyme still produces site-specific cleavage, but at only 1% of the original rate. Although TT and CT mismatches at this position are cleaved at a low rate (approximately 1%), mismatches with other bases (such as GA and AC) and Watson-Crick base pairs are not cleaved by the enzyme. There is also no cleavage when the mismatched T is replaced with difluorotoluene. PMID- 10871404 TI - HMG I/Y regulates long-range enhancer-dependent transcription on DNA and chromatin by changes in DNA topology. AB - The nature of nuclear structures that are required to confer transcriptional regulation by distal enhancers is unknown. We show that long-range enhancer dependent beta-globin transcription is achieved in vitro upon addition of the DNA architectural protein HMG I/Y to affinity-enriched holo RNA polymerase II complexes. In this system, HMG I/Y represses promoter activity in the absence of an associated enhancer and strongly activates transcription in the presence of a distal enhancer. Importantly, nucleosome formation is neither necessary for long range enhancer regulation in vitro nor sufficient without the addition of HMG I/Y. Thus, the modulation of DNA structure by HMG I/Y is a critical regulator of long-range enhancer function on both DNA and chromatin-assembled genes. Electron microscopic analysis reveals that HMG I/Y binds cooperatively to preferred DNA sites to generate distinct looped structures in the presence or absence of the beta-globin enhancer. The formation of DNA topologies that enable distal enhancers to strongly regulate gene expression is an intrinsic property of HMG I/Y and naked DNA. PMID- 10871405 TI - Identification of proteins of Escherichia coli and Saccharomyces cerevisiae that specifically bind to C/C mismatches in DNA. AB - The pathways leading to G:C-->C:G transversions and their repair mechanisms remain uncertain. C/C and G/G mismatches arising during DNA replication are a potential source of G:C-->C:G transversions. The Escherichia coli mutHLS mismatch repair pathway efficiently corrects G/G mismatches, whereas C/C mismatches are a poor substrate. Escherichia coli must have a more specific repair pathway to correct C/C mismatches. In this study, we performed gel-shift assays to identify C/C mismatch-binding proteins in cell extracts of E. COLI: By testing heteroduplex DNA (34mers) containing C/C mismatches, two specific band shifts were generated in the gels. The band shifts were due to mismatch-specific binding of proteins present in the extracts. Cell extracts of a mutant strain defective in MutM protein did not produce a low-mobility complex. Purified MutM protein bound efficiently to the C/C mismatch-containing heteroduplex to produce the low mobility complex. The second protein, which produced a high-mobility complex with the C/C mismatches, was purified to homogeneity, and the amino acid sequence revealed that this protein was the FabA protein of E.COLI: The high-mobility complex was not formed in cell extracts of a fabA mutant. From these results it is possible that MutM and FabA proteins are components of repair pathways for C/C mismatches in E.COLI: Furthermore, we found that Saccharomyces cerevisiae OGG1 protein, a functional homolog of E.COLI: MutM protein, could specifically bind to the C/C mismatches in DNA. PMID- 10871406 TI - Identification and characterization of negative regulatory elements of the human telomerase catalytic subunit (hTERT) gene promoter: possible role of MZF-2 in transcriptional repression of hTERT. AB - Human telomerase reverse transcriptase (hTERT) is a catalytic subunit of human telomerase and is a critical determinant of the enzymatic activity of telomerase. Expression of hTERT is known to be regulated mainly at the transcriptional level. In the present study, using transient expression assays, we identified a 400 bp silencer of the hTERT promoter between -776 and -378 upstream of the proximal core promoter. The inhibitory effects of this silencer were enhanced with cellular differentiation. A computer-assisted homology search identified multiple binding motifs for myeloid-specific zinc finger protein 2 (MZF-2) within this region. Mutation introduced in these sites resulted in significant activation of hTERT transcription. Gel shift assays demonstrated that MZF-2 proteins specifically bound to these sites. Overexpression of MZF-2 in cells led to down regulation of hTERT transcription as well as telomerase activity. These findings suggest that the 400 bp region upstream of the hTERT core promoter that we identified functions as a negative regulatory region and that MZF-2 may be an effector of negative regulation of hTERT. PMID- 10871407 TI - The role of region II in the RNA polymerase sigma factor sigma(N) (sigma(54)). AB - Bacterial RNA polymerase holoenzymes containing the sigma subunit sigma(N) (sigma(54)) can form a stable closed complex with promoter DNA but only undergo transition to an open complex and transcription initiation when acted on by an activator protein. Proteins of the sigma(N) family have a conserved N-terminal region of 50 amino acids (Region I) that is separated from a conserved C-terminal region of around 360 amino acids (Region III) by a much more variable sequence of between 30 and 110 residues (Region II). We have investigated the role of Region II in Klebsiella pneumoniae sigma(N) by studying the properties of deletions of all or part of the region both in vivo and in vitro. We found that whilst Region II is not essential, deletion of all or part of it can significantly impair sigma(N) activity. Deletions have effects on DNA binding by the isolated sigma factor and on holoenzyme formation, but the most marked effects are on transition of the holoenzyme from the closed to the open complex in the presence of the activator protein. PMID- 10871408 TI - The complete nucleotide sequence of the mitochondrial genome of sugar beet (Beta vulgaris L.) reveals a novel gene for tRNA(Cys)(GCA). AB - We determined the complete nucleotide sequence of the mitochondrial genome of an angiosperm, sugar beet (Beta vulgaris cv TK81-O). The 368 799 bp genome contains 29 protein, five rRNA and 25 tRNA genes, most of which are also shared by the mitochondrial genome of Arabidopsis thaliana, the only other completely sequenced angiosperm mitochondrial genome. However, four genes identified here (namely rps13, trnF-GAA, ccb577 and trnC2-GCA) are missing in Arabidopsis mitochondria. In addition, four genes found in Arabidopsis (ccb228, rpl2, rpl16 and trnY2-GUA) are entirely absent in sugar beet or present only in severely truncated form. Introns, duplicated sequences, additional reading frames and inserted foreign sequences (chloroplast, nuclear and plasmid DNA sequences) contribute significantly to the overall size of the sugar beet mitochondrial genome. Nevertheless, 55.6% of the genome has no obvious features of information. We identified a novel tRNA(Cys) gene (trnC2-GCA) which shows no sequence homology with any tRNA(Cys) genes reported so far in higher plants. Intriguingly, this tRNA gene is actually transcribed into a mature tRNA, whereas the native tRNA(Cys) gene (trnC1-GCA) is most likely a pseudogene. PMID- 10871409 TI - Multiple mutations and frameshifts are the hallmark of defective hPMS2 in pZ189 transfected human tumor cells. AB - Two HeLa variants defective in the mismatch repair protein hPMS2 were isolated by selection for methylation tolerance. Neither variant expressed detectable hPMS2 protein as determined by western blotting. Cell extracts were defective in correcting a single base mispair and were unable to perform mismatch repair dependent processing of a methylated DNA substrate. Correction of the repair defect and restoration of sensitivity to a methylating agent was achieved by introducing a wild-type copy of chromosome 7 on which the hPMS2 gene is located. Loss of hPMS2 function in the HeLa variants was associated with a 5-fold increase in mutation frequency in the supF gene of the pZ189 shuttle vector. Wild-type levels of mutagenesis were restored by the transferred chromosome 7. Comparisons of mutational spectra identified multiple base substitutions, frameshifts and, to a lesser extent, single base pair changes as the types of mutation which are selectively increased in a hPMS2-defective background. The location of multiple mutations and frameshifts indicates that misalignment-mediated mutagenesis could underlie most of these events. Thus the mutator phenotype associated with loss of hPMS2 most likely arises because of the failure to correct replication slippage errors. Our data also suggest that a considerable fraction of mutagenic intermediates are recognized by the hMutSbeta complex and processed via the hMLH1/hPMS2 heterodimer. PMID- 10871410 TI - DNA double-strand break repair in cell-free extracts from Ku80-deficient cells: implications for Ku serving as an alignment factor in non-homologous DNA end joining. AB - Non-homologous DNA end joining (NHEJ) is considered the major pathway of double strand break (DSB) repair in mammalian cells and depends, among other things, on the DNA end-binding Ku70/80 hetero-dimer. To investigate the function of Ku in NHEJ we have compared the ability of cell-free extracts from wild-type CHO-K1 cells, Ku80-deficient xrs6 cells and Ku80-cDNA-complemented xrs6 cells (xrs6 Ku80) to rejoin different types of DSB in vitro. While the two Ku80-proficient extracts were highly efficient and accurate in rejoining all types of DNA ends, the xrs6 extract displayed strongly decreased NHEJ efficiency and accuracy. The lack of accuracy is most evident in non-homologous terminus configurations containing 3'-overhangs that abut a 5'-overhang or blunt end. While the sequences of the 3'-overhangs are mostly preserved by fill-in DNA synthesis in the Ku80 proficient extracts, they are always completely lost in the xrs6 extract so that, instead, small deletions displaying microhomology patches at their breakpoints arise. In summary, our results are consistent with previous results from Ku deficient yeast strains and indicate that Ku may serve as an alignment factor that not only increases NHEJ efficiency but also accuracy. Furthermore, a secondary NHEJ activity is present in the absence of Ku which is error-prone and possibly assisted by base pairing interactions. PMID- 10871411 TI - Theoretical design of antisense genes with statistically increased efficacy. AB - Endogenous expression of antisense RNA represents one major way of applying antisense nucleic acids. To express antisense RNA intracellularly, recombinant antisense genes have to be designed and introduced into cells where the target RNA is encountered. Efficient annealing between the antisense RNA and the target RNA is crucial for efficacy and is strongly influenced by RNA structure. Here we extend structural rules for the design of in vitro transcribed antisense RNAs to the design of recombinant antisense genes. Intracellularly expressed antisense RNA transcripts contain a central antisense portion and additional flanking vector-derived sequences. A computer algorithm was generated to compose large sets of antisense genes, to calculate secondary structures of the transcribed sequences and to select for favorable structures of antisense RNA in terms of annealing and efficacy. The biological test system to measure efficiency of antisense genes was human immunodeficiency virus type 1 (HIV-1) replication in 293T cells. When considering the lower intracellular steady-state levels of favorably structured endogenous transcripts, an antisense effect against HIV-1 replication was observed that was up to 60-fold stronger than that measured for predicted unfavorable species. The computational selection was successful for antisense portions of 300 nt but not 100 nt in length. This theoretical design of antisense genes supports their improved application under time- and labor-saving conditions. PMID- 10871412 TI - A novel functional genomics approach identifies mTERT as a suppressor of fibroblast transformation. AB - As a tool for functional genomics, a hairpin ribozyme gene library with randomized target recognition sequences was constructed in a retroviral vector. This library has the potential to target and cleave any possible RNA substrate. Mouse fibroblasts transduced with this ribozyme gene vector library were selected in a focus formation assay to isolate in vivo functional ribozymes that promote cell transformation in tissue culture. After two successive rounds of selection by focus formation assay, a transforming ribozyme (Rz007) was identified. The sequence of this ribozyme was used to identify the putative target genes responsible for the transformation. A candidate gene target for Rz007 encodes telomerase reverse transcriptase (mTERT). Both mRNA level and enzymatic activity of mTERT were down-regulated in Rz007-transformed cells. Furthermore, newly designed ribozymes, recognizing other potential ribozyme cleavage sites unique to the mTERT mRNA, also cause cell transformation, thus validating the role of mTERT in suppressing the transformation phenotype. These surprising results suggest that the commonly accepted role of telomerase in maintaining cellular immortalization is more complicated than previously thought. These studies also demonstrate the utility of this novel 'reverse' functional genomics approach, enabling the targeted discovery of genes, whether previously known or not, that are involved in any selectable phenotype. PMID- 10871414 TI - New specificity and yield enhancer of polymerase chain reactions. AB - Tetramethylammonium (TMA) chloride, dimethyl sulfoxide and formamide are known to increase, under certain conditions, the specificity and efficiency of the polymerase chain reaction (PCR). We compared the ability of several TMA derivatives and some other reagents to increase the specificity of PCR and to improve the yield of amplification. A novel combination of the enhancer TMA and oxalate as anion is demonstrated to be a powerful enhancer of PCR. Addition of 2 mM TMA oxalate to the PCR mixture decreases the formation of non-specific DNA fragments and increases the yield of specific PCR products. PMID- 10871413 TI - New insights into the structure of abasic DNA from molecular dynamics simulations. AB - Abasic (AP) sites constitute a common form of DNA damage, arising from the spontaneous or enzymatic breakage of the N-glycosyl bond and the loss of a nucleotide base. To examine the effects of such damage on DNA structure, especially in the vicinity of the abasic sugar, four 1.5 ns molecular dynamics simulations of double-helical DNA dodecamers with and without a single abasic (tetrahydrofuran, X) lesion in a 5'-d(CXT) context have been performed and analyzed. The results indicate that the abasic site does not maintain a hole or gap in the DNA, but instead perturbs the canonical structure and induces additional flexibility close to the abasic site. In the apurinic simulations (i.e., when a pyrimidine is opposite the AP site), the abasic sugar flipped in and out of the minor groove, and the gap was water filled, except during the occurrence of a novel non-Watson-Crick C-T base pair across the abasic site. The apyrimidinic gap was not penetrated by water until the abasic sugar flipped out and remained extrahelical. Both AP helices showed kinks of 20-30 degrees at the abasic site. The Watson-Crick hydrogen bonds are more transient throughout the DNA double helices containing an abasic site. The abasic sugar displayed an unusually broad range of sugar puckers centered around the northern pucker. The increased motion of the bases and backbone near the abasic site appear to correlate with sequence-dependent helical stability. The data indicate that abasic DNA contorts more easily and in specific ways relative to unmodified DNA, an aspect likely to be important in abasic site recognition and hydrolysis. PMID- 10871415 TI - Experimental kinetic rates of food-chain and waterborne radionuclide transfer to freshwater fish: a basis for the construction of fish contamination charts. AB - A standardized procedure is proposed to obtain from laboratory experiments the kinetic accumulation and release rates necessary to calibrate dynamic models to quantify radionuclide direct and trophic transfer in fish. The model takes into account the food-chain effect, the feeding rate, and the growth of organisms. It takes as examples (54)Mn, (60)Co, and (137)Cs transfer dynamics through a simple pelagic food-chain (phytoplankton, zooplankton, prey fish, and predator fish). The estimated kinetic rates used in quantifying all the transfers of the three radioactive pollutants through the pelagic food chain are compared from the radioecological point of view. For fish, comparison was based on the calculation of concentration factors referring to direct transfer from water and trophic transfer factors. For the prey fish and the predator fish, direct transfer gave the following order for accumulation (60)Co < (137)Cs < (54)Mn. Values reached at equilibrium in L/kg WW were respectively for the prey fish and the predator fish: 8.7 < 27.4 < 107 and 4.14 < 6.59 < 13.4. For the trophic route, (137)Cs is the most accumulated (TTF(eq) = 0.485 in 291 days for the prey fish and TTF(eq) = 1.45 in 17 years for the predator fish). A sensitivity analysis adapted to the case of a chronic contamination scenario of a watercourse was run. It showed that the phytoplankton biomass, the contact time of these drifting particles from a release point to the station where they are ingested and the feeding rates of the fish are the most influential parameter with regard to the concentration in fish, whatever the trophic level. Contamination charts are constructed for the predator fish to illustrate the relationship between the most influential ecological parameters and the radionuclide concentration in fish for simple contamination scenarios. They are shown to be effective tools for helping in the choice of the most relevant value of aggregated concentration factors (ACFs: radionuclide concentration ratio between the organism and the water, referred to steady-state and to all possible transfer pathways) for a given key ecological situation in a given ecosystem. An example is given of a simple chronic release scenario of 1 Bq/L and a phytoplanktonic bloom period. For (137)Cs, the ACF increases with increasing contact time and increasing feeding rate, to nearly 550 L/kg WW at equilibrium. For (54)Mn, ACF reaches 65 L/kg WW. For (60)Co, the general pattern of the relationship is due to the rapid kinetic rates governing the distribution of the radionuclide between dissolved and solid (phytoplankton) phases with a maximum value for ACF of 7.2 L/kg WW for the case study. Analysis of these charts provides a basis for overall guidelines for chronic releases in a given watercourse. PMID- 10871416 TI - Membrane lipid composition of Bacillus stearothermophilus as affected by lipophilic environmental pollutants: an approach to membrane toxicity assessment. AB - The thermophilic eubacterium Bacillus stearothermophilus is used as a model to identify membrane perturbing effects of lipophilic compounds. A parallelism has been established between the toxicity of the organochlorine insecticide DDT and its metabolite, DDE, in bacterial growth and the effects on cell functions and physical perturbations induced at the membrane (Donato et al. 1997a, Arch Environ Contam Toxicol 33:109-116; Donato et al. 1997b, Appl Environ Microbiol 63:4948 495). In the present work, the use of B. stearothermophilus as a model of screening for chemical toxicity has been implemented. Because the regulation of the lipid composition of the membrane is a common strategy in response to adverse growth conditions, we studied the effects of DDE on the lipid composition and the consequent alterations of membrane physical properties in comparison to the parental compound DDT. As expected, different adaptation responses were induced by the compounds, being DDT more effective as compared with DDE. Collected data are consistent with the stronger perturbations induced by DDT on growth and membrane functions. It is concluded that the membrane lipid composition of the bacterium is a very sensitive criterium to detect membrane-mediated toxic effects at low concentrations of lipophilic xenobiotics. PMID- 10871417 TI - A phytotoxicity test using transpiration of willows. AB - A short-term acute toxicity assay for willow trees growing in contaminated solution or in polluted soil was developed and tested. The test apparatus consists of an Erlenmeyer flask with a prerooted tree cutting growing in it. Growth and reduction of transpiration are used to determine toxicity. Transpiration is closely related to photosynthesis and growth, but is easier and faster to measure and can be measured without disturbance of the test system. Plants are grown for 24 h in uncontaminated nutrient solution before the toxicant is added to determine the initial transpiration. The loss of weight is expressed as % decrease after 48 and 72 h or longer compared to the initial transpiration, divided by the transpiration of control plants. More toxicity parameters are growth and water use efficiency of the plants. The sensitivity of the test was evaluated with 3,5-dichlorophenol. EC(50) values between 5.8 and 9.6 mg/L were found. This is similar to the results from algal growth rate tests. The willow tree toxicity test may be useful for determining the site-specific toxicity of polluted soils and for terrestrial risk assessment of new chemicals and pesticides. PMID- 10871418 TI - Monitoring of urban traffic emissions using some physiological indicators in Ricinus communis L. plants. AB - Plants of Ricinus communis L. in the city of Porto Alegre, in southern Brazil, were exposed to urban traffic exhaust emissions for 5 months and compared with controls kept at a site essentially free of direct motor vehicle emissions. No symptomatic visible injuries were observed, but significant differences could be measured in growth, enzymatic activities of total peroxidase and nitrate reductase, chlorophyll content, leaf buffering capacity, and N contents in leaves. Additionally, these data were compared with results from fumigation experiments under controlled conditions. The study showed that some physiological parameters in R. communis L. plants could be used as an appropriate bioindicator system for urban traffic contamination, and therefore it is recommended that dose response relationships should be developed. PMID- 10871419 TI - Heavy metal toxicity and differential effects on the hyperglycemic stress response in the shrimp Palaemon elegans. AB - Agricultural and industrial activities cause heavy metal pollution of the aquatic environment. The sensitivity of crustaceans to heavy metals is well documented. However, the hormonal and metabolic target of physiological functions affected by sublethal toxicity and stress responses have been scarcely investigated. Exposure of Palaemon elegans to increasing concentrations of heavy metals dissolved in artificial sea water resulted in an order of toxicity tested by LC(50) for 96 h in intact and eyestalkless animals in which Hg is the most toxic, followed by Cd, Cu, Zn, and Pb. Eyestalkless animals were found to be more sensitive than intact individuals. Heavy metals affect the blood glucose levels, yet manipulative stress does not. The intermediate sublethal concentrations of Hg, Cd, and Pb produced significant hyperglycemic responses within 3 h, while the highest concentrations elicited no hyperglycemia in 24 h. In contrast, animals exposed to Cu and Zn showed hyperglycemia even at high concentrations. This difference in response between Cu or Zn and the nonessential heavy metals Cd, Hg, or Pb can probably be explained by the physiological roles of the former in crustaceans and by tolerance adaptations. Involvement of the crustacean hyperglycemic hormone (cHH) was tested by routine bioassay on eyestalkless individuals; each group was injected with a two-eyestalk-equivalent extract from control animals or from shrimp exposed to high concentrations of Cd, Hg, Pb, or low concentrations of Cu or Zn. All showed a hyperglycemic response within 2 h. In contrast, extracts of eyestalk removed from animals that had developed a full hyperglycemic reaction after exposure to low concentrations of Hg, Cd, Pb, or high concentrations of Cu and Zn were depleted of cHH as shown by the attenuation of the response after injection of the extracts into eyestalkless animals. This generalized and predictable sublethal response can be used as a quantitative physiological biomarker for water quality monitoring assessment. PMID- 10871420 TI - Effects of sustained-release methoprene and a combined formulation of liquid methoprene and Bacillus thuringiensis israelensis on insects in salt marshes. AB - Aquatic insects are an important component of the food web in salt marshes, therefore it is necessary to test whether pesticides used to control mosquitoes in salt marshes are safe for nontarget insects. We tested the nontarget effects of a combined formulation (duplex) of Bacillus thuringiensis israelensis (B.t.i.) and liquid methoprene (an insect development regulator) or sustained-release methoprene pellets (Altosid(R) pellets) by applying these materials to replicated salt marsh ponds at maximum label rates. Untreated ponds served as controls. We measured effects of the pesticides by rearing immature mosquitoes (Aedes dorsalis) and water boatmen (Trichocorixa reticulata) in predator-exclusion cages and by monitoring uncaged populations of invertebrates using replicated sweep-net samples. Both pesticides killed caged mosquitoes, and the activity of the Altosid(R) pellets continued through 99 days. There were no detectable effects of either pesticide on the survival or maturation of T. reticulata, or on abundances of uncaged invertebrates. The long-term activity of the pellets could help minimize mosquito abatement activity in salt marshes where there are breeding birds or endangered species. However, other studies suggest that this advantage needs to be balanced against the risks that sustained-release formulations could lead to development of resistance in mosquitoes or that initially undetected nontarget effects could build over time. PMID- 10871421 TI - Accumulation of nitrite in the tissues of Penaeus monodon exposed to elevated ambient nitrite after different time periods. AB - Penaeus monodon (29.42 +/- 0.39 g) that had been exposed individually to 0.001 (control), 0.07, 0.36, 0.72, and 1.44 mM nitrite in 25 ppt sea water were examined for the nitrite accumulation in hemolymph, gill, eyestalk, heart, foregut, midgut, hepatopancreas, and muscle and nitrite uptake after 1, 3, 6, 12, 24, and 48 h, respectively. Concentration of nitrite in the tissues increased directly with ambient nitrite and exposure time except for muscle. P. monodon following 48-h exposure to 0.36 mM nitrite, nitrite concentration progressively increased from the muscle (0.40 micromol/g), hepatopancreas (1.24 micromol/g), gill (1.82 micromol/g), foregut (2.03 micromol/g), hemolymph (0.39 micromol/mL), heart (2.43 micromol/g), eyestalk (3.07 micromol/g), and to the midgut (4.14 micromol/g), which is 1.1, 3.4, 5.0, 5.6, 6.6, 6.8, 8.5, and 11.4 times the ambient nitrite concentration, respectively. It is concluded that when P. monodon is exposed to ambient nitrite, nitrite is immediately incorporated in the hemolymph and midgut via branchial chloride uptake of NO(2)(-), and accumulated in the tissues. PMID- 10871422 TI - Detrimental effects associated with trace element uptake in lake chubsuckers (Erimyzon sucetta) exposed to polluted sediments. AB - Lake chubsuckers (Erimyzon sucetta) were exposed to coal ash-polluted sediments under conservative experimental conditions (filtered artificial soft water and abundant uncontaminated food). After 4 months of exposure, fish grazing the polluted sediments had significantly elevated body burdens of Se, Sr, and V. Selenium levels were particularly elevated, reaching mean whole body concentrations of 5.6 microg/g dry mass by the end of experimental manipulations. Twenty-five percent of fish exposed to pollutants died during the study. All surviving fish exposed to ash exhibited substantial decreases in growth and severe fin erosion. Total nonpolar lipids were two times higher in fish from the control treatment, but percent lipid did not differ between treatments. Because fish were presented with the same amount of food during the study, it appears fish exposed to ash utilized more energy for daily activities and/or were less efficient at converting available energy to tissues for growth and storage. The results were particularly interesting because we were unable to detect differences in standard metabolic rate (SMR) of fish between treatments. Increased energy expenditures not detectable in estimates of maintenance based on SMR, such as costs of digestion or activity, may have contributed to decreased energetic efficiency. Our findings corroborate previous studies which have documented the toxicity of ash-derived pollutants in fish. PMID- 10871423 TI - Mercury in feathers of Audouin's gull (Larus audouinii) chicks from northeastern Mediterranean colonies. AB - Feathers of Audouin's gull chicks from three Aegean island areas (north Dodecanese, Cyclades, Kythera) Greece, were sampled in 1997 and 1998 and analyzed for mercury. Mean concentrations varied from 0. 94 microg/g (Lipsos, Dodecanese, 1998) to 2.14 microg/g (Paros, Cyclades, 1998). Significant differences between years occurred in some regions (Lipsos, Fourni) but not in others (Paros). Within each year, especially in 1998, mean mercury concentrations differed among colonies. Results did not support the prediction that mercury levels would be higher in the north Dodecanese area due to the proximity of the polluted Menderes delta. There was no relationship between estimated chick age and feather mercury contents (r = -0.04, NS). Detected mercury levels do not seem to pose any toxic hazard to the Aegean Audouin's gull populations. However, the ease of sampling from gulls indicates that they may be a useful biomonitor of mercury contamination in this region. PMID- 10871424 TI - Transfer of DDT and metabolites from fruit orchard soils to American robins (Turdus migratorius) twenty years after agricultural use of DDT in Canada. AB - Wildlife contamination studies found high levels of DDT and associated metabolites in bird eggs from Canadian orchard sites during the early 1990s. The present study investigated local dietary uptake of DDT and geographic variability in tissue concentrations in the same orchards. A soil-earthworm-robin food chain was chosen for study, as early surveys showed that robins contained the highest levels of DDT of several avian species and because published research indicated that earthworms were a probable dietary exposure route. Organochlorine pesticides and PCBs were measured in soil, earthworm, robin egg, and robin nestling samples collected from fruit orchards and reference sites. High average DDE (soil: 5.2 mg/kg; earthworm: 52 mg/kg; robin egg: 484 mg/kg dry weight) and DDT (soil: 9.2 mg/kg; earthworm: 21 mg/kg; robin egg: 73 mg/kg dry weight) concentrations in Okanagan (British Columbia) samples confirmed that previously recorded contamination was common in the region. Concentrations detected in Simcoe, Ontario, orchards were not as high but were still significantly elevated relative to levels in soils and robins from reference areas. Significant positive linear regressions between soil and earthworm concentrations and consistent trends in food chain accumulation suggested that robins were acquiring DDT and metabolite (DDTr) burdens locally. Low concentrations of DDT and DDTr in robin eggs collected from nests in nearby nonorchard and post-DDT orchard habitats suggested that the local sources were in orchards. Persistence of DDT in orchard food chains is likely due to a combination of retarded degradation rates for DDT in soil and its extensive use historically. DDT concentrations in some robin eggs and earthworms were at levels comparable to those observed in field studies where mortality or reproductive effects occurred. PMID- 10871425 TI - Developmental toxicity of lead-contaminated sediment to mallard ducklings. AB - Sediment ingestion has been identified as an important exposure route for toxicants in waterfowl. The toxicity of lead-contaminated sediment from the Coeur d'Alene River Basin (CDARB) in Idaho was examined on posthatching development of mallard (Anas platyrhynchos) ducklings for 6 weeks. Day-old ducklings received either untreated control diet, clean sediment (24%) supplemented control diet, CDARB sediment (3,449 microg/g lead) supplemented diets at 12% or 24%, or a positive control diet containing lead acetate equivalent to that found in 24% CDARB. The 12% CDARB diet resulted in a geometric mean blood lead concentration of 1.41 ppm (WW) with over 90% depression of red blood cell ALAD activity and over threefold elevation of free erythrocyte protoporphyrin concentration. The 24% CDARB diet resulted in blood lead of 2.56 ppm with over sixfold elevation of protoporphyrin and lower brain weight. In this group the liver lead concentration was 7.92 ppm (WW), and there was a 40% increase in hepatic reduced glutathione concentration. The kidney lead concentration in this group was 7.97 ppm, and acid fast inclusion bodies were present in the kidneys of four of nine ducklings. The lead acetate positive control group was more adversely affected in most respects than the 24% CDARB group. With a less optimal diet (mixture of two thirds corn and one third standard diet), CDARB sediment was more toxic; blood lead levels were higher, body growth and liver biochemistry (TBARS) were more affected, and prevalence of acid-fast inclusion bodies increased. Lead from CDARB sediment accumulated more readily in duckling blood and liver than reported in goslings, but at given concentrations was generally less toxic to ducklings. Many of these effects are similar to ones reported in wild mallards and geese within the CDARB. PMID- 10871426 TI - Differential toxicities of organophosphate and carbamate insecticides in the nestling European starling (Sturnus vulgaris). AB - The concept of B-esterase buffering against anti-cholinesterase (ChE) insecticide toxicity has been extensively researched in mammalian species. Presumably due to relatively low levels of anti-ChE detoxifying enzyme activity in birds, however, avian species are often more susceptible to the toxic effects of these compounds. We quantified B-esterase buffering of organophosphate (diazinon and methyl parathion) and carbamate (aldicarb and oxamyl) toxicity in nestling European starlings (Sturnus vulgaris). The differential toxicities were studied using mortality, behavioral observation, and inhibitor affinity data. The toxicities of diazinon, methyl parathion, and oxamyl were affected by the removal of butyrylcholinesterase (BChE) using the specific inhibitor tetraisopropylpyrophosphoramide (iso-OMPA). When BChE was absent, aldicarb toxicity was not affected. Theoretically, compounds affected by BChE removal would have a higher affinity for BChE or carboxylesterase (CaE) than acetylcholinesterase (AChE). However, this was only the case for diazoxon, which had a 1,000-fold higher affinity for plasma BChE and CaE than AChE. Methyl paraoxon and aldicarb had a higher affinity for plasma AChE than for BChE or CaE. Oxamyl had similar IC50 values for all three enzymes studied. The generation of IC50 curves for each inhibitor revealed the presence of nonsensitive forms of CaE in both the plasma and brain. Based on the results of this research, there appears to be no strict correlation between mortality data and inhibitor affinities for each esterase that alone can explain the differential toxicities of these compounds. PMID- 10871427 TI - Increased production of proinflammatory cytokines by murine macrophages following oral exposure to sodium selenite but not to seleno-L-methionine. AB - Selenium (Se) is an essential as well as a toxic trace element in animal and human nutrition. The immune system is a known target of Se intoxication. The objectives of the present study were to determine the effects of oral exposure to inorganic and organic forms of Se on the murine immune system and to compare the relative toxicity of the different chemical forms. Male BALB/c mice, 6-7 weeks of age, were exposed continuously to 0, 1, 3 or 9 ppm of Se as sodium selenite or seleno-L-methionine in the drinking water for 14 days. Following the treatment period mice were euthanized; trunk blood, spleen, thymus, liver and kidney were aseptically collected and organs weighed. Single-cell splenocyte cultures were made from the spleens and used to determine the effects of Se treatment on mitogen-induced lymphocyte blastogenesis and cytokine production. There were no changes in the 0 and 1 ppm Se groups as selenite. The thymus/body weight ratio was significantly reduced at 3 ppm Se as sodium selenite, and all other parameters remained unaffected. Exposure to 9 ppm of Se as sodium selenite resulted in marked decrease in body weight gain and relative organ weights. Treatment of mice with 9 ppm Se as sodium selenite increased erythrocyte counts in peripheral blood, reduced splenic cellularity, but increased the basal rate of splenocyte proliferation and induced a dose-dependent increase in phytohemagglutinin-P-induced lymphocyte proliferation. Sodium selenite at this dose increased the production of proinflammatory cytokines, tumor necrosis factor alpha and interleukin-1 beta, in lipopolysaccharide-stimulated splenic macrophages. Mice exposed to Se as seleno-L-methionine in the drinking water did not display any effects on the parameters examined at the dose range in this study. Results indicated that splenic macrophages and lymphocytes are sensitive to Se intoxication and there is a disparity in the immune system toxicity of inorganic and organic forms of Se administered via the drinking water, inorganic Se being more toxic. PMID- 10871428 TI - A role for oxidative stress in suppressing serum immunoglobulin levels in lead exposed Fisher 344 rats. AB - Evidence implicating oxidative stress in toxicity during lead intoxication in vivo has opened new avenues for investigation of the mechanisms of lead-induced immunosuppression. The current study explores the possibility that lead-induced oxidative stress contributes to the immunosuppression observed during lead poisoning. Fisher 344 rats were exposed to 2,000 ppm lead acetate in their drinking water for 5 weeks. One week following removal of lead from the drinking water, significant reductions in serum levels of IgA, IgM, and IgG were detected. Significant increases in oxidative damage, based on malondialdehyde (MDA) content, were observed in peripheral blood mononuclear cells (PMCs) collected during the same experiments. In addition, MDA content increased in livers from lead-exposed rats. Following 5 weeks of lead exposure, administration of either 5.5 mmol/kg N-acetylcysteine (NAC) or 1 mmol/kg meso-2,3-dimercaptosuccinic acid (DMSA) in the drinking water for 1 week significantly reversed the inhibitory effects of lead on serum immunoglobulin (Ig) levels. Also, all parameters indicative of oxidative stress returned to control levels. These results suggest that oxidative stress contributes to suppressed serum Ig levels during lead intoxication in vivo, and that intervention with either a thiol antioxidant (NAC) or a metal chelator (DMSA) will alleviate this lead-induced suppression by correcting the prooxidant/antioxidant imbalance caused by lead exposure. PMID- 10871429 TI - The effect of sulfur dioxide inhalation on visual evoked potentials, antioxidant status, and lipid peroxidation in alloxan-induced diabetic rats. AB - The aim of the study was to investigate the effect of 10 ppm sulfur dioxide (SO(2)) exposure on visual evoked potentials (VEPs), thiobarbituric acid reactive substances (TBARS), and the activities of Cu,Zn superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in diabetes mellitus. Forty healthy male albino rats, aged 3 months, were divided into four equal groups: control (C), sulfur dioxide + control (CSO(2)), diabetic (D), and sulfur dioxide + diabetic (DSO(2)) groups. Experimental diabetes mellitus was induced by IV injection of alloxane monohydrate in a dose of 50 mg/kg body weight. Ten ppm sulfur dioxide was administered to the animals of sulfur dioxide-exposed groups in an exposure chamber for 1 h/day x 7 days/week x 6 weeks while control and diabetic groups were exposed to filtered air in the same condition. SO(2) exposure, though markedly decreasing retina CAT and GSH-Px activities, significantly increased retina Cu,Zn-SOD activity in the diabetic and nondiabetic groups. In contrast to SO(2)-related increase in the activity of Cu,Zn-SOD, decrease in GSH-Px activity was observed in the brain of those groups. Brain CAT activity was unaltered. SO(2) exposure caused the significant elevation in brain TBARS levels of CSO(2) and DSO(2) groups, whereas only in the retina TBARS level of the CSO(2) group. SO(2) exposure caused the significant prolongations of P(1), N(1), P(2), and P(3) components of VEPs in the nondiabetic and all components of VEPs in the diabetic groups. SO(2) exposure also resulted in significant amplitude reductions in both experimental groups. PMID- 10871430 TI - Comparison of persistent organic pollutant residues in serum and adipose tissue in a female population in Belgium, 1996-1998. AB - This study was performed to determine and compare persistent organic pollutant (POP) levels in different matrices in a female population. A total of 96 serum and 46 adipose tissue samples were collected from infertile women (n = 101) attending Centers for Reproductive Medicine in Belgium from 1996 to 1998. Gas chromatography with electron-capture detection was used to quantify residue levels on a lipid basis of seven organochlorine pesticides (OCPs) and seven polychlorinated biphenyls (PCBs). There was a strong association between adipose tissue and serum residues. The adipose tissue levels of CB-138, 153, 180 and p,p' DDE were explained by the serum residues. Besides, the accumulation pattern for CB-153 and CB-180 in serum and adipose tissue are mirror images of each other. PMID- 10871431 TI - Transient increase in alveolar epithelial permeability induced by volatile anesthesia with isoflurane. AB - Fifteen patients undergoing surgery and receiving volatile anesthesia with isoflurane were enrolled as the study group. At the same time, 15 patients undergoing surgery with intravenous anesthesia drugs were included as a control group to compare each other. Before surgery, 1 h after surgery, and 1 week after surgery, we investigated these two groups of patients with technetium-99m-labeled diethylene triamine pentaacetic acid radioaerosol inhalation lung scan (DTPA lung scan), a test to evaluate lung ventilation (LV), which was evaluated by the first and equilibrium lung ventilation image and alveolar epithelial permeability (AP) which was evaluated by the half time (T1/2, minutes) of Tc-99m DTPA radioaerosol lung clearance. No significant change and abnormality of LV before surgery, 1 h after surgery, or 1 week after surgery was found in either group of patients. In addition, no significant change in AP before surgery (T1/2 = 64.0 +/- 17.3 min), 1 h after surgery (64.5 +/- 19.6 min), or 1 week after surgery (63.6 +/- 17.6 min) was found among the control group patients (p values > 0.05). However, a significant transient increase in AP was found in the study group 1 h after surgery (71.7 +/- 17.5 versus 51.2 +/- 16.4 min), but it recovered 1 week after surgery (51.2 +/- 16.4 versus 70.9 +/- 16.0 min) (p values < 0.05). We conclude that volatile anesthesia with isoflurane can induce transient increase of AP. PMID- 10871432 TI - Leukotoxin, 9,10-epoxy-12-octadecenoate, causes pulmonary vasodilation by stimulation of vascular eNOS and iNOS. AB - We have previously reported that leukotoxin, 9,10-epoxy-12-octadecenoate (Lx) dilates rat pulmonary arteries by means of nitric oxide synthase (NOS) activation. In this study, we investigated if Lx stimulates constitutive and/or inducible NOS. We studied the effect of the NOS inhibitors, N(G)-monomethyl-L arginine and aminoguanidine, as well as endothelium denudation on Lx-induced rat pulmonary arterial dilation and that of aminoguanidine on Lx-induced endothelium denuded lipopolysaccharide (LPS)-treated rat pulmonary arterial dilation and tissue cGMP content. Furthermore, we assessed the effect of aminoguanidine, an inducible NOS (iNOS) inhibitor, on the cGMP content increase induced by Lx in LPS treated human pulmonary artery smooth muscle cells (HPASMC). The NOS inhibitors and endothelium denudation significantly attenuated Lx-induced vasodilation. Aminoguanidine also significantly attenuated Lx-induced vasodilation in LPS treated rat denuded pulmonary arteries, and attenuated Lx-induced cGMP content increase in denuded pulmonary arterial rings from LPS-treated rats and in LPS treated HPASMC. These results suggest that Lx causes pulmonary vasodilation by stimulation of vascular endothelial NOS (eNOS) and iNOS. PMID- 10871433 TI - Interleukin-9 receptor expression in asthmatic airways In vivo. AB - Inflammation of the airway wall is a defining feature in asthma and is likely the cause of the hyperreactivity and variable airflow limitation found in asthma. Immune response biased toward production of Th2 cytokines has been proposed as a mechanism in the pathogenesis of airway inflammation in asthma. The Th2 cytokine interleukin-9 (IL-9) is one candidate gene for asthma on the basis of position cloning and animal models of airway inflammation. To determine whether IL-9 is involved in the chronic inflammation of the asthmatic airway, we investigated the expression of IL-9 and the IL-9 specific receptor chain in asthmatic airways compared with healthy airways. IL-9 and IL-9 receptor expression in airway epithelial cells and bronchoalveolar lavage cells obtained at bronchoscopy of healthy (n = 9) and mild intermittent asthmatic individuals (n = 7) were studied by Northern analyses and reverse-transcription polymerase chain reaction technique. Primary and transformed human airway epithelial cells were also evaluated for IL-9 specific receptor chain expression in vitro. IL-9 was not detected in airways of healthy or mild asthmatic individuals. In contrast, IL-9 specific receptor chain expression was found in asthmatic airway samples but not in healthy controls. In vitro, airway epithelial cells did not express IL-9 specific receptor chain until stimulation with interferon gamma. Our results support that IL-9 may play a role in the mechanism leading to chronic airway inflammation and asthma. PMID- 10871434 TI - Thoracic involvement in Behcet's disease and its correlation with multiple parameters. AB - In Behcet's disease (BD), controversy has existed over the incidence of thoracic involvement, which may be a direct threat to the patient's life. The aim of this study is to evaluate the incidence of thoracic involvement in BD and its correlation with the number of diagnostic BD criteria of The International Study Group (ISG), gender, disease duration, and the presence of symptoms. Forty-two BD patients, who had consecutively applied to different clinics in Turgut Ozal Medical Center Research Hospital, were included in the study. They were either newly diagnosed or already under treatment. All patients were examined by standard chest roentgenogram, spirometry, and thorax CT. Perfusion scintigraphies were obtained in patients with thoracic involvement. Thoracic pathologic conditions were found in five patients (11.9%). All thoracic pathologic conditions appeared in patients with at least four diagnostic criteria (26 patients) of the ISG for BD. In this subgroup, the rate of thoracic involvement was 19.2%. Also, 25% of the patients with pulmonary symptoms (12 patients) had thoracic lesions. Gender and the duration of the disease did not correlate with thoracic involvement. Our findings suggest that the rate of thoracic involvement in BD is greater than is generally believed. An increased number of diagnostic BD criteria of the ISG may indicate other organ system involvement and an increased risk of thoracic pathosis. All BD patients with at least four diagnostic criteria or any pulmonary symptoms should be evaluated for thoracic involvement, which is a major menace to life and necessitates early intervention. PMID- 10871435 TI - Elevation of anti-cytokeratin 18 antibody and circulating cytokeratin 18: anti cytokeratin 18 antibody immune complexes in sera of patients with idiopathic pulmonary fibrosis. AB - In this study, we hypothesize that anti-cytokeratin 18 (CK18) antibody and CK18:anti-CK18 immune complex increase in sera in patients with idiopathic pulmonary fibrosis (IPF). To prove the existence of anti-CK18 antibodies in patients' sera, bovine CK18 was stained with patients' sera using a Western blotting. In patients with IPF, anti-CK18 antibodies were clearly demonstrated in sera by Western blotting. Then, we tried to establish an enzyme-linked immunosorbent assay (ELISA) to quantify anti-CK18 antibodies and CK18:anti-CK18 immune complexes in sera of patients with IPF. Levels of anti-human CK18 antibodies in sera of patients with IPF (0.81 +/- 0.31, mean +/- SD) measured by ELISA were significantly high compared with that of normal volunteers (0.45 +/- 0.06, p < 0.01). In addition, levels of CK18:anti-CK18 antibody complexes in patients' sera (0.64 +/- 0.35, man +/- SD) significantly increased compared with those of normal subjects (0.40 +/- 0.10, p < 0.01). These results suggest that anti-CK18 antibody and its immune complex may have played a role in the process of lung injury in IPF. PMID- 10871436 TI - Detection of elastase from Pseudomonas aeruginosa in sputum and its potential role in epithelial cell permeability. AB - Most clinical strains of Pseudomonas aeruginosa produce elastase, a zinc metalloprotease that is implicated in the pathogenesis of infections related to these organisms. To better understand the physiologic role of this protease in the regulation of airway permeability, we developed a panel of specific monoclonal antibodies (mAb) against purified Pseudomonas elastase (PE) that do not react with either neutrophil elastase or porcine pancreatic elastase. These mAbs were used in a competitive enzyme-linked immunosorbent assay to determine the concentrations of PE in sputum samples from patients with pulmonary infections. Sputum from patients infected with P. aeruginosa showed a varying amount of PE, whereas others indicated no signals. We also found that the mAbs blocked the effect of PE on epithelial barrier function in vitro on the basis of measurement of transmonolayer electrical resistance of polarized epithelial cells as an index of paracellular permeability. PMID- 10871438 TI - Go, girl!: presidential address. PMID- 10871439 TI - The significance of atypical glandular cells on routine cervical cytologic testing in a community-based population. AB - OBJECTIVES: We sought to determine the follow-up rate of women with glandular atypia on routine Papanicolaou smears in a community-based population and to describe the associated pathologic findings. STUDY DESIGN: Over a 12-month period, all patients with Papanicolaou smears with atypical glandular cells of undetermined significance were reviewed for demographic and clinical characteristics and followed up for a period of 12 to 24 months. RESULTS: Of the 48,890 Papanicolaou smears examined, 141 (0.29%) were diagnosed with atypical glandular cells of undetermined significance. Of these, 22 (17.6%) had no record of any subsequent investigation, and only 64 (51.2%) were monitored with both colposcopy and biopsy. Of the 64 biopsy specimens, 39 (60.9%) were positive for disease. Twenty-six (66.7%) were of squamous origin, with the most advanced lesion being cervical intraepithelial neoplasia 3. An additional patient had a combined cervical intraepithelial neoplasia and adenocarcinoma in situ lesion. Four (10.3%) additional patients had glandular cervical lesions, 2 benign polyps and 2 adenocarcinoma in situ lesions. Seven (17.9%) patients had endometrial lesions (benign polyps, 2 patients; complex atypical endometrial hyperplasia, 1 patient; and endometrial carcinoma, 4 patients). One patient had ovarian cystadenocarcinoma. Postmenopausal women were 5 times more likely to have a glandular lesion. Women with abnormal vaginal bleeding were also more likely to have a glandular lesion. These same patient groups were also more likely to have endometrial disease. CONCLUSION: The incidence of atypical glandular cells of undetermined significance on Papanicolaou smears in this community-based population was 0.29%, which is consistent with estimates from institution-based populations. Nearly 50% of women studied were not followed up with tissue biopsy. Of those with a tissue biopsy, 61% had positive findings, including 5 with cancer. Although postmenopausal status and abnormal vaginal bleeding were associated with endometrial or glandular disease, studies of larger patient populations should be conducted to examine potential risk factors for these conditions. PMID- 10871440 TI - Benefits, risks, costs, and patient satisfaction associated with insulin pump therapy for the pregnancy complicated by type 1 diabetes mellitus. AB - OBJECTIVE: Glycemic control, perinatal outcome, and health care costs were evaluated among women with type 1 diabetes mellitus who began insulin pump therapy during pregnancy (group 1, n = 24), were treated with multiple insulin injections (group 2, n = 24), or were already using an insulin pump before pregnancy (group 3, n = 12). Patient satisfaction and continuation of pump therapy post partum were assessed. STUDY DESIGN: A retrospective review of maternal and neonatal medical records was performed, and a questionnaire was sent to patients after delivery. Patients in groups 1 and 2 were matched for age, age at onset and duration of diabetes mellitus, White class, and date of delivery. RESULTS: No differences in glycosylated hemoglobin A levels were observed among groups 1, 2 or 3 in the first, second, or third trimester. Patients in group 1 started pump therapy at a mean of 16.8 weeks' gestation, and 17 (70.8%) began therapy as outpatients. No deterioration in glycemic control was noted during the 2- to 4-week period after the start of pump treatment. Among the women in group 1 eight had at least one episode of severe hypoglycemia before starting pump therapy, but only one had such an episode after this treatment was begun. Two episodes of ketoacidosis occurred in group 1, and no episodes occurred in groups 2 and 3. No significant differences in perinatal outcomes or health care costs were observed among groups 1, 2, and 3. After delivery 94. 7% of the women in group 1 continued to use the pump because it provided better glycemic control and a more flexible lifestyle. Postpartum glycosylated hemoglobin A values were 7.2% in group 1 and 9.1% in group 2, a significant difference. CONCLUSIONS: Insulin pump therapy was initiated during pregnancy without a deterioration of glycemic control and was associated with maternal and perinatal outcomes and health care costs comparable to those among women who were already using the pump before pregnancy or who received multiple-dose insulin therapy. Women who began pump therapy in pregnancy were highly likely to continue pump use after delivery and preferred the flexible lifestyle that this treatment allowed. PMID- 10871441 TI - Acceptability of suction curettage and mifepristone abortion in the United States: a prospective comparison study. AB - OBJECTIVE: We sought to compare the acceptability of suction curettage abortion with that of medical abortion with mifepristone and misoprostol in American women. STUDY DESIGN: We performed a prospective, serially enrolled, cohort analysis. The study population consisted of 152 subjects receiving mifepristone and misoprostol and 174 subjects undergoing suction curettage abortion aged > or =18 years with intrauterine pregnancies of up to 63 days' estimated gestation. Questionnaires regarding expectations and experiences were administered before the abortion and at the 2-week follow-up visit. RESULTS: Subjects undergoing medical abortions reported significantly greater satisfaction than those undergoing surgical abortions (mean rank, 121 vs 149; P <.01) but were no more likely to recommend the method they had just experienced to a friend (97% vs 93.3%). If a future abortion was required, however, 41.7% of subjects undergoing surgical abortions indicated they would opt for a medical abortion, whereas only 8.6% of subjects receiving medical abortions would choose a surgical abortion (P <.001). Failure of the abortion decreased satisfaction in the medical group and increased the likelihood of choosing a surgical abortion for a subsequent procedure (P <.001). Surgical subjects who experienced more anxiety than expected during the abortion were more likely to choose a medical procedure for a subsequent abortion (P <.01). CONCLUSION: Women receiving mifepristone and misoprostol were more satisfied with their method and more likely to choose the same method again than were subjects undergoing surgical abortion. Failure of a medical abortion and increased anxiety during surgical abortion were associated with preference for the alternative technique in a future procedure. PMID- 10871442 TI - Lack of utility in clinical practice of cytologic examination of nonbloody cyst fluid from palpable breast cysts. AB - OBJECTIVE: This study was undertaken to answer the following question: Does cytologic evaluation of nonbloody fluid aspirated from breast cysts contribute to appropriate clinical management? STUDY DESIGN: A retrospective review of palpable breast cyst fluid cytologic reports and associated medical records was undertaken to determine whether the cytologic findings affected patient management. Breast cyst size, fluid volume, fluid color, and patient age were abstracted from 689 medical records (1988-1999) of women whose palpable cysts had been aspirated at the Breast Diagnostic Center, Women's and Children's Hospital, Los Angeles. These observations were correlated with the fluid cytologic reports. RESULTS: Except for frankly bloody fluid, all breast fluid cytologic reports listed the results as acellular, inadequate for cytologic diagnosis, or no malignant cells identified. CONCLUSION: In clinical practice only frankly bloody fluid should be submitted for cytologic analysis. All other cyst fluid should be discarded. PMID- 10871443 TI - Obstetric management of a protracted labor in a captive western lowland gorilla. AB - This article discusses the cooperative efforts of a team of physicians and veterinarians resulting in the successful assisted vaginal delivery of a Western lowland gorilla at the Woodland Park Zoo in Seattle, Washington. A 10-year-old, captive-born female gorilla, gravida 3, para 0, aborta 2, was observed to be in labor at term after spontaneous rupture of membranes. After 36 hours of observation, she had not yet delivered her infant. A team of physicians and veterinarians intervened. After induction of general anesthesia, an assessment of fetal and maternal status was made. With ultrasonographic monitoring of fetal cardiac activity, labor was augmented with administration of intravenous oxytocin. A vaginal delivery was performed with a vacuum extractor, resulting in the birth of a viable, 2.4-kg female infant. The infant survived the neonatal period and was hand reared until she was successfully introduced to the gorilla troop at the age of 1 year. Although there are several cases of cesarean delivery in captive gorillas, this is the first reported use of labor augmentation and assisted vaginal delivery in this captive species. Captive breeding is the key to survival of the Western lowland gorilla. The collaborative work of physicians and veterinarians is an integral part of this successful program. It is reported that in the captive population approximately 30% of newborns die in the first year, with more than half of the deaths occurring just before birth or within the first day of life. This does not include early spontaneous pregnancy losses. With aggressive management of pregnancy and labor complications, it may be possible to reduce perinatal morbidity and mortality in the captive gorilla population. Because gorillas are closely related to humans, with the only closer relative being the chimpanzee, it is probable that the basic principles of obstetric management in humans can be safely and appropriately applied to the captive gorilla population. With the exception of this report and the cited cesarean delivery reports, there are no other references to cross-species studies of active intervention in gorillas with human obstetric techniques. It is hoped that an increased awareness on the part of obstetricians of the importance of their knowledge and skill to zoos will lead to more successful outcomes in the captive gorilla population. PMID- 10871444 TI - The relationship between maternal age and uterine dysfunction: a continuous effect throughout reproductive life. AB - OBJECTIVE: In a selected low-risk population with spontaneous term labor we sought to determine whether there was a continuous effect of maternal age on uterine function. STUDY DESIGN: With our comprehensive computerized database and medical record system, we identified 8496 patients who were nulliparous and in spontaneous labor at term (> or =37 weeks' gestation) with singleton fetuses in vertex presentation. This group was then analyzed according to maternal age for measures of labor dysfunction and rates of operative delivery. Analysis of variance and chi(2) statistics were used. RESULTS: Use of oxytocin, duration of second stage of labor, cesarean delivery, cesarean delivery for failure to progress, and operative vaginal delivery rates were significantly increased with advancing maternal age (P <.0001). These increases appeared to be continuous functions beginning during the early 20s rather than new phenomena beginning after age 35 years. CONCLUSION: Among nulliparous patients with uncomplicated labor there is a continuously increasing risk of uterine dysfunction related to maternal age. PMID- 10871445 TI - Second-look laparotomy after modified posterior exenteration: patterns of persistence and recurrence in patients with stage III and stage IV ovarian cancer. AB - OBJECTIVE: The purpose of this study was to determine patterns of persistence and recurrence in patients with advanced ovarian cancer (stage IIIC and stage IV) after modified posterior exenteration. STUDY DESIGN: Retrospective chart review was used to determine patterns of persistence and recurrence of disease in patients undergoing modified posterior exenteration. From January 1, 1987, to September 15, 1998, 151 of 212 (71.2%) patients undergoing modified posterior exenteration in addition to other cytoreductive surgical procedures for stage IIIC and stage IV ovarian cancer underwent second-look laparotomy. The average age of the patients was 60.3 years (range, 20.3-86.3). A total of 207 of the 212 (97.6%) had grade 2 or 3 disease. Papillary serous carcinoma (113/212; 53.3%) and adenocarcinoma (75/212; 35.4%) were the most frequent cell types encountered. After initial cytoreductive surgery, minimal disease (<5 mm) was present in 206 of the 212 (96.2%) patients with 153 of 212 (72.2%) having no visible residual disease. There were 4 (1.9%) postoperative deaths. In 13 patients (6.1%) progressive disease was noted. Second-look laparotomy was not undertaken in 61 of the 212 (28%) patients. Fluid for cytologic testing was obtained from all four intra-abdominal quadrants, and biopsies of previously noted sites of disease were performed, in addition to random biopsies of diaphragmatic peritoneum, colonic gutters, and pelvic peritoneum. If present, the retroperitoneal lymph nodes were resected; biopsy specimens of these sites were obtained if there was no evidence of intraperitoneal disease. RESULTS: Findings at second-look laparotomy were negative for cancer in 85 of 151 (56.3%) and positive for cancer in 66 of 151 (43.7%). Only 8 of 151 (5.3%) patients had persistent disease in the pelvis. In the remainder (58/151; 38.4%) disease was found either in the upper abdomen or in the bowel mesentery. Recurrence was documented in the upper abdomen only (71/212; 33.5%), upper abdomen and pelvis (18/212; 8.5%), multiple sites excluding the pelvis (22/212; 10.4%), pelvis only (2/212; 0. 9%), chest alone (5/212; 2.4%), head alone (4/212; 1.9%), or groin alone (2/212; 0.9%). Median survival in the overall group was 51.1 months, with estimated 5- and 10-year survival rates of 44.2% and 32. 9%, respectively. CONCLUSIONS: Modified posterior exenteration is an effective surgical means of eliminating pelvic disease in patients with advanced ovarian cancer. Results of second-look laparotomy confirmed that only 8 of 151 (5.3%) had persistent disease in the pelvis. PMID- 10871446 TI - Preoperative CA 125 in endometrial cancer: is it useful? AB - OBJECTIVE: We sought to determine the clinical utility of preoperative CA 125 measurement in determining the need for lymphadenectomy in patients with endometrial carcinoma. STUDY DESIGN: A prospective nonrandomized study was performed over a 2-year period. Patients referred with the diagnosis of endometrial carcinoma had CA 125 levels determined before surgical staging. Operative findings were then correlated with preoperative CA 125 values. Standard statistical calculations were used to determine sensitivity, specificity, positive predictive value, and false-positive and false-negative rates. The Student t test was used to determine differences between mean values. RESULTS: Either a CA 125 level of >20 U/mL or a grade 3 tumor or both of these correctly predicted 87% of patients requiring surgical staging. In patients with a preoperative diagnosis of stage I, grade 1 or 2 tumors, a CA 125 level of >20 U/mL correctly identified 75% (9/12) of patients requiring lymphadenectomy compared with only 50% (6/12) identified when a CA 125 level of >35 U/mL was used. Two of 16 low-risk patients with preoperative grade 1 tumors and CA 125 levels of <20 U/mL had occult extrauterine disease at surgery. CONCLUSION: Measurement of preoperative CA 125 is a clinically useful test in endometrial cancer. CA 125 levels of >35 U/mL strongly predicted extrauterine disease but lacked sensitivity in identifying patients needing staging. Either a CA 125 level of >20 U/mL or a grade 3 tumor or both of these correctly identified 75% to 87% of patients requiring lymphadenectomy. Until more data are collected, abdominal hysterectomy should be the procedure of choice for patients with grade 1 tumors and CA 125 levels of <20 U/mL. PMID- 10871447 TI - Performance of a group B streptococcal prophylaxis protocol combining high-risk treatment and low-risk screening. AB - OBJECTIVE: This study was undertaken to evaluate a group B streptococcal protocol in a large community hospital that combined treatment of high-risk patients with rapid screening of low-risk patients. STUDY DESIGN: In a prospective cohort study from 1994 through 1996 laboring patients in a level III community hospital were considered to be at high risk for neonatal group B streptococcal transmission if they were at <37 weeks' gestation, if they had rupture of membranes >12 hours, if they were known carriers of group B streptococci, if they had a temperature > or =100 degrees F, if the gestation was complicated by fetal growth restriction or was a multiple gestation, or if they had a previous neonate infected with group B streptococci. High-risk patients were treated intravenously with antibiotics during labor. Low-risk patients were screened for group B streptococcal antigen by means of a rapid optical immunoassay. Patients with positive screening results were treated. Neonatal morbidity and mortality were evaluated. RESULTS: Two of 9932 infants delivered had group B streptococcal sepsis diagnosed. In the 2 previous years without a protocol 9 cases of neonatal group B streptococcal sepsis had been diagnosed in 8188 deliveries (P =.0287 by Fisher exact test). The 2 cases of group B streptococcal sepsis during the protocol were as follows: 1 infant born to a high-risk mother with delay in treatment and 1 infant born to a low-risk mother with negative results of both culture and rapid screen during labor. During the previous period 7 infected infants had been born to high-risk mothers and 2 had been born to low-risk mothers. The maternal group B streptococcal carriage rate during the study was 18%. Group B streptococcal rapid optical immunoassay sensitivity was 81%. Elapsed time from screening to treatment was < or =2(1/2) hours for 93% of patients. No maternal anaphylaxis, no increase in bacterial neonatal sepsis caused by organisms other than group B streptococci, and no protocol-related group B streptococcal antibiotic resistance were noted. CONCLUSION: Successful implementation and maintenance of a protocol combining treatment of high-risk patients with rapid screening of low-risk patients during labor reduced neonatal group B streptococcal sepsis. PMID- 10871448 TI - Prevention of early-onset invasive neonatal group B streptococcal disease in a private hospital setting: the superiority of culture-based protocols. AB - OBJECTIVE: In a large private tertiary care hospital we compared the two different approaches to group B streptococcal screening and intrapartum chemoprophylaxis suggested by The American College of Obstetricians and Gynecologists, the American Academy of Pediatrics, and the Centers for Disease Control and Prevention: risk factor-based protocol and culture-based protocol. STUDY DESIGN: A 2-year baseline period was followed by sequential prospective observational studies of the impacts of two different group B streptococcal management protocols, 3 years with the risk-based approach and 2 years with the culture-based approach of universal screening at 35 to 37 weeks' gestation. RESULTS: During the baseline period the rate of early-onset group B streptococcal infection was 1. 1 cases per 1000 births (n = 8 cases per 6829 births). With the risk-based strategy the rate was also 1.1 cases per 1000 births (15 cases/13,270 births). After we switched to the culture-based protocol for 2 years, there were no cases of early-onset group B streptococcal infections among 9304 births (P =.001; chi(2) = 10.9). There were no increases in other early-onset infections or in antibiotic resistance. CONCLUSIONS: In our setting, which included good prenatal care and good communication between laboratories and the hospital, the approach based on maternal culture at 35 to 37 weeks' gestation and treatment during labor of all patients with positive results significantly reduced early onset group B streptococcal infections without increasing infections from resistant organisms. PMID- 10871449 TI - The epidemiology of labor induction: Arizona, 1997. AB - OBJECTIVE: This study was undertaken to describe labor induction risk factors and consequences among women with term singleton gestations with vertex presentation. STUDY DESIGN: Arizona births in 1997 (N = 65,607) were studied by means of stratified analysis and logistic regression. RESULTS: Labor induction occurred in 20.3% (n = 13,288). Labor induction risk factors were as follows: race or ethnicity (white non-Hispanic 25.3%; Hispanic, 13.9%; foreign-born Hispanic, 10.3%; and US-born Hispanic, 18.5%), education (<12 years, 14.1%; >12 years, 24.6%), payor (private insurance, 24.5%; Medicaid, 16.7%), hospital type (government controlled, 13.7%; investor owned, 30.5%). Race or ethnicity and hospital type remained important determinants of labor induction in the multivariate analysis. Relative risks of cesarean delivery with labor induction were as follows: nulliparous, 1.38; parous with no previous cesarean delivery, 1.00; and parous with previous cesarean delivery, 0.50. CONCLUSION: Large variations in labor induction were noted across maternal ethnicity and hospital type categories. Labor induction increased cesarean delivery rates among nulliparous women, whereas no increase was seen among parous women with no previous cesarean delivery. Labor induction was used less often among those with previous cesarean delivery; when it was used in this group, however, it was associated with a lower cesarean delivery rate. PMID- 10871450 TI - Maternal birth weight and cesarean delivery in four race-ethnic groups. AB - OBJECTIVE: We hypothesized that maternal birth weight was associated with the risk of cesarean delivery for nulliparous women. STUDY DESIGN: In a population based cohort study, maternal birth data were linked to a Washington State database, including the birth certificates of 18,905 first-born singleton infants (1987-1995). RESULTS: Among non-Hispanic white subjects, maternal birth weight of 2500 to 3999 g was associated with a 20.9% risk of cesarean delivery, which was the lowest risk, compared with 24.5% for a maternal birth weight <2500 g (P <.05) and 24.0% for a maternal birth weight > or =4000 g (P <.05). Similar patterns of risk were noted among Hispanic and Native American subjects, although the associations did not reach statistical significance. Risk of cesarean delivery was not associated with maternal birth weight among African American subjects. Among non-Hispanic white subjects, the risk of cesarean delivery was 3.23 times greater with a maternal birth weight <2500 g and an infant birth weight > or =4000 g compared with pregnancies with both maternal and infant birth weights between 2500 and 3999 g (P <. 001). Adjustment for socioeconomic factors did not alter these results. CONCLUSION: Low and high maternal birth weights exert an intergenerational risk of cesarean delivery in nulliparous non-Hispanic white women. PMID- 10871451 TI - Bone mineral density in grand multiparous women with extended lactation. AB - OBJECTIVE: We sought to assess whether the accumulation of multiple, frequent pregnancies and the accompanying repeated extended lactation events was a risk factor for low bone mineral density and osteoporosis. STUDY DESIGN: The study population consisted of 30 grand multiparous women who had borne at least 6 children and lactated for at least 6 months with each child, as well as 6 nulliparous, premenopausal women from a population of Finnish American women associated with the Laestadian Church in Washington State. The Church membership has not embraced contraception or extensive bottle-feeding, resulting in a group of women who are either pregnant or lactating during most of their adult reproductive lives. The medical history included the delivery date, birth outcome, infant birth weight, and number of months lactated for each pregnancy, as well as other health information. Bone mineral density of the lumbar spine, femoral neck, and radius was measured with the Hologic QDR 4500-C dual-energy x ray absorptiometry scanner. Proc Genmod, SAS version 6.14 (Statistical Analysis Systems, Inc, Cary, NC), was used to perform a Wilcoxon test for a nonparametric analysis of covariance and significance adjusted for age and body size. RESULTS: The 2 study groups did not differ in terms of body mass index, history of smoking, or family history of osteoporosis and fracture, although the parous group was, on average, 8 years older than the nulliparous group (P <.05). The accumulation of repeated pregnancy and lactation events without a recovery interval was not associated with lowered bone mineral density or the presence of osteoporosis or osteopenia. CONCLUSIONS: This study suggests that bone mineral density levels can be sustained in the presence of the rapidly changing hormone environment associated with multiple pregnancies accompanying lactation events without a "recovery" interval. PMID- 10871452 TI - Urodynamic outcome after surgery for severe prolapse and potential stress incontinence. AB - OBJECTIVE: Women with severe prolapse may be paradoxically continent because of kinking of the urethra. It is currently a common practice to perform urethropexy in women who demonstrate stress incontinence on preoperative reduction of the prolapse with a pessary. We compared the urodynamic outcomes after reconstructive operations that included suspending urethropexy with outcomes after those that did not. STUDY DESIGN: A review was performed of the charts of the Gynecologic Urology Clinic at Los Angeles County-University of Southern California Women's and Children's Hospital from 1991-1997 of patients with grade III uterovaginal prolapse or procidentia in whom the pessary test was used to determine whether urethropexy was included in the reconstructive operation. Urodynamic outcomes were compared statistically with the Fisher exact test, and P < or =.05 denoted statistical significance. RESULTS: Fifty-five patients underwent urethropexy in addition to repair of the prolapse, and 70 underwent reconstruction alone. Twenty three patients in the first group and 20 in the second were available for a mean urodynamic follow-up of 3.5 years. In the urethropexy group 7 (30%) patients had de novo detrusor instability and 1 (4%) had stress incontinence. In the reconstruction-only group 1 (5%) patient had detrusor instability and none had stress incontinence. CONCLUSIONS: Preoperative barrier testing is useful in identifying patients who do not require an antiincontinence procedure. Prophylactic Burch retropubic urethropexy increases the incidence of bladder instability. PMID- 10871453 TI - Transrectal ultrasonographically guided drainage of gynecologic pelvic abscesses. AB - OBJECTIVE: This study assessed the feasibility of ultrasonographically guided transrectal aspiration of gynecologic pelvic abscesses to treat patients for whom intravenous antibiotic therapies failed and whose abscesses were not optimally amenable to colpotomy drainage or transabdominal or transvaginal ultrasonographically guided aspiration. STUDY DESIGN: This was a retrospective review of the first 15 women with pelvic abscesses that resulted from salpingitis or complications of gynecologic surgery who underwent transrectal pelvic abscess drainage after failure of antibiotic therapy. RESULTS: Purulent material was aspirated from the abscesses in 14 of the 15 women. All 14 women with aspirated material were successfully treated with real-time ultrasonographically guided transrectal drainage; only 4 of the 14 had indwelling catheter placement. CONCLUSION: Ultrasonographically guided transrectal drainage of gynecologic pelvic abscesses is a safe and effective treatment of pelvic abscesses for women who do not have an adequate response to antibiotic therapy. PMID- 10871454 TI - Preeclampsia into eclampsia: toward a new paradigm. AB - OBJECTIVE: This study was undertaken to characterize aspects of the natural history of eclampsia. STUDY DESIGN: A retrospective analysis was performed on the records of patients with eclampsia who were delivered at two tertiary care hospitals. RESULTS: Fifty-three pregnancies complicated by eclampsia were identified. Thirty-seven of the women were nulliparous. The mean age was 22 years (range, 15-38 years). Mean gestational age at the time of seizures was 34.2 weeks' gestation (range, 22-43 weeks' gestation). Twenty-eight women had antepartum seizures (53%); 23 of the 28 had seizures at home. Nineteen women had intrapartum seizures (36%). Eight of these women had seizures while receiving magnesium sulfate, and 7 had therapeutic magnesium levels. Six women had postpartum seizures (11%), 4 >24 hours after delivery. Headache preceded seizures in 34 cases. Visual disturbance preceded seizures in 16 cases. The uric acid level was elevated to >6 mg/dL in 43 women. There were no maternal deaths or permanent morbidities. There were 4 perinatal deaths. Two patients had intrauterine fetal deaths at 28 and 36 weeks' gestation. These mothers had seizures at home. One infant died of complications of prematurity at 22 weeks' gestation and one died of respiratory complications at 26 weeks' gestation. There were 4 cases of abruptio placentae, 1 of which resulted in fetal death. Of the 53 cases of eclampsia, only 9 were potentially preventable. One of these was that of a woman who was being observed at home. The other 8 women were hospitalized and had hypertension and proteinuria. Only 7 women could be considered to have severe preeclampsia before seizure (13%), and 4 of these 7 women were receiving magnesium sulfate. CONCLUSIONS: Eclampsia was not found to be a progression from severe preeclampsia. In 32 of 53 cases (60%) seizures were the first signs of preeclampsia. In this series eclampsia appeared to be more of a subset of preeclampsia. Only 9 cases of eclampsia were potentially preventable with current standards of practice. Our paradigm for this disease, as well as our approach to seizure prophylaxis, should be reevaluated. PMID- 10871455 TI - The safety and utility of pulmonary artery catheterization in severe preeclampsia and eclampsia. AB - OBJECTIVE: The objective of this research was to study the safety and utility of pulmonary artery catheterization in the management of severe preeclampsia and eclampsia. STUDY DESIGN: In a retrospective chart review from January 1, 1995, through December 31, 1997, a total of 115 patients admitted to the obstetric intensive care unit at Groote Schuur Hospital were found to have required placement of a pulmonary artery catheter. From this population 100 maternal charts were examined for medical and pregnancy history, including indication for pulmonary artery catheter placement, hemodynamic readings, complications, and subsequent management. RESULTS: The initial indications for pulmonary artery catheter placement in cases of severe preeclampsia or eclampsia were renal failure in 53 cases (53%), pulmonary edema in 30 (30%), and eclampsia in 17 (17%). Subjective evaluation demonstrated that the pulmonary artery catheter was helpful in determining management in 93 cases (93%). There was a 4.0% complication rate, which included three venous thromboses and one case of cellulitis. Eleven patients required dialysis, and 3 women died. The mean (+/-SE) duration of catheter placement was 2.1 +/- 0.1 days and the mean (+/-SE) intensive care unit and hospital stays were 3.4 +/- 0.2 days and 11.4 +/- 0.8 days, respectively. The pulmonary artery catheter measurements of pulmonary artery wedge pressure and central venous pressure were increased in the cases of pulmonary edema (21.0 +/- 2.0 mm Hg and 9. 6 +/- 1.2 mm Hg, respectively) but were normal in the cases of renal failure and eclampsia. CONCLUSION: Despite significant maternal morbidity and mortality, pulmonary artery catheter use in cases of severe preeclampsia or eclampsia was subjectively beneficial in 93 of 100 cases (93%), with an acceptable complication rate (4.0%). PMID- 10871456 TI - Intrauterine infection and preterm delivery: evidence for activation of the fetal hypothalamic-pituitary-adrenal axis. AB - OBJECTIVE: We studied pregnant women in preterm labor with and without intrauterine infection to determine whether fetal hypothalamic-pituitary-adrenal axis activation occurs in the setting of infection-induced preterm parturition. STUDY DESIGN: Amniotic fluid collected by amniocentesis and maternal blood from patients in preterm labor with intact membranes at 24 to 34 weeks' gestation were analyzed by radioimmunoassay for the steroid hormones estrone, estradiol, progesterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone sulfate, and cortisol. Amniotic fluid was also obtained for microbial culture and for interleukin 6 measurements by enzyme immunoassay. RESULTS: Patients with intrauterine infection (n = 11) had significantly higher amniotic fluid concentrations of dehydroepiandrosterone (539 +/- 79 pg/mL) and of cortisol (5.28 +/- 1.0 microg/dL) than did patients with preterm labor and preterm delivery without infection (n = 11; 273 +/- 82 pg/mL and 1.61 +/- 1.05 microg/dL, respectively) or patients with preterm labor and subsequent term delivery (n = 11; 202 +/- 79 pg/mL and 1.82 +/- 1.0 microg/dL, respectively). Furthermore those patients who were delivered within 7 days after enrollment (who were also more likely to have intrauterine infection) had higher amniotic fluid concentrations than did those who were not delivered within 7 days of both estrone (586 +/- 101 pg/mL vs 314 +/- 98 pg/mL) and estradiol (238 +/- 44 pg/mL vs 91 +/- 43 pg/mL). CONCLUSION: Intrauterine infection was associated with increased fetal adrenal androgen and cortisol biosynthesis, and delivery within 7 days after the onset of preterm labor was associated with increased placental estrogen synthesis. These data are consistent with fetal hypothalamic-pituitary-adrenal axis activation in the setting of infection-associated preterm delivery. PMID- 10871457 TI - Hysteroscopic treatment of the uterine septum: a clinician's experience. AB - OBJECTIVE: I reviewed my experience with the diagnosis and hysteroscopic treatment of uterine septa. STUDY DESIGN: This article is a retrospective review of cases from 1992-1999. A septate uterus was diagnosed in a total of 40 patients, and all were treated by hysteroscopic resection. RESULTS: The rate of preoperative pregnancy loss was 77.4%, and the uncomplicated delivery rate was 6.5%. After hysteroscopic septum resection 21 patients reported a total of 22 pregnancies with an 18.2% miscarriage rate and a 77.3% uncomplicated delivery rate. CONCLUSION: Hysteroscopic treatment of uterine septa is a safe, simple, and effective procedure. It can be used for all types of uterine septa, it attains optimal obstetric outcomes, and it should be undertaken whenever a uterine septum is diagnosed. PMID- 10871458 TI - Subcutaneous human menopausal gonadotropin administration for controlled ovarian hyperstimulation with intrauterine insemination cycles. AB - OBJECTIVE: This study was undertaken to determine the feasibility of administering human menopausal gonadotropin subcutaneously for controlled ovarian hyperstimulation with intrauterine insemination. STUDY DESIGN: This was a prospective nonrandomized matched-group comparison. Study patients (n = 25) undergoing controlled ovarian hyperstimulation with intrauterine insemination infertility treatment between June 1998 and March 1999 self-administered human menopausal gonadotropin subcutaneously for ovulation induction. Cycles (n = 39) were analyzed for duration of human menopausal gonadotropin treatment, total number of ampules of human menopausal gonadotropin used, peak serum estradiol level, number of mature follicles (> or =15 mm), cycle fecundity, and acceptability of the subcutaneous route of human menopausal gonadotropin administration. Age-matched historical control subjects who followed the same protocol except for the route of human menopausal gonadotropin administration, which was instead intramuscular, were used for comparison. RESULTS: Study and control cycles did not differ with respect to duration of human menopausal gonadotropin treatment (7.49 vs 8.18 d), total number of ampules of human menopausal gonadotropin used (17.44 vs 19.55), peak serum estradiol level (881 vs 769 pg/mL), number of mature follicles (>/=15 mm; 3.39 vs 2.92), or cycle fecundity rate (15.4% vs 17.9%). Two study patients were switched from subcutaneous to intramuscular administration because of minor local injection site reactions. CONCLUSION: Subcutaneous human menopausal gonadotropin administration for controlled ovarian hyperstimulation with intrauterine insemination treatment cycles was generally well tolerated and yielded stimulation parameters and pregnancy rates similar to those associated with the intramuscular route. Patients subjectively preferred subcutaneous human menopausal gonadotropin administration because of the ability to self-administer the injections, the use of a smaller injection needle, and reduced muscular pain at the injection site. PMID- 10871459 TI - Cefotetan-induced hemolysis associated with antibiotic prophylaxis for cesarean delivery. AB - We describe 3 cases of antibiotic-induced hemolysis associated with cefotetan prophylaxis during cesarean delivery. Each of the 3 patients showed development of significant anemia with documented cefotetan-induced hemolysis. When postpartum anemia is associated with antibiotic use, immune hemolytic anemia should be considered and included in the differential diagnosis. PMID- 10871460 TI - Factors influencing obstetric and gynecologic patients' decisions toward medical student involvement in the outpatient setting. AB - OBJECTIVE: We sought to determine the reasons for obstetric and gynecologic patients' acceptance or refusal of medical student participation in their outpatient care. STUDY DESIGN: A descriptive and analytic cross-sectional study of 180 patients at the University of California, San Francisco, was done to identify factors involved in patient acceptance or refusal of medical student participation in their outpatient obstetric-gynecologic visit. Responses were analyzed by Cochran-Mantel-Haenszel tests for rank order tests of factors involved in the decision to accept or decline medical student participation and chi(2) or Fisher exact tests for comparison of data among different groups. RESULTS: Reasons for accepting medical student involvement included the desire to contribute to the training of future physicians and the desire for the highest standard of care. Reasons for refusing medical student involvement included the protection of patient privacy and the low comfort level with the examination. The acceptance rate for medical students during the obstetric visits was 89.1%, and that during the gynecologic visits was 81.4%. CONCLUSION: Private faculty patients, as well as Medicaid patients, have a high acceptance of both male and female medical students in the obstetric-gynecologic outpatient setting. PMID- 10871461 TI - Longitudinal evaluation of activated protein C resistance among normal pregnancies of Hispanic women. AB - OBJECTIVE: This study was undertaken to describe pregnancy-associated activated protein C resistance and the presence of the factor V Leiden mutation in a sample population of pregnant Hispanic women. STUDY DESIGN: Twenty healthy Hispanic women with single intrauterine pregnancies were randomly selected. Blood samples were taken before 8 weeks' gestation, every 4 weeks during pregnancy, and at 6 weeks post partum. Samples were collected, separated, and stored at -70 degrees C until assay. Standard and modified partial thromboplastin time-based assays were used to evaluate response to activated protein C. A sensitivity ratio < or =2 indicated resistance to activated protein C. Repeated measures analysis of variance and unpaired t tests were used as appropriate. P <.05 was considered significant. RESULTS: Mean (+/-SEM) maternal age was 29 +/- 5 years, and most women were multiparous. Mean gestational age at delivery was 38 weeks' gestation, and the mean birth weight was 3000 g. According to the standard assay, 10 women (50%) acquired activated protein C resistance by 13 weeks' gestation, and this condition persisted through delivery and resolved post partum. Another two had preexisting activated protein C resistance. Results of the standard assay were significantly different for women with preexisting and pregnancy-associated activated protein C resistance (1.55 vs 2.18; P =.01). The modified assay distinguished between women with preexisting and pregnancy-associated activated protein C resistance at 8 weeks' gestation, 24 weeks' gestation, and post partum. The pregnancies of the women with preexisting activated protein C resistance were complicated by oligohydramnios at 34 weeks' gestation and required delivery at 36 weeks' gestation. One infant was small for gestational age. Allele-specific polymerase chain reaction analysis demonstrated that both patients with preexisting activated protein C resistance carried one copy of the factor V Leiden mutation. CONCLUSION: The incidences of pregnancy-associated and factor V Leiden mutation-associated activated protein C resistances in our cohort of gravid Hispanic women was higher than previously reported. Factor V Leiden associated activated protein C resistance in two patients was associated with adverse perinatal outcome. PMID- 10871462 TI - Limited clinical utility of blood and urine cultures in the treatment of acute pyelonephritis during pregnancy. AB - OBJECTIVE: The purpose of this study was to determine the utility of urine and blood cultures in the clinical management of pregnant women with acute pyelonephritis. STUDY DESIGN: Data were pooled from three randomized controlled trials that were conducted at two university-based tertiary care centers and included 391 pregnant women with pyelonephritis. The results of urine and blood cultures were correlated with clinical management decisions, outcome, length of hospital stay, and cost. RESULTS: Results of 98% of urine cultures (382/391) and 99% of blood cultures (388/391) were available for analysis. The most common pathogen isolated was Escherichia coli, which was found in 79% of the urine cultures (300/382) and in 77% of the blood cultures (27/35). Susceptibility testing revealed 46% resistance to ampicillin; 7%, 2%, and 0% resistances to first-, second-, and third-generation cephalosporins, respectively; and 1% resistance to gentamicin. Six percent of the participants (25/391) required changes in antibiotic therapy, most commonly for persistent fever (6/25, 25%). Positive blood culture results directly influenced management by prolonging the duration of hospitalization, with means of 4.6 +/- 2.6 hospital days for women with bacteremia and 2.6 +/- 1.5 hospital days for women without bacteremia (P <.001) despite similar durations of symptoms. CONCLUSION: Urine and blood cultures with sensitivity testing had limited utility in the clinical management of pregnant women with pyelonephritis. Decisions to change antibiotic treatment were affected more by clinical course than by culture results. We suggest that elimination of blood and urine cultures might simplify management and result in significant cost savings without compromising patient care. PMID- 10871463 TI - It was the best of times, it was the worst of times: medicine in the 1990s: presidential address. PMID- 10871464 TI - The legendary superior strength of the Pfannenstiel incision: a myth? AB - OBJECTIVE: This study was undertaken to determine whether there is a difference in the frequency of fascial dehiscence between midline vertical lower abdominal and Pfannenstiel incisions among women undergoing obstetric and gynecologic operations. STUDY DESIGN: A case-control study of 48 cases of fascial dehiscence complicating 17, 995 major operations (8950 cesarean deliveries and 9405 gynecologic procedures) during a 6-year period at Wayne State University Hutzel Hospital, Detroit, was performed. Univariate analysis identified significant independent variables related to fascial dehiscence. Stepwise logistic regression analysis then identified those risk factors that were independently associated with fascial dehiscence. RESULTS: Among the 48 patients who underwent repair of fascial dehiscence after a major obstetric or gynecologic operation, 27 were from the obstetric service and 21 were from the benign and cancer gynecologic services. Wound dehiscence occurred in 10 vertical incisions and 17 Pfannenstiel incisions among the obstetric patients and in 12 vertical and 9 Pfannenstiel incisions among the gynecologic patients. The risk for dehiscence with vertical lower abdominal incisions was not increased with respect to that associated with Pfannenstiel incisions (P =.39, 2-tailed). This finding was true for all patients (odds ratio, 1.3; 95% confidence interval, 0.7-2.6), obstetric patients (odds ratio, 1.3; 95% confidence interval, 0.5-3.4), and gynecologic patients (odds ratio, 1.5; 95% confidence interval, 0.5-4.0). Forty-seven of the 48 case patients had documented wound infections, compared with 1 of the 144 control subjects (P <.0001, odds ratio, 37.8; 95% confidence interval, 14.8-96.8). CONCLUSION: Wound infection was the most important risk factor for fascial dehiscence among women who underwent major obstetric and gynecologic operations. Our results do not support the long-held belief that Pfannenstiel incisions are stronger than lower abdominal vertical incisions and reduce the risk for fascial dehiscence. PMID- 10871465 TI - The relationship between nucleated red blood cell counts and early-onset neonatal seizures. AB - OBJECTIVE: This study was undertaken to better define the timing of neurologic insult in neonates with early-onset seizures through evaluation of neonatal nucleated red blood cell levels. STUDY DESIGN: Medical records and the International Classification of Diseases, Ninth Revision codes were used to identify all term neonates with neonatal convulsions who were delivered at our institution (January 1, 1990-December 31, 1995). Each neonate with early-onset seizures was matched to the next 3 neonates who met the following criteria: gestational age > or =37 weeks, no early-onset seizures, birth weight > or =800 g, umbilical artery pH > or =7.25, and a 5-minute Apgar score >7. Demographic characteristics, clinical factors, and mean initial nucleated red blood cell counts were compared between groups. RESULTS: During the 6-year study period, there were a total of 36, 490 singleton term deliveries of infants who were alive at birth. Forty-five (0.1%) of these neonates had early-onset seizures. Thirty neonates with early-onset seizures met the inclusion criteria. Mean nucleated red blood cell counts (number of nucleated red blood cells per 100 white blood cells) for neonates with early-onset seizures were significantly increased compared with those of control neonates (18.4 +/- 22.0 vs 4.6 +/- 4.5; P <.0008). CONCLUSIONS: Our findings are suggestive of the hypothesis that neurologic injury leading to early-onset seizures often occurs before the intrapartum period. PMID- 10871466 TI - Patients with an ultrasonographic cervical length < or =15 mm have nearly a 50% risk of early spontaneous preterm delivery. AB - OBJECTIVE: The aim of this study was to determine the value in the prediction of spontaneous preterm delivery of ultrasonographically measured cervical length measured between 14 and 24 weeks' gestation. STUDY DESIGN: A retrospective cohort study examined cervical length by means of a two-stage procedure, transabdominal ultrasonography followed by transvaginal ultrasonography if cervical length was <30 mm. RESULTS: A total of 6877 patients met inclusion criteria. Mean cervical length was 37.5 mm. Odds ratios for early preterm delivery (< or =32 weeks' gestation) for patients with cervical lengths < or =10, < or =15, < or = 20, < or =25, and < or =30 mm were, respectively, 29.3 (95% confidence interval, 11.3 75.8), 24.3 (95% confidence interval, 12. 9-45.9), 18.3 (95% confidence interval, 10.8-31.0), 13.4 (95% confidence interval, 8.8-20.6), and 3.2 (95% confidence interval, 2. 4-4.4). For early preterm delivery a cervical length of < or =15 mm had a positive predictive value of 47.6%, a negative predictive value of 96.7%, a sensitivity of 8.2%, and a specificity of 99.7%. CONCLUSIONS: A short cervix seen on a second-trimester sonogram was a powerful predictor of early spontaneous preterm delivery (< or =32 weeks' gestation). Nearly 50% of patients with a cervical length < or =15 mm had an early spontaneous preterm delivery, which suggests that clinical trials of interventions (eg, cerclage) in this population are urgently needed. PMID- 10871467 TI - Programmed cell death (apoptosis) as a possible pathway to metalloproteinase activation and fetal membrane degradation in premature rupture of membranes. AB - OBJECTIVE: Increased matrix metalloproteinase 2 expression and activity are associated with premature rupture of fetal membranes. A proapoptotic protein produced in response to deoxyribonucleic acid fragmentation, p53, can bind to the matrix metalloproteinase 2 gene promoter and cause increased gene expression. It promotes apoptosis by inducing the expression of the proapoptotic bax gene and inhibiting the antiapoptotic bcl-2 gene. This study was undertaken to investigate the expression pattern of apoptotic elements in pregnancy complications that may cause increased expression of the gene for matrix metalloproteinase 2. STUDY DESIGN: Amniochorial membranes were collected from the following groups of women: (1) women with premature rupture of fetal membranes, (2) women with preterm labor and intact membranes, and (3) women with term labor after vaginal delivery. Deoxyribonucleic acid fragmentation was tested with ligation-mediated polymerase chain reaction and the terminal deoxynucleotidyl transferase-mediated biotinylated deoxyribonucleoside triphosphate end-labeling assay. Matrix metalloproteinase 2, p53, bcl-2, and bax gene expression patterns were studied with quantitative competitive polymerase chain reaction. Statistical analysis was performed with the Tukey-Kramer multiple comparison test. RESULTS: Quantitative competitive polymerase chain reaction documented a 10-fold increase in the expression of the gene for matrix metalloproteinase 2 in premature rupture of fetal membranes with respect to term and preterm labor. This induction coincided with an increase in the expressions of the proapoptotic genes p53 and bax and a drop in the expression of the antiapoptotic gene bcl-2. Ligation-mediated polymerase chain reaction revealed deoxyribonucleic acid fragmentation in specimens from premature rupture of fetal membranes and not in those from preterm labor or labor at term. Histochemical analysis documented fragmented deoxyribonucleic acid in chorionic and amniotic cells. CONCLUSION: This study suggests that apoptosis is associated with premature rupture of fetal membranes. Deoxyribonucleic acid fragmentation, associated with elevations in the levels of the two proapoptotic gene products evaluated (p53 and bax ) and a drop in the level of the antiapoptotic bcl-2, was seen in premature rupture of the fetal membranes. Induction of matrix metalloproteinase 2 may be a function of p53 gene expression increase in premature rupture of fetal membranes. PMID- 10871468 TI - A prospective, randomized, controlled trial of high and low maintenance doses of magnesium sulfate for acute tocolysis. AB - OBJECTIVE: This study was undertaken to compare a high-dose protocol for magnesium sulfate tocolytic therapy with a low-dose regimen with respect to time needed to achieve tocolysis. STUDY DESIGN: Patients between 24 and 34 weeks' gestation with preterm labor were included. Patients with ruptured membranes or nonreassuring fetal assessments were excluded. Gravid women received a 4-g loading dose of magnesium sulfate and were prospectively randomly assigned to receive a maintenance dose of 2 or 5 g/h. RESULTS: The median times to tocolysis were 120 minutes (semi-interquartile range, 30 minutes) in the low-dose group and 90 minutes (semi-interquartile range, 28 minutes) in the high-dose group (P <.001). CONCLUSION: Patients treated with a higher maintenance dose of magnesium sulfate had a higher frequency of side effects; however, tocolysis was achieved more rapidly and they required shorter admissions to the labor and delivery unit without increased maternal or neonatal morbidity. PMID- 10871469 TI - Quantification of endometriosis-associated pain and quality of life during the stimulatory phase of gonadotropin-releasing hormone agonist therapy: a double blind, randomized, placebo-controlled trial. AB - OBJECTIVE: The purpose of this study was a quantification of changes in endometriosis-associated pain and quality of life during the stimulatory phase of gonadotropin-releasing hormone agonist therapy. STUDY DESIGN: One hundred twenty women with significant endometriosis-associated pain participated in a 1-month double-blind, randomized, placebo-controlled trial. Pain was measured at baseline and at 2 and 4 weeks with visual analog scales and the Endometriosis Symptom Severity score. Quality of life was measured with the SF-36 instrument. Group means and SEMs were calculated. Paired t tests were used after determination of data normality. RESULTS: Compared with placebo-treated control subjects women treated with gonadotropin-releasing hormone agonist had a statistically (P <. 0001) and clinically significant temporary increase in pain and a concomitant decrease in quality of life. CONCLUSION: The stimulatory phase of gonadotropin releasing hormone agonist therapy is associated with an increase in endometriosis associated pain and a decrease in quality of life. PMID- 10871470 TI - Correctly identifying the macrosomic fetus: improving ultrasonography-based prediction. AB - OBJECTIVE: Our goal was to improve the accuracy of estimating fetal weights among macrosomic fetuses with the traditional measurements of abdominal circumference, femur length, and head circumference. STUDY DESIGN: We used 4831 cases without anomalies from an ultrasonography laboratory database with an estimated fetal weight obtained a maximum of 14 days before delivery. Abdominal circumference, femur length, and head circumference were each regressed on birth weight to obtain estimated fetal weight by abdominal circumference, femur length, and head circumference, respectively. We compared the individual variation for estimated fetal weight by abdominal circumference, femur length, and head circumference by calculating a within-subject standard deviation to quantify the level of disparity. We adjusted the estimated fetal weight to the date of delivery and for dependencies on maternal diabetes mellitus, weight, and height. We then weighted cases with birth weight >4500 g and diabetic cases with birth weight >4000 g 20 fold (weighted estimated fetal weight) for the purpose of creating a favorable bias for classifying these cases. The equation of Hadlock et al, with abdominal circumference, femur length, and head circumference, was applied as a benchmark estimated fetal weight. RESULTS: Of the 4831 newborns, 308 (6.4%) had a birth weight >4000 g, and 56 (1.2%) had a birth weight >4500 g. There were 154 pregnancies complicated by diabetes mellitus; 26 (16.9%) of the resulting infants weighed >4000 g, and 5 (3.2%) weighed >4500 g. At 95% specificity, the weighted estimated fetal weight had a sensitivity of 85.7% at a cut point of 3912 g, compared with a sensitivity of 71.4% at 3604 g by use of the estimated fetal weight of Hadlock et al. CONCLUSIONS: We were able to improve the accuracy of identifying the macrosomic fetus compared to reliance on the equation by Hadlock et al. A fetus was found to be at significantly increased risk for birth weight >4000 g when the estimated fetal weight based on abdominal circumference is larger than that based on either head circumference or femur length or when there is a large within-subject variance in estimated fetal weight based on abdominal circumference, femur length, and head circumference. We also found that there were significantly different groups of patients whose estimated fetal weights require different equations for better estimates. Even given ultrasonographic measurements, taking into account maternal height, weight, and presence of diabetes mellitus can improve macrosomia detection. Although these findings remain to be optimized and validated, the approach used here appears to yield better predictions than the current "one function fits all" approach. PMID- 10871472 TI - Panniculectomy at the time of gynecologic surgery in morbidly obese patients. AB - OBJECTIVE: Our goal was to demonstrate that panniculectomy performed at the time of gynecologic surgery aids in reducing the operative time and exposure and does not increase the wound infection rate in morbidly obese patients. STUDY DESIGN: A retrospective survey was performed of massively obese patients who underwent panniculectomy at the time of gynecologic surgery at Northeastern Ohio Universities College of Medicine consortium hospitals from 1990-1999. Data collected during surgery included the patient's weight, operative opening and closing times, blood loss, and weight of the removed panniculus adiposus. Postoperative wound infection rates were monitored, and patients were followed up for 6 months. RESULTS: Seventy-eight patients underwent the following operations: radical hysterectomy (n = 19), extrafascial hysterectomy (n = 18), standard hysterectomy (n = 32), or other gynecologic surgery (n = 9). The average blood loss was 71 mL. Opening and closing times were 27 and 33 minutes, respectively, adding a minimal amount of operative time to the required gynecologic surgery. The average removed panniculus adiposus weighed 4745 g. Efficiency in obtaining exposure to the operative site was noted. A total of 2 wound infections were recorded in the postoperative period. In 1 case debridement was required, and in the other healing occurred by secondary intention. Minimal separation occurred in 4 other cases and required no intervention. CONCLUSION: Massively obese patients can safely undergo panniculectomy simultaneously with a gynecologic procedure. The difficulty with operative exposure is reduced, and these patients are better served intraoperatively. Postoperatively, the wound infection rates quoted for this population were markedly improved from prior studies and involved a larger group of patients. PMID- 10871471 TI - Intrauterine endotoxin infusion in rat pregnancy induces preterm delivery and increases placental prostaglandin F2alpha metabolite levels. AB - OBJECTIVE: This study was designed to examine the effects of intrauterine endotoxin (lipopolysaccharide) on rat pregnancy. STUDY DESIGN: Pregnant Sprague Dawley rats (N = 26) were implanted with uterine catheters on day 15 or 16 of a 22-day gestation. Animals were randomly assigned to receive either lipopolysaccharide (25 or 50 microg) or sodium chloride solution (1 mL) on day 17 and then were either sacrificed on day 19 or observed until delivery. Placentas were harvested at the time of death, homogenates were prepared, and prostaglandin F(2)(alpha) metabolite levels were determined by means of radioimmunoassay. Data were analyzed by analysis of variance, Student-Newman-Keuls, and Mann-Whitney tests. RESULTS: Lipopolysaccharide-treated groups (25 and 50 microg) displayed a shorter interval to delivery (mean +/- SE, 82 +/- 13 and 63 +/- 8 hours, respectively) than control animals (117 +/- 3 hours). Pups of lipopolysaccharide treated (25 and 50 microg) female animals had lower live birth weights (4.92 +/- 0.01 and 5.12 +/- 0. 24 g, respectively) compared with control animals (6.04 +/- 0.07 g). Placental homogenates from lipopolysaccharide-treated female animals contained higher levels of prostaglandin F(2)(alpha) metabolite (1567 +/- 64 and 1475 +/- 59 pg/mL) than those from sodium chloride solution-infused control animals (804 +/- 68 pg/mL). CONCLUSION: Bacterial products induce the preterm delivery of low-birth-weight pups in rats, possibly by increasing local prostaglandin biosynthesis. PMID- 10871473 TI - Low-risk corpus cancer: is lymphadenectomy or radiotherapy necessary? AB - OBJECTIVE: The objective of this study was to find readily ascertainable intraoperative pathologic indicators that would discriminate a subgroup of early corpus cancers that would not require lymphadenectomy or adjuvant radiotherapy. STUDY DESIGN: Between 1984 and 1993, a total of 328 patients with endometrioid corpus cancer, grade 1 or 2 tumor, myometrial invasion < or =50%, and no intraoperative evidence of macroscopic extrauterine spread were treated surgically. Pelvic lymphadenectomy was performed in 187 cases (57%), and nodes were positive in nine cases (5%). Adjuvant radiotherapy was administered to 65 patients (20%). Median follow-up was 88 months. RESULTS: The 5-year overall cancer-related and recurrence-free survivals were 97% and 96%, respectively. Primary tumor diameter and lymphatic or vascular invasion significantly affected longevity. No patient with tumor diameter < or =2 cm had positive lymph nodes or died of disease. CONCLUSION: Patients who have International Federation of Gynecology and Obstetrics grade 1 or 2 endometrioid corpus cancer with greatest surface dimension < or =2 cm, myometrial invasion < or =50%, and no intraoperative evidence of macroscopic disease can be treated optimally with hysterectomy only. PMID- 10871474 TI - Lack of utility of standard labor curves in the prediction of progression during labor induction. AB - OBJECTIVE: This study was undertaken to determine whether patients undergoing labor induction can be reliably evaluated by means of standard labor assessment curves. STUDY DESIGN: In this retrospective chart review of 123 patients who underwent cervical ripening and induction of labor, Friedman's standard labor curves were used for comparison. Statistical analysis was performed with the Student t test. RESULTS: Nulliparous and parous patients undergoing cervical ripening spent more time in active-phase labor than standard expectations of labor progression would indicate (12.7 +/- 7.8 vs 5. 9 +/- 3.4 hours for nulliparous women, P <.001; 7.9 +/- 6.4 vs 2.5 +/- 1.5 hours for parous women, P <.001). Nulliparous and parous patients who were delivered vaginally spent more time in active labor than did their respective standard historical control populations (10.3 +/- 8.0 vs 5.9 +/- 3.4 hours for nulliparous women, P <.001; 7.0 +/- 6.0 vs 2.5 +/- 1.5 hours for parous women, P <. 001). CONCLUSION: Standard methods for the evaluation of labor adequacy and prediction of the likelihood of vaginal delivery may not apply to patients undergoing cervical ripening. PMID- 10871475 TI - Is vaginal delivery preferable to elective cesarean delivery in fetuses with a known ventral wall defect? AB - OBJECTIVE: We sought to test the hypothesis that vaginal delivery compared with elective cesarean delivery results in improved neonatal outcome in fetuses with a known isolated ventral wall defect. STUDY DESIGN: We performed a retrospective chart review. RESULTS: Between 1989 and 1999, we identified 102 infants with a confirmed antenatal diagnosis of an isolated ventral wall defect with either the diagnosis of an omphalocele or gastroschisis. Sixty-six infants were delivered by cesarean and 36 were delivered vaginally. There were no significant demographic differences between the study groups or between the two sites except that one center (Cincinnati) usually delivered these fetuses by cesarean whereas the other (Louisville) usually delivered such fetuses vaginally. Overall, there were a greater number of infants with gastroschisis than omphalocele (gastroschisis, n = 71; omphalocele, n = 31). After we controlled for primary versus staged closure of ventral wall defect and gestational age at delivery; the medians and interquartile ranges for cesarean and vaginal delivery were 39 (25, 63) days versus 42 (26, 75) days, respectively (P =.32), for neonatal length of stay and 13 (9, 18) days versus 13 (9, 26) days, respectively (P =.16), for days to enteral feeding. After we controlled for the size of the defect and the amount of bowel resected, the odds of primary closure given a vaginal delivery was about half that given a cesarean delivery (odds ratio, 0.56; 95% confidence interval, 0.18-1. 69), but this was not statistically significant. There was no statistically significant difference in the rates of neonatal death (2 [3%] vs 2 [6%]; P =.61) and neonatal sepsis (2 [3%] vs 4 [11%]; P =.18) for cesarean versus vaginal delivery. Maternal length of stay after delivery was found to be 1 day less after vaginal delivery [vaginal, 2 (2, 2) days; cesarean, 3 (2, 3) days; P =.0001]. There were 5 instances of maternal complications, and all 5 pregnancies were delivered by cesarean (P =.16). CONCLUSION: Fetuses with an antenatal diagnosis of an isolated ventral wall defect may safely be delivered vaginally, and cesarean delivery should be performed for obstetric indications only. PMID- 10871476 TI - Pretreatment assessment of prognostic indicators in endometrial cancer. AB - OBJECTIVE: The object of this study was to assess the association of histologic, cytokinetic, and molecular variables in preoperative endometrial samples with extrauterine disease, recurrence, and survival among patients with endometrial cancer. STUDY DESIGN: In a case-cohort study of 125 women, ploidy, S-phase fraction, proliferative index, deoxyribonucleic acid index, proliferating cell nuclear antigen, MIB-1 proliferation marker, p53 tumor suppressor gene, and cytoplasmic HER-2/neu oncogene and bcl-2 expressions were quantitated. RESULTS: A model with only one independent term predicted progression-free survival; that variable was p53 (P <. 0001; relative risk, 5.60). A model with two independent terms predicted disease-related survival; these variables were p53 (P =. 0002; relative risk, 7.39) and MIB-1 (P =.03; relative risk, 3.27). Among patients with tumors with both p53 and MIB-1 expression exceeding 33%, a total of 32% had died of disease by 2 years. A model for predicting extrauterine disease selected two independent variables: p53 (odds ratio, 3.20; P =.01) and ploidy (odds ratio, 2. 16; P =.04). An advanced surgical stage was encountered in 26% to 35% of cases in which either the p53 expression exceeded 33% or the deoxyribonucleic acid content was nondiploid and in 53% of cases in which both variables were unfavorable. CONCLUSIONS: Preoperative evaluation of quantifiable cytokinetic and molecular variables can assist in identifying tumor types that are predisposed toward a more aggressive clinical course. PMID- 10871477 TI - Matrilysin (matrix metalloproteinase 7) in parturition, premature rupture of membranes, and intrauterine infection. AB - OBJECTIVE: Matrix metalloproteinases are enzymes capable of degrading extracellular matrix components. Matrilysin (matrix metalloproteinase 7), a novel member of this family, degrades fibronectin and proteoglycans. The objective of this study was to determine whether parturition (either term or preterm), premature rupture of the membranes, and microbial invasion of the amniotic cavity are associated with changes in the amniotic fluid concentration of matrilysin. STUDY DESIGN: A cross-sectional study was conducted with 275 women in the following categories: (1) second trimester, (2) term not in labor, (3) term in labor, (4) term with microbial invasion of the amniotic cavity, (5) preterm labor with intact membranes without microbial invasion of the amniotic cavity who delivered at term, (6) preterm labor without microbial invasion of the amniotic cavity who delivered preterm, (7) preterm labor with microbial invasion of the amniotic cavity, (8) preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, and (9) term premature rupture of membranes not in labor and without microbial invasion of the amniotic cavity. Matrilysin concentrations were measured with a sensitive specific immunoassay that was validated for amniotic fluid. RESULTS: Matrilysin was detectable in 97.4% (268/275) of the samples. The concentration of matrilysin increased with advancing gestational age (r = 0.8; P <.001). Parturition at term was not associated with a significant increase in amniotic fluid concentration of matrilysin. Preterm parturition in the absence of microbial invasion of the amniotic cavity was associated with a significant increase in amniotic fluid concentration of matrilysin (preterm labor with preterm delivery: median, 1.7 ng/mL; range, 0.45-21.6 mg/mL; vs preterm labor with term delivery: median, 1.2 ng/mL; range, 0.17-42. 1 ng/mL; P <.05). Premature rupture of membranes without microbial invasion of the amniotic cavity (either term or preterm) was not associated with a significant change in the amniotic fluid matrilysin concentration. Intra-amniotic infection was associated with a significant increase in amniotic fluid matrilysin among both patients with preterm labor and patients with preterm premature rupture of membranes (preterm labor with microbial invasion of the amniotic cavity: median, 3.2 ng/mL; range, 0.16-21.9 ng/mL; vs preterm labor and delivery without microbial invasion of the amniotic cavity: median, 1.7 ng/mL; range, 0.45-21.6 ng/mL; vs preterm labor with term delivery: median, 1.2 ng/mL; range, 0.17-42. 1 ng/mL; P <.01 for each comparison; and preterm premature rupture of membranes without microbial invasion of the amniotic cavity: median, 1.7 ng/mL; range, 0.29-13.9 ng/mL; vs preterm premature rupture of membranes with microbial invasion of the amniotic cavity: median, 3.6 ng/mL; range, 0.59-20.3 ng/mL; P <.01). CONCLUSION: Matrilysin is a physiologic constituent of amniotic fluid, and its concentration increases with advancing gestational age. Microbial invasion of the amniotic cavity in preterm gestations was associated with a significant increase in amniotic fluid concentration of matrilysin. Matrilysin therefore may play a role in the host defense mechanism. PMID- 10871478 TI - A comprehensive program to improve safety for pregnant women and fetuses in motor vehicle crashes: a preliminary report. AB - OBJECTIVE: A program was developed to study the mechanisms of abruptio placentae and pregnancy loss caused by motor vehicle crashes. The results were intended to be used to develop strategies to improve protection of the fetus in this setting. STUDY DESIGN: Four integrated projects were conducted: (1) seated anthropometric measurements and belt fit determination during pregnancy, (2) development of new models of traumatic abruptio placentae, (3) investigations of crashes involving pregnant women, and (4) the development of the second-generation pregnant crash dummy from these data and others. RESULTS: Twenty-two different pregnant subjects in five different height groups underwent serial measurements of abdominal surface contours, seat belt fit, and distances between the subjects and various landmarks in the automobile interior with a laboratory-designed "automobile seat" (seating buck). The abdomen was significantly closer to the steering wheel in the shorter stature group than among the taller women. Beginning at approximately 20 weeks' gestation the fundus of the uterus was above the lower rim of the steering wheel. Lap belts fit properly over the anterior superior iliac spine throughout gestation, but the lap belt overlapped the uterus in the midsagittal plane. Two separate mechanisms for traumatic abruptio placentae were tested: shear failure and tensile failure. In the shear failure model large circumferential strains in the uterine wall induce a shear strain across the uteroplacental interface, and the model predicts placental separation at a mean circumferential strain of -58% +/- 8%. By means of finite-element modeling, it was demonstrated that tensile failure might also be a mechanism that causes abruptio placentae during rapid deceleration of the uterus. Crash investigations were performed in 43 cases involving pregnant women beyond 20 weeks' gestation. There were a total of 8 fetal losses and 8 major complications (fetal survival with abruptio placentae, direct fetal injury, or preterm delivery before 34 weeks' as a result of the accident). The best predictors of fetal loss or adverse outcome were impact severity and proper seat belt use. With these newly acquired data a second generation crash dummy, known as the Maternal Anthropomorphic Measurement Apparatus version 2b (MAMA-2b), is being developed. It incorporates strain gauges in the fundal region of the fluid-filled uterus plus pressure transducers in both the anterior and posterior uterus. Criteria are being developed to associate the likelihood of abruptio placentae with measurements from these instruments that correspond to the two major hypothesized mechanisms of abruptio placentae. CONCLUSION: An improved understanding of the elements of automobile crashes that cause fetal loss and other major pregnancy complications has been gained through this series of investigations. PMID- 10871479 TI - Urethral collagen morphologic characteristics among women with genuine stress incontinence. AB - OBJECTIVE: This was a study of the morphologic characteristics of urethral collagen in women with stress incontinence and continent control women. STUDY DESIGN: Urethral needle biopsy specimens were obtained from 31 women. Fifteen women were continent, and the other 16 had undergone full urogynecologic assessment for symptoms of urinary incontinence. Biopsy specimens were assessed under electron microscopy. Mean collagen fibril diameter was measured and collagen morphologic characteristics were assessed. RESULTS: The biopsy specimens from 30 women were included in the analysis. Collagen fibril diameter did not vary with continence status, the presence of pelvic organ prolapse, age, race or hormonal status. Alterations in collagen fibril morphologic characteristics were evident in the biopsy specimens from nine patients with incontinence. The alterations in collagen morphologic characteristics fell into three patterns, which for convenience were referred to as the obscured pattern, the dense pattern, and the degenerative pattern. CONCLUSION: Altered collagen morphologic characteristics are found in some patients with stress incontinence, and possible causes for those alterations are suggested by their appearances. PMID- 10871480 TI - Multifetal pregnancy reduction: perinatal and fiscal outcomes. AB - OBJECTIVE: This study was undertaken to compare the birth outcomes of a multifetal pregnancy reduction population with those of other patients delivered at Hutzel Hospital, Detroit, and to determine the fiscal impact of the multifetal pregnancy reduction program. STUDY DESIGN: In a retrospective review patients who were delivered after multifetal pregnancy reduction were compared with a general obstetric population who were delivered at Hutzel Hospital from January 1, 1986, through June 30, 1998. Outcome data were determined through a comprehensive perinatal database. The chi(2) analysis was used to examine the relationship between gestational age and delivery group. Financial data were estimated from published reports of neonatal intensive care unit admissions, cost estimates for neonatal intensive care unit care, and charges for multifetal pregnancy reduction. RESULTS: Pregnancies reduced to triplets, twins, and singletons had outcomes at least comparable to unreduced pregnancies starting at these numbers and substantially better than unreduced pregnancies with the same starting number. Financial estimates of hospitalization costs averted in the multifetal pregnancy reduction population exceeded $28 million. CONCLUSION: Use of multifetal pregnancy reduction improved obstetric outcomes for pregnancies with multiple gestations and also was associated with significant fiscal savings. PMID- 10871481 TI - The amniotic fluid index, single deepest pocket, and two-diameter pocket in normal human pregnancy. AB - OBJECTIVE: This study was undertaken to determine normative values for amniotic fluid index, single deepest pocket, and 2-diameter pocket across gestation. STUDY DESIGN: Fifty patients with normal pregnancies at each gestational age between 14 and 41 weeks' gestation were recruited prospectively and scanned once. Data were transformed into logarithmic (base 10) values for analysis. Polynomial regression equations were used to predict the normal values for amniotic fluid index, single deepest pocket, and 2-diameter pocket across gestational age and to predict the weekly percentage changes. RESULTS: The mean amniotic fluid index, single deepest pocket, and 2-diameter pocket values were significantly lower among patients at <37 weeks' gestation (n = 1150) than among those at > or =37 weeks' gestation (n = 250; P <.001 for all comparisons). The calculated prevalences of oligohydramnios (amniotic fluid index < or =5 cm, single deepest pocket <2 cm, or 2-diameter pocket <15 cm(2)) were significantly different (P <.0001) for the three techniques (8%, 1%, and 30%, respectively). Hydramnios (amniotic fluid index >24 cm, single deepest pocket >8 cm, or 2-diameter pocket >50 cm(2)) was also diagnosed with significantly different (P <.0001) frequencies (0%, 0.7%, and 3%, respectively). CONCLUSIONS: This is the largest prospective study to date to provide normative data for each of three ultrasonographic techniques used to assess amniotic fluid volume. The single deepest pocket appears to be the preferable method, because its use is least likely to lead to the false-positive diagnosis of either oligohydramnios or hydramnios. PMID- 10871482 TI - Predicting preterm birth: a cost-effectiveness analysis. AB - OBJECTIVE: The objective of this study was to compare the cost-effectiveness of 9 strategies for the management of threatened preterm labor. STUDY DESIGN: We derived 6 management options from the literature. These were (1) to treat all women with tocolytics and corticosteroids ("treat all"); (2) to treat all women while awaiting results of the "traditional" fetal fibronectin test results, then discontinue treatment on those with negative results; (3) to treat only those with abnormal cervical length measurements as detected by ultrasonography; (4) to treat only those with abnormal "rapid" fetal fibronectin test results; (5) to perform rapid fetal fibronectin testing and cervical length measurements and treat those with a positive result on either or both; (6) not to treat any women ("treat none"). To assess the contributions of tocolytics and corticosteroids to our outcomes, we analyzed 3 additional treatment options: (7) to treat all women with outpatient corticosteroids but not give tocolytics, (8) to administer corticosteroids to all but give tocolytics only to those with abnormal rapid fetal fibronectin test results, and (9) to administer corticosteroids to all but give tocolytics only to those with abnormal cervical length. We used decision analytic techniques to perform a cost-effectiveness analysis. RESULTS: A decision tree was constructed on the basis of these strategies. We reviewed the literature to derive all probability information. We derived sensitivity and specificity for delivery <37 weeks for fetal fibronectin and for abnormal cervical length. Outcomes of interest were respiratory distress syndrome and neonatal death. We derived cost variables from institutional statistics and from values quoted in the literature. Total costs, cases of respiratory distress syndrome, neonatal deaths, and cost-effectiveness ratios were calculated for each of the strategies. We conducted sensitivity analyses on all variables. Universal administration of outpatient corticosteroids was the least expensive option, but it resulted in more cases of respiratory distress syndrome and deaths than "treat all." Rapid fetal fibronectin plus corticosteroids, traditional fetal fibronectin, and cervical length plus corticosteroids were the next least expensive options and resulted in numbers of cases of respiratory distress syndrome and deaths that were similar to those in the "treat all" strategy. The "rapid" fetal fibronectin test, cervical length measurement, rapid fetal fibronectin test plus cervical length measurement, and "treat none" strategies resulted in more respiratory distress syndrome, more deaths, and higher costs. Treating all patients resulted in the fewest number of cases of respiratory distress syndrome and deaths but the greatest costs. CONCLUSION: Risk prediction strategies with the fetal fibronectin assay or corticosteroids plus rapid fetal fibronectin testing or cervical length assessment may offer cost savings compared with treatment of all women with threatened preterm labor and may prevent similar numbers of cases of respiratory distress syndrome and neonatal deaths. PMID- 10871483 TI - Haplogroup-associated differences in neonatal death and incidence of low birth weight at elevation: a preliminary assessment. AB - OBJECTIVE: We sought to assess reproductive fitness differences between mitochondrial deoxyribonucleic acid haplogroups at high altitude. STUDY DESIGN: This study considers differences in outcomes of conception, birth weight, and neonatal mortality rates for 62 women classified according to haplogroups (B or non-B). RESULTS: The number of low-weight births (<2500 g) for the non-B group was significant (P =.019). Mothers in the non-B group reported more spontaneous abortions (P =.171) and stillbirths (P =.301). The difference in conceptions per woman between groups was significant (P =.036). However, no difference in infants alive at 1 month of age was evident. Neonatal death was significant (P =.017). The odds of an unsuccessful outcome among mothers in the B group was compared with mothers in the non-B group and was significant (P =.029). The chance of an adverse outcome, that is, fetal or infant death before 1 month, for mothers in the B group was between 11.1% and 88.7% lower than for mothers in the non-B group. CONCLUSIONS: The neonatal mortality rate for the non-B group was significantly elevated relative to the B group. The molecular basis for these observations is not clear. PMID- 10871484 TI - Determination of amniotic fluid volume in twin pregnancies: ultrasonographic evaluation versus operator estimation. AB - OBJECTIVE: We sought to determine the accuracy of amniotic fluid volume estimation (visually) in diamniotic twin pregnancy versus ultrasonography techniques. STUDY DESIGN: In this prospective study the volume of each sac in 23 sets of diamniotic twin pregnancies was subjectively and objectively estimated by a second-year obstetric resident, nurse sonographer, maternal-fetal medicine fellow, and maternal-fetal medicine staff. The actual volume was confirmed by amniocentesis and a dye-dilution technique. RESULTS: There was no difference in the total number of correct estimates of volume by level of operator experience (P =.98), ultrasonography technique (P =.87), or combined subjective versus objective correct estimates (P =.87). Identification of low volume was not different among the four evaluators (P =.48), but the percentage of correct estimates was poor (7%-29%). The 2-diameter pocket was a better predictor of oligohydramnios (57%) than the amniotic fluid index or the largest vertical pocket (12.5%; P =.002). CONCLUSION: The extremes of volume (low or high) are poorly identified by the subjective or objective assessment of volume. PMID- 10871485 TI - Sexual function and vaginal anatomy in women before and after surgery for pelvic organ prolapse and urinary incontinence. AB - OBJECTIVE: We sought to describe sexual function in women before and after surgery for either prolapse or urinary incontinence, or both. STUDY DESIGN: Women completed questionnaires, and vaginal dimensions were measured before and at least 6 months after surgery for prolapse or incontinence. Comparisons were made with signed-rank tests or the McNemar test. RESULTS: Eighty-one (49%) of 165 women were sexually active before and after surgery; their mean age was 54. 0 +/- 9.9 years. Mean frequency of intercourse did not change. Dyspareunia was reported by 6 (8%) women preoperatively and 15 (19%) women after surgery; dyspareunia persisted postoperatively in 1 woman, developed in 14, and resolved in 5 (P =.04). Dyspareunia occurred in 14 (26%) of 53 women after posterior colporrhaphy (P =. 01) and in 8 (38%) of 21 women who had Burch colposusupension and posterior colporrhaphy performed together (P =.02). Vaginal dimensions decreased slightly after surgery; however, this did not correlate with any change in sexual function. Preoperatively, 66 (82%) women were satisfied with their sexual relationships, compared with 71 (89%) who were satisfied postoperatively. CONCLUSION: Sexual function and satisfaction improved or did not change in most women after surgery for either prolapse or urinary incontinence, or both. However, the combination of Burch colposusupension and posterior colporrhaphy was especially likely to result in dyspareunia. PMID- 10871486 TI - Effect of vaginal pH on efficacy of misoprostol for cervical ripening and labor induction. AB - OBJECTIVE: We sought to evaluate whether vaginal pH has an effect on the relative efficacy of misoprostol for cervical ripening and labor induction. STUDY DESIGN: Thirty-seven gravid women with an unfavorable cervix and indication for labor induction were enrolled in this prospective, double-blind, observational study. Baseline assessments of cervicovaginal pH and Bishop score were made at the time of enrollment by an independent examiner. All patients received 50 microg misoprostol intravaginally every 6 hours for 12 hours. After the initial 12 hours of preinduction, a repeat Bishop score assessment was made by the same initial examiner. Patients not in active labor at 12 hours were placed on a standardized oxytocin induction regimen. Labor was managed by the on-call obstetric team, who remained blinded to pH assessment. Clinical outcomes were evaluated. Statistical analyses were made by the Student t test, the Fisher exact test, and linear regression analysis. RESULTS: Average initial vaginal pH was 4.8 +/- 0.5 (range, 3.5-7.0) for the study cohort. No significant differences were noted between those patients with low vaginal pH (< or =4.5) compared with those with high pH vaginal (>4.5) with respect to maternal age, parity, gestational age, or initial Bishop score. Similarly, Bishop score change over preinduction interval (5.6 vs 4.9), time to active labor (16.3 vs 17. 1 hours), time to complete dilatation (20.0 vs 19.9 hours), and time to delivery (21.0 vs 21.6 hours) were not significantly different between the low and high pH groups, respectively. Linear regression analysis revealed no significant association between vaginal pH and Bishop score change during preinduction interval, time to active labor, time to complete dilatation, or time to delivery. CONCLUSION: Vaginal pH does not appear to influence the efficacy of intravaginally administered misoprostol for cervical ripening and labor induction. PMID- 10871487 TI - Benefits associated with harmonic tissue imaging in the obstetric patient. AB - OBJECTIVE: We sought to determine the impact of harmonic tissue imaging on image resolution and visualization of fetal structures during obstetric ultrasonography. STUDY DESIGN: Patients with singleton second- or third-trimester fetuses were recruited. Prospective comparisons of conventional fundamental imaging and harmonic tissue imaging were made. Visualization rates and frequencies of improvement in resolution were calculated. Discriminate function analysis evaluated determinants of improved visualization. RESULTS: Harmonic tissue imaging improved resolution of at least one fetal structure in 51.4% of patients studied. Differences were most marked for 4-chamber views of the heart with improvement in resolution in 30.5% of patients and change in ability to visualize in 9.5%. Maternal weight and gestational age had a significant influence on whether improvements were noted with harmonic tissue imaging, accounting for 27% of the variance. CONCLUSIONS: Harmonic tissue imaging offers significant improvements over fundamental imaging in image resolution and structure visualization in obese patients during the second trimester of pregnancy. PMID- 10871488 TI - Minimal precycle testing and ongoing cycle monitoring for in vitro fertilization and fresh pre-embryo transfer do not compromise fertilization, implantation, or ongoing pregnancy rates. AB - OBJECTIVE: We sought to assess the fertilization, implantation, and ongoing pregnancy rates with a minimal precycle and ongoing cycle monitoring protocol for in vitro fertilization and embryo transfer. STUDY DESIGN: Retrospective review was conducted of 103 consecutive cycles of fresh in vitro fertilization and embryo transfer from 1996 to 1998. Precycle screening included semen analysis without strict morphologic analysis, and hysterosalpingography-sonohysterography within the last year. Serum prolactin, serum thyroid-stimulating hormone, reactive plasma reagin, human immunodeficiency virus, rubella titer, blood type, hepatitis B surface antigen, and hepatitis C antibody testing was performed on all patients within 3 months of cycle initiation. Women > or =37 years old underwent clomiphene challenge testing. The monitoring protocol included the following: baseline transvaginal ultrasonography after 12 to 14 days of midluteal gonadotropin-releasing hormone agonist down-regulation to assess endometrial thickness and adnexal appearance, transvaginal ultrasonography for follicle evaluation at 7 and 10 days, serum estradiol assay if > or =20 follicles, quantitative beta-human chorionic gonadotropin 12 to 14 days after pre-embryo transfer, repeat quantitative beta-human chorionic gonadotropin 3 to 5 days later, and transvaginal ultrasonography for intrauterine gestational sac confirmation 7 to 9 days after the initial positive pregnancy test result. The dose of gonadotropin used remained constant unless the sonogram obtained on day 7 indicated a suboptimal response (<3 follicles each, with an average diameter of 10 to 12 mm) or hyperresponse (> or =15 follicles with an average diameter of 10 to 12 mm). RESULTS: The per embryo implantation rate (fetal cardiac activity) was 13.1%, and the live birth rate per 100 pre-embryo transfers was 31.5 for patients < or =40 years old. The average number of pre-embryos transferred was 3.1. The singleton pregnancy rate was 71%, and there were no multiple gestations greater than twins. The mean number of oocytes fertilized was 66%. There was 1 case of failed fertilization with intracytoplasmic sperm injection. There were two other cases of failed fertilization. One case of severe ovarian hyperstimulation occurred in spite of cryopreservation of all embryos. CONCLUSIONS: In vitro fertilization and embryo transfer can be accomplished with minimal precycle testing and ongoing cycle monitoring without compromising fertilization, implantation, and ongoing pregnancy rates. This results in reduced overall costs for couples. PMID- 10871489 TI - Obstetric outcomes in 102 pregnancies after preimplantation genetic diagnosis. AB - OBJECTIVE: We sought to determine whether preimplantation genetic diagnosis is associated with particular pregnancy or delivery complications. STUDY DESIGN: A total of 102 consecutive pregnancies after preimplantation genetic diagnosis by polar body removal performed at Illinois Masonic Medical Center resulting in 114 live births were analyzed. All patients were given a delivery and newborn questionnaire, and attempts were made to contact and question them regarding any pregnancy complications and type of delivery. Permission was obtained to examine medical records and discuss the patient's pregnancy with her obstetrician when questions existed with respect to complications or indication for cesarean delivery. RESULTS: Delivery and newborn questionnaires were completed or telephone contact was achieved for 100 of the 102 pregnancies. There were 85 singleton, 9 twin, and 7 triplet pregnancies. Of the 7 triplet gestations, 3 couples elected multifetal pregnancy reduction to twins and healthy triplets were born to 4 couples between 32 and 36 weeks by cesarean delivery. Of the 80 singleton deliveries, 60 (75%) progressed to term. Of these 60 term singleton deliveries, 34 were vaginal, 23 were cesarean (40%), and 3 delivery types were unknown. The incidence of small-for-gestational-age infants was 3% for neonates in the 60 term singleton deliveries and 7% in the entire cohort of 80 singleton deliveries. Only 3 pregnancy complications (other than premature delivery) were reported more than once. There were 3 instances each of gestational diabetes, intrauterine growth restriction, and pregnancy-induced hypertension. There was 1 case each of HELLP (hemolysis, elevated liver enzymes, and low platelets) syndrome, congestive heart failure, mild oligohydramnios, and abruptio placentae. The indications for cesarean delivery were (in descending order) failure of labor to progress (n = 7), fetal distress (n = 4), placenta previa (n = 4), elective repeat cesarean delivery (n = 4), triplets (n = 3), uterine scarring (n = 3), 1 twin in the breech position (n = 3), failed forceps delivery (n = 2), and a variety of other indications that occurred in only 1 patient each. All preimplantation genetic diagnoses were confirmed by prenatal or postnatal testing. No diagnostic errors were made in this cohort of patients or in any patients undergoing preimplantation genetic diagnosis having polar body removal in our center. CONCLUSIONS: Preimplantation genetic diagnosis is associated with a risk of multiple gestations, cesarean delivery, and placenta previa. Cesarean delivery rates and multiple gestation rates are comparable to those of patients undergoing in vitro fertilization in general. The preimplantation genetic diagnosis itself does not seem to cause an increased risk for any particular pregnancy complication, with the possible exception of placenta previa, which was seen in 4% of patients. PMID- 10871490 TI - A study of the relationship between placenta growth factor and gestational age, parturition, rupture of membranes, and intrauterine infection. AB - OBJECTIVE: Placenta growth factor is a potent angiogenic factor produced by the human placenta that has been implicated in the pathogenesis of preeclampsia and intrauterine growth restriction. Placenta growth factor belongs to the vascular endothelial growth factor family and is capable of inducing proliferation, migration, and activation of endothelial cells. The objective of this study was to determine the relationship between amniotic fluid concentration of placenta growth factor and gestational age, parturition (term and preterm), spontaneous rupture of the membranes, and intra-amniotic infection. STUDY DESIGN: Amniotic fluid samples obtained from 273 pregnant patients were assayed in the following clinical groups: midtrimester pregnancy, preterm labor who delivered at term, preterm labor without microbial invasion of the amniotic cavity who delivered preterm, preterm labor with microbial invasion of the amniotic cavity, term not in labor, term in labor, term with microbial invasion of the amniotic cavity, preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, and term with premature rupture of membranes without microbial invasion of the amniotic cavity. The placenta growth factor concentrations were determined by an immunoassay that is both sensitive and specific. RESULTS: Placenta growth factor was detectable in 96.3% (263/273) of samples. Amniotic fluid placenta growth factor concentration decreased with advancing gestational age (r = -0.42; P <.001). Amniotic fluid placenta growth factor concentrations were significantly higher in women in midtrimester pregnancy than in those at term not in labor (midtrimester pregnancy: median, 43.1 pg/mL; range, 22.9-69.8 pg/mL; vs term not in labor: median, 28.7 pg/mL; range, 16.1-82.7 pg/mL; P <.01). Neither term nor preterm parturition was associated with a change in amniotic fluid placenta growth factor concentrations. Term premature rupture of membranes was associated with a significant decrease in amniotic fluid placenta growth factor concentration (term premature rupture of membranes: median, 16.5 pg/mL; range <5.2-195.1 pg/mL; vs term intact membranes: median, 28.7 pg/mL; range, 16.1 822.7 pg/mL; P <.005). Preterm premature rupture of membranes was not associated with changes in amniotic fluid placenta growth factor concentrations. Intra amniotic infection in preterm labor, term labor with intact membranes, and preterm premature rupture of membranes were not associated with changes in amniotic fluid placenta growth factor concentrations. CONCLUSION: Placenta growth factor is a physiologic constituent of amniotic fluid. Amniotic fluid concentrations of placenta growth factor decrease with advancing gestational age. Neither parturition nor infection affects amniotic fluid placenta growth factor concentrations. PMID- 10871491 TI - Neonatal pulmonary hypoplasia and perinatal mortality in patients with midtrimester rupture of amniotic membranes--a critical analysis. AB - OBJECTIVE: We sought to critically assess the risk factors for neonatal pulmonary hypoplasia and perinatal death in patients with preterm rupture of the amniotic membranes from 15 to 28 weeks' gestation. STUDY DESIGN: This was a prospective cohort study. The study patients had preterm rupture of the amniotic membranes at 15 to 28 weeks' gestation and were without fetal anomalies, multiple gestation, and oligohydramnios before rupture of the membranes. The amniotic fluid volume index was determined at admission and weekly afterward until delivery. RESULTS: The incidence of pulmonary hypoplasia was 12.9% (21/163). The overall perinatal mortality rate was 54% (88/163). Logistic regression analysis revealed the following: (1) Gestational age at rupture of the membranes, the latency period, and either the initial or the average amniotic fluid index have significant influence on the development of pulmonary hypoplasia; (2) gestational age at rupture of the membranes and latency period are significant factors in predicting perinatal death. CONCLUSIONS: In this large population of patients with rupture of membranes at 15 to 28 weeks' gestation, gestational age at rupture of the membranes, latency period, and amniotic fluid index were important independent predictors of neonatal pulmonary hypoplasia. In addition, gestational age at rupture of the membranes and latency period were important independent determinants of perinatal death. Expectant management of patients with preterm rupture of the amniotic membranes during this gestational age interval was associated with improved perinatal survival, even though it may increase the risk of pulmonary hypoplasia. PMID- 10871492 TI - Nucleated red blood cells as a marker of acidemia in term neonates. PMID- 10871493 TI - Nucleated red blood cells in term infants. PMID- 10871495 TI - Dietary sodium restriction in rats and human beings. PMID- 10871497 TI - Laparoscopic sentinel lymph node detection in patients with cervical cancer. PMID- 10871498 TI - The return of subtotal hysterectomy. PMID- 10871500 TI - About singleton breech deliveries. PMID- 10871502 TI - Not all cases in a reported series were acute fatty liver of pregnancy. PMID- 10871532 TI - Quality of care issues and nursing research that gives voice to the vulnerable. PMID- 10871533 TI - Oncology patients' perceptions of quality nursing care. AB - The purpose of this study was to analyze theoretically oncology patients' perceptions of the attributes and outcomes of quality nursing care. The grounded theory method as described by Strauss and Corbin (1998) was used. The purposive sample comprised 22 oncology patients being treated at an urban medical center; they were interviewed using a semistructured schedule. Eight attributes of quality nursing care emerged from the data. From the patient's perspective, excellent care was characterized by professional knowledge, continuity, attentiveness, coordination, partnership, individualization, rapport, and caring. In addition, two outcomes of quality care included increased fortitude and a sense of well-being with its constituents of trust, optimism, and authenticity. These findings can inform investigations of how oncology patients may experience "being well cared-for by nurses. PMID- 10871535 TI - Living on the edge: a phenomenological study of medically uninsured working Americans. AB - An estimated 35 to 50 million Americans have no medical insurance; the vast majority are employed persons and their dependents. This phenomenological study was developed to make visible the experience of working Americans living on the edge-forced to walk a fine line between health and illness without the safety net of medical insurance. A purposive sample of 12 individuals was asked, "What is it like to be working and without medical insurance? Based on textual analysis, using an adaptation of Colazzi's method, themes were grouped into four theme clusters: A Marginalized Life, Up Against Rocks and Hard Places, Making Choices Chancing It, and Getting By-More or Less. These are illustrated through commentary and direct quotation to depict an overall sense of the experience. Implications for nurses charged with addressing the needs of the medically uninsured and for nursing as a whole are discussed. PMID- 10871534 TI - Family caregiving skill: development of the concept. AB - Families increasingly are expected to provide complex care at home to ill relatives. Such care requires a level of caregiving knowledge and skill unprecedented among lay persons, yet family caregiving skill has never been formally developed as a concept in nursing. The purpose of the study reported here was to develop the concept of family caregiving skill systematically through qualitative analysis of interviews with patients (n = 30) receiving chemotherapy for cancer and their primary family caregivers (n = 29). Open coding and constant comparison constituted the analytic methods. Sixty-three indicators of caregiving skill were identified for nine core caregiving processes. Family caregiving skill was defined as the ability to engage effectively and smoothly in these nine processes. Properties of family caregiving skill also were identified. Conceptualizing skill as a variable and identifying indicators of varying levels of skill provides a basis for measurement and will allow clinicians to more precisely assess family caregiving skill. PMID- 10871536 TI - Self-reported weapon ownership, use, and violence experience among clients accessing an inner-city sexually transmitted disease clinic. AB - Little is known about the extent to which people who access public health care settings own/carry weapons and experience/perpetrate acts of violence. The purpose of this study was to describe weapon ownership and violence experiences of persons attending an inner-city sexually transmitted disease clinic. Face-to face interviews were administered to 245 clients to assess weapon ownership, types of weapons carried, and experiences as victims or perpetrators of violent acts. Overall, 43.7% reported experience of carrying a weapon at some point in their lives. More men chose to carry guns; more women chose to carry knives or mace. Participants reported experiencing alarming levels of violence in the previous year: 30.5% experienced beatings, 23.9% reported being threatened with a gun, and 18.9% reported forced, unwanted sex. Persons with a history of carrying weapons were significantly more likely to report being both victims and perpetrators of violence. Persons who experienced violence in the previous month were significantly more likely to be diagnosed with an STD. Results show that STD clinics represent yet another setting wherein interventions to curb the extent of violence might be appropriate, and strategies to assist and protect those experiencing violence are needed. PMID- 10871537 TI - Tumor necrosis factor alpha and interleukin-6 have differential effects on food intake and gastric emptying in fasted rats. AB - Interleukin-1 (IL-1), IL-6, and tumor necrosis factor (TNF) are thought to mediate the onset of anorexia with infection. Animal studies suggest that gastric stasis accompanies IL-1-induced anorexia, and that food intake and gastric emptying of IL-1-injected rats are improved by pretreatment with ibuprofen (ibu), an inhibitor of prostaglandin (PG) synthesis. The purpose of the present study was to determine if gastric stasis accompanies the reduced food intake induced by intraperitoneal injection of TNFa or IL-6 and whether these effects are mediated by PG. Injection of TNFa reduced food intake of fasted rats, but did not affect gastric emptying; injection of IL-6 reduced both food intake and gastric emptying. Pretreatment with 10 mg/kg ibu improved food intake of TNFa-injected animals, but did not affect food intake or gastric emptying of IL-6-injected animals. These data indicate that although IL-6 and TNFa have overlapping effects on food intake, the mechanisms of action are not identical. Delayed gastric emptying does not play a major role in the anorexigenic effects of TNFa, and PG synthesis does not play a major role in the anorexigenic effects of IL-6. These findings may be helpful in the development of interventions to improve nutritional intake during infection. PMID- 10871538 TI - Pretty and powerless: nurses in advertisements, 1930-1950. AB - Images of nurses in pictorial advertisements from all issues of hospital administration journals published in 1930, 1940, and 1950 (N = 598) were examined. Content analysis of the data was based on Goffman's classic 1979 study on gender advertisements. Nurses also were compared with other figures in the advertisements and nursing activities were described. Nurses were predominantly portrayed as female, young, eager to please, and without the appearance of wisdom. In group scenes, nurses were placed as subordinate to physicians and hospital administrators. Nurses in 1940 performed more complex, autonomous activities than in 1930 and 1950. These findings support previous research focused on more recent portrayals of women and nurses in communication media. The overt and subtle subordinate representation of nurses in these advertisements, compared with physicians and administrators, reveals one facet of nursing's heritage as a woman's profession. PMID- 10871539 TI - Development of a measure of resident satisfaction with the nursing home. AB - A satisfaction instrument specifically designed for use with nursing home residents, the Satisfaction with the Nursing Home Instrument (SNHI), was developed and tested with a sample of 110 nursing home residents from three proprietary facilities in Minnesota. As hypothesized, significant relationships were found between SNHI scores and measures of affect (negatively associated with depression and positively associated with morale), providing support for the construct validity of the scale. The lack of a significant relationship between SNHI scores and both age and mental status confirmed the predicted divergent validity of the instrument. The alpha coefficient for the 29-item scale was 0.81. PMID- 10871541 TI - Introduction to histological localization of prion proteins. PMID- 10871540 TI - Combining qualitative and quantitative sampling, data collection, and analysis techniques in mixed-method studies. AB - Researchers have increasingly turned to mixed-method techniques to expand the scope and improve the analytic power of their studies. Yet there is still relatively little direction on and much confusion about how to combine qualitative and quantitative techniques. These techniques are neither paradigm- nor method-linked; researchers' orientations to inquiry and their methodological commitments will influence how they use them. Examples of sampling combinations include criterion sampling from instrument scores, random purposeful sampling, and stratified purposeful sampling. Examples of data collection combinations include the use of instruments for fuller qualitative description, for validation, as guides for purposeful sampling, and as elicitation devices in interviews. Examples of data analysis combinations include interpretively linking qualitative and quantitative data sets and the transformation processes of qualitizing and quantitizing. PMID- 10871542 TI - Variant Creutzfeldt-Jakob disease: immunocytochemical studies and image analysis. AB - Variant Creutzfeldt-Jakob disease (vCJD) is a recently identified human prion disease that appears to arise from exposure to the bovine spongiform encephalopathy agent. The clinical features and neuropathology of vCJD are distinctive, particularly the patterns of PrP(sc) accumulation in the brain. PrP immunocytochemistry has also demonstrated the accumulation of PrP(sc) in tissues outside the central nervous system, including sensory ganglia and lymphoid tissues. These observations have allowed the use of tonsillar biopsy as an investigation to aid the diagnosis of vCJD, since accumulation of PrP(sc) in lymphoid tissues does not occur in other forms of human prion disease. The patterns of PrP(sc) accumulation in vCJD can be studied by image analysis techniques, using both quantitative and qualitative approaches. Preliminary results of textural analysis are presented, which indicate that this approach can be used to discriminate and study the unique features of PrP(sc) accumulation in the brain in vCJD. This technique has major potential as a research tool in human prion diseases, particularly for the characterisation of disease phenotype in large series of cases. PMID- 10871543 TI - Neuropathology of Gerstmann-Straussler-Scheinker disease. AB - Gerstmann-Straussler-Scheinker disease is a familial neurodegeneration characterized clinically by adult-onset ataxia, postural abnormalities, and cognitive decline, and pathologically by amyloid deposits mostly localized in the cerebral and cerebellar cortices and the basal ganglia. The disease is due to mutations in the prion protein gene. Processing of the mutant proteins originates the amyloidogenic fragments that accumulate in the tissue. PrP-immunoreactive amyloid deposits are the morphological hallmark of the disease. Hypertrophic astrocytes, activated microglia, and nerve cell loss are consistently associated with PrP-amyloid deposits, while spongiosis, diffuse PrP immunoreactivity, neurofibrillary tangles, Lewy bodies, and long fiber tracts degeneration are occasionally associated. The clinical and pathological variability observed in GSS families is related to both mutations and the M/V polymorphism at codon 129 of the mutated gene. PMID- 10871544 TI - Intracerebral distribution of the abnormal isoform of the prion protein in sporadic Creutzfeldt-Jakob disease and fatal insomnia. AB - Molecular genetics and protein chemistry have led to major advances in our understanding of the molecular basis of phenotypic variability of prion diseases. A large body of evidence indicates that a common methionine/valine polymorphism at codon 129 in the prion protein gene (PRNP), alone or in conjunction with PRNP mutations, modulates both disease susceptibility and phenotypic expression of human prion diseases. In addition, there are physicochemical properties of the abnormal isoform of the prion protein (PrP(sc)), such as relative molecular mass and glycosylation, that correlate with distinct phenotypes even in subjects carrying the same PRNP genotype. Different PrP(sc) "type"-PRNP genotype combinations are found associated with pathological phenotypes that differ in the relative severity of lesions among distinct brain regions, the presence and morphology of certain lesions such as amyloid plaques, and the pattern of intracerebral and tissue deposition of PrP(sc). This review summarizes the currently available data on the molecular pathology of sporadic Creutzfeldt-Jakob disease, the most common human prion disease, and fatal insomnia, a more recently defined entity that has rapidly become one of the best characterized of the human prion diseases. PMID- 10871545 TI - PrP immunohistochemistry: different protocols, including a procedure for long formalin fixation, and a proposed schematic classification for deposits in sporadic Creutzfeldt-Jakob disease. AB - The use of immunohistochemistry on formalin-fixed and paraffin-embedded tissue has greatly improved the neuropathological diagnosis of Creutzfeldt-Jakob disease and the other subacute spongiform encephalopathies in human and animals. Two pitfalls of this technique, however, currently exist: low sensitivity after long formalin fixation and difficulties in interpreting some images. Here we review the protocols currently in use for the pretreatment of sections allowing PrP detection by immunohistochemistry. In addition, a technique useful after long formalin fixation is reported: enzymatic digestion with proteinase K (24 degrees C, 1/100 for 8 minutes) was employed in addition to the usual autoclaving (121 degrees C for 10 minutes) followed by formic acid (99% for 5 minutes) and 4M guanidine thiocyanate (4 degrees C for 2 hours). This allowed a substantial increase in the sensitivity of 3F4 immunohistochemistry on paraffin-embedded tissue, especially after prolonged formalin fixation. In addition, we suggest a simple method for classification of PrP immunolabelling in sporadic Creutzfeldt Jakob disease that would allow easy comparisons. PMID- 10871546 TI - Applicability of three anti-PrP peptide sera including staining of tonsils and brainstem of sheep with scrapie. AB - Three rabbit antibodies (R521, R505, R524) were produced, and raised to synthetic peptides corresponding to residues 94-105, 100-111, and 223-234, respectively, of the sheep prion protein (PrP). Epitope mapping analysis revealed the monospecific character of antisera R505 and R524. In addition to the amino acid sequence against which it was raised, R521 also recognized other small epitopes. ELISA and radio-immunoprecipitation were used to assess the relative immunoreactivities of the antisera to the normal sheep prion protein (PrP(c)). Highest reactivity was found for R521, followed by R505 and R524. According to Western blot analysis, all three sera specifically reacted with the prion proteins PrP(Sc) and PrP27-30, extracted from the brain stem of a scrapie-affected sheep. Yet, with R505 not all of the lower molecular weight deglycosylated forms could be detected. Contrary to the immunoreactivities found with the PrP(Sc) and PrP27-30 isoforms, only R521 recognised PrP(c) from a healthy sheep. The usefulness of all three anti-peptide sera in the immunohistochemical detection of PrP(Sc) in brain stem and tonsils of scrapie-affected sheep was demonstrated and compared with an established rabbit anti-PrP serum. PMID- 10871547 TI - Detection of PrP in extraneural tissues. AB - Transmissible spongiform encephalopathies (TSEs) or "prion diseases" are a group of unconventional fatal diseases. TSEs are characterised by the accumulation of a modified form of the normal host glycoprotein, PrP (PrP(c)). In the course of infection PrP(c) is converted to an abnormally protease resistant form, PrP(Sc). The exact nature of the infectious agent responsible for these diseases remains controversial. While there is compelling evidence that TSE agents contain an informational molecule, possibly a nucleic acid, some believe that the infectious agent or "prion" is solely composed of PrP(Sc). Nevertheless, PrP is required for TSE pathogenesis, as mice devoid of the PrP gene (PrP(-/-)) remain healthy when challenged with TSE isolates and are unable to replicate infectivity within the central nervous system (CNS) or in other tissues. In recent years immunocytochemistry has been used to pinpoint which cells are associated with abnormal accumulations of PrP, providing important information on the cellular targeting of TSE infection. In uninfected and scrapie-infected mice, PrP protein is found in the CNS and in extraneural tissues such as spleen and lymph nodes. In the peripheral lymphoid system, PrP is associated with follicular dendritic cells that are known to be important for replication of infectivity for at least one TSE strain. This review will focus on current methods for the immunocytochemical detection of PrP in murine extraneural tissues, mainly lymphoid tissues, and will discuss recent findings on the role of the peripheral lymphoid system in TSE pathogenesis. PMID- 10871548 TI - Tubulovesicular structures: what are they really? AB - The tubulovesicular structures (TVS) are the only structures unique at the level of thin-section electron microscopy for all TSEs so far examined. They were first described in NIH Swiss mice infected intracerebrally with the "Chandler" strain of scrapie by David-Ferreira et al. in 1968 [Proc. Soc. Exp. Biol. Med. 127:313 320]. TVS were described as "particles and rods ranging in diameter from 320 to 360 A(o)." The exact topology of TVS is not entirely clear. In most published electron micrographs, TVS appeared as spheres measuring between 20 and 40 nm in diameter. The number of neuritic processes containing TVS increases through the incubation period and has been shown to correlate with the incubation period and titre of infectivity in three longitudinal disease studies of scrapie and CJD. These studies, therefore, suggest that TVS may represent a primary pathogenetic event rather than a pathological product of disease. The predominant theory of the scrapie agent is now the "prion hypothesis" and its derivatives, which implies that a conformationally altered abnormal isoform (PrP(Sc) or PrP*) of a normal cellular membrane glycoprotein (PrP(c)) is the agent and its accumulation merely mimicks replication. If an abnormal fraction of PrP is indeed the infectious agent, (although it is no longer suggested in some quarters that protease resistant fraction of PrP(Sc) is the agent). The absence of stainable PrP in TVS, however, would indicate that they are not the ultrastructural correlate of the agent. However, TVS appear to be specific and unique to the TSEs, appearing before the earliest pathological changes and increasing in line with incubation period or titre. The very existence of TVS and their correlation with infectivity, therefore, urgently needs an explanation. PMID- 10871549 TI - Immunolocalization of the cellular prion protein in normal brain. AB - We examined the localization of PrP(c) in normal brain using free-floating section immunohistochemistry and monclonal antibody 3F4. In the mature hamster and baboon brain, PrP(c) is localized to the neuropil with a synaptic distribution and the PrP(c) immunoreactivity is denser in regions known for ongoing plasticity. Cell bodies and major fiber tracts have little or no PrP(c) immunoreactivity. At the electron microscopic level, PrP(c) immunoreactivity decorates synaptic profiles, both pre- and postsynaptically. Results obtained with two additional antibodies, 3B5 and Pri-304, showed similar patterns of PrP(c) bands on Western blots, although Pri-304 was less sensitive. On sections through the adult hamster hippocampus, 3B5 and Pri-304 both stained the synaptic neuropil while cell bodies in the pyramidal and dentate granule cell layers were not immunoreactive. Pri-304 differentiated between synaptic layers in the hippocampus and closely resembled the pattern of staining obtained with 3F4. Preliminary results of developing brain showed that PrP(c) is initially localized along fiber tracts in the neonate brain. These results show that PrP(c) has a synaptic distribution in the adult brain and suggest that there are important changes in its distribution during brain development. These results also characterize two additional reagents for studies of PrP(c) localization. PMID- 10871550 TI - Synaptic prion protein immuno-reactivity in the rodent cerebellum. AB - The cellular prion protein PrP(c) is a neurolemmal glycoprotein essential for the development of the transmissible spongiform encephalopathies. In these neurodegenerative diseases, host PrP(c) is converted to infectious protease resistant isoforms PrP(res) or prions. Prions provoque predictable and distinctive patterns of PrP(res) accumulation and neurodegeneration depending on the prion strain and on regional cell-specific properties modulating PrP(c) affinity for infectious PrP(res) in the host brain. Synaptolysis and synaptic accumulation of PrP(res) during PrP-related diseases suggests that the synapses could be primary sites able to propagate PrP(res) and neurodegeneration in the central nervous system. In the rodent cerebellum, the present light and electron microscopic immuno-cytochemical analysis shows that distinct types of synapses display differential expression of PrP(c), suggesting that synapse-specific parameters could influence neuroinvasion and neurodegeneration following cerebral infection by prions. Although the physiological functions of PrP(c) remain unknown, the concentration of PrP(c) almost exclusively at the Purkinje cell synapses in the cerebellum suggests its critical involvement in the synaptic relationships between cerebellar neurons in agreement with their known vulnerability to PrP deficiencies. PMID- 10871551 TI - Ultrastructural localization of prion proteins: physiological and pathological implications. AB - The transmissible spongiform encephalopathies (TSE) or prion diseases are fatal neurodegenerative disorders in which the central event is the conversion of a normal host-encoded protein (PrP(c)) into an abnormal isoform (PrP(sc)) which accumulates as amyloid in TSE brain. The two PrP(c) and PrP(sc) prion protein isoforms are membrane sialoglycoproteins synthesized in the central nervous system and various peripheral organ tissues. In this review, we describe the ultrastructural localization of prion proteins in human and animal cerebral and non-cerebral tissues whether or not infected by TSE agents. In addition to the plasma membrane of several cells, PrP(c) was found in association with cytoplasmic organelles of central and nerve-muscle synapses, and secretory granules of epithelial cells. Fibrils of amyloid plaques, synaptic structures, and lysosome-like organelles constitute the subcellular sites harboring PrP(sc). These findings have led to discussions on the physiological role of PrP(c) and the pathological mechanisms underlying prion spongiform encephalopathies. PMID- 10871552 TI - When does hyporetinolemia mean vitamin A deficiency? PMID- 10871553 TI - Is impaired folate absorption a factor in neural tube defects? PMID- 10871554 TI - Colostrum and milk-derived peptide growth factors for the treatment of gastrointestinal disorders. AB - Colostrum is the specific first diet of mammalian neonates and is rich in immunoglobulins, antimicrobial peptides, and growth factors. In this article we review some of these constituents of human and bovine colostrum in comparison with those of mature milk. Recent studies suggest that colostral fractions, or individual peptides present in colostrum, might be useful for the treatment of a wide variety of gastrointestinal conditions, including inflammatory bowel disease, nonsteroidal antiinflammatory drug-induced gut injury, and chemotherapy induced mucositis. We therefore discuss the therapeutic possibilities of using whole colostrum, or individual peptides present in colostrum, for the treatment of various gastrointestinal diseases and the relative merits of the 2 approaches. PMID- 10871555 TI - Diet composition and body composition in preschool children. AB - BACKGROUND: In studies of adult humans and in animal models, dietary intakes of the macronutrients, particularly fat, are related to body composition; however, data on children are more scarce. OBJECTIVE: We sought to determine whether diet composition is related to percentage body fat in children aged 1.5-4.5 y. DESIGN: In 77 preschool children, a 4-d weighed-food record was used to determine intakes of total energy and energy from each macronutrient. An oxygen-18 dilution method was used to calculate percentage body fat. Habitual physical activity level was determined by calculating the ratio of total energy expenditure (from stable isotope analyses) to predicted basal metabolic rate. Dietary intake and body composition data were analyzed to evaluate whether diet composition was related to body fat. Further analyses incorporating physical activity level were performed. RESULTS: Percentage body fat was not significantly correlated with dietary intake variables (total energy or percentage of energy from fat, carbohydrate, or protein) and did not differ significantly among 3 increasing levels of each dietary intake variable by analysis of variance. In multiple regression analysis, physical activity level was related to body fat whereas diet composition was not. CONCLUSIONS: We found no relations between dietary intakes of total energy, fat, carbohydrate, or protein and percentage body fat in children. The relation between fat intake and body fat may develop over time and may not be evident in preschool children. Energy expenditure, in particular physical activity level, may have a greater influence on body composition in early childhood. PMID- 10871556 TI - Sex-related differences in methionine metabolism and plasma homocysteine concentrations. AB - BACKGROUND: Elevated fasting homocysteine concentrations are considered a risk factor for vascular disease. Homocysteine, which is produced by the transmethylation of methionine, can be either remethylated back to methionine or metabolized via transsulfuration to cystathionine. It has been speculated that the lower risk of vascular disease among premenopausal women may be related to lower homocysteine concentrations in women than in men. OBJECTIVE: This study was designed to determine whether sex-related differences exist in methionine cycle kinetics, which may account for the reportedly lower fasting homocysteine concentrations in premenopausal women. DESIGN: Eleven healthy young men and 11 premenopausal women without cardiac risk factors were studied by using stable isotope-labeled L-[methyl-(2)H(3),1-(13)C]methionine and L-[methyl- (2)H(3)]leucine. After 3 h of tracer infusion, 100 mg unlabeled L-methionine/kg body wt was ingested. Blood and breath samples were obtained at timed intervals. Fat-free mass was estimated by dual-energy X-ray absorptiometry and muscle mass by urinary creatinine excretion. RESULTS: No significant sex-related differences were found in fasting homocysteine concentrations, responses to the oral methionine load, or rates of methionine flux based on carboxyl or methyl labels. However, women had significantly higher remethylation rates than did men (P < 0.005) and a tendency toward higher transmethylation (P < 0.10). Whereas adjustment of remethylation rates for fat-free mass tended to attenuate the sex related effect (P = 0.08), adjustment for muscle mass did not (P < 0.04). In contrast, significant sex-related differences in leucine flux (P < 0.02) were eliminated after adjustment for either fat-free mass or muscle mass. CONCLUSION: Reported differences between men and women in homocysteine concentrations may be partially explained by differences in rates of homocysteine remethylation. PMID- 10871557 TI - Cocoa inhibits platelet activation and function. AB - BACKGROUND: Epidemiologic studies have shown inverse associations between dietary polyphenols and mortality from coronary heart disease. However, the basis for this protective association is uncertain. Food polyphenols reportedly have antioxidant properties and decrease platelet function in vitro. OBJECTIVE: This study sought to evaluate whether consumption of a polyphenol-rich cocoa beverage modulates human platelet activation and primary hemostasis. DESIGN: Peripheral blood was obtained from 30 healthy subjects before and 2 and 6 h after ingestion of a cocoa beverage (n = 10), a caffeine-containing control beverage (n = 10), or water (n = 10). Platelet activation was measured in terms of expression of activation-dependent platelet antigens and platelet microparticle formation by using fluorescent-labeled monoclonal antibodies and flow cytometry. Primary platelet-related hemostasis was measured with a platelet function analyzer. RESULTS: Ex vivo epinephrine- or ADP-stimulated expression of the fibrinogen binding conformation of glycoprotein IIb-IIIa was lower 2 and 6 h after consumption of cocoa than before consumption. Cocoa consumption also decreased ADP-stimulated P-selectin expression. In contrast, epinephrine-induced platelet glycoprotein IIb-IIIa expression increased after consumption of the caffeine containing beverage but not after water consumption. Platelet microparticle formation decreased 2 and 6 h after cocoa consumption but increased after caffeine and water consumption. Primary hemostasis in response to epinephrine in vitro was inhibited 6 h after cocoa consumption. The caffeine-containing beverage inhibited ADP-induced primary hemostasis 2 and 6 h after consumption. CONCLUSIONS: Cocoa consumption suppressed ADP- or epinephrine-stimulated platelet activation and platelet microparticle formation. Cocoa consumption had an aspirin like effect on primary hemostasis. PMID- 10871558 TI - Low-fat and high-monounsaturated fatty acid diets decrease plasma cholesterol ester transfer protein concentrations in young, healthy, normolipemic men. AB - BACKGROUND: Cholesterol ester transfer protein (CETP) mediates the transfer of cholesteryl esters from HDL to apolipoprotein (apo) B-containing lipoproteins. The possible atherogenic role of this protein is controversial. Diet may influence plasma CETP concentrations. OBJECTIVE: The objective was to determine whether the changes in plasma lipids observed after consumption of 2 lipid lowering diets are associated with changes in plasma CETP concentrations. DESIGN: : We studied 41 healthy, normolipidemic men over 3 consecutive 4-wk dietary periods: a saturated fatty acid-rich diet (SFA diet: 38% fat, 20% saturated fat), a National Cholesterol Education Program Step I diet (NCEP Step I diet: 28% fat, 10% saturated fat), and a monounsaturated fatty acid-rich diet (MUFA diet: 38% fat, 22% monounsaturated fat). Cholesterol content (27.5 mg/MJ) was kept constant during the 3 periods. Plasma concentrations of total, LDL, and HDL cholesterol; triacylglycerol; apo A-I and B; and CETP were measured at the end of each dietary period. RESULTS: Compared with the SFA diet, both lipid-lowering diets significantly decreased plasma total and LDL cholesterol, apo B, and CETP. Only the NCEP Step I diet lowered plasma HDL cholesterol. Positive, significant correlations were found between plasma CETP and total (r = 0.3868, P < 0.0001) and LDL (r = 0.4454, P < 0.0001) cholesterol and also between changes in CETP concentrations and those of total (r = 0.4543, P < 0.0001) and LDL (r = 0.4554, P < 0.0001) cholesterol. CONCLUSIONS: The isoenergetic substitution of a high saturated fatty acid diet with an NCEP Step I or a high-monounsaturated fatty acid diet decreases plasma CETP concentrations. PMID- 10871559 TI - Biochemical effects of a diet containing foods enriched with n-3 fatty acids. AB - BACKGROUND: Results of many studies indicate that consumption of n-3 fatty acids can benefit persons with cardiovascular disease and rheumatoid arthritis. However, encapsulated fish oil is unlikely to be suited to lifetime daily use and recommendations to increase fish intake have not been effective. OBJECTIVE: The objective was to examine the effectiveness of a diet that incorporates foods rich in n-3 fatty acids in elevating tissue concentrations of eicosapentaenoic acid and in suppressing the production of inflammatory mediators. DESIGN: Healthy male volunteers were provided with foods that were enriched in alpha-linolenic acid (cooking oil, margarine, salad dressing, and mayonnaise) and eicosapentaenoic and docosahexaenoic acids (sausages and savory dip) and with foods naturally rich in n-3 fatty acids, such as flaxseed meal and fish. Subjects incorporated these products into their food at home for 4 wk. Fatty acid intakes, cellular and plasma fatty acid concentrations, and monocyte-derived eicosanoid and cytokine production were measured. RESULTS: Analyses of dietary records indicated that intake of eicosapentaenoic acid plus docosahexaenoic acid averaged 1.8 g/d and intake of alpha-linolenic acid averaged 9. 0 g/d. These intakes led to an average 3-fold increase in eicosapentaenoic acid in plasma, platelet, and mononuclear cell phospholipids. Thromboxane B(2), prostaglandin E(2), and interleukin 1beta synthesis decreased by 36%, 26%, and 20% (P < 0.05), respectively. CONCLUSIONS: Foods that are strategically or naturally enriched in n-3 fatty acids can be used to achieve desired biochemical effects without the ingestion of supplements or a change in dietary habits. A wide range of n-3-enriched foods could be developed to support large-scale programs on the basis of the therapeutic and disease preventive effects of n-3 fatty acids. PMID- 10871560 TI - Modest doses of beta-glucan do not reduce concentrations of potentially atherogenic lipoproteins. AB - BACKGROUND: In 1997, the US Food and Drug Administration passed a unique ruling that allowed oat bran to be registered as the first cholesterol-reducing food at a dosage of 3 g beta-glucan/d. OBJECTIVE: The effects of a low dose of oat bran in the background diet only were investigated in volunteers with mild-to-moderate hyperlipidemia. DESIGN: The study was a double-blind, placebo-controlled, randomized, parallel study. Sixty-two healthy men (n = 31) and women (n = 31) were randomly allocated to consume either 20 g oat bran concentrate (OBC; containing 3 g beta-glucan) or 20 g wheat bran (control) daily for 8 wk. Fasting blood samples were collected at weeks -1, 0, 4, 8, and 12. A subgroup (n = 17) was studied postprandially after consumption of 2 meals (containing no OBC or wheat bran) at baseline and after supplementation. Fasting plasma samples were analyzed for total cholesterol, HDL cholesterol, triacylglycerol, glucose, and insulin. LDL cholesterol was measured by using the Friedewald formula. The postprandial samples were anlayzed for triacylglycerol, glucose, and insulin. RESULTS: No significant difference was observed in fasting plasma cholesterol, LDL cholesterol, glucose, or insulin between the OBC and wheat-bran groups. HDL cholesterol concentrations fell significantly from weeks 0 to 8 in the OBC group (P = 0.05). There was a significant increase in fasting glucose concentrations after both OBC (P = 0.03) and wheat-bran (P = 0.02) consumption. No significant difference was found between the OBC and wheat-bran groups in any of the postprandial variables measured. CONCLUSIONS: A low dosage of beta-glucan (3 g/d) did not significantly reduce total cholesterol or LDL cholesterol in volunteers with plasma cholesterol concentrations representative of a middle-aged UK population. PMID- 10871561 TI - Plasma lipid and lipoprotein responsiveness to dietary fat and cholesterol in premenopausal African American and white women. AB - BACKGROUND: Premenopausal African American women have a 2-3 times greater incidence of coronary heart disease (CHD) than do white women. The plasma lipid responsiveness to dietary fat, which may be associated with CHD, has not been adequately studied in premenopausal African American or white women. OBJECTIVE: The objective of our study was to compare the effect of diet on fasting plasma lipids and lipoproteins and postprandial lipemia in premenopausal African American and white women. DESIGN: Thirteen African American and 9 white healthy premenopausal women were fed a low-fat, high-fiber diet and a high-fat, low-fiber diet for 4 wk each in a randomized crossover design. Fasting plasma lipid and lipoprotein concentrations and the 24-h plasma triacylglycerol response to a standard fatty test meal were measured at the end of each dietary period. RESULTS: Plasma total and LDL-cholesterol concentrations were higher after the high-fat, low-fiber diet in both white and African American women (P < 0.0001). The 24-h area under the plasma triacylglycerol curve after the test meal was lower after the low-fat diet than after the high-fat diet (P < 0.04). CONCLUSIONS: African American and white women had lower fasting plasma total and LDL-cholesterol concentrations and less postprandial lipemia after the low-fat than the high-fat diet. Diets low in total and saturated fat and cholesterol and high in fiber may reduce the risk of CHD by lowering fasting plasma total and LDL cholesterol concentrations and by reducing the lipemic response to fatty meals. PMID- 10871562 TI - Simple pediatric nutritional risk score to identify children at risk of malnutrition. AB - BACKGROUND: Although hospitalized children are at risk of malnutrition, routine screening of nutritional status has been hindered by lack of a validated nutritional assessment tool. OBJECTIVE: Our aim was to develop a simple pediatric nutritional risk score that could be used at hospital admission to identify patients at risk of acute malnutrition during hospitalization. DESIGN: Nutritional risk was assessed prospectively in 296 children. Anthropometric measurements, food intake, ability to eat and retain food, medical condition, and symptoms interfering with feeding (pain, dyspnea, and depression) were evaluated within 48 h of admission. Pathology was classified as mild (grade 1), moderate (grade 2), or severe (grade 3). The risk of weight loss was investigated with stepwise logistic regression. RESULTS: Weight loss during hospitalization occurred in 65% of the children and was >2% of admission weight in 45% of patients. Multivariate analysis indicated that food intake <50%, pain, and grade 2 and 3 pathologic conditions (P = 0.0001 for all) were associated with weight losses of >2%. The nutritional risk score ranged from 0 to 5 and was calculated by adding the values for the significant risk factors as follows: 1 for food intake <50%, 1 for pain, 1 for grade 2 pathologic condition, and 3 for grade 3 pathologic condition. A score of 1 or 2 indicated moderate risk and a score >/=3 indicated high risk of malnutrition. CONCLUSIONS: This simple score is suitable for routine use to identify patients at risk of malnutrition during hospitalization. Implementation may prevent hospital-acquired malnutrition. PMID- 10871563 TI - Body composition in children with celiac disease and the effects of a gluten-free diet: a prospective case-control study. AB - BACKGROUND: Celiac disease is the most common cause of malnutrition in children of Western countries. OBJECTIVE: The objective was to measure body composition in children at the time celiac disease was diagnosed and after consumption of a gluten-free diet (GFD). DESIGN: We assessed body composition by dual-energy X-ray absorptiometry in 29 children and adolescents with a mean (+/-SD) age of 9.5 +/- 3.4 y at the time celiac disease was diagnosed and in a subset of 20 patients after 1.2 +/- 0.2 y of a GFD. We also studied 23 patients aged 21.2 +/- 4.6 y who consumed a GFD for 10.6 +/- 4.5 y. Each patient was matched with a healthy control subject of the same age and sex. RESULTS: Untreated patients weighed less than control subjects (P = 0.04). Fat mass and bone mineral content were lower in the patients than in the control subjects (P < 0.01), as was lean mass of the limbs (P = 0.0013). After approximately 1 y of the GFD, there were no significant differences in body-composition values between patients and control subjects. Similarly, body-composition values of celiac disease patients who consumed the GFD long term were comparable with those of healthy subjects. CONCLUSIONS: Remarkable abnormalities in body composition were found in children at the time of diagnosis of celiac disease. Appropriate dietary treatment reverses body composition abnormalities quickly and the beneficial effects of gluten withdrawal are persistent. Because these results are harder to achieve if celiac disease is first diagnosed in adulthood, efforts to encourage early diagnosis of celiac disease should be made. PMID- 10871564 TI - Changes in body composition, substrate oxidation, and resting metabolic rate in adult celiac disease patients after a 1-y gluten-free diet treatment. AB - BACKGROUND: The incidence of celiac disease has been on the rise in both Europe and the United States. Celiac disease patients are at high risk of undernutrition because of nutrient malabsorption. OBJECTIVE: The aim of the present study was to evaluate changes in body composition and energy metabolism in a group of patients with celiac disease before and after consumption of a gluten-free diet (GFD). DESIGN: Body composition (by anthropometry and isotopic dilution), resting metabolic rate (RMR), and substrate oxidation rates (by indirect calorimetry) were assessed in 39 adult celiac disease patients (16 men and 23 women) with a mean (+/-SD) age of 29. 9 +/- 7.6 y, weight of 58.3 +/- 6.6 kg, and percentage body fat of 20.1 +/- 6.7%, and in 63 (29 men and 34 women) age- and height matched control subjects (age: 33.2 +/- 8.1 y; weight: 66.8 +/- 6.6 kg; and percentage body fat: 25.4 +/- 3.7%). Celiac disease patients were studied twice, at diagnosis and 1 y after treatment with a GFD. RESULTS: Before treatment, celiac disease patients had a lower body weight (P < 0.05) and a higher carbohydrate oxidation rate (P < 0.01) than did control subjects. Carbohydrate oxidation rates correlated positively with fecal lipid loss in untreated celiac disease patients (r = 0.80, P < 0.0001). After the GFD, percentage body fat was higher in celiac disease patients than in control subjects (P < 0.01), and lipid intakes tended to be higher than before treatment. CONCLUSIONS: This longitudinal study showed that the GFD treatment significantly increased body fat stores. Untreated patients preferentially utilized carbohydrates as a fuel substrate, probably as a consequence of both lipid malabsorption and a high carbohydrate intake, and lipid utilization increased with the restoration of the intestinal mucosa. PMID- 10871565 TI - Introducing a new component of the metabolic syndrome: low cholesterol absorption. AB - BACKGROUND: Weight reduction is the recommended treatment of obese type 2 diabetes, but the effects of weight reduction on cholesterol metabolism are poorly understood. OBJECTIVE: We investigated glucose, cholesterol, and lipoprotein metabolism at baseline and 2 y after weight reduction in obese patients with type 2 diabetes consuming an isoenergetic diet. DESIGN: Sixteen subjects were randomly chosen to consume a very-low-energy or low-energy diet for 3 mo, after which they consumed a weight-maintenance diet for up to 2 y. Cholesterol absorption and metabolism, LDL and HDL kinetics, and variables of glucose metabolism were studied at baseline and 2 y. RESULTS: Baseline serum sex hormone binding globulin (SHBG) was significantly related to cholesterol absorption efficiency, and serum glucose and insulin concentrations were associated with cholesterol synthesis. After 2 y, body weight was reduced by 6 +/ 1 kg (P < 0.01), body mass index by 6% (P < 0.05), and blood glucose by 14% (P < 0.01); the ratio of serum SHBG to insulin increased by 66% (P < 0.05). Serum and VLDL, LDL, and HDL triacylglycerol were significantly reduced by 13-24%. Despite unchanged serum concentrations of cholesterol, cholesterol absorption efficiency and the ratio of serum plant sterols to cholesterol-indicators of cholesterol absorption-increased by 28% (P < 0.01) and 20-31% (P < 0. 05 for both), respectively; the fractional removal of LDL apolipoprotein B decreased. Fecal excretion of cholesterol as neutral sterols decreased significantly by 11%. Changes in body weight were significantly negatively correlated with changes in ratios of cholesterol to serum plant sterols and cholestanol. CONCLUSIONS: Baseline cholesterol absorption and synthesis were related to respective serum SHBG, glucose, and insulin values. Weight reduction increased cholesterol absorption and improved variables of glucose metabolism. These results suggest that low cholesterol absorption and high synthesis may be part of the insulin resistance syndrome. PMID- 10871566 TI - Protein nutritional status and function are associated with type 2 diabetes in Hispanic elders. AB - BACKGROUND: Hispanic elders have a high prevalence of diabetes and poor glycemic control, leading to inadequate nutritional status, muscle wasting, and impaired function. OBJECTIVE: We examined the association of type 2 diabetes with nutritional status measured by serum albumin concentrations and midupper arm muscle area (MAM) and with function measured by difficulty with at least one activity of daily living. DESIGN: : Health history and disability were assessed by self report in 556 Hispanics with a mean (+/- SD) age of 69 +/- 7 y and 158 non-Hispanic whites (NHW; aged 71 +/- 7 y) from the Massachusetts Hispanic Elders Survey. Energy intake (in MJ/d) and protein intake (in g/d) were estimated with use of a food-frequency questionnaire. Body mass index (BMI; in kg/m(2)) and C reactive protein concentrations (in mg/L) were also measured. Multiple logistic regression models by ethnic group were used. RESULTS: There were no significant differences between Hispanics and NHWs in the proportion of those with low albumin concentrations or low MAM. Hispanic diabetic women had the lowest proportion of low MAM. The risk of low serum albumin concentration was twice as high in Hispanics taking insulin as in their NHW counterparts. Among Hispanics, low albumin concentration and low BMI were associated with low MAM; female sex, low albumin concentration, high BMI, and insulin use were significantly associated with risk of functional impairment. CONCLUSION: Type 2 diabetes is associated with poor nutritional status, muscle loss, and functional impairment among Hispanic elders. PMID- 10871567 TI - Plasma insulin responses after ingestion of different amino acid or protein mixtures with carbohydrate. AB - BACKGROUND: Protein induces an increase in insulin concentrations when ingested in combination with carbohydrate. Increases in plasma insulin concentrations have been observed after the infusion of free amino acids. However, the insulinotropic properties of different amino acids or protein (hydrolysates) when co-ingested with carbohydrate have not been investigated. OBJECTIVE: The aim of this study was to define an amino acid and protein (hydrolysate) mixture with a maximal insulinotropic effect when co-ingested with carbohydrate. DESIGN: Eight healthy, nonobese male subjects visited our laboratory, after an overnight fast, on 10 occasions on which different beverage compositions were tested for 2 h. During those trials the subjects ingested 0.8 g*kg(-)(1)*h(-)(1) carbohydrate and 0.4 g*kg(-)(1)*h(-)(1) of an amino acid and protein (hydrolysate) mixture. RESULTS: A strong initial increase in plasma glucose and insulin concentrations was observed in all trials, after which large differences in insulin response between drinks became apparent. After we expressed the insulin response as area under the curve during the second hour, ingestion of the drinks containing free leucine, phenylalanine, and arginine and the drinks with free leucine, phenylalanine, and wheat protein hydrolysate were followed by the largest insulin response (101% and 103% greater, respectively, than with the carbohydrate-only drink; P < 0.05). CONCLUSIONS: Insulin responses are positively correlated with plasma leucine, phenylalanine, and tyrosine concentrations. A mixture of wheat protein hydrolysate, free leucine, phenylalanine, and carbohydrate can be applied as a nutritional supplement to strongly elevate insulin concentrations. PMID- 10871569 TI - Human adult amino acid requirements: [1-13C]leucine balance evaluation of the efficiency of utilization and apparent requirements for wheat protein and lysine compared with those for milk protein in healthy adults. AB - BACKGROUND: There is considerable debate about the human lysine requirement and the consequent nutritional value of wheat protein. OBJECTIVE: We used a novel [1 (13)C]leucine balance protocol to examine whether adaptive mechanisms to conserve lysine allow wheat to be utilized more efficiently than expected according to current estimates of lysine requirements and wheat utilization. DESIGN: Wheat and milk proteins were compared in 6 adults infused for 9 h with L-[1-(13)C]leucine in the postabsorptive state (0-3 h), who were fed half-hourly with low-protein (2% of energy, 3-6 h) and isoenergetic higher-protein (12-13% of energy, 6-9 h) meals providing maintenance energy intakes. From acute measurements of [1 (13)C]leucine balance, we predicted nitrogen balance, the metabolic demand for protein, the efficiency of postprandial protein utilization (PPU), and the requirements for wheat protein and lysine. RESULTS: Leucine balance was higher after the milk than after the wheat feeding because of the greater inhibition of proteolysis by milk. PPU, calculated as the ratio of Deltanitrogen balance to Deltanitrogen intake between the low-protein and higher-protein periods, was 0.68 +/- 0.06 for wheat and 1.00 +/- 0.09 for milk (P +/- SD) was 27. 9 +/- 8.8 and 27.3 +/- 17.6 mg lysine * kg(-)(1) * d( )(1) for the low- and intermediate-intake groups, respectively (NS). Daily lysine balance was -12.4 +/- 92 and 1.8 +/- 17.7 mg * kg(-)(1) * d(-)(1), respectively (P < 0.025), for the low and intermediate intakes. The balance was significantly less than zero (P < 0.001) for the low intake. CONCLUSION: The FAO/WHO/UNU lysine requirement value is not sufficient to maintain lysine homeostasis in healthy adults. From the results of this and tracer studies done by others, the mean lysine requirement of healthy adults was determined to be 30 mg * kg(-)(1) * d( )(1). PMID- 10871571 TI - Concurrent physical activity increases fat oxidation during the shift to a high fat diet. AB - BACKGROUND: It takes several days to adapt to a high-fat diet. In an earlier study, we observed a large degree of interindividual variation in the capacity to adapt to a high-fat diet. We hypothesized that concurrent physical activity would accelerate fat oxidation during an isoenergetic high-fat diet. OBJECTIVE: The objective of this study was to determine the effect of increased physical activity on the ability of young healthy men to increase fat oxidation during the shift to a high-fat diet. DESIGN: Six young healthy men participated in a randomized, single-blind crossover study. The volunteers consumed a diet contributing 37% of energy as fat, 14% as protein, and 49% as carbohydrate for 4 d. Energy expenditure and macronutrient balance were then measured in a respiration chamber as the energy content of the isoenergetic diet was changed to 50% fat, 14% protein, and 36% carbohydrate. Treadmill walking, as the physical activity, was used to increase total daily energy expenditure to 1.8 times the resting metabolic rate during 1 of 2 stays in the metabolic chamber. Total daily energy expenditure was maintained at 1.4 times the resting metabolic rate for the other stay. RESULTS: Energy balance was not significantly different between the 2 conditions. The 24-h respiratory quotient decreased more rapidly and to a greater extent under conditions of increased energy expenditure. Further, there was a decrease in the interindividual variability in the response of the respiratory quotient to a high-fat diet with increased energy expenditure (physical activity). Cumulative carbohydrate and protein balances were greater under conditions of increased physical activity. Conversely, cumulative fat balance was greater under sedentary conditions. CONCLUSION: Concurrent physical activity increases fat oxidation during the shift to a high-fat diet. PMID- 10871572 TI - Vitamin C status and mortality in US adults. AB - BACKGROUND: Low vitamin C status may increase the risk of mortality from cancer and cardiovascular disease. OBJECTIVE: The objective was to test whether an association existed between serum ascorbate concentrations and mortality and whether the association was modified by cigarette smoking status or sex. DESIGN: Serum ascorbate concentrations were measured in adults as part of the second National Health and Nutrition Examination Survey (1976-1980). Vital status was ascertained 12-16 y later. RESULTS: The relative risk (RR) of death, adjusted for potential confounders, was estimated by using Cox proportional hazards models. Men in the lowest (<28.4 micromol/L) compared with the highest (>/=73.8 micromol/L) serum ascorbate quartile had a 57% higher risk of dying from any cause (RR: 1.57; 95% CI: 1.21, 2.03) and a 62% higher risk of dying from cancer (RR: 1.62; 95% CI: 1.01, 2.59). In contrast, there was no increased risk among men in the middle 2 quartiles for these outcomes and no increased risk of cardiovascular disease mortality in any quartile. There was no association between serum ascorbate quartile and mortality among women. These findings were consistent when analyses were limited to nonsmokers or further to adults who never smoked, suggesting that the observed relations were not due to cigarette smoking. CONCLUSIONS: These data suggest that men with low serum ascorbate concentrations may have an increased risk of mortality, probably because of an increased risk of dying from cancer. In contrast, serum ascorbate concentrations were not related to mortality among women. PMID- 10871573 TI - Hyporetinolemia and acute phase proteins in children with and without xerophthalmia. AB - BACKGROUND: The relations among hyporetinolemia, acute phase proteins, and vitamin A status in children are unclear. OBJECTIVE: The objective was to examine the relations between acute phase proteins and plasma retinol concentrations in children with and without clinical vitamin A deficiency (Bitot spots and night blindness). DESIGN: The study was a nonconcurrent analysis of acute phase protein concentrations and other data from a previous clinical trial. Preschool children, 3-6 y of age, with (n = 118) and without (n = 118) xerophthalmia were assigned to receive oral vitamin A (60 mg retinol equivalent) or placebo and were seen at 5 wk. All children received oral vitamin A (60 mg retinol equivalent) at 5 wk. RESULTS: At baseline, alpha(1)-acid glycoprotein (AGP) was elevated in 42.9% and 23.5% (P < 0.003) and C-reactive protein (CRP) was elevated in 17.7% and 13.7% (NS) of children with and without xerophthalmia, respectively. Hyporetinolemia (retinol < 0.7 micromol/L) occurred in 61.0% and 47.4% (P < 0.04) of children with and without xerophthalmia, respectively. A history of fever, a history of cough, and nasal discharge noted on examination were each associated with elevated acute phase proteins. Vitamin A supplementation increased plasma retinol at 5 wk but had no significant effect on concentrations of acute phase proteins. CONCLUSIONS: Elevated acute phase protein concentrations and infectious disease morbidity are closely associated during vitamin A deficiency. PMID- 10871574 TI - Folate absorption in women with a history of neural tube defect-affected pregnancy. AB - BACKGROUND: The risk of neural tube defects (NTDs) is significantly reduced by supplemental folic acid. NTD risk may be associated with impaired absorption of polyglutamyl folate, the primary form of naturally occurring food folate, and of folic acid in supplements or fortified food. Stable-isotope methods provide the specificity needed to test this hypothesis. OBJECTIVE: We determined whether women who had an NTD-affected pregnancy had a reduced ability compared with control women to absorb polyglutamyl folate relative to folic acid. DESIGN: Healthy, nonpregnant women with a history of an NTD-affected pregnancy (cases; n = 11) and control women (n = 11) were administered an oral dose containing a mixture of [(2)H]pteroylpentaglutamate ([(2)H(2)]PteGlu(5); 233 nmol) and [(13)C]pteroylmonoglutamate ([(13)C(5)]PteGlu(1); 567 nmol) after a 30-d saturation protocol (2 mg unlabeled folic acid/d). Relative extents of absorption were evaluated by urinary excretion of (2)H(2)- and (13)C(5)-labeled folates 48 h postdose. RESULTS: During the first 24 h postdose, cases excreted less (f1.gif" BORDER="0"> +/- SD) [(2)H(2)]PteGlu(5) (21 +/- 12% compared with 37 +/- 19%; P = 0.01) and [(13)C(5)]PteGlu(1) (17 +/- 8% compared with 31 +/- 14%; P = 0.007) than did controls. No significant differences between cases and controls were detected in the percentage of [(2)H(2)]PteGlu(5) or [(13)C(5)]PteGlu(1) excreted during the second 24 h postdose or when the data were averaged over 48 h. However, excretion of the [(2)H(2)]folates tended to be lower in cases than in controls over the 48-h period (33 +/- 13% compared with 45 +/- 26%; P = 0.21). A similar trend (P = 0.29) for lower excretion of [(13)C(5)]folates in cases was also observed (31 +/- 16% compared with 39 +/- 17%). The ratio of urinary [(2)H(2)]folates to [(13)C(5)]folates did not differ significantly between cases and controls. CONCLUSION: These data suggest the need for a larger-scale study using stable-isotope methods to further investigate this hypothesis. PMID- 10871575 TI - Early infant feeding and growth status of US-born infants and children aged 4-71 mo: analyses from the third National Health and Nutrition Examination Survey, 1988-1994. AB - BACKGROUND: There is controversy over what growth references to use in evaluating breast-fed infants and concern about whether never-breast-fed infants are at risk of overweight in childhood. OBJECTIVE: The objective of this study was to determine whether infants who are exclusively breast-fed for 4 mo differ in average size from infants who are fed in other ways and whether such differences persist through age 5 y. DESIGN: Data from the third National Health and Nutrition Examination Survey (NHANES III) were linked to birth certificates of US born infants and children. Feeding groups were defined on the basis of feeding patterns over the first 4 mo of life: exclusively breast-fed for 4 mo, partially breast-fed, breast-fed for <4 mo, and never breast-fed. Growth status, indexed as internally derived z scores (SD units) for weight, length (height), weight-for length (height), midupper arm circumference, and triceps skinfold thickness, was compared among feeding groups. RESULTS: The final sample consisted of 5594 non Hispanic white, non-Hispanic black, and Mexican American infants and children aged 4-71 mo. Of these, 21% were exclusively breast-fed for 4 mo, 10% were partially breast-fed, 24% were breast-fed for <4 mo, and 45% were never breast fed. At 8-11 mo, infants who were exclusively breast-fed for4 mo had adjusted mean z scores for weight (-0.21; -0.2 kg), weight-for-length (-0.27), and midupper arm circumference (-0.15) that differed significantly from zero (P < 0. 05). By 12-23 mo, the differences had dissipated; there were no significant differences subsequent to 5 y. Triceps skinfold thickness was not related to early infant feeding. CONCLUSION: Infants who were exclusively breast-fed for 4 mo weighed less at 8-11 mo than did infants who were fed in other ways, but there were few other significant differences in growth status through age 5 y associated with early infant feeding. PMID- 10871576 TI - A threshold for low-protein-diet-induced elevations in parathyroid hormone. AB - BACKGROUND: We reported previously that lowering dietary protein intake in young healthy women to 0.7 g/kg depressed intestinal calcium absorption and was accompanied by elevations in parathyroid hormone (PTH). Moderate amounts of dietary protein (1.0 g/kg) did not appear to perturb calcium homeostasis. OBJECTIVE: The purpose of this study was to evaluate the effect of graded intakes of dietary protein (0.7, 0.8, 0.9, and 1.0 g/kg) on calcium homeostasis. DESIGN: The experiment consisted of 2 wk of a well-balanced diet containing moderate amounts of calcium, sodium, and protein followed by 4 d of an experimental diet containing 1 of 4 amounts of protein. Eight young healthy women received the 4 amounts of protein in random order. The average age of the subjects was 23.1 +/- 2.3 y, their weight was 64 +/- 3 kg, and their body mass index (in kg/m(2)) was 24.3 +/- 0.9. RESULTS: Elevations in PTH developed by day 4 of the diets containing 0.7 and 0.8 g protein/kg but not during the diets containing 0.9 or 1.0 g protein/kg. By day 4 of the 0.7- and 0. 8-g/kg diets, midmolecule PTH, calcitriol, and nephrogenous cyclic adenosine monophosphate were 1.5-3.5-fold higher than on day 0. Calcitropic hormones on day 4 of the diets containing 0.8 and 0.9 g protein/kg were within the normal range and 23-57% lower than values observed with the 0.7- and 0.8-g/kg diets (P < 0.005). Mean 24-h urinary calcium was 3.29 +/- 0.35 mmol with the diet containing 0.7 g protein/kg and 3.54 +/- 0.46 mmol with the diet containing 1.0 g protein/kg. CONCLUSIONS: Our data suggest that in young healthy women consuming a well-balanced diet, the current recommended dietary allowance for protein (0.8 g/kg) results in short-term perturbations in calcium homeostasis. PMID- 10871577 TI - Detection of impaired intestinal absorption of long-chain fatty acids: validation studies of a novel test in a rat model of fat malabsorption. AB - BACKGROUND: Classic fat balance studies detect fat malabsorption but do not discriminate between the potential causes of malabsorption, such as impaired intestinal lipolysis or reduced uptake of fatty acids. OBJECTIVE: We aimed to validate a novel test for the specific, sensitive detection of impaired intestinal uptake of long-chain unesterified fatty acids in an appropriate rat model of fat malabsorption. DESIGN: The absorption and appearance in plasma of [(13)C]palmitic acid were determined in control rats and in rats with fat malabsorption due either to chronic bile deficiency (permanent bile diversion) or to oral administration of the lipase inhibitor orlistat (200 mg/kg diet). [(13)C]Palmitic acid results were compared with the percentage absorption of ingested dietary fat determined by fat balance. RESULTS: Between 1 and 6 h after intraduodenal administration, plasma [(13)C]palmitate concentrations in control rats were 4-10-fold higher than in bile-deficient rats (P < 0.05) but were not significantly different between orlistat-supplemented rats and their controls. In control and bile-deficient rats, plasma [(13)C]palmitate concentrations allowed complete discrimination between normal (>92%) and reduced (<92%) fat absorption, whereas the percentage absorption of [(13)C]palmitate over 48 h appeared to be highly correlated with the percentage absorption of ingested dietary fat (r = 0.89, P < 0.001). CONCLUSIONS: The [(13)C]palmitic acid absorption test detects impaired intestinal absorption of long-chain fatty acids selectively and sensitively in a rat model of fat malabsorption due to bile deficiency. Our data strongly support the use of the [(13)C]palmitic acid absorption test for the diagnosis of clinical fat malabsorption syndromes. PMID- 10871578 TI - Plasma antioxidant status after high-dose chemotherapy: a randomized trial of parenteral nutrition in bone marrow transplantation patients. AB - BACKGROUND: Chemotherapy and radiation therapy result in increased free radical formation and depletion of tissue antioxidants. It is not known whether parenteral nutrition (PN) administered during bone marrow transplantation (BMT) supports systemic antioxidant status. OBJECTIVE: The aims of the study were to determine 1) whether high-dose chemotherapy decreases concentrations of major circulating antioxidants in patients undergoing BMT and 2) whether administration of standard PN maintains systemic antioxidant concentrations compared with PN containing micronutrients and minimal lipids alone. DESIGN: Twenty-four BMT patients were randomly assigned to receive either standard PN containing conventional amounts of dextrose, amino acids, micronutrients, and lipid (120 kJ/d) or a solution containing only micronutrients (identical to those in standard PN) and a small amount of lipid (12 kJ/d). Plasma antioxidant status was measured before conditioning therapy and serially at days 1, 3, 7, 10, and 14 after BMT. RESULTS: Plasma glutathione (GSH) and alpha- and gamma-tocopherol concentrations decreased and the GSH redox state became more oxidized after conditioning chemotherapy. Plasma cysteine concentrations were unchanged, whereas cystine concentrations increased. Plasma vitamin C and zinc concentrations and GSH peroxidase activity increased over time. Plasma alpha-tocopherol concentrations were lower in patients given standard PN. There were no differences in other plasma antioxidants between groups. CONCLUSIONS: A significant decline in GSH-glutathione disulfide, cysteine-cystine, and vitamin E status occurs after chemotherapy and BMT. Standard PN does not improve antioxidant status compared with administration of micronutrients alone. Further evaluation of PN formulations to support patients undergoing high-dose chemotherapy and BMT are needed. PMID- 10871579 TI - Effects of chronic alcohol treatment on the synthesis, sialylation, and disposition of nascent apolipoprotein E by peritoneal macrophages of rats. AB - BACKGROUND: Plasma apolipoprotein (apo) E, a sialoprotein, plays an important role in reverse cholesterol transport. Previously, we showed that chronic alcohol consumption impairs glycosylation of apo E in rat liver. Peritoneal macrophages are another significant apo E synthesis site. OBJECTIVE: The main purpose of this study was to determine the effects of chronic alcohol feeding of rats on the synthesis, sialylation, and sialic acid content of macrophage apo E and its ability to bind to the HDL(3) molecule in vitro. DESIGN: Rats were fed an alcoholic diet or an isoenergetic control diet for 8 wk, after which peritoneal macrophages isolated from them were cultured and analyzed for apo E metabolism. RESULTS: Macrophages from alcohol-fed rats accumulated 33.3% more (P < 0.05) cholesterol than did those from control rats when incubated with acetylated LDL. These macrophages showed a 51-57% lower relative sialylation rate of apo E (P < 0.001) but no significant difference in relative protein synthetic rate. The sialic acid content of the intracellular and secreted forms of apo E was reduced by 41.8% (P < 0.001) and 50.3% (P < 0.001), respectively, with chronic alcohol treatment. Secretion of newly synthesized apo E was impaired by 53.7% (P < 0.001) and 26. 1% (P < 0.001) in the absence and presence of HDL in the medium, respectively. Macrophages of alcohol-treated rats secreted apo E with 47.6-67.2% lower (P < 0.001) HDL(3) binding ability; binding ability was restored completely by resialylation of the desialylated apo E. CONCLUSION: In rats, an alcohol mediated decrease in sialylation rate resulting in loss of sialic acid residues in apo E impairs the ability of apo E to bind to HDL and consequently in defective reverse cholesterol transport. PMID- 10871580 TI - Salt consumption during the nutrition transition in South Korea. PMID- 10871582 TI - On the biological activity of vitamin E. PMID- 10871587 TI - Introduction PMID- 10871584 TI - Is fat intake important in the public health control of obesity? PMID- 10871588 TI - Nutrition and maternal mortality in the developing world. AB - This review relates nutritional status to pregnancy-related death in the developing world, where maternal mortality rates are typically >/=100-fold higher than rates in the industrialized countries. For 3 of the central causes of maternal mortality (ie, induced abortion, puerperal infection, and pregnancy induced hypertension), knowledge of the contribution of nutrition is too scanty for programmatic application. Hemorrhage (including, for this discussion, anemia) and obstructed labor are different. The risk of death is greatly increased with severe anemia (Hb <70 or 80 g/L); there is little evidence of increased risk associated with mild or moderate anemia. Current programs of universal iron supplementation are unlikely to have much effect on severe anemia. There is an urgent need to reassess how to approach anemia control in pregnant women. Obstructed labor is far more common in short women. Unfortunately, nutritional strategies for increasing adult stature are nearly nonexistent: supplemental feeding appears to have little benefit after 3 y of age and could possibly be harmful at later ages, inducing accelerated growth before puberty, earlier menarche (and possible earlier marriage), and unchanged adult stature. Deprived girls without intervention typically have late menarche, extended periods of growth, and can achieve nearly complete catch-up growth. The need for operative delivery also increases with increased fetal size. Supplementary feeding could therefore increase the risk of obstructed labor. In the absence of accessible obstetric services, primiparous women <1.5 m in height should be excluded from supplementary feeding programs aimed at accelerating fetal growth. The knowledge base to model the risks and benefits of increased fetal size does not exist. PMID- 10871589 TI - Maternal mortality in the past and its relevance to developing countries today. AB - High maternal mortality was a feature of the Western world from the mid-19th century, when reliable record keeping commenced, to the mid-1930s. During this time, maternal mortality rates tended to remain on a high plateau, although there was wide disparity between countries in the height of the plateau. From approximately 1937, maternal mortality rates began to decline everywhere, and within 20 y, the intercountry differences had almost disappeared. The decline in maternal mortality rates was so dramatic that current rates for developed countries are between one-fortieth and one-fiftieth of the rates that prevailed 60 y ago. In this paper, the reasons for the high mortality before 1937 and its decline since that date are discussed. It is suggested that the main determinant of maternal mortality was the overall standard of maternal care provided by birth attendants. Poverty and associated malnutrition played little part in determining the rate of maternal mortality. This view is supported by much evidence, including the fact that, unlike for infant mortality rates, maternal mortality rates tended to be higher in the upper than in the lower social classes. The potential relevance of these findings to developing countries is discussed. PMID- 10871590 TI - Etiology of anemia in pregnancy in south Malawi. AB - BACKGROUND: Anemia in pregnancy is a major public health problem in developing countries. In sub-Saharan Africa, such anemia is generally accepted as resulting from nutritional deficiencies, particularly iron deficiency. OBJECTIVE: We comprehensively assessed the full spectrum of nutritional and nonnutritional factors associated with pregnancy anemia. DESIGN: Iron, folate, vitamin B-12, and vitamin A were measured in serum in a cross-sectional study of 150 pregnant women in Blantyre, Malawi. Bone marrow aspirates were evaluated, peripheral blood films were examined for malaria parasites, stool and urine samples were examined for helminthic infection, and tests were done for genetic disorders and for HIV infection. C-reactive protein (CRP) concentrations and erythrocyte sedimentation rates were measured as markers of inflammation. RESULTS: Of the 150 anemic women, 23% were iron deficient with no evidence of folate, vitamin B-12, or vitamin A deficiencies; 32% were deficient in iron and one or more of the other micronutrients; 26% were not iron deficient but had evidence of one of the other micronutrient deficiencies, most often vitamin A; and 19% were not deficient in any of the micronutrients studied. CRP concentrations were notably high in 54% of the anemic women with no nutritional deficiencies and in 73.5% of the anemic women who were iron replete by bone marrow assessment. CONCLUSION: The role of chronic inflammation as a possible contributing factor to anemia in pregnancy has important implications for the clinical evaluation and treatment of women. PMID- 10871591 TI - Iron requirements in pregnancy and strategies to meet them. AB - Iron requirements are greater in pregnancy than in the nonpregnant state. Although iron requirements are reduced in the first trimester because of the absence of menstruation, they rise steadily thereafter; the total requirement of a 55-kg woman is approximately 1000 mg. Translated into daily needs, the requirement is approximately 0.8 mg Fe in the first trimester, between 4 and 5 mg in the second trimester, and >6 mg in the third trimester. Absorptive behavior changes accordingly: a reduction in iron absorption in the first trimester is followed by a progressive rise in absorption throughout the remainder of pregnancy. The amounts that can be absorbed from even an optimal diet, however, are less than the iron requirements in later pregnancy and a woman must enter pregnancy with iron stores of >/=300 mg if she is to meet her requirements fully. This is more than most women possess, especially in developing countries. Results of controlled studies indicate that the deficit can be met by supplementation, but inadequacies in health care delivery systems have limited the effectiveness of larger-scale interventions. Attempts to improve compliance include the use of a supplement of ferrous sulfate in a hydrocolloid matrix (gastric delivery system, or GDS) and the use of intermittent supplementation. Another approach is intermittent, preventive supplementation aimed at improving the iron status of all women of childbearing age. Like all supplementation strategies, however, this approach has the drawback of depending on delivery systems and good compliance. On a long-term basis, iron fortification offers the most cost-effective option for the future. PMID- 10871592 TI - Iron needs during pregnancy: do we need to rethink our targets? AB - This paper argues that current estimates of the need for absorbed iron, estimates of iron absorption, and hence estimates of iron requirements for pregnant women greatly depend on what is determined as the desirable or target hemoglobin concentration (goal). The existing goal appears to be based on the maximal hemoglobin concentration that can be achieved with iron supplementation of well nourished women; this is a situation that can be expected to minimize iron absorption efficiency. I am unaware of attempts to define hemoglobin or anemia goals based on functional criteria (health of infant or mother). The current approach may seriously overestimate iron need and discourage food-based programs; furthermore, it may declare operational iron supplementation programs to be failures when, in fact, many programs may be successful in preventing functional effects of iron deficiency anemia. This is illustrated with data from a completed comparative study of daily and weekly iron supplementation. The final plea is to set aside existing traditions and, instead, attempt to develop functional criteria for anemia and establish functional goals of hemoglobin concentrations to be achieved during pregnancy. PMID- 10871593 TI - Significance of an abnormally low or high hemoglobin concentration during pregnancy: special consideration of iron nutrition. AB - An association between moderate anemia and poor perinatal outcomes has been found through epidemiologic studies, although available evidence cannot establish this relation as causal. Anemia may not be a direct cause of poor pregnancy outcomes, except in the case of maternal mortality resulting directly from severe anemia due to hypoxia and heart failure. Preventing or treating anemia, whether moderate or severe, is desirable. Because iron deficiency is a common cause of maternal anemia, iron supplementation is a common practice to reduce the incidence of maternal anemia. Nevertheless, the effectiveness of large-scale supplementation programs needs to be improved operationally and, where multiple micronutrient deficiencies are common, supplementation beyond iron and folate can be considered. High hemoglobin concentrations are often mistaken as adequate iron status; however, high hemoglobin is independent of iron status and is often associated with poor health outcomes. Very high hemoglobin concentrations cause high blood viscosity, which results in both compromised oxygen delivery to tissues and cerebrovascular complications. Epidemiologic studies have also found an association between high maternal hemoglobin concentrations and an increased risk of poor pregnancy outcomes. Evidence does not suggest that this association is causal; it could be better attributed to hypertensive disorders of pregnancy and to preeclampsia. The pathophysiologic mechanism of these conditions during pregnancy can produce higher hemoglobin concentrations because of reduced normal plasma expansion and cause fetal stress because of reduced placental-fetal perfusion. Accordingly, higher than normal hemoglobin concentrations should be regarded as an indicator of possible pregnancy complications, not necessarily as a sign of adequate iron nutrition, because iron supplementation does not increase hemoglobin higher than the optimal concentration needed for oxygen delivery. PMID- 10871594 TI - Nutrition in pregnancy: mineral and vitamin supplements. AB - Pregnancy is associated with physiologic changes that result in increased plasma volume and red blood cells and decreased concentrations of circulating nutrient binding proteins and micronutrients. In many developing countries, these physiologic changes can be aggravated by undernutrition, leading to micronutrient deficiency states, such as anemia, that can have disastrous consequences for both mothers and newborn infants. Multiple micronutrients are often taken by pregnant women in developed countries, but their benefits are limited, except for prophylactic folic acid taken during the periconceptional period. Women in developing countries may benefit from multiple-micronutrient prophylaxis in pregnancy, but the underlying basis and rationale for changing from supplementation with iron and folate to supplementation with multiple micronutrients has not been debated in the context of existing program objectives. There is an urgent need for this discussion so that both program effectiveness and program efficacy can be improved. PMID- 10871595 TI - Nutrition and obstructed labor. AB - Obstructed labor is one of the most common preventable causes of maternal and perinatal morbidity and mortality in developing countries. Among the common causes are cephalopelvic disproportion, malpresentation, and malposition. Recognizing the causes of obstructed labor is important if the complications are to be prevented. Adequate prevention, however, can be achieved only through a multidisciplinary approach aimed in the short term at identifying high-risk cases and in the long term at improving nutrition. Early motherhood should be discouraged, and efforts are needed to improve nutrition during infancy, childhood, early adulthood, and pregnancy. Improving the access to and promoting the use of reproductive and contraceptive services will help reduce the prevalence of this complication. PMID- 10871596 TI - Role of nutrition in the prevention of toxemia. AB - Toxemia of pregnancy is called the disease of theories because, over decades of research, numerous causes have been proposed but none proved. Although many nutritional factors have been suggested as playing a causal role in the etiology of toxemia, mortality from this disease has not varied over time or between circumstances as one would expect a nutritional disease to do. This does not mean that there is no nutritional influence, but it does mean that the available evidence does not show that nutrition makes a major difference in maternal mortality from toxemia of pregnancy. PMID- 10871597 TI - Discussion on program implications PMID- 10871598 TI - Discussion on research needs and designs PMID- 10871599 TI - Stabilization of circular rpsT mRNA demonstrates the 5'-end dependence of RNase E action in vivo. AB - RNase E is the major intracellular endonuclease in Escherichia coli. Its ability to cleave susceptible substrates in vitro depends on both the cleavage site itself and the availability of an unstructured 5' terminus. To test whether RNase E activity is 5'-end-dependent in vivo in the presence of all the components of the RNA degradative machinery, a known substrate, the rpsT mRNA, has been embedded in a permuted group I intron to permit its efficient, precise circularization in E. coli. Circular rpsT mRNAs are 4-6-fold more stable in vivo than their linear counterparts. Even partial inactivation of RNase E activity further enhances this stability 6-fold. However, the stabilization of circular rpsT mRNAs depends strongly on their efficient translation. These results show unambiguously the importance of an accessible 5'-end in controlling mRNA stability in vivo and support a two-step ("looping") model for RNase E action in which the first step is end recognition and the second is actual cleavage. PMID- 10871600 TI - Protease inhibitor 10 inhibits tumor necrosis factor alpha -induced cell death. Evidence for the formation of intracellular high M(r) protease inhibitor 10 containing complexes. AB - Protease inhibitor 10 (PI10) is a member of the ovalbumin family of serine protease inhibitors (ov-serpin) that is expressed at elevated levels in patients with acute myeloid leukemia and chronic myelomonocytic leukemia. Based upon the ability of the related serpin plasminogen activator inhibitor 2 (PAI-2) to protect cells against tumor necrosis factor alpha (TNFalpha)-induced cell death, this study was initiated to investigate the potential cytoprotective activity of PI10. Two different expression systems (i.e. plasmids encoding either PI10 alone or PI10 fused to the tag: enhanced green fluorescent protein, EGFP) were utilized to stably transfect an eukaryotic model cell system (i.e. HeLa cells) that neither expresses PAI-2 nor PI10. The level of PI10 expression in the stable transfectants was found to correlate with their resistance to TNFalpha-induced cell death. Immunoprecipitation/immunoblotting experiments demonstrated that PI10 is able to form SDS-stable complexes (i.e. M(r) >100,000) with a cytosolic protein(s). Increased levels of the PI10-containing complexes can be detected by TNFalpha treatment by preventing intracellular degradative activities with the proteasome inhibitor N-carbobenzyloxy-leucine-leucine-norvalinal. PI10-containing complexes are dissociated with conditions known to separate classical protease serpin complexes (i.e., 1.5 m ammonium hydroxide in the presence of SDS). These data support a role for the regulation of intracellular protease activities by ov serpins. PMID- 10871601 TI - Cloning and functional characterization of a cation-Cl- cotransporter-interacting protein. AB - To date, the cation-Cl(-) cotransporter (CCC) family comprises two branches of homologous membrane proteins. One branch includes the Na(+)-K(+)-Cl(-) cotransporters (NKCCs) and the Na(+)-Cl(-) cotransporter, and the other branch includes the K(+)-Cl(-) cotransporters. Here, we have isolated the first member of a third CCC family branch. This member shares approximately 25% identity in amino acid sequence with each of the other known mammalian CCCs. The corresponding cDNA, obtained from a human heart library and initially termed WO(3.3), encodes a 914-residue polypeptide of 96.2 kDa (calculated mass). Sequence analyses predict a 12-transmembrane domain (tm) region, two N-linked glycosylation sites between tm(5) and tm(6), and a large intracellular carboxyl terminus containing protein kinase C phosphorylation sites. Northern blot analysis uncovers an approximately 3.7-kilobase pair transcript present in muscle, placenta, brain, and kidney. With regard to function, WO(3. 3) expressed either in HEK-293 cells or Xenopus laevis oocytes does not increase Rb(+)-, Na(+) , and Cl(-)-coupled transport during 5- or 6-h fluxes, respectively. In the oocyte, however, WO(3.3) specifically inhibits human NKCC1-mediated (86)Rb(+) flux. In addition, coimmunoprecipitation studies using lysates from WO(3. 3) transfected HEK-293 cells suggest a direct interaction of WO(3.3) with endogenous NKCC. Thus, we have cloned and characterized the first putative heterologous CCC interacting protein (CIP) known at present. CIP1 may be part of a novel family of proteins that modifies the activity or kinetics of CCCs through heterodimer formation. PMID- 10871602 TI - Structural basis of hematopoietic prostaglandin D synthase activity elucidated by site-directed mutagenesis. AB - Hematopoietic prostaglandin (PG) D synthase (PGDS) is the first identified vertebrate ortholog in the Sigma class of the glutathione S-transferase (GST) family and catalyzes both isomerization of PGH(2) to PGD(2) and conjugation of glutathione to 1-chloro-2, 4-dinitrobenzene. We introduced site-directed mutations of Tyr(8), Arg(14), Trp(104), Lys(112), Tyr(152), Cys(156), Lys(198), and Leu(199), which are presumed to participate in catalysis or PGH(2) substrate binding based on the crystallographic structure. Mutants were analyzed in terms of structure, GST and PGDS activities, and activation of the glutathione thiol group. Of all the mutants, only Y8F, W104I, K112E, and L199F showed minor but substantial differences in their far-UV circular dichroism spectra from the wild type enzyme. Y8F, R14K/E, and W104I were completely inactive. C156L/Y selectively lost only PGDS activity. K112E reduced GST activity slightly and PGDS activity markedly, whereas K198E caused a selective decrease in PGDS activity and K(m) for glutathione and PGH(2) in the PGDS reaction. No significant changes were observed in the catalytic activities of Y152F and L199F, although their K(m) for glutathione was increased. Using 5,5'-dithiobis(2-nitrobenzoic acid) as an SH selective agent, we found that only Y8F and R14E/K did not accelerate the reactivity of the glutathione thiol group under the low reactivity condition of pH 5.0. These results indicate that Lys(112), Cys(156), and Lys(198) are involved in the binding of PGH(2); Trp(104) is critical for structural integrity of the catalytic center for GST and PGDS activities; and Tyr(8) and Arg(14) are essential for activation of the thiol group of glutathione. PMID- 10871603 TI - Regulation of the Ca2+-sensitive domains of the maxi-K channel in the mouse myometrium during gestation. AB - Large conductance Ca(2+)-activated K(+) channels (maxi-K channels) are known to modulate uterine activity during gestation. Electrophysiological recordings demonstrate that myometrial maxi-K current is suppressed in term-pregnant compared to non-pregnant mice. We sought to determine whether maxi-K current suppression is due to reduction of maxi-K channel protein or differential expression of maxi-K channel isoforms that vary in their Ca(2+) and voltage sensitivities. Immunoblot analyses show an increase of maxi-K channel protein throughout gestation. Polymerase chain reaction of mouse myometrial cDNA identified four alternatively spliced sites within the maxi-K transcript and three within the Ca(2+)-sensitive "tail" domain. Ribonuclease protection analyses demonstrate that total channel transcript levels mimic protein expression; however transcript levels of alternatively spliced regions of regulatory domains that alter sensitivity to voltage and Ca(2+) differ in their gestational expression. An insert that increases the maxi-K channel sensitivity to voltage and Ca(2+) is present at steady levels throughout gestation, differing from total channel transcript regulation. The insert-less form of this transcript, which reduces the channel voltage and Ca(2+) sensitivity, is not detected until midterm pregnancy. These findings verify that multiple isoforms of the maxi-K channel are present in the mouse myometrium and are regulated differentially during gestation, which is a likely mechanism for modulation of myometrial excitability during pregnancy. PMID- 10871604 TI - Distinct self-oligomerization activities of synaptotagmin family. Unique calcium dependent oligomerization properties of synaptotagmin VII. AB - Synaptotagmins constitute a large protein family, characterized by one transmembrane region and two C2 domains, and can be classified into several subclasses based on phylogenetic relationships and biochemical activities (Fukuda, M., Kanno, E., and Mikoshiba, K. (1999) J. Biol. Chem. 274, 31421 31427). Synaptotagmin I (Syt I), a possible Ca(2+) sensor for neurotransmitter release, showed both Ca(2+)-dependent (via the C2 domain) and -independent (via the NH(2)-terminal domain) self-oligomerization, which are thought to be important for synaptic vesicle exocytosis. However, little is known about the relationship between these two interactions and the Ca(2+)-dependent oligomerization properties of other synaptotagmin isoforms. In this study, we first examined the Ca(2+)-dependent self-oligomerization properties of synaptotagmin family by co-expression of T7- and FLAG-tagged Syts (full-length or cytoplasmic domain) in COS-7 cells. We found that Syt VII is a unique class of synaptotagmins that only showed robust Ca(2+)-dependent self-oligomerization at the cytoplasmic domain with EC(50) values of about 150 micrometer Ca(2+). In addition, Syt VII preferentially interacted with the previously described subclass of Syts (V, VI, and X) in a Ca(2+)-dependent manner. Co-expression of full-length and cytoplasmic portion of Syts VII (or II) indicate that Syt VII cytoplasmic domain oligomerizes in a Ca(2+)-dependent manner without being tethered at the NH(2)-terminal domain, whereas Ca(2+)-dependent self oligomerization at the cytoplasmic domain of other isoforms (e.g. Syt II) occurs only when the two molecules are tethered at the NH(2)-terminal domain. PMID- 10871605 TI - Evidence of a role for SHP-1 in platelet activation by the collagen receptor glycoprotein VI. AB - The Src homology (SH)2 domain-containing protein-tyrosine phosphatase SHP-1 is tyrosine phosphorylated in platelets in response to the glycoprotein VI (GPVI) selective agonist collagen-related peptide (CRP), collagen, and thrombin. Two major unidentified tyrosine-phosphorylated bands of 28 and 32 kDa and a minor band of 130 kDa coprecipitate with SHP-1 in response to all three agonists. Additionally, tyrosine-phosphorylated proteins of 50-55 and 70 kDa specifically associate with SHP-1 following stimulation by CRP and collagen. The tyrosine kinases Lyn, which exists as a 53 and 56-kDa doublet, and Syk were identified as major components of these bands, respectively. Kinase assays on SHP-1 immunoprecipitates performed in the presence of the Src family kinase inhibitor PP1 confirmed the presence of a Src kinase in CRP- but not thrombin-stimulated cells. Lyn, Syk, and SLP-76, along with tyrosine-phosphorylated 28-, 32-, and 130 kDa proteins, bound selectively to a glutathione S-transferase protein encoding the SH2 domains of SHP-1, suggesting that this is the major site of interaction. Platelets isolated from motheaten viable mice (mev/mev) revealed the presence of a heavily tyrosine-phosphorylated 26-kDa protein that was not found in wild-type platelets. CRP-stimulated mev/mev platelets manifested hypophosphorylation of Syk and Lyn and reduced P-selectin expression relative to controls. These observations provide evidence of a functional role for SHP-1 in platelet activation by GPVI. PMID- 10871606 TI - Mitogen-induced expression of the fibroblast growth factor-binding protein is transcriptionally repressed through a non-canonical E-box element. AB - The fibroblast growth factor-binding protein (FGF-BP) stimulates FGF-2-mediated angiogenesis and is thought to play an important role in the progression of squamous cell, colon, and breast carcinomas. 12-O-Tetradecanoylphorbol-13-acetate (TPA) induction of the FGF-BP gene occurs through transcriptional mechanisms involving Sp1, AP-1, and CCAATT/enhancer-binding protein sites in the proximal FGF-BP gene promoter. The level of TPA induction, however, is limited due to the presence of a repressor element that shows similarity to a non-canonical E-box (AACGTG). Mutation or deletion of the repressor element led to enhanced induction by TPA or epidermal growth factor in cervical squamous cell and breast carcinoma cell lines. Repression was dependent on the adjacent AP-1 site, without discernible alteration in the binding affinity or composition of AP-1. We investigated the following two possible mechanisms for E-box-mediated repression: 1) CpG methylation of the core of the E-box element, and 2) binding of a distinct protein complex to this site. Point mutation of the CpG methylation site in the E box showed loss of repressor activity. Conversely, in vitro methylation of this site significantly reduced TPA induction. In vitro gel shift analysis revealed distinct and TPA-dependent binding of USF1 and USF2 to the repressor element that required nucleotides within the E-box. Furthermore, chromatin immunoprecipitation assay showed that USF, c-Myc, and Max proteins were associated with the FGF-BP promoter in vivo. Overall, these findings suggested that the balance between trans-activation by AP-1 and repression through the E-box is an important control mechanism for fine-tuning the angiogenic response to growth factor-activated pathways. PMID- 10871607 TI - The RAD51 family member, RAD51L3, is a DNA-stimulated ATPase that forms a complex with XRCC2. AB - The Rad51 protein in eukaryotic cells is a structural and functional homolog of Escherichia coli RecA with a role in DNA repair and genetic recombination. Several proteins showing sequence similarity to Rad51 have previously been identified in both yeast and human cells. In Saccharomyces cerevisiae, two of these proteins, Rad55p and Rad57p, form a heterodimer that can stimulate Rad51 mediated DNA strand exchange. Here, we report the purification of one of the representatives of the RAD51 family in human cells. We demonstrate that the purified RAD51L3 protein possesses single-stranded DNA binding activity and DNA stimulated ATPase activity, consistent with the presence of "Walker box" motifs in the deduced RAD51L3 sequence. We have identified a protein complex in human cells containing RAD51L3 and a second RAD51 family member, XRCC2. By using purified proteins, we demonstrate that the interaction between RAD51L3 and XRCC2 is direct. Given the requirements for XRCC2 in genetic recombination and protection against DNA-damaging agents, we suggest that the complex of RAD51L3 and XRCC2 is likely to be important for these functions in human cells. PMID- 10871608 TI - Global gene expression profiling in Escherichia coli K12. The effects of integration host factor. AB - We have used nylon membranes spotted in duplicate with full-length polymerase chain reaction-generated products of each of the 4,290 predicted Escherichia coli K12 open reading frames (ORFs) to measure the gene expression profiles in otherwise isogenic integration host factor IHF(+) and IHF(-) strains. Our results demonstrate that random hexamer rather than 3' ORF-specific priming of cDNA probe synthesis is required for accurate measurement of gene expression levels in bacteria. This is explained by the fact that the currently available set of 4,290 unique 3' ORF-specific primers do not hybridize to each ORF with equal efficiency and by the fact that widely differing degradation rates (steady-state levels) are observed for the 25-base pair region of each message complementary to each ORF specific primer. To evaluate the DNA microarray data reported here, we used a linear analysis of variance (ANOVA) model appropriate for our experimental design. These statistical methods allowed us to identify and appropriately correct for experimental variables that affect the reproducibility and accuracy of DNA microarray measurements and allowed us to determine the statistical significance of gene expression differences between our IHF(+) and IHF(-) strains. Our results demonstrate that small differences in gene expression levels can be accurately measured and that the significance of differential gene expression measurements cannot be assessed simply by the magnitude of the fold difference. Our statistical criteria, supported by excellent agreement between previously determined effects of IHF on gene expression and the results reported here, have allowed us to identify new genes regulated by IHF with a high degree of confidence. PMID- 10871609 TI - Hyaluronan binding by cell surface CD44. AB - CD44 is the primary cell surface receptor for the extracellular matrix glycosaminoglycan hyaluronan. Here we determined the relative avidities of unlabeled hyaluronan preparations for cell surface CD44 by their ability to block the binding of fluorescein-conjugated hyaluronan to a variety of cells. We show that hyaluronan binding at the cell surface is a complex interplay of multivalent binding events affected by the size of the multivalent hyaluronan ligand, the quantity and density of cell surface CD44, and the activation state of CD44 as determined by cell-specific factors and/or treatment with CD44-specific monoclonal antibody (mAb). Using low M(r) hyaluronan oligomers of defined sizes, we observed monovalent binding between 6 and 18 sugars. At approximately 20 to approximately 38 sugars, there was an increase in avidity (approximately 3x), suggesting that divalent binding was occurring. In the presence of the inducing mAb IRAWB14, monovalent binding avidity was similar to that of noninduced CD44, but beginning at approximately 20 residues, there was a dramatic and progressive increase in avidity with increasing oligomer size ( approximately 22 < 26 < 30 < 34 < 38 sugars). Kinetic studies of binding and dissociation of fluorescein conjugated hyaluronan indicated that inducing mAb treatment had little effect on the binding kinetics, but dissociation from the cell surface was greatly delayed by inducing mAb. PMID- 10871610 TI - Increased thermal resistance and modification of the catalytic properties of a beta-glucosidase by random mutagenesis and in vitro recombination. AB - The bglB gene from Paenibacillus polymyxa was subjected to random mutagenesis mediated by error prone polymerase chain reaction amplification and DNA shuffling. After this treatment, mutant variants of the encoded beta-glucosidase with enhanced thermal resistance were selected. We identified five amino acid substitutions at four different positions of the sequence that increased the resistance of the enzyme to heat denaturation. Four of the mutations, H62R, M319V, M319I, and M361I, did not change the kinetic parameters of the enzyme. However, mutant N223Y, which caused only a marginal increase in thermoresistance, showed an 8-fold decrease in K(m). Copies of the bglB gene carrying each one of the individual mutations were recombined in vitro by DNA shuffling. As a result, we obtained an enzyme that simultaneously exhibited a 20-fold increase in heat resistance and an 8-fold increase in the catalytic efficiency. The structural basis of the properties conferred by the mutations was analyzed using homology based structural models. The four mutations causing a more pronounced effect on thermoresistance were located in loops, on the periphery of the (alpha/beta)(8) barrel that conforms the structure of the protein. Mutation N223Y, which modifies the catalytic properties of the enzyme, was on one of the barrel beta-strands that shape the active center. PMID- 10871611 TI - Structural requirements for catalysis and membrane targeting of mammalian enzymes with neutral sphingomyelinase and lysophospholipid phospholipase C activities. Analysis by chemical modification and site-directed mutagenesis. AB - The sequence similarity with bacterial neutral sphingomyelinase resulted in the isolation of putative mammalian counterparts and, subsequently, identification of similar molecules in a number of other eukaryotic organisms. Based on sequence similarities and previous characterization of the mammalian enzymes, we have chemically modified specific residues and performed site-directed mutagenesis in order to identify critical catalytic residues and determinants for membrane localization. Modification of histidine residues and the substrate protection experiments demonstrated the presence of reactive histidine residues within the active site. Site directed mutagenesis suggested an essential role in catalysis for two histidine residues (His-136 and His-272), which are conserved in all sequences. Mutations of two additional histidines (His-138 and His-151), conserved only in eukaryotes, resulted in reduced neutral sphingomyelinase activity. In addition to sphingomyelin, the enzyme also hydrolyzed lysophosphatidylcholine. Exposure to an oxidizing environment or modification of cysteine residues using several specific compounds also inactivated the enzyme. Site-directed mutagenesis of eight cysteine residues and gel-shift analysis demonstrated that these residues did not participate in the catalytic reaction and suggested the involvement of cysteines in the formation/breakage of disulfide bonds, which could underlie the reversible inactivation by the oxidizing compounds. Cellular localization studies of a series of deletion mutants, expressed as green fluorescent protein fusion proteins, demonstrated that the transmembrane region contains determinants for the endoplasmic reticulum localization. PMID- 10871612 TI - Interleukins 4 and 13 increase intestinal epithelial permeability by a phosphatidylinositol 3-kinase pathway. Lack of evidence for STAT 6 involvement. AB - Interleukins 4 and 13 can affect their target cells by activation of signal transducer and activator of transcription 6 (STAT 6) or phosphatidylinositol 3 kinase (PI3K). We examined the signal transduction events involved in IL-4 and IL 13 regulation of epithelial paracellular permeability using T84 cells, a model human colonic epithelium. T84 cells treated with IL-4 or IL-13 displayed virtually identical dose- and time-dependent STAT 6 activation as assessed by electrophoretic mobility shift assay (EMSA) and decreases in transepithelial resistance (TER). STAT 6 DNA binding activity was maximal in nuclear extracts 30 min after exposure to IL-4 or IL-13, and TER was maximally reduced by 24 h post treatment. Pretreatment of epithelia with transcription factor decoys (phosphorothioated DNA oligonucleotides containing the STAT 6 binding site) dramatically reduced STAT 6 activation as detected by EMSA, but did not attenuate the TER reduction by IL-4 or IL-13. In contrast, although the PI3K inhibitors wortmannin and LY294002 did not affect IL-4 or IL-13 STAT 6 activation, they significantly inhibited the ability of either cytokine to lower TER. Thus, we provide evidence for PI3K as the major proximal signaling event in IL-4 and IL-13 regulation of TER and speculate that pharmacological targeting of enterocytic PI3K activity may represent a means to manipulate epithelial permeability. PMID- 10871613 TI - Complex structure and regulation of expression of the rat gene for inward rectifier potassium channel Kir7.1. AB - Genomic organization of the rat inward rectifier K(+) channel Kir7.1 was determined in an attempt to clarify how multiple species of its mRNA are generated in a tissue-specific manner and how its expression is regulated. The rat Kir7.1 gene spans >40 kilobases (kb) and consists of eight exons; the first four exons encode the 5'-untranslated region that is unusually long ( approximately 3 kb). The coding region is located in exons 5 and 6. In the testis, exon 4 is processed as four exons (4a-4d), whereas it is recognized as a single exon in the small intestine. The three major species of rat Kir7.1 mRNA (1.4, 2.2, and 3.2 kb) were found to arise from alternative usage of the two promoters and polyadenylation signals and by alternative splicing of the 5' noncoding exons. The splicing pattern of the 5'-noncoding exons is quite complex and highly tissue-specific, suggesting that complex mechanisms may operate to regulate the Kir7.1 expression. Deletion and mutational analysis of the promoter activity indicated that the rat Kir7.1 gene is regulated by cAMP through a CCAAT element. The cAMP induction was also demonstrated using the rat follicular cell line FRTL-5 endogenously expressing Kir7.1. PMID- 10871614 TI - D-Alanine substitution of teichoic acids as a modulator of protein folding and stability at the cytoplasmic membrane/cell wall interface of Bacillus subtilis. AB - The extracytoplasmic folding of secreted proteins in Gram-positive bacteria is influenced by the microenvironment of the compartment into which they are translocated, namely the negatively charged matrix of the cell wall polymers. In this compartment, the PrsA lipoprotein facilitates correct post-translocational folding or prevents misfolding of secreted proteins. In this study, a secretion mutant of B. subtilis (prsA3) encoding a defective PrsA protein was mutagenized and screened for restored secretion of the AmyQ alpha-amylase. One mini-Tn10 insertion, which partially suppressed the secretion deficiency, was found to interrupt dlt, the operon involved in the d-alanylation of teichoic acids. The inactivation of dlt rescued the mutant PrsA3 protein from degradation, and the increased amount of PrsA3 was shown to enhance the secretion of PrsA-dependent proteins. Heterologous or abnormal secreted proteins, which are prone to degradation after translocation, were also stabilized and secreted in increased quantities from a dlt prsA(+) strain. Furthermore, the dlt mutation partially suppressed the lethal effect of PrsA depletion, suggesting that the dlt deficiency also leads to stabilization of an essential cell wall protein(s). Our results suggest that main influence of the increased net negative charge of the wall caused by the absence of d-alanine is to increase the rate of post translocational folding of exported proteins. PMID- 10871615 TI - Interaction of the bacteriophage T4 gene 59 helicase loading protein and gene 41 helicase with each other and with fork, flap, and cruciform DNA. AB - Bacteriophage T4 gene 59 helicase loading protein accelerates the loading of T4 gene 41 DNA helicase and is required for recombination-dependent DNA replication late in T4 phage infection. The crystal structure of 59 protein revealed a two domain alpha-helical protein, whose N-terminal domain has strong structural similarity to the DNA binding domain of high mobility group family proteins (Mueser, T. C., Jones, C. E., Nossal, N. G., and Hyde, C. C. (2000) J. Mol. Biol. 296, 597-612). We have previously shown that 59 protein binds preferentially to fork DNA. Here we show that 59 protein binds to completely duplex forks but cannot load the helicase unless there is a single-stranded gap of more than 5 nucleotides on the fork arm corresponding to the lagging strand template. Consistent with the roles of these proteins in recombination, we find that 59 protein binds to and stimulates 41 helicase activity on Holliday junction DNA, and on a substrate that resembles a strand invasion structure. 59 protein forms a stable complex with wild type 41 helicase and fork DNA in the presence of adenosine 5'-O-(thiotriphosphate). The unwinding activity of 41 helicase missing 20 C-terminal amino acids is not stimulated by 59 protein, and it does not form a complex with 59 protein on fork DNA. PMID- 10871616 TI - The binding of substrate analogs to phosphotriesterase. AB - Phosphotriesterase (PTE) from Pseudomonas diminuta catalyzes the detoxification of organophosphates such as the widely utilized insecticide paraoxon and the chemical warfare agent sarin. The three-dimensional structure of the enzyme is known from high resolution x-ray crystallographic analyses. Each subunit of the homodimer folds into a so-called TIM barrel, with eight strands of parallel beta sheet. The two zinc ions required for activity are positioned at the C-terminal portion of the beta-barrel. Here, we describe the three-dimensional structure of PTE complexed with the inhibitor diisopropyl methyl phosphonate, which serves as a mimic for sarin. Additionally, the structure of the enzyme complexed with triethyl phosphate is also presented. In the case of the PTE-diisopropyl methyl phosphonate complex, the phosphoryl oxygen of the inhibitor coordinates to the more solvent-exposed zinc ion (2.5 A), thereby lending support to the presumed catalytic mechanism involving metal coordination of the substrate. In the PTE triethyl phosphate complex, the phosphoryl oxygen of the inhibitor is positioned at 3.4 A from the more solvent-exposed zinc ion. The two structures described in this report provide additional molecular understanding for the ability of this remarkable enzyme to hydrolyze such a wide range of organophosphorus substrates. PMID- 10871617 TI - 31P NMR spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major. Evidence for high levels of condensed inorganic phosphates. AB - High resolution (31)P nuclear magnetic resonance spectra at 303.6 MHz (corresponding to a (1)H resonance frequency of 750 MHz) have been obtained of perchloric acid extracts of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major, the causative agents of African sleeping sickness, Chagas' disease, and leishmaniasis. Essentially complete assignments have been made based on chemical shifts and by direct addition of authentic reference compounds. The results indicate the presence of high levels of short chain condensed polyphosphates: di , tri-, tetra-, and pentapolyphosphate. (31)P NMR spectra of purified T. brucei, T. cruzi, and L. major acidocalcisomes, calcium and phosphorus storage organelles, indicate that polyphosphates are abundant in these organelles and have an average chain length of 3.11-3.39 phosphates. In the context of the recent discovery of several pyrophosphate-utilizing enzymes in trypanosomatids, the presence of these inorganic polyphosphates implies a critical role for these molecules in these parasites and a potential new route to chemotherapy. PMID- 10871618 TI - Bovine papillomavirus E1 protein is sumoylated by the host cell Ubc9 protein. AB - Papillomavirus E1 protein is the replication initiator that recognizes and binds to the viral origin and initiates DNA strand separation through its ATP-dependent helicase activity. The E1 protein also functions in viral DNA replication by recruiting several cellular proteins to the origin, including host DNA polymerase alpha and replication protein A. To identify other cellular proteins that interact with bovine papillomavirus E1, an HeLa cDNA library was screened using a yeast two-hybrid assay. The host cell sumoylating enzyme, Ubc9, was found to interact specifically with E1 both in vitro and in vivo. Mapping studies localized critical E1 sequences for interaction to amino acids 315-459 and strongly implicated leucine 420 as critical for E1.Ubc9 complex formation. In addition to binding E1, Ubc9 catalyzed the covalent linkage of the ubiquitin-like protein, SUMO-1, to E1. An E1 mutant unable to bind Ubc9 showed normal intracellular stability, but was impaired for intranuclear distribution. Failure to accumulate in appropriate nuclear subdomains may account for the previously demonstrated replication defect of a human papillomavirus 16 E1 protein that was also unable to bind Ubc9 and suggests that sumoylation is a functionally important modification with regulatory implications for papillomavirus replication. PMID- 10871619 TI - Identification of the (183)RWTNNFREY(191) region as a critical segment of matrix metalloproteinase 1 for the expression of collagenolytic activity. AB - Matrix metalloproteinase 1 (MMP-1) cleaves types I, II, and III collagen triple helices into (3/4) and (1/4) fragments. To understand the structural elements responsible for this activity, various lengths of MMP-1 segments have been introduced into MMP-3 (stromelysin 1) starting from the C-terminal end. MMP-3/MMP 1 chimeras and variants were overexpressed in Escherichia coli, folded from inclusion bodies, and isolated as zymogens. After activation, recombinant chimeras were tested for their ability to digest triple helical type I collagen at 25 degrees C. The results indicate that the nine residues (183)RWTNNFREY(191) located between the fifth beta-strand and the second alpha-helix in the catalytic domain of MMP-1 are critical for the expression of collagenolytic activity. Mutation of Tyr(191) of MMP-1 to Thr, the corresponding residue in MMP-3, reduced collagenolytic activity about 5-fold. Replacement of the nine residues with those of the MMP-3 sequence further decreased the activity 2-fold. Those variants exhibited significant changes in substrate specificity and activity against gelatin and synthetic substrates, further supporting the notion that this region plays a critical role in the expression of collagenolytic activity. However, introduction of this sequence into MMP-3 or a chimera consisting of the catalytic domain of MMP-3 with the hinge region and the C-terminal hemopexin domain of MMP 1 did not express any collagenolytic activity. It is therefore concluded that RWTNNFREY, together with the C-terminal hemopexin domain, is essential for collagenolytic activity but that additional structural elements in the catalytic domain are also required. These elements probably act in a concerted manner to cleave the collagen triple helix. PMID- 10871620 TI - Importance of stalk segment S5 for intramolecular communication in the sarcoplasmic reticulum Ca2+-ATPase. AB - Sixteen residues in stalk segment S5 of the Ca(2+)-ATPase of sarcoplasmic reticulum were studied by site-directed mutagenesis. The rate of the Ca(2+) binding transition, determined at 0 degrees C, was enhanced relative to wild type in mutants Ile(743) --> Ala, Val(747) --> Ala, Glu(748) --> Ala, Glu(749) --> Ala, Met(757) --> Gly, and Gln(759) --> Ala and reduced in mutants Asp(737) --> Ala, Asp(738) --> Ala, Ala(752) --> Leu, and Tyr(754) --> Ala. In mutant Arg(762) --> Ile, the rate of the Ca(2+) binding transition was wild type like at 0 degrees C, whereas it was 3.5-fold reduced relative to wild type at 25 degrees C. The rate of dephosphorylation of the ADP-insensitive phosphoenzyme was increased conspicuously in mutants Ile(743) --> Ala and Tyr(754) --> Ala (close to 20-fold in the absence of K(+)) and increased to a lesser extent in Asn(739) --> Ala, Glu(749) --> Ala, Gly(750) --> Ala, Ala(752) --> Gly, Met(757) --> Gly, and Arg(762) --> Ile, whereas it was reduced in mutants Asp(737) --> Ala, Val(744) - > Gly, Val(744) --> Ala, Val(747) --> Ala, and Ala(752) --> Leu. In mutants Ile(743) --> Ala, Tyr(754) --> Ala, and Arg(762) --> Ile, the apparent affinities for vanadate were enhanced 23-, 30-, and 18-fold, respectively, relative to wild type. The rate of Ca(2+) dissociation was 11-fold increased in Gly(750) --> Ala and 2-fold reduced in Val(747) --> Ala. Mutants with alterations to Arg(751) either were not expressed at a significant level or were completely nonfunctional. The findings show that S5 plays a crucial role in mediating communication between the Ca(2+) binding pocket and the catalytic domain and that Arg(751) is important for both structural and functional integrity of the enzyme. PMID- 10871621 TI - Regulation of the balance of one-carbon metabolism in Saccharomyces cerevisiae. AB - One-carbon metabolism in yeast is an essential process that relies on at least one of three one-carbon donor molecules: serine, glycine, or formate. By a combination of genetics and biochemistry we have shown how cells regulate the balance of one-carbon flow between the donors by regulating cytoplasmic serine hydroxymethyltransferase activity in a side reaction occurring in the presence of excess glycine. This control governs the level of 5,10-methylene tetrahydrofolate (5,10-CH(2)-H(4)folate) in the cytoplasm, which has a direct role in signaling transcriptional control of the expression of key genes, particularly those encoding the unique components of the glycine decarboxylase complex (GCV1, GCV2, and GCV3). Based on these and other observations, we propose a model for how cells balance the need to supplement their one-carbon pools when charged folates are limiting or when glycine is in excess. We also propose that under normal conditions, cytoplasmic 5,10-CH(2)-H(4)folate is mainly directed to generating methyl groups via methionine, whereas one-carbon units generated from glycine in mitochondria are more directed to purine biosynthesis. When glycine is in excess, 5, 10-CH(2)-H(4)folate is decreased, and the regulation loop shifts the balance of generation of one-carbon units into the mitochondrion. PMID- 10871623 TI - Establishment of lysogeny in bacteriophage 186. DNA binding and transcriptional activation by the CII protein. AB - The CII protein of bacteriophage 186 is a transcriptional activator of the helix turn helix family required for establishment of the lysogenic state. DNA binding by 186 CII is unusual in that the invertedly repeated half sites are separated by 20 base pairs, or two turns of the DNA helix, rather than the one turn usually associated with this class of proteins. Here, we investigate quantitatively the DNA binding properties of CII and its interaction with RNA polymerase at the establishment promoter, p(E). The stoichiometry of CII binding was determined by sedimentation equilibrium experiments using a fluorescein-labeled oligonucleotide and purified CII. These experiments indicate that the CII species bound to DNA is a dimer, with additional weak binding of a tetrameric species at high concentrations. Examination of the thermodynamic linkages between CII self association and DNA binding shows that CII binds to the DNA as a preformed dimer (binding free energy, 9.9 kcal/mol at 4 degrees C) rather than by association of monomers on the DNA. CII binding induces in the DNA a bend of 41 (+/- 5) degrees. The spacing between the binding half sites was shown to be important for CII binding, insertion or removal of just 1 base pair significantly reducing the affinity for CII. Removal of 5 or 10 base pairs between binding half sites eliminated binding, as did insertion of an additional 10 base pairs. CII binding at p(E) was improved marginally by the presence of RNA polymerase (DeltaDeltaG = 0.5 (+/- 0.3) kcal/mol). In contrast, the binding of RNA polymerase at p(E) was undetectable in the absence of CII but was improved markedly by the presence of CII. Thus, CII appears to recruit RNA polymerase to the promoter. The nature of the base pair changes in mutant phage, selected by their inability to establish lysogeny, are consistent with this mechanism of CII action. PMID- 10871622 TI - Stereoisomeric specificity of the retinoid cycle in the vertebrate retina. AB - Understanding of the stereospecificity of enzymatic reactions that regenerate the universal chromophore required to sustain vision in vertebrates, 11-cis-retinal, is needed for an accurate molecular model of retinoid transformations. In rod outer segments (ROS), the redox reaction involves all-trans-retinal and pro-S NADPH that results in the production of pro-R-all-trans-retinol. A recently identified all-trans-retinol dehydrogenase (photoreceptor retinol dehydrogenase) displays identical stereospecificity to that of the ROS enzyme(s). This result is unusual, because photoreceptor retinol dehydrogenase is a member of a short chain alcohol dehydrogenase family, which is often pro-S-specific toward their hydrophobic alcohol substrates. The second redox reaction occurring in retinal pigment epithelium, oxidation of 11-cis-retinol, which is largely catalyzed by abundantly expressed 11-cis-retinol dehydrogenase, is pro-S-specific to both 11 cis-retinol and NADH. However, there is notable presence of pro-R-specific activities. Therefore, multiple retinol dehydrogenases are involved in regeneration of 11-cis-retinal. Finally, the cellular retinaldehyde-binding protein-induced isomerization of all-trans-retinol to 11-cis-retinol proceeds with inversion of configuration at the C(15) carbon of retinol. Together, these results provide important additions to our understanding of retinoid transformations in the eye and a prelude for in vivo studies that ultimately may result in efficient pharmacological intervention to restore and prevent deterioration of vision in several inherited eye diseases. PMID- 10871624 TI - Distinct mRNAs that encode La autoantigen are differentially expressed and contain internal ribosome entry sites. AB - Analysis by reverse transcription-polymerase chain reaction has suggested the existence of at least two La autoantigen-encoding mRNAs that contain different 5' noncoding regions (NCRs) linked to the same La coding region (Troster, H., Metzger, T. E., Semsei, I., Schwemmle, M., Winterpacht, A., Zabel, B., and Bachmann, M. (1994) J. Exp. Med. 180, 2059-2067). La-encoding transcripts La1 and La1' contain 115- and 483-nucleotide 5' NCRs, respectively. To determine whether the various La transcripts are functional mRNAs, the expression and polysomal association of natural La1 and La1' RNAs were examined. Although La1 transcripts were ubiquitously expressed in human tissues, La1' transcripts were predominantly expressed in peripheral blood leukocytes, especially in B, T, and natural killer cells. Both La1 and La1' transcripts associated with polysomes in natural killer cells, suggesting that these transcripts were functional mRNAs. Upon activation of B cells with the mitogens phorbol 12-myristate 13-acetate and ionomycin, the amount of La1' mRNA, but not La1, declined. In contrast, after chemical activation of T cells, the amount of La 1 mRNA, but not La1', declined. The mechanism by which the La1 and La1' 5' NCRs initiate translation initiation was tested in cultured human HeLa cells and in two different in vitro translation systems. It was found that both 5' NCRs can mediate translation initiation by internal initiation. These findings indicate that the constitutive expression of La1 mRNA and the tissue-specific expression of La1' mRNA can both allow La protein synthesis under conditions when cap-dependent translation is compromised, such as inflammation, apoptosis, or certain viral infections. PMID- 10871625 TI - The C termini of constitutive nitric-oxide synthases control electron flow through the flavin and heme domains and affect modulation by calmodulin. AB - The sequences of nitric-oxide synthase flavin domains closely resemble that of NADPH-cytochrome P450 reductase (CPR). However, all nitric-oxide synthase (NOS) isoforms are 20-40 residues longer in the C terminus, forming a "tail" that is absent in CPR. To investigate its function, we removed the 33 and 42 residue C termini from neuronal NOS (nNOS) and endothelial NOS (eNOS), respectively. Both truncated enzymes exhibited cytochrome c reductase activities without calmodulin that were 7-21-fold higher than the nontruncated forms. With calmodulin, the truncated and wild-type enzymes reduced cytochrome c at approximately equal rates. Therefore, calmodulin functioned as a nonessential activator of the wild type enzymes and a partial noncompetitive inhibitor of the truncated mutants. Truncated nNOS and eNOS plus calmodulin catalyzed NO formation at rates that were 45 and 33%, respectively, those of their intact forms. Without calmodulin, truncated nNOS and eNOS synthesized NO at rates 14 and 20%, respectively, those with calmodulin. By using stopped-flow spectrophotometry, we demonstrated that electron transfer into and between the two flavins is faster in the absence of the C terminus. Although both CPR and intact NOS can exist in a stable, one electron-reduced semiquinone form, neither of the truncated enzymes do so. We propose negative modulation of FAD-FMN interaction by the C termini of both constitutive NOSs. PMID- 10871627 TI - Regulation of intra-Golgi membrane transport by calcium. AB - Calcium cations play a critical role in regulating vesicular transport between different intracellular membrane-bound compartments. The role of calcium in transport between the Golgi cisternae, however, remains unclear. Using a well characterized cell-free intra-Golgi transport assay, we now show that changes in free Ca(2+) concentration in the physiological range regulate this transport process. The calcium-chelating agent 1,2-bis(2-aminophenoxy)ethane-N,N,N',N' tetraacetic acid blocked transport with an IC(50) of approximately 0.8 mm. The effect of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid was reversible by addition of fresh cytosol and was irreversible when performed in the presence of a Ca(2+) ionophore that depletes calcium from lumenal stores. We demonstrate here that intra-Golgi transport is stimulated by low Ca(2+) concentrations (20 100 nm) but is inhibited by higher concentrations (above 100 nm). Further, we show that calmodulin antagonists specifically block intra-Golgi transport, implying a role for calmodulin in mediating the effect of calcium. Our results suggest that Ca(2+) efflux from intracellular pools may play an essential role in regulating intra-Golgi transport. PMID- 10871626 TI - Receptor clustering drives polarized assembly of ankyrin. AB - Expression of the L1 family cell adhesion molecule neuroglian in Drosophila S2 cells leads to cell aggregation and polarized ankyrin accumulation at sites of cell-cell contact. Thus neuroglian adhesion generates a spatial cue for polarized assembly of ankyrin and the spectrin cytoskeleton. Here we characterized a chimera of the extracellular and transmembrane domains of rat CD2 fused to the cytoplasmic domain of neuroglian. The chimera was used to test the hypothesis that clustering of neuroglian at sites of adhesion generates the signal that activates ankyrin binding. Abundant expression of the chimera at the plasma membrane was not a sufficient cue to drive ankyrin assembly, since ankyrin remained diffusely distributed throughout the cytoplasm of CD2-neuroglian expressing cells. However, ankyrin became highly enriched at sites of antibody induced capping of CD2-neuroglian. Spectrin codistributed with ankyrin at capped sites. A green fluorescent protein-tagged ankyrin was used to monitor ankyrin distribution in living cells. Enhanced green fluorescent protein-ankyrin behaved identically to antibody-stained endogenous ankyrin, proving that the polarized accumulation of ankyrin was not an artifact of fixing and staining cells. We propose a model in which clustering of neuroglian induces a conformational change in the cytoplasmic domain that drives polarized assembly of the spectrin cytoskeleton. PMID- 10871628 TI - Entrapping intermediates of thermal aggregation in alpha-helical proteins with low concentration of guanidine hydrochloride. AB - Aggregation of proteins is a problem with serious medical implications and economic importance. To develop strategies for preventing aggregation, the mechanism(s) and pathways by which proteins aggregate must be characterized. In this study, the thermally induced aggregation processes of three alpha-helix proteins (myoglobin, cytochrome c, and lysozyme) in the presence and absence of 1.0 m guanidine hydrochloride (GdnHCl) were investigated by means of infrared spectroscopy. In the absence of GdnHCl, intensities of the alpha-helix bands (approximately 1656 cm(-1)) decrease as a function of temperature at above 50 degrees C. With myoglobin and cytochrome c, the loss of helix bands was accompanied by the appearance of two new bands at 1694 and 1623 cm(-1), indicative of the formation of intermolecular beta-sheet aggregates. For lysozyme, bands indicative of intermolecular beta-sheet aggregates did not appear in any significant intensity. In the presence of 1.0 m GdnHCl, two major intermediate states rich in 3(10)-helix (represented by the band at 1663 cm(-1)) and beta-turn structure (represented by the band at 1667 cm(-1)), respectively, were observed. These findings demonstrated that IR spectroscopic studies of protein aggregation using a combination of thermal and chemical denaturing factors could provide a means to populate and characterize aggregation intermediates. PMID- 10871629 TI - Purification and characterization of N-acetylgalactosamine 4-sulfate 6-O sulfotransferase from the squid cartilage. AB - N-Acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S-6ST), which transfers sulfate from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to position 6 of N-acetylgalactosamine 4-sulfate in chondroitin sulfate and dermatan sulfate, was purified 19,600-fold to apparent homogeneity from the squid cartilage. SDS polyacrylamide gel electrophoresis of the purified enzyme showed a broad protein band with a molecular mass of 63 kDa. The protein band coeluted with GalNAc4S-6ST activity from Toyopearl HW-55 around the position of 66 kDa, indicating that the active form of GalNAc4S-6ST may be a monomer. The purified enzyme transferred sulfate from PAPS to chondroitin sulfate A, chondroitin sulfate C, and dermatan sulfate. The transfer of sulfate to chondroitin sulfate A and dermatan sulfate occurred mainly at position 6 of the internal N-acetylgalactosamine 4-sulfate residues. Chondroitin sulfate E, keratan sulfate, heparan sulfate, and completely desulfated N-resulfated heparin were not efficient acceptors of the sulfotransferase. When a trisaccharide or a pentasaccharide having sulfate groups at position 4 of N-acetylgalactosamine was used as acceptor, efficient sulfation of position 6 at the nonreducing terminal N-acetylgalactosamine 4-sulfate residue was observed. PMID- 10871631 TI - Torsin A and its torsion dystonia-associated mutant forms are lumenal glycoproteins that exhibit distinct subcellular localizations. AB - Early-onset torsion dystonia is an autosomal dominant hyperkinetic movement disorder that has recently been linked to a 3-base pair deletion in the DYT1 gene. The DYT1 gene encodes a 332-amino acid protein, torsin A, that bears low but significant homology to the Hsp100/Clp family of ATPase chaperones. The deletion in DYT1 associated with torsion dystonia results in the loss of one of a pair of glutamic acid residues residing near the C terminus of torsin A (DeltaE torsin A). At present, little is known about the expression, subcellular distribution, and/or function of either the torsin A or DeltaE-torsin A protein. When transfected into mammalian cells, both torsin A and DeltaE-torsin A were found to behave as lumenally oriented glycoproteins. Immunofluorescence studies revealed that torsin A localized to a diffuse network of intracellular membranes displaying significant co-immunoreactivity for the endoplasmic reticulum resident protein BiP, whereas DeltaE-torsin A resided in large spheroid intracellular structures exclusive of BiP immunoreactivity. These results initially suggested that DeltaE-torsin A might exist as insoluble aggregates. However, both torsin A and DeltaE-torsin A were readily solubilized by nonionic detergents, were similarly accessible to proteases, and displayed equivalent migration patterns on sucrose gradients. Collectively, these data support that both the wild type and torsion dystonia-associated forms of torsin A are properly folded, lumenal proteins of similar oligomeric states. The potential relationship between the altered subcellular distribution of DeltaE-torsin A and the disease-inducing phenotype of the protein is discussed. PMID- 10871630 TI - Evolution of C4 phosphoenolpyruvate carboxylase in Flaveria, a conserved serine residue in the carboxyl-terminal part of the enzyme is a major determinant for C4 specific characteristics. AB - C4 phosphoenolpyruvate carboxylases have evolved from ancestral C3 isoforms during the evolution of angiosperms and gained distinct kinetic and regulatory properties compared with the C3 isozymes. To identify amino acid residues and/or domains responsible for these C4-specific properties the C4 phosphoenolpyruvate carboxylase of Flaveria trinervia (C4) was compared with its orthologue in the closely related C3 plant Flaveria pringlei. Reciprocal enzyme chimera were constructed and the kinetic constants, K(0.5) and k(cat), as well as the Hill coefficient, h, were determined for the substrate phosphoenolpyruvate both in the presence and absence of the activator glucose 6-phosphate. By this approach two regions were identified which determined most of the kinetic differences of the C4 and C3 ppcA phosphoenolpyruvate carboxylases with respect to the substrate PEP. In addition, the experiments suggest that the two regions do not act additively but interact with each other. The region between amino acids 296 and 437 is essential for activation by glucose 6-phosphate. The carboxyl-terminal segment between amino acids 645 and 966 contains a C4 conserved serine or a C3 invariant alanine at position 774 in the respective enzyme isoform. Site-directed mutagenesis shows that this position is a key determinant for the kinetic properties of the two isozymes. PMID- 10871632 TI - Erythrocytes reduce extracellular ascorbate free radicals using intracellular ascorbate as an electron donor. AB - Ascorbate is readily oxidized in aqueous solution by ascorbate oxidase. Ascorbate radicals are formed, which disproportionate to ascorbate and dehydroascorbic acid. Addition of erythrocytes with increasing intracellular ascorbate concentrations decreased the oxidation of ascorbate in a concentration-dependent manner. Concurrently, it was found, utilizing electron spin resonance spectroscopy, that extracellular ascorbate radical levels were decreased. Control experiments showed that these results could not be explained by leakage of ascorbate from the cells, inactivation of ascorbate oxidase, or oxygen depletion. Thus, this means that intracellular ascorbate is directly responsible for the decreased oxidation of extracellular ascorbate. Exposure of ascorbate-loaded erythrocytes to higher levels of extracellular ascorbate radicals resulted in the detection of intracellular ascorbate radicals. Moreover, efflux of dehydroascorbic acid was observed under these conditions. These data confirm the view that intracellular ascorbate donates electrons to extracellular ascorbate free radical via a plasma membrane redox system. Such a redox system enables the cells to effectively counteract oxidative processes and thereby prevent depletion of extracellular ascorbate. PMID- 10871633 TI - c-Jun inhibits transforming growth factor beta-mediated transcription by repressing Smad3 transcriptional activity. AB - Transforming growth factor beta (TGF-beta) is a pleiotropic cytokine that exerts its effects through a heteromeric complex of transmembrane serine/threonine kinase receptors. At least two intracellular pathways are activated by TGF-beta as follows: the SAPK/JNK, involving the MEKK1, MKK4, and JNK cascade, and the Smad pathway. Here, we report that the SAPK/JNK pathway inhibits the Smad3 pathway. Expression of dominant negative or constitutively active mutants of kinases of the SAPK/JNK pathway, respectively, activates or represses a TGF-beta induced reporter containing Smad3-binding sites. This effect is not dependent on blocking of Smad3 nuclear translocation but involves a functional interaction between Smad3 and c-Jun, a transcription factor activated by the SAPK/JNK pathway. Overexpression of constitutively active MEKK1 or MKK4 mutants stabilizes the physical interaction between Smad3 and c-Jun, whereas dominant negative mutants inhibit this interaction. Moreover, overexpression of wild-type c-Jun inhibits Smad3-dependent transcription. However, c-Jun does not inhibit Smad3 binding to DNA in vitro. The repression obtained with a c-Jun mutant unable to activate transcription through AP-1 sites indicates that the inhibitory mechanism does not rely on the induction of a Smad3 repressor by c-Jun, suggesting that c Jun could act as a Smad3 co-repressor. The inhibition of the Smad3 pathway by the SAPK/JNK pathway, both triggered by TGF-beta, could participate in a negative feedback loop to control TGF-beta responses. PMID- 10871635 TI - Abstracts PMID- 10871634 TI - The interaction of a neutral ryanoid with the ryanodine receptor channel provides insights into the mechanisms by which ryanoid binding is modulated by voltage. AB - In an earlier investigation, we demonstrated that the likelihood of interaction of a positively charged ryanoid, 21-amino-9alpha-hydroxyryanodine, with the sarcoplasmic reticulum Ca(2+)-release channel (ryanodine receptor, RyR) is dependent on holding potential (Tanna, B., W. Welch, L. Ruest, J.L. Sutko, and A. J. Williams. 1998. J. Gen. Physiol. 112:55-69) and suggested that voltage dependence could result from either the translocation of the charged ligand to a site within the voltage drop across the channel or a voltage-driven alteration in receptor affinity. We now report experiments that allow us to assess the validity of these alternate mechanisms. Ryanodol is a neutral ryanoid that binds to RyR and induces modification of channel function. By determining the influence of transmembrane potential on the probability of channel modification by ryanodol and the rate constants of ryanodol association and dissociation, we demonstrate that the influence of voltage is qualitatively the same for both the neutral and positively charged ryanoids. These experiments establish that most, if not all, of the modification of ryanoid interaction with RyR by transmembrane holding potential results from a voltage-driven alteration in receptor affinity. PMID- 10871636 TI - Gap junction voltage dependence. A clear picture emerges. PMID- 10871637 TI - Stoichiometry of transjunctional voltage-gating polarity reversal by a negative charge substitution in the amino terminus of a connexin32 chimera. AB - Gap junctions are intercellular channels formed by the serial, head to head arrangement of two hemichannels. Each hemichannel is an oligomer of six protein subunits, which in vertebrates are encoded by the connexin gene family. All intercellular channels formed by connexins are sensitive to the relative difference in the membrane potential between coupled cells, the transjunctional voltage (Vj), and gate by the separate action of their component hemichannels (Harris, A.L., D.C. Spray, and M.V. Bennett. 1981. J. Gen. Physiol. 77:95-117). We reported previously that the polarity of Vj dependence is opposite for hemichannels formed by two closely related connexins, Cx32 and Cx26, when they are paired to form intercellular channels (Verselis, V.K., C.S. Ginter, and T.A. Bargiello. 1994. Nature. 368:348-351). The opposite gating polarity is due to a difference in the charge of the second amino acid. Negative charge substitutions of the neutral asparagine residue present in wild-type Cx32 (Cx32N2E or Cx32N2D) reverse the gating polarity of Cx32 hemichannels from closure at negative Vj to closure at positive Vj. In this paper, we further examine the mechanism of polarity reversal by determining the gating polarity of a chimeric connexin, in which the first extracellular loop (E1) of Cx32 is replaced with that of Cx43 (Cx43E1). The resulting chimera, Cx32*Cx43E1, forms conductive hemichannels when expressed in single Xenopus oocytes and intercellular channels in pairs of oocytes (Pfahnl, A., X.W. Zhou, R. Werner, and G. Dahl. 1997. Pflugers Arch. 433:733-779). We demonstrate that the polarity of Vj dependence of Cx32*Cx43E1 hemichannels in intercellular pairings is the same as that of wild-type Cx32 hemichannels and is reversed by the N2E substitution. In records of single intercellular channels, Vj dependence is characterized by gating transitions between fully open and subconductance levels. Comparable transitions are observed in Cx32*Cx43E1 conductive hemichannels at negative membrane potentials and the polarity of these transitions is reversed by the N2E substitution. We conclude that the mechanism of Vj dependence of intercellular channels is conserved in conductive hemichannels and term the process Vj gating. Heteromeric conductive hemichannels comprised of Cx32*Cx43E1 and Cx32N2E*Cx43E1 subunits display bipolar Vj gating, closing to substates at both positive and negative membrane potentials. The number of bipolar hemichannels observed in cells expressing mixtures of the two connexin subunits coincides with the number of hemichannels that are expected to contain a single oppositely charged subunit. We conclude that the movement of the voltage sensor in a single connexin subunit is sufficient to initiate Vj gating. We further suggest that Vj gating results from conformational changes in individual connexin subunits rather than by a concerted change in the conformation of all six subunits. PMID- 10871638 TI - Biophysical and molecular mechanisms underlying the modulation of heteromeric Kir4.1-Kir5.1 channels by CO2 and pH. AB - CO2 chemoreception may be related to modulation of inward rectifier K+ channels (Kir channels) in brainstem neurons. Kir4.1 is expressed predominantly in the brainstem and inhibited during hypercapnia. Although the homomeric Kir4.1 only responds to severe intracellular acidification, coexpression of Kir4.1 with Kir5.1 greatly enhances channel sensitivities to CO2 and pH. To understand the biophysical and molecular mechanisms underlying the modulation of these currents by CO2 and pH, heteromeric Kir4. 1-Kir5.1 were studied in inside-out patches. These Kir4.1-Kir5.1 currents showed a single channel conductance of 59 pS with open-state probability (P(open)) approximately 0.4 at pH 7.4. Channel activity reached the maximum at pH 8.5 and was completely suppressed at pH 6.5 with pKa 7.45. The effect of low pH on these currents was due to selective suppression of P(open) without evident effects on single channel conductance, leading to a decrease in the channel mean open time and an increase in the mean closed time. At pH 8.5, single-channel currents showed two sublevels of conductance at approximately 1/4 and 3/4 of the maximal openings. None of them was affected by lowering pH. The Kir4.1-Kir5.1 currents were modulated by phosphatidylinositol 4,5-bisphosphate (PIP2) that enhanced baseline P(open) and reduced channel sensitivity to intracellular protons. In the presence of 10 microM PIP2, the Kir4.1-Kir5.1 showed a pKa value of 7.22. The effect of PIP2, however, was not seen in homomeric Kir4.1 currents. The CO2/pH sensitivities were related to a lysine residue in the NH2 terminus of Kir4.1. Mutation of this residue (K67M, K67Q) completely eliminated the CO2 sensitivity of both homomeric Kir4.1 and heteromeric Kir4.1-Kir5.1. In excised patches, interestingly, the Kir4.1-Kir5.1 carrying K67M mutation remained sensitive to low pHi. Such pH sensitivity, however, disappeared in the presence of PIP2. The effect of PIP2 on shifting the titration curve of wild-type and mutant channels was totally abolished when Arg178 in Kir5.1 was mutated. Thus, these studies demonstrate a heteromeric Kir channel that can be modulated by both acidic and alkaline pH, show the modulation of pH sensitivity of Kir channels by PIP2, and provide information of the biophysical and molecular mechanisms underlying the Kir modulation by intracellular protons. PMID- 10871639 TI - The role of Na,K-ATPase alpha subunit serine 775 and glutamate 779 in determining the extracellular K+ and membrane potential-dependent properties of the Na,K pump. AB - The roles of Ser775 and Glu779, two amino acids in the putative fifth transmembrane segment of the Na,K-ATPase alpha subunit, in determining the voltage and extracellular K+ (K+(o)) dependence of enzyme-mediated ion transport, were examined in this study. HeLa cells expressing the alpha1 subunit of sheep Na,K-ATPase were voltage clamped via patch electrodes containing solutions with 115 mM Na+ (37 degrees C). Na,K-pump current produced by the ouabain-resistant control enzyme (RD), containing amino acid substitutions Gln111Arg and Asn122Asp, displayed a membrane potential and K+(o) dependence similar to wild-type Na,K ATPase during superfusion with 0 and 148 mM Na+-containing salt solutions. Additional substitution of alanine at Ser775 or Glu779 produced 155- and 15-fold increases, respectively, in the K+(o) concentration that half-maximally activated Na,K-pump current at 0 mV in extracellular Na+-free solutions. However, the voltage dependence of Na,K-pump current was unchanged in RD and alanine substituted enzymes. Thus, large changes in apparent K+(o) affinity could be produced by mutations in the fifth transmembrane segment of the Na,K-ATPase with little effect on voltage-dependent properties of K+ transport. One interpretation of these results is that protein structures responsible for the kinetics of K+(o) binding and/or occlusion may be distinct, at least in part, from those that are responsible for the voltage dependence of K+(o) binding to the Na,K-ATPase. PMID- 10871640 TI - Electrogenic sodium-sodium exchange carried out by Na,K-ATPase containing the amino acid substitution Glu779Ala. AB - Na,K-ATPase containing the amino acid substitution glutamate to alanine at position 779 of the alpha subunit (Glu779Ala) supports a high level of Na-ATPase and electrogenic Na+-Na+ exchange activity in the absence of K+. In microsomal preparations of Glu779Ala enzyme, the Na+ concentration for half maximal activation of Na-ATPase activity was 161 +/- 14 mM (n = 3). Furthermore, enzyme activity with 800 mM Na+ was found to be similar in the presence and absence of 20 mM K+. These results showed that Na+, with low affinity, could stimulate enzyme turnover as effectively as K+. To gain further insight into the mechanism of this enzyme activity, HeLa cells expressing Glu779Ala enzyme were voltage clamped with patch electrodes containing 115 mM Na+ during superfusion in K+-free solutions. Electrogenic Na+-Na+ exchange was observed as an ouabain-inhibitable outward current whose amplitude was proportional to extracellular Na+ (Na+(o)) concentration. At all Na+(o) concentrations tested (3-148 mM), exchange current was maximal at negative membrane potentials (V(M)), but decreased as V(M) became more positive. Analyzing this current at each V(M) with a Hill equation showed that Na+-Na+ exchange had a high-affinity, low-capacity component with an apparent Na+(o) affinity at 0 mV (K0(0.5)) of 13.4 +/- 0.6 mM and a low-affinity, high-capacity component with a K0(0.5) of 120 +/- 13 mM (n = 17). Both high- and low-affinity exchange components were V(M) dependent, dissipating 30 +/- 3% and 82 +/- 6% (n = 17) of the membrane dielectric, respectively. The low-affinity, but not the high-affinity exchange component was inhibited with 2 mM free ADP in the patch electrode solution. These results suggest that the high-affinity component of electrogenic Na+-Na+ exchange could be explained by Na+(o) acting as a low-affinity K+ congener; however, the low-affinity component of electrogenic exchange appeared to be due to forward enzyme cycling activated by Na+(o) binding at a Na+-specific site deep in the membrane dielectric. A pseudo six-state model for the Na,K-ATPase was developed to simulate these data and the results of the accompanying paper (Peluffo, R.D., J.M. Arguello, and J.R. Berlin. 2000. J. Gen. Physiol. 116:47-59). This model showed that alterations in the kinetics of extracellular ion-dependent reactions alone could explain the effects of Glu779Ala substitution on the Na,K-ATPase. PMID- 10871642 TI - Heat stress reduces poly(ADPR)polymerase expression in rat testis. AB - Poly(ADPR)polymerase (PARP) is a chromatin-associated enzyme with a presumptive role in DNA repair during replication and recovery from strand breaks caused by genotoxic agents. It catalyses the attachment and elongation of ADPribose polymers (pADPR) to a variety of acceptor proteins (including PARP itself, and histones) and is particularly active in the testis where its expression varies according to the stage of germ cell differentiation. PARP degradation is also one of the classic indicators of apoptosis. In this investigation we have examined the effects of heat stress on the adult rat testis with respect to the concentration and activity of PARP, the nature of the pADRP nuclear acceptor proteins, the length of ADPR polymers and the activity of the ADPR depolymerizing enzyme, poly(ADPR)glycohydrolase (PARG). Our results show a significant reduction in the concentration and activity of PARP 4 and 8 days after artificial cryptorchidism, but no significant changes were observed in PARG activity or in pADPR length. Unexpectedly, the apoptotic degradation of PARP was not detected following heat stress. These results confirm that PARP gene expression is developmentally regulated during spermatogenesis and indicate that it is suppressed coincidentally with the loss of meiotic spermatocytes during artificial cryptorchidism. PMID- 10871641 TI - Voltage and Ca2+ activation of single large-conductance Ca2+-activated K+ channels described by a two-tiered allosteric gating mechanism. AB - The voltage- and Ca2+-dependent gating mechanism of large-conductance Ca2+ activated K+ (BK) channels from cultured rat skeletal muscle was studied using single-channel analysis. Channel open probability (Po) increased with depolarization, as determined by limiting slope measurements (11 mV per e-fold change in Po; effective gating charge, q(eff), of 2.3 +/- 0.6 e(o)). Estimates of q(eff) were little changed for intracellular Ca2+ (Ca2+(i)) ranging from 0.0003 to 1,024 microM. Increasing Ca2+(i) from 0.03 to 1,024 microM shifted the voltage for half maximal activation (V(1/2)) 175 mV in the hyperpolarizing direction. V(1/2) was independent of Ca2+(i) for Ca2+(i) < or = 0.03 microM, indicating that the channel can be activated in the absence of Ca2+(i). Open and closed dwell time distributions for data obtained at different Ca2+(i) and voltage, but at the same Po, were different, indicating that the major action of voltage is not through concentrating Ca2+ at the binding sites. The voltage dependence of Po arose from a decrease in the mean closing rate with depolarization (q(eff) = -0.5 e(o)) and an increase in the mean opening rate (q(eff) = 1.8 e(o)), consistent with voltage-dependent steps in both the activation and deactivation pathways. A 50-state two-tiered model with separate voltage- and Ca2+-dependent steps was consistent with the major features of the voltage and Ca2+ dependence of the single-channel kinetics over wide ranges of Ca2+(i) (approximately 0 through 1,024 microM), voltage (+80 to -80 mV), and Po (10(-4) to 0.96). In the model, the voltage dependence of the gating arises mainly from voltage-dependent transitions between closed (C-C) and open (O-O) states, with less voltage dependence for transitions between open and closed states (C-O), and with no voltage dependence for Ca2+-binding and unbinding. The two-tiered model can serve as a working hypothesis for the Ca2+- and voltage-dependent gating of the BK channel. PMID- 10871644 TI - Preliminary investigation of follistatin gene mutations in women with polycystic ovary syndrome. AB - Strong evidence for a link between the follistatin gene and polycystic ovary syndrome (PCOS) has recently been found in a well-designed large-scale study. Follistatin binds to activin and affects its functions, e.g. stimulation of FSH synthesis and secretion. Thus, it may play a role in the functional impairment of the FSH-granulosa cell axis in PCOS. In this study, we screened 64 Chinese patients with PCOS for mutations in the entire coding region (including the region encoding alternative carboxy-terminals) of the follistatin gene using polymerase chain reaction (PCR)-based single-stranded conformational polymorphism (SSCP) and DNA sequencing. However, we could not identify a single mutation of either the activating or inhibiting type, using these techniques. Therefore, it would appear that PCOS in the local Chinese population is not caused by mutations in the coding regions of the follistatin gene. PMID- 10871643 TI - Expression of GnRH receptor in mouse and rat testicular germ cells. AB - The expression of gonadotrophin-releasing hormone receptor (GnRH-R) in germinal cells of mouse testis, whole testes, pituitary glands, and mouse ovaries was determined by means of Northern hybridization using a mouse GnRH-R [(32)P] labelled cDNA probe. Also, the expression of GnRH-R in rat germinal cells, testis and pituitary gland was determined by Northern blot analysis using the same mouse specific probe. Three receptor transcripts were detected in all cases. In mouse pituitary, ovary and testis, we found two GnRH-R transcripts in the proximity of 4.6-4.7 and 3.4 kb, as well as a 1.6 kb transcript in the pituitary and a 2.0-2.1 kb transcript in both the ovary and testis. Mouse germ cells also exhibited three GnRH-R transcripts of 4.7, 3.5 and 2.2 kb. Two distinct GnRH-R transcripts were also detected in the rat pituitary (4.6 and 2.1 kb), testis (4. 7 and 3.5 kb) and germ cells (4.5 and 3.5 kb). In addition, a third transcript was detected in rat pituitary (1.9 kb) and in rat testis and germinal cells (2.1 kb). The present study demonstrates that GnRH-R mRNA is expressed in rat and mouse testicular germ cells. We suggest that GnRH-R present in these cells may interact with GnRH or GnRH-like peptides produced in the testis and may be part of a paracrine system. The presence of multiple GnRH-R encoding transcripts is also of interest and warrants further studies to evaluate their regulation and function. PMID- 10871645 TI - X inactivation-specific transcript expression in mouse oocytes and zygotes. AB - Expression of the X inactivation-specific transcript (XIST:) gene has previously been shown by reverse transcription-polymerase chain reaction (RT-PCR) to be present at the 4-cell stage of female mouse embryos. This early expression, which is followed by X inactivation in the extra-embryonic tissues, is maternally imprinted. By the blastocyst stage, as the embryonic lineages begin to form, the imprint is lost and expression becomes random. By applying in-situ RT-PCR, we showed that XIST: is expressed even earlier in development, in unfertilized mouse oocytes as well as in pronuclei stage zygotes. Our data demonstrate XIST: expression in oocytes and suggest that XIST: transcripts may occur in both XX and XY zygotes. A difference in the pattern of expression (rod-like or rounded punctate signal) is found among pronuclear-stage embryos. Early expression is in agreement with findings reported in human embryos. PMID- 10871646 TI - The endometrium as a novel target for leptin: differences in fertility and subfertility. AB - Leptin and its receptor are involved in endocrine and paracrine regulation of metabolism, obesity and reproduction. Here, we describe the detection of the functional long isoform receptor of leptin in human endometrium. The leptin receptor protein was shown to be expressed in glandular and luminal epithelium and is periodically regulated throughout the menstrual cycle, demonstrating main expression in follicular and mid-luteal phase. In contrast, leptin receptor mRNA is detectable by reverse transcription-polymerase chain reaction (RT-PCR) as a constitutive component. Since RT-PCR analyses showed that leptin is not expressed in this tissue, the present study suggests that the human endometrium is a novel target for leptin. Therefore, we investigated 11 subfertile patients who underwent two biopsies in one menstrual cycle. The patients presented with a repetitive endometrial maturation defect, but showed adequate serum hormone concentrations and normal steroid hormone receptor expression and down-regulation in the endometrium. These patients were, however, deficient for expression of the functional leptin receptor. These analyses provide evidence that the lack of the leptin receptor in an ovulatory cycle may contribute to subfertility by a yet undefined 'endometrial factor'. PMID- 10871647 TI - Kappa opioids and TGFbeta1 interact in human endometrial cells. AB - The transforming growth factor beta1 (TGFbeta1) is a major regulator of human endometrial function. Human endometrium possesses specific opioid binding sites, the majority of which belong to the kappa type, for which the prodynorphin derived opioids are the endogenous ligands. Since these two systems interact in several other tissues we postulated that opioids may affect the production of TGFbeta1 in human endometrium. We have found that kappa opioids exerted a time- and dose-dependent inhibitory effect on TGFbeta1 production from endometrial stromal and epithelial cells and from the Ishikawa human endometrial adenocarcinoma cell line. This effect was reversible by the specific opioid antagonist diprenorphine. To examine if this effect represents a paracrine endometrial response to locally produced kappa opioids we searched for the presence of the endogenous kappa opioid receptor ligands. Indeed, the prodynorphin transcript was detectable on Northern blots from normal and tumoral human endometrial cells; its size was that of the pituitary transcript, i.e. approximately 2.4 kb long. Most immunoreactive dynorphin from human endometrium had a molecular weight of 8 kDa. Finally, immunofluorescence staining of normal and tumoral human endometrial cells revealed the presence of dynorphin-positive cytoplasmic secretory granules. Taken together, our data suggest that in human endometrium, kappa opioids and the TGFbeta1 form a paracrine network which appears to be retained by the Ishikawa human endometrial adenocarcinoma cell line. PMID- 10871648 TI - Reduced proliferation and cell adhesion in endometriosis. AB - Endometriosis is defined as endometriotic tissues growing outside the uterine cavity. The cell biological processes responsible for the pathogenesis of this disease are not well understood. In order to detect differences in proliferative activity between endometria and endometriotic lesions, Ki67 staining was analysed. In addition, expression of epidermal growth factor (EGF) and its receptor was examined using immunohistochemistry. For dedifferentiation processes pointing to invasive properties of the uterine epithelium, the presence of the adhesion complex E-cadherin with the associated alpha- and beta-catenin was investigated. Specimens of endometrium in the proliferative phase of 36 patients without, and 79 patients with, endometriosis together with endometriotic lesions were studied. The study revealed a significantly reduced proliferation activity in uterine epithelium within the ectopic lesions but no differences between eutopic endometria of non-affected and affected patients. Furthermore, a lower expression of both EGF and its receptor in the epithelial cells of the ectopic glands was observed. The adhesion complex E-cadherin, together with alpha-, and beta-catenin, was slightly reduced in uterine epithelial cells of women with endometriosis and less expressed in endometriotic lesions. The results indicate that epithelial cells of endometriotic lesions are not hyperproliferative, but do appear to dedifferentiate, displaying an invasive character. PMID- 10871649 TI - Ovarian hormones modulate monocyte chemotactic protein-1 expression in endometrial cells of women with endometriosis. AB - Endometriosis, a frequent oestrogen-dependent disease believed to result from an aberrant proliferation of endometrial tissue outside the uterine cavity, is associated with an increased expression of monocyte chemotactic protein-1 (MCP-1) in the intrauterine endometrium. This makes it plausible that migrating endometrial cells are intrinsically able to initiate monocyte chemoattraction and activation, a phenomenon which has been consistently observed in the peritoneal cavity of patients and recently in their eutopic endometrium. To elucidate the mechanisms involved in the regulation of MCP-1 expression in eutopic endometrial cells, we studied the effects of ovarian hormones and found that oestradiol (10( 9) and 10(-8) mol/l) markedly increased MCP-1 mRNA steady-state levels and protein secretion by endometrial cells in response to interleukin-1beta (IL 1beta) (0.1 ng/ml). The IL-1beta-induced MCP-1 expression was even higher following pretreatment of cells with both oestradiol (10(-9) mol/l) and progesterone (5x10(-8) mol/l). This did not seem to be due to increased MCP-1 mRNA stability, but rather to a higher level of gene transcription. Our results provide evidence that ovarian steroids regulate, indirectly, the synthesis and the secretion of a potent chemotactic and activating factor for monocytes/macrophages by endometrial cells of women with endometriosis and reveal a new mechanism for oestradiol action. PMID- 10871650 TI - Regulated expression of cytokines in human endometrium throughout the menstrual cycle: dysregulation in habitual abortion. AB - It is widely assumed that, after ovulation, the human endometrium undergoes specific changes and becomes receptive to the implantation of embryo during the mid-secretory phase. When implantation does not take place, further changes occur which eventually result in the shedding of human endometrium. The present study was carried out to examine whether there are changes in the cytokine gene expression in human endometrium which are correlated with endometrial function in various phases of the menstrual cycle. The RNase protection assay was performed on carefully dated endometria from normal subjects to characterize the expression of cytokines which potentially contribute to endometrial function. These included: tumour necrosis factor (TNF), interleukin (IL)-1beta, IL-6, IL-8, leukaemia inhibitory factor (LIF), transforming growth factor beta1 (TGF-beta1), macrophage colony stimulating factor (MCSF or colony stimulating factor-1), and vascular endothelial growth factor (VEGF) mRNAs. A low level of expression of these cytokine mRNAs was found during the proliferative and early secretory phase. Expression of cytokine mRNA increased during the mid-secretory phase and rose to a peak in the late secretory phase. The level of cytokine mRNA expression during gestation was most akin to that observed during the mid-secretory phase. Individuals with habitual abortion presented with an abnormal expression of IL 1beta and IL-6 mRNA in endometrium, during the mid-secretory phase. Taken together, these findings are consistent with a progressive rise in the expression of cytokines in human endometrium during the secretory phase in natural cycles. Furthermore, the findings show that habitual abortion is associated with the abnormal expression of IL-1beta and IL-6 in the mid-secretory phase. PMID- 10871651 TI - Cytotoxic effects of tumour necrosis factor (TNF)-alpha and interferon-gamma on cultured human trophoblast are modulated by fibronectin. AB - Tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma, produced by maternal inflammatory cells, may compromise trophoblast survival at the trophoblast-maternal interface and notably in the placental bed which is invaded by trophoblast. Extracellular matrix components, e.g. fibronectin, may enhance trophoblast survival. A possible protective effect of fibronectin against toxic effects of TNF-alpha and IFN-gamma was investigated in cultured trophoblasts isolated from six human term placentas, grown on uncoated and fibronectin-coated plastics. IFN-gamma and increasing doses of TNF-alpha resulted in decreasing viability of trophoblast on uncoated as well as fibronectin-coated dishes, as shown by 3-[4, 5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assays, but for each TNF/IFN treatment condition viability on fibronectin was higher (P < 0.001). Epidermal growth factor (EGF), a growth factor reported to protect against TNF-alpha/IFN-gamma induced toxicity, resulted in further increased viability, but not if IFN-gamma was included in the treatment. EGF caused increased fibronectin secretion into the medium (P < 0.001), and double cytokeratin/fibronectin immunostaining confirmed the trophoblastic nature of fibronectin secreting cells. We conclude that fibronectin increases viability, but does not completely abolish the cytotoxic action of TNF-alpha and IFN-gamma on trophoblast. The protective effect of EGF may be related to stimulation of fibronectin secretion by trophoblast. PMID- 10871652 TI - Decreased superoxide dismutase expression and increased concentrations of lipid peroxide and prostaglandin F(2alpha) in the decidua of failed pregnancy. AB - To study the possible role of the superoxide radical and its scavenging system in the decidua of early pregnancy, superoxide dismutase (SOD) values and concentrations of lipid peroxide and prostaglandin F(2alpha) (PGF(2alpha)) were analysed in the decidua obtained from normal pregnancy and failed pregnancy. Failed pregnancy was divided into two groups; spontaneous abortion with or without vaginal bleeding. In the spontaneous abortion with vaginal bleeding, total SOD activities, Cu,Zn-SOD activities and Cu,Zn-SOD mRNA values in the decidua were significantly lower, and concentrations of lipid peroxide and PGF(2alpha) were significantly higher, than those in the normal pregnancy and the spontaneous abortion without vaginal bleeding. In contrast, activities and mRNA values of Mn-SOD were significantly higher in the spontaneous abortion with vaginal bleeding than the other two groups. There was no significant difference in all of these parameters between the normal pregnancy and the spontaneous abortion without vaginal bleeding. In conclusion, the decrease in Cu,Zn-SOD expression and the increase in lipid peroxide in the decidua could be involved in the termination of spontaneous abortion, mediated through the increase in PGF(2alpha) synthesis. In other words, Cu,Zn-SOD may contribute to the maintenance of pregnancy by preventing the accumulation of superoxide radicals that cause PGF(2alpha) synthesis. PMID- 10871653 TI - Expression of the cyclic AMP-dependent transcription factors, CREB, CREM and ATF2, in the human myometrium during pregnancy and labour. AB - Elevated concentrations of cyclic AMP (cAMP) in the human myometrium may promote uterine quiescence during pregnancy by protein kinase A (PKA)-mediated phosphorylation and subsequent inactivation of myosin light-chain kinase, as well as by the phosphorylation and activation of cAMP-dependent transcription factors. In this context, we show that the altered expression of cAMP response-element binding protein (CREB), cAMP response-element modulator protein (CREM) and activating transcription factor 2 (ATF2) are implicated in the maintenance of myometrial quiescence during fetal maturation and the switch to uterine activation at term. Using electrophoretic mobility shift and super shift assays, as well as immunoblotting of paired myometrial tissue samples from non-pregnant, pregnant non-labouring and spontaneous labouring women, we defined the patterns of expression of various isoforms of these proteins in the human uterus. Here, we report spatio-temporal changes in the expression of a 43 kDa form of CREB, a 28 kDa CREM-like protein, and a novel 28 kDa ATF2-like protein which are differentially expressed, depending on the gestational state of the uterus. Changes in the pattern of expression of these potent transcription factors may have an important role in the control of uterine activity throughout pregnancy. PMID- 10871654 TI - Effects of progesterone on prostaglandin E(2)-induced changes in glycosaminoglycan synthesis by human cervical fibroblasts in culture. AB - Prostaglandins are known to induce cervical ripening and this effect may be mediated by an increase in glycosaminoglycan (GAG) concentration. The aim of this study was to assess the effects of progesterone on prostaglandin E(2) (PGE(2)) induced changes in GAG synthesis by human cervical cells in culture. Human cervical fibroblasts were obtained by cervical biopsies in hormonally active women and cultured. Cells were submitted to an incubation with progesterone or control medium. A second incubation was then performed with increasing concentrations of PGE(2). GAG synthesis by the cervical cells was assayed after extraction, by incorporation of [(3)H]-glucosamine and [(35)S]-sulphate into GAGs. It was found that progesterone alone induced a dose-dependent increase in GAG synthesis. After pre-incubation with progesterone, PGE(2) further increased [(3)H]-glucosamine and [(35)S]-sulphate uptake. However, when expressed as percentage of stimulation, the stimulatory effect of PGE(2) on GAG synthesis was inhibited at high progesterone concentrations. Therefore we concluded that, although high concentrations of progesterone increase the overall synthesis of GAG, they may also play a preventative role against PGE(2)-induced changes in GAG production during pregnancy. PMID- 10871655 TI - Fluorescence in-situ hybridization analysis of chromosomal constitution in spermatozoa from a mosaic 47,XYY/46,XY male. AB - Sex-chromosome mosaicism in spermatozoa from a mosaic 47,XYY[20%]/46, XY[80%] male with fertility problems was assessed using triple-probe fluorescence in-situ hybridization (FISH) studies. Chromosome-specific probes for X, Y and 18 were used, and the possible outcomes were deduced. In normal haploid spermatozoa of the patient and a normal 46,XY male control, the X:Y ratio was close to 1:1. There was a significant difference in the total incidence of karyotypically abnormal spermatozoa between the patient and the 46, XY male control (2.31% versus 1.46%, P < 0.0001). The incidence of some types of disomic spermatozoa X+Y+18 (24,XY) and X+18+18 (24,X, +18), or diploid X+Y+18+18 (46,XY) spermatozoa was significantly increased in the patient's semen sample. There was, however, no significant difference in the incidence of disomic Y+Y+18 (24,YY) spermatozoa. Because the majority of the patient's spermatozoa was karyotypically normal, the aetiology of his fertility problems was unclear. These results add to the growing body of information regarding chromosome abnormalities in spermatozoa from men who are mosaic for sex chromosome abnormalities. In these men, FISH analysis of spermatozoa may be warranted to determine the relative percentages of abnormal cells, and to determine if in-vitro fertilization with preimplantation genetic diagnosis may increase the likelihood of a successful pregnancy. PMID- 10871656 TI - Homozygosity of the cystic fibrosis (CF) gene allele IVS8-(5T) in a Tamil male with congenital bilateral absence of the vas deferens (CBAVD). PMID- 10871657 TI - Nursing, medication education and the new policy agenda: the evidence base. AB - Current social and demographic trends, combined with 'the new policy agenda', highlight the importance of nurses' role in educating patients about medication. In the absence of previous research investigation, this study set out to explore nurses' current contribution to medication education and the clinical contextual factors that influence current practice. The evidence base for effective medication education was established from reviews of literature and focus groups with key informants. Nurses' practice was investigated using a case study approach in seven clinical areas representing adult, care of the older person, mental health and community nursing contexts. Methods used to collect data were: audio-recordings (n=37) and observation (n=48) of nurse-patient interactions about medication, post-interaction interviews with nurses (n=29), post interaction interviews with patients (n=39), analysis of relevant written documentation and researcher observation and field notes. Data sources within each case were subjected to systematic content analysis in order to identify current practice and contextual influences within each case. Cross-case analysis was also employed in order to identify explanations for any differentiation in practice. Findings indicate that nurses' contribution to medication education is commonly limited to simple information giving about medicines, involving the name, purpose, colour, number of tablets and the time and frequency that medications should be administered. Nurses' practice in two of the seven clinical areas was characterised by interactions that more closely demonstrated features of what is known to constitute more comprehensive and effective medication education. Analysis of contextual influences within and between cases allowed explanations to be derived for the types of medication education interactions observed. These concerned: patient characteristics, perceived and expressed preferences of patients for information, characteristics of the nurse-patient relationship, lack of time and high workload, and the philosophy of care within the clinical area. In all clinical areas, nurses were not explicitly and judiciously using available evidence to inform their medication-related interactions. The paper concludes with discussion and implications of the findings. PMID- 10871658 TI - A comparison of an international experience for nursing students in developed and developing countries. AB - Theoretical perspectives and approaches to transcultural nursing have been developed and there are different options for teaching transcultural care. The opportunity for nursing students to gain healthcare experience in another country is one option. This article reports a study undertaken in Northern Ireland to evaluate outcomes of a 3-month international experience for undergraduate nursing students (n=74) and to assess differences between the experiences in developed and developing countries. Data were collected by questionnaire. The findings indicate a high impact on students' international perspective and career development. Students' understanding of cultural and political issues within Northern Ireland was enhanced. PMID- 10871659 TI - Psychosocial factors influencing personal control in pain relief. AB - A questionnaire was administered to 100 women (50 primigravidae, 50 multigravidae) to investigate the influence of psychosocial factors on personal control in pain relief. Personal control was measured using a 36-item scale based on the concept of 'Rule'. The women were asked to rate each rule on a 7-point Likert scale. Measures of psychosocial factors included assessment of the women's expectations of labour pain, maternal confidence, pain intensity, antenatal training and partner support. Demographic variables including parity were also recorded. The questionnaires were completed prior to and within 48 h following the women's delivery (whilst they were inpatients on the postnatal ward).Two variables, antenatal training and pain intensity, emerged as predictors of personal control in pain relief following stepwise multiple regression analysis. These findings have implications for clinical practice particularly in the area of antenatal care and planning care during labour. PMID- 10871660 TI - An exploratory pilot study of nurse-midwives' attitudes toward active euthanasia and abortion. AB - Over the past three decades, active euthanasia and abortion have received increasing international attention. Since both these practices are relevant to the role of the nurse-midwife, it is important to know what influences their attitudes towards them. Therefore, the purpose of this study was: 1, to survey the attitudes of nurse-midwives' to active euthanasia and its legalization; 2, to determine the relationship between nurse-midwives' attitudes toward active euthanasia and its legalization, and attitudes toward abortion, self-reported religiosity and religious affiliation. The study setting was an international midwifery conference and the sample consisted of 139 nurse-midwives attending the conference. The majority of nurse-midwives displayed a positive attitude toward active euthanasia and its legalization. In addition, there was a positive relationship between their attitude to abortion and active euthanasia. Self reported religiosity and religious affiliation were significantly related to attitudes toward active euthanasia and its legalization. An interesting positive relationship between country of practice and attitudes to euthanasia was also found. Nurse-midwives practicing in countries with more liberal euthanasia and assisted suicide legislation were more supportive of active euthanasia. With the increasing acceptance of active euthanasia's legalization, the results of this study pose some ethical questions that nurse-midwives internationally will have to consider. PMID- 10871661 TI - The lived experience of the oncology clinical nurse specialist. AB - The purpose of this study was to explore what was important and what was unique in the experience of the oncology clinical nurse specialist (OCNS). The sample included eight clinical nurse specialists from one geographical area. One to one interviews focused on reflection on two critical incidents from the OCNSs' own clinical experience: one which they thought had gone particularly well and one that had not gone well. Thematic analysis of the OCNS narratives revealed the main themes of uncertain ground, boundaries, support, and reflective practice. Issues which evolved from thematic analysis were related to the developmental needs of the OCNSs involved in the study. Some aspects of the uniqueness of the role was apparent. Recommendations for further research are made. PMID- 10871662 TI - Education for advanced nursing practice: an evolving framework. AB - This paper presents and critiques a framework for advanced health care practice. The framework is a set of professional attributes which underpin the delivery of an inter-disciplinary postgraduate course in the United Kingdom (UK). It enables students to review and develop knowledge and skills required to lead and advance nursing and health care practice. The framework is reviewed in relation to UK policy discussion on advanced or higher levels of practice. Brief comparisons are made with international concepts and literature on advanced nursing practice. The potential contribution of this framework to nursing practice and education is discussed. In particular, the framework provides a degree of clarity and coherence in specifying the nature and scope of advanced and higher level practice in the UK. PMID- 10871663 TI - When nurses cry: coping with occupational stress in Thailand. AB - Anecdotal reports of people feeling better after they cry support theories that link crying to the reduction of stress after a period of prolonged sympathetic activation. A sample of 200 nurses were asked to rate their occupational stress, job satisfaction, and crying as a coping strategy. Crying was found to be an important symptom of home/work conflicts and pressures related to dealing with patients, but did not substantially reduce these sources of stress. Supporting the stress-buffering hypothesis, nurses with lower intrinsic job satisfaction seemed to benefit from emotional crying whereas dissatisfied nurses who cry infrequently reported the highest levels of stress. PMID- 10871664 TI - An international methodology to describe clinical nursing phenomena: a team approach. AB - The development of a structured and standardized clinical language for nursing is of major importance to the profession for both practice and science. This paper describes a methodological approach which has been developed for the refinement and extension of the NANDA taxonomy in the Nursing Diagnosis Extension and Classification (NDEC) project. The paper proposes that this method could be used by nurses in all countries to facilitate the identification, development and validation of terms and labels which can be incorporated into each country's own emerging data systems, translated and cross mapped between systems, and eventually incorporated into international data sets. PMID- 10871691 TI - Preface PMID- 10871692 TI - Brain atrophy and neuronal loss in alcoholism: a role for DNA damage? AB - Chronic alcohol abuse has deleterious effects on several organs in the body including the brain. Neuroradiological studies have demonstrated that the brains of chronic alcoholics undergo loss of both gray and white matter volumes. Neuropathological studies using unbiased stereological methods have provided evidence for loss of neurons in specific parts of the brain in chronic alcoholics. The purpose of this paper is to propose a mechanism for this alcohol related neuronal loss. The hypothesis is based on the neurodegeneration observed in patients with the genetic disorder xeroderma pigmentosum (XP), who lack the capacity to carry out a specific type of DNA repair called nucleotide excision repair (NER). Some XP patients develop a progressive atrophic neurodegeneration, termed XP neurological disease, indicating that endogenous DNA damage that is normally repaired by NER has the capacity to cause neuronal death. Accumulating evidence indicates that the neurodegenerative DNA damage that is responsible for neuronal loss in XP patients results from reactive oxygen species (ROS) and lipid peroxidation products, and has the capacity to inhibit gene expression by RNA polymerase II. Therefore, the following model is proposed: chronic alcohol abuse results in increased levels of ROS and lipid peroxidation products in neurons, which results in an overwhelming burden on the NER pathway, and increased steady state levels of DNA lesions that inhibit gene expression. This results in neuronal death either by reduction in the levels of essential gene products or by apoptosis. The implications of this model for future studies are discussed. PMID- 10871693 TI - Role of the GABA(A)beta2, GABA(A)alpha6, GABA(A)alpha1 and GABA(A)gamma2 receptor subunit genes cluster in drug responses and the development of alcohol dependence. AB - gamma-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter of the central nervous system and it acts at the GABA(A) and GABA(B) receptors. A possible role for the GABA(A) receptors in alcohol action has been derived from in vitro cell models, animal studies and human research. GABA(A) subunit mRNA expression in cell models has suggested that the long form of the gamma2 subunit is essential for ethanol enhanced potentiation of GABA(A) receptors, by phosphorylation of a serine contained within the extra eight amino acids. Several animal studies have demonstrated that alterations in drug and alcohol responses may be caused by amino-acid differences at the GABA(A)alpha6 and GABA(A)gamma2 subunits. An Arg(100)/Glu(100) change at the GABA(A)alpha6 subunit conferring altered binding efficacy of the benzodiazepine inverse agonist Ro 15-4513, was found between the AT (alcohol tolerance) and ANT (alcohol non-tolerance) rats. Several loci related to alcohol withdrawal on mouse chromosome 11 which corresponds to the region containing four GABA(A) subunit (beta2, alpha6, alpha1 and gamma2) genes on human chromosome 5q33-34, were also identified. Gene knockout studies of the role of GABA(A)alpha6 and GABA(A)gamma2 subunit genes in mice have demonstrated an essential role in the modulation of other GABA(A) subunit expression and the efficacy of benzodiazepine binding. Absence of the GABA(A)gamma2 subunit gene has more severe effects with many of the mice dying shortly after birth. Disappointingly few studies have examined the effects of response to alcohol in these gene knockout mice. Human genetic association studies have suggested that the GABA(A)beta2, alpha6, alpha1 and gamma2 subunit genes have a role in the development of alcohol dependence, although their contributions may vary between ethnic group and phenotype. In summary, in vitro cell, animal and human genetic association studies have suggested that the GABA(A)beta2, alpha6, alpha1 and gamma2 subunit genes have an important role in alcohol related phenotypes (300 words). PMID- 10871694 TI - Genotype effects on neurodegeneration and neuroadaptation in monoaminergic neurotransmitter systems. AB - Neuroadaptation and neurodegeneration in central dopaminergic and serotonergic systems are central to vulnerability, process and consequences of addictive behavior. Serotonergic dysfunction has been associated with behavior disinhibition and negative mood states that may predispose to excessive alcohol intake, while alcohol-induced stimulation of dopaminergic neurotransmission may encode the reinforcing properties of alcohol consumption. Chronic alcohol intake induces neuroadaptive reductions in striatal dopamine transporter (DAT) and D2 receptor availability, which were reversible during early abstinence. A polymorphism of the DAT gene (SLC6A3) was associated with the in vivo transporter availability in the putamen of abstinent alcoholics and control subjects. The same genotype was associated with severity of alcohol withdrawal symptoms, hypothetically due to interactions of genotype and alcohol-induced neuroadaptation. Reduction in raphe serotonin transporter (5-HTT) availability was observed in abstinent male alcoholics and it may be the result of neurodegeneration rather than reversible neuroadaptation. Neurotoxic reduction in 5-HTT protein expression seems to be limited to homozygous carriers of a long, more transcriptionally active allele of a promoter repeat polymorphism of the 5 HTT gene (SCL6A4). This genotype was also associated with a low level of acute unpleasant effects of alcohol consumption, a factor predisposing to excessive alcohol intake. The time course of neuroadaptation and recovery of monoaminergic neurotransmission in alcohol intake and withdrawal imply that monoamine transporter genotype could profoundly influence alcohol-induced reinforcement and, perhaps, contribute to neurochemical changes which are long lasting or permanent. PMID- 10871695 TI - Ovariectomy has minimal effects on neuroadaptations associated with ethanol dependence in female rats. AB - We previously found gender selective alterations in gene expression for GABA(A) and NMDA receptors associated with the development of ethanol dependence. Males and females have a differing hormonal environment, including steroid hormone derivatives (neuroactive steroids) that exert effects at GABA(A) and NMDA receptors. Therefore, we explored whether the removal of ovarian steroids would alter gender differences in response to chronic ethanol exposure. We found that ovariectomy reduced ethanol drinking levels by 15%, comparable to earlier observations between intact female and male rats. However, investigation of the effects of chronic ethanol exposure on intact versus ovariectomized female rats uncovered few differences in chronic ethanol-induced alterations in selected GABA(A) or NMDA receptor subunit peptide levels. In general, findings for both groups of females were similar to previous observations. There was no reduction in GABA(A) receptor alpha1 subunit levels in cerebral cortex in either intact or ovariectomized female rats, in contrast to the significant reduction observed in male rats. In addition, both intact and ovariectomized female rats had increased levels of the NMDA NR1 subunit in cerebral cortex and hypothalamus, but not in hippocampus, whereas ethanol dependent male rats displayed significant increases in the NR1 subunit only in hippocampus. Radioligand binding analysis with [35S]TBPS found no differences in modulation of the GABA(A) receptor by neuroactive steroids between ethanol dependent male, intact female or ovariectomized female rats. Seizure susceptibility was not different between intact or ovariectomized female rats during ethanol withdrawal. We did observe differential effects on brain allopregnanolone and plasma corticosterone levels between ethanol dependent intact and ovariectomized female rats, suggesting that ovarian steroids influence HPA axis adaptations to prolonged ethanol exposure. Overall, these data suggest that ovarian steroids do not significantly impact the gender selective alterations of GABA(A) and NMDA receptors associated with ethanol dependence. PMID- 10871697 TI - Regional variations in the effects of chronic ethanol administration on GABA(A) receptor expression: potential mechanisms. AB - Gamma-aminobutyric acid type A (GABA(A)) receptors in brain adapt to chronic ethanol exposure via changes in receptor function and subunit expression. The present review summarizes currently available data regarding changes in GABA(A) receptor subunit mRNA and peptide expression. Data are presented from various different brain regions and the variations between specific brain regions used to draw conclusions about mechanisms that may underlie GABA(A) receptor adaptations during chronic ethanol exposure. In the whole cerebral cortex, chronic ethanol exposure leads to a reduction of GABA(A) receptor alpha1 subunit mRNA and peptide levels and a near equivalent increase in alpha4 subunit mRNA and peptide levels. This observation is the primary support for the hypothesis that altered receptor composition is a mechanism for GABA(A) receptor adaptation produced by chronic ethanol exposure. However, other brain regions do not display similar patterns of subunit changes. Moreover, subregions within cortex (prefrontal, cingulate, parietal, motor, and piriform) exhibit patterns of changes in subunit expression that differ from whole cortex. Therefore, regional differences in GABA(A) receptor subunit expression are evident following chronic ethanol administration, thus suggesting that multiple mechanisms contribute to the regulation of GABA(A) receptor expression. These mechanisms may include the involvement of other neurotransmitter systems, endogenous steroids and second or third messenger cross talk. PMID- 10871696 TI - GABA(A) receptor beta(2) subunit mRNA content is differentially regulated in ethanol-dependent DBA/2J and C57BL/6J mice. AB - Chronic ethanol treatment is known to alter gene expression and function of gamma aminobutyric acid type-A (GABA(A)) receptors. Here we focus on the beta(2) subunit which is widely expressed in the mammalian brain, and plays a key role in the GABA binding site. Previous studies using rodent models of ethanol dependence show either increased or no change of beta(2) subunit mRNA and peptide content following chronic ethanol administration. In humans, polymorphism at the beta(2) subunit is associated with ethanol dependence in some, but not all, populations. In the present study we measured mRNA content in the cerebellum and cerebral cortex using ethanol-naive and ethanol-dependent DBA/2J and C57BL/6J mice. The DBA/2J strain displays severe ethanol withdrawal severity, while the C57BL/6J strain shows milder withdrawal reactions. RNase protection analysis demonstrated that the DBA/2J strain is more sensitive to ethanol-induced increases in beta(2) subunit mRNA content in the cerebellum, showing significant increases at lower blood ethanol concentrations than C57BL/6J mice. The ethanol-induced regulation in C57BL/6J mice appears to be more complex, with decreases in beta(2) subunit mRNA content at low blood ethanol concentrations, and increases at higher concentrations. These data suggest that differences between C57BL/6J and DBA/2J mice in the degree of physical dependence (withdrawal) on ethanol may be related to differential sensitivity to ethanol regulation of beta(2) subunit expression. PMID- 10871698 TI - Elevated prodynorphin expression associated with ethanol withdrawal convulsions. AB - The hypothesis that kappa-opioid system activity may in part mediate convulsions exhibited during ethanol withdrawal was tested by exposing Withdrawal Seizure Prone (WSP) and Withdrawal Seizure-Resistant (WSR) mice to chronic ethanol. Whole brain was harvested for RNA isolation and prodynorphin mRNA steady-state levels in whole brain were examined using Northern blot analysis. The data revealed significantly increased levels of prodynorphin mRNA expression in mice susceptible to ethanol withdrawal convulsions after withdrawal, with no corresponding increase in prodynorphin steady-state levels in mice resistant to ethanol withdrawal convulsions. These findings were not due to basal differences in prodynorphin expression between the WSP and WSR mice. To verify that the differences observed were not due to an ethanol-induced global alteration in gene transcription, mRNA levels of the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase were measured. Glyceraldehyde-3-phosphate dehydrogenase expression was unchanged following both chronic exposure to ethanol and chronic exposure followed by withdrawal. These results extend our understanding of prodynorphin's role in generalized seizure activity to include ethanol withdrawal-induced convulsions. Our findings suggest that prodynorphin expression is modulated during ethanol withdrawal convulsions, or alternatively, prodynorphin may mediate the severity of ethanol withdrawal convulsions. PMID- 10871699 TI - Chronic ethanol has region-selective effects on Egr-1 and Egr-3 DNA-binding activity and protein expression in the rat brain. AB - This study focused on the DNA-binding activity and protein expression of the transcription factors Egr-1 and Egr-3 in the rat brain cortex and hippocampus after chronic or acute ethanol exposure. DNA-binding activity was reduced in both regions after chronic ethanol exposure and was restored to the level of the pair fed group at 16 h of withdrawal. Cortical Egr-1 protein levels were not altered by chronic ethanol exposure but increased 16 h after withdrawal, thus mirroring DNA-binding activity. In contrast, Egr-3 protein levels did not undergo any change. There was no change in the level of either protein in the hippocampus. Immunohistochemistry revealed a region-selective change in immunopositive cells in the cortex and hippocampus. Finally, an acute bolus dose of ethanol did not affect Egr DNA-binding activity and ethanol treatment did not alter the DNA binding activity or protein levels of the transcription factor Sp1. These observations suggest that chronic exposure to ethanol has region-selective effects on the DNA-binding activity and protein expression of Egr-1 and Egr-3 transcription factors in the rat brain. These changes occur after prolonged ethanol exposure and may thus reflect neuroadaptive changes associated with physical dependency and withdrawal. These effects are also transcription factor selective. Clearly, protein expression is not the sole mediator of the changes in DNA-binding activity and chronic ethanol exposure must have effects on modulatory agents of Egr DNA-binding activity. PMID- 10871700 TI - Acute ethanol administration induces changes in TRH and proenkephalin expression in hypothalamic and limbic regions of rat brain. AB - Thyrotropin releasing hormone (TRH) present in several brain areas has been proposed as a neuromodulator. Its administration produces opposite effects to those observed with acute ethanol consumption. Opioid peptides, in contrast, have been proposed to mediate some of the effects of alcohol intoxication. We measured TRH content and the levels of its mRNA in hypothalamic and limbic zones 1-24 h after acute ethanol injection. We report here fast and transient changes in the content of TRH and its mRNA in these areas. The levels of proenkephalin mRNA varied differently from those of proTRH mRNA, depending on the time and region studied. Wistar rats were administered one dose of ethanol (intraperitoneal, 3 g/kg body weight) and brains dissected in hypothalamus, hippocampus, amygdala, n. accumbens and frontal cortex, for TRH quantification by radioimmunoassay or for proTRH mRNA measurement by RT-PCR. After 1 h injection, TRH levels were increased in hippocampus and decreased in n. accumbens; after 4 h, it decreased in the hypothalamus, frontal cortex and amygdala, recovering to control values in all regions at 24 h. ProTRH mRNA levels increased at 1 h post-injection in total hypothalamus and hippocampus, while they decreased in the frontal cortex. The effect of ethanol was also studied in primary culture of hypothalamic cells; a fast and transient increase in proTRH mRNA was observed at 1 h of incubation (0.001% final ethanol concentration). Changes in the mRNA levels of proTRH and proenkephalin were quantified by in situ hybridization in rats administered ethanol intragastrically (2.5 g/kg). Opposite alterations were observed for these two mRNAs in hippocampus and frontal cortex, while in n. accumbens and the paraventricular nucleus of the hypothalamus, both mRNA levels were increased but with different kinetics. These results give support for TRH and enkephalin neurons as targets of ethanol and, as possible mediators of some of its observed behavioral effects. PMID- 10871701 TI - Effects of NMDA and ferrous sulfate on oxidation and cell death in primary neuronal cultures. AB - Excessive oxidative radical production has been implicated in a variety of neurodegerative processes including NMDA (N-methyl-D-aspartate) mediated excitotoxicity. To determine the relationship of oxidation to NMDA-receptor mediated neuronal death, we exposed rat primary cortical neuronal cultures to ferrous sulfate and the fluorescent dyes dichlorofluorescin diacetate (H(2)DCF) and propidium iodide (PI) to monitor reactive oxygen species (ROS) and cell death, respectively in the same cultures. Ferrous sulfate (FeSO(4)) caused a dose dependent increase in cellular oxidation with an ED(50) of approximately 136 microM. Levels of oxidation increased over time reaching maximum levels between 15 and 25 min. Ferrous sulfate (ED(50) approximately 241 microM) treatment for 25 min caused a delayed and progressive neuronal death that was comparable to NMDA (100 microM, 25 min) delayed neuronal death. NMDA (100 microM, 25 min) alone did not result in measurable increases of DCF fluorescence. However, when combined with 40 microM FeSO(4), NMDA dose-dependently increased H(2)DCF fluorescence. Despite the increase in DCF oxidation, combinations of FeSO(4) with NMDA did not synergize or accelerate NMDA-receptor mediated or glutamate-mediated excitotoxicity. Although excessive amounts FeSO(4) induced oxidation can cause delayed neuronal death, these findings suggest that oxidative stress is not the key factor in triggering the NMDA mediated excitotoxic cascade. PMID- 10871702 TI - Glutamate-mediated transmission, alcohol, and alcoholism. AB - Glutamate-mediated neurotransmission may be involved in the range of adaptive changes in brain which occur after ethanol administration in laboratory animals, and in chronic alcoholism in human cases. Excitatory amino acid transmission is modulated by a complex system of receptors and other effectors, the efficacy of which can be profoundly affected by altered gene or protein expression. Local variations in receptor composition may underlie intrinsic regional variations in susceptibility to pathological change. Equally, ethanol use and abuse may bring about alterations in receptor subunit expression as the essence of the adaptive response. Such considerations may underlie the regional localization characteristic of the pathogenesis of alcoholic brain damage, or they may form part of the homeostatic change that constitutes the neural substrate for alcohol dependence. PMID- 10871703 TI - The antidepressive-like effect of oxcarbazepine: possible role of dopaminergic neurotransmission. AB - It has been previously shown that oxcarbazepine (OXCBZ), a keto-analogue of carbamazepine, exhibits an antidepressive-like effect profile in the learned helplessness and forced swimming test (FST). Since carbamazepine possesses dopaminergic effect, the present study was carried out to evaluate the extent to which the antidepressive effect of OXCBZ might be mediated by dopaminergic system. Thus, the effects of OXCBZ in haloperidol-induced catalepsy and apomorphine-induced stereotypy were studied. The anti-immobility effect of OXCBZ in the FST was also evaluated in haloperidol pre-treated rats. OXCBZ (40 and 80 mg/kg, i.p.) dose-dependently reduced the catalepsy induced by haloperidol (2.0 mg/kg, i.p.). Moreover, OXCBZ (80 mg/kg, but not 20 or 40 mg/kg, i.p.) increased the intensity of apomorphine-induced stereotypy (0.6 mg/kg, s.c.). Finally, it was observed that the combination of OXCBZ (80 mg/kg, i. p.) and haloperidol (0.5 mg/kg, i.p.) antagonized the anti-immobility effect of OXCBZ and further increased the immobility time when compared to haloperidol alone. Haloperidol alone (0.5 or 1. 0 mg/kg) did not change the immobility time. Thus, these results suggest that OXCBZ could enhance dopaminergic neurotransmission, which might mediate its antidepressive-like effect. PMID- 10871704 TI - Subchronic fluoxetine administration to rats: effects on 5-HT autoreceptor activity as measured by in vivo microdialysis. AB - Subchronic administration of fluoxetine to rats has been shown to induce subsensitivity of presynaptic 5-HT(1A) and 5-HT(1B) autoreceptors, and also postsynaptic 5-HT(1A) receptors in the hypothalamus. We investigated the effects of administration of fluoxetine (10 mg/kg i.p.) to rats for 6 days on presynaptic 5-HT(1A) receptor activity in the hypothalamus, postsynaptic 5-HT(1A) receptor activity in the hippocampus, and presynaptic 5-HT(1B) autoreceptor activity in both areas, using in vivo microdialysis. The effect of the 5-HT(1B/1D) antagonist (N-[4-methoxy-3-(4-methyl-1-piperizinyl)phenyl]-2'-methyl-4'-(5- methyl-1,2,4 oxadiazole-3-yl)[1,1'-biphenyl]-carboxamide (GR 127935) (5 mg/kg s.c.) to elevate 5-hydroxytryptamine (5-HT) levels was reduced in hippocampus but not hypothalamus of fluoxetine-treated rats. Fluoxetine did not alter either presynaptic 5-HT(1A) autoreceptor activity, as measured by the effect of injection of 8-hydroxy-2(di-n propylamino)tetralin (8-OH-DPAT) (0.2 mg/kg or 50 microg/kg s.c.) on 5-HT levels in the hypothalamus, or postsynaptic 5-HT(1A) receptor activity, as measured by the effect of 8-OH-DPAT (0.2 mg/kg s.c.) on cyclic AMP accumulation, in the hippocampus. PMID- 10871705 TI - N(6)-2-(4-aminophenyl)ethyl-adenosine enhances the anticonvulsive action of conventional antiepileptic drugs in the kindling model of epilepsy in rats. AB - APNEA [(N(6)-2-(4-aminophenyl)ethyl-adenosine; a non-selective adenosine A(3) receptor agonist; 2-4 mgkg(-1)] had no significant effect on seizure parameters (seizure severity, seizure duration and afterdischarge duration) in amygdala kindled rats. Subsequently, APNEA was combined with antiepileptic drugs administered at doses ineffective in fully kindled rats. Co-administration of APNEA (0.5-2 mg kg(-1)) with carbamazepine (2.5-20 mg kg(-1)) resulted in the significant reduction of all studied seizure parameters. Moreover, 8-cyclopentyl 1,3-dimethylxanthine 8-CPX (a selective adenosine A(1) receptor antagonist; 5 mg kg(-1)) partially reduced the anticonvulsive activity of a combination of APNEA (2 mg kg(-1)) with carbamazepine (20 mg kg(-1)), but not that of carbamazepine (20 mgkg(-1))+APNEA (0.5 mg kg(-1)). When APNEA (2 mg kg(-1)) was combined with phenobarbital (20 mg kg(-1)), valproate (75 mg kg(-1)) or clonazepam (0.003 mg kg(-1)), seizure and afterdischarge durations were significantly shortened. 8-CPX (5 mg kg(-1)) totally reversed the APNEA (2 mg kg(-1))-induced enhancement of the anticonvulsive action of valproate. However, when the non-selective adenosine A(3) receptor agonist was administered together with diphenylhydantoin, no protection was observed in the kindling model of epilepsy. The interaction at the pharmacokinetic level can be excluded because APNEA did not interfere with the free plasma level of antiepileptics used in this study. It may be concluded that the interaction of APNEA with carbamazepine involves A(3) adenosine receptor dependent events. PMID- 10871706 TI - Effects of atypical antipsychotic drugs on dopamine output in the shell and core of the nucleus accumbens: role of 5-HT(2A) and alpha(1)-adrenoceptor antagonism. AB - The effects of acute intravenous administration of several new, atypical antipsychotic drugs (APDs): olanzapine (0.05 and 1.0 mg/kg), sertindole (0.1 and 1.0 mg/kg) and quetiapine (0.25 and 2.5 mg/kg), a selective 5-HT(2A) receptor antagonist, M100907 (0.03 and 0.3 mg/kg), and an alpha(1)-adrenoceptor antagonist, prazosin (0.3 mg/kg), on regional dopamine output were examined in the two subdivisions of the nucleus accumbens (NAC), the core and shell, which seem associated with motor control and limbic functions, respectively, by using in vivo differential normal pulse voltammetry in anaesthetised, pargyline pretreated rats. Both quetiapine and sertindole, in the two doses used, caused a more pronounced dopamine release in the shell than in the core region of the NAC. In contrast, the low dose of olanzapine increased dopamine output almost to the same extent in both regions, whereas the high dose increased dopamine output to a greater extent in the core. M100907 selectively increased dopamine output in the shell. Also, prazosin significantly increased dopamine output in the shell, but not in the core. The results indicate that both 5-HT(2A) and alpha(1) adrenoceptor antagonism may play an important role in the preferential effect of atypical APDs on dopamine output in the shell versus the core of the NAC. PMID- 10871708 TI - Unaltered alpha(2)-noradrenergic/imidazoline receptors in suicide victims: a postmortem brain autoradiographic analysis. AB - In vitro quantitative autoradiography of alpha(2)-adrenergic/imidazoline receptors, using [(125)I]iodoclonidine as a ligand, was performed on 24 human brains postmortem. Twelve brains were obtained from suicide victims and 12 from matched controls. We found no significant, region-dependent alterations in the density of alpha(2)-adrenergic receptors in brains of suicide victims as compared to matched controls. We also report age-dependent reductions in binding in the prefrontal cortex and hippocampus, as well as significant recent alcohol ingestion-dependent reductions in binding in the prefrontal cortex. Sex and time from death to autopsy did not affect iodoclonidine binding in our sample. PMID- 10871707 TI - Effects of nefazodone on the immune system of mice. AB - Mice exposed to a chronic auditory stressor and treated with nefazodone (10 mg/kg/day s.c.), showed a reduction in stress-induced suppression of thymus and spleen cellularity, and in peripheral T-Iymphocyte population. The in vitro blastogenic response of spleen lymphoid cells to mitogen concanavalin A, the in vitro and in vivo activity of phagocytosis, both measured using the zymosan and carbon clearance tests, respectively, were also assessed and nefazodone was found to partially reverse the inhibitory effect of stress on those parameters. Nefazodone did not significantly affect those parameters in unstressed mice. In conclusion, this report provides evidence on the immunoprotective effects of this novel antidepressant drug against the adverse effects of stress in mice. PMID- 10871709 TI - The GABA-B antagonist CGP 36742 prevent PTZ-kindling-provoked amnesia in rats. AB - Deficit in active and inhibitory avoidance behaviour has been found in pentylenetetrazole (PTZ)-kindled rats. This supports the view that memory deficit is an integral part of epilepsy. In the present study we examined the effect of the GABA B antagonist CGP 36742 on memory deficit induced by PTZ-kindling in shuttle-box- and step-down-trained rats. The retention in CGP 36742-treated animals was significantly improved compared to the kindled controls. The mechanisms of action of CGP 36742 is considered. The favourable effect of the GABA B antagonist in cases of amnesia provoked by PTZ-kindling might be of interest in clinical practice. PMID- 10871710 TI - Oral administration of losartan influences aminopeptidase activity in the frontal cortex. AB - Although there is a brain renin-angiotensin system, its mechanisms of control are not fully understood. We studied the effect of oral administration of the AT(1) receptor antagonist losartan on brain aminopeptidase (AP) activity, which plays a major role in neuropeptide metabolism. Six AP activities, related and non-related with the angiotensin (Ang) metabolism, were measured in their soluble and membrane-bound forms in the frontal cortex of control animals and rats treated with losartan, chronically administered via the drinking water. The results demonstrate that soluble pGluAP and membrane-bound AspAP and GluAP increased significantly in losartan-treated animals, indicating that the blockade of the AT(1) receptor stimulates the activity of AP involved in the Ang metabolism. Moreover, the blockade of the AT(1) receptor induces changes not only in the brain angiotensin metabolism, but probably also in that of other neuropeptides. PMID- 10871711 TI - Altered brain protein kinase C in depression: a post-mortem study. AB - [(3)H]Phorbol 12,13-dibutyrate (PDBu) binding was measured in soluble and particulate fractions of frontal cortex and hippocampus from suicides, with a firm retrospective diagnosis of depression, and individually matched controls. Suicides were divided into those who had been free of antidepressant drugs for at least 3 months and those in whom prescription of antidepressants was clearly documented. In frontal cortex, there was a significantly higher number (by 75%) of [(3)H]PDBu binding sites in the soluble fraction in antidepressant-free suicides compared to controls; significant differences were also seen in the proportion of sites in the soluble and particulate fractions. Higher numbers of [(3)H]PDBu binding sites in the particulate fraction of hippocampus in antidepressant-free suicides was restricted to those who died by violent means. No significant differences in the number of [(3)H]PDBu binding sites were found in antidepressant-treated suicides compared to controls. This study provides evidence for the involvement of protein kinase C in the pathophysiology of depression. PMID- 10871712 TI - Acute psychological and physiological effects of MDMA ("Ecstasy") after haloperidol pretreatment in healthy humans. AB - 3,4-Methylenedioxymethamphetamine (MDMA, "Ecstasy") releases serotonin and dopamine. The role for dopamine in mediating the effects of MDMA has not yet been examined in humans. We investigated the effect of pretreatment with the dopamine D(2) antagonist haloperidol (1.4 mg i.v.) on psychological and physiological responses to MDMA (1.5 mg/kg p.o.) in 14 healthy volunteers using a double-blind placebo-controlled within-subject design. Subjective peak effects were rated using standardised scales. The physiological effects measured were blood pressure, heart rate and body temperature. Side effects were assessed during the session, and after 1 and 3 days. Haloperidol attenuated MDMA-induced positive and mania-like mood but had no reducing effect on other subjective changes or on cardiovascular effects. Results are consistent with a partial dopaminergic mediation of the euphoriant effects of MDMA. In contrast, dopamine does not seem to contribute to the physiological effects of MDMA, indicating a role for serotonin and norepinephrine. PMID- 10871713 TI - ECNP consensus meeting, March 5-6, 1999, Nice. Post traumatic stress disorder: guidelines for investigating efficacy of pharmacological intervention. ECNP and ECST. PMID- 10871714 TI - Body image dimensions and cancer: a heuristic cognitive behavioural model. AB - The term body image has been associated with a multitude of definitions within psychosocial oncology. It is well known that cancer and cancer treatments often have a negative impact on appearance-related variables. A growing literature has emerged in recent years on the psychological aspects of changed appearance. This work has mainly addressed weight-related appearance and the psychology of eating disorders. A number of themes have emerged from this work. These themes have been strongly influenced by a cognitive behavioural perspective. There seems, however, to have been few attempts to integrate findings from such work with attempts to understand cancer-related appearance changes. This paper outlines some of the key developments within body image psychology and suggests a heuristic cognitive behavioural model that could be applied to the assessment, conceptualisation and treatment of body image disturbance among cancer patients. PMID- 10871715 TI - Factors implicated in the decision whether or not to join the tamoxifen trial in women at high familial risk of breast cancer. AB - Why, given similar medical circumstances-high familial risk of breast cancer-will some women elect to join a trial of drugs designed to reduce that risk but others choose not to take part? The aim of this study was to identify measurable differences between women who elect to join a placebo-controlled, double-blind randomised trial of the drug tamoxifen and women who elect not to join. One hundred and six women attending a breast care clinic completed questionnaires covering demographic details, health locus of control, perception of risk and adequacy of medical communication. All were eligible for inclusion in the tamoxifen trial. Only half (n=53) of the sample elected to join, the other half (n=53) declined. Those who declined the trial were significantly more aware of lifestyle factors thought to influence the development of cancers-diet, exercise and oestrogen-prolonging activities (p<0.001), and they also appeared to find the information given by the hospital about tamoxifen harder to understand than did the group who had joined the trial (p=0.01). They could think of significantly fewer positive reasons for joining (p<0.001) and were significantly younger (p=0.001). Participants in both groups significantly overestimated the relative risks of breast cancer. The average estimation of risk for women aged 30 with a family history of breast cancer was 22 times higher than that given by their consultant. This gives rise to considerable concerns about the information underpinning informed consent. PMID- 10871716 TI - Impact of computerized quality of life screening on physician behaviour and patient satisfaction in lung cancer outpatients. AB - The purpose of this paper was to determine if providing patient specific Quality of Life (QL) information to clinic staff before a clinic appointment improved patient care in a lung cancer outpatient clinic. Patients were sequentially assigned to either a usual care control group or the experimental group, which completed a computerized version of the European Organization for Research and Treatment of Cancer (EORTC) QLQ-C30 questionnaire in order to provide the clinic staff with QL information prior to the clinic appointment. The control group completed the EORTC QLQ-C30 paper version after the clinic appointment. Outcome measures were patient satisfaction, the degree to which issues identified on the QL questionnaire were addressed in the appointment, and a chart audit, which measured charting of QL issues and actions taken by the clincian relating to QL. In the experimental group, more QL issues identified by the patient on the EORTC QLQ-C30 were addressed during the clinic appointment than in the control group. As well, marginally more categories were charted and a trend towards more actions being taken was seen in the experimental group. Patients reported being equally and highly satisfied with the treatment in both groups. The clinical implication is that the computerized administration of the EORTC QLQ-C30 questionnaire and providing staff with a report highlighting patient-specific QL deficits is a simple, time-effective and acceptable means of improving patient-provider communication in a busy outpatient clinic. Large trials studying its effectiveness in different patient populations and regions would further elucidate the nature of this effect and potentially improve the overall quality of care that patients receive. PMID- 10871717 TI - Surviving cancer; what does it mean for you? An evaluation of a clinic based intervention for survivors of childhood cancer. AB - BACKGROUND: To evaluate a clinic based intervention designed to improve attitude to follow-up, increase self-efficacy or confidence to care for health, and raise awareness of possible vulnerability to future health issues among survivors of childhood cancer. The intervention included an information booklet, treatment summary and separate information sheets, which were explained to survivors as part of routine follow-up care. PROCEDURE: Survivors (n=263; mean age=21 years; >5 years since diagnosis) attending one of seven United Kingdom Children's Cancer Study Group (UKCCSG) late-effects follow-up clinics completed questionnaires before and after the intervention. Outcome measures included self-ratings of importance of follow-up, readiness to change behaviour, self-efficacy, perceived vulnerability, and ratings of informativeness and emotional content of the written material. RESULTS: Responders were more likely to be female than non responders, held more positive views about the importance of follow-up and perceived themselves to be more vulnerable to health risk. After the intervention, responders reported that they were more prepared to change their behaviour, had increased self-efficacy, but also perceived themselves to be more vulnerable to future health problems. CONCLUSIONS: We conclude that the intervention is a relatively simple way to enhance awareness among survivors about the importance of follow-up and need for vigilance in their healthcare. Difficulties in recruiting survivors who failed to attend are considered. PMID- 10871718 TI - Physical, psychological and social well-being of women with breast cancer: the influence of disease phase. AB - While research exists on the well-being of women during a specific phase of breast cancer, little research exists in which researchers utilized the same instruments to examine differences in women's well-being, based on the phase of their breast cancer. Using a trajectory framework, the purpose of this study is to examine the differences in the physical and social well-being of women during the following breast cancer states: newly diagnosed, adjuvant therapy, stable disease and recurrent disease. The convenience sample consisted of 35 women newly diagnosed with breast cancer, 52 women with breast cancer undergoing adjuvant therapy, 84 women whose breast cancer was considered stable and 64 women with recurrent breast cancer. Participants completed a packet of questionnaires which contained a demographic questionnaire, Short Form-36 (SF-36) Health Survey, a researcher designed (RD) questionnaire, Cancer Rehabilitation Evaluation System Short Form (CARES-SF) and the Brief Symptom Inventory (BSI). Descriptive statistics, analysis of variance, and general linear F-tests were used to analyze the data. Differences were found across phases of disease on various subscales, including those representing perceived health states, overall impact, medical interactions, physical function, role function, fatigue, pain, social function and satisfaction with health. No significant differences were found between groups on the BSI subscales with the exception of somatization, global psychosocial measures, sexual and marital relation subscales. While individuals with recurrent disease often experienced more difficulties with their well-being than women in the other groups, women newly diagnosed and in the adjuvant group experienced more difficulties in select areas of well-being when compared with women in the stable group. Health care professionals need to recognize differences between groups to better meet the needs of patients with a breast cancer diagnosis. PMID- 10871719 TI - Couples dealing with cancer: role and gender differences regarding psychological distress and quality of life. AB - The goal of the present study was to further knowledge on gender and role (i.e. patient versus partner) differences in psychological distress and quality of life as a consequence of dealing with cancer. There is some evidence that being the patient or the caregiver makes more difference for men than for women. In total, 173 couples facing various forms of cancer (two samples) and a control group of 80 couples completed the CES-D and Cantril's Ladder. Analyses of variance revealed that both female patients and female partners of patients perceived more psychological distress and a lower quality of life than women in healthy couples. In contrast, role did have an effect on men. Specifically, male patients scored as high on psychological distress and as low on quality of life as female patients and female partners, but psychological distress and quality of life did not differ between male partners of patients and their healthy controls. However, this effect was found in only one patient sample. The finding that female partners perceived more psychological distress and a lower quality of life than male partners could not be accounted for by differences in the physical condition of the patient or the partner. PMID- 10871720 TI - A supportive-expressive group intervention for women with a family history of breast cancer: results of a phase II study. AB - BACKGROUND: Evidence suggests that there are significant psychological and behavioural sequelae associated with having a family history of breast cancer (BC) which can interfere with comprehension of risk estimates. PURPOSE: The purpose of this study was to develop, standardize and do preliminary testing of a group intervention designed to address the emotional impact of having a family history of BC. METHODS: This study is a single-arm pilot design with pre- and post-measures of perceived risk, psychosocial distress, knowledge and screening practices. RESULTS: The primary study outcome measure of risk comprehension was significantly improved by 70%, according to our predetermined criteria for success. In addition, the most important secondary measures of psychosocial functioning, such as cancer-related distress (p=0.025), depression (p=0.05), anxiety (p=0.005) and unresolved grief (p=0.034) were significantly improved. CONCLUSION: The results of this initial pilot study are encouraging; however, further research is required, using a randomized controlled study design to evaluate the relative contribution of this intervention to the successful modification of risk comprehension, enhanced psychological functioning, and to promote optimal screening adherence. PMID- 10871721 TI - Panic disorder in hospitalized cancer patients. AB - Cancer is commonly complicated by psychiatric comorbidity, particularly depression. However, the effects of panic on cancer treatment and cancer patients' quality of life are not well understood. To examine more closely the occurrence of panic attacks and panic disorder in cancer patients, we retrospectively reviewed charts of 106 consecutive psychosomatic consultations of inpatients with cancer at a regional cancer center. Approximately one-fifth of the patient sample presented with panic attacks or panic disorder at the time of the consultation. We present four case examples to demonstrate the potential effect of panic on cancer patients, including requests for cancer treatment discontinuation. We report resolution of panic with benzodiazepine and selective serotonin reuptake inhibitors. PMID- 10871722 TI - A dramatic loss of non-verbal intelligence following a right parietal ependymoma: brief case report. AB - Virtually all reports on the effects of focal brain lesions upon specific neuropsychological functions are based upon estimates of cognitive loss in persons following a lesion, without proof of premorbid capacity. This report presents the case of a 19-year-old left-handed male with assessment of Verbal and Performance I.Q. testing 1 year prior to a subsequent brain tumor for reasons unrelated to the neoplasm. The patient was then re-tested 23 months later, following the diagnosis and treatment of a supratentorial ependymoma in the right parietal region. His multi-modal treatment regimen included a partial surgical resection of the tumor, cranio-spinal irradiation with focal boost to the primary site, and chemotherapy. Results demonstrate a striking 58-point decline in the Wechsler Adult Intelligence Scale - Revised (WAIS - R) Performance I. Q. with no significant change in Verbal I.Q. These findings clearly document the important cognitive functions associated with the right parietal lobe. PMID- 10871723 TI - Clinical update. PMID- 10871724 TI - Analysis of prognostic factors for allogeneic marrow transplantation following busulfan and cyclophosphamide in myelodysplastic syndrome and after leukemic transformation. AB - Prognostic factors in 42 patients aged 11 to 62 (median 46) years, with myelodysplastic syndrome (MDS) or after leukemic transformation, who underwent allogeneic marrow transplantation between 1984 and 1999 were analyzed. Thirty-six had advanced disease morphology; 19 had leukemic transformation. Twenty-nine received a preparative regimen of BuCy2 and 13 busulfan 14 mg/kg, etoposide 50 mg/kg and cyclophosphamide 120 mg/kg. Severe hepatic veno-occlusive disease (VOD) occurred in three patients all of whom received anti-leukemic chemotherapy prior to transplantation. Fifteen patients (36%) died from early transplant-related complications; nine patients relapsed. The estimated 4 year disease-free survival (DFS) was 35% (95% CI 26-44%). Older age was the most significant adverse prognostic factor. Patients with leukemic transformation who underwent early transplantation had significantly better DFS than those treated first with chemotherapy (P = 0.002). Delayed toxicity was rare in these patients; no late relapses occurred. Bone Marrow Transplantation (2000) 25, 1219-1222. PMID- 10871725 TI - Allogeneic peripheral blood stem cell transplantation following CD34+ enrichment by density gradient separation. AB - GVHD is a significant cause of morbidity and mortality following allogeneic peripheral blood stem cell transplantation (AlloPBSC). CD34+ cell selection could reduce GVHD by negative selection of T cells. In an attempt to reduce the T cell content of alloPBSC we carried out a trial in which 11 patients with hematologic malignancies received alloPBSC from HLA-matched siblings following density gradient separation using an isotonic colloidal silica solution (BDS 60; Dendreon Corporation). Cyclosporine and methylprednisone were used for GVHD prophylaxis. The mean yield of CD34+ cells was 69 +/- 15.6% with a purity of 2.9 +/- 1.7%. The mean number of CD3+ cells infused was 1.0 +/- 1.2 x 107/kg, representing a 1.3 log depletion. A high risk of acute GVHD was observed: grade II-IV in 7/11 (64%) and grade III-IV GVHD in 5/11 (45%) patients. Nine of the 11 (82%) patients died with a median survival of 68 days. Cytokine expression in PBSC was compared pre and post processing. Interferon-gamma was detected only following density gradient separation while IL-8 expression increased 3- to 6-fold post processing. Therefore, processing with this device may augment production of pro-inflammatory cytokines. Bone Marrow Transplantation (2000) 25, 1223-1228. PMID- 10871726 TI - Allogeneic peripheral blood stem cell transplantation for standard risk leukemia: experience of Ibni Sina Hospital. AB - Fifty-three patients with standard risk leukemia who underwent allogeneic peripheral blood stem cell transplantation (alloPBSCT) from their HLA-identical siblings were analyzed for engraftment, incidence and severity of GVHD, and relapse rate. Standard risk leukemia was defined as AML in first complete remission or CML in first chronic phase within the first year after diagnosis. The median age was 34.5 years (range 13-47). Stem cells were mobilized by using 10 microg/kg G-CSF subcutaneously for 5 days. A median of 5. 7 (2.1-21.4) x 106/kg CD34+ cells was collected over a median of 2 (range 1-5) apheresis procedures. Cyclosporin A (CsA) plus short-course MTX were used for GVHD prophylaxis. Recovery to granulocytes >0.5 x 109/l and platelets >20 x 109/l occurred at a median of day +13 (range 8-32) and +13 (range 8-51), respectively. Day +100 transplant-related mortality was 13.2% (7/53). Acute GVHD occurred in 20 of 49 (41%) evaluable patients and only six (12.3%) of them had severe disease (grade III-IV). Chronic GVHD occurred in 30 of 42 (71.4%) evaluable patients. Relapse rate at 2 years was 7. 5%. The median overall and leukemia-free survivals were 22 (4-44) and 20 (3-44) months, respectively. Estimated 4 year leukemia-free and overall survival rates were 60% and 62%, respectively. In conclusion, alloPBSCT in standard risk leukemia seems to be associated with a low relapse rate and no increased risk of acute GVHD, but there is a trend for higher incidence of cGVHD. Bone Marrow Transplantation (2000) 25, 1229-1232. PMID- 10871727 TI - Clinical outcome of breast and ovarian cancer patients treated with high-dose chemotherapy, autologous stem cell rescue and THERATOPE STn-KLH cancer vaccine. AB - The purpose of this study was to evaluate the toxicity and potential efficacy of administering the THERATOPE STn-KLH cancer vaccine to ovarian and breast cancer patients after an autologous stem cell transplant. Forty patients (11 high-risk stage II/III breast cancer, 22 stage IV breast cancer, and seven stage III/IV ovarian cancer patients) were treated with high-dose chemotherapy followed by autologous/syngeneic stem cell rescue and vaccination with THERATOPE STn-KLH (Sialyl-Tn-KLH with Detox-B Stable Emulsion). Each patient was scheduled to receive a total of five vaccinations beginning on days 30-151 after stem cell infusion. The vaccine was well tolerated. Induration and erythema at the site of injection were the most common side-effects. When one compares the outcome of patients vaccinated with 66 breast and ovarian cancer patients who were not, following risk-adjustment analysis, vaccinated patients appeared more likely to survive (P = 0.07) and less likely to relapse (P = 0. 10). Vaccinated patients with the greatest specific lytic activity against STn+OVCAR tumor cells relative to nonspecific killing of Daudi cells tended to remain in remission longer than patients who displayed less specific immune activity (P = 0.057). We conclude that the THERATOPE STn-KLH cancer vaccine is well tolerated in breast and ovarian cancer patients after autologous transplant and, while not statistically significant, the trends in data support the concept that THERATOPE vaccine may decrease the risk for relapse and death and thus warrants further study. Bone Marrow Transplantation (2000) 25, 1233-1241. PMID- 10871728 TI - Autotransplantation following busulfan, etoposide and cyclophosphamide in patients with non-Hodgkin's lymphoma. AB - The purpose of the study was to determine the toxicities and effectiveness of a novel preparative regimen of busulfan (Bu) 14 mg/kg, etoposide 50 or 60 mg/kg, and cyclophosphamide (Cy) 120 mg/kg in non-Hodgkin's lymphoma (NHL) and to analyze results using doses based on different body weight parameters and the two different etoposide doses. Three hundred and eighty-two patients aged 16 to 72 underwent first autologous transplantation with mobilized peripheral blood progenitor cells between August 1992 and December 1998 at either of two transplant centers. Mucositis was the most common toxicity. Hepatic toxicity was the most common life-threatening toxicity; severe hepatic VOD occurred in 11 patients (2.9%). Ten patients (2.6%) died from treatment-related toxicity. The 3 year progression-free survival (PFS) for the entire group was 46.9% (95% CI, 40.5 53.3%). Elevated LDH, resistance to chemotherapy, and intermediate/aggressive histology were significant adverse prognostic factors. For patients in sensitive first relapse PFS was 47.0% (95% CI, 37-57%). Neither etoposide dose nor body weight parameter utilized significantly affected outcome. In conclusion, the novel preparative regimen of Bu, etoposide and Cy results in a low incidence of treatment-related mortality and is effective in the treatment of patients with NHL. Bone Marrow Transplantation (2000) 25, 1243-1248. PMID- 10871729 TI - Recovery of lymphocyte and dendritic cell subsets after autologous CD34+ cell transplantation. AB - Following high-dose chemotherapy (HDC) and peripheral blood progenitor cell transplantation (PBPCT), there are profound changes in leukocyte homeostasis and the immune system is compromised. Transplantation of purified CD34+ cells may further compromise immune recovery because the grafts are depleted of mature immune cells. However, a detailed monitoring of immune cell reconstitution has not been done. We monitored blood levels of antigen-presenting cells (APC) and of lymphocytes by multi-color flow cytometry at different times post CD34+ PBPCT. We found a rapid normalization of circulating levels of the antigen-presenting CD11c+ dendritic cells (defined as lineage- HLA-DR+ CD11c+ cells). There was a slight over-representation of lin- DR+ CD11c- cells at day 42 post transplantation suggesting that the composition of the APC population might be affected. Normal levels of total T, B and NK lymphocytes were rapidly achieved but the composition of the T cell population was abnormal. Patients had elevated levels of CD8+ T cells at early times and a persistent reduction in levels of naive CD8+ T cells (CD8+ CD4- CD45RA+ CD27+) and of naive CD4+ T cells (CD4+CD3+ CD8- CD45RA+). Thus, we found a rapid recovery of DC after CD34+ PBPCT but the specific numerical defects in naive T cells are likely to be a major cause of immune dysfunction in the patients. Bone Marrow Transplantation (2000) 25, 1249 1255. PMID- 10871730 TI - Diagnosis of toxoplasmosis in bone marrow transplant recipients: comparison of PCR-based results and immunohistochemistry. AB - Toxoplasmosis in bone marrow transplant recipients is a rare but serious complication and if untreated, almost uniformly fatal. The diagnosis, however, remains difficult. We therefore compared serial determination of antibody titers specific for T. gondii before and after transplantation, serial PCR for T. gondii DNA in serum, PCR and nested PCR for T. gondii DNA in various tissues, conventional histology and immunohistochemistry for detection of parasites in three patients with autopsy-confirmed toxoplasmosis after bone marrow transplantation. Immunohistochemistry demonstrated the presence of parasites in 13 out of 20 organs investigated (65%), whereas PCR detected T. gondii-specific DNA in 15 out of 20 organs (75%). Immunohistochemistry revealed concordant results to PCR data in 60% of the specimens. With the use of a nested PCR protocol, eight out of nine samples (89%) were positive for T. gondii-specific DNA. The combination of both methods detected the presence of parasites in 90% of the specimens. Serial PCR in serum did not yield positive results. Neither PCR nor immunohistochemistry was able to detect parasites in all organs investigated, but both methods together improved sensitivity to 90% and consequently, should be used jointly to maximize diagnostic precision. Bone Marrow Transplantation (2000) 25, 1257-1262. PMID- 10871731 TI - Non-infectious lung complications are closely associated with chronic graft versus-host disease: a single center study of incidence, risk factors and outcome. AB - Non-infectious lung complications (NILC) are frequent, influencing morbidity and mortality of patients after allogeneic BMT. Although the term NILC encompasses a number of different entities, an association with GVHD has been noted for almost all of them. Our study was directed towards assessing the incidence and risk factors for developing NILC, as well as the response to treatment and long-term outcome. Forty (14.7%) out of 272 patients surviving for more than 3 months after allogeneic BMT, developed lung complications fulfilling the criteria for NILC. The evaluation was based on clinical investigation, radiologic imaging, lung function tests, broncho-alveolar lavage and biopsies. Risk factors were assessed by univariate and multiple statistical regression models, where chronic GVHD proved to be the only significant risk factor for the development of NILC (P = 0.011). In three patients NILC developed in direct association with donor lymphocyte infusions. The majority of patients responded well to treatment with corticosteroids and immunosuppressive drugs. NILC had no adverse effect on survival. The frequency of NILC was low in autologous (5%) as compared with allogeneic transplants (14.7%) but this difference was not statistically significant. Bone Marrow Transplantation (2000) 25, 1263-1268. PMID- 10871732 TI - Mucosal barrier injury: biology, pathology, clinical counterparts and consequences of intensive treatment for haematological malignancy: an overview. AB - Mucositis is an inevitable side-effect of the conditioning regimens used for haematopoietic stem cell transplantation. The condition is better referred to as mucosal barrier injury (MBI) since it is primarily the result of toxicity and is a complex and dynamic pathobiological process manifested not only in the mouth but also throughout the entire digestive tract. A model has been proposed for oral MBI and consists of four phases, namely inflammatory, epithelial, ulcerative and healing phases. A variety of factors are involved in causing and modulating MBI including the nature of the conditioning regimen, the elaboration of pro inflammatory and other cytokines, translocation of the resident microflora and their products, for example, endotoxins across the mucosal barrier, exposure to antimicrobial agents and whether or not the haematopoietic stem cell graft is from a donor. Neutropenic typhlitis is the most severe gastrointestinal manifestation of MBI, but it also influences the occurrence of other major transplant-related complications including acute GVHD, veno-occlusive disease and systemic infections. The pathobiology, clinical counterparts and the means of measuring MBI are discussed together with potential approaches for prevention, amelioration and, perhaps, even cure. Bone Marrow Transplantation (2000) 25, 1269 1278. PMID- 10871733 TI - Granisetron (Kytril) plus dexamethasone for antiemetic control in bone marrow transplant patients receiving highly emetogenic chemotherapy with or without total body irradiation. AB - This prospective trial evaluated the efficacy and toxicity of granisetron for antiemetic control in patients receiving high-dose cyclophosphamide (CY) containing regimens with/without TBI for bone marrow (BM) or peripheral blood stem cell (PBSC) transplantation or PBSC mobilization. Granisetron 1 mg i.v. plus dexamethasone 10 mg i. v. were administered daily 30 min before chemotherapy or radiation for a median of 5 days. Response was defined as the number of emetic episodes per 24 h: complete response, 0 and no emetic rescue; major response, 1 2; minor response, 3-5; failure, >5. One hundred patients were enrolled. Ninety eight received CY-containing regimens and 26 of these additionally received TBI (12 Gy divided over 4 days). Response was complete on 216 (47%) of a total 456 patient days, major on 222 (49%), minor on 14 (3%), and failure on 4 (1%). Mean number of emetic episodes per patient per day and breakthrough medication required per patient per day was 0.24 (range 0-8) and 0. 40 (range 0-8), respectively. Adverse effects were minimal, with headache (20%) reported most frequently. Based on these results, granisetron plus dexamethasone is an effective and well-tolerated antiemetic regimen in BMT/PBSCT recipients conditioned with high-dose chemotherapy with/without TBI. Bone Marrow Transplantation (2000) 25, 1279-1283. PMID- 10871734 TI - Neurological events associated with the infusion of cryopreserved bone marrow and/or peripheral blood progenitor cells. AB - Reports of neurological toxicity of cryopreserved stem cell infusion are infrequent. Three of 179 consecutive patients experienced significant neurological events in this context. Transient global amnesia developed following infusion in one patient and in the other two, cerebral infarction occurred. Profound hypotension, bradyarrhythmias or hypoxia were not associated with any of these episodes. These events may have been related to infused DMSO, which in the non-transplant setting has been associated with neurological toxicity and local infusion of which has resulted in acute vasospasm in animal models. These cases suggest that infusion of cryopreserved stem cells may result in cerebrovascular ischaemia. Bone Marrow Transplantation (2000) 25, 1285-1287. PMID- 10871735 TI - Effects of bone marrow transplantation on the cardiovascular abnormalities in canine mucopolysaccharidosis VII. AB - The genetic mucopolysaccharidoses (MPS) are a family of lysosomal storage diseases resulting from defective catabolism of glycosaminoglycans (GAGs). Echocardiographic abnormalities in dogs with MPS type VII (Sly syndrome, beta glucuronidase deficiency) included mitral valve thickening and insufficiency, large aortic dimensions in both the long and short axes, and thickened aortic valves. Grossly, at post mortem examination, there was nodular thickening of the mitral valve, a prominent ductus diverticulum, and a dilated aorta with thickened walls. Histologically, cytoplasmic vacuolation was seen in cells of the mitral valves, coronary arteries, and aorta. By electron microscopy, the cells of the mitral valve were packed with electron-lucent cytoplasmic vacuoles. The mean residual activity of beta-glucuronidase in the aorta and myocardium was <1% of normal, the mean hexosaminidase A activity >2. 5 times normal, and the mean GAG concentrations more than twice normal. In three MPS VII dogs that received heterologous BMT at 6 weeks of age, the echocardiographic abnormalities were improved, and the histopathologic and ultrastructural pathology was reduced. In the aorta and myocardium, the mean beta-glucuronidase activity of the BMT group was 4.5% and 11% of normal, respectively, and the hexosaminidase A activity and GAG concentrations were normalized. Bone Marrow Transplantation (2000) 25, 1289 1297. PMID- 10871736 TI - Severe respiratory depression after dimethylsulphoxide-containing autologous stem cell infusion in a patient with AL amyloidosis. AB - Adverse reactions with DMSO-cryopreserved stem cell infusion are well-recognized. However, severe, life-threatening anaphylactic reactions with DMSO are very rarely described in the literature. We report here a 58-year-old female with AL amyloidosis who developed an unexpected episode of respiratory arrest a few seconds after the beginning of thawed stem cell product infusion. Fortunately, the patient was resuscitated successfully without the need for intubation. The prompt development of the reaction just a few seconds after the stem cell infusion convincingly implicates DMSO as the potential suspect. The presence of amyloid cardiomyopathy might have also contributed to this adverse event. Bone Marrow Transplantation (2000) 25, 1299-1301. PMID- 10871738 TI - Prolonged remission state of refractory adult onset Still's disease following CD34-selected autologous peripheral blood stem cell transplantation. AB - We report a 38-year-old patient affected by refractory adult onset Still's disease who achieved a prolonged remission following CD34-selected ABMT. The conditioning regimen was based on the use of CY and anti-thymocyte globulin. A 3.0 and 2.0 log reduction of T (CD3+) and B (CD19+) lymphocytes, respectively, was obtained using a Ceprate device to select CD34+ cells from PBSC. In the pre transplant period (1994-1998) the patient had a chronic persistent disease course with frequent and recurrent systemic articular flares and loss of some functional abilities, despite daily prednisone, pulses of CY and immunosuppressive therapy (CYA or MTX). At the time of ABMT the patient had become non-ambulatory. Within 3 weeks of ABMT the patient showed a marked decrease in joint swelling, and morning stiffness. Joint pain and systemic symptoms disappeared, the patient was able to walk and run and gained general well being. ESR, C-reactive protein and WBC count were significantly decreased, while Hb level increased. This partial remission persisted for at least 1 year after ABMT, although at 15 months of follow-up a reappearance of moderate synovitis in the knees and wrists was noted. Our data further showed that both patient BM microenvironment and stem-progenitor cell function (as assessed by LTC-IC assay) were damaged even 1 year after CD34 selected ABMT, suggesting that the persistence of these alterations could have facilitated the favorable outcome of the disease following ABMT. Bone Marrow Transplantation (2000) 25, 1307-1310. PMID- 10871737 TI - Bone marrow transplantation does not correct the hyper IgE syndrome. AB - Congenital immunodeficiency in hyper IgE syndrome is characterised by a markedly raised IgE level, recurrent staphylococcal skin infection and pneumatoceles. Standard treatments include anti-staphylococcal antibiotics. We report a severely affected patient in whom successful bone marrow transplantation was followed by reappearance of the immunodeficiency. We conclude that bone marrow transplantation does not cure the immunological features of the hyper IgE syndrome. Bone Marrow Transplantation (2000) 25, 1303-1305. PMID- 10871739 TI - Successful treatment of relapsed Burkitt's lymphoma using unrelated cord blood transplantation as consolidation therapy. AB - We report a 10-year-old male with widespread recurrent Burkitt's lymphoma who underwent successful mismatched unrelated cord blood transplantation (UCBT) following salvage chemotherapy. He was conditioned with TBI, antithymocyte globulin (ATG) and high-dose VP-16 and achieved full donor engraftment. He experienced grade II skin and grade I gastrointestinal acute GVHD with no chronic GVHD. He is alive with no evidence of disease 24 months following UCBT. Bone Marrow Transplantation (2000) 25, 1311-1313. PMID- 10871740 TI - Effective high-dose chemotherapy combined with CD34+-selected autologous peripheral blood stem cell transplantation in a patient with cutaneous CD30 negative large T cell lymphoma. AB - Generalized multiple cutaneous tumors developed in a 60-year-old Japanese man. Skin biopsy revealed atypical large T lymphocytes infiltrating the dermis. CD30 staining was negative in the tumor cells. The diagnosis of CD30-negative cutaneous large T cell lymphoma was made. Axial and inguinal lymphadenopathy was present, but there was no evidence of bone marrow involvement. Seven cycles of chemotherapy and local electron beam irradiation were administered and complete remission (CR) was attained. As CD30-negative cutaneous large T cell lymphoma has a poor prognosis despite intensive chemotherapy, high-dose chemotherapy followed by CD34+-selected autologous peripheral blood stem cell transplantation (CD34+ APBSCT) was prescribed. The clinical course after CD34+-selected APBSCT was complicated with CMV infection occurring twice but administration of ganciclovir resolved the symptoms. He has remained in CR for 16 months after CD34+-APBSCT. This appears to be the first case report of CD34+-APBSCT in a patient with CD30 negative cutaneous large T cell lymphoma. Bone Marrow Transplantation (2000) 25, 1315-1317. PMID- 10871741 TI - Successful treatment of multiple myeloma relapsing after high-dose therapy and autologous transplantation with thalidomide as a single agent. AB - A 52-year-old dentist with kappa light chain multiple myeloma relapsed 6 months after 180 mg/m2 melphalan and an autograft. A partial remission had been attained after the autograft. Relapse occurred while he was on dexamethasone maintenance therapy. Chemotherapy was not an option due to low blood counts. Thalidomide was administered at relatively high doses (escalated up to 700 mg daily and continued for 4 months). There was a prompt decline in urine protein from 6067 mg/day to 2177 mg/day within a month. The response continued to improve with achievement of near-complete remission within 6 months and a decline in urine protein to 413 mg/day. Subsequently, grade 3 neutropenia and peripheral neuropathy required dose reduction to 200 mg/day. Disease activity parameters continued to improve on the lower dose of thalidomide. Nine months after starting thalidomide, the patient is in near-complete remission, enjoys an excellent quality of life, and has returned to work. We conclude that thalidomide can effectively control myeloma relapsing after high-dose chemotherapy, and may be especially useful in resistant cases or those unable to tolerate further chemotherapy. Bone Marrow Transplantation (2000) 25, 1319-1320. PMID- 10871742 TI - Results of allogeneic BMT in 16 patients with Fanconi's anemia. PMID- 10871745 TI - Keyword index to Volume 25. PMID- 10871743 TI - Mismatched related cord blood transplantation in a severe thalassemia patient. PMID- 10871746 TI - B7.1 expression by the weakly immunogenic F98 rat glioma does not enhance immunogenicity. AB - Enhanced immunogenicity has been reported following transfection of a variety of immunogenic tumors with the B7.1 co-stimulatory molecule. The purpose of the present study was to determine if transfection of a weakly immunogenic rat brain tumor, the F98 glioma, with the gene encoding B7.1 could enhance its immunogenicity. F98 cells were transfected with a plasmid containing the B7.1 gene, and stable transfectants (F98/B7.1) were obtained. Flow cytometric analysis confirmed the expression of B7.1 and MHC class I antigens on the cell surface. To investigate the effects of B7.1 expression on the tumorigenicity of the F98 glioma, Fischer rats were implanted intracerebrally with either F98 (wild-type) or F98/B7.1 transfected cells. No significant differences in survival times were noted. Mean survival times of 21.8 and 24.0 days were observed for the respective groups at a challenge dose of 103 cells. These differences in survival time were not significant. To determine if expression of B7.1 enhanced the immunogenicity of the F98 glioma, rats were vaccinated weekly for 3 weeks with 107 mitomycin C treated F98 or F98/B7.1 cells injected subcutaneously and then challenged intracerebrally with F98 cells 1 week later. Unvaccinated animals or those that received wild-type F98 cells as a vaccine had a survival time (mean +/- s.d.) of 22.3 +/- 1.5 days following tumor challenge versus 20.0 +/- 1.7 days for rats that had been vaccinated with F98/B7.1. Although we recognize that it might be possible to design more effective vaccination regimes, nevertheless, our data indicate that transfection of the B7.1 gene into the F98 rat glioma did not enhance its immunogenicity, and that other approaches will be required. PMID- 10871747 TI - Therapeutic application of T cell receptor mimic peptides or cDNA in the treatment of T cell-mediated skin diseases. AB - An 8-amino acid peptide encoding a sequence of the transmembrane region of the T cell receptor alpha chain (TCR-alpha) was shown to inhibit T cell function by preventing functional assembly of the T cell receptor (mimic peptide). To avoid systemic immunosuppression by peptide application in vivo, we used a topical application of the peptide. In the system of murine contact sensitivity, topical application of the peptide inhibited the elicitation of contact sensitivity following application of a contact allergen in sensitized animals. Alternatively, when naked DNA encoding the peptide sequence was injected into skin before application of a contact allergen to sensitized animals, local immunosuppression was also observed. To investigate the effects of this peptide in humans, patients with psoriasis, atopic eczema, lichen planus, or contact dermatitis were treated topically with mimic peptide or control peptide. All patients except for one reported a marked improvement or cure of their skin disease following application of the TCR-alpha peptide, but not controls. These data indicate that TCR-alpha peptide or cDNA treatment might be a proper treatment for human T cell-mediated dermatoses substituting for corticosteroids. PMID- 10871748 TI - Diffusion MRI detects early events in the response of a glioma model to the yeast cytosine deaminase gene therapy strategy. AB - Detection of a therapeutic response early in the course of cancer treatment, before tumor growth delay or regression, is not currently possible in experimental models or clinical medicine. New interim measures of therapeutic response would be particularly useful in the development of cancer chemosensitization gene therapy by facilitating optimization of gene transfer protocols and prodrug dosing schedules. Diffusion MRI is a sensitive technique producing quantitative and noninvasive images of the apparent mobility of water within a tissue. We investigated the utility of diffusion MRI for detecting early changes associated with a refined cytosine deaminase (CD)/5-fluorocytosine (5FC) chemosensitization gene therapy paradigm in orthotopic 9L gliomas stably expressing the recently cloned S. cerevisiae CD gene. Mean tumor diffusion increased 31% within 8 days of initiating 5-FC treatment, preceding tumor growth arrest and regression. Complete regression of the intracranial tumor was observed in four of five treated animals, and recurrent tumor in the remaining animal exhibited water diffusion behavior similar to primary, untreated tumors. These results demonstrate the efficacy of the yCD/5FC strategy for glioma and suggest that increased tumor water diffusion is an indicator of active therapeutic intervention. PMID- 10871749 TI - Intratracheal injection of manganese superoxide dismutase (MnSOD) plasmid/liposomes protects normal lung but not orthotopic tumors from irradiation. AB - To determine whether intratracheal (IT) lung protective manganese superoxide plasmid/liposomes (MnSOD-PL) complex provided 'bystander' protection of thoracic tumors, mice with orthotopic Lewis lung carcinoma-bacterial beta-galactosidase gene (3LL-LacZ) were studied. There was no significant difference in irradiation survival of 3LL-LacZ cells irradiated, then cocultured with MnSOD-PL-treated compared with control lung cells (D0 2.022 and 2.153, respectively), or when irradiation was delivered 24 h after coculture (D0 0.934 and 0.907, respectively). Tumor-bearing control mice showed 50% survival at 18 days and 10% survival at 21 days. Mice receiving liposomes with no insert or LacZ-PL complex plus 18 Gy had 50% survival at 22 days, and a 20% and 30% survival at day 50, respectively. Mice receiving MnSOD-PL complex followed by 18 Gy showed prolonged survival of 45% at 50 days after irradiation (P < 0.001). Nested RT-PCR assay for the human MnSOD transgene demonstrated expression at 24 h in normal lung, but not in orthotopic tumors. Decreased irradiation induction of TGF-beta1, TGF-beta2, TGF-beta3, MIF, TNF-alpha, and IL-1 at 24 h was detected in lungs, but not orthotopic tumors from MnSOD-PL-injected mice (P < 0.001). Thus, pulmonary radioprotective MnSOD-PL therapy does not provide detectable 'bystander' protection to thoracic tumors. PMID- 10871750 TI - Fractionated radiation therapy in combination with adenoviral delivery of the cytosine deaminase gene and 5-fluorocytosine enhances cytotoxic and antitumor effects in human colorectal and cholangiocarcinoma models. AB - Radiosensitization of human gastrointestinal tumors by 5-fluorouracil (5-FU) has been studied in vitro and clinically in human cancer therapy trials. The bacterial enzyme cytosine deaminase (CD) converts the nontoxic prodrug 5 fluorocytosine (5-FC) into 5-FU. Human colon cancer cells stably expressing CD have been shown by other investigators to be sensitized to radiation following treatment with 5-FC. We previously used an adenoviral vector under control of the cytomegalovirus promoter (AdCMVCD) encoding the CD gene in combination with 5-FC and a single fraction of radiation exposure to enhance cytotoxicity to human cholangiocarcinoma cells in vitro and in vivo. The purpose of this study was to determine whether AdCMVCD infection and 5-FC with multiple fraction low-dose radiotherapy results in enhanced cytotoxicity. In the present study, we utilized AdCMVCD and 5-FC with single fraction radiotherapy to demonstrate enhanced cytotoxicity to WiDr human colon carcinoma cells in vitro. Additionally, we tested this gene therapy/prodrug treatment strategy employing a fractionated radiation dosing schema in animal models of WiDr colon carcinoma and SK-ChA-1 cholangiocarcinoma. A prolonged WiDr tumor regrowth delay was obtained with AdCMVCD infection in combination with systemic delivery of 5-FC and fractionated external beam radiation therapy compared with control animals treated without radiation, without 5-FC, or without AdCMVCD. The results of treatment with AdCMVCD + 5-FC + radiation therapy to cholangiocarcinoma xenografts were equivalent to those obtained with systemic 5-FU administration + radiation. Thus, the use of AdCMVCD can be effectively combined with clinically relevant 5-FC and radiation administration schemes to achieve enhanced tumor cell killing and increased control of established tumors of human gastrointestinal malignancies. PMID- 10871751 TI - Soluble Flt-1 gene therapy for peritoneal metastases using HVJ-cationic liposomes. AB - Many studies have reported a close association between VEGF and tumor angiogenesis. The aim of the present study was to evaluate the effectiveness of gene therapy against cancer, including peritoneal metastasis, using a cDNA encoding a soluble type of Flt-1, one of the VEGF receptors. In a peritoneal metastasis model of MKN45 human gastric cancer cells, mice repetitively treated with intraperitoneal injections of HVJ-Fex, a type of HVJ-cationic liposome encapsulating a plasmid expressing soluble mFlt-1, exhibited smaller disseminated foci with fewer microvessels, thus resulting in a significantly longer survival period than the control mice. In another peritoneal metastasis model using HT1080S cells, a clone of HT1080 human fibrosarcoma cells stably transfected with hVEGF, treatments with HVJ-Fex also reduced the growth of disseminated foci without ascites formation. In conclusion, this study demonstrated that the peritoneal metastases of some cancers were largely dependent on VEGF, and that the repeated intraperitoneal transduction of a soluble flt-1 gene using HVJ cationic liposomes suppressed peritoneal metastases, thereby contributing to a longer survival period. PMID- 10871752 TI - A simple method for the rapid generation of recombinant adenovirus vectors. AB - Recombinant adenoviruses are useful vectors for basic research. When the vectors are used for delineating protein function, several viruses, each containing a mutated version of the transgene are compared at the same time. However, methods to generate multiple vectors simultaneously within a short time period are cumbersome. In this report, we show that a novel backbone plasmid, when cotransfected with routinely used shuttle vectors into HEK293 cells allowed for production of recombinant viruses in an average of 14 days. The recombinant viruses had no detectable wild-type virus contamination by A549 plaque assay and only three to 300 E1a copies per 109 adenovirus genomes by a sensitive PCR-based assay. Further culturing or serial amplification did not result in wild-type revertants nor did cultures show increased levels of E1a copy number by quantitative PCR. Thus, recombinant adenovirus vectors can be produced very simply, rapidly and with little to no contaminating wild-type particles. This system should facilitate the generation of multiple genetic variants by eliminating the need for time-consuming plaque purification and the need to manipulate and screen very large plasmids. We call this the RAPAd.I system. PMID- 10871753 TI - Ligand-mediated retargeting of recombinant adenovirus for gene transfer in vivo. AB - The development of efficient and safe methods for in vivo gene transfer is central to the success of gene therapy. Recombinant adenoviral vectors, although highly efficient, are limited by the host immune response, potential safety hazards due to obligatory cotransfer of viral proteins, and their broad tissue tropism. Here, we demonstrate in an animal model that host range and tissue tropism of a recombinant adenovirus from a distant species can be modified by complexing adenovirus with a cell-specific ligand. Thus, a replication-deficient lacZ recombinant human adenovirus, which naturally does not infect avian cells, allowed highly efficient and specific gene transfer to the liver of ducks in vivo when complexed with N-acetylglucosamine, a ligand for the chicken hepatic lectin. This combination of ligand-mediated receptor targeting with adenoviral uptake and intracellular processing of a given gene represents a novel approach to gene therapy of inherited and acquired liver diseases. PMID- 10871754 TI - Effect of transgenic GDNF expression on gentamicin-induced cochlear and vestibular toxicity. AB - Gentamicin administration often results in cochlear and/or vestibular hair cell loss and hearing and balance impairment. It has been demonstrated that adenovirus mediated overexpression of glial cell line-derived neurotrophic factor (GDNF) can protect cochlear hair cells against ototoxic injury. In this study, we evaluated the protective effects of adenovirus-mediated overexpression of GDNF against gentamicin ototoxicity. An adenovirus vector expressing the human GDNF gene (Ad.GDNF) was administered into the scala vestibuli as a rescue agent at the same time as gentamicin, or as a protective agent, 7 days before gentamicin administration. Animals in the Rescue group displayed hearing thresholds that were significantly better than those measured in the Gentamicin or Ad.LacZ/Gentamicin groups. In the Protection group, Ad.GDNF afforded significant preservation of utricular hair cells. The data demonstrated protection of the inner ear structure, and rescue of the inner ear structure and function against ototoxic insults. These experiments suggest that inner ear gene therapy may be developed as a clinical tool for protecting the ear against environmentally induced insults. PMID- 10871755 TI - An improved anion-exchange HPLC method for the detection and purification of adenoviral particles. AB - We have developed an anion-exchange high-performance liquid chromatography (HPLC) method using Q Sepharose XL (Amersham Pharmacia Biotech) as adsorbent to analyze samples containing adenovirus. This method has several major advantages over the HPLC method previously described for quantitating particles, namely (1) a >10 fold improvement in the detection limit of adenovirus in crude preparations; (2) absence of interferences originating from nucleic acids and proteins which usually contaminate crude samples; (3) unprecedented sharpness and symmetry of adenovirus peak, rendering the identification of the viral peak unambiguous, even in extremely crude and dilute preparations; and (4) no enzymatic treatment required even for crude samples. This assay was used to quantitate particles in samples taken at the transfection and amplification stages of production of various recombinant adenovirus, and in cultures of wild-type adenovirus of different serotypes. A modification of this analytical method was also developed for the purification of infectious adenovirus particles, including fiber-modified and third-generation recombinant viruses, giving highly purified preparations from low-titer crude lysates with an excellent overall recovery (50-74%). PMID- 10871756 TI - Plat-E: an efficient and stable system for transient packaging of retroviruses. AB - A potent retrovirus packaging cell line named Platinum-E (Plat-E) was generated based on the 293T cell line. Plat-E is superior to existing packaging cell lines regarding efficiency, stability and safety. The novel packaging constructs utilized in establishment of Plat-E ensure high and stable expression of viral structural proteins. Conventional packaging constructs made use of the promoter of MuLV-LTR for expression of viral structural genes gag-pol and env, while our packaging constructs utilized the EF1alpha promoter, which is 100-fold more potent than the MuLV-LTR in 293T cells in combination with the Kozak's consensus sequence upstream of the initiation codon resulting in high expression of virus structural proteins in Plat-E cells. To maintain the high titers of retroviruses under drug selection pressure, we inserted the IRES (internal ribosome entry site) sequence between the gene encoding gag-pol or env, and the gene encoding a selectable marker in the packaging constructs. Plat-E cells can stably produce retroviruses with an average titer of 1 x 107/ml for at least 4 months. In addition, as we used only the coding sequences of viral structural genes to avoid inclusion of unnecessary retrovirus sequences in the packaging constructs, the probability of generating the replication competent retroviruses (RCR) by recombination can virtually be ruled out. PMID- 10871757 TI - T cell activation by recombinant FcepsilonRI gamma-chain immune receptors: an extracellular spacer domain impairs antigen-dependent T cell activation but not antigen recognition. AB - T cells can be endowed with antigen specificity by grafting with a chimeric receptor consisting of an extracellular antigen binding moiety (scFv) derived from an antibody and an intracellular signaling domain. Conflicting data exist on the impact of an extracellular spacer domain between the antigen binding and the signaling domain with respect to cellular activation. Here, we recorded conjugate formation and antigen-driven cellular activation of T cells grafted with receptor molecules that contain the same antigen binding site (anti-CD30 HRS3-scFv) and signaling domain (FcepsilonRI gamma-chain), however, with and without an IgG1 CH2CH3 (Fc) spacer domain between the scFv and transmembrane moiety. Receptors of both configurations mediate equally efficient conjugate formation between receptor grafted T cells and antigen-positive target cells. Specific signaling by the spacer containing receptor, however, is blocked by five- to 10-fold lower concentrations of soluble antigen than by the spacer-less receptor indicating a higher avidity of the spacer containing receptor to soluble antigen. In contrast, cellular activation upon binding to antigen-positive cells is mediated more efficiently by the spacer-less receptor. This demonstrates that the extracellular spacer domain impairs antigen-dependent cellular activation by the chimeric immune receptor, but not intercellular conjugate formation. PMID- 10871758 TI - The artificial zinc finger coding gene 'Jazz' binds the utrophin promoter and activates transcription. AB - Up-regulation of utrophin gene expression is recognized as a plausible therapeutic approach in the treatment of Duchenne muscular dystrophy (DMD). We have designed and engineered new zinc finger-based transcription factors capable of binding and activating transcription from the promoter of the dystrophin related gene, utrophin. Using the recognition 'code' that proposes specific rules between zinc finger primary structure and potential DNA binding sites, we engineered a new gene named 'Jazz' that encodes for a three-zinc finger peptide. Jazz belongs to the Cys2-His2 zinc finger type and was engineered to target the nine base pair DNA sequence: 5'-GCT-GCT-GCG-3', present in the promoter region of both the human and mouse utrophin gene. The entire zinc finger alpha-helix region, containing the amino acid positions that are crucial for DNA binding, was specifically chosen on the basis of the contacts more frequently represented in the available list of the 'code'. Here we demonstrate that Jazz protein binds specifically to the double-stranded DNA target, with a dissociation constant of about 32 nM. Band shift and super-shift experiments confirmed the high affinity and specificity of Jazz protein for its DNA target. Moreover, we show that chimeric proteins, named Gal4-Jazz and Sp1-Jazz, are able to drive the transcription of a test gene from the human utrophin promoter. PMID- 10871759 TI - HIV and the nervous system: emerging issues. Proceedings of a symposium. Washington DC, USA. April 14-16, 1999. PMID- 10871760 TI - Current approaches to treatment for HIV-1 infection. AB - The last 3 years have seen a dramatic fall in mortality and morbidity from HIV infection. Four factors have contributed to this: an improved understanding of the pathogenesis of HIV infection; the availability of tests that could measure plasma viral burden; the development of new and more powerful drugs such as the protease and non-nucleoside reverse transcriptase inhibitors; and the completion of large clinical endpoint trials that conclusively demonstrated that potent antiretroviral combinations significantly delayed the progression of HIV disease and improved survival. Typical antiretroviral regimen now consist of at least three agents: one or two protease inhibitors or a non-nucleoside reverse transcriptase inhibitor combined with two nucleoside analogs. The goal of therapy is to reduce measurable plasma viral burden to undetectable levels. Viral load testing has made it possible to individualize therapy and to more accurately determine the best time to initiate or change therapy, long before declining CD4+ counts would have given evidence of active viral replication. However, despite the impressive progress to date, there remain significant shortcomings with current treatment. Even with the most potent regimens available, there exists a proportion of patients (perhaps 20 - 50% of treated individuals) who fail to have complete and durable virologic responses to therapy. The shortcomings of current regimens are particularly evident in patients with high plasma HIV-1 RNA levels, extensive prior treatment, and advanced disease. Complexity, short- and long-term toxicities, cross-resistance, and drug-drug interactions all complicate current regimens. Viral resistance is increasingly encountered in clinical practice and transmission of resistant virus is well-documented. In addition, there remain concerns about the ability of the virus to evade current therapies, whether in viral reservoirs in non-lymphoid compartments or in lymphoid tissue, such as resting memory T cells. Thus there remains a need for new therapies as well as new strategies using existing drugs. PMID- 10871761 TI - HIV-1 infected mononuclear phagocyte secretory products affect neuronal physiology leading to cellular demise: relevance for HIV-1-associated dementia. AB - Viral and cellular products from HIV-1-infected and/or immune competent mononuclear phagocytes (MP) (brain macrophages and microglia) affect neuronal function during HIV-1-associated dementia (HAD). Neurotoxic MP factors include, but are not limited to, pro-inflammatory cytokines, chemokines, platelet activating factor, arachidonic acid and its metabolites, nitric oxide, progeny virions and viral structural and regulatory proteins. The mechanisms for immune mediated neural injury in HAD, only now, are being unraveled. In this regard, we reviewed the current knowledge of how postmitotic neurons, which can neither divide nor be replaced, are damaged by MP secretory activities. Linking neuronal function with brain MP activation was made possible by placing viral and/or immune products onto neurons and measuring cell signaling events or through ex vivo electrophysiological tests on MP-treated brain slices. Such linkages are shown, in this report, by select demonstrations of MP factors which cause neuronal dysfunction in HAD. PMID- 10871762 TI - Chemokine receptors and mechanisms of cell death in HIV neuropathogenesis. AB - Several chemokine receptors are used as coreceptors for HIV-1 entry in the central nervous system (CNS). CCR5 is the major coreceptor together with CD4 for HIV-1 infection of microglia, the major target cells for HIV-1 infection in the CNS. CXCR4 and CCR3 are also expressed on microglia and can mediate infection by certain HIV-1 isolates but at lower efficiency than CCR5. Additional chemokine coreceptors are expressed in the brain, but their role in HIV-1 neuropathogenesis has not been defined. The expression of CXCR4, and possibly other chemokine receptors, on subpopulations of neurons and glial cells may render neurons vulnerable to mechanisms of CNS injury induced by the HIV-1 gp120 Env protein. HIV-1 viruses which use CXCR4 and emerge during the late stages of HIV-1 infection may impact disease progression in the CNS by inducing apoptosis of neurons and other cell types. The neurodegenerative mechanisms may involve infection of microglia by certain CXCR4 tropic viruses in addition to cellular dysfunction and apoptosis induced by HIV-1 gp120 binding to CXCR4. Understanding the role of CXCR4 and other chemokine receptors in HIV-1 neuropathogenesis will help to advance the development of new therapeutic strategies for the prevention and treatment of neurologic disorders associated with HIV-1 infection. PMID- 10871763 TI - Neurologic disease in injection drug users: therapeutic issues. AB - Illicit drug use may cause nervous system impairment as a result of direct and indirect effects on the integrity and function of nervous system tissue and, potentially, through immune effects. HIV-1 infection poses an additional risk of impairment, and this risk may be decreased as a result of antiretroviral drug treatment. Obviously, the goal of such therapy is to improve the potential clinical course of infection. However, interactions between antiretroviral drugs, abused drugs, and hepatic metabolic enzyme systems may result in impaired or more efficient drug clearance and, consequently, antiretroviral or substance abuse treatment failure. The clinical outcome of this interaction may potentially include drug-related neurotoxicity or neurologic disease induced by HIV infection. The actual impact of these interactions on the occurrence of neurologic impairment and disease are unknown at this time, and, therefore, require study. PMID- 10871764 TI - Neuronal apoptosis in human immunodeficiency virus infection. AB - Neuronal apoptosis has been shown to occur in HIV infection by a number of in vivo and in vitro studies, however, the cause of neuronal damage in AIDS is still unclear and its relationships with the cognitive disorders characteristic of HIV dementia remain a matter of debate. In this review, based on our experience, we analyse the techniques used to identify neuronal apoptosis on post-mortem AIDS brains and describe the relationships of neuronal apoptosis with the stage of disease, a history of HIV-dementia, the degree of productive HIV infection, microglial activation, blood-brain barrier involvement and axonal damage. We conclude that the severity of neuronal apoptosis in the cerebral cortex correlates with the presence of cerebral atrophy, but not with the cognitive disorders. There is no global quantitative correlation between neuronal apoptosis and HIV encephalitis, microglial activation or axonal damage. However we found some topographical correlation between these changes. We conclude that neuronal apoptosis and consequent neuronal loss, in HIV infected patients, are probably not related to a single cause. It seems likely that microglial activation, directly or indirectly related to HIV infection of the CNS, plays a major role in its causation possibly through the mediation of oxidative stress. Axonal damage, either secondary to microglial activation, or to the intervention of systemic factors may also contribute to neuronal apoptosis. PMID- 10871765 TI - Biphasic and regionally-restricted chemokine expression in the central nervous system in the Theiler's virus model of multiple sclerosis. AB - Intracerebral infection of susceptible strains of mice with Theiler's murine encephalomyelitis virus (TMEV) induces a biphasic disease characterized by acute polioencephalitis followed by chronic demyelination and viral persistence in the spinal cord white matter. There has been limited study of soluble mediators responsible for the initial recruitment of inflammatory cells into the gray matter, and the secondary influx into the white matter during infection with TMEV. We used sensitive and specific RT - PCR/dot blot hybridization assays to quantitate the relative levels of chemokine mRNA in the brains and spinal cords during the acute and chronic phases of TMEV infection in mice susceptible (B10.M, H-2f) and resistant (B10, H-2b) to virus-induced demyelination. TMEV infection resulted in robust expression of mRNA for IP-10, RANTES, and MCP-1, but not GRO alpha, in brains and spinal cords in both strains of mice within 5 days. By day 21, virus was cleared, inflammation reduced, and expression of all three chemokines subsided to baseline levels in the brains and spinal cords of resistant mice, and the brains of susceptible mice. Chemokine expression was also reduced in the spinal cords of susceptible mice, corresponding to a shift in TMEV replication from the gray to the white matter. During the chronic, demyelinating phase of infection, there was a resurgence in IP-10, RANTES, and MCP-1 mRNA in spinal cords of susceptible B10.M mice. This study demonstrates the coordinated regulation and regionally restricted expression of chemokines in a biphasic disease of the central nervous system and provides greater understanding of the mechanism by which inflammation is established and maintained in the CNS. PMID- 10871766 TI - Characterization of cultured microglia that can be infected by HIV-1. AB - Parenchymal microglia are targets of HIV infection. We, as well as others, have used in vitro microglia culture systems to study the tropism and replication of HIV. Characterization of perivascular and parenchymal microglia surface markers in vivo, in vitro, and ex vivo, has led to the understanding that these cell populations are different, and data from both the HIV and SIV models support the hypothesis that they may play different roles in infection of the CNS. We determined that human adult parenchymal microglia cultured from temporal lobe tissue for use in HIV replication studies, were CD11c+, CD45+, CD68+, CD14- when cultured with standard serum/cytokine-supplemented media. To determine the influence of serum and cytokines on HIV replication in microglia, we designed a new protocol for culturing microglia, and compared the results obtained with this protocol with the standard approach previously described. Microglia cultured in the presence of a 'feeder' layer of glial cells and in the absence of serum and cytokines expressed the same surface markers as pure microglia (>95%) cultured in supplemented media. However, pure microglia cultured in the absence of both serum/cytokines supplements and other glial cells, did not have characteristic microglial morphology and did not support HIV replication to as high a level. Lastly, we determined that unlike monocytes, ex vivo parenchymal microglia were capable of supporting HIV replication. PMID- 10871767 TI - Functional expression of the seven-transmembrane HIV-1 co-receptor APJ in neural cells. AB - APJ is a recently described seven-transmembrane (7TM) receptor that is abundantly expressed in the central nervous system (CNS). This suggests an important role for APJ in neural development and/or function, but neither its cellular distribution nor its function have been defined. APJ can also serve as a co receptor with CD4 for fusion and infection by some strains of human immunodeficiency virus (HIV-1) in vitro, suggesting a role in HIV neuropathogenesis if it were expressed on CD4-positive CNS cells. To address this, we examined APJ expression in cultured neurons, astrocytes, oligodendrocytes, microglia and monocyte-derived macrophages utilizing both immunocytochemical staining with a polyclonal anti-APJ antibody and RT - PCR. We also analyzed the ability of a recently identified APJ peptide ligand, apelin, to induce calcium elevations in cultured neural cells. APJ was expressed at a high level in neurons and oligodendrocytes, and at lower levels in astrocytes. In contrast, APJ was not expressed in either primary microglia or monocyte-derived macrophages. Several forms of the APJ peptide ligand induced calcium elevations in neurons. Thus, APJ is selectively expressed in certain CNS cell types and mediates intracellular signals in neurons, suggesting that APJ may normally play a role in signaling in the CNS. However, the absence of APJ expression in microglia and macrophages, the prinicpal CD4-positive cell types in the brain, indicates that APJ is unlikely to mediate HIV-1 infection in the CNS. PMID- 10871768 TI - Analysis of human immunodeficiency virus type 1 gp160 sequences from a patient with HIV dementia: evidence for monocyte trafficking into brain. AB - Towards understanding the pathogenesis of HIV dementia, we molecularly cloned and sequenced human immundeficiency virus type 1 (HIV-1) gp160 genes from uncultured post-mortem tissues collected from a patient with HIV dementia. Sequences from bone marrow, lymph node, lung, and four regions of brain - the deep white matter, head of caudate, choroid plexus and meninges - were compared. Also included were gp160 sequences recovered from blood monocytes collected 5 months prior to death. Phylogenetic analyses showed that the sequences from deep white matter were more closely related to those from bone marrow, than to those from the other tissues, and moreover, were most closely related to sequences from the blood monocytes. These findings suggest trafficking of bone marrow-derived monocytes into the deep white matter during this late stage of infection. Another cluster included sequences from choroid plexus, meninges and lymph node, and interestingly, identical patterns of four or nine stop codons were shared among these tissues. These mutations appear to be the consequence of G-->A hypermutation, and could reflect independent events, or the movement of virions or infected cells, from the choroid plexus into the cerebrospinal fluid and ultimately, into the lymph node. We propose that a critical step towards the development of HIV dementia is an increase in monocyte trafficking into the brain, and that this process is either initiated and/or accelerated during late-stage infection, which could explain why dementia occurs primarily during this time. PMID- 10871769 TI - Mechanisms of leukocyte trafficking into the CNS. AB - HIV-1 encephalitis occurs in up to one-third of HIV-1-infected individuals. The mechanisms through which this pathology develops are thought to involve viral passage across the blood-brain barrier (BBB), as well as entry of HIV-infected and/or uninfected inflammatory cells into the central nervous system (CNS). Viral proteins and cytokines may also contribute to the pathogenesis of encephalitis. We show that the chemokines SDF-1 and MCP-1 induce transmigration of uninfected human lymphocytes and monocytes across our model of the BBB, a co-culture of human fetal astrocytes and endothelial cells. We also demonstrate that the HIV-1 protein Tat induces adhesion molecule expression and chemokine production by human fetal astrocytes and microglia, which could further contribute to leukocyte entry into the CNS. Finally, our data indicate that inflammatory cytokines modulate the expression of CXCR4, a co-receptor for HIV-1, on human fetal astrocytes, suggesting that these cytokines may potentially modulate the infectability of astrocytes by HIV-1. These findings support the hypothesis that there may be several different mechanisms that contribute to the development and progression of HIV-1 encephalitis. PMID- 10871770 TI - Persistent pathogens in the parenchyma of the brain. AB - It has recently been shown that bacteria and viruses can be delivered to the brain parenchyma without evoking an immune response. These experiments demonstrate that there are no cells within the brain parenchyma that can initiate a primary immune response, and that the drainage of pathogens from the brain parenchyma is distinct from that documented for soluble proteins. A persistent pathogen in the brain parenchyma can become a target for the immune system following peripheral sensitisation, and this may lead to bystander tissue damage. These observations may have consequences for vaccination of persons with central nervous system HIV infection. PMID- 10871771 TI - Stages of restricted HIV-1 infection in astrocyte cultures derived from human fetal brain tissue. AB - The predominant cell types infected by HIV-1 in AIDS associated encephalopathy are cells of the macrophage/microglial lineage. There has been consistent evidence, however, that astrocytes also become infected although not at the same frequency or level of multiplication as microglial cells. HIV-1 antigens and/or nucleic acid have been identified in astrocytes in brain autopsy tissue from both adult and pediatric AIDS cases. In cell cultures, HIV-1 infection of astrocytes results in an initial productive but non-cytopathogenic infection that diminishes to a viral persistence or latent state. Understanding the nature of HIV-1 infection of astrocytes, which represents the largest population of cells in the brain, will contribute to the understanding of AIDS encephalopathy and the dementia that occurs in nearly one-quarter of all AIDS patients. PMID- 10871772 TI - Molecular analysis of cerebrospinal fluid: potential for the study of HIV-1 infection of the central nervous system. AB - The molecular analysis of cerebrospinal fluid (CSF) provides an inestimable tool for the study of HIV infection of the central nervous system (CNS). Current nucleic acid amplification techniques enable the measurement of CSF HIV-1 RNA levels which can be predictive of HIV-associated neurological damage. CSF HIV-1 RNA levels do not necessarily correlate with the corresponding plasma levels, thus supporting the possibility of an intrathecal virus production, i.e., from brain macrophages. However, in early stages of HIV infection, as well as during some opportunistic CNS diseases, CNS or CSF infiltrating lymphocytes might be the main source of CSF virus. A drastic decrease in CSF viral load is usually observed along with a decrease in plasma levels in patients receiving highly active antiretroviral therapy (HAART), with durable suppression of CSF viral load over months. However, during the first weeks of therapy, the dynamics of response may differ in the CSF as compared to plasma, again suggesting that virus replication may be compartmentalised in the CSF. A number of mechanisms are likely to be involved in the response to therapy in CSF, including among the others the trafficking of cell populations supporting viral replication between blood, CNS and CSF, and the role of the anatomical brain barriers in limiting the access of antiretroviral drugs into the CSF. A potential risk associated with compartmentalisation of HIV infection is of an incomplete suppression of virus replication in the CSF, thus creating the ground for local development of anti HIV drug resistance. In order to assess this occurrence, long-term studies of viral load and genotypic analyses on paired CSF and plasma will be necessary and these will also help elucidate the complex interrelationship between viral replication in these compartments. PMID- 10871773 TI - Neuronal damage - recent issues and implications for therapy. AB - The spectrum of structural damage in the brain associated with HIV is now more fully understood. Such changes include inflammatory disorders (such as HIV encephalitis, leukoencephalopathy, and diffuse poliodystrophy), dendritic and synaptic damage, and neuronal loss. However, the relationship between neuronal damage and loss and clinical variables is still not clear. In my laboratory my research group has been addressing three separate areas, which will be the subject of this presentation. (1) The relationship of neuronal damage and loss to various clinical features such as cognitive symptoms, and risk group. (2) The cellular site of production of neurotoxic factors in the HIV-infected brain. (3) Assessing potential neuroprotective strategies using appropriate in-vitro models. PMID- 10871774 TI - The puzzling natural history of multiple sclerosis: a challenge for the research and care. PMID- 10871775 TI - History and activities of the Don Carlo Gnocchi multiple sclerosis center. PMID- 10871776 TI - The genetics of multiple sclerosis. AB - Epidemiological studies implicate an interplay between genetic and environmental factors in the aetiology of multiple sclerosis. The classical genetic observations suggest that multiple sclerosis is a complex trait in which susceptibility is determined by several genes acting independently or epistatically. The main dividend from understanding the genetic basis of susceptibility in multiple sclerosis will be an improved understanding of the pathogenesis. To date, candidate gene approaches have proved relatively unrewarding other than in establishing the association with alleles of the major histocompatibility complex (MHC). In common with most other complex traits, no major susceptibility gene has been identified through full genome screens but regions of interest have provisionally been identified. An important part of future studies in the genetics of multiple sclerosis will be to resolve the question of disease heterogeneity. PMID- 10871777 TI - The complex genetic aetiology of multiple sclerosis. AB - A large body of immunologic, epidemiologic, and genetic data indicate that tissue injury in multiple sclerosis (MS) results from an abnormal immune response to one or more myelin antigens that develops in genetically susceptible individuals after exposure to an as-yet undefined casual agent. A genetic component in MS is indicated by an increased relative risk to siblings compared to the general population and an increased concordance rate in monozygotic compared to dizygotic twins. The past few years have seen real progress in defining the genetic basis of MS setting the stage for new approaches for the final characterisation of the genes involved in MS susceptibility and pathogenesis. Whole genome screens conducted in different populations identified discrete chromosomal regions potentially harbouring MS susceptibility genes, however, with the exception of the Major Histocompatibility Complex (MHC) on 6p21, no single locus generated overwhelming evidence of linkage. These results suggest a complex genetic aetiology, including multiple genes of small to moderate effect and probable genetic heterogeneity. The identification and characterisation of MS susceptibility genes and their correlation with disease phenotypes is likely to define the basic aetiology of the disease, improve risk assessment and influence therapeutics. PMID- 10871778 TI - MS genetics: recent Scandinavian efforts. AB - As a potential founder population of MS, the Scandinavian ethnic group is of special interest in MS genetics. Project in these countries have recently led to several reports, including associations with HLA class I, CTLA-4 and suggestive linkage to several chromosomal candidate loci. The analysis of isolated populations within Scandinavia may also prove to be rewarding. PMID- 10871779 TI - An attempt of identifying MS-associated loci as a follow-up of a genomic linkage study in the Italian population. AB - Subsequent to a genomic linkage study on Sardinian and Continental Italian families, we considered the possibility that some of the tested microsatellite markers showed association to MS. Markers selected on the basis of the data obtained in the original set of 70 multiplex families were tested for MS association in an additional set of 154 simplex families. A limited set of markers were further tested on an additional set of 100 simplex families. The results indicate the presence of a putative MS gene in 19q13.13. PMID- 10871780 TI - Genetic risk factors in multiple sclerosis and approaches to their identification. AB - Development of multiple sclerosis (MS) is believed to involve genetic as well as environmental factors. A complicating aspect to the study of aetiological factors in MS concerns the possible existence of genetically different subtypes of the disease. In addition, a relatively large number of susceptibility genes could be involved. Most likely, the contribution of the single genes to the susceptibility to MS is modest. However, interactions between different genes could result in a dramatic increase in disease susceptibility (synergistic gene effects). In this short review we focus upon genetic heterogeneity and gene interactions in MS. We also outline approaches to the genetic analysis of complex disease traits such as MS. PMID- 10871781 TI - A polymorphism in the repetitive (TGGA)n sequence 5' to the human myelin basic protein gene in Italian multiple sclerosis patients. AB - Human myelin basic protein (hMBP) gene is one of the candidate genes in the complex mosaic of multiple sclerosis (MS) susceptibility. In this study we verified the distribution of the polymorphism of the region 5' flanking the first exon of the hMBP gene, in 97 relapsing remitting, 74 primary progressive Italian MS patients, and in 236 healthy controls, using polymerase chain reaction (PCR) and gel electrophoresis analysis in this region from 1116 - 1540 nt. Three different band patterns were observed: one homozygote with a 354 bp long fragment, one homozygote with 424 bp long fragment and one heterozygote with both bands present. The short fragment was statistically more frequent in RRMS patients than in HC (P<0.05). The long fragment was more present in HC. Similarly the short homozygous pattern (354 bp/354 bp) was significantly higher in the RRMS patients versus the healthy controls (P<0.01). The sequence analysis of the hMBP alleles showed that while the long fragments matched the prototype sequence completely, all the short fragments showed a deletion of 70 bp from nt 1177 to nt 1247, which explains the short 354 bp allele detected by PCR. Moreover two single mismatches in positions 1386 (T-->C) and 1431 (G-->A), were present only in the short hMBP fragment. PMID- 10871783 TI - Augmented type 1 cytokines and human endogenous retroviruses specific immune responses in patients with acute multiple sclerosis. AB - In vitro antigen- and mitogen-stimulated cytokine production were analysed in multiple sclerosis (MS) patients with either acute (AMS) or stable (SMS) disease and in healthy controls (HC). We also investigated whether immune responses to human endogenous retroviruses (HERV) could be detected in MS and whether these immune responses would be correlated with disease status by analysing cytokine production after stimulation of PBMC with HERV peptides. Results showed that mitogen-stimulated IL-2 and IFN-gamma was augmented and IL-10 was decreased in AMS compared to both SMS and healthy controls. Whereas the production of the metabolically active IL-12 (p70 heterodimer), was comparable in SMS, AMS and HC, production of the total IL-12 (p70 heterodimer and the p40 chain) were augmented in SMS compared to both AMS and HC. HERV-peptides IL-2 and IFN-gamma production was more frequent and more potent in AMS compared to both SMS patients and HC. HERV-specific type 2 cytokine production was more frequent and potent in SMS compared to AMS and HC. Thus a prevalent type 1 cytokine profile was seen in AMS patients, while IL-10 production predominated in SMS individuals. PMID- 10871782 TI - Induction of IL-1 receptor antagonist by interferon beta: implication for the treatment of multiple sclerosis. AB - IFNbeta has been the first drug approved for the treatment of multiple sclerosis patients, but we still lack a full understanding of the mechanisms underlying its clinical effects and the great variability of its therapeutic efficacy among different patients. Serum levels of the anti-inflammatory cytokine IL-1 receptor antagonist increase after IFNbeta administration in MS patients. We now report that IFNbeta induced IL-1ra mRNA and mature protein in three myelomonocytic cell lines. The induction of IL-1ra was already visible after 2 h of stimulation and persisted at least for 24 h. The amounts of induced IL-1ra were equal or higher than those obtained using other IL-1ra stimuli (LPS, IL-1beta, IFNgamma, IL-4, dexamethasone). This prolonged and quantitatively elevated induction of IL-1ra may contribute to the anti-inflammatory effect of IFNbeta and partially account for the reduction of exacerbation rate shown in most IFNbeta-treated MS patients. PMID- 10871784 TI - Presence of autoantibodies against complement regulatory proteins in relapsing remitting multiple sclerosis. AB - Complement was proposed to play an important role in the onset of Multiple Sclerosis (MS) lesions by inducing physical damage to myelin-producing cells. Every somatic cell is however endowed with a repertoire of membrane-bound molecules which normally down-regulate the complement activation cascade (Regulators of Complement Activation, RCA) and therefore protect cells from complement-dependent lysis. We show here that antibodies against two complement regulatory molecules expressed in the membrane of human cells (CD46 and CD59) are present in sera from relapsing-remitting MS patients in the acute phase, that they are directed against the active site of the RCA molecules and that they inactivate their regulatory function, thus providing a mechanism by which cells of the nervous system might be damaged in a complement-dependent fashion during the acute MS phase. Moreover, we found that most of these sera also contain antibodies reacting with an epitope of the transmembrane glycoprotein of HIV which is conserved in most retroviruses; this may support the hypothesis that self-reacting antibodies might have arisen in these patients as an immune response after retroviral infection or expression of endogenous retroviral proteins. PMID- 10871785 TI - Effects of rIFN-beta-1b on serum circulating ICAM-1 in relapsing remitting multiple sclerosis and on the membrane-bound ICAM-1 expression on brain microvascular endothelial cells. AB - rIFN-beta reduces the frequency of the gadolinium-enhancing (Gd+) magnetic resonance imaging (MRI) lesions in relapsing remitting (RR) MS. Its mechanism of action on improving the integrity of the blood-brain barrier (BBB) remains unclear. We investigated the effect of rIFN-beta-1b on the soluble intercellular adhesion molecule-1 (sICAM-1) serum levels (ELISA) in 36 RR MS patients receiving treatment with rIFN-beta for 1 year, and also the TNF-alpha - induced membrane bound ICAM-1 (mICAM-1) expression on cultured rat brain microvascular endothelial cells (BMECs). In vivo data showed that sICAM-1 serum levels at baseline significantly increased (P<0.01) in 12 months of rIFN-beta-1b treatment. The increase paralleled a clinical and MRI improvement. In the second semester of the treatment the integrated area under the curve of Expanded Disability Status Score normalised to entry baseline (DeltaEDSS AUC) was significantly (P<0.05) smaller than in the first semester. The percentage of patients with Gd+MRI decreased significantly (P<0.05) in the first (33%) and second (29%) semesters of treatment compared to baseline (62%). In vitro experiments showed that the incubation of BMEC monolayer with 100 u/ml of TNF-alpha for 24 h significantly (P<0.05) increased mICAM-1 expression, whereas 2000 u/ml of rIFN-beta-1b for 72 h did not modify the baseline levels. The incubation of BMEC with 2000 u/ml of rIFN-beta-1b for 48 h followed by combined IFN-beta-1b and TNF-alpha for 24 h significantly (P<0.05) downregulated TNF-alpha-induced mICAM-1 expression. These results suggest that the effect of rIFN-beta-1b on the BBB may be mediated by changes in both sICAM-1 serum levels and mICAM-1 BMEC expression. PMID- 10871786 TI - Dynamics of the reactivity to MBP in multiple sclerosis. AB - Though many lines of evidence support the importance of myelin basic protein (MBP) in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), its role in multiple sclerosis (MS) is still debated as well as the significance of epitope spreading in disease progression. We characterised the response to MBP in eight MS subjects and three of these were followed over time. In one case, the follow up lasted over a 6-year period. Clonal expansion, clonal persistence and epitope spreading against other MBP determinants was detected irrespective of disease course. In one patient we identified a novel T-cell receptor variable gene (BV28S2) which may be involved in the selection of MBP determinants, as suggested by experiments performed in the presence of mismatched antigen presenting cells (APC) between two subjects compatible for HLA-DR2 subtype but differing for the epitope recognised. Our findings do not sustain a role for the response to MBP effecting on clinical course and suggest that a novel TCR gene may be involved in the recognition of unusual self antigens. PMID- 10871787 TI - Serum auto antibodies presence in multiple sclerosis patients treated with beta interferon 1a and 1b. AB - To verify the possible effect of IFN-beta treatment on auto antibodies development in multiple sclerosis (MS) we studied 69 MS patients before and during the treatment with IFN-beta 1b (n=35) and IFN-beta 1a (n=20) for 27 and 12 months respectively, and, as controls, 14 untreated MS patients. The serum, collected every 3 months from all the patients, was investigated for the presence of antinuclear (ANA), anti-smooth muscle (ASMA), anti-mitochondrial (AMA), anti native DNA (nDNA) anti-cardiolipin (aCL), anti-parietal cells (APCA), anti microsomal (AMC) and anti-tireoglobulin (ATG) antibodies. Among the IFN-beta 1b treated MS patients an increase of the frequency and of the level of ANA, AMC and ATG was observed. ASMA and ANA antibodies were already present in about 45% of the MS patients before the treatment and fluctuated over the time. In one patient the treatment was interrupted after 6 months because of the occurrence of high ASMA level and of an autoimmune hepatitis. The data obtained in the smaller number of MS patients treated with IFN-beta 1a were very similar. No increase in aCL level was observed during both the IFN treatments. Our results indicate that the treatment with IFN-beta induces an increase of AMC and ATG antibodies in MS patients and confirm that, although rare, autoimmune diseases could be observed. The possible effect of these auto antibodies on the treatment efficacy and on MS clinical course need to be further investigated. PMID- 10871788 TI - Endothelin and nitric oxide levels in cerebrospinal fluid of patients with multiple sclerosis. AB - In order to investigate the potential role of endothelins (ETs) and nitric oxide (NO) in the pathogenesis of multiple sclerosis (MS) we evaluated the levels of these vasoactive mediators in cerebrospinal fluid (CSF) of relapsing remitting MS patients and in a group of subjects with other neurological diseases (OND) and in a control group of subjects without neurological disease. Eighty patients affected from clinically diagnosed MS were selected, 44 of them were studied during an acute clinical attack and 36 in a stable phase. The OND group included 21 subjects affected by degenerative non inflammatory (n=9) and inflammatory (n=12) neurological disease while the control group included 22 subjects with cancer of the prostate (n=11) and with bladder disease (n=11). ET levels were significantly increased in CSF of relapsing remitting MS patients with an acute clinical attack in comparison with those in a stable phase, the OND group and the control group. Moreover significant differences were observed among the four groups with regard to the NO levels: MS patients in a stable and acute phase like OND group have high levels of NO compared to the control group. Since the blood brain barrier index values did not differ significantly between the three groups, the data of this study suggest an important role for NO and ET in cerebral microcirculation in MS patients. PMID- 10871789 TI - Particle-associated retroviral RNA and tandem RGH/HERV-W copies on human chromosome 7q: possible components of a 'chain-reaction' triggered by infectious agents in multiple sclerosis? AB - Different groups have observed retrovirus particle (RVP) production in cell cultures from patients with multiple sclerosis (MS). This in vitro production appeared relatively specific for MS versus healthy controls, but was likely to be enhanced or activated by infectious triggers such as Herpesviruses (e.g. HSV, EBV). Independent molecular analysis of retroviral RNA associated with RVP revealed two different genetic families of endogenous retroviral elements (HERV): MSRV/HERV-W and RGH/HERV-H. Interestingly, these sequences were detected by mutually exclusive primers in RT - PCR amplifications. Surprisingly, these two HERV families both contain an ancestral proviral copy inserted in chromosome 7q21 22 region at about 1 kb of distance of each other. Another HERV-W proviral sequence is located within a T-cell alpha-delta receptor (TCR) gene in chromosome 14q11.2 region. Interestingly, these two regions correspond to genetic loci previously identified as potentially associated with 'multigenic' susceptibility to MS and TCR alpha chain genetic determinants have been reported to be statistically associated with MS. A plausible role for infectious agents triggering a co-activation of the chromosome 7q HERV tandem (replicative retrovirus and/or other virus and/or intracellular bacteria) and, eventually, other HERV copies, is discussed. The role of particular HERV polymorphism and the production of pathogenic molecules (gliotoxin and superantigen) possibly associated with retroviral expression are also evoked. An integrative concept of pathogenic 'chain-reaction' in MS involving several step-specific pathogenic 'agents' and 'products' somewhat interacting with particular genetic elements would federate most partial data obtained on MS, including retroviral expression. PMID- 10871790 TI - The association between multiple sclerosis and infection with Epstein-Barr virus and retrovirus. AB - B-lymphoblastoid cell-lines may develop spontaneously in mononuclear cells from patients with multiple sclerosis, an observation rarely seen in healthy individuals. Examination of such spontaneously established B-cell lines reveal the presence of Epstein-Barr virus and retrovirus particles. We have speculated that in predisposed individuals, a dual infection with retrovirus and late acquired Epstein-Barr virus plays an aetiological role in the development of multiple sclerosis. This hypothesis is supported by a number of observations, including the finding that infection with Epstein-Barr virus may be a prerequisite for developing multiple sclerosis. The association between multiple sclerosis and infection with Epstein-Barr virus and retrovirus is evaluated in this study. PMID- 10871791 TI - Endogenous retroviruses and multiple sclerosis. AB - Endogenous retroviruses are normal constituents in vertebrate genomes. They have been associated with various diseases of presumed autoimmune etiology. However, conclusive evidence of their significance as susceptibility factors in these diseases is still lacking. In our laboratory we have focused attention upon endogenous retroviruses as candidate genes in multiple sclerosis. In this communication we describe general properties of endogenous retroviruses and we present observations from some of our studies. PMID- 10871792 TI - Extended observations on the association of HHV-6 and multiple sclerosis. AB - Throughout the years, a long list of viruses has been associated with multiple sclerosis (MS), however no virus to date has been definitively identified as the etiologic agent of this disease. Recently, human herpesvirus 6 (HHV-6), a newly described herpesvirus, has been suggested to play a role in MS based on: immunohistochemical demonstration of HHV-6 in MS plaques, increased antibodies response to HHV-6 in sera and CSF of MS patients, and the demonstration of HHV-6 DNA in the serum of MS patients but not in normal individuals. To extend these observations we have focused our research in multiple directions. We have increased the number of MS patients tested for HHV-6 serum DNA providing confirmation of our previous study. Additionally we have investigated a possible correlation between HHV-6 viremia and clinical activity. Finally to provide insight into the pathogenesis of this disease, we have begun to characterize the cellular immune response of MS patients to HHV-6. Collectively these studies will help to define the role that HHV-6 may play in the pathogenesis of MS. PMID- 10871793 TI - Novel human herpesviruses and multiple sclerosis. AB - It has been suggested that human herpesvirus 6 (HHV-6) might be involved in the pathogenesis of multiple sclerosis (MS). However, studies of the association between HHV-6 and MS are hindered by the difficulty in discriminating between latent and active infection. We undertook a study to determine whether HHV-6 establish a systemic active infection in the course of MS, and to investigate possible roles of HHV-7, a herpesvirus closely related to HHV-6. To discriminate between latent and active infection, we analysed viral transcription. The results indicate that both viruses are prevalent in PBMCs of MS patients as in healthy controls, and that viral sequences are maintained in a non-transcriptional state. These observations indicate that further studies should define the state of viral persistence in the central nervous system. PMID- 10871794 TI - Human polyomavirus JCV and expression of myelin genes. AB - Myelin basic protein (MBP) is a major component of the myelin sheath of both the central and peripheral nervous systems. A number of neurological diseases in humans are associated with demyelination of the central and/or peripheral nervous systems, including multiple sclerosis and its variants such as acute disseminated encephalomyelitis (AD), acute hemorrhagic leukoencephalopathy, and idiopathic polyneuritis (Guilliame-Barre syndrome), as well as tropical spastic paraparesis (TSP), and progressive multifocal leukoencephalopathy (PML). Multiple sclerosis (MS) is perhaps the most common demyelinating disease and is one of great importance to the clinical neurologist. The underlying cause of the demyelination seen in multiple sclerosis patients is unknown. However, patients frequently have unusually high antibody titers to a number of common viruses, leading to speculation that viral infections may participate in the pathogenesis of MS. On the other hand, studies on maternal and paternal twins have suggested the involvement of genetic factors in the predisposition of an individual toward developing MS. PML, once a rare demyelinating disease of elderly patients with lymphoproliferative disorders, is now a much more common disease affecting patients of all ages due to the increasingly widespread use of immunosuppressive chemotherapy and the prevalence of AIDS. PML is the result of productive infection of oligodendrocytes, the myelin producing cells of the CNS, with the human polyomavirus, JCV. In this article, we have focused our attention on PML, and the role of JCV in disrupting myelin sheaths by affecting myelin basic gene expression, ultimately leading to demyelination. PMID- 10871795 TI - Progressive multifocal leukoencephalopathy: JC virus induced demyelination in the immune compromised host. AB - Progressive multifocal leukoencephalopathy (PML) is a fatal demyelinating disease of the central nervous system that predominantly affects immunocompromised individuals. The etiologic agent, JCV, is a widespread polyomavirus with a very specific target, the myelin-producing oligodendrocytes of the brain. During periods of immune suppression, the virus can be reactivated from lymphoid tissues and kidney, causing targeted myelin destruction and corresponding neurological deficits. The incidence of PML has increased in recent years, due in large part to the advent of AIDS and the growing number of immunodeficient individuals. Furthermore, previous serological studies have shown that greater than 80% of the human population has antibodies to JCV in circulation. When combined, these statistics highlight an increasing need to establish effective treatment regimens for infected individuals as well as strategies to identify those at risk for developing PML. PMID- 10871796 TI - Influence of JC virus coding region genotype on risk of multiple sclerosis and progressive multifocal leukoencephalopathy. AB - Two features of the biology of JC virus make it a particularly suitable candidate for an agent in MS-like disease: its neurotropic capability targeting glial cells as evidenced in progressive multifocal leukoencephalopathy lesions, and its capacity for latency and persistence as illustrated by its behaviour in the kidney. JC virus is chronically or intermittently excreted in the urine by some 40% of the population. The existence of JC virus in multiple coding-region genotypes provides a unique approach to the study of JC virus-induced neurological disease. We have previously shown that a genotype originating in Asia but also present in Europe and the US, called Type 2B, is more frequently found in PML brain than expected based on its prevalence in urine samples from a control population. In contrast, we find that the excretion of JCV in MS patients is similar in both genotype and frequency to that of control individuals, and appears to be regulated by factors unrelated to those that control CNS disease activity. PMID- 10871797 TI - Molecular evidences for a role of HSV-1 in multiple sclerosis clinical acute attack. AB - To verify the possible role of human herpesviruses as triggering or aggravating factors in relapsing-remitting multiple sclerosis (RRMS) clinical acute attack, we studied the prevalence of some herpesviruses in the peripheral blood mononuclear cells (PBMCs) collected from 22 MS patients during an MS relapse and in a stable phase and from 18 healthy controls (HC). DNA belonging to Herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), Human cytomegalovirus (HCMV), Epstein-Barr virus (EBV) and Human Herpes virus 6 (HHV-6) has been searched by specific nested polymerase chain reaction (n-PCR). EBV and HHV6 DNA has been detected with high frequency in acute and stable MS and in healthy controls without significant differences. HCMV DNA was observed both in acute and stable MS but not in HC, and, more interestingly, HSV-1 DNA was only found in 13% of acute MS, while both stable MS and healthy controls were negative. On the basis of these results we focused on HSV-1, and to confirm them and to demonstrate that HSV-1 is actively replicating in MS patients during clinical relapse, we searched both messenger RNA (mRNA) and DNA of HSV-1 in the PBMCs of 15 acute MS patients and 15 healthy controls. We found HSV-1 mRNA and DNA in a significant number of acute MS patients but not in the control group. On the whole these data indicate that HSV-1 reactivate in the peripheral blood of MS patients during clinical acute attack and probably play a role in the triggering of MS relapses. PMID- 10871798 TI - Magnetisation transfer imaging in multiple sclerosis. AB - Magnetisation transfer imaging (MTI) is a magnetic resonance imaging (MRI) technique that has a higher specificity than conventional T2-weighted scans to the heterogeneous pathological substrates of multiple sclerosis (MS) lesions. This review outlines the contribution of MTI in the study of lesion evolution and in the assessment of disease burden in MS. MTI studies of individual MS lesions confirm the pathological heterogeneity of T2-weighted MRI abnormalities and the potential role of unenhanced T1-weighted hypointensities as specific markers of localised severe white matter disruption. Correlative cross-sectional and longitudinal studies using MTI and gadolinium (Gd)-enhanced MRI reveal that MTI findings may vary in lesions with different patterns of enhancement, and that MTI abnormalities are closely related to the onset and recovery of blood-brain barrier disruption in new MS plaques. Measures obtained from MTI scans using whole-brain histogram analysis are highly correlated with the extent of MS abnormalities on conventional MRI scans and predict patients' clinical disability well, since they are sensitive to the amounts of both macro- and microscopic MS disease burden in the whole brain and in specific regions. PMID- 10871799 TI - Proton MR spectroscopy to assess axonal damage in multiple sclerosis and other white matter disorders. AB - Proton MR spectroscopy allows in vivo measurement of N-acetylaspartate in white matter, providing a biochemical index of axonal integrity. Several recent studies of patients with multiple sclerosis and other white matter disorders have shown both transient and sustained decreases in N-acetylaspartate in white matter lesions and in brain regions appearing normal on conventional MRI. These data have emphasised that a substantial amount of axonal damage or loss (presumably secondary to myelin pathology) is consistently present in most of these disorders. Recent post-mortem studies support these results. In contrast to changes seen with conventional MR imaging, decreases in N-acetylaspartate have shown a close correlation with changes in neurological status. This suggests that axonal damage may be more relevant than demyelination for determining chronic functional impairments in primary white matter diseases. Thus, serial measurement of brain N-acetylaspartate with proton MR spectroscopy can provide a reliable and clinically-relevant monitor of disease evolution. As pathological changes responsible for long-term morbidity are logically important targets for therapeutic agents, early treatment directed at axonal protection should be useful in these disorders. PMID- 10871800 TI - MRI of spinal cord in MS. AB - Over the last 10 - 15 years, magnetic resonance imaging techniques have had a major impact in understanding and managing multiple sclerosis. The present review briefly summarises the current usefulness of spinal cord MRI in MS disease, examining the frequency, distribution and main characteristics of spine MS plaques; the differential diagnosis with other spinal cord disease was also described. Finally we considered how newer imaging sequences when added to semi automated quantitative methods, may give us a putative tool to reliably quantify subtle changes which develop on the spinal cord of MS patients over time. PMID- 10871801 TI - Multiple sclerosis in time and space--geographic clues to cause. AB - Geographically MS describes three frequency zones. High frequency areas (prevalence 30+ per 100 000) now comprise most of Europe, Israel, Canada, northern US, southeastern Australia, New Zealand, and easternmost Russia. Medium frequency areas include southern US, most of Australia, South Africa, the southern Mediterranean basin, Russia into Siberia, the Ukraine and parts of Latin America. Prevalence rates under 5 per 100 000 are found in the rest of Asia, Africa and northern South America. Migrants from high to lower risk areas retain the MS risk of their birth place only if they are at least age 15 at migration. Those from low to high increase their risk even beyond that of the natives, with susceptibility extending from about age 11 to 45. Thus MS is ordinarily acquired in early adolescence with a lengthy latency before symptom onset. MS occurred in epidemic form in North Atlantic islands: probably in Iceland and the Shetland Orkneys; clearly in the Faroe Islands. In the Faroes first symptom onset was in 1943, heralding the first of four successive epidemics at 13 year intervals. The disease was presumably introduced by occupying British troops during World War II, with the postwar occurrences representing later transmissions to and from consecutive cohorts of Faroese. What was transmitted is thought to be a specific, widespread, persistent infection called PMSA (the primary multiple sclerosis affection) which only rarely leads years later to clinical MS. Search for PMSA is best attempted on the Faroes where there are regions still free of MS after 50 years. PMID- 10871802 TI - Exogeneous factors in the aetiology of multiple sclerosis. AB - Neuroepidemiology has undoubtedly played a fundamental role in the study of multiple sclerosis (MS) by providing some aetiologic clues, although a definitive basis for the conclusive resolution of its enigma is still lacking. Epidemiological and genetic studies have indicated that MS is probably caused by multiple factors, both genetic and environmental, none of which is individually sufficient, which appear to act before adolescence - or possibly later - in genetically susceptible individuals. This unifying hypothesis emphasizes, on the one hand, the role of a genetic-racial susceptibility and the importance of environmental factors and, on the other, a possible aetiologic heterogeneity and lack of specificity of the unknown endogenous and exogeneous agents. In this context, several environmental factors may be involved in the aetiopathogenesis of MS in individuals who are susceptible to the effect of exposure to these factors. Situations or events with biological plausibility, such as childhood or adolescent infectious diseases, exposures to geographic and socio-cultural factors, nutritional habits, hypersensitivity, significant head and spinal trauma, and other factors may contribute, at different times, to the putative acquisition of MS, trigger its onset, and modify its subsequent course. However, additional empirical evidence is needed to clarify the complex interplay of genetic and environmental factors. PMID- 10871803 TI - Clinical infections and multiple sclerosis: contribution from analytical epidemiology. AB - Epidemiological studies have suggested that exogenous factors may play a role in the etiology of multiple sclerosis and that the environmental component may be viral, but, as yet, there is insufficient evidence to draw any definite conclusions concerning any of the viruses so far proposed. The case-control approach failed to give any definitive conclusion. While the frequency of each common childhood illness is not significantly different between cases and controls, there are more consistent data suggesting that cases do report a later age at infections: this applies particularly to measles, rubella, mumps and EBV infection. Several studies have proved that viral or bacterial infections or reactivations could trigger the clinical attacks in relapsing-remitting MS. PMID- 10871804 TI - Clinico-immunogenetic characteristics of multiple sclerosis with optic neuritis in children. AB - The frequency of multiple sclerosis (MS) with clinical onset before 16 years of age in different regions of Russia fluctuates from 2 to 10% of all MS patients. One of the most frequent signs of MS manifestation and/or exacerbation at this age is optic neuritis (ON). Forty-seven children with MS were observed in Moscow. Diagnosis of MS in every case was clinically definite and proved by serial MRI. Clinico-tomographic dissociation was noticed: numerous large lesions, typical for MS on T2 images were often seen in children with mild or moderate residual neurological symptoms. All patients had relapsing/remitting MS course, mean EDSS was 2.24+/-0.26. Thirty-eight children (80%) had ON at least once, ten (21.3%) - twice or more times. In several cases ON had subclinical course or might be missed and the damage of the optic nerve with partial atrophy was found only after complex ophthalmological investigation including visual evoked potentials. Thus, the clinical course of MS and ON have some peculiarities in children and may be genetically based. Analyses of allelic polymorphisms of HLA-DR and TNF loci on chromosome 6 was performed. Data from children with MS were compared with data from their parents, healthy controls and other MS patients from the same ethnic group. Children with MS had increased frequency of DR2(15) and TNF-a11, but not TNF-a9 as adult MS patients from the same ethnic group. The presence of TNF-a7, rare in adult patients, could be proposed as a marker of early MS onset. PMID- 10871806 TI - Mood disorders in multiple sclerosis: diagnosis and treatment. AB - Emotional disturbances are common in MS and consist of disturbances of mood and disturbances of affect. The important mood disorders are major depressive disorder, dysthymic disorder, bipolar disorder, panic disorder, and generalized anxiety disorder. Their relationship to MS is multi-factorial and complex, and the extent to which they are direct consequences of the disease process or psychological reactions to it remains unclear. Whatever their cause, however, the symptoms of mood disorders in people with MS are no different from the symptoms of mood disorders in people without MS, and respond just as well to standard treatments. The disorders of affect are euphoria, pathological laughing and weeping, and other frontal lobe syndromes. These disorders result from demyelination, are some of the most characteristic symptoms of MS, and have the same implications for treatment as do other aspects of the disease. Mood and affective disturbances can cause enormous pain and suffering and lead to significant disruption of family, work, and social life. Physicians who can identify, diagnose, treat, and manage mood and affective disturbances effectively and who can help their patients and family members acknowledge these difficulties, talk about them, and accept psychiatric consultation and treatment can have a dramatic impact on the quality of their lives. This paper outlines the symptoms and diagnostic criteria for mood disorders and affective disturbances, reviews current treatment options, summarizes data from epidemiologic and pathophysiological studies, and suggests areas for future research. PMID- 10871805 TI - Grading brainstem involvement in multiple sclerosis - by means of electro oculography. AB - One of the most frequent disorders of the brainstem in multiple sclerosis (MS) is internuclear opthalmoplegia (INO). The aim of this study is to show how it is possible to monitor the course of MS grading INO on the basis of electro oculographic findings. We selected 130 patients with a diagnosis of clinically defined multiple sclerosis (78 males and 52 females, mean age 43.5 years) from a population of 354 MS patients. Both saccadic eye movements and spontaneous, vestibular (VOR), visuo-vestibular (VVOR) and optokinetich nystagmus (OKN) were assessed. Slowing of the adducting eye was considered as a sign of lesion of the interocular pathways. Statistical analyses showed that the most sensitive test was VVOR, the least sensitive being randomised saccades. An impairment of random saccades was always associated with abnormal results on all other tests. It seems thus possible to grade the involvement of the medial longitudinal fasciculi (MLF) in MS from an abnormality limited to the VVOR test up to an impairment of randomised saccadic movements. Grading brainstem involvement in MS is particularly important in therapeutic trials and during rehabilitation. PMID- 10871807 TI - Group psychotherapy experiences for people with multiple sclerosis and psychological support for families. AB - In this article we describe our experience of group psychotherapy for patients afflicted with multiple sclerosis in the care of the Centre of Multiple Sclerosis of the Don Gnocchi Foundation in Milan, and an experience of psychological support to the patients' families. Multiple sclerosis, with its disability, chronic and unforeseeable features, brings about a series of changes in the patient's life. Considering the youthful age during which the disease arises, patients are obliged to review their own life plans in their family, social and working circles and have to face living with the chronicity and the loss of their own autonomy. Group psychotherapy was qualified as a preferred ambit to express and to share individual problems. The disease also involves the family: relatives must revise certains plans and projects made before. Psychological support could help relatives not only with the emotional load following on from the disease so that they do not feel alone in the patient's everyday care, but also with their own emotional management, in order to give them a listening space also for their distresses and fears. PMID- 10871808 TI - MRI correlates of cognitive dysfunction in multiple sclerosis patients. AB - Studies with conventional magnetic resonance imaging (MRI) support the hypothesis that cognitive impairment in multiple sclerosis (MS) patients is related with the lesion burden. Patterns of frontal lobe cognitive decline were also found to be related with the corresponding regional lesion load, although the total lesion load on T2-weighted MRI scans of the brain seems to be more relevant in determining frontal lobe deficits. Other non-conventional MRI techniques with a higher specificity to the heterogeneous substrates of MS pathology, such as the assessment of hypointense lesion load on T1-weighted scans and the histogram analysis of magnetisation transfer ratio (MTR) maps, have recently been applied to MS cognitive studies. Results from these studies suggest that three factors play a role in the pathogenesis of MS dementia: the burden of MS lesions, the severity of the pathological damage within individual lesions and that of the normal-appearing white matter. PMID- 10871809 TI - Medical rehabilitation of chronic progressive disseminated encephalomyelitis (MS). AB - Eight years after diagnosis, 40% of MS patients develop a chronically progressive form. Annually we treat approximately 200 patients with progressive MS. Treatment consists of medication, i.e. agents that help to prevent future impairment, or interferon-beta injections, and intervals of mitoxantrone infusions (Novantrone(R)), and in some cases cyclic cyclophosphamide (Endoxan(R)) or nucleoside analogue cladribin (Leustatin(R)). Without clear scientific evidence, we recommend unsaturated fatty acids (thistle or sunflower oil), sufficient protein, and freshly prepared fruits and vegetables as a sound basis for remyelination. Remyelination profits from general prophylaxis in the use of ascorbic acid to help prevent urinary infections via acidification, autogenic training to reduce fatigue, improve ventilation of deeper airways, and stimulate vagotonic regeneration, and prevention of unnecessary immune stimulation caused by insects and some food. We recommend the use of sun hats and disencourage blood donation (Allain 1998). Physiotherapy can improve strength, reduce spasticity, and train the patient to compensate for dysbalance and ataxia; supported by beta blockers and good antispastics, tremor and gait disturbances can be positively influenced. Music and motion, speech therapy, realistic training of daily activities, and prudent psychotherapy complete the range of measurements to reconstitute as much as possible of the patient's individual freedom. In the individual, we eventually provide prudent technical aids and careful prognostic estimations. Cooperating with local and regional patient networks, we reinforce long-term disease management and spread up-to-date medical research results, and finally gather valuable contextual information and clinical data on an increasingly frequent idiopathic disease of the human central nervous system. PMID- 10871810 TI - Update on multiple sclerosis rehabilitation. AB - The neuromotor rehabilitative treatment should be joined in the global individualised project. The rehabilitation project is developed by an interdisciplinary. Hence, the clinicians have to promote an appropriate compensation process in the daily living activities with a rapid insertion of new or recovered motor-postural patterns. The model of neuromotor intervention on patients suffering from MS provides a series of important moments: (1) a specific evaluation of the motor and postural behaviour; (2) the identification of targets and consequently the treatment planning; (3) the carrying out of a therapeutic program in different kinds of management; (4) the dynamic monitoring; (5) the experimentation of new therapeutic models and new rehabilitative hypothesis; (6) the placing of a neuro-psycho-motor treatment in a global therapeutic project. Recently we have investigated the performances and the efficiency of a particular therapeutic manoeuvre for the evocation of absent motor components in an experimental group of patients with paretic-atactic features and we have studied the correlations with the results observed in a control group of patients with similar pictures, treated with stretching exercises: the study is described and the first results are reported. Furthermore we have studied the therapeutical use of a stabilometric platform, 'BIOGP', on patients with ataxia. The platform is described and the first clinical experience is reported with the protocol of evaluation. PMID- 10871811 TI - Cognitive function and quality of life in multiple sclerosis patients. AB - This is a study on the longitudinal evaluation of cognitive functions in multiple sclerosis (MS) patients and their quality of life (QoL). The study follow-up lasted for 3 years and the evaluation was performed every 9 months for four times altogether. We present data on the first and second session, when we evaluated the frontal component of cognitive functions, behavioural memory and quality of life. We administered the Luria Frontal Lobe Syndrome test (LFLS), the Rivermead Behavioural Memory Test (RBMT), the State-Trait Anxiety Inventory (STAI), the Beck Depression Inventory (BDI), SF-36 for QoL evaluation. The frontal component of cognitive functions and behavioural memory involvement is related to a worsening of QoL, in particular in the Physical Functioning and the Mental Health of SF-36. PMID- 10871812 TI - A cost evaluation of multiple sclerosis. AB - As a chronic and disabling disease, multiple sclerosis (MS) is extremely costly, both for the individual and the family, as well as for the society. Early onset, long duration and effects on employment contribute to the extensive costs related to the illness. Thus far, studies conducted in developed countries have demonstrated that direct costs, including treatment (prior to the approval of beta interferon), medical visits, hospitalization, assistance, etc., are much lower in respect to indirect costs, such as loss of income from reduction of work activity for patients and carers, which account for up to 75% of the total cost. Informal care represents a heavy burden for the families of disabled persons and little is known about the 'intangible' costs of MS, such as those related to the influence of the disease on quality of life. In addition, the cost/benefit ratio for expensive new therapies, such as beta interferon, remains to be determined. PMID- 10871813 TI - Viewpoint on the impact of interferon in the treatment of multiple myeloma: benefit for a small proportion of patients? AB - Interferon-alpha (IFN-alpha) generally inhibits myeloma cell growth. However, a growth stimulatory effect for myeloma cells has also been reported. In patients with untreated multiple myeloma (MM) IFN-alpha, used as a single agent, produced an objective response rate ranging from 10 to 25%. In previously untreated patients: (1) the time to response is short, (2) the median duration of response is similar to the duration of response observed in patients given chemotherapy, and (3) the patients who are more likely to benefit are those with IgA myeloma type. Concerning the results of IFN-alpha given as a single agent in relapsing and resistant MM, they are poor, with a response rate ranging between 10-20%. The combination of high-dose glucocorticoids and IFN-alpha for relapsing/resistant patients produced controversial results. Some studies showed an increased response rate and/or longer survival with chemotherapy plus IFN-alpha versus chemotherapy alone in previously untreated patients. In contrast, most reports did not show a significant increase in response rate or survival benefit by adding IFN-alpha to the initial chemotherapy. Perhaps the most encouraging role for IFN in MM is as maintenance therapy in patients responding to first line treatment (ie conventional chemotherapy followed or not by high-dose intensification/autotransplantation). In spite of that, several reports failed to show longer response duration. The majority of studies have shown a modest but significant prolongation in response duration in favour of the IFN arm. However, most of these studies have failed to show a significant survival advantage with IFN maintenance. A meta-analysis, by the Myeloma Trialists' Collaborative Group in Oxford, based on the individual data from 4012 patients included in 24 randomized trials (induction and/or maintenance) has shown that IFN produced a moderate improvement in relapse-free survival and a minor improvement in overall survival. In summary, the only role of IFN in MM is as maintenance treatment after a response is achieved. However, looking at the published data, it seems that the vast majority of patients do not benefit from IFN maintenance, while a small proportion of them, in the range of 5-10%, obtain a significant prolongation in event-free survival and overall survival. Unfortunately, there are no predictive factors that can identify the patients who are likely to benefit from IFN maintenance. PMID- 10871815 TI - Phase II study of temozolomide in patients with relapsing high grade glioma and poor performance status. AB - Temozolomide (SCHS2.365), an oral alkylating agent which penetrates the blood brain barrier, evolved as an alternative to dacarbazine. The aim of this study was to evaluate the efficacy and safety of temozolomide in terms of overall survival, progression-free survival, clinical benefit and health related quality of life in symptomatic patients with relapsing malignant glioma and a poor performance status. Eleven patients were enrolled in the study. The median age was 44.6 years. Patients were treated with temozolomide per os at a dose of 150 200 mg/m2 daily for 5 consecutive days. Each cycle was repeated every 28 days. The median number of courses given per patient was 3.5. Nine patients were assessable for response. All patients were evaluable for toxicity. Based on radiographic findings 4 patients had stable disease (2 patients after a total of 16 cycles, and 2 patients after a total of 10 cycles). Four patients had progressive disease after 2 to 4 cycles. Of these 3 patients demonstrated a clinical benefit and one patient died after 3 cycles of treatment. Six patients had a significant clinical benefit even after 2 cycles of treatment with improvement of their neurological and performance status. Hematologic toxicity Gr II-III occurred in 3/9 patients. Nonhematologic toxicity consisted of Gr I nausea, and vomiting. In conclusion temozolomide appears to be a useful alternative for patients with relapsing malignant glioma after radiation and surgery and a poor performance status with little or no toxicity and considerable clinical benefit. PMID- 10871816 TI - Infections in acute leukemia: an analysis of 240 febrile episodes. AB - Infections are the major cause of morbidity and mortality in acute leukemia patients. Case records of 91 consecutive patients (AML-48, ALL-40, RAEB-t/AML-3) treated between January 1997 and July 1999 were studied to determine the type, frequency and severity of infections. Patients' median age was 36 y (range 6-66) and male to female ratio was 2.5:1. A total of 240 febrile episodes were recorded; of them, 162 were associated with neutropenia (absolute neutrophil count, ANC<500/mm3) and 78 were without neutropenia. Among the neutropenic episodes, an infectious etiology could be documented in 52%; the remainder (48%) were defined as isolated febrile episodes. Chest was the most common site of infection (35. 7%) followed by skin, soft tissue (13%), GIT (7%) and genitourinary tract (6%) infections in order of decreasing frequency. Microbiologically, gram positive organisms (staphylococcus aureus, coagulase negative staphylococcus, streptococcus, enterococcus) were the most common isolates (52.8%) followed by gram negative organisms (E. coli, klebsiella, pseudomonas) in 42.8% of isolates. Two patients had pulmonary tuberculosis and three patients had fungal infections (candida-2, aspergillus-1). Among non neutropenic patients, infection could be documented in 36%; the remaining 64% were isolated febrile episodes. Gram negative infections were documented in 50%, gram positive in 30% and fungal infections (candida-4, aspergillus-1, mucormycosis-1) in 20% of them. A combination of third generation cephalosporin and an aminoglycoside were used in 79% of episodes initially; a combination of a newer penicillin and aminoglycoside (4.6%), double betalactums (4.1%), oral antibiotics (9.8%) and others were used in the remaining episodes. Fever resolved in 38% of episodes using the above combinations; in the remainder second line antibiotics (mainly vancomycin) and antifungals (amphotericin-B) were added empirically or depending on culture and sensitivity. In 52.5% of episodes fever resolved after addition of second line antibiotics and antifungals. 11 of 91 patients died of infectious complications in this study. There is a need for improvised diagnostic tests to detect infections early as well as for new therapies to overcome antimicrobial resistance. PMID- 10871814 TI - Prostate cancer: a comprehensive review. PMID- 10871817 TI - Urinary albumin excretion and transcapillary escape rate of albumin in malignancies. AB - Transcapillary escape rate of albumin was determined in 22 patients with different malignancies. In addition, urinary albumin excretion rate was measured in 24-h urine samples using a sensitive immunoassay. Increased urinary albumin excretion was defined as >/=20 microg/min according to conventional standards. Renal glomerular filtration and tubular function was estimated by 51Cr-EDTA plasma clearance and urinary beta 2-microglobulin, respectively. Median urinary albumin excretion rate was 15.0 microg/min (range 6-510 microg/min) and the frequency of increased urinary albumin excretion was 41%. This agrees with other studies showing increased albuminuria in several types of malignant diseases. Patients with advanced disease (tumour, node, metastasis (TNM) stage II-IV) had a significantly higher urinary albumin excretion rate than patients with localized disease (TNM stage I). Serum creatinine, glomerular filtration rate and urinary beta 2-microglobulin were all within normal limits. Median transcapillary escape rate of albumin was 5.5 %/h (range 2-8 %/h) and this level is comparable with values in healthy subjects. There was no significant difference in transcapillary escape rate between patients with elevated urinary albumin excretion and the normoalbuminuric group. Median value of the absolut outflux of albumin was 10.6 g/h with similar levels in patients with increased urinary albumin excretion and patients with normoalbuminuria. Our results indicate a high prevalence of minor glomerular dysfunction with a slightly elevated urinary albumin excretion in patients with malignancies. The normal endothelial function, as estimated by the transcapillary escape rate of albumin, suggests an overall unaffected capillary permeability and increased urinary albumin loss appears to be an isolated renal phenomenon in cancer patients. PMID- 10871818 TI - Prevalence of hepatitis-G virus and hepatitis-C virus infection in patients with non-Hodgkin's lymphoma. AB - Several studies have reported that hepatitis-C virus may have a role in the development of non- Hodgkin's lymphoma. Hepatitis-G virus has hepatitis-C virus like characteristics. The possible association between hepatitis-G virus infection and non-Hodgkin's lymphoma is not clear. The aim of this study was to determine the prevalence of hepatitis-G virus and hepatitis-C virus infection in patients with non-Hodgkin's lymphoma without blood transfusion. Forty-four patients with non-Hodgkin's lymphoma were enrolled in the study. Serum samples derived from the patients were tested for antibodies against hepatitis-C virus by ELISA. Hepatitis-G virus and hepatitis-C virus RNA were detected by reverse transcription-polymerase chain reaction. Only two of 44 patients (5%) with non Hodgkin's lymphoma were positive for Anti- HCV and hepatitis C virus RNA. One patient had low grade non-Hodgkin's lymphoma with follicular mixed histopathology while the other had intermediate grade with diffuse large cell histopathology. Hepatitis-G virus infection was detected in none of the patients. We concluded that hepatitis-G virus does not seem to be in association with non-Hodgkin's lymphoma. PMID- 10871819 TI - Restaging with gallium scan identifies chemosensitive patients and predicts survival of poor-prognosis mediastinal Hodgkin's disease patients. AB - Following treatment of mediastinal Hodgkin's disease (HD), residual masses are frequent and gallium scanning has proven to be of value in the evaluation of their specificity (fibrosis or active disease). This study assessed, for relapse and survival, the predictive value of restaging gallium scan of patients with a residual mass on computed tomography scan after induction chemotherapy. Between 1/89 and 12/97, in 53 newly diagnosed HD patients with a residual mediastinal mass, a gallium scan was performed after chemotherapy (3 or 4 courses) and always before consolidative radiotherapy. Characteristics at diagnosis were: nodular sclerosis histology, 89%; bulky mediastinal disease, 79%; B-symptoms, 51%. RESULTS: gallium scan was positive in 16 patients (30%) and negative in 37 (70%). At median follow-up period of 36 months, freedom-from-progression rate was 86% versus 19% (P<0.0001) for patients with negative vs positive gallium scans, respectively. The 5-year overall survival (OS) rate was 68% and differed significantly (P<0.0001) between negative (91%) and positive (25%) gallium scanning groups. The specificity of gallium scanning was 91% and the sensitivity 72% with a positive predictive value of 81% and a negative predictive value of 86%. Evaluation with gallium scan after induction chemotherapy identifies chemosensitive patients among those with poor-prognosis mediastinal HD. Although relapse may occur in patients with negative gallium scan, a postive gallium scan is highly predictive of failure and poor outcome, and treatment should thus be modified. PMID- 10871820 TI - Phase I/II dose escalation study of docetaxel and carboplatin combination supported with amifostine and GM-CSF in patients with incomplete response following docetaxel chemo-radiotherapy: additional chemotherapy enhances regression of residual cancer. AB - Taxanes have been shown to interact with anti-apoptotic proteins. In the present study we investigated whether the addition of taxane in combination with DNA damaging drugs can further enhance tumor shrinkage in cases with incomplete response to radiotherapy. Since the dose of docetaxel in combination with carboplatin is not known, the above hypothesis was tested in the context of a dose escalation phase I study. Twenty-eight patients with locally advanced chest or pelvic tumors, showing residual disease on CT scans performed 40 d following docetaxel radio-chemotherapy, were recruited in a dose escalation protocol of docetaxel/carboplatin supported with amifostine and GM-CSF. The starting dose of docetaxel was 40 mg/m2 every 2 weeks. Carboplatin dose was calculated using the Calvert formula and was escalated in cohorts of 4 patients (starting dose AUC2 every two weeks; AUC0.5 increments up to AUC3). Thereafter the docetaxel dose was increased to 50 and 60 mg/m2, while carboplatin was escalated (by AUC0.5 increments) starting from AUC3 and AUC4 respectively. Amifostine (600 mg/m2) was administered i.v. before carboplatin and GM-CSF (480 microg) was injected s.c. on days 5, 6 and 10, 11 of each cycle. Six cycles were given and response was assessed 2 weeks after the end of chemotherapy. None out of four patients treated in the 6th dose level cohort (50 mg/m2 of docetaxel and AUC4 of carboplatin every 2 weeks) showed any grade 2-4 hematologic toxicity. Mild non-hematologic toxicity such as neuropathy, leg edema, pleural effusion, pyrexia, alopecia grade 2 and hypersensitivity was observed in 4-12% of patients. Out of four patients treated in a 7th cohort (docetaxel 60 mg/m2 and carboplatin AUC4), one developed grade IV neutropenia and two developed grade 3 severe asthenia requiring treatment delay for 2 weeks. Out of 11 patients with PR following docetaxel radio-chemotherapy, 7 (63%) showed CR after docetaxel/carboplatin additional chemotherapy. Eight out of 17 patients with MR following docetaxel radio-chemotherapy showed PR (47%) and one showed CR (6%) after additional chemotherapy. High dose combined docetaxel (50 mg/m2) and carboplatin (AUC4) chemotherapy can be safely administered on a two-weekly basis if supported with amifostine and GM-CSF. Such an additional therapy may be important in patients with incomplete response after chemo-RT. Broad spectrum cytoprotection with amifostine and GM-CSF may also contribute to the reduction of incidence of neurosensory reactions and asthenia in patients treated with taxanes. PMID- 10871821 TI - Granular acute lymphoblastic leukemia in a 15-year-old boy. AB - We report a case of a boy with acute lymphoblastic leukemia expressing granular inclusions in the cytoplasm of blastic cells. Granular lymphoblasts had mostly L2 morphology but azurophilic granules were also present in part of the cells with L3 morphology. Immunophenotyping clearly indicated a lymphoid origin of the blasts and showed positivity of HLA-DR, CD10, CD19 and CD24 markers. Cytogenetic analysis brought normal karyotype 46 XY. Molecular genetic analysis showed immunoglobulin heavy chain rearrangement (IgH R/R) and T-cell receptor gamma rearrangement pattern (TCR-gamma C/R). TCR-beta and TCR-delta did not show rearrangements. The course of the disease was favourable. The patient achieved initial complete remission within 4 weeks of protocolar treatment and the remission remains until now, 5 years from the diagnosis and three years after finishing the treatment. PMID- 10871822 TI - Cardiac metastasis from a transitional cell carcinoma: a case report. AB - We report the case of a patient with a metastatic tumor in the right ventricle, apparently derived from a transitional cell carcinoma. The patient presented with severe hypoxemia as a result of right-to-left shunt due to the position of the tumor and a patent foramen ovale. The clinical course of this case is presented and the pathophysiology of the physiological effects caused by the metastatic tumor is discussed. The literature concerning cardiac metastases is reviewed. PMID- 10871823 TI - Antinuclear antibodies and response to IFNbeta-1a therapy in relapsing-remitting multiple sclerosis. AB - We determined whether positive ANA was related to response to rIFNss-1a in 62 relapsing-remitting MS patients. According to the presence of antinuclear antibodies (ANA) at baseline and during the first 6 months of treatment, patients were sorted in different groups. The clinical and MRI outcome during short-term (6 months) and long-term (24 months) treatment period was not statistically different between the groups. Therefore, the response to IFNbeta-1a seems not to be influenced by ANA occurrence either before or during treatment. When the analysis was extended to other autoantibodies (i. e. antithyroid, anticardiolipin) similar results were obtained. PMID- 10871824 TI - T cell receptor beta chain genotyping in Australian relapsing-remitting multiple sclerosis patients. AB - This study focused on susceptibility to MS within the beta-chain of the T-cell antigen receptor (TCRB locus, 7q35) in a cohort of 122 RR-MS patients compared with 96 normal individuals using biallelic polymorphisms across the bv8s1(Vbeta8.1) to bv11s1 (Vbeta11) TCRB subregion. The markers bv6s5, bv8s1, bv10s1, bv15s1 and bv3s1 were studied for allele and genotype frequencies; haplotypes were assigned with combinations of two of these markers and stratification for HLA-DR15 was also performed. Linkage disequilibrium was found between alleles of the bv8s1, bv10s1/bv15s1 and bv3s1 loci in both patients and controls. An increase among RR-MS patients in the allele frequency of bv8s1*2 (P=0.03) and the haplotype bv8s1*2/bv3s1*1 (P=0.006) was noted and both were found to be statistically significant. In the DR15-positive group, the association between TCRB and MS was seen with the bv8s1*2 allele (Puc=0.05) and the bv8s1*2/bv10s1 haplotypes (Puc=0.048), while the haplotype associations seen among DR15-negative RR-MS patients included the bv3s1*1 allele (bv10s1*1/ bv3s1*1, Puc=0.022; bv8s1*2/bv3s1*1, Puc=0.048). These results support the involvement of the TCRB region in MS susceptibility and encourage further study of the variable gene segments in this region. PMID- 10871826 TI - Lesion pattern in patients with multiple sclerosis and depression. AB - To assess if a specific lesion pattern or changes of the basal limbic system as seen in primary depression and depression associated with neurodegenerative disorders might be identified in depressive multiple sclerosis (MS) patients, we submitted 78 MS patients to a MRI examination consisting of a quantitative measurement of lesions and of hyperintense signals from the pontomesencephalic midline (raphe). Furthermore relaxometry of the pontomesencephalic midline, a transcranial ultrasound examination rating its echogenicity semiquantitatively and a standardized neurological, neuropsychiatric and neuropsychological assessment were obtained. Thirty-one patients fulfilled the DSM-IV criteria for depression. Depressed MS patients had a significantly larger temporal lesion load than non-depressed MS patients, especially on the right side. A trend of difference was detected for lesions of the right parietal lobe, the right frontal lobe, the cerebellum and the total lesion load. Neither hyperintense signals or relaxometry nor echogenicity of the region at the level of the pontomesencephalic midline were significantly different between the groups. We conclude that depression in MS patients is not associated with an alteration of the basal limbic system at the brainstem as seen in Parkinson's disease or unipolar depression but with an increased lesion load of the projection areas of the basal limbic system. PMID- 10871825 TI - 1H MRSI comparison of white matter and lesions in primary progressive and relapsing-remitting MS. AB - OBJECTIVE: To compare brain metabolite levels in patients with primary progressive (PP) and relapsing remitting (RR) MS and controls. HYPOTHESES: (1) creatine (Cr), a putative marker of gliosis, is elevated and N-acetylaspartate (NAA), a putative marker of axonal density and functional integrity, is reduced in PPMS lesions and normal appearing white matter (NAWM) compared to control white matter; (2) The pattern of metabolite change in PPMS is different than in RRMS. METHODS: MRI and proton magnetic resonance spectroscopic imaging (1H MRSI) were collected from 15 PPMS patients, 13 RRMS patients, and 20 controls. RESULTS: Cr was increased in PPMS NAWM compared to controls (P=0.035), and compared to RRMS NAWM (P=0.038). Cr was increased in focal MRI lesions from PPMS compared to lesions from RRMS (P=0.044) and compared to control white matter (P=0.041). NAA was similarly reduced in PPMS and RRMS NAWM compared to control. NAA was similarly reduced in PPMS and RRMS lesions, compared to control white matter. CONCLUSIONS: Creatine is higher in PPMS than RRMS NAWM and focal lesions. This observation is consistent with the notion that progressive disability in PPMS reflects increased gliosis and axonal loss whereas disability in RRMS reflects the cumulative effects of acute inflammatory lesions and axonal loss. PMID- 10871827 TI - New low-contrast vision charts: reliability and test characteristics in patients with multiple sclerosis. AB - The quantitative assessment of visual function in multiple sclerosis (MS) clinical trials has been limited to Snellen visual acuity. The purpose of this study was to examine the inter-rater reliability and test characteristics of a new visual outcome measure, the Low-Contrast Sloan Letter Charts, in patients with MS and visually-asymptomatic volunteers. Contrast letter acuity scores (letter scores) were measured at each of four contrast levels (100, 5, 1.25 and 0.6%) by two independent raters. Inter-rater agreement was described with the intraclass correlation coefficient (ICC) and comparison of mean scores. Excellent inter-rater agreement (ICC=0.86 - 0.95) was demonstrated at each contrast level among MS patients (n=100) and visually-asymptomatic volunteers (n=33). Average letter scores at the lowest contrast level (0.6%) were highly variable in the MS group, even among patients with visual acuities of 20/20 or better, and among those who required no assistance for ambulation. Low-Contrast Sloan Letter Chart testing is a highly reliable method of visual assessment, and provides information on an aspect of neurologic impairment in MS which is not captured by Snellen visual acuity or ambulation status. This new method demonstrates excellent potential as a visual function outcome measure for future MS clinical trials. PMID- 10871828 TI - The Danish National Project of interferon-beta treatment in relapsing-remitting multiple sclerosis. The Danish Multiple Sclerosis Group. AB - After approval by the European Union in 1996 of interferon-beta for treatment of relapsing-remitting multiple sclerosis (RRMS), 862 patients in Denmark have received treatment (by April 1999), and 304 of those were enrolled into an open labelled randomised trial to compare the efficacy and tolerability of treatment with interferon-beta-1b (Betaferon) 8 MIU subcutaneous every other day and interferon-beta-1a (Rebif) 6 MIU subcutaneous once a week. Primary and secondary end-points included: (1) the relapse rate, (2) tolerability, (3) neutralizing antibody formation using the same specialised laboratory and validated assay, (4) time to first relapse, (5) time to sustained progression, and (6) number of gadolinium-enhancing lesions in T1-weighted MRI and the total lesion load in T2 weighted MRI. All the records are kept in a nationwide clinical database connected with the Danish Multiple Sclerosis Registry. The randomised trial will be completed by the end of 1999, and the results are not yet available. The annual relapse rate in all treated patients until April 1999 was 0.73, the annual rate of steroid treated relapses was 0.21. By 1 April 1999 8.8% of the patients had discontinued treatment and 10.4% had changed to another interferon-beta preparation. It has been planned to extend the follow-up of all patients treated according to the Danish National Protocol for a period of several years. The continuous and nation-wide registration of the course of the disease in interferon-beta treated RRMS patients may provide valuable knowledge of the treatment-modified long-term course of the disease in a cohort of patients, selected on the basis of well-defined criteria. PMID- 10871829 TI - Effect of precooling on physical performance in multiple sclerosis. AB - Many individuals with MS experience heat sensitivity that may be associated with transient increases in the frequency of clinical signs and symptoms. Although physical activity may be beneficial for those with MS, induced thermal loads may preclude participation in exercise and other daily activities. This project was designed to evaluate the effects of precooling on physical function. Six thermosensitive MS patients were studied. Participants performed a graded exercise test to determine maximal oxygen uptake (VO2max) on a combined arm-leg ergometer. Thermal load was induced by 30 min of exercise under noncooled and precooled conditions at a workrate corresponding to 60% VO2max. Precooling consisted of 30 min lower body immersion in 16 - 17 degrees C water. Fatigue and 25-ft walk performance were assessed before, immediately after, and 30 min following exercise. No treatment differences in VO2 were observed. Rectal temperature, heart rate, and rating of perceived exertion (RPE) were significantly lower during the precooled exercise trial compared to the noncooled trial. Immediately following exercise, 25-ft walk performance and fatigue scores showed significantly greater deterioration in the noncooled condition. Precooling was effective in preventing gains in core temperature with physical work and may allow heat-sensitive individuals with MS to exercise with greater physical comfort. PMID- 10871830 TI - Fatigue in multiple sclerosis and its relationship to depression and neurologic disability. AB - We studied multiple sclerosis fatigue (MSF) and its relationship to depression and disability. Seventy-one patients [50 relapsing-remitting, 21 secondary progressive] were grouped by Fatigue Severity Scale (FSS) into MS-fatigue (MSF) (FSS>/=5; n=46) or MS-nonfatigue (MSNF) (FSSAlanine) is completely localized in the nucleus while another Sam68 (Proline439-->Arginine) mutant is found in the cytoplasm. The localization of these Sam68 mutant proteins also correlates with their function in RRE mediated reporter gene expression, i.e. Sam68 mutant protein that is localized in the cytoplasm failed to enhance RRE-mediated transactivation. Furthermore, we demonstrate that Sam68 P439-->R inhibited the transactivation of RRE-mediated gene expression by both wild type Sam68 and Rev. These results indicate that the proline residue at position 439 unlike arginine at position 429, may play a critical role in targeting Sam68 protein to nucleus. We propose that these negative dominant mutants of Sam68 may have potential as anti-viral agents to combat AIDS. PMID- 10871865 TI - p27KIP1 is down-regulated by two different mechanisms in human lymphoid cells undergoing apoptosis. AB - The cyclin-dependent kinase inhibitor p27KIP1 is a crucial component of the mammalian restriction point, and as such is subject to multiple regulatory mechanisms. It has recently been shown that the abundance of p27KIP1 is also regulated during apoptosis; p27KIP1 is cleaved by a Z-VAD-fmk-sensitive caspase during apoptosis induced by growth factor deprivation in endothelial cells, and also following exposure of myeloid leukaemia cells to etoposide. Here, we investigate p27KIP1 regulation in B- and T-lymphoid cells undergoing apoptosis. We observe that p27KIP1 is down-regulated following exposure to a variety of apoptotic stimuli including an agonistic anti-Fas antibody, cycloheximide and etoposide. Further investigation revealed the existence of two different routes of p27KIP1 regulation in lymphoid cells undergoing apoptosis. The first pathway is utilized by lymphoid cells stimulated through Fas, is abrogated in a caspase-8 deficient T-cell line, and is blocked by the caspase inhibitors Z-VAD-fmk and Boc D-fmk. In contrast, the loss of p27KIP1 in cells exposed to cycloheximide and etoposide occurs in the absence of caspase-8 or any Z-VAD-fmk- or Boc-D-fmk sensitive caspase activities. Thus the down-regulation of p27KIP1 is a common occurrence in lymphoid cells undergoing apoptosis but, depending on the apoptotic trigger, this can be affected by two different mechanisms. PMID- 10871866 TI - Multiple endocrine neoplasia type 2B mutation in human RET oncogene induces medullary thyroid carcinoma in transgenic mice. AB - Multiple endocrine neoplasia type 2B (MEN 2B) is a familial cancer syndrome, in which the cardinal feature is medullary thyroid carcinoma (MTC), a malignant tumor arising from the calcitonin producing thyroid C-cells. MEN 2B is associated with a germline point mutation in the RET proto-oncogene, leading to a Met-->Thr substitution at codon 918 in the kinase domain, which alters the substrate specificity of the protein. We used the human calcitonin gene (CALC-I) promoter to generate transgenic mice expressing either the human RET oncogene with the MEN2B-specific 918 Met-->Thr mutation (CALC-MEN2B-RET) or the human non-mutated RET proto-oncogene (CALC-WT-RET) in the C-cells. At 20 - 22 months of age three out of eight CALC-MEN2B-RET transgenic founders presented with macroscopic bilateral MTC. In two founders nodular C-cell hyperplasia (CCH) was observed. Thyroid abnormalities were never observed in CALC-WT-RET transgenic mice or control non-transgenic mice analysed at this age. In some mice from established CALC-MEN2B-RET transgenic lines nodular CCH was observed from 8 months on whereas MTC was detected in 13% of mice from one CALC-MEN2B-RET line, from the age of 11 months on. These results show for the first time that the MEN2B mutation in the RET oncogene predisposes mice for MTC. PMID- 10871867 TI - p51A (TAp63gamma), a p53 homolog, accumulates in response to DNA damage for cell regulation. AB - p51A, or TAp63gamma, a translation product of gene p51, or p63, was identified as a homolog of p53 in its primary structure and transactivating function. p53 plays a decision-making role in inducing either cell cycle arrest or apoptosis in response to DNA damage, and thereby preserves genome integrity of living cells. To compare the biological activities between p51A and p53, cell lines with low level, constitutive expression of each protein were obtained by cDNA transfection of mouse erythroleukemic cells. Production of p51A with an apparent molecular mass of 57-kilodalton (kD) accompanied induction of p21waf1 and appearance of hemoglobin-producing cells. After DNA-damaging treatment either with ultraviolet light (UV) irradiation or with actinomycin D, the p51A protein accumulated in time courses corresponding to those of wild-type p53, and caused an increase in the hemoglobin-positive cell count. In contrast, p53-accumulated cells underwent apoptosis without exhibiting the feature of erythroid differentiation. The mode of p21waf1 and Bax-alpha upregulations varied between p51A- and p53-expressing cells and between the types of DNA damage. These results suggest the possibility that p51A induces differentiation under genotoxic circumstances. There may be cellular factors that control p51A protein stability and transactivating ability. PMID- 10871868 TI - A conformational change in the Fc precludes the binding of two Fcgamma receptor molecules to one IgG. PMID- 10871869 TI - Interleukin 18: a pleiotropic participant in chronic inflammation. PMID- 10871870 TI - A homing selection hypothesis for T-cell trafficking. PMID- 10871871 TI - The human and mouse MHC class III region: a parade of 21 genes at the centromeric segment. AB - The human major histocompatibility complex (MHC) class III region contains 57-60 structural genes spanning 654-759 kb of genomic DNA. Analysis of the sequence identities of the human and mouse genomic regions between NOTCH4 and complement C2 yields important information on the locations of the coding and regulatory sequences. It also provides insights into the relationship between protein function and level of sequence conservation, and on the clustering of genes with related functions. PMID- 10871873 TI - Authors' correction. PMID- 10871872 TI - The mouse as a model for the effects of MHC genes on human disease. AB - As mice are often used to model human major histocompatibility complex (MHC) associated diseases, it is important to understand how their MHC regions differ at the DNA level. The sequencing of the mouse MHC (H2 region) has enabled a detailed map of this region to be assembled for comparison with the human MHC. Here, Richard Allcock and colleagues outline the similarities between the human and mouse MHC regions and discuss notable differences that might affect disease models. PMID- 10871874 TI - Co-stimulation and selection for T-cell help for germinal centres: the role of CD28 and OX40. AB - Given the importance of responding to infections with the right defensive strategy, much interest has focused on cytokine differentiation in CD4+ T cells. However, relatively little is known of the logistics of T-cell help for B cells. Here, Lucy Walker and colleagues propose key roles for CD28 and OX40 in coordinating the selection, expansion and migration of CD4+ T cells to B-cell follicles. PMID- 10871875 TI - IFN-alpha and IFN-beta: a link between immune memory and chronic inflammation. AB - The majority of expanded T cells generated during an immune response are cleared by apoptosis. Prevention of death in some activated T cells enables the persistence of a memory T-cell pool. Here, observations that IFN-alpha and IFN beta inhibit activated T-cell apoptosis are described. Although this enables memory T cells to persist without antigen, excessive IFN-alpha or IFN-gamma secretion might lead to chronic inflammation. PMID- 10871876 TI - Twins: mirrors of the immune system. AB - Twin studies are a powerful tool to assess genetic and nongenetic factors in multifactorial, immune-mediated diseases. Here, Marco Salvetti and colleagues review important results from such studies and highlight their potential value. Future developments that should help to realize the potential of twin studies are discussed. PMID- 10871877 TI - The causative H+/K+ ATPase antigen in the pathogenesis of autoimmune gastritis. AB - The gastric H+/K+ ATPase is the causative autoantigen recognized by CD4+ T cells that mediate autoimmune gastritis. Pathogenic CD4+ T cells are regulated by CD25+CD4+ T cells. Here, it is proposed that waves of activation and migration of antigen presenting cells and CD4+ T cells to and from the target organ and draining lymph node result in tissue damage. PMID- 10871878 TI - Serpins and other serine protease inhibitors. PMID- 10871879 TI - Calcium, a signaling molecule in the endoplasmic reticulum? AB - For many years now, it has been known that Ca2+ is an important signaling molecule in the cytosol of the cell, but emerging evidence suggests that Ca2+ might also play a signaling role in the endoplasmic reticulum. For example, agonist-induced fluctuations in free Ca2+ concentration in the endoplasmic reticulum can affect many functions of the endoplasmic reticulum, including protein synthesis and modification, and interchaperone interactions. PMID- 10871880 TI - The adaptor hypothesis revisited. AB - As originally postulated in Crick's Adaptor hypothesis, the faithful synthesis of proteins from messenger RNA is dependent on the presence of perfectly acylated tRNAs. The hypothesis also suggested that each aminoacyl-tRNA would be made by a unique enzyme. Recent data have now forced a revision of this latter point, with an increasingly diverse array of enzymes and pathways being implicated in aminoacyl-tRNA synthesis. These unexpected findings have far-reaching implications for our understanding of protein synthesis and its origins. PMID- 10871881 TI - L27, a novel heterodimerization domain in receptor targeting proteins Lin-2 and Lin-7. PMID- 10871882 TI - The internal structure of mitochondria. AB - Electron microscopic (EM) tomography is providing important new insights into the internal organization of mitochondria. The standard baffle model for cristae structure, called into question years ago, has now clearly been shown to be inaccurate. Depending on source and conformational state, cristae can vary from simple tubular structures to more complex lamellar structures merging with the inner boundary membrane through tubular structures 28 nm in diameter. The structural information provided by EM tomography has important implications for mitochondrial bioenergetics, biogenesis and the role of mitochondria in apoptosis. The structural paradigm defined by EM tomography is helping in the design of new experimental approaches to mitochondrial function. PMID- 10871883 TI - Turning genes off by Ssn6-Tup1: a conserved system of transcriptional repression in eukaryotes. AB - The Ssn6-Tup1 repressor forms one of the largest and most important gene regulatory circuits in budding yeast. This circuit, which appears conserved in flies, worms and mammals, exemplifies how a 'global' repressor (i.e. a repressor that regulates many genes in the cell) can be highly selective in the genes it represses. It also explains how, given the appropriate signal, specific subsets of these genes can be derepressed. Ssn6-Tup1 seems especially robust, bringing about a high level of repression irrespective of its precise placement on DNA or of specific features of the DNA control regions of its target genes. This high degree of repression probably results from several distinct mechanisms acting together. PMID- 10871884 TI - Understanding protein folding via free-energy surfaces from theory and experiment. AB - The ability of protein molecules to fold into their highly structured functional states is one of the most remarkable evolutionary achievements of biology. In recent years, our understanding of the way in which this complex self-assembly process takes place has increased dramatically. Much of the reason for this advance has been the development of energy surfaces (landscapes), which allow the folding reaction to be described and visualized in a meaningful manner. Analysis of these surfaces, derived from the constructive interplay between theory and experiment, has led to the development of a unified mechanism for folding and a recognition of the underlying factors that control the rates and products of the folding process. PMID- 10871885 TI - Automated annotation of GPI anchor sites: case study C. elegans. PMID- 10871886 TI - The introduction of Escherichia coli and biochemical genetics to the study of oxidative phosphorylation. PMID- 10871887 TI - Adiposity signals and brain reward mechanisms. PMID- 10871888 TI - Reconstructing one of nature's designs. PMID- 10871889 TI - How accurately can we image inositol lipids in living cells? PMID- 10871890 TI - The p75 receptor: first insights into the transduction mechanisms leading to either cell death or survival. PMID- 10871891 TI - NK1 (substance P) receptor antagonists--why are they not analgesic in humans? AB - Tachykinin NK1 receptor antagonists have failed to exhibit efficacy in clinical trials of a variety of clinical pain states. By contrast, in preclinical studies in animals NK1 receptor antagonists have been shown to attenuate nociceptive responses sensitized by inflammation or nerve damage, although they exhibit little effect on baseline nociception. Other agents with this profile of activity in animal tests, typically nonsteroidal anti-inflammatory drugs (NSAIDs), are analgesic in humans. Thus, NK1 receptor antagonists appear able to block behavioural responses to noxious and other stressful sensory stimuli at a level detectable in animal tests but fail to provide the level of sensory blockade required to produce clinical analgesia in humans. PMID- 10871892 TI - Ethical considerations in clinical pharmacogenomics research. AB - In recent years there have been unprecedented advances in our understanding of the involvement of genetic polymorphisms in the response to drug therapies. Polymorphisms have been identified that lead to variable patient responses to several medications including cardiovascular, psychiatric, anti-infective and analgesic therapies. The potential for the development of customized, genotype based therapies is scientifically and clinically attractive. However, these developments, although bearing scientific promise, raise ethical concerns for the conduct of research with human subjects, particularly with respect to confidentiality, risk-benefit analysis, DNA-banking and pharmacoeconomic issues. This article discusses some of the ethical considerations that are related to the use of pharmacogenomics in clinical research protocols. PMID- 10871893 TI - Glutamate receptors in glia: new cells, new inputs and new functions. AB - Functional glutamate receptors are expressed on the majority of glial cell types in the developing and mature brain. Although glutamate receptors on glia are activated by glutamate released from neurons, their physiological role remains largely unknown. Potential roles for these receptors in glia include regulation of proliferation and differentiation, and modulation of synaptic efficacy. Recent anatomical and functional evidence indicates that glutamate receptors on immature glia are activated through direct synaptic inputs. Therefore, glutamate and its receptors appear to be involved in a continuous crosstalk between neurons and glia during development and also in the mature brain. PMID- 10871894 TI - Potential new treatments for type 2 diabetes. AB - The heterogeneous pathogenesis and progressive natural history of type 2 diabetes mellitus (T2DM) contrive a formidable therapeutic challenge. Dual endocrine deficits of impaired insulin action (insulin resistance) and inadequate insulin secretion create an environment of chronic hyperglycaemia and general metabolic disarray. This inflicts a heavy burden of morbidity and premature mortality from cardiovascular diseases, microvascular disorders (e.g. retinopathy and nephropathy) and neuropathic conditions. Improving glycaemic control delays the onset and reduces the severity of these long-term complications. However, even with intensive use of current antidiabetic agents more than 50% of T2DM patients suffer poor glycaemic control and 18% develop serious complications within six years of diagnosis. Clearly, there is a need for new antidiabetic agents. PMID- 10871895 TI - Fluorescence techniques: shedding light on ligand-receptor interactions. AB - The ability of organisms, or individual cells, to react to external chemical signals, which are detected and transduced by cell-surface receptors, is crucial for their survival. These receptors are the targets of the majority of clinically used medicines. Combinatorial genetics can provide almost unlimited numbers of mutant receptor proteins and combinatorial chemistry can produce large libraries of potential therapeutic compounds that act on these membrane receptors. What is missing for the fundamental understanding of receptor function and for the discovery of new medicines are efficient procedures to screen both ligand receptor interactions and the subsequent functional consequences. Ultrasensitive fluorescence spectroscopic approaches, in combination with efficient labelling protocols, offer enormous possibilities for highly parallel functional bioanalytics at the micro- and nanometer level. PMID- 10871896 TI - Restenosis: a challenge for pharmacology. AB - The quest for an anti-restenotic drug continues to be a major challenge in the field of cardiovascular pharmacology because most therapies with proven efficacy in experimental neointima models have failed to limit restenosis. Some drug classes, including glycoprotein IIb/IIIa antagonists, nitric oxide donors and the antioxidant probucol, have recently demonstrated potential benefits in clinical trials. Progress in the development of local delivery systems for administration of drugs, antisense oligonucleotides or genes, in combination with an improved understanding of the pathogenesis of restenosis holds promise for ultimate pharmacotherapy of this condition. PMID- 10871897 TI - Molecular 'pharming' with plant P450s. PMID- 10871898 TI - Removing selectable marker genes: taking the shortcut. PMID- 10871899 TI - Molecular aspects of leaf senescence. AB - Senescence is the last stage of leaf development and one type of programmed cell death that occurs in plants. The relationships among senescence programs that are induced by a variety of factors have been addressed at a molecular level in recent studies. Furthermore, an overlap between the pathogen-response and senescence programs is beginning to be characterized. The complexity of the senescence program is also evident in studies of senescence-specific gene regulation and the role of photosynthesis and plant hormones in senescence regulation. New molecular-genetic approaches are expected to be useful in unraveling the molecular mechanisms of the leaf senescence program. PMID- 10871900 TI - Sugar transporters in higher plants--a diversity of roles and complex regulation. AB - Sugar-transport proteins play a crucial role in the cell-to-cell and long distance distribution of sugars throughout the plant. In the past decade, genes encoding sugar transporters (or carriers) have been identified, functionally expressed in heterologous systems, and studied with respect to their spatial and temporal expression. Higher plants possess two distinct families of sugar carriers: the disaccharide transporters that primarily catalyse sucrose transport and the monosaccharide transporters that mediate the transport of a variable range of monosaccharides. The tissue and cellular expression pattern of the respective genes indicates their specific and sometimes unique physiological tasks. Some play a purely nutritional role and supply sugars to cells for growth and development, whereas others are involved in generating osmotic gradients required to drive mass flow or movement. Intriguingly, some carriers might be involved in signalling. Various levels of control regulate these sugar transporters during plant development and when the normal environment is perturbed. This article focuses on members of the monosaccharide transporter and disaccharide transporter families, providing details about their structure, function and regulation. The tissue and cellular distribution of these sugar transporters suggests that they have interesting physiological roles. PMID- 10871901 TI - Engineering starch for increased quantity and quality. AB - The characterization and production of starch variants from mutation studies and transgene technology has been invaluable for our understanding of the synthesis of the starch granule. The knowledge gained has allowed for genetic manipulation of the starch biosynthetic pathway in plants. This in vivo approach can be used to generate novel starches and diminishes the need for post-harvest chemically and enzymatically treated starches. Thus, the modification of the starch biosynthetic pathway is a plausible means by which starches with novel properties and applications can be created. PMID- 10871902 TI - The Rrop GTPase switch turns on polar growth in pollen. AB - Pollen-tube growth not only represents an essential stage of plant reproduction but also provides an attractive model for studying cell polarity and morphogenesis. For many years, pollen-tube growth has been known to require a tip focused Ca2+ gradient and dynamic F actin, but the way that these are controlled remained a mystery until recently. Rop appears to be activated at growth sites by a tip-localized growth cue, acting as a central switch that controls the polar growth of pollen tubes, probably having its effect through phosphoinositides and Ca2+. These findings have begun to shed light on the molecular basis of pollen tube growth and cell morphogenesis in plants. PMID- 10871903 TI - Are microorganisms more effective than plants at competing for nitrogen? AB - Plant scientists have long debated whether plants or microorganisms are the superior competitor for nitrogen in terrestrial ecosystems. Microorganisms have traditionally been viewed as the victors but recent evidence that plants can take up organic nitrogen compounds intact and can successfully acquire N from organic patches in soil raises the question anew. We argue that the key determinants of 'success' in nitrogen competition are spatial differences in nitrogen availability and in root and microbial distributions, together with temporal differences in microbial and root turnover. Consequently, it is not possible to discuss plant-microorganism competition without taking into account this spatiotemporal context. PMID- 10871904 TI - Overcompensation of plants in response to herbivory and the by-product benefits of mutualism. AB - Plants that overcompensate for herbivory are relatively healthier when damaged. In this mutualistic association, the herbivore benefits from the plant, and the plant benefits from the herbivore's actions. As long as the benefit to the plant outweighs the costs imposed by browsing herbivores, this interaction should remain stable. Many apparently parasitic associations can be mutually beneficial under some environmental conditions. PMID- 10871905 TI - Interpretation of bone mineral density measurement and its change. AB - Bone mineral density (BMD), an important measurable predictor of osteoporotic fractures, is used as a surrogate definition of osteoporosis, and often as an end point in clinical trials. However, BMD is measured with random error and random fluctuation within individuals. This study addresses two specific questions: Given an observed level of BMD for an individual, what is the individual's likely "true" level? To what extent does an observed BMD change reflect a real change? A Bayesian model was formulated to address these questions, using data from the Dubbo Osteoporosis Epidemiology Study, past clinical trials, and a short-term reliability study in individuals ages 60 yr and older. Measurements of lumbar spine and femoral neck BMD by dual X-ray absorptiometry are highly reliable, with coefficients of reliability ranging from 0.90 to 0.99. Consequently, for an individual, there is good agreement between observed and "true" BMD values. However, the 90% confidence interval for the true level in elderly people and those with low measured BMD values is particularly wide. Using the cutoff of 2.5 standard deviations below the young normal mean as a definition of osteoporosis, the rates of false positives and false negatives can be as high as 20% among individuals ages 80 yr and older. In a typical clinical trial with an overall average increase in BMD of 2%, for a subject whose baseline femoral neck BMD is 0.80 g/cm(2), no conclusion of significant change could be drawn until an observed increase of at least 5.5% or an observed decrease of at least 7.5%. If two measurements were taken at baseline and follow-up, the true change could be detected with an observed increase of 3.5% or an observed decrease of 5%. On the other hand, there needs to be an observed increase of 6.2 and 8.5% before one can be 90% certain that a true increase of 2 and 5%, respectively, has occurred in an individual. This analysis suggests that the diagnosis of osteoporosis based on a single measurement of BMD and point estimates of changes in BMD may be inappropriate and unreliable. We propose that the current practice of informing individuals who have BMD measurements about their actual T- and Z-scores be replaced with a system of reporting in which their osteoporosis probability risk category is conveyed. Also, assessment of change in an individual should take into account the overall change in a population. PMID- 10871906 TI - Evaluation of error bounds on calcaneal speed of sound caused by surrounding soft tissue. AB - For absorptiometry measurements, soft tissue may have an impact on quantitative ultrasound (QUS) measurements. In the present study, we focused primarily on the quantification of measurement error on speed of sound (SOS) caused by surrounding soft tissue. The relevant soft tissue parameters affecting the inherent SOS inaccuracies are thickness and sound velocity. To meet our goal, SOS measurements were taken at the right heel using a QUS imaging device in 21 healthy subjects. Site-matched measurements of soft tissue thickness (STT) and bone width were performed using magnetic resonance imaging of the heel. Several bone velocities were calculated either by accounting for bone width (SOSBW) only or by taking into account the exact path lengths of all major components traversed by ultrasound &lapr;V(b)). Given that soft tissue composition is difficult to determine in vivo, we chose to estimate lower and upper error bounds on bone velocity (V(b lower) and V(b upper)) by spanning the full range of available values in the literature. The mean BW was 30.7 +/- 2.7 mm and the mean medial and external STTs were 8.8 +/- 1.7 and 8.5 +/- 1.5 mm, respectively. Accounting for true BW only resulted in no significant difference between SOS (1533 +/- 37) and SOSBW (1531 +/- 33). By contrast, accounting for both true BW and surrounding soft tissue resulted in an increase in the calculated bone velocity and statistically significant differences between SOS and V(b upper) (1568 +/- 36) and V(b lower) (1542 +/- 34). Root mean square errors between SOS and the calculated velocities were 0.34, 2. 32, and 0.70% for SOSBW, V(b upper), and V(b lower), respectively. We report here measurement errors caused by soft tissue to be 3 to 20 times higher than the SOS short-term precision (SOS coefficient of variation of 0.1%). Our results suggest that inaccuracies in SOS measurement caused by overlying soft tissue cannot be neglected. Overlying soft tissues may influence outcomes of longitudinal studies, especially if variations in tissue thickness and composition occur during the longitudinal follow-up. A practical way of minimizing the measurement error could be to perform an adequate correction for the overlying soft tissue. However, ideally, this should require knowing both the thickness and sound velocity in soft tissue. One might preferably conduct experimental investigations that directly control soft tissue thickness and composition to resolve this problem. PMID- 10871907 TI - Bilateral measurement of femoral bone mineral density. AB - Both femora were measured on 61 normal adults using dual X-ray absorptiometry (DXA). In a subset of 31 subjects, each femur was scanned once using the conventional leg-positioning device supplied with the densitometer, and once using a new positioning device and software that allowed both legs to be measured simultaneously. In another subgroup (n = 30), subjects were measured three times using the new dual-femur approach to better assess precision error. The data were analyzed for differences owing to the different positioning devices and for differences between right and left sides. The correlation between results with the old and new positioners was high (r > 0.99, standard error of the estimate [SEE] = 0.01-0.02 g/cm(2)). There was no significant difference in the average bone mineral density (BMD) values between the old and new positioner. The precision errors for each femur alone with the dual-femur approach were similar to those reported for the single-femur scans (1 to 2%), but the precision errors for the combined femora were reduced by 30% as expected. The correlation between right and left sides was high (r = 0.94-0.96), and the SEE in predicting one side from the other was moderate for total, trochanteric, and femoral neck BMD (0.05, 0. 05, and 0.06 g/cm(2), respectively). These SEE equate to about 0.5 standard deviation in terms of T-score. Differences in many individual cases between the right and left sides were significantly greater than the precision error. The new dual-femur software and leg positioner allows rapid measurement and analysis of both femora, thereby eliminating the uncertainty between sides. PMID- 10871908 TI - Quantitative ultrasound: an indicator of osteoporosis in perimenopausal women. AB - The key to effective treatment of osteoporosis is early detection; however, the disease in perimenopausal women is frequently undiagnosed. To assess the utility of quantitative ultrasound (QUS) at the calcaneus in perimenopausal women, broadband ultrasound attenuation (BUA); speed of sound (SOS); quantitative ultra sound index (QUI), an algorithm of BUA and SOS; and bone mineral density by dual X-ray absorptiometry (DXA) of the posteroanteiror spine, femoral neck, and total hip were measured in 420 women (ages 45-55 yr). Thirty (7.1%) of the women were found to be osteoporotic by DXA. All QUS measurements were predictors of osteoporosis. QUS values did not differ between postmenopausal women on estrogen replacement therapy (ERT) and those not on ERT. There were no differences among BUA, SOS, and QUI in the area under the receiver operating characteristic curves for predicting osteoporotic vs nonosteoporotic cases. At a QUI of 89, ultrasound had an 80% sensitivity for the diagnosis of osteoporosis, but only a 74% specificity. The use of QUS in perimenopausal women will facilitate the identification of women with osteoporosis. However, the high false-positive rate (26%) limits the utility of QUS as the sole diagnostic technique on which to base therapeutic decisions. Nevertheless, low QUS measurements may provide a means for targeting those women who would benefit most from more extensive evaluation (e. g., DXA). PMID- 10871909 TI - Monitoring skeletal response to treatment which site to measure in the femur? AB - In the past it was usual to interpret bone mineral density (BMD) scans of the femur using the femoral neck, trochanter, or Ward's triangle sites. Recently, a study by the International Committee for Standards in Bone Measurement recommended that the total hip should be the preferred site for the interpretation of femur BMD, and another study described a new central hip site that may offer improved precision. This article compares the longitudinal sensitivities of the different femur BMD sites for monitoring patient response to treatment. The study population was 152 postmenopausal women enrolled in a trial of a bisphosphonate therapy. Spine and hip BMD scans were performed at 0, 1, and 2 yr. The mean percentage change at 2 yr was calculated for six sites in the hip, and the spine was also included for comparison. Treatment effect was defined as the difference in the BMD change between the treated and placebo groups. Although the data analysis incorporated a term for a calibration change caused by a repair of the dual X-ray absorptiometry scanner, the effect of this event on the estimation of treatment effect was negligible. Longitudinal sensitivity was derived by dividing the treatment effect by the root mean square error (RMSE) of the statistical model. Results (and standard errors) normalized to the ratio of treatment effect: RMSE for femoral neck BMD were as follows: femoral neck: 1.00; trochanter: 1.33 (0.38); intertrochanteric: 0.84 (0.41); total hip: 1.20 (0.38); Ward's triangle: 1.03 (0.27); central hip: 1.09 (0.30); spine: 2.08 (0. 45). At none of the femur sites was the change in BMD large enough to allow monitoring of response to treatment in individual patients. However, for studies involving the follow-up of a group of subjects, the longitudinal sensitivities of the different femur sites were equal within the statistical errors of the study. In particular, total hip BMD appears to be as effective as femoral neck BMD for detecting response to treatment in the femur in the setting of a clinical trial or similar research study. PMID- 10871910 TI - Differences between dual X-ray absorptiometry using pencil beam and fan beam modes and their determinants in vivo and in vitro. AB - The aim of this study was to determine the influence of individual factors on differences in bone mineral density (BMD) using dual X-ray absorptiometry pencil beam (PB) and fan beam (FB) modes in vivo and in vitro. PB.BMD and FB.BMD of 63 normal Caucasian females ages 21-80 yr were measured at the lumbar spine and hip. Residuals of the FB/PB regression were used to assess the impact of height, weight, adiposity index (AI) (= weight/height(3/2)), back tissue thickness, and PB.BMD, respectively, on FB/PB difference. The Hologic Anthropomorphic Spine Phantom (ASP) was measured using the PB and FB modes at two different levels to assess the impact of scanning mode and focus distance. The European Spine Phantom (ESP) prototype, a geometrically well-defined phantom with known vertebral densities, was measured using PB and FB modes and analyzed manually to determine the impact of bone density on FB/PB difference and automatically to determine the impact of edge detection on FB/PB difference. Population BMD results were perfectly correlated, but significantly overestimated by 1.5% at the lumbar spine and underestimated by 0.7% at the neck, 1.8% at the trochanter, and 2.0% at the total hip, respectively, when using the FB compared with PB mode. At the lumbar spine, the FB/PB residual correlated negatively with height (r = 0.34, p < 0.01) and PB.BMD (r = 0.48, p <: 0. 0001) and positively with AI (r = 0.26, p < 0.05). At the hip, residual of trochanter correlated positively with weight (r = 0.36, p < 0.01) and AI (r = 0.36, p < 0.01). The FB mode significantly increased ASP BMD by 0.7% compared with PB. Using the FB mode, increasing focus distance significantly (p < 0.001) decreased area and bone mineral content, but not BMD. By contrast, increasing focus distance significantly decreased PB.BMD by 0.7%. With the ESP, the PB mode supplied accurate projected are of the bone (AREA) results but significant underestimation of specified BMD in the manual analysis. The FB mode significantly underestimated PB. AREA by 2.9% but fitted specified BMD quite well. FB/PB overestimation was larger for the low-density (+8.7%) than for the high-density vertebra (+4. 9%). The automated analysis resulted in more than 14% underestimation of PB. AREA (low-density vertebra) and an almost 13% overestimation of PB.BMD (high-density vertebra) using FB. In conclusion, FB and PB measurements are highly correlated at the lumbar spine and hip with small but significant BMD differences related to height, adiposity, and BMD. In clinical practice, it can be erroneous to switch from one method to another, especially in women with low bone density. PMID- 10871911 TI - Bone mineral density in diagnosis of osteoporosis: reference population, definition of peak bone mass, and measured site determine prevalence. AB - A population-based study was performed in order to compare different definitions of peak bone mass, and to apply the corresponding T-scores for different skeletal sites to a cohort of 70-yr-old women for studying the prevalence of osteoporosis. Bone mineral density (BMD) of the hip, lumbar spine, and forearm was measured by dual X-ray absorptiometry (Hologic 4500) in 296 women ages 16-31 yr and 210 women age 70 yr. Peak bone mass occurred in women in their early 20s at the proximal femur and at 28 and 31 yr at the spine and forearm, respectively. BMD cutoff levels were compared to machine-specific cutoff values for the different sites. When applied to our cohort of 70-yr-old women, the prevalence of osteoporosis at the total hip was 9-25%, depending on which peak bone mass the T-score of -2.5 was based. The prevalence in the spine was 28-33% and in the forearm 45-67%. Osteoporosis in at least one of the three measured sites was documented in 49-72% of the population sample. Our results show that the use of T-score to define osteoporosis results in a highly different prevalence rate in a given population depending on the reference population and the skeletal sites chosen for measurement. PMID- 10871912 TI - Prevention and treatment of osteoporosis: efficacy of combination of hormone replacement therapy with other antiresorptive agents. AB - Osteoporosis is a debilitating disease characterized by decreased bone mineral density (BMD) leading to fractures. It primarily affects postmenopausal women and elderly men. Prevention of osteoporosis is very important because present therapies do not have the potential to mend damage to the bone microarchitecture caused by osteoporosis. The first line of prevention and treatment of osteoporosis is hormone replacement therapy (HRT). All of the approved drugs for the prevention and treatment of osteoporosis act as inhibitors of bone resorption; these drugs include HRT, selective estrogen receptor modulators, calcitonin, and bisphosphonates. The latter two drugs have also been shown to prevent fractures. This article discusses data from nine controlled prospective clinical studies. Study 1 was designed to assess the efficacy of combined HRT and bisphosphonate in preventing osteoporosis during the early stages of menopause. This combined therapy increased the lumbar spine BMD by 10.9% and femoral BMD by 7.3% over 4 yr, compared with 6.8 and 4.0% with HRT alone, and 6.8 and 1.2% with bisphosphonate alone. Study 2 was conducted on postmenopausal women with established osteoporosis. These results showed a 10.4 and 7.0% increase in BMD in vertebrae and femora, respectively, compared with 7.3 and 4.8% increases in the HRT group, and 6.8 and 0.9% in the bisphosphonate group. Data from study 3 demonstrated similar findings in that the combination of alendronate and HRT also enhanced BMD values. Studies 4 and 5 assessed the efficacy of the combined therapy of HRT and calcitonin in the prevention of early postmenopausal bone loss. Both studies demonstrated a significant increase in BMD over and above that observed with either HRT or calcitonin alone. Studies 6, 7, and 8 demonstrated that the addition of testosterone to estrogen therapy further increased BMD when compared to estrogen therapy alone, and also prevented the expected decreases in markers of bone formation in early postmenopausal women. Study 9 demonstrated a synergistic effect on BMD in postmenopausal women, when HRT was coadministered with monofluorophosphate. Other combination therapies may also enhance BMD (e.g., the combination of alendronate and parathyroid hormone [PTH]). However, some agents either lose their efficacy or have no added effects on BMD when they are coadministered (e.g., tiludronate and PTH, calcitonin and PTH, calcitonin and anabolic steroids). These studies illustrate that in a subgroup of patients (i.e., patients with high bone turnover and/or severe osteoporosis), specific combination treatments such as HRT with bis-phosphonates, calcitonin, or androgens (and perhaps also with PTH, fluoride, nitric oxide donors) provide additional beneficial effects over a single-drug therapy. Whether these combination therapies are more effective than individual drugs in reducing fractures still needs to be determined. PMID- 10871913 TI - Osteolysis of the pelvis presenting as insufficiency fracture in a patient with rheumatoid arthritis. AB - Physician awareness of the risk of osteoporosis and subsequent fractures in a patient with a history of long-term steroid treatment is high. The tendency to assume that a fracture is owing to steroid-induced osteoporosis may result in an unnecessarily intense antiresorptive treatment regimen for a patient who may not have osteoporosis. I report here about a patient with rheumatoid arthritis who presented with bone fracture despite antiresorptive therapy and without evidence of osteoporosis by bone mineral density testing. PMID- 10871914 TI - Development of a live varicella vaccine--past and future. AB - Background of the development of a live varicella vaccine, including studies on the attenuation of measles and polioviruses, and transformation experiments of cultured hamster and human cells with conditional lethal mutants of adenovirus and herpes simplex virus were described. Varicella-zoster virus (Oka strain) was passaged in guinea pig cells, and the resulting virus (vaccine virus) was found to have a higher affinity to guinea pig cells. It was recently proved that variations of base sequence occurred exclusively in gene 62 (immediate-early gene) in comparison of vaccine Oka virus and parent Oka virus. This variation is presumed to have occurred during passage in guinea pig cells. Live varicella vaccine (Oka strain) has increasingly been used throughout the world. It was also found in a preliminary study that giving the vaccine to the elderly enhanced humoral and cell-mediated immunity, leading to a prevention of post herpetic neuralgia. A large field trial is now going on in the United States to immunize the elderly for the purpose of prevention of herpes zoster, particularly post herpetic neuralgia. PMID- 10871915 TI - Japanese scrapie cases. AB - Worldwide attention has been given to scrapie, because bovine spongiform encephalopathy (BSE) could be experimentally transmitted to sheep. This ovine form of BSE was clinically identical to scrapie. In Japanese scrapie cases, a majority of the diseased sheep were from Suffolk, while 8 cases were from Corriedale. It is very likely that sheep-to-sheep transmission of scrapie has taken place in Obihiro, Hokkaido. Normal prion protein may play a role in the morphoregulatory signaling pathway, which orchestrates the specificity of a particular cellular response. Over-expression of normal prion protein in mice cause neurodegenerative disorders. Recently, Prnd was identified downstream of the mouse prion protein gene (Prnp), and encodes 179 amino acids and a prion protein (PrP)-like protein designated doppel (Dpl). Dpl was upregulated in the central nervous system of two PrP-deficient lines of mice, as well as in prionless cell lines. Dpl caused neurodegeneration similar to that caused by PrP. Linked expression of Prnp and Prnd may cause several neurodegenerative disorders. PMID- 10871916 TI - Effect of sulfated colominic acid on enteric virus (rotavirus, poliovirus and coxsackievirus) infections in vitro. AB - Sulfated colominic acid exhibited suppressive effects on SA11 (simian rotavirus)- and MO (human rotavirus)-infections, but not on Wa (human rotavirus)-, Sabin 1 (poliovirus 1)-, and Nancy (coxsackie B3 virus)-infections, in vitro. The infection of SA11 was found to be inhibited by mixed treatment and early posttreatment with sulfated colominic acid, but not by pretreatment, by plaque assay and multiple growth assay. The results were confirmed by the infectivity titer, RNA polyacrylamide gel electrophoresis, and electron microscopic analysis. The mechanism of the suppressive effect was suggested to be adsorption inhibition at an early stage of the infection. PMID- 10871917 TI - Viral load in Indonesian patients with chronic liver disease and in blood donors infected with different subtypes of hepatitis C virus. AB - The viral load of different hepatitis C virus (HCV) subtypes, including the globally distributed HCV-1b and the unique Indonesian subtype HCV-1c, was analyzed using serum samples obtained from Indonesian blood donors and patients with chronic liver disease. The mean viral load of HCV-1c was comparable with that of HCV-1b, suggesting that HCV-1c is as pathogenic as HCV-1b. On the other hand, the mean viral load of HCV-2a was lower than that of HCV-1b or HCV-1c, with this result being consistent with previous observations. Interestingly, some HCV 2a strains were associated with a high viral load that was almost equivalent to that of HCV-1b and HCV-1c. This result implies the possibility that there exists a minor fraction of HCV-2a strains that cause higher levels of viremia compared with the majority of ordinary HCV-2a strains. PMID- 10871918 TI - Simultaneous detection of hepatitis B, C, and G viral genomes by multiplex PCR method. AB - We established a multiplex polymerase chain reaction (PCR) method for simultaneous detection of hepatitis B, C, and G viral genomes. The levels of concordance with the data obtained by conventional single PCR method were 100% for single infection, 98 to 100% for double infections, and 92% for triple infections. This method is not only suited to rapid, large-scale epidemiological screening and clinical diagnosis of those virus infections occurring alone or in combination, but is also time- and cost-effective. PMID- 10871919 TI - Japanese spotted fever in Shimane Prefecture - outbreak and place of infection. PMID- 10871920 TI - Sporadic cases of Yersinia pseudotuberculosis serotype 5 infection in Shodo Island, Kagawa Prefecture. PMID- 10871921 TI - The use of colony hybridization in the isolation of thermostable direct hemolysin producing Vibrio parahaemolyticus from foods implicated in an incidence of food poisoning. PMID- 10871922 TI - Occurrence of scrub typhus (Tsutsugamushi) in Kanagawa Prefecture and types of Orientia tsutsugamushi involved. PMID- 10871923 TI - An electronic system combining MIC2000(TM) and antibiogram cluster analysis for surveillance of methicillin-resistant Staphylococcus aureus in hospitals. PMID- 10871924 TI - Fatal rapidly progressing streptococcal toxic shock-like syndrome; case report. PMID- 10871925 TI - Epidemiological analysis of methicillin-resistant Staphylococcus aureus outbreaks in a neonatal intensive care unit by genomic DNA fingerprinting using pulsed field gel electrophoresis. PMID- 10871926 TI - Epidemiological analysis of a methicillin-resistant Staphylococcus aureus outbreak in a surgery ward by genomic DNA fingerprinting using pulsed-field gel electrophoresis. PMID- 10871927 TI - Comparison of genomic DNA fingerprinting using pulsed-field gel electrophoresis and antibiotic susceptibility of clinical isolates of methicillin-resistant Staphylococcus aureus between Chiang Mai and Tokyo. PMID- 10871928 TI - Fv-1 restriction of murine leukemia virus may not necessarily be at cytoplasmic nuclear transport phase. PMID- 10871929 TI - Surveillance of poliovirus-isolates in Japan, 1999. PMID- 10871930 TI - Regression of deeply infiltrating giant condyloma (Buschke-Lowenstein tumor) following long-term intralesional interferon alfa therapy. PMID- 10871931 TI - Previously undiagnosed sarcoidosis in a patient presenting with leonine facies and complete heart block. PMID- 10871932 TI - Clinical significance of skin biopsies in the diagnosis and management of graft vs-host disease in early postallogeneic bone marrow transplantation. AB - OBJECTIVE: To determine the value of skin biopsies in the management of suspected graft-vs-host disease (GVHD) within 30 days of allogeneic bone marrow transplantation (BMT). DESIGN: Retrospective study based on review of a BMT database. SETTING: Leukemia/BMT ward of a tertiary care, university teaching hospital. PATIENTS: One hundred and eighty-seven consecutive patients who received allogeneic BMT between January 1, 1994, and June 30, 1997, at Vancouver General Hospital, Vancouver, British Columbia. MAIN OUTCOME MEASURES: (1) Skin biopsy frequency for patients with rashes suggestive of acute GVHD; (2) clinical significance of skin biopsy in the management of patients with suspected acute GVHD after BMT; (3) relationship between severity of clinical GVHD and the likelihood to receive GVHD therapy; and (4) relationship between biopsy status or biopsy result and outcome of BMT (acute and chronic GVHD, transplant-related mortality, and overall and event-free survival). RESULTS: During the early post BMT period (<30 days after BMT), 88 patients had rashes suggestive of acute GVHD; of these, 51 (58%) underwent skin biopsy to confirm the diagnosis. Skin biopsies were performed more often for higher clinical stages of cutaneous GVHD. There was no significant difference between the patients with positive biopsy findings and those with negative findings, either in the clinical severity of acute GVHD or in likelihood to receive treatment for GVHD. Most (85%) of the patients who underwent biopsies and received GVHD therapy had treatment initiated before skin biopsies were performed or before the results were available. The higher the clinical grade of overall acute GVHD, the more likely it was that the patients were treated for GVHD (P<.001). The outcome of BMT was not influenced by the skin biopsy status or biopsy result. CONCLUSIONS: The biopsy findings correlated poorly with the clinical severity of skin rash suggestive of acute GVHD soon after BMT. The decision to treat suspected acute GVHD depended not on skin biopsy findings but rather on clinical severity of acute GVHD. In this regard, skin biopsy has a limited role in the management of patients early after allogeneic BMT. PMID- 10871933 TI - Treatment of perioral rhytides: a comparison of dermabrasion and superpulsed carbon dioxide laser. AB - OBJECTIVE: To directly compare the cosmetic outcome and adverse effects of dermabrasion and superpulsed carbon dioxide laser for the treatment of perioral rhytides. DESIGN: Subjects were randomly assigned to receive treatment with carbon dioxide laser resurfacing to one side of the perioral area and dermabrasion to the other side in a prospective, comparative clinical study. The duration of follow-up by blinded observers was 4 months. SETTING: University hospital-based dermatologic surgery clinic. PATIENTS: Fifteen healthy fair skinned volunteers with moderate to severe perioral rhytides and no history of prior cosmetic surgical procedures to the same anatomic area. INTERVENTIONS: One half of the perioral area was treated with the LX-20SP Novapulse carbon dioxide laser (Luxar Corp, Bothell, Wash), and the other half was treated with dermabrasion using either a hand engine-driven diamond fraise or a medium-grade drywall sanding screen (3M Corp, St Paul, Minn). MAIN OUTCOME MEASURES: Improvement in rhytides, patients' subjective reports of postoperative pain, time to reepithelialization, degree of postoperative crusting, and duration of postoperative erythema were observed for both methods. Standardized scoring systems were used to quantify outcome measures. Paired t tests were used for statistical comparisons of the 2 resurfacing methods. RESULTS: The difference in rhytide scores for the 2 methods was not statistically significant (P= .35) at 4 months. Less postoperative crusting and more rapid reepithelialization were noted with the dermabrasion-treated skin. Postoperative erythema was of longer duration on laser-treated skin. Patients reported less pain with dermabrasion treatment. Subtle differences that were difficult to quantify were also noted between the methods. CONCLUSIONS: Both dermabrasion and carbon dioxide laser resurfacing are effective in the treatment of perioral rhytides. Both methods have unique advantages and disadvantages. PMID- 10871934 TI - Value of capillary microscopy in the diagnosis of hereditary hemorrhagic telangiectasia. AB - BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is a hereditary disorder, leading to easily bleeding telangiectases on the skin and mucosal surfaces. The disease is associated with arteriovenous malformations in multiple organs. Potentially serious complications warrant an early diagnosis. Telangiectases are the hallmark of the disease, but may be difficult to distinguish. OBJECTIVES: To evaluate the value of capillary microscopy in the diagnosis of HHT and to compare the capillary pattern of the fingernail folds in patients with HHT and healthy persons. SETTING: Outpatient department of a general hospital. PARTICIPANTS: A random sample of 54 patients with HHT and a volunteer sample of 40 healthy persons. MAIN OUTCOME MEASURE: The difference in the capillary pattern between patients with HHT and healthy volunteers. RESULTS: Forty-five (83%) of 54 patients with HHT had giant loops between the normal capillaries in the nail fold. Two patients had only enlargement of the draining limb of the capillary. Seven patients (13%) had no vascular abnormalities in the nail fold. Seven of 9 patients with HHT but without cutaneous telangiectases had microvascular abnormalities. None of the volunteers had vascular abnormalities. The difference between both groups was significant (chi2, P<.001). CONCLUSION: Capillary microscopy can be a valuable tool in diagnosing HHT. PMID- 10871935 TI - Methicillin-resistant Staphylococcus aureus. Nosocomial acquisition and carrier state in a wound care center. AB - OBJECTIVE: To assess methicillin-resistant Staphylococcus aureus (MRSA) nosocomial acquisition and carrier state in a wound care center. DESIGN AND SETTING: The results of an intervention to control MRSA were compared with those of historical controls at the wound care center of university-based Hopital Broussais, Paris, France. PATIENTS: Patients admitted for specific care of chronic ulcers and surgical wounds. MAIN OUTCOME MEASURES: Incidence rates of MRSA carriers and acquisition in wounds. RESULTS: Of 88 patients admitted during a 3-month preintervention period in 1993, 18 (21%) were MRSA carriers. Of 334 patients admitted in 1994 and 395 in 1996, 65 (19.5%) and 81 (20.5%) were MRSA carriers, respectively (P=.80). In 1993, 6 (9%) of 70 patients without MRSA acquired MRSA wound infections; the corresponding numbers were 6 (2.2%) of 269 in 1994 and 3 (0.9%) of 314 in 1996. Despite that the number of MRSA carriers remained stable at admission to the wound care center, the rate of MRSA infections in wounds per 100 noncarriers decreased significantly between the preintervention period and subsequent years: 1994 (P=.02) and 1996 (P=.002). CONCLUSIONS: Although our results are limited by the use of historical controls, they showed that simple infection control measures, such as the use of soap and water and barrier precautions associated with staff education, seemed to significantly reduce MRSA infection rates in patients with chronic skin breaks. PMID- 10871936 TI - News and Notes: June 2000. PMID- 10871937 TI - Columnar epidermal necrosis: a unique manifestation of transfusion-associated cutaneous graft-vs-host disease. AB - BACKGROUND: In 1978, the first case of columnar epidermal necrosis was reported in a 6-year-old boy. There were scaly, partially vesicular or crusty, erythematous lesions mainly involving the extremities that histopathologically showed peculiar features of focal, total epidermal necrosis accompanied by a lichenoid tissue reaction. He developed the skin eruption after receiving a blood transfusion from his mother when he showed debility induced by vaccination with an alternated live measles virus vaccine. The lesions rapidly regressed after sun exposure. To our knowledge, there has been no report of a similar case despite such unique features. OBSERVATION: We encountered a similar case of columnar epidermal necrosis in a 15-year-old Japanese girl with chronic graft-vs-host disease; the lesions occurred 3 months after the transfusion of peripheral blood stem cells from her HLA antigen-matched brother. However, there was no exacerbation of liver dysfunction, diarrhea, or bone marrow aplasia. The peculiar cutaneous lesions responded well to topical phototherapy. CONCLUSION: These 2 patients shared a similarity in their lesions and circumstances under which the blood transfusion was performed to a debilitated patient from a close family member. We believe that focal epidermal necrosis observed in patients with this condition represents a variant of blood transfusion-associated lichenoid graft-vs host disease that occurs uniquely in a skin-targeted fashion. PMID- 10871938 TI - Narrowband TL-01 phototherapy for patch-stage mycosis fungoides. AB - BACKGROUND: Although patch-stage mycosis fungoides (MF) has a generally good prognosis, and long-term survival rates with current therapies (UV-B, photochemotherapy, topical nitrogen mustards, electron-beam therapy) are similar, there is concern regarding their potential adverse effects. Narrowband or TL-01 UV-B phototherapy (311 nm), in use for more than 10 years, is more effective than broadband UV-B for the treatment of psoriasis, with an efficacy approaching that of psoralen UV-A. This open study assesses TL-01 as an alternative therapy for patch-stage MF. OBSERVATIONS: Eight white patients (4 men, 4 women; age range, 66 83 years) with histologically proven patch-stage MF received TL-01 phototherapy 3 times weekly using a standard protocol. Complete clearance of MF was achieved in 6 cases in a mean of 9 weeks or 26 treatments (range, 20-37 weeks) and 4 patients have had prolonged remissions. Mean duration of clinical improvement has been 20 months (range, 11-40 months). Partial response to TL-01 or poor histologic improvement was associated with rapid relapse. CONCLUSIONS: TL-01 is an effective, convenient therapy that may have less risk of long-term adverse effects than current alternatives. Although larger prospective studies are necessary, for some patients intermittent courses of TL-01 may offer effective long-term therapy. PMID- 10871939 TI - Necrolytic acral erythema associated with hepatitis C: effective treatment with interferon alfa and zinc. AB - BACKGROUND: Necrolytic acral erythema is a recently described necrolytic erythema that is unique in its exclusive acral location and strong association with hepatitis C. OBSERVATION: We report the first case of necrolytic acral erythema in the United States. The patient is a 43-year-old black woman who presented with a 4-year history of tender, flaccid blisters localized to the dorsal aspect of her feet. Serum zinc and glucagon levels were normal. Serum antibodies were positive for hepatitis C, and a liver biopsy specimen showed chronic hepatitis. She was successfully treated with interferon alfa-2b and zinc. We review all previously reported cases. CONCLUSIONS: Necrolytic acral erythema is a distinct entity. In a review of the literature, most patients were between 35 and 55 years of age, although 1 patient was 12 years old. Five of 8 patients were female. Four of 7 patients described previously were treated with variable success using oral zinc sulfate and amino acids, whereas 2 were successfully treated with interferon alfa. All patients were infected with hepatitis C. Necrolytic acral erythema appears to be a skin disorder linked to infection with hepatitis C virus that responds to treatment with interferon alfa and oral zinc. PMID- 10871940 TI - The prevalence of seborrheic keratoses in people aged 15 to 30 years: is the term senile keratosis redundant? AB - BACKGROUND: Seborrheic keratoses (SKs) are common skin lesions that have been shown to occur with increasing age, although the age of onset is not well recorded. OBJECTIVE: To determine the prevalence, nature, and distribution of SKs in young people. METHODS: One hundred seventy people aged 15 to 30 years were given a total body examination during which the presence, number, site, and size of SKs were recorded. Biopsy specimens were taken from the first 22 people who had lesions clinically diagnosed as SKs. Data on age, skin type, eye color, and hair color were recorded for all respondents. RESULTS: Forty (23.5%) of 170 respondents had at least one SK, with no significant difference between the sexes. There was an increase in prevalence with age from 15.7% in 15- to 19-year olds to 32.3% in those aged 25 to 30 years. The size of the SKs also increased with age. A total of 77.5% of SKs were found on the trunk and 22.5% on the limbs, head, and neck. There was no correlation between SKs and any particular hair and eye color or skin type. CONCLUSIONS: These findings confirm that SKs are common lesions in young Australians, appearing in a substantial proportion of people younger than 30 years. The term senile keratosis is no longer appropriate for these lesions. PMID- 10871941 TI - Outcome measures of disease severity in atopic eczema. AB - BACKGROUND: An essential component of evidence-based medicine is the use of valid and reliable outcome measures in clinical trials. There is much confusion in the field of atopic eczema regarding how to best measure disease severity objectively. OBJECTIVE: To establish the extent to which existing objective clinical scales for atopic eczema have been tested for validity, reliability, sensitivity to change, and acceptability. DESIGN: An electronic bibliographic search was performed for published data on all currently available named atopic eczema scales. RESULTS: Thirteen scales were identified in total. Data on construct or criterion validity were available for 10 scales. Only 5 scales had been tested for reliability (interobserver variability, intraobserver variability, or internal consistency). Data on responsiveness to change were available for 8 scales. An estimated time to administer the measure had been given for 3 scales. The only severity scale for which published data could be found on validity, reliability, sensitivity, and acceptability testing was the Severity Scoring of Atopic Dermatitis index, although problems occurred with interobserver variation of the index. CONCLUSION: The rapidly increasing number of severity scales for atopic eczema, many of which have been inadequately tested, has made the interpretation of patient outcomes confusing, and comparison of results between studies almost impossible. Consensus among clinicians and researchers on the use of severity scales for atopic eczema should be based on evidence of adequate validity), reliability, sensitivity to change, and ease of use. PMID- 10871943 TI - Advantage of Smaller Trials for the Widespread Use of Sedative Drugs. PMID- 10871942 TI - Odds ratios and relative risks. PMID- 10871944 TI - The Relationship Between Melanoma and Continuous or Intermittent Exposure to UV Radiation. PMID- 10871945 TI - Biologic Treatment of Basal Cell Carcinoma. PMID- 10871946 TI - A Remarkable Result of a Double-Masked, Placebo-Controlled Trial of Erythromycin in the Treatment of Pityriasis Rosea. PMID- 10871947 TI - Why a skin biopsy? PMID- 10871948 TI - Resurfacing procedures: how do you choose? PMID- 10871949 TI - Controlling nosocomial infection: everyone is concerned. PMID- 10871951 TI - Tender nodules on the legs of a cardiac transplant recipient. PMID- 10871950 TI - Giving "scale" new meaning in dermatology: measurement matters. PMID- 10871952 TI - Multiple light-yellow papules. PMID- 10871953 TI - Off-Center Fold: Symmetrical Black Plaques on the Toes. PMID- 10871954 TI - Off-Center Fold: Diffuse and Progressive Papules and Nodules. PMID- 10871955 TI - The appropriateness of curettage and electrodesiccation for the treatment of basal cell carcinomas. PMID- 10871956 TI - Accuracy of diagnosis of seborrheic keratoses in a dermatology clinic. PMID- 10871957 TI - Preliminary evidence for an association of measles virus with recurrent aphthous ulceration. PMID- 10871958 TI - A cycle: recurrent gram-negative folliculitis with Citrobacter diversus (koseri) following eradication of recurrent staphylococcal pyoderma. PMID- 10871959 TI - Candida parapsilosis chrondritis successfully treated with oral fluconazole. PMID- 10871960 TI - Nutria itch. PMID- 10871961 TI - Genetic mosaicism in an acquired inflammatory dermatosis following the lines of Blaschko. PMID- 10871962 TI - Double-blind, right/left comparison of calcipotriol ointment and betamethasone ointment in the treatment of Prurigo nodularis. PMID- 10871963 TI - Comparison of urinary 8-hydroxy-2'-deoxyguanosine in patients treated with topical corticosteroids, UV-B, and psoralen UV-A therapies. PMID- 10871964 TI - Does depression kill? PMID- 10871965 TI - Prayer and medical science: a commentary on the prayer study by Harris et al and a response to critics. PMID- 10871966 TI - Practical guidelines for clinicians who treat patients with amiodarone. Practice Guidelines Subcommittee, North American Society of Pacing and Electrophysiology. AB - Amiodarone has become an important drug for the treatment of supraventricular and ventricular arrhythmias, in short-term inpatient and outpatient settings. It may also have a role in affecting outcome in patients at high risk for arrhythmic events and sudden death; its place among available therapies is being established in clinical trials. PMID- 10871967 TI - Quality of life in patients with atrial fibrillation. AB - Atrial fibrillation (AF) is a frequent and costly health care problem. In patients with AF, the restoration and maintenance of sinus rhythm is the primary therapeutic goal. The most frequent strategy for maintaining sinus rhythm after restoration is the use of antiarrhythmic drugs. The efficacy of therapy in AF has been predominantly measured using objective criteria such as mortality and morbidity. In recent years, the importance of quality of life (QoL) as an outcome measure has been recognized. However, few studies in the literature have examined QoL in patients with AF using properly validated tools. In addition, the specific impact of antiarrhythmic drug treatment on QoL in patients with AF has not been assessed. These issues are now being addressed in several ongoing studies. This article attempts to define QoL, makes recommendations on how QoL might be assessed, reviews our current knowledge regarding QoL in patients with AF, and discusses new clinical trials currently assessing QoL in patients with AF. PMID- 10871968 TI - Association between depression and mortality in older adults: the Cardiovascular Health Study. AB - BACKGROUND: Studies of the association between depressive symptoms and mortality in elderly populations have yielded contradictory findings. To address these discrepancies, we test this association using the most extensive array of sociodemographic and physical health control variables ever studied, to our knowledge, in a large population-based sample of elderly individuals. OBJECTIVE: To examine the relation between baseline depressive symptoms and 6-year all-cause mortality in older persons, systematically controlling for sociodemographic factors, clinical disease, subclinical disease, and health risk factors. METHODS: A total of 5201 men and women aged 65 years and older from 4 US communities participated in the study. Depressive symptoms and 4 categories of covariates were assessed at baseline. The primary outcome measure was 6-year mortality. RESULTS: Of the 5201 participants, 984 (18.9%) died within 6 years. High baseline depressive symptoms were associated with a higher mortality rate (23.9%) than low baseline depression scores (17.7%) (unadjusted relative risk [RR], 1.41; 95% confidence interval [CI], 1.22-1.63). Depression was also an independent predictor of mortality when controlling for sociodemographic factors (RR, 1.43; 95% CI, 1.23-1.66), prevalent clinical disease (RR, 1.25; 95% CI, 1.07-1.45), subclinical disease indicators (RR, 1.35; 95% CI, 1.15-1.58), or biological or behavioral risk factors (RR, 1.42; 95% CI, 1.22-1.65). When the best predictors from all 4 classes of variables were included as covariates, high depressive symptoms remained an independent predictor of mortality (RR, 1.24; 95% CI, 1.06 1.46). CONCLUSIONS: High levels of depressive symptoms are an independent risk factor for mortality in community-residing older adults. Motivational depletion may be a key underlying mechanism for the depression-mortality effect. PMID- 10871969 TI - Clinical outcome and cost of hospital vs home treatment of proximal deep vein thrombosis with a low-molecular-weight heparin: the Vascular Midi-Pyrenees study. AB - BACKGROUND: Low-molecular-weight heparins have been shown to be effective and safe in the treatment of deep vein thrombosis. To our knowledge, there have been no direct comparisons of such treatment on an outpatient vs an inpatient basis. OBJECTIVE: To conduct a randomized, comparative, multicenter trial to evaluate the clinical outcomes and treatment costs of deep vein thrombosis in the outpatient and inpatient settings. METHODS: Two hundred one patients presenting with proximal deep vein thrombosis, without known risk factors for pulmonary embolism or hemorrhagic complications, were randomized to receive a low-molecular weight heparin at the registered dose followed by an oral anticoagulant for up to 6 months, either in the hospital for the first 10 days followed by treatment at home (n=102) or at home from the outset (n=99). The primary clinical outcome was the incidence of venous thromboembolism recurrence, pulmonary embolism, or major bleeding. The economic analysis was performed from the point of view of the health insurance company. Total costs of the 2 management strategies were calculated to compare the cost consequences during the first 10 days. RESULTS: No differences in clinical outcome were detectable between the 2 groups. There was no increase in the rates of primary efficacy outcome in the patients treated at home vs in the hospital (3.0% vs 3.9%), while a cost reduction of 56% was demonstrated for outpatient management. CONCLUSION: For patients with proximal deep vein thrombosis and no symptoms of pulmonary embolism or increased risk of major bleeding, home treatment using a low-molecular-weight heparin is an effective, safe, and cost-saving strategy. PMID- 10871970 TI - Association of unstable angina guideline care with improved survival. AB - BACKGROUND: An unstable angina guideline was published in 1994 by the Agency for Health Care Policy and Research, Bethesda, Md. However, the relationship between guideline-concordant care and patient outcomes is unknown. OBJECTIVE: To determine whether guideline-concordant care is associated with improved outcomes. METHODS: The study sample consisted of 275 consecutive nonreferral patients hospitalized with primary unstable angina. One-year survival and survival free of myocardial infarction were compared between patients who received care concordant with 8 selected guideline recommendations and patients who received discordant care. RESULTS: Care concordant with the 8 key guideline recommendations was associated with improved 1-year survival (95% vs 81%; log-rank P<.001) and survival free of myocardial infarction (91% vs 74%; P<.001), compared with guideline-discordant care. Patients in high-risk subgroups had the largest survival benefit associated with guideline-concordant care (aged -65 years, 91% vs 74% [P=.005]; heart failure at presentation, 91% vs 68% [P=.10]). Aspirin therapy was the single recommendation most strongly associated with improved 1 year survival (94% vs 78%; P=.002). CONCLUSIONS: Care as outlined in the unstable angina clinical practice guideline is associated with improved 1-year outcomes. Subgroups of patients at highest risk and recommendations firmly based on randomized clinical trial data were most strongly associated with better outcomes. These findings support the use of an evidence-based approach to guideline development and assessment of quality of care in patients with primary unstable angina. PMID- 10871971 TI - A randomized trial of the efficacy and tolerability of the COX-2 inhibitor rofecoxib vs ibuprofen in patients with osteoarthritis. Rofecoxib/Ibuprofen Comparator Study Group. AB - BACKGROUND: Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit both cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2). It is not known whether a specific inhibitor of COX-2 will provide efficacy in osteoarthritis (OA) comparable with NSAIDs. Therefore, we compared the efficacy and safety of the rofecoxib, which specifically inhibits COX-2, with those of the NSAID ibuprofen in patients with OA. OBJECTIVE: To compare the clinical efficacy and tolerability of rofecoxib (12.5 and 25 mg once daily) with ibuprofen (800 mg 3 times daily). METHODS: A randomized, double-blind trial of 809 adults with OA was conducted. Patients with OA in whom the knee or hip was the primary source of pain were randomized to 1 of 4 treatment groups on demonstration of disease activity: placebo; rofecoxib, 12.5 or 25 mg once daily; or ibuprofen, 800 mg 3 times daily. Clinical efficacy and safety were monitored during a 6-week treatment period. RESULTS: Both doses of rofecoxib demonstrated efficacy clinically comparable with ibuprofen as assessed by 3 primary end points (pain walking on a flat surface [Western Ontario and McMaster Universities Osteoarthritis Index], patient global assessment of response to therapy, and investigator global assessment of disease status) according to predefined comparability criteria. Both rofecoxib doses and the ibuprofen dose provided significantly (P<.001) greater efficacy than placebo on all primary end points. Results from secondary end points were consistent with those of the primary end points. All treatments were well tolerated; the overall incidence rates of clinical adverse experiences were not significantly different (P>.05) among the treatment groups. CONCLUSION: Rofecoxib was well tolerated and provided clinical efficacy comparable with a high dose of the NSAID ibuprofen. PMID- 10871972 TI - Patient satisfaction with screening flexible sigmoidoscopy. AB - BACKGROUND: Screening flexible sigmoidoscopy is an underused cancer prevention procedure. Physicians often cite patient discomfort as a reason for not requesting sigmoidoscopy, but patient experiences and attitudes toward sigmoidoscopy have not been well studied. OBJECTIVE: To measure patient satisfaction and the determinants of satisfaction with screening sigmoidoscopy. METHODS: An instrument to assess satisfaction with screening sigmoidoscopy was developed. Responses were evaluated with a factor analysis, tested for reproducibility and internal consistency, and validated against an external standard. RESULTS: A total of 1221 patients (666 men and 555 women; mean age, 61.8 years) were surveyed after sigmoidoscopy. Examinations were performed by a nurse practitioner (n = 668), internist (n = 344), or gastrointestinal specialist (n= 184). More than 93% of the participants strongly agreed or agreed they would be willing to undergo another examination, and 74.9% would strongly recommend the procedure to their friends. Regarding pain and discomfort, 76.2% strongly agreed or agreed that the examination did not cause a lot of pain, 78.1% stated that it did not cause a lot of discomfort, and 68.5% thought that it was more comfortable than they expected. Fifteen percent to 25% of the patients indicated they had a lot of pain, great discomfort, or more discomfort than expected. Women were more likely to have significant pain or discomfort than men (adjusted odds ratio, 2.9; 95% confidence interval, 1.9-4.3; P<.001). CONCLUSIONS: Approximately 70% of individuals who undergo screening sigmoidoscopy are satisfied and find the procedure more comfortable than expected, whereas only 15% to 25% find the procedure unpleasant. Physicians should not project discomfort onto patients as a reason for not requesting screening sigmoidoscopy. PMID- 10871973 TI - Relief of cardiorespiratory symptoms and increased physical activity after surgically induced weight loss: results from the Swedish Obese Subjects study. AB - BACKGROUND: Obese people frequently suffer from shortness of breath and chest discomfort on exertion, and they often have a sedentary lifestyle. In the present study of patients with severe obesity, we investigated the effects of surgically induced weight loss on cardiorespiratory symptoms and leisure-time physical activity. METHODS: The Swedish Obese Subjects study is an ongoing intervention trial of obesity consisting of 1 surgically treated group and 1 matched control group. Information on smoking habits, hypertension, diabetes, and sleep apnea was obtained from 1210 surgical cases and 1099 controls who were observed for 2 years. Patients were also asked about symptoms of breathlessness and chest pain and their levels of leisure-time physical activity. RESULTS: The surgically treated group displayed a mean weight loss of 28 kg (23%) compared with the control group in which the average weight remained unchanged (P<.001). The rates of hypertension, diabetes, and apneas during sleep decreased in surgical cases compared with controls (P<.001), while smoking habits remained largely the same. The surgical group also displayed highly significant improvements in dyspnea and chest pain and increases in physical activity compared with the control group (P<.001). The odds ratio for self-reported breathlessness, chest discomfort, or sedentary behavior after 2 years decreased progressively with the degree of weight loss. Furthermore, patients who recovered from apneas during sleep reduced their odds of having dyspnea and chest discomfort at follow-up, independent of changes in weight. CONCLUSIONS: Surgically induced weight loss in patients with severe obesity is associated with a marked relief in symptoms of dyspnea and chest pain and promotes increased leisure-time physical activity. Sleep disordered breathing may be involved in the pathophysiology of breathlessness and chest discomfort in obese subjects. PMID- 10871974 TI - Lansoprazole compared with ranitidine for the treatment of nonerosive gastroesophageal reflux disease. AB - BACKGROUND: Traditionally, proton pump inhibitors are used primarily for patients with esophagitis. However, patients with nonerosive reflux disease may also benefit from these powerful medications. OBJECTIVE: To compare the safety and symptom relief efficacy of lansoprazole with ranitidine therapy and with placebo. METHODS: In 2 randomized, double-blind, multicenter trials of 901 patients with symptomatic reflux disease, which was confirmed by endoscopy to be nonerosive, received lansoprazole, 15 or 30 mg once daily; ranitidine, 150 mg twice daily; or placebo for 8 weeks. RESULTS: Analysis of daily diary data during the first 4 weeks and for the entire 8 weeks of treatment revealed that patients who were treated with either dosage of lansoprazole reported significantly (P<.05) lower percentages of days and nights with heartburn, less pain severity of both day and night heartburn, fewer days of antacid use, and smaller amounts of antacid use compared with patients who were treated with ranitidine or placebo. The incidence of possible or probable treatment-related adverse reactions was comparable among the treatment groups; abdominal pain and diarrhea were the most commonly reported adverse events. No statistically significant differences were noted between treatment groups in laboratory analyses. CONCLUSION: Lansoprazole therapy is more effective than standard dosages of ranitidine or placebo in relieving symptoms in patients with endoscopically confirmed non-erosive reflux esophagitis. PMID- 10871975 TI - Efficacy of omeprazole for the treatment of symptomatic acid reflux disease without esophagitis. AB - BACKGROUND: Up to three quarters of patients with gastroesophageal reflux disease (GERD) have symptoms, such as heartburn, but no macroscopic evidence of erosive esophagitis, making symptomatic GERD a common clinical problem in the primary care setting. OBJECTIVE: To compare the efficacy and safety of omeprazole, 20 mg once daily; omeprazole, 10 mg once daily; and placebo in the treatment of symptomatic GERD without erosive esophagitis. METHODS: Patients with a history of heartburn (> or =12 months) and episodes of moderate to severe heartburn on 4 or more of the 7 days before endoscopy were eligible to participate in this 4-week, randomized, double-blind, placebo-controlled trial. The absence of erosive esophagitis was established through endoscopy. Eligible patients were randomized to 1 of 3 treatment groups: omeprazole, 20 mg once daily; omeprazole, 10 mg once daily; or placebo. Patients were assessed at weeks 2 and 4. The efficacy of omeprazole for the treatment of heartburn was determined mainly through the following diary card data: daily resolution of heartburn and complete resolution of heartburn every day during 1 week of treatment. The efficacy of omeprazole for the treatment of acid regurgitation, dysphagia, epigastric pain, and nausea was also assessed. RESULTS: Of 359 randomized patients, 355 were included in the statistical analysis (intention-to-treat population). Daily proportions of patients with no heartburn were consistently greater in the 20-mg omeprazole group (62%, day 7; 74%, day 27) than in the 10-mg omeprazole group (41%, day 7; 49%, day 27) or the placebo group (14%, day 7; 23%; day 27). Complete resolution of heartburn every day during the last treatment week was significantly (P< or =.002) higher in the 20-mg omeprazole group (48%) than in the 10-mg omeprazole (27%) or placebo (5%) group. Omeprazole was significantly (P< or =.003) more effective than placebo for the treatment of acid regurgitation, dysphagia, epigastric pain, and nausea. CONCLUSIONS: Patients with symptomatic GERD require profound acid suppression to achieve symptomatic relief. Omeprazole, 20 mg once daily, was superior to omeprazole, 10 mg once daily, and to placebo in providing early and sustained resolution of heartburn, as well as treatment of other troublesome GERD symptoms. PMID- 10871976 TI - Patients with depression are less likely to follow recommendations to reduce cardiac risk during recovery from a myocardial infarction. AB - BACKGROUND: Patients with depression are at greater risk of cardiac death in the first few months after a myocardial infarction (MI). This study was performed to determine whether depression affects adherence to recommendations intended to reduce the risk of cardiac events after an MI. METHODS: All consenting patients admitted to a university-affiliated teaching hospital during an 18-month period were interviewed 3 to 5 days following an acute MI using the Beck Depression Inventory to assess symptoms of depression and using the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, Revised Third Edition, to determine the presence of major depression and/or dysthymia. Accessible survivors (n=204; 116 men and 88 women) were interviewed by telephone 4 months later using the Medical Outcomes Study Specific Adherence Scale to measure self-reported adherence to recommendations to modify cardiac risk. RESULTS: Patients who were found in the hospital to have symptoms of at least mild to moderate depression (Beck Depression Inventory score > or =10, n=35 [17.2%]) or to have major depression and/or dysthymia (n=31 [15.2%]) reported lower adherence to a low-fat diet, regular exercise, reducing stress, and increasing social support 4 months later. Those with major depression and/or dysthymia also reported taking medications as prescribed less often than those without major depression and/or dysthymia. Diabetic patients with major depression and/or dysthymia were less likely to follow a diet for patients with diabetes than diabetic patients without depression. CONCLUSIONS: Patients with depression following an acute MI are less likely to adhere to recommended behavior and lifestyle changes intended to reduce the risk of subsequent cardiac events. This finding could explain why depression in the hospital is related to long-term prognosis in patients recovering from an MI. PMID- 10871977 TI - Physician, nurse, and social worker collaboration in primary care for chronically ill seniors. AB - OBJECTIVES: To examine the impact of an interdisciplinary, collaborative practice intervention involving a primary care physician, a nurse, and a social worker for community-dwelling seniors with chronic illnesses. METHODS: A concurrent, controlled cohort study of 543 patients in 18 private office practices of primary care physicians was conducted. The intervention group received care from their primary care physician working with a registered nurse and a social worker, while the control group received care as usual from their primary care physician. The outcome measures included changes in number of hospital admissions, readmissions, office visits, emergency department visits, skilled nursing facility admissions, home care visits, and changes in patient self-rated physical, emotional, and social functioning. RESULTS: From 1992 (baseline year) to 1993, the two groups did not differ in service use or in self-reported health status. From 1993 to 1994, the hospitalization rate of the control group increased from 0.34 to 0.52, while the rate in the intervention group stayed at baseline (P= .03). The proportion of intervention patients with readmissions decreased from 6% to 4%, while the rate in the control group increased from 4% to 9% (P=.03). In the intervention group, mean office visits to all physicians fell by 1.5 visits compared with a 0.5-visit increase for the control group (P=.003). The patients in the intervention group reported an increase in social activities compared with the control group's decrease (P=.04). With fewer hospital admissions, average per patient savings for 1994 were estimated at $90, inclusive of the intervention's cost but exclusive of savings from fewer office visits. CONCLUSIONS: This model of primary care collaborative practice shows potential for reducing utilization and maintaining health status for seniors with chronic illnesses. Future work should explore the specific benefit accruing from physician involvement in the collaborative practice team. PMID- 10871978 TI - Serum homocysteine concentration as an indicator of survival in patients with acute coronary syndromes. AB - BACKGROUND: Circulating homocysteine levels are predictive of survival in patients with stable coronary artery disease. The prognostic value of serum homocysteine levels, obtained in the acute phase in patients with myocardial infarction or unstable angina, is unknown. OBJECTIVES: To test the hypothesis that circulating homocysteine levels, obtained during the first 24 hours following hospital admission in patients with acute coronary syndromes, are predictive of long-term mortality. METHODS: To test this hypothesis we performed a prospective inception cohort study at a teaching hospital in Gothenburg, Sweden. A total of 579 patients (179 women and 400 men; median age, 67 years) were included (Q-wave myocardial infarction in 163 patients, non-Q-wave myocardial infarction in 210 patients, unstable angina pectoris in 206 patients). MAIN OUTCOME MEASURE: All-cause mortality. RESULTS: During a median follow-up of 628 days, 65 patients died. The serum homocysteine level (mean [SD]) was significantly lower in long-term survivors (n = 514) than in nonsurvivors (n=65) (12.3 [7.0] vs 14.3 [5.9] pmol/L; P=.003). The relative risk (all-cause mortality) for patients with homocysteine levels in the upper quartile was 2.4 (95% confidence interval, 1.5-4.0) compared with that of patients in the 3 lower quartiles. After adjustment for relevant confounders, the relative risk estimate remained significant (relative risk= 1.69; 95% confidence interval, 1.02-2.80). In a stepwise model the homocysteine level provided prognostic information additional to that of patient age, diabetes mellitus, and diuretic usage prior to hospital admission (P=.03). CONCLUSION: The serum homocysteine level on hospital admission is an independent predictor of long-term survival in patients with acute coronary syndromes. PMID- 10871979 TI - The rapidity of drug dose escalation influences blood pressure response and adverse effects burden in patients with hypertension: the Quinapril Titration Interval Management Evaluation (ATIME) Study. ATIME Research Group. AB - BACKGROUND: Antihypertensive medication doses are typically increased within several weeks after initiation of therapy because of inadequate blood pressure (BP) control and/or adverse effects. METHODS: We conducted a parallel-group clinical trial with 2935 subjects (53% women, n=1547) aged 21 to 75 years, with Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure VI stages 1 to 2 hypertension, recruited from 365 physician practices in the southeastern United States. Participants were randomized either to a fast (every 2 weeks; n=1727) or slow (every 6 weeks; n=1208) drug titration. Therapy with quinapril, an angiotensin-converting enzyme inhibitor, was initiated at 20 mg once daily. The dose was doubled at the next 2 clinic visits until the BP was lower than 140/90 mm Hg or a dose of 80 mg was reached. RESULTS: Pretreatment BP averaged 152/95 mm Hg. Patients with stage 2 hypertension reported more symptoms than those with stage 1. The BP averaged 140/86, 137/84, and 134/83 mm Hg in the slow group compared with 141/88, 137/85, and 135/84 mm Hg in the fast group at the 3 respective clinic visits. The BP control rates to lower than 140/90 mm Hg at the 3 clinic visits were (slow, fast, respectively) 41.3%, 35.7% (P<.001); 54.3%, 51.5% (P=.16); and 68%, 62.3% (P=.02). In the fast group, 10.7% of participants experienced adverse events vs 10.8% in the slow group; however, 21.0% of adverse events in the fast group were "serious" vs only 12% in the slow group. CONCLUSION: Slower dose escalation of the angiotensin converting enzyme inhibitor quinapril provides higher BP control rates and fewer serious adverse events than more rapid drug dose escalation. PMID- 10871980 TI - Use of calcium channel blockers and risk of hospitalized gastrointestinal tract bleeding. AB - BACKGROUND: We conducted a case-control study of the association between calcium channel blocker use and gastrointestinal (GI) tract bleeding in hypertensive members of a health maintenance organization. METHODS: Case patients (n=174) were treated hypertensive health maintenance organization members hospitalized for GI tract bleeding between January 1992 and December 1994. Case patients were identified using computerized diagnosis codes and were confirmed by medical record review. Control subjects (n=771) were treated hypertensive members selected from ongoing studies at the health maintenance organization. Computerized pharmacy data and medical records were used to assess medication use and other risk factors for GI tract bleeding. RESULTS: Compared with beta-blocker users, calcium channel blocker users had an age-, sex- and calendar year-adjusted relative risk (RR) of GI tract bleeding of 2.60 (95% confidence interval [CI], 1.71-3.96). The RR associated with calcium channel blocker use was 2.05 (95% CI, 1.33-3.17) after further adjustment for number of recent visits, diastolic blood pressure, chronic congestive heart failure, and duration of hypertension. No significant dose-response relationship was observed. Compared with beta-blocker users, angiotensin-converting enzyme inhibitor users had an RR of 1.22 (95% CI, 0.75-1.97). Calcium channel blocker use tended to be more strongly associated with risk of lower GI tract bleeding (RR, 2.56; 95% CI, 1.08-6.05) than with risk of upper GI tract bleeding (RR, 1.54; 95% CI, 0.91-2.59) or peptic ulcer-related bleeding (RR, 1.17; 95% CI, 0.62-2.21), although these results were compatible with chance. CONCLUSIONS: Calcium channel blocker use might be associated with an elevated risk of GI tract bleeding. These findings require confirmation in randomized studies. PMID- 10871981 TI - The cause of delirium in patients with hip fracture. AB - OBJECTIVES: To ascertain the most common causes of delirium, to establish the initiation and timing of delirium, and to determine the duration of delirium in patients with hip fracture. METHODS: Five hundred seventy-one (88%) of 650 patients with hip fracture admitted to 4 New York City hospitals were prospectively interviewed on a daily basis, 5 days a week, with the Confusion Assessment Method for the presence of delirium. The patients were enrolled within 48 hours of admission. Their medical charts and the data collected by the study staff were reviewed and summarized. Two of us (R.S.M. and A.L.S.) reviewed the case summaries independently and assigned a cause based on a previously developed classification system, estimated the onset of the delirious episode, and determined whether the delirium had cleared, improved, or persisted at discharge. Subsequently, discrepancies in cause, timing of initiation, and mental status on discharge between the 2 physicians reviewers were discussed until consensus was reached. RESULTS: The prevalence of delirium was 9.5% (54/ 571; 95% confidence interval, 7.0-11.9). Seven percent of episodes were assigned a definite cause, 20% a probable cause, 11% a possible cause, and 61% were attributable to 1 or more comorbid conditions. Twenty-eight (53%) of 54 subjects developed delirium after surgery. The delirium had cleared or improved in 40 (74%) of 54 subjects at the time of discharge. CONCLUSIONS: Delirium in patients with hip fracture appears to be a different syndrome from that observed in patients who are otherwise medically ill; it also appears to follow a different clinical course. These results have important implications for the management of delirium in patients with hip fracture. PMID- 10871982 TI - Increasing rates of ischemic heart disease in the native population of Ontario, Canada. AB - BACKGROUND: The prevalence of ischemic heart disease (IHD) has been declining in North America since the 1960s. Over this time, Native populations, which have traditionally had low rates of IHD, have undergone striking lifestyle changes that may have had health consequences. In this context, IHD trends in the Native communities of Ontario, Canada, were evaluated. OBJECTIVE: To assess trends in admission rates for IHD in the Native population of Ontario compared with the general population of Ontario. METHODS: A comprehensive administrative database of all hospital admissions in Ontario 1981 to 1997, was used. Age- and sex adjusted rates of hospital admissions with IHD-related diagnostic or procedure codes were determined in all residents of Ontario communities that had regular census participation and at least 95% of their population claiming Native origins (N=16,874 in 1991). Comparison was made with all residents of the surrounding northern Ontario region (N=822,450) and of the whole province (N = 10,084,885). RESULTS: In 1981, the rate of IHD admissions was similar in all groups, at 99 to 124 per 10,000 persons. By 1997, it decreased to 82 per 10,000 in the province (slope, -1.09; 95% confidence interval, -1.26 to -0.91), with a similar trend in northern Ontario. However, in the Native communities, it increased to 155 per 10,000 (slope, 5.6; 95% confidence interval, 3.8-7.5). A similar trend was seen for acute myocardial infarction admissions, a more precisely coded subset of IHD. Spurious causes of increasing rates were ruled out. CONCLUSIONS: Hospitalizations for IHD have doubled in the Native population despite declining rates in the general population. These findings document an alarming trend in Native health and support the need for further research and targeted intervention. PMID- 10871983 TI - Sweet syndrome associated with Pasteurella multocida bronchitis. PMID- 10871984 TI - Data without a prayer. PMID- 10871985 TI - Intercessory prayer. PMID- 10871986 TI - Waiving informed consent for research on spiritual matters? PMID- 10871987 TI - A randomized, controlled trial of prayer? PMID- 10871988 TI - P value out of control. PMID- 10871989 TI - No effect of intercessory prayer has been proven. PMID- 10871990 TI - Does prayer really set one apart? PMID- 10871991 TI - Does prayer need testing? PMID- 10871992 TI - Ethical and practical problems in studying prayer. PMID- 10871993 TI - Therapeutic efficacy of prayer. PMID- 10871994 TI - Is it prayer, or is it parity? PMID- 10871995 TI - Questions on the design and findings of a randomized, controlled trial of the effects of remote, intercessory prayer on outcomes in patients admitted to the coronary care unit. PMID- 10871996 TI - The effect of remote intercessory prayer on clinical outcomes. PMID- 10871997 TI - Prayer can help. PMID- 10871998 TI - God, prayer, and coronary care unit outcomes: faith vs works? PMID- 10872000 TI - JAMA 100 years ago: HEALTH AND SUMMER RESORTS PMID- 10872001 TI - Patient-physician relationship critical even during brief "medication checks". PMID- 10872002 TI - Alliance anxious about children's mental health. PMID- 10872003 TI - Human papillomavirus detection to screen for cervical cancer. PMID- 10872004 TI - Human papillomavirus detection to screen for cervical cancer PMID- 10872005 TI - Adjunctive therapies for wound healing. PMID- 10872006 TI - Adjunctive therapies for wound healing. PMID- 10872007 TI - Adjunctive therapies for wound healing PMID- 10872008 TI - Political solicitation of physicians. PMID- 10872009 TI - Early risks of hormone therapy in patients with coronary heart disease. PMID- 10872010 TI - Mechanical ventilation as a mediator of multisystem organ failure in acute respiratory distress syndrome. PMID- 10872011 TI - Differentiating Parkinson disease from multiple-system atrophy by measuring cardiac iodine-123 metaiodobenzylguanidine accumulation. PMID- 10872012 TI - Survival in end-stage dementia following acute illness. AB - CONTEXT: Little is known about the prognosis of acutely ill patients with end stage dementia or about the type of care that these patients receive. If their prognosis is poor, then emphasis should be placed on palliative care for these patients rather than on curative interventions. OBJECTIVES: To examine survival for patients with end-stage dementia following hospitalization for hip fracture or pneumonia and to compare their care with that of cognitively intact older adults. DESIGN: Prospective cohort study with 6 months of follow-up. SETTING AND PATIENTS: Patients aged 70 years or older who were hospitalized with hip fracture (cognitively intact, n=59; with end-stage dementia, n=38) or pneumonia (cognitively intact, n=39; with end-stage dementia, n=80) in a large hospital in New York, NY, between September 1, 1996, and March 1, 1998. MAIN OUTCOME MEASURES: Mortality, treatments directed at symptoms, and application of distressing and painful procedures in cognitively intact patients vs those with end-stage dementia. RESULTS: Six-month mortality for patients with end-stage dementia and pneumonia was 53% (95% confidence interval [CI], 41%-64%) compared with 13% (95% CI, 4%-27%) for cognitively intact patients (adjusted hazard ratio, 4.6; 95% CI, 1.8-11.8). Six-month mortality for patients with end-stage dementia and hip fracture was 55% (95% CI, 42%-75%) compared with 12% (95% CI, 5%-24%) for cognitively intact patients (adjusted hazard ratio, 5.8; 95% CI, 1.7-20.4). Patients with end-stage dementia received as many burdensome procedures as cognitively intact patients and only 8 (7%) of 118 patients with end-stage dementia had a documented decision made to forego a life-sustaining treatment other than cardiopulmonary resuscitation. Only 24% of patients with end-stage dementia and hip fracture received a standing order for analgesics. CONCLUSIONS: In this study, patients with advanced dementia and hip fracture or pneumonia had a very poor prognosis. Given the limited life expectancy of patients with end stage dementia following these illnesses and the burdens associated with their treatment, increased attention should be focused on efforts to enhance comfort in this patient population. JAMA. 2000;284:47-52 PMID- 10872013 TI - Effect of a monetary sanction on immunization rates of recipients of aid to families with dependent children. AB - CONTEXT: Immunization rates among low-income families have lagged behind those for the general community, with several possible barriers cited in the literature. OBJECTIVE: To evaluate the effect of an initiative aimed at improving immunization rates among low-income preschool children by imposing a sanction on families who failed to provide proof of up-to-date immunization status. DESIGN AND SETTING: Randomized, controlled before-after trial conducted from January 1, 1993, through December 31, 1996, in Muscogee County, Georgia. PARTICIPANTS: A total of 2500 families with children aged 6 years or younger who received Aid to Families with Dependent Children assistance. INTERVENTION: Families in the intervention group (n=1500) were informed that receipt of the welfare benefit for any preschool-aged children was contingent on provision of proof of up-to-date immunization status at the beginning of welfare eligibility and, subsequently, semiannually or annually. Case families in the control group (n=1000) were encouraged to immunize their preschool children but were not informed of any aid sanctions nor did such sanctions apply to them. MAIN OUTCOME MEASURE: Age appropriate rates of 5 immunizations (measles-mumps-rubella; poliovirus; diphtheria and tetanus toxoids and pertussis; Haemophilus influenzae type b; and hepatitis B), based on examination (with family's written consent) of medical provider records, compared among intervention-group vs control-group families. RESULTS: There were no significant differences at baseline between intervention and control families in immunization rates of preschool children. Families in the intervention group were significantly more likely than families in the control group to have up-to-date immunization status in all 4 years of the study for all 5 immunizations (with 3 exceptions). At age 2 years, 72.4% of children in the intervention group vs 60.6% of those in the control group achieved vaccine series completion, which included 4 diphtheria and tetanus toxoids and pertussis, 3 poliovirus, and 1 measles-mumps-rubella (P<.001). Sanctions were implemented only 11 times. There was relatively little increased burden on the part of families to comply with requirements. CONCLUSION: In our study, a monetary sanction in a population receiving welfare benefits stimulated a significant increase in childhood immunization rates, suggesting that when welfare recipients are given an incentive to keep their children's immunizations up-to-date, most are able to do so. JAMA. 2000;284:53-59 PMID- 10872014 TI - Effect of a community intervention on patient delay and emergency medical service use in acute coronary heart disease: The Rapid Early Action for Coronary Treatment (REACT) Trial. AB - CONTEXT: Delayed access to medical care in patients with acute myocardial infarction (AMI) is common and increases myocardial damage and mortality. OBJECTIVE: To evaluate a community intervention to reduce patient delay from symptom onset to hospital presentation and increase emergency medical service (EMS) use. DESIGN AND SETTING: The Rapid Early Action for Coronary Treatment Trial, a randomized trial conducted from 1995 to 1997 in 20 US cities (10 matched pairs; population range, 55,777-238,912) in 10 states. PARTICIPANTS: A total of 59,944 adults aged 30 years or older presenting to hospital emergency departments (EDs) with chest pain, of whom 20,364 met the primary population criteria of suspected acute coronary heart disease on admission and were discharged with a coronary heart disease-related diagnosis. INTERVENTION: One city in each pair was randomly assigned to an 18-month intervention that targeted mass media, community organizations, and professional, public, and patient education to increase appropriate patient actions for AMI symptoms (primary population, n=10,563). The other city in each pair was randomly assigned to reference status (primary population, n=9801). MAIN OUTCOME MEASURES: Time from symptom onset to ED arrival and EMS use, compared between intervention and reference city pairs. RESULTS: General population surveys provided evidence of increased public awareness and knowledge of program messages. Patient delay from symptom onset to hospital arrival at baseline (median, 140 minutes) was identical in the intervention and reference communities. Delay time decreased in intervention communities by -4.7% per year (95% confidence interval [CI], -8.6% to -0.6%), but the change did not differ significantly from that observed in reference communities (-6. 8% per year; 95% CI, -14.5% to 1.6%; P=.54). EMS use by the primary study population increased significantly in intervention communities compared with reference communities, with a net effect of 20% (95% CI, 7%-34%; P<.005). Total numbers of ED presentations for chest pain and patients with chest pain discharged from the ED, as well as EMS use among patients with chest pain released from the ED, did not change significantly. CONCLUSIONS: In this study, despite an 18-month intervention, time from symptom onset to hospital arrival for patients with chest pain did not change differentially between groups, although increased appropriate EMS use occurred in intervention communities. New strategies are needed if delay time from symptom onset to hospital presentation is to be decreased further in patients with suspected AMI. JAMA. 2000;284:60-67 PMID- 10872015 TI - Physicians' recommendations to patients for use of antibiotic prophylaxis to prevent endocarditis. AB - CONTEXT: The American Heart Association recommendations for infectious endocarditis (IE) prophylaxis, published in June 1997, sought to improve patient and physician compliance by simplifying the dosing regimen and clarifying endocarditis risk. Adherence to these updated recommendations in patients with echocardiographic verification of their endocarditis risk profile is unknown. OBJECTIVE: To determine the recommended and actual use of IE prophylaxis as reported by patients undergoing echocardiography. DESIGN, SETTING, AND PARTICIPANTS: All patients who underwent outpatient transthoracic echocardiography at a university-based tertiary hospital in Boston, Mass, during December 1997 were contacted 6 to 9 months later to respond to a survey, completed by 218 (80%) eligible subjects. MAIN OUTCOME MEASURE: Patients' report of their physicians' instructions on actual use of IE prophylaxis in accordance with patient risk category, determined by echocardiographic data. RESULTS: One hundred eight patients (49.5%) had clinical or echocardiographic findings for which prophylaxis was indicated. Of these 108 patients, 71 (65.7%) reported that they were instructed to take IE prophylaxis. Sixteen high-risk patients (88. 9%) but only 55 moderate-risk patients (61.1%) reported that they were instructed to take prophylaxis. Among the 110 negligible-risk patients, 29 (26.4%) reported that they had been instructed to take IE prophylaxis. Overall, 100 patients (45.9%) reported that they received physician instructions to take IE prophylaxis. Of those who subsequently underwent a procedure for which IE prophylaxis was indicated (n=68), 9 (13.2%) elected not to follow their physician's advice to take prophylaxis. CONCLUSIONS: We found that although most patients reported receiving instructions for IE prophylaxis use consistent with American Heart Association guidelines, IE prophylaxis overuse among negligible risk patients and underuse among moderate-risk patients was common. Continued physician and patient education may lead to improved adherence to the current American Heart Association recommendations. JAMA. 2000;284:68-71 PMID- 10872016 TI - Ischemic stroke risk with oral contraceptives: A meta-analysis. AB - CONTEXT: The relationship between ischemic stroke and oral contraceptive (OC) use has been studied for 40 years, but disagreement about an association persists. OBJECTIVE: To review available literature to determine whether OC use is associated with increased stroke risk. DATA SOURCES: Studies published from January 1960 through November 1999 were identified from electronic databases (MEDLINE, BIOSIS, and Dissertation Abstracts Online), Index Medicus, bibliographies of pertinent review articles and pertinent original articles, textbooks, and expert consultation. STUDY SELECTION: From 804 potentially relevant references retrieved, 73 were studies investigating risk of ischemic stroke with OC use. Two reviewers independently applied the following inclusion criteria: more than 10 stroke cases sampled, clear stroke subtype differentiation, concurrent controls included, adequate data included to determine relative risks (RRs) and confidence intervals (CIs), analysis controlled for age, and no later publication of identical data. A third investigator adjudicated disagreements. Sixteen studies met all inclusion criteria and were included in the meta-analysis. DATA EXTRACTION: Two investigators independently extracted data, with disagreements resolved through discussion. DATA SYNTHESIS: The 16 studies were analyzed using random effects modeling. Current OC use was associated with increased risk of ischemic stroke (RR, 2.75; 95% CI, 2.24-3.38). Smaller estrogen dosages were associated with lower risk (P=.01 for trend), but risk was significantly elevated for all dosages. Studies that did not control for smoking (P=.01) and those using hospital-based controls (P<.001) found higher RRs, but no other patient characteristics or elements of study design were important. The summary RR was 1.93 (95% CI, 1.35-2.74) for low-estrogen preparations in population-based studies that controlled for smoking and hypertension. This translates to an additional 4.1 ischemic strokes per 100,000 nonsmoking, normotensive women using low-estrogen OCs, or 1 additional ischemic stroke per year per 24,000 such women. The RR of stroke due to OC use was not different in women who smoked, had migraines, or had hypertension. CONCLUSIONS: Summary results indicate that risk of ischemic stroke is increased in current OC users, even with newer low-estrogen preparations. However, the absolute increase in stroke risk is expected to be small since incidence is very low in this population. JAMA. 2000;284:72-78 PMID- 10872017 TI - Users' guides to the medical literature: XXII: how to use articles about clinical decision rules. Evidence-Based Medicine Working Group. AB - Clinical experience provides clinicians with an intuitive sense of which findings on history, physical examination, and investigation are critical in making an accurate diagnosis, or an accurate assessment of a patient's fate. A clinical decision rule (CDR) is a clinical tool that quantifies the individual contributions that various components of the history, physical examination, and basic laboratory results make toward the diagnosis, prognosis, or likely response to treatment in a patient. Clinical decision rules attempt to formally test, simplify, and increase the accuracy of clinicians' diagnostic and prognostic assessments. Existing CDRs guide clinicians, establish pretest probability, provide screening tests for common problems, and estimate risk. Three steps are involved in the development and testing of a CDR: creation of the rule, testing or validating the rule, and assessing the impact of the rule on clinical behavior. Clinicians evaluating CDRs for possible clinical use should assess the following components: the method of derivation; the validation of the CDR to ensure that its repeated use leads to the same results; and its predictive power. We consider CDRs that have been validated in a new clinical setting to be level 1 CDRs and most appropriate for implementation. Level 1 CDRs have the potential to inform clinical judgment, to change clinical behavior, and to reduce unnecessary costs, while maintaining quality of care and patient satisfaction. JAMA. 2000;284:79-84 PMID- 10872018 TI - Ethical considerations in the public policy laboratory. PMID- 10872019 TI - Hospital care of patients with dementia. PMID- 10872020 TI - The contributions of authors. PMID- 10872021 TI - Deaths due to medical errors are exaggerated in Institute of Medicine report. PMID- 10872022 TI - Institute of Medicine medical error figures are not exaggerated. PMID- 10872023 TI - A piece of my mind: A view from there. PMID- 10872025 TI - Perspectives in Olfactory Loss Following Viral Infections of the Upper Respiratory Tract. PMID- 10872027 TI - A Criterion for Distinguishing Level V Nodes From Clavicular Nodes. PMID- 10872026 TI - A Perplexing Olfactory Loss. PMID- 10872028 TI - Invited Critique. PMID- 10872029 TI - Invited Critique. PMID- 10872030 TI - Alosetron (Lotronex) for treatment of irritable bowel syndrome. PMID- 10872031 TI - Beta-blockers for heart failure. PMID- 10872032 TI - Drug interactions with St. John's wort. PMID- 10872033 TI - Morphometric analysis of AgNORs in tubular and papillary parts of canine mammary gland tumors. AB - OBJECTIVE: To test the value of the silver staining nucleolar organizer regions (AgNORs) technique on canine mammary gland tumors using image analysis and to estimate differences in AgNOR parameters in structurally different parts of canine mammary gland tumors. STUDY DESIGN: Analysis was performed on 13 complex type and 10 simple type malignant canine mammary gland tumors containing tubular and/or papillary structures. Ten normal mammary glands were used as controls. Morphometric analysis was done by a computer-assisted image analysis system and consisted of evaluation of nuclear area, number and area of AgNORs per nuclear area, ratio of nuclei with five or more AgNORs, nuclear perimeter, area fraction between nuclear area and area of AgNORs, and area, equivalent diameter, volume equivalent sphere, perimeter and circularity of a singular AgNOR. RESULTS: Distinct differences were detected between normal and malignant mammary gland tissue for all measured parameters. There were no significant differences between the tubular and papillary parts of the same tumor or between the tubular and papillary parts of complex and simple type tumors. CONCLUSION: Despite the fact that no significant differences were found for AgNOR parameters between papillary and tubular structures of mammary gland tumors, the results of grouping tumors by the number of AgNORs indicate that this might help with classification of canine mammary gland tumors. PMID- 10872034 TI - Grading of ductal breast carcinoma by cytomorphology and image morphometry with histologic correlation. AB - OBJECTIVE: To investigate the relevance of image analysis for grading breast carcinoma. STUDY DESIGN: Twenty-five ductal breast carcinoma cases were chosen randomly from routine fine needle aspiration clinics. The results of cytomorphologic grading and image morphometry were correlated with those of histologic grading. The five image morphometric parameters studied were nuclear diameter, nuclear area, nuclear roundness, nuclear perimeter and grey level to compare with chromatin texture. RESULTS: Cytologic grading alone had a high correlation with histologic grading. The lowest correlation was found in grade 2 tumors. When cytologic grading was supplemented with image morphometric parameters, the correlation was higher than that of cytologic grading alone. CONCLUSION: Cytologic grading has a high correlation with histologic grading. The correlation improves further on supplementation with image morphometric parameters. PMID- 10872035 TI - Image cytometry detection of breast cancer cells with > 5C DNA content and minor DNA stemlines. AB - OBJECTIVE: To determine if the presence of cells having a DNA content > 5c and occurring at very low frequency is related to breast cancer outcome. STUDY DESIGN: Feulgen-stained imprints of fresh tumors used for routine standard DNA image cytometry were reanalyzed, with the aim of detecting hyperploid (> 5c) cells or minor stemlines. Specially adapted software was used. RESULTS: The new DNA analysis showed discordance of 47.3% with standard DNA cytometry. Minor stemline or rarely occurring 5c exceeding cells were found. These were not detected by the first DNA analysis. The presence of both DNA hyperploid cells occurring as rare events and a DNA hyperploid stemline was related to outcome. CONCLUSION: The detection of DNA hyperploid cells, even in very small numbers, appears essential to outcome, particularly in diploid or single DNA aneuploid breast cancers. PMID- 10872036 TI - AgNOR analysis of atypical ductal hyperplasia and intraductal carcinoma of the breast. AB - OBJECTIVE: To determine the diagnostic and prognostic value of argyrophilic nucleolar organizer regions (AgNORs) in atypical ductal hyperplasia (ADH), ductal carcinoma in situ (DCIS) and microinvasive ductal carcinoma (MDCA) of the breast. STUDY DESIGN: Image analysis of histologic sections from biopsies of 46 breast ADH and DCIS and 18 cases of MDCA. Determination of morphometric features of cell nuclei and nucleolar organizer regions by using AMBA software system. Data were compared with the estrogen receptor/progesterone receptor (ER/PR) content as well as with the growth fraction, determined immunohistochemically. RESULTS: AgNOR number and total AgNOR area increased from ADH to DCIS. The highest values were recorded in cases of DCIS with microinvasion. Differences between ADH and intraductal or microinvasive ductal carcinoma were statistically significant. Within the group of intraductal carcinomas, the lowest values were measured in the solid type and highest values in the comedo type. A correlation was found between AgNOR features and growth fraction but not between these features and ER/PR status. CONCLUSION: Selected AgNOR features are relevant for differentiation between ADH and DCIS as well as between low and high grade DCIS and microinvasive ductal carcinoma. Therefore, objective and reproducible data obtained by AgNOR analysis may allow better evaluation of the prognostic significance of these lesions. PMID- 10872037 TI - Real-time quantification of the proliferative state in astrocytomas. AB - OBJECTIVE: To evaluate proliferative activity in a set of gliomas and to compare the quantitative data obtained by a real-time processor with the labelling index (LI) and mitotic index (MI). STUDY DESIGN: Ki-67 immunostaining was performed on paraffin-embedded specimens from 42 cases of glioblastomas, 17 cases of anaplastic astrocytomas and 14 cases of low grade astrocytomas. Nuclear positivity was calculated as LI and by a real-time image processor for quantitative evaluation. MI was also calculated at 10 high-power fields. The data obtained from glioblastomas were compared with those from anaplastic and low grade astrocytomas. To all the data was applied the Pearson test to verify the correlation between counting and quantitative values and between proliferative markers and survival. RESULTS: A positive trend from low grade astrocytomas to glioblastomas was found for Ki-67 (LI and quantitative values) and MI, with highly significant differences between the three grades of gliomas considered. A good correlation between LI and quantitative values of Ki-67 was found. Very little relationship resulted between survival and Ki-67 LI. No relationship was found between survival and quantitative values of Ki-67. CONCLUSION: Ki-67 allowed effective separation of astrocytic tumors with different grades of malignancy. Quantitative evaluation of color information by means of a real-time processor proved to be a useful, objective and fast way to obtain readings, useful for grading purposes but not for prognostic evaluation. PMID- 10872038 TI - Sensitivity of Seescan TV image analysis in discriminating between normal, dysplastic and malignant oral smears. AB - OBJECTIVE: Smears from premalignant and malignant lesions may contain large proportions of normal cells together with atypical cells; that could reduce the sensitivity of cytologic diagnosis. The present study assessed the performance of the Seescan TV image analysis system (TVIAS) in distinguishing between normal, premalignant and malignant oral smears. The sensitivity of Seescan TVIAS was tested using both white and monochromatic light. STUDY DESIGN: Nuclear area (NA) and corrected integrated optical density (IOD) of 50 Feulgen-stained nuclei were measured in smears collected from normal oral mucosa (n = 6), lesions displaying epithelial dysplasia (n = 5) and invasive squamous cell carcinoma (n = 5) using a Seescan TVIAS with both white and monochromatic light. RESULTS: There was a significant increase (P < .001) in mean IOD for nuclei in smears from dysplastic lesions and carcinomas as compared with normal smears. For smears from carcinomas, the mean NA was significantly elevated as compared with dysplastic (P < .001) and normal smears (P < .01). Mean NA for dysplastic smears was significantly reduced as compared with normal smears. While all smears from premalignant and malignant lesions contained mostly normal nuclei, a significant proportion of abnormal nuclei was identified in each smear. CONCLUSION: Although oral smears contain large amounts of normal cells, the Seescan TVIAS could successfully identify dysplastic and malignant cells on the basis of both IOD and NA values with or without the use of a monochromatic filter. PMID- 10872039 TI - p53 protein expression and proliferative activity in imprints of superficial transitional cell carcinoma of the bladder. AB - OBJECTIVE: To investigate p53 protein expression and proliferative activity in imprints of tumor biopsies from superficial transitional cell carcinoma of the bladder in relation to the histologic grade of malignancy and recurrence status. STUDY DESIGN: The study group consisted of 70 cases of superficial transitional cell carcinoma of the bladder. In order to investigate p53 protein expression and Ki-67 expression, an immunocytochemical avidin-extravidin complex technique was performed using monoclonal antibodies p53 D0-7 and proliferating cells correspondingly. RESULTS: Thirty-seven percent of superficial transitional cell carcinoma cases showed positive expression of p53 protein. No correlation was found between p53 protein expression and grade of malignancy (P = .45). p53 Protein expression was statistically correlated with a high Ki-67 labeling index (LI) (P < .001) and recurrence status (P < .001). Forty-seven percent of cases showed a Ki-67 LI > 25%. No correlation was found between a high Ki-67 LI and grade of malignancy (P = .703). A significant difference in high Ki-67 LI between recurrent and nonrecurrent tumors of the same grade (P < .001) and between recurrent and nonrecurrent tumors was found independently of grade (P < .001). CONCLUSION: These results on cytologic material could provide useful information on the biologic behavior of superficial transitional cell carcinoma of the bladder at the time of diagnosis. PMID- 10872040 TI - MIB-1 immunohistometry of follicular adenoma and follicular carcinoma of the thyroid gland. AB - OBJECTIVE: To analyze cellular proliferative activity, MIB-1 immunopositivity of normal tissue (n = 20), follicular adenoma (n = 30) and follicular carcinoma (n = 32) of the thyroid gland was analyzed by means of immunohistometry. STUDY DESIGN: Immunohistochemical reactions were performed on 3-micron sections from routinely formalin fixed and paraffin embedded surgical specimens using an indirect peroxidase method. The rate of immunostained cells was determined using the CM-2 TV image analysis system (Hund, Wetzlar, Federal Republic of Germany). Forty viewing fields (1.94 mm2) were measured with 20:1 objective magnification. An average of 5,965 cells were assessed in each case. RESULTS: Mean MIB-1 immunopositivity was higher in follicular carcinoma (average, 2.30%) and follicular adenoma (0.58%) than in normal thyroid tissue (0.14%). The distribution of single values differed significantly between groups (P < .001). To test the suitability of MIB-1 immunohistometry for the differential diagnosis of follicular adenoma and follicular carcinoma, different four-field tables with varying thresholds were calculated. Using a threshold of 0.9%, follicular carcinoma could be detected with a sensitivity of 75% (24/32) and a specificity of 83% (25/30). If a specificity of 90% is required (27/30), the sensitivity of the test decreases to 69% (22/32), based on a threshold of 1.1%. CONCLUSION: As some overlap of single values has to be considered, MIB-1 immunohistometry, although presenting new insights into the proliferative potential of thyroid lesions, is of only limited value for the differential diagnosis of follicular lesions in routine surgical pathology. PMID- 10872041 TI - Chromatin texture analysis of cortical adrenal gland adenomas, including incidentalomas, and adjacent normal-appearing cortical tissue. AB - OBJECTIVE: To describe, by morphometric and chromatin texture analysis, a series of adrenal gland lesions, including Cushing's and Conn's adenomas and incidentalomas. STUDY DESIGN: The material for the study consisted of five consecutive cases of incidentaloma, three cases of Conn's adenoma and three cases of Cushing's adenoma. Also included were five cases of adrenal carcinoma. Sections were stained according to the Feulgen procedure. Measurements were taken from the nodules and from two different zones, identified as outer and inner parts, of the normal-appearing adrenal cortex adjacent to the tumor. Data on approximately 50 nuclei were recorded for each of these three sites (tumor and outer and inner normal-appearing adrenal cortex). The nuclei were subjected to feature extraction and were analyzed by identification procedures--i.e., establishing nuclear and lesion signatures. RESULTS: The total optical density (OD) distributions of the nuclei from the normal-appearing adrenal cortex pointed to their diploid or near-diploid nature. In incidentalomas there was a very small increase in the number of nuclei, with increased total OD. In Conn's adenoma there was a noticeable but modest extension of the total OD distribution into the higher OD range. This trend continued for Cushing's adenoma. The pixel OD histograms for nuclei from normal-appearing tissue and from incidentalomas were hardly distinguishable. Starting with nuclei from Conn's adenoma, a shift toward lower pixel OD values began. The trend continued for nuclei from Cushing's adenoma and was very pronounced for nuclei from carcinoma. The nuclear signatures showed no appreciable difference between nuclei from normal-appearing cortex and from incidentaloma. Nuclei from Conn's adenoma were more similar to those from normal tissue in their signatures than nuclei from Cushing's adenoma. In fact, the nuclear signatures from Cushing's adenoma were almost identical to those of carcinoma. The lesion signatures for normal tissue, incidentaloma and Conn's adenoma confirmed the results seen in the nuclear signatures. There was a very modest increase in the number of nuclei with greater deviation from normal in incidentalomas, and the trend was more obvious in Conn's adenoma. However, in Cushing's adenoma there was a very substantial increase in the number of nuclei, with large deviations of their nuclear chromatin texture from normal. CONCLUSION: Computer-assisted analysis of nuclear characteristics proved useful in identifying and describing differences between groups of tumors arising in the adrenal cortex and highlighted the similarity between incidentalomas and adjacent normal-appearing cortex and between Cushing's adenoma and adrenal carcinoma. PMID- 10872042 TI - Measurement of erythrocytes on diagnostic slides by scanning electron microscopy. AB - OBJECTIVE: To determine whether the measured sizes of erythrocytes in both paraffin-embedded sections and air-dried blood smears differ from values published in standard texts. STUDY DESIGN: Routinely prepared surgical pathology slides as well as an air-dried blood smear were viewed with a scanning electron microscope. Erythrocytes were measured using the instrument software. RESULTS: Erythrocyte size in the peripheral blood smear correlated well with textbook values, 7.2-7.9 microns. However, red blood cells within sectioned material from several laboratories showed a prominent decrease, ranging from 25% to 35%, as compared to textbook values, about 7 microns. CONCLUSION: Since cytologists and surgical pathologists often use the erythrocyte as a convenient marker on diagnostic slides, attention should be given to these observations in making sizing judgments. PMID- 10872043 TI - Automated surface and volume measurement of fused cells. AB - OBJECTIVE: To design and evaluate an algorithm to automatically calculate membrane area, volume and derived values from grey scale microscopic images of pairs of spheroid fused cells, especially human erythrocytes. STUDY DESIGN: Pairs of fused cells ("doublets") were identified by their high optical contrast, which resulted from their unhemolysed state. Global thresholding and noise-removing algorithms were applied to the image and resulted in a binary, "8"-shaped contour. The contour was used as a base for the calculation of a two-dimensional weighted distance histogram, the hilltops of which could be identified as center points of the contour's spheres. This allowed calculation of the distance of both center points and of the spheres' radii. With these three values, calculation of membrane area, volume and other derived values of the doublets became possible. High-speed time series were created based on consecutive images of the postfusion swelling and hemoglobin ejection from erythrocyte doublets at different temperatures. RESULTS: The influence of observation temperature on the dynamics of electrofused erythrocytes was measured with the algorithms given, and results were in agreement with physical changes in cell plasma viscosity. Images taken by optical and electron microscopy were in agreement with the two-spheres model of a doublet. The algorithm was not affected by fragmented contours. CONCLUSION: The velocity of hemoglobin ejection from electrofused erythrocytes and the relative change in static membrane area increase with temperature. The algorithm delivered reliable geometric values of fused cell configurations. PMID- 10872044 TI - Nuclear volume estimation using different sampling, measurement and calculation methods. AB - OBJECTIVE: To study the reliability of volume parameter measured on tissue sections through different sampling, measurement and calculation methods. STUDY DESIGN: The largest nuclear profile image under a 100x, NA 1.30 oil immersion objective of primary spermatocytes and spherical spermatoblasts on 11-micron thick seminiferous tubule sections and tissue images, under a 20x objective, on 4 micron sections were captured. Their volumes were measured and calculated by the five methods provided by the Technology for Image and Graphics Engineering Research cell image analysis system. RESULTS: The nuclear volumes obtained by nucleator and area equivalent diameter on the largest nuclear profile image were almost the same, including binary images by automated and manual interactive nucleator and grey scale images only by the latter. Nuclear volumes, calculated by random Feret diameter and equivalent diameter of the perimeter, the minimal circumference of the largest nuclear profile binary image, were obviously larger than those of the nucleator and area equivalent diameter. Due to different-sized nuclear slices entrapped in the same section, those nuclear volumes from the seminiferous tubule tissue images were strikingly lower than that of the largest nuclei profile image. The shape factors of primary spermatocytes and spherical spermatoblast nuclei under 100x and 20x objectives were approximately the same. CONCLUSION: The sample preparation, sampling methods and calculation formulas suitable to nuclear form are necessary to obtain reproducible volume parameters. PMID- 10872045 TI - Quantitative chromatics analysis for computer imaging of cytologic subtypes of lung cancer stained by Papanicolaou stain. AB - OBJECTIVE: To determine the role of quantitative chromatics analysis in the classification of subtypes of lung cancer stained by Papanicolaou stain. STUDY DESIGN: By means of computer image analysis, 60 keratinized squamous carcinoma cells (KSCC), 88 nonkeratinized squamous carcinoma cells (NKSCC) and 150 adenocarcinoma cells (ACC) from lung cancer in sputum smears stained by Papanicolaou stain were analyzed and distinguished based on quantitative colorimetry. The features measured were the content of three primary colors, red (R), green (G) and blue (B) and the coefficients of R, G and B (r, g and b, respectively). Hue, saturation, brightness and gray level were also measured. A stepwise discriminant analysis was carried out. RESULTS: The values of R, G and B and r, g and b, hue and saturation in NKSCC and ACC were significantly different from those of KSCC, and the changes in the three primary colors were more sensitive than those in the gray level. Computer assessment based on three primary color coefficients, hue and saturation yielded accuracy of distinguishing KSCC from NKSCC and KSCC from ACC of 95.2% and 95%, respectively. CONCLUSION: Quantitative analyses of R, G and B and r, g, b and hue and saturation are valuable in distinguishing KSCC from NKSCC and ACC. PMID- 10872046 TI - Correlation of bFGF, FGFR-1 and VEGF expression with vascularity and malignancy of human astrocytomas. AB - OBJECTIVE: To investigate the correlation of angiogenic factor expression levels with the degrees of malignancy and vascularity and their clinicopathologic significance in astrocytomas. STUDY DESIGN: Factor VIII-related antigen (FVIII RAg) was used as the marker of endothelia and basic fibroblast growth factor (bFGF); FGF receptor (FGFR)-1 and vascular endothelial growth factor (VEGF) were qualitatively and quantitatively detected with immunohistochemistry and image analysis in 61 brain astrocytomas. The correlation with tumor grades, angiogenesis and prognosis was studied. RESULTS: Measurement of FVIIIRAg expression could describe endothelial proliferation and vascularity, which were related to grade of tumor and prognosis. bFGF and VEGF expression levels in neoplastic astrocytes and endothelia were significantly different in various grades of astrocytoma. These angiogenic factors affected the positive reaction areas and integral optical densities of FVIII-RAg as well as survival time. In contrast, the expression of FGFR-1 was related to neither bFGF nor FVIIIRAg and had no significant effect on tumor malignancy. CONCLUSION: Positive regulation by bFGF and autocrine/paracrine VEGF contributes to the growth and angiogenesis of astrocytomas. Measurement of endothelial cell proliferation with FVIIIRAg in tumor stroma and quantitative detection of angiogenic factor levels in neoplastic cells had prognostic value in brain astrocytomas. The results also indicate that inhibiting bFGF and VEGF expression and/or blocking their effects could be a very useful therapeutic strategy for malignant gliomas. PMID- 10872047 TI - Low- and high-magnification image analysis of cytologic material to discriminate between benign and malignant breast lesions. PMID- 10872048 TI - [The properties of liquid water in electric and magnetic fields]. AB - A new model of liquid water is proposed, which involves the liquid-crystalline phase, namely, the presence of linearly ordered chains of water molecules and large clusters of these molecules. The presence of linearly ordered chains and clusters in liquid water makes itself evident after the exposure of water to low frequency electromagnetic radiation. The kinetics of the linearly ordered chains is described by nonlinear excitations (solitons). A hydrodynamic equation describing the behavior of this model system was derived. On the basis of the model, the dimensions of clusters were determined (approximately 600 A). The calculated values of dielectric permeability in the range of low frequencies appeared to be close to their experimentally obtained values. A tenfold increase in the stationary value of dielectric permeability compared with the known value of the dielectric constant in the frequency range of 10(4)-10(6) Hz was predicted. The frequency dependence of dielectric permeability in this range was studied. PMID- 10872049 TI - [Statistical distribution of dipeptides in protein structures and dynamic characteristics of some protein fragments]. AB - Statistical distributions of the occurrence of dipeptide fragments in proteins were studied. Various algorithms of ordering of files of frequency distribution were used. A correlation of occurrence of pairs of amino acid residues in various classes of proteins was established. The problem of the dynamic compatibility of amino acid residues in protein structures is discussed. The dynamic properties of frequently and seldom occurring dimers of amino acids are compared. PMID- 10872050 TI - [About the effect of the amplitude of fluctuations on the coefficient of rubbing of the brownian oscillator in aqueous media]. AB - The methods of molecular dynamics and correlation functions of coordinates were applied to investigate molecular oscillations in water solution at a variation of the amplitude of fluctuations. It was established, that the amplitude of fluctuation exceeds the threshold value (in water of the order of 0.3-0.45 A), of the force of friction sharply increases, and the vibration mode changes to the mode of limited diffusion. The consistency of the results with the theory of Mossbauer effect and the model of Brownian oscillator for conformational mobility in biopolymers is discussed. PMID- 10872051 TI - [Protein folding with molecular chaperones:stochastic process under control]. AB - Protein folding in a living cell occurs with the participation of specialized proteins, molecular shaperons. The functional role and molecular mechanism of action of shaperons are discussed. It is shown that shaperons can be considered as proteins that, upon interaction with the folding peptide chain, transform the spontaneous folding to a process controlled and regulated by cellular factors. Models describing these controlled phenomena are discussed. PMID- 10872052 TI - [Characterization of sensitivity and specificity of fluorescent markers of structural state and binding ability of proteins]. AB - Methods for determining the sensitivity of fluorescent probes and their classification with respect to sensitivity and specificity using human serum albumin as a reference protein and 8-anili-nonaphthalenesulfonate, K35 and pyron red as probes are proposed. The indicators of sensitivity and specificity of fluorescent probes as markers of the structural state of proteins are the activation of the probe fluorescence in excess of protein and the constant of its binding to the strong highly affine center of protein sorption. Fluorescent probes as markers of the binding ability are assessed by a protein-dependent increase in fluorescence under the conditions of excess probe and from the constant of its binding to weak secondary binding sites of the protein. PMID- 10872053 TI - [Regularities in the kinetic of photoactivation of lactate dehydrogenase by the action of UV light in different microenvironment]. AB - The kinetics of photoinactivation of cardiac (H4) and muscular (M4) isoforms of lactate dehydrogenase irradiated by UV light (240-390 nm) in the free form and in the presence of sodium azide, D-mannitol, and serotonin was studied. It was shown that the decrease in the catalytic activity of both enzymes can be described by the kinetics of the first-order monomolecular reaction. The inactivation rate constant of lactate dehydrogenase M4 is considerably higher than that of lactate dehydrogenase H4, indicating a greater photochemical lability of the isoform M4. It was shown that sodium azide has a different protective action on the proteins studied. The irradiation of the muscular isoform in the presence of serotonin and D-mannitol did not change the character of the "dose-effect" curve and only led to a decrease in the photoinactivation rate constant of the protein. PMID- 10872054 TI - [Analysis of peculiarities in nucleotides disposition in the origin of chromosome replication-oriC from Escherichia coli]. AB - Along with symmetrical features (palindromes, direct and inverted repeats), periodicities in the disposition of nucleotides in the origin of chromosome replication oriC from E. coli were studied by means of Fourier analysis. Peaks corresponding to the periods T = 2, 17, 95-100 nucleotides are the highest in the Fourier spectrum of oriC. Peaks corresponding to the periods T = 3, 11, 19, 13, 24, 27, 28, 41, 79-81 nucleotides are also prominent, but not so high. Thus, the main periodicities of the oriC spectrum of are not multiple of the B-DNA sugar phosphate backbone period, which destabilize DNA at oriC and contributes to the spontaneous unwinding of DNA. The differences between the Fourier spectrum of oriC and those of regions adjacent to oriC are demonstrated. PMID- 10872055 TI - [Kinetic-thermodynamic aspects of catalysis of polysaccharides by native end immobilized amylases]. AB - It was shown that covalent immobilization of 1.4-alpha-glucanohydrolase (glucoamylase) and 2.1-beta-D-fructanfructanohydrolase (inulase) on ionites leads to an increase in the activation energy Eact of hydrolysis of polysaccharides and a change in entalphy delta H as compared with native enzymes. During binding to the matrix, multipoint interactions of polypeptide chains with active groups take place, which are accompanied by an increase in the Michaelis constant KM, a decrease in the maximum rate of hydrolysis Vmax, and a substantial decrease in the constant of conformational transition L0. It was also shown that the kinetics of the hydrolysis of starch and inulin upon immobilization of glucoamylase and inulase on ionites does not correspond to the Michaelis-Menten equiation and is characterized by a shift of equilibrium from state R to state T. PMID- 10872056 TI - [Study of molecular structure of polyhydroxybutyrate-a termoplastic anddegradable biopolymer]. AB - The molecular structure of polyhydroxybutyrate from hydrogen-oxidizing bacteria Alcaligenes eutrophus was studied by X-ray diffraction analysis. It was shown that the degree of crystallinity of various samples depends little on the conditions of their preparation and is equal to 0.62-0.76. The molecular structure of solid samples and solution of polyhydroxybutyrate in chloroform were studied by the NMR and EPR methods. The conclusion is made that the molecular structure of polyhydroxybutyrate does not depend on the features of the strain and conditions of carbon nutrition of microorganisms producing polyhydroxybutyrate. Defects induced by gamma-radiation in polyhydroxybutyrate were studied. Free radicals were isolated, and their structure was decoded. PMID- 10872057 TI - [Regulation levels of photosynthesis processes]. AB - The basic mechanisms of kinetic regulation of photosynthetic processes are considered, which provide a strict light regulation of electron transfer in photosynthetic reaction centers and a more flexible regulation at the level of interaction of photosystems, transmembrane ion fluxes and coupling with dark reactions of the Calvin cycle. A generalized model was developed, which integrates the modern knowledge about photosynthetic processes of higher plants. The general principles of multilevel regulation in photosynthetic systems are discussed. PMID- 10872058 TI - [Relation between structural-dynamic organization of reaction centers in Rhodobacter sphaeroides and picosecond steps of photosynthesis]. AB - The effect of deuteration, and the addition of glycerol and dimethylsulfoxide on the redox midpoint potential Em of bacteriochlorophyll of the special pair ?PMPL?, the rate of energy migration from bacteriopheophytin HM to ?PMPL?, and electron transfer from ?PMPL? to HL and from HL to quinone QA in reaction centers of Rhodobacter sphaeroides was studied. It was shown that H2O-->D2O substitution did not change Em of the special pair, while the addition of 70% glycerol and 35% dimethylsulfoxide (v/v) increased the Em value by 30 and 45 mV, correspondingly. The rate constants of energy migration [formula: see text], charge separation [formula: see text], electron transfer to QA kQ remained unchanged upon the addition of glycerol. The isotopic substitution of water and addition of dimethylsulfoxide led to a 2-3-fold increase in km, ke and kQ values. The dependence of the potential of redox center on the dielectric constant epsilon was analyzed. It was shown that replacement of H2O by dimethylsulfoxide can increase Em by tens of millivolt. There was no correlation between changes in Em and the values of km, ke and kQ upon deuteration and addition of cryoprotectors. It was concluded that the processes of energy migration, charge separation, and electron transfer to the quinone acceptor are preceded by the solvation of states H*M, ?P+MP-L?* and [formula: see text]. PMID- 10872059 TI - [Possible states of the photosystem II reaction centerand thermoluminescence of higher plants. II. Application of the theory to experimental data analysis. Relation of thermoluminescence peaks to initial states of reaction center and excitation conditions]. AB - Initial conditions for the thermoluminescence theory described in the first part of the present work were determined. The initial populations of possible states of the photosystem II reaction center and thermoluminescence of higher plants for different excitation conditions were estimated. The estimation enables one to explain the known experimental conditions favoring the appearance of certain thermoluminescence peaks. The possibility of applying the theory to the deconvolution of experimental curves into components was shown. PMID- 10872060 TI - [Photoinduced reactivation of photosystem II in Chlorella after prolong incubation without light]. AB - The light-dependent reactivation of photosystem II in Chlorella pyrenoidosa Chick, CALU-175 cells, inactivated with supraoptimal temperatures (40-43 degrees C) in the dark or during heterotrophic growth was studied. It was shown that the inactivation of photosystem II after incubation in the dark at 41-42 degrees C, which showed up in the suppression of relative yield of variable chlorophyll fluorescence Fv due to an increase in yield F0 could be completely reversed by light. The inactivation of photosystem II at 43 degrees C in the dark could not be reversed by subsequent irradiation. In this case, the suppression of Fv/Fm was related not only to the growth of F0 but also with the decrease in Fm. The light dependences of the rate and extent of reactivation of yield Fv after heterotrophic growth or incubation of chlorella at 41 degrees C in the dark completely coincided. The full light-induced reactivation of photosystem II took place as the rate of photoinduced electron transport reached the rate of nonphotochemical reduction of plastoquinone in the dark. These results suggest that the light-reversed inactivation of photosystem II after heterotrophic growth or incubation at 41 degrees C in the dark is due to the redox-interaction of the primary quinone acceptor with plastoquinone reduced by the electron flux from the substrates of chlororespiration. PMID- 10872061 TI - [The use of fluorescent methods for measuring thr photosystem II activity in biomonitoring of phytoplankton]. AB - The regulation of activity of the photosystem II in microalgae under the influence of environmental factors was studied. Luminescent methods for biomonitoring the phytoplankton in situ are proposed. PMID- 10872062 TI - [The effect of electric field on the spatial-time patterns in the reaction diffusion system]. AB - A model of electrodiffusion processes in the vicinity of cell membrane was developed. The model takes into account chemical reactions, Coulomb interactions between charged particles and the effect of external electric field. It was concluded that the applied electric field can change the characteristics of space time patterns in the system. Dissipative structures slowly move and this is accompanied by a change in the number of structure elements. The characteristic equation includes odd powers of the wavenumber, which can lead to the appearance of soliton-like structures. The dissipative structures can appear not only due to the Turing diffusion instability but due to the disperse instability under electric field the applied. PMID- 10872063 TI - [The study of microviscosity of plasma membranes of Nitella cells during rest and excitation]. AB - Changes in the microviscosity of excitable membranes was investigated using resonance Raman spectroscopy of carotenoids. The Raman resonance spectra of carotenoids in Nitella cells were excited by 514.5 nm line of an argon ion laser. The bands at 1525 cm-1, 1160 cm-1 and 1008 cm-1 were observed and they were assigned to C=C, C-C and C-CH vibrations, respectively. The rhythmic excitation of cell reduced the intensity and increased the ratios of intensity of major carotenoid bands with no noticeable shift in the position of peaks. The Arrhenius plot of relative intensity ratios of 1525 cm-1 and 1160 cm-1 bands versus reciprocal temperature showed a change of the slope in the range of 13-18 degrees C. This indicates a membrane phase transitions in which a reorientation of carotenoids species takes place. The interpretation was supported by parallel microcalorimetric and EPR measurements. The decrease of microviscosity with increasing temperature is probably caused by changes in polyene chain conformation. It is suggested that membrane microviscosity during NH4(+) stimulated rhythmic excitation of algal cells increases, and membrane-associated carotenoids act as microviscosity-sensitive "potential sensor" for the channel. PMID- 10872064 TI - [Anaerobic formation of succinate and facilitation of its oxidation -- possible mechanisms of cell adaptation to oxygen deficiency]. AB - An important role of anaerobic formation of succinate in anoxic and hypoxic states and the activation of succinate oxidation under hypoxia were shown. It was concluded that, for maintaining the energetics of animal cells under conditions of oxygen deficiency, it is advisable to use substrates capable of participating in the anaerobic formation of succinate, whereas under hypoxia it is reasonable to use succinate itself. PMID- 10872065 TI - [Motor activity of paramecium]. AB - A systemic study of the mechanisms of motor activity of paramecium was carried out. The movements of paramecium responding to various influences were photographed. The analysis of the data revealed the time dependences of the rate of movement, rate of rotation, and the radius and the pitch of the helix trajectory. Mathematical models of the membrane and a unit that transforms the calcium signal to programs of regulating the effectors were constructed. A system of equations for constructing the trajectory of movement was proposed. It is concluded that the biomolecular system that involves calmodulin, calmodulin dependent ionic channels, adenylate cyclase, guanylate cyclase, phosphodiesterases, Ca(2+)-calmodulin, cAMP, cGMP-dependent protein kinases, and phosphoprotein phosphatases is capable of regulating motor reactions necessary for complex maneuvering of paramecium under various conditions. PMID- 10872066 TI - [Problems in estimating the dimensions of strange attractors in the analysis of biophysical data]. AB - Basic algorithms of calculating the dimensions of strange attractors from experimental data are considered. A special emphasis is placed on difficulties arising when the methods of solution of the reverse nonlinear dynamic problem are used in the analysis of the behavior of biological systems. These difficulties are associated with a poor convergence and weak stability of estimated values as well as a low degree of statistical confidence. Factors that hamper the estimation of strange attractor dimensions with reasonable accuracy are discussed by the example of analysis of human electrocardiograms. A method for the statistcal estimation of dimensions of attractors is proposed based on multidimensional imitational modeling. PMID- 10872067 TI - [The mechanism of formation of Liesegang structures around Dictyostelium discoideum]. AB - A theoretical study of the phenomenon of Liesegang structure formation induced by the Dictyostelium discoideum population in a medium containing folic acid was carried out. Using a "reaction-diffusion" model proposed in this work, it was shown that the formation of Liesegang structures around the Dictyostelium discoideum population depends on two competing processes: (a) inactivation of folic acid by vegetative amoebae and (b) the chemical reaction of folic acid with the products of amoeba metabolism, which results in the formation of insoluble sediment. The dependence of the model solutions on the geometric and functional parameters was studied. The results are in good agreement with experimental data. PMID- 10872068 TI - [Regulation of nitrate metabolism through anion polycompartmentation in plant roots]. AB - A new concept illustrated by a corresponding mathematical model of nitrate metabolism regulation is proposed. The model is based on root nitrate compartmentation in several functional pools: storage, metabolic and a mobile pool which is intended for translocation to shoots. Data on nitrate uptake, compartmentation, reduction in intact roots and translocation to shoots were obtained on steady-state wheat seedlings grown at 25 and 12 degrees C in the root zone. The net uptake, influx/efflux ratio, mobile pool size and translocation changed depending on the medium temperature. The oscillations of the net uptake rate, nitrate tissue concentration were revealed and the effect of temperature on these changes was demonstrated. The scheme of regulation is based on the idea that net uptake through nitrate influx/efflux is under the control of the nitrate the mobile pool whose size was dependent on the nitrate translocation into shoots. The mathematical model is represented by a system of ordinary differential equations simplified according to the time hierarchy of reactions. It has a limit cycle at definite values of the parameters. The model postulates the mechanism of a positive feed-back regulation of the transfer of newly absorbed nitrate into translocated pool formed in the root cortex. Theoretical results are verified experimentally. PMID- 10872069 TI - [The study of role of membrane-bound Ca2+ in the regulation of relationship between neuron and glia during rhythmic excitation]. AB - The role of membrane-bound Ca2+ in the regulation of Ca2+ transport through voltage-gated Ca2+ channel, and NMDA-glutamate and n-acetylcholine receptors upon interaction of a neuron with glia during rhythmic excitation was studied. It was found that the redistribution and transport of Ca2+ play a crucial role in the conductance of rhythmic excitation in both a "neuron-neuron" system and the processes providing the maintenance of a stationary level of rhythmic excitation in the system "neuron-glia". PMID- 10872070 TI - [A biophysical approach to the investigation of physiological processes]. AB - It was shown that understanding the mechanism of rhythmic excitation in cells and tissues requires the combination of physiological and biophysical approaches. Systemic studies of changes in the physicochemical characteristics of the object were carried out by a protocol that takes into account the mode of rhythmic excitation and the functional sate of the object being studied. The validity of the approach was proved in studies of rhythmic excitation in somatic nonmyelinic and myelinic nerves, and in model systems. The approach can be used in studies of many physiological processes. PMID- 10872071 TI - [A model of blood flow distribution during filtration-reabsorption processes in capillaries]. AB - A model of blood flow in a capillary was constructed, which takes into account the movement of plasma through its porous wall. The functions of changes in pressure and the rate of blood flow along the capillary were calculated. It was shown that, in the general case, the distribution of hemodynamic parameters as a result of filtration-reabsorption processes is nonlinear. Possible mechanism of tissue edema resulting from the disturbance of the filtration-reabsorption equilibrium were analyzed. PMID- 10872072 TI - [Changes in the asynchronism of transmitter release in a neuromuscular synapse and the time course of evoke postsynaptic signals during growth and branching og frog nerve ending]. AB - The influence of both growth and branching of a nerve terminal on the asynchronism of transmitter release and the time-course of evoked postsynaptic responses was investigated using a model of a frog neuromuscular synapse in which the nerve terminal represents a population of spatially isolated active zones. It was shown that the appearance of additional branching in proximal parts of the nerve ending leads to decrease in the asynchronism of transmitter release, an increase in quantum content and the amplitude of the postsynaptic signal, and the shortening of its phase of growth. It was found that the asynchronism of transmitter release has a much stronger influence on the time-course of end plate currents compared with end plate potentials. The factors strengthening and weakening the asynchronism of transmitter release in a neuromuscular synapse and the reasons for various length and branching of vertebrate nerve terminals are considered. PMID- 10872073 TI - [Molecular evolution more than three billion years ago]. AB - The earlier stages of life evolution at the molecular level were reconstructed. A set of triplets incorporated into the earliest RNA molecules (mRNA) was determined. It is believed that tRNA is one of the descedants of the earliest protein-encoding sequences. It was shown that tRNA has a hidden periodic structure of base sequences that conforms to the rule (GNY)n, indicating it ancient age. A set of the earliest amino acids was established and their distribution with respect to age was made. It was shown that genomes were built from elementary units (primary genes) by recombination or binding some time prior to division into eukaryotes and prokaryotes. Mean dimensions of the elementary unit for eukaryotes and prokaryotes were determined. PMID- 10872074 TI - [Phase transition in lipids and the problem of homoiothermia]. AB - Why such a high stability of temperature is necessary for warm-blooded animals? Why the range of body temperatures of various warm-blooded species is several fold narrower than the range of environmental temperatures? What were the physicochemical factors that determined such results of the biological evolution? A hypothesis presented in this short communication provides answers to these basic questions of the problem of homoiothermia. The hypothesis implies that the Ca(2+)-induced chain-ordering phase transition in the lipid component of synaptic membranes plays a key role at the last step of the mechanism of synaptic transmission, namely, the step of neurotransmitter release. The physicochemical substantiation of a possible molecular mechanism of the release involving the phase transition is presented, and the main kinetic and evolutionary issues of the mechanism are considered in brief. PMID- 10872075 TI - Bacterial metabolism of fluorene, dibenzofuran, dibenzothiophene, and carbazole. AB - Fluorene and its three heteroatomic analogs, dibenzofuran, dibenzothiophene, and carbazole, are environmental contaminants in areas impacted by spills of creosote. In addition, dibenzofuran has been used as an insecticide, and it is formed from the photolysis of chlorinated biphenyl ethers. Many biodegradation studies of dibenzofuran have considered it as a model for chlorinated dibenzofurans, which are of greater environmental concern. This paper reviews the bacterial degradation of fluorene and its analogs. These compounds are susceptible to three different modes of initial oxidation: (i) the naphthalene like attack, in which one of the aromatic rings is oxidized to a dihydrodiol; (ii) an angular dioxygenase attack, in which the carbon bonded to the methylene group in fluorene or to the heteroatoms in the analogs, and the adjacent carbon in the aromatic ring are both oxidized; and (iii) the five-membered ring attack, in which the methylene carbon atom in fluorene or the sulfur atom in dibenzothiophene is oxidized. The metabolites, enzymology, and genetics of these transformation are summarized. Literature data are presented, indicating that the electronegativity of the atom connecting the two aromatic rings influences the attack of the angular dioxygenase. In dibenzofuran and carbazole, the connecting atoms, O and N respectively, have high electronegativities, and these compounds serve as substrates for angular dioxygenases. In contrast, the connecting atoms in dibenzothiophene and fluorene, S and C respectively, have lower electronegativities, and these atoms must be oxidized before the angular dioxygenases attack these compounds. PMID- 10872076 TI - Segregation following interspecific transfer of isolated nuclei between Phytophthora parasitica and P. capsici. AB - Nuclei isolated from metalaxyl-resistant (MR) protoplasts of Phytophthora parasitica were transferred into chloroneb-resistant (CnR) protoplasts of Phytophthora capsici and vice versa, with an average success rate of 2.6 x 10(-4) (protoplasts with donor nuclei/regenerated protoplasts), using a selective medium containing only the fungicide tolerated by the nuclear donor. No colonies appeared when self-fusion products of donor nuclei or recipient protoplasts were exposed to the selective medium. Colonies produced by the nuclear transfer formed sectors commonly, and differed from the parental types in appearance. All the zoospores produced by the nuclear hybrids were of normal size, and one-fifth of them contained both MR and CnR genes. Since zoospores are mostly uninucleate, these results indicated the occurrence of chromosome re-assortment or mitotic crossing-over following the production of transitory tetraploids, followed by diploidization during zoosporogenesis, thus suggesting the completion of events leading to a parasexual cycle. Hyphal fragment cultures from a nuclear hybrid tested showed considerable variation in growth rate, mycelial morphology, and level of resistance to metalaxyl, indicating uneven distribution and continuous segregation of different types of nuclei in mycelia during vegetative growth. PMID- 10872077 TI - Detection of adenoviruses and enteroviruses in tap water and river water by reverse transcription multiplex PCR. AB - A reverse transcription (RT) multiplex polymerase chain reaction (PCR) assay was developed to simultaneously detect adenoviruses and enteroviruses, both of which have attracted much attention as molecular indices of viral pollution in environmental samples. The method involves a reverse transcription step, followed by a multiplex nested PCR in which the combination of primers amplifies cDNA from enteroviruses and adenoviruses. The sensitivity of this assay was found to be similar to that of each monoplex PCR or RT-PCR assay, and to be consistent regardless of relative concentrations of adenoviruses and enteroviruses. To assess suitability and environmental application of the RT multiplex PCR assay, a total of 12 river water samples and 4 tap water samples were analyzed by RT multiplex PCR, each monoplex PCR or RT-PCR, and cell culture assay on the Buffalo Green Monkey kidney cell line. The sensitivity of the RT multiplex PCR was also found to be similar to that of each monoplex PCR in environmental samples. This suggests the RT multiplex PCR assay could be applied to the routine monitoring of viral pollution in environmental waters. PMID- 10872078 TI - The potential of soil microorganisms to mineralize atrazine as predicted by MCH PCR followed by nested PCR. AB - The potential of soil microorganisms to mineralize atrazine was studied in soil samples collected from fields with various histories of atrazine application. In contrast to many previous studies, which showed no atrazine mineralization activity, all the tested soils mineralized atrazine regardless of their atrazine application history. However, the delay before mineralization and the variation in the subsequent mineralization rate were in agreement with the initial copy number of the atrazine dechlorinaze gene, and the proliferation rate of the degraders. Soils from corn fields, which had up to 100 copies of the atzA gene per gram of soil, had a lag period of 4-5 days before atrazine mineralization started, and final mineralization percentages ranged from 40% to 54%. However, soils from fields that were never amended with atrazine had much longer lag periods (more than 17 days), which decreased after enrichment of the degrader population with high concentrations of atrazine for 15 days. Generally the mineralization rate and the atzA gene copy number increased after the enrichment period. The atrazine mineralization potential was measured by PCR of genes from the atrazine mineralization pathway. Magnetic capture hybridization was the most efficient of the two tested methods for purifying target DNA of PCR inhibitors, without reducing the copy number of the required fragment. Nested PCR proved to be the most effective method for predicting the exact potential of the soil to mineralize the pollutant even without enrichment of a small population with the target genes. This method can complement microcosm studies and eliminate futile efforts when the potential to mineralize the pollutant does not exist in the soil. PMID- 10872079 TI - Monitoring by laser-flow-cytometry of the polycyclic aromatic hydrocarbon degrading Sphingomonas sp. strain 107 during biotreatment of a contaminated soil. AB - A flow cytometric method (FCM) was used to detect and accurately enumerate a polycyclic aromatic hydrocarbon-degrading bacterial strain, Sphingomonas sp. 107, inoculated into a soil sample artificially contaminated with pyrene. To compare the FCM method with colony forming unit (CFU) assays, a rifampicin-resistant Sphingomonas sp. 107 was obtained which could be distinguished from the indigenous microflora, since there was no organism resistant to rifampicin in the soil that could transform indole to indigo (naphthalene dioxygenase activity). By combining light-scattering profiles (i.e., morphological properties), ethidium bromide influx (i.e., cell wall permeability), and fluorescence in situ hybridization against the 16S rRNA (i.e., detection specificity), we could enumerate the bacterial population of interest from the indigenous microflora and soil debris during the biotreatment. The FCM technique revealed that the number of inoculated Sphingomonas cells decreased gradually for 15 days of incubation before reaching a steady level of 7 to 12 x 10(5) cells.g-1 of soil. Similar values were obtained with the CFU assay. During this period, pyrene concentration decreased from 632 to 26 mg.kg-1 of dry soil. The FCM detection was improved by adding blocking reagent to the hybridization buffer to minimize the non-specific attachment of the fluorescent probe to soil particles. Combined with the improvements in probe technology, FCM detection was shown to be a good alternative to the conventional culture methods for the analysis of bacterial populations in environmental samples. This technique could be potentially useful for the detection of microorganisms that grow poorly in culture. PMID- 10872080 TI - Community dynamics of a mixed-bacterial culture growing on petroleum hydrocarbons in batch culture. AB - The effects of various hydrocarbon substrates, and a chemical surfactant capable of enhancing crude-oil biodegradation, on the community structure of a mixed bacterial inoculum were examined in batch culture. Of 1000 TSA-culturable isolates, 68.6% were identified at the genus level or better by phospholipid fatty acid analysis over 7-day time course experiments. Cultures were exposed to 20 g/L Bow River crude oil with and without 0.625 g/L Igepal CO-630 (a nonylphenol ethoxylate surfactant), 5 g/L saturates, 5 g/L aromatics, or 125 g/L refinery sludge. A group of six genera dominated the cultures: Acinetobacter, Alcaligenes, Ochrobactrum, Pseudomonas/Flavimonas, Stenotrophomonas, and Yersinia. Species from four of the genera were shown to be capable of hydrocarbon degradation, and counts of hydrocarbon degrading and total heterotrophic bacteria over time were nearly identical. Pseudomonas/Flavimonas and Stenotrophomonas normally dominated during the early portions of cultures, although the lag phase of Stenotrophomonas appears to have been increased by surfactant addition. Acinetobacter calcoaceticus was the most frequently isolated microorganism during exposure to the saturate fraction of crude oil. Regardless of substrate, the culture medium supported a greater variety of organisms during the latter portions of cultures. Understanding the community structure and dynamics of mixed bacterial cultures involved in treatment of heterogeneous waste substrates may assist in process development and optimization studies. PMID- 10872081 TI - Effects of Pinus sylvestris root growth and mycorrhizosphere development on bacterial carbon source utilization and hydrocarbon oxidation in forest and petroleum-contaminated soils. AB - The hypothesis that Pinus sylvestris L. root and mycorrhizosphere development positively influences bacterial community-linked carbon source utilization, and drives a concomitant reduction in mineral oil levels in a petroleum hydrocarbon- (PHC-) contaminated soil was confirmed in a forest ecosystem-based phytoremediation simulation. Seedlings were grown for 9 months in large petri dish microcosms containing either forest humus or humus amended with cores of PHC contaminated soil. Except for increased root biomass in the humus/PHC treatment, there were no other significant treatment-related differences in plant growth and needle C and N status. Total cell and culturable bacterial (CFU) densities significantly increased in both rhizospheres and mycorrhizospheres that actively developed in the humus and PHC-contaminated soil. Mycorrhizospheres (mycorrhizas and extramatrical mycelium) supported the highest numbers of bacteria. Multivariate analyses of bacterial community carbon source utilization profiles (Biolog GN microplate) from different rhizosphere, mycorrhizosphere, and bulk soil compartments, involving principal component and correspondence analysis, highlighted three main niche-related groupings. The respective clusters identified contained bacterial communities from (i) unplanted bulk soils, (ii) planted bulk PHC and rhizospheres in PHC-contaminated soils, and (iii) planted bulk humus and rhizosphere/mycorrhizosphere-influenced humus, and mycorrhizosphere-influenced PHC contaminated soil. Correspondence analysis allowed further identification of amino acid preferences and increased carboxylic/organic acid preferences in rhizosphere and mycorrhizosphere compartments. Decreased levels of mineral oil (non-polar hydrocarbons) were detected in the PHC-contaminated soil colonized by pine roots and mycorrhizal fungi. These data further support our view that mycorrhizosphere development and function plays a central role in controlling associated bacterial communities and their degradative activities in lignin-rich forest humus and PHC-contaminated soils. PMID- 10872082 TI - Heterotrophic nitrification by a thermophilic Bacillus species as influenced by different culture conditions. AB - The nitrification activity of a thermophilic heterotrophic bacterium, Bacillus MS30 isolated from a deep-sea hydrothermal vent, was studied under various growth conditions. Nitrification was estimated from the nitrogen balance calculations in the culture media. The results showed that this isolate actively nitrified in culture conditions similar to those prevailing in hydrothermal sites. Therefore, its ecological significance was considered. In standard aerobic conditions, MS30 produced nitrite from ammonia and acetate (1.13 mumol NO2-.mg-1 dry wt), but nitrate was never produced, and a low nitrite reduction was often observed. Higher nitrification activities were observed in defined optimal conditions (simple carbon substrate, 65 degrees C, pH 7.5, and 15 g sea salts.L-1). In addition, discrepancies between the optima for growth and nitrification were observed, showing the ability of MS30 to adapt to changing environmental conditions typical of hydrothermal sites. PMID- 10872083 TI - Overproduction in Escherichia coli of the pectin methylesterase A from Erwinia chrysanthemi 3937: one-step purification, biochemical characterization, and production of polyclonal antibodies. AB - Pectin methylesterase A (EC 3.1.1.11), one of the pathogenicity factors of Erwinia chrysanthemi strain 3937, was purified to homogeneity using one-step chromatography on cross-linked pectate. The purified protein showed maximum activity at pH 8-9, 50 degrees C, 50-100 mM monovalent cations or 5-10 mM divalent cations, and on a 50% esterified pectin. A particular effect of Ca2+ and Zn2+ on PMEA activity, due to the formation of a pectin gel, was observed. A Km value of 0.03% and 0.051% was determined at pH 6 and 7.6, respectively, using the same substrate. Polyclonal antibodies raised against the PMEA from E. chrysanthemi strain 3937 were produced. It recognized PMEs from Erwinia species, but did not cross-react with PME of fungal or plant origin, and will therefore be a useful tool to immunolocalize the protein during plant-pathogen interactions. PMID- 10872084 TI - Arsenic and cadmium resistance in environmental isolates of Yersinia enterocolitica and Yersinia intermedia. AB - Environmental strains of Yersinia enterocolitica representing biotype 1A lack virulence plasmid (pYV) and are regarded as non-pathogenic. Though these occupy a diverse range of environmental niches, nothing is known about their resistance to heavy metals. The minimal inhibitory concentrations (MICs) of various metal ions, namely Ag+, Cu2+, Zn2+, Cd2+, As5+, and As3+, for strains of Yersinia enterocolitica (biotype 1A) and Yersinia intermedia (biotypes 1, 2, and 4), isolated from sewage effluents or pork, were determined. All isolates were resistant (MICs 2.5-5 mM) to Cd2+. The MICs of arsenic varied with bacterial strain and the chemical species of the arsenic used. For the majority of the strains, however, it was between 5-10 mM of Na2HAsO4.7H2O and NaAsO2, and 0.625 2.5 mM of As2O3. Except for one isolate, MICs of Ag+, Cu2+, and Zn2+ for these strains were in the range of 0.3-0.625 mM. PMID- 10872085 TI - Deletions of mob and tra pJP4 transfer functions after mating of Ralstonia eutropha JMP134 (pJP4) with Escherichia coli harboring F'::Tn10. AB - One-tenth of Escherichia coli transconjugants resulting from the transfer of the catabolic plasmid pJP4 from Ralstonia eutropha JMP134 to E. coli XL1Blue, contained pJP4 derivatives with deletions (approximately 15-30 kb). The occurrence of these deletions is probably associated with the presence of Tn10 in the recipient. DNA endonuclease restriction analysis of the pJP4 deletion derivatives showed the absence of SphI and EcoRI fragments previously reported to hybridize with IncP Tra DNA probes. Moreover, these pJP4 deletion derivatives are not able to self-transfer, nor are they able to be mobilized. Accordingly, these pJP4 deletion derivatives lack transfer functions. PMID- 10872086 TI - Evidence for the occurrence of nucleotide-activated oligosaccharides in Saccharomyces cerevisiae. AB - Recently, nucleotide-activated oligosaccharides have been found to be involved in the biosynthesis of certain glycoconjugates in archaeal and bacterial procaryotes. This paper describes the isolation and partial chemical characterization of nucleotide-activated oligosaccharides from the eucaryotic microbe Saccharomyces cerevisiae. We purified four different nucleotide-activated oligosaccharides from cell extracts of Saccharomyces cerevisiae. Three of the oligosaccharides were UDP, and one was TDP-activated. D-Glucose was the only carbohydrate constituent, except for one oligosaccharide, which also contained glucosamine. The chain length varied between two and four sugar residues. PMID- 10872087 TI - Osmotically controlled oral drug delivery. AB - It is advantageous to deliver some drugs with short half-life, and which are to be given frequently for chronic ailments, in the form of controlled-release (CR) formulations. The orally administered drugs, in the form of conventional matrix or reservoir type formulations, pose problems of bioavailability fluctuations due to gastric pH variations. Moreover, the release of drug(s) from these systems is affected by the hydrodynamic conditions of the body. Osmotically controlled drug delivery systems utilize the principles of osmotic pressure for the controlled delivery of active agent(s). The release rate of drug(s) from these systems is independent of the physiological factors of the gastrointestinal (GI) tract to a large extent. In the present review, theory underlying the delivery of drugs from osmotic systems is presented. Different types of oral osmotic systems, their advantages over conventional matrix and reservoir types of systems, and their applications are also discussed. Finally, some of the limitations, adverse effects, and patent and market status of these systems are reviewed. These systems form a major segment of drug delivery products. Because of their advantages and strong market potential, it appears that the future of osmotic systems in rate-controlled oral drug delivery is promising. PMID- 10872088 TI - Dermal delivery of ETH-615, a zwitterionic drug. AB - ETH-615 is an amphoteric drug that forms a water-insoluble zwitterion at intermediate pH values. Increasing the aqueous solubility of ETH-615 through cyclodextrin complexation did not enhance transdermal delivery of the drug from saturated aqueous solutions. However, increasing the lipophilicity of the drug through masking of the anionic group with a pro-moiety increased the dermal and transdermal delivery of the drug. Furthermore, masking the anionic group enhanced the chemical stability of the drug, resulting in significant improvement of the shelf life of the drug in both aqueous and nonaqueous solutions. PMID- 10872089 TI - Functionalization of hydrocolloids: principal component analysis applied to the study of correlations between parameters describing the consistency of hydrogels. AB - This work was part of a pure research project on the functionalization of three families of hydrocolloids: cellulose derivatives, carrageenates, and alginates. Principal component analysis (PCA), a powerful statistical method, was used to demonstrate the relations existing among these different parameters that describe the consistency of hydrogels and their spreadability. This approach therefore provides a basis for modeling hydrogel consistency. PCA also afforded a classification of hydrogels that demonstrated the remarkable adhesiveness of very stiff gels based on cellulose derivatives and sodium or potassium alginates. The corresponding semi-fluid gels and all the gels based on carrageenates and mixed sodium-calcium alginates, whatever their spreadability, were found to be very poorly adhesive. Generalized to all the many colloids currently marketed, this approach can be used to set up a databank for the formulation of mucoadhesive excipients. PMID- 10872090 TI - Application of similarity factor in development of controlled-release diltiazem tablet. AB - Controlled-release grade hydroxypropylmethylcellulose (HPMC) or xanthan gum (XG) and microcrystalline cellulose (MCC) were employed to prepare controlled-release diltiazem hydrochloride tablets. The similarity factor f2 was used for dissolution profile comparison using Herbesser 90 SR as a reference product. Drug release could be sustained in a predictable manner by modifying the content of HPMC or XG. Moreover, the drug release profiles of tablets prepared using these matrix materials were not affected by pH and agitation rate. The f2 values showed that only one batch of tablets (of diltiazem HCl, HPMC or XG, and MCC in proportions of 3.0:3.0:4.0) was considered similar to that of the reference product, with values above 50. The unbiased similarity factor f2* values were not much different from the f2 values, ascribing to a small dissolution variance of the test and reference products. The amount of HPMC or XG incorporated to produce tablets with the desired dissolution profile could be determined from the curves of f2 versus polymer content. Hence, the f2 values can be applied as screening and optimization tools during development of controlled-release preparations. PMID- 10872091 TI - Formulation of activated charcoal for per os administration to addicted subjects. AB - The objective of this work was to develop a galenic form of activated charcoal appropriate for the needs of clinical toxicology. To preserve the adsorption capacity of charcoal, we developed an extemporaneous preparation of activated charcoal intended for clinical toxicology. To improve the wettability of activated charcoal, we used densification by wet granulation. The presence of a viscosity agent is necessary to ensure the homogeneity of the suspension and its adhesiveness on gastric mucous membrane. Five formulations with different viscosity agents were prepared, and their adsorption capacity, wettability, viscosity, and adhesiveness were studied. PMID- 10872092 TI - A new in vitro/in vivo kinetic correlation method for nitrofurantoin matrix tablet formulations. AB - The kinetic distributions of in vitro percentage release and in vivo percentage urinary excretion rates of nitrofurantoin from matrix tablets were plotted using a kinetic program. In vitro release rates were determined using the USP paddle and half-change methods. Urinary excretion curves of the drug were characterized by means of the statistical moments. The individual linear correlations between each in vitro and in vivo kinetic distribution were established, and regression equations were calculated. The application results of the best correlations obtained were evaluated according to in vivo results. A reversed kinetic procedure was applied for transformation of the correlated kinetic values to the drug percentage release rates. The modified Langenbucher kinetic showed excellent linear correlation (r = .9985). The method that is proposed in this study, the kinetic correlation program, is simple, independent of time, and suggests that it is possible to use kinetic distributions in the in vitro/in vivo correlation. This study also suggests using kinetic correlation to investigate the suitability of the in vitro dissolution methods with the in vivo drug dissolution. PMID- 10872093 TI - Influence of tableting forces and lubricant concentration on the adhesion strength in complex layer tablets. AB - The strength of adhesion in complex two-layer tablets is assessed using statistical methods with respect to the applied tableting forces for the first layer and for applying the second layer on the first, as well as regarding the fraction of the lubricant. These results, obtained on a single-punch tablet press, are compared with the results for three-layer tablets produced on a rotary press at production scale. The strongest negative influence on adhesion strength was exerted by the amount of lubricant in the central layer. As expected, compression forces for central-layer tableting also had a negative effect, whereas the compression forces for complex layer tableting exerted a positive effect on layer adhesion. The validity of the derived model equation was proved by experiments: It was shown that the adhesion strength in complex layer tablets produced in production scale can be predicted from laboratory-scale experiments. This makes optimization of the formulation and parameter settings at an early stage of development possible. PMID- 10872094 TI - Transdermal absorption of L-dopa from a new system composed of two separate layers of L-dopa and hydrogel in rats. AB - To maintain the stability of L-dopa in hydrogel, a new system composed of two separate layers of L-dopa and hydrogel was developed. L-Dopa sheets were made by immersing L-dopa solution into wiper sheets and by lyophilizing them. Examination for stability of L-dopa in the L-dopa sheet revealed that its stability was maintained for at least 12 weeks, providing the sheet was kept at room temperature in a dark box. In a cutaneous absorption study of L-dopa in rats, an L-dopa sheet was attached to the shaved abdominal skin. A hydrogel composed of cutaneous absorption enhancers, water and ethanol, was spread on vinyl tape (hydrogel sheet), and this sheet was placed over the L-dopa sheet. L-Dopa that was administered transdermally effectively penetrated through the skin: The plasma level of L-dopa peaked at 30 min and remained high between 60 and 180 min after the cutaneous application. Our system, composed of two separated layers of L-dopa and hydrogel, enabled the stability of L-dopa to be maintained without losing transdermal absorption of L-dopa. PMID- 10872095 TI - Effect of diluents on tablet integrity and controlled drug release. AB - The objective of this study was to evaluate the effect of diluents and wax level on tablet integrity during heat treatment and dissolution for sustained-release formulations and the resultant effect on drug release. Dibasic calcium phosphate dihydrate (DCPD), microcrystalline cellulose (MCC), and lactose were evaluated for their effect on tablet integrity during drug dissolution and heat treatment in wax matrix formulations. A newly developed direct compression diluent, dibasic calcium phosphate anhydrous (DCPA), was also evaluated. Compritol 888 ATO was used as the wax matrix material, with phenylpropanolamine hydrochloride (PPA) as a model drug. Tablets were made by direct compression and then subjected to heat treatment at 80 degrees C for 30 min. The results showed that MCC, lactose, and DCPA could maintain tablets intact during heat treatment above the melting point of wax (70 degrees C-75 degrees C). However, DCPD tablets showed wax egress during the treatment. MCC tablets swelled and cracked during drug dissolution and resulted in quick release. DCPD and lactose tablets remained intact during dissolution and gave slower release than MCC tablets. DCPA tablets without heat treatment disintegrated very quickly and showed immediate release. In contrast, heat-treated DCPA tablets remained intact through the 24-hr dissolution test and only released about 80% PPA at 6 hr. In the investigation of wax level, DCPD was used as the diluent. The drug release rate decreased as the wax content increased from 15% to 81.25%. The dissolution data were best described by the Higuchi square-root-of-time model. Diluents showed various effects during heat treatment and drug dissolution. The integrity of the tablets was related to the drug release rate. Heat treatment retarded drug release if there was no wax egress. PMID- 10872096 TI - Optimization and development of a core-in-cup tablet for modulated release of theophylline in simulated gastrointestinal fluids. AB - A triple-layer core-in-cup tablet that can release theophylline in simulated gastrointestinal (GI) fluids at three distinct rates has been developed. The first layer is an immediate-release layer; the second layer is a sustained release layer; and the last layer is a boost layer, which was designed to coincide with a higher nocturnal dose of theophylline. The study consisted of two stages. The first stage optimized the sustained-release layer of the tablet to release theophylline over a period of 12 hr. Results from this stage indicated that 30% w/w acacia gum was the best polymer and concentration to use when compressed to a hardness of 50 N/m2. The second stage of the study involved the investigation of the final triple-layer core-in-cup tablet to release theophylline at three different rates in simulated GI fluids. The triple-layer modulated core-in-cup tablet successfully released drug in simulated fluids at an initial rate of 40 mg/min, followed by a rate of 0.4085 mg/min, in simulated gastric fluid TS, 0.1860 mg/min in simulated intestinal fluid TS, and finally by a boosted rate of 0.6952 mg/min. PMID- 10872097 TI - Studies of ion-exchange resin complex of chloroquine phosphate. AB - High-potency adsorbates of chloroquine phosphate (CQP) were prepared by the batch method using a polyacrylic acid ion-exchange resin. Taste evaluation of the adsorbates shows significant masking of the bitterness of the drug. The complex formation was complete at pH 6.0. Stability studies at 37 degrees C, 45 degrees C, and 60 degrees C indicated that the complex was stable at all conditions for 1 month. In vitro release studies revealed complete drug elution from the complex at pH 1.2 and 2.0. PMID- 10872098 TI - Stability of extemporaneous norfloxacin suspension. AB - The stability of norfloxacin as extemporaneous suspensions compounded from two brands of film-coated tablets (formulas I and II) was studied. The vehicle consisted of tragacanth, saccharin sodium, sorbitol solution, glycerin, paraben concentrate, peppermint spirit BP, purified water, and syrup USP. The final concentration of norfloxacin in the suspensions was 20 mg/ml. Formulas I and II were chemically stable for 28 days when stored in amber glass bottles at ambient temperature; however, their physical characteristics were different. PMID- 10872099 TI - Preformulation investigation of the novel proton pump inhibitor lansoprazole. AB - Some technologically important physicochemical properties of lansoprazole were investigated. This compound is very unstable, especially in aqueous solutions with low pH. It has one acidic and two basic dissociation constants. Lansoprazole has relatively high solubility in solutions with high pH and is well partitioned from aqueous to n-octanol phase. Under conditions examined in this study, lansoprazole was not hydroscopic and did not decompose at higher relative humidities. PMID- 10872100 TI - Dissolution of omeprazole from delayed-release solid oral dosage forms. AB - The evaluation of the biopharmaceutical quality of omeprazole enteric-coated products (granules in capsules) with respect to its dissolution characteristics is not specifically regulated in any of the most common official pharmacopoeia. USP 23 includes a general monograph for enteric-coated products. This paper reports the evaluation of the medium pH effect on the dissolution rates of omeprazole from four omeprazole-containing products of different manufacturers. It is concluded that the USP 23 recommended dissolution procedure for enteric coated products is not suitable due to the degradation of omeprazole under such conditions. Furthermore, the medium with pH 8.0 showed different dissolution rates not observed at pH 7.4, allowing discrimination between products of different manufacturers. PMID- 10872101 TI - Formulation and release behavior of diclofenac sodium in Compritol 888 matrix beads encapsulated in alginate. AB - Sustained-release polymer beads containing diclofenac sodium (DNa) dispersed in Compritol 888 and encapsulated in calcium alginate shell were prepared utilizing 2(3) factorial design. The effect of sodium alginate concentration, drug: Compritol 888 weight ratio and CaCl2 concentration on drug content (%), time for 50% and 80% of the drug to be released, and mean dissolution time (MDT) were evaluated with analysis of variance (ANOVA). An increase in the level of all these factors caused retardation in the release, and t50%, t80%, and MDT were increased. The drug release was dependent on the pH of the release media. A formula that gives a release comparable to commercial products was prepared. PMID- 10872102 TI - A study of the spread surface of polymeric dispersions of Eudragit. AB - The rheology of non-Newtonian fluid systems is complex. Their experimental study is difficult because of the existence of many dependent variables. In the present work, a mathematical equation is proposed to calculate the spread surface from a simple viscosimetric study in a theoretical way. It was determined for multiple parameters. The spread surface has to be marked because of its direct relation to the shear stress; this fact enabled us to connect one variable dependent on the compression deformation with another dependent on the shear deformation. At the same time, the viscoelastic phenomenon can be evidenced by applying this mathematical equation. PMID- 10872104 TI - Childhood stroke--beyond re-inventing the wheel. PMID- 10872103 TI - Comparative bioavailability study of two controlled-release pentoxifylline tablet preparations. AB - The bioavailability of a generic preparation of pentoxifylline sustained-release (SR) tablet was evaluated in comparison with a proprietary product (Trental 400). For the study, 12 healthy male volunteers participated; the study was conducted according to a randomized, two-way crossover design. The bioavailability was compared using the parameters total area under the plasma level-time curve AUC0 infinity, peak plasma concentration Cmax, and time to reach peak plasma concentration Tmax. No statistically significant difference was observed between the values of the two products in all three parameters. The 90% confidence interval for the ratio of the logarithmic transformed AUC0-infinity values of the generic pentoxifylline over those of Trental 400 was found to lie between 0.83 and 1.00, while that of the parameter Cmax was between 0.91 and 1.29. In addition, elimination half-life t1/2 and apparent volume of distribution Vd were calculated. There was no statistically significant difference between the t1/2 Vd values obtained from the data of the two preparations. PMID- 10872105 TI - Brain magnetic resonance imaging abnormalities in merosin-positive congenital muscular dystrophy. AB - We report the brain magnetic resonance imaging (MRI) findings in 23 patients with merosin-positive congenital muscular dystrophy (CMD). Twelve patients had normal scans. Eight other children had essentially normal scans but showed mild non specific periventricular white matter changes. Three children had structural abnormalities on imaging. The first patient, a 15-month-old boy with hypotonia, muscle weakness and global development delay, had moderate cerebellar atrophy and mild dilatation of the lateral ventricles. The second child, a 3-year-old ambulant girl with subtle learning problems, had mild cerebellar hypoplasia and a large subarachnoid space when scanned at 16 months. The third patient, a 15-year old ambulant male with normal intelligence and complex partial seizures, had polymicrogyria of both temporoparietal lobes on brain MRI. The clinical features and motor ability of children with merosin-positive CMD are variable, although usually milder than merosin-deficient CMD. Our findings confirm that central nervous system involvement can occur in some merosin-positive cases. We suggest performing brain MRI in children with merosin-positive CMD, as this may help in our understanding of this very heterogeneous disease. PMID- 10872106 TI - Outcome of thiamine treatment in a child with Leigh disease due to thiamine responsive pyruvate dehydrogenase deficiency. AB - We describe a child with severe psychomotor retardation, peripheral neuropathy and bilateral abnormal signal in basal ganglia on magnetic resonance imaging, consistent with Leigh disease. Fibroblast pyruvate dehydrogenase assayed with routine method was normal. However, because of neurological improvement after treatment with thiamine, pyruvate dehydrogenase activity was studied again with thiamine pyrophosphate concentration adjusted to the normal human tissue level and found to be deficient. We report here on diagnostic difficulties and clinical follow-up of this patient. PMID- 10872107 TI - An established case of dentatorubral pallidoluysian atrophy (DRPLA) with unusual features on muscle biopsy. AB - Dentatorubral pallidoluysian atrophy (DRPLA) belongs to the group of autosomal dominant ataxias. Central nervous system pathology and inheritance are both well characterized, although the illness is rare. The presentation of a European child affected by this illness is described. He presented at 9 years of age with intractable progressive myoclonus epilepsy against a background of learning difficulties and developed progressive hypertonicity and dementia before his death at 15 years of age. Significant histological changes in a muscle biopsy were found. There was an absence of type IIB fibres and a predominance of type I fibres. Mean fibre diameter of all the fibre types was markedly reduced. All type I fibres showed an increase in lipid droplets. No previous descriptions exist of muscle histology in DRPLA. Although at least five adult family members have symptoms consistent with a diagnosis of DRPLA, their condition had not been recognized. We therefore describe the clinical picture and histological findings. PMID- 10872108 TI - An unusual complication of tapping a ventriculoperitoneal shunt. AB - A case is reported describing a complication of an unsuccessful attempt to aspirate the reservoir of a ventriculoperitoneal shunt system with a suspected shunt infection. This arose due to a misunderstanding of the anatomy of the shunt and resulted in an intracerebral haematoma. The complications of cerebrospinal fluid shunting and the difficulty in the diagnosis thereof are outlined. We discuss the role and method of shunt tapping in diagnosing shunt problems before reviewing the literature describing the rationale. The variation in shunt design is emphasized. Guidelines are then proposed not to dissuade physicians from tapping shunts but to ensure that the procedure is performed safely and in collaboration with neurosurgical units. PMID- 10872109 TI - Menkes kinky hair disease: an unusual case. AB - Menkes disease is a rare X-linked recessive disease of copper metabolism. Clinical manifestations begin in the first few months of life or even in the neonatal period. Hypothermia, hypotonia, poor weight gain, seizures and neurodevelopmental delay or regression are seen. Outcome is poor, with death occurring usually by 3 years of age. A characteristic facial appearance with steely hair suggest the diagnosis. Neuroimaging usually shows cortical atrophy, extra-axial fluid collections and progressive and extensive degeneration of grey matter with secondary demyelination. We describe an atypical, but biochemically proven case of Menkes disease with atypical clinical and radiological features. Our patient had a large head, atypical electron microscopy appearances of the hair and predominant diffuse white matter involvement on neuroimaging, but a low serum copper level and a high 64CU uptake in fibroblasts (89.5 ng/mg of protein) confirmed the diagnosis. PMID- 10872110 TI - Gene table: Inherited retinal diseases in humans. PMID- 10872111 TI - Orphan drugs and orphan diseases. Round table proceedings, 3rd European Pediatric Neurology Society Congress. Nice, November 1999. PMID- 10872112 TI - [Is there new information on necrotizing otitis externa?]. PMID- 10872113 TI - [Otitis media--a hereditary disease?]. PMID- 10872114 TI - [Kinetoses]. AB - Motion sickness is a well known nausea and vomiting syndrome whose physical signs occur during travel by sea, automobile, airplane, and space. This review describes current concepts concerning the aetiology, nature and therapy of this common phenomenon. Motion sickness involve a neural mismatch or confusion between the vestibular, visual, and proprioceptive systems. Therapy is directed towards decreasing conflicting sensory input, controlling nausea, and speeding the process of adaptation. PMID- 10872115 TI - [Disorders of auditory processing and perception. Consensus statement]. PMID- 10872116 TI - [Cranialization of the frontal sinus. Indications, technique and results]. AB - The osteoplastic frontal sinus surgery with obliteration of the sinus has been established in the therapy of frontal sinus diseases that can not be drainaged permanently or healed through an endonasal access. The obliteration of the frontal sinus is endangered in cases of multiple fracturing of the posterior frontal sinus wall or if it has been destroyed by an inflammatory process. In these problematic cases obliteration bears the danger of complications and cranialization of the frontal sinus is therefore the method of choice. We review 8 patients who were operated on using the cranialization technique. Indications for surgery were a combined fracture of the anterior and posterior frontal sinus wall (3), a pyocele of the frontal sinus with extensive destruction of the posterior wall (4) and a large osteoma of the posterior frontal sinus wall (1). The frontal sinus was exposed through a coronal incision, the mucosa and the posterior wall were completely removed and the frontal sinus obliterated with fat tissue. The anterior sinus wall was replaced after obliteration of the sinus or reconstructed with calvarian bone transplants. The follow up period was 1.8 years (11 months to 8 years). All patients underwent postoperatively a clinical ENT examination and radiological assessment by CT-Scan or MRI. The overall functional and esthetic outcome was excellent. There were no serious complications nor any recurrence. The cranialization of the frontal sinus is a reliable and safe variation of the classical osteoplastic frontal sinus surgery with fat obliteration. PMID- 10872117 TI - [Detection of numerical chromosome aberrations in leukoplakia and squamous epithelial carcinomas of the head-neck area using fluorescence in situ hybridization]. AB - The tumorigenesis of head and neck squamous cell carcinoma (HNSCC) has been proposed to represent a multistep process characterized by an accumulation of genetic alterations. To study numerical chromosomal aberrations and chromosomal imbalances, biopsies of 11 malignant tumours and biopsies of 16 oral premalignant lesions (leukoplakias) were analyzed by fluorescence in situ hybridization (FISH) using centromeric probes for chromosomes, 1, 7, 9, 10 and 17. The comparison of the alterations observed in simple leukoplakias (group 1, n = 8), dysplastic leukoplakias (group 2, n = 8) and malignant tumours (group 3, n = 11) by the Cochran-Armitage Trend Test revealed an increasing number of numerical chromosomal abberations. This difference was statistically highly significant (p < 0.001). The data open up the possibility that FISH analysis might help to better characterize the progression of premalignant oral leukoplakias. PMID- 10872118 TI - [Effect of microneurosurgical operation in acoustic neurinoma on symptoms of vertigo and tinnitus]. AB - After exstirpation of an acoustic neuroma one trends to concentrated on the preservation of hearing and facial function. But how does the surgical removal for an acoustic neuroma, influence the symptoms of tinnitus and vertigo? Our report follows a retrospective evaluation of 78 patients. Based on these results we will also discuss aspects of the origin of these symptoms. Our patients suffered from tinnitus at a rate of 73% preoperatively and 59% postoperatively. With increasing tumor size tinnitus occurred less often, therefore we considered the size of the tumor exclusively. Also, an intraoperative dissection of the cochlear nerve often did not result in an improvement of the symptoms. Vertigo was stated prooperatively in 44% of the cases, postoperatively in 22%. Again as with tinnitus, there was an inverse relationship between the size of the tumor and the severity of the symptoms. In conclusion a deafferention-like syndrome has to be considered as the major causative factor for the tinnitus in patients with acoustic neuromas. However, in the vertigo cases, the symptoms seem to be caused vestibularily due to the increasing tumor and then after a temporary compensation a cerebellar ataxia occurred which the patients often perceive as vertigo. PMID- 10872119 TI - [Regeneration after delayed nerve suture]. AB - The influence of the time delay of a nerve suture on axonal regeneration after nerve lesion is still unknown. We used the rat as an animal model and studied the influence of a 14- and 224-day delayed hypoglossal-facial nerve anastomosis (HFA) and compared results to immediate HFA. After injection of horseradish peroxidase into the whiskerpad we counted the retrogradely labelled neurons in the hypoglossal nucleus 7-112 days after operation to quantify the axonal reinnervation. Additionally, the amplitude of the evoked compound action potential of the whiskerpad was measured after hypoglossal nerve stimulation. By 112 days after immediate HFA only 931 +/- 27 hypoglossal neurons reinnervated the muscle (for 75% of normal innervation). After 14 days delayed HFA reinnervation was accelerated and enhanced with 1293 +/- 81 (104%) neurons. Even after 224 days delayed HFA reinnervation was possible, with 760 +/- 80 neurons (61%) present. We conclude that short-term delayed nerve suture improves axonal reinnervation. PMID- 10872120 TI - [Physiological effects of destruction of the tip links of cochlear hair cells. Significance for noise-induced hearing loss]. AB - Sound overexposure is known to cause damage to cochlear structures and can induce permanent or temporary hearing loss and tinnitus. Perhaps the most sensitive of these structures to sound overexposure are the tip links. In this paper the electrophysiological effects of pharmacological destruction of the tip links of outer hair cells was investigated. Outer hair cells treated with elastase (20 U/mL) or BAPTA (5 mM) no longer responded to sinusoidal stimuli. In contrast to common belief, transduction channels opened due to loss of tip links. Such opened channels can allow K+ and Ca2+ to enter the cell from the endolymphatic space and cold lead to permanent depolarization. This influx of cations caused by loss of tip links, together with the subsequent hair-cell depolarization, might be a source of sensorineural hearing loss and tinnitus associated with acoustic trauma. PMID- 10872121 TI - [Comparison between the old and new evaluation mode in the dichotic discrimination test]. AB - The dichotic discrimination test for children could make an important contribution to the diagnosis of central auditory processing disorders. With previous test procedures and their interpretation it was not possible to get the auditory discrimination ability in only one grade. To evaluate this ability, it was necessary to measure the degree of comprehension in each ear as well as the required noise level. The evaluation cannot be used for statistical results because there are too many parameters. Moreover, results apparently similar can come from varying abilities. We are therefore working on a new method to evaluate dichotic hearing more accurately. By using a constant noise level and only evaluating positively those hearing pairs which have been repeated correctly, we arrive at one single grade. The dichotic discrimination test was administered to 46 children with auditory perception disorders and the results evaluated according to both, the previous and new evaluation system. According to the new evaluation, a maximum of 20 correct responses is possible. The average number of correct responses was significantly lower than by the previous evaluation system. The standard deviation increased from 16% on the left side and 13% in the right to 24%, or five word-pairs. The difference is even clearer when the test is subdivided into four performance ranges, each of 25%. Of all the children examined, 85% fell in the upper performance range according to the previous evaluation. No child was in the lowest performance range. According to the new evaluation, only 56.5% of the children were in the upper performance range, while three children, that is 6.5%, were in the lowest performance range. This difference, P = 0.0002, is highly significant. Our results demonstrate clearly that on the basis of the new evaluation system the measurement of dichotic ability is possible, for the first time. We consider the new evaluation method to be an improvement in determining dichotic discrimination and in comparing it with other abilities. To check the validity, further independent testing should be carried out and results compared with those obtained from children without auditory perception disorders. PMID- 10872122 TI - [Carcinoid of the middle ear. Further therapy after primary tumor excision?]. AB - The carcinoid tumor of the middle ear is a very rare neoplasm which in general is regarded as benign but may be mistaken for an adenocarcinoma because of its histological heterogeneity. Typical, however, is its neuroendocrine and mucinous differentiation so that an unequivocal diagnosis is possible by means of immunohistochemistry and electron microscopy. As the tumor is very rare, there is no statistical evidence as to whether further treatment is necessary after primary exstirpation of the tumor. Therefore, a review of the literature was performed. We report about a 28-year-old male patient with a carcinoid tumor of the left tympanic cavity. Without any further treatment there has been no evidence for either recurrence or metastases 32 months after surgical resection. As treatment of choice we recommend conservative surgery with complete removal of the tumor and a clinical follow-up on a regular basis. PMID- 10872123 TI - [Mouth floor cyst with increasing dyspnea. Thyroglossal duct cyst in the area of the mouth floor]. PMID- 10872124 TI - [Emergency management in general ENT practice. 2: Special emergencies A]. PMID- 10872125 TI - [B-image ultrasound. 2: Technical principles]. PMID- 10872126 TI - The effects of positive end-expiratory pressure on the splanchnic circulation. PMID- 10872127 TI - The inflammatory balance in human sepsis. PMID- 10872128 TI - Effect of positive end-expiratory pressure on splanchnic perfusion in acute lung injury. AB - OBJECTIVE: To evaluate the acute effects of an increased positive end-expiratory pressure (PEEP) on splanchnic tissue perfusion. DESIGN: Clinical prospective study. SETTING: Intensive care unit in a university clinic. PATIENTS: Six patients with severe acute lung injury (ALI) requiring mechanical ventilation. All patients had bilateral infiltrates in chest X-ray, PaO2/FiO2 < 200 mmHg and stable hemodynamics without vasoactive drugs. INTERVENTIONS: PEEP was increased by 5 cmH2O from a clinically selected PEEP level (8/6-11 cmH2O) up to (13/10-14 cmH2O) followed by a return to baseline. MEASUREMENTS AND MAIN RESULTS: Splanchnic blood flow was measured using primed continuous infusion of indocyanine green dye with hepatic venous sampling and systemic hemodynamics by routine monitoring. In addition, we estimated gastric mucosal-arterial PCO2 difference and splanchnic lactate/pyruvate exchange. After a baseline measurement, PEEP was increased. After 60 min all measurements were repeated. PEEP was returned to the baseline level and a third measurement followed. PEEP had no effect on cardiac index (baseline I: 3.2/6.1-2.5 l/min/m2; PEEP: 3.3/5.7 2.3 l/min/m2; baseline II: 3.4/6.0-2.5 l/min/m2); neither did PEEP have any effect on splanchnic blood flow (baseline I: 0.91/1.39-0.62 l/min/m2; PEEP: 1.04/1.75-0.54 l/min/m2; baseline II: 1.07/1.42-0.68 l/min/m2, respectively) or perfusion (gastric mucosal-arterial PCO2 difference baseline I: 2.1/12.8-0.6 kPa; PEEP: 1.7/14.5-0.7 kPa; baseline II: 1.7/8.8-0.1 kPa; lactate uptake baseline I: 0.5/1.1-0.3 mmol/min/m2; PEEP: 0.4/1.0-0.3 mmol/min/m2; baseline II: 0.5/0.9-0.3 mmol/min/m2; hepatic venous lactate/pyruvate baseline I: 9.7/10.6-5.7; PEEP: 9.7/14.2-6.4; baseline II: 8.4/12.4-7.3; respectively). CONCLUSION: PEEP by itself does not have a consistent effect on splanchnic blood flow and metabolism when cardiac index is stable and patients are ventilated within the linear part of the pv curve. PMID- 10872129 TI - Non-invasive ventilatory approach to treatment of acute respiratory failure in neuromuscular disorders. A comparison with endotracheal intubation. AB - OBJECTIVE: Prospectively to investigate the efficacy of non-invasive positive pressure ventilation (NPPV) combined with cricothyroid "mini-tracheostomy" (CM) as a first-line intervention in patients with acute respiratory failure (ARF) of neuromuscular origin, in comparison with positive pressure ventilation (PPV) via endotracheal intubation (ETI). DESIGN: Prospective analysis of the short-term outcomes of 14 non-consecutive patients suffering from ARF of neuromuscular origin who were administered NPPV and comparison with the outcomes of 14 matched historical control patients receiving conventional mechanical ventilation (MV) via ETI. SETTING: Adult five-bedded respiratory intensive care unit in a university hospital. PATIENTS AND INTERVENTIONS: Fourteen neuromyopathic patients who developed hypercapnic ARF and were submitted to NPPV (group A) and fourteen matched historical control patients, who were administered PPV via ETI (group B). Seven subjects receiving NPPV also underwent CM. OUTCOME MEASURES: Mortality during ICU stay and treatment failure were evaluated; treatment failure was defined as death or the need for ETI for the NPPV group and as death or the inability to wean from MV for the control group. Length of stay in the ICU and time to improvement, defined as the time required for a significant relief of dyspnea and neurologic impairment and for correction of arterial blood gases, were also compared. RESULTS: Intra-hospital mortality and treatment failure were lower in the NPPV group than in the conventional PPV via ETI group (2 vs 8 cases and 4 vs 11 cases, respectively). In addition, the duration of ICU stay for subjects who underwent NPPV was shorter than for patients who were intubated (13.6 +/- 9.7 vs 47.1 +/- 51.9 days). "Mini-tracheostomy" was well tolerated and no significant side effects were encountered. Two patient were excluded from the study because they showed a severe inability to swallow and needed to be intubated to protect the upper airway from the risk of aspiration. CONCLUSIONS: Non-invasive positive pressure ventilation in combination with CM may be considered as a safer and more effective alternative to ETI in the treatment of patients with neuromuscular disorders (NMD) who develop ARF and require MV; nevertheless, patient selection remains important, since a significant proportion of neuromyopathic patients might have to be excluded from NPPV because of severe risk of aspiration. PMID- 10872130 TI - Nasal, pulmonary and autoinhaled nitric oxide at rest and during moderate exercise. AB - OBJECTIVE: To investigate nasal nitric oxide (NO) excretion, pulmonary NO excretion, and autoinhalation of nasally released NO at rest compared with that during moderate exercise in smokers and non-smokers. DESIGN: Prospective observational study. SETTING: University laboratory. PARTICIPANTS: Fourteen healthy adult volunteers. INTERVENTIONS: Breathing of NO-purified air supplied via a tube system at rest and during a bicycle-ergometer workload of 60 Watt over a time of 10 min. MEASUREMENT AND RESULTS: We examined nasal and pulmonary NO excretion in smoking (n = 7) and non-smoking (n = 7) adult human volunteers. At rest, we measured constant nasal NO excretion rates of 311 +/- 89 nl/min for non smokers and 261 +/- 142 nl/min for smokers (mean +/- SD, n.s.). During 60 W exercise, nasal NO release remained unchanged, while pulmonary NO excretion doubled compared with the rates at rest (non-smokers: 40 +/- 21 nl/min versus 23 +/- 14 nl/min, p < 0.05; smokers: 41 +/- 8 nl/min versus 22 +/- 8 nl/min, p < 0.05). The differences between smokers and non-smokers in nasal or pulmonary NO excretion were not significant. To determine the autoinhaled amount of nasally released NO, we also measured the NO concentration within the nasopharynx of five volunteers during nasal breathing. The average inhaled NO concentration was 17.8 +/- 3.1 ppb at rest and this decreased to 9.3 +/- 1.8 ppb during exercise of 60 W, while minute ventilation approximately doubled from 9 +/- 2 to 21 +/- 3 l/min. CONCLUSION: Our results demonstrate that moderate exercise increased exclusively pulmonary NO excretion. Nasal NO release, which is 10 times higher at rest, was not changed. The decrease in autoinhaled NO concentration during exercise results from dilution of the continuous nasal release by the increased respiratory gas flow. The individual NO release allows no conclusion about smoking habits. PMID- 10872131 TI - Acute renal failure in patients over 80 years old: 25-years' experience. AB - OBJECTIVE: To determine the epidemiological trends, spectrum of etiologies, morbidity and mortality of acute renal failure (ARF) in patients over 80 years old. DESIGN: Historical cohort analysis. SETTING: Intensive care unit (ICU) of nephrology, Tenon Hospital, Paris. PATIENTS AND PARTICIPANTS: The criteria of inclusion was ARF, defined on the basis of a creatinine value over 120 mumol/l, in patients over 80 years of age admitted between October 1971 and September 1996. When moderate chronic nephropathy was pre-existing, ARF was defined by the increase of at least 50% over the basal creatininemia. MEASUREMENTS AND RESULTS: Three hundred and eighty-one patients over 80 years of age were included. The etiology and mechanism of ARF are detailed. 29% of the patients received dialysis. Global mortality at the hospital was 40%. Factors significantly associated with a poor prognosis are identified. Mean survival after hospitalization was 19 months. CONCLUSION: The frequency of admission to ICUs for ARF in patients older than 80 years seems to be on the increase. Mortality is less severe than expected. These patients could benefit from the renal replacement therapy of modern intensive care medicine. PMID- 10872132 TI - Bioethical issues related to continuous renal replacement therapy in intensive care patients. AB - OBJECTIVE: To examine the ethical approach of intensivists and nephrologists to continuous renal replacement therapy (CRRT). DESIGN: A questionnaire. SETTING: The First International Course on Critical Care Nephrology. PARTICIPANTS: The participants in the course (around 500). RESULTS: Most participants think that establishing ethical criteria for managing CRRT is a medical task, as clinicians have adequate criteria for defining futility. However, many responders would grant the request of starting futile CRRT or would maintain it if requested by the family. Only 55% believe that informed consent is necessary for initiating CRRT; one out of four would start or maintain unwanted life-saving CRRT. In case of lack of equipment, the majority would select the patients, excluding the worst one or on a "first-come, first-served" basis. Withholding and withdrawing are regarded differently by most responders. Again, most think that every vital support should be withdrawn when futile, but practical and psychological aspects still influence the final decision. Responders think that ethics critical care committees can help in the management of ethical problems in ICU. CONCLUSIONS: Our results show that several ethical questions are still unsolved and that practical and psychological aspects of the treatment process can be stronger than bioethical principles. PMID- 10872133 TI - Clostridium difficile-associated diseases. The clinical courses of 18 fatal cases. AB - OBJECTIVE: Severe cases of Clostridium difficile-associated diseases with sepsis seem to be rare, as are case reports about the pathogen involved and sepsis. Our objective was to investigate the frequency and the clinical courses of severe cases of C. difficile-associated diseases with a fatal outcome in our hospital. SETTING: Teaching hospital of the University of Kiel (650 beds). DESIGN: We reviewed retrospectively all deceased patients' charts who had prior histological or microbiological evidence of C. difficile-associated diarrhea (CDAD) and revised the available histological specimens of the autopsies. PATIENTS: Over a 4 year period (November 1994-October 1998) we diagnosed 304 cases of C. difficile associated diseases in our hospital. RESULTS: Eighteen of our cases with C. difficile-associated diseases had a fatal outcome. C. difficile was not likely to be the cause of death in two of the cases. Four of the fatal infections were community-acquired and the reason for admission to the hospital. CDAD is most prevalent in elderly patients with multiple or severe underlying diseases and tends to be overlooked. Sepsis was diagnosed in 15 of our 18 patients with C. difficile-associated diseases. CONCLUSION: Our study shows that severe cases of CDAD or cases with C. difficile-associated sepsis are probably not rare. Routine testing of fecal specimens for the presence of C. difficile toxins should be considered not only in nosocomial gastrointestinal infections but also in community-acquired gastrointestinal infections of elderly people. PMID- 10872134 TI - Atropine test and circulatory arrest in the fossa posterior assessed by transcranial Doppler. AB - OBJECTIVE: To evaluate whether a negative atropine test (i.e., increase in heart rate of less than 3% after intravenous administration of 3 mg atropine) correctly predicts circulatory arrest in the fossa posterior during craniocaudal herniation in patients with primary supratentorial lesions. MATERIAL AND METHODS: Prospective, observational clinical study. SETTING: Two surgical intensive care units in a university hospital. PATIENTS: In 45 consecutive patients with suspected brain death, an atropine test (AT) and a transcranial Doppler sonography were performed simultaneously and, if necessary, repeatedly. MEASUREMENTS AND RESULTS: Forty-four patients fulfilled the typical criteria of a supratentorial and infratentorial circulatory arrest as the atropine test became negative. In one patient, who had undergone a decompressive craniectomy for uncontrollable intracranial pressure 4 h prior to the AT testing, we found a negative AT in the presence of an antegrade supratentorial and infratentorial flow. CONCLUSION: A negative atropine test indicates a circulatory arrest in the fossa posterior in patients with primary supratentorial lesions and craniocaudal herniation. In patients with brain-stem lesions, however, a negative atropine test does not unequivocally indicate a circulatory arrest. PMID- 10872135 TI - Sleep-related breathing disorders following discharge from intensive care. AB - OBJECTIVES: To determine the incidence of sleep-related breathing disorders and nocturnal hypoxaemia in patients discharged from ICU following prolonged mechanical ventilation. DESIGN: Prospective, consecutive patient observational study. SETTING: The medical and surgical wards of a University Hospital. PATIENTS AND PARTICIPANTS: Fifteen consecutive, adult patients discharged from the ICU who had received more than 48 h of mechanical ventilation were studied. Ten healthy volunteers acted as controls. MEASUREMENTS AND RESULTS: Overnight, multi-channel pneumographic studies were performed on all patients and controls. Chest and abdominal wall movement, air flow, oxygen saturation and snoring were continuously recorded. Data was analysed by both visual inspection of the traces and by computer-based algorithms. An apnoea/hypopnoea index was calculated for each patient and volunteer. Volunteers had an apnoea/hypopnoea index of less than 5 and had no episodes of nocturnal oxygen desaturation (SaO2 < 90%). Despite oxygen therapy 13/15 patients had episodes of desaturation and 9/15 spent more than 2 h with an SaO2 < 90%. Eleven patients had an abnormal apnoea/hypopnoea index (range 5-34 events/h). Four patients had predominantly obstructive events while 7 primarily had hypopnoeas. CONCLUSIONS: Significant overnight oxygen desaturation is common in patients discharged from ICU who have received prolonged mechanical ventilation. This group also has a significant incidence of sleep-related breathing disorders and this mechanism is likely to be important in the pathogenesis of the hypoxaemia. PMID- 10872136 TI - Dose-dependent effects of almitrine on hemodynamics and gas exchange in an animal model of acute lung injury. AB - OBJECTIVE: To determine the dose-response relationship of almitrine (Alm) on pulmonary gas exchange and hemodynamics in an animal model of acute lung injury (ALI). DESIGN: Prospective, randomized, controlled study. METHODS: Twenty anesthetized, tracheotomized and mechanically ventilated (FIO2 1.0) pigs underwent induction of ALI by repeated saline washout of surfactant. Animals were randomly assigned to either receive cumulating doses of Alm intravenously (0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 micrograms.kg-1.min-1) for 30 min each (treatment; n = 10) or to receive the solvent malic acid (controls; n = 10). MEASUREMENTS AND RESULTS: Measurements of pulmonary gas exchange and hemodynamics were performed at the end of each infusion period. Alm < 4.0 micrograms.kg-1.min-1 improved arterial oxygen pressure (PaO2) (105 +/- 9 mmHg for Alm 1.0 vs 59 +/- 5 mmHg) and decreased intrapulmonary shunt (Qs/Qt) (32 +/- 4% for Alm 1.0 vs 46 +/- 4%) (P < 0.05). Alm > or = 8.0 micrograms.kg-1.min-1 did not improve pulmonary gas exchange compared to controls. When compared to low doses of Alm < 4.0 micrograms.kg-1.min-1, high doses > or = 8.0 micrograms.kg1.min-1 decreased PaO2 (58 +/- 11 mmHg for Alm 16.0) and increased Qs/Qt (67 +/- 10% for Alm 16.0) (P < 0.05). CONCLUSIONS: In experimental ALI, effects of almitrine on oxygenation are dose-dependent. Almitrine is most effective when used at low doses known to mimic hypoxic pulmonary vasoconstriction. PMID- 10872137 TI - Functioning of ICU ventilators under hyperbaric conditions--comparison of volume- and pressure-controlled modes. AB - OBJECTIVE: To evaluate the function of four currently available, not specifically modified time-cycled ICU ventilators (EVITA 4, Oxylog 2000 HBO and Microvent from Dragerwerk, Germany and Servo 900C, Siemens-Elema, Sweden) under hyperbaric conditions using volume-controlled ventilation (VCV) and, if available, pressure controlled ventilation (PCV). DESIGN: All ventilators were studied on an electromechanical lung simulator consisting of a motor driven bellows (LS 1500, Dragerwerk, Germany) at normobaric (1 bar) and hyperbaric ambient pressures (1.3, 1.6, 1.9, 2.8 bar). Servo 900C and Microvent were additionally tested at 6 bar. SETTINGS: Hyperbaric chamber. MEASUREMENTS AND RESULTS: During VCV the tidal volume (VT) was set at 750 ml at normobaric conditions prior to starting hyperbaric exposure. During PCV the same VT setting was achieved by adjusting the inspiratory pressure level. At each ambient pressure we registered airway pressure (measured inside the bellows) and flow (derived from the linear displacement of the bellows) for a period of 1 min. From these data we calculated off-line VT, inspiratory airway peak and plateau pressure (Ppeak and Pplateau) and, during PCV only, peak inspiratory flow (Vmax) and the time delay between onset of and peak inspiratory flow (Vdelay). During VCV inspiratory flow and, consequently, VT consistently decreased with increasing ambient pressure. In contrast, during PCV VT remained stable at each condition despite a slight decrease in Vmax. CONCLUSIONS: Whenever available, PCV should be preferentially used during hyperbaric oxygen therapy due to the stability of ventilator functioning. Based on the specific ventilator properties at increasing ambient pressures, appropriate corrections should be possible which will allow the safe use of ICU ventilators even during VCV. PMID- 10872138 TI - The hemodynamic effects of prolonged respiratory alkalosis in anesthetized newborn piglets. AB - OBJECTIVE: To test the hypothesis that prolonged alkalosis decreases cardiac output and, furthermore, exacerbates hypoxic pulmonary vasoconstriction, as respiratory alkalosis is frequently induced as a therapy for persistent pulmonary hypertension of the newborn despite a lack of controlled evidence of improved outcomes. Potential adverse effects of prolonged alkalosis have been demonstrated. METHOD: Two groups (control, n = 6, and hypocapnic alkalosis, n = 6) of 1-3 day old fentanyl-anesthetized, vecuronium-paralyzed piglets were instrumented to measure cardiac index (CI) and mean systemic (MAP) and pulmonary (PAP) arterial pressures. Baseline values were recorded. Alveolar hypoxia was then induced to achieve an arterial oxygen saturation of between 50 and 60% for 15 min. Respiratory alkalosis was then induced, by increasing ventilation to achieve a pH between 7.55-7.60, and was continued for 240 min. Inspired carbon dioxide was used with hyperventilation in the control group to maintain pressure of arterial carbon dioxide (PaCO2) at 35-45 mmHg and pH of 7.35-7.45. Hypoxia was induced again at 15 and 240 min. Pulmonary and systemic vascular resistances (PVR and SVR) were calculated. RESULTS: Prolonged alkalosis led to a significant and progressive fall in mean MAP from 61 (SD 7) mmHg at the start of the study falling to 50 (SD 6.9, p = 0.043), with no effect on CI. Calculated SVR decreased (0.45 SD 0.03 vs 0.36 SD 0.05). There were no statistically significant changes in any of the variables in the control group. Neither acute nor prolonged respiratory alkalosis had a significant effect on hypoxic pulmonary vasoconstriction. CONCLUSIONS: Prolonged hyperventilation leads to systemic hypotension, however it does not exacerbate hypoxic pulmonary vasoconstriction. PMID- 10872139 TI - Translaryngeal tracheostomy: two modified techniques versus the basic technique- early experience in 75 critically ill adults. AB - OBJECTIVES: Elective tracheostomy is an established procedure in the management of ICU patients on long-term ventilation. In addition, percutaneous tracheostomy techniques are increasingly being used. In 1997, Fantoni's translaryngeal technique (TLT), another minimally invasive procedure, was introduced. While clinical studies of TLT showed that the technique is safe and can be performed rapidly, technical difficulties which sometimes led to prolonged operating times were also noted. Our study compared the basic TLT technique to a modified TLT approach and to TLT performed with the manufacturer's new, improved "Straight Cannula" set. Twenty-five patients were enrolled in each group, and the advantages and disadvantages of the respective techniques were evaluated. SETTING: Surgical ICU of a university hospital. PATIENTS: Seventy-five adult, surgical intensive care patients. MEASUREMENTS AND RESULTS: Average operating times with the modified TLT techniques were significantly shorter, 4 and 5 min respectively, as compared to 11 min for the basic TLT technique. Initially, use of the new, improved TLT set resulted in a complete passage of the tracheal cannula in two patients; uneventful Griggs's tracheostomy was performed instead. Regardless of the technique used, no other perioperative complications were noted and the perioperative gas exchange remained unaffected by the tracheostomy procedure. CONCLUSIONS: The modified TLT procedures are safer and more readily performed than the basic technique. Difficulty in the retrograde passage of the guide wire was seen only occasionally with the modified techniques. Based on our data we conclude that the modified techniques are superior to the basic technique and represent significant progress in TLT. PMID- 10872140 TI - Pressure signal transmission of five commercially available oesophageal balloon catheters. AB - OBJECTIVE: Five commercially available oesophageal balloon catheters (OBCs) were tested to evaluate the accuracy in transmitting fast-changing pressure signals which can be observed, for example, during phrenic nerve stimulation. SETTING: Research laboratory of a university hospital. METHOD: The OBCs tested varied in length (900-1390 mm) and inner diameter (0.9-1.5 mm) as well as in balloon material [latex or polyvinylchloride (PVC)]. A 180-cm tube served as a control. A sudden pressure drop was generated by the explosion of a pressurized latex balloon. The time between the pressure drop and 75, 50, 25 and 10% of the maximal pressure was measured. RESULTS: The time intervals required to transduce a pressure drop of 90% varied between the different OBCs from 85 to 476 ms (control 32 ms). Transmission time was lower in OBCs with a larger inner diameter. Shortening the OBCs resulted in a further decrease in transmission time. CONCLUSION: The type of OBC used has an impact on signal processing. An OBCs with a short transmission time should be chosen, especially if fast pressure changes are to be evaluated such as during phrenic nerve stimulation. PMID- 10872141 TI - Unilateral pulmonary oedema complicating mitral regurgitation: diagnosis and demonstration by transoesophageal echocardiography. AB - One aetiology of unilateral pulmonary oedema is mitral valve disease. We report three cases of right pulmonary oedema caused by acute mitral regurgitation. These reports underline the diagnostic value of transoesophageal echocardiography, which rapidly visualised severe mitral regurgitation with retrograde jet directed toward the right pulmonary veins. Two patients underwent prompt cardiac surgery. PMID- 10872142 TI - Meningococcal septicaemia: treatment with protein C concentrate. AB - Meningococcal septicaemia is a devastating disease with the potential to develop severe vascular complications. The incidence in Northern Ireland has risen from 27 cases notified in 1992 to 56 notified in 1997. We describe the first use of protein C concentrate in addition to antithrombin III infusion in the management of a life-threatening case of meningococcal septicaemia in the Regional Intensive Care Unit, Royal Group of Hospitals, Belfast, UK. The rationale and the evidence to support the use of protein C concentrate are discussed. Despite the apparent efficacy and safety of this treatment, subsequent cases of meningococcal septicaemia have not received protein C concentrate due to a lack of availability. PMID- 10872143 TI - Oxygen free radicals in ARDS, septic shock and organ dysfunction. PMID- 10872144 TI - Mechanisms of pulmonary edema clearance: from basic research to clinical implication. PMID- 10872145 TI - Acute severe asthma after immunotherapy with Friedmann's vaccine. PMID- 10872146 TI - Treatment of brain metastasis in patients with significant impairment of consciousness. PMID- 10872147 TI - Transient complete heart block during pulmonary artery catheter removal. PMID- 10872148 TI - Chylous ascites following radical nephrectomy: efficiency of octreotide as treatment of a ruptured thoracic duct. PMID- 10872149 TI - Decreased soluble L-selectin levels do not indicate leukocyte activation. PMID- 10872150 TI - MRA versus digital subtraction angiography in acute subarachnoid haemorrhage: a blinded multireader study of prospectively recruited patients. AB - We performed a blinded multireader study comparing MR angiography (MRA) with digital subtraction angiography (DSA) in 34 prospectively recruited patients who presented with acute subarachnoid haemorrhage (SAH). Two observers independently reviewed the MRA and DSA studies some months after clinical presentation. Presence of an aneurysm was rated on a 4-point confidence scale. Cases in which the initial interpretation of the observers varied were jointly reviewed to reach a consensus opinion. DSA was deliberately chosen not to represent the reference standard and the clinical course and surgical findings were used to explain significant differences between the consensus readings of MRA and DSA. Diagnostic confidence and interobserver agreement were, overall, higher on DSA than on MRA studies (kappa DSA = 0.64 versus kappa MRA = 0.52 with 95% CI for delta = kappa DSA-kappa MRA [-0.06, 0.31]). With both methods, discrepancies between observers were due to aneurysms overlooked rather than false-positive readings by one observer. Diagnostic accuracy therefore improved when the readings of the two observers were combined, particularly for MRA. Intermethod agreement was only fair and similar for both readers (kappa reader 1 = 0.37 versus kappa reader 2 = 0.32 with 95% CI for delta = kappa reader 1-kappa reader 2 [-0.02, 0.11]). Both interobserver and intermethod agreements improved when the data were analysed on a per-study (positive or negative study) rather than on a per-aneurysm basis. Differences in the consensus reading were due to five aneurysms (four single and one multiple) detected only with MRA and five (two single and three multiple) detected only with DSA. MRA and DSA should be regarded as complementary in the investigation of patients with acute SAH. DSA can no longer be regarded as the reference standard. PMID- 10872151 TI - The natural history of familial cerebral cavernomas: a retrospective MRI study of 40 patients. AB - Our objective was to determine the natural history and prognostic factors of familial forms of cerebral cavernous malformations (CCM). Cavernomas are one of the most common central nervous system vascular malformations. Familial CCM is increasingly diagnosed, but little is known about its natural history. In a national survey, we analysed clinical and MRI features of 173 patients from 57 unrelated French families. Of these 40 had undergone at least two clinical and MRI examinations. Occurrence of haemorrhage, new lesions, change in signal intensity and size of lesions have been studied by comparison between first and last MRI studies. The CCM were classified according to Zabramski et al. Mean follow-up was 3.2 years (range 0.5-6.5 years). We followed 232 cavernomas (mean 5.9 per patient, range 1-17). Serial MRI demonstrated changes in 28 patients (70%). Bleeding occurred in 21 lesions (9.1%) in 14 patients (35%). The haemorrhagic risk was 2.5% per lesion-year, higher in type I and brain-stem CCM. We saw 23 new lesions appear in 11 patients (27.5%), with an incidence of 0.2 lesions per patient year. Signal change was observed in 11 patients (27.5%), in 14 lesions (6%), while 9 lesions (3.9%) in 9 patients (22.5%) changed significantly in size. PMID- 10872152 TI - Proton magnetic resonance spectroscopy reflects cellular proliferative activity in astrocytomas. AB - We examined whether proton magnetic resonance spectroscopy (MRS) could provide accurate information on histological grade and cell proliferation in astrocytomas. We studied 23 patients with astrocytomas: five grade II, 10 grade III and eight with grade IV (glioblastoma multiforme). We performed proton MRS and determined the Ki-67 labeling index (LI), a tumour proliferation marker, in the same areas of the astrocytomas, and examined the statistical relationship between proton MRS and Ki-67 LI. The N-acetylaspartate (NAA)/creatine phosphocreatine (Cr) and NAA/choline (Cho)-containing compound ratios were always significantly lower and the Cho/Cr ratios significantly higher than those for normal brain. The Cho/Cr ratio correlated positively and the NAA/Cho ratio inversely with Ki-67 LI. These findings suggest that the Cho signal in proton MRS reflects cellular proliferation. In Kaplan-Meier survival analysis, there was no significant difference between high (> 2.0, 14 cases) and low (< 2.0, 9 cases) Cho/cr ratio groups. PMID- 10872153 TI - Hippocampal malrotation with normal corpus callosum: a new entity? AB - Among 527 MRI examinations of patients with a suspicion of epilepsy in 5 years, we found 32 cases of hippocampal malrotation (HIMAL). The characteristic features are: incomplete inversion of the hippocampus with and abnormally round shape; unilateral involvement of the whole hippocampus; normal signal intensity and size; blurred internal structure; an abnormal angle of collateral sulcus; abnormal position and size of the fornix; normal size of the temporal lobe; enlargement and particular configuration of the temporal horn, typical of corpus callosum agenesis; and a normal corpus callosum. In 7 cases (22%) HIMAL occurred together with developmental disorders. It was predominantly seen in men. The clinical features were varied. Based on some MRI features, the presence of developmental disorders, the male predominance, the frequently positive family history, and a review of the literature, we think HIMAL may be the consequence of a mild hemisphere developmental disorder. It is probably not the basic cause of epilepsy in such varied clinical setting, but may be a sign of a developmental disorder and can help in selecting patients for more meticulous investigation. It also may give some new understanding of brain development. PMID- 10872154 TI - Thalamic changes with mesial temporal sclerosis: MRI. AB - We reviewed the preoperative images of 28 patients with pathologically proven mesial temporal sclerosis, to assess thalamic asymmetry and signal change. A further 25 nonsurgical patients with temporal lobe epilepsy and unequivocal, unilateral changes of mesial temporal sclerosis, and 20 controls, were also reviewed. None of the control group had unequivocal asymmetry of the thalamus. There was an ipsilateral asymmetrically small thalamus in five (18%) of the surgical group and in three (12%) of the nonsurgical patients. In four cases there was thalamic signal change. In three patients with thalamic volume loss there was ipsilateral hemiatrophy. All patients with an asymmetrically small thalamus had an asymmetrically small fornix and all but one a small ipsilateral mamillary body. PMID- 10872155 TI - Regression after biopsy of a pilocytic opticochiasmatic astrocytoma in a young adult without neurofibromatosis. AB - Serial MRI over 60 months demonstrated regression after biopsy of a pilocytic opticochiasmatic astrocytoma in a 20-year-old woman with no signs of neurofibromatosis, together with improvement in vision. The patient did not receive radio- or chemotherapy. Close MRI follow-up of optic gliomas is recommended. Aggressive treatment should be limited to cases with clear clinical and radiological progression. PMID- 10872156 TI - Spontaneously regressing infundibular cyst: a case report. AB - A 74-year-old man reported headaches and blurring of vision for 1 month. MRI showed a nonenhancing infundibular cyst. Neurologic findings, blood and cerebrospinal fluid examinations, and chest and abdominal CT were all normal. MRI 4 months later showed no change. The patient was without any medication other than simple analgesics. One year later, the stalk had returned to its normal size and configuration on MRI. PMID- 10872157 TI - Spontaneous regression of a cyst of the cavum septi pellucidi. AB - A 20-year-old woman with secondary amenorrhoea and an empty sella turcica was found to have a cyst of the cavum septi pellucidi (CSP) on MRI. The cyst had regressed spontaneously on follow-up MRI. PMID- 10872158 TI - Primary malignant rhabdoid tumour of the brain in an adult. AB - We report a mass in the left cerebral hemisphere of a 20-year-old man. Histological, ultrastructural and immunohistochemical features of the tumour were consistent with primary malignant rhabdoid tumour. The age of presentation, imaging features prior to histological examination, and prognosis in this case were unusual. PMID- 10872159 TI - MRI in decompression illness. AB - We report a case of decompression illness in which the patient developed paraparesis during scuba diving after rapid ascent. MRI of the spine revealed a focal intramedullary lesion consistent with the symptoms. The pathophysiological and radiological aspects of spinal decompression illness are discussed. PMID- 10872160 TI - Needle diameter in outpatient myelography: rates of adverse effects and current practice trends. AB - Telephone calls were made to 1251 consecutive patients one day following outpatient myelography. Data were available on 518 patients punctured with 22 gauge (g) (large-diameter) and 465 with 25-g (small-diameter) spinal needles. We surveyed 48 academic and private practice groups regarding needle diameter use in myelography; data were obtained from 34 private practice and 14 academic radiology departments. Patients reported adverse effects including mild and severe headache, back pain and nausea. The percentage of total adverse effects was significantly greater in the 22-g than in the 25-g needle group. The percentage of patients with headache was higher in the 22-g than in the 25-g group, but this difference was not statistically significant. Only 19% of private practice groups and 17% of academic centers use 25-g needles; the remainder use 20-g or 22-g needles. PMID- 10872161 TI - Nasu-Hakola disease in two Tunisian siblings: new radiological findings. AB - We report radiological features of a biopsy-proven early infantile form of Nasu Hakoka disease in two Tunisian sisters with new bony and cerebral findings. PMID- 10872162 TI - Diffusion-weighted MRI and selection of patients for fibrinolytic therapy of acute cerebral ischaemia. AB - Treatment of patients with acute cerebral ischaemic events remains controversial. We investigated the reversibility of high signal intensity on diffusion-weighted (DW) MRI after acute local intra-arterial fibrinolysis (LIF) and the feasibility of DW MRI for selecting patients for acute LIF. Nine patients with acute middle cerebral artery embolic occlusion underwent single-photon emission computed tomography (SPECT) and DW MRI followed by acute LIF using tissue plasminogen activator. Recanalisation was observed in all patients, and eight improved clinically. The area of high signal intensity on pretreatment DW MRI was smaller than the low-uptake area on SPECT in all patients, and went on to infarction, as detected by MRI or CT 3 days after onset in all patients. It appears to correlate, at least clinically, with irreversible brain damage. Therefore, acute LIF should not be performed in patients with areas of high signal intensity in the cortex responsible for the symptoms. SPECT remains important, because areas normal on DW MRI with low uptake on SPECT often contribute to functional prognosis. PMID- 10872163 TI - Coil embolisation for ruptured vertebral artery dissection distal to the origin of the posterior inferior cerebellar artery. AB - Although many surgical or endovascular treatments for ruptured vertebral artery dissection have been reported, the best treatment is controversial. We treated five cases of ruptured vertebral artery dissection distal to the origin of the posterior inferior cerebellar artery (PICA), using retrievable platinum coils packed in the dissection site and the immediately proximal vertebral artery. All patients had a contralateral vertebral artery of the same calibre or larger. All dissections were occluded completely, together with the portion of the vertebral artery distal to the PICA origin. No complications related to the procedure were seen. The purpose of the treatment is to isolate the dissection from the cerebral circulation. Occlusion of the rupture site, preserving perforating arteries arising from the vertebral artery, would be ideal. Short-segment occlusion by retrievable platinum coils is close to the ideal. PMID- 10872164 TI - Selective transparenchymal venous embolisation for dural arteriovenous fistulae. PMID- 10872165 TI - Serial CT and MRI of ischaemic cerebral infarcts: frequency and clinical impact of haemorrhagic transformation. AB - The frequency, predisposing factors and clinical consequences of haemorrhagic infarcts and damaged blood-brain barrier as shown by contrast enhancement (CE) in ischaemic cerebral infarcts are controversial. We prospectively compared the sensitivity of CT and MRI to haemorrhagic transformation (HT) and CE. We also wished to investigate the clinical significance of HT and factors possibly associated with it. We studied 36 patients with acute ischaemic infarcts in the middle cerebral artery territory during the first 2 weeks after the ictus. After CT and rating of the neurological deficit on admission, serial examinations with clinical neuromonitoring, contrast-enhanced CT and MRI were done on the same day. The occurrence and severity of HT were correlated with CE, stroke mechanism, infarct size, development of neurological deficits and antithrombotic treatment. The frequency of HT detected by MRI was 80%. CE usually preceded HT or was seen simultaneously. MRI had a higher sensitivity than CT to HT and CE. Severity of HT was positively correlated with infarct size (P < 0.01). HT had no influence on patient's neurological status. Neither the type of antithrombotic treatment nor the stroke mechanism was associated with the severity of HT. No parenchymal haemorrhage occurred. PMID- 10872166 TI - Contrast-enhanced MR angiography in patients with carotid artery stenosis: comparison of two different techniques with an unenhanced 2D time-of-flight sequence. AB - Conventional time-of-flight (TOF) MR angiography (MRA) in carotid artery stenosis relies on flow-related enhancement to produce signal from vascular structures. Intravoxel phase dispersion, due to vortices, causes loss of signal and is the reason for the tendency to overestimate the degree of stenosis. In contrast enhanced MRA, intravascular signal is mainly dependent on T1 shortening of the blood. We compared first-pass contrast-enhanced MRA (contrast-enhanced 3D gradient echo, ce3D GRE) and contrast-enhanced 2D TOF (ce2D TOF) sequences with an unenhanced 2D TOF in 13 patients with carotid artery stenosis, assessing delineation of the carotid bifurcation, enhancement of veins and grade of stenosis. The contrast-enhanced techniques produced more morphological detail, the ce3D GRE being superior to the ce2D TOF. Four carotid arteries were reclassified into lesser stenosis categories using the ce3D GRE technique. However, seven carotid arteries (27%) were rated as nondiagnostic on the ce3D GRE, mainly due to masking of the carotid bifurcation by veins. The latter can be avoided by decreasing the acquisition time; on our 1.5-T system we could achieve a minimum time of 23 s per 3D GRE. Further reduction of acquisition time would be necessary to incorporate this method into clinical routine, requiring higher performance gradients, which are not available in many UK hospitals. PMID- 10872167 TI - Diagnosing CADASIL using MRI: evidence from families with known mutations of Notch 3 gene. AB - Clinical data and MRI findings are presented on 18 subjects from two families with neuropathologically confirmed CADASIL. DNA analysis revealed mutations in exon 4 of Notch 3 gene in both families. All family members with mutations in Notch 3 gene had extensive abnormalities on MRI, principally lesions in the white matter of the frontal lobes and in the external capsules. Of several family members in whom a diagnosis of CADASIL was suspected on the basis of minor symptoms, one had MRI changes consistent with CADASIL; none of these cases carried a mutation in the Notch 3 gene. MRI and clinical features that may alert the radiologist to the diagnosis of CADASIL are reviewed. However, a wide differential diagnosis exists for the MRI appearances of CADASIL, including multiple sclerosis and small-vessel disease secondary to hypertension. The definitive diagnosis cannot be made on MRI alone and requires additional evidence, where available, from a positive family history and by screening DNA for mutations of Notch 3 gene. PMID- 10872168 TI - Triple-dose contrast-enhanced images in neurologically symptomatic HIV-positive patients. AB - Our purpose was to determine whether triple-dose delayed contrast-enhanced images would improve lesion detection in patients with symptomatic human immunodeficiency virus (HIV) infection. We reviewed 33 MRI studies on 29 patients. Single-dose immediate T1-weighted spin-echo (1x-T1) images were compared with delayed triple-dose images (D3x-T1). Two neuroradiologists decided which technique showed more lesions, increased lesion conspicuity and/or altered the radiologic diagnosis. The D3x-T1 technique improved lesion detection in 14 of 29 patients (48%). In two patients (7%), the improvement changed the radiologic diagnosis by showing new meningeal lesions. PMID- 10872169 TI - Multiple infarcts in a patient with cerebral phaeohyphomycosis: CT and MRI. AB - Phaeohyphomycosis is an uncommon disorder caused by a variety of saprophytic fungi having distinctive morphologic features. Central nervous system infection typically occurs in the absence of predisposing factors and usually manifest symptoms and signs of abscess formation. We describe an otherwise healthy young man whose presentation with cerebral phaeohyphomycosis was subacute meningitis and stroke. Neuroimaging studies revealed multiple parenchymal lesions having the characteristics of recent infarcts; several vascular territories were involved. The nature of these lesions was confirmed histologically at autopsy. To our knowledge, such radiologic appearances have not previously been reported in this condition. PMID- 10872170 TI - Aqueduct stenosis due to venous ectasia with a dural arteriovenous fistula. AB - We report aqueduct compression by venous ectasia in a 65-year-old man with a dural arteriovenous fistula in the posterior cranial fossa draining into a superior vermian vein. Conventional and phase-contrast MRI showed the aqueduct stenosis and the causative dilated vein. PMID- 10872171 TI - Neuroradiological and clinical findings in rhombencephalosynapsis. AB - Rhombencephalosynapsis is an unusual disorder characterised by maldevelopment of the rhombencephalon, sometimes with supratentorial midline anomalies. We report MRI findings in a 39-year-old woman, the oldest in the literature. MRI demonstrated hypoplasia of the cerebellar vermis, with fusion of the cerebellar hemispheres and abnormally oriented folia. Supratentorial anomalies were also seen. PMID- 10872172 TI - MRI of intramedullary spinal schwannomas: case report and review of the literature. AB - Intramedullary spinal schwannomas are uncommon. We report a solitary cervical intramedullary schwannoma shown by MRI and treated surgically, and review 12 previous cases with MRI. MRI findings and pathogenesis are discussed. PMID- 10872173 TI - Transient oedema of the cervical spinal cord. AB - Transient but very intense oedema of the cervical spinal cord was observed in two patients with obstruction of the cerebrospinal fluid (CSF) pathways. Both presented with hydrocephalus, one due to an infratentorial obstructing mass and the other due to postmeningitic adhesive obstruction of the outlet foramina of the fourth ventricle. In animal experiments with obstruction of CSF pathways (due to outlet foramina obstruction or to downward tentorial herniation) flattening and stretching of the ependymal cells along the central canal is observed, followed by disruption and splitting of the ependymal lining and then by extracellular oedema of the subependymal tissue. Without treatment, frank cavity formation develops in a fourth stage. In our two patients, however, most probably because of appropriate decompressive therapy, the oedema disappeared completely without a residual spinal cord lesion. PMID- 10872174 TI - Primary malignant lymphoma of the maxillary sinus: CT and MRI. AB - We reviewed the CT and MRI of seven patients with primary malignant lymphoma of the maxillary sinus to find if there are characteristic imaging findings suggestive of the disease. The images were analysed for appearance, size, signal, internal characteristics, extent of tumour, bone change and lymph node enlargement. In two patients, the tumour first presented with mucosal thickening. In the remaining five, the tumours were an expansile mass 4-6 cm in diameter at the time of detection. Although it was difficult to distinguish tumour from mucosa or obstructed fluid on CT, T2-weighted MRI enabled us to separate tumour from normal mucosa or fluid. In two patients, the tumours were heterogeneous. Calcification and haemorrhage were observed in one patient. Periantral soft tissue infiltration was always present, even when tumour appeared as slight mucosal thickening. Posterior extension was seen in all patients. Permeative and lytic bone destruction accompanied most cases of periantral soft-tissue infiltration; mixed destruction and sclerosis was also observed. Mucosal thickening with periantral soft-tissue infiltration may suggest malignant lymphoma of the maxillary sinus in its early form. Various types of bone change may accompany the periantral soft-tissue infiltration. PMID- 10872175 TI - Blake's pouch cyst: an entity within the Dandy-Walker continuum. AB - Abnormal cerebrospinal fluid (CSF) collections within the posterior fossa are defined by the Dandy-Walker complex (DWC) and by arachnoid cysts (AC). The DWC includes the Dandy-Walker malformation (DWM), the Dandy-Walker variant (DWV) and the mega-cisterna magna (MCM). In addition, Tortori-Donati et al. added persistent Blake's pouch cyst (BPC) as an independent entity within the DWC. BPC represents a posterior ballooning of the superior medullary velum into the cisterna magna. All of these malformations are overlapping developmental anomalies characterized by varying degrees of malformation of the medullary vela, the cerebellar vermis and hemispheres, the fourth ventricle choroid plexus, the posterior fossa subarachnoid cisterns and the enveloping meningeal structures. We present two cases of persistent BPC detected in two adult women without history of gestational or subsequent growth problems. They underwent neuroradiological investigation because of headache and because of recurrent episodes of loss of consciousness, respectively. The MRI findings included tetraventricular hydrocephalus, wide communication of the fourth ventricle and the cystic posterior fossa (i.e. BPC), inferior posterior fossa mass effect with or without hypoplasia of both the cerebellar vermis and the medial aspects of the cerebellar hemispheres, and absence of communication between fourth ventricle and the basal subarachnoid space in the midline posteriorly. Persistent BPC is defined by a failure of embryonic assimilation of the area membranacea anterior within the tela choroidea associated with imperforation of the foramen of Magendie. Typically this condition becomes symptomatic early in life. In the current cases the normal function of the laterally positioned foramina of Luschka probably helped to maintain some CSF flow between intraventricular and subarachnoid spaces, with the establishment of a precarious equilibrium characterized by a compensatory enlargement of the cerebral ventricular system (i.e. hydrocephalus). PMID- 10872176 TI - Treatment of restenosis after percutaneous transluminal angioplasty for internal carotid artery stenosis. AB - The efficacy of repeated percutaneous transluminal angioplasty (PTA) and carotid endarterectomy (CEA) was examined in patients with restenosis after PTA for carotid stenosis. After percutaneous transluminal angioplasty (PTA) for 63 cases of internal carotid stenoses 13 cases of restenosis appeared. They were treated by PTA or carotid endarterectomy. The treatment was chosen by the patient after explanation of each treatment. We initially treated seven patients by repeat PTA and six by carotid endarterectomy. The degree of stenosis improved from 82% to 30% on average after repeated PTA. However, one patient in the PTA group had restenosis, and carotid endarterectomy was then performed. The other cases also had restenosis and were treated by PTA. The six cases treated by carotid endarterectomy were successfully treated without difficulty. The success rate of PTA was 5/7 (71%) in the restenosis cases. Patients with a greater residual stenosis after initial PTA had significantly more frequent restenosis. Repeat PTA and CEA both appeared effective treatment for restenosis after initial PTA, although PTA had a restenosis rate similar to that of initial PTA. PMID- 10872177 TI - Adjuvant use of epsilon-aminocaproic acid (Amicar) in the endovascular treatment of cranial arteriovenous fistulae. AB - We report our experience with the use of the antifibrinolytic agent epsilon aminocaproic acid (EACA), Amicar, as an adjuvant to endovascular treatment of cranial arteriovenous fistulae. We also review applications of antifibrinolytic agents to neurovascular disorders and discuss the mechanism of action, dosing strategy, contraindications, and possible complications associated with the use of EACA. We identified 13 patients with cranial arteriovenous fistulae (five direct carotid cavernous fistulae [CCF], seven dural arteriovenous fistulae [DAVF], and one vein of Galen malformation) who received EACA as an adjunct to endovascular treatment. In all cases embolic coils were the primary embolic agent. We reviewed the modes of initial endovascular therapy and angiographic findings immediately thereafter and the response to EACA. Two direct CCF and two DAVF were completely thrombosed on follow-up angiography, and two DAVF demonstrated diminished flow after EACA therapy. Seven fistulae did not respond to EACA. Four of eight tightly coiled fistulae thrombosed, while none of five loosely coiled fistulae thrombosed. None of four cases with a residual fistula separate from the coil mass underwent thrombosis with EACA, while four of nine cases without a separate fistula thrombosed. There was no morbidity related to EACA therapy. EACA may thus be useful as an adjunct to endovascular treatment of cranial arteriovenous fistulae. Loose or incomplete coil packing of the fistula predicts a poor response to EACA therapy. PMID- 10872178 TI - [Pharmacological and clinical aspects of fluvoxamine (Depromel), the first selective serotonin reuptake inhibitor approved for clinical use employed in Japan]. AB - Fluvoxamine (Depromel), a selective serotonin reuptake inhibitor (SSRI), was launched in May 1999 in Japan with more than 10 years' delay from the marketing in Europe and the United States. Fluvoxamine has been approved in about 80 countries as the indication to "depression" since 1983. As the indication to obsessive-compulsive disorder (OCD), fluvoxamine was first approved in the United States in 1994 and then in about 30 countries. Efficacy of the drug on "depression and depressed state" was found to be comparable to traditional tricyclic antidepressants (TCAs) by the clinical studies in Japan. Indication to OCD was first approved for fluvoxamine in Japan. The antidepressant and the anti OCD action are considered the result of the serotonin reuptake inhibition at the serotonergic neurons. Fluvoxamine has little affinity for muscarinic, adrenergic alpha 1- and histamine H1-receptors, which TCAs have. Therefore, fluvoxamine possesses less side effects such as dry mouse, disuria, dizziness, orthostatic hypotension and drowsiness, etc.; and it is useful for elderly patients and long term treatments for depression and OCD. PMID- 10872179 TI - [Pharmacological profiles of latanoprost (Xalatan), a novel anti-glaucoma drug]. AB - Latanoprost is a novel prostaglandin F2 alpha (PGF2 alpha) derivative. Topically applied latanoprost into the glaucomatous monkey eyes lowered intraocular pressure (IOP). Latanoprost, however, failed to produce the hypotensive effect in either rabbit or cat eyes. This species difference may be attributed to its high selectivity for the FP receptor and differences in prostaglandin receptor subtypes existed in the eye amongst these species. In ligand binding studies with bovine corpus luteum cell membranes, the Kd value for the FP receptor of latanoprost was the same as that for PGF2 alpha, 2.8 nM. Latanoprost augmented uveoscleral outflow (Uv) in monkeys without affecting trabecular outflow or outflow facility like PGF2 alpha. Although the precise mechanism of the increase in Uv is not fully understood, it is suggested that a decrease in extracellular matrix components in ciliary muscle may contribute to the increase in Uv. On the other hand, an increase in blood flow at the optic nerve head and neuroprotective action in addition to the IOP lowering effect may contribute to the efficacy of latanoprost in glaucoma therapy. Only tolerable conjunctival hyperemia was seen in rabbits. A phase III clinical trial revealed latanoprost (0.005%) once daily produced sustained reduction of IOP in ocular hypertension or primary open-angle glaucoma patients to a greater extent than timolol did. Furthermore, the effects of latanoprost on aqueous humor dynamics in normal human volunteers were similar to those in monkeys, indicating that latanoprost lowers IOP by the increase in Uv in humans. PMID- 10872180 TI - [Vascular effects of insulin]. AB - Insulin as a vascular hormone, apart from its effect on intermediary metabolism, has been considered to play an important role in cardiovascular regulation and pathophysiology of cardiovascular diseases such as essential hypertension, congestive cardiac failure and atherosclerosis. Insulin induces pressor effects by mechanisms of increased sympathetic activity, renal sodium retention and proliferation of vascular smooth muscle cells. On the other hand, accumulating evidence indicates that insulin decreases vascular resistance and increases organ blood flow especially in skeletal muscle tissue, indicating that insulin is a vasodilator. Several mechanisms underlying insulin-induced vasodilation have been proposed. Insulin enhances calcium efflux from vascular smooth muscle cells by activating the plasma membrane Ca(2+)-ATPase and causes hyperpolarization by stimulating Na+, K(+)-ATPase and sodium/potassium pump. Insulin also stimulates nitric oxide (NO) synthase and increases release of NO from vascular endothelium to cause vasodilation. An increase in cyclic AMP levels is induced by insulin, via activation of insulin receptors, beta-adrenoceptors and calcitonin gene related peptide receptors. However, main cause of mechanisms mediating the vasodilation remain obscure. Hypertension is associated with insulin resistance and hyperinsulinemia. Insulin resistance may contribute to hypertension by sympathetic overactivity, endothelium dysfunction and decreased vasodilator action of insulin. Therefore, insulin must be considered a vasoactive peptide and more investigations are needed to better understand the full significance of the hemodynamic effect of insulin. PMID- 10872181 TI - [Antiarrhythmic effects of pilsicainide hydrochloride and effects on cardiac function and ECG in dogs: comparison with disopyramide]. AB - Antiarrhythmic effects and cardiovascular effects of pilsicainide hydrochloride were compared with those of disopyramide in a canine model of coronary ligation induced ventricular arrhythmias and anesthetized dogs. Pilsicainide (1.25, 2.5 and 5 mg/kg) and disopyramide (2.5 and 5 mg/kg) decreased the arrhythmic ratio ?(ventricular arrhythmias/total heart rate) x 100? dose-dependently. Pilsicainide at 2.5 and 5 mg/kg and disopyramide at 5 mg/kg suppressed ventricular arrhythmias more than 50%. The effective dose of pilsicainide was lower than that of disopyramide, but the effective plasma concentration of pilsicainide was between 3 and 8 micrograms/ml, which was almost the same as that of disopyramide. In anesthetized dogs, both drugs decreased LV dP/dt max in almost the same concentration-dependent manner. PQ-interval was prolonged by pilsicainide, but not by disopyramide. QRS and QTc were prolonged by both drugs in a concentration dependent manner. However, the prolongation of QTc by disopyramide was provoked at lower plasma concentrations than by pilsicainide. Because the excessive prolongation of QTc lead to the lethal arrhythmias such as torsades de pointes, pilsicainide may be useful as an injectable antiarrhythmic agent superior to disopyramide. PMID- 10872182 TI - Pretransplant blood transfusions with cyclosporine in pediatric renal transplantation. AB - Pretransplant transfusions were repeatedly shown to be associated with improved graft survival in the "pre-cyclosporine era," and have recently been shown to be beneficial in patients on modern immuno-suppressive regimes. In an attempt to improve this transfusion effect and minimize the potential development of cytotoxic antibodies, we have given these transfusions, with concomitant cyclosporine cover, prior to transplantation. Ninety-two renal transplantations were performed in 91 children in the study group (group 1) and all received pretransplant transfusions with cyclosporine cover. Results were compared with a preceding group of 102 children (104 transplantations) who had received pretransplant transfusions without cyclosporine cover (group 2). There were 70 cadaver and 22 living-related donor (LRD) transplants in group 1, and 88 cadaver and 16 LRD transplants in group 2. Graft survival rates (1- and 5-year) for cadaver transplantation were 96% and 90% in group 1 compared with 78% and 64% in group 2 (P = 0.001). For LRD transplantation, these figures were 95% and 87% in group 1 and 81% and 69% in group 2. There was no difference between the two groups in terms of age at transplantation, sex, donor age, HLA-A, -B, -DR mismatches, or cold and warm ischemia times. All cadaver graft recipients received quadruple, sequential immunosuppression post transplant. However, 9 patients in group 1 were changed to tacrolimus for recurrent rejection episodes. No patient developed persistent lymphocytotoxic antibodies post transfusion or side effects of cyclosporine. Cyclosporine can be safely given with whole blood prior to transplantation with no adverse effect and no sensitization. Graft survival was significantly improved in this group of patients and graft loss due to rejection was exceptional. This effect should be further evaluated in prospective studies. PMID- 10872183 TI - Experience with deflazacort in children and adolescents after renal transplantation. AB - Deflazacort (DFZ) has been proposed as an alternative drug for immunosuppression after renal transplantation (TX), with fewer side effects than conventional glucocorticoids. We investigated renal function, body growth, body fat, and bone mineral density (BMD) after switching from oral methylprednisolone (MPR) to equivalent doses of DFZ 1-9 years after TX in 20 patients aged 5-20 years, selected because of severe adverse effects from previous steroid therapy. At conversion the patients received a mean dose of 7.4 +/- 2.4 mg DFZ/m2 per day. The drug was continued for a mean of 3.7 (1.2-5.5) years. Under DFZ, the glomerular filtration rate dropped slightly (NS). A single rejection episode occurred. Growth velocity significantly improved in the 1st year on DFZ treatment and height standard deviation score (SDS) increased steadily after introduction of DFZ (from -2.64 to -1.96 after 4 years, P = 0.06). However, in 10 prepubertal children the height gain (+0.20 SDS in 2 years on DFZ) was not significant and the overall mean annual growth rate after TX was similar to that in 10 matched prepubertal TX children on continued MPR treatment. Relative obesity, estimated from mean body mass index corrected for height, was reduced from +1.11 SDS at the start of DFZ to +0.71 SDS after 2 years (P = 0.03) and to +0.39 SDS after 4 years (NS). BMD-SDS of the lumbar spine (L2-4) increased after 1 year on DFZ (P = 0.005). In conclusion, DFZ is well tolerated and safe in pediatric patients after TX. It improves relative obesity and bone mineralization. However, body growth is not significantly influenced pre puberty. PMID- 10872184 TI - Autosomal dominant hemolytic uremic syndrome: variable phenotypes and transplant results. AB - Autosomal dominant hemolytic uremic syndrome (ADHUS) is a rare disorder with a poor prognosis that was considered to present mainly in adults. Recurrent episodes of ADHUS were also thought to be uncommon. However, increasing reports suggest that children are often affected, that recurrent episodes may occur pretransplantation, and that post-transplant recurrences occur in about 50% of cases. We describe the occurrence of ADHUS in two unrelated families with different outcomes. It is apparent that there are several types of ADHUS (with and without serum complement abnormalities) and that there may be variable expression of the phenotype. There was variable penetration of HUS in four (possibly five) adults and two children in four generations of one kindred. There was also one definite unaffected carrier. Two patients had successful renal transplants without recurrences; the HUS recurred in a third patient soon after transplantation. In a second family, the father had three episodes of HUS at 18, 26 and 29 years of age; his son had one episode of HUS at 5 years of age. Both recovered completely. Evaluation of these patients, previous reports, and follow up contacts of previous reports suggests that post-transplant recurrence of ADHUS is more common than previously reported and may not be prevented by prior nephrectomy of native kidneys. PMID- 10872185 TI - Non-compliance and transfer from paediatric to adult transplant unit. AB - Adolescents and young adults appear to be a particularly high-risk group for problems of non-compliance and associated graft loss. We reviewed the progress of 20 young adults (9 female) who had been transferred to three different adult centres at a mean age of 17.9 years (range 15.7-20.9 years) having been transplanted at a mean age of 14.3 years (range 9.6-18.1 years) in the paediatric unit. Eight transplants failed within 36 months of transfer, and in 7 of 20 (35%) the transplant failure was unexpected (3 < 12 months, 3 12-24 months, 1 31 months post transfer). Although many of the patients had recognised problems in family dynamics, only 1 had had a major rejection episode prior to transfer due to admitted non-compliance. In 3 others low cyclosporin levels had been noted. Two young men had been transplanted preemptively in the paediatric unit at 15.3 and 16.7 years, and 3 patients had been transferred to the adult unit via the recently established transition clinic. The results suggest that close attention needs to be paid to this group of patients who require ongoing education and support. Improved dialogue between staff of the paediatric and adult units about transition issues is also essential. PMID- 10872186 TI - Growth hormone and markers of immune function in children with renal transplants. AB - Lymphocyte subsets and T cell activation markers were measured in ten children with renal transplants for up to 1 year before and during their 1st year of recombinant human growth hormone (rhGH) treatment. The number of lymphocytes, helper or cytotoxic T cells or natural killer cells, and the T cell expression of CD25, CD26 and HLA-DR antigens were not altered by rhGH. B cell numbers declined both before and during treatment. There was no difference in lymphocyte subset numbers between children with and without rejection episodes. PMID- 10872187 TI - Torasemide is an effective diuretic in the newborn rabbit. AB - The effect of intravenous (i.v.) torasemide on diuresis and renal function was evaluated in three groups of normoxemic, 5- to 10-day-old, newborn New Zealand White rabbits. The animals of group 1 received 0.2 mg/kg of torasemide i.v., whereas in group 2 an i.v. dose of 1.0 mg/kg was given. The third group of animals received a bolus i.v. dose of 1.0 mg/kg torasemide with continuous i.v. replacement of estimated urinary fluid and electrolyte losses. Torasemide proved to be an effective, potassium-sparing diuretic, without significant effect on glomerular filtration rate (GFR). Renal blood flow (RBF) fell and the renal vascular resistance (RVR) rose in all three groups of animals, although the rise in RVR in group 3 was not significant. These changes in renal hemodynamics were most pronounced in the animals of group 2 and are probably secondary to torasemide-induced hypovolemia (2.8% loss of body weight) and accompanying humoral reactions, such as an increase in angiotensin II (not measured). When the latter is prevented by simultaneous re-infusion of an electrolyte solution (group 3), replacing urinary losses, GFR increases and the changes in RBF and RVR are blunted. We conclude that torasemide is an effective, potassium-sparing diuretic in newborn rabbits. No evidence was found for a vasodilatory action of the drug. PMID- 10872188 TI - A novel G472R mutation in a Turkish family with X-linked Alport syndrome. AB - Alport syndrome (AS) is a hereditary disorder of progressive nephritis. Most cases are X-linked, but autosomal forms have been reported. The X-linked form is associated with mutations in the COL4A5 gene that encodes the alpha 5 chain of type IV collagen. More than 200 mutations have been reported in X-linked AS. We report a novel 1616 G > A mutation resulting in glycine substitution to arginine at position 472 in a Turkish family with a severely affected man and several variably affected women. This is the first Turkish family in whom the molecular basis of the disease has been reported. PMID- 10872189 TI - Steroid-sensitive nephrotic syndrome associated with Kimura disease. AB - We report an 11-year-old Japanese boy with Kimura disease and associated nephrotic syndrome. Before the diagnosis of Kimura disease was established, the patient had three episodes of swelling on the left cheek with subsequent nephrotic syndrome. Steroids were effective for both conditions. However, both conditions recurred within months of discontinuation of steroids. For the fourth episode of swelling on the left cheek, cyclosporine (CsA) was used. The subcutaneous tumor responded to CsA and disappeared within a few days. There has been no subsequent relapse of the nephrotic syndrome to date. PMID- 10872190 TI - Nephrotic syndrome associated with Kimura disease. AB - Kimura disease presents as benign subcutaneous swelling predominantly around the head and neck region. It has a high incidence of renal involvement. However, the pathogenesis of this association remains elusive. Only 2 pediatric cases and 11 adult cases of Kimura disease with renal involvement have been reported in the literature. In recent years many immunopathogenetic features suggesting an underlying T-cell and related cytokine defect have been noted in Kimura disease. We describe a unique case of an Asian boy who presented with nephrotic syndrome resistant to steroid and cytotoxic therapy, and 5 years later developed cervical lymphadenopathy consistent with Kimura disease. We also review the literature, summarizing the presentation, differential diagnosis, incidence of renal disease, prognosis, immunopathogenetic features, and therapy. PMID- 10872191 TI - Rickets in an infant with Williams syndrome. AB - Calcium homeostasis is altered in patients with Williams syndrome. We report an infant in whom Williams syndrome was diagnosed at 4 weeks who presented with hypercalcemia, hypercalciuria, and medullary nephrocalcinosis. Fluorescence in situ hybridization demonstrated a deletion of the elastin gene on chromosome 7. This infant was treated with a low-calcium/vitamin D-deficient infant formula that resulted in the development of rickets. Replacement of the low calcium/vitamin D-deficient formula with standard formula led to resolution of the rickets. PMID- 10872192 TI - Subcutaneous nodules in Henoch-Schonlein purpura. AB - A 6.5-year-old boy with active Henoch-Schonlein purpura developed subcutaneous nodules (SCN), a vasculitic manifestation previously unreported in children with this disease. The authors suggest that pressure played a role in the pathogenesis of the SCN. PMID- 10872193 TI - Renal Fanconi syndrome: first sign of partial respiratory chain complex IV deficiency. AB - A 2-year-old boy who developed hypophosphatemic rickets without signs of muscular weakness or neurological disturbances is presented. Biochemical findings included hypophosphatemia, metabolic acidosis, hypouricemia, hyperphosphaturia, severe glucosuria, generalized hyperaminoaciduria, hypercalciuria, proteinuria with elevated excretion of IgG, transferrin, albumin and high levels of alpha-1 microglobulin. Urine concentration capacity and creatinine clearance were normal. Lactaturia without elevated levels of plasma lactate and a high urinary excretion of beta-hydroxybutyrate were suggestive for mitochondriopathy. Partial deficiency of cytochrome c oxidase (complex IV of the respiratory chain) was found in skeletal muscle. A renal biopsy specimen demonstrated enlarged mitochondria with abnormal arborization and disorientation of the cristae in the proximal tubular cells. Reduced activity of mitochondrial cytochrome c oxidase in tubular cells could be demonstrated by ultracytochemistry. In conclusion, rickets due to the renal Fanconi syndrome can be the first clinical sign of mitochondrial cytopathies without extra-renal symptoms. Elevated excretion of lactate and ketone bodies in urine may serve as a diagnostic marker. PMID- 10872194 TI - Maxillomandibular brown tumor--a rare complication of chronic renal failure. AB - We report a 17-year-old hemodialysis patient with a rapidly growing maxillary mass diagnosed as a brown tumor. Although successful control of the parathyroid hormone (PTH) levels was achieved by treatment with vitamin D pulse therapy, the lesion progressed, invaded the maxillary sinus, and caused severe eating and speech disabilities. No recurrence was observed following surgical excision. The differential diagnosis and considerations regarding the causes of the disease in a child, therapy options, and review of the literature are presented. PMID- 10872195 TI - Alport syndromes: phenotypic heterogeneity of progressive hereditary nephritis. AB - Alport syndrome is a primary genetic disease of basement membranes, manifested clinically as a progressive nephropathy variably associated with sensorineural deafness and a plethora of ocular abnormalities. The long-recognized phenotypic heterogeneity of Alport syndrome may be considered on several levels, including basement membrane biochemistry, basement membrane ultrastructure, the natural history of the nephropathy, and the occurrence of extrarenal abnormalities. This review discusses the possible molecular bases for the heterogeneity. The discussion draws upon recent insights into the molecular genetics of Alport syndrome, and the biochemistry of normal and Alport syndrome basement membranes, in order to provide a framework for understanding the variable renal and extrarenal manifestations of the disease. PMID- 10872196 TI - Transplantation of renal precursor cells: a new therapeutic approach. AB - The number of kidney transplantations performed per year is limited due to availability of donor organs. One possible solution to the organ shortage is the use of renal xenografts. However, the transplantation of xenografts is complicated by hyperacute and acute rejection. It has been postulated that the host immune response might be attenuated following the transplantation of renal precursor cells or embryonic kidneys (metanephroi) instead of developed (adult) kidneys. Transplanted metanephroi become chimeric organs in that their blood supply originates, at least in part, from the host. It is possible to transplant a developing metanephros, without the use of immunosuppression, from one rat to another. Transplanted metanephroi grow, develop, become vascularized, and function in host rats. Transplantation of metanephroi may be a promising novel therapeutic approach for the treatment of chronic renal failure. PMID- 10872197 TI - In defense of altruistic kidney donation by strangers. AB - A shortage of cadaveric donor kidneys has created waiting lists for patients on chronic dialysis. Despite many ethical issues, donor kidneys are obtained from cadavers, first-degree living relatives, second-degree relatives (uncles, aunts), emotionally related persons such as spouses, and non-genetic altruistic donors who have a close relationship with the recipient. Most centers do not accept kidneys from minors, persons who have no genetic or personal relationship with the recipient, organs offered by altruistic strangers, or those that are purchased. The pros and cons of using kidneys from donors who are altruistic strangers (donors who have no genetic or personal relationship with the recipient) are reviewed. It may seem that organ acquisition for renal transplantation has moved down a slippery slope from cadaver donors to living non related but emotionally related donors. However, it can also be argued that the approach to obtaining kidneys has evolved with improvements in safety to the donor and an increasing shortage of organs. It may also be argued that the approach should evolve from a paternalistic physician-centered role to a position in which the patient has more autonomy in deciding whether or not to accept a kidney from an altruistic donor. PMID- 10872198 TI - In defense of altruistic kidney donation by strangers: a commentary. PMID- 10872199 TI - Factor V Leiden: a risk factor for renal vein thrombosis in renal transplantation. PMID- 10872200 TI - Steroid-resistant nephrotic syndrome at 4 months: no complications over 10 years. PMID- 10872201 TI - Maxillary brown tumor caused by secondary hyperparathyroidism in a boy. PMID- 10872202 TI - Podophyllotoxin. AB - Podophyllin, an ethanolic extract of Podophyllum peltatum L. or P. emodi Wall (syn. P. hexandnum Royle), is a good source of the aryltetralin-type lignan, podophyllotoxin. The latter compound, as well as its congeners and derivatives exhibit pronounced biological activity mainly as strong antiviral agents and as antineoplastic drugs. The podophyllotoxin derivatives etoposide, etopophos (etoposide phosphate), and teniposide are thus successfully utilized in the treatment of a variety of malignant conditions. Continued research on the Podophyllum lignans is currently focused on structure optimization to generate derivatives with superior pharmacological profiles and broader therapeutic scope, and the development of alternative and renewable sources of podophyllotoxin. PMID- 10872203 TI - Plant aromatic L-amino acid decarboxylases: evolution, biochemistry, regulation, and metabolic engineering applications. AB - A comprehensive survey of the extensive literature relevant to the evolution, physiology, biochemistry, regulation, and genetic engineering applications of plant aromatic L-amino acid decarboxylases (AADCs) is presented. AADCs catalyze the pyridoxal-5'-phosphate (PLP)-dependent decarboxylation of select aromatic L amino acids in plants, mammals, and insects. Two plant AADCs, L-tryptophan decarboxylase (TDC) and L-tyrosine decarboxylase (TYDC), have attracted considerable attention because of their role in the biosynthesis of pharmaceutically important monoterpenoid indole alkaloids and benzylisoquinoline alkaloids, respectively. Although plant and animal AADCs share extensive amino acid homology, the enzymes display striking differences in their substrate specificities. AADCs from mammals and insects accept a broad range of aromatic L amino acids, whereas TDC and TYDC from plants exhibit exclusive substrate specificity for L-amino acids with either indole or phenol side chains, but not both. Recent biochemical and kinetic studies on animal AADCs support basic features of the classic AADC reaction mechanism. The catalytic mechanism involves the formation of a Schiff base between PLP and an invariable lysine residue, followed by a transaldimination reaction with an aromatic L-amino acid substrate. Both TDC and TYDC are primarily regulated at the transcriptional level by developmental and environmental factors. However, the putative post-translational regulation of TDC via the ubiquitin pathway, by an ATP-dependent proteolytic process, has also been suggested. Isolated TDC and TYDC genes have been used to genetically alter the regulation of secondary metabolic pathways derived from aromatic amino acids in several plant species. The metabolic modifications include increased serotonin levels, reduced indole glucosinolate levels, redirected shikimate metabolism, increased indole alkaloid levels, and increased cell wall-bound tyramine levels. PMID- 10872204 TI - Enzyme kinetics and chemical modification of alpha-1,4-glucan lyase from Gracilariopsis sp. AB - The kinetic properties and active site amino acids of alpha-1,4-glucan lyase from the marine red macroalga Gracilariopsis sp. were examined. Using 1H NMR spectroscopy the alpha-1,4-glucan lyase was found to degrade alpha- and beta maltose at different rates. The effect of pH on the kinetic constants suggested the presence of two catalytically important amino acids in the active site with pKa values of 3.5 and 6.2. The former indicated the presence of an ionised aspartate or glutamate residue in the active site. This was tested using the carboxyl specific reagent EDAC, which inhibited enzyme activity in a time dependent manner when an external nucleophile was added. No protection against the inactivation was obtained by addition of amylopectin, maltitol or 1 deoxinojirimycin. Inactivation decreased Vmax over 2.5-fold with little effect on Km which supports the direct involvement of a carboxyl group in catalysis. PMID- 10872205 TI - Purification and characterization of polygalacturonase from banana fruit. AB - Polygalacturonase isoenzyme 3 (PG-3) was purified to homogeneity with a specific activity of 0.7 mu katal mg-1 protein from banana fruit pulp. The purified enzyme was a glycoprotein with ca. 8% carbohydrate. The molecular weight of the native enzyme was found to be 90 +/- 10 kDa with a subunit molecular weight of 29 +/- 2 kDa. The enzyme exhibited optimum activity at pH 4.3 and temperature 40 degrees C with activation energy 35.4 kJ mol-1. A unique property of the enzyme was the requirement of -SH groups for the enzyme activity. The enzyme was inhibited by p CMB and activated by 2-ME and DTT. The inhibition of p-CMB could be reversed by DTT. The enzyme contained eight free -SH groups. The Km of the enzyme was 0.15% for polygalacturonic acid. PMID- 10872206 TI - Biosynthesis of avenacins and phytosterols in roots of Avena sativa cv. Image. AB - In keeping with the proposal that avenacin biosynthesis is restricted to the tips of primary roots of oat seedlings, the incorporation of radioactivity from R-[2 (14)C]mevalonic acid (MVA) into avenacins and beta-amyrin by serial sections of primary roots was found to be more-or-less restricted to root tip sections. Squalene synthase (SQS) (EC 2.5.1.21) and 2,3-oxidosqualene:beta-amyrin cyclase (OS beta AC) (EC 5.4.99) were also most active in these sections. The incorporation of radiolabel from R-[2-(14)C]MVA into cycloartenol and 24 methylene cycloartanol by, and the 2,3-oxidosqualene:cycloartenol cyclase (OSCC) (EC 5.4.99) activity in, the various serial sections were consistent with phytosterol biosynthesis occurring in all the sections of the root with some tailing-off in the rate of synthesis in the more distal sections. PMID- 10872207 TI - Dehydrozaluzanin C: a potent plant growth regulator with potential use as a natural herbicide template. AB - The natural product dehydrozaluzanin C (former DHZ) is a sesquiterpene lactone obtained from different weeds of the Compositae family. Its potential as a plant growth regulator has been evaluated by using a phytotoxic allelopathic bioassay, where the commercial herbicide Logran is used as internal reference. The evaluation is made based on their effects on germination and growth over several dicotyledon and monocotyledon species. The activity was tested in the range of 1000-0.001 microM. In almost all cases, DHZ was more active than the internal reference at 1000 microM, and its activity fell below the level of the internal reference at 100 microM. These results confirm DHZ as a potent plant growth regulator and a good candidate for the development of new herbicide models. PMID- 10872208 TI - The structure of cranberry proanthocyanidins which inhibit adherence of uropathogenic P-fimbriated Escherichia coli in vitro. AB - Ethyl acetate extracts of Sephadex LH20-purified proanthocyanidins of American cranberry (Vaccinium macrocarpon Ait.) exhibited potent biological activity by inhibiting adherence of uropathogenic isolates of P-fimbriated Escherichia coli bacteria to cellular surfaces containing alpha-Gal(1-->4)beta-Gal receptor sequences similar to those on epithelial cells in the urinary tract. The chemical structures of the proanthocyanidins were determined by 13C NMR, electrospray mass spectrometry, matrix-assisted laser absorption time-of-flight mass spectrometry and by acid catalyzed degradation with phloroglucinol. The proanthocyanidin molecules consisted predominantly of epicatechin units with mainly DP of 4 and 5 containing at least one A-type linkage. The procyanidin A2 was the most common terminating unit occurring about four times as frequently as the epicatechin monomer. PMID- 10872209 TI - Antifungal compounds from idioblast cells isolated from avocado fruits. AB - (E,Z,Z)-1-Acetoxy-2-hydroxy-4-oxo-heneicosa-5,12,15-triene was isolated from avocado, Persea americana Mill., idioblast cells. It inhibited spore germination of the fungal pathogen Colletotrichum gloeosporioides. Full characterization is also reported for two additional compounds that have been described and partially characterized previously. PMID- 10872210 TI - Iridoid glycosides from Gmelina philippensis. AB - Six new iridoids, 6-O-alpha-L-(2"-O-, 3"-O-, 4"-O tribenzoyl)rhamnopyranosylcatalpol, 6-O-alpha-L-(2"-O-, 3"-O-dibenzoyl, 4"-O-cis p-coumaroyl)rhamnopyranosylcatalpol, 6-O-alpha-L-(2"-O-, 3"-O-dibenzoyl, 4"-O trans-p-coumaroyl)rhamnopyranosylcatalpol, 6-O-alpha-L-(2"-O-benzoyl, 3"-O-trans p-coumaroyl)rhamnopyranosylcatalpol, 6-O-alpha-L-(2"-O-, 3"-O dibenzoyl)rhamnopyranosylcatalpol, and gmephiloside as well as five known monoacyl and diacyl rhamnopyranosylcatalpol derivatives were isolated from the aerial parts of Gmelina philippensis. Their structures were established by spectroscopic means. Additionally, the known iridoids catalpol, geniposidic acid, gardoside, and 8-epi-loganic acid were identified and quantified by GC and GC-MS. The taxonomic significance of rhamnopyranosylcatalpol derivatives and iridoid acids as chemical characters is discussed. PMID- 10872211 TI - A sesquiterpene acid and flavonoids from Polygonum viscosum. AB - 4-Isobutyl-6-methyl-5-oxo-3a,4,5,7a-tetrahydro-1H-inden-13-oic acid (named viscosumic acid) and quercetin 3-O-(6"-feruloyl)-beta-D-galactopyranoside, and the known 3',5-dihydroxy-3,4',5',7-tetramethoxyflavone have been isolated from Polygonum viscosum. The structures of these isolates were determined primarily on the basis of extensive 1D and 2D NMR spectral analyses, notably, 13C PENDANT, COSY45, TOCSY, GOESY, NOESY, HMQC and HMBC. PMID- 10872212 TI - Prenylated sulfonyl amides from Glycosmis species. AB - Nine new sulfur containing amides were isolated from the lipophilic leaf extracts of different varieties of Glycosmis chlorosperma and G. ex aff. pseudoracemosa mainly collected in Thailand. Their structures were elucidated by spectroscopic methods. All amides were shown to be characterized by a methylsulfonylpropenoic acid moiety linked to a p-geranyloxy- or p-prenyloxy-phenethylamide rest. The compounds differ by different states of oxidation (i) at the 2-position of the ethylamine unit, (ii) at the aromatic m-position of phenethylamine, or (iii) at the terminal methyl group of the geranyloxy side chain. PMID- 10872213 TI - Normethyl pentacyclic and lanostane-type triterpenes from Adiantum venustum. AB - Phytochemical studies on the aerial parts of Adiantum venustum resulted in the isolation of normethyl lupane-type, a normethyl oleanane-type and a lanostane type triterpene. Structures of these triterpenes have been established as 30 normethyl lupane-20-one, 30-normethyl olean-3-one-30 beta-ol and lanost-20(22) ene-30-ol on the basis of spectral data analyses and chemical methods. PMID- 10872214 TI - Taxoids from the needles of the Canadian yew. AB - Systematic characterization of the taxoids in the needles of Taxus canadensis led to the discovery of seven taxanes along with three known congeners. Their structures were rigorously established by spectroscopic methods as 15-benzoyl-10 deacetyl-2-debenzoyl-10-dehydro-abeo-baccat in III; 15-benzoyl-2-debenzoyl-7, 9 dideacetyl-abeo-baccatin VI; N-acetyl-N-debenzoyltaxol; 7,9,13 trideacetylbaccatin VI; 10-deacetyl-10-glycolylbaccatin IV; 1 beta-hydroxy-10 deacetyl-10-glycolylbaccatin I; and 7-deacetyltaxuspine L. These taxanes, specific to the Canadian yew, were co-isolated with taxacustin, taxagifine and 2 deacetyl-7,10-diacetyl-5-deaminoacyl taxine A previously found in Taxus cuspidata, baccata, and yunnanensis, respectively. PMID- 10872215 TI - The hypersensitive response is associated with host and nonhost resistance to Phytophthora infestans. AB - The interaction between Phytophthora infestans (Mont.) de Bary and Solanum was examined cytologically using a diverse set of wild Solanum species and potato (S. tuberosum L.) cultivars with various levels of resistance to late blight. In wild Solanum species, in potato cultivars carrying known resistance (R) genes and in nonhosts the major defense reaction appeared to be the hypersensitive response (HR). In fully resistant Solanum species and nonhosts, the HR was fast and occurred within 22 h. This resulted in the death of one to three cells. In partially resistant clones, the HR was induced between 16 and 46 h, and resulted in HR lesions consisting of five or more dead cells, from which hyphae were occasionally able to escape to establish a biotrophic interaction. These results demonstrate the quantitative nature of the resistance to P. infestans. The effectiveness of the HR in restricting growth of the pathogen differed considerably between clones and correlated with resistance levels. Other responses associated with the defense reaction were deposition of callose and extracellular globules containing phenolic compounds. These globules were deposited near cells showing the HR, and may function in cell wall strengthening. PMID- 10872216 TI - Immunolocalization of LeAGP-1, a modular arabinogalactan-protein, reveals its developmentally regulated expression in tomato. AB - Arabinogalactan-proteins (AGPs) are highly glycosylated cell surface proteins that are thought to function in plant growth and development. The developmentally regulated expression of LeAGP-1, a novel and major AGP in tomato, was examined in different organs and tissues of tomato (Lycopersicon esculentum Mill. cv. UC82B) plants with an anti-peptide antibody (i.e. the PAP antibody) directed specifically against the lysine-rich subdomain of the LeAGP-1 core protein. During cell differentiation in tomato plants, LeAGP-1 was associated with cell wall thickening and lignification of particular cell types. Specifically, LeAGP-1 was detected in secondary wall thickenings of maturing metaxylem and secondary xylem tracheary elements in roots and stems, and in thickened cell walls of phloem sieve elements. However, LeAGP-1 was also present in thin-walled, cortical parenchyma cells of seedling roots as well as thick-walled collenchyma cells in young stems, both of which are not lignified. Based on these observed patterns, possible roles for LeAGP-1 in plant growth and development are discussed. PMID- 10872217 TI - The genes ABI1 and ABI2 are involved in abscisic acid- and drought-inducible expression of the Daucus carota L. Dc3 promoter in guard cells of transgenic Arabidopsis thaliana (L.) Heynh. AB - The ABA INSENSITIVE1 (ABI1) and ABI2 genes encode homologous type-2C protein phosphatases with redundant yet distinct functions in abscisic acid (ABA) responses. Results from Northern blot analysis showed that ABA- and mannitol inducible expression of the COR47 and COR78/LTI78/RD29A (COR78) genes was more impaired in the abi2 mutant of Arabidopsis thaliana (L.) Heynh than in the abi1 mutant. Furthermore, ABA-plus-mannitol treatments were additive towards COR47 gene expression; however, the ABA-deficient aba1 mutant showed reduced COR expression relative to the wild type in response to mannitol and ABA-plus mannitol treatments. These results support the notion that drought- and ABA signalling pathways are separate yet overlapping. To facilitate quantitative analysis of the genetic control of tissue-specific ABA- and desiccation-response pathways, we analyzed ABA- and mannitol-inducible expression of a carrot (Daucus carota L.) Dc3 promoter:uidA (beta-glucuronidase; GUS) chimaeric reporter (Dc3 GUS) in transgenic wild-type, ABA-deficient aba1, and ABA-insensitive abi1 and abi2 mutants. The Dc3 promoter directed ABA- and mannitol-inducible GUS expression in Arabidopsis guard cells and the two treatments were additive. The aba1, abi1, and abi2 mutant genotypes had reduced GUS expression in guard cells of cotyledons in response to mannitol, whereas abi1 and abi2 mutants were reduced in ABA-inducible GUS expression, consistent with overlapping ABA- and drought response pathways. Quantitative fluorometric GUS assays of leaf extracts showed that abi2 mutants responded less to exogenous ABA than did abi1 mutants, and abi2 mutants responded more to mannitol than did abi1 mutants. We conclude that Dc3 GUS Arabidopsis is a tractable system in which to study tissue-specific ABA and drought signalling and suggest that ABI2 functions predominantly over ABI1 in COR78 and COR47 gene expression and guard-cell Dc3-GUS expression. PMID- 10872218 TI - Ectopic expression of maize knotted1 results in the cytokinin-autotrophic growth of cultured tobacco tissues. AB - The ectopic expression of knotted homologues has cytokinin-like effects on plant morphology. The functional relationship between knotted and cytokinins was investigated in cultures of leaf tissue established from tobacco (Nicotiana tabacum L. cv. Havana 425) plants transformed with the maize knotted1 (kn1) gene regulated by cauliflower mosaic virus 35S RNA expression signals. In contrast to leaf tissues of untransformed plants, leaf tissues of kn1 transformants were capable of sustained, cytokinin-autotrophic growth on auxin-containing medium and resembled the tobacco cytokinin-autotrophic mutants Hl-1 and Hl-2. The concentration of 18 cytokinins was measured in cultures initiated from leaves of three independent kn1 transformants and the Hl-1 and Hl-2 mutants. Although cytokinin contents were variable, the content of several cytokinins in Kn1, Hl-1 and Hl-2 tissue lines was at least 10-fold higher than that of wild-type tobacco tissues and in the range reported for other cytokinin-autotrophic tobacco tissues. These results suggest that the cytokinin-autotrophic growth of Kn1 lines could result from elevated steady-state levels of cytokinins. PMID- 10872219 TI - A cytoskeletal basis for wood formation in angiosperm trees: the involvement of microfilaments. AB - The cortical microfilament (MF) component of the cytoskeleton within axial elements of the secondary vascular system of the angiosperm tree, Aesculus hippocastanum L. (horse-chestnut) was studied using transmission electron microscopy of ultrathin sections and indirect immunofluorescence microscopy of actin in thick sections. As seen by electron microscopy, MF bundles have a net axial orientation within fusiform cambial cells and their secondary vascular derivatives (i.e. in the axial xylem and phloem parenchyma, xylem fibres, vessel and sieve elements, and companion cells). Immunofluorescence studies, however, reveal that this axial orientation can be more accurately described as a helix of extremely high pitch; it is a persistent feature of all axial secondary vascular elements during their development. Helical MF arrays are the only arrangement seen in secondary phloem cells. However, in addition to helices, other MF arrays are seen in secondary xylem cells. For example, fibres possess ellipses of MFs associated with simple-pit formation, and vessel elements possess circular arrays of MFs that associate with the developing inter-vessel bordered pits, ray-vessel contact pits, and with the perforation plate. Linear MF arrays are seen co oriented with the developing tertiary wall-thickenings in vessel elements. The possible roles of MFs during the cytodifferentiation of secondary vascular cells is discussed, and compared with that of microtubules. PMID- 10872220 TI - Vacuolar granules in Chlamydomonas reinhardtii: polyphosphate and a 70-kDa polypeptide as major components. AB - The alga Chlamydomonas reinhardtii contains cytoplasmic vacuoles that are often filled with a dense granule that is released from the cell by exocytosis. Purified granules contained polyphosphate, complexed with calcium and magnesium, as the predominant inorganic components. Antiserum was raised against the major 70-kDa protein in granules purified from wall-deficient (cw15) mutants, which reacted on immunoblots with larger glycoprotein complexes in purified cell wall fractions from wild-type cells. Confocal fluorescence microscopy detected binding of these antibodies predominantly at the periphery of wall-containing C. reinhardtii y1 cells but primarily to loci in the interior of cells of the cw15 strain. Immunoelectron microscopy demonstrated that the 70-kDa protein was localized in vacuolar granules and the trans-Golgi network in sections of cw15 cells but not in the cytosol or chloroplast. Treatment of cells with a dye, fluorescent in its protonated form, indicated that the pH within vacuoles was lower than that in the cytosol, which suggested that the vacuoles are similar to lysosomes. Thus, the vacuoles may serve a dual function to provide an environment for degradation within the cell and also serve as a vehicle for secretion of specific proteins. PMID- 10872221 TI - The auxin-resistant diageotropica mutant of tomato responds to gravity via an auxin-mediated pathway. AB - Hypocotyls of the diageotropica (dgt) mutant of tomato (Lycopersicon esculentum Mill.) do not elongate in response to exogenous auxin, but can respond to gravity. This appears paradoxical in light of the Cholodny-Went hypothesis, which states that shoot gravicurvature results from asymmetric stimulation of elongation by auxin. While light-grown dgt seedlings can achieve correct gravitropic reorientation, the response is slow compared to wild-type seedlings. The sensitivity of dgt seedlings to inhibition of gravicurvature by immersion in auxin or auxin-transport inhibitors is similar to that of wild-type plants, indicating that both an auxin gradient and auxin transport are required for the gravitropic response and that auxin uptake, efflux, and at least one auxin receptor are functional in dgt. Furthermore, dgt gravicurvature is the result of asymmetrically increased elongation as would be expected for an auxin-mediated response. Our results suggest differences between elongation in response to exogenous auxin (absent in dgt) and elongation in response to gravistimulation (present but attenuated in dgt) and confirm the presence of two phases during the gravitropic response, both of which are dependent on functional auxin transport. PMID- 10872222 TI - A DEFICIENS homologue is down-regulated during apomictic initiation in ovules of Hieracium. AB - Hieracium is a member of the Asteraceae family, and contains sexual species in addition to apomictic species that reproduce by apospory and produce seed without fertilization. A homologue of the floral organ-identity gene DEFICIENS (DEF) was isolated from an apomictic line of Hieracium piloselloides (Vill.) following differential display between mature ovules and those initiating autonomous embryogenesis. The gene termed HPDEF has 93% amino acid identity with GDEF2, a DEF homologue isolated from Gerbera hybrida (D. Yu et al., 1999, Plant J. 17: 51 62), another member of the Asteraceae. In-situ analysis showed that early in floral development HPDEF is expressed in stamen and petal primordia, indicating expected B-function activity, according to the ABC model of floral organ identity (J. L. Bowman et al., 1991, Development 112: 1-20; E. S. Coen and E. M. Meyerowitz, 1991, Nature 353: 31-37). However, HPDEF expression was also observed in ovule primordia and expression continued in developing ovules until anthesis, indicating that this gene may have a role in ovule development. Expression of HPDEF was not detected in megaspore mother cells, or in sexual or aposporous embryo sacs. In sexual Hieracium, HPDEF was uniformly expressed throughout the ovule integument until anthesis. In most ovules of the apomict, however, HPDEF expression was transiently down-regulated in a specific zone in the chalazal region where cells initiating aposporous embryo sac formation differentiate. Uniform low-level HPDEF expression was subsequently observed prior to anthesis in ovules from sexual and apomictic plants. HPDEF may be down-regulated as a consequence of apomictic initiation and/or its down-regulation may facilitate progression of apomictic events. PMID- 10872223 TI - A survey of proteinases active during meiotic development. AB - Microsporogenesis in Lilium longiflorum Thunb. is a naturally synchronous process and affords a system in which to study stage-specific events of meiosis and anther development. Zymogram gel analyses were conducted with extracts from a variety of stages of anther development to identify proteinases which likely play roles in anther metabolism. These experiments revealed that several proteinases are present at different stages of anther development, and class-specific inhibitors were used to classify these enzymes. Proteolytic activities increased as anther development proceeded and these activities were temporally correlated with the apoptotic events which precede dehiscence, as well as with events crucial for the maturation of viable pollen. PMID- 10872224 TI - Changes in carotenoids, tocopherols and diterpenes during drought and recovery, and the biological significance of chlorophyll loss in Rosmarinus officinalis plants. AB - Two-year-old rosemary (Rosmarinus officinalis L.) plants were subjected to severe stress by exposure to prolonged drought during a Mediterranean summer. Severely stressed plants recovered completely after the autumn rainfalls although the relative water content remained below 35% for 3 months and the chlorophyll content of leaves was reduced by up to 85% during the drought. In severe stress: (i) alpha-tocopherol increased 9-fold per g dry weight and 20-fold per unit of chlorophyll; (ii) lutein and beta-carotene contents decreased on a dry-weight basis, but an 80% increase in lutein and constant levels of beta-carotene were observed on a chlorophyll basis; (iii) there were transient and sustained increases in the de-epoxidation state of the xanthophyll cycle; and (iv) the highly oxidised abietane diterpene isorosmanol increased 8-fold as a result of the oxidation of carnosic acid. With the autumn rainfalls, water status, alpha tocopherol and violaxanthin recovered first and the levels of photosynthetic pigments and abietane diterpenes increased later. The photoprotection conferred by the xanthophyll cycle and the antioxidant function of tocopherols, lutein and diterpenes may help to avoid irreversible damage in severe drought, making possible the recovery of functional membranes after the autumn rainfalls. Besides, chlorophyll loss reduces the amount of photons absorbed by leaves, which enhances the photoprotective and antioxidant capacity of leaves per amount of photons absorbed, since the ratios of xanthophylls, alpha-tocopherol and abietane diterpenes to chlorophyll increase. PMID- 10872225 TI - Intracellular chloroplast photorelocation in the moss Physcomitrella patens is mediated by phytochrome as well as by a blue-light receptor. AB - The light-induced intracellular relocation of chloroplasts was examined in red light-grown protonemal cells of the moss Physcomitrella patens. When irradiated with polarized red or blue light, chloroplast distribution in the cell depended upon the direction of the electrical vector (E-vector) in both light qualities. When the E-vector was parallel to the cross-wall (i.e. perpendicular to the protonemal axis), chloroplasts accumulated along the cross-wall; however, no accumulation along the cross-wall was observed when the E-vector was perpendicular to it (i.e. parallel to the protonemal axis). When a part of the cell was irradiated with a microbeam of red or blue light, chloroplasts accumulated at or avoided the illumination point depending on the fluence rate used. Red light of 0.1-18 W m-2 and blue light of 0.01-85.5 W m-2 induced an accumulation response (low-fluence-rate response; LFR), while an avoidance response (high-fluence-rate response; HFR) was induced by red light of 60 W m-2 or higher and by blue light of 285 W m-2. The red-light-induced LFR and HFR were nullified by a simultaneous background irradiation of far-red light, whereas the blue-light-induced LFR and HFR were not affected at all by this treatment. These results show, for the first time, that dichroic phytochrome, as well as the dichroic blue-light receptor, is involved in the chloroplast relocation movement in these bryophyte cells. Further, the phytochrome-mediated responses but not the blue-light responses were revealed to be lost when red-light-grown cells were cultured under white light for 2 d. PMID- 10872226 TI - Blue-light regulation of phytoene dehydrogenase (carB) gene expression in Mucor circinelloides. AB - The carB gene, encoding the phytoene dehydrogenase of Mucor circinelloides, was isolated by heterologous hybridisation with a probe derived from the corresponding gene of Phycomyces blakesleeanus. The cDNA and genomic copies complemented phytoene dehydrogenase defects in Escherichia coli and in carB mutants of M. circinelloides, respectively. Fluence-response curves for transcript accumulation were constructed after different blue-light pulses. The level of carB mRNA accumulation reached values up to 150-fold higher than basal levels in darkness. Several elements in the promoter of this gene resemble a consensus sequence identified in Neurospora crassa (APE) which is essential for blue-light regulation. Comparison of the available phytoene dehydrogenase sequences from plants, fungi, algae and bacteria suggests that the two known types of phytoene dehydrogenase are more closely related to each other than previously thought. PMID- 10872227 TI - Light-harvesting complex II pigments and proteins in association with Cbr, a homolog of higher-plant early light-inducible proteins in the unicellular green alga Dunaliella. AB - Like higher plants, unicellular green algae of the genus Dunaliella respond to light stress by enhanced de-epoxidation of violaxanthin and accumulation of Cbr, a protein homologous to early light-inducible proteins (Elips) in plants. Earlier studies indicated that Cbr was associated with the light-harvesting complex of photosystem II (LHCII) and suggested it acted as a zeaxanthin-binding protein and fulfilled a photo-protective function (Levy et al. 1993, J. Biol. Chem. 268: 20892-20896). To characterize the protein-pigment subcomplexes containing Cbr in greater detail than attained so far, thylakoid membranes from Dunaliella salina grown in high light or normal light were solubilized with dodecyl maltoside and fractionated by isoelectric-focusing. Analysis of the resolved LHCII subcomplexes indicated preferred associations among the four LHCIIb polypeptides and between them and Cbr: subcomplexes including Cbr contained one or two of the more acidic of the four LHCIIb polypeptides as well as large amounts of lutein and zeaxanthin relative to chlorophyll a/b. After sucrose gradient centrifugation, Cbr free of LHCIIb polypeptides was detected together with released pigments; this Cbr possibly originated in subcomplexes dissociated in the course of the analysis. These results agree with the conclusion that Cbr is part of the network of LHCIIb protein-pigment complexes and suggest that the role played by Cbr involves the organization and/or stabilization of assemblies highly enriched in zeaxanthin and lutein. Such assemblies may function to protect PSII from photodamage due to overexcitation. PMID- 10872228 TI - Submergence induces expansin gene expression in flooding-tolerant Rumex palustris and not in flooding-intolerant R. acetosa. AB - Ethylene-enhanced leaf elongation upon submergence is part of the survival mechanism of Rumex palustris Sm. plants that grow in frequently flooded areas. Other Rumex species, like R. acetosa L., do not possess this ability and can therefore only survive in habitats that are not frequently inundated. Expansins are proteins that induce extension of isolated cell walls, and therefore might play a role in the stimulation of petiole elongation, also in Rumex. We report here on the identification of several gene sequences encoding for alpha-expansins in R. palustris and R. acetosa plants. The pattern of transcript accumulation of one of these genes, Rp-EXP1, could be correlated with the pattern of leaf elongation in R. palustris after submergence or ethylene treatment. Induction of expansin gene activity was not found in R. acetosa upon these treatments, indicating that ethylene induces the expression of expansin genes in leaves of species that exhibit flooding-induced shoot elongation. PMID- 10872229 TI - Artificially increased ascorbate content affects zeaxanthin formation but not thermal energy dissipation or degradation of antioxidants during cold-induced photooxidative stress in maize leaves. AB - Infiltrating detached maize (Zea mays L.) leaves with L-galactono-1,4-lactone (L GAL) resulted in a 4-fold increase in the content of leaf ascorbate. Upon exposure to high irradiance (1000 mumol photons m-2 s-1) at 5 degrees C, L-GAL leaves de-epoxidized the xanthophyll-cycle pigments faster than the control leaves; the maximal ratio of de-epoxidized xanthophyll-cycle pigments to the whole xanthophyll-cycle pool was the same in both leaf types. The elevated ascorbate content, together with the faster violaxanthin de-epoxidation, did not affect the degree of photoinhibition and the kinetics of the recovery from photoinhibition, assayed by monitoring the maximum quantum efficiency of photosystem II primary photochemistry (Fv/Fm). Under the experimental conditions, the thermal energy dissipation seems to be zeaxanthin-independent since, in contrast to the de-epoxidation, the decrease in the efficiency of excitation energy capture by open photosystem II reaction centers (F'v/F'm) during the high irradiance treatment at low temperature showed the same kinetic in both leaf types. This was also observed for the recovery of the maximal fluorescence after stress. Furthermore, the elevated ascorbate content did not diminish the degradation of pigments or alpha-tocopherol when leaves were exposed for up to 24 h to high irradiance at low temperature. Moreover, a higher content of ascorbate appeared to increase the requirement for reduced glutathione. PMID- 10872230 TI - Isolation and characterization of a jacalin-related mannose-binding lectin from salt-stressed rice (Oryza sativa) plants. AB - A novel plant lectin was isolated from salt-stressed rice (Oryza sativa L.) plants and partially characterized. The lectin occurs as a natural mixture of two closely related isoforms consisting of two identical non-covalently linked subunits of 15 kDa. Both isoforms are best inhibited by mannose and exhibit potent mitogenic activity towards T-lymphocytes. Biochemical analyses and sequence comparisons further revealed that the rice lectins belong to the subgroup of mannose-binding jacalin-related lectins. In addition, it could be demonstrated that the lectins described here correspond to the protein products of previously described salt-stress-induced genes. Our results not only identify the rice lectin as a stress protein but also highlight the possible importance of protein-carbohydrate interactions in stress responses in plants. PMID- 10872231 TI - 12-Oxophytodienoate reductase 3 (OPR3) is the isoenzyme involved in jasmonate biosynthesis. AB - In addition to OPR1 and OPR2, two isoenzymes of 12-oxophytodienoate reductase, a third isoform (OPR3) has recently been identified in Arabidopsis thaliana (L.) Heynh. The expression of the OPR3 gene is induced not only by a variety of stimuli, such as touch, wind, wounding, UV-light and application of detergent, but also by brassinosteroids. The three enzymes were expressed in a functional form in Escherichia coli, and OPR2 was additionally expressed in insect cell cultures and overexpressed in A. thaliana. Substrate conversion was analyzed using a stereospecific assay. The results show that OPR3 effectively converts the natural (9S,13S)-12-oxophytodienoic acid [Km = 35 microM, Vmax 53.7 nkat (mg protein)-1] to the corresponding 3-2(2'(Z)-pentenyl) cyclopentane-1-octanoic acid (OPC-8:0) stereoisomer while OPR1 and OPR2 convert (9S,13S)-12-oxophytodienoic acid with greatly reduced efficiency compared to OPR3. Thus, OPR3 is the isoenzyme relevant for jasmonate biosynthesis. PMID- 10872232 TI - Metabolic responses in cucumber (Cucumis sativus L.) roots under Fe-deficiency: a 31P-nuclear magnetic resonance in-vivo study. AB - The metabolic responses occurring in cucumber (Cucumis sativus L.) roots (a strategy-I plant) grown under iron-deficiency conditions were studied in-vivo using 31P-nuclear magnetic resonance spectroscopy. Iron starvation induced activation of metabolism leading to the consumption of stored carbohydrates to produce the NAD(P)H, ATP and phosphoenolpyruvate necessary to sustain the increased activity of the NAD(P)H:Fe(3+)-reductase, the H(+)-ATPase (EC 3.6.1.35) and phosphoenolpyruvate carboxylase (EC 4.1.1.31). Activation of catabolic pathways was supported by the enhancement of glycolytic enzymes and concentrations of the metabolites glucose-6-phosphate and fructose-6-phosphate, and by enhancement of the respiration rate. Moreover, Fe-deficiency induced a slight increase in the cytoplasmic (pHc) and vacuolar (pHv) pHs as well as a dramatic decrease in the vacuolar phosphate (Pi) concentration. A comparison was done using fusicoccin (FC), a fungal toxin which stimulates proton extrusion. Changes in pHc and pHv were measured after addition of FC. Under these conditions, a dramatic alkalinization of the pHv of -Fe roots was observed, as well as a concomitant Pi movement from the vacuole to the cytoplasm. These results showed that Fe starvation was indeed accompanied by the activation of metabolic processes useful for sustaining the typical responses occurring at the plasma-membrane level (i.e. increases in the NAD(P)H:Fe(3+)-reductase and H(+) ATPase activities) as well as those involved in the homeostasis of pHc. The decrease in vacuolar Pi levels induced by Fe-deficiency and FC and movement of Pi from the vacuole to the cytoplasm suggest a possible involvement of this compound in the cellular pH-stat system. PMID- 10872233 TI - Isoforms of chalcone synthase in Daucus carota L. and their differential expression in organs from the European wild carrot and in ultraviolet-A irradiated cell cultures. AB - Two isoforms of chalcone synthase (CHS) were isolated from cDNA libraries derived from UV-A-irradiated anthocyanin-accumulating (DCb) and non-accumulating (DCs) cell cultures of carrot (Daucus carota L.). The clones designated as DcCHS1, which were present only in the DCb library, had a deduced primary sequence of 389 amino acids and an expected molecular mass of 42.7 kDa, and seem to be alleles of those cloned by Ozeki et al. (1993). The second isoform (DcCHS2) was present in both libraries. It had the highest degree of similarity (97.7%) to parsley CHS over all 397 amino acids. The expected molecular mass of the corresponding protein was 43.6 kDa. Results obtained from Southern blot analysis indicated the existence of at least two CHS genes in carrot. A transient enhancement of the DcCHS1 mRNA level after continuous irradiation with UV-A light could only be observed in anthocyanin-accumulating cultures, whereas an increase in DcCHS2 mRNA was seen in both cell lines. The maximum accumulation of CHS mRNA occurred 48 h after the onset of UV-A irradiation. In the European wild carrot the accumulation of DcCHS1 mRNA was restricted to the red central flowers, whereas the DcCHS2 mRNA was detectable in all red and white petals, as well as leaves, but was absent in stems and roots. The expression of DcCHS1 was restricted to anthocyanin accumulating cells or organs. The heterologous expression of both cDNAs in Escherichia coli resulted in immunostainable bands of different sizes on the Western blot and high levels of catalytic CHS activity. PMID- 10872235 TI - Cloning and expression of UDP-glucose: flavonoid 7-O-glucosyltransferase from hairy root cultures of Scutellaria baicalensis. AB - A cDNA encoding UDP-glucose: baicalein 7-O-glucosyltransferase (UBGT) was isolated from a cDNA library from hairy root cultures of Scutellaria baicalensis Georgi probed with a partial-length cDNA clone of a UDP-glucose: flavonoid 3-O glucosyltransferase (UFGT) from grape (Vitis vinifera L.). The heterologous probe contained a glucosyltransferase consensus amino acid sequence which was also present in the Scutellaria cDNA clones. The complete nucleotide sequence of the 1688-bp cDNA insert was determined and the deduced amino acid sequences are presented. The nucleotide sequence analysis of UBGT revealed an open reading frame encoding a polypeptide of 476 amino acids with a calculated molecular mass of 53,094 Da. The reaction product for baicalein and UDP-glucose catalyzed by recombinant UBGT in Escherichia coli was identified as authentic baicalein 7-O glucoside using high-performance liquid chromatography and proton nuclear magnetic resonance spectroscopy. The enzyme activities of recombinant UBGT expressed in E. coli were also detected towards flavonoids such as baicalein, wogonin, apigenin, scutellarein, 7,4'-dihydroxyflavone and kaempferol, and phenolic compounds. The accumulation of UBGT mRNA in hairy roots was in response to wounding or salicylic acid treatments. PMID- 10872234 TI - Identification of possible signal transduction components mediating light induction of the Gsa gene for an early chlorophyll biosynthetic step in Chlamydomonas reinhardtii. AB - Light-induced expression of the Gsa gene encoding the heme and chlorophyll biosynthetic enzyme glutamate 1-semialdehyde aminotransferase in Chlamydomonas reinhardtii was previously shown to involve Ca2+ and calmodulin (CaM) (C. lm et al. 1996, Plant Cell 8: 2245-2253). To further analyze the signal transduction pathway for light-induced Gsa expression, the effects of several pharmacological agents were examined. Treatment of light-dark synchronized cells with the heterotrimeric G-protein agonist Mas-7 caused partial induction of Gsa in the dark. The phospholipase C inhibitor U73122 inhibited light induction of Gsa. Exposure of cells to light caused a sustained 3-fold increase in cellular D inositol 1,4,5-trisphosphate (InsP3) concentration. KN-93, a specific inhibitor of Ca2+/CaM-dependent protein kinase II, inhibited light induction of Gsa. In contrast, cyclosporin A, a specific inhibitor of the Ca2+/CaM-dependent phosphoprotein phosphatase calcineurin, did not affect light induction of Gsa. These results, together with the earlier results, suggest the involvement of a canonical signal transduction pathway for light-regulated Gsa expression that involves a heterotrimeric G-protein activation, phospholipase C-catalyzed InsP3 formation, InsP3-dependent Ca2+ release, and activation of a downstream signaling pathway through a Ca2+/CaM-dependent protein kinase. PMID- 10872237 TI - Expansion of transgenic tobacco protoplasts expressing pumpkin ascorbate oxidase is more rapid than that of wild-type protoplasts. AB - When pumpkin (Cucurbita spp., cv. Ebisu Nankin) ascorbate oxidase cDNA was introduced into cultured cells of tobacco BY-2 (Nicotiana tabacum L. cv. Bright Yellow No. 2) by Agrobacterium-mediated transformation, the transgenic cells expressed and secreted the recombinant pumpkin ascorbate oxidase into the culture medium. These transgenic cells showed no morphological difference from wild-type cells. However, in the presence of applied hormones protoplasts prepared from the transgenic cells elongated more rapidly than those of wild-type cells. We propose that ascorbate oxidase may play a key role in the regulation of cell expansion perhaps by controlling transport processes through the plasma membrane, but not by affecting the cell wall. PMID- 10872236 TI - Stomatal opening by fusicoccin is accompanied by depolymerization of actin filaments in guard cells. AB - Actin in guard cells is assembled in a radial pattern when stomata are induced to open under light, but the filaments are disassembled when stomata are closed under darkness or by abscisic acid (S.-O. Eun and Y. Lee, 1997, Plant Physiol. 115: 1491-1498). To test if signals that open stomata commonly generate the polymerized form of actin in guard cells, leaves of Commelina communis L. were treated with a potent stomatal opening agent, fusicoccin, and the actin organization examined by immunolocalization techniques. When stomata were induced to open by fusicoccin, hardly any of the filamentous form of actin was detected; instead, the actin resembled that present in guard cells that had been treated with an antagonist to actin filaments, cytochalasin D, and showed a sharp contrast to the long filaments developed in illuminated guard cells. Furthermore, treatment of illuminated leaves with fusicoccin disintegrated actin filaments that had already been formed in the guard cells. Preincubation of leaves with phalloidin, which interferes with fusicoccin-induced actin depolymerization, delayed fusicoccin-induced opening during the early phase. These observations suggest that the prevention of actin filament formation and/or depolymerization of actin filaments may accelerate the stomatal opening process in response to fusicoccin. PMID- 10872238 TI - An actuarial procedure for assessing risk with juvenile sex offenders. AB - Assessments of juvenile sexual offenders that are intended to aid in dispositional decisions occur at a multitude of decision points within the juvenile justice system. Despite the ubiquity of decisions that include considerations of risk, relatively little empirical work has been done on the development and validation of a risk assessment procedure for these young offenders. In this article, we discuss our initial efforts in developing and validating an actuarial risk assessment protocol for juvenile sex offenders using a sample of 96 adolescents that had been admitted, treated, and discharged from the Joseph J. Peters Institute. We conclude with a critical discussion of problems associated with evaluating risk in this population, and of deficiencies and revision requirements in the present protocol. PMID- 10872239 TI - Problems with the DSM-IV diagnosis of pedophilia. AB - This paper examines the taxonomic adequacy of the Diagnostic and Statistical Manual, 4th ed., DSM-IV (American Psychiatric Association, 1994) diagnostic category of pedophilia. This diagnosis, as well as the other sexual disorders, have been ignored in DSM field trials. There is no empirical information about the reliability or validity of this diagnosis. Moreover, because the vagueness of the diagnostic criteria, clinicians would need to make inferences that would likely lead to reliability problems in diagnosis. Further, the DSM diagnostic criteria include constructs that are not intersubjectively verifiable and for which there are no valid measures. This can also lead to lack of diagnostic reliability and accuracy. Most problematical however, there are aspects of the diagnostic criteria, most notably the presence of an "ego dystonic sexual attraction to children," that are incorrect exclusion criteria. Suggestions for improvement are provided. PMID- 10872240 TI - Psychometric analysis of the Sexual Interest Cardsort Questionnaire. AB - Reliability, measured by Cronbach's coefficient alpha, and concurrent validity, measured by Pearson's r and polychoric correlation coefficients, were evaluated in this study. A sample of 371 sexual offenders referred to the Behavioral Medicine Institute of Atlanta for evaluation of sexual interests and behaviors by the courts were assessed using the Sexual Interest Cardsort Questionnaire (SI), a self-report measure of deviant and nondeviant sexual interest, as well as indicator variables obtained from classifications assigned by clinicians as a result of 2 hour-long, semistructured clinical interviews. Internal consistency of 75 items from the SI ranged from 0.71 to 0.96, across 15 categories of sexual interest and behavior. Additionally, the SI was shortened utilizing Cronbach's alphas to maintain a high level of internal consistency. The resulting questionnaire, the shortened SI (SIS), had 45 items and 15 categories. Cronbach's alpha ranged from 0.78 to 0.97. Utilizing Pearson's r and polychoric correlation coefficients, significant correlations were found for the 11 sexually deviant categories of the SI and indicator variables, and the 10 sexually deviant categories of the SIS and indicator variables. The SI and SIS showed a high level of reliability and concurrent validity. Clinical and research issues pertaining to the clinical assessment of male sexual offenders utilizing self-report and clinical interview data, both obtained as the result of comprehensive evaluations, are discussed. PMID- 10872241 TI - The impact of polygraphy on admissions of victims and offenses in adult sexual offenders. AB - Sexual offenders are extremely reluctant to disclose their offending histories for a variety of psychosocial and legal reasons. The polygraph has shown promise as a intervention for eliciting admissions of past sexual offending behaviors. For 60 adult male sexual offender (35 inmates and 25 parolees), the number of victims and offenses were recorded from the Presentence Investigative Report, Sexual History Disclosure form, and 2 consecutive polygraph examination reports. Dramatic increases in the number of admitted victims and offenses were found for inmates, but not for parolees, across each source. However, there was a substantial decline in the number of victim and offense admissions by the second polygraph examination for both groups, even though 80% of the examination results reveled deception about sexual offending behaviors. Standardized use of sanctions and privileges for deceptive and nondeceptive polygraph results, respectively, are proposed as a way of eliciting full disclosure of offending histories for these offenders. PMID- 10872242 TI - Outcome of an institutional sexual offender treatment program: a comparison between treated and matched untreated offenders. AB - Data from a sexual offender treatment program operated by the Correctional Service of Canada at the Regional Psychiatric Center (Saskatoon) supported the conclusion that cognitive behavioral treatment can reduce sexual offense recidivism. The study compared 296 treated and 283 untreated offenders followed for a mean of 6 years after their release. An untreated comparison subject was located for each treated offender on three dimensions: (a) age at index offense, (b) date of index offense, and (c) prior criminal history. Data were analyzed using tests of proportion, survival analysis, and analysis of offender Criminal Career Profiles. Over a mean follow-up period of almost 6 years, convictions for new sexual offenses among treated offenders were 14.5% versus 33.2% for untreated offenders. During the follow-up period, 48% of treated offenders remained out of prison compared to 28.3% of untreated offenders. Time series comparisons of treated and comparison samples also showed that treated men reoffended at significantly lower rates after 10 years. A Criminal Career Profile (CCP) was constructed by taking the Age at First Conviction and plotting the offender's successive lengths of time free against time incarcerated. Pre- and posttreatment slopes of the CCP were lower for both groups posttreatment; however, the degree of change was significantly greater for the treated group, indicating a greater reduction in criminal activity among these offenders. Taken together, the results of all three analytic techniques supported the efficacy of appropriate correctional treatment for effective reduction of recidivism. PMID- 10872243 TI - The importance of meeting research standards: a reply to Fischer and Smith's articles on the Abel assessment for sexual interest. PMID- 10872244 TI - [Molecular genetic principles of tumor development and progression]. PMID- 10872245 TI - [Molecular genetic and cell biology principles for the development of malignant tumors]. AB - The development of cancer is one of the most intensively studied areas of medical research resulting in an immense quantity of data. Therefore, the purpose of this article is to give an overview of the basic principles of cancer development. Key words such as multi-step carcinogenesis, cell cycle, protooncogene, tumor suppressor gene, DNA repair gene, apoptosis and telomeres are explained and described in examples. This paper aims to connect recent information of molecular and cellular biology in an overview of cancer origin and development. PMID- 10872246 TI - [Molecular genetic principles of progression of malignant diseases]. AB - During the past decade, the molecular mechanisms in the process of tumor progression, including metastasis and angiogenesis, have become better understood. Cancer metastasis consists of multiple, complex interacting steps. Each of these steps is crucial and limiting, since a failure to complete any one prevents the tumor cell from producing a metastasis. Detachment from the solid tumor by loosening the intercellular junctions and proteolysis of the extracellular matrix enables tumor cells to enter blood- and lymph vessels. The intravasation into the circulation is supported by the secretion of angiogenic factors, which induce degradation of the basal membrane in blood vessels. Adhesion to endothelial cells, extravasation from the circulation, and induction of angiogenesis are further essential steps for completing the metastatic process. Furthermore, it is well known that once a tumor cell has entered circulation, it will survive only by evasion of the immune system. The systematic identification of tumor antigens opens up new possibilities for immunotherapeutic approaches. PMID- 10872247 TI - [Pathogenetic and clinical aspects of low estradiol level in the man]. AB - To elucidate a possible role of low estradiol (E2) levels in blood serum of men, normal values were determined in 91 healthy men (age 20-75 years), classified as high or low complaint-index due to a psychological questionnaire. Statistical analysis gave no correlation of estradiol levels to age or complaint index in normal men whereas testosterone (T) could be significantly correlated to complaint-index (p < 0.01) and free testosterone (fT) could be significantly correlated to age (p < 0.001) and complaint-index (p < 0.01). T and E2 were determined in 1370 clinical patients with various urological diseases, T, fT and E2 in 1261 ambulant patients. In 72/1370 (5.2%) and 76/1261 (6%) patients, low E2 levels (< 10 pg/ml) were found in blood serum. In 56/76 (74%) patients with low E2-levels, T or fT was simultaneously low. Isolated low E2-levels were found in 20/1261 (1.6%) patients. In clinical patients, no special urological disease correlated to low E2-levels. Due to these results, low E2 levels in men are in most cases the result of low testosteron levels. The adequate hormonal treatment in men is therefore the replacement of testosteron. Substitution of E2 in men is at that time an experimental therapy, that is limited on selected cases. PMID- 10872248 TI - [Volumetry of the urinary bladder with implantable ultrasound sensors]. AB - Experimental studies revealed that the contractile response of the urinary bladder to sacral anterior root stimulation depends on the actual bladder volume. Furthermore, no clinical relevant technique is available for continuous monitoring of the bladder wall distension respectively bladder volume in paraplegic patients. The presented study investigates the reliability of especially developed implantable ultrasound sensors as a sensoric system for continuous monitoring of the bladder volume. In six anaesthesized pigs two ultrasound sensors, one transmitter and one receiver, were implanted on the bladder wall at different locations (latero-lateral, dorsal-ventral, rostral caudal). After closing the abdominal wall, the bladder was filled in 50 ml steps up to 250 ml. After each filling step the running time of the ultrasound signal was measured. In all experiments reproducible results and a high correlation of the measured running times with bladder volume were observed. The latero-lateral configuration of the sensors seemed to be most confidential. The presented study indicates that bladder volumetry with implantable ultrasound sensors is possible with minimal technical prerequisites. This promising technique for continuous bladder volumetry could play an important role in the development of an intelligent and autoadaptive neurostimulator of the urinary bladder in paraplegic patients. PMID- 10872249 TI - [Microsurgical vasovasostomy in the age of modern reproduction medicine. A cost benefit analysis]. AB - WS represents the standard procedure of choice for the treatment of obstructive azoospermia following vasectomy. However, recently, ICSI has been suggested by some to represent the solution for all cases of male factor infertility regardless of its etiology based on its success rates. Therefore, we compared VVS to MESA/TESE and ICSI in terms of pregnancy, complications, and costs. Between 1/93 and 6/98 157 VVS was performed microsurgically using the 2-layer technique in 157 patients following prior vasectomy. Between 9/94 and 9/97 69 couples underwent MESA/ICSI for epididymal obstruction not amenable to micro-surgical reconstruction such as post-inflammatory obstruction and congenital absence of the vas deferens; in the same time period 42 couples underwent TESE/ICSI for azoospermia of testicular origin due to cryptorchidism, testicular atrophy, obstruction of the rete testis. In most cases MESA or TESE and ICSI were performed metachronously. Mean intervall of vasal obstruction was 7.6 (0.5-18) years; patency after VVS was 77%, pregnancy rate was 52%. Local complication rate was 4.7%, no major complications were observed. Costs per life birth after VVS were as high as 5,447,-DM or 2,800 Euro. Pregnancy rates after MESA/TESE and ICSI were 22.5% and 19.5%, respectively with 16 singletons, 3 twins and 3 abortions; local complications occurred in 3.9% of the men. Multiple birth were noticed in 15.8% following ICSI, but only in 0.7% following VVS. 5.7% and 1.4% of the female partners experienced serious complications as a mild or severe ovarian hyperstimulation-syndrome, respectively. Costs per life birth after MESA/TESE cycle were as high as 28,804,-DM or 14,100 Euro. Even in the era of ICSI microsurgical vasovasostomy represents the standard approach for obstructive azoospermia following vasectomy. Based on a cost-benefit analysis VVS is more successful in terms of pregnancy rates (52% vs. 22.5%). We conclude that MESA/ICSI should be reserved for patients not amenable for microsurgical reconstruction. PMID- 10872250 TI - [Long-term outcome of endoscopic treatment of vesicoureteral reflux]. AB - For more than 10 years VUR has been treated by endoscopic injections with biocompatible substances. This study presents the results of this procedure with bovine collagen (Zyplast) in 42 children with 66 refluxing ureters. The follow-up period comprises an average time of 6.5 years (2-10 years). In 16 ureters a second and in 4 a third injection was given. In 5 kidneys scarring occurred during the years of follow up. Growth, kidney function and blood pressure remained normal in all children. 6 ureters were reimplanted because of recurrences, in 13 ureters (23%) the reflux improved and since no infection recurred no further management became necessary. In 36 (65.4%) ureters reflux was cured and in further 11 this was supposedly the case. However this was not proven by MCU, because parents refused this investigation. Earlier MCU have shown no reflux. We think that the endoscopic injection with collagen is a excellent option in management of VUR grade I-IV. PMID- 10872251 TI - ["False positive" testicular perfusion in testicular torsion in power Doppler ultrasound]. AB - We report about a patient with acute scrotal pain. Power Doppler sonography demonstrated arterial testicular perfusion. As no amelioration was achieved by conservative treatment, surgical exploration was done after 7 days. Intraoperatively, we found a partial testicular torsion with a 180 degrees rotation of the testis at the rete testis. This case demonstrates, that the detection of intratesticular arterial blood flow cannot exclude testicular torsion. PMID- 10872253 TI - [Contralateral biopsy in testicular tumor. A plea in favor of the procedure]. PMID- 10872252 TI - [Rational therapy]. AB - In a time of scarce resources, the demand for a rational therapy is important in the present discussion to reform the health system. This article describes rational therapy and introduces practical approaches like drug therapy, evidence based medicine, and clinical practice guidelines. The development of efficient cost-benefit relations may lead to more rationalization in the health sector. It is important to attain acceptance also among the patients that rationing in the health system is a necessary measure in diagnosis and therapy. PMID- 10872254 TI - [Therapeutic options in hormone refractory prostate carcinoma]. PMID- 10872255 TI - [Acute scrotum]. PMID- 10872256 TI - [Report at the Annual Meeting of the Kidney Transplantation Working Group of the Graduate and Continuing Education Committee of German Urologists. Lubeck 24-26 June 1999]. PMID- 10872257 TI - e-health: will we be victims once again? PMID- 10872258 TI - Reducing time delay in the thrombolysis of myocardial infarction: an internal quality improvement project. ARIAM Project Group. Analisis del Retraso en Infarto Agudo de Miocardio. AB - The objectives of this study were to improve thrombolytic therapy in acute myocardial infarction by reducing the "door-to-needle" time in a 285-bed university hospital in Spain. A quality management approach was used involving all the relevant staff. Target standard was set at 35 minutes. Baseline data, intervention effect, and continuous monitoring were analyzed using x control charts. Analysis of baseline data showed a wide out-of-control variation and 72 minutes' average delay. Cause analysis revealed organizational and clinical problems that were subjected to intervention. Postintervention data showed a stable process, with an average of 30 minutes. Continuous monitoring showed further improvement in average time and predictable variation. The template of the current control chart has an average of 26 minutes. Quality management methods, particularly staff involvement in problem analysis and intervention design, and the use of control charts were useful to understand, solve, and continuously monitor an important clinical problem whose existence was evident only after it was measured. PMID- 10872259 TI - Patient satisfaction as an indicator of quality care in independent health facilities: developing and assessing a tool to enhance public accountability. AB - The objective of this research was to examine the performance of a brief patient survey about quality of care received in community-based diagnostic and therapeutic facilities. The survey was administered to patients in 44 facilities that were also scheduled for a formal external assessment. The response rate was 53%. Patients generally rated their care positively; 18.5% of patients rated at least 1 item as fair or poor. The amount of information received about risks and complications was rated least favorably; concern and caring shown by staff was rated most favorably. The 10 items which patients rated regarding aspects of quality formed an internally consistent scale (alpha = .93). Patients' ratings were not useful predictors of assessor ratings. Although patients' ratings cannot substitute for expert on-site assessments, they are an important part of a quality management program. The patient survey provides additional, complementary information about components of quality care that are important to them. PMID- 10872260 TI - What's happening in quality improvement at the local hospital: a state-wide study from the Cooperative Cardiovascular Project. AB - The objective of this study was to investigate what happened to improve the quality of care for acute myocardial infarction (AMI) at all 32 nonfederal hospitals in Connecticut and to assess the impact of the Cooperative Cardiovascular Project (CCP) on quality improvement (QI) activities for AMI. We performed a questionnaire study with secondary analyses using the CCP database. On-site interviews were conducted with QI directors at all 32 Connecticut nonfederal hospitals that participated in the Health Care Financing Administration's Cooperative Cardiovascular Project (CCP) in 1992-93 and 1995. The interviews sought information about the makeup of QI departments, specific approaches used to improve the care of patients with AMI, and the perceived value of the CCP to each individual hospital. Results showed that the number of full time equivalents (FTEs) and FTEs per beds employed in QI departments ranged from 1 to 30 and from 0.4 to 7.9, respectively, with a registered nurse most often serving as the department head (27/32). Over half of the departments (17/32) had additional responsibilities. The majority (25/32) used some combination of physician champions, multidisciplinary QI teams, standing orders, or critical pathways to effect change in AMI care. Finally, 26 of the 32 hospitals believed the CCP was valuable because it provided credible benchmark data, a catalyst for change, or a specific focus on processes of care needing improvement in AMI. Despite great variability in institutional resources, all 32 hospitals used a similar combination of QI approaches to effect change in AMI care. However, there is variable scientific evidence supporting these approaches. Externally sponsored projects such as the CCP appear to play a useful role for individual hospitals. Defining the optimal methods of QI is difficult given that hospitals are using complex combinations of nonstandardized improvement interventions. PMID- 10872261 TI - Commentary: quality of care and cost containment are the hospital-based ambulatory surgery challenges for the future. AB - Cost containment and quality of care represent the most important objectives of all health care professionals. Because of its progressive growth over the past decade, ambulatory surgery has become an area where these 2 issues need to be addressed. The goal of this paper is to discuss the economic and quality of care challenges faced by hospitals as they strive to become competitive in the 21st century. The quality of care in ambulatory surgery has been improving because of multidisciplinary activities. Hospitals tend to hire the staff on the basis of their expertise in certain areas, and those personnel do not have to cover other hospital roles. Moreover, the hospital staff is able to seek information at any time from coworkers in other areas of specialty. Ambulatory surgery in a hospital offers advantages, such as multiple operating rooms, multiple skilled health care providers, and the ability to stay overnight if needed. The consolidation of supplies makes it easier to contract for a better price. Aggressive contract negotiations and implementation of cost-effective and cost-efficient strategies are the keys to success in the future. Quality improvement (QI) initiatives and quality of care (QC) indicators need to be developed to address various problems in the ambulatory surgery setting such as unnecessary admissions, inadequate staffing, efficient operating room (OR) utilization, quality of care, and assessment outcome. These initiatives should be addressed at regular meetings where opportunities to improve the ambulatory services are discussed. The number of ambulatory surgery procedures performed each year will continue to increase, although perhaps not at the rate we experienced in the past. Procedures that once were performed in an inpatient setting can now be accomplished on an outpatient basis or even in the physician's office. We will continue to see this shift of volume as technologic advancements and anesthetic agents allow more complex procedures to be performed on an outpatient basis. PMID- 10872262 TI - Clinical performance measurements proposed to compare diabetes care among health care systems. PMID- 10872263 TI - Major Medicaid victory for physicians. PMID- 10872264 TI - The right to unite. PMID- 10872265 TI - Bring back the humanism. PMID- 10872266 TI - Medicine and health care of Bosnia and Herzegovinia on WWW. PMID- 10872267 TI - [Late effects of experimental ablation of the pineal gland on ultrastructural morphometric changes in thymic epithelial cells]. AB - The late effect of surgical ablation of pineal gland on the morphometric changes in epithelial cells of rat thymus were investigated. The aim of this study is to determine a possible existence of sex-different changes in composition of the epithelial cell component unit long after pinealecotmy what could be important for the subtle understanding of mutual correlation between pincal body and thymus. This article presents results of stereological ultrastructure parameters of thymuscortical and medullar epithelial cells of male and female rats two month after pinealectomy. The experimental animals were divide into two groups: an experimental one (pinealectomized) and the control group (shampinealectomized). Pinealectomized animals were submitted to surgical ablation of pineal gland while the control were undertaken the same surgical treatment but without removal of the pineal gland. Animals were sacrificed 60 th days following the surgery treatment. Parasagital pieces of thymus tissue were fixed by means of immersion in glutaraldehyde and prepared for transmission electron microscopy. Using Weibls multipurpose test system and multilevel sampling technique on electron micrographs the volume and surface density of nucleus (Vu, Sv) and cytoplasm of cortical and medullar thymus epithelial cells were calculated. At the different magnification level were established the volume and surface density of mitochondria (Vvm, Svm) endoplasmic reticulum (Vvr, Svr), vacoule(Vuv) as well as numerical density of mitochondria (Nvm). Our analysis has conformed statistically significant increase in Vv of reticulum and vacuole in both sex of pinealectomized rats. Sv of plasmalema, reticulum and mitochondrial membrane are markedly increased in thymus medullar epithelial cells of pienalectomized rats. Vv of mitochondria is significantly increased in cortical epithelial cells of pienalectomized animals. Results allow us to confirm that mutual correlation between pineal gland and thymus exists but present findings seem to support the concept of sex independent inhibitory action of pineal gland on thymus cortical and medullar epithelial cells. PMID- 10872268 TI - [Cytomegalovirus disease in immunocompromised patients]. AB - Human Cytomegalovirus are a ubiquitous herpesvirus establishing virus-infections which are usually asymptomatic in immunocompetent individuals. In patients with Acquired Immunodeficiency Syndrome (AIDS) and immunocompromised hosts, the virus causes primary, latent or chronic persistent infection. Primary CMV infection is very severe in immunocompromised patients as well as among healthy population. Among patients with AIDS Cytomegalovirus (CMV) is usually isolated from patients specimen in association with other pathogens (Pneumocystis carinii, Candida albicans). The prevalence of serious CMV (chorioretinitis, gastrointestinal disease, interstitial pneumonia and central nervous system disease), in AIDS population are respectable. PMID- 10872269 TI - Occurrence of surgical pathogens and their antimicrobial susceptibility patterns. AB - OBJECTIVE: To assess the occurrence frequency of bacterial pathogens at the Surgical Wards, Casualty and Outpatient Department (OP) of major Kuwaiti hospital and to compare their antimicrobial susceptibility patterns. METHODS: The Automicrobic System (bioMerieux-Vitek) with respective ready-to-use cards were used for identification of isolates and their susceptibility testing. Vitek DataTrac software automatically tabulated the occurrence rate of pathogens or their antimicrobial susceptibility percentage--for defined periods of time and specified patient locations. RESULTS: The most common organisms for surgical inpatient isolates were E. coli, S. aureus, K. pneumoniae and P. aeruginosa; but for blood culture and sputum, by far, the most common were coagulase-negative staphylococci and K. pneumoniae, respectively. E. coli from wounds were less susceptible (for 12-26%) to ampicillin, co-amoxiclave, cephalothin and cefuroxime than surgical inpatient, OP or casualty E. coli isolates. Cephalothin and piperacillin susceptibility rates of K. pneumoniae of surgical in- and outpatients were twice higher than that of respective E. coli isolates. CONCLUSION: The occurrence frequency of bacterial pathogens were dependent on the surgical services. Overall, antimicrobial susceptibility rates were high for different surgical subcategories, especially for casualty and inpatients. The lowest susceptibility rate showed the wound isolates versus beta-lactams, except third generation cephalosporins. PMID- 10872270 TI - [Postoperative left ventricular function in patients with transposition of the great blood vessels]. AB - During August 1996 to August 1998 at the Paediatric Clinic in Sarajevo, 8 patients (pts) have been diagnosed to have a Transposition of the Great Arteries (TGA), age 10 hrs to 31 days. First Group (n = 4) had a simple TGA and in II Group in 3 pts TGA was associated with double inlet left ventricle (DILV) and subpulmonary artery stenosis and in 1 with double outlet right ventricle (DORV) and subpulmonary artery stenosis. Anatomical correction of TGA-arterial switch has been performed in Group I, mean age 15.5 days (7-18). In Group II palliative correction has been completed in: atrioseptectomy (1/4), pulmonary artery banding (3/4), right Blalock-Tausing modified shunt and also partial correction: Glenn anastomosis, mean age 4.7 months. Pts have been followed from 3 to 19 months postoperatively. All pts are well, except 1 pt who died following the arterial switch (mortality rate 12.5%). The aim of this study is to evaluate left ventricle (LV) function pre and postoperatively with electrocardiographic monitoring. Electrocardiographically there was no significant rhythm disorders. Using M mode echocardiography techniques, LV function was measured including internal dimensions of the LV as well as a wall thickness and than compared with the others comparable to the age and body weight. LV function in pts post anatomical correction has returned to normal values faster, with statistically significant difference of p = 0.02, than in pts post palliative-partial correction. CONCLUSION: Echocardiographically LV function in pts with TGA post arterial switch returned faster to normal values than in pts following the palliative-partial correction. PMID- 10872272 TI - [Reasons for reoperation after laparoscopic cholecystectomy]. AB - AIMS: Analysis of the reasons for reoperation after laparoscopic cholecystectomy. METHODS AND PATIENTS: Retrospective-prospective analyses of the first 250 patients who undergone laparoscopic cholecystectomy. In 86% cases indication for operation was chronic calculosis of gallbladder. RESULTS: Reoperation was performed at 6 patients (2.4%). The reasons of reoperation were: haematoma of gallbladder's loge (1), biliary fistulas (1), biliary peritonitis (1), abdominal abscesses (2), and perforated peptic ulcer (1). At 2 patients with intraabdominal abscesses, it was solved by laparoscopic drainage. The other complications were solved with laparotomy, also. We did not have lethal cases after reoperation. CONCLUSION: Rate of postoperative complications was 2.4%, and all of them required reoperation. Our results are similar with results of the other authors. PMID- 10872271 TI - [Effect of pantoprazole, amoxicillin and metronidazole treatment on the level of H. pylori eradication and the histological image of antral gastritis in patients with duodenal ulcer]. AB - BACKGROUND: Relationships between Helicobacter pylori infection, inflammatory changes in antral region of gastric mucosa, and duodenal ulcer is well known and documented in a large number of studies. This trial is designed to examine effect of one week regimen of Pantoprazole, Amoxycillinum and Metronidazol to eradication of H. pylori, duodenal ulcer healing and histological changes on gastric mucosa. PATIENTS AND METHODS: 30 patients with active duodenal ulcer, H. pylori-positive, 16 male, with average age 47.12 +/- 13.13 yrs (AVG +/- STD) and 14 female patients with average age 44.47 +/- 12.29 yrs were included in trial. Biopsy of gastric antral region were performed in each patient. Patients were given Pantoprazole 40 mg bid, Amoxycillinum 1000 mg bid, Metronidazolum 500 mg bid for 7 days. After 7 days of treatment, control endoscopy was performed with repeated rapid ureasa test for H. pylori and antral biopsy and with verification of duodenal ulcer healing. Patients were followed up for 24.3 +/- 9.7 weeks for occasion of subjective symptoms. RESULTS: 96.67% patients were presented with eradicated H. pylori, complete ulcer healing was found in 83.34% patients after one week regimen (13.33% patients with ulcer reduced on one third of previous described), 73.33% of patients were presented with histologically feature of chronic gastritis turned from active to stationary phase. CONCLUSION: One week regiment with Pantoprazole, Amoxycilline and Metronidazole is effective, and beside a high rate ulcer healing and eradication of H. pylori it provides an improvement of histological feature of antral gastritis. PMID- 10872273 TI - [Changes in fibrinogen, fibrin-fibrinogen degradation products and fibrinolytic activity during pregnancy, labor and the puerperium in pregnant women with chronic hypertension]. AB - In this study 79 pregnant women were analyzed, during the pregnancy, delivery and postnatal period. The control group consisted of 41 healthy women, while in the studied group there were 38 patients with severe chronic hypertension. Analyzed parameters: fibrinogen, fibrinolysis and FDP. Parameters were performed in all cases of pregnancies during third trimester (I), during the first stage of labor (II), immediately after delivery of placenta (III), two hours after placenta was delivered (IV), eight hours (V), 24 hours (VI), 48 hours (VII) and 72 hours (VIII) after placenta was delivered. Comparing certain measures in fibrinogen's value, high statistical significance was obtained for control group (healthy women), ANOVA (F = 5.17; p < 0.001) and for studied group (women for chronic hypertension), (F = 3.17; p < 0.001. Analyses of FDP values showed high statistically significant difference for control group, F = 15.03; p < 0.00001, and for studied group, F = 10.56; p < 0.00001. Statistically significant difference in fibrinolytic activity was obtained for control group, F = 60.72; p < 0.0001 and studied group, F = 5.70; p < 0.0001. PMID- 10872274 TI - [Molecular mechanisms in apoptosis]. AB - Apoptosis is evolutionary conserved form of cell suicide. Tumor necrosis factor alpha (TNF-alpha) or Fas Ligand activated apoptosis by binding of the plasma membrane receptor. The activation of TNF Receptor 1 or Fas-Ligand Receptor lead to activate of caspase 8. The activation of the caspase-8 lead to activate the cell-death machinery cascade. The inhibitor of cell death machinery is Bcl-2 also fails to prevent Bax-induced cytochrome c release, activation of caspase-3, membrane blebbing, nuclear fragmentation, and cell death. Bcl-2 is important cell live-death regulator. Cleavage of specific protein subsets is a key event in the execution of apoptosis. Protein degradation may serve for the structural alterations in the process of cell self-destruction, but it may also function as a switch in the decisions between apoptosis and necrosis or apoptosis and cell proliferation. PMID- 10872275 TI - [Eye injuries caused by an eclipse of the sun]. AB - Two patients with macula damage following sungazing are reported. Visual acuity was damaged, gentle central scotoma in automated perimetry (Octopus) was presented same as lower A wave in ERG and small macular hyperfluorescency in fluorescein angiography. Funduscopy findings were macular changes similar to macular semirupture. In one month all pathologic symptoms disappeared. The only safe prevention is that by Mylar folia that completely prevent eye injury from sungazing. PMID- 10872276 TI - [Sarcoidosis activity in a comparison of the tuberculin test and the endoscopic picture in a randomized clinical sample]. AB - The study population consisted of 100 patients with sarcoidosis, selected by method of accidental selection. According to clinical form of sarcoidosis, the sample was divided, in three groups: group of patients with chronic form of sarcoidosis (SHFS)--36 patients, group of patients with acute form of sarcoidosis (SAFS)--46 patients, and group of patients with asymptomatic form of sarcoidosis (SAsFS)--18 patients. The aim of study was to determine whether there were statistically significant differences between the groups of different sarcoidosis clinical forms in diagnosis time in connection with tuberculin reaction, endoscopic picture and index of sarcoidosis activity? Which level of correlation was between index of sarcoidosis activity and tuberculin reaction, respectively endoscopic picture on observed groups of sarcoidosis patients? On the basis of our results we reached the following conclusions: there was significant correlation between magnitude of tuberculin reaction and index of sarcoidosis activity in SAsFS, in SAFS correlation there was no significance and in SHFS there was no correlation. In all of three groups the most frequent was endoscopic picture of enlarged and irregularly arranged capillaries with yellowish plaques on bronchial mucous with average index of activity (6.7). PMID- 10872277 TI - [TENS in the treatment of muscle spasm]. AB - This study deals with 60 patients with muscle spasm after the lesion of upper motor neurone. Thirty patients were treated by battery operated TENS unit, which produce biphasic impulses, with possibility of individual determination of impulse duration (0.05-0.25 msec), frequency (2-100 Hz) and intensity (0-80 mA). The electrodes were put on the motor points of m. tibialis anterior and m. extensor digitorum. The parameters of stimulation where determined individually, using the impulses that elicit dorsal flexion of the ankle joint and fingers. In the control group, consisted of 30 patients, passive exercises were performed (increasing range of motion, stretching of the agonistic and antagonistic muscles). At the end of the research, the authors report statistically significant decrease of muscles spasm in patients treated by TENS (p < 0.05), as well as improvement of the passive range of motion in ankle joint in both groups. Reducing of the muscle spasm is more pronounced if longer period of stimulation is applied. PMID- 10872278 TI - [Differences in non-verbal behavior in individuals with various types of aphasia]. AB - The aim of this research was to investigate if there were differences in non verbal behaviour at persons with different forms of aphasia while resolving certain problems, and time spent for finding solutions to them. The research involved 30 patients with motoric, sensory and senso-motoric aphasia, in the area of 7 non-verbal variables. Using the statistical method (discriminatory analysis) we came up with the results confirming that there is a difference in the quality of problem solution at patients with different forms of aphasia, but the time frame for problem solving does not differ significantly with the same patients. PMID- 10872279 TI - The use of DCS and info systems in protection of environmental health. AB - New millennium will foster advances in different face of information age. The world is already prepared for global Large Seale Networking. Existing PC-Based DCS could be able to achieve higher intelligence, integrity, speed, memory capacity. Currently in this climate IT will develop and demonstrate revolutionary applications in basic science, medicine, environment protection, etc. The main goal of this paper is to show basic architecture and system software functionality at DCS. Totally specific principle and approach to system philosophy needs to be used for next generations at QC with HECC. DCS are helping and it will do in future to overcome limitations of traditional technology to make better environment. In same time IT allow to gain high speed access to scientific and medicine instruments, surgical and other therapeutic interventions and better organisation of primary health care what Bosnian people have to do! The people at Bosnia and Herzegovina needs to design move competitive education and training system. PMID- 10872280 TI - [Use of information technology in the 21st century in the medical sciences]. AB - Development of health-care system depends of using modern IT. The world is already prepared for global Large Seale Networking. Existing PC-Based DCS could be able to achieve higher intelligence, integrity, speed, memory capacity. Currently in this climate IT will develop and demonstrate revolutionary applications in basic science, medicine, environment protection, etc. Totally specific principle and approach to system philosophy needs to be used for next generations at QC with HECC. DCS are helping and it will do in future to overcome limitations of traditional technology to make better environment. In same time IT allow to gain high speed access to scientific and medicine instruments, surgical and other therapeutic interventions and better organisation of primary health care what Bosnian people have to do! PMID- 10872281 TI - [The properties of Pragia fontium bacteria isolated on the territories of Ukraine and the Czech Republic]. AB - The results are presented of the study of the properties (morphological, tinctorial, cultural, biochemical) and primary DNA structure of 28 Pragia fontium strains, 18 of which were isolated in the territory of Ukraine and 10 in the territory of Czechia. The scheme of differentiation of genera Pragia, Budvicia and Leminorella is presented. The GC (guanine + cysteine) content of DNA is 47 +/ 1.5 mol.%. The levels of DNA relatedness of P. fontium strains and the type strain (CNCTC Eb11/82) varied from 84 to 95%. The levels of DNA relatedness of P. fontium and Escherichia coli K-12 strain is 3 to 5%. PMID- 10872282 TI - [The effect of glucose on the fatty acid level in Saccharomyces cerevisiae and Schwanniomyces occidentalis cells]. AB - The influence of glucose on fatty acid contents in the cells of yeast Saccharomyces cerevisiae and Schwanniomyces occidentalis has been studied. It was shown that fatty acid contents in Schw. occidentalis was enhanced with the increase of glucose concentration in the medium. The increase of glucose concentration in the medium during respiro-fermentative growth phase reduces the level of C18 in unsaturated fatty acids of S. cerevisiae; the increase of fatty acids content in total lipids and nonetherified fatty acids was observed under the same conditions in the respiratory phase. It was established that during the growth of S. cerevisiae in the medium with 0.1% glucose the level of nonetherified fatty acids was higher and with 4% glucose--lower in comparison with Schw. occidentalis. The correlation of nonetherified and etherified fatty acid levels in S. cerevisiae is higher in comparison with Schw. occidentalis grown under the same conditions. PMID- 10872283 TI - [The effect of glucose on the biosynthesis of extracellular enzymes by Streptomyces recifensis var. lyticus 2435 and its mutants]. AB - Different glucose concentrations have been studied for their effect on biosynthesis of bacterio- and proteolytic enzymes in Streptomyces recifensis var. lyticus 2435 and strains II-29 and 2P-15 obtained from the latter. It has been established that synthesis of enzymes by the parent strain 2435 was subjected to catabolite repression: synthesis of proteolytic and staphylolytic enzymes was the most sensitive to glucose, synthesis of glucosidases was less sensitive. The role of catabolite repression in regulation of synthesis of bacteriolytic enzymes to the latter in strain II-29 and rifamycin-resistant mutant 2P-15 is essentially decreased. As compared with the initial strains, the high level of activity of lytic enzymes in the strain 2P-15 is achieved at higher glucose concentrations in the fermentation medium, synthesis of staphylolysins and glucosideases are inhibited inconsiderably at 2% and 4% content of glucose, in medium respectively and it does not decrease under further increase of glucose concentration. PMID- 10872284 TI - [LPS mutants of Sinorhizobium meliloti and their nodulation competitiveness]. AB - Four Tn5-transposon LPS mutants of Sinorhizobium meliloti (Tb9, Tb29, Ts22 and Ts32) have been studied. Each of four mutants has been established to contain a single insertion of Tn5-transposon in its genome. All mutations are located on a chromosome. Nodulation competitiveness (NC) of mutants towards the parent strain of S. meliloti CXM1-188 was investigated by resistant method using coinoculation of mutant and parent strain in the ratio 1:1. It was shown that NC was only 19 31% and 8-10%, for two strains Tb29 and Ts22, respectively which had lost the capability to synthesize higher molecular weight form of lipopolysaccharide (LPS1). Nodulation competitiveness of two other strains (Tb9 and Ts32) which retained the capability to synthesize LPS1 although in modified form varied from 49 to 62% and did not differ from NC of strain CXM1-188. The investigation of nodule formation rate has shown that four LPS-mutants did not differ from the parent strain by the number of root nodules. However the appearance of nodules induced by the mutant Tb29 was registered 7 days later than the nodules formed by other LPS-mutants and CXM1-188 strain. Obtained data concerning a single Tn5 insertion in genome of each of four S. meliloti LPS-mutants testify to the fact that both the disturbance of lipopolysaccharide synthesis and change of nodulation competitiveness in mutants Tb29 and Ts22 are results of a single mutation. PMID- 10872285 TI - [The isolation, purification and cloning of the pSM1 plasmid from the cyanobacterium Plectonema boryanum]. AB - Three methods of plasmids isolation from the cells of cyanobacteria: Plectonema boryanum 465, P. nostocorum 636, P. nostocorum 638, P. edaphicum 262, Plectonema sp. 456, Phormidium uncinatum 528, Anabaena variabilis 458 and Synechococcus cedrorum 750 have been approved. The method of plasmids isolation by 1 M NaCl salting out was used for plasmid screening. All the strains studied contained large, above 55 KB plasmids, small plasmids (1.5 KB) were found in the cells of P. boryanum 465 and P. nostocorum 636. After the restriction analysis a small plasmid of P. boryanum 465 was chosen to be used in the experiments on the vector construction. The above plasmid was called pSM1 was cloned into the vector pBluescri ptIISK(+) on the unique ClaI site. The obtained construction pBSMI may be used as the basic one for creation of cyanobacterial vectors. PMID- 10872286 TI - [The interaction of Bradyrhizobium japonicum with clay minerals]. AB - It is shown that such clay minerals as palygorskite and montmorillonite stimulate growth activity of Bradyrhizobium japonicum. The bacteria come into contact with the above minerals. Granulated preparations of rhizobia have been developed on the basis of the results obtained. These preparations are characterized by the high yield of viable cells and a possibility of long-term storage. The use of montmorillonite in production of granulated preparations provides the higher yield of viable bacteria in the preparations and stability of their composition under long-term storage. PMID- 10872287 TI - [Associative nitrogen-fixing microorganisms]. AB - State of the study of associative nitrogen-fixing microorganisms has been evaluated. The paper also deals with the problem of spatial and functional interrelations of diasotrophs with the higher plants, formation peculiarities of nitrogen-fixing associations under inoculation of annual and perennial plants. PMID- 10872288 TI - [Optimizing oral anticoagulation. Free flow in the blood stream]. PMID- 10872289 TI - [Managing one's own fate. Is every patient his own general practitioner?]. PMID- 10872290 TI - [MDR-1 gene changes effectiveness of drugs. DNA analysis instead of dose schedule F?. Interview by Petra Eiden]. PMID- 10872291 TI - [New trend in type 2 diabetes moves toward short-term insulins! Better metabolic control, less weight gain]. AB - Owing to the fact that the regulation of glucose is usually inadequate, and that the constituents of the metabolic syndrome are frequently present, type 2 diabetics are classed as high-risk patients. The prime importance of disordered insulin secretion for the pathogenesis and clinical presentation of the disease is currently receiving ever stronger confirmation from the results of animal experiments and clinical studies. Also, the noxae for macroangiopathy and insulin resistance originating in endothelial dysfunction are presently attracting attention. Damage to the vascular endothelium can be caused simply by postprandial hyperglycemia. Despite formerly held views to the contrary, insulin must clearly be considered a protective factor against arteriosclerosis. For this reason alone, a central hypothesis of the UKPDS published in 1998 should be taken to heart: 'more insulin will be needed'. PMID- 10872292 TI - [Therapy of type 2 diabetes. Critical evaluation of oral antidiabetic drugs]. AB - Changes in lifestyle and eating habits always form the initial phase of treatment of type 2 diabetes. Sooner or later, type 2 diabetes is associated with a lack of insulin that makes treatment with insulin necessary. Today, however, modern oral antidiabetics make it possible to exert a selective positive influence on both insulin resistance and disturbed insulin secretion. Three main groups of substances are available. Among the stimulators of insulin secretion are the sulfonylureas (e.g. glibenclamide, glibonuride, glisoxepid, glimepiride), and the so-called prandial glucose regulators, such as repaglinide, which differ from the sulfonylureas both chemically and in their pharmacodynamic properties. The group of insulin sensitizers includes the biguanide, metformin and the thiazolidinediones or glitazones (rosiglitazone, pioglitazone). The third group are the alpha-glucosidase inhibitors, e.g. acarbose. In principle, these oral antidiabetics can be combined with one another. PMID- 10872293 TI - [Insulin resistance. Frequently administration errors are the source]. AB - The term insulin resistance includes both physiological and pathological metabolic changes. Physiological insulin resistance, for example, is to be found in puberty or in advanced old age; additionally, it may be caused by endocrinological diseases. A pathologically elevated insulin requirement is to be found in the insulin resistance syndrome, type A (genetic) or type B (insulin receptor antibodies). In recent times, however, a massively elevated insulin requirement has been observed in increasing numbers of type 2 diabetics, in whom a subcutaneous insulin resistance is suspected--a condition that can be verified by various tests. The treatment of choice for breaking down this resistance is the intravenous administration of sufficiently high-dosed insulin that is as closely matched to the sugar profile as possible. In many cases the return to subcutaneous insulin--which again must be adequately high dosed--becomes possible during the further course of the disease. In addition, a case of insulin resistance caused by patient manipulation is described. PMID- 10872295 TI - [Intraperitoneal insulin therapy. High tech for treatment of type 1 diabetes]. PMID- 10872294 TI - [Intensified insulin therapy. References for general practice for increased control]. AB - Today, intensified conventional insulin therapy (ICT) is the standard treatment for type 1 diabetes, but patients with type 2 diabetes younger than 70 also profit it. Within an international framework too, the DCCT study has confirmed the usefulness of intensified insulin therapy. Factors that have a decisive influence on the success of such treatment are the choice of a suitable insulin, its dose, site of injection, and prandial adaptation. The various possibilities of carrying out ICT are discussed. PMID- 10872296 TI - [Alpha-glucosidase inhibitor for type 2 diabetic patients. Clinical study with miglitol]. PMID- 10872297 TI - [Travel pharmaceuticals for the physician. 2: Trip to the (sub-)tropics]. PMID- 10872298 TI - [Flu-like infection with background illness. Hairy cell leukemia]. PMID- 10872299 TI - [Is evidence-based medicine tyranny?]. PMID- 10872300 TI - [Tackling insulin resistance. Control of etiology in type 2 diabetes]. PMID- 10872301 TI - [Antihypertensive for hypertensive patients without discipline. The threat of senile dementia!]. PMID- 10872302 TI - [Infectious diarrhea. Probiotic agent can be useful]. PMID- 10872303 TI - [Dyslipidemia is a risk for patients with kidney transplants. Do statins protect the kidney?]. PMID- 10872304 TI - [Solutions to questions on the topic of ultrasound diagnosis]. PMID- 10872305 TI - [Allergy diagnosis and treatment in the year 2000]. PMID- 10872306 TI - [Diagnostic clinical trial with allergen extract from Blomia tropicalis in allergic adults and healthy volunteers]. AB - BACKGROUND: There are no antecedents in Cuba of studies developed to prove the efficacy of an allergy test to Blomia tropicalis despite of being one of the main mites which cause this ailment which affects more than 2,200,000 people. OBJECTIVES: Evaluate the allergenic extract Blomia tropicalis elaborated at the Centro Nacional de Biopreparados (BioCen), in the prick skin test, as a diagnose of allergy to this allergen which exists in all the tropical and subtropical countries. MATERIAL AND METHODS: We selected 50 patients with a positive clinic history of allergy to house dust and 50 healthy persons. Al the persons selected were skin prick tested with dilution's of the extracts (20,000 UB/mL) and positive and negative control. We measured the average diameter of the wheal and the perpendicular diameter, then we calculated the average. We also calculated size of the wheal in the duplicated, which constituted the size of the reaction. We estimated the validity of the test calculating the sensitivity and specificity. We used the EPITABLE program to do this. RESULTS: The skin prick tested positive in 78% of the patients and 6% in the healthy persons, for a sensitivity of 78% the interval of confidence (IC) 63.7-88.0 and a specificity of 94%, IC 82.5-98.4 the predictive negative value 81%, IC 68.2-89.7. CONCLUSIONS: The allergenic extract of Blomia tropicalis elaborated in BioCen is very sharp to test the allergy to this mite. PMID- 10872307 TI - [Procedure for bronchodynamic tests with inhaled allergens]. AB - The asthma is one of the most frequent chronic illnesses in the children. It is characterized by hyperreactivity from the air roads to inspecific stimuli (metacholin and histamin) and specific (allergens). The broncodynamics tests determines the broncolability asthmatic boy's and they help to settle down the I diagnose functional. They should be carried out with antigenic extracts of high quality, lyophilized watery and standardized; you can use an open or closed circuit and a dosimeter, the initial dose of the allergen it is lower than with the one that the positive cutaneous reactivity was obtained, carrying out successive increments until the VEF-1 descend 15% or more. The patient's security is fundamental, they should be carried out for specialized personnel. PMID- 10872308 TI - [Clinical effect of a sustained-release theophylline formulation in moderate or severe asthmatic patients with intercrisis]. AB - The clinical effect of the formulation of theophylline of sustained action was evaluated (Aristegui 300 mg, capsules) to the dose of 9 weight/day mg/kg (every 12 hours) in the consultation of bronchial asthma of Hospital Docente Provincial Clinico Quirurgico de Santa Clara, in the understood period of February to September of 1997. The time of treatment was of one month and they were carried out five consultations. The sample was constituted by 30 asthmatic, moderate or severe patients. The variables were analyzed: attendance to guard's body, salbutamol use in spray and they were carried out objective mensurations from the lung function to all the patients in the different consultations. With the use of the formulation they diminished the flow expiratory pick significantly, the expiratory volume forced in one second and the flow half expiratory maximum and low incidence of adverse effects existed. PMID- 10872309 TI - [Effect of 2 pesticides indoors on the respiratory function of a Mexican population]. AB - INTRODUCTION: The utilisation of pesticides in the large cities has been increased. Exist few studies that determine the damage in the respiratory appliance. OBJECTIVE: Determined on a healthy population of Mexico City with two known products used for the control of the insecticides in household, with the main objective to determine the clinical and functional manifestations that its use implies. MATERIAL AND METHODS: They were included 70 sound volunteers with residence in the City of Mexico that lived in apartments with 100 square meters of construction, tobacco negative and without previous antecedents of cardiovascular disease, without acute infectious process. They were split into 2 groups one that inhaled pesticides for combustion and the group b in electric form, In them I accomplished study espirometric to the beginning and during 3 exposed hours. The analysis by T Student. RESULTS: In group A was observed a meaningful difference of 0.01 in the first hour and of p < 0.028 in the second and third hours, with collateral effects as irritation eyes (74%) disnea (57%) cough (43%) and headaches (28%). Group B with a small difference of the VEF-1 in the first hour p < 0.03 and without differences on the following hours. We haven't observed secondary effects according to this group. The analysis en both group we have only differences in the first hours with p < 0.004. CONCLUSION: The exposition to pesticides our group for combustion produces a lot of clinical and functional alterations than the electric pesticides. PMID- 10872310 TI - [Effect of heating on FEV-1 in children with asthma challenged with exercise]. AB - AIMS: The exercise-induced bronchoespasm, also called asthma for exercise, it corresponds to a frequent clinical entity that commonly accompanies the asthmatic patient. Their frequency is of around the 80 to 90% for the patients with asthma; it is presented among 40 to 50% of the children with allergic rhinitis, in the athletes 14% and in the population's 12% in general. OBJECTIVE: To determine the effectiveness of a heating routine like preventive agent of the exercise-induced bronchoespasm. METHOD: 30 patients were included of between 10 and 16 years of age, asthmatic and with exercise-induced bronchoespasm by diagnosed by means of challenge test to the exercise. All the patients carried out an exercise routine with duration of 20 minutes during which movements of elasticity, calisthenics and light activity were made directed to the exercise to develop, taking as objective parameter 60% of the frequency heart submax. Did all the patients carry out challenge at once, again to the exercise, in which a band numberless was used, with speeds that were increased progressively from 1 to 8 km/h and with inclination of 0? 10 0. The spirometrics registrations was made before the challenge and at the 2, 5, 10, 15, 20, 25, 30 and 60 minutes after the same one. RESULTS: The average of the patient's age was of 12.8 +/- 2 years with size of 157 +/- 10 cm. CONCLUSION: The heating in children with broncoespasmo induced by exercise acts as a protective agent against the decrement of the VEF-1. Although the heating can be a good control method in the asthmatic patient, it is necessary to keep in mind that not all the patients have a benefit of this protection. This can only be used in patient with programmed physical activity, since in the preschoolers it is not possible to implant a heating routine for the characteristics characteristic of their activity. PMID- 10872311 TI - [Rhinitis and nasal polyps. Their association with domestic acari]. AB - BACKGROUND: In persons older than one year, respiratory allergies can be studied from the etiologic viewpoint using skin test with specific allergens. MATERIAL AND METHOD: The objective of this research is to make an assessment of the skin reactivity to mites Dermatophagoides pteronyssinus, Dermatophagoides siboney and Blomia tropicalis in 76 patients with a diagnosis of allergic and nasal polyps which had been treated in Otorhinolaryngologie and sent to the service of allergologie in Esmeralda Camaguey. Every patient received a skin puncture test twice. Fifteen minutes later the hives began to appear and we measured the longer diameter and the perpendicular diameter of every hive and we calculated an average. We also took the average size of the hives in the second test, considering them positive when the medium diameter was < 3 mm and negative when it was < 3 mm. In those patients suffering from rhinitis the greatest positive frequency was for those with Dermatophagoides pteronyssinus and the greatest negative frequency to Dermatophagoides siboney. In patients with nasal polyps there was a positive predominance (p < 0.05) to Dermatophagoides pteronyssinus and a negative predominance (p < 0.05) to Dermatophagoides siboney. So, we many conclude that the mite Dermatophagoides pteronyssinus causes more positive reactions than Dermatophagoides siboney and Blomia tropicalis in those patients with allergic rhinitis and nasal polyps. PMID- 10872312 TI - Procollagen alpha 2(I) mRNA in dermatosparactic fibroblasts: evidence for post transcriptional regulation. AB - Dermatosparaxis is a genetic defect of connective tissues found in man and animals. Dermatosparactic sheep fibroblasts fail to cleave off the aminopropeptide of procollagen, this makes the feedback regulation less effective and leads to an elevated production of collagen. We have isolated the total RNA from dermatosparactic and normal sheep fibroblasts and then compared the amount of mRNA sequences complementary to alpha 2(I) genomic DNA. Using methyl mercury gels, the Northern hybridization procedure and dotting hybridization, we were able to show that the levels of mRNA specific for alpha 2(I) collagen were only marginally different in both fibroblast strains. Furthermore, the amount of collagen mRNA translatable in a cell-free translation system was the same in affected and non-affected sheep cells. Our data provide further evidence for a post-transcriptional control of collagen synthesis in fibroblasts. PMID- 10872313 TI - Microtubule-associated protein MAP2 preferentially binds to a dA/dT sequence present in mouse satellite DNA. AB - Microtubule-associated protein MAP2 binds to the Sau96.1 restriction monomer fragment of mouse satellite DNA. This fragment is also present in a lower proportion in bulk DNA. The digestion of MAP2-Sau96.1 fragment complex by DNase results in the protection of certain nucleotide sequences. The sequence poly(dA)4/poly(dT)4 is mainly protected against DNase digestion. PMID- 10872314 TI - Nucleotide sequence and intron structure of the apocytochrome b gene of Neurospora crassa mitochondria. AB - The sequence of the apocytochrome b (cob) gene of Neurospora crassa has been determined. The structural gene is interrupted by two intervening sequences of approximately 1260 bp each. The polypeptide encoded by the exons shows extensive homology with the cob proteins of Aspergillus nidulans and Saccharomyces cerevisiae (79% and 60%, respectively). The two introns are, however, located at sites different from those of introns in the cob genes of A. nidulans and S. cerevisiae (which contain highly homologous introns at the same site within the gene). The introns share several short regions of sequence homology (10-12 bp long) with each other and with other fungal mitochondrial introns. Moreover, the second intron contains a 50 nucleotide long sequence that is highly homologous with sequences within every ribosomal intron of fungal mitochondria sequenced to date. The conserved sequences may allow the formation of a core secondary structure, which is nearly identical in many mitochondrial introns. The conserved secondary structure may be required for intron splicing. The second intron contains an open reading frame, continuous with the preceding exon, of approximately 290 codons. Two stretches of 10 amino acid residues, conserved in many introns, are present in the open reading frame. PMID- 10872315 TI - Microheterogeneity of tubulin proteins in neuronal and glial cells from the mouse brain in culture. AB - The microheterogeneity of the alpha and beta isoforms of tubulin in brain cells in culture was studied. The cells were prepared from two precise regions of the embryonic mouse brain (ED15), the striatum and the mesencephalon. It was possible to maintain virtually pure cultures of neuronal or glial cells up to 1 and 4 weeks in vitro, respectively. The tubulin heterogeneity of striatal and mesencephalic neurons was found to be very similar after a few days in culture. More precise examination of pure neurons from the striatum revealed that their tubulin content after 7 days in vitro exhibited the same degree of complexity as a control extract from a 4 day-old mouse brain. In fact, we could detect the presence of at least six alpha and nine beta tubulin isoforms. Among these isoforms a specific family of beta proteins (beta' tubulin) and the more acidic alpha proteins were present. Since these isoforms have, up to now, been found only in tubulin extracts prepared from the nervous system, our experiments suggest that they belong to the neuronal subpopulation of this tissue. This point is reinforced by their complete absence from the tubulin proteins extracted from pure glial cells even after several weeks in vitro. These results lead us to propose that brain tubulin microheterogeneity is associated with the presence of neurons and not of glia and may, therefore, play a specific role in maintaining neuronal shape and function. PMID- 10872316 TI - Control-mechanisms acting at the transcriptional and post-transcriptional levels are involved in the synthesis of the arginine pathway carbamoylphosphate synthase of yeast. AB - In Saccharomyces cerevisiae, the synthesis of the arginine pathway enzyme carbamoylphosphate synthase (CPSase A) is subject to two control mechanisms. One mechanism, the general control of amino acid biosynthesis, influences the expression of both CPA1 and CPA2 genes, the structural genes for the two subunits of the enzyme. The second mechanism, the specific control of arginine biosynthesis, only affects the expression of CPA1. To study these mechanisms in more detail, we have cloned the CPA1 and CPA2 genes and used their DNA to measure the CPA1 and CPA2 mRNA content of cells grown under various conditions. A close coordination was observed in the variation of the levels of CPA1 and CPA2 mRNAs and polypeptide products under conditions where the general control of amino acid biosynthesis operates. In contrast, little correlation was found between the levels of CPA1 mRNA and the corresponding protein for conditions affecting repression by arginine: the total amplitude of variation was 6-fold higher for the CPA1 protein than for the CPA1 messenger transcript. Such findings are consistent with the conclusion that the general control operates at the transcriptional level and that the specific arginine control acts primarily at a post-transcriptional level. PMID- 10872317 TI - Catenation of DNA by eucaryotic topoisomerase II associated with simian virus 40 minichromosomes. AB - After incubation of purified SV40 minichromosomes with superhelical DNA molecules either of SV40 or plasmid origin, a catenation of monomeric DNA via dimers and multimers to large networks was observed. The catenation reaction was stimulated by the DNA condensing agent spermidine with ATP as an energy donor and was dependent on the presence of magnesium ions. The reaction could be blocked by inhibitors of topoisomerase II such as novobiocin and nalidixic acid. Relaxed covalently closed circular DNA was catenated to networks in the presence of ATP as the energy donor. PMID- 10872318 TI - In vitro synthesis and post-translational insertion into microsomes of the integral membrane protein, NADH-cytochrome b5 oxidoreductase. AB - RNA extracted from a free polysome fraction from rat liver was used to direct translation in nuclease-treated rabbit reticulocyte lysates, and the [35S]methionine-labelled, in vitro-synthesized, cytochrome b5 reductase was isolated with specific antibodies. Analysis by SDS-polyacrylamide gel electrophoresis, non-equilibrium pH gradient electrophoresis and one-dimensional peptide mapping failed to reveal any difference between the in vitro-synthesized reductase and the enzyme endogenous to rat liver microsomes. To study the integration of the in vitro-synthesized reductase into membranes, carboxypeptidase Y was used as a proteolytic probe. The reductase endogenous to rat liver microsomes was resistant to attack by carboxypeptidase Y, but was degraded to a smaller form when the microsomes were solubilized by detergent. Likewise, the enzyme synthesized in vitro was attacked by carboxypeptidase Y, but became largely resistant after post-translational incubation with dog pancreatic microsomes, indicating that an integration into membranes similar to the physiological one had occurred. It is concluded that cytochrome b5 reductase is probably not synthesized as a precursor and inserts post-translationally into the membrane. The results are discussed in relation to the particular subcellular distribution of the reductase and to the possible topology in the lipid bilayer of its C-terminal non-polar membrane-binding segment. PMID- 10872319 TI - 72 residues of gal repressor fused to beta-galactosidase repress the gal operon of E. coli. AB - An active gene has been constructed which produces a chimera consisting of the N terminal domain of the gal repressor and all but the first five residues of beta galactosidase. Seventy two residues of gal repressor fused to beta-galactosidase as tetrameric core are sufficient to repress the gal operon in vivo and to bind to the gal operator in vitro. PMID- 10872320 TI - The ultimate localization of an outer membrane protein of Escherichia coli K-12 is not determined by the signal sequence. AB - To study the role of the signal sequences in the biogenesis of outer membrane proteins, we have constructed two hybrid genes: a phoE-ompF hybrid gene, which encodes the signal sequence of outer membrane PhoE protein and the structural sequence of outer membrane OmpF protein, and a bla-phoE hybrid gene which encodes the signal sequence as well as 158 amino acids of the structural sequence of the periplasmic enzyme beta-lactamase and the complete structural sequence of PhoE protein. The products of these genes are normally transported to and assembled into the outer membrane These results show: (i) that signal sequences of exported proteins are export signals which function independently of the structural sequence, and (ii) that the information which determines the ultimate location of an outer membrane protein is located in the structural sequence of this protein, and not in the signal sequence. PMID- 10872321 TI - Stage-specific protein synthesis during early embryogenesis in Drosophila melanogaster. AB - The changes in protein species synthesized during early Drosophila embryogenesis were characterized by two-dimensional electrophoresis. Of the 261 proteins scored, 68 (26%) show dramatic changes in rates of synthesis during the first 8 h of embryogenesis. These stage-specific proteins can be classified into three categories: early, detected at 1, 2 and 3 h but not later; late, not detected at 1 h, but appearing later; and discontinuous, detected before and after, but not at 3 and 4 h. RNA was extracted from three representative stages, translated in vitro, and the translation products separated on two-dimensional gels. There was a strong correlation between the patterns of synthesis in vivo and in vitro, suggesting that the early proteins are translated from maternal mRNA, and the late proteins from zygotic mRNA. A thorough comparison was made between the proteins synthesized in wild-type and dorsal embryos, in which virtually only dorsal hypoderm differentiates. The first observed difference was a reduced synthesis of actin I at 8 h, indicating that the absence of mesodermal and endodermal tissues is not detectable at the level of moderately abundant protein until the onset of differentiation. PMID- 10872322 TI - Archaebacteria and eukaryotes possess DNA-dependent RNA polymerases of a common type. AB - DNA-dependent RNA polymerases of archaebacteria not only resemble the nuclear RNA polymerases of eukaryotes rather than the eubacterial enzymes in their complex component patterns but also show striking immunochemical, i.e., structural, homology with the eukaryotic polymerases at the level of single components. Thus, eukaryotic and archaebacterial RNA polymerases are indeed of the same type, distinct from the eubacterial enzymes, which, however, are also derived from a common ancestral structure. PMID- 10872323 TI - Neurofilament architecture combines structural principles of intermediate filaments with carboxy-terminal extensions increasing in size between triplet proteins. AB - Mammalian neurofilament triplet proteins (68 K, 160 K and 200 K) have been correlated by a biochemical, immunological and protein chemical study. The 160 K and 200 K triplet proteins are intermediate filament proteins in their own right, since they reveal the alpha-helical coiled-coil rod domain analyzed in detail for the 68 K protein. Triplet proteins display two distinct arrays. Their amino terminal region built analogously to non-neuronal intermediate filament proteins should allow a co-polymerization process via the interaction of coiled-coil domains. The extra mass of all triplet proteins is allocated to carboxy terminally located extensions of increasing size and unique amino acid sequences. These may provide highly charged scaffolds suitable for interactions with other neuronal components. Such a domain of 68 K reveals, in sequence analysis, 47 glutamic acids within 106 residues. The epitope recognized by a monoclonal antibody reacting probably with all intermediate filament proteins has been mapped. It is located within the last 20 residues of the rods, where six distinct intermediate filament proteins point to a consensus sequence. PMID- 10872324 TI - Purification and characterization of Xenopus laevis topoisomerase II. AB - We have purified to apparent homogeneity a type II DNA topoisomerase from Xenopus laevis oocyte nuclei (germinal vesicles, or GV). The most pure preparations contain a single polypeptide of 175,000 daltons as determined by SDS-gel electrophoresis. The enzyme changes the linking number of DNA circles in steps of two and reversibly knots or catenates DNA rings. No gyrase activity is detectable and ATP is required. PMID- 10872325 TI - A model for the tertiary structure of mammalian mitochondrial transfer RNAs lacking the entire 'dihydrouridine' loop and stem. AB - The mammalian mitochondrial tRNA(AGY)Ser is unique in lacking the entire dihydrouridine arm. This reduces its secondary structure to a 'truncated cloverleaf'. Experimental evidence on the tertiary structure has been obtained by chemically probing the conformation of both the bovine and human species in their native conformation and at various stages of denaturation. A structural model of the bovine tRNA is presented based on the results of this chemical probing, on a comparison between nine homologous 'truncated cloverleaf' secondary structures and on analogies with the crystal structure of yeast phenylalanine tRNA. The proposed structure is very similar in shape to that of yeast tRNA(Phe) but is slightly smaller in size. It is defined by a unique set of tertiary interactions. Structural considerations suggest that other mammalian mitochondrial tRNAs have smaller dimensions as well. PMID- 10872326 TI - Puffing activities and binding of ecdysteroid to polytene chromosomes of Drosophila melanogaster. AB - Salivary glands of third instar Drosophila melanogaster larvae were incubated in vitro in the presence of 5 x 10(-6) M 20-hydroxy-ecdysone. Steroid hormone was localized on the polytene chromosomes of the salivary gland by a combination of photoaffinity-labeling and indirect immunofluorescence microscopy. Steroid hormone binding to chromosomal loci and their puffing activity was correlated for the larval/prepupal puffing cycle characterized by puff stages 1-10. In general, there was a good correlation between the sequential and temporal puffing activity induced by 20-hydroxy-ecdysone and the binding of ecdysteroid hormone to these puffs. Ecdysteroid hormone was detected at intermolt, and at early and late puffs with two notable exceptions. Ecdysteroid was not detected at the two well-studied puffs at 23E and at 25AC, the former being an early puff, which is activated in the presence of 20-hydroxy-ecdysone, and the latter being an intermolt puff, which regresses more rapidly in the presence of hormone. Ecdysteroid hormone was present at puffs as long as the respective puff was active. Also, it apparently accumulated at late puff sites after induction. Since ecdysteroid binding to chromosomal loci is temporal as well as sequential during the larval/prepupal puffing cycle, additional factors besides steroid hormone are necessary for sequentially regulating puffing and concomitant gene activity during development from larvae to prepupae. PMID- 10872327 TI - Latent and lytic cycle promoters of Epstein-Barr virus. AB - Four RNA polymerase II promoters have been mapped in the DNA sequence of the EcoRI-H and -Dhet fragments of B95-8 Epstein-Barr virus. RNAs transcribed from three of these promoters are dramatically induced by treatment of B95-8 cells with 12-O-tetradecanoylphorbol-13-acetate (TPA). The other promoter is active with or without TPA treatment of the cells and is thus active in the latent virus cycle. Deletion mapping suggests that DNA sequence homologies between some of the promoters lie in the same region as essential upstream promoter elements. PMID- 10872328 TI - The primary structure of the duck alpha D-globin gene: an unusual 5' splice junction sequence. AB - The complete nucleotide sequence of the duck minor alpha D-globin gene including the flanking regions has been determined. A unique structural feature of the alpha D-globin gene is a GC instead of the invariant GT dinucleotide at the 5' end of the second intervening sequence. The 1013 base pair long gene has otherwise all the characteristics normally attributed to a functional globin gene. Indirect evidence suggests that the alpha D-globin gene is expressed in vivo. PMID- 10872329 TI - The nucleotide sequence of the first externally suppressible--1 frameshift mutant, and of some nearby leaky frameshift mutants. AB - Nine mutants within a 23 nucleotide sequence of the trpE gene of Salmonella typhimurium have been characterized. trpE91, a mutant which is externally suppressible has a single base deletion. Eight (or nine) nucleotides upstream of this deletion, two independently isolated mutations have the same transversion. In combination with trpE91 these mutations lead to partial restoration of synthesis of anthranilate synthetase in the absence of external suppressors. In the transversion the sequence A CA is changed to A AA and this new sequence may be the site where frameshifting occurs to allow leakiness. Leakiness is displayed by two further mutants of the same sign as trpE91, and one of the opposite sign, in the absence of any base substitution or external suppressors. Specific sequences, e.g., UUUC, may be especially prone to frameshifting and this sequence is created at the site of the +1 frameshift mutant which displays leakiness. In the new reading frame generated by the two -1 frame leaky mutants, a tryptophan codon is encountered. Leakiness is necessarily detected in the absence of tryptophan and under these conditions there will be a shortage of charged tryptophan tRNA. The possibility of such functional imbalance leading to frameshifting in these mutants is discussed. PMID- 10872330 TI - Codon-specific missense errors in vivo. AB - We have developed a simple method for measuring the missense substitution of amino acids at specified positions in proteins synthesized in vivo. We find that the frequency of cysteine substitution for the single arginine in Escherichia coli ribosomal protein L7/L12 is close to 10(-3) for wild-type bacteria, decreases to 4 x 10(-4) in streptomycin-resistant bacteria containing mutant S12 (rpsL), and is virtually unchanged in Ram bacteria containing mutant S4 (rpsD). We have also found that the frequency of the cysteine substitution for the single tryptophan in E. coli ribosomal protein S6 is 3-4 x 10(-3) for wild-type bacteria, decreases to 6 x 10(-4) in streptomycin-resistant bacteria and is elevated to nearly 10(-2) in Ram bacteria. PMID- 10872331 TI - Evidence for a repeating cross-beta sheet structure in the adenovirus fibre. AB - The amino acid sequence of the adenovirus fibre protein reveals an approximately repeating motif of 15 residues. A diagonal comparison matrix established that these repeats extended from residue 43 to residue 400 of the 581 residue sequence. Assignment of secondary structure combined with model building showed that each 15-residue segment contained two short beta-strands and two beta-bends, one of which incorporated an extra residue in a beta-bulge of the Gx type. The 44 strands together gave a long (210 A) narrow, amphipathic beta-sheet, which could be stabilised by dimer formation to give the shaft of the fibre. The knob could arise from a dimer of the C-terminal 180 residue segment, predicted to be an 8-10 stranded beta-sandwich. This model is consistent with the electron micrographs of the fibre and it was supported by measurements of c.d. and of electron diffraction from microcrystals. The latter gave a pair of wide angle arcs, corresponding to a repeat of 4.7 A, oriented appropriately for a cross-beta structure. The relation of this structure to globular structures is discussed and a folding pathway is proposed. In its general features the structure resembles that proposed for the tail fibre of bacteriophage T4. PMID- 10872332 TI - Higher-order structure of nucleosome oligomers from short-repeat chromatin. AB - Sedimentation measurements and electron microscopy at a series of ionic strengths suggest that chromatin from neurons of the cerebral cortex is able to form condensed structures in vitro that are probably several turns of a solenoid with about six nucleosomes per turn. Since neuronal chromatin has a short nucleosomal repeat (approximately 165 bp) allowing virtually no linker DNA between nucleosomes, and yet forms apparently 'normal' elements of solenoid, the packing of nucleosomes in the solenoid must be highly constrained. This permits only a limited number of possible models, and enables tentative suggestions to be made about the location of the linker DNA in the typical solenoid. PMID- 10872333 TI - Expression and regulation of immunoglobulin heavy chain gene transfected into lymphoid cells. AB - A plasmid including a mouse immunoglobulin mu gene was transfected into the IgG secreting human lymphoid line HMy2 and mouse B- and pre-B-cell lines WEHI 231 and 18-81; stably transfected cells were selected. Transfected HMy2 cells synthesized mouse immunoglobulin mu chains as a major secreted protein but the WEHI 231 and 18-81 transfectants transcribed the introduced mu gene at lower levels. In HMy2 transfectants, most of the transcription of the introduced heavy chain gene initiated 40 and 62 bp upstream of the beginning of the VH exon translation start, although a small proportion of transcripts initiating further upstream was detected. WEHI 231 and 18-81 transfectants gave a much higher proportion of upstream initiation. Transient expression of the VH exon was monitored following transfection of mouse myeloma with the VH gene DNA in various plasmid constructs. VH transcription was only observed if the plasmids contained a segment derived from the large VH-CH intron of the immunoglobulin heavy chain locus. This segment, located between JH and switch regions, functioned both downstream of the VH exon and upstream in either orientation. The existence of a transcription enhancer element in this region is therefore proposed. PMID- 10872334 TI - Isolation and properties of the native chromoprotein halorhodopsin. AB - The native chromoprotein of the light-driven chloride pump halorhodopsin (HR) was isolated from Halobacterium halobium strain L-33 which lacks bacteriorhodopsin but contains 'slow cycling rhodopsin-like pigment' (SR). A membrane fraction was prepared in low salt and dissolved in a high salt medium by the detergents Lubrol PX or octylglucoside. These conditions destroyed the chromophore of SR but not the HR pigment. Chromatography on phenyl-Sepharose and hydroxylapatite produced, in 60% yield, a 230-fold enriched monomeric chromoprotein with an apparent mol. wt. of 20,000. The chromoprotein was stable in 1 M NaCl and 1% octylglucoside and remained stable upon removal of detergent. It reacted with borohydride in the dark and with hydroxylamine in the light. The absorption maximum of the light adapted state is at 580 + 2 nm and its molar extinction approximately 50,000/M/cm. Upon illumination in the presence of detergent it was converted into a 410 nm absorbing species with concomitant release of protons. A thermal reconversion to the 580 nm species occurred with a half time of 76 s at -6 degrees C. Blue light absorbed by the photoproduct accelerated the re-conversion as well as the re-uptake of protons. Removal of the detergent prevented the light induced formation of the 410 nm species. Under these conditions a photochemical behaviour similar to that in intact cells and cell vesicles, i.e., a photocycle in the 10-20 ms range was observed. These findings form the basis for functional reconstitution of HR. PMID- 10872336 TI - Clathrin heavy chain, light chain interactions. AB - Purified pig brain clathrin can be reversibly dissociated and separated into heavy chain trimers and light chains in the presence of non-denaturing concentrations of the chaotrope thiocyanate. The isolated heavy chain trimers reassemble into regular polygonal cage structures in the absence of light chains. The light chain fraction can be further resolved into its two components L alpha and L beta which give different one-dimensional peptide maps. Radiolabelled light chains bind with high affinity (KD < 10(-10) M) to heavy chain trimers, to heavy chain cages and to a 110,000 mol. wt. tryptic fragment of the heavy chain. Both light chains compete with each other and with light chains from other sources for the same binding sites on heavy chains and c.d. spectroscopy shows that the two pig brain light chains possess very similar structures. We conclude that light chains from different sources, despite some heterogeneity, have a highly conserved, high affinity binding site on the heavy chain but are not essential for the formation of regular cage structures. PMID- 10872335 TI - Translational products of mRNAs coding for non-epidermal cytokeratins. AB - Total RNA and poly(A)+ RNA were isolated from tissues and cultured cells of various mammalian species (bovine muzzle epidermis and bladder urothelium; rat hepatoma cells; human cell lines HeLa, MCF-7 and A-431) and examined by translation in vitro using the reticulocyte lysate system. Polypeptides were separated and identified by two-dimensional electrophoresis and cytokeratins were selectively enriched from the translation assays by co-polymerization with added heterologous cytokeratins. In all three species, non-epidermal cytokeratins A, D and mol. wt. 40,000 (corresponding to numbers 8, 18 and 19 of the human cytokeratin catalog of Moll et al., 1982) were identified as translation products capable of co-polymerization with epidermal keratins. Several other basic and other acidic cytokeratins were also identified as translational products. In addition, two unidentified polypeptides (mol. wt. 52,000 and 43,000) which were minor polypeptides in cytoskeletons and translation assays were found to be specifically enriched in co-polymers with bovine epidermal keratins. The results indicate that many, perhaps all, non-epidermal cytokeratins characteristic of simple epithelia are genuine products of translation and that their diversity is not due to post-translational modification or processing. These findings, taken together with observations of in vitro translation of epidermal mRNAs, suggest that the diversity of cell type-specific expression of the different members of the cytokeratin polypeptide family is largely due to the cell type-specific synthesis of diverse mRNAs. PMID- 10872337 TI - Biochemical and immunological studies on clathrin light chains and their binding sites on clathrin triskelions. AB - Clathrin light chains from bovine brain tissue (LC alpha and LC beta) are monomeric proteins with an average mol. wt. of approximately 33,000, as determined by sedimentation equilibrium. Solution studies on purified light chains indicate a large Stokes radius (Re = 3.3 nm) and little defined secondary structure. Both light chains bind specifically and with high affinity (KA approximately 5 x 10(7)/M) to overlapping sites on clathrin heavy chains. These binding sites are contained within a 125,000 dalton heavy chain fragment that forms truncated triskelions with legs, 15 nm shorter than those of intact triskelions. As judged by immuno-electron microscopy, light chain-specific IgG molecules bind mostly to the center of triskelions, but there are also sites that are scattered some 16 nm along the proximal part of triskelion legs. From heterologous binding experiments using human placenta light chains and heavy chain fragments from bovine brain clathrin, it is concluded that the domains of light and heavy chains that are involved in the interaction are conserved across tissue and species boundaries. PMID- 10872338 TI - Electron microscopy and single molecule averaging of subunit-deficient F1-ATPases from Escherichia coli and spinach chloroplasts. AB - The morphology of F1-ATPases lacking one or more small subunits has been investigated by minimal-beam electron microscopy of close-packed monolayers of molecules. Computer-based rotational analyses of single molecules were performed on reconstituted 3-subunit F1-ATPase (-delta epsilon) from Escherichia coli and both 3-subunit (-delta epsilon) and 4-subunit (-delta) F1-ATPase from chloroplasts. Optical diffraction measurements of close-packed arrays revealed maximal dimensions of 122 +/- 4 A and 129 +/- 9 A for 3-subunit ECF1 and 4 subunit CF1, respectively. Molecules which displayed either hollow or solid hexagonal morphologies were observed in all preparations. Averaged reconstructions were obtained from molecules with hollow morphologies in 3 subunit preparations and demonstrated strong hexagonal symmetry in projection with a central, stain-filled cavity. The average reconstruction obtained from molecules with the solid morphology in 4-subunit CF1 preparations, was also strongly hexagonal with six peripheral units ringed about a central subunit. Differences between hollow and solid morphologies cannot be attributed solely to the presence or absence of the delta and epsilon subunits; therefore, the two image types may represent staining variants of a common structure. Overall, the reconstructions are consistent with an alpha 3 beta 3 gamma stoichiometry for the coupling factors from both E. coli and chloroplasts. PMID- 10872339 TI - Molecular cloning and sequencing of OAX DNA: an abundant gene family transcribed and activated in Xenopus oocytes. AB - OAX DNA codes for a 181 nucleotide long RNA whose transcription is strongly activated in somatic nuclei after their injection into a Xenopus oocyte nucleus. OAX RNA can be transcribed in vitro using an extract of Xenopus oocyte nuclei and total genomic DNA. Hybridization with OAX RNA as a probe indicates that OAX DNA is abundant in the Xenopus genome (at least 10(4) copies per genome). OAX DNA is present in tandemly repeated HindIII units of 752 bp. The complete DNA sequence of one of these OAX HindIII units is reported here. The OAX RNA transcript has been mapped within the OAX HindIII unit using S1 nuclease. Microinjection into Xenopus oocyte nuclei of either the OAX HindIII unit or a subclone containing only the RNA coding portion of the OAX HindIII unit both produce OAX RNA transcripts. This shows that the OAX promoter lies within the coding region of the RNA. The OAX RNA sequence has two elements which fit the RNA polymerase III promoter consensus sequence, and shows homology with dispersed RNA polymerase III transcription units in mammals. PMID- 10872342 TI - Pruritic lesions on the trunk. Red scaly papules form around hair follicles and are resistant to hydrocortisone cream. PMID- 10872340 TI - Glucocorticoid regulation of mouse mammary tumor virus: identification of a short essential DNA region. AB - Transcription of mouse mammary tumor virus (MMTV) DNA is stimulated by steroid hormones. To determine the DNA sequences involved in this regulation, we constructed a plasmid containing the MMTV long terminal repeat (LTR) in front of the coding region of the herpes simplex thymidine kinase gene, from which the promoter had been removed. Portions of the LTR were removed by the nuclease Ba/31, and the deleted molecules were recloned and tested for transcriptional activity in transfections of Ltk-aprt- cells. Stably transfected cell clones were selected and hormone-dependent transcription from the MMTV promoter was studied by the S1 nuclease mapping method. The results show that DNA sequences between 105 and -204 base pairs upstream from the initiation site of viral transcription are required for glucocorticoid stimulation. PMID- 10872341 TI - 'Anti-aging' elixirs. PMID- 10872343 TI - Insulin resistance and hypertension. Patients in double jeopardy for cardiovascular disease. AB - Essential hypertension is prevalent among older individuals, and approximately 50% of persons with hypertension can be considered to have insulin resistance and hyperinsulinemia. It appears likely that insulin resistance and hyperinsulinemia predispose to, rather than result from, hypertension. Insulin resistance is associated with abnormalities in lipoprotein metabolism, hypercoagulability, and endothelial function, which probably account in part for the increased cardiovascular risk among hypertensive patients. To identify this subset of patients, all hypertensive patients should be screened for diabetes and lipid abnormalities. The presence of impaired glucose tolerance, diabetes, or hypertriglyceridemia and low HDL suggest the presence of insulin resistance. Insulin resistant patients, in particular, will benefit from exercise and weight loss. PMID- 10872344 TI - Anti-aging medicine. What makes it different from geriatrics? AB - The growth in popularity of so-called "anti-aging" medicine challenges physicians to examine their attitudes about aging. Does one define aging as a predisposition to pathology or as part of the life cycle? Is longevity without the chronic diseases associated with aging a realistic goal? Anti-aging modalities being prescribed by some practitioners include hormone replacement therapies, vitamin and mineral supplements, diet, and exercise. Although diet, exercise, and some vitamin and mineral supplements are well-recognized as preventive measures, unproven hormone, mega-vitamin, and herbal therapies are controversial. Both the patient and the physician bring biases and values to the discussion of anti-aging medicine, and that combination will influence the treatment decisions. PMID- 10872345 TI - H pylori infection. Review of the guideline for diagnosis and treatment. AB - Helicobacter pylori infection is prevalent among persons over age 60, is strongly associated with peptic and duodenal ulcer, and is caused by a microbe classified as a carcinogen. These factors combine to make the primary care physician key to proper diagnosis and treatment of H pylori infection. In 1998, the American College of Gastroenterology published an evidence-based guideline for the management of this infection. The guideline produced several fundamental recommendations that help clarify the management process: asymptomatic persons should not undergo testing, but testing should be performed on certain persons, testing should only be done if the intention is to treat; the choice of test is governed by the need for endoscopy; and several triple-therapy regimens are effective for eradication. PMID- 10872346 TI - Brain aging and memory: new findings help differentiate forgetfulness and dementia. AB - Geriatrics is pleased to highlight the clinical implications of research topics supported by the American Federation for Aging Research (AFAR). AFAR is a leading private organization supporting research on the aging process and diseases of older populations. More than 900 physicians, scientists, and students have received AFAR grants totaling more than $20 million since AFAR was founded by Irving S. Wright, MD, in 1981. The articles in the New Frontiers series are designed to provide primary care physicians with insight into the pathogenesis, diagnosis, prevention, and treatment of the diseases of aging. PMID- 10872347 TI - Severe trauma to the extremities. PMID- 10872348 TI - Joint diseases. PMID- 10872349 TI - Spinal disorders and low back pain. PMID- 10872350 TI - Osteoporosis. PMID- 10872351 TI - Lateral shelf acetabuloplasty in Perthes' disease. A review of the end of growth. AB - The surgical treatment of Perthes' disease by femoral or innominate osteotomy is not as effective in those over the age of eight years as it is in the younger child. This has prompted the search for other types of management in those who are older. The preliminary results of the use of a lateral shelf acetabuloplasty for such cases have shown encouraging results at two years. The concern with such an operation is that it might interfere with the growth of the outer aspect of the acetabulum and so prejudice the long-term outcome. We describe a review at maturity of 26 children presenting with early disease after the age of eight years who were treated by lateral shelf acetabuloplasty. The results suggest that the outcome is improved; 22 of 27 hips were rated as Stulberg groups 1 to 3. Poor results occurred in children, particularly girls, presenting with Group-4 disease over the age of 11 years. PMID- 10872352 TI - Soft-tissue interposition after closed reduction in developmental dysplasia of the hip. The long-term effect on acetabular development and avascular necrosis. AB - We reviewed 98 children (133 hips) with developmental dysplasia of the hip who underwent arthrography immediately after closed reduction by overhead traction. We followed the patients to skeletal maturity to investigate whether soft-tissue interposition influences acetabular development and avascular necrosis over the long term. The shape of the limbus and the thickness of the soft-tissue interposition at the acetabular floor, as shown on arthrograms at the time of reduction, were not directly related to the final radiological results or to the incidence of avascular necrosis. Even if marked soft-tissue interposition was found on the initial arthrogram, spontaneous disappearance was noted in 71% up to the age of five years. The final radiological results showed no difference between those in which the interposition disappeared and those with none at the time of closed reduction. However, the requirement for secondary surgery at the age of five years was significantly higher in those with more than 3.5 mm of soft tissue interposition. In the no-disappearance group (group C) further operation was necessary in 100% and the results were significantly worse at maturity according to Severin's classification. We suggest that the indications for open reduction should not be based solely on the arthrographic findings at the time of closed reduction. PMID- 10872353 TI - The use of dynamic interventional MRI in developmental dysplasia of the hip. AB - Conventional methods of imaging in the investigation of developmental dysplasia of the hip all have disadvantages, either in definition or in exposure to radiation. We describe a new open-configuration MR scanner which is unique in that it allows anaesthesia and access to the patient within the imaging volume for surgical procedures and application of casts. We performed 13 scans in eight anaesthetised infants. Dynamic imaging revealed two dislocated hips which were then visualised during reduction. Hip spicas were applied without removing the patient from the scanner. In one hip, an adductor tenotomy was carried out. In all patients, stressing the hips during dynamic imaging allowed an assessment of stability. This was particularly useful in two hips in which an analysis of stability in different positions facilitated the planning of femoral osteotomies. This method of imaging provides new and important information. It has great potential in the investigation of developmental dysplasia of the hip and, with ultrasound, may allow management without the need for radiography. PMID- 10872354 TI - Implantation of a soft-tissue expander before operation for club foot in children. AB - Primary skin closure after surgery for club foot in children can be difficult especially in revision operations. Between 1990 and 1996 a soft-tissue expander was implanted in 13 feet before such procedures. Two were primary operations and 11 were revisions. A standard technique was used for implantation of the expander. Skin augmentation was successful in 11 cases. There was failure of one expander and one case of wound infection. Sufficient stable skin could be gained at an average of five weeks. Primary skin closure after surgery was achieved in 12 cases. We conclude that soft-tissue expansion can be used successfully before extensive surgery for club foot. The method should be reserved for revision procedures and for older children. The technique is not very demanding, but requires experience to achieve successful results. PMID- 10872355 TI - Extension of the elbow and supracondylar fractures in children. AB - We tested the hypothesis that children who sustain a supracondylar fracture have a greater range of elbow hyperextension than those with a fracture of the distal radius. Three observers made 358 measurements in 183 children (114 boys and 69 girls). There were 119 fractures of the distal radius and 64 supracondylar fractures. Initially, the group with a supracondylar fracture appeared to have extension 1.7 degrees greater than that of the group with fracture of the distal radius. On average, there was a maximum variation of 3 degrees between observers. After allowing for age, gender and observer, there was no significant difference between the groups. Our study had greater than 80% power to detect a difference in hyperextension of 2 degrees at the 5% level with the above observer variability. When age and gender are taken into account, any variation in the amount of hyperextension at the elbow is not sufficient to explain the occurrence of a supracondylar fracture. PMID- 10872356 TI - The anterior band of the inferior glenohumeral ligament. Assessment of its permanent deformation and the anatomy of its glenoid attachment. AB - Surgical treatment for traumatic, anterior glenohumeral instability requires repair of the anterior band of the inferior glenohumeral ligament, usually at the site of glenoid insertion, often combined with capsuloligamentous plication. In this study, we determined the mechanical properties of this ligament and the precise anatomy of its insertion into the glenoid in fresh-frozen glenohumeral joints of cadavers. Strength was measured by tensile testing of the glenoid-soft tissue-humerus (G-ST-H) complex. Two other specimens of the complex were frozen in the position of apprehension, serially sectioned perpendicular to the plane containing the anterior and posterior rims of the glenoid, and stained with Toluidine Blue. On tensile testing, eight G-ST-H complexes failed at the site of the glenoid insertion, representing a Bankart lesion, two at the insertion into the humerus, and two at the midsubstance. For those which failed at the glenoid attachment the mean yield load was 491.0 N and the mean ultimate load, 585.0 N. At the glenoid region, stress at yield was 7.8 +/- 1.3 MPa and stress at failure, 9.2 +/- 1.5 MPa. The permanent deformation, defined as the difference between yield and ultimate deformation, was only 2.3 +/- 0.8 mm. The strain at yield was 13.0 +/- 0.7% and at failure, 15.4 +/- 1.2%; therefore permanent strain was only 2.4 +/- 1.1%. Histological examination showed that there were two attachments of the anterior band of the inferior glenohumeral ligament at the site of the glenoid insertion. In one, poorly organised collagen fibres inserted into the labrum. In the other, dense collagen fibres were attached to the front of the neck of the glenoid. PMID- 10872357 TI - Entrapment of the suprascapular nerve. AB - Operative release for entrapment of the suprascapular nerve was carried out in 35 patients. They were assessed at an average of 30 months (12 to 98) after operation using the functional shoulder score devised by Constant and Murley. The average age at the time of surgery was 40 years (17 to 67). Entrapment was due to injury in ten patients and no cause was found in three; 34 had diffuse posterolateral shoulder pain. The strength of abduction was reduced in all the patients. The average Constant score, unadjusted for age or gender, before operative release was 47% (28 to 53). In 25 of the patients both the supraspinatus and infraspinatus muscles were atrophied and seven had isolated atrophy of the infraspinatus muscle. The average conduction time from Erb's point to the supraspinatus muscle and to the infraspinatus muscle was 5.7 ms (2.8 to 12.8) and 7.4 ms (3.4 to 13.4), respectively. In two patients MRI revealed a ganglion in the infraspinatus fossa and, in another, a complete rupture of the rotator cuff. The average time from the onset of symptoms to operation was ten months (3 to 36). A posterior approach was advocated. The average Constant score, after operative release, unadjusted for age or gender was 77% (35 to 91). The overall result was excellent in ten of the patients, very good in seven, good in 14, fair in two, and poor in two. The symptomatic and functional outcome in our series confirmed the usefulness and safety of operative decompression for entrapment of the suprascapular nerve. PMID- 10872358 TI - The assessment of shoulder instability. The development and validation of a questionnaire. AB - We have developed a 12-item questionnaire for completion by patients presenting with shoulder instability. A prospective study of 92 patients was undertaken involving two assessments, approximately six months apart, performed in an outpatient department. Each patient completed the new questionnaire and the SF36 form. An orthopaedic surgeon completed the Constant shoulder score and the Rowe assessment. The new questionnaire and the Rowe clinical score each achieved a large standardised effect size (> or = 0.8) and compared favourably with relevant items on the SF36. By contrast, the Constant score barely registered any effect, confirming that it may be relatively insensitive to changes in clinical status for this particular condition. The questionnaire provides a measurement of outcome for shoulder instability which is short, practical, reliable, valid and sensitive to changes of clinical importance. PMID- 10872359 TI - Charnley low-frictional torque arthroplasty of the hip. 20-to-30 year results. AB - We reviewed 261 patients with 320 Charnley low-friction arthroplasties who had a mean follow-up of 22 years 10 months (20 to 30). Of these, 93.9% considered the operation to be a success; 82.3% were free from pain and 11.6% had occasional discomfort. Satisfactory function was achieved in 59.6% and 62% had an excellent range of movement. The clinical results did not correlate well with the radiological appearance; radiologically loose components did not affect the clinical outcome. The main long-term problem was wear and loosening of the UHMWPE cup. Our findings suggest that the radiological appearance of the arthroplasty is a more reliable indication of the state of the arthroplasty than the clinical results. PMID- 10872360 TI - Planar anteversion of the acetabular cup as determined from plain anteroposterior radiographs. AB - In total hip replacement, orientation of the cup is critical to the stability of the prosthesis. A new method to determine the angle of planar anteversion is described. A simple mathematical formula uses the measurements taken from anteroposterior radiographs to calculate the planar anteversion without reference to tables or charts. An experimental study in vitro has shown the efficacy of the formula in giving results which are within a clinically acceptable range. PMID- 10872361 TI - Total hip arthroplasty in patients with Down's syndrome. AB - Hip disease occurs in between 8% and 28% of patients with Down's syndrome, many of whom develop disabling pain. We have carried out total hip replacement in six adult patients (9 hips) with severe arthritis of the hip. The mean follow-up was 7.75 years (2 to 14). At the latest review, all had relief of pain and full hip function. Increasing longevity and a high incidence of hip disease in these patients suggest a greater role for total hip arthroplasty in the future. PMID- 10872362 TI - Effectiveness of bone cement containing tobramycin. An in vitro susceptibility study of 99 organisms found in infected joint arthroplasty. AB - We used 99 strains of organisms representative of orthopaedic infections to examine the effectiveness of a bone cement containing tobramycin, employing a modified in vitro Kirby-Bauer susceptibility model. The spectrum was broad, including Gram-positive and Gram-negative aerobic organisms, anaerobes and mycobacteria. Simplex P with added tobramycin was effective against most of the strains, including those which are resistant to typical systemic levels of tobramycin. Although direct correlation between in vitro and in vivo results is difficult, the study showed that tobramycin is stable to the exothermic polymerisation of the cement, and that it is released from the surface of the cement at concentrations high enough to inhibit the growth of most organisms which may be encountered after joint arthroplasty. PMID- 10872363 TI - Open-wedge osteotomy by hemicallotasis or the closed-wedge technique for osteoarthritis of the knee. A randomised study of 50 operations. AB - We describe the results of 50 operations carried out on 46 patients with medial osteoarthritis of the knee of Ahlback grade 1 to 3. Patients were randomised either to a closed-wedge high tibial osteotomy (HTO) or an open-wedge procedure based on the hemicallotasis technique (HCO). Their median age was 55 years (38 to 68). The preoperative median hip-knee-ankle (HKA) angle was 171 degrees (164 to 176) in the HTO group and 173 degrees (165 to 179) in the HCO group. After six weeks, the median HKA angle was 185 degrees (176 to 194) in the HTO group and 184 degrees (181 to 188) in the HCO group. In the HTO group, seven patients were within the range of 182 degrees to 186 degrees compared with 21 in the HCO group (p < 0.001). One year later, ten HTO patients were within this range while the HKA angulation in the HCO group was unchanged. At two years the numbers were 11 and 18, respectively. We evaluated the clinical results on the Hospital for Special Surgery, Lysholm and Wallgren-Tegner activity scores, and patients completed part of the Nottingham Health Profile questionnaire. An impartial observer at the two-year follow-up concluded that all scores had improved, but found no clinical differences between the groups. PMID- 10872364 TI - Hemicallotasis open-wedge osteotomy for osteoarthritis of the knee. Complications in 308 operations. AB - We studied the complications after open-wedge osteotomy by hemicallotasis in 308 consecutive patients, most of whom had osteoarthritis of the knee. The participating surgeons, who worked at 17 hospitals, used their discretion in selecting patients, operating techniques and external fixators. The general complications included 11 cases of deep-vein thrombosis (4%), six of nonunion (2%) and one of septic arthritis of the knee. There were technical complications in 13 patients (4%). In 157 patients (51%) pin-site infections were recorded; of these, 96% were minor and responded to wound toilet and antibiotic treatment. A total of 18 revision procedures was carried out. PMID- 10872365 TI - Anatomy and surface geometry of the patellofemoral joint in the axial plane. AB - We studied the anatomy of the patellofemoral joint in the axial plane on cryosections from a cadaver knee and on MR arthrotomograms from 30 patients. The cryosections revealed differences in the geometry and anatomy of the surface of the articular cartilage and corresponding subchondral osseous contours of the patellofemoral joint. On the MR arthrotomograms the surface geometry of the cartilage matched the osseous contour of the patella in only four of the 30 knees. The articular cartilaginous surface of the intercondylar sulcus and corresponding osseous contour of the femoral trochlea matched in only seven knees. Since MR arthrotomography can distinguish between the surface geometry of the articular cartilage and subchondral osseous anatomy of the patellofemoral joint, it allows the surgeon and the radiologist to appraise the true articulating surfaces. We therefore recommend MR arthrotomography as the imaging technique of choice. PMID- 10872366 TI - Cryptococcal osteomyelitis of the spine. AB - We have treated seven patients with cryptococcal spondylitis. Five presented with a neurological deficit and one was HIV-positive. Amphotericin-B and 5-flucytosine were used in five patients and ketoconazole was given orally in the remaining two. Three patients made a complete neurological recovery. Since these lesions mimic spinal tuberculosis, which is commonly seen in our environment, we draw attention to the importance of obtaining a tissue diagnosis. PMID- 10872367 TI - High cervical disc herniation and Brown-Sequard syndrome. A case report and review of the literature. AB - We describe a rare herniation of the disc at the C2/C3 level in a 73-year-old woman. It caused hemicompression of the spinal cord and led to the Brown-Sequard syndrome. The condition was diagnosed clinically and by MRI six months after onset. Discectomy and fusion gave complete neurological resolution. PMID- 10872368 TI - Short-course chemotherapy for tuberculosis of the spine. A comparison between ambulant treatment and radical surgery--ten-year report. AB - We performed a randomised, controlled clinical trial to compare ambulant short course chemotherapy with anterior spinal fusion plus short-course chemotherapy for spinal tuberculosis without paraplegia. Patients with active disease of vertebral bodies were randomly allocated to one of three regimens: a) radical anterior resection with bone grafting plus six months of daily isoniazid plus rifampicin (Rad6); b) ambulant chemotherapy for six months with daily isoniazid plus rifampicin (Amb6); or c) similar to b) but with chemotherapy for nine months (Amb9). Ten years from the onset of treatment, 90% of 78 Rad6, 94% of 78 Amb6 and 99% of 79 Amb9 patients had a favourable status. Ambulant chemotherapy for a period of six months with daily isoniazid plus rifampicin (Amb6) was an effective treatment for spinal tuberculosis except in patients aged less than 15 years with an initial angle of kyphosis of more than 30 degrees whose kyphosis increased substantially. PMID- 10872369 TI - Fixation of odontoid fractures by an anterior screw. AB - We have reviewed 81 patients with fractures of the odontoid process treated between May 1983 and July 1997, by anterior screw fixation. There were 29 patients with Anderson and D'Alonzo type-II fractures and 52 with type III. Roy Camille's classification identified the direction and instability of the fracture. Operative fixation was carried out on 48 men and 33 women with a mean age of 57 years. Associated injuries of the cervical spine were present in 15 patients, neurological signs in 13, and 18 had an Injury Severity Score of more than 15. Nine patients died and 11 were lost to follow-up. Of 61 patients, 56 (92%) achieved bony union at an average of 14.1 weeks. Two patients required a secondary posterior fusion after failure of the index operation. A full range of movement was restored in 43 patients; only six had a limitation of movement greater than 25%. We conclude that anterior screw fixation is effective and practicable in the treatment of fractures of the dens. PMID- 10872370 TI - Biomechanical comparison of fixation of type-I fractures of the lateral tibial plateau. Is the antiglide screw effective? AB - Type-I fractures of the lateral tibial plateau were simulated by osteotomy in 18 pairs of unembalmed cadaver tibiae. One fracture of each pair was fixed with two lag screws whereas the contralateral site was stabilised with three lag screws, or two lag screws plus an antiglide screw. The lateral plateau was displaced downwards using a servohydraulic materials testing machine and the resulting force and articular surface gap were recorded. Yield load was defined as the maximum load needed to create a 2.0 mm articular offset at the fracture line. The yield loads of the three-lag-screw (307 +/- 240 N) and antiglide constructs (342 +/- 249 N) were not significantly different from their two-screw control constructs (231 +/- 227 and 289 +/- 245 N, respectively). We concluded that adding an antiglide screw or a third lag screw did not provide any biomechanical advantage in stabilising these fractures. PMID- 10872371 TI - Dropped hallux after the intramedullary nailing of tibial fractures. AB - We made a prospective study of 208 patients with tibial fractures treated by reamed intramedullary nailing. Of these, 11 (5.3%) developed dysfunction of the peroneal nerve with no evidence of a compartment syndrome. The patients with this complication were significantly younger (mean age 25.6 years) and most had closed fractures of the forced-varus type with relatively minor soft-tissue damage. The fibula was intact in three, fractured in the distal or middle third in seven, with only one fracture in the proximal third. Eight of the 11 patients showed a 'dropped hallux' syndrome, with weakness of extensor hallucis longus and numbness in the first web space, but no clinical involvement of extensor digitorum longus or tibialis anterior. This was confirmed by nerve-conduction studies in three of the eight patients. There was good recovery of muscle function within three to four months in all cases, but after one year three patients still had some residual tightness of extensor hallucis longus, and two some numbness in the first web space. No patient required further treatment. PMID- 10872372 TI - The quadriceps myocutaneous flap for operation on the distal femur. AB - We describe a U-shaped approach to the distal femur which, having divided the extensor mechanism and elevated the entire quadriceps muscle, gives excellent exposure and allows a number of reconstructive options. It was used in 14 patients, 13 of whom were followed up for a mean of 3.5 years (1 to 11). There was no case of flap necrosis, and complications related to the reconstruction were acceptable. PMID- 10872373 TI - Endoprosthetic replacement of the proximal tibia. AB - We have performed endoprosthetic replacement after resection of tumours of the proximal tibia on 151 patients over a period of 20 years. During this period limb salvage surgery was achieved in 88% of patients with tumours of the proximal tibia. Both the implant and the operative technique have been gradually modified in order to reduce complications. An initial rate of infection of 36% has been reduced to 12% by the use of a flap of the medial gastrocnemius, to which the divided patellar tendon is attached. Loosening and breakage of the implant have been further causes of failure. We found that the probability of further surgical procedures being required was 70% at ten years and the risk of amputation, 25%. The development of a new rotating hinge endoprosthesis may lower the incidence of mechanical problems. Limb salvage for tumours of the proximal tibia is fraught with complications, but the good functional outcome in successful cases justifies its continued use. PMID- 10872374 TI - Extensible endoprostheses of the humerus after resection of bone tumours. AB - We carried out extensible endoprosthetic replacement of the proximal or total humerus in 18 children aged between six and 12 years, after resection of primary bone tumours mainly for osteosarcoma and Ewing's sarcoma. In 11 patients we performed 44 lengthening procedures, with an average of two per child annually and a mean total extension of 29.9 mm per patient. We were able to achieve lengthening of the operated limb with few complications and a mean functional rating of 79.3% according to the Enneking system. Progressive lengthening of these prostheses does not adversely affect the overall function of the arm, and superior subluxation of the head of the prosthesis has not been a problem. PMID- 10872375 TI - The Gorham-Stout syndrome (Gorham's massive osteolysis). A report of six cases with histopathological findings. AB - The Gorham-Stout Syndrome (Gorham's massive osteolysis) is a rare condition in which spontaneous, progressive resorption of bone occurs. The aetiology is poorly understood. We report six cases of the condition and present evidence that osteolysis is due to an increased number of stimulated osteoclasts. This suggests that early potent antiresorptive therapy such as with calcitonin or bisphosphonates may prevent local progressive osteolysis. PMID- 10872376 TI - Spatial and temporal distribution of CD44 and osteopontin in fracture callus. AB - The multifunctional adhesion molecule CD44 is a major cell-surface receptor for hyaluronic acid (HUA). Recent data suggest that it may also bind the ubiquitous bone-matrix protein, osteopontin (OPN). Because OPN has been shown to be a potentially important protein in bone remodelling, we investigated the hypothesis that OPN interactions with the CD44 receptor on bone cells participate in the regulation of the healing of fractures. We examined the spatial and temporal patterns of expression of OPN and CD44 in healing fractures of rat femora by in situ hybridisation and immunohistochemistry. We also localised HUA in the fracture callus using biotinylated HUA-binding protein. OPN was expressed in remodelling areas of the hard callus and was found in osteocytes, osteoclasts and osteoprogenitor cells, but not in cuboidal osteoblasts which were otherwise shown to express osteocalcin. The OPN signal in osteocytes was not uniformly distributed, but was restricted to specific regions near sites where OPN mRNA positive osteoclasts were attached to bone surfaces. In the remodelling callus, intense immunostaining for CD44 was detected in osteocyte lacunae, along canaliculi, and on the basolateral plasma membrane of osteoclasts, but not in the cuboidal osteoblasts. HUA staining was detected in fibrous tissues but little was observed in areas of hard callus where bone remodelling was progressing. Our findings suggest that OPN, rather than HUA, is the major ligand for CD44 on bone cells in the remodelling phase of healing of fractures. They also raise the possibility that such interactions may be involved in the communication of osteocytes with each other and with osteoclasts on bone surfaces. The interactions between CD44 and OPN may have important clinical implications in the repair of skeletal tissues. PMID- 10872377 TI - Cytotoxicity and macrophage cytokine release induced by ceramic and polyethylene particles in vitro. AB - Although the response of macrophages to polyethylene debris has been widely studied, it has never been compared with the cellular response to ceramic debris. Our aim was to investigate the cytotoxicity of ceramic particles (Al2O3 and ZrO2) and to analyse their ability to stimulate the release of inflammatory mediators compared with that of high-density polyethylene particles (HDP). We analysed the effects of particle size, concentration and composition using an in vitro model. The J774 mouse macrophage cell line was exposed to commercial particles in the phagocytosable range (up to 4.5 microns). Al2O3 was compared with ZrO2 at 0.6 micron and with HDP at 4.5 microns. Cytotoxicity tests were performed using flow cytometry and macrophage cytokine release was measured by ELISA. Cell mortality increased with the size and concentration of Al2O3 particles. When comparing Al2O3 and ZrO2 at 0.6 micron, we did not detect any significant difference at the concentrations analysed (up to 2500 particles per macrophage), and mortality remained very low (less than 10%). Release of TNF-alpha also increased with the size and concentration of Al2O3 particles, reaching 195% of control (165 pg/ml v 84 pg/ml) at 2.4 microns and 350 particles per cell (p < 0.05). Release of TNF alpha was higher with HDP than with Al2O3 particles at 4.5 microns. However, we did not detect any significant difference in the release of TNF-alpha between Al2O3 and ZrO2 at 0.6 micron (p > 0.05). We saw no evidence of release of interleukin-1 alpha or interleukin-1 beta after exposure to ceramic or HDP particles. PMID- 10872378 TI - Intracellular biogenesis of collagen fibrils in 'activated fibroblasts' of tendo Achillis. An ultrastructural study in the New Zealand rabbit. AB - We have studied the formation of collagen fibrils in 'activated fibroblasts' of tendo Achillis of rabbits. The tendon was in the process of regeneration after experimental partial tenotomy. Samples were taken from the peri-incisional region and analysed by transmission electron microscopy. Ultrastructural examination showed the presence of a 'fine dense granular substance' inside the rough endoplasmic reticulum and procollagen filaments. These come together to form collagen fibrils in the dilated vacuoles of the rough endoplasmic reticulum. The possible intra- and extracellular origin of collagen fibrils is suggested. Within the cell biosynthesis of collagen fibrils take place with the formation of collagen substance which gives rise to procollagen filaments. These make contact in parallel apposition to produce striated 'spindle-shaped bodies' which elongate by the longitudinal attachment of more procollagen filaments and form intracellular nascent collagen fibrils. PMID- 10872379 TI - Nitric oxide in fracture repair. Differential localisation, expression and activity of nitric oxide synthases. AB - Our aim was to investigate whether nitric oxide synthase (NOS) isoforms, responsible for the generation of NO, are expressed during the healing of fractures. To localise the sites of expression compared with those in normal bone we made standardised, stabilised, unilateral tibial fractures in male Wistar rats. Immunostaining was used to determine the precise tissue localisation of the different NOS isoforms. Western blotting was used to assess expression of NOS isoform protein and L-citrulline assays for studies on NOS activity. Control tissue was obtained from both the contralateral uninjured limb and limbs of normal rats. Immunohistochemistry showed increased expression of endothelial NOS (eNOS) to be strongest in the cortical blood vessels and in osteocytes in the early phase of fracture repair. Western blot and image analysis confirmed this initial increase. Significantly elevated calcium-dependent NOS activity was observed at day 1 after fracture. Inducible NOS (iNOS) was localised principally in endosteal osteoblasts and was also seen in chondroblasts especially in the second week of fracture healing. Western blotting showed a reduction in iNOS during the early healing period. Significantly reduced calcium-independent NOS activity was also seen. No neuronal NOS was seen in either fracture or normal tissue. Increased eNOS in bone blood vessels is likely to mediate the increased blood flow recognised during fracture healing. eNOS expression in osteocytes may occur in response to changes in either mechanical or local fluid shear stress. The finding that eNOS is increased and iNOS reduced in early healing of fractures may be important in their successful repair. PMID- 10872380 TI - Human bone allografts can induce T cells with high affinity for donor antigens. AB - We analysed the cellular immune response in ten transplantations of different massive bone allografts, of which five had a poor clinical outcome. Cytotoxic T lymphocytes (CTL) and T helper lymphocytes (TH) against mismatched donor antigens were found in all patients. More importantly, CTL with a high affinity for donor antigens were found in five cases. High-affinity CTL need no CD8 molecule to stabilise the antigen binding and are strongly associated with rejection of heart and corneal transplants. Even after removal of most of the bone-marrow cells, we found high-affinity CTL and high TH frequencies. This T-cell response could be detected over a period of years. We conclude that frozen bone allografts can induce high-affinity donor-specific CTL. The present assay allows qualification and quantification of the levels of CTL and TH in the blood. This approach may be helpful in studying the effect of the immune response on the outcome of the graft. PMID- 10872381 TI - Cytotoxic effect of methotrexate and its solvent on osteosarcoma cells in vitro. AB - Bone tumours may recur locally even after wide surgical excision and systemic chemotherapy. Local control of growth may be accomplished by the addition of cytostatic drugs such as methotrexate (MTX) to bone cement used to fill the defect after surgery and to stabilise the reconstructive prosthesis. We have studied the elution kinetics of MTX and its solvent N-methyl-pyrrolidone (NMP) from bone cement and their biological activities in five cell lines of osteosarcoma and in osteoblasts, and compared them with the effects of the parent compounds alone and in combination. Our findings show that MTX is released continuously over months at concentrations highly cytotoxic to osteosarcoma cells and suggest that the impregnated bone cement would be effective in the long term. Proliferating osteoblasts, however, were much less sensitive towards MTX. The dose-response relationship for NMP and experiments with MTX/NMP-mixtures show that the eluted concentrations of solvent are not toxic and do not influence the effects of MTX. We suggest that bone cement containing MTX dissolved in NMP releases the drug in a suitable and effective way and may be of value in the treatment of bone tumours. PMID- 10872382 TI - Deposition of calcium pyrophosphate in tissue after revision arthroplasty of the hip. AB - We reviewed histologically the incidence and pathogenesis of the deposition of calcium pyrophosphate dihydrate (CPPD) crystals in the pseudocapsule, femoral and acetabular membranes and periprosthetic tissue at revision of 789 cases of failed total hip replacement. In 13, periprosthetic tissues were found to have deposits of CPPD crystals in areas of cartilaginous metaplasia; four also showed evidence of localised deposition of amyloid. None of the patients had a history of chondrocalcinosis in the hip or other joints. Cartilaginous metaplasia and other changes in periprosthetic tissues may predispose to the deposition of CPPD and associated localised amyloid. PMID- 10872383 TI - Comparison of the Wrightington FC hip with the Charnley low-friction arthroplasty. PMID- 10872384 TI - The control of new prosthetic implants. PMID- 10872385 TI - The control of new prosthetic implants. PMID- 10872386 TI - Femoral stem fixation. PMID- 10872387 TI - Trauma and orthopaedic surgery on the Internet. PMID- 10872388 TI - Total knee arthroplasty with the PFC system. PMID- 10872389 TI - The protective effect of a cut-resistant glove liner. PMID- 10872390 TI - Determining the sagittal dimensions of the canal of the cervical spine. PMID- 10872391 TI - Neonatal detection of developmental dysplasia of the hip (DDH) PMID- 10872392 TI - The influence of salivary flow rate on diffusion of potassium chloride from artificial plaque at different sites in the mouth. AB - The rate at which substances diffuse from dental plaque influences the rate of clearance of acid and bacterial toxins from plaque into saliva. The aim of this study was to compare the rates of clearance of potassium chloride, as a model substance, from artificial plaque of 3-, 4-, and 6-mm-diameter, positioned bilaterally at different locations in the mouth. The diffusant was KCl (1 mol/L) in a 1.0% agarose matrix, placed in wells 1.5 mm deep, in small acrylic devices 3 mm thick, which could be fastened to the teeth with dental floss and removed after different time periods. The half-time for clearance was determined from the best-fitting least-squares line of the potassium concentration remaining in the gel plotted against the square root of time. For 14 subjects, half-times for the lower anterior lingual and upper posterior lingual regions averaged about 2.5 times greater than those for clearance into a large, stirred volume in vitro, whereas those for the upper and lower anterior buccal regions averaged about 12.8 times greater. This difference may be due to the fact that anterior buccal sites are exposed only to minor rather than to major salivary gland secretions. When salivary flow was stimulated by the sucking of sour lemon drops, all in vivo half times were reduced by about one-half. The half-times were also directly related to the surface areas of the chambers, which implies that rates of diffusion from plaque of substances such as acid or bacterial toxins are inversely related to the surface area of the plaque at a particular site. PMID- 10872393 TI - Sialochemistry of whole, parotid, and labial minor gland saliva in patients with oral lichen planus. AB - This study was undertaken to determine whether oral lichen planus in otherwise healthy patients is associated with sialochemical abnormalities. Unstimulated and stimulated whole saliva, stimulated parotid saliva, and stimulated labial minor gland saliva were collected from 25 patients with oral lichen planus and from 25 age- and sex-matched controls. Flow rate and salivary concentrations of immunoglobulins A and G, albumin, amylase, lysozyme, lactoferrin, and total protein were determined by standard analytical techniques. Concentrations of inorganic components including sodium, potassium, calcium, chloride, and phosphate were also measured. No significant differences were found between the lichen planus patients and the controls. These findings do not support an association between oral lichen planus and salivary dysfunction in otherwise healthy patients. PMID- 10872394 TI - Developmental changes of esteroprotease and androgen receptors in the mouse submandibular gland. AB - The activities of p-tosyl-L-arginine methyl ester (TAMEase) and cytosolic and nuclear androgen receptors in the submandibular glands of male and female mice were determined at various developmental stages. In males, the activity of TAMEase was detectable at four weeks after birth, and thereafter it increased rapidly. Cytosolic androgen receptor increased gradually with age, whereas nuclear androgen receptor, which was minimal one week after birth, increased remarkably four weeks after birth. In females, the minimal activity of TAMEase was detectable at five weeks after birth and increased very slowly with age. Cytosolic receptor increased with age, but nuclear receptor level was unchanged. These findings suggest that the appearance of TAMEase was in accordance with an elevation of nuclear androgen receptor in mice submandibular glands. PMID- 10872395 TI - Immunocytochemical evidence for the coexistence of catecholestrogen and catechol O-methyltransferase in the rat parotid gland. AB - Catechol-O-methyltransferase (COMT) (EC 2.1.1.6) and catecholestrogen were localized in the parotid gland of the rat by immunocytochemical methods. Specific immunoreactive deposits for COMT and catecholestrogen were found in the cytoplasm of duct cells, but only those for COMT in myo-epithelial cells. The pattern of localization of COMT and catecholestrogen in the parotid gland suggests a functional relationship between COMT and catecholestrogen. PMID- 10872396 TI - Cost-effectiveness analysis of periodontal disease control. AB - Cost-effectiveness analysis was used to evaluate alternative methods of periodontal disease control. The alternatives considered included non-surgical and surgical procedures as well as the use of antimicrobial agents. Data on costs were obtained from American Dental Association publications of average charges for periodontal services. The concept of quality-adjusted tooth-years (QATYs) was developed to provide an outcome measure which could be compared across treatments. The conclusions of this analysis are as follows: (1) Conservative non surgical treatments for periodontal disease control not only have costs lower than surgical alternatives, as would be expected, but also maximize expected quality-adjusted tooth-years over a wide range of estimates; (2) antimicrobial therapy used as an adjunct to non-surgical treatment is likely to be both effective and cost-effective; and (3) quality of tooth-years is a critical consideration in the determination of outcome of periodontal treatment. For example, when tooth-years are not adjusted for quality, differences between treatments are diminished, and surgical treatment becomes as good as or better than more conservative treatments for some levels of disease severity. PMID- 10872397 TI - Setting stress in composite resin in relation to configuration of the restoration. AB - The setting stress in composite resins was studied as a function of restoration shape. The shape is described by the configuration factor, C, the ratio of the restoration's bonded to unbonded (free) surfaces. In an experimental set-up, the shape of the restoration was simulated by cylindrical forms of various dimensions. The shrinkage stress was measured continuously. It was shown that in most of the clinically relevant cavity configurations, the stress-relieving flow is not sufficient to preserve adhesion to dentin by dentin-bonding agents. PMID- 10872399 TI - The effect of baseline lesion mineral loss on in situ remineralization. AB - The effect of baseline lesion mineral loss on the remineralization of enamel lesions by a sodium fluoride dentifrice was studied in situ by means of an appliance carrying enamel sections. Artificial lesions of various sizes were created, by means of acidified gelatin, and were then mounted on the appliances of five volunteers. Each brushed twice daily for two min with a 1000 ppm F sodium fluoride dentifrice. Measurements of mineral content were made at baseline and at weekly intervals by microradiographic/microdensitometric techniques. Data from all five volunteers showed a linear increase in remineralization rate with increasing lesion size. Thus, in studies which compare the effects of different remineralizing formulations, care must be taken to ensure that initial lesion sizes are matched, or that the results are expressed as a percentage change in mineral content. PMID- 10872398 TI - The use of saturated DCPD in remineralization of artificial caries lesions in vitro. AB - Dicalcium phosphate dihydrate (DCPD) may play a significant role in the caries lesion since it is a stable calcium phosphate phase under acidic conditions. The reaction of DCPD and fluoride, forming fluorapatite (FAP), may provide a potentially promising treatment regimen for remineralization of caries lesions in vivo. The purpose of this study was to determine whether a two-step DCPD and inorganic wash with fluoride can remineralize artificial caries-like lesions in vitro. We used the single-section technique to facilitate quantitation of the same tissue before and after the experimental regimen. The two-step remineralizing treatment was repeated three times and consisted of a two-minute saturated DCPD treatment (pH 2.1) followed by a 24-hour inorganic wash. Lesion parameters were recorded before and after treatment by the taking of polarized light photomicrographs of each section after imbibition in several media. The changes in the tissue following treatment were expressed as a percent change in the area of the initial pre-treatment lesion. Significant reductions (p < 0.02) in lesion pore volume were observed in all aqueous media examined. In the lesions after imbibition in quinoline, remineralization was also apparent from the significant increase in the area of the dark zone following treatment. This two step DCPD treatment appears to remineralize artificial caries-like lesions effectively, but additional work is needed to determine whether it affords any protection against subsequent cariogenic challenges. PMID- 10872400 TI - Intracrystalline structure of enamel crystals affected by caries. AB - Enamel crystals in the demineralized zones in early caries lesions of human teeth were observed by high-resolution electron microscopy. The enamel crystals frequently exhibited perforations in their centers and defects of various sizes on their lateral surfaces. There were a number of small electron-lucent spots, suggesting that the dissolution of crystals had taken place there. These spots were in especially large numbers near the central dark line. The central perforations, the lateral defects, and the small spots had a common habit which formed regularly along the crystalline a- and b-axes. In many cases, when the central dark line was seen, the perforations were located a few unit cells away from the line. The perforations seem to result from a fusion of small spots, which enlarge by involving other small spots. The lateral defect seemed to enlarge by removal of unit cells and progression along the a- and b-axes. In the regions where the small spots were present, however, the enlargement of the defects also progressed involving the spots. The central dark line seems to be rather resistant to dissolution. One of the main factors for the central perforation of the crystals is thought to be the presence there of especially large numbers of defective sites. PMID- 10872401 TI - Early microbial colonization of permucosal implants in edentulous patients. AB - In edentulous patients, the microbial colonization of permucosal implants of sintered hydroxyapatite was studied. Samples were taken from mucosa and dentures before insertion of implants and from supra- and subgingival sites two to 10 weeks after insertion. In total, five patients and 10 implants with clinically healthy peri-implant tissues were studied. The samples were investigated by dark field microscopy and anaerobic culture. The supragingival plaque of the implants was dominated by Gram-positive cocci and rods, the subgingival plaque by Haemophilus spp. and Veillonella parvula. A group of bacteria was found specifically related to the implants: Actinomyces odontolyticus, Peptostreptococcus micros, Haemophilus actinomycetemcomitans, Eikenella corrodens, Capnocytophaga sputigena, and Leptotrichia buccalis. Black-pigmented Bacteroides was not found in any of the examined samples. Spirochetes were observed in denture plaque samples and in supragingival plaque of the implants. It is concluded that bacteria known as potential periodontal pathogens colonize the permucosal implants in the first weeks after insertion. The presence of these species seems to be dependent on the ecological factors provided by the artificial gingival crevice of the permucosal implants in the edentulous mouth. PMID- 10872402 TI - Comparison of the effects of fluoride and the ionophore nigericin on acid production by Streptococcus mutans and the resultant in vitro enamel demineralization. AB - In an in vitro demineralization experiment with a plaque of S. mutans C180-2 over enamel, the effect of 0.5 mmol fluoride/L was compared with that of 10 nmol nigericin/L. Since the effects of both substances on the acid production in a dense cell suspension were of the same magnitude, and since nigericin did not affect the enamel solubility, the present demineralization experiment may differentiate between the effect of fluoride on bacterial metabolism alone and the combined effect of fluoride on both the bacterial metabolism and enamel solubility. In our experiments, the effect on bacterial metabolism accounted for 75% of the caries-inhibiting effect of fluoride. Microradiographic data showed subsurface lesions in the control and nigericin group and a slight surface softening in the fluoride group. In addition, the effects of fluoride and nigericin on the cellular internal pH during glycolysis were studied. Analysis of these data suggested that glycolysis was inhibited when the intracellular pH reached about pH 5.2, which supports the view that acidification of the cell cytoplasm is a mechanism by which fluoride inhibits glycolysis. PMID- 10872403 TI - Cross-inhibition between black-pigmented Bacteroides species. AB - Cross-inhibition within the group of black-pigmented Bacteroides, including both oral and non-oral strains, was studied by means of a membrane filter technique. It was found that B. gingivalis possessed the most extended inhibitory capacity among all species tested. B. gingivalis showed inhibitory activity against B. intermedius, B. endodontalis, B. loescheii, and B. melaninogenicus. B. endodontalis was active against some B. intermedius strains. Among the saccharolytic species, some B. melaninogenicus strains were inhibitory for some B. endodontalis strains, some B. gingivalis strains, and some B. intermedius strains. These inhibitory activities observed in vitro may play a role in the colonization of the periodontal pocket. PMID- 10872404 TI - Characteristics of trypsin-like activity in subgingival plaque samples. AB - Previous studies have demonstrated that the hydrolysis of the trypsin substrate N benzoyl-DL-arginine-2-naphthylamide (BANA), by subgingival plaque obtained from a single site, correlates best with the numbers and proportions of spirochetes in plaque samples and may serve as an indicator of clinical disease. In this investigation, we determined whether the association between BANA hydrolysis and spirochetes could be obtained in pooled subgingival plaque samples. Concomitantly, the characteristics of this reaction in terms of substrate type and concentration, microbial numbers needed to give a positive reaction as assessed by microscopic counts, rapidity of hydrolysis, and the effect of pH and various additives on the plaque BANA hydrolytic activity have been studied in pooled plaque samples from patients who were periodontally healthy or diseased. In addition, it was determined whether BANA hydrolytic activity found in subgingival plaque reflected contributions from saliva and supragingival plaque. Results indicated that the assay can best be performed with 0.67 mmol/L BANA at pH 7.0. EDTA and CaCl2 gave a slight inhibition and DTT a slight enhancement of the BANA reaction by the pooled plaque suspensions. The majority of the reactions (85%) developed their full color after overnight incubation. BANA hydrolysis was not found in saliva and occurred with much greater frequency in subgingival plaque as opposed to supragingival plaque. Analysis of the data indicated that BANA hydrolysis by pooled subgingival plaque samples is a suitable test for the detection of spirochetes when two or three spirochetes per high microscopic field are present in the sample. PMID- 10872405 TI - Regional lymph node metastasis created by partial excision of carcinomas induced in hamster cheek pouch with 9,10-dimethyl-1,2-benzanthracene. AB - The feasibility of using the hamster cheek pouch/dimethyl-benzanthracene (DMBA) system as an experimental model of lymphatic metastasis was investigated. Forty male Syrian golden hamsters treated with DMBA were divided into two equal groups- one with surgical excision of their tumors and a control group without tumor excision. In the excision group, the animals received three applications/week to the left cheek pouch of 0.3% DMBA in acetone for 14 weeks. Following a three-week observation period, the tumors in the pouch were excised at their base, and the animals were killed after four weeks of further observation. In the control group, the animals were treated for 14 weeks in a manner similar to that used for the excision group, left for seven weeks without treatment, and then killed. Cheek pouches with tumors and cervical lymph nodes were processed for histological examination. All of the animals, both with and without metastasis, had borne squamous cell carcinomas (SCC) in their treated cheek pouches. Histologically, seven out of 16 animals in the excision group showed metastatic deposits of SCC confined to the left cervical lymph nodes, while in the control group, metastasis was not found in any of the 19 animals with SCC in their cheek pouches. The results demonstrate that surgical excision of the hamster cheek pouch carcinoma is efficient in producing unequivocal lymph node metastasis. PMID- 10872406 TI - Comparison of pain associated with mechanical and chemomechanical removal of caries. AB - Previous studies have indicated that a chemomechanical caries removal system (CRS) has been effective in minimizing the use of conventional mechanical instruments and that it may reduce the need for local anesthesia. In the present study, a comparison of the pain experienced both during treatment with a chemomechanical technique and during conventional caries removal (control) was made for each of 47 patients who initially were not given a local anesthetic. One of two dentists was randomly selected to examine and treat patients with a matched pair of carious teeth, and each pair of teeth was treated in a randomized order with the CRS or control procedure. Responses to the McGill Pain Questionnaire (MPQ) revealed a significantly higher level of pain (p < 0.025) associated with the conventional treatment compared with the chemomechanical procedure. A significantly greater number of patients (p < 0.05) requested local anesthetic for the tooth subjected to the control procedure than for the tooth subjected to the CRS procedure. However, 72.3% of the patients did not request local anesthesia for either treatment, although pain was experienced by these patients in 46.8% of the control teeth and 27.7% of the teeth which received the CRS treatment. Of the 20 pain descriptor categories listed on the MPQ, the sensory categories accounted for the greatest mean number of pain descriptors selected for both the CRS (4.4) and control procedures (8.0), compared with the mean number of descriptors selected from the affective pain categories for the CRS (0.8) and control procedures (1.0). The results suggest that sensory pain factors are more important than affective pain factors in controlling the overall discomfort of patients and the need for local anesthetic during caries removal and subsequent restorative procedures. PMID- 10872407 TI - A cephalometric study of young marmosets (Callithrix jacchus). AB - Linear and angular measurements of young marmosets taken at three-month intervals from a series of cephalograms are presented. They show that linear craniofacial development was largely complete by six months of age, subsequent angular changes being mainly related to dental development. No sexual dimorphism was apparent, and there was substantial homogeneity in results for animals of the same age. Thus, marmosets, with their low cost, ease of handling, and rapid maturation, may provide a useful animal model for craniofacial research. PMID- 10872409 TI - Micro-electrode techniques for the analysis of oral fluids. AB - This paper describes the use of micro-electrodes for the analysis of small fluid volumes recovered from the oral environment. The analysis has several advantages: (1) It directly measures the activity of ions, a quantity more relevant to mineral saturation than the conventionally measured concentration, (2) minimum fluid volume for analysis is usually less than 0.005 microL, small enough to avoid sample pooling in most analyses, (3) numerous ions can be measured simultaneously, (4) the analysis time is very short, and (5) the use of mineral oil to isolate specimens provides a simple method for controlling the CO2 tension and humidity over the specimens. PMID- 10872408 TI - Lack of genotoxic effects of fluoride in the mouse bone-marrow micronucleus test. AB - The purpose of this investigation was to examine the potential genotoxic influence of sodium fluoride (NaF) on mammalian cells by means of a mouse bone marrow micronucleus test. Mice of genotype B6C3F1 were obtained at about eight weeks of age and maintained on a low-fluoride diet (< 0.2 ppm F) and distilled water ad libitum throughout the experiment. At approximately 12 weeks of age, the animals were randomly assigned to seven groups, with multiple sampling schedules. The animals were intubated with various doses of NaF, ranging from 0.1 mg/kg to the Maximum Tolerable Dose (MTD), with sampling at 30, 48, and 72 hours after treatment. Negative (distilled water) and positive (cyclophosphamide) controls were included. Coded slides of femur marrow cells were prepared and examined, without identification as to treatments, for the frequency of micronucleated polychromatic erythrocytes (MN-PCE). The humera were analyzed for fluoride for monitoring of the absorption of fluoride following oral intubation. The results indicated that doses of NaF up to the MTD did not significantly increase the frequencies of MN-PCE when compared with the negative controls, although the bone fluoride content increased as the dose of NaF was increased. PMID- 10872410 TI - Setting the record straight. PMID- 10872411 TI - Hereditary pancreatitis. Historical perspectives. AB - Clinically, hereditary pancreatitis was not distinguishable from any other cause of pancreatitis. But astute clinical observations demonstrated an evolution toward chronic pancreatitis that could develop into carcinoma in some patients. A chromosomal abnormality was identified on chromosome 7q35, and then three separate genetic abnormalities were identified. It is now understood that a defect in trypsinogen is at the basis of the anomaly, and further developments should help identify new therapeutic approaches. PMID- 10872412 TI - Genetic predispositions to acute and chronic pancreatitis. AB - Advances in molecular genetics have provided the powerful tools necessary to identify the key molecules and mechanisms that underly the disease process. Continued work in this area promises to reveal new insights as new disease genes are discovered. This article focuses on the insights into the cause of acute and chronic pancreatitis gained by investigation of the HP genes, the diagnosis of the known mutations, the fascinating observation of nonpenetrance, and a look at future directions. PMID- 10872413 TI - Early trypsinogen activation in acute pancreatitis. AB - This article discusses zymogen activation within the acinar cell. The authors review advances in respect to the cellular mechanism involved in a premature intrapancreatic protease activation. Critical factors that determine the onset of premature protease activation appear to be the molecular structure of trypsinogen, the presence or absence of functionally intact lysosomal hydrolases, the pH in intracellular compartments, and the calcium signaling cascade in the pancreatic acinar cell. PMID- 10872414 TI - Risk factors for cancer in hereditary pancreatitis. International Hereditary Pancreatitis Study Group. AB - Hereditary pancreatitis is a rare form of pancreatitis, accounting for approximately 1% of all types of pancreatitis. It is inherited as an autosomal dominant disease, with incomplete penetrance. The genetic defect is believed to be caused by mutations in the trypsinogen gene. Patients who inherit the disorder suffer from a form of pancreatitis that resembles other types of pancreatitis, but the age of onset is much earlier. Sixteen biopsy-proven pancreatic cancers have developed in a cohort of 412 patients with a median follow-up period of 18 years (interquartile range, 7 to 30 years) since the onset of symptoms. Compared with the background population, the risk of pancreatic cancer is approximately 50 to 60 times greater than expected. Smoking appears to be an additional risk factor in these patients: Smoking increases the risk of developing pancreatic cancer and lowers the age of onset by approximately 20 years. Patients with hereditary pancreatitis are urged to avoid smoking because it greatly increases the risk of pancreatic cancer and to avoid alcohol, a known risk factor for all forms of pancreatitis. PMID- 10872415 TI - Genetic testing. Counseling, laboratory, and regulatory issues and the EUROPAC protocol for ethical research in multicenter studies of inherited pancreatic diseases. AB - This article highlights several of the important issues and illustrates a European protocol that should be considered when offering genetic testing on a research or clinical basis for HP, as well as for other inherited disorders of the pancreas. PMID- 10872416 TI - Preventive strategies and therapeutic options for hereditary pancreatitis. AB - Much has been learned about hereditary pancreatitis. Much still remains to be explained, including the characteristic 20% nonpenetrance, variable expressivity, and factors affecting risk for pancreatic cancer. There is much work to be done. PMID- 10872417 TI - Genotype-phenotype relationships in cystic fibrosis. AB - The genotype-phenotype relationship in CF is complex despite its being a monogenic disorder. Factors that contribute to variability among individuals with the same genotype are an area of intense study. Nevertheless, certain conclusions can be derived from these studies. First, mutations in both CFTR alleles cause the CF phenotype. Homozygosity for delta F508 or compound heterozygosity for delta F508 and another severe mutation (e.g., G551D, W1282X) cause classic CF: obstructive pulmonary disease, exocrine pancreatic deficiency, male infertility, and elevated sweat chloride concentrations. Clinical variability is observed among patients with the classic form of CF, especially with regards to the severity of lung disease. Although understanding of the role of other genes and environment in the development of lung disease is incomplete, evidence that other factors are important raises the possibility that therapeutic intervention may be possible at several levels. Second, genotype correlates more closely with certain features of the CF phenotype than others. Mutations that allow partial function of CFTR are often associated with pancreatic sufficiency, occasionally identified with normal sweat gland function, and sporadically correlated with mild lung disease. Partially functioning mutants rarely prevent maldevelopment of the male reproductive tract; an exception is 3849 + 10 Kb C-->T. These observations suggest that certain tissues require different levels of CFTR function to avoid the pathologic manifestations typical of CF. The genetic cause of several disorders that clinically overlap CF can be attributed, in part, to mutations in CFTR. Finally, molecular analysis of disease-associated mutations identified through genotype-phenotype studies provides a mechanistic framework for genotype based therapeutic approaches and pharmaceutical interventions. PMID- 10872418 TI - Pancreatic aspects of cystic fibrosis and other inherited causes of pancreatic dysfunction. AB - Causes of pancreatic dysfunction in childhood can be divided into two general categories: (1) hereditary conditions that directly affect the pancreas and (2) acquired disorders in which loss of pancreatic function is a secondary phenomenon. This article discusses genotypes and phenotypes, clinical features, Shwachman-Diamond syndrome, isolated enzyme deficiencies, and other topics. PMID- 10872419 TI - Cystic fibrosis mutations and genetic predisposition to idiopathic chronic pancreatitis. AB - Idiopathic chronic pancreatitis is a leading cause of chronic pancreatitis. Work from this and other groups has shown that idiopathic chronic pancreatitis is associated with mutations of the cystic fibrosis gene (CFTR). Many idiopathic pancreatitis patients have compound heterozygote genotypes in which both copies of the CFTR gene are abnormal. In these patients, the pancreatic disease can be viewed as a mild variant of cystic fibrosis, in which there is sufficient residual CFTR function to prevent lung disease. This article summarizes the evidence associating these abnormal CFTR genotypes with idiopathic chronic pancreatitis and reviews the implications of this association for the pathogenesis, classification, and prevention of pancreatitis. PMID- 10872420 TI - Mechanisms underlying regulated CFTR trafficking. AB - Stimulation of membrane capacitance and cell surface labeling of epitope-tagged CFTR provide evidence of cAMP-regulated CFTR trafficking. Co-expression of syntaxin 1A inhibits cAMP-stimulated current and capacitance changes in CFTR expressing cells and blocks cAMP-induced increases in cell surface CFTR. Inhibition of CFTR trafficking by syntaxin over-expression suggests a role for SNARE proteins in this process. CFTR phosphorylation may alter physical interactions with SNARE proteins to regulate plasma membrane CFTR density. PMID- 10872421 TI - How cystic fibrosis affects pancreatic ductal bicarbonate secretion. AB - Pancreatic bicarbonate secretion is impaired in patients with cystic fibrosis. This article reviews recent advances in bicarbonate dependent transporters in pancreatic duct cells and discusses their regulation in cystic fibrosis. PMID- 10872422 TI - Mechanisms to explain pancreatic dysfunction in cystic fibrosis. AB - This article focuses on three potential mechanisms by which pancreatic dysfunction occurs in cystic fibrosis. These include (1) obstruction of pancreatic ducts by inspissated plugs, (2) inhibition of endocytosis in acinar cells, and (3) imbalance in membrane lipids in cystic fibrosis regulated cells. Any of these abnormalities alone or in combination may explain the development of pancreatic exocrine insufficiency. PMID- 10872423 TI - Hereditary pancreatic adenocarcinoma. A clinical perspective. AB - Although the total number of patients in these various high-risk groups is relatively small, they nevertheless provide excellent models for studying the cause, natural history, pathogenesis, and treatment of pancreatic cancer. These patients would also benefit greatly from procedures capable of detecting cancer at an early stage. This knowledge would be useful for the much commoner sporadic form of pancreatic cancer, in which diagnosis is almost always late and prognosis fatal. With early diagnosis, surgical resection before the cancer's extension beyond the organ's anatomic confines could be curative. The establishment of a National Familial Pancreatic Cancer Registry is essential and would increase the availability of these invaluable families for medical research. PMID- 10872424 TI - Inheritance of pancreatic cancer in pancreatic cancer-prone families. AB - Families are being increasingly recognized as carrying an inherited susceptibility for pancreatic cancer, apparently unrelated to any currently recognized syndrome. The authors provide a review of the current evidence for familial susceptibility to pancreatic cancer. A formal segregation analysis of the pattern of inheritance of pancreatic cancer in 70 families from the National Registry for Familial Pancreatic Cancer is described. This analysis suggests a single major gene with an autosomal dominant mode of inheritance controlling susceptibility for pancreatic cancer in these families. PMID- 10872425 TI - Molecular genetic alterations in ductal pancreatic adenocarcinomas. AB - Pancreatic ductal adenocarcinomas are known to harbor a distinct variety of genetic alterations in oncogenes, tumor-suppressor genes, and occasionally genes that carry out DNA mismatch repair. Although this malignancy occurs at an elevated frequency in patients with familial recurrent acute pancreatitis, the genetic alterations of these particular tumors have not been reported. The changes are likely to be similar to those of sporadic pancreatic cancer; if so, this would provide useful clues for studying the progression of early and advanced neoplasia in such pancreatitis patients to aid their clinical monitoring and provision of therapeutic recommendations. PMID- 10872426 TI - Growth factors, receptors, and molecular alterations in pancreatic cancer. Putting it all together. AB - Because of the dismal prognosis of advanced ductal pancreatic adenocarcinoma, recent investigational strategies have focused on improved detection and therapeutic intervention in early-stage pancreatic cancer. The obvious cost constraints of screening populations at risk but with a low tumor yield will restrict screening protocols to only the highest risk groups (hereditary pancreatitis = age 50, certain hereditary pancreatic cancer kindreds). The vast majority of patients, either lacking or exhibiting an inherited predisposition to pancreatic cancer, will continue to present with disease not resectable for cure. The authors believe that the best hope for these patients lies in the further delineation of the integrative pathophysiology driving tumor growth; this would facilitate the future development of a computer program or other modality that would predict the dominant pathways driving the growth and spread of each tumor based on its "molecular profile." This article reviews the authors' current knowledge regarding the growth factors, receptors, and molecular alterations driving uncontrolled proliferation, local invasion, and metastatic spread of these tumors. The current and potential contributions of studies in cohorts with an inherited predisposition to pancreatic cancer to this pathophysiologic model are also discussed. The future strategy for incorporating this information into a working pathophysiologic road map with clinical relevance is subsequently outlined. PMID- 10872427 TI - Cancer surveillance of patients from familial pancreatic cancer kindreds. AB - The family history can be used to determine which family members warrant surveillance and when to start it. Surveillance should be started at least 1 decade before the earliest age of pancreatic cancer in the family. EUS is the basic, least-invasive surveillance tool; however, findings are similar to those seen in chronic pancreatitis. All patients who have a positive EUS or who have symptoms warrant ERCP. Changes on ERCP of ductal stricturing and clubbed or saccular side branches are suggestive of patients who may need pancreatectomy in the setting of hereditary pancreatic cancer. The goal for surveillance of familial pancreatic cancer patients is to diagnose them before the development of cancer, when they have dysplasia or carcinoma in situ, and to perform a complete pancreatectomy. Timing is crucial for determining when a patient warrants surgery; if performed too early, the patient is put at risk for the morbidity and mortality of a total pancreatectomy, which is not inconsequential. If the patient survives the operation, he or she is often left a brittle diabetic. The alternative of diagnosing too late is more worrisome because the patient dies of pancreatic cancer. An essential ingredient to a good patient outcome is a team approach to these patients, using gastroenterologists, surgeons, and pathologists who have expertise and interest in pancreatic disease. PMID- 10872428 TI - Screening for early pancreatic ductal adenocarcinoma in hereditary pancreatitis. AB - Patients with hereditary pancreatitis have a 40% lifetime risk of developing pancreatic ductal adenocarcinoma. Existing methods of diagnosing pancreatic cancer such as tumor markers, endoscopy, and radiological imaging lack the sensitivity and specificity for early diagnosis, particularly in a background of chronic pancreatitis. Molecular based strategies offer new avenues of screening for pancreatic ductal adenocarcinoma in these high-risk patients, which may allow the development of highly sensitive and specific diagnostic tests for the early detection of cancer. PMID- 10872429 TI - Pancreatic cancer surveillance in a high-risk cohort. Is it worth the cost? AB - Pancreatic adenocarcinoma is the 10th most common malignancy and 4th largest cancer killer in adults. Earlier tumor detection through screening of high risk groups, presumably to increase the percentage of cases resectable for cure in these cohorts, has emerged as a prominent strategy to combat this disease. This article examines the feasibility of this strategy in patients with hereditary pancreatic cancer (HPC) and hereditary pancreatitis (HP). Because of a variety of factors, specific cost projections for screening with HPC kindreds are problematic at best. Patients with HP exhibit a 53-fold increased risk of pancreatic cancer, with a cumulative risk of 40% by age 70. The authors discuss the modalities available to screen this cohort and subsequently perform a theoretical cost analysis. The authors' findings suggest that screening has the potential to be cost-effective only in hereditary pancreatitis patients = 50 years-of-age. The most cost-effective option will likely combine an initial serologic test with high sensitivity and a subsequent serologic or pancreatic juice test with sufficient specificity to act as a "gatekeeper" to imaging with endoscopic ultrasound (EUS). Banking of blood and pancreatic juice samples should be mandatory in any screening protocol. The lower tumor yield in other high-risk groups (e.g., non-hereditary chronic pancreatitis) will effectively preclude the use of such screening protocols. The vast majority of patients will continue to present with unresectable disease. PMID- 10872430 TI - Surgical management of hereditary pancreatic cancer. AB - Pancreatic cancer continues to be a leading cause of cancer death in the United States. Seven genetic syndromes are now known to be associated with an increased incidence of pancreatic cancer. Other familial forms of pancreatic cancer exist although the genetic basis for this predisposition remains elusive. The similarities in the genetic and clinical manifestations of the sporadic and familial forms of pancreatic cancer suggest that pretreatment staging and management of patients with established pancreatic cancer should be similar. For carcinomas of the pancreatic head, pancreaticoduodenectomy should be performed according to current surgical practice, whereas the use of total pancreatectomy should be limited to cases in which margins are found to be positive or if the anatomy precludes a safe pancreaticojejunostomy. Total pancreatectomy may be considered in high-risk kindreds who strongly desire prophylactic surgery and in those with premalignant lesions. Identification of the precise genetic basis for inherited pancreatic cancer will someday make it possible to examine scientifically the effectiveness of specific management strategies. PMID- 10872431 TI - A turbulent decade: lessons from the 'health reforms'. PMID- 10872432 TI - Internet use amongst New Zealand general practitioners. AB - AIMS: To assess how extensively New Zealand doctors are using medical information on the Internet, and to examine how the new technology is affecting their practice of medicine. METHODS: All general practitioners (GPs) known to be working in Otago and Southland were asked to complete a postal questionnaire regarding their use of the Internet and their impressions of patient use of online medical resources. RESULTS: Of 259 questionnaires mailed out, 168 (65%) were returned by GPs currently in practice. Of those, 114 (68%) said they used the Internet at least monthly. A total of 71% of GPs had patients who indicated they had sought medical information from the Internet. Nearly half of respondents expressed concerns that the Internet could have unwelcome effects on the doctor patient relationship. CONCLUSIONS: Internet use among New Zealand doctors and patients is widespread, and is likely to have significant impact on medical practice now and in the future. While the potential benefits of the new technology are numerous, the Internet may become a source of conflict between doctors and patients. PMID- 10872433 TI - A meta-analysis of 25 hydroxyvitamin D in older people with fracture of the proximal femur. AB - AIMS: To perform a meta-analysis on the published studies of serum levels of 25 hydroxyvitamin D in older people with fractures of the proximal femur compared to control groups. METHODS: A 'Medline' literature search using key words vitamin D' and 'hip fractures' for the years January 1966 to June 1999, seeking only papers published in English and available from New Zealand medical libraries, was performed. Bibliographies of identified papers were also searched. Studies which compared 25 hydroxyvitamin D levels in people with a fracture of the proximal femur to an older control group were eligible for inclusion. 30 studies were identified and 28 papers could be found in New Zealand. The method of weighted Z statistics was used in the meta-analysis. RESULTS: The pooled reduction in serum 25 hydroxyvitamin D for the fracture group compared to the controls was 0.66 of a standard deviation with a 95% confidence interval of 0.74 to 0.59. CONCLUSIONS: Although there may be publication bias in this meta-analysis and there was some evidence of heterogeneity in the studies, there is very good evidence that older people with fracture of the proximal femur have reduced levels of vitamin D compared to controls. Older people with fracture of the proximal femur should be treated with vitamin D. PMID- 10872434 TI - Glycaemic index of New Zealand foods. AB - AIM: To determine the glycaemic index values to a range of foods that are unique to New Zealand, and those that are grown and/or manufactured locally. METHODS: We determined the glycaemic index of 28 carbohydrate foods in both healthy subjects and those with type 2 diabetes. Venous blood samples were collected over two to three hours, and the incremental area under the blood glucose curve was used to calculate glycaemic index values. RESULTS AND CONCLUSIONS: This study has identified the glycaemic index values for a range of New Zealand foods which will be clinically useful in the nutritional management of individuals with diabetes mellitus. People with diabetes are recommended to choose foods with a low glycaemic index which is associated with optimal blood glucose control and lipid levels. PMID- 10872435 TI - Willingness to pay for new chemotherapy for advanced ovarian cancer. AB - AIMS: To investigate whether health care professionals had differing attitudes about cost and survival issues in regard to the treatment of advanced ovarian cancer. METHODS: A questionnaire designed for the study was mailed to hospital doctors, hospital nurses, Crown Health Enterprise (CHE) managers and health authority managers who were working in the same geographical area. A total of 391 questionnaires were mailed and 186 were completed reflecting an overall response rate of 47%. Replies were received from 46% of the health authority managers, 61% of the CHE managers, 59% of the doctors and 27% of nurses. RESULTS: Results indicated a threshold for how much individuals were willing to pay. Group differences were evident with the nurses and CHE managers willing to spend less than the other two groups. In addition, those with experience of cancer treatment were willing to spend less than those with no such experience. CONCLUSIONS: There are differences in attitudes to costs and benefits of treatment between individuals working in different areas of health care. Health sector workers value treatments which extend life expectancy without necessarily being curative. PMID- 10872436 TI - Trauma form documentation in major trauma. AB - AIMS: To examine the impact of a standardised trauma form for documentation in cases of major trauma, a prospective study was undertaken. METHODS: Records written by medical staff pertaining to the assessment and treatment of major trauma patients in the resuscitation room were scored against a panel of parameters derived from advanced trauma life support guidelines. Demographics, aetiology, trauma scores and outcome data were obtained from a trauma registry. Attitudes of medical staff involved in major trauma to the trauma form were assessed using a questionnaire. RESULTS: The trauma form was used in 53 of 69 (76.8%) consecutive cases of major trauma seen over a three month period. No significant differences existed in demographics, aetiology, trauma scores or outcome between form and formless groups. In the form group, a median of 44 of 51 (86.3%) relevant information parameters were documented versus 32 of 51 (62.7%) in the formless group, p < 0.0001. A positive approach to the trauma form was indicated by the questionnaire results. CONCLUSION: The use of a standardised form improves documentation in major trauma. PMID- 10872437 TI - Right idea, wrong model. Why general practitioners are not succeeding with preventive care. PMID- 10872438 TI - Research in general practice or general practice research. PMID- 10872439 TI - What isn't medical practice? PMID- 10872441 TI - Excessive or inappropriate claims and treatment. PMID- 10872440 TI - Gallstones in males. PMID- 10872442 TI - It all started on a streetcar in Boston. AB - We first met on a Boston streetcar in 1940, being introduced by a mutual friend. Celia was returning from research work at the Massachusetts General Hospital as part of her senior thesis at Radcliffe College, and Herb was returning from a concert by the Boston Symphony. We were married in 1946 after Celia had finished her medical training. We started working together in 1952, and we are still actively collaborating in our studies on various aspects of the biosynthesis and function of polyamines. We are honored to have been invited by the editors of the Annual Review of Biochemistry to summarize our activities in biochemical research over the past 60 years. During most of this time we have been at the National Institutes of Health in Bethesda, Md., and we have witnessed the enormous expansion of biomedical research that has occurred during this period. In addition to summarizing our research, Herb summarizes his association with the Journal of Biological Chemistry and the remarkable developments that have occurred recently in electronic publication and dissemination of scientific literature. PMID- 10872443 TI - Catalysis by metal-activated hydroxide in zinc and manganese metalloenzymes. AB - The metal-activated hydroxide ion is a critical nucleophile in metalloenzymes that catalyze hydrolysis or hydration reactions. The most common metal used is zinc; occasionally, other transition metals such as manganese are required. Human carbonic anhydrase II and rat liver arginase serve as well-studied paradigms of zinc and manganese metalloenzymes, respectively. Comparative structure-function relationships between these two metalloenzymes highlight parallels in the chemistry of metal-activated hydroxide: (a) the protein environment of metal bound hydroxide modulates its reactivity; (b) a hydrogen bond with metal-bound hydroxide holds it in the proper orientation for catalysis; (c) nonmetal substrate-binding sites are implicated in both enzyme mechanisms; and (d) regeneration of metal-bound hydroxide ion from a metal-bound water molecule requires proton transfer to bulk solvent mediated by a histidine proton shuttle residue. Interestingly, the electrostatics of catalysis differ between the two enzymes, in that the first step of catalysis requires formation of a negatively charged transition state in the carbonic anhydrase II mechanism, whereas a neutral transition state is approached in the arginase mechanism. This electrostatic feature may contribute to the differences in the chemistry, the metal binding sites, and the metal specificity between these two enzymes. PMID- 10872444 TI - Conus peptides targeted to specific nicotinic acetylcholine receptor subtypes. AB - The venoms of predatory cone snails represent a rich combinatorial-like library of evolutionarily selected, neuropharmacologically active peptides. A major fraction of the venom components are conotoxins--small, disulfide-rich peptides that potently and specifically target components of the neuromuscular system, particularly ligand- and voltage-gated ion channels. This review focuses on Conus peptides, which act at nicotinic acetylcholine receptors. These nicotinic antagonist peptides from Conus are broadly divided into two groups: those that act at the neuromuscular junction and those that act at subtypes of neuronal nicotinic acetylcholine receptors. The latter include peptides specific for the alpha 7, alpha 3 beta 2, and alpha 3 beta 4 nicotinic receptor subtypes. The degree of specificity exhibited by these peptides is remarkable, particularly given their relatively small size. As a group the nicotinic acetylcholine receptor-targeted Conus peptides represent an increasingly well-defined set of tools for probing the structure, function, and physiological role of nicotinic acetylcholine receptors. PMID- 10872445 TI - Inorganic polyphosphate: a molecule of many functions. AB - Inorganic polyphosphate (poly P) is a chain of tens or many hundreds of phosphate (Pi) residues linked by high-energy phosphoanhydride bonds. Despite inorganic polyphosphate's ubiquity--found in every cell in nature and likely conserved from prebiotic times--this polymer has been given scant attention. Among the reasons for this neglect of poly P have been the lack of sensitive, definitive, and facile analytical methods to assess its concentration in biological sources and the consequent lack of demonstrably important physiological functions. This review focuses on recent advances made possible by the introduction of novel, enzymatically based assays. The isolation and ready availability of Escherichia coli polyphosphate kinase (PPK) that can convert poly P and ADP to ATP and of a yeast exopolyphosphatase that can hydrolyze poly P to Pi, provide highly specific, sensitive, and facile assays adaptable to a high-throughput format. Beyond the reagents afforded by the use of these enzymes, their genes, when identified, mutated, and overexpressed, have offered insights into the physiological functions of poly P. Most notably, studies in E. coli reveal large accumulations of poly P in cellular responses to deficiencies in an amino acid, Pi, or nitrogen or to the stresses of a nutrient downshift or high salt. The ppk mutant, lacking PPK and thus severely deficient in poly P, also fails to express RpoS (a sigma factor for RNA polymerase), the regulatory protein that governs > or = 50 genes responsible for stationary-phase adaptations to resist starvation, heat and oxidant stresses, UV irradiation, etc. Most dramatically, ppk mutants die after only a few days in stationary phase. The high degree of homology of the PPK sequence in many bacteria, including some of the major pathogenic species (e.g. Mycobacterium tuberculosis, Neisseria meningitidis, Helicobacter pylori, Vibrio cholerae, Salmonella typhimurium, Shigella flexneri, Pseudomonas aeruginosa, Bordetella pertussis, and Yersinia pestis), has prompted the knockout of their ppk gene to determine the dependence of virulence on poly P and the potential of PPK as a target for antimicrobial drugs. In yeast and mammalian cells, exo- and endopolyphosphatases have been identified and isolated, but little is known about the synthesis of poly P or its physiologic functions. Whether microbe or human, all species depend on adaptations in the stationary phase, which is truly a dynamic phase of life. Most research is focused on the early and reproductive phases of organisms, which are rather brief intervals of rapid growth. More attention needs to be given to the extensive period of maturity. Survival of microbial species depends on being able to manage in the stationary phase. In view of the universality and complexity of basic biochemical mechanisms, it would be surprising if some of the variety of poly P functions observed in microorganisms did not apply to aspects of human growth and development, to aging, and to the aberrations of disease. Of theoretical interest regarding poly P is its antiquity in prebiotic evolution, which along with its high energy and phosphate content, make it a plausible precursor to RNA, DNA, and proteins. Practical interest in poly P includes many industrial applications, among which is the microbial removal of Pi in aquatic environments. PMID- 10872446 TI - The molecular basis of hypertension. AB - More than 50 million Americans display blood pressures outside the safe physiological range. Unfortunately for most individuals, the molecular basis of hypertension is unknown, in part because pathological elevations of blood pressure are the result of abnormal expression of multiple genes. This review identifies a number of important blood pressure regulatory genes including their loci in the human, mouse, and rat genome. Phenotypes of gene deletions and overexpression in mice are summarized. More detailed discussion of selected gene products follows, beginning with proteins involved in ion transport, specifically the epithelial sodium channel and sodium proton exchangers. Next, proteins involved in vasodilation/natriuresis are discussed with emphasis on natriuretic peptides, guanylin/uroguanylin, and nitric oxide. The renin angiotensin aldosterone system has an important role antagonizing the vasodilatory cyclic GMP system. PMID- 10872447 TI - Sterols and isoprenoids: signaling molecules derived from the cholesterol biosynthetic pathway. AB - Compounds derived from the isoprenoid/cholesterol biosynthetic pathway have recently been shown to have novel biological activities. These compounds include certain sterols, oxysterols, farnesol, and geranylgeraniol, as well as the diphosphate derivatives of isopentenyl, geranyl, farnesyl, geranylgeranyl, and presqualene. They regulate transcriptional and post-transcriptional events that in turn affect lipid synthesis, meiosis, apoptosis, developmental patterning, protein cleavage, and protein degradation. PMID- 10872448 TI - Ciliate telomerase biochemistry. AB - Telomerase is a cellular reverse transcriptase specialized for use of a template carried within the RNA component of the enzyme ribonucleoprotein complex. Substrates for telomerase are single-stranded oligonucleotides in vitro and chromosome ends in vivo. In vitro, a bound substrate is extended by an initial round of DNA synthesis on the internal RNA template and in some cases by multiple rounds of template copying before product dissociation. In vivo, de novo synthesis of one strand of a telomeric repeat sequence by telomerase balances the sequence loss resulting from incomplete replication of linear chromosome ends by RNA primer-requiring DNA polymerases. Telomerase biochemistry has been studied extensively by using partially purified cell extracts. Telomerase components are being identified and beginning to be produced in recombinant form. This review focuses on the enzyme mechanism of telomerases from ciliate species, thus far the most intensively studied systems. PMID- 10872449 TI - Tolerance and specificity of polyketide synthases. AB - Polyketide synthases catalyze the assembly of complex natural products from simple precursors such as propionyl-CoA and methylmalonyl-CoA in a biosynthetic process that closely parallels fatty acid biosynthesis. Like fatty acids, polyketides are assembled by successive decarboxylative condensations of simple precursors. But whereas the intermediates in fatty acid biosynthesis are fully reduced to generate unfunctionalized alkyl chains, the intermediates in polyketide biosynthesis may be only partially processed, giving rise to complex patterns of functional groups. Additional complexity arises from the use of different starter and chain extension substrates, the generation of chiral centers, and further functional group modifications, such as cyclizations. The structural and functional modularity of these multienzyme systems has raised the possibility that polyketide biosynthetic pathways might be rationally reprogrammed by combinatorial manipulation. An essential prerequisite for harnessing this biosynthetic potential is a better understanding of the molecular recognition features of polyketide synthases. Within this decade, a variety of genetic, biochemical, and chemical investigations have yielded insights into the tolerance and specificity of several architecturally different polyketide synthases. The results of these studies, together with their implications for biosynthetic engineering, are summarized in this review. PMID- 10872450 TI - Initiation of base excision repair: glycosylase mechanisms and structures. AB - The base excision repair pathway is an organism's primary defense against mutations induced by oxidative, alkylating, and other DNA-damaging agents. This pathway is initiated by DNA glycosylases that excise the damaged base by cleavage of the glycosidic bond between the base and the DNA sugar-phosphate backbone. A subset of glycosylases has an associated apurinic/apyrimidinic (AP) lyase activity that further processes the AP site to generate cleavage of the DNA phosphate backbone. Chemical mechanisms that are supported by biochemical and structural data have been proposed for several glycosylases and glycosylase/AP lyases. This review focuses on the chemical mechanisms of catalysis in the context of recent structural information, with emphasis on the catalytic residues and the active site conformations of several cocrystal structures of glycosylases with their substrate DNAs. Common structural motifs for DNA binding and damage specificity as well as conservation of acidic residues and amino groups for catalysis are discussed. PMID- 10872451 TI - Structural motifs in RNA. AB - An RNA motif is a discrete sequence or combination of base juxtapositions found in naturally occurring RNAs in unexpectedly high abundance. Because all the motifs examined so far have three-dimensional structures independent of the context in which they are embedded, they are important components of the "kit" of structural elements from which RNAs are constructed. This review discusses the structures of the motifs that have been identified so far and speculates on the importance of their role in determining RNA conformation and their evolutionary origin. PMID- 10872452 TI - Transcription elongation and human disease. AB - Eukaryotic mRNA synthesis is catalyzed by multisubunit RNA polymerase II and proceeds through multiple stages referred to as preinitiation, initiation, elongation, and termination. Over the past 20 years, biochemical studies of eukaryotic mRNA synthesis have largely focused on the preinitiation and initiation stages of transcription. These studies led to the discovery of the class of general initiation factors (TFIIB, TFIID, TFIIE, TFIIF, and TFIIH), which function in intimate association with RNA polymerase II and are required for selective binding of polymerase to its promoters, formation of the open complex, and synthesis of the first few phosphodiester bonds of nascent transcripts. Recently, biochemical studies of the elongation stage of eukaryotic mRNA synthesis have led to the discovery of several cellular proteins that have properties expected of general elongation factors and that have been found to play unanticipated roles in human disease. Among these candidate general elongation factors are the positive transcription elongation factor b (P-TEFb), eleven-nineteen lysine-rich in leukemia (ELL), Cockayne syndrome complementation group B (CSB), and elongin proteins, which all function in vitro to expedite elongation by RNA polymerase II by suppressing transient pausing or premature arrest by polymerase through direct interactions with the elongation complex. Despite their similar activities in elongation, the P-TEFb, ELL, CSB, and elongin proteins appear to play roles in a diverse collection of human diseases, including human immunodeficiency virus-1 infection, acute myeloid leukemia, Cockayne syndrome, and the familial cancer predisposition syndrome von Hippel Lindau disease. here we review our current understanding of the P-TEFb, ELL, CSB, and elongin proteins, their mechanisms of action, and their roles in human disease. PMID- 10872453 TI - Control of carpel and fruit development in Arabidopsis. AB - The fruit is a highly specialized plant organ that occurs in diverse forms among the angiosperms. Fruits of Arabidopsis thaliana, which are typical of the > 3000 species of Brassicaceae, develop from a gynoecium that consists of two fused carpels. The mature gynoecium of Arabidopsis is composed of an apical stigma, a short style, and a basal ovary that contains the developing ovules. After the ovules are fertilized, the fruit elongates and differentiates a number of distinct cell types, allowing for the successful maturation and the eventual dispersal of the seeds. Although the processes involved in carpel and fruit morphogenesis are not well understood, recent studies have identified a large number of mutants that display abnormal gynoecium and fruit development. The detailed phenotypic description of these mutants together with recent cloning of many of these genes has begun to shed light on this interesting and complex developmental process. Here we review the growing collection of Arabidopsis genes known to control the initiation and development of the gynoecium and resulting fruit. PMID- 10872454 TI - Tetrahydropterin-dependent amino acid hydroxylases. AB - Phenylalanine hydroxylase, tyrosine hydroxylase, and tryptophan hydroxylase constitute a small family of monooxygenases that utilize tetrahydropterins as substrates. When from eukaryotic sources, these enzymes are composed of a homologous catalytic domain to which are attached discrete N-terminal regulatory domains and short C-terminal tetramerization domains, whereas the bacterial enzymes lack the N-terminal and C-terminal domains. Each enzyme contains a single ferrous iron atom bound to two histidines and a glutamate. Recent mechanistic studies have begun to provide insights into the mechanisms of oxygen activation and hydroxylation. Although the hydroxylating intermediate in these enzymes has not been identified, the iron is likely to be involved. Reversible phosphorylation of serine residues in the regulatory domains affects the activities of all three enzymes. In addition, phenylalanine hydroxylase is allosterically regulated by its substrates, phenylalanine and tetrahydrobiopterin. PMID- 10872455 TI - Mammalian caspases: structure, activation, substrates, and functions during apoptosis. AB - Apoptosis is a genetically programmed, morphologically distinct form of cell death that can be triggered by a variety of physiological and pathological stimuli. Studies performed over the past 10 years have demonstrated that proteases play critical roles in initiation and execution of this process. The caspases, a family of cysteine-dependent aspartate-directed proteases, are prominent among the death proteases. Caspases are synthesized as relatively inactive zymogens that become activated by scaffold-mediated transactivation or by cleavage via upstream proteases in an intracellular cascade. Regulation of caspase activation and activity occurs at several different levels: (a) Zymogen gene transcription is regulated; (b) antiapoptotic members of the Bcl-2 family and other cellular polypeptides block proximity-induced activation of certain procaspases; and (c) certain cellular inhibitor of apoptosis proteins (cIAPs) can bind to and inhibit active caspases. Once activated, caspases cleave a variety of intracellular polypeptides, including major structural elements of the cytoplasm and nucleus, components of the DNA repair machinery, and a number of protein kinases. Collectively, these scissions disrupt survival pathways and disassemble important architectural components of the cell, contributing to the stereotypic morphological and biochemical changes that characterize apoptotic cell death. PMID- 10872456 TI - Cellular and molecular biology of the aquaporin water channels. AB - The high water permeability characteristic of mammalian red cell membranes is now known to be caused by the protein AQP1. This channel freely permits movement of water across the cell membrane, but it is not permeated by other small, uncharged molecules or charged solutes. AQP1 is a tetramer with each subunit containing an aqueous pore likened to an hourglass formed by obversely arranged tandem repeats. Cryoelectron microscopy of reconstituted AQP1 membrane crystals has revealed the three-dimensional structure at 3-6 A. AQP1 is distributed in apical and basolateral membranes of renal proximal tubules and descending thin limbs as well as capillary endothelia. Ten mammalian aquaporins have been identified in water permeable tissues and fall into two groupings. Orthodox aquaporins are water selective and include AQP2, a vasopressin-regulated water channel in renal collecting duct, in addition to AQP0, AQP4, and AQP5. Multifunctional aquaglyceroporins AQP3, AQP7, and AQP9 are permeated by water, glycerol, and some other solutes. Aquaporins are being defined in numerous other species including amphibia, insects, plants, and microbials. Members of the aquaporin family are implicated in numerous physiological processes as well as the pathophysiology of a wide range of clinical disorders. PMID- 10872457 TI - Regulation of the cytoskeleton and cell adhesion by the Rho family GTPases in mammalian cells. AB - Members of the Rho family of small Ras-like GTPases--including RhoA, -B, and -C, Rac1 and -2, and Cdc42--exhibit guanine nucleotide-binding activity and function as molecular switches, cycling between an inactive GDP-bound state and an active GTP-bound state. The Rho family GTPases participate in regulation of the actin cytoskeleton and cell adhesion through specific targets. Identification and characterization of these targets have begun to clarify how the Rho family GTPases act to regulate cytoskeletal structure and cell-cell and cell-substratum contacts in mammalian cells. The Rho family GTPases are also involved in regulation of smooth muscle contraction, cell morphology, cell motility, neurite retraction, and cytokinesis. However, the molecular mechanisms by which the Rho family GTPases participate in the regulation of such processes are not well established. PMID- 10872458 TI - Mutagenesis of glycosidases. AB - Enzymatic hydrolysis of glycosides can occur by one of two elementary mechanisms identified by the stereochemical outcome of the reaction, inversion or retention. The key active-site residues involved are a pair of carboxylic acids in each case, and strategies for their identification and for probing the details of their roles in catalysis have been developed through detailed kinetic analysis of mutants. Similarly the roles of other active-site residues have also been probed this way, and mutants have been developed that trap intermediates in catalysis, allowing the determination of the three-dimensional structures of several such key species. By manipulating the locations or even the presence of these carboxyl side chains in the active site, the mechanisms of several glycosidases have been completely changed, and this has allowed the development of "glycosynthases," mutant glycosidases that are capable of synthesizing oligosaccharides but unable to degrade them. Surprisingly little progress has been made on altering specificities through mutagenesis, although recent results suggest that gene shuffling coupled with effective screens will provide the most effective approach. PMID- 10872459 TI - Charting the fate of the "good cholesterol": identification and characterization of the high-density lipoprotein receptor SR-BI. AB - Risk for cardiovascular disease due to atherosclerosis increases with increasing concentrations of low-density lipoprotein (LDL) cholesterol and is inversely proportional to the levels of high-density lipoprotein (HDL) cholesterol. The receptor-mediated control of plasma LDL levels has been well understood for over two decades and has been a focus for the pharmacologic treatment of hypercholesterolemia. In contrast, the first identification and characterization of a receptor that mediates cellular metabolism of HDL was only recently reported. This receptor, called scavenger receptor class B type I (SR-BI), is a fatty acylated glycoprotein that can cluster in caveolae-like domains on the surfaces of cultured cells. SR-BI mediates selective lipid uptake from HDL to cells. The mechanism of selective lipid uptake is fundamentally different from that of classic receptor-mediated endocytic uptake via coated pits and vesicles (e.g. the LDL receptor pathway) in that it involves efficient receptor-mediated transfer of the lipids, but not the outer shell proteins, from HDL to cells. In mice, SR-BI plays a key role in determining the levels of plasma HDL cholesterol and in mediating the regulated, selective delivery of HDL-cholesterol to steroidogenic tissues and the liver. Significant alterations in SR-BI expression can result in cardiovascular and reproductive disorders. SR-BI may play a similar role in humans; thus, modulation of its activity may provide the basis of future approaches to the treatment and prevention of atherosclerotic disease. PMID- 10872460 TI - Nuclear-receptor ligands and ligand-binding domains. AB - The determination of several structures of nuclear receptor ligand binding domains (LBD) has led to new insights into the mechanism of action of this very important class of receptors. This review describes and compares the different LBD structures and their relationship to the function of the nuclear receptors. The role of the ligand in the LBD structures and the implications of ligand structure on receptor activity are also discussed. Structural information regarding interactions between the LBD and coactivator proteins and the potential role of these interactions in ligand agonism and antagonism is reviewed. Different pathways for nuclear receptor signaling and the use of new ligands to investigate these pathways are also described. PMID- 10872461 TI - The anaphase-promoting complex: new subunits and regulators. AB - Ubiquitin-mediated proteolysis of cell cycle regulators is a crucial process during the cell cycle. The anaphase-promoting complex (APC) is a large, multiprotein complex whose E3-ubiquitin ligase activity is required for the ubiquitination of mitotic cyclins and other regulatory proteins that are targeted for destruction during cell division. The recent identification of new APC subunits and regulatory proteins has begun to reveal some of the intricate mechanisms that govern APC regulation. One mechanism is the use of specificity factors to impose temporal control over substrate degradation. A second mechanism is the APC-mediated proteolysis of specific APC regulators. Finally, components of both the APC and the SCF E3 ubiquitin-ligase complex contain several conserved sequence motifs, including WD-40 repeats and cullin homology domains, which suggest that both complexes may use a similar mechanism for substrate ubiquitination. PMID- 10872462 TI - In vitro selection of functional nucleic acids. AB - In vitro selection allows rare functional RNA or DNA molecules to be isolated from pools of over 10(15) different sequences. This approach has been used to identify RNA and DNA ligands for numerous small molecules, and recent three dimensional structure solutions have revealed the basis for ligand recognition in several cases. By selecting high-affinity and -specificity nucleic acid ligands for proteins, promising new therapeutic and diagnostic reagents have been identified. Selection experiments have also been carried out to identify ribozymes that catalyze a variety of chemical transformations, including RNA cleavage, ligation, and synthesis, as well as alkylation and acyl-transfer reactions and N-glycosidic and peptide bond formation. The existence of such RNA enzymes supports the notion that ribozymes could have directed a primitive metabolism before the evolution of protein synthesis. New in vitro protein selection techniques should allow for a direct comparison of the frequency of ligand binding and catalytic structures in pools of random sequence polynucleotides versus polypeptides. PMID- 10872463 TI - MCM proteins in DNA replication. AB - The MCM proteins are essential replication initiation factors originally identified as proteins required for minichromosome maintenance in Saccharomyces cerevisiae. The best known among them are a family of six structurally related proteins, MCM2-7, which are evolutionally conserved in all eukaryotes. The MCM2-7 proteins form a hexameric complex. This complex is a key component of the prereplication complex that assembles at replication origins during early G1 phase. New evidence suggests that the MCM2-7 proteins may be involved not only in the initiation but also in the elongation of DNA replication. Orchestration of the functional interactions between the MCM2-7 proteins and other components of the prereplication complex by cell cycle-dependent protein kinases results in initiation of DNA synthesis once every cell cycle. PMID- 10872464 TI - Structural mechanism of muscle contraction. AB - X-ray crystallography shows the myosin cross-bridge to exist in two conformations, the beginning and end of the "power stroke." A long lever-arm undergoes a 60 degrees to 70 degrees rotation between the two states. This rotation is coupled with changes in the active site (OPEN to CLOSED) and phosphate release. Actin binding mediates the transition from CLOSED to OPEN. Kinetics shows that the binding of myosin to actin is a two-step process which affects ATP and ADP affinity. The structural basis of these effects is not explained by the presently known conformers of myosin. Therefore, other states of the myosin cross-bridge must exist. Moreover, cryoelectronmicroscopy has revealed other angles of the cross-bridge lever arm induced by ADP binding. These structural states are presently being characterized by site-directed mutagenesis coupled with kinetic analysis. PMID- 10872465 TI - Functions of cell surface heparan sulfate proteoglycans. AB - The heparan sulfate on the surface of all adherent cells modulates the actions of a large number of extracellular ligands. Members of both cell surface heparan sulfate proteoglycan families, the transmembrane syndecans and the glycosylphosphoinositide-linked glypicans, bind these ligands and enhance formation of their receptor-signaling complexes. These heparan sulfate proteoglycans also immobilize and regulate the turnover of ligands that act at the cell surface. The extracellular domains of these proteoglycans can be shed from the cell surface, generating soluble heparan sulfate proteoglycans that can inhibit interactions at the cell surface. Recent analyses of genetic defects in Drosophila melanogaster, mice, and humans confirm most of these activities in vivo and identify additional processes that involve cell surface heparan sulfate proteoglycans. This chapter focuses on the mechanisms underlying these activities and on the cellular functions that they regulate. PMID- 10872466 TI - De novo design and structural characterization of proteins and metalloproteins. AB - De novo protein design has recently emerged as an attractive approach for studying the structure and function of proteins. This approach critically tests our understanding of the principles of protein folding; only in de novo design must one truly confront the issue of how to specify a protein's fold and function. If we truly understand proteins, it should be possible to design receptors, enzymes, and ion channels from scratch. Further, as this understanding evolves and is further refined, it should be possible to design proteins and biomimetic polymers with properties unprecedented in nature. PMID- 10872467 TI - CREB: a stimulus-induced transcription factor activated by a diverse array of extracellular signals. AB - Extracellular stimuli elicit changes in gene expression in target cells by activating intracellular protein kinase cascades that phosphorylate transcription factors within the nucleus. One of the best characterized stimulus-induced transcription factors, cyclic AMP response element (CRE)-binding protein (CREB), activates transcription of target genes in response to a diverse array of stimuli, including peptide hormones, growth factors, and neuronal activity, that activate a variety of protein kinases including protein kinase A (PKA), pp90 ribosomal S6 kinase (pp90RSK), and Ca2+/calmodulin-dependent protein kinases (CaMKs)[corrected]. These kinases all phosphorylate CREB at a particular residue, serine 133 (Ser133), and phosphorylation of Ser133 is required for CREB-mediated transcription. Despite this common feature, the mechanism by which CREB activates transcription varies depending on the stimulus. In some cases, signaling pathways target additional sites on CREB or proteins associated with CREB, permitting CREB to regulate distinct programs of gene expression under different conditions of stimulation. This review discusses the molecular mechanisms by which Ser133 phosphorylated CREB activates transcription, intracellular signaling pathways that lead to phosphorylation of CREB at Ser133, and features of each signaling pathway that impart specificity at the level of CREB activation. PMID- 10872468 TI - Membrane fusion and exocytosis. AB - Membrane fusion involves the merger of two phospholipid bilayers in an aqueous environment. In artificial lipid bilayers, fusion proceeds by means of defined transition states, including hourglass-shaped intermediates in which the proximal leaflets of the fusing membranes are merged whereas the distal leaflets are separate (fusion stalk), followed by the reversible opening of small aqueous fusion pores. Fusion of biological membranes requires the action of specific fusion proteins. Best understood are the viral fusion proteins that are responsible for merging the viral with the host cell membrane during infection. These proteins undergo spontaneous and dramatic conformational changes upon activation. In the case of the paradigmatic fusion proteins of the influenza virus and of the human immunodeficiency virus, an amphiphilic fusion peptide is inserted into the target membrane. The protein then reorients itself, thus forcing the fusing membranes together and inducing lipid mixing. Fusion of intracellular membranes in eukaryotic cells involves several protein families including SNAREs, Rab proteins, and Sec1/Munc-18 related proteins (SM-proteins). SNAREs form a novel superfamily of small and mostly membrane-anchored proteins that share a common motif of about 60 amino acids (SNARE motif). SNAREs reversibly assemble into tightly packed helical bundles, the core complexes. Assembly is thought to pull the fusing membranes closely together, thus inducing fusion. SM-proteins comprise a family of soluble proteins that bind to certain types of SNAREs and prevent the formation of core complexes. Rab proteins are GTPases that undergo highly regulated GTP-GDP cycles. In their GTP form, they interact with specific proteins, the effector proteins. Recent evidence suggests that Rab proteins function in the initial membrane contact connecting the fusing membranes but are not involved in the fusion reaction itself. PMID- 10872469 TI - eIF4 initiation factors: effectors of mRNA recruitment to ribosomes and regulators of translation. AB - Eukaryotic translation initiation factor 4F (eIF4F) is a protein complex that mediates recruitment of ribosomes to mRNA. This event is the rate-limiting step for translation under most circumstances and a primary target for translational control. Functions of the constituent proteins of eIF4F include recognition of the mRNA 5' cap structure (eIF4E), delivery of an RNA helicase to the 5' region (eIF4A), bridging of the mRNA and the ribosome (eIF4G), and circularization of the mRNA via interaction with poly(A)-binding protein (eIF4G). eIF4 activity is regulated by transcription, phosphorylation, inhibitory proteins, and proteolytic cleavage. Extracellular stimuli evoke changes in phosphorylation that influence eIF4F activity, especially through the phosphoinositide 3-kinase (PI3K) and Ras signaling pathways. Viral infection and cellular stresses also affect eIF4F function. The recent determination of the structure of eIF4E at atomic resolution has provided insight about how translation is initiated and regulated. Evidence suggests that eIF4F is also implicated in malignancy and apoptosis. PMID- 10872470 TI - AKT/PKB and other D3 phosphoinositide-regulated kinases: kinase activation by phosphoinositide-dependent phosphorylation. AB - The protein kinase Akt/PKB is activated via a multistep process by a variety of signals. In the early steps of this process, PI-3 kinase-generated D3 phosphorylated phosphoinositides bind the Akt PH domain and induce the translocation of the kinase to the plasma membrane where it co-localizes with phosphoinositide-dependent kinase-1. By binding to the PH domains of both Akt and phosphoinositide-dependent kinase-1, D3-phosphorylated phosphoinositides appear to also induce conformational changes that permit phosphoinositide-dependent kinase-1 to phosphorylate the activation loop of Akt. The paradigm of Akt activation via phosphoinositide-dependent phosphorylation provided a framework for research into the mechanism of activation of other members of the AGC kinase group (p70S6K, PKC, and PKA) and members of the Tec tyrosine kinase family (TecI, TecII, Btk/Atk, Itk/Tsk/Emt, Txk/Rlk, and Bm/Etk). The result was the discovery that these kinases and Akt are activated by overlapping pathways. In this review, we present our current understanding of the regulation and function of the Akt kinase and we discuss the common and unique features of the activation processes of Akt and the AGC and Tec kinase families. In addition, we present an overview of the biosynthesis of phosphoinositides that contribute to the regulation of these kinases. PMID- 10872471 TI - The 26S proteasome: a molecular machine designed for controlled proteolysis. AB - In eukaryotic cells, most proteins in the cytosol and nucleus are degraded via the ubiquitin-proteasome pathway. The 26S proteasome is a 2.5-MDa molecular machine built from approximately 31 different subunits, which catalyzes protein degradation. It contains a barrel-shaped proteolytic core complex (the 20S proteasome), capped at one or both ends by 19S regulatory complexes, which recognize ubiquitinated proteins. The regulatory complexes are also implicated in unfolding and translocation of ubiquitinated targets into the interior of the 20S complex, where they are degraded to oligopeptides. Structure, assembly and enzymatic mechanism of the 20S complex have been elucidated, but the functional organization of the 19S complex is less well understood. Most subunits of the 19S complex have been identified, however, specific functions have been assigned to only a few. A low-resolution structure of the 26S proteasome has been obtained by electron microscopy, but the precise arrangement of subunits in the 19S complex is unclear. PMID- 10872472 TI - The population genetics of the haemoglobinopathies. AB - The haemoglobinopathies are the commonest single-gene disorders known, almost certainly because of the protection they provide against malaria, as attested by a number of observations. The geographical distributions of malaria and haemoglobinopathies largely overlap, and microepidemiological surveys confirm the close relationship between them. For two of the commonest disorders, haemoglobin S and alpha(+)-thalassaemia, there is also good clinical evidence for protection against malaria morbidity. However, not all the evidence appears to support this view. In some parts of the world malaria and haemoglobinopathies are not, and never have been, coexistent. It is also difficult to explain why the majority of haemoglobinopathies appear to be recent mutations and are regionally specific. Here we argue that these apparent inconsistencies in the malaria hypothesis are the result of processes such as genetic drift and migration and of demographic changes that have occurred during the past 10,000 years. When these factors are taken into account, selection by malaria remains the force responsible for the prevalence of the haemoglobinopathies. PMID- 10872473 TI - Alpha-thalassaemia. AB - alpha-Thalassaemias are genetic defects extremely frequent in some populations and are characterized by the decrease or complete suppression of alpha-globin polypeptide chains. The gene cluster, which codes for and controls the production of these polypeptides, maps near the telomere of the short arm of chromosome 16, within a G + C rich and early-replicating DNA region. The genes expressed during the embryonic (zeta) or fetal and adult stage (alpha 2 and alpha 1) can be modified by point mutations which affect either the processing-translation of mRNA or make the polypeptide chains extremely unstable. Much more frequent are the deletions of variable size (from approximately 3 to more than 100 kb) which remove one or both alpha genes in cis or even the whole gene cluster. Deletions of a single gene are the result of unequal pairing during meiosis, followed by reciprocal recombination. These unequal cross-overs, which produce also alpha gene triplications and quadruplications, are made possible by the high degree of homology of the two alpha genes and of their flanking sequences. Other deletions involving one or more genes are due to recombinations which have taken place within non-homologous regions (illegitimate recombinations) or in DNA segments whose homology is limited to very short sequences. Particularly interesting are the deletions which eliminate large DNA areas 5' of zeta or of both alpha genes. These deletions do not include the structural genes but, nevertheless, suppress completely their expression. Larger deletions involving the tip of the short arm of chromosome 16 by truncation, interstitial deletions or translocations result in the contiguous gene syndrome ATR-16. In this complex syndrome alpha thalassaemia is accompanied by mental retardation and variable dismorphic features. The study of mutations of the 5' upstream flanking region has led to the discovery of a DNA sequence, localized 40 kb upstream of the zeta-globin gene, which controls the expression of the alpha genes (alpha major regulatory element or HS-40). In the acquired variant of haemoglobin H (HbH) disease found in rare individuals with myelodysplastic disorders and in the X-linked mental retardation associated with alpha-thalassaemia, a profound reduction or absence of alpha gene expression has been observed, which is not accompanied by structural alterations of the coding or controlling regions of the alpha gene complex. Most probably the acquired alpha-thalassaemia is due to the lack of soluble activators (or presence of repressors) which act in trans and affect the expression of the homologous clusters and are coded by genes not (closely) linked to the alpha genes. The ATR-X syndrome results from mutations of the XH2 gene, located on the X chromosome (Xq13.3) and coding for a transacting factor which regulates gene expression. The interaction of the different alpha-thalassaemia determinants results in three phenotypes: the alpha-thalassaemic trait, clinically silent and presenting only limited alterations of haematological parameters, HbH disease, characterized by the development of a haemolytic anaemia of variable degree, and the (lethal) Hb Bart's hydrops fetalis syndrome. The diagnosis of alpha-thalassaemia due to deletions is implemented by the electrophoretic analysis of genomic DNA digested with restriction enzymes and hybridized with specific molecular probes. Recently polymerase chain reaction (PCR) based strategies have replaced the Southern blotting methodology. The straightforward identification of point mutations is carried out by the specific amplification of the alpha 2 or alpha 1 gene by PCR followed by the localization and identification of the mutation with a variety of screening systems (denaturing gradient gel electrophoresis (DGGE), single strand conformation polymorphisms (SSCP)) and direct sequencing. PMID- 10872474 TI - Beta-thalassaemia. AB - A complete spectrum of genetic lesions affecting the beta-globin gene giving rise to a complete spectrum of phenotypic severity is described. Although most of the molecular lesions involve the structural beta gene directly, some down regulate the gene through in-cis effects at a distance while trans-acting factors are implicated in a few cases. The remarkable phenotypic diversity can be related ultimately to the degree of alpha-globin-beta-globin chain imbalance and arises from variability of mutations affecting the beta gene itself and from interactions with other genetic loci, such as the alpha- and gamma-globin genes. The presence of other interacting loci is implicated by their interactions in increasing gamma gene expression or by an increased proteolytic capacity of the erythroid precursors. It is hoped that observations from the genotype-phenotype relationship might form the basis for a comprehensive diagnostic database that will be useful not only for genetic counselling and prenatal diagnosis but also for providing prognostic information for decision making in bone marrow transplantation and gene therapy programmes in the future. However, it is clear from recent analyses that, apart from the two categories of triplicated alpha genes with heterozygous beta-thalassaemia and inheritance of mild beta(+) thalassaemia alleles, it is still not possible to predict consistently phenotype from alpha and beta genotypes alone owing to the influence of the other modulating factors, some implicated (such as inheritance of hereditary persistence of fetal haemoglobin) and others as yet unidentified. PMID- 10872475 TI - Pathophysiology of thalassaemia. AB - Most of the major clinical manifestations of the beta-thalassaemias can be related to the deleterious effects of imbalanced globin chain synthesis on erythroid maturation and red cell survival. The destruction of red cell progenitors and their progeny results from an extremely complex series of mechanisms all related to the presence of excess alpha-globin chain production. These include mechanical damage, interference with cell division and oxidative destruction of both organelles and components of the red cell membrane. The unequal distribution of gamma-globin chains between different precursors, and the intense selection of those with relatively higher levels of gamma chain production, lead to an extremely heterogeneous cell population in the peripheral blood. Iron overload, due to increased gastrointestinal absorption and blood transfusion is the major cause of tissue damage, morbidity and death. PMID- 10872476 TI - Thalassaemia: clinical management. AB - Advances in the management of thalassaemia major have greatly improved the prognosis for patients with this disease. In countries able to afford programmes of regular transfusion and iron-chelating therapy, survival to the fourth decade is now common, and most complications associated with the primary disease are now infrequently observed. This situation stands in contrast to that in emerging countries, where the widespread implementation of these expensive treatment regimens is still awaited. This review will focus on recent advances in the treatment of thalassaemia and briefly review the progress in experimental approaches to treatment of this disorder. PMID- 10872477 TI - Pathophysiology of sickle cell disease. AB - Sickle cell disease is caused by a mutation in the beta-globin chain of the haemoglobin molecule. Sickle haemoglobin, the result of this mutation, has the singular property of polymerizing when deoxygenated. Exactly how normal tissue perfusion is interrupted by abnormal sickle cells is complex and poorly understood. Despite genetic identity at the site of the sickle haemoglobin mutation, all patients with sickle cell anaemia are not affected equally by this disease. Secondary genetic determinants and acquired erythrocyte and vascular damage are likely to be central components of the pathophysiology of sickle cell anaemia. PMID- 10872478 TI - Sickle cell disease: clinical management. AB - Sickle cell syndromes are a group of inherited disorders of haemoglobin structure that have no cure in adults at the present time. Bone marrow transplantation in children has been shown to be curative in selected patients. The phenotypic expression of these disorders and their clinical severity vary greatly among patients and longitudinally in the same patient. They are multisystem disorders and influence all aspects of the life of affected individuals including social interactions, family relations, peer interaction, intimate relationships, education, employment, spiritual attitudes and navigating the complexities of the health care system, providers and their ancillary functions. The clinical manifestations of these syndromes are protean. In this review emphasis is placed on four sets of major complications of these syndromes and their management. The first set pertains to the management of anaemia and its sequelae; the second set addresses painful syndromes both acute and chronic; the third set discusses infections; the fourth section deals with organ failure. New experimental therapies for these disorders are briefly mentioned at the end. Efforts were made to include several tables and figures to clarify the message of this review. PMID- 10872479 TI - Prenatal diagnosis and screening of the haemoglobinopathies. AB - This paper reviews the most important aspects of carrier detection procedures, genetic counselling, population screening and prenatal diagnosis of the thalassaemias and sickle cell anaemia. Carrier detection can be made retrospectively, following the birth of an affected child, or prospectively. Carrier detection and genetic counselling in at-risk populations for alpha thalassaemia and sickle cell anaemia is carried out mostly retrospectively. However prospective carrier screening is ongoing in Cuba and Guadeloupe for sickle cell anaemia and, in a very limited way, in some South East Asian populations, for alpha-thalassaemia. For beta-thalassaemia, several programmes, based on carrier screening and counselling of couples at marriage, preconception or early pregnancy, are operating in several Mediterranean at-risk populations. These programmes have been very effective, as indicated by increasing knowledge on thalassaemia and its prevention by the target population and by the marked decline of the incidence of thalassaemia major. Carrier detection is carried out by haematological methods followed by mutation detection by DNA analysis. Prenatal diagnosis is accomplished by mutation analysis on PCR-amplified DNA from chorionic villi. Future prospects include automation of the process of mutation detection, simplification of preconception and preimplantation diagnosis and fetal diagnosis by analysis of fetal cells in maternal circulation. PMID- 10872480 TI - Pharmacological therapy. AB - Collectively sickle cell disease and beta-thalassaemia are the most commonly inherited single-gene defects world-wide and were the first group of diseases for which DNA-based detection strategies were utilized. Although genotypically distinct, these two groups of diseases exhibit several common clinical features: moderate-to-severe haemolytic anaemia, acute and progressive tissue damage, disease- or treatment-related organ failure and premature death. Within the last two decades, a striking improvement in life expectancy in the two patient populations has been observed, by dint of primary and secondary prevention strategies. However, apart from bone marrow transplantation, a generally applicable, specific and non-toxic form of treatment remains unavailable for these disorders. Nonetheless, a greater appreciation of the developmental control of human globin gene expression coupled with observations of the effects of certain classes of agents to 'reverse' erythroid cellular phenotype in in vitro and animal models have led to pharmacological trials to obtain meaningful increases in haemoglobin F production in patients affected by these two severe beta-globin disorders. Contemporary understanding of the quantitative relationship between the abnormal molecules in the red cells (aggregates of sickle haemoglobin) in the sickle cell syndromes and aggregated alpha-globin polypeptides in the beta-thalassemia syndromes, and the extent of the red cell and/or organ involvement, has now enabled investigators to predict how much inhibition of these intracellular pathogenic processes might be necessary to achieve partial or total abrogation of disease manifestations. The results of the Multicenter Study of Hydroxyurea and other controlled trials now bear out these predictions. PMID- 10872481 TI - Current status of stem cell therapy and prospects for gene therapy for the disorders of globin synthesis. AB - Sickle cell anaemia and beta-thalassaemia are today curable through the use of stem cell transplantation. Nevertheless, the disadvantages inherent in stem cell transplantation underscore the need for better therapies. A recent finding of potentially major importance is that complete eradication of host haematopoiesis is not an absolute requirement for achieving therapeutic effects in thalassaemia and sickle cell anaemia. Future stem cell transplantation protocols will use less toxic conditioning regimens in an effort to achieve a state of stable mixed chimerism between donor and host haematopoietic elements. An improved understanding of globin gene regulation and stem cell biology will allow for the first gene therapy trials for sickle cell anaemia and beta-thalassaemia in the relatively near future. Initial gene therapy protocols will emphasize safety, are likely to target progenitor cells, and will involve repeated cycles of mobilization, transduction and reinfusion, with little or no conditioning. These first generation gene therapy trials are unlikely to confer major therapeutic benefits, but will provide the foundation upon which subsequent, more effective protocols will be based. PMID- 10872482 TI - NIH and NSF funding mechanisms applicable to interventional radiology. PMID- 10872483 TI - Custom-made stent-graft of polytetrafluoroethylene-covered Wallstents: technique and applications. PMID- 10872484 TI - Treatment of inferior vena cava anastomotic stenoses with the Wallstent endoprosthesis after orthotopic liver transplantation. AB - PURPOSE: To evaluate the efficacy of the Wallstent endoprosthesis for treatment of stenotic or occlusive inferior vena cava (IVC) lesions refractory to balloon angioplasty in patients after orthotopic liver transplantation. MATERIALS AND METHODS: Wallstent endoprostheses were implanted in six patients with IVC anastomotic stenoses or occlusions that were refractory to balloon angioplasty. Follow-up included both duplex ultrasound (US) and clinical evaluations. RESULTS: Ten stents were successfully implanted in six patients. Five of six patients (83%) demonstrated primary patency on duplex US for a mean period of 11 months (range, 4-17 months). One patient's symptoms recurred within 3 weeks after intervention. This patient underwent repeated stent placement. Follow-up duplex US in this patient demonstrated primary assisted patency at 7 months. Mean clinical follow-up was 12 months (range, 7-18 months). Other than the previously described case, no patient developed recurrent symptoms of IVC stenosis or occlusion. Two patients who experienced hemorrhagic complications secondary to anticoagulation were treated successfully. CONCLUSIONS: The Wallstent endoprosthesis is a useful adjunct for treatment of IVC stenosis or occlusions in patients who have undergone orthotopic liver transplantation when these lesions are refractory to simple balloon angioplasty. PMID- 10872485 TI - The nitinol vascular occlusion plug: preliminary experimental evaluation in peripheral veins. AB - PURPOSE: To compare the nitinol occlusion plug with standard stainless steel coils for the occlusion of moderate-size peripheral veins. MATERIALS AND METHODS: The nitinol plug is a braided multilayered vascular occlusion device filled with thrombogenic polyester fibers. It is self-expanding and can be recaptured into its 6-F introducing sheath for repositioning prior to detachment. Ten occlusion procedures were performed in five dogs from a retrograde transjugular venous approach. Five nitinol plugs (diameter: 7.4 mm +/- 0.5) were deployed in five femoropopliteal veins (diameter: 6.5 mm +/- 0.7; mean oversizing 14.6%). Two sequential Gianturco coils (diameter: 7.4 mm +/- 0.9) were deployed in the corresponding contralateral veins (diameter: 6.2 mm +/- 0.8; mean oversizing 19.6%). Follow-up venography was performed at 1 month, following which the animals were killed and the vessels were explanted. RESULTS: Time-to-occlusion was significantly shorter with the nitinol plug as compared to two Gianturco coils (4.2 minutes +/- 3.4 vs 25.6 minutes +/- 14.1, respectively [P < .03]). At 1 month all but one coil-doublet (80%) had recanalized or migrated, compared to only one nitinol occluder (20%, P < .04). Histopathologic examination of plug occluded veins showed a uniform organized matrix and underlying intimal proliferative response. CONCLUSION: A single nitinol occluder resulted in significantly faster occlusion time and significantly lower recanalization or migration rate than two Gianturco coils, in moderate-size peripheral veins. PMID- 10872486 TI - Superior mesenteric artery stent placement in a patient with acute mesenteric ischemia. PMID- 10872487 TI - Use of a collagen plug device to seal a subclavian artery puncture secondary to intraarterial dialysis catheter placement. PMID- 10872488 TI - Fluoroscopic landmarks for optimal visualization of the proximal renal arteries. AB - PURPOSE: To accurately determine the in vivo orientation of the origin of the renal arteries from the aorta relative to a fluoroscopic bony landmark for optimal diagnostic arteriography and renal artery stent placement. MATERIALS AND METHODS: One hundred sixty abdominal computed tomography (CT) scans of patients in eight age groups (20-90 years) were reviewed to determine the angle of the origins of the renal arteries from the aorta relative to the long axis of the L-1 spinous process (L1SP). RESULTS: The right renal artery arises ventrally at an angle of 30 degrees (standard deviation [SD] = 15 degrees) from a plane orthogonal to the long axis of the L1SP. The left renal artery arises dorsally at an angle of 7 degrees (SD = 13 degrees) relative to the same plane. CONCLUSIONS: The optimal initial angle for angiographic evaluation of the origin of the renal artery and for renal artery stent placement is 30 degrees left anterior oblique (LAO) relative to the L1SP for the right renal artery and 7 degrees LAO for the left renal artery. Unfortunately, there is variability in the angle of the renal artery origins from the aorta which cannot be controlled for using this technique. In some patients, additional views will be necessary to optimally depict the origins of the renal arteries. PMID- 10872490 TI - Renal artery stents: indications and techniques. PMID- 10872489 TI - Carbon dioxide angiography: effect of injection parameters on bolus configuration. AB - PURPOSE: Predict the intravascular distribution of carbon dioxide during angiography. MATERIALS AND METHODS: Mathematical modeling was used to predict the flow pattern of CO2 in a pulsatile system as a function of the CO2 flow rate. Findings were validated in an in vitro pulsatile circuit. RESULTS: The annular flow pattern with filling of nearly the entire lumen with CO2 is the most desirable, followed by intermittent bubble flow (provided individual bubbles are large). Stratified flow relates to a continuous floating CO2 bubble. Configuration of the CO2 bolus depends on fluid properties, fluid velocity, flow rates, mean intraluminal pressure, pressure amplitude, pulse rate, and vessel diameter. In vessels with less than 10-mm inner diameter, annular flow can be achieved relatively easily with injection rates above 20-30 mL/sec. Higher rates are not expected to produce superior results. When imaging a 2-cm artery, the best that can be realized clinically is intermittent flow with large bubbles. Bubbles size increases with increasing CO2 flow rate. In aneurysms, only stratified flow can be achieved with reasonable injection rates. Periodicity of the flow patterns is determined by the pulsatile circuit and can produce indentations in the CO2 bolus, which can be mistaken for stenoses. CONCLUSIONS: Flow regime maps can be used to optimize bolus configuration during CO2 angiography. PMID- 10872491 TI - Experimental treatment of early chronic iliac vein thrombosis with a modified hydrodynamic thrombectomy catheter: preliminary animal experience. AB - PURPOSE: To evaluate the efficacy and safety of a new hydrodynamic catheter for removal of chronic iliac vein thrombus. METHODS: Unilateral iliac vein thrombosis was induced in seven pigs by combining permanent coil and temporary balloon occlusion. Thrombectomy was performed with a new hydrodynamic catheter (10 F S.E.T.) 3 days after thrombus induction. After thrombectomy, the animals were killed and the iliac veins were examined histologically. RESULTS: Complete thrombectomy (100% thrombus removal) was achieved in three of seven animals, 75% removal in three of seven animals, and only 30% removal in one animal. The average thrombus removal was 75%. Successful re-establishment of flow was achieved in five of seven cases. Histologically, the thrombi were partially organized, meeting the histologic criteria for early chronic venous thrombosis. Minor venous wall damage caused by the thrombectomy procedure without acute hemodynamic consequences was observed in four of seven cases. CONCLUSION: The 10 F S.E.T. catheter was reasonably effective in removing chronic iliac vein thrombus with no hemodynamically significant complications. PMID- 10872492 TI - Injury potential to venous valves from the Amplatz thrombectomy device. AB - PURPOSE: To evaluate the acute effects of the Amplatz thrombectomy device (ATD) on peripheral venous valves in a canine model. MATERIALS AND METHODS: ATD thrombectomy was performed in 17 veins, and control experiments with use of an 8 F sheath-dilator were performed in four veins. Prethrombectomy ascending venography was performed, followed by device passage across the vein segment. Post-thrombectomy ascending venography was then performed, followed by heparinization and euthanasia. The treated veins were carefully explanted and stored in formaldehyde for histopathologic examination. Severity of valve injury was graded on a scale of 0 to 4. RESULTS: In ATD-treated veins: 10 veins sustained no injury [grade 0] (diameter, 6.7 mm +/- 1.7; antegrade/retrograde approach, 5/5), five veins sustained mild injury [grade 1-2] (diameter, 5.2 mm +/ 0.8; antegrade/retrograde, 3/2), while the remaining two veins sustained moderate-to-severe injury [grade 3-4] (diameter, 5 and 6 mm; antegrade/retrograde, 1/1). In sheath-dilator treated veins: no injury [grade 0] in any of the four treated veins (mean diameter, 5.5 mm +/- 0.6; all retrograde). In ATD-treated veins, valve injury (of any grade) was significantly more frequent in veins 6 mm or less in diameter than in veins at least 7 mm in diameter (seven of 12 vs zero of five; P < .03). There was no significant association between thrombectomy approach and injury grade. CONCLUSION: Veins 7 mm or greater in diameter were associated with no significant valve injury during ATD thrombectomy. However, long-term and short-term effects on valvular function will need to be assessed. PMID- 10872493 TI - Infectious complications of 393 peripherally implantable venous access devices in HIV-positive and HIV-negative patients. AB - PURPOSE: To compare and investigate the rate of infection in patients with and without human immunodeficiency virus (HIV) who have implantable venous access devices placed by interventional radiologists. MATERIALS AND METHODS: Three hundred ninety-one patients undergoing radiologically guided placement of peripheral arm ports were grouped according to their HIV serologic status. Findings were prospectively reviewed in 393 peripherally placed arm ports that were implanted in the basilic, cephalic, or brachial vein under fluoroscopic or sonographic guidance over a 4-year span. Infectious complications were categorized according to severity (local or systemic) and time (periprocedural or late). RESULTS: Three hundred ninety-three ports have been indwelling for a total of 97,256 patient days (range, 1-694; mean duration, 247 days). Among the 30 catheter placements in 29 HIV-positive patients with a total exposure time of 7,242 days, five (one local and four systemic) infections occurred, resulting in a 16.6% overall infection rate, yielding 0.069 infections per 100 catheter days at risk (95% confidence interval [CI], 0.032-0.127). In the remaining 362 HIV negative patients, 27 (14 local and 13 systemic) infectious complications (7.4%) occurred, translating into 0.030 infections per 100 catheter days (95% CI, 0.021 0.042). The odds ratio of getting an infection from the implantable arm ports in the HIV-positive group was 2.5 times higher than that of the HIV-negative group. The relative risk was similar and was calculated to be 2.3. The P value was .084 (P < .05 required to be considered significant). CONCLUSIONS: These results suggest a significant difference in the infectious complication rate encountered in HIV-positive patients compared with the general population. However, the HIV positive peripheral arm port infection rate compares favorably with the surgically placed catheters and ports. Many more arm ports in HIV-positive patients must be evaluated for the data to achieve an acceptable level of statistical significance. PMID- 10872494 TI - Interventional radiology placement of twin Tesio catheters for dialysis access: review of 75 patients. AB - PURPOSE: To evaluate the safety and efficacy of modern interventional radiology techniques and imaging guidance for placement of jugular vein twin Tesio hemodialysis catheters. MATERIALS AND METHODS: Eighty-two sets (75 patients) of twin Tesio catheters were percutaneously placed in the right (n = 70) and left (n = 12) internal jugular veins with use of ultrasound (US) and fluoroscopic guidance. Immediate procedural and late complications were recorded. The efficacy of the Tesio system was also evaluated. RESULTS: With US and fluoroscopic guidance, the technical success for access and catheter placement was 100%. Measured dialysis blood flow rate of greater than 375 mL/min was obtained in 95% of the patients and recirculation averaged 4.6% +/- 5%. An inadvertent common carotid artery puncture occurred in one (0.6%) patient and prolonged exit site bleeding occurred in another five patients (3%). Each of these was successfully controlled with compression. More chronically, catheter thrombosis and exit site infection occurred each at the rate of 0.16 episodes per 100 catheter days. All thrombosis and exit site infections responded to local thrombolysis and antibiotic therapy, respectively. Bacteremia occurred in 20 patients and required catheter removal in five patients. There was no clinical evidence of upper extremity or superior central vein thrombosis. CONCLUSION: Placement of internal jugular, twin Tesio catheters with use of imaging and interventional techniques provides a safe and efficacious means of either short or long-term hemodialysis. PMID- 10872495 TI - Ultrasonically guided inferior vena cava stent placement: experience in 83 cases. AB - PURPOSE: Traditionally, inferior vena cava (IVC) stent placement is performed with fluoroscopic guidance. The object of this study was to evaluate use of ultrasound (US) as guidance for IVC stent placement for the management of Budd Chiari syndrome. MATERIALS AND METHODS: Eighty-three patients with IVC membranous stenosis (n = 30), membranous occlusion (n = 19), segmental stenosis (n = 21), or segmental occlusion (n = 13) underwent IVC recanalization, balloon dilation, and stent placement under US guidance. Among the 83 patients, 67 had at least one patent hepatic vein, while 16 patients had three occluded hepatic veins. RESULTS: IVC stents were successfully placed in 79 of 83 patients, with a success rate of 95%. After the procedure, the symptoms and signs of IVC obstruction disappeared or markedly improved in all patients, and the blockage of hepatic outflow was alleviated in 67 patients. Pericardial effusion, complete atrial ventricular block, and stent migration into the right atrium occurred, respectively, in one patient. During 1-46-month follow-up, stent restenosis occurred in one patient; the other stents remained open and functioned effectively. CONCLUSION: Because of the absence of nonionizing radiation and iodinated contrast material, and its low cost, US is well suited and often preferred for guidance of IVC stent placement. PMID- 10872496 TI - Experimental nonfluoroscopic placement of inferior vena cava filters: use of an electromagnetic navigation system with previous CT data. PMID- 10872497 TI - Hemodialysis graft declotting with the "lyse and wait" technique: a new name for an old recipe. PMID- 10872498 TI - Mesenteric venous thrombosis: successful treatment by intraarterial lytic therapy. PMID- 10872499 TI - Contrast angiography and MR angiography: still not optimum. PMID- 10872500 TI - Carbon dioxide angiography: complications and pseudocomplications. PMID- 10872501 TI - Femoropopliteal graft entrapment: angiographic demonstration with MR correlation. PMID- 10872502 TI - Mutation detection in the breast cancer gene BRCA1 using the protein truncation test. AB - About 400 distinct mutations have been defined in the BRCA1 gene, and these are spread fairly evenly through the 5592 bp of coding DNA. This circumstance presents a formidable challenge for mutation screening. Apart from total direct sequencing, the preferred screening method has been single-strand conformation polymorphism (SSCP) gels, with a smaller input from constant denaturant gradient electrophoresis (CDGE), heteroduplex (HD) analysis, and mismatch cleavage. The protein truncation test (PTT) was used early in BRCA1 mutation screening but has not been widely adopted, perhaps because a straightforward analysis of the whole BRCA1 gene requires working with RNA and all its perceived problems. The present work was undertaken to assess the practicality of using the PTT under routine conditions for the screening of long genes such as BRCA1 that are not highly expressed in lymphocytes. We conclude that, provided RNA preparation is carried out effectively and consistently, the PTT approach has significant advantages over other methodologies such as SSCP gels. PMID- 10872503 TI - In planta expression of HIV-1 p24 protein using an RNA plant virus-based expression vector. AB - Plant viruses show significant potential as expression vectors for the production of foreign proteins (e.g., antigens) in plants. The HIV-1 p24 nucleocapsid protein is an important early marker of HIV infection and has been used as an antigen in the development of HIV vaccines. Toward developing a plant-based expression system for the production of p24, we have investigated the use of a (positive)-strand RNA plant virus, tomato bushy stunt virus (TBSV), as an expression vector. The HIV p24 open reading frame (ORF) was introduced into a cloned cDNA copy of the TBSV genome as an in-frame fusion with a 5'-terminal portion of the TBSV coat protein ORF. In vitro-generated RNA transcripts corresponding to the engineered virus vector were infectious when inoculated into plant protoplasts; Northern and Western blot analyses verified the accumulation of a predicted p24-encoding viral subgenomic mRNA and the production of p24 fusion product. Whole-plant infections with the viral vector led to the accumulation of p24 fusion protein in inoculated leaves, which cross-reacted with p24-specific antibodies, thus confirming the maintenance of key antigenic determinants. This study is the first to demonstrate that TBSV can be engineered to express a complete foreign protein of clinical importance. Strategies for optimizing protein yield from this viral vector are discussed. PMID- 10872505 TI - Detection of recombinant P-glycoprotein in multidrug resistant cultured cells. AB - The MDR1 multidrug resistance gene encodes a high molecular weight membrane spanning cell surface protein, P-glycoprotein, that confers multidrug resistance by pumping various cytotoxic drugs, including vinblastine, doxorubicin or paclitaxel, out of cells. Overexpression of P-glycoprotein in human tumors has been recognized as a major obstacle for successful chemotherapy of cancer. Thus, P-glycoprotein represents an important drug target for pharmacological chemosensitizers. Initially, cell culture models to study the multidrug resistance phenotype were established by selecting drug-sensitive cells in step wise increasing, sublethal concentrations of chemotherapy agents. P-glycoprotein was found to be overexpressed in many of these models. Multidrug resistant cells can also be generated by transfection of cultured cells with the MDR1 gene, followed by selection with cytotoxic drug at a concentration that kills all untransfected host cells. Transfectants expressing wild-type or mutant recombinant P-glycoprotein have significantly contributed to our understanding of the structure of P-glycoprotein and its molecular and cellular functions. Additionally, the MDR1 gene has also been used as a selectable marker for the transfer and coexpression of non-selectable genes. This article details means for detection of P-glycoprotein in DNA-transfected or retrovirally transduced, cultured cells. Different experimental approaches are described that make use of specific antibodies for detection of P-glycoprotein. Strategies to visualize P glycoprotein include metabolic labeling using 35S-methionine, labeling with a radioactive photoaffinity analog, and non-radioactive immunostaining after Western blotting. PMID- 10872504 TI - Principles of affinity-based biosensors. AB - Despite the amount of resources that have been invested by national and international academic, government, and commercial sectors to develop affinity based biosensor products, little obvious success has been realized through commercialization of these devices for specific applications (such as the enzyme biosensors for blood glucose analysis). Nevertheless, the fastest growing area in the biosensors research literature continues to involve advances in affinity based biosensors and biosensor-related methods. Numerous biosensor techniques have been reported that allow researchers to better study the kinetics, structure, and (solid/liquid) interface phenomena associated with protein-ligand binding interactions. In addition, potential application areas for which affinity based biosensor techniques show promise include clinical/diagnostics, food processing, military/antiterrorism, and environmental monitoring. The design and structural features of these devices--composed of a biological affinity element interfaced to a signal transducer--primarily determine their operational characteristics. This paper, although not intended as a comprehensive review, will outline the principles of affinity biosensors with respect to potential application areas. PMID- 10872506 TI - Microassay analyses of protein glycosylation. AB - To accurately characterize the carbohydrate moieties of oligosaccharide chains in glycosylated proteins, it is necessary to distinguish exactly which types of oligosaccharides are present at which site. We describe lectin overlay assays, which take advantage of the ability of lectins to distinguish between different types of glycoproteins via recognition of terminal sugars, thus allowing the chain type and peripheral antigenic components to be determined. Three microassays involving lectins are reported in this paper: non-protease-treated intact glycoproteins; glycopeptides released by prior digestion of the glycoprotein and then separated by HPLC; and release of sugars from glycoproteins by hydrazinolysis and then coupling them to a multivalent support. PMID- 10872507 TI - Modulation of human interferon-gamma biosynthesis by antisense oligodeoxynucleotides. AB - We investigated the inhibition of human interferon-gamma (HuIFN-gamma) production in cultures of lymphocytes with the use of the antisense strategy. Out of a series of antisense oligodeoxynucleotides (ODN) complementary to different regions of the HuIFN-gamma gene, a 16-mer specific for a sequence including the translation initiation codon was the most effective. Here we describe a detailed protocol for the isolation of lymphocytes from buffy coats, the rational design of antisense ODN, and the monitoring of HuIFN-gamma production of the antisense ODN-treated cells. PMID- 10872508 TI - High-copy cDNA amplification of minimal total RNA quantities for gene expression analyses. AB - This protocol describes a PCR-based cDNA amplification technique of small total RNA quantities, optimized for determination and verification of gene expression variations in cells or tissue specimen. A proportional amplification of rare and abundant transcripts is thereby achieved by initial random hexamer-primed reverse transcription of total RNA. Compared to established oligo(dT)-primed techniques, this approach generates shorter than full length copies of long RNAs which leads to a normalized cDNA pool for a more adequate PCR-amplification. Subsequent double oligo(dA) tailing of the synthesized total cDNA strands and the utilization of heteropolymeric primers allow a highly specific, up to 500-fold PCR-amplification of the total cellular RNA amount. Thus, obstacles in availability of RNA from limited sources, such as human biopsies or microdissected histological sections, can be overcome. The amplified total cDNA (atcDNA) is shown to be applicable for confirmation of differential gene expression, as demonstrated in this protocol by expression analysis of the multidrug resistance-associated genes mdr1, mrp1 and lrp, using human cell lines as well as microdissected human tissue sections. PMID- 10872509 TI - Ligninolysis. A very peculiar microbial process. AB - Lignin, the most abundant renewable source of aromatic carbon on earth, consists in a highly irregular three dimensional biopolymer of oxygenated phenylpropanoid units. In natural environments, lignin is only degraded efficiently by some basidioamycetes that secrete an array of enzymes for this purpose. Recent advances in our understanding of the mechanism of lignin breakdown have revealed several features that make this process highly unique from a biochemical standpoint. This article summarizes some of them. PMID- 10872510 TI - Pathogenesis of Cryptococcus neoformans is associated with quantitative differences in multiple virulence factors. AB - Two isolates of Cryptococcus neoformans were previously described as being highly divergent in their level of capsule synthesis in vivo and in their virulence for mice. The highly virulent isolate (NU-2) produced more capsule than a weakly virulent isolate (184A) in vitro under tissue culture conditions and in vivo. This investigation was done to determine if there were differences between the two isolates in other factors that might also contribute to virulence. Growth rate was not a factor as NU-2 grew more slowly than 184A. Based on PCR fingerprinting the two isolates were genetically different providing an opportunity to examine differences in multiple virulence traits. Quantitative analysis revealed that NU-2 expressed significantly more melanin and mannitol than did 184A. Although the isolates expressed the same capsular chemotype, NU-2 produced an additional structure reporter group (SRG) under tissue culture conditions that was not present when grown in glucose salts/urea/basal medium (GSU). Capsular polysaccharide SRGs of 184A were unaffected by shifting the growth conditions from GSU to tissue culture conditions. Our results suggest that pathogenesis of a C. neoformans strain is dictated by the quantitative expression of the strain's combined virulence traits. Regulators of the expression of these genes may be playing key roles in virulence. PMID- 10872511 TI - Isolation and identification of a 92-kDa stress induced protein from Candida albicans. AB - It was previously shown that the presence of estrogen enhances survival of Candida albicans under heat and oxidative stresses. A 92-kDa protein is inducible by heat shock and estrogen in C. albicans. Previous studies have described this protein as hsp90 because of its molecular size and heat inducibility as seen on electrophoretic gels and Western blots. In this study, ion exchange, hydroxyapatite and size exclusion chromatography were used to isolate a 92-kDa protein band. The N-terminal sequence of isolated protein blotted onto a PVDF membrane was determined to be V-Q-S-?-V-L-G-F-P-R. This sequence is homologous to the N-terminal sequence of the MET6 gene product, cobalamin-independent methionine synthase, from Saccharomyces cerevisiae. The results of this study suggest that a cobalamin-independent methionine synthase homolog is inducible by heat and estrogen in C. albicans. This study also suggests that Candida hsp90 is more likely to exist as an 82-kDa protein as predicted by a previously described cDNA and not as a 92-kDa protein as reported in the literature. PMID- 10872512 TI - Testing the association between residential fungus and health using ergosterol measures and cough recordings. AB - Questionnaire surveys in several countries have consistently detected an association between symptoms and residential mould growth. Confirmation by objective measures would strengthen the argument for causality. To address this issue, quantitative and qualitative fungal measures (airborne ergosterol and viable fungi in dust) were compared to respiratory symptoms (n = 403) and nocturnal cough recordings (n = 145) in Canadian elementary schoolchildren during the winter of 1993-1994. There was a 25 percent to 50 percent relative increase in symptom prevalence when mould was reported to be present (p < 0.05). However, neither symptoms nor recorded cough was related to objective measures of mould. In conclusion, the inability to find an association between objective measures of fungus and health suggest that either these objective measures, or the traditionally used questionnaire data are inaccurate. This discrepancy limits the acceptance of a causal relation between indoor fungal growth and illness. PMID- 10872513 TI - Onychomycosis in Malaysia. AB - The common etiological agents of onychomycosis are dermatophytes, molds and yeasts. A mycological nail investigation of onychomycosis using direct microscopy and culture was conducted by the Mycology Unit, Department of Medical Microbiology, University of Malaya from March 1996 to November 1998. The study involved 878 nail clippings or subungal scrapings from subjects with onychomycosis. On direct microscopy examination, 50% of the specimens were negative for fungal elements. On culture, 373 specimens had no growth; bacteria were isolated from 15 nail specimens. Among the 490 specimens with positive fungal cultures, 177 (36.1%) were dermatophytes, 173 (35.5%) were molds and 130 (26.5%) were Candida. There were 2% (10/490) mixed infections of molds, yeasts and dermatophytes. Trichophyton rubrum (115/177) and Trichophyton mentagrophytes (59/177) were the main dermatophytes isolated. The molds isolated were predominantly Aspergillus niger (61/173), Aspergillus nidulans (30/173), Hendersonula toruloidea (26/173) and Fusarium species (16/173). 96.9% of the Candida species identified were Candida albicans. PMID- 10872514 TI - Occurrence of Aspergillus section flavi and aflatoxin B1 in corn genotypes and corn meal in Argentina. AB - A study has been carried out in Argentina on samples of corn genotypes from a breeding station as well as in commercially available corn meal. All samples were analyzed for fungal infection and aflatoxin B1. Mycological analysis of corn genotypes showed the presence of three principal genera of filamentous fungi Fusarium (100%), Penicillium (67%) and Aspergillus (60%). In the genus Fusarium three species were identified, F. moniliforme (42%), F. nygamai (56%) and F. proliferatum (1.8%). Eight species of Penicillium were identified, the predominant species isolated were P. minioluteum, P. funiculosum and P. variabile. In the genus ranked third in isolation frequency, two species were identified, A. flavus and A. parasiticus, the percentage of infection was 78% and 21%, respectively. Only one corn genotype was contaminated with aflatoxin B1 at a level of 5 ppb. The corn meal samples showed great differences in fungal contamination, the values ranging from 1 x 10(1) to 7 x 10(5) cfu g-1. Fusarium (68%), Aspergillus (35%) and Penicillium (21%) were the most frequent genera isolated. Among the genus, Aspergillus, A. parasiticus (38%) was the most frequent species isolated. All the samples of corn meal were negative to aflatoxin B1. These results indicate a low degree of human exposure to aflatoxins in Argentina through the ingestion of maize or corn meal. PMID- 10872515 TI - Mycotoxins of fungal strains from stored herbal plants and mycotoxin contents of Nigerian crude herbal drugs. AB - The ability of fungi isolated from stored herbal drug plants to produce mycotoxins in semisynthetic media was studied. The results obtained show that aflatoxins and ochratoxin A, were produced by Aspergillus flavus, A. parasiticus and A. ochraceus isolates. The time-production courses of aflatoxins B1, B2, 1 and ochratoxin A in crude herbal drug preparations show that more of these toxins were produced with increase in time of storage of the drugs. The results indicate that the potential exists for the toxigenic strains to elaborate mycotoxins in a large quantity in herbal drug substrates than in semisynthetic media. PMID- 10872516 TI - A survey of Fusarium toxins in cereal-based foods marketed in an area of southwest Germany. AB - A total of 237 commercially available samples of cereal-based foods including bread and related products, noodles, breakfast cereals, baby and infant foods, rice and other foods were randomly collected in southwest Germany during the first six months of 1998. The trichothecenes deoxynivalenol (DON), 3- and 15 acetyl-deoxynivalenol (3-,15-ADON), nivalenol (NIV), fusarenon-X (FUS-X), T-2 toxin (T-2) and HT-2 toxin (HT-2) were determined by gas chromatography/mass spectrometry following clean-up by a two stage solid-phase extraction. Detection limits ranged between 2 and 12 micrograms/kg. Based on all samples, the incidence of DON, HT-2, T-2, 3-ADON, 15-ADON, and NIV was at 71, 18, 4, 4, 4 and 2%, respectively; the average contents in positive samples were at 103, 16, 14, 17, 24 and 109 micrograms/kg, respectively. Fus-X was not detected in any sample. A lower (P < 0.05) DON content was found in baby and infant foods as well as in cookies and cakes compared to bread. Overall, based on the incidence and level of all six toxins, the degree of contamination was lowest in baby and infant foods. Foods produced from either white or whole grain flour did not differ (P > 0.05) with regard to the incidence and level of DON. In foods produced from cereals of organic production both the incidence and median content of DON was lower compared to conventional production. Zearalenone, alpha- and beta-zearalenol were determined by high performance liquid chromatography in 20 selected samples, mostly baby and infant foods. These toxins were not present in excess of the detection limit in any sample. PMID- 10872517 TI - Scarless endoscopic thyroidectomy: breast approach for better cosmesis. AB - An original technique for performing endoscopic thyroidectomy using a breast approach to avoid an operative scar in the neck was developed. The subcutaneous space in the breast area and the subplatysmal space in the neck were bluntly dissected through a 15-mm incision between the nipples, and CO2 was insufflated at 6 mm Hg to create the operative space. Three trocars were inserted at the breast, and dissection of the thyroid and division of the thyroid vessels and parenchyma were performed endoscopically using an ultrasonically activated scalpel. Four hemithyroidectomies and one partial resection of the thyroid for five female patients with thyroid adenomas 5 to 7 cm in diameter were successfully performed using this procedure. There were no conversions to open surgery or complications. No scars were apparent in the neck, and all patients were fully satisfied with the cosmetic results. Endoscopic thyroidectomy using a breast approach and low-pressure subcutaneous CO2 insufflation is a feasible and safe procedure, which results in satisfactory cosmetic results. PMID- 10872518 TI - Endoscopic thoracic sympathectomy for treatment of essential hyperhidrosis syndrome: experience with 650 patients. AB - Patients with essential hyperhidrosis (EH) syndrome may experience subjective suffering and social/occupational challenges. We examined the safety and efficacy of minimally invasive endoscopic surgery for treating EH. Single bilateral incisions, followed by endoscopic thoracic sympathectomy (ETS)-mediated bilateral ablation of the T2 sympathetic ganglia, were used to treat 650 patients with a primary diagnosis of palmar (90%) or facial hyperhidrosis (10%). Palmar and facial hyperhidrosis were resolved in 584 of 585 (>99%) and 62 of 65 (95%) patients, respectively. Surgery required less than 1 hour, and no patient experienced a life-threatening adverse event. Compensatory sweating was observed in 83% of patients and was considered mild or moderate in approximately 67% of those patients. Innovations in ETS have resulted in minimally invasive, highly efficient, safe treatment of EH. Surgery is minimally intrusive to patients, who were usually discharged within 2 hours after surgery and able to resume normal activities within 1 week. PMID- 10872519 TI - The management of Mirizzi syndrome in the laparoscopic era. AB - Mirizzi syndrome is a rare complication of long-standing gallstone disease resulting in obstructive jaundice. Careful perioperative management is of utmost importance because of an increased risk of bile duct injury intraoperatively. Experience with Mirizzi syndrome over a period of 3 years, from January 1996 to December 1998, was reviewed. Twenty-seven patients were operated upon, which constituted 0.9% of 2840 patients who underwent laparoscopic cholecystectomy in the authors' department. There were 12 patients with Mirizzi type I syndrome and 15 patients with Mirizzi type II syndrome, according to McSherry classification. Six (22%) conversions were reported, all because of unclear anatomy and inherent limitations of the laparoscopic approach. For the remaining 21 (78%) patients, the procedure was completed laparoscopically. No bilioenteric anastomosis was required. A preoperative stent insertion in the common bile duct (CBD) during endoscopic retrograde cholangiopancreatography (ERCP) enabled us to achieve primary closure of CBD in every case. There was no perioperative mortality, and patients remained well for an average 2.1-year follow-up. It is highly desirable to have a preoperative diagnosis of Mirizzi syndrome, and the laparoscopic approach is not a contraindication in specialized centers. Our current management protocol to treat Mirizzi syndrome consists of a high degree of suspicion at ERCP, with stenting preoperatively and a complete stone clearance with subtotal cholecystectomy intraoperatively. PMID- 10872520 TI - Laparoscopic treatment of Mirizzi syndrome. AB - Mirizzi syndrome is a rare disorder and remains a surgical challenge. It is generally considered as a contraindication to laparoscopic surgery. Three patients with Mirizzi type II syndrome and two patients with Mirizzi type I syndrome were treated laparoscopically. Partial cholecystectomy with fundus-first dissection of the gallbladder was performed, and closure of the fistula in type II syndrome was achieved over a T-tube. The common bile duct (CBD) was explored in one patient using a choledochoscope through the fistula. The procedure was completed laparoscopically in all five patients. The three patients with type II syndrome had residual CBD stones, which were associated with significant morbidity and mortality. Laparoscopic treatment of Mirizzi type I syndrome is technically feasible and safe. For Mirizzi type II syndrome, laparoscopic CBD exploration is demanding and experience, skill, and the full spectrum of modern technology are required. PMID- 10872521 TI - Laparoscopic wedge resection of gastric submucosal tumors. AB - Minimally invasive surgery has revolutionized the treatment of gastrointestinal tumors. Submucosal tumors (SMTs) of the stomach can be resected using laparoscopic techniques. Between 1993 and 1997, laparoscopic wedge resection was performed in 34 patients with an SMT of the stomach. The tumors ranged from 8 to 60 mm in diameter. All surgical margins were clear. The average operative time was 131 minutes. Most of the patients began eating on the first postoperative day and were discharged within 5 to 7 days. Histopathologic examination of the tumors showed gastrointestinal stromal tumor (n = 14), ectopic pancreas (n = 7), leiomyosarcoma (n = 4), schwannoma (n = 3), carcinoid (n = 2), leiomyoma (n = 2), an inflammatory lesion caused by parasites (n = 1), and cyst (n = 1). No recurrences were observed over the 5-year follow-up period. A solid SMT of the stomach larger than 20 mm in diameter can be treated using laparoscopic wedge resection. PMID- 10872522 TI - Sutureless laparoscopic extraperitoneal inguinal herniorrhaphy using reusable instruments: two hundred three repairs without recurrence. AB - Laparoscopic extraperitoneal hernia repair has several distinct advantages over the anterior repair and the laparoscopic transabdominal preperitoneal method. Laparoscopic extraperitoneal hernia repair allows detection and repair of occult contralateral defects with minimal risk of intraabdominal injury or adhesion formation and is associated with less pain and a quicker recovery. However, there are disadvantages. Circumferential mobilization of the spermatic cord and the use of staples to secure the mesh have been associated with injury to the spermatic cord and nerves. The cost of the laparoscopic approach is higher than that of open herniorrhaphy. Additionally, it is more difficult to do because there is a poor understanding of the preperitoneal fascial anatomy. A method of totally extraperitoneal inguinal herniorrhaphy emphasizing anatomic dissection and landmarks is described. The authors use only reusable instruments, no balloon dissector, and no fixation of the mesh. The wide dissection of the myopectineal orifice allows placement of a large mesh and utilizes intraabdominal pressure alone to secure the mesh on the posterior aspect of the abdominal wall, as described by Stoppa et al. (1). Operative costs are minimized. From experience with 203 sutureless extraperitoneal repairs, a low incidence of complications and no recurrences are demonstrated. It is extrapolated that the cost of this laparoscopic repair will approximate more closely that of open anterior herniorrhaphy. PMID- 10872523 TI - Intraoperative and postoperative complications of totally extraperitoneal laparoscopic inguinal hernioplasty. AB - Inguinal hernioplasty using extraperitoneal laparoscopy is a new surgical option but still controversial because of the great technical difficulty involved. To analyze the clinical factors that could be related to intraoperative and postoperative morbidity, a prospective study was performed of 131 patients (153 repairs) undergoing totally extraperitoneal endoscopic surgery for inguinal hernia in an Outpatient Surgery Unit. Clinical parameters (age, sex, associated diseases, prior abdominal surgery, site and type), intraoperative complications (detachment of epigastric vessels, preperitoneal bleeding, rupture of the peritoneal sac, subcutaneous emphysema, problems with extending the mesh, visceral or deferential lesions, and rate of reconversion), postoperative complications (haematomas, urinary retention, transitory pain, neuralgias, and infections), and rate of recurrence were evaluated. Follow-up averaged 18 months (range, 1-3 years) and was complete in 100% of the patients. Intraoperative morbidity was 47%; postoperative, 16%; and the rate of reconversion, 4%. The rate of readmissions was 0%. One patient underwent reoperation for suspected early recurrence. The following statistically significant relations were shown: bleeding to recurrent hernias; presence of pain to hematomas; peritoneal rupture to female sex, diabetes, prior infraumbilical surgery and bilateral site; detachment of epigastric vessels to absence of prior surgery and hernia type 3a; and hematomas to age older than 50 years (P < 0.05). The preperitoneal laparoscopic technique is a difficult surgical operation, which often requires added interventions to resolve unexpected problems. The complications are acceptable, and the rate of recurrence is low (0.65%). We establish a standard for selecting patients during a program of apprenticeship. PMID- 10872524 TI - Incisional hernia and fascial defect following laparoscopic surgery. AB - Complications involving the abdominal wall, particularly incisional hernias, were not expected when laparoscopic procedures were first introduced. With the increasing number of laparoscopies in abdominal surgery, more incisional hernias are observed. The authors report 13 cases of umbilical incisional hernia, which occurred late after laparoscopic cholecystectomy, and one case of omental procidentia through a lateral port, which occurred early after laparoscopic hernia repair with the transabdominal preperitoneal technique. There are 4 men and 10 women (mean age, 59.8 years; range, 40-74 years). Between March 1991 and December 1997, a total of 1,287 patients underwent laparoscopic operations at the Surgical Department of the Gradenigo Hospital in Turin, Italy. Incisional hernia incidence is 1%. Risk factors, such as chronic bronchitis or weight increase, which give rise to endoabdominal pressure, are present in some cases. Malnutrition may have a major role in many cases. Calculi larger than 15 mm are also seen frequently. Postlaparoscopy incisional hernia is generally a minor complication--only once did its occurrence cause a strangulated hernia. All precautions, including fascial suturing, must be taken to reduce the 1% incidence of postoperative incisional hernias. PMID- 10872525 TI - The use of liposucker for spleen retrieval after laparoscopic splenectomy. AB - The retrieval of spleen after laparoscopic splenectomy has long been a problem. Frequently, it is necessary to extend the wound for retrieving the spleen intact and to prevent potential spillage of splenic tissue into the peritoneal cavity. We describe the application of the liposuction unit to remove the spleen piecemeal after laparoscopic splenectomy. We have found this technique easy to apply and safe, without the necessity of excessive wound extension, while preserving splenic tissue for histologic examination. PMID- 10872526 TI - Granulation stenosis caused by a Dumon stent placed for endobronchial tuberculous stenosis. AB - Two patients with cicatric tracheobronchial stenosis caused by tuberculosis who suffered granulation stenosis after placement of a Dumon stent are reported. Dumon stents, which were long enough to cover the stenotic sites, were placed in the trachea and left main bronchus of each patient. Granulation tissue grew at both edges of the stent 3 or 4 months after stent placement, which caused restenosis and necessitated removal of the stents. The authors conclude that a Dumon stent for treatment of tracheobronchial stenosis caused by tuberculosis can cause granulation stenosis at the edges of the stent. PMID- 10872527 TI - Tension pneumopericardium during laparoscopy for trauma. AB - A case involving a patient with multiple stab wounds to the thoracoabdomen is reviewed. Laparoscopy was employed to determine peritoneal penetration. Hemodynamic collapse occurred secondary to tension pneumopericardium, which resulted from a 15 mm Hg pneumoperitoneum. The relevant literature is reviewed. PMID- 10872528 TI - Improving craft through science in health care. PMID- 10872529 TI - Audit on laparoscopic cholecystectomy: a lesson to learn. PMID- 10872530 TI - The quality of clinical care in diabetes in Britain--could do better. PMID- 10872531 TI - The relevance of the glycaemic index to our understanding of dietary carbohydrates. AB - AIMS: To review the evidence for the importance of glycaemic index of dietary carbohydrate in disease prevention and control. METHODS: A critical appraisal of the literature published in English between and cited on Medline between January 1966 and October 1999. RESULTS: Using basic, intervention and epidemiological studies from experienced teams, evidence that the glycaemic index of diet may influence outcome in terms of cardiovascular risk, risk of metabolic syndrome diseases and pregnancy was found. CONCLUSIONS: Consideration of glycaemic indices in making dietary recommendations may be expected to produce additional health benefit. PMID- 10872532 TI - Effect of ageing and diabetes on glucose-dependent insulinotropic polypeptide and dipeptidyl peptidase IV responses to oral glucose. AB - AIMS: Glucose-dependent insulinotropic polypeptide (GIP) acts on the pancreas to potentiate glucose-induced insulin secretion (enteroinsular axis). GIP is rapidly inactivated in vivo by the enzyme dipeptidyl dipeptidase IV (DPP-IV). The current studies were designed to examine the effect of ageing, obesity and diabetes on GIP and DPP-IV responses to oral glucose. METHODS: Healthy controls (nine middle aged, age 42 +/- 2 years, body mass index (BMI) 33 +/- 1 kg/m2; nine elderly, age 71 +/- 1 years, BMI 30 +/- 1 kg/m2) and patients with Type 2 diabetes (12 middle aged, age 44 +/- 2 years, BMI 34 +/- 2 kg/m2; 19 elderly, age 74 +/- 1 years, BMI 31 +/- 1 kg/m2) underwent a 3-h oral glucose tolerance test (OGTT) (glucose dose 40 g/m2). RESULTS: Insulin responses were similar in elderly controls and patients with diabetes, but were lower in middle-aged patients with diabetes than in controls (308 +/- 65 vs. 640 +/- 109 pM, P < 0.05). GIP responses were similar in controls and patients with diabetes in each age group, but were higher in elderly controls (middle-aged 45 +/- 13; elderly 112 +/- 13 pM, P < 0.01) and patients with diabetes (middle-aged 55 +/- 10; elderly 99 +/- 10 pM, P < 0.01). DPP-IV levels were lower in patients with diabetes in both middle-aged (control 0.241 +/- 0.015; diabetes 0.179 +/- 0.017 delta OD/20 min, P < 0.05) and elderly groups (control 0.223 +/- 0.019; diabetes 0.173 +/- 0.010 delta OD/20 min, P < 0.05). CONCLUSIONS: It was concluded that ageing in obese subjects is associated with enhanced GIP responses to oral glucose. In addition, DPP-IV activity is reduced in middle-aged and elderly obese patients with diabetes. PMID- 10872534 TI - Non-dipping circadian blood pressure and renal impairment are associated with increased mortality in diabetes mellitus. AB - AIMS: To assess the relevance of circadian blood pressure variation to future morbidity and mortality in patients with diabetes mellitus. METHODS: A retrospective descriptive 4 year follow-up study of data collected after ambulatory blood pressure monitoring in a clinic setting. RESULTS: Seventy-five patients (46 male; 29 female) of whom 41 % had Type 1 diabetes and 59% Type 2 were followed up for a median of 42 months (11-56). The median creatinine for the whole group at baseline was 101 (56-501) micromol/l. The median circadian blood pressures for the total study population were 147 (110-194)/87 (66-109) mmHg during daytime and 132 (86-190)/77 (50-122) mmHg during night-time. Half of the patients exhibited a fall in night-time pressures to 10% lower than daytime pressures (dippers). Dippers were younger, 47 (32-75) years, than non-dippers, 57 (35-79) years, P = 0.03. Over time, dippers had a lower mortality than non dippers, with 8% deaths in the cohort of dippers, 26% deaths in the cohort of non dippers, P = 0.04. Cox regression analysis revealed significant contributions from age, duration of diabetes and baseline renal function to subsequent mortality in non-dippers. Analysing current degree of renal impairment and original dipper status together revealed that, of those patients whose creatinine remained normal, 7% of patients whose blood pressure dipped had subsequently died and 10% of non-dipping patients had died; of those patients whose creatinine unequivocally rose, 10% of dipping patients had died and 42% of non-dipping patients had died, P = 0.03 CONCLUSIONS: Loss of circadian variation in blood pressure is associated with an increased mortality rate, regardless of diabetes type. The combination of non-dipping and subsequent renal impairment leads to the highest mortality rate. The study suggests a role for ambulatory blood pressure monitoring in day-to-day clinical practice to select patients with nephropathy who are at greatest risk, in an effort to alter outcome. PMID- 10872533 TI - Carbohydrate sources and glycaemic control in Type 1 diabetes mellitus. EURODIAB IDDM Complications Study Group. AB - AIMS: Little information is available on the relationship between glycated haemoglobin levels and the source or amount of dietary carbohydrate. The present study compares the association of carbohydrate intake with HbA1c between European individuals with Type 1 diabetes mellitus injecting insulin once or twice per day and those with > or = 3 daily injections. METHODS: The relation of carbohydrate intake (total, cereal, fruit, vegetable, milk, and potato carbohydrate assessed by a 3-day dietary record) to HbA1c was examined in 2084 patients (age 32.6 +/- 10.2 years, duration of diabetes 14.8 +/- 9.5 years) included in the EURODIAB Complications Study. RESULTS: In both insulin injection regimens, an increased intake of total carbohydrate (% of energy) and a higher consumption of potato carbohydrates (g) were associated with higher levels of HbA1c, whereas an increased intake of vegetable carbohydrate (g) was inversely related to HbA1c. These tendencies were all more pronounced in persons with one or two daily insulin injections. Consumption of cereal and fruit carbohydrates (g) was not related to HbA1c, irrespective of the insulin injection regimen. A trend of HbA1c to increase with higher intakes of milk carbohydrate was confined to those with one or two insulin injections per day (test for interaction: P = 0.01). CONCLUSIONS: In particular, subjects with only 1 or 2 daily insulin injections per day should receive specific advice to correctly consider milk and potato carbohydrates. On the other hand, people with Type 1 diabetes may profit from a higher consumption of vegetable carbohydrates for their levels of HbA1c. PMID- 10872535 TI - Low birthweight and metabolic abnormalities in twins with increased susceptibility to Type 2 diabetes mellitus. AB - AIMS: To evaluate the role of environmental intra-uterine factors in determining the birthweights of twins with increased susceptibility to diabetes and discordant for abnormal responses to the oral glucose tolerance test (OGTT) and verify the possible association of within-pair birthweight differences and metabolic abnormalities in adult life. METHODS: Forty-six monozygotic (MZ) and 32 dizygotic (DZ) twins were enrolled; 13 MZ twins were discordant for impaired glucose tolerance (IGT) and/or hyperinsulinaemia compared to their co-twins. RESULTS: The 13 MZ discordant twins showed significantly lower birthweights than their normal co-twins (P < 0.001). When dividing all twins in those with the highest birthweights within the couple and those with the lowest, all subjects with abnormal OGTT were found in the latter group (P < 0.0001). Within-pair birthweight difference was significantly higher in MZ twins with abnormal OGTT and the metabolic syndrome compared to normal MZ twins. The relative risk of developing the metabolic syndrome was 8.7 (1.6-46.9) when comparing the higher tertile of within-pair birthweight differences (> or = 0.450 kg) to the two lower tertiles (< 0.450 kg). Logistic regression analysis confirmed within-pair birthweight difference as a significant predictor of abnormal responses to the OGTT and the metabolic syndrome. CONCLUSIONS: These data suggest a causative role for environmental intrauterine factors on the determination of birthweight and support the hypothesis that within-pair birthweight difference, rather than an absolute low birthweight, is responsible for the metabolic abnormalities in the adult life. PMID- 10872536 TI - Post-prandial administration of the insulin analogue insulin aspart in patients with Type 1 diabetes mellitus. AB - AIMS: In intensified insulin therapy, the recent development of short-acting insulin analogues with a very rapid onset of action forces a new discussion in terms of the optimal injection-meal interval. This study evaluated prandial glycaemia in patients with Type 1 diabetes following the subcutaneous injection of soluble human insulin (HI) and the insulin analogue insulin aspart (IAsp) at different injection-meal intervals and investigated whether administration of IAsp after the meal might provide satisfactory metabolic control. METHODS: In a randomized, double-blind, double-dummy, four-period crossover study, 20 Type 1 diabetic patients were investigated. Prandial insulin was administered 15 min before the start of the meal (HI(-15min)), immediately before the meal (HI(0min); IAsp(0min)) and 15 min after the start of the meal (IAsp(+15min)). RESULTS: Plasma glucose excursions from baseline levels during the 4 h (PGexc) were highest with HI(0min) (17.9 mmol.l(-1).h; P < 0.05 vs. other treatments) and were not statistically different for HI(-15min), IAsp(0min) and IAsp(15min) (13.6, 11.9 and 14.2 mmol.l(-1).h, respectively). Maximum concentration of plasma glucose (PGmax) was lowest with IAsp(0min) (11.2 mmol/l; P < 0.05 vs. other treatments). PGmax was comparable with HI(-15min), HI(0min) and IAsp(+15min) (13.3, 14.1 and 13.2 mmol/l, respectively). CONCLUSIONS: With regard to prandial glycaemia IAsp(+15min) is as effective as HI(-5min) and superior to HI(0min). Thus, post-prandial dosing of the insulin analogue IAsp offers an attractive and feasible therapeutic option for well-controlled patients with Type 1 diabetes mellitus. PMID- 10872538 TI - Application of the new ADA criteria for the diagnosis of diabetes to population studies in sub-Saharan Africa. American diabetes association. AB - AIMS: To examine the implications for epidemiological studies of the American Diabetes Association (ADA) recommendation that the fasting blood glucose at a lowered level becomes the main diagnostic test for diabetes on cross-sectional based data from sub-Saharan Africa. METHODS: Data from 11 surveys conducted in rural, peri-urban and urban Cameroon (n = 1804), South Africa (n = 3799) and Tanzania (n = 10013) which measured fasting (ADA criteria) and 2-h blood glucose concentrations during a standard 75 g OGTT (old WHO criteria) were analysed. RESULTS: The prevalence of diabetes was higher in eight of the 11 surveys when applying the new ADA compared to the old WHO criteria. With the exception of one population (Mara, Tanzania) the absolute difference in prevalence between the two classifications tended to be small (< 2%). There was considerable variation in the categorization of individuals using the ADA and old WHO criteria. The level of agreement between the two ranged from fair to good (Kappa statistic 0.17 0.86). The prevalence of impaired fasting glycaemia (IFG) was lower than that of impaired glucose tolerance (IGT) in 10 of the surveys and the agreement between the two was fair, < or = 0.26 in all the surveys. CONCLUSIONS: Although the use of the new ADA fasting criteria for prevalence surveys is an attractive and practical option, particularly in Africa, further information is required on the characteristics and prognosis of individuals classified as IFG or diabetic by the fasting criteria, prior to wide adoption of the ADA criteria. Ideally measurement of both fasting and two low glucose concentrations should remain the standard for epidemiological studies. PMID- 10872537 TI - Prevalence of gestational diabetes mellitus--do the new WHO criteria make a difference? Brazilian Gestational Diabetes Study Group. AB - AIMS: To describe the prevalence of gestational diabetes mellitus (GDM) according to the 1998 WHO provisional recommendations and compare it to that found with previous 1985 WHO criteria. METHODS: A total of 5564 consecutive women aged 20 years or more without diagnosis of diabetes mellitus outside of pregnancy in general prenatal care clinics of the National Health Service in 6 state capitals of Brazil, between their 20th and 28th gestational weeks were enrolled. RESULTS: Of the 5004 women who completed a 75-g oral glucose tolerance test, 379 (7.6%, 95% confidence interval (CI) 6.9% to 8.4%) had GDM by the 1998 criteria (fasting glucose > or = 7.0 mmol/l or 2 h glucose > or = 7.8 mmol/l). Of these 379 cases, only 21 (5.5%) had hyperglycaemia in the range considered diabetes mellitus outside pregnancy (fasting glucose > or = 7.0 mmol/l or 2 h glucose > or = 11.1 mmol/l); the remaining 358 (94.5%) had hyperglycaemia in the impaired glucose tolerance range (fasting glucose < 7.0 and 2 h glucose > or = 7.8 mmol/l and < 11.1 mmol/l). Using the 1985 criteria (fasting or 2 h glucose > or = 7.8 mmol/l), 378 cases of GDM were found, 15 in the diabetes range and 363 in the impaired glucose tolerance range. CONCLUSIONS: Prevalence of GDM is minimally altered by the new WHO definition. Although GDM is a common condition, the vast majority of the cases have hyperglycaemia in the range considered impaired glucose tolerance outside pregnancy. PMID- 10872539 TI - Evaluation of an on-call diabetes service in a large teaching hospital. AB - AIMS: To assess prospectively the activity and effectiveness of a diabetes specialist on-call service. METHODS: All requests for specialist advice received by the doctor on-call for diabetes in a large teaching hospital were recorded and analysed over a period of 3 weeks. RESULTS: The total number of calls was 135 (mean 45 per week) of which 48.1% were outside normal working hours. Requests for advice from surgical and medical ward staff accounted for 43% of calls, the remainder were from staff in the maternity ward (12.6%), staff in other hospitals in the city (3.7%), general practitioners (13.3%) and patients (27.4%). The time spent by the on-call doctor for diabetes responding to calls was a mean of 8.6 h per week. The number of acute admissions prevented by this service was estimated to be 11 (3.6 per week). CONCLUSIONS: Open access to specialist advice provided by a doctor with expertise in diabetes is an effective adjunct to the delivery of diabetes care in the setting of a large teaching hospital. PMID- 10872540 TI - A novel mutation in the hepatocyte nuclear factor-1alpha/MODY3 gene in Chinese subjects with early-onset Type 2 diabetes mellitus in Taiwan. AB - AIMS: The goal of this study was to determine the frequency of mutation in hepatic nuclear factor (HNF)-1alpha, a gene recently implicated as causing maturity-onset diabetes of the young (MODY) and to analyse the respective clinical presentations in an ethnically Chinese population. METHODS: Fifteen unrelated subjects (nine females and six males) aged less than 35 years who had early-onset diabetes were analysed to test the possibility that mutation of the HNF-1alpha gene was responsible for this disorder. Genomic DNA extraction, polymerase chain reaction and DNA sequence analysis were performed accordingly. RESULTS: One patient with MODY had a novel missense mutation in exon 3 of the HNF 1alpha gene (Y218C) in a region of the protein that corresponds to a predicted DNA binding domain. CONCLUSIONS: A Y218C mutation in HNF-1alpha gene was identified in one family in Taiwan. PMID- 10872541 TI - Bilateral optic atrophy following diabetic ketoacidosis. AB - Diabetic ketoacidosis (DKA) can result in neuropathic abnormalities of the somatic and the autonomous nervous systems. We report the case of a 50-year-old man with Type 1 diabetes of 20-year duration who after severe DKA lost vision in his right eye and only retain partial vision in his left. This case demonstrates that optic neural tissue is vulnerable to haemodynamic and metabolic complications of DKA. PMID- 10872542 TI - Why ask questions? PMID- 10872543 TI - Obesity and Type 2 diabetes mellitus. PMID- 10872544 TI - Evidence-based screening for gestational diabetes? PMID- 10872545 TI - Insulin resistance: a problem for clinicians and scientists. PMID- 10872546 TI - Persistent vomiting in patients with diabetes. PMID- 10872547 TI - Erythropoietin depletion and anaemia in diabetes mellitus. PMID- 10872548 TI - Clinical value of microalbuminuria in hypertension. AB - Microalbuminuria (MA) is a well recognized marker of cardiovascular complications in hypertension, but whether MA can predict adverse outcome in this clinical condition is still a subject for debate. The fact that in hypertensive cohorts those patients who showed an increase in albumin excretion rate also manifested an increased incidence of morbid events indicates that the presence of MA in hypertension may carry an increased cardiovascular risk. However, the prognostic significance of MA remains controversial because no results of prospective studies performed in hypertensive subjects without diabetes mellitus are available. Several factors can affect the prevalence of MA in hypertension, including severity of the disease, selection procedures, concomitant risk factors, degree of obesity, age, and sex distribution. This accounts for the large differences in the prevalence of MA that can be found in the literature, with prevalence rates going from a low of 4.7% to a high of 40%. There is still conflict over whether MA in hypertension is due to increased intraglomerular pressure or to glomerular damage. The data from the literature suggest that in subjects with mild hypertension the main determinant of albumin excretion rate is the haemodynamic load. In subjects with more severe hypertension and hypertensive complications, the augmented urinary albumin leak is probably the consequence of a systemic microvascular disturbance which involves the glomeruli. In this respect, the insulin resistance state often associated to high blood pressure appears as one of the main pathogenetic factors. Whether management of hypertensive populations may be improved by monitoring of albumin excretion rate and whether antihypertensive drugs which are more effective in decreasing urinary albumin can be more beneficial in patients with MA remains to be determined. PMID- 10872549 TI - Role of oxidative stress in cardiovascular diseases. AB - OBJECTIVES: In view of the critical role of intracellular Ca2 overload in the genesis of myocyte dysfunction and the ability of reactive oxygen species (ROS) to induce the intracellular Ca2+-overload, this article is concerned with analysis of the existing literature with respect to the role of oxidative stress in different types of cardiovascular diseases. OBSERVATIONS: Oxidative stress in cardiac and vascular myocytes describes the injury caused to cells resulting from increased formation of ROS and/or decreased antioxidant reserve. The increase in the generation of ROS seems to be due to impaired mitochondrial reduction of molecular oxygen, secretion of ROS by white blood cells, endothelial dysfunction, auto-oxidation of catecholamines, as well as exposure to radiation or air pollution. On the other hand, depression in the antioxidant reserve, which serves as a defense mechanism in cardiac and vascular myocytes, appears to be due to the exhaustion and/or changes in gene expression. The deleterious effects of ROS are mainly due to abilities of ROS to produce changes in subcellular organelles, and induce intracellular Ca2+-overload. Although the cause-effect relationship of oxidative stress with any of the cardiovascular diseases still remains to be established, increased formation of ROS indicating the presence of oxidative stress has been observed in a wide variety of experimental and clinical conditions. Furthermore, antioxidant therapy has been shown to exert beneficial effects in hypertension, atherosclerosis, ischemic heart disease, cardiomyopathies and congestive heart failure. CONCLUSIONS: The existing evidence support the view that oxidative stress may play a crucial role in cardiac and vascular abnormalities in different types of cardiovascular diseases and that the antioxidant therapy may prove beneficial in combating these problems. PMID- 10872550 TI - Adrenaline and hypertension: new evidence for a guilty verdict? PMID- 10872551 TI - Sleep-related breathing disorder is an independent risk factor for uncontrolled hypertension. AB - OBJECTIVE: To test the hypothesis that sleep-related breathing disorder (SRBD) is associated with poor blood pressure control in hypertensive patients independent from confounding factors such as age, body mass index, alcohol, smoking and daytime blood gases. DESIGN AND METHODS: This cross-sectional study of a sleep laboratory cohort was carried out at the University Hospital Sleep Disorders Centre, Marburg. The study comprised 599 patients referred for a sleep study, all of them with a documented history of systemic hypertension and/or previously initiated antihypertensive therapy. Data were obtained from a clinical interview, two unattended sleep studies and assessment of clinic blood pressure, cholesterol level, alcohol and nicotine consumption and daytime blood gases. The main outcome measure was a post hoc analysis of predictors for poor blood pressure control. RESULTS: Respiratory disturbance index (RDI) was significantly higher in patients with uncontrolled hypertension (blood pressure > or = 160 and/or 95 mmHg, n = 463) than in those with controlled hypertension (n = 136) (34.0 +/- 26.8 versus 27.0 +/- 23.5, P < 0.01). The relative proportion of patients with uncontrolled hypertension increased significantly as SRBD activity increased (chi2, P< 0.05). Body mass index was the only independent predictor (P = 0.006) of uncontrolled hypertension in the whole study sample. However, in the subset of patients aged < or = 50 years, RDI (P= 0.006) and age (P = 0.016) were the only independent predictors. The probability of uncontrolled hypertension increased by approximately 2% (B = 0.019, P= 0.006) for each RDI unit. CONCLUSION: SRBD should be considered, in addition to traditional confounders, as a risk factor for poor blood pressure control in younger hypertensive patients (< or = 50 years of age). PMID- 10872552 TI - beta2-adrenoceptor gene polymorphisms and blood pressure variations in East Anglian Caucasians. AB - OBJECTIVE: The amino-terminal polymorphisms, Arg16Gly and Gln27Glu, of the beta2 adrenergic receptor (beta2AR) have been shown to affect regulation of the receptor expression by an agonist in cell culture studies. The Arg16Gly polymorphism has also been recently shown to be associated with essential hypertension. We therefore evaluated whether the amino-terminal polymorphisms of beta2AR are associated with hypertension in a Caucasian population. SUBJECTS AND METHODS: We performed an association study in 298 hypertensive patients and an equal number of age-matched normotensive controls from the East Anglian region, with blood pressure assessed categorically and quantitatively. We also examined the influence of the amino-terminal polymorphisms on blood pressure response to beta-blockade in 144 of the patients randomly assigned to this class of drug. Genotyping of the Arg16Gly polymorphism was undertaken by a newly designed mismatched polymerase chain reaction (PCR) and digestion with Nde I, whereas the Gln27Glu polymorphism was genotyped by PCR followed by Fnu4H I cleavage. RESULTS: We found no differences in the genotype or allele frequencies of the beta2AR polymorphisms between hypertensive and normotensive participants. There was also no association between the beta2AR genotypes and variations in either basal blood pressure or the blood pressure response to a beta-blocker. CONCLUSION: These findings suggest that the amino-terminal polymorphisms of the beta2AR gene are unlikely to constitute major susceptibility for essential hypertension in the East Anglian population. PMID- 10872553 TI - The tissue renin-angiotensin system in rats with fructose-induced hypertension: overexpression of type 1 angiotensin II receptor in adipose tissue. AB - OBJECTIVE: Fructose feeding induces hypertension, insulin-resistance and hypertriglyceridemia in Sprague-Dawley rats. The mechanisms of fructose-induced hypertension are as yet unknown. Here we investigate the effects of fructose feeding and of varying salt intake on blood pressure, glucose tolerance, plasma renin activity, and tissue angiotensinogen, renin, and AT1 receptor mRNA levels in this model of hypertension. DESIGN AND METHODS: To investigate the role of the renin-angiotensin system in fructose-induced hypertension we measured angiotensinogen, renin and angiotensin II type 1 (AT1) receptor mRNA levels in tissues of Sprague-Dawley rats that were fed either standard rat chow or a diet containing 66% fructose. RESULTS: Blood pressure (P < 0.05) and triglyceride (P < 0.01) levels were significantly greater in the fructose-fed animals. Plasma glucose and insulin responses to an oral glucose load were significantly greater (P< 0.05) in fructose-fed than control rats. Angiotensinogen mRNA levels in liver and fat, and renin mRNA levels in kidney did not differ between fructose-fed and control animals. Levels of AT1 receptor mRNA were significantly greater in the fat obtained from fructose-fed rats than in that from control rats (P< 0.05), but this was not so in the kidney. To determine whether fructose-induced hypertension is dependent on dietary salt content, rats were fed standard rat chow and a fructose-enriched diet with low and high sodium chloride concentrations. Blood pressure increased significantly (P< 0.05) only in the fructose-fed rats receiving the high-salt diet Similarly, increased AT1 receptor mRNA levels were observed only in the fructose-fed rats that were maintained on the high-salt diet CONCLUSIONS: Fructose feeding induces hypertension in normal- or high-salt fed animals and it is associated with an increased expression of the AT1 receptor in adipose tissue. These findings suggest that AT1 receptors might play a role in the pathophysiology of metabolic and hemodynamic abnormalities induced by fructose feeding. PMID- 10872554 TI - Polymorphonuclear leukocytes (PMNs) functions in SHR, L-NAME- and DOCA/salt induced hypertensive rats. AB - OBJECTIVES: To clarify ex-vivo polymorphonuclear leukocytes (PMNs) functions, we examined superoxide anion (O2-) production and adhesion to a plastic plate of isolated PMNs obtained from spontaneously hypertensive rats (SHR/lzm), NG-nitro-L arginine methyl ester (L-NAME)- and deoxycorticosterone acetate (DOCA)/salt induced hypertensive rats. DESIGN: Sixteen week-old male SHR/Izm and Wistar-Kyoto rats (WKY/Izm) were used as a model of hypertension and its control, respectively. L-NAME-hypertension was induced by oral administration of 100 mg/kg per day of L-NAME twice daily for 4 weeks using 4-week-old male Wistar rats. DOCA/salt-hypertension was induced by once daily subcutaneous injection of 1 mg DOCA with 1% NaCl drinking water for 2 weeks using 8-week-old male Wistar rats with heminephrectomy. METHODS: Heparinized whole blood was obtained from abdominal aorta. PMNs were isolated by density gradient following dextran sedimentation. A production of superoxide anion (O2-) by PMNs stimulated with phorbol ester myristate acetate (PMA, 100 ng/ml) was determined by a superoxide dismutase (SOD)-inhibitable cytochrome-C reduction method. Adhesion of PMNs was evaluated by their protein content on a plastic plate measured by Lowry method. RESULTS: SHR/Izm showed a significant enhancement of O2- production by isolated PMNs compared with WKY/Izm. Rats treated with L-NAME showed a lower O2- production by PMNs compared to control animals. In DOCA/salt hypertensive rats, O2- production was not different from that in the control rats. Adherent function of isolated PMNs did not differ significantly among these hypertensive animal models. CONCLUSIONS: These results suggest that O2- production by circulatory PMNs is augmented in SHR, but not in L-NAME and DOCA/salt hypertensive rats. This enhanced function, which is also observed in human essential hypertension, might contribute to the development of cardiovascular damage in genetically determined hypertension. PMID- 10872555 TI - Lipoprotein (a), haemostatic variables and cardiovascular damage in hypertensive patients. AB - OBJECTIVE: Lipoproteins and coagulation factors are independent predictors of atherothrombotic events in the general population and their interaction may contribute to the development of cardiovascular damage. This study was designed to assess relationships between lipoproteins, haemostatic variables, and atherosclerotic complications in hypertensive patients. METHODS: In 389 untreated essential hypertensive patients recruited at a hypertension clinic, we measured plasma lipids, apolipoproteins, lipoprotein (a), apolipoprotein (a) isoforms, fibrinogen, and parameters that directly reflect the coagulation activation. Hypertensive patients were compared to 92 normotensive controls. RESULTS: Univariate analysis showed log lipoprotein (a) concentrations to be significantly correlated with age (P< 0.02), apolipoprotein B (P< 0.02), plasma fibrinogen (P< 0.001), and fibrin D-dimer (P< 0.001) levels, but not with body mass index, blood pressure, dietary fat intake, cholesterol, triglycerides, apolipoprotein Al, prothrombin fragment 1 + 2, and antithrombin III. The relationship of lipoprotein (a) with fibrinogen and D-dimer was present in both sexes, whereas the relationship of lipoprotein (a) with age and apolipoprotein B was found only in males. Multiple regression analysis showed that both fibrinogen and D-dimer were independently related with lipoprotein (a). Elevated fibrinogen, D-dimer, and lipoprotein (a) levels were significantly and independently associated with clinical evidence of atherosclerotic disease. To investigate whether the relationships of lipoprotein (a) with coagulation parameters are genetically determined, we analysed apolipoprotein (a) phenotypes in a subset of 188 hypertensive patients. While lipoprotein (a) levels were inversely correlated with apolipoprotein (a) isoform protein size, both fibrinogen and D-dimer concentrations were comparable in patients with apolipoprotein (a) isoforms of different size. CONCLUSIONS: This study demonstrates a relationship between lipoprotein (a) and clotting variables in hypertensive patients that may contribute to atherosclerotic damage in these patients. There is no evidence of a genetic background for this relationship. PMID- 10872556 TI - The 'adrenaline hypothesis' of hypertension revisited: evidence for adrenaline release from the heart of patients with essential hypertension. AB - OBJECTIVE: Whether adrenaline acts as a sympathetic nervous cotransmitter in humans and stimulates beta2-adrenoceptors to augment neuronal noradrenaline release remains a subject of considerable dispute. The aim of this study was to test if adrenaline is released from regional sympathetic nerves (in the heart) in patients with essential hypertension, and to investigate whether locally released adrenaline might enhance cardiac noradrenaline release. METHODS: Using dual isotope dilution methodology, adrenaline and noradrenaline plasma kinetics was measured for the whole body and in the heart in 13 untreated patients with essential hypertension and 27 healthy volunteers. All research participants underwent cardiac catheterization under resting conditions. RESULTS: At rest, there was negligible adrenaline release from the sympathetic nerves of the heart in healthy subjects, 0.27 +/- 1.62 ng/min. In contrast, in patients with essential hypertension, adrenaline was released from the heart at a rate of 1.46 +/- 1.73 ng/min, equivalent on a molar basis to approximately 5% of the associated cardiac noradrenaline spillover value. Cardiac noradrenaline spillover was higher in hypertensive patients, 24.9 +/- 17.0 ng/min compared to 15.4 +/- 11.7 ng/min in healthy volunteers (P< 0.05). Among patients, rates of cardiac adrenaline and noradrenaline spillover correlated directly (r= 0.59, P< 0.05). CONCLUSIONS: This study, in demonstrating release of adrenaline from the heart in patients with essential hypertension, and in disclosing a proportionality between rates of cardiac adrenaline and noradrenaline release, provides perhaps the most direct evidence to date in support of the 'adrenaline hypothesis' of essential hypertension. PMID- 10872557 TI - Reduction of plasma angiotensin II to normal levels by antisense oligodeoxynucleotides against liver angiotensinogen cannot completely attenuate vascular remodeling in spontaneously hypertensive rats. AB - OBJECTIVE: The exact role of angiotensinogen (AGT) in vascular remodeling has yet to be determined. In the present study, we examined the effects of reducing plasma AGT by intravenous injections with antisense oligodeoxynucleotides (ODNs) against AGT targeted to the liver on vascular remodeling in spontaneously hypertensive rats (SHRs). DESIGN AND METHODS: The ODNs against rat AGT were coupled to asialoglycoprotein (ASOR) carrier molecules, which serve as an important method for regulating liver gene expression. Male SHRs (n = 18) and age matched male Wistar- Kyoto (WKY) rats (n = 4) were used for this study. All animals were fed a standard rat diet throughout the experiment At 10 weeks of age, the SHRs were divided into three groups (n = 6); systolic blood pressure (SBP) was similar in each group. The control group received saline, the sense group was injected with the sense ODN complex and the antisense group was injected with the antisense ODN complex. WKY rats were fed for the same period of time. The ASOR-poly(L)lysine-ODN complex was injected into the tail veins twice a week. RESULTS: At the end of the treatment, a reduction in AGT mRNA levels in the liver and plasma AGT was observed only in the animals injected with antisense ODNs. Antisense ODNs significantly reduced the plasma angiotensin II (Ang II) concentrations to levels similar to those observed in WKY rats. Antisense ODNs significantly reduced the SBP (180.7 +/- 4.4 mmHg) and media cross-sectional areas of the aorta (1.11 +/- 0.02 mm2), which were still larger than those seen in WKY rats (140.3 +/- 2.1 mmHg, 0.84 +/- 0.02 mm2), compared with the SHRs injected with sense ODNs (225.2 +/- 4.4 mmHg, 1.24 +/- 0.02 mm2) and control SHRs (223.7 +/- 4.8 mmHg, 1.25 +/- 0.02 mm2). The aortic angiotensin-converting enzyme (ACE) activity and collagen concentrations, which were significantly higher than those seen in WKY rats, did not significantly change among the SHR groups. The aortic AGT, ACE, angiotensin II type 1 (AT1) receptor and angiotensin II type 2 (AT2) receptor mRNA also did not significantly change among the SHR groups. CONCLUSION: On the basis of these findings, plasma AGT is thus considered to play a role in the development of hypertrophy of smooth muscle in the aorta of SHRs, it is thought to have only a slight effect, however, on the remodeling of the matrix tissue when the suppression of hypertension is insufficient. PMID- 10872558 TI - Transforming growth factor-beta1 modulates angiotensin II-induced calcium release in vascular smooth muscle cells from spontaneously hypertensive rats. AB - OBJECTIVES: To investigate the role of transforming growth factor-beta1 (TGF beta1) on Ca2+-dependent mechanisms elicited by angiotensin II in aortic vascular smooth muscle cells (VSMC) of Wistar- Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). METHODS: Cai2+ release induced by angiotensin II (1 micromol/ l) was studied in cultured VSMC isolated from the aortas of 6-week-old WKY rats and SHR. Intracellular Ca2+ (Cai2+) was assessed in Fura-2 loaded cells using fluorescent imaging microscopy. Angiotensin II receptors were analysed by binding studies. RESULTS: Pretreatment of VSMC for 24 h with TGF-beta1 significantly increased angiotensin II-induced Cai2+ mobilization from internal stores in SHR, while Ca2+ influx was not altered. This effect involves tyrosine kinase and is not due to an increase in angiotensin II binding sites, or a change in the affinity of the receptors. By contrast, TGF-beta1 did not modify the response of VSMC from WKY rats to angiotensin II. CONCLUSIONS: These results help our understanding of the interactions between the pathways activated by TGF-beta1 and the G protein-coupled receptor signalling pathway, and their role in genetic hypertension. PMID- 10872559 TI - Insulin resistance and insulin pulsatility in essential hypertension. AB - OBJECTIVE: Studies in normal humans and in patients with type 2 diabetes mellitus have demonstrated a close inverse relationship between peripheral insulin sensitivity and the frequency of short-term insulin secretory pulses in the systemic circulation. Our objective was to study this relationship in essential hypertension. DESIGN: Study of insulin sensitivity and insulin pulse characteristics in hypertensive subjects and normotensive controls using well established techniques. METHODS: Twelve subjects with essential hypertension and 12 age- and sex-matched normotensive controls were recruited. Insulin action was measured using the glucose clamp technique combined with isotope dilution methodology. Insulin pulsatility in the peripheral circulation was assessed by sampling every 2 min for 90 min after an overnight fast Pulses were identified using the computer program Pulsar. RESULTS: Insulin sensitivity index (glucose infusion rate/ serum insulin) was lower in the hypertensive patients (P= 0.01) and fasting insulin was increased (P= 0.008) compared to controls. The frequency and amplitude of insulin pulses were similar in the two groups. Insulin pulse frequency and insulin sensitivity were inversely related in the normotensive group (r= -0.68, P= 0.015), but not in the hypertensive group (r= -0.23, P= 0.48). Insulin clearance was reduced in the hypertensive group (P= 0.03), and was inversely related to insulin pulse frequency in the two groups combined (r = 0.51, P= 0.01). CONCLUSIONS: Insulin action was not related to insulin pulse frequency in essential hypertension, in contrast to the situation in normal man. PMID- 10872560 TI - Reversal of cardiac hypertrophy and fibrosis by S21402, a dual inhibitor of neutral endopeptidase and angiotensin converting enzyme in SHRs. AB - OBJECTIVE: The major advantage of dual inhibitors of neutral endopeptidase (NEP) and angiotensin converting enzyme (ACE) is their ability to lower blood pressure irrespective of renin or volume status. The aim of this study was to determine whether dual NEP/ACE inhibition produces different effects on cardiovascular structure and fibrosis, hormonal parameters and inhibition of tissue enzymes compared with selective inhibition of ACE and NEP in the spontaneously hypertensive rat (SHR). METHODS: Male SHRs received the dual NEP/ACE inhibitor (S21402, 100 mg/kg per day), the ACE inhibitor (captopril, 50 mg/kg per day), the NEP inhibitor (SCH42495, 60 mg/kg per day) or vehicle for 2 weeks. RESULTS: S21402 produced equivalent blood pressure lowering effects to captopril (vehicle, 220 +/- 1 mmHg; S21402, 189 +/- 2 mmHg; captopril, 187 +/- 3 mmHg), but was a more effective antihypertensive agent than SCH42495 (214 +/- 2 mmHg, P< 0.01). All treatments reduced left ventricular mass (P< 0.05) and cardiac fibrosis (P< 0.01). S21402 inhibited renal NEP and ACE (P< 0.01), SCH42495 inhibited renal NEP (P < 0.01), and captopril inhibited renal ACE (P< 0.01). Captopril and S21402 increased plasma renin activity (P< 0.05), but the rise with S21402 was attenuated compared with that caused by captopril (P< 0.01). All treatments reduced plasma aldosterone levels (P< 0.01), and NEP inhibition with SCH42495 and S21402 increased plasma atrial natriuretic peptide (ANP; P< 0.05). CONCLUSIONS: These results indicate that selective NEP inhibition has major benefits in the regression of cardiac hypertrophy and reduction of fibrosis but has limited antihypertensive effects. The dual NEP/ACE inhibitor S21402 offered no advantage over the selective ACE inhibitor in terms of blood pressure reduction, or attenuation of cardiac hypertrophy and fibrosis, but did increase plasma ANP and blunted the reactive rise in renin with ACE inhibition. Further studies are needed to determine whether more complete blockade of the renin-angiotensin system with dual NEP/ACE inhibition results in additional benefits in terms of morbidity and mortality in cardiovascular disease. PMID- 10872561 TI - Increased left ventricular mass is not associated with impaired left ventricular diastolic filling in normal individuals. AB - BACKGROUND: Hypertensive left ventricular (LV) hypertrophy has been associated with diastolic dysfunction. However, the underlying physiological relationship between LV size and diastolic function remains to be clarified. The aim of this study was to evaluate the relationship between several measures of diastolic filling and LV mass in a population sample. METHODS: We used M-mode and Doppler echocardiography to compare left ventricular mass index (LVMI) and wall thickness with five measures of ventricular diastolic filling (ratio of the peak early mitral inflow velocity to the peak atrial mitral inflow velocity, deceleration time of early mitral inflow, isovolumetric relaxation time, ratio of the peak pulmonary venous systolic to diastolic flow and difference between the durations of the pulmonary venous and mitral inflow atrial waves) in 159 healthy volunteers. RESULTS: LVMI was significantly (P< 0.0001) greater in men (81.3 g/m2, interquartile range: 67-94) than women (59.7 g/m2, interquartile range: 49 74), but no gender differences were observed in diastolic filling. Higher age, blood pressure and heart rate showed significant correlation with diminished diastolic filling. However, no measure of diastolic filling correlated with LVMI or wall thickness in either univariate or multiple regression analyses that adjusted for relevant covariates. CONCLUSIONS: LVMI does not explain physiological differences in diastolic filling. The significant decline in diastolic filling with age reflects changes in the quality rather than the quantity of myocardial tissue. PMID- 10872562 TI - Markers of electrical instability in hypertensive patients with and without ventricular arrhythmias. Are they useful in identifying patients with different risk profiles? AB - BACKGROUND: Markers of electrical instability of the ventricular myocardium, namely abnormal repolarization and late potentials, are frequently observed in patients with hypertension when both ventricular arrhythmias and left ventricular hypertrophy are present. This information cannot be extrapolated to the population of hypertensive patients with ventricular arrhythmias but without left ventricular hypertrophy. OBJECTIVE: To evaluate QT duration, QT dispersion and the incidence of ventricular late potentials in patients with essential hypertension, already on anti-hypertensive therapy, both with and without non sustained ventricular arrhythmia. DESIGN: The study population consisted of 49 patients with essential hypertension who were compared to 89 control normotensive subjects both with and without frequent (> 30 per h) ventricular ectopic beats (VPBs). Patients were divided into four groups: (1) hypertensive patients without VPBs (H, n = 19), (2) hypertensive patients with VPBs (HA, n = 30), (3) normotensive subjects without VPBs (C, n = 28), and (4) normotensive subjects with VPBs (CA, n=61). METHODS: Echocardiographic parameters, QT interval, QT dispersion and signal-averaged ECG were evaluated without withdrawing anti hypertensive drugs. RESULTS: In no case was left ventricular hypertrophy documented. The number of VPBs during 24 h Holter recording (median 11 343 versus 7617) and the incidence of repetitive VPBs (37 versus 46% of patients) were similar in the two groups of patients (HA versus CA). Signal-averaged ECG parameters were normal and not different between the four groups. QT interval was longer in hypertensive patients compared to controls irrespective of the presence of VPBs. QT dispersion was slightly greater in subjects with VPBs, both hypertensive and normotensive, compared to subjects without arrhythmias. CONCLUSIONS: In patients with hypertension well-controlled by drug therapy and without left ventricular hypertrophy, frequent VPBs are not associated with markers indicating an electrophysiological substrate for re-entrant arrhythmias. However, QT prolongation suggests the persistence of a higher risk of cardiovascular mortality that is independent of the presence of VPBs. PMID- 10872563 TI - Determinants of left ventricular structure and mass in young subjects with sympathetic over-activity. The Tecumseh Offspring Study. AB - OBJECTIVE: In this study, we tested the hypothesis that sympathetic over-activity may cause metabolic abnormalities and affect left ventricular (LV) structure and mass early in life. SUBJECTS AND SETTING: The study population consisted of 111 healthy adolescents and young adults living in Tecumseh, Michigan (USA). MAIN OUTCOME MEASURES: Correlations of LV mass and structure with several clinical variables in relation to the activity of the sympathetic nervous system. METHODS: Power spectrum density estimates of heart rate variability were calculated with an auto-regressive method, and subjects were divided by cluster analysis into two groups according to low-frequency and high-frequency components. LV data were obtained by echocardiographic assessment RESULTS: Subjects with signs of sympathetic over-activity (n = 38, group 1) had higher heart rate, blood pressure (BP), waist/hip ratio and cholesterol levels than the rest of the group (n = 73, group 2). In group 1 subjects, insulin emerged as the strongest univariate correlate of interventricular septum and posterior wall thicknesses (P< 0.001 for both) and of LV mass (P= 0.009). These relationships remained significant when body mass index was accounted for. By contrast, the marginal univariate relationship with diastolic BP did not remain significant in multivariate analysis. In group 2 subjects, BP was strongly correlated with LV wall thickness and mass both in univariate (P values from 0.03 to < 0.001) and multivariate analyses, while insulin was not. The interactive effect of sympathetic activity and insulin on echocardiographic data was confirmed by multivariate analyses performed in the subjects grouped together (P values from 0.02 to 0.001 for the sympathetic activity x insulin interaction term). CONCLUSIONS: In young subjects with heightened sympathetic activity and initial metabolic abnormalities, insulin is a strong determinant of LV wall thickness and geometry, while in subjects with normal autonomic nervous system activity, the main determinant of left ventricular size is the haemodynamic load. PMID- 10872564 TI - Effect of high NaCl diet on spontaneous hypertension in a genetic rat model with reduced nephron number. AB - OBJECTIVE: An inherited reduction in nephron number has been implicated in the development of salt-sensitive hypertension and end stage renal disease. The Munich Wistar Fromter (MWF) rat represents a genetic model with a 30-50% reduction of nephrons compared with normal rats. MWF rats develop spontaneous hypertension and increased urinary albumin excretion (UAE). We addressed the question whether the inherited defect in this model leads to salt-sensitive hypertension. METHODS: At the age of 6 weeks, we started male and female MWF/Fub rats and salt-resistant Lewis (Lew) reference rats on either a normal NaCl (0.2%) or a high NaCl (8%) diet (n = 8, each group). Systolic blood pressure (SBP) and UAE were measured at 14 weeks. RESULTS: Under a normal diet, MWF/Fub rats demonstrated significantly elevated SBP compared to Lew rats both in male (165 +/ 2 versus 133 +/- 3 mmHg, P < 0.0001) and female (156 +/- 3 versus 134 +/- 3 mmHg, P < 0.0001) rats. After high NaCl treatment, SBP was significantly higher in both male and female MWF/Fub rats (+55 mmHg and +36 mmHg, P < 0.0001, respectively) compared with MWF/ Fub under a normal diet UAE was also significantly higher in male and female MWF/Fub rats after high NaCl excess (P < 0.0005, respectively). In contrast, both SBP and UAE remained unchanged in response to high NaCl in Lew rats. CONCLUSIONS: Our findings demonstrate that both the hypertension and UAE are sensitive to high NaCl loading in female and male MWF/Fub rats. Thus, an inborn nephron deficit may lead to salt-sensitive hypertension and renal dysfunction. PMID- 10872565 TI - Renal vascular morphology and haemodynamics in Dahl salt-sensitive rats on high salt-low potassium diet: neural and genetic influences. AB - OBJECTIVE: A dietary combination of high salt and low potassium (HS-LK) exacerbates hypertension in Dahl salt-sensitive (DS) rats and renders Dahl salt resistant (DR) rats hypertensive. In both strains, the hypertension is accompanied by remodelling of the renal resistance vasculature, and is attenuated by peripheral chemical sympathectomy. In the current study, we sought to determine whether the sympathetic nervous system is causally involved in mediating the renal vascular and haemodynamic alterations associated with HS-LK feeding in Dahl rats. DESIGN: Two groups each of DS and DR rats were maintained on HS-LK diet (8% NaCl, 0.2% KCl) for 8 weeks. One group of DS (n = 9) and DR (n = 8) were treated with 6-hydroxydopamine (6-OHDA) in 0.001 N HCl vehicle to chemically ablate peripheral sympathetic nerve terminals. The two remaining groups (n = 8 each) received equivalent injections of vehicle. METHODS: At the end of the dietary regimen, arterial blood pressure (ABP), glomerular filtration rate (GFR) and renal blood flow (RBF) were measured, and the structure of intra renal resistance vessels was examined by planar morphometric analysis of coronal sections prepared from perfusion-fixed kidneys. RESULTS: Both 6-OHDA-treated and untreated DS rats presented a greater degree of intra-renal vessel remodelling characterized by reduced lumen diameter in the absence (eutrophic) or presence (hypertrophic) of cross-sectional area expansion, higher renal vascular resistance (RVR) and lower GFR and RBF than DR rats. Chemical sympathectomy increased lumen diameters and reduced vascular wall expansion, resulting in a decrease in RVR and a concomitant increase in RBF and GFR in both strains; however, the effect was more prominent in the DS rats. CONCLUSIONS: We conclude that HS-LK-induced changes in intra-renal vessel structure and renal haemodynamic function in Dahl rats are, at least in part, dependent on the activity of the sympathetic nervous system. PMID- 10872566 TI - The efficacy and tolerability of losartan versus atenolol in patients with isolated systolic hypertension. Losartan ISH Investigators Group. AB - OBJECTIVE: To compare the efficacy and tolerability of angiotensin II (Ang II) antagonist losartan and the beta-blocker atenolol in the treatment of patients with isolated systolic hypertension (ISH) after 16 weeks of treatment. METHODS: A double-blind, randomized, multi-country study was carried out in 273 patients with ISH. Patients with a sitting systolic blood pressure (SiSBP) of 160-205 mmHg, and a sitting diastolic blood pressure (SiDBP) < 90 mmHg at screening and at placebo baseline were subjected to a 4-week placebo period and then randomly grouped to receive 50 mg losartan or 50 mg atenolol once daily for 16 weeks. At 8 and 12 weeks, patients not controlled (SiDBP > or = 160 mmHg) were given additional treatment of 12.5 mg hydrochlorothiazide (HCTZ) once daily. RESULTS: Similar significant reductions in SiSBPs (mean +/- SD) were obtained with 50 mg losartan and 50 mg atenolol, from 173.7 +/- 10.3 and 173.5 +/- 10.7 mmHg at baseline to 149.0 +/- 15.5 and 148.2 +/- 15.3 mmHg after 16 weeks of losartan or atenolol treatment respectively. Sixty-seven percent of the losartan-treated and 64% of the atenolol-treated patients remained on monotherapy throughout the study. Only 1.5% of the losartan-treated patients withdrew because of a clinical adverse event (CAE) compared with 7.2% in the atenolol-treatment group (P= 0.035). Drug-related CAEs were observed significantly more frequently with atenolol than with losartan treatment (20.3 versus 10.4%; P = 0.029). CONCLUSION: It is concluded that 50 mg losartan and 50 mg atenolol produced comparable reductions in SiSBP in patients with ISH but losartan was better tolerated. This is the first demonstration of the therapeutic value of selective Ang II receptor blockade with losartan in the treatment of ISH. PMID- 10872567 TI - Prevalence of target organ damage in treated hypertensive patients: different impact of clinic and ambulatory blood pressure control. AB - OBJECTIVES: First, to evaluate the prevalence of left ventricular (LV) hypertrophy, LV concentric remodelling and microalbuminuria in a selected sample of treated hypertensive patients with effective and prolonged clinic blood pressure (BP) control (BP < 140/90 mmHg). Second, to compare the prevalence of these markers of organ damage in patients with and without ambulatory BP (ABP) control, defined as average daytime BP < 132/85 mmHg). DESIGN AND METHODS: Fifty eight consecutive hypertensive patients who attended our hypertension outpatient clinic over a period of 3 months and were regularly followed up by the same medical team were included in the study. Obesity, diabetes mellitus, history or signs of cardiovascular or renal complications and major noncardiovascular diseases were the exclusion criteria from the study. Each patient underwent 24 h ABP monitoring, echocardiography and 24 h urine collection for albumin measurement. RESULTS: The prevalence of LV hypertrophy (LV mass index > 125 g/m2 in both sexes), LV concentric remodelling (relative wall thickness > 0.45) and microalbuminuria (urinary albumin excretion < 300 mg/ 24 h) in this selected group of patients (32 men, 26 women; mean age 53 +/- 9 years; mean clinic BP 122 +/- 9/ 78 +/- 6 mmHg) was markedly low (6.9, 8.6 and 5.1%, respectively). The 26 patients with effective ABP control (group I) were similar to the 32 patients without effective ABP control (group II) in age, gender, body surface area, clinic BP, smoking habit, glucose, cholesterol and creatinine plasma levels. Prevalence of LV hypertrophy, LV concentric remodelling and microalbuminuria was lower in group I than in group II (0 versus 12.9% P< 0.01, 7.7 versus 9.4% NS, 3.8 versus 6.2% NS, respectively). CONCLUSIONS: This study demonstrates that nonobese, nondiabetic hypertensive patients with an effective clinic BP control have a very low prevalence of target organ damage and that LVH is present only in individuals with insufficient ABP control. PMID- 10872568 TI - Effects of pyridinium chlorochromate adulterant (urine luck) on testing for drugs of abuse and a method for quantitative detection of chromium (VI) in urine. AB - Pyridinium chlorochromate (PCC) as an adulterant is popular for concealing drug positive results. When 11-nor-delta9-THC-9-carboxylic acid (THC-acid) in urine was treated with 2 mmol/L of PCC (Cr6+ 104 microg/mL), 58-100% of the THC-acid was lost. The loss increased with decreasing pH and increasing reaction time (0-3 days). Free codeine and free morphine remained unaffected by PCC at pH within the physiological range of the urine (pH 5-7). At lower pH, the loss of free morphine varied from 0 to 100%. Amphetamine, methamphetamine, benzoylecgonine, and PCP remained unaffected by PCC when exposed to the oxidant for three days in urine pH of 3-7. Chromium (VI) from PCC in a urine solution was detected by a color reaction with 1,5-diphenylcarbazide (DPC). When the reagent was added to the urine, an immediate red-violet color appeared. The chromium-DPC complex showed a characteristic absorption peak at wavelength 544 nm with a shoulder at wavelength 575 nm. The ratio of absorption was used to identify the chromium compound. The concentration of chromium (VI) was determined by measuring absorption at wavelength 544 nm and was linear over 0.5-20 microg/mL. The limit of detection of the procedure was 0.37 microg/mL. PMID- 10872569 TI - Determination of enantiomeric metabolites of l-deprenyl, d-methamphetamine, and racemic methamphetamine in urine by capillary electrophoresis: comparison of deprenyl use and methamphetamine use. AB - The enantiomeric analysis of urine collected from rats administered l-deprenyl, d methamphetamine (MA), or dl-MA and from healthy male volunteers who ingested l deprenyl by capillary electrophoresis (CE) using carboxy methylated-beta cyclodextrin (CMCD) as a chiral selector was investigated to compare the metabolic pattern of l-deprenyl with the metabolism of d- or dl-MA. Urine from illegal drug abusers was also analyzed for the comparison of therapeutic drug (l deprenyl) use with illicit drug (d-MA) use. MA enantiomers (l-, d-), amphetamine (AM) enantiomers (l-, d-), l-deprenyl, and desmethylselegiline (DMS) enantiomers (l-, d-) were simultaneously separated and detected with clear resolution. L deprenyl and its metabolites, l-MA, l-AM, and l-DMS, were detected in rat urine sample collected up to 24 h after oral administration of l-deprenyl (10 mg/kg), and the urinary l-AM/l-MA ratio was 2.45 +/- 0.55. This AM/MA ratio was significantly higher than the ratios obtained from rats administered with d-MA (5 mg/kg) and dl-MA (10 mg/kg). The d-AM/d-MA ratio was 0.98 +/- 0.25 for the d-MA treatment, and the d-AM/d-MA and l-AM/l-MA ratios were 0.72 +/- 0.24 and 0.71 +/- 0.21, respectively, for the dl-MA treatment. Analysis of human urine revealed that, unlike in rat urine, the MA content was much greater than the AM content, resulting in the AM/MA ratios being far lower in cases of healthy adult men treated with l-deprenyl (10 mg) and MA abusers. The AM/MA ratio from l-deprenyl users (0.33 +/- 0.03) was significantly higher than the ratio from MA abusers (0.20 +/- 0.12). Results indicate that although metabolic patterns of the drugs in rat and humans may be different, the AM/MA ratio from l-deprenyl use is significantly higher than the ratio from MA use in both rat and human urine. This ratio, however, cannot give conclusive proof of deprenyl or MA use in humans. The simultaneous chiral separation for all the metabolites of l-deprenyl and MA by CE analysis used in this study could provide rapid and simple discrimination between therapeutic drug use and illegal drug abuse. PMID- 10872570 TI - Solid-phase extraction of 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid from urine drug-testing specimens with the cerex polycrom-THC column. AB - Confirmation of drugs of abuse by gas chromatography-mass spectrometry (GC-MS) is the most time-consuming process used by drug-testing laboratories. Cost effectiveness and competitive turnaround times for testing results demand fast, efficient, and reliable extraction methods. We applied the Cerex Polycrom-THC solid-phase extraction (SPE) column to the extraction of 11-nor-delta9 tetrahydrocannabinol carboxylic acid (9-THCA) from urine. This column uses an anion exchange divinyl-benzene copolymer, which requires no pH adjustment after hydrolysis of the THC-glucuronide with base, as is necessary with many other SPE columns. With urine, no preconditioning of the SPE column was necessary. After extraction, trimethylsilation derivatization was performed with N-methyl-N (trimethylsilyl) trifluoroacetamide. This further improved efficiency because no heated incubation was required. A method correlation to an existing liquid-liquid extraction was performed. Analysis was on a Finnigan Voyager GC-MS with a 2.5-min run time per injection. Using 9-THCA-d9 deuterated internal standard, the assay was linear from 2 to 2000 ng/mL. Total precision at the 15-ng/mL cutoff concentration was 4.4%. The Cerex column was also evaluated for interference using two common drug-test adulterants, Klear and Urine Luck. Considerably less interference was observed when compared to an existing liquid-liquid method. PMID- 10872571 TI - The use of low-resolution FT-IR spectrometry for the analysis of alcohols in breath. AB - Fast and reliable diagnostic methods are needed for detection or exclusion of industrial solvents as a cause of intoxication. Analyzing human breath reveals the presence of any volatile substance. A portable Fourier transform infrared (FT IR) multicomponent point-of-care analyzer was developed for exhaled breath. The analyzer proved to be accurate and precise in laboratory tests for simultaneous measurement of methanol and ethanol in water. Ethanol, in addition to normal contents of breath, was simultaneously analyzed in human experiments, and the results correlated well with blood samples. FT-IR method has a traceable calibration to physical properties of the analyte. The measured spectra can also be saved and analyzed later. Breath analysis with FT-IR is fast and easy, and no preparation of the sample is needed. PMID- 10872572 TI - Simple and rapid determination of amphetamine, methamphetamine, and their methylenedioxy derivatives in urine by automated in-tube solid-phase microextraction coupled with liquid chromatography-electrospray ionization mass spectrometry. AB - A simple and rapid method for the determination of amphetamine, methamphetamine, and their 3,4-methylenedioxy derivatives in urine samples was developed using automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS). In-tube SPME is an extraction technique for organic compounds in aqueous samples in which analytes are extracted from the sample directly into an open tubular capillary by repeated draw/eject cycles of sample solution. LC-MS analyses of stimulants were initially performed by liquid injection onto an LC column to determine spectra. Five stimulants tested in this study gave very simple ESI mass spectra, and strong signals corresponding to [M+H]+ were observed for all stimulants. The stimulants were well separated with a Supelcosil LC-CN column using acetonitrile/50mM ammonium acetate (15:85) as a mobile phase. In order to optimize the extraction of stimulants, several in-tube SPME parameters were examined. The optimum extraction conditions were 15 draw/eject cycles of 35 microL of sample in 50mM Tris-HCI (pH 8.5) at a flow rate of 100 microL/min using an Omegawax 250 capillary column. The stimulants extracted by the capillary were easily desorbed by mobile phase flow, and carryover of stimulants was not observed. Using in-tube SPME-LC-ESI-MS with selected ion monitoring, the calibration curves of stimulants were linear in the range from 2 to 100 ng/mL with correlation coefficients above 0.9985 (n = 18) and detection limits (S/N = 3) of 0.38-0.82 ng/mL. This method was successfully applied to the analysis of human urine samples without interference peaks. The recoveries of stimulants spiked into urine samples were above 81%. PMID- 10872573 TI - A new spectrophotometric method for the toxicological diagnosis of cyanide poisoning. AB - A spectrophotometric method for the determination of hydrogen cyanide in biological fluids based on the release of cyanide ion by the addition of a strong acid and its subsequent specific reaction with hydroxocobalamin to give cyanocobalamin is proposed. The release of cyanide ion is accelerated by aeration with a stream of an inert gas (nitrogen) that carries it into the hydroxocobalamin solution. Although the in vitro reaction develops to completion within 20 min, reproducible quantitation in biological media takes 45 min. The cyanocobalamin formed is quantitated by second-derivative visible spectrophotometry from the absorbance difference between 333 and 361 nm, the measured signal being proportional to the cyanide ion concentration in the sample. PMID- 10872574 TI - Blood concentrations of amitriptyline and its metabolite in rats after acute oral administration of amitriptyline. AB - Amitriptyline (AMT), a tricyclic antidepressant that is a dibenzocycloheptadine derivative, is frequently used. However, the case reports of AMT-related fatalities are increased, nowadays, due to the low levels of toxic and fatal concentration in blood. So, this study was carried out to determine the concentrations of AMT and its demethylated metabolite, nortriptyline (NTR), after acute single oral administration of AMT in rats. Blood samples were collected five times from the ophthalmic venous plexus at 0, 1, 2, 4, and 8 h after acute single oral administration of AMT in toxic doses of 10 (Group I) or 20 mg/kg (Group II), and the concentrations of AMT and NTR and the mean ratios of AMT to NTR (AMT/NTR) in the blood were periodically determined at designated times. The blood concentrations of AMT and NTR were identified and quantitated by gas chromatography with thermionic specific detection and gas chromatography-mass spectrometry after solid-phase extraction with a Clean Screen DAU column. The peak blood concentrations of AMT and NTR in Group I were 0.34 and 0.28 microg/mL, respectively, and those of AMT and NTR in Group II were 0.59 and 0.43 microg/mL, respectively, and were reached at 1 h after single oral administration. PMID- 10872575 TI - Chemical derivatization and the selection of deuterated internal standard for quantitative determination--methamphetamine example. AB - Use of an isotopic analogue of the analyte as the internal standard in a quantitative gas chromatography-mass spectrometry targeted-compound-analysis protocol is often hindered by the availability of an adequate number (typically three for the drug/metabolite and two for the isotopic analogue) of sufficiently high mass ions that can be attributed to each member of the pair and are sufficiently free of interference by the contribution from the other component of the pair, a phenomenon termed "cross-contribution". Methamphetamine (MA) is selected as the exemplar compound to examine the effectiveness in using different chemical derivatization routes to produce derivatized analyte-isotopic analogue pairs that can generate more favorable mass spectrometric data to meet this analytical requirement. Trimethylsilyl-, trichloroacetyl-, and pentafluoropropionyl-derivatization and MA-d5, MA-d8, and MA-d9 are studied. Data resulting from this study indicate that the number of ion pairs suitable for quantitation and the degree of cross-contribution of these ions vary significantly. These data empirically demonstrate that derivatization methods play a significant role in deciding which deuterated analogue of the analyte provides the most suitable ion pairs that cause the least cross-contribution. The most suitable internal standard varies with the derivatization route adapted for an analytical protocol. PMID- 10872576 TI - Development of analytical methods for the detection of metaraminol in the horse. AB - Aramine (metaraminol bitartrate) has been found in the possession of horse trainers and veterinarians who have been investigated for possible inappropriate drug administration to racing horses. Metaraminol (3 hydroxyphenylisopropanolamine) is a sympathomimetic amine that directly and indirectly affects adrenergic receptors, with alpha effects being predominant. Because it has the potential to affect the performance of a racing horse, its use is prohibited. In the present study, methods for the detection of metaraminol were developed. Metaraminol was found to be extracted with poor recovery (< 50%) from aqueous solutions by routine basic extraction or cation exchange/reversed phase solid-phase extraction techniques. However, an extractive acetylation method gave good (> 90%) recovery of metaraminol from aqueous samples. Sequential urine samples collected from horses administered metaraminol intramuscularly at 0.02, 0.10, and 0.23 mg/kg were extracted by the developed extractive acetylation procedure and analyzed by gas chromatography-mass spectrometry (GC-MS) in full scan and selected ion monitoring modes. Norphenylephrine was used as an internal standard for quantitative analysis. The maximum concentration of metaraminol occurred between 1 and 2 h postadministration. Metaraminol was detected in the 0.23 mg/kg administration urine for 24 h postadministration. Metaraminol was detected for the 0.10 and 0.02 mg/kg doses for approximately 8 h postadministration. No apparent biotransformation products were observed in a reaction mixture of metaraminol and horse liver microsomal reaction mixture. Comparison of gas chromatograms of the extracts of the postadministration urine samples with those of the pre-administration samples failed to reveal any exogenous compound other than metaraminol. PMID- 10872577 TI - An automated and simultaneous solid-phase extraction of delta 9 tetrahydrocannabinol and 11-nor-9-carboxy-delta 9-tetrahydrocannabinol from whole blood using the Zymark RapidTrace with confirmation and quantitation by GC-EI-MS. AB - A sensitive, reliable, and automated solid-phase extraction (SPE) method was developed for the simultaneous extraction, confirmation, and quantitation of delta9-tetrahydrocannabinol (THC) and 11-nor-9-carboxy-delta9 tetrahydrocannabinol (THCCOOH) from whole blood. The extraction was performed on the Zymark RapidTrace SPE Workstation with a reduced solvent volume SPE copolymer cartridge. Quantitative analysis was completed on a benchtop gas chromatograph mass spectrometer using electron ionization mode with selected ion monitoring of three ions for each analyte. The limits of quantitation for THC and THCCOOH were 2 ng/mL and 1 ng/mL, respectively. The limits of detection for THC and THCCOOH were 1.6 ng/mL and 0.8 ng/mL, respectively. Extensive method validation is presented including extraction recoveries, within-run precision, day-to-day precision, linearity, and carryover. This procedure is used routinely in the laboratory on blood samples screened positive for cannabinoids. PMID- 10872578 TI - A sensitive immunoassay for flunitrazepam and metabolites. AB - The objective of this study was to develop an immunoassay that would be capable of detecting flunitrazepam and/or cross-reacting metabolites in urine and comparing the results with those obtained by gas chromatography-mass spectrometry. Doses of Rohypnol varying between 0.5 and 4 mg were given to volunteers, and urine was collected for up to two weeks postingestion. These samples were analyzed by an ELISA that was developed using an antibody raised to flunitrazepam and a drug-enzyme conjugate prepared by attaching 7 aminoflunitrazepam to horseradish peroxidase. Significant levels of flunitrazepam and/or cross-reacting metabolites were detected in urine for up to one week after ingestion. The immunoassay is selective with only diazepam cross-reacting at a level of 1000 microg/L. PMID- 10872579 TI - Quetiapine (Seroquel) concentrations in seven postmortem cases. AB - Quetiapine is a new antipsychotic drug that has been available in the United States since September 1997. It belongs to a new chemical class of drugs called the dibenzothiazepine derivatives and is easily detected with a basic drug screen. The Los Angeles County Department of Coroner Toxicology Laboratory has encountered quetiapine in seven postmortem cases. Tissue distributions were determined in each of the seven cases. The analysis of quetiapine from postmortem specimens consisted of an n-butylchloride basic extraction with presumptive identification and quantitation on a gas chromatograph-nitrogen-phosphorus detector. Linearity was achieved from 0.10 to 3.0 mg/L with a limit of quantitation of 0.10 mg/L. Confirmation of quetiapine was performed on a gas chromatograph-mass spectrometer by comparison with a pure analytical standard. The tissue distribution of quetiapine was as follows: heart blood present, but less than (+<) 0.10-49 mg/L (seven cases); femoral blood +< 0.10-1.4 mg/L (five cases); liver +< 0.10-112 mg/kg (five cases); spleen 4.0 mg/kg (one case); urine 0-3.0 mg/L (two cases); bile 0.60-7.5 mg/L (three cases); and gastric contents +< 0.01-18 mg total (five cases). To our knowledge, this is the first report of the presence of quetiapine in postmortem specimens. PMID- 10872580 TI - A fatality due to injection of tiletamine and zolazepam. AB - A 22-year-old male with more than 28 needle marks on his right arm was found dead. First, he was suspected as a drug abuser. Blood, urine, spleen, and injection-site tissue was collected during autopsy. The blood and urine specimens were screened for drugs. Immunoassay studies did not show any illegal drugs. However, two unidentified peaks were isolated in both of these biological fluids by routine gas chromatography-flame-ionization detection (GC-FID) and thermionic specific detection. Additional gas chromatography-mass spectrometry analysis determined these two peaks to be tiletamine and zolazepam. These two agents are used in combination as veterinary anesthesia. The concentrations of these drugs in blood were quantitated by GC-FID and found to be 0.85 mg/L of tiletamine and 3.3 mg/L of zolazepam. In urine, tiletamine and its metabolite, 2-(ethylamino)-2 (2-thionyl) cyclohexanol, were identified to be present along with zolazepam. The concentrations of tiletamine and zolazepam in spleen were revealed to be 0.92 and 3.5 mg/kg, respectively. Injection-site tissue concentrations were determined to be 25.1 mg/kg tiletamine and 23.3 mg/kg for zolazepam. The cause of death in this case was determined to be due to the multiple drug intoxication of tiletamine and zolazepam. PMID- 10872581 TI - Polymer displacement/shielding in protein chromatography. AB - An overview of different applications of polymer interactions with ion-exchange and dye-affinity chromatographic matrices is presented here. The strength of interaction between the ligand and the polymer plays a crucial role in deciding the mode of chromatographic application. Charged, non-ionic and thermosensitive polymers such as poly(ethylene imine), poly(N-vinyl pyrrolidone) and poly(vinyl caprolactam) respectively, show different degrees of interaction with the dye molecules in dye ligand chromatography. Polymers, with their ability of multipoint and hence strong attachment to the chromatographic matrices, were used as efficient displacers in displacement chromatography. The polymer displacement resulted in better recoveries and sharper elution profiles than traditional salt elutions. The globule-coil transition of the thermosensitive reversible soluble insoluble polymer, poly(vinyl caprolactam), can be exploited in dye-affinity columns for the temperature induced displacement of the bound protein. In another situation, prior to the column chromatography of crude protein extract, polymers formed complexes with the dye matrix and "shielded" the column. The polymer shielding decreased the nonspecific interactions without affecting the specific interactions of the target protein to the dye matrix. PMID- 10872582 TI - High-throughput single-strand conformation polymorphism analysis by capillary electrophoresis. AB - Mutation detection plays a great role in genetic and medical research and clinical diagnosis of inherited diseases and particular cancers. Single-strand conformation polymorphism (SSCP) analysis is one of the most popular methods for detection of mutations. Recently, automated capillary electrophoresis (CE) systems have been used in SSCP analysis instead of conventional slab gel electrophoresis. SSCP analysis in combination with CE is a rapid, simple, sensitive and high-throughput mutation screening tool, and has been successfully applied for mutation detection involving human tumor suppressor genes, oncogenes and disease-causing genes. The new technique has a great potential for mutation screening of large numbers of samples in clinical diagnosis. This review discusses basic issues about the methodology of SSCP analysis based on CE and summarizes several key applications. PMID- 10872583 TI - Gemfibrozil and its oxidative metabolites: quantification of aglycones, acyl glucuronides, and covalent adducts in samples from preclinical and clinical kinetic studies. AB - A gradient reversed-phase HPLC analysis for the direct measurement of gemfibrozil (GEM) and four oxidative metabolites in plasma and urine of humans and in tissue homogenates of rats was developed. The corresponding acyl glucuronides and the covalently bound protein adducts (in protein precipitates) were determined after liberation from the respective conjugates via alkaline hydrolysis. The limits of detection for the covalent adducts in human plasma are: 10 ng ml(-1) (GEM), 20 ng ml(-1) (M1), 0.5 ng ml(-1) (M2, M4), and 5 ng ml(-1) (M3). The method was validated with respect to selectivity, recovery, linearity, precision, and accuracy. It has been applied to the analysis of preclinical and clinical studies. Pharmacokinetic profiles of gemfibrozil, its metabolites, and covalent adducts in human plasma and rat tissue homogenates are given. PMID- 10872584 TI - Analysis of hydroxylated and N-dealkylated metabolites of terfenadine in microsomal incubates by liquid chromatography--mass spectrometry. AB - This report describes an assay for the H(1)-receptor antagonist, terfenadine, and its two primary metabolites, terfenadine alcohol (TOH) and azacyclonol (AZ), using positive-ion, electrospray ionization-liquid chromatography-mass spectrometry. The assay was developed in support of kinetic studies of terfenadine oxidative metabolism in human liver and intestinal microsomes, which required quantification of incubate metabolites at low nanomolar concentrations. Terfenadine metabolites were extracted from basified microsomal incubates into methylene chloride. Reconstituted extracts were subject to liquid chromatographic separation on a cyano-reverse phase column. The [M+H]+ ions of terfenadine, terfenadine metabolites, and internal standard were monitored in the effluent by quadrupole mass spectrometry. The assay demonstrated linearity over an incubate concentration range of 5-250 and 12.5-1250 ng/ml for the metabolites and the parent drug, respectively. The respective limits of detection and quantitation for all three analytes were 1.5 and 5 ng/ml of microsomal incubate. Replicate analysis of quality control samples exhibited intra-day coefficients of variation ranging from 3.3% to 7.8% for the three analytes. The corresponding inter-day coefficients of variation ranged from 4.2% to 8.6%. The reproducibility and sensitivity of the assay, combined with the selectivity of mass spectrometric detection, should allow an accurate kinetic characterization of terfenadine oxidation mediated by the high affinity CYP3A enzymes in human liver and intestinal microsomes. PMID- 10872585 TI - Measurement of chloramphenicol by capillary zone electrophoresis following end column amperometric detection at a carbon fiber micro-disk array electrode. AB - Capillary zone electrophoresis was employed for the measurement of chloramphenicol using end-column amperometric detection with a carbon fiber micro disk array electrode, at a constant potential of -1.00 V vs. saturated calomel electrode. The effect of oxygen in the buffer has been investigated. It is found that when the area of the carbon fiber electrode is smaller than 1.1 mm2, the interference of oxygen can be overcome. In this procedure deoxygenation is not necessary. The effect of pH, the concentration of the buffer and the high separation voltage across the capillary on the migration time, electrophoretic peak current and separation efficiency has been studied. The optimum conditions of separation and detection are 8.4x10(-4) mol/l HOAc-3.2x10(-3) mol/l NaOAc for the buffer solution, 20 kV for the separation voltage, 5 kV and 5 s for the injection voltage and the injection time, respectively. The calibration plot was found to be linear in the range 5x10(-6) to 1x10(-3) mol/l and the limit of detection is 9.1x10(-7) mol/l or 1.4 fmol (S/N=2). The relative standard deviation is 1.1% for the migration time and 2.3% for the electrophoretic peak current. The method was applied to the determination of chloramphenicol in human serum. PMID- 10872586 TI - Simultaneous determination of dimethylamphetamine and its metabolites in rat hair by gas chromatography-mass spectrometry. AB - In order to study the disposition of dimethylamphetamine (DMAP) and its metabolites, DMAP N-oxide, methamphetamine (MA) and amphetamine (AP), from plasma to hair in rats, a simultaneous determination method for these compounds in biological samples using gas chromatography-mass spectrometry with selected ion monitoring (GC-MS-SIM) was developed. As DMAP N-oxide partially degrades to DMAP and MA during GC-MS analysis, it was necessary to avoid conditions which co extract the N-oxide in the sample preparation so as to assure no contribution of artifactual products from DMAP N-oxide in the detection of the other compounds. For confirmation of the satisfactory separation of DMAP N-oxide from the others, the internal standards used for quantification were labeled with different numbers of deuterium atoms. Determination of unchanged DMAP was performed without any derivatization, that of DMAP N-oxide was carried out after conversion into trifluoroacetyl-MA by reaction with trifluoroacetic anhydride, and MA and AP were quantified after trifluoroacetyl-derivatization. After intraperitoneal administration of DMAP HCI to pigmented hairy rats (5 mgkg(-1) day(-1), 10 days, n=3), concentrations of DMAP and its metabolites in urine, plasma and hair were measured by GC-MS-SIM. The area under the concentration versus time curves (AUCs) of DMAP, DMAP N-oxide, MA and AP in the plasma were 397.2+/-97.5, 279.7+/-68.3, 18.4+/-1.2 and 15.9+/-2.2 microg min ml(-1), while their concentrations in the hair newly grown for 4 weeks after administration were 4.82+/-0.67. 0.45+/-0.09, 3.25+/-0.36 and 0.89+/-0.05 ng mg(-1), respectively. This fact suggested that the incorporation tendency of DMAP N-oxide from plasma into hair was distinctly low in comparison with the other compounds. PMID- 10872587 TI - High-performance liquid chromatography-tandem electrospray mass spectrometry for the determination of lidocaine and its metabolites in human plasma and urine. AB - A sensitive, selective and accurate high-performance liquid chromatographic tandem mass spectrometric assay was developed and validated for the determination of lidocaine and its metabolites 2,6-dimethylaniline (2,6-xylidine), monoethylglycinexylidide and glycinexylidide in human plasma and urine. A simple sample preparation technique was used for plasma samples. The plasma samples were ultrafiltered after acidification with phosphoric acid and the ultrafiltrate was directly injected into the LC system. For urine samples, solid-phase extraction discs (C(18)) were used as sample preparation. The limit of quantification (LOQ) was improved by at least 10 times compared to the methods described in the literature. The LOQ was in the range 1.6-5 nmol/l for the studied compounds in plasma samples. PMID- 10872588 TI - Simultaneous separation of atovaquone, proguanil and its metabolites on a mixed mode high-performance liquid chromatographic column. AB - An isocratic high-performance liquid chromatographic (HPLC) method for simultaneous separation of the components in the antimalarial combination drug Malarone with UV detection is described. An HPLC system using a mixed mode column composed of 50% C(18) phase and 50% strong cation-exchanger has been optimised for the simultaneous separation of atovaquone, proguanil and its two main metabolites. The mobile phase was optimised for factors such as pH, counter ion concentration and acetonitrile. Elimination of interferences from other antimalarial drugs was achieved by adding sodium perchlorate to the mobile phase. With a mobile phase of acetonitrile-phosphate buffer (60:40, v/v) pH 6.8, 50.7 mmol l(-1) K+ and 10 mmol l(-1) Na x ClO4, separation was achieved within a run time shorter than 17 min. PMID- 10872589 TI - Amounts and variation in grapefruit juice of the main components causing grapefruit-drug interaction. AB - A method for the determination of three furocoumarins containing two new chemicals (GF-I-1 and GF-I-4) in commercially available grapefruit juice and grapefruit itself was developed using high-performance liquid chromatography (HPLC). These components isolated from grapefruit juice have 5 geranyloxyfurocoumarin dimer structures showing extremely high affinities for a form of cytochrome P450 (CYP3A4). Considerable differences were observed on the contents among commercial brands and also batches. The contents were determined to be 321.4+/-95.2 ng/ml GF-I-1, 5641.2+/-1538.1 ng/ml GF-I-2 and 296.3+/-84.9 ng/ml GF-I-4 in twenty-eight white grapefruit juices. These chemicals were not detected in beverages from orange, apple, grape and tangerine, except that trace amount of GF-I-2 and GF-I-4 were found in lemon juice. The average levels of these furocoumarins were lower in the juice from red grapefruit than a white one. The highest level of these components were found in the fruit meat. PMID- 10872590 TI - Identification of the principal circulating metabolite of a synthetic 5,4' diaminoflavone (NSC 686288), an antitumor agent, in the rat. AB - During the course of our study to develop analytical methodology for quantitating the investigative antitumor agent 5-amino-2-(4-amino-3-fluorophenyl)-6,8-difluoro 7-methyl-4H-1-benzopyran -4-one (DAF; NSC 686288) in plasma, a significant concentration of a metabolite was observed in a post-dosed rat. The results of electron-ionization (EI) mass spectrometric analysis of the metabolite suggested that N-acetylation had occurred, but, interestingly, that only one of the compound's two primary amino groups had been transformed. Comparing the mass spectra and gas chromatographic retention times of a mono-acetylated sample of DAF and that of the metabolite showed both to be the same. A retro-Diels-Alder (RDA) fragmentation of the B ring of DAF results in formation of two abundant product ions, each retaining one of the amino groups. The EI mass spectrum of mono-N-acetamido-d3 DAF shows loss of ketene-d2, leading to formation of an -NHD group. The ensuing RDA fragmentation easily identifies which of the two product ions contains the deuterium, thereby allowing us to assign the site of N acetylation as the amino group on ring C (the 4' position) of DAF. PMID- 10872591 TI - Use of short high-performance liquid chromatography columns and tandem-mass spectrometry for the rapid analysis of a prostaglandin analog, fluprostenol, in rat plasma. AB - A short reversed-phase HPLC column and a tandem mass spectrometer were used to develop a stable-isotope-dilution assay for the rapid and sensitive analysis of fluprostenol, a prostaglandin analog, in rat plasma. A Waters Symmetry ODS column (2.1x10 mm) afforded rapid isocratic elution of fluprostenol (t(R)=40 s) but still provided a relatively large k' value of 4. The use of tandem mass spectrometry allowed the interference-free detection of fluprostenol under the rapid elution conditions, with a limit of quantitation of 25 pg ml(-1) fluprostenol, using 0.2 ml plasma sample volumes. The method was linear over three orders of magnitude, yielded accurate and precise results and allowed the pharmacokinetic profile of fluprostenol to be defined following intravenous administration in rats. PMID- 10872592 TI - Determination of enantiomeric amphetamines as metabolites of illicit amphetamines and selegiline in urine by capillary electrophoresis using modified beta cyclodextrin. AB - The determination of enantiomeric amphetamine and methamphetamine in urine samples is important in order to distinguish use of the prescription drug selegiline (metabolized to R(-)-A and R(-)-MA) from the illicit use of S(+)-A and S(+)-MA. For the analysis of enantiomeric amphetamine (A) and methamphetamine (MA) in biological samples, the optimization of analytical condition was performed by capillary electrophoresis using chiral selectors including beta cyclodextrin, carboxymethyl-beta-cyclodextrin and 2-hydroxypropyl-beta cyclodextrin. We have examined the factors to obtain the best chiral resolutions, separation efficiency and sensitivity, and wide concentration linearity. Optimum resolutions were achieved using 100 mM phosphate buffer, pH 2.5, containing 10 mM of carboxymethyl-beta-cyclodextrin. This method was applied for the quantitative determination of enantiomeric amphetamine and methamphetamine in urine samples obtained from patients taking illicit amphetamines or from rats and patients taking selegiline. Acceptable quantitative results in terms of resolution, precision, sensitivity and linearity were obtained from the real urine samples containing wide-ranging concentrations of A and MA by using two concentrations of internal standards, alpha(+)- (1 microg/ml) and beta-phenylethylamine (50 microg/ml). PMID- 10872593 TI - Optimization and performance of a rapid gas chromatography-mass spectrometry analysis for methylmalonic acid determination in serum and plasma. AB - We have developed a rapid and sensitive GC-MS assay for methylmalonic acid determination in serum and plasma utilizing an anion exchange solid-phase extraction and trimethylsilyl derivatization. Each step of the procedure was optimized by the experimental design methods to assure the assay reliable performance. The limit of detection and limit of quantitation were 0.025 and 0.1 micromol/l. The total coefficient of variation for the method was 9.8, 4.4, and 4.6% at the concentration of 0.2, 3.1, and 6.2 micromol/l methylmalonic acid concentration, respectively. The assay was linear up to 9.0 micromol/l, and showed good correlation with a reference method. The method has proven to be reliable in routine production, producing clean chromatography, unique ion fragments, and consistent ion mass ratio. PMID- 10872594 TI - Analysis of gamma radiation-induced damage to plasmid DNA using dynamic size sieving capillary electrophoresis. AB - Bacterial plasmids and the chromosomal DNA of many organisms adopt naturally the negatively supercoiled conformation. Therefore, the irradiation of such plasmids could be used to model conformational changes of chromosomal DNA associated with externally-induced damage. We have applied dynamic size-sieving capillary electrophoresis (CE) to monitor the damage of three DNA plasmids, over an unprecedented base pair (bp) size range (2870-27 500 bp), upon exposure to gamma radiation (20-400 Gy). Predominantly, CE with UV absorbance detection in the absence of DNA intercalating dyes was employed to preclude undesirable, induced plasmid conformational changes. Plasmid samples and their enzymatic digestion products were analyzed using both CE and slab gel electrophoresis (SGE) in order to verify the conformation of sample components. Relative to SGE, CE analyses revealed more fine structural features of plasmid degradation. PMID- 10872595 TI - Development, validation and application of assays to quantify metrifonate and 2,2 dichlorovinyl dimethylphosphate in human body fluids. AB - Gas chromatographic procedures [GC with electron-capture detection (ECD) and GC MS] for the quantitative analysis of metrifonate and its active metabolite 2,2 dichlorovinyl dimethylphosphate (DDVP) in human blood and urine were developed, validated, and applied to the analysis of clinical study samples. Analysis of metrifonate involved extraction of acidified blood with ethyl acetate followed by solid-phase clean-up of the organic extract. Acidified urine was extracted with dichloromethane and the residue of evaporated organic phase was reconstituted in toluene. ECD and diethyl analogue of metrifonate internal standard (I.S.) were used for quantitation of metrifonate. The metrifonate lower limit of quantitation (LOQ) was 10.0 microg/l. The DDVP metabolite was chromatographed separately after cyclohexane extraction of acidified blood and urine using d6-DDVP I.S. and MS detection. The LOQ of DDVP was 1 microg/l. Stability studies have confirmed that the matrix should be acidified prior to storage at -20 degrees C or -80 degrees C to inhibit chemical and enzymatic degradation of the analytes and to avoid overestimation of DDVP concentrations. Metrifonate was found to be stable in acidified human blood after 20 months of storage at -20 degrees C and after 23 months of storage at -80 degrees C. Under these conditions DDVP was found to be stable after 12 months of storage. Both assay procedures were cross-validated by different world-wide laboratories and found to be accurate and robust during analyses of clinical study samples. PMID- 10872596 TI - Stable isotope dilution analysis of human urinary metabolites of 17alpha methyltestosterone. AB - A method based on gas chromatography-mass spectrometry-selected-ion monitoring was developed to measure the main metabolites of 17alpha-methyltestosterone, 17alpha-methyl-5alpha-androstan-3alpha,17beta-di ol and 17alpha-methyl-5beta androstan-3alpha,17beta-dio l, in human urine. 17alpha-Methyl-[(2)H3]-5alpha androstan-3alpha,1 7beta-diol and 17alpha-methyl-[(2)H3]-5beta-androstan 3alpha,17 beta-diol were used as internal standards. The methods involved purification using a Sep-Pak C(18) cartridge, hydrolysis by beta-glucuronidase from Ampullaria and derivatization with N-methyl-N-trimethylsilyl trifluoroacetamide/dithioerythriol/ammon ium iodide. Quantitation was achieved by selected-ion monitoring of the characteristic fragment ions ([(M+H)-2xTMSOH]+) of the di-TMS derivatives on the chemical ionization mode. The method provides a specific, sensitive and reliable technique to determine the urine levels of 17alpha-methyl-5alpha-androstan-3alpha,17beta-di ol and 17alpha-methyl-5beta androstan-3alpha,17beta-dio l, and can be applied to pharmacokinetic studies of 17alpha-methyltestosterone. PMID- 10872597 TI - Convenient method of threonine, methionine and their related amino compounds by high-performance liquid chromatography and its application to rumen fluid. AB - A high-performance liquid chromatographic procedure for the quantitative determination of cysteine (Cys), homocysteine (Hcys), methionine sulfoxide (MSO), methionine sulfone (MSO2), homoserine (Hser), glycine (Gly), threonine (Thr), 2 aminobutyric acid (2AB), methionine (Met), cystathionine (Cysta) and its application to rumen fluid are described. The samples containing Thr, Met and other related amino compounds were derivatized with 9-fluorenylmethyl chloroformate. The separation of compounds was accomplished with a methanol gradient in 25 mM sodium citrate buffer (obtaining pH 6.40 and 3.80 by addition of 25 mM citric acid). All derivatized compounds were separated on a Mightysil RP 18 GP (150x4.6 mm I.D., 5 microm particle size) column. All analytes were detected at 265 nm with UV detection. The limits of detection (microM) (S/N ratio, 3:1) and quantification (microM) (S/N ratio, 10:1) of Cys, Hcys, MSO, MSO2, Hser, Gly, Thr, 2AB, Met and Cysta were 0.50 and 1.68; 1.76 and 5.85; 0.85 and 2.88; 0.92 and 3.09; 1.04 and 3.52; 0.76 and 2.52; 0.65 and 2.18; 0.39 and 1.36; 0.31 and 1.03; 0.17 and 0.58, respectively. The recoveries of all compounds in rumen fluid were 97.93-102.3% in the within-day study and 94.52-98.69% on different day (6 days) studies. The average contents (microM) of Cys, Gly, Thr, 2AB, Met and Cysta were 1.72, 45.6, 20.0, 4.3, 2.11 and 3.42 before morning feeding. The concentration of Thr, 2AB and Cysta in rumen fluid tended to increase with time after feeding whereas Met showed the opposite tendency. PMID- 10872598 TI - Validated high-performance liquid chromatographic method for the determination of lamotrigine in human plasma. AB - A high-performance liquid chromatographic (HPLC) procedure for lamotrigine was developed and validated. Lamotrigine (LTG) and an internal standard were extracted from plasma using liquid-liquid extraction under alkaline conditions into an organic solvent. The method was linear in the range 0.78-46.95 micromol/l, with a mean coefficient of correlation (r)> or =0.99923. The limit of detection (LOD) and limit of quantification (LOQ) were 0.19 and 0.58 micromol/l, respectively. Within- and between-run precision studies demonstrated C.V.<3% at all tested concentrations. LTG median recovery was 86.14%. Antiepileptic drugs tested did not interfere with the assay. The method showed to be appropriate for monitoring LTG in plasma samples. PMID- 10872599 TI - Determination of grepafloxacin in plasma and urine by a simple and rapid high performance liquid chromatographic method. AB - A rapid, specific, sensitive and economical method has been developed and validated for the determination of grepafloxacin in human plasma and urine. The assay consisted of reversed-phase HPLC with UV detection. Plasma proteins were removed by a fast and efficient procedure that has eliminated the need for costly extraction and evaporation. For the urine samples, the only required sample preparation was dilution. Separation was achieved on a reversed-phase TSK gel column with an isocratic mobile system. The method had a quantification limit of 0.05 microg/ml in plasma and 0.5 microg/ml in urine. The coefficients of variation (C.V.) were less than 4% for within- and between-day analyses. The method was successfully applied to a pharmacokinetic study, and was proved to be simple, fast and reproducible. PMID- 10872600 TI - Purge-and-trap gas chromatographic determination of styrene in urine and blood. Application to exposed workers. AB - A simple purge-and-trap gas chromatographic method with flame ionization detection was developed for the determination of styrene in urine and blood. Styrene present in a 5 ml sample at room temperature was swept by helium at 40 ml/min for 11 min, trapped on a Tenax trap, desorbed by heating, cryofocused, and injected by flash heating into a DB-5 capillary GC column. The oven temperature program was from 80 degrees C, held for 8 min, to 120 degrees C at 5 degrees C/min, and then held for 2 min. The detector temperature was 250 degrees C. The calibration curves were linear in the range of 2.5-15 ppb styrene in urine and 25 150 ppb in blood. The detection limits calculated were 0.4 microg/l in urine and 0.6 microg/l in blood. The coefficients of variations within the day and day-to day were 3 and 3.1%, respectively, for 2.5 ppb of styrene in urine, and 1 and 1.6% for 25 ppb of styrene in blood. The results obtained from samples taken from workers exposed to styrene were reported. PMID- 10872601 TI - Simple method for determination of terbutaline plasma concentration by high performance liquid chromatography. AB - A method is described in which low nanomolar concentrations of terbutaline in plasma can be quantitated by use of a standard isocratic high-performance liquid chromatography system with electrochemical detection. Samples were prepared for injection by solid-phase extraction and preserved from degradation by addition of glutathione. Terbutaline and internal standard metaproterenol were resolved from plasma constituents on a single C(18) column by ion-pairing chromatography. The method is precise and accurate for measurement of freebase concentrations as low as 4.4 nmol/l (1 ng/ml). PMID- 10872602 TI - The role of amniotic fluid interleukin-6, and cell adhesion molecules, intercellular adhesion molecule-1 and leukocyte adhesion molecule-1, in intra amniotic infection. AB - PROBLEM: To determine amniotic fluid concentrations and correlations of interleukin-6 (IL-6), intercellular adhesion molecule-1 (ICAM-1), and leukocyte adhesion molecule-1 (LAM-1) in patients with and without intra-amniotic infection. METHOD OF STUDY: Fourteen specimens with intra-amniotic infection and 45 without intra-amniotic infection were studied. Intra-amniotic infection was defined as the presence of a positive amniotic fluid culture. Amniotic fluid IL 6, ICAM-1, and LAM-1 levels were determined by an enzyme-linked immunoassay, and normalized by amniotic fluid creatinine levels. RESULTS: Amniotic fluid concentrations of IL-6 and LAM-1 were significantly higher in patients with than without intra-amniotic infection. However, amniotic fluid ICAM-1 concentrations were not significantly different between two groups. Amniotic fluid IL-6, LAM-1, and ICAM-1 were positively correlated. CONCLUSIONS: Our data indicate that amniotic fluid IL-6 is significantly associated with an increased adhesion molecule expression in intra-amniotic infection. However, LAM-1 plays a more important role than ICAM-1 in intra-amniotic infection. PMID- 10872603 TI - Increased apoptosis in human amnion is associated with labor at term. AB - PROBLEM: To characterize whether increased apoptosis in human amnion was associated with labor at term. METHOD OF STUDY: Human amnion were obtained from term patients with vaginal delivery (n = 5) or who underwent elective Cesarean section (C/S) without labor (n = 5). Apoptosis was performed by the TUNEL (Terminal dUTP Nuclear End Labeling) assay. All nucleated cells stained with propidium iodide in the amnion epithelial cells were identified in red fluorescence. TUNEL positive apoptotic nuclei were identified in green fluorescence. Five random fields of each specimen were blindly counted by investigators. The percentage of apoptotic nuclei of total nuclei (apoptotic index) was calculated and compared between the two groups (25 microscopic fields for each group, respectively). RESULTS: Patients with term labor had a significantly higher mean apoptotic index in amnion epithelial cells than that with elective C/S without labor (27.3 +/- 4.1% versus 3.6 +/- 1.6%, P < 0.001). CONCLUSIONS: Our data indicate that apoptosis in human amnion is significantly increased and associated with labor at term. PMID- 10872604 TI - Fetal endothelial cells express vascular cell adhesion molecule in the setting of chorioamnionitis. AB - PROBLEM: In intrauterine infection, inflammatory mediators may be released into the fetal circulation prior to fetal infection. We hypothesize that, in chorioamnionitis, inflammation alters fetal blood vessels. To test this, fetal endothelial cells were examined for vascular cell adhesion molecule (VCAM). METHOD OF STUDY: Umbilical cords (n = 9) from placentas with chorioamnionitis were immunostained for VCAM. Controls from preterm preeclamptic pregnancies (n = 7) without histologic inflammation were selected, and matched for gestational age and method of delivery. VCAM sections were reviewed by a pathologist blinded to clinical diagnoses. RESULTS: All endothelial cells from each of the nine cords from placentas with chorioamnionitis had strong VCAM staining. Two of nine samples also had acute cord vasculitis. No cord endothelial cells from preeclamptic placentas demonstrated similar VCAM staining (p < 0.01). CONCLUSION: Histologic chorioamnionitis was associated with VCAM expression of the umbilical cord vessels. In chorioamnionitis, inflammatory mediators may have entered the fetal circulation to activate endothelial cells. Intrauterine inflammation was not restricted to the chorioamnion, but also involved the fetal circulation. PMID- 10872605 TI - Intercellular adhesion molecule-1 (ICAM-1) in cervicovaginal fluid of women presenting with preterm labor: predictive value for preterm delivery. AB - PROBLEM: Clinically useful tests for the prediction and diagnosis of preterm labor and delivery remain to be established. We have hypothesized that soluble intercellular adhesion molecule-1 (sICAM-1) in the cervicovaginal fluid of women with preterm labor may be a useful diagnostic tool. METHOD OF STUDY: The cervicovaginal fluid of 103 women between 24(0) and 33(6) weeks gestation with preterm contractions and intact membranes was assayed for sICAM-1. RESULTS: Elevated sICAM-1 concentrations predicted short intervals to delivery (area under receiver operator characteristic (ROC) curves, 0.70-0.72 for delivery within 3, 7 and 10 days), with high specificity. Characteristics for delivery within 3 days at a 3 ng/mL threshold for a positive test were sensitivity 33.3%, specificity 98.9%, and positive and negative predictive values of 75.0% and 93.9%, respectively. Predictive ability was independent of and complementary to that of fetal fibronectin (fFN). CONCLUSIONS: Measurement of sICAM-1 in cervicovaginal fluid has potential as a predictor of preterm delivery in women with symptoms of preterm labor, particularly in conjunction with fFN testing. PMID- 10872606 TI - Interleukin 8 expression in human myometrium: changes in relation to labor onset and with gestational age. AB - PROBLEM: Preterm labor remains the major cause of perinatal mortality and morbidity in normally formed babies. The mechanisms involved in the onset of preterm labor are poorly understood, mainly because the mechanisms initiating term labor remain ill-defined. METHOD OF STUDY: Human myometrial samples were collected at cesarean delivery from preterm (26-36 weeks gestation) and term (37 41 weeks) women. Women at term were either non-laboring or laboring. The expressions of interleukin-8 (IL-8) mRNA and protein were measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. RESULTS: The expression of both IL-8 mRNA and protein significantly increased in the term laboring group, compared with either the term non-laboring or preterm groups. Levels of IL-8 expression did not alter with advancing gestational age. CONCLUSIONS: The increased expression of IL-8 in laboring myometria at term supports the hypothesis that up-regulation of IL-8 has a role in the initiation of labor in association with an influx of neutrophils and the release of their collagenolytic enzymes into uterine tissues. PMID- 10872607 TI - Production of protein hormones by cultured trophoblast cells isolated from term and early placentae. AB - PROBLEM: To compare the capacity of de novo hormone synthesis by cultured trophoblast cells isolated from early and term placenta as cytotrophoblast, and to determine the ability of these cells to proliferate in culture. METHOD OF STUDY: Cytotrophoblast cells were isolated from term (TP, 38-42 weeks) and early placentae (EP, 8-13 weeks) by enzymatic digestion and subsequent purification on a percoll gradient. The net synthesis of the hormones human placental lactogen (hPL) and human chorionic gonadotropin (hCG) was determined as the release during culture + cell content after culture - cell content before culture. Proliferation was determined using a dedicated colorimetric reagent (CellTiter 96). RESULTS: Using a percoll gradient we were able to isolate three cell bands with densities of 1.051, 1.058, and 1.063 g/mL, which were predominantly cytotrophoblast cells as shown by immunocytochemical analysis. The cytotrophoblast cells with the highest density (1.063 g/mL) were used because they were found to release the highest amount of hormones and have shown the lowest rate of cell death after 6 days in culture. Both hCG and hPL showed different patterns of release during the first 2-3 days of culture between TP and EP. While the release by EP cytotrophoblast cells continued during 6 days of culture (n = 4), the concentrations for TP cytotrophoblast (n = 4) reached a plateau between 4 and 6 days. Net de novo synthesis calculated for 3 x 10(4) TP trophoblast cells cultured for 6 days (mean +/- SD, n = 4) was 8.65 +/- 9.05 mU for hCG and 0.95 +/ 0.45 ng for hPL. For EP, it was 395.5 +/- 265.5 mU for hCG and 148.8 +/- 84.2 ng for hPL. Net synthesis of hCG was > 10-fold (TP) and > 70-fold (EP) higher than the initial cell content. While at term, hPL synthesis was only a fraction of the initial cell content, production by EP cytotrophoblast was 106 times the initial cell content. The extent of cell death after 6 days in culture was significantly (P < 0.02) higher for term (30-40%) than for early trophoblast (10-20%). Using a proliferation detection agent during the first 3 days of culture with first trimester cytotrophoblast cells, we did not find any changes in the proliferative activity. CONCLUSIONS: There are differences in the functional activity between trophoblast cells obtained from first and third trimester. The in vitro findings are difficult to reconcile with the different patterns of plasma concentrations of the two hormones observed in vitro during the course of pregnancy. PMID- 10872608 TI - Transforming growth factor beta isoforms production by human peritoneal mesothelial cells after exposure to hypoxia. AB - PROBLEM: Although human mesothelial cells (HMC) line nearly the entire abdominal cavity, little is known about their role in adhesion formation. This study determines the effect of hypoxia and transforming growth factor (TGF)-beta1 on the ability of HMC to produce TGF-beta1-3, which have been implicated as mediators of the healing process. METHOD OF STUDY: HMC were cultured under normal and hypoxic conditions, and treated with and without TGF-beta1 for 24 hr. RNA from each group was subjected to multiplex reverse transcription-polymerase chain reaction to quantitate TGF-beta1-3 mRNA levels. RESULTS: Hypoxia resulted in 2- and 3.3-fold increase, while TGF-beta1 treatment resulted in 1.4- and 1.2-fold increase (normoxia) and 0- and 4.8-fold increase (hypoxia) in TGF-beta1 and TGF beta2 mRNA levels, respectively. There was no detectable TGF-beta3 mRNA in HMC before or after treatments. CONCLUSION: TGF-beta1 treatment under hypoxia further extenuates endogenous TGF-beta2 but blocks TGF-beta1 production, thereby decreasing the TGF-beta1/TGF-beta2 ratio, which may result in the reduction of scarring and fibrosis. PMID- 10872609 TI - Does leptin exhibit cytokine-like properties in tissues of pregnancy? AB - PROBLEM: To determine whether leptin exhibits cytokine-like properties in gestational tissues in light of its homologies with the class I family of cytokines. METHOD OF STUDY: WISH and JEG3 cells, and amnion and choriodecidua explants, were treated inflammatory modulators (interleukin-1beta [IL-1beta], tumor necrosis factor-alpha [TNF-alpha] and bacterial lipopolysaccharide [LPS]) and leptin production was measured by immunoassay. Other agents known to regulate adipocyte leptin production were also tested for comparative purposes. In addition, WISH cells, JAR cells and placental explants were treated with leptin to assess its effects on production of IL-8, IL-6 and prostaglandin E2 (PGE2). RESULTS: Leptin production by all cells and tissues studied was unaffected by treatment with IL-1beta (2.5 ng/mL), TNF-alpha (25 ng/mL) and LPS (2.5 microg/mL). Dexamethasone stimulated leptin production over two-fold by WISH and JEG3 cells, whereas insulin also stimulated a two-fold increase in leptin production in JEG3 cells. IL-6 production by JAR cells and placental explants was stimulated (two- to three-fold) by leptin (300 ng/mL). PGE2 production was unaffected. CONCLUSIONS: Leptin derived from gestational tissues is unlikely to play a role in inflammatory reactions within the placenta, but may regulate placental cytokine production. The physiological significance of amnion-derived leptin remains to be established. PMID- 10872610 TI - Altered expressions of VEGF mRNA splice variants during progression of uterine peritoneal adhesions in the rat. AB - PROBLEM: Postoperative pelvic adhesions contribute to infertility, pelvic pain, bowel obstruction, and difficult reoperative procedures. METHOD OF STUDY: In the present study, a rat uterine-peritoneal adhesion model was developed to study the progression of adhesion formation during a course of 7 days following pelvic surgery. The distal 1 cm of each uterine horn and its adjacent peritoneum was abraded by six scratches with a scalpel blade, producing punctate bleeding. The scratched portion of uterine horn and the peritoneum was then held with Vicryl 3 0 to promote adhesion. The uterine tissue and the portion of peritoneum, held with suture, were then excised from a group of four rats, each at 6, 12, 24, 48, 72 hr and 5 and 7 days following surgery. Total RNA was isolated from these tissues and the expression pattern of different splice variants of vascular endothelial growth factors (VEGF) was examined using relative abundance reverse transcriptase polymerase chain reaction (RA-RT-PCR) method. RESULTS: Three known splice variants of VEGF mRNA (VEGF120, VEGF164 and VEGF188), as well as an additional band (approximately 510 bp), were amplified from these tissues. The relative abundance of known VEGF isoforms demonstrated altered expression during adhesion progression. When compared with noninjured uterine tissues, VEGF120 and VEGF188 demonstrated up-regulation during early stages of adhesion formation, whereas VEGF164 rather demonstrated down-regulation 24 and 48 hr following surgery. CONCLUSIONS: The up-regulation of VEGF isoforms during the progression of uterine-peritoneal adhesion may be a compensatory mechanism regulating angiogenesis in order to provide nutrients and oxygen to the injured tissues. PMID- 10872611 TI - Acute intrauterine infection results in an imbalance between pro- and anti inflammatory cytokines in the pregnant rabbit. AB - PROBLEM: Intrauterine infection results in an increase in cytokines. This study compared the time courses for the pro- and anti-inflammatory cytokine responses in 33 pregnant rabbits at 70% gestation. Pro-inflammatory markers were activated nuclear factor-kappa B (NF-kappaB) in placenta and tumor necrosis factor-alpha (TNF-alpha) in amniotic fluid. These were compared to the anti-inflammatory cytokine, interleukin-1 receptor antagonist (IL-1ra), in placenta and uterus. METHOD OF STUDY: Does were endoscopically inoculated with Escherichia coli through their cervices and sacrificed at six intervals between 0 and 30 hr post inoculation. RESULTS: Activated NF-kappaB, determined by electromobility gel shift assay, increased significantly 16 hr after bacterial inoculation (P < or = 0.05). This was directly mirrored by TNF-alpha concentrations, determined by bioassay, in the amniotic fluid. However, IL-1ra levels, determined by enzyme linked immunosorbent assay, did not increase in response to infection. CONCLUSIONS: Intrauterine infection results in an imbalance between pro- and anti inflammatory cytokines that may potentiate infection-induced preterm delivery. PMID- 10872612 TI - Effect of Saireitoh on rabbit smooth muscle cell proliferation and experimental atherosclerosis. AB - Saireitoh is a traditional Chinese medicine that is often given to patients with nephrotic syndrome or glomerulonephritis. Studies have reported that Saireitoh stimulates intrinsic steroid secretion in rats and suppresses the proliferation of fibroblasts in vitro. We examined the effects of Saireitoh on vascular smooth muscle cell proliferation and migration in vitro and experimental atherosclerosis in vivo. Saireitoh rabbit serum obtained from New Zealand White rabbits which were given a diet containing 2% Saireitoh for 3 days significantly inhibited [3H] thymidine incorporation by smooth muscle cells, which were isolated from thoracic aorta explants of rabbits. The addition of 10% Saireitoh rabbit serum to a culture medium containing smooth muscle cells inhibited DNA synthesis by 50% as compared with a control culture to which 10% normal rabbit serum was added. We also found that the number of smooth muscle cells in the culture containing Saireitoh rabbit serum was decreased. When PDGF was used as a chemoattractant, we demonstrated that Saireitoh rabbit serum slightly inhibits the migration of smooth muscle cells. In in vivo experiments, Saireitoh did not suppress the development of atherosclerosis but tended to reduce the damage. We concluded that although Saireitoh inhibited the proliferation of smooth muscle cells, the effect of prevention on the development of atherosclerosis is weak in the in vivo condition. PMID- 10872613 TI - Late results of acute medial necrosis in rabbit aorta. AB - To clarify whether acute medial necrosis of the aorta induces aneurysms and intimal thickening at a later stage, we first attempted to induce acute aortic medial necrosis in 47 normal rabbits by the administration of Russell's viper venom intraperitoneally and of angiotensin II intravenously as used in a previous study and then followed the rabbits for 1 and 2 months respectively. As a control, 18 adult normal rabbits were used. Six control and 20 treated rabbits were sacrificed after aortagraphy at the end of one month, while the remaining 12 control and 27 treated rabbits were sacrificed at the end of 2 months. We evaluated the aortic lesions by gross observations and both light and electron microscopic examinations. In addition, at the end of one month, aortagraphy was performed to measure the luminal diameter of the aorta of the 6 control and 20 treated rabbits. We macroscopically found the saccular lesions to be surrounded by small crater like lesions mainly at the thoracic aortas in 18 out of 47 treated rabbits. These lesions consisted of the necrosis and calcification of the aortic media and the destruction of the elastic fiber along with intimal thickening. However, no aneurysmal dilatation was found in the aortagraphy findings. We thus conclude that acute medial necrosis produced saccular and crater like lesions but these lesions were not confirmed by aortagraphy. PMID- 10872614 TI - Antioxidative effects of Choi-oki-to and its ability to inhibit the progression of atheroma in KHC rabbits. AB - Agents which inhibit the oxidative modification of low density lipoprotein (LDL) have been thought to be helpful in preventing the formation of atherosclerotic lesions; the so called "oxidation hypothesis". To test this hypothesis, we examined the antioxidative activities of 127 Kampo medicines in vitro and their inhibitory effects on the development of atheromatous plaque formation in KHC rabbits, a model of spontaneous familial hypercholesterolemia. Some of the 127 Kampo medicines showed scavenging or antioxidative effects equal to or stronger than those of probucol in vitro. Choi joki to, which had the strongest antioxidative effects on LDL in vitro, was chosen for a study in vivo. After 24 weeks, 1 g/kg of Choi joki to successfully inhibited the progression of atherosclerotic lesions in KHC rabbits (P < 0.01). Further investigations regarding the antioxidative effects of Kampo medicines are expected. PMID- 10872615 TI - Coronary atherosclerosis in youths in Kyushu Island, Japan: histological findings and stenosis. AB - We histologically examined the coronary arteries of 52 autopsied cases of the youths (3 to 39 years of age, mean 28.5 years) in Kyushu island, Japan, without clinical events of coronary artery diseases. The coronary artery specimens were taken from the proximal portions of the right coronary artery (Seg. 1), the left anterior descending artery (Seg. 6), and the macroscopically most stenotic region (ST). Atherosclerotic lesions were histologically classified into four types: concentric fibrous, eccentric fibrous, concentric lipid rich, or eccentric lipid rich type. The degrees of stenosis (< 25%, 25-50%, 50 75%, > 75%) were morphometrically evaluated. The majority of coronary arteries with under 50% stenosis were of the concentric fibrous type. Lipid-rich types of coronary atherosclerosis increased in the coronary arteries with over 50% stenosis and were observed in the Seg. 6 and ST, while 70% of Seg. 1 lesions with over 50% stenosis were of a fibrous type. Serum cholesterol levels of patients with a lipid rich type of coronary atherosclerosis were significantly higher than those with a fibrous type. These results suggested that the early stage of coronary atherosclerosis in Japanese youths is mainly of a concentric fibrous type, which later develops to a lipid rich type. Hypercholesterolemia would promote the progression of atherosclerosis. PMID- 10872616 TI - The Framingham Study: ITS 50-year legacy and future promise. AB - The Framingham Study was initiated in 1948 to investigate an epidemic of coronary disease in the USA, using a prospective epidemiological approach. Insights were provided into the prevalence, incidence, full clinical spectrum and predisposing factors. The major "risk factors" (a term coined by the Framingham Study) for coronary disease, stroke, peripheral artery disease and heart failure were identified and clinical misconceptions dispelled about isolated systolic hypertension, left ventricular hypertrophy, dyslipidemia, atrial fibrillation and glucose intolerance. Average values for blood lipids, blood pressure, body weight, glucose and fibrinogen were shown to be dangerously suboptimal and to have a continuous graded relationship to cardiovascular disease without critical values. Dyslipidemia, glucose intolerance and elevated fibrinogen were shown to have smaller hazard ratios in the elderly, but this was offset by a higher absolute risk. Diabetes was shown to operate more strongly in women, eliminating their advantage over men. Serum total cholesterol was shown to derive its atherogenic potential from its LDL component and also to reflect cholesterol being removed in the HDL fraction. The total/HDL-cholesterol ratio was demonstrated to be the most efficient lipid profile for predicting coronary disease. LDL was shown to be correlated with hemostatic factors, suggesting that there would be additional benefits to lowering LDL. High triglyceride associated with reduced HDL, indicating insulin resistance and small dense LDL, was shown to be associated with excess coronary disease. All the risk factors tended to cluster, and this was shown to be promoted by insulin resistance induced by weight gain. Multivariate risk profiles were produced to facilitate risk stratification of candidates for coronary disease, stroke, peripheral artery disease and heart failure. The Framingham Study is now engaged in quantifying the independent contributions of homocysteine Lp(a), insulin resistance, small dense LDL, C reactive protein, clotting factors and genetic determinants of cardiovascular disease. We are now able to estimate the lifetime risk of all the atherosclerotic cardiovascular disease outcomes. PMID- 10872617 TI - The effect of the learning curve on the outcome of laparoscopic treatment for gastroesophageal reflux. AB - The laparoscopic treatment for gastroesophageal reflux (GR) by partial (PF) or total (TF) fundoplication is the current surgical treatment of choice after failure of appropriate medical treatment. The overall results with fundoplication include the initial learning period, during which the rate of complications, conversions, and duration of surgery and hospitalization are assumed to be greater. The aim of this study was to compare the results of laparoscopic treatment for GR in three groups of consecutive patients to determine the effect of the learning period on outcome. One hundred and fifty-six patients (88 men and 68 women) with an average age of 52.3 years (range, 18-78) were included. Surgery was indicated for failure or early relapse after the end of medical treatment or a symptomatic sliding hernia. The preoperative workup (endoscopy, barium meal, or esophageal pH monitoring) was governed by the clinical picture. The choice between TF and PF was based on the results of pH monitoring. Three groups of patients were chronologically defined. The parameters that were examined were the type of preoperative exploration, the type of fundoplication, the operative technique, the conversion rate, the mortality and morbidity rates, the duration of surgery and hospitalization, and the results at short- and medium-term follow up. The three groups were comparable with respect to patient characteristics and the nature of their GR. All patients had an endoscopy, 91% had a barium meal, 77.5% underwent esophageal manometry, and 67% had pH monitoring. One hundred and thirty-six patients had a TF and 20 had a PF. Rossetti type TF became the reference procedure (67% in group III) and closure of the diaphragmatic crura was performed systematically in group III (100%). The duration of surgery was significantly reduced between groups I and groups II and III (140, 100, 80 minutes, respectively). The rate of conversion, due to a variety of causes, decreased from 9.8% to 3.8%, and then to 0%. The average duration of hospitalization decreased from 5.8 to 4.2 days (p = 0.01). There was no mortality and the morbidity rate decreased from 15% to 3.8%, and then to 0%. There were seven cases of relapse (4.6%), five in group I (10%) and two in group II (4%), with no cases in group III, although the follow-up in group III was shorter. There is an effect of the learning curve on the outcome of treatment for GR, and this must be taken into account in the training of surgeons (training within experienced departments and guidance during their initial interventions) and also in publications to allow a more accurate comparison of this technique with other treatments for GR. PMID- 10872618 TI - Laparoscopic cholecystectomy for gallbladder dyskinesia: clinical outcome and patient satisfaction. AB - The clinical outcome of laparoscopic cholecystectomy in 63 patients with gallbladder dyskinesia (GD) and 60 patients with proven gallstone (GS) disease was compared. Patients were contacted to determine the extent of symptom relief and satisfaction after surgery. Patients with GD underwent significantly more diagnostic procedures than patients with GS and were found to have higher prevalence of other gastrointestinal motor disorders. Only 47% of patients with GD became completely asymptomatic after surgery, compared with 81% of patients with GS (p = 0.002). This was reflected in the satisfaction scores, which were 79% and 91%, respectively (p < 0.01). There was a significant difference in pathologic findings among the two groups; chronic cholecystitis was more frequently found in the GS group and the incidence of normal gallbladder was higher in the GD group. It is concluded that patients with GD have a good response to laparoscopic cholecystectomy, but they commonly experience continuing gastrointestinal symptoms related to other gastrointestinal motor disorders. PMID- 10872619 TI - Dye-enhanced selective laser ablation for surgical mucosectomy. AB - The diode laser operates at a wavelength of 805 nm; indocyanine green (ICG) has a maximum energy absorption of a wavelength of approximately 800 nm. The effect of the diode laser as a laser knife can be significantly enhanced with an injection of ICG. In the present study, this dye-enhanced photothermal effect was investigated in the field of surgical endoscopy. A 9-cm2 region of the canine gastric mucosa was removed by the laser after injection of 5 ml of ICG solution at a concentration of 0.5 mg/mL into the submucosal space. The diode laser was used at a power output of 10 watts. The canine stomach was removed 10 days after the operation to investigate the site histologically. Clinical application using transanal endoscopic microsurgery (TEM) was employed using a dye-enhanced laser in five patients with a rectal adenoma. The pathological changes in the canine gastric wall resected 10 days after the operation showed that the low-power laser enhanced by ICG produced less fibrosis in the submucosal space than electrocautery. Mucosal resection using a dye-enhanced laser was easily performed in these five patients. It was concluded that this easy removal of the mucosa by a dye-enhanced laser was due to its ability to produce hemostasis of the vessels and its excellent tissue-cutting effect. PMID- 10872620 TI - Efficiency of ultrasound in the detection of the viability of hydatid cysts in the liver. AB - The aim of the present study was to establish the relationship between viability of the hydatid cyst and its ultrasonic appearance (Gharbi classification). To evaluate cyst viability, the criteria that were reported by the World Health Organization in 1982 for both the microbiological evaluation of the cystic fluid and the pathological evaluation of the cyst wall were used. In this study, the possibility of being viable was high in Type I cysts; the possibility of being dead was high in Type IV cysts. It is concluded that there is a relationship between ultrasonic appearance and the evolution of hydatid cysts. PMID- 10872621 TI - Laparoscopic colorectal procedures: a multicenter Brazilian experience. AB - An evaluation of the results of the Brazilian experience in colorectal laparoscopic procedures in a multicenter prospective protocol done by the Brazilian Society of Colo-Proctology is presented. From December 1991 to August 1998, 1,161 patients (583 men and 578 women; mean age, 49.8 years), were operated on laparoscopically. Most of the procedures (40.5%) were for cancer, and the most common procedure was anterior resection (22.5%). The mean operative time was 189 minutes (3.1 hours). There were 42 (3.6%) perioperative complications; visceral injuries were the most common (1.4%). Conversions occurred in 122 (10.5%) cases. There were 148 (12.7%) postoperative complications; wound infections were the most common (5.2%). A liquid diet was started at a mean time of 1.4 days after the operation, and the mean hospitalization period was 6.4 days. PMID- 10872622 TI - Laparoscopic low anterior resection using a triple stapling technique. AB - Laparoscopic low anterior resections using a triple stapling technique in five patients with rectal cancers (four Dukes A and one Dukes C) were performed. The location of the tumors was between 5 and 18 cm from the anal verge. For easy maneuverability, a 33-mm suprapubic port was used. In this technique, the Endo TA (the first stapler) is applied at the distal margin of the rectum to occlude the bowel. The bowel is irrigated with povidone-iodine solution and transected by an endolinear (the second) stapler. Anastomosis is completed by firing the circular (the third) stapler. The operative time was 177 +/- 28.0 minutes, estimated blood loss was 41.7 +/- 28.6 g, and flatus appeared 1.8 +/- 0.8 days after surgery. Follow-up studies have showed no local recurrence or distant metastasis. This procedure is safe and useful for performing laparoscopic low anterior resection. PMID- 10872623 TI - Long-term benefits for the quality of life after video-assisted thoracoscopic lobectomy in patients with lung cancer. AB - Quality of life (QOL) after video-assisted thoracic surgical (VATS) lobectomy remains to be defined. Forty-four consecutive patients with clinical stage I lung cancer underwent lobectomy by the VATS approach (n = 22 patients) or thoracotomy approach (n = 22 patients). Acute pain was quantitated by postoperative narcotic requirements and the need for epidural anesthesia. Long-term QOL was assessed by questioning patients about the presence of chronic chest pain, ongoing limitations in arm or shoulder function, time until return to preoperative activity, and satisfaction with the operation. Patients who underwent VATS lobectomy had significant decreases in both acute and chronic chest pain and time until return to preoperative activity. Patients also had more confidence regarding wound size and their overall impression of the operation. In this series, VATS lobectomy was associated with long-term benefits for the QOL in patients with lung cancer. However, the exact role of this approach should be defined by carefully-designed controlled trials studying long-term survival. PMID- 10872624 TI - Video-assisted thoracoscopic surgery in the diagnosis and treatment of chest diseases. AB - Video-assisted thoracoscopic surgery (VATS) has been used recently in the diagnosis and management of thoracic diseases. In this report, VATS experience with 95 cases, focusing on indications, surgical procedures, complications, and failure rates, are reviewed. Over the past 5 years, 95 VATS procedures for diagnostic and therapeutic purposes were performed in 59 men and 36 women. The specific indications for VATS were lung biopsy for undiagnosed diffuse lung disease (48), mediastinal biopsy (12) and cyst (2), pleural effusion (10), empyema (5), pneumothorax and bullous lung disease (6), pericardial effusion (2) and cyst (2), paravertebral abscess (2), solitary pulmonary nodules (3), and thoracic trauma (3). In all patients, postoperative pain was controlled with non narcotic analgesics and was measured according to the visual analogue scale (VAS). There was no surgical mortality. Postoperative nonfatal complications were seen in seven cases (7.5%). The overall median duration of chest tube drainage was 2.7 days and the mean postoperative hospital stay was 3 days. For diffuse lung disease, a tissue diagnosis was obtained in all the cases. Definitive diagnosis in the patients with undiagnosed pleural effusion was obtained in 90% of cases, and the overall diagnostic rate was 98.5%. The success rate of the therapeutic procedures was 100% after a mean follow-up of 12 months (range, 6-30 months). Conversion to thoracotomy was needed in six cases (6.6%). All patients scored postoperative pain <50% according to the VAS. Video-assisted thoracoscopic surgery should be considered as a procedure of choice, with exceptional results in the following chest diseases: (a) undiagnosed pleural effusions; (b) recurrent, post-traumatic, or complicated spontaneous pneumothorax; (c) stage II empyema; (d) accurate staging of lung cancer; (e) emergency traumatic injuries of the chest; (f) peripheral solitary pulmonary nodule <3 cm; and (g) lung biopsy for pulmonary diffuse disease. PMID- 10872625 TI - Incidental gallbladder cancer at laparoscopy: a review of two cases. AB - Early peritoneal seeding and trocar site metastasis from gallbladder cancer have been reported after laparoscopic cholecystectomy. Nevertheless, the role of laparoscopy in gallbladder cancer remains controversial. Two cases of early recurrence of carcinoma of the gallbladder after laparoscopic cholecystectomy are described. In the first case, the use of a gasless technique did not prevent an early, diffuse peritoneal dissemination of the disease. In the second case, despite the use of a retrieval bag to extract the gallbladder, multiple metastases around the gallbladder bed and local peritoneal seeding developed. These cases demonstrate that factors other than bile spillage, CO2 inflation, and the use of a retrieval bag are responsible for early dissemination of gallbladder cancer. PMID- 10872626 TI - Purely laparoscopic pylorus-preserving gastrectomy with extraperigastric lymphadenectomy for early gastric cancer: a case and technical report. AB - For the purpose of prevention of postgastrectomy syndrome and a less invasive and yet curative oncological resection, a purely laparoscopic pylorus-preserving gastrectomy with extraperigastric lymphadenectomy was performed for a patient with early gastric cancer located in the middle third of the stomach. The patient's postoperative course was uneventful. During his postoperative recovery, the patient experienced very little pain and used analgesic medication only one time. This operation appeared to be oncologically adequate. As of the seventh postoperative month, the patient never experienced dumping syndrome or alkaline reflux gastritis. This procedure is technically feasible and an excellent option because of its reduced surgical invasiveness and better postoperative quality of life. PMID- 10872627 TI - Laparoscopic repair of Bochdalek hernia in an adult. AB - Diaphragmatic hernias of the Bochdalek type are rare in adults. The diagnosis may be made with radiography of the chest in an asymptomatic person or in a person with respiratory and/or gastrointestinal symptoms. It has been mistaken for pleural effusion, empyema, lung cysts, and pneumothorax. A 38-year-old woman presented with epigastric pain and a persistent cough of 2 months' duration. A chest radiograph showed bowel loops in the left side of the chest. On laparoscopy, two defects, measuring 10 and 4 cm, respectively, were seen in the left hemidiaphragm. The herniated fundus of the stomach was reduced and the defect repaired with Gore-Tex mesh (W. L. Gore & Associates, Inc., Flagstaff, AZ, U.S.A.). The patient had an uneventful recovery. Laparoscopic repair of the rare Bochdalek hernia is feasible. PMID- 10872628 TI - Laparoscopic thrombendarterectomy of the infrarenal aorta. AB - The aim was to perform a totally laparoscopic thrombendarterectomy (TEA) of the infrarenal aorta to reduce the trauma connected to the surgical approach. A 52 year-old man was referred to our institution with severe claudication. Angiography revealed a subtotal stenosis of the infrarenal aorta. Because the lesion was not suitable for an interventional procedure, a TEA was planned. The surgery was performed through six ports using a transperitoneal approach with pneumoperitoneum. The laparoscopic TEA was carried out according to the standards of open vascular surgery. The surgery time was 285 minutes, the crossclamping lasted 105 minutes, and the blood loss was 100 mL. The angiographic and functional results were excellent. The patient experienced a rapid recovery and was discharged after 6 days. This case report shows the feasibility of totally laparoscopic TEA of the infrarenal aorta. The well-known advantages of minimally invasive techniques in abdominal surgery with regard to the decrease of surgical trauma may also be valid in aortic surgery for occlusive disease. PMID- 10872629 TI - Mesh fixation with the helical fastener in laparoscopic herinorraphy. PMID- 10872630 TI - Effects of hormone replacement therapy on plasma nitric oxide and total thiol levels in postmenopausal women. AB - Improvement in endothelial function may be an important mechanism by which hormone replacement therapy (HRT) protects postmenopausal women against coronary artery disease. Our aim was to assess the effects of HRT on plasma nitric oxide (NOx) (nitrate plus nitrite) and total thiols in postmenopausal women, as these parameters are associated with enhanced endothelial functions. Thirty-five healthy postmenopausal volunteers (mean age 50.5 +/- 4.7 yr) in an academic and hospital research environment were involved in the study. Blood samples were collected, one at baseline and the second after 6 mo of HRT. Plasma NOx and total thiol levels were significantly elevated in the subjects after HRT. NOx may be of importance in the protective effects of HRT. Further, the increase of plasma antioxidant thiol levels might also contribute to the beneficial effects of HRT. PMID- 10872631 TI - A small dose of ethanol increases the exhalation of mercury in low-level-exposed humans. AB - Inorganic mercury is mainly eliminated by urinary and fecal excretion, but it is also eliminated by exhalation and sweat. There are only a few reports on exhalation of mercury in humans. In volunteers with short-term mercury exposure, an increased exhalation of mercury was found after alcohol intake. The aim of this study was to determine mercury in end-exhaled air and the influence of ethanol on mercury exhalation in subjects with long-term mercury exposure from diet, amalgam fillings, or the work environment. Fourteen subjects, with different grades of mercury exposure, were given 0.2 g ethanol/kg body weight. Measurements of mercury in end-exhaled air were performed before and after alcohol intake. Mercury in end-exhaled air could be detected in all subjects. In 10 individuals without amalgam fillings the mercury concentration was 3 to 12 pg/L. A marked increase, in general about fivefold, in mercury concentrations in end-exhaled air was seen in all subjects 30 min after intake of alcohol, regardless of the level of mercury exposure. Higher ethanol doses resulted in higher mercury levels in end-exhaled air and longer time periods before a return to background levels. An increase was seen even after an ethanol dose of only 0.1 g ethanol/kg body weight (about 0.08 L wine). The decrease in exhaled mercury at higher alcohol doses followed approximately zero-order kinetics and probably reflects the elimination of ethanol in tissues. In conclusion, low levels of mercury can be detected in end-exhaled air also in individuals without amalgam fillings. About a fivefold increase was seen 30 min after alcohol intake, and the relative increase seemed to be independent of the body burden of mercury. Exhalation of mercury represents only a small percentage of the total elimination of mercury. PMID- 10872633 TI - Respiratory toxicity of fabric softener emissions. AB - To determine whether there is any biological basis for complaints that fabric softener emissions can cause acute adverse effects in certain individuals, screening tests were performed in which groups of mice were exposed to the emissions of 5 commercial fabric softener products (antistatic pads used in laundry dryers) for 90 min. Pneumotachographs and a computerized version of ASTM test method E-981 were used to measure acute changes in several respiratory cycle parameters, especially the pause after inspiration, the pause after expiration, and the midexpiratory airflow velocity. From these changes, sensory irritation (SI), pulmonary irritation (PI), and airflow limitation (AFL) of differing intensities were measured with each of the five brands tested. At the peak effect, SI ranged from 21 to 58% of the breaths, PI ranged from 4 to 23% of the breaths, and AFL ranged from 6 to 32% of the breaths. After three exposures, histopathology revealed mild inflammation of interalveolar septae of the lungs. Gas chromatography/ mass spectroscopy (GC/MS) analysis of the emissions of one pad identified several known irritants (isopropylbenzene, styrene, trimethylbenzene, phenol, and thymol). Laundry that had been dried with one the fabric softener pads emitted sufficient chemicals to elicit SI in 49% of breaths at the peak effect Placing one fabric softener pad in a small room overnight resulted in an atmosphere that caused marked SI (61% of breaths). These results demonstrate that some commercial fabric softeners emit mixtures of chemicals that can cause SI, PI, and reduce midexpiratory airflow velocity in normal mice. The results provide a toxicological basis to explain some of the human complaints of adverse reactions to fabric softener emissions. PMID- 10872632 TI - Induction of cytochrome P-450 1A1 in human hepatoma HepG2 and lung carcinoma NCI H322 cells by motorcycle exhaust particulate. AB - The effects of motorcycle exhaust particulate (MEP) on human cytochrome P-450 (P 450)-dependent monooxygenases were determined using human hepatoma cell line HepG2 and lung carcinoma cell line NCI-H322 treated with organic extracts of MEP from a two-stroke engine. Gas chromatography and mass spectrometry analysis of MEP extract revealed the presence of carcinogens benzo[a]pyrene, benz[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[g,h,i]perylene, chrysene, and indeno[1,2,3-c,d]pyrene in the chemical mixture. Treatment with MEP extract produced concentration- and time-dependent increases of monooxygenase activity in HepG2 cells. Treatment of the cells with 100 microg/ ml MEP extract for 24 h markedly increased benzo[a]pyrene hydroxylation, 7-ethoxycoumarin, and 7-ethoxyresorufin O-deethylation activities in microsomes. Immunoblot analysis of microsomal proteins using mouse monoclonal antibody 1-12-3 against P-450 1A1 revealed that MEP extract induced a P-450 immunorelated protein in the hepatoma cells. RNA blot analysis of cellular total RNA using a human P-450 1A1 3'-end cDNA probe showed that MEP extract increased the level of a hybridizable P-450 mRNA. These P-450 1A1 inductive effects of MEP extract were similar to those from treatment with 10 microM benzo[a]pyrene or 3 methylcholanthrene (3-MC) in HepG2 cells. Treatment of lung carcinoma NCI-H322 cells with 100 microg/ml MEP extract, 10 microM benzo[a]pyrene, or 3-MC resulted in induction of monooxygenase activity, protein, and mRNA of P-450 1A1, similar to the induction observed with the hepatoma cells. The present study demonstrates that MEP extract has the ability to induce human hepatic and pulmonary P-450 1A1 in the liver- and lung-derived cell lines, and the induction involves a pretranslational mechanism. Induction of the human hepatic and pulmonary P-450 1A1 in vitro may provide important information in the assessment of MEP metabolism and toxicity in humans. PMID- 10872634 TI - Evaluating the efficiency of toxicity abatement in a constructed wetland with Ceriodaphnia dubia. AB - Constructed wetlands are becoming increasingly popular as low-cost, high efficiency means of treating agricultural and municipal wastewaters. Monitoring programs for constructed wetlands usually measure physical and chemical characteristics of wetland treatment, including hydraulic residence time and removal of nutrients (N, P), suspended solids, and biochemical oxygen demand (BOD). However, toxicity abatement is seldom measured as evidence of wetland treatment efficiency. In this study, toxicity tests combined with chemical measurements were employed to measure the efficiency of a constructed wetland in treating swine wastes during fall and winter sampling periods. Although the wetland system operated at three wastewater loading rates, only the high-loading rate cells were tested because of their year-round flows. Wastewater samples were collected prior to, during, and following wetland treatment to track treatment progress as effluents passed through the wetland cells. Toxicity tests with Ceriodaphnia dubia showed significant toxicity abatement of wastewater as it progressed through the constructed wetland system; however, residual toxicity was still observed in the final wetland effluent. No seasonal differences were observed in toxicity abatement between fall and winter wastewater samples, although nitrate and BOD were removed more efficiently during the fall. Results suggest that, while the constructed wetland system is effective in reducing toxicity in swine wastewater, further pre- or posttreatment or additional dilution is necessary before treated effluents are discharged into surface water. PMID- 10872635 TI - Cognitive resource limitations during sentence comprehension in Parkinson's disease. AB - Patients with idiopathic Parkinson's disease (PD) were asked to identify the agent of the action in orally presented sentences with subject-relative or object relative center-embedded clauses while simultaneously performing a secondary task that was less resource-demanding (finger tapping) or more resource-demanding (recognition span). We found that a subgroup of PD patients with impaired sentence comprehension at baseline (no secondary task) did not differ from random in their accuracy understanding all types of sentences during the more demanding (recognition span) condition and also had difficulty understanding the most complex sentences during the less demanding (finger tapping) condition. Control subjects and PD patients without baseline sentence comprehension difficulty were random only in their comprehension of the most complex sentences under the more demanding (recognition span) secondary task condition. Examination of response latencies for accurately understood sentences revealed only an effect for the type of sentence, and this was equally evident across all groups of subjects and regardless of the condition under which the sentences were administered. The sensitivity of PD patients' sentence comprehension accuracy to secondary task resource demands is most consistent with the hypothesis that limited cognitive resources contribute to sentence comprehension difficulty in PD. PMID- 10872636 TI - The rise and fall of frequency and imageability: noun and verb production in semantic dementia. AB - This study examines the impact of progressive degeneration of conceptual knowledge on the content words used in connected speech elicited using the Cookie Theft picture description (Goodglass & Kaplan. 1983). We began with an analysis of control subjects' descriptions with regard to word types and their frequency and imageability. Because the impairment of conceptual knowledge in semantic dementia is graded by concept familiarity, we created a model of a standardized normal Cookie Theft description that was then progressively degraded by the systematic removal of lower bands of word frequency. We drew two main predictions from this model: reduced availability of the lower bands of word frequency should result in (a) an apparent deficit for noun retrieval in relation to verb retrieval and (b) an apparent reverse imageability effect. Results from a longitudinal study. in which three patients with semantic dementia each described the Cookie Theft picture on three occasions during the progression of their disease, confirmed these predictions. An additional cross-sectional analysis, adding narratives from a larger number of cases, demonstrated that the decline in ability to produce suitable words for the picture description is closely related to the extent of semantic impairment as measured in tests of word comprehension and production. Both verbs and nouns are affected by the degradation of semantic memory; the fact that the impairment to noun production is manifested earlier and more catastrophically may be attributed to the relatively lower frequency of these terms. PMID- 10872637 TI - Sentence and word outline shape as co-primes for target words presented to the two visual hemifields. AB - A lexical decision experiment tested visual field stimulation of word targets after priming the central visual field by the target word outline shape and/or an incomplete sentence. In general, RT was shorter and accuracy better for target words presented to the RVF. Responses were quicker and more accurate to target words presented to either visual hemifield after priming by either a congruent incomplete sentence or a congruent word outline shape (WOS). However, the joint effect of WOS and an incomplete sentence as co-primes was different when the succeeding word target appeared in the RVF than when it appeared in the LVF. While a congruent WOS and incomplete sentence acting as co-primes reduced RT to LVF targets orthogonally. the two variables operated interactively as co-primes on target words presented to the RVF. PMID- 10872638 TI - A model-driven analysis of severity, response characteristics, and partial recovery in aphasics' picture naming. AB - Dell, Schwartz, Martin, Saffran, and Gagnon (DSMSG; 1997) presented a computational analysis of aphasic naming that, among other things, purports to explain why some error types correlate with naming severity while others do not. It does so in terms of chance response opportunities, which differ among error types and which come into play particularly when activation levels are small. The present study looks at error frequencies in relation to severity at two points in time: at study entry and after a period of partial recovery. Results support the model's distinction between severity-sensitive errors (nonwords. formal paraphasias, and unrelated errors) and those that are severity insensitive (semantic; mixed). Additionally, we show that the degree of target overlap in nonwords is sensitive to severity but various measures of monitoring and error correction are not. While these results generally support DSMSG, effects at the level of individual patients underscore the difficulties that their model encounters in explaining some pure error dissociations. PMID- 10872639 TI - Semantic representation and ease of predication. AB - Jones' (1985) Ease of Predication hypothesis, which states that underlying differences in the semantic representation of concrete and abstract words can be explained in terms of disproportionate numbers of semantic predicates, is explored in two experiments. The results suggest that (1) the advantage shown by concrete words in terms of greater number of predicates is only apparent for words of low frequency, and (2) Jones' case of predication variable does not accurately reflect predicate distributions, or differences in imageability. Rather, it appears to represent differences in concreteness. As such, the validity of this concept as the basis of theories of semantic representation is questioned. Models based on the assumption of a "richer" semantic representation for concrete words are therefore not supported. PMID- 10872640 TI - Return of stuttering after stroke. AB - The pathophysiology of developmental or acquired stuttering still remains an enigma. In a few cases, the developmental stuttering that had disappeared spontaneously or as a result of therapy reoccurred following a brain lesion. We report on a patient with return of developmental stuttering following a left hemispheric stroke. This case supports the theory that acquired brain lesions may cause a return of stuttering, possibly by interfering with the compensatory mechanism(s) that once had relieved the developmental stuttering. PMID- 10872641 TI - Ethanol and production of the hepatotoxic metabolite of acetaminophen in healthy adults. AB - BACKGROUND: Recent case reports suggest that consumption of ethanol may increase the risk of liver injury induced by acetaminophen (INN, paracetamol). However, this possibility is at odds with previous clinical studies that showed that acute ethanol ingestion could protect against hepatotoxicity by inhibiting CYP-mediated acetaminophen oxidation. We tested the hypothesis that ethanol ingestion can increase susceptibility to acetaminophen toxicity if acetaminophen ingestion occurs shortly after ethanol is cleared from the body. METHODS: Ten healthy volunteers each received a 6-hour intravenous infusion of ethanol (to achieve a blood concentration of 100 mg/dL ethanol) or 5% dextrose in water, administered in random order. Acetaminophen (500 mg) was ingested 8 hours after the end of the infusion. Blood and urine were collected for assessment of formation of N-acetyl p-benzoquinone imine (NAPQI), the hepatotoxic metabolite of acetaminophen. RESULTS: Mean NAPQI formation was enhanced by 22% (range, 2% to 38%; P < .03) when the acetaminophen dose was given after an ethanol infusion, compared with after 5% dextrose in water infusion. This mean increase was similar in magnitude to that predicted by a mathematical model describing the induction of CYP2E1, the main enzyme catalyzing NAPQI formation, by a mechanism of enzyme stabilization. CONCLUSIONS: Consumption of up to one 750-mL bottle of wine, six 12-ounce cans of beer, or 9 ounces of 80-proof liquor over the course of a single evening modestly increases the fraction of an acetaminophen dose converted to its toxic metabolite, NAPQI, when acetaminophen is ingested soon after ethanol has been cleared from the body. This change in acetaminophen metabolism may present an incremental increase in the risk of acetaminophen hepatotoxicity. PMID- 10872642 TI - Population pharmacokinetic analysis of netilmicin in neonates and infants with use of a nonparametric method. AB - BACKGROUND: Although the therapeutic and toxic effects of netilmicin are related to its plasma concentration, its pharmacokinetics in neonates and infants and the influence of clinical and biological variables have been only partially assessed. METHODS: Therapeutic drug monitoring data collected from 186 neonates and 95 infants receiving netilmicin were analyzed with a nonparametric population approach. The influence of gestational and postnatal age, weight, Apgar score, and creatinine and urea plasma concentrations on the pharmacokinetic parameters was assessed. The neonate and infant groups were each randomly divided into a learning sample and a validation sample. The population analysis was performed on each learning subgroup with the nonparametric maximum likelihood (NPML) method. In the validation group, the data were used to assess the concentration predictability. Because there is no specific netilmicin formulation for neonates and infants, an error model was proposed to account for errors attributable to dilution processes when preparing the infusion. RESULTS: In neonates, the covariates that reduced expected variance of plasma clearance by more than 10% were postnatal age, body weight, and plasma creatinine, as well as plasma urea and creatinine in infants. Body weight and sex played a significant role in explaining the variability of the volume of distribution. The accuracy of the concentration predictability assessed in the validation samples was satisfactory, and no significant bias was found. CONCLUSION: These findings help explain the large interindividual variability of the pharmacokinetics of netilmicin and the influence of the clinical and laboratory covariates in neonates and infants. PMID- 10872643 TI - Population pharmacokinetics of levodopa in patients with Parkinson's disease treated with tolcapone. AB - OBJECTIVE: To use pharmacostatistical models to evaluate the overall exposure of patients with Parkinson's disease to levodopa in the presence and absence of tolcapone. METHODS: Four hundred twelve patients with Parkinson's disease with fluctuating and nonfluctuating responses to levodopa participated in three multicentered, parallel, double-blind, placebo-controlled dose-finding studies and received either placebo or tolcapone in addition to levodopa-decarboxylase inhibitor therapy. Sparse blood samples were obtained from 393 patients for levodopa and 3-O-methyldopa assay, and the data were analyzed with use of the NONMEM program. RESULTS: The fraction of levodopa metabolized to 3-O-methyldopa was substantially reduced by the co-administration of tolcapone (by 65%, 74%, and 84% with tolcapone doses of 50, 200, and 400 mg, respectively, in fluctuators, and by 50% and 90% with doses of 200 and 400 mg, respectively, in nonfluctuators). This led to an overall reduction in levodopa clearance (CL) of approximately 15% to 25% in fluctuators and 20% to 30% in nonfluctuators. Because this was partly compensated for by a reduction in levodopa dose in these studies, the total daily exposure of patients to levodopa was only slightly increased (11% to 16%). The peak-trough fluctuations of plasma levodopa (Cmax-Cmin) were reduced in both populations in a dose-dependent fashion. CONCLUSIONS: Tolcapone effectively inhibited the formation of 3-O-methyldopa and resulted in a decrease in levodopa CL. The consequent increase in levodopa bioavailability was mostly offset by reductions in levodopa dose. It is possible that decreased fluctuations in plasma levodopa concentrations rather than increased levodopa exposure may explain the clinical benefits obtained with tolcapone. PMID- 10872644 TI - Validation of techniques for the prediction of carboplatin exposure: application of Bayesian methods. AB - OBJECTIVE: Several methods have been developed for the prediction of carboplatin exposure to facilitate pharmacokinetic guided dosing. The aim of this study was to develop and validate sparse data Bayesian methods for the estimation of carboplatin exposure and to validate other commonly applied techniques, such as the Chatelut formula, the Sorensen limited sampling model, and the Calvert formula, in which glomerular filtration rate was estimated with the Cockcroft Gault, the Jelliffe, and the recently proposed Wright formulas. METHODS: Complete concentration-time curves were available for a total of 43 patients (45 courses) receiving carboplatin (265 or 400 mg/m2/day) in a 1-hour infusion for 4 consecutive days in combination with thiotepa and cyclophosphamide. A population two-compartment model was developed on an index set of 12 courses. The other 33 courses served as validation set. Bayesian estimates were generated with the population parameters by use of either one or two randomly timed samples or two samples at optimal time points determined with the D-optimality theory. RESULTS: The Bayesian methods provided an accurate and precise prediction of the area under the concentration-time curve (bias <4% and precision <18%). The other formulas (Sorensen model, Chatelut, and Calvert with Jelliffe, Cockcroft-Gault, and Wright) resulted in a precision >18%, whereas the Jelliffe formula and the Sorensen model resulted in a bias >12%. CONCLUSION: The applicability of a Bayesian method for the prediction of the carboplatin exposure by use of one or two samples without the necessity for exact timing of infusion duration and sampling was demonstrated. The Bayesian method may be very instrumental to execute pharmacokinetic guided dosing for carboplatin. PMID- 10872645 TI - Pharmacokinetics of methotrexate in cerebrospinal fluid and serum after osmotic blood-brain barrier disruption in patients with brain lymphoma. AB - OBJECTIVE: To evaluate the pharmacokinetics of methotrexate in ventricular cerebrospinal fluid and serum after osmotic blood-brain barrier disruption and intra-arterial administration compared with intravenous or simple intra-arterial infusion in patients with primary central nervous system lymphoma. METHODS: Serum and ventricular cerebrospinal fluid were sampled after methotrexate administration in 12 patients. Blood-brain barrier disruption was induced on 2 sequential days by mannitol (25%) infusion delivered to the vertebral or internal carotid artery territories followed by intra-arterial methotrexate (dose, 1.4 g/m2; 47 treatments). Sixteen treatments were given without barrier disruption by intravenous (3.5 g/m2; nine treatments) or intra-arterial (2.8 g/m2; seven treatments) infusion. RESULTS: Ventricular cerebrospinal fluid-methotrexate peak levels after blood-brain barrier disruption of the vertebral and the internal carotid arteries territories were 19.3 +/- 2.9 and 8.5 +/- 0.7 micromol/L (P < .001), and the area under the curve from time 0 to infinity was 178.0 +/- 21.3 and 110.0 +/- 12.4 [micromol/L x h, respectively (P < .01). No significant differences were observed in serum levels. After intra-arterial infusion was performed without disruption, the serum peak level was higher than that achieved by intravenous treatment (518.2 +/- 67.7 versus 180.6 +/- 31.8 micromol/L; P < .001). No differences were observed in cerebrospinal fluid concentrations, which dropped below 1 micromol/L at 6 hours. The cerebrospinal fluid/serum ratio [AUC(%)] of methotrexate after blood-brain barrier disruption was three to four times greater than that by systemic administration. CONCLUSION: Enhanced methotrexate delivery to the central nervous system can be attained by intra arterial administration combined with osmotic disruption of the blood-brain barrier compared with simple intra-arterial or intravenous administration. PMID- 10872646 TI - Population pharmacokinetics of long-term oral amiodarone therapy. AB - BACKGROUND: Amiodarone is an increasingly popular and uniquely effective antiarrhythmic agent for which population pharmacokinetic parameters in patients receiving long-term oral therapy have not been defined previously. METHODS: We collected 605 observations of serum amiodarone and desethylamiodarone metabolite concentrations from 77 patients (mean follow-up, 2 years). Mixed-effects modeling (NONMEM) was used to determine the typical population pharmacokinetic parameters, their respective variabilities, and a simple oral dosing regimen to rapidly achieve and maintain a target concentration of 1.5 mg/L. Individual serum concentration versus time curves were simulated for the study population based on regimens outlined in the product monograph and were compared with those for the proposed dosing regimen. The relationship between the duration of amiodarone therapy and the rate of decrement in serum concentration after discontinuation was explored. RESULTS: Amiodarone concentrations were best described by a two compartment model with the typical parameters +/- interindividual coefficients of variation (where applicable) as follows: volumes of distribution/bioavailability (V1/F = 882 L; V2/F = 12,700 L +/- 58%) and clearances/bioavailability (CL1/F = 229 L/day +/- 31%; and CL2/F = 599 L/day +/- 56%). Rapid distribution half-life was 17 hours, and terminal half-life was 55 days. A practical dosing regimen of 1600 mg/d for 2 days, 1,200 mg/d for 5 days, 1,000 mg/d for 7 days, 800 mg/d for 7 days, 600 mg/d for 7 days, and 400 mg/d for 62 days followed by a maintenance dose of 343 mg/d (400 mg/d for 6 of 7 days) is proposed. After steady state is reached, cessation of dosing produces a 25% serum concentration decrement in 3 days and 50% in 36 days. CONCLUSIONS: Population pharmacokinetics confirm that amiodarone has an extraordinarily long half-life. The slow elimination rate makes anticipating the timing of adjustments in amiodarone therapy to avoid toxicity unusually perplexing. However, based on the estimated variability, the proposed dosing regimen would produce steady-state concentrations within the therapeutic window for 90% of patients. PMID- 10872647 TI - Effects of cigarette smoking and carbon monoxide on nicotine and cotinine metabolism. AB - OBJECTIVES: To examine the effects of cigarette smoking on the disposition kinetics of nicotine and cotinine, to determine the effects of cigarette smoking on pathways of nicotine and cotinine metabolism, and to test the hypothesis that carbon monoxide inhibits the metabolism of nicotine. STUDY DESIGN: Twelve cigarette smokers were studied in three treatment conditions, each lasting 7 days, during which they smoked cigarettes, breathed carbon monoxide to achieve carboxyhemoglobin levels similar to cigarette smoking, or breathed air. In each treatment condition, subjects received a combined infusion of deuterium-labeled nicotine (d2) and cotinine (d4), with measurement of disposition kinetics and urine metabolite profile. RESULTS: Cigarette smoking significantly inhibited the metabolism of nicotine but had no effect on cotinine metabolism. Cigarette smoking markedly induced the O-glucuronidation of trans-3'-hydroxycotinine but had no effect on the N-glucuronidation of nicotine or cotinine. Carbon monoxide had no effect on nicotine or cotinine kinetics or metabolic profile. CONCLUSIONS: This study confirms previous observations that cigarette smoking inhibits nicotine metabolism but disproves the hypothesis that this effect is due to carbon monoxide. Induction of glucuronidation must be considered in understanding the effects of cigarette smoking on drug metabolism. PMID- 10872648 TI - Pituitary and gonadal endocrine effects and pharmacokinetics of the novel luteinizing hormone-releasing hormone antagonist teverelix in healthy men--a first-dose-in-humans study. AB - BACKGROUND. Teverelix is a novel synthetic peptidic luteinizing hormone-releasing hormone (LHRH) antagonist. METHODS: Single subcutaneous morning doses of teverelix acetate (either 0.5, 1, 2, 3, or 5 mg base) were investigated in a randomized, single-blind, placebo-controlled, dose-escalating parallel-group design in healthy men. Six subjects received teverelix, and two subjects received placebo per dose level. Blood samples for lutropin, luteinizing hormone (LH), and follitropin, follicle-stimulating hormone (FSH), and testosterone, as well as for pharmacokinetics, were withdrawn up to 120 hours after dosing. Serum hormone levels were determined by electrochemicoluminescence immunoassays, and plasma teverelix concentrations were determined by radioimmunoassay. RESULTS: Teverelix led to a rapid, marked suppression of LH, testosterone and, to a lesser extent, FSH. Median maximum suppressions compared with predose levels were -93% for LH and -54% for FSH after teverelix 5 mg, and -93% for testosterone after teverelix 3 mg, respectively. After 5 mg teverelix, testosterone suppression <1 ng/mL started a median of 12 hours after dosing and lasted for a median of 33 hours. The duration of testosterone suppression increased with dose. Geometric means of peak teverilix plasma concentrations were 4.5 ng/mL (0.5 mg teverelix) to 49.0 ng/mL (5 mg teverelix) and tmax occurred between 1 and 4 hours after dosing. Geometric means of the area under the teverelix plasma concentration-time course from zero to time of the last quantifiable plasma concentration [AUC(O-tlast)] were 54.9 ng x h/mL (0.5 mg teverelix) to 881.8 ng x h/mL (5 mg teverelix). Median values for apparent terminal half-lives ranged from 24 to 75 hours. The most frequently reported adverse events were short-lasting mild injection-site reactions. CONCLUSIONS: Teverelix showed pronounced LH and testosterone suppressive effects after single subcutaneous doses in healthy men. Duration of hormone suppression increased with dose. Teverelix was well tolerated. This profile indicates potential for further clinical use. PMID- 10872649 TI - Human beta2-adrenergic receptor polymorphisms: no association with essential hypertension in black or white Americans. AB - BACKGROUND AND OBJECTIVES: The most common polymorphisms of the human beta2 adrenergic receptor--Arg16-->Gly and Gln27-->Glu--are associated with alterations in beta2-adrenergic receptor responses, both in vitro and in vivo. beta2 Adrenergic receptor-mediated vascular responses are affected by ethnicity, blood pressure, and genotype. We tested the hypothesis that these two common beta2 adrenergic receptor genetic variants are associated with essential hypertension in black or white Americans. SUBJECTS AND METHODS: In a population-based case control association study, the relationship between beta2-adrenergic receptor genotypes and hypertension was examined in 307 normotensive subjects (128 black and 179 white) and 356 hypertensive subjects (155 black and 201 white). A polymerase chain reaction-based single-stranded conformational polymorphism method with direct sequencing of the bands of interest was used to detect the two frequently occurring beta2-adrenergic receptor variants (Arg16-->Gly, Gln27- >Glu). RESULTS: No significant differences in the distributions of alleles and genotypes of the tested beta2-adrenergic receptor variants were found between normotensive and hypertensive groups from either black or white Americans (all P > .05). There was a marked interethnic difference in the frequency of the Gln27- >Glu beta2-adrenergic receptor polymorphism in both normotensive and hypertensive subjects. In normotensive white subjects, the variant Glu27 allele (35.2% versus 18.0%; P < .0001) and Glu27 homozygous genotype (14.0% versus 4.7%; P < .01) were more common than in black subjects. Similarly, in hypertensive white subjects, the variant Glu27 allele (35.8% versus 18.4%; P < .0001) and the Glu27 homozygous genotype (15.9% versus 2.6%; P < .0001) were more common than in black subjects. CONCLUSIONS: These data suggest that although there are marked ethnic differences in their distribution, the common genetic polymorphisms of the human beta2 adrenergic receptor gene do not cosegregate with the presence of hypertension in either black or white Americans. PMID- 10872650 TI - Effects of prophylactic ibuprofen on cerebral and renal hemodynamics in very preterm neonates. AB - OBJECTIVE: To evaluate the effects on cerebral and renal blood flow velocities of ibuprofen when used as prophylaxis for patent ductus arteriosus in preterm neonates (gestational age <30 weeks). METHODS: Blood flow velocities in the anterior cerebral artery and the renal artery were measured with Doppler ultrasonography in 17 neonates before, during, and 10, 30, and 60 minutes after administration of 10 mg/kg ibuprofen lysine. RESULTS: In four (23.6%) neonates without echocardiographic patency of the ductus, no significant modifications in blood flow velocities and Doppler indexes were found either in the anterior cerebral artery or in the renal artery. In 13 (76.4%) neonates, cardiac echocardiographic Doppler showed patency of the ductus and left-to-right shunt. In these neonates diastolic and mean blood velocities rapidly increased both in the anterior cerebral artery and the renal artery (P < .0001). Resistance and pulsatility index decreased during the study period (P < .0001 and P < .001, respectively, in the anterior cerebral artery; P < .0001 in the renal artery). CONCLUSIONS: Data suggest that ibuprofen does not determine any direct effect on cerebral and renal blood flow velocities; hemodynamic modifications observed in neonates with patency of ductus can be related to closure of the ductus induced by the drug. PMID- 10872651 TI - Cure of refractory duodenal ulcer and infection caused by Helicobacter pylori by high doses of omeprazole and amoxicillin in a homozygous CYP2C19 extensive metabolizer patient. AB - A 53-year old female patient with duodenal ulcer and Helicobacter pylori infection was treated three times with a proton pump inhibitor-based triple therapy, such as lansoprazole-clarithromycin-amoxicillin (INN, amoxicilline) and lansoprazole-minocycline-cefaclor. However, the H pylori infection was not cured. A culture test revealed that her infection was a clarithromycin-resistant but amoxicillin-sensitive strain of H pylori. Moreover, a polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis revealed that she was a homozygous extensive metabolizer of cytochrome P450 (CYP) 2C19 (wt/wt). The usual dose of the proton pump inhibitor was therefore assumed to be insufficient for her and then she was treated with a high dose of omeprazole (120 mg/day) and amoxicillin (2,250 mg/day) for 2 weeks. The H pylori infection and the ulcer lesion were then cured. One of the factors associated with success or failure of cure of H pylori infection by the proton pump inhibitor-based triple therapy appeared to be CYP2C19 genotype status. Dual treatment with a sufficient dose of a proton pump inhibitor plus amoxicillin could cure H pylori infection even after the failure to cure H pylori infection by a usual proton pump inhibitor-based triple therapy in patients with the wt/wt homozygous extensive metabolizer genotype of CYP2C19. PMID- 10872652 TI - Grapefruit juice-lovastatin interaction. PMID- 10872653 TI - DNA topoisomerase expression in tumors--a novel target for chemotherapy. PMID- 10872654 TI - Low-grade intraosseous osteosarcoma in northern Japan: advantage of AgNOR and MIB 1 staining in differential diagnosis. AB - Low-grade intraosseous osteosarcoma is an uncommon form of bone cancer. It is occasionally difficult to recognize as a malignant tumor and is commonly misdiagnosed as a benign fibrous lesion. We retrospectively studied the records of 8 patients with low-grade intraosseous osteosarcoma in the files of the Tohoku Musculoskeletal Tumor Society in Japan. All tumors arose in the lower limb. The most common symptom was pain, with a duration exceeding 2 years in 4 patients. Radiologic findings, including those at magnetic resonance imaging (MRI), suggested malignancy in 5 lesions, whereas 3 were diagnosed as benign. Two patients initially presented with pathological fracture. The initial pathological diagnosis was malignant in 5 patients and benign in 3. All eight tumors were grade 1 in Broders' classification. The tumor showed a permeative pattern in all eight cases, but this pattern could not be confirmed in the multiple tiny fragments obtained as biopsy specimens in 3 cases. The number of silver-staining nucleolar organizer regions (AgNOR) per nucleus and MIB-1-positive rate were significantly higher in low-grade intraosseous osteosarcoma than in fibrous dysplasia, offering an advantage in differential diagnosis. Three patients (38%) developed high-grade sarcoma at the site of local recurrence after multiple intralesional excisions, and one of them died of the disease. The other 5 patients had a good clinical course after surgery with a wide margin. These findings indicate that preoperative diagnosis with radiologic investigation, including magnetic resonance (MR) imaging and histologic examination of biopsy specimens is essential in preparation for surgery with a wide margin, assuring a good clinical course, and the results of AgNOR and immunohistochemical MIB-1 staining might be helpful in differentiating low-grade intraosseous osteosarcoma from fibrous dysplasia. PMID- 10872655 TI - Collagen and elastin degradation by matrix metalloproteinases and tissue inhibitors of matrix metalloproteinase in aortic dissection. AB - The degradation of collagen fibrils and elastic fibers in 21 cases of acute aortic dissection (AD) was ultrastructurally and immunohistochemically investigated; and the expression of the catabolic matrix metalloproteinases (MMPs)-1, -2, -3, and -9 and their inhibitors, the tissue inhibitors of matrix metalloproteinase (TIMPs)-1 and -2, was studied. The features of the entry site of the dissection (ES; 21 ascending aortas) were compared with those of fully remote sites (RS; 19 nondissected abdominal aortas) and the ascending aortas from 10 control cases. By electron microscopy, the medial layer at the ES and adjacent intact aortic wall demonstrated spirally thickened collagen fibrils with a typical banding pattern that were almost always colocalized with elastic lamellae, which often exhibited attenuation, fragmentation, or disruption. In addition, the basement membrane surrounding the smooth muscle cells (SMCs) comprising the media was frequently thinned or lost at the ES. These findings were rarely seen at the RS or in the aortas of controls. Immunohistochemically, the expression of MMP-1 was significantly in the cytoplasm of SMCs of both the intima and media at the ES and adjacent intact wall, and significant expression of MMP-2 and -9 was found in SMCs of the intima compared with the RS and controls. Significant expression of TIMP-1 and -2 was demonstrated in the cytoplasm of SMCs at the ES and adjacent intact wall compared with that at the RS and the control specimens. These findings suggest that the degradation of proteins associated with fibrosis and the occurrence of AD are not merely coincident, but rather that AD is induced by alterations of the extracellular matrix caused by changes of SMCs at a segment of the ascending aorta made vulnerable through hemodynamic stress, especially that caused by hypertension. PMID- 10872656 TI - Thomsen-Friedenreich glycotope is expressed in developing and normal kidney but not in renal neoplasms. AB - The Thomsen-Friedenreich glycotope (TF) is considered a general carcinoma autoantigen and is therefore of importance in cancer diagnosis and immunotherapy. We report the distribution of the TF glycotope in developing and adult human kidney and renal neoplasms. A monoclonal antibody and the lectin amaranthin were used to study the TF and its sialylated, masked form by immunohistochemistry and immunoblotting. In developing kidney, the TF was restricted to the loop of Henle, distal tubules, and peripheral collecting ducts, whereas its sialylated form was detectable in all epithelial differentiations derived from the 2 embryonic anlagen, the metanephrogenic blastema being unreactive. This pattern was essentially preserved in adult kidney, with TF labeling beginning in the thick ascending limb and extending into the collecting ducts of outer medulla. The sialylated TF glycotope was additionally observed in ascending thin limbs. The TF was exclusively expressed in the luminal cell surface and hence was inaccessible to immune reactions. Analysis of a spectrum of renal neoplasms failed to detect the TF, with the exception of occasional staining of tubules in nephroblastoma. Moreover, the sialylated TF was only detectable in oncocytoma, chromophobe renal cell carcinoma, cystic nephroma, nephroblastoma, and nephroblastomatosis complex and occasionally in type 1 papillary renal cell carcinoma. Thus, the TF and its sialylated form are expressed in normal developing and adult kidney. However, the TF does not seem to represent a tumor-associated glycotope in human kidney, nor does it appear to be of value in diagnosis and immunotherapy of renal neoplasms. PMID- 10872657 TI - Endocervical intraepithelial glandular atypia (dysplasia): a histopathologic, human papillomavirus, and MIB-1 analysis of 25 cases. AB - The purpose of this study was to determine the likelihood that intraepithelial endocervical glandular atypias that are less severe than adenocarcinoma in situ (AIS) are precursors to AIS and, if so, whether they can be recognized by morphological or other means. We first assessed the frequency of atypias found in association with either AIS or invasive adenocarcinoma (ACA) and then tested these cases and additional randomly encountered cases for the presence of human papillomavirus (HPV) and for their proliferative (Ki-67) index. Lesions not fulfilling the classic criteria for AIS were subdivided into high-grade (HGGA) and low-grade glandular atypias (LGGA). Atypias and controls were microdissected and tested for HPV by the polymerase chain reaction. Serial sections were labeled for Ki-67 by immunohistochemistry with the MIB-1 antibody. Eight cases (6.8%) containing glandular atypia were found in a search of 117 consecutive cone biopsy or hysterectomy specimens that also had either AIS, ACA, or both. An additional 17 cases were either randomly encountered or were received in consultation. In 3 cases, both HGGA and LGGA were present, yielding a total of 28 lesions for study. Of the 9 HGGA cases that were associated with either AIS, ACA, or CIN II/III, 6 were positive for HPV; MIB-1 reactivity in all 6 was present in greater than 25% of the nuclei. Of the 3 HPV-negative HGGA cases in this group, the 2 that were tested showed low MIB-1 reactivity. All 3 cases of HGGA that were not associated with a diagnostic lesion were HPV-negative and had low MIB-1 reactivity. Of the 6 LGGAs associated with either AIS, ACA, or CIN II/III, 1 was positive for HPV; MIB 1 was nonreactive in this case and was low in all of the HPV-negative cases in this group that were tested. Of 10 LGGAs not associated with a diagnostic lesion, or with a low-grade squamous lesion as the only other abnormality, 2 were positive for HPV. Of these 2, one had an MIB-1 reactivity of greater than 25% and also had intestinal differentiation. MIB-1 reactivity was elevated in 2 of the 8 HPV-negative LGGAs from this group. All 10 ciliated atypias (3 HGGA, 7 LGGA) were HPV-negative and had low MIB-1 reactivity. One HPV-positive AIS control case was focally ciliated. Six of 7 foci with apoptotic bodies (5 HGGA, 2 LGGA) were HPV positive. The infrequent occurrence of glandular atypias with AIS and ACA and the low rate of HPV DNA positivity when they are found in isolation are evidence that most AIS lesions do not evolve through morphologically identifiable antecedents and that most isolated atypias are not AIS precursors. HGGAs associated with AIS or CIN II/III maybe either precursors to or subtle variants of AIS. However, LGGAs similarly encountered are unlikely to be either. Elevated MIB-1 reactivity may be helpful diagnostically in selected cases, but it is not reliable as an independent criterion. The presence of cilia in isolated glandular atypias favors a nonneoplastic process, whereas intestinal differentiation and apoptotic bodies each suggest neoplasia. PMID- 10872658 TI - Microsatellite instability and K-ras mutations in patients with ulcerative colitis. AB - Patients with inflammatory bowel disease (IBD), particularly ulcerative colitis (UC), have an increased incidence of colorectal carcinoma. The underlying mechanism is unknown, but we postulated that microsatellite instability (MSI) might predispose the colonic mucosa of UC patients to mutations, thereby increasing their cancer risk. We also sought to determine the frequency of K-ras mutations, to determine whether MSI predisposed to K-ras mutations and to compare the molecular phenotype of biopsy and resection specimens in the same patient. We also sought to determine whether molecular alterations found in biopsy specimens presaged their presence in subsequent resection specimens. Two hundred fifty eight specimens from 52 patients were examined for K-ras mutations by direct sequencing. Seventy-one of the specimens were neoplastic. MSI was evaluated after polymerase chain reaction (PCR) amplification using primers directed at 8 microsatellite loci. Of the patients, 18.2% had K-ras mutations, and 30.8% had MSI in at least 1 locus. Of K-ras mutations, 81.8% were G to A substitutions involving the second nucleotide of codons 12 or 13. Only 0.7% of the samples showed a high level of MSI. No relationship existed between MSI and K-ras mutations, even in the 2 samples with high-level MSI. The numbers are small, but it appeared that MSI in biopsies failed to predict its presence in resection specimens. In contrast, K-ras mutations present in biopsy specimens tended to predict their presence in resections. K-ras mutations were found predominantly in neoplastic mucosae, whereas MSI was found predominantly in regenerative mucosae. The lack of any relationship between MSI and K-ras mutations suggests that MSI in the UC replicative mucosa does not predispose to colonic neoplasia via a K-ras mediated pathway. This is probably related to the fact that the MSI is generally low-level MSI. PMID- 10872659 TI - Differential expression of MUC2 and MUC5AC mucin genes in primary ovarian and metastatic colonic carcinoma. AB - Colonic adenocarcinoma, the most common tumor metastatic to the ovary, may closely mimic primary ovarian adenocarcinoma, especially that of mucinous or endometrioid histology. The differential diagnosis is important for therapeutic considerations. Mucin gene expression is relatively organ-specific and may therefore have use in distinguishing between colonic carcinomas metastatic to the ovary and primary ovarian tumors. In this study, we compared the expression of MUC2 and MUC5AC apomucins in 10 colonic adenocarcinomas metastatic with the ovary, 10 ovarian endometrioid carcinomas (4 primary, 6 metastatic), and 32 primary mucinous ovarian tumors (12 cystadenomas, 10 borderline tumors, and 10 cystadenocarcinomas). Monoclonal antibodies CCP58 and 45M1 were used for immunostains of MUC2 and MUC5AC apomucin, respectively. All but 1 of the 10 metastatic colon adenocarcinomas expressed MUC2, whereas none expressed MUC5AC. None of the 10 endometrioid carcinomas expressed MUC2, and only 2 showed weak immunoreactivity with MUC5AC. All 32 primary mucinous ovarian tumors expressed MUC5AC. The percentages of MUC2-positive immunostaining for cystadenomas, borderline tumors, and cystadenocarcinomas were 0% (0/12), 50% (5/10), and 70% (7/10) respectively. These studies show that MUC2 and MUC5AC are useful markers in the distinction between colonic carcinoma metastatic to the ovary and primary ovarian carcinoma. PMID- 10872660 TI - Radiation-induced pseudocarcinomatous proliferations of the urinary bladder: a report of 4 cases. AB - Four cases of radiation cystitis that caused diagnostic difficulty because of an epithelial proliferation with architectural complexity and reactive cytologic atypia are described. The patients, 2 male, 2 female, were from 43 to 77 years of age. Two presented with hematuria. Cystoscopy disclosed abnormalities in 3 patients. Microscopic examination showed irregularly shaped and arranged aggregates of epithelial cells in the upper and mid zones of the lamina propria. The cells, which typically showed at least mild, and sometimes severe, pleomorphism, were usually transitional, but squamous differentiation was seen focally in 3 cases. Ulceration of the overlying epithelium was present in all cases and was prominent and associated with conspicuous fibrin and hemorrhage in one of them. Edema of the lamina propria was present in 3 cases, whereas lamina propria fibrosis and chronic inflammation were present in all cases. The presence in all 4 cases of vascular ectasia and other changes characteristic of radiation injury, such as atypical fibroblasts, prompted investigation of the clinical history in 2 cases in which the pathologist was unaware that the patient had received radiation. Pseudocarcinomatous proliferations in the bladder caused by radiation injury have received limited attention in the literature. Our cases illustrate the potential diagnostic errors with which these lesions may be associated. PMID- 10872661 TI - Parietal cell protrusions and fundic gland cysts during omeprazole maintenance treatment. AB - Parietal cell protrusion (PCP), swelling and bulging of parietal cells, has been observed in the oxyntic mucosa of patients receiving omeprazole. The frequency of this event and the underlying mechanisms remain to be clarified. As such, it is unknown whether there is a relation with either serum gastrin or Helicobacter pylori infection, and whether PCP predisposes to the development of fundic gland cysts (FGC). We therefore investigated the development of PCP and FGC in gastroesophageal reflux disease (GERD) patients treated with omeprazole and correlated findings to duration of therapy, gastrin, and H pylori infection. In a randomized, double-blinded study, GERD patients were evaluated by endoscopy with biopsy sampling for histology and culture at baseline, and after 3 and 12 months' therapy with omeprazole 40 mg daily. H pylori-positive patients were randomized to additional eradication therapy or placebo antibiotics at baseline. All histological slides were scored blinded for time and outcome of culture for the presence of PCP and FGC. Fasting serum samples from all visits were used for gastrin measurements. The prevalence of PCP increased during omeprazole therapy from 18% at baseline to 79% and 86% at 3 and 12 months (P < .001, baseline v both 3 and 12 months). The prevalence of FGC increased from 8% to 17% and 35% (P < .05, baseline v 12 months). The prevalence of PCP and FGC did not differ among the H pylori-positive and H pylori-negative patients at baseline (PCP 16% v 20% and FGC 7% v 8%, respectively). Whereas H pylori eradication did not significantly affect development of PCP (P = .7), FGC developed significantly more often in the H pylori-eradicated patients when compared with persistent H pylori-positive patients (P < .05). PCP development was related to serum gastrin rise during therapy. In conclusion, PCP occurs in most patients within the first months of omeprazole treatment and is related to increased gastrin levels. FGC develops more gradually and is enhanced by H pylori eradication. PMID- 10872662 TI - Clusterin/apoJ expression during the development of hemangioma. AB - Hemangioma is the most common tumor of infancy. This vascular tumor is characterized by an initial rapid proliferation followed by an inevitable regression. The life cycle of hemangioma is divided into proliferative, involuting, and involuted phases. The cellular and molecular mechanisms responsible for controlling the biological behavior of hemangioma are largely unknown. Differential display analysis using mRNA isolated from biopsy specimens representative of the 3 different phases showed increased expression of clusterin/apoJ (clust/apoJ) in the involuting samples. Clust/apoJ is a multifunctional glycoprotein that has been associated with apoptosis. Reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry showed that both the transcription and protein expression of clust/apoJ were increased in hemangioma as the tumor progressed from the proliferative to the involuting and involuted phases. This suggests that clust/apoJ is involved in regulating apoptosis during the spontaneous regression of hemangioma. It has been suggested that mast cells (MC) play a role in the regression of hemangioma. The increase in the number and proportion of clust/ apoJ-positive MC with progression of hemangioma, along with the localization of clust/apoJ to MC granules, supports this hypothesis. We suggest that MC may be synthesizing/releasing this apoptotic modulator, leading to the regression of the tumor. Better understanding of the pathogenesis of hemangioma by identification of the relevant factors involved in its regression such as clust/apoJ will result in the development of novel therapies for this condition and tumors that do not undergo spontaneous regression. PMID- 10872664 TI - Progression to invasive melanoma from malignant melanoma in situ, lentigo maligna type. AB - We have previously hypothesized that lesions that have been termed lentigo maligna can be divided into 2 categories: 1 represents a pigmented lesion that is a precursor to melanoma, and the other melanoma in situ. We and others have hypothesized that there is a progressive acquisition of attributes in pigmented lesions that results in malignant melanoma. Based on these 2 hypotheses, we have predicted that the intraepidermal component of invasive malignant melanomas, lentigo maligna type, should be similar to those lesions that we have termed malignant melanoma in situ, lentigo maligna type rather than lentigo maligna. The intraepidermal component of 42 consecutive cases of invasive malignant melanoma, lentigo maligna type was evaluated by all of the authors. Malignant melanoma in situ, lentigo maligna type is characterized by pagetoid spread, confluence, and nesting of atypical melanocytes. All of the cases evaluated showed features diagnostic of malignant melanoma in situ, lentigo maligna type, in the epidermis overlying the invasive dermal component. We conclude that invasive lentigo maligna melanoma arises in association with those lesions that we have termed malignant melanoma in situ, lentigo maligna type, which may represent a step in the progression between atypical melanocytic hyperplasia (lentigo maligna) and invasive melanoma. This finding supports the distinction of these entities and may have therapeutic implications. PMID- 10872663 TI - Overexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline ovarian tumors. AB - Ovarian epithelial tumors are classically divided into benign, malignant, and borderline or of low malignant potential. It is controversial whether this last group of tumors should be considered benign or malignant. Expression of cell cycle markers has recently been linked to tumor behavior and response to treatment. It has been shown that one of the pathways through which the p53 gene controls the cell cycle is by transactivating p21WAF1/CIP1, a cyclin-dependent kinase (cdk) inhibitor. By inhibiting cdks, p21WAF1/CIP1 blocks the G-1 to S phase transition in the cell cycle. p53 can be regulated by MDM2 (murine double minute-2) through direct inactivation or promotion of its cytoplasmic degradation. In an attempt to investigate the cell cycle checkpoint mechanisms of these tumors, we studied the expression of p53, Ki-67, MDM2, and p21WAF1/CIP1 by immunohistochemistry. We analyzed the expression of these proteins in 19 cystadenomas (8 serous and 11 mucinous), 40 borderline tumors (31 serous and 9 mucinous), and 18 serous carcinomas of the ovary. p21WAF1/CIP1 was expressed in 7 of 19 (37%) benign cystadenomas, 32 of 40 (80%) borderline tumors (93.5% of serous and 33% of mucinous), and in 9 of 18 (50%) serous carcinomas. Ki-67 was only weakly expressed in 8 of 19 (42%) benign cystadenomas, all borderline tumors showed Ki-67 staining in less than 50% of the cells, and 55% of serous carcinomas stained in more than 50% of tumor cells. p53 was absent in all but 1 of the cystadenomas, was expressed in 9 of 40 (22.5%) borderline tumors (25.8% of serous and 11% of mucinous), and in 10 of 18 (55%) carcinomas. All 11 implants of serous borderline tumors expressed p21WAF1/CIP1. Most serous borderline tumors expressed higher levels of MDM2 compared with the benign cystadenomas and carcinomas. Four of the serous borderline implants (40%) expressed MDM2. Coexpression of p21WAF1/CIP1 and MDM2 characterizes serous borderline tumors of the ovary and their implants, which suggests that these cell cycle control proteins are important in these tumors and may be related to tumor progression. Low expression of p53 protein in serous borderline tumors might be in part mediated by MDM2. This suggests that the p53 pathway is intact in most of these tumors, in contrast with carcinomas, in which high expression of p53 has been related to mutations of this gene. PMID- 10872665 TI - Comparison of histological changes and changes in nm23 and c-MET expression between primary and metastatic sites in osteosarcoma: a clinicopathologic and immunohistochemical study. AB - Changes in morphological features between the primary and metastatic sites in osteosarcoma and the role of nm23 protein and c-MET oncogene product have remained controversial. In addition to histological studies, we evaluated the expression of nm23, c-MET, p53, and MDM2 immunohistochemically using 25 osteosarcomas in which both primary and concordant metastatic specimens were available. Moreover, we assessed proliferative activity using the monoclonal antibody MIB-1. Among these 25 cases, 4 tumors that were osteoblastic type (16%) in the primary site had changed morphologically to MFH-like type in the metastatic site, whereas 2 MFH-like type and 1 small cell-type tumors had changed to osteoblastic type. MIB-1 LI was significantly higher in the metastatic site than in the primary site (primary, 20.02; metastatic, 26.72; P = .0209). Seventeen cases (68%) showed increased nm23 expression in the metastatic site, whereas 2 cases showed reduced expression. nm23 expression was significantly increased in the metastatic site, compared with the primary site (P = .0009). Seven cases (28%) showing negative reaction for c-MET in the primary site showed immunuoreactivity for c-MET in the metastatic site. Although there was no statistical significance, c-MET expression seemed to be more frequent in the metastatic site, compared with the primary site. Among the overall tumors, c-MET positive tumors showed significantly higher MIB-1 LI, compared with c-MET negative tumors (negative, 20.99; positive, 27.65; P = .0292). No significant change was observed regarding p53 and MDM2 between the primary and metastatic site. Our results suggest that rather than being a metastasis-suppressor gene, nm23 is in fact correlated with metastatic progression in osteosarcoma. Positive correlation between c-MET expression and proliferative activity also suggests that c-MET expression may play an important role in tumor progression in osteosarcomas. PMID- 10872666 TI - Dedifferentiated liposarcoma of retroperitoneum and mesentery: varied growth patterns and histological grades--a clinicopathologic study of 32 cases. AB - The purpose of this study was to obtain further information regarding cellular differentiation and proliferative characteristics of dedifferentiated liposarcoma (DDL) arising in the retroperitoneum and mesentery for accurate diagnosis and prognostic criteria. The patients included 20 men and 12 women, mean age, 60 years (range, 33 to 80 years). Twenty-seven tumors were located in the retroperitoneum and 5 in the mesentery. Tumor size ranged from 9 to 51 cm (mean, 24 cm). Follow-up was available on all patients and ranged from 4 to 243 months (mean, 64 months). Twenty-four (75%) patients developed local recurrences, 3 (9%) had distant metastasis, and 16 (50%) died of the disease. The predominant histology of dedifferentiation (DD) included fibrosarcoma or malignant fibrous histiocytoma (MFH) in 15 (47%), myxofibrosarcoma (myxoid MFH) in 5 (16%), mixed type in 10 (31%), and a whorling pattern in 2 (6%). Divergent differentiation, such as osseous, osteosarcomatous, chondrosarcomatous, and leiomyosarcomatous, was observed in 9 (28%). Immunoreactivity for vimentin, desmin, CD34, neurofilament, alpha-smooth muscle actin, p53, and MDM2 was observed in 32 (100%), 14 (44%), 8 (25%), 13 (41%), 14 (44%), 19 (59%), and 18 (56%) of DD areas, respectively. On the basis of a histological grading using MIB-1 (MIB-1 index range, 3% to 80%; mean, 27%) to replace mitosis counts (1 to 35/10 high power fields [HPF]; mean, 13/10 HPF), 16 tumors each were classified as low-grade (grade 2) and high-grade (grade 3). The mixed type with poorly differentiated areas including scattered lipoblasts could be mistaken for myxoid liposarcoma but lacked the C/EBP-homologous protein-translocated in liposarcomat (CHOP-TLS) fusion genes specific for myxoid liposarcoma. Period to the first recurrence and histological grade using the MIB-1 index were associated with overall survival. Identification of DD areas, especially a poorly recognized variant of the mixed type, careful follow-up to detect early recurrence, and histological malignancy grading combined with proliferation indices are important in providing an accurate prognosis for all patients with retroperitoneal and mesenteric liposarcoma. PMID- 10872667 TI - Expression of DNA toposiomerase I and DNA topoisomerase II-alpha in testicular seminomas. AB - DNA topoisomerase I (topo I) is the molecular target of the camptothecin group of anticancer drugs. These drugs are S-phase specific and require elevated topo I for tumor cell killing. To determine whether increased topo I expression occurs in testicular seminomas, 20 cases of testicular seminoma were retrieved from the surgical pathology files at the University of Utah Health Sciences Center and stained with an antibody that recognizes topo I in paraffin embedded human tissue sections. Topo I elevation was observed in 30% (6/20) of the seminomas. Because the response to topo I targeted drugs requires cell proliferation, the proliferative index of the seminomas was determined by immunohistochemical staining for DNA topoisomerase II-alpha (topo II-alpha) a new marker of cell proliferation. AU seminomas had easily detectable topo II-alpha. The average topo II-alpha index of the 20 cases was 52.1 +/- 15.3. Seminomas with elevated topo I had an average topo II-alpha proliferative index of 60.8 +/- 17.5 and seminomas with normal topo I expression had a topo II-alpha proliferative index of 48.4 +/- 13.2. This is significantly different at the 0.05% confidence level. Focal expression of CD30 was seen in 60% (12/20) of the neoplasms. None of the cases showed positive staining for CD15 and c-erbB-2. Our results suggest that chemotherapeutic protocols involving topoisomerase targeting drugs might be useful against testicular seminomas. PMID- 10872668 TI - Erdheim-Chester disease: evidence for a disease entity different from Langerhans cell histiocytosis? Three cases with detailed radiological and immunohistochemical analysis. AB - Erdheim-Chester (EC) disease is a rare pathological entity with a highly specific and characteristic pattern of radiographic bone changes. Histologically it resembles Langerhans cell histiocytosis (LCH), and it is still a matter of discussion whether EC disease is a distinct entity or a type of LCH. In this study, 3 cases of Erdheim-Chester disease were followed up over years and examined in detail both radiologically and immunohistochemically. All 3 cases showed the pathognomonic skeletal features for EC disease as well as an identical immunohistochemical phenotype quite different from LCH. Macrophages and Touton cells reacted strongly positive with the histiocytic marker CD 68, whereas staining with S100 and CD1a, markers for Langerhans cells, were negative. Both the immunohistochemical phenotype and the bone changes were clearly distinct from LCH. PMID- 10872669 TI - Adenoid basal carcinomas of the cervix: a unique morphological evolution with cell cycle correlates. AB - Adenoid basal carcinoma (ABC) is a rare cervical carcinoma of postmenopausal women composed of small basal-type (basaloid) cells with focal endocervical ("adenoid") differentiation. ABCs are associated with high-grade squamous intraepithelial lesions (HSIL) and contain integrated human papillomavirus type 16 DNA. However, ABCs have a favorable prognosis and do not metastasize. Five (5) ABCs were analyzed histologically for a marker distinguishing basal/ squamous from columnar (adenoid) differentiation (p63) and cell cycle activity (Ki-67), and compared with 20 cervical (CC) carcinomas. In contrast to other CCs, ABCs contained 4 distinct components, including (1) a classic HSIL; (2) a limited invasive component with squamoid maturation, often with a discrete layer of peripheral basal cells; (3) outgrowth of small basal cells from either HSIL or squamoid areas; (4) focal endocervical (adenoid) differentiation. ABCs showed distinct differences in cell cycle activity relative to CCs. Ki-67 positivity was high in associated HSILs but remained high and concentrated in the suprabasal cells of the invasive squamoid component of ABC. Moreover, proliferative index was variable to sharply reduced in areas of basaloid and adenoid differentiation, in contrast to conventional CCs. ABC is a unique neoplasm, not only by its transition through multiple phenotypes during invasion, but also by a proliferative index that is high in more mature neoplastic cells during the infiltrative process and reduced with progressive basal differentiation. The precise mechanism underlying this unique process of tumor evolution is unclear. However, the postmenopausal status of these patients suggests that host factors related to aging may influence tumor evolution and morphology after HPV 16 infection. PMID- 10872670 TI - Assessment of basal cell status and proliferative patterns in flat and papillary urothelial lesions: a contribution to the new WHO classification of the urothelial tumors of the urinary bladder. AB - In 1999, the World Health Organization (WHO) published a new classification of papillary urothelial tumors of the urinary bladder. Intended to represent a reproducible, easy-to-use classification system that better separates patients with true malignancies (bladder cancer) from those patients who are at an increased risk for developing bladder cancer, problems in the differential diagnosis of various lesions remained. Probably the most critical distinction is between papillomas, papillary urothelial neoplasms of low malignant potential (lmp), and grade I papillary carcinomas. Conversely, problems in the distinction between reactive atypia, atypia of unknown significance, and dysplasia, as well as the distinction of dysplasia from carcinoma in situ (CIS), are unresolved. Whether urothelial basal cell status assessment on hematoxylin and eosin-stained slides completed by cytokeratin immunohistochemistry with anticytokeratin clone 34betaE12 may help to improve some of the previously mentioned diagnostic dilemmas was investigated. Basal cell status assessment was helpful in the differentiation between dysplasia and CIS. In dysplasia, CK IHC showed a predominantly basal labeling pattern, whereas in CIS, labeling of all urothelial layers was seen. Basal cell status assessment could separate 2 groups of pTa GIb papillary carcinoma. Group 1 with a continuous basal CK labeling and a low MIB-1 labeling index (LI) was compared with group 2, with a diffuse labeling pattern and a significantly higher MIB-1 LI. Whether group 1 carcinomas should better be assigned to the group of papillary urothelial neoplasms of lmp is discussed. PMID- 10872671 TI - Cell cycle regulators in bladder cancer: a multivariate survival study with emphasis on p27Kip1. AB - Cyclin-dependent kinase inhibitors (CKIs) prevent cyclin-dependent kinases from phosphorylating critical substrates such as retinoblastoma gene protein (pRb), hence blocking the cascade of events leading to cell proliferation. Currently, the list of CKIs includes p21WAF1/Cip1, p27Kip1, p57Kip2 (the Cip/Kip family), p15/ INK4b, p16/INK4a, p18/INK4c, and p19/INK4d (the INK4 family). Among them, p27 plays a crucial role linking extracellular growth-regulatory signals to progression to or exit from the cell cycle. Unlike p53, p16, and Rb, mutations in Kip1 and WAF1 genes are distinctly rare in bladder cancer. We analyzed immunohistochemically the expression of p27 and other interacting G1 proteins (ie, p21, p16, pRb, p53) in 120 consecutive cases of transitional cell carcinomas (TCCs) and related it to proliferation rate, clinicopathologic parameters, and survival. p27 levels were significantly higher in low-grade (P = .001), superficial (Ta-T1) (P = .001), papillary (P < .001), and slowly proliferating TCCs (rs = -0.235, P = .05). p27 also positively correlated with p16 expression (rs = 0.212, P = .05). In univariate analysis, decreased p27 expression was associated with poor overall (P = .0109) and postrelapse (P = .0344) survival, especially if combined to increased Ki-67 expression (P = .0004 and P = .036, respectively). Furthermore, in multivariate analysis, Ki-67/p27 status had the strongest bearing on the overall survival of muscle-invasive TCCs (P = .0019). Our results indicate that low p27 expression is more common in poorly differentiated muscle-invasive TCCs and is a major player in cell cycle control in these neoplasms. More importantly, the combined Ki-67/p27 expression provides prognostic information beyond that provided by conventional parameters or other cell cycle-related proteins, concerning overall survival in muscle-invasive TCCs. PMID- 10872672 TI - Pure cystic nephroblastoma of the ovary with a review of extrarenal Wilms' tumors. AB - A second case of pure ovarian extrarenal Wilms' tumor (EWT) is presented. A clinical stage Ic tumor occurred in the right ovary of a 21-year-old female and corresponded to a 19-cm multilocular mass which histologically was a cystic, partially differentiated Wilms' tumor, closely resembling the highly differentiated metanephric adenoma. This pattern is reported for the first time in an ectopic location. At the interface between epithelial nests and ovarian tissue, plaques of alpha-inhibin positive cells were detected that corresponded to foci of peripheral stromal luteinization. Differential diagnosis with entities such as retiform Sertoli-Leydig cell tumors and with adenosarcoma should be made. The literature on EWT is also reviewed. PMID- 10872673 TI - Dilated cardiomyopathy in Noonan's syndrome: a first autopsy case. AB - The first autopsy case of dilated cardiomyopathy associated with Noonan's syndrome is described. A 9-month-old girl with Noonan's syndrome died from cardiac failure. Autopsy showed biatrial and biventricular enlargement of the heart. The posterior half of the ventricular septum and right ventricular wall were remarkably thin. Other parts of the wall were nearly normal in thickness, but both ventricular cavities were dilated. Microscopically, the myocardium of both ventricles consisted of mainly abnormally thinned, elongated, and loosely arranged myocardial fibers lacking immunoreactivity to anti-dystrophin antibody. Myocardial disarray was not found except for where it normally existed. The abnormal changes of the myocardial fibers were considered to be primary. Common cardiomyopathy associated with Noonan's syndrome is a hypertrophic type. Various types of cardiovascular abnormality associated with Noonan's syndrome, including dilated cardiomyopathy, might be disclosed by further investigation and precise diagnosis of Noonan's syndrome. PMID- 10872674 TI - Vacuolated cell mesothelioma of the pericardium resembling liposarcoma: a case report. AB - We report a case of localized pericardial mesothelioma with unusual histological features in a 44-year-old woman. Her radiological imagings showed an 11-cm pericardial tumor, between the heart and aortic arch. Microscopically, the tumor was predominantly composed of vacuolated cells and vaguely reminiscent of well differentiated "lipoma-like" liposarcoma, but only small foci of the tumor showed the papillotubular configuration. Histochemically, the tumor cells contained hyaluronic acid in the vacuoles but no lipids. Immunohistochemically, they showed immunoreactivity for cytokeratin, calretinin, vimentin, and epithelial membrane antigen. Ultrastructural study showed that the vacuoles of the tumor cells were intracytoplasmic lumina. The intracytoplasmic lumina and the surface membranes of the tumor cells had many long and slender microvilli with focal bush-like appearance. Desmosomes between adjacent cells were occasionally observed. To our knowledge, this is the first case report of epithelial type mesothelioma predominantly composed of vacuolated tumor cells, microscopically mimicking liposarcoma. PMID- 10872675 TI - Chromosomal translocations are common in natural killer-cell lymphoma/leukemia as shown by spectral karyotyping. AB - Natural killer (NK)-cell lymphoma/leukemia is a group of rare but highly aggressive neoplasms. The associated genetic aberrations, as defined by conventional cytogenetics, include 6q deletion and chromosome X copy gain, while translocations have been suggested to be uncommon. In this study, three cases of NK cell lymphoma/ leukemia were investigated by spectral karyotyping (SKY). SKY permitted reinterpretation of the chromosomal alterations defined by G-banding and identified several cryptic translocations. In agreement with G-band, 6q deletion was detected in all 3 cases. Structural rearrangement involving chromosome X was observed in 2 cases, and fluorescence in situ hybridization (FISH) analysis indicated that both translocations involved Xp21-pter. Chromosome 8 translocation was also identified in 2 cases and shared a common breakpoint, 8p23. The present study shows the value of SKY in providing additional information on karyotypic abnormalities. The novel findings of recurring Xp21 pter rearrangements and 8p23 translocation should provide basis for further investigations into the tumorigenesis of NK cell lymphoma/leukemia. PMID- 10872676 TI - ASAP. PMID- 10872677 TI - Cytological recognition of invasive squamous cancer of the uterine cervix. PMID- 10872678 TI - Proliferation center cells in the lymph nodes of B-cell chronic lymphatic leukemia express relatively higher levels of CD20. PMID- 10872679 TI - Role of IL-15 on monocytic resistance to human herpesvirus 6 infection. AB - Interleukin-15 (IL-15) is a cytokine that possesses a variety of biological functions, including stimulation and maintenance of cellular immune responses. Recently, it has been demonstrated that Human Herpes virus type 6 (HHV-6) enhances NK activity of human PBMC by inducing IL-15. HHV-6 is a typical immunosuppressive agent, as suggested by its tropism for both CD4+ and CD8+ T cells, B cells, monocytes/macrophages, megakaryocytes and NK cells. Moreover, several studies have indicated that mononuclear phagocyte resistance to virus infection is influenced by the cellular differentiation state. This paper describes the effect of pretreatment "in vitro" with IL-15 on the resistance of human monocytes (HM) to HHV-6 infection. Our results demonstrate that undifferentiated HM were highly resistant to HHV-6 infection, whereas HM pretreated with human recombinant IL-15 showed an increased permissiveness for HHV-6 infection. This permissiveness was characterised by higher release of extracellular virus as well as an increased percentage of antigen positive cells. Moreover, we evaluated IL-15 production after the addition of HHV-6 to monocytes precultured in different experimental conditions. Our data indicate that HHV-6 induced IL-15 production by human monocytes is not affected by the condition of "in vitro" precultivation/differentiation. Furthermore, the neutralization of IL 15 induced by HHV-6 in differentiated monocytes did not affect viral replication. These findings suggest that IL-15 acts only on the mechanisms of cellular differentiation, rendering HM more susceptible to HHV-6 infection, without interfering with virus replication. PMID- 10872680 TI - Characterization of immune response to synthetic peptides derived from the hemagglutinin of measles virus. AB - We describe here four synthetic peptides derived from the hemagglutinin of measles virus. The peptides were predicted by a computer program combining hydrophilicity, flexibility, surface probability, secondary structure and antigenic index parameters of the amino acid sequence of measles virus hemagglutinin. Rabbits were immunized with the synthesized peptides conjugated to purified protein derivative using immunostimulating complex as adjuvant. Anti peptide antisera raised in rabbits against the peptide conjugates reacted well with the homologous peptides and with measles virus antigen as tested with plate ELISA. None of these sera had either neutralizing or hemagglutination inhibiting antibody or reacted with measles hemagglutinin protein in Western blot and reacted weakly in immunofluorescence. Human sera positive for measles virus antibody reacted with the synthesized peptides indicating that the selected locations function as partial antigenic sites. PMID- 10872681 TI - Expression of measles virus antigens in Streptococcus gordonii. AB - The measles virus proteins haemagglutinin (HA) and fusion protein (F), which together mediate attachment and penetration of the virus in the host cell and can elicit production of neutralising antibodies in the course of natural infection were expressed in the vaccine vector Streptococcus gordonii, a Gram-positive bacterium normally present in the human oral cavity. HA and F were expressed as fusion proteins attached to the bacterial surface, and were both found to be immunogenic when the recombinant S. gordonii were inoculated subcutaneously in mice. PMID- 10872682 TI - Transfection of bovine cell culture with bovine herpesvirus 4 DNA obtained by cell nuclear extraction. AB - Bovine herpes virus 4 (BHV-4) is a gamma-herpesvirus not associated with clearly defined clinical entities in cattle. The BHV-4 genome consists of a linear dsDNA of approximately 145 Kbp which is only partially characterized and sequenced. We set up a rapid and practical method to isolate BHV-4 DNA from infected cell culture. Microfuged infected cells after exposure to high salt concentration and detergent allowed viral DNA to be purified. Electrophoretic analysis of the digested DNA showed a complete digestion, corresponding to a classical EcoRI banding pattern of strains Movar 33/63, LVR and DN 599. Moreover the biological integrity of viral DNA here obtained, was demonstrated by transfection experiments. BHV-4 DNA was capable of forming CPE after transfection into BAE 7372 cells. Transfected cells specifically reacted with a BHV-4 infected cow serum demonstrating the presence of viral particles. The possibility of obtaining infectious viral DNA using this method may facilitate the construction of recombinant viruses. Specifically, through the use of cotransfection experiments with deleted or mutated viral DNA sequences, the infectious clones isolated could provide the basis for an increased understanding of BHV-4 viral gene expression, replication and pathogenesis. PMID- 10872683 TI - Influence of antimicrobial subinhibitory concentrations on hemolytic activity and bacteriocin-like substances in oral Fusobacterium nucleatum. AB - Fusobacterium nucleatum is considered for its role in colonization of initial and late microorganisms in dental plaque and for its coaggregation with other bacterial species. It is known that action of different antimicrobial substances may interfere with either virulence factors or with host-bacteria interaction. The goal of this study was to examine the influence of subinhibitory concentrations of chlorhexidine, triclosan, penicillin G and metronidazole on hemolytic activity and bacteriocin-like substance production of oral F. nucleatum. A high resistance to penicillin G was observed and 63% of the isolates were beta-lactamase positive. All the tested isolates were susceptible to metronidazole. F. nucleatum isolates grown with or without antimicrobials were alpha-hemolytics. Bacteriocin-like substance production was increased in isolates grown with penicillin G. Impaired production of hemolytic or antagonic substances can suggest a role in the regulation of oral microbiota. PMID- 10872684 TI - Isolation of Arcobacter spp. from a brackish environment. AB - Arcobacter spp. were isolated from water and mussels of two brackish lakes near Messina (Italy). The isolates were phenotypically characterized on the basis of a large battery of cultural and biochemical tests. By comparison with the reference strains Arcobacter butzleri ATCC 49616 and A. cryaerophilus ATCC 43157 they may be considered Arcobacter butzleri-like bacteria. The current isolation suggests that the brackish environment may play an important role in the survival and transmission of Arcobacter spp. also by seafood cultured in the examined waters. PMID- 10872685 TI - Comparison of the mycobacteria growth indicator tube with radiometric and solid culture for isolation of mycobacteria from clinical specimens and susceptibility testing of Mycobacterium tuberculosis. AB - We compared the mycobacteria growth indicator tube (MGIT) system with the BACTEC 460 TB and Loewenstein-Jensen (LJ) systems for the recovery of mycobacteria (acid fast bacilli [AFB]) from 600 clinical specimens. A total of 50 AFB (32 Mycobacterium tuberculosis complex, 10 M. avium complex, 3 M. gordonae, 3 M. xenopi, 1 M. terrae and 1 M. fortuitum) were detected. MGIT recovered 50 isolates of AFB (100% sensitivity), and BACTEC 460 TB and LJ recovered 49 (98% sensitivity) and 19 (38% sensitivity) AFB isolates, respectively. The mean times to detect mycobacteria were 10, 10 and 25 days for MGIT, BACTEC 460, and LJ slants. All isolates of M. tuberculosis complex were tested for susceptibility to streptomycin, isoniazid, rifampin, and ethambutol with the MGIT and BACTEC 460 TB. Both systems yielded identical susceptibility data with different mean times to report (5.38 days for MGIT versus 7.33 days for BACTEC 460 TB, P<0.05). The results suggest that MGIT is equivalent to BACTEC 460 TB in its ability to support the growth of mycobacteria, but significantly more efficient than LJ. MGIT may also be used for susceptibility testing of primary antituberculosis drugs. PMID- 10872686 TI - Characterization of iap gene in Listeria monocytogenes strains isolated in Japan. AB - Variation of the iap gene region (407bp) encoding an invasion-associated protein p60 was studied on 12 strains of Listeria monocytogenes of different origin in Japan. These 12 strains are known to have 2 types of serotype (1/2a and 4b) and have a diversity among the strains (Saito et al., 1998). The dye-primer cycle sequencing method was employed to determine the genomic structure, and the nucleotide sequences obtained were compared with those of reference strain SV 1/2a EGD. Differences found in the nucleotides were as follows; point mutations of 33 variations in 32 places; an insertion and 3 deletions of 3 bases; AAT position (po.) 1282-1283, and GCA po. 1307-1309, ACA po. 1412-1414, AAT po. 1439 1444, respectively. Different repeating numbers by 6 base unit, ACA AAT, were also found in the tandem repeat region (po. 1394-1423). Classification of 12 strains was attempted, then 8, 4 and 5 types were obtained from the point mutations, the insertions and deletions, and the repeating numbers, respectively. Consequently, 8 patterns were profiled regardless of each serotype. From these results, genomic structures were partially clarified in the iap gene 407bp of L. monocytogenes isolated in Japan. Then, the possibility of detailed epidemiology for L. monocytogenes infection using a combination of serotype and genome structure was suggested because of the previous polymorphism thought to be due to the nucleotide differences in the region. PMID- 10872687 TI - Interactions between cytokines and growth hormone for resistance to experimental. AB - Cytokine-activated human vein endothelial cells (HUVEC) may play an important role in resistance to Toxoplasma gondii infection. In this study, it was investigated the role of rTNF-alpha and GH in the induction of antitoxoplasmal activities in HUVEC. Co-treatment of HUVEC with rTNF-alpha plus GH induced both toxoplasmastatic activity and the intracellular killing of T. gondii (p <0.01 each vs untreated cells). Thus, these functions were inhibited by both neutralizing antibodies to IL-6 and GM-CSF (but not to IL-3) suggesting that these cytokines participate in the inhibitory process. Consistent with this hypothesis, the treatment of HUVEC with rIL-6 or rGM-CSF in the presence of rTNF alpha, limited T. gondii multiplication in a dose-dependent manner (p <0.01 each vs untreated cells). In order to elucidate the inhibitory mechanism of HUVEC, it was assessed by L-arginine analogs (e.g., NG-monomethyl-arginine) whether NO2 molecules originating from HUVEC were directly or indirectly involved in the rTNF alpha/GH-dependent induction of toxoplasmastatic activity. A good correlation was found between toxoplasmastatic activity and NO2 release during the activation phase, before infection of the HUVEC with T. gondii, but no correlation was found between the parasitostatic activity and NO2 release during the infection phase. These data indicate that NO2- itself does not directly affect toxoplasmastatic activity. Besides, the reduction of intracellular killing by monoclonal antibodies to ICAM-1 suggest that this adhesin plays a role in controlling T. gondii entry into cells. PMID- 10872688 TI - Interactions of invasive and noninvasive strains of Neisseria meningitidis with monkey epithelial cells, mouse monocytes and human macrophages. AB - Adherence and phagocytosis of invasive and noninvasive Neisseria meningitidis strains was investigated using light, fluorescence and electron microscopy. Invasive strains were isolated from the cerebrospinal fluid and/or blood of the patients with invasive meningococcal disease and noninvasive strains from the nasopharynx and/or larynx of healthy carriers. Adherence/endocytosis was studied on monkey kidney cells (the LLC-MK2 cell line) and phagocytosis on mouse monocytes and human macrophages (the P388D1 and U-937 cell lines, respectively). Although invasive and noninvasive meningococci isolated in the same cluster showed identical genotype and phenotype markers, they were found to interact differently with epithelial cells as well as with monocytes/macrophages. Invasive isolates displayed higher adherence to the surface of LLC-MK2 cells compared to noninvasive ones. Phagocytosis by P388D1 cells of noninvasive strains was effective and the bacteria were damaged by cytolysis. In contrast, invasive bacteria frequently persisted in "coiling" vacuoles and in effect could destroy the host cell. This is the first demonstration of coiling phagocytosis induced by meningococci. Efficiency of phagocytosis by U-937 cells was significantly higher for the noninvasive than invasive strains. Different behaviour of invasive and noninvasive strains of N. meningitidis observed during 4 hours of interactions with epithelial cells and monocytes/macrophages reflects well the higher pathogenic potential of invasive bacteria. PMID- 10872689 TI - Staphylococcus sciuri: recommendation for simple identification. AB - We report the isolation of Staphylococcus sciuri, primarily animal species, from samples taken from hospitalised patients. Considering that Staphylococcus sciuri often remains unrecognised in routine laboratory practice, we propose the criteria for simple identification of this bacterium. PMID- 10872690 TI - Microbiological investigation on black crusts from open-air stone monuments of Bologna (Italy). AB - Samples of black crusts taken from open-air stone monuments in Bologna dating back to between the twelfth and the nineteenth century were considered. The microbiological test procedures isolated from a high number of chemoorganotrophic bacteria, each of the samples although it was expected to find chemo-lithotrophic (specially sulfur-oxidizing and nitrifying) bacteria on account of the great amounts of sulphur, carbon, nitrogen dioxides and ammonia in the air. Sporogenous and asporogenous species were found among the isolated bacteria: the first were recognized as Bacillus subtilis, Bacillus brevis, Bacillus licheniformis, Bacillus mycoides and Bacillus megaterium; the latter were Corynebacterium glutamicum, Actinomyces sp., Alcaligenes denitrificans subsp. denitrificans, Flavobacterium breve, Acinetobacter calcoaceticus, Enterobacter cloacae, Pseudomonas stutzeri. These species were differently associated in the samples studied with a frequency of more than sixty-four per cent. This finding leads us to believe that their presence in the black crusts is not occasional, but the result of an ecologically significant process of colonization. Although behavioural tests (solubilization of calcium carbonate, of calcium sulphate and marble dust) did not reveal lithic activity among the bacteria studied, one can presume that their behaviour in nature - as a more or less complex microbial consortium - can be totally different and exert a decomposing action with regard to several minerals, most of them acidify the media in which they develop. PMID- 10872691 TI - Interpretation and diagnostic implications of an immunoblotting (INNOLIA) test for HCV serology. AB - The association analysis of antibodies versus HCV, carried out with INNOLIA test, prevented a clear determination of the existence of specific serological patterns. In this respect, it may be of interest to monitor the immune response to the non-structural genomic regions (NS3, NS4, NS5). The INNOLIA kit is reliable, but susceptible to improvement in terms of specificity, sensitivity and biological standardization. PMID- 10872692 TI - Who determines the research agenda? PMID- 10872693 TI - Prevention of cardiovascular diseases--a scientific dilemma. PMID- 10872694 TI - Brain protection during aortic arch surgery. PMID- 10872695 TI - Intermittent retrograde cerebral perfusion during prolonged period of hypothermic circulatory arrest: a study in a chronic porcine model. AB - Previous studies have shown that although retrograde cerebral perfusion (RCP) improves cerebral outcome during hypothermic circulatory arrest (HCA), RCP exposes the brain to subsequent edema. In this study, we have compared intermittent RCP (I-RCP) with continuous RCP (C-RCP) and HCA alone to determine whether the rate of fluid sequestration can be decreased without losing the beneficial effects of RCP. Eighteen pigs were randomly assigned to undergo 75 min of I-RCP, C-RCP or HCA at 20 degrees C. Hemodynamic and metabolic measurements were carried out for upto 20 h. Behavioral assessments were examined until day 7, when histopathologic analysis of the brain was performed. The median amount of fluid sequestered was 145 ml after C-RCP and -50 ml after I-RCP (p = 0.04). The mean brain weight of the animals that died within the first postoperative day was significantly higher than that in electively sacrificed animals in the C-RCP group (p = 0.04). These data suggest that if RCP is implemented intermittently, the rate of cerebral edema can be decreased, without compromising the benefits of this strategy. PMID- 10872696 TI - Management of extensive lesions of the aortic arch: review of 42 consecutive cases. AB - This retrospective study reviews the long-term results in aortic arch surgery. Forty-two consecutive patients (44-74 years) were operated on between 1980 and 1995. Nineteen patients had acute dissections of the aortic arch, 11 had chronic dissections and 12 had aneurysms. Twenty patients were given emergency surgery and 22 were operated on electively. Reconstruction of the aortic arch alone (31%) or with other aortic segments (69%) was performed during antegrade cerebral perfusion (81%) or circulatory arrest (17%). Sixteen patients died during the first 30 days (38%). Early mortality rates were 60% in emergencies and 18% in elective cases. Early mortality in the group with aneurysms was 33% and 40% in patients in the dissection group. Long-term follow-up to July 1998 is now complete. Twelve patients (28%) died during the follow-up period; 4 of these deaths were due to a rupture of the descending aorta. Three patients (11.5%) underwent late reoperations on the remaining aorta, without operative mortality. We recommend careful follow-up of patients with aortic arch disease and, when indicated, surgery to avoid the rupture of the remaining aorta. PMID- 10872697 TI - The value of predischarge ICD tests in patients with a successful peroperative test. AB - An internal cardioverter defibrillator (ICD) is normally extensively tested during implantation. The necessity of retesting prior to discharge of the patient is a matter of debate. In our material of 30 patients undergoing first-time implantation of a transvenous internal defibrillator system, we retrospectively compare the predischarge defibrillation test with the peroperative test. A successful peroperative defibrillation test with no failed shocks at 10 J below maximal energy level was followed by a successful predischarge test with the same safety margin in 18/19 patients, while one patient required a maximal energy ICD shock for conversion at the predischarge test. We conclude that the predischarge defibrillation test can be omitted if the peroperative test was successful, with no failed shocks at 10 J below maximal energy level and if the shock therapy is set to maximal energy level. PMID- 10872698 TI - Adaptation to myocardial ischemia during repeated ventricular pacing in patients with coronary artery disease. AB - OBJECTIVE: The purpose of our study was to evaluate whether repeated ventricular pacing is able to induce adaptation against ischemia in coronary artery disease patients. DESIGN: Fifteen patients with documented coronary artery disease were subjected to two successive periods of rapid ventricular pacing (150 bpm) of equal length (295+/-33 s), the first being limited by intolerable anginal pain. The second pacing period, of the same length as the first, was initiated after the disappearance of angina and ST depression, the mean resting time being 433+/ 30 s. Blood samples for the determination of transcardiac differences in glucose, lactate, free fatty acids, K+, pCO2, pH, oxygen saturation and noradrenaline were taken from the femoral artery and coronary sinus before and at the end of each pacing period. The mechanical performance of the hearts was followed by continuous monitoring of intra-arterial blood pressure and pulmonary capillary wedge pressure, and the observed adaptation in the measured variables during the successive pacing tests was correlated with the duration of angina, severity of coronary artery disease and degree of collateralization. RESULTS: Changes in the transcardiac pH and K+ differences, ST segment and pulmonary capillary wedge pressure were less pronounced during the second pacing period. The subgroup with net lactate production before or after the first pacing period demonstrated metabolic adaptation manifested as improved lactate extraction during the second pacing period. Rate-pressure product and oxygen extraction, and thus presumably also overall oxygen consumption and oxygen delivery, were similar during both tests. The magnitude of adaptation did not correlate with the duration of angina, severity of coronary artery disease or overall collateral score. CONCLUSION: Rapid ventricular pacing is able to induce adaptation to myocardial ischemia, but the exact mechanisms in this process remain to be elucidated. PMID- 10872699 TI - Assessment of myocardium at risk in pigs with single photon emission computed tomography and computerized vectorcardiography during transient coronary occlusion. AB - Since myocardium at risk (MAR) is the major prognosticator of final infarct size and outcome in patients with acute myocardial infarction, it is highly desirable to estimate the size of the acutely ischemic myocardium, that is the MAR, in these patients. We assessed MAR size by Tc-99m-sestamibi-SPECT and computerized vectorcardiography using autoradiography as reference method. Transient myocardial ischemia was achieved in 12 pigs by coronary artery occlusion with PTCA catheters. During the procedure, computerized vectorcardiography was continuously recorded. After injection of Tc-99m-sestamibi and gadolinium-153 labelled microspheres, MAR size was estimated by SPECT and post-mortem autoradiography. Different cut-off levels (50-70%) were compared with respect to MAR-SPECT. Tc-99m-sestamibi-SPECT showed a good correlation with autoradiography (r = 0.94). Computerized vectorcardiography showed a good correlation with autoradiography as well as with Tc-99m-sestamibi-SPECT (STC-VM: r = 0.75 and 0.80, respectively, ST-VM: 0.75 and 0.87, respectively). It was found that 1) MAR assessed by Tc-99m-sestamibi-SPECT correlates closely with the autoradiographic reference; 2) a lower cut-off point of 60% of maximum uptake for MAR by Tc-99m sestamibi-SPECT gives the closest correlation with the autoradiographic reference; and 3) ST-VM and STC-VM correlate well with MAR assessed by Tc-99m sestamibi-SPECT and autoradiography. PMID- 10872700 TI - Postoperative nitric oxide therapy in children with congenital heart disease. Can the need be predicted? AB - The necessity for postoperative inhaled nitric oxide (NO) therapy and predictive factors for that need were retrospectively analysed in 457 paediatric patients at risk of pulmonary hypertensive events following open-heart surgery for congenital heart disease. Inhaled NO was given postoperatively to 46% of the study group and to 23% of all patients undergoing open-heart surgery during the study period. Factors associated with increased need for postoperative NO were age <1 year, Down's syndrome, preoperative pulmonary hypertension and increased pulmonary vascular resistance. Using a multivariate model based on these factors, 73% of the patients who were given NO were identified. Thus, in a setting with unrestricted access to NO therapy, almost half of the patients with cardiac lesions that commonly give rise to postoperative pulmonary hypertension were given postoperative NO. Seventy-three percent of postoperative NO treatment was associated with a relatively small number of pre- and perioperative patient related risk factors. PMID- 10872701 TI - Inhaled nitric oxide to newborns and infants after congenital heart surgery on cardiopulmonary bypass. A dose-response study. AB - Twelve patients (median age 3.8 months) with pulmonary hypertension in the postoperative period after congenital heart surgery on cardiopulmonary bypass were given inhaled nitric oxide. Effects on cardiovascular and respiratory systems were measured. Mean pulmonary artery pressure decreased from 33+/-2 to 28+/-2 mmHg (p < 0.001) and arterial oxygen tension increased from 13.3+/-2.3 to 16.7+/-2.7 kPa (p < 0.05). The mean change in arterial oxygen tension in percent was 29.8+/-6.3% (p < 0.05). The response was significant only in the first step from 0 to 3 or 5 ppm with no further significant changes in mean pulmonary artery pressure or oxygenation at higher doses. The decrease in mean pulmonary artery pressure was concomitant with a significant increase in arterial oxygen tension. No dose-response relationship was found with increasing doses to 80 ppm. PMID- 10872702 TI - Elevated urinary albumin excretion is not linked to the angiotensin I-converting enzyme gene polymorphism in clinically healthy subjects. AB - An elevated urinary albumin excretion (UAE) in non-diabetic subjects without renal or cardiovascular disease has been shown to be predictive of ischaemic heart disease. An insertion (I)/deletion (D) polymorphism in the angiotensin I converting enzyme (ACE) gene has been identified and the D allele may be associated with cardiovascular disease. The aim of this study was to find a potential linkage between this polymorphism and elevated UAE. For studies of UAE and cardiovascular pathophysiology, a highly selected population sample has been identified comprising all clinically healthy subjects aged 40-65 years with elevated UAE in a dipstick negative urinary sample (n = 27) from The Copenhagen City Heart Study. Neither the ACE genotype distribution (p = 0.12) nor the D and I allele frequencies (p = 0.69) differed significantly between subjects with elevated UAE and a matched normoalbuminuric control group (n = 46). Elevated UAE in clinically healthy subjects is not linked to the ACE gene polymorphism. PMID- 10872703 TI - Has interest in secondary prevention increased among physicians after the 4S study? AB - This study evaluates the occurrence of various risk indicators, with particular emphasis on serum lipids one year after a coronary event (development of acute mycoardial infarction (AMI); exposure to either coronary artery bypass grafting (CABG) or percutaneous transluminal coronary angioplasty (PTCA), prior to and after presentation of the main results from the 4S study. Patients under 70 years of age either hospitalized for AMI or undergoing CABG or PTCA at Sahlgrenska University Hospital in Goteborg were evaluated one year after the event. Patients who had an event during the period January 1, 1993 until December 31, 1993 were evaluated one year later (Period I) and those who had an event during the period September 1, 1995 until August 31, 1996 were evaluated one year thereafter (Period II). In total, 293 patients were evaluated during Period I and 284 during Period II. Mean total serum cholesterol levels fell from 6.2 mmol/l during Period I to 5.3 mmol/l during Period II (p < 0.001). The proportion of patients with serum cholesterol < or =5.0 mmol/l increased from 15% during Period I to 40% during Period II (p < 0.001). The mean low-density lipoprotein (LDL) levels fell from 4.0 mmol/l during Period I to 3.2 mmol/l during Period II (p < 0.001). The proportion of patients with LDL < or =3.5 mmol/l increased from 32% during Period I to 68% during Period II (p < 0.001). The proportion of patients using lipid lowering drugs increased from 25% during Period I to 57% during Period II (p < 0.001). Among patients with coronary artery disease who had either developed AMI or undergone CABG or PTCA, a marked increase in the use of lipid-lowering drugs has been observed in a university hospital in Sweden after presentation of the results of the 4S study. Parallel to the increased use of lipid-lowering drugs, we observed a substantial lowering of serum lipids. PMID- 10872704 TI - Effects of warfarin, aspirin and the two combined, on mortality and thromboembolic morbidity after myocardial infarction. The WARIS-II (Warfarin Aspirin Reinfarction Study) design. AB - The efficacy and safety of warfarin, aspirin, and the two combined are compared in a long-term, randomized, open, multicentre study involving 3606 patients after acute myocardial infarction (1202 in each treatment group). In this trial three groups receive either warfarin, aimed at a therapeutic level of the International Normalized Ratio (INR) 2.8-4.2, or 160 mg aspirin daily, or 75 mg aspirin daily combined with warfarin with INR 2.0-2.5. A placebo group is not included. Patients are screened before randomization and are given major examinations at 4 weeks and at the end of the study. In addition, all patients are given a questionnaire every 6 months. Composite endpoints include death, non-fatal reinfarction and cerebral stroke. All analyses are conducted on the intention-to treat principle and on on-efficacy basis. The analyses control for recruiting centre, use of beta blockade, use of thrombolytic therapy and use of angiotensin converting enzyme (ACE) inhibitors. PMID- 10872705 TI - Early diagnosis and exclusion of acute myocardial infarction by two hours' vector ECG and determination of either myoglobin or CK-mb. BIOMACS-study. BIOchemical Markers in Acute Coronary Syndromes. AB - This retrospective study assesses the early diagnostic potential of a combination of multilead continuous vectorcardiography (VCG) and biochemical markers (myoglobin, troponin-t and CK-mb mass) in patients with chest pain who present with suspected acute myocardial infarction (AMI), but without ST-elevation on resting 12-lead ECG on admission. Within a multicenter study 56 patients admitted for chest pain (< 12 h) and with a non-diagnostic 12-lead ECG on admission and a VCG recording were included. Venous blood samples were drawn on admission and the continuous VCG was monitored for 2 h. The results were related to the clinical diagnosis of AMI. Neither the biochemical markers nor VCG alone permitted the diagnosis or exclusion of AMI at admission. However, if either analysis of myoglobin on admission or 2 h of VCG recording were positive, they would have a sensitivity for detection of AMI of 100% and specificity of 69%. In a subset of patients with more than 4 h delay since start of chest pain, CK-mb could replace myoglobin and give a sensitivity of 100% and a specificity of 81%. Determination of myoglobin or CK-mb at admission and VCG monitoring for 2 h can reliably confirm or exclude AMI within 2 h. This combination seems useful for early stratifications of patients in chest pain or coronary care units. PMID- 10872706 TI - Clinical outcome after successful coronary angioplasty: impact of intracoronary stent implantation. AB - The clinical outcome after successful conventional coronary balloon angioplasty is compared with that of stent implantation after 30 days and 12 months. The study took place at the Divisions of Cardiology and Thoracic Radiology, Norrland University Hospital, Umea, a referral centre for northern Sweden. The first 100 consecutive patients with stable or unstable angina undergoing successful percutaneous transluminal coronary angioplasty (PTCA) in 1994 and the first 100 consecutive patients undergoing successful coronary stent implantation in 1995 were included. The cardiac endpoints studied were death, myocardial infarction, need for repeat PTCA or coronary artery bypass grafting (CABG). Significantly more adverse cardiac events were observed in the PTCA group compared with the stent group. Event-free 12 months' follow-up (no deaths, myocardial infarction or re-intervention) was 64% in the PTCA group and 86% in the stent group (p < 0.005). The main explanation for the observed difference was a reduction in the need for a repeat PTCA (7 vs 18, p < 0.05) or CABG (4 vs 12, p < 0.05) in the stent group. Patients with stable or unstable angina who can be treated with a stent have a better clinical outcome than those treated with coronary balloon angioplasty only. PMID- 10872707 TI - Six months' clinical and angiographic follow-up of a flexible, coiled stainless steel stent in long, native coronary artery lesions. AB - This study was conducted to evaluate the short- and long-term clinical and angiographic results of implantation of a flexible, coiled stainless steel stent, the Freedom Coronary Stent. During the study period this stent was used as an alternative to the Palmaz-Schatz PS153 coronary stent in long or tortuous lesions. The study was designed as a prospectively planned outcome analysis. Implantation of Freedom stents was attempted in 62 consecutive patients (56% males, mean age 63+/-10 years) with a total of 65 coronary lesions. Indications for stent implantation were: restenosis, 8%; recoil, 26%; visible dissection, 32%; threatening occlusion, 15%; chronic total occlusion, 18%. The average stent length was 30+/-16 mm and 67% of the lesions were type C. Rate of successful stent implantation, acute complications, angiographic restenosis after 6 months and major cardiac events (death, myocardial infarction, target vessel revascularization) during follow-up were assessed. The success rate of stent implantation was 94%. One patient died after an emergency bypass operation and one patient suffered a subacute stent thrombosis, which was successfully treated with re-percutaneous transluminal coronary angioplasty (PTCA). There were no Q- or non-Q myocardial infarctions. Clinical follow-up was carried out in 56 patients (97%) and 57 vessels were assessed by angiography (93%). Mean length of the follow-up period was 6.8+/-2.3 months. During the 6 months' follow-up period, one patient died, two patients suffered an acute non-Q myocardial infarction and eight patients had revascularization of the target vessel. Major cardiac event rate for all patients where stent implantation was intended was 23%. Angina CCS class declined from 3.0+/-0.9 to 1.1+/-0.8 (p < 0.01) before PTCA to follow-up. Overall restenosis rate was 28%. In 14 lesions with a stented segment length of <20 mm, the restenosis rate was 21%; in 31 lesions with a stented segment length > or =20 and <30 mm, the restenosis rate was 26%; and in 13 lesions with a stented segment length of > or =30 mm, the restenosis rate was 42%. Although there was a high procedural success rate after implantation of the Freedom stent in long or tortuous lesions, problems with high restenosis rates in long lesions remain unresolved. PMID- 10872708 TI - Duration of preoperative electrocardiographic QRS complex and the incidence of heart arrest after aorto coronary bypass surgery. AB - Sudden heart arrest (HA) in the early phase after aorto coronary bypass surgery represents a serious event necessitating resuscitation, and for those who survive usually also an extra stay in the coronary care unit. Since such episodes of heart standstill may be related to conduction defects, a study was conducted to determine whether the duration of the QRS complex on the preoperative ECG is a marker for this morbid event. A cohort of 1011 consecutive patients operated on between 1982 and 1986 and followed to January 1st, 1993 were included in the study. Incidence of lethal or non-lethal HA during the first 4 weeks after surgery was considered as the primary endpoint and total mortality as the secondary endpoint. The incidence of HA was 40/1011 = 4%, with the majority of events (60%) being lethal. Independent risk factors of HA using the multivariate logistic model were previous coronary artery bypass surgery, presence of mitral regurgitation, left ventricular ejection fraction and the intraoperative cross clamp time of aorta. Adjusting for the effect of confounder variables showed that the gradient effect of QRS complex duration on the endpoint HA was still present (p = 0.012). The duration of the QRS complex taken from the preoperative ECG had a gradient effect on the incidence of HA. With a baseline level of QRS <70 ms, the following odds ratios (OR) for HA were found: OR = 1.38 (95% CI 0.60-3.31) for QRS 70-80 ms; OR = 2.27 (95% CI 0.87-5.90) for QRS >90-120 ms; and OR = 3.38 (95% CI 1.06-11.50) for QRS > 120 ms, when adjusting for the risk factors. Cumulative survival at 5 years after surgery was 28+/-7.1% for patients experiencing HA versus 87+/-1.2% for patients free from this event. Our results underline the importance of the QRS complex duration as a preoperative marker for HA after aorta coronary bypass surgery, when adjusting for other risk factors. Although the one-year survival is poor for patients experiencing HA, there is no increase in mortality during the late follow-up. PMID- 10872709 TI - Mitroflow pericardial bioprosthesis in the aortic position. Low incidence of structural valve deterioration in elderly patients during an 11-year follow-up. AB - Aortic valve replacement with Mitroflow pericardial bioprosthesis, with or without concomitant bypass surgery was performed on 403 consecutive patients between March 1984 and December 1994. The mean age was 74 (range 10-92) years, with male/female ratio 183/220. Early mortality was 3.5% (14/403) and late mortality 25% (99/389). Actuarial survival was 35.4%+/-9.4% at 10 years and freedom from valve-related mortality 86.2%+/-9.6% at 8 years (total follow-up 1 270 years). The thromboembolic rate was 6.5% (fatal 0.7%)/patient year. The incidence of prosthetic valve endocarditis was 0.6% (fatal 0.3%)/patient year. Structural valve deterioration was found in 0.8%/patient year (no fatality). Actuarial freedom from all reoperations at 8 years was 92.4+/-8%. The Mitroflow pericardial heart valve is proposed as a good choice for aortic valve replacement in elderly patients, in view of the low rates of reoperation, anticoagulant treatment and valve-related thromboembolism. PMID- 10872710 TI - Off-pump bypass surgery--experience of 250 cases. AB - From April 1996 to October 1998, 250 patients with a mean age of 63 years (31-86 years) underwent coronary artery bypass grafting using the off-pump technique. The prime reason for using this technique was the need to minimize the surgical trauma by avoiding extracorporeal circulation. Fifty-seven percent of the patients had 1-vessel disease, 39% had 2-vessel disease and 4% 3-vessel disease. Sternotomy was performed in 196 patients and an anterior mini-thoracotomy in 54 patients. The mean number of coronary anastomoses was 1.5. Perioperative mortality was 0.4%. The first consecutive 87 patients underwent an early postoperative coronary angiography (days 1-5) revealing a graft patency of 96.5%. Five out of the 7 patients with occluded grafts subsequently underwent another intervention (surgical revascularization in 4 patients and percutaneous transluminal coronary angioplasty in one); 1.2% developed transmural myocardial infarction and 2.8% were reoperated upon for bleeding. The mean time of ventilatory support was 2.5+/-0.5 h. The mean ICU time for all patients was 12 h (0-10 days). The mean in-hospital time was 7 days (2-30 days). Coronary artery bypass surgery without the use of extracorporeal circulation is a safe procedure that can be performed with limited need for intensive care resources. However, long-term results remain to be investigated. PMID- 10872711 TI - Captopril improves oxygen and glucose extraction in pig hearts during reperfusion after cold cardioplegic storage. AB - The purpose of this study was to evaluate the haemodynamic and metabolic effects of captopril during reperfusion of pig hearts following 360 min global hypothermic cardioplegia and storage (HCS). The hearts were perfused with one litre of cold crystalloid cardioplegia (Bretschneider solution no. 3), excised and stored in saline at 4 degrees C for 360 min. The hearts were then reperfused with blood in a modified Langendorff model for 60 min. Left ventricular function, myocardial blood flow, and arteriovenous differences in oxygen, glucose and lactate were monitored intraoperatively and during reperfusion. Two groups of hearts were studied. Group I (captopril treated, n = 9): the pigs were pre medicated with increasing oral doses of captopril for 3 weeks (12.5 mg-150 mg daily) and an intravenous dose (25 mg) upon arrival at the laboratory. Captopril was added to the cardioplegia (1000 microg/l) and to the reperfusion media (1000 microg/l). Group II (controls, n = 8): the pigs were given no premedication, captopril-free cardioplegia and the hearts were reperfused with captopril-free blood. Captopril increased myocardial oxygen and glucose extraction during reperfusion (p < 0.05 for both) while lactate remained unchanged after 360 min HCS. Treatments with captopril increased developed left ventricular pressure (DLVP) and relaxation (-dP/dtmax) during reperfusion (p < 0.05 for both), while contractility (+dP/dtmax) was unchanged. Heart rate was reduced in captopril treated hearts (p < 0.05) while myocardial blood flow (MBF) was similar in the two groups. Captopril administration prior to and during HCS and postcardioplegic reperfusion improves oxygen and glucose extraction in large spontaneously beating porcine hearts during reperfusion. The underlying mechanisms seem to involve metabolic modulation, since myocardial uptake of oxygen and glucose was increased in the absence of changes in myocardial blood flow. PMID- 10872712 TI - Improvement of ischaemia-reperfusion injury by lazaroid U74389G in rat lung transplantation model. AB - The effect of lazaroid 74389G on ischaemia-perfusion injury in a rat lung transplantation model was investigated using three administration methods. In all groups, the University of Wisconsin (UW) solution was used as a flush and preservation solution at 4 degrees C, and lungs were stored for 12 h. Group I rats (controls) were not given any lazaroid treatment. In group II, lazaroid U74389G was added to the UW solution (100 micromol/l). In group III, lazaroid (10 mg/kg) was intravenously injected in the donors 30 min before lung ischaemia. Group IV received lazaroid treatment by the combined methods of groups II and III. In all the experimental groups (II-IV), recipient rats were given lazaroid (6 mg/kg) intravenously 30 min before reperfusion. Lazaroid improved the gas exchange function (groups II, III and IV), reduced the tissue lipid peroxides (group II) and ameliorated histologic lung damage (group II). The results thus seemed to be better in group II than in groups III and IV. PMID- 10872713 TI - Bronchial artery revascularization improves tracheal anastomotic healing after lung transplantation. AB - The study aimed to clarify the role of direct bronchial artery revascularization (BAR) after en bloc double-lung (DLT) and heart-lung transplantation (HLT). Group I comprised eight patients with en bloc DLT or HLT and successful BAR, while group II included 14 DLT or HLT cases without BAR or with failed BAR. From these groups, 2 subgroups were extracted: group III, including 6 cases of en bloc DLT with successful BAR and group IV 10 HLT cases without or with failed BAR. Airway healing was evaluated at bronchoscopy and patency of BAR with angiography. Pulmonary viral, bacterial and fungal infections, rejections and bronchiolitis obliterans syndrome (BOS) were registered. Tracheal healing at 2 weeks and 3 months was better in group I than in group 1 (p = 0.003 and p = 0.05, respectively). Compared with group IV, tracheal anastomotic healing at 2 weeks was better in group III (p = 0.007) and tended to be better also after 3 months (p = 0.07). The incidence of infections, rejection or BOS did not differ between groups I and II. BAR thus improved healing of tracheal anastomosis. PMID- 10872714 TI - Home access to hospital discharge information on audiotape reduces sick leave and readmissions in patients with first-time myocardial infarction. AB - A prospective, randomized study was carried out to investigate whether repeated hospital discharge information on audiotape at home could improve knowledge and rehabilitation in patients with first-time myocardial infarction. The study comprised 50 patients (12 females and 38 males) of up to 75 years of age. At discharge, 26 patients were given tape recordings to use at home for one week, while 24 patients represented controls. High scores in patients' knowledge about their own disease were noted, as well as differences in long-term sick leave between the two groups and readmissions or need for emergency visits. No difference in knowledge was observed, but fewer patients in the tape group had long-term sick leave, readmissions or need for emergency visits. We found that baseline characteristics were similar in the two groups. Since acquired knowledge did not appear to differ between the groups, an effect of the taped information on the patients' family network is considered. PMID- 10872715 TI - Influenza and its vaccination in HIV1-AIDS patients. AB - Influenza vaccination of HIV1-seropositive subjects generally protects against specific flu, especially if the patients have not reached a too-advanced stage. Influenza vaccination, as spontaneous influenza, can increase viral load or induce a rebound at a PCR-imperceptible phase of HIV1. Recombinant canarypox vectors encoding HIV1 antigens can stimulate HIV1-specific cytotoxic T lymphocytes. PMID- 10872716 TI - Pathogenesis of and immunity to avian influenza A H5 viruses. AB - In 1997 in Hong Kong, 18 human cases of respiratory illness were caused by an avian influenza A H5N1 virus. Although avian influenza viruses had not previously been known to cause respiratory illness in humans, the H5N1 viruses caused severe illness and death, primarily in individuals aged > 12 years. The introduction of H5N1 viruses into humans raised concerns about the potential of these viruses to cause a pandemic. We have used the BALB/c mouse to better understand the pathogenesis of and immunity to the H5N1 viruses in a mammalian model. Previously, we demonstrated that H5N1 viruses isolated from humans replicated efficiently in the lungs of mice without prior adaptation to this host. Two general phenotypes of pathogenicity of H5N1 viruses, based on high and low lethality for mice, were observed. We now demonstrate that in addition to a lethal outcome, H5N1 viruses with a high pathogenicity phenotype exhibit additional features that include rapid and uncontrolled replication in the lungs of infected mice, dissemination and replication of the virus in other organs, and depletion of peripheral blood leukocytes. The BALB/c mouse model was also used to better understand the parameters of protective immunity to the H5N1 viruses. Prior infection with H5N1 viruses of low pathogenicity or an antigenically related non-pathogenic H5N3 virus protected mice from death by infection with a highly pathogenic HK/483 virus. Serum hemagglutination-inhibition antibody titers of 40 or greater were associated with protection of mice from death. Immunization of mice with baculovirus-expressed recombinant H5 hemagglutinin protein or a previously defined HS-specific synthetic peptide induced MHC class II restricted CTL activity. Mice that had CTL activity but no serum hemagglutination-inhibition antibody were not protected from a lethal challenge with H5N1 virus. These results suggest that antibody is required for protection of mice against lethal challenge with H5N1 viruses of the high pathogenicity phenotype. PMID- 10872717 TI - The epidemiology and history of influenza. AB - Disease caused by influenza virus has been known from ancient times. However, in spite of the progress in pharmacology and vaccinology, this pathogen still affects morbidity and mortality in all age groups all over the world. This article gives some essential information on the history of influenza, its pandemics and main features of influenza viruses. Data on the epidemiologic situation of influenza in Poland in the period 1946-1999 are presented. Influenza surveillance issues and particularly prevention of influenza by vaccinations are discussed. PMID- 10872718 TI - Influenza virus genetics. AB - Significant progress has been made in understanding the process of influenza A virus replication in cell culture; however, much less is known about the genetic control of virus-host interactions in disease. This review provides an overview of the genetic analysis of influenza virus biology. The functional map of the individual genes of influenza A virus is presented as well as the status of our current understanding of pathogenesis. Influenza has a segmented genome so it is possible to obtain reassortants that contain novel combinations of genome segments derived from different viruses. This is a very useful genetic tool and is also an important aspect of influenza evolution and biology. Human influenza viruses originate from avian strains of influenza virus so that influenza infection is at its basis a zoonosis. Influenza virus strains are host restricted, however, and avian strains must be adapted to the human host. So questions of host-range and interaction with host factors are important determinants of the ability of influenza virus to cause disease in humans. Host range is restricted primarily due to host-specific interactions of the ribonucleocapsid and the viral receptor. There are two classes of drugs for inhibiting influenza infection, amantadine HCl and neuraminidase inhibitors. The mode of action and basis for resistance to these drugs are presented. Prospective targets for antiviral therapy are also discussed. PMID- 10872719 TI - Promises and challenges of live-attenuated intranasal influenza vaccines across the age spectrum: a review. AB - Despite the availability of inactivated influenza vaccines, influenza continues to cause considerable mortality in the elderly, and morbidity in all age groups. Cold-adapted, live-attenuated, intranasally administered influenza vaccines, first developed in the 1960s, have been tested in more than 10,000 volunteers and have been shown to be safe, well-tolerated, and immunogenic. Recent trials suggest that efficacy in children may be superior to that of inactivated vaccines, and efficacy in healthy adults may be similar to that of inactivated vaccines, although there are limited data comparing the two vaccines directly. Advantages of the live-attenuated vaccines include acceptability, ease of administration, and the potential for mass immunization. The possibility of substantially higher vaccination rates across all age groups brings promise for the development of herd immunity and greatly improved control of influenza in the future. PMID- 10872720 TI - Correlation between levels of autoantibodies to nerve growth factor and the clinical features of schizophrenia in children. PMID- 10872721 TI - Experimental studies of the ability of ants to add and subtract small numbers. AB - Ants had to communicate information to each other in an apparatus consisting of a horizontal "trunk" with "branches" in order to obtain food, the information being used to identify which of 40 branches had a feeder. The feeder was placed on two preselected branches significantly more frequently than on the other branches. The ants were able to tune their communication system such that the duration of communications was related to the frequency with which feeders were located on branches, and the ants could add and subtract small numbers during communication of information on the feeder identification number. PMID- 10872722 TI - Functional characteristics of the input-output correlation in the vestibular nuclear complex of the frog. AB - Experiments on perfused frog brains were used to record focal and intracellular potentials of neurons in the vestibular nuclear complex produced in response to stimulation of the anterior branch of the ipsilateral vestibular nerve and the spinal cord. Stimulation of the vestibular nerve evoked mono- and polysynaptic EPSP with orthodromic action potentials. These were accompanied by recordings of antidromic activation (with a mean latent period of 0.75 sec) of neurons which send their axons into the labyrinth. Antidromic action potentials from vestibular neurons arose with latent periods of the order of 1.43 msec in response to stimulation of the cervical thickening and 2.19 msec in response to stimulation of the lumbar thickening of the spinal cord. Bursts from the spinal cord often evoked EPSP with orthodromic action potentials in vestibular neurons. The characteristics of the functional correlation between the vestibular input and the vestibulospinal system are discussed. PMID- 10872723 TI - Synchronization of evoked secretion of quanta of mediator as a mechanism facilitating the action of sympathomimetics. AB - Experiments on frog neuromuscular junction preparations with extracellular recording of nerve terminal action potentials and single-quantum end-plate currents (EPC) were used to assess the time course of evoked quantum secretion of mediator by analyzing histograms of the distribution of true synaptic delays. These studies showed that noradrenaline, isoproterenol, and dobutamine change the kinetics of secretion of quanta, leading to synchronization of the process of mediator release; substances blocking beta-adrenoceptors (atenolol, propranolol) blocked this effect. Clonidine and phenylephrine, which activate alpha-receptors, had no effect on the kinetics of secretion, while the alpha-blocker phentolamine had no effect on the synchronizing action of noradrenaline. Reconstruction of multiquantum EPC from changes in the level of synchronization in the release of individual quanta, showed that EPC amplitude increased in response to noradrenaline by 17%, and that this was due only to alterations in the time course of secretion. These data led to the conclusion that there is a special presynaptic mechanism which facilitates the action of sympathomimetics, acting via beta-adrenoceptors. PMID- 10872724 TI - Dopaminergic mechanisms of neostriatum in the regulation of adaptive behavior by corticoliberin. AB - A conditioned active avoidance response was developed in rats with high (KHA) and low (KLA) rates of learning and the effects of injection of corticoliberin into the dorsal striatum on orientational-investigative and avoidance behavior were studied in normal animals and after depletion of striatal dopamine by preliminary injection of 6-hydroxydopamine. These studies showed that corticoliberin, like 6 hydroxydopamine, produced similar trends in the animals' behavior. Their effects were mediated by opposite mechanisms in animals with initial active and passive learning strategies for adaptive behavior. The role of dopaminergic structures of the striatum in mediating the behavioral effects of corticoliberin is discussed. PMID- 10872725 TI - NO-producing compounds transform neuron responses to glutamate. AB - We have previously shown that NO increases the excitatory effects of glutamate and blocks the desensitization of neurons to glutamate in the brain of the common snail. The aim of the present work was to identify the possible effect of NO on inhibitory responses to glutamate in the neurons of this mollusk. Electrophysiological investigations were performed on three identified neurons. The results showed that glutamate (0.05-0.1 mM) initially induced hyperpolarization and blocked the spike activity of these neurons. Simultaneous exposure to glutamate and the NO donor nitroprusside or preincubation with an NO donor had the effect that cells again responded to glutamate with depolarization and excitation. The transformed excitatory response lasted several minutes and could be reproduced even after 24 h of washing. The NO synthase blocker monomethylarginine blocked the excitatory response to glutamate. Another agonist of glutamate receptors, N-methyl-D-aspartate (NMDA, 0.1-1 mM), initially had excitatory effects on these neurons; this effect was significantly enhanced after transformation of the response to glutamate by NO donors. The results obtained here show that NO is involved in transforming the inhibitory responses to glutamate to excitatory responses, and that this effect may be mediated by NMDA type receptors. PMID- 10872726 TI - The role of the posterior hypothalamus in controlling the paradoxical phase of sleep. AB - Chronic experiments were performed on seven cats to study the effects of high frequency electrical stimulation of the posterior hypothalamic area on the characteristics of paradoxical sleep; the excitability of this structure at different stages of paradoxical sleep was determined. These studies showed that at the stage showing ECoG desynchronization and phasic events (stage 1), the response threshold for behavioral arousal resulting from stimulation of posterior hypothalamus was 20-30% higher than at the stage characterized by alpha-like activity in the ECoG and the absence of phasic phenomena (stage 2). Transient stimulation of the posterior hypothalamus at stage 1, at a level which was subthreshold for arousal from this state, led to a transition to stage 2 or a reduction in phasic events in paradoxical sleep without altering the qualitative characteristics of phase 1. Stimulation of the posterior hypothalamus at a level subthreshold for arousal from stage 2 and applied continuously during paradoxical sleep led to a reduction in the duration of this stage by 25-50% and to an increase in the proportion of stage 2 in the structure of paradoxical sleep. These results provide evidence that the posterior hypothalamus is involved in the inhibitory control of the 'executive' mechanisms of paradoxical sleep responsible for the ECoG desynchronization and the phasic manifestations of this state. It is suggested that the functional activity of the posterior hypothalamus at stage 1 also increases at stage 2, thus evidently fulfilling a 'guard' function. PMID- 10872728 TI - The effects of nitrite on the excitability of brain neurons in the common snail. AB - Electrophysiological studies were performed on the effects of sodium nitrite (0.01-1 mM) on identified command neurons in the brain of the common snail. These studies showed that short periods of exposure to nitrite (from a few minutes up to 30 min) had little effect, producing a low level of depolarization and increases in neuron spike activity in only a few cases. Incubation of isolated brains with nitrite (1 mM) for periods ranging from 2 h to several days reduced excitation thresholds, significantly increased spike activity, increases responses to stimulation, and increases the levels of synaptic activity of the neurons studied. These effects increased with time and were stable, but were reversible on washing and were accompanied by depolarization and increased input resistance. The action of nitrite was imitated by known NO donors, and blockers of NO synthesis had the opposite effect. The possible mediation of the effects of nitrite by NO synthesis and the development of hypoxia is discussed. PMID- 10872727 TI - Mechanisms of the interaction of the angular and linear components of the horizontal vestibulo-ocular reflex in the pigeon. AB - Intact pigeons (Columba livia, n = 30) were rotated in a horizontal plane in the dark at different orientations relative to the axis of rotation. A total of 24 birds showed different directions of changes in the duration of contrarotatory nystamus (on transition from central rotation to eccentric), along with displacement of the otolith membranes in both the frontal and sagittal planes. These pigeons showed a direct relationship between changes in the duration of the primary phase of nystagmus and the peak rate of the slow component on the background of increasing centrifugal force, while no such relationship was seen in conditions of decreasing centrifugal force. Increases in the duration of the primary phase were accompanied by decreases in the duration of the secondary phase (i.e., the reversive phase) and vice versa. These data provide evidence that the otolith component is not decreased to zero by rotation at constant angular rates or immediately after this stopped; in conditions of negative angular acceleration, this component was biphasic. The results are in good agreement with a hypothesis [2] suggesting that the otolith component represents asymmetric (different in paired brain structures) neuronal activity modifying the canal component even when the level of asymmetry is itself insufficient to initiate eye movements. PMID- 10872729 TI - The effects of microwave radiation from mobile telephones on humans and animals. AB - This article presents a brief review of current mobile telecommunications systems, which represent a source of microwave pollution of the environment. It has been shown that the biological effects of radiation from cellular telephones involve the time factor for the real effects on the body. Results of studies of the biological effects of low-intensity modulated microwave irradiation, including that from cellular telephones, lead to the conclusion that irradiation does not have pathological effects on the body, but does induce the usual non specific adaptive reactions. It is only in conditions of serious derangements of the immune system and prolonged exposure with cumulative effects that cancerogenic effects can occur in the body; as in other examples of external influences on the body, this is mediated by disruption of the balance between cellular repair systems and damage, the latter being favored. Several methods for studying low-intensity microwave irradiation are presented; these can be used for investigating its influence on psychophysiological functions in humans. PMID- 10872730 TI - Co-housing in a stable hierarchical group is not aversive for dominant and subordinate individuals. AB - The behavior of individuals and their responses to external stimuli are controlled by the microsocial environment, which for most mammals is associated with dominant-subordinate relationships. Physiological and behavioral differences between dominant and subordinate individuals may be 'primary' (genetically determined) or 'secondary' (due to position in the group's hierarchical structure). A series of experiments was conducted to investigate the physiological (pain response threshold), immunological (thymus, spleen weights, primary immune response), and behavioral (motor activity, behavior in a shuttle box test) characteristics of dominant and subordinate individuals in groups of three laboratory mice formed on the basis of linear hierarchy. Assessment of the effects of group conditions was made using a conditioned reflex location preference test. The results showed: 1) there are no statistically significant differences in physiological and behavioral (except for motor activity) parameters between dominant and subordinate mice; 2) co-housing of dominant and subordinate individuals in groups with stable hierarchical relationships was not aversive for them. PMID- 10872731 TI - Immunohistochemical studies of the structural bases of inhibition in the central cerebellar nuclei in mice. AB - The distribution of glutamate decarboxylase-immunoreactive structures in the central nuclei of the cerebellum, its first afferent component, was studied at the light and electron microscope levels. Axosomatic, axodendritic, and axospinous synapses were detected, in which the presynaptic parts contained glutamate decarboxylase (GDC); this enzyme is involved in GABA synthesis. Additionally, these investigations revealed axoaxonal synapses in which both poles were GDC-reactive. The central nuclei of the cerebellum were found to have an intrinsic GABAergic system. PMID- 10872732 TI - The effects of stimulation and lesioning of afferent nerves on blood glucose and free fatty acid contents in rats in conditions of changing glycemia. AB - Studies were performed on the effects of activation of afferent nerves with capsaicin (5 mg/kg i.p.) and lesioning of these nerves with neurotoxic doses of capsaicin (50 mg/kg, s.c. in two-day-old rats and 200 mg/kg s.c. in adult rats) on serum glucose and free fatty acid (FFA) concentrations in conditions of changing glucose levels induced by administration of insulin and glucose to starved (16 h) Wistar rats. These studies showed that capsaicin stimulation of intact rats decreased the hypoglycemic action of insulin, increased hyperglycemia following glucose dosage, increased FFA levels, and prevented the FFA-lowering effect of insulin. Neonatal treatment with capsaicin decreased the hypoglycemic effect of insulin but had no effect on hyperglycemia following glucose doses, but decreased FFA levels. Treatment of adult rats with neurotoxic doses of capsaicin did not alter the effect of insulin on glucose levels and decreased FFA concentrations. Capsaicin stimulation in rats following treatment with neurotoxic doses had no effect on the hypoglycemic action of insulin, but prevented it from affecting the FFA concentration. PMID- 10872733 TI - Fragmentary control of vestibuloocular responses. AB - Intact pigeons (n = 64) were rotated in the dark in the horizontal plane at different orientations relative to the axis of rotation. The overall patterns of changes in nystagmus of the eyes arising as a result of displacement of the otolith membranes in several directions were analyzed. In ten pigeons, all changes in nystagmus (type 1 general patterns) could be explained in terms of the dynamics of peripheral neuron activity and non-specific (identical for all combinations of interacting inputs) central influences. In the remaining pigeons, part of the change in nystagmus (type 2 general patterns) was associated with central influences which were not identical for different combinations of interacting inputs. Repeated unusual combinations of vestibular stimuli and subsequent treatment with Nembutal transformed type 2 general patterns into type 1 general patterns. These data provide evidence for the fragmentary control of eye movements, whereby there is selective (fragmentary) modification of only some (individually specific) combinations of canal and otolith signals out of the whole set of vestibuloocular responses arising in response to stimulation of paired vestibular inputs; modification is mediated by changes in the sign of otolith influences on the canal components of these responses. PMID- 10872734 TI - Improvements in the selective perception and training of rats using an original analog of the C-terminal fragment of vasopressin. AB - This report presents studies on the effects of intranasal administration of five doses (0.001, 0.01, 0.1, 1, and 10 microg/kg) of a new analog of arginine vasopressin fragment AVP(6-9), i.e., D-MPRG, on the learning ability of rats with positive and negative reinforcement. All doses of the peptide improved learning. The most effective dose was 0.01 microg/kg, at which the peptide accelerated the acquisition of a conditioned active avoidance reflex both when given 1 h before and when given immediately after training sessions. The peptide had greater effects when animals were trained with negative reinforcement. Analysis of the results suggests that the action of D-MPRG is mainly on perception processes, i.e., extraction of the conditioned stimulus from the environmental surroundings and evaluation and enhancement of its biological significance. In addition, this peptide prevented extinction of the acquired habit and improved the processes of consolidation, though this effect was weaker than its effect on perception. PMID- 10872736 TI - Prolonged reductions in the thresholds of evoked epileptiform discharges in slices of rat hippocampal field CA1 induced by periodic removal of Mg2+. PMID- 10872735 TI - Studies of the role of the central nucleus of the amygdala in controlling cardiovascular functions. AB - Studies on cats anesthetized with a mixture of chloralose and Nembutal addressed the effects of high-frequency stimulation (100 impulses/sec) of the central nucleus of the amygdala on bioelectrical activity in two postganglionic sympathetic nerves-the inferior cardiac nerve and the vertebral branch of the stellate ganglion, which innervate the coronary vessels and the vessels of the anterior thorax respectively. The central nucleus of the amygdala was found to have differential, selective effects, in most experiments producing increases in the amplitude of integrated activity in the inferior cardiac nerve and decreases in the amplitude of biopotentials in the vertebral nerve. In a few experiments, a second type of modulation of the activities of these two postganglionic nerves was seen, with selective inhibition of activity in the inferior cardiac nerve and an accompanying increase in activity in the vertebral nerve. Stimulation of the central nucleus of the amygdala induced significant increases in systemic arterial blood pressure. The role of the central nucleus of the amygdala in the development of experimental neurogenic hypertension was studied in a series of chronic experiments on rats; these established that rats subjected to bilateral electrolytic lesioning of the central nucleus of the amygdala prevented the development of neurogenic hypertension induced by daily imposition of stress for four weeks for induction of operant aversive conditioned reflexes, which was not the case in control rats. The role of the central nucleus of the amygdala in the regulation of vascular tone is discussed. PMID- 10872737 TI - The neurochemical basis of cognitive deficits induced by brain iron deficiency: involvement of dopamine-opiate system. AB - Iron is an essential element in maintaining normal structure and functions of the central nervous system. Dangerous effects of decreases in the bioavailability of iron in the brain are shown to affect brain biochemistry, neurotransmitters production and function, mainly in the dopamine-opiate systems well as cognitive functions (learning and memory) and a number of physiological variables such motor activity and thermoregulation. Recent research has shown the added complications and deficits that are introduced in the endocrine and the immune system activity. While iron deficiency is not perceived as a life threatening disorder, it is the most prevalent nutritional disorder in the world and a better understanding of the modes and sites of action, can help devise better treatment programs for those who suffer from it. PMID- 10872738 TI - Dietary iron and the integrity of the developing rat brain: a study with the artificially-reared rat pup. AB - Inadequate iron nutrition is thought to affect many aspects of brain development. Iron is a component of enzyme systems in DNA synthesis, the respiratory chain, neurotransmitter and lipid metabolism. The iron content of the striatum increases post-natally, with neuronal differentiation, myelin lipid and receptor formation: Seventy percent of the iron in the brain is associated with myelin. In an attempt to dissociate the global effects of under-and/or malnutrition and to produce exclusively an iron deficiency, we have used the gastrostomy-reared rat pup fed milk substitutes which vary only in their iron content. To ensure the pups did not have adequate iron reserves at birth, dams were fed a meal diet of low iron content (3 ppm) throughout gestation. The pups were then artificially reared on milk with (43 ppm), and without added iron (2.5 ppm) from 6 up to 21 days after birth. At 21 days of age, body weights of iron deficient pups were about 90% those of control animals. At 21 days of age, the pups were weaned, then fed standard laboratory rat chow. Brain was examined at 42 days of age (for young adults) and up to 6 months of age (180 days as mature adults). Morphometric analysis of sagittal sections of the cerebellum at 21 and 63 days of age revealed a deficit in white matter formation in pups fed low-iron at 21 days of age when compared to controls. This deficit was partially recouped by age 63 days. By contrast, animals fed milk supplemented with iron showed greater definition in white matter formation than controls at 21 days of age; indicative of precocious maturation of the white matter tracts. Our findings indicate that iron deficiency, without under/mal-nutrition and other variables, does not result in extensive growth deficits in body and brain weight. However, the iron status profoundly influences the development of myelination in that the process is delayed in iron deficiency. PMID- 10872739 TI - Iron deficiency alters H- and L-ferritin expression in rat brain. AB - Ferritin (Ft) H and L subunits are independently regulated proteins with both transcriptional and translational regulation in response to cellular iron levels. While the heterogeneous distribution of ferritin and iron in the brain is now well established, the relative response of each subunit to iron deficiency and iron supplementation, is not well defined. Weanling male Sprague-Dawley rats (n=12 per group) were randomly assigned to an iron deficient (3.5 mg Fe/kg diet), control (35 mg Fe/kg diet) or supplemented (350 mg Fe/kg diet) diet for six weeks. The H-/L-ferritin subunit ratio and mRNA levels were determined. Overall, the protein ratio in control rats of H to L was approximately 45:1 compared to a ratio >60:1 in iron deficiency but the absolute amounts of each subunit varied greatly from one brain region to another. The ratio of H-:L-ferritin mRNA was 6:1 and was not affected by dietary iron deficiency in contrast to a potent effect on mRNA levels in liver. Severe iron deficiency reduced brain ferritin H protein levels significantly in all regions, whereas only ferritin L levels in striatum, substantia nigra and pons were affected by iron deficiency. Supplemental dietary iron increased both ferritin subunits, with the largest increase (50%) in the hippocampus. These data indicate that ferritin H and L subunits within the brain respond differently to iron status and suggest post transcription regulation as a key event. PMID- 10872740 TI - Oligodendroglial cell differentiation in rat brain is accelerated by the intracranial injection of apotransferrin. AB - In the present paper we first studied the brain distribution and the time and dose dependent effects of apotransferrin, after its intracranial injection into young rats and at different post-natal ages. Its action upon the transferrin receptor (TfR) and upon the expression of brain transferrin, as well as its effect on the proliferation and differentiation of oligodendroglial cells (OLGc) was one of the main objectives of our investigation. Total DNA and BrdU labeling, as an index of cellularity and proliferation, respectively, were the same in the control and experimental groups of rats. A significant increase in the MBP+ and CA II+ OLGc, and a decrease in the more immature (A2B5+) OLGc were found in the aTf injected rats. At 10 and 17 days of age, Tf-mRNA decreased to around 20% of the amount present in control animals. The TfR-mRNA in the animals receiving a single dose of aTf at 3 days of age showed an increase in its expression at 10 and 17 days of age, coincident with a higher immunoreactivity of the TfR itself of neurons, choroid plexus and brain capillaries in different brain areas. Although TfR+ OLGc were present up to 7 days of age in controls and in the Tf injected rats, no positive cells were observed at 17 days of age, even in the aTf injected rats. Our results give support to the hypothesis that aTf is an important factor necessary for the maturation of the OLGc, and that the effects that it produces in the OLGc-myelin unit after its intracranial injection in young rats are not due to an increase in proliferation, but to an accelerated differentiation of Tf-sensitive OLGc. PMID- 10872741 TI - Transferrin-bound and transferrin free iron uptake by cultured rat astrocytes. AB - Previously we had demonstrated the presence of transferrin receptor (TfR) on the plasma membrane of cultured rat cortical astrocytes. In this study, we investigated the roles of TfR in transferrin-bound iron (Tf-Fe) as well as transferrin-free iron (Fe II) uptake by the cells. The cultured rat astrocytes were incubated with 1 microM of double-labelled transferrin (125I-Tf-59Fe) in serum- free DMEM F12 medium or 59Fe II in isotonic sucrose solution at 37 degrees C or 4 degrees C for varying times. The cellular Tf-Fe, Tf and Fe II uptake was analyzed by measuring the intracellular radioactivity with gamma counter. The result showed that Tf-Fe uptake kept increasing in a linear manner at least in the first 30-min. In contrast to Tf-Fe uptake, the internalization of Tf into the cells was rapid initially but then slowed to a plateau level after 10 min. of incubation. The addition of either NH4Cl or CH3NH2, the blockers of Tf-Fe uptake via inhibiting iron release from Tf within endosomes, decreased the cellular Tf Fe uptake but had no significant effect on Tf uptake. Pre-treated cells with trypsin inhibited significantly the cellular uptake of Tf-Fe as well as Tf. These findings suggested that Tf-Fe transport across the membrane of astrocytes is mediated by Tf-TfR endocytosis. The results of transferrin-free iron uptake indicated that the cultured rat cortical astrocytes had the capacity to acquire Fe II. The highest uptake of Fe II occurred at pH 6.5. The Fe II uptake was time and temperature dependent, iron concentration saturable, inhibited by several divalent metal ions, such as Co2+, Zn2+, Mn2+ and Ni2+ and not significantly affected by phenylarsine oxide treatment. These characteristics of Fe II uptake by the cultured astrocytes suggested that Fe II uptake is not mediated by TfR and implied that a carrier-mediated iron transport system might be present on the membrane of the cultured cells. PMID- 10872742 TI - Cellular distribution of ferric iron, ferritin, transferrin and divalent metal transporter 1 (DMT1) in substantia nigra and basal ganglia of normal and beta2 microglobulin deficient mouse brain. AB - We examined whether high levels of circulatory iron may cause iron accumulation in the brain. In particular, we focussed on the substantia nigra and basal ganglia as several papers have indicated that iron may accumulate here and cause death of dopaminergic neurons. Normal mice and a mouse model of hereditary haemochromatosis, the beta2-microglobulin (beta2m) knock out [beta2m (-/-)] mouse, which has high levels of circulating iron due to increased iron absorption, were examined. The iron concentration in livers were: 170+/-15 microg/g (mean +/- SD) in controls and 1010+/-50 microg/g in beta2m (-/-) mice (p<0.001), whereas in the brain the respective values were 47 +/-1 microg/g and 53+/-2 microg/g (p<0.02). Hence, the difference between cerebral iron levels of normal and beta2m (-/-) mice was small. Histological examination of the brains revealed an unequivocal distribution of ferric iron, ferritin, transferrin and divalent metal transporter 1 (DMT1), which were indistinguishable when normal and beta2m (-/-) mice were compared. In the substantia nigra and basal ganglia, ferric iron and the iron-binding proteins were present in identical cell types, which mainly comprised oligodendrocytes and microglia. Neurons were lightly labelled with transferrin and DMT1. The virtual lack of an increase in cerebral iron in beta2m (-/-) mice clearly shows that the blood-brain barrier (BBB) is capable of restricting the transport of excess plasma iron into the brain. PMID- 10872743 TI - Regulation and developmental expression of the divalent metal-ion transporter in the rat brain. AB - Divalent metal ion transporter 1 (DMT1) is a recently identified metal-ion transporter that appears to mediate the absorption of iron in the intestine. DMT1 mRNA is also present in discrete areas of the brain. In this study, we examined the expression of DMT1 mRNA in developing rat brain. DMT1 mRNA was found by in situ hybridization in the striatum, cortex, hippocampus and cerebellum. During development, DMT1 mRNA was found in Purkinje and granule cells in the cerebellum at post-natal day (PND) 14 and PND 30. DMT1 mRNA was also expressed in the external granular layer of the cerebellum at PND 14. No change in the level of DMT1 mRNA was observed by Northern analysis in the cerebellum at different ages between PND 1 and 21. DMT1 was found by Northern analysis in cultures of rat astrocytes. Activation of protein kinase C increased the expression of DMT1 in kidney epithelial cells but not astrocytes from newborn rats. Because DMT1 is expressed in a wide variety of types of cells, we suggest that it plays an important role in metal homeostasis in the brain. PMID- 10872744 TI - The heme oxygenase system and its functions in the brain. AB - The heme oxygenase (HO) system was identified in the early 1970s as a distinct microsomal enzyme system that catalyzes formation of bile pigments (Maines and Kappas, 1974). Up to the early 1990s the system was considered only as a "molecular wrecking ball" (Lane, 1998) for degradation of the heme molecule and production of toxic waste products, CO and bile pigments. For those years, the HO system remained relatively unknown to the research community. In a rather short span of the past 10 years following the discovery of high levels of a second form of the enzyme, HO-2, in the brain, suggesting that "heme oxygenase in the brain has functions aside from heme degradation" (Sun et al., 1990); concomitant with finding that another toxic gas, NO, is a signal molecule for generation of cGMP (Ignarro et al., 1982), the system was propelled into main stream research. This propulsion was fueled by the realization of the multiple and diverse functions of heme degradation products. Heme oxygenase has now found relevance in all kinds of human pathophysiology ranging from stroke, cancer, multiple sclerosis, and malaria to transplantation and immune response. As it turns out, its potential benefits are mesmerizing investigators in diverse fields (Lane, 1998). The most recent findings with HO-2 being a hemoprotein and potentially an intracellular "sink" for NO (McCoubrey et al., 1997a; Ding et al., 1999), together with the discovery of the third form of the enzyme, HO-3 (McCoubrey et al., 1997b), are likely to insure the widespread interest in the enzyme system in the coming years. The present review is intended to highlight molecular properties of HO isozymes and their likely functions in the brain. Extended reviews of the system are found in Maines (1992, 1997). PMID- 10872745 TI - Heme oxygenase-1 induction and mitochondrial iron sequestration in astroglia exposed to amyloid peptides. AB - The mechanisms responsible for pathological iron deposition and mitochondrial insufficiency that have been documented in the brains of Alzheimer (AD) patients remain poorly understood. In the present study, we demonstrate that low micromolar concentrations of amyloid1-40 (A40) and amyloid 1-42 (A42), peptides implicated in the pathogenesis of AD, increase levels of heme oxygenase-1 (HO-1) mRNA and protein in cultured rat astroglia. Furthermore, 6 days of exposure to amyloid augments the sequestration of 55FeCl3-derived iron by astroglial mitochondria without affecting the disposition of this metal in whole-cell and lysosomal compartments. Mitochondrial iron deposition was not observed in the amyloid-treated glia when diferric-transferrin served as the metal donor. We had previously shown that inhibitors of HO-1 and the mitochondrial permeability transition pore (MTP) block the uptake of mitochondrial iron in astrocytes exposed to the pro-oxidant effects of dopamine and several pro-inflammatory cytokines. Similarly, in the current study, amyloid-induced mitochondrial iron trapping was significantly attenuated by co-administration of the HO-1 transcriptional suppressor, dexamethasone (DEX) or the MTP blocker, cyclosporin A (CSA). Thus, the marked enhancement of HO-1 expression previously demonstrated in AD-affected neurons and astroglia may transduce amyloid (oxidative) stress into the abnormal patterns of iron deposition and mitochondrial insufficiency characteristic of this disease. Finally, in experiments employing cytotoxic concentrations of A40, we provide evidence that inhibition of HO-1 transcription and related mitochondrial iron deposition may be an important mechanism by which DEX protects tissues subjected to amyloid stress. PMID- 10872746 TI - Tin-mesoporphyrin, a potent heme oxygenase inhibitor, for treatment of intracerebral hemorrhage: in vivo and in vitro studies. AB - Spontaneous intracerebral hemorrhage (ICH) is the stroke subtype with highest mortality and morbidity. ICH can also occur following traumatic brain injury and thrombolysis for ischemic stroke and myocardial infarction. Development of ICH induced hemispheric edema can elevate intracranial pressure and cause death. In survivors, edema-related white matter injury can lead to life-long neurological deficits. At present, there are no scientifically proven treatments for ICH. Heme oxygenase products, particularly iron and bilirubin, can be toxic to cells. In cerebral ischemia models, metalloporphyrins that are potent heme oxygenase inhibitors, reduce edema and infarct size. Tin-mesoporphyrin (SnMP) is a neuroprotectant that has also been used clinically to treat hyperbilirubinemia. Presently, we tested the hypothesis that SnMP treatment would reduce edema development following experimental ICH. We produced hematomas in pentobarbital anesthetized pigs (9-11 kg) by infusing autologous blood into the frontal white matter. To maximize tissue concentrations, SnMP (87.5 microM in DMSO) or DMSO (vehicle controls) was included in the infused blood. Pig brains were frozen in situ at 24 hrs. following ICH and hematoma and edema volumes were determined on coronal sections by computer-assisted image analysis. We also examined the effects of SnMP in vitro on ferritin iron release, the formation of iron-induced thiobarbituric acid reactive substances (TBARS) and initial clot formation and hemolysis. SnMP treatment significantly reduced intracerebral mass following ICH. This was due to significant decreases in hematoma (0.68+/-0.08 vs. 1.39+/-0.30 cc, vehicle controls p<0.025) and edema volumes (edema = 1. 16+/-0.33 vs. 1.77+/ 0.31 cc, p<0.05). In vitro, SnMP did not stabilize ferritin iron against reductive release nor did it decrease iron-induced TBARS formation in brain homogenates. SnMP or DMSO added to pig blood did not alter clot weights. In conclusion, SnMP reduced intracerebral mass in an ICH model by decreasing both hematoma and edema volumes SnMP's mechanism of action is presently unknown but may involve its potent inhibition of heme oxygenase activity. SnMP's effect appears unrelated to ferritin iron release, antioxidant activity or initial clot formation. Since SnMP treatment could be brain protective following ICH, further investigations into neurological and neuropathological outcomes and as well as into its mechanism of action are warranted. PMID- 10872747 TI - Nitric oxide induces heme oxygenase-1 via mitogen-activated protein kinases ERK and p38. AB - Heme oxygenase-1 (HO), the heat shock/stress cognate of the heat shock protein 32 (HSP32) family of proteins, is postulated to be a component of cellular defense mechanisms against oxidative stress-mediated injury. Nitric oxide (NO) is among the extensive array of stimuli that induce HO-1. The cellular signaling mechanisms that regulate the induction of HO-1 by NO are not understood. In the present study, we have demonstrated that exposure of HeLa cells to the NO donor, sodium nitroprusside (SNP), results in concentration and time-dependent increase in HO-1 mRNA and activation of MAPKs: ERK (ERK1 and ERK2) and p38 pathways, but not SAPK/JNK pathway. Pre-treatment of the cells with PD98059, a selective ERK pathway inhibitor, and SB203580, a p38 MAPK inhibitor, blocked the induction of HO-1 by the NO donor in a dose-dependent manner. In addition, an increase in HO-1 mRNA level that was detected as early as 2 hrs.following SNP treatment preceded c jun and c-fos induction. These transcription factors are downstream of SAPK/JNK pathway, and their increased expression was detected at 3hr. and 6hr. after SNP treatment. Similarly, AP-1 DNA binding activity was not increased when measured 6 hrs. after SNP treatment. ERK and p38 inhibitors also suppressed induction of HO 1 by SNAP and GSNO. The increase in HO-1 mRNA was inhibited by actinomycin D and cycloheximide, but not by NAC, and was not mimicked by the lipophilic cGMP analogue, 8-bromo-cGMP, suggesting that NO-mediated induction required de novo RNA and protein synthesis and was unrelated to cGMP and redox signaling. Collectively, the findings suggest that MAP kinase ERK and p38 pathways are involved in the NO-mediated induction of HO-1 and that SAPK/JNK pathway and increased DNA binding of AP-1 transcription factor are not involved in HO-1 gene activation by NO. A plausible mechanism by which the NO donors cause HO-1 induction may involve HO-1 gene regulation by its substrate, heme. PMID- 10872748 TI - Effects of hypoxia preconditioning on expression of metallothionein-1,2 and heme oxygenase-1 before and after kainic acid-induced seizures. AB - Global hypoxia preconditioning provides neuroprotection against a subsequent, normally damaging challenge. While the mechanistic pathways are unknown, changes in the expression of stress-related proteins are implicated. Hypoxia preconditioning attenuates the brain edema and neuropathology associated with kainic acid-induced status epilepticus in a protein synthesis-dependent manner when a kainic acid challenge is given up to one week post-preconditioning. Kainic acid initiates a glutamate-driven status epilepticus causing a Ca2+ and oxidative stress, resulting in injury to the piriform cortex and hippocampus. Stress related gene expression [e.g. metallothioneins (MTs), heme oxygenase-1 (HO-1)] is enhanced during seizures in vulnerable brain areas, (e.g. piriform cortex). This study explores the effects of hypoxia preconditioning on expression of MT-1, MT-2 and HO-1 before and after kainic acid-induced seizures. Analysis of MT-1, MT-2 and HO-1 expression, through Western and Northern blotting, indicates that there is a variable pattern of induction and suppression of these two genes following hypoxia preconditioning alone as well as after kainic acid-induced seizures compared to non-preconditioned animals. These findings suggest that hypoxia preconditioning induces an adaptive response that prevents kainic acid seizure associated neuropathology even when robust seizures occur. This may involve a variety of stress-related proteins, working in concert, each with their own individual expression profiles. Induction of this type of neuroprotection pharmacologically, or through preconditioning, will provide a better understanding of the stress response in brain. PMID- 10872749 TI - Free radical scavenging actions of hippocampal metallothionein isoforms and of antimetallothioneins: an electron spin resonance spectroscopic study. AB - The high concentration of zinc in the hippocampal mossy fiber axon boutons is localized in the vesicles and is mobilized by exocytosis of the zinc-laden vesicles. Furthermore, the mammalian hippocampi contain metallothionein (MT) isoforms which regulate the steady state concentration of zinc, an important antioxidant. Indeed, zinc deprivation leads to an increased lipid peroxidation, reduces the activity of Cu++-Zn++ superoxide dismutase, and protect against oxidative stress such as exposure to ultraviolet A irradiation. By employing electron spin resonance (ESR) spectroscopy, we have demonstrated that rat hippocampal MT isoforms 1 and 2 were able to scavenge 1,1-diphenyl-2 picrylhydrazyl radicals (DPPH), hydroxyl radicals (*OH) generated in a Fenton reaction, and superoxide anions (O2*-) generated by the hypoxanthine and xanthine oxidase system. In addition, MT-1 isoform protected the isolated hepatocytes from lipid peroxidation as determined by thiobarbituric acid bound malondialdehyde. MT antibodies scavenged DPPH radicals, hydroxyl radicals and reactive oxygen species but not superoxide anions. The results of these studies suggest that although both isoforms of MT are able to scavenge free radicals, the MT-1 appears to be a superior scavenger of superoxide anions and 1,1-diphenyl-2-picrylhydrazyl radicals. Moreover, antibodies formed against MT isoform retain some, but not all, free radical scavenging actions exhibited by MT-1 and MT-2. PMID- 10872750 TI - Iron, metalloenzymes and cytotoxic reactions. AB - There is considerable evidence implicating iron and other redox-active transition metals as progenitors of reactive intermediates of oxygen (ROI), molecules which lead to oxidative stress and contribute to various neurodegenerative processes. An important aspect of such metal-mediated damage to biomolecules is the site specific nature of such pathological activity. Iron sequestering molecules, such as ferritin, transferrin, lactotransferrin, melanotransferrin, hemosiderin and heme can serve as cytoprotectants against metal-mediated oxidant damage. Metalloenzymes also constitute an important group of iron sequestering molecules. Metalloenzyme-catalyzed reactions in which metal ions at the enzyme active site undergo redox-cycling in association with O2 are site-specific in nature, and may represent a potential source of ROI-mediated damage to biomolecules. Dysregulation of brain iron and alterations in the levels of metalloenzymes involved in reactions with O2 derived molecules can contribute to neuronal damage. Iron may increase the cytotoxicity of neuronal dopamine by increasing its rate of oxidation to quinones and semiquinones, thereby reducing the level of this neurotransmitter. Interestingly, dopamine also may play an important role in the maintenance of transition-metal homeostasis as an iron chelator, since it can form both catecholate and hydroxamate groups, molecules employed by many microorganisms to sequester iron. PMID- 10872751 TI - Measurement of loosely-bound iron in brain regions using redox cycling and salicylate. AB - A sensitive iron assay was developed for measuring non-heme and loosely bound iron in regions of rat brain. The method is based on the salicylate trapping of hydroxyl radicals generated from ascorbate-driven redox cycling of Fe3+-EDTA. This assay has high sensitivity (about 20 nM) because of amplification obtained with redox-cycling and fluorescent detection of the salicylate hydroxylation product, 2,5-dihydroxybenzoate. The assay detects iron as Fe2+ and Fe3+ combined. Values of non-heme and loosely bound iron are given for three areas of cortex, caudate, hippocampus, thalamus and brainstem of the rat brain. PMID- 10872752 TI - Iron and free radical oxidations in cell membranes. AB - Brain tissue being rich in polyunsaturated fatty acids, is very susceptible to lipid peroxidation. Iron is well known to be an important initiator of free radical oxidations. We propose that the principal route to iron-mediated lipid peroxidations is via iron-oxygen complexes rather than the reaction of iron with hydrogen peroxide, the Fenton reaction. To test this hypothesis, we enriched leukemia cells (K-562 and L1210 cells) with docosahexaenoic acid (DHA) as a model for brain tissue, increasing the amount of DHA from approximately 3 mole % to 32 mole %. These cells were then subjected to ferrous iron and dioxygen to initiate lipid peroxidation in the presence or absence of hydrogen peroxide. Lipid-derived radicals were detected using EPR spin trapping with alpha-(4-pyridyl-1-oxide)-N-t butylnitrone (POBN). As expected, lipid-derived radical formation increases with increasing cellular lipid unsaturation. Experiments with desferal demonstrate that iron is required for the formation of lipid radicals from these cells. Addition of iron to DHA-enriched L1210 cells resulted in significant amounts of radical formation; radical formation increased with increasing amount of iron. However, the exposure of cells to hydrogen peroxide before the addition of ferrous iron did not increase cellular radical formation, but actually decreased spin adduct formation. These data suggest that iron-oxygen complexes are the primary route to the initiation of biological free radical oxidations. This model proposes a mechanism to explain how catalytic iron in brain tissue can be so destructive. PMID- 10872753 TI - Chemical changes in the photoreceptor outer segments due to iron induced oxidative stress: analysis by Fourier transform infrared (FT-IR) microspectroscopy. AB - Oxidative stress is thought to be an important pathogenic mechanism in many diseases of the retina. The purpose of this study was to investigate the chemical changes that are present in the photoreceptor outer segments of the retina following exposure to oxidative stress. Fourier transform infrared (FT-IR) microspectroscopy enables the characterization and semi-quantitation of chemical functional groups in microscopic regions of tissue sections. This technique was used to evaluate the chemical changes in the outer segments following exposure to ferrous sulfate, which promotes oxidative tissue damage. A reduction of C=C-H and C=O functional groups was observed in the outer segments of iron-injected eyes compared to vehicle-injected eyes at 3 days following injection, which is prior to major histological changes that occur by 7 days. These functional groups are found in docosahexaenoic acid (DHA), which is present at a high concentration in the outer segments. DHA contains a series of six cis-conjugated double bonds, which are vulnerable to free radical attack, and the reduction of these unsaturation group absorptions suggests that DHA was degraded and/or removed from the outer segments. An unexpected finding was that several other chemical functional groups increased in concentration over time in the outer segments of vehicle-injected eyes compared to non-injected eyes. These increases generally did not include C=C-H or C=O, which suggests that either DHA was being degraded while other organic molecules were being concentrated, or that production of DHA failed to be upregulated in vehicle-injected eyes. In summary, there was a loss of both C=C-H and C=O functional group concentrations in the outer segments of iron-injected eyes, and there was an increased concentration of several other chemical functional groups following trauma induced by vehicle injection. PMID- 10872754 TI - Protein oxidation and enzyme susceptibility in white and gray matter with in vitro oxidative stress: relevance to brain injury from intracerebral hemorrhage. AB - Intracerebral hemorrhage (ICH) is a devastating stroke sub-type with high mortality and morbidity. ICH frequently occurs in subcortical white matter generating hematomas that contain high heme iron levels. In this study, we examined the consequences of iron-induced oxidation (1-100 microM Fe2+ for 30 min. or 50 microM Fe2+ for 1-120 min.) on the activities of two oxidatively sensitive enzymes, creatine kinase (CK) and glutamine synthetase (GS), and on an oxidative stress marker, protein carbonyl formation, in porcine cerebral cortical white and gray matter. In vitro iron oxidation produced time and concentration dependent decreases in both CK [maximum decreases of 49.3+/-1.2% and 44.3+/-4.1% (average +/- SEM, N=3) for white and gray matter, respectively] and GS activities (maximum decreases of 16.9+/-1.7% and 13.2+/-1.0% for white and gray matter, respectively) and increases in protein carbonyl formation. Interestingly, protein carbonyl concentrations were significantly greater (p<0.05) in white vs. gray matter at 100 microM iron (30 min.) and 50 microM iron (120 min.). Additionally, CK and GS activities were lower for white versus gray matter at several time points and iron concentrations. It is our hypothesis that iron induced oxidative stress contributes to the pathogenesis of perihematomal brain injury following ICH. PMID- 10872755 TI - Metal-catalyzed oxidation of brain-derived neurotrophic factor (BDNF): selectivity and conformational consequences of histidine modification. AB - We have studied the metal-catalyzed oxidation (MCO) of brain-derived neurotrophic factor (BDNF) with regard to target sites and potential conformational changes of the protein. The exposure of BDNF to three different levels of ascorbate/Cu(II)/O2 [20 microM Cu(II), 2 mM ascorbate (level 1); 20 microM Cu(II), 4 mM ascorbate (level 2); 40 microM Cu(II), 4 mM ascorbate (level 3)], chosen based on the extent of chemical modification of Met and His, respectively, resulted in the exclusive oxidation of a buried Met residue, Met92, at level 1 but in the predominant oxidation of His at level 3. His modification had a significant impact on the structure of BDNF, as quantified by CD and ANSA fluorescence measurements, while Met oxidation had not, also assessed through complementary oxidation of BDNF through hydrogen peroxide. Our ultimate objective was the correlation of the surface exposure of an oxidized His residue in a protein with potential effects on the conformational integrity of the oxidized protein. In a series of three proteins, human growth hormone (hGH), human relaxin (hR1x), and BDNF, we have now observed that His oxidation is paralleled by significant conformational changes when the target His residue is more surface exposed (hR1x, BDNF) while conformational consequences of His modification are less significant when the target His residues are more buried in the interior of the protein (hGH). PMID- 10872756 TI - A critical appraisal of the reporting of the National Acute Spinal Cord Injury Studies (II and III) of methylprednisolone in acute spinal cord injury. AB - From the beginning, the reporting of the results of National Acute Spinal Cord Injury Studies (NASCIS) II and III has been incomplete, leaving clinicians in the spinal cord injury (SCI) community to use or avoid using methylprednisolone in acute SCI on the basis of faith rather than a publicly developed scientific consensus. NASCIS II was initially reported by National Institutes of Health announcements, National Institutes of Health facsimiles to emergency room physicians, and the news media. The subsequent report in the New England Journal of Medicine implied that there was a positive result in the primary efficacy analysis for the entire 487 patient sample. However, this analysis was in fact negative, and the positive result was found only in a secondary analysis of the subgroup of patients who received treatment within 8 hours. In addition, that subgroup apparently had only 62 patients taking methylprednisolone and 67 receiving placebo. The NASCIS II and III reports embody specific choices of statistical methods that have strongly shaped the reporting of results but have not been adequately challenged or or even explained. These studies show statistical artifacts that call their results into question. In NASCIS II, the placebo group treated before 8 hours did poorly, not only when compared with the methylprednisolone group treated before 8 hours but even when compared with the placebo group treated after 8 hours. Thus, the positive result may have been caused by a weakness in the control group rather than any strength of methylprednisolone. In NASCIS III, a randomization imbalance occurred that allocated a disproportionate number of patients with no motor deficit (and therefore no chance for recovery) to the lower dose control group. When this imbalance is controlled for, much of the superiority of the higher dose group seems to disappear. The NASCIS group's decision to admit persons with minor SCIs with minimal or no motor deficit not only enables statistical artifacts it complicates the interpretation of results from the population actually sampled. Perhaps one half of the NASCIS III sample may have had at most a minor deficit. Thus, we do not know whether the results of these studies reflect the severely injured population to which they have been applied. The numbers, tables, and figures in the published reports are scant and are inconsistently defined, making it impossible even for professional statisticians to duplicate the analyses, to guess the effect of changes in assumptions, or to supply the missing parts of the picture. Nonetheless, even 9 years after NASCIS II, the primary data have not been made public. The reporting of the NASCIS studies has fallen far short of the guidelines of the ICH/FDA and of the Evidence-based Medicine Group. Despite the lucrative "off label" markets for methylprednisolone in SCI, no Food and Drug Association indication has been obtained. There has been no public process of validation. These shortcomings have denied physicians the chance to use confidently a drug that many were enthusiastic about and has left them in an intolerably ambiguous position in their therapeutic choices, in their legal exposure, and in their ability to perform further research to help their patients. PMID- 10872757 TI - Spinal fusion for lumbar instability: does it have a scientific basis? AB - The validity of spinal fusion for lumbar instability is considered. Some difficulty lies in the interpretation of the term instability. The differing interpretations in the clinical, radiologic, and biomechanical contexts are discussed. These interpretations may only be reconciled if the confusion between hypermobility and instability is removed and some recourse is made to soft tissue integrity. Fusion is considered in the context of the functioning spine as a whole. Although the aim of the surgery is usually to produce a solid arthrodesis, some studies show that this can compromise the functioning of the rest of the spine. Furthermore, there is some evidence that pseudarthrosis itself may not be detrimental. Dynamic imaging offers the potential for improved diagnosis and assessment, but further work is needed to pave the way for better selection criteria and treatment strategies. PMID- 10872758 TI - The prevalence of low back pain: a systematic review of the literature from 1966 to 1998. AB - A systematic literature review of population prevalence studies of low back pain between 1966 and 1998 was conducted to investigate data homogeneity and appropriateness for pooling. Fifty-six studies were analyzed using methodologic criteria that examined sample representativeness, data quality, and pain definition. Acceptable studies were assessed for homogeneity and appropriateness for pooling. Thirty were methodologically acceptable. Of these there were significant differences in study design, patient age, mode of data collection, potential temporal effects, and prevalence results. Point prevalence ranged from 12% to 33%, 1-year prevalence ranged from 22% to 65%, and lifetime prevalence ranged from 11% to 84%. A limited number of studies were left for analysis, making the pooling of data difficult. A model using uniform best-practice methods is proposed. PMID- 10872759 TI - Cervical pedicle screw insertion: assessment of safety and accuracy with computer assisted image guidance. AB - We used a commercially available computer-assisted image-guidance system for cervical pedicle screw insertion in both the laboratory and in a preliminary clinical setting. Nine plastic cervical spine models (C2-C7) were used in the laboratory test. The StealthStation was used to create the preoperative plan for each screw such that it would be inserted down the center of the pedicle, parallel to the long axis. Using a light-emitting diode-equipped drill guide, 2 mm holes were drilled in 108 pedicles. A total of 108 pedicle holes were drilled. The mean trajectory deviation from the surgical plan in the axial plane was 1.7+/ 1.7 degrees (range, 0 to 8 degrees), and the mean deviation of the position of the hole was 1.7+/-0.6 mm (range, 0.1 to 2.9 mm). Eighty-three drill holes (76.9%) were contained within the pedicles, whereas partial cortical perforation was noted in 25 pedicles (23.1%). In the clinical setting, 36 cervical pedicle screws were inserted in nine patients using the image-guided system. Within the limits of imaging artifact, all 36 pedicle screws appear to have been inserted accurately by postoperative computed tomographic examination. No neurologic or vascular complications were encountered. PMID- 10872760 TI - In vivo pedicle screw placement: image-guided virtual vision. AB - Near-real-time frameless stereotaxy registering intraoperative anatomy to a preoperative three-dimensional computer model has been developed for use with in vivo pedicle screw placement. Eight patients underwent thoracolumbar and lumbar spine stabilization surgery using this new technology, and 32 pedicle screws were placed. Three additional patients had 12 pedicle screws removed during revision surgery, and they allowed the authors to estimate the accuracy of this navigational system. Accuracy was determined by comparing pedicle screw position on postoperative computed tomographs for the first eight patients and on preoperative computed tomographs for the latter three patients, with the intraoperative computer trajectory data gathered during operation. In the group of eight patients, all screws were intrapedicular. In evaluating all 11 patients, the overall accuracy was +/- 2 mm, but the greatest error of 5.4 mm was noted in the sagittal plane measurement. During the development phase of this technology, time constraints prolong surgery, but this may be addressed once the tool's accuracy has been confirmed and intraoperative radiographic confirmation becomes unnecessary. In vivo real-time frameless stereotaxy for pedicle screw placement offers promise for the future. Refinements are needed to improve accuracy and address time constraints. PMID- 10872761 TI - Biomechanical evaluation and preliminary clinical experience with an expansive pedicle screw design. AB - The advantages of pedicle screw fixation depend on their ability to retain bony purchase until the fusion mass is stable. Osteoporotic bone and removal and replacement of pedicle screws in revision procedures substantially reduce screw mechanical fixation strength and can lead to clinical failure. The objective of this study was to determine if an expansive pedicle screw design could be used to improve biomechanical fixation in bone of compromised quality. Axial mechanical pullout testing was performed on paired expansive and conventional pedicle screws placed in fresh, unembalmed cadaveric vertebrae. Bone mineral density measurements (made using a dual-energy X-ray absorption meter) were used to characterize bone quality. A preliminary clinical and radiographic evaluation of 14 patients was also performed at a minimum 2-year follow-up. The mean axial pullout force in bone of all qualities was increased 30% when the expansive pedicle screw design was used. This included an appropriate 50% increase in pullout force in bone of poor quality (low bone mineral density). The preliminary clinical and radiographic results were supportive of the biomechanical design rationale and mechanical testing. The results were similar to those expected for spinal instrumentation using pedicle screws, even though compromised bone was present in two thirds of the cases in which the expansive screw was used. PMID- 10872762 TI - Quality of life before and after microsurgical decompression in lumbar spinal stenosis. AB - Twenty consecutive patients (10 men and 10 women; median age, 68 years) with lumbar spinal stenosis were studied before and after microsurgical decompression without laminectomy. Fourteen of the patients had pure stenosis symptoms, whereas six had intercurrent diseases that could exacerbate the symptoms of stenosis. The mean duration of symptoms was 4.5 years (range, 1 to 15 years). All patients were interviewed before operation, and an assessment form based on and modified from the Oswestry Low Back Pain Disability Questionnaire was completed. The ability to perform physical activities including house work, gardening, going to the post office, and so forth was markedly reduced before operation for nearly all patients, and social life such as traveling, meeting friends, and participating in hobbies was similarly restricted. Sleeping was also greatly affected before operation, as were psychological parameters including irritability, depression, infirmity, energy, patience, and concentration. At follow-up 2.8 years after surgery, 13 of the 14 patients with pure stenosis evaluated their quality of life as much improved and principally normal. Among the patients with intercurrent diseases, only two of six judged the quality of their lives as much improved. PMID- 10872764 TI - An experimental study of porcine lumbar segmental stiffness by the distraction compression principle using a threaded interbody cage. AB - The objectives of this study were to quantify changes in stiffness and disk height of porcine functional spinal units (FSUs) by installation of a threaded interbody cage and those by gradual resection of the annulus fibrosus. Flexion, extension, bending, and torsion to the FSUs were performed in four sequential stages: stage I, intact FSU; stage II, the FSUs are fitted with a threaded fusion cage; stage III, the FSUs are fitted with a threaded fusion cage with the anterior one third of the annulus fibrosus excised, including excision of the anterior longitudinal ligament; and stage IV, in addition to stage III, the bilateral annulus fibrosus is excised. Segmental stiffness in each loading in the four stages and a change of disk height induced by the instrumentation were measured. After instrumentation, stiffnesses in all loading modes (p < 0.005) and disk height (p = 0.002) increased significantly. The stiffnesses of FSUs fixed by the cage decreased with gradual excision of the annulus fibrosus in flexion, extension, and bending. These results suggest that distraction of the annulus fibrosus and posterior ligamentous structures by installation of the cage increases the soft-tissue tension, resulting in compression to the cage and a stiffer motion segment. PMID- 10872763 TI - Postoperative enteroparesis by patient-controlled analgesia combined with continuous epidural block for patients after posterior lumbar surgery. AB - This study evaluated postoperative enteroparesis influenced by patient-controlled analgesia combined with continuous epidural block in patients who underwent posterior lumbar surgery. One hundred nine patients were divided into three groups at random (group 1, controls (18 patients); group 2, postoperative patient controlled analgesia and continuous epidural block (45 patients); group 3, one shot epidural analgesia, postoperative patient-controlled analgesia, and continuous epidural block (46 patients). The patients in groups 2 and 3 had more satisfactory pain relief and needed analgesics less frequently. However, their clinical abdominal findings the morning after surgery were worse than those in control patients. The times when patients could take any nourishment and eat solid food (rice) were delayed by patient-controlled analgesia with continuous epidural block. PMID- 10872765 TI - Reliability in grading the severity of lumbar spinal stenosis. AB - Stenosis of the lumbar spinal canal is a major cause of disability and lost productivity. Computed tomography (CT) is used commonly to assess the presence and severity of spinal stensosis, because it is relatively inexpensive, readily available, and has few adverse effects. The ability of four surgeons to agree about the presence and severity of lumbar spinal stenosis based on plain CT scans was evaluated from 30 scans of varying stenosis severity (normal to severe). Kappa, a measure of chance-corrected agreement, was calculated. Surgeons exhibited moderate agreement for the presence or absence of spinal stenosis (kappa = 0.58+/-0.06). Agreement regarding the severity of stenosis, when present, was poor (kappa = 0.26+/-0.04). The ability of surgeons to agree was not improved when individual features of the CT scans were assessed (facet joint arthrosis, ligamentum flavum hypertrophy, disk protrusion, and nerve root impingement). This study suggests that CT scans are not a reliable method by which to examine the severity of lumbar spinal stenosis. PMID- 10872766 TI - Comparison of the in vitro holding strengths of conical and cylindrical pedicle screws in a fully inserted setting and backed out 180 degrees. AB - Previous investigations have suggested that conical and cylindrical pedicle screws have comparable holding strengths. So far, the remaining performance in screws turned back or loose as a result of other reasons has not been determined. Twenty-four cadaveric spines from 6- to 8-week-old calves were examined. After bone mineral density was determined, four pedicle screws (two conical and two cylindrical screws) were inserted. The screws were fully inserted and half of them turned back 180 degrees. Twenty-four axial pullout and 24 cyclic loading tests with subsequent pullout tests were conducted. The pullout strengths of conical screws turned back 180 degrees are significantly smaller (1.8 kN) than those of cylindrical screws (4.3 kN). After cyclic loading, the displacement of conical screws is significantly greater (6.9 mm) than that of cylindrical screws (4.7 mm). Pedicle screws, especially conical ones, need to be placed to a correct depth, and they should not have to be backed out. PMID- 10872767 TI - Thoracic epidural desmoplastic fibroma. AB - Desmoplastic fibroma is a relatively uncommon tumor and rarely involves the spine. The authors describe a 20-year-old woman with a thoracic epidural desmoplastic fibroma treated by complete resection and posterior spinal fusion. Four years after surgery, neither the tumor nor clinical symptoms have recurred. Thus, complete resection is considered necessary to treat this tumor. PMID- 10872768 TI - Unilateral blindness as a complication of intraoperative positioning for cervical spinal surgery. AB - The authors report a case of unilateral blindness after surgical vertebral stabilization for C5-C6 subluxation. The blindness resulted from ischemia of the retina caused by prolonged compression of the eyeball on the surgical bed. This injury can be serious and irreversible, so it must be prevented by placing the patient in the proper position. The anesthetist must pay particular attention to avoid the consequences of possible intraoperative movement. PMID- 10872769 TI - Fusion outcome. PMID- 10872770 TI - Preparation, characterization, and performance of magnetic iron-carbon composite microparticles for chemotherapy. AB - Magnetic microcarrier particles useful for delivering chemotherapeutic drug molecules are described. The particles are formed by joint deformation of iron and carbon in a ball mill. Physical, chemical, and functional characterization has been carried out on the particles. Physical characteristics include microscopy, particle size analysis (0.5-5 microm), surface area (250 m2/g), water vapor adsorption isotherm (hydrophobic surface), and analysis of the iron-carbon interface by Mossbauer spectroscopy, X-ray diffraction, and differential thermal analysis. Chemical analysis was used to identify elements in the particles other than carbon and iron. Functional characteristics measured included the particles' ability to adsorb and desorb doxorubicin, cytotoxicity, and their magnetic susceptibility. PMID- 10872771 TI - Dynamic viscoelastic properties and the age changes of long-term soft denture liners. AB - The dynamic viscoelastic properties of long-term soft denture liners were measured over a wide range of frequencies using a dynamic viscoelastometer based on a non-resonance-forced vibration principle. Changes in properties over a 3 yr period have also been monitored. One acrylic material, one fluoroelastomer, one heat cured silicone and one self-curing addition silicone were used. Complex dynamic tensile modulus (E*), tensile storage modulus (E'), tensile loss modulus (E") and loss tangent (tan delta) were determined over the frequency range from 0.01 to 100 Hz on administration of a 0.27% strain at 37 degrees C. The dynamic viscoelasticity of the acrylic and fluoroelastomer products was more sensitive to changes in frequency than that of silicone products. The acrylic material and fluoroelastomer exhibited viscoelastic behaviour whilst silicones exhibited elastic behaviour. The silicone products remained unchanged after soaking for 3 yr whilst the acrylic and fluoroelastomer products underwent significant change. PMID- 10872772 TI - Synthesis of biomimetic Ca-hydroxyapatite powders at 37 degrees C in synthetic body fluids. AB - An important inorganic phase for synthetic bone applications, calcium hydroxyapatite (HA, Ca10(PO4)6(OH)2), was prepared as a nano-sized (approximately 50 nm), homogeneous and high-purity ceramic powder from calcium nitrate tetrahydrate and diammonium hydrogen phosphate salts dissolved in modified synthetic body fluid (SBF) solutions at 37 degrees C and pH of 7.4 using a novel chemical precipitation technique. The synthesized precursors were found to easily reach a phase purity >99% after 6 h of calcination in air atmosphere at 90 degrees C, following oven drying at 80 degrees C. There was observed, surprisingly, no decomposition of HA into the undesired beta-TCP phase even after heating at 1,600 degrees C in air for 6 h. This observation showed the superior high-temperature stability of such 'biomimetic' HA powders as compared to those reported in previous studies. The former powders were also found to contain trace amounts of Na and Mg ions, originating from the use of SBF solutions instead of pure water during their synthesis. Characterization and chemical analysis of the synthesized powders were performed by X-ray powder diffraction, energy-dispersive X-ray spectroscopy, Fourier-transform infra-red spectroscopy, scanning electron microscopy, and inductively coupled plasma atomic emission spectroscopy. PMID- 10872773 TI - Dynamic behavior of glucose oxidase-containing microparticles of poly(ethylene glycol)-grafted cationic hydrogels in an environment of changing pH. AB - Poly(diethylaminoethyl-g-ethylene glycol) microparticles were prepared by suspension polymerization of diethylaminoethyl methacrylate, poly(ethylene glycol) monomethacrylate and the crosslinking agent tetra(ethylene glycol) dimethacrylate in silicone oil using redox initiators. Particles of different sizes, crosslinking ratios and graft molecular weights were prepared. The changes in the swelling of the particles were studied as the pH was changed between 3.0 and 7.4. The particles showed rapid swelling/deswelling dynamics in response to changes in pH. It was evident that faster response could be obtained from smaller particles. Changing the crosslinking ratio resulted in changes in the extent of swelling, as well as the speed of response. It was also found that longer graft lengths were responsible for increasing the effect of relaxation of the swelling of the network. PMID- 10872774 TI - Degradation of mechanical behavior in UHMWPE after natural and accelerated aging. AB - Ultra-high molecular weight polyethylene (UHMWPE) is known to degrade during natural (shelf) aging following gamma irradiation in air, but the mechanical signature of degradation remains poorly understood. Accelerated aging methods have been developed to reproduce the natural aging process as well as to precondition total joint replacement components prior to joint simulator wear testing. In this study, we compared the mechanical behavior of naturally (shelf) aged and accelerated aged tibial inserts using a previously validated miniature specimen testing technique known as the small punch test. Tibial inserts made-of GUR 1120 and sterilized with 25 to 40 kGy of gamma radiation (in air) in 1988, 1993, and 1997 were obtained; a subset of the 1997 implants were subjected to 4 weeks of accelerated aging in air at 80 degrees C. To determine the spatial variation of mechanical properties within each insert, miniature disk shaped specimens were machined from the surface and subsurface regions of the inserts. Analysis of variance of the test data showed that aging significantly affected the small punch test measures of elastic modulus, initial load, ultimate load, ultimate displacement, and work to failure. The accelerated aging protocol was unable to reproduce the spatial mechanical profile seen in shelf aged components, but it did mechanically degrade the surface of GUR 1120 tibial components to an extent comparable to that seen after 10 years of natural aging. Test specimens showed a fracture morphology consistent with the decreased ductility and toughness which was corroborated by the small punch test metrics of this study. Our data support the hypothesis that UHMWPE undergoes a spatially nonuniform change towards a less ductile (more brittle) mechanical behavior after gamma irradiation in air and shelf aging. PMID- 10872775 TI - Adhesion of bioactive glass coating to Ti6A14V oral implant. AB - Bioactive glass (BAG) is a bioactive material with a high potential as implant material. Reactive plasma spraying produces an economically feasible BAG-coating for Ti6A14V oral implants. This coating is only functional if it adheres well to the metal substrate and if it is strong enough to transfer all loads. To examine these two properties an appropriate mechanical adhesion test, the moment test, is developed. This test quantifies under a realistic loading condition the corresponding functional adhesion strength to be >84 MPa in tensile. To get a qualitative insight in the BAG-coating behavior during loading the mechanical test was combined with finite element analysis, acoustic emission and microscopic analysis. These analyses showed that the coating withstands without any damage an externally generated tensile stress of 47 MPa. Not only the initial adhesion is determining for the implant quality, but more important is the coating functionality after reaction of the BAG. Adhesion testing after two months of in vitro reaction in a simulated body fluid showed that coating adhesion strength decreased with 10%, but the implant system was still adequate for load-bearing applications. PMID- 10872776 TI - Blood compatible aspects of poly(2-methoxyethylacrylate) (PMEA)--relationship between protein adsorption and platelet adhesion on PMEA surface. AB - Platelet adhesion and spreading is suppressed when a poly(2-methoxyethylacrylate) (PMEA) surface is used, compared with other polymer surfaces. To clarify the reason for this suppression, the relationship among the amount of the plasma protein adsorbed onto PMEA, its secondary structure and platelet adhesion was investigated. Poly(2-hydroxyethylmethacrylate) (PHEMA) and polyacrylate analogous were used as references. The amount of protein adsorbed onto PMEA was very low and similar to that absorbed onto PHEMA. Circular dichroism (CD) spectroscopy was applied to examine changes in the secondary structure of the proteins after adsorption onto the polymer surface. The conformation of the proteins adsorbed onto PHEMA changed considerably, but that of proteins adsorbed onto PMEA differed only a little from the native one. These results suggest that low platelet adhesion and spreading are closely related to the low degree of the denaturation of the protein adsorbed onto PMEA. PMEA could be developed as a promising material to produce a useful blood-contacting surface for medical devices. PMID- 10872777 TI - Neutrophil activation by plasma opsonized polymeric microspheres: inhibitory effect of pluronic F127. AB - The phagocytosis of drug-loaded polymeric microspheres by white blood cells, such as neutrophils or mononuclear cells, represents the major clearance mechanism by which this foreign material is eliminated from the body. The process of phagocytosis requires the activation of the white blood cells by the microsphere surface, followed by binding and engulfment. Phagocytosis may result in the removal of the microspheres from the blood or the disease site and an inflammatory response. Therefore, we have studied the level of neutrophil activation by microspheres ( +/- opsonization) manufactured from various biomaterials or polymers. Polymer microspheres with equivalent size distributions were made from poly (DL-lactic acid) (PLA), poly(epsilon-caprolactone) (PCL), poly(methyl methacrylate) (PMMA) or a 50 : 50 blend of PLA: poly(ethylene-co vinyl acetate) (PLA: EVA). Neutrophils were isolated from human blood and activation of these cells by microspheres was measured by chemiluminescence (CL). All four types of microspheres induced only low levels of CL, however these levels were enhanced significantly if the microspheres were pretreated with plasma or IgG suggesting an opsonization effect. The adsorption of IgG or proteins from plasma was confirmed by polyacrylamide gel electrophoresis (SDS PAGE). The poloxamer Pluronic F127 inhibited the opsonization effect of IgG and plasma on all four types of microspheres and inhibited protein adsorption as measured by SDS-PAGE. Since neutrophil activation is part of the inflammation process in vivo, these in vitro data suggest that all four types of microspheres are likely to be inflammatory if injected into body compartments containing plasma-derived fluids. Pretreatment of the microspheres with Pluronic F127 may reduce the inflammatory potential of the microspheres. PMID- 10872778 TI - Controlled release of swine semen encapsulated in calcium alginate beads. AB - A quick and successful encapsulation method of swine spermatozoa is described: hydroxypropylmethylcellulose and calcium chloride were added to the sampled ejaculate swine sperm (sperm-rich fraction: creamy white) and then this suspension was dropped into an aqueous solution of sodium alginate. In order to obtain different capsule thicknesses, different calcium chloride concentrations were used. The influence of different formulations on in vitro spermatozoa release behavior and on the mechanical properties has been studied. In vitro sperm kinetics (motility and average velocity) have been determined. The results obtained from motility and average velocity tests of treated seminal material are promising, especially if the difficulty of preservation of swine spermatozoa compared to bovine sperm is considered. The different membranes obtained from the different calcium concentrations have had an influence on mechanical properties and on the release profile of spermatozoa from the capsules, and therefore, it is possible to modulate the release rate of the cells. PMID- 10872779 TI - Oral peptide drug delivery: polymer-inhibitor conjugates protecting insulin from enzymatic degradation in vitro. AB - A drug-carrier matrix has been developed which protects embedded insulin from degradation by the luminally secreted serine-proteases trypsin (EC 3.4.21.4), chymotrypsin (EC 3.4.21.1) and elastase (EC 3.4.21.36) in vitro. Increasing amounts of the Bowman-Birk inhibitor (BBI) and elastatinal, respectively, were thereby covalently bound to the mucoadhesive polymer sodium carboxymethylcellulose (Na-CMC). The inhibitory efficacy of resulting polymers was evaluated. On the one hand, all polymer-BBI conjugates showed a strong inhibitory activity towards trypsin and chymotrypsin whereas it was markedly lower towards elastase. The polymer-elastatinal conjugates, on the other hand, displayed a comparatively higher inhibitory activity towards elastase. In an artificial intestinal fluid containing trypsin, chymotrypsin and elastase in physiological concentrations insulin, being incorporated in unmodified Na-CMC, was rapidly degraded at 37 degrees C. Within 1 h 98.7 +/- 0.4% (mean +/- SD, n = 3) of the peptide drug were thereby metabolized. On the contrary, the incorporation of insulin in a mixture of the two polymer-inhibitor conjugates CMC BBI (40%; w/w) and CMC-elastatinal conjugate (60%; w/w) led to a peptide degradation of 22.3 +/- 2.5% (mean +/- SD, n = 3) within the same time period. Even after 4 h of incubation, 33.6 +/- 3.2% (mean +/- SD, n = 3) of the therapeutic agent remained stable towards enzymatic attack. Hence, the polymer inhibitor conjugates described in this study seem to be a useful tool in overcoming the luminal enzymatic barrier in peroral insulin delivery. PMID- 10872780 TI - Physical-mechanical, moisture absorption and bioadhesive properties of hydroxypropylcellulose hot-melt extruded films. AB - The objective of this study was to investigate the moisture absorption, physical mechanical and bioadhesive properties of hot-melt extruded hydroxypropylcellulose (HPC) films containing polymer additives. These additives included polyethylene glycol (PEG) 5%, polycarbophil 5%, carbomer 5%, Eudragit E-100 5%, and sodium starch glycolate (SSG) 5%. Relative humidity (RH) and temperature parameters of the films studied included 25 degree C at 0, 50, 80 and 100% RH, and 40 degrees C at 0 and 100% RH, stored for 2 weeks. Tensile strength and percent elongation were determined on an Instron according to the ASTM standards. The bioadhesive properties of the HPC/PEG 3350 5% film and the polycarbophil 5% containing films, with and without PEG, were investigated in vivo on the human epidermis. Although all films studied exhibited an increase in percent water content as the percent RH increased, the SSG containing film exhibited an almost three-fold increase in percent water content compared to that of the HPC/PEG film. The temperature storage condition of 40 degrees C/100% RH (versus 25 degrees C/100% RH) increased the percent water content of the SSG containing film. Percent elongation was highest for films containing polycarbophil 5% (without PEG). In addition, the HPC film containing polycarbophil 5% exhibited a greater force of adhesion and elongation at adhesive failure in vivo, and a lower modulus of adhesion when compared to the HPC/PEG film. A novel approach to determine bioadhesion of films to the human epidermis is presented. PMID- 10872781 TI - Government criticised over handling of withdrawal of OP dips. PMID- 10872783 TI - Induction of zinc deficiency in sheep and its correction with a soluble glass bolus containing zinc. AB - Balance studies were carried out on four Suffolk-cross lambs which were fed a diet containing only 1.2 mg zinc/kg dry matter; zinc deficiency was induced within three weeks. After a period during which the deficiency was relieved by a pica, the zinc deficient state was re-established. Each sheep was then treated with a soluble glass bolus containing zinc, cobalt and selenium. The plasma zinc concentration of the sheep rapidly increased and was maintained for between six and 10 weeks. The bolus was able to supply the daily requirement of the sheep for zinc, with no detrimental effect on their copper status. PMID- 10872782 TI - Central nervous system pathology in 25 dogs with chronic degenerative radiculomyelopathy. AB - The neuropathology of 20 German shepherd dogs and five German shepherd dog crosses with chronic degenerative radiculomyelopathy were analysed by conventional techniques, immunocytochemistry and electron microscopy. There were previously unrecognised changes in brain nuclei. In the spinal cord, both motor and sensory tracts were involved, principally in their more distal regions. Wallerian degeneration affected the corticorubrospinal pathways in the lateral columns and the ventral funiculi, predominantly in the caudal thoracic and lumbar segments, although more cranial involvement was also observed. The dorsal columns were affected in the caudal lumbar region and the cervical fasciculus gracilis. The regional distribution was variable between cases. Within the brain, abnormalities, including chromatolysis, gliosis and neuronal loss were observed in the red nucleus, lateral vestibular nucleus and, occasionally, in the dentate nucleus. The changes in brain nuclei were compared with those found in dogs at various times after a focal spinal injury. The neuronal changes in the brain may be related to the primary site of damage, and possible aetiological mechanisms are discussed. PMID- 10872784 TI - Detection and differentiation of porcine epidemic diarrhoea virus and transmissible gastroenteritis virus in clinical samples by multiplex RT-PCR. AB - A multiplex reverse-transcriptase-PCR (RT-PCR) procedure was developed for the simultaneous detection of porcine epidemic diarrhoea virus (PEDV) and transmissible gastroenteritis virus (TGEV) in preweaning pigs with diarrhoea. The membrane gene of PEDV and the nucleocapsid gene of TGEV were chosen as targets. The PCR products of PEDV and TGEV had molecular sizes of 412 and 612 base pairs, respectively. Primers from PEDV did not react with any TGEV tested and vice versa. In addition, the primers did not react with other pig viruses. The multiplex RT-PCR was able to detect 10 tissue culture-infective doses 50 per cent (TCID50)/ml of PEDV or TGEV with each of the primer sets for PEDV and TGEV, respectively. The RNAS of PEDV and TGEV were detected directly in intestinal and faecal samples from pigs infected experimentally with either virus. The results of the assay correlated well with the results of virus isolation. None of the five control specimens was positive. PEDV was detected in 10 intestinal and nine faecal samples, and among the nine positive faecal samples two were culture negative. TGEV was also detected in 10 intestinal and nine faecal samples, and among the nine positive faecal samples, three were culture-negative. PMID- 10872785 TI - Collateral ligament prosthesis for the repair of subluxation of the metatarsophalangeal joint in a jersey cow. AB - A two-and-a-half-year-old Jersey cow had been moderately lame in its right hindlimb for one month. Clinical and radiographic examinations revealed an incomplete lateral luxation of the metatarsophalangeal joint. The medial condyle of the metatarsal bone was locked in the condylar groove of the medial first phalanx. The subluxation was reduced, the joint carefully debrided and irrigated, and the torn ligament was replaced with a synthetic prosthesis which was fixed with titanium alloy staples. One year after surgery, the joint was slightly swollen but the cow was not lame. PMID- 10872786 TI - Isolation of Mycoplasma bovis from cattle in Northern Ireland, 1993 to 1998. PMID- 10872787 TI - Histology in recovered cases of grass sickness. PMID- 10872788 TI - Hydronephrosis and renal failure in two Friesian cows. PMID- 10872789 TI - Seroprevalence of Neospora caninum infection in domestic dogs in Turkey. PMID- 10872790 TI - PDNS, PMWS and porcine circovirus type 2 in Scotland. Porcine dermatitis and nephropathy syndrome. Post-weaning multisystemic wasting syndrome. PMID- 10872791 TI - Unseasonal snake bites in dogs. PMID- 10872792 TI - Urinary incontinence in rabbits. PMID- 10872793 TI - Request for UK avian tick samples. PMID- 10872794 TI - Antithrombin III, a serpin family protease inhibitor, is a major heparin binding protein in porcine aqueous humor. AB - Our hypothesis is that the proteins in aqueous humor may be involved in the regulation of outflow facility through the trabecular meshwork and uveoscleral meshwork. In this study, we analyzed the profile of heparin-binding proteins present in porcine aqueous humor to identify and characterize secretory proteins with a binding affinity for heparin. A single step involving heparin-sepharose affinity chromatography of porcine aqueous humor yielded a approximately 60 kDa protein as the major heparin-binding species. This protein was specifically eluted from the column by heparin. The N-terminal sequence and immunological cross reactivity of this protein confirmed its identity as antithrombin III. Aqueous humor from different species, as well as cells from human trabecular meshwork, Schlemm's canal, and lens epithelium, contained detectable amounts of antithrombin III. Based on its known anticoagulative function in endothelial cells and effects on the production of prostacyclin, it is reasonable to speculate that antithrombin III present in aqueous humor might influence the physiology of the trabecular and uveoscleral meshwork and thereby regulate intraocular pressure. PMID- 10872795 TI - Characterization of large plasmids encoding resistance to toxic heavy metals in Salmonella abortus equi. AB - Salmonella abortus equi vaccine strains were found to be resistant to high levels of toxic heavy metals--arsenic, chromium, cadmium, and mercury. The two strains 157 and 158 were resistant to ampicillin also. Curing of these strains resulted in loss of one or more resistance marker indicating plasmid borne resistance. Plasmid profile of strain 157 showed presence of three plasmids of 85, 54, and 0.1 Kb, whereas 158 strain showed presence of 85 Kb and 2 Kb plasmids. Plasmids were isolated from strain 157 and introduced into E. coli DH5alpha with a transformation efficiency of 2 x 10(3) transformants/microg DNA. Interestingly the transformants were resistant to antibiotics, heavy metals (As, Cr, Cd, Hg) and was also able to utilize citrate, a trait specific to Salmonella species. We report and establish for the first time the transferable large plasmids encoding resistance to various heavy metals, antibiotics and biochemical nature of S. abortus equi. PMID- 10872796 TI - Distinctive structural and kinetic properties of an unusual juvenile hormone hydrolyzing esterase. AB - The insect juvenile hormone specific esterases (JHEs), related to acetylcholinesterases but exhibiting substrate specificity for juvenile hormone (JH), are essential enzymes for normal insect development, making them attractive targets for biorationally designed, environmentally safe pesticides. We examine here a new enzyme, JHER, related to, but yet structurally, biochemically, and kinetically distinct from, the classical JHE. Both classical JHE and baculovirus expressed JHER hydrolyze JH show disproportionately higher catalytic rates at higher substrate concentrations (in contrast to substrate inhibition reported for acetylcholinesterase) and are similarly inhibited by an organophosphate. However, JHER, which possesses an unusual cysteine residue at +1 to the catalytic serine, is less sensitive to trifluoromethyl ketone transition state analogs designed around the structure of JH. We propose a model in which JHER is expressed just prior to metamorphosis for hydrolysis of a JH-like substrate with hydrophobic backbone, a proximal ester, and a terminal expoxide or related substitution. PMID- 10872797 TI - Apomorphine up-regulates NGF and GDNF synthesis in cultured mouse astrocytes. AB - Apomorphine, a D1/D2 dopamine agonist, is an anti-parkinsonian drug. We examined the effects of apomorphine on synthesis of neurotrophic factors in cultured mouse astrocytes. After 24 h incubation with apomorphine, NGF and GDNF contents in the culture medium increased to 122-fold and 1.8-fold of the control, respectively; whereas the BDNF content did not change significantly. In Northern blot analysis, expression of NGF mRNA in astrocytes reached the maximum level at 6 h after addition of the drug. By semiquantitative RT-PCR analysis, the GDNF transcript level was found to reach 2.9-fold of the control level at 15 h. These results suggest that apomorphine may exert neuroprotective effects by stimulation of NGF and GDNF synthesis in astrocytes. PMID- 10872798 TI - Analogs of lactam derivatives of alpha-melanotropin with basic and acidic residues. AB - A role of the aromatic and of the basic residues of the potent agonist (MTII) and antagonist (SHU9119) at the human melanocortin receptors 4 in the formation and stabilization of ligand-receptor complexes was examined. Analogs of MTII and SHU9119 with glutamic acid replacing one amino acid at a time were synthesized and tested for their ability to bind to and activate human melanocortin receptors 3, 4, and 5. Replacement of Phe (Nal) or Trp with Glu resulted in analogs of MTII and SHU9119 which were practically inactive at the receptors studied. The rather large (and unexpected) tolerance toward the presence of Glu in the position of His or Arg of MTII and SHU9119 clearly suggested that in the ligand receptor complexes these basic residues are not in contact with the receptors but probably face the extracellular environment. This identified the aromatic residues of MTII and SHU9119 as the primary structural features determining interactions of the agonist/antagonist with hMCR3-5. PMID- 10872799 TI - Zinc-histidine as nucleation centers for growth of ZnS nanocrystals. AB - Histidine is a chelator of zinc, most notably in zinc-finger proteins (zinc coordinated by cysteine and histidine) and in hyperaccumulator plants. Sulfide incorporation into molecules containing metal-cysteinyl complexes has been shown to occur in vivo in certain yeasts, leading to enhanced metal tolerance. Demonstrated here for the first time is incorporation of sulfide into zinc histidine, resulting in histidine-ZnS nanocrystals (NCs) having unique optical properties. Sulfide complexation occurred optimally at alkaline pH into zinc (histidine)2 species, and UV/Vis absorption maxima were red-shifted as increasing sulfide addition occurred. Intermediate sulfide concentrations led to multiple, thermodynamically preferred NC species within a sample. Fluorescence of histidine ZnS NCs was greater than ZnS prepared previously with cysteinyl peptides. Transmission electron microscopy and selected-area electron diffraction indicated hexagonal ZnS crystals having an average size of 4.2 nm. A photocatalytic application of histidine-ZnS NCs was shown by efficient degradation of p nitrophenol and paraquat in the presence of UV irradiation. PMID- 10872800 TI - Association of functional microsatellites in the human type I collagen alpha2 chain (COL1A2) gene with systemic sclerosis. AB - Systemic sclerosis (SSc) or scleroderma is a generalized disorder of connective tissue. The etiology is poorly understood; however, both genetic and environmental factors have been implicated. To investigate the disease susceptible gene for SSc, we examined the association of the disease with a gene (COL1A2) for type I collagen, which accumulates excessively in the affected organs. The COL1A2 gene containing a specific combination of the two dinucleotide repeats, repeat-haplotype, is involved in the regulation of gene expression. Homozygotes for a 5'-(CA)13CGCACA(CG)6(CA)8 -(GT)12 -3' were found with significantly higher frequency (P = 0.029, relative risk, RR > 6.93) in SSc patients than in controls, and association was prominent (P = 0.0042, RR > 32.0) in the male patients positive for SSc-specific antinuclear antibodies (ANAs). This repeat-haplotype showed the highest stimulative activity for the transcription of the COL1A2 promoter among the reporter gene constructs tested. The results indicate that a portion of the patients having a specific dinucleotide repeat-haplotype homozygously and expressing the ANAs have a significantly higher risk for SSc than those individuals with other combinations of the repeat-haplotypes. PMID- 10872801 TI - Distinct hormone stimulation and counteraction by insulin of the expression of the two components of glucose 6-phosphatase in HepG2 cells. AB - We found recently (J. Biol. Chem. 274, 33866-33869, 1999) that the expression of the catalytic subunit (p36) and putative glucose 6-phosphate translocase (p46) of the liver glucose 6-phosphatase system was stimulated by cyclic AMP and glucose and repressed by insulin. We now further show in HepG2 cells that whereas insulin (0.01-10 nM) suppressed p36 mRNA, it only reduced p46 mRNA by half at 1 microM. Cyclic AMP (0.01-100 microM) caused a 2.7-fold increase in p36 mRNA but barely increased p46 mRNA. In contrast, dexamethasone (0.1-100 nM) increased both p36 and p46 mRNA by more than 3-fold. The effects of cyclic AMP and dexamethasone were counteracted by 1 microM insulin. The endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin (1-100 nM) increased p36 mRNA by 2-fold but not p46 mRNA. It thus appears that the hormonal changes which affect p36 alone concur with known modifications in glucose production; those that affect both p36 and p46 are rather consistent with glucose storage. PMID- 10872803 TI - Dynamic measurement of single protein's mechanical properties. AB - Dimerized (tandemly repeated) protein was constructed, and the stretching force during the unfolding of the single protein molecule was measured using an atomic force microscope. In quasistatic measurements using normal force-distance curve measurements, each monomer unit was unfolded step by step. To elucidate the conformational state at each extension length, we measured the relax-stress response of the protein using short stroke sinusoidal movements of the sample stage. This allowed us to investigate the dynamic response of the protein repeatedly without full stretching or rupturing. Although the protein molecule responded in-phase to the applied movement in most cases, we found a novel out-of phase response around the stretching length where the second monomer unit unfolded. Applying the spring constant measured in the quasistatic experiment, the out-of-phase response was reproduced in the simple calculation, which suggested the folding and the unfolding at the second monomer unit were taking place repeatedly during the relax-stress response measurement. PMID- 10872802 TI - Molecular cloning of the mouse APS as a member of the Lnk family adaptor proteins. AB - Engagement of cell-surface receptors leads to activation of protein tyrosine kinases, which in turn phosphorylate various downstream enzymes and adaptor proteins. Lnk is an adaptor protein that appears to be involved in signal transduction in lymphocytes, and forms an adaptor protein family with SH2-B. We tried to identify another member of the adaptor protein family and isolated the mouse APS (adaptor molecule containing PH and SH2 domains). APS contains a proline-rich region, PH and SH2 domains, and a putative tyrosine phosphorylation site at the C-terminal, and the overall structure resembles those of Lnk and SH2 B. APS is expressed in brain, kidney, muscle, and mature B cells in spleen. Mouse APS gene consists of 8 coding exons and is deduced to map to chromosome 5. APS is tyrosine phosphorylated at the C-terminal phosphorylation site conserved among the Lnk family adaptor proteins by stimulation of IL-5 or IL-3 as well as by crosslinking of B cell receptor complex. These results suggest that APS is a member of the Lnk family adaptor protein and likely plays a role in signaling in B cells. PMID- 10872804 TI - 1H NMR conformational study of antiherpetic C5-substituted 2'-deoxyuridines: insight into the nature of structure-activity relationships. AB - 1H NMR study and conformational analysis of a broad series of biologically important C5-substituted 2'-deoxyuridines, including alkyl, halogen, vinyl, hydroxymethyl, and hydroxy derivatives as well as nitro, formyl, trifluoromethyl, and dimethylamino substituents, is presented. A thorough analysis of chemical shifts in correlation with C5-substituent electronegativity as well as calculations by SCF semi-empirical method of the formal charge localized on C6 carbon is discussed in terms of charge distribution for electron attracting and electron donating groups. Conformation of the sugar ring is determined from proton-proton coupling constants and described in terms of pseudorotation between two main puckering domains C2'endo (S) and C3'endo (N). Generally, electron donating groups destabilise the N conformation, simultaneously decreasing the mean pseudorotation amplitude. Absolute assignments of the H5' and H5'' methylene protons in 1H NMR spectra permitted the unequivocal determination of molar fractions of the three classical exocyclic C4'-C5' rotamers gauche+, trans, and gauche-, and correlation of them with the sugar ring puckering domains. Conformation about the glycosidic bond is described in terms of equilibrium between two conformational regions, anti and syn. Finally, the role of the C5 substituent in the creation of cytotoxic activity is considered on the basis of a simplified model assuming that compound activity is a function of substituent polar surface, its molecular volume, and its molecule polarity defined at the relative partition of the polar atoms. PMID- 10872805 TI - 4-Hydroxynonenal-induced MEL cell differentiation involves PKC activity translocation. AB - 4-Hydroxynonenal (HNE) is a highly reactive aldehyde, produced by cellular lipid peroxidation, able to inhibit proliferation and to induce differentiation in MEL cells at concentrations similar to those detected in several normal tissues. Inducer-mediated differentiation of murine erythroleukemia (MEL) cells is a multiple step process characterized by modulation of several genes as well as by a transient increase in the amount of membrane-associated protein kinase C (PKC) activity. Here we demonstrate that a rapid translocation of PKC activity from cytosol to the membranes occurs during the differentiation induced by HNE. When PKC is completely translocated by phorbol-12-myristate-13-acetate (TPA), the degree of HNE-induced MEL cells differentiation is highly decreased. However, if TPA is washed out from the culture medium before the exposition to the aldehyde, HNE gradually resumes its differentiative ability. The incubation of cells with a selective inhibitor of PKC activity, bisindolylmaleimide GF 109203X, partially prevents the HNE-induced differentiation in MEL cells. In conclusion, our results demonstrate that HNE-induced MEL cell differentiation is preceded by a rapid translocation of PKC activity, and that the inhibition of this phenomenon prevents the onset of terminal differentiation. PMID- 10872806 TI - Single mutation induces a metal-dependent subunit association in dimeric Cu,Zn superoxide dismutase. AB - Tryptophan 83, a residue strongly involved in the intersubunit interaction of the Cu,Zn superoxide dismutases from Photobacterium leiognathi, has been selectively mutated to phenylalanine or tyrosine. The recombinant mutant enzymes expressed in Escherichia coli were purified in two well distinct and stable forms, one dimeric and fully active and the other monomeric and devoid of metals. In agreement, in vitro experiments indicate that the removal and addition of zinc in the mutant enzymes induces monomerization and dimerization, respectively, while does not perturb the dimeric association of the native protein. This is the first unambiguous experimental proof of a direct communication between the intersubunit interface and the metal active site. PMID- 10872807 TI - Molecular basis of competition between HSF2 and catalytic subunit for binding to the PR65/A subunit of PP2A. AB - We recently identified the existence of a novel interaction between heat shock transcription factor 2 (HSF2) and the PR65/A subunit of protein phosphatase 2A (PP2A) and showed that HSF2 is able to compete with the PP2A catalytic subunit for binding to PR65. To elucidate the mechanistic basis of this competition between HSF2 and catalytic subunit at the molecular level we have sought to characterize sequences within PR65 that are important for interaction with HSF2. The results identify the intra-repeat loop within HEAT repeat 11 of PR65 as critical for interaction with HSF2. Analysis of point mutants within this loop region of PR65 identify lysine 416 as a residue critical for interaction with HSF2. Interestingly, this same lysine residue of PR65 is important for its binding to catalytic subunit. These results suggest that HSF2's ability to interfere with catalytic subunit binding to PR65 is due to competition between HSF2 and catalytic subunit for at least one amino acid residue of PR65, lysine 416. These data support the hypothesis that HSF2 represents a new type of PP2A regulatory protein. PMID- 10872808 TI - Regulated expression of endothelial cell-derived lipase. AB - A lipoprotein lipase-like gene was recently cloned from endothelial cells. In vitro functional experiments have suggested that this endothelial-derived lipase (EDL) has phospholipase activity, and preliminary in vivo studies have suggested a role in the regulation of high-density lipoprotein metabolism. To investigate local control of lipase activity and lipid metabolism in the blood vessel wall, we have examined the regulation of EDL expression in cultured human umbilical vein and coronary artery endothelial cells. EDL mRNA levels were upregulated in both cell types by inflammatory cytokines implicated in vascular disease etiology, including TNF-alpha and IL-1beta. In addition, both fluid shear stress and cyclic stretch were found to increase the EDL mRNA levels in these cultured cells. This highly regulated expression of EDL in vascular endothelial cells suggests that this recently identified lipase is intricately involved in modulating vessel wall lipid metabolism and may play a role in vascular diseases such as atherosclerosis. PMID- 10872809 TI - Redefined substrate specificity of ST6GalNAc II: a second candidate sialyl-Tn synthase. AB - The acceptor substrate specificities of ST6GalNAc I and II, which act on the synthesis of O-linked oligosaccharides, were reexamined using ovine submaxillary mucin, [Ala-Thr(GalNAc)-Ala]n polymer (n = 7-11). It has been suggested that only ST6GalNAc I can synthesize carbohydrate structures of sialyl-Tn-antigen; i.e., NeuAc alpha2-6GalNAc-O-Thr/Ser [Kurosawa et al., J. Biol. Chem. 269, 19048-19053 (1994)] based on the result that ST6GalNAc I, not ST6GalNAc II, exhibited activity toward asialoagalacto-fetuin. In this study, we present evidence that both ST6GalNAc I and II exhibit activity toward asialo-OSM (ovine submaxillary mucin) and [Ala-Thr(GalNAc)-Ala]n polymer (n = 7-11) which have only the GalNAc-O Thr/Ser-structures. These results strongly indicate that not only ST6GalNAc I but also II are candidates for sialyl-Tn synthases. PMID- 10872810 TI - Retinoic acid switches differential expression of FGF8 isoforms in LNCaP cells. AB - Retinoic acid (RA) is described as an inhibitor of prostate cancer cell growth. We utilized reverse transcription-polymerase chain reaction (RT-PCR) to analyze expression of different isoforms of fibroblast growth factor 8 (FGF8) in response to RA. Results in the prostate cancer cell line LNCaP show that whereas overall expression levels of FGF8 appear to remain constant, RA addition induces an inversion of the ratio between FGF8a and -b mRNAs. Along with this observed "isoform switch," unexpected expression of retinoic acid receptor alpha was detected. Although preliminary, these data allow one to hypothesize on the existence of a possible link between the morphogenic hormone RA and the regulation of the potent mitogen FGF8. PMID- 10872811 TI - Effect of troponin I phosphorylation by protein kinase A on length-dependence of tension activation in skinned cardiac muscle fibers. AB - We examined the effect of troponin I (TnI) phosphorylation by cAMP-dependent protein kinase (PKA) on the length-dependent tension activation in skinned rat cardiac trabeculae. Increasing sarcomere length shifted the pCa (-log[Ca2+]) tension relation to the left. Treatment with PKA decreased the Ca2+ sensitivity of the myofilament and also decreased the length-dependent shift of the pCa tension relation. Replacement of endogenous TnI with phosphorylated TnI directly demonstrated that TnI phosphorylation is responsible for the decreased length dependence. When MgATP concentration was lowered in the absence of Ca2+, tension was elicited through rigorous cross-bridge-induced thin filament activation. Increasing sarcomere length shifted the pMgATP (-log[MgATP])-tension relation to the right, and either TnI phosphorylation or partial extraction of troponin C (TnC) abolished this length-dependent shift. We conclude that TnI phosphorylation by PKA attenuates the length-dependence of tension activation in cardiac muscle by decreasing the cross-bridge-dependent thin filament activation through a reduction of the interaction between TnI and TnC. PMID- 10872812 TI - Anchorage-independent activation of mitogen-activated protein kinase through phosphatidylinositol-3 kinase by insulin-like growth factor I. AB - Insulin-like growth factor I (IGF-I) is a well-established mitogen in human breast cancer cells. We show here that human breast cancer MCF-7 cells, which were prevented from attaching to the substratum and were floating in medium, responded to IGF-I and initiated DNA synthesis. The addition of IGF-I to floating cells induced activation of protein kinase B (PKB)/Akt, as to cells attached to the substratum. In addition, mitogen-activated protein kinase (MAPK)/extracellular response kinase (ERK) and its upstream kinases, ERK kinase (MEK) and Raf-1, were activated by IGF-I in floating cells. While the IGF-I induced activation of PKB/Akt was inhibited by PI3-K inhibitor LY294002 but not by MEK inhibitor PD98059, the activation of both MEK and ERK by IGF-I was inhibited by both. These findings suggest that the IGF-I signal that leads to stimulation of DNA synthesis of MCF-7 cells is transduced to ERK through PI3-K, only when they are anchorage-deficient. PMID- 10872813 TI - Regulation of collagenolytic protease secretion through c-Src in osteoclasts. AB - The role of pp60c-src activity in the synthesis and secretion of the collagenolytic cysteine proteases (CCPs), cathepsin K (CAK), cathepsin L (CAL), and cathepsin B (CAB), by osteoclasts was investigated. Synthesis and secretion of CAL were up-regulated by 1alpha,25-(OH)2D3, but neither those of CAK, dominant relative to CAL, nor CAB, barely detectable, levels changed in the experiments. Though PP1, a pp60c-src inhibitor, had no effect on CCPs synthesis, suppressed the CAK and CAL secretion. Wortmannin, a phosphatidylinositol 3-kinase (PI3 kinase) inhibitor that works as a second messenger for pp60c-src, and cytochalasin B, an inhibitor of actin polymerization, suppressed the secretion of both CAK and CAL without suppressing synthesis. Hydroxyproline release, an indicator of degradation of type-I collagen, and F-actin ring formation, a structure linked to osteoclastic bone resorption, were suppressed by PP1, cytochalasin B or wortmannin. These results suggested inhibition of pp60c-src activity affected the osteoclastic cytoskeleton, which in turn reflected the suppression of bone resorption. PMID- 10872814 TI - Spermatocyte-specific gene excision by targeted expression of Cre recombinase. AB - Transgenic mice carrying the coding sequence of the Cre recombinase, whose expression was driven by the spermatocyte-specific Pgk-2 promoter, were generated. These mice were crossed with a reporter transgenic line, which produces beta-galactosidase depending on the occurrence of loxP-mediated DNA recombination. When DNA of the offspring was analyzed by PCR and Southern blotting, signals that appear after the recombination were detectable only in the testis. Histochemical analyses revealed that beta-galactosidase was present in spermatocytes and spermatogenic cells at later differentiation stages. However, the distribution of the protein was not uniform in all spermatocytes. Analyses of genomic DNA of the next generation indicated that recombination took place in about 70% of spermatogenic cells. From these results, we concluded that this transgenic line possessing Pgk-2-driven expression of the Cre recombinase should be useful for identifying spermatogenic genes that function at or after the spermatocyte stage. PMID- 10872815 TI - Direct evidence of nitric oxide presence within mitochondria. AB - Nitric oxide (NO) has been implicated in the modulation of mitochondrial respiration, membrane potential, and subsequently in apoptosis. Although the presence of a mitochondrial NO synthase (mtNOS) has been described, there is no direct evidence in vivo of the presence of NO within mitochondria. It was the aim of this study to demonstrate the in vivo production of NO within mitochondria. Using the novel fluorometric NO detection system, 4,5-diaminofluorescein diacetate (DAF-2/DA), we observed the presence of NO production in PC12 and COS-1 cells by conventional and confocal fluorescence microscopy. Part of the overall NO signal was colocalized within a subpopulation of mitochondria, labeled with the potential-dependent probe MitoTracker red. These findings demonstrate for the first time that the subcellular distribution of NO production is consistent with the presence of a mitochondrial NOS. Our results provide a new tool to directly study the modulatory role of NO in mitochondrial respiration and membrane potential, in vivo. PMID- 10872816 TI - Visual imaging of ion distribution in human epidermis. AB - The distribution of calcium, magnesium, potassium, sodium, and hydrogen ions in the human epidermis was visualized by blotting to gel containing chemical indicators and the effects of skin barrier disruption were examined. In normal skin, both calcium and magnesium were localized with high concentration in the upper epidermis. EDTA blocked these imaging. The hydrogen ion was also high in the upper epidermis. Sodium did not show obvious gradation in the epidermis. The potassium concentration was the lowest in the upper epidermis. After the barrier disruption, the gradients of calcium, magnesium, and potassium disappeared while the pH gradation was not altered. Observation at a high magnification revealed lower calcium and sodium concentrations in the nucleus. The concentration of magnesium was slightly higher in the nucleus. The novel method of the present study could show the visual image of the ions in frozen tissue without further preparation. PMID- 10872817 TI - Skeletal muscle regeneration is not impaired in Fgf6 -/- mutant mice. AB - FGF6 is a member of the fibroblast growth factor family. The Fgf6 gene is almost exclusively expressed in adult and developing skeletal muscle. We have obtained mice deficient in FGF6 by targeting the Fgf6 gene by homologous recombination. We studied regeneration of adult skeletal muscle in Fgf6 -/- mice derived on a standard inbred background. Muscle degeneration was induced by notexin drug or crush injury. The defect in FGF6 did not modify the kinetics of muscle regeneration. We bred Fgf6 -/- mice with mdx dystrophin deficient mice; Fgf6 -/ :mdx and mdx muscles were similar. Our study suggests that FGF6 does not play a role in muscle regeneration, i.e., in satellite cell proliferation and fusion, or that this role is strictly compensated by other factors, possibly other FGFs. PMID- 10872818 TI - Axin-induced apoptosis depends on the extent of its JNK activation and its ability to down-regulate beta-catenin levels. AB - Axin is a multidomain protein that coordinates a variety of critical factors in Wnt signaling and JNK activation. In this study, we found that overexpression of Axin leads to apoptosis in several cell lines. A mutant Axin (Axin-deltaMID) that does not contain the MEKK1-interacting domain and is not capable of activating JNK, has less apoptotic effect. Together with the observations that dominant negative forms of MEKK1 and JNK1 can attenuate Axin-induced apoptosis, we suggest that JNK activation is required for Axin-mediated apoptosis. Wild-type Axin proteins that can lead to destabilization of beta-catenin are more effective at causing cell death than those constructs (Axin-deltaGSK/beta-cat, Axin deltaRGS/GSK/beta-cat) that are defective in regulation of beta-catenin but still fully capable of JNK activation. Furthermore, enhanced beta-catenin signaling by coexpression of beta-catenin or PP2C alpha attenuate cell death. Taken together, we suggest that the ability of Axin to induce apoptosis is determined by its ability to activate JNK and destabilize beta-catenin. PMID- 10872819 TI - Inhibition of intracellular cathepsin activities and suppression of immune responses mediated by helper T lymphocyte type-2 by peroral or intraperitoneal administration of vitamin B6. AB - We reported that pyridoxal phosphate (PAP), a coenzyme form of vitamin B6, strongly inhibits activities of cathepsin B and weakly inhibits those of cathepsins S, K, and C in vitro. Either intraperitoneal injection or peroral administration of medication doses of vitamin B6 in the diet caused dose dependent inhibition of hepatic cathepsins B, L, S, and C, and the inhibition was exhibited much more significantly in the case of a high protein diet than in a low protein diet. Administration of vitamin B6 induced the suppression of immune responses against ovalbumin (OVA) mediated by helper T lymphocyte type-2, based on the suppression of antigen processing by cathepsin B inhibition, as in the case of CA-074 administration, a cathepsin B specific inhibitor. Ovalbumin dependent production of immunoglobulins IgE, IgG1 and interleukin IL-4 was suppressed by administration of medication doses of pyridoxal (PA) or pyridoxine (PI), while the production of IgG2alpha and interferon (INF)-gamma mediated by helper T lymphocyte type 1 was not changed. Administration of medication doses of vitamin B6 caused the inhibition of intracellular cathepsin B activity due to suppression of the functions of helper T lymphocyte type-2. PMID- 10872820 TI - Down syndrome critical region gene 2: expression during mouse development and in human cell lines indicates a function related to cell proliferation. AB - The isolation of the genes located in chromosome 21 and the characterisation of their function are essential steps towards the understanding of the physiopathological mechanisms involved in Down syndrome. We have used two complementary approaches to characterise the function of the novel gene DSCR2 (Down Syndrome Critical Region gene 2): the isolation and characterisation of the mouse gene homologue to the human DSCR2 gene, and the analysis of the expression of the gene in different human cell lines. We have isolated and characterised a 1012 bp of a mouse cDNA having a high homology to the human DSCR2 gene. The predicted mouse dscr2 protein has an identity of 85.4% as compared to the human protein, indicating that the DSCR2 protein has been conserved during the evolution. However, the amino acid sequence is not homologous to other known proteins, or to known protein domains. The dscr2 gene is expressed throughout all the stages of the mouse embryo development. In the adult mouse the gene is expressed in testis, kidney, liver, brain, heart, skeletal muscle, and pancreas. The expression analysis of the DSCR2 gene in different human tumour derived cell lines indicates that the gene is expressed in all proliferating cell lines tested. The levels of the DSCR2 mRNA correlate with cellular growth of T98G and Jurkat cells in response to different treatments. The expression pattern throughout the foetal development together with the correlation observed with the cell cycle indicates a possible function for the DSCR2 gene related to cell proliferation. PMID- 10872821 TI - A region in the 3' UTR of MnSOD RNA enhances translation of a heterologous RNA. AB - The studies reported in this paper were designed to test the hypothesis that a cis element located in the 3' UTR of manganese superoxide dismutase (MnSOD) RNA, designated MnSOD-response element (MnSOD-RE), is a translational enhancer in vivo. NIH/3T3 cells were transfected with a posttranscriptional reporter construct in which MnSOD-RE was placed 3' of the coding region of chloramphenicol acetyltransferase (CAT); this construct is designated CAT-RMS. Transient transfection of CAT-RMS did not change the concentration of CAT mRNA but increased CAT activity by approximately 400% compared to a control construct, CAT V, which contains approximately the same size of non-MnSOD 3' UTR sequence. Transfection of CAT-RMS had no effect on endogenous MnSOD protein, mRNA, or MnSOD RNA-binding protein activity. Because of its ability to increase translation of a heterologous RNA, MnSOD-RE may be useful in designing expression vectors for in vitro expression systems and in vivo gene therapy. PMID- 10872822 TI - L-152,804: orally active and selective neuropeptide Y Y5 receptor antagonist. AB - Neuropeptide Y (NPY) elicits food intake through the action of hypothalamic G protein-coupled receptors. Previous publications indicate that the Y5 receptor may represent one of these postulated hypothalamic "feeding" receptors. Using a potent and orally available Y5 antagonist L-152,804, we evaluated the involvement of the Y5 receptor in feeding regulation. L-152,804 displaced [125I]peptide YY (PYY) binding to human and rat Y5 receptors with Ki values of 26 and 31 nM, respectively, and inhibited NPY (100 nM)-induced increase in intracellular calcium levels via human Y5 receptors (IC50 = 210 nM). L-152,804 did not show significant affinity for human Y1, Y2, and Y4 receptors at a dose of 10 microM. Intracerebroventricular (i.c.v.) (30 microg) or oral (10 mg/kg) administration of L-152,804 significantly inhibited food intake evoked by i.c.v.-injected bovine pancreatic peptide (bPP, 5 microg; a moderately selective Y4, Y5 agonist) in satiated SD rats. However L-152,804 did not significantly inhibit i.c.v. NPY (5 microg; a Y1, Y2, Y5 agonist)-induced food intake. These findings suggest that L 152,804 is a selective and potent non-peptide Y5 antagonist with oral bioavailability and brain penetrability. In addition, the anorexigenic effects of L-152,804 on bPP-induced feeding revealed participation of the Y5 receptor in feeding regulation, while i.c.v. administration of NPY does not appear to significantly contribute to Y5 stimulated food intake. We conclude that the potent and orally active Y5 antagonist, L-152,804, represents a useful tool to address the physiological role of the Y5 receptor. PMID- 10872823 TI - Nuclear and chloroplast poly(A) polymerases from plants share a novel biochemical property. AB - Poly(A) polymerases are centrally involved in the process of mRNA 3' end formation in eukaryotes. In animals and yeast, this enzyme works as part of a large multimeric complex to add polyadenylate tracts to the 3' ends of precursor RNAs in the nucleus. Plant nuclear enzymes remain largely uncharacterized. In this report, we describe an initial analysis of plant nuclear poly(A) polymerases (nPAPs). An enzyme purified from pea nuclear extracts possesses many features that are seen with the enzymes from yeast and mammals. However, the pea enzyme possesses the ability to polyadenylate RNAs that are associated with polynucleotide phosphorylase (PNP), a chloroplast-localized enzyme involved in RNA turnover. Similar behavior is not seen with the yeast poly(A) polymerase (PAP). A fusion protein consisting of glutathione-S-transferase and the active domain of an Arabidopsis-encoded nuclear poly(A) polymerase was also able to utilize PNP, indicating that the activity of the pea enzyme was due to an interaction between the pea nPAP and PNP, and not to other factors that might copurify with the pea enzyme. These results suggest the existence, in plant nuclei, of factors related to PNP, and an interaction between such factors and poly(A) polymerases. PMID- 10872824 TI - Enhancement by homocysteine of plasminogen activator inhibitor-1 gene expression and secretion from vascular endothelial and smooth muscle cells. AB - In order to elucidate the relationship between homocysteine and the fibrinolytic system, we examined the effect of homocysteine on plasminogen activator inhibitor 1 (PAI-1) and tissue-type plasminogen activator (tPA) gene expression and protein secretion in cultured human vascular endothelial and smooth muscle cells in vitro. PAI-1 mRNA and secreted protein levels were both enhanced by homocysteine in a dose dependent manner, with significant stimulation of PAI-1 secretion observed at concentrations greater than 0.5 mM homocysteine. In contrast, secretion and mRNA expression of tPA were not significantly altered by homocysteine stimulation. Secretion of TGFbeta (transforming growth factor beta) and TNFalpha (tumor necrosis factor alpha), possible regulators of PAI-1 expression and secretion, were not stimulated by treatment with 1.0 mM homocysteine. These results suggests that hyperhomocysteinemia-induced atherosclerosis and/or thrombosis may be caused by homocysteine-induced stimulation of PAI-1 gene expression and secretion in the vasculatures by a mechanism independent from paracrine-autocrine activity of TGFbeta and TNFalpha. PMID- 10872825 TI - Cloning and characterization of the human and mouse PDE7B, a novel cAMP-specific cyclic nucleotide phosphodiesterase. AB - We have identified and characterised a novel member of the PDE7 family of cyclic nucleotide phosphodiesterases (PDE), which we have designated PDE7B. Mouse and human full-length cDNAs were isolated encoding a protein of 446 and 450 amino acids, respectively. The predicted protein sequence of PDE7B showed highest homology (70% identity) to that of PDE7A. Northern blot analysis identified a single 5.5-kb transcript with highest levels detected in brain, heart, and liver. Kinetic analysis of the mouse and human purified recombinant enzymes show them to specifically hydrolyse cAMP with a Km of 0.1 and 0.2 microM respectively. Inhibitor studies show sensitivity to dipyridamole, IC50 of 0.51 and 1.94 microM, and IBMX, IC50 of 3.81 and 7.37 microM, for the mouse and human enzymes, respectively. This shows that dipyridamole is not selective for cGMP over cAMP PDEs as previously believed. Other standard PDE inhibitors including zaprinast, rolipram, and milrinone do not significantly inhibit PDE7B. PMID- 10872826 TI - Gadd45 family proteins are coactivators of nuclear hormone receptors. AB - Gadd45 family genes encode nuclear acidic proteins composed of Gadd45, MyD118, and CR6. Sequence analysis showed that Gadd45 family proteins (Gadd45, MyD118, and CR6) contain LXXLL signature motifs considered necessary and sufficient for the binding of several coactivators to nuclear receptors. Interaction between Gadd45 or CR6 and RXR alpha was confirmed by a two-hybrid test in yeast. Results from a series of GST pulldown assays showed that these Gadd45 family proteins interact with several nuclear hormone receptors including RXR alpha, RAR alpha, ER alpha, PPAR alpha, PPAR beta, and PPAR gamma2 in vitro. Interaction between Gadd45 family proteins and nuclear hormone receptors resulted in modest activation of transactivating function of nuclear hormone receptors in reporter systems. When fused to DNA binding domain of GAL4, Gadd45 and CR6 activated the UAS-mediated transcription in mammalian cells. These results suggest that Gadd45 family proteins bind to nuclear hormone receptors and act as nuclear coactivators. PMID- 10872827 TI - Identification of a new active site for autocatalytic processing of penicillin acylase precursor in Escherichia coli ATCC11105. AB - Penicillin acylase (PA) from Escherichia coli ATCC11105 is a periplasmic heterodimer consisting of a 24 kDa small subunit and a 65 kDa large subunit. It is synthesized as a single 96 kDa precursor and then matures to functional PA via a posttranslational processing pathway. The GST-PA fusion protein expression system was established for monitoring the precursor PA processing in vitro. The purified PA precursor was processed into mature PA the same way as in vivo, but pH dependently. From the primary sequence analysis, we identified a putative conserved lysine residue (K299) responsible for the pH dependent processing. The substitution of K299 residue by site-directed mutagenesis affected both the enzyme activity and the precursor PA processing in vivo. Furthermore, it was shown that the processing rates of wild-type and mutant precursor PAs depended on the pKa values of their side chain R group. These results demonstrated that the lysine residue (K299) was involved in the precursor processing of PA together with N-terminal serine residue (S290) of the large subunit. PMID- 10872828 TI - A novel isoform of a kallikrein-like protease, TLSP/hippostasin, (PRSS20), is expressed in the human brain and prostate. AB - cDNAs encoding two splicing variants of a serine protease, termed hippostasin, were isolated by a PCR-based cloning strategy. The difference of 5' nucleotide sequence resulted in the variation in the amino terminal ends of the two, brain and prostate, types of human hippostasin. The longest ORF of the brain-type was 250 amino acids with a putative signal peptide, while that of the prostate-type was 282 amino acids. Homology search using the amino acid sequence revealed that prostate-type hippostasin was identical to TLSP (PRSS20), which is expressed in human primary keratinocytes (1). Transient expression analysis showed that both brain- and prostate-type TLSP/hippostasin were secreted into the conditioned medium as about 40 kDa proteins. Human TLSP/hippostasin showed 47% and 45% identity to trypsinogen II and kallikrein, respectively. In fact, the recombinant human TLSP/hippostasin efficiently cleaved Bz-Phe-Arg-4-methylcoumaryl-7-amide, a kallikrein substrate, and weakly cleaved other substrates for kallikrein and trypsin. Northern blot analysis detected a 1.3 kb band in the whole brain and a 1.4 kb band in the prostate and the lung. In situ hybridization revealed that it was expressed preferentially by the pyramidal neurons in the human hippocampus and secretory epithelial cells in the prostate. These results indicated that TLSP/hippostasin is involved in the functions of the human central nervous system and prostate and that it is a multifunctional protease present in various organs. PMID- 10872829 TI - Resveratrol is absorbed in the small intestine as resveratrol glucuronide. AB - We have studied the absorption and metabolism of resveratrol in the jejunum in an isolated rat small intestine model. Only small amounts of resveratrol were absorbed across the enterocytes of the jejunum and ileum unmetabolised. The major compound detected on the serosal side was the glucuronide conjugate of resveratrol (96.5% +/- 4.6 of the amount absorbed) indicating the susceptibility of resveratrol to glucuronidation during transfer across the rat jejunum. The presence of the glucuronide was confirmed using HPLC-PDA and nanoES-MS/MS techniques. These findings suggest that resveratrol is most likely to be in the form of a glucuronide conjugate after crossing the small intestine and entering the blood circulation. This will have important implications for the biological functions of resveratrol in vivo. PMID- 10872830 TI - Noninsecticidal parasporal proteins of a Bacillus thuringiensis serovar shandongiensis isolate exhibit a preferential cytotoxicity against human leukemic T cells. AB - A Bacillus thuringiensis isolate, 89-T-34-22, belonging to the serovar shandongiensis (H22) produced noninsecticidal and nonhemolytic proteins crystallizing into irregular-shaped parasporal inclusions. The proteins showed in vitro cytotoxicity to human cells, including cancer cells, only when activated by protease treatment. The human leukemic T (MOLT-4) cells were > 100 times more susceptible than HeLa and normal T cells to the proteins of 89-T-34-22. The cytotoxicity was dose dependent and the median effective concentration for the MOLT-4 was 3.5 microg/ml. The cytopathy induced by the 89-T-34-22 proteins was characterized by remarkable condensation of the nucleus and cell-ballooning. Five major parasporal proteins of 89-T-34-22, with molecular masses in the range of 16 160 kDa, shared no similarity with the previously reported proteins in terms of the N-terminal sequence. PMID- 10872831 TI - Molecular cloning and characterization of CRLM-2, a novel type I cytokine receptor preferentially expressed in hematopoietic cells. AB - A murine expressed sequence tag (EST) showing homology with erythropoietin receptor (EPOR) was identified in the EST database. Cloning of the full-length cDNA revealed a 359 amino acid novel type I cytokine receptor, designated cytokine receptor like molecule-2 (CRLM-2). While CRLM-2 lacks typical WSXWS motif, it has a significant homology with EPOR, IL-2 receptor beta and gamma, and IL-9 receptor alpha. The murine CRLM-2 gene is composed of 8 exons, and an alternative mRNA splicing generates a variant transcript encoding a soluble CRLM 2. CRLM-2 is preferentially expressed in hematopoietic cells, particularly in hematopoietic progenitors and myeloid cells. Furthermore, CRLM-2 is constitutively associated with JAK2, a well-known tyrosine kinase that transmits signals from cytokine receptors. These data strongly suggest that CRLM-2 is a novel cytokine receptor involved in the regulation of hematopoietic system. PMID- 10872832 TI - Improved expression of vascular endothelial growth factor by naked DNA in mouse skeletal muscles: implication for gene therapy of ischemic diseases. AB - We have constructed an expression vector, pCK, that is able to drive high levels of gene expression in the skeletal muscles of mice. pCK contains not only the full length immediate-early (IE) promoter of human cytomegalovirus but also its entire 5' untranslated region upstream from the start codon of the IE gene. In addition, pCK contains the kanamycin resistance gene, but lacks nucleotide sequences unnecessary for its function as a gene delivery vector, allowing the plasmid size to be 3.7 kb. pCK produced significantly higher levels of vascular endothelial growth factor 165 both in vitro and in vivo than the control vector, the structure of which is similar to naked DNA vectors employed in previous gene therapy trials. pCK would not only significantly increase the therapeutic effects of naked DNA gene therapy but also dramatically cut down the costs for production and treatment. PMID- 10872833 TI - Decomposition of cocoa procyanidins in the gastric milieu. AB - There is considerable interest in the bioavailability of flavonoids and phenolic components of the diet and their bioactivity in vivo. However, little is known of pre-absorption events in the gastric lumen. The effects of the acidic environment, as found in the gastric milieu, on procyanidin oligomers of catechin polyphenols has been investigated. The results show that under these conditions the procyanidin oligomers (trimer to hexamer) are hydrolysed to mixtures of epicatechin monomer and dimer, thus enhancing the potential for their absorption in the small intestine. PMID- 10872834 TI - Enzymatic properties, tissue-specific expression, and lysosomal location of two highly homologous rat SULT1C2 sulfotransferases. AB - We have isolated two highly homologous but distinct rat sulfotransferase cDNAs termed ratSULT1C2 and ratSULT1C2A encoding polypeptides of 297 amino acids each. The amino acid sequence of ratSULT1C2 is 84% identical to the human SULT1C2 and 81% identical to a rabbit SULT1C2 sulfotransferase. ratSULT1C2 and ratSULT1C2A are 92% identical but differ in 22 amino acids. The majority of these amino acid substitutions in ratSULT1C2A is not found in the human and rabbit SULT1C2, which identifies ratSULT1C2 as the orthologue of these sulfotransferases, whereas SULT1C2A is a closely related but distinct enzyme. ratSULT1C2 and 2A sulfotransferases do not sulfonate steroids, dopamine, acetaminophen, or alpha naphthol, but only p-nitrophenol. Prokaryotically expressed ratSULT1C2A is less active than ratSULT1C2. ratSULT1C2/2A mRNAs are abundant in kidney and less abundant in stomach and liver. The enzymes are expressed as 34-kDa polypeptides in rat kidney, liver, and stomach. In addition, a 28-kDa cross-reacting polypeptide is found in kidney only. Immunohistochemistry revealed expression of ratSULT1C2/2A in the epithelial cells of the proximal tubules of the kidney, bile duct epithelia, hepatocytes, and the epithelium of the gastric mucosal glands. Although the cDNA predicted amino acid sequence identifies both sulfotransferases as cytosolic enzymes, in tissue sections, in the kidney cell line NRK 52, and in transiently transfected BHK cells a considerable fraction of the enzyme was found in a granular perinuclear compartment. Costaining with a lysosomal marker in gastric mucosa tissue sections and cultured cells identifies these structures as lysosomes. PMID- 10872835 TI - DNA annealing and DNA-protein interactions by capillary electrophoresis. AB - This work deals with annealing of single-stranded DNA and the binding of a serum respond factor to a DNA probe containing specific binding site. Capillary electrophoresis (CE) method is explored and compared with the mobility-shift gel electrophoresis (GE) procedure. The results indicate the CE method offers direct and rapid annealing of the DNA strands. It requires no prior incubation with additives (polynucleotides, proteins) to reduce nonspecific DNA-protein interactions. Unwanted nonspecific interactions are not observed in the CE method. The presence of a fluorescein tag to the DNA probe yields identical results to those with the radioactive label. A fluorescein tag in the CE work can be used without any adverse effects. The dissociation constant (Kd) of this protein-DNA complex by the CE method was similar to those determined by the GE method (approximately 10(-6) M). The proposed method is extremely powerful, highly sensitive, quantitative, and fast. It can determine even very small conformational differences of the DNA probe. PMID- 10872836 TI - Chimeric synthetic peptide as antigen for immunodiagnosis of HIV-1 infection. AB - One chimeric peptide incorporating antigenic sequences from the gp41 transmembrane region (peptide H-18) and the gp120 envelope region (peptide H-15) corresponding to amino acids (587-617) on gp41 and (495-516) on gp120 of human immunodeficiency virus (HIV 1) was synthesized. Both sequences were separated by two glycine residues. This peptide was evaluated as antigen in an ultramicro enzyme-linked immunosorbent assay (UMELISA) with samples derived from HIV-1 (n = 30) with different titers of antibodies and healthy blood donors (n = 30). The results were compared to plates coated with monomeric peptides and to plates coated with two monomeric peptides together. Results demonstrated that monomeric peptides gp41 (H-18) and gp120 (H-15) were good as antigens with samples that present antibodies to these regions. The chimeric peptide was the most antigenic. Those results may be related to the peptide structure, adsorption to the solid surface, and epitope accessibility to the antibodies. This chimeric peptide would be very useful for HIV-1 diagnostics. PMID- 10872837 TI - A palmitoyl-CoA-specific delta9 fatty acid desaturase from Caenorhabditis elegans. AB - Biosynthesis of polyunsaturated fatty acids in C. elegans is initiated by the introduction of a double bond at the delta9 position of a saturated fatty acid. We identified three C. elegans fatty acid desaturase genes related to the yeast delta9 desaturase OLE1 and the rat stearoyl-CoA desaturase SCD1. Heterologous expression of all three genes rescues the fatty acid auxotrophy of the yeast delta9 desaturase mutant ole1. Examination of the fatty acid composition of the transgenic yeast reveals striking differences in the substrate specificities of these desaturases. Two desaturases, FAT-6 and FAT-7, readily desaturate stearic acid (18:0) and show less activity on palmitic acid (16:0). In contrast, the other desaturase, FAT-5, readily desaturates palmitic acid (16:0), but shows nearly undetectable activity on the common delta9 substrate stearic acid. This is the first report of a palmitoyl-CoA-specific membrane fatty acid desaturase. PMID- 10872838 TI - Identification of a 26S proteasome-associated UCH in fission yeast. AB - We have identified a 26S proteasome-associated ubiquitin carboxyl-terminal hydrolase (UCH) in Schizosaccharomyces pombe. The gene (designated uch2+) encodes a protein containing a UCH catalytic domain at its N-terminus and a short extension at its C-terminus. uch2+ is nonessential as the uch2 null mutant strain showed no significant difference from the wild-type strain. The GFP-tagged Uch2p is localized predominantly to the nuclear periphery, which is similar to the 26S proteasome localization. Deletion of the C-terminal extension of Uch2p resulted in a drastic change of its subcellular localization: it showed a generally diffused distribution instead of a perinuclear pattern. Glycerol gradient centrifugation analysis and coimmunoprecipitation studies of fission yeast extracts using anti-Mts4p antiserum suggest that Uch2p is associated with the 26S proteasome and the association of Uch2p with the 26S proteasome is mediated by its C-terminal extension. PMID- 10872839 TI - Activation of the chemotactic peptide receptor FPRL1 in monocytes phosphorylates the chemokine receptor CCR5 and attenuates cell responses to selected chemokines. AB - FPRL1 is a seven-transmembrane (STM), G-protein coupled receptor which was originally identified as a low affinity receptor for the bacterial chemotactic formyl peptide and a high affinity receptor for the lipid metabolite lipoxin A4. We recently discovered that a number of peptides, including several synthetic domains of the HIV-1 envelope proteins and the serum amyloid A, use FPRL1 to induce migration and calcium mobilization in human monocytes and neutrophils. In this study, we report that a synthetic peptide domain of the V3 region of the HIV 1 envelope gp120, activates the FPRL1 receptor in monocytes and neutrophils. Furthermore, monocytes prestimulated with V3 peptide showed reduced response to several chemokines that use multiple cell receptors. This is associated with a rapid phosphorylation of the chemokine receptor CCR5 on the serine residues. Our study suggests that FPRL1, as a classical chemoattractant receptor, may play an important role in modulating monocyte activation in the presence of multiple stimuli. PMID- 10872841 TI - Phytanyl-pyrophosphate-linked substrate for a bacterial alpha mannosyltransferase. AB - The biochemical characterization of bacterial glycosyltransferases involved in the assembly of cell-wall-associated polysaccharides is often hindered by the lack of the appropriate undecaprenyl-pyrophosphate-linked acceptor substrate. In order to find a suitable synthetic substrate for the alpha1,3-mannosyltransferase AceA from Acetobacter xylinum, phytanyl-pyrophosphate-linked cellobiose was prepared. In the presence of GDP-[14C]mannose and recombinant AceA, the phytanyl pyrophosphate-linked cellobiose afforded a 14C-labeled trisaccharide that was sensitive to alpha-mannosidase degradation in a fashion analogous to the natural undecaprenyl-pyrophosphate-linked cellobiose substrate. These results suggest that phytanyl-pyrophosphate-linked oligosaccharides may be useful substrates for other important bacterial glycosyltransferases. PMID- 10872840 TI - Multiple copies of beta-lactoglobulin promoter do not function as LCR. AB - Increasing the number of transcription factor binding sites within a construct can enhance expression. In an attempt to create a synthetic locus control region for mammary expression, we have generated beta-lactoglobulin-reporter constructs with multiple copies of the cluster of transcription sites normally located within the proximal promoter. These constructs were functionally tested by stable transfection of mammary epithelial cells in vitro and in transgenic mice in vivo. Rather than enhancing expression, multimerisation of the promoter region acted neither in vivo nor in vitro to enhance expression. Indeed, its presence reduced expression. This failure to enhance expression was reflected in the inability of this region to form a DNaseI hypersensitive site autonomously in mammary chromatin in vivo. It is implicit from our study that not all combinations of transcription factor binding sites will enhance transcription. PMID- 10872842 TI - Identification of novel membrane and secreted proteins upregulated during adipocyte differentiation. AB - Adipose tissue is the largest organ in the body that secretes soluble proteins such as cytokines. A preadipocyte cell line 3T3-L1 has been widely used for investigations of mechanisms of adipocyte differentiation. 3T3-L1 cells convert to adipocytes in the presence of 1-methyl-3-isobutylxanthine, dexamethasone, and insulin. We screened a cDNA library derived from differentiated 3T3-L1 cells, using the SST-REX method (signal sequence trap by retrovirus-mediated expression screening method). Screening of 4 x 10(5) clones gave rise to 63 known and 8 novel clones. The known clones represented 28 independent proteins, 21 of which were secreted proteins and 7 were membrane proteins. The novel clones represented 7 independent proteins, 5 of which had no similarity to known proteins. Interestingly, most of these novel genes showed differentiation- and tissue specific expression. The present results indicate that adipocytes specific genes or adipocyte differentiation-related genes encoding membrane and secreted proteins can be readily identified if signal sequence trap screening of differentiated adipocyte-derived cDNAs is done. PMID- 10872843 TI - Control of smooth muscle cell proliferation and phenotype by integrin signaling through focal adhesion kinase. AB - Extracellular matrix proteins such as fibronectin (FN) and laminin (LM) are known to help control the growth and phenotype of vascular smooth muscle cells (VSMCs). Here we have analyzed the relationship between growth factor and integrin signaling pathways in VSMCs. Culturing porcine coronary artery smooth muscle cells (PCASMCs) on FN and LM leads to distinct effects on cell proliferation and contractile protein expression. PCASMCs cultured on FN proliferate at a higher rate than cells cultured on LM, regardless of the growth factor used to support proliferation. Moreover, cells cultured on LM show higher levels of expression of smooth muscle myosin heavy chain (a marker of smooth muscle cell differentiation) than cells cultured on FN. In contrast to the effects on proliferation and contractile protein expression, both FN and LM supported cell migration in response to PDGF. Also, both FN and LM supported activation of ERK1 and ERK2 in response to PDGF and bFGF. However, FN and LM did show a difference in their ability to support signaling through the focal adhesion kinase (FAK). PCASMCs cultured on FN show robust activation of FAK in response to either PDGF or bFGF, however, cells cultured on LM show little-to-no activation of FAK in response to the growth factors. The results show that integrin signaling pathways have a profound effect on VSMC proliferation and phenotype, and that FAK is an important intermediate in these signaling pathways. The implications of our findings on the mechanisms controlling VSMC proliferation and phenotype in pathological states such as atherosclerosis and restenosis are discussed. PMID- 10872844 TI - Monospecific antibodies against the three mammalian fast limb myosin heavy chains. AB - Skeletal muscle fibres in mammalian limb muscles are of four types: slow, 2A, 2X, and 2B, each characterized by a distinct myosin heavy chain (MyHC) isoform. Existing monoclonal antibodies (mabs) against fast MyHCs lack fibre-type specificity across species and could not positively identify 2X fibres. In this work, mabs were raised against each of the fast MyHCs. These mabs were shown to be monospecific by Western blots and immunohistochemistry in the rat. The advantages of using these mabs for identifying the three fast fibre types and hybrid fibres expressing multiple isoforms were illustrated using rat tibialis anterior muscle. Immunohistochemical analyses confirmed the monospecificity of these mabs in the following additional species: mouse, guinea pig, rabbit, cat, and baboon. 2B fibres were absent in limb muscles of the cat and baboon. These mabs constitute a set of powerful tools for studying muscle fibre types and their transformations. PMID- 10872845 TI - Troponins in renal dysfunction. PMID- 10872846 TI - Selection of graft material for prosthetic arteriovenous access. PMID- 10872847 TI - Membrane compatibility, flux and HCV infection in dialysis patients: newer evidence. PMID- 10872848 TI - Experimental model for studying the effects of 2-ethylhexyl-phthalate and dialysate on connective tissue. AB - In order to have a model for studying the possible implications of 2-ethylhexyl phthalate and dialysate on connective tissue, we evaluated their direct effects on the air pouch lining tissue and on fibroblast cultures. Air pouches were formed on the backs of 60 ten-week-old Wistar rats by subcutaneous injections of 10 ml sterile air. On the tenth day 2 ml sterile air, or 2 ml 5 microg/L or 2 ml 10 microg/L 2-ethylhexyl-phthalate in olive oil, or 2 ml olive oil alone, or 2 ml 5 mg/ml or 12 mg/ml lyophilized dialysate were injected into the air pouches. After sampling at seven or twenty-one days, the rats were killed. The biochemical data showed an increase in sulphated glycosaminoglycans with 2-ethylhexyl phthalate and dialysate. Electron microscopy findings revealed cellular alterations such as vacuolation and cell remnants with 2-ethylhexyl-phthalate, while the cells of the air pouches treated with dialysate showed regular organelles with increased and dilated cisternae of rough endoplasmic reticulum. Moreover, an increase in collagen fibres surrounding the damaged zones was noticed in 2-ethylhexyl-phthalate and dialysate treated rats. The glycosaminoglycan modifications and collagen fibre increase seem to suggest that the morphological changes, with the features of fibrosis, could be the result of 2-ethylhexyl-phthalate and dialysate action on connective tissue. Moreover, the air pouch technique can be considered a good model for studying the direct effects of 2-ethylhexyl-phthalate and other substances, such as uremic toxins, on connective tissue. PMID- 10872849 TI - Biochemical assessment and clinical evaluation of a non-ionic adsorbent resin in patients with intractable jaundice. AB - We investigated in vitro and in vivo the ability of a non-ionic adsorbing resin (styrenedivinylbenzene copolymer) to remove bilirubin and bile acids from human plasma. In preliminary experiments, human plasma from healthy donors, enriched in conjugated bile acids and bilirubin, and pooled plasma from jaundiced patients were recirculated through the resin column. The removal of bilirubin and bile acids was evaluated at two different flow rates (200 ml/min and 40 ml/min), and compared to an activated charcoal column. Four patients with severe jaundice were subsequently treated by 4-hour plasmaperfusion through the resin. The in vitro studies showed that after 1 hour the removal of bile acids was almost complete and bilirubin level decreased significantly, reaching a plateau after 4 hours. In the in vivo study, all treatments were well tolerated. After plasmaperfusion, serum bile acid levels decreased by 64.9-94.6% and total bilirubin by 35.3-57.7%. No clinical or biochemical side effects were observed. Our data suggest that plasmaperfusion through this resin is safe and efficient for removal of bilirubin and bile acids in jaundiced patients. Thus, it may serve as a method of artificial liver support in the treatment of cholestatic syndromes. PMID- 10872850 TI - Trillium biopassive surface: a new biocompatible treatment for extracorporeal circulation circuits. AB - 139 patients undergoing cardiac surgery were included in a prospective, randomized trial. Patients were randomly allocated to receive cardiopulmonary bypass (CPB) with Trillium Biopassive Surface (TBS Group) coated oxygenators or conventional circuits (control group). 112 patients were studied with respect to postoperative biochemical profile; a subgroup of 27 patients was studied with respect to perioperative complement (C3a) activation. Patients in the TBS group demonstrated a significantly lower white blood cell count at the end of the operation (p=0.036) and a significantly higher platelet count the day after the operation (p=0.023) when compared to the control group. C3a was significantly higher (p=0.02) in the TBS group after 30 minutes of CPB, but the C3a increase after protamine administration was significantly less pronounced in the TBS group vs. the control group. Further studies involving platelet and leukocyte activation are required to better elucidate the action of this new coating in the setting of routine CPB. PMID- 10872851 TI - Development of acute ischemic heart failure in sheep. AB - The goal of the present study was to develop a large animal model of acute ischemic left ventricular heart failure (LVHF) that can be used to assess the influence of the PUCA pump on the heart and circulatory system under realistic conditions. We tested the hypothesis that mild stenosis of the coronary artery in combination with mild ventricular pacing induces an acute heart failure condition, whereas the separate phenomena themselves do not lead to impaired heart function. Mean aortic pressure (AoP), left ventricular end-diastolic pressure (LVEDP), stroke volume (SV) and myocardial systolic shortening (MSS) were compared 30 minutes after a pacemaker (PM) induced tachycardia in anaesthetized sheep (n=3) without and with +/- 50% stenosis of the proximal LCx. All parameters measured restored to basic levels when stenosis was absent. When the LCx was partially occluded, mild PM-induced tachycardia resulted in decreased AoP (P=0.045) as well as in decreased SV (P=0.048); the LVEDP remained high (P=0.002). Also the recovery of MSS was impaired when stenosis was present (P=0. 002). These values indicate that acute heart failure conditions were present. The technique used proved to be safe and allowed fine-tuning of the demand ischemia by adapting heart frequency to the required heart failure conditions. The model can be used to study the effect of LV mechanical support during acute heart failure conditions. PMID- 10872852 TI - A resorbable biomaterial shaped as a tubular chamber and containing stem cells: a pilot study on artificial bone regeneration. AB - In a previous study, we showed how healing of non-union defects in rabbit radii can be achieved by means of a tubular resorbable chamber, in comparison with untreated defects. In the present study, we placed bone marrow stem cells inside the chamber. Bone marrow was obtained by percutaneous aspiration from the iliac crest in 9 adult New Zealand rabbits. Stem cells were separated by the centrifugation technique. In the same animals, a defect of 10 mm was created in both radii. On the left side, the defect was treated with the poly-DL-Lactide chamber, in which a suspension of autologous cells was injected; on the right side, only autologous cells were used. Radiological and histomorphometric data were compared within this study as well as with the results of our previous study. At 3, 6 and 9 months, there was no healing on the right side..On the left side, progressive bone formation with reunion of the stumps was observed in the chamber. We conclude that stem cells can accelerate bone healing when contained in the tubular chamber. PMID- 10872853 TI - Evaluation of microparticle leakage during DALI LDL adsorption in a simulated clinical setting. AB - The aim of the present study was to investigate microparticle (Mp) leakage during simulated LDL-hemoperfusion using 12 DALI 750 adsorbers and the original DALI hardware under conditions strictly comparable to the clinical situation. Thus, the sessions were divided into 4 sections, i.e. priming and preparation of the adsorber followed by treatment (6-7 L at a flow rate of 60 ml/min) and reinfusion. As Mp counts can be performed only in clear, cell-free media, blood was replaced by normal saline in sections 2-4 of the simulated sessions. Mp counts were analysed for > or = 2, > or = 5, > or = 10 and > or = 25 microm particle sizes in the efferent line post adsorber using a standard light blockage method. As there are no official thresholds for particle release in extracorporeal circuits, the limits for infusion of large fluid volumes of 500, 100 (80), 25 and 5 (3) Mp/ml according to the Europaische Arzneimittelbuch, the British and American Pharmacopoeias were used. Mean particle counts for the sections 3 and 4 in which the patient is connected to the efferent line were 19, 7, 2 and 0 Mp/ml and amounted to < 10% of the above mentioned limits. Modifications of the standard simulation procedure by inserting additional pump stops or using different flow rates during the treatment phase slightly increased Mp leakage, but never exceeded the prescribed limits. In summary, no undue particle release could be detected during simulations of the clinical DALI LDL adsorption procedure. PMID- 10872854 TI - Aachen-Keratoprosthesis as temporary implant. Case report on first clinical application. AB - BACKGROUND: The Aachen-Keratoprosthesis was designed to serve as a permanent keratoprosthesis, modeling natural corneal physical properties as closely as possible. Prior to permanent application in patients, keratoprostheses are commonly tested in animal models to assess biochemical and biomechanical compatibility. However, immune and inflammatory responses acquired through animal experimentation are difficult to extrapolate in order to develop a predictable and generalized outcome in humans. Therefore, this preliminary report includes results following a temporary implantation of the Aachen-Keratoprosthesis in a patient during vitreoretinal surgery to assess the long-term prospect of application as a permanent artificial cornea. PATIENT AND METHODS: A 43 year old man presented with a ruptured right eye resulting in an opaque cornea and retinal detachment. A soft silicone rubber keratoprosthesis, the Aachen-Keratoprosthesis, was temporarily implanted. Subretinal membranes were removed and the total retinal detachment was reattached. Liquid perfluorocarbon and silicone oil were used. The temporary keratoprosthesis was replaced by a 7 mm corneal graft after completion of surgery. RESULTS: The Aachen-Keratoprosthesis was securely positioned into the trephined hole. It allowed complete visualisation of vitreous, retina and thus controlled manipulations in the vitreous cavity up to the extreme periphery. Leakage across the trephination-prosthesis interface was minimal. CONCLUSION: We report the first temporary application of an innovative keratoprosthesis. Its flexibility and good optical qualities allow visualisation and control of intraoperative procedures. This temporary pilot study of the Aachen KPro encourages further investigation of the Aachen KPro as a permanent replacement for a diseased cornea. PMID- 10872855 TI - Sampling methods for phlebotomine sandflies. AB - A review is presented of methods for sampling phlebotomine sandflies (Diptera: Psychodidae). Among approximately 500 species of Phlebotominae so far described, mostly in the New World genus Lutzomyia and the Old World genus Phlebotomus, about 10% are known vectors of Leishmania parasites or other pathogens. Despite being small and fragile, sandflies have a wide geographical range with species occupying a considerable diversity of ecotopes and habitats, from deserts to humid forests, so that suitable methods for collecting them are influenced by environmental conditions where they are sought. Because immature phlebotomines occupy obscure terrestrial habitats, it is difficult to find their breeding sites. Therefore, most trapping methods and sampling procedures focus on sandfly adults, whether resting or active. The diurnal resting sites of adult sandflies include tree holes, buttress roots, rock crevices, houses, animal shelters and burrows, from which they may be aspirated directly or trapped after being disturbed. Sandflies can be collected during their periods of activity by interception traps, or by using attractants such as bait animals, CO2 or light. The method of trapping used should: (a) be suited to the habitat and area to be surveyed, (b) take into account the segment of the sandfly population to be sampled (species, sex and reproduction condition) and (c) yield specimens of appropriate condition for the study objectives (e.g. identification of species present, population genetics or vector implication). Methods for preservation and transportation of sandflies to the laboratory also depend on the objectives of a particular study and are described accordingly. PMID- 10872856 TI - Insecticide-treated cattle for tsetse control: the power and the problems. AB - Trypanosomiasis control increasingly involves financial input from livestock owners and their active participation. If control is carried out on smaller scales than in the past, methods such as aerial and ground spraying and sterile insect techniques will have reduced application. There will be increased reliance on trypanocidal drugs, and bait methods of tsetse control--where flies are attracted to point sources and killed. If drug resistance develops, cheap and simple bait methods offer the only means of disease control that might be applied, and paid for, by stockowners themselves. The methods have been effective in some circumstances, but not in others, and it is important to understand the reasons for the successes and the failures. Analysis is presented of the results of two Tanzanian tsetse control campaigns involving the use of insecticide treated cattle. Between 1991 and 1996, following the introduction of widespread dipping in the Kagera Region, trypanosomiasis declined from >19000 cases to <2400 and deaths from >4000 to 29. On four ranches in the region, tsetse have been almost eliminated and trypanosomiasis prophylaxis is no longer used. Similarly aggressive use of pyrethroids on Mkwaja Ranch in Tanga Region has not had such dramatic effects. Tsetse and trypanosomiasis are still common, despite high levels of prophylaxis and the deployment of approximately 200 odour-baited targets. The difference in the results is attributed to a combination of the much smaller area covered by treated animals at Mkwaja, a greater susceptibility to re invasion and a more suitable habitat for the flies. A better understanding of the dynamics of the use of insecticide-treated cattle is needed before we can predict confidently the outcome of particular control operations. PMID- 10872857 TI - Life-cycle stage morphology of Psoroptes mange mites. AB - Detailed life-cycle stage descriptions for the ectoparasitic mite Psoroptes ovis (Hering) (Acari: Psoroptidae) from rabbit hosts (syn. Psoroptes cuniculi) are presented. The results resolve a number of contradictions in the literature relating to the recognition of the life cycle stages of these mites. This study supports the view that there are two distinct male nymphal stages, both lacking dorsoposterior tubercles. The male tritonymph is significantly larger than the protonymph and has five pairs of metapodosomal setae rather than three. In addition, male tritonymphs have two pairs of cuticular pits on the central metapodosoma rather than the single pair of the protonymph. The results also show that the female protonymph can be distinguished from the male nymphal stages and the female tritonymph. Both female nymphal stages possess dorsoposterior tubercles, but the protonymph is significantly smaller than the tritonymph. In addition, the protonymph possesses three pairs of metapodosomal setae rather than five, one pair of cuticular pits rather than two, and a pulvillus on leg IV which is absent in the female tritonymph. The presence of dorsoposterior tubercles enables the female nymphs to be distinguished easily from the males. By contrast, distinguishing between the nymphal stages of the same sex relies on the identification of both the number of metapodosomal setae and cuticular pits. These descriptions are used to produce a key, which allows the various stages of both sexes to be distinguished. PMID- 10872858 TI - Blood-feeding in mosquitoes: probing time and salivary gland anti-haemostatic activities in representatives of three genera (Aedes, Anopheles, Culex). AB - Mosquitoes (Diptera: Culicidae) face their hosts' haemostatic mechanisms when attempting to feed on blood. Accordingly, they antagonize haemostasis by salivary agents that include anti-clotting, anti-platelet and vasodilatory compounds. Because haemostasis is a complex and redundant physiological response that varies between vertebrates, it is to be expected that haematophagous animals have a salivary armoury that most efficiently counteracts their preferred hosts. The mosquito Culex quinquefasciatus Say, which has a strong tendency to ornithophagy, appears to have only recently adapted to mammals and may not have evolved efficient mechanisms to counteract mammalian platelet responses, while birds only have relatively inefficient thrombocytes. Accordingly, we compared the probing behaviour of Cx. quinquefasciatus with two other mosquito species from different backgrounds: Aedes aegypti (L.) and Anopheles albimanus Weidemann, that have apparently had a longer evolutionary association with mammals. Culex takes much more time to find blood on a mammalian host (human or mouse) when compared to the two other mosquito species, but does not differ in probing behaviour when feeding on a chicken. Salivary anti-haemostatic components were also measured in those three species of mosquito and results are discussed in context with the probing behaviour. PMID- 10872859 TI - Cytogenetics of the Anopheles gambiae complex in Sudan, with special reference to An. arabiensis: relationships with East and West African populations. AB - The species composition of malaria vector mosquitoes belonging to the Anopheles gambiae complex (Diptera: Culicidae) from >40 localities in Sudan, representing most ecological situations, was determined by analysis of ovarian polytene chromosomes. Of 2162 females, 93% were identified as An. arabiensis Patton and 7% were An. gambiae Giles sensu stricto. No hybrids were found between the two species. Anopheles arabiensis occurred in all but two sites, whereas An. gambiae s.s. was effectively limited to the southernmost, more humid localities. For chromosomal paracentric inversions, the degree of polymorphism was low in An. gambiae s.s. (inversions 2La, 2Rb and 2Rd), higher in An. arabiensis (inversions Xe, 2Ra, b, bc, d1, s; 3Ra, d). Anopheles gambiae samples from Sudan were all apparently panmictic, i.e. they did not show restricted gene flow such as observed among West African populations (interpreted as incipient speciation). Chromosomal inversion patterns of An. gambiae in southern Sudan showed characteristics of intergrading Savanna/Forest populations similar to those observed in comparable eco-climatic situations of West Africa. Anopheles arabiensis was polymorphic for inversion systems recorded in West Africa (2Ra, 2Rb, 2Rdl, 3Ra) and for a novel 2Rs polymorphism, overlapping with inversion systems 2Rb and 2Rd1. Samples carrying the 2Rs inversion were mostly from Khashm el-Girba area in central-eastern Sudan. In the great majority of the samples all polymorphic inversions were found to be in Hardy-Weinberg equilibrium. Sudan populations of An. arabiensis should therefore be considered as generally panmictic. Anopheles arabiensis shows more inversion polymorphism in west than in east African populations. Sudan populations have more evident similarities with those from westwards than those from eastwards of the Great Rift Valley. The possible influence of the Rift on evolution of An. arabiensis is discussed. PMID- 10872860 TI - Interspecific competition between sibling species larvae of Anopheles arabiensis and An. gambiae. AB - Mosquito larvae of the sibling species Anopheles arabiensis Patton and An. gambiae Giles sensu stricto (Diptera: Culicidae) were investigated for interspecific competition. Single-species and mixed-species populations were reared at 27 degrees C from the first instar to pupation at different densities (100, 200 or 400 larvae/200 cm2 tray) with a constant amount of food, 0.2 mg/larva/day. Pupae obtained from mixed populations were identified to species using PCR. Both species had a 1:1 sex ratio at pupation. Development time to pupation averaged about one day less for An. arabiensis compared to An. gambiae, ranging from 0.93-1.49d for males and from 0.44-0.84 d for females in single populations. In mixed species populations the difference for males ranged from 0.99-1.58d and for females from 0.93-1.62d. Survival rates of An. gambiae s.s. were significantly higher than those of An. arabiensis in both the single-species and mixed-species populations. Mixed-species rearing did not have an effect on the survival of An. gambiae, whereas the mortality rate of An. arabiensis was significantly higher in mixed populations than when only this species was reared at the same densities, suggesting a competitive disadvantage for An. arabiensis in mixed populations. High proportions of larvae (4-35%) were lost during development; these losses could not be accounted for by corpses found in the rearing pans. The possibility of cannibalism was investigated by rearing each species separately in small containers (five per 50 ml), inspected every 6h, but no cannibalism was detected at any stage of development in either species. It was concluded that, under these experimental circumstances, interspecific competition between both species did occur but with a detrimental effect on An. arabiensis only. Relevance of these findings to the ecology of both species in the field is discussed briefly. PMID- 10872861 TI - Anopheles arabiensis and An. funestus are equally important vectors of malaria in Matola coastal suburb of Maputo, southern Mozambique. AB - Transmission characteristics of malaria were studied in Matola, a coastal suburb of Maputo, the capital City, in southern Mozambique, from November 1994 to April 1996. The local climate alternates between cool dry season (May-October) and hot rainy season (November-April) with mean annual rainfall 650-850 mm. Saltmarsh and freshwater pools provide mosquito breeding sites in Matola. Malaria prevalence reached approximately 60% among people living nearest to the main breeding sites of the vectors. Plasmodium falciparum caused 97% of malaria cases, others being P. malariae and P. ovale. Potential malaria vector mosquitoes (Diptera: Culicidae) collected at Matola during daytime indoor-resting (n = 1021) and on human bait at night (n = 5893) comprised 12% Anopheles coustani Laveran (93% biting outdoors), 46% An. funestus Giles (68% biting indoors) and 42% An. gambiae Giles sensu lato (60% biting outdoors). All 215 specimens of An. gambiae s.l. identified genetically were An. arabiensis Patton. Anopheles funestus populations remained stable throughout the year, whereas densities of the An. gambiae complex fluctuated considerably, with An. arabiensis peaking during the rainy season. No concomitant rise in malaria incidence was observed. Human landing indices of An. funestus and An. arabiensis averaged 1.8 and 3.8 per man-night, respectively. Overall Plasmodium sporozoite rates were 2.42+/-1.24% in 2181 An. funestus and 1.11+/-1.25% in 1689 An. arabiensis dissected and examined microscopically. Mean daily survival rates were 0.79 for both vector species. Estimated infective bites/person/year were 15 An. funestus and 12 An. arabiensis. Biting rates were greatest at 2100-24.00 hours for An. funestus (68% endophagic) and 21.00-03.00 hours for An. arabiensis (40% endophagic). The entomological inoculation rate (EIR) declined sharply over very short distances (50% per 90m) away from breeding sites of the vectors. Consequently, P. falciparum prevalence among Matola residents was halved 350 m within the town. Implications for the protective effectiveness of a 'cordon sanitaire' by residual house-spraying and/or the use of insecticide-treated bednets are discussed. PMID- 10872862 TI - Anopheles funestus resistant to pyrethroid insecticides in South Africa. AB - Northern Kwazulu/Natal (KZN) Province of South Africa borders on southern Mozambique, between Swaziland and the Indian Ocean. To control malaria vectors in KZN, houses were sprayed annually with residual DDT 2 g/ m2 until 1996 when the treatment changed to deltamethrin 20-25 mg/m2. At Ndumu (27 degrees 02'S, 32 degrees 19'E) the recorded malaria incidence increased more than six-fold between 1995 and 1999. Entomological surveys during late 1999 found mosquitoes of the Anopheles funestus group (Diptera: Culicidae) resting in sprayed houses in some sectors of Ndumu area. This very endophilic-vector of malaria had been eliminated from South Africa by DDT spraying in the 1950s, leaving the less endophilic An. arabiensis Patton as the only vector of known importance in KZN. Deltamethrin sprayed houses at Ndumu were checked for insecticide efficacy by bioassay using susceptible An. arabiensis (laboratory-reared) that demonstrated 100% mortality. Members of the An. funestus group from Ndumu houses (29 males, 116 females) were identified by the rDNA PCR method and four species were found: 74 An. funestus Giles sensu stricto, 34 An. parensis Gillies, seven An. rivulorum Leeson and one An. leesoni Evans. Among An. funestus s.s. females, 5.4% (4/74) were positive for Plasmodium falciparum by ELISA and PCR tests. To test for pyrethroid resistance, mosquito adults were exposed to permethrin discriminating dosage and mortality scored 24h post-exposure: survival rates of wild-caught healthy males were 5/10 An. funestus, 1/9 An. rivulorum and 0/2 An. parensis; survival rates of laboratory-reared adult progeny from 19 An. funestus females averaged 14% (after 1h exposure to 1% permethrin 25:75cis:trans on papers in WHO test kits) and 27% (after 30 min in a bottle with 25 microg permethrin 40:60cis:trans). Anopheles funestus families showing >20% survival in these two resistance test procedures numbered 5/19 and 12/19, respectively. Progeny from 15 of the families were tested on 4% DDT impregnated papers and gave 100% mortality. Finding these proportions of pyrethroid-resistant An. funestus, associated with a malaria upsurge at Ndumu, has serious implications for malaria vector control operations in southern Africa. PMID- 10872863 TI - Resistance to dieldrin + fipronil assorts with chromosome inversion 2La in the malaria vector Anopheles gambiae. AB - Cyclodiene insecticide resistance in malaria vector mosquitoes of the Anopheles gambiae species complex (Diptera: Culicidae) has been reported previously from several parts of Africa. We report resistance to dieldrin, a cyclodiene, in two laboratory strains of An. gambiae Giles sensu stricto code-named Ian P20 from Nigeria (1979) and CIG from Cote d'Ivoire (1997). Dieldrin resistance levels were high in adult female mosquitoes (40-75% survived exposure to 4% dieldrin for 1 h) and was closely linked with chromosomal paracentric inversion 2La. This inversion did not occur in Hardy-Weinberg proportions, but showed an excess of heterozygotes in both strains, which may account for the high levels of resistance. This linkage also suggests that dieldrin resistance in Ian P20 is dominant. After subsamples of strain Ian P20 were exposed for 1 h to dieldrin 4% or fipronil 2% (discriminating concentrations), the resultant mortality-rates (61% and 65%) were not significantly different. Most survivors after fipronil treatment also survived subsequent exposure to dieldrin (46/50=92%). This apparent cross-resistance between insecticides of two classes (cyclodiene and phenyl pyrazole) has implications for the management of insecticide resistance in wild populations of the An. gambiae complex. PMID- 10872864 TI - Human sweat and 2-oxopentanoic acid elicit a landing response from Anopheles gambiae. AB - A wind tunnel bioassay and video to observe mosquitoes landing on heated glass cylinders were used to test sweat and some derivatives for responses of Anopheles gambiae Giles (Diptera: Culicidae), a highly anthropophilic African species of malaria vector. Filter papers impregnated with human sweat and a diethyl ether extract from the filter papers elicited significantly more landings than a water control (P<0.001). The concentration of lactic acid in the extract was determined by GLC assay, but bioassays of an equivalent dose of lactic acid (from a commercial supplier) did not elicit landings. Chemical analysis of the extract by combined GLC/mass spectrometry indicated the presence of 73 compounds, of which 40 were tentatively identified. The major components of the extract were aliphatic carboxylic acids. An artificial blend of 22 carboxylic acids did not elicit landings. Bioassays of 2-oxopentanoic acid elicited significantly more, landings (P<0.001). The possible importance of oxo-carboxylic acids for host seeking by anthropophilic mosquitoes is discussed and their use for trapping is suggested. PMID- 10872865 TI - Survival, ovarian development and bloodmeal size for the horn fly Haematobia irritans irritans reared in vitro. AB - Horn flies, Haematobia irritans irritans (Linneaus) (Diptera: Muscidae) were reared in vitro using cattle, pig, horse, rabbit, sheep, goat or chicken blood. The highest survival, bloodmeal size and rate of ovarian development were recorded for both female and male flies fed cattle blood. Flies fed pig, rabbit, sheep and goat blood showed intermediate survival. Flies fed chicken blood showed the lowest survival rates, ingested the smallest bloodmeals and did not develop ovaries. The relationship between dietary factors and host specificity of the horn fly, and the efficiency of vertebrate blood source of several animals for laboratory colonization of horn fly are discussed. PMID- 10872866 TI - Courtship and mating by the sandfly Phlebotomus duboscqi, a vector of zoonotic cutaneous leishmaniasis in the Afrotropical region. AB - Courtship behaviour of males of the Afrotropical sandfly Phlebotomus duboscqi Neveu-Lemaire (Diptera: Psychodidae) involved mounting the female and clasping her 'waist' with the male coxites placed between the female's thorax and abdomen. This behaviour, which we call 'piggy-backing', was preceded by male wing beating, perhaps involving mate recognition and contact pheromones. It did not seem to be pre- or postcopulatory mate guarding. Piggy-backing was attempted by P. duboscqi males on females of other species (P. papatasi and P. perniciosus) and even on other male P. duboscqi. The majority of female P. duboscqi piggy-backed by males were already inseminated, and most of the courting did not lead to copulation. This, coupled with the presence of a mating plug (semen) in each spermatheca of inseminated females, suggests that female P. duboscqi are monogamous for at least the first gonotrophic cycle. Male courtship with piggy-backing was more intense when females could feed on a hamster than when a hamster was present but the females were denied access to the host. It is suggested that, when a hamster was available to the females, the conditions in the laboratory are similar to those in rodent holes, the natural habitat of P. duboscqi. PMID- 10872867 TI - Permethrin resistance ratios compared by two methods of testing nymphs of the German cockroach, Blattella germanica. AB - For the German cockroach, Blattella germanica L. (Dictyoptera: Blattellidae), the permethrin resistance ratio (RR) was assessed by topical application and by tarsal contact tests, using first-instar nymphs of five strains from Tehran, Iran. Each test was replicated three or four times with 10 nymphs aged 2-3 days; mortality was scored 24h post-treatment. The reference susceptible strain showed LD50 permethrin 0.0175 microl/nymph from topical application, KT50 of 8.41 min and LT50 of 12.82 following tarsal contact with permethrin 15 mg/m2. In four wild strains (F1 generation) the RR varied from 4.14 to 4.7 for mortality after topical application, from 4.2 to 6.45 for mortality and 17-27 for knockdown following tarsal contact tests. Hence, overall knockdown results gave much higher RRs than for mortality data. Resistance ratios based on both methods of treatment were very similar: one strain showed a slightly higher value by topical application (RR 4.6 vs. 4.2, i.e. 1.1-fold difference) whereas the other three strains gave slightly greater RR (1.2-1.4 fold) by tarsal contact. Resistance was abolished by cotreatment with the synergist piperonyl butoxide plus permethrin (ratio 3:1 required for full efficacy), indicating that mixed-function oxidases were inhibited as a major metabolic pathway in all four resistant strains. PMID- 10872868 TI - DNA in situ hybridization on polytene chromosomes of Simulium sanctipauli at loci relevant to insecticide resistance. AB - A DNA technique for in situ hybridization developed by Kumar & Collins (1994) for use on polytene chromosomes of adult Anopheles mosquitoes (Diptera: Culicidae) was modified for use with Simulium larval salivary gland chromosomes (Diptera: Simuliidae). Cloned fragments of several Simulium genes (coding for aspartate amino transferase, cytochrome P450 and DNA polymerase) were successfully mapped physically by assigning specific band locations in Simulim sanctipauli V. & D. This represents the first attempt at locating genes beyond the resolution of linkage to inversions in any blackfly species. PMID- 10872869 TI - Further evidence that deltamethrin-impregnated collars protect domestic dogs from sandfly bites. AB - In many foci of zoonotic visceral leishmaniasis (ZVL), domestic dogs are important reservoir hosts of the causative Leishmania parasites transmitted by phlebotomine sandflies (Diptera: Psychodidae). We tested the protective value of impregnated dog collars (20 g plastic containing deltamethrin 800 mg ai) against Phlebotomus papatasi (Scopoli) sandflies in Iran. For each assay, the dog was sedated and caged in a net with 70-100 wild-caught sandflies overnight (23.30 06.30 hours). Dogs wearing the collars were bitten by approximately 80% fewer sandflies than before collars were fitted, i.e. 51% vs. 11% of hungry female flies exposed. Sandfly mortality rates following 20 h exposure to dogs with collars (18%) or without collars (17%) were not significantly different. Effects of collars were tested when dogs had been wearing them for 8 days. A previous trial against the sandfly P. perniciosus Newstead in France, using smaller dogs, showed that effects of such collars were not fully realized until they had been worn for 2 weeks or more; they remained effective for at least 8 months and killed significant proportions of the sandflies exposed. Present results with P. papatasi, confirming that this simple device provides effective protection against sandflies, are considered sufficiently encouraging to justify a community wide field trial of deltamethrin-impregnated dog collars against ZVL vector sandflies in Iran. PMID- 10872870 TI - New cases of Mokola virus infection in South Africa: a genotypic comparison of Southern African virus isolates. AB - Mokola virus, one of the six genotypes within the Lyssavirus genus of the Rhabdoviridae family, is believed to be exclusive to the African continent, where infections in various mammal species have been reported. After an isolation of Mokola virus at Umhlanga on the east coast of South Africa in 1970, the virus was not reported in South Africa until its reappearance in 1995. Since then a total of six new isolates of the virus were made, three from the East London region in 1995 and 1996, two near Pinetown in 1997 and a further isolate in a residential suburb of the city of Pietermaritzburg, in 1998. These isolation sites are respectively about 500 km (East London region) and 23 to 60 km from the site of the 1970 isolation Phylogenetically the three isolates from the East London area were similar and could be distinguished from the four KwaZulu-Natal isolates, which formed a defined group of their own. The viruses comprising these two clusters were also found to be distant from another southern African isolate, made in 1982 in Zimbabwe, Mokola virus isolates thus conforms to a pattern of virus evolution strongly influenced by geographical determinants. In comparison to Rabies virus, of which at least two different biotypes are known and a vast array of different wildlife species contribute to its complex epidemiology on the sub-continent, Mokola viruses have only been isolated form one species, i.e. domestic cats, in South Africa. Nevertheless, the heterogeneity among the Mokola virus isolates is far greater than the degree of variation among the Rabies virus populations of the region. PMID- 10872871 TI - Nucleotide sequence of the gene encoding the major capsid protein of herpesvirus of turkeys. AB - The gene encoding the major capsid protein (MCP) VP5 of herpesvirus of turkeys (HVT) was identified and sequenced. It has a single open reading frame of 4236 nucleotides encoding 1412 aa protein. The gene is flanked by VP23 and UL20 sequences and is localized in the unique long region (UL) within the BamHI-B fragment. Comparison of amino acid homology has shown its clear position among the alpha-herpesviruses rather than beta- or gamma-herpesviruses. The VP5 is expressed from polycistronic mRNA together with the UL20 and the VP23 genes. The 7,2 kb RNA transcript is lacking any promoter elements or polyA signal in intergenomic regions between VP5 and UL20 or VP5 and VP23 genes, respectively. Multiple alignment of known major capsid protein sequences of all herpesvirus groups revealed presence of seven highly homologous clusters suggesting-that the corresponding protein domains might play an important role in folding of MCP and assembly of herpesvirus capsid. PMID- 10872872 TI - Differential effect of TPA on cell growth and Epstein-Barr virus reactivation in epithelial cell lines derived from gastric tissues and B cell line Raji. AB - We characterized the cell growth and Epstein-Barr virus (EBV) reactivation for EBV infected epithelial cell lines, GT38, GT39, and GTC-4 using 12-O tetradecanoylphorbol-13-acetate (TPA). These cell lines grew similarly in liquid medium, and formed colonies in soft agar. The cell growth was inhibited with TPA, dose-dependently in liquid medium. The colony formation was enhanced with low concentrations of TPA, but was inhibited with high concentrations. The latent EBV was reactivated with high concentrations of TPA as shown by the expression of EBV BZLF1 gene product ZEBRA. The effects of TPA on GTC-4 were compared with a Burkitt's lymphoma cell line Raji. The mode of actions of TPA in GTC-4 was different from Raji in terms of cell growth and EBV reactivation. The effective concentrations of TPA for cell growth inhibition and EBV reactivation were higher in Raji than GTC-4. Cell cycle analysis showed that TPA (20 ng/ml) induced cell cycle arrest to Raji but not to GTC-4; however, the rate of trypan blue stained cells increased in the TPA treated GTC-4 but not Raji. These results demonstrated that TPA affects differentially for the stimulation and inhibition of cell growth, and also EBV reactivation depends on TPA concentrations and cell types. PMID- 10872873 TI - Cloning and secreted expression of the extracellular domain of the mumps virus fusion protein in Pichia pastoris. AB - The extracellular globular domain of the mumps virus fusion (F) protein (amino acids 28-481) has been overexpressed from GS115 his4 Pichia pastoris cells following the generation of a recombinant clone. The heterologous protein was directed for secreted expression by in-frame cloning with the S. cerevisiae alpha factor secretion signal. The expressed protein was observed to secrete into the culture medium. An expressing clone was obtained initially by small-scale induction, metabolic labeling and immunoprecipitation. Expression analysis of the chosen clone was confirmed by western blotting with F protein specific polyclonal serum. The effects of culture volume, temperature and methanol concentration on the levels of expression, were studied. The results indicate that there is a balance required between the induction temperature and methanol concentration to achieve maximal expression. In addition, the presence of designated monomeric (47 K), dimeric (85-90 K) and trimeric (140 K) forms are dependent upon the induction conditions. Estimated secreted protein expression levels of > 1 mg/L were obtained in these studies. Further, the experiments demonstrate that the complete reconstruction of the KEX2 protease cleavage site is not necessary to facilitate secretion. PMID- 10872874 TI - The ORF RTL1 transcript of fowl adenovirus type-8 is spliced and truncated at late stages of the virus replication cycle. AB - Two transcription products were found for the open reading frame (ORF) RTL1 located near the right terminus of the fowl adenovirus type-8 genome. The larger transcript, which was transcribed mostly during the early stage of the virus infection, contains the complete sequence (933 nucleotides) of the predicted ORF from the genomic DNA sequence encoding a 311 amino acid (aa) polypeptide. In contrast, the shorter transcript, which was more predominant at the late stage of the infection, was missing 580 nucleotides (from nucleotide 117 to 696). A premature stop codon was introduced at 210 nucleotides downstream from the start codon and the shorter transcript would encode a 70 aa polypeptide. This observation indicates that the ORF RTL1 may produce two different proteins, which function differently at different stages of the virus infection. Another possibility is that the virus may use alternative splicing as a mechanism to control the expression of the ORF, since the spliced transcript was prematurely terminated at the late stage of the infection. PMID- 10872875 TI - An outbreak of African Swine Fever in Nigeria: virus isolation and molecular characterization of the VP72 gene of a first isolate from West Africa. AB - The isolation of 98/ASF/NG, a strain of African Swine Fever Virus (ASFV) associated with a 1998 epizootic in Nigeria, is reported. This first isolate of the virus from West Africa was identified through a successful polymerase chain reaction (PCR) amplification and sequencing of a 280 base pair (bp) fragment of the Major Capsid Protein (VP72) gene. Further amplification and sequence analysis of a 1.9 kilobase pair (kbp) fragment encompassing the complete VP72 gene showed that the isolate has a 92.2%, 92.4%, and 97.2% homology with previously sequenced Ugandan, Dominican Republican and Spanish isolates respectively. Of the 50 nucleotide changes observed in this highly conserved gene, 45 were found to result in 40 amino acid changes clustered around the central region (position 426 to 516) of the VP 72 protein while changes at the remaining 5 positions were silent. These changes also led to the loss of two out of the seven potential N glycosylation sites which are in this gene conserved among all isolates. The possible epizootiological implications of such mutations in a highly conserved gene of a DNA virus is discussed in relation to this outbreak. PMID- 10872876 TI - Induction of apoptosis by equine arteritis virus infection. AB - Equine arteritis virus (EAV) is the etiological agent of equine viral arteritis, a contagious viral disease of equids. EAV is the prototype virus of the arteriviruses, a group of small enveloped viruses with positive single-stranded RNA genomes. Because apoptosis or programmed cell death is believed to play an important role in the biogenesis of several cytopathogenic viruses, we examined whether EAV was able to induce cell apoptosis in vitro. To do this, Vero cells were infected with EAV at a multiplicity of infection of 0.1 tissue culture infectious dose (TCID50) per cell, and analyzed at various time intervals for the appearance of apoptotic signs. Fragmentation of chromosomal DNA into nucleosomal oligomers and caspase activation were observed in the infected cells at the time (e.g. 24h postinfection) where a noticeable cytopathic effect was observed. The kinetics of the DNA fragmentation correlated with that of the production of progeny virus, so that viral multiplication was not interrupted by the apoptotic cell damage. All these data provide evidence that EAV is able to induce apoptotic cell death in vitro. PMID- 10872877 TI - Cloning, sequencing, expression and promoter analysis of a structural protein of bacteriophage MB78. AB - Bacteriophage MB78, a virulent phage of Salmonella typhimurium isolated in our laboratory. It is different from the well-known temperate phage P22 and 9NA. A detailed physical map has been constructed. To understand more about the physiology and genetics of this interesting phage it has become necessary to fragment the phage genome, clone the fragments and analyze in depth. A number of promoters of bacteriophage MB78 have been cloned and characterized recently. As a part of this program, in this investigation, we report cloning, sequencing and expression and promoter analysis of the ClaI G fragment. We identified the expressed protein as phage structural. Phage structural proteins play a vital role in forming the core head of the phage particle. PMID- 10872878 TI - Screening predicted coding regions in poxvirus genomes. AB - The amino acid composition and pI values were calculated for the predicted proteins of a series of complete poxvirus genomes. Many of the vaccinia virus (strain Copenhagen) minor ORFs, thought not to be functional genes, were found to have significantly more or less of several amino acids than a set of the largest 150 vaccinia virus proteins. Very high isoelectric point (pI) values were also correlated with a group of the minor ORFs. Analysis of molluscum contagiosum virus ORFs by amino acid composition and pI identified a number of ORFs previously denoted as doubtful and highlighted several others that could be similarly classified. The use of amino acid composition and pI appears to be a generally applicable tool to aid identification of viral ORFs that are unlikely to be functional genes. PMID- 10872879 TI - Acquisition of multiple virulence/avirulence determinants by potato virus X (PVX) has occurred through convergent evolution rather than through recombination. AB - Resistance to potato virus X (PVX) is determined by the product of a host resistance gene and a viral determinant specifying either virulence (resistance breaking ability) or avirulence (resistance sensitivity). The viral coat protein is the determinant of resistance mediated by the host Nx gene while the 25 kDa movement protein is the determinant of Nb-mediated resistance. Group 1 and group 4 strains of PVX are avirulent or virulent respectively for both these determinants while group 2 and group 3 strains are virulent for one but avirulent for the other determinant. There are two alternative evolutionary mechanisms by which the various strain groups might have evolved: either by recombination between strains carrying virulence (or avirulence) determinants that evolved once only, or alternatively, by independent evolution of at least one virulence (or avirulence) determinant in distinct phylogenetic branches. These alternative hypotheses were investigated by (i) determining the complete genomic sequence of a group 1 and a group 4 strain and (ii) comparing the completely sequenced genomes of six isolates representative of the four strain groups. The analysis revealed the same phylogeny for all five PVX genes. Thus, there is no evidence that the PVX strain groups evolved by recombination. PMID- 10872880 TI - Sequence determination and molecular analysis of two strains of bovine parainfluenza virus type 3 that are attenuated for primates. AB - The Kansas/15626/84 (Ka) and Shipping Fever (SF) strains of bovine parainfluenza virus type 3 (BPIV3) replicate less efficiently than human PIV3 (HPIV3) in the upper and lower respiratory tract of rhesus monkeys, and BPIV3 Ka is also highly attenuated in humans and is in clinical trials as a candidate vaccine against HPIV3. To initiate an investigation of the genetic basis of the observed attenuation phenotype of BPIV3 in primates, the complete genomic sequences of Ka and SF genomes were determined and compared to those of BPIV3 strain 910N and two HPIV3 strains, JS and Wash/47885/57. There is a high degree of identity between the five PIV3 viruses in their 55 nucleotide (nt) leader (83.6%) and 44 nt trailer (93.2%) sequences. The five viruses display amino acid sequence identity ranging from 58.6% for the phosphoprotein to 89.7% for the matrix protein. Interestingly, the majority of amino acid residues found to be variable at a given position in a five-way protein alignment are nonetheless identical within the viruses of either host species (BPIV3 or HPIV3). These host-specific residues might be products of distinct selective pressures on BPIV3 and HPIV3 during evolution in their respective hosts. These host-specific sequences likely include ones which are responsible for the host range differences, such as the efficient growth of BPIV3 in bovines compared to its restricted growth in primates. It should now be possible using the techniques of reverse genetics to import sequences from BPIV3 into HPIV3 and identify those nt or protein sequences which attenuate HPIV3 for primates. This information should be useful in understanding virus-host interactions and in the development of vaccines to protect against HPIV3-induced disease. PMID- 10872881 TI - Full-genome nucleotide sequence of a hepatitis C virus variant (isolate name VAT96) representing a new subtype within the genotype 2 (arbitrarily 2k). AB - Hepatitis C virus (HCV), a single-stranded RNA virus of the family Flaviviridae, has a wide range of genetic heterogeneity: 6-11 genotypes (or 6 clades) have been known and each genotype comprises multiple subtypes. Here we report the entire nucleotide sequence of an HCV isolate from a patient in Moldova with chronic hepatitis (isolate name VAT96). The genetic organization of VAT96 was, from 5' to 3' ends, 5'UTR (341 nt), polyprotein ORF (9099 nt), 3'UTR (38 nt except for the poly-U and poly-pyrimidine stretch), and X-tail (98 nt). Comparison of the polyprotein amino acid sequence of VAT96 with those of known full-genome isolates assigned VAT96 to the genotype 2 (or clade 2), and further phylogenetic analysis based on a 447-nt sequence that covers part of the C and El regions suggested that VAT96 represents a new subtype within the genotype 2, arbitrarily designated "2k" VAT96 was unique in that it possessed a U residue prior to GCC at the 5' end of its genome while all the other full-genome HCV sequences start with GCC or ACC. In addition, the polyprotein ORF of HCV-VAT96, like HCV-BEBE1 of 2c, encoded several additional amino acids in excess, compared to 2a and 2b sequences. Despite these characteristics that may be unique to VAT96, the 98-nt sequence of the X-tail of VAT96 was highly homologous to those of other isolates with different genotypes so far reported. PMID- 10872882 TI - Identification and characterization of the Trichoplusia ni single capsid nuclear polyhedrosis virus p10 gene. AB - The p10 gene was identified and characterized from the Trichoplusia ni single capsid nuclear polyhedrosis virus (TniSNPV). The p10 open reading frame (ORF) sequence was identified following sequencing of the ends of the EcoRI-G clone. Subsequent sequencing of an EcoRI-SmaI subclone identified the entire p10 and a portion of a p26 homologue. The p10 ORF of 264 basepairs (bps), encoded a predicted protein of 88 amino acids (aas) with Mr 9527 Da. The putative late transcription initiation motif (TAAG) was found upstream of the translation initiation codon at position -46. Downstream of the translation stop codon, a putative poly(A) signal was identified. The p10 amino acid sequence contained the three conserved domains reported for all other p10 genes. The p10 amino acid sequence was most homologous (85% similarity and 67% identity) to that of Buzura suppresaria NPV p10 sequence. PMID- 10872883 TI - Completion of the four large gene sequences of porcine group C Cowden rotavirus. AB - The terminal nucleotide sequences of group C Cowden rotavirus gene segments 1-4 were determined. When compared with the published sequences, we found 14 to 29 additional nt at the 5' ends of the four reported gene sequences. For the 3' ends, we observed an additional 16 nt in gene 2 and 14 fewer nt in gene 4. PMID- 10872884 TI - Temporal relationship between immune cell influx and the expression of inducible nitric oxide synthase, interleukin-4 and interferon-gamma in pancreatic islets of NOD mice following adoptive transfer of diabetic spleen cells. AB - Beta cell destruction in NOD mice can be accelerated by adoptive transfer of diabetic spleen cells into irradiated adult NOD mice. Here mice receiving diabetic spleen cells were examined at days 0, 7, 14, 21 and at onset of diabetes for the resulting insulitis and the number of intra-islet CD4 and CD8 cells and macrophages. The progression of insulitis and the number of intra-islet CD4 and CD8 cells and macrophages were correlated with the expression and co-localization of inducible nitric oxide synthase, interferon-gamma and interleukin-4 by dual label light and confocal immunofluorescence microscopy. Diabetes developed in 7/8 mice by 27 days following cell transfer. The insulitis score increased slightly by day 7 but rose sharply at day 14 (p = 0.001) and was maintained until diabetes. The mean number of intra-islet CD4 and CD8 cells and macrophages showed a similar trend to the insulitis scores and were present in almost equal numbers within the islets. Immunolabelling for inducible nitric oxide synthase was observed at day 7 in only some cells of a few islets but increased sharply from day 14. It was restricted to islets with insulitis and was co-localized in selective macrophages. Weak intra-islet interleukin-4 labelling was observed at days 7 and 14 but became more pronounced at day 21 and at onset of diabetes, being present in selective CD4 cells. Intra-islet labelling for interferon-gamma was first observed at day 21, but became more intense at onset of diabetes and was co-localized in a proportion of macrophages. Both cytokines were expressed in islets with advanced insulitis. Interferon-gamma staining was also observed within endothelial cells located in the exocrine pancreas. We conclude that transfer of diabetic spleen cells results in a rapid influx of CD4 and CD8 cells and macrophages within the pancreas of recipient mice. During the period of heightened insulitis, selective immune cells begin to express inducible nitric oxide synthase and the opposing cytokines, interferon-gamma and interleukin-4. Expression of these molecules becomes more pronounced immediately prior to and during the onset of diabetes. PMID- 10872885 TI - Glycosylation profiles of airway epithelium after repair of mechanical injury in guinea pigs. AB - Glycosylated structures on the cell surface have a role in cell adhesion, migration, and proliferation. Repair of the airway epithelium after injury requires each of these processes, but the expression of cell surface glycosylation of airway epithelial cells after injury is not known. We examined cell surface glycosylation using lectin-binding profiles of normal and repairing epithelia in Hartley guinea pigs from 0 to 14 days after mechanical injury. The epithelium regenerated completely-over 7 days. In normal trachea, galactose- or galactosamine-specific lectins (14 of 20 tested) labelled epithelial cells, but fucose, mannose, and other sugar-specific lectins (15 tested) did not. GSA-2, a glucosamine-specific lectin, labelled epithelial cells weakly in uninjured tracheas, but intense labelling was noted in basal and non-ciliated columnar cells adjacent to the injury site over 3 h to 14 days after injury. Labelling of these cells peaked at 12 h and 5 days after injury respectively. Similar patterns were seen with lectins AlloA and HAA but not with CPA during repair. The binding of the lectin DSA to proteins collected from primary cultures of airway epithelial cells decreased substantially after treatment for 24 h with either transforming growth factor-beta or interleukin-1beta, but that of the CPA lectin did not. We demonstrate changes in glycosylation profiles of airway epithelial cells coordinate with repair after mechanical injury. These changes may be useful to study mechanisms by which repair is regulated. PMID- 10872886 TI - Detection of different mRnas expressed in the thyro-parathyroid complex of the rat by in situ hybridization using digoxigenin-labelled oligonucleotide probes. AB - The effects have been examined of different methods and regimens for tissue fixation, preservation, permeabilization and immunostaining of different mRNAs detected by in situ hybridization in paraffin-embedded samples. The three main hormone mRNAs expressed in the thyro-parathyroid glands, namely thyroglobulin, calcitonin and parathyroid hormone mRNAs, were chosen as the target nucleic acid sequences to be detected using digoxigenin-labelled probes. Our results suggest that chemical fixation and permeabilization of tissue samples are restrictive steps. Thus, paraformaldehyde fixation provides excellent signal intensities and non-detectable background levels whereas routine formalin and Bouin's solution give unsatisfactory results. A clear linear correlation was also found between signal intensity and proteinase K permeabilization. Moreover, the optimization of immunohistochemical steps, such as anti-digoxigenin antibody concentration and colour development times, enhance the intensity and specificity of hybrid signals. Furthermore, our results show that, in contrast to some data in the literature, paraffin-embedded tissue is suitable for detection of mRNAs by in situ hybridization. It gives equivalent intensities of specific signal and superior histological and cellular resolutions when compared to cryopreserved tissue. PMID- 10872887 TI - Ghost cells in calcifying odontogenic cyst express enamel-related proteins. AB - The so-called ghost cell is a unique cell type occurring in a variety of odontogenic and non-odontogenic lesions. However, the true nature of ghost cells has not been determined. In the present study, we examined the immunoreactivity of ghost cells in calcifying odontogenic cysts and dermal calcifying epitheliomas, with antibodies against amelogenin, enamelin, sheath protein (sheathlin) and enamelysin, in an attempt to clarify the nature of this unique cell. The cytoplasm of ghost cells in calcifying odontogenic cysts demonstrated distinct immunolocalization of the enamel-related proteins, while similar in the calcifying epitheliomas of the skin showed a negative reaction. The results indicate that the ghost cells in calcifying odontogenic cysts, as opposed to ghost cells in dermal calcifying epitheliomas, contain enamel-related proteins in their cytoplasm accumulated during the process of pathological transformation. PMID- 10872888 TI - Ultracytochemical detection of guanylate cyclase C activity in alimentary tract and associated glands of the rat. Influence of pH, ATP and the ions Mg2+ and Mn2+. AB - Intestinal guanylate cyclase C is activated by guanylin, an endogenous peptide. This activity seems to be modulated by adenine nucleotides, the ions Mg2+ and Mn2+, and pH. In this study, we report an ultracytochemical method for the localization of guanylate cyclase C activity at the electron microscope level. We studied the enzymatic activity in the presence or absence of guanylin and/or ATP, in the presence of the ions Mg2+ or Mn2+, and at different pH levels. The greatest distribution of enzymatic activity was detected in samples incubated at pH 8 and 7.4 in the presence of guanylin, Mg2+ and ATP. Guanylate cyclase C activity was detected at the surface epithelium of stomach and intestine, and in liver, exocrine pancreas and parotid gland. In the intestine, enzymatic activity was more widely distributed in the duodenum than in the jejunum-ileum and colon. In the small intestine, activity was more evident in the upper portion than in the basal portion of the villus. In samples incubated at pH 8 and 7.4 in the absence of ATP, enzymatic activity was detected only in small intestine, liver and exocrine pancreas. Enzymatic activity was present in duodenum incubated at pH 8 and 7.4 in the presence of Mn2+ and in the presence or absence of ATP. No samples incubated in all these experimental conditions but at pH 5 or samples incubated in the presence of guanylin only or in the absence of guanylin, displayed guanylate cyclase C activity. Our results suggest that a complete ultracytochemical detection of guanylate cyclase C activity requires guanylin as stimulator, and incubation in the presence of Mg2+ and ATP at pH 8 and 7.4. PMID- 10872889 TI - The presence of pericytes and transitional cells in the vasculature of the human dental pulp: an ultrastructural study. AB - The aim of this study was to determine the ultrastructural characteristics of the microvasculature of healthy human dental pulp, with particular reference to pericytes. Pulp tissue was taken from healthy impacted third molars following extraction. Eight teeth were obtained from 17- to 25-year-old patients and pulp tissue was processed for examination using standard techniques for transmission electron microscopy. The pulp was rich in capillaries composed of endothelial and peri-endothelial cells in a 4: 1 ratio. Endothelial cells contained typical and abundant Weibel-Palade bodies. Three types of peri-endothelial cells were identified: pericytes, transitional cells and fibroblasts. Pericytes were embedded within the capillary basement membrane. Transitional cells were partly surrounded by basement membrane, but separated from the endothelium by collagen fibrils; fibroblasts were outside, but adjacent to the basement membrane and closely associated with collagen fibrils. Pericytes and transitional cells, but not peri-endothelial fibroblasts, contained low numbers of dense bodies similar to the endothelial Weibel-Palade bodies. Our observations are consistent with the hypothesis that, during normal tissue turnover, some pericytes may originate from endothelium and migrate away from the vessel wall to undergo transition to a fibroblastic phenotype. PMID- 10872890 TI - Transforming growth factor-beta isoform expression in mature human healthy and carious molar teeth. AB - Transforming growth factor (TGF)-beta isoforms have been implicated in cellular signalling during tooth development and repair, but little is known of their cellular localisation or distribution within the dental tissues in the mature tooth. This study investigated the presence of TGF-beta1, beta2 and beta3 isoforms in tissues of sound and carious human molar teeth, to understand better the expression of TGF-betas during health and disease. In healthy tissues, odontoblasts, cells of the cell rich layer, pulpal fibroblasts and endothelial cells were stained to varying degrees for all isoforms, with TGF-beta3 showing the greatest intensity and TGF-beta1 the weakest intensity. Similar patterns of staining were observed in carious teeth; however, TGF-beta1 showed significantly increased staining intensity within odontoblasts and pulpal cells of carious teeth (p < 0.001). Biochemical analysis showed greater amounts of TGF-beta1 in tertiary dentine than in primary dentine samples. The expression of TGF-betas in odontoblasts and the increased presence of TGF-beta1 in tertiary dentine suggest that these isoforms may be important in odontoblast behaviour and the modulation of the tissue response to injury. PMID- 10872891 TI - Cryopreservation and image enhancement of juvenile and adult dentine mineral. AB - The inorganic component of bone and related hard tissues is generally described as sheets of uniform needle- and plate-like crystals. However, cryofixation has become the method of choice for ultrastructural studies of bone mineral when ladder-like arrangements of filaments contained within deformable microspheres about 1 microm in diameter are apparently the prime structural feature and are consistent with the optical image. The same methodology has now been applied to mature human dentine in caries-free juvenile and adult teeth. These were fixed, sliced, stained for mineral and examined optically or were snap frozen, fragmented under liquid nitrogen, freeze-substituted with methanol or acetone and embedded without thawing in Lowicryl K4M for electron microscopy. Others were processed by traditional transmission electron microscopy methods. To obtain maximum resolution, the electron micrographs were photographically printed as negatives and image-enhanced by digitisation using a Polaroid Sprint Scan 45 and laser printer. In both optical and cryopreparations of juvenile and adult dentine, mineral microspheres up to 1 microm in diameter, were present in the dentinal tubules and peritubular dentine. Within these objects, the mineral was primarily in the form of sinuous electron dense filaments, 5 nm thick, which had a characteristic periodicity. In these preparations needle-like and plate-like structures were rare. In contrast, after traditional transmission electron microscopy preparation although similar filamentous structures remained, the mineral more generally had the familiar form of needles measuring approximately 50 nm in the long axis. The cryopreserved calcified filaments were apparently particularly densely distributed in the intertubular dentine where their parallel ladder-like arrays often formed highly orientated struts and stays. It was concluded that early dentine mineral has the form of filamentous microspheres and as in bone (and other calcifying tissues and cells) has no specific association with collagen. It was also concluded that these structures compact and deform with maturity into a sub-structural framework which may relate to powerful biomechanical forces transmitted through the tissue. Needle- or plate-like mineral is probably rare in vivo in dentine, only becoming commonplace after extensive chemical processing. PMID- 10872892 TI - Immunoreceptor tyrosine-based inhibitory motifs on activating molecules. AB - Immunoreceptor tyrosine-based inhibitory motifs (ITIMs) have the restricted consensus sequence V/I/xYxxL/V, but may be more broadly defined by the sequence V/I/L/SxYxxL/V/I/S. If one includes the ITIM of CTLA-4, then the sequence becomes psixYxxpsi, where psi represents amino acids with nonpolar side chains. Aside from their presence in various inhibitory molecules, ITIMs are also found on many activating receptors and pathways. ITIMs with the restricted consensus sequence occur on IL-4Ralpha, IL-3Rbeta type II, gp130 cytokineR, OB-R (leptinR), LIF Rbeta TNF-RI, G-CSF-R, PDGF-R, Blk, Ctk/Ntk, Lsk, Zap-70, PKB/RACalpha, PKC alpha, PKC-beta, PKC-gamma, PKC-delta, PKC-zeta, PKC-epsilon, PKC-eta, PKC-phi, PKC-mu, calmodulin-dependent kinase IIdelta, SLP-76-associated protein, FYN binding protein, Shc binding protein, RasGRF2, CDC25 homologue, Jak2, Jak3, PLCbeta1, and PLCbeta3. If ITIMs are defined by a broader consensus sequence, the list of ITIMs on activating molecules becomes even larger. In some instances, these ITIMs have been shown to associate with inhibitory phosphatases. Whether these ITIMs on activating receptors/pathways are necessary and sufficient for negative control of activating events and for immunologic tolerance is not yet known. In some instances, ITIMs on coinhibitory receptors are also required for appropriate negative regulation. By studying events leading to negative control during activation and to immunologic tolerance, it should be possible to discern the balance between antigen receptor-based negative events and coinhibition. PMID- 10872893 TI - Tolerance and immunity in the intestinal immune system. AB - The intestinal immune system must guard the body against invasion by pathogens while avoiding a response to the many potential antigens present in food. In the absence of the inflammatory stimuli necessary to elicit an immune response, oral administration of soluble protein antigens induces antigen-specific systemic nonresponsiveness. Recent studies have shown that peripheral nonresponsiveness to orally administered antigen is preceded by transient T-cell activation and is due primarily to the induction of functional T-cell anergy. The microenvironment of the gut-associated lymphoid tissue plays a central role in orally induced nonresponsiveness by supporting the growth of regulatory T cells that maintain intestinal homeostasis in the face of constant antigenic challenge. The transfer of nonresponsiveness by peripheral T cells from antigen-fed mice suggests that these-gut-derived regulatory cells also function in peripheral sites. When oral antigens are presented with adjuvants (microbial products that activate the innate immune system) an adaptive immune response is induced to this normally tolerogenic form of antigen. This review examines recent work that has provided new insight into the regulation of tolerance and immunity in the intestinal immune system. PMID- 10872894 TI - Phosphorylation-Based signaling in Fas receptor-mediated apoptosis. AB - Apoptosis or programmed cell death plays an essential role during development of the immune system, in immune responses, and in the control of tissue homeostasis in the adult. An important physiological mediator of apoptosis is the Fas/APO 1/CD95 receptor (FasR), a surface receptor belonging to the tumor necrosis factor receptor family. Apoptosis consists of a series of characteristic features that occur following activation of caspases, a collective term for apoptosis-specific proteases. The focus in FasR research has been on determining the mechanisms resulting in caspase activation. However, the role of phosphorylation-based signaling has received increasing attention both as an outcome of FasR activation and as a factor regulating FasR responses. Tyrosine-directed phosphorylation has been implicated to be induced and required during FasR stimulation. The FasR also activates all major signaling pathways that belong to the family of mitogen activated protein kinase (MAPK) pathways, by either caspase-independent or dependent mechanisms. Furthermore, phosphorylation-based signaling serves as a potent modifier of FasR responses. In this respect, especially the extracellular signal-regulated kinase and the phosphoinositide 3-kinase signaling pathways have been established as important regulators. This type of control seems to be directly phosphorylation-mediated without the requirement of newly synthesized proteins. Signaling through phosphorylation also regulates the expression of the Fas ligand (FasL), the FasR, as well as various other proteins that affect the outcome of receptor stimulation. While the involvement of phosphorylation has been established in FasR responses, the targets, molecular mechanisms, and biological significance of this aspect of the FasR signaling machinery still require further elucidation. PMID- 10872895 TI - Structure of IL-10 and its role in autoimmune exocrinopathy. AB - Cytokines are implicated in the pathogenesis of a variety of autoimmune disorders. Whether the presence of cytokines is primary or secondary and the extent to which these factors may contribute to the development and progression of clinicopathologic alterations in these disorders remain largely unknown but are highly important questions. Our research focuses on evaluating these issues by using a transgenic approach to direct the constitutive expression of cytokines to epithelial cells in the intact exocrine glands of mice. Our recent studies have focused on the potential of the regulatory cytokine IL-10 produced by type 2 T-helper cells to inhibit inflammation and the development of autoimmue diseases. Using targeted introduction of this molecule into the exocrine gland epithelial cells, we found that IL-10 induced apoptosis of glandular tissues destruction and lymphocyte infiltration consisting primarily of Fas-ligand+ CD4+ T cells, as well as in vitro upregulation of FasL expression on T cells. These results suggest that IL-10 is necessary and sufficient for exocrine gland dysfunction in the transgenic mice. This article discusses recent advances in IL-10 and its role in the pathological conditions of autoimmune exocrinopathy. PMID- 10872896 TI - A perspective on fat intake in athletes. AB - Performance in endurance events is dependent upon the maximal aerobic power, the percentage of that power that can be sustained and the availability of substrates (carbohydrates [CHO] and fats). The purpose of this paper is to present a perspective of recent studies that demonstrate the role of fat intake and oxidation on endurance performance. Studies have shown that fatigue is associated with reduced muscle glycogen and that increasing muscle glycogen or blood glucose prolongs performance while increasing fat and decreasing CHO decreases performance. This has led to an emphasis on CHO intake in athletes in endurance sports, which quite often leads to low caloric intake. It is well known that trained subjects have higher levels of fat oxidative capacity, which spares glycogen during endurance sports. Data from recent studies in trained athletes, who were fed iso-caloric high-fat diets (42% to 55%) that maintained adequate CHO levels, have shown an increase in endurance in both men and women when compared to diets composed of low fat intake (10% to 15%). The magnitude of the effect on endurance was significant at high percentages of maximal aerobic power and increased as the percentage of maximal aerobic power decreased. Based on this review, a baseline diet comprising 20% protein, 30% CHO and 30% fat, with the remaining 20% of the calories distributed between CHO and fat based on the intensity and duration of the sport, is recommended for discussion and future research. PMID- 10872897 TI - Incorporation of lean red meat into a National Cholesterol Education Program Step I diet: a long-term, randomized clinical trial in free-living persons with hypercholesterolemia. AB - OBJECTIVE: Clinicians often recommend that intake of all meat, particularly red meat, be reduced in conjunction with a low-fat, low-cholesterol diet to reduce low-density lipoprotein (LDL) cholesterol. This study was designed to determine the long-term effects of lean red meat (beef, veal and pork) compared to lean white meat (poultry and fish) consumption on lipoprotein concentrations in free living hypercholesterolemic subjects consuming a National Cholesterol Education Program (NCEP) Step I diet. METHODS: A randomized, crossover design was utilized. Hypercholesterolemic men and women (LDL cholesterol between 3.37 and 4.92 mmol/L) (triglycerides <3.96 mmol/L) (n = 145) were counseled to consume > or =80% of their 170 g/d meat intake as either lean red meat or lean white meat for two 36 week phases, separated by a four-week washout period of free meat selection. Subjects were instructed to follow an NCEP Step I diet throughout the study. RESULTS: There were no significant differences in lipid concentrations between the lean red meat and lean white meat phases. LDL cholesterol was 4.02+/-0.04 (SEM) and 4.01+/-0.04 mmol/L in the white and red phases, respectively; this represented a decrease of approximately 2% from baseline concentrations (p < 0.01). Total cholesterol also declined by 1% from baseline (p < 0.05), and high density lipoprotein (HDL) cholesterol rose over the study period by approximately 2% to approximately 3% from baseline to reach concentrations of 1.37+/-0.03 mmol/L and 1.38+/-0.03 mmol/L in the white and red phases, respectively (p < 0.001). Triglycerides were not altered by treatment. CONCLUSIONS: Consumption of lean red meat or lean white meat, as part of an NCEP Step I diet, is similarly effective for reducing LDL cholesterol and elevating HDL cholesterol concentrations in free-living persons with hypercholesterolemia. PMID- 10872898 TI - The effect of vitamin E and vitamin C supplementation on LDL oxidizability and neutrophil respiratory burst in young smokers. AB - OBJECTIVE: The purpose of this study was to determine the effect of vitamin E and/or vitamin C supplementation on low-density lipoprotein (LDL) oxidizability and neutrophil (PMN) superoxide anion production in young smokers. METHODS: Thirty smokers with a <5 pack-year history were randomly assigned to take placebo; vitamin C (1 g/day); vitamin E (400 IU/day), or both vitamins in a double-blind fashion. Subjects took the supplements for 8 weeks. At weeks 0 and 8, blood was collected for isolation of LDL and PMN, and for antioxidant vitamin analysis. LDL was oxidized with a copper (Cu) catalyst, and oxidation was measured by formation of conjugated dienes over a 5-hour time course. Lag times and maximum oxidation rates were calculated from the time course data. PMN superoxide anion release was assessed by respiratory burst after stimulation with phorbol ester and opsonized zymosan, and their ability to oxidize autologous LDL following treatment with the above stimuli was measured with the conjugated diene assay. RESULTS: Subjects who received vitamin E alone had a significant increase in the lag phase of Cu-catalyzed LDL oxidation (week 0, 118+/-31 min vs. week 8, 193+/-80 min, mean +/- SD, p < 0.05), whereas the vitamin C and placebo groups had no changes in LDL oxidation kinetics. The group receiving both vitamins E and C had a significant reduction in oxidation rate (week 0. 7.4+/-2.3 vs. week 8, 5.1+/-2.1, p < 0.05). There were no significant changes for any group in PMN superoxide anion production or PMN LDL oxidation after stimulation with either phorbol ester or opsonized zymosan. Plasma and LDL vitamin E concentrations were significantly increased in both groups that received vitamin E. The subjects who received vitamin C alone had no significant change in plasma vitamin C concentrations; however, when data were pooled from both groups who received vitamin C, the increases were significant. CONCLUSION: Vitamin E supplementation of young smokers was effective in reducing Cu-catalyzed LDL oxidizability; however, vitamin E and/or C supplementation showed few significant effects on the more physiologically relevant PMN function. This casts doubt on the ability of antioxidant supplementation to reduce oxidative stress in smokers in vivo. Therefore, smoking cessation remains the only means by which young smokers can prevent premature coronary heart disease. PMID- 10872899 TI - Lactoferrin levels in term and preterm milk. AB - OBJECTIVE: Even though there is no doubt that human milk is the best nourishment for the neonate, there is still controversy regarding its suitability for preterm infants. The aim of this study was to contribute to the knowledge of the anti infective properties of preterm milk, measuring lactoferrin levels, which are a non-specific protective factor. METHODS: Samples from 26 preterm and 20 term mothers (mean gestational age +/- standard deviation, 30.9+/-2.6 and 39.5+/-1.1 weeks, respectively) were collected during the first month post-partum. Milk samples were obtained by total expression of one breast between 10 a.m. and noon. An aliquot was kept at -20 degrees C until analyzed by SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Each sample was run in duplicate. RESULTS: Lactoferrin levels (X +/- SD) in colostrum and mature milk varied from 575.0+/-218.2 mg/dL to 459.4+/-190.7 mg/dL in preterm samples and from 970.6+/ 288.6 mg/dL to 292.0+/-167.4 mg/dL in term samples. No significant differences were observed between preterm and term groups, in spite of the trend observed in colostrum, where term milk tended to show higher levels than preterm milk. Decreasing values were observed in both groups along time (ANOVA, p<0.05). However, in the preterm group, lactoferrin levels seemed to maintain rather constant values from the eighth post-partum day onwards. CONCLUSIONS: The trend to higher levels of lactoferrin in preterm mature milk would allow maintenance of the protective effect of human milk in preterm infants in spite of the small volumes ingested by these neonates. These findings support the practice of feeding premature infants with their own mothers' milk at a time when their immune systems have not completely developed. PMID- 10872900 TI - Magnesium status and parameters of the oxidant-antioxidant balance in patients with chronic fatigue: effects of supplementation with magnesium. AB - OBJECTIVE: Magnesium deficiency and oxidative stress have both been identified as pathogenic factors in aging and in several age-related diseases. The link between these two factors is unclear in humans although, in experimental animals, severe Mg deficiency has been shown to lead to increased oxidative stress. METHODS: The relationship between Mg body stores, dietary intakes and supplements on the one hand and parameters of the oxidant-antioxidant balance on the other was investigated in human subjects. RESULTS: The study population consisted of 93 patients with unexplained chronic fatigue (median age 38 years, 25% male, 16% smokers and 54% with Chronic Fatigue Syndrome (CFS). Mg deficient patients (47%) had lower total antioxidant capacity in plasma (p=0.007) which was related to serum albumin. Mg deficient patients whose Mg body stores did not improve after oral supplementation with Mg (10 mg/kg/day) had persistently lower blood glutathione levels (p=0.003). In vitro production of thiobarbituric acid reactive substances (TBARS) by non-HDL lipoproteins incubated with copper was related to serum cholesterol (p<0.001) but not to Mg or antioxidants and did not improve after Mg supplementation. In contrast, velocity of formation of fluorescent products of peroxidation (slope) correlated with serum vitamin E (p<0.001), which was, in turn, related to Mg dietary intakes. Both slope and serum vitamin E improved after Mg supplementation (p<0.001). CONCLUSIONS: These results show that the lower antioxidant capacity found in moderate Mg deficiency was not due to a deficit in Mg dietary intakes and was not accompanied by increased lipid susceptibility to in vitro peroxidation. Nevertheless, Mg supplementation was followed by an improvement in Mg body stores, in serum vitamin E and its interrelated stage of lipid peroxidation. PMID- 10872901 TI - Fasting lipoprotein and postprandial triacylglycerol responses to a low carbohydrate diet supplemented with n-3 fatty acids. AB - BACKGROUND: The effects of a prolonged low-carbohydrate diet rich in n-3 fatty acids on blood lipid profiles have not been addressed in the scientific literature. OBJECTIVE: This study examined the effects of an eight-week ketogenic diet rich in n-3 fatty acids on fasting serum lipoproteins and postprandial triacylglycerol (TG) responses. DESIGN: Ten men consumed a low-carbohydrate diet rich in monounsaturated fat (MUFA) and supplemented with n-3 fatty acids for eight weeks. Fasting blood samples were collected before and after one week of habitual diet and on two consecutive days after 2, 4, 6 and 8 weeks of the intervention diet. Postprandial TG responses to a fat-rich test meal were measured prior to and after the intervention diet. RESULTS: Compared to the habitual diet, subjects consumed significantly (p < or = 0.05) greater quantities of protein, fat, MUFA and n-3 fatty acids and significantly less total energy, carbohydrate and dietary fiber. Body weight significantly declined over the experimental period (-4.2+/-2.7 kg). Compared to baseline, fasting total cholesterol, LDL cholesterol and HDL cholesterol were not significantly different after the intervention diet (+1.5%, +9.7% and +10.0%, respectively). Fasting TG were significantly reduced after the intervention diet (-55%). There was a significant reduction in peak postprandial TG (-42%) and TG area under the curve (-48%) after the intervention diet. CONCLUSIONS: A hypocaloric low-carbohydrate diet rich in MUFA and supplemented with n-3 fatty acids significantly reduced postabsorptive and postprandial TG in men that were not hypertriglyceridemic as a group before the diet. This may be viewed as a clinically significant positive adaptation in terms of cardiovascular risk status. However, transient increases in total cholesterol and LDL cholesterol were also evident and should be examined further in regard to which particular subfractions are elevated. PMID- 10872902 TI - Folate status worsens in recently institutionalized elderly people without evidence of functional deterioration. AB - OBJECTIVE: To follow folate status, hematological and cognitive changes during the first year of institutionalization among elderly subjects. DESIGN: Prospective study. SETTING: Long-stay unit of the Dijon University Geriatric Hospital. SUBJECTS: Twenty women and four men older than 65 years admitted consecutively. MAIN OUTCOME MEASURES: Folate and vitamin B-6 dietary intake was evaluated by a five-day record on admission (day 1 or d 1), at day 45 (d 45), day 90 (d 90), day 135 (d 135), day 180 (d 180), day 360 (d 360). Circulating levels of folate, vitamin B-6, total homocysteine (tHcy), blood counts and cognitive performance were determined in parallel. RESULTS: From d 1 to d 360, mean folate and vitamin B-6 intakes remained below the French RDA and mean folate intakes decreased significantly (delta = 10.2%, p < 0.05). Mean plasma or erythrocyte folate decreased significantly (delta = 33.7%, p <0.05 and delta = -30.2%, p < 0.001, respectively) from d 1 to d 360; no significant change was observed for the other blood parameters. The incidence of folate deficiency increased (8% vs. 37% for plasma folate <6.8 nmol/L and 8% vs. 17% for erythrocyte folate <340 nmol/L) from d I to d 360. Mean plasma pyridoxal 5'-phosphate (PLP) remained <20 nmol/L during the one-year follow-up. There was no difference between genders for plasma tHcy. Although mean plasma tHcy was <14 micromol/L. plasma tHcy was >14 micromol/L in about one-third of the subjects. At each period, 50% or more subjects were anemic (Hct <35% in women and Hct <40% in men), but the anemia was normocytic (MCV <100 fL). Subjects had a moderate dementia at admission, and no change was observed during the study. CONCLUSIONS: Subjects were already vitamin B-6 deficient at admission. Folate status was impaired during the study. Low vitamin intakes were the main cause of vitamin B-6 deficiency and folate status deterioration. Hematology and mental status capacity were not aggravated by folate status deterioration. Plasma tHcy didn't appear to be an earlier predictor of folate deficiency. PMID- 10872903 TI - Addition of supplementary foods and infant growth (2 to 24 months). AB - OBJECTIVE: To determine the effect of adding supplementary foods on infant growth 2 to 8 and 12 to 24 months. METHODS: Length (cm/month) and weight (kg/month) of white infants (n = 94) were measured five to nine times from 2 to 24 months of age. Mothers reported birth weights, infants' ages at first introduction of supplementary food, illnesses and information sources about infant feeding. Simple linear regression equations were used to compute slopes for each child (unit changes in length and in weight by age). Stepwise linear regression was used to determine the effect on weight and length slopes by the introduction of supplementary foods (e.g., an infant's age when cereal, fruit, juice, vegetables and a meat cluster were first added) to the diet. Breast feeding (months duration or ever fed), illness scores and gender were covariates in the regression models. RESULTS: A significant model (F = 10.09, p = .002) for weight gain (2 to 8 months) showed that gender explained 10% of the variance; for length slope, the model was non-significant and gender explained 3% of the variance. Females had a slower weight gain compared to that of males. None of the covariates or supplementary foods were retained in the models. Weight prior to 12 months was the best predictor (p = .0001, 54% of the variance) of weight gain 12 to 24 months. CONCLUSIONS: Unit changes in weight or length for an infant's age were not statistically associated with the timing of when supplementary foods were first added to the diet 2 to 8 or 12 to 24 months. Weight prior to 12 months was a significant predictor of weight gain 12 to 24 months. PMID- 10872904 TI - Basal urinary zinc/creatinine ratio as an indicator of dietary zinc intake in healthy adult women. AB - OBJECTIVE: To study, in healthy women, the correlation between the basal urinary zinc/creatinine ratio and dietary zinc intake. SUBJECTS: A group of 36 healthy female University students was evaluated. Mean age and body weight were, respectively, 25.6+/-3.3 years and 54.4+/-7.0 kg. METHODS: Basal urine was collected; Zn was determined by AAS and Creatinine (Creat) by the Jaffe method. A nutritional survey of seven days was recorded. Mean daily dietary intake of energy (DE) and zinc (DZn) were calculated according to the INCAP and English or German Food Composition Tables, respectively. RESULTS: Mean dietary daily intake were as follows (x +/- SD): Energy (kcal): 1606+/-570; zinc (mg): 9.1+/-3.8; basal urine Zn/Creat ratio: 0.41+/-0.24. Individual values of the Zn/Creat ratio correlated with dietary Zn (r=0.481, p=0.0339); data grouped according to ranges of dietary Zn fit the following equation: Zn/Creat=0.160+/-0.034 DZn (mg/day); (r=0.870, p=0.00497). CONCLUSIONS: These results showed that the basal urinary Zn/Creat ratio could be a useful indicator of dietary Zn intake in healthy adult women. PMID- 10872905 TI - Anaerobic bacteria and non-gonococcal urethritis. AB - Anaerobic bacteria are frequent inhabitants of the urethra of both normal men and men with non-gonococcal urethritis. All microbiologically-based studies have shown them not to have a role in the aetiology of the condition. However, Bacteroides ureolyticus continues to be an enigma having been isolated more commonly from men with urethritis in some studies, not confirmed by others, as well as in treatment-based studies in which the organism has been implicated by some authors. Few studies related to anaerobic organisms in the male genital tract have been conducted during the last decade. PMID- 10872906 TI - Actinomycosis in HIV infection: a review of a rare complication. AB - The emergence of the human immunodeficiency virus (HIV) and the onset of the AIDS epidemic has been associated with the frequent presentation of otherwise rare opportunistic infections and neoplasms. Despite the impairments of cellular and humoral immunity that accompany HIV infection, the prevalence of actinomycosis in the HIV-infected population has remained low. This article reviews previously reported cases of actinomycosis in HIV-positive and AIDS patients. Microbiological, pathological, diagnostic, clinical and therapeutic aspects of actinomycosis in this population are discussed. Clinicians should be aware of the possibility of actinomycosis as the cause of a persistent inflammatory lesion in these patients and know the correct techniques for collecting and submitting tissue specimens for anaerobic culture. PMID- 10872907 TI - The association of Mycoplasma hominis, Ureaplasma urealyticum and Mycoplasma genitalium with bacterial vaginosis: observations on heterosexual women and their male partners. AB - The prevalence of 3 mycoplasmas (Mycoplasma hominis, Ureaplasma urealyticum and Mycoplasma genitalium) was determined in a cohort of women with or without bacterial vaginosis (BV) and in their respective male partners. Heterosexual women with or without BV and their male partners were recruited and genital sampling for these microorganisms was performed. Seventeen women with BV and 21 women with normal flora, and their respective male partners, were recruited. M. hominis was present in 9 (53%) of 17 women with BV compared with none of 21 women without BV (P=0.0001). Of the 17 male partners of women with BV, 8 (47%) had M. hominis compared to 5 (24%) of 21 male partners of women without BV (not significant [n/s]). U. urealyticum was detected in 11 (65%) of 17 women with BV in comparison with 10 (48%) of 21 women without BV (n/s). U. urealyticum was present in 4 (24%) of 17 male partners of women with BV compared to 6 (29%) of 21 male partners of women without BV (n/s). M. genitalium was not detected in any of 15 women with BV and in only 2 (12%) of 17 women without BV (n/s). M. genitalium was present in 4 (25%) male partners of 16 women with BV in comparison with 3 (16%) male partners of 19 women without BV (n/s). Thus, M. hominis was the only mycoplasma detected significantly more often in women with, rather than in those without, BV. None of the mycoplasmas was found significantly more often in male partners of women with, rather than those without, BV. Overall, M. genitalium behaved somewhat similar to Chlamydia trachomatis. It was the least commonly occurring mycoplasma, a reflection perhaps of the relatively low incidence of partner change in this study population. PMID- 10872908 TI - The reasoning behind decisions not to take up antiretroviral therapy in Australians infected with HIV. AB - A substantial minority of HIV-infected Australians are not taking antiretroviral drugs. This study investigated the reasons behind their decision not to do so. Anyone who was HIV-infected but not taking antiretroviral drugs could participate. A self-administered, anonymous questionnaire was used, the principal recruitment method being through insertion of the questionnaire into gay community newspapers in Sydney and Melbourne. All respondents were asked questions covering demographics, previous AIDS-defining illnesses, T-cell and viral load monitoring, and previous use of antiretroviral drugs. In addition, respondents who had considered going on antiretroviral treatment, but then decided not to do so, were given a list of possible reasons for their decision and asked to indicate how much each played a role in their thinking. Of the 270 respondents, the great majority were gay men. One-eighth had experienced AIDS defining illnesses. Two-thirds had recently had T-cell and viral load tests. One third had taken antiretroviral drugs previously. Over two-thirds had considered antiretroviral therapy, most having given the matter quite some thought. Reasons for not taking up therapy did not differ greatly at different stages of HIV disease. The most common individual reason was fear of side effects. Important themes that emerged from factor analysis of the reasons data included distrust of conventional medical approaches to treatment, practical problems associated with taking antiretroviral drugs, unpleasant thoughts that being on therapy would evoke, and acceptance of the idea of dying. The findings can be used by doctors and counsellors to help patients clarify and evaluate their concerns about antiretroviral therapy. PMID- 10872910 TI - Factors influencing delay in treatment seeking by first-time attenders at a genitourinary clinic. AB - We examined treatment-seeking behaviours amongst a consecutive sample of 188 first-time attenders at a busy genitourinary (GU) clinic in a general hospital. Participants were interviewed and completed a battery of questionnaires prior to receiving diagnosis or treatment. Delay in treatment seeking was measured in 3 ways: utilization delay, illness behaviour delay and appraisal delay. Appraisal delay and illness behaviour were significantly related, but were unrelated to utilization delay. Substantial delays were reported before seeking treatment, with a median of 30 days elapsing between first noticing symptoms and attending the clinic. Analysis of age groups indicated that the oldest groups--45 and above were likely to delay longest, on all 3 measures of delay. PMID- 10872909 TI - Management of intravaginal warts in women with 5-fluorouracil (1%) in vaginal hydrophilic gel: a placebo-controlled double-blind study. AB - The purpose of this placebo-controlled, double-blind study was to determine the safety, tolerability and clinical efficacy of 5-fluorouracil (1%) in a vaginal hydrophilic gel (hydroxyethylcellulose, 1%) to cure intravaginal papillomas in women. Pre-selected, 60 women ranging between 18 and 50 years of age (mean 24.6), having 312 vaginal condylomas (mean 5.2) joined the study. The diagnosis of human papillomavirus (HPV) was established with clinical, histopathological and polymerase chain reaction (PCR) techniques. Subjects were randomized into 2 parallel groups. Each patient was allocated a pre-coded tube 15 g (active or placebo) with graduated vaginal applicators (disposable), and instructions how to insert 4 g of the trial medication deep into the vagina once at bedtime on every other day (1, 3 and 5) per week, to visit the clinic on day 7 for clinical evaluations and to receive the same pre-coded replacement to continue the regimen for another week. A maximum 12 applications were to be used in 4 weeks. Cure was defined as absence of clinical signs of infection, re-confirmed by PCR and Southern blot hybridization negative HPV DNA. By the end of the treatment 48.4% patients and 51.9% lesions were cured. Breaking the code revealed that 5 fluorouracil (1%) gel had cured 83.3% patients and 87% intravaginal warts. Placebo resolved 13.3% patients and 14% condylomas; (active gel versus placebo; P < 0.001). Twelve patients (20%) mostly in the active gel experienced mild erythema, erosion and oedema, with no drop-outs. Among cured patients 3 had a relapse after 16 months. In conclusion, the clinical results of the study demonstrate that 5-fluorouracil (1%) in a vaginal hydrophilic gel is safe, tolerable and significantly more effective than placebo to cure intravaginal warts in women. PMID- 10872911 TI - Serological prevalence of herpes simplex virus type 2 amongst GUM clinic attenders in a district general hospital setting. AB - Our objective was to determine the seroprevalence of herpes simplex virus (HSV) type 2 infection amongst genitourinary medicine (GUM) clinic attenders at a district general hospital using a commercially available enzyme immunoassay (EIA). In a prospective study, heterosexual patients attending the Department of GUM at Trafford General Hospital attending with a new clinical problem and having a blood sample taken for routine syphilis serology had the same sample tested for HSV type 2 antibodies. The prevalence of HSV type 2 seropositivity amongst participants was 9.9% (24/242) for men and 18.7% (46/246) for women. With respect to undiagnosed, asymptomatic infection the seroprevalence was 8.6% and 17% respectively. For those attenders locally resident the seroprevalence was 10.1% and 17.5% respectively, and undiagnosed, asymptomatic infection 8.5% and 17.1% respectively. Although seroprevalence figures in this study are lower than the only previous report in the UK, these results, nevertheless, show that seropositivity is not confined to large urban centres. Patients attending GUM clinics are likely to have high rates of undiagnosed HSV type 2 infection. PMID- 10872912 TI - Sexual behaviour and infection rates for HIV, blood-borne and sexually transmitted infections among patients attending drug treatment centres in Rio de Janeiro, Brazil. AB - A survey was carried out in 2 drug use treatment centres (TCs) in Rio de Janeiro, Brazil, to assess risk behaviours, HIV infection and other sexually transmitted infections/blood-borne infections (STIs/BBIs). Two hundred and twenty-five drug users (195 males and 30 females) were interviewed and clinically examined, and their blood and urine were tested for STIs/BBIs. Prevalences (%) for these infections were as follows--HIV: 0.9, hepatitis B virus (HBV): 14.7, hepatitis C virus (HCV): 5.8, syphilis: 5.3, gonorrhoea/chlamydia (CT/NG): 4.7. In bivariate analyses CT/NG infection was associated with younger age (P=0.003); current genitourinary symptoms (odds ratio [OR]=6.2) and a mainly illegal source of income (OR=9.1). Hepatitis C infection was associated with a history of ever having injected any drug (OR=19.6), and with each one of the injected drugs. After multiple logistic regression, lower educational level (adjusted odds ratio [AOR]=3.70) and 'ever having injected drugs' (AOR=3.69) remained as independent risk factors for hepatitis B infection. In conclusion, TCs must implement programmes directed towards the prevention of STIs/BBIs. PMID- 10872913 TI - Risk factors for HIV infection among asymptomatic pregnant women attending an antenatal clinic in western Kenya. AB - Our objective was to evaluate HIV prevalence and identify risk factors for HIV infection among women attending the antenatal clinic (ANC) at a large public hospital in Kisumu town, western Kenya. Between June 1996 and November 1997, in the context of a study to determine the effect of placental malaria on mother-to child transmission of HIV in western Kenya, HIV-1 antibody testing was offered to women with a singleton uncomplicated pregnancy of > or =32 weeks' gestation attending the ANC. Women were interviewed using a structured questionnaire and had a fingerstick blood sample collected for haemoglobin (Hb), malaria smears, and HIV antibody testing. Overall HIV seroprevalence was 26.1% (743/2844) (95% confidence interval (CI): 24.5-27.7) and in bivariate evaluation was significantly associated with anaemia (Hb <11 g/dl) (risk ratio (RR) 1.8), malarial parasitaemia (RR 1.6), fever (axillary temperature > or =37.5 degrees C at screening) (RR 1.6), a history of being treated for either vaginal discharge (RR 1.5) or tuberculosis (RR 1.6), reported alcohol consumption (RR 1.6), being an unmarried multigravida (RR 2.2) or a history of the most recent child having died (RR 2.0). Poisson regression analysis for all women identified 5 significant factors independently associated with HIV seropositivity: anaemia (adjusted RR 1.7; 95% CI 1.3-2.0), malarial parasitaemia (adjusted RR 1.7; 95% CI 1.4-2.0), a history of being treated for vaginal discharge (adjusted RR 1.5; 95% CI 1.1-2.0), fever (adjusted RR 2.0; 95% CI 1.3-3.2) and reported alcohol consumption (adjusted RR 1.6; 95% CI 1.1-2.5). Multigravidae women whose most recent child had died were also more likely to be HIV seropositive (adjusted RR 1.9; 95% CI 1.7-2.8). Only 5.5% (156/2844) of the women had none of these risk factors, of whom 12% (18/156) were HIV(+). Even though the model containing the 5 identified factors fitted the data well (goodness-of-fit chi2=18.41, P=0.10), its collective capacity to predict HIV infection was poor; while 74% of the truly positive women were correctly predicted positive by the model, 52% of the truly negative women were misclassified. Among pregnant women attending the ANC in western Kenya, we were unable to identify a subgroup at risk of HIV infection using non-serological information, indicating that wherever possible universal access to voluntary HIV counselling and testing would be preferable to targeted screening. PMID- 10872914 TI - The prevalence of urethral infections amongst asymptomatic young men in Hat Yai, southern Thailand. AB - The aim of this study was to survey sexual behaviour and estimate the prevalence of urethral infections amongst male vocational college students. A cross sectional survey was performed among 479 young men attending 2 vocational colleges in Hat Yai, southern Thailand. Polymerase chain reaction (PCR) tests of first-void urine (FVU) samples were used to detect infection with Chlamydia trachomatis, Neisseria gonorrhoeae, Ureaplasma urealyticum, Mycoplasma genitalium and Mycoplasma hominis. Girlfriends were the usual sexual partners for 89% of men with only 11% regularly patronizing sex workers. Condom usage was low. The prevalence of any urethral infection was 15.9% with: C. trachomatis 4%, N. gonorrhoeae 0.2%, U. urealyticum 10.9%, M. genitalium 2.3% and M. hominis 1.3%. Infection with more than one organism was found in 2% of men. While the prevalence of infection with chlamydia or gonorrhoea was relatively low, the prevalence of 'any urethral infection' was moderately high and suggests that unprotected sexual intercourse is commonly occurring. As girlfriends were the most usual sexual partners, they must be at significant risk of pelvic infection. There is a need for programmes targeting this group of people. PMID- 10872915 TI - Factors affecting attitudes towards mother-to-child transmission of HIV among pregnant women in a maternal and child hospital in Thailand. AB - This study determines the factors which correlate with attitudes towards mother to-child transmission of HIV in pregnant women. Using a structured questionnaire, 527 pregnant women who visited a hospital to have prenatal checkups were interviewed. The survey items were: sociodemographic characteristics, experiences of pre-test counselling, knowledge of mother-to-child transmission, and attitude towards termination of pregnancy. Results showed that many pregnant women (80%) did not have proper knowledge of the possibility of mother-to-child transmission. Logistic regression analysis also indicates that age and knowledge of the possibility of mother-to-child transmission were the significant determinants of attitudes towards termination of pregnancy. Older women who believe that all the babies of pregnant women with HIV will be infected are most likely to terminate their pregnancy when they are diagnosed as HIV positive. Considering the importance of informed decisions regarding pregnancy, this study must have important implications for future support programmes for HIV-positive pregnant women. PMID- 10872916 TI - Silent oophoritis due to cytomegalovirus in a patient with advanced HIV disease. AB - A case of isolated necrotizing cytomegalovirus (CMV) oophoritis disclosed only by necropsy studies in a patient with AIDS, is described. This unusual case report is discussed with a review of the literature dealing with CMV involvement of genital organs in the immunocompromised host, and in patients with HIV infection and AIDS. PMID- 10872917 TI - Metastatic tufted angioma. PMID- 10872918 TI - Effect of SR121463, a selective non-peptide vasopressin V2 receptor antagonist, in a rabbit model of ocular hypertension. AB - The activity on intraocular pressure (IOP) of SR121463, a selective non-peptide arginin-vasopressin (AVP) V2 receptor antagonist, was investigated in a rabbit model of ocular hypertension. We first demonstrated that, in vitro, SR121463 displayed high competitive affinity for rabbit vasopressin V2 receptors (Ki = 2.1 +/- 1.2 nM). In vivo, SR121463 was instilled once (at concentrations ranging from 0.1 to 3%), or for 10 days (20 instillations) at 1% concentration, in the eye of ocular hypertensive rabbits (intraocular injection of 0.14 mg alpha chymotrypsin). SR121463 also was instilled at 1% in the normotensive eye or intravenously injected (100 microg/kg) to ocular hypertensive rabbits. SR121463 was compared to timolol 0.5% or to clonidine 0.25%. Additionally, local and systemic safety aspects were examined. Results showed that SR121463 was locally well-tolerated and had no anesthetic effect. A significant decrease in IOP of the hypertensive eye was observed for concentrations of SR121463 > or =1%. This decrease was comparable to that obtained with reference compounds. A similar activity was found after intravenous administration. No tachyphylaxis was observed after 10 days, and no contralateral or systemic effect was noted. Also, when applied on the normotensive eye or when intravenously injected, SR121463 had no effect on the normotensive eye. These results on IOP and the good local and systemic safety profile, suggest that a potent vasopressin V2 receptor antagonist, SR121463, could be of value for the treatment of glaucoma, through a mechanism of action that remains to be elucidated. PMID- 10872919 TI - Pharmacology of the intraocular pressure (IOP) lowering effect of systemic dexanabinol (HU-211), a non-psychotropic cannabinoid. AB - The purpose of this study was to characterize the intraocular pressure (IOP) lowering activity and possible mechanism of action of the synthetic, non psychotropic cannabinoid dexanabinol (HU-211) [(+)(3S,4S), 7-hydroxy-delta-6 tetrahydrocannabinol 1,1 dimethylheptyl], following intravenous (i.v.) administration in the rabbit. IOP (pneumatonometry), aqueous humor inflow rate (fluorophotometry), blood pressure, and heart rate (computerized physiograph system connected to central ear artery cannula) were measured in unanesthetized albino rabbits. Intravenous administration of HU-211 resulted in a dose-related reduction in IOP; a maximal IOP reduction of 5.0 +/- 0.2 mmHg was observed 4 hr after a 0.5 mg/kg dose. No significant changes in blood pressure or heart rate were observed during the first hr following this dose of HU-21 1. Pupil diameter did not change significantly during the 5 hr following the 0.5 mg/kg i.v. dose. No significant change in the rate of aqueous humor inflow occurred during the 6 hr after a 0.5 mg/kg dose of HU-211, thereby implicating outflow changes as the major source of IOP reduction. IOP reduction by HU-211 following pre-treatment with the alpha2 adrenergic antagonist, yohimbine (1 mg/kg, i.v.), was only 30% of that of HU-211 alone. IOP reduction following pretreatment with the alpha2 agonist, clonidine (0.5 mg/kg i.v.), was twice as large as that of HU-211 alone. Pretreatment with the beta-adrenergic antagonist, propranolol (0.5 mg/kg i.v.), resulted in a 50% reduction in the IOP-lowering effect of HU-211. In summary, HU 211, administered i.v., is an effective IOP-lowering agent, devoid of any significant side effects (blood pressure, heart rate or pupil diameter, all of which have been reported previously for cannabinoids). Involvement of the adrenergic system is indicated in mediating the IOP-lowering effects of HU-211 that appear to reflect a change in fluid outflow from the eye. PMID- 10872920 TI - Effects of semotiadil, a novel calcium antagonist, on the retina and optic nerve head circulation. AB - The effects of semotiadil, a novel benzothiazine calcium antagonist, on the retinal and optic nerve head (ONH) tissue circulation were evaluated using the noninvasive laser speckle method. In urethane-anesthetized Dutch or albino rabbits, before and up to 90 min following intravenous injection of 400 microg/kg semotiadil fumarate (semotiadil group) or vehicle (control group), normalized blur value, a quantitative index of tissue blood velocity, in the retina (NB(retina)) or ONH (NB(onh)), was serially obtained with monitoring intraocular pressure (IOP) and systemic parameters: arterial pressure, pulse rate, arterial blood gas, and body temperature. There were no significant differences in IOP and the systemic parameters except arterial pressure between semotiadil and control groups during the experiments. Arterial pressure showed an acute and transient drop during the first 5 min after semotiadil administration. The time courses of the normalized blur value were significantly different between semotiadil and control groups in the retina (P = 0.0001, repeated measures two-way ANOVA), but not in the ONH (P = 0.6724). Changes in NB(retina) from the baseline in the semotiadil group was significantly greater than those in the control group 50 min or later after the administration (P < 0.0500, Mann-Whitney test). NB(onh) showed no significant differences between the two groups except during the first few min when arterial pressure acutely decreased in the semotiadil group. In conclusion, intravenously injected semotiadil increased the tissue blood velocity in the retina, but not in the ONH. This vascular selectivity in the ocular neural tissues differs from those of other calcium antagonists, such as nicardipine. PMID- 10872921 TI - Effect of ifenprodil on aqueous humor dynamics and optic nerve head circulation in rabbits. AB - The purpose of this study was to study the effects of ifenprodil, a cerebral vasodilator with alpha and N-methyl-D-aspartate (NMDA) receptor antagonistic activities, on aqueous humor dynamics and optic nerve head (ONH) circulation in rabbits. Experiments were performed during the dark phase in rabbits conditioned to a schedule of alternating 12-hr periods of light and dark. Effects on blood aqueous barrier permeability (K(d)), aqueous flow rate (F), outflow facility to general blood circulation (C(gen)), and uveoscleral outflow (F(u) were determined fluorophotometrically. Effects on ONH tissue circulation were estimated using the laser speckle method. Unilateral topical administration of 0.5% ifenprodil solution decreased intraocular pressure (IOP) with a maximum reduction of 3.4 mmHg and an effect duration of 3 hr without effects on the contralateral eye. A single instillation of 0.5% ifenprodil had no significant effect on K(d), F, or C(gen), whereas it substantially increased F(u). Twenty-day, twice-daily unilateral 0.5% ifenprodil instillation significantly increased tissue blood velocity in the ONH only in the treated eye. PMID- 10872922 TI - Comparison of the efficacy and safety of latanoprost 0.005% compared to brimonidine 0.2% or dorzolamide 2% when added to a topical beta-adrenergic blocker in patients with primary open-angle glaucoma or ocular hypertension. AB - The purpose of this study was to evaluate the ocular hypotensive efficacy and safety of latanoprost 0.005% (Xalatan, Pharmacia & Upjohn), brimonidine (Alphagan, Allergan), and dorzolamide (Trusopt, Merck Inc.) when added to a beta blocker in patients with ocular hypertension or primary open-angle glaucoma. This was a multicenter, retrospective analysis which included all reviewed patient records in which latanoprost, brimonidine or dorzolamide were added to a beta blocker for at least three months. Patients who were treated for less than three months, who failed therapy due to ineffectiveness of the medicine or an adverse event also were included. The study included 141 patients. Latanoprost (n = 50) showed an intraocular pressure of 16.7 +/- 3.3 mm Hg (-6.3 +/- 4.1 mm Hg, P < 0.001), brimonidine (n = 24) 17.4 +/- 4.9 mm Hg (-4.2 +/- 4.5 mm Hg, P < 0.001), and dorzolamide (n = 67) 20.1 +/- 6.1 mm Hg (-3.1 +/- 5.1 mm Hg, P < 0.001) at three months. A significant difference was observed in the absolute level of intraocular pressure (P < 0.005) and the change from baseline between groups (P < 0.005) at three months. A significant difference was observed between groups in the success rate of therapy between latanoprost (70%), brimonidine (58%) and dorzolamide (40%) (P = 0.008). No significant differences were observed between groups for rate or type of adverse events leading to discontinued therapy. This study showed that latanoprost, when added to beta-blockers, compares favorably in ocular hypotensive efficacy and is similar in safety to brimonidine and dorzolamide. PMID- 10872923 TI - Chitosan as tear substitute: a wetting agent endowed with antimicrobial efficacy. AB - A cationic biopolymer, chitosan, is proposed for use in artificial tear formulations. It is endowed with good wetting properties as well as an antibacterial effect that are desirable in cases of dry eye, which is often complicated by secondary infections. Solutions containing 0.5% w/v of a low molecular weight (M(w)) chitosan (160 kDa) were assessed for antibacterial efficacy against E. coli and S. aureus by using the usual broth-dilution technique. The in vitro evaluation showed that concentrations of chitosan as low as 0.0375% still exert a bacteriostatic effect against E. coli. Minimal inhibitory concentration (MIC) values of chitosan were calculated to be as low as 0.375 mg/ml for E. coli and 0.15 mg/ml for S. aureus. Gamma scintigraphic studies demonstrated that chitosan formulations remain on the precorneal surface as long as commonly used commercial artificial tears (Protagent collyrium and Protagent SE unit-dose) having a 5-fold higher viscosity. PMID- 10872924 TI - Involvement of cysteine proteases in bFGF-induced angiogenesis in guinea pig and rat cornea. AB - Overexpression and activation of matrix metalloprotease (MMP) have been implicated in angiogenesis. However, the involvement of cysteine proteases, such as calpains (EC 34.22.17), is obscure. Thus, the purpose of this experiment was to study the involvement of cysteine proteases in angiogenesis induced by basic fibroblast growth factor (bFGF) in guinea pig and rat corneas using cysteine protease inhibitors. Sustained-release polymers containing bFGF were implanted into guinea pig and rat corneas to induce angiogenesis. For treatment of corneal angiogenesis, polymers containing cysteine protease inhibitors, leupeptin or SJA6017, were also implanted into corneas. Using the slit lamp, the corneas were observed for nine days after polymer implantation. Soluble proteins and albumin levels were used as markers of corneal injury by angiogenesis. bFGF induced angiogenesis in guinea pig and rat corneas. In guinea pig cornea, wet weight, the amount of soluble protein and albumin was highest at four days after bFGF containing pellet implantation. In rat cornea, the amount of soluble protein and albumin was highest at six days, and wet weight increased within four days. One hundred nmole of leupeptin showed a tendency to reduce bFGF-induced angiogenesis in guinea pig cornea, and 10 nmole of SJA6017 was effective in reducing bFGF induced angiogenesis in rat cornea, although SJA6017 showed a stronger effect than leupeptin. Ten nmole of SJA6017 significantly reduced the number of new blood vessels. These data suggested involvement of cysteine proteases in angiogenesis in guinea pig and rat cornea. PMID- 10872925 TI - Ultraviolet radiation and cataract. AB - While solar radiation falling on earth comprises light in the infrared, visible, UVA, UVB, and even UVC ranges, the light incident on, and thus important to the biology of, the eye lens is essentially in the visible and UVA regions. Thus, direct photochemical damage to the lens from UVB radiation is minor, though long term UVA (and even visible range) irradiation is seen to lead to lens malfunction. Short-term exposure of the lens in vivo to UVA light leads to compromised optical and biochemical properties which are repaired in time, while higher doses affect permanent damage. Such longer wavelength light-mediated changes in the lens occur through photodynamic means, affected by some of the compounds that accumulate in the lens over a period of time, which act as sensitizers. Isolation and chemical identification of over a dozen such compounds has been done, and their photoactive properties have been studied. While several of these are photodynamic and generate reactive oxygen species when UVA light is shone on them, other compounds that accumulate in the lens act as antioxidants. PMID- 10872926 TI - Features of macular hole closure in the early postoperative period using optical coherence tomography. AB - PURPOSE: Studying the retina in the early postoperative period after macular hole surgery is difficult because of the limitation of imaging of the fundus through a gas tamponade. Silicone oil was shown recently to be an effective alternative to gas for macular hole repair. The authors hypothesized that optical coherence tomography (OCT) could be performed through silicone oil to study early macular hole closure. METHODS: Fourteen patients with idiopathic full-thickness macular holes underwent repair, including vitrectomy and silicone fill of the vitreous cavity. Silicone oil was removed 5 to 18 weeks later in a second operative procedure. Optical coherence tomography images were obtained preoperatively (n = 14), on the first postoperative day (n = 7), after 1 week (n = 4), after 1 month (n = 14), and after silicone oil removal (n = 14). RESULTS: By the first postoperative day, the retina was flat and the hole was open or closed in all patients. By 1 month, 9 of 14 patients had flat retinas and closed holes, whereas the other 5 patients had flat retinas and open holes. All patients who had an open hole after 1 month had a reopening of the hole after silicone oil removal. CONCLUSIONS: Flattening of the retina with resolution of the foveal cysts after macular hole repair occurs by the first postoperative day. Closure of the horizontal retinal separation is variable but occurs by the first postoperative month in cases of successful macular hole repair. Persistent retinal separation after 1 month may be predictive of hole reopening. PMID- 10872927 TI - Evaluation of patients with visual field defects following macular hole surgery using multifocal electroretinography. AB - PURPOSE: To investigate patients with visual field defects following macular hole surgery to determine the cause of such defects, specifically with reference to ischemic damage versus mechanical trauma. METHODS: Five patients with known visual field defects following macular hole surgery were studied with Goldmann perimetry, Humphrey automated perimetry, and multifocal electroretinography (MERG). Three patients returned at a later date for nerve fiber layer analysis. RESULTS: None of the five patients demonstrated evidence of a- or b-wave loss on MERG in the regions corresponding to the visual field defects. Two of three patients studied with the nerve fiber layer analyzer demonstrated significant loss of nerve fiber layer thickness in the quadrant corresponding to the field defect. CONCLUSION: The normal MERG results indicate that the possibility of an arteriolar occlusion as the principal cause for the defects is unlikely in most cases. Data suggest that the site of damage is in the nerve fiber layer, although the specific cause of this damage remains to be determined. PMID- 10872929 TI - Intravitreal injection of tissue plasminogen activator and gas in subretinal hemorrhage caused by age-related macular degeneration. AB - PURPOSE: To assess the efficacy and safety of intravitreal injection of recombinant tissue plasminogen activator and sulfur hexafluoride gas for displacement of subretinal hemorrhages in patients with age-related macular degeneration. METHODS: The authors injected 25 microg of recombinant tissue plasminogen activator and 0.5 mL sulfur hexafluoride gas intravitreally in 11 patients with subretinal hemorrhages of less than 3 weeks duration. Anatomic and functional results were evaluated. RESULTS: Displacement of subretinal blood was successful within the first week after surgery in 10 of 11 patients. This was accompanied by visual improvement in eight patients. After 1 year, visual acuity was better than before surgery in five patients. Diagnosis of a choroidal neovascularization by fluorescein angiography was possible in all patients, and was treated with laser photocoagulation in five. The authors observed no adverse effects of treatment. CONCLUSION: Recombinant tissue plasminogen activator and gas effectively displace subretinal blood in patients with age-related macular degeneration. Randomized studies are necessary to prove the benefit of this simple and safe method in patients with subretinal hemorrhage due to age-related macular degeneration. PMID- 10872928 TI - Intravitreal triamcinolone acetonide in exudative age-related macular degeneration. AB - PURPOSE: To examine the effects of intravitreal injection of 4.0 mg triamcinolone acetonide on the visual and clinical course of exudative age-related macular degeneration. METHODS: A randomized clinical trial of a single injection of triamcinolone acetonide into the vitreous cavity of experimental eyes at baseline versus observation of untreated subjects was performed in 27 patients followed up for 6 months. Inclusion criteria included exudative age-related macular degeneration with subfoveal or occult choroidal neovascularization, and visual acuity between 20/40 and 20/400. Examination, acuity assessment, fundus photography, and fluorescein angiography were performed at baseline and at 3 and 6 months after enrollment. LogMAR visual acuity was compared between groups by a repeated measures analysis of variance model. Masked assessment of photographic studies was performed and groups were compared with Fisher's exact test. RESULTS: Visual acuity was significantly better in the treated group compared with control subjects at 3 and 6 months (P < 0.005). Fundus photography and angiography were more likely to show stability or improvement at 3 and 6 months in the treated group (P = 0.05). Intraocular pressure elevation was seen in 25% of treated patients, but was controlled with topical medications. Progression of cataract was more frequently seen in the treated group. CONCLUSIONS: Intravitreal triamcinolone acetonide may provide short-term improvement in visual acuity and fundus findings in exudative macular degeneration. These findings must be considered preliminary and should be followed by multicenter, masked, placebo controlled trials with long-term follow-up. PMID- 10872930 TI - Combined cataract surgery and vitrectomy for recurrent retinal detachment. AB - PURPOSE: To report our experience with combined cataract surgery, posterior chamber intraocular lens implantation, and pars plana vitrectomy in the management of recurrent retinal detachment (RD) and visually significant cataract. METHODS: Retrospective chart review of patients with cataract and recurrent RD who underwent combined cataract extraction, posterior chamber intraocular lens implantation, and pars plana vitrectomy between January 1991 and September 1998 at the Bascom Palmer Eye Institute. Sixteen eyes were included. All eyes had visually significant cataract and had undergone primary repair of the RD with encircling scleral buckle; eight eyes also had undergone pars plana vitrectomy during the primary repair. The technique of cataract extraction included phacoemulsification (10 eyes), extracapsular cataract extraction (5 eyes), and pars plana lensectomy (1 eye). All eyes underwent pars plana vitrectomy, membrane peeling, fluid-air exchange, endolaser treatment, and placement of a retinal tamponade. Perfluoropropane (C3F8) gas was used in 14 eyes, and silicone oil was placed in two eyes. RESULTS: The postoperative follow up interval ranged from 4 to 64 months (mean, 16 months). Preoperative visual acuity ranged from 20/60 to hand motions and was better than 20/200 in 3 (19%) eyes. Postoperatively, 9 (56%) eyes improved to better than 20/200. Anatomic success was achieved after the initial reoperation in 13 (81 %) eyes. With further surgery, the overall success rate was 94%. CONCLUSIONS: Combined cataract surgery, posterior chamber intraocular lens implantation, and pars plana vitrectomy in selected patients with cataract and recurrent RD was successful in improving visual acuity and achieving retinal reattachment in most of these reoperated patients. PMID- 10872931 TI - Fungal endophthalmitis after a single intravenous administration of presumably contaminated dextrose infusion fluid. AB - PURPOSE: To report fungal endophthalmitis in nonimmunocompromised patients, each of whom received a single intravenous administration of presumably contaminated dextrose infusion fluid for minor ailments in rural settings. METHODS: This noncomparative case series included 12 nonimmunocompromised patients (12 eyes) with culture-positive fungal endophthalmitis. All eyes underwent initial vitreous tap with injection of intravitreal antibiotics. Eleven eyes required pars plana vitrectomy and oral fluconazole or itraconazole for 4 to 6 weeks. One patient with panophthalmitis was treated with intravenous amphotericin B. To support the hypothesis that contaminated intravenous fluid was the possible risk factor, samples from 72 sealed bottles of 5% dextrose were subjected to fungal culture. RESULTS: Patients presented 1 to 11 weeks (mean, 4.6 weeks) after the intravenous infusion. All eyes had a positive smear and cultures for fungi. Aspergillus specimen was isolated in nine eyes, Candida in two eyes, and Mucor in one eye. Final visual acuity was 20/80 or better in 8 (66.6%) eyes. Eleven of the 72 samples from dextrose bottles were culture-positive for fungi: six for Aspergillus fumigatus, three for Aspergillus niger, and two for Candida albicans. CONCLUSION: A presumed contaminated intravenous infusion administered in a rural setting was found as a new risk factor for development of endogenous fungal endophthalmitis. These patients were successfully treated with pars plana vitrectomy and oral fluconazole and itraconazole therapy. PMID- 10872933 TI - Grading, image analysis, and stereopsis of digitally compressed fundus images. AB - PURPOSE: To investigate the effects of image digitization and compression on the ability to identify and quantify features in color fundus photographs. METHODS: Color fundus photographs were digitized as tagged image file format (TIFF) and high-compression (80:1) and low-compression (30:1) joint photographic experts group (JPEG) images. Rerendered images were subjected to standard grading protocols developed for a clinical trial, and digitized images were subjected to image analysis software for drusen identification and quantitation. Re-created stereoscopic images were compared subjectively with originals. RESULTS: Original, TIFF, and low-compression (30:1) JPEG images were virtually indistinguishable when subjected to close scrutiny with magnification. The overall quality of high compression (80:1) JPEG images and images digitized at 500 dots per inch was markedly reduced. Protocol grading of original and digitized images was highly concordant within the repeatability of multiple grading of original images. The area subtended by drusen differed by less than 1.0% for all uncompressed and compressed image pairs quantified. Stereoscopic information was accurately preserved when compared with originals for TIFF and low-compression JPEG images. CONCLUSIONS: Fundus images can be digitized and stored with significant compression while preserving stereopsis and image quality suitable for quantitative image analysis and semiquantitative grading. Low-compression (30:1) JPEG images may be suitable for archiving and telemedical applications. PMID- 10872932 TI - Subretinal abscess due to Nocardia farcinica infection. AB - PURPOSE: Nocardia infection of the eye is uncommon. A case of choroidal abscess due to Nocardia farcinica infection is presented, and the literature is reviewed. METHODS: A 41-year-old immunocompromised man with chronic myeloid leukemia developed a unilateral choroidal abscess. N. farcinica was isolated from a simultaneous subcutaneous abscess and both infections responded to systemic sulfonamide therapy. RESULTS: Three weeks after discontinuation of the sulfonamides, the choroidal abscess recurred with involvement of the vitreous. The infection was brought under control after reinstitution of the same drug. CONCLUSIONS: Nocardiosis is a multisystem disease that has high mortality and ocular morbidity rates. The eyes of immunocompromised patients should be examined frequently as early detection and administration of the proper antibiotics may reduce the risk of this life-threatening infection. PMID- 10872934 TI - Estimate and measure of the region of view and myopia resulting from vitreous gas. AB - PURPOSE: After vitreous gas injection, patients notice better acuity in downward gaze than in horizontal gaze. The authors evaluated the refractive error and the size of the region within which vision improves. METHODS: For the vitreous fluid gas interface, the authors calculated the angle of total internal reflection and the expected myopic shift and then measured them in nine consecutive patients. The volume of gas, declination angle at which perception of small targets occurred, and preoperative and postoperative refractive error in downward gaze were measured. RESULTS: Total internal reflection occurs at 41.5 degrees declination. Patients perceived a region of improved acuity below 41 degrees (+/ 5 degrees) declination. The difference between the calculated and clinically measured gas-induced myopia was less than 25% for five of nine patients. The largest measured induced myopia was -23.9 diopters (60% gas volume). In eight eyes, patients read 5-point type or smaller. CONCLUSIONS: Patients accurately perceive that their acuity improves in downward gaze; the boundary of this region corresponds with the angle of total internal reflection. Calculations predict that vitreous gas produces a myopic shift and aberration. These data support the notion that ocular positioning by patients with vitreous gas can be enhanced by instructing them to regard near targets in downward gaze. PMID- 10872935 TI - Ultrastructural and immunohistochemical findings in five patients with vitreomacular traction syndrome. AB - OBJECTIVE: To evaluate pathologic features of vitreomacular traction syndrome (VMT). METHODS: Preretinal membranes removed from five patients during vitreous surgery for VMT syndrome were evaluated by electron microscopy (n = 4) and immunohistochemistry (n = 1). RESULTS: Electron microscopic examination revealed large segments of internal limiting lamina (ILL) in three of the four cases. Other extracellular features included two types of abnormal collagen fibrils, determined to be type I and fibrous long-spacing collagen. The myofibrocyte was the predominant cell type in all cases. Collagen types I, II, and III, as well as glial fibrillary acidic protein and vimentin, were identified by immunohistochemistry. CONCLUSIONS: Epiretinal membranes in eyes with VMT syndrome adhered tightly to the ILL with abnormal collagen and contractile elements that included myofibrocytes. PMID- 10872936 TI - Diagnostic and therapeutic challenges. A 56-year-old man, had a 1-year history of blurred vision in his left eye. PMID- 10872937 TI - Bilateral vitreous hemorrhages in a patient with relapsing polychondritis and high levels of type II collagen antibodies. PMID- 10872938 TI - Unilateral central retinal artery occlusion followed by contralateral anterior ischemic optic neuropathy in giant cell arteritis. PMID- 10872939 TI - Cilioretinal artery occlusion in posterior scleritis. PMID- 10872940 TI - Occurrence of retinoblastoma and uveal melanoma in the same patient. PMID- 10872941 TI - Occult choroidal hemangioma revealed after vitrectomy for macular pucker. PMID- 10872942 TI - Indocyanine green angiography in Behcet's uveitis. PMID- 10872943 TI - Visualization of choroidal rupture with indocyanine green angiography. PMID- 10872944 TI - Retinal transscleral photocoagulation under endoscopic control. PMID- 10872945 TI - TPA-assisted vitrectomy for proliferative diabetic retinopathy. PMID- 10872946 TI - Suggested terminology for different phases of indocyanine green angiogram. PMID- 10872947 TI - Alveolar bone height and postcranial bone mineral density: negative effects of cigarette smoking and parity. AB - BACKGROUND: Our objective was to test the association between cemento-enamel junction, alveolar-crest distance (CEJ-AC, as measured on digitized vertical bite wing radiographs) and postcranial bone mineral density (BMD) relative to clinical, dietary, and demographic variables. METHODS: Data were collected in a cross-sectional study of 134 postmenopausal women. CEJ-AC distances were determined from digitized vertical bite-wing radiographs. Lumbar spine and proximal femur BMDs were determined from dual-energy x-ray absorptiometric scans. Correlation analysis and Student t tests were used to identify those variables most associated with CEJ-AC distance. The selected variables were modeled with a backward stepwise regression analysis, with CEJ-AC distance as the dependent variable. RESULTS: Parity (number of pregnancies to term), cigarette smoking, and the interaction of lateral spine BMD with cigarette smoking were independent predictors of CEJ-AC distance (P < or =0.05). Statistical models containing these variables accounted for 19% of the variation in CEJ-AC distances. CONCLUSIONS: CEJ-AC distance in postmenopausal women is the result of a complicated interaction of many effects, including but not limited to, parity, cigarette smoking, and skeletal BMD. PMID- 10872948 TI - Use of a fluorogenic septapeptide matrix metalloproteinase assay to assess responses to periodontal treatment. AB - BACKGROUND: Quantification of gingival crevicular fluid matrix metalloproteinase activity may provide improved assessment of periodontal disease status and response to treatment. A fluorogenic matrix metalloproteinase substrate assay (FSA) has been developed using a methoxycoumarin-containing septapeptide analog of the alpha2(I) collagen cleavage site. This substrate exhibits increased fluorescence following cleavage by many matrix metalloproteinases, and the enzyme activity can be readily estimated with a fluorimeter. Here we compared this assay with classical methods of periodontal assessment including bleeding on probing, crevicular fluid flow, and probing depth to assess its utility as an indicator of changes in periodontal status and treatment response. METHODS: Complete measurements of probing depth were obtained for Ramfjord teeth on subjects who had been previously treated for periodontitis. Subjects were subsequently divided into groups based on existing periodontal disease severity: gingivitis (n = 21), stable periodontitis (n = 41), and severe periodontitis (n = 50). Crevicular fluid volume, bleeding on probing, and FSA were measured at each Ramfjord tooth or substitute. After baseline measurements, subjects received subgingival scaling and prophylaxis; 3 months later, they were reassessed. RESULTS: FSA measurements were positively associated with severity of disease at baseline. After treatment there were substantial reductions of FSA in gingivitis (approximately 51%; P <0.01) and severe periodontitis (approximately 45%; P <0.001), but not in stable periodontitis (13%; P >0.2). All groups showed a positive association between FSA measurements and higher bleeding scores at individual sites. FSA measurements were also positively associated with crevicular fluid flow at baseline, but after treatment there was a approximately 67% decrease (P <0.01) in the highest crevicular fluid flow class. There were significant reductions of FSA at follow up for sites with probing depths between 0 to 3 mm (23%; P <0.05) and 4 to 6 mm (31%; P <0.05). However, the largest reduction was for sites with probing depth between 7 to 9 mm (49%; P <0.001). CONCLUSIONS: These results indicate that monitoring patients by measurement of matrix metalloproteinase levels in gingival crevicular fluid with the quenched fluorescent substrate assay provides estimates of inflammatory status, periodontal destruction, and response to treatment, especially in more severe periodontitis lesions. PMID- 10872949 TI - The detection and comparison of angiogenesis-associated factors in pyogenic granuloma by immunohistochemistry. AB - BACKGROUND: Pyogenic granuloma is a benign inflammatory lesion demonstrating obvious activity of angiogenesis. Female steroid hormones are believed to play important roles in the etiology because the lesion is frequently found in females with high levels of sex hormones. Few molecular mechanisms of the pathogenesis have been proposed and proven. The purpose of this study was to detect and compare the expression of angiogenesis-associated factors among healthy gingiva, gingiva from periodontitis, and pyogenic granuloma to clarify the pathogenesis of pyogenic granuloma. METHODS: Fifteen specimens were collected from each of 3 groups of gingiva (healthy gingiva, periodontitis, and pyogenic granuloma). The subjects were age and gender matched. The specimens were processed for immunohistochemistry to detect and compare the expression of 2 angiogenesis enhancers, i.e., vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), 2 angiogenesis inhibitors, i.e., angiostatin and thrombospondin-1 (TSP-1), and estrogen receptor (ER). Using the subject as the unit of statistical analysis, either analysis of variance or chi-square analysis was employed to show the statistically significant difference at a level P <0.05. RESULTS: The pyogenic granuloma group expressed significantly more VEGF and bFGF than healthy gingiva and periodontitis. The positive staining of VEGF was mostly localized in the cytoplasm of macrophages and fibroblasts while that of bFGF was in the extracellular matrix of lamina propria. Angiostatin was expressed significantly less in pyogenic granuloma than the other 2 groups and was mostly localized in the nuclei of endothelial cells and epithelial cells. There was no significant difference in the expression of TSP-1 and ER among the 3 groups. CONCLUSIONS: The results of this research suggest that the etiology of pyogenic granuloma is due to the imbalance between angiogenesis enhancers and inhibitors. Whether and how the angiogenesis-associated factors are regulated by female steroid hormones remain to be answered. PMID- 10872950 TI - Proximal caries in juvenile periodontitis patients. AB - BACKGROUND: Caries is recognized as the prevalent proximal dental disease in adolescents, while proximal bone loss is minimal to non-existent in this population. Adolescents demonstrating an inverse disease pattern, i.e., minimal caries and active periodontitis, could provide powerful clues with regard to both diseases. However, data are inconsistent. This study was designed to clarify this relationship by comparing proximal caries prevalence in a juvenile periodontitis (JP) group to a matched non-periodontally diseased control group. METHODS: Two groups (cases [JPs] and control patients [CPs]) were matched for age, sex, and race and evaluated for decayed, missing, filled teeth and surfaces (DMFS) by radiographic analysis. Statistical analysis was performed by ANOVA and Student t test. The study consisted of four phases. Phase I was based on data from a previous study that failed to include race in the analysis. Thus, the original 23 JP patients (mostly African-Americans from New York City) were rematched for race as well as sex and age with CPs from Newark, NJ. The effect of water fluoridation (found in NYC) was evaluated in Phase II by matching the 23 original CPs (mostly Caucasian from NYC) with 23 CPs from NJ. Since differences were seen, we rematched our original JPs from NYC with a new set of race-matched CPs from NYC (Phase III). Finally, 13 JP patients from the University of Medicine and Dentistry of New Jersey (UMDNJ) were matched with CPs from NJ (Phase IV). RESULTS: Phase I and III indicated that JP patients had significantly less proximal caries than their matched CPs (P < or =0.05). Phase II confirmed the role of fluoride in caries reduction. Phase IV (NJ sample) supported our previous data and suggested that JP patients had less proximal caries than CPs (P < or =0.05). CONCLUSIONS: JP patients had significantly less proximal caries than their matched CPs when groups were balanced and radiographic evaluations were performed. In-depth studies of JP patients could provide important clues about both caries and periodontal disease etiology and pathogenesis. PMID- 10872951 TI - Effects of nicotine on the strength of attachment of gingival fibroblasts to glass and non-diseased human root surfaces. AB - BACKGROUND: The aim of this study was to evaluate the effect of nicotine on the strength of attachment of human gingival fibroblast cells to glass and non diseased human root surfaces. METHODS: Human gingival fibroblast cells (HGF) were trypsinized, suspended in RPMI 1640 medium, and incubated with autoclaved human root and glass sections and nicotine (NIC) concentrations of 0 (control), 25, 50, and 100 ng/ml for 1 week. HGF attached and grew on glass and root surfaces for 4 weeks at all NIC concentrations. HGF cultures were subjected to a rotary shaker machine for 30 minutes to test the strength of attachment of these cells at 100, 150, and 200 rpm. The root and glass sections were examined at 48 hours by light microscopy. RESULTS: Control groups exhibited a monolayer of long, spindle-shaped fibroblasts with a parallel alignment and minimal overlapping. With a concentration of NIC of 50 or 100 ng/ml as well as with increasing "speeds," the number of cells attached to these surfaces decreased dramatically. When 200 rpm was used for both groups at all NIC concentrations, very few HGF remained attached to these surfaces. CONCLUSIONS: This study showed that the nature of cell attachment to either glass or root surfaces is altered by nicotine, and marked detachment was noted when nicotine exposure was coupled with vigorous agitation at different rpm. Marked detachment noted in all specimens at 200 rpm indicates that this speed is excessive for use in subsequent experimentation. PMID- 10872952 TI - Genetic polymorphisms of the IL-1alpha and IL-1beta genes in African-American LJP patients and an African-American control population. AB - BACKGROUND: A functional polymorphism of the interleukin-1 beta (IL-1beta) gene has been proposed to be a risk factor for periodontitis. In adult forms of periodontitis, non-smokers of northern European heritage carrying the "2" allele of the IL-1alpha-889 and the IL-1beta +3953 RFLPs in either the heterozygous or the homozygous state at both loci were observed to have a greater risk for developing severe periodontitis. Studies of early-onset periodontitis (EOP) found that allele "1" of both IL-1alpha-889 and IL-1beta +3953 was transmitted more frequently with the EOP phenotype. The purpose of the present study was to determine the prevalence of the IL-1alpha and IL-1beta genotype polymorphisms in an African-American (AA) control population and in 37 African-Americans with localized juvenile periodontitis (LJP). METHODS: The IL-1alpha +4845 and IL-1beta +3953 loci were genotyped by PCR amplification, followed by restriction enzyme digestion and gel electrophoresis. The IL-1alpha +4845 locus, in linkage disequilibrium (>99%) with IL-1alpha-889, was genotyped because it is technically easier. Data were analyzed using r x c contingency tables. RESULTS: The IL-1beta +3953 allele "1" was carried by >99% of the AA control population and by 100% of the AA LJP group, with most individuals being homozygous 1,1. The prevalence of the composite genotype with at least one allele "2" at each of the IL-1beta +3953 and IL-1alpha +4845 loci was 14% (AA control group) and 8% (AA LJP group). CONCLUSIONS: Given the high frequency of the IL-1beta allele "1" in the African American population, it would appear that knowledge of this +3953 polymorphism would provide little diagnostic or predictive information for LJP. PMID- 10872953 TI - Bone probing measurement as a reliable evaluation of the bone level in periodontal defects. AB - BACKGROUND: Evaluation of periodontal regeneration is usually made by clinical measurements despite their limitations in determining the precise nature of the healing response. In the present study, the possibility of using bone probing measurements under local anesthesia to determine bone level changes without a re entry procedure was investigated. METHODS: Thirty-eight (38) first molars in 28 patients with chronic periodontitis who were scheduled to have periodontal surgery were included in this study. A custom-made acrylic resin stent was used for proper orientation of the probe for the bone probing depth measurement as well as probing depth measurements and surgical and radiographic bone level evaluations. The mesial, distal, and middle sites in the buccal aspect of each tooth were used. The sites were divided into 2 groups according to probing depth: those with a probing depth < 4 mm and those with a probing depth > or = 4 mm. RESULTS: The probing depth was not significant in the difference between actual bone level (SBL) and bone probing depth (BP) (P >0.05). The greatest correlation to SBL was found with BP (gamma = 0.92), followed by radiographic bone level (RBL) (gamma = 0.69). The morphology of the defects had no significant effect on the difference between SBL and other measurements, while tooth surface and probing depth had significant effects on the difference between RBL and SBL. CONCLUSIONS: The results of this study suggest that, regardless of probing depth, probing surface, and the presence of intrabony defects, there is a minimal difference between the BP and SBL. Determining the bone probing depth measurement is a kind of reliable method to estimate the regenerated bone level following periodontal treatment. PMID- 10872954 TI - Non-surgical periodontal therapy and tooth loss. A cohort study. AB - BACKGROUND: No reliable evidence is available regarding the effect of periodontal therapy on major disease endpoints such as tooth loss, edentulism, or quality of life. The primary objective of this study was to assess the association between tooth loss and the non-surgical periodontal treatment history of 1,021 members of the Kaiser Permanente Dental Care Program. METHODS: Tooth loss rates were estimated using Poisson regression models, adjusting for some of the potentially confounding variables such as initial disease severity and extent. RESULTS: Continuous non-surgical therapy (one or more non-surgical procedures performed during 3 successive years), as opposed to no therapy during such a 3-year period, reduced the subsequent tooth mortality rate by 58% (relative rate, 0.42; 95% confidence interval, 0.29-0.61). Intermittent non-surgical therapy reduced the tooth mortality rate by 48% (RR = 0.52; 95% confidence interval, 0.34-0.80). As the number of non-surgical procedures increased, tooth loss rates decreased. CONCLUSIONS: These findings suggest that non-surgical periodontal therapy may be associated with a substantial reduction in tooth mortality. Different study designs and populations are needed to confirm these findings. PMID- 10872955 TI - Smoking-attributable periodontitis in the United States: findings from NHANES III. National Health and Nutrition Examination Survey. AB - BACKGROUND: The principal objectives of this study were to examine the relationship between cigarette smoking and periodontitis and to estimate the proportion of periodontitis in the United States adult population that is attributable to cigarette smoking. METHODS: Data were derived from the Third National Health and Nutrition Examination Survey, a nationally representative multipurpose health survey conducted in 1988 to 1994. Participants were interviewed about tobacco use and examined by dentists trained to use standardized clinical criteria. Analysis was limited to dentate persons aged > or =18 years with complete clinical periodontal data and information on tobacco use and important covariates (n = 12,329). Data were weighted to provide U.S. national estimates, and analyses accounted for the complex sample design. We defined periodontitis as the presence of > or =1 site with clinical periodontal attachment level > or =4 mm apical to the cemento-enamel junction and probing depth > or =4 mm. Current cigarette smokers were those who had smoked > or =100 cigarettes over their lifetime and smoked at the time of the interview; former smokers had smoked > or =100 cigarettes but did not currently smoke; and never smokers had not smoked > or =100 cigarettes in their lifetime. RESULTS: We found that 27.9% (95% confidence interval [CI]: +/-1.8%) of dentate adults were current smokers and 23.3% (95% CI: +/-1.2%) were former smokers. Overall, 9.2% (95% CI: +/-1.4%) of dentate adults met our case definition for periodontitis, which projects to about 15 million cases of periodontitis among U.S. adults. Modeling with multiple logistic regression revealed that current smokers were about 4 times as likely as persons who had never smoked to have periodontitis (prevalence odds ratio [ORp] = 3.97; 95% CI, 3.20-4.93), after adjusting for age, gender, race/ethnicity, education, and income:poverty ratio. Former smokers were more likely than persons who had never smoked to have periodontitis (ORp = 1.68; 95% CI, 1.31-2.17). Among current smokers, there was a dose-response relationship between cigarettes smoked per day and the odds of periodontitis (P <0.000001), ranging from ORp = 2.79 (95% CI, 1.90-4.10) for < or =9 cigarettes per day to ORp = 5.88 (95% CI, 4.03-8.58) for > or =31 cigarettes per day. Among former smokers, the odds of periodontitis declined with the number of years since quitting, from ORp = 3.22 (95% CI, 2.18-4.76) for 0 to 2 years to ORp = 1.15 (95% CI, 0.83-1.60) for > or =11 years. Applying standard epidemiologic formulas for the attributable fraction for the population, we calculated that 41.9% of periodontitis cases (6.4 million cases) in the U.S. adult population were attributable to current cigarette smoking and 10.9% (1.7 million cases) to former smoking. Among current smokers, 74.8% of their periodontitis was attributable to smoking. CONCLUSIONS: Based on findings from this study and numerous other reports, we conclude that smoking is a major risk factor for periodontitis and may be responsible for more than half of periodontitis cases among adults in the United States. A large proportion of adult periodontitis may be preventable through prevention and cessation of cigarette smoking. PMID- 10872956 TI - Histologic evaluation of periodontal healing in humans following regenerative therapy with enamel matrix derivative. A 10-case series. AB - BACKGROUND: Enamel matrix derivative (EMD) has been developed as a stimulus of periodontal regeneration. Human histology following its use has not been evaluated on pathologically altered root surfaces. METHODS: Ten intrabony defects in 8 patients were evaluated at 2 centers. Teeth with advanced adult periodontitis that were treatment planned for extraction were treated with sulcular incisions; full-thickness flap reflection; debridement of granulomatous tissue from the defect; placement of a notch in the root at the apical extent of calculus; mechanical root planing; conditioning with citric acid; application of EMD; wound closure with sutures; and placement of a periodontal dressing. Biweekly to monthly recalls were made until removal of small block section biopsies at about 6 months. The biopsies were fixed, decalcified, step-serial sectioned at 6 microns to 8 microns, and stained with hematoxylin and eosin or Masson's trichrome. RESULTS: Histologic evaluation of the region coronal to the base of the calculus notch showed evidence of regeneration (new cementum, new bone, and new periodontal ligament) in 3 specimens, new attachment (connective tissue attachment/adhesion only) in 3 specimens, and a long junctional epithelium in 4 specimens. No evidence of root resorption, ankylosis, or untoward inflammation was seen. CONCLUSIONS: The results of this study fulfill the proof of principle that use of EMD can result in periodontal regeneration on previously diseased root surfaces in humans, but on an inconsistent basis. PMID- 10872957 TI - A rapid DNA probe method for detection of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans. AB - BACKGROUND: The objective of the present study was to develop a rapid DNA probe method for the microbiological detection of periodontitis that can be used in dental clinics. By using the DNA probe, we also investigated the correlation between the occurrence of putative periodontopathic bacteria and clinical parameters. METHODS: This rapid DNA probe method minimizes the use of a water bath for ordinary hybridization and washing in order to shorten the total reaction time. The detection process could be completed within 2 hours. In order to evaluate the clinical application of the DNA probe, subgingival plaque samples were taken from patients with periodontitis before initial therapy. After the therapy, the patients were microbiologically and clinically evaluated. RESULTS: When the DNA probe method was compared with the culture method, the agreement was 88% for Porphyromonas gingivalis and 67% for Actinobacillus actinomycetemcomitans. A statistically significant association was found between the detection of P. gingivalis and probing depth, bleeding on probing (chi2 test: P <0.001, P <0.05). A significant association was also shown between the detection of A. actinomycetemcomitans and probing depth in patients aged 35 or older (chi2 test: P <0.001). The detection rate of A. actinomycetemcomitans was highest in teenagers. At shallow periodontal pocket sites (PD < or =3 mm) in teenagers, no P. gingivalis was found, while 22% of the sites harbored A. actinomycetemcomitans. After the therapy, the frequency of detection of P. gingivalis decreased significantly only in the clinically improved sites (chi2 test: P <0.001). CONCLUSIONS: The rapid DNA probe method appears promising as an efficient tool for rapid clinical detection of periodontopathic bacteria. PMID- 10872958 TI - The effects of sustained release doxycycline on the anaerobic flora and antibiotic-resistant patterns in subgingival plaque and saliva. AB - BACKGROUND: The purpose of this study was to determine the effects of periodontal treatment with a sustained-release, biodegradable gel containing 8.5% doxycycline on the anaerobic flora and on antibiotic susceptibility patterns associated with subgingival plaque and saliva. METHODS: Forty-five subjects with adult periodontitis were entered into a parallel design, single-blind study of 6 months' duration. The subjects were randomized to receive either doxycycline treatment (n = 23) or oral hygiene instruction/reinforcement (n = 22). Saliva and subgingival plaque samples were collected prior to and at 7, 21, 91, and 182 days after initiation of treatment. The proportion of the cultivable flora resistant to 10 microg doxycycline/ml was determined relative to total anaerobic counts, and the 3 most predominant colony types resistant to doxycycline were individually enumerated. A representative of each was subcultured, identified to genus and species level, and tested for its susceptibilities to 6 antibiotics. RESULTS: A significant decrease (P <0.01) in total anaerobic counts following doxycycline treatment caused a transient increase in the proportion, but not in the actual counts, of doxycycline-resistant bacteria recovered from both plaque and saliva at 7 and 21 days but not at 91 or 182 days. The same doxycycline resistant taxa were recovered at all sample periods including baseline. Regardless of treatment, the isolates were similarly distributed and belonged to the same bacterial groups. CONCLUSIONS: Doxycycline treatment significantly reduced the anaerobic population in plaque but did not result in a change in either the number of resistant bacteria present or the acquisition of antibiotic resistance. PMID- 10872959 TI - Regeneration of Class III furcation defects with basic fibroblast growth factor (b-FGF) associated with GTR. A descriptive and histometric study in dogs. AB - BACKGROUND: The poor predictability of periodontal regenerative treatment of Class III furcation defects stimulates the study of alternatives to improve its results, such as the use of polypeptide growth factors. The objective of this study was to evaluate, both histologically and histometrically, the effects of topical application of basic fibroblast growth factor (b-FGF) associated with guided tissue regeneration (GTR) in the treatment of Class III defects surgically induced in dogs. METHODS: All second and fourth premolars of 5 mongrel dogs were used and randomly assigned to one of three treatment groups: group 1 (control), treated with scaling and root planing, tetracycline hydrochloride (125 mg/ml) conditioning, and GTR with a collagen membrane; group 2, same treatment as group 1 plus 0.5 mg of b-FGF; group 3, same treatment as group 1 plus 1.0 mg of b-FGF. After a 90-day healing period, routine histologic processing and staining with hematoxylin and eosin and Masson trichrome were performed. RESULTS: The descriptive analysis indicated better regenerative results in both groups treated with b-FGF while the histometric data, analyzed by means of analysis of variance (ANOVA), showed greater filling of the defects in group 2 in comparison to the defects in groups 3 and 1, respectively, which was represented by a smaller area of plaque-occupied space (P = 0.004) as well as a greater amount of newly formed cementum (P = 0.002). CONCLUSIONS: These results indicate that b-FGF, especially in smaller doses, may enhance the regenerative results in Class III furcation lesions, leading to greater filling of these defects with both mineralized and non-mineralized tissues. PMID- 10872960 TI - An evaluation of the efficacy of a curved bristle and conventional toothbrush. A comparative clinical study. AB - BACKGROUND: The aim of this study was to determine the plaque-removing ability of a curved bristle toothbrush compared to a conventional, straight bristle, manual toothbrush. METHODS: The study group consisted of 100 volunteers 16 to 24 years of age from a professional engineering college. A four-week post-prophylaxis, parallel, longitudinal, double-blind clinical study was conducted; all volunteers were instructed in specific oral hygiene techniques. Plaque was assessed at baseline and at the end of 1, 2, 3, and 4 weeks using the Quigley-Hein plaque index after disclosing with erythrosin red. Gingival status was assessed at baseline and at the end of 1, 2, 3 and 4 weeks by using the gingival index of Loe and Silness. RESULTS: Comparative assessment showed a mean of 2.11 +/- 0.086 mm for group 1 and 2.37 +/- 0.216 mm for group 2, indicating a significant difference between the plaque-removing efficacy of the curved bristle and straight bristle toothbrush. CONCLUSIONS: The curved bristle toothbrush was significantly more effective in removing plaque overall than the conventional toothbrush. PMID- 10872961 TI - A histopathological investigation on the effects of the bisphosphonate alendronate on resorptive phase following mucoperiosteal flap surgery in the mandible of rats. AB - BACKGROUND: The present study was designed to examine histopathologically whether local delivery of aminobisphosphonate (alendronate) could be effective in preventing the alveolar bone resorption associated with mucoperiosteal flaps. METHODS: Following mucoperiosteal flap elevation in the molar region of the rat mandible, a surgical pellet soaked with aminobisphosphonate was locally applied on the exposed bone surface and covered by flap. The determined parameters with a semi-quantitative subjective method for the histopathological evaluation were as follows: existing inflammatory cell infiltration of the related periodontal tissue; fibrotic component content and bundles of collagen fibers; the number and morphology of osteoclasts of the alveolar bone and interdental septum; existing resorption lacunae (osteoclast surfaces); and existing osteoblastic activity (forming surfaces). RESULTS: The results showed that while there were no detectable statistically significant differences between the saline and alendronate-treated groups on the existing inflammatory cell infiltration (ICI), number of osteoclasts, and osteoblastic activity, the results for the fibrotic and collagen component, osteoclast morphologies, and existing resorption lacunae were statistically significant. CONCLUSIONS: These results suggest that local application of the aminobisphosphonate alendronate can be used as an adjunct in therapy for reducing bone resorption following surgery. It can also be suggested for consideration that, even for the surgical approaches in dentistry where bone graft materials and/or dental implants are needed, using bisphosphonate may achieve a new dimension in periodontal therapy in the near future. PMID- 10872962 TI - Defective calcium influx factor activity in neutrophils from patients with localized juvenile periodontitis. AB - BACKGROUND: Localized juvenile periodontitis (LJP) is an early-onset periodontal disease associated with neutrophil dysfunction, including defective chemotaxis, reduced protein kinase C (PKC) activity, and reduced calcium entry. These observations are important because reduced availability of cytosolic-free calcium concentration in the cell will have detrimental consequences for the numerous cytosolic calcium concentration-dependent pathways. In particular, there is a direct relationship between Ca2+ flux and the cell activation enzyme PKC. In this report, we focused on the mechanism of calcium entry, investigating a newly described molecule, calcium influx factor (CIF). CIF is thought to be a second messenger for the opening of membrane calcium channels when intracellular calcium stores are depleted. We examined CIF activity in neutrophils from normal subjects and LJP patients. METHODS: Neutrophils from 11 LJP patients, 3 adult periodontitis (AP) patients, and 12 normal subjects were isolated from peripheral venous blood. CIF was extracted with thapsigargin, a Ca2+-ATPase inhibitor, from isolated neutrophils and CIF activity measured using a 45CaCl2 uptake assay. RESULTS: The CIF activity in neutrophils from LJP patients ranged from 98.9 to 281.5 units/mg protein (mean = 180.2 +/- 56.3) and from 291.9 to 755.5 units/mg protein (mean = 528.8 +/- 153.8) in non-periodontal disease controls. CIF activity in AP patients was also measured and found to be similar to controls. The CIF activity in LJP patients was statistically significantly reduced compared to that in normal subjects (P <0.001). CONCLUSIONS: This study suggests that CIF activity may be an important determinant in neutrophil abnormalities in LJP. PMID- 10872963 TI - Pluronic polyol effects on human gingival fibroblast attachment and growth. AB - BACKGROUND: Enhanced speed of human gingival fibroblast (HGF) spreading and attachment, as affected by ionic bonding interactions, may facilitate cell orientation and subsequent collagen synthesis to promote early wound healing. The purpose of this study was to determine the in vitro effects of pluronic polyols, a family of widely used surfactants currently used as drug carriers for antibiotic, anti-inflammatory, and anti-neoplastic agents, on the attachment and growth of human gingival fibroblasts (HGF) to dentin and plastic surfaces using established tissue culture techniques. METHODS: Plastic culture wells containing Eagle's minimal essential media (EMEM) with 10% fetal calf serum and Pluronic F 68 or F-127 in concentrations from 1.2 x 10(-2) to 1.2 x 10(-10) M were incubated with HGF and run in replicates of ten. Attached cells were quantified by measuring the optical density of methylene blue-stained cells. Additional experiments were conducted using human dentin sections as a substrate and Pluronic F-68 or F-127 at a concentration of 1.2 x 10(-8) M. In these experiments, HGF were stained with acridine orange and quantified per unit area of dentin by fluorescence microscopy. RESULTS: Attachment and growth of HGF to both plastic and dentin were significantly increased over serum controls by very low concentrations of Pluronic F-68 and F-127 by 30 minutes, with attachment reaching a plateau at 2 hours. CONCLUSIONS: Pluronic polyols, a family of widely used surfactants, in very low dosages may be beneficial in early postsurgical wound healing by facilitating early attachment and enhancing the growth rate of human gingival fibroblasts. PMID- 10872964 TI - Effects of diode and Nd:YAG laser irradiation on titanium discs: a scanning electron microscope examination. AB - BACKGROUND: Dental lasers have been recommended for uncovering submerged implants as well as decontaminating implant surfaces when treating peri-implantitis. The aim of this study was to show the possible alterations in titanium disc surfaces using an Nd:YAG or a diode laser. METHODS: Three different titanium discs were used (sandblasted, titanium plasma-sprayed [TPS], and hydroxyapatite [HA] coated) to determine the effects of laser irradiation on these surfaces using a scanning electron microscope (SEM). The discs were either irradiated with a pulsed Nd:YAG laser with a contact handpiece and power settings of 2.0, 4.0, and 6.0 W or with a diode laser at 5.0, 10.0, and 15.0 W power settings and continuous wave (cw) in the contact handpiece. Irradiated areas were compared with control titanium sites which were not lased. The specimens were prepared for SEM examination after the disc irradiation. RESULTS: The SEM examination demonstrated extensive melting in all of the Nd:YAG laser irradiated areas. Damage was seen in all TPS- and HA coated discs even at the lowest power setting. Loss of porosity, coating microfractures, and a relatively smooth surface were observed. In contrast, the diode laser did not cause any damage or modify the disc surface. Regardless of the power setting, there was no visible difference between lased and non-lased surfaces after cw irradiation with the diode laser. CONCLUSIONS: From these findings, it was concluded that the diode laser (980 nm) does not damage titanium surfaces, which should be of value when uncovering submerged implants and treating peri-implantitis. PMID- 10872965 TI - Features of severe periodontal disease in a teenager with Chediak-Higashi syndrome. AB - BACKGROUND: Chediak-Higashi syndrome (C-HS) is a rare congenital disease characterized by defective neutrophil function with abnormal lysosomal inclusions, neutropenia, and reduced chemotaxis. The complete syndrome includes oculocutaneous albinism with photophobia, neurologic features, recurrent infections, and enterocolitis. METHODS: A 14-year-old male C-HS patient was referred to us because of serious periodontal destruction with acute inflamed gingiva and ulcers. Clinical and biological investigations were performed, leading to the diagnosis of C-HS. RESULTS: Laboratory findings included neutropenia and hypergammaglobulinemia. Peripheral blood smears showed giant granules in neutrophils, eosinophils, and granulocytes. Bone marrow smears showed giant inclusions in leukocyte precursor cells. These granules and inclusions were characteristic of Chediak-Higashi syndrome. Oral radiographic status showed extensive loss of alveolar bone leading, in most cases, to tooth exfoliation. Bacteria often associated with periodontitis were detected in subgingival plaque samples, including Fusobacterium nucleatum, Campylobacter rectus, Prevotella melaninogenica, Peptostreptococcus anaerobius, and Clostridium sp. Biopsies of periodontal tissues for light and electronic microscopic examinations revealed massive bacterial invasion of the epithelial tissue, epithelial cells, and connective tissue. Ultrastructural observations of periodontal polymorphonuclear leukocytes showed defective granulation, with abnormal granules not discharging their lysosomal content against engulfed bacteria. Viable dividing bacteria were found in the cytoplasm. CONCLUSIONS: In this case, early-onset periodontitis seems to be the expression of C-HS granulocyte deficiency. Periodontal treatment of these patients is often unsuccessful. This case report illustrates the importance of the dentist in initiating clinical and biological investigations in such early aggressive periodontitis in young patients. PMID- 10872966 TI - Case report of regional alveolar bone actinomycosis: a juvenile periodontitis like lesion. AB - BACKGROUND: Cervicofacial actinomycosis infection most often involves the mandibular bone and rarely the alveolar crest. METHODS: We describe a 14-year-old patient who had actinomycosis involving the alveolar bone at the left lower dental quadrant region. Resembling juvenile periodontitis, it was difficult to diagnose properly and resulted in devastating dental and periodontal consequences: loss of one tooth with most of its adjacent regional alveolar bone, severely compromising the support of two other teeth. RESULTS: With the diagnosis came successful treatment, including surgical removal of the soft and hard tissues with concomitant prolonged penicillin administration. CONCLUSIONS: We feel that this case should raise the interest and concern of both the periodontist and the general practitioner so that early diagnosis can be obtained, significantly improving the clinical outcome. PMID- 10872967 TI - Gingival peripheral odontoma in an adult: case report. AB - BACKGROUND: Odontoma arising in the extraosseous soft tissue is extremely uncommon. We describe our experience of gingival peripheral odontoma in which the initial presentation was a small asymptomatic nodule. METHODS: Case study. RESULTS: A 44-year-old man reported with a firm gingival mass of the anterior maxilla which had been gradually enlarging over 5 years. Radiographic examination showed a dense radiopaque mass occupying most of the tumor and no evidence of underlying intraosseous lesion. The pathology was reported as odontoma. CONCLUSIONS: This is the fourth reported case of peripheral odontoma in the gingiva. PMID- 10872968 TI - Immediate loading of implants in partially and fully edentulous jaws: a series of 27 case reports. AB - BACKGROUND: The concept of immediate loading of dental implants has been researched in animal and man. High predictability with titanium screw implants can be expected under certain circumstances. Delayed loading, which is empirically based, may offer roadblocks to success in the complex case. Micromotion under a denture, retention of compromised teeth, and the necessity for multiple implant surgeries complicate the situation. The present study was undertaken to demonstrate whether in partial and fully edentulous patients, titanium screw implants may be installed and loaded within 72 hours. METHODS: The study included 27 jaws (23 mandibles and 4 maxillas) in patients who refused to wear a denture or were told of the possibility of immediately loading their implants. Criteria similar to delayed loading was utilized. The patient had to have adequate volume and density for a minimum of 4 (10 mm) implants in the mandible and 6 in the maxilla, in addition to other requirements. After a thorough presurgical evaluation by the restorative dentist, a template and heat cured/metal-reinforced provisional were fabricated. Following fixture installation, either the fixed provisional was placed or an impression was taken, and the provisional was seated within 72 hours. Once the temporary was placed, it remained until osseointegration was complete. RESULTS: Success rates similar to those in delayed loaded cases may be expected (95%). Fewer sandblasted/acid etched titanium screws were lost than machined titanium screws. All patients went on to completion of their original prosthetic prescription. CONCLUSIONS: Patients who are partially or fully edentulous may predictably be restored with fixed implant prostheses immediately upon fixture placement if certain parameters are met. PMID- 10872969 TI - The clinical effectiveness of implants placed immediately into fresh extraction sites of molar teeth. AB - BACKGROUND: Studies concerning immediate implantation describe its use in the anterior and premolar regions. However, its clinical effectiveness in immediately replacing molar teeth has rarely been challenged. The purpose of this study was to evaluate the survival rate of implants placed immediately after extraction of molar teeth to support a fixed ceramo-metal prosthesis. METHODS: From 1989 to 1996, 56 immediate implants were placed in 43 patients following extraction of 51 molars; 46 molars were replaced by 1 implant and 5 molars replaced by 2 implants. All implants were restored with fixed prostheses (4 single crowns and 52 splinted). Mean follow-up period was 15 months (range, 4 to 60 months). The influence of the following parameters on implant failure was evaluated: gender, arch, smoking, pre-extraction vertical bone loss, implant length, and severity of complications between the two stages of surgery. RESULTS: The 5-year cumulative survival rate (5-year CSR) was 89%. The 5-year CSR among men was 84% compared to 93.5% among women. The maxillary 5-year CSR was 82% and the mandibular 92%. Among non-smokers (50 implants), the 5-year CSR was 90% compared to 83% among smokers (6 implants). Complications were evident in 8 (6 minor, 2 major) out of 50 non failing implants compared to 2 (minor) of the 6 failing implants. No differences were evident in the other study variables. CONCLUSIONS: Immediate implantation in the molar region is an alternative, predictable surgical treatment. Immediate implantation in the posterior mandible has a better prognosis than in the posterior maxilla. PMID- 10872970 TI - Tooth loss significantly reduced with non-surgical therapy. PMID- 10872971 TI - Laparoscopic fundoplication for gastroesophageal reflux: effects on esophageal motility. AB - Laparoscopic Nissen-Rossetti fundoplication is now recognized as a valid therapy for the treatment of gastroesophageal reflux disease. This retrospective study evaluates the effects of laparoscopic fundoplication on esophageal motility and correlates these effects to postsurgical symptoms. A total of 123 patients underwent laparoscopic fundoplication at our institution. Pre- and postoperative esophageal manometric data were analyzed with regard to the effect of surgery and postsurgical outcome. Postoperative lower esophageal sphincter pressure was significantly increased compared wtih preoperative values (1.7 +/- 0.8 kPa vs 0.9 +/- 0.7 kPa). Duration and amplitude of esophageal body contractions were not modified. The percentage of deglutition-induced complete peristaltic waves and the velocity of propagation were significantly decreased after surgery (P < 0.05). Postoperative symptoms were significantly correlated with postoperative lower esophageal sphincter pressure only. Laparoscopic fundoplication significantly increases lower esophageal sphincter pressure. It significantly decreases esophageal body peristaltic efficiency, a decrease that is most likely of minor clinical significance. PMID- 10872972 TI - The laparoscopic finding of pericholedochitis at cholecystectomy predicts the presence of unsuspected bile duct stones. AB - Routine laparoscopic cholangiography and sonography have been recommended for identification of unsuspected bile duct stones in laparoscopic cholecystectomy. The increased prevalence of retained stones seems, however, to confirm that cholangiography has been used rather selectively. The aim of this study was to investigate the kind and extent of a possible correlation between inflammatory changes in the bile duct mucosa and the hepatoduodenal ligament in patients with and without unsuspected bile duct stones. Sixty-eight patients, without symptoms or signs of bile duct stones in their case histories, laboratory findings, and preoperative sonography results, underwent laparoscopic cholecystectomy for symptomatic gallstone disease and diagnostic transcystic cholangiography and cholangioscopy. The initial step at laparoscopy revealed edema of the periductal loose tissue, vascular dilation, and petechiae on the external surface of the distal portion of the extrahepatic bile duct in 47 of 68 patients. Cholangioscopy revealed inflammatory changes in the mucosa of the distal and sphincteric portions of the extrahepatic bile duct in 45 of 47 patients, and unsuspected ductal stones were identified in 41 of these 45 patients. Neither the external surface of the bile duct nor its mucosa exhibited any signs of inflammation in the 21 remaining patients (controls), all of whom showed stone-free bile ducts. Inflammatory changes in the bile duct mucosa, occurring along with unsuspected mobile ductal stones in this study, were reflected on the external surface of the bile duct with a specificity of 87%. The changes in the external surface of the bile duct can be recognized at the initial inspection in laparoscopic cholecystectomy, as in this study, and indicate that more accurate diagnostic procedures to identify unsuspected ductal stones should be used. PMID- 10872973 TI - Use of a cauterizing laparoscopic linear stapler in intestinal anastomosis. AB - The purpose of this study was to assess the usefulness of a cauterizing laparoscopic linear stapler for intestinal anastomosis. In a porcine model, intestinal anastomoses performed with a standard laparoscopic linear stapler, a cauterizing laparoscopic linear stapler (RF stapler), and a two-layer, hand-sewn technique were compared by measuring bursting pressures at 4 and 7 days after surgery. During surgery, the RF stapler provided better hemostasis than the regular stapler for mesenteric transection. At 4 days, one leak occurred in the RF stapler group, and the bursting pressure in the RF stapler group was significantly lower than the bursting pressures in the regular stapler group and the hand-sewn group. In addition, the bursting pressure was significantly greater in the hand-sewn group than in the regular stapler group at 4 days. By 7 days, there were no differences in bursting pressure among the groups. We recommend that the RF stapler not be used for intestinal anastomosis. However, the device may be beneficial for controlling vasculature. PMID- 10872975 TI - Practice parameters for sigmoid diverticulitis. The Standards Task Force, American Society of Colon and Rectal Surgeons PMID- 10872974 TI - Two-stage laparoscopic management of generalized peritonitis due to perforated sigmoid diverticula: eighteen cases. AB - The classic treatment of generalized peritonitis due to perforation of sigmoid diverticula is based on the principle of a two-stage surgery with a temporary derivation of the colonic transit. This procedure is associated with a prohibitively high immediate and delayed morbidity, especially associated with the abdominal wound. The laparoscopic approach to this complication is less aggressive and allows a second-stage elective laparoscopic resection. Eighteen consecutive patients (ten women and eight men; average age, 53.7 years) underwent emergency laparoscopic treatment for generalized peritonitis due to perforated diverticula. Eight of these patients had previously had diverticulitis attacks. By peritoneal cavity exploration and full peritoneal lavage (average, 15 L), the infected sigmoid lesion was stuck with biologic glue. A drain was inserted at the site of the lesion and in some cases also in other abdominal zones. No colostomy was necessary. Antibiotic treatment was started at diagnosis and continued for a minimum of 7 days. There was no mortality. Morbidity was limited to three patients (two cases of lymphangitis and one of pulmonary disease). No patient had a wound abscess or residual deep collections. The mean hospitalization was 8 days. Fourteen patients underwent elective laparoscopic sigmoid resection with a delay of 3.5 months. One conversion to laparotomy was necessary. The laparoscopic treatment of generalized peritonitis due to perforated sigmoid diverticula is an interesting alternative to the traditional treatment. It is associated with a lower morbidity, a shorter postoperative hospital stay, and an improvement in the patient's quality of life, because colostomy is avoided. It is also associated with economic savings. PMID- 10872976 TI - Laparoscopic hernia repair: the learning curve. AB - The performance of a laparoscopic inguinal hernia repair requires unique technical and cognitive skills which, until recently, were not routinely taught to general surgeons. The initial experience of three surgeons with laparoscopic hernia repair was audited prospectively to assess the learning curve for the technique. From March 1992 to June 1994, transabdominal preperitoneal (TAP) mesh repair was attempted on 172 consecutive inguinal hernias. Three procedures were converted to traditional repairs. The three independent surgeons that performed the repairs had minimal or no prior clinical experience with the technique in the role as primary surgeon. The hernia repairs were divided into two groups. Group 1 consisted of the first 90 hernia repairs in the series, 30 repairs per surgeon. This group was compared to the subsequent 82 repairs (group 2), approximately 27 repairs per surgeon. Patients were followed up for a median of 31 months. Group 1 had more patients who were hospitalized overnight (37% versus 31%), a greater rate of conversion (2.2% versus 1.2%), a higher complication rate (11.7% versus 0%), a higher recurrence rate (12.2% versus 0%), and a longer delay in the return to full activity (11 weeks versus 8 weeks). Also, overall patient satisfaction with their hernia repair was slightly greater in group 2 (score, 9.0/10 versus 8.2/10). The lack of prior experience with the TAP technique (one surgeon) was associated with a marked increase in the number of conversions (two of three total conversions), complications (four of eight total), and hernia recurrences (8 of 11 total). This study demonstrates that a surgeon's initial experience with laparoscopic herniorrhaphy is associated with an identifiable learning curve. Significant improvements in complication and recurrence rates and overall patient satisfaction can be expected after the initial learning phase. Also, a complete lack of prior experience with laparoscopic herniorrhaphy is associated with a higher rate of conversion and significant increases in complications and hernia recurrences. PMID- 10872977 TI - Variations of the cystic artery in Chinese adults. AB - The origin and course of the cystic artery related to the Calot triangle were studied in 72 autopsies. The cystic artery arises from many possible origins; the right hepatic artery is the most common origin (76.6%). The Calot triangle (hepatocystic triangle), which is an important imaginary referent area for biliary surgery, is bounded by the common hepatic duct (CHD), the cystic duct, and the cystic artery. Of all the cystic arteries, 86.1% coursed through the Calot triangle, and 100% of the cystic arteries originating from the right hepatic artery coursed through the Calot triangle. However, only 54% of the cystic arteries that originated from the left, bifurcation, proper, and common hepatic arteries ran through the triangle. None of the cystic arteries that originated from the gastroduodenal, celiac, superior mesentery, or superior pancreaticoduodenal arteries passed through the triangle. Furthermore, 72.7% of the cystic arteries that originated from the right hepatic artery ran beneath the CHD as they entered the Calot triangle; the others ran anterior to the CHD. Of the cystic arteries that arose from locations other than the right hepatic artery, 29.4% ran posterior to the CHD, and 11.8% ran anterior to the CHD. The current study provides detailed information about anatomic variance in Chinese adults that may help avoid injury during open or laparoscopic cholecystectomies. PMID- 10872978 TI - Femoral venous flow during laparoscopic gynecologic surgery. AB - The lower-limb venous return, assessed by the peak systolic venous velocities (PSVV) of the left common femoral vein, was recorded at different stages of operation for five patients undergoing major gynecologic operative laparoscopy. The average baseline PSVV was 23.1 cm/s. After positioning the patient in the Trendelenburg position, the PSVV increased to an average of 31.5 cm/s; this was a statistically significant increase. Creation of the pneumoperitoneum changed the waveform from a normal phasic pattern to a dampened, continuous, monophasic waveform. The average PSVV was reduced to 15.9 cm/s; this dampening was statistically significant. Further dampening was evident 1 hour intraoperatively, and the flow became intermittent, with cycles of dampened flow followed by periods of absent flow; these changes in PSVV were not statistically significant. Calf compressors did not increase the femoral PSVV at the beginning of operation, nor at I hour intraoperatively; the decrease was not statistically significant. After release of the pneumoperitoneum, the baseline waveform pattern and velocity returned. The Trendelenburg position used for gynecologic operative laparoscopy was associated with a statistically significant increase in the lower-limb PSVV. This increase did not fully counteract the dampening effect of a pneumoperitoneum on lower-limb PSVV. The authors' study did not support the benefit previously reported on the use of pneumatic calf compressors. The authors therefore recommend continuing the practice of antithrombotic measures for patients undergoing gynecologic operative laparoscopy. PMID- 10872979 TI - A comparative experimental study evaluating the performance of surgical robots aesop and endosista. AB - The aim of this study was to experimentally assess and compare the accuracy of the surgical robots Aesop and Endosista as camera holders for use in laparoscopic surgery. The performance of these two robotic systems was examined for linear (upwards, downwards, diagonal), complex, and "in and out" movements using laparoscopic training boxes. Standard distances and tests were used for each system, and the time required to achieve each task was measured. The majority of the linear movements of the verbal and preprogrammed modes of Aesop were quicker than those of Endosista. Diagonal movements were significantly faster with the preprogrammed-mode Aesop. Complex or three-dimensional movements were also significantly faster with both modes of Aesop than with Endosista. Under the experimental conditions, Aesop, particularly in the preprogrammed mode, is quicker and more accurate than Endosista. PMID- 10872980 TI - Ultrasound-guided laparoscopic resection of pancreatic islet cell tumors. AB - Pancreatic islet cell tumors represent a diverse group of neuroendocrine lesions. These tumors may be singular or multiple, benign or malignant, sporadic, or part of the constellation of multiple endocrine neoplasia type 1. Tumors such as insulinomas and gastrinomas produce gastrointestinal peptides that lead to diagnosis. Nonfunctioning lesions may be found incidentally or by screening patients at high risk for such tumors. Successful management of patients with pancreatic islet cell tumors relies on accurate localization and sound operative technique. With proper preoperative localization, advanced laparoscopic methods can be used to manage patients with these pancreatic neoplasms. Preoperative localization of pancreatic islet cell tumors was difficult in the past. Standard imaging and localizing modalities, such as computed tomography scanning, magnetic resonance imaging, angiography, transabdominal sonography, and portal venous sampling, yield only 24% to 75% accuracy. Consequently, many biochemically suspected lesions cannot be imaged with current techniques. Decreased tactile sensation of laparoscopy adds complexity to intraoperative identification. Endoscopic sonography and laparoscopic sonography provide accurate preoperative and intraoperative localization to enhance laparoscopic and open resection. The authors treated two patients with islet cell neoplasms using endoscopic sonography to preoperatively visualize the tumors and laparoscopic sonography to guide laparoscopic enucleation. Their approach and difficulties are discussed. PMID- 10872981 TI - Alternative technique of laparoscopic hepaticojejunostomy for advanced pancreatic head cancer. AB - Only 20% of patients with pancreatic cancer can undergo curative resection. Therefore, palliative treatment of pancreatic cancer assumes the utmost clinical importance. The aim of the palliative treatment of pancreatic head carcinoma is to relieve the jaundice and/or duodenal obstruction. Endoscopic or transparietal decompression of the obstructed bile duct can be accomplished in most cases, but the durability of these techniques is not as great as that of a surgically created bypass. On the other hand, hepaticojejunostomy carries higher morbidity and mortality rates than the former nonsurgical methods. In order to promote long lasting palliation with low morbidity and mortality rates, minimally invasive techniques of biliary and gastric bypass have been described. However, laparoscopic Roux-en-Y hepaticojejunostomy seems to be a complex surgical procedure. With an aim to simplify the construction of a laparoscopic hepaticojejunostomy, the authors suggest an alternative technique. PMID- 10872982 TI - Liver hematoma after laparoscopic nissen fundoplication: a case report and review of retraction injuries. AB - Laparoscopic fundoplication is a safe and effective alternative to long-term medical therapy in select patients with gastroesophageal reflux disease. Among the technical challenges of laparoscopic fundoplication, retraction of the left lobe of liver can cause significant morbidity. Intraoperative complications from retraction injuries have been reported in the literature, but postoperative complications arising from liver retraction have not been published. The authors present a case of a symptomatic liver hematoma requiring hospital readmission for diagnosis and pain control and a review of retraction injuries. PMID- 10872983 TI - Laparoscopic repair of a chronic diaphragmatic hernia. AB - Diaphragmatic injuries that remain undetected after an acute traumatic event may lead to the formation of a diaphragmatic hernia. Symptoms of a chronic diaphragmatic hernia are related to the incarceration of abdominal contents in the defect or to impingement of the lung, heart, or thoracic esophagus by abdominal viscera. A 49-year-old woman with a symptomatic chronic diaphragmatic hernia from an unrecognized iatrogenic injury to the left hemidiaphragm sought treatment. The diaphragmatic injury occurred 2 years earlier when a low, left sided chest tube was placed for a persistent pleural effusion 2 weeks after a lower lobectomy for an aspergilloma. The patient's diaphragmatic hernia was diagnosed after an upper gastrointestinal series and an esophagogastroduodenoscopy. Approximately 75% of her stomach was incarcerated in the diaphragmatic defect. The diaphragmatic hernia was repaired laparoscopically using a 9 cm x 10-cm polytetrafluoroethylene patch sewn with nonabsorbable, interrupted, horizontal mattress sutures. Improvement of video technology, laparoscopic instruments, and surgical skills has allowed surgeons to expand the boundaries of advanced therapeutic laparoscopy. These factors facilitated the authors' standard tension-free prosthetic repair of a chronic diaphragmatic hernia using minimally invasive techniques. PMID- 10872984 TI - The use of double-straight needle device in laparoscopic incisional and ventral hernia repair. AB - Incisional and ventral hernias are good indications for laparoscopic hernia repair. A successful repair requires complete covering of the hernia defect, adequate tension of the prosthesis, and secure stapling by a hernia stapler. The authors introduce their technique using a double-straight needle device. This technique is easy and quick and achieves adequate fixation between the prosthesis and the abdominal wall, which reduces operating time and provides cosmetic benefit. PMID- 10872985 TI - Antimicrobial resistance with focus on oral beta-lactamases. AB - Over the last 10-15 yr antibiotic resistance has increased in the oral microflora. The beta-lactam antibiotics, i.e., penicillins and cephalosporins, are the most frequently used antimicrobial agents. Unfortunately, the efficiency of these drugs is increasingly being challenged by the emergence of resistant bacteria, which is mainly due to their production of beta-lactamases. In this paper, mechanisms of antibiotic resistance are reviewed, with emphasis on beta lactamases. This review also discusses how the presence of beta-lactamases in oral microorganisms may affect the treatment of oral diseases. Dentists can influence the emerging global crisis of antibiotic resistance by carefully evaluating the indications for antibiotic treatment. General guidelines for when and how to use antibiotics in dentistry are reviewed. PMID- 10872986 TI - Factors related to missed and cancelled dental appointments among adolescents in Norway. AB - The aim of this study was to explore possible explanatory factors related to high frequency of missed/cancelled dental appointments during the age group 12-18 yr. A total of 754 20 yr olds completed a questionnaire including variables measuring demographics, occupation (school/job), attendance pattern, attitudes to dentists, opinion about importance of dental treatment, and the psychometric scales Dental Fear Scale (DFS), Dental Beliefs Survey (DBS) and Geer Fear Scale (GFS). Based on written consents, the following data were recorded from their dental records: the total number of scheduled appointments, the number of missed and cancelled appointments and the individual caries experience of those in the age group 12-18 yr. A total of 124 subjects who had missed/cancelled 20% or more of their dental appointments during this age were defined as a target group. A stepwise regression model indicated that the likelihood of being included in the target group increased by a factor of 6.0 if the subject had forgotten dental appointments during the last 5 yr, by a factor of 3.5 for working or without specified occupation (as opposed to attending school), by a factor of 2.7 for negative beliefs of dentists, and by a factor of 2.1 for high caries experience. PMID- 10872987 TI - Advanced dental maturity in children with juvenile rheumatoid arthritis. AB - The subjects of the investigation comprised 95 girls and 73 boys with juvenile rheumatoid arthritis (JRA), and 102 girls and 66 boys representing healthy controls, all with a chronological age from 6.3 to 14.4 yr. The dental development was assessed from panoramic radiographs using a seven-tooth model. The radiographs were evaluated on three separate occasions with a minimum interval of one month in a randomized order, and blind with respect to absence or presence of JRA. In both JRA patients and healthy controls, dental maturity was ahead of chronological age. In addition, dental maturity was significantly advanced in JRA patients with 0.26 yr in girls and 0.28 yr in boys. It is tentatively suggested that the advanced dental development in JRA patients compared with healthy children was partly an effect of treatment with cortisone, while the influence of the disorder per se remains to be elucidated. PMID- 10872988 TI - Characterization of a family with dominant hypophosphatasia. AB - A kindred with dominant hypophosphatasia resulting from an alanine to threonine substitution at position 99 of the alkaline phosphatase protein is described. The clinical findings of individual members of the kindred were assessed by oral and physical examinations, or from the descriptions of multiple family members. The proband displayed enamel hypoplasia and premature loss of fully rooted primary anterior teeth, which were shown by histological examination to lack cementum. Serum alkaline phosphatase (ALP) and a vitamin B6 panel, and urine phosphoethanolamine (PEA) were measured on 21 family members. Based upon the clinical and laboratory tests, affected and unaffected status was assigned. Parametric linkage analysis of the kindred using different dominant models and frequency distributions for the disease allele and the mutation gave lodscores > 4.2 and confirmed the strong linkage between the disease and the mutation. Assuming the defined mutation causes the disease, the reliability of clinical and laboratory tests is assessed. PMID- 10872990 TI - Treatment of inflamed ferret dental pulps with recombinant bone morphogenetic protein-7. AB - Recombinant human BMP-7 (bone morphogenetic protein-7, osteogenic protein-1) is osteogenic, dentinogenic and cementogenic when implanted into the appropriate tissue in vivo. However, most studies characterizing the induction of these tissues have implanted BMP-7 into freshly surgerized, clinically healthy tissues. To determine if BMP-7 is dentinogenic in inflamed dental pulps, we applied BMP-7 to inflamed ferret pulps. A single application of 5 microg of a commercial preparation of lipopolysaccharide (LPS) from Salmonella typhimurium directly to the coronal pulp induced a reversible mixed inflammatory exudate of moderate intensity within 3 d. Treatment with a single application of 2.5, 7.5 or 25 microg recombinant human BMP-7/mg collagen (2 mg total mass/tooth) induced reparative dentinogenesis in controls but not LPS treated dental pulps. These data reveal that a single application of up to 50 microg/tooth of exogenous recombinant BMP-7 is insufficient to induce reparative dentinogenesis in ferret teeth with reversible pulpitis. Given that pulp cells in the inflamed tissues likely retain the capacity to respond to exogenous BMP-7, it is possible that insufficient active recombinant protein is available to induce tissue formation in experimentally inflamed dental pulps. PMID- 10872989 TI - Cytokine expression in periapical granulation tissue as assessed by immunohistochemistry. AB - The aims of this study were to investigate the expression of pro-inflammatory, anti-inflammatory and immune-related cytokines present in periapical lesions. We investigated the expression of cytokines: namely interleukins IL-2, IL-4, IL-6, IL-10 and interferon-gamma (IFN-gamma) in formalin-fixed, paraffin-embedded sections of periapical granulation tissue. The study samples were biopsies from 24 patients with periapical lesions: 12 with periapical granulomas and 12 patients with radicular cysts. Immunohistochemistry was also performed on tonsillar tissue which served as a control. We utilised a set of specific monoclonal antibodies and polyclonal monospecific antibodies to detect cells that expressed the different cytokines within the tissues. We also considered the nature of the periapical immune response by investigation of the T-helper 1 (Th 1) and T-helper 2 (Th-2) lymphocyte subsets using their cytokine profile, i.e., Th-1: IL-2 and IFN-gamma and Th-2: IL-4, IL-5 and IL-6. Only a few cells were weakly positive for the IL-2 protein in each of the tissue sections. Cells that expressed IL-4 or IL-6 were far more numerous than cells that expressed either IL 2 or IFN-gamma. Thus, we demonstrated a greater number of Th-2 cells in periapical lesions. This relative ratio of the T-cell subsets underlines the importance of the anti-inflammatory mechanisms taking place in the diseased tissue manifested by the wide array of IL-10-expressing cells: B cells, T suppressor cells (CD8 (+)) and tissue macrophages. The numbers of inflammatory cells expressing the anti-inflammatory molecules far outnumbered the cells that expressed pro-inflammatory cytokines. Thus, the downregulation of the inflammatory response and the predominant Th-2 or humoral immune response in periapical periodontitis may be important features that dictate the outcome of the disease process in the periapical lesion. PMID- 10872991 TI - Enamel demineralization under driving forces found in dental plaque fluid. AB - The present study was carried out to examine the kinetics of enamel demineralization in vitro under driving forces for demineralization (i.e., the degree of saturation with respect to enamel, DS(En)) similar to those found in dental plaque fluid. Thin sections of human enamel were exposed at 25 degrees C to lactic acid solutions with DS(En) values (DS(En) = [(Ca2+)5(OH-)(PO4(3 ))3/K(En)]1/9; K(En) = 5.5 x 10(-55)) ranging from 0.28 to 0.79. Lesion development was monitored by quantitative microradiography. Enamel mineral loss in solutions with DS(En) values of 0.28, 0.32 and 0.36 was first detected after 3, 3, and 7 wk of continuous exposure, respectively. Consistent with previous findings, subsurface demineralization was observed and rates of mineral loss increased significantly with decreasing DS(En) values. However, no mineral loss was observed in sections of enamel exposed to solutions with DS(En) values of 0.41 and 0.79, even after 11 months. These results suggest that (outer) enamel mineral behaves as a mineral phase that is less soluble than that dictated by the solubility product constant (K(En)) used in this study. Furthermore, these results indicate that the kinetics and general features of the demineralization process are maintained over a wide range of DS(En) values, including conditions that better reflect those found in the oral cavity. These findings are particularly relevant to the assessment of the cariogenic potential of dental plaque fluids. PMID- 10872992 TI - In situ induced demineralization in irradiated and non-irradiated human dentin. AB - The objective of this study was to evaluate the onset of initial demineralization in irradiated and non-irradiated human dentin. Dentin specimens were prepared from the cervical regions of 48 third molars. Either the lingual or the buccal dentin specimen of each tooth was irradiated fractionally up to 60 Gy (2 Gy/d, 5 d/wk). The remaining dentin sample was not irradiated. Two irradiated and two non irradiated dentin specimens were inserted into both buccal aspects of each 12 intraoral mandibular appliances. The appliances were worn by 12 persons for 5 wk day and night. One side was brushed daily with a fluoride-free toothpaste. On the other side, plaque was allowed to grow. Individual oral hygiene techniques were performed without any fluorides. During meals, the appliance was stored in 10% sucrose solution. After the in situ period, slabs (150 microm) were ground and studied by means of transversal microradiography and microscopic techniques. Concerning mineral loss and lesion depth, ANOVA revealed significant differences between brushed and non-brushed specimens, whereas no differences between irradiated and non-irradiated dentin lesions were found. It is concluded that (in vitro) irradiated dentine is not more susceptible to caries than non-irradiated, if adequate oral hygiene techniques are implemented. PMID- 10872993 TI - Distribution of non-collagenous dentin matrix proteins and proteoglycans, and their relation to calcium accumulation in bisphosphonate-affected rat incisors. AB - It has been reported that multiple injections of 1-hydroxyethylidene- 1,1 bisphosphonate (HEBP) to rats prevent mineralization of incisor dentin, thereby revealing high concentrations of calcium in the non-mineralized matrix of circumpulpal dentin. To identify the molecules responsible for calcium accumulation in circumpulpal dentin matrix, rats were injected daily with HEBP (8 mg P/kg) for 7 d, and the incisors processed for various histochemical and immunohistochemical staining of non-collagenous matrices of dentin. Cuprolinic blue reactions for proteoglycans (PGs) were equally distributed in non mineralized matrix of mantle and circumpulpal dentin layers. Dentin sialoprotein (DSP) and osteopontin (OPN) immunoreactions were found in non-mineralized circumpulpal dentin matrix, but not in mantle dentin. In normal incisors, however, predentin matrix showing significant DSP immunoreactivity was negative for Ca-GBHA reactions. HEBP-affected, non-mineralized OPN immunopositive bone matrix was also non-reactive for calcium. From these observations, neither PGs, OPN nor DSP appear to be responsible for calcium accumulation in HEBP-affected circumpulpal dentin. Stains-all reactive component, possibly dentin phosphoprotein (DPP), only showed the same distribution as that of Ca-GBHA in both HEBP-affected and normal dentin matrix, implicating a possible contribution of DPP to calcium accumulation in circumpulpal dentin and, hence, to appositional mineralization of dentin. PMID- 10872994 TI - Determination of changes on tooth-colored cervical restorations in vivo using a three-dimensional laser scanning device. AB - The present study aimed at the determination of changes of tooth-colored cervical restorations in vivo using an optical 3-dimensional laser scanning device. The study was performed on 197 cervical restorations placed on incisors, canines, and premolars. Four different tooth-colored restoration materials, a composite, a polyacid-modified resin composite, and two resin-modified glass ionomer cements, were used for the restoration of the lesions. For the determination of changes, images were taken at baseline and 15, 24 and 36 months after the placement of the fillings using a 3D-laser scanning device. The images were superimposed automatically, and digital subtraction was made by a specially developed image analysis software. The total substance loss on the entire filling surface at 36 months for the resin-modified glass ionomer Photac-Fil was 44 (+/-23) microm, for Fuji II LC 45 (+/-26) microm, for Dyract 71 (+/-47) microm and for Tetric 18 (+/ 12) microm. Differentiating between the class of lesion, a higher wear rate was observed at 36 months on restorations which had been placed in erosion/non carious cervical cavities (66 (+/-33) microm). In conclusion, the composite material demonstrated a distinctly lower surface wear rate over time in comparison to the resin-modified glass ionomer cements and the polyacid-modified resin composite. PMID- 10872995 TI - An 8-year evaluation of sintered ceramic and glass ceramic inlays processed by the Cerec CAD/CAM system. AB - The purpose of this study was to evaluate Cerec CAD/CAM inlays processed of two industrially made machinable ceramics during an 8-yr follow-up period. Each of 16 patients received two similar ceramic inlays. Half the number of the inlays were made of a feldspathic (Vita Mark II) and the other of a glass ceramic (Dicor MGC) block. The inlays were luted with a dual resin composite and evaluated clinically using modified USPHS criteria at baseline, 8 months, 2, 3, 5, 6 and 8 yr, and indirectly using models. At baseline, 84% of the inlays were estimated as optimal and 16% as acceptable. Postoperative sensitivity was reported by one patient for 8 months. Of the 32 inlays evaluated during the 8 yr, 3 failed due to fracture of the material. No secondary caries was found adjacent to the inlays. No significant differences in the clinical performance were found between inlays made of the two ceramics. It can be concluded that the CAD/CAM inlays processed of the two ceramics functioned well during the 8-yr follow-up period. PMID- 10872996 TI - Calcium leaching from dentin and shear bond strength after etching with phosphoric acid of different concentrations. AB - Based on the H2O-P2O5-CaO phase diagram, we hypothesize that a phosphoric acid concentration around 27 wt% leaches most calcium from dentin. We also hypothesize that bond strength is affected by resin infiltration, and that resin infiltration becomes incomplete when calcium leakage exceeds a certain value. Dentin disks were cut from human molars. Eight phosphoric acid concentrations were prepared (15.7-51.2 wt%). For each acid group, there were four etch time subgroups (15, 30, 60 and 120 s). The dentin disks were etched in acid and rinsed in water for times corresponding to 15 s, 30 s, 60 s and 120 s. The calcium concentrations were analyzed using atomic absorption spectroscopy. Composite cylinders were bonded to the remaining parts of the teeth using the same etching protocol, and shear bond strength was determined. The 29.2 wt% group demonstrated the highest and the 15.7 wt% group the lowest calcium leaching value. Even though there were trends towards lower bond strength for longer etch times, a statistically significant difference was only found between 30 and 120 s. There was no significant correlation between calcium leaching and bond strength. The results support the tested hypothesis that the highest leaching value would be around 27 wt% phosphoric acid. PMID- 10872997 TI - Decreasing prevalence of salivary lactobacilli in Swedish schoolchildren 1987 1998. AB - The prevalence and number of salivary lactobacilli was determined in all schoolchildren starting the 7th grade (12-13 yr; n = 1,578) in comprehensive school located in Kumla, Sweden between 1987 and 1998. Whole saliva samples were collected and transferred to dip-slides (Dentocult-LB) and incubated in room temperature for 7 d. Caries data were collected from the dental records and from bitewing radiographs. Both the number of lactobacilli and the prevalence of manifest caries and restorations exhibited a decreasing tendency and were significantly lower in 1998 than in 1987. The mean caries prevalence (DFS) declined from 5.2 to 1.8. In 1987, 45% of the children harboured high or very high lactobacilli counts ( > or =10(5) CFU/ml) in their saliva compared to approximately 10-15% at the later examinations. The proportion of children with no detectable counts varied between 4-29% during the study period, while children with low counts (10(3)-10(4) CFU/ml saliva) constituted a clear majority at the recent samplings. With such low levels in the population, the use of lactobacilli counts as a didactic tool in dietary counselling must be called into question. PMID- 10872998 TI - Laparoscopic ventral and incisional hernia repair in 407 patients. AB - BACKGROUND: Recurrence rates after primary repair of ventral and incisional hernias range from 25% to 52%. Recurrence after open surgery is less likely if mesh is used, but the wide fascial dissection and required flap creation increase complication rates. Laparoscopic techniques offer an alternative. STUDY DESIGN: To assess the safety and efficacy of laparoscopic ventral and incisional herniorrhaphy, we reviewed the records of all our patients who underwent such a procedure from November 1993 to August 1999. A laparoscopic approach was attempted in all patients considered to require a mesh repair. Patient demographic characteristics, operative details, and outcomes were recorded. RESULTS: Of 415 patients scheduled to undergo laparoscopic ventral or incisional herniorrhaphy, conversion to an open procedure was necessary in 8. All the remaining 407 patients (205 men and 202 women; mean age 53.2 years; range 13 to 88 years) were included in the study. Mean fascial defect size was 100.1 cm2 (range 1 to 480 cm2). In 97% of patients, expanded polytetrafluoroethylene mesh was used. Mean operating time was 97 minutes (range 11 to 270 minutes). Mean estimated blood loss was 35 mL (range 10 to 150 mL). Average hospital stay was 1.8 days (range 0 to 17 days). There were 53 complications (13.0%), including cellulitis of a trocar site, infection requiring mesh removal, prolonged suture pain, persistent seroma, intestinal injury, hematoma or postoperative bleeding, prolonged ileus, urinary retention, respiratory distress, fever, intraabdominal abscess, and trocar site herniation. There were no deaths. During a mean followup time of 23 months (range 1 to 60 months), there were 14 hernia recurrences (3.4%), 6 in patients in whom only a stapling device (no sutures) had been used to secure the mesh to the abdominal wall. CONCLUSIONS: Laparoscopic repair was completed in 98.1% of patients in whom it was attempted. The complication rate was acceptable. A short hospital stay and minimal blood loss were documented. The recurrence rate was 3.4%. Laparoscopic ventral and incisional hernia repair appear to be safe and effective. PMID- 10873000 TI - The utility of venous lactate to triage injured patients in the trauma center. AB - BACKGROUND: Field triage criteria for trauma patients results in over-triage rates of 30% to 50% to achieve under-triage rates of 10%. This large number of patients may stress trauma center resources. Elevated arterial lactate (ALAC) levels have been shown to be a marker of serious injury but the need for arterial sampling limits the utility of the determination. The goal of this study was: 1) to determine the correlation between venous lactate (VLAC) and ALAC; 2) to determine whether VLAC could identify those patients with serious injuries; and 3) to compare an elevated VLAC level against standard triage criteria (STC) in their ability to identify major injury. STUDY DESIGN: Arterial and venous samples for blood gas and lactate analyses were obtained in 375 patients within 10 minutes of patient arrival to the trauma center. Arterial and venous samples were drawn within 2 minutes of each other, placed on ice, and analyzed within 10 minutes of sampling. The location of sampling was left to physician discretion. Data collected included injury mechanism, demographics, admission vital signs, emergency department disposition, length of stay, and injury severity scores (ISS). Admission to the ICU, need for emergency operation, length of stay, and death were noted. Emergency medical service staff were queried to determine which standard triage criteria (STC) were fulfilled. RESULTS: The mean ALAC was 3.11 mmol/L (SD 3.45, 95% confidence interval [CI] 2.67 to 3.55) and mean VLAC was 3.43 mmol/L (SD 3.41, 95% CI 2.96 to 3.90). There was no significant difference between ALAC and VLAC. The correlation between ALAC and VLAC was 0.94 (95% CI 0.94 to 0.96, p = 0.0001). An elevated VLAC predicted moderate to severe injury and there was a significant association between an increased lactate and maximum Abbreviated Injury Score (AIS) of 4 and 5 (ANOVA, F = 8.26, p < 0.001). Patients with VLAC > or =2 mmol/L had significantly increased relative risks of ISS > or = 13, death, admission to the ICU, and length of stay > 2 days. In comparison with STC, a VLAC > or = 2 mmol/L decreased undertriage in patients with ISS > or = 13 by one half (11% versus 24%) for patients with ISS > or = 13 and decreased over triage by 28% (46% versus 64%). These data were most pronounced for patients injured in motor vehicle collisions. CONCLUSIONS: VLAC is an excellent approximation for ALAC. A VLAC > or = 2 mmol/L appears to predict an ISS > or = 13, the need for ICU resources, and prolonged hospital stays. VLAC was significantly better than STC in all patients and was most useful in victims of blunt trauma, especially motor vehicle collisions. PMID- 10872999 TI - Recurrence after endoscopic transperitoneal hernia repair (TAPP): causes, reparative techniques, and results of the reoperation. AB - BACKGROUND: Even though the introduction of endoscopic surgical techniques to inguinal hernia therapy dates back 10 years, only a few data exist concerning the problem of development of a recurrence after endoscopic repair. Similarly there are only anecdotal reports on the feasibility of an endoscopic reintervention for this situation. For the first time we are able to present data of a prospective study on both issues. STUDY DESIGN: We analyzed the data of a prospectively documented series of 46 transperitoneal hernia repair reinterventions after endoscopic hernia repair. In 33 patients from our own clinic we evaluated the cause of recurrence after transperitoneal hernia repair. Together with these and 13 more patients sent to us from external clinics we examined the efficiency of an endoscopic reoperation. RESULTS: When implanting a 13 x 8-cm mesh with an incision (phase I) we found the main cause of recurrence to be that the mesh was too small (47.4%) and the region of the mesh incision was insufficient (42.1%). After a change to a 15 x 10-cm implant without incision (phase II) the main cause of recurrence was found to be a mesh dislocation (38.9%) and the rate of recurrence dropped from 2.8% (phase I) to 0.36% (phase II). The transperitoneal reoperation lasted for a median of 75 minutes (range 45 to 170 minutes) for the medial recurrence and a median of 110 minutes (range 65 to 190 minutes) for the lateral recurrence (p = 0.009). The total rate of complications was 10.9%, and the rate of re-recurrence was 0% after a median followup of 26 months (range 2 to 72 months). CONCLUSIONS: To avoid hernia recurrence after transperitoneal hernia repair operations a sufficiently large mesh (at least 15 x 10 cm) has to be implanted, preferably without an incision, after an extensive parietalization. The endoscopic reoperation for recurrence can be done only in a transperitoneal way and is effective with comparably low complication rates. The procedure is significantly easier for a medial recurrence compared with a lateral recurrence. This method of reoperation should be reserved for endoscopically experienced surgeons. PMID- 10873001 TI - Myocardial infarction as a complication of injury. AB - BACKGROUND: MI is a rare complication of trauma. We anticipate that the aging of the population and the concomitant rise in geriatric trauma will result in an increase in acute illnesses of the elderly (such as MI) complicating recovery from injury. The purpose of this article is to define the presentation of MI in the immediate postinjury period. STUDY DESIGN: Medical records of all trauma patients in whom MI developed during their hospitalizations at a single Level I trauma center, the Barnes Hospital/Washington University Medical Center, between 1990 and 1999 were screened through the trauma registry. Nineteen patients with possible postinjury MI were identified. Of these, five had bona fide cases of postinjury MI, five had ambiguity about whether MI preceded or followed trauma, one had an MI resulting in trauma, and eight were excluded because they did not meet strict diagnostic criteria for MI. RESULTS: The five patients with posttraumatic MI were older than the general trauma population with ages ranging from 51 to 81 years (mean +/- SD = 72 +/- 14 years). Each had preexisting medical illnesses, some of which are recognized to predispose to coronary artery disease. There were no identifiable precipitants other than the recent injury. Importantly, only one of the five patients had chest pain as a presenting symptom and each of the five cases was complicated by acute congestive heart failure. CONCLUSIONS: MI remains a rare but important complication of injury and may increase owing to the changing demographics of trauma victims. Methods for thorough history-gathering to identify preexisting conditions, for early hemodynamic monitoring and anticoagulation for MI in the setting of trauma, and for identifying preexisting conditions should be defined. The presentation of MI in the setting of injury is atypical and complications are frequent. PMID- 10873002 TI - Bacteremia associated with central venous catheter infection is not an independent predictor of outcomes. AB - BACKGROUND: Infection is the leading complication of central venous catheters. In the setting of suspected line infection, the CDC recognizes only catheter-related bloodstream infection but not catheter infection without bacteremia, which is designated "colonization." To evaluate the hypothesis that catheter-related bloodstream infection has worse outcomes than catheter infection without bacteremia, we compared demographics, clinical data, and outcomes. STUDY DESIGN: Analysis of catheter infections was performed on data collected prospectively for all episodes of infection occurring from December 1996 to September 1999 on the surgical services at a university hospital. Catheter tips were cultured only when infection was suspected. Catheter infection without bacteremia was defined as systemic evidence of infection, the presence of at least 15 colony-forming units on the catheter tip by a semiquantitative technique, and absence of bloodstream infection with the same organism as the catheter. Catheter-related bloodstream infection required the presence of bacteremia with the same organism as the catheter tip. RESULTS: The 59 patients with catheter-related bloodstream infection had more coexistent infections than the 91 patients with catheter infection without bacteremia (2.9+/-0.1 versus 1.7+/-0.1; p=0.0001), most commonly pneumonia (37.3% versus 16.5%, p = 0.004) and urinary tract infections (28.8% versus 8.8%, p = 0.001). Catheter-related bloodstream infection was associated with an increased proportion of gram-negative organisms compared with catheter infections without bacteremia (29.5% versus 16.9%, p = 0.04) and a trend toward fewer gram-positive organisms (61.5% versus 73.7%, p = 0.07). There were no differences in APACHE II score, WBC, length of hospital stay, time from admission to fever, time from fever to treatment, normalization of WBC, days of antibiotics, defervescence, gender, presence of comorbidities, occurrence of colonization while in an ICU, or mortality rate (18.6% with bacteremia, 24.2% without; p = 0.42). CONCLUSIONS: The presence of bloodstream infection in addition to catheter infection does not appear to alter outcomes. The definition of catheter infection perhaps should be extended to include catheter infections without bloodstream infection in the presence of systemic illness without another source. PMID- 10873003 TI - Measurement of endotracheal tube cuff leak to predict postextubation stridor and need for reintubation. AB - BACKGROUND: The purpose of this study was to determine the predictive value of an endotracheal tube cuff leak for the development of postextubation stridor and the need for reintubation. STUDY DESIGN: Consecutive trauma patients who required intubation at a level I trauma center from July 1997 to July 1998 were studied prospectively. Pediatric patients and those who did not meet the standard weaning protocol criteria established by the Division of Trauma and Surgical Critical Care were excluded. Injury Severity Score, endotracheal tube size, reason for intubation, and the number of days intubated before the initial extubation attempt were recorded. At the time of extubation, the difference in exhaled tidal volume from before to after endotracheal tube cuff deflation was calculated. This number was then divided by the exhaled tidal volume before cuff deflation and was recorded as the percent cuff leak. Patients were followed for 24 hours after extubation for the development of stridor or need for reintubation. Statistical analysis to compare subgroups of patients was performed using ANOVA with Scheffe post hoc analysis. RESULTS: Among the 110 patients analyzed, the most common reason for intubation was closed-head injury. Seven patients (6.4%) developed stridor alone and had a mean cuff leak of 5 8 mL (8.4% of tidal volume before cuff deflation). Six patients (5.5%) experienced stridor that required reintubation and had a mean cuff leak of 68 mL (9.2% of tidal volume before cuff deflation). Patients who developed stridor or needed reintubation had been intubated for a significantly greater length of time than those not developing stridor or requiring reintubation (2.6 versus 3.0 days, p < 0.001). There were no differences in Injury Severity Score, endotracheal tube size, or reason for intubation between these groups. CONCLUSIONS: A cuff leak of less than 10% of tidal volume before cuff deflation is useful in identifying patients at risk for stridor or reintubation (96% specificity). It appears that the amount of cuff leak decreases after intubation for more than 3 days, increasing the risk of stridor and need for reintubation. This information may be helpful in identifying those patients who need treatment for laryngotracheal edema, ie, use of steroids or anesthesia during extubation, the efficacy of which remains to be determined. PMID- 10873004 TI - Pediatric postoperative abdominal wound dehiscence: transverse versus vertical incisions. AB - BACKGROUND: Fascial dehiscence is uncommon in children but can have serious consequences when it occurs. There are multiple risk factors for fascial dehiscence, including the type of incision used. Pediatric surgeons often use a supraumbilical transverse incision particularly in infants because of the access this incision provides to the entire abdomen. This article details the experience with fascial wound dehiscence at a large tertiary children's hospital and focuses on problems with the types of incision used. STUDY DESIGN: This is a retrospective review of 2,785 intraabdominal operations performed over a 5-year period at Children's Hospital and Regional Medical Center in Seattle. Risk factors for dehiscence were reviewed for each case of fascial dehiscence. Statistical analysis using chi-square was used to examine for differences in complication rates between transverse and vertical incisions. RESULTS: In this series, 2,442 children (88%) had transverse incisions and 343 (12%) had vertical incisions. Twelve children had abdominal fascial dehiscence post-operatively. Six cases involved transverse incisions and six involved vertical incisions. Five of the children suffered evisceration. One child died as a direct result of the dehiscence. There were multiple risk factors for dehiscence in 10 of the 12 children. Vertical incisions were found to be much more likely to dehisce than were transverse incisions, especially in children under 1 year of age (p < 0.001). CONCLUSIONS: Vertical incisions are more apt to dehisce than transverse incisions in children, particularly babies. We recommend the use of transverse incisions whenever possible in babies less than 1 year of age, especially when other risk factors for dehiscence are present. PMID- 10873005 TI - Use of technetium-99m-labeled colloid albumin for preoperative and intraoperative localization of nonpalpable breast lesions. AB - BACKGROUND: Management of clinically occult breast lesions is still a major point of debate. Several techniques (eg, skin projection, guidewire localization) have been proposed, but all of them have technical limitations. STUDY DESIGN: The aim of this study was to assess the efficacy of a new method to locate occult breast lesions using technetium-99m (99mTc)-labeled colloid particles of human serum albumin (radioguided occult lesion localization). We studied 647 consecutive patients (mean age 51.3 years; range 25 to 77 years) with nonpalpable breast lesions detected mammographically or by ultrasonography. Within 24 hours before operation, 3.7 MBq (0.1 mCi) of 99mTc-labeled colloid was injected directly into the center of the lesion using stereotactic mammographic guidance (when only microcalcifications were present) or ultrasonographic guidance (for opacities). Excision biopsy was performed with a gamma-detecting probe. After excision, the area was checked for residual radioactivity and the specimen was radiographed to verify complete removal of the lesion. The material was then sent for pathologic examination. The absorbed dose to the inoculated area and the external irradiation to staff were also determined. RESULTS: In all 647 patients, the "hot spot" was located easily and quickly. X-ray and scintigraphy of the specimen verified the presence and centricity of the lesion in all patients but three (99.5%). Pathologic examination revealed 340 cancer lesions (52.6%). Of these patients, 339 (99.7%) were treated by breast-conserving operations and one (0.3%) received a modified radical mastectomy. No major surgical or postoperative complications were encountered. No recurrences were documented during follow-up. The absorbed dose to the breast and other tissue was negligible (0.03 +/- 0.02 mGy/MBq), as was the dose to the surgeon's hands (7.5 +/- 5.0 microSv/h). The latter dose represents 0.015% and 0.002% of the recommended limits of the European Community for the general population and for exposed workers, respectively. CONCLUSIONS: Radioguided occult lesion localization seems to offer a simple and reliable method to locate occult breast lesions with a gamma detecting probe, allowing complete removal of the lesion in 99.5% of patients. Because of the small quantity of radioactivity, the procedure is safe for both patients and medical staff. PMID- 10873006 TI - CT in the selection of patients with abdominal or pelvic sarcoma for reoperative surgery. AB - BACKGROUND: Retroperitoneal or visceral sarcoma may recur with disease limited to the abdomen and pelvis. In this clinical situation, further surgical treatments in an attempt to control the disease may be appropriate. CT is used to help select patients for additional surgical interventions. STUDY DESIGN: Preoperative abdominal and pelvic CT scans of 33 patients with recurrent sarcoma were reviewed retrospectively. All patients underwent reoperative surgery and, when appropriate, perioperative intraperitoneal chemotherapy. Patients were divided into two groups according to survival and disease status: alive with no evidence of disease (n = 7) and alive with disease or dead of disease (n = 26). Twenty-two CT indices were studied retrospectively for each patient and evaluated statistically. RESULTS: The presence of large (greater than 5 cm) tumor volume in 3 of the 13 abdominopelvic regions resulted in a significant difference in the prognosis between the groups of patients. These findings included tumor in the left lower quadrant (p = 0.032), tumor in the pelvis (p = 0.008), and tumor in the distal jejunum (p = 0.032). Two other CT indices that showed a significant difference in survival between the groups were involvement of five abdominopelvic regions or fewer (p = 0.008) and a peritoneal cancer index of 15 or less (p = 0.03). A statistical approach using a tree-structured diagram showed that patients with tumor diameter greater than 5 cm in the pelvis accompanied by tumor involvement of more than one segment of small bowel had a 0% probability of postoperative disease-free survival. In contrast, patients with tumor diameter less than 5 cm in the pelvis on CT had an 86% probability of disease-free survival. CONCLUSIONS: For patients with recurrent sarcoma, selection criteria were generated by a preoperative CT of the abdomen and pelvis. In this disease, CT was a reliable diagnostic test for predicting benefit from further surgical interventions and should be used in the future to help select patients for an aggressive versus a palliative approach. PMID- 10873007 TI - Median pancreatectomy for tumors of the neck and body of the pancreas. AB - BACKGROUND: When enucleation is too risky because of possible damage of the main pancreatic duct, benign tumors located in the neck or body of the pancreas are usually removed by a left (spleno)-pancreatectomy or by a pancreatoduodenectomy. But standard pancreatic resection results in an important loss of normal pancreatic parenchyma and may cause impairment of exocrine and endocrine function. The aim of this study was to evaluate early and longterm results of median pancreatectomy, a limited resection of the midportion of the pancreas, in selected patients with benign or borderline tumors of the pancreas. STUDY DESIGN: Records of patients at Ospedale Busonera between November 1985 and September 1998 were reviewed. Ten patients with tumors of the neck or body of the pancreas underwent median pancreatectomy; the cephalic stump was sutured and the distal stump was anastomosed with a Roux-en-Y jejunal loop. Followup included clinical evaluation and routine laboratory tests: abdominal ultrasonography, exocrine and endocrine pancreatic function with fecal chymotrypsin, and an oral glucose tolerance test. RESULTS: Pathologic examination showed: insulinoma (n = 3), mucinous cystadenoma (n = 3), nonfunctioning endocrine tumor (n = 1), papillary cystic neoplasm (n= 1), serous cystadenoma (n = 1), and intraductal mucinous tumor (n = 1). Operative mortality and morbidity were 0% and 40%, respectively; pancreatic fistula occurred in three patients. At mean followup of 62.7 months, no recurrence was found and no patient had exocrine insufficiency or glucose metabolism impairment. CONCLUSIONS: Median pancreatectomy is a safe and effective alternative to major pancreatic resection in selected patients with benign or low malignant lesions of the pancreas. This procedure carries a surgical risk similar to that of the standard operation, but avoids extensive pancreatic resection and pancreatic function impairment. PMID- 10873008 TI - Median pancreatectomy: do the risks justify the effort? PMID- 10873009 TI - Sigmoid volvulus in children and adolescents. AB - BACKGROUND: Sigmoid volvulus is an exceptionally rare and potentially life threatening condition in the pediatric age group. STUDY DESIGN: We report our experience with three children treated for sigmoid volvulus and review the cases reported in the medical literature since 1940. RESULTS: Since 1940, 63 cases of sigmoid volvulus in children (including this series) have been reported. The median age was 7 years and the male to female ratio was 3.5:1. Two distinct presentations (acute and recurrent) were identified. Abdominal symptoms dominated the clinical picture. Barium enemas either confirmed or were highly suggestive of sigmoid volvulus. Reduction by barium enema was successful in 77% (10 of 13) of the attempts. Forty-nine patients underwent operative treatment, with sigmoidectomy (with or without primary anastomosis) being the most common. The overall mortality rate was 6%, operative mortality was 8.1%, and neonatal mortality was 14%. Associated conditions were frequent. Particular emphasis should be placed on ruling out Hirschsprung's disease (present in 11 of 63 patients). CONCLUSIONS: Sigmoid volvulus remains a rare occurrence in children, but it should be included in the differential diagnosis of pain in children when colonic distention is present. An algorithm for treatment is proposed. PMID- 10873010 TI - What's new in the management of short gut syndrome in children. PMID- 10873011 TI - Postoperative colorectal cancer surveillance. PMID- 10873012 TI - Cavernous hemangioma of the rectosigmoid. PMID- 10873013 TI - Images for surgeons. Concomitant open surgical repair of an abdominal aortic aneurysm and endovascular repair of a thoracic aortic aneurysm. PMID- 10873014 TI - A method for easier laparoscopic cholangiography and common bile duct exploration. PMID- 10873015 TI - An alternative repair technique for anastomotic leakage after total gastrectomy. PMID- 10873016 TI - Trendelenburg patient positioning: a reevaluation. PMID- 10873017 TI - Intermediate-term results of the free tracheal autograft for long segment congenital tracheal stenosis. AB - BACKGROUND/PURPOSE: Since 1996 a new procedure--the free tracheal autograft--has been used to repair long segment congenital tracheal stenosis (LSCTS). The purpose of this report is to examine the intermediate-term results of that technique. METHODS: Between January 1996 and July 1999, 10 infants underwent repair of LSCTS using a free tracheal autograft. Age ranged from 10 days to 23 months (mean age, 6.6 months). Six infants had a pulmonary artery (PA) sling; 5 had intracardiac anomalies. On cardiopulmonary bypass (CPB) the trachea was incised anteriorly through the area of stenosis. The midportion of the stenotic trachea was excised, and an end-to-end anastomosis was made posteriorly. The excised tracheal segment was trimmed and sutured in place anteriorly as a free autograft. In 5 patients the autograft was not long enough, and the upper portion of the tracheal opening was patched with pericardium. RESULTS: There was 1 death 26 days postoperatively in a child that had simultaneous repair of tetralogy of Fallot and required extracorporeal membrane oxygenation postoperatively for cardiac failure. The other 9 children are alive and well at 2 to 44 months postoperatively (mean follow-up, 24 months). One child had autograft dehiscence and required replacement of the autograft with an aortic homograft. Two children have tracheostomies at 6 and 36 months postoperatively. All children have had serial postoperative bronchoscopic examinations. Most recent bronchoscopies have shown widely patent tracheal lumina in all survivors. CONCLUSION: Intermediate term follow-up of children with a free tracheal autograft continues to support our use of this technique as our procedure of choice for infants with LSCTS. PMID- 10873018 TI - One-stage Soave pull-through for Hirschsprung's disease: a comparison of the transanal and open approaches. AB - PURPOSE: The authors reviewed their experience using the transanal Soave technique, to determine (1) if it offers any advantages over the standard open approach and (2) whether routine laparoscopic visualization is necessary. METHODS: The case reports of 37 consecutive children less than 3 years old undergoing Soave pull-through were reviewed. Patients were excluded from analysis if they had total colon disease or had a previous colostomy. The patients were divided into 3 groups: open Soave (OS, n = 13), transanal Soave with routine laparoscopic visualization (LVS, n = 9), and transanal Soave with selective laparoscopy or minilaparotomy (TAS, n = 15). Cost was calculated based on hospital stay, operating room time, and use of laparoscopic equipment. RESULTS: In the TAS group, suspicion of a longer segment led to the selective use of laparoscopy with or without biopsy in 2 children, and the use of a small umbilical incision for mobilization of the splenic flexure in 2. There were no differences among groups with respect to age, weight, gender, transition zone, operating time, blood loss, intraoperative complications, enterocolitis, or stricture or cuff narrowing. Hospital stay was significantly longer in the OS group (median, 7 days; range, 3 to 47) than the LVS (median, 1; range 1 to 6) or TAS (median, 1, range, 1 to 3) groups. Cost (in thousands of dollars) was also higher in the OS group (median, 6.9; range, 3.9-25.7) than the LVS (median, 3.9; range, 3.6 to 6.4) or TAS (median, 3.4; range, 2.2 to 9.4) groups. Repeat surgery was necessary for 4 OS patients: 2 adhesive small bowel obstructions (1 of whom died), 1 twisted pull-through, and 1 recurrent aganglionosis. Three TAS patients required repeat surgery: 1 twisted pull-through, 1 anastomotic leak, and 1 cuff narrowing. CONCLUSIONS: These data suggest that the transanal pull-through is associated with a significantly shorter hospital stay and lower cost than the open approach, without an increased risk of complications. Because there is no intraabdominal dissection, there probably is a lower incidence of adhesive bowel obstruction. Routine laparoscopic visualization or minilaparotomy is not necessary but should be used in children who are at higher risk for long segment disease. PMID- 10873019 TI - Immunolocalization of the gap junction protein Connexin43 in the interstitial cells of Cajal in the normal and Hirschsprung's disease bowel. AB - BACKGROUND: Interstitial cells of Cajal (ICC) are pacemaker cells between gastrointestinal smooth muscles; they generate spontaneous slow waves of the smooth muscle layers and mediate neurotransmission. The cellular network of ICC is connected by Gap junctions to each other and to the smooth muscle cells. Although there have been several studies reporting distribution of ICC in the normal bowel and pathological conditions such as Hirschsprung's disease, there is little information on the crucial role of Gap junctions in the intercellular communication in the gut musculature. The aim of this study was to investigate the immunolocalization of the Gap junction protein Connexin43 in the normal and Hirschsprung's disease (HD) bowel using whole-mount preparation technique and confocal laser scanning microscopy. METHODS: Full-thickness bowel specimens were collected at pull-through operation from 8 patients diagnosed as having HD. Normal control large bowel specimens were collected from 12 patients during bladder augmentation operation. Whole-mount preparation was performed on all specimens and double immunostaining was carried out using anti c-kit and antiConnexin43 antibodies. The immunolocalization was detected with the help of confocal laser scanning microscopy. RESULTS: Connexin43 immunoreactivity appeared in and between the c-kit-positive cells and along the smooth muscle fibers of the normal bowel and ganglionic part of HD bowel. In the aganglionic part of HD bowel there was no expression of Connexin43. In the transitional zone of HD the Connexin43 staining was weak and colocalized only in the processes of the c-kit positive Cajal cells. CONCLUSIONS: Results of this study show for the first time that Gap junctional protein Connexin43 is present in the ICCs, which form a 3 dimensional network in the normal bowel wall. The lack of expression of Connexin43 in the aganglionic bowel and reduced expression in the transitional zone of HD suggest that the impaired intercellular communication between ICCs and smooth muscle cells may partly be responsible for the motility dysfunction in HD. PMID- 10873020 TI - Long-term follow-up of redo pull-through procedures for Hirschsprung's disease: efficacy of the endorectal pull-through. AB - BACKGROUND/PURPOSE: The purpose of this study was to review the authors' 25-year experience with redo pull-through procedures for Hirschsprung's disease including surgical technique and long-term outcome. METHODS: From 1974 to now, over 325 patients with Hirschsprung's disease have been treated at C.S. Mott Children's Hospital. This includes 30 patients referred after an unsuccessful pull-through at another hospital and 2 patients with an unsuccessful pull-through from C.S. Mott. All redo pull-throughs (n = 19) were performed in these patients, and their clinical courses are reviewed. RESULTS: Twelve patients required reoperation secondary to a mechanical problem with their first pull-through. The other 7 patients had evidence of residual segments of dilated colon leading to functional failure of their initial operation including 5 patients with documented aganglionic bowel present at the second pull-through. Ten of the patients requiring reoperation initially had an endorectal pull-through (ERPT), 5 had a Duhamel procedure, 3 had a Swenson procedure, and 1 had a Rehbein procedure. Choice of revision was an ERPT in 8 patients in whom an adequate rectal cuff could be developed. Additional redo procedures included a Duhamel in 8 patients and a Swenson in 3 patients. Follow-up ranges from 3 months to 23 years (mean, 13.8 years). There were no deaths in the series, and 1 patient required a third pull-through. All patients who are not neurologically impaired and are over age 3 are continent except one (94%). Stools per day range from 1 to 10 (mean, 3.2). CONCLUSIONS: Redo pull-through operations for Hirschsprung's disease appear to be as effective as primary procedures in terms of continence and stooling frequency. Distinct from other series, we found an ERPT to be the procedure of choice if an adequate rectal cuff was present. PMID- 10873021 TI - Glial-derived growth factor signaling pathway in infantile hypertrophic pyloric stenosis. AB - BACKGROUND/PURPOSE: Glial-derived growth factor (GDNF), which is the ligand of RET is reported to be essential for the development of enteric nervous system. A GDNF knockout mouse model has shown that the gastric region is a critical passing site between GDNF-RET-independent neuroblasts (colonizing the esophagus) and GDNF RET-dependent neuroblasts (colonizing the small and large bowel). The earliest GDNF site of production is the mesenchyme and the outer smooth muscle cell (SMC) layer of the developing bowel. In the mature gastrointestinal tract the presence of GDNF is restricted to enteric glial cells. The aim of this study was to investigate the expression of GDNF and RET in infantile hypertrophic pyloric stenosis (IHPS). METHODS: Full-thickness muscle biopsy specimens were obtained from 8 IHPS patients at pyloromyotomy and from 8 age-matched controls without gastrointestinal disease. Indirect immunohistochemistry was performed using avidin-biotin-peroxidase complex method with anti-GDNF and anti-RET antibodies. Quantitative analysis was performed using sandwich-type enzyme-linked immunosorbent assay (ELISA) for GDNF. RESULTS: GDNF- and RET-positive nerve fibers were absent or markedly reduced in IHPS compared with controls. GDNF was expressed strongly by smooth muscle cells of both muscular layers in IHPS, whereas no GDNF expression was detected in pyloric muscle of controls. The quantity of total GDNF in IHPS was significantly higher than in controls (P < .01). CONCLUSIONS: The lack or markedly decreased number of GDNF-positive nerve fibers in IHPS supports the hypothesis of a selective immaturity of the enteric glia in the muscular layers in IHPS. The strong expression of GDNF in smooth muscle cells in IHPS and the increased levels of GDNF in IHPS suggest a compensatory mechanism by which the smooth muscle cells continue to produce GDNF until maturation of the enteric glial cells occurs. PMID- 10873022 TI - Abdominal compartment syndrome in children: experience with three cases. AB - BACKGROUND/PURPOSE: Abdominal compartment syndrome (ACS) is defined as cardiopulmonary or renal dysfunction caused by an acute increase in intraabdominal pressure. Although the condition is well described in adults, particularly trauma patients, little is known about ACS in children. METHODS: Three girls, ages 4, 5, and 5 years, were treated for ACS by silo decompression. Each child presented in profound shock, required massive fluid resuscitation, and had tremendous abdominal distension. The first child sustained a thoracoabdominal crush injury, underwent immediate celiotomy for splenic avulsion and a liver laceration, and required decompression 5 hours postoperatively. The second underwent ligation of her bluntly transected inferior vena cava; because of massive edema, her abdominal wall could not be closed, and prophylactic decompression had to be performed. The third presented with shock of unknown etiology, and ACS developed acutely with a bladder pressure of 26 mm Hg. RESULTS: Respiratory, renal, and hemodynamic function improved immediately in all 3 patients after decompression. Subsequently, each child underwent abdominal wall reconstruction and recovered uneventfully. CONCLUSIONS: ACS is a potentially lethal complication of severe trauma and shock in children. To prevent the development of renal or cardiopulmonary failure in these patients, decompression should be considered for acute, tense abdominal distension. PMID- 10873023 TI - Routine insertion of a silastic spring-loaded silo for infants with gastroschisis. AB - BACKGROUND/PURPOSE: Gastroschisis traditionally is managed by emergency operating room closure (EC), with a silo reserved for cases that cannot be closed primarily. The authors recently began using routine insertion of a SILASTIC (Dow Corning, Midland, MI) spring-loaded silo (SLS), followed by elective closure. METHODS: A total of 43 consecutive neonates with gastroschisis were treated between 1993 and 1998. RESULTS: Thirty patients underwent EC, and 13 underwent closure after insertion of a SLS (10 at bedside, 3 in the operating room). Eight infants treated by EC required staged repair. There were no differences with respect to gestational age, birth weight, gender, Apgar score, maternal age, or mode of delivery. Median length of stay was 32 days for EC and 25 days for SLS (P = .05). The SLS group required fewer days on a ventilator (4 v 6 days, P = .03) and had lower intraoperative (28 v 21, P = .02) and early postoperative peak airway pressures. The time to tolerate full feedings was 21 days for SLS and 27 days for EC (P = .07). The SLS group had fewer complications and a lower median hospital charge ($71,498 v $85,147; P = .05). CONCLUSION: SLS followed by elective repair permits gentle, gradual reduction of the viscera. When compared with EC, SLS is associated with significantly lower airway pressures, earlier extubation, fewer complications, and decreased length of stay and hospital charges. PMID- 10873024 TI - Treatment of inflammatory bowel disease in a rodent model with the intestinal growth factor glucagon-like peptide-2. AB - BACKGROUND/PURPOSE: Microinjection of a Fisher (F344) rat zygote with human HLA B27 and beta2-microglobulin genes induces spontaneous chronic gastrointestinal (GI) inflammation similar to lesions seen in patients with inflammatory bowel disease (IBD). This study was designed to evaluate the potential therapeutic benefit of GLP-2, an intestinal growth factor, in this transgenic rat model of IBD. METHODS: Five F344 (control) and 10 HLA-B27 (on a F344 background) rats at 25 weeks of age were used. Rats were divided into the following 3 groups: group 1, F344 rats, no treatment (n = 5); group 2, HLA-B27, no treatment (n = 5); and group 3, HLA-B27, treated with a 14-day systemic infusion (via the jugular vein) of GLP-2 at 50 microg/kg/d (n = 5). After infusion, all rats underwent laparotomy, and the intestine from the ligament of Treitz to the rectum was harvested. Total mucosal damage (percent surface area) was measured using image analysis software (Sigmascan 2.0). Microscopic analysis was performed by a blinded reviewer and scored as follows: 0, no inflammation; 1, mild inflammation; 2, moderate inflammation; and 3, severe inflammation. Colonic mucosal total RNA was assayed for tumor necrosis factor alpha (TNF-alpha), interferon-gamma (IFN gamma), interleukin-2 (IL-2), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), internal standard, mRNA by reverse transcriptase polymerase chain reaction. Statistical analysis was performed using analysis of variance (ANOVA) and expressed as mean +/- SEM. RESULTS: Normal F344 rats did not show evidence of gross or histological lesions in the small or large intestine. GLP-2 reduced total mucosal damage from 9.0% +/- 0.7% in group 2 to 0.9% +/- 0.5% in group 3 (P < .01). The histological lesion score was reduced from 7.0 +/- 0.6 in group 2 to 4.4 +/- 0.8 in group 3 (P < .01). Furthermore, GLP-2 reduced the mean band intensity (MBI) of TNF-alpha (0.4 +/- 0.04 in group 2 to 0 in group 3, P < .01) and IFN-gamma (0.3 +/- 0.02 in group 2 to 0 in group 3, P < .01). CONCLUSIONS: These data show for the first time that GLP-2 significantly reduces gross (90% decrease) and histological (40% decrease) lesions in this rat model of IBD. This is further supported by a significant decrease in gene expression of the inflammatory mediators TNF-alpha (100% decrease) and IFN-gamma (100% decrease). These data suggest a potential therapeutic role for GLP-2 in IBD. PMID- 10873025 TI - The role of anti-tumor necrosis factor-alpha and interleukin-10 in protecting murine neonates from Escherichia coli sepsis. AB - BACKGROUND/PURPOSE: The neonate is at much higher risk for septic complications and death than the adult. Although some aspects of the infant's immune response are immature, others are fully functional. Many models of septic death are caused by an overexpression of proinflammatory cytokines. If there were inadequate down regulatory mechanisms, this could lead to an overexpression of proinflammatory cytokines. The authors hypothesized that the high mortality rate of the newborn was caused by overexpression of tumor necrosis factor (TNF-alpha) and that interleukin-10 (IL-10) would attenuate this response. The aim of this study was to determine if TNF-alpha plays an important role in early death from Escherichia coli sepsis in the newborn animal and if blocking TNF improves survival. METHODS: A dose response curve was determined for 1 day old C3H/HEN mice using 10(5) intraperitoneal E coli resulting in a 30% to 50% mortality rate. Litters of newborn (1 day old) C3H/HEN mice received a subcutaneous injection of either 25 or 50 ng of murine IL-10 or 20 microL of anti-TNF-alpha 4 hours before a bacterial challenge. Control animals received nothing. Animals were observed for 5 to 7 days. At least 6 litters (18 pups per group) were used for each regimen. RESULTS: Anti-TNF-alpha resulted in a significant improvement in survival rate compared with controls (100% v 53%, P < .001). In separate experiments, IL-10 at a dose of 25 ng failed to produce any improvement in survival; however, a 50-ng dose resulted in a significant improvement in treated animals compared with controls (95% v 65%, P < .01). CONCLUSIONS: TNF-alpha plays an important role in neonatal sepsis, suggesting that the newborn mouse is capable of mounting a significant proinflammatory response to gram-negative bacteria. Newborn mice may respond to bacterial challenge with an overexpression of proinflammatory cytokines or an underproduction of downregulating cytokines. Future attempts at immunomodulation in human infants must be undertaken with caution until the inflammatory response is better defined. PMID- 10873026 TI - Salvage laparotomy for failure of peritoneal drainage in necrotizing enterocolitis in infants with extremely low birth weight. AB - BACKGROUND/PURPOSE: Peritoneal drainage is a temporizing procedure for infants with extremely low birth weight (ELBW) who have perforated necrotizing enterocolitis (NEC). "Salvage" laparotomy is advocated when patients worsen after drainage. Some patients have survived with intact gastrointestinal functional after drainage alone. The purpose of this study is to determine if these salvage laparotomies are beneficial. METHODS: The authors reviewed the records of ELBW infants treated at Stanford University with perforated NEC from 1993 through 1998. Data collected included demographic makeup, type of operation, survival rate, postoperative complications, length of stay (LOS), and cost. RESULTS: The authors treated 26 patients, 9 with laparotomy and 17 with peritoneal drainage. The peritoneal drainage group had lower birth weight and more comorbid conditions. Survival rate was similar between laparotomy and drainage: 55.6% versus 41.2%. Four patients in the drainage group underwent salvage laparotomy for perceived clinical deterioration. All of these patients died. The clinical status of patients who had salvage laparotomy and died was similar to those who did not and lived. Seven of 13 patients treated with drainage followed only by supportive care and antibiotics survived. Cost and LOS for patients undergoing salvage laparotomy were much greater than for nonsurviving patients undergoing only peritoneal drainage: 84 +/- 20 days and $660,000 compared with 34 +/- 11 days and $306,000. CONCLUSIONS: Both primary peritoneal drainage and laparotomy should be considered primary therapy for perforated NEC. Patients undergoing peritoneal drainage typically experience clinical deterioration after operation. In this limited experience, salvage laparotomy did not appear beneficial. PMID- 10873027 TI - Cognitive deficits in school-age children with severe short bowel syndrome. AB - BACKGROUND/PURPOSE: Improved therapies for the management of short bowel syndrome (SBS) have resulted in the prolonged survival of many children. By early childhood, the physiological sequelae of severe SBS include delayed physical development and metabolic imbalances. However, little is known about how SBS affects brain development. Although many parents report school problems, no controlled study has evaluated the integrity of the central nervous system in SBS children. The purpose of this study was to investigate the neuropsychological status of school-aged SBS children to determine if there were characteristic cognitive impairments that might be amenable to early therapeutic intervention. METHODS: SBS children (n = 8; mean age, 116.9 +/- 21 months) were compared with an age-matched cystic fibrosis (CF) control group (n = 8; mean age, 118.1 +/- 14 months). Groups did not differ in age, grade, or absences. Neuropsychological tests with established sensitivity to CNS integrity compared performance over 6 cognitive domains. Emotional status also was measured. Analyses were completed with 2-tailed t tests. RESULTS: Groups did not differ on tests of intellectual ability and emotional function. Language, memory and learning, and problem solving testing results indicated no significant group differences. However, the SBS group performed more poorly on measures assessing visual-spatial ability, with P values ranging from .002 to .045. In a subset of subjects, we noted significantly slower left-handed, but not right-handed, performance on measures of finger dexterity and psychomotor speed. CONCLUSIONS: Although emotional status did not differ from that of children with CF, SBS patients showed visual-spatial deficits in the company of preserved language, attention and memory, and executive skills. The specificity and consistency of these findings suggests that right hemisphere CNS changes may occur in children with SBS. This unexpected finding, coupled with the indication of left-sided psychomotor slowing in right handed subjects, raises the possibility that actual brain impairment, rather than developmental delay accompanying slowed physical growth, accounts for these findings. Longitudinal studies are needed to further clarify this issue. The educational significance of the results is discussed. PMID- 10873028 TI - Fetal wound repair results in scar formation in interleukin-10-deficient mice in a syngeneic murine model of scarless fetal wound repair. AB - BACKGROUND: Fetal dermal wound healing is characterized by minimal inflammation, restoration of normal dermal architecture, and scarless repair. The authors have shown that proinflammatory cytokines interleukin-6 (IL-6) and interleukin-8 (IL 8) are diminished during fetal wound repair. Interleukin-10 (IL-10) is an antiinflammatory cytokine that decreases production of IL-6 and IL-8. The authors hypothesized that diminished IL-6 and IL-8 and minimal inflammation may be caused by IL-10. METHODS: To test this hypothesis, the authors developed a new syngeneic murine model of fetal wound repair in which 15-day-gestation skin from either normal C57BL/6 or transgenic C57BL/6 IL-10 knockout mice was grafted to the back of the same strain adult mice. The grafts were incisionally wounded after 5 days, harvested at 1 week, and analyzed for inflammatory response and scar formation. RESULTS: Wounds in normal fetal skin grafts showed minimal inflammation and normal dermal reticular collagen pattern at the site of the wound, consistent with scarless repair. In contrast, wounds in IL-10 knockout fetal skin grafts showed significant inflammation and scar formation. CONCLUSIONS: Fetal skin grafts on adult syngeneic mice heal without inflammation or scar formation. The absence of IL-10 in fetal skin results in scar formation. Intrinsic lack of IL-10 may result in continued amplification of the inflammatory cytokine cascade, continued stimulation of fibroblasts, and abnormal collagen deposition. IL-10 is necessary for scarless wound repair to occur. PMID- 10873029 TI - Proinflammatory cytokines differentially regulate hyaluronan synthase isoforms in fetal and adult fibroblasts. AB - BACKGROUND/PURPOSE: Fetal wound healing is a relatively scarless process that occurs in an hyaluronan-rich environment. Understanding the regulation of hyaluronan expression may provide insight into the process of fetal repair. Therefore, the purpose of this study was to compare the regulation of hyaluronan and hyaluronan synthase transcripts by the proinflammatory cytokines interleukin 1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) in human adult and fetal fibroblasts. METHODS: Hyaluronan deposited in the medium of untreated fibroblasts or fibroblasts treated with either IL-1beta or TNF-alpha was determined by an assay utilizing iodine I 125-hyaluronan binding protein. HAS transcript levels were compared in using a ribonuclease protection assay. RESULTS: IL-1beta induced an increase in hyaluronan accumulation by both fetal and adult fibroblasts. In contrast, TNF-alpha induced higher levels of hyaluronan only in fetal fibroblasts. HAS-2 and HAS-3 transcript levels were constitutively expressed by both fetal and adult fibroblasts. Proinflammatory cytokines induced a differential increase in HAS-1 and HAS-3 transcript levels. CONCLUSIONS: Differential regulation was observed in hyaluronan accumulation and for HAS transcript levels in fetal and adult dermal fibroblasts. The muted response of fetal fibroblasts to cytokines may be relevant to the minimal inflammation associated with fetal repair. PMID- 10873030 TI - The role of preoperative chemotherapy in the treatment of infantile fibrosarcoma. AB - Infantile fibrosarcoma (IFS) is a rare tumor most often affecting the extremities of infants and young children. Unlike its adult counterpart, IFS has a low potential for metastatic spread, and surgical extirpation alone has therefore resulted in an excellent prognosis. The amputation rate, however, exceeds 50%. The dramatic response in 2 recent cases to preoperative chemotherapy, given in an attempt to avoid amputation, prompted this report and a review of the literature. PMID- 10873031 TI - Receptor tyrosine kinase inhibition suppresses growth of pediatric renal tumor cells in vitro. AB - PURPOSE: Children who undergo standard therapy for renal tumors are at an increased risk for treatment sequelae such as congestive heart failure, abnormal trunk development, and secondary malignancies. Therefore, research on the use of novel chemotherapeutic agents with fewer side effects is justified. Recent experimental evidence suggests that growth factor receptors such as epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor (PDGFR) play an important role in growth and development of pediatric renal tumors especially that of Wilms' tumor. In this study we investigated the effects of genistein, AG1478, and AG1295, from the class of growth factor receptor tyrosine kinase (GFR-TK) inhibitors, on proliferation and colonigenic growth of 2 pediatric renal tumor cell lines. METHODS: The authors studied the effect of genistein (broad-spectrum GFR-TK inhibitor), AG1478 (EGFR-specific GFR-TK inhibitor), and AG1295 (PDGFR-specific GFR-TK inhibitor) on proliferation and colonigenic growth of rhabdoid tumor of the kidney and Wilms' tumor cell lines: G 401 and SK-NEP-1, respectively. The effect of genistein at concentrations of 0 to 200 micromol/L, and AG1478 and AG1295 at 0 to 10,000 nmol/L were tested on proliferation by using a growth inhibition assay. Viable cell counts at each concentration were obtained by hemocytometer and trypan blue exclusion, and percent growth inhibition was calculated based on control cultures at the same time-point. As a measure of colonigenic survival, the percent inhibition of colony formation in drug-treated dishes was calculated based on the number of colonies (>50 cells) in control dishes. RESULTS: Genistein at concentrations of 25 and 50 micromol/L inhibited the colonigenic growth of G-401 by 37% and 79% (P = .01 and 5E-06, 2-tailed t test, respectively) and that of SK-NEP-1 by 44% and 74% (P = .0001 and 9.9E-07). The mean percent growth inhibition at the above doses was 57% +/- 7.9% and 96% +/- 0.2% for G-401, and 47% +/- 11.2% and 60% +/- 2.7% for SK-NEP-1. AG1478 at concentrations of 1,000 and 5,000 nmol/L inhibited the colonigenic growth of G401 by 75% and 78% (P = .0005 and 7.38E-06, respectively) and that of SK-NEP-1 by 19% and 40% (P = .02 and .0001). The percent growth inhibition at the mentioned concentrations for G-401 were 53% +/- 9.3% and 63% +/- 6.3%, and for SK-NEP-1 were 55% +/- 14.5% and 65% +/- 20.1%, respectively. AG1295 did not appear to be as effective as AG1478. CONCLUSIONS: This is the first experimental study on the use of GFR-TK inhibitors as a potential treatment for pediatric renal tumors. GFR-TK inhibitors such as genistein occur naturally in soybean foods and have been shown to reach therapeutic levels in blood after consuming a soybean-based diet. Considering the significance of growth factor receptor activity in Wilms' tumor development, inhibition of GFR-TKs should be investigated as effective and potentially nontoxic adjunctive treatment for this childhood tumor. Furthermore, GFR-TK inhibitors may offer an effective alternative to the treatment of commonly fatal rhabdoid tumor of the kidney in children. PMID- 10873032 TI - 16q loss of heterozygosity and microsatellite instability in Wilms' tumor. AB - BACKGROUND/PURPOSE: Wilms' tumor is the most common renal malignancy of childhood. Loss of heterozygosity (LOH) at 16q is seen in about 17% of cases and has been associated with a poor prognosis. To more precisely define the pattern of 16q deletion exhibited by Wilms' tumor, the authors performed a detailed LOH analysis of 96 specimens using polymorphic microsatellite repeat markers. The authors also evaluated the neoplasms for the presence of microsatellite instability (MSI). METHODS: A total of 96 DNA samples were studied using polymerase chain reaction-based LOH analyses amplifying polymorphic microsatellite repeat markers. Screening for MSI using 2 additional genetic markers also was carried out. RESULTS: The authors found 16q LOH in 14 of the specimens evaluated. Comprehensive analysis of these LOH-positive specimens showed a region of loss spanning 16p11.2-q22.1 and a separate distal region of LOH at 16q23.2-24.2. The distal region of deletion is very small, estimated to be approximately 2.4 megabases. In addition to the observed LOH, 2 specimens were found to consistently exhibit MSI, which has not been reported previously in Wilms' tumor. CONCLUSIONS: The smallest consensus region of deletion in our analysis of Wilms' tumor 16q LOH measures 2.4 megabases at 16q23.2-q24.2. Additionally, MSI was present in a subset of tumor specimens suggesting that defects in DNA mismatch repair may contribute to the pathogenesis of Wilms' tumor. PMID- 10873033 TI - Effect of dexamethasone on insulin-like growth factor-1 expression in a rabbit model of growth retardation. AB - BACKGROUND/PURPOSE: The maternal administration of steroids promotes fetal maturative effects in the gastrointestinal tract. To determine if fetal insulin like growth factor-1 (IGF-1) expression is altered in response to maternal dexamethasone administration, this rabbit model of intrauterine growth retardation (IUGR) was utilized. METHODS: Eight pregnant rabbits received either dexamethasone (Dex 0.1 mg/kg/d intramuscular), or normal saline (Cont) on gestational days 26 and 27. Fetuses were harvested on gestational day 28 or 29 and were identified as favored (Fav) or runt (Runt): DexFav, DexRunt, ContFav, and ContRunt. Fetal weight was recorded and the serum, amniotic fluid, liver, kidney, and small intestine (SI) were collected. Reverse transcription polymerase chain reaction (RT-PCR) was used to measure IGF-1/beta-actin mRNA densitometric band ratios in all tissues. Radioimmunoassay (RIA) was used to measure IGF-1 protein levels in the serum and amniotic fluid. RESULTS: Weight was decreased in the Runt fetuses at all time-points (P < .08). The percent weight accretion from day 28 to 29, was greatest in the DexRunt fetus (P < .001), suggesting "catch-up" growth. All Dex fetuses (Fav and Runt) had increased liver and proximal, middle and distal SI IGF-1 mRNA at day 28 and elevated levels in the liver, proximal and distal SI at day 29 compared with control fetuses. The DexRunt fetuses had serum IGF-1 protein surpassing that of the DexFav fetus at day 28. CONCLUSIONS: This report provides the first description of maternal steroid administration effecting a marked increase in fetal IGF-1 mRNA expression and IGF-1 protein levels in an in vivo rabbit model of IUGR. The growth-retarded fetus appears to be particularly responsive. PMID- 10873034 TI - Stretch-induced upregulation of VEGF gene expression in murine pulmonary culture: a role for angiogenesis in lung development. AB - BACKGROUND/PURPOSE: The authors' have shown pulmonary alveolarization (capillary and alveolar growth) both after fetal tracheal occlusion and postnatal pulmonary distension. The trophic and developmental mechanisms responsible for this growth remain largely unknown; however, experimental systems have defined an enhanced expression of angiogenic proteins in response to tissue stretch. The authors hypothesize that the stimulation of pulmonary alveolarization after stretch is secondary to upregulation of the potent endothelial cell mitogen vascular endothelial growth factor (VEGF) and that the endothelial cell represents the central stimulus of parenchymal growth. METHODS: A mixed primary pulmonary cell culture obtained by enzymatic digestion of fetal, neonatal, and adult mouse lung was plated on Bioflex elastomer bottom plates, grown to confluence, rendered quiescent, and subjected to continuous cycles of stretch-relaxation with nonstretched cells as controls. Cells were harvested at time-points 0, 30 minutes, 2 hours, 4 hours, 8 hours, and 24 hours. RNA was extracted and VEGF gene expression analyzed by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). Similar cell groups were harvested, processed, and analyzed utilizing Western Blot techniques. VEGF PCR of mRNA isolated from fetal sheep subjected to surgical creation of diaphragmatic hernia both with (DH-TL) and without (DH) tracheal ligation also was analyzed. RESULTS: VEGF mRNA isoforms 120, 164, and 188 showed increased expression in all stretched groups, which was noted by 30 minutes with maximal expression seen at 2 to 4 hours and a return to baseline expression by 24 hours. VEGF protein was similarly elevated in all stretched cell groups. In preliminary studies, DH/TL sheep showed upregulation of VEGF compared with DH sheep alone. CONCLUSIONS: These data show in an in vitro system that "pulmonary stretch" upregulates VEGF mRNA and protein expression supporting the role of angiogenesis in the stretch-induced pulmonary alveolarization. The authors speculate that such angiogenic activity is a rate limiting factor in stimulating alveolar epithelial development, and as a treatment modality, therapeutic angiogenesis may provide a noninvasive method with which to treat pulmonary hypoplasia. PMID- 10873035 TI - In vitro effects of growth factors on lung hypoplasia in a model of congenital diaphragmatic hernia. AB - BACKGROUND/PURPOSE: Pulmonary hypoplasia, a leading contributor to the lethality of congenital diaphragmatic hernia (CDH), precedes diaphragmatic malformation in the nitrofen model and persists to allow experimental manipulations in organ culture. Fibroblast growth factors (FGFs) are crucial to early lung development. Acidic FGF (FGF-1) binds to all FGF receptors and enhances in vitro branching morphogenesis. Basic FGF (FGF-2) is localized to developing airway epithelium, basement membrane, and extracellular matrix. Heparin (HEP) modulates FGF kinetics and inhibits smooth muscle proliferation in lung primordia. The aim of this study was to examine the morphological effects of fibroblast growth factors and heparin on lung hypoplasia in an organ culture model. METHODS: Sprague-Dawley rats were fed nitrofen on day 9.5 of pregnancy to induce lung hypoplasia and CDH in newborns. Control rats received olive oil. Normal and hypoplastic lung primordia were microdissected on day 13.5 of gestation and cultured up to 78 hours in plain media with or without FGF-1 or FGF-2, with or without HEP. In vitro morphological development was studied by serial measurements of terminal bud count, lung area, and lung perimeter. RESULTS: Over 120 fetal lung specimens were studied (n > or = 4 per group). Significant increases in area, perimeter, and bud count were seen in normal lungs cultured with FGF-1 plus HEP compared with control media (P < .05). In the nitrofen lungs, FGF1 plus HEP yielded reductions in all parameters compared with those in control media (P < .05), whereas FGF-2 produced significant expansion in lung area but marked reductions in bud count and lung perimeter divided by square root of area (P < .05). Heparin did not produce substantial or sustained alteration of morphology in normal or hypoplastic lungs. CONCLUSIONS: These observations may indicate an intrinsic abnormality of FGF processing in the hypoplastic nitrofen lung before diaphragmatic malformation. Heparin did not rescue abnormal lung development. Mechanisms underlying the differential effects of these agents now need to be explored to target fetal lung growth and improve the dismal prognosis of human CDH. PMID- 10873036 TI - Perforated appendicitis: prospective outcome analysis for 150 children. AB - BACKGROUND/PURPOSE: Controversy persists in the management of perforated appendicitis with regard to antibiotic choice and duration, operative timing, drain utilization, and wound closure. For 2 decades at the authors' institution, patients were treated with ampicillin, gentamicin, and clindamycin for 10 inpatient days, with drains in the abdomen, resulting in lower complication rates than most other published series. Managed care pressures have led to less aggressive medical management regimens with length of stay and financial factors viewed as principal outcome measures with little emphasis on clinical outcomes. In addition, there are little prospective data on clinical outcomes. The authors sought to determine whether our previously documented excellent quality outcomes could be maintained when modifications aimed at decreasing cost and length of stay in our protocol were instituted. METHODS: The authors monitored prospectively clinical outcomes in patients with perforated appendicitis treated according to their clinical practice guidelines over a 43-month period. Patients received a single antibiotic, piperacillin-tazobactam, intravenously for 10 days. They were permitted to go home with a percutaneous intravenous catheter for the final 5 days if medical and social criteria were met. Other practices from our earlier protocol were continued, including immediate operation, placement of Penrose drains, and primary wound closure. RESULTS: Of 150 patients treated on our protocol, major complications included intraabdominal abscess in 5 (3.3%), cecal fistula in 2 (1.3%), phlegmon in 3 (2.0%), wound infection in 4 (2.7%), and no small bowel obstructions requiring operation. None of these complications, nor their aggregate, were significantly more common than those reported in 373 patients treated over 11 years on the authors' prior protocol (chi2, P > .05). CONCLUSIONS: Prospective outcome analysis of our protocol shows that a single broad-spectrum antibiotic (allowing portions of therapy to be delivered less expensively on an outpatient basis) effectively can treat postoperative appendicitis with very few infectious complications. These outcome data provide baseline against which future protocols can be compared. All treatment modifications aimed at decreasing costs must be analyzed to ensure quality of care is not unduly compromised. PMID- 10873037 TI - Laparoscopically assisted anorectal pull-through for high imperforate anus--a new technique. AB - BACKGROUND/PURPOSE: This report describes a new technique of laparoscopically assisted anorectal pull-through (LAARP) for repair of high imperforate anus. The procedure utilizes minimal perineal dissection, preservation of the distal rectum, and accurate placement of the rectum within the levator ani and external anal sphincter muscle complex. METHODS: Sharp dissection and cautery was used laparoscopically to expose the rectal pouch down to the urethral or vaginal fistula, which was clipped distally and divided. The pelvic floor musculature was then assessed and the levator sling identified. Externally, electrostimulation was used to define the center of the anal dimple. An 8-mm skin incision was made, centered at the strongest cephalad contraction. Using a hemostat, minimal blunt dissection on the perineum was guided by transillumination from the laparoscopic light source. A trocar, consisting of a radially expandable sheath over a Varess needle, was passed through this defined plane in the external sphincter muscle complex and advanced into the pelvis between the 2 bellies of the pubococcygeus muscle, guided by laparoscopic visualization. This perineal trocar therefore formed a passage through the center of the striated muscle complex and levators. The rectal fistula, which had been dissected out laparoscopically, was grasped using the perineal trocar and exteriorized to the perineum. Anorectal anastomosis was performed with absorbable interrupted suture. RESULTS: Seven patients were treated with initial colostomy in the newborn period followed by delayed LAARP 2 to 12 months later. In 4 newborn infants, the LAARP was performed as a primary procedure without prior colostomy. Laparoscopic mobilization has been possible on all cases attempted. All of the patients have a brisk and symmetric anal contraction with perineal electrostimulation. CONCLUSIONS: Lack of long-term follow-up precludes accurate assessment of the potential for fecal continence. However, short-term experience has been that this new method of pull-through for imperforate anus offers many advantages, including excellent visualization of the rectal fistula and surrounding structures, accurate placement of the bowel through the anatomic midline and levator sling, and minimally invasive abdominal and perineal wounds. PMID- 10873038 TI - Cloacal exstrophy: a unified management plan. AB - BACKGROUND/PURPOSE: The belief that patients with cloacal exstrophy have a short and therefore useless colon is all too common. Frequently, the colon is used for urinary or vaginal reconstruction, and the possibility of a pull-through is lost. In the authors' experience, the use of a unified management plan allowed most patients to undergo pull-through and avoid a permanent stoma. METHODS: Twenty five patients were treated for cloacal exstrophy in the authors' institution from 1985 through 1999. In all patients, bladder closure, omphalocele repair, and creation of a colostomy were performed at birth. All available colon, no matter how small, was incorporated into the fecal stream. After at least 1 year, patients were assessed for the ability to form solid stool through their stoma. Normal colonic length, capacity to form solid stool, or success with a bowel management regimen through the stoma were considered indications for pull through. Genitourinary reconstruction was contingent on the colorectal plan. RESULTS: Colonic length ranged from normal in 12 patients, 40 to 70 cm in 3 patients, 10 to 30 cm in 4 patients, and less than 10 cm in 2 patients. All 25 patients underwent pull-through. Three are totally continent, 4 are continent with occasional soiling, 11 remain clean with a bowel management regimen, and 4 are too young to assess. One patient was clean, but now refuses bowel management. Two early patients, both with less than 10 cm of colon, now have ileostomies. CONCLUSIONS: During neonatal repair, a colostomy should be formed incorporating all pieces of colon, no matter how small. With time, most patients will be able to form solid stool, and a pull-through should be undertaken if that ability exists. Decisions regarding genitourinary reconstruction should be made only after the gastrointestinal plan is established to achieve the optimal use of available bowel. PMID- 10873039 TI - The nonoperative management of fistula-in-ano. AB - BACKGROUND/PURPOSE: Fistulotomy is the accepted treatment for infants with perianal fistula. Although recurrence rates range from 0% to 68%. Based on the experience of a senior colleague who noted that babies suffering from perianal fistula follow a self-limited course the authors decided to determine if this observation was accurate. METHODS: A conservative approach to perianal abscess and fistula was used prospectively in 18 male infants. Abscesses were to be drained only if the baby was very uncomfortable or febrile. Once a fistula developed the authors continued observation until the fistula healed. Data are expressed as mean +/- SD. Mean follow-up period was 37 months. RESULTS: Mean age at onset of symptoms was 4 +/- 3 months. Fistulas developed in 14 patients (77%). All fistulas healed without operation. Four patients had abscesses drained for discomfort (n = 3) or fever (n = 1). No patient required antibiotics. Mean duration of symptoms was 6 +/- 4 months. Four patients in whom fistulas did not form healed after incision (n = 3) or spontaneous drainage (n = 1). All patients currently are asymptomatic. CONCLUSIONS: In healthy neonates, perianal abscess and fistula are self-limited conditions rarely requiring surgical drainage and not requiring antibiotics. The conservative management of perianal abscess and fistula in healthy infants appears to be safe and effective. PMID- 10873040 TI - Characterization and treatment of biliary anastomotic stricture after segmental liver transplantation. AB - BACKGROUND/PURPOSE: Biliary anastomotic strictures (BAS) after left lateral segment liver transplantation (LLST) may cause graft dysfunction, sepsis, and patient mortality. A review of the authors' experience was performed to better characterize the risk factors and corrective management. METHODS: The medical records of 9 children who underwent a LLST in whom a BAS developed from 1989 to the present were reviewed retrospectively. RESULTS: Seventy-five of 199 liver transplants (38%) at the authors' institution since 1989 have been LLST. BAS developed in 12% of these cases. BAS were diagnosed less than 12 months after transplantation in 4 children (mean, 7.5 months; range, 5 to 11 months) and greater than 12 months in 5 children (mean, 37 months; range, 14 to 72 months). Early strictures (<12 months) were associated with hepatic artery thrombosis (n = 1), and posttransplant bile leak (n = 1) and ducts from segment II and III exiting separately from the left lateral segment (n = 2). The diagnosis of BAS was heralded by episodes of liver biopsy-proven cholangitis in all patients and confirmed radiographically. Seven children underwent successful biliary exploration and revision of the hepaticojejunostomy. Two of these children ultimately required retransplantation secondary to chronic graft rejection. CONCLUSIONS: BAS in LLST are a source of significant morbidity and should be considered in children after LLST who present with cholangitis. Surgical correction is possible in most cases. PMID- 10873041 TI - Percutaneous cannulation for pediatric venovenous extracorporeal life support. AB - BACKGROUND/PURPOSE: The objective of this study was to show the safety and efficacy of a method of percutaneous cannulation for venovenous extracorporeal life support (ECLS) access in nonneonatal (>10 kg) pediatric patients. METHODS: Between June 1992 and October 1998, 26 pediatric patients (age range, 3 to 17 years; weight range, 19 to 100 kg) underwent attempted percutaneous cannulation for venovenous ECLS at our institution. Venous drainage access was attempted using a modified Seldinger technique via the right internal jugular vein (RIJ, n = 22) or right femoral vein (RFV, n = 4). Reinfusion access was attempted via the RFV (n = 19), RIJ (n = 4), or left femoral vein (n = 3). RESULTS: The percutaneous technique was successful in 24 of 26 patients (92.3%). Maximum blood flow during ECLS was 80.1 +/- 30.0 mL/kg/min, generating a postmembrane lung outlet pressure of 138 +/- 54.8 mm Hg. Adequate gas exchange was achieved in all patients, and survival to discharge was 79.2%. There was no procedure-related mortality. Complications potentially related to the percutaneous technique included RIJ thrombosis (n = 1) detected after decannulation and cannula site bleeding (n = 3). CONCLUSION: Percutaneous access may be used safely and effectively for venovenous ECLS in pediatric patients. PMID- 10873042 TI - Nonrhabdomyosarcoma soft tissue sarcomas in children: is age at diagnosis an important variable? AB - PURPOSE: The associations between age at diagnosis, tumor characteristics, and outcome in children diagnosed with nonrhabdomyosarcoma soft tissue sarcoma (NRSTS) were studied. METHODS: Retrospective review was conducted of 192 children from 1962 through 1996. Patients were divided into groups: birth to 1 year (n = 13), 1 to 5 years (n = 26), 5 to 10 years (n = 49), 10 to 15 years (n = 55), and older than 15 years (n = 49) of age at diagnosis. Characteristics including IRS group, histological grade and pattern, tumor size, and invasiveness were investigated. Survival rate was estimated by age group. The median follow-up was 8.8 years (range, 2 to 28 years). RESULTS: There were 81 group I patients, 40 group II, 41 group III, and 30 group IV. A significant difference of IRS groups among the age groups was seen (P = .034). There were no IRS group IV patients less than 1 year of age; 50% of IRS group IV patients were older than 15 years. A significant difference in the distribution of histological grade among the age groups (P = .032) was seen. Ten of 13 (77%) children less than 1 year of age had low-grade tumors, whereas 42%, 45%, 60%, and 37% of patients aged 1 to 5, 5 to 10, 10 to 15, and older than 15 years, respectively, had low-grade tumors. Patients older than 15 years had the highest incidence of invasive tumors (59%). Histological pattern also varied with age. The most prevalent histology in the less-than-1-year age group was infantile fibrosarcoma. No predominant histology was seen in the 1- to 5-year age group. Malignant fibrous histiocytoma was the most frequent histological subtype in children between 5 and 10 years of age. In the 10- to 15-year age group and children older than 15 years the malignant peripheral nerve sheath tumor and synovial sarcoma were the most prevalent subtypes. Without adjusting for any other factors, age group was prognostic of survival (P = .007). Patients less than 1 year at diagnosis had the best outcome, with a 5-year survival rate of 92% +/- 9%. Five-year estimates were lowest for patients older than 15 years (49% +/- 7%). CONCLUSIONS: Significant differences in IRS group, histological grade, and histological subtype were observed in different age groups. Infants with NRSTS were more likely to have low grade, less invasive, and lower stage tumors. These characteristics may account for their improved prognosis. PMID- 10873044 TI - Lymphatic mapping with sentinel node biopsy in pediatric patients. AB - BACKGROUND/PURPOSE: Lymphatic mapping with sentinel node biopsy is used widely in adult melanoma and breast cancer to determine nodal status without the morbidity associated with elective lymph node dissection. This technique can be used in children to determine lymph node status with limited dissection and accurate interpretation. The authors report their initial experience. METHODS: The charts of patients who underwent lymphatic mapping with sentinel node biopsy were reviewed retrospectively. Lymphoscintigraphy was performed in patients with truncal lesions 24 hours before surgery to determine the draining nodal basin (for surgical mapping). The tumors were injected 1 hour preoperatively with technetium sulfur colloid and in the operating room with Lymphazurin blue. The draining basin was examined using a radioisotope detector. The blue nodes with high counts were localized and removed. If nodal metastases were identified, lymph node dissection was recommended. Four patients were injected only with Lymphazurin blue. RESULTS: Thirteen children (7 girls, 6 boys; mean age, 7 years) underwent lymphatic mapping with sentinel node biopsy. The tumor types were as follows: 8 malignant melanoma (6 extremity, 2 truncal), 1 malignant peripheral nerve sheath tumor, 1 alveolar soft part sarcoma, and 3 rhabdomyosarcoma. A mean of 2.4 nodes (range, 1 to 6) were removed from each patient. Six patients had a positive sentinel node. Formal lymph node dissection was performed on 4 of the 6 patients, 1 of whom had further nodal disease with 2 of 13 nodes containing micrometastases. One of the 6 patients refused lymph node dissection and adjuvant therapy; the final patient had rhabdomyosarcoma, a malignancy for which lymph node dissection is not indicated. Pulmonary metastasis developed 26 months after diagnoses in the patient with alveolar soft part sarcoma and a negative sentinel node. This patient was injected only with Lymphazurin blue at the time of sentinel node biopsy and refused adjuvant therapy. There have been no other recurrences. There were no complications related to lymphatic mapping or sentinel node biopsy. CONCLUSIONS: Lymphatic mapping with sentinel node biopsy, using both technetium-labeled sulfur colloid and Lymphazurin blue, can be performed safely in pediatric skin and soft tissue malignancies. Further study with long-term follow-up will determine the utility and accuracy of this technique in pediatric malignancies. PMID- 10873043 TI - Differentiated thyroid cancer: clinical characteristics, treatment, and outcome in patients under 21 years of age who present with distant metastases. A report from the Surgical Discipline Committee of the Children's Cancer Group. AB - BACKGROUND/PURPOSE: Young patients with differentiated thyroid cancer typically present with regional lymph node involvement (60% to 80%), and 10% to 20% have distant metastases. This study characterizes the clinical presentation, treatment, and outcome in patients with differentiated thyroid cancer who were less than 21 years of age at diagnosis and who presented with distant parenchymal metastases. METHODS: A retrospective, multi-institutional data base that included 327 patients in this age group with differentiated thyroid carcinoma was searched for patients who presented with distant metastases, and 83 cases (25%) were found. The median time to first disease progression was 2.4 years (range, 0.1 to 12.4 years) and the overall median follow-up was 10.9 years (range, 1.0 to 42.1 years). RESULTS: The median age at diagnosis was 14.6 years (range, 6.6 to 20.8 years); 69% were girls and 92% were white. In 12%, there was a history of prior head and neck irradiation, and 10% of these patients had a family history of carcinoma. Preoperative needle biopsies were performed in 25%. Regional lymph nodes were positive in 90%, and extrathyroidal extension occurred in 48%. The site of distant metastases included the lungs in all patients. Total thyroidectomy, subtotal thyroidectomy, lobectomy, and nodule excision was done in 66%, 24%, 3%, and 8% of patients, respectively. There was no residual cervical disease after surgery in 75%, whereas 14% had microscopic and 11% had gross residual. Histopathologic subtypes included papillary-follicular (48%), papillary (42%), and follicular (10%). The median tumor size was 3.0 cm (range, 0.4 to 11.0 cm). In this group, 100% of patients received adjuvant iodine 131I therapy, and the overall survival rate at 10 years was 100%. The progression-free survival rate was 76% at 5 years and 66% at 10 years from diagnosis. CONCLUSIONS: A significant number of young patients with thyroid cancer present with distant metastases and will require radioiodine therapy. This should be considered when planning the surgical approach because total or subtotal thyroidectomy facilitates 131I imaging and treatment. Although about one third of these patients will experience relapse or disease progression, the overall mortality rate is low. PMID- 10873045 TI - Sentinel lymph node biopsy for melanoma in young children. AB - Lymphoscintigraphy and sentinel lymph node biopsy techniques can be applied successfully to young children with melanoma to detect nodal disease. The authors describe their methods of lymphoscintigraphy and sentinel node biopsy and its application in 2 young children with malignant melanoma of the head and neck. PMID- 10873046 TI - The response to splenectomy in pediatric patients with idiopathic thrombocytopenic purpura who fail high-dose intravenous immune globulin. AB - BACKGROUND: A recent article by Law et al concluded that patients with idiopathic thrombocytopenic purpura (ITP) who have a poor response to intravenous immune globulin (IgG) are unlikely to have a good or excellent response to surgical splenectomy. METHODS: The authors studied retrospectively 23 pediatric patients age 11.7 +/- 1.0 years with ITP who had been treated with IgG before undergoing splenectomy. As in the aforementioned article, the responses to the 2 treatments were classified on the basis of the platelet count as poor (<50,000/mm3), good (50,000 to 150,000/mm3), or excellent (>150,000/mm3). For patients who received multiple IgG treatments, both initial and final treatment responses were analyzed. RESULTS: Sixteen patients had an excellent or good initial response to IgG. Of these 16 patients, 14 had an excellent or good response to splenectomy. Among the 7 patients who had a poor response to IgG there were 3 who had an excellent or good response to splenectomy (43%), and 4 patients who had a poor response to splenectomy. A good or excellent response to initial treatment with IgG was associated with a significant probability of a good or excellent response to splenectomy (P = .045). CONCLUSIONS: A good or excellent response to IgG may be predictive of a favorable response to splenectomy. However, a poor response to IgG does not preclude a satisfactory response to splenectomy in pediatric patients with ITP. PMID- 10873047 TI - The effects of prebiopsy corticosteroid treatment on the diagnosis of mediastinal lymphoma. AB - BACKGROUND/PURPOSE: For children with probable mediastinal lymphoma and a high risk of cardiorespiratory morbidity, many centers recommend delaying the diagnostic biopsy for 24 to 48 hours while corticosteroids are administered to reduce tumor size and morbidity. This study was undertaken to determine the effect of preoperative steroid use on the accuracy of the pathological diagnosis and incidence of perioperative cardiorespiratory morbidity. METHODS: From 1988 to 1998, 86 children were treated for mediastinal lymphoma. Twenty-three received steroid before biopsy (study group) because of clinical evidence of respiratory compromise, and the remaining 63 served as controls. Clinical parameters, steroid use, and detailed pathological findings obtained at initial and subsequent biopsies were reviewed. Steroid treatment was considered to have had an adverse effect on the pathological diagnosis if (1) a definitive diagnosis was delayed more that 1 month, (2) a definitive diagnosis could not be made, or (3) the extent of disease could not be staged with certainty. RESULTS: Steroid treatment had an adverse effect on the pathological diagnosis in 5 of 23 (22%) children: 1 diagnostic delay, 3 failures of a definitive diagnosis, and 1 possible failure of staging. A definitive diagnosis was made in all control patients. Perioperative survival was 100% in both groups. At biopsy, only 3 children in the steroid treatment group and 2 children in the control group had moderate, nonfatal cardiorespiratory instability. Parameters of steroid use among children who had inaccurate pathological diagnoses or cardiorespiratory morbidity were not significantly different from those who did not. CONCLUSIONS: Steroid treatment before biopsy of mediastinal lymphoma may adversely affect the pathological accuracy or cause a delay in definitive diagnosis in a minority of cases. The dose and duration of steroid use was not related to outcome. Prebiopsy steroid appears to minimize the likelihood of cardiorespiratory morbidity in high-risk patients. PMID- 10873048 TI - Suppression of primary tumor growth in a mouse model of human neuroblastoma. AB - BACKGROUND/PURPOSE: Neuroblastoma is the most common tumor of the abdomen in children. Consistently effective treatments are lacking for aggressive disease. The authors previously reported that therapy with anti-vascular endothelial growth factor (VEGF) antibodies suppresses both growth and metastasis in an experimental model of Wilms' tumor. The authors hypothesized that, in a parallel model of neuroblastoma, anti-VEGF treatment would inhibit (1) growth and (2) metastasis. METHODS: Primary tumors were established in the kidneys of nude mice. In cohort 1 (n = 42), mice were killed at 3 time-points, and tissues were evaluated histologically. Tumors were assayed for VEGF. In cohort 2 (n = 28), anti-VEGF antibody or vehicle was administered. Tumor weights and the incidence of metastases in the 2 groups were compared. VEGF deposition was evaluated by immunohistochemistry. RESULTS: Mice displayed large tumors with liver and lung metastases. VEGF levels in tumors increased over time. Antibody-treated animals displayed significantly smaller tumors, but incidence and size of metastases were unaffected. VEGF was localized to tumor stroma immunohistochemically, with no difference in pattern observed in control and antibody-treated tumors. CONCLUSIONS: Anti-VEGF antibodies inhibit primary tumor growth in experimental neuroblastoma, but not metastasis. This may contrast with the effect of the same antibody in a parallel model of Wilms' tumor. PMID- 10873049 TI - Endorectal pull-through abates gastrointestinal hemorrhage from colorectal venous malformations. AB - BACKGROUND/PURPOSE: Lower intestinal venous malformations are rare anomalies resulting from errors in vascular morphogenesis. These lesions may cause significant chronic and acute gastrointestinal hemorrhage. Venous malformations are unresponsive to angiogenesis inhibitors. Although these anomalies generally are incompletely resectable because of diffuse pelvic and mesenteric involvement, the authors sought to abate bleeding by excluding the lesion from the gastrointestinal lumen. METHODS: Three patients with circumferential transmural venous malformations of the colorectum, pelvis, and mesentery were identified. Imaging findings were similar among the patients and included circumferential septated bright signal on T2-weighted magnetic resonance imaging (MRI) contrast enhancement, and multiple phleboliths, seen best on computed tomography (CT). The lesion extended from the anus to the splenic flexure in 2 patients and throughout the entire colorectum in the other. Each had daily hematochezia for many years and required transfusions and chronic iron therapy. Although bleeding began in childhood in each patient, no therapy was successful until ages 7, 24, and 45. Colectomy, anorectal mucosectomy (through the pelvic venous malformation), and endorectal pull-through and anastomosis was performed (coloanal in 2 and ileoanal in 1). RESULTS: Bleeding essentially has been eradicated in all 3 patients with 10- to 57-month follow-up. One patient received a 3-unit transfusion intraoperatively, and the other 2 received none. The most recent patient to undergo surgery, who has residual venous malformation in the remaining 1 cm of anal mucosa, has some mild difficulty with fecal control if her diet results in loose stool. CONCLUSION: Colectomy with mucosectomy and endorectal pull-through should be considered for diffuse venous malformations of the colorectum before the development of large transfusion requirements. PMID- 10873050 TI - Changing patterns of treatment for blunt splenic injuries: an 11-year experience in a rural state. AB - PURPOSE: The aim of this study was to perform a population-based study evaluating the trend in management of pediatric blunt splenic injuries in a rural state and assess differences in the management of those injuries at a level I pediatric trauma center (PTC) and regional hospitals (RH) from 1985 through 1995. METHODS: ICD-9-CM diagnosis and procedure codes for children (age less than 19) discharged from all hospitals in a rural state with splenic injuries from 1985 through 1995 were reviewed. Hospital charges, age, and nonoperative management (NOM) rates were calculated for PTC and RH and compared using chi2 and linear regression. (P < .05 is statistically significant.) Patients were divided into 2 groups; G1, 1985 through 1989 (127 children); G2, 1990 through 1995 (140 children). RESULTS: The overall NOM rate increased from 21% (G1) to 64.2% (G2), P < .001. A total of 114 patients were treated at PTC and 153 patients received care at RH. PTC had a NOM rate of 54.3% versus 35.9% at RH (P = .003). There was no statistical difference in ages or ISS within the groups or between PTC and RH. NOM in RH rose from 7.7% in G1 to 56.9% in G2 (P < .000), and from 35.5% in G1 to 76.9% in G2 (P < .001) for PTC. Hospital charges were lower for patients receiving NOM versus those with surgical treatment of their injury, $8,094 versus $10,862 (P = .018). However, a higher percentage of children were treated at RH than PTC in G2 versus G1 (68.2% v 51.2%, P = .0541). CONCLUSIONS: Over the 10-year period studied, the NOM rate for splenic injuries significantly decreased. This trend was seen at both the PTC and RH, but the PTC maintained a higher rate of NOM. Unfortunately, more children were treated at RH in G2. Educational programs increased NOM in RH but not to a level equal to PTC. These programs had the negative effect of allowing more children to be treated at RH, actually increasing the splenic operation rate for this population. PMID- 10873051 TI - Free fluid on abdominal computed tomography scan after blunt trauma does not mandate exploratory laparotomy in children. AB - BACKGROUND/PURPOSE: The isolated finding of free intraperitoneal fluid on abdominal computed tomography (CT) scan after blunt trauma in adults is considered an indication for laparotomy by many trauma surgeons. The authors wished to determine if these guidelines are applicable to children. METHODS: A retrospective chart review was conducted. The authors included all children (< or =12 years of age) sustaining blunt abdominal trauma who were admitted to our institution between January 1, 1994 and November 1, 1998. RESULTS: There were 814 children admitted, and 437 had abdominal CT scans. Thirty-four studies showed free fluid associated with solid organ injuries, spine or pelvic fractures, or pneumoperitoneum, and were excluded. Thirty-two children had free fluid without associated injuries and formed the basis for the study. Five of these children underwent laparotomy based on the CT finding alone. The remaining 27 were observed with serial abdominal examinations and did not require surgical intervention. Only 1 of the 5 children who underwent surgery for the finding of isolated free fluid had a therapeutic laparotomy. In comparison, during the same period, 38 children underwent laparotomy after blunt injury based only on physical examination findings with a therapeutic laparotomy rate of 68%. The therapeutic laparotomy rate was significantly higher when the procedure was based solely on clinical examination as compared with the isolated finding or free fluid on the abdominal CT (26 of 38 v 1 of 5, P < .05). CONCLUSION: In contrast to adults, finding isolated free fluid on abdominal CT scans in children after blunt trauma does not dictate immediate surgical exploration. PMID- 10873052 TI - The pediatric trauma C-spine: is the 'odontoid' view necessary? AB - BACKGROUND/PURPOSE: The "odontoid view" is a difficult and often hazardous film to obtain in young children. The aim of this study was to determine if the transoral roentgenogram is necessary in the evaluation of the pediatric cervical spine. METHODS: A retrospective, multiinstitutional review was performed of all patients 16 years of age and under with documented cervical spine injury in a large metropolitan area during the past decade. Fifty-one children with cervical spine injury were identified from the medical records at 4 hospitals. RESULTS: The 0- to 8-year-old age group had a significantly higher incidence of upper (occiput to C3) cervical injury than the 9- to 16-year-old age group (67% v 39%; P < .05). In the 0- to 8-year-old group the initial lateral/anteroposterior radiograph made the diagnosis of cervical spine injury in 13 of 15 patients (87%), and in no patients was the transoral odontoid view used to make the diagnosis of cervical spine injury. In only 1 patient in the 9- to 16-year-old age group with a type III odontoid fracture was this view deemed useful. The overall mortality rate in this series was 7.8% with all deaths secondary to associated head injury. CONCLUSIONS: In the 0- to 8-year-old age group in whom the incidence of cervical spine injury is rare but frequently involves the upper cervical spine, the transoral odontoid roentgenogram may be of little value in the evaluation of the spine and should not be considered necessary in "clearing" the pediatric cervical spine. An alternative evaluation of these patients would include an initial lateral and AP radiograph, followed by computed tomography scan. PMID- 10873053 TI - Supplemental oxygen improves resolution of injury-induced pneumothorax. AB - BACKGROUND/PURPOSE: Traditionally, supplemental oxygen is administered to patients with asymptomatic pneumothorax to accelerate spontaneous resolution. However, this practice is based on models that did not include injury to the visceral pleura and ongoing pleural air leak. This study evaluated the effects of increased inspired oxygen concentration on pneumothorax resolution in a visceral pleural injury model. METHODS: A total of 27 New Zealand white rabbits were divided randomly into 3 groups: room air (RA), 40% FIO2 (40%), and 60% FIO2 (60%). A complete unilateral pneumothorax was created in each animal by a thoracoscopically guided visceral pleural puncture. The animals were then housed in designated oxygen chambers, and observers were blinded to the inspired oxygen concentration. Cross table anteroposterior chest x-rays were obtained preoperatively, postoperatively, and twice a day until the pneumothorax resolved. Time to resolution between the 3 groups was analyzed with 1-way analysis of variance (ANOVA). RESULTS: Twenty of 27 (74%) of the animals completed the study successfully. Mean time to resolution in the RA group (n = 7; 111.2 +/- 30.8 hours) was longer than in the 40% group (n = 6; 71.8 +/- 22.3 hours) and the 60% group (n = 7; 39.4 +/- 14.2 hours). The time to resolution also was longer in the 40% group than in the 60% group. Seven rabbits died before completion of the study of tension pneumothorax (3 of 7) or anesthetic complications (4 of 7). CONCLUSIONS: Supplemental oxygen improves resolution of injury-induced pneumothorax. The tradition of administering supplemental oxygen to patients with asymptomatic pneumothorax should be continued even if there is ongoing pleural air leak. PMID- 10873054 TI - Early functional outcome in children with pelvic fractures. AB - BACKGROUND/PURPOSE: Although the mortality, morbidity, and spectrum of associated injuries in children with pelvic fractures have been extensively studied, little is known about the functional outcomes in these patients. The authors examined retrospectively functional independence measurement (FIM) at discharge in children with pelvic fractures to determine how it should influence their management protocol. METHODS: The authors reviewed the records of all patients who sustained pelvic fractures between 1993 and 1998 in the trauma registry of a level I pediatric trauma center. Patients were stratified according to demographics, type of pelvic fracture, functional independence measurement, and discharge disposition. Fractures graded 1, 2, or 3 were defined as stable, whereas grade 4 fractures were deemed unstable. RESULTS: A total of 88 children sustained pelvic fractures. Seventy-four percent had stable fractures, whereas 26% sustained unstable fractures. There was no difference in age or sex between the groups; boys were more commonly injured than girls. Motor vehicle-related crashes accounted for most injuries. The mean injury severity score (ISS) for patients with a stable fracture was 17 +/- 14 and 20 +/- 13 for unstable fractures. There was no difference in overall hospitalization nor intensive care unit stay between the unstable and stable fracture patients. Eighty percent of the patients with unstable and 52% of the patients with stable pelvic fractures were dependent based on locomotion, and similar proportions were seen for the transfer category. CONCLUSIONS: Short-term function appears to be significantly impaired in a high percentage of children with stable and unstable pelvic fractures. Therefore, aggressive rehabilitation should be instituted early in all children with pelvic fractures to achieve optimal functional outcome. PMID- 10873055 TI - Bile duct size does not predict success of portoenterostomy for biliary atresia. AB - BACKGROUND/PURPOSE: Presence of large bile ducts (>200 microm) at the portal end plate has been suggested to predict success after portoenterostomy. The authors reviewed their patients with biliary atresia to test the hypothesis that bile duct size in patients with successful portoenterostomy was no different than in the patients with unsuccessful portoenterostomy. METHODS: The authors reviewed the patients at their institution from 1989 to 1998 who had the diagnosis of biliary atresia (n = 38). A pathologist blinded to the results of the operation confirmed the measurements of the bile duct remnants. RESULTS: Five of the 38 patients did not have a portoenterostomy. They underwent cholangiogram and liver biopsy and were evaluated for liver transplantation. All patients who underwent surgery (n = 33) had a Roux-en-y hepaticojejunostomy. Twenty-one patients had successful surgery (64%) and 12 patients (36%) had unsuccessful surgery. The average age at operation in the successful group was 50.9 +/- 3 days and in failures, 57.9 +/- 4 days (P = .16). Duct size at the portal end-plate was not different between the successes and failures. Two of the patients in the success group had no evidence of bile ducts grossly or histologically. CONCLUSION: Children presenting early in infancy (<3 months) with biliary atresia should undergo a portoenterostomy regardless of the size of the bile ducts at the time of exploration. PMID- 10873056 TI - Routine insertion of a silastic springloaded silo for infants with gastroschisis. PMID- 10873057 TI - The use of a spur valve in a Roux loop to prevent reflux into the biliary tract. PMID- 10873058 TI - Case report of dysphagia and esophageal web in an anemic child. PMID- 10873059 TI - Molecular imaging in oncology: the diagnostic imaging "revolution". PMID- 10873060 TI - Are we ready to use surrogate end points and surrogate tissues to evaluate response to chemopreventive and therapeutic intervention? PMID- 10873061 TI - Detection of mutated KRAS2 sequences as tumor markers in plasma/serum of patients with gastrointestinal cancer. AB - Mutated KRAS2 commonly can be detected in the plasma/serum of patients with pancreatic or colorectal cancers possessing this mutated gene. Positive assays are more common in patients with higher stage tumors but some smaller cancers can also be detected; occasionally, patients with large tumors have negative assays. Because relatively few patients with low-stage tumors have been evaluated, more studies in patients with smaller tumors are needed to further define the clinical usefulness of these assays. The reasons for variable results, particularly in patients with larger tumors, is unclear, although a variety of factors may be involved. More sensitive assays need to be developed that will increase the detection rates, although the problem of producing false positives must be minimized. The presence of mutated KRAS2 sequences in the plasma/serum seems to be quite specifically associated with the presence of cancer containing this mutated gene. This is an important feature of KRAS2 as a tumor marker. Preliminary studies in patients with pancreatic cancer suggest that assays for mutated KRAS2 can complement the commonly used CA19-9 assay and provide additional clinically useful information. The results from currently completed studies on the detection of mutated KRAS2 in patients with colorectal and pancreatic cancer are promising, and the potential usefulness of KRAS2 as a clinically important tumor marker should encourage future research. PMID- 10873062 TI - Gain-of-function mutations in the tumor suppressor gene p53. AB - The tumor suppressor protein p53 is a multifunctional transcription factor involved in the control of cell cycle progression, DNA integrity, and cell survival. p53 is mutated in half of all tumors and has a wide spectrum of mutation types. p53 mutants show different degrees of dominance over coexpressed wild-type p53, and loss of the wild-type p53 allele has been observed frequently. Several p53 mutants can exert oncogenic functions beyond their negative domination over the wild-type p53 tumor suppressor functions. These so-called gain-of-function effects, such as enhancement of tumorigenicity and therapy resistance, were investigated in p53-null cells. The possible mechanisms by which p53 mutants exert their gain-of-function effects are reviewed. The existence of functional gains of certain p53 mutants has important ramifications for tumor prognosis and cancer therapies. PMID- 10873063 TI - Rapid in vivo monitoring of chemotherapeutic response using weighted sodium magnetic resonance imaging. AB - A novel pulse sequence strategy uses sodium magnetic resonance imaging to monitor the response to chemotherapy of mouse xenograft tumors propagated from human prostate cancer cell lines. An inversion pulse suppresses sodium with long longitudinal relaxation times, weighting the image toward intracellular sodium nuclei. Comparing these weighted sodium images before and 24 h after administration of antineoplastics, we measured a 36 +/- 4% (P < 0.001; n = 16) increase in signal intensity. Experiments with these same drugs and cells, treated in culture, detected a significant intracellular sodium elevation (10-20 mM) using a ratiometric fluorescent dye. Flow cytometry studies showed that this elevation preceded cell death by apoptosis, as determined by fluorescent end labeling of apoptotic nuclei or Annexin V binding. Histopathology on formalin fixed sections of explanted tumors confirmed that drug administration reduces proliferation (2.2 versus 8.6 mitotic figures per high power field; P < 0.0001), an effect that inversely correlates with the sodium magnetic resonance image response on a tumor-to-tumor basis (P < 0.02; n = 10). Morphological features, such as central zones of nonviable cells, rims of active apoptosis, and areas of viable tumor, could be distinguished by comparing weighted and unweighted images. Advantages of this sodium imaging technique include rapid determination of drug efficacy, improved diagnosis of lesions, ease of coregistration with high resolution proton magnetic resonance imaging, and absence of costly or toxic reagents. PMID- 10873064 TI - Intratumoral administration of recombinant circularly permuted interleukin-4 Pseudomonas exotoxin in patients with high-grade glioma. AB - Human glioblastoma but not normal brain cells express numerous receptors for the cytokine interleukin (IL)-4. To target these receptors, we have investigated the safety and activity of directly infusing IL-4(38-37)-PE38KDEL, a chimeric protein composed of circularly permuted IL-4 and a truncated form of Pseudomonas exotoxin (PE), into recurrent malignant high-grade gliomas. IL-4(38-37)-PE38KDEL (IL-4 toxin) was infused over a 4-8-day period into gliomas of nine patients by one to three stereotactically placed catheters. No apparent systemic toxicity occurred in any patient. The infusion of IL-4-toxin in six of nine patients showed glioma necrosis as evidenced by diminished gadolinium enhancement on magnetic resonance imaging. Seven of nine patients underwent craniotomy because of increased intracranial pressure at 16-101 days after the beginning of infusion. In six of these seven patients, partial-to-extensive tumor necrosis with edema was confirmed pathologically. No histological evidence of neurotoxicity to normal brain was identified in any patient. Two patients were not operated on; by magnetic resonance imaging, one showed mottled gadolinium enhancement, and the other showed extensive necrosis of tumor leading to complete remission; this patient remains disease-free > 18 months after the procedure. We conclude that direct glioma injection of IL-4(38-37)-PE38KDEL is safe without systemic toxicity. Local toxicity seemed attributable mainly to tumor necrosis or occasionally to the volume of infusion. Histological evidence of toxicity to normal brain was not observed and in many patients, could be pathologically excluded. Additional patients are being treated to determine the maximal tolerated concentration and volume of IL-4(38-37)-PE38KDEL. PMID- 10873065 TI - Modulation of radiation response after epidermal growth factor receptor blockade in squamous cell carcinomas: inhibition of damage repair, cell cycle kinetics, and tumor angiogenesis. AB - We have recently demonstrated that molecular blockade of the epidermal growth factor receptor with the anti-epidermal growth factor receptor (EGFR) monoclonal antibody C225 enhances the in vitro radiosensitivity of human squamous cell carcinomas (SCCs) derived from the head and neck. In the present study, we further investigated the capacity of C225 to modulate the in vitro and in vivo radiation response of human SCC tumor cells and xenografts, and we examined several potential mechanisms that may contribute to the enhanced radiation response induced by C225. Tumor xenograft studies demonstrated complete regression of both newly established (20 mm3) and well-established (100 mm3) SCC tumors over a 55-100 day follow-up period in athymic mice treated with the combination of C225 (i.p. injection) and radiation. Cell cycle analysis via flow cytometry confirmed that combined treatment with C225 and radiation induced an accumulation of cells in the more radiosensitive cell cycle phases (G1, G2-M) with concurrent reduction in the proportion of cells in the more radioresistant S phase. Results from sublethal damage repair and potentially lethal damage repair analyses in cultured SCC cells demonstrated a strong inhibitory effect of C225 on postradiation damage repair. Further, exposure of SCC cells to C225 induced a redistribution of DNA-dependent protein kinase from the nucleus to the cytosol, suggesting one potential mechanism whereby C225 may influence the cellular response to radiation. Immunohistochemical analysis of SCC tumor xenografts after systemic administration of C225 demonstrated inhibition of the in vivo expression of tumor angiogenesis markers, including vascular endothelial growth factor and Factor VIII. Taken together, the collective data suggest that the profound in vivo antitumor activity identified in the xenograft setting when C225 is combined with radiation derives from more than simply the antiproliferative and cell cycle effects of EGFR system inhibition. In addition to antiproliferative growth inhibition, EGFR blockade with C225 appears to influence the capacity of human SCCs to effect DNA repair after exposure to radiation, and to express classic markers of tumor angiogenesis. PMID- 10873066 TI - Priming tissue-specific cellular immunity in a phase I trial of autologous dendritic cells for prostate cancer. AB - We attempted to induce therapeutic immunity against prostate-derived tissues in patients suffering from progressive hormone-refractory metastatic prostate carcinoma. Thirteen patients were treated with two infusions, 1 month apart, of autologous dendritic cells (APC8015) preexposed ex vivo to PA2024, a fusion protein consisting of human granulocyte/macrophage-colony stimulating factor (GM CSF) and human prostatic acid phosphatase (PAP). The infusions were followed by three s.c. monthly doses of PA2024 without cells. Three groups of patients each received PA2024 at 0.3, 0.6, or 1.0 mg/injection. All Ps were two-sided. Treatment was well tolerated. After infusions of APC8015, patients experienced only mild (grade 1-2) short-lived fever and/or chills, myalgia, pain, and fatigue. One patient developed grade 3 fatigue. Four patients developed mild local reactions to s.c. PA2024. Twelve patients were evaluable for response to treatment. Circulating prostate-specific antigen levels dropped in three patients. T cells, drawn from patients after infusions of APC8015, but not before, could be stimulated in vitro by GM-CSF (P = 0.0004) and PAP (P = 0.0001), demonstrating broken immune tolerance against these two normal proteins. Injections of PA2024 did not influence the reactivity of T cells against PAP and GM-CSF. However, antibodies to GM-CSF and, to a much lesser extent, to PAP reached maximum titers only after two or even three injections of PA2024, showing that directly injected PA2024 was involved in stimulation of humoral immunity. Dendritic cells exposed to antigen ex vivo can induce antigen-specific cellular immunity in prostate cancer patients, warranting further studies of this mode of immunotherapy. PMID- 10873067 TI - Quantitative analysis of circulating tumor cells in peripheral blood of osteosarcoma patients using osteoblast-specific messenger RNA markers: a pilot study. AB - Metastasis is a major cause of mortality and morbidity in osteosarcoma (OS) patients. To monitor tumor dissemination, we assessed the circulating tumor burden in OS patients by semiquantitative reverse transcription-PCR using osteocalcin, osteonectin, osteopontin, and type I collagen (COLL) mRNAs as molecular markers. We distinguished levels of the mRNAs in peripheral blood between OS patients and healthy subjects using an OS-derived cell line (Saos-2) as a reference standard. We prospectively analyzed 40 peripheral blood samples from 11 OS patients at diagnosis and 29 healthy subjects. In all 29 (100%) healthy subjects, we detected osteocalcin, osteonectin, and osteopontin mRNAs that were most likely attributed to illegitimate transcription in normal hematopoietic cells. In contrast, we found low COLL mRNA levels in only 35% (10 of 29) of healthy subjects, but significantly higher COLL mRNA levels in 91% (10 of 11) of OS patients (P < 0.0001). The reverse transcription-PCR assay for COLL mRNA was sensitive down to the detection of 10 Saos-2 cells among 10(6) normal peripheral blood nucleated cells. The upper limit of COLL mRNA determined among the healthy subjects was found exceeded by six OS patients. The substantially elevated COLL mRNA levels in peripheral blood seemed to originate from circulating malignant cells in these six OS patients, all of whom subsequently developed clinical metastases within 12 months of diagnosis (P = 0.002). Conversely, no metastases were detected in the remaining OS patients with normal COLL mRNA levels. Quantification of COLL mRNA may prove valuable for diagnosing OS micrometastasis and assessing prognosis. PMID- 10873068 TI - High microvascular blood volume is associated with high glucose uptake and tumor angiogenesis in human gliomas. AB - The purpose of this investigation was to elucidate the association between microvascular blood volume and glucose uptake and to link these measures with tumor angiogenesis. We demonstrate a regionally specific correlation between tumor relative microvascular blood volume (CBV), determined in vivo with functional magnetic resonance imaging techniques, and tumor glucose uptake determined with fluorodeoxyglucose positron emission tomography. Regions of maximum glucose uptake were well matched with maximum CBV across all patients (n = 21; r = 0.572; P = 0.023). High-grade gliomas showed significantly elevated CBV and glucose uptake compared with low-grade gliomas, (P = 0.009 and 0.008, respectively). Correlations between CBV and glucose uptake were then determined on a voxel-by-voxel basis within each patient's glioma. Correlation indices varied widely, but in 16 of 21 cases of human glioma, CBV and glucose uptake were correlated (r > 0.150). These measures were well correlated in all cases when comparing healthy brain tissue in these same patients. Tumor vascularity, as determined immunohistochemically and morphometrically on clinical samples, revealed statistically significant relationships with functional imaging characteristics in vivo. Regional heterogeneities in glucose uptake were well matched with functional magnetic resonance imaging CBV maps. Our findings support the concept that there is an association of microvascular density and tumor energy metabolism in most human gliomas. In addition, the findings are likely to have important clinical applications in the initial evaluation, treatment, and longitudinal monitoring of patients with malignant gliomas. PMID- 10873069 TI - A phase II pilot trial of concurrent biochemotherapy with cisplatin, vinblastine, dacarbazine, interleukin 2, and interferon alpha-2B in patients with metastatic melanoma. AB - In an effort to develop a biochemotherapy regimen for metastatic melanoma suitable for testing in a cooperative group setting, we modified the concurrent biochemotherapy regimen of S. S. Legha et al. (J. Clin. Oncol., 16: 1752-1759, 1998) by providing enhanced supportive care and developing a strict, conservative approach to the management of treatment-related toxicities. Patients received cisplatin, vinblastine, and dacarbazine (CVD: cisplatin (20 mg/m2) and vinblastine (1.2 mg/m2) on days 1-4, dacarbazine (800 mg/m2) on day 1 only) concurrently with interleukin 2 (9 MIU/m2/day) by continuous i.v. infusion on days 1-4 and IFN-alpha (5 MU/m2/day) on days 1-5, 8, 10, and 12. Prophylactic antibiotics and a maximum of four cycles were administered. Routine granulocyte colony-stimulating factor and aggressive antiemetics were initiated after patients 7 and 14, respectively. Forty-four patients were enrolled in this study. No patients had received prior chemotherapy or interleukin 2; however, 23 (53%) had received prior IFN-alpha, mostly in the adjuvant setting. A total of 131 treatment cycles was administered. Significant toxicities requiring dose modification included: hypotension requiring pressors (15 episodes in 11 patients), grades 3/4 vomiting (12 episodes in 15 cycles; 5 episodes in 12 patients (6 episodes in 9 cycles after initiation of the modified antiemetic regimen), transient renal insufficiency (5 episodes in 5 patients), grade 4 thrombocytopenia (24 episodes, 1 associated with bleeding), neutropenia with or without fever (15 instances, only 11 in 112 cycles after routine use of granulocyte colony-stimulating factor), and catheter-related bacteremia (2 patients). Five (16%) of 30 patients who were treated after the last protocol modification experienced what we defined as unacceptable toxicity for a cooperative group setting. Responses were seen in 19 of 40 evaluable patients (relative risk, 48%) with 8 complete responses (20%). The median response duration was 7 months (range, 1-17+ months) with one currently ongoing. The central nervous system was the initial site of relapse in 11 responding patients. The median survival duration was 11 months (range, 2-31 months). This modified, concurrent biochemotherapy regimen is active and tolerable for use in a cooperative group setting. Central nervous system relapse, however, remains a concern for responders. This regimen is being compared with CVD in a Phase III Intergroup Trial (Eastern Cooperative Oncology Group/Southwest Oncology Group 3695). PMID- 10873070 TI - T cell adoptive immunotherapy of newly diagnosed gliomas. AB - Patients with newly diagnosed gliomas were treated with adoptive transfer of ex vivo activated T lymphocytes, derived from lymph nodes (LNs) draining autologous tumor vaccines, to determine the long-term toxicity of this treatment. Twelve consecutive patients were enrolled: 2 with grade II astrocytoma, 4 with anaplastic gliomas, and 6 with glioblastoma multiforme. Patients were injected intradermally with short-term cultured autologous irradiated tumor cells, admixed with granulocyte macrophage colony-stimulating factor, to stimulate draining LNs. The LN cells were activated with staphylococcal enterotoxin A for 48 h and then cultured in medium containing interleukin 2 for an additional 6-8 days and subsequently transferred i.v. to the patients. The number of cells obtained from the LNs ranged from 9 x 10(7) to 1.1 x 10(9), and the median cell proliferation was 41-fold. The dose of T cells infused ranged from 0.6 to 5.5 x 10(10) with a median of 1.1 x 10(10), the majority of which were CD 4+ (mean, 71%). The entire treatment was performed as outpatient therapy and was associated with a toxicity of grade 2 or less, consisting mainly of fever, nausea, and myalgias during the first 24 h. There were no indications of late adverse events from this treatment even among three patients with follow-up greater than 2 years post T cell transfer. Moreover, four patients demonstrated partial regression of residual tumor. This Phase I clinical trial of adoptive immunotherapy for patients with newly diagnosed malignant gliomas demonstrates feasibility, lack of long-term toxicity, and several objective clinical responses. PMID- 10873071 TI - Pilot study of a dual gene recombinant avipox vaccine containing both carcinoembryonic antigen (CEA) and B7.1 transgenes in patients with recurrent CEA expressing adenocarcinomas. AB - Coordinated presentation of antigen and costimulatory molecules has been shown to result in the induction of an antigen-specific T-cell response rather than the development of anergy. This study evaluated the vaccine ALVAC-CEA B7.1, a canary pox virus that has been engineered to encode the gene for the tumor-associated antigen carcinoembryonic antigen (CEA) and B7.1, a T-cell costimulatory molecule. Patients with CEA-expressing tumors were immunized with 2.5 x 10(7) (n = 3), 1.0 x 10(8) (n = 6), and 4.5 x 10(8) (n = 30) plaque-forming units intradermally every other week for 8 weeks. Patients with stable or responding disease received monthly boost injections. Biopsies of vaccine sites were obtained 48 h after vaccination to evaluate leukocytic infiltration and CEA expression. Induction of CEA-specific T-cell precursors was assessed by an ELISPOT assay looking for the production of IFN-gamma. Therapy was well tolerated, without significant toxicity attributable to vaccine. All patients had evidence of leukocytic infiltration and CEA expression in vaccine biopsy sites. Six patients with elevated serum CEA values at baseline had declines in their levels lasting 4-12 weeks. These patients all had stable disease after four vaccinations. After four vaccinations, patients who were HLA-A-2-positive demonstrated increases in their CEA-specific T cell precursor frequencies to a CEA-A2-binding peptide from baseline. The number of prior chemotherapy regimens was inversely correlated with the ability to generate a T-cell response. ALVAC-CEA B7.1 is safe in patients with advanced, recurrent adenocarcinomas that express CEA, and it is associated with the induction of a CEA-specific T-cell response. PMID- 10873072 TI - The effects of neoadjuvant anastrozole (Arimidex) on tumor volume in postmenopausal women with breast cancer: a randomized, double-blind, single center study. AB - Anastrozole, an orally active, nonsteroidal aromatase inhibitor, was evaluated in a randomized, double-blind, single-center study to determine its efficacy as neoadjuvant therapy in postmenopausal women with newly diagnosed, estrogen receptor-rich, locally advanced or large (>3 cm), operable breast cancers. Twenty four eligible patients were recruited into the study and received either 1 mg (n = 12) or 10 mg (n = 12) of anastrozole daily over a 3-month period. Tumor volumes were estimated clinically, by using caliper measurements and ultrasound (at baseline and after 1, 2, and 3 months' treatment) and by mammography (at baseline and after 3 months). Tumor volume was also measured in surgical specimens. Twenty one patients were classified as T2, two patients as T3, and one patient as T4B at baseline. Three patients had clinical evidence of lymph node involvement. When considering the difference between the volume as measured by each assessment and the actual pathological volume, the interquartile range and the difference between the maximum and minimum values were smaller for ultrasound when compared with those measured with calipers and mammography. Therefore, of the three clinical assessments of tumor volume used in this study, the data suggest that ultrasound may be the most accurate. The median reductions in tumor volumes as measured by ultrasound for those patients with a measurable 12-week assessment were 80.5 and 69.6% for anastrozole (1 and 10 mg, respectively) after 12 weeks of treatment and 75.5% when both doses were grouped together. Moreover, of these patients, 11 of 12 given 1 mg and 7 of 11 given 10 mg of anastrozole were found on ultrasound to have a >50% reduction in tumor volume after 12 weeks of treatment. Of the 17 patients who would have required a mastectomy at initiation of treatment, 15 were suitable for breast conservation after anastrozole treatment. These results suggest that anastrozole is highly effective as neoadjuvant therapy in postmenopausal women with estrogen receptor-rich, large, operable breast cancer. Future studies comparing anastrozole with tamoxifen as a neoadjuvant treatment should be considered. PMID- 10873073 TI - Phase I dose-finding and pharmacokinetic trial of irinotecan hydrochloride (CPT 11) using a once-every-three-week dosing schedule for patients with advanced solid tumor malignancy. AB - A Phase I study was performed to determine the maximum tolerated dose (MTD), toxicities, and pharmacokinetic profile of irinotecan (CPT-11) and its active metabolites when given on a once-every-3-week schedule. Thirty-four patients with advanced refractory solid malignancies were treated with CPT-11 (240-340 mg/m2) administered as a 90-min i.v. infusion every 3 weeks. Patients were divided into two groups: those with and those without prior abdominal/pelvic (AP) radiotherapy. Gastrointestinal toxicity (nausea, vomiting, and diarrhea) and hematological toxicity (leukopenia and neutropenia) were dose-limiting side effects. Other common toxicities included anorexia, asthenia, and acute cholinergic symptoms (abdominal cramps, diaphoresis, and lacrimation). For patients with no prior AP radiation therapy, the MTD was determined to be 320 mg/m2, whereas those with prior AP radiation therapy had a MTD of 290 mg/m2. Dose proportional increases in the mean area under the concentration-time curves for CPT-11, SN-38, and SN-38G were not observed over the narrow dose range studied. Mean values of terminal phase half-life, clearance, terminal phase volume of distribution, and steady-state volume of distribution for CPT-11 were 12.4 +/- 1.8 h, 13.0 +/- 3.8 liters/h/m2, 234 +/- 83 liters/m2, and 123 +/- 38 liters/m2, respectively. The pharmacodynamic analyses indicated the strongest correlation to be between SN-38 area under the concentration-time curves and neutropenia (p = 0.60; P = 0.001). A total of five responses (one complete response and four partial responses) were observed in the cohort of 32 patients with previously treated metastatic colorectal carcinoma. In conclusion, gastrointestinal toxicity and hematological toxicity were the dose-limiting toxicities of CPT-11 when administered as a 90-min infusion every 3 weeks. In this trial, the recommended Phase II starting dose for patients with no prior AP radiation therapy was found to be 320 mg/m2; for patients with prior AP radiation, the recommended Phase II starting dose was 290 mg/m2. This once-every-3-week schedule has been incorporated into a Phase I trial of CPT-11 combined with 5-fluorouracil and leucovorin. PMID- 10873074 TI - Determination of intermediate biomarker expression levels by quantitative reverse transcription-polymerase chain reaction in oral mucosa of cancer patients treated with liarozole. AB - Liarozole is a 1-substituted imidazole derivative that inhibits cytochrome P450 activity and increases endogenous plasma concentrations of retinoid acid (RA). We have previously demonstrated that RA down-modulates transforming growth factor (TGF)-alpha and epidermal growth factor receptor (EGFR) levels in head and neck squamous cell carcinoma by decreasing the transcription rate of these two genes. Previous reports suggest that RA receptor (RAR)-beta levels are down-modulated in head and neck cancer and are restored by RA therapy. Cellular RA-binding protein (CRABP)-II is up-regulated by RA and appears to modulate intracellular RA metabolism. In conjunction with a Phase I clinical trial, total intact RNA was extracted from oral cavity mucosa biopsied from 17 patients with advanced malignancies, before and after treatment with a 4-week course of liarozole. To analyze these limited quantities of total RNA (as little as 0.6 microg/sample), a quantitative reverse transcription-PCR assay was developed using delayed dropping of the 5' beta-actin primer to amplify the highly abundant beta-actin gene as an internal control. We used this method to determine the expression levels of TGF alpha, EGFR, RAR-beta, and CRABP-II before and after treatment. There was a trend toward elevation of RAR-beta levels in oral mucosa after liarozole therapy (P = 0.107), whereas TGF-alpha, EGFR, and CRABP-II were not modulated by systemic liarozole treatment. These results suggest that liarozole may up-regulate RAR beta in tissues from cancer patients and that expression levels of potential intermediate biomarkers may be determined in small tissue biopsies using a quantitative reverse transcription-PCR assay. PMID- 10873075 TI - Imaging of soft-tissue tumors using L-3-[iodine-123]iodo-alpha-methyl-tyrosine single photon emission computed tomography: comparison with proliferative and mitotic activity, cellularity, and vascularity. AB - The radiolabeled amino acid L-3-[123I]-iodo-alpha-methyltyrosine (IMT) is a new tumor tracer that accumulates in many tumors and is suitable for single photon emission computed tomography (SPECT) imaging. Using IMT SPECT, we studied 32 patients with a soft-tissue tumor suspected to be a soft-tissue sarcoma to determine whether: (a) tumors can be visualized; (b) benign and malignant lesions can be distinguished; and (c) IMT uptake is related to tumor grade and proliferation. Whole-body imaging was performed 15 min after administration of 300 MBq IMT, biopsy, or resection 1-2 weeks later. IMT uptake was quantified using a region-of-interest method resulting in tumor:background (T:B) ratios. These were compared with tumor grade, mitotic index, tumor cellularity, vascularity, and the Ki-67 proliferation index. Eleven patients had a benign tumor, and 21 patients had a soft-tissue sarcoma. Six benign tumors demonstrated minor IMT uptake, and five lipomas had no uptake. All malignant tumors had high uptake and were clearly visualized. T:B ratios in malignant tumors (3.83 +/- 1.16) were higher (P < 0.001) than in benign tumors (1.52 +/- 0.60). Small (<5 mm) metastases in two patients were not detected. Taking the T:B ratio 2.0 as the cutoff level, the sensitivity for detection of malignancy was 100%, and specificity was 88%. IMT uptake correlated with histological grade (r = 0.82; P < 0.001), mitotic index (r = 0.75; P < 0.001), tumor cellularity (r = 0.73; P < 0.01), and with the Ki-67 proliferation index (r = 0.63; P < 0.01). In conclusion, IMT SPECT visualized all soft-tissue sarcomas. Uptake in sarcomas was clearly higher than in benign lesions, yielding 100% sensitivity for detection of malignancy at 88% specificity. Uptake increased with higher tumor grade and higher proliferation rate. PMID- 10873076 TI - Antiproliferative effects of idoxifene in a placebo-controlled trial in primary human breast cancer. AB - Idoxifene is a novel selective estrogen receptor modulator. It has reduced agonist activity on breast and uterine cells compared with tamoxifen and antiproliferative effects in tamoxifen-resistant breast cancer cells. Previous studies have shown that a short course of treatment with other antiestrogens prior to surgery caused a significant reduction of the growth fraction when measured by immunohistological staining using the mouse monoclonal antibody Ki67. In this study, we assessed the effect of idoxifene on biological markers of cell proliferation (Ki67) and apoptosis (TdT-mediated dUTP-biotin nick end labeling), and estrogen and progesterone receptor (ER/PR) expression was also evaluated. Core-cut biopsies were obtained in 77 postmenopausal patients with primary breast cancer at diagnosis. Patients were randomized to 40 mg/day idoxifene or placebo for 14-21 days prior to obtaining a second biopsy sample at surgical resection. The percentage of Ki67-positive cells fell from a mean 19.7 +/- 2.7% (SE) to 13.4 +/- 3.4% in idoxifene-treated ER-positive tumors (n = 30; P = 0.0043), but there was no significant effect in placebo-treated ER-positive tumors (n = 27). No effect was seen on ER-negative tumors in either group. Idoxifene had no significant effect on apoptotic index but produced a statistically significant fall in idoxifene-treated ER immunohistochemical score and a small increase in PR that did not reach statistical significance (0.05 < P < 0.10). Idoxifene was well tolerated in all patients. Idoxifene has an antiproliferative effect in ER positive but not ER-negative breast cancers, and no significant effect on apoptosis in the short-term. PMID- 10873077 TI - Clinical and biological effects of intraperitoneal injections of recombinant interferon-gamma and recombinant interleukin 2 with or without tumor-infiltrating lymphocytes in patients with ovarian or peritoneal carcinoma. AB - To identify strategies that enhance tumor-specific immunity in patients with ovarian carcinoma, 22 patients received four to six doses of i.p. recombinant IFN gamma (rIFN-gamma), 200 microg/m2 on days 1, 3, 5, 8, 10, and 12, and i.p. recombinant interleukin 2 (rIL-2), either 6.0 x 10(5) IU/m2 (group A) or 1.0 x 10(5) IU/m2 (group B), on days 9, 10, and 11. Two patients in group A also received T-cell lines expanded from peritoneal tumor-infiltrating lymphocytes (TILs) obtained after i.p. rIFN-gamma/rIL-2 administration. Toxicity was manageable and included five nonhematological grade 3 or 4 events in 22 patients (23%). A patient had normalization of CA-125 values and a progression-free interval of 18 months, after receiving i.p. rIFN-gamma/rIL-2 without TILs. Another patient who received i.p. rIFN-gamma/rIL-2 plus TILs had stabilization of ascites and intra-abdominal tumors and >50% reduction in serum CA-125 values over 6 months. A third patient who received i.p. rIFN-gamma/rIL-2 had stabilization of intra-abdominal tumors and ascites accompanied by CA-125 values of 50 to 100 units over 6 months. T-cell lines for adoptive immunotherapy were developed for only 3 of 20 patients who were treated with rIFN-gamma/rIL-2. Large numbers of CD3- CD56+ adherent cells were expanded in rIL-2 in the remaining patients, precluding the development of T-cell lines. i.p. rIFN-gamma, either alone or followed by rIL-2, increased proportions of human leukocyte antigen (HLA) class I+ and class II+ tumor cells and increased HLA class I staining intensity on peritoneal carcinoma cells. i.p. rIFN-gamma plus rIL-2 also enhanced cytotoxic activity against Daudi and K562 cells and against allogeneic ovarian tumor cells. Increased cytotoxic activity was associated with an increase in the proportion of CD56+ cells. IFN-gamma and IL-2 transcripts were expressed more frequently after rIFN-gamma and rIL-2 treatment. In addition, the proportions of CD45RA+ (naive lymphocytes) were increased, and CD8+ DR+ lymphocytes were increased relative to CD8+ CD69+ cells, which were decreased. IL-10 concentrations in peritoneal fluids were increased after treatment with rIFN-gamma and the higher rIL-2 dosing (group A) but not in those treated with rIFN-gamma and the lower rIL-2 dosing (group B). These results demonstrated that patients with ovarian carcinoma can tolerate treatment with rIFN-gamma and rIL-2 and that rIFN-gamma alone or rIFN-gamma combined with rIL-2 enhances the expression of HLA class I and class II antigens on ovarian tumor cells, although immunosuppressive cytokines, such as transforming growth factor-beta and IL-10, may persist. Treatment with rIFN gamma/rIL-2 i.p. did not facilitate the production of TIL-derived T-cell lines ex vivo. PMID- 10873078 TI - Dose-finding and pharmacologic study of chronic oral idarubicin therapy in metastatic breast cancer patients. AB - Oral idarubicin (IDA) is an active drug in metastatic breast cancer, but its role in the management of this tumor is yet not established completely. To investigate a new modality of IDA administration, a dose-finding study was designed with hyperfractionated doses. The purpose was to determine the maximum tolerated dose (MTD), the dose-limiting toxicity (DLT), and the pharmacokinetics of this schedule. IDA was administered twice daily as outpatient therapy in cycles of 3 weeks followed by a 1-week rest. Thirty-one patients with progressive metastatic breast cancer and pretreated with chemotherapy (including epirubicin and doxorubicin) were enrolled. DLT was defined as G4 hematological toxicity or any other toxicity G3 or higher (Bloom and Richardson grading). Inter- and intrapatient dose increases were studied. Pharmacokinetics of IDA and its metabolite idarubicinol (IDOL) were evaluated. IDA dose was increased from 2 mg/day to 10 mg/day, by steps of 1 mg/day, with the larger dose given in the evening. MTD was reached at 10 mg/day. Overall, the therapy cycles were 69 (median/patient, 2; range, 1-6). DLTs were G4 neutropenia associated with leukopenia and thrombocytopenia in one patient and G3 diarrhea in another of the 5 patients in the 10 mg/day cohort. The two patients developing DLT at the daily dose of 10 mg received a dose normalized for body surface of 6.85 and 5.65 mg/m2/day, respectively. We considered 5.5 mg/m2/day to be the MTD. Other toxicities were nausea, vomiting, neutropenia, and diarrhea, grades G1 to G2. By univariate analysis, significant correlations were observed between absolute neutrophil count at nadir and IDA area under the curve (P = 0.022; r = -0.33), IDA Cmax (P = 0.0067; r = -0.38), IDOL area under the curve (P = 0.0009; r = 0.43), and IDOL Cmax (P = 0.0016; r = -0.41), respectively. By multivariate analysis, IDA Cmax was the strongest determinant for neutropenia (R2 = 0.14; P = 0.01). Among the 21 patients evaluable for response, 3 (14.3%) had partial response (lasting 3, 6, and 8 months, respectively), and 6 (28.6%) had a complete arrest of disease progression (lasting 2-6 months). In conclusion, the MTD of this schedule is 10 mg/day and the DLTs are neutropenia and diarrhea. Tolerance was good, and the treatment is feasible as home therapy. Some objective measurable responses were documented in this group of anthracycline-pretreated patients. IDOL could have a role for the pharmacological effect. Further evaluation of this schedule is warranted to assess the activity and toxicity of prolonged oral IDA administration. PMID- 10873079 TI - Phase I trial of oral 2'-deoxy-2'-methylidenecytidine: on a daily x 14-day schedule. AB - 2'-deoxy-2'-methylidenecytidine (DMDC) is a potent deoxycytidine analogue. Preclinical studies of DMDC demonstrated activity against a variety of murine and human tumors in cell cultures and murine models and indicate enhanced antitumor activity of DMDC when it was administered in a manner that provided prolonged systemic exposure. In view of this observation, this study was designed to determine the toxicities, maximum-tolerated dose, and pharmacokinetic profile of DMDC. DMDC was given p.o. under fasting conditions for 14 consecutive days every 4 weeks in patients with advanced solid tumors. The starting dose was 12 mg/m2/day. Pharmacokinetic studies were carried out on days 1 and 14 of the first cycle. Fourteen patients received 22 courses of DMDC. The dose-limiting toxicities were anorexia, leukopenia, thrombocytopenia, and anemia. General fatigue was the common nonhematological toxicity. The maximum-tolerated dose was 18 mg/m2/day, at which two of six patients developed grade 3 toxicities. This dose level could also be considered for Phase II testing with this schedule. At the 18-mg/m2/day dose level, the mean terminal half-life, maximum plasma concentration (Cmax), the area under the plasma drug concentration-time curve (AUC(0-infinity)) on day 1 were 1.7496 h, 112.9 ng/ml, and 399.8 ng x h/ml, respectively. Forty to 50% of the administered dose was recovered in the urine, indicating a good bioavailability and resulting significant systemic exposure to the drug, which may enable chronic oral treatment. PMID- 10873080 TI - Relative expression of type IV collagenase, E-cadherin, and vascular endothelial growth factor/vascular permeability factor in prostatectomy specimens distinguishes organ-confined from pathologically advanced prostate cancers. AB - The tumor grade (Gleason score) in the biopsy and pretherapy prostate-specific antigen level do not accurately predict disease outcome of individual patients' prostate cancer. We used a rapid colorimetric in situ hybridization technique to evaluate the expression level of E-cadherin (which affects cell cohesion); matrix metalloproteinases (MMPs) types 2 and 9 (which affect invasion); and vascular endothelial growth factor/vascular permeability factor (which affects angiogenesis) in archival prostatectomy specimens from 40 patients. Intratumoral heterogeneity for gene expression (edge versus center versus perineural area) was more pronounced in advanced cancers than in those that were organ confined. Regardless of Gleason score, the highest expression level for E-cadherin was found in the center or perineural area of the tumors, whereas the highest expression levels for MMP-2 and MMP-9 were associated with the invasive edge. The relationship between advancing pathological stage and expression of all four metastasis-related genes was highly significant. Decreased expression of E cadherin and increased expression of MMP-2, MMP-9, and vascular endothelial growth factor/vascular permeability factor were associated with the Gleason score of the tumors. Irrespective of serum prostate-specific antigen level or Gleason score, the ratio between expression of MMPs and E-cadherin at the invasive edge of tumors exhibited the strongest association with nonorgan-confined prostate cancer. These data suggest that the relative expression of metastasis-related genes in radical prostatectomy specimens can distinguish between organ-confined and advanced prostate cancers and provides the rationale for a prospective study correlating gene expression in pretherapy core biopsies with outcome. PMID- 10873081 TI - Extracellular catalytic subunit activity of the cAMP-dependent protein kinase in prostate cancer. AB - The role of cAMP in cell growth and differentiation, gene expression, and neuronal function is mediated by the cAMP-dependent protein kinase (PKA). Differential expression of type I and type II PKA has been correlated with neoplastic transformation and differentiation, respectively. PKA is primarily an intracellular enzyme. However, it has been demonstrated that PKA may be associated with the plasma membrane and is exposed to the extracellular environment. Here we report the first evidence for the presence of a free extracellular kinase activity of PKA in the growth media of cultured prostate and other cancer cells, as well as in plasma samples from prostate cancer patients. This PKA activity is specific due to its phosphorylation of the PKA-specific substrate kemptide and its inhibition by the potent and specific PKA inhibitor PKI, but not by other protein kinase-inhibitory peptides. Intriguingly, this exoprotein kinase activity is cAMP independent, suggesting that only the catalytic subunit is secreted, and therefore the kinase activity is not modulated by the regulatory subunit of PKA. Western blot analysis of the culture supernatant from prostate cancer cells indicates the presence of the catalytic subunit. This increase in extracellular PKA catalytic subunit activity in prostate cancer may have profound effects on the tumorigenesis of prostate cancer and may serve as a novel marker and therapeutic target for the disease. PMID- 10873082 TI - Comparison of potential markers of farnesyltransferase inhibition. AB - Farnesyltransferase inhibitors (FTIs) were developed to target abnormal signaling pathways that are commonly activated in neoplastic cells. Five FTIs have recently undergone Phase I testing; and two are currently in Phase II clinical trials. As part of the development of these agents, there has been interest in determining their cellular effects in the clinical setting. Several approaches have been proposed, including measurement of FT enzymatic activity, evaluation of the processing of FT polypeptide substrates, and assessment of the accumulation of p21waf1. In the present study, a number of these assays have been compared in four cultured human neoplastic cell lines of different histology (A549, HCT116, BxPC-3, and MCF-7) after treatment with the nonpeptidomimetic FTI SCH66336 and the peptidomimetic inhibitor FTI-277. Immunoblotting studies failed to demonstrate a mobility shift in ras proteins or increased accumulation of p21waf1 after treatment with these agents. In contrast, drug-induced increases in the slower migrating, unprocessed species of the chaperone protein HDJ-2 and the intranuclear intermediate filament protein lamin A were detected in all four cell lines after treatment with either agent. Unprocessed forms of both polypeptides accumulated in noncycling as well as cycling cells. The precursor peptide that is present in prelamin A but absent from mature lamin A could be readily detected by immunohistochemistry in noncycling cells with a peptide-specific antiserum. Our results indicate that unprocessed HDJ-2 and prelamin A should be suitable markers of FT inhibition in clinical samples. PMID- 10873084 TI - Enhanced urinary gelatinase activities (matrix metalloproteinases 2 and 9) are associated with early-stage bladder carcinoma: a comparison with clinically used tumor markers. AB - Matrix metalloproteinases (MMPs) are involved in tumor growth and metastasis, promoting the migration and invasion of cells. In this study, the amount of MMP-2 and MMP-9 activity was measured in urine from superficial bladder carcinoma patients (pTa, pT1) to evaluate their possible diagnostic value. The active and total amount of MMP-2 and MMP-9, respectively, in urine from tumor patients were compared with the levels in urine from age- and gender-matched healthy volunteers. Both MMP-2 and MMP-9 activity levels were significantly enhanced in urine from patients with high invasive cancers (pT2, PT3), whereas in urine from healthy controls no or very low MMP activities were found. More importantly, a substantial number of urine samples from patients with superficial tumors contained elevated MMP-2 and MMP-9 activities, suggesting that enhanced urinary MMP activity levels, indeed, might be indicative for early-stage bladder cancer. Overall, urinary MMP-2 and MMP-9 activity levels were significantly correlated to each other, with some individual exceptions. A comparison between urinary MMP-9 activity and a recently proposed urinary marker for bladder cancer, NMP-22, showed slightly lower numbers of patients with elevated levels for MMP-9. But because MMP-9 and NMP-22 levels were not correlated, enhanced urinary MMP activity might be useful as a marker for superficial bladder carcinoma like, or especially in combination with, other markers. PMID- 10873083 TI - Genetic detection for micrometastasis in lymph node of biliary tract carcinoma. AB - The presence of regional lymph node metastasis is one of the most significant poor-prognosis factors in patients with biliary tract carcinoma. To establish a sensitive reverse transcription (RT)-PCR assay to detect micrometastases in lymph nodes of biliary tract carcinoma, we first investigated the optimal markers in biliary tract carcinoma. The expressions of the six candidates for a suitable RT PCR marker [mammaglobin B, carcinoembryonic antigen (CEA), cytokeratin (CK) 20, prostate-specific antigen, and melanoma antigens (MAGE-1 and MAGE-3)] were evaluated in two bile duct cancer cell lines and human biliary tract carcinoma tissues. Of 32 carcinoma tissues, mammaglobin B, CEA, prostate-specific antigen, MAGE-1, MAGE-3, and CK 20 were expressed in 28 (88%), 26 (81%), 4 (13%), 5 (16%), 7 (22%), and 9 (28%), respectively. Mammaglobin B and CEA were considered to be good markers of the six candidates. We then examined 209 lymph nodes obtained from 15 patients with biliary tract carcinoma by RT-PCR assay using both mammaglobin B and CEA and compared the results with those of histological examination. All of 20 histologically positive lymph nodes for metastasis displayed the PCR product(s) of marker genes. Of 189 histologically negative nodes, 24 (13%) nodes expressed mammaglobin B and/or CEA mRNA, suggesting the presence of micrometastasis. Our findings suggest that mammaglobin B and CEA could be useful RT-PCR markers for the detection of lymph node micrometastases in biliary tract carcinomas. Our RT-PCR assay allows accurate clinical staging necessary for patient stratification with respect to adjuvant therapy after surgery. PMID- 10873085 TI - Clinicopathological significance of fragile histidine triad transcription protein expression in endometrial carcinomas. AB - Abnormalities in structure and expression of the fragile histidine triad transcription (FHIT) gene have been reported in a variety of cancers, including endometrial cancers. A good correlation between FHIT gene alteration and loss of Fhit expression was observed in endometrial cancers, although those are the selected cases. Therefore, we investigated the association of Fhit expression with clinicopathological features in 111 cases of endometrial cancer. Loss of Fhit expression was associated with high malignant potential, including extensive muscular invasion, advanced surgical stage, high histological grade, nonendometrioid types of adenocarcinoma, negative estrogen receptor status, and p53 overexpression. The presence of personal cancer history was also related to the loss of Fhit with a marginal significance. Survival curves determined by the Kaplan-Meier method and univariate analysis demonstrated that decreased expression of Fhit was associated with a poor outcome. However, multivariate analysis using the stepwise Cox proportional hazard model showed that whereas lymph node metastasis, advanced stage, and high tumor grade were related to poor survival rates, loss of Fhit expression was not. Consequently, loss of Fhit expression is associated with advanced surgical stage and does not appear to be an independent prognostic factor in endometrial cancers, although a still larger sample of patients will be required to asses this issue definitively. PMID- 10873086 TI - Matrix metalloproteinase 9 and the epidermal growth factor signal pathway in operable non-small cell lung cancer. AB - Matrix metalloproteinase (MMP)-9 is an endopeptidase that digests basement membrane type IV collagen. Enhanced expression has been related to tumor progression both in vitro and in vivo. The control of MMP transcription is complex, but recently, epidermal growth factor receptor (EGFR) expression has been implicated in up-regulation of MMP-9 in tumor cells in vitro. Our objective was to evaluate the relationship between MMP-9 and EGFR expression in non-small cell lung cancer (NSCLC) and to assess the impact of expression on clinicopathological parameters and survival. This is a retrospective study of 169 patients who underwent resection for stage I-IIIa NSCLC with a postoperative survival >60 days. Minimum follow-up was 2 years. Standard avidin-biotin complex immunohistochemistry was performed on 4-microm paraffin-embedded sections from the tumor periphery using monoclonal antibodies to EGFR and MMP-9. MMP-9 was expressed in the tumor cells of 88 of 169 (52%) cases. EGFR expression was found in 94 of 169 (56%) cases [membranous, 55 of 169 (33%); cytoplasmic, 39 of 169 (23%)]. MMP-9 expression was associated with poor outcome in univariate (P = 0.0023) and multivariate (P = 0.027) analysis. Membranous, cytoplasmic, and overall EGFR expression were not associated with outcome (P = 0.13, 0.99, and 0.17, respectively). MMP-9 expression showed a strong correlation with EGFR expression (P < 0.0001) and EGFR membranous expression (P = 0.002) but not with cytoplasmic EGFR expression (P = 0.18). Co-expression of MMP-9 and EGFR (37%) conferred a worse prognosis (P = 0.0001). Subset analysis revealed only MMP-9 and membranous EGFR co-expression (22%) was associated with poor outcome (P = 0.0019). Our results show that a significant proportion of NSCLC tumors co express MMP-9 and EGFR. The co-expression of these markers confers a poor prognosis. This finding suggests that EGFR signaling pathway may play an important role in the invasive behavior of NSCLC via specific up-regulation of MMP-9. PMID- 10873087 TI - Circulating HER2 extracellular domain and resistance to chemotherapy in advanced breast cancer. AB - To test the hypothesis of an association between HER2 and chemotherapy resistance, we performed a prospective assessment of the predictive value of the circulating HER2 extracellular domain (ECD) in patients with advanced breast carcinoma in the setting of a multicenter Phase II trial using paclitaxel and doxorubicin. Serum samples were collected from 58 patients with metastatic breast carcinoma before first-line chemotherapy for advanced disease, and the levels of circulating HER2 ECD were measured using an enzyme immunoassay. Immunohistochemistry with anti-HER2 monoclonal antibody CB11 was used to assess the overexpression of HER2 in the primary tumors. When 450 fmol/ml was used as a cutoff, 24 cases (41%) had elevated HER2 ECD levels. Elevated levels of circulating HER2 ECD were associated with the expression of HER2 in the primary tumor tissue and with the metastatic tumor burden (evaluated with the marker CA 15-3; P = 0.032 and P = 0.002, respectively) but not with variables such as menopausal status, stage at diagnosis, previous adjuvant therapy, or the number of metastatic sites. The levels of circulating HER2 ECD correlated inversely with the response to treatment. The probability of obtaining a complete response to chemotherapy was significantly lower (P = 0.021) in patients with elevated HER2 ECD levels (0%; 95% confidence interval, 0-13%) compared with patients with nonelevated HER2 (26%; 95% confidence interval, 12-45%). In addition, the duration of clinical response was significantly shorter in patients with elevated HER2 ECD, compared with the cases with nonelevated HER2 (7.5 versus 11 months; P = 0.035). In conclusion, elevated levels of circulating HER2 ECD in patients with metastatic breast cancer correlate with reduced efficacy of a paclitaxel doxorubicin chemotherapy combination. We suggest that the poor response rate associated with HER2 expression in advanced breast cancer may not be reversed by aggressive chemotherapy alone. PMID- 10873088 TI - The presence of soluble c-erbB-2 in saliva and serum among women with breast carcinoma: a preliminary study. AB - The protein c-erbB-2, also known as Her2/neu, is a prognostic breast cancer marker assayed in tissue biopsies from women diagnosed with malignant tumors. Present studies suggest that soluble fragments of the c-erbB-2 oncogene may be released from the cell surface and become detectable in patients with carcinoma of the breast. Consequently, the purpose of this study was to assay the c-erbB-2 protein in the saliva and serum of women with and without carcinoma of the breast and to determine whether the protein possesses any diagnostic value. To determine the diagnostic utility of this oncogene, the soluble form of the c-erbB-2 protein was assayed in the saliva and serum using ELISA in three different groups of women. The three groups consisted of 57 healthy women, 41 women with benign breast lesions, and 30 women diagnosed with breast cancer. To compare the relative diagnostic utility of the c-erbB-2 protein, CA 15-3 was also measured. The CA 15-3 measurements served as a "gold standard" by which to compare the c erbB-2 protein's diagnostic effectiveness. We found c-erbB-2 protein in the saliva and serum of all three groups of women. The salivary and serological levels of c-erbB-2 in the cancer patients, however, were significantly higher (P < 0.001) than the salivary and serum levels of healthy controls and benign tumor patients. Additionally, the c-erbB-2 protein was found to be equal to or to surpass the ability of CA 15-3 to detect patients with carcinoma. The results of the pilot study suggest that the c-erbB-2 protein may have potential use in the initial detection and/or follow-up screening for the recurrence of breast cancer in women. PMID- 10873089 TI - Expression of BAX in plasma cell dyscrasias. AB - Several studies demonstrate that the BCL-2 and BCL-XL antiapoptotic genes are variably expressed in plasma cells of patients with multiple myeloma (MM). However, the plasma cell expression of BAX protein, their major proapoptotic partner, has not been investigated. Our initial Western blot analysis of myeloma cell extracts also suggested patient variability in the expression of BAX, which was not altered by exposure to interleukin 6. To further investigate the significance of BAX expression, we performed immunohistochemistry on archival bone marrow biopsies and compared BAX staining to BCL-2 immunostaining. Expression was first evaluated in 104 patients with reactive plasmacytosis, monoclonal gammopathy of undetermined significance/smoldering MM, or active MM. An increase (P < 0.05) in expression of both BAX and BCL-2 was detected in MM patients compared with patients with reactive plasmacytosis. Patients with monoclonal gammopathy of undetermined significance/smoldering MM had intermediate values. For correlations with outcome, expression was assessed in 43 patients at diagnosis who were treated with melphalan and prednisone; 30 at diagnosis who were treated with vincristine, Adriamycin, and dexamethasone; and 29 at relapse who were treated with second-line therapy. There was no correlation between BAX or BCL-2 expression and response to chemotherapy or duration of response or between BCL-2 expression and survival. However, patients who demonstrated extremely low plasma cell BAX expression had significantly increased survival. This was true for patients initially treated with melphalan and prednisone or vincristine, Adriamycin, and dexamethasone, as well as patients studied at relapse. BAX expression did not correlate with expression of proliferating cell nuclear antigen used as a marker of proliferation. These data indicate a myeloma specific increase in BAX expression in plasma cells and suggest that low BAX expression identifies a cohort of patients with long survival, which is not specifically associated with low proliferating cell nuclear antigen expression. PMID- 10873090 TI - Noninvasive diagnosis of bladder carcinoma by enzyme-linked immunosorbent assay detection of CD44 isoforms in exfoliated urothelia. AB - The expression of variant isoforms of the adhesion molecule CD44 is correlated with the onset of neoplasia in many carcinomas. We have previously shown that noninvasive detection of bladder carcinoma is possible by analysis of anomalous CD44 expression in exfoliated urothelia. Although the sensitivity and specificity values obtained for the detection of bladder tumors using RT-PCR and Western blotting methods were superior to those obtained using urine cytology, the application of such techniques is inconvenient for routine diagnostic use. We now report the design and development of a sandwich-ELISA system for the reliable detection of CD44 protein extracted from sedimented urothelial cells in voided urine. Naturally micturated urine samples were obtained from 53 patients with newly diagnosed bladder cancer and from 65 subjects with no evidence of disease; patients with gross hematuria were excluded because of interference with the assay. To demonstrate the diagnostic potential of the system, a "gate" was imposed at N (max), i.e., the highest absorbance value obtained from a sample known to be tumor free. All values above this value were assumed to be indicative of the presence of a tumor. Using this parameter, 42 of 53 (81.1%) patients with histologically confirmed bladder tumors were correctly diagnosed. Correspondingly, under these conditions, the assay is 100% specific for tumor detection, with a sensitivity of 81.1%, which equates to a positive predictive value of 100% and a negative predictive value of 81.1%. A further 54 patients who had previously received treatment for bladder cancer but were currently clinically disease-free were also investigated. Of these, 47 of 54 (87%) were correctly diagnosed to be tumor-free, which in this group equates to a positive predictive value of 87% and a negative predictive value of 100%. The data presented demonstrate that the rapid and accurate detection of elevated levels of CD44 protein isoforms in exfoliated urothelial cells is applicable to the identification and monitoring of primary and recurrent bladder cancer. PMID- 10873091 TI - Detection by denaturant gradient gel electrophoresis of tumor-specific mutations in biopsies and relative bronchoalveolar lavage fluid from resectable non-small cell lung cancer. AB - A PCR-denaturant gradient gel electrophoresis (DGGE) method was developed for the detection of p53 and K-ras mutations in primary operable tumors and paired BAL samples of non-small cell lung cancer. Among 36 patients, 9 showed p53 exon V mutations in biopsies and in three paired bronchoalveolar lavage (BAL) specimens with a 33% concordance. Five patients presented p53 exon VI mutations in biopsies and in two paired BALs with a 40% concordance. No mutations were found in p53 exon VII either in biopsies or in paired BAL samples with 100% concordance. Exon VIII mutations were found in six primary tumors and in two BALs with a 33% concordance. Of 36 patients, we detected 7 (19.4%) with K-ras exon I mutations on tumor samples. DGGE analysis of DNA from BAL samples revealed three mutations distributed on K-ras exon I with a 42% overall concordance with respect to tumor tissue. Molecular screening by DGGE of p53-amplified DNA from BAL had cumulative 46.6% sensitivity, 100% specificity, and 77.7% accuracy. DGGE K-ras detection showed 43% sensitivity, 100% specificity, and 88.8% test accuracy. The method proposed demonstrated to be specific, accurate, and at relatively low cost but limited by low sensitivity in detecting the presence of neoplastic cells in patients with resectable non-small cell lung cancer. PMID- 10873092 TI - Increased expression of an ATP-binding cassette superfamily transporter, multidrug resistance protein 2, in human colorectal carcinomas. AB - The expression of ATP-binding cassette superfamily transporter genes, such as P glycoprotein/multidrug resistance (MDR) 1 and MDR protein (MRP) 1, is often up regulated in various tumor types and is involved in responses to some anticancer chemotherapeutic agents. Five human MRP subfamily members (MRP2-6) with structural similarities to MRP1 have been identified. The relationships between MRP2-6 mRNA levels and drug resistance are not well understood. Data on 45 patients with colorectal cancer were analyzed. Of the ATP-binding cassette superfamily genes, we asked whether mRNA levels of MDR1, MRP1, MRP2, and MRP3 correlated with drug resistance to anticancer agents. For this analysis, we used quantitative reverse transcription-PCR, and the sensitivity to anticancer agents in surgically resected colon carcinomas was determined using the in vitro succinate dehydrogenase inhibition test. MDR1, MRP1, and MRP3 were highly expressed in normal colorectal mucosa, and the relative mRNA levels of MDR1, MRP1, and MRP3 in cancerous tissues compared with noncancerous tissues were decreased or unchanged. By contrast, MRP2 mRNA expression was low in normal colorectal mucosa and specifically increased in cancer regions compared with noncancerous regions. Of the anticancer agents prescribed for patients with colorectal cancers, including doxorubicin, mitomycin C, cisplatin, 5 fluorouracil, etoposide, and a camptothecin derivative, mRNA expression of MRP2 was significantly associated with resistance to cisplatin. MRP2 may be important for resistance to cisplatin treatment in colorectal cancer. PMID- 10873093 TI - Inducible nitric oxide synthase expression, apoptosis, and angiogenesis in in situ and invasive breast carcinomas. AB - In this investigation, we studied the expression of inducible nitric oxide synthase (iNOS) and its association to apoptosis and angiogenesis in 43 in situ and 68 invasive breast carcinomas. Its expression was studied immunohistochemically using a polyclonal iNOS antibody, and the staining was evaluated both in tumor and stromal cells. Apoptosis was detected by 3' end labeling of fragmented DNA (terminal deoxynucleotidyl transferase-mediated nick end labeling method). Vascularization was detected immunohistochemically using an antibody to the FVIII-related antigen, and calculated microvessel densities were determined. In addition to strong iNOS expression in stromal cells, iNOS positivity was observed in tumor cells in 46.5% of in situ and 58.8% of invasive carcinomas. In invasive carcinomas, there were more cases with iNOS positivity both in tumor and stromal cells compared to in situ carcinomas (0.007). The proportion of cases with iNOS-positive tumor cells increased in in situ carcinomas from grade I to III (20.0%, 46.2%, and 73.3%). In invasive ductal carcinomas, there were more cases with iNOS-positive tumor cells than with in situ carcinomas (P = 0.04). Carcinomas with both iNOS-positive tumor and stromal cells had a higher apoptotic index (P = 0.02) and a higher calculated microvessel densities index (P = 0.02). A high number of iNOS-positive stromal cells associated with metastatic disease (P = 0.05). The results show that breast carcinoma cells, in addition to stromal cells, express iNOS and are capable of producing NO. Carcinomas with iNOS-positive tumor and stromal cells have a higher apoptotic indices and increased vascularization, suggesting that iNOS contributes to promotion of apoptosis and angiogenesis in breast carcinoma. The association of the number of iNOS-positive stromal cells with metastatic disease might be attributable to stimulation of angiogenesis, resulting in a higher vascular density and consequently a higher probability for tumor cells to invade. PMID- 10873094 TI - Immunohistochemical analyses of focal adhesion kinase expression in benign and malignant human breast and colon tissues: correlation with preinvasive and invasive phenotypes. AB - The focal adhesion kinase (FAK) is a protein tyrosine kinase linked to signaling events between cells and the extracellular matrix. Studies at the Western blot level have demonstrated up-regulation of FAK expression in invasive breast and colon cancers. To assess p125FAK expression at the cellular level, we developed monoclonal antibodies that specifically detected FAK in formalin-fixed, paraffin embedded tissue sections and analyzed the levels of FAK expression in human breast and colon tissues. Monoclonal antibody 4.47 demonstrated FAK-specific focal adhesion staining by immunofluorescence assays on BT-474 breast cancer cells and detected a Mr 125,000 protein by both Western blotting and immunoprecipitation analyses. Using immunohistochemical techniques, the expression of p125FAK was analyzed in 36 normal and 43 preinvasive or invasive human breast and colon tissues from individual patients. FAK was weakly expressed in most benign breast epithelium but was up-regulated at moderate or strong levels in 14 of 18 invasive breast carcinomas. In seven samples of ductal carcinoma-in situ, FAK was overexpressed. Borderline-to-weak expression of FAK was detected in the normal colonic epithelium. In the invasive colon cancers, FAK was overexpressed at moderate or strong levels in 13 of 15 tumors. Furthermore, FAK expression was up-regulated in areas of dysplastic, premalignant colon epithelium. These results provide the first evidence at the cellular level that FAK expression is variably overexpressed in breast and colon cancer and suggest that up-regulation occurs at an early stage of tumorigenesis. PMID- 10873095 TI - Cyclooxygenase-2 expression is up-regulated in transitional cell carcinoma and its preneoplastic lesions in the human urinary bladder. AB - Cyclooxygenase (COX) is a key enzyme in the synthesis of prostaglandins from arachidonic acid. Much evidence, including that from epidemiological and experimental studies, suggests that the inducible form of COX, COX-2, is increased in colon tumor tissues and is involved in colon cancer tumorigenesis. To determine the significance of COX-2 in tumorigenesis in the urinary bladder, the expression of COX-2 in transitional cell carcinoma and preneoplastic lesions of the bladder was examined. Tumor specificity of COX-2 immunoblotting was 100% in 12 of 35 (34%) tumors, but in 0 of the 10 normal urothelia samples. COX-2 expression was significantly correlated with tumor stage in 9 of 20 (45%) muscle invasive (pT2-4) tumors and in 3 of 15 (20%) superficially invasive (pT1) tumors (P < 0.05). Immunohistochemical examination revealed that 13 of 14 (93%) samples of carcinoma in situ (CIS), which may be the precursor of muscle-invasive-type tumors, expressed COX-2, whereas 10 of 21 (48%) samples of dysplasia, which may be the precursor of both superficially invasive and muscle-invasive tumors, expressed COX-2. From the expression profile of COX-2 in these various urothelia, it is suggested that COX-2 is involved in the development of transitional cell carcinoma of the urinary bladder, especially that of muscle-invasive tumors via CIS. Furthermore, COX-2 may be a therapeutic target for CIS because of the high expression rate of COX-2 in CIS lesions. PMID- 10873096 TI - Expression of vascular endothelial growth factors A, B, C, and D and their relationships to lymph node status in lung adenocarcinoma. AB - Vascular endothelial growth factors (VEGFs) C and D are novel members of the VEGF family that show some selectivity toward lymphatic endothelial cells. Recent studies suggest that VEGF-C may be involved in lymphangiogenesis and spread of cancer cells via lymphatic vessels. However, whether other VEGF family members play a role in lymph node metastasis is largely unknown. The aim of the present study was to explore whether expressions of VEGF-A, VEGF-B, VEGF-C, and VEGF-D are correlated with lymph node status in lung adenocarcinoma. Total RNA was isolated from 60 surgical specimens of lung adenocarcinoma with (n = 27) or without (n = 33) lymph node metastasis. The relative mRNA abundance of VEGF-A, VEGF-B, VEGF-C, and VEGF-D was measured by real-time reverse transcription-PCR analysis based on TaqMan fluorescence methodology. We found that, as single factors, expression of none of the four VEGF family members clearly correlated with the presence of lymph node metastasis. The only tendency noted was for higher VEGF-B and VEGF-C and lower VEGF-D levels in the node-positive group. However, two-way scatterplot analysis revealed that tumors with lymph node metastasis were associated with a pattern of low VEGF-D and high VEGF-A, VEGF-B, or VEGF-C, such that the ratios of VEGF-D:VEGF-A, VEGF-D:VEGF-B, or VEGF-D:VEGF-C were significantly lower in the node-positive group. Strikingly, none of the 11 tumors with high VEGF-D levels metastasized to lymph nodes. Furthermore, a low VEGF-D:VEGF-C ratio correlated with the presence of lymphatic invasion, and six of seven tumors with a pattern of very high expression of VEGF-C and low expression of VEGF-D displayed lymph vessel invasion that extended along the bronchovascular tree beyond the main tumor. Finally, levels of VEGF-A, but not VEGF-B or VEGF-C, were higher in tumors with large nodal metastasis (> or = 1 cm) than in those with small (< 1 cm) nodal metastasis. These results support the hypothesis that two VEGF family members are involved in lymph node metastasis at two distinct steps; VEGF-C facilitates entry of cancer cells into the lymph vasculature, whereas VEGF-A promotes the growth of metastatic tumor through angiogenesis. The results also suggest that the balance between VEGF-C and VEGF-D could be important rather than the level of VEGF-C alone. Whether a low VEGF-D level plays a causative role in lymph node metastasis requires further investigation. PMID- 10873097 TI - Association between polymorphism in p21(Waf1/Cip1) cyclin-dependent kinase inhibitor gene and human oral cancer. AB - The cyclin-dependent kinase inhibitor gene p21(Waf1/Cip1) plays a central role in inducing cellular growth arrest, terminal differentiation, and apoptosis. Alterations in this gene may adversely affect regulation of these processes and increase susceptibility for cancer. We have recently reported a novel polymorphism in the p21(Waf1/Cip1) gene in the Indian population and its association with esophageal cancer. An A-->G transition at codon 149 resulted in amino acid substitution from aspartate to glycine in the proliferating cell nuclear antigen binding COOH-terminal domain of p21(Waf1/Cip1) that may affect PCNA-p21(Waf1/Cip1) interactions, thereby affecting regulation of cellular proliferation, and may increase susceptibility for development of cancer. In a parallel study in our laboratory, we searched for putative p21(Waf1/Cip1) mutations in oral premalignant and malignant lesions. No somatic mutation was detected in exon 2 of p21(Waf1/Cip1). Interestingly, a codon 149 polymorphism variant (A-->G) was identified in 11 of 30 (37%) premalignant lesions (7 of 19 hyperplastic lesions and 4 of 11 dysplastic lesions) and 11 of 30 (37%) squamous cell carcinomas (SCCs). This codon 149 variant was also identified in paired lymphocytes of all of the patients with premalignant lesions and SCCs harboring the variant allele, suggesting the occurrence of a polymorphism. Lymphocyte DNA isolated from 50 unrelated age- and gender-matched healthy subjects was screened for this polymorphism. Seven of 50 (14%) normal controls harbored the A-->G codon 149 variant allele. Immunohistochemical analysis of p21(Waf1/Cip1) protein expression showed immunoreactivity in 19 of these 30 (63%) oral premalignant lesions and 16 of 30 (53%) SCCs. The most intriguing features of the study were: (a) the significant increase in frequency of this polymorphism not only in patients with oral SCCs (P = 0.038), but also in patients with premalignant lesions (P = 0.038), compared with normal controls; and (b) the significantly higher frequency of p21(Waf1/Cip1) variants (codon 149) in oral premalignant lesions (10 of 11 cases) and SCCs (11 of 11 cases) with wild-type p53 (P = 0.045) than in lesions with p53 mutations, suggesting that this polymorphism affects the p53 pathway and may play a vital role in oral tumorigenesis. Furthermore, overexpression of p21 protein in oral lesions harboring missense mutations in the p53 gene suggest a p53-independent role for p21 in the pathogenesis of oral cancer. PMID- 10873098 TI - Production and pro-apoptotic activity of soluble CD95 ligand in pancreatic carcinoma. AB - We report here that the progression of pancreatic carcinomas in tumor patients is associated with increased serum levels of both the soluble forms of CD95 ligand (CD95L/FasL) and its receptor, CD95 (Fas). Shedding of proteolytically processed soluble CD95L was also observed in pancreatic carcinoma cells in vitro, thus identifying one possible source of CD95L in patients' sera. Because the secreted forms of both CD95 and CD95L have been implicated previously in protection of cells from CD95-mediated cell death, we assessed the effect of soluble CD95L in supernatants of pancreatic carcinoma cells on viability of Jurkat T lymphocytes. We describe that (a) supernatants derived from cultured pancreatic carcinoma cells caused apoptosis of Jurkat cells; (b) soluble tumor-derived CD95L contributed significantly to this effect; and (c) in comparison to Jurkat cells, pancreatic carcinoma cells themselves revealed increased resistance to apoptosis induction by autocrine soluble CD95L. These results are consistent with the notion that in the microenvironment of pancreatic tumors, tumor-derived shed CD95L exerts paracrine pro-apoptotic effects. In addition, because it is released at high levels into the bloodstream, soluble CD95L may have systemic effects in tumor patients that reach beyond the microenvironment of the tumor site. PMID- 10873099 TI - Calpain inhibitor II induces caspase-dependent apoptosis in human acute lymphoblastic leukemia and non-Hodgkin's lymphoma cells as well as some solid tumor cells. AB - Calpain is a calcium-dependent cysteine protease that is implicated in calcium dependent cell death, and calpain inhibitors are generally considered as inhibitors of apoptosis. To the contrary, in the present study, we found that calpain inhibitor II (CPI-2) triggers rapid apoptosis in acute lymphoblastic leukemia (ALL) and non-Hodgkin's lymphoma (NHL) cells. All target cell lines were killed by CPI-2, including: ALL-1, a multidrug-resistant BCR-ABL fusion transcript-positive t(9;22) pro-B ALL cell line; RS4;11, a highly radiation resistant MLL-AF4 fusion transcript-positive t(4;11) pre-pre B ALL cell line; RAMOS, a highly radiation-resistant and p53-deficient Burkitt's lymphoma cell line; DAUDI, a Burkitt's leukemia/lymphoma cell line; NALM-6, a pre-B ALL cell line; and JURKAT and MOLT-3, two T-lineage ALL/NHL cell lines. CPI-2-induced apoptosis in LYN-deficient and BTK-deficient subclones of the DT-40 lymphoma B cell line as effectively as it did in wild-type DT-40 cells. Thus, CPI-2-induced apoptosis is not dependent on the protein tyrosine kinases LYN or BTK. Notably, caspase inhibitor I effectively inhibited CPI-2-induced apoptosis, suggesting that the inhibition of a CPI-2-susceptible protease results in caspase activation, leading to apoptosis in ALL/NHL cells. Unlike the high calpain expressing ALL/NHL cell lines, myeloid leukemia cell lines HL-60/AML, K562/CML, and U937/AMML, or solid tumor cell lines BT-20/breast cancer, PC-3/prostate cancer, U373/glioblastoma, and HeLa/epitheloid cancer, were not susceptible to the cytotoxicity of CPI-2. Taken together, our results identify calpain as a new molecular target for the treatment of ALL and NHL. CPI-2 and its analogues represent a promising new class of antileukemia/lymphoma agents that deserves further development. PMID- 10873100 TI - Antimetastatic intraoperative chemotherapy of human colon tumors in the livers of nude mice. AB - We have developed a new antimetastatic chemotherapeutic strategy for combination with hepatic resection of human colon cancers in a high-metastasis nude mouse model. The new procedure involves i.p. administration of 5-fluorouracil (5-FU) 2 h before hepatic resection of the human colon tumors, with therapy continued postoperatively for 4 consecutive days. We termed this strategy neo-neoadjuvant chemotherapy. The regime significantly prolonged animal survival compared with preoperative 5-FU neoadjuvant therapy, 5-FU postoperative adjuvant therapy, surgery alone, 5-FU without surgery, or the untreated control. The median survival of neo-neoadjuvant i.p. 5-FU-treated group was 81 days, compared with 27 days for the control group (P < 0.009). The median survival of animals in the neoadjuvant group was 37 days (P < 0.021 compared with the control group). There was also a significant difference between the median survival of neo-neoadjuvant, and the neoadjuvant group (P < 0.031). When all animals in the control group had died, 70% of animals with neo-neoadjuvant and 60% of animals with neoadjuvant 5 FU were still alive (P < 0.003 and P < 0.011, respectively). When all animals with neoadjuvant 5-FU treatment had died, 70% of animals with neo-neoadjuvant treatment were still alive (P < 0.003). Survival of all other treatment groups, including 5-FU without surgery, surgery alone, and adjuvant postoperative chemotherapy, was not significantly different from the untreated control group. Two animals in the neo-neoadjuvant group were free of tumors when sacrificed at days 154 and 165 post surgery. Whereas 100% of animals in the control, 90% in the 5-FU alone, 70% in the surgery alone, 60% in the 5-FU adjuvant, and 40% in the neoadjuvant groups had metastases in the lymph nodes draining the liver, only 10% of animals in the neo-neoadjuvant group had metastases. These data suggest that the neo-neoadjuvant therapy increased survival by preventing metastasis of cancer cells not removed in the liver resection procedure. The results of this study indicate that the neo-neoadjuvant treatment strategy for resection of colon cancer liver metastasis should be explored clinically. PMID- 10873101 TI - Successful treatment of intracranial gliomas in rat by oligodeoxynucleotides containing CpG motifs. AB - Phosphorothioate oligodeoxynucleotides with CpG motifs (CpG-ODNs) activate various immune cell subsets and induce production of numerous cytokines. To evaluate whether CpG-ODNs can induce rejection of established tumors, Lewis rats were inoculated intracerebrally with syngeneic CNS-1 glioma cells and subsequently injected with CpG-ODNs into the tumor bed. Although all of the control rats (n = 14) died within 23 days, 88% of the animals (n = 8) treated with a single CpG-ODN injection 5 days after tumor inoculation showed long-term survival (>90 days; P < 0.002). CpG-ODNs increased tumoral infiltration with macrophage/microglial cells, CD8, and natural killer lymphocytes. CpG-ODN-cured animals were further protected against a second tumor challenge. CpG-ODNs had no effect on a s.c. CNS1 tumor in nude mice, which suggested that CpG-ODN is not directly cytotoxic and that immunostimulation is required for the antitumoral effect. These findings suggest that intratumoral injections of CpG-ODNs represent a new immunotherapeutic approach in human gliomas, which overcome the need for the selection and purification of a tumoral antigen. PMID- 10873102 TI - Cytotoxic chemotherapy regimens that increase dose per cycle (dose intensity) by extending daily dosing from 5 consecutive days to 28 consecutive days and beyond. AB - Dose intensity, defined as dose administered per unit time, has emerged as a potentially important measurement of anticancer drug exposure and determinant of efficacy. There are several strategies for increasing dose intensity, one being a protracted daily dosing strategy without major dose reduction for toxicity. This strategy involves continued therapy during periods of recovery from reversible toxicity, and it inherently challenges our understanding that renewing tissues cannot repopulate (recover) in the continued presence of cytotoxic drug. We have tested this idea directly in a murine preclinical trial. Specifically, we have tested whether acutely myelotoxic doses of gemcitabine (i.p. injection, 6.0 mg/m2/day), acetyldinaline [CI-994; GOE 5549; PD 123 654; 4-acetylamino-N-(2' aminophenyl)-benzamide, 150 mg/m2/day p.o.], and/or melphalan (i.p. injection, 7.2 mg/m2/day) can be tolerated for 28 consecutive days and whether suppressed bone marrow function recovers despite this protracted daily therapy. The three drugs all caused acute neutropenia and suppression of medullary hematopoiesis. Damage to progenitor populations exposed to acetyldinaline and gemcitabine was not as severe as that caused by melphalan, in which case absolute neutrophil count, mature progenitors (colony-forming unit granulocyte/macrophage), and immature progenitors (colony-forming unit-S) progressively declined to severely depressed levels. Marrow recovery was observed during continued daily treatment with acetyldinaline and gemcitabine but not melphalan, and marrow function completely recovered after finishing the 28-day course. Pharmacology studies proved that protracted therapy causes little, if any, change in cellular drug tolerance or systemic exposure. PMID- 10873103 TI - Constitutive and lysophosphatidic acid (LPA)-induced LPA production: role of phospholipase D and phospholipase A2. AB - Ascitic fluid and plasma from ovarian cancer patients, but not from patients with nongynecological tumors, contain elevated levels of the bioactive phospholipid lysophosphatidic acid (LPA). We show that ovarian cancer cells constitutively produce increased amounts of LPA as compared with normal ovarian epithelium, the precursor of ovarian epithelial cancer, or breast cancer cells. In addition, LPA, but not other growth factors, increases LPA production by the OVCAR-3 ovarian cancer cell line but not by normal ovarian epithelium or breast cancer cell lines. We show that phospholipase D activity contributes to both constitutive and LPA-induced LPA production by ovarian cancer cells. Constitutive and LPA-induced LPA synthesis by ovarian cancer cells is differentially regulated with respect to the requirement of specific phospholipase A2 (PLA2) subgroups. Group IB (pancreatic) secretory PLA2 plays a critical role in both constitutive and LPA induced LPA formation, whereas group IIA (synovial) secretory PLA2 contributes to LPA-induced LPA production only. Calcium-dependent and/or -independent cytosolic PLA2s are required for constitutive LPA synthesis but do not play a role in LPA induced LPA formation. LPA increases the proliferation of ovarian cancer cells, decreases sensitivity to cisplatin, the most commonly used drug in ovarian cancer, decreases apoptosis and anoikis, increases protease production, and increases production of neovascularization mediators. Thus, an understanding of the source and regulation of LPA production in ovarian cancer patients could identify novel targets for therapy. PMID- 10873104 TI - Eradication of human non-Hodgkin's lymphoma in SCID mice by BCL-2 antisense oligonucleotides combined with low-dose cyclophosphamide. AB - Cancers overexpressing Bcl-2 protein, which prevents programmed cell death (apoptosis), are less sensitive to stresses that produce cellular damage, including chemotherapy. If the level of Bcl-2 protein can be reduced sufficiently using antisense oligonucleotides (ASOs) targeting the gene message, then cytotoxic agents may be rendered more effective in eliminating disease and increasing cure rate. Preclinical studies in SCID mice bearing Bcl-2 overexpressing systemic human B-cell lymphoma (DoHH2) were undertaken to support development of a clinical trial. These data confirm that a combination of an ASO (5 mg/kg) targeting bcl-2 and a low dose of cyclophosphamide (35 mg/kg) was an effective strategy, leading to the eradication of the DoHH2 cells in vivo and cure of the animals. When mice deficient in natural killer cell activity were treated with an ASO, similar results were observed, suggesting that ASO stimulation of the host immune system was not a significant factor in elimination of lymphoma cells. These studies indicate that therapeutic strategies involving the use of an ASO targeting bcl-2 in combination with a cytotoxic agent may improve clinical outcomes. PMID- 10873105 TI - The relationship between intracellular and extracellular pH in spontaneous canine tumors. AB - Recently, it has been suggested that the cellular uptake of chemotherapeutic drugs may be dependent on the pH gradient between the intracellular (pHi) and extracellular (pHe) compartments. It has been demonstrated in murine tumor models that the extracellular environment is acidic, relative to the intracellular environment, thus favoring preferential accumulation of drugs that are weak acids into cells. However, concomitant measurements of pHi and pHe in spontaneous tumors have not been reported, so it is not certain how well the murine results translate to the clinical scenario. In this study, both types of measurements were performed in dogs with spontaneous malignant soft tissue tumors. On average, pHe was more acidic than pHi, with maintenance of a more physiologically balanced intracellular tumor environment. However, the magnitude of the gradient varied widely, and individual tumors had both positive and negative pH gradients (pHi - pHe). These data suggest that the magnitude and direction of the pH gradient may need to be measured for individual patient tumors and/or that manipulation of pHe may be required if exploitation of the pH gradient is to be achieved for tumor selective augmentation of intracellular drug delivery. PMID- 10873106 TI - Oral antisense that targets protein kinase A cooperates with taxol and inhibits tumor growth, angiogenesis, and growth factor production. AB - Protein kinase A type I (PKAI) transduces mitogenic signals from different growth factors and oncogenes and is overexpressed in the majority of human cancers. We and other investigators previously have reported that different PKAI inhibitors, including antisense oligonucleotides, have antitumor activity. In this study, we used a novel hybrid DNA/RNA mixed-backbone oligonucleotide (MBO) targeting the PKAI subunit RIalpha. We demonstrated that after oral administration, the MBO antisense RIalpha inhibited the growth of human colon cancer xenografts in nude mice and showed a cooperative antitumor effect with Taxol, which outlasted treatment withdrawal and significantly prolonged survival of mice compared with untreated controls or to single-agent-treated mice. Immunohistochemical analysis of tumor specimens showed inhibition of target protein RIalpha and of growth factor expression along with a marked inhibition of angiogenesis and an increase in p27 expression. In conclusion, a novel MBO that targets PKAI, administered p.o., is effective and cooperates with the anticancer drug Taxol on both tumor growth and expression of factors involved in the control of cell proliferation, cell cycle, and angiogenesis. Because the MBO described has completed a phase I trial involving i.v. injection in cancer patients, these results provide the biological rationale of its activity after oral administration and may be translated into a therapeutic strategy in a clinical setting. PMID- 10873107 TI - Enhancement of intrinsic tumor cell radiosensitivity induced by a selective cyclooxygenase-2 inhibitor. AB - The antitumor effects of the selective cyclooxygenase (COX)-2 inhibitor SC-236 alone and in combination with radiation were investigated using the human glioma cell line U251 grown in monolayer culture and as tumor xenografts. On the basis of Western and Northern blot analyses, these cells express COX-2 protein and mRNA to levels similar to those in the human colon carcinoma cell line HT29. Treatment of U251 cells in monolayer culture with 50 microM SC-236 resulted in a time dependent decrease in cell survival as determined by a clonogenic assay. The cell death induced by SC-236 was associated with apoptosis and the detachment of cells from the monolayer. After 2 days of drug treatment, the cells that remained attached were exposed to graded doses of radiation, and the clonogenic assay was performed. Comparison of the survival curves for drug-treated and untreated cultures revealed that SC-236 enhanced radiation-induced cell death. In these combination studies, SC-236 treatment resulted in a dose-enhancement factor of 1.4 at a surviving fraction of 0.1, with the surviving fraction at 2 Gy (SF2) reduced from 0.61 to 0.31. These data indicate that in vitro SC-236 induces U251 apoptotic cell death and enhances the radiosensitivity of the surviving cells. To extend these investigations to an in vivo situation, U251 glioma cells were grown as tumor xenografts in the hind leg of nude mice, and SC-236 was administered in drinking water. SC-236 alone slowed tumor growth rate, and when administered in combination with local irradiation, SC-236 caused a greater than additive increase in tumor growth delay. These in vitro and in vivo results suggest that the selective inhibition of COX-2 combined with radiation has potential as a cancer treatment. PMID- 10873108 TI - Purging of epithelial tumor cells from peripheral blood stem cells by means of the bispecific antibody BIS-1. AB - Peripheral blood stem cell (PBSC) support in breast cancer patients allows high dose chemotherapy, but tumor cell contamination of the PBSCs is a potential source of relapse. Specific carcinoma cell killing can be obtained by retargeting activated T cells with bispecific antibody BIS-1, directed against epithelial glycoprotein-2 and CD3. To purge epithelial tumor cells from the PBSCs of breast cancer patients, activation of T cells in PBSCs and T-cell retargeting by BIS-1 was studied. PBSCs, obtained by leukapheresis after chemotherapy and recombinant human granulocyte colony-stimulating factor, were cultured in the presence of PBS, interleukin-2, OKT3, or interleukin-2/OKT3 for induction of T-cell activation. Subsequently, lysis of epithelial tumor cell lines by activated T cells of PBSCs in the presence or absence of BIS-1 was assessed with the 51Cr release assay or immunocytochemical staining. The effect on PBSC hematopoietic colony formation (HCF) was evaluated by the granulocyte macrophage colony stimulating units assay. Prior to activation, PBSCs from breast cancer patients contained higher levels of CD8+ T cells than peripheral blood from healthy volunteers (P < 0.05). The potential of PBSCs to sustain tumor cell lysis was increased after all prior activations and was further enhanced by BIS-1. Maximal BIS-1 effect was observed after OKT3 activation of PBSCs for 72 h (P < 0.0005), inducing a >3 log depletion of tumor cells. HCF was not affected by prior OKT3 activation and/or BIS-1. In conclusion, specific tumor cell lysis by PBSCs can be obtained in vitro by OKT3 activation and BIS-1 retargeting of T cells, without affecting HCF. At present, studies are evaluating this format for future clinical application. PMID- 10873109 TI - Pegylated liposomes have potential as vehicles for intratumoral and subcutaneous drug delivery. AB - The potential value of intratumoral or s.c. injections of pegylated liposomes as locoregionally targeted therapy of tumors and their draining lymph nodes was assessed in nude mice as part of an ongoing program aimed at developing pegylated liposomal radiosensitizers for the treatment of head and neck cancers. Animals received (111)In-labeled diethylenetriaminepentaacetic acid (DTPA), either encapsulated in pegylated liposomes (IDLPL) or in the unencapsulated form ((111)In-DTPA), as intratumoral or s.c. injections, and the local retention, locoregional nodal drainage, and systemic biodistribution were measured. After intratumoral injections, IDLPL were effectively retained in the tumor with an area under the curve (AUC) between 1 and 96 h of 2,574.4% injected dose per gram hours (%ID/g x h). The corresponding value for (111)In-DTPA was 204.4%ID/g x h. Accumulation of IDLPL was seen in ipsilateral lymph nodes. The maximal ipsilateral:contralateral node ratios were 8:1 (2.2 versus 0.27%ID/g) for inguinal nodes at 24 h and 19:1 (2.5 versus 0.13%ID/g) for axillary nodes at 48 h. Unencapsulated (111)In-DTPA showed no evidence of accumulation in locoregional nodes. After s.c. injection, IDLPL were cleared slowly from the injection site with an AUC between 1 and 192 h of 24,051.1%ID/g x h. Unencapsulated (111)In-DTPA was cleared rapidly with an AUC between 1 and 192 h of 46.4%ID/g x h. Again, significant levels of IDLPL were detected in the ipsilateral locoregional nodes, with ipsilateral:contralateral ratios of 121:1 (57.9 versus 0.48%ID/g) at 24 h (inguinal nodes) and 17:1 (5.2 versus 0.3%ID/g) at 72 h (axillary nodes). There was no retention of unencapsulated (111)In-DTPA in the draining nodes. Locoregional administration of pegylated liposomal radiosensitizers may be a useful approach for targeted therapy of head and neck tumors and their nodal metastases. PMID- 10873110 TI - Comparison of thymidylate synthase (TS) protein up-regulation after exposure to TS inhibitors in normal and tumor cell lines and tissues. AB - Thymidylate synthase (TS) is an important target for cancer chemotherapy. However, several mechanisms of resistance to TS inhibitors have been described. One mechanism that may be relevant to short-term exposure to TS inhibitors occurs as a result of disruption of the autoregulatory loop, which allows TS to control its own translation. This disruption leads to up-regulation of TS protein and is generally thought to decrease efficacy. This study has investigated TS protein up regulation using a range of TS inhibitors in both tumor and nonmalignant cell lines in vitro and in vivo. Up-regulation of TS protein showed a time-, dose-, and cell-type-specific response to treatment with ZD9331. This response was observed in W1L2 cells treated for 24 h at equitoxic doses of raltitrexed (6 fold), ZD9331 (10-fold), fluorouracil (5-fold), LY231514 (7-fold), AG337 (7 fold), and BW1843U89 (3-fold). Up-regulation was observed over a range of doses. Elevation of TS protein only persisted up to 12 h after removal of drug. The extent of induction does not depend on basal TS levels. Nontransformed human fibroblasts showed significantly greater up-regulation of TS protein than tumor cells exposed to an equitoxic dose of ZD9331. In vivo experiments using the L5178Y thymidine kinase -/- mouse lymphoma implanted into DBA2 mice also showed greater up-regulation of TS protein in normal intestinal epithelial cells compared with tumor cells. These results confirm that TS up-regulation is a common feature of TS inhibition in tumor cells and that it may occur to a greater extent in normal tissues, although the clinical implications of these findings remain to be determined. PMID- 10873111 TI - A novel bispecific antisense oligonucleotide inhibiting both bcl-2 and bcl-xL expression efficiently induces apoptosis in tumor cells. AB - Bcl-2 and Bcl-xL are inhibitors of apoptosis frequently overexpressed in solid tumors. The bcl-2 and bcl-xL mRNAs share a region of homology comprising nucleotides 605-624 and 687-706, respectively, which differs by only three nucleotides. This sequence does not occur in the proapoptotic splice variant bcl xS. To test the possibility that oligonucleotides targeting this region have the potential to down-regulate bcl-2 and bcl-xL expression simultaneously, three 2'-O methoxy-ethoxy-modified phosphorothioate oligonucleotides were designed. These oligonucleotides differed in the number of mismatches to bcl-2 and bcl-xL and in the number of nucleotides to which the modifications were made. The effects of these oligonucleotides on bcl-2 and bcl-xL expression, as well as their abilities to induce apoptosis, were assessed in small cell and non-small cell lung cancer cell lines expressing different basal levels of bcl-2 and bcl-xL. Although all oligonucleotides down-regulated bcl-2 and bcl-xL expression, oligonucleotide 4625, which has no mismatching nucleotides to bcl-2 but three to bcl-xL, two of which were modified by 2'-O-methoxy-ethoxy residues, showed the strongest bispecific activity on the transcript and protein level. In all cell lines this bispecific activity induced apoptotic cell death, as demonstrated by increased uptake of propidium iodide, a 10-100-fold increase in caspase-3-like protease activity, and nuclear condensation and fragmentation. This is the first report of a bcl-2/bcl-xL bispecific antisense oligonucleotide that deserves attention as a therapeutic compound in lung cancer and other malignancies in which bcl-2 and/or bcl-xL are overexpressed. PMID- 10873112 TI - An orthotopic mouse model of remetastasis of human colon cancer liver metastasis. AB - Whether liver metastases from colon cancer are capable of metastasizing to other sites is an important question in surgical oncology. To answer this question, we have developed a highly metastatic orthotopic transplant model of a liver metastasis from a human colon cancer patient in nude mice that targets the liver and lymph nodes. The metastatic human tumor was transplanted in athymic nude mice by surgical orthotopic implantation (SOI) of a liver metastasis from a colon cancer patient. The human colon tumor was then subsequently implanted in the colon by SOI or, in an additional series of nude mice, in the liver by surgical hepatic implantation (SHI). The mice were then explored over time for lymph node involvement beginning 10 days after implantation. After SOI, 100% of the animals had liver metastasis within 10 days, and subsequently, 19 days after SOI, all lymph nodes draining the liver were involved with metastasis without any retroperitoneal or lung tissue involvement. After SHI, all sites of lymphatic drainage of the liver, including portal, celiac, and mediastinal lymph nodes, were massively involved by metastasis in 100% of the animals as early as 10 days after tumor implantation on the liver. The results of this study demonstrate that liver metastases from colon cancer are capable of remetastasizing to other sites. This study thus suggests that in colon cancer patients with liver metastasis, mediastinal, celiac, and portal lymph node metastases originate from the liver metastasis and not, as previously thought, from primary colon cancer. PMID- 10873113 TI - The relevance of cell proliferation, vascular endothelial growth factor, and basic fibroblast growth factor production to angiogenesis and tumorigenicity in human glioma cell lines. AB - Tumor growth is partially dependent on angiogenesis, a process that relies on angiogenic factors. Tumorigenicity of cancer cells is thought to be associated with the production of various angiogenic factors that stimulate or inhibit the rate of endothelial cell migration and proliferation. However, the relative importance of specific individual factors originally studied in cancer cell lines has yet to be determined in vivo. In this study, we examined seven human glioma cell lines for dynamic changes of two major angiogenic factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), and for doubling time and tumorigenicity in nude mice. Various correlation studies demonstrated that in these glioma cell lines, VEGF expression correlated well with RBC density in tumor sections (r2 = 0.804) and with average tumor weight (r2 = 0.987). In contrast, bFGF expression in the observed glioma cell lines did not correlate with tumorigenicity (r2 = 0.001) or with VEGF expression (r2 = 0.255). Furthermore, there was no correlation between doubling time and tumorigenicity in these cell lines (r2 = 0.160). Taken together, these results suggest that VEGF plays a major role in glioma formation and that down-regulation of VEGF, rather than bFGF, would be a more effective choice for glioma gene therapy. PMID- 10873114 TI - Nuclear factor-kappaB activity correlates with growth, angiogenesis, and metastasis of human melanoma cells in nude mice. AB - The purpose of this study was to determine the role of nuclear factor (NF) kappaB/relA activity in the induction of angiogenesis and production of metastasis by human melanoma cells. Highly metastatic melanoma variant cells expressed high levels of constitutive NF-kappaB activity. Transfection of highly metastatic human melanoma variant cells with a dominant-negative mutant inhibitor of nuclear factor-kappaB alpha (Ikappabeta alpha) expression vector (Ikappabeta alphaM) decreased the level of constitutive NF-kappaB activity, inhibited s.c. tumor growth, and prevented lung metastasis in nude mice. Furthermore, the slow growing s.c. tumors formed by the IkappaB alphaM-transfected cells exhibited a decrease in microvessel density (angiogenesis), which correlated with a decrease in the level of interleukin-8 expression. Collectively, these results demonstrate that NF-kappaB/reLA activity significantly contributes to tumorigenicity, angiogenesis, and metastasis of human melanoma cells implanted in nude mice. PMID- 10873116 TI - Polycystic ovary syndrome: a single gene mutation or an evolving set of symptoms. AB - Polycystic ovary syndrome is one of the most common endocrinopathies among women. Nevertheless, there is no one single acceptable definition for this syndrome, its pathophysiology is not completely understood and its etiology remains an enigma. Several studies have examined the genetic basis of polycystic ovary syndrome with varying results, probably caused by the different criteria used to define the syndrome. These studies, together with studies that deal with reclassification of polycystic ovary syndrome, are described in the following review. Furthermore, a simplified alternative approach to this symptom complex is proposed. PMID- 10873115 TI - Insulin-like growth factors in obstetrics. AB - Data from a number of studies reported during the past two decades indicate that the insulin-like growth factor (IGF) system, including IGF-I and IGF-II, their receptors and six high-affinity binding proteins, is involved in the control of foetal and placental growth and development. Recent studies that addressed the role of the IGF system in pregnancy and the clinical usefulness of IGF and IGF binding protein measurements in obstetrics are reviewed and discussed. PMID- 10873117 TI - Emergency contraception: methods and efficacy. AB - A number of effective and safe methods for emergency contraception are now available. High doses of oestrogens, although effective, are seldom used nowadays because of the high incidence of nausea and vomiting, and the need for administration for 5 days. The Yuzpe regimen, consisting of administration of two doses of combined oral contraceptive pills with a 12-h interval, can prevent more than 74% of expected pregnancies, but the incidence of side effects, mainly gastrointestinal side effects, is high. Levonorgestrel and mifepristone are more effective than the Yuzpe regimen and have a lower incidence of side effects. They can prevent about 85% of pregnancies. The efficacy of both the Yuzpe regimen and levonorgestrel decreased with increase in the intercourse-treatment interval. The dose of mifepristone can be reduced to 10 mg without loss of efficacy. Both levonorgestrel or mifepristone are not yet widely available, and the Yuzpe regimen remains the only hormonal method in many countries. The postcoital insertion of an intrauterine contraceptive device is also a highly effective method, which can prevent over 90% of pregnancies. PMID- 10873118 TI - Direct vascular effects of estrogens and selective estrogen receptor modulators. AB - The aim of this review is to provide an update on the latest advancements in the field of the action of estrogens on the cardiovascular system, and particularly on the molecular mechanisms of the direct effects of these hormones and of some of the new synthetic selective estrogen receptor modulators on the vascular wall. PMID- 10873119 TI - Postmenopausal osteoporosis management. AB - Osteoporosis is perhaps the widest-ranging social, physical, and economic impact of estrogen deficiency. Postmenopausal bone loss is the major determinant of osteoporosis. Osteoporotic risk can be determined by measuring bone mineral density using dual X-ray absorptiometry. The radiation free quantitative bone ultrasound is emerging in the assessment of bone structure giving reliable estimates of fracture risk. Diet and exercise are important in determining a woman's risks for osteoporosis. Hormone replacement therapy clearly decreases bone turnover and prevents postmenopausal bone loss and reduces fractures. Tibolone as well as raloxifene prevent bone loss and solid data demonstrate a reduction of vertebral fractures after raloxifene administration. PMID- 10873120 TI - Current status of subfertility care: time to rationalize treatment options. PMID- 10873121 TI - New concepts in superovulation strategies for assisted conception treatments. AB - Controlled ovarian stimulation for in-vitro fertilization treatment using preparations that contain follicle-stimulating hormone has been routinely performed since the 1980s. The early preparations were urinary human menopausal gonadotrophins, containing follicle-stimulating hormone and luteinizing hormone. In the early 1990s, highly purified follicle-stimulating hormone preparations were introduced because of a desire to provide drugs for subcutaneous administration with a lower risk of allergic reactions. Intensive research resulted in the discovery of recombinant follicle-stimulating hormone, which is more potent that the highly purified follicle-stimulating hormone, resulting in significantly higher clinical pregnancy rates. The two recombinant follicle stimulating hormone preparations available appear to be equally effective and provide comparable results. Gonadotrophin-releasing hormone antagonists, which have recently been introduced, appear to be effective in preventing a premature rise in luteinizing hormone during ovarian stimulation for in-vitro fertilization, as well as improved response to lower doses of gonadotrophins. It is envisaged that the availability of recombinant gonadotrophins and gonadotrophin-releasing hormone antagonists will ultimately lead to shorter, cheaper and safer treatments, using reduced dosages. PMID- 10873122 TI - Genetics and male infertility. AB - This article reviews chromosomal and genetic disorders in the context of male fertility. Particular emphasis is on those disorders, which are encountered, in clinical practice including Klinefelter's syndrome, Kallman's syndrome, Androgen insensitivity, Y microdeletions, Y fertility gene deletions, and cystic fibrosis gene mutations. These disorders are discussed in relation to the aetiology of male fertility and also risks to children who are born of fathers with these disorders. A list of fathers' categories is proposed for outcome studies for children born after IVF-ICSI. Finally a question is proposed to catalyse debate about germ line therapy. PMID- 10873123 TI - Advanced ultrasound examination in the management of subfertility. AB - The development of new ultrasound techniques such as colour power angiography (CPA) and three-dimensional ultrasound may expand the role of ultrasound in the management of the infertile couple. Measurements of follicular blood flow velocities appears to predict the development of a healthy oocyte while high ovarian stromal vascularity is associated with polycystic ovary and the risk of ovarian hyperstimulation syndrome. Three-dimensional ultrasound with CPA provides spatial evaluation of volume flow in pelvic organs which has been used to measure vascular changes in the endometrium and improve the evaluation of tubal patency. PMID- 10873124 TI - Cost-effective, evidence-based infertility care. AB - Economic evaluation of health care interventions is becoming increasingly important as the demand for services increases. For unexplained infertility, in vitro fertilization is not a cost-effective first-line treatment option compared with ovarian stimulation and intrauterine insemination. In clomiphene citrate resistant polycystic ovarian syndrome, in-vitro fertilization may be cost effective. PMID- 10873125 TI - Bibliography. Current world literature. Reproductive endocrinology. PMID- 10873126 TI - Bibliography. Current world literature. Fertility. PMID- 10873127 TI - Epidemiology of transmissible diseases after elimination. AB - Elimination of an infectious disease is often understood to mean the total absence of cases in a population. This situation can occur only if the entire population is immune as a result of either natural disease or vaccination. However, this costly and unrealistic scenario is not necessary to ensure elimination, more appropriately defined as a situation in which sustained transmission cannot occur and secondary spread from importations of disease will end naturally, without intervention. The authors describe the size and duration of outbreaks caused by imported infections after indigenous transmission has been eliminated. They show that the status of the elimination process can be monitored by assessing the proportion of cases imported and the distribution of outbreak sizes. Measles in Canada, the United States, and the United Kingdom provides a good example of the relevance of these criteria. Surveillance of the size and duration of these outbreaks enables maintenance of elimination to be monitored. PMID- 10873128 TI - Is systemic lupus erythematosus, amyotrophic lateral sclerosis, or fibromyalgia associated with Persian Gulf War service? An examination of Department of Defense hospitalization data. AB - Since the Persian Gulf War ended in 1991, veterans have reported diverse, unexplained symptoms. Some have wondered if their development of systemic lupus erythematosus, amyotrophic lateral sclerosis, or fibromyalgia might be related to Gulf War service. The authors used Cox proportional hazard modeling to determine whether regular, active-duty service personnel deployed to the Persian Gulf War (n = 551,841) were at increased risk of postwar hospitalization with the three conditions compared with nondeployed Gulf War era service personnel (n = 1,478,704). All hospitalizations in Department of Defense facilities from October 1, 1988, through July 31, 1997, were examined. With removal of personnel diagnosed with any of the three diseases before August 1, 1991, and adjustment for multiple covariates, Gulf War veterans were not at increased risk of postwar hospitalization due to systemic lupus erythematosus (risk ratio (RR) = 0.94, 95% confidence interval (CI): 0.65, 1.35). Because of the small number of cases and wide confidence limits, the data regarding amyotrophic lateral sclerosis were inconclusive. Gulf War veterans were slightly at risk of postwar hospitalization for fibromyalgia (RR = 1.23, 95% Cl: 1.05, 1.43); however, this risk difference was probably due to the Gulf War veteran clinical evaluation program beginning in 1994. These data do not support Gulf War service and disease associations. PMID- 10873129 TI - Cigarette smoking and suicide: a prospective study of 300,000 male active-duty Army soldiers. AB - The authors examined the relation between cigarette smoking and suicide by conducting a cohort study of 300,000 male US Army personnel followed prospectively from January 1987 through December 1996 for 961,657 person-years. They found that the risk of suicide increased significantly with the number of cigarettes smoked daily (p for trend < 0.001). In multivariable-adjusted analyses, smokers of more than 20 cigarettes a day, compared with never smokers, were more than twice as likely to commit suicide. For male active-duty army personnel, the dose-related association between smoking and suicide was not entirely explained by the greater tendency of smokers to be White, drink heavily, have less education, and exercise less often. PMID- 10873130 TI - Education and the risk for Alzheimer's disease: sex makes a difference. EURODEM pooled analyses. EURODEM Incidence Research Group. AB - The hypothesis that a low educational level increases the risk for Alzheimer's disease remains controversial. The authors studied the association of years of schooling with the risk for incident dementia and Alzheimer's disease by using pooled data from four European population-based follow-up studies. Dementia cases were identified in a two-stage procedure that included a detailed diagnostic assessment of screen-positive subjects. Dementia and Alzheimer's disease were diagnosed by using international research criteria. Educational level was categorized by years of schooling as low (< or =7), middle (8-11), or high (> or =12). Relative risks (95% confidence intervals) were estimated by using Poisson regression, adjusting for age, sex, study center, smoking status, and self reported myocardial infarction and stroke. There were 493 (328) incident cases of dementia (Alzheimer's disease) and 28,061 (27,839) person-years of follow-up. Compared with women with a high level of education, those with low and middle levels of education had 4.3 (95% confidence interval: 1.5, 11.9) and 2.6 (95% confidence interval: 1.0, 7.1) times increased risks, respectively, for Alzheimer's disease. The risk estimates for men were close to 1.0. Finding an association of education with Alzheimer's disease for women only raises the possibility that unmeasured confounding explains the previously reported increased risk for Alzheimer's disease for persons with low levels of education. PMID- 10873131 TI - Body mass index and delayed conception: a European Multicenter Study on Infertility and Subfecundity. AB - Obesity has become a health problem in affluent societies, but few studies have investigated its effect on subfertility. Previous studies were based on select groups of women, focused mainly on ovulatory dysfunctions, and yielded controversial results. The authors evaluated the effect of body mass index on delayed conception by using a European population-based survey of pregnant women from five countries. Delayed conception was defined as a time to pregnancy that exceeded 9.5 months of unprotected intercourse. During 1992, 4,035 pregnant women from well-defined geographic areas were recruited consecutively at antenatal clinics or hospitals after at least 20 weeks of gestation. For women smokers, after adjustment for sociodemographic, biologic, and lifestyle-related factors, there was a strong association between obesity (body mass index of > or =30 kg/m2) and delayed conception (odds ratio = 11.54, 95% confidence interval: 3.68, 36.15) and also an increased risk for women whose body mass index was <20 kg/m2 (odds ratio = 1.70; 95% confidence interval: 1.01, 2.83). The same analysis conducted for women nonsmokers showed no association. The authors concluded that for women who achieve a clinically detectable pregnancy, those who are underweight or obese require a longer time to conceive only if they also smoke. PMID- 10873132 TI - Enigma of maternal race and infant birth weight: a population-based study of US born Black and Caribbean-born Black women. AB - The authors used 1985-1990 Illinois' vital records to determine the low birth weight components of infants delivered to US-born Black women, Caribbean-born Black women, and US-born White women. The moderately low birth weight rate (1,500 2,499 g) was 10% for infants with US-born Black mothers (n = 67,357) and 6% for infants with Caribbean-born mothers (n = 2,265) compared with 4% for infants with US-born White mothers (n = 34,124); the relative risk equaled 2.7 (95% confidence interval (CI): 2.5, 2.8) and 1.7 (95% CI: 1.4, 2.0), respectively. The very low birth weight rate (<1,500 g) was 2.6% for infants delivered to US-born Black women and 2.4% for infants to Caribbean-born women compared with 0.7% for infants to US-born White women; the relative risk equaled 3.6 (95% CI: 3.1, 4.1) and 3.3 (95% CI: 2.5, 4.4), respectively. Among the lowest risk mothers, the relative risk of moderately low birth weight for infants with US-born Black mothers and Caribbean-born mothers (compared with US-born White mothers) was 2.7 (95% CI: 2.1, 3.4) and 1.2 (95% CI: 0.4, 3.1), respectively; the relative risk of very low birth weight for infants with US-born Black mothers and Caribbean-born mothers was 6.7 (95% CI: 3.8, 12) and 4.2 (95% CI: 1.0, 18), respectively. The authors conclude that Caribbean-born women and US-born Black women have disparate moderate rates but equivalent very low birth weight rates. PMID- 10873133 TI - Long term relations between earthquake experiences and coronary heart disease risk factors. AB - The authors analyzed the relations between a variety of earthquake-related experiences incurred in 1983-1984 (financial loss, evacuation, indices of disruption of social networks) and coronary heart disease risk factors (heart rate, blood pressure, total serum cholesterol) assessed in 1987 among 693 Italian male factory workers. Multivariate analyses (adjusting for age, body mass index, smoking, and educational level) revealed no long term relations between the quake related experiences and blood pressure or cholesterol level. However, higher resting heart rates were observed for individuals who reported financial loss, increased distance from family/friends, or decreased visiting as a result of relocation after the quakes. Findings were unchanged after further adjustment for self-reported psychological distress (assessed using the global symptom index of the Symptom Checklist). These findings, while limited by the cross-sectional nature of the data, suggest that a number of psychosocial consequences of relocation due to a natural disaster are unrelated in the long term to coronary heart disease risk factors, except for small but significant differences in heart rate among individuals who have experienced financial loss and/or social network disruptions. PMID- 10873134 TI - Residential radon gas exposure and lung cancer: the Iowa Radon Lung Cancer Study. AB - Exposure to high concentrations of radon progeny (radon) produces lung cancer in both underground miners and experimentally exposed laboratory animals. To determine the risk posed by residential radon exposure, the authors performed a population-based, case-control epidemiologic study in Iowa from 1993 to 1997. Subjects were female Iowa residents who had occupied their current home for at least 20 years. A total of 413 lung cancer cases and 614 age-frequency-matched controls were included in the final analysis. Excess odds were calculated per 11 working-level months for exposures that occurred 5-19 years (WLM(5-19)) prior to diagnosis for cases or prior to time of interview for controls. Eleven WLM(5-19) is approximately equal to an average residential radon exposure of 4 pCl/liter (148 Bq/m3) during this period. After adjustment for age, smoking, and education, the authors found excess odds of 0.50 (95% confidence interval: 0.004, 1.81) and 0.83 (95% percent confidence interval: 0.11, 3.34) using categorical radon exposure estimates for all cases and for live cases, respectively. Slightly lower excess odds of 0.24 (95 percent confidence interval: -0.05, 0.92) and 0.49 (95 percent confidence interval: 0.03, 1.84) per 11 WLM(5-19) were noted for continuous radon exposure estimates for all subjects and live subjects only. The observed risk estimates suggest that cumulative ambient radon exposure presents an important environmental health hazard. PMID- 10873135 TI - Exposure to electromagnetic fields from use of electric blankets and other in home electrical appliances and breast cancer risk. AB - Exposure to electromagnetic fields (EMFs) from use of electric blankets and other in-home electrical appliances has been hypothesized to increase breast cancer risk. To test the hypothesis, the authors analyzed data from a case-control study of female breast cancer conducted in Connecticut in 1994-1997. A total of 608 incident breast cancer patients and 609 age frequency-matched controls, 31-85 years old, were interviewed by trained study interviewers using a standardized, structured questionnaire to obtain information on lifetime use of various in-home electrical appliances. A total of 40% of the cases and 43% of the controls reported regular use of electric blankets in their lifetime, which gave an adjusted odds ratio of 0.9 (95% confidence interval (CI): 0.7, 1.1). For those who reported using electric blankets continuously throughout the night, the adjusted odds ratio was 0.9 (95% CI: 0.7, 1.2) when compared with never users. The risk did not vary according to age at first use, duration of use, or menopausal and estrogen receptor status. The authors also did not find an association between use of other major in-home electrical appliances and breast cancer risk. In conclusion, exposure to EMFs from in-home electrical appliance use was not found to increase breast cancer risk in this study. PMID- 10873136 TI - Relation of adult height to cause-specific and total mortality: a prospective follow-up study of 31,199 middle-aged men and women in Finland. AB - The purpose of this study was to analyze the association of adult height with cause-specific and total mortality. The study included 31,199 men and women aged 25-64 years who participated in a risk factor survey in 1972, 1977, 1982, or 1987 in eastern Finland. The cohorts were followed until the end of 1994. The relation between height and mortality was assessed by using Cox proportional hazard models. The authors found that height was associated inversely with most of the measured risk factors and directly with socioeconomic status. For both genders, height was inversely associated with cardiovascular and total mortality; the age- and birth-cohort-adjusted risk ratios per 5 cm increase in height were 0.89 and 0.91 for men and 0.86 and 0.90 for women, respectively. The inverse association also remained after adjustment for the other known risk factors. For men, an independent inverse association also was found between height and mortality from chronic obstructive pulmonary disease and from violence and accidents. Cancer mortality was not associated with height. Thus, genetic factors, and environmental factors during the fetal period, childhood, and adolescence, which determine adult height, appear to be related to a person's health later in life. PMID- 10873137 TI - Familial aggregation of environmental risk factors and familial aggregation of disease. AB - Almost all human diseases have been shown to aggregate familially to some degree. This familiality is generally taken as evidence for the existence of a genetic etiologic mechanism or environmental factors common to family members, or a combination of both. It has been argued that for a disease with strong familial aggregation, environmental risk factors alone are unlikely to account for such strong aggregation, unless the presumed environmental risk factors are associated with enormous risk. This paper revisits this issue through the use of a novel statistical model. Ascertainment bias aside, the author demonstrates that familial aggregation could be explained by multiple interactive risk factors, each of which may confer a low disease risk and thus contribute only a minuscule portion to disease familiality. For example, two correlated risk factors (r=0.5), each with a relative risk of 5 and acting multiplicatively, could give rise to a sibling relative risk of 1.96. Therefore, it may not be sufficient to argue for a genetic component for a disease based solely on the notion that no high risk environmental factors have been found. In view of this, there is a need to examine carefully the roles of multiple environmental risk factors in disease familiality. PMID- 10873138 TI - Re: "Neighborhood social environment and risk of death: multilevel evidence from the Alameda County study". PMID- 10873139 TI - Re: "Sex ratios, family size, and birth order". PMID- 10873140 TI - Nuremberg forgotten. The deplorable origins of informed consent. PMID- 10873141 TI - Bluish lesions of the anterior maxillary vestibule. PMID- 10873142 TI - The history of dentistry. Could it be a cure for dental phobias? PMID- 10873143 TI - The need for cultural competency in dentistry. PMID- 10873144 TI - Tobacco-induced oral lesions. PMID- 10873145 TI - Perennial goals for the millennium. PMID- 10873146 TI - Fetal airway smooth-muscle contractility and lung development. A player in the band or just someone in the audience? PMID- 10873147 TI - A cytokine reborn? Endothelin-1 in pulmonary inflammation and fibrosis. PMID- 10873148 TI - Spontaneous peristaltic airway contractions propel lung liquid through the bronchial tree of intact and fetal lung explants. AB - Spontaneous contractions of the fetal airways are a well recognized but poorly characterized phenomenon. In the present study spontaneous narrowing of the airways was analyzed in freshly isolated lungs from early to late gestation in fetal pigs and rabbits and in cultured fetal mouse lungs. Propagating waves of contraction traveling proximal to distal were observed in fresh lungs throughout gestation which displaced the lung liquid along the lumen. In the pseudoglandular and canalicular stages (fetal pigs) the frequency ranged from 2.3 to 3.3 contractions/min with a 39 to 46% maximum reduction of lumen diameter. In the saccular stage (rabbit) the frequency was 10 to 12/min with a narrowing of approximately 30%. In the organ cultures the waves of narrowing started at the trachea in whole lungs, or at the main bronchus in lobes (5.2 +/- 1.5 contractions/min, 22 +/- 8% reduction of lumen diameter), and as they proceeded distally along the epithelial tubes the luminal liquid was shifted toward the terminal tubules, which expanded the endbuds. As the tubules relaxed the flow of liquid was reversed. Thus the behavior of airway smooth muscle in the fetal lung is phasic in type (like gastrointestinal muscle) in contrast to that in postnatal lung, where it is tonic. An intraluminal positive pressure of 2.33 +/- 0.77 cm H(2)O was recorded in rabbit fetal trachea. It is proposed that the active tone of the smooth muscle maintains the positive intraluminal pressure and acts as a stimulus to lung growth via the force exerted across the airway wall and adjacent parenchyma. The expansion of the compliant endbuds by the fluid shifts at the airway tip may promote their growth into the surrounding mesenchyme. PMID- 10873149 TI - Pulmonary fibrosis and chronic lung inflammation in ET-1 transgenic mice. AB - The pulmonary endothelin (ET) system has been implicated in the pathogenesis of chronic lung diseases such as pulmonary hypertension, asthma, chronic obstructive lung disease, idiopathic pulmonary fibrosis, and bronchiolitis obliterans. However, the etiologic role of ET-1 in these diseases has not yet been established. We recently demonstrated that ET-1 transgenic mice, generated using the human prepro-ET-1 expression cassette including the cis-acting transcriptional regulatory elements, had predominant transgene expression in lung, brain, and kidney. We used these mice in the present study to analyze the pathophysiologic consequences of long-term pulmonary overexpression of ET-1. We found that ET-1 overexpression in the lungs did not result in significant pulmonary hypertension, but did result in development of a progressive pulmonary fibrosis and recruitment of inflammatory cells (predominantly CD4-positive cells). Our study provides evidence that a long-term activated pulmonary ET system, without any other stimuli, produces chronic lymphocytic inflammation and lung fibrosis. This suggests that overexpression of ET-1 may be a central event in the pathogenesis of lung diseases associated with fibrosis and chronic inflammation, such as pulmonary fibrosis and bronchiolitis. PMID- 10873150 TI - Heterogeneity of clara cell glutathione. A possible basis for differences in cellular responses to pulmonary cytotoxicants. AB - Clara-cell populations show a high degree of variation in susceptibility to injury by bioactivated cytotoxicants. Because glutathione (GSH) is critical for detoxification of electrophilic metabolites, heterogeneity in Clara cell GSH levels may lead to a wide range of cytotoxic responses. This study was designed to define the distinct GSH pools within Clara cells, characterize heterogeneity within the population, and examine whether heterogeneity contributes to susceptibility. Using fluorescent imaging combined with high-performance liquid chromatography analysis, semiquantitative measurements were obtained by evaluation of GSH using monochlorobimane and monobromobimane. In steady-state conditions, the GSH measured in isolated cells was in the femtomole range, but varied 4-fold between individual cells. Clara cells analyzed in situ and in vitro confirmed this heterogeneity. The response of these cells to compounds that modulate GSH was also variable. Diethylmaleate depleted GSH, whereas GSH monoethylester augmented it. However, both acted nonuniformly in isolated Clara cells. The depletion of intracellular GSH caused a striking decrease in cell viability upon incubation with naphthalene (NA). The sulfhydryl-binding fluorochrome BODIPY, which colocalized with tetramethylrosamine, a mitochondrial dye, demonstrated by confocal microscopy that cellular sulfhydryls are highest in the mitochondria, next-highest in cytoplasm, and lowest in the nucleus. These pools responded differently to modulators of GSH. We concluded that the steady state intracellular GSH of Clara cells exists in distinct pools and is highly heterogeneous within the population, and that the heterogeneity of GSH levels corresponds closely to the response of Clara cells to injury by NA. PMID- 10873151 TI - Cell-specific expression of group X and group V secretory phospholipases A(2) in human lung airway epithelial cells. AB - Secretory phospholipase A(2) (sPLA(2)) enzymes contribute to inflammatory injury in human lungs by several mechanisms, including eicosanoid production and hydrolytic damage to surfactant phospholipids. Several distinct sPLA(2) genes have been described in human tissue but little is known regarding their presence, localization, or function(s) within lungs. We hypothesized that sPLA(2)s would have cell-specific distributions within lung. We used reverse transcriptase/polymerase chain reaction to identify sPLA(2) messenger RNAs (mRNAs) in adult human lung tissue. Resulting complementary DNA (cDNA) sequences indicated that total lung extracts contained mRNA for Groups IB, IIA, V, and X sPLA(2). An epithelial cell line, BEAS cells, expressed only Groups IIA, V, and X. We used these cDNAs to clone these enzymes, especially the recently described Group X and Group V enzymes. Digoxigenin-labeled complementary RNA probes were used to determine localization of each sPLA(2) by in situ hybridization of human lung. Hybridization was strongly positive for Group X and Group V in airway epithelial cells, which failed to hybridize Group IB or IIA probes. Although four known mammalian sPLA(2) isotypes were expressed in lung, only Group X and Group V sPLA(2) mRNAs appear uniquely expressed in airway epithelium, suggesting they could provide a mechanism of pulmonary surfactant hydrolysis during lung injury. PMID- 10873152 TI - Ciliogenesis and left-right axis defects in forkhead factor HFH-4-null mice. AB - Cilia have been classified as sensory or motile types on the basis of functional and structural characteristics; however, factors important for regulation of assembly of different cilia types are not well understood. Hepatocyte nuclear factor-3/forkhead homologue 4 (HFH-4) is a winged helix/forkhead transcription factor expressed in ciliated cells of the respiratory tract, oviduct, and ependyma in late development through adulthood. Targeted deletion of the Hfh4 gene resulted in defective ciliogenesis in airway epithelial cells and randomized left-right asymmetry so that half the mice had situs inversus. In HFH-4-null mice, classic motile type cilia with a 9 + 2 microtubule ultrastructure were absent in epithelial cells, including those in the airways. In other organs, sensory cilia with a 9 + 0 microtubule pattern, such as those on olfactory neuroepithelial cells, were present. Ultrastructural analysis of mutant cells with absent 9 + 2 cilia demonstrated that defective ciliogenesis was due to abnormal centriole migration and/or apical membrane docking, suggesting that HFH 4 functions to direct basal body positioning or anchoring. Evaluation of wild type embryos at gestational days 7.0 to 7.5 revealed Hfh4 expression in embryonic node cells that have monocilium, consistent with a function for this factor at the node in early determination of left- right axis. Analysis of the node of HFH 4 mutant embryos revealed that, in contrast to absent airway cilia, node cilia were present. These observations indicate that there are independent regulatory pathways for node ciliogenesis compared with 9 + 2 type ciliogenesis in airways, and support a central role for HFH-4 in ciliogenesis and left-right axis formation. PMID- 10873153 TI - Intraepithelial vagal sensory nerve terminals in rat pulmonary neuroepithelial bodies express P2X(3) receptors. AB - The neurotransmitters/modulators involved in the interaction between pulmonary neuroepithelial bodies (NEBs) and the vagal sensory component of their innervation have not yet been elucidated. Because P2X(3) purinoreceptors are known to be strongly expressed in peripheral sensory neurons, the aim of the present study was to examine the localization of nerve endings expressing P2X(3) purinoreceptors in the rat lung in general and those contacting pulmonary NEBs in particular. Most striking were intraepithelial arborizations of P2X(3) purinoceptor-immunoreactive (IR) nerve terminals, which in all cases appeared to ramify between calcitonin gene-related peptide (CGRP)- or calbindin D28k (CB) labeled NEB cells. However, not all NEBs received nerve endings expressing P2X(3) receptors. Using CGRP and CB staining as markers for two different sensory components of the innervation of NEBs, it was revealed that P2X(3) receptor and CB immunoreactivity were colocalized, whereas CGRP-IR fibers clearly formed a different population. The disappearance of characteristic P2X(3) receptor positive nerve fibers in contact with NEBs after infranodosal vagal crush and colocalization of tracer and P2X(3) receptor immunoreactivity in vagal nodose neuronal cell bodies in retrograde tracing experiments further supports our hypothesis that the P2X(3) receptor-IR nerve fibers contacting NEBs have their origin in the vagal sensory nodose ganglia. Combination of quinacrine accumulation in NEBs, suggestive of the presence of high concentrations of adenosine triphosphate (ATP) in their secretory vesicles, and P2X(3) receptor staining showed that the branching intraepithelial P2X(3) receptor-IR nerve terminals in rat lungs were exclusively associated with quinacrine-stained NEBs. We conclude that ATP might act as a neurotransmitter/neuromodulator in the vagal sensory innervation of NEBs via a P2X(3) receptor-mediated pathway. Further studies are necessary to determine whether the P2X(3) receptor-expressing neurons, specifically innervating NEBs in the rat lung, belong to a population of P2X(3) receptor-IR nociceptive vagal nodose neurons. PMID- 10873154 TI - Type V collagen modulates alloantigen-induced pathology and immunology in the lung. AB - Perivascular and peribronchiolar tissues are targets of the immune response during lung allograft rejection. Collagen type V (col[V]) is located within these tissues. Col(V) may be major histocompatibility complex (MHC)-like, and MHC derived peptides have been used to induce immunologic tolerance and prevent rejection in allografts other than the lung. The current study tests the hypothesis that col(V) could be used to downregulate immune responses to lung alloantigen in vivo. We developed a murine model in which instillations of allogeneic bronchoalveolar lavage (BAL) cells (C57BL/6, I-a(b), H-2(b)) into lungs of BALB/c mice (I-a(d), H-2(d)) induce histology similar to grades 1 and 2 acute lung allograft rejection, apoptosis of airway epithelium and vascular endothelium, and upregulate tumor necrosis factor (TNF)-alpha production locally. The current study reports that instillations of col(V) into lungs before allogeneic BAL cells prevent development of rejection pathology and apoptosis, downregulate alloantigen-induced T-lymphocyte proliferation, and abrogate local TNF-alpha production. In addition, instillation of col(V)-pulsed autologous BAL cells into lungs of mice primed with allogeneic BAL cells perpetuates rejection pathology. Collectively, these data show that col(V) is a novel antigen involved in the rejection process, and suggest that col(V) could be used to modulate the rejection response to lung allografts. PMID- 10873155 TI - Surfactant protein A binding to cytomegalovirus proteins enhances virus entry into rat lung cells. AB - The role of surfactant protein (SP)-A in cytomegalovirus (CMV) infection of the lung was investigated. We found that SP-A binds to various immobilized human CMV proteins and those exposed on the surface of infected embryonal lung fibroblasts. The interaction between SP-A and immobilized CMV proteins was found to be calcium dependent and inhibited by mannan, suggesting involvement of the carbohydrate recognition domain of SP-A and high-mannose carbohydrate residues of viral envelope glycoproteins. Using flow cytometry and confocal laser fluorescence microscopy in the rat model we showed that preincubation of rat CMV with SP-A stimulates its binding and internalization by rat type II pneumocytes and alveolar tissue macrophages. This effect was concentration- and Ca(2+)-dependent but was not inhibited by mannan. Therefore, the domains of SP-A involved in SP-A CMV interaction and in interaction of the SP-A/virus complex with rat lung cells are distinct. Additionally, in the human CMV model, sheep as well as human proteinosis SP-A did not significantly affect human CMV replication in embryonal lung fibroblasts. Thus, SP-A may contribute to CMV-associated pathology of the lung by increasing the efficiency of target cell infection. PMID- 10873156 TI - Synergistic inhibition by beta(2)-agonists and corticosteroids on tumor necrosis factor-alpha-induced interleukin-8 release from cultured human airway smooth muscle cells. AB - We have previously reported that human airway smooth-muscle (ASM) cells produce abundant interleukin (IL)-8, a major neutrophil chemoattractant involved in asthma exacerbations. Here, we tested the effects of the beta(2)-agonists salbutamol (Salbu) and salmeterol (Salme) on IL-8 release and tumor necrosis factor (TNF)-alpha-induced IL-8 release from ASM cells. We found that TNF-alpha strongly enhanced IL-8 release in a time- and concentration-dependent manner, whereas Salbu, Salme, the direct adenylyl cyclase activator forskolin (FSK), and the cyclic monophosphate (cAMP) analogue 8-bromoadenosine 3',5'-cAMP (8-Br-cAMP) alone weakly stimulated IL-8 release. TNF-alpha (10 ng/ml)-induced IL-8 release was markedly inhibited by the steroids dexamethasone (Dex) (0.1 to 10 microM) and fluticasone (Flut) (0.01 to 1 microM) but unaffected by Salbu, Salme, FSK, or 8 Br-cAMP. However, a combination of Dex (1 microM) or Flut (0.1 microM) with Salbu (10 microM), Salme (1 microM), FSK (10 microM), or 8-Br-cAMP (10 and 100 microM) significantly enhanced the inhibition by Dex or Flut alone. Experiments with KT5720, a selective inhibitor of cAMP-dependent protein kinase A; rolipram, a selective inhibitor of type IV phosphodiesterase; and ICI-118,551, a beta(2) receptor antagonist, suggested that the synergistic inhibition was mediated by beta(2)-receptor in a cAMP-dependent manner. This novel synergistic interaction of beta(2)-agonists and steroids may partly explain the benefits that result when these agents are combined to treat asthma. PMID- 10873157 TI - Mitogen-activated protein kinases regulate cytokine gene expression in human airway myocytes. AB - The signal transduction pathways regulating smooth-muscle gene expression and production of cytokines in response to proinflammatory mediators are undefined. Cultured human bronchial smooth-muscle cells were treated for 20 h with a cytokine cocktail containing interleukin (IL)-1beta, tumor necrosis factor-alpha, and interferon-gamma. A complementary DNA expression array containing 588 genes was used to follow cytokine-stimulated gene expression. The expression and secretion of the cytokines IL-1beta, IL-6, and IL-8 significantly increased after 20 h of stimulation as measured by relative reverse transcriptase/ polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blotting techniques. Expression of IL-6 and IL-8 was sensitive to SB203580, the specific inhibitor of p38 mitogen-activated protein (MAP) kinase and PD98059, an inhibitor of MAP kinase kinase. Expression of IL-1beta was sensitive only to PD98059. Together, these results demonstrate that the p38 and extracellular signal regulated protein kinase MAP kinase pathways are required for proinflammatory mediator- induced cytokine expression in airway myocytes. The generation of chemokines and cytokines in airway smooth muscle also provides evidence that smooth-muscle cells have the ability to contribute to the inflammatory response. PMID- 10873159 TI - Tumor necrosis factor-alpha enhances mRNA expression and secretion of interleukin 6 in cultured human airway smooth muscle cells. AB - Airway smooth muscle (ASM) is considered to be an end-target cell for the effects of mediators released during airway wall inflammation. Several reports suggest that activated ASM may be capable of generating various proinflammatory cytokines. We investigated the effects of tumor necrosis factor (TNF)-alpha, a potent proinflammatory cytokine, on cultured human ASM cells by examining the expression and release of the cytokine interleukin (IL)-6, cell proliferation, and the expression pattern of c-fos and c-jun, two nuclear proto-oncogenes constituting the activator protein-1 transcription factor. Growth-arrested cell monolayers were stimulated with human recombinant TNF-alpha in a concentration- and time-dependent manner. TNF-alpha stimulated the expression of IL-6 messenger RNA (mRNA), which was detected after 15 min, reaching a maximum at 1 h. IL-6 protein was readily detected in ASM cell-conditioned medium after 2 h of TNF alpha stimulation. Protein levels increased in a time- and concentration dependent manner. Release of IL-6 elicited by TNF-alpha was significantly inhibited by dexamethasone, cycloheximide, and nordihydroguaiaretic acid (NDGA). TNF-alpha did not alter DNA biosynthesis up to 48 h or cell numbers up to 120 h. Northern blot analysis of proto-oncogene expression revealed that c-fos and c-jun mRNA levels were elevated after 30 min of TNF-alpha incubation with maximum levels at 1 h and 45 min, respectively. Expression of c-fos mRNA was downregulated by NDGA. Four hours of TNF-alpha treatment resulted in translocation of c-jun immunofluorescence from the cytoplasm to the nucleus in human ASM cells. Our results suggest that despite the lack of a mitogenic response to TNF-alpha, upregulation of primary response genes in human ASM cells may account for the induction of proinflammatory cytokines, such as IL-6, in human airways. PMID- 10873158 TI - Vanadium-induced kappaB-dependent transcription depends upon peroxide-induced activation of the p38 mitogen-activated protein kinase. AB - Activation of nuclear factor (NF)-kappaB and subsequent proinflammatory gene expression in human airway epithelial cells can be evoked by oxidative stress. In this study we examined signal transduction pathways activated by vanadyl sulfate (V(IV))-induced oxidative stress in normal human bronchial epithelial cells. Both nuclear translocation of NF-kappaB and enhanced kappaB-dependent transcription induced by V(IV) were inhibited by overexpression of catalase, but not Cu,Zn superoxide dismutase (Cu,Zn-SOD), indicating that peroxides rather than superoxides initiated signaling. Catalase selectively blocked the response to V(IV) because it inhibited neither NF-kappaB translocation nor kappaB-dependent transcription evoked by the proinflammatory cytokine tumor necrosis factor (TNF) alpha. The V(IV)-induced kappaB-dependent transcription was dependent upon activation of the p38 mitogen-activated protein kinase because overexpression of dominant-negative mutants of the p38 MAPK pathway inhibited V(IV)-induced kappaB dependent transcription. This inhibition was not due to suppression of NF-kappaB nuclear translocation because NF-kappaB DNA binding was unaffected by the inhibition of p38 activity. Overexpression of catalase, but not Cu,Zn-SOD, inhibited p38 activation, indicating that peroxides activated p38. Catalase failed to block V(IV)- induced increases in phosphotyrosine levels, suggesting that the catalase-sensitive signaling components were independent of V(IV) induced tyrosine phosphorylation. The data demonstrate that V(IV)-induced oxidative stress activates at least two distinct pathways, NF-kappaB nuclear translocation and p38-dependent transactivation of NF-kappaB, both of which are required to fully activate kappaB-dependent transcription. Moreover, V(IV) induced oxidative stress activated these pathways in bronchial epithelial cells by upstream signaling cascades that were distinct at some level from those used by the proinflammatory cytokine TNF-alpha. PMID- 10873160 TI - Iron is a regulatory component of human IL-1beta production. Support for regional variability in the lung. AB - The human lung accumulates iron with senescence. Smoking escalates the accumulation of iron, and we have demonstrated regional variability in the accumulation of iron in smokers' lungs. Iron has been reported to influence the production of a number of proinflammatory mediators, including human interleukin (IL)-1beta. We postulated that we could (1) demonstrate regional differences in the release of IL-1beta from human alveolar macrophages and (2) influence the production of IL-1beta in human macrophages by altering intracellular iron concentrations. To test these hypotheses, alveolar macrophages were obtained by independent lavage of the upper and lower lobes of healthy volunteers (both smokers and nonsmokers), after which the ability of each population to secrete IL 1beta was quantified, together with their ability to produce tumor necrosis factor-alpha, IL-6, and IL-8. Additionally, we established an in vitro model of "iron-loaded" cells of the human myelomonocytic cell line THP-1 in order to examine more directly the effect of iron and its chelation on the secretion of IL 1beta. We report here that an intracellular, chelatable pool of iron expands with exogenous iron-loading as well as with lipopolysaccharide (LPS) stimulation and appears to suppress transcription of IL-1beta, whereas shrinkage of this pool by early chelation augments transcription of IL-1beta beyond that induced by LPS alone. And finally, we demonstrate a regional relationship in the lung between excess alveolar iron and the production of human alveolar macrophage-derived IL 1beta, suggesting a partnership between iron and inflammation that may have clinical significance, especially in relation to lung diseases with a regional predominance. PMID- 10873161 TI - Early motor and mental development in very preterm infants with chronic lung disease. AB - BACKGROUND: The increased incidence of neurological deviations in preterm infants with chronic lung disease (CLD) has been linked to severe brain haemorrhage (intraventricular haemorrhage (IVH)) and periventricular leucomalacia (PVL) rather than to CLD per se. AIM: To evaluate whether CLD without concomitant brain lesions constitutes a risk factor for adverse developmental outcome. METHOD: Forty three very low birthweight infants with CLD, but without IVH or PVL, and 43 very low birthweight infants without CLD, IVH, or PVL were evaluated at 5 and 10 months of corrected age using the movement assessment of infants (MAI) scale. The Griffiths' developmental test was carried out at 10 months of age. RESULTS: The overall motor assessments (MAI) in infants with CLD and controls were not significantly different. However, differences were observed in the execution of volitional movements (MAI), the total sum, hand and eye coordination, and perception and intelligence (measured by the performance scale of the Griffiths' test). CONCLUSIONS: CLD has a deleterious effect on the control of hand and eye coordination and on perception and intelligence. These results thus re-emphasise the necessity for careful neurodevelopmental follow up of infants with CLD whether or not they suffered IVH or PVL. PMID- 10873163 TI - Midwivery PMID- 10873162 TI - Increasing rates of cerebral palsy across the severity spectrum in north-east England 1964-1993. The North of England Collaborative Cerebral Palsy Survey. AB - OBJECTIVES: To report epidemiological trends in cerebral palsy including analyses by severity. DESIGN: Descriptive longitudinal study in north-east England. Every child with suspected cerebral palsy was examined by a developmental paediatrician to confirm the diagnosis. Severity of impact of disability was derived from a parent completed questionnaire already developed and validated for this purpose. SUBJECTS: All children with cerebral palsy, not associated with any known postneonatal insult, born 1964-1993 to mothers resident at the time of birth in the study area. MAIN OUTCOME MEASURES: Cerebral palsy rates by year, birth weight, and severity. Severity of 30% and above defines the more reliably ascertained cases; children who died before assessment at around 6 years of age are included in the most severe group (70% and above). RESULTS: 584 cases of cerebral palsy were ascertained, yielding a rate that rose from 1.68 per 1000 neonatal survivors during 1964-1968 to 2.45 during 1989-1993 (rise = 0.77; 95% confidence interval 0.2-1.3). For the more reliably ascertained cases there was a twofold increase in rate from 0.98 to 1.96 (rise = 0.98; 95% confidence interval 0.5-1.4). By birth weight, increases in rates were from 29.8 to 74.2 per 1000 neonatal survivors < 1500 g and from 3.9 to 11.5 for those 1500-2499 g. Newborns < 2500 g now contribute one half of all cases of cerebral palsy and just over half of the most severe cases, whereas in the first decade of this study they contributed one third of all cases and only one sixth of the most severe (chi(2) and chi(2) for trend p < 0.001). CONCLUSIONS: The rate of cerebral palsy has risen in spite of falling perinatal and neonatal mortality rates, a rise that is even more pronounced when the mildest and least reliably ascertained are excluded. The effect of modern care seems to be that many babies < 2500 g who would have died in the perinatal period now survive with severe cerebral palsy. A global measure of severity should be included in registers of cerebral palsy to determine a minimum threshold for international comparisons of rates, and to monitor changes in the distribution of severity. PMID- 10873164 TI - Increased leptin concentration in preterm infants of pre-eclamptic mothers. AB - AIM: To study the effect of maternal pre-eclampsia on cord plasma leptin concentrations in preterm infants. METHODS: Leptin concentration was analysed in cord plasma of 74 preterm infants, gestational age 24 to 32 weeks. Of these, 14 were born to pre-eclamptic mothers, in 10 intrauterine growth retardation (IUGR) was present, and 59 had been exposed antenatally to corticosteroids. RESULTS: The mean (SD) concentration of cord plasma leptin was 1.31 (0.88) microg/l. A significant correlation was found between leptin concentration and gestational age (r = 0.336; p = 0.0037). Leptin levels were higher in infants of pre eclamptic mothers (p = 0.0007), in those with IUGR (p = 0.0005), and in infants exposed antenatally to corticosteroids (p = 0.02). In multiple regression analysis, leptin was associated with gestational age and maternal pre-eclampsia (both p < 0.05), but not with antenatal corticosteroids. CONCLUSIONS: Increased fetal leptin in maternal pre-eclampsia may reflect a physiological adaptation to fetal stress such as hypoxia. PMID- 10873165 TI - Markers of collagen metabolism and insulin-like growth factor binding protein-1 in term infants. AB - AIM: To study the relation between fetal growth and markers of collagen metabolism and insulin-like growth factor binding protein-1 (IGFBP-1) in term infants. METHODS: Cord vein plasma was obtained from 67 term infants of gestational age 37.1-41.7 weeks (39 appropriate for gestational age (AGA), 11 large for gestational age (LGA; relative birth weight >/= 2.0 SD), and 17 small for gestational age (SGA; relative birth weight 0.05). CONCLUSIONS: In the term fetus, collagen metabolism is primarily dependent on maturity and not on intrauterine growth status, whereas IGFBP-1 reflects intrauterine growth independently of maturity. PMID- 10873166 TI - Malignant infantile osteopetrosis presenting with neonatal hypocalcaemia. AB - Presentation characteristics were reviewed in 14 children from 12 families with malignant infantile osteopetrosis seen at two large referral centres for bone marrow transplantation. Children from six of these families presented initially with symptoms of hypocalcaemia. These comprised early or late neonatal convulsions in six cases (corrected serum calcium < 1.5 mmol/l), and vomiting and irritability (serum calcium 1.68 mmol/l) in another. One other related child had severe and persistent jittering episodes almost certainly attributable to hypocalcaemia. In seven of eight cases, these symptoms developed during the first 14 days of life. Although occasionally reported previously, malignant infantile osteopetrosis remains essentially unrecognised as a cause of neonatal hypocalcaemia, often resulting in diagnostic confusion and delay. This is important in the context of curative haemopoietic stem cell transplantation where preservation of sight may depend on early intervention. PMID- 10873167 TI - Controlled trial of immune response of preterm infants to recombinant hepatitis B and inactivated poliovirus vaccines administered simultaneously shortly after birth. AB - AIM: The study was conducted to evaluate the immunogenicity of an early, extra dose of enhanced inactivated poliovirus vaccine (IPV) administered simultaneously with recombinant hepatitis B vaccine (HBV) to preterm infants shortly after birth. METHODS: Three groups were studied. Fifty preterm infants received IPV intramuscularly within 24 hours of birth, in addition to routine recommended childhood immunisations. Fifty two preterm infants and 35 full term infants received routine immunisations only (routine vaccination timing: HBV at birth, 1 and 6 months of age; IPV at 2 and 4 months; oral polio vaccine (OPV) at 4 and 6 months; diphtheria-tetanus-pertussis (DTP) at 2, 4, and 6 months; and Haemophilus influenzae B vaccine at 2 and 4 months). Blood samples were taken at birth, 3 and 7 months of age from all infants, and at 1 month of age from preterm infants only. RESULTS: At birth, a lower percentage of both study and control preterm infants had antipoliovirus type 3 titres >/= 1:8 than full term infants. At 1 and 3 months of age significantly more early IPV infants had antipoliovirus type 3 titres >/= 1:8 than routinely vaccinated preterm infants (p < 0.05). At 7 months of age there were no significant differences in percentage of antipoliovirus titres >/= 1:8 or geometric mean times (GMTs) between the early IPV group and the routinely vaccinated preterm group. At 3 and 7 months of age, the percentage of positive antihepatitis B titres (>/= 1:10) and the GMT of the early IPV preterm group did not differ significantly from those of preterm controls. There was no significant difference in percentage of positive antihepatitis B titres between the early IPV group and full term controls at any time. GMTs for hepatitis B antibodies were significantly lower in the early IPV preterm group than in full term controls at 3 and 7 months of age. CONCLUSIONS: Administration of an additional dose of IPV simultaneously with routine HBV to preterm infants shortly after birth provides early protection from poliovirus and hepatitis B infection, and does not interfere with poliovirus antibody production at the age of 7 months. PMID- 10873168 TI - Effects of glucagon on in vitro liquid production by lungs from fetal guinea pigs. AB - BACKGROUND: Lung liquid reabsorption in newborns with respiratory distress syndrome can be deficient. Respiratory distress syndrome is often seen in infants of diabetic mothers, in whom the neonatal surge of glucagon is suppressed. AIM: To investigate the possible effects of glucagon on lung liquid reabsorption. METHODS: Lungs from near term fetal guinea pigs (62 (2) days gestation; term = 67 days) were supported in vitro for three hours; lung liquid production and reabsorption were monitored by a dye dilution method. RESULTS: Untreated control preparations produced fluid at 1.75 (0.33) ml/h per kg body weight, and did not change significantly in three hours; those immersed in 10(-12) M glucagon during the middle hour showed no significant change, but those given higher concentrations all showed significant reductions in fluid production or even reabsorption (65.6 (10.3)% fall at 10(-11) M, 70.0 (6.3)% fall at 10(-10) M, and 90.6 (11.1)% fall at 10(-9) M; based on 54 preparations). At 10(-9) M glucagon, 12 out of 30 preparations reabsorbed fluid. The linear log dose-response curve (r(2) = 0.94) gave a theoretical threshold at 4 x 10(-15) M glucagon. Responses appeared to involve the amiloride sensitive Na(+) based reabsorptive system: responses to 10(-9) M glucagon appeared to be reduced by 10(-6) M amiloride, and were abolished by 10(-5) M amiloride (based on 72 preparations). CONCLUSIONS: The results suggest that the surge of glucagon at birth may help to drain the lungs of fluid. As glucagon liberates cAMP, which also stimulates surfactant, glucagon is worth consideration for possible use in neonatal respiratory distress. PMID- 10873169 TI - Oxygen saturation during the first 24 hours of life. AB - AIM: To determine normative data for arterial oxygen saturation, measured by pulse oximetry (SpO2), in healthy full term infants throughout their first 24 hours of life. METHODS: Long term recordings of SpO2, pulse waveform, and breathing movements were made on 90 infants. Recordings were analysed for baseline SpO(2), episodes of desaturation (SpO2 /= four seconds, and periodic apnoea (>/= three apnoeic pauses, each separated by /= 20 seconds) were identified in six recordings. Four desaturations fell to 15 g. There was no correlation between alcohol consumption and severity of hepatotoxicity (mean INR and the serum creatinine levels over the first 7 days after the overdose). The significantly lower platelet count in heavy drinkers was probably the consequence of direct alcohol toxicity to the marrow. Overall there was a greater incidence of heavy alcohol consumption amongst therapeutic misadventure compared to deliberate overdose cases, but there was no difference between the two groups when amounts of <10 g/day were involved. Eleven (29%) patients in the therapeutic misadventure group were depressed, 10 of whom had previously attempted suicide. In conclusion, we were unable to demonstrate that heavy drinkers develop more severe hepatotoxicity following paracetamol overdose than non-drinkers, and from the material reported in this study, accidental overdose is a better defining term than therapeutic misadventure. PMID- 10873184 TI - Doctors who kill themselves: a study of the methods used for suicide. AB - Medical practitioners have a relatively high rate of suicide. Death entry data for doctors who died by suicide or undetermined cause between 1979 and 1995 in England and Wales were used to compare methods used for suicide by doctors with those used by the general population. Methods used were analysed according to gender, occupational status and speciality, to assess the extent to which access to dangerous means influences the pattern of suicide. Self-poisoning with drugs was more common in the doctors than in general population suicides (57% vs. 26.6%; OR=3.65, 95% CI 2.85-4. 68), including in retired doctors. Barbiturates were the most frequent drugs used. Half of the anaesthetists who died used anaesthetic agents. Self-cutting was also more frequently used as a method of suicide. The finding that the greater proportion of suicide deaths in doctors were by self-poisoning may reflect the fact that doctors have ready access to drugs, and have knowledge of which drugs and doses are likely to cause death. The specific finding that a large proportion of suicides in anaesthetists involved anaesthetic agents supports this explanation. Availability of method may be a factor contributing to the relatively high suicide rate of doctors. This fact might influence clinical management of doctors who are known to be depressed or suicidal. PMID- 10873185 TI - Randomized trial of graded compression stockings for prevention of deep-vein thrombosis after acute stroke. AB - Graded compression stockings are commonly used to prevent deep-vein thrombosis (DVT) after stroke, but their efficacy in this setting has not been evaluated. Extrapolation of effectiveness from trials in patients undergoing elective surgery may be inappropriate. We undertook a randomized, controlled trial, with blinded data review, in a University hospital Acute Stroke Unit. Patients were allocated to graded compression stockings or to standard care alone. DVT incidence was determined at baseline and at day 7+/-2 by colour-flow Doppler ultrasound. Ninety-eight patients with acute, immobilizing stroke were randomized; 97 had full outcome data. One patient had clinically manifest DVT, and no patient had pulmonary thromboembolism. DVT was detected in 7/65 patients allocated stockings, and 7/32 controls (odds ratio 0.43, 95% CI 0.14-1.36); DVT involving femoral veins was detected in 3/65 and 2/32. In the first week after stroke, radiologically-detected DVT remains common, but is usually clinically silent. Proximal DVT is less common. Graded compression stockings produced a reduction in DVT incidence comparable to that in other patient groups, but the reduction was not statistically significant, and the magnitude of effect size requires confirmation. There is greater doubt over efficacy in early prevention of proximal DVT. PMID- 10873186 TI - Primary hyperparathyroidism with normal serum intact parathyroid hormone levels. AB - To evaluate the features of primary hyperparathyroidism (HPT) with normal serum intact parathyroid hormone (iPTH) levels, we studied 271 consecutive patients undergoing surgery for primary HPT. In 20 patients, serum iPTH levels were within the normal range (10-65 ng/l). In their records, the most common clinical features were fatigue (n=13), polyuria (n=6), renal stone (n=5), and hypertension (n=5). Mean serum calcium and phosphorus were 2.78 and 0.85 mmol/l, respectively: 14 had serum phosphorus within the normal range. Mean serum iPTH was 48.5 ng/l, and was <45 ng/l in nine patients. Cervical ultrasound demonstrated a parathyroid adenoma in nine, and was normal in four. Tc sestamibi parathyroid scintigraphy always demonstrated an adenoma (9/9). In eight patients, normal iPTH values delayed diagnosis. Physicians should be aware of the possibility of HPT in patients with hypercalcaemia, even when serum phosphorus and iPTH levels are within the normal limits. Particularly, HPT cannot be excluded when serum iPTH levels are below the upper part of the normal range. In such cases, cervical imaging, which has the same sensitivity as in other HPT, should be undertaken. These explorations are useful, because many patients are symptomatic and can take advantage of surgery. PMID- 10873187 TI - Poor glycaemic control in type 2 diabetes: a conspiracy of disease, suboptimal therapy and attitude. PMID- 10873188 TI - Respiratory infection and coronary heart disease: progression of a paradigm. AB - We have developed a previously published paradigm concerning causation of coronary heart disease, based on the probability that the fundamental cause is a microbe, probably Chlamydia pneumoniae, and that the progress of the disease is influenced by number of accelerating and inhibiting factors. We propose that cigarette smoking acts via respiratory infection, this itself being influenced by immunocompetence resulting from sunlight exposure. We also propose an immuno enhancing effect of oestrogen and an anti-inflammatory effect of statin therapy. In respect of the geographical variation of coronary heart disease, we emphasize that this must be viewed as part of the bigger picture of a high mortality from all causes in countries of North-west Europe that have a particularly low level of sunlight exposure. Finally, we draw attention to the Albanian, French, Italian, Northern Ireland and Scottish paradoxes which should lead to a major review of the conventional wisdom concerning the aetiogenesis of coronary heart disease. PMID- 10873189 TI - Seasonal variation in coronary heart disease and seasonal mood changes. PMID- 10873190 TI - Behcet's syndrome. PMID- 10873191 TI - Cardiovascular risk in women: the cardiologist's perspective. PMID- 10873192 TI - Vegetarian diet. PMID- 10873193 TI - Acute cholecystitis and septic shock due to salmonella virchow. PMID- 10873194 TI - Dual resonant birdcage coils for 1H detected 13C microscopic imaging at 11.7 T. AB - Liquid state, rotating frame cross polarisation experiments are very sensitive to RF field inhomogeneity. In this work, we present an easily fabricated, co resident high- and low-pass linear birdcage resonator, optimised to perform liquid state rotating frame polarisation transfer at 1H and 13C frequencies. Both the RF fields have been experimentally mapped, and used to validate the spatial signal dependence of a proton detected, 13C image. The predicted performance was then confirmed using PRAWN-based, cyclic J-cross polarisation (CYCLCROP) imaging. A novel variant of a B(1)-field mapping approach is also presented, using the signal enhancement of the CYCLCROP sequence to generate proton detected, 13C field maps. PMID- 10873195 TI - Open access birdcage coils for microscopic imaging of plants at 11.7 T. AB - The use of a U-shaped high-pass birdcage coil for microscopic imaging at 11.7 T has been investigated. The study was motivated by the requirement for a side access coil, permitting higher filling factors for the in-vivo imaging of plant petioles and stems. The performance of a U-shaped coil (with a cross section consisting of a 16 mm diameter semi-circle plus two 12 mm length straight sections) has been experimentally assessed, and compared both in terms of homogeneity and sensitivity to a 16 mm diameter conventional (linear) birdcage and a saddle coil of the same diameter. The U-shaped coil, which offers 12 mm width side access, has a significantly better performance than the saddle coil, whilst providing 57% of the B(1) sensitivity of the bird-cage. PMID- 10873196 TI - An efficient geometric image distortion correction method for a biplanar planar gradient coil. AB - Since the spatial field non-linearity of gradient coils translates into image geometric distortion in MRI, in many applications, such as cardiac function analysis and interventional MR-based device tracking/guidance, where the precise geometric information is needed, the presence of geometric image distortion can not be simply ignored. To address the concern for geometric image distortion, we have developed and validated a general and efficient numerical technique for parameterizing the global image distortion for a bi-planar gradient coil as well as accomplishing image restoration as a post-imaging processing. This image correction methodology is based on a global distortion coordinate mapping function which can be systematically defined directly from the gradient field non linearity in 3-dimension (3D) of a given gradient coil. The image correction was carried out in two steps: (1) map each pixel of the corrected image representation onto its distorted image according to the distortion mapping; (2) interpolate the pixel intensity in the distorted image using its neighboring points via a bi-linear interpolation procedure. The results showed clearly that the distortion correction method was robust in term of the capability of reducing image geometric distortion dramatically. Also it is shown that the magnetic field non-linearity or the image distortion of a typical bi-planar gradient coil can be adequately parameterized using a finite Taylor series expansion based on its design parameters. Furthermore, this image distortion correction method is very efficient in practice for performing 3D correction for any image orientation since a compact parameterized field expression contains non-zero terms. PMID- 10873197 TI - Determination of appropriate RF blocking impedance for MRI surface coils and arrays. AB - Surface and phased array receiving coils in MRI typically require that RF excitation be accomplished using the body coil. This process requires that the receiving coils contain blocking circuitry to increase the overall circuit impedance during RF excitation and withstand the electromotive force induced by the applied electromagnetic field. The aim of this study was to determine the optimal impedance range required during RF excitation based on an assessment of image quality. The experimental results are fit by an exponential model and establish criteria that can be applied for general receiver coil design. PMID- 10873198 TI - Moment method analysis of mutual interaction in MRI phased array coils. AB - We describe the use of a computational method for determining the overlap distance minimizing the mutual interaction between two adjacent coils that constitute a part of a phased array system used in MRI. The method is based upon the method of moments, and the analysis is carried out at a target imaging frequency to obtain the overlap distance. For a variety of complex RF phased array coils, we can determine, using the proposed approach, the overlap distance between nearest neighbor block element RF coils such that the mutual interaction is nullified or minimized. We give experimental results to validate the proposed approach. When compared with the experimental data, our theoretical prediction is in excellent agreement. PMID- 10873199 TI - SMASH imaging with an eight element multiplexed RF coil array. AB - SMASH (SiMultaneous Acquisition of Spatial Harmonics) is a technique which can be used to acquire multiple lines of k-space in parallel, by using spatial information from a radiofrequency coil array to perform some of the encoding normally produced by gradients. Using SMASH, imaging speed can be increased up to a maximum acceleration factor equal to the number of coil array elements. This work is a feasibility study which examines the use of SMASH with specialized coil array and data reception hardware to achieve previously unattainable accelerations. An eight element linear SMASH array was designed to operate in conjunction with a time domain multiplexing system to examine the effectiveness of SMASH imaging with as much as eightfold acceleration factors. Time domain multiplexing allowed the multiple independent array elements to be sampled through a standard single-channel receiver. SMASH-reconstructed images using this system were compared with reference images, and signal to noise ratio and reconstruction artifact power were measured as a function of acceleration factor. Results of the imaging experiments showed an almost constant SNR for SMASH acceleration factors of up to eight. Artifact power remained low within this range of acceleration factors. This study demonstrates that efficient SMASH imaging at high acceleration factors is feasible using appropriate hardware, and that time domain multiplexing is a convenient strategy to provide the multiple channels required for rapid imaging with large arrays. PMID- 10873200 TI - A multicoil array designed for cardiac SMASH imaging. AB - Recently, several partially parallel acquisition (PPA) techniques have been presented which use spatial information inherent in an RF coil array to reconstruct an image from a reduced set of phase encoding steps. PPAs represent a change in paradigm for the RF coil designer since the focus for arrays to be used with PPAs is to optimize the spatial encoding that is provided by the array. One of the first practical implementations of PPA imaging was demonstrated using the SMASH technique. In this study, we present our results from the construction of the first array designed specifically for cardiac SMASH imaging. Additional design criteria are presented for SMASH arrays that are not considered in conventional array design. Using these design criteria, a four-element array was constructed and then tested in SMASH imaging experiments in the heart. This array has been used in all of our initial cardiac and head SMASH studies with good results. PMID- 10873201 TI - MR imaging of RF heating using a paramagnetic doped agarose phantom. AB - In this paper, we present the first description of a technique to visualize and quantitate radiofrequency (RF) heating of a tissue phantom during a magnetic resonance imaging (MRI) procedure. We evaluated the heating patterns of four 10 cm diameter transmit/receive surface coils with differing degrees of distributed capacitance. The tissue phantom was a 6% agarose gel doped with 40 mM Na(4)HTm[DOTP], and possesses a conductivity intermediate to human muscle and fat. Heating was discerned via phase difference mapping using the large temperature dependent chemical shift coefficient for 23Na in Na(4)HTm[DOTP]. This coefficient is -0.5 ppm/ degrees C. Heating was highest where the phantom was closest to the surface coils, dropping off towards the center of the coil. No significant difference was observed in the heating patterns between the different surface coils. For the experimental setups used in this study, electric field 'hot spots' at the areas corresponding to the placement of the capacitor gaps were not observed. PMID- 10873202 TI - Investigation of complex phased array coil designs for cardiac imaging. AB - In this study we present a method to simulate complex phased array coil designs for cardiac imaging. It is based on the combination of numerically calculated B(1) field vectors for each coil of the array and a noise resistance data set, which is acquired only once with a set of test coils. This technique allowed fast assessment of the SNR performance of arbitrary geometries of single coils to be used as building blocks in complex array configurations. In addition, since clinical scanners usually provide only four receiver channels, we used this method to investigate the use of hardware combiners for different array configurations, consisting of up to eight coils. Simulated array geometries resulted in up to approximately 30% gain in SNR for deep cardiac structures, compared to a conventional linear four coil array. This was confirmed by phantom experiments with implemented coils. PMID- 10873203 TI - Design and fabrication of a three-axis multilayer gradient coil for magnetic resonance microscopy of mice. AB - There is great interest in the non-destructive capabilities of magnetic resonance microscopy for studying murine models of both disease and normal function; however, these studies place extreme demands on the MR hardware, most notably the gradient field system. We designed, using constrained current minimum inductance methods, and fabricated a complete, unshielded three-axis gradient coil set that utilizes interleaved, multilayer axes to achieve maximum gradient strengths of over 2000 mT m(-1) in rise times of less than 50 micros with an inner coil diameter of 5 cm. The coil was wire-wound using a rectangular wire that minimizes the deposited power for a given gradient efficiency. Water cooling was also incorporated into the coil to assist in thermal management. The duty cycle for the most extreme cases of single shot echo planar imaging (EPI) is limited by the thermal response and expressions for maximum rates of image collection are given for burst and continuous modes of operation. The final coil is capable of the collection of single shot EPI images with 6 mm field of view and 94 microm isotropic voxels at imaging rates exceeding 50 s(-1). PMID- 10873204 TI - Principles of active acoustic control in gradient coil design. AB - The new principles of active acoustic control in gradient coil design are introduced and theoretical expressions are developed for the far field acoustic output for a coil system comprising four or more flat rectangular coil sectors. Each sector consists of a split plate arrangement in which are embedded two windings, an outer primary winding and a narrow inner re-entrant loop control winding immediately adjacent to and surrounding the split or air gap. The wire spacing of the control winding is made small so as not to affect substantially the magnetic field created by the primary winding. Experimental results are produced for one sector which show an average difference in acoustic output of 34.9 dB when the control winding is appropriately driven. The theoretical expressions developed are used to fit the experimental data and indicate good agreement with regard to the form of the output response. PMID- 10873205 TI - Proton MR spectroscopy of prostatic tissue focused on the detection of spermine, a possible biomarker of malignant behavior in prostate cancer. AB - To investigate whether polyamines may be valuable diagnostic and prognostic markers in prostate cancer, the presence of polyamines was studied in various human prostatic tissues using both proton magnetic resonance (MR) spectroscopy and high-pressure liquid chromatography (HPLC). The HPLC results showed that normal and benign hyperplastic prostatic tissues were characterized by a high content of spermine. Spermine levels were reduced in tumor tissue, especially in prostatic carcinoma with metastases, and in xenografts of human prostatic carcinoma cells. These preliminary results indicate that spermine may be used as a biomarker for malignant behavior. The MR spectroscopy study showed that it is possible to detect spermine resonances in prostatic biopsy material by one dimensional and two-dimensional J-resolved MR spectroscopy at high field (600 MHz). Localized one-dimensional in vitro MR spectra obtained at the clinical field strength of 1.5 T showed spermine signals in the region between 3.0 and 3.3 ppm. In in vivo MR spectra of the human prostate, however, these signals were obscured by resonances of choline (3.2 ppm) and creatine (3.0 ppm). PMID- 10873206 TI - Hyperammonemia and chronic hepatic encephalopathy: an in vivo PMRS study of the rat brain. AB - The brain energy metabolism of rats affected by chronic hepatic encephalopathy due to portacaval shunting was monitored by in vivo 31P-nuclear magnetic resonance spectroscopy before and after ammonium acetate administration. With respect to healthy unoperated and to sham operated controls, portacaval shunting decreased the levels of the nuclear magnetic resonance (NMR) visible brain phosphocreatine and nucleoside phosphates, and the intracellular [free Mg(2+)]. Ammonium acetate induced a further decrease of the levels of the NMR detectable phosphocreatine and nucleoside triphosphates and of the [free Mg(2+)], while the PMR spectra of the brain of non-shunted rats did not show any significant change even after treatment with ammonium acetate. PMID- 10873207 TI - Artifacts in CSI-measurements caused by the drift of the static magnetic field. AB - In chemical shift resolved spectroscopic imaging (CSI) temporal changes in the static magnetic field (drift) can lead to distortions of the phase encoding process. This can result in localization artifacts. The extent of the artifact depends on the size of the drift, the number of acquisitions, as well as on the combination of the size of the field of view and the number of phase encoding gradient steps. Furthermore, it is affected by the succession of the phase encoding gradients. Precautions are described which allow substantial minimization of the artifact. PMID- 10873208 TI - Semi-quantitative approach to estimating GFR by magnetic resonance imaging. AB - OBJECTIVE: The purpose of this study was to compare a semi-quantitative approach to estimating glomerular filtration rate (GFR) by magnetic resonance imaging with radionuclide calculation of GFR, and to investigate whether spin echo or gradient echo is more suitable for estimating GFR. METHODS AND PATIENTS: Fourteen kidneys of seven patients (GFR ranging from 26 to 57 ml/min) were studied. Spin echo and gradient echo sequences interleaving each other at every excitation were used. After intravenous injection of gadopentetate dimeglumine, serial scans were performed. The signal intensities measured in the regions of interest were converted to time-transverse relaxation rate changes for both spin echo (DeltaR2) and gradient echo (DeltaR2*). The areas under the time-DeltaR2 and time-DeltaR2* curves were calculated as a semi-quantitative index of GFR for both spin echo and gradient echo images, and the results were compared by GFR measured by radionuclide imaging. RESULTS: The semi-quantitative index of the GFR calculated from spin echo images showed a significant correlation with the GFR measured by radionuclide imaging (r=0.85, P<0.001). On the other hand, the semi-quantitative index of the GFR calculated from gradient echo images showed no such correlation (r=0.46, P=0.10). CONCLUSION: Spin echo sequences may be more suitable than gradient echo sequences for the evaluation of GFR. PMID- 10873209 TI - Valproic acid intoxication identified by 1H and 1H-(13)C correlated NMR spectroscopy of urine samples. AB - Analysis of biological fluids by proton and carbon nuclear magnetic resonance spectroscopy (1H and 13C NMR) is a promising tool in clinical biology. We used this method for rapid toxicological screening in the case of two suicide attempts. For each case, a urine sample was analysed at 300 MHz by 1D and 2D sequences (TOCSY and HMBC) in a short experimental time. Quantification was performed by peak integration on the 1D 1H NMR spectrum. For the two patients, results showed the same resonances of the major metabolite, valproyl-O glucuronide at concentrations of 121 and 44 mmol/l. PMID- 10873210 TI - MR CAT scan: a modular approach for hybrid imaging. AB - In this study, a modular concept for NMR hybrid imaging is presented. This concept essentially integrates different imaging modules in a sequential fashion and is therefore called CAT (combined acquisition technique). CAT is not a single specific measurement sequence, but rather a sequence design concept whereby distinct acquisition techniques with varying imaging parameters are employed in rapid succession in order to cover k-space. The power of the CAT approach is that it provides a high flexibility toward the acquisition optimization with respect to the available imaging time and the desired image quality. Important CAT sequence optimization steps include the appropriate choice of the k-space coverage ratio and the application of mixed bandwidth technology. Details of both the CAT methodology and possible CAT acquisition strategies, such as FLASH/EPI-, RARE/EPI- and FLASH/BURST-CAT are provided. Examples from imaging experiments in phantoms and healthy volunteers including mixed bandwidth acquisitions are provided to demonstrate the feasibility of the proposed CAT concept. PMID- 10873211 TI - Enhancement characteristics of hepatic focal nodular hyperplasia and its scar by dynamic magnetic resonance imaging. AB - RATIONALE AND OBJECTIVES: To determine the relationship between the lesion and the scar enhancement characteristic in a series of hepatic Focal Nodular Hyperplasia (FNH) lesions studied with dynamic MR imaging. METHODS: Nine patients with FNH were studied. The slice showing the largest scar was selected for the dynamic single slice T1-weighted Gradient-echo sequence before and after contrast administration (15 images, one every 20 s). Analysis was performed with ROI measurements in the lesion and the scar. Signal-intensity and enhancement curves were obtained from both structures. RESULTS: Dynamic MRI showed the typical homogeneous early enhancement of the lesion with delayed enhancement of the scar. The scar enhanced early and vigorously in all cases. Two patterns of enhancement curves were defined. In the parallel pattern, both curves started early, quickly reaching a plateau maintained over time (77.8%). In the divergent pattern the curve of the scar was above the curve of the FNH (22.2%), after the maximum slope was reached, with progressive separation of the curves. CONCLUSION: There is a hypervascular scar enhancement within FNH lesions with either a parallel or divergent course after the maximum early enhancement. PMID- 10873212 TI - MR imaging of Baker cysts --prevalence and relation to internal derangements of the knee. AB - RATIONALE AND OBJECTIVES: To evaluate the prevalence of popliteal cysts in patients studied with magnetic resonance (MR) imaging. Cyst presence and volume will be related to other internal derangement of the knee. MATERIALS AND METHODS: Three hundred and eighty-two consecutive patients with a MR study of the knee were included. Images were obtained in the three spatial orthogonal planes and evaluated through consensus. Four categories were established for the Baker cyst and synovial fluid (absence, minimum, moderate and massive), and the presence of meniscal, cruciate ligaments and cartilage lesions were recorded. Statistical analysis was carried out with bivariate analysis (chi(2) of Pearson and Gamma tests). RESULTS: From the 382 patients, 145 had Baker cysts (38.0%). Cyst content was minimum in 99, moderate in 34 and massive in 12. Joint effusion was observed in 269 patients (70.4%), being minimal in 140 patients, moderate in 119 and massive in ten. Meniscal lesions were observed in 195 patients (51%), while 58 patients (15%) had a cruciate ligament lesion. Baker cyst had a statistically significant direct relationship with the presence and quantity of synovial fluid (P=0.002) and with the presence and type of meniscal lesion (P=0.01) but not with cruciate ligaments or cartilage lesions. CONCLUSIONS: The prevalence of Baker cysts in MR studies of the knee is high. Its presence and volume are related to the quantity of synovial fluid, and to the presence and severity of meniscal lesions. PMID- 10873213 TI - Anatomic variations of anterior cerebral artery cortical branches. AB - The anterior cerebral artery (ACA) is a major vessel responsible for the blood supply to the interhemispheric region. The ACA segment after the anterior communicating artery (AComA) origin is called the distal ACA and has central and cortical branches. The cortical branches are distributed in the different regions of the orbital and medial part of the brain. The objects of this study are the anatomical variations found in the distal ACA. In 76 hemispheres the ACA distal branches were injected with latex and dissected under microscope magnification. Vessel diameters and distances between vessel origins and anterior communicating artery were recorded and analyzed. Microsurgical dissection was carried out to demonstrate anatomic variations of these vessels. Average diameter of ACA at origin was 2.61 +/- 0.34 mm and average diameter of cortical branches diameter ranged from 0.79 +/- 0.27 mm to 1.84 +/- 0.3 mm. Distances between vessel origin and AComA ranged from 7.68 +/- 3.91 mm (orbitofrontal) to 112.6 +/- 11.63 mm (inferior internal parietal). This study found anatomical variations: a single (azygos) ACA was present in one case and three in three cases. Crossing branches of the distal ACA to the contralateral hemisphere were present in 26% of the cases. In some cases a single ACA may supply the posterior hemispheric region through crossing branches. This calls attention to potential bilateral brain infarcts due to a single unilateral ACA occlusion. PMID- 10873214 TI - Anatomical evaluation of the groove for the vertebral artery in the axis vertebrae for atlanto-axial transarticular screw fixation technique. AB - Anatomical measurements were studied on 40 dry axis vertebrae to determine the suitability of the groove for the vertebral artery for atlanto-axial transarticular screw fixation technique. We measured 13 parameters including three angular and 10 linear dimensions related to the groove of the vertebral artery, pedicle, and pars interarticularis and evaluated 80 measurements for each parameter. All measurements were done after placing a Kischner guide wire through the pedicle. We found that differences between measurements on the left and right sides of each vertebra were nonsignificant. In spite of the variability in measurements such as height, width, and median angle of the pedicle, the decline angle for instrumentation, the depth of the groove for the vertebral artery, and the internal height of the pars interarticularis, all of these had good symmetry. However, there were statistically significant differences between the sides in measurements for both the width (P=0.05) and the angle (P<0.02) of the pedicle allowing instrumentation and they did not show good symmetry. The risk of vertebral artery injury was found to be 22.5% per specimen, or 16.25% per screw inserted because the internal height of the pars interarticularis at point of fixation was 2)-mannosidase activity converts the glycoprotein to its Man(5) form, identifiable by its mass of 14,899 [M + H](+); this species subsequently is converted, by the actions of alpha-(1 - > 3) and alpha-(1 --> 6)-mannosidases, to the Man(1) form via Man(4), Man(3), and Man(2). The Man(1) glycoform (which is readily isolated) has then similarly been used for identifying beta-(1 --> 4)-mannosidase and the derived Man(0) form has served in turn as a natural substrate for beta-(1 --> 4) N-acetylglucosaminidase producing a species possessing a single asparagine-linked GlcNAc residue (mass 13,886). Mannose liberated from the actions of mannosidases can, if desired, be quantified by, for example, chromatography. The actions and specificities of endoglycosidases such as a peptide-N-glycosidase F (PNGase F) and of endo-N acetlyglucosaminidases (e.g., endo-F and endo-H), which respectively cleave between the GlcNAc&bond;Asn and GlcNAc&bond;GlcNAc bonds of N-linked glycoproteins, are also demonstrable by MALDI-ToF analysis of RNase B (and derived products). From these digests the completely deglycosylated polypeptide corresponding to RNase A in which Asn has been converted to Asp (mass 13,684) and a species corresponding to RNase A + GlcNAc (mass 13,886) are produced, together with their corresponding free oligosaccharides which are amenable to analysis by both MALDI-ToF and by HPLC. PMID- 10873271 TI - Biotinylated steroid derivatives as ligands for biospecific interaction analysis with monoclonal antibodies using immunosensor devices. AB - Systematic ligand-binding studies of the biospecific interaction between steroids and antisteroid antibodies can be performed in real time using biosensor techniques. In this study, quartz crystal microbalance (QCM) and surface plasmon resonance (SPR) biosensor systems were applied. Different biotinylated testosterone (T) and 17beta-estradiol (E2) derivatives were preincubated with streptavidin and immobilized on the sensor surfaces. We obtained low matrix densities of antigen enabling the investigation of the binding kinetics and position specificities of various anti-E2 and anti-T monoclonal antibodies (mAbs) to these steroidal compounds. The highest immunoreactivity of anti-E2 and anti-T mAbs is not necessarily for the specific modified steroid that was used as a protein-coupled hapten for immunization. The kinetic data confirm that both 3- and 19-specific anti-T mAbs do not discriminate between the 3- and 19 biotinylated T derivatives, whereas the 7alpha-biotinylated T probe showed no affinity to these two anti-T mAbs. In the case of the 3-specific anti-E2 mAb, comparable interaction data were found for 3- and 6alpha-biotinylated E2 compounds. The 6-specific anti-E2 mAb showed comparable ligand binding, but a significant higher dissociation rate to the position-specific antigen. The QCM and SPR results correspond well to the data from cross-reactivity studies in solution as well as to enzyme immunoassay equilibrium measurements. PMID- 10873272 TI - Method enabling pyrosequencing on double-stranded DNA. AB - Pyrosequencing is a new nonelectrophoretic, single-tube DNA sequencing method that takes advantage of co-operativity between four enzymes to monitor DNA synthesis (M. Ronaghi, M. Uhlen, and P. Nyren, Science 281, 363-365). Pyrosequencing has so far only been performed on single-stranded DNA. In this paper different enzymatic strategies for template preparation enabling pyrosequencing on double-stranded DNA were studied. High quality data were obtained with several different enzyme combinations: (i) shrimp alkaline phosphatase and exonuclease I, (ii) calf intestine alkaline phosphatase and exonuclease I, (iii) apyrase and inorganic pyrophosphatase together with exonuclease I, and (iv) apyrase and ATP sulfurylase together with exonuclease I. In many cases, when the polymerase chain reaction was efficient exonuclease I could be omitted. In certain cases, additives such as dimethyl sulfoxide, single stranded DNA-binding protein, and Klenow DNA polymerase improved the sequence quality. Apyrase was the fastest and most efficient of the three different nucleotide degrading enzymes tested. The data quality obtained on double-stranded DNA was comparable with that on single-stranded DNA. Pyrosequencing data for more than 30 bases could be generated on both long and short templates, as well as on templates with high GC content. PMID- 10873273 TI - Development of a stable isotope approach for the inductively coupled plasma-mass spectrometry determination of oxidized metallothionein in biological materials. AB - The use of isotope dilution analysis (IDA) with inductively coupled plasma-mass spectrometry (ICP-MS) for the determination of oxidized metallothionein (MT) by a Cd-saturation method is investigated. The method developed here is a modification of an earlier methodology which used a radioactive Cd isotope ((109)Cd). While retaining the many advantages of this previous approach, the procedure presented here uses stable isotope ratio measurements ((114)Cd/(111)Cd) for the determination of MT. Experimental parameters governing the instrumental precision and accuracy for isotope ratio measurements of Cd by ICP-MS were characterized. Systematic errors, including mass bias, detector dead time, and spectroscopic interferences, could be easily corrected. The isotope dilution ICP-MS method was validated by the determination of very low levels of cadmium in biological certified reference materials (NIST SRM 2670 freeze-dried urine, IAEA H-8 horse kidney, and BCR TP-25 lichens). Finally, the IDA procedure was evaluated for the determination of oxidized MT by a Cd-saturation method previously developed using radioactive (109)Cd. The final procedure was applied to the quantification of MT in Long-Evans Cinnamon rat liver cytosol samples and the results were compared with data obtained for the same samples using the reference (109)Cd methodology. A good agreement between the analytical values obtained by both methods was observed. PMID- 10873274 TI - A streptavidin surface on planar glass substrates for the detection of biomolecular interaction. AB - Based on the requirements of biomolecular interaction analysis on direct optical transducers, a streptavidin surface is examined. A general protocol was developed allowing the immobilization of biotinylated compounds using the rife biotin streptavidin system. This type of surface modification can be applied to all biosensors using glass surfaces as sensor devices. Reflectometric interference spectroscopy (RIfS), a label-free, direct optical method was used to demonstrate the quality of the transducer surfaces. The surface modification is based on an aminofunctionalized polyethylene glycol layer covalently bound to the silica surface of the transducer and shows very little nonspecific binding. Biotin molecules can be easily coupled on such layers. Streptavidin followed by a biotinylated estrone derivative was immobilized by incubation of the biotinylated transducer surface. For the streptavidin layer we obtained interference signals corresponding to a protein monolayer. Finally, using a surface prepared as described above, biomolecular interaction experiments with an antibody against estrone were carried out to show the quality of the transducer surface. With RIfS all of the affinity-based surface modifications can be detected online and time resolved. PMID- 10873275 TI - Simultaneous detection of ubiquinol-10, ubiquinone-10, and tocopherols in human plasma microsamples and macrosamples as a marker of oxidative damage in neonates and infants. AB - A HPLC-method was developed for the simultaneous detection of the lipophilic antioxidants ubiquinol-10, ubiquinone-10, alpha-tocopherol, and gamma-tocopherol in plasma macrosamples (100 microl) as well as microsamples (10 microl) using electrochemical detection with internal standardization (gamma-tocotrienol, ubiquinol-9, ubiquinone-9). The use of a mobile phase containing ammonium formate instead of lithium perchlorate as conductivity salt and the use of a PEEK column instead of a steel column improved the reproducibility of the method. The components were separated on a RP C18 column. The detection limits for all components were between 1 and 28 fmol. The within-day precision varied between 3 and 13% for all measured substances. The analytical recovery was between 93 and 109%. The plasma levels from 10 healthy donors were determined in microsamples and macrosamples taken by micropuncture or venous puncture, respectively. A good correlation of the antioxidant levels for both methods strengthened the reliability and the transferability of the results. The present method can be used to assess the role of these antioxidants in diseases associated with oxidative damage in childhood. PMID- 10873276 TI - Rapid SLT gene detection on polyethylene-coacrylic acid film without molecular labels or surface-fouling agents. AB - The commercially available copolymer of 10 mol% acrylic acid and polyethylene is easily formed into a nonfluorescing, non-polynucleotide-adsorbing film. The film has surface carboxylate functions whose concentration can be increased by heating to 80 degrees C in 30% NaOH. The carboxylate groups will react at pH approximately 7 with commercially available, oligo-DNA, 2-8 ng/microl, that has been synthesized with a C(12)-alkylamino tail on the 5'-end. The reaction is mediated with water-soluble carbodiimide reagent and is assumed to result in a primary amide bond between the polymer film and the modified oligo-DNA. The tethered oligo-DNA retains its hybridization activity, and its surface concentration is sufficient to permit qualitative, labelless detection of hybridized target by fluorescence after brief staining with ethidium bromide. The film is used to detect Shiga-like toxin gene II (SLT-II) from Escherichia coli O157:H7 after asymmetric, capillary, PCR amplification, and a 4-h hybridization. Captured target may be removed from the film using distilled water, after which the film can be used again without noticeable loss of activity. The method provides relatively rapid detection of PCR amplimers without having to use molecular labels, or surface-fouling agents. PMID- 10873277 TI - Analysis of isoaspartate in peptides and proteins without the use of radioisotopes. AB - A rapid and sensitive HPLC-based method for quantitating isoaspartate levels in peptides and proteins is described. The analyte is incubated for 40 min with S adenosyl-l-methionine and the commercially available enzyme protein l-isoaspartyl methyltransferase. Methylation of isoaspartyl sites results in stoichiometric production of S-adenosyl-l-homocysteine that is separated from the other components of the reaction by reversed-phase HPLC and quantitated online by absorbance at 260 nm. This method can accurately detect 5 pmol or less of isoaspartate and works with tryptic digests as well as intact proteins. Using a commercially available isoaspartyl peptide, the relationship between isoaspartate levels and S-adenosyl-l-homocysteine production was found to be linear and stoichiometric over a range of 5-250 pmol. Compared to methods that measure [(3)H]methanol production after methylation with S-adenosyl-l-[methyl (3)H]methionine, the HPLC method is safer, faster, less expensive, and equally sensitive. PMID- 10873278 TI - Quantitation of the blocking effect of tween 20 and bovine serum albumin in ELISA microwells. AB - ELISA provides a highly sensitive procedure for quantitating antigens and antibodies. In that assay, microwells are coated initially with a specific ligand and then saturated with inert molecules to minimize nonspecific background. Coating can be improved by pretreating the microwells with poly-l-lysine (PLL). Proteins and Tween 20 are most often used to block vacant binding sites in enzyme linked immunosorbent assay (ELISA). In the present study the blocking effects of Tween 20 and bovine serum albumin (BSA) were estimated using an original novel approach. In the assay the magnitude of saturation of the microwells was quantitated by measuring the enzymatic activity of alkaline phosphatase adsorbed to residual vacant sites in the microwell. Tween 20 completely saturated ELISA microwells at concentrations higher than 2 microg/ml. If the microwells were pretreated with PLL, even high concentrations of the detergent did not completely saturate the wells. In contrast, BSA completely saturated both PLL-treated and nontreated microwells at 5 microg/ml. Complementation of Tween 20-induced saturation of PLL-treated microwells was achieved only by addition of BSA at concentration required for BSA alone to reach complete saturation. This approach is applicable for assessing binding to ELISA microwells of any reagent of choice either as a ligand or as a blocking reagent. PMID- 10873279 TI - Monitoring equilibria and kinetics of protein folding/unfolding reactions by capillary zone electrophoresis. AB - A method is described here for studying conformational transitions of proteins due to denaturing agents: capillary zone electrophoresis (CZE) in acidic, isoelectric buffers. The sample is run in 50 mM isoelectric glutamic acid (pH = pI = 3.2) added with 1 mM oligoamine (tetraethylene pentamine) for quenching protein interaction to the capillary wall (final pH = 3.3). Muscle acylphosphatase (AcP), in this buffer, exhibited a free solution mobility of 2.63 x 10(-4) cm(2) V(-1) s(-1). By studying the unfolding kinetics, as a function of time of incubation in 7 M urea, it was possible to measure the rate constant of the unfolding reaction, estimated to be 0.00030+/-0.00006 s(-1). The same measurements, when repeated via spectroscopic monitoring of intrinsic fluorescence, gave a value of 0.00034+/-0.00002 s(-1), thus in excellent agreement with CZE data. By equilibrium unfolding CZE studies, it was possible to construct the typical sigmoidal transition of unfolding vs urea molarity: the midpoint of this transition, at which the folded and unfolded states should be equally populated, was estimated to be at 4.56 M urea. Similar experiments by fluorometric analysis gave a value of 4.60 M urea as midpoint of the unfolding curve. PMID- 10873280 TI - Separation and purification of sphingomyelin diastereomers by high-performance liquid chromatography. AB - All naturally occurring sphingomyelins have the d-erythro-(2S,3R) configuration of the sphingoid base. We have developed a normal-phase HPLC method for the separation of this natural stereoisomer from the l-threo-sphingomyelin, which is the other stereoisomer commonly present in semisynthetic preparations of acyl chain defined sphingomyelins. The chromatographic method was developed by modification of a previously reported method for phospholipid separation on a normal-phase diol column. The separation was accomplished by a binary gradient of solvent mixtures (A) hexane:isopropanol:acetic acid (82:17:1.0 by vol) and (B) isopropanol:water:acetic acid (85:14:1.0 by vol) with 0.08 vol% triethylamine added to both solvent mixtures. The program of gradient elution was optimized for maximal separation of sphingomyelin diastereomers. For detection of the lipids, a light-scattering detector was used. This analytical scale HPLC method was also used for purification of the stereoisomers (up to 0.5 mg of N-oleoyl sphingomyelin in a single injection). The purified stereoisomers were at least 99% pure according to high-performance thin-layer chromatography and analytical HPLC. PMID- 10873281 TI - Comparison of two direct methods for the determination of fatty acids in infant feces. AB - We have validated and compared two direct methods for the determination of fatty acids in feces by capillary gas chromatography. Method I consisted of esterification of fatty acids using acetyl chloride. Method II used boron trifluoride-methanol as esterification reagent. The two methods were assayed with and without previous freeze-drying of the fecal sample. We found that the two methods could be carried out without sample freeze-drying. Precision and recovery rates were determined and the results were satisfactory. Both methods gave similar results, but Method II has certain advantages over Method I, such as speed, safety, and better recovery rates. PMID- 10873282 TI - Simultaneous synthesis of all partially methylated alditol acetates of glucosamine and galactosamine for gas chromatography-mass spectrometry analysis. PMID- 10873283 TI - A gel elution apparatus with high sample recovery. PMID- 10873284 TI - Formation of chimeric cDNAs during suppression subtractive hybridization and subsequent polymerase chain reaction. PMID- 10873285 TI - In vivo gene transfer by low-volume jet injection. PMID- 10873287 TI - Diversification of gap junction proteins (connexins) in the central nervous system and the concept of functional compartments. AB - This review describes recent progress in the identification of the molecular composition of gap junction proteins (connexins) in brain tissue. First, a general overview of gap junction function and the composition of the hemichannels (connexons) is given. Then the question of diversity of gap junction channels in the nervous system is addressed and its functional implications are discussed. Based on recent findings of gap junction mediated cell-to-cell communication we attempt to refine the concept of functionally coupled compartments in the brain. This conceptual introduction passes on to a detailed description of the coupling efficiency in glial and neuronal compartments with emphasis on innovative perspectives that allow a better understanding of gap junction function in the brain. PMID- 10873288 TI - Ultrastructure, histological distribution, and freeze-fracture immunocytochemistry of gap junctions in rat brain and spinal cord. AB - The historical development of concepts of gap junctions as sites for electrical, ionic, and metabolic coupling is reviewed, from the initial discovery of gap junctions linking heart cells, to the current concepts that gap junctions represent 'electrotonic synapses' between neurons. The ultrastructure and immunocytochemistry of gap junctions in heart, brain, and spinal cord of adult rats is examined using conventional thin sections, negative staining, grid-mapped freeze-fracture replicas, and immunogold-labeled freeze-fracture replicas. We review evidence for neuronal gap junctions at 'mixed' (combined electrical and chemical) synapses throughout adult rat spinal cord. We also show immunogold labeling of connexin43 in astrocyte and ependymocyte gap junctions and of connexin32 in oligodendrocyte gap junctions. Ultrastructural and freeze-fracture immunocytochemical methods have provided for definitive determination of the number, size, histological distribution, and connexin composition of gap junctions between neurons in all regions of the central nervous systems of vertebrate species. PMID- 10873289 TI - Gap junctional communication in the developing central nervous system. AB - The development of the central nervous system is a complex process involving multiple interactions between various cell types undergoing mitosis, migration, differentiation, axonal outgrowth, synaptogenesis and programmed cell death. For example, neocortical development is characterized by a series of transient events that ultimately leads to the formation of a discrete pattern of laminar and columnar organization. While neuron-glial cell-cell interactions have been shown to be involved in neuronal migration, recent observations that neurons are extensively coupled by gap junctions in the developing neocortex have implicated this phenomenon in the process of neocortical differentiation. The present review will examine the putative role of gap junctional intercellular communication in development of the central nervous system, with specific reference to recent studies in the development of the cerebral cortex. PMID- 10873290 TI - Synaptic modulation of neuronal coupling. AB - Electrotonic coupling among neurons in the vertebrate, and more specifically the mammalian, brain has now been demonstrated to exist in all major brain subdivisions and in the spinal cord. For many of these brain areas, recent studies have investigated the possibilities of modulation of that coupling by synaptically released transmitters and/or neuromodulators. Reviewed here is the evidence for coupling, the synaptically related factors that play roles in up- or downregulation of this type of intercellular interaction and, to the extent that they have been investigated, the intracellular mechanisms operative in changing the extent of coupled networks in the brain. The functional significance of coupling and its modulation is discussed for some of these areas. PMID- 10873291 TI - Cell coupling in the retina: patterns and purpose. AB - Gap junction channels (electrical synapses) are a major component of the central nervous system mediating both electrical and metabolic coupling between neurons and glia. Their roles are as diverse as the cell types in which they are expressed and only some of these are reviewed here. In the adult the plastic nature of the gap junction channel allows for changes in the writing of the retinal circuitry that optimize visual processing to suit ambient lighting conditions. Gap junctional communication has been proposed to play a key role in embryonic development in general and in particular during the development of the retina where its roles may include control of neurogenesis, cell specification, synaptogenesis, and the synchronization of the spontaneous electrical activity required for the sharpening of central visual projections. Here we review gap junctional coupling within the retina and present data correlating gap junction expression with development events in the chick retina. PMID- 10873292 TI - 'Non-synaptic' mechanisms in seizures and epileptogenesis. AB - The role of 'non-synaptic' mechanisms (i.e. those mechanisms that are independent of active chemical synpases) in the synchronization of neuronal activity during seizures and their possible contribution to chronic epileptogenesis are summarized. These 'non-synaptic' mechanisms include electrotonic coupling through gap junctions, electrical field effects (i.e. ephaptic transmission), and ionic interactions (e.g. increases in the extracellular concentration of K(+)). Several lines of evidence indicate that granule cells and pyramidal cells of the hippocampus, and probably other cortical neurons, can generate synchronized electrical activity after active chemical synaptic transmission has been blocked. This synchronized activity is sensitive to alterations in the size of the extracellular space, thus suggesting that electrical field effects and ionic mechanisms contribute to this synchronized activity. Recent studies also indicate that 'non-synaptic' synchronization is quite prominent early in development. Electrophysiological data from hippocampal and neocortical slices have led to a re-interpretation of the fast prepotentials (i.e. partial spikes) recorded in cortical pyramidal cells, suggesting that they may not be due to dendritic spike generation. Improvement in freeze-fracture ultrastructural techniques have led to a re-assessment of previous data on gap junctions in the nervous system and opened new approaches to the quantitative analysis and characterization of gap junctions on glia and neurons. Finally, new methods of dye/tracer coupling have the potential to provide a more rigorous basis for evaluating gap junctions and electrotonic communication between neurons in the mammalian central nervous system. Therefore, recent data continue to suggest that gap junctions and electrotonic coupling play an important role in neural integration, although additional studies using new techniques will be needed to address some of the controversial issues that have arisen over the last several decades. PMID- 10873293 TI - X-linked Charcot-Marie-Tooth disease and connexin32. AB - We studied 29 families with X-linked dominant CMT (CMTX1) neuropathy. Twenty-five families showed mutations in the coding region of the connexin32 (Cx32) gene. The mutations included five nonsense mutations, 17 missense mutations, two medium size deletions and one insertion. Most missense mutations showed a mild clinical phenotype and slowing of motor nerve conduction velocities. All five nonsense mutations, the larger deletion and the insertion showed severe clinical phenotype. Four CMTX1 families with mild clinical phenotype showed no point mutations of the Cx32 gene coding region. Two mutations of the non-coding region were identified. The first mutation was located in the nerve specific Cx32 promoter, the second mutation was located in the 5' untranslated region of the mRNA. PMID- 10873294 TI - Common themes in peripheral neuropathy disease genes. AB - After a century of study, mutations in connexin32, peripheral myelin protein22, and protein zero are now known to culminate in the prototypical phenotype of Charcot-Marie-Tooth disease. Many of these mutations have been modeled in rodents and in tissue culture. Consequently, structure-function predictions for these mutations are now possible and detailed analyses of many of them are ongoing. Despite the marked differences in the functions of these three proteins, it is profitable to consider the many similarities between them, including the types of mutational mechanisms and their effects on myelin structure and function. Accordingly, the biology and genetics of Charcot-Marie-Tooth disease and other inherited peripheral neuropathies due to mutations in these proteins are reviewed. PMID- 10873295 TI - Connexin expression in Huntington's diseased human brain. AB - In Huntington's diseased human brain, it is in the caudate nucleus (CN) and globus pallidus (GP) of the basal ganglia where nerve cell death is seen most dramatically. The distribution of five gap junction proteins (connexins 26, 32, 40, 43 and 50) has been examined in these areas in normal and Huntington's diseased human brain using immunohistochemical techniques. There was no Cx50 expression observed and Cx40 was localized in the endothelial cells of blood vessels, with the Huntington's diseased brains having more numerous and smaller blood vessels than normal tissue. Cx26 and Cx32 revealed a similar distribution pattern to each other in both normal and diseased brains with little labelling in the CN but clear labelling in the GP. Cx43, expressed by astrocytes, was the most abundant connexin type of those studied. In both normal and diseased brains Cx43 in the GP was homogeneously distributed in the neuropil. In the CN, however, Cx43 density was both increased with Huntington's disease and became located in patches. Glial fibrillary acidic protein(GFAP) staining of astrocytes was also highly increased in the CN compared with normal brains. These labelling patterns indicate a reactive astrocytosis around degenerating neurons with an increased expression of astrocytic gap junctions. The enhanced coupling state between astrocytes, assuming the junctions are functional, could provide an increased spatial buffering capacity by the astrocytes in an attempt to maintain a proper environment for the neurons, helping promote neuronal survival in this neurodegenerative disorder. PMID- 10873296 TI - Growth hormone receptor gene expression on human lymphocytic and monocytic cell lines. AB - The potential effect of growth hormone (GH) in tumorigenesis, particularly in acute leukemia is controversial. Human growth hormone has the ability to influence certain immune functions; the majority of immune cells express growth hormone receptor (GHR) on plasma membranes. We determined GHR gene expression on different human lymphocyte (JURKAT, CESS) and monocyte (U937, THP1) cell lines by reverse transcriptase polymerase chain reaction analysis of GHR mRNA after stimulating the cells with phytohaemagglutinin or phorbolester, human growth hormone and with a combination of these. The receptor gene expression showed differences; in the U937 and CESS cell lines only the stimulants were able to induce GHR mRNA expression; in the case of JURKAT cells even the hormone alone had the ability to express its own receptor gene. Both the increased TNF-alpha production of U937 (but not that of THP1 cells), and the decreased proliferation of JURKAT cells in response to GH stimuli also prove the presence of biologically active GHR on the cell surface. Our data suggest asymmetric interaction between GH or phorbolester-induced signal pathways in U937 cells sharply depending on the temporal sequence of treatments. THP1 monocytes showed no gene expression in response to any of the stimulants. The phenomenon that certain human lymphoid and monocytoid cell lines at different levels of cell differentiation are able to express the GH receptor gene could have importance in the rhGH therapy. PMID- 10873297 TI - Inhibition of antigen and calcium ionophore induced secretion from RBL-2H3 cells by phosphatase inhibitors. AB - The role of serine/threonine protein phosphatases PP1 and PP2A in mast cell secretion was investigated using the phosphatase inhibitors okadaic acid and calyculin A. Calyculin A (5-25 nm) inhibited antigen-induced secretion from a rat mucosal mast cell line (RBL-2H3) when added in conjunction with the activator. Okadaic acid (250-1000 nm) inhibited secretion only when added before activation and did so in a time- and concentration-dependent manner. Both inhibitors caused the cells to become rounder, but only calyculin A induced membrane blebbing and a loss of adherence. Okadaic acid also inhibited secretion induced by the calcium ionophore A23187, in the presence or absence of PMA, indicating that the phosphatase inhibitors act on a component of the secretory pathway downstream of calcium mobilization. Okadaic acid increased the phosphorylation of a number of proteins, as did an analogue methyl okadaate, which also inhibited secretion, but less effectively. Okadaic acid induced the phosphorylation of triton-insoluble proteins of 55, 18 and 16 kDa. The 55 kDa protein was identified as vimentin and okadaic acid induced its partial translocation to the triton-soluble fraction. Our data indicate that full secretory function in mucosal mast cells requires phosphatase activity. PMID- 10873298 TI - Induction of premature mitosis in S-blocked onion cells. AB - Onion root-tip cells were blocked at S-phase by treating them with 5-aminouracil (5AU). These cells were then further treated with caffeine/2-aminopurine (Caf/2AP) or a combination of both in the presence of 5AU. These tyrosine kinase inhibitors were able to induce premature mitosis in the S-blocked cells as evident from the breaks and gaps in the metaphase chromosomes and the presence of laggards and fragments in anaphase. Immunofluorescence showed normal spindle formation in these cells. Immunoblotting of cyclin B revealed that the level of cyclin B was slightly higher in the recovered and treated samples than the S blocked one. The level of p(34)was found to be almost equal in all three samples as expected. We failed to observe any significant difference in the level of p(34)containing phosphorylated tyrosine. Such premature induction of mitosis by the purine derivatives has also been reported in BHK cells. However, those cells failed to progress through mitosis. A comparative analysis indicates that the plant cells and the animal cells, perhaps, follow identical pathway for the initiation of mitosis. The possible causes for differential behaviour in mitotic progression in these cells have been discussed. PMID- 10873299 TI - Steroid hormone (hydrocortisone, oestradiol and testosterone) uptake, storage or induced synthesis in tetrahymena. AB - After cyclodextrin-coated 10(-6) m steroid hormone treatment for 3 days (hormonal imprinting), Tetrahymena cells and their media were analysed by radioimmunoassay for the same hormone and for the presence of the other two. In the absence of hormone treatment, the cells contained no detectable levels of the three steroids. By 2 days in fresh medium following exposure of cells to a 72 h pretreatment of each specific hormone, correspondingly high quantities of hydrocortisone and oestradiol, but lesser quantities of testosterone, were found in both the media and the cells. One week after treatment only traces of hydrocortisone were found, exclusively within the cells themselves. Oestradiol was present in measurable quantities in both cells and media, whereas testosterone was only present in the medium. The presence of the other two hormones to the one used in the pretreatment were not usually present, except that when testosterone had been given, some oestradiol was also detected at 48 h, suggesting Tetrahymena has a functional cytochrome P(450)aromatase. PMID- 10873300 TI - Mouse homolog of Saccharomyces cerevisiae spo11 is induced in normal mu(+)B-cells by stimuli that cause germline C(H) transcription and subsequent class switch recombination. AB - The first step of Ig heavy chain class switch recombination (CSR) is considered to be DNA double strand break (DSB) formation in the two switch (S) regions (S(mu) and downstream S(H)), although the underlying mechanism is unknown. Recently, it has been demonstrated that at least Spo11, a homolog of the novel type II topoisomerase (topo VI) that catalyzes DSB formation, is involved in the initiation of meiotic recombination of Saccaromyces cerevisiae. In the present study, we examined whether the mouse homolog of Spo11 is induced in normal mouse mu(+)B-cells by stimuli that cause an early step of CSR, germline C(H) transcription, and subsequent CSR. Two CSR systems were used: IgA CSR induced by all-trans retinoic acid, IL-5, and LPS, and IgG1 CSR induced by IL-4 and LPS. Germline transcript and mouse Spo11 expression were analyzed by RT-PCR. In both systems, first germline transcripts were clearly detected on day 2 and then Spo11 was detected on day 3, increasing thereafter with time. The time course of changes in Spo11 expression coincided with that of CSR. Spo11 seems to be induced by CSR-inducing stimuli, regardless of the direction of CSR. These results suggested that mouse Spo11 might participate in the initiation step of CSR. PMID- 10873301 TI - Role of T cell subsets in the modulation of Mycobacterium avium growth within human monocytes. AB - Mycobacterium avium frequently causes disseminated disease in patients with advanced AIDS with low CD4 counts. The effects of T lymphocyte on intracellular M. avium replication were examined. Plastic adherent monocytes and nonadherent lymphocytes were separated from peripheral blood mononuclear cells. After infection with M. avium, monocytes were cultured with or without autologous lymphocytes (1-10 cells/monocyte) for up to 7 days. Addition of lymphocytes to M. avium-infected monocytes significantly decreased intracellular M. avium growth after 7 days culture (n = 11, P < 0.01, paired t test) and increased IFN-gamma production compared to monocytes alone. Neutralizing IFN-gamma partially abrogated lymphocyte activity. CD4 depletion diminished anti-mycobactericidal effects and CD8(+) lymphocytes increased intracellular M. avium growth (P < 0.05, n = 5, t test). These data suggest that interactions between monocytes and nonadherent cell fractions such as CD4(+) T cells and NK cells are important in intracellular M. avium growth modulation in monocytes from healthy humans. PMID- 10873302 TI - CD8(+) T cells secreting type 2 lymphokines are defective in protection against viral infection. AB - Effector T cells secreting type 1 and/or type 2 lymphokines (Tc1, Tc0, Tc2) were generated in vitro from CD8(+) T cells of mice with a transgenic TCR recognizing lymphocytic choriomeningitis virus (LCMV) glycoprotein to compare their effector function in vitro and in vivo. Tc1, Tc2, and Tc0 showed similar Fas- and perforin mediated cytotoxicity in vitro. Upon adoptive transfer, Tc2 and Tc0 effectors were less efficient than Tc1 at controlling LCMV or recombinant vaccinia virus expressing the LCMV glycoprotein in vivo. Tc2 and Tc0 had decreased surface VLA-4 density and deficient activation-induced LFA-1/ICAM-1-dependent homotypic adhesion in vitro. Therefore, the reduced antiviral activity in vivo of Tc2 and Tc0 compared with Tc1 is not due to reduced cytotoxic activity or IFN-gamma secretion but may be explained by defective homing to the target organ due to decreased expression and/or lower activity of adhesion molecules. PMID- 10873303 TI - The endothelium and cytokine secretion: the role of peroxidases as immunoregulators. AB - The endothelium is frequently exposed to many proinflammatory mediators. The present study was done to determine the effects of human recombinant myeloperoxidase (MPO) and porcine eosinophil peroxidase (EPO) on certain endothelial cell (HUVEC) functions. The following areas were evaluated: (1) production of reactive oxygen intermediates (ROI), (2) cytokine secretion, and (3) regulation of mRNA cytokine transcripts. Both MPO and EPO induced the production of ROI, but an enzymatically inactive form of MPO (iMPO) was the most effective. Enzymatically inactive MPO, but not MPO, induced the secretion of interleukins 6 and 8 and granulocyte-monocyte colony-stimulating factor. A ribonuclease protection assay indicated that both iMPO and MPO upregulated mRNA cytokine transcripts; however, the former was markedly more effective. The simultaneous addition of EPO and iMPO resulted in a decrease in cytokine-specific mRNA. These data indicate a major role for peroxidases in the regulation of inflammation. PMID- 10873304 TI - Differences in interleukin-12 and -15 production by dendritic cells at the early stage of Listeria monocytogenes infection between BALB/c and C57 BL/6 mice. AB - The mechanisms responsible for the resistance of C57BL/6 mice and for the susceptibility of BALB/c mice to infection with Listeria monocytogenes were studied by comparing early IL-12 and IL-15 production by dendritic cells (DC) after infection with L. monocytogenes. Splenic DC expressing CD11b(low) and CD11c(+) obtained from C57BL/6 mice at 3 and 6 h after L. monocytogenes infection expressed higher levels of IL-12 p40 mRNA and IL-12 p40 protein than did those from BALB/c mice. Concurrently, a larger amount of IFN-gamma was produced by the splenic T cells from C57BL/6 mice in response to immobilized anti-TCRalphabeta mAb than by those from BALB/c mice, while the splenic T cells from BALB/c mice produced a higher level of IL-4 upon TCR alphabeta stimulation than did those of C57BL/6 mice. IL-15 mRNA and intracellular IL-15 protein were detected more abundantly in the DC from C57BL/6 mice than in those from BALB/c mice on day 3 after infection. CD3(+) IL2Rbeta (+) cells in the spleen were increased in C57BL/6 mice but not in BALB/c mice at the early stage after infection. Furthermore, IL-12Rbeta2 gene expression was up-regulated in T cells from C57BL/6 mice but not in those from BALB/c mice at the early stage after listerial infection. These results suggest that the difference in early production of IL-12 and IL-15 by DC may at least partly underlie the difference in susceptibility to L. monocytogenes between C57BL/6 and BALB/c mice. PMID- 10873305 TI - Preferential proliferation and differentiation of double-positive thymocytes into CD8(+) single-positive thymocytes in a novel cell culture medium. AB - The identification of factors that regulate the proliferation and differentiation of double-positive (DP) into CD4(+) and CD8(+) single-positive (SP) thymocytes has proven difficult due to the inability of DP thymocytes to proliferate, expand, and differentiate into SP thymocytes in available cell culture media. Here we report on the ability of DP thymocytes to differentiate in a novel conditioned medium, termed XLCM, derived from the supernatant of mitogen activated human cord blood mononuclear cells. During a 5-day culture in XLCM in the absence of thymic stromal cells, DP thymocytes from normal mice and MHC double knockout mice (lack SP thymocytes) proliferate, expand, and differentiate into several (alphabetaTCR(+), NK1.1(+)alphabetaTCR(+), and gammadeltaTCR(+)) subsets of CD4(+) and predominantly CD8(+) SP thymocytes. These studies suggest that the use of XLCM may aid in the characterization of factors that regulate the differentiation of DP thymocytes into CD8(+) SP thymocytes. PMID- 10873306 TI - Immunosuppressive effect on T cell activation by interleukin- 16-cDNA-transfected human squamous cell line. AB - It is well known that it is difficult to induce an immunotolerance with allogeneic skin transplantation. We attempted to find the immunosuppressive protocol for prolonging skin allograft rejection by using interleukin-16 because IL-16 is considered one of the natural ligands to CD4 molecules. First we examined whether synergistic immunosuppressive effects of recombinant IL-16 plus anti-CD4 mAbs are induced in mixed lymphocyte reaction (MLR). Next we used IL-16 cDNA-transfected OSC-20 (human oral squamous cell carcinoma cell line) as an in vitro model of the epidermal keratinocyte equivalent and examined whether this transfectant could inhibit the activation of allogeneic T cells. Our data indicated that IL-16 clearly inhibited human MLR and that IL-16 increased synergistically the immunosuppressive effect of anti-CD4 mAb. We also used IL-16 transfectant and this produced more than 50 ng/ml of IL-16 in the supernatant by which human MLR was significantly inhibited. Furthermore, this transfectant also inhibited the activation of allogeneic lymphocytes stimulated directly with transfectant cells. These results indicated that the IL-16-producing allogeneic skin graft might have a local immunosuppressive action that would prolong graft survival. PMID- 10873309 TI - Modeling Orthokinetic Coagulation in Spatially Varying Laminar Flow. AB - An orthokinetic coagulation model including the effects of agglomeration and local stress-induced aggregate breakup was developed. This model was used to simulate coagulation in the flow between two eccentrically located and rotating cylinders. Four methods of modeling coagulation in the flow system were examined. The first technique used a volume-weighted average of the local strain rates, while a second method used an equivalent volume-weighted power (G). A third method treated each volume element as a separate batch reactor and determined a final volume-averaged floc population. The final modeling technique applied mass transfer between each of the elements. Results indicated that substantial differences in average particle diameters and populations were generated with each of the methods, especially where mass transfer between the elements was considered. It was concluded that mass transfer between regions of varying flow strain rate and/or velocity gradient should be included in accurate coagulation modeling. Copyright 2000 Academic Press. PMID- 10873307 TI - gammadelta T cells may dichotomously modulate infection with avirulent Salmonella choleraesuis via IFN-gamma and IL-13 in mice. AB - To investigate the roles of gammadelta T cells in Salmonella infection, we examined the resolution of an intraperitoneal infection with avirulent Salmonella choleraesuis 31N-1 in mice lacking T-cell-receptor (TCR) alphabeta T cells by disruption of the TCRbeta chain gene (TCRbeta(-/-)). The bacteria in TCRbeta(-/-) mice decreased with kinetics similar to that seen in control mice (TCRbeta(+/+)) after infection. The number of natural killer (NK) cells in the peritoneal cavity increased on day 6 after infection and thereafter decreased in both TCRbeta(-/-) and TCRbeta(+/+) mice, whereas the number of gammadelta T cells, in place of alphabeta T cells, increased remarkably in the peritoneal cavity of TCRbeta(-/-) mice on day 6 after infection. The NK cells from Salmonella-infected TCRbeta(-/-) mice produced interferon-gamma (IFN-gamma) but neither interleukin-4 (IL-4) nor IL-13 in response to immobilized anti-NK1.1 monoclonal antibody (mAb). The gammadelta T cells produced IFN-gamma but neither IL-4 nor IL-13 in response to heat-killed Salmonella, whereas both IFN-gamma and IL-13 but no IL-4 was produced by the gammadelta T cells stimulated with immobilized anti-TCRgammadelta mAb. In vivo administration of anti-NK1.1 mAb inhibited the reduction of Salmonella, whereas anti-TCRgammadelta mAb treatment did not affect the bacterial growth in TCRbeta(-/-) mice after Salmonella infection. However, neutralization of endogenous IL-13 with anti-IL-13 mAb enhanced the bacterial clearance in TCRbeta( /-) mice after infection. These results suggest that NK1.1(+) cells serve mainly to protect against avirulent Salmonella infection in the absence of alphabeta T cells, whereas gammadelta T cells may play dichotomous roles in Salmonella infection through IFN-gamma and IL-13 in TCRbeta(-/-) mice. PMID- 10873310 TI - Multivariate Screening Analysis of Water-in-Oil Emulsions in High External Electric Fields as Studied by Means of Dielectric Time Domain Spectroscopy. AB - The effect of crude oil resins with various polar characters on the stability of w/o model emulsions containing asphaltenes is investigated using a mixture design. The resins were extracted using an adsorption-desorption technique. One asphaltene fraction and four different resin fractions from one European crude oil were used. The stabilities are measured using time-domain dielectric spectroscopy in high external electric field. It is found that resins with different polar character have different effects on the emulsion stability. At asphaltene/resin ratios of 1 and 5 : 3 the resins in some cases lead to an emulsion stability higher than that of a similar emulsion stabilized by asphaltenes only, while at low asphaltene/resin ratios ( approximately 1 : 3) the emulsion stability is reduced by the resins. The effect on emulsion stability of combining two different resin fractions depended on the resin types combined as well as the relative amount of resins and asphaltenes. Also, an increase in the stability of some of the emulsions containing resins and asphaltenes for a period of 50-300 min after the emulsification was observed. This time-dependence of emulsion stability is attributed to the mobility of resins at the oil-water interface and the slow buildup of a stabilizing interfacial film consisting of resins and asphaltenes. Copyright 2000 Academic Press. PMID- 10873311 TI - Hydrophobic Flocculation of Galena Fines in Aqueous Suspensions. AB - The hydrophobic flocculation of galena fines induced by potassium amyl xanthate (PAX) in aqueous suspensions has been studied using laser diffraction, electrophoretic light scattering, contact angle, and microflotation measurements. The measurements were performed on <2 um, 2-5 um, 5-10 um, and <30 um size galena by varying several parameters, including PAX concentration, pH, original particle size, kerosene concentration, and suspension stirring. The experimental results have demonstrated that the hydrophobic flocculation was closely correlated with the particle hydrophobicity, but was not lowered upon increasing the particle surface charges due to PAX adsorption, which is contrary to the DLVO theory. This flocculation has been observed to increase with a reduction of the original particle size and an increase in kerosene concentration, and to require sufficient stirring strength and magnitudes of kinetic energy input to achieve the maximum aggregation degree. From the microflotation results, it has been found that the flotation response of galena fines is markedly improved due to the formation of hydrophobic flocs, suggesting that floc flotation is a promising means to recover galena in the fine size range. Copyright 2000 Academic Press. PMID- 10873312 TI - A New Method for the Characterization of Electrically Conducting Liquid Bridges. AB - An electrical conductance technique is employed in investigating the behavior of constant volume liquid bridges when their length is altered. The liquid bridges are edge-pinned between two vertical, identical rods with a variable separation distance. Rods of different radius, material, and edge geometry are examined as they play a role in the response of the system. It is shown that liquid bridge volume and rod radius are the parameters that mainly influence the conductance signal. A mathematical framework is developed for the identification of the geometrical characteristics of liquid bridges explicitly from conductance data. The role of gravity is discussed in both the experiments and the theoretical analysis. The theoretical predictions obtained show a close agreement with measurements. Copyright 2000 Academic Press. PMID- 10873313 TI - Montmorillonites Modified with Carbonaceous Deposits. AB - The mechanism of formation of carbonaceous deposits on montmorillonites was studied for systems prepared by sorption of polymers on pillared montmorillonites and subsequent carbonization. Temperature-programmed desorption of ammonia and X ray photoelectron spectroscopy showed that polymer was attached to Al-OH acidic sites of smectite layers. Pillars were not influenced to a greater extent. Islands of carbonaceous material were formed, but the amount of deposited carbon depended strongly upon carbonization conditions. The hydrophylic and acidic properties of carbon-covered silica aluminas differed strongly from those of the starting material. Copyright 2000 Academic Press. PMID- 10873314 TI - Synthesis of Microporous Silica Spheres. AB - Spherical microporous silica powders with a narrow size distribution have been prepared by a precipitation technique involving the hydrolysis reaction of a silicon alkoxide in ethanol. The formation of the important microporosity has been investigated following two templating methods: the co-hydrolysis and condensation of two alkoxides, one of which presents porogen function, and the adsorption of an organic compound (glycerol) as the porogen. In both processes, the organic porogen is removed by a simple calcination. In the first method, the addition of more than 20 mol% of the porogen alkoxide, necessary for generating enough microporosity, disturbs completely the condensation process resulting in microporous, nonuniform silica particles of large size distribution. The best result has been obtained with the glycerol method where submicrometer-sized silica spheres with a very narrow size distribution and about 40 vol% porosity have been synthesized. The presence of glycerol during the synthesis considerably affects the precipitation mechanism, resulting in a larger mean particle size. The use of an aggregative growth model has successfully been employed to explain the effect of the porogen during particle formation. The precipitation mechanism of silica involves the aggregation between particles of similar size until a critical size is reached, resulting in a uniform particle size distribution. In the presence of glycerol, it has been shown that a second aggregative growth between still-nucleating primary particles and large particles occurred with increasing reaction time. This second aggregative growth appears at an intermediate stage of the precipitation process and is due to both the precipitation of smaller primary particles and the destabilization of the colloidal stability of the system. This explains why the final particle size reached in this system is larger compared to silica particles synthesized without glycerol and shows how glycerol is incorporated in the silica particles. The synthesis of silica microporous spheres of narrow size distribution, by varying particle size and porosity, should yield a wide range of aqueous silica slurries for particular chemical mechanical polishing applications. Copyright 2000 Academic Press. PMID- 10873315 TI - The Effect of Poly(methyl vinyl ether-alt-maleic acid) Stabilizer on the Stability of Polyaniline-Poly(methyl vinyl ether-alt-maleic acid) Dispersions. AB - The polyaniline (PANI) dispersions have been prepared in acidic aqueous media by oxidative dispersion polymerization in the presence of a polymeric stabilizer. The polymeric stabilizer used in this study is the poly(methyl vinyl ether-alt maleic acid) (PMVEMA) which contains acid groups (-COOH). The PANI-PMVEMA particles have a uniform size and a spherical shape. The PANI-PMVEMA dispersions show almost no desorption of the PMVEMA, even though the sonication at 500 W for 20 min and the centrifugation at 500 rpm for 60 min are performed 10 times. The existence of the PMVEMA on the surface is confirmed by X-ray photoelectron spectroscopy. The dispersion stability of the PANI-PMVEMA dispersions is extensively influenced by zeta potential which was governed by the acid group ( COOH) of the PMVEMA on the PANI-PMVEMA particle surface. Copyright 2000 Academic Press. PMID- 10873316 TI - Adsorption and Desorption of Triton X-100 in Polystyrene Particles with Different Functionality. AB - A complete adsorption study of Triton X-100 onto latexes with different functionality is presented. Three surfactant-free polystyrene latices with different chemical surface groups (sulfate, carboxyl, and amidine) were prepared. The pH of the liquid phase controls the surface charge, and hence this factor can be considered an important variable in this study. The adsorption isotherms show that an increase in the surface charge yields a decrease in the amount of adsorbed surfactant. A descriptive mechanism of the adsorption process which considers the presence of holes in the layer of adsorbed surfactant is presented for explaining the experimental results. On the other hand, the adsorption isotherms have been analyzed with two classical adsorption theories, those of Langmuir and Kronberg et al. Both theories give a good description of the results, but the latter offers more information on the adsorption phenomena. Copyright 2000 Academic Press. PMID- 10873317 TI - Adsorption and Desorption of Triton X-100 in Polystyrene Particles with Different Functionality. AB - An experimental desorption study of Triton X-100 from latices with different functionality is presented. Two different strategies to follow the desorption experiment have been used: first, a discontinuous method based on the wash step (centrifugation-removal-redispersion); and second, a continuous method based on the replacement of the disperse media. The desorption results show that in all cases a residual amount of Triton X-100 remains adsorbed, indicating irreversibility of the surfactant adsorption. Finally, the effect of desorption on the colloidal stability of one type of latex-surfactant complex has been analyzed. Copyright 2000 Academic Press. PMID- 10873318 TI - Adsorption and Exchange Dynamics in Aging Hydroxyethylcellulose Layers on Silica. AB - The adsorption kinetics of hydroxyethylcellulose (HEC) on silica and relaxations in adsorbed HEC layers were probed using total internal reflectance fluorescence and near-Brewster reflectivity. Like many random-coil polymers, HEC was found to adsorb at the transport-limited rate. Relaxations occurred at nearly constant interfacial mass when HEC layers were exposed to aqueous solvent, causing the subsequent exchange of chains between the layer and the free solution to become increasingly hindered. Eventually, on the time scale of a day, layers became immobilized and unable to accommodate chains from free solution. A continued fluorescence decay, beyond time scales that could be probed with self exchange, suggested further relaxations of the adsorbed HEC. The polydisperse HEC system (with an average molecular weight near 450,000) behaved qualitatively similar to molecular weight standard polyethylene oxide (PEO) layers on silica. For instance, relaxations in PEO layers occurred on a time scale of 10-20 h, like the HEC layers. Young layers of the latter, however, exhibited self-exchange kinetics that were an order of magnitude slower than PEO layers of similar age. This difference in adsorbed layer dynamics was attributed to HEC's stiffer backbone, compared with flexible PEO. Copyright 2000 Academic Press. PMID- 10873319 TI - Electric Potential (psi(delta) and psi(d)) at Outer Helmholtz Plane and Midplane on the Clay Colloid Surface with Overlapping Flat Double Layers. AB - An anion negative adsorption equation in the condensed colloidal suspension with overlapping flat double layers was derived according to Gouy-Chapman theory. The electric potential at the outer Helmholtz plane (OHP), psi(delta), and the electric potential at the midplane, psi(d), were numerically solved by computer using the anion negative adsorption equation on the basis of experiments. The results showed that psi(delta) and psi(d) increase with the decrease of the distance between two clay plates, lambda, at first in the given electrolyte concentration. When lambda is smaller than 50-70 A, psi(d) remains almost unchanged while psi(delta) declines remarkably with the further decrease of lambda. The change of psi(d)/psi(delta) with lambda can explain and manifest overlapping degree of flat double layers more appropriately than psi(d) in previous works. Due to compression of the flat double layer on the clay colloid surface at increasing electrolyte concentration, the magnitude of the electrical potentials at OHP and midplane is considerably reduced at a given lambda. Copyright 2000 Academic Press. PMID- 10873320 TI - Effect of Added Salts or Polyols on the Cloud Point and the Liquid-Crystalline Structures of Polyoxyethylene-Modified Silicone. AB - The effect of added salts (NaCl, Na(2)SO(4), and NaSCN) or polyols (glycerin (Gly), 1,3-butanediol (1,3-BD), ethylene glycol (EG), and polyethylene glycol (PEG400)) on the hexagonal liquid-crystalline structure of polyoxyethylene modified silicone was investigated by means of small angle X-ray scattering (SAXS). The effective cross-sectional area of the lipophilic part of the aggregate, a(s), in the hexagonal phase decreases upon the addition of salts, on one hand, lowering the cloud point in the dilute aqueous siloxane surfactant solutions. On the other hand, if added salt raises the cloud point, the a(s) increases. Similar results were obtained in the case of the addition of polyols. Since the a(s) mainly depends on the EO chain length, the above results are direct evidence that the hydration or dehydration of the EO chain is affected by these additives. The static fluorescence probe method was applied to the Gly and 1,3-BD systems using 8-anilino-1-naphthalene-sulfonic acid, ANS, to know the change in hydration of the EO chains. In the Gly system, the hydration of the EO chain monotonically decreases whereas 1,3-BD first increases the hydration and then decreases it at high 1,3-BD content. These results are very consistent with the SAXS and cloud temperature results. Copyright 2000 Academic Press. PMID- 10873321 TI - A Calorimetric Study of the Influence of Temperature on the Self-Association of Amphiphilic Antidepressant Drugs in Aqueous Solution. AB - Relative apparent molar enthalpies have been determined as a function of concentration (0.0001 to 0.2 mol kg(-1)) by heat conduction calorimetry for aqueous solutions of the structurally related antidepressant drugs imipramine and clomipramine in water over the temperature range 288 to 308 K. Critical concentrations determined from inflections in these plots for both drugs had minimum values at 298 K. The concentration dependence of the relative apparent molar enthalpy could be quantitatively described using a mass action model of association based on the Guggenheim equations for the activity coefficients of mixed electrolytes. Derived values of the monomer-counterion interaction coefficient for imipramine became increasingly negative with an increase in temperature over the temperature range 293 to 303 K, indicative of an increasing degree of premicellar association. In contrast, negative monomer-counterion interaction coefficients were obtained for clomipramine at only 303 and 308 K, suggesting an absence of premicellar association at lower temperatures. Values derived for the molar enthalpy of micellization of both drugs from the mass action model indicate an increasingly exothermic process with increase in temperature; positive values at 288 and 293 K arise from hydrophobic interactions while the negative values at higher temperatures suggest the importance of London dispersion interactions as the major driving force for aggregation. Copyright 2000 Academic Press. PMID- 10873322 TI - On the Measurement of Phase Transition Temperatures in Physical Molecular Organogels. AB - Methods of measurement of the phase transition temperatures in physical molecular gels are analyzed. The "falling ball" technique, NMR, and rheology are compared. Temperature versus concentration phase diagrams can be easily obtained, and the reliability and workability of the methods are discussed. It appears that rheology is the more accurate and convenient technique while the falling ball method provides acceptable DeltaH, DeltaS values, but the melting temperature values are significantly altered. With NMR, kinetic information associated with the molecular aggregation process can also be extracted. Copyright 2000 Academic Press. PMID- 10873323 TI - Dewetting Revisited: New Asymptotics of the Film Stability Diagram and the Metastable Regime of Nucleation and Growth of Dry Zones. AB - Stability properties of a nonwetting film are discussed. Assuming a general form of the disjoining pressure, accurate asymptotic formulas for the upper thickness range of the film instability/metastability are derived. This analysis is applied to two particular cases: a nonionic liquid film with the (m, n) power form of the disjoining pressure and an ionic liquid film with an exponentially decaying electrostatic part of the disjoining pressure. The metastable regime of dewetting is considered, and an expression for the critical radius of a hole is derived. A new Fokker-Planck kinetic model of metastable dewetting, applicable at early stages of the process, is developed. It yields a relationship between the number of viable holes (per unit area and unit time) moving in steady-state regime to the supercritical part of the "embryo size space" and the equilibrium number of "critical" holes determined from thermodynamics. The dynamics of metastable dewetting is quantitatively described in terms of the surface fraction of holes in the film. Continuous dynamic models of the metastable dewetting applicable in the entire range of times have to include the thermal noise, as proposed by V. S. Mitlin (1994, Colloids Surf. A 89, 97). Copyright 2000 Academic Press. PMID- 10873324 TI - Static and Dynamic Wetting Behavior of Triglycerides on Solid Surfaces. AB - Triglyceride wetting properties on solid surfaces of different hydro-phobicities were investigated using three different methods, namely, the sessile drop method for static contact angle measurements, the Wilhelmy method for dynamic contact angle measurements, and the captive bubble method to investigate thin triglyceride film stability. For solid surfaces having a surface free energy higher than the surface tension of triglycerides (tributyrin, tricaprylin, and triolein), a qualitative correlation was observed between wetting and solid/triglyceride relative hydrophobicities. On surfaces presenting extreme hydrophobic or hydrophilic properties, medium-chain triglycerides had a behavior similar to that of long-chain unsaturated ones. On a high-energy surface (glass), tricaprylin showed an autophobic effect subsequent to molecular adsorption in trident conformation on the solid, observed with the three methods. Thin triglyceride films between an air bubble and a solid surface were stable for a short time, for solids with a surface free energy larger than the triglyceride surface tension. If the solid surface had a lower surface free energy, the thin film collapsed after a time interval which increased with triglyceride viscosity. Copyright 2000 Academic Press. PMID- 10873325 TI - Flocculation and Coalescence of Oil-in-Water Poly(dimethylsiloxane) Emulsions. AB - The stability of poly(dimethylsiloxane) (PDMS) oil-in-water emulsions has been investigated in the presence of added NaCl as well as in the presence of added surfactant. The emulsions were prepared using a combination of nonionic (C(x)E(y), x and y represent the number of methylene (C) and ethylene oxide (E) groups, respectively) and cationic (quarternary alkylammonium) surfactants. The droplets were observed to exhibit weak flocculation in the presence of high NaCl concentration (1 M). Phase separation and optical microscopic observations revealed that the principal mechanism for emulsion destabilization at high salt concentration was coalescence, which was accelerated at elevated temperature (50 degrees C). The effective coalescence rate for diluted emulsions was investigated using photon correlation spectroscopy. The small effective Hamaker constant for PDMS is the primary reason for the slow rate of coalescence observed for the emulsions at neutral pH in the presence of NaCl. The stability of PDMS emulsions to flocculation is qualitatively similar to that reported for low Hamaker constant dispersions (e.g., microgel particles). Addition of cationic surfactants (cetyltrimethylammonium chloride and dodecyl dimethylbenzylammonium chloride) to the negatively charged droplets after preparation was shown to decrease the emulsion stability once the surfactant concentration exceeded the CMC. Electrophoretic mobility measurements showed that added cationic surfactant changed the sign of the droplet charge from negative to positive at concentrations well below the CMC. Charged micelles of the same sign as the droplets are electrostatically excluded from close approach to the droplet surface within a distance (varepsilon) which results in depletion flocculation. Copyright 2000 Academic Press. PMID- 10873326 TI - Effect of the Supporting Electrolyte on the Adsorption of Octanoic Acid at the Mercury/Electrolyte Interface. AB - The adsorption and the changes in the interfacial composition of octanoic acid at the mercury/electrolyte interface was studied by measuring the differential capacitance at different concentrations of the supporting electrolyte, at various supporting electrolyte systems and at various temperatures. The adsorption was followed by means of capacity-potential curves in the short-term region and capacity-time transients in the long-term region at selected potentials, in all the potential ranges. A decrease of the capacitance with time was observed in most cases, followed either by a constant capacitance value or by its increase. In the short-term region, anion-surfactant complexes are formed, where the anions act as bridges between the perpendicularly oriented surfactant molecules. The larger is the negative charge of the anion, the more negative will be the charge of the anion-surfactant complex leading to a shift of the potential of maximal adsorption to more positive values. The formation of metastable condensed films is best when the hydration of the anion and its size are not too large. In the long-term region the observed increase of the capacity with time can be explained as an exchange of the metastable condensed film by a hemimicellar surface state. Here, the anions act as cores of the hemimicelles, and the hydrophilic acid groups of the amphiphiles contact the solution. Two contrary effects determine the formation of the hemimicelles. The greater is the specific adsorption of the anions, the larger is the formation of hemimicelles and the increase of the capacity. With an increase in the ability of the anions to break the water structure (lyotropic or Hofmeister series), the formation of hemimicelles will be decreased. Copyright 2000 Academic Press. PMID- 10873327 TI - Properties of Poly(N-isopropylacrylamide)-Grafted Colloidal Silica. AB - Poly(N-isopropylacrylamide-co-3-trimethoxysilylpropyl methacrylate) was prepared by radical polymerization and was grafted onto the surface of spherical colloidal silica. The copolymer, which had on average 1 silyl group per chain, condensed on the silica dispersed in THF at 60 degrees C. The resulting particles had a critical coagulation concentration of calcium chloride of 500 mM at room temperature, indicating robust colloidal stability. By contrast, heating the suspension to 50 degrees C lowered the critical coagulation concentration by more than three orders of magnitude, giving a value of 0.1 mM. Thus, the PNIPAM shell induced temperature-sensitive colloidal stability in the silica dispersion. The coated silica particles were also surface active. However, the surface tension of 50 mJ/m(2) at 25 degrees C is 15% higher than the corresponding value for PNIPAM solutions. Copyright 2000 Academic Press. PMID- 10873328 TI - On the Behavior of Nonionic Surfactants at the N-Heptane/Silica Gel Interface: Influence of the Presence of Interfacial Water Inferred from Adsorption Isotherms and Calorimetric Data. AB - The behavior of two polydisperse nonionic surfactants, poly (oxyethylene) glycol alkylphenyl ether TX-35 and TX-100, at the prewetted silica gel/n-heptane and dried silica gel/n-heptane interfaces has been compared by the determination of the average adsorption isotherms of the polydisperse surfactants and of displacement enthalpies. From HPLC experiments, we could also separately quantify the adsorption of each ethyleneoxide (EO) fractions for silica gel from the polydisperse surfactant solution. The adsorption isotherms clearly indicate an incomplete preferential adsorption of the large (EO) chains over the small ones, as well on dried silica gel as on a prehydrated sample. This preferential adsorption and its driving force follow the solubility rules of the poly(oxyethylene) glycol alkylphenyl ether in an apolar solvent and support the idea of a solubility-limited adsorption: solubility in organic solvents of the smaller (EO) chains is much more significant than that of the longer ones and hence prevents adsorption of the smaller species. Consequently, it is observed that the presence of interfacial water decreases the affinity of TX-35 molecules for the hydrophilic silica surface due to the hydration of (EO) chains. In contrast, for TX-100 adsorption after the prewetting treatment the clearest trend is a drastic increase of the adsorption ascribed to the additional solubilization (and micellization) of the TX-100 molecules in the interfacial aqueous phase. The differential molar enthalpies of displacement show a change in the adsorption mechanism, depending on the presence of molecular water on the surface. In the initial part of the adsorption isotherm, a prevailing exothermic process is obtained with prehydrated silica and suggests that hydration of the polar heads of TX-35 and the solubilization of the TX-35 in interfacial water are occurring. For higher equilibrium concentrations, the enthalpies of displacement observed with the prehydrated adsorbent become slightly lower than those obtained with dry silica gel. It may be that this difference is due to the micellization phenomenon of the surfactant species with longer EO chains in interfacial water. These features emphasize the influence of interfacial water on the adsorption of EO fractions from organic solvent. Copyright 2000 Academic Press. PMID- 10873329 TI - Preparation and Characterization of a Phospholipid Membrane-Bound Tetrapeptide That Corresponds to the C-Terminus of the Gastrin/Cholecystokinin Hormone Family. AB - The present work deals with the synthesis of a hydrophobized peptide and its localization at the membrane surface, after its incorporation into phospholipid vesicles. The tetrapeptide, Trp-Met-Asp-Phe-NH(2), which corresponds to the C terminus of the cholecystokinin/gastrin hormone family, is conjugated to N glutaryldioleoylphosphatidylethanolamine using a carbodiimide-catalyzed reaction method. Sonication of the lipophilized hormone in the presence of dimyristoylphosphatidylcholine results in a strong sequestration of the conjugate in the artificial membrane structures that are formed. More detailed information on the localization of the peptide moiety with respect to the membrane surface is gathered from fluorescence measurements. Both the observed blue shift in the fluorescence spectra and the quenching of Trp emission in the presence of potassium iodide point to a partial screening of the hormone moiety from the surrounding aqueous phase. The different parameters that may influence the physicochemical behavior of a hydrophobized peptide in a membrane structure are briefly discussed Copyright 2000 Academic Press. PMID- 10873330 TI - Physicochemical Properties of Dodecylammonium Picrate. AB - A novel surfactant, dodecylammonium picrate (DDAP), was synthesized and its crystal structure was determined by X-ray diffraction analysis. DDAP's physicochemical properties were examined by spectral (infrared and nuclear magnetic resonance), thermal, microscopic, and conductometric studies. The results revealed the influence of counterion specificity on thermal solid-state transitions and solution properties: the Krafft point, the aqueous solubility, the critical micelle concentration, the degree of counterion binding, and thermodynamic parameters of micellization. Copyright 2000 Academic Press. PMID- 10873331 TI - Mechanism of Adsorption of Dyes and Phenols from Water Using Activated Carbons Prepared from Plum Kernels. AB - The kinetics and mechanism of adsorption of two commercial dyes (BR22, AB25), phenol, and 3-chlorophenol from water on activated carbons were studied at 30 degrees C. The activated carbons were prepared from plum kernels, and the activation temperature and time tested were in the ranges 750-900 degrees C and 1 4 h, respectively. Three simplified kinetic models including a pseudo-first order, a pseudo-second-order, and an intraparticle diffusion model were tested. It was shown that the adsorption of both phenols could be fitted to a pseudo second-order rate law, and that of both dyes could be fitted to an intraparticle diffusion model. Kinetic parameters were calculated and correlated with the physical properties of the adsorbents. Copyright 2000 Academic Press. PMID- 10873332 TI - Covalent Grafting of Ethylene Glycol into the Zn-Al-CO(3) Layered Double Hydroxide. AB - Zn-Al-CO(3) layered double hydroxide was synthesized at room temperature using a procedure reported elsewhere. After characterization, 0.5 g of the Zn-Al-CO(3) layered compound was reacted under air with 15 cm(3) of ethylene glycol at 80 degrees C for a period of 5 days. After washing with acetone and drying at 50 degrees C, the resulting white powder was characterized by X-ray powder diffraction, thermal analysis (simultaneous TG/DSC), elemental analysis (C, H, N, Al, and Zn content), and FTIR spectroscopy. All of the experimental data were consistent with the bidentade grafting of ethylene glycol into the interlayer surface of the Al-Zn double hydroxide. The stoichiometry obtained [Zn(0.66)Al(0.34)(OCH(2)CH(2)O)](OH)(0.34).0.4H(2)O] showed that all of the surface hydroxide groups were replaced by ethylene glycol through Al-(Zn)-O-C bonds. Carbonate could not be detected by FTIR, proving that OH(-) was probably the actual intercalated counteranion. Copyright 2000 Academic Press. PMID- 10873333 TI - Interfacial Behavior of beta-Lactoglobulin at a Stainless Steel Surface: An Electrochemical Impedance Spectroscopy Study. AB - The electrochemical impedance spectroscopy technique was used to investigate the interfacial behavior of beta-lactoglobulin at an austenitic stainless steel surface over the temperature range 299 to 343 K at an open circuit potential. The electrode/electrolyte interface and corresponding surface processes were successfully modeled by applying an equivalent-electrical-circuit approach. A charge-transfer resistance value was found to be very sensitive to the amount of adsorbed protein (surface concentration), thus indicating that the adsorption of the protein (i) was accompanied by the transfer of the charge, via chemisorption, and (ii) influenced the mechanism and kinetics of the corrosion reaction. This was also apparent from the large decrease in the corrosion activation energy (16 kJ mol(-1)) caused by the adsorption of the protein. Adsorption of beta lactoglobulin onto the stainless steel surface at an open circuit potential resulted in a unimodal isotherm at all the temperatures studied and the adsorption process was described with a Langmuir adsorption isotherm. From the calculated Gibbs free energies of adsorption it was confirmed that beta lactoglobulin molecules adsorb strongly onto the stainless steel surface. The enthalpy and entropy values indicated that the molecule partially unfolds at the surface upon adsorption. The adsorption process was found to be entirely governed by the change in entropy. Copyright 2000 Academic Press. PMID- 10873334 TI - Roles of gamma-Globulin in the Dynamic Interfacial Behavior of Mixed Dipalmitoyl Phosphatidylcholine/gamma-Globulin Monolayers at Air/Liquid Interfaces. AB - This study investigated the roles of gamma-globulin in the dynamic interfacial behavior of dipalmitoyl phosphatidylcholine (DPPC)/gamma-globulin monolayers at air/liquid interfaces at 25 degrees C. The surface tension behavior demonstrated that gamma-globulin had a large adsorption time scale. Moreover, the surface pressure-area hysteresis behavior of adsorbed gamma-globulin monolayers suggested that no significant desorption occurred during the compression stage, and the respreading of gamma-globulin molecules at the interface during the expansion stage was slow. From the hysteresis behavior of adsorbed gamma-globulin monolayers with spread DPPC molecules, it was found that gamma-globulin molecules were expelled from the interface as DPPC molecules were in a condensed state. The squeeze-out of gamma-globulin molecules seemed to induce the loss of DPPC molecules at the interface with the extent depending on the initial gamma globulin surface concentration. Furthermore, the expelled gamma-globulin molecules re-entered the monolayer and participated in the surface pressure increase during the following expansion stage. The exclusion of gamma-globulin associated with the removal of DPPC during monolayer compression and the re-entry of gamma-globulin during subsequent monolayer expansion represented a mechanism for DPPC depletion and gamma-globulin enrichment at the interface, which may explain the inhibitory effect of certain proteins on the surface activity of DPPC. Copyright 2000 Academic Press. PMID- 10873335 TI - Characterization of the Structure and Catalytic Activity of Pt/Sepiolite Catalysts. AB - Four catalysts containing 1% w/w Pt deposited on various sepiolite supports were prepared. Two natural sepiolites and another two obtained by acid treatment of one of them were used. Both the sepiolites used as metal supports and the catalysts were characterized for structure and surface properties, using (1)H and (29)Si MAS NMR spectroscopy, and (29)Si CP/MAS NMR spectroscopy, in addition to hydrogen, pyridine, and CO(2) chemisorption measurements. The activity of the catalysts in the hydrogenation of benzene to cyclohexane and the dehydrogenation of cyclohexane to benzene was examined and the catalyst supported on the natural sepiolite called Pangel was found to be the most active of all. Copyright 2000 Academic Press. PMID- 10873336 TI - Photophysical Behavior of a New Gemini Surfactant in Neat Solvents and in Micellar Environments. AB - A novel fluorescent gemini surfactant, 1,4-bis-(2'-(N-dodecyl pyridinio-4" yl)ethenyl)benzene dibromide, abbreviated BDPEBB, has been synthesized and its photophysical properties have been studied in different environments. BDPEBB has a limited solubility in alcohols where it is found in aggregate form at concentrations>/=1 mM. In other solvents, e.g., water, it is only found in aggregate form, even at much lower concentrations. Solvent polarity has a small and insignificant solvatochromic effect but alcohols give a specific interaction with BDPEBB, causing a significant hypsochromic shift in absorption maxima and a large increase in relative fluorescence efficiency. Pyrene fluorescence is effectively quenched by BDPEBB. Pyrene also forms associative complexes with BDPEBB in water. These complexes are partly dissociated in the presence of surfactant micelles. Triton X-100 micelles provide a favorable environment for BDPEBB solubilization well distinguished from the behavior of ionic surfactants. Small quantities of BDPEBB have a large influence on the behavior of aqueous sodium dodecylsulfate (SDS) and sodium decylsulfate (SDeS) micelles, inducing the formation of large aggregates, visible by the naked eye. These large aggregates are most probably microcrystals of BDPEBB(2+)/2DS(-) or BDPEBB(2+)/2DeS(-). The aggregation number of SDS and SDeS micelles in the absence and in the presence of BDPEBB has been calculated by exploitation of the static luminescence quenching kinetics of Ru(bpy)(3)(2+) by 9-methylanthracene, both solubilized in the micellar phase. It has been observed that Ru(bpy)(3)(2+) inhibits the precipitation of SDeS micelles in the presence of BDPEBB. Our results suggest that double-chain surfactant chromophores should be employed with particular care if they are to be used as probes of the micellar phase. Copyright 2000 Academic Press. PMID- 10873337 TI - The Normalization of the Micropore-Size Distribution Function in the Polanyi Dubinin Type of Adsorption Isotherm Equations. AB - The problem of the normalization of the micropore-size distribution (MSD) based on the gamma-type function is presented. Three cases of the integration range (widely known in the literature) of MSD, characterizing the geometric heterogeneity of a solid, are considered (val( identical withB, E(0), and/or x)) i.e., from zero to infinity, from val(min) to infinity, and the finite range from val(min) up to val(max)-due to the boundary setting of an adsorbate-adsorbent system. The physical meaning of the parameters of the gamma-type function (rho and nu) is investigated for the mentioned intervals. The behavior and properties of this MSD function are analyzed and compared with the fractal MSD proposed by Pfeifer and Avnir. The general conclusion is that if adsorption proceeds by a micropore filling mechanism and the structural heterogeneity is described in the finite region (val(min), val(max)), for all cases of the possible values of the parameters of the MSD functions, the generated isotherms belong to the first class of the IUPAC classification (i.e., Langmuir-type behavior is observed). For the other cases (valin<0, infinity) and valin> satisfaction with the operations. RESULTS: The women found it difficult to translate the genetic information transmitted to them, although they were satisfied by the way it was given. At some stage during the pre operative and post-operative period nearly all women stated that they lacked psychological support from the different caregivers. No woman regretted her choice to undergo PM and IBR. By far, the most important issue was the actual risk reduction. However, the result exceeded all patients' initial expectations. When performing PM and IBR, a multidisciplinary team approach, including a psychologist, seems mandatory. It will facilitate the overall management of this group of women. PMID- 10873355 TI - Indicators of loco-regional recurrence in breast cancer. The South East Swedish Breast Cancer Group. AB - AIM: The aim of the investigation was to contribute to the identification of patients who have increased or decreased risk of loco-regional recurrence. METHODS: Six hundred and twenty-nine consecutive patients with primary breast cancer diagnosed between 1988 and 1990 were studied. Two-thirds of the patients underwent mastectomy. Radiotherapy was administered if patients were node positive or breast conserved. The Nottingham histological grading protocol was used and presence of lymphovascular invasion was assessed. Investigated parameters were: age, size, grade, steroid receptor content, surgical radicality, vascular invasion and nodal status. Statistically significant risk factors for loco-regional recurrence using univariate or Cox proportional hazard analysis were grade and lymphovascular invasion. RESULTS: Women with grade 1-2, node negative tumours without vascular invasion had a very low loco-regional recurrence rate-3.1%. Seventeen percent of patients with grade 3 tumours and vessel invasion had loco-regional recurrence. CONCLUSIONS: Our findings, and those of others, indicate that the use of adjuvant radiotherapy should be influenced to a greater extent by grade and lymphovascular invasion. PMID- 10873356 TI - Antibiotic prophylaxis for post-operative wound infection in clean elective breast surgery. AB - Antibiotic prophylaxis has been used to good effect in the prevention of post operative wound infections in patients undergoing gastrointestinal operations. We have assessed the use of a single dose of intravenous antibiotic (Augmentin 1.2 g), given with induction of anaesthesia as prophylaxis, against post-operative wound infection in women undergoing clean, elective breast surgery. Three hundred and thirty-four patients were recruited. Of the 164 receiving antibiotic prophylaxis 29 (17.7%) had wound infections compared with 32 (18.8%) in the placebo group (P=0.79). There were no significant differences in any other post operative infective complications. Antibiotic prophylaxis is probably not required in clean, elective breast surgery. PMID- 10873357 TI - Envelope mastectomy with immediate reconstruction (EMIR). AB - AIMS: To develop an oncologically safe and aesthetically acceptable technique for mastectomy, using a muscle flap and tissue expander through one incision. METHODS: Twelve consecutive patients (mean age 40) underwent an envelope mastectomy (skin and nipple sparing), with immediate reconstruction with a latissimus dorsi muscle flap and tissue expander. Assessment of cosmesis was by review of pre- and post-operative photographs by an independent observer. RESULTS: During follow-up there have been no recurrences. Assessment of cosmesis gave a score of 44 out of 48 (92%). One prosthesis was removed due to erosion of the prosthesis through the skin. CONCLUSIONS: In patients with large lesions, multi-focal lesions (both invasive and in situ) and recurrent phyllodes tumours may undergo an oncologically safe mastectomy with immediate reconstruction through a single incision that is inconspicuous being in the mid-axillary line. Although follow up is only 8.5 months, long-term studies are being undertaken. PMID- 10873358 TI - Surgical treatment of gastrointestinal carcinomas in octogenarians: risk factors for complications and long-term outcome. AB - BACKGROUND: The aims of this retrospective study were to determine the factors predictive of morbidity and mortality, and to evaluate the probability of long term survival in octogenarians with carcinomas of the gastrointestinal tract. PATIENTS AND METHODS: Out of a total of 194 patients, aged 80 years or over, with histologically diagnosed carcinoma of the stomach or colon-rectum, observed between 1987 and 1995, 167 underwent surgery and were included in this study. The relationship between a series of clinico-pathological variables and morbidity/mortality rates was investigated by univariate and multivariate analysis. Complete follow-up data were available in 161 patients. RESULTS: Fifty nine patients (35.3%) experienced complications and 14 (8.4%) died during hospitalization. Statistical analysis identified hypoalbuminaemia (P<0.01, relative risk (RR)=2.92) and hypercreatininaemia (P<0.05, RR=3.59) as independent predictors of post-operative complications. Hypercreatininaemia (P<0.05, RR=5.22) and non-curative surgery (P<0. 05, RR=3.99) significantly affected operative mortality. Crude 5-year survival rate, including operative mortality after curative surgery, was 41% in gastric cancer and 39% in colorectal cancer patients. CONCLUSION: These results indicate that surgery for gastrointestinal carcinomas yields an acceptable operative risk in octogenarians, and provides good long-term results if oncological radicality can be obtained. Pre-operative evaluation of tumour stage and patient's general condition is useful to identify subgroups of patients at high risk of surgical complications and mortality. PMID- 10873359 TI - The progression of invasiveness regarding the role of transforming growth factor beta receptor type II in gastric cancer. AB - AIMS: Transforming growth factor beta (TGF beta) is a potent growth inhibitor of epithelial cells. The expression of TGF beta receptors is required for the effect of TGF beta. In this study, we used immunohistochemistry to demonstrate the roles of the expression of TGF beta type I (T beta R-I) and type II (T beta R-II) receptors in the progression of gastric carcinoma. METHODS: To evaluate the potential prognostic value of T beta R-I and T beta R-II, 158 consecutive gastric cancer tissues specimens obtained over a 3-year period were examined. RESULTS: A total of 50 (32%) and 28 (18%) patients had T beta R-I(+) and T beta R-II(+), respectively. The 5-year survival rates of the patients with T beta R-I(+) and those with T beta R-I(-) were 74% and 71%, respectively. In contrast, the 5-year survival rates of the patients with T beta R-II(+) and those with T beta R-II(-) were 57% and 75%, respectively, and the difference was statistically significant (P<0.05). The extent of T beta R-II was closely correlated to the macroscopic types based on the Borrmann classification (P<0.01), and curability (P<0.05). However, a significant difference between the 5-year survival rates of the patients with T beta R-II(+) and those with T beta R-II(-) was only obtained in advanced cases (P<0.05) not in either curative cases, non-curative cases, or early cases. CONCLUSIONS: Our data suggest that when T beta R-II expression correlates with the progression of invasiveness in gastric cancer, it may lead to a non-curative resection and a poor prognosis. PMID- 10873360 TI - Significance of duplex/colour Doppler sonography in hepatic arterial chemotherapy for patients with liver metastases from colorectal carcinoma. AB - AIMS: The aim of the study was to evaluate the importance of duplex/colour Doppler ultrasound in a protocol of hepatic regional chemotherapy, by measuring the blood flow in the hepatic artery, portal vein, hepatic veins, and inferior caval vein of patients with unresectable liver metastases from colorectal carcinoma. METHODS: Thirty-nine consecutive subjects were selected for this study, including 21 patients who had unresectable histologically confirmed liver metastases from colorectal carcinoma (Group A), and 18 asymptomatic volunteers as normal controls (Group B). All subjects of Groups A and B were examined using duplex/colour Doppler sonography. After the ultrasound study, all patients of Group A were submitted to the administration of high dose mitomycin C into the hepatic artery, with concomitant detoxication of post-hepatic venous blood. RESULTS: The mean value of the hepatic artery blood flow was significantly higher (P=0.0009) in liver metastases patients (361 ml/min, SEM=24 ml/min) than in normal controls (245 ml/min, SEM=20 ml/min). Also, the mean Doppler perfusion index was higher in liver metastases patients than in normal controls. For each patient of Group A, the total dose of mitomycin C to be infused was calculated based on the blood flow in the hepatic artery. In this way the concentration of mitomycin C in the hepatic artery was always greater than 3 microg/ml. The duration of detoxication was calculated based on the blood flow in the inferior caval vein. For two patients only, the blood flow was lower than 1000 ml/min, with the necessity to protract the duration of detoxication over 2 hours. CONCLUSIONS: The measurement of the blood flow in hepatic vessels of patients with liver metastases seems to be very important in establishing the total dose of drug which has to be infused in hepatic arterial chemotherapy, and to determine the duration of concomitant detoxication of post-hepatic venous blood. PMID- 10873361 TI - Factors affecting survival and long-term outcome in the cirrhotic patient undergoing hepatic resection for hepatocellular carcinoma. AB - AIMS: Prognostic analysis of hepatocellular carcinoma (HCC) in the cirrhotic patient undergoing hepatic resection is necessary in order to determine the clinical effect of hepatectomy on prognosis. PATIENTS AND METHODS: Univariate and multivariate retrospective analyses were performed in 51 cirrhotic patients (38 men, 13 women; mean age 65 years, range 43-81 years) with supervening HCC undergoing hepatic resection between January 1993 and December 1997. RESULTS: Segmental liver resection was performed in 39 patients (76%) with non-anatomical (wedge) resections in the remainder of cases. The post-operative mortality rate was 8%. The tumours recurred in 23 patients (45%), with 12 patients (52% of recurrences) recurring within 1 year of surgery and 22 patients (96% of recurrences) within 3 years. Recurrent disease was most frequently intrahepatic (22 patients). Significant risk factors for recurrence were micro/macro vascular invasion, and symptoms. CONCLUSIONS: The recurrence rate of hepatocellular carcinoma in patients with cirrhosis undergoing surgical resection alone is high and actuarial survival at 4 years is low. Other approaches to the treatment of hepatocellular carcinoma in patients with cirrhosis require consideration. PMID- 10873362 TI - Description of an intraperitoneal tumour xenograft survival model in the pig. AB - AIMS: Experimental animal studies are necessary if the results of minimally invasive oncological surgery are to be improved. In particular the influence of surgical technique on tumour implantation needs further assessment. Small animals such as rodents are inappropriate for such laparoscopic surgical studies. There is a requirement for another animal tumour model with animals greater in size. METHODS: Accordingly we developed an intraperitoneal tumour xenograft survival model using the domesticated pig. After creating a 12 mmHg pneumoperitoneum, 10(7)human HeLa cells were injected into the peritoneal cavity of nine non syngeneic animals to induce tumour xenograft. Resection of the sigmoid colon using four trocars and a transanal double-stapling technique was performed. The mean operating time was 69 min. No signs of post-operative pain symptoms were observed, and all the animals survived the procedure and gained weight. After 4 weeks, the animals were sacrified and all incision sites and anastomoses were excised. RESULTS: Immunohistochemical staining with antihuman pancytokeratin antibodies confirmed tumour implants in 25 out of 36 port-sites (63.8%). No peritoneal carcinosis nor tumour implants at anastomosis sites were observed. CONCLUSION: This intraperitoneal xenograft tumour model in the pig can be applied in survival studies to check the quality of surgical techniques and its influence on tumour implantation following laparoscopy for cancer. PMID- 10873363 TI - Internal hemipelvectomy for bone sarcomas in children and young adults: surgical considerations. AB - AIMS: Pelvic bone sarcomas in children and young adults are rare, and associated with a poor prognosis and a high rate of local recurrence. Primary goals of treatment include prevention of local recurrence and distant metastases. A secondary goal is maintenance of quality of life by avoiding major amputative surgery. This is why internal hemipelvectomy (a limb-sparing surgery) is advocated whenever possible. The focus of our presentation is surgical issues in the context of resection and reconstruction of the pelvis in the first two decades of life. MATERIALS AND METHODS: Between January 1988 and June 1998, 27 patients were treated and operated on (follow-up time 1.5-12 years). There were 17 males and 10 females. Their age ranged between 2 and 22 years. There were 24 patients with Ewing's sarcoma (ES) and three with other bone sarcomas. In 19 patients the tumour involved the entire or part of the iliac bone (in some cases with extension to the sacrum). In five patients the tumour involved the pubis and/or ischium. In three patients the tumour involved the sacrum with some extension to the posterior iliac bone. All patients received neoadjuvant and adjuvant chemotherapy and radiotherapy with different protocols (related to the origin of referral). RESULTS: Twenty-seven patients underwent internal hemipelvectomy. According to Enneking's classification there were: type I-10; type II-one; type III-six; type IV-five (including one localized sacrectomy); type I+IV-five patients. In 15 patients some kind of reconstruction was needed and in 12 no reconstruction was done. Four wound infections occurred that were managed successfully by surgical debridement, antibiotics and local wound care. In one case removal of the 'implant' was needed. No primary or secondary amputations were performed in the series. The rate of local recurrence was 22%. Functional status at the last follow-up visit or before death, according to the AMSTS functional rating system: excellent-six; good-17; fair-three and poor-one. All patients except the one poor result maintained their walking ability during the follow-up time. CONCLUSIONS: Internal hemipelvectomy is achievable in most cases and justified for better quality of life in children, adolescents and young adults with sarcomas. Further efforts are needed to improve the reconstructive options in the pelvis. PMID- 10873364 TI - Cell production rates in human tissues and tumours and their significance. Part II: clinical data. AB - This paper reviews the available data for cell production rates of human tissues and tumours, measured in vivo using halogenated pyrimidine labelling and laser cytometry. The technique has now been widely evaluated, and we draw general inferences from the proliferative data over a broad range of tumour and tissue types. Estimates of the S-phase duration, the time taken for DNA synthesis in cycling cells, are consistent over a narrow range with a median value of around 10 hours, notwithstanding the constraints of the experimental and statistical technique, in normal tissues and tumours. This suggests that Ts values may be a species-specific constant. The more easily measured labelled S-phase fraction, or labelling index, shows much greater intra and intertumour variation within any one tumour class. It may thus be a surrogate for time dependent measurements to a first order approximation. The cell production rate, described by the potential doubling time (Tpot), is remarkably rapid in most tumours, a median value of the order of 5 days, and much faster than clinical volume doubling times for most lesions. The rapid cell production rates in normal tissues and tumours highlight the importance of cell loss in the growth and modelling of biological structures. Cell production rate measurements do not adequately describe the biological aggressiveness of tumours. They may be used to refine adjuvant strategies for radiotherapy and chemotherapy in experimental research. Dynamic halogenated pyrimidine labelling has provided unique and valuable insights into the living biology of human tissues and tumours. PMID- 10873365 TI - Tumour immunotherapy: the adjuvant treatment of the 21st century? AB - In the course of a century, tumour immunology has revealed a picture of a very complex immune system involving the recognition and eradication of malignancies. Many tumours evade the immune system, and understanding of tumour escape mechanisms is the key to a successful immunotherapy for cancer. A wide array of tumour immunotherapy modalities have been developed, many of which have reached the phase of clinical trials, with some satisfactory results. Based on the available clinical, data and the techniques available for further improvement, we analyse the prospects for the different treatment modalities, and predict an important role for tumour immunotherapy in the near future. PMID- 10873366 TI - Secondary achalasia due to a mesenchymal tumour of the oesophagus. AB - A rare case of a secondary achalasia or pseudoachalasia due to a mesenchymal tumour of the oesophagus is presented. A 67-year-old Caucasian man had symptoms including dysphagia, odynophagia, and weight loss for 8 months. Radiological examination revealed no signs of neoplasia but an exploratory laparotomy revealed the presence of a mesenchymal tumour of the oesophagus. Tucker's criteria constitute an important tool in the differential diagnosis of secondary achalasia from primary achalasia with clinical value, but in this case, exploratory laparotomy rather than non-invasive diagnostic procedures provided the final and definite diagnosis. PMID- 10873367 TI - Ampullary carcinoid and jejunal stromal tumour associated with von Recklinghausen's disease presenting as gastrointestinal bleeding and jaundice. AB - We report a very rare case of a 36-year-old woman with von Recklinghausen's disease, synchronous carcinoid of the ampulla of Vater and stromal tumour of the jejunum, who presented with gastrointestinal bleeding and jaundice. PMID- 10873368 TI - Primary non-Hodgkin's lymphoma of the female breast masquerading as a breast abscess. AB - We report the case of a primary non-Hodgkin's lymphoma of the breast, masquerading as a breast abscess. PMID- 10873369 TI - Mammary hibernoma. AB - Hibernomas are rare benign tumours of brown fat. We report the first account in the English literature of a mammary hibernoma, presenting in the post-lactational period. PMID- 10873370 TI - Anaphylaxis to patient blue during sentinel node biopsy. PMID- 10873372 TI - A Policy on Reporting of Effect Sizes. PMID- 10873371 TI - Implementation of the sentinel node biopsy: a survey among surgeons in the Netherlands. PMID- 10873373 TI - Measures of Effect Size for Comparative Studies: Applications, Interpretations, and Limitations. AB - Although dissatisfaction with the limitations associated with tests for statistical significance has been growing for several decades, applied researchers have continued to rely almost exclusively on these indicators of effect when reporting their findings. To encourage an increased use of alternative measures of effect, the present paper discusses several measures of effect size that might be used in group comparison studies involving univariate and/or multivariate models. For the methods discussed, formulas are presented and data from an experimental study are used to demonstrate the application and interpretation of these indices. The paper concludes with some cautionary notes on the limitations associated with these measures of effect size. Copyright 2000 Academic Press. PMID- 10873374 TI - Motivational Beliefs, Study Strategies, and Mathematics Attainment in High- and Low-Achieving Chinese Secondary School Students. AB - In order to examine the relationship between cognitive and motivational variables and their relationship to mathematics attainment, Hong Kong-Chinese students enrolled in schools for high-, average-, and low-achievers completed questionnaires in Year 10 and in Year 11. Low-achievers perceived academic learning as being less useful over time and reported spending less time studying in Year 10 than in Year 11 but high- and low-achievers did not differ on their use of self-regulated learning strategies. Performance on the public examination in mathematics was predicted by prior achievement and Self-Concept of Mathematics Ability. Results underscore the importance of considering cultural beliefs systems and educational systems in models of academic motivation. Copyright 2000 Academic Press. PMID- 10873375 TI - How Organizational Signals, Need for Cognition, and Verbal Ability Affect Text Recall and Recognition. AB - This study investigated the effects of organizational signals, need for cognition, and verbal ability on recall and recognition of information from an expository text. Ninety-two undergraduate students completed the Need for Cognition scale (Cacioppo, Petty, & Kao, 1984) and read a text that either: (a) contained organizational signals in the form of an overview, headings, and a summary or (b) contained no signals. Consistent with our primary hypothesis, there was a marginal tendency for organizational signals to interact with need for cognition to influence conditional recall of expository text information. Specifically, need for cognition was related marginally to conditional recall only in the "no signals" condition. Organizational signals, need for cognition, and verbal ability contributed significantly to prediction of performance on three measures of text recall. By contrast, performance on the recognition test was influenced by an interaction between organizational signals and verbal ability. Implications of the findings are discussed. Copyright 2000 Academic Press. PMID- 10873376 TI - Effects of Informal Cooperative Learning and the Affiliation Motive on Achievement, Attitude, and Student Interactions. AB - The purpose of this study was to investigate the effect of informal cooperative learning and the affiliation motive on achievement, attitude, and student interactions. Participants classified as high or low need for affiliation used either an informal cooperative learning strategy or an individual strategy while receiving information, examples, practice and feedback from an instructional television lesson. Results indicated that participants who used the individual strategy acquired significantly more knowledge from the lesson and indicated significantly more continuing motivation for working alone than those who used the informal cooperative strategy. Instructional strategy did not influence performance on the application portion of the test. Results also revealed that high affiliation participants expressed significantly more continuing motivation than low affiliation participants for working with another person. Low affiliation participants expressed significantly more continuing motivation than high affiliation participants for working alone. Finally, results indicated that high affiliation dyads exhibited significantly more on-task group behaviors (taking turns, sharing materials, group discussion of content) and significantly more off-task behaviors than low affiliation dyads. Copyright 2000 Academic Press. PMID- 10873377 TI - Identification of Ankrd2, a novel skeletal muscle gene coding for a stretch responsive ankyrin-repeat protein. AB - Mechanically induced hypertrophy of skeletal muscles involves shifts in gene expression leading to increases in the synthesis of specific proteins. Full characterization of the regulation of muscle hypertrophy is a prerequisite for the development of novel therapies aimed at treating muscle wasting (atrophy) in human aging and disease. Using suppression subtractive hybridization, cDNAs corresponding to mRNAs that increase in relative abundance in response to mechanical stretch of mouse skeletal muscles in vivo were identified. A novel 1100-bp transcript was detected exclusively in skeletal muscle. This exhibited a fourfold increase in expression after 7 days of stretch. The transcript had an open reading frame of 328 amino acids encoding an ATP/GTP binding domain, a nuclear localization signal, two PEST protein-destabilization motifs, and a 132 amino-acid ankyrin-repeat region. We have named this gene ankyrin-repeat domain 2 (stretch-responsive muscle) (Ankrd2). We hypothesize that Ankrd2 plays an important role in skeletal muscle hypertrophy. PMID- 10873378 TI - A novel conserved cochlear gene, OTOR: identification, expression analysis, and chromosomal mapping. AB - We have identified a novel cochlear gene, designated OTOR, from a comparative sequence analysis of over 4000 clones from a human fetal cochlear cDNA library. Northern blot analysis of human and chicken organs shows strong OTOR expression only in the cochlea; very low levels are detected in the chicken eye and spinal cord. Otor and Col2A1 are coexpressed in the cartilaginous plates of the neural and abneural limbs of the chicken cochlea, structures analogous to the mammalian spiral limbus, osseous spiral lamina, and spiral ligament, and not in any other tissues in head and body sections. The human OTOR gene localizes to chromosome 20 in bands p11.23-p12.1 and more precisely to STS marker WI-16380. We have isolated cDNAs orthologous to human OTOR in the mouse, chicken, and bullfrog. The encoded protein, designated otoraplin, has a predicted secretion signal peptide sequence and shows a high degree of cross-species conservation. Otoraplin is homologous to the protein encoded by CDRAP/MIA (cartilage-derived retinoic acid sensitive protein/melanoma inhibitory activity), which is expressed predominantly by chondrocytes, functions in cartilage development and maintenance, and has growth inhibitory activity in melanoma cell lines. PMID- 10873379 TI - An algorithm for clustering cDNA fingerprints. AB - Clustering large data sets is a central challenge in gene expression analysis. The hybridization of synthetic oligonucleotides to arrayed cDNAs yields a fingerprint for each cDNA clone. Cluster analysis of these fingerprints can identify clones corresponding to the same gene. We have developed a novel algorithm for cluster analysis that is based on graph theoretic techniques. Unlike other methods, it does not assume that the clusters are hierarchically structured and does not require prior knowledge on the number of clusters. In tests with simulated libraries the algorithm outperformed the Greedy method and demonstrated high speed and robustness to high error rate. Good solution quality was also obtained in a blind test on real cDNA fingerprints. PMID- 10873380 TI - A novel androgen-regulated gene, PMEPA1, located on chromosome 20q13 exhibits high level expression in prostate. AB - Biologic effects of androgen on target cells are mediated in part by transcriptional regulation of androgen-regulated genes (ARGs) by androgen receptor. Using serial analysis of gene expression (SAGE), we have identified a comprehensive repertoire of ARGs in LNCaP cells. One of the SAGE-derived tags exhibiting homology to an expressed sequence tag was maximally induced in response to synthetic androgen R1881 treatment. The open reading frame of the androgen-induced RNA (PMEPA1) was characterized as a 759-bp nucleotide sequence coding for a 252-amino-acid protein. The analysis of PMEPA1 protein sequence indicated the existence of a type Ib transmembrane domain between residues 9 and 25. Analysis of multiple-tissue Northern blots revealed the highest level of PMEPA1 expression in prostate tissue. PMEPA1 expression was predominately detected in glandular epithelial cells of prostate by in situ hybridization analysis. The expression of PMEPA1 in LNCaP cells was induced by androgen in a time- and dose-specific manner. Evaluation of PMEPA1 expression in androgen dependent/independent tumors of the CWR22 xenograft model revealed that PMEPA1 was overexpressed in three of four androgen-independent tumor tissues. These observations define PMEPA1 as a novel androgen-regulated gene exhibiting abundant expression in prostate tissue. The increased expression of PMEPA1 in relapsed tumors of the CWR22 model suggests activation of androgen signaling in hormone refractory disease. PMEPA1, along with other highly androgen-induced prostate specific genes, has potential to serve as an androgen signaling read-out biomarker in prostate tissue. PMID- 10873381 TI - Identification and expression of a novel type I procollagen C-proteinase enhancer protein gene from the glaucoma candidate region on 3q21-q24. AB - A novel human Type I procollagen C-proteinase enhancer protein-like gene, PCOLCE2, was identified by sequencing an EST in the primary open-angle glaucoma (POAG) region on 3q21. The total cDNA encoded a 415-amino-acid protein that has 43% identity to the Type I procollagen C-proteinase enhancer protein (PCOLCE1). PCOLCE2 contains two CUB domains, which are thought to be involved in protein protein interactions, and an NTR module. PCOLCE2 message is expressed in the trabecular meshwork, lungs, heart, brain, liver, skeletal muscle, kidney, pancreas, and placenta as a 2-kb message. PCOLCE2, a 52-kDa protein, is expressed in the trabecular meshwork. A novel gene, PCOLCE2, has been identified and characterized. Based upon its homology with collagen-binding proteins, its expression in the trabecular meshwork, and its chromosome location, PCOLCE2 is a candidate gene for GLC1C. However, no coding sequence mutations were detected in PCOLCE2 in a POAG patient from the GLC1C family. PMID- 10873382 TI - Identification and validation of a gene involved in anchorage-independent cell growth control using a library of randomized hairpin ribozymes. AB - We have developed a library of hairpin ribozyme genes that can be delivered and expressed in mammalian cells with the purpose of identifying genes involved in a specific phenotype. By applying the appropriate phenotypic selection criteria in tissue culture, we can enrich for ribozymes that knock down expression of an unknown gene or genes in a particular pathway. Once specific ribozymes are selected, their target binding sequence is used to identify and clone the target gene. We have applied this technology to identify a putative tumor suppressor gene that has been activated in HF cells, a nontransformed revertant of HeLa cells. Using soft agar growth as the selection criteria for gain of transformation, we have isolated ribozymes capable of triggering anchorage independent growth. Isolation of one of these ribozymes, Rz 568, led to the identification and cloning of the human homologue of the Drosophila gene ppan, a gene involved in DNA replication, cell proliferation, and larval development. This novel human gene, PPAN, was verified as the biologically relevant target of Rz 568 by creating five additional "target validation" ribozymes directed against additional sites in the PPAN mRNA. Rz 568 and all of the target validation ribozymes reduced the level of PPAN mRNA in cells and promoted anchorage independent growth. Exogenous expression of PPAN in HeLa and A549 tumor cells reduced their ability to grow in soft agar, underscoring its role in regulating anchorage-dependent growth. This study describes a novel method for gene discovery where the intracellular application of hairpin ribozyme libraries was used to identify a novel gene based solely on a phenotype. PMID- 10873383 TI - Sequence analyses of the olfactory receptor gene cluster mOR37 on mouse chromosome 4. AB - The olfactory receptor multigene family is organized in clusters spread throughout the genome. In the present study, we have sequenced two subregions of the mOR37 gene cluster on mouse chromosome 4. The resulting 100 kb of sequence revealed seven odorant receptor coding regions and one gene fragment. Sequence analyses reveal that the mOR37 gene cluster may represent a rather ancient cluster. The mOR37 genes exhibit a complex intron/exon structure, and some appear to be differentially spliced. All genes in the cluster share conserved sequence motifs 5' of their putative initial exons, which represent potential binding sites for transcription factors. The clustered organization and conserved sequence motifs suggest common expression control mechanisms for these genes. PMID- 10873384 TI - The human fatty acid transport protein-1 (SLC27A1; FATP-1) cDNA and gene: organization, chromosomal localization, and expression. AB - Uptake of fatty acids into cells is a controlled process in part regulated by fatty acid transport proteins (FATPs), which facilitate the transport of fatty acids across the cell membrane. In this study the structure of the human FATP-1 (HGMW-approved symbol SLC27A1) cDNA and gene was determined, and the expression of its mRNA in human was characterized. Muscle and adipose tissue have the highest levels of FATP-1 mRNA, small intestine has intermediate levels, and FATP 1 mRNA is barely detectable in liver. The human FATP-1 gene has 12 exons and extends over more than 13 kb of genomic DNA. The FATP gene maps to chromosome 19p13.1 by fluorescence in situ hybridization, a region previously suggested to be implicated in the determination of small dense low-density lipoprotein (LDL). Knowledge of the gene structure and chromosomal localization will allow screening for FATP mutations in humans with metabolic disorders, whereas knowledge of its expression pattern and factors regulating its expression could be of importance in understanding its biology. PMID- 10873385 TI - Cloning and characterization of the murine pkd2 promoter. AB - Pkd2, the mouse homologue of PKD2, the gene responsible for the second form of autosomal dominant polycystic kidney disease, is highly expressed in fetal and adult mouse tissues. The expression of Pkd2 is developmentally regulated. To begin to dissect out the regulatory mechanism of Pkd2 expression, we characterized the basic features of the gene structure and identified potential cis-regulatory elements of Pkd2 transcription. Pkd2 spans 42 kb with a transcription start site 165 bp upstream of the translation start codon. Exon 1 of Pkd2 is 755 bp long, and the full-length transcript is 5215 bp. The Pkd2 promoter region is GC-rich and lacks a consensus TATA or CCAAT box. Consensus binding sites for the transcription factors Sp-1, NF-1, and Ap-2 lie in the 5' upstream region of Pkd2. The Sp-1 binding site is conserved in 5' upstream sequences of both the mouse and the human genes. The CAT activity of a series of upstream segments from +178 to -2749 was assessed in MDCK, LLCPK1, COS-7, and HEK293 cells. Deletion analysis identified a 409-bp fragment from position -221 to +178 responsible for basal promoter activity. A 922-bp fragment from -744 to +178 showed the highest level of CAT activity in the cell lines tested. These data define a functional promoter candidate region for Pkd2. PMID- 10873386 TI - The mouse Psma1 gene coding for the alpha-type C2 proteasome subunit: structural and functional analysis, mapping, and colocalization with Pde3b on mouse chromosome 7. AB - We have isolated and functionally characterized the mouse gene for the C2 subunit of the 20S proteasome. The gene contains 10 exons distributed over a region of 12 kb on the distal end of mouse chromosome 7. Its exon-intron structure differs from those of the other few known proteasome genes. Transfection assays revealed that 1.5 kb of 5' flanking sequence is active as promoter in cultured myoblasts. Deletion reporter constructs narrowed this presumptive promoter region to within 450 bp upstream of the translation initiation site. Several consensus motifs for transcription factor binding sites were identified in this upstream region of the gene. Psma1 was mapped to mouse chromosome 7 using the interspecific backcross DNA panels from The Jackson Laboratory. Additional mapping studies showed that the mouse genes Psma1 and Pde3b are closely linked, residing between cM 53 and 53.3 in a region syntenic to human chromosome 11p15. Our results extend the structural and functional analysis of genes encoding the 20S proteasome subunits and provide the basis for the study of their regulation. PMID- 10873387 TI - Isolation and characterization of a calcium channel gene, Cacna1f, the murine orthologue of the gene for incomplete X-linked congenital stationary night blindness. AB - The mutant L-type calcium channel alpha(1)-subunit gene, CACNA1F, was recently identified as the gene responsible for incomplete X-linked congenital stationary night blindness. The 6070-bp mRNA transcript is predicted to encode a 1977-amino acid pore-forming protein with cytoplasmic amino- and carboxyl-termini separated by four homologous repeat domains, each consisting of six transmembrane segments. CACNA1F has been shown to be preferentially expressed in the retina, indicative of a specific functional role in visual processing. We have established the complete sequence of the murine orthologue of CACNA1F, namely Cacna1f. The total length of the mRNA transcript of the murine gene was established to be 6080 bp with an open reading frame that translates into a 1985-amino-acid protein. Cacna1f is highly homologous to the human sequence, with 90% identity at the amino acid level and almost perfect conservation between the functional domains. Furthermore, as in the human gene, the 3' end of the Cacna1f gene maps within 5 kb of the 5' end of the mouse synaptophysin gene in a region orthologous to Xp11.23. Using in situ hybridization, Cacna1f was found to be expressed in the inner and outer nuclear layers and the ganglion cell layer of the retina. PMID- 10873388 TI - Molecular cloning, expression characterization, and mapping of a novel putative inhibitor of rho GTPase activity, RTKN, to D2S145-D2S286. AB - The Rho proteins are a class of small molecular GTPases that regulate multiple fundamental cellular processes by mediating the G-protein-coupled receptor signaling pathway. Rhotekin, which is one of the downstream target molecules of Rho with a Rho binding motif class I domain, can inhibit endogenous or RhoGAP stimulating Rho GTPase activity to regulate the signaling pathway. Here, a novel human cDNA containing an intact open reading frame that encodes 544 amino acids has been identified. As this putative protein shares 84. 6% amino acid identity with mouse Rhotekin, and has a tandem Rho binding domain class 1 and Pleckstrin homology domain, it was regarded as a human homologue of the mouse Rhotekin and assigned a symbol of RTKN. With the human Rhotekin cDNA as a probe, Northern hybridization revealed that a 4.0-kb transcript was expressed at a high level in prostate and at a middle level in 13 of 16 tissues examined, but it cannot be detected in liver and lung tissues. Meanwhile, a 2.4-kb transcript was expressed at a middle level in prostate and another 3.0-kb transcript in kidney. In addition, the RTKN gene was localized to chromosome 2p13 between markers D2S145 at 6.94 cR (LOD > 12) and D2S286 at 8.12 cR (LOD > 9.7) by radiation hybrid panel mapping. Compared with BAC clone AC005041 sequence, there were 12 exons for the RTKN gene and it spanned a 16.5-kb genomic region. PMID- 10873389 TI - YAC/BAC-based physical and transcript mapping around the gracile axonal dystrophy (gad) locus identifies Uchl1, Pmx2b, Atp3a2, and Hip2 genes. AB - We generated a yeast artificial chromosome (YAC)/bacterial artificial chromosome (BAC)-based physical and transcript map of a region containing the gracile axonal dystrophy (gad) locus on mouse chromosome 5. The YAC/BAC contig consists of 13 YAC and 49 BAC clones onto which 4 genes, 40 expressed sequence tags, and 7 new DNA polymorphisms were ordered. Using this physical map, we mapped Uchl1 encoding ubiquitin carboxyl-terminal hydrolase I, whose deletion has been determined to cause the gad mutation. We also mapped three other recently identified genes: Hip2, encoding Huntingtin interacting protein 2; Atp3a2, encoding a P-type ATPase; and Pmx2b, encoding PHOX2b. PMID- 10873390 TI - P53 and IGFBP-3: apoptosis and cancer protection. AB - p53, perhaps the single most important human tumor suppressor, is commonly mutated in human cancers. Normally genotoxic stress and hypoxia activate p53, which, through DNA-specific transcription activation, transcriptional repression, and protein-protein interactions, triggers cell cycle arrest and apoptosis. One of the genes induced by p53 was identified as that encoding the insulin-like growth factor binding protein (IGFBP)-3. IGFBP-3 was originally defined by the somatomedin hypothesis as the principal carrier of IGF-I in the circulation and the primary regulator of the amount of free IGF-I available to interact with the IGF-1 receptor. However, there is accumulating evidence that IGFBP-3 can also cause apoptosis in an IGF-independent manner. Thus, IGFBP-3 induction by p53 constitutes a new means of cross-talk between the p53 and IGF axes, and suggests that the ultimate function of IGFBP-3 may be to serve a protective role against the potentially carcinogenic effects of growth hormone and IGF-I. PMID- 10873391 TI - Fragile sites and minisatellite repeat instability. PMID- 10873392 TI - Reconstitution of TCP80/NF90 translation inhibition activity in insect cells. AB - Acid beta-glucosidase (GCase) is the enzyme deficient in Gaucher disease, an inherited metabolic prototype for enzyme and gene therapy. An 80-kDa mammalian cytoplasmic protein (TCP80/NF90) was discovered to interact with the GCase mRNA coding region and inhibit its translation in vitro and ex vivo. Human TCP80/NF90 is identical to NF90, an IL-2 enhancer protein, and MPP4, an M-phase phosphoprotein. The interaction of recombinant TCP80/NF90 with GCase mRNA was evaluated using the baculovirus/Sf9 insect cell system since these cells lack this protein. Purified recombinant and isolated mammalian cytoplasmic TCP80/NF90 had identical functions including binding of coding regions of selected RNAs and inhibition of their in vitro translation. Individual baculoviruses containing the human TCP80/NF90 cDNA (vSf9/TCP80) and GCase cDNA (vSf9/GCase) were used to co infect Sf9 cells. The presence of preformed TCP80/NF90 significantly (>87%) inhibited wild-type GCase mRNA translation in these cells, but baculovirus containing a mutant GCase did not. Sf9 cells co-infected with vSf9/TCP80 showed a major reduction of GCase RNA polysome association. These results show that the multifunctional protein, TCP80/NF90, can function in vivo as a translation inhibitory protein and include alterations of mRNA binding to polysomes as a component of its mechanism of action. PMID- 10873393 TI - Biochemical and molecular investigations of patients with nonketotic hyperglycinemia. AB - The investigation of 14 unrelated patients with nonketotic hyperglycinemia led to the identification of mutations in 4 cases. Patients were initially categorized into probable P- or T-protein defects of the glycine cleavage enzyme complex, by the use of the glycine exchange assay without supplemental H-protein, then screened for mutations in the P-protein and T-protein genes, respectively. PMID- 10873394 TI - Ataxia-telangiectasia: phenotype/genotype studies of ATM protein expression, mutations, and radiosensitivity. AB - Previous studies on a limited number of ataxia-telangiectasia (A-T) patients with detectable levels of intracellular ATM protein have suggested a genotype/phenotype correlation. We sought to elucidate this possible correlation by comparing ATM protein levels with mutation types, radiosensitivity, and clinical phenotype. In this study, Western blot analysis was used to measure ATM protein in lysates of lymphoblastoid cell lines (LCLs) from 123 unrelated A-T patients, 10 A-T heterozygotes, and 10 patients with phenotypes similar to A-T. Our Western blot protocol can detect the presence of ATM protein in as little as 1 microg of total protein; at least 25 microg of protein was tested for each individual. ATM protein was absent in 105 of the 123 patients (85%); most of these patients had truncating mutations. The remaining subset of 18 patients (15%) had reduced levels of normal-sized ATM protein; missense mutations were more common in this subset. We used a colony survival assay to characterize the phenotypic response of the LCLs to radiation exposure; patients with or without detectable ATM protein were typically radiosensitive. Nine of 10 A-T heterozygotes also had reduced expression of ATM, indicating that both alleles contribute to ATM protein production. These data suggest that although ATM specific mRNA is abundant in A-T cells, the abnormal ATM protein is unstable and is quickly targeted for degradation. We found little correlation between level of ATM protein and the type of underlying mutation, the clinical phenotype, or the radiophenotype. PMID- 10873395 TI - A variable myopathy associated with heterozygosity for the R503C mutation in the carnitine palmitoyltransferase II gene. AB - Adult-onset carnitine palmitoyltransferase II (CPT II) deficiency is an autosomal recessive disease characterized by muscle pain and stiffness with rhabdomyolysis and myoglobinuria in severe cases. Exercise, fasting, viral infection, anesthesia, or extremes in temperature may trigger symptoms. A 54-year-old woman exhibited a 35-year history of progressive weakness and myopathic symptoms. CPT II activity in the patient's lymphoblasts, cultured skin fibroblasts, and skeletal muscle was reduced to 47, 43, and 13% of normal, respectively. Respiratory chain enzymes were also reduced in muscle ranging from 22 to 49% of their respective normal reference means. beta-oxidation enzymes in fibroblasts ranged from 29 to 63% of normal. The patient, her father, and her 26-year-old son were all heterozygous for the R503C mutation. The patient's son has a lifelong history of myopathic symptoms while his grandfather only had mild weakness during childhood. Analysis of the V368I and M647V polymorphisms in the CPT2 gene showed that the mutant allele is linked to 368I and 647M in this family and that the normal allele is linked to 647V in the affected patient and her son, and to 647M in the patient's father. While the variability in CPT2 gene haplotypes may contribute to the phenotypic complexities in this family, it is also possible that an additional gene defect in the transport of mitochondrial proteins contributes to the complex phenotype in the patient. We present biochemical and molecular evidence for vertical transmission of a variable myopathy caused by heterozygosity for a single mutation, R503C, in the CPT2 gene. PMID- 10873396 TI - Prevalence of AIPL1 mutations in inherited retinal degenerative disease. AB - Leber congenital amaurosis (LCA) is the most severe form of inherited retinal dystrophy and the most frequent cause of inherited blindness in children. LCA is usually inherited in an autosomal recessive fashion, although rare dominant cases have been reported. One form of LCA, LCA4, maps to chromosome 17p13 and is genetically distinct from other forms of LCA. We recently identified the gene associated with LCA4, AIPL1 (aryl-hydrocarbon interacting protein-like 1) and identified three mutations that were the cause of blindness in five families with LCA. In this study, AIPL1 was screened for mutations in 512 unrelated probands with a range of retinal degenerative diseases to determine if AIPL1 mutations cause other forms of inherited retinal degeneration and to determine the relative contribution of AIPL1 mutations to inherited retinal disorders in populations worldwide. We identified 11 LCA families whose retinal disorder is caused by homozygous or compound heterozygous AIPL1 mutations. We also identified affected individuals in two apparently dominant families, diagnosed with juvenile retinitis pigmentosa or dominant cone-rod dystrophy, respectively, who are heterozygous for a 12-bp AIPL1 deletion. Our results suggest that AIPL1 mutations cause approximately 7% of LCA worldwide and may cause dominant retinopathy. PMID- 10873397 TI - Functional analyses of amino acid substitutions Arg883Ser and Asp905Tyr of protein phosphatase-1 G-subunit. AB - The PPP1R3 gene encoding the G-subunit of protein phosphatase-1 has three polymorphisms in linkage disequilibrium in the Pima Indians: an mRNA destabilizing element in the 3'-untranslated region (ARE1/ARE2 alleles), Arg883Ser, and Asp905Tyr substitutions. The ARE2 allele, Arg883, and Asp905 variants are associated with insulin resistance and higher prevalence of type 2 diabetes in the Pima Indians. The ARE2 allele is associated with lower PPP1R3 transcript and protein levels in muscle tissue. Here we determined the functional contribution of the amino acid substitutions independent of the ARE alleles to insulin-stimulated glycogen synthesis by adenoviral-mediated gene expression in L6 myotubes. Similar overexpression levels of the G-subunit variants increased glycogen synthase fractional activity in the presence ( approximately 1. 5-fold) of insulin compared to control myotubes transduced with adenovirus encoding beta galactosidase. The glycogen synthesis rate of myotubes overexpressing the G subunit variants also increased by approximately 1.7-fold over the control with and without insulin. However, these measures were not significantly different among the variants. This study does not support a role for Arg883 and Asp905 variants independent of the ARE2 allele in the impaired insulin-stimulated glycogen synthesis in the muscle of Pima Indians. PMID- 10873398 TI - The peroxisome proliferator-activated receptor gamma 2 Pro12Ala mutation is associated with early onset extreme obesity and reduced fasting glucose. AB - We screened the peroxisome proliferator activated receptor gamma 2 (PPAR gamma 2) for sequence variants in 165 unrelated obese (BMI >/= 30 kg/m(2)) Caucasian women, and 49 normal weight Caucasian female controls (BMI < 27 kg/m(2)). The allele frequency of the Pro12Ala mutation was higher in obese(18.18%) than in normal weight women (8. 16%) (chi(2)((1)) = 5.68, P = 0.017). Among obese women, the Pro12Ala mutation lowered age of obesity onset (Pro/Pro, 13.2 +/- 9. 4 years; Pro/Ala+Ala/Ala 8.6 +/- 7.1 years, P = 0.005), was associated with lower fasting glucose and was protective against type II diabetes. PMID- 10873400 TI - Analysis of the cost effectiveness of concurrent cisplatin-based chemoradiation in cervical cancer: implications from five randomized trials. AB - PURPOSE: Five recent phase III trials provide strong evidence that a new alternative therapy, cisplatin-based chemoradiation, is more effective than standard therapy using radiation alone in the treatment of advanced cervical cancer. We conducted a pharmacoeconomic analysis to determine whether the alternative cisplatin-based chemoradiation is cost effective as compared with standard therapy using radiation alone. METHODS: Using an economic model, we applied costs to resource utilization data derived from the cisplatin-based chemoradiation arms of these five randomized trials. We examined the cisplatin based chemoradiation benefits in terms of increased median survival time. Incremental costs were divided by difference in survival to determine the cost per patient benefited. Incremental cost per year of life gained (IC/YLG) was calculated based on both published and estimated survival figures. RESULTS: Cost per year of life gained for cisplatin-based chemoradiation regimens varied from $2384 to $28,770 based on published survival and from $308 to $3712 based on estimated survival. Variations in regimen cost were largely dependent on treatment setting. Administration costs per patient for cisplatin and fluorouracil in the inpatient setting were $8839 compared with $3590 in the outpatient setting. CONCLUSION: The increased median survival cost per year of life gained with cisplatin-based chemoradiation (inpatient and outpatient settings) adds a substantial benefit at an acceptable cost compared with radiation therapy alone. PMID- 10873399 TI - Better treatments that cost more: the dilemma. PMID- 10873401 TI - Contemporary surgical management of borderline ovarian tumors: a survey of the Society of Gynecologic Oncologists. AB - OBJECTIVE: The objective of this study was to review the current practice patterns regarding the surgical management of borderline ovarian tumors. METHODS: A one-page survey was mailed to the members of the Society of Gynecologic Oncologists (SGO), using the directory of the Society. The survey addressed the demographics of the respondent and the recommended staging procedure for presumed early-stage disease. RESULTS: Of the 660 surveys mailed, 274 (42%) were returned. Ninety-seven percent (267/274) of respondents advocate surgical staging. Of this group, 96% (257/267) perform peritoneal washings, 97% (259/267) sample the omentum, and 92% (245/267) submit random peritoneal biopsies. Eighty-eight percent (235/267) perform lymph node sampling: paraaortic biopsies by 89% (210/235) and pelvic biopsies by 97% (228/235). Of this latter group, 91% sample the external iliac chain, 82% submit hypogastric nodal tissue, and 70% remove obturator lymph nodes. CONCLUSION: Diversity exists in the surgical management of borderline ovarian tumors among members of the SGO who responded to this survey. Efforts to ensure a consistent approach to the management of borderline ovarian tumors are warranted. PMID- 10873402 TI - Frequent down-regulation and lack of mutation of the KAI1 metastasis suppressor gene in epithelial ovarian carcinoma. AB - OBJECTIVE: KAI1 is a recently identified metastasis suppressor gene on human chromosome 11p11.2. It belongs to a structurally distinct family of cell surface glycoproteins. Decreased KAI1 expression seems to be involved in the progression of human prostate, lung, pancreatic, and possibly breast cancer, and recently a reduced KAI1 protein expression has been demonstrated in several ovarian carcinoma cell lines. The aim of this study is to determine whether the KAI1 gene is altered in human epithelial ovarian carcinomas. In addition, its prognostic significance in this tumor is also evaluated. METHODS: To detect KAI1 expression, 102 tumor samples from benign, borderline, primary invasive, metastatic, and recurrent epithelial ovarian tumors were prepared for immunohistochemical study with C-16, an anti-KAI1 polyclonal antibody. In addition, cellular RNA from 24 primary invasive and 7 recurrent tumors was also analyzed for KAI1 expression by using a reverse transcriptase PCR (RT-PCR) technique. The PCR single-strand conformation polymorphism method and direct DNA sequencing were used to detect KAI1 mutation in the 44 primary invasive and 8 recurrent ovarian carcinomas. RESULTS: In immunohistochemical study, decrease of KAI1 protein expression was associated with the progression of ovarian tumor. However, it had no relation to the stage of primary invasive cancers because of its frequent occurrence in early stage tumors. KAI1 expression was also frequently down-regulated in primary invasive and recurrent tumors in RT-PCR analysis. Except for a missense change at codon 241 (ATC to GTC), which causes the substitution of a valine for an isoleucine in the amino acid sequence and occurs in both normal and tumor tissues, no mutation of the KAI1 gene was found in any of the 52 carcinomas. Although there was a trend for deteriorating survival from patients with KAI1 preserved tumors to those with KAI1-decreased and -negative tumors, statistically it was not significant (P = 0.079). CONCLUSION: KAI1 may play a role in the malignant progression of epithelial ovarian carcinoma through the down-regulation of expression rather than gene mutation. Since the decreased expression presented frequently in early stage tumors, it may be an early event in the progression of this tumor and its prognostic significance needs further investigation with a larger number of cases. PMID- 10873403 TI - Serum CA 125, carcinoembryonic antigen, and CA 19-9 as tumor markers in borderline ovarian tumors. AB - OBJECTIVES: The goals of this study were to analyze preoperative serum levels of CA 125, carcinoembryonic antigen (CEA), and CA 19-9 in patients with borderline ovarian tumors and to investigate if routine assessment of these markers in follow-up may lead to earlier detection of recurrence. METHODS: For patient identification a database was used, in which data from all patients treated for gynecologic malignancies in the Department of Gynecologic Oncology, University Hospital Groningen, The Netherlands, are compiled. Between 1982 and 1997, 44 patients with borderline ovarian tumors were identified. Clinical data and serum CA-125 and CEA levels were retrieved from the database. CA 19-9 levels were determined in retrospect in available stored preoperative (24 patients) and follow-up (43 patients) serum samples. RESULTS: Preoperative CA 125 levels were elevated in 8 of 33 (24%), CEA levels in 3 of 32 (9%), and CA 19-9 levels in 11 of 24 (46%) cases. In patients with mucinous tumors preoperative CA 19-9 was more frequently elevated (8/14, 57%) than CA 125 (3/20, 15%) (P = 0.02) or CEA (2/18, 11%) (P = 0.02). Complete follow-up serum CA 125, CEA, and CA 19-9 levels were available for 43 of 44 patients. Median follow-up was 84 months (range, 22-204). During follow-up two patients (5%) had recurrent disease. In one patient CA 125 became elevated at the time of recurrence; in the other patient (in retrospect) the CA 19-9 level did not return to normal after surgery, but kept rising, preceding clinical symptoms of recurrence for 13 months. CONCLUSIONS: If one chooses to use serum markers in follow-up of mucinous borderline ovarian tumors CA 19-9 should be included. Measurement of serum tumor markers in the follow-up of patients with borderline ovarian tumors may lead to earlier detection of recurrence in only a very small proportion of patients, while the clinical value of earlier detection of recurrence remains to be established. PMID- 10873404 TI - Long-term results and prognostic factors in patients with epithelial ovarian cancer. AB - OBJECTIVES: The aim of this study was to evaluate long-term results and to assess prognostic factors which have an impact on overall survival in patients with epithelial ovarian cancer. METHODS: A retrospective analysis of 287 patients treated between 1975 and 1995 was performed. All operations were performed by senior surgeons. Histologic sections were reviewed by the same pathologist. Successive adjuvant chemotherapy regimens are described. Survival was evaluated in 1997. Follow-up lasted 25-260 months (median 90). Statistical methods included Kaplan-Meier survival curves, log-rank test, and multivariate analysis. RESULTS: The 5-year survival rates were 76, 42, 21, and 6% for patients with stage I, II, III, and IV disease, respectively. Age, FIGO stage, cytology of ascites, histologic type and grade, extent of surgery, and number of residual tumors were significant prognostic indicators in univariate analysis. Multivariate analysis showed that the risk of mortality according to FIGO stage was 2.8, 95% CI [1.2 6.3], P = 0.01 for FIGO II, 5.6, 95% CI [2.9-10.8], P < 0.001 for FIGO III, and 10.5, 95% CI [4.9-22. 1], P < 0.001 for FIGO IV in comparison with FIGO I. Patients with a serous epithelial carcinoma had a 1.7-fold higher risk of mortality than patients with other histologic types: RR = 1.7, 95% CI [1.1-2. 8], P < 0.001. Patients whose tumors distribution permitted optimal surgery had a 2.3 fold lower risk of mortality than patients treated with sub- or nonoptimal surgery: RR = 0.43, 95% CI [0.29-0.64], P < 0.001. The risk of mortality for patients treated with alkylating agents, platinum-based combination chemotherapy without taxanes, or carboplatin plus paclitaxel regimens compared with patients who did not receive treatment was reduced by 47%, 95% CI [8-69%], P = 0.025, 55%, 95% CI [22-74%], P = 0.005, and 70%, 95% CI [35-86%], P = 0.002, respectively. CONCLUSION: Our study confirms the benefit of cytoreductive surgery and the efficacy of platinum plus paclitaxel first-line chemotherapy which has recently been recognized as the standard treatment for advanced epithelial ovarian cancer. PMID- 10873405 TI - Combination chemotherapy with methotrexate, etoposide, and actinomycin D for high risk gestational trophoblastic tumors. AB - OBJECTIVES: The goal of this study was to evaluate the efficacy, toxicity, and survival of patients with high-risk gestational trophoblastic tumors (GTTs) treated with a methotrexate-etoposide-actinomycin D (MEA) regimen without cyclosphosphamide or vincristine. METHODS: Thirty-nine consecutive patients with high-risk GTTs (28 were defined high risk by WHO criteria) were treated with primarily the MEA regimen. Among them, 27 patients had received no prior chemotherapy and 12 had received prior chemotherapy. Survival, causes of treatment failure, and toxicity were analyzed retrospectively. RESULTS: After treatment with the MEA regimen, 29 of 39 patients achieved primary remission (74.4%), 8 developed resistance (20.5%), and 2 died of widespread metastases and chemotherapy-related toxicity. All 8 patients who developed resistance were treated with high-dose 5-fluorouracil and actinomycin D (FA); 6 were salvaged and 2 died of refractory disease. Three patients relapsed; 2 were controlled with FA or cisplatin-based chemotherapy and 1 who refused further treatment died. The disease-free survival rate was 87%. WHO grade 4 leukocytopenia and thrombocytopenia with the MEA regimen occurred in 5.3 and 6.4%, respectively, of the cycles; other toxic effects were acceptable and manageable. CONCLUSIONS: At present, MEA chemotherapy (without cyclophosphamide or vincristine) is our treatment of choice for patients with high-risk GTT. Its toxicity is predictable and manageable. For patients who become resistant to MEA, new salvage chemotherapy regimens are needed. PMID- 10873406 TI - One versus two intracavitary brachytherapy applications in early-stage cervical cancer patients undergoing definitive radiation therapy. AB - PURPOSE: The purpose of this study was to compare the outcomes of early stage cervical cancer patients undergoing definitive radiation therapy (RT) with one versus two low-dose-rate intracavitary brachytherapy (ICB) applications. METHODS AND MATERIALS: Between 1983 and 1993, 140 stage IB-IIA patients underwent whole pelvis RT (WPRT) and ICB. Prior to 1988, 56 patients (40%) received two ICB applications. After 1988, our policy was modified and subsequently 84 (60%) patients underwent one application. Patient, tumor, and treatment characteristics, outcome, and complications of the two groups were compared. RESULTS: The groups were balanced in terms of race, hemoglobin level, histology, grade, treatment duration, chemotherapy, and follow-up. The single-application group, however, had more stage IB disease, had small (< or =4 cm) tumors, and received higher WPRT and lower point A doses. Overall, the two groups had similar 5-year local control (P = 0.83) and disease-free (P = 0.23) and cause-specific (P = 0.29) survival rates. Moreover, no differences were seen when analyzed by tumor size or stage. On multivariate analysis, the number of applications was not correlated with recurrence (P = 0.59, hazard rate = 1.1, 95% confidence interval = 0.6-2.2). Chronic complications were similar in the two groups. CONCLUSION: Our nonselected comparison of one versus two ICB applications in early-stage cervical cancer patients reveals comparable outcomes and complication rates for the two approaches. These results support the use of a single application in early-stage patients undergoing definitive RT. PMID- 10873407 TI - Measurement of CA-125 in trophoblastic disease. AB - OBJECTIVES: Physicians treating hydatidiform mole are still seeking means of identifying those patients who will require chemotherapy. The standard accepted method is to follow human chorionic gonadotropin levels but CA-125 measurement has been suggested as a supplement that may be clinically useful. This study was undertaken to validate or refute the one previous study that addresses this issue. CA-125 was measured at the time of hydatidiform mole evacuation to determine (1) whether it would predict the need for chemotherapy and (2) whether it correlated with human chorionic gonadotropin and tumor load in following patients with hydatidiform mole and metastatic gestational trophoblastic disease. PATIENTS AND METHODS: CA-125 was measured in serial weekly samples selected from diagnostic groups of patients with trophoblastic disease. Sixteen patients had hydatidiform mole with spontaneous resolution, fourteen had nonmetastatic gestational trophoblastic tumor, and four had low-risk metastatic disease. Six patients had high-risk metastatic disease. Ten patients had partial hydatidiform mole and one of these required chemotherapy. One patient had primary ovarian choriocarcinoma and three had placental site tumor. RESULTS: The mean preevacuation CA-125 among the 15 patients with complete hydatidiform mole was 40.9 U/ml: 52.5 U/ml for 5 patients who required chemotherapy and 36.2 U/ml for 10 patients who did not require chemotherapy. There was no statistical difference between these values. There was no correlation of CA-125 with hCG. Frequently CA 125 became negative when hCG was still elevated. Among six patients with high risk disease, CA-125 was elevated in four but in all six patients hCG remained elevated when CA-125 became negative. In nine patients with partial hydatidiform mole CA-125 was elevated prior to mole evacuation and then became negative. The patient with a tetraploid conceptus who required chemotherapy had negative CA 125. With placental site tumor CA-125 was negative, but it was elevated with ovarian choriocarcinoma. CONCLUSION: CA-125 levels do not provide reliable information in the management of patients with gestational trophoblastic disease. PMID- 10873408 TI - The antiemetic efficacy of oral ondansetron plus intravenous dexamethasone in patients with gynecologic malignancies receiving carboplatin-based chemotherapy. AB - PURPOSE: The purpose of this study was to develop a cost-effective prophylactic antiemetic regimen for the prevention of carboplatin-induced emesis. METHODS: Patients being treated in the Gynecologic Cancer Program of the Cleveland Clinic Taussig Cancer Center with a carboplatin-based chemotherapy regimen received a prophylactic antiemetic program consisting of a single dose of oral ondansetron (16 mg) plus intravenous dexamethasone (20 mg) approximately 30 min prior to chemotherapy. Evaluation of the effectiveness of this antiemetic regimen was performed during a single treatment course. RESULTS: A total of 27 patients (median age, 62; range, 41-83) participated in this phase 2 trial. Three patients received single-agent carboplatin, and 24 were treated with either a carboplatin/paclitaxel or carboplatin/docetaxel regimen. The carboplatin AUC dosing level was 4, 5, or 6 in 6, 5, and 16 individuals, respectively. No patient developed vomiting; 2 (7%) individuals experienced nausea during the 24-h period following chemotherapy administration. CONCLUSION: The combination of a single dose of oral ondansetron (16 mg) plus intravenous dexamethasone (20 mg) is an effective prophylactic antiemetic regimen for patients receiving carboplatin based chemotherapy. PMID- 10873409 TI - Combined colposcopy, loop conization, and laser vaporization reduces recurrent abnormal cytology and residual disease in cervical dysplasia. AB - OBJECTIVES: Loop electrosurgical excision of the transformation zone (LEETZ) was recently associated with relatively high failure rates. We evaluated whether the combination of LEETZ with laser vaporization is superior to LEETZ alone in reducing the rates of recurrent abnormal cytology and residual disease. METHODS: The study population included 426 women with histologic diagnosis of cervical intraepithelial neoplasia (CIN) 2-3, of whom 289 (study group) were treated by LEETZ followed by laser vaporization of the crater base and walls and 137 (control group) were treated by LEETZ alone. All women were followed scrupulously at regular intervals for recurrent abnormal cytology and residual disease. The mean follow-up periods were 43 and 59 months for the study and control groups, respectively. RESULTS: Both groups were derived from the same community and were similar in epidemiologic characteristics and disease severity. Although the incidence of positive surgical margins was similar in both groups (10.4 and 9.5% for the study and control groups, respectively), recurrent abnormal cytology (10.2% vs 5.5%, P = 0.07) and histologic residual disease (21.4% vs 0%, P = 0.05) were more frequent among women in the control group. This applied to women with both negative and positive surgical margins. Both study and control women with positive surgical margins, especially at the endocervix, were at higher risk for recurrence. CONCLUSION: The addition of laser vaporization to LEETZ may improve outcome of both women with positive margins and women with negative margins. Our results support conservative management for all treated women, regardless of cone margin status. PMID- 10873410 TI - Paclitaxel and cisplatin in advanced or recurrent carcinoma of the endometrium: long-term results of a phase II multicenter study. AB - OBJECTIVES: Both paclitaxel and cisplatin have moderate activity in patients with metastatic or recurrent carcinoma of the endometrium, and the combination of these two agents has shown activity in a variety of solid tumors. We administered this combination to patients with metastatic or recurrent carcinoma of the endometrium to evaluate its activity and to define its toxicity. METHODS: Twenty four consecutive patients were treated on an outpatient basis with paclitaxel 175 mg/m(2) administered intravenously over a 3-h period followed by cisplatin 75 mg/m(2) administered intravenously with granulocyte colony-stimulating factor (G CSF) support. The chemotherapy was repeated every 3 weeks for a maximum of six courses. RESULTS: Sixteen patients (67%; 95% confidence interval, 45-84%) achieved an objective response, including seven complete responses and nine partial responses. The median duration of response was 7 months, and the median times to progression and survival for all patients were 8.4 and 17.6 months, respectively. Some degree of neurotoxicity occurred in 44% of patients. Grade 3 or 4 toxicity included granulocytopenia in 22% of patients and peripheral neuropathy in 9%. CONCLUSION: The combination of paclitaxel with cisplatin with G CSF support appears active in patients with metastatic or recurrent carcinoma of the endometrium. The significant incidence of neurotoxicity is of concern and alternative methods of administration of the two agents could be evaluated. PMID- 10873411 TI - Effect of surgeon's experience on the surgical outcome of laparoscopic surgery for women with endometrial cancer. AB - OBJECTIVE: The purpose of this study was to assess the effect of increasing surgeon's experience in the laparoscopic management of women with endometrial cancer on the surgical outcome of these patients. METHODS: Seventy-five consecutive women with clinical stage I endometrial cancer who underwent laparoscopically assisted vaginal hysterectomy, bilateral salpingo-oophorectomy, and pelvic lymph node sampling by the same surgeon using the same technique and instruments over a period of 2 years were divided into three equal groups based on the date of surgery. The three groups were compared in patient characteristics and surgical outcome using one-way analysis of variance and Pearson chi(2) tests. RESULTS: The three groups were similar in patient characteristics. There was no significant difference in estimated amount of blood loss, rate of conversion to laparotomy, complications, and length of hospital stay among the three groups. There was a significant decrease in operating time (means: 231.0 min for group 1, 175.0 min for group 2, and 167.7 min for group 3, P < 0.001) and a significant increase in the number of pelvic lymph nodes harvested (7.8 for group 1, 10.6 for group 2, and 11.9 for group 3, P < 0.05) with increasing surgeon's experience. CONCLUSIONS: A learning curve is demonstrated in the laparoscopic management of women with endometrial cancer. With increasing surgeon's experience, there is significant decrease in operating time and increase in the number of pelvic lymph nodes removed. PMID- 10873412 TI - Immunoreactivity of recombinant squamous cell carcinoma antigen and leupin/SCCA 2: implications for tumor marker detection. AB - OBJECTIVES: Squamous cell carcinoma antigen (SCCA) is a member of the serpin superfamily, and has been used as a serological tumor marker for cervical squamous cell carcinomas. We have identified a closely related serpin gene, leupin (SCCA-2), which may be the fraction previously thought to be the acidic isoform of SCCA. The purpose of this study is to isolate the individual recombinant proteins, to examine their reactivity with current immunological detection methods, and to use a gene-specific method to examine their expression in the uterine cervix. METHODS: We have expressed and purified recombinant forms of SCCA and leupin individually. The proteins were characterized with respect to their isoelecric points and their reactivity with the monoclonal antibody from the current tumor marker diagnostic immunoassay (IMx SCC). Reverse transcription polymerase chain reaction (RT-PCR) with gene-specific primers was used to examine expression of both genes. RESULTS: Isoelectric focusing shows that leupin is the more acidic antigen with a determined pI for recombinant leupin (rLeupin) of 6.01, with rSCCA having a pI of 6.17. The IMx SCC monoclonal antibody recognized both rSCCA and rLeupin in immunoassays and immunoblots and both genes are expressed in normal cervix and in cervical carcinoma tissue. CONCLUSIONS: The findings from this study suggest that all previous clinical studies examining SCCA expression have used methodology that detects two gene products. The confirmation that leupin or SCCA-2 is the more acidic protein and that its expression is significantly elevated in cervical cancer suggests that this gene product may be the more important tumor marker. PMID- 10873413 TI - Inability of preoperative computed tomography scans to accurately predict the extent of myometrial invasion and extracorporal spread in endometrial cancer. AB - OBJECTIVE: The purpose of this study was to assess the value of computed tomography (CT) scans in predicting preoperatively the depth of invasion and extrauterine spread in patients with endometrial cancer. METHODS: The records of 54 patients with endometrial cancer who underwent a preoperative CT scan and surgical treatment (36 of whom had complete surgical staging) were reviewed. Final pathological findings were compared with those of the CT scan. The ability of the CT scan to detect the depth of invasion of the tumor into the myometrium and extrauterine spread was assessed. RESULTS: The sensitivity of CT scans at predicting the depth of myometrial invasion (none, inner half, outer half) and cervical and parametrial spread was 10, 9, and 17%, respectively, and sensitivity in predicting any degree of myometrial invasion, lymph node metastasis, adnexal involvement, and the presence of malignant cells in peritoneal cytology was 61, 50, 60 and 57%, respectively. CONCLUSION: CT scan has limited usefulness in determining the depth of myometrial invasion or extent of tumor spread in patients with endometrial cancer. Its routine preoperative use is difficult to justify. PMID- 10873414 TI - Primary peritoneal carcinoma presenting on routine papanicolaou smear. AB - BACKGROUND: Primary peritoneal carcinoma is an uncommon disease, characterized by peritoneal carcinomatosis without other identifiable primary tumor. It typically presents resembling ovarian cancer, with abdominal pain and distention and in an advanced stage. We report a unique presentation of this disease. CASE: A 76-year old woman had severe glandular dysplasia on a screening Papanicolaou smear. An ectocervical lesion was biopsied, revealing moderately differentiated adenocarcinoma. Cervical stenosis prevented sampling of the endocervix and endometrium. Colonoscopy and mammography did not reveal malignancy. Total abdominal hysterectomy with bilateral salpingo-oophrectomy was performed. Surgery revealed surface implants on the pelvic organs, with minimal involvement of the ovaries. Histologic examination revealed adenocarcinoma with papillary serous differentiation. Surgical and microscopic findings were consistent with a diagnosis of primary peritoneal carcinoma. CONCLUSION: Primary peritoneal carcinoma usually presents in a manner similar to that of ovarian cancer, but atypical presentations also occur. While a Papanicolaou smear suggestive of carcinoma usually represents a primary cervical malignancy, this case serves as a reminder that other metastatic malignancies should be considered. PMID- 10873415 TI - Dermatofibrosarcoma protuberans of the vulva: a case report and review of the literature. AB - OBJECTIVE: The purpose of this study is to describe the management of a patient with dermatofibrosarcoma protuberans (DFSP) of the vulva and to review the literature. METHODS: A 39-year-old was referred by a district hospital for incomplete excision of a vulval mass. The lesion involved the left labium major and measured 8 x 12 cm. The lesion was reexcised with a 3-cm margin of normal skin. RESULTS: The patient made an uneventful postoperative recovery. Histology confirmed a diagnosis of DFSP with clear margins. Immunohistochemistry was positive for CD34 glycoprotein. CONCLUSIONS: DFSP of the vulva is a rare fibrous tumor of intermediate grade malignancy, with a tendency for local recurrence. However, it rarely metastasizes. Management should be multidisciplinary and Mohs' micrographic surgery is generally advocated to ensure precise margin control. Survival rates range from 91 to 100%, and local recurrence rates of 20 to 49% have been reported. Therefore close follow-up is recommended. PMID- 10873416 TI - Gemcitabine is ineffective in recurrent, preirradiated cervical cancer. PMID- 10873417 TI - Spread of ovarian cancer after laparoscopic surgery. PMID- 10873418 TI - Fractal analysis. PMID- 10873419 TI - Reply PMID- 10873420 TI - Reply PMID- 10873421 TI - Good "negative results". PMID- 10873422 TI - CD1-Restricted T cells: T cells with a unique immunological niche. AB - Until recently, antigen presentation to T cells was defined only by proteins encoded within the MHC locus. That definition has now been expanded to include proteins encoded outside the MHC locus, most notably the CD1 family of proteins. The pathway of CD1-presented antigens diverges from that of MHC processing, indicating that the CD1 antigen-processing pathway may be complementary to the MHC pathways. The most surprising finding of the CD1 antigen-presenting system is that the antigens presented by CD1 are not peptides, but rather lipid and glycolipid in nature. The most compelling evidence for the role of CD1-restricted T cells in immune homeostasis stems from studies of mycobacterial infection and autoimmunity. These studies suggest that CD1-restricted T cells promote cell mediated immune responses to intracellular infection and protect against anti self responses. PMID- 10873423 TI - The antiviral 2',5'-oligoadenylate synthetase is persistently activated in type 1 diabetes. AB - Type 1 diabetes results from autoimmune destruction of the pancreatic beta-cells. Although viruses have been implicated as etiologic factors, specific pathogenic mechanisms have not been identified. Recently, increased attention has focused on the role of the innate antiviral defense system in directing adaptive immune responses. In this context, the pathogenesis of type 1 diabetes may involve an aberrant response to endogenous or exogenous viruses or their products. The family of 2',5' oligoadenylate synthetases (2', 5' AS) are IFN-alpha-inducible, RNA-dependent effector molecules in the antiviral defense system. We show that lymphocytic 2',5' AS activity is significantly increased in type 1 diabetes, both in recent-onset and in long-standing type 1 diabetes, and in diabetic twins from monozygotic twin pairs. The activity of 2',5' AS was not elevated in patients with type 2 diabetes or multiple sclerosis thus excluding hyperglycemia or autoimmunity per se as inducing upregulation of enzyme activity. In recent-onset diabetic patients, lymphocyte levels of protein kinase p68 and MxA, two other IFN alpha-inducible antiviral proteins, were similar to control levels. These data suggest that the increased 2',5' AS activity may reflect an aberrant response to viruses or RNA molecules originating from exogenous or endogenous sources. PMID- 10873424 TI - MHC-Class I antigens regulate both the function and the survival of human peripheral blood NK cells: role of endogenously secreted TNF-alpha. AB - The cytotoxic activity and survival of NK cells are negatively regulated in the presence of antibodies directed against the FCgammaRIII (CD16) surface receptor. The addition of MHC-class I monoclonal antibody (mAb) in combination with CD16 mAb resulted in a significant potentiation of the inhibition of NK cell cytotoxic function and both accelerated kinetics and synergistic induction of cell death in both resting and IL-2-treated human peripheral blood purified NK cells. The potentiating effect of class I MHC monoclonal antibody was specific as monoclonal antibodies directed against other cell surface antigens on NK (e.g., LFA-3, LFA 1, and CD56) had no effect. A correlation was observed between the levels of secreted TNF-alpha and the frequency of NK cell death and the addition of anti TNF-alpha antibody inhibited NK cell death. The levels of death promoting gene products such as the Fas receptor and the Fas ligand were upregulated in NK cells in the presence of anti-class I and anti-CD16 antibodies. However, anti-Fas antibodies did not block cell death. These findings demonstrate that MHC-class I antigens regulate both TNF-alpha secretion and cell death of anti-CD16 mAb treated NK cells. These results also suggest that MHC-class I antigens regulate the function and survival of activated NK cells. PMID- 10873425 TI - Intradermal granulocyte-macrophage colony-stimulating factor alters cutaneous antigen-presenting cells and differentially affects local versus distant immunization in humans. AB - We hypothesized that intradermal delivery of granulocyte-macrophage colony stimulating factor (GM-CSF) would alter the number and differentiation state of local antigen-presenting cells and thereby alter immunization strength at that site in humans. GM-CSF or placebo was administered intradermally on consecutive days prior to contact sensitization at that site. In GM-CSF-treated skin, epidermal CD1a(+)S100(+) Langerhans cells were reduced in number and had altered morphology, while the number of dermal CD1a(+), HLA-DR(+), and S100(+) cells was increased. In the deep dermis CD68(+) macrophages were increased. Expression of the APC activation markers CD40 and ICAM-1 was also increased in the dermis. Subjects were sensitized to DNCB through GM-CSF- or placebo-pretreated skin and to DPCP through untreated skin. Subjects immunized through GM-CSF-treated sites exhibited 64% greater elicitation responses to DNCB than placebo-treated subjects. GM-CSF-treated subjects also showed 43% lower responses to DPCP than placebo-treated subjects. The difference between DNCB (local) and DPCP (distant) responses was significantly greater for GM-CSF-treated subjects than for placebo responses (n = 8, P < 0.05). Therefore, local immunization site pretreatment with intradermal GM-CSF enhances immunization efficiency at that site. PMID- 10873426 TI - Bacterial superantigen staphylococcal enterotoxin B induces interstitial pneumonia in SCID mice reconstituted with peripheral blood mononuclear cells from collagen vascular disease patients. AB - To investigate whether superantigens induce interstitial pneumonia associated with collagen vascular disease (CVD), staphylococcal enterotoxin B (SEB) was intratracheally administered to SCID mice reconstituted with peripheral blood mononuclear cells (PBMCs) from CVD patients that suffered lung complications. Although a slight accumulation of inflammatory cells into the perivascular area was seen in the lungs of SCID mice injected with PBMCs from CVD patients or healthy donors, SEB administration significantly increased the severity of inflammation in the lungs of SCID mice that received CVD patient PBMCs. Furthermore, human leukocytes were detected by immunohistochemistry in the lungs of SCID mice that received SEB after reconstitution with PBMCs from CVD patients but not in other groups of SCID mice. CD45RO(+) memory T cells comprised the majority of infiltrating human leukocytes. These results suggest the possibility that external superantigens may induce the development of interstitial pneumonia in patients that have a genetic background predisposition to autoimmune disease. PMID- 10873427 TI - Interactions between the alpha(2)-adrenergic and the prostaglandin response in the regulation of macrophage-derived tumor necrosis factor. AB - Mediators such as prostaglandin E(2) (PGE(2)) and norepinephrine (NE) regulate macrophage (Mφ) responsiveness. Activation of alpha(2)-adrenergic receptors on Mφ potentiates lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNFalpha) production. PGE(2) inhibits LPS-stimulated TNFalpha production and gene expression, a response that can be desensitized by pretreatment of Mφ with PGE(2). We have determined that concomitant pretreatment of Mφ with PGE(2) and the alpha(2)-adrenergic agonist UK-14304 (UK) can prevent the PGE(2) induced desensitization. PGE(2) concentration-effect curves have been determined for the inhibition of LPS-stimulated TNFalpha production by murine peritoneal Mφ. The addition of 10 nM UK to Mφ in culture significantly shifts the PGE(2) concentration-effect curve to the right; pretreatment of Mφ with UK significantly shifts the PGE(2) concentration-effect curve to the left; and pretreatment with the cyclooxygenase inhibitor, indomethacin, increases the maximum response of PGE(2). Preincubation of Mφ with PGE(2) (0.5 h) followed by washing significantly shifts the subsequent PGE(2) concentration-effect curve to the right. Concomitant preincubation of Mφ with PGE(2) and UK prevents this rightward shift, an effect that is blocked by the alpha(2)-adrenergic receptor antagonist yohimbine. Northern blot analysis demonstrates that UK increases LPS-induced TNFalpha mRNA accumulation, and this is blocked by yohimbine, while PGE(2) decreases TNFalpha mRNA accumulation. Preincubation of Mφ with PGE(2) prevents PGE(2) regulation of TNFalpha mRNA, and concomitant preincubation of Mφ with PGE(2) and UK reverses this effect. These investigations support the role of NE as a regulator of Mφ TNFalpha production, a response that has functional interactions with Mφ sensitivity to PGE(2). PMID- 10873428 TI - Induction of global anergy rather than inhibitory Th2 lymphokines mediates posttrauma T cell immunodepression. AB - Depressed mitogen-induced IL-2 and IFN-gamma responses after severe mechanical or thermal injury are postulated to result from an expansion of Th2 lymphocytes with concomitant excessive production of IL-4 and/or IL-10. Here, we simultaneously assessed proliferation and Th1 (IFN-gamma) versus Th2 (IL-10, IL-4) lymphokine production in trauma patients' isolated T cells stimulated in a costimulation sufficient, antigen presenting cell independent system (anti CD3 + anti-CD4). T cells with depressed proliferation and IL-2 production simultaneously lost IL-4, IL-10, and IFN-gamma protein and mRNA responses. Exogenous IL-12 addition did not restore IFNgamma responses, but exogenous IL-2 partially restored IL-4, IFN gamma, and IL-10 production. Although initially partially restored by exogenous IL-2 or stimulation with PMA + ionomycin, patient T cells with persisting anergy progressively lost even these lymphokine and proliferative responses. Development of global T cell anergy was not a result of lost T cell viability or protein synthesis, since it corresponded to predominance of anergic T cells with upregulated expression of CD11b, but downregulated CD28 and CD3 expression. Thus, the subset of posttrauma patients whose isolated T cells become unresponsive experienced progressively worsening global anergy, mediated not by an increased production of Th2 lymphokines, but possibly by T cell incapacity to be activated through TCR triggering or Ca(2+) mobilization. PMID- 10873429 TI - Characterization of gammadelta T cells expressing CD158b, a killer cell inhibitory receptor, in a patient with chronic CD4(+) lymphocytopenia and disseminated Mycobacterium intracellulare infection. AB - A population of Vdelta1(+)Vgamma9(-) gammadelta T cells that represented almost the totality (84%) of circulating lymphocytes in a patient with chronic, non-HIV related, CD4 lymphocytopenia complicated by a disseminated Mycobacterium intracellulare infection was characterized. These gammadelta(+) T cells expressed a single killer inhibitory receptor (CD158b) and their phenotype (CD8(+)CD57(+)CD27(-)CD28(-)) indicated that, although CD45RA(+), they were not naive. However, the absence of large granular lymphocyte morphology, the impaired proliferative activity, the high susceptibility to apoptosis, and the total lack of cytotoxic ability suggested that these gammadelta cells were in a resting state. A high percentage of the cells did not harbor the CD11b integrin alpha chain and exhibited a decreased capability to bind endothelial cells. This defect might represent the mechanism whereby they remained trapped in the circulation. PMID- 10873430 TI - Mortality-rate crossovers and maximum lifespan in advantaged and disadvantaged populations: accelerated-mortality and sudden-death models. AB - One population is advantaged relative to another by our definition if its survival function is greater at all ages. A population has a lifespan maximum if there is an age at which its survival function becomes exactly zero. Earlier work concerned conditions under which the mortality-rate functions of advantaged and disadvantaged populations displaying lifespan maxima always crossed. Here two survival models of populations having lifespan maxima are presented in which mortality-rate crossings between advantaged and disadvantaged subpopulations may fail to appear. One, the accelerated-mortality model, has a continuous survival function; in the other, the sudden-death model, the survival function is discontinuous. Both differ from examples examined previously in that their mortality-rate functions become infinite at their lifespan maxima. PMID- 10873431 TI - Spontaneous secondary spiking in excitable cells. AB - Kepler & Marder (1993, Biol. Cybern.68, 209-214) proposed a model describing the electrical activity of a crab neuron in which a train of directly induced action potentials is sometimes followed by one or more spontaneous action potentials, referred to as spontaneous secondary spikes. We reduce their five-dimensional model to three dimensions in two different ways in order to gain insight into the mechanism underlying the spontaneous spikes. We then treat a slowly varying current as a parameter in order to give a qualitative explanation of the phenomenon using phase-plane and bifurcation analysis. We demonstrate that a three-dimensional model, consisting of a two-dimensional excitable system plus a slow inward current, is sufficient to produce the behaviour observed in the original model. The exact dynamics of the excitable system are not important, but the relative time constant and amplitude of the slow inward current are crucial. Using the numerical bifurcation analysis package AUTO (Doedel & Kernevez, 1986, AUTO: Software for Continuation and Bifurcation Problems in Ordinary Differential Equations. California Institute of Technology), we compute bifurcation diagrams using the maximum amplitude of the slow inward current as the bifurcation parameter. The full and reduced models have a stable resting potential for all values of the bifurcation parameter. At a critical value of the bifurcation parameter, a stable tonic firing mode arises via a saddle-node of periodics bifurcation. Whether or not the models can exhibit transient or continuous spontaneous spiking depends on their position in parameter space relative to this saddle-node of periodics. PMID- 10873432 TI - Lessons on pattern formation from planet WATOR. AB - It is well known that if reacting species experience unequal diffusion rates, then dynamics that lead to a constant steady state in a "well-mixed" environment can in a spatial setting lead to interesting patterns. In this paper, we focus on complementary pattern formation mechanisms that operate even when the diffusion rates are equal. In particular, we can say that when the mean-field ODE has an attracting periodic orbit then the stochastic spatial model will have large-scale spatial structures in equilibrium. We explore this mechanism in depth through the dynamics of the simulator WATOR. PMID- 10873433 TI - Potential mechanisms for the regulation of growth factor binding by heparin. AB - Heparin and heparan sulfate proteoglycans (HSPG) bind many soluble growth factors and this binding is now recognized as an important mechanism for modulation of cell activity. Fibroblast growth factor-2 (FGF-2) is one of the best characterized of the heparin-binding growth factors and it has been shown experimentally that heparin regulation of FGF-2 activity is dependent on the level of cell HSPG and the concentration of heparin. In this paper, we explore, using mathematical modeling, proposed mechanisms for heparin regulation and determine how they impact FGF receptor binding. We demonstrate that the experimentally observed receptor binding phenomena can be reproduced if cells (1) express heparin-binding cell surface molecules and if either (2) these heparin binding sites are FGFR and bind heparin and FGF-2-heparin complexes or (3) are surface molecules able to bind FGF-2 and couple with FGF-2 receptors to form high affinity FGF-2-bound surface complexes. The ability of heparin to directly interact with the FGFR and bind FGF-2 in the absence of this coupling function was not sufficient to explain heparin activity. These findings have implications with regard to regulation of heparin-binding growth factors and could help guide the development of highly specific growth regulatory molecules through specific regulation by heparin and HSPG. PMID- 10873434 TI - Antibacterial activity of flavonoids against methicillin-resistant Staphylococcus aureus strains. AB - An experimental and theoretical study was performed on the anti-staphylococcal activity of 18 natural and synthetic flavonoids against methicillin-resistant Staphylococcus aureus strains. The analysed flavonoids belong to three well differentiated structural patterns: chalcones, flavanones and flavones. The quantitative analysis of the anti-staphylococcal activity of the compounds was carried out by determining their percent inhibition degree. The hierarchical cluster analysis method was used to analyse the anti-MRSA activity of the compounds. With this methodology, the flavonoids were classified into four groups according to their anti-staphylococcal activity (high, sufficient, intermediate and low). The carbonylic region is of importance because it is part of the bioactive region inducing anti-MRSA activity in the flavonoid molecules. The introduction of OH groups in positions 2' of chalcones and 5 of flavanones (or flavones) increases flavonoid activity, while the OCH(3)groups produce the reverse effect. Using the experimental anti-MRSA activity data of flavonoids and six quantum chemical parameters calculated by means of the AM1 semiempirical molecular orbital method, a very good quantitative structure-activity relationship was obtained (confidence range: 95%; significance level for tests: 0.05; correlation coefficient=0.9842). The selected parameters explain 96.86% of the percent inhibition degree. The obtained relation is consistent with the conclusions formulated in this paper and serves as a theoretical support for some of them. Finally, it is concluded that the flavonoids chalcone, 2'(OH)-chalcone, 2',4'(OH)(2)-chalcone and 2',4(OH)(2)-chalcone might constitute promising therapeutic agents against infections with methicillin-resistant S. aureus strains. PMID- 10873435 TI - A mathematical model for assessing changes in neurofilament protein levels in neurites and cell bodies of differentiating neuroblastoma cells. AB - A mathematical model which allows the calculation of the level of neurofilament protein in the cell body (x) and in the neurites (y) of differentiating SK-N-SH cells is presented. The model considers the changes in cell number (proliferating cells) and the number of cells with neurites (differentiating cells). It takes into account the fact that (i) when cells are cultured in differentiating conditions, an increase in cell number is initially observed and (ii) in a non synchronized population of differentiating cells, the length of neurite extended by individual cells varies within the population. Total neurofilament protein levels in a population of cells were measured by enzyme-linked immunoabsorbant assay and application of the model to the data allowed values for x and y to be calculated. The validity of the model is supported by the fact that the predicted total neurofilament protein levels are highly correlated with the experimentally derived neurofilament protein levels. The model should be of use in temporal studies of cytoskeletal proteins involved in neuronal growth/differentiation and also in studies in which the system is a target of toxic insult. PMID- 10873436 TI - A single theory explains two empirical laws applicable to plant populations. AB - Two empirical laws, formulated independently, are known to be well satisfied in even-aged plant monocultures. One relates yield to plant density in different plant populations, and the other relates cumulative plant mass to cumulative plant number from the largest individual within a population. In this paper, we construct a mathematical model of plant growth under asymmetric competition between individuals in a population, where large individuals grow larger than small individuals because they pre-empt resources, especially light. The model categorizes influences on the growth of a plant into those from individuals larger than itself, and those from all individuals in the population. We derive the two laws from our model. Thus, individual growth, determined by asymmetric interaction between individuals in a population, can explain the two different laws relating to plant populations. PMID- 10873437 TI - A class of flow bifurcation models with lognormal distribution and fractal dispersion. AB - We report a quantitative analysis of a simple dichotomous branching tree model for blood flow in vascular networks. Using the method of moment-generating function and geometric Brownian motion from stochastic mathematics, our analysis shows that a vascular network with asymmetric branching and random variation at each bifurcating point gives rise to an asymptotic lognormal flow distribution with a positive skewness. The model exhibits a fractal scaling in the dispersion of the regional flow in the branches. Experimentally measurable fractal dimension of the relative dispersion in regional flow is analytically calculated in terms of the asymmetry and the variance at local bifurcation; hence the model suggests a powerful method to obtain the physiological information on local flow bifurcation in terms of flow dispersion analysis. Both the fractal behavior and the lognormal distribution are intimately related to the fact that it is the logarithm of flow, rather than flow itself, which is the natural variable in the tree models. The kinetics of tracer washout is also discussed in terms of the lognormal distribution. PMID- 10873438 TI - Multiplicative moments and measures of persistence in ecology. AB - Ecologists and epidemiologists have begun focusing on demographic stochasticity and spatial heterogeneity as important biological factors. With high-powered computers simulation of such systems is a common modelling technique; however we lack a detailed understanding of the processes involved. Moment closure approximations provide a simple method which can be used to capture the main features of a wide variety of stochastic models and to gain a more intuitive understanding. In this paper we give an alternative variation based on multiplicative moments which is equivalent to taking a novel third-order cumulant approximation. The differential equations for these multiplicative moments are far more robust than their additive counterparts. We use this technique to consider the behaviour and persistence of finite metapopulations for two common ecological systems. PMID- 10873440 TI - The ligand-receptor-G-protein ternary complex as a GTP-synthase. steady-state proton pumping and dose-response relationships for beta -adrenoceptors. AB - Steady-state solutions are developed for the rate of G alpha.GTP production in a synthase model of the ligand-receptor-G-protein ternary complex activated by a ligand-receptor proton pumping mechanism. The effective rate, k(31), defining the proton transfer, phosphorylation and G alpha.GTP release is a controlling rate of the synthase in the presence of a ligand with an efficient mode of signal activation, the ligand-receptor interaction taking place under effectively equilibrium conditions. The composite rate, however, becomes an amplifying factor in any dose-response relationship. The amplification is a triple product of the rate, k(31), the equilibrium constant associated with the activation of the proton signal, K(act)and the fraction of agonist conformer transmitting the signal, f(*). Where the rate of activation of the proton signal becomes critically inefficient, the rate of activation, k(act 1)replaces k(31)K(act). A correlation between beta(1)-adrenergic receptor-stimulated GDP release and adenylate cyclase activation shows that this correlation is not unique to an exchange reaction. Within the initiating Tyr-Arg-Tyr receptor proton shuttle mechanism, the position of Arg(r156) paralleldictates the high-(R(p)) and low (R(u)) ligand-binding affinities. These states are close to R(*)and R(0)of the equilibrium model (De Lean et al., 1980, J. Biol. Chem.255, 7108-7117). An increased rate of hydrogen ion diffusion into a receptor mutant can give rise to constitutive activity while increased rates of G-protein release and changes in receptor state balance can contribute to the resultant level of action. Constitutive action will arise from a faster rate of G-protein release alone if proton diffusion in the wild-type receptor contributes to a basal level of G protein activation. Competitive ligand-receptor occupancy for constitutive mutants shows that, where the rate of G-protein activation from the proportion of ligand-occupied receptors is less than the equivalent rate that would be generated from this fraction by proton diffusion, inverse agonism will occur. Rate-dependent dose-responses developed for the proposed synthase mechanism give explicit definition to the operational model for partial agonism (Black & Leff, 1983, Proc. Roy. Soc. Lond. B220, 141-162). When comparable ligands have effectively identical conformational states at the transition state for signal activation, the antagonist component of the binding "in vitro" can be derived by multiplying the apparent binding constant by (1-e) where e is the maximum stimulatory response. This component should be consistent throughout the tissues. PMID- 10873441 TI - Modeling the differential fitness of cyanobacterial strains whose circadian oscillators have different free-running periods: comparing the mutual inhibition and substrate depletion hypotheses. AB - In a recent experimental study, Ouyang et al. (1998, Proc. Natl. Acad. Sci. U.S.A.95, 8660-8664) have shown that, in direct competition, cyanobacterial strains whose circadian clocks have free-running periods (FRPs) which match the period of an imposed light/dark (LD) cycle exclude strains whose FRPs are out of resonance with the LD cycle. These differences in competitive fitness are observed despite the lack of measurable differences in monoculture growth rates between the strains. Here we show that the experimental results are consistent with a mathematical model in which cells rhythmically produce a metabolic inhibitor to which they display a sensitivity modulated by their circadian rhythm. We argue that models in which there is a circadian modulation of nutrient uptake kinetics cannot account for the results of these experiments. We discuss possible experiments to further characterize this phenomenon. The experimental protocol we propose can be used to distinguish between mutual inhibition and substrate depletion as underlying causes of the competitive advantage of circadian resonance. PMID- 10873442 TI - Aspartyl tRNA-synthetase from Escherichia coli: flexibility and adaptability to the substrates. AB - The crystal structure of aspartyl-tRNA synthetase from Escherichia coli has been determined to a resolution of 2.7 A. The structure is compared to the same enzyme co-crystallized with tRNA(Asp) and containing aspartyl adenylate or ATP. The asymmetric unit contains three monomers of the enzyme. While most parts of the protein show no significant differences in the three monomers, a few regions cannot be superimposed. Those regions are characterized by a high B-factor, and consist mostly of loops that make contacts with the tRNA in the complexes. The flexibility of the protein is seen at a global level, by the observation of a 10 to 15 degrees rotation of the N-terminal and insertion domains upon tRNA binding, and at the level of the individual amino acid residues, by main-chain and side chain rearrangements. In contrast to these induced-fit conformational changes, a few residues essential for the tRNA anticodon or aspartyl-adenylate recognition exist in a predefined conformation, ensured by specific interactions within the protein. PMID- 10873443 TI - C-terminal domains of Escherichia coli topoisomerase I belong to the zinc-ribbon superfamily. AB - Detection of remote evolutionary connections is increasingly difficult with sequence and structural divergence. A combination of sequence and structural analysis, in which statistically supported sequence similarity had a crucial impact, revealed that Escherichia coli topoisomerase I C-terminal fragment is evolutionarily related to the three tetracysteine zinc-binding domains of the enzyme. Spatial structure analysis of this C-terminal fragment indicates that it consists of two structurally similar domains and suggests homology between them. Sequence similarity between the zinc-binding domains of type Ia topoisomerases and transcription regulators of known spatial structure helps to conclude that E. coli topo I contains five copies of a zinc ribbon domain at the C terminus. Two of these domains, corresponding to the C-terminal fragment, lost their cysteine residues and are probably not able to bind zinc. Present analyses lead to the classification of the C-terminal fragment of E. coli topoisomerase I as a member of zinc ribbon superfamily, despite the absence of zinc-binding sites. PMID- 10873439 TI - Quantifying the dynamics of prion infection: a bifurcation analysis of Laurent's model. AB - Laurent (1996a, Medecine/sciences12, 774-785; 1996b, Biochem. J.318, 35-39; 1998, Bio-phys. Chem.72, 211-222) proposed a model for the dynamics of diseases of the central nervous system caused by prions. It is based on the protein-only hypothesis (Prusiner et al., 1981, Proc. Natl. Acad. Sci. U.S.A.78, 6675-6679), which assumes that infection can be spread by particular proteins (prions) that can exist in two forms that share the same sequence, but have a different structure. The normal form is harmless, while the infectious isoform of the prion protein catalyses a transconformation from the native isoform to itself within a specialized compartment of the brain cells. This paper systematically explores the model behavior with the aim of quantifying the fundamental parameters characterizing the dynamics of prion infection. To this end we use data from the literature to fix orders of magnitude for the rates of synthesis and degradation of the native form of prion protein and for the shape of the autocatalytic function. The dynamical behavior is classified with respect to two unknown parameters (bifurcation analysis): the rate of spontaneous transconformation and the rate of output of the infectious isoform from the specialized compartment. We thus find that the bistability properties evidenced by Laurent are confined to a certain range of parameters and that permanent oscillations of the two isoforms concentrations are possible. The bifurcation analysis allows us to estimate approximate ranges for the values of the two unknown parameters and consequently to derive incubation times and compare them with actual data for hamster. Also, our study predicts that the output rate of the infectious isoform is relatively insensitive to variations of model parameters. PMID- 10873444 TI - Rapid induction of nuclear transcripts and inhibition of intron decay in response to the polymerase II inhibitor DRB. AB - The transcriptional inhibitor 5, 6-dichloro-1-beta-d-ribofuranosylbenzimidazole (DRB) is an adenosine analog that has been shown to cause premature transcriptional termination and thus has been a useful tool to identify factors important for transcriptional elongation. Here, we establish an efficient system for studying DRB-sensitive steps of transcriptional elongation. In addition, we establish two novel effects of DRB not previously reported: intron stabilization and the induction of long transcripts by a mechanism other than premature termination. We found that DRB had a biphasic effect on T-cell receptor-beta (TCRbeta) transcripts driven by a tetracycline (tet)-responsive promoter in transfected HeLa cells. In the first phase, DRB caused a rapid decrease (within five minutes) of pre-mRNA and its spliced intron (IVS1(Cbeta1)), consistent with the known ability of DRB to inhibit transcription. In the second phase (which began ten minutes to two hours after treatment, depending on the dose), DRB dramatically increased the levels of IVS1(Cbeta1)-containing transcripts by a mechanism requiring de novo RNA synthesis. DRB induced the appearance of short 0.4 to 0.8 kb TCRbeta transcripts in vivo, indicating DRB enhances premature transcriptional termination. A approximately 475 nt prematurely terminated transcript (PT) was characterized that terminated at an internal poly(A) tract in the intron IVS1(Cbeta1). We identified three other effects of DRB. First, we observed that DRB induced the appearance of heterodisperse TCRbeta transcripts that were too long ( approximately 1 kb to >8 kb) to result from the type of premature termination events previously described. Their production was not promoter-specific, as we found that long transcripts were induced by DRB from both the tet-responsive and beta-actin promoters. Second, DRB upregulated full length normal-sized c-myc mRNA, which provided further evidence that DRB has effects besides regulation of premature termination. Third, DRB stabilized lariat forms of the intron IVS1(Cbeta1), indicating that DRB exerts post-transcriptional actions. We propose that our model system will be useful for elucidating the factors that regulate RNA decay and transcriptional elongation in vivo. PMID- 10873445 TI - Dissection of bacteriophage lambda site-specific recombination using synthetic peptide combinatorial libraries. AB - A wide variety of tools have been used to dissect biochemical pathways, inhibitors being chief among them. Combinatorial approaches have made the search for inhibitors much more efficient. We have applied such an approach to identify hexapeptides which inhibit different steps in a site-specific recombination reaction mediated by the bacteriophage lambda integrase protein. Integrase's mechanism is still incompletely understood, in large part because several pathway intermediates remain hard to isolate. Integrase-catalyzed recombination is very efficient, but if blocked, it is highly reversible to substrates; this combination makes some intermediates exceedingly transient. We have used synthetic peptide combinatorial libraries to screen for hexapeptides that affect the recombination pathway at different stages, and have identified two families of peptides: one probably blocks DNA cleavage, the other may stabilize the Holliday junction intermediates. These peptides do not resemble parts of integrase or any of the other helper functions in the pathway. The deconvolution of hexapeptide libraries based both on inhibition of an enzymatic reaction as well as on accumulation of reaction intermediates is a novel approach to finding useful tools for dissecting a biochemical pathway. PMID- 10873446 TI - Peptide inhibitors of DNA cleavage by tyrosine recombinases and topoisomerases. AB - The study of biochemical pathways requires the isolation and characterization of each and every intermediate in the pathway. For the site-specific recombination reactions catalyzed by the bacteriophage lambda tyrosine recombinase integrase (Int), this has been difficult because of the high level of efficiency of the reaction, the highly reversible nature of certain reaction steps, and the lack of requirements for high-energy cofactors or metals. By screening synthetic peptide combinatorial libraries, we have identified two related hexapeptides, KWWCRW and KWWWRW, that block the strand-cleavage activity of Int but not the assembly of higher-order intermediates. Although the peptides bind DNA, their inhibitory activity appears to be more specifically targeted to the Int-substrate complex, insofar as inhibition is resistant to high levels of non-specific competitor DNA and the peptides have higher levels of affinity for the Int-DNA substrate complex than for DNA alone. The peptides inhibit the four pathways of Int-mediated recombination with different potencies, suggesting that the interactions of the Int enzyme with its DNA substrates differs among pathways. The KWWCRW and KWWWRW peptides also inhibit vaccinia virus topoisomerase, a type IB enzyme, which is mechanistically and structurally related to Int. The peptides differentially affect the forward and reverse DNA transesterification steps of the vaccinia topoisomerase. They block formation of the covalent vaccinia topoisomerase-DNA intermediate, but have no apparent effect on DNA religation by preformed covalent complexes. The peptides also inhibit Escherichia coli topoisomerase I, a type IA enzyme. Finally, the peptides inhibit the bacteriophage T4 type II topoisomerase and several restriction enzymes with 2000-fold lower potency than they inhibit integrase in the bent-L pathway. PMID- 10873447 TI - Aromatic amino acids in region 2.3 of Escherichia coli sigma 70 participate collectively in the formation of an RNA polymerase-promoter open complex. AB - Formation of an initiation-competent RNA polymerase-promoter complex involves DNA melting over a region of about 12 base-pairs, which includes the start site of transcription, thus enabling the template strand to base-pair with the initiating nucleoside triphosphates. By studying the effects of alanine substitutions, we have investigated the role of the aromatic amino residues in the Escherichia coli sigma(70) conserved region 2.3 in promoter strand separation. The resulting mutants were assessed for their activity in vivo in the context of a sigma(70)/sigma(32) hybrid sigma factor that could be targeted to a specific hybrid promoter in the cell. All substitutions lead to an at least twofold reduction in expression of the hybrid promoter-driven reporter gene. The in vitro assay of single substitutions indicated cold sensitivity similar to that previously observed with analogous substitutions in Bacillus subtilis sigma(A). Kinetic assays showed that these substitutions slowed the rate of open complex formation at 37 degrees C as well. RNA polymerase reconstituted with a sigma(70) containing multiple alanine substitutions readily binds to promoter DNA, but then proceeds slowly beyond the first intermediate complex on the pathway to formation of the transcription-competent complex. These data demonstrate that together the aromatic residues in region 2.3 of E. coli sigma(70) ensure that DNA strand separation proceeds efficiently, even if no individual residue may be essential for accomplishment of the process. PMID- 10873448 TI - Construction of new ribozymes requiring short regulator oligonucleotides as a cofactor. AB - A hairpin loop and an oligonucleotide bound to the loop form one-half of the pseudoknot structure. We have designed an allosteric hammerhead ribozyme, which is activated by the introduction of this motif by using a short complementary oligonucleotide as a cofactor. Stem II of the hammerhead ribozyme was substituted with a non-self-complementary loop sequence (loop II) to abolish the cleavage activity. The new ribozyme had almost no cleavage activity of the target RNA. However, it exhibited the cleavage activity in the presence of a cofactor oligoribonucleotide, which is complementary to loop II of the ribozyme. The activity is assumed to be derived from the formation of a pseudo-stem structure between the cofactor oligonucleotide and loop II. The structure including the loop may be similar to the pseudo-half-knot structure. The activation efficiencies of the cofactor oligonucleotides were decreased as the lengths of the oligonucleotides increased, and the ribozyme with a longer loop II was more active than that with a short loop II. Oligoribonucleotides with 3'-dangling purine bases served as efficient cofactors of the ribozyme, and a 2'-O methyloligonucleotide enhanced the cleavage activity of the ribozyme most efficiently, by as much as about 750-fold as compared with that in the absence of the oligonucleotide. Cofactor oligonucleotides with a cytidine base at the 3'-end also activated a ribozyme with the G10.1.G11.1 mutation, which eliminates the cleavage activity in the wild-type. The binding sites of the oligonucleotide were identified by photo-crosslinking experiments and were found to be the predicted sites in the loop. This is the first report of a design aimed at positively controlling the activity of ribozymes by employing a structural motif. This method can be applied to control the activities of other functional RNAs with hairpin loops. PMID- 10873449 TI - Probing the Escherichia coli transcriptional activator MarA using alanine scanning mutagenesis: residues important for DNA binding and activation. AB - The MarA transcriptional activator binds to a 20 bp asymmetric degenerate sequence (marbox) located at different positions and orientations within the promoters of the genes of the Escherichia coli mar regulon. Solution of the MarA marbox X-ray crystallographic structure suggested the presence of base-specific and non-specific interactions between the marbox and two helix-turn-helix (HTH) motifs on the monomeric MarA. Here, we use alanine-scanning mutagenesis and DNA retardation analysis to: (i) evaluate the contacts between MarA and the marboxes of five differently configured mar regulon promoters; (ii) assess the role of conserved hydrophobic amino acid residues for MarA activity; and (iii) identify residues required for RNA polymerase activation. These analyses revealed that the phosphate-backbone contacts and hydrogen bonds with the bases of the marbox are more significant for DNA binding than are the van der Waals interactions. While both N and C-terminal HTH motifs make essential contributions to binding site affinity, MarA is more sensitive to alterations in the N-terminal HTH. In a similar way, the activity of MarA is more sensitive to alterations in the hydrophobic core of this HTH. Solvent-exposed amino acid residues located at many positions on the MarA surface are important for activity. Some of these residues affect activity on all promoters and thus, are implicated in maintaining MarA structure whereas several solvent-exposed amino acids not involved in DNA binding were important for MarA activity on specific promoters. The pattern of activation defects defined a class II promoter-specific activating region. However, a localized class I activating region was not apparent. These results suggest that MarA activates transcription by at least two distinct mechanisms. Furthermore, the important role of phosphate contacts in marbox affinity suggests that indirect readout contributes to binding site recognition by MarA. PMID- 10873451 TI - The aquaporin sidedness revisited. AB - Aquaporins are transmembrane water channel proteins, which play important functions in the osmoregulation and water balance of micro-organisms, plants, and animal tissues. All aquaporins studied to date are thought to be tetrameric assemblies of four subunits each containing its own aqueous pore. Moreover, the subunits contain an internal sequence repeat forming two obversely symmetric hemichannels predicted to resemble an hour-glass. This unique arrangement of two highly related protein domains oriented at 180 degrees to each other poses a significant challenge in the determination of sidedness. Aquaporin Z (AqpZ) from Escherichia coli was reconstituted into highly ordered two-dimensional crystals. They were freeze-dried and metal-shadowed to establish the relationship between surface structure and underlying protein density by electron microscopy. The shadowing of some surfaces was prevented by protruding aggregates. Thus, images collected from freeze-dried crystals that exhibited both metal-coated and uncoated regions allowed surface relief reconstructions and projection maps to be obtained from the same crystal. Cross-correlation peak searches along lattices crossing metal-coated and uncoated regions allowed an unambiguous alignment of the surface reliefs to the underlying density maps. AqpZ topographs previously determined by AFM could then be aligned with projection maps of AqpZ, and finally with human erythrocyte aquaporin-1 (AQP1). Thereby features of the AqpZ topography could be interpreted by direct comparison to the 6 A three-dimensional structure of AQP1. We conclude that the sidedness we originally proposed for aquaporin density maps was inverted. PMID- 10873450 TI - A single amino acid substitution in the C terminus of OmpR alters DNA recognition and phosphorylation. AB - In bacteria and lower eukaryotes, adaptation to changes in the environment is often mediated by two-component regulatory systems. Such systems provide the basis for chemotaxis, nitrogen and phosphate regulation and adaptation to osmotic stress, for example. In Escherichia coli, the sensor kinase EnvZ detects a change in the osmotic environment and phosphorylates the response regulator OmpR. Phospho-OmpR binds to the regulatory regions of the porin genes ompF and ompC, and alters their expression. Recent evidence suggests that OmpR functions as a global regulator, regulating additional genes besides the porin genes. In this study, we have characterized a previously isolated OmpR2 mutant (V203M) that constitutively activates ompF and fails to express ompC. Because the substitution was located in the C-terminal DNA-binding domain, it had been assumed that the substitution would not affect phosphorylation of the N-terminal domain of OmpR. Our results indicate that this substitution completely eliminates phosphorylation by a small phosphate donor, acetyl phosphate, but not phosphorylation by the kinase EnvZ. The mutant OmpR has altered dephosphorylation kinetics and altered binding affinities to both ompF and ompC sites compared to the wild-type. Thus, a single amino acid substitution in the C-terminal DNA-binding domain has dramatic effects on the N-terminal phosphorylation domain. Most strikingly, we have identified a single base change in the OmpR binding site of ompC that restores high-affinity binding activity by the mutant. We interpret our results in the context of a model for porin gene expression. PMID- 10873452 TI - Three-dimensional reconstruction of transcription termination factor rho: orientation of the N-terminal domain and visualization of an RNA-binding site. AB - The Escherichia coli rho transcription termination protein is a hexameric helicase, and is believed to function by separating an RNA-DNA hybrid. Unlike hexameric DNA helicases, where a single strand of DNA passes through the central channel, it has been proposed that the RNA wraps around the outside of the ring. We have generated a three-dimensional reconstruction of rho, and localized a tRNA molecule bound to the primary RNA-binding site to the outside of the ring. An atomic structure of the N-terminal domain of rho fits into our reconstruction uniquely, with the residues involved in RNA-binding on the outside of the ring. Although rho shares a common structural core with the F1-ATPase and other hexameric helicases, there has been a divergence in function due to rho's N terminal domain, which has no homology to other helicases. PMID- 10873453 TI - Reconstruction of protein form with X-ray solution scattering and a genetic algorithm. AB - We have reconstructed, from experimental approximately 2 nm resolution X-ray solution scattering profiles, the corresponding shapes and sizes of myoglobin, troponin C, spermadhesin PSP-I/PSP-II, chymotrypsinogen A, superoxide dismutase, ovalbumin, tubulin, nitrite reductase, catalase, the structural change of troponin C upon dissociation of the two high affinity Ca(2+), and the solution model structure of a tandem pair of fibronectin type III cytoplasmic domains of integrin alpha6beta4 before determination of its crystal structure. To this purpose we have designed a new genetic algorithm which gradually explores a discrete search space and evolves convergent models made of several hundred beads (down to 0.3 nm radius) best fitting the scattering profile upon Debye calculation, without geometrical constraints or penalty for loose beads. This is a procedure of effective numerical transformation of the one-dimensional scattering profiles into three-dimensional model structures. The number of beads in models is correlated with the protein molecular mass (with one exception). The shape and approximate dimensions of each protein have been retrieved by a set of ten solution models, essentially superimposable with the available crystal structures. PMID- 10873454 TI - A revised mechanism for the alkaline phosphatase reaction involving three metal ions. AB - Here, X-ray crystallography has been used to investigate the proposed double in line displacement mechanism of Escherichia coli alkaline phosphatase in which two of the three active-site metal ions have a direct role in catalysis. Two new X ray crystal structures of the wild-type enzyme in the absence and presence of inorganic phosphate have been refined at 1.75 A to final working R-factors of 15.4% and 16.4%, respectively. In the refinement of both structures, residues in the active sites were treated anisotropically. The ellipsoids resulting from the partial anisotropic refinement show a clear route for the binding and release of substrate/product. In addition, a direct comparison of the refined structures with and without phosphate reveal a strong correlation between the occupancy of the third metal-binding site and the conformation of the Ser102 nucleophile. These findings clarify two important and unresolved aspects of the previously proposed catalytic mechanism, how Ser102 is activated for nucleophilic attack and why a magnesium ion in the third metal site is required for catalysis. Analysis of these results suggest that three metal-ion assisted catalysis is a more accurate description of the mechanism of the alkaline phosphatase reaction. A revised mechanism for the catalytic reaction of alkaline phosphatase is proposed on the basis of the two new X-ray crystal structures reported. PMID- 10873455 TI - The free yeast aspartyl-tRNA synthetase differs from the tRNA(Asp)-complexed enzyme by structural changes in the catalytic site, hinge region, and anticodon binding domain. AB - Aminoacyl-tRNA synthetases catalyze the specific charging of amino acid residues on tRNAs. Accurate recognition of a tRNA by its synthetase is achieved through sequence and structural signalling. It has been shown that tRNAs undergo large conformational changes upon binding to enzymes, but little is known about the conformational rearrangements in tRNA-bound synthetases. To address this issue the crystal structure of the dimeric class II aspartyl-tRNA synthetase (AspRS) from yeast was solved in its free form and compared to that of the protein associated to the cognate tRNA(Asp). The use of an enzyme truncated in N terminus improved the crystal quality and allowed us to solve and refine the structure of free AspRS at 2.3 A resolution. For the first time, snapshots are available for the different macromolecular states belonging to the same tRNA aminoacylation system, comprising the free forms for tRNA and enzyme, and their complex. Overall, the synthetase is less affected by the association than the tRNA, although significant local changes occur. They concern a rotation of the anticodon binding domain and a movement in the hinge region which connects the anticodon binding and active-site domains in the AspRS subunit. The most dramatic differences are observed in two evolutionary conserved loops. Both are in the neighborhood of the catalytic site and are of importance for ligand binding. The combination of this structural analysis with mutagenesis and enzymology data points to a tRNA binding process that starts by a recognition event between the tRNA anticodon loop and the synthetase anticodon binding module. PMID- 10873456 TI - Phosphorylated and dephosphorylated structures of pig heart, GTP-specific succinyl-CoA synthetase. AB - Succinyl-CoA synthetase (SCS) catalyzes the reversible phosphorylation/dephosphorylation reaction:???rm succinyl ?hbox ? ?CoA+NDP+P_i?leftrightarrow succinate+CoA+NTP??where N denotes adenosine or guanosine. In the course of the reaction, an essential histidine residue is transiently phosphorylated. We have crystallized and solved the structure of the GTP-specific isoform of SCS from pig heart (EC 6.2.1.4) in both the dephosphorylated and phosphorylated forms. The structures were refined to 2.1 A resolution. In the dephosphorylated structure, the enzyme is stabilized via coordination of a phosphate ion by the active-site histidine residue and the two "power" helices, one contributed by each subunit of the alphabeta-dimer. Small changes in the conformations of residues at the amino terminus of the power helix contributed by the alpha-subunit allow the enzyme to accommodate either the covalently bound phosphoryl group or the free phosphate ion. Structural comparisons are made between the active sites in these two forms of the enzyme, both of which can occur along the catalytic path. Comparisons are also made with the structure of Escherichia coli SCS. The domain that has been shown to bind ADP in E. coli SCS is more open in the pig heart, GTP-specific SCS structure. PMID- 10873457 TI - NMR characterization of residual structure in the denatured state of protein L. AB - Triple-resonance NMR experiments were used to assign the (13)C(alpha), (13)C(beta), (15)N and NH resonances for all the residues in the denatured state of a destabilized protein L variant in 2 M guanidine. The chemical shifts of most resonances were very close to their random coil values. Significant deviations were observed for G22, L38 and K39; increasing the denaturant concentration shifted the chemical shifts of these residues towards theory random coil values. Medium-range nuclear Overhauser enhancements were detected in segments corresponding to the turn between the first two strands, the end of the second strand through the turn between the second strand and the helix, and the turn between the helix and the third strand in 3D H(1), N(15)-HSQC-NOESY-HSQC experiments on perdeuterated samples. Longer-range interactions were probed by measuring the paramagnetic relaxation enhancement produced by nitroxide spin labels introduced via cysteine residues at five sites around the molecule. Damped oscillations in the magnitude of the paramagnetic relaxation enhancement as a function of distance along the sequence suggested native-like chain reversals in the same three turn regions. The more extensive interactions within the region corresponding to the first beta-turn than in the region corresponding to the second beta-turn suggests that the asymmetry in the folding reaction evident in previous studies of the protein L folding transition state is already established in the denatured state. PMID- 10873458 TI - New structural insights into the molecular deciphering of mycobacterial lipoglycan binding to C-type lectins: lipoarabinomannan glycoform characterization and quantification by capillary electrophoresis at the subnanomole level. AB - Lipoarabinomannans are key molecules of the mycobacterial envelopes involved in many steps of tuberculosis immunopathogenesis. Several of the biological activities of lipoarabinomannans are mediated by their ability to bind human C type lectins, such as the macrophage mannose receptor, the mannose-binding protein and the surfactant proteins A and D. The lipoarabinomannan mannooligosaccharide caps have been demonstrated to be involved in the binding to the lectin carbohydrate recognition domains. We report an original analytical approach, based on capillary electrophoresis monitored by laser-induced fluorescence, allowing the absolute quantification, in nanomole quantities of lipoarabinomannan, of the number of mannooligosaccharide units per lipoarabinomannan molecule. Moreover, this analytical approach was successful for the glycosidic linkage determination of the mannooligosaccharide motifs and has been applied to the comparative analysis of parietal and cellular lipoarabinomannans of Mycobacterium bovis BCG and Mycobacterium tuberculosis H37Rv, H37Ra and Erdman strains. Significant differences were observed in the amounts of the various mannooligosaccharide units between lipoarabinomannans of different strains and between parietal and cellular lipoarabinomannans of the same strain. Nevertheless, no relationship was found between the number of mannooligosaccharide caps and the virulence of the corresponding strain. The results of the present study should help us to gain more understanding of the molecular basis of lipoarabinomannan discrimination in the process of binding to C-type lectins. PMID- 10873459 TI - Determination of intramolecular distance distribution during protein folding on the millisecond timescale. AB - A method for determination of transient (on the millisecond timescale) intramolecular distance distributions (IDDs) by time-resolved dynamic non radiative excitation energy transfer measurements was developed. The time-course of the development of the IDD between residues 73 and 203 in the CORE domain of Escherichia coli adenylate kinase throughout refolding from the GuHCl-induced denatured state was determined. The mean of the apparent IDD reduced to a value close to its magnitude in the native protein, within 2 ms (the dead-time of the instrument). At that time the width of that distribution was rather large (16+/-2 A). The large width implies that the intramolecular diffusion coefficient of the labeled segment does not exceed 10(-7) cm(2)/second. In a second slower phase of the refolding transition, the width was reduced to its native value (6+/-4 A). PMID- 10873460 TI - The thermodynamic stability of the proteins of the ccd plasmid addiction system. AB - The two opponents, toxin (CcdB, LetB or LetD, protein G, LynB) and antidote (CcdA, LetA, protein H, LynA), in the plasmid addiction system ccd of the F plasmid were studied by different biophysical methods. The thermodynamic stability was measured at different temperatures combining denaturant and thermally induced unfolding. It was found that both proteins denature in a two state equilibrium (native dimer versus unfolded monomer) and that CcdA has a significantly lower thermodynamic stability. Using a numerical model, which was developed earlier by us, and on the basis of the determined thermodynamic parameters the concentration dependence of the denaturation transition temperature was obtained for both proteins. This concentration dependence may be of physiological significance, as the concentration of both ccd addiction proteins cannot exceed a certain limit because their expression is controlled by autoregulation. The influence of DNA on the thermal stability of the two proteins was probed. It was found that cognate DNA increases the melting temperature of CcdA. In the presence of non-specific DNA the thermal stability was not changed. The melting temperature of CcdB was not influenced by the applied double-stranded oligonucleotides, neither cognate nor unspecific. PMID- 10873462 TI - Protein crystallization by design: chymotrypsinogen without precipitants. AB - Protein crystals are usually obtained by an empirical approach based on extensive screening to identify suitable crystallization conditions. In contrast, we have used a systematic predictive procedure to produce data-quality crystals of bovine chymotrypsinogen A and used them to obtain a refined X-ray structure to 3 A resolution. Measurements of the osmotic second virial coefficient of chymotrypsinogen solutions were used to identify suitable solvent conditions, following which crystals were grown for approximately 30 hours by ultracentrifugal crystallization, without the use of any precipitants. Existing structures of chymotrypsinogen were obtained in solutions including 10-30 % ethanol, whereas simple buffered NaCl solutions were used here. The protein crystallized in the tetragonal space group P4(1)2(1)2, with one molecule per asymmetric unit. The quality of the refined map was very high throughout, with the main-chain atoms of all but four residues clearly defined and with nearly all side-chains also defined. Although only minor differences are seen compared to the structures previously reported, they indicate the possibility of structural changes due to the crystallization conditions used in those studies. Our results show that more systematic crystallization of proteins is possible, and that the procedure can expand the range of conditions under which crystals can be grown successfully and can make new crystal forms available. PMID- 10873461 TI - Use of intrinsic binding energy for catalysis by a cofactor-independent DNA enzyme. AB - The concept of the Circe effect, according to which an enzyme's substrate-binding energy is utilized to destabilize the substrate towards the reaction transition state, has been shown to be a relevant catalytic strategy for naturally occurring protein enzymes and for two ribozymes that use nucleotide-based substrates and metal ion cofactors. We wished to investigate whether such a catalytic strategy extends even to divergent and unevolved catalysts constructed from biopolymers. We examined the properties of a small, in vitro selected, and cofactor independent DNA enzyme, PS5.M, which catalyzes porphyrin metallation. The metallation reaction is unique, in that the energies for binding and for metallation of both the substrate and of a transition-state analogue (TSA) can be measured. We report that PS5.M, originally selected for binding to the TSA, displays the Circe effect in channeling a significant component of entropy-rich "intrinsic" binding energy to distort and to alter the basicity of the bound substrate. The study demonstrates that nucleic acids are, by themselves, capable of creating active sites for the catalysis of chemical reactions involving non nucleotide substrates. Furthermore, the study of the metallation of the TSA provides a quantitative estimate of the effectiveness of such a compound in mimicking the true transition state for porphyrin metallation. PMID- 10873463 TI - Modeling of substrate specificity of the Alzheimer's disease amyloid precursor protein beta-secretase. AB - The enzyme BACE (beta-site APP-cleaving enzyme) has recently been identified as the beta-secretase that cleaves the amyloid precursor protein (APP) to produce the N terminus of the Abeta peptide found in plaques in the brains of Alzheimer's disease patients. BACE is an aspartic protease similar to pepsin and renin. Comparative modeling of the three-dimensional structure of BACE in complex with its substrate shows that several residues confer specificity of the enzyme for APP. In particular, Arg296 forms a salt-bridge with the P1' Asp of the APP substrate, explaining the unusual preference of BACE among aspartic proteases for a P1' residue that is negatively charged. Several hydrophobic residues in the enzyme form a pocket for the P1 hydrophobic residue (Met in wild-type APP and Leu in APP with the "Swedish mutation" associated with early-onset of Alzheimer's disease). Inhibitors that can bind to the BACE active site may prove useful for drugs to treat and prevent Alzheimer's disease. PMID- 10873464 TI - Short TpA-rich segments of the zeta-eta region induce DNA methylation in Neurospora crassa. AB - The mechanisms that establish DNA methylation in eukaryotes are poorly understood. In principle, methylation in a particular chromosomal region may reflect the presence of a "signal" that recruits methylation, the absence of a signal that prevents methylation, or both. Experiments were carried out to address these possibilities for the 1.6 kb zeta-eta (zeta-eta) region, a relict of repeat-induced point mutation (RIP) in the fungus Neurospora crassa. The zeta eta region directs its own de novo methylation at a variety of chromosomal locations. We tested the methylation potential of a nested set of fragments with deletions from one end of the zeta-eta region, various internal fragments of this region, chimeras of eta and the homologous unmutated allele, theta (theta), and various synthetic variants, integrated precisely in single copy at the am locus on linkage group (LG) VR or the his-3 locus on LG IR. We found that: (1) the zeta eta region contains at least two non-overlapping methylation signals; (2) different fragments of the region can induce different levels of methylation; (3) methylation induced by zeta-eta sequences can spread far into flanking sequences; (4) fragments as small as 171 bp can trigger methylation; (5) methylation signals behave similarly, but not identically, at different chromosomal sites; (6) mutation density, per se, does not determine whether sequences become methylated; and (7) neither A:T-richness nor high densities of TpA dinucleotides, typical attributes of methylated sequences in Neurospora, are essential features of methylation signals, but both promote de novo methylation. We conclude that de novo methylation of zeta-eta sequences does not simply reflect the absence of signals that prevent methylation; rather, the region contains multiple, positive signals that trigger methylation. These findings conflict with earlier models for the control of DNA methylation, including the simplest version of the collapsed chromatin model. PMID- 10873465 TI - Stable binding of human XPC complex to irradiated DNA confers strong discrimination for damaged sites. AB - Nucleotide excision repair (NER) of DNA damage requires an efficient means of discrimination between damaged and non-damaged DNA. Cells from humans with xeroderma pigmentosum group C do not perform NER in the bulk of the genome and are corrected by XPC protein, which forms a complex with hHR23B protein. This complex preferentially binds to some types of damaged DNA, but the extent of discrimination in comparison to other NER proteins has not been clear. Recombinant XPC, hHR23B, and XPC-hHR23B complex were purified. In a reconstituted repair system, hHR23B stimulated XPC activity tenfold. Electrophoretic mobility shift competition measurements revealed a 400-fold preference for binding of XPC hHR23B to UV damaged over non-damaged DNA. This damage preference is much greater than displayed by the XPA protein. The discrimination power is similar to that determined here in parallel for the XP-E factor UV-DDB, despite the considerably greater molar affinity of UV-DDB for DNA. Binding of XPC-hHR23B to UV damaged DNA was very fast. Damaged DNA-XPC-hHR23B complexes were stable, with half of the complexes remaining four hours after challenge with excess UV-damaged DNA at 30 degrees C. XPC-hHR23B had a higher level of affinity for (6-4) photoproducts than cyclobutane pyrimidine dimers, and some affinity for DNA treated with cisplatin and alkylating agents. XPC-hHR23B could bind to single-stranded M13 DNA, but only poorly to single-stranded homopolymers. The strong preference of XPC complex for structures in damaged duplex DNA indicates its importance as a primary damage recognition factor in non-transcribed DNA during human NER. PMID- 10873466 TI - Acetyl phosphate-dependent activation of a mutant PhoP response regulator that functions independently of its cognate sensor kinase. AB - The two-component system is a signal communication network generally consisting of a sensor kinase that receives inputs from the environment and modifies the phosphorylated state of a response regulator that executes an adaptive behavior. PhoP is a response regulator that controls virulence gene expression in Salmonella enterica. Transcription of PhoP-regulated genes is modulated by the Mg(2+) levels detected by the sensor PhoQ. Here, we describe a PhoP mutant protein, PhoP*, that functions in the absence of its cognate sensor, thereby allowing transcription of PhoP-activated genes independently of the Mg(2+ )concentration in the environment. The PhoP* protein harbors a S93N substitution in the response regulator receiver domain. PhoP*-mediated transcription is abolished by either mutation of the aspartate residue that is conserved among response regulators as the site of phosphorylation or inactivation of the pta encoded phosphotransacetylase. This enzyme mediates the production of acetyl phosphate, which has been shown to serve as a low molecular mass phosphate donor for certain response regulators. The purified PhoP* protein autophosphorylated from acetyl phosphate more efficiently than the wild-type PhoP protein in vitro. The PhoP* protein retained the capacity to interact with the PhoQ protein, which promoted phosphorylation of the PhoP* protein in vitro and abolished PhoP* mediated transcription under high Mg(2+ )concentrations in vivo. Cumulatively, our results uncover a role of PhoQ in transcriptional repression during growth in millimolar Mg(2+ )and define a mutant response regulator form with an increased capacity to be phosphorylated by acetyl phosphate. PMID- 10873467 TI - Homodimeric peptides displayed by the major coat protein of filamentous phage. AB - Peptide libraries displayed by filamentous bacteriophage have proven a powerful tool for the discovery of novel peptide agonists, antagonists and epitope mimics. Most phage-displayed peptides are fused to the N terminus of either the minor coat protein, pIII, or the major coat protein, pVIII. We report here that peptides containing cysteine residues, displayed as N-terminal fusions to pVIII, can form disulfide-bridged homodimers on the phage coat. Phage clones were randomly selected from libraries containing one or two fixed Cys residues, and surveyed for the presence of peptide-pVIII homodimers by SDS-PAGE analysis that involved pretreatment of the phage with reducing or thiol-modifying agents. For all phage whose recombinant peptide contained a single Cys residue, a significant fraction of the peptide-pVIII molecules were displayed as dimers on the phage coat. The dimeric form was in greater abundance than the monomer in almost all cases in which both forms could be reliably observed. Occasionally, peptides containing two Cys residues also formed dimers. These results indicate that, for a given pVIII-displayed peptide bearing a single Cys residue, a significant fraction of the peptide (>40 %) will dimerize regardless of its sequence; however, sequence constraints probably determine whether all of the peptide will dimerize. Similarly, only occasionally do peptides bearing two Cys residues form intermolecular disulfide bridges instead of intramolecular ones; this indicates that sequence constraints may also determine dimerization versus cyclization. Sucrose-gradient analysis of membranes from cells expressing pVIII fused to a peptide containing a single Cys residue showed that dimeric pVIII is present in the cell prior to its assembly onto phage. A model of the peptide-pVIII homodimer is discussed in light of existing models of the structure and assembly of the phage coat. The unique secondary structures created by the covalent association of peptides on the phage surface suggest a role for homo- and heterodimeric peptide libraries as novel sources of bioactive peptides. PMID- 10873468 TI - A thermodynamic and structural analysis of DNA minor-groove complex formation. AB - As part of an effort to develop a better understanding of the structural and thermodynamic principles of DNA minor groove recognition, we have investigated complexes of three diphenylfuran dications with the d(CGCGAATTCGCG)(2) duplex. The parent compound, furamidine (DB75), has two amidine substituents while DB244 has cyclopentyl amidine substituents and DB226 has 3-pentyl amidines. The structure for the DB244-DNA complex is reported here and is compared to the structure of the DB75 complex. Crystals were not obtained with DB226 but information from the DB75 and DB244 structures as well as previous NMR results on DB226 indicate that all three compounds bind in the minor groove at the AATT site of the duplex. DB244 and DB75 penetrate to the floor of the groove and form hydrogen bonds with T8 on one strand and T20 on the opposite strand while DB226 forms a complex with fewer interactions. Binding studies by surface plasmon resonance (SPR) yield -delta G degrees values in the order DB244>DB75>DB226 that are relatively constant with temperature. The equilibrium binding constants for DB244 are 10-20 times greater than that for DB226. Isothermal titration calorimetric (ITC) experiments indicate that, in contrast to delta G degrees, delta H degrees varies considerably with temperature to yield large negative delta Cp degrees values. The thermodynamic results, analyzed in terms of structures of the DNA complexes, provide an explanation of why DB244 binds more strongly to DNA than DB75, while DB266 binds more weakly. All three compounds have a major contribution to binding from hydrophobic interactions but the hydrophobic term is most favorable for DB244. DB244 also has strong contributions from molecular interactions in its DNA complex and all of these factors combine to give it the largest-delta G degrees for binding. Although the factors that influence the energetics of minor groove interactions are varied and complex, results from the literature coupled with those on the furan derivatives indicate that there are some common characteristics for minor groove recognition by unfused heterocyclic cations that can be used in molecular design. PMID- 10873469 TI - Evaluation of uranyl photocleavage as a probe to monitor ion binding and flexibility in RNAs. AB - In order to evaluate uranyl photocleavage as a tool to identify and characterize structural and dynamic properties in RNA, we compared uranyl cleavage sites in five RNA molecules with known X-ray structures, namely the hammerhead and hepatitis delta virus ribozymes, the P4-P6 domain of the Tetrahymena group I intron, as well as tRNA(Phe) and tRNA(Asp) from yeast. Uranyl photocleavage was observed at specific positions in all molecules investigated. In order to characterize the sites, photocleavage was performed in the absence and in increasing amounts of MgCl(2). Uranyl photocleavage correlates well with sites of low calculated accessibility, suggesting that uranyl ions bind in tight RNA pockets formed by close approach of phosphate groups. RNA foldings require ion binding, usually magnesium ions. Thus, upon the adoption of the native structure, uranyl ions can no longer bind well except in flexible and open to the solvent regions that can undergo induced-fit without disrupting the native fold. Uranyl photocleavage was compared to N-ethyl-N-nitrosourea and lead-induced cleavages in the context of the three-dimensional X-ray structures. Overall, the regions protected from ENU attack are sites of uranyl cleavage, indicating sites of low accessibility which can form ion binding sites. On the contrary, lead cleavages occur at flexible and accessible sites and correlate with the unspecific cleavages prevalent in dynamic and open regions. Applied in a magnesium-dependent manner, and only in combination with other backbone probing agents such as N ethyl-N-nitrosourea, lead and Fenton cleavage, uranyl probing has the potential to reveal high-affinity metal ion environments, as well as regions involved in conformational transitions. PMID- 10873470 TI - A TOPRIM domain in the crystal structure of the catalytic core of Escherichia coli primase confirms a structural link to DNA topoisomerases. AB - Primases synthesize short RNA strands on single-stranded DNA templates, thereby generating the hybrid duplexes required for the initiation of synthesis by DNA polymerases. We present the crystal structure of the catalytic unit of a primase enzyme, that of a approximately 320 residue fragment of Escherichia coli primase, determined at 2.9 A resolution. Central to the catalytic unit is a TOPRIM domain that is strikingly similar in its structure to that of corresponding domains in DNA topoisomerases, but is unrelated to the catalytic centers of other DNA or RNA polymerases. The catalytic domain of primase is crescent-shaped, and the concave face of the crescent is predicted to accommodate about 10 base-pairs of RNA-DNA duplex in a loose interaction, thereby limiting processivity. PMID- 10873471 TI - The crystal structure of tetrameric methionine adenosyltransferase from rat liver reveals the methionine-binding site. AB - Most of the transmethylation reactions use the same methyl donor, S adenosylmethionine (SAM), that is synthesised from methionine and ATP by methionine adenosyltransferase (MAT). In mammals, two MAT enzymes have been detected, one ubiquitous and another liver specific. The liver enzyme exists in two oligomeric forms, a tetramer (MAT I) and a dimer (MAT III), MAT I being the one that shows a higher level of affinity for methionine but a lower SAM synthesis capacity. We have solved the crystal structure of rat liver MAT I at 2.7 A resolution, complexed with a methionine analogue: l-2-amino-4-methoxy-cis but-3-enoic acid (l-cisAMB). The enzyme consists of four identical subunits arranged in two tight dimers that are related by crystallographic 2-fold symmetry. The crystal structure shows the positions of the relevant cysteine residues in the chain, and that Cys35 and Cys61 are perfectly oriented for forming a disulphide link. This result leads us to propose a hypothesis to explain the control of MAT I/III exchange and hence, the effects observed on activity. We have identified the methionine-binding site into the active-site cavity, for the first time. The l-cisAMB inhibitor is stacked against Phe251 aromatic ring in a rather planar conformation, and its carboxylate group coordinates a Mg(2+), which, in turn, is linked to Asp180. The essential role of the involved residues in MAT activity has been confirmed by site-directed mutagenesis. Phe251 is exposed to solvent and is located in the beginning of the flexible loop Phe251-Ala260 that is connecting the N-terminal domain to the central domain. We postulate that a conformational change may take place during the enzymatic reaction and this is possibly the reason of the unusual two-step mechanism involving tripolyphosphate hydrolysis. Other important mechanistic implications are discussed on the light of the results. Moreover, the critical role that certain residues identified in this study may have in methionine recognition opens further possibilities for rational drug design. PMID- 10873472 TI - Effects of side-chain characteristics on stability and oligomerization state of a de novo-designed model coiled-coil: 20 amino acid substitutions in position "d". AB - We describe the de novo design and biophysical characterization of a model coiled coil protein in which we have systematically substituted 20 different amino acid residues in the central "d" position. The model protein consists of two identical 38 residue polypeptide chains covalently linked at their N termini via a disulfide bridge. The hydrophobic core contained Val and Ile residues at positions "a" and Leu residues at positions "d". This core allowed for the formation of both two-stranded and three-stranded coiled-coils in benign buffer, depending on the substitution at position "d". The structure of each analog was analyzed by CD spectroscopy and their relative stability determined by chemical denaturation using GdnHCI (all analogs denatured from the two-stranded state). The oligomeric state(s) was determined by high-performance size-exclusion chromatography and sedimentation equilibrium analysis in benign medium. Our results showed a thermodynamic stability order (in order of decreasing stability) of: Leu, Met, Ile, Tyr, Phe, Val, Gln, Ala, Trp, Asn, His, Thr, Lys, Ser, Asp, Glu, Arg, Orn, and Gly. The Pro analog prevented coiled-coil formation. The overall stability range was 7.4 kcal/mol from the lowest to the highest analog, indicating the importance of the hydrophobic core and the dramatic effect a single substitution in the core can have upon the stability of the protein fold. In general, the side-chain contribution to the level of stability correlated with side-chain hydrophobicity. Molecular modelling studies, however, showed that packing effects could explain deviations from a direct correlation. In regards to oligomerization state, eight analogs demonstrated the ability to populate exclusively one oligomerization state in benign buffer (0.1 M KCl, 0.05 M K(2)PO(4)(pH 7)). Ile and Val (the beta-branched residues) induced the three stranded oligomerization state, whereas Tyr, Lys, Arg, Orn, Glu and Asp induced the two-stranded state. Asn, Gln, Ser, Ala, Gly, Phe, Leu, Met and Trp analogs were indiscriminate and populated two-stranded and three-stranded states. Comparison of these results with similar substitutions in position "a" highlights the positional effects of individual residues in defining the stability and numbers of polypeptide chains occurring in a coiled-coil structure. Overall, these results in conjunction with other work now generate a relative thermodynamic stability scale for 19 naturally occurring amino acid residues in either an "a" or "d" position of a two-stranded coiled-coil. Thus, these results will aid in the de novo design of new coiled-coil structures, a better understanding of their structure/function relationships and the design of algorithms to predict the presence of coiled-coils within native protein sequences. PMID- 10873474 TI - Introduction to flow cytometry. PMID- 10873473 TI - The role of steric hindrance in 3TC resistance of human immunodeficiency virus type-1 reverse transcriptase. AB - Treating HIV infections with drugs that block viral replication selects for drug resistant strains of the virus. Particular inhibitors select characteristic resistance mutations. In the case of the nucleoside analogs 3TC and FTC, resistant viruses are selected with mutations at amino acid residue 184 of reverse transcriptase (RT). The initial change is usually to M184I; this virus is rapidly replaced by a variant carrying the mutation M184V. 3TC and FTC are taken up by cells and converted into 3TCTP and FTCTP. The triphosphate forms of these nucleoside analogs are incorporated into DNA by HIV-1 RT and act as chain terminators. Both of the mutations, M184I and M184V, provide very high levels of resistance in vivo; purified HIV-1 RT carrying M184V and M184I also shows resistance to 3TCTP and FTCTP in in vitro polymerase assays. Amino acid M184 is part of the dNTP binding site of HIV-1 RT. Structural studies suggest that the mechanism of resistance of HIV-1 RTs carrying the M184V or M184I mutation involves steric hindrance, which could either completely block the binding of 3TCTP and FTCTP or allow binding of these nucleoside triphosphate molecules but only in a configuration that would prevent incorporation. The available kinetic data are ambiguous: one group has reported that the primary effect of the mutations is at the level of 3TCTP binding; another, at the level of incorporation. We have approached this problem using assays that monitor the ability of HIV-1 RT to undergo a conformational change upon binding a dNTP. These studies show that both wild-type RT and the drug-resistant variants can bind 3TCTP at the polymerase active site; however, the binding to M184V and M184I is somewhat weaker and is sensitive to salt. We propose that the drug-resistant variants bind 3TCTP in a strained configuration that is salt-sensitive and is not catalytically competent. PMID- 10873475 TI - Concurrent measurement of antigen- and antibody-dependent oxidative burst and phagocytosis in monocytes and neutrophils. AB - The current study aims to review flow cytometric (FCM) parameters for the quantification of phagocytosis. A limitation of existing methods is their difficulty with accurate quantification of the phagocytic index, i.e., number of beads per phagocyte, in individual cell lines in mixed cell suspensions. We have quantified phagocytosis and the oxidative burst simultaneously using fluorescent beads coated with meningococcal outer membrane vesicles (OMV beads) by the conversion of dihydrorhodamine 123 (DHR-123) to rhodamine 123 (R-123). Both these processes depend on specific serum opsonins. After the incubation, staining with a fluorescent anti-CD14 monoclonal antibody succeeded in discriminating phagocytosing monocytes from neutrophils. The spectral overlaps between OMV beads, R-123, and anti-CD14 could be completely compensated. Percentage of phagocytosis and the phagocytic index were similar in monocytes and neutrophils, but the oxidative burst behaved differently. Two monocyte subpopulations were observed. Both subpopulations spontaneously converted some DHR-123 into R-123, whereas the reaction was triggered by phagocytosis in neutrophils. The total oxidative response increased with increasing phagocytic index in both cell types, but the oxidative burst in monocytes was about twice that of neutrophils. The oxidative ratio (mean R-123 fluorescence value divided by the phagocytic index) declined with time in monocytes, but increased in neutrophils. Our results demonstrate the need for careful attention to technical details. This single laser, three-color FCM method facilitates the comparative research of phagocytosis and the oxidative burst in monocytes and neutrophils and provides a basis for a number of applications in hematology, infectious medicine, and immunology. PMID- 10873476 TI - Analysis of free intracellular calcium by flow cytometry: multiparameter and pharmacologic applications. AB - Flow cytometry offers numerous advantages over traditional techniques for measuring intracellular Ca(2+) in lymphoid and nonlymphoid cells. In particular, the heterogeneity of cell responses can be defined by flow cytometry, and multiparameter analyses permit the determination of intracellular Ca(2+) in surface-marker-defined target cells as well as correlation of changes in Ca(2+) with other biochemical markers, including ligand binding. This article presents several established methods for measuring intracellular Ca(2+) by flow cytometry in lymphoid and nonlymphoid cells. Examples are provided for determination of Ca(2+) in human peripheral blood leukocytes and two human epithelial cell lines grown in monolayer. In addition, applications are reviewed or presented for correlating changes in intracellular Ca(2+) with other cell parameters, including cell cycle analysis, changes in cell membrane integrity, and the induction of apoptosis markers. Finally, a number of novel sample handling capabilities useful for performing kinetic analyses of Ca(2+) changes by flow cytometry are now available and one application is presented which is finding utility in pharmacologic studies. PMID- 10873477 TI - Flow cytometric analysis of microorganisms. AB - The application of flow cytometry to microorganisms is as old as the technique itself, but it has historically been underexploited for microbial applications. This is now being reversed and microbiologists are ideally placed to benefit from recent technological advances. While earlier papers demonstrated the use of flow cytometry for studies of viability and taxonomy, recent developments in bioinformatics and reporter gene technologies are leading to novel applications in microbiology. Variants of green fluorescent protein have been used for the study of conditional microbial gene regulation in medically important host pathogen interactions and fluorescence-activated cell sorting is being applied to the isolation of novel mutants in directed evolution studies. This paper reviews the reasons for the delay in the application of flow cytometry to microbial problems, the range of applications, and their limitations and considers the progress made in developing new strategies for use in microbiological investigations. PMID- 10873478 TI - Alveolar macrophage-environmental particle interaction: analysis by flow cytometry. AB - Inhaled particulates such as pollutant particles, allergens, and microorganisms are rapidly cleared by alveolar macrophages (AMs). Methods for analysis of AM particle interaction have been hindered by the lack of a convenient assay. Flow cytometry offers rapid, sensitive, and reproducible measurements of single cells in suspension. Multiple parameters can be measured in real time. Here we will review the application of flow cytometry to the study and characterization of AM receptors for unopsonized environmental particles. We will discuss the role of this technique in identifying a key AM receptor system involved in lung defense. Multiparametric flow cytometry to analyze intracellular functional parameters, though a powerful and unique tool, needs to be interpreted with caution. We will also discuss the advantages and limitations of flow cytometry in analysis of AM particle interaction. PMID- 10873479 TI - Analysis of virus-infected cells by flow cytometry. AB - Flow cytometry has been used to study virus-cell interactions for many years. This article critically reviews a number of reports on the use of flow cytometry for the detection of virus-infected cells directly in clinical samples and in virus-infected cultured cells. Examples are presented of the use of flow cytometry to screen antiviral drugs against human immunodeficiency virus (HIV), human cytomegalovirus, and herpes simplex viruses (HSV) and to perform drug susceptibility testing for these viruses. The use of reporter genes such as green fluorescent protein incorporated into HIV or HSV or into cells for the detection of the presence of virus, for drug susceptibility assay, and for viral pathogenesis is also covered. Finally, studies on the use of flow cytometry for studying the effect of virus infection on apoptosis and the cell cycle are summarized. It is hoped that this article will give the reader some understanding of the great potential of this technology for studying virus cell interactions. PMID- 10873480 TI - Evaluation of platelet function by flow cytometry. AB - Platelet function in whole blood can be comprehensively evaluated by flow cytometry. Flow cytometry can be used to measure platelet reactivity, circulating activated platelets, platelet-platelet aggregates, leukocyte-platelet aggregates, procoagulant platelet-derived microparticles, and calcium flux. Clinical applications of whole blood flow cytometric assays of platelet function in disease states (e.g., acute coronary syndromes, angioplasty, and stroke) may include identification of patients who would benefit from additional antiplatelet therapy and prediction of ischemic events. Circulating monocyte-platelet aggregates appear to be a more sensitive marker of in vivo platelet activation than circulating P-selectin-positive platelets. Flow cytometry can also be used in the following clinical settings: monitoring of GPIIb-IIIa antagonist therapy, diagnosis of inherited deficiencies of platelet surface glycoproteins, diagnosis of storage pool disease, diagnosis of heparin-induced thrombocytopenia, and measurement of the rate of thrombopoiesis. PMID- 10873482 TI - Flow cytometry assay for counting micronucleated erythrocytes: development process. AB - Development of any new assay proceeds in several phases. When an assay is intended for regular use to support regulatory decision-making, there are significant additional stages in the development process beyond the initial description of the method. In this paper we discuss some of the studies related to the development of a flow cytometric method for counting micronuclei in rodent erythrocytes. Studies related to fixation methods and conditions, standardization of DNA staining, and antibody staining are discussed. These studies, while not part of the formal description of the method, are needed as part of the preparation for the formal validation of the method. In addition, the lessons learned in transferring the method to other laboratories are briefly discussed in relation to defining the final protocol. PMID- 10873481 TI - Membrane potential estimation by flow cytometry. AB - Membrane potential (delta psi) is generated and maintained by concentration gradients of ions such as sodium, potassium, chloride, and hydrogen. Changes in cytoplasmic delta psi in the course of surface-receptor-mediated processes related to the development, function, and pathology of many cell types often play a role in transmembrane signaling. Cytoplasmic delta psi is also reduced to zero when the membrane is ruptured by chemical or physical agents. Mitochondrial delta psi is reduced when energy metabolism is disrupted, notably in apoptosis. In bacteria, which lack mitochondria, delta psi reflects both the state of energy metabolism and the physical integrity of the cytoplasmic membrane. Flow cytometry can be used to estimate membrane potential in eukaryotic cells, mitochondria in situ, isolated mitochondria, and bacteria. Older methods, using lipophilic cationic dyes such as the cyanines and rhodamine 123 or lipophilic anionic dyes such as the oxonols can detect relatively large changes in delta psi and identify heterogeneity of response in subpopulations comprising substantial fractions of a cell population. Newer ratiometric techniques allow precise measurement of delta psi to within 10 mV or less. Among other factors, action of efflux pumps, changes in membrane structure, and changes in protein or lipid concentration in the medium in which cells are suspended can produce changes in cellular fluorescence which may be misinterpreted as changes in delta psi. Techniques for estimation and measurement of Delta Psi therefore typically require careful control of cell and reagent concentrations and incubation times and selection of appropriate controls if they are to provide accurate information. PMID- 10873483 TI - Fluorescence intensity calibration for immunophenotyping by flow cytometry. AB - Fluorescence intensity (FI) is the basis for classifying phenotypes by fluorescence-label flow cytometry. FI is customarily recorded as an arbitrary relative value, but with proper calibration it can be expressed in stoichiometric units called molecules of equivalent soluble fluorochrome (MESF) that reflect the concentrations of the fluorescent conjugates and the receptors they stain. Forthcoming availability of authoritative standards and consensus methods will alleviate many of the difficulties encountered in making valid MESF measurements. FI calibration establishes the true values for the critical parameters of the fluorescence measurement, a useful feature for quality control. It further allows the establishment of a comparable window of analysis across different times and laboratories, and it permits numeric assessment of antibody-binding capacity (ABC) values in selected cell populations. The relation between ABC values and receptor expression is complicated by several factors, but careful assessment of the binding chemistry can establish the actual number of receptors on cells stained by fluorescent conjugates. PMID- 10873484 TI - Evaluation of adenoviral vectors by flow cytometry. AB - Biological assays for adenoviral gene therapy vectors have included conventional procedures initially developed to detect wild-type adenoviruses. Standard virological assays to quantitate adenoviruses rely on the virus to infect and replicate in the host cell until a cytopathic effect is observed. The appearance of plaques, colonies of rounded, enlarged cells containing infectious virions, usually takes 2 to 3 weeks to reach an endpoint. We describe a flow cytometric bioassay for adenovirus which shortens the time from when the infection takes place to the time that biological titer is determined. A fluorescent focus forming assay was one of the first rapid adenoviral bioassays developed. Virus titer was determined using fluorescence immunocytochemistry to detect adenovirus proteins and microscopy to count fluorescent foci in cultures of adenovirus infected cells. In this study, we describe a flow cytometric assay performed on cells stained for adenovirus hexon capsid protein, where virus titer is determined based on the dose-dependent appearance of hexon-positive cells. Adenovirus hexon detection in infected cells can provide data to determine virus titer, inducible promoter function in vector-complementing cells, and vector replication in complementation-deficient cells. PMID- 10873485 TI - Identification of a CHO cell-elongating factor produced by Vibrio cholerae O1. AB - Vibrio cholerae strains with all known toxin genes deleted or inactivated still cause diarrhoea in some volunteers, suggesting the presence of an unknown virulence factor or factors. Lysozyme-EDTA treated cells of JBK70, a genetically manipulated cholera toxin negative strain of Vibrio cholerae O1, biotype El Tor, release a factor that causes elongation of Chinese hamster ovary (CHO) cells. CHO cell-elongating toxin (Cef) was purified by FPLC chromatography (anion exchange; Q Sepharose High Performance) followed by 2D electrophoresis (isoelectric focusing gel, IEF; pH 3-9 and SDS-PAGE, 8-25% gradient gel). Partly purified toxin (anion exchange or IEF-eluted concentrate) caused fluid accumulation in sealed infant mice suggesting that Cef shows some properties of an enterotoxin. On SDS-PAGE (8-25%) and IEF (pH 2.5-5.0) gels, CHO cell activity was associated with a single band at 85 kDa and a pI of 3.8, respectively. A unique amino terminal sequence, XGDETNSSGASTEVVYESYIQQ, was determined by automated Edman degradation of gel-purified protein. The unique molecular mass, N-terminal sequence and activity on CHO cells indicate that this factor is not zonula occludens toxin (Zot) or accessory cholera enterotoxin (Ace) or the Hly A haemolysin. Partly purified Cef did not increase cyclic AMP or prostaglandin E(2)levels in CHO cells which suggests that its mechanism of action differs from that of cholera toxin. PMID- 10873486 TI - Exposure of human peripheral blood mononuclear cells to total lipids and serovar specific glycopeptidolipids from Mycobacterium avium serovars 4 and 8 results in inhibition of TH1-type responses. AB - Previous studies have suggested that large quantities of bacterial lipids may accumulate and persist within host cells during chronic stages of Mycobacterium avium infections. This study intended to assess the ability of purified M. avium lipids to affect TH-1-type responses in human peripheral blood mononuclear cells (PBMC) from healthy donors. PBMC were exposed to total lipids and serovar specific glycopeptidolipids (GPL) extracted from M. avium serovars 4 and 8, which have been reported to predominate as opportunistic infection among AIDS patients. After 24 h exposure to lipids followed by PHA/PMA treatment, IL-2 and IFN-gamma were assayed in the supernatants. Reverse transcriptase polymerase chain reaction (RT-PCR) was used for a semiquantitative estimation of mRNA for IL-2 and IFN gamma in cell pellets at various time points. Exposure of PBMC to M. avium total lipids significantly suppressed PHA/PMA-induced secretion of IL-2 and IFN-gamma as determined by ELISA. The GPL antigens from serovar 4 were more efficient at inhibiting TH-1 responses than GPL from serovar 8. CD4(+)T-lymphocyte enrichment of PBMC demonstrated that suppression by M. avium lipids was intact without the presence of other cell populations such as monocytes and B-cells. Preliminary RT PCR experiments showed that the secretion of TH-1 cytokines was partially affected at the transcriptional level. The results obtained showed that M. avium lipids are indeed able to modify the induction of TH-1-type cytokines by human PBMC, and suggest that accumulation of M. avium lipids in the chronic stages of infection may play an important role in the pathogenesis of HIV infection. PMID- 10873487 TI - Chlamydia pneumoniae present in the human synovium are viable and metabolically active. AB - We demonstrated that chromosomal DNA from Chlamydia pneumoniae is present in synovial tissue in at least some patients with reactive arthritis/Reiter's syndrome and other arthritides. Here, we provide initial molecular evidence that the bacterium is viable and metabolically active when present in the synovium. We used reverse transcription-polymerase chain reaction (RT-PCR) assays targeting primary transcripts from the chlamydial rRNA operons, and mRNA from several C. pneumoniae genes (hsp60, ompA, KDO transferase, Mr=76000 protein), to analyse RNA preparations from synovial tissue of 10 patients with various forms of arthritis; each patient was known to be PCR-positive for C. pneumoniae DNA in synovium prior to RT-PCR assays. Two PCR-negative patients served as controls for RT-PCR assays. In the 10 patients PCR-positive for C. pneumoniae DNA, RT-PCR assays targeting primary transcripts from the rRNA operons of the organism showed that these molecules were present in each sample, as were transcripts from the bacterial hsp60 gene. Assays targeting mRNAs from the Mr=76000 protein and the KDO transferase genes of C. pneumoniae gave positive results for 6/10 preparations. We were unable to identify mRNA from the chlamydial major outer membrane protein gene (ompA) in any preparation. RNA preparations from the two control patients were negative in all RT-PCR assays targeting C. pneumoniae transcripts. These results indicate that in patients infected with the organism, synovial C. pneumoniae are viable and metabolically active, as are C. trachomatis cells in the same context. Such viability is consistent with a role in long-term contribution to pathogenesis in joint disease. PMID- 10873488 TI - Identification of Pasteurella multocida virulence genes in a septicemic mouse model using signature-tagged mutagenesis. AB - P. multocida is the causative agent of several economically significant veterinary diseases occurring in numerous species worldwide. Signature-tagged mutagenesis (STM) is a powerful genetic technique used to simultaneously screen multiple transposon mutants of a pathogen for their inability to survive in vivo. We have designed an STM system based on a mini-Tn10 transposon, chemiluminescent detection and semi-quantitative analysis and have identified transposon insertions into genes of Pasteurella multocida that attenuate virulence in a septicemic mouse model. A bank of 96 transposons containing strongly-hybridizing tags was used to create 19 pools of P. multocida transposon mutants containing approximately 70-90 mutants/pool. A total of 62 mutants were attenuated when checked individually, and 25 unique single transposon insertion mutations were identified from this group. The sequence of the disrupted ORF for each attenuated mutant was determined by either cloning or PCR-amplifying and sequencing the flanking regions. The attenuated mutants contained transposon insertions in genes encoding biosynthetic enzymes, virulence factors, regulatory components and unknown functions. This study should contribute to an understanding of the pathogenic mechanisms by which P. multocida and other pathogens in the Pasteurellaceae family cause disease and identify novel live vaccine candidates and new potential antibiotic targets. PMID- 10873489 TI - Identification of Actinobacillus pleuropneumoniae virulence genes using signature tagged mutagenesis in a swine infection model. AB - Actinobacillus pleuropneumoniae is a significant respiratory pathogen of swine causing a severe and often fatal fibrinous hemorrhagic bronchopneumonia with significant economic losses resulting from chronic as well as acute infections. This study describes the application of a signature-tagged mutagenesis (STM) system to identify in vivo critical genes of A. pleuropneumoniae. Twenty pools representing over 800 A. pleuropneumoniae mutants were screened in a natural-host porcine infection model and presumptive attenuated mutants were selected. The identity of the disrupted gene in each mutant was determined using an inverse PCR approach to amplify DNA sequences adjacent to the transposon insertion, followed by sequencing of the PCR product and comparison to bacterial databases. In vitro and in vivo competitive indices were determined for each unique mutant, and a total of 20 unique, attenuating gene disruptions were identified including insertions into homologues of genes involved in biosynthesis, virulence determinants, regulation, translation and unknown functions. Three of the genes required for virulence of A. pleuropneumoniae in this study were also identified in a previous STM study of Pasteurella multocida. Seven of the STM-derived mutants were also evaluated for their potential as live vaccine strains and provided good protection against homologous challenge. PMID- 10873490 TI - Use of confocal microscopy to detect Salmonella typhimurium within host cells associated with Spv-mediated intracellular proliferation. AB - The major limitation in histological examination of orally inoculated mice of Salmonella typhimurium has been the difficulty of attaining high enough levels for immunochemical detection. This problem has been solved by the use of confocal laser scanning microscopy (CLSM) analysis, which allows detection of bacteria in the immunostained sections of mouse spleens at a minimum rate of approximately 1000 colony-forming units (cfu)/spleen. Here, we demonstrate that over 80% of salmonellae of the wild type of S. typhimurium were detected intracellularly within Mac-1 positive cells by the CLSM analysis of immunostained sections from spleens in orally or subcutaneously inoculated mice. Only 40% of salmonellae of the spv -deleted strain were detected inside Mac-1 positive cells. These data suggest that the spv genes play a key role in intracellular proliferation within phagocytes in the mouse spleen. PMID- 10873491 TI - Tunneling spectroscopy from magnetization evolution in a tilted rotating frame of nuclear spins. AB - The time evolution of the proton Zeeman magnetization in the rotating frame at the magic angle theta(M) = cos(-1)(1/3) is calculated for an isolated tunneling methyl group and its Fourier transform is given. The calculation compares well with the experimental spectra of CH(3)CD(2)I and methylmalonic acid. It is shown that Fourier transform spectroscopy of the magnetization evolution in a tilted RF frame represents an excellent alternative to the analogous experiment performed at exact resonance, resulting in improved resolution and a much better signal-to noise ratio. PMID- 10873492 TI - Cross correlations in the longitudinal relaxation of strongly coupled spins AB - A generalized set of magnetization modes for quantifying cross-correlation contributions to longitudinal relaxation in strongly coupled spin systems is described in this paper. Such a set of modes (called longitudinal multiple quantum modes) is used to unravel cross-correlation information in strongly coupled systems, where the strength of the J coupling tends to obscure such effects. The applicability of such methods is demonstrated for a small molecule which exhibits some strong coupling effects even at high magnetic field strengths. The contribution of "remote" cross correlations to the longitudinal relaxation of strongly coupled spins is detailed. Copyright 2000 Academic Press. PMID- 10873493 TI - NMR spin echoes for molecular rotators with C(3) symmetry AB - NMR spin echoes are calculated for a rotator consisting of three spin-(1/2) nuclei whose three-particle wave function obeys C(3) symmetry. On decomposing the nuclear dipole-dipole interactions in terms of irreducible operators of the representations of the group C(3) it is found that for spins belonging to the A representation, the echo amplitude is maximized for a RF pulse sequence. The dependences of the echo formation on the orientation of the rotor and on the strengths of the dipolar interactions and the magnetic field inhomogeneities are discussed. Copyright 2000 Academic Press. PMID- 10873494 TI - Quantitative interpretation of magnetization transfer in spoiled gradient echo MRI sequences. AB - A method for analyzing general pulsed magnetization transfer (MT) experiments in which off-resonance saturation pulses are interleaved with on-resonance excitation pulses is presented. We apply this method to develop a steady-state signal equation for MT-weighted spoiled gradient echo sequences and consider approximations that facilitate its rapid computation. Using this equation, we assess various experimental designs for quantitatively imaging the fractional size of the restricted pool, cross-relaxation rate, and T(1) and T(2) relaxation times of the two pools in a binary spin bath system. From experiments on agar gel, this method is shown to reliably and accurately estimate the exchange and relaxation properties of a material in an imaging context, suggesting the feasibility of using this technique in vivo. PMID- 10873495 TI - Nonlinear identification of NMR spin systems by adaptive filtering. AB - In this paper, we present two new methods for identifying NMR spin systems. These methods are based on nonlinear adaptive filtering. The spin system is assumed to be time-invariant with memory. The first method uses a truncated discrete Volterra series to describe the nonlinear relationship between excitation (input) and system response (output). First-, second-, and third-order kernels of this series are estimated employing the least mean square (LMS) algorithm. Three parallel filters can then model the NMR spin system so that its output is no more than simple sum of three convolution products between combinations of the input signal and filters coefficients. It is also shown that the contribution of the Volterra second-order term to the total system response is neglected compared with the contributions of the first- and the third-order terms. In the second identification method, the output signal is related to the input signal through a recursive nonlinear difference equation with constant coefficients. The LMS algorithm is used again to estimate the equation coefficients. The two methods are validated with a simulated NMR system model based on Bloch equations. The results and the performances of these methods are analyzed and compared. It is shown that our methods permit a simple identification of NMR spin systems. The field of applications of this study is promising in the optimization of NMR signal detection, especially in the cases of low signal-to-noise ratios where optimum signal filtering and analysis must be performed. PMID- 10873496 TI - Efficient deuterium-carbon REDOR NMR spectroscopy. AB - Phase modulated pulses for deuterium recoupling in (2)H-(13)C REDOR NMR spectroscopy have been introduced to improve dephasing of the detected (13)C nuclei. The deuterium inversion properties of phase modulated recoupling pulses have been studied experimentally on l-alanine-2-d(1) and theoretically using average Hamiltonian theory and exact simulations of the equation of motion of the density matrix. The best (13)C dephasing was observed when XYXYX (PM5) deuterium recoupling pulses were applied. A comparison to the 90 degrees -180 degrees -90 degrees (CPL) composite pulse scheme revealed an improvement of recoupling on the order of 2.5. Simple CW recoupling pulses of the same length of PM5 and CPL pulses showed the weakest (13)C dephasing. Simulations have shown that the (2)H recoupling efficiency of PM5 REDOR experiments approach the very efficient REAPDOR results. However, in our case a REAPDOR study of l-alanine-2-d(1) resulted in a significant decrease of the (13)C signal intensity due to pulse imperfections of (13)C pi-pulses. The new PM5-REDOR technique has been employed to study the torsion angle between C1/2 and C5 in ethylmalonic acid-4-d(2). PMID- 10873497 TI - Thermally excited multiplet states in macerals separated from bituminous coal AB - Electron paramagnetic resonance searches of thermally excited multiplet states in macerals, exinite, vitrinite, and inertinite of Polish medium-rank coal (85.6 wt% C), were performed. Numerical analysis of lineshape indicates a multicomponent structure of the EPR spectra of macerals heated at 300 degrees and 650 degrees C. EPR spectra of exinite and vitrinite are a superposition of broad Gauss, broad Lorentz (Lorentz 1), and narrow Lorentz (Lorentz 3) lines. Two narrow Lorentz (Lorentz 2 and Lorentz 3) lines were observed in the resonance absorption curves of inertinite. The influence of the measuring temperature (100-300 K) on the EPR lines of the macerals was also studied. The experimentally obtained temperature dependence of the EPR line intensities were fitted by the theoretical functions characteristic for paramagnetic centers with ground doublet state (S = 12) and paramagnetic centers with thermally excited triplet (S = 1) and quadruplet (S = 32) states. Thermally excited multiplet states were found in exinite and vitrinite. Both paramagnetic centers with doublet ground state (S = 12) and paramagnetic centers with thermally excited states, probably quadruplet states (S = 32), exist in the group of paramagnetic centers of exinite and vitrinite with the broad Lorentz 1 lines. Intensities (I) of the broad Gauss and the narrow Lorentz 3 lines of exinite and vitrinite changes with temperature according to the Curie law (I = C/T). The existence of thermally excited multiplet states was not stated for inertinite. The two groups of paramagnetic centers of inertinite with Lorentz 2 and Lorentz 3 lines obey the Curie law. Copyright 2000 Academic Press. PMID- 10873498 TI - Compound radiofrequency-driven recoupling pulse sequences for efficient magnetization transfer by homonuclear dipolar interaction under magic-angle spinning conditions AB - The maximum of the transferred magnetization in rotating powdered solids under the radiofrequency-driven recoupling (RFDR) pulse sequence is enhanced by reducing the orientation dependence of the effective recoupled homonuclear dipolar interaction. The compound RFDR (CRFDR) pulse sequence for this enhancement consists of RFDR pulse units (tau(i)-pi-tau(R)-pi-1171;tau(i)) with different tau(i), where tau(R) is the sample rotation period, tau(i) and 1171;tau(i) (=tau(R) - tau(i)) are delays, and pi is a 180 degrees pulse. The delay tau(i) modifies the zero-quantum spin operators and the sample rotation angle dependence of the recoupled dipolar Hamiltonian. The CRFDR pulse sequences were optimized for mixing by varying tau(i). Numerical simulation for the two spin system only with a dipolar interaction and isotropic chemical shifts indicates that the transfer efficiency of CRFDR averaged over the powder is about 70%, which is 30% higher than the efficiency of the RFDR pulse over a broad range of about 1/tau(R) in resonance frequency difference. The CRFDR sequences need about 60% longer mixing times to maximize the transferred magnetizaion in comparison with the original RFDR sequence. Chemical shift anisotropy, the other dipolar interactions, and relaxation generally reduce the enhancement by CRFDR. Experiments for fully (13)C-labeled alanine, however, show that the maximum of the magnetization transferred with CRFDR from the carboxyl to alpha carbon is about 15% greater than that with RFDR. Copyright 2000 Academic Press. PMID- 10873499 TI - Spin relaxation measurements using first-harmonic out-of-phase absorption EPR signals: rotational motion effects. AB - A recent survey of nonlinear continuous-wave (CW) EPR methods revealed that the first-harmonic absorption EPR signal, detected 90 degrees out of phase with respect to the Zeeman modulation (V(1)(')-EPR), is the most appropriate for determining spin-lattice relaxation enhancements of spin labels (V. A. Livshits, T. Pali, and D. Marsh, 1998, J. Magn. Reson. 134, 113-123). The sensitivity of such V(1)(')-EPR spectra to molecular rotational motion is investigated here by spectral simulations for nitroxyl spin labels, over the entire range of rotational correlation times. Determination of the effective spin-lattice relaxation times is less dependent on rotational mobility than for other nonlinear CW EPR methods, especially at a Zeeman modulation frequency of 25 kHz which is particularly appropriate for spin labels. This relative insensitivity to molecular motion further enhances the usefulness of the V(1)(')-EPR method. Calibrations of the out-of-phase to in-phase spectral intensity (and amplitude) ratios are given as a function of spin-lattice relaxation time, for the full range of spin-label rotational correlation times. Experimental measurements on spin labels in the slow, intermediate, and fast motional regimes of molecular rotation are used to test and validate the method. PMID- 10873500 TI - Efficient double-quantum excitation in rotational resonance NMR. AB - We present a new technique for double-quantum excitation in magic-angle-spinning solid-state NMR. The method involves (i) preparation of nonequilibrium longitudinal magnetization; (ii) mechanical excitation of zero-quantum coherence by spinning the sample at rotational resonance, and (iii) phase-coherent conversion of the zero-quantum coherence into double-quantum coherence by frequency-selective spin inversion. The double-quantum coherence is converted into observable magnetization by reversing the excitation process, followed by a pi/2 pulse. The method is technically simple, does not require strong RF fields, and is feasible at high spinning frequencies. In [(13)C(2),(15)N]-glycine, with an internuclear (13)C-(13)C distance of 0.153 nm, we achieve a double-quantum filtering efficiency of approximately 56%. In [11, 20-(13)C(2)]-all-E-retinal, with an internuclear (13)C-(13)C distance of 0.296 nm, we obtain approximately 45% double-quantum filtering efficiency. PMID- 10873501 TI - Exact ML estimation of spectroscopic parameters AB - In a paper on spectroscopic imaging published in this journal Spielman et al. (J. Magn. Reson. 79, 66-77 (1988)) made the important point that a priori information about the compounds present can and should be incorporated into the estimation of spectroscopic signal parameters. They proposed using the maximum likelihood (ML) approach for parameter estimation, but failed to incorporate properly the full a priori information that was assumed to be available. Consequently they ended up with a spectroscopic imaging method that is only a suboptimal approximation of the ML method. In this paper we derive the exact ML method, present a computationally efficient implementation of it (which is much faster than the direct implementation suggested by Spielman et al. for their suboptimal method), and illustrate numerically the performance gain that can be achieved over the method of Spielman et al. Copyright 2000 Academic Press. PMID- 10873502 TI - Two-dimensional ENDOR-ESEEM correlation spectroscopy AB - A two-dimensional (2D) experiment that correlates electron-nuclear double resonance (ENDOR) and electron spin-echo envelope modulation (ESEEM) frequencies, useful for unraveling and assigning ENDOR and ESEEM spectra from different paramagnetic centers with overlapping EPR spectra, is presented. The pulse sequence employed is similar to the Davies ENDOR experiment with the exception that the two-pulse echo detection is replaced by a stimulated echo detection in order to enhance the resolution in the ESEEM dimension. The two-dimensional data set is acquired by measuring the ENDOR spectrum as a function of the time interval T between the last two microwave pulses of the stimulated echo detection scheme. This produces a series of ENDOR spectra with amplitudes that are modulated with T. Fourier transformation (FT) with respect to T then generates a 2D spectrum with cross peaks connecting spectral lines of the ESEEM and ENDOR spectra that belong to the same paramagnetic center. Projections along the vertical and horizontal axes give the three-pulse FT-ESEEM and ENDOR spectra, respectively. The feasibility of the experiment was tested by simulating 2D ENDOR ESEEM correlation spectra of a system consisting of an electron spin (S = (1/2)) coupled to two nuclei (I(1) = I(2) = (1/2)), taking into account experimental conditions such as pulse durations and off-resonance irradiation frequencies. The experiment is demonstrated on a single crystal of Cu(2+) doped l-histidine (Cu His), containing two symmetrically related Cu(2+) sites that at an arbitrary orientation exhibit overlapping ESEEM and ENDOR spectra. While the ESEEM spectrum is relatively simple and arises primarily from one weakly coupled (14)N, the ENDOR spectrum is very crowded due to contributions from two nonequivalent nitrogens, two chlorides, and a relatively large number of protons. The simple ESEEM projection of the 2D ENDOR-ESEEM correlation spectrum is then used to disentangle the ENDOR spectrum and resolve two sets of lines corresponding to the different sites. Copyright 2000 Academic Press. PMID- 10873503 TI - Thermal convection currents in NMR: flow profiles and implications for coherence pathway selection AB - Rayleigh-Benard convection currents are visualized in a vertical cylindrical tube by means of magnetic resonance imaging. Axially antisymmetric flow, multiple vertical rolls, and twisted node planes are observed. The flow can also be induced by strong RF irradiation. Its effects on the coherence pathways in NMR experiments employing field gradients are discussed. Copyright 2000 Academic Press. PMID- 10873504 TI - NH-NH vector correlation in peptides by solid-state NMR. AB - We present a novel solid-state magic angle-spinning NMR method for measuring the NH(i)-NH(i+1) projection angle θ(i,i+1) in peptides. The experiment is applicable to uniformly (15)N-labeled peptides and is demonstrated on the chemotactic tripeptide N-formyl-l-Met-l-Leu-l-Phe. The projection angle θ(i,i+1) is directly related to the peptide backbone torsion angles φ(i) and psi(i). The method utilizes the T-MREV recoupling scheme to restore (15)N-(1)H interactions, and proton-mediated spin diffusion to establish (15)N (15)N correlations. T-MREV has recently been shown to increase the dynamic range of the (15)N-(1)H recoupling by gamma-encoding, and permits an accurate determination of the recoupled NH dipolar interaction. The results are interpreted in a quasi-analytical fashion that permits efficient extraction of the structural parameters. PMID- 10873505 TI - Hydrogen bonding effects on the (15)N and (1)H shielding tensors in nucleic acid base pairs. AB - The results of systematic ab initio calculations of (15)N and (1)H chemical shielding tensors in the GC base pair as a function of hydrogen bond length are presented for the first time. The hydrogen bond length characterized by the distance r(N...N) between purine N1 and pyrimidine N3 was varied between 2.57 and 3.50 A and the chemical shift tensors were calculated by the sum-over-states density functional perturbation theory. It is shown that the hydrogen bond length has a strong effect on the chemical shielding tensor of both imino proton and nitrogen, on their orientation, and, as a consequence, on the relaxation properties of both nuclei. For a nitrogen nucleus not involved in hydrogen bonding, the shielding tensor is nearly axially symmetric and almost collinear with the bond vector. As the length of the hydrogen bond decreases, the least shielding component sigma(11) deflects from the N-H vector and the shielding tensor becomes increasingly asymmetric. The significance of the presented results for the analysis of relaxation data and the efficiency of TROSY effects together with a summary of the relevant shielding parameters are presented and discussed. PMID- 10873507 TI - Spin-1/2 nucleus scalar coupled to quadrupolar nuclei: analysis of (77)Se spectra in 2,1,3-benzoselenadiazole PMID- 10873506 TI - Sensitivity-enhanced MQ-HCN-CCH-TOCSY and MQ-HCN-CCH-COSY pulse schemes for (13)C/(15)N labeled RNA oligonucleotides. AB - Sensitivity enhanced multiple-quantum 3D HCN-CCH-TOCSY and HCN-CCH-COSY experiments are presented for the ribose resonance assignment of (13)C/(15)N labeled RNA sample. The experiments make use of the chemical shift dispersion of N1/N9 of pyrimidine/purine to distinguish the ribose spin systems. They provide a complementary approach for the assignment of ribose resonance to the currently used HCCH-COSY and HCCH-TOCSY type experiments in which either (13)C or (1)H is utilized to separate the different ribose spin systems. The pulse schemes have been demonstrated on a 23-mer (13)C/(15)N-labeled RNA aptamer complexed with neomycin and tested on a 32-mer RNA complexed with a 23-residue peptide. PMID- 10873508 TI - Carbon-proton dipolar decoupling in REDOR AB - Dipolar decoupling of protons with radiofrequency field amplitudes comparable to those used for the rare-spin refocusing and dephasing pi pulses results in accurate, high-sensitivity determinations of internuclear distances in rotational echo double-resonance experiments and simulations performed on (13)C and (15)N labeled l-alanine. Copyright 2000 Academic Press. PMID- 10873509 TI - Endothelial cells scraped from the luminal surface of bovine pulmonary artery give rise to nonmuscle cells. PMID- 10873510 TI - Can streptokinase produce beneficial effects additional to coronary recanalization? Quantitative microvascular analysis of critically injured reperfused myocardium. AB - The aim of this study was to evaluate whether streptokinase (SK) may produce beneficial effects at the level of microvascular circulation in addition to coronary recanalization. Twenty mongrel dogs weighing 28.4 +/- 4.6 kg were randomized to receive SK (16,000-72,000 U/kg) or 20 ml saline (control) in an open-chest anterior descending artery occlusion (3 h) and reperfusion (2 h) model. Myocardial blood flow was measured by the radioactive microsphere technique and the state of microvascular circulation (red blood cell containing capillary counts and tissue red blood cell content) was evaluated in the infarcted subendocardial zone using computerized image analysis. The percentage (mean +/- SE) of cell-containing vessels normalized to nonischemic control areas was 166.5 +/- 7.5 in SK-treated infarcts while in untreated control infarcts it was more variable (130.0 +/- 15.6%) (2P > 0.1). The red blood cell content of infarcts treated with 2.0 megaunits SK was 3.9 +/- 0.6% compared with 6.7 +/- 0.9% in untreated control infarcts (2P = 0.029). Plasma viscosity was slightly reduced in SK-treated dogs (2P = 0.05), but no significant changes in blood fibrinogen, hemoglobin, blood flow, and high energy phosphate levels between control and SK-treated infarcts were observed. SK reduces congestion and results in more even reperfusion of the microvasculature in severely ischemic myocardium to which blood flow has been restored. This effect may be beneficial in the salvage and healing of clinical infarctions. PMID- 10873511 TI - Constriction of resistance arteries determines l-NAME-induced hypertension in a conscious hamster model. AB - The influence of infusion of a nitric oxide (NO) synthase inhibitor, N(omega) nitro-l-arginine methyl ester (l-NAME), on resistance arteries (diameter, 150 +/- 8 microm) and its relationship with hypertension were examined in conscious hamsters fitted with a dorsal skinfold window. After infusing l-NAME (10 and 30 mg/kg), hamsters showed immediate hypertension of +13 +/- 9 and +21 +/- 9 mm Hg, respectively, relative to basal values, and a maximum of +44 +/- 4 mm Hg at 30 min for the high-dose group. There was simultaneous significant vasoconstriction of the resistance arteries (A(0)) which reduced to 60 +/- 5% of baseline diameter at 3 h; however, there was no significant vasoconstriction in large and small arterioles with diameters diameters less than 70 microm. Blood flow rate in all the vessels decreased in consonance with the vasoconstriction of the resistance artery, irrespective of microvessel classification. These results indicate that the resistance artery plays a key role as a regulator and microvascular resistance in determining blood flow distribution and hypertension when a NO synthase inhibitor is infused. PMID- 10873512 TI - Relationship between pericardial pressure and lymphatic pericardial fluid transport in sheep. AB - We investigated the relationship between pericardial pressure and the volumetric lymphatic clearance rate of pericardial fluid in sheep. A single catheter perfusion system was established to deliver tracer to the pericardial cavity and control pericardial pressure. In addition, catheters were placed into the thoracic duct and into the jugular vein at the base of the neck. (125)I-human serum albumin (HSA) was administered into the pericardial perfusate to serve as the lymph flow marker and its concentration monitored in the effluent from the outflow end of the perfusion system. (131)I-HSA was injected intravenously to permit calculation of plasma tracer loss and tracer recirculation into lymphatics. From mass balance equations, estimates of total pericardial clearance into lymphatics increased significantly as pericardial pressures were elevated in 2. 5 cm H(2)O increments from 2.5 to 12.5 cm H(2)O (P = 0.018). Pericardial lymph transport ranged from 0.89 +/- 0.10 to 3.09 +/- 0. 66 ml/h at 2.5 and 12.5 cm H(2)O pericardial pressure, respectively. The majority of transport occurred through mediastinal vessels with a small proportion (10.3 to 23.9%) being cleared into lymphatics leading to the thoracic duct. We conclude that lymphatic pericardial fluid transport increases approximately 3.5-fold over a pericardial pressure range that encompasses the transition between the shallow and steep portions of the pericardial pressure-volume relationship. PMID- 10873513 TI - Impairment of flow-induced dilation of skeletal muscle arterioles with elevated oxygen in normotensive and hypertensive rats. AB - The effects of elevated PO(2) on flow-induced dilation of in situ skeletal muscle arterioles was assessed in cremaster muscle preparations from spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats. Blood flow increases in selected arterioles were initiated by occlusion of a parallel daughter branch from a parent arteriole. Changes in the diameter of the perfused arteriole were measured with a video micrometer and erythrocyte velocity was measured using optical Doppler velocimetry. Superfusate PO(2) was controlled by changing the O(2) concentration (0% O(2) or 21% O(2)) of the equilibration gas mixture. The increase in arteriolar diameter during occlusion was reduced in SHR compared to WKY rats, resulting in an elevated wall shear rate in SHR. Elevated PO(2) decreased flow-induced dilation in both groups and increased wall shear rate during parallel occlusion. An inhibitor of the formation of 20-HETE via cytochrome P450-4A enzymes (P450), dibromododecenyl methylsulfimide, minimized O(2)-induced constriction of arterioles and prevented the O(2)-induced decrease in flow-induced dilation and the increase in wall shear rate in both SHR and WKY rats. These results suggest that: (1) flow-induced dilation of in situ skeletal muscle arterioles is impaired in SHR compared to WKY, (2) elevated O(2) compromises flow-induced dilation in both groups, (3) 20-HETE contributes to both the O(2)-induced increases in resting tone and the reduced flow-induced dilation of cremasteric arterioles with elevated PO(2). PMID- 10873514 TI - Endothelin inhibits histamine-induced cyclic AMP accumulation in bovine brain vessels. AB - We have studied whether endothelin isopeptides have any effects on histamine induced cyclic AMP in [(3)H]adenine-prelabeled brain vessels isolated from bovine brain. Basal levels of [(3)H]cyclic AMP were enhanced by histamine in a concentration-dependent manner (EC(50) = 1.1 +/- 0.3 microM). Endothelin-1 inhibited histamine-elicited [(3)H]cyclic AMP generation with an IC(50) value of 3 +/- 2.5 nM. Sarafotoxin 6c, an ET-B receptor agonist, had no effect. ET-1 inhibition of histamine-induced [(3)H]cyclic AMP was reversed by the ET-A receptor antagonist BQ-123 while the ET-B receptor antagonist BQ-788 had no effect. The levels of [(3)H]cyclic AMP induced by isoprenaline were not altered by endothelin-1. Taken together, these results show that endothelins modulate the actions of histamine on the blood-brain barrier, probably by type A endothelin receptors. PMID- 10873515 TI - Pericyte migration from the vascular wall in response to traumatic brain injury. AB - Any perturbation of the blood brain barrier, whether from changes in cell physiology or from direct injury, may result in microvascular dysfunction and disease. We examined, at the ultrastructural level, microvascular pericyte responses in a well-defined model of traumatic brain injury in the rat. In areas close to the site of impact cortical pericytes underwent a number of changes within the first hour. Approximately 40% of pericytes migrated from their microvascular location. Migration occurred concomitant with a thinning of the abluminal surface of the basal lamina and an accumulation of the receptor for the urokinase plasminogen activator on the leading surface of the migrating cell. Migrated pericytes appeared viable and remained in a perivascular location in the adjacent neuropil. Nonmigrating pericytes in the same section displayed cytoplasmic alterations and nuclear chromatin changes consistent with a rapid degenerative process. PMID- 10873516 TI - Endothelial monocyte activating polypeptide II induces endothelial cell apoptosis and may inhibit tumor angiogenesis. AB - Endothelial monocyte activating polypeptide II (EMAP-II) is a tumor-derived cytokine with potent effects on endothelial cells in vitro and in vivo including upregulation of tissue factor and the sensitization of human melanoma to systemic TNF treatment via its effects on the tumor vasculature. We investigated the effects of EMAP-II on tumor growth, angiogenesis, vasculogenesis, and apoptosis. EMAP-II inhibited endothelial cell proliferation, vasculogenesis, and neovessel formation. In vivo growth of human melanoma lines expressing high amounts of EMAP II demonstrated slower growth, smaller tumors, and increased amounts of tumor necrosis than those expressing lower amounts of EMAP-II. EMAP-II induced endothelial-cell-specific apoptosis via a pathway that includes upregulation of the Fas-associated death domain and downregulation of Bcl-2. EMAP-II appears to have important effects on angiogenesis and may play a role in regulating tumor vascular growth. PMID- 10873517 TI - Pattern vision of the honeybee (Apis mellifera): blue and green receptors in the discrimination of translocation. AB - The visual discrimination of horizontal gratings by the honeybee (Apis mellifera) was studied in a Y-choice apparatus with fixed patterns presented vertically at a set range. Translocation in this context is the exchange of the positions of two different colored or black areas. This paper investigates what cues the bees have learned in this task. The patterns, made from combinations of calibrated colored papers, are designed to explore the parts played by the blue and green receptors when the boundary between the two colors provides contrast to only one receptor type. Horizontal translocation is not discriminated without contrast to the green receptors, but up/down translocation can be discriminated whatever the contrast at the boundary. The trained bees were tested on the same patterns made with different papers that included extreme changes in contrast. The results show that discrimination of up/down translocation involves green receptors and also blue receptors. When bees discriminate a translocation that shows contrast to only one type of receptor, they do not use the apparent brightness or the direction of the contrast to that receptor type acting alone. Instead, they discriminate the locations of colored areas irrespective of intensity differences or directions of contrasts. They use some measure of the photon flux at both receptor types and remember the difference between the colors and their locations. PMID- 10873518 TI - Chronic propranolol induces deficits in retention but not acquisition performance in the water maze in mice. AB - Agents that alter adrenergic receptors, such as "beta-blockers," also alter memory storage. However, reports suggest that beta-adrenergic receptor antagonists, such as propranolol, have conflicting behavioral effects with acute vs chronic dosing. This study was designed to evaluate the effects of chronic propranolol on retention for a spatial learning task. Adult male ICR mice were given daily injections of propranolol (2, 4, 8, or 12 mg/kg ip) or 0. 9% NaCl for 15 days prior to, and during, trials in a Morris water maze. Mice received five massed acquisition (escape) trials in each of three daily sessions, followed by a single 60-s probe trial on the fourth day. The location of the submerged platform was constant for each animal over acquisition trials, but varied across animals; starting position varied across trials. A 5 (dose) x 3 (trial blocks) mixed factorial ANOVA for escape time yielded a significant trial blocks effect only (p <.001), showing performance improving over sessions. Time spent in the target quadrant on the probe trial was shorter under all doses of propranolol when compared to vehicle group (all p <.001), indicating poorer retention of prior platform location. This effect, however, was not dose-related. Swim speed was not significantly affected by propranolol. These data demonstrate that chronic dosing with propranolol can impair retention of spatial learning, which cannot be attributed to reduced arousal or motor function. PMID- 10873519 TI - Effects of repetitive motor training on movement representations in adult squirrel monkeys: role of use versus learning. AB - Current evidence indicates that repetitive motor behavior during motor learning paradigms can produce changes in representational organization in motor cortex. In a previous study, we trained adult squirrel monkeys on a repetitive motor task that required the retrieval of food pellets from a small-diameter well. It was found that training produced consistent task-related changes in movement representations in primary motor cortex (M1) in conjunction with the acquisition of a new motor skill. In the present study, we trained adult squirrel monkeys on a similar motor task that required pellet retrievals from a much larger diameter well. This large-well retrieval task was designed to produce repetitive use of a limited set of distal forelimb movements in the absence of motor skill acquisition. Motor activity levels, estimated by the total number of finger flexions performed during training, were matched between the two training groups. This experiment was intended to evaluate whether simple, repetitive motor activity alone is sufficient to produce representational plasticity in cortical motor maps. Detailed analysis of the motor behavior of the monkeys indicates that their retrieval behavior was highly successful and stereotypical throughout the training period, suggesting that no new motor skills were learned during the performance of the large-well retrieval task. Comparisons between pretraining and posttraining maps of M1 movement representations revealed no task-related changes in the cortical area devoted to individual distal forelimb movement representations. We conclude that repetitive motor activity alone does not produce functional reorganization of cortical maps. Instead, we propose that motor skill acquisition, or motor learning, is a prerequisite factor in driving representational plasticity in M1. PMID- 10873520 TI - Background illumination effects upon in vitro conditioning in Hermissenda. AB - In the marine snail Hermissenda, associative learning can be accomplished by paired presentations of light and vestibular stimulation. It is generally assumed that associative learning depends upon the intensity or salience of the conditioned or unconditioned stimulus (CS and US, respectively). Accordingly, during Hermissenda conditioning a stronger dark adaptation is expected to render the CS (the light) more salient and hence facilitate association. We studied the influence of background illumination level using an in vitro pairing procedure in Hermissenda. This procedure allows one to assess the effect of conditioning upon a single cell, the B photoreceptor, which is implicated in this learning process. After 15 min of adaptation to a dim background light, B photoreceptors maintained a basal rate of firing, while after adaptation to complete darkness, they stopped firing. Paired and unpaired groups received 10 training trials in either a completely dark or a dim light environment. Although a trial to trial cumulative increase in excitability was found in the paired group trained in darkness, only the paired group trained under dim background light showed a higher input resistance and cell excitability 10 min after training. These results suggest that the background dim illumination was not needed for the induction but played a role in the maintenance of the pairing effect. Possible mechanisms for such a modulatory effect are discussed. PMID- 10873521 TI - Effects of glucose on scopolamine-induced learning deficits in rats performing the Morris water maze task. AB - In order to assess the effects of glucose on drug-induced spatial learning deficits, three experiments were conducted using the Morris water maze. Scopolamine and glucose were injected ip at various stages of training. Rats of Wistar strain served as subjects. In Experiment 1, scopolamine (0.4 mg/kg) and 10, 100, or 500 mg/kg of glucose were administered every day from the start of training, and the effect on acquisition was evaluated. In Experiment 2, scopolamine and 100 or 500 mg/kg of glucose were administered after 6 days of training, and the effect on performance was assessed. In Experiment 3, scopolamine and 500 mg/kg of glucose were injected after 2 days of training, and the effect on the following trial was tested. In all experiments, scopolamine impaired acquisition/performance of the task. Glucose at 500 mg/kg showed a significant enhancing effect on acquisition regardless of scopolamine injection only when injected daily from the start of training (Experiment 1). Glucose injected after the performance has reached asymptote (Experiment 2) did not affect performance, and glucose in the middle of training showed a slight but insignificant enhancing effect (Experiment 3). These results may suggest that the effect of glucose changes as a function of the degree of learning of the spatial learning task. The possibility of task specificity of the glucose effect was also discussed in relation to the cholinergic systems and local cerebral glucose utilization. PMID- 10873522 TI - pT3.2I, the smallest plasmid of Thiobacillus T3.2. AB - The whole nucleotide sequence of pT3.2I, the smallest plasmid of the acidophilic bacterium Thiobacillus T3.2, has been determined. pT3.2I is 15,390 bp long with a 53.7% GC content. Different regions can be defined in it: one 2569-bp putative insertion sequence similar to other insertion sequences of some Agrobacterium Ti plasmids; and a longer sequence, which occurs in two almost identical copies, differing only in a 1-bp deletion (6406 and 6405 bp). Several open reading frames and some smaller sequences were found in this duplicated region: ORFA and ORFG, encoding a putative polyol dehydrogenase and a putative RepA replication protein, respectively, an 83-bp sequence which could code for an antisense RNA, and a 36 bp region highly homologous to ori sequences of ColE2- and ColE3-related plasmids. Another putative gene, ORFH, is only present in the longer copy of this region (it is deleted in the short copy) and might encode a 90-amino-acid polypeptide which could act as a second replication protein, RepB. Based on sequence comparisons, pT3. 2I can be related to plasmids in the pColE2-CA42 incB incompatibility group. PMID- 10873523 TI - Nucleotide sequence of a 7-kb fragment of pACM1 encoding an IncM DNA primase and other putative proteins associated with conjugation. AB - A 7-kb fragment of pACM1 (fragment 90?91) containing one or more kor (kill override) loci was sequenced, and 28 open reading frames (ORFs; >/=50 codons) were identified. The nucleotide sequence has no significant homologs in the GenBank database except for a 1.3-kb region 98.6% identical to the iml (insensitivity to phage PhiM-mediated lysis) determinant fragment of IncM plasmid R446. Deduced amino acid sequences for several ORFs are homologous to those of known proteins, including the Sog DNA primases of IncI1 plasmids R64 and ColIb-P9 and the TraL, TraM, and TraN products of ColIb-P9. Two protein products of the putative primase ORF (ORF 1, 1100 amino acids) were detected by SDS-PAGE. The 158 and 107-kDa proteins were designated PriL and PriS, respectively. PriS is apparently produced by an in-frame reinitiation of the ORF 1 transcript at a second start codon located between a Sau96I site and a PstI site. The motif EGYATA, conserved among primases and associated with primase function, occurs in the first one-third of the deduced amino acid sequence of PriL and is not included in PriS. Partial suppression of the temperature-sensitive dnaG3 mutation in BW86 was demonstrated by recombinants that overexpressed both PriL and PriS, but not by constructs overexpressing only PriS. Therefore, primase function can be assigned to PriL. Fragment 90/91 represents a portion of the IncM tra region, which has not previously been examined in detail. PMID- 10873524 TI - The virulence plasmid of Salmonella typhimurium contains an autoregulated gene, rlgA, that codes for a resolvase-like DNA binding protein. AB - The virulence plasmid of Salmonella typhimurium contains a gene, rlgA, that shows strong homology to several reported resolvase-like proteins. This gene maps 5 kb upstream of spv locus, the major virulence determinant on the plasmid. Regulation of rlgA was studied using a lacZ transcriptional reporter fusion. The rlgA gene was found to be repressed at the level of transcription by its own product and to be expressed maximally in the late exponential phase of growth. The transcription start site of the rlgA gene was determined and the RlgA binding site was mapped and found to overlap with the transcription initiation signals. A derivative of the virulence plasmid was constructed with a knockout mutation in rlgA. This mutation did not alter the stability of the virulence plasmid nor did it affect the ability of S. typhimurium to cause systemic disease in mice. PMID- 10873525 TI - In Rhizobium etli symbiotic plasmid transfer, nodulation competitivity and cellular growth require interaction among different replicons. AB - Bacteria belonging to the genus Rhizobium are able to develop two different lifestyles, in symbiotic association with plant roots or through saprophytic growth. The genome of Rhizobium strains is constituted by a chromosome and several large plasmids, one of them containing most of the genes involved in symbiosis (symbiotic plasmid or pSym). Our model strain Rhizobium etli CFN42 contains six plasmids. We have constructed multiple plasmid-cured derivatives of this strain and used them to analyze the contribution of these plasmids to free living cellular viability, competitivity for nodulation, plasmid transfer, and utilization of diverse carbon sources. Our results show that the transfer of the pSym is strictly dependent on the presence of another plasmid; consequently under conditions where pSym transfer is required, nodulation relies on the presence of a plasmid devoid of nodulation genes. We also found a drastic decrease in competitivity for nodulation in multiple plasmid-cured derivatives when compared with single plasmid-cured strains. Cellular growth and viability were greatly diminished in some multiple plasmid-cured strains. The utilization of a number of carbon sources depends on the presence of specific plasmids. The results presented in this work indicate that functional interactions among sequences scattered in the different plasmids are required for successful completion of both lifestyles. PMID- 10873526 TI - The novel insertion sequences IS1417, IS1418, and IS1419 from Burkholderia glumae and their strain distribution. AB - Three insertion sequences, IS1417, IS1418, and IS1419, were isolated from Burkholderia glumae (formerly Pseudomonas glumae), a gram-negative rice pathogenic bacterium, on the basis of their abilities to activate the expression of the neo gene of the entrap vector pSHI1063. The 1335-bp IS1417 element with 17 bp imperfect terminal inverted repeats was found to be flanked by 5-bp direct repeats of the vector sequence. IS1418 is 865 bp in length and carries 15-bp inverted repeats with a target duplication of 3 bp. The 1215-bp IS1419 sequence is bounded by the 36-bp terminal inverted repeats of the element and 7-bp direct repeats of the vector sequence. IS1417 and IS1418 belong to the IS2 subgroup of the IS3 family and the IS427 subgroup of the IS5 family, respectively, whereas IS1419 does not appear to be a member of any known IS family. Southern blot analysis of DNAs from B. glumae field isolates indicated that those IS elements are widely distributed, but the host range of the three IS elements appears to be limited to B. glumae and some other related species such as B. plantarii. The polymorphisms exhibited in B. glumae isolates suggest that those elements are useful for molecular epidemiological studies of B. glumae infections. PMID- 10873527 TI - pUB2380: characterization of a ColD-like resistance plasmid. AB - A detailed analysis of the mobilizable, ColE1-like resistance plasmid, pUB2380, is reported. The 8.5-kb genome encodes six (possibly seven) major functions: (1) a ColD-like origin of replication, oriV, with associated replication functions, RNAI and RNAII; (2) a set of active mobilization functions highly homologous to that of ColE1, including the origin of transfer, oriT; (3) a ColE1-like multimer resolution site (cer); (4) a kanamycin-resistance determinant, aph, encoding an aminoglycoside-3'-phosphotransferase type 1; (5) an insertion sequence, IS1294; and (6) two genes, probably cotranscribed, of unknown function(s). The GC content of the various parts of the genome indicates that the plasmid is a hybrid structure assembled from DNA from at least three different sources, of which the replication region, the mobilization functions, and the resistance gene are likely to have originated in the enterobacteriaceae. PMID- 10873529 TI - Sequencing and characterization of the cryptic plasmid QpRS from Coxiella burnetii. AB - Plasmid QpRS from Coxiella burnetii, isolate Priscilla Q177, phase I, has been completely sequenced. DNA for sequencing was amplified with "extra-long" (XL) PCR. The size of the QpRS plasmid sequence was determined to be 39,280 bp, with a G + C content of 39.3%. Putative proteins associated with replication and recombination were identified. The sequence of QpRS plasmid was analyzed for shared and unique regions among C. burnetii plasmids. PMID- 10873528 TI - IS1294, a DNA element that transposes by RC transposition. AB - IS1294, found on the ColD-like resistance plasmid pUB2380, is IS91-like. It is an active 1.7-kb insertion sequence that lacks terminal inverted repeats, displays insertion-site specificity, and does not generate direct repeats of the target site. The element has one large open reading frame, tnp(1294), encoding a transposase of 351 amino acids, related to members of the REP family of replication proteins used by RC-plasmids of gram-positive bacteria. IS1294 transposes using rolling-circle replication, initiated at one end of the element, oriIS, and terminated at the other, terIS. oriIS and terIS are highly conserved among like IS elements. oriIS resembles the leading strand replication origins of RC-plasmids; terIS resembles a rho-independent transcription terminator. IS1294 mediates not only its own transposition, but also sequences adjacent to terIS. A transposition model for IS1294 and related elements, involving rolling-circle replication and single-strand DNA intermediates, is presented. PMID- 10873530 TI - Tungsten particle-induced nicking of supercoiled plasmid DNA. AB - Small particles of metallic tungsten, known also as tungsten microprojectiles, are routinely used for biotechnological purposes. In such applications, tungsten was observed to affect the integrity of plasmid DNA. Here we present evidence that interaction between tungsten particles and intact circular plasmids pU19, pUC119, and ColE1 may result in generation of a limited number of single-strand DNA breaks. As a consequence, supercoiled DNA is converted into its open circular form and no fragmentation products can be detected. The rate of the tungsten mediated reaction depends on pH but is not influenced by ascorbate, Tris, or EDTA. No DNA nicking can be observed when the tungsten particles are replaced by substances that can be leached out from these particles with water or incubation buffers. Likewise, commercial sodium tungstate, tungsten (VI) oxide, and tungsten (VI) chloride and products of its decomposition remain DNA undamaged. Native plasmid DNA molecules, upon adsorption on the surface of tungsten microparticles, may undergo some nicking without a need for participation of external catalysts. PMID- 10873531 TI - Highly conserved DNA sequence present in small plasmids from Selenomonas ruminantium. AB - Plasmid pJW1 from Selenomonas ruminantium subsp. lactilytica strain JW13 has been cloned in Escherichia coli vector pBluescriptSK(-) and completely sequenced. The plasmid is only 1410 bp with an overall GC content of 42.2%. Computer analysis of sequence data revealed a single open reading frame (ORF1, 146 amino acids, MW 16,525.5 Da) encoding a putative replication protein which is similar to the Rep protein of Ruminobacter amylophilus plasmid pRAO1. ORF1 is followed by a long AT rich (75%) region and a region abundant in direct and inverted repeats. Comparison of DNA sequences revealed the presence of a short (<250 bp) DNA segment which is highly conserved between several small S. ruminantium plasmids including pJDB21. PMID- 10873532 TI - A method for demonstrating gene essentiality in Staphylococcus aureus. AB - A method for demonstrating whether a gene of Staphylococcus aureus is essential for growth in a rich medium is described. We have used this method to determine whether the murE gene, which encodes the UDP-N-acetylmuramyl tripeptide synthetase required for peptidoglycan synthesis, is essential for growth in S. aureus. In this study, strain CYL368 was constructed from S. aureus RN4220 by placing the murE gene in the chromosome under the control of the spac promoter (a hybrid promoter of the Escherichia coli lac operator and the Bacillus subtilis SPO1 phage promoter). To regulate the murE gene in CYL368, the E. coli lacI gene was expressed from the B. licheniformis penicillinase gene (pcn) promoter in plasmid pMJ8426. Strain CYL368(pMJ8426) grew normally in the presence of isopropyl-beta-d-thiogalactopyranoside but could not grow in the absence of the inducer. These results indicate that the murE gene expressed from the spac promoter in CYL368(pMJ8426) is needed for bacterial growth. We concluded that murE is an essential gene of S. aureus. PMID- 10873533 TI - Genetic analysis and complete nucleotide sequence of a 2-kb cryptic plasmid from the marine methylotroph Methylophaga thalassica S1. AB - A small cryptic plasmid (pMTS1) was isolated from the marine methylotroph Methylophaga thalassica S1. Sequence analysis showed that pMTS1 has a 1995-bp genome encoding three putative polypeptides and containing two intergenic regions. The longest open reading frame encodes a polypeptide of 325 amino acid (37,079 Da). This polypeptide shares similarity and signature amino acids with the rep proteins of the pC194 group replicons, suggesting that pMTS1 replicates by the rolling-cycle mechanism. Insertional mutagenesis confirmed that the repA gene product is obligatory for replication of pMTS1 in M. thalassica S1. A pC194 type dso was identified in the second intergenic area. It appears that this intergenic region contains both the nic and bind sites. It is hypothesized that sso of pMTS1 is located in the first intergenic region. Two smaller open reading frames (orf-1 and orf-2) encode transcriptionally coupled polypeptides of 123 and 101 amino acids (14,486 and 11,241 Da, respectively) with unknown biological function. PMID- 10873534 TI - Refolding and characterization of rat liver methionine adenosyltransferase from Escherichia coli inclusion bodies. AB - Methionine adenosyltransferase (MAT) catalyzes the synthesis of S adenosylmethionine, the major methyl donor for transmethylation reactions. Attempts to perform structural studies using rat liver MAT have met with problems because the protein purified from cellular extracts is heterogeneous. Overexpression of the enzyme in Escherichia coli rendered most of the protein as inclusion bodies. These aggregates were purified by specific washes using urea and Triton X-100 and used for refolding. Maximal activity was obtained when chaotropic solubilization included the structural cation Mg(2+), the protein concentration was kept below 0.1 mg/ml, and denaturant removal was carried out in a two-step process, namely, a fast dilution followed by dialysis in the presence of 10 mM DTT or GSH/GSSG redox buffers. Refolding by this procedure generated the oligomeric forms, MAT I and III, which were basically indistinguishable from the purified rat liver forms in secondary structure and catalytic properties. PMID- 10873535 TI - Expression, purification, and functional analysis of the human serine protease HtrA2. AB - HumHtrA2 or Omi is a recently described member of a novel family of mammalian serine proteases homologous to the Escherichia coli htrA gene product. Although the physiological function of members of this new family is unclear, the current understanding is that as well as being involved with the degradation aberrantly folded proteins during conditions of cellular stress, they may possess a chaperone-like role under normal conditions. In this report we describe the overexpression of humHtrA2 in two heterologous systems comparing the merits of each. We found that molecular analysis of processing events in Sf9 cells allowed us to revisit E. coli expression systems which were initially unsuccessful. Using E. coli we were able to produce milligram amounts of >90% pure recombinant enzyme as determined by SDS-PAGE gels. By means of fluorescently labeled substrates alpha- and beta-casein and zymography, the proteolytic activity of recombinant HumHtrA2 was also demonstrated. PMID- 10873536 TI - Overexpression and biosynthetic deuterium enrichment of TEM-1 beta-lactamase for structural characterization by magnetic resonance methods. AB - An expression system has been developed that allows high levels of production of TEM-1 beta-lactamase with ease of biosynthetic incorporation of nuclear isotopes. The gene for mature TEM-1 beta-lactamase fused to the leader sequence of the ompA protein was subcloned into the pET-24a(+) vector by introduction of an NdeI restriction site at the first codon of the fused genes and transformed into Escherichia coli BL21 (DE3) cells. With protein induction at 25 degrees C supported by LB medium supplemented with osmolytes (300 mM sucrose and 2.5 mM betaine), the extracellular, mature form of wild-type TEM-1 beta-lactamase was recovered at a level of 140 mg/L. The production level of E166N, E240C, E104C, and M272C mutants depended on the mutation but was invariably higher than reported by others for expression systems of the wild-type enzyme. Comparison of different carbon sources on the efficiency of biosynthetic incorporation of covalent deuterium showed maximal (90%) incorporation with minimal medium containing 99% (2)H(2)O and sodium d(3)-acetate (99 atom% (2)H). The yield of deuterium-enriched wild-type enzyme was 80 mg/L with yields for mutants proportionally reduced. The high level of protein deuteration achieved with this system allowed detection of the hyperfine coupling between the paramagnetic nitroxyl group of a spin-labeled penicillin substrate and hydrogens on the penicillin moiety in a cryokinetically isolated acylenzyme reaction intermediate because of the decrease in overlapping resonances of active site residues. The overexpression system is readily adaptable for other target proteins and facilitates studies requiring large quantities of protein in isotopically enriched forms. PMID- 10873537 TI - An analysis of two refolding routes for a C-terminally truncated human collagenase-3 expressed in Escherichia coli. AB - We describe here the expression of a C-terminally truncated form of human procollagenase-3 in Escherichia coli. The protein was found almost exclusively in inclusion bodies that were solubilized and refolded by two separate methods and then purified on Ni-NTA agarose. The purified proenzyme could be activated with either trypsin or APMA and active enzyme could be purified on a peptidic hydroxamate affinity column. Competitive elution from the affinity matrix yielded a highly purified preparation. PMID- 10873538 TI - Expression of the Arabidopsis thaliana AtJ2 cochaperone protein in Pichia pastoris. AB - A vector was constructed for intracellular expression of the Arabidopsis thaliana DnaJ homologue AtJ2 in the methylotrophic yeast Pichia pastoris. The vector includes DNA encoding an amino-terminal histidine-tag, to simplify protein purification. Shake-flask cultures could be induced to produce approximately 250 mg/ L of AtJ2. Purified recombinant AtJ2 was able to stimulate the ATPase activities of both the Escherichia coli and Zea mays cytoplasmic Stress70 chaperone proteins five- to ninefold. The carboxy terminus of AtJ2 is -CAQQ, a protein farnesylation motif. When transformed P. pastoris was induced to synthesize AtJ2 in the presence of [(3)H]mevalonolactone, radioactivity was incorporated into the protein, suggesting farnesylation. PMID- 10873539 TI - Expression and characterization of recombinant Rhodocyclus tenuis high potential iron-sulfur protein. AB - The high potential iron-sulfur protein (HiPIP) from Rhodocyclus tenuis strain 2761 has been overproduced in Escherichia coli from its structural gene, purified to apparent homogeneity, and then characterized by an array of methods. UV visible spectra of the reduced and oxidized recombinant protein were similar to those of the native protein. EPR of the oxidized protein shows g values of 2. 11, 2.03, and 2.03. ESI-MS gave a mass difference of 350 Da between the holoprotein and acid-treated protein, consistent with incorporation of a [Fe(4)S(4)] cluster in the holoprotein. The observed mass of the apoprotein was 6296.6 Da compared to the expected average molecular mass of 6297.2 Da of the apoprotein. The reduction potential was determined using cyclic and square-wave voltammetry to be 321 and 314 mV versus NHE, respectively. All the observed properties of the recombinant protein parallel those of the native protein or those of native HiPIPs in general, indicating correct folding and incorporation of the iron-sulfur cluster. PMID- 10873540 TI - High-level expression of Escherichia coli and Bacillus subtilis thymidylate synthases. AB - Procedures are described for the preparation of highly purified thymidylate synthases from Escherichia coli and Bacillus subtilis. The yields in each case are quite high with about 350 mg of pure protein obtained from 1 liter of cells. Basically all that is required to obtain pure enzyme is an induction step from a high-expression vector, followed by a DE-52 column elution. Both enzymes appeared to be fairly stable in that incubation at 43 degrees C for 10 min resulted in the loss of 50% of the E. coli thymidylate synthase activity, while 50 degrees C for 10 min was required to obtain the same effect with the B. subtilis enzyme. In the presence of the substrate, dUMP, each protein was stabilized further by 6 to 7 degrees C, which was increased to 9 to 10 degrees C on addition of dihydrofolate. It was shown also that the E. coli thymidylate synthase could be maintained at 4 degrees C for at least 4 months with little or no loss in activity provided that mercaptoethanol was not present. The presence of the latter led to a progressive loss in activity until little activity could be detected after 18 weeks, which was due, in part, to the formation of a disulfide bond with the active site cysteine. Addition of dithiothreitol restored the enzyme activity to its original state. PMID- 10873541 TI - Expression and purification of recombinant neurotensin in Escherichia coli. AB - An expression system has been designed for the rapid and economic expression of recombinant neurotensin for biophysical studies. A synthetic gene for neurotensin (Glu(1)-Leu(2)-Tyr(3)-Glu(4)-Asn(5)-Lys(6)-Pro(7)-Arg(8)-Arg(9)-Pro(1 0)-Tyr(11) Ile(12)-Leu(13)) was cloned into the pGEX-5X-2 vector to allow expression of neurotensin as a glutathione S-transferase (GST) fusion protein. The inclusion of a methionine residue between the glutathione S-transferase and the neurotensin has facilitated the rapid cleavage of the neurotensin from its carrier protein. Purification of recombinant neurotensin was performed by reverse-phase HPLC. This method produced a relatively high yield of peptide and offers the potential for economic partial or uniform labeling of small peptides (<15 amino acids) with isotopes for NMR or other biophysical techniques. PMID- 10873542 TI - Pisum sativum mitochondrial pyruvate dehydrogenase can be assembled as a functional alpha(2)beta(2) heterotetramer in the cytoplasm of Pichia pastoris. AB - Pea (Pisum sativum) mitochondrial pyruvate dehydrogenase (E1) was produced by coexpression of the mature alpha and beta subunits in the cytoplasm of the yeast Pichia pastoris. Size-exclusion chromatography of recombinant E1, using a Superose 12 column, yielded a peak at M(r) 160,000 that contained both alpha and beta subunits as well as E1 activity. This corresponds to the size of native alpha(2)beta(2) E1. Recombinant E1 alpha (His(6))-E1 beta was purified by affinity chromatography using immobilized Ni(+), with a yield of 2.8 mg L(-1). The pyruvate-decarboxylating activity of recombinant E1 was dependent upon added Mg(2+) and thiamin-pyrophosphate and was enhanced by the oxidant potassium ferricyanide. Native pea mitochondrial E1-kinase catalyzed phosphorylation of Ser residues in the alpha-subunit of recombinant E1, with concomitant loss of enzymatic activity. Thus, mitochondrial pyruvate dehydrogenase can be assembled in the cytoplasm of P. pastoris into an alpha(2)beta(2) heterotetramer that is both catalytically active and competent for regulatory phosphorylation. PMID- 10873543 TI - Heterodimeric complex of RAR and RXR nuclear receptor ligand-binding domains: purification, crystallization, and preliminary X-ray diffraction analysis. AB - Both the human retinoic acid receptor alpha (hRARalpha) and a constitutively active mutant (F318A) of the mouse retinoid X receptor alpha (mRXR alpha F318A) ligand-binding domains were separately overexpressed in Escherichia coli, copurified as a heterodimer in a two-step procedure, and cocrystallized with an RAR alpha-specific antagonist by using polyethylene glycol 10,000 as precipitant. The crystals grew in the hexagonal space group P6(1)22 displaying the unit cell parameters a = b = 116.6 A and c = 207.8 A. They diffracted X-ray to a limit of 2.2-A resolution. The asymmetric unit comprises one heterodimer and the crystal contains 60% solvent. The structure was determined by molecular replacement and is currently being refined. PMID- 10873544 TI - Identification, purification, and characterization of a thermophilic imidase from pig liver. AB - This study investigates thermophilic imidase activity of the liver. We demonstrate that imidase catalyzes the hydrolysis of imides at a temperature substantially higher than that of its native environment. Then, a thermophilic imidase is purified to homogeneity from pig liver, and its thermoproperties are studied. About 2500-fold of purification and 15% yield of imidase activity are obtained after ammonium sulfate precipitation, octyl, DEAE, chelation, and gel filtration chromatography. While avoiding heat treatment for the protein purification, this study also indicates that only one enzyme is responsible for the imidase activity. This homogenous enzyme prefers to catalyze hydrolysis of imides at above 60 degrees C rather than at the body temperature of a pig. Although stable at below 50 degrees C, imidase quickly loses its activity at above 65 degrees C. Thus, the temperature effect on imidase activity is limited mainly by its thermostability. Substrate specificity of imidase is also temperature dependent. Our results demonstrate that the hydrolysis of physiological substrates is the most temperature dependent and that of hydantoins is the least temperature dependent. When increasing the reaction temperature from 25 to 60 degrees C, specific activities increase 50- and 60-fold for dihydrouracil and dihydrothymine, respectively. The temperature effect on the K(m) and V(max) of imidase is substrate dependent. PMID- 10873545 TI - High expression of glycogen-debranching enzyme in Escherichia coli and its competent purification method. AB - Glycogen-debranching enzyme (GDE) gene from Saccharomyces cerevisiae was cloned and expressed into Escherichia coli. A 99.3% homology was found between the nucleotide sequences of GDE gene harbored in the recombinant E. coli plasmid (pTrc99A) and the open reading frame (902039-906646 position) of the 4608-bp fragment of S. cerevisiae chromosome XVI. We investigated the best conditions for GDE expression. When the cultivation temperature of recombinant E. coli strains was lowered to 25 degrees C and the isopropyl-beta-d-thiogalactopyranoside (IPTG) concentration used for induction was decreased to as low as 0.02 mM, a total of about 33 mg of recombinant GDE can be isolated from a liter culture as estimated by amylo-1,6-glucosidase activity. Consecutively, we developed a new method for purifying GDE. The method requires only a single-step purification via beta cyclodextrin-immobilized Sepharose 6B (beta-CD Sepharose 6B) affinity chromatography and renders a 90% recovery of the enzyme. Moreover, the purified recombinant GDE is a homogeneous protein and possesses the same characteristics as those of S. cerevisiae. With the highly expressed GDE in recombinant E. coli and a rapid and effective purification method, we successfully resolved the hurdle always faced for obtaining an ample amount of purified GDE. The availability of GDE, hence, may allow advancement on GDE studies and provide new prospects for GDE on biotechnological application. PMID- 10873547 TI - Controlled intracellular processing of fusion proteins by TEV protease. AB - Here we describe a method for controlled intracellular processing (CIP) of fusion proteins by tobacco etch virus (TEV) protease. A fusion protein containing a TEV protease recognition site is expressed in Escherichia coli cells that also contain a TEV protease expression vector. The fusion protein vector is an IPTG inducible ColE1-type plasmid, such as a T7 or tac promoter vector. In contrast, the TEV protease is produced by a compatible p15A-type vector that is induced by tetracyclines. Not only is the TEV protease regulated independently of the fusion protein, but its expression is highly repressed in the absence of inducer. Certain fusion partners have been shown to enhance the yield and solubility of their passenger proteins. When CIP is used as a purification step, it is possible to take advantage of these characteristics while both eliminating the need for large amounts of pure protease at a later stage and possibly simplifying the purification process. Additionally, we have observed that in some cases the timing of intracellular proteolysis can affect the solubility of the cleaved passenger protein, allowing it to be directed to either the soluble or the insoluble fraction of the crude cell lysate. This method also makes it possible to quickly gauge the efficiency of proteolysis in vivo, before protein purification has begun and in vitro processing is attempted. PMID- 10873546 TI - Expression of an anti-CD3 single-chain immunotoxin with a truncated diphtheria toxin in a mutant CHO cell line. AB - ADP-ribosylating immunotoxins are generally expressed in Escherichia coli and then refolded in vitro. Because the efficiency of the in vitro refolding process decreases with the number of protein domains and internal disulfide bonds, these immunotoxins have been generally limited to single-chain monovalent structures. We now show that using the hamster cell line CHO K1 RE1.22c (J. M. Moehring and T. J. Moehring, 1979, Somat. Cell Genet. 5, 453-468) that has been mutated to ADP ribosylation insensitivity, a level of 4 microg/ml of a truncated anti-T cell immunotoxin, DT390-scFvUCHT1, can be secreted into the medium. This immunotoxin is glycosylated at the two potential N-linked glycosylation sites in the toxin moiety: positions 16-18 in the A chain and residues 235-237 in the B chain. The glycosylated immunotoxin is relatively nontoxic (IC(50) 4.8 x 10(-10) M). Removal of the N-linked oligosaccharides by N-glycosidase F treatment or mutations at the two N-linked glycosylation sites results in a highly active immunotoxin with an IC(50) of 4 x 10(-12) M toward CD3(+) Jurkat cells. This is a 12-fold increase in toxicity over the same immunotoxin harvested from E. coli periplasm without refolding. A single Asn(235) Ala mutation that removed the B chain glycosylation was nearly as toxic as the double mutant. This suggests that B chain glycosylation is the major cause for the loss of toxicity. PMID- 10873548 TI - Potential role of EDG receptors and lysophospholipids as their endogenous ligands in the respiratory tract. AB - The role of lipid mediators derived from membrane glycerophospholipids and sphingolipids as intracellular messenger has been studied intensively during the last two decades, but with the recent discovery of high affinity G-protein coupled receptors for the lysophospholipids lysophosphatidic acid (LPA), sphingosine-1-phosphate (S1P) and sphingosylphosphorylcholine (SPC), increasing attention has been paid to the role of these lipid mediators as extracellular mediators. This review will summarize the biosynthesis and metabolism of lysophospholipids and describe the family of endothelial differentiation gene (EDG) receptors as high affinity receptors for lysophospholipids. Furthermore, an overview of the numerous biological effects of lysophospholipids which might be mediated by EDG receptors will be given together with an outlook on the potential role of such mechanisms in pulmonary physiology and pathophysiology. PMID- 10873549 TI - Molecular mechanisms of glucocorticosteroid actions. PMID- 10873550 TI - Importance of beta(2)adrenergic receptor genotype, gender and race on albuterol evoked bronchodilation in asthmatics. AB - To determine whether race and gender affect beta(2)receptor-stimulated bronchodilation, we quantified FEV(1)and plasma concentrations of albuterol at various times following the oral administration of a single 8-mg dose of albuterol in 15 black and 15 white male and female asthmatics. No important racial or gender differences in albuterol-evoked FEV(1)or percent-predicted FEV(1)were evident, although females tended to be more sensitive compared to males. Pharmacodynamic (PD) models were fitted to data in 19 patients (63%); FEV(1)was too erratic to fit in three, and a clockwise hysteresis in the FEV(1)vs. albuterol concentration relationship was observed in eight asthmatics. Mean +/- SD baseline (E(0)), maximal FEV(1)(E(max)) and C(50)were: 3.18 + 1.03 l, 4.00 +/- 1.12 l, 7.84 +/- 10.2 microg/l, respectively. beta(2)receptor genotype was determined in 16 patients. All Arg 16 homozygotes exhibited proportional FEV(1)response vs. plasma albuterol concentration relationships, and thus were fitted by PD models. All those having a poor FEV(1)vs. albuterol concentration relationship carried the Gly 16 allele. We conclude that receptor genotype, but not race or gender, is an important determinant of albuterol pharmacodynamics. PMID- 10873551 TI - Importance of local production of endothelin-1 and of the ET(B)Receptor in the regulation of pulmonary vascular tone. AB - Interaction between locally released endothelin-1 (ET-1) and the endothelial ET(B)receptor could modulate pulmonary vascular tone. We evaluated pulmonary ET-1 clearance and ET-1-ET(B)receptor interaction in the modulation of pulmonary vascular tone. Controls and rats with Monocrotaline (MCT)-induced pulmonary hypertension (PH) were studied. Lungs were isolated and perfused under constant pressure. The effect of the selective ET(B)antagonist BQ-788 (10(-12)-10(-8)mole) on perfusion flow rate and(125)I-ET-1 extraction was determined. Baseline(125)I ET-1 extraction was reduced from 62+/-5% in controls to 49+/-10% in PH (P=0.012). BQ-788 inhibited extraction with a higher half-inhibitory dose in the MCT group ( Log ID(50)= 8.9+/-0.4 vs. 9.5+/-0.1, P=0.03). BQ-788 induced a mild reduction in perfusion flow rate of 0.7+/-0.3 ml/min in controls. In the MCT group, this occurred at a lower dose and was more pronounced with a maximal reduction of 3.3+/-0.7 ml/min (P<0.01 vs. control). ET-1 was undetectable in the effluent at baseline but was present in similar concentrations in both groups after ET(B)blockade. Addition of 2 pg/ml ET-1 to lung perfusate did not modify pulmonary ET-1 clearance or the effect of BQ788 on perfusion flow rate in control lungs. In normal rat lungs, the ET(B)receptor plays a minor regulatory role on vascular tone. In MCT hypertension however, despite a reduction in ET(B)mediated extraction, luminal production of ET-1 attenuates the increase in pulmonary vascular tone. PMID- 10873552 TI - Phosphodiesterase inhibitors and forskolin Up-regulate arginase activity in rabbit alveolar macrophages. AB - Alveolar macrophages (AMsmall ef, Cyrillic) express considerable arginase activity which can be modulated by various mediators. As inhibitors of phosphodiesterase (PDE) play an increasing role in the treatment of chronic inflammatory and obstructive airway disease, we tested whether PDE inhibitors affect arginase activity in AMsmall ef, Cyrillic. Isolated rabbit AMsmall ef, Cyrillic were cultured for 20 h in the absence or presence of bacterial lipopolysaccharides (LPS) and/or different test substances. Thereafter arginase activity was determined by measuring the formation of [(3)H]-L-ornithine during 1 h incubation with [(3)H]-L-arginine. Lipopolysaccharide-enhanced (0. 01-5 microg/ml) maximal arginase activity by about 2.5-fold. The non-selective PDE inhibitor IBMX and the PDE4 selective inhibitor rolipram (each up to 30 microM) caused a 2.4-fold increase in arginase activity, and these effects were additive to those of LPS. The PDE3-selective inhibitor siguazodan had only marginal effects. Forskolin (10 microM) also enhanced arginase activity in the absence and presence of LPS. The effect of forskolin was almost prevented by cycloheximide (30 microM) and largely attenuated by the protein kinase A inhibitor KT 5720 (300 nM). In conclusion, inhibition of the cAMP-specific PDE4, like direct activation of adenylyl cyclase, causes an up-regulation of arginase activity in rabbit AMsmall ef, Cyrillic. PMID- 10873553 TI - Transcriptional homeostatic control of membrane lipid composition. AB - Plasma membranes have a structural property, commonly referred to as membrane fluidity, that is compositionally regulated. The two main features of plasma membrane lipid composition that determine membrane fluidity are the ratio of cholesterol to phospholipids and the ratio of saturated to unsaturated fatty acids that are incorporated into the phospholipids. These ratios are determined, at least in part, by regulation of membrane lipid biosynthesis-particularly that of cholesterol and oleate. It now appears that cholesterol and oleate biosynthesis are feedback regulated by a common transcriptional mechanism which is governed by the maturation of the SREBP transcription factors. In this article, we briefly review our current understanding of transcriptional regulation of plasma membrane lipid biosynthesis by sterols and oleate. We also discuss studies related to the mechanism by which the physical state of membrane lipids signals the transcriptional regulatory machinery to control the rates of synthesis of these structural components of the lipid bilayer. PMID- 10873554 TI - Activation of NADPH oxidase in Alzheimer's disease brains. AB - The present study is the first to show that superoxide (O(-)(2)) forming NADPH oxidase is activated in Alzheimer's disease (AD) brains by demonstrating the marked translocation of the cytosolic factors p47-phox and p67-phox to the membrane. In conjunction with a recent in vitro study showing that amyloid beta activates O(-)(2) forming NADPH oxidase in microglia, where these phox proteins are localized in this study, the present results suggest that, in AD, NADPH oxidase is activated in microglia, resulting in the formation of reactive oxygen species which can be toxic to neighboring neurons in AD. PMID- 10873555 TI - Alterations in outward K(+) currents on removal of external Ca(2+) in human atrial myocytes. AB - External divalent cations are known to play an important role in the function of voltage-gated ion channels. The purpose of this study was to examine the sensitivity of the voltage-gated K(+) currents of human atrial myocytes to external Ca(2+) ions. Myocytes were isolated by collagenase digestion of atrial appendages taken from patients undergoing coronary artery-bypass surgery. Currents were recorded from single isolated myocytes at 37 degrees C using the whole-cell patch-clamp technique. With 0.5 mM external Ca(2+), voltage pulses positive to -20 mV (holding potential = -60 mV) activated outward currents which very rapidly reached a peak (I(peak)) and subsequently inactivated (tau = 7.5 +/- 0.7 msec at +60 mV) to a sustained level, demonstrating the contribution of both rapidly inactivating transient (I(to1)) and non-inactivating sustained (I(so)) outward currents. The I(to1) component of I(peak), but not I(so), showed voltage dependent inactivation using 100 msec prepulses (V(1/2) = -35.2 +/- 0.5 mV). The K(+) channel blocker, 4-aminopyridine (4-AP, 2 mM), inhibited I(to1) by approximately 76% and reduced I(so) by approximately 33%. Removal of external Ca(2+) had several effects: (i) I(peak) was reduced in a manner consistent with an approximately 13 mV shift to negative voltages in the voltage-dependent inactivation of I(to1). (ii) I(so) was increased over the entire voltage range and this was associated with an increase in a non-inactivating 4-AP-sensitive current. (iii) In 79% cells (11/14), a slowly inactivating component was revealed such that the time-dependent inactivation was described by a double exponential time course (tau(1) = 7.0 +/- 0.7, tau(2) = 90 +/- 21 msec at +60 mV) with no effect on the fast time constant. Removal of external Ca(2+) was associated with an additional component to the voltage-dependent inactivation of I(peak) and I(so) (V(1/2) = -20.5 +/- 1.5 mV). The slowly inactivating component was seen only in the absence of external Ca(2+) ions and was insensitive to 4-AP (2 mM). Experiments with Cs(+)-rich pipette solutions suggested that the Ca(2+)-sensitive currents were carried predominantly by K(+) ions. External Ca(2+) ions are important to voltage-gated K(+) channel function in human atrial myocytes and removal of external Ca(2+) ions affects I(to1) and 4-AP-sensitive I(so) in distinct ways. PMID- 10873556 TI - Mitochondrial inhibitors evoke catecholamine release from pheochromocytoma cells. AB - Quantal catecholamine secretion evoked from individual pheochromocytoma (PC12) cells by exposure to mitochondrial inhibitors and uncouplers was monitored in real time using amperometry. Cyanide (0.05-5 mM) caused a concentration-dependent increase in the frequency of amperometric events. This secretory response was abolished by removal of extracellular Ca(2+) and by the application of Cd(2+) (200 microM), a nonselective blocker of voltage-gated Ca(2+) channels. Secretion was also inhibited by ca. 75% following pretreatment of cells with omega conotoxin GVIA to inhibit N-type Ca(2+) channels selectively. Secretion was also detected when cells were exposed to rotenone (10 microM), dinitrophenol (250 microM) and p-trifluoromethoxyphenyl hydrazone (1 microM) and, as for cyanide, these secretory responses were abolished by removal of extracellular Ca(2+) or application of 200 microM Cd(2+). These results indicate that, like hypoxia, mitochondrial inhibitors and uncouplers evoke catecholamine secretion from PC12 cells which is wholly dependent on Ca(2+) influx through voltage-gated Ca(2+) channels. PMID- 10873557 TI - S-nitrosothiol formation in blood of lipopolysaccharide-treated rats. AB - The administration of the gram-negative bacterial cell wall component lipopolysaccharide (LPS) to experimental animals results in the dramatic up regulation of the inducible form of nitric oxide synthase (iNOS). The resulting sustained overproduction of nitric oxide (NO) is thought to contribute to the septic shock-like state in these animals. Numerous studies have characterized the kinetics and magnitude of expression of iNOS as well as the production of NO derived nitrite and nitrate. However, little is known regarding the ability of iNOS-derived NO to interact with physiological substrates such as thiols to yield biologically active S-nitrosothiols during endotoxemia. It has been hypothesized that these relatively stable, vaso-active compounds may serve as a storage system for NO and they may thus play an important role in the pathophysiology associated with endotoxemia. In the present study, we demonstrate that 5 h after i.p. administration of LPS in rats, circulating S-nitrosoalbumin was increased by approximately 3. 4-fold over control. S-nitrosohemoglobin was increased by approximately 25-fold over controls and by threefold over S-nitrosoalbumin. No increase in low molecular weight S-nitrosothiols (i.e., S-nitrosoglutathione and S-nitrosocysteine) could be detected under our experimental conditions. Taken together these data demonstrate that endotoxemia dramatically enhances circulating S-nitrosothiol formation. PMID- 10873558 TI - Identification, gene structure, and expression of human frizzled-3 (FZD3). AB - We report the identification, genomic structure, chromosomal localization, and expression analysis of human frizzled-3 (FZD3), a 7-transmembrane receptor belonging to the frizzled family. The cDNA obtained from adult human brain shows 91% identity at the nucleotide level and 98% at the amino acid level to mouse frizzled-3 (fzd3). The FZD3 locus is located on chromosome 8p21, spans 48 Kb and its coding sequence is distributed in 6 exons intercalated by 5 introns. FZD3 is expressed in all analyzed human tissues, with quantitatively higher expression in the CNS and in urogenital structures. PMID- 10873559 TI - Phosphoprotein phosphatases regulate steroidogenesis by influencing StAR gene transcription. AB - The rate-limiting step in steroidogenesis is the transport of cholesterol into the mitochondria, and this is controlled by the steroidogenic acute regulatory (StAR) protein. We have previously shown that inhibition of phosphoprotein phosphatase 1 and 2A (PP1/2A) activities with the PP1/2A inhibitor calyculin A selectively reduces StAR protein expression and thus inhibits the synthesis of steroid hormones. The aim of this study was to determine whether this inhibition of StAR protein expression occurs at the level of transcription of StAR mRNA. We have used a competitive reverse transcription-polymerase chain reaction (RT-PCR) technique to determine whether inhibition of PP1/2A activities has any effect on the levels of StAR mRNA. Exposure of Y1 cells to forskolin significantly increased the expression of StAR mRNA and this forskolin-induced increase was reduced after exposure to Cal A at levels similar to those seen in the controls. These results suggest that cyclic AMP-induced increases in StAR mRNA levels are dependent upon phosphoprotein phosphatase activities. PMID- 10873560 TI - Analysis of tamoxifen-DNA adducts by high-performance liquid chromatography using postcolumn online photochemical activation. AB - Tamoxifen, a widely used nonsteroidal antiestrogen in the treatment of breast cancer, forms several metabolites. 4-Hydroxytamoxifen (4-OHTam), a metabolite found in the bloodstream, has much higher affinity for the estrogen receptor than tamoxifen itself. Oxidative activation of 4-OHTam induces DNA damage. DNA isolated from HL-60 cells exposed to 10 microM 4-OHTam in the presence of 1 microM hydrogen peroxide was digested enzymatically to release both normal and modified nucleosides. The modified nucleosides were enriched by butanol extraction. Using UV detection, HPLC analysis of the butanol extract from 200 microg DNA digest detected approximately 4 4-OHTam-dG adducts per 10(7) nucleotides (n = 3). Online postcolumn UV irradiation in HPLC and fluorescence detection improved the detection sensitivity by 3 x 10(2) times. Using 4-OHTam as an example, this report demonstrated for the first time the power of the technique to assay tamoxifen-DNA adducts directly in the DNA digest without relying on postlabeling. PMID- 10873561 TI - Bile acids reduce the apoptosis-inducing effects of sodium butyrate on human colon adenoma (AA/C1) cells: implications for colon carcinogenesis. AB - Butyrate is produced in the colon by fermentation of dietary fibre and induces apoptosis in colon adenoma and cancer cell lines, which may contribute to the protective effect of a high fibre diet against colorectal cancer (CRC). However, butyrate is present in the colon together with unconjugated bile acids, which are tumour promoters in the colon. We show here that bile acids deoxycholate (DCA) and chenodeoxycholate (CDCA), at levels present in the colon, gave a modest increase in cell proliferation and decreased spontaneous apoptosis in AA/C1 adenoma cells. Bile acids significantly inhibited the induction of apoptosis by butyrate in AA/C1 cells. However, the survival-inducing effects of bile acids on AA/C1 cells could be overcome by increasing the concentration of sodium butyrate. These results suggest that dysregulation of apoptosis in colonic epithelial cells by dietary factors is a key factor in the pathophysiology of CRC. PMID- 10873562 TI - Artifacts in cell culture: rapid generation of hydrogen peroxide on addition of ( )-epigallocatechin, (-)-epigallocatechin gallate, (+)-catechin, and quercetin to commonly used cell culture media. AB - There is considerable current interest in the possible beneficial health effects of quercetin, catechins, epigallocatechins, epigallocatechin gallates, and related phenolic compounds found in teas, wines, and other plant products. As a result, many laboratories are studying the effects of these compounds on cells in culture. The present paper shows that addition of these compounds to commonly used cell culture media leads to generation of substantial amounts of hydrogen peroxide (H(2)O(2)). Dulbecco's modified Eagle medium gives the highest H(2)O(2) level for all the compounds tested, with levels reaching >400 microM within 2 h for addition of 1 mM concentrations of gallic acid, epigallocatechin gallate, and epigallocatechin. Catechin and quercetin produced lower, but still significant, levels of H(2)O(2). McCoy's 5A and RPMI 1640 media also promoted H(2)O(2) production from the above phenolic compounds. This rapid generation of H(2)O(2) could account for some or all of the reported effects of phenolic compounds on cells in culture. PMID- 10873563 TI - Downregulation of proapoptotic proteins Bax and Bcl-X(S) in p53 overexpressing hepatocellular carcinomas. AB - As the occurrence of structural p53 mutations in hepatocellular carcinoma (HCC) in Thailand was previously reported to be much lower than that found in other high-incidence HCC areas, we analyzed 16 HCC samples from Thailand to determine the expression and functionality of p53 protein. We observed the overexpression of p53 protein in 69% of HCC, despite the prevalence of the wild-type p53 gene. However, the overexpressed p53 protein was nonfunctional as suggested by its inability to modulate the expressions of several p53 effector proteins (p21 and Bcl-2 family proteins). In addition, we observed significant underexpression of two proapoptotic proteins, Bax and Bcl-X(S), in 81% (P = 0.02) and 64% (P = 0.03) of HCC, respectively. Consequently, the ratios of proapoptotic to antiapoptotic BCL-2 family proteins were reduced in 88% of the HCC tumor tissues when compared to normal tissues, such that the rheostat between BCL-2 family proteins is strongly skewed toward enhanced cell survival in the tumor cells. PMID- 10873564 TI - Inhibition of alanyl-aminopeptidase suppresses the activation-dependent induction of glycogen synthase kinase-3beta (GSK-3beta) in human T cells. AB - Inhibition of alanyl-aminopeptidase (APN, CD13) gene expression or enzymatic activity compromises T cell proliferation and function. Molecular mechanisms mediating these effects are not known as yet. Recently, we found the expression of the proto-oncogen Wnt-5a to be strongly affected by APN-inhibition. Wnt-5a and other members of the Wnt family of secreted factors are implicated in cell growth and differentiation. Here, we analyzed by quantitative RT-PCR and immunoblotting the expression in mitogen-activated T cells of a major constituent of the Wnt-5a pathway, glycogen synthase kinase-3beta (GSK-3beta). T cell activation by phytohaemagglutinin or pokeweed mitogen results in a strong increase of GSK-3beta mRNA amounts. At the protein level, we observed an up-regulation of both GSK 3beta and phosphorylated GSK-3beta. This induction-dependent increase of GSK 3beta is markedly reduced in response to inhibitors of alanyl-aminopeptidase, actinonin, leuhistin, and RB3014. These findings may provide a rational for the growth inhibition resulting from a diminished expression or activity of alanyl aminopeptidase. PMID- 10873565 TI - Shear stress enhances glutathione peroxidase expression in endothelial cells. AB - Hemodynamic forces have profound effects on vasculature. Laminar shear stress upregulates superoxide dismutase (SOD) expression in endothelial cells. SOD converts superoxide anion to H(2)O(2), which, however, promotes atherosclerosis. Therefore, defense against H(2)O(2) may be crucial in reducing oxidative stress. Since glutathione peroxidase (GPx-1) reduces H(2)O(2) to H(2)O, the regulation of GPx-1 expression by mechanical stress was examined. Cultured bovine aortic endothelial cells (BAECs) were subjected to laminar shear stress and stretch force. Shear stress upregulated GPx-1 mRNA expression in a time- and force dependent manner in BAECs, whereas stretch force was without effect. Furthermore, shear stress increased GPx activity. L-NAME, an inhibitor of nitric oxide synthase, did not affect shear stress-induced GPx-1 mRNA expression. The ability of laminar shear stress to induce GPx-1 expression in endothelial cells may be an important mechanism whereby shear stress protects vascular cells against oxidative stress. PMID- 10873566 TI - Lipid peroxidation in small and large phospholipid unilamellar vesicles induced by water-soluble free radical sources. AB - The susceptibility of small and large egg yolk phosphatidylcholine unilamellar vesicles to Fe(2+)/histidine-Fe(3+)- and Fenton reagent (Fe(2+)-H(2)O(2))-induced lipid peroxidation was evaluated by measuring the formation of thiobarbituric acid reactive substances (TBARS). It has been found that surface curvature or phospholipid packing exerts significant effect on the oxidative susceptibility of the unsaturated lipid bilayers and the highly curved and loosely packed small unilamellar vesicles (SUVs) exhibit much less resistance to the oxidative stress induced by the water-soluble free radical sources. The presence of lipid hydroperoxides in sonicated vesicles was excluded as the cause for higher level of lipid peroxidation in the phospholipid SUVs. Instead, the experimental results can be explained by the difference in ability of the water-soluble oxidants to penetrate the two types of lipid membranes. This hypothesis is supported by data obtained from fluorescence lifetime and quenching studies. PMID- 10873568 TI - Characterization of novel and identified genes in guinea pig organ of corti. AB - A number of proteins are expressed in the organ of Corti and are considered to be responsible for hearing. However, most of them have not been identified. Therefore, to achieve a better understanding of the genetic factors influencing these traits, the first step is to characterize the genes expressed in the organ of Corti. In the present study, a cDNA library was constructed from the guinea pig organ of Corti. After sequencing isolated clones, 196 expressed sequence tags (ESTs) were identified with FASTA analysis: 65 ESTs showed significant sequence homology to previously identified genes in guinea pig, human or other species, and 131 ESTs showed no significant matches to sequences already present in the DNA database DDBJ/GenBank/EMBL. A variety of matching sequences, some of which were known to be cochlea-specific, were found through FASTA analysis of the 65 clones. RT-PCR with a panel of 10 different tissue mRNA revealed the restricted expression of 13 unknown clones. The results of our analysis allowed the establishment of a list of genes expressed in the guinea pig organ of Corti. PMID- 10873567 TI - Mitogenic phospholipase D activity is restricted to caveolin-enriched membrane microdomains. AB - Phospholipase D (PLD) activity is elevated in response to the oncogenic stimulus of several signaling oncogenes. PLD activity is also elevated in response to peptide growth factors, indicating that PLD likely plays an important role in mitogenic signaling. Many proteins that mediate mitogenic signaling are localized in caveolin-enriched membrane microdomains (CEMMs). We report here that the elevated PLD activity in NIH 3T3 cells transformed by activated oncogenic forms of Src, Ras, and Raf is largely restricted to the CEMMs. Likewise, the PLD activity stimulated by epidermal growth factor is also restricted to the CEMMs. Although both PLD1 and PLD2 were found in CEMMs, neither was particularly enriched in the CEMMs of the transformed relative to the parental cells, indicating that it is the specific activity of PLD that is increased in the CEMMs. An apparent PLD substrate specificity in transformed cells for phosphatidylcholine lacking arachidonate acyl groups is also explained by the localization of activity in the CEMMs where [(3)H]arachidonate-labeled PC was excluded. These data indicate that mitogenic signals through PLD are initiated in CEMMs where many signaling molecules colocalize. PMID- 10873569 TI - Characterization of five novel human genes in the 11q13-q22 region. AB - The redundancy of sequences in dbEST has approached a level where contiguous cDNA sequences of genes can be assembled, without the need to physically handle the clones from which the ESTs are derived. This is termed EST based in silico gene cloning. With the availability of sequence chromatogram files for a subset of ESTs, the quality of EST sequences can be ascertained accurately and used in contig assembly. In this report, we performed a study using this approach and isolated five novel human genes, C11orf1-C11orf5, in the 11q13-q22 region. The full open reading frames of these genes were determined by comparison with their orthologs, of which four mouse orthologs were isolated (c11orf1, c11orf2, c11orf3 and c11orf5). These genes were then analyzed using several proteomics tools. Both C11orf1 and C11orf2 are nuclear proteins with no other distinguishing features. C11orf3 is a cytoplasmic protein containing an ATP/GTP binding site, a signal peptide located in the N-terminus and a similarity to the C. elegans protein "Probable ARP 2/3 complex 20kD subunit." C11orf4 is a peptide which displays four putative transmembrane domains and is predicted to have a cytoplasmic localization. It contains signal peptides at the N- and C-termini. C11orf5 is a putative nuclear protein displaying a central coiled coil domain. Here, we propose that this purely EST-based cloning approach can be used by modestly sized laboratories to rapidly and accurately characterize and map a significant number of human genes without the need of further sequencing. PMID- 10873570 TI - 1,25 dihydroxyvitamin D(3) activates sphingomyelin turnover in ROS17/2.8 osteosarcoma cells without sphingolipid-induced changes in cytosolic Ca(2+). AB - 1,25-Dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] initiates the hydrolysis of sphingomyelin in ROS 17/2.8 osteosarcoma cells with the resultant generation of cell-associated ceramide. Increases in ceramide levels were detectable at 15 min and maximal one hour after exposure of cells to 1,25(OH)(2)D(3). Neither 1,25(OH)(2)D(3) nor exogenous ceramide elicited a change in cytosolic free Ca(2+) ([Ca(2+)](i)). Transient elevations in [Ca(2+)](i) were observed when cells were exposed to exogenous sphingosine, but there was no detectable conversion of ceramide to sphingosine in 1, 25(OH)(2)D(3)-treated cells. Ceramide also did not stimulate Ca(2+) uptake across ROS 17/2.8 cell plasma membranes. Collectively, these results suggest that 1,25(OH)(2)D(3) activates sphingomyelin turnover in ROS 17/2.8 osteosarcoma cells but that the sphingolipid metabolite ceramide is not responsible for 1,25(OH)(2)D(3)-induced activation of plasma membrane Ca(2+) channels. PMID- 10873571 TI - Inhibition of Fas-mediated fulminant hepatitis in CrmA gene-transfected mice. AB - Hyperimmune response via Fas/Fas-ligand and perforin/granzyme pathways may be essential in pathogenesis of virus-induced fulminant hepatitis. CrmA inhibits activation of caspases and granzyme B, suggesting it may block these pathways. We investigated whether CrmA expression would inhibit Fas-associated lethal hepatitis in mice. We successfully generated AxCALNLCrmA, a recombinant adenovirus expressing CrmA gene with a Cre-mediated switching cassette. We increased CrmA expression level in the liver transfected with AxCALNLCrmA (10(9) pfu) by increasing administration dose (10(7)-10(9) pfu) of AxCANCre, a recombinant, adenovirus-expressing Cre gene. Injection of anti-Fas antibody into the control mice rapidly led to animal death due to massive liver apoptosis, while the apoptosis was dramatically reduced in the CrmA-expressed mice. The animal survival increased with an increase of CrmA expression. The formation of active caspase-3 was markedly inhibited in the crmA-transfected hepatocytes in vitro. These results suggest that crmA is an effective gene that can inhibit immune-related liver apoptosis. PMID- 10873572 TI - Identification of calponin as a novel substrate of Rho-kinase. AB - Calponin, an F-actin-associated protein implicated in the regulation of smooth muscle contraction, is known to be phosphorylated in vitro by protein kinase C (PKC) and Ca(2+)/calmodulin dependent protein kinase II (CaM kinase II). Unphosphorylated calponin binds to F-actin and inhibits the actin-activated myosin ATPase activity; these properties are lost on phosphorylation. In the present study, we found that Rho-kinase phosphorylated basic calponin stoichiometrically in vitro. We identified the sites of phosphorylation of calponin by Rho-kinase as Thr-170, Ser-175, Thr-180, Thr-184, and Thr-259, and prepared antibodies that specifically recognized calponin phosphorylated at Thr 170 and Thr-184. We showed that the phosphorylation of calponin by Rho-kinase inhibited the binding of calponin to F-actin. Taken together, these results suggest that calponin is a substrate of Rho-kinase and that Rho-kinase regulates the interaction of calponin with F-actin. PMID- 10873573 TI - Comparative analysis of CD137 and LPS effects on monocyte activation, survival, and proliferation. AB - CD137 (ILA/4-1BB), a member of the TNF receptor family, regulates activation, survival and proliferation of primary human monocytes. Here we compare the activities of lipopolysaccharide (LPS), a classical and potent monocyte activator to that of CD137. LPS is a more potent activator of monocytes, as evidenced by a stronger induction of the proinflammatory cytokine IL-8. However, CD137 could further increase maximal cytokine induction by LPS, which points to separate signaling pathways for LPS and CD137. Also, expression of myc was only induced by the combination of CD137 and LPS. Expression of macrophage colony-stimulating factor is induced more potently by CD137, but an additive effect is obtained by the combination of CD137 and LPS. Monocyte/macrophage survival and proliferation is only induced by CD137. LPS counteracts both activities of CD137 via activation induced cell death. While LPS has a role in activation of monocytes in innate immunity, the CD137 receptor/ligand system seems to deliver an activating signal to monocyte in acquired immunity. PMID- 10873575 TI - Differential contribution of superoxide dismutase activity by prion protein in vivo. AB - Normal prion protein (PrP(C)) is a copper binding protein and may play a role in cellular resistance to oxidative stress. Recently, copper-bound recombinant PrP(C) has been shown to exhibit superoxide dismutase (SOD)-like activity. However, as PrP(C) affinity for copper is low in comparison to other cupro proteins, the question remains as to whether PrP(C) could contribute SOD activity in vivo. To unravel this enigma, we compared the SOD activity in lysates extracted from different regions of the brain from wild-type mice before and after the depletion of PrP(C). We found that removal of PrP(C) from the brain lysates reduced the levels of total SOD activity. The level of contribution to the total SOD activity was correlated to the level of PrP expressed and to the predominant form of PrP present in the specific brain region. Collectively, these results provide strong evidence that PrP(C) differentially contributes to the total SOD activity in vivo. PMID- 10873576 TI - NF-kappaB activation is related to the resistance of lung cancer cells to TNF alpha-induced apoptosis. AB - In diverse cell types, NF-kappaB transcription factors have been shown to have a role in regulating the apoptotic program, either as essential for the induction of apoptosis or, perhaps more commonly, as blockers of apoptosis. We investigated the role of NF-kappaB activation in the TNF-alpha-mediated apoptosis in lung cancer cells. TNF-alpha-resistant NCI-H157 cells became sensitized to TNF-alpha by prior treatment with cycloheximide, suggesting the presence of newly synthesized antiapoptotic protein(s). We next evaluated whether the transcription of antiapoptotic protein(s) depends on the activation of NF-kappaB. NF-kappaB activation was blocked by either adenovirus-mediated overexpression of IkappaBalpha superrepressor or pretreatment with proteasome inhibitor, MG132. Both methods of blocking NF-kappaB activation enhanced TNF-alpha-induced apoptosis in NCI-H157 cells. These results suggest that NF-kappaB activation confers resistance to TNF-alpha-mediated apoptosis in lung cancer cells. PMID- 10873574 TI - Inhibition of signal transducer and activator transcription factor 3 in rats with acute hepatic failure. AB - In fulminant hepatic failure, survival is not possible without recovery of sufficient hepatocyte mass. Remarkably, only a few studies exist that provide insight into the mechanisms that control proliferation of residual hepatocytes after extensive hepatocyte loss. In this regard, the role of growth-regulatory factors, including pro-inflammatory cytokines such as interleukin-6 (IL-6), is not well understood. In the present study we show that in rats with critically low (10%) hepatocyte mass, whether with or without ongoing liver cell necrosis, inhibition of liver regeneration is associated with early and sustained increase in blood IL-6 levels. Under these conditions, the signal transducer and activator of transcription (Stat3) DNA binding activity was lowered at the time of G1/S cell-cycle transition. We further demonstrate that the protein inhibitor of activated Stat3 (PIAS3) and the suppressor of cytokine signaling (SOCS-1) were up regulated early after induction of liver failure (6-12 h). In vitro, IL-6 induced PIAS3 expression in HGF stimulated rat hepatocytes. These findings suggest that after massive hepatocyte loss, an early and rapid rise in blood IL-6 levels may weaken the hepatic regenerative response through up-regulation of Stat3 inhibitors PIAS3 and SOCS-1. PMID- 10873577 TI - CEA is the major PHA-L-reactive glycoprotein in colon carcinoma cell lines and tumors: relationship between K-ras activation and beta1-6 branching of N-linked carbohydrate on CEA. AB - Previously we have shown that a positive correlation existed between the presence of beta1-6 branching of N-linked carbohydrate (detected as PHA-L reactivity) and the level of Ras activation in colon carcinoma cell lines. In these cell lines the major PHA-L-reactive species was found to be 180 kDa. Here we identified this species to be carcinoembryonic antigen (CEA) by demonstrating that: (a) CEA immunoreactivity and PHA-L reactivity colocalized on blots of crude cellular membranes from these cell lines, and that (b) immunoprecipitation of CEA resulted in quantitative coprecipitation of PHA-L reactivity at 180 kDa. Metabolic labeling of cell line HTB39 with [(3)H]mannose revealed that CEA was the predominantly labeled glycoprotein. This indicated that CEA was the major PHA-L reactive species due its high level of expression. The amount of PHA-L reactivity present on CEA, expressed as the PHA-L/CEA ratio, was found to vary between cell lines. This ratio was found to correlate closely with the level of Ras activation in these cells. In cellular membrane isolated from primary colon carcinoma, the major PHA-L-reactive species was also 180 kDa. This reactivity colocalized with CEA immunoreactivity, indicating that the major beta1-6-branching glycoprotein in membranes from primary colon carcinoma was CEA. Similar to that seen in cell lines, the amount of PHA-L reactivity on CEA in human tumor samples varied, suggesting that a similar paradigm of Ras-induced expression of beta1-6 branching may occur in human colon carcinoma. PMID- 10873578 TI - TSH and cAMP do not signal mitogenesis through Ras activation. AB - Ras activation by receptor tyrosine kinases or serpentine receptors is generally considered to be essential for G1 phase progression and mitogenesis. In the physiologically relevant model of primary dog thyrocytes, the accumulation of the GTP-bound form of Ras constituted an early convergence point of various mitogenic or comitogenic stimuli including EGF, HGF, phorbol esters, insulin and carbachol. By contrast, the basal level of GTP-Ras was slightly reduced by TSH and forskolin and did not increase during the TSH/cAMP-dependent progression into G1 phase. This rules out a role for the activation of Ras as a signal in the mitogenesis elicited by TSH via cAMP in these cells. PMID- 10873579 TI - Glycosylation affects translocation of integrin, Src, and caveolin into or out of GEM. AB - Endogenous GM3 synthesis and full N-glycosylation in membrane receptors occurred in "4-epimerase-less" ldlD (Krieger's CHO mutant) cells cultured in Gal containing medium, whereby components of detergent-insoluble, low-density, buoyant membrane fraction, termed "glycolipid-enriched microdomain (GEM)," varied significantly by translocation into or out of GEM. Integrins alpha3 and alpha5 were translocated into GEM in the presence of 0.5 or 0.25% Triton X-100, particularly in the absence of Gal, whereby integrins are underglycosylated and GlcCer is the major glycolipid component in GEM. Src family kinase was translocated into and enriched in GEM fractions when prepared in 0.5 or 0.25% Triton X-100 from cells grown in Gal-containing medium, whereby GM3 synthesis is induced. In contrast, caveolin is highly enriched in GEM when GM3 synthesis does not occur, and is translocated into high-density membrane fraction when GM3 synthesis occurs. The results suggest that levels of key molecules controlling cell adhesion and signaling are defined by translocation into or out of GEM, which depends on glycosylation state. PMID- 10873580 TI - Cytokine-inducing macromolecular glycolipids from Enterococcus hirae: improved method for separation and analysis of its effects on cellular activation. AB - Previously, we showed that several minor macromolecular glycolipids accounting for less than 5% of the lipoteichoic acid (LTA) fraction from Enterococcus hirae ATCC 9790 possess cytokine-inducing activity, whereas the purified LTA does not. In other words, the immunobiological activity of the LTA fraction reported in the 1980s was not attributable to LTA itself, but to other glycolipids coexisting in the fraction. In the present study, we improved the procedure of separation of the active glycolipids and evaluated their effects on cellular activation. The immunobiologically active glycolipids were separated from the crude glycolipid fraction obtained by hot phenol-water extraction of the cells. The total yield of active glycolipids was about fivefold higher than that separated by the previous method. Interleukin-6-inducing activities of the active glycolipids from 1,25 dihydroxy vitamin D(3)-differentiated human monocytic leukemia cells, THP-1, were inhibited by anti-CD14 mAbs in a dose-dependent manner. Macrophages from Toll like receptor (TLR)-2-deficient or -4-deficient mice completely lacked the ability to produce tumor necrosis factor-alpha on stimulation with active glycolipids. These observations indicated that the cellular activation by the active glycolipids from E. hirae is mediated by CD14 and by both TLR2 and TLR4. PMID- 10873581 TI - Possible role of calpain in normal processing of beta-amyloid precursor protein in human platelets. AB - Abnormal proteolytic processing of beta-amyloid precursor protein (APP) underlies the formation of amyloid plaques in aging and Alzheimer's disease. The proteases involved in the process have not been identified. Here we found that spontaneous proteolysis of intact APP in detergent-lysed human platelets generated a N terminal fragment that was immunologically indistinguishable from secreted APP, reminiscent of the action of a putative alpha-secretase. This proteolysis of APP was inhibited by EDTA, suggesting that a metal-dependent protease was involved. Among the several metals tested, calcium was the only one that enhanced APP proteolysis and the reaction was blocked by EGTA as well as by several calpain inhibitors. The APP fragments generated by spontaneous proteolysis in platelet lysates were identical to those produced by exposure of partially purified APP to exogenous calpain. Finally, the secretion of APP from intact platelets was inhibited by cell-permeable calpain inhibitors. Taken together, these results suggest that normal processing of APP in human platelets is mediated by a calcium dependent protease that exhibits calpain-like properties. PMID- 10873582 TI - Characterization of Cop I coat proteins in plant cells. AB - Membrane traffic in eukaryotic cells is mediated by COP (coat protein)-coated vesicles. Their existence in plant cells has not yet been unequivocally demonstrated, although coated vesicles (probably with a COP coat) can be seen by electron microscopy. At the gene level, plant cells seem to contain all the components necessary to form COP-coated vesicles. In this paper, we have used antibodies raised against mammalian COPI coat proteins to detect putative homologues in rice (Oryza sativa) cells. Using these antibodies, we have found that rice cells contain alpha-, beta-, beta'-, and gamma-COP, as well as ADP ribosylation factor (ARF) 1 protein. In addition, we show that antibodies against mammalian beta'-COP can immunoprecipitate not only beta'-COP but also alpha-, beta-, and gamma-COP, suggesting that COPI components in rice cells exist as a complex (or coatomer) in the cytosol, as in mammalian cells. Finally, we show that COP binding to membranes is GTP-dependent, and that ARF1 also binds to membranes in a GTP-dependent manner. PMID- 10873583 TI - The 300-kDa intermediate filament-associated protein (IFAP300) is a hamster plectin ortholog. AB - Plectin is a high-molecular-weight cytoskeleton-associated protein that was initially identified in intermediate filament (IF)-enriched fractions of rat C6 glioma cells. At the cellular level, plectin has been found to associate with IF networks and IF-associated structures that are involved in cell-cell and cell substrate adhesions. IFAP300 is an IF-associated protein that was initially identified in hamster cells by a monoclonal antibody directed against a high molecular weight protein present in IF-enriched cytoskeletal preparations. Plectin and IFAP300 display similar distribution patterns within cells as determined by immunofluorescence. Based upon this and the finding that their biochemical properties are similar, it has been suggested that they may actually be orthologous proteins. In this paper we demonstrate that this is the case. Cloning and sequencing of most of the hamster plectin cDNA demonstrates that plectin is found in hamster cells and that its sequence is highly conserved between species. Using immunological cross-reactivity, epitope mapping, and immunoelectron microscopy, we show that IFAP300 is actually the hamster ortholog of plectin. PMID- 10873584 TI - PC2 and 7B2 null mice demonstrate that PC2 is essential for normal pro-CCK processing. AB - Analysis of CCK content in extracts of whole forebrain from PC2 and 7B2 null mouse brain showed a significant decrease relative to wild-type brains. More detailed analysis revealed that CCK 8 amide levels in cerebral cortex and forebrain regions were more decreased than in hypothalamus. CCK 8 content in PC2 null mouse intestines was identical to control. Null mutant brains contained less CCK 8 than wild type and no other forms were seen when analyzed by gel filtration chromatography. No brain area examined was completely devoid of CCK, suggesting that other enzymes can partially compensate for the loss of PC2. This is the first demonstration that any endoprotease is important for CCK processing but also suggest the presence of a redundant system to ensure production of active CCK in the brain. PMID- 10873585 TI - Deficiency in mitochondrial aldehyde dehydrogenase increases the risk for late onset Alzheimer's disease in the Japanese population. AB - Mitochondrial aldehyde dehydrogenase 2 (ALDH2) deficiency is caused by a mutant allele in the Mongoloids. To examine whether genetic constitutions affecting aldehyde metabolism influence the risk for late-onset Alzheimer's disease (LOAD), we performed a case-control study in the Japanese population on the deficiency in ALDH2 caused by the dominant-negative mutant allele of the ALDH2 gene (ALDH2*2). In a comparison of 447 patients with sex, age, and region matched nondemented controls, the genotype frequency carrying the ALDH2*2 allele was significantly higher in the patients than in the controls (48.1% vs 37.4%, P = 0.001). Logistic regression analysis indicates that carriage of the ALDH2*2 allele is an independent risk for LOAD of the epsilon4 allele of the apolipoprotein E gene (APOE-epsilon4) (P = 0.002). Moreover, the odds ratio for LOAD in carriers of the ALDH2*2 allele was almost twice that in noncarriers, irrespective of status with regard to the APOE-epsilon4 allele. Among patients homozygous for the APOE epsilon4 allele, age at onset of LOAD was significantly lower in those with than without the ALDH2*2 allele. In addition, dosage of the ALDH2*2 allele significantly affected age at onset of patients homozygous for the APOE-epsilon4 allele. These results indicate that the ALDH2 deficiency is a risk for LOAD, synergistically acting with the APOE-epsilon4 allele. PMID- 10873586 TI - Functional and structural studies of alpha-crystallin from galactosemic rat lenses. AB - Chaperone-like activity and structural changes of lens alpha-crystallin from rats fed with galactose at various time intervals have been studied using high performance liquid chromatograph (HPLC), circular dichroism (CD), and 1 anilinonaphthalene-8-sulfonic acid (ANS) fluorescence emission. It was found that chaperone-like activity of alpha-crystallin from galactose-fed rats toward dithiothreitol (DTT)-induced insulin B aggregation started to decrease after 3 weeks and decreased significantly after 5 weeks. Consistent results were observed in lens morphology, and lens opacity slightly developed after 3 weeks and became obvious after 5 weeks. HPLC analysis for chaperone function showed that the formation of high molecular weight aggregates (HMWA) of alpha-/gamma-crystallins decreases with the increase of galactose-feeding time, revealing that chaperone like activity is concomitant with the formation of HMWA. Circular dichroism results showed the reduction of beta-sheet structure and loss of microenvironment of aromatic-type amino acids for opaque lenses, indicating alpha-crystallin's secondary and tertiary structure changed with the development of the lens opacity. ANS binding site estimated by Klotz equation showed it is 1.5 times higher at room temperature and is 2.4 times higher at 58 degrees C for age matched normal alpha-crystallin than for 5-week galactose-fed lens alpha crystallin, indicating opaque lens alpha-crystallin loses the ability to assemble into an appropriately placed hydrophobic regions. The overall results accordingly indicated that galactose-induced cataractous alpha-crystallin has disordered structure, leading to the loss of its chaperone-like activity. PMID- 10873587 TI - The frequency of point mutations in mitochondrial DNA is elevated in the Alzheimer's brain. AB - Using a PCR-based strategy, we found that point mutation frequencies in mitochondrial DNA (mtDNA) were 2- to 3-fold higher in the parietal gyrus, hippocampus, and cerebellum from subjects with Alzheimer's disease (AD) compared to normal controls. In contrast, levels of a commonly studied deletion mutation, mtDNA(4977), were not elevated in AD. The frequency of point mutations did not vary significantly among the three brain areas, whereas the frequency of mtDNA(4977) was 15- to 25-fold lower in the cerebellum in comparison to the cortex; this regional variation was seen in both the normal and Alzheimer's brain. In blood mtDNA, point mutation frequencies were not elevated in AD patients. The elevated frequency of point mutations in all three brain regions is consistent with the idea that increased oxidant stress is associated with AD. PMID- 10873588 TI - Dual effects of PKNalpha and protein kinase C on phosphorylation of tau protein by glycogen synthase kinase-3beta. AB - We analyzed the effects of PKNalpha and protein kinase C (PKC) on phosphorylation of tau protein by glycogen synthase kinase (GSK)-3beta using monoclonal antibodies (AT8, AT180, and AT270). These antibodies are highly specific for phosphorylated tau in Alzheimer paired helical filaments, and recognize phosphorylated Ser202/Thr205, Thr231, and Thr181 of tau protein, respectively. Immunoblot analysis demonstrated that PKNalpha and PKC did not directly phosphorylate their sites, whereas GSK-3beta efficiently did so. Incubating GSK 3beta with PKNalpha or PKC subtypes inhibited subsequent GSK-3beta-induced AT8 and AT270 immunoreactivity. However, the constitutive active form of the GSK 3beta(S9A) mutant was almost totally inert to each enzyme. Incubating tau with PKNalpha increased the GSK-3beta-induced AT180 immunoreactivity, which was further enhanced when the S9A mutant was used instead of the wild type GSK-3beta. These results suggest that PKNalpha and PKC directly inhibit GSK-3beta activity at least in part by phosphorylating Ser9 of GSK-3beta, and that they indirectly suppress GSK-3beta-stimulated phosphorylation of tau at amino acids Ser202/Thr205 and Thr181, but enhanced phosphorylation at Thr231 through phosphorylation at other sites of tau. PMID- 10873589 TI - Expression of autocrine motility factor/phosphohexose isomerase in Cos7 cells. AB - Autocrine motility factor (AMF) is identical to the glycolytic enzyme phosphohexose isomerase (PHI) and overexpression of AMF/PHI is associated with tumor malignancy. In order to study the overexpression of AMF/PHI, an HA-tagged AMF construct was transiently transfected into Cos7 cells. Expression of a tagged AMF-HA allowed us to determine that over a period of 16 hours only a small amount (0.1-1%) of total cellular AMF-HA was secreted into the cell medium. Cell associated AMF-HA was exclusively cytosolic as it could be completely extracted with Triton X-100 and concentrated within actin rich pseudopodial domains. Treatment of the cells with the glycolysis inhibitor oxamate disrupted the association of AMF-HA with actin concentrations demonstrating that glycolysis regulates the formation of these AMF/PHI-associated actin-rich protrusions. AMF/PHI is a well-characterized tumor cell secreted cytokine and we identify here an alternate intracellular function for this glycolytic enzyme/cytokine in cell motility. PMID- 10873590 TI - Specific impairment of cardiogenesis in mouse ES cells containing a human chromosome 21. AB - Down syndrome (DS) leads to cardiac defects which are common and significant in babies with DS. We recently generated chimeric mice carrying a human chromosome (hChr) 21. The contribution ratio of embryonic stem (ES) cells containing a hChr 21 was specifically low in the heart, compared to other organs, and cardiovascular malformations were observed, suggesting that an additional copy of hChr 21 also disrupts the normal development of heart in mice. Here we describe that the presence of hChr 21 in ES cells delays the appearance of beating cardiomyocyte during differentiation, whereas differentiation into other cell types is not disrupted. Furthermore, the defect in cardiogenesis was restored following the deletion of a specific region of hChr 21. Therefore, we conclude that the imbalance of specific gene(s) on hChr 21 may lead to the disturbance of cardiogenesis and that this may be a useful system to model and investigate the cardiac defects of human DS. PMID- 10873591 TI - Expression of guanylin is downregulated in mouse and human intestinal adenomas. AB - Guanylin is a pro-secretory hormone that is expressed in intestinal epithelia. Previously, we mapped the guanylin gene to mouse and human chromosomal regions containing multiple intestinal tumor-modifying loci. Here, we investigate whether guanylin expression is downregulated in precancerous human and mouse intestinal adenomas and whether diminished guanylin expression increases adenoma susceptibility in an animal model of intestinal cancer, the multiple intestinal neoplasia (Min) mouse. In situ hybridization analysis indicated diminished guanylin expression in both mouse and human adenomas. Northern analysis of mouse intestinal tissues showed strain-specific levels of guanylin expression but no correlation with the resistance or susceptibility of each strain to adenoma formation. Similarly, cDNA sequence analysis indicated no inactivating mutations or polymorphisms common to either the high or low adenoma-risk groups. Nonetheless, we have shown that significant loss of guanylin RNA in adenomas of mouse and human is a marker of intestinal epithelial cell transformation. PMID- 10873592 TI - The role of ARNT2 in tumor angiogenesis and the neural response to hypoxia. AB - The Hypoxia-Inducible Factor-1 (HIF-1) activates the transcription of many genes required for cellular and organismal responses to oxygen deprivation. The HIF-1 complex is composed of the ubiquitously expressed basic helix-loop-helix/PAS (bHLH/PAS) proteins HIF-1alpha and Arylhydrocarbon Receptor Nuclear Translocator (ARNT). ARNT2 is a conserved ARNT homolog that is highly expressed in neurons, suggesting that ARNT2/HIF-1alpha heterodimers mediate transcriptional responses to oxygen deprivation in the nervous system. We show here that ARNT2 forms functional HIF complexes in vivo, and that ARNT2 restores hypoxia-induced gene expression to ARNT-deficient ES cells and hepatocytes. Formation of neural ARNT2/HIF-1alpha complexes in Arnt(-/-) ES cell-derived teratocarcinomas may explain why these tumors express VEGF, vascularize and grow efficiently, in contrast to ARNT-deficient hepatomas. Interestingly, all neural cell types studied accumulate both ARNT- and ARNT2-containing HIF complexes. We conclude that ARNT2 forms functional HIF complexes in neurons and plays an integral role in hypoxic responses in the CNS. PMID- 10873593 TI - Transcription factor GATA-2 gene is located near 3q21 breakpoints in myeloid leukemia. AB - Rearrangements affecting chromosome band 3q21 are observed in a subgroup of patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). However, little is known about the molecular consequences of such aberrations. We therefore established a PAC contig in the 3q21 breakpoint region and identified potential protein coding sequences by exon trapping. One of the exons isolated was from the human GATA-2 gene, which we showed to be transcribed from telomere to centromere. The majority of 3q21 breakpoints are located telomeric to the transcribed portion of this gene in a region that in mice appears to be necessary for proper promoter function. Results of GATA-2 expression analyses in leukemic cell lines as well as primary patient samples are compatible with the hypothesis that 3q21 aberrations contribute to leukemogenesis through deregulation of the hematopoietic transcription factor GATA-2. PMID- 10873594 TI - Convulxin binding to platelet receptor GPVI: competition with collagen related peptides. AB - Convulxin (CVX), a potent platelet aggregating protein from the venom of the snake Crotalus durissus terrificus, is known to bind to the platelet collagen receptor, glycoprotein VI (GPVI). CVX binding to human platelets was investigated by flow cytometry, using fluorescein labeled convulxin (FITC-CVX). Scatchard analysis indicated high and low affinity binding sites with Kd values of 0.6 and 4 nM and Bmax values of 1200 and 2000 binding sites per platelet. FITC-CVX binding was inhibited by collagen related peptides (CRPs) comprising a repeated GPO sequence, namely GCO(GPO)(10)GCOGNH(2) and GKO(GPO)(10)GKOGNH(2), which also bind to receptor GPVI. These peptides (monomeric or cross-linked forms) gave a high affinity inhibition of 10-20% for concentrations between 10 ng/ml and 5 microg/ml, followed by a second phase of inhibition at concentrations greater than 5 microg/ml. It was shown also that the inhibition of FITC-CVX binding by CRPs was independent on the time of preincubation of platelets with CRPs, and the same percentage of inhibition was seen with various concentrations of convulxin. Confocal microscopy of the distribution of FITC-CVX binding sites on platelets showed an homogeneous distribution of FITC-CVX bound to GPVI, although some limited clustering may exist. PMID- 10873595 TI - Molecular identification and characterization of novel members of the human organic anion transporter (OATP) family. AB - We identified three novel transporters structurally belonging to the organic anion transporting polypeptide (OATP) family in humans. Since previously known rat oatp1 to 3 do not necessarily correspond to the human OATPs in terms of either tissue distribution or function, here we designate the newly identified human OATPs as OATP-B, -D and -E, and we rename the previously known human OATP as OATP-A. OATP-C proved to be identical with the recently reported LST1/OATP-2. Expression profiles of the five OATPs and the prostaglandin transporter PGT (a member of OATP family) in human tissues showed that OATP-C is exclusively localized in liver, OATP-A and PGT are expressed in restricted ranges of tissues, and OATP-B, -D and -E show broad expression profiles. OATP-B, -C, -D and -E exhibited transport activity for [(3)H]estrone-3-sulfate as a common substrate. OATP-C has a high transport activity with broad substrate specificity. PMID- 10873596 TI - Interaction of albumin mRNA with proteins from rat liver with CCl(4)-induced injury. AB - Acute phase responses to intragastric administration of a single dose of CCl(4) were examined with albumin mRNA turnover as a marker. Based on the combination of the changes in stability of albumin mRNA and activity of transcription of its gene, the entire course of liver injury was classified into three stages, the first stage for aggravation of injury until 9 h, the second from 9 to 24 h, and the third for repair of injury or regeneration of liver after 48 h. Liver S100 fractions from normal and CCl(4)-treated rats contained, in total, 11 polypeptides cross-linked with part of albumin mRNA, although they did not appear to be specific to this mRNA. Their profiles were altered together with the changes in stability of albumin mRNA in different stages. These findings suggest that the polypeptides with distinct properties play roles in physiologically significant processes involved in utilization and turnover of albumin mRNA, apparently altering its stability. PMID- 10873597 TI - Kaposi's sarcoma cells of different etiologic origins respond to HIV-Tat through the Flk-1/KDR (VEGFR-2): relevance in AIDS-KS pathology. AB - Kaposi's sarcoma (KS) is an hyperplastic lesion whose main histological features are typical spindle shaped cells with a mixed endothelial-mesenchymal-macrophage phenotype, an intense vascularization and an inflammatory infiltrate. The etiology of KS appears to be linked to activation of a latent HHV8 infection. Sporadic and iatrogenic KS are slow progressing lesions that can undergo spontaneous regression. In contrast, KS, which is frequently associated with HIV infection, is found in a highly aggressive form in AIDS patients. The HIV-1 Tat has been shown to activate the VEGF receptor KDR in endothelial and KS spindle cells, suggesting this HIV protein could contribute to KS pathogenesis. We used primary 'reactive' KS cell culture from sporadic and epidemic KS, and an immortal KS-line (KS-Imm) isolated in our laboratory from a iatrogenic KS lesion, to verify if Tat-induced cell signaling is able to mediate cellular responses. We demonstrate that KS cells migrated in response to Tat and that VEGF is able to compete with the Tat chemotactic activity towards these cells. A function blocking anti-KDR antibody was able to abrogate both VEGF and Tat-induced KS chemotactic response, indicating a direct involvement of this receptor. Our data show that HIV-Tat can also activate KS cells derived from sporadic or iatrogenic lesions, suggesting that in AIDS patients Tat could cooperate with VEGF in activation of KDS on KS precursor spindle and endothelial cells, and contribute to the aggressiveness of AIDS-KS lesions. PMID- 10873598 TI - Restructured transactivation domain in hamster AH receptor. AB - Hamsters and Han/Wistar (Kuopio; H/W) rats show peculiarly selective responsiveness to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). They are extremely resistant to its acute lethality but sensitive to, e.g. , enzyme induction. The biological effects of TCDD are mediated by the AH receptor (AHR). Recent studies on H/W rat AHR discovered a remodelled transactivation domain which appears to be critical for the TCDD resistance of these animals. Here, molecular cloning and sequencing of hamster AHR reveals another type of restructured transactivation domain. In hamsters, the functionally pivotal Q-rich region is substantially expanded and enriched in glutamine compared with all other AHRs cloned to date. By contrast, the amino-terminal end is highly conserved, which is in agreement with the H/W rat AHR. Because of the additional material in the transactivation domain, hamster AHR protein is larger than that in rats or mice, but the pattern of AHR mRNA expression in tissues is similar. PMID- 10873599 TI - Phenotypic expression of IGF binding protein transcripts in muscle, in vitro and in vivo. AB - The actions of the insulin-like growth factors (IGF-I and IGF-II) which are essential components of skeletal muscle growth are mediated via their receptors and modulated by six binding proteins (IGFBPs). We studied IGFBP transcripts in C2C12 cell cultures and in adult control and denervated gastrocnemius muscle. IGFBP-2, -4, -5, and -6 were detected in C2C12 cells. IGFBP-6 mRNA levels remained unchanged, IGFBP-2 levels decreased and IGFBP-4 and -5 increased over 1, 5, and 9 days after serum reduction. In a range of adult muscles studied, IGFBP-4 mRNA levels were similar, IGFBP-5 was present at different levels in slow and fast muscles and IGFBP-6 had the lowest expression in the tibialis anterior. Denervation resulted in dramatic up-regulation of IGFBP-4 and -5 transcripts but there was no change in IGFBP-6. These results suggest that either lack of neural input and/or mechanical loading, both of which contribute to muscle atrophy, affect IGFBP expression. PMID- 10873600 TI - Glioma inhibition by HGF/NK2, an antagonist of scatter factor/hepatocyte growth factor. AB - Strategies that antagonize growth factor signaling are attractive candidates for the biological therapy of brain tumors. HGF/NK2 is a secreted truncated splicing variant and potential antagonist of scatter factor/hepatocyte growth factor (SF/HGF), a multifunctional cytokine involved in the malignant progression of solid tumors including glioblastoma. U87 human malignant glioma cells that express an autocrine SF/HGF stimulatory loop were transfected with the human HGF/NK2 cDNA and clonal cell lines that secrete high levels of HGF/NK2 protein (U87-NK2) were isolated. The effects of HGF/NK2 gene transfer on the U87 malignant phenotype were examined. HGF/NK2 gene transfer had no effect on 2 dimensional anchorage-dependent cell growth. In contrast, U87-NK2 cell lines were approximately 20-fold less clonogenic in soft agar and approximately 4-fold less migratory than control-transfected cell lines. Intracranial tumor xenografts derived from U87-NK2 cells grew much slower than controls. U87-NK2 tumors were approximately 50-fold smaller than controls at 21 days post-implantation and HGF/NK2 gene transfer resulted in a trend toward diminished tumorigenicity. This report shows that the predominant effect of transgenic HGF/NK2 overexpression by glioma cells that are autocrine for SF/HGF stimulation is to inhibit their malignant phenotype. PMID- 10873601 TI - Characterization of the tyrosine kinase Tnk1 and its binding with phospholipase C gamma1. AB - Tnk1 is a nonreceptor tyrosine kinase cloned from CD34+/Lin-/CD38- hematopoietic stem/progenitor cells. The cDNA predicts a 72-kDa protein containing an NH(2) terminal kinase, a Src Homology 3 (SH3) domain, and a proline-rich (PR) tail. We generated rabbit antiserum to a GST-Tnk1(SH3) fusion protein. Affinity-purified anti-Tnk1 antibodies specifically recognized a 72-kDa protein in Tnk1-transfected COS-1 cells and cells which express Tnk1 mRNA. Western blot analysis indicated that Tnk1 is expressed in fetal blood cells, but not in any other hematopoietic tissues examined. Tnk1 immunoprecipitated from cell lysates possessed kinase activity and was tyrosine phosphorylated. In binding experiments with a panel of GST-fusion constructs, only GST-PLC-gamma1(SH3) interacted with in vitro translated Tnk1. GST-protein precipitations from cell lysates confirmed that GST PLC-gamma1(SH3) associated with endogenously expressed Tnk1. Conversely, GST Tnk1(PR) protein constructs complexed with endogenously expressed PLC-gamma1. The association of Tnk1 with PLC-gamma1 suggests a role for Tnk1 in phospholipid signal transduction. PMID- 10873603 TI - Expression of leptin receptors and induction of IL-1beta transcript in glial cells. AB - To examine the role of leptin in the immune function of the brain, we examined the effect of leptin on interleukin-1beta (IL-1beta) expression in mouse primary cultured glial cells. The expression of leptin receptor isoforms Ob-Ra and Ob-Rb mRNA was detected by RT-PCR analysis of total RNA from primary cultured glial cells. Protein of leptin receptor was also expressed in mouse primary cultured glial cells as evaluated by Western blotting analysis. Leptin increased the expression of IL-1beta mRNA evaluated by RT-PCR. The expression of IL-1beta transcript peaked 2 to 6 h after leptin application. These results indicate that leptin could induce IL-1beta transcript in the brain and that one of the target cells of the leptin-induced IL-1beta transcript may be a glial cell. PMID- 10873602 TI - Direct inhibition of in vitro PLD activity by 4-(2-aminoethyl)-benzenesulfonyl fluoride. AB - While conducting a purification protocol of phospholipase D (PLD) from human granulocytes, we observed that PLD activity was inhibited by a commonly-used protease inhibitor cocktail. Of the six inhibitors present in the cocktail, the serine protease inhibitor, 4-(2-aminoethyl)-benezensulfonyl fluoride (AEBSF), was found to be the sole inhibitor of PLD. AEBSF caused a loss of neutrophil and purified plant PLD activities in vitro, but not in intact cells at the concentrations used, nor did it affect the related phospholipases A(2) and C, that were utilized as specificity controls. The compound AEBSNH(2), which has the fluoride replaced by an -NH(2) group, failed to affect PLD activity as did other compounds structurally related to AEBSF with known protease inhibitory capabilities. Finally, basal- and agonist-stimulated PLD activity was inhibited in phosphatidylcholine-specific anti-PLD immunoprecipitates (IC(50) = 75 microM). These results suggest that AEBSF, in an effect probably unrelated to its anti proteolytic ability, directly interferes with PLD enzymatic activity, making it a significant compound to begin analyzing the role of PLD in mammalian cell signaling. PMID- 10873604 TI - The 5'-end of hTERT mRNA is a good target for hammerhead ribozyme to suppress telomerase activity. AB - Because the expression level of hTERT, a catalytic subunit of human telomerase, is a rate-limiting determinant of telomerase activity, hTERT mRNA would be an excellent target of hammerhead ribozymes for the regulation of telomerase activity. We studied the efficiency of several hammerhead ribozymes targeting hTERT mRNA by transient and stable transfection procedures. To screen the potency of the ribozymes, transient ribozyme transfection and telomerase determination were performed. The ribozyme targeting 13 nucleotides downstream from the 5'-end of hTERT mRNA (13-ribozyme) exhibited the strongest telomerase-inhibitory activity, and the ribozyme to target 59 nucleotides upstream from the poly(A) tail showed clear activity. A stable transfection study confirmed that the 13 ribozyme suppressed telomerase. These observations suggest that the 13-ribozyme can regulate telomerase activity and may possess potential for cancer therapy. PMID- 10873605 TI - IGF-1 regulates cardiac fibroblast apoptosis induced by osmotic stress. AB - In this study we have determined the ability of IGF-1 to protect cardiac fibroblasts against osmotic-induced apoptosis and investigated the potential mechanism(s) underlying this protection. Treatment with IGF-1 (1-100 ng/ml) promoted a dose dependent increase in cell survival against osmotic cell death. Both Akt and ERK1/2 were rapidly phosphorylated by IGF-1 and blocked by wortmannin and PD98059, inhibitors of their upstream activators respectively. However, IGF-1-induced protection was mediated via a wortmannin-dependent but PD98059-independent pathway as determined by cell survival assay suggesting a role of PI3-K/Akt. Furthermore, IGF-1 appeared to reduce the activation of a number of early components in the apoptotic pathway in a wortmannin dependent manner including the osmotic stress-induced perturbation in mitochondrial membrane potential, cleavage and activation of caspase-3 and DNA fragmentation. Thus, the results suggest that IGF-1 regulates osmotic stress-induced apoptosis via the activation of the PI3-K/Akt pathway at a point upstream of the mitochondria and caspase-3. PMID- 10873606 TI - Protein kinase A-dependent phosphorylation of aquaporin-1. AB - The molecular mechanisms for regulating water balance in many tissues are unknown. Like the kidney, the eye contains multiple water channel proteins (aquaporins) that transport water through membranes, including two (AQP1 and AQP4) in the ciliary body, the site of aqueous humor production. Previous results from our laboratory demonstrated that water channel activity of AQP1 was significantly increased by protein kinase A (PKA) activators such as cyclic-AMP (cAMP) and forskolin. The purpose of this study is to determine whether PKA dependent protein phosphorylation is involved in the regulation of water channel activity of AQP1. Results presented here suggest that catalytic subunit of protein kinase A significantly increased the amount of phosphorylated AQP1 protein. In addition, these results indicated that cAMP-responsive redistribution of AQP1 may be regulated by phosphorylation of AQP1. Moreover, they provide new insights on the molecular mechanisms for regulating water balance in several tissues involving rapid water transport such as ciliary epithelium. In addition, they suggest important potential roles for AQP1 in several clinical disorders involving rapid water transport such as glaucoma. PMID- 10873607 TI - Induction of cytochrome P450 (CYP)1A1, CYP1A2, and CYP3A4 but not of CYP2C9, CYP2C19, multidrug resistance (MDR-1) and multidrug resistance associated protein (MRP-1) by prototypical inducers in human hepatocytes. AB - Human hepatocytes cultured serum-free for up to 6 weeks were used to study expression and induction of enzymes and membrane transport proteins involved in drug metabolism. Phase I drug metabolizing enzymes cytochrome P450 (CYP)1A1, CYP1A2, CYP2C9, CYP2C19, CYP2E1, and CYP3A4 were detected by Western blot analyses and, when appropriate, by enzymatic assays for ethoxyresorufin-O deethylase(EROD)-activity and testosterone-6beta-hydroxylase(T6H)-activity. Expression of the membrane transporter multi-drug resistance protein (P glycoprotein, MDR-1), multidrug resistance-associated protein (MRP-1), and lung resistance protein (LRP) was maintained during the culture as detected by RT-PCR and Western blot analyses. Model inducers like rifampicin, phenobarbital, or 3 methylcholanthrene and beta-naphtoflavone were able to induce CYP1A or CYP3A4 as well as EROD or T6H activities for up to 30 days. CYP2C9, CYP2C19 and CYP2E1 expression was maintained but not inducible for 48 days. Also, rifampicin and phenobarbital were unable to increase MDR-1 and MRP-1 protein levels significantly. PMID- 10873608 TI - p63 and p73 transactivate differentiation gene promoters in human keratinocytes. AB - p53 and its two homologues, p73 and p63, share considerable structural similarities, an ability to interact between themselves and to transactivate the same promoters, including for example p21. Furthermore, p73 can induce cell death via its interaction with c-Abl. In contrast, p63 has been demonstrated to be essential for limb and skin formation. We evaluated the expression of p63 and p73 in differentiating human keratinocytes in vitro. Skin biopsy and primary cultures of normal human epidermal keratinocytes (NHEK) express both p73 and p63. NHEK induced to differentiate in vitro by high calcium exposure show induction of p73 delta and downregulation of all isoforms of p63. This latter gene is predominantly expressed in its transcriptionally inactive form, DeltaNp63. We further evaluated the effect of either p73s or p63 transfected in either NHEK or transformed human keratinocytes (HaCat cells). p73 gamma, delta, and p63 were able to transactivate the promoters of loricrin and involucrin in both NHEK and HaCat cells. These results suggest the involvement of both p73 and p63 genes in keratinocyte terminal differentiation. PMID- 10873609 TI - Cardiac calcineurin during transition from hypertrophy to heart failure in rats. AB - We studied an alteration of calcineurin expression in the heart and its modification by cyclosporin A and an ACE inhibitor, temocapril, using Dahl salt sensitive (DS) rats with hypertensive left ventricular hypertrophy (LVH) and congestive heart failure (CHF). Calcineurin protein expression in the LV myocardium was increased in the LVH stage, but then decreased during CHF transition. Chronic cyclosporin A treatment (10 mg/kg/day), which inhibits calcineurin activity, could not block the increases of LV weight and dimensions and did not improve the LV systolic function during the CHF transition. In contrast, chronic temocapril treatment (20 mg/kg/day) restored the downregulation of calcineurin expression, but progression of the hypertrophic process was inhibited. Therefore, cardiac calcineurin is increased in the hypertensive LVH and may be involved in the development of the adaptive hypertrophic process. However, calcineurin expression is downregulated during CHF transition and may no longer play a major role in the pathogenesis of myocardial hypertrophy in the failing hearts. PMID- 10873610 TI - Cloning and characterization of the 5'-flanking region for the mouse phospholipase C-delta1 gene. AB - To date, little is known about the molecular mechanisms controlling the regulation of phospholipase C-delta1 (PLC-delta1) gene expression. To understand the mechanisms responsible for the regulation of PLC-delta1 gene expression, the 5'-flanking region of the mouse PLC-delta1 gene was isolated from a mouse genomic DNA library. Primer extension analysis revealed that there is a single transcriptional start site located at 127 bases upstream from the translation start codon in the mouse PLC-delta1 gene. DNA sequence analysis showed that the sequence around the transcriptional start site is very GC-rich and has no TATA or CAAT boxes. Transient expression of a luciferase reporter gene under the control of serially deleted 5'-flanking sequences revealed that the 160-base-pair region from -622 to -462 upstream of the transcriptional start site includes a positive cis-acting element(s) for the efficient expression of the PLC-delta1 gene. Gel retardation analysis suggests that multiple transcription factors bind to separate sites on the promoter region. Based on these results, our study suggests that the minimal essential region located at -622 to +70 is fully sufficient to confer high-level transcriptional activity and contains high-affinity binding elements for multiple transcription factors. PMID- 10873611 TI - Manganese superoxide dismutase levels are elevated in a proportion of amyotrophic lateral sclerosis patient cell lines. AB - The most frequent genetic causes of amyotrophic lateral sclerosis (ALS) determined so far are mutations occurring in the gene for copper/zinc superoxide dismutase (CuZnSOD). The mechanism may involve inappropriate formation of hyroxyl radicals, peroxynitrite or malfunctioning of the SOD protein. We hypothesized that undiscovered genetic causes of sporadically occurring amyotrophic lateral sclerosis might be found in the mechanisms that create and destroy oxygen free radicals within the cell. After determining that there were no CuZnSOD mutations present, we measured superoxide production from mitochondria and manganese superoxide dismutase (MnSOD), glutathione peroxidase, NFkappaB, Bcl-2 and Bax by immunoblot. Of the ten sporadic patients we tested we found three patients with significantly increased concentrations of MnSOD. These patients also had lower levels of superoxide production from mitochondria and decreased expression of Bcl 2. No mutations were found in the cDNA sequence of either MnSOD in any of the sporadic patients. A patient with a CuZnSOD mutation (G82R) used as a positive control showed none of these abnormalities. The patients displaying the MnSOD aberrations showed no specific distinguishing features. This result suggests that the cause of ALS in a subgroup of ALS patients (30%) is genetic in origin and can be identified by these markers. The alteration in MnSOD and Bcl-2 are likely epiphenomena resulting from the primary genetic defect. It suggests also that the oxygen free radicals are part of the cause in this subgroup and that dysregulation of MnSOD or increased endogenous superoxide production might be responsible. PMID- 10873612 TI - Isolation of two novel human RhoGEFs, ARHGEF3 and ARHGEF4, in 3p13-21 and 2q22. AB - RhoGEFs play an important role in various signaling cascades and are implicated in human conditions like cancer and mental retardation. A database search combined with screening of a human neuronal teratocarcinoma library identified two novel RhoGEFs, ARHGEF3 and ARHGEF4 (HGMW-approved symbols). The widely expressed ARHGEF3 transcript of 3561 nucleotides encodes a polypeptide of 526 amino acids with homology to NET1. The ARHGEF4 gene generates two transcripts of 3665 and 4000 nucleotides that translate into 720 amino acid residues. Expression of ARHGEF4 is restricted to brain and the encoded protein shows homology to collybistin. FISH analysis of genomic clones mapped ARHGEF3 to 3p13-21 and ARHGEF4 to 2q22. PMID- 10873613 TI - Expression and transcriptional regulation of the human alpha1, 3 fucosyltransferase 4 (FUT4) gene in myeloid and colon adenocarcinoma cell lines. AB - In fucosyltransferase genes, mRNA expression is regulated in a cell-type-specific manner. The expression level of human fucosyltransferase 4 (FUT4) mRNA is high in both colon adenocarcinoma and myeloid cell lines. We will demonstrate here cell specific expression and transcriptional regulation of the FUT4 gene. FUT4 has two different transcription initiation sites that respectively produce long- and short-form mRNAs. To determine the major FUT4 transcript in colon adenocarcinoma and myeloid cell lines, we analyzed the transcriptional starting sites of the FUT4 gene in myeloid and colon adenocarcinoma cell lines, using 5'-RACE, RT-PCR, and luciferase analysis. The results suggested that the expression level of short form mRNA is higher than the long-form transcript in the colon adenocarcinoma cell lines and that the expression level of long-form mRNA is higher than the short-form transcript in the myeloid cell lines. Using a luciferase assay, we identified a functional DNA portion within FUT4 genomic DNA that confers a colon adenocarcinoma cell line-specific enhancer, located in nucleotide number (nt) 256 to -44, and a myeloid cell line-specific enhancer, located in nt -686 to 582. The present results suggest that these elements play a critical role in the colon adenocarcinoma and leukemia cell-specific transcriptional regulation of the FUT4 gene. PMID- 10873614 TI - Enhanced secretion of ApoB by transfected HepG2 cells overexpressing fibrinogen. AB - HepG2 cells stably transfected with cDNA-encoding single fibrinogen chains overexpress fibrinogen and have increased (4-fold) secretion of apolipoprotein B. Overexpression of fibrinogen does not affect the secretion of three representative acute-phase proteins but causes a small increase in albumin secretion. Enhanced apolipoprotein B secretion is due to less intracellular degradation and not to increased expression. The increased secretion of apolipoprotein B is independent of the acute-phase response, since stimulation of fibrinogen gene expression by interleukin 6 did not affect secretion. HepG2 cells overexpressing fibrinogen chains had increased 3-hydroxy-3-methylglutaryl coenzyme A reductase mRNA levels, enhanced cholesterol production but normal levels of triglyceride and phospholipid synthesis and of sterol response binding proteins. These results, that associate overexpression of fibrinogen with enhance apolipoprotein B secretion, may be significant since epidemiological studies indicate that elevated levels of fibrinogen and lipids are independent risk factors in coronary artery disease. PMID- 10873615 TI - Myoneurin, a novel member of the BTB/POZ-zinc finger family highly expressed in human muscle. AB - Initially characterized as Drosophila developmental regulators, the BTB/POZ and zinc finger proteins (BTB/POZ-ZF) constitute a growing family of proteins with gene expression regulatory functions since they have been shown to be involved in both transcriptional activation and repression of various genes in a broad range of species, including mammals. Here we report the cloning of a novel human transcript, coding for a 68-kDa deduced BTB/POZ-ZF protein. This molecule, called myoneurin on the basis of its prevalent expression in the neuromuscular system, contains an amino-terminal BTB/POZ domain and eight tandemly repeated zinc-finger motifs of the C(2)H(2) type. The murine myoneurin, identified in the mouse embryo, is highly homologous to the human protein. PMID- 10873616 TI - Identification of dipeptidyl peptidase III in human neutrophils. AB - We have found activity of dipeptidyl peptidase (DPP) III, one of the most important enkephalin-degrading enzymes in the central nervous system, in human neutrophils. HPLC analysis of the peptide fragments produced by treatment of leucine-enkephalin with isolated neutrophils in the presence of inhibitors of other enkephalin-degrading enzymes revealed that the enzyme in human neutrophils cleaved dipeptides from the NH(2) terminus of leucine-enkephalin, suggesting the presence of DPPIII activity in human neutrophils. Using a specific synthesized substrate and proteinase inhibitors, it was found that the neutrophils have 19.2 +/- 3.6 microM/h/5 x 10(6) cells of beta-naphthylamine for the enzyme. It was also confirmed that spinorphin and tynorphin, both reported to inhibit the activities of enkephalin-degrading enzymes, had potent inhibitory activities (IC(50): 4.0 and 0.029 microg/ml, respectively) against the enzyme. The presence of DPPIII activity in human neutrophils suggests that the biologically active peptides which are associated with enkephalin play a physiological role in regulating enkephalin or inflammatory mechanisms in peripheral tissues. PMID- 10873617 TI - The transcript for a novel protein with a zinc finger motif is expressed at specific stages of mouse spermatogenesis. AB - The cDNA for an RNA that is expressed predominantly in mouse spermatogenic cells was cloned and characterized. It was found to encode novel zinc finger protein. We first generated a cDNA fragment from mouse osteoblastic cells by the differential display method. To our surprise, Northern blot analysis revealed that the corresponding transcript was expressed at high levels in the testis rather than in osteoblastic cells. Therefore, using this fragment as a probe, we isolated the full-length cDNA (3340 bp) from a mouse testis cDNA library. Analysis of the open reading frame of the cDNA indicated that the encoded protein was a polypeptide of 942 amino acids residues that included three distinct domains, namely, a zinc finger domain of the Cys(2)-His(2) type, four basic amino acid-rich domains, and a myosin II-homology domain. In situ hybridization indicated that the transcript was present in seminiferous tubules of adult mice. Elevated expression of the transcript during testicular development in mice was restricted to spermatocytes at the pachytene stage of meiotic prophase and to round and elongated spermatids, as indicated by Northern blot analysis and RT PCR. Our results suggest that this novel zinc finger protein might act as a transcriptional regulator during spermatogenesis and, in particular, during meiotic division. PMID- 10873618 TI - Dual anticancer activity of 8-Cl-cAMP: inhibition of cell proliferation and induction of apoptotic cell death. AB - 8-Cl-cAMP induces apoptotic cell death in human cancer cells. To look at this more closely, we examined the changes in the levels of Bcl-2 family proteins during 8-Cl-cAMP-induced apoptosis of SH-SY5Y human neuroblastoma cells. Following the treatment with 8-Cl-cAMP, Bcl-2 was transiently down-regulated and Bad was increased continuously up to day 5. In addition, overexpression of Bcl-2 efficiently blocked the 8-Cl-cAMP-induced apoptosis, suggesting Bcl-2 family proteins may be involved in the 8-Cl-cAMP-induced apoptosis. The contribution of the apoptotic cell death and the inhibition of cell proliferation in the 8-Cl cAMP-induced growth inhibition was closely monitored in the Bcl-2-overexpressing cells. Though the apoptosis was reduced significantly, no significant difference was observed in the inhibition of cell proliferation up to day 2 of 8-Cl-cAMP treatment. These results suggest that 8-Cl-cAMP exerts anticancer activity by two distinct mechanisms, i.e. , through the inhibition of cell proliferation as well as the induction of apoptosis. Supporting this notion was the observations that (1) suppression of apoptosis by zVAD did not abrogate 8-Cl-cAMP-induced inhibition of cell proliferation, and (2) 8-Cl-cAMP did not show additive inhibition of cell proliferation in RIIbeta-overexpressing cells. PMID- 10873619 TI - Influence of bcl-2-related proteins on matrix metalloproteinase expression in a rat glioma cell line. AB - The expression of bcl-2-related proteins has been shown to be a key element in tumoral malignancy. The degradation of the extracellular matrix (ECM) by specialized matrix metalloproteinases (MMPs) is another major step in tumor invasion and metastasis. We have examined, in a rat glioma cell line A15A5, the effect of the stable transfection of human bcl-2, bax and bcl-xl on MMPs expression. Using a zymographic assay, we found that all transfected cell lines expressed a gelatinase activity which is predominantly associated with MMP-9. In bcl-2 and bcl-xl transfected cells, the transcription of MMP-9 was decreased compared to that of control or bax transfected cells. In addition, in bax transfected A15A5, we observed a down regulation of TIMP-1, the inhibitor of MMP 9. These results suggest that the ratio between MMP-9 and its inhibitor TIMP-1 is tightly controlled in cells overexpressing bcl-2 related proteins (i.e., high ratio in bax transfected A15A5 and low ratio in bcl-2 transfected A15A5). However, MMPs secreted by bcl-2 transfected cells were still capable of hydrolyzing FasL present on human lymphocytes. Our results suggest that the expression of bcl-2 related proteins could participate in the regulation of MMP 9/TIMP-1 in gliomas. PMID- 10873620 TI - Losartan inhibits cellular uptake of oxidized LDL by monocyte-macrophages from hypercholesterolemic patients. AB - Angiotensin II (Ang II) and oxidized LDL (Ox-LDL) are risk factors for atherosclerosis, and both of them contribute to macrophage cholesterol accumulation, the hallmark of early atherosclerosis. As Ang II was shown to increase macrophage uptake of Ox-LDL, we investigated the effect of losartan, an Ang II receptor antagonist with antiatherogenic properties, on the cellular uptake of Ox-LDL by human monocyte-derived macrophages (HMDM) from hypercholesterolemic patients. Eight normotensive hypercholesterolemic patients were treated with losartan (50 mg/day) for a period of 4 weeks. Losartan therapy did not significantly affect the degradation of native LDL by the patients' HMDM. However, losartan therapy significantly reduced HMDM uptake of Ox-LDL as shown by a 78% reduction in Ox-LDL cell-association and a 21% reduction in Ox-LDL degradation. CD36 (an Ox-LDL receptor) mRNA expression in HMDM obtained after losartan treatment was decreased by 54% compared to HMDM obtained before treatment. The ability of losartan to inhibit HMDM CD36 mRNA expression and, hence, Ox-LDL uptake and macrophage foam cell formation is probably related to the blockage of Ang II binding to the cell surface and thus to the prevention of Ang II atherogenic effects. PMID- 10873621 TI - Structure-activity relationships for selected sulfur-rich antithrombotic compounds. AB - We assessed the antithrombotic activity of some simple organosulfur compounds which have some of the functionality found in the disulfide ajoene, a pharmacologically active compound isolated from garlic. The results establish that antithrombotic activity is associated with disulfides directly attached to a phenyl ring and is further enhanced by an alpha-sulfonyl group. CH(3)SO(2)CH(2)SSPh proved to be a potent inhibitor of platelet aggregation with an IC(50) of 5 microM. PMID- 10873622 TI - Deregulation of the cytoplasmic tyrosine kinase cSrc in the absence of a truncating mutation at codon 531 in human bladder carcinoma. AB - The involvement of the cytoplasmic tyrosine kinase cSrc was investigated in human bladder carcinogenesis. Kinase activity was determined in tissue lysates from bladder transitional cell carcinoma (TCC) relative to normal epithelia. Strong kinase activation was observed at all stages of carcinogenesis with a peak at the stage pT1, where tumor cells disrupt the basement membrane and invade the submucosa. In agreement with a role for cSrc in cell invasion, immunocytochemistry analysis showed a strong staining of invading cells. An increase in cSrc protein level were also found in most tumor samples, however, it did not correlate with an increase in activity (r = 0.44) suggesting that cSrc is deregulated in these tumors. Indeed, high Src activity was affinity-purified from a column (IRSVSSDGHE(p)YIYVDP-Affigel 10) that specifically retains active Src. Enzymatic regulation involves the C-terminus, recently found mutated at codon 531 in a subset of advanced human colon cancers. However, no such mutations were detected in TCC, suggesting the existence of other mechanisms for kinase activation. PMID- 10873623 TI - Molecular cloning and characterization of a new insulin/IGF-like peptide of the nematode Caenorhabditis elegans. AB - Diapause, aging, and fat accumulation in Caenorhabditis elegans are regulated by DAF-2, a homolog of mammalian insulin/insulin-like growth factor-I (IGF-I) receptors. We have cloned and characterized a C. elegans gene encoding a new insulin/IGF-like peptide. The gene containing three exons encodes a precursor protein 95 residue long. Although the putative precursor contains a signal peptide, B chain, C peptide, and A chain like the preproinsulin, the mature peptide consists of one polypeptide-like IGF. The predicted tertiary structure seems similar to crystal structure of insulin. Therefore, the peptide may be a hybrid molecule of insulin and IGF. The peptide expression was detected at the embryonic and several larval stages. Disruption of the peptide production led to an extended life span like the daf-2 mutation, suggesting that the peptide should be one of the ligands of the DAF-2. This is the first description of the peptide that mediates animal longevity. PMID- 10873624 TI - 4-hydroxy-2-nonenal, the end product of lipid peroxidation, is a specific inducer of cyclooxygenase-2 gene expression. AB - Cyclooxygenase-2 (COX-2), an enzyme responsible for catalyzing the committed step in prostanoid biosynthesis, is the product of an immediate early gene capable of being upregulated by diverse stimuli. Based on the experimental evidence that oxidative stress is associated with the upregulation of COX-2, we evaluated the effect of the oxidized fatty acid metabolites on COX-2 induction in rat liver epithelial RL34 cells. Among the compounds tested, only 4-hydroxy-2-nonenal (HNE), a highly mutagenic and genotoxic aldehyde generated during oxidative stress, dramatically induced COX-2. Enhanced gene expression of COX-2 by treatment with HNE was evident as a drastic elevation of the mRNA level. We also found that intracellular glutathione status was strictly related to HNE-induced COX-2 expression. These findings suggest the presence of a signaling pathway in the cellular response mediated by locally produced lipid peroxidation products under oxidative stress. PMID- 10873625 TI - Molecular cloning of a cyclin-like protein associated with cyclin-dependent kinase 3 (cdk 3) in vivo. AB - cdk3 has been considered to be rate-limiting for cell cycle progression of mammalian cells while its precise function remains to be elucidated. To assess cdk3 function, a cDNA coding for a cyclin-like protein (designated as ik3-1 from an interactor-1 with cdk3) was isolated with the yeast two-hybrid system using a cyclin-dependent kinase 3 (cdk3) cDNA as bait. p70(ik3-1) (a 70-kDa protein designated as p70(ik3-1)) seems to belong to the cyclin family as its C-terminal domain composed of 124 amino acids resembles the highly conserved cyclin box. Coimmunoprecipitation indicated that p70(ik3-1) binds to p35(cdk3) in vivo. The ik3-1 gene may belong to a multigene family and is highly conserved during evolution. mRNA expression of ik3-1 was low in the early G1 phase, upregulated during G1 progression, maximal at a mid-late G1 point, and declined gradually thereafter, suggesting that it may work mainly in G1 phase. PMID- 10873626 TI - Stimulation of cell invasion and migration by alcohol in breast cancer cells. AB - Increasing epidemiological studies suggest that alcohol consumption confers a high risk for development of breast cancer. In this study, we found that biologically relevant concentrations of alcohol elicited a significant stimulation of cell adhesion, migration, and invasion in MCF-7 human breast cancer cells. Moreover, the promotion of invasion and migration potential by alcohol was associated with the significant decrease of E-cadherin, alpha, beta, and gamma three major catenin, and BRCA1 expression. In addition, an enhanced expression of BRCA1 significantly blocked alcohol-stimulated cell invasion. Thus, our present study suggests that alcohol as a breast cancer risk factor plays an important role not only in carcinogenesis, but also in promotion of cell invasion and migration. PMID- 10873627 TI - Quantitative determination of the expression of xeroderma pigmentosum F gene in human nonmelanoma skin cancers. AB - Nonmelanoma skin cancers (NMSC) has been evidenced with an impaired function in nucleotide excision repair (NER). However, malfunction of NER elements in NMSC has not been identified. Xeroderma pigmentosum F (XPF) is an essential subunit in NER and functions as a 5'-incision enzyme when repairing damaged DNA. So far, neither XPF's protein nor antibody is commercially available. To explore the expression of XPF in NMSC, the gene was determined by quantitative reverse transcription-polymerase chain reaction (RT-PCR). All the designed primers specifically amplified XPF cDNA as demonstrated by nested PCR, and one set of the primers was mimic constructed to form a controlled cDNA for the semiquantification of XPF gene in NMSC. The results indicated that the quantities of XPF expression of BCC and SCC specimens were approximately 57.0 and 76.4% less than that of normal skins, respectively. This paper indicates that the decrease expression of XPF gene may be one of mechanisms for impaired NER in NMSC, and the feasible and quantitative primers used in the experiments may explore the study of XPF in etiology of carcinogenesis. PMID- 10873628 TI - Human histo-blood group ABO glycosyltransferase genes: different enhancer structures with different transcriptional activities. AB - The enhancer element of the human histo-blood group ABO glycosyltransferase gene has been demonstrated to be located -3.7 kb upstream from the transcription start site and to be composed of four tandem repeats of a 43-bp unit. Recently we identified three different enhancer structures among the allelic A, B, and O glycosyltransferase genes. The enhancer structure with four 43-bp units is present in the B and O genes, but not in the A gene. The corresponding enhancer region of the A gene contains only one 43-bp unit, and within this unit a nucleotide substitution exists when compared with the consensus sequence. Through transient transfection assays, the transcriptional activity of the A-gene enhancer region was demonstrated to be less than 1% of that of the B-gene enhancer. The difference between the transcriptional activities of the two enhancers became more significant when acting in concert with the ABO-gene's native promoter. The different repeat numbers of the 43-bp unit possessed by the two allelic genes were shown to be the main reason for the vast difference in the transcriptional activities between the A-gene and B-gene enhancers. PMID- 10873629 TI - Hemin is kinetically trapped in cytochrome b(5) from rat outer mitochondrial membrane. AB - Cytochrome b(5) from the outer mitochondrial membrane of rat liver (OM cyt b(5)) is substantially more stable to thermal and chemical denaturation than cytochrome b(5) from the endoplasmic reticulum of bovine liver (microsomal, or Mc cyt b(5)). In contrast, the corresponding apoproteins have similar stability, suggesting stronger interactions between hemin and the polypeptide in OM cyt b(5). Whereas complete transfer of hemin from bovine Mc cyt b(5) to apomyoglobin at pH 5.2 takes less than 1 h, hemin transfer from OM cyt b(5) is unmeasurably slow. Coupled with the previously reported 1:1 ratio of hemin orientational isomers in OM cyt b(5), this finding suggests that the cofactor is kinetically trapped under physiologically relevant conditions. This conclusion is confirmed by (1)H NMR studies which show that the hemin isomeric ratio changes when the protein is incubated for several hours at 68 degrees C. Interestingly, the orientational isomer favored in OM cyt b(5) is the form less favored in all other known cytochromes b(5). PMID- 10873630 TI - Site-specific integration of a transgene mediated by a hybrid adenovirus/adeno associated virus vector using the Cre/loxP-expression-switching system. AB - As vectors, adenoviruses (Ads) have many attractive advantages for in vivo gene therapy. However, Ads do not usually integrate into the host genome and gene expression is, thus, transient. Adeno-associated virus (AAV) integrates into a specific locus (AAVS1) on the human host's chromosome 19, while conventional recombinant AAV (rAAV) vectors do not possess this property because such vectors lack the rep gene. AAV vectors carrying the rep gene do not have enough space for insertion of a transgene. We have constructed a hybrid adenovirus/adeno associated virus (Ad/AAV) vector which has the advantages of both Ads and AAVs. Given that the rep gene products inhibit propagation of Ads, we used the Cre/loxP expression-switching system to regulate the expression of the rep gene. The Ad/AAV vector easily propagates, can efficiently infect a broad range of cell types, and can integrate into a specific locus on host chromosomes. PMID- 10873631 TI - Complex formation between hepatitis C virus core protein and p21Waf1/Cip1/Sdi1. AB - The core protein (Core) of hepatitis C virus (HCV) has been known to play an important role in hepatocarcinogenesis. By using glutathione S-transferase (GST) pull-down assay, we show here that Core formed a complex with p21Waf1/Cip1/Sdi1 (p21) cell cycle regulator. The deletion-mapping analysis revealed that a portion near the N-terminus of Core (amino acids 24-52) and a C-terminal portion of p21 (amino acids 139-164) were involved in the complex formation. The complex formation was not impaired by point mutations of p21 at residues 147, 149, and 150, which have been reported to abrogate interaction of p21 with proliferating cell nuclear antigen (PCNA), discriminating the Core-binding sequence from the PCNA-binding sequence. Due to the close vicinity of the binding sites, however, Core and PCNA competed with each other when interacting with p21. The distinct interaction between Core and p21 may provide a new aspect to the studies of HCV pathogenesis. PMID- 10873632 TI - Prostaglandins up-regulate vascular endothelial growth factor production through distinct pathways in differentiated U937 cells. AB - We previously reported that cyclooxygenase (COX)-2 was predominantly expressed in macrophages of human colonic adenomas (Int. J. Cancer 83, 470-475.). The role of prostaglandins (PGs) produced by COX-2-expressing macrophages in colon carcinogenesis is still unclear. Here we show that PGs up-regulate vascular endothelial growth factor (VEGF) production by activated macrophages through their specific receptors. mRNAs of both PGE-specific receptors and peroxisome proliferator-activated receptor gamma (PPARgamma), a member of the nuclear receptor superfamily of ligand-dependent transcription factors, were expressed in phorbol 12-myristate 13-acetate-differentiated U937, a human macrophage model (H Mac). Prostaglandin E(1) (PGE(1)) and 15-deoxy-Delta(12,14)-PGJ(2) (a potent PPARgamma ligand, 15d-PGJ(2)) dramatically increased VEGF production. The combination of PGE(1) and 15d-PGJ(2) additively increased VEGF production. In addition, PGE(1) significantly increased cAMP formation, whereas 15d-PGJ(2) did not affect cAMP formation. The effect of the combination of PGE(1) and 15d-PGJ(2) on cAMP formation was similar to that of PGE(1) alone. Unexpectedly, 15d-PGJ(2) also drastically increased IL-1beta production, an indicator of macrophage activation, although PGE(1) only mildly increased it. Additional enhancement of IL-1beta production was observed in the combination of PGE(1) and 15d-PGJ(2). These results suggest that PGs dramatically increased VEGF production by activated macrophages through specific PGE receptor and PPARgamma-mediated processes and that PGs may thereby promote tumor growth through VEGF production. PMID- 10873633 TI - Collateral existence of folded and extended conformations of the beta-Ala moiety in a model peptide. AB - The single-crystal X-ray diffraction analysis of a nonchiral beta-Ala-containing model peptide, Boc-beta-Ala-Aib-OCH(3) 1 (beta-Ala, 3-aminopropionic acid; Aib, alpha-aminoisobutyric acid), establishes the coexistence of distinctly different backbone conformations in two crystallographically independent molecules, A and B, in the asymmetric unit. Interestingly, the central mu torsion angle around the -C(beta)-C(alpha)- bond of the conformationally flexible beta-Ala residue appears to be critical in dictating the overall distinct structural features, i.e., in molecule A it adopts a folded gauche conformation: mu = -71.0 degrees, whereas it favors an extended trans conformation, mu = 161.2 degrees, in molecule B. As expected, the stereochemically constrained Aib residue preferred an energetically favorable folded backbone conformation, the torsion angles being phi = 46.2 degrees and psi = 48.3 degrees for molecule A and phi = -43.6 degrees and psi = 45.5 degrees for molecule B, lying in the left-handed and right-handed helical regions of the Ramachandran map, respectively. Considering the signs as well as the magnitudes of the backbone torsional angles, molecule A typically folds into a pseudo type III' beta-turn-like structure while molecule B prefers an overall extended conformation. Entrapping the two dramatically distinct conformational characteristics in the crystalline state clearly suggests that the gauche and the trans effects of the beta-Ala moieties are indeed energetically accessible to a short linear peptide and receive strong experimental support. The analyses permitted us to emphasize that in addition to conformational constraints of the neighboring residue, the chemical nature of the side-chain acyclic substituents and the "local environments" collectively seem to influence the stabilization of the folding-unfolding behavior of the two methylene units (-CONH-CH(2)-CH(2)-CONH ) in 1. PMID- 10873634 TI - Cloning of the Microcystis aeruginosa M228 lectin (MAL) gene. AB - We have cloned and characterized the gene encoding Microcystis aeruginosa (strain M228) lectin (MAL). The gene contains 1551 nucleotides and an open reading frame for a protein of 517 amino acids with a predicted molecular weight of 55,159 Da. The carboxy-terminal region of MAL has three tandemly repeated homologous domains composed of 61 amino acids. These regions show similarity to the corresponding regions of the alpha-amylase of Clostridium beijerinckii (23% identity). The mal gene lies adjacent to an ORF that display homology to cytochrome P-450 and polyketide synthase. Southern hybridization showed that the genomic DNA of the strain M228 contained, in addition to MAL gene (mal), at least two other mal like gene. PMID- 10873635 TI - Estradiol-17beta sulfotransferase activity in canine osteosarcoma D17 cells. AB - Estrogen sulfatase and sulfotransferase (EST) activities are present in breast cancer tissues but there are no reports on EST in cancerous bone cells. We incubated [(3)H]estradiol-17beta with cells from a canine osteosarcoma D17 line for periods up to 24 h. Radioactive steroids were recovered from the media and separated into unconjugated and conjugated fractions using Sep-Pak C18 cartridges. The conjugate fraction was solvolyzed and the resulting free steroids were obtained from a second C18 cartridge. Little metabolism was apparent in 4 h of incubation, but by 24 h as much as one half of the radioactivity was seen in the conjugate fraction. Most of the conjugates were recovered as sulfates in all three experiments. HPLC profiles showed a limited metabolism of estradiol to other compounds except for estrone, which was clearly present in both free and sulfate fractions. These results suggest that EST may have a role in the local metabolism of estrogens in bone. PMID- 10873636 TI - Rapid isolation and characterization of the yeast proteasome regulatory complex. AB - The 26S proteasome, which catalyzes degradation of ubiquitinated proteins, is composed of the 20S proteasome and the 19S complex. Recently, it has been reported that the 26S complex can be dissociated into the lid complex and the 20S proteasome-base complex in a mutant yeast and that the lid complex is required for ubiquitin-dependent proteolysis. In the present study, we established methods for rapid isolation of the 19S complex, the lid complex, and the base complex from wild-type yeast. The isolated 19S complex was capable of binding to the 20S proteasome to reconstitute the 26S proteasome. In contrast with the previously reported result showing that Rpn10, a multiubiquitin chain binding subunit, is a component of the base complex, we present evidence that the lid complex isolated from wild-type yeast contains Rpn10. PMID- 10873637 TI - Prostaglandin I(2), a possible thermo-sensory mediator in Paramecium. AB - Thermo-sensory mechanisms are little understood. The protozoan, Paramecium multimicronucleatum, is sensitive and responsive to a cooling stimulus. We found that inhibitors of prostaglandin (PG) biosynthesis reduced the response to the cooling stimulus. Inversely, the response suppressed by the inhibitors was recovered by application of stable PGI(2) analogs, including iloprost. Paramecium cells showed binding activity specific for [(3)H]iloprost. Moreover, an arachidonic acid metabolite, possibly PGI(2), was rapidly increased in response to the cooling stimulus, suggesting that prostaglandin biosynthesis plays a crucial role in the cooling-sensory transduction. Paramecium may be a useful model for the investigation of the molecular basis of thermo-sensory mechanisms. PMID- 10873638 TI - Involvement of the Ras/MAPK signaling pathway in the modulation of urokinase production and cellular invasiveness by transforming growth factor-beta(1) in transformed keratinocytes. AB - Transformed PDV keratinocytes respond to TGF-beta(1) by stimulating cell motility and invasiveness concomitantly to enhancement of the urokinase-type plasminogen activator (uPA) expression/secretion. Depletion of extracellular signal-regulated kinase (ERK1, 2) proteins by treatment of PDV cells with antisense oligonucleotides reduced basal uPA production and abolished stimulation of uPA secreted levels and cell motility by TGF-beta(1). PD098059, an inhibitor of mitogen-activated protein kinase (MAPK) kinase (MEK), decreased TGF-beta(1) induced uPA mRNA expression, secreted activity in a dose-dependent manner, and abrogated TGF-beta(1)-stimulated cell motility and invasiveness. PDV-derived dominant-negative RasN17 cell transfectants secreted similar amounts of uPA and exhibited similar invasive abilities as the parental cells or control clones, but were unable to respond to TGF-beta(1) for stimulation of uPA-secreted levels and invasiveness. These results suggest that a Ras/MAPK transduction pathway is involved in the invasive response of transformed keratinocytes to TGF-beta(1). PMID- 10873639 TI - Epsilon as an initiator of translation of CAT mRNA in Escherichia coli. AB - Epsilon sequence (UUAACUUUA) has originally been found in the bacteriophage T7 gene 10 leader region. It enhances translation in Escherichia coli via base pairing with nucleotides 458-466 located in the helical domain #17 of 16S rRNA. We have recently reported that when the complementarity to 16S rRNA is extended, the epsilon is converted from an enhancer to an independent initiator of translation. Here we report the effect of two other structural parameters, positioning in mRNA and the degree of complementarity to 16S rRNA on the translation initiation activity of epsilon in E. coli cells. Our results show that epsilon displays maximal activity as a translational initiator at its natural 9-nucleotide-long complementarity to 16S rRNA and at a 16-nucleotide-long distance to the initiation codon. Under these conditions its efficiency is comparable with that of the consensus Shine-Dalgarno sequence. PMID- 10873640 TI - Identification of a novel human sterol-sensitive ATP-binding cassette transporter (ABCA7). AB - We report the identification of the full-length cDNA for a novel ATP-binding cassette (ABC) transporter from human macrophages. The mRNA is of 6.8 kb size and contains an open reading frame encoding a polypeptide of 2146 amino acids with a calculated molecular weight of 220 kDa. The predicted protein product is composed of two transmembrane domains and two nucleotide binding folds indicating that it pertains to the group of full-size ABC transporters. The novel transporter shows highest protein sequence homology with the recently cloned human cholesterol and phospholipid exporter ABCA1 (54%) and the human retinal transporter ABCR (49%), both members of the ABC transporter subfamily A. In accordance with the currently proposed classification, the novel transporter was designated ABCA7. ABCA7 mRNA was detected predominantly in myelo-lymphatic tissues with highest expression in peripheral leukocytes, thymus, spleen, and bone marrow. Expression of ABCA7 is induced during in vitro differentiation of human monocytes into macrophages. In macrophages, both the ABCA7 mRNA and protein expression are upregulated in the presence of modified low density lipoprotein and downregulated by HDL(3). Our results suggest a role for ABCA7 in macrophage transmembrane lipid transport. PMID- 10873641 TI - Adhesion to fibronectin enhances MKP-1 activation in human endothelial cells. AB - Integrin-mediated substrate adhesion of endothelial cells leads to intracellular signaling, including the activation of ERK 1/2 (extracellular regulated kinases 1 and 2), members of the mitogen-activated protein kinase (MAPK) family. MKP-1 is a dual-specificity protein phosphatase that may play an important role in regulating MAPK activity through dephosphorylation of threonine and tyrosine. Adhesion of human umbilical vein endothelial cells to fibronectin increased MKP-1 protein and mRNA levels, which reached a maximum at 60 min, while MAPK activity was maximal at 30 min. The MEK inhibitor PD98059 blocked activation of MAPK as well as the induction of MKP-1 during adhesion. The transcription inhibitor actinomycin D blocked MKP-1 induction and produced prolonged MAPK activation during adhesion. In contrast, endothelial adhesion to poly-L-lysine did not alter MAPK activity or MKP-1 levels. These findings demonstrate that integrin-mediated adhesion of endothelial cells to fibronectin results in transcriptional activation of MKP-1 through a MAPK-dependent mechanism. Regulation of MKP-1 by MAPK likely represents an important negative-feedback mechanism. PMID- 10873642 TI - Involvement of p65 in the regulation of NF-kappaB in rat hepatic stellate cells during cirrhosis. AB - We have examined the NF-kappaB binding and functional activities in two stellate cell lines derived from normal (NFSC) and cirrhotic (CFSC) rat liver. Gel mobility shift assays revealed two bands in NFSC nuclear extracts that correspond to p65/p50 heterodimers and p50/p50 homodimers. In contrast, a single and more intense band that migrates faster was detected in CFSC nuclear extracts. This band supershifts with either p65 or p50 antibody. The differential NF-kappaB binding observed in these two cell lines appears to depend on the phosphorylation of the p65 subunit rather than the expression levels of either p65 or p50. The nonphosphorylated NF-kappaB form, present in CFSC cells, possesses significantly lower transcriptional activity compared to phosphorylated NF-kappaB, found in NFSC cells. To our knowledge, this is the first report on the NF-kappaB regulation at the p65 protein in hepatic stellate cells. It is likely that this regulation affects IL-6 expression and may represent a mechanism regulating hepatocyte death during fibrogenesis. PMID- 10873643 TI - Fibroblast growth factors are translocated to the nucleus of human endothelial cells in a microtubule- and lysosome-independent pathway. AB - Exogenous acidic and basic fibroblast growth factors undergo rapid nuclear translocation in human umbilical vein endothelial cells. When nuclear translocation reaches saturation, more than 70% of the internalized growth factors are in the nuclear fraction. Lysosomal inhibitors, such as leupeptin and chloroquine, and microtubule inhibitors including colchicine and 2-methoxyl-beta estradiol neither increase nor decrease nuclear translocation. The results suggest that nuclear translocation of fibroblast growth factors does not require cytosolic accumulation or lysosomal processing and that the transportation of exogenous growth factors across the cytoplasm is independent of microtubules. PMID- 10873644 TI - Efficiency of erythropoietin's signal peptide for HIV(MN)-1 gp 120 expression. AB - The HIV-1 gp120 gene with natural signal sequence expressed in eukaryotic expression systems showed extremely low levels of synthesis and secretion. Several expression systems have been used to improve the secretion levels of gp 120. In mammalian cells, the efficient expression of gp120 fused to t-PA signal peptide has been previously reported. Here, the effects of t-PA and EPO signal peptides were compared as secretion sequences for expression of gp120 in COS-7 cells. The EPO's signal peptide is used for the first time as leader sequence for secretion of foreign proteins. Our results indicated that higher amounts of secreted gp 120 were obtained when vectors containing EPO signal peptide were used. PMID- 10873646 TI - Quantification of platelet activation status by analyzing P-selectin expression. AB - Platelet activation status (PAS) is used for characterizing quality and function of platelets in various experimental and clinical settings. In this study, we created a set of platelet populations differing in PAS, using stimulation of platelets with thrombin in a wide range of concentrations, and analyzed a number of flow cytometric parameters, which characterize PAS by measuring P-selectin (CD62) expression. We found that PAS of a platelet population depends significantly on the specific parameters used for detecting CD62 expression and can differ several fold. We revealed the parameters which are more sensitive for distinguishing the differences between populations with similar low and similar high PAS. Selection of valid and sensitive flow cytometric parameters for PAS evaluation and distinguishing the differences between platelet populations with similar PAS can serve for diagnosis of platelet-associated disorders and monitoring their course and therapeutic interventions. PMID- 10873645 TI - Synergistic activation of UCP-3 expression in cultured fetal rat brown adipocytes by PPARalpha and PPARgamma ligands. AB - Rat brown adipocytes express mRNAs for Uncoupling Proteins (UCP) 1, 2 and 3 and the Peroxisome Proliferator Activated Receptors (PPAR) alpha and gamma. We have examined the effects of selective PPARalpha or -gamma activation on changes in UCP-1 and UCP-3 mRNA levels in cultured fetal rat brown adipocytes (FBA). Rosiglitazone (1.0 microM), a selective PPARgamma agonist, elicited 5- and 3-fold increases in UCP-1 and UCP-3, respectively. The PPARalpha ligand, Wy14643 (10.0 microM) increased UCP-3 tenfold, but decreased UCP-1. A synergistic effect on UCP 3 expression (30-fold increase; P < 0. 05) was observed when FBA were exposed to a combination of Wy14643 (10.0 microM) and rosiglitazone (10.0 microM). Thus, activation of PPARgamma increases UCP-1 and UCP-3 levels which are differentially regulated by PPARalpha. A synergistic interaction occurs between PPARalpha and PPARgamma in the regulation of UCP-3 in FBA, probably via co-activator recruitment, suppression of co-repressor proteins or through a direct interaction at the level of the PPRE. PMID- 10873647 TI - Molecular cloning and genomic organization of a second probable allatostatin receptor from Drosophila melanogaster. AB - We (C. Lenz et al. (2000) Biochem. Biophys. Res. Commun. 269, 91-96) and others (N. Birgul et al. (1999) EMBO J. 18, 5892-5900) have recently cloned a Drosophila receptor that was structurally related to the mammalian galanin receptors, but turned out to be a receptor for a Drosophila peptide belonging to the insect allatostatin neuropeptide family. In the present paper, we screened the Berkeley "Drosophila Genome Project" database with "electronic probes" corresponding to the conserved regions of the four rat (delta, kappa, mu, nociceptin/orphanin FQ) opioid receptors. This yielded alignment with a Drosophila genomic database clone that contained a DNA sequence coding for a protein having, again, structural similarities with the rat galanin receptors. Using PCR with primers coding for the presumed exons of this second Drosophila receptor gene, 5'- and 3'-RACE, and Drosophila cDNA as template, we subsequently cloned the cDNA of this receptor. The receptor cDNA codes for a protein that is strongly related to the first Drosophila receptor (60% amino acid sequence identity in the transmembrane region; 47% identity in the overall sequence) and that is, therefore, most likely to be a second Drosophila allatostatin receptor (named DAR-2). The DAR-2 gene has three introns and four exons. Two of these introns coincide with two introns in the first Drosophila receptor (DAR-1) gene, and have the same intron phasing, showing that the two receptor genes are clearly evolutionarily related. The DAR-2 gene is located at the right arm of the third chromosome, position 98 D-E. This is the first report on the existence of two different allatostatin receptors in an animal. PMID- 10873648 TI - Characterization of an ECF sigma factor protein from Pseudomonas aeruginosa. AB - The PvdS protein is essential for synthesis of the siderophore pyoverdine by Pseudomonas aeruginosa. PvdS has some sequence similarity to a family of alternative sigma factor proteins (the ECF [extracytoplasmic factor] family) that direct bacterial RNA polymerases to transcribe genes encoding extracytoplasmic factors. PvdS was purified as a His-tagged protein (hPvdS) and this was used to test the hypothesis that PvdS is a sigma factor protein. The purified protein caused core RNA polymerase from Escherichia coli to bind specifically to the promoters of pyoverdine synthesis genes and enabled transcription from these promoters in vitro. In addition, PvdS was found to co-purify with RNA polymerase from P. aeruginosa, indicating that PvdS associates with RNA polymerase inside the bacteria. These results show that PvdS is a sigma factor protein. PMID- 10873649 TI - Functional analysis of heme regulatory elements of the transcriptional activator Hap1. AB - Heme regulation of the activity of diverse proteins was thought to be mediated by heme-responsive motifs (HRMs). The yeast transcriptional activator Hap1 contains seven HRMs: HRM1-7. Three copies of a 17-amino-acid repeat are also located in the region encompassing HRM1 to -6. We examined the effects of these HRMs and repeats on heme regulation of Hap1 activity by deletion analysis and by Ala substitutions of key residues. We found that the effect of mutation or deletion of one HRM or 17-amino-acid repeat on Hap1 heme responsiveness is different from the effect of mutation or deletion of another HRM or repeat. Our data suggest that HRM7 plays a dominant role in mediating heme activation of Hap1 in heme sufficient cells while HRM1-6 may scavenge heme and cause a low level of Hap1 activation in heme-deficient cells. These results may help in understanding the roles of HRMs in other hemoproteins. PMID- 10873650 TI - PQBP-1/Npw38, a nuclear protein binding to the polyglutamine tract, interacts with U5-15kD/dim1p via the carboxyl-terminal domain. AB - PQBP-1 was identified as a binding protein to the polyglutamine tract present in various transcription-related factors and causative genes for neurodegenerative disorders. This novel gene contains at least two functional domains, WW domain and carboxyl-terminal domain (CTD), strictly conserved beyond species. Although human PQBP-1 additionally contains the polar amino acid-rich domain by which it binds to the polyglutamine tract, genuine physiological function(s) have not been clarified. In this study, we showed that U5-15kD, human homologue of fission yeast dim1p, is a partner molecule of PQBP-1 binding to CTD. This finding suggests physiological functions of PQBP-1 in splicing, cell cycle, and ubiquitination, through which we can speculate the pathological roles of PQBP-1 in triplet repeat diseases. PMID- 10873651 TI - Cloning of a coproporphyrinogen oxidase promoter regulatory element binding protein. AB - Coproporphyrinogen oxidase [CPO] gene promoter regulatory element (CPRE) plays an important role in CPO gene regulation. To isolate a CPRE binding protein, we performed Southwestern screening of K562 cDNA expression library using CPRE as a probe and isolated a cDNA clone which encoded a novel protein, Klp1 (K562 cell derived leucine-zipper-like protein 1). Klp1 mRNA was highly expressed in K562 cells, HeLa cells, and brain as a single transcript (1.4 kb). Gel mobility shift assays revealed that Klp1 specifically binds to CPRE. Computational analysis revealed that Klp1 has a leucine-zipper-like structure, a Leu-X-X-Leu-Leu motif, and a putative nuclear localization signal in the basic amino acid rich region. Transfection of the Klp1 expression vector into THP-1 cells resulted in transcriptional activation of a reporter construct containing CPRE. These results indicate that Klp1 is a DNA sequence-specific transcription factor that regulates gene expression of genes that contain CPRE in their regulatory region. PMID- 10873652 TI - Aging enhances the activation of the permeability transition pore in mitochondria. AB - Aging is associated with mitochondrial dysfunction in several tissues. However, it is not known how the observed mitochondrial dysfunction is related to aging associated tissue degeneration. We have shown previously that the activation of the permeability transition pore (PTP), which is believed to play a critical role in cell necrosis and apoptosis, is enhanced in spleen lymphocytes from old mice. Here we show that the threshold for calcium-induced, cyclosporin-sensitive, calcium release was significantly lower in isolated brain and liver mitochondria from aging mice. Thus, aging mice exhibit enhanced PTP activation in lymphocytes, brain, and liver. These results suggest that aging increases the susceptibility to calcium-dependent cell death (e.g., excitotoxicity, ischemia-reperfusion damage) in the brain, liver, and possibly other tissues. In addition, other pathways to apoptosis or necrosis that depend on PTP activation are also likely to be enhanced by aging. PMID- 10873653 TI - Disruption of actin cytoskeleton induces chondrogenesis of mesenchymal cells by activating protein kinase C-alpha signaling. AB - Disruption of actin cytoskeleton with cytochalasin D has been known to induce chondrogenic differentiation of chick embryo limb bud mesenchymal cells. However, the mechanism(s) for the induction of chondrogenesis by cytochalasin D is not yet clearly known. In the present study, we examined possible involvement of protein kinase C (PKC) and extracellular signal-regulated protein kinase (Erk-1) in chondrogenesis of mesenchymal cells induced by disruption of actin cytoskeleton. Disruption of actin cytoskeleton with cytochalasin D or latrunculin B induced chondrogenesis of mesenchymal cells cultured at subconfluent cell density, as determined by type II collagen expression. Among the expressed PKC isoforms, cytochalasin D dramatically increased expression and activation of PKCalpha in a dose-dependent manner, and inhibition or downregulation of PKCalpha blocked cytochalasin D-induced chondrogenesis. Cytochalasin D also downregulated Erk-1 phosphorylation that is associated with chondrogenesis. Our results, therefore, suggest that disruption of actin cytoskeleton induces chondrogenesis of mesenchymal cells by activating PKCalpha and by inhibiting Erk-1 signaling. PMID- 10873654 TI - Molecular basis for differing antineurogenic effects of GATA-1a and GATA-1b in Xenopus. AB - The erythroid transcription factor GATA-1 in Xenopus has been cloned as a pair of presumably duplicated genes designated as xGATA-1a and xGATA-1b. Although both xGATA-1a and xGATA-1b are able to stimulate erythropoiesis, only xGATA-1b is capable of inhibiting neurogenesis in Xenopus embryos. Chimeras of these two genes were constructed by permuting coding and untranslated regions (UTR) on both ends of these two xGATA-1, and their neurogenesis-inhibitory effects were studied. These results reveal that (1) sequence variations between the coding regions alone do not account for the neurogenesis effect; (2) 3' UTR of xGATA-1a causes the loss of the neurogenesis inhibition of xGATA-1b; (3) 3' UTR of xGATA 1b is essential to inhibit neurogenesis. In addition, the presence of either UTR does not affect the stability of the mRNA in vitro. These observations suggest the influence of 3' UTR in xGATA-1 on the inhibition of neurogenesis. PMID- 10873655 TI - Inefficient function of the signal sequence of PTHrP for targeting into the secretory pathway. AB - Parathyroid hormone-related peptide (PTHrP) is not only secreted out of cells, but also targeted to the nucleoli due to a nucleolar targeting signal (NTS). We assessed the molecular mechanism underlying the dual targeting of PTHrP by constructing a series of truncated forms of rat PTHrP cDNA and expressing them in CHO cells. Immunostaining was observed in both the Golgi apparatus and nucleoli in the same cell expressing PTHrP with the N-terminal full-length signal sequence. When PTHrP molecules were translated from CUGs downstream of the AUG initiator codon in the signal sequences, potential alternative initiators of the translation, they were exclusively localized in the nucleoli. In contrast, when a construct containing only the ATG-initiator codon was expressed, PTHrP was found to localize in both the nucleolus and the Golgi apparatus. No nucleolar staining of PTHrP was observed in the CHO cells transfected with PTH/PTHrP receptors even after incubating with a conditioned medium containing PTHrP, ruling out a possibility that PTHrP is, once secreted, internalized via receptor-mediated endocytosis and subsequently conveyed to nucleoli. Compatible with these morphological observations, a preproform of PTHrP was found in the cells expressing PTHrP in addition to proPTHrP, indicative of molecules along the secretory pathway. These results strongly indicate that the signal sequence of PTHrP is not sufficient to direct all the newly synthesized molecules across the endoplasmic reticulum, resulting in part of it being delivered to the nucleoli due to the NTS. PMID- 10873656 TI - Multilineage differentiation of Cbfa1-deficient calvarial cells in vitro. AB - We characterized calvaria-derived cells of Cbfa1-deficient mice to determine their stages of differentiation. In long-term culture, Cbfa1-deficient calvarial cells did not acquire osteoblastic phenotypes, but numerous adipocyte foci appeared with an increase in the expression of marker genes for adipocyte differentiation. In culture with BMP-2, Cbfa1-deficient calvarial cells still failed to generate bone nodules but differentiated into chondrocytes and further to terminal hypertrophic chondrocytes, and adipocyte foci were decreased. Cbfa1 deficient calvarial cells transplanted into the peritoneal cavity of athymic mice using BMP-2-coated diffusion chambers generated cartilage but not bone. These data indicate that Cbfa1-deficient calvarial cells completely lack the ability to differentiate into mature osteoblasts and Cbfa1 has an inhibitory function in adipocyte differentiation. As Cbfa1-deficient calvarial cells were enriched with immature mesenchymal cells, which can differentiate into adipocytes and chondrocytes, it is suggested that Cbfa1 plays an essential role in determining the lineage of multipotential mesenchymal precursor cells. PMID- 10873657 TI - Neovascularization with blood-brain barrier breakdown in delayed neuronal death. AB - Various kinds of acute pathological events in the central nervous system, such as ischemia, hemorrhage, and trauma, often cause brain edema. The edema may advance for days or weeks while inducing extensive damage in neural function, regardless of the extent of the original damage, and often results in death. Delayed edema is thought to be vasogenic; however, the mechanism underlying edema induction remains unknown. We found delayed vascular cell proliferation with a blood-brain barrier breakdown in and around the gerbil CA1 hippocampus, a region known to be involved in delayed apoptotic neuronal death 2-6 days after transient ischemia. Vascular cell proliferation, assessed by (3)H-thymidine incorporation, was most prominent 4-6 days after ischemia, and extravasation of exogenously applied dye or endogenous serum albumin from blood vessels was observed concomitantly. We propose neovascularization in delayed neuronal death as a cause of brain edema advancing days after neurological events. PMID- 10873658 TI - Molecular cloning and expression of the mouse N-acetylneuraminic acid 9-phosphate synthase which does not have deaminoneuraminic acid (KDN) 9-phosphate synthase activity. AB - A cDNA of the mouse homologue of Escherichia coli N-acetylneuraminic acid (Neu5Ac) synthase (neuB gene product) was cloned by the PCR-based method. The mouse homologue consists of 359 amino acids, and the cDNA sequence displays 33% identity to that of the E. coli Neu5Ac synthase. The recombinant mouse homologue which is transiently expressed in HeLa cells does not exhibit the Neu5Ac synthase activity, which catalyzes condensation of phosphoenolpyruvate (PEP) and N acetylmannosamine (ManNAc) to synthesize Neu5Ac, but the Neu5Ac 9-phosphate (Neu5Ac-9-P) synthase activity, which catalyzes condensation of PEP and ManNAc 6 phosphate (ManNAc-6-P) to synthesize Neu5Ac-9-P. Thus, the mouse homologue of E. coli Neu5Ac synthase is the Neu5Ac-9-P synthase. The Neu5Ac-9-P synthase is a cytosolic enzyme and ubiquitously distributed in mouse various tissues. Notably, the Neu5Ac-9-P synthase can not catalyze the synthesis of deaminoneuraminic acid (KDN) or KDN-9-P from PEP and Man or ManNAc-6-P, thus suggesting that the enzyme is not involved in the synthesis of KDN. This is consistent with the previous observation that only a very low activity to synthesize KDN is found in mouse B16 cells [Angata, T., et al. (1999) Biochem. Biophys. Res. Commun. 261, 326-331]. PMID- 10873659 TI - In vitro creation of amyloid fibrils from native and Arg124Cys mutated betaIGH3((110-131)) peptides, and its relevance for lattice corneal amyloid dystrophy type I. AB - BetaIGH3 protein has been recently involved in the pathogenesis of blinding corneal diseases, some of which have characteristic amyloid corneal deposits. The 124 codon of the betaig-h3 gene seems to be crucial for the amyloidogenicity of the protein product. We presently report an in vitro system that reproducibly forms amyloid fibrils from betaIGH3((110-131)) derived peptides. We also assessed the differences in fibril formation of two 22-amino acid peptides centered on the 124 residue: the native form and the Arg124Cys peptide (mutation linked to lattice corneal amyloid dystrophy type 1). After dialysis of Arg124Cys peptide against PBS 1/15 M pH 7.4 for 72 hours, Congo red staining and electron microscopy demonstrated the presence of abundant material fulfilling the criteria of amyloid. Quantitative analysis with thioflavine T fluorescence studies confirmed the high capacity of Arg124Cys peptide to form amyloid fibrils when compared to the native form. PMID- 10873660 TI - Promoter analysis and chromosomal mapping of human EBAG9 gene. AB - The human EBAG9 was previously identified as an estrogen responsive gene using CpG-genomic binding site cloning (Watanate et al., (1998) Mol. Cell. Biol. 18: 442-449). Recently it was revealed that the EBAG9 is identical with RCAS1 which is a cancer cell surface antigen implicated in immune escape. Here, we isolated and analyzed the 5'-flanking region of human EBAG9 gene. We determined transcription initiation site, which has a homology with an initiator element YYCAYYYY, and found that TATA motif was absent. Deletion analysis of the 5' flanking region using MCF-7 breast cancer cells indicated that the sequences -86 to -36 containing the ERE had the basal level of promoter activity and the upstream GC-rich region positively regulated the activity. EBAG9 promoter luciferase reporters containing the ERE could respond to estrogen, and electrophoretic mobility shift assay showed that ERalpha bound to the ERE. Moreover, fluorescent in situ hybridization analysis has shown that the human EBAG9 gene is located at chromosome 8q23 which is frequently amplified in tumors. These findings suggest that the human EBAG9 might be involved in carcinogenesis as an estrogen responsive gene. PMID- 10873662 TI - Systemic administration of HVJ viral coat-liposome complex containing human insulin vector decreases glucose level in diabetic mouse: A model of gene therapy. AB - In this study, we examined the feasibility of a systemic administration of HVJ liposome complex containing human insulin construct into the blood in mice via the tail vein. Transfection of human insulin vector resulted in a transient decrease in serum glucose in streptozotocin (SZT)-induced diabetic mice, accompanied by the detection of human insulin in the liver and spleen. In accordance with the decreased glucose, plasma immunoreactive insulin could be detected up to 14 days after a single transfection in mice transfected with insulin vector. Repeated intravenous injection of human insulin vector every week resulted in a sustained decrease in serum glucose over a 4-week period, accompanied by the detection of C-peptide fragments and a significant decrease in BUN and creatinine. Here, we demonstrated the feasibility of intravenous systemic administration of an insulin vector that results in a sustained improvement of diabetic glucose metabolism. PMID- 10873661 TI - Specificity and efficiency of Cre-mediated recombination in Emx1-Cre knock-in mice. AB - Emx1 is a mouse homologue of the Drosophila homeobox gene empty spiracles and its expression is restricted to the neurons in the developing and adult cerebral cortex and hippocampus. We reported previously the creation of a line of transgenic mice in which the cre gene was placed directly downstream of the putative Emx1 promoter using ES cell technology. We showed that Cre protein was present in the cerebral cortex of the transgenic mice and was able to mediate loxP-specific recombination in vitro. In the present study, the specificity and efficiency of the cre-mediated recombination were determined using three independent lines of reporter mice and a combination of histochemical staining, neuronal culture, and Southern detection of the genomic DNA. Our results showed that the recombination was highly efficient in all three lines of reporter mice tested and confirmed that the deletion was restricted to the neurons in the cerebral cortex and hippocampus. Furthermore, we have determined that the recombination efficiency in the cerebral cortex was 91%. Our results suggest that Emx1 is not expressed in every neuron in the developing and adult cerebral cortex. This line of cre mice should contribute to the studies of cortical development and plasticity. PMID- 10873663 TI - Vitamin D induced up-regulation of keratinocyte growth factor (FGF-7/KGF) in MCF 7 human breast cancer cells. AB - Keratinocyte growth factor (FGF-7/KGF) is a secreted member of the fibroblast growth factor family, which functions primarily as an important paracrine mediator of cell growth and differentiation. Inhibitory pathways of vitamin D may also involve participation of some growth factors. To determine whether vitamin D may play a role in the expression of FGF-7, we investigated FGF-7 expression in human breast cancer cells treated with 1,25-dihydroxyvitamin D3, which inhibited the growth of the cells. By means of cDNA microarray, RT-PCR, and Western blot analysis, we have shown an increase in expression of FGF-7 on both mRNA and protein levels after vitamin D exposure. This is the first demonstration of vitamin D regulation of FGF-7 expression and its possible involvement in mediating growth and differentiation by vitamin D. PMID- 10873664 TI - Enantiospecific recognition of DNA sequences by a proflavine Troger base. AB - The DNA interaction of a chiral Troger base derived from proflavine was investigated by DNA melting temperature measurements and complementary biochemical assays. DNase I footprinting experiments demonstrate that the binding of the proflavine-based Troger base is both enantio- and sequence-specific. The (+)-isomer poorly interacts with DNA in a non-sequence-selective fashion. In sharp contrast, the corresponding (-)-isomer recognizes preferentially certain DNA sequences containing both A. T and G. C base pairs, such as the motifs 5' GTT. AAC and 5'-ATGA. TCAT. This is the first experimental demonstration that acridine-type Troger bases can be used for enantiospecific recognition of DNA sequences. PMID- 10873665 TI - The mouse ZFH-4 protein contains four homeodomains and twenty-two zinc fingers. AB - We isolated the mouse zfh-4 cDNA which is 12 kb long and capable of encoding a 3,550-amino acid protein containing four homeodomains and 22 zinc fingers including two pseudo zinc finger motifs. The mouse ZFH-4 is 51% homologous to the mouse ATBF1 and 23% to the Drosophila ZFH-2. The homeodomain and zinc finger regions are highly conserved between ZFH-4 and ATBF1 except that one zinc finger is missing in ZFH-4. Analysis of partial genomic sequences showed that the mouse zfh-4 and ATBF1 genes are similar in exon-intron organization. RT-PCR analysis of zfh-4 transcripts in adult mouse tissues showed that zfh-4 expression was low but reproducibly detectable in brain, heart, lung and muscle. In these mouse tissues, ATBF1 transcripts were poorly amplified by PCR under the conditions where zfh-4 transcripts were amplified, suggesting that the expression of zfh-4 mRNA is higher than that of ATBF1 mRNA. Other comparative analysis suggests functional similarities and dissimilarities between ZFH-4 and ATBF1. PMID- 10873666 TI - 6,3'-dibromoflavone and 6-nitro-3'-bromoflavone: new additions to the 6,3' disubstituted flavone family of high-affinity ligands of the brain benzodiazepine binding site with agonistic properties. AB - 6,3'-dibromoflavone and 6-nitro-3'-bromoflavone inhibited [(3)H]flunitrazepam binding to the benzodiazepine binding site of the gamma amino butyric acid receptor complex with K(i) values between 17 and 36 nM in different brain regions. Their gamma amino butyric acid ratio for [(3)H]flunitrazepam binding to cerebral cortex membranes indicated partial agonistic properties. Both compounds had similar pharmacological effects: they produced anxiolytic-like effects at low doses but did not alter locomotor activity or muscle tonicity; sedation was caused only at doses higher than 30 mg/kg in mice. These synthetic flavone derivatives join an existing family of 6,3'-disubstituted flavone compounds with high affinity for the benzodiazepine binding site and partial agonistic profiles. PMID- 10873668 TI - Expression and binding properties of a soluble chimeric protein containing the N terminal domain of the Duffy antigen. AB - The blood group Duffy antigen of human erythrocytes, which exists in two allelic forms, Fy(a) and Fy(b), is a promiscuous chemokine receptor. In this report we describe the expression and purification of a chimeric protein composed of the amino-terminal extracellular domain of the Duffy antigen (aa 3-60), C-terminal intracellular fragment of glycophorin A (GPA, aa 104-131), and the hexahistydyl tag. We obtained two forms of the recombinant protein containing the Fy(a) or Fy(b) epitope, denoted Fy(a)/GPA and Fy(b)/GPA, respectively. These constructs were expressed in Escherichia coli as periplasmic proteins and were purified by affinity chromatography on the Ni-NTA-agarose. Both proteins bound the monoclonal antibodies recognizing the common Fy6 epitope of the Duffy antigen and an epitope of the C-terminal fragment of GPA, and only the Fy(a)/GPA bound anti-Fy(a) antibody. However, binding of IL-8 to the recombinant proteins was not detected, which indicated that an N-terminal domain of the Duffy antigen is not sufficient for an effective chemokine binding. The lack of the chemokine binding was not likely to be due to the lack of glycosylation of the Fy/GPA, since IL-8 was effectively bound to de-N-glycosylated erythrocytes. PMID- 10873667 TI - COX-2 inhibition prevents insulin-dependent diabetes in low-dose streptozotocin treated mice. AB - Insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease believed to be caused by an inflammatory process in the pancreas leading to selective destruction of the beta cells. Inducible cyclooxygenase (COX-2) is expressed under inflammatory conditions and its product prostaglandin E(2) (PGE(2)) is an important inflammation mediator. We report here that administration of the selective COX-2 inhibitor NS-398 prevents the onset of diabetes in mice brought on by multiple low-doses of streptozotocin (STZ). Histological observations indicated that STZ-mediated destruction of beta cells was prevented by NS-398 treatment. Delayed (day 3) administration of NS-398 was also protective in this model. No protective effect was observed when NS-398 was administered prior to a high, toxic dose of STZ. These results demonstrate the critical importance of COX 2 activity in autoimmune destruction of beta cells, and point to the fact that COX-2 inhibition can potentially develop into a preventive therapy against IDDM. PMID- 10873669 TI - Identification of a novel beta-catenin-interacting protein. AB - Cadherin is a well-known cell-cell adhesion molecule, and it binds to beta catenin, which in turn binds to alpha-catenin. However, little is known about the regulatory mechanism underlying the cadherin-mediated cell-cell adhesion. Here we purified two novel beta-catenin-interacting proteins with molecular masses of 180 kDa (p180) and 150 kDa (p150) from bovine brain cytosol by using glutathione S transferase (GST)-beta-catenin affinity column chromatography. Mass spectral analysis revealed p180 to be identical to KIAA0313 which has a putative Rap1 guanine nucleotide exchange factor (GEF) domain and p150 to be the same as KIAA0705 which has a high degree of sequence similarity to the synaptic scaffolding molecule (S-SCAM), which binds beta-catenin and KIAA0313 in the yeast two-hybrid system and overlay assay, respectively (Ide et al., Biochem. Biophys. Res. Commun. 256, 456-461, 1999; Ohtsuka et al., Biochem. Biophys. Res. Commun. 265, 38-44, 1999). beta-Catenin was coimmunoprecipitated with KIAA0313 in Madin Darby canine kidney II (MDCKII) cells, bovine brain cytosol, and EL cells. KIAA0313 and beta-catenin were partly colocalized at sites of cell-cell contact in MDCKII cells. Taken together, our data suggest that KIAA0313 associates with beta-catenin through KIAA0705 in vivo at sites of cell-cell contact. PMID- 10873670 TI - Stress-activated protein kinase-dependent induction of c-fos by Cd(2+) is mediated by MKK7. AB - Exposure of mesangial cells to ionic Cd(2+) induces the proto-oncogene c-fos, while activating both Erk and stress-activated protein kinase (SAPK) MAP kinase pathways. While we have previously used a pharmacological inhibitor of Erk activation to implicate involvement of this pathway in the induction of c-fos by Cd(2+), the consequences of SAPK activation remained unknown. Here we use dominant negative inhibitors of the SAPK kinases, SEK1 and MKK7, to show that Cd(2+) activates SAPK through MKK7, but that partial inhibition of SAPK alone is insufficient to significantly affect the magnitude of the Cd(2+)-dependent increase in c-fos mRNA. However, inhibition of Erk and SAPK pathways together abrogates the increase, suggesting that these pathways act in concert in the induction of c-fos by this toxic metal. PMID- 10873671 TI - Reciprocal expression of infant- and adult-preferring transcripts of CDCrel-1 septin gene in the rat neocortex. AB - We report here the isolation and characterization of cDNA clones for a novel isoform of CDCrel-1 septin, termed CDCrel-1A, with a different 5' end sequence from the transcripts encoding the known CDCrel-1 (designated as CDCrel-1F) in the developing rat neocortex. Alternative polyadenylation site selections resulted in various transcripts for CDCrel-1A including the fusion forms with another gene, platelet glycoprotein Ibbeta (GPIbbeta). Expression of the distinct transcripts encoding CDCrel-1A and CDCrel-1F increased and decreased, respectively, from the infant to adult period. Therefore CDCrel-1A might be a major form of the CDCrel-1 septin in the adult neocortex of mammals. PMID- 10873672 TI - The effects of vitamin C supplementation on protein oxidation in healthy volunteers. AB - We have investigated vitamin C supplementation effects on immunoglobulin oxidation (carbonyls) and total plasma protein sulfhydryls in healthy human volunteers. After receiving placebo, plasma ascorbate and oxidation markers were unchanged. Following 5 weeks supplementation with vitamin C (400 mg/day), plasma ascorbate increased but no significant effect on protein oxidation was observed. At 10 and 15 weeks supplementation, carbonyl levels were significantly reduced (P < 0.01) in subjects with low baseline ascorbate (29.51 +/- 5.3 microM) but not in those with normal baseline ascorbate (51.81 +/- 2.3 microM). To eliminate any effect from seasonal variation in dietary antioxidant intake, a second phase was undertaken. Subjects on vitamin C for 15 weeks were randomly assigned to receive either placebo or vitamin C. No difference in plasma sulfhydryl content was observed. Subjects withdrawn from supplementation showed an increase in immunoglobulin carbonyl content (P < 0.01). This demonstrates that dietary vitamin C supplementation can reduce certain types of oxidative protein damage in subjects with low basal antioxidant. PMID- 10873673 TI - Extracellular regulated kinase, but not protein kinase C, is an antiapoptotic signal of insulin-like growth factor-1 on cultured cardiac myocytes. AB - This study aims to elucidate the signaling pathway for insulin-like growth factor 1 (IGF-1) in cultured neonatal rat cardiomyocytes and particularly the role of IGF-1 in cardiac apoptosis. IGF-1 stimulated polyphosphoinositide turnover, translocation of protein kinase C (PKC) isoforms (alpha, epsilon, and delta) from the soluble to the particulate fraction, activation of phospholipid-dependent and Ca(2+)-, phospholipid-dependent PKC, and activation of the extracellular regulated kinase (ERK). IGF-1 attenuated sorbitol-induced cardiomyocyte viability and nuclear DNA fragmentation. These antiapoptotic effects of IGF-1 were blocked by PD-098059 (an MEK inhibitor) but not by bisindolylmaleimide I (BIM, a specific PKC inhibitor). The ERK pathway may therefore be an important component in the mechanism whereby IGF-1 exerts its antiapoptotic effect on the cardiomyocyte. PMID- 10873674 TI - Involvement of nitric oxide in endothelium-dependent arterial relaxation by leptin. AB - Leptin is a polypeptide, mainly produced in white adipose tissue, and increases sympathetic nerve activity. A few studies investigated leptin's effect on peripheral vessels. We examined the vasorelaxant effects of human leptin on rat arteries. Arterial rings were precontracted with 1 x 10(-6) mol/l of phenylephrine, and leptin was superfused. Leptin relaxed phenylephrine precontracted arterial rings in a dose-dependent manner. ED50 was calculated to 8.4 microg/ml. Removal of endothelium abolished the effects of leptin. Indomethacin (1 x 10(-5) mol/l) did not affect the vasorelaxation by leptin, whereas 1 x 10(-4) mol/l of N(omega)-nitro-L-arginine methyl ester (L-NAME) completely suppressed it. The inhibition was antagonized by 1 x 10(-4) mol/l of L arginine. Leptin normally relaxed arterial rings during superfusion of K channel blockers, including 3 x 10(-5) mol/l of glibenclamide, 1 x 10(-6) of mol/l apamin, and 5 x 10(-7) mol/l of charybdotoxin. Low Cl(-) solution (8. 3 mmol/l) inhibited leptin-induced relaxation, but endothelium-independent vasodilatation by nitroprusside was not impaired at low Cl(-) solution. These results suggest that arterial relaxation by leptin is mediated by nitric oxide released from endothelium, and Cl(-) plays an important role in leptin-induced nitric oxide release. PMID- 10873675 TI - Optical biosensor study of ternary complex formation in a cytochrome P4502B4 system. AB - The optical biosensor method was used for the revelation of ternary complexes, formed by the full-length NADPH-cytochrome P450 reductase (d-Fp) and cytochromes P4502B4 (d-2B4) and b5 (d-b5) in the course of their interactions within the reconstituted d-2B4-containing system. Based on the lack of competition between d b5 and d-Fp for the binding sites on immobilized 2B4 (3) as well as on the analysis of data obtained in the three proteins' dissociation reactions, the possibility of formation of ternary complexes through the interactions between membranous hydrophobic fragments of proteins was substantiated. All the complexes obtained were productive. PMID- 10873676 TI - Cloning and characterization of the NADH pyrophosphatases from Caenorhabditis elegans and Saccharomyces cerevisiae, members of a Nudix hydrolase subfamily. AB - Two genes from Caenorhabditis elegans and Saccharomyces cerevisiae, coding for enzymes homologous to the Nudix hydrolase family of nucleotide pyrophosphatases, have been cloned and expressed in Escherichia coli. The purified enzymes are homodimers of 39.1 and 43. 5 kDa, respectively, are activated by Mg(2+) and Mn(2+), and are 30 to 50 times more active on NADH than on NAD(+). They both have a conserved array of amino acids downstream of the Nudix box first seen in the orthologous enzyme from E. coli which designates them as members of an NADH pyrophosphatase subfamily of the Nudix hydrolases. PMID- 10873677 TI - Cloning and expression of mouse deafness dystonia peptide 1 cDNA. AB - Complementary DNA of mouse deafness dystonia peptide 1 (DDP1) was isolated from adipocyte cDNA library and expressed in mammalian cells. The sequence shares homology of 92 and 97% on the nucleic acid and the amino acid levels with human DDP1. In comparison to mouse Bruton's tyrosine kinase (Btk) locus, the coding region spans 2 exons and the splice point is the same as human DDP1. Northern blot analysis suggests that mouse DDP1 expresses ubiquitously. In vitro transcription/translation study showed that the cDNA of mouse DDP1 codes about 11 kDa peptide. DDP1 tagged with FLAG localized in mitochondria and cytoplasm of COS7 cells. P19 embryonal carcinoma cells transfected with anti sense DDP1 cDNA were frequently dead after subculture and all the survivals expressed endogenous DDP1 mRNA. Therefore, mouse DDP1 may play an important role to survive in contrast to Tim8p, a yeast homologue, which was unnecessary in yeast. PMID- 10873678 TI - Polymorphisms in FcepsilonRI beta chain do not affect IgE-mediated mast cell activation. AB - Genetic polyymorphisms that result in three amino acid changes in FcepsilonRI beta chain (Ile(181)-->Leu, Val(183)-->Leu, and Glu(237)-->Gly) have been identified as candidates that associate with allergic disorders such as atopy and asthma. To elucidate the biological significance of these polymorphisms in regulating the expression and function of FcepsilonRI, we generated four types of transfectants that express wild-type or mutant mouse beta chains corresponding to these human variants by retrovirus-mediated gene transfer into beta chain deficient mouse-derived mast cells. No significant functional differences between the wild-type beta chain transfectant and any of the mutant beta chain transfectants were observed in beta-hexosaminidase release, intracellular calcium mobilization, or cytokine and leukotriene C(4) production in response to FcepsilonRI crosslinking. Our results suggest that these polymorphisms in FcepsilonRI beta chain do not affect FcepsilonRI-mediated mast cell activation at least in our mouse in vitro system. PMID- 10873679 TI - Inhibition of cyclooxygenase-2 improves cardiac function in myocardial infarction. AB - Induction of cyclooxygenase-2 (COX-2) in ischemic myocardium is thought to increase the production of proinflammatory prostanoids and contribute significantly to the ischemic inflammation. Left ventricular myocardial infarction (MI) was created by ligating the left coronary artery in Lewis rats. Hemodynamic measurements at 4 weeks showed better cardiac function in the group treated with a selective COX-2 inhibitor (DFU; 5 mg/kg/day) for 2 weeks after induction of MI compared to the vehicle treated group. These results suggest that induction of COX-2 contributes to myocardial dysfunction, and that selective inhibition of COX-2 could constitute an important therapeutic target for the treatment of MI. PMID- 10873680 TI - The structure of the human LIM protein ACT gene and its expression in tumor cell lines. AB - We describe the human ACT genomic and cDNA sequence which like its murine counterpart contains the defining secondary structure of the FHL (Four-and-a-Half LIM-domain) LIM-protein family. The coding region of the human ACT gene spans five exons. This distribution is very similar to the FHL1 gene and includes the arrangement of split codons across exon boundaries suggesting that these genes share a common ancestor. The human ACT gene was not detected by Northern analysis in the adult testis although this is the only known site of expression found with its murine counterpart. However, the human ACT gene was found to be expressed in a panel of human tumor cell lines derived from squamous cell carcinomas, melanomas, and leukemias. Interestingly, FHL1, FHL2, and FHL3 were also found to be expressed in some of these cell lines and the results suggest an important role for FHLs in tumor biology. PMID- 10873681 TI - Epolactaene, a novel neuritogenic compound in human neuroblastoma cells, selectively inhibits the activities of mammalian DNA polymerases and human DNA topoisomerase II. AB - We chemically synthesized epolactaene, a neuritogenic compound in human neuroblastoma cells, and investigated its biochemical action in vitro. Epolactaene and its derivatives selectively inhibited the activities of mammalian DNA polymerase alpha and beta and human DNA topoisomerase II, with IC(50) values of 25, 94, and 10 microM, respectively. By comparison with its structural derivatives, the long alkyl side chain in epolactaene seemed to have an important role in this inhibitory effect. The compound did not influence the activities of plant or prokaryotic DNA polymerases or of other DNA metabolic enzymes such as telomerase, RNA polymerase, and deoxyribonuclease I. Epolactaene did not intercalate into DNA. These results suggested that the neuritogenic compound epolactaene influences both DNA polymerases and topoisomerase II despite the dissimilarity in both structure and properties of these two enzymes and that inhibition of these enzymes could be related to the neuritogenic effect in human neuroblastoma cells. The relationship between the neuritogenic mechanism and cell cycle regulation by epolactaene was also discussed. PMID- 10873682 TI - The deuterium isotope effect on the NMR signal of the low-barrier hydrogen bond in a transition-state analog complex of chymotrypsin. AB - The participation of a low-barrier hydrogen bond (LBHB) in the mechanism of action of chymotrypsin introduces a new role for Asp 102 and His 57 in catalysis [C. S. Cassidy, J. Lin, and P. A. Frey (1997) Biochemistry 36, 4576-4584]. It is postulated that the LBHB increases the basicity of His 57-N(epsilon2) in the transition state, thereby facilitating the abstraction of a proton from Ser 195, and stabilizes the tetrahedral intermediate in the acylation step. Evidence for this mechanism includes the downfield chemical shift of the proton bridging His 57 and Asp 102 in transition-state analog complexes and the low deuterium fractionation factors for this proton in the same complexes. We present additional spectroscopic evidence supporting the assignment of an LBHB between His 57 and Asp 102. The tetrahedral addition complex between Ser 195 of chymotrypsin and N-acetyl-l-leucyl-l-phenylalanyl trifluoromethylketone is regarded as a close structural analog of a tetrahedral intermediate. The deuterium NMR signal for the downfield deuteron bridging His 57 and Asp 102 in D(2)O has now been observed as a broad band centered at 17.8 +/- 0.5 ppm. The proton NMR signal in H(2)O is centered at 18.9 +/- 0.05 ppm. The two signals are clearly separated corresponding to a deuterium isotope effect of Delta[delta(H) - delta(D)] = 1.1 +/- 0.5 ppm. Deuterium isotope effects in this range are characteristic of LBHBs, and this observation provides further support for the assignment of the proton bridging His 57 and Asp 102 in transition-state analog complexes as an LBHB. PMID- 10873683 TI - Phylogenetic classification of prokaryotic and eukaryotic Sir2-like proteins. AB - Sirtuins (Sir2-like proteins) are present in prokaryotes and eukaryotes. Here, two new human sirtuins (SIRT6 and SIRT7) are found to be similar to a particular subset of insect, nematode, plant, and protozoan sirtuins. Molecular phylogenetic analysis of 60 sirtuin conserved core domain sequences from a diverse array of organisms (including archaeans, bacteria, yeasts, plants, protozoans, and metazoans) shows that eukaryotic Sir2-like proteins group into four main branches designated here as classes I-IV. Prokaryotic sirtuins include members of classes II and III. A fifth class of sirtuin is present in gram positive bacteria and Thermotoga maritima. Saccharomyces cerevisiae has five class I sirtuins. Caenorhabditis elegans and Drosophila melanogaster have sirtuin genes from classes I, II, and IV. The seven human sirtuin genes include all four classes: SIRT1, SIRT2, and SIRT3 are class I, SIRT4 is class II, SIRT5 is class III, and SIRT6 and SIRT7 are class IV. PMID- 10873684 TI - Impact factors or common sense. PMID- 10873685 TI - Chest drains: does size matter? PMID- 10873686 TI - Asbestos-related benign pleural disease. AB - Benign pleural disease is the commonest manifestation of asbestos exposure encountered by radiologists. Benign pleural thickening can appear as circumscribed parietal pleural plaques or as more diffuse thickening of the visceral pleura. Benign-asbestos induced pleural effusions are a significant and under-recognized manifestation of asbestos exposure with important sequelae, such as diffuse pleural thickening which may be associated with functional impairment and for which compensation may be sought. This review concentrates on the strengths and weaknesses of chest radiography and computed tomography for the detection and characterization of benign asbestos-related pleural disease and the relevance of imaging abnormalities to compensation and functional impairment.Peacock, C. (2000). Clinical Radiology55, 422-432. PMID- 10873687 TI - Ultrasound. PMID- 10873688 TI - Osteoma of the inner table of the skull--CT diagnosis. AB - AIM: The purpose of this study was to ascertain CT criteria for the differentiation of osteoma of the internal table of the skull (OIT) from meningioma. MATERIAL AND METHODS: Thirty-eight patients with proven OIT by operation or by post-mortem examination and 100 patients with proven meningioma were studied. RESULTS: Unenhanced computed tomography (UCT) and enhanced CT (ECT) was performed in each case and images on brain and bone window settings were evaluated. Measurements were taken of the widest and narrowest diameters of the OIT on the bone window images. The OIT appeared as an ivory-density, mushroom like mass with well-defined borders attached to the inner table of the skull by a bony stalk or neck. The ratio between the widest diameter of the mass and the narrowest area of the stalk or neck ranged from 1.6 to 6. The CT features of 100 meningiomas were not consistent with OIT, using the following parameters: contrast enhancement, surrounding vasogenic oedema, dural lucent line, osteolytic bone lesion and cystic component. Hard meningiomas without contrast enhancement did not present with a bony stalk or neck. No meningioma had the CT features of OIT, and no OIT had the CT features of meningioma. No patient operated on for OIT showed any clinical improvement following surgery. CONCLUSIONS: Computed tomography examination can be sufficient for the diagnosis of OIT and may therefore prevent unnecessary surgery.Avrahami, E., Even, I. (2000). Clinical Radiology55, 435-438. PMID- 10873689 TI - Magnetic resonance imaging of spinal plasmacytoma. AB - AIM: To describe the magnetic resonance imaging (MRI) features of spinal plasmacytoma. MATERIALS AND METHODS: The clinical records and MRI findings in six patients (five men, one woman; age range 41-61 years) with histologically proven plasmacytoma of the spine were reviewed. All studies included sagittal T1- and T2 weighted spin-echo sequences and axial T1-weighted spin-echo sequences. Intravenous gadolinium DTPA was administered in four cases. RESULTS: MRI showed a characteristic appearances in four cases of low signal intensity curvilinear areas within the vertebra or cortical irregularity. CONCLUSION: Recognition of these imaging features can initiate the appropriate investigation as the commonest differential diagnosis for such lesions is metastasis.Shah, B. K. (2000). Clinical Radiology55, 439-445. PMID- 10873690 TI - Early report: comparison of breath-hold MR excretory urography, Doppler ultrasound and isotope renography in evaluation of symptomatic hydronephrosis in pregnancy. AB - AIM: To compare assessment by MR excretory urography (MREU), Doppler ultrasound and isotope renography of women with symptomatic hydronephrosis in pregnancy and to define its cause. MATERIALS AND METHODS: Eleven women at 19-34 weeks of gestation were studied prospectively with gadolinium-enhanced breath-hold gradient echo MREU and transabdominal Doppler ultrasound compared with a 'gold standard' of isotope renography employing frusemide challenge. All studies were performed within 24 h, were reported independently in a blinded fashion and employed clearly defined criteria. Obstetric and infant outcomes were obtained. RESULTS: There were no adverse reactions to gadolinium administration in pregnancy and no adverse obstetric or infant outcomes. Three of the 11 women were unable to tolerate the complete MREU protocol. Ultrasound indices could not be used to predict ureteric obstruction as shown by isotope renography. MREU agreed with renographic findings in five of the six cases with obstruction and in two without obstruction. MREU directly demonstrated hydronephrosis to result from extrinsic compression of the ureter between the gravid uterus and iliopsoas muscle. CONCLUSION: MR excretory urography is a promising technique which affords equivalent functional and additional anatomical information to isotope renography. It is more accurate than Doppler ultrasound in the assessment of ureteric obstruction in pregnancy.Spencer, J. A. (2000). Clinical Radiology55, 446-453. PMID- 10873691 TI - An investigation into why two-view mammography is better than one-view in breast cancer screening. AB - AIM: To determine why two-view mammography in screening for breast cancer is more effective than using a single medio-lateral oblique view. MATERIALS AND METHODS: In the United Kingdom Coordinating Committee on Cancer Research randomized trial of one- vs two-view mammography in breast cancer screening the oblique view was assessed by one radiologist and two views (oblique and cranio-caudal) assessed by another. For the present study the mammographic films were retrieved from the screening centres and assessed by three consultant radiologists. Mammographic films were available from 110 women; 87 had their breast cancer detected by both one and two views and in 23 it was missed by one view but detected using two views. Outcome measures were breast size, location and size of the cancer, mammographic features, presence of microcalcification and overall radiological assessment. RESULTS: Although 23 cancers were missed in the original trial when one view was used, only two were not visible on the oblique view. Cancers missed using a single oblique view (and only detected if the cranio-caudal view was available with the oblique) tended to be smaller by about 4 mm (P = 0.05), centrally located in the breast (P = 0.16), not spiculated or round, (P 70% internal carotid artery stenosis, underwent inverted surgical timing (aortic reconstruction first and carotid endarterectomy second). Preoperative Transcranial Doppler (TCD) with and without acetazolamide was used to evaluate cerebrovascular reserve capacity (CRC). Intraoperatively, middle cerebral artery flow velocity (mean MCAv) and systemic blood pressure (SBP) were recorded. RESULTS: preoperatively, all 16 patients had good CRC (increase in mean MCAv: 66% right and 72% left). Intraoperatively, the mean MCAv (from 49+/-13 to 45+/-14 cm/s p=0.0249) and SBP decreased (from 127+/-25 to 113+/ 22 mmHg p=0.0016). In patients with unilateral carotid disease, declamping had no effect on left mean MCAv despite a significant decrease of SBP (129+/-44 to 113+/ 21 mmHg p=0.0211). In those with bilateral disease, declamping decreased both mean MCAv: from (48+/-12 to 39+/-10 cm/s p=0.011) and SBP (123+/-26 to 111+/-25 mmHg p=0.0479). No perioperative neurological deficit occurred. CONCLUSIONS: if CRC is normal or still effective, aortoiliac reconstruction does not impair cerebral perfusion. PMID- 10873724 TI - Outcome in patients with symptomatic occlusion of the internal carotid artery. AB - OBJECTIVES: to assess whether the risk of recurrent ischaemic stroke in patients with symptomatic internal carotid artery (ICA) occlusion has changed over the past decades, to determine risk factors for the occurrence of ischaemic stroke and to assess the risk of endarterectomy (CEA) of a severe contralateral ICA stenosis. DESIGN: retrospective cohort study. PATIENTS AND METHODS: patients with symptomatic ICA occlusion were identified from duplex registry files between 1991 and 1995. Information was obtained on vascular risk factors, performance of CEA for a contralateral ICA stenosis and on recurrence of ischaemic stroke. The rate of complications occurring within 30 days after CEA of the contralateral ICA in patients with symptomatic ICA occlusion was compared with the risk of CEA in patients with asymptomatic ICA occlusion and severe contralateral ICA stenosis (symptomatic or asymptomatic). RESULTS: ninety-seven patients were identified. Mean follow-up time was 26 months. The annual risk of (non-)fatal stroke was 5.3% for all strokes (95% CI 2. 9%-9.6%) and 3.8% for ipsilateral stroke (95% CI 1.9% 7.7%). Hyperlipidaemia and severe stenosis of the contralateral ICA were independent risk factors. Twenty-two of 32 patients with a severe stenosis of the contralateral ICA underwent CEA, of which one patient died and three suffered a minor ischaemic stroke. The perioperative risk of CEA in the control group of 20 patients with asymptomatic contralateral ICA occlusion was 0% (0 of 20). CONCLUSIONS: outcome in patients with symptomatic ICA occlusion has not substantially improved over the years. CEA for severe stenosis of the contralateral ICA carried a relatively high risk in our series, but deserves to be studied in a controlled design. PMID- 10873725 TI - Spinal cord stimulation in diabetic lower limb critical ischaemia: transcutaneous oxygen measurement as predictor for treatment success. AB - OBJECTIVES: to evaluate whether transcutaneous oxygen tension (TcpO(2)) measurements could be used as a specific prognostic parameter in selecting diabetic patients for permanent device implantation. METHODS: sixty consecutive diabetic patients (28 with autonomic neuropathy), classified as Fontaine stage III or IV, underwent spinal cord stimulation (SCS) for ischaemic pain, after failed conservative or surgical treatment. Pedal TcpO(2)on the dorsum of the foot and ankle-pressure Doppler measurements were performed before, and 2 and 4 weeks after implantation. RESULTS: limb salvage and good pain relief were achieved in 35 patients, while in 12 partial pain relief and limb salvage for at least 6 months were obtained. In 13 patients the method failed and the ischaemic limbs were amputated. Only 3 of the 28 patients with neuropathy had any long-term benefit. Limb salvage was achieved in those patients with a significant increase in TcpO(2)within 2 weeks of stimulation. The stage of the neuropathy was inversely related to the success of SCS therapy. The ankle-brachial pressure index (ABPI) did not change after stimulation. CONCLUSIONS: diabetic patients with significant increase of TcpO(2)and pain relief during a 2-week test period may be successfully treated by long-term SCS unless they have advanced autonomic neuropathy. PMID- 10873726 TI - Optimising the performance of intermittent pneumatic compression devices. AB - OBJECTIVES: this study was designed to determine whether an intermittent pneumatic compression device (IPC) with an increased maximal inflation pressure, a decreased time to maximal pressure and a longer duration of compression would improve venous return compared to a standard IPC device. METHODS: thirty limbs in 15 volunteers without evidence of venous disease were studied using duplex scanning at rest and during the application of two different IPC devices with different compression parameters. The first device IPC-1 (SCD 5325, Kendall) has a six-chambered cuff applying 45 mmHg after 12 s, sequentially from ankle to thigh followed by 60 s of non-compression. The second device IPC-2 (Vena Assist(R), ACI Medical) has a foot, ankle and calf cuff, applies a pressure of 80 mmHg, has a pressure rise time of 0.3 s, maintains inflation for 5.5 s, and has a cycling time of 1 min. Peak venous velocity and acceleration time were measured at rest and during the IPC application. Measurements were obtained in supine position from the common femoral vein 1 cm above the saphenofemoral junction to include the entire venous outflow from the limb. RESULTS: peak venous velocity at rest was significantly higher in the right limb than in the left limb (26+/-7.2 vs. 22+/-5.7 cm/s, p<0.01). Peak venous velocity was significantly increased by both IPC devices (p <0.0001). IPC-2 achieved significantly higher peak venous velocity than IPC-1 (55.1+/-17.8 vs. 37.4+/-6.9 cm/s, p<0.0001). Acceleration time was also found to be significantly shorter (370+/-93.4 vs. 560+/-83.5 ms, p<0.0001) in IPC-2 than in IPC-1, respectively. CONCLUSIONS: we have demonstrated that progressive inflation at the foot, ankle and calf, increasing maximal inflation pressure and decreasing time to maximal pressure result in increased venous return. These changes may improve the efficacy of IPC devices in the prevention of deep-venous thrombosis (DVT) formation. PMID- 10873727 TI - Transcapillary forces and the development of oedema in the lower limb of patients with chronic critical limb ischaemia (CLI). AB - OBJECTIVE: factors regulating transcapillary fluid transport were investigated to elucidate the causes of oedema in CLI. MATERIAL: sixteen patients, 6 men and 10 women (mean age of 79+/-10.3 years) with unilateral CLI and peripheral pitting oedema. METHODS: measurements were performed in both limbs. Interstitial fluid was collected by applying blister suction cups on the dorsolateral part of the foot and colloid osmotic pressure of this fluid (COP if) was measured in a colloid oncometer. Plasma colloid osmotic pressure (COP pl) was obtained from venous blood. Interstitial fluid pressure (P if) was measured by wick-in-needle technique. RESULTS: mean COP if in the limbs with CLI was 2.3 S.D. 0.5 mmHg, significantly lower than in the limbs without CLI (3.1 S.D. 0.7 mmHg, p<0.0001). Mean COP pl was 21.1 S.D. 1.8 mmHg, which was lower than in healthy controls. Mean plasma albumin concentration was 30 S.D. 6 g/l which was lower than the reference values. Mean P if in the limbs with CLI was 0.7 S.D. 1.6 mmHg, significantly higher than in the limbs without CLI (-1.4 S.D. 1.4 mmHg, p<0.0001). The calculated mean reabsorption pressure (P r) in the limbs with CLI was 19.6 S.D. 1.7 mmHg, significantly higher than in the contralateral limbs (16.7 S.D. 2.1 mmHg, p<0.001). CONCLUSION: a low plasma albumin concentration in patients with CLI agrees with the reduction in COP pl but cannot explain the oedema formation, since it is unilateral. The high P r may cause a high transcapillary filtration pressure, resulting in a relatively great net filtration and subsequent oedema formation. PMID- 10873728 TI - Continuous assessment of local metabolism by microdialysis in critical limb ischaemia. AB - OBJECTIVE: to investigate the feasibility of using microdialysate glucose, lactate and pyruvate concentrations for grading the severity of blood flow reduction in patients with critical limb ischaemia. PATIENTS AND METHODS: microdialysis catheters were inserted (two subcutaneously and one intramuscularly) in the symptomatic limb of ten patients. To further reduce limb perfusion, the lower leg was elevated during part of the experiment. RESULTS: elevation reduced ankle and toe blood pressure and transcutaneous oxygen tension. Microdialysate glucose concentration decreased at all three catheter sites, while lactate increased in the intramuscular catheter. Two patients interrupted the elevated position prematurely due to severe pain in the foot. They had among the highest lactate levels in the horizontal position and the most marked increases following provocation. Neither initial metabolite concentrations nor concentration changes during elevation were shown to correlate to conventional methods used to assess limb perfusion. CONCLUSIONS: in patients with critical limb ischaemia microdialysis can be used without complications. A significant decrease in glucose concentration may reflect lowered blood flow in the elevated position. Metabolic response, i.e. increase in lactate concentration during profoundly reduced limb perfusion was heterogeneous, indicating an overestimation of the presence of ischaemia in some patients using current diagnostic methods. PMID- 10873729 TI - Management of synchronous infrarenal aortic disease and large bowel cancer: a North-east of Scotland experience. AB - OBJECTIVES: to review our experience of combined aortic and colonic surgery. DESIGN: retrospective review. METHODS: synchronous aortic and colorectal procedures were identified from prospective computerised audit and archival vascular records. Clinical parameters were used as surrogates for measuring clinical outcome. RESULTS: six patients (F:M=2:1), median age 75.6 years (range 70-80 years) were identified with infrarenal aortic pathology (5 aneurysms, median AP diameter 6 cm, 1 occluded aortoiliac segment) and colonic carcinoma. All carcinomas were Dukes stage B and moderately well differentiated. Synchronous aortic and colonic resections were performed in five cases, bypass for aortoiliac occlusion was deferred in preference to colonic resection in one case. Operating time ranged between 3-6.5 hours (median 4 h), transfusion requirements 2-5 units (median 3 units). One anastomotic dehiscence was reported. With follow-up between 6 months to 6 years all patients remain alive; no patient has re-presented with graft sepsis or symptomatic tumour recurrence. CONCLUSION: synchronous resections of aortic and colonic lesion may be a treatment option in selected cases. PMID- 10873730 TI - Intestinal manipulation during elective aortic aneurysm surgery leads to portal endotoxaemia and mucosal barrier dysfunction. AB - OBJECTIVES: to investigate the effect of intestinal manipulation on intestinal permeability and endotoxaemia during elective abdominal aortic aneurysm (AAA) surgery. DESIGN: prospective randomised controlled study. PATIENTS AND METHODS: fourteen patients undergoing elective infrarenal AAA repair were randomised into either the transperitoneal (n=7) or extraperitoneal approach (n=7). Intestinal permeability was measured preoperatively (PO), and at day 1 (D1) and day 3 (D3) after surgery using the lactulose/mannitol absorption test. Portal and systemic blood samples were taken before clamping, at completion of proximal and distal anastomoses and immediately before abdominal wound closure, for endotoxin measurement using the chromogenic limulus amoebocyte lysate assay. RESULTS: intestinal permeability was significantly increased at D1 (0.107+/-0.04 (mean+/ S.E.M.)) in the transperitoneal group compared to the PO level (0.020+/-0.004, p<0.05) and to the extraperitoneal group at D1 (0.020+/-0.004, p<0.05) which showed no change in comparison with the PO level. No correlation was seen between increased intestinal permeability and aortic clamp time, operation time, amount of blood lost or transfused. However, a significantly higher concentration of portal endotoxin was detected intraoperatively in the transperitoneal group of patients in comparison to the extraperitoneal group (p<0.05). There was a significant positive correlation between portal endotoxaemia and intestinal permeability (r(s)=0.955 p=0.001). CONCLUSION: an increase in intestinal permeability and a greater degree of portal endotoxaemia are observed during transperitoneal approach to the aorta. This suggests that intestinal manipulation may impair gut mucosal barrier function and contribute to the systemic inflammatory response seen in AAA surgery. PMID- 10873731 TI - Is non-specific aneurysmal disease of the infrarenal aorta also a peripheral microvascular disease? AB - OBJECTIVES: to examine whether aneurysmal disease of the aorta has a functional component in the peripheral microIvasculature. MATERIALS: ten normal persons; and 15 patients who had been operated on for ruptured non-specific infrarenal aortic aneurysm months to years previously were studied. METHODS: blood flow rates were measured: (a) in the subcutaneous adipose tissue of the forefoot by the(133)xenon local washout method (perfusion through nutritive capillaries supplied by arterioles with elastin in the tunica media); and (b) in the arteriovenous anastomoses of the pulp of the first toe as measured by the heat washout method (perfusion predominantly through thick-walled tubes without elastin). Perfusion rates were measured in supine subjects at heart level, at 30 cm above and at 30 cm below heart level. RESULTS: in subcutaneous adipose tissue, the capillary blood flow rate was four times higher in patients with aneurysmal disease than in normal subjects. Both groups exhibited autoregulation of blood flow and a normal veno-arteriolar sympathetic axon reflex. Blood flow rates in the arteriovenous anastomoses of the pulp did not differ between aneurysm patients and normal subjects. Autoregulation and the axon reflex were absent in the arteriovenous anastomoses of normal subjects as well as in aneurysm patients. CONCLUSIONS: non specific aneurysmal disease of the infrarenal aorta has a peripheral functional component affecting arterioles but not arteriovenous anastomoses. PMID- 10873732 TI - Detection of viable Chlamydia pneumoniae in abdominal aortic aneurysms. AB - OBJECTIVES: to investigate the presence of Chlamydia pneumoniae in the wall of abdominal aortic aneurysms (AAAs) and in the aortas of patients without a history of cardiovascular disease. DESIGN: case-control study. MATERIALS: twenty-six consecutive patients operated for AAA were compared to 17 controls. METHODS: aorta was obtained at surgery or autopsy (controls) and prepared for immunohistochemical (IHC) analysis and culture for C. pneumoniae. Throat swabs from 14/26 patients were analysed by PCR for C. pneumoniae. Blood was obtained from 24/26 patients and from 178 70-year-old males. RESULTS: C. pneumoniae was detected in the aortic aneurysms of 20/26 patients by IHC. C. pneumoniae was cultured from 10 of the 20 IHC-positive patients. Only 1/17 controls was positive for C. pneumoniae by IHC (p=0.0001). PCR was positive for C. pneumoniae in 5/14 patients. Serological analysis by microimmunofluoresence (MIF) showed significantly more high titres of the specific antibodies to C. pneumoniae in patients than in age-matched male controls. CONCLUSIONS: we conclude that C. pneumoniae is often present in AAAs in a viable form and that C. pneumoniae is linked to the pathogenesis of AAA. PMID- 10873733 TI - Smoking, lung function and the prognosis of abdominal aortic aneurysm. The UK Small Aneurysm Trial Participants. AB - BACKGROUND: the UK Small Aneurysm Trial was established to test the benefit of prophylactic elective surgery for small abdominal aortic aneurysms (4.0-5.5 cm in diameter) and identify prognostic risk factors, including smoking. PATIENTS, METHODS AND OUTCOMES: one thousand and ninety patients (902 men and 188 women, mean age 69.3 years) were randomised to either early elective surgery or ultrasonography surveillance until the aneurysm diameter exceeded 5. 5 cm, mean follow-up was 4.6 years. Baseline assessments included lung function tests and cotinine (a smoking marker). The principal outcome measures were all-cause mortality and aneurysm rupture. RESULTS: during the course of the trial, aneurysm rupture was diagnosed in 25 patients and 309 patients died. Whereas self-reported smoking status was not significantly associated with survival, patients without any trace of plasma cotinine had a significantly improved long-term (6-year) survival, p=0.02. Current smokers had a lower FEV(1)than past- and never-smokers. FEV(1)was the most powerful predictor of long-term (6-year) survival, the crude death rates per 100 person-years were 9.1, 6.9 and 4.6 for those with FEV(1)<1.9 l, 1.9-2.5 l and >2.5 l respectively, p=0.001. Moreover, the rupture rate was 1.9% per year for patients positive for plasma cotinine compared with 0.5% in those without trace of plasma cotinine, p=0.004. CONCLUSIONS: self-reported smoking status underestimates the effect of continued smoking on the prognosis of patients with small abdominal aortic aneurysm. Patients with high plasma cotinine concentrations (smokers) have an increased risk of aneurysm rupture and poorer long-term survival. PMID- 10873734 TI - Changes of the infrarenal aortic segment after conventional abdominal aortic aneurysm repair. AB - OBJECTIVES: to delineate the natural history of the residual infrarenal aortic segment after conventional abdominal aortic aneurysm (AAA) repair. DESIGN: open prospective study. PATIENTS AND METHODS: between 1990 and 1997, 100 patients, who underwent conventional infrarenal AAA repair at our department, were followed annually by means of colour duplex ultrasonography. Data from 76 patients who had at least 3 scans were analysed. RESULTS: mean duration of follow-up was 4.7 years (range: 3-8 years). The residual infrarenal aorta dilated a mean of 0.57 mm annually. No patient required reoperation. There was no significant correlation between dilatation and any of the recorded risk factors except for the initial neck diameter (p=0.03). CONCLUSIONS: conventional AAA surgery is durable so that surveillance, during the first 5 postoperative years, is not justified in terms of cost-effectiveness. The impact of such a dilatation on endovascular AAA repair requires further investigation. PMID- 10873735 TI - Life-table analysis of primary and assisted success following endoluminal repair of abdominal aortic aneurysms: the role of supplementary endovascular intervention in improving outcome. AB - AIM: the aim of this study was to analyse the effect of supplementary endovascular intervention on the outcome of primary endoluminal repair of abdominal aortic aneurysm (AAA). METHODS: between May 1992 and December 1998, 266 patients underwent endoluminal repair of AAA. Minimum period of follow-up was 6 months. Those patients in whom the endoprosthesis could not be deployed were converted to open repair at the primary operation. Patients developing an early endoleak, within 31 days, were treated by a period of observation and secondary endovascular intervention in persistent cases. Patients developing a late endoleak were treated similarly, without a period of observation. Outcome was analysed by the life-table method. Primary success was defined as exclusion of the aneurysm from the circulation resulting from the original operation. Assisted success occurred when aneurysms with endoleaks became excluded from the circulation as a result of supplementary endovascular intervention. RESULTS: endoluminal repair failed in 17 patients requiring conversion to open repair at the original operation. Supplementary endovascular intervention was undertaken in 26 patients, with early endoleaks (n=6) and late endoleaks (n=20). Interventions involved deployment of secondary endoluminal grafts within the primary grafts (n=22), and coil embolisation (n=4). Successful exclusion of the aneurysm sac was achieved in 22 of 26 (85%) patients undergoing supplementary endovascular procedures. Conditional cumulative incidence of primary graft failure and secondary graft failure in the presence of all-cause mortality at 6 years was 47% and 25% respectively. CONCLUSIONS: supplementary endovascular intervention is an important adjunct to endoluminal AAA repair with the potential to improve outcome and avoid conversion to open repair. Successful supplementary endovascular intervention was achieved in 85% of patients in whom it was attempted. Life-table analysis showed these supplementary procedures to be durable in the long term. PMID- 10873736 TI - Fogarty balloon dilatation for intraoperative arterial spasm. AB - OBJECTIVE: to describe an intraoperative technique using a Fogarty balloon to treat arterial spasm following vascular bypass and endarterectomy. DESIGN: prospective case control study. SUBJECTS AND TREATMENT: twenty-two patients following femorodistal bypass surgery and one patient following carotid endarterectomy, with arterial spasm in the distal run-off on completion angiography, were treated with Fogarty balloon dilatation. MATERIALS: Fogarty balloon catheter (Baxtertrade mark). RESULTS: twenty-three patients (100%) with arterial spasm were successfully treated by Fogarty balloon as demonstrated on completion angiography. No complications were seen. CONCLUSION: this simple technique removes vascular spasm rapidly and produces an excellent angiographic result. PMID- 10873737 TI - Acute lower paraplegia as a possible sign of a concomitant injury of the major intra-abdominal blood vessels. PMID- 10873738 TI - Bilateral brachial artery fibromuscular dysplasia. PMID- 10873739 TI - Primary aortogastric fistula. PMID- 10873741 TI - Volume 19 contents PMID- 10873740 TI - Epidural abscess formation as a complication of femoropopliteal bypass graft infection. PMID- 10873743 TI - Acute and persistent viral life strategies and their relationship to emerging diseases. PMID- 10873744 TI - Optimization of regulated LTR-mediated expression. AB - Retroviral vectors are ideally suited to the study of gene function, allowing efficient, stable expression. Many biological systems (e.g., cell cycle, apoptosis) require the use of regulated expression systems. We therefore developed a regulated retroviral vector system, TRA99, based on a tetracycline transactivator-dependent LTR, where the MMLV enhancer was replaced with a tetracycline-response element. Using fluorescence-activated flow cytometric analysis of a destabilized green fluorescent protein to monitor expression levels, we optimized the minimal promoter configuration with respect to both activated and repressed transcription. The TRA99 vectors demonstrate regulated expression with activated levels comparable to those of standard retroviral vectors and repressed levels indistinguishable from background. This was achieved without using an internal promoter cassette, thus retaining the cis-packaging elements requisite for helper-mediated transfer. PMID- 10873745 TI - Reovirus mu2 protein determines strain-specific differences in the rate of viral inclusion formation in L929 cells. AB - Reovirus infection induces the formation of large cytoplasmic inclusions that serve as the major site of viral assembly. Reovirus strains type 3 Dearing (T3D) and type 1 Lang (T1L) differ in the rate of inclusion formation in L929 cells. The median time of inclusion formation is 18 h in cells infected with T3D and 39 h in cells infected with T1L. Using reassortant viruses that contain combinations of gene segments derived from T1L and T3D, we found that the M1 gene, which encodes the mu2 protein, is the primary determinant of the rate of inclusion formation. The S3 gene, which encodes the nonstructural protein sigmaNS, plays a secondary role in this process. The subcellular location of the mu2 protein was determined by confocal laser scanning microscopy using dual-fluorescence labeling of mu2 and the outer-capsid protein mu1/mu1C. In virus-infected cells, mu2 protein colocalized with other viral proteins in inclusions and was also distributed diffusely in the cytoplasm and nucleus. Expression of recombinant T1L and T3D mu2 proteins resulted in the formation of protein complexes resembling inclusions in both the cytoplasm and the nucleus with kinetics that reflected the strain of origin. The median time of mu2 protein complex formation was 22 h in cells transfected with the T3D M1 gene and 43 h in cells transfected with the T1L M1 gene. These findings suggest that the mu2 protein influences the rate of inclusion formation and contributes to inclusion morphogenesis. The requirement of mu2 protein in inclusion formation was tested by determining the subcellular localization of mu2 in cells infected with temperature-sensitive (ts) mutants that are defective in viral assembly. In contrast to infection with wild-type virus, mu2 did not colocalize with mu1/mu1C protein in subcellular structures that formed in cells infected at nonpermissive temperature with ts mutants tsH11.2, tsC447, and tsG453 with mutations in the M1, S2, and S4 genes, respectively. These results suggest that despite the role of the mu2 protein in controlling the rate of inclusion formation, this process is a concerted function of several reovirus proteins. PMID- 10873746 TI - Further characterization of the coronavirus infectious bronchitis virus 3C-like proteinase and determination of a new cleavage site. AB - Coronavirus infectious bronchitis virus (IBV) encodes a trypsin-like proteinase (3C-like proteinase) by ORF 1a, which has been demonstrated to play a pivotal role in proteolytic processing of gene 1-encoded polyproteins. In our previous studies, the proteinase was identified as a 33-kDa protein in IBV-infected cells, and its catalytic center was shown to consist of H(2820) and C(2922) residues. It is released from the 1a and 1a/1b polyproteins by autoprocessing at two Q-S dipeptide bonds (Q(2779)-S(2780) and Q(3086)-S(3087)). In this report, further characterization of the two cleavage sites demonstrates that the N-terminal Q(2779)-S(2780) site is tolerant to mutations at the P1 position. Deletion of the C-terminal region of the proteinase shows that a significant amount of the enzymatic activity is maintained upon deletion of up to 67 amino acids, suggesting that the extreme C-terminal region may be dispensable for the proteolytic activity of the proteinase. Analysis of the autoprocessing kinetics in vitro reveals that proteolysis at the Q(2779)-S(2780) site is the first cleavage event mediated by this proteinase. This is followed by cleavage at the Q(3086)-S(3087) site. The occurrence of both cleavage events in intact cells is potentially rapid and efficient, as no intermediate cleavage products covering the proteinase were detected in either IBV-infected or transfected cells. Immunofluorescence microscopy and subcellular fractionation studies further show differential subcellular localization of the proteinase in IBV-infected cells and in cells expressing the 3C-like proteinase alone, indicating that additional roles in viral replication might be played by this protein. Finally, a Q-A (Q(3379)-A(3380)) dipeptide bond encoded by nucleotides 10,663 to 10,668 was demonstrated to be a cleavage site of the proteinase. PMID- 10873747 TI - A functional NF-kappaB binding site in the human papillomavirus type 16 long control region. AB - By computer search, we identified one potential NF-kappaB binding site in the HPV16 long control region (LCR) at position 7554-7563 having two mismatches in comparison to the consensus NF-kappaB binding site of the Igkappa L promoter. Bandshift experiments with nuclear extracts from HeLa cells or purified glutathione S-transferase-p65 fusion protein clearly demonstrated that NF-kappaB is able to bind to this region of the LCR. However, in comparison to NF-kappaB binding on a consensus probe, the affinity of NF-kappaB for this site is about 250-fold reduced. When mutations were introduced into this NF-kappaB binding site, the activity of the LCR was increased, strongly suggesting that NF-kappaB was acting as a transcriptional repressor in the context of the HPV16 LCR. In addition, overexpression of NF-kappaB p65 repressed the activity of the HPV16 LCR, strengthening this conclusion. PMID- 10873748 TI - Pathogenesis of granulocytopenia and bone marrow atrophy during classical swine fever involves apoptosis and necrosis of uninfected cells. AB - Granulocytopenia, a hematological hallmark of classical swine fever, is partially responsible for the suppression of innate immune defenses during classical swine fever. The present report demonstrates that this depletion was apparent as early as 3 days postinfection (p.i.). Both mature peripheral and bone marrow neutrophils were affected, whereas immature neutrophils increased absolutely in the periphery and coincidentally immature myeloid progenitors in the bone marrow. These data suggest that a pathogenic relationship exists between these compartments. The central event was not the arrest of hematopoietic cell proliferation or of the mobilization process, but instead apoptosis and possibly also necrosis were shown to play a role. This increase in apoptotic and dead cells was detected as early as 1-3 days p.i. In contrast, viral RNA in bone marrow hematopoietic cells (BMHC) was first detected 5 days p.i., and significant amounts of infected BMHC were detected only 7 days p.i., with the major target being the myeloid compartment. The increased caspase-3 activity observed supported a role for apoptotic cell death. Furthermore, the elevated caspase-9 activity indicated the involvement of the mitochondrial apoptotic pathway. Taken together, the results demonstrate that granulocytopenia and bone marrow atrophy are mediated by hematopoietic cell death and that indirect virus-host-mediated mechanisms are likely to be responsible. PMID- 10873749 TI - A mutant of Sindbis virus that is resistant to pyrazofurin encodes an altered RNA polymerase. AB - Pyrazofurin (PZF), a cytidine analog and an inhibitor of orotate monophosphate decarboxylase, has been shown to decrease the levels of UTP and CTP in treated cells. When Sindbis virus (SV)-infected Aedes albopictus cells were treated with PZF, the yield of virus was reduced 100- to 1000-fold. By serial passage of our standard SV(STD) in Ae. albopictus cells in the presence of increasing concentrations of PZF, a mutant, SV(PZF), was derived, which was not inhibited by PZF. SV(PZF) is also resistant to adenosine, guanosine, and phosphono-acetyl-N aspartate, all of which have been shown to decrease levels of UTP and CTP. Analysis of chimeric viruses containing sequences from the SV(PZF) and parental genomes showed that the sequence between nt 5262 and 7999 conferred the PZF resistant phenotype. Sequencing of this region identified four mutations (nt 5750, 6627, 7543, and 7593), which are predicted to lead to amino acid changes: opal550L in nsP3 and M287L, K592I, and P609T in nsP4. Characterization of viruses containing one or more of these mutations demonstrated that all three mutations in the nsP4 coding region are required to produce full resistance to PZF. Using a molecular model of nsP4 based on the structure of HIV reverse transcriptase, we located amino acid change M287L at the tip of the fingers domain and K592I and P609T at the base of the thumb domain of the viral RNA polymerase. We suggest that these three amino acid changes in nsP4 alter the geometry of the NTP binding pocket so as to increase the affinity of the enzyme for CTP and UTP. PMID- 10873750 TI - Studies on the attenuation phenotype of polio vaccines: poliovirus RNA polymerase derived from Sabin type 1 sequence is temperature sensitive in the uridylylation of VPg. AB - Determinants of temperature sensitivity and/or attenuation in Sabin type 1 poliovirus reside in the 5' NTR and coding sequences of the capsid proteins and viral RNA polymerase, 3D(pol). Previous studies have implicated at least two mutations in 3D(pol) of Sabin 1 vaccine strain [PV1(S)], including a Y73H change, as contributing to these phenotypes. We have used an in vitro assay to test the first step in RNA synthesis, the uridylylation of the terminal protein VPg with 3D(pol) isolated from PV1(S). Wt and two mutant 3D(pol) proteins (Y73H, D53N/Y73H) were expressed in Escherichia coli and were purified, and their activities were measured in the synthesis of VPgpU(pU) and of VPg-linked poly(U) at 30 and 39.5 degrees C. Our results show that at 39.5 degrees C the Y73H mutation leads to a defect in the synthesis of VPgpUp(U) and of VPg-poly(U) but not in the elongation of a (dT)(15) primer. The double mutant protein had the same activities as Y73H 3D(pol). Using the yeast two-hybrid assay, we detected a reduced interaction between 3D(pol) molecules carrying either the single or double mutations. Tyrosine-73 maps to the finger domain in the three-dimensional structure of 3D(pol). A model will be presented in which a change of Y73 to H73 may interfere with an interaction between two polymerase molecules that, in turn, may interfere with VPg uridylylation. Alternative explanations, however, cannot be excluded at the present time. PMID- 10873751 TI - Characterization of the lysogenic repressor (c) gene of the Pseudomonas aeruginosa transposable bacteriophage D3112. AB - Bacteriophage D3112 is a Mu-like temperate transposable phage of Pseudomonas aeruginosa. Genetic mapping and DNA sequence analysis have identified the left end of the phage genome as encoding the transposase enzyme (A) and the lysogenic (c) repressor. The c open reading frame (ORF), located at the leftmost end of the phage genome and transcribed from right to left, has four possible GTG initiation codons. Using site-directed mutagenesis, each of the four GTG codons was modified to GTA, which cannot serve as an initiation codon. Plasmids were constructed expressing either the wild-type repressor ORF or the ORFs containing the mutated GTA codons. When introduced into Pseudomonas aeruginosa, no immunity to superinfection by D3112 was observed when the second GTG had been mutated. Northern blotting analysis demonstrated that the D3112 c repressor is transcribed as a 900-nt mRNA. The promoter region was defined by transcriptional lacZ fusions and primer extension analyses to bp 972-940 from the left end of the phage genome. When the D3112 c repressor was overexpressed and purified as a fusion protein with a C-terminal six-histidine extension (cts15-His6), it showed high affinity for a 261-bp PvuII fragment localized directly upstream of the c repressor ORF. Our results indicate that although D3112 c shows higher amino acid similarity to the lambda family of repressors than it does to those of Mu and D108, it appears that its structure and function more accurately reflect an evolutionary ancestry with those from transposable coliphages Mu and D108. PMID- 10873752 TI - Protection against measles virus-induced encephalitis by anti-mimotope antibodies: the role of antibody affinity. AB - Synthetic peptides mimicking a conformational B-cell epitope (M2) of the measles virus fusion protein (MVF) were used for the immunization of BALB/c mice and the anti-peptide and anti-virus antibody titers induced were compared. Of the panel of tested peptides, a chimeric peptide consisting of two copies of a T-helper epitope (residues 288-302 of MVF) and one copy of the mimotope M2 (TTM2) and a multiple antigen peptide with eight copies of M2 (MAP-M2) induced the highest titers of anti-M2 and anti-MV antibodies. Furthermore, peptides TTM2 and MAP-M2 induced antibodies with highest affinity for the mimotope and highest avidity for measles virus. Immunization with the MAP-M2 construct induced high titers of high affinity anti-M2 antibody despite the absence of a T-helper epitope, and lymphocyte proliferation data suggest that the addition of M2 to the MAP resulted in the generation of a structure capable of stimulating T-cell help. Sera with anti-M2 reactivity were pooled according to affinity values for binding to M2, and high- and low-affinity pools were tested for their ability to prevent MV induced encephalitis in a mouse model. The high-affinity serum pool conferred protection in 100% of mice, whereas the lower affinity pool conferred protection to only 50% of animals. These results indicate the potential of mimotopes for use as synthetic peptide immunogens and highlight the importance of designing vaccines to induce antibodies of high affinity. PMID- 10873753 TI - Hypervariability in the envelope genes of subgroup J avian leukosis viruses obtained from different farms in the United States. AB - Avian leukosis virus, subgroup J (ALV-J), has a wide host range, preferentially infecting meat-type birds, and produces a high incidence of myelocytomatosis and nephromas. Using the published sequences from HPRS-103 (ALV-J isolated in 1989 in Great Britain), we designed a set of PCR primers that amplified proviral DNA from nine U.S. field samples. The primers were specific for ALV-J, not amplifying DNA from uninfected cells or cells infected with ALV subgroups A-E. These primers expanded a 2.4-kb fragment that encompasses gp85, gp37, the E element, and most of the 3' LTR. We also developed a set of PCR primers that amplified a 2.1-kb fragment from ALV-J-infected cells and a 1.6-kb fragment from uninfected ev- chicken embryo fibroblasts (Line 0). Upon cloning and DNA sequencing, we determined that the 2.1- and 1.6-kb fragments contained ALV-J gp85- and gp37-like sequences. Comparison of the amino acid sequences demonstrated that the Line 0 sequences were 97.5% identical with the gp85 and gp37 of HPRS-103 and somewhat less identical with the other nine U.S. isolates. This suggests that the envelope genes of ALV-J may have arisen as a result of a recombination event between exogenous ALV and Line 0-like sequences in the chicken. Phylogenetic analysis also showed that the U.S. field isolates were closely related to one another and more distantly related to the European HPRS-103. The pattern of mutations in the U.S. field isolates suggests that the U.S. strains are slowly drifting away from their progenitor Line 0-like sequences. The development of effective vaccines and diagnostic tests is likely to become more problematic as the viruses continue to mutate. PMID- 10873754 TI - A molecular clone of simian-human immunodeficiency virus (DeltavpuSHIV(KU 1bMC33)) with a truncated, non-membrane-bound vpu results in rapid CD4(+) T cell loss and neuro-AIDS in pig-tailed macaques. AB - We report on the role of vpu in the pathogenesis of a molecularly cloned simian human immunodeficiency virus (SHIV(KU-1bMC33)), in which the tat, rev, vpu, env, and nef genes derived from the uncloned SHIV(KU-1b) virus were inserted into the genetic background of parental nonpathogenic SHIV-4. A mutant was constructed (DeltavpuSHIV(KU-1bMC33)) in which 42 of 82 amino acids of Vpu were deleted. Phase partitioning studies revealed that the truncated Vpu was not an integral membrane protein, and pulse-chase culture studies revealed that cells inoculated with DeltavpuSHIV(KU-1bMC33) released viral p27 into the culture medium with slightly reduced kinetics compared with cultures inoculated with SHIV(KU-1bMC33). Inoculation of DeltavpuSHIV(KU-1bMC33) into two pig-tailed macaques resulted in a severe decline of CD4(+) T cells and neurological disease in one macaque and a more moderate decline of CD4(+) T cells in the other macaque. These results indicate that a membrane-bound Vpu is not required for the CD4(+) T cell loss and neurological disease in SHIV-inoculated pig-tailed macaques. Furthermore, because the amino acid substitutions in the Tat and Rev were identical to those previously reported for the nonpathogenic SHIV(PPc), our results indicate that amino acid substitutions in the Env and/or Nef were responsible for the observed CD4(+) T cell loss and neurological disease after inoculation with this molecular clone. PMID- 10873755 TI - Genetic diversity of the Junin virus in Argentina: geographic and temporal patterns. AB - RNA was purified from 39 strains of cell-cultured Junin virus (JUN) from central Argentina, which included both human- and rodent-derived isolates (a total of 26 and 13, respectively), as well as from 2 laboratory JUN strains, XJ Cl3 and XJ #44. JUN-specific primers were used to amplify a 511-nucleotide (nt) fragment of the nucleocapsid protein gene and a 495-nt fragment of the glycoprotein 1 (GP1) gene. Genetic diversity among JUN strains studied was up to 13% at the nt level and up to 9% at the amino acid (aa) level for the GP1 gene and up to 9% (nt) and 4% (aa) for the NP gene. Phylogenetic analyses of both genes revealed three distinct clades. The first clade was composed of the JUN strains from the center of the endemic area and included the majority of JUN strains analyzed in the current study. The second clade contained 4 JUN strains isolated between 1963 and 1971 from Cordoba Province, the western-most edge of the known endemic area. The third clade contained 4 JUN strains that originated from Calomys musculinus trapped in Zarate, the northeastern edge of the known endemic area. Certain JUN sequences, which were obtained from GenBank and identified as XJ, XJ #44, and Candid #1 strains, appeared to form a separate clade. Over 400 nt of the GP1 and GP2 genes were additionally sequenced for 7 JUN strains derived from patients with different clinical presentations and outcomes of Argentine hemorrhagic fever. Analysis of the corresponding aa sequences did not allow us to attribute any particular genetic marker to the changing severity or clinical form of the human disease. PMID- 10873756 TI - Identification of domains of the HPV11 E1 protein required for DNA replication in vitro. AB - The HPV E1 and E2 proteins along with cellular factors, are required for replication of the viral genome. In this study we show that in vitro synthesized HPV11 E1 can support DNA replication in a cell-free system and is able to cooperate with E2 to recruit the host polymerase alpha primase to the HPV origin in vitro. Deletion analysis revealed that the N-terminal 166 amino acids of E1, which encompass a nuclear localization signal and a cyclin E-binding motif, are dispensable for E1-dependent DNA replication and for recruitment of pol alpha primase to the origin in vitro. A shorter E1 protein lacking the N-terminal 190 amino acids supported cell-free DNA replication at less than 25% the efficiency of wild-type E1 and was active in the pol alpha primase recruitment assay. An even shorter E1 protein lacking a functional DNA-binding domain due to a truncation of the N-terminal 352 amino acids was inactive in both assays despite the fact that it retains the ability to associate with E2 or pol alpha primase in the absence of ori DNA. We provide additional functional evidence that E1 interacts with pol alpha primase through the p70 subunit of the complex by showing that p70 can be recruited to the HPV origin by E1 and E2 in vitro, that the domain of E1 (amino acids 353-649) that binds to pol alpha primase in vitro is the same as that needed for interaction with p70 in the yeast two-hybrid system, and that exogenously added p70 competes with the interaction between E1 and pol alpha primase and inhibits E1-dependent cell-free DNA replication. On the basis of these results and the observation that pol alpha primase competes with the interaction between E1 and E2 in solution, we propose that these three proteins assemble at the origin in a stepwise process during which E1, following its interaction with E2, must bind to DNA prior to interacting with pol alpha primase. PMID- 10873757 TI - Analysis of RNA secondary structure in replication of human parainfluenza virus type 3. AB - The terminal RNA regions of the genomic and antigenomic RNAs of the paramyxoviruses and rhabdoviruses are known to contain sequences essential for RNA replication and transcription. The 3'- and 5'-termini of human parainfluenza virus type 3 (HPIV3) genomic RNA, termed leader and trailer sequences, respectively, are capable of forming stable stem-loop structures. Additionally, the 17 terminal bases of the leader and trailer are complementary and therefore also capable of forming a helical structure. We investigated the roles of the stem-loop structure and terminal complementarity in HPIV3 RNA replication and transcription in vivo using a minigenome containing all RNA elements necessary for these processes. By mutational analysis, we show that the RNA secondary structure features present at the termini of HPIV3 have no discernible role in replication or transcription. Rather, the primary sequence of these regions is what is critical in promoting replication. Interestingly, a mutation at leader base 24 was found to revert a mutation at leader position 5 but probably not via RNA secondary structure restoration. PMID- 10873758 TI - Circumvention of vector-specific neutralizing antibody response by alternating use of human and non-human adenoviruses: implications in gene therapy. AB - To determine whether non-human adenovirus-specific antibodies are cross neutralizing, rabbit and mouse anti-human adenovirus type 5 (HAd5), anti-bovine adenovirus type 3 (BAd3), and anti-porcine adenovirus type 3 (PAd3) sera were used in cross-virus neutralization assays. Adenovirus neutralizing antibodies were found to be virus-specific, suggesting that virus neutralizing epitope differs significantly in HAd5, BAd3, and PAd3. To further investigate whether immunity to an HAd5-derived vector could be circumvented by the use of non-human adenoviruses in vivo, mice were first immunized either intranasally or intraperitoneally with HAd5, BAd3, PAd3, or BAd3 + PAd3, and after development of adenovirus-specific antibodies, animals were inoculated with the HAd5 recombinant (AdCA36lacZ) containing the bacterial beta-galactosidase gene under the control of murine cytomegalovirus immediate-early promoter. Virus-inoculated animals developed virus-specific IgG and IgA antibodies. LacZ expression in animals initially primed with HAd5 was significantly reduced (P < 0.05), suggesting that the immune response against the vector could prevent the transgene expression following subsequent inoculation of the same vector, whereas LacZ expression in mice initially primed with BAd3, PAd3, or BAd3 + PAd3 was significantly higher (P > 0.05) than that obtained in HAd5-primed animals. Our results suggest that HAd5 , BAd3-, or PAd3-based vectors may be used sequentially for human gene therapy or vaccine production as a means to avoid immunity to the vector. PMID- 10873759 TI - Sequencing and characterization of A-2 plaque virus: A new member of the Picornaviridae family. AB - A-2 plaque virus (A2 virus) was originally isolated from the icteric-phase sera of US servicemen with viral hepatitis in the 1960s, but apart from a preliminary characterization little is known about the agent. We have now successfully cloned and sequenced the complete viral genome. A2 viral RNA consists of 7312 nucleotides, excluding the 62 nucleotide poly(A) tract at the 3' end, with one large open reading frame. Although clearly a member of the Picornaviridae, there is low homology to the available sequences, suggesting it is only loosely related to the classic rhino/enterovirus genus. In addition, there was no reactivity with group specific monoclonal antibody blends against polioviruses, enteroviruses 70 and 71, coxsackievirus B, and echoviruses. Two tamarins were inoculated with A2 virus to study viral pathogenesis. Both animals that received A2 virus became transiently viremic 1 week after the infection, as determined by RT-PCR, and they developed an antibody response to A2 virus. However, no physical signs or biochemical abnormalities, including elevated liver transaminases, were observed. In addition, no liver samples from patients with fulminant hepatitis (n = 7) or controls (n = 7) were positive for A2 viral RNA nor was anti-A2 neutralizing antibody detected in sera from hepatitis patients (n = 14), healthy laboratory donors (n = 14), or US blood donors (n = 33); however, most sera contained antibodies reactive with A2 virus proteins. These results suggest that A2 virus is a new member of the Picornaviridae but that its pathogenicity in nonhuman primates and association with human disease still need to be determined. PMID- 10873760 TI - Different immunological requirements for protection against acute versus persistent Friend retrovirus infections. AB - The propensity of retroviruses to rapidly establish persistent infections poses a formidable problem in vaccination strategies. In the current study, we use a live attenuated vaccine to study protection against acute and persistent Friend virus infections in mice. Adoptive transfers of immune CD8(+) T cells combined with passive immunizations with virus-neutralizing antibodies increased protection against acute disease compared with either treatment alone, but there was no protection against the establishment of persistent infection. In addition, the protection against acute disease elicited by the combination treatment was dependent on endogenous CD4(+) T cells as no protection was achieved in CD4(+) T cell-depleted mice. Quantitative studies showed that doubling the numbers of immune lymphocytes used in adoptive transfer experiments increased protection against acute disease depending on the type of lymphocyte subset used in the transfer. CD8(+) T cells were the most potent subset for the transfer of such protection. However, even high numbers of immune CD8(+) T cells gave no protection against the establishment of persistent infections. The data indicate that strengthening the numbers of specific immune cell subsets may have a beneficial effect on protection against acute disease, but protection from establishment of persistence requires complex immune responses involving multiple lymphocyte subsets. PMID- 10873761 TI - A cellular protein with an RNA-binding activity co-purifies with viral dsRNA from mycovirus-infected Helminthosporium victoriae. AB - A cellular protein that co-purifies with mycoviral dsRNA was isolated from the plant pathogenic fungus Helminthosporium victoriae (telomorph: Cochliobolus victoriae) infected with two viruses, the totivirus Helminthosporium victoriae 190S virus and the chrysovirus-like Helminthosporium victoriae 145S virus (Hv145SV). The cellular protein, which was, designated Hv-p68, accumulated to higher levels in virus-infected isolates compared to virus-free ones. The majority of the Hv145S dsRNAs were found in association with Hv-p68 and not packaged in virions. Hv-p68 could also be detected as a minor component of the virus capsid. Evidence is presented that Hv-p68 occurs in vivo as an octamer and that it possesses RNA-binding activities. Based on partial amino acid sequence analysis, Hv-p68 was shown to share significant sequence identity with alcohol oxidases from methylotrophic yeasts. Hv-p68 is proposed to play a role in viral RNA packaging/replication and in regulating viral pathogenesis. PMID- 10873762 TI - Two leaky-late HSV-1 promoters differ significantly in structural architecture. AB - The HSV-1 VP5 and VP16 transcripts are expressed with leaky-late (gamma1) kinetics and reach maximal levels after viral DNA replication. While the minimal VP5 promoter includes only an Sp1 site at -48, a TATA box at -30, and an initiator (Inr) element at the cap site, here we show that elements upstream of 48 can functionally compensate for the mutational loss of the critical Sp1 site at -48. To determine whether this is a general feature of leaky-late promoters, we have carried out a detailed analysis of the VP16 promoter in the context of the viral genome at the gC locus. Sequence analysis suggests a great deal of similarity between the two. Despite this, however, mutational analysis revealed that the 5' boundary of the VP16 promoter extends to ca. -90. This region includes an Sp1 binding site at -46, CAAT box homology at -77, and "E box" (CACGTG) at -85. Mutational and deletional analyses demonstrate that the proximal Sp1 site plays little or no role in promoter strength; despite this it can be shown to bind Sp1 protein using DNA mobility shift assays. Like the VP5 promoter, the VP16 promoter also requires an initiator element at the cap site. The VP16 Inr element differs in sequence from that of the VP5 promoter, and its deletion or mutation has a significantly smaller effect on promoter strength. The difference between these two Inr elements was confirmed by our finding that the VP16 initiator element binds to the 65-kDa YY1 transcription factor, and the VP5 Inr element competes poorly for the binding between the VP16 element and infected cell proteins in comparative bandshift assays. While the VP16 Inr sequence is identical to that of several murine TATA-less promoters, the VP16 Inr requires a TATA box for measurable activity. PMID- 10873763 TI - Identification of functional differences between prototype Epstein-Barr virus encoded LMP1 and a nasopharyngeal carcinoma-derived LMP1 in human epithelial cells. AB - The contribution of Epstein-Barr virus (EBV) strain variation to the pathogenesis of virus-associated tumours remains unknown. Given the central role of LMP1 in EBV-induced transformation, much interest has focused on the influence of LMP1 sequence variation on the signaling pathways and multiple downstream phenotypic consequences of LMP1 expression. The identification of LMP1 variants with a common 10-amino-acid deletion and additional point mutations (typified by the CAO LMP1 isolate) in EBV strains associated with nasopharyngeal carcinoma prompted us to examine the effect of stable prototype B95.8-LMP1 and CAO-LMP1 expression on the phenotype and differentiation of SCC12F human epithelial cells. Both forms of LMP1 were able to induce expression of the antiapoptotic A20 protein and provide protection from tumour necrosis factor-alpha-induced cytotoxicity. Although B95.8 LMP1 induced growth inhibition, expression of certain cell surface molecules (CD40, CD44, and CD54), and secretion of interleukin-6 and -8 in SCC12F cells, stable CAO-LMP1 expression failed to elicit these effects. Furthermore, B95. 8 LMP1, but not CAO-LMP1, induced alterations in cell morphology and blocked epithelial cell differentiation. Both B95.8-LMP1 and CAO-LMP1 induced similar levels of nuclear factor-kappaB activation, but the ability of CAO-LMP1 to activate the AP-1 pathway was relatively impaired. These data highlight significant functional differences between the prototype B95.8-LMP1 and the CAO LMP1 variant when stably expressed in human epithelial cells and suggest that continued analysis of LMP1 variants will help to further dissect the signaling pathways activated by LMP1 as well as provide insights into the contribution of LMP1 sequence variation to the pathogenesis of EBV-associated tumours. PMID- 10873764 TI - Characterization of phi8, a bacteriophage containing three double-stranded RNA genomic segments and distantly related to Phi6. AB - The three double-stranded RNA genomic segments of bacteriophage Phi8 were copied as cDNA, and their nucleotide sequences were determined. Although the organization of the genome is similar to that of Phi6, there is no similarity in either the nucleotide sequences or the amino acid sequences, with the exception of the motifs characteristic of viral RNA polymerases that are found in the presumptive polymerase sequence. Several features of the viral proteins differ markedly from those of Phi6. Although both phages are covered by a lipid containing membrane, the protein compositions are very different. The most striking difference is that protein P8, which constitutes a shell around the procapsid in Phi6, is part of the membrane in Phi8. The host attachment protein consists of two peptides rather than one and the phage attaches directly to the lipopolysaccharide of the host rather than to a type IV pilus. The host range of Phi8 includes rough strains of Salmonella typhimurium and of pseudomonads PMID- 10873765 TI - Long nucleotide insertions between the HN and L protein coding regions of human parainfluenza virus type 3 yield viruses with temperature-sensitive and attenuation phenotypes. AB - Recombinant parainfluenza virus 3 (rPIV3) is being developed as a vector to express foreign genes as a bivalent or multivalent live attenuated virus vaccine. In the present study, we examined the effect of inserted foreign sequence on virus replication in vitro and in vivo, focusing on the parameter of insert length. In one type of construct, foreign sequence of increasing length was flanked by PIV3 transcription signals and inserted as an additional gene unit (GU insert) between the HN and L genes, so that one additional mRNA would be made. In a second type of construct, foreign sequence was inserted into the downstream NCR (NCR insert) of the HN gene, so that the number of encoded mRNAs remained unchanged. In each case, the foreign sequence was designed to lack any significant open reading frame, which permitted an evaluation of the effect of insert length on replication independent of an effect of an expressed protein. The GU or NCR insert sizes ranged from 168 nucleotides (nt) to 3918 nt. rPIV3s containing GU insertions of up to 3918 nt in length, the largest size tested, were viable and replicated efficiently at permissive temperatures in vitro, but a reduction in plaque size was seen at 39 degrees C and 40 degrees C. The rPIV3 with a 3918-nt GU insertion was restricted in replication in the upper (fivefold) and lower (25-fold) respiratory tracts of hamsters. Although a 1908-nt GU insertion did not significantly modify replication of wild-type PIV3 in vitro or in vivo, its introduction significantly augmented the level of temperature sensitivity (ts) and attenuation (att) specified by three mutations in the L protein of a cold-passaged attenuated PIV3 vaccine virus. rPIV3s bearing a 3126- or 3894-nt NCR insertion exhibited in vitro and in vivo phenotypes like those of the rPIV3s bearing similar-sized GU insertions. These findings indicate that rPIV3s whose genome length has been increased by more than 3000 nt by either a GU or an NCR insertion exhibit an unexpected host-range phenotype, that is, efficient replication in vitro but restricted replication in hamsters, especially in the lower respiratory tract. Furthermore, these effects were greatly enhanced when the rPIV3 backbone contained other ts or att mutations. The implications of these findings for the use of single-stranded, negative-sense RNA viruses as vectors for vaccines are discussed. PMID- 10873766 TI - Inhibition of antiviral CTL responses by virus-infected cells: line item veto (cells) revisited. PMID- 10873767 TI - The role of CD4 and CD8 T cells in recovery and protection from retroviral infection: lessons from the Friend virus model. PMID- 10873768 TI - Poliovirus induces apoptosis in the human U937 promonocytic cell line. AB - The human promonocytic U937 cell line, which is moderately susceptible to poliovirus infection, has been used to investigate the induction of apoptosis by this virus. Infection of U937 cells with poliovirus induces morphological changes typical of apoptosis. Poliovirus-resistant U937 cells (PRU) have been isolated that are resistant to apoptosis induced by poliovirus, but that undergo apoptosis after treatment with TNF plus cycloheximide. Despite the fact that poliovirus triggers nitric oxide production in U937 cells, the inhibitor of inducible nitric oxide (NO) synthase, N(omega)-monomethyl-l-arginine, did not hinder apoptosis after infection, suggesting that NO does not play a direct role in this process. Finally, poliovirus infection of U937 cells led to the cleavage of pro-caspase-3 and poly(ADP-ribose)polymerase, indicating the activation of the CPP32 ICE-like cysteine protease in the induction of apoptosis. Our findings suggest that cellular death takes place in U937 cells productively infected by poliovirus as a result of apoptosis and involves caspase activation. PMID- 10873769 TI - Detection of the human herpesvirus 8-encoded cyclin protein in primary effusion lymphoma-derived cell lines. AB - The human herpesvirus 8 (HHV8/KSHV), along with certain other herpesviruses, encodes a gene with cyclin homology. Although the functional significance of the encoded cyclin is not clear at present, various lines of evidence propose a role for this cyclin in latently infected cells and possibly in the induction of tumors that arise in HHV8-infected individuals. We provide evidence here that the cyclin protein is expressed in HHV8 positive primary effusion lymphoma (PEL) derived cell lines and that its level of expression varies greatly between different lines. Our analysis indicates that the level of cyclin protein expression in different PEL cell lines may correlate with the level of transcript expression during latency but not in cells induced to undergo lytic replication. In highly expressing BC-3 cells the cyclin is complexed with cdk6, cdk4, cdk2, and cdk5 under both latent and lytic conditions, although subtle changes in the level of cdk association are seen after induction of the lytic cycle. Altogether our findings support the notion that the cyclin is a latency-associated gene product expressed in PEL tumor cells. They furthermore indicate that after lytic cycle induction, the level of cyclin transcript expression may not be a reliable indicator for the level of cyclin protein expression. PMID- 10873770 TI - The cowpox virus serpin SPI-3 complexes with and inhibits urokinase-type and tissue-type plasminogen activators and plasmin. AB - The orthopoxvirus serpin SPI-3 is N-glycosylated and suppresses fusion between infected cells. Although SPI-3 contains motifs conserved in inhibitory serpins, no proteinase inhibition by SPI-3 has been demonstrated, and mutations within the serpin reactive center loop (RCL) do not affect the ability to regulate cell fusion. We demonstrate here that SPI-3 protein expressed by transcription/translation in vitro is able to form SDS-stable complexes with the serine proteinases plasmin, urokinase-type plasminogen activator (uPA), and tissue-type plasminogen activator (tPA), consistent with inhibitory activity of the serpin. Weaker complexes were noted with factor Xa and thrombin. Mutation of Arg-340/Ser-341 at the predicted P1/P1' sites within the RCL prevented the formation of complexes between SPI-3 and plasmin, uPA, or tPA, suggesting that the arginine at the P1 position was required for complex formation. SPI-3 protein lacking the N-terminal signal peptide was purified by means of an N-terminal His(10)-tag and gave complete inhibition in vitro of plasmin, uPA, and tPA and partial inhibition of factor Xa. SPI-3 is therefore a bifunctional protein that acts as a proteinase inhibitor and suppresses infected cell-cell fusion. As a proteinase inhibitor, SPI-3 has similar specificity to the leporipoxvirus SERP1 protein of myxoma virus, although the two serpins are less than 30% identical overall. The inhibition constants of SPI-3 for plasmin, uPA, and tPA were determined to be 0.64, 0.51, and 1.9 nM, respectively, very similar to the corresponding K(i) values of SERP1. PMID- 10873771 TI - The cowpox virus SPI-3 and myxoma virus SERP1 serpins are not functionally interchangeable despite their similar proteinase inhibition profiles in vitro. AB - The myxoma virus (MYX) serpin SERP1 is a secreted glycoprotein with anti inflammatory activity that is required for full MYX virulence in vivo. The cowpox virus (CPV) serpin SPI-3 (vaccinia virus ORF K2L) is a nonsecreted glycoprotein that blocks cell-cell fusion, independent of serpin activity, and is not required for virulence of vaccinia virus or CPV in mice. Although SPI-3 has only 29% overall identity to SERP1, both serpins have arginine at the P1 position in the reactive center loop, and SPI-3 has a proteinase inhibitory profile strikingly similar to that of SERP1 [Turner, P. C., Baquero, M. T., Yuan, S., Thoennes, S. R., and Moyer, R. W. (2000) Virology 272, 267-280]. To determine whether SPI-3 and SERP1 were functionally equivalent, a CPV variant was constructed where the SPI-3 gene was deleted and replaced with the SERP1 gene regulated by the SPI-3 promoter. Cells infected with CPVDeltaSPI-3::SERP1 secrete SERP1 and show extensive fusion, suggesting that SERP1 is unable to functionally substitute for SPI-3 in fusion inhibition. In the reciprocal experiment, both copies of SERP1 were deleted from MYX and replaced with SPI-3 under the control of the SERP1 promoter. Cells infected with the MYXDeltaSERP1::SPI-3 recombinant unexpectedly secreted SPI-3, suggesting either that the cellular secretory pathway is enhanced by MYX or that CPV encodes a protein that prevents SPI-3 secretion. MYXDeltaSERP1::SPI-3 was as attenuated in rabbits as MYXDeltaSERP1::lacZ, indicating that SPI-3 cannot substitute for SERP1 in MYX pathogenesis. PMID- 10873772 TI - A mutation in the DE loop of the VP1 protein that prevents polyomavirus transcription and replication. AB - Natural mutants of the DE loop of the Polyomavirus (Py) major coat protein VP1 have been previously shown to display an altered host specificity (L. Ricci, R. Maione, C. Passananti, A. Felsani, and P. Amati, 1992, J. Virol. 66, 7153-7158). To better understand the role of this outfacing loop of the VP1 protein in Py infectivity, we constructed and characterized a Py mutant (Py M17) harboring a deletion of 7 AA within the tip of the DE loop. The mutant virions obtained after DNA transfection were unable to replicate and initiate early transcription in fibroblast cells. Complementation experiments performed to rescue the deficient M17 replication by means of wt functions revealed the cis-dominance of the mutation. In situ cell fractionation experiments demonstrated that the Py mutant, like the Py wt, enters the cells, reaches the nucleus and that both the viral DNA and VP1 protein are found tightly bound to the nuclear matrix. These data suggest that the VP1 protein, associated to the viral DNA, conditions early viral gene expression and that the DE loop of the protein must be involved in this process. PMID- 10873773 TI - The RGD sequence in the cytomegalovirus DNA polymerase accessory protein can mediate cell adhesion. AB - The murine cytomegalovirus (MCMV) polymerase processivity factor ppM44 (also referred to as pp50) is an abundant phosphoprotein found in MCMV-infected cells. Sequence analysis of the MCMV M44 open reading frame revealed an "RGD" motif that is also present in the human cytomegalovirus (HCMV) UL44 open reading frame. In this report, histidine-tagged M44 protein produced in Escherichia coli or the vaccinia/T7 expression system was purified to near homogeneity by metal chelation affinity chromatography using His*Bind resins. We demonstrated that recombinant M44 protein could mediate cell adhesion via its conserved "RGD" motif, because a single amino acid change (RGD to RGE) abolished cell attachment. In addition, cell adhesion was abolished in the presence of EDTA. We next showed that recombinant HCMV UL44, but not human herpesvirus type 6 p41, which lacks the RGD motif, could mediate cell adhesion in a similar manner. We also provided evidence that ppM44 was present in the culture medium during virus infection. Thus these results suggested that in addition to its primary role as the polymerase processivity factor, MCMV ppM44 may serve as a substrate for integrin-binding via its conserved RGD motif, with the potential for a novel role in the MCMV replication cycle. PMID- 10873774 TI - A mutagenic analysis of the E5 protein of human papillomavirus type 16 reveals that E5 binding to the vacuolar H+-ATPase is not sufficient for biological activity, using mammalian and yeast expression systems. AB - The E5 gene of human papillomavirus type 16 encodes a highly hydrophobic membrane protein previously shown to inhibit endosomal acidification, presumably by binding to the 16-kDa pore-forming subunit of the vacuolar H(+)-ATPase (v ATPase). The role of this interaction in the disruption of v-ATPase activity was explored through extensive mutagenesis of E5 to identify residues that mediate binding to the 16-kDa subunit. Coimmunoprecipitations revealed that the hydrophobic span between residues 41 and 54 is primarily responsible for this interaction and can be replaced with random hydrophobic amino acids. Studies using mutated 16-kDa proteins indicated that the fourth transmembrane domain of the pore subunit mediates binding to E5. Analysis of the E5 mutants in a yeast expression system revealed that several mutants that retained the capacity to bind to the 16-kDa subunit in COS-1 cells failed to disrupt vacuolar acidification. These data argue that E5 binding to the pore subunit is not sufficient for the associated activity of disruption of v-ATPase function. PMID- 10873775 TI - Genetic diversity of primary HIV-1 isolates and their sensitivity to antibody mediated neutralization. AB - Wide differences exist among primary isolates of HIV-1 in their sensitivity to antibody-mediated neutralization. While it is well documented that even short term tissue culture amplification of HIV-1 leads to a reduction in the genetic diversity of the viral quasispecies seen in vivo, viral isolates, while relatively homogeneous, are generally not clonal. We investigated whether the extent of genetic diversity within primary viral isolates correlates with their general susceptibility to neutralization. We compared the number of V1V2 and V3 V5 envelope variants detectable within 16 primary isolates selected to represent the extremes of the neutralization sensitive and resistant phenotypes. Using DNA heteroduplex tracking assays to estimate the extent of genetic diversity in these two regions of the envelope locus, we found that these primary isolates were made up of one to five distinguishable V1V2 and V3-V5 sequence variants. We found that higher levels of env genetic diversity did not correlate with increased resistance to antibody neutralization. PMID- 10873776 TI - The three-dimensional structure of bacteriophage PP7 from Pseudomonas aeruginosa at 3.7-A resolution. AB - The three-dimensional structure of phage PP7 from Pseudomonas aeruginosa has been determined to 3.7-A resolution. A comparison with distantly related small RNA phages showed that the biggest differences were found in the FG loops, forming the contacts around the fivefold and threefold axes. In contrast to the situation in other phages, the FG loops of phage PP7 are very similar in all three subunits. This supports the hypothesis that no switches are needed for the assembly control in these viruses. Some of the most conserved residues lie within the region involved in RNA binding in the related phages MS2 and seem to have the same function. PMID- 10873777 TI - Ubiquitous human adeno-associated virus type 2 autonomously replicates in differentiating keratinocytes of a normal skin model. AB - Since its discovery in 1966, adeno-associated virus type 2 (AAV) has been described as a helper-dependent parvovirus. However, in this study we demonstrate that AAV undergoes its complete life cycle, devoid of helper viruses or genotoxic agents, in the organotypic epithelial raft tissue culture system, a model of normal skin. AAV progeny production directly correlated with epithelial differentiation, as nondifferentiating keratinocytes were defective for this activity. Large nuclear virus arrays of particles of approximately 26 nm (parvovirus size) were observed in the granular layers of the raft epithelium by electron microscopy. Additionally, dosage-dependent histologic changes, some of which might be interpreted as cytopathology, were induced in the AAV-infected epithelial tissues. These data suggest a new biological model for AAV; that is, AAV is an epithelial-tropic autonomous parvovirus that can alter normal squamous differentiation. PMID- 10873778 TI - Direct measurement of CD8+ T cell responses in macaques infected with simian immunodeficiency virus. AB - The simian immunodeficiency virus (SIV) macaque model system has been used extensively to study AIDS pathogenesis and to test candidate vaccines for their ability to protect against homologous or heterologous challenge with pathogenic SIV or SHIV. Recent studies suggest that stimulation of HIV-1-specific CTL responses is important for effective vaccination against HIV-1. While quantitative measurements of SIV-specific cytotoxic T lymphocyte (CTL) responses have been facilitated by the use of tetrameric peptide complexes, this technique is currently limited to the study of Mamu-A*01-positive rhesus macaques. Furthermore, very few SIV-specific CTL epitopes have been identified, and there is limited identification of other MHC alleles in macaques. In this study, cytokine flow cytometry (CFC) was used to quantify SIV-specific CD8+ antigen reactive T cells in macaques infected with SIV. We found a strong correlation (r = 0.96, P < 0.001) between CD8+ antigen-reactive T cells stained with the Mamu A*01 p11C, C-M tetramer and production of intracellular TNF-alpha in the CFC assay. Furthermore, the CFC assay was used to identify a novel SIV-specific CTL epitope in Envelope (SIV Env, a.a. 486-494, sequence AEVAELYRL). The use of the CFC assay facilitates the study of antigen-reactive T cell responses in SIV infection and vaccination. PMID- 10873779 TI - A novel link between stress and human cytomegalovirus (HCMV) infection: sympathetic hyperactivity stimulates HCMV activation. AB - Recently, inflammatory mediators such as TNFalpha were identified as triggering active human cytomegalovirus (HCMV) infection. Here, we demonstrate that a highly stressful event in the absence of systemic inflammation, as observed in patients with acute myocardial infarction, leads to the development of an active HCMV infection in latently infected patients. Elucidating the molecular mechanism of virus activation, we could show that catecholamines directly stimulate the HCMV immediate-early (IE) enhancer/promoter in monocytic cells via beta-2 adrenergic receptors. Subsequent activation of the cAMP/PK-A-signaling pathway results in enhanced synthesis and binding of the transcription factor CREB-1/ATF-1 to the cAMP-responsive elements within the IE enhancer. Epinephrine also enhanced HCMV gene expression in infected THP-1 cells by about 50% in three of four experiments. These data suggest that HCMV, like HSV-1 and VZV, can be (re)activated under stress conditions. PMID- 10873780 TI - In vitro processing of human immunodeficiency virus type 1 Gag virus-like particles. AB - Human immunodeficiency virus (HIV) Gag proteins are assembled into virus particles and then cleaved by the virion-associated HIV protease. Concomitant with Gag processing, doughnut-like HIV particles (the immature form) are converted to particles containing condensed cores (the mature form). Here we describe the in vitro processing of immature HIV Gag virus-like particles (VLP) by exogenously added HIV protease. Following delipidization, sequential processing of immature VLP showed that the matrix (MA)/capsid (CA) junction was cleaved faster than the CA/nucleocapsid (NC) junction, an altered order of processing when compared with authentic processing. When the in vitro processed VLP were analyzed on density gradients, most of the MA, CA-p15 intermediate, and NC were detected as a highly multimeric form, equivalent to the unprocessed VLP. In contrast, CA was found as a monomer dissociated from the multimeric CA-p15 following cleavage of the CA/NC junction. Electron microscopy revealed that the in vitro processing was accompanied by conversion of the doughnut-like particles to particles containing condensed cores and spherical outer shells. The cores, however, lacked core shells, which are normally observed for authentic HIV, suggesting that the in vitro processing of immature VLP failed to produce core shells. PMID- 10873781 TI - Physical interaction between two varicella zoster virus gene regulatory proteins, IE4 and IE62. AB - Transfection assays demonstrate that the varicella zoster virus (VZV) immediate early 62 (IE62) protein is a major transactivator of VZV gene expression, whereas a second immediate-early protein, IE4, can act as a major coactivator of transactivation mediated through IE62. To test whether IE62 and IE4 interact physically, we performed several protein-protein interaction assays. Coimmunoprecipitation analyses using VZV-infected cell lysates as well as purified protein mixtures demonstrate that IE62 and IE4 form stable complexes in solution under stringent salt conditions. Enzyme-linked immunosorbent assay protein-protein interaction assays and maltose-binding protein capture assays demonstrate that IE62 binds IE4 in a concentration- and dose-dependent manner. Far Western blot analyses show that IE4 binds to an undermodified form of IE62, and the use of calf intestinal phosphatase and protein kinases suggests that the interaction with IE4 is dependent on the phosphorylation state of IE62. An IE4 binding domain on IE62 has been mapped using a set of truncated IE62 fusion peptides. Collectively, these results imply a direct and specific physical interaction between IE4 and less-phosphorylated forms of IE62. These data have implications for virion assembly, as well as for the regulation of gene expression in VZV-infected cells. PMID- 10873782 TI - BPV1 E2 protein enhances packaging of full-length plasmid DNA in BPV1 pseudovirions. AB - We studied determinants of efficient encapsidation of circular DNA, incorporating a PV early region DNA sequence (nt 584-1978) previously shown to enhance packaging of DNA within papillomavirus (PV)-like particles (VLPs). Insect coelomic cells (Sf-9) and cultured monkey kidney cells (Cos-1) were transfected with an 8-kb reporter plasmid incorporating the putative BPV packaging sequence and infected with BPV1 L1 and L2 recombinant baculovirus or vaccinia virus. Heavy (1.34 g/ml) and light (1.30 g/ml) VLPs were produced, and each packaged some of the input plasmid. In light VLPs, truncated plasmids, which nevertheless incorporated the PV-derived DNA packaging sequence, were more common than full length plasmids. Packaging efficiency of the plasmid was estimated at 1 plasmid per 10(4) VLPs in both Cos-1 and Sf-9 cells. In each cell type, expression of the BPV1 early region protein E2 in trans doubled the quantity of heavy but not light VLPs and also increased the packaging efficiency of full-length circular plasmids by threefold in heavy VLPs. The resultant pseudovirions incorporated significant amounts of E2 protein. Pseudovirions, comprising plasmids packaged within heavy VLPs, mediated the delivery of packaged plasmid into Cos-1 cells, whereby "infectivity" was blocked by antisera to BPV1 L1, but not antisera to BPV1 E4. We conclude that (a) packaging of DNA within PV L1+L2 pseudovirions is enhanced by BPV1 E2 acting in trans, (b) E2 may be packaged with the pseudovirion, and (c) E2 mediated enhancement of packaging favors 8-kb plasmid incorporation over incorporation of shorter DNA sequences. PMID- 10873783 TI - Characterization of human herpesvirus 7 U27 gene product and identification of its nuclear localization signal. AB - A monoclonal antibody, 5H4, that recognizes human herpesvirus 7 (HHV-7) was used in Western analysis to probe HHV-7-infected SupT1 cells. This antibody recognizes a 40-kDa virus-specific polypeptide that is expressed in the absence of viral DNA synthesis. By screening a lambdagt11 HHV-7 cDNA library, the gene encoding the protein was identified as the U27 open reading frame previously reported [J. Virol. (1996) 70, 5975-5989]. Immunofluorescent studies showed a punctate nuclear localization of the protein in both HHV-7-infected cells and transfected cells. A computer program predicted two classic nuclear localization signals (NLSs) in the middle and C-terminal regions of the protein. A C-terminal deletion mutant of the protein could not enter the nucleus, whereas green fluorescent protein or maltose binding protein fused to the C-terminal region of the protein was transported into the nucleus. These findings demonstrate that the predicted C-terminal, but not middle, NLS of the protein actually function as NLS. In addition, nuclear transport of a maltose binding protein-fusion protein containing the C-terminal NLS of the U27 protein was inhibited by both wheat germ agglutinin and a Q69L Ran GTP mutant, indicating that the U27 protein is transported into the nucleus from the cytoplasm by means of classic nuclear transport machinery. Interestingly, this NLS motif is highly conserved at the C-termini of all herpesvirus DNA polymerase processivity factors that have been examined. PMID- 10873784 TI - Inhibition of the RNA-dependent transactivation and replication of human immunodeficiency virus type 1 by a fluoroquinoline derivative K-37. AB - Human immunodeficiency virus type 1 (HIV-1) is unique in that it encodes its own transcriptional activator Tat, which specifically binds to the viral mRNA sequence TAR (transactivation response) element and activates viral transcription at the step of elongation as well as initiation. We recently reported that fluoroquinoline derivatives inhibited HIV-1 replication most likely by blocking viral transcription. In this report, we investigated the mechanism of action of one such compound 7-(3, 4-dehydro-4-phenyl-1-piperidinyl)-1, 4-dihydro-6-fluoro-1 methyl-8-trifluoromethyl-4-oxoquinoline-3-carbox ylic acid (K-37). We demonstrated that K-37 inhibited not only Tat but also other RNA-dependent transactivators. No effect was observed with DNA-dependent transactivators such as p65 (NF-kappaB) and Gal4VP16. Moreover, K-37 did not inhibit carboxyl-terminal domain (CTD)-kinase activities of CDK-activating kinase (CAK) and positive transcription elongation factor b (P-TEFb), which are known to be involved in Tat mediated transactivation at the step of transcriptional elongation. It is suggested that RNA-mediated transactivation may involve a common unknown factor to which K-37 directly interacts. Since K-37 did not appear to block DNA-mediated transactivation and thus did not show strong nonspecific cytotoxicity as reported previously, K-37 and its derivative compounds are considered to be feasible candidates for a novel AIDS therapy. PMID- 10873785 TI - SNDV, a novel virus of the extremely thermophilic and acidophilic archaeon Sulfolobus. AB - We describe a novel virus, SNDV (Sulfolobus neozealandicus droplet-shaped virus), of the crenarchaeotal archaeon Sulfolobus, which was found in a carrier state in a Sulfolobus strain isolated from a field sample from New Zealand. SNDV particles are droplet-shaped and densely covered by thin tail fibers at their pointed ends. The virion consists of a core and a coat. The latter has the appearance of a beehive and has a surface that is either helically ribbed or a stack of hoops. The genome is cccDNA of 20 kb, which is modified by dam-like methylation. It is cleaved by only a few type II restriction enzymes e.g., DpnI but not MboI, demonstrating an N(6)-methylation of the adenine residue in GATC sequences. The DNA-modifying system differentiates between virus and host. We postulate a virus encoded methylase that is active on hemimethylated DNA. The host range of SNDV is confined to few Sulfolobus strains from New Zealand. The virus persists in an unstable carrier state rather than as a prophage. Due to its uniqueness we propose to assign it to a novel virus family termed Guttaviridae. PMID- 10873786 TI - The HIV-1 Env protein signal sequence retards its cleavage and down-regulates the glycoprotein folding. AB - Secretory proteins and most membrane proteins are synthesized with a signal sequence that is usually cleaved from the nascent polypeptide chain, during its transport, into the lumen of the endoplasmic reticulum (ER). We have analyzed the kinetics of the cleavage of the HIV-1 Env protein signal sequence from gp160 and gp120 in HeLa, BHK, and Jurkat cells. Furthermore, we have determined the effects of this cleavage on the association of the gp160 and gp120 glycoproteins with the ER protein calnexin and the effects of the signal sequence cleavage on protein folding. The cleavage of the HIV-1 Env protein signal sequence on both gp160 and gp120 occurred very slowly in all three cell lines with a t(1/2) of 45-60 min. The core glycosylated and signal-sequence-retained forms of gp160 and gp120 associated with calnexin while the signal-sequence-cleaved forms of gp160 and gp120 had disassociated from calnexin and correctly folded as determined by their ability to associate with the CD4 cellular receptor. Further analysis of the folding state of gp160 and gp120 in nonreducing SDS-PAGE revealed that the signal sequence-retained and calnexin-associated forms of gp160 and gp120 migrated as broad, diffuse bands, whereas the signal-sequence-cleaved or CD4-associated forms of gp160 and gp120 migrated as single sharper bands. The cause of this retardation in the rate of folding and intracellular transport of HIV-1 glycoproteins was localized to their signal sequences by fusing the vesicular stomatitis virus G protein with the HIV-1 Env protein signal sequence and expressing this chimeric protein in mammalian cells. The HIV-1 Env protein signal sequence on the VSV-G protein also confers a reduced rate of cleavage and slow intracellular transport and folding of the chimeric G protein. These results provide direct evidence that in vivo the HIV-1 glycoprotein signal sequence inhibits the folding of HIV-1 Env protein. Our data also suggest a direct correlation between the rate of the signal sequence cleavage and protein folding. PMID- 10873787 TI - Characterization of low virulent strains of highly pathogenic A/Hong Kong/156/97 (H5N1) virus in mice after passage in embryonated hens' eggs. AB - Avian influenza A H5N1 viruses were isolated from humans for the first time in Hong Kong in 1997. The virulence of A/Hong Kong/156/97 (HK156) strain in mice was found to change significantly depending on the passage history of the virus. Madin-Darby canine kidney (MDCK) cell-grown parental virus and three of its clones derived from mouse brain showed high pathogenicity in mice after intranasal or intracerebral infection. In contrast, the egg-derived parental virus HK156-E3 and its cloned viruses were markedly less pathogenic in mice. It appeared that differences in pathogenicity among viruses derived from MDCK cells and eggs were due to their ability or inability to disseminate from the lungs to the brain. Sequence analysis of the entire protein coding regions of all eight RNA genome segments revealed a total of six conserved amino acid differences in the HA1 domain (residue 211) of the HA protein, as well as the PB1 (residues 456 and 712), PA (residue 631), NP (residue 127), and NS1 (residue 101) proteins that correlated with observed changes in virulence and neurovirulence of HK156 virus in mice. Thus it was evident that the passaging of HK156 in embryonated eggs led to the adaptation and selection of variants demonstrating markedly decreased pathogenicity and neurovirulence in mice that appeared to be attributable to specific amino acid changes in the HA and internal proteins. PMID- 10873788 TI - Anti-Gag cytolytic T lymphocytes specific for an alternative translational reading frame-derived epitope and resistance versus susceptibility to retrovirus induced murine AIDS in F(1) mice. AB - Murine AIDS (MAIDS) develops in susceptible mouse strains after infection with the LP-BM5 murine leukemia virus complex that contains causative defective, and ecotropic helper, retroviruses. We previously demonstrated that the MAIDS resistant H-2(d) strains BALB/cByJ and C57BL/KsJ generate MHC class I (K(d)) restricted virus-specific CD8(+) cytolytic T lymphocytes (CTLs) that lyse cells expressing either defective or ecotropic gag proteins. In contrast, the congenic BALB.B and closely related C57BL/6J MAIDS-susceptible H-2(b) strains were unable to serve as a source of gag-specific CTLs (Schwarz and Green, 1994), suggesting that anti-gag CTLs might provide a basis for resistance to MAIDS. Although its susceptibility to MAIDS was unknown, the (BALB/c x C57BL/6J) F(1) (CBY6F(1)) strain could also produce H-2(d)-, but not H-2(b)-, restricted, anti-gag CTLs (Schwarz and Green, 1994). Because of this correlation between anti-gag CTLs and resistance to MAIDS, it was important to provide more direct evidence in support of CTL-mediated protection and to determine both the fine specificity of CByB6F(1) anti-gag CTLs, in comparison with the resistant C57BL/Ks and BALB/c strains, and the susceptibility of this F(1) strain to LP-BM5-induced MAIDS. We report here that no symptoms of MAIDS were observed in CBY6F(1) (H-2(dxb)) mice. For F(2) mice, in contrast to the high susceptibility of H-2(b/b) mice, 77% of H 2(d/d) and 81% of H-2(b/d) F(2) mice did not exhibit MAIDS after LP-BM5 infection. These results are in contrast to other published studies that concluded that susceptibility, rather than resistance, is dominant in F(1) (resistant x susceptible or susceptible x resistant) mice. We also show that CBY6F(1) anti-gag CTLs exhibit a fine specificity shared by the MAIDS-resistant BALB/c and C57BL/Ks strains, that is, the immunodominant gag epitope, SYNTGRFPPL, encoded by an alternative open reading frame. Together with our direct demonstration here that in vivo monoclonal antibody (mAb) depletion of CD8(+) T cells converts genetically resistant mice to MAIDS susceptibility, these data on the ability to mount anti-ORF2/SYNTGRFPPL, gag-specific CTL responses strongly suggest that CTLs are a primary factor in determining MAIDS resistance. Accordingly, given the K(d)-restricted nature of the CTLs, the main genetic determinant of resistance appeared to be the codominant expression of the resistant H-2(d) haplotype. Interestingly, however, 19% of H-2(d/b) and 23% of the H-2(d/d) F(2) mice had at least one clinical aspect of MAIDS, suggesting that a non-MHC genetic determinant(s) can negatively influence T-cell protection and thus disease outcome PMID- 10873789 TI - A sensitive denaturing gradient-Gel electrophoresis assay reveals a high frequency of heteroplasmy in hypervariable region 1 of the human mtDNA control region. AB - A population study of heteroplasmy in the hypervariable region 1 (HV1) portion of the human mtDNA control region was performed. Blood samples from 253 randomly chosen individuals were examined using a sensitive denaturing gradient-gel electrophoresis (DGGE) system. This method is capable of detecting heteroplasmic proportions as low as 1% and virtually all heteroplasmy where the minor component is > or = 5%. Heteroplasmy was observed in 35 individuals (13.8%; 95% confidence interval [CI] 9.6-18.0). Of these individuals, 33 were heteroplasmic at one nucleotide position, whereas 2 were heteroplasmic at two different positions (a condition known as "triplasmy"). Although heteroplasmy occurred at a total of 16 different positions throughout HV1, it was most frequently observed at positions 16093 (n=13) and 16129 (n=6). In addition, the majority of heteroplasmic variants occurred at low proportions and could not be detected by direct sequencing of PCR products. This study indicates that low-level heteroplasmy in HV1 is relatively common and that it occurs at a broad spectrum of sites. Our results corroborate those of other recent reports indicating that heteroplasmy in the control region is more common than was previously believed-a finding that is of potential importance to evolutionary studies and forensic applications that are based on mtDNA variation. PMID- 10873791 TI - Trisomic pregnancy and earlier age at menopause. AB - We tested the hypothesis that the connection between advanced maternal age and autosomal trisomy reflects the diminution of the oocyte pool with age. Because menopause occurs when the number of oocytes falls below some threshold, our hypothesis is that menopause occurs at an earlier age among women with trisomic pregnancies than it does among women with chromosomally normal pregnancies. To determine their menstrual status, we interviewed women from our previous study of karyotyped spontaneous abortions who, in 1993, were age >/=44 years. Premenopausal women completed interviews every 4-5 mo, until menopause or until the study ended in 1997. The primary analyses compare 111 women whose index pregnancy was a trisomic spontaneous abortion with two groups: women whose index pregnancy was a chromosomally normal loss (n=157) and women whose index pregnancy was a chromosomally normal birth (n=226). We used a parametric logistic survival analysis to compare median ages at menopause. The estimated median age at menopause was 0.96 years earlier (95% confidence interval -0.18 to 2.10) among women with trisomic losses than it was among women with chromosomally normal losses and chromosomally normal births combined. Results were unaltered by adjustment for education, ethnicity, and cigarette smoking. Our results support the hypothesis that trisomy risk is increased with decreased numbers of oocytes. Decreased numbers may indicate accelerated oocyte atresia or fewer oocytes formed during fetal development. PMID- 10873792 TI - Motion vision: are 'speed lines' used in human visual motion? AB - Motion analysis poses problems for any visual system, not least because of the ambiguities inherent in motion signals. Recent studies suggest that the human motion system may exploit 'motion streaks' - analogous to the cartoonist's speed lines - to help resolve the direction of ambiguous motion. PMID- 10873793 TI - Non-coding RNA's: lightning strikes twice. AB - A second case has been found of a nematode gene involved in developmental timing that encodes a short, non-coding RNA. Both RNAs are expressed at specific times and appear to repress target genes by interacting with their 3' untranslated regions. A coincidence? Or does this pathway attract small RNA regulators? PMID- 10873790 TI - The ancestry of Brazilian mtDNA lineages. AB - We have analyzed 247 Brazilian mtDNAs for hypervariable segment (HVS)-I and selected restriction fragment-length-polymorphism sites, to assess their ancestry in different continents. The total sample showed nearly equal amounts of Native American, African, and European matrilineal genetic contribution but with regional differences within Brazil. The mtDNA pool of present-day Brazilians clearly reflects the imprints of the early Portuguese colonization process (involving directional mating), as well as the recent immigrant waves (from Europe) of the last century. The subset of 99 mtDNAs from the southeastern region encompasses nearly all mtDNA haplogroups observed in the total Brazilian sample; for this regional subset, HVS-II was analyzed, providing, in particular, some novel details of the African mtDNA phylogeny. PMID- 10873794 TI - DNA helicases: one small step for PcrA, one giant leap for RecBC? AB - One might imagine that the mechanism of helicases would relate to the number of base pairs that are unwound for each ATP that is hydrolysed. Recent studies, however, suggest the situation can be more complicated than this. PMID- 10873795 TI - Topographic mapping: organising by repulsion and competition? AB - The establishment of topographic maps of neuronal connections is believed to involve graded repulsion mediated by EphA receptors and ephrin-A ligands. Gene knockouts show that ephrin-A ligands do indeed have a crucial role in mapping, and that mechanisms in addition to graded repulsion must also be at work. PMID- 10873796 TI - Hemispheric asymmetries: a brain in two minds. AB - The two cerebral hemispheres are specialised for different cognitive functions, and which hemisphere's strategy is superior depends on the nature of the task. A new study of split-brain patients has provided another unexpected insight: the two hemispheres use different strategies when performing a guessing task. PMID- 10873797 TI - Neurobiology: the acid test for resting potassium channels. AB - The K(+) channels that generate the resting potential of mammalian neurons have been difficult to identify and characterize. Recent experiments on hypoglossal motoneurons and cerebellar granule cells suggest that the resting current in these neurons is carried by TASK-1, a member of the twin-pore family of K(+) channels. PMID- 10873798 TI - Evolution: hox genes and the cellared wine principle. AB - Two Drosophila Hox genes involved in segmentation, fushi tarazu and bicoid, appear to have acquired these roles by functional divergence from classical homeotic genes. Recent results indicate how genes with critical functions in development can evolve completely different functions among species. PMID- 10873799 TI - Transcriptional control: imprinting insulation. AB - Recent studies on the transcriptional regulation of two linked, imprinted genes, Igf2 and H19, have provided evidence for a novel mechanism of epigenetic control. DNA methylation controls the activity of an insulator element located between the two linked genes by regulating the binding of the zinc-finger protein CTCF. PMID- 10873800 TI - A novel Akt/PKB-related kinase is essential for morphogenesis in Dictyostelium. AB - BACKGROUND: Dictyostelium Akt/PKB is homologous to mammalian Akt/PKB and is required for cell polarity and proper chemotaxis during early development. The kinase activity of Akt/PKB kinase is activated in response to chemoattractants in neutrophils and in Dictyostelium by the chemoattractant cAMP functioning via a pathway involving a heterotrimeric G protein and PI3-kinase. Dictyostelium contains several kinases structurally related to Akt/PKB, one of which, PKBR-1, is investigated here for its role in cell polarity, movement and cellular morphogenesis during development. RESULTS: PKBR-1 has a kinase and a carboxy terminal domain related to those of Akt/PKB, but no PH domain. Instead, it has an amino-terminal myristoylation site, which is required for its constitutive membrane localization. Like Akt/PKB, PKBR-1 is activated by cAMP through a G protein-dependent pathway, but does not require PI3-kinase, probably because of the constitutive membrane localization of PKBR-1. This is supported by experiments demonstrating the requirement for membrane association for activation and in vivo function of PKBR-1. PKBR-1 protein is found in all cells throughout early development but is then restricted to the apical cells in developing aggregates, which are thought to control morphogenesis. PKBR-1 null cells arrest development at the mound stage and are defective in morphogenesis and multicellular development. These phenotypes are complemented by Akt/PKB, suggesting functional overlap between PKBR-1 and Akt/PKB. Akt/PKB PKBR-1 double knockout cells exhibit growth defects and show stronger chemotaxis and cell polarity defects than Akt/PKB null cells. CONCLUSIONS: Our results expand the previously known functions of Akt/PKB family members in cell movement and morphogenesis during Dictyostelium multicellular development. The results suggest that Akt/PKB and PKBR-1 have overlapping effectors and biological function: Akt/PKB functions predominantly during aggregation to control cell polarity and chemotaxis, whereas PKBR-1 is required for morphogenesis during multicellular development. PMID- 10873801 TI - A putative ubiquitin ligase required for efficient mRNA export differentially affects hnRNP transport. AB - BACKGROUND: In the nucleus, mRNAs are bound by hnRNP proteins. A subset of hnRNP proteins shuttle between the nucleus and cytoplasm and are believed to promote mRNA export by acting as adaptors between mRNA and the transport machinery. The existence of multiple shuttling hnRNP proteins raises the question of whether differentially regulated, hnRNP-specific mRNA export pathways exist. RESULTS: We have determined that Tom1p, a conserved protein with a hect (homology to E6-AP carboxyl terminus) E3 ubiquitin ligase domain, is required for efficient mRNA export in S. cerevisiae, yet differentially affects hnRNP protein localization and export. Mutations in tom1 predicted to abolish ubiquitin ligase activity block efficient export of Nab2p and mRNA, causing Nab2p-mRNA complexes to accumulate in a punctate pattern coincident with the nuclear pore complex (NPC). Notably, the subcellular distribution of several other hnRNP proteins is not affected. In particular, Np13p remains mRNA-associated and continues to be efficiently exported in tom1 mutants. CONCLUSION: Our results demonstrate that mutations predicted to affect the enzymatic activity of the Tom1p ubiquitin ligase differentially affect export of hnRNP proteins in association with mRNA. We propose the existence of multiple mRNA export pathways, with export of Nab2p associated mRNAs dependent on a branch of the ubiquitin protein modification pathway. PMID- 10873802 TI - A human homolog of the C. elegans polarity determinant Par-6 links Rac and Cdc42 to PKCzeta signaling and cell transformation. AB - BACKGROUND: Rac and Cdc42 are members of the Rho family of small GTPases. They modulate cell growth and polarity, and contribute to oncogenic transformation by Ras. The molecular mechanisms underlying these functions remain elusive, however. RESULTS: We have identified a novel effector of Rac and Cdc42, hPar-6, which is the human homolog of a cell-polarity determinant in Caenorhabditis elegans. hPar 6 contains a PDZ domain and a Cdc42/Rac interactive binding (CRIB) motif, and interacts with Rac1 and Cdc42 in a GTP-dependent manner. hPar-6 also binds directly to an atypical protein kinase C isoform, PKCzeta, and forms a stable ternary complex with Rac1 or Cdc42 and PKCzeta. This association results in stimulation of PKCzeta kinase activity. Moreover, hPar-6 potentiates cell transformation by Rac1/Cdc42 and its interaction with Rac1/Cdc42 is essential for this effect. Cell transformation by hPar-6 involves a PKCzeta-dependent pathway distinct from the pathway mediated by Raf. CONCLUSIONS: These findings indicate that Rac/Cdc42 can regulate cell growth through Par-6 and PKCzeta, and suggest that deregulation of cell-polarity signaling can lead to cell transformation. PMID- 10873803 TI - A myosin light chain mediates the localization of the budding yeast IQGAP-like protein during contractile ring formation. AB - Cytokinesis in animal cells is accomplished through constriction of an actomyosin ring [1] [2] [3], which must assemble at the correct time and place in order to ensure proper division of genetic material and organelles. Budding yeast is a useful model system for determining the biochemical pathway of contractile ring assembly. The budding yeast IQGAP-like protein, Cyk1/Iqg1p, has multiple roles in the assembly and contraction of the actomyosin ring [4] [5] [6]. Previously, the IQ motifs of Cyk1/Iqg1p were shown to be required for the localization of this protein at the bud neck [6]. We have investigated the binding partner of the IQ motifs, which are predicted to interact with calmodulin-like proteins. Mlc1p was originally identified as a light chain for a type V myosin, Myo2p; however, a cytokinesis defect associated with disruption of the MLC1 gene suggested that the essential function of Mlc1p may involve interactions with other proteins [7]. We show that Mlc1p binds the IQ motifs of Cyk1/Iqg1p and present evidence that this interaction recruits Cyk1/Iqg1p to the bud neck. Immunofluorescence staining shows that Mlc1p is localized to sites of polarized cell growth as well as the bud neck before and independently of Cyk1p. These results demonstrate that Mlc1p is important for the assembly of the actomyosin ring in budding yeast and that this function is mediated through interaction with Cyk1/Iqg1p. PMID- 10873804 TI - Dynamics of phosphatidylinositol 4,5-bisphosphate in actin-rich structures. AB - Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) is known to regulate a wide range of molecular targets and cellular processes, from ion channels to actin polymerization [1] [2] [3] [4] [5] [6]. Recent studies have used the phospholipase C-delta1 (PLC-delta1) pleckstrin-homology (PH) domain fused to green fluorescent protein (GFP) as a detector for PI(4,5)P(2) in vivo [7] [8] [9] [10]. Although these studies demonstrated that PI(4,5)P(2) is concentrated in the plasma membrane, its association with actin-containing structures was not reported. In the present study, fluorescence imaging of living NIH-3T3 fibroblasts expressing the PLC-delta1 PH domain linked to enhanced green fluorescent protein (PH-EGFP) reveals intense, non-uniform fluorescence in distinct structures at the cell periphery. Corresponding fluorescence and phase contrast imaging over time shows that these fluorescent structures correlate with dynamic, phase-dense features identified as ruffles and with microvillus-like protrusions from the cell's dorsal surface. Imaging of fixed and permeabilized cells shows co-localization of PH-EGFP with F-actin in ruffles, but not with vinculin in focal adhesions. The selective concentration of the PH-EGFP fusion protein in highly dynamic regions of the plasma membrane that are rich in F-actin supports the hypothesis that localized synthesis and lateral segregation of PI(4,5)P(2) spatially restricts actin polymerization and thereby affects cell spreading and retraction. PMID- 10873805 TI - Enforced polarisation and locomotion of fibroblasts lacking microtubules. AB - The polarisation and locomotion of fibroblasts requires an intact microtubule cytoskeleton [1]. This has been attributed to an influence of microtubule mediated signals on actin cytoskeleton dynamics, either through the generation of active Rac to promote protrusion of lamellipodia [2], or through the modulation of substrate adhesion via microtubule targeting events [3] [4]. We show here that the polarizing role of microtubules can be mimicked by externally imposing an asymmetric gradient of contractility by local application of the contractility inhibitor ML-7. Apolar fibroblasts lacking microtubules could be induced to polarize and to move by application of ML-7 by micropipette to one side of the cell and then to the trailing vertices that developed. The release and retraction of trailing adhesions could be correlated with a relaxation of traction on the substrate and a differential shortening of stress-fibre bundles, with their distal tips relaxed. Although retraction and protrusion in these conditions resembled control cell locomotion, the normal turnover of adhesion sites that form behind the protruding cell front was blocked. These findings show that microtubules are dispensable for fibroblast protrusion, but are required for the turnover of substrate adhesions that normally occurs during cell locomotion. We conclude that regional contractility is modulated by the interfacing of microtubule-linked events with focal adhesions and that microtubules determine cell polarity via this route. PMID- 10873806 TI - Active maintenance of mHDA2/mHDAC6 histone-deacetylase in the cytoplasm. AB - The intracellular localization, and thereby the function, of a number of key regulator proteins tagged with a short leucine-rich motif (the nuclear export signal or NES) is controlled by CRM1/exportin1, which is involved in the export of these proteins from the nucleus [1]. A common characteristic of these regulators is their transient action in the nucleus during either a specific phase of the cell cycle or in response to specific signals [1]. Here, we show that a particular member of the class II histone-deacetylases mHDA2/mHDAC6 [2] belongs to this family of cellular regulators that are present predominantly in the cytoplasm, but are also capable of shuttling between the nucleus and the cytoplasm. A very potent NES present at the amino terminus of mHDAC6 was found to play an essential role in this shuttling process. The sub-cellular localization of mHDAC6 appeared to be controlled by specific signals, since the arrest of cell proliferation was found to be associated with the translocation of a fraction of the protein into the nucleus. Data presented here suggest that mHDAC6 might be the first member of a functionally distinct class of deacetylases, responsible for activities not shared by other known histone deacetylases. PMID- 10873807 TI - Integrin engagement suppresses RhoA activity via a c-Src-dependent mechanism. AB - The Rho family GTPases Cdc42, Rac1 and RhoA control many of the changes in the actin cytoskeleton that are triggered when growth factor receptors and integrins bind their ligands [1] [2]. Rac1 and Cdc42 stimulate the formation of protrusive structures such as membrane ruffles, lamellipodia and filopodia. RhoA regulates contractility and assembly of actin stress fibers and focal adhesions. Although prolonged integrin engagement can stimulate RhoA [3] [4] [5], regulation of this GTPase by early integrin-mediated signals is poorly understood. Here we show that integrin engagement initially inactivates RhoA, in a c-Src-dependent manner, but has no effect on Cdc42 or Rac1 activity. Additionally, early integrin signaling induces activation and tyrosine phosphorylation of p190RhoGAP via a mechanism that requires c-Src. Dynamic modulation of RhoA activity appears to have a role in motility, as both inhibition and activation of RhoA hinder migration [6] [7] [8]. Transient suppression of RhoA by integrins may alleviate contractile forces that would otherwise impede protrusion at the leading edge of migrating cells. PMID- 10873808 TI - Enteropathogenic E. coli translocated intimin receptor, Tir, interacts directly with alpha-actinin. AB - Enteropathogenic Escherichia coli (EPEC) triggers a dramatic rearrangement of the host epithelial cell actin cytoskeleton to form an attaching and effacing lesion, or pedestal. The pathogen remains attached extracellularly to the host cell through the pedestal for the duration of the infection. At the tip of the pedestal is a bacterial protein, Tir, which is secreted from the bacterium into the host cell plasma membrane, where it functions as the receptor for an EPEC outer membrane protein, intimin [1]. Delivery of Tir to the host cell results in its tyrosine phosphorylation, followed by Tir-intimin binding. Tir is believed to anchor EPEC firmly to the host cell, although its direct linkage to the cytoskeleton is unknown. Here, we show that Tir directly binds the cytoskeletal protein alpha-actinin. alpha-Actinin is recruited to the pedestal in a Tir dependent manner and colocalizes with Tir in infected host cells. Binding is mediated through the amino terminus of Tir. Recruitment of alpha-actinin occurs independently of Tir tyrosine phosphorylation. Recruitment of actin, VASP, and N WASP, however, is abolished in the absence of this tyrosine phosphorylation. These results suggest that Tir plays at least three roles in the host cell during infection: binding intimin on EPEC; mediating a stable anchor with alpha-actinin through its amino terminus in a phosphotyrosine-independent manner; and recruiting additional cytoskeletal proteins at the carboxyl terminus in a phosphotyrosine-dependent manner. These findings demonstrate the first known direct linkage between extracellular EPEC, through the transmembrane protein Tir, to the host cell actin cytoskeleton via alpha-actinin. PMID- 10873809 TI - Evidence for cognition without grammar from causal reasoning and 'theory of mind' in an agrammatic aphasic patient. AB - Understanding the inter-relationship between language and thought is fundamental to the study of human cognition [1] [2] [3]. Some investigators have proposed that propositions in natural language serve to scaffold thinking, by providing, for example, a sequential structure to a massively parallel process [4]. Others have maintained that certain thoughts, such as inferring the mental states of others, termed 'theory of mind' (ToM) reasoning, and identifying causal relationships, necessarily involve language propositions [5]. It has been proposed that ToM reasoning depends upon the possession of syntactic structures such as those that permit the embedding of false propositions within true statements ('Mary knows that John (falsely) thinks chocolates are in the cupboard') [6]. The performance on reasoning tasks of individuals with severe agrammatic aphasia (an impairment of language following a lesion of the perisylvian areas of the language-dominant hemisphere) offers novel insights into the relation between grammar and cognition. We report the unusual case of a patient with agrammatic aphasia of such severity that language propositions were not apparently available at an explicit processing level in any modality of language use. Despite this profound impairment in grammar, he displayed simple causal reasoning and ToM understanding. Thus, reasoning about causes and beliefs involve processes that are independent of propositional language. PMID- 10873811 TI - Hard cases (contd) PMID- 10873810 TI - Form and motion coherence activate independent, but not dorsal/ventral segregated, networks in the human brain. AB - There is much evidence in primates' visual processing for distinct mechanisms involved in object recognition and encoding object position and motion, which have been identified with 'ventral' and 'dorsal' streams, respectively, of the extra-striate visual areas [1] [2] [3]. This distinction may yield insights into normal human perception, its development and pathology. Motion coherence sensitivity has been taken as a test of global processing in the dorsal stream [4] [5]. We have proposed an analogous 'form coherence' measure of global processing in the ventral stream [6]. In a functional magnetic resonance imaging (fMRI) experiment, we found that the cortical regions activated by form coherence did not overlap with those activated by motion coherence in the same individuals. Areas differentially activated by form coherence included regions in the middle occipital gyrus, the ventral occipital surface, the intraparietal sulcus, and the temporal lobe. Motion coherence activated areas consistent with those previously identified as V5 and V3a, the ventral occipital surface, the intraparietal sulcus, and temporal structures. Neither form nor motion coherence activated area V1 differentially. Form and motion foci in occipital, parietal, and temporal areas were nearby but showed almost no overlap. These results support the idea that form and motion coherence test distinct functional brain systems, but that these do not necessarily correspond to a gross anatomical separation of dorsal and ventral processing streams. PMID- 10873812 TI - Bristling all over. PMID- 10873813 TI - Notch. PMID- 10873814 TI - Coliform confusion. PMID- 10873815 TI - Dynactin. PMID- 10873816 TI - Bax, Bid and the permeabilization of the mitochondrial outer membrane in apoptosis. AB - Mitochondria provide a key amplification step in the apoptotic pathway of many cells by releasing apoptogenic proteins into the cytosol. Recent studies have provided insights into how Bax and Bid may operate synergistically to recruit mitochondria into the pathway and how GD3 ganglioside, a metabolite of the sphingomyelin pathway, may also be used. In ischaemic disease, activation of the mitochondrial permeability transition pore may bypass the requirement for these factors. PMID- 10873817 TI - Membrane traffic in polarized epithelial cells. AB - Epithelial cells contain apical and basolateral surfaces with distinct compositions. Sorting of certain proteins to the basolateral surface involves the epithelial-specific mu 1b clathrin adaptor subunit. Recent results have shown that targeting to the basolateral surface utilizes the exocyst, whereas traffic to the apical surface uses syntaxin 3. Endocytosis at the apical surface is regulated by ARF6. Transcytosis of IgA is regulated by the p62Yes tyrosine kinase. PMID- 10873818 TI - Messenger proteins: homeoproteins, TAT and others. AB - Some proteins are internalized by live cells by a process that does not involve classical endocytosis and thus gain direct access to the cytoplasm and nucleus. These same proteins are often secreted, despite the absence of a signal peptide. Recent studies of this unexpected mode of intercellular signaling have opened the way for biotechnological developments. PMID- 10873819 TI - Unique features of the plant vacuolar sorting machinery. AB - Multiple types of vacuoles can exist within the same plant cell, and different vesicle-trafficking pathways transport proteins to each of them. Recent work has identified proteins unique to each vacuole type, and the transport pathways have begun to be elucidated. Plant trafficking proteins are usually encoded by small gene families, the different members of which have distinct functions in the endomembrane system. PMID- 10873820 TI - Penetration of protein toxins into cells. AB - AB toxins deliver their enzymatically active A domain to the cytosol. Some AB toxins are able to penetrate cellular membranes from endosomes where the low pH triggers their translocation. One such toxin is diphtheria toxin and important features of its translocation mechanism have been unraveled during the last year. Other toxins depend on retrograde transport through the secretory pathway to the ER before translocation, and recent findings suggest that these toxins take advantage of the ER translocation machinery normally used for transport of cellular proteins. In addition, the intracellular targets of many of these toxins have been identified recently. PMID- 10873821 TI - Sorting and transport in C. elegans: aA model system with a sequenced genome. AB - In the past few years, yeast and cultured cells have been the model systems of choice for the study of protein sorting and transport. Recently, there has been a surge in research in these areas in Caenorhabditis elegans, with advances in experimental techniques and genomics. New in vivo assays that monitor endocytosis and neuronal transport have been used to delineate roles for several genes in these processes. PMID- 10873822 TI - Transport between ER and Golgi. AB - In the last eighteen months, it has become clear that some classes of proteins are actively recruited into endoplasmic reticulum export carriers, whereas others are exported as bulk-flow cargo. Subsequent transport to the Golgi is mediated by tubulovesicular membranes. The anterograde membrane flow is compensated for by a retrograde pathway, which, in addition to the recycling of membrane and proteins to the endoplasmic reticulum, may play a role in anterograde cargo concentration. PMID- 10873823 TI - Connecting vesicle transport to the cytoskeleton. AB - The cytoskeleton is crucial for the efficient and polarized transport of vesicles in intracellular membrane-sorting pathways. Recent studies have identified specific kinesin, dynein, and myosin motor proteins that mediate defined membrane transport steps. Important clues have also been uncovered about the nature of motor-protein receptors on vesicular cargoes and the molecular mechanisms of motor-protein regulation. PMID- 10873825 TI - Lipid trafficking and sorting: how cholesterol is filling gaps. AB - Recent research has highlighted a role for cholesterol homeostasis in the regulation of trafficking and sorting of sphingolipids. This sorting may dictate the nature of the acyl chain species of phospholipids in the plasma membrane which, in turn, may govern the selective partitioning of these lipids into lateral domains. Recently, several proteins have been identified that play a role in the flow and sorting of all major lipid classes. PMID- 10873824 TI - Divide and multiply: organelle partitioning in yeast. AB - The mechanisms ensuring accurate partitioning of yeast vacuoles and mitochondria are distinct, yet they share common elements. Both organelles move along actin filaments, and both organelles require fusion and fission to maintain normal morphology. Recent studies have revealed that while vacuolar inheritance requires a processive myosin motor, mitochondrial inheritance requires controlled actin polymerization. Distinct sets of proteins required for the fusion and fission of each organelle have also been identified. PMID- 10873826 TI - Organization of the Golgi apparatus. AB - Investigators are revisiting basic concepts of the structure-function relationships of the Golgi apparatus. A key issue is the properties of the transport carriers that operate within the secretory pathway. Golgi morphology and dynamics differ between species but data from various model systems are pointing toward an integrated view of Golgi organization. PMID- 10873827 TI - Sodium transport and salt tolerance in plants. AB - The ability of plant cells to maintain low cytosolic sodium concentrations is an essential process associated with the ability of plants to grow in high salt concentrations. Recent results have identified pathways for Na(+) entry, and the cloning of vacuolar Na(+)/H(+) antiporters has demonstrated the role of intracellular Na(+) compartmentation in plant salt tolerance. PMID- 10873828 TI - Understanding the insertion of transporters and other membrane proteins. AB - Recent studies show that transporters integrate into the lipid bilayer using topogenic sequences present throughout the entire polypeptide chain. These topogenic sequences can act in unpredictable ways with new translocation/stop transfer activities. In addition, a new membrane-insertion pathway has been identified in bacteria with homologs in mitochondria and chloroplasts. PMID- 10873829 TI - Structural insights into clathrin-mediated endocytosis. AB - The process of clathrin-mediated endocytosis from the plasma membrane has been the subject of many biological and biochemical investigations. Recent atomic resolution structures determined by X-ray crystallography now enable the molecular basis for the interactions of some components of the endocytic machinery to be understood in detail. PMID- 10873830 TI - Multiple pathways allow protein secretion across the bacterial outer membrane. AB - Secretion of proteins across the bacterial outer membrane takes place via a variety of mechanisms from simple one-component systems to complex multicomponent pathways. Secretion pathways can be organized into evolutionarily and functionally related groups, which highlight their relationship with organelle biogenesis. Recent work is beginning to reveal the structure and function of various secretion components and the molecular mechanisms of secretion. PMID- 10873831 TI - Regulators and effectors of the ARF GTPases. AB - The small G proteins of the ARF family are key regulators of membrane dynamics. Many functions of ARF proteins in cells are being revealed by studies of their regulators and effectors. Significant progress has been made over the past year, with the identification of a surprisingly large family of novel ARF GTPase activating proteins. In addition, two new classes of effectors, the PIP kinases and a novel family of monomeric coat-like proteins have been discovered. PMID- 10873832 TI - Sorting in the endosomal system in yeast and animal cells. AB - The endosomal system is a major membrane-sorting apparatus. New evidence reveals that novel coat proteins assist specific sorting steps and docking factors ensure the vectorial nature of trafficking in the endosomal compartment. There is also good evidence for ubiquitin regulating passage of certain proteins into multivesicular late endosomes, which mature by accumulating invaginated membrane. Lipids play a central role in this involution process, as do the class E vacuolar protein-sorting proteins. PMID- 10873833 TI - The structures of caspases-1, -3, -7 and -8 reveal the basis for substrate and inhibitor selectivity. AB - BACKGROUND: Peptide inhibitors of caspases have helped define the role of these cysteine proteases in biology. Structural and biochemical characterization of the caspase enzymes may contribute to the development of new drugs for the treatment of caspase-mediated inflammation and apoptosis. RESULTS: The crystal structure of the previously unpublished caspase-7 (Csp7; 2.35 A) bound to the reversible tetrapeptide aldehyde inhibitor acetyl-Asp-Glu-Val-Asp-CHO is compared with crystal structures of caspases-1 (2.3 A), -3 (2.2 A), and -8 (2.65 A) bound to the same inhibitor. Csp7 is a close homolog of caspase-3 (Csp3), and these two caspases possess some quarternary structural characteristics that support their unique role among the caspase family. However, although Csp3 and Csp7 are quite similar overall, they were found to have a significantly different substitution pattern of amino acids in and around the S4-binding site. CONCLUSIONS: These structures span all three caspase subgroups, and provide a basis for inferring substrate and inhibitor binding, as well as selectivity for the entire caspase family. This information will influence the design of selective caspase inhibitors to further elucidate the role of caspases in biology and hopefully lead to the design of therapeutic agents to treat caspase-mediated diseases, such as rheumatoid arthritis, certain neurogenerative diseases and stroke. PMID- 10873835 TI - Brevetoxin derivatives that inhibit toxin activity. AB - BACKGROUND: The brevetoxins are marine neurotoxins that interfere with the normal functions of the voltage-gated Na(+) channel. We have identified two brevetoxin derivatives that do not exhibit pharmacological properties typical of the brevetoxins and that function as brevetoxin antagonists. RESULTS: PbTx-3 and benzoyl-PbTx-3 elicited Na(+) channel openings during steady-state depolarizations; however, two PbTx-3 derivatives retained their ability to bind to the receptor, but did not elicit Na(+) channel openings. alpha-Naphthoyl-PbTx 3 acted as a PbTx-3 antagonist but did not affect Na(+) channels that were not exposed to PbTx-3. beta-Naphthoyl-PbTx-3 reduced openings of Na(+) channels that were not exposed to PbTx-3. CONCLUSIONS: Some modifications to the brevetoxin molecule do not alter either the binding properties or the activity of these toxins. Larger modifications to the K-ring sidechain do not interfere with binding but have profound effects on their pharmacological properties. This implies a critical function for the K-ring sidechain of the native toxin. PMID- 10873834 TI - Intracellular targets of cyclin-dependent kinase inhibitors: identification by affinity chromatography using immobilised inhibitors. AB - BACKGROUND: Chemical inhibitors of cyclin-dependent kinases (CDKs) have great therapeutic potential against various proliferative and neurodegenerative disorders. Olomoucine, a 2,6,9-trisubstituted purine, has been optimized for activity against CDK1/cyclin B by combinatorial and medicinal chemistry efforts to yield the purvalanol inhibitors. Although many studies support the action of purvalanols against CDKs, the actual intracellular targets of 2,6, 9 trisubstituted purines remain unverified. RESULTS: To address this issue, purvalanol B (95. ) and an N6-methylated, CDK-inactive derivative (95M. ) were immobilized on an agarose matrix. Extracts from a diverse collection of cell types and organisms were screened for proteins binding purvalanol B. In addition to validating CDKs as intracellular targets, a variety of unexpected protein kinases were recovered from the 95. matrix. Casein kinase 1 (CK1) was identified as a principal 95. matrix binding protein in Plasmodium falciparum, Leishmania mexicana, Toxoplasma gondii and Trypanosoma cruzi. Purvalanol compounds also inhibit the proliferation of these parasites, suggesting that CK1 is a valuable target for further screening with 2,6,9-trisubstituted purine libraries. CONCLUSIONS: That a simple batchwise affinity chromatography approach using two purine derivatives facilitated isolation of a small set of highly purified kinases suggests that this could be a general method for identifying intracellular targets relevant to a particular class of ligands. This method allows a close correlation to be established between the pattern of proteins bound to a small family of related compounds and the pattern of cellular responses to these compounds. PMID- 10873836 TI - Negative selectivity and the evolution of protease cascades: the specificity of plasmin for peptide and protein substrates. AB - BACKGROUND: Understanding the networks of selective proteolysis that regulate complex biological systems requires an appreciation of the molecular mechanisms used to maintain substrate specificity. Human plasmin, a serine protease that promotes the dissolution of blood clots and is essential in maintaining normal hemostasis, is usually described as having broad substrate specificity. Recent evidence that plasmin also plays a key role in a variety of other important biological and pathological processes, however, has suggested that this description might need to be re-evaluated. RESULTS: We used substrate phage display to elucidate optimal subsite occupancy for substrates of plasmin. We identified a peptide substrate that is cleaved 710,000-fold more efficiently by plasmin than a peptide containing the activation sequence of plasminogen. Plasmin achieves this unexpected, large differential activity even though both target sequences possess an arginine residue in the P1 position. We also demonstrate that proteolysis by plasmin can be targeted to an engineered protein substrate and that introduction of substrate sequences identified by phage display into plasminogen increases plasmin-mediated cleavage of the mutant 2000-fold. CONCLUSIONS: The specificity of plasmin is more tightly controlled than previously recognized; interactions with substrates at all subsites between S4 and S2' contribute to catalysis. Furthermore, in contrast to most enzymes that exhibit positive selectivity for substrate, the evolution of substrate specificity by plasmin has apparently been dominated by a strong negative selection against development of autoactivation activity. This 'negative selectivity' avoids short-circuiting regulation of the fibrinolytic system and other important biological processes, and might be an important general mechanism for controlling protease cascades. PMID- 10873837 TI - Tagging DNA mismatches by selective 2'-amine acylation. AB - BACKGROUND: Widespread characterization of genetic variation and disease at the gene-sequence level has inaugurated a new era in human biology. Techniques for the molecular analysis of these variations and their linkage with measurable phenotypes will profoundly affect diverse fields of biological chemistry and biology. RESULTS: A chemical tagging method has been developed to detect point mutations and other defects in nucleic acid sequences. The method employs oligodeoxynucleotide probes in which one 2'-ribose position (-H) is substituted with an amine (-NH(2)) group. 2'-Amine-substituted nucleotides are specifically acylated by succinimidyl esters to form a 2'-amide product. The mutation detection method exploits our observation that 2'-amine groups at the site of a mismatch are acylated more rapidly than amine substitutions at base-paired nucleotides. 2'-Amine acylation is governed primarily by local, rather than global, differences in nucleotide dynamics, such that site-specific tagging of DNA mismatches does not require discriminatory hybridization conditions to be determined. CONCLUSIONS: 2'-Amine mismatch tagging offers an approach for chemically interrogating the base-paired state of individual nucleotides in a hybridized duplex and for quantifying nucleicacid hybridization with single-base specificity. PMID- 10873838 TI - Simulated molecular evolution in a full combinatorial library. AB - BACKGROUND: The Darwinian concept of 'survival of the fittest' has inspired the development of evolutionary optimization methods to find molecules with desired properties in iterative feedback cycles of synthesis and testing. These methods have recently been applied to the computer-guided heuristic selection of molecules that bind with high affinity to a given biological target. We describe the optimization behavior and performance of genetic algorithms (GAs) that select molecules from a combinatorial library of potential thrombin inhibitors in 'artificial molecular evolution' experiments, on the basis of biological screening results. RESULTS: A full combinatorial library of 15,360 members structurally biased towards the serine protease thrombin was synthesized, and all were tested for their ability to inhibit the protease activity of thrombin. Using the resulting large structure-activity landscape, we simulated the evolutionary selection of potent thrombin inhibitors from this library using GAs. Optimal parameter sets were found (encoding strategy, population size, mutation and cross over rate) for this artificial molecular evolution. CONCLUSIONS: A GA-based evolutionary selection is a valuable combinatorial optimization strategy to discover compounds with desired properties without needing to synthesize and test all possible combinations (i.e. all molecules). GAs are especially powerful when dealing with very large combinatorial libraries for which synthesis and screening of all members is not possible and/or when only a small number of compounds compared with the library size can be synthesized or tested. The optimization gradient or 'learning' per individual increases when using smaller population sizes and decreases for higher mutation rates. PMID- 10873839 TI - Dipeptide formation on engineered hybrid peptide synthetases. AB - BACKGROUND: Nonribosomal peptide synthetases (NRPSs) are modular 'megaenzymes' that catalyze the assembly of a large number of bioactive peptides using the multiple carrier thiotemplate mechanism. The modules comprise specific domains that act as distinct units to catalyze specific reactions associated with substrate activation, modification and condensation. Such an arrangement of biosynthetic templates has evoked interest in engineering novel NRPSs. RESULTS: We describe the design and construction of a set of dimodular hybrid NRPSs. By introducing domain fusions between adenylation and thiolation (PCP) domains we designed synthetic templates for dipeptide formation. The predicted dipeptides, as defined by the specificity and arrangement of the adenylation domains of the constructed templates, were synthesized in vitro. The effect of the intramolecular fusion was investigated by determining kinetic parameters for substrate adenylation and thiolation. The rate of dipeptide formation on the artificial NRPSs is similar to that of natural templates. CONCLUSIONS: Several new aspects concerning the tolerance of NRPSs to domain swaps can be deduced. By choosing the fusion site in the border region of adenylation and PCP domains we showed that the PCP domain exhibits no general substrate selectivity. There was no suggestion that selectivity of the condensation reaction was biased towards the donor amino acid, whereas at the acceptor position there was a size determined selection. In addition, we demonstrated that a native elongation module can be converted to an initiation module for peptide-bond formation. These results represent the first example of rational de novo synthesis of small peptides on engineered NRPSs. PMID- 10873840 TI - The ribosome--a macromolecular machine par excellence. AB - The ribosome is the site in the cell where proteins are synthesized. Cryo electron microscopy and X-ray crystallography have revealed the ribosome as a particle made of two subunits, each formed as an intricate mesh of RNAs and many proteins. Ligand-binding experiments followed by cryo-electron microscopy have helped to determine some of the key stages of interaction between the ribosome and the main ligand molecules. PMID- 10873841 TI - Biosynthesis of the polyene antifungal antibiotic nystatin in Streptomyces noursei ATCC 11455: analysis of the gene cluster and deduction of the biosynthetic pathway. AB - BACKGROUND: The polyene macrolide antibiotic nystatin produced by Streptomyces noursei ATCC 11455 is an important antifungal agent. The nystatin molecule contains a polyketide moiety represented by a 38-membered macrolactone ring to which the deoxysugar mycosamine is attached. Molecular cloning and characterization of the genes governing the nystatin biosynthesis is of considerable interest because this information can be used for the generation of new antifungal antibiotics. RESULTS: A DNA region of 123,580 base pairs from the S. noursei ATCC 11455 genome was isolated, sequenced and shown by gene disruption to be involved in nystatin biosynthesis. Analysis of the DNA sequence resulted in identification of six genes encoding a modular polyketide synthase (PKS), genes for thioesterase, deoxysugar biosynthesis, modification, transport and regulatory proteins. One of the PKS-encoding genes, nysC, was found to encode the largest (11,096 amino acids long) modular PKS described to date. Analysis of the deduced gene products allowed us to propose a model for the nystatin biosynthetic pathway in S. noursei. CONCLUSIONS: A complete set of genes responsible for the biosynthesis of the antifungal polyene antibiotic nystatin in S. noursei ATCC 11455 has been cloned and analyzed. This represents the first example of the complete DNA sequence analysis of a polyene antibiotic biosynthetic gene cluster. Manipulation of the genes identified within the cluster may potentially lead to the generation of novel polyketides and yield improvements in the production strains. PMID- 10873842 TI - Resisting resistance: new chemical strategies for battling superbugs. AB - As microbes become increasingly resistant to antibiotics, and in many cases to several drugs simultaneously, the search is on to find new therapies. One method to combat resistance is to use inhibitors of resistance mechanisms to potentiate existing antibiotics. Recent efforts are encouraging and highlight the importance of research at the chemistry-microbiology interface in developing new approaches to tackle resistance. PMID- 10873843 TI - Nonhost resistance and nonspecific plant defenses. AB - In the past year, most of the advances in our understanding of nonhost resistance to plant pathogens have been incremental. Highlights include the discovery of a general bacterial elicitor of plant defenses, the description of more similarities between the hypersensitive response and animal programmed cell death, and a growing appreciation of the cell wall as the site of initiation and expression of nonhost resistance towards fungi. PMID- 10873844 TI - Structure, function and evolution of plant disease resistance genes. AB - Gene-for-gene plant disease resistance involves two basic processes: perception of pathogen attack, followed by responses to limit disease. Perception involves receptors with high degrees of specificity for pathogen strains, which are encoded by disease resistance genes. Large repertoires of distantly related resistance (R) genes with diverse recognitional specificities are found within a single plant species. The generation of R-gene polymorphism involves gene duplication, followed by DNA-sequence divergence by point mutation, and by deletion and duplication of intragenic DNA repeats encoding blocks of leucine rich elements. Recombination between related genes reassorts this variation to further diversify gene sequences. Pathogen pressure selects functional resistance specificities and results in the maintenance of R-gene diversity. Recent genome sequence data reveal that the NBS-LRR (i.e. nucleotide-binding site-leucine-rich repeat) class of R genes represents as much as 1% of the Arabidopsis genome. Experimental data have shown that the LRR has a role in determination of specificity. Mutation experiments, in which R-gene signaling has been dissociated from specificity in constitutive signal mutants, have provided the potential for non-specific resistance to be expressed from pathogen-infection-induced promoters in transgenic plants. PMID- 10873845 TI - Mechanisms, ecological consequences and agricultural implications of tri-trophic interactions. AB - Recent research bridging mechanistic and ecological approaches demonstrates that plant attributes can affect herbivores, natural enemies of herbivores, and their interaction. Such effects may be genetically variable among plants and/or induced in individual plants by herbivore attack, and are mediated by primary plant attributes (i.e. nutritional quality and physical structure) and defense-related products (i.e. secondary chemicals and plant volatiles), and may be modified by human activity (e.g. by the introduction of Bacillus thuringiensis). The study of tri-trophic interactions is important in order to understand natural species interactions and to manipulate these interactions in pest control. PMID- 10873847 TI - Intracellular accommodation of microbes by plants: a common developmental program for symbiosis and disease? AB - Plant cells engage in mutualistic and parasitic endosymbioses with a wide variety of microorganisms, ranging from Gram-negative (Rhizobium, Nostoc) and Gram positive bacteria (Frankia), to oomycetes (Phytophthora), Chytridiomycetes, Zygomycetes (arbuscular mycorrhizal fungi) and true fungi (Erysiphe, ascomycete; Puccinia, basidiomycete). Endosymbiosis is characterised by the 'symbiosome', a compartment within host cells in which the symbiotic microorganism is either partially or completely enclosed by a host-derived membrane. The analysis of plant mutants indicates that the genetic requirements for the interaction with rhizobia and arbuscular mycorrhiza fungi are partially overlapping. The extent to which plants use similar or identical developmental programs for the intracellular accommodation of different microorganisms is, however, not clear. For example, plant cells actively weaken their cell wall to facilitate bacterial colonisation, whereas penetration by fungal symbionts appears not to be assisted in this manner. Moreover, different transport requirements are imposed on the symbiotic interface of different interactions indicating that additional system specific components are likely to exist. PMID- 10873846 TI - Costs of resistance. AB - Studies of the reduction of fitness in plants expressing resistance characteristics have always been popular. New techniques for manipulating defense expression have recently resulted in a greater understanding of the mechanisms through which different types of resistance strategies produce costs, especially those costs associated with inducible defenses. PMID- 10873848 TI - The genetic architecture of resistance. AB - Plant resistance genes (R genes), especially the nucleotide binding site leucine rich repeat (NBS-LRR) family of sequences, have been extensively studied in terms of structural organization, sequence evolution and genome distribution. These studies indicate that NBS-LRR sequences can be split into two related groups that have distinct amino-acid motif organizations, evolutionary histories and signal transduction pathways. One NBS-LRR group, characterized by the presence of a Toll/interleukin receptor domain at the amino-terminal end, seems to be absent from the Poaceae. Phylogenetic analysis suggests that a small number of NBS-LRR sequences existed among ancient Angiosperms and that these ancestral sequences diversified after the separation into distinct taxonomic families. There are probably hundreds, perhaps thousands, of NBS-LRR sequences and other types of R gene-like sequences within a typical plant genome. These sequences frequently reside in 'mega-clusters' consisting of smaller clusters with several members each, all localized within a few million base pairs of one another. The organization of R-gene clusters highlights a tension between diversifying and conservative selection that may be relevant to gene families that are unrelated to disease resistance. PMID- 10873849 TI - Plant-pathogen arms races at the molecular level. AB - Advances in research into the genetics of plant-pathogen interactions include an embracing of evolutionary ideas and methodologies. Recent studies reveal positive selection and selective maintenance of variation in plant resistance and defense related genes. Coevolution between plants and their enemies involves many interactions at the molecular level. PMID- 10873850 TI - Prospects for understanding avirulence gene function. AB - Avirulence genes are originally defined by their negative impact on the ability of a pathogen to infect their host plant. Many avirulence genes are now known to represent a subset of virulence factors involved in the mediation of the host pathogen interaction. Characterization of avirulence genes has revealed that they encode an amazing assortment of proteins and belong to several gene families. Although the biochemical functions of the avirulence gene products are unknown, studies are beginning to reveal the features and interesting relationships between the avirulence and virulence activities of the proteins. Identification of critical virulence factors and elucidation of their functions promises to provide insight into plant defense mechanisms, and new and improved strategies for the control of plant disease. PMID- 10873851 TI - Trade-offs between pathogen and herbivore resistance. AB - During the past year genetic and pharmacological experiments have revealed a molecular basis for the cross-talk between signaling pathways mediating pathogen and herbivore resistance. These findings provide considerable insight into the apparently contradictory results reported for trade-offs between pathogen and herbivore resistance. PMID- 10873852 TI - Insect transmission of plant viruses: a constraint on virus variability. AB - Genetic diversity in viruses is shaped by high rates of recombination and is constrained by host defenses and the requirements of transmission. Recent studies of insect-transmitted plant viruses demonstrate highly conserved molecular motifs in viral genomes that regulate the specificity of insect transmission. In contrast, advances in our understanding of host plant response to virus infection reveal some generalized patterns of host defense to a diversity of viruses. PMID- 10873853 TI - Insights into the phosphoryltransfer mechanism of human thymidylate kinase gained from crystal structures of enzyme complexes along the reaction coordinate. AB - BACKGROUND: Thymidylate kinase (TMPK) is a nucleoside monophosphate kinase that catalyzes the reversible phosphoryltransfer between ATP and TMP to yield ADP and TDP. In addition to its vital role in supplying precursors for DNA synthesis, human TMPK has an important medical role participating in the activation of a number of anti-HIV prodrugs. RESULTS: Crystal structures of human TMPK in complex with TMP and ADP, TMP and the ATP analog AppNHp, TMP with ADP and the phosphoryl analog AlF(3), TDP and ADP, and the bisubstrate analog TP(5)A were determined. The conformations of the P-loop, the LID region, and the adenine-binding loop vary according to the nature of the complex. Substitution of ADP by AppNHp results in partial closure of the P-loop and the rotation of the TMP phosphate group to a catalytically unfavorable position, which rotates back in the AlF(3) complex to a position suitable for in-line attack. In the fully closed state observed in the TP(5)A and the TDP-ADP complexes, Asp15 interacts strongly with the 3'-hydroxyl group of TMP. CONCLUSIONS: The observed changes of nucleotide state and conformation and the corresponding protein structural changes are correlated with intermediates occurring along the reaction coordinate and show the sequence of events occurring during phosphate transfer. The low catalytic activity of human TMPK appears to be determined by structural changes required to achieve catalytic competence and it is suggested that a mechanism might exist to accelerate the activity. PMID- 10873854 TI - Structure of bovine mitochondrial F(1)-ATPase inhibited by Mg(2+) ADP and aluminium fluoride. AB - BACKGROUND: The globular domain of the membrane-associated F(1)F(o)-ATP synthase complex can be detached intact as a water-soluble fragment known as F(1)-ATPase. It consists of five different subunits, alpha, beta, gamma, delta and epsilon, assembled with the stoichiometry 3:3:1:1:1. In the crystal structure of bovine F(1)-ATPase determined previously at 2.8 A resolution, the three catalytic beta subunits and the three noncatalytic alpha subunits are arranged alternately around a central alpha-helical coiled coil in the gamma subunit. In the crystals, the catalytic sites have different nucleotide occupancies. One contains the triphosphate form of the nucleotide, the second contains the diphosphate, and the third is unoccupied. Fluoroaluminate complexes have been shown to mimic the transition state in several ATP and GTP hydrolases. In order to understand more about its catalytic mechanism, F(1)-ATPase was inhibited with Mg(2+)ADP and aluminium fluoride and the structure of the inhibited complex was determined by X ray crystallography. RESULTS: The structure of bovine F(1)-ATPase inhibited with Mg(2+)ADP and aluminium fluoride determined at 2.5 A resolution differs little from the original structure with bound AMP-PNP and ADP. The nucleotide occupancies of the alpha and beta subunits are unchanged except that both aluminium trifluoride and Mg(2+)ADP are bound in the nucleotide-binding site of the beta(DP) subunit. The presence of aluminium fluoride is accompanied by only minor adjustments in the surrounding protein. CONCLUSIONS: The structure appears to mimic a possible transition state. The coordination of the aluminofluoride group has many features in common with other aluminofluoride-NTP hydrolase complexes. Apparently, once nucleotide is bound to the catalytic beta subunit, no additional major structural changes are required for catalysis to occur. PMID- 10873855 TI - Crystal structure of decameric 2-Cys peroxiredoxin from human erythrocytes at 1.7 A resolution. AB - BACKGROUND: The peroxiredoxins (Prxs) are an emerging family of multifunctional enzymes that exhibit peroxidase activity in vitro, and in vivo participate in a range of cellular processes known to be sensitive to reactive oxygen species. Thioredoxin peroxidase B (TPx-B), a 2-Cys type II Prx from erythrocytes, promotes potassium efflux and down-regulates apoptosis and the recruitment of monocytes by endothelial tissue. RESULTS: The crystal structure of human decameric TPx-B purified from erythrocytes has been determined to 1.7 [corrected)] A resolution. The structure is a toroid comprising five dimers linked end-on through predominantly hydrophobic interactions, and is proposed to represent an intermediate in the in vivo reaction cycle. In the crystal structure, Cys51, the site of peroxide reduction, is oxidised to cysteine sulphinic acid. The residue Cys172, lies approximately 10 A away from Cys51 [corrected]. CONCLUSIONS: The oxidation of Cys51 appears to have trapped the structure into a stable decamer, as confirmed by sedimentation analysis. A comparison with two previously reported dimeric Prx structures reveals that the catalytic cycle of 2-Cys Prx requires significant conformational changes that include the unwinding of the active-site helix and the movement of four loops. It is proposed that the stable decamer forms in vivo under conditions of oxidative stress. Similar decameric structures of TPx-B have been observed by electron microscopy, which show the protein associated with the erythrocyte membrane. PMID- 10873856 TI - A new allosteric site in glycogen phosphorylase b as a target for drug interactions. AB - BACKGROUND: In muscle and liver, glycogen concentrations are regulated by the coordinated activities of glycogen phosphorylase (GP) and glycogen synthase. GP exists in two forms: the dephosphorylated low-activity form GPb and the phosphorylated high-activity form GPa. In both forms, allosteric effectors can promote equilibrium between a less active T state and a more active R state. GP is a possible target for drugs that aim to prevent unwanted glycogen breakdown and to stimulate glycogen synthesis in non-insulin-dependent diabetes. As a result of a data bank search, 5-chloro-1H-indole-2-carboxylic acid (1-(4 fluorobenzyl)-2-(4-hydroxypiperidin-1-yl)-2-oxoethy l)amide, CP320626, was identified as a potent inhibitor of human liver GP. Structural studies have been carried out in order to establish the mechanism of this unusual inhibitor. RESULTS: The structure of the cocrystallised GPb-CP320626 complex has been determined to 2.3 A resolution. CP320626 binds at a site located at the subunit interface in the region of the central cavity of the dimeric structure. The site has not previously been observed to bind ligands and is some 15 A from the AMP allosteric site and 33 A from the catalytic site. The contacts between GPb and CP320626 comprise six hydrogen bonds and extensive van der Waals interactions that create a tight binding site in the T-state conformation of GPb. In the R state conformation of GPa these interactions are significantly diminished. CONCLUSIONS: CP320626 inhibits GPb by binding at a new allosteric site. Although over 30 A from the catalytic site, the inhibitor exerts its effects by stabilising the T state at the expense of the R state and thereby shifting the allosteric equilibrium between the two states. The new allosteric binding site offers a further recognition site in the search for improved GP inhibitors. PMID- 10873857 TI - Structure at 2.3 A resolution of the cytochrome bc(1) complex from the yeast Saccharomyces cerevisiae co-crystallized with an antibody Fv fragment. AB - BACKGROUND: The cytochrome bc(1) complex is part of the energy conversion machinery of the respiratory and photosynthetic electron transfer chains. This integral membrane protein complex catalyzes electron transfer from ubiquinol to cytochrome c. It couples the electron transfer to the electrogenic translocation of protons across the membrane via a so-called Q cycle mechanism. RESULTS: The cytochrome bc(1) complex from the yeast Saccharomyces cerevisiae was crystallized together with a bound antibody Fv fragment. The structure was determined at 2.3 A resolution using multiple isomorphous replacement, and refined to a crystallographic R factor of 22.2% (R(free) = 25.4%). The complex is present as a homodimer. Each 'monomer' of the refined model includes 2178 amino acid residues of subunits COR1, QCR2, COB, CYT1, RIP1, QCR6, QCR7, QCR8 and QCR9 of the cytochrome bc(1) complex and of the polypeptides V(H) and V(L) of the Fv fragment, the cofactors heme b(H), heme b(L), heme c(1), the [2Fe-2S] cluster and 346 water molecules. The Fv fragment binds to the extrinsic domain of the [2Fe 2S] Rieske protein and is essential for formation of the crystal lattice. CONCLUSIONS: The approach to crystallize membrane proteins as complexes with specific antibody fragments appears to be of general importance. The structure of the yeast cytochrome bc(1) complex reveals in detail the binding sites of the natural substrate coenzyme Q6 and the inhibitor stigmatellin. Buried water molecules close to the binding sites suggest possible pathways for proton uptake and release. A comparison with other cytochrome bc(1) complexes shows features that are specific to yeast. PMID- 10873858 TI - Re-engineering ketoacyl synthase specificity. PMID- 10873860 TI - The third dimension for biologists PMID- 10873859 TI - A conserved structural motif for lipopolysaccharide recognition by procaryotic and eucaryotic proteins. AB - BACKGROUND: Lipopolysaccharide (LPS), a lipoglycan from the outer membrane of Gram-negative bacteria, is an immunomodulatory molecule that stimulates the innate immune response. High levels of LPS cause excessive release of inflammatory mediators and are responsible for the septic shock syndrome. The interaction of LPS with its cognate binding proteins has not, as yet, been structurally elucidated. RESULTS: The X-ray crystallographic structure of LPS in complex with the integral outer membrane protein FhuA from Escherichia coli K-12 is reported. It is in accord with data obtained using mass spectroscopy and nuclear magnetic resonance. Most of the important hydrogen-bonding or electrostatic interactions with LPS are provided by eight positively charged residues of FhuA. Residues in a similar three-dimensional arrangement were searched for in all structurally known proteins using a fast template-matching algorithm, and a subset of four residues was identified that is common to known LPS-binding proteins. CONCLUSIONS: These four residues, three of which form specific interactions with lipid A, appear to provide the structural basis of pattern recognition in the innate immune response. Their arrangement can serve to identify LPS-binding sites on proteins known to interact with LPS, and could serve as a template for molecular modeling of a LPS scavenger designed to reduce the septic shock syndrome. PMID- 10873861 TI - Crystal structure of Urtica dioica agglutinin, a superantigen presented by MHC molecules of class I and class II. AB - BACKGROUND: Urtica dioica agglutinin (UDA), a monomeric lectin extracted from stinging nettle rhizomes, is specific for saccharides containing N acetylglucosamine (GlcNAc). The lectin behaves as a superantigen for murine T cells, inducing the exclusive proliferation of Vbeta8.3(+) lymphocytes. UDA is unique among known T cell superantigens because it can be presented by major histocompatibility complex (MHC) molecules of both class I and II. RESULTS: The crystal structure of UDA has been determined in the ligand-free state, and in complex with tri-acetylchitotriose and tetra-acetylchitotetraose at 1.66 A, 1.90 A and 1.40 A resolution, respectively. UDA comprises two hevein-like domains, each with a saccharide-binding site. A serine and three aromatic residues at each site form the principal contacts with the ligand. The N-terminal domain binding site can centre on any residue of a chito-oligosaccharide, whereas that of the C terminal domain is specific for residues at the nonreducing terminus of the ligand. We have shown previously that oligomers of GlcNAc inhibit the superantigenic activity of UDA and that the lectin binds to glycans on the MHC molecule. We show that UDA also binds to glycans on the T cell receptor (TCR). CONCLUSIONS: The presence of two saccharide-binding sites observed in the structure of UDA suggests that its superantigenic properties arise from the simultaneous fixation of glycans on the TCR and MHC molecules of the T cell and antigen-presenting cell, respectively. The well defined spacing between the two binding sites of UDA is probably a key factor in determining the specificity for Vbeta8.3(+) lymphocytes. PMID- 10873862 TI - The first crystal structure of a phospholipase D. AB - BACKGROUND: The phospholipase D (PLD) superfamily includes enzymes that are involved in phospholipid metabolism, nucleases, toxins and virus envelope proteins of unknown function. PLD hydrolyzes the terminal phosphodiester bond of phospholipids to phosphatidic acid and a hydrophilic constituent. Phosphatidic acid is a compound that is heavily involved in signal transduction. PLD also catalyses a transphosphatidylation reaction in the presence of phosphatidylcholine and a short-chained primary or secondary alcohol. RESULTS: The first crystal structure of a 54 kDa PLD has been determined to 1.9 A resolution using the multiwavelength anomalous dispersion (MAD) method on a single WO(4) ion and refined to 1.4 A resolution. PLD from the bacterial source Streptomyces sp. strain PMF consists of a single polypeptide chain that is folded into two domains. An active site is located at the interface between these domains. The presented structure supports the proposed superfamily relationship with the published structure of the 16 kDa endonuclease from Salmonella typhimurium. CONCLUSIONS: The structure of PLD provides insight into the structure and mode of action of not only bacterial, plant and mammalian PLDs, but also of a variety of enzymes as diverse as cardiolipin synthases, phosphatidylserine synthases, toxins, endonucleases, as well as poxvirus envelope proteins having a so far unknown function. The common features of these enzymes are that they can bind to a phosphodiester moiety, and that most of these enzymes are active as bi-lobed monomers or dimers. PMID- 10873863 TI - Structure and self-association of the Rous sarcoma virus capsid protein. AB - BACKGROUND: The capsid protein (CA) of retroviruses, such as Rous sarcoma virus (RSV), consists of two independently folded domains. CA functions as part of a polyprotein during particle assembly and budding and, in addition, forms a shell encapsidating the genomic RNA in the mature, infectious virus. RESULTS: The structures of the N- and C-terminal domains of RSV CA have been determined by X ray crystallography and solution nuclear magnetic resonance (NMR) spectroscopy, respectively. The N-terminal domain comprises seven alpha helices and a short beta hairpin at the N terminus. The N-terminal domain associates through a small, tightly packed, twofold symmetric interface within the crystal, different from those previously described for other retroviral CAs. The C-terminal domain is a compact bundle of four alpha helices, although the last few residues are disordered. In dilute solution, RSV CA is predominantly monomeric. We show, however, using electron microscopy, that intact RSV CA can assemble in vitro to form both tubular structures constructed from toroidal oligomers and planar monolayers. Both modes of assembly occur under similar solution conditions, and both sheets and tubes exhibit long-range order. CONCLUSIONS: The tertiary structure of CA is conserved across the major retroviral genera, yet sequence variations are sufficient to cause change in associative behavior. CA forms the exterior shell of the viral core in all mature retroviruses. However, the core morphology differs between viruses. Consistent with this observation, we find that the capsid proteins of RSV and human immunodeficiency virus type 1 exhibit different associative behavior in dilute solution and assemble in vitro into different structures. PMID- 10873864 TI - Conformations of the rhodopsin third cytoplasmic loop grafted onto bacteriorhodopsin. AB - BACKGROUND: The third cytoplasmic loop of rhodopsin (Rho EF) is important in signal transduction from the retinal in rhodopsin to its G protein, transducin. This loop also interacts with rhodopsin kinase, which phosphorylates light activated rhodopsin, and arrestin, which displaces transducin from light activated phosphorylated rhodopsin. RESULTS: We replaced eight residues of the EF loop of bacteriorhodopsin (BR) with 24 residues from the third cytoplasmic loop of bovine Rho EF. The surfaces of purple membrane containing the mutant BR (called IIIN) were imaged by atomic force microscopy (AFM) under physiological conditions to a resolution of 0.5-0.7 nm. The crystallinity and extracellular surface of IIIN were not perturbed, and the cytoplasmic surface of IIIN increased in height compared with BR, consistent with the larger loop. Ten residues of Rho EF were excised by V8 protease, revealing helices E and F in the AFM topographs. Rho EF was modeled onto the BR structure, and the envelope derived from the AFM data of IIIN was used to select probable models. CONCLUSIONS: A likely conformation of Rho EF involves some extension of helices E and F, with the tip of the loop lying over helix C and projecting towards the C terminus. This is consistent with mutagenesis data showing the TTQ transducin-binding motif close to loop CD, and cysteine cross-linking data indicating the C-terminal part of Rho EF to be close to the CD loop. PMID- 10873865 TI - Integrin-collagen complex: a metal-glutamate handshake. AB - The recently determined crystal structure of the complex between an integrin I domain and a synthetic collagen peptide shows a collagen triple-helix engaged in specific macromolecular recognition and binding. This structure confirms a previously proposed binding mechanism for integrin I domains and has important implications for integrin activation and signalling. PMID- 10873866 TI - Methods used to evaluate the quality of evidence underlying the National Kidney Foundation-Dialysis Outcomes Quality Initiative Clinical Practice Guidelines: description, findings, and implications. AB - This report describes the approach the National Kidney Foundation-Dialysis Outcomes Quality Initiative (NKF-DOQI) used to assess the strength of published evidence pertinent to individual NKF-DOQI Clinical Practice Guidelines, as well as the relationship between that approach and methods used by the US Preventive Services Task Force, the Cochrane Collaboration, and the Agency for Health Care Policy and Research to rate the quality and/or strength of evidence. We also present the results of an analysis of the strength of evidence underlying the NKF DOQI Guidelines showing that one cannot infer the quality of evidence reported in a study (rated either on a 0-to-1 scale or categorically as excellent, very good, good, fair, or poor) simply by knowing the type of study design used (randomized trial, nonrandomized trial, natural experiment, cohort study, cross-sectional study, case-control study, case report). Issues related to assessment of the strength of evidence underlying a practice guideline opposed to that reported in an individual study are highlighted. PMID- 10873867 TI - Economic evaluation and end-stage renal disease: from basics to bedside. AB - Economic evaluation is the comparative analysis of alternative health care interventions in terms of their relative costs (resource use) and effectiveness (health effects). High-quality studies of economic evaluation have been increasingly published in medical journals and read by clinicians, although publication of these studies in nephrology journals has been a more recent phenomenon. This article shows how the basic principles of economics can be applied to health care through the use of economic evaluation. Different types of economic evaluation are discussed, and pitfalls common to such studies are identified. A simple framework is introduced that can be used to interpret the results of economic evaluations. Using this framework, selected therapies for patients with end-stage renal disease (ESRD) are categorized to highlight therapies that are very efficient, encourage their use, and draw attention to therapies in current use that are less effective and more expensive (ie, less efficient) than alternative therapy. Using examples pertinent to care of the patient with ESRD, we show how economic evaluation can be used to link medical outcomes, quality of life, and costs in a common index for multiple therapies with disparate outcome measures. This article highlights the need for clinical studies and economic evaluations of therapies in ESRD for which the effects of the therapy on health outcomes and/or costs are unknown. PMID- 10873868 TI - Serum cystatin C as a new marker for noninvasive estimation of glomerular filtration rate and as a marker for early renal impairment. AB - Cystatin C is a nonglycosylated basic protein produced at a constant rate by all investigated nucleated cells. It is freely filtered by the renal glomeruli and primarily catabolized in the tubuli (not secreted or reabsorbed as an intact molecule). Because serum cystatin C concentration is independent of age, sex, and muscle mass, it has been postulated to be an improved marker of glomerular filtration rate (GFR) compared with serum creatinine level. We compared serum cystatin C level with other markers of GFR, such as serum creatinine level and creatinine clearance, and analyzed their variations based on iothalamate labeled with iodine 125 ((125)I-iothalamate) clearance ((125)I-ICl), used as the gold standard for GFR. The concentrations of the two different markers of GFR in patients with impaired renal function were classified according to (125)I-ICl. Twenty individuals with normal renal function ((125)I-ICl, 128 +/- 23 mL/min/1.73 m(2)) were used as the control group. Serum cystatin C level showed a greater sensitivity (93.4%) than serum creatinine level (86.8%). Also, serum cystatin C showed the greatest proportion of increased values in patients with impaired renal function (100%) compared with serum creatinine level (92.15%). Serum cystatin C levels started to increase to greater than normal values when GFR was 88 mL/min/1.73 m(2), whereas serum creatinine level began to increase when GFR was 75 mL/min/1.73 m(2). These data suggest that measurement of serum cystatin C may be useful to estimate GFR, especially to detect mild reductions in GFR, and therefore may be important in the detection of early renal insufficiency in a variety of renal diseases for which early treatment is critical. PMID- 10873869 TI - Timing of nephrology referral: influence on mortality and morbidity. AB - To assess the influence of the timing of nephrology referral on the short- and long-term outcome of hemodialysis patients, we retrospectively studied 309 patients who had end-stage renal failure and entered the chronic hemodialysis program in Sainte-Marguerite University Hospital between January 1, 1989, and December 31, 1996. We excluded from the analysis five patients without available data on referral pattern and 34 patients with irreversible acute renal failure. Of the remaining 270 patients, 177 patients (58%) had an early referral (ER) 16 or more weeks before the start of dialysis, and 93 patients (31%) had a late referral (LR) of less than 16 weeks before dialysis. Short-time morbidity (initial emergent dialysis, pulmonary edema, severe hypertension, temporary vascular access placement for first dialysis, prolonged initial hospitalization) was significantly more frequent in LR patients. Long-term evolution (mean follow up, 26.5 +/- 26 months) did not differ between the two groups. The number of days of hospitalization per patient-year at risk beyond the third month was 21.5 +/- 33.7 days for ER and 21.1 +/- 36 days for LR patients. Survival analysis showed no difference between the two groups: 3-month survival rates were 96% in both groups, 1-year survival rates were 90% in the ER and 89% in the LR group, and 5 year survival rates were 52% in the ER and 56% in the LR group. In a Cox hazards regression model, referral pattern was not associated with a greater risk for death. In conclusion, delayed nephrology referral generated strikingly greater initial morbidity, but long-term outcome of hemodialysis patients was not modified by delayed nephrological care. PMID- 10873870 TI - Platelet-activating factor acetylhydrolase gene mutation in Japanese children with Escherichia coli O157-associated hemolytic uremic syndrome. AB - Platelet-activating factor (PAF) may be involved in the pathogenesis of Escherichia coli O157-associated hemolytic uremic syndrome (HUS). PAF is degraded to inactive products by PAF acetylhydrolase. In this study, we investigated whether a PAF acetylhydrolase gene mutation (G-->T transversion at position 994) is involved in HUS in Japanese children. A point mutation in the PAF acetylhydrolase gene (G994T) was identified using polymerase chain reaction in 50 Japanese children with E coli O157-associated HUS and 100 healthy Japanese. We then determined the relationship between the PAF acetylhydrolase G994T gene mutation and clinical features of HUS. There was no difference in genotype and allele frequencies between patients with HUS and healthy controls. The mean duration of oligoanuria was significantly longer in patients with the GT genotype than in those with the GG genotype (P = 0.012). Although 11 of 15 patients (73%) heterozygous for the mutant allele (GT) required dialysis, only 13 of the 35 wild type homozygotes (GG; 37%) required dialysis (P = 0. 030). Mean plasma PAF acetylhydrolase activity was significantly less in patients with the GT genotype than in those with the GG genotype (P < 0.0001). In conclusion, we have shown an association between the G994T PAF acetylhydrolase gene mutation and the severity of renal damage in E coli O157-associated HUS. Our study suggests that analysis of the PAF acetylhydrolase gene mutation in Japanese children with E coli O157 associated HUS may allow the prediction of the severity of HUS. PMID- 10873871 TI - Haemophilus parainfluenzae antigen and antibody in children with IgA nephropathy and Henoch-Schonlein nephritis. AB - Although the pathogenesis of immunoglobulin A (IgA) nephropathy and Henoch Schonlein nephritis (HSN) remains uncertain, there is substantial evidence that they are immune complex-mediated diseases. Recently, Haemophilus parainfluenzae antigens were shown in the glomerular mesangium of adult patients with IgA nephropathy, and greater levels of IgA antibody against H parainfluenzae were also shown in the sera of adult patients with IgA nephropathy. The present study was performed to detect H parainfluenzae antigens and antibody against H parainfluenzae in children with IgA nephropathy and HSN. H parainfluenzae antigens in the mesangium were examined by indirect immunofluorescence, and antibody against H parainfluenzae was examined by enzyme-linked immunosorbent assay. Diffuse global staining of the mesangium with rabbit antisera against H parainfluenzae was shown in 10 of the 32 patients (31%) with IgA nephropathy and 12 of the 34 patients (35%) with HSN. Conversely, only 2 of the 47 patients (4%) with other renal diseases showed staining of glomeruli with rabbit antisera against H parainfluenzae (IgA nephropathy versus other renal diseases, P = 0.003; HSN versus other renal diseases, P = 0.0006). Patients with IgA nephropathy and those with HSN showed significantly greater levels of plasma IgA1 antibody against H parainfluenzae than patients with other renal diseases (IgA nephropathy versus other renal diseases, P = 0.008; HSN versus other renal diseases, P = 0.025). These findings suggest that H parainfluenzae has a role in the cause of these two conditions in a subset of patients. PMID- 10873872 TI - Anatomic and metabolic risk factors for nephrolithiasis in patients with autosomal dominant polycystic kidney disease. AB - The prevalence of nephrolithiasis is considerably greater in patients with autosomal dominant polycystic kidney disease (ADPKD) than in the general population. We evaluated anatomic and metabolic factors that may be associated with an increased prevalence of nephrolithiasis in patients with ADPKD. We compared anatomic parameters among ADPKD patients with or without nephrolithiasis as diagnosed by ultrasonography, whereas metabolic factors were determined by 24 hour urinary chemical analysis. Patients with ADPKD and nephrolithiasis had more renal cysts (P < 0.05) and a larger predominant renal cyst size (P < 0.005) than patients without nephrolithiasis. Concurrently, individual stone-forming kidneys had a greater cyst number (P < 0.05) and a significantly larger predominant cyst size (P < 0.01) compared with kidneys without stones. Patients with ADPKD and nephrolithiasis had a significantly lower creatinine clearance than those without nephrolithiasis (68.7 +/- 8.6 versus 94.8 +/- 5.4 mL/min, respectively; P < 0.05). Twenty-four-hour urinary analysis showed that patients with ADPKD and nephrolithiasis had significantly lower urinary volumes (P < 0. 05), and levels of urinary phosphate (P < 0.05), magnesium (P < 0. 005), and potassium (P < 0.05). Although not statistically significant, patients with ADPKD with stones tended to have lower levels of urinary citrate, and both groups showed a high percentage (range, 49% to 60%) of patients with hypocitraturia. Our data are consistent with the hypothesis that patients with ADPKD who develop nephrolithiasis do so because of increased intrarenal anatomic obstruction, as well as lower levels of such urinary inhibitors of stones as magnesium and citrate. PMID- 10873873 TI - Hybrid dialysis: recirculation peritoneal dialysis revisited. AB - In a crossover trial, eight patients were studied during one treatment each of automated peritoneal dialysis (APD) and hybrid dialysis (HyD). During HyD, a fixed quantity of peritoneal dialysis fluid (PDF) was continuously removed at a flow rate of 141.3 +/- 23. 7 mL/min, dialyzed against the secondary dialysate (250 +/- 53.5 mL/min) generated by the hemodialysis delivery system with single needle dialysis capability, and the regenerated PDF (PDF(HyD)) was reinfused into the peritoneal cavity. Despite using a smaller volume (6,195 +/- 737 versus 13,321 +/- 1,201 mL; P < 0. 0001) of PDF(HyD) with a lower glucose concentration (729 +/- 562 versus 1,659 +/- 373 mg/dL; P < 0.0001) and osmolality (331 +/- 79 versus 387 +/- 184 mOsm/kg; P < 0.001) during HyD compared with APD (PDF(APD)), weight loss was similar with both treatments (1.4 +/- 1. 0 versus 1.6 +/- 1.2 kg). Lactate levels were lower (3.2 +/- 2.5 versus 11.4 +/- 5.4 mEq/L), but pH (7.5 +/- 1.3 versus 5.6 +/- 0.9; P < 0.001) and bicarbonate concentration (22.6 +/- 8.0 versus 11.9 +/- 7.9 mEq/L; P < 0.0001) were greater in PDF(HyD) than PDF(APD). Although the mean dialysate calcium level was lower (6.0 +/- 0.5 versus 6.9 +/- 1.1 mg/dL; P < 0.001) in PDF(HyD), it was more stable throughout the dialysis compared with PDF(APD). A steeper concentration gradient between the blood and dialysate resulted in greater clearance of urea (26.5 +/- 9.1 versus 11.0 +/- 4.7 mL/min; P = 0.04), creatinine (24.1 +/- 11.4 versus 12.0 +/- 7.9 mL/min; P = 0.03), phosphate (19.2 +/- 4.3 versus 9.8 +/- 7.2 mL/min; P = 0.01), and uric acid (15.6 +/- 6.9 versus 9.1 +/- 2.7 mL/min; P = 0.04) and a greater percentage of reduction in values for blood urea nitrogen (20.7% +/- 7.7% versus 11.6% +/- 5.5%; P = 0.02), serum creatinine (16.1% +/- 5.3% versus 6.6% +/- 3.0%; P < 0.001), phosphate (22.7% +/- 8.9% versus 9.8% +/- 4.5%; P = 0.004), and uric acid (15.8% +/- 2.9% versus 6.3% +/- 3.4%; P < 0.001) during HyD than APD. To conclude, HyD is a novel dialytic technique that uses biocompatible bicarbonate based dialysate to achieve excellent clearance of uremic toxins and ultrafiltration with minimal glucose load. PMID- 10873874 TI - Natural history of arteriovenous grafts in hemodialysis patients. AB - Most hemodialysis patients in the United States have an arteriovenous graft as their vascular access. Grafts have a relatively short life span and are prone to recurrent stenosis and thrombosis, requiring multiple salvage procedures to maintain their patency. There is little information in the literature regarding the clinical factors that determine graft survival and complications. We evaluated prospectively the outcomes of 256 grafts placed at a single institution during a 2-year period. A salvage procedure to maintain graft patency (thrombectomy, angioplasty, or surgical revision) was required in 29% of the grafts at 3 months, 52% at 6 months, 77% at 12 months, and 96% at 24 months. Thus, primary graft survival (time from graft placement to the first intervention) was only 23% at 1 year and 4% at 2 years. Primary graft survival was significantly less among patients with hypoalbuminemia compared with patients with a normal serum albumin level (P = 0.003). Secondary graft survival (time from graft placement to permanent graft failure) was 65% at 1 year and 51% at 2 years. Neither primary nor secondary graft survival was significantly correlated with patient age, sex, diabetic status, body mass index, or graft site. A mean of 1.22 interventions per graft-year were required to maintain access patency, including 0.51 thrombectomies, 0.54 angioplasties, and 0.17 surgical revisions. In conclusion, hypoalbuminemia is a strong predictor of the requirement for an early graft intervention. Patients with hypoalbuminemia may require a heightened index of suspicion in monitoring their grafts for evidence of stenosis. PMID- 10873875 TI - Efficacy of tissue plasminogen activator administration on patency of hemodialysis access catheters. AB - Patients with end-stage renal disease use hemodialysis catheters for either temporary or permanent blood access. Recurrent thrombosis and fibrin sheath formation are common causes of poor or inadequate blood flow rates that require intervention. We studied the effect of tissue plasminogen activator (tPA) in reestablishing adequate blood flow rates through nonfunctional vascular catheters in 22 consecutive chronic hemodialysis patients. From January 1, 1999, to May 20, 1999, there were 56 instances in which tPA was used in an attempt to improve blood flow rates. In all instances, 2 mg of tPA was infused into each port of a dual-lumen internal jugular catheter. Dwell time ranged between 2 and 96 hours (median, 24 hours), and patient follow-up ranged between 47 and 140 days (median, 133.5 days). tPA was effective in establishing adequate blood flow rates (>/=200 mL/min) during the next dialysis session in 49 of 56 cases (87.5%). Seven additional interventions were required because of early or late tPA failure (one fibrin sheath stripping, one catheter replacement for kinking, one catheter replacement for central venous stenosis, and four catheter replacements for persistently poor blood flow rates), and eight catheters were replaced for infection. Thus, further interventions to achieve adequate blood flow rates were required in 12.5% of the cases because of early or late tPA failure. tPA appears to be as effective as urokinase for reestablishing adequate blood flow rates through hemodialysis catheters that are thrombosed or have low blood flow rates. PMID- 10873876 TI - Antiplatelet therapy alters iron requirements in hemodialysis patients. AB - Hemodialysis (HD) patients are prone to develop iron deficiency because of consumption of iron stores during erythropoietin (EPO) therapy. Data are needed to establish the factors involved in the different iron needs among these patients. Sixty-five HD patients were prospectively studied during a year. The subjects were dialyzed through polytetrafluoroethylene (PTFE) grafts (n = 23), arteriovenous native fistulae (n = 41), and a Permcath (n = 1). Twenty-four patients were administered aspirin; 23 patients, ticlopidine; 1 patient, dipyridamole; and 4 patients, anticoagulation with acenocoumarol. Iron supplementation (oral or parenteral) and laboratory parameters were recorded monthly. Significant differences in iron requirements, depending on the use of antiplatelet and/or anticoagulation agents, were found. Total parenteral iron supplements were greater in patients on antiplatelet therapy with either native or graft vascular accesses compared with the rest (2,406 +/- 1,445 versus 1,562 +/- 858 mg; P = 0.0081). Twelve of 52 patients on antiplatelet therapy required oral iron and only 1 of 13 patients not on antiplatelet therapy was administered oral iron supplements (P < 0.05). Patients on antiplatelet therapy were administered more transfusions (1.9 +/- 3.8 transfusions/y) than individuals not on antiplatelet therapy (0.15 +/- 0.3 transfusions/y; P = 0.0015). However, only patients with PTFE grafts on antiplatelet therapy had a post-HD bleeding time longer than patients not on antiplatelet therapy (9.1 +/- 3.6 versus 5.7 +/- 3.9 minutes; P < 0.0001). Multiple logistic regression analysis showed that the use of antiplatelet agents (P < 0.05) is an independent factor that increased the probability of requiring greater parenteral iron supplements (>2.5 g/y). Patients with PTFE grafts required more EPO than those with autologous fistulae (160 +/- 93 versus 100 +/- 63 U/kg/wk; P = 0.012). No differences between groups were found that could explain this finding. Antiplatelet and/or anticoagulation therapy implied the use of greater amounts of iron supplements in HD patients. Although these greater requirements of iron occurred in parallel with bleeding from the vascular access, additional data favor the existence of other factors, eg, interdialytic blood losses. The present study suggests that antiplatelet therapy may be an important factor in determining iron requirements in HD patients. Moreover, our data relate for the first time the use of prosthetic grafts with increased EPO requirements, an issue of great potential importance in the debate about vascular access policy in dialysis units. PMID- 10873877 TI - Safety and efficacy of iron sucrose in patients sensitive to iron dextran: North American clinical trial. AB - Sensitivity to iron dextran is a potent obstacle to maintaining optimum iron status in patients with dialysis-associated anemia. As part of the North American clinical trials for iron sucrose injection, we examined the effect of intravenous (IV) iron sucrose in 23 hemodialysis patients with documented sensitivity to iron dextran, ongoing epoetin alfa therapy, and below-target-range hemoglobin (Hgb) levels (<11.0 g/dL). We assigned patients to treatment groups according to whether reactions they had experienced to iron dextran were judged to be mild (n = 16; group A) or severe (n = 7; group B). We prospectively examined adverse events and vital signs after administering 100 mg of IV iron sucrose in each of 10 consecutive dialysis treatment sessions and compared results with those recorded in each of three consecutive dialysis sessions without iron treatment. We administered iron sucrose by IV push over 5 minutes to group A patients and by IV push over 5 minutes or IV infusion over 15 to 30 minutes to group B patients. We did not administer a test dose. Results showed no serious adverse drug reactions after a total of 223 doses of iron sucrose (184 doses by IV push, 39 doses by IV infusion). Intradialytic blood pressure changes after IV iron sucrose injection did not differ from those recorded during dialysis sessions without treatment. An increase in values for Hgb, hematocrit, transferrin saturation, and ferritin, coupled with no significant change in epoetin dose and a decrease in total iron-binding capacity, confirmed the efficacy of iron sucrose injection in managing anemia. We conclude that iron sucrose injection is safe and effective in the management of anemia in patients sensitive to iron dextran and can be administered without a test dose by IV push or infusion. PMID- 10873878 TI - Logarithmic extrapolation of a 15-minute postdialysis BUN to predict equilibrated BUN and calculate double-pool Kt/V in the pediatric hemodialysis population. AB - Blood urea nitrogen (BUN) concentration rebounds logarithmically for 1 hour after a hemodialysis treatment. We have previously devised and evaluated an equilibrated Kt/V (eqKt/V) estimation method using logarithmic extrapolation of the BUN increase from 30 seconds to 15 minutes postdialysis in six pediatric hemodialysis patients. The current study evaluates logarithmic extrapolation in 15 additional pediatric patients. Mean measured equilibrated BUN (eqBUN) and estimated BUN at equilibrium (estBUN) using logarithmic extrapolation were 23.1 +/- 9.2 and 23.0 +/- 9.4 mg/dL, respectively. The mean absolute difference between estBUN and eqBUN was 0.7 +/- 0. 4 mg/dL (range, 0.1 to 1.55 mg/dL). All treatments had an absolute difference less than the SD of the laboratory measurement itself. The mean absolute percentage of difference between eqKt/V using eqBUN and estimated double-pool equilibrated Kt/V (estKt/V) using estBUN from logarithmic extrapolation was 3.4% +/- 2.3% and did not vary as a function of patient size, urea generation rate, dialyzer urea clearance, Kd/V, or ultrafiltration fraction. Mean absolute percentages of difference between eqKt/V and Kt/V estimated by the Tattersall, Daugirdas, or Maduell formulas were 4.5% +/ 3.9%, 4.4% +/- 3.7%, and 6.7% +/- 8.3%, respectively. Total percentages of error (absolute mean percentage of error + 2 SD) between eqKt/V and estKt/V by logarithmic extrapolation or the Tattersall, Daugirdas, or Maduell formulas were 8.0%, 12.3%, 11.8%, and 22.3%, respectively. The greater accuracy of logarithmic extrapolation compared with other methods of double-pool Kt/V estimation held true for patients weighing less than 35 kg. We have validated the use of an easy and accurate method requiring only an additional 15-minute posttreatment BUN level to estimate double-pool Kt/V in children. PMID- 10873879 TI - Randomized prospective study of the effect of increased dialytic dose on nutritional and clinical outcome in continuous ambulatory peritoneal dialysis patients. AB - Cohort studies have shown that greater urea (Kt/V) and creatinine clearances (CCr) were associated with better survival in patients on continuous ambulatory peritoneal dialysis (CAPD). The possibility of improved patient outcome with increased dialytic dose remains unknown. We prospectively studied over 1 year the effects of an extra 2-L bag on the outcome of 82 patients undergoing three daily 2-L exchanges for at least 12 months. At 1 year, 36 patients were undergoing 6-L exchanges, whereas 30 patients underwent 8-L exchanges. The increased dialytic dose resulted in increased total weekly Kt/V (TKt/V; 1.82 to 2.02), whereas total weekly CCr (TCCr) was maintained (63.2 to 61.9 L/1.73 m(2)). Control patients had reduced solute clearances (TKt/V, 1.87 to 1.67; TCCr, 64.8 to 54.6 L/1.73 m(2)). The fourth bag exchange resulted in a significant increase in net ultrafiltration (0.83 to 1.51 L/d), whereas the control group also had greater ultrafiltration (0.68 to 1.01 L/d) after 1 year. Although the normalized protein equivalent of nitrogen appearance (nPNA) was stable in the controls, the patients using 8-L exchanges achieved a greater nPNA (1.10 to 1.24 g/kg/d). There was no associated change in serum albumin levels (3.79 to 3.48 g/dL). The hospitalization rate increased in the controls (0.9 to 1.8 admissions/12 mon), whereas it was unchanged in the patients using 8-L exchanges. In conclusion, a 33% increase in dialytic prescription led to increased peritoneal and total clearances. Despite achieving increased nPNA (13%), the serum albumin level was unchanged. However, the increased hospitalization rate observed in the controls was avoided in the group using 8-L exchanges. PMID- 10873880 TI - Ultrafiltration profiling and measurement of relative blood volume as strategies to reduce hemodialysis-related side effects. AB - Hemodialysis (HD) side effects, such as hypotension and muscle cramps, may be related to excessive ultrafiltration (UF) in relation to refilling of fluids from the extravascular space, resulting in hemoconcentration and reduction of relative blood volume (RBV). This study examines the suitability of RBV measurements and UF modeling to reduce the incidence of dialysis side effects. We followed up 188 dialysis sessions in 53 patients. RBV and incidence of side effects were evaluated. Six treatment regimens were examined: UF profile 0, with a constant UF rate; UF profile 1, with a linear decreasing UF rate; UF profile 2, with a stepwise decreasing UF rate; and UF profiles 3 through 5, with intermittent high UF rates interrupted by UF pauses. During dialyses with a constant UF rate (UF profile 0), 10.6% of the treatments were associated with symptomatic hypotension. UF profiles 2 through 5, intermittently using high UF rates, caused a marked increase in hypotensive episodes (18.4%). In contrast, UF profile 1, providing a continuously decreasing UF rate, showed a reduced incidence of hypotension at only 5.7%. Symptomatic hypotension occurred in 13 of 53 patients during one or more dialysis sessions. With the help of RBV measurements, a subgroup of 8 patients with hypovolemia-induced hypotension could be identified. In these patients, an individual threshold of RBV could be defined, below which 92.3% of all hypotensive episodes occurred. In the remaining 5 hypotension-prone patients, there was no correlation between the occurrence of symptomatic hypotension and low RBV during HD treatments. In conclusion, UF profiles intermittently using high UF pulses cannot be recommended. RBV measurements help define a subgroup of patients at risk for hypovolemia-induced hypotension. Only these patients may benefit from blood volume-controlled UF. The incidence of symptomatic hypotension can likely be reduced if an individual threshold of RBV is avoided during HD treatments, eg, using lower UF rates in these patients. PMID- 10873881 TI - Diagnostic efficacy of (13)C-urea breath test for Helicobacter pylori infection in hemodialysis patients. AB - The noninvasive urea breath test (UBT) avoids the discomforts and risks of invasive endoscopic methods of Helicobacter pylori detection. This study investigated the diagnostic efficacy of carbon 13 ((13)C)-labeled UBT for H pylori detection in 70 patients with end-stage renal disease (ESRD) undergoing hemodialysis (HD) and 70 dyspeptic controls without renal impairment. With H pylori infection defined as a positive result on either histological examination or culture of gastric biopsy specimen, we evaluated the reliability of the (13)C UBT in detecting H pylori infection in both groups. To ascertain whether HD therapy affects the diagnostic efficacy of the UBT, the test was performed twice in patients with ESRD (before and after HD) at least 72 hours apart. In each UBT session, the baseline, 10-minute, and 15-minute (Delta15) gas samples were obtained to analyze excess (13)CO(2)/(12)CO(2) ratio (ECR). Histological stain and/or culture studies found that 33 of the patients with ESRD (47. 1%) and 42 of the control patients (60%) had H pylori infection. (13)C-UBT for H pylori detection in patients with ESRD was found to be only 93.8% sensitive and 85.3% specific. These results were achieved by gas sampling (Delta15) after HD therapy with a cutoff ECR value greater than 5. Conversely, the UBT in the control group achieved the greatest diagnostic efficacy (sensitivity, 97.6%; specificity, 96.4%) with a comparatively lower ECR cutoff value of 4. We conclude that the diagnostic accuracy for H pylori detection in patients with ESRD could be improved by performing (13)C-UBT (Delta15) after HD therapy and assessing the UBT with a cutoff ECR value greater than 5. However, the diagnostic efficacy of the UBT for patients with ESRD remained less accurate than that for dyspeptic patients without renal impairment. PMID- 10873882 TI - Cardiopulmonary events during hemodialysis: effects of dialysis membranes and dialysate buffers. AB - Adverse cardiac and pulmonary events are frequently observed during hemodialysis and contribute to significant morbidity and mortality. The temporal relationship between these events during the intradialytic period has not been well defined. To examine the event rate and timing of silent ischemia, cardiac ectopy, and hypoxemia, we conducted a prospective, single-blind, randomized study of 10 subjects undergoing maintenance hemodialysis with four contiguous combinations of dialysis membranes (cuprammonium or polysulfone) and dialysates (acetate or bicarbonate). The frequency of oxygen desaturation events peaked during the first 2 hours, whereas silent myocardial ischemia and supraventricular ectopies occurred more often in the later hours. Ventricular ectopy occurred steadily throughout the intradialytic period. The combination of acetate dialysis and cuprammonium membrane is associated with the most frequent events. We conclude that cardiopulmonary events can occur frequently during hemodialysis, and the frequency is dependent on the type of dialysis membrane and dialysate buffer used. PMID- 10873883 TI - Dialysis discontinuation and palliative care. AB - Little attention has been accorded to the terminal course and end-of-life care of patients after dialysis discontinuation. This prospective cohort observational study involves six dialysis clinics in the United States and two clinics in Canada. Data were collected on 131 patients who were undergoing maintenance dialysis and died after treatment discontinuation. Seventy-nine of the patients (60%) were prospectively studied until their deaths. Caregivers and families provided information about the symptoms and treatment provided in the final 24 hours of life, and structured interviews were conducted at the time of stopping dialysis with patients and families. The patient population was primarily white (73%), elderly (70 +/- 1.2 years), and diabetic (46%). Three quarters of the subjects had between three and seven comorbid conditions. Pain and agitation were the most common symptoms during the last day of life. Terminal treatment was generally considered to be satisfactory, and most people had good deaths. Although dialysis prolongs life, the integration of palliative medicine into dialysis programs offers opportunities to improve the quality of end-of-life care, especially for those patients who elect to stop treatment. Recommendations include making advance care planning an expectation at all clinics and using quality-of-dying measures to establish benchmarks for the provision of terminal care. PMID- 10873884 TI - Long-term survival of renal transplant recipients in the United States after acute myocardial infarction. AB - Cardiac disease is a major cause of death in renal transplant recipients. One third of the cardiac deaths are attributed to acute myocardial infarction (AMI). Few data exist on predictors of long-term survival of renal transplant recipients after AMI. The purpose of this study is to determine predictors of survival (including treatment era) for renal transplant recipients in the United States after AMI. The US Renal Data System database of 783, 171 patients was used to retrospectively examine outcomes of renal transplant recipients hospitalized during 1977 to 1996 for a first AMI after initiation of renal replacement therapy. Long-term survival was estimated by life-table method, and independent predictors of survival were examined in a comorbidity-adjusted Cox model. There were 4,250 renal transplant recipients with AMI. The in-hospital death rate was 12.8%. Overall 2-year cardiac and all-cause mortality rates were 11.8% +/- 0.6% (SE) and 33.6% +/- 0. 8%, respectively. The poorest survival after AMI occurred in patients with diabetic end-stage renal disease (ESRD), with 2-year cardiac and all-cause mortality rates of 14.9% +/- 1.1% and 40.5% +/- 1.4%, respectively. In the Cox model, the risks for cardiac and all-cause death from AMI were 51% (P = 0.0003) and 45% less (P < 0. 0001) in 1990 to 1996 compared with 1977 to 1984, respectively. The long-term survival of renal transplant recipients in the United States after AMI has markedly improved in the modern treatment era. Patients with diabetic ESRD experience the worst outcome. PMID- 10873885 TI - Impact of pre-existing donor hypertension and diabetes mellitus on cadaveric renal transplant outcomes. AB - Hypertension (HTN) and diabetes mellitus (DM) predispose to systemic atherosclerosis with renal involvement. The prevalence of HTN and DM in cadaveric renal donors (affected donors) and the results of transplantation are unknown. We investigated these issues with national data from the US Renal Data System. A total of 4,035 transplants from affected donors were matched 1:1 with unaffected controls according to donor age and race, recipient race, and year of transplantation. Graft and patient survival were estimated. Among the 25,039 solitary renal transplantations performed between July 1, 1994, and June 30, 1997, cadaveric renal transplants from donors with HTN accounted for 15%, and donors with DM, 2%. Programs with 1-year cadaveric renal graft survival rates greater than 90% had 50% less affected donors compared with programs having 1 year cadaveric renal graft survival rates of 85% or less. Compared with donor-age matched controls, transplants from affected donors were at minimally increased risk for primary nonfunction, delayed graft function, and acute rejection. Three year graft survival rates were 71% in affected donor organs and 75% in controls (P = 0.001). Compared with controls, duration of HTN was an independent risk factor for graft survival (3-year graft survival rates, 75% versus 65%; relative risk = 1.36 for HTN >10 years; P < 0.001). A substantial fraction of cadaveric renal donors have preexisting HTN. Programs transplanting fewer affected donor kidneys had better than average results. Because the negative impact of donor HTN and DM on transplant outcome was of moderate degree except when the duration of donor HTN was greater than 10 years, use of affected donors should not be discouraged, but graft and patient survival analyses should account for their presence. PMID- 10873886 TI - Bone disease in patients with long-term renal transplantation and normal renal function. AB - Renal osteodystrophy may persist during the early years after renal transplantation. However, information on bone status after a successful long-term renal transplantation is limited. We examined biochemical parameters, bone mineral density (BMD), and bone histomorphometry in 25 asymptomatic men with normal renal function after 7.5 +/- 5.7 years of a renal transplantation. Serum calcium, phosphorus, alkaline phosphatase, and 1,25(OH)(2)D(3) levels and urinary calcium level and cyclic andenosine monophosphate excretion were within normal range in all patients. Serum intact parathyroid hormone (PTH) level was elevated in 11 subjects (133.6 +/- 78 pg/mL) and normal in the other 14 subjects (47.9 +/- 13.6 pg/mL). Mean BMD at the lumbar spine and femoral neck was low in the entire group. However, it progressively increased as time after transplantation increased, approaching normal values after 10 years. Bone histomorphometric analysis showed bone resorption, osteoid volume, and osteoid surface greater than normal range in the majority of patients. Bone formation rate and mineralization surface were low, and mineralization time was delayed in most patients. These lesions were more severe in patients after 3 to 4 years of transplantation but improved with time and approached normal values after a period of 10 years. PTH values did not correlate with bone histological characteristics or BMD. These results show that the bone alterations observed after long-term renal transplantation consist of a mixed bone disease in which features of high bone turnover coexist with altered bone formation and delayed mineralization. These findings may result from the combined effect of preexisting bone disease and immunosuppressive therapy. PMID- 10873887 TI - Clinical risk factors associated with nonmelanoma skin cancer in renal transplant recipients. AB - A single-center, cross-sectional, longitudinal study was conducted to determine the prevalence, annual incidence, and clinical risk factors for skin cancer in a white renal transplant population. One hundred eighty-two white patients (95% of population) with functioning allografts, a mean age at transplantation of 38.9 +/ 15. 6 (SD) years, and a mean follow-up of 8.5 +/- 6.3 years were interviewed and examined between May 1997 and June 1999. All case notes were carefully reviewed. Since transplantation, 16.5% of the patients had developed nonmelanoma skin cancer; 15.4%, actinic keratoses (AK); 53%, viral warts; and 1.6%, lentigo maligna melanoma (n = 3). Thirty-nine percent of the tumors were detected as a consequence of this study, and 20% of these occurred on covered body sites. The squamous cell (SCC)-basal cell carcinoma (BCC) ratio was 3.8:1. Eighty-two percent of the patients were examined a second time 12 months after the initial assessment. Using these data to identify new lesions, the annual incidence was calculated at 6.5%, increasing to 10.5% at more than 10 years posttransplantation. Duration of immunosuppression, older age at transplantation, presence of AK, male sex, and outdoor occupation were significantly associated with both SCC and BCC; SCC alone was associated with a history of having smoked tobacco. Early identification of those at greatest risk using a clinical risk profile may allow the development of more structured preventative and surveillance strategies than currently exist. PMID- 10873888 TI - Plasma exchange for the removal of digoxin-specific antibody fragments in renal failure: timing is important for maximizing clearance. AB - Life-threatening digoxin toxicity may be effectively treated with digoxin specific antibody fragments (Fab). However, in end-stage renal disease, the digoxin-Fab complexes persist in the circulation and dissociate, potentially resulting in rebounding free digoxin levels and the recurrence of symptomatic toxicity. To prevent this rebound phenomenon, plasma exchange (PE) has been implemented for the removal of the digoxin-Fab complexes in renal failure. However, there is only one case report describing its use in this setting. To better determine the optimal timing of PE after Fab administration, we performed two PE treatments (each preceded by Fab) in a patient with acute renal failure and acute digoxin poisoning. The admission serum digoxin level was 21 ng/mL. The timing of the PE treatments relative to Fab dosing was as follows: the first PE was performed 26 hours post-Fab, and the second PE was performed 2.5 hours post Fab. The plasma ultrafiltrate digoxin concentration was 2.5-fold greater when PE was performed 2.5 hours versus 26 hours after Fab administration (19.9 versus 8.1 ng/mL). The combined total amount of digoxin removed in the ultrafiltrate plasma was minimal (0.13 mg), less than 1% of the total amount of ingested drug. We conclude that the optimal timing of PE is within the first 3 hours after Fab administration. Although PE is efficacious for removing digoxin-Fab complexes, thus preventing rebound digoxin toxicity, it is not efficacious for improving total digoxin clearance because of the large apparent volume of distribution of digoxin (5 to 8 L/kg). PMID- 10873889 TI - Bilateral renal infarction in a black man with medial fibromuscular dysplasia. AB - We report a case of bilateral renal infarction in a patient with medial fibrous dysplasia of both renal arteries and a thrombosed aneurysmal dilatation of the right renal artery. A previously healthy 40-year-old black man presented to the emergency department with acute onset of bilateral flank pain. Computerized tomography of the abdomen showed bilateral renal infarction, predominantly affecting the anterior distribution of both renal arteries. Estimated loss of renal mass was 50% on the right and 25% on the left. The patient was treated with intravenous heparin, oral warfarin, and antihypertensive therapy with labetolol and long-acting nifedipine. By day 3, his abdominal pain resolved; however, the serum creatinine level increased to a maximum value of 2.6 mg/dL. The serum creatinine level slowly improved and stabilized at 1.9 mg/dL, and he was subsequently discharged on the seventh hospital day. Magnetic resonance angiography performed 2 months later showed "beading2 of both renal arteries consistent with medial fibromuscular dysplasia, a finding confirmed by conventional angiography. To our knowledge, bilateral renal infarction complicating medial fibrous dysplasia of the renal arteries has not been previously reported, nor has medial fibrous dysplasia been reported in blacks. PMID- 10873890 TI - Congenital focal segmental glomerulosclerosis associated with beta4 integrin mutation and epidermolysis bullosa. AB - We report the occurrence of congenital nephrotic-range proteinuria secondary to focal segmental glomerulosclerosis in an infant with epidermolysis bullosa and pyloric atresia. A homozygous missense mutation, R1281W, in exon 31 of the beta4 integrin gene, ITGB4, was identified. By immunofluorescence, beta4 integrin expression was reduced in both dermal keratinocytes and glomerular podocytes. This is the first demonstration of beta4 integrin expression in human glomeruli. We postulate a role for altered beta4 integrin function in the mediation of the glomerular permeability defect. PMID- 10873891 TI - Prevention of intraocular pressure elevations during hemodialysis. AB - Intraocular pressure (IOP) may rise during hemodialysis sessions in predisposed patients because of a rapid drop in osmolality at the blood compartment. A patient with diabetes had painful ocular episodes during hemodialysis that were associated with an IOP increase. We modified the dialysis parameters to prevent a rapid decrease in osmolality by creating conductivity and ultrafiltration profiles and adding a colloid solution at the beginning of the procedure. After instituting these changes, the patient became asymptomatic and did not have variations in IOP during the dialysis sessions. PMID- 10873892 TI - An outbreak of Burkholderia cepacia bacteremia in hemodialysis patients: an epidemiologic and molecular study. AB - The risk of blood stream infections increases in patients undergoing chronic hemodialysis. Outbreaks of infection are usually caused by contamination of the water supply, water treatment, distribution system, or dialyzer reprocessing. We report an outbreak of subclavian catheter-related Burkholderia cepacia bacteremia in nine patients undergoing hemodialysis. Using randomly amplified polymorphic DNA (RAPD) analysis, the bacterial isolates were clonally identical to Burkholderia cepacia isolated from residue of the diluted chlorhexidine-cetrimide solution used to disinfect the transfer forceps. These forceps were used to pick up cotton balls and gauze for dressing the subclavian catheter. Antibiotic therapy failed to cure the infections, and all patients required catheter removal. Pathology showed numerous bacilli embedded in the biofilm on the inner surface of the removed catheters. In conclusion, our study showed that contaminated chlorhexidine-cetrimide solution was the source of a bacteremic outbreak in nine patients who developed catheter-related Burkholderia cepacia infection. PMID- 10873893 TI - Cystatin C measurement: improved detection of mild decrements in glomerular filtration rate versus creatinine-based estimates? PMID- 10873894 TI - Selection bias impacts outcome reports of uremia therapy. PMID- 10873895 TI - Survival of renal transplant patients after first myocardial infarction: a look to the past and promises for the future. PMID- 10873896 TI - New insights into the pathogenesis of proteinuria. PMID- 10873898 TI - Henoch-Schonlein purpura and myocardial necrosis. PMID- 10873897 TI - Nephrotic syndrome and arthritis in a 12-year-old girl. PMID- 10873899 TI - No need for immediate dialysis after administration of low-osmolarity contrast medium in patients undergoing hemodialysis. PMID- 10873900 TI - Cyclosporine toxicity PMID- 10873901 TI - Continuing medical education exercise, july 2000 PMID- 10873902 TI - Glomerular and extraglomerular immune complex deposits in a bone marrow transplant recipient. AB - A 44-year-old man developed nephrotic syndrome 9 months after HLA-identical sibling bone marrow transplantation. Membranous changes consisted mainly of alterations of glomeruli, which were interpreted as chronic graft-versus-host disease (GVHD) caused by lodging of the circulating immune complex. In the tubules, a lumpy deposition of IgG and complement breakdown products was distributed along the tubular basement membrane, which coincided with the peculiar deposits ascertained by electron microscopy. These findings suggest that an extraglomerular reaction should be considered in evaluating renal involvement of GVHD. PMID- 10873903 TI - Renal sarcoidosis with superimposed postinfectious glomerulonephritis presenting as acute renal failure. AB - We describe two patients with sarcoidosis with lesions of granulomatous interstitial nephritis (GIN) and postinfectious glomerulonephritis (GN). Both patients presented with heavy proteinuria, hematuria, and renal failure. Renal histology in both showed GIN and glomerular changes of proliferative GN with hump like subepithelial deposits by electron microscopy of postinfectious GN. Antecedent history of pneumonia was present in one, and ASO titer was elevated in the other. The proteinuria and azotemia improved in both with steroid therapy. Reports of "postinfectious" or diffuse proliferative GN in patients with sarcoidosis are rare. The authors are unaware of reports of concomitant sarcoid GIN and postinfectious GN. Although acute renal insufficiency or failure can occur with GIN or other more common renal lesions primary glomerular disease should be considered in patients with sarcoidosis who present with renal dysfunction. This is a US government work. There are no restrictions on its use. PMID- 10873904 TI - Renal thrombotic microangiopathy associated with interferon-alpha treatment of chronic myeloid leukemia. AB - Recent reports have documented the development of renal thrombotic microangiopathy in patients with chronic myeloid leukemia (CML) who have undergone treatment with interferon-alpha. The pathogenesis of the renal lesion in such cases remains unclear. We report the case of a patient with chronic myeloid leukemia who developed renal failure and nephrotic syndrome while being treated with hydroxyurea and interferon-alpha. The renal biopsy showed features of chronic thrombotic microangiopathy. The patient had serologic and functional evidence of anti-phospholipid antibody. Interferon-alpha is known to cause induction of multiple autoantibodies. We propose that in the context of CML, interferon-alpha treatment can induce pathogenic anti-phospholipid antibodies that result in renal thrombotic microangiopathy. This has important implications for patients with CML receiving immune-stimulating therapy because it suggests that prospective monitoring of such patients for anti-phospholipid antibody might identify those at risk of developing thrombotic microangiopathy. Furthermore, patients with established anti-phospholipid antibody syndrome in this context might benefit from intervention such as early anticoagulation. PMID- 10873905 TI - Marked hyperlactatemia associated with severe alkalemia in a patient with thrombotic thrombocytopenic purpura. AB - This report describes a case of severe alkalemia associated with a blood lactate level greater than 13 mEq/L in a patient without evidence of hypotension or hypoxemia. The patient, who had the clinical manifestations of thrombotic thrombocytopenic purpura (TTP), developed the alkalemia from an acute respiratory alkalosis superimposed on an existing metabolic alkalosis. Profound alkalemia may impair oxygen delivery because of stronger hemoglobin-oxygen affinity, vasoconstriction, and alterations in the redox potential of cytochrome c. We suggest that the synergistic effects of a sudden, extreme alkalemia and the localized tissue hypoxia that resulted from extensive microvascular thrombi secondary to TTP caused the patient's hyperlactatemia. PMID- 10873906 TI - Intrarenal endothelin-1 and hypertension. PMID- 10873907 TI - Genetics of psychiatric disorders: where have we been and where are we going? PMID- 10873908 TI - Toward reformulating the diagnosis of schizophrenia. AB - OBJECTIVE: The authors assess implications of DSM criteria for schizophrenia by reviewing the criteria's 1) emphasis on psychotic features, 2) dissociation of symptoms from their etiology, 3) exclusive reliance on clinical features but exclusion of biological indicators, and 4) classification of schizophrenia as a discrete category. The authors then discuss alternative conceptions of schizophrenia that take into account recent data concerning its genetic and neurodevelopmental origins and its pathophysiological substrates. METHOD: The historical development of diagnostic criteria for schizophrenia is reviewed in the context of recent published data on the biology and development of schizophrenia. RESULTS: Growing evidence suggests that symptoms of psychosis may be a common end-state in a variety of disorders, including schizophrenia, rather than a reflection of the specific etiology of schizophrenia. Features occurring before the advent of psychosis that are clinical, biological, and/or neuropsychological in nature may constitute evidence of a genetic predisposition toward schizophrenia ("schizotaxia") and may provide more specific information about the genetic, pathophysiological, and developmental origins of schizophrenia. CONCLUSIONS: The success of efforts to treat and prevent schizophrenia will depend to an important extent on an accurate understanding of its causes. This goal can be furthered by conducting field trials to develop research criteria to assess the value of a developmentally sensitive, biologically informed approach to classification that would consider schizotaxia with psychosis (schizophrenia) and schizotaxia alone as distinct diagnostic conditions. PMID- 10873909 TI - Cognitive behavioral therapy for the treatment of binge eating disorder: what constitutes success? PMID- 10873911 TI - Mitochondrial DNA sequence diversity in bipolar affective disorder. AB - OBJECTIVE: Point mutations in mitochondrial DNA (mtDNA) are one mechanism that could explain the apparent excess maternal transmission of bipolar affective disorder observed in some families. The authors sequenced the mtDNA from probands with bipolar disorder and tested nucleotide variants for association with the disorder. METHOD: The entire 16.5 kilobase mitochondrial genome was sequenced in nine unrelated probands selected from large pedigrees with exclusively maternal transmission of bipolar affective disorder. Compared to a reference sequence, variants were detected at 107 nucleotide positions. Fifteen variants of possible pathogenic significance were selected for further study. These variants were assayed in 93 unrelated probands with bipolar I, bipolar II, or schizoaffective manic disorder and 63 comparison subjects, all of whom were classified into the major groups comprising the European mtDNA haplotype structure (haplogroups). RESULTS: The major European haplogroups were represented at the expected frequencies among both probands and comparison subjects. There was no significant difference between probands and comparison subjects in the frequency of any variant, although odds ratios >2 or <0.5 were observed for four variants. Frequencies of these four variants were similar in probands and haplogroup matched comparison subjects. The results of all comparisons were essentially unchanged when probands from families with an apparently paternal transmission pattern were excluded. CONCLUSIONS: The results demonstrate that bipolar affective disorder occurs across all of the major European mtDNA haplogroups but do not reveal any point mutations that explain excess maternal transmission of the disorder. PMID- 10873913 TI - Sibling correlation of deficit syndrome in the Irish study of high-density schizophrenia families. AB - OBJECTIVE: The deficit syndrome is a subtype of schizophrenia characterized by primary and enduring negative features of psychopathology. It appears to reflect a distinct subtype within the syndrome of schizophrenia. Little is known about the familial or genetic aspects of the deficit syndrome. The purpose of this study was to determine whether deficit versus nondeficit subtypes are correlated in sibling pairs affected with schizophrenia. METHOD: The present study was based on the Irish Study of High-Density Schizophrenia Families. From the earlier study the authors selected a subset of patients who were members of sibling pairs in which both siblings had been diagnosed with "core" schizophrenia, which included schizophrenia, simple schizophrenia, and schizoaffective disorder with poor outcome. The Schedule for the Deficit Syndrome was used to make deficit versus nondeficit diagnoses, which were based on chart examinations by reviewers blind to sibling status. This method resulted in 65 patients being diagnosed with the deficit syndrome and 401 patients diagnosed as nondeficit (prevalence=13.9%). This group included 347 full sibling pairs, which were analyzed for resemblance with respect to deficit versus nondeficit subtype by means of logistic regression. RESULTS: Deficit versus nondeficit subtypes were significantly correlated in sibling pairs concordant for core schizophrenia. CONCLUSIONS: Familial factors contribute significantly to whether a person has the deficit subtype of schizophrenia. This familial contribution could be genetic or environmental. PMID- 10873912 TI - Low prevalence of psychoses among the Hutterites, an isolated religious community. AB - OBJECTIVE: The authors estimated the prevalence of psychoses among the Hutterites in Manitoba, Canada, who lived in 102 communal farms or colonies. The study stemmed from an earlier epidemiological survey of North American Hutterite colonies (1950-1953), in which a low prevalence of psychoses was documented. METHOD: Psychiatrically ill individuals identified during the previous survey were rediagnosed with DSM-IV criteria. A current provincial health insurance claims database was queried anonymously for the period June 1992-May 1997, and the prevalence rate of disease among Hutterites, identified by distinctive surnames and unique postal addresses, was compared with the rate in the entire population of the province of Manitoba and in a comparison group of persons with Hutterite surnames but with addresses outside the Hutterite colonies. RESULTS: The annual prevalence of schizophrenia among the communal Hutterites, estimated from the database search by using ICD-9 criteria, was consistent with the prevalence found in the prior epidemiological survey (annual mean of 1.2/1,000 population, compared with 1.3/1,000 in the prior survey). The database search yielded a significantly lower prevalence for schizophrenia and other functional psychoses among communal Hutterites as well as among the comparison group, compared to the total Manitoba population. There was also lower prevalence for affective psychoses and adjustment reaction disorders among the communal Hutterites, compared to the total Manitoba population. Rates for neurotic disorders were elevated both among the communal Hutterites and the comparison group. CONCLUSIONS: The prevalence of specific psychoses was reduced among the Hutterites, although neurotic disorders were more prevalent. These findings suggest some specificity, although possible artifacts such as ascertainment bias must be considered. Further research is needed to examine genetic and environmental factors that may contribute to reduced prevalence of specific psychoses among the Hutterites. PMID- 10873914 TI - Family study of girls with attention deficit hyperactivity disorder. AB - OBJECTIVE: Because attention deficit hyperactivity disorder (ADHD) is relatively infrequent among girls, little is known about the causes of ADHD in girls. To help fill this gap in the literature, the authors assessed the familial transmission of ADHD in families ascertained through girls. METHOD: Interviewers who were blind to diagnosis administered structured psychiatric interviews to 140 girls with ADHD and their 417 first-degree relatives and to 122 girls without ADHD and their 369 first-degree relatives. RESULTS: The relatives of the ADHD girls had a significantly higher prevalence of ADHD, according to either the DSM III-R or DSM-IV definition, than the relatives of the comparison girls. However, this did not differ from the prevalence the authors reported previously for families of boys with ADHD. Like the boys' families, the relatives of the girl probands also had significantly higher prevalences of antisocial, mood, anxiety, and substance use disorders, although the prevalence of familial antisocial disorders was lower than had been observed in the boys' families. There was no association between the DSM-IV subtypes of the probands and relatives. CONCLUSIONS: The familial transmission of ADHD and comorbid disorders generalizes to families of girls with ADHD. Neither proband gender nor subtype influences the familial transmission of ADHD. PMID- 10873915 TI - Protective factors against suicidal acts in major depression: reasons for living. AB - OBJECTIVE: Over 30,000 people a year commit suicide in the United States. Prior attempted suicide and hopelessness are the most powerful clinical predictors of future completed suicide. The authors hypothesized that "reasons for living" might protect or restrain patients with major depression from making a suicide attempt. METHOD: Inpatients with DSM-III-R major depression were assessed for depression, general psychopathology, suicide history, reasons for living, and hopelessness. Of the 84 patients, 45 had attempted suicide and 39 had not. RESULTS: The depressed patients who had not attempted suicide expressed more feelings of responsibility toward family, more fear of social disapproval, more moral objections to suicide, greater survival and coping skills, and a greater fear of suicide than the depressed patients who had attempted suicide. Scores for hopelessness, subjective depression, and suicidal ideation were significantly higher for the suicide attempters. Reasons for living correlated inversely with the combined score on these measures, considered an indicator of "clinical suicidality." Neither objective severity of depression nor quantity of recent life events differed between the two groups. CONCLUSIONS: During a depressive episode, the subjective perception of stressful life events may be more germane to suicidal expression than the objective quantity of such events. A more optimistic perceptual set, despite equivalent objective severity of depression, may modify hopelessness and may protect against suicidal behavior during periods of risk, such as major depression. Assessment of reasons for living should be included in the evaluation of suicidal patients. PMID- 10873916 TI - Marked differences in antidepressant use by race in an elderly community sample: 1986-1996. AB - OBJECTIVE: Prescriptions of antidepressant medications have increased significantly over the past 15 years across the life cycle. One overall correlate of medication use in older adults is race, with African Americans using fewer medications than whites. Given the frequency of depressive symptoms among elderly populations, as well as the increased potential for adverse side effects from antidepressants, the relative contribution of race in the use of antidepressants is critical for determining well-designed studies. The authors analyzed data from a community-based cohort of elders followed for 10 years to determine the association of race to the use of antidepressants between 1986 and 1996, with control for known correlates of depression in late life. METHOD: Information on antidepressant use and demographic and health characteristics were obtained from a stratified, probability-based sample of 4,162 elders (equally distributed between African American and white community-dwelling subjects) in the Piedmont region of North Carolina during four in-person interviews spanning 10 years. Descriptive statistics were calculated. Logistic regression was used for the final models. RESULTS: A total of 4.6% of whites and 2.3% of African Americans used antidepressants in 1986. Approximately 14.3% of whites and 5.0% of African Americans used antidepressants in 1996. In controlled analyses, the prevalence odds ratio for antidepressant use in whites, compared to African Americans, was 1. 76 in 1986 and 3.77 in 1996. CONCLUSIONS: African American elders are much less likely to take antidepressants, and the difference in use increased over the 10 years of the survey. PMID- 10873917 TI - Neuropsychological deficits in psychotic versus nonpsychotic major depression and no mental illness. AB - OBJECTIVE: At least three studies have indicated that patients with psychotic major depression studied under non-drug-free conditions differ from patients with nonpsychotic major depression and healthy comparison subjects on several measures of neuropsychological performance. The current study explored specific impairments in cognitive function in subjects with psychotic major depression, subjects with nonpsychotic major depression, and healthy comparison subjects studied under drug-free conditions. METHOD: A battery of neuropsychological tests was administered to 11 patients with psychotic major depression, 32 patients with nonpsychotic major depression, and 23 normal comparison subjects under drug-free conditions. The three groups did not differ statistically in age, sex, or level of education. To ensure that participants had minimal levels of severity and endogenicity, all patients were required to have a score of at least 20 on the 21 item Hamilton Depression Rating Scale and a score of at least 7 on the Core Endogenomorphic Scale, which uses eight items from the Hamilton depression scale. RESULTS: Patients with psychotic major depression demonstrated significantly greater impairment than patients with nonpsychotic major depression and/or comparison subjects in attention and response inhibition (as measured by the Stroop color-word subscale score) as well as in verbal declarative memory (as measured by the Paragraph Recall Test). CONCLUSIONS: These data indicate that patients with psychotic major depression demonstrate impairment in functions thought to be mediated by the frontal cortex and mediotemporal lobes. PMID- 10873918 TI - Influence of panic-agoraphobic spectrum symptoms on treatment response in patients with recurrent major depression. AB - OBJECTIVE: The authors tested the hypothesis that a lifetime history of panic agoraphobic spectrum symptoms predicts a poorer response to depression treatment. METHOD: A threshold for clinically meaningful panic-agoraphobic spectrum symptoms was defined by means of receiver operating characteristic curve analysis of total scores on the Structured Clinical Interview for Panic-Agoraphobic Spectrum in a group of 88 outpatients with and without panic disorder. This threshold was then applied to a group of 61 women with recurrent major depression, who completed a self-report version of the same instrument, in order to compare treatment outcomes for patients above and below this clinical threshold. RESULTS: Women with high scores (> or =35) on the Panic-Agoraphobic Spectrum Self-Report were less likely than women with low scores (<35) to respond to interpersonal psychotherapy alone (43.5% versus 68.4%, respectively). Women with high scores also took longer (18.1 versus 10.3 weeks) to respond to a sequential treatment paradigm (adding a selective serotonin reuptake inhibitor when depression did not remit with interpersonal psychotherapy alone). This effect was only partially accounted for by the higher likelihood that patients with high scores required the addition of antidepressants. Although four domains from the Panic-Agoraphobic Spectrum Self-Report were individually associated with a longer time to remission, only stress sensitivity emerged as significant in multivariate regression analyses. CONCLUSIONS: A lifetime burden of panic-agoraphobic spectrum symptoms predicted a poorer response to interpersonal psychotherapy and an 8-week delay in sequential treatment response among women with recurrent depression. These results lend clinical validity to the spectrum construct and highlight the need for alternate psychotherapeutic and pharmacologic strategies to treat depressed patients with panic spectrum features. PMID- 10873919 TI - Brain SPECT imaging of amphetamine-induced dopamine release in euthymic bipolar disorder patients. AB - OBJECTIVE: Increased dopaminergic neurotransmission has been implicated in the pathophysiology of bipolar disorder. However, it remains unclear whether the abnormality is due to increased dopamine release or enhanced postsynaptic receptor sensitivity. In this study, dopamine receptor imaging combined with a pharmacological challenge of amphetamine was used to assess both pre- and postsynaptic aspects of dopamine neurotransmission in euthymic bipolar disorder patients. METHOD: Thirteen patients with bipolar disorder (seven medication free and six receiving mood stabilizer therapy) who had been euthymic for more than 4 weeks and 13 age- and gender-matched healthy comparison subjects were included in the study. Single photon emission computed tomography scans were obtained with the striatal dopamine (D(2)/D(3)) receptor radiotracer iodobenzamide ([(123)I]IBZM) before and after an intravenous amphetamine challenge (0.3 mg/kg). Reduction in striatal [(123)I]IBZM binding potential from the first scan to the second scan was used as an indirect measure of the amount of dopamine released. Behavioral response to amphetamine was measured with the Brief Psychiatric Rating Scale, Young Mania Rating Scale, and visual analogue scales. RESULTS: Bipolar patients and healthy subjects did not differ in terms of mood state or striatal D(2) receptor binding at baseline. Amphetamine challenge led to a significantly greater behavioral response in bipolar patients than in healthy subjects. However, there was no significant difference between the two groups in the amphetamine-induced decrease in striatal [(123)I]IBZM binding. CONCLUSIONS: In a group of euthymic patients with bipolar disorder, this study did not find evidence for increased striatal dopamine release. Instead, these data are consistent with enhanced postsynaptic dopamine responsivity in patients with bipolar disorder. PMID- 10873920 TI - Predeployment personality traits and exposure to trauma as predictors of posttraumatic stress symptoms: a prospective study of former peacekeepers. AB - OBJECTIVE: The authors' goal was to study the contribution of predeployment personality traits and exposure to traumatic events during deployment to the development of symptoms of posttraumatic stress disorder (PTSD) in individuals involved in military peacekeeping activities. METHOD: Five hundred seventy-two male veterans who participated in the United Nations Protection Force mission in the former Yugoslavia completed a short form of the Dutch MMPI before deployment. Following deployment, they participated in a survey of all Dutch military veterans who had been deployed in the years 1990-1995 and completed the Self Rating Inventory for PTSD. RESULTS: Exposure to traumatic events during deployment had the highest unique contribution to the prediction of PTSD symptom severity, followed by the personality traits of negativism and psychopathology, followed by age. CONCLUSIONS: Both pretrauma vulnerabilities and exposure to traumatic events were found to be important factors in the etiology of posttraumatic stress symptoms. The current study replicates in a non-American sample of peacekeepers findings obtained among American Vietnam veterans. Particularly, there is accumulating evidence for an etiological role of the personality trait of psychoneuroticism in the development of posttraumatic stress symptoms. PMID- 10873921 TI - Decreased benzodiazepine receptor binding in prefrontal cortex in combat-related posttraumatic stress disorder. AB - OBJECTIVE: Animals exposed to stress exhibit a decrease in benzodiazepine receptor binding in the frontal cortex. No studies have examined central benzodiazepine receptor binding in patients with posttraumatic stress disorder (PTSD). The purpose of this study was to examine measures of benzodiazepine receptor binding in PTSD. METHOD: From 13 patients with Vietnam combat-related PTSD and 13 case-matched healthy comparison subjects, a quantitative measure related to benzodiazepine receptor binding (distribution volume) was obtained with single photon emission computed tomography (SPECT) imaging of [(123)I]iomazenil binding and measurement of radioligand concentration in plasma. Distribution volume image data were analyzed by means of statistical parametric mapping. RESULTS: Lower distribution volumes were found in the prefrontal cortex (Brodmann's area 9) of PTSD patients than in comparison subjects. CONCLUSIONS: These findings of lower values for the benzodiazepine receptor binding measure of distribution volume are consistent with fewer benzodiazepine receptors and/or reduced affinity of receptor binding in the medial prefrontal cortex in patients with PTSD. Alterations in benzodiazepine receptor function in this area may underlie many of the symptoms of PTSD. PMID- 10873922 TI - Pathological gambling among cocaine-dependent outpatients. AB - OBJECTIVE: The authors investigated the occurrence of pathological gambling among cocaine-dependent outpatients, its influence on short-term outcome of treatment, and comparative characteristics of patients with and without pathological gambling. METHOD: The subjects were 313 cocaine-dependent (200 also opiate dependent) outpatients in clinical trials of medication for cocaine dependence. Pathological gambling (DSM-III-R criteria) was assessed with the Diagnostic Interview Schedule, and sociodemographic and socioeconomic characteristics were determined with the Addiction Severity Index. Outcome was defined as time in treatment (proportion of maximum scheduled time) and proportion of cocaine positive urine samples during treatment. RESULTS: Pathological gambling had a lifetime occurrence rate of 8.0% and a current (past month) occurrence of 3.8%. Onset preceded the onset of cocaine dependence in 72.0% of the patients (and preceded onset of opiate dependence in 44.4%). Patients with pathological gambling (lifetime or current) did not differ significantly from other patients in length of treatment or proportion of cocaine-positive urine samples. Those with lifetime pathological gambling were significantly more likely to have tobacco dependence (84.0% versus 61.1%) and antisocial personality disorder (56.0% versus 19.8%), to be unemployed (84.0% versus 49.3%), to have recently engaged in illegal activity for profit (64.0% versus 38.5%), and to have been incarcerated (62.5% versus 33.9%). CONCLUSIONS: Pathological gambling is substantially more prevalent among cocaine-dependent outpatients than in the general population. Patients with pathological gambling differ from other cocaine dependent outpatients in some sociodemographic characteristics but not in short term outcome of treatment for cocaine dependence. PMID- 10873924 TI - Ionotropic glutamate receptors and expression of N-methyl-D-aspartate receptor subunits in subregions of human hippocampus: effects of schizophrenia. AB - OBJECTIVE: Multiple quantifiable biologic abnormalities have been localized to the hippocampus in schizophrenia. Alterations in glutamate-mediated transmission at N-methyl-D-aspartic acid (NMDA)-sensitive receptors in hippocampus have been implicated in the pathophysiology of the illness. The authors tested the hypothesis that glutamatergic transmission within and efferent from hippocampus is altered in schizophrenia. METHOD: The authors analyzed postmortem hippocampal tissue from individuals with schizophrenia and from healthy individuals. The tissue samples had been collected by two brain tissue banks, one in Maryland and the other in Melbourne, Australia. lonotropic receptor binding for the NMDA, kainate, and (3)H-amino-3-hydroxy-5-methylisoxazol-4-propionate (AMPA) receptors was quantified by using usual radioligand techniques. In situ hybridization autoradiography was used to quantify mRNA for the NMDA receptor subunits NR1, NR2A, and NR2B. RESULTS: Ligand binding to the ionotropic glutamate receptors (NMDA, kainate, and AMPA) did not differ significantly overall or in any subregion between the schizophrenia tissue and the healthy comparison tissue. The only exception was AMPA receptor binding in hippocampal subregion CA2, which was slightly but significantly less in schizophrenia. However, the level of mRNA for the NMDA receptor subunits NR1 and NR2B was significantly different between groups; in several hippocampal subregions, the level of NR1 mRNA was lower and the level of NR2B mRNA higher in schizophrenia. CONCLUSIONS: Because the NR1 subunit of the NMDA receptor is critical to full receptor activity, a reduction of NR1 in hippocampus in schizophrenia suggests a functional impairment in glutamatergic transmission at the NMDA receptor, resulting in reduced glutamatergic transmission within and possibly efferent from the hippocampus in schizophrenia. This defect could underlie a hypoglutamatergic state in regions of limbic cortex, consistent with published results from other lines of research in schizophrenia. PMID- 10873923 TI - Elevated central serotonin transporter binding availability in acutely abstinent cocaine-dependent patients. AB - OBJECTIVE: Recent work has underscored the role of serotonergic neurotransmission in chronic neural adaptations to cocaine dependence. The authors tested for evidence of serotonergic dysfunction during acute abstinence from cocaine, a period of high risk for relapse in cocaine dependence. METHOD: Binding availability of dopamine transporters and serotonin transporters was measured in 15 cocaine-dependent subjects during acute abstinence and in 37 healthy comparison subjects by using [(123)I]beta-CIT and single photon emission computed tomography. RESULTS: Significant increases in diencephalic and brainstem serotonin transporter binding (16.7% and 31.6%, respectively) were observed in cocaine-dependent subjects. Brainstem serotonin transporter binding was significantly inversely correlated with age across diagnostic groups. CONCLUSIONS: These findings provide further evidence of serotonergic dysfunction during acute abstinence from chronic cocaine use. Age-related decline in brainstem serotonin transporter binding may underlie the poor response to selective serotonin reuptake inhibitor antidepressants seen in some elderly depressed patients. PMID- 10873926 TI - Use of self-ratings in the assessment of symptoms of attention deficit hyperactivity disorder in adults. AB - OBJECTIVE: The purpose of this research was to determine if adults can provide a true rating of their own childhood and current symptoms of attention deficit hyperactivity disorder (ADHD). METHOD: The authors conducted two studies. In study 1, 50 adult subjects completed a questionnaire assessing their ADHD symptoms in childhood. In addition, a parent of each subject completed a questionnaire rating the subject's childhood ADHD symptoms. In study 2, 100 adult subjects completed a questionnaire rating their own current ADHD symptoms. The subject's partner also completed a questionnaire rating the subject's current ADHD symptoms. The correlation between subject and observer ratings was measured in each study. Inattentive symptoms, hyperactive-impulsive symptoms, and total symptoms were analyzed. RESULTS: Good correlations were found between subject and observer scores in both studies. CONCLUSIONS: The diagnosis of ADHD in adults relies on an accurate recall of childhood behavior and an accurate account of current behavior. The results of this study suggest that adults can give a true account of their childhood and current symptoms of ADHD. PMID- 10873925 TI - Low incidence of persistent tardive dyskinesia in elderly patients with dementia treated with risperidone. AB - OBJECTIVE: The authors studied the incidence of tardive dyskinesia in elderly institutionalized patients with dementia being treated with risperidone. METHOD: After participating in a 12-week multicenter double-blind study during which they received placebo or one of three doses of risperidone, 330 patients (mean age=82.5 years) with Alzheimer's, vascular, or mixed dementia were enrolled in a 1-year open-label study during which they received flexible doses of risperidone. Persistent emergent tardive dyskinesia was defined according to scores on the dyskinesia subscale of the Extrapyramidal Symptom Rating Scale. RESULTS: The mean modal risperidone dose was 0.96 mg/day (SD=0.53), and the median length of risperidone use was 273 days. The 1-year cumulative incidence of persistent emergent tardive dyskinesia among the 255 patients without dyskinesia at baseline was 2.6%. Patients with dyskinetic symptoms at baseline experienced significant reductions in the severity of dyskinesia. Patients who received 0.75-1.5 mg/day of risperidone showed a significant improvement in psychopathologic symptoms over the 1-year period. CONCLUSIONS: Although there was no control group, the observed incidence of persistent tardive dyskinesia with risperidone seemed to be much lower than that seen in elderly patients treated with conventional neuroleptics. The average optimal dose of risperidone in elderly dementia patients was found to be 0.75-1.5 mg/day. PMID- 10873927 TI - Is the 5-HT(1Dbeta) receptor gene implicated in the pathogenesis of obsessive compulsive disorder? AB - OBJECTIVE: Obsessive-compulsive disorder (OCD) is a psychiatric condition for which strong evidence of a genetic component and serotonergic system involvement exists. Recent studies have shown that sumatriptan, a selective ligand of the serotonin (5-HT)(1Dbeta) autoreceptor, modifies OCD symptoms. The aim of this study was to investigate the presence of linkage disequilibrium between the 5 HT(1Dbeta) receptor gene, which has a variant caused by a silent G to C substitution at nucleotide 861 of the coding region, and OCD. METHOD: DNA was collected from 67 probands who met DSM-IV criteria for OCD and from their living parents or siblings. Transmission Disequilibrium Test/sib-Transmission Disequilibrium Test analyses were then conducted with the DNA data. RESULTS: Thirty-two families were informative for the analysis, which showed a preferential transmission of the G allele to the affected subjects. CONCLUSIONS: If the results are confirmed, there may be important implications for the 5 HT(1Dbeta) receptor gene in the pathogenesis and treatment of OCD. PMID- 10873928 TI - MDMA ("Ecstasy") abuse and high-risk sexual behaviors among 169 gay and bisexual men. AB - OBJECTIVE: The authors explored the association between abuse of 3, 4 methylenedioxymethamphetamine (MDMA, or "Ecstasy") and high-risk sexual behaviors among gay and bisexual men. METHOD: An anonymous questionnaire was completed by 169 gay and bisexual men at three New York City dance clubs. The questionnaire covered demographic indices, use of MDMA and other drugs, and history of high risk sexual behaviors. RESULTS: About one-third of the respondents reported MDMA use at least monthly. MDMA use was strongly and significantly associated with a history of recent unprotected anal intercourse. This association remained equally strong even after controlling for age, ethnicity, and all other forms of drug use, including alcohol. CONCLUSIONS: MDMA abuse, and its strong association with high-risk sexual behaviors, appears to represent important unexplored public health problems among some gay or bisexual men. PMID- 10873929 TI - Lack of interaction of buprenorphine with flunitrazepam metabolism. AB - OBJECTIVE: The authors' goal was to determine if the reported clinical adverse interaction of flunitrazepam and buprenorphine was caused by inhibition of drug metabolism. METHOD: Inhibition of flunitrazepam metabolism by buprenorphine and norbuprenorphine were determined in three human liver microsome preparations carrying the CYP2C19*1/*1 allele. Omeprazole metabolism mediated by CYP2C19 and CYP3A4 was used as a control reaction. Apparent K(i) values were determined. RESULTS: Norbuprenorphine did not inhibit the metabolism of flunitrazepam or omeprazole. Buprenorphine inhibited the formation of CYP3A4-mediated pathways of 3-hydroxyflunitrazepam and omeprazole sulfone formation (K(i) 118 and 16 microM) in human liver microsomes. Corresponding values were 38 and 90 microM in cDNA expressed CYP3A4 microsomes. Projected in vivo inhibition of CYP3A4-mediated metabolism of flunitrazepam by buprenorphine is 0. 1%-2.5%. Estimated inhibition of buprenorphine N-dealkylation by flunitrazepam in vivo is 0.08%. CONCLUSIONS: The clinical interaction of flunitrazepam and buprenorphine is likely based on a pharmacodynamic mechanism. PMID- 10873930 TI - Smaller brain size associated with unawareness of illness in patients with schizophrenia. AB - OBJECTIVE: Although several neuropsychological studies have supported the notion of frontal and parietal lobe involvement in unawareness of illness in schizophrenia, neuroanatomic differences have not been examined. METHOD: Thirty patients with schizophrenia spectrum disorder were rated by means of a structured interview assessing awareness of illness and performance on clinical rating scales. With 13 healthy comparison subjects, they underwent neuropsychological assessment and a scan using three-dimensional, spoiled gradient recall acquisition volumetric magnetic resonance imaging. RESULTS: Patients who were relatively unaware of their illness had smaller brain and intracranial volumes (brain tissue plus CSF) than either aware patients or normal comparison subjects, who did not differ significantly from each other. CONCLUSIONS: These findings suggest that unawareness of illness is an important phenomenological feature with neurological correlates that is seen in at least one subgroup of patients with schizophrenia. PMID- 10873931 TI - Prenatal exposure to famine and brain morphology in schizophrenia. AB - OBJECTIVE: The authors assessed the effects of nutritional deficiency during the first trimester of pregnancy on brain morphology in patients with schizophrenia. METHOD: Nine schizophrenic patients and nine healthy comparison subjects exposed during the first trimester of gestation to the Dutch Hunger Winter were evaluated with magnetic resonance brain imaging, as were nine schizophrenic patients and nine healthy subjects who were not prenatally exposed to the famine. RESULTS: Prenatal famine exposure in patients with schizophrenia was associated with decreased intracranial volume. Prenatal Hunger Winter exposure alone was related to an increase in brain abnormalities, predominantly white matter hyperintensities. CONCLUSIONS: Nutritional deficiency during the first trimester of gestation resulted in an increase in clinical brain abnormalities and was associated with aberrant early brain development in patients with schizophrenia. Stunted brain development secondary to factors that affect brain growth during the first trimester of gestation may thus be a potential risk factor for developing schizophrenia. PMID- 10873932 TI - Human leukocyte antigen and season of birth in Japanese patients with schizophrenia. AB - OBJECTIVE: Five Japanese studies, to the authors' knowledge, without exception, have consistently shown an increased frequency of human leukocyte antigen (HLA) DR1 in patients with schizophrenia. This suggests an association between HLA-DR1 and schizophrenia in the Japanese population. The mechanism of the association is unknown; however, prenatal infections may be involved. The present study explored factors, including winter birth, that might correlate with this mechanism. Age at onset and gender were also studied. METHOD: Factors were compared between Japanese patients with schizophrenia with and in those without HLA-DR1 (N=60 and N=307, respectively). RESULTS: A significantly higher incidence of births in February and March was observed in patients with (31.7%) than those without (15. 6%) HLA-DR1. No association was found between the presence of HLA-DR1 and other variables. CONCLUSIONS: Although this result is preliminary, it may suggest an interaction between HLA and winter birth in the development of schizophrenia in the Japanese population. PMID- 10873933 TI - N-Acetylaspartate concentration in the anterior cingulate of maltreated children and adolescents with PTSD. AB - OBJECTIVE: Anterior cingulate dysfunction has been implicated in the pathophysiology of posttraumatic stress disorder (PTSD). The authors hypothesized that integrity of the anterior cingulate may be affected in childhood PTSD. METHOD: Single voxel proton magnetic resonance spectroscopy (proton MRS) was used to measure the relative concentration of N-acetylaspartate and creatine, a marker of neural integrity, in the anterior cingulate of 11 children and adolescents who met DSM-IV criteria for PTSD secondary to maltreatment and 11 healthy matched comparison subjects. RESULTS: The ratio of N-acetylaspartate to creatine was significantly lower in the maltreated subjects with PTSD than in the comparison subjects. CONCLUSIONS: The lower N-acetylaspartate/creatine ratio in subjects with PTSD suggests that anterior cingulate neuronal metabolism may be altered in childhood PTSD. PMID- 10873934 TI - Optimal risperidone dose in drug-naive, first-episode schizophrenia. PMID- 10873935 TI - Aggression in dementia with lamotrigine treatment. PMID- 10873937 TI - Psychiatrists' attitudes toward dissociative disorders diagnoses. PMID- 10873941 TI - Cognitive deficits in obsessive-compulsive disorder. PMID- 10873942 TI - Recurrence of geriatric depression. PMID- 10873945 TI - Sex differences in cerebral metabolism among abstinent cocaine users. PMID- 10873955 TI - Rheumatoid arthritis and Epstein-Barr virus: a case of living with the enemy? PMID- 10873956 TI - Teaching rheumatology in primary care. PMID- 10873957 TI - Consensus statement on the initiation and continuation of tumour necrosis factor blocking therapies in rheumatoid arthritis. PMID- 10873959 TI - Clinical assessment of joints and spine PMID- 10873958 TI - Synovial biopsy in arthritis research: five years of concerted European collaboration. PMID- 10873960 TI - Distribution of distal femoral osteophytes in a human skeletal population. AB - OBJECTIVES: To examine objectively spatial patterns of osteophytes around the distal end of the femur and to identify distinct subgroups. METHODS: A sample of 107 human femora from a large skeletal population were selected for study. These femora included subjects with evidence of late stage osteoarthritis (that is, with eburnation present) and those with no such evidence. The location of osteophytes was recorded using a video camera and digitised computer images were extracted. Multidimensional scaling was used to identify clusters of femora based upon osteophyte location. RESULTS: A distinct subgroup of femora was identified with osteophytes present only within the intercondylar notch region. None of these subjects had any evidence of eburnation. CONCLUSIONS: This finding adds to an earlier study based on radiographs. Osteophytes located within the intercondylar notch of the femur appear to be a distinct subset, which may occur either as an early stage of knee osteoarthritis or for some independent reason. PMID- 10873961 TI - Magnetic resonance imaging, radiography, and scintigraphy of the finger joints: one year follow up of patients with early arthritis. The TIRA Group. AB - OBJECTIVES: To evaluate synovial membrane hypertrophy, tenosynovitis, and erosion development of the 2nd to 5th metacarpophalangeal (MCP) and proximal interphalangeal (PIP) joints by magnetic resonance imaging in a group of patients with rheumatoid arthritis (RA) or suspected RA followed up for one year. Additionally, to compare the results with radiography, bone scintigraphy, and clinical findings. PATIENTS AND METHODS: Fifty five patients were examined at baseline, of whom 34 were followed up for one year. Twenty one patients already fulfilled the American College of Rheumatology (ACR) criteria for RA at baseline, five fulfilled the criteria only after one year's follow up, whereas eight maintained the original diagnosis of early unclassified polyarthritis. The following MRI variables were assessed at baseline and one year: synovial membrane hypertrophy score, number of erosions, and tenosynovitis score. RESULTS: MRI detected progression of erosions earlier and more often than did radiography of the same joints; at baseline the MRI to radiography ratio was 28:4. Erosions were exclusively found in patients with RA at baseline or fulfilling the ACR criteria at one year. At one year follow up, scores of MR synovial membrane hypertrophy, tenosynovitis, and scintigraphic tracer accumulation had not changed significantly from baseline; in contrast, swollen and tender joint counts had declined significantly (p<0.05). CONCLUSIONS: MRI detected more erosions than radiography. MR synovial membrane hypertrophy and scintigraphy scores did not parallel the changes seen over time in clinically assessed swollen and tender joint counts. Although joint disease activity may be assessed as quiescent by conventional clinical methods, a more detailed evaluation by MRI may show that a pathological condition is still present within the synovium. PMID- 10873962 TI - Effect of interleukin 17 on proteoglycan degradation in murine knee joints. AB - OBJECTIVE: To evaluate the effect of murine interleukin 17 (IL17) on cartilage catabolism and joint inflammation by direct intra-articular injection of the cytokine into murine knee joints. METHODS: Knees of normal C57 Bl mice were injected once or repeatedly with recombinant IL17 or IL1beta. Inflammation was estimated by technetium-99m pertechnetate ((99)Tc) uptake and histological scoring of tissue sections. Proteoglycan depletion was evaluated by histological scoring of safranin O stained sections. Effects on proteoglycan synthesis were studied by (35)SO(4) incorporation. RESULTS: A single intra-articular injection of IL17 (10 ng/knee) produced effects very similar to those of IL1beta (10 ng/knee). No inflammation was detected at six or 24 hours by (99)Tc uptake. However, safranin O staining showed depletion of proteoglycan at 48 hours. Repeated injections of IL17 induced joint inflammation and cartilage proteoglycan depletion as shown by histological scoring. Unlike IL1beta, proteoglycan depletion induced by IL17 seemed to be the result of increased degradation only, as no suppression of (35)SO(4) incorporation was seen. CONCLUSION: These findings confirm, in vivo, the catabolic effects of IL17 on cartilage. IL17 is thus the first T cell cytokine showing a direct catabolic effect on cartilage in addition to stimulatory effects on macrophages and synoviocytes, making it a potentially important cytokine in the pathogenesis of arthritis. PMID- 10873963 TI - Decreased T cell precursor frequencies to Epstein-Barr virus glycoprotein Gp110 in peripheral blood correlate with disease activity and severity in patients with rheumatoid arthritis. AB - OBJECTIVES: Rheumatoid arthritis (RA) is a chronic joint disease associated with certain HLA-DR alleles expressing the QK/RRAA motif or shared epitope. The Epstein-Barr virus (EBV) has been suspected to be a causative factor for RA. The EBV gp110, a glycoprotein of the replicative cycle that contains a copy of the shared epitope, constitutes an important target in the immune control of EBV replication. This study evaluated the specific T cell response to EBV gp110 in patients with RA expressing or not the shared epitope and examined whether this immune cellular response might be related to disease activity and severity. METHODS: 25 patients with RA were studied and compared with 25 healthy controls. Disease activity was assessed by biochemical markers of inflammation (erythrocyte sedimentation rate (ESR) and C reactive protein (CRP) levels). Disease severity was defined by extra-articular disease (vasculitis, subcutaneous nodules, or other organ disease). The frequencies of peripheral blood T cells specific for EBV gp110 and a control protein (total protein extract from Escherichia coli) were determined by direct limiting dilution analysis without preliminary bulk culture. RESULTS: The gp110 precursor frequencies ranged from 0 to 20 x 10(-6) in patients with RA and controls. The mean gp110 T cell precursor frequency was lower in patients with RA (SD 3.2 (4.4) x 10(-6)) than in healthy controls (4.1 (3.8) x 10(-6)) (p = 0.02). No difference was found for the control protein (p = 0.09). Both shared epitope positive and negative patients with RA responded to gp110, without significant difference. A negative correlation between both ESR and CRP levels and the gp110 T cell response was found (r = -0.71, p<0.0001 and r = -0.42, p = 0.038, respectively). Finally, patients with extra-articular disease displayed the lowest immune cellular response to EBV gp110. CONCLUSION: These results suggest that patients with RA have a decreased T cell response to EBV gp110. Since gp110 is an important protein in the control of EBV replication, this might lead to a poor control of EBV infection, chronic exposure to other EBV antigens, and thus to a chronic inflammatory response in patients with RA. PMID- 10873964 TI - Beneficial effects of weight loss associated with moderate calorie/carbohydrate restriction, and increased proportional intake of protein and unsaturated fat on serum urate and lipoprotein levels in gout: a pilot study. AB - OBJECTIVES: Insulin resistance (IR) has been increasingly implicated in the pathogenesis of gout. The lipoprotein abnormalities described in hyperuricaemic subjects are similar to those associated with IR, and insulin influences renal urate excretion. In this study it was investigated whether dietary measures, reported to be beneficial in IR, have serum uric acid (SU) and lipid lowering effects in gout. METHODS: Thirteen non-diabetic men (median age 50, range 38-62) were enrolled. Each patient had had at least two gouty attacks during the four months before enrollment. Dietary recommendations consisted of calorie restriction to 6690 kJ (1600 kcal) a day with 40% derived from carbohydrate, 30% from protein, and 30% from fat; replacement of refined carbohydrates with complex ones and saturated fats with mono- and polyunsaturated ones. At onset and after 16 weeks, fasting blood samples were taken for determination of SU, serum cholesterol (C), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), and triglycerides (TGs). Results were expressed as median (SD). RESULTS: At onset, the body mass index (BMI) was 30.5 (8.1) kg/m(2). Dietary measures resulted in weight loss of 7.7 (5.4) kg (p=0.002) and a decrease in the frequency of monthly attacks from 2.1 (0.8) to 0.6 (0.7) (p=0.002). The SU decreased from 0.57 (0.10) to 0.47 (0.09) mmol/l (p=0.001) and normalised in 7 (58%) of the 12 patients with an initially raised level. Serum cholesterol decreased from 6.0 (1.7) to 4.7 (0. 9) mmol/l (p=0.002), LDL-C from 3.5 (1.2) to 2.7 (0.8) mmol/l (p=0. 004), TGs from 4.7 (4.2) to 1.9 (1.0) mmol/l (p=0.001), and C:HDL-C ratios from 6.7 (1.7) to 5.2 (1.0) (p=0.002). HDL-C levels increased insignificantly. High baseline SU, frequency of attacks, total cholesterol, LDL-C and TG levels, and total C:HDL-C ratios correlated with higher decreases in the respective variables upon dietary intervention (p<0.05). CONCLUSION: The results suggest that weight reduction associated with a change in proportional macronutrient intake, as recently recommended in IR, is beneficial, reducing the SU levels and dyslipidaemia in gout. Current dietary recommendations for gout may need re-evaluation. PMID- 10873965 TI - Raised human cartilage glycoprotein-39 plasma levels in patients with rheumatoid arthritis and other inflammatory conditions. AB - OBJECTIVE: To evaluate plasma human cartilage glycoprotein (HC gp-39) as a possible marker for the presence and/or activity of rheumatoid arthritis (RA) and other inflammatory conditions. BACKGROUND: HC gp-39 is a secretory product of chondrocytes, synovial cells, macrophages, and neutrophils. HC gp-39, also described as YKL-40, was found to be a marker of joint disease and tissue injury in RA and various other diseases. METHODS: Levels of HC gp-39 were determined by a sandwich enzyme linked immunosorbent assay (ELISA) in 47 patients with RA, 47 with osteoarthritis (OA), 24 with systemic lupus erythematosus (SLE), 24 with inflammatory bowel disease (IBD), and in 47 healthy controls. A disease activity score was assessed in the patients with RA, SLE, and IBD. RESULTS: The plasma level of HC gp-39 in the RA patient group was significantly higher than in the other patient groups and healthy controls. The level in patients with OA, SLE, and IBD was also significantly higher than the HC gp-39 level found in the healthy control group. HC gp-39 levels in patients with RA correlated positively with the ESR and IgM rheumatoid factor level but not with other variables of disease activity. In the patients with SLE and IBD no correlation was found with the disease activity score. CONCLUSION: The plasma level of HC gp-39 is increased in inflammatory conditions with and without joint disease (SLE, IBD, OA, and RA). Thus increased levels of HC gp-39 do not only reflect joint disease but also reflect inflammation or tissue degradation in various conditions. Notably, the highest level of HC gp-39 was found in patients with RA. Only in the RA patient group was a correlation between HC gp-39 plasma levels and some laboratory variables of disease activity found. PMID- 10873966 TI - Labour force participation among patients with rheumatoid arthritis. AB - OBJECTIVES: To assess work history and labour force participation among patients with rheumatoid arthritis (RA) in the Netherlands. METHODS: A random sample of 1056 patients with RA aged 16-59 years from 17 rheumatology practices in the Netherlands was examined. Data on disease status and outcome were obtained by a questionnaire including standardised instruments, such as the Rapid Assessment of Disease Activity in Rheumatology (RADAR) and RAND-36 questionnaires. Labour force participation was defined as having a paid job. RESULTS: Of the study group with a mean disease duration of 12 years, 35.7% held a paid job (men 56.7%; women 27.7%). When standardised for age, sex, and educational level, the labour force participation of patients with RA was 61.2% compared with 65.5% for the general population, which was not statistically significant. Disease duration of six years and more was negatively associated with labour force participation. CONCLUSIONS: After controlling for the confounding effects of age, sex, and education, the labour force participation of patients with RA in the Netherlands is only slightly lower than that of the general population. PMID- 10873967 TI - Neutrophil function in pregnancy and rheumatoid arthritis. AB - BACKGROUND: Pregnancy exerts suppressive effects on rheumatoid arthritis (RA). An attenuation in neutrophil function in late pregnancy which may explain this amelioration has previously been reported. OBJECTIVE: A longitudinal investigation of neutrophil activity in healthy pregnant women (n=9) and pregnant patients with RA (n=9), compared with age matched non-pregnant patients with RA (n=12) and healthy controls (n=22). METHODS: Neutrophil activation was measured in response to the physiological receptor agonists, n-formyl-methionyl-leucyl phenylalanine (fMLP) and zymosan activated serum (ZAS). Superoxide anion production (respiratory burst) was determined by lucigenin enhanced chemiluminescence (LUCL); secondary granule lactoferrin release by enzyme linked immunosorbent assay (ELISA); and CD11b, CD18, and CD62L expression by flow cytometric analysis. RESULTS: Stimulated neutrophil LUCL was significantly reduced in both pregnant women with RA and healthy pregnant women in the second (fMLP 43% and 69%, ZAS 43% and 59%, respectively) and third trimesters (fMLP 24% and 44%, ZAS 32% and 38%, respectively). Responses returned to normal within eight weeks of delivery and unstimulated levels remained unchanged throughout pregnancy. Basal and stimulated CD11b, CD18, and CD62L expression showed no variations throughout gestation for both pregnancy groups. Likewise, stimulated lactoferrin release and plasma lactoferrin remained unchanged. Certain morphological differences in RA neutrophils were highlighted by the flow cytometric analysis. Moreover, resting neutrophils and stimulated cells from patients with RA, including pregnant subjects, showed a marked increase in LUCL, but a reduction in CD11b, CD18, and CD62L. Low dose prednisolone and methylprednisolone had no effect on neutrophil parameters over the period of treatment with non-steroidal anti-inflammatory drugs. CONCLUSION: The attenuation to neutrophil respiratory burst in both healthy and RA pregnancies may offer an explanation for the pregnancy induced remission of this inflammatory disorder. PMID- 10873968 TI - Effect of a three month course of ciprofloxacin on the outcome of reactive arthritis. AB - BACKGROUND: Treatment of reactive arthritis (ReA) with antibiotics has so far remained controversial. Eradication of the causative microbe appears logical, but short term antibiotic treatment has no beneficial effect on the outcome of ReA. OBJECTIVE: To evaluate the effect of a three month course of ciprofloxacin on ReA. METHODS: In a randomised, double blind, placebo controlled trial, between December 1992 and February 1996, 71 patients with acute ReA triggered by a gastrointestinal or a urogenital infection were randomly assigned to receive ciprofloxacin 500 mg or placebo twice daily for three months. Patients were assessed at study entry, at 6 weeks, 3 months, 6 months, and 12 months. Sixty two patients were valid for the efficacy analysis. The primary outcome measures were erythrocyte sedimentation rate, number of swollen joints, patients self assessment, and complete recovery. RESULTS: Adverse events were mostly mild and occurred in both treatment groups. There were no statistically significant differences in any of the primary or secondary efficacy variables between the study groups at baseline or during the 12 month follow up. All primary outcome measures indicated that the condition of the patients improved during the study. CONCLUSION: Both groups tended to recover. Ciprofloxacin, given as a three month course, had no advantage over placebo treatment. PMID- 10873969 TI - Ankylosing spondylitis in north Jordan: descriptive and analytical study. AB - OBJECTIVE: A study of ankylosing spondylitis in Jordan, which has been under investigated in this region. METHODS: Twenty two patients were studied according to standard methodology during a period of four years. Information on HLA, the presence of uveitis, cardiac disease, and peripheral arthritis was recorded. Other variables such as age, sex, employment, and level of disability were also recorded. RESULTS: The results reflected the characteristics of the illness and the impact of the disease on the patients and their quality of life and were consistent with the findings of other workers in the region. PMID- 10873970 TI - Care to share? PMID- 10873971 TI - Selective ganglion cell death in glaucoma. PMID- 10873972 TI - What is Sorsby's fundus dystrophy? PMID- 10873973 TI - Sorsby fundus dystrophy without a mutation in the TIMP-3 gene. AB - AIMS: To examine a large family with an autosomal dominant fundus dystrophy and to investigate whether or not mutations in TIMP-3 gene were involved. METHODS: A large family of 58 individuals with an autosomal dominant fundus dystrophy was examined ophthalmologically. A DNA linkage analysis in the 22q12.1-q13.2 region was performed. The TIMP-3 gene was screened for mutations in all five exons. RESULTS: In this large family 15 individuals were affected. All other individuals were found to be clinically unaffected. Pisciform flecks in the midperiphery and drusen-like deposits were the most typical ophthalmological finding in this family and were encountered from the fifth decade on. Chorioretinal atrophy and neovascularisation with disciform lesions characterised the disease from the sixth decade on. Linkage analysis using an affected only analysis, showed a maximum positive lod score of 3.94 at theta = 0.0 with marker D22S283. No mutations possibly causing Sorsby fundus dystrophy were found in either the exonic sequences, the promotor region, or the 3'UTR. CONCLUSION: The family in this pedigree has an autosomal dominant fundus dystrophy, which is most probably Sorsby fundus dystrophy. Although, in the linkage analysis, significant positive lod scores were found with the region 22q12.1-q13.2, no causative mutations could be identified in the TIMP-3 gene. PMID- 10873974 TI - In vitro antibiotic resistance in bacterial keratitis in London. AB - AIM: To document changes in the profile of bacterial isolates from cases of keratitis and changes in their susceptibility to first line antibiotic therapies. METHODS: A retrospective review was performed of all bacterial isolates from cases of keratitis seen between 1984 and 1999. In vitro laboratory susceptibilities to antibiotics were determined by the Kirby-Bauer disc diffusion method. The number of isolates, changes in the proportion of bacterial types, and the number that were fully resistant to monotherapy (ofloxacin), dual therapy (gentamicin and cefuroxime), and prophylactic treatment (chloramphenicol) were calculated. RESULTS: There were 1312 bacterial isolates over 16 years. Gram positive bacteria accounted for 54.7% of isolates and Staphylococcus species (33.4%) were the most frequently isolated organisms. During the study period there has been an increase in the proportion of Pseudomonas species isolates but no overall increase in the proportion of Gram negative isolates. There has not been an increase in the proportion of isolates resistant to ofloxacin since 1995 or an increase in resistance to the combination of gentamicin and cefuroxime. However, since 1984 there has been a significant increase in proportion of Gram negative organisms resistant to chloramphenicol (p=0.0019). CONCLUSIONS: An increase in the in vitro resistance of organisms to first line therapies for bacterial keratitis has not been observed. An increased resistance to chloramphenicol indicates that this drug is unlikely to provide prophylactic cover when Gram negative infection is a risk. Continued monitoring for the emergence of antibiotic resistance is recommended. PMID- 10873975 TI - Reassessment of the corneal endothelial cell organisation in children. AB - AIM: To assess uniformity of the corneal endothelial cell mosaic in children. METHODS: 36 healthy children (5-11 years old, 16 boys, 20 girls) were assessed by specular microscopy. Endothelial cell density (ECD) was calculated from measured cell areas, and the number of sides/cell noted. RESULTS: Average values for ECD and cell areas were 3987 cells/mm(2) (95% CI 3806 to 4168 cells/mm(2)) and 278 (SD 85) mm(2) respectively, with normal distribution (COV 28. 2%, range 17.4 to 39.2%) and with the average percentage of six sided cells being 66.6% (8.8%). Cell area was positively correlated to number of cell sides (p <0.01, r(2)=0.993), but the percentage of six sided cells was negatively correlated to ECD (p <0.01, r=0.493). CONCLUSION: A high ECD occurs in children, but this does not mean there is a high percentage of "hexagons". PMID- 10873976 TI - The lens in hereditary hyperferritinaemia cataract syndrome contains crystalline deposits of L-ferritin. AB - BACKGROUND/AIM: Hereditary hyperferritinaemia cataract syndrome (HHCS) is an autosomal dominant disorder characterised by elevated serum L-ferritin and bilateral cataracts. The ocular manifestations of this disorder are poorly studied. This study therefore sought to determine the origin of cataracts in HHCS. METHODS: L-ferritin ELISA, immunohistochemical and ultrastructural analysis of a lens nucleus from an HHCS individual. RESULTS: The HHCS lens L-ferritin content was 147 microg/g dry weight of lens compared with <16 microg/g for a non HHCS control cataract lens. The cataract comprised discrete crystalline inclusions with positive staining with anti-L-ferritin but not anti-H-ferritin. CONCLUSIONS: This unusual finding of crystalline opacities in the lens may be unique to HHCS and is likely to result from disturbed metabolism of L-ferritin within the lens or an abnormal interaction between L-ferritin and lens proteins. PMID- 10873977 TI - Graft failure in human donor corneas due to transmission of herpes simplex virus. AB - AIM: To report the clinical consequences of contamination of human donor corneas by herpes simplex virus (HSV) in organ culture. METHODS: Two patients without previous history of ocular HSV infection underwent penetrating keratoplasty (PK), one for keratoconus and the other for Fuchs' endothelial dystrophy. One patient suffered primary graft failure while the other developed a persistent epithelial defect, ultimately resulting in graft failure. Viral culture of swabs taken from both corneas during the early postoperative period was undertaken. The failed donor corneas were examined histopathologically by immunohistochemistry (IHC) for HSV-1 antigens, transmission electron microscopy (TEM), and by polymerase chain reaction (PCR) for HSV DNA. Both failed corneas were replaced within 6 weeks of the initial surgery. The records of the fellow donor corneas were also examined for evidence of infection. RESULTS: HSV was cultured from both corneas during the early postoperative period. Histology of both donor corneas demonstrated a thickened corneal stroma with widespread necrosis of keratocytes and loss of endothelial cells. IHC showed keratocytes positive with antibodies to HSV-1 antigens. TEM demonstrated HSV-like viral particles within degenerating keratocytes. PCR performed on the failed corneal grafts was positive for HSV-1 DNA, whereas PCR performed on the excised host corneal buttons was negative in both patients. Records of the fellow donor corneas showed that one cornea was successfully transplanted into another recipient after 18 days in organ culture, whilst the other was discarded because of extensive endothelial cell necrosis noted after 15 days in organ culture. CONCLUSION: HSV within a donor cornea may cause endothelial destruction in organ culture and both primary graft failure and ulcerative keratitis after transplantation. Endothelial necrosis of a donor cornea in culture also raises the possibility of HSV infection within the fellow cornea. PMID- 10873978 TI - Population based assessment of uveitis in an urban population in southern India. AB - AIM: To assess the prevalence of active and inactive uveitis unrelated to previous surgery or trauma in an urban population in southern India. METHODS: As part of the Andhra Pradesh Eye Disease Study, 2522 subjects (85.4% of those eligible), a sample representative of the population of Hyderabad city in southern India, underwent interview and detailed dilated eye examination. Presence of sequelae of uveitis without current active inflammation was defined as inactive uveitis. RESULTS: Unequivocal evidence of active or inactive uveitis unrelated to previous surgery or trauma was present in 21 subjects, an age-sex adjusted prevalence of 0.73% (95% confidence interval (CI) 0.44-1.14%). Active uveitis was present in eight subjects, an age-sex adjusted prevalence of 0.37% (95% CI 0. 19-0.70), of which 0.06% was anterior, 0.25% intermediate, and 0.06% posterior. The 0.36% (95% CI 0.17-0.68%) prevalence of inactive uveitis included macular chorioretinitis scars (0.26%), anterior (0. 07%) and previous vasculitis involving the whole eye (0.03%). The prevalence of visual impairment due to uveitis of less than 6/18 in at least one eye was 0.27%, less than 6/60 in at least one eye was 0. 16%, and less than 6/60 in both eyes was 0.03%. CONCLUSION: These population based cross sectional data give an estimate of the prevalence of various types of uveitis in this urban population in India. Active or past uveitis that might need treatment at some stage was present in one of every 140 people in this population. PMID- 10873979 TI - Effects of glaucoma medications on the cardiorespiratory and intraocular pressure status of newly diagnosed glaucoma patients. AB - AIMS: To evaluate the short term cardiovascular, respiratory, and intraocular pressure (IOP) effects of four glaucoma medications in newly diagnosed glaucoma patients. METHODS: 141 newly diagnosed glaucoma patients were recruited and underwent a full ocular, cardiovascular, and respiratory examination, including an electrocardiogram (ECG) and spirometry. They were prescribed one of four topical glaucoma medications and reviewed 3 months later. One eye of each patient was randomly chosen for analysis, performed using analysis of variance and the chi(2) test. RESULTS: Latanoprost had the greatest mean IOP lowering effect in both the primary open angle glaucoma (POAG) (p = 0.005) and the "presumed" normal tension glaucoma (NTG) groups (p = 0.33), reducing the IOP by 8.9 mm Hg and 4.1 mm Hg respectively. Timolol was associated with lowered pulse rates and reductions in the spirometry measurements. 41% of patients using brimonidine complained of systemic side effects and over 55% of patients using betaxolol complained of ocular irritation. 28% of patients required an alteration in their glaucoma management. CONCLUSIONS: Latanoprost appears to be a useful primary treatment for glaucoma patients, in view of superior IOP control and a low incidence of local and systemic side effects. Timolol causes a reduction in measurements of respiratory function, a concern in view of the potential subclinical reversible airways disease in the elderly glaucoma population. Brimonidine is associated with substantial, unpredictable systemic side effects and betaxolol causes ocular irritation and weak IOP control. Spirometry is advised in all patients receiving topical beta blocker therapy to control their glaucoma. PMID- 10873980 TI - Trabeculectomy with mitomycin C for post-keratoplasty glaucoma. AB - AIM: To investigate the effect of trabeculectomy with and without mitomycin C in post-keratoplasty glaucoma. METHODS: A retrospective study was performed on patients who underwent trabeculectomy for glaucoma after penetrating keratoplasty. 34 eyes of 32 patients were included in this study. 26 eyes received trabeculectomy with mitomycin C and eight eyes without mitomycin C. The procedure was deemed successful if the intraocular pressure was maintained below 21 mm Hg with or without use of additional antiglaucoma medication (mean follow up time 22.3 (SD 10.3) months). RESULTS: At the last examination trabeculectomy was successful in 19 of 26 eyes (73.0%) with mitomycin C (+) and two of eight (25.0%) without (p=0.0219). When the prognosis was analysed by Kaplan-Meier curve, the mitomycin C (+) group showed a better prognosis (p=0.0182). Mean intraocular pressure and average number of glaucoma medications improved in the group with mitomycin C without severe side effects on the graft. Graft rejection after trabeculectomy was seen in two eyes in the mitomycin C group. Final graft clarity rate was 69.2% (18/26) in the mitomycin C (+) group and 37.5% (3/8) in the mitomycin C (-) group. Complications such as persistent epithelial defect, cystoid macular oedema, choroidal detachment, leakage from bleb were seen in four eyes in the mitomycin C (+) group and in one eye in the mitomycin C (-) group. CONCLUSIONS: Trabeculectomy with mitomycin C showed better results for glaucoma following keratoplasty. PMID- 10873981 TI - Efficacy and safety of the Ahmed glaucoma valve implant in Chinese eyes with complicated glaucoma. AB - AIMS: To evaluate the efficacy and safety of the Ahmed glaucoma valve implant in Chinese eyes with complicated glaucomas. METHODS: This retrospective study reviewed the final intraocular pressure, visual outcome, and incidence of complications in all patients with the Ahmed glaucoma valve implant performed at the Prince of Wales Hospital, Hong Kong, between June 1996 and November 1998. RESULTS: A total of 65 eyes from 60 patients were treated with the Ahmed glaucoma implant. At a mean follow up (SD, median) of 21.8 (9.2, 28. 0) months (range 6-37 months), the mean intraocular pressure was reduced from 37.0 (SD 12.1) mm Hg before the implant surgery to 16.1 (12.4) mm Hg at the last follow up after surgery. The success rate of intraocular pressure control of <22 mm Hg was achieved in 73.8% of operated eyes. Transient postoperative hypotony with shallow anterior chamber occurred in 10.8% of cases. The most common postoperative complication was the formation of encapsulated bleb (24.6%). CONCLUSIONS: The Ahmed glaucoma valve implant appears to be effective and relatively safe for treating complicated glaucomas in Chinese eyes. The success rate is comparable with those reported in non-Asian eyes. Formation of postoperative encapsulated bleb is, however, more commonly encountered. PMID- 10873982 TI - Genetic screening in a large family with juvenile onset primary open angle glaucoma. AB - AIMS: A number of genetic loci have been implicated in the pathogenesis of primary open angle glaucoma (POAG). The aim of this study was to identify the genetic cause of POAG in a large Scottish family and, if possible, offer genetic screening and advice to family members. METHODS: Family members were examined to determine their disease status. Base excision sequence scanning was carried out in order to test for the presence of a POAG causing mutation at known genetic loci. Direct DNA sequencing was performed in order to determine the mutation sequence. RESULTS: All family members of known affected disease status and two family members of unknown disease status were found to have a mutation in the TIGR gene. The mutation resulted in the substitution of a glycine residue with an arginine residue at codon 252 (Gly252Arg). No other sequence variations were present in any members of the family. CONCLUSION: The Gly252Arg mutation in the TIGR gene results in the development of POAG in this family. It was possible to identify younger, currently unaffected, members of the family who carry the mutation and who are therefore at a very high risk of developing POAG themselves. This is the first demonstration that Gly252Arg can be a disease causing mutation rather than a benign polymorphism. The possible pathogenic mechanisms and wider implications of the mutation are considered. PMID- 10873983 TI - Increased polymorphonuclear leucocyte rigidity in HIV infected individuals. AB - AIM: Individuals with human immunodeficiency virus (HIV) infection were evaluated for evidence of abnormal polymorphonuclear leucocyte (PMN) rigidity, which can alter capillary blood flow. METHODS: The transit time of individual PMN through 8 microm pores in a cell transit analyser was used as a measure of cell rigidity. PMN transit time was compared between HIV infected individuals (n=45) with and without CMV retinitis and HIV negative controls (n=17). RESULTS: Transit times were longer for PMN from HIV infected individuals than for PMN from controls (p<0.001). PMN from HIV infected individuals with CMV retinitis (n=13) had longer transit times than PMN from those without CMV retinitis (n=32, p<0.001). Transit times were longer in HIV infected individuals with lower CD4+ T lymphocyte counts (p<0.001). Regression analysis indicated that the relation between transit times and the presence of CMV retinitis could not be explained solely on the basis of low CD4+ T lymphocytes. In HIV infected individuals, mean transit time was not correlated with age, blood pressure, or serum creatinine, cholesterol, or triglycerides. CONCLUSIONS: HIV infected individuals appear to have increased PMN rigidity, a cellular change that might be involved in the pathogenesis of HIV related retinal microvasculopathy. PMN rigidity appears to be related to severity of immune dysfunction. PMN rigidity may remain high in patients with CMV retinitis after elevations of CD4+ T lymphocyte counts that result from potent antiretroviral therapy. PMID- 10873984 TI - Hepatocyte growth factor in vitreous and serum from patients with proliferative diabetic retinopathy. AB - BACKGROUND: Hepatocyte growth factor (HGF) is an endothelium specific growth factor that has been implicated in angiogenesis, a crucial event for the development of proliferative diabetic retinopathy (PDR). The aim of the study is to determine the intravitreous concentrations of HGF in diabetic patients with PDR, and to investigate whether its serum levels could contribute to its intravitreous concentration. METHODS: 17 diabetic patients and seven non-diabetic patients in whom a vitrectomy was performed were studied. Both groups were matched by serum levels of HGF. Venous blood and vitreous samples were collected simultaneously at the time of vitreoretinal surgery. Vitreous and serum HGF were determined by ELISA. RESULTS: Intravitreous concentrations of HGF (median and range) were higher in diabetic patients (17.04 ng/ml (9.98-80)) in comparison with non-diabetic patients (5.88 ng/ml (2.57-14.20); p=0. 003). Intravitreous HGF concentrations were strikingly higher than serum HGF concentrations both in diabetic patients (17.04 ng/ml (9. 98-80) v 0.66 ng/ml (0.26-1.26); p<0.001) and in the control group (5.88 ng/ml (2.57-14.20) v 0.68 ng/ml (0.49-0.96); p=0.003). No correlation was found between serum and vitreous levels of HGF in both groups (diabetic patients, r= -0.31; p=0.5 and control subjects r= -0.15; p=0.5). CONCLUSION: The high vitreous levels of HGF observed in diabetic patients with PDR cannot be attributed to serum diffusion across the blood-retinal barrier. Therefore, intraocular synthesis appears to be the main contributing factor for the high vitreous HGF concentrations in diabetic patients, a cytokine that seems to be directly involved in the pathogenesis of PDR. PMID- 10873985 TI - Stabilisation of diabetic retinopathy following simultaneous pancreas and kidney transplant. AB - BACKGROUND/AIMS: Simultaneous pancreas and kidney transplantation (SPK) has become an important option in selected IDDM patients with end stage renal disease (ESRD). Successful SPK transplants are associated with long term normoglycaemic control and improved quality of life. However, debate still continues on the benefit to patients in terms of stabilisation or amelioration of diabetic retinopathy. The progression of diabetic retinopathy (DR) in a cohort of 20 SPK transplant patients is reported. METHODS: All patients were reviewed postoperatively with corrected visual acuity, slit lamp examination, and fundal biomicroscopy. Preoperative data were collected retrospectively and DR was considered unstable if there had been a drop in Snellen acuity greater than three lines or a need for laser photocoagulation or vitrectomy in the 2 years preoperatively. RESULTS: 20 patients who received SPK transplants between March 1983 and April 1994 were reviewed (mean age 35.1 years; mean duration of IDDM = 24.6 years). 17 patients still had functioning grafts at a mean follow up of 5.1 years. Nine of these patients had unstable DR before transplantation. Of these, 89% (8/9) had stabilised DR following transplantation with only a single case requiring laser photocoagulation. Of the eight patients that had stable DR before transplantation all had stable DR following transplantation. 41% of cases (7/17) required cataract surgery during the follow up period. CONCLUSIONS: Advanced diabetic retinopathy is present in a high proportion of cases managed with SPK transplant as a consequence of the duration of IDDM and the presence of ESRD. More than 90% of cases have stable DR following transplant. PMID- 10873986 TI - Quantitative evaluation of fundus autofluorescence imaged "in vivo" in eyes with retinal disease. AB - AIM: To describe a new method of evaluating the topographic distribution of fundus autofluorescence in eyes with retinal disease. METHODS: Images of fundus autofluorescence were obtained in five patients and 34 normal volunteers using a confocal scanning laser ophthalmoscope (cSLO). To evaluate the topographic distribution of fundus autofluorescence throughout the posterior pole a rectangular box, 10 x 750 pixels, was used as the area of analysis. The box was placed, horizontally, across the macular region. The intensity of fundus autofluorescence of each pixel within the rectangular box was plotted against its degree of eccentricity. Profiles of fundus autofluorescence from patients were compared with those obtained from the age matched control group and with cSLO images. RESULTS: Profiles of fundus autofluorescence appeared to represent the topographic distribution of fundus autofluorescence throughout the posterior pole appreciated in the cSLO images, and allowed rapid identification and quantification of areas of increased or decreased fundus autofluorescence. CONCLUSIONS: Fundus autofluorescence profiles appear to be useful to study the spatial distribution of fundus autofluorescence in eyes with retinal disease. PMID- 10873987 TI - Outcome in refractive accommodative esotropia. AB - AIM: To examine outcome among children with refractive accommodative esotropia. METHODS: Children with accommodative esotropia associated with hyperopia were included in the study. The features studied were ocular alignment, amblyopia, and the response to treatment, binocular single vision, requirement for surgery, and the change in refraction with age. RESULTS: 103 children with refractive accommodative esotropia were identified. Mean follow up was 4.5 years (range 2 9.5 years). 41 children (39.8%) were fully accommodative (no manifest deviation with full hyperopic correction). The remaining 62 children (60.2%) were partially accommodative. At presentation 61.2% of children were amblyopic in one eye decreasing to 15.5% at the most recent examination. Stereopsis was demonstrated in 89.3% of children at the most recent examination. Mean cycloplegic refraction (dioptres, spherical equivalent) remained stable throughout the follow up period. The mean change in refraction per year was 0.005 dioptres (D) in right eyes (95% CL -0. 0098 to 0.02) and 0.001 D in left eyes (95% CL -0.018 to 0.021). No patients were able to discard their glasses and maintain alignment. CONCLUSIONS: Most children with refractive accommodative esotropia have an excellent outcome in terms of visual acuity and binocular single vision. Current management strategies for this condition result in a marked reduction in the prevalence of amblyopia compared with the prevalence at presentation. The degree of hyperopia, however, remains unchanged with poor prospects for discontinuing glasses wear. The possibility that long term full time glasses wear impedes emmetropisation must be considered. It is also conceivable, however, that these children may behave differently with normal and be predestined to remain hyperopic. PMID- 10873988 TI - Expression of vascular endothelial growth factor in uveal melanoma and its correlation with metastasis. AB - AIMS: To evaluate the expression of vascular endothelial growth factor (VEGF) in uveal melanomas and correlate its presence with tumour characteristics and systemic metastasis. METHODS: 47 cases of ciliochoroidal melanoma enucleated between 1983 and 1993 were retrieved from the pathology archives at the University of Western Ontario. Paraffin sections stained with haematoxylin and eosin, periodic acid Schiff, and periodic acid Schiff without haematoxylin after bleaching of melanin were examined. The expression of VEGF protein was examined by an immunoalkaline phosphatase method following antigen retrieval, using an antibody to VEGF and vector red as the chromogen. The intensity of VEGF immunoreactivity was graded on a scale of 0-7 and correlated with tumour cell type, tumour size, presence or absence of necrosis, pigmentation, mitotic activity, microvascular density, and microvascular pattern. RESULTS: VEGF immunoreactivity was present in 44/47 tumours (94%): the intensity was graded as very weak (1-2) in 29/47 (62%) and as weak or greater in 15/47 (32%). VEGF was also found in the ciliary epithelium, smooth muscle of the ciliary body and iris, retinal ganglion cells, inner photoreceptor segments, and the retinal pigment epithelium. Follow up data were available in 43/47 patients (91.5%), with a median follow up time of 10 years. 16/43 (37%) patients developed metastases. VEGF expression in melanoma was linked to the presence of tumour necrosis and the degree of pigmentation but no statistically significant relation with microvascular pattern, tumour size, or microvascular density was found. There was no statistically significant correlation between VEGF expression and metastasis. CONCLUSIONS: Most ciliochoroidal melanomas express VEGF and expression is correlated with the presence of necrosis but not with the occurrence of systemic metastasis or tumour angiogenesis. PMID- 10873989 TI - Radiotherapy for age related macular degeneration causes transient lens transparency changes. AB - AIM: Evaluation of potential side effects of photon radiotherapy on the transparency of the lens. METHODS: The anterior segments of 14 phakic eyes from patients suffering from subfoveal neovascularisation as a result of age related macular degeneration (AMD) were documented by Scheimpflug photography (Topcon SL 45, Kodak Tmax 400) before the start of radiotherapy as well as 6 and 12 months afterwards. All negatives were evaluated by microdensitometry, and peak heights for distinct layers of the lens were used for statistical comparison. External beam radiotherapy (6 MeV photons) consisted of a total dose of 20 Gy, delivered as 10 fractions of 2 Gy. RESULTS: Six and 12 months following irradiation statistical comparison of the ratios in density change of lenses from irradiated versus non-irradiated fellow eyes revealed statistically significant (pretinal ->retinoic acid. The latter retinal-->retinoic acid step is irreversible and eventually marks disposal of this essential nutrient, through cytochrome P450 dependent oxidative steps. Mutant mice deficient in aryl hydrocarbon receptor (AHR) accumulate retinyl palmitate, retinol and retinoic acid. This suggests a direct connection between the AHR and retinoid homeostasis. Retinoids control gene expression through the nuclear retinoic acid receptors (RARs) alpha, beta and gamma and 9-cis-retinoic acid receptors alpha, beta and gamma, which bind with high affinity the natural ligands all-trans-retinoic acid and 9-cis-retinoic acid, respectively. Retinoids are effective chemopreventive agents against skin, head and neck, breast, liver and other forms of cancer. Differentiation therapy of acute promyelocytic leukemia (APL) is based on the ability of retinoic acid to induce differentiation of leukemic promyelocytes. Patients with relapsed, retinoid-resistant APL are now being treated with arsenic oxide, which results in apoptosis of the leukemic cells. Interestingly, induction of differentiation in promyelocytes and consequent remission of APL following retinoid therapy depends on expression of a chimeric PML-RAR alpha fusion protein resulting from a t(15;17) chromosomal translocation. This protein functions as a dominant negative against the function of both PML and RARs and its overexpression is able to recreate the phenotypes of the disease in transgenic mice. The development of new, more effective and less toxic retinoids, alone or in combination with other drugs, may provide additional avenues for cancer chemoprevention and differentiation therapy. PMID- 10874004 TI - The relationship between genetic damage from polycyclic aromatic hydrocarbons in breast tissue and breast cancer. AB - A number of polycyclic aromatic hydrocarbons (PAH) are widespread environmental contaminants that cause mammary cancer experimentally. We investigated whether exposure and susceptibility to PAH, as measured by PAH-DNA adducts in breast tissue, are associated with human breast cancer. We carried out a hospital-based case-control study using immunohistochemical methods to analyze PAH-DNA adducts in tumor and nontumor breast tissue from cases and benign breast tissue from controls. The subjects were white, African-American and Latina women without prior cancer or treatment, including 119 women with breast cancer and 108 with benign breast disease without atypia. PAH-DNA adducts measured in breast tumor tissue of 100 cases and in normal tissue from 105 controls were significantly associated with breast cancer (OR=4.43, 96% CI 1.09-18.01) after controlling for known breast cancer risk factors and current active and passive smoking, and dietary PAH. There was substantial interindividual (17-fold) variability in adducts overall, with 27% of cases and 13% of controls having elevated adducts. The odds ratio for elevated adducts in tumor tissue compared with control tissue was 2.56 (1. 05-6.24), after controlling for potential confounders. Adduct levels in tumor tissue did not vary by stage or tumor size. Among 86 cases with paired tumor and nontumor tissue, adducts levels in these two tissues were highly correlated (r=0.56, P<0.001). However, the corresponding associations between case-control status and adducts measured in nontumor tissue from 90 cases and in normal tissue from 105 controls were positive but not statistically significant. Overall, neither active nor passive smoking, or dietary PAH were significantly associated with PAH-DNA adducts or breast cancer case-control status. These results suggest that genetic damage reflecting individual exposure and susceptibility to PAH may play a role in breast cancer; but more research is needed to determine whether the findings are relevant to causation or progression of breast cancer. PMID- 10874005 TI - Mutagenesis in PMS2- and MSH2-deficient mice indicates differential protection from transversions and frameshifts. AB - DNA mismatch repair (MMR) deficiency leads to an increased mutation frequency and a predisposition to neoplasia. 'Knockout' mice deficient in the MMR proteins Msh2 and Pms2 crossed with mutation detection reporter (supF, lacI and cII) transgenic mice have been used to facilitate a comparison of the changes in mutation frequency and spectra. We find that the mutation frequency was consistently higher in Msh2-deficient mice than Pms2-deficient mice. The lacI target gene, which is highly sensitive to point mutations, demonstrated that both Msh2- and Pms2-deficient mice accumulate transition mutations as the predominant mutation. However, when compared with Msh2(-/-) mice, lacI and cII mutants from Pms2 deficient mice revealed an increased proportion of +/-1 bp frameshift mutations and a corresponding decrease in transversion mutations. The supF target gene, which is sensitive to frameshift mutations, and the cII target gene revealed a strong tendency for -1 bp deletions over +1 bp insertions in Msh2(-/-) compared with Pms2(-/-) mice. These data indicate that Msh2 and Pms2 deficiency have subtle but differing effects on mutation avoidance which may contribute to the differences in tumor spectra observed in the two 'knockout' mouse models. These variances in mutation accumulation may also play a role, in part, in the differences seen in prevalence of MSH2 and PMS2 germline mutations in hereditary non-polyposis colorectal cancer patients. PMID- 10874006 TI - Allelotype analysis of chemically induced squamous cell carcinomas in F(1) hybrids of two inbred mouse strains with different susceptibility to tumor progression. AB - Loss of heterozygosity (LOH) at specific chromosomal loci is generally considered indirect evidence for the presence of putative suppressor genes. Allelotyping of tumors using polymorphic markers distributed throughout the entire genome allows the analysis of specific allelic losses. In the field of chemical carcinogenesis, the outbred SENCAR mouse has been commonly used to analyze the multistage nature of skin tumor development. In the study reported here we generated F(1) hybrids between two inbred strains (SENCARB/Pt and SSIN/Sprd) derived from the SENCAR stock that differ in their susceptibility to tumor progression. We typed 24 7, 12 dimethylbenz[a]anthracene and 12-O-tetradecanoylphorbol-13-acetate-induced squamous cell carcinomas for LOH using 56 microsatellite markers distributed among all autosomal chromosomes. The highest percentage of LOH, 78%, was found on chromosome 7, but there was no preferential loss of one particular allele, indicating that the putative suppressor genes found in this area are not involved in genetic susceptibility. High levels of LOH were also found on chromosomes 16 (39%), 6 (29%), 4 (25%), 9 (25%), 14 (22%), 10 (20%) and 19 (20%), but with no preferential loss of the alleles of one strain. The chromosomal regions with LOH on mouse chromosomes 4, 6, 7, 9, 10, 14, 16 and 19 correspond to regions in the human genome where LOH has been reported and have been suggested to harbor tumor suppressor genes. PMID- 10874008 TI - Expression of Jun family members in human colorectal adenocarcinoma. AB - The products of the Jun family genes, c-Jun, JunB and JunD, are essential components of the activating protein-1 transcription factor complexes that are critically important in the control of cell growth, differentiation and neoplastic transformation. Although increased c-Jun expression has been reported in human colorectal tumors, expression of JunB and JunD in these tumors has not previously been characterized. In the current study, we examined 24 cases of human colorectal adenocarcinoma by western immunoblotting analysis and immunohistochemical staining for the expression of c-Jun, JunB and JunD proteins. Normal-appearing colonic mucosa distant from the tumors in the same colectomy specimens were used as a reference for comparison. The results showed that both c Jun and JunB proteins were undetectable or barely detectable in normal mucosa but their expression levels were significantly increased in human colorectal adenocarcinomas. In contrast, JunD protein was present at high levels in normal mucosa and only showed a minimal increase in adenocarcinomas. These observations suggest that different Jun proteins may serve different roles in regulating colonic epithelial cell growth and in colorectal tumorigenesis. PMID- 10874007 TI - Phorbol ester-induced production of cytostatic factors by normal and oncogenic Ha ras-transformed human breast cell lines. AB - The effects of 12-O-tetradecanoylphorbol-13-acetate (TPA) on cell cycle progression were examined in the human breast cell line MCF10A-Neo and a derivative line which expresses a Ha-ras oncogene (MCF10A-NeoT cells). Exposure of MCF10A-Neo cultures to TPA induced a G(1) arrest that lasted approximately 16 24 h (IC(50) approximately 0.5 nM). TPA-treated cultures produced a cytostatic conditioned medium. Cytostatic activity was detectable within 1 h of TPA treatment, peaked 3-7 h later and disappeared between 16 and 24 h post-treatment. However, cytostatic conditioned medium could be quickly regenerated by re-feeding previously treated cultures with new medium. Removal of latent transforming growth factor beta (TGF beta) from the culture medium, supplementing the culture medium with anti-TGFbeta or soluble TGF beta(II) receptor, or pre-absorption of conditioned medium with anti-TGF beta all reduced the cytostatic effects of TPA or conditioned medium on MCF10A-Neo proliferation by approximately 50%. Co treatment with the serine protease inhibitors aprotinin or plasminogen activator inhibitor-1 also suppressed the cytostatic activity of TPA approximately 50%. Conditioned medium isolated from TPA-treated MCF10A-Neo cultures was transiently cytostatic to MCF10A-NeoT cells. The proliferation of MCF10A-NeoT cultures, in contrast to MCF10A-Neo cells, was suppressed at least 72 h following TPA exposure. Conditioned medium isolated from TPA-treated MCF10A-NeoT cultures also suppressed MCF10A-NeoT proliferation for approximately 72 h, but suppressed MCF10A-Neo proliferation for <24 h. These studies suggest that TPA quickly activates proteolytic processes in MCF10A-Neo cells leading to the activation of latent TGF beta supplied by the serum in the culture medium. TPA also stimulates the production of an additional cytostatic factor(s) which signals via a mechanism not involving the TGF beta(II) receptor. Lastly, expression of an activated Ha-ras oncogene alters both the types of cytostatic factors produced following TPA treatment and responsiveness to these factors. PMID- 10874009 TI - Altered expression of beta-catenin, inducible nitric oxide synthase and cyclooxygenase-2 in azoxymethane-induced rat colon carcinogenesis. AB - Activation of the beta-catenin/T cell factor-mediated transcription pathway through mutations of the APC or beta-catenin gene is suggested to play an important role in colon carcinogenesis and there is great interest in the target genes. We have described the frequent mutation and an altered cellular localization of beta-catenin in rat colon adenocarcinomas induced by azoxymethane (AOM), along with up-regulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2. In the present study, the relation between beta-catenin alteration and expression of iNOS and COX-2 in AOM-induced rat colon carcinogenesis was examined in hyperplastic and dysplastic type aberrant crypt, adenoma and adenocarcinoma samples. K-ras gene mutations were also investigated. Mutation analysis by the PCR-single strand conformation polymorphism method and direct sequencing demonstrated the beta-catenin gene to be mutated in two of three dysplastic aberrant crypt foci (ACF), two of six adenomas and 20 of 26 adenocarcinomas, while K-ras was mutated in seven of 10 hyperplastic ACF and seven of 26 adenocarcinomas. Immunohistochemical staining showed an alteration in cellular localization of beta-catenin in all dysplastic ACF, adenomas and adenocarcinomas examined. iNOS expression was also observed in all but one of the lesions in which beta-catenin alterations were observed. Neither iNOS expression nor beta-catenin alterations were observed in any hyperplastic ACF. COX-2 expression in stromal elements was found even in normal colon mucosa and increased in adenomas and adenocarcinomas, while epithelial cells were only positive in large adenocarcinomas. These results show that beta-catenin alterations may be related to induction of iNOS expression, these being early events in AOM-induced colon tumorigenesis which may play important roles in causing dysplastic changes. PMID- 10874010 TI - Requirement for human AP endonuclease 1 for repair of 3'-blocking damage at DNA single-strand breaks induced by reactive oxygen species. AB - The major mammalian apurinic/apyrimidinic (AP) endonuclease (APE1) plays a central role in the DNA base excision repair pathway (BER) in two distinct ways. As an AP endonuclease, it initiates repair of AP sites in DNA produced either spontaneously or after removal of uracil and alkylated bases in DNA by monofunctional DNA glycosylases. Alternatively, by acting as a 3' phosphoesterase, it initiates repair of DNA strand breaks with 3'-blocking damage, which are produced either directly by reactive oxygen species (ROS) or indirectly through the AP lyase reaction of damage-specific DNA glycosylases. The endonuclease activity of APE1, however, is much more efficient than its DNA 3' phosphoesterase activity. Using whole extracts from human HeLa and lymphoblastoid TK6 cells, we have investigated whether these two activities differentially affect BER efficiency. The repair of ROS-induced DNA strand breaks was significantly stimulated by supplementing the reaction with purified APE1. This enhancement was linearly dependent on the amount of APE1 added, while addition of other BER enzymes, such as DNA ligase I and FEN1, had no effect. Moreover, depletion of endogenous APE1 from the extract significantly reduced the repair activity, suggesting that APE1 is essential for repairing such DNA damage and is limiting in extracts of human cells. In contrast, when uracil-containing DNA was used as the substrate, the efficiency of repair was not affected by exogenous APE1, presumably because the AP endonuclease activity was not limiting. These results indicate that the cellular level of APE1 may differentially affect repair efficiency for DNA strand breaks but not for uracil and AP sites in DNA. PMID- 10874011 TI - Comparison between smoking-related DNA adduct analysis in induced sputum and peripheral blood lymphocytes. AB - We investigated the applicability of induced sputum (IS), a non-invasive derivative from the lower respiratory tract, for smoking-related DNA adduct analysis and its comparability with peripheral blood lymphocytes (PBL). Lipophilic DNA adducts were quantified by the (32)P-post-labeling assay in IS and PBL of smokers (n = 9) with stable smoking status at three time points (one week intervals) and non-smokers (n = 9) at one time point. The success rate for sputum induction was 100% at all time points. There was no significant difference in total cell count, cell viability, squamous cell count and DNA yield between smokers and non-smokers. Within the smokers, there was no significant difference in IS cytology at the three time points: overall (mean of three measurements) total cell count, 9.0 +/- 2.4 x 10(6); cell viability, 77 +/- 4%; squamous cell count, 28 +/- 5%; non-squamous cell count, 72 +/- 4% (bronchoalveolar macrophages, 75 +/- 6%; neutrophils, 17 +/- 3%; bronchoepithelial cells, 7 +/- 2%; lymphocytes, 0.7 +/- 0.2%; metachromatic cells, 0.3 +/- 0.2%). IS DNA yield did not differ significantly at the three time points [overall (mean of three extractions) DNA yield, 66 +/- 20 microg]. A typical smoking-associated diagonal radioactive zone was observed in the adduct maps of IS and PBL of all and five smokers, respectively, and of none of the non-smokers. Lipophilic DNA adduct levels in both IS and PBL of smokers were higher than those of non-smokers (3.7 +/- 0. 9 versus 0.7 +/- 0.2/10(8) nt, P = 0.0005, and 2.1 +/- 0.3 versus 0. 6 +/- 0.1/10(8) nt, P = 0.0001, respectively). In smokers the level of adducts in IS was non-significantly higher than that in PBL (3.7 +/- 0.9 versus 2.1 +/- 0.3/10(8) nt, P = 0.1), whilst in non-smokers the difference was not appreciable (0.7 +/- 0.2 versus 0.6 +/- 0. 1/10(8) nt). Within the smokers there was no significant change in the level of adducts at the three time points either in IS or in PBL (coefficients of variation 34 and 29%, respectively). Adduct levels in IS at each time point were higher than those in PBL, leading to a significantly higher overall (mean of three quantifications) level of adducts in IS than PBL (3.3 +/- 0.2 versus 2.1 +/- 0.1/10(8) nt, P = 0.02). The overall levels of adducts in both IS and PBL were dose-dependently related to smoking indices. We conclude that IS is a preferable matrix as compared with PBL for molecular dosimetry of (current) exposure to inhalatory carcinogens as its analysis reveals both the existence and the magnitude of exposure more explicitly. PMID- 10874012 TI - Conceptually new deltanoids (vitamin D analogs) inhibit multistage skin tumorigenesis. AB - Development of vitamin D analogs (deltanoids) as chemopreventive agents requires separation of desirable antiproliferative and pro-differentiating activities from the undesirable calcemic activity also found in the hormone calcitriol (1 alpha, 25-dihydroxyvitamin D(3)). Therefore, several conceptually new deltanoids were synthesized with modifications to the 1alpha- and/or 25-hydroxyl groups, positions traditionally considered essential for stimulating biological responses. In this study, 1 beta-hydroxymethyl-3-epi-25-hydroxyvitamin D(3), a non-calcemic CH(2) homolog of the natural hormone with antiproliferative activity in vitro, was ineffective as an inhibitor of 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced induction of ornithine decarboxylase activity in mouse epidermis. However, a hybrid analog incorporating not only the calcemia-ablating 1 beta hydroxymethyl alteration, but potentiating C,D ring 16-unsaturation and side chain 24,24-fluorination and 26, 27-homologation was found to be as effective as calcitriol. Several non-calcemic 24- or 25-t-butyl sulfones, some containing side chain fluorination but all lacking the 25-hydroxyl group, were also shown to be active in this assay. Three sulfones and the 1 beta-hydroxymethyl hybrid were evaluated as inhibitors of multistage carcinogenesis in mouse skin. Female CD-1 mice were initiated with a single dose of 7,12-dimethylbenz[a]anthracene and then promoted twice weekly for 20 weeks with TPA. Deltanoids were applied topically 30 min before TPA. Unlike calcitriol, none of the atypical deltanoids affected body weight gain in these animals. Minimal effects on urinary calcium excretion were observed following chronic treatment with these analogs. All deltanoids inhibited the incidence and multiplicity of papilloma formation, with the hybrid analog showing the greatest efficacy. With this deltanoid, tumor incidence was significantly reduced by 28% and tumor multiplicity by 63%. These results, coupled with the rich chemical diversity available in side chain sulfur containing deltanoids, particularly when combined with A ring modifications such as 1 beta-hydroxylalkyl groups, provide important new advances in the fundamental understanding of chemical structure-biological activity relationships as well as more potent and safe vitamin D analogs for cancer chemoprevention and other medicinal uses. PMID- 10874013 TI - In vitro bioactivation of N-hydroxy-2-amino-alpha-carboline. AB - 2-Amino-alpha-carboline (A alpha C) is a mutagenic and carcinogenic heterocyclic amine present in foods cooked at high temperature and in cigarette smoke. The mutagenic activity of A alpha C is dependent upon metabolic activation to N hydroxy-A alpha C (N-OH-A alpha C); however, the metabolism of N-OH-A alpha C has not been studied. We have synthesized 2-nitro-alpha-carboline and N-OH-A alpha C and have examined in vitro bioactivation of N-OH-A alpha C by human and rodent liver cytosolic sulfotransferase(s) and acetyltransferase(s) and by recombinant human N-acetyltransferases, NAT1 and NAT2. The sulfotransferase-dependent bioactivation of N-OH-A alpha C by human liver cytosol exhibited large inter individual variation (0.5-75, n = 14) and was significantly higher than bioactivation of N-hydroxy-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH PhIP). Correlation and inhibition studies suggested that the isoform of sulfotransferase primarily responsible for bioactivation of N-OH-A alpha C in human liver cytosol is SULT1A1. O-Acetyltransferase-dependent bioactivation of N OH-A alpha C by human liver cytosol also exhibited large inter-individual variation (16-192, n = 18). In contrast to other N-hydroxy heterocyclic amines, which are primarily substrates only for NAT2, both NAT1 and NAT2 catalyzed bioactivation of N-OH-A alpha C. The rate of bioactivation of N-OH-A alpha C by both NAT1 and NAT2 was significantly higher than that for N-OH-PhIP. In rat and mouse liver cytosols, the level of sulfotransferase-dependent bioactivation of N OH-A alpha C was similar to the level in the high sulfotransferase activity human liver cytosol. The level of O-acetyltransferase-dependent bioactivation of N-OH-A alpha C in rat liver cytosol was also comparable with that in the high acetyltransferase activity human liver cytosol. However, the level of O acetyltransferase-dependent bioactivation of N-OH-A alpha C in mouse liver cytosol was comparable with that in the low acetyltransferase activity human liver cytosol. In contrast to N-OH-PhIP, bioactivation of N-OH-A alpha C was not inhibited by glutathione S-transferase activity; however, DNA binding of N acetoxy-A alpha C was inhibited 20% in the presence of GSH. These results suggest that bioactivation of N-OH-A alpha C may be a significant source of DNA damage in human tissues after dietary exposure to AalphaC and that the relative contribution of each pathway to bioactivation or detoxification of N-OH-A alpha C differs significantly from other N-hydroxy heterocyclic or aromatic amines. PMID- 10874014 TI - Proliferative lesions and reproductive tract tumors in male descendants of mice exposed developmentally to diethylstilbestrol. AB - Prenatal exposure to diethylstilbestrol (DES) is associated with reproductive tract abnormalities, subfertility and neoplasia in experimental animals and humans. Studies using experimental animals suggest that the carcinogenic effects of DES may be transmitted to succeeding generations. To further evaluate this possibility and to determine if there is a sensitive window of exposure, outbred CD-1 mice were treated with DES during three developmental stages: group 1 was treated on days 9-16 of gestation (2.5, 5 or 10 microg/kg maternal body weight) during major organogenesis; group II was treated once on day 18 of gestation (1000 microg/kg maternal body weight) just prior to birth; and group III was treated on days 1-5 of neonatal life (0.002 microg/pup/day). DES-exposed female mice (F(1)) were raised to maturity and bred to control males to generate DES lineage (F(2)) descendants. The F(2) males obtained from these matings are the subjects of this report; results in F(2) females have been reported previously [Newbold et al. (1998) CARCINOGENESIS:, 19, 1655-1663]. Reproductive performance of F(2) males when bred to control females was not different from control males. However, in DES F(2) males killed at 17-24 months, an increased incidence of proliferative lesions of the rete testis and tumors of the reproductive tract was observed. Since these increases were seen in all DES treatment groups, all exposure periods were considered susceptible to perturbation by DES. These data suggest that, while fertility of the DES F(2) mice appeared unaltered, increased susceptibility for tumors is transmitted from the DES 'grandmothers' to subsequent generations. PMID- 10874015 TI - Areca nut extract up-regulates prostaglandin production, cyclooxygenase-2 mRNA and protein expression of human oral keratinocytes. AB - There are about 600 million betel quid (BQ) chewers in the world. BQ chewing is associated with increased incidence of oral cancer and submucous fibrosis. In this study, areca nut (AN) extract (200-800 microg/ml) induced the prostaglandin E(2) (PGE(2)) production by 1. 4-3.4-fold and 6-keto-PGF(1 alpha) production by 1.1-1.7-fold of gingival keratinocytes (GK), respectively, following 24 h of exposure. Exposure of GK to AN extract (>400 microg/ml) led to cell retraction and intracellular vacuoles formation. At concentrations of 800 and 1200 microg/ml, AN extract induced cell death at 21-24 and 32-52% as detected by MTT assay and cellular lactate dehydrogenase release, respectively. Interestingly, AN induced morphological changes of GK are reversible. GK can still proliferate following exposure to AN extract. Cytotoxicity of AN extract cannot be inhibited by indomethacin (1 microM) and aspirin (50 microM), indicating that prostaglandin (PG) production is not the major factor responsible for AN cytotoxicity. PGE(2) exhibited little effect on the growth of GK at concentrations ranging from 100 1000 pg/ml. Stimulating GK production of PGs by AN extract could be due to induction of cyclooxygenase-2 (COX-2) mRNA expression and protein production. These results suggest that AN ingredients are critical in the pathogenesis of oral submucous fibrosis and oral cancer via their stimulatory effects on the PGs, COX-2 production and associated tissue inflammatory responses. AN cytotoxicity to GK is not directly mediated by COX-2 stimulation and PG production. PMID- 10874016 TI - Cyclooxygenase-2 expression is abundant in alveolar type II cells in lung cancer sensitive mouse strains and in premalignant lesions. AB - Overexpression of cyclooxygenase-2 (COX-2) is seen in a high percentage of human colon tumors, lung adenocarcinomas and other cancers. Inhibition of this enzyme represses human colon tumorigenesis and decreases lung tumor multiplicity in 4 (methylnitrosamino)-1-(3-pyridyl)-1-butanone-exposed A/J mice. The purpose of this investigation was to characterize the expression of cyclooxygenase-2 (COX-2) during tumor progression in the A/J mouse lung and to compare the results with expression in other cancer-susceptible and several cancer-resistant mouse strains. Analysis of normal A/J mouse lung showed that type II alveolar epithelial cells express high levels of COX-2 protein and mRNA, indicating that COX-2 is present constitutively in this tumor progenitor cell prior to any carcinogen exposure. Examination of lung-cancer-resistant (C3H/HeJ, C57BL/6J, DBA/2J) and other lung-cancer-susceptible (A/WySnJ, SWR/J) strains showed similar levels of COX-2 mRNA expression in the three susceptible strains and lower levels of expression in two of the resistant strains, indicating a possible correlation between COX-2 expression in type II cells and lung cancer susceptibility. COX-2 protein expression was observed in A/J lung tumors at all stages of development. Variation and occasional absence of protein expression were also observed in A/J lung tumors, particularly in adenomas and adenocarcinomas, suggesting that COX-2 is not obligatory for maintenance of the malignant phenotype. In support of this conclusion, treatment of xenografted cell lines derived from malignant murine pulmonary tumors with COX-2 inhibitors produced only a slight repression of growth. However, the frequent expression of COX-2 in early lesions in the A/J mouse lung combined with the known reduction in tumor number in animals treated with COX-2 inhibitors prior to carcinogen exposure indicate that COX-2 could be a promising target for lung cancer chemoprevention. In addition, high levels of COX 2 expression in the normal tumor-progenitor cells of lung-cancer-sensitive mice indicate that COX-2 may play a role in lung cancer susceptibility. PMID- 10874017 TI - Hematopoietic neoplasia in C57BL/6 mice exposed to split-dose ionizing radiation and circularly polarized 60 Hz magnetic fields. AB - This study assessed the effect of chronic exposure to a 60 Hz circularly polarized magnetic field on the occurrence of ionizing radiation-induced lymphoma and other hematopoietic neoplasia in mice. Female C57BL/6 mice received lifetime exposure to either a magnetic field flux density of 1.42 mT for 18 h/day, or an ambient magnetic field of 0.13 microT. Beginning on the first day of magnetic field exposure, 1710 mice were treated with one of three levels of split-dose Cobalt-60 gamma-radiation (cumulative 3.0, 4.0 or 5.1 Gy). The remaining 570 mice received sham irradiation treatment. Sections from 10 lymphoid tissues were evaluated histopathologically for hematopoietic neoplasia. The primary statistical analysis used the Poly3 method to compare lymphoma incidences in magnetic field (MF)-exposed and control mice. Secondary analyses used the Cox proportional hazards model to analyze incidence rates for mortality and development of specific types of neoplasia. The mortality incidence rate was increased by ionizing radiation treatment, and all neoplasms were observed sooner in irradiated mice. However, the lifetime incidence of hematopoietic neoplasia was similar in all experimental groups, including those that were not exposed to ionizing radiation. Chronic exposure to MFs did not affect the mortality incidence rates and did not change the relative incidences of hematopoietic neoplasia in mice that received the same ionizing radiation treatment, with the exception of a marginally significant reduced relative risk of 0.97 (P = 0.05) for lymphoblastic lymphoma in mice exposed to a magnetic field and treated with 5.1 Gy. Lymphomas and histiocytic sarcomas were first observed approximately 50 days sooner in mice that were exposed to magnetic fields but not ionizing radiation, although this comparison was not statistically significant and the incidence of hematopoietic neoplasia in these mice was not different from that of mice in the 0 T/0 Gy group. PMID- 10874018 TI - Transgenic rats carrying copies of the human c-Ha-ras proto-oncogene exhibit enhanced susceptibility to N-butyl-N-(4-hydroxybutyl)nitrosamine bladder carcinogenesis. AB - We have established a transgenic rat line carrying three copies of the human c-Ha ras proto-oncogene with its own original promoter region, Jcl/SD TgN(HrasGen)128Ncc (Hras128) rat. c-Ha-ras protein from expression of transduced and endogenous c-Ha-ras genes could be detected in the bladder epithelium of untreated transgenic rats. To examine their susceptibility to N-butyl-N-(4 hydroxybutyl)nitrosamine (BBN)-induced urinary bladder carcinogenesis, male transgenic and wild-type littermates were treated with 0.05% BBN in their drinking water for 10 weeks and then killed at week 20. The numbers and volumes of total macroscopic bladder tumors including both transitional cell papillomas and carcinomas (TCC) per rat were much greater in Hras128 rats than in their wild type counterparts. The numbers of carcinomas per rat were also significantly greater in Hras128 rats. Two cases of TCC exhibiting invasion of the bladder muscle layer, which is extremely rare in the wild-type animals under the experimental conditions used, were also observed in Hras128 rats. The GGC-->GAC mutations at codon 12 of the transgene were observed in only two TCC out of 21 bladder tumors (9.5%), assessed by RFLP analysis and direct sequencing. SSCP analysis did not show any endogenous c-Ha-ras gene mutations. One of 25 tumors (4.0%) in wild-type rats had an endogenous c-Ha-ras gene mutation at codon 12 that was detected (GGA-->GAA) by single-strand conformation polymorphism and direct sequencing. These results indicate that the Hras128 rat is highly susceptible to BBN carcinogenesis and may be utilized as a rat model for analysis of bladder tumor development. The mutation findings indicate that the enhanced tumor development is not primarily due to mutations occurring in the transgene. PMID- 10874019 TI - Enhanced in vivo repair of O(4)-methylthymine by a mutant human DNA alkyltransferase. AB - The repair of O(6)-methylguanine (m(6)G) by human O(6)-alkylguanine-DNA alkyltransferase (hAGT) is approximately 5000-fold greater than that for O(4) methylthymine (m(4)T). To evaluate each adduct's contribution to mutagenesis, we previously created a mutant hAGT with increased specificity for m(4)T in vitro. The mutant and wild-type (WT) hAGT have now been expressed in bacterial strains that allow for the specific detection of A:T-->G:C and G:C-->A:T mutations induced by m(4)T and m(6)G, respectively. After exposure to the mutagenic methylating agent, N-methyl-N'-nitro-N-nitrosoguanidine, A:T-->G:C substitutions were reduced >4-fold in cells expressing the mutant hAGT compared with 1. 1-fold for WT hAGT. G:C-->A:T substitutions were decreased >2.5-fold in cells expressing the mutant hAGT, whereas WT hAGT totally prevented G:C-->A:T mutations. These results demonstrate that the altered substrate specificity of hAGT observed in vitro also occurs in vivo, and that it is responsible for the observed differences in mutations. PMID- 10874020 TI - The role of cyclooxygenase enzymes in the growth of human gall bladder cancer cells. AB - Information suggests that the cyclooxygenase (COX) metabolites, the prostanoids, play a role in gall bladder physiology and disease. Non-steroidal anti inflammatory drugs which inhibit COX enzymes have been shown in vivo and in vitro to alter the growth patterns of intestinal epithelial cells, and specific COX-2 inhibitors have been shown to decrease mitogenesis in intestinal epithelial cells. The present study was intended to evaluate the effect of specific COX inhibitors on the growth patterns of gall bladder cancer cells. Employing a human gall bladder cancer cell line, mitogenesis, apoptosis and prostaglandin E(2) (PGE(2)) formation were evaluated in response to serum and hepatocyte growth factor and transforming growth factor alpha stimulation in the presence and absence of specific COX-1 and -2 inhibitors. The effect of the mitogens on COX enzyme expression was also evaluated. Serum and the growth factors increased COX enzyme expression and mitogenesis, and decreased apoptosis as evaluated by the percentage of cells that were floating in culture media rather than attached. There was more DNA degradation in floating than in attached cells. The specific COX-2 inhibitor, but not the COX-1 inhibitor, decreased mitogenesis and increased gall bladder cell apoptosis as evaluated by the number of floating versus attached cells and the number of floating cells in the terminal phase of apoptosis or dead. The inhibition of mitogenesis and the increased apoptosis produced by the COX-2 inhibitor was associated with decreased PGE(2) production. The inhibition of replication of gall bladder cancer cells and the increase in apoptosis produced by the selective COX-2 inhibitor suggests that the COX enzymes and the prostanoids may play a role in the development of gall bladder cancer and that the COX-2 inhibitors may have a therapeutic role in the prevention of gall bladder neoplasms. PMID- 10874021 TI - Quantitative analysis of tumor initiation in rat liver: role of cell replication and cell death (apoptosis). AB - The formation and development of initiated cells has been studied at the beginning of hepatocarcinogenesis. Rats received the genotoxic carcinogen N nitrosomorpholine (NNM); placental glutathione S-transferase was used as a marker of initiated cells (G+ cells). Single G+ cells appeared within 24 h after NNM; their frequency increased steeply for approximately 2 weeks, then decreased and finally remained constant. G+ foci consisting of >/=2 G+ cells appeared successively after the single cells. Histological determination of DNA replication and apoptosis revealed that: the formation of single G+ cells may not depend on DNA replication of precursor cells; single G+ cells showed considerably lower DNA replication than G- normal hepatocytes; from the 2-cell stage onwards G+ foci displayed enhanced DNA replication and apoptosis. Data from histological sections were transformed into the third dimension by a new stereological method which considers the non-spherical shape of many G+ lesions. Rates of division and death of G+ cells and of formation and growth of G+ foci were estimated by a stochastic model: initially G+ clones appeared at a rate of 12 000 per day and liver until a maximal number of 176 000 (phase I) was reached; thereafter they declined to 134 000 (phase II); they then remained constant (phase III). Estimated division rates of G+ cells decreased from phase I to phase III, while the death rate increased in phase II, when every third G+ clone disappeared. As a result, at day 50 after NNM only 0.3% of G+ single cells had formed a clone containing >/=5 cells. In conclusion, experimental and computed parameters provide direct evidence that hepatocarcinogenesis evolves clonally and that initiated hepatocytes have a selective proliferation advantage, associated with an enhanced potential to undergo apoptosis. Thereby, depending on the conditions, initiated clones expand or become extinct. Extinction may lead to reversion of the biological effects of initiation. PMID- 10874022 TI - Roles of JNK, p38 and ERK mitogen-activated protein kinases in the growth inhibition and apoptosis induced by cadmium. AB - Cadmium (Cd), a human carcinogen, can induce apoptosis in various cell types. Three major mitogen-activated protein kinases (MAPKs), c-JUN N-terminal kinase (JNK), p38 and extracellular signal-regulated kinase (ERK), have been shown to regulate apoptosis. In this study we explore the ability of Cd to activate JNK, p38 and ERK, including their effects on Cd-mediated growth inhibition and apoptosis in a human non-small cell lung carcinoma cell line, CL3. The kinase activity of JNK was induced dose-dependently by 30-160 microM CdCl(2). High cytotoxic doses of Cd (130-160 microM) markedly activated p38, but low Cd doses did not. Conversely, the activities of ERK1 and ERK2 were decreased by low cytotoxic doses of Cd (A and C-->T transitions. The identified p53 point mutations occurred at or near (within three codons) of the corresponding hot-spot codons (175, 245, 248 and 273) of human oral SCC. The proportion of group A and group B tumors analyzed exhibiting Ha-ras mutations was 1/15 (7%) and 7/45 (16%), respectively. Only four of the observed eight Ha-ras mutations occurred in codons known to result in activation of this gene. Telomerase activation was demonstrated in 11 of 13 group A tumors (85%) and in 23 of 24 (96%) group B tumors analyzed. The alterations in p53, Ha-ras and telomerase activity observed in this HBP-MBN model are similar in many respects to those observed in the analogous human lesions of the head and neck. This model may be particularly useful for development of cancer chemoprevention regimens and multimodality cancer therapies. PMID- 10874025 TI - APC truncation and increased beta-catenin levels in a human breast cancer cell line. AB - Mutations in the Adenomatous Polyposis Coli (APC) tumor suppressor gene or the beta-catenin gene are present in most colon cancers and less frequently in other tumor types. In this study, we screened 24 human breast cancer cell lines and three immortalized human breast epithelial cell lines for alterations in beta- and gamma-catenin and APC by western blotting, protein truncation assay and DNA sequence analysis. In one cell line (DU 4475), an APC mutation was identified (E1577stop) that resulted in expression of truncated APC. This mutation was associated with elevated cytosolic beta-catenin levels, probably due to loss of APC function, as in colon cancers. No mutations were found in exon 3 of the beta- or gamma-catenin genes. We conclude that APC mutations and beta-catenin upregulation may occur with low frequency in human breast cancer cells. PMID- 10874026 TI - Polymorphisms in the DNA repair genesXRCC1 and ERCC2 and biomarkers of DNA damage in human blood mononuclear cells PMID- 10874027 TI - Dual regulation of platelet protein kinase B. AB - Protein kinase B (PKB) is a serine/threonine kinase that is activated by growth hormones and implicated in prevention of apoptosis, glycogen metabolism, and glucose uptake. A key enzyme in PKB activation is phosphatidylinositide 3-kinase (PI-3K), which triggers the dual phosphorylation of PKB by phosphatidylinositol dependent kinases (PDKs). Here we report that the major PKB subtype in platelets is PKBalpha, which is activated by phosphorylation of Thr(308) and Ser(473) and has a constitutively phosphorylated Thr(450) that does not contribute to PKB activation. alpha-Thrombin and thrombopoietin activate PKBalpha via PI-3K and trigger the concurrent phosphorylation of Thr(308) (via PDK1) and Ser(473) (via a not yet identified PDK2). In addition, alpha-thrombin activates a PI-3K independent pathway involving phospholipase Cbeta and calcium-dependent protein kinase C subtypes (PKCalpha/beta). This route is specific for phosphorylation of Ser(473) and can be initiated by direct PKC activation with phorbol ester or purified active PKC catalytic fragment in platelet lysate. Different degrees of Ser(473) and Thr(308) phosphorylation correlate with different degrees of enzyme activity. These data reveal a PI-3K-independent PKB activation in which PKCalpha/beta regulates the phosphorylation of Ser(473) in PKBalpha. The independent control of the two phosphorylation sites may contribute to fine regulation of PKBalpha activity. PMID- 10874028 TI - Direct functional interactions between insulin-like growth factor-binding protein 3 and retinoid X receptor-alpha regulate transcriptional signaling and apoptosis. AB - Insulin-like growth factor-binding protein (IGFBP)-3 regulates apoptosis in an IGF-independent fashion and has been shown to localize to nuclei. We cloned the nuclear receptor retinoid X receptor-alpha(RXR-alpha) as an IGFBP-3 protein partner in a yeast two-hybrid screen. Multiple methodologies showed that IGFBP-3 and RXR-alpha bind each other within the nucleus. IGFBP-3-induced apoptosis was abolished in RXR-alpha-knockout cells. IGFBP-3 and RXR ligands were additive in inducing apoptosis in prostate cancer cells. IGFBP-3 enhanced RXR response element and inhibited RARE signaling. Thus, RXR-alpha-IGFBP-3 interaction leads to modulation of the transcriptional activity of RXR-alpha and is essential for mediating the effects of IGFBP-3 on apoptosis. PMID- 10874029 TI - Ouabain interaction with cardiac Na+/K+-ATPase initiates signal cascades independent of changes in intracellular Na+ and Ca2+ concentrations. AB - We have shown previously that partial inhibition of the cardiac myocyte Na(+)/K(+)-ATPase activates signal pathways that regulate myocyte growth and growth-related genes and that increases in intracellular Ca(2+) concentration ([Ca(2+)](i)) and reactive oxygen species (ROS) are two essential second messengers within these pathways. The aim of this work was to explore the relation between [Ca(2+)](i) and ROS. When myocytes were in a Ca(2+)-free medium, ouabain caused no change in [Ca(2+)](i), but it increased ROS as it did when the cells were in a Ca(2+)-containing medium. Ouabain-induced increase in ROS also occurred under conditions where there was little or no change in [Na(+)](i). Exposure of myocytes in Ca(2+)-free medium to monensin did not increase ROS. Increase in protein tyrosine phosphorylation, an early event induced by ouabain, was also independent of changes in [Ca(2+)](i) and [Na(+)](i). Ouabain-induced generation of ROS in myocytes was antagonized by genistein, a dominant negative Ras, and myxothiazol/diphenyleneiodonium, indicating a mitochondrial origin for the Ras-dependent ROS generation. These findings, along with our previous data, indicate that increases in [Ca(2+)](i) and ROS in cardiac myocytes are induced by two parallel pathways initiated at the plasma membrane: One being the ouabain altered transient interactions of a fraction of the Na(+)/K(+)-ATPase with neighboring proteins (Src, growth factor receptors, adaptor proteins, and Ras) leading to ROS generation, and the other, inhibition of the transport function of another fraction of the Na(+)/K(+)-ATPase leading to rise in [Ca(2+)](i). Evidently, the gene regulatory effects of ouabain in cardiac myocytes require the downstream collaborations of ROS and [Ca(2+)](i). PMID- 10874030 TI - Involvement of Src and epidermal growth factor receptor in the signal-transducing function of Na+/K+-ATPase. AB - Nontoxic concentrations of ouabain, causing partial inhibition of the cardiac myocyte Na(+)/K(+)-ATPase, induce hypertrophy and several growth-related genes through signal pathways that include the activation of Ras and p42/44 mitogen activated protein kinase (MAPK). The aim of this work was to examine the ouabain induced events upstream of the Ras/MAPK cascade. Treatment of myocytes with genistein antagonized ouabain-induced activation of the MAPK, suggesting that protein tyrosine phosphorylation has a role. Tyrosine phosphorylation of several myocyte proteins was increased rapidly upon cell exposure to ouabain. Lowering of extracellular K(+) had a similar ouabain-like effect. Ouabain also increased protein tyrosine phosphorylation in A7r5, HeLa, and L929 cells. In cardiac myocytes and A7r5 cells, herbimycin A antagonized the ouabain-induced increase in protein tyrosine phosphorylation and MAPK activation. In both cell types, ouabain stimulated Src kinase activity, Src translocation to the Triton-insoluble fraction, Src association with the epidermal growth factor receptor, and the tyrosine phosphorylation of this receptor on site(s) other than its major autophosphorylation site, Tyr(1173). The findings suggest that (a) the ouabain induced activation of Src and the Src-induced phosphorylation of the growth factor receptor provide the scaffolding for the recruitment of adaptor proteins and Ras and the activation of Ras/MAPK cascade; and (b) the activation of such pathways may be a common feature of the signal-transducing function of Na(+)/K(+) ATPase in most cells. PMID- 10874031 TI - Inhibition of neuronal nitric-oxide synthase by calcium/ calmodulin-dependent protein kinase IIalpha through Ser847 phosphorylation in NG108-15 neuronal cells. AB - We have previously demonstrated that phosphorylation of neuronal nitric-oxide synthase (nNOS) at Ser(847) by Ca(2+)/calmodulin-dependent protein kinases (CaM kinases) attenuates the catalytic activity of the enzyme in vitro (Hayashi Y., Nishio M., Naito Y., Yokokura H., Nimura Y., Hidaka H., and Watanabe Y. (1999) J. Biol. Chem. 274, 20597-20602). In the present study we determined that CaM kinase IIalpha (CaM-K IIalpha) can directly phosphorylate nNOS on Ser(847), leading to a reduction of nNOS activity in cells. The phosphorylation abilities of purified CaM kinase Ialpha (CaM-K Ialpha), CaM-K IIalpha, and CaM-kinase IV (CaM-K IV) on Ser(847) were analyzed using the synthetic peptide nNOS-(836-859) (Glu-Glu-Arg Lys-Ser-Tyr-Lys-Val-Arg-Phe-Asn-Ser-Val-Ser-Ser-Tyr-Ser- Asp-Ser-Arg-Lys-Ser-Ser Gly) from nNOS as substrate. The relative V(max)/K(m) ratios of CaM kinases for nNOS-(836-859) were found to be as follows: CaM-K IIalpha, 100; CaM-K Ialpha, 54.5; CaM-K IV, 9.1. Co-transfection of constitutively active CaM-K IIalpha1-274 but not inactive CaM-K IIalpha1-274, generated by mutation of Lys(42) to Ala, with nNOS into NG108-15 cells, resulted in increased Ser(847) phosphorylation in the presence of okadaic acid, an inhibitor of protein phosphatase (PP)1 and PP2A, with a concomitant inhibition of NOS enzyme activity. In addition, this latter decrease could be reversed by treatment with exogenous PP2A. Cells expressing mutant nNOS (S847A) proved resistant to phosphorylation and a decrease of NOS activity. Thus, our results indicate that Ca(2+) triggers cross-talk signal transduction between CaM kinase and NO and CaM-K IIalpha phosphorylating nNOS on Ser(847), which in turn decreases the gaseous second messenger NO in neuronal cells. PMID- 10874032 TI - Cryptic dimer interface and domain organization of the extracellular region of metabotropic glutamate receptor subtype 1. AB - Previously, we produced the whole extracellular region of metabotropic glutamate receptor subtype 1 (mGluR1) in a soluble form. The soluble receptor retained a ligand affinity comparable with that of the full-length membrane-bound receptor and formed a disulfide-linked dimer. Here, we have identified a cysteine residue responsible for the intermolecular disulfide bond and determined domain organization of the extracellular region of mGluR1. A mutant, C140A, was a monomer under nonreduced conditions by SDS-polyacrylamide gel electrophoresis; however, C140A was eluted at the position similar to that of mGluR113, the wild type soluble receptor, by size exclusion column chromatography. Furthermore, C140A bound a ligand, [(3)H]quisqualate, with an affinity similar to that obtained by mGluR113. Oocytes injected with RNA for full-length mGluR1 containing C140A mutation showed responses to ligands at magnitudes similar to those with wild type full-length RNA. Thus, elimination of the disulfide linkage did not perturb the dimer formation and ligand signaling, suggesting that cryptic dimer interface(s) possibly exist in mGluR1. Limited proteolysis of the whole extracellular fragment (residue 33-592) revealed two trypsin-sensitive sites, after the residues Arg(139) and Arg(521). A 15-kDa NH(2)-terminal proteolytic fragment (residue 33-139) was associated with the downstream part after the digestion. Arg(521) was located before a cysteine-rich stretch preceding the transmembrane region. A new shorter soluble receptor (residue 33-522) lacking the cysteine-rich region was designed based on the protease-sensitive boundary. The purified receptor protein gave a K(d) value of 58.1 +/- 0.84 nm, which is compatible to a reported value of the full-length receptor. The B(max) value was 7.06 +/- 0. 82 nmol/mg of protein. These results indicated that the ligand binding specificity of mGluR1 is confined to the NH(2)-terminal 490-amino acid region of the mature protein. PMID- 10874033 TI - Neelaredoxin, an iron-binding protein from the syphilis spirochete, Treponema pallidum, is a superoxide reductase. AB - Treponema pallidum, the causative agent of venereal syphilis, is a microaerophilic obligate pathogen of humans. As it disseminates hematogenously and invades a wide range of tissues, T. pallidum presumably must tolerate substantial oxidative stress. Analysis of the T. pallidum genome indicates that the syphilis spirochete lacks most of the iron-binding proteins present in many other bacterial pathogens, including the oxidative defense enzymes superoxide dismutase, catalase, and peroxidase, but does possess an orthologue (TP0823) for neelaredoxin, an enzyme of hyperthermophilic and sulfate-reducing anaerobes shown to possess superoxide reductase activity. To analyze the potential role of neelaredoxin in treponemal oxidative defense, we examined the biochemical, spectroscopic, and antioxidant properties of recombinant T. pallidum neelaredoxin. Neelaredoxin was shown to be expressed in T. pallidum by reverse transcriptase-polymerase chain reaction and Western blot analysis. Recombinant neelaredoxin is a 26-kDa alpha(2) homodimer containing, on average, 0.7 iron atoms/subunit. Mossbauer and EPR analysis of the purified protein indicates that the iron atom exists as a mononuclear center in a mixture of high spin ferrous and ferric oxidation states. The fully oxidized form, obtained by the addition of K(3)(Fe(CN)(6)), exhibits an optical spectrum with absorbances at 280, 320, and 656 nm; the last feature is responsible for the protein's blue color, which disappears upon ascorbate reduction. The fully oxidized protein has a A(280)/A(656) ratio of 10.3. Enzymatic studies revealed that T. pallidum neelaredoxin is able to catalyze a redox equilibrium between superoxide and hydrogen peroxide, a result consistent with it being a superoxide reductase. This finding, the first description of a T. pallidum iron-binding protein, indicates that the syphilis spirochete copes with oxidative stress via a primitive mechanism, which, thus far, has not been described in pathogenic bacteria. PMID- 10874034 TI - Regulation of glycogen synthase. Identification of residues involved in regulation by the allosteric ligand glucose-6-P and by phosphorylation. AB - The major yeast glycogen synthase, Gsy2p, is inactivated by phosphorylation and activated by the allosteric ligand glucose-6-P. From studies of recombinant proteins, the control can be accommodated by a three-state model, in which unphosphorylated enzyme has intermediate activity (state II). Glucose-6-P increased V(max)/K(m) by about 2-fold (state III), whereas phosphorylation by the cyclin-dependent protein kinase Pcl10p/Pho85p decreased V(max)/K(m) by approximately 30-fold (state I). In the presence of glucose-6-P, state III is achieved regardless of phosphorylation state. The enzyme forms complexes in solution with the yeast glycogenin Glg2p, but this interaction appears not to affect control either by glucose-6-P binding or by phosphorylation. Scanning mutagenesis was applied to identify residues potentially involved in ligand binding. Of 22 mutant enzymes analyzed, seven were essentially inactive. Five mutant proteins were altered in their activation by glucose-6-P, and two were completely unaffected by the hexose phosphate. One of these, R586A/R588A/R591A (all three of the indicated Arg residues mutated to Ala), had wild-type activity and was normally inactivated by phosphorylation. A second mutant, R579A/R580A/R582A, had somewhat reduced V(max), but its activity was not greatly reduced by phosphorylation. The Arg residues in these two mutants are restricted to a highly conserved, 13-residue segment of Gsy2p that we propose to be important for glucose-6-P binding and/or the ability of the enzyme to undergo transitions between activity states. PMID- 10874035 TI - One of two genes encoding glycyl-tRNA synthetase in Saccharomyces cerevisiae provides mitochondrial and cytoplasmic functions. AB - In the yeast Saccharomyces cerevisiae, two genes (GRS1 and GRS2) encode glycyl tRNA synthetase (GlyRS1 and GlyRS2, respectively). 59% of the sequence of GlyRS2 is identical to that of GlyRS1. Others have proposed that GRS1 and GRS2 encode the cytoplasmic and mitochondrial enzymes, respectively. In this work, we show that GRS1 encodes both functions, whereas GRS2 is dispensable. In addition, both cytoplasmic and mitochondrial phenotypes of the knockout allele of GRS1 in S. cerevisiae are complemented by the expression of the only known gene for glycyl tRNA synthetase in Schizosaccharomyces pombe. Thus, a single gene for glycyl-tRNA synthetase likely encodes both cytoplasmic and mitochondrial activities in most or all yeast. Phylogenetic analysis shows that GlyRS2 is a predecessor of all yeast GlyRS homologues. Thus, GRS1 appears to be the result of a duplication of GRS2, which itself is pseudogene-like. PMID- 10874036 TI - Tumor necrosis factor receptors types 1 and 2 differentially regulate osteoclastogenesis. AB - The potent osteoclastogenic agent, tumor necrosis factor-alpha (TNF), exerts its biological effects via two receptors, namely TNF receptors 1 (p55r) and 2 (p75r), each present on osteoclast precursors. Thus, we asked if p55r and p75r differentially impact the osteoclastogenic process. Marrow derived from mice expressing only p55r generates substantially more osteoclasts, in the basal state, than does wild type, while marrow expressing only p75r, produces substantially fewer. Reflecting its preferential activation of p55r, exogenous TNF stimulates osteoclast formation by p55r(+/+)p75r(-/-), but not p55r(-/ )p75r(+/+), marrow. Consistent with the fact that NF-kappaB is essential for osteoclastogenesis, this transcription complex is activated, relative to wild type, in p55r(+/+)p75r(-/-) osteoclast precursors and suppressed in those expressing only p75r. Because p55r enhances, and p75r suppresses, osteoclastogenesis, we asked if their principal ligands, namely soluble and membrane-residing TNF, respectively, differentially impact basal osteoclast recruitment. We find, in contrast to the significant level of osteoclast formation in wild type marrow, osteoclastogenesis by that derived from mice expressing membrane, but not soluble, TNF, is negligible. Thus, optimal therapeutic inhibition of bone resorption may entail selective TNF receptor modulation and/or arrested cleavage of membrane TNF to its soluble form. PMID- 10874037 TI - Rapid dephosphorylation of H1 histones after apoptosis induction. AB - H1 histones are involved in the formation of higher order chromatin structures and in the modulation of gene expression. Changes in chromatin structure are a characteristic initial feature of apoptosis. We therefore have investigated the histone H1 pattern of the human leukemic cell line HL60 undergoing programmed cell death, as induced by topoisomerase I inhibition. Histone H1 proteins were isolated and analyzed by high performance liquid chromatography and capillary zone electrophoresis. DNA fragmentation after apoptosis induction was monitored by agarose gel electrophoresis. The patterns of the three H1 histone subtypes extractable from apoptotic HL60 cells significantly differed from those of control cells in showing a decrease of phosphorylated H1 subtypes and an increase of the respective dephosphorylated forms. This dephosphorylation of H1 histones could be observed already 45 min after apoptosis induction and preceded internucleosomal DNA cleavage by approximately 2 h. We conclude that during apoptotic DNA fragmentation, the H1 histones become rapidly dephosphorylated by a yet unknown protein phosphatase. PMID- 10874038 TI - CLIC-1 functions as a chloride channel when expressed and purified from bacteria. AB - CLIC-1 is a member of a family of proteins related to the bovine intracellular chloride channel p64 which has been proposed to function as a chloride channel. We expressed CLIC-1 as a glutathione S-transferase fusion protein in bacteria. The fusion protein was purified by glutathione affinity, and CLIC-1 was released from its fusion partner by digestion with thrombin. After further purification, CLIC-1 was reconstituted into phospholipid vesicles by detergent dialysis. Chloride permeability of reconstituted vesicles was assessed using a valinomycin dependent chloride efflux assay, demonstrating increased vesicular chloride permeability with CLIC-1 compared with control. CLIC-1-dependent chloride permeability was inhibited by indanyloxyacetic acid-94 with an apparent IC(50) of 8.6 micrometer. The single channel properties of CLIC-1 were determined using the planar lipid bilayer technique. We found that CLIC-1 forms a voltage-dependent, Cl-selective channel with a rectifying current-voltage relationship and single channel conductances of 161 +/- 7.9 and 67.5 +/- 6.9 picosiemens in symmetric 300 and 150 mm KCl, respectively. The anion selectivity of this activity is Br approximately Cl > I. The open probability of CLIC-1 channels in planar bilayers was decreased by indanyloxyacetic acid-94 with an apparent IC(50) of 86 micrometer at 50 mV. These data convincingly demonstrate that CLIC-1 is capable of forming a novel, chloride-selective channel in the absence of other subunits or proteins. PMID- 10874039 TI - The plastid ribosomal proteins. Identification of all the proteins in the 30 S subunit of an organelle ribosome (chloroplast). AB - Identification of all the protein components of a plastid (chloroplast) ribosomal 30 S subunit has been achieved, using two-dimensional gel electropholesis, high performance liquid chromatography purification, N-terminal sequencing, polymerase chain reaction-based screening of cDNA library, nucleotide sequencing, and mass spectrometry (electrospray ionization, matrix-assisted laser desorption/ionization time-of-flight, and reversed-phase HPLC coupled with electrospray ionization mass spectrometry). 25 proteins were identified, of which 21 are orthologues of all Escherichia coli 30 S ribosomal proteins (S1-S21), and 4 are plastid-specific ribosomal proteins (PSRPs) that have no homologues in the mitochondrial, archaebacterial, or cytosolic ribosomal protein sequences in data bases. 12 of the 25 plastid 30 S ribosomal proteins (PRPs) are encoded in the plastid genome, whereas the remaining 13 are encoded by the nuclear genome. Post translational transit peptide cleavage sites for the maturation of the 13 cytosolically synthesized PRPs, and post-translational N-terminal processing in the maturation of the 12 plastid synthesized PRPs are described. Post translational modifications in several PRPs were observed: alpha-N-acetylation of S9, N-terminal processings leading to five mature forms of S6 and two mature forms of S10, C-terminal and/or internal modifications in S1, S14, S18, and S19, leading to two distinct forms differing in mass and/or charge (the corresponding modifications are not observed in E. coli). The four PSRPs in spinach plastid 30 S ribosomal subunit (PSRP-1, 26.8 kDa, pI 6.2; PSRP-2, 21.7 kDa, pI 5.0; PSRP-3, 13.8 kDa, pI 4.9; PSRP-4, 5.2 kDa, pI 11.8) comprise 16% (67.6 kDa) of the total protein mass of the 30 S subunit (429.3 kDa). PSRP-1 and PSRP-3 show sequence similarities with hypothetical photosynthetic bacterial proteins, indicating their possible origins in photosynthetic bacteria. We propose the hypothesis that PSRPs form a "plastid translational regulatory module" on the 30 S ribosomal subunit structure for the possible mediation of nuclear factors on plastid translation. PMID- 10874040 TI - Evidence that type I, II, and III inositol 1,4,5-trisphosphate receptors can occur as integral plasma membrane proteins. AB - A number of previous reports have suggested that inositol 1,4, 5-trisphosphate receptors (IP(3)Rs) are present in the plasma membranes of cells. We confirm this directly in the present study by demonstrating that a significant proportion of the IP(3)Rs found in A431 cells, Jurkat cells, and rat parotid acini can be biotinylated by the extracellular application of sulfo-N-hydroxysuccinimide biotin to intact cells. This labeling cannot be accounted for by the reaction of sulfo-N-hydroxysuccinimide-biotin with intracellular IP(3)Rs since calnexin and the SERCA2 ATPase, both integral membrane proteins of the endoplasmic reticulum, are not labeled under the same experimental conditions. Individual IP(3)R subtypes were detected using subtype-specific antibodies. A431 cells expressed only the type-3 IP(3)R, and 23% of this protein was in the biotinylated (plasma membrane) fraction. Jurkat cells and rat parotid cells expressed all three IP(3)R subtypes. Contrary to earlier results suggesting that only the type-3 IP(3)R might localize to the plasma membrane, we found that significant amounts (5-14%) of all three subtypes could be identified in the biotinylated fractions of Jurkat and rat parotid cells. Our results suggest a role for IP(3)Rs in plasma membrane as well as intracellular membrane function. PMID- 10874041 TI - Mullerian inhibiting substance inhibits breast cancer cell growth through an NFkappa B-mediated pathway. AB - Mullerian inhibiting substance (MIS), a member of the transforming growth factor beta superfamily, induces regression of the Mullerian duct in male embryos. In this report, we demonstrate MIS type II receptor expression in normal breast tissue and in human breast cancer cell lines, breast fibroadenoma, and ductal adenocarcinomas. MIS inhibited the growth of both estrogen receptor (ER)-positive T47D and ER-negative MDA-MB-231 breast cancer cell lines, suggesting a broader range of target tissues for MIS action. Inhibition of growth was manifested by an increase in the fraction of cells in the G(1) phase of the cell cycle and induction of apoptosis. Treatment of breast cancer cells with MIS activated the NFkappaB pathway and selectively up-regulated the immediate early gene IEX-1S, which, when overexpressed, inhibited breast cancer cell growth. Dominant negative IkappaBalpha expression ablated both MIS-mediated induction of IEX-1S and inhibition of growth, indicating that activation of the NFkappaB signaling pathway was required for these processes. These results identify the NFkappaB mediated signaling pathway and a target gene for MIS action and suggest a putative role for the MIS ligand and its downstream interactors in the treatment of ER-positive as well as negative breast cancers. PMID- 10874042 TI - Characterization of the interaction between protein 4.1R and ZO-2. A possible link between the tight junction and the actin cytoskeleton. AB - Multiple isoforms of the red cell protein 4.1R are expressed in nonerythroid cells, including novel 135-kDa isoforms. Using a yeast two-hybrid system, immunocolocalization, immunoprecipitation, and in vitro binding studies, we found that two 4.1R isoforms of 135 and 150 kDa specifically interact with the protein ZO-2 (zonula occludens-2). 4.1R is colocalized with ZO-2 and occludin at Madin Darby canine kidney (MDCK) cell tight junctions. Both isoforms of 4.1R coprecipitated with proteins that organize tight junctions such as ZO-2, ZO-1, and occludin. Western blot analysis also revealed the presence of actin and alpha spectrin in these immunoprecipitates. Association of 4.1R isoforms with these tight junction and cytoskeletal proteins was found to be specific for the tight junction and was not seen in nonconfluent MDCK cells. The amino acid residues that sustain the interaction between 4.1R and ZO-2 reside within the amino acids encoded by exons 19-21 of 4.1R and residues 1054-1118 of ZO-2. Exogenously expressed 4.1R containing the spectrin/actin- and ZO-2-binding domains was recruited to tight junctions in confluent MDCK cells. Taken together, our results suggest that 4.1R might play an important role in organization and function of the tight junction by establishing a link between the tight junction and the actin cytoskeleton. PMID- 10874043 TI - Inhibitory serpins from wheat grain with reactive centers resembling glutamine rich repeats of prolamin storage proteins. Cloning and characterization of five major molecular forms. AB - Genes encoding proteins of the serpin superfamily are widespread in the plant kingdom, but the properties of very few plant serpins have been studied, and physiological functions have not been elucidated. Six distinct serpins have been identified in grains of hexaploid bread wheat (Triticum aestivum L.) by partial purification and amino acid sequencing. The reactive centers of all but one of the serpins resemble the glutamine-rich repetitive sequences in prolamin storage proteins of wheat grain. Five of the serpins, classified into two protein Z subfamilies, WSZ1 and WSZ2, have been cloned, expressed in Escherichia coli, and purified. Inhibitory specificity toward 17 proteinases of mammalian, plant, and microbial origin was studied. All five serpins were suicide substrate inhibitors of chymotrypsin and cathepsin G. WSZ1a and WSZ1b inhibited at the unusual reactive center P(1)-P(1)' Gln-Gln, and WSZ2b at P(2)-P(1) Leu-Arg-one of two overlapping reactive centers. WSZ1c with P(1)-P(1)' Leu-Gln was the fastest inhibitor of chymotrypsin (k(a) = 1.3 x 10(6) m(-1) s(-1)). WSZ1a was as efficient an inhibitor of chymotrypsin as WSZ2a (k(a) approximately 10(5) m(-1) s(-1)), which has P(1)-P(1)' Leu-Ser-a reactive center common in animal serpins. WSZ2b inhibited plasmin at P(1)-P(1)' Arg-Gln (k(a) approximately 10(3) m(-1) s( 1)). None of the five serpins inhibited Bacillus subtilisin A, Fusarium trypsin, or two subtilisin-like plant serine proteinases, hordolisin from barley green malt and cucumisin D from honeydew melon. Possible functions involving interactions with endogenous or exogenous proteinases adapted to prolamin degradation are discussed. PMID- 10874045 TI - Substituting selenocysteine for catalytic cysteine 41 enhances enzymatic activity of plant phospholipid hydroperoxide glutathione peroxidase expressed in Escherichia coli. AB - The citrus phospholipid hydroperoxide glutathione peroxidase (cit-PHGPx) was the first plant peroxidase demonstrated to exhibit PHGPx-specific enzymatic activity, although it was 500-fold weaker than that of the pig heart analog. This relatively low activity is accounted for the catalytic residue of cit-PHGPx, which was found to be cysteine and not the rare selenocysteine (Sec) present in animal enzymes. Sec incorporation into proteins is encoded by a UGA codon, usually a STOP codon, which, in prokaryotes, is suppressed by an adjacent downstream mRNA stem-loop structure, the Sec insertion sequence (SECIS). By performing appropriate nucleotide substitutions into the gene encoding cit-PHGPx, we introduced bacterial-type SECIS elements that afforded the substitution of the catalytic Cys(41) by Sec, as established by mass spectrometry, while preserving the functional integrity of the peroxidase. The recombinant enzyme, whose synthesis is selenium-dependent, displayed a 4-fold enhanced peroxidase activity as compared with the Cys-containing analog, thus confirming the higher catalytic power of Sec compared with Cys in cit-PHGPx active site. The study led also to refinement of the minimal sequence requirements of the bacterial-type SECIS, and, for the first time, to the heterologous expression in Escherichia coli of a eukaryotic selenoprotein containing a SECIS in its open reading frame. PMID- 10874044 TI - Mechanism of heme oxygenase-1 gene activation by cadmium in MCF-7 mammary epithelial cells. Role of p38 kinase and Nrf2 transcription factor. AB - The mouse heme oxygenase-1 (HO-1) gene, ho-1, contains two inducible enhancers, E1 and E2. Of several cell lines tested, induction of an E1/luciferase fusion construct, pE1-luc, by CdCl(2) is most pronounced in MCF-7 cells. In these cells, E1, but not E2, is necessary and sufficient for ho-1 gene activation. Exposure of MCF-7 cells to 10 micrometer CdCl(2) stimulates phosphorylation of ERK, JNK, and p38 mitogen-activated protein kinases, implicating one or more of these signaling pathways in ho-1 gene induction. SB203580, an inhibitor of p38, diminishes cadmium-stimulated pE1-luc expression and HO-1 mRNA levels by up to 70-80%. PD098059, an ERK pathway inhibitor, does not affect HO-1 mRNA induction at the highest concentration (40 micrometer) tested. Similarly, co-expression of a dominant-negative mutant of p38alpha, but not of ERK1, ERK2, JNK1, or JNK2, reduces basal and cadmium-induced pE1-luc activity. E1 contains binding sites for the activator protein-1 (Fos/Jun), Cap'n'Collar/basic leucine zipper (CNC-bZIP), and CCAAT/enhancer-binding protein (C/EBP) families of transcription factors. A dominant-negative mutant of Nrf2 (a CNC-bZIP member), but not of c-Jun or C/EBPbeta, inhibits pE1-luc activation by cadmium. Induction of the endogenous ho 1 gene is also inhibited by the Nrf2 mutant. Mutations of E1 that inhibit cadmium inducibility also suppress the trans-activation and DNA binding activities of Nrf2, and SB203580, but not PD098059, attenuates Nrf2-mediated trans-activation of pE1-luc. Taken together, these results indicate that cadmium induces ho-1 gene expression via sequential activation of the p38 kinase pathway and Nrf2. PMID- 10874046 TI - The plastid ribosomal proteins. Identification of all the proteins in the 50 S subunit of an organelle ribosome (chloroplast). AB - We have completed identification of all the ribosomal proteins (RPs) in spinach plastid (chloroplast) ribosomal 50 S subunit via a proteomic approach using two dimensional electrophoresis, electroblotting/protein sequencing, high performance liquid chromatography purification, polymerase chain reaction-based screening of cDNA library/nucleotide sequencing, and mass spectrometry (reversed-phase HPLC coupled to electrospray ionization mass spectrometry and electrospray ionization mass spectrometry). Spinach plastid 50 S subunit comprises 33 proteins, of which 31 are orthologues of Escherichia coli RPs and two are plastid-specific RPs (PSRP 5 and PSRP-6) having no homologues in other types of ribosomes. Orthologues of E. coli L25 and L30 are absent in spinach plastid ribosome. 25 of the plastid 50 S RPs are encoded in the nuclear genome and synthesized on cytosolic ribosomes, whereas eight of the plastid RPs are encoded in the plastid organelle genome and synthesized on plastid ribosomes. Sites for transit peptide cleavages in the cytosolic RP precursors and formyl Met processing in the plastid-synthesized RPs were established. Post-translational modifications were observed in several mature plastid RPs, including multiple forms of L10, L18, L31, and PSRP-5 and N terminal/internal modifications in L2, L11 and L16. Comparison of the RPs in gradient-purified 70 S ribosome with those in the 30 and 50 S subunits revealed an additional protein, in approximately stoichiometric amount, specific to the 70 S ribosome. It was identified to be plastid ribosome recycling factor. Combining with our recent study of the proteins in plastid 30 S subunit (Yamaguchi, K., von Knoblauch, K., and Subramanian, A. R. (2000) J. Biol. Chem. 275, 28455-28465), we show that spinach plastid ribosome comprises 59 proteins (33 in 50 S subunit and 25 in 30 S subunit and ribosome recycling factor in 70 S), of which 53 are E. coli orthologues and 6 are plastid-specific proteins (PSRP-1 to PSRP-6). We propose the hypothesis that PSRPs were evolved to perform functions unique to plastid translation and its regulation, including protein targeting/translocation to thylakoid membrane via plastid 50 S subunit. PMID- 10874047 TI - A reaction-induced fourier transform-infrared spectroscopic study of the lactose permease. A transmembrane potential perturbs carboxylic acid residues. AB - In chemiosmotic coupling, a transmembrane ion gradient is used as the source of energy to drive reactions. This process occurs in all cells, but the microscopic mechanism is not understood. Here, Escherichia coli lactose permease was used in a novel spectroscopic method to investigate the mechanism of chemiosmotic coupling in secondary active transporters. To provide a light-triggered electrochemical gradient, bacteriorhodopsin was co-reconstituted with the permease, and reaction-induced Fourier transform-infrared spectra were obtained from the co-reconstituted samples. The bacteriorhodopsin contributions were subtracted from these data to give spectra reflecting permease conformational changes that are induced by an electrochemical gradient. Positive bands in the 1765-1730 cm(-1) region are attributable to carboxylic acid residues in the permease and are consistent with changes of pK(a), protonation state, or environment. This is the first direct information concerning gradient-induced structural changes in the permease at the single amino acid level. Ultimately, these structural changes facilitate galactoside binding and may be involved in the storage of free energy. PMID- 10874048 TI - Interaction and functional cooperation of NF-kappa B with Smads. Transcriptional regulation of the junB promoter. AB - The transforming growth factor-beta (TGF-beta) family of cytokines regulates diverse cellular processes through control of the expression of target genes. Smad proteins are a recently identified family of signal transducers for members of the TGF-beta family. Smads act as transcriptional regulators through binding to DNA and interacting with a variety of transcription factors. Here, we identified a kappaB site as a TGF-beta-responsive region in the 3'-downstream junB promoter region. We also demonstrate that kappaB sites alone are sufficient to mediate immediate transcriptional activation by TGF-beta. Transactivation of kappaB sites by TGF-beta requires an intact NF-kappaB pathway, cooperates with known activators of this pathway, and is mediated by Smad family members. Furthermore, we show that Smad3 interacts with p52 in vivo. These data expand the model in which Smad proteins undergo multiple interactions with several transcription factors that could induce either activation or repression of gene expression. PMID- 10874049 TI - Molecular mechanism and energetics of clamp assembly in Escherichia coli. The role of ATP hydrolysis when gamma complex loads beta on DNA. AB - Escherichia coli DNA polymerase III holoenzyme is a multisubunit composite containing the beta sliding clamp and clamp loading gamma complex. The gamma complex requires ATP to load beta onto DNA. A two-color fluorescence spectroscopic approach was utilized to study this system, wherein both assembly (red fluorescence; X-rhodamine labeled DNA anisotropy assay) and ATP hydrolysis (green fluorescence; phosphate binding protein assay) were simultaneously measured with millisecond timing resolution. The two temporally correlated stopped-flow signals revealed that a preassembled beta. gamma complex composite rapidly binds primer/template DNA in an ATP hydrolysis independent step. Once bound, two molecules of ATP are rapidly hydrolyzed (approximately 34 s(-1)). Following hydrolysis, gamma complex dissociates from the DNA ( approximately 22 s(-1)). Once dissociated, the next cycle of loading is severely compromised, resulting in steady-state ATP hydrolysis rates with a maximum of only approximately 3 s(-1). Two single-site beta dimer interface mutants were examined which had impaired steady-state rates of ATP hydrolysis. The pre-steady-state correlated kinetics of these mutants revealed a pattern essentially identical to wild type. The anisotropy data showed that these mutants decrease the steady state rates of ATP hydrolysis by causing a buildup of "stuck" binary-ternary complexes on the primer/template DNA. PMID- 10874050 TI - Impact of genomics on drug discovery and clinical medicine. AB - Genomics, particularly high-throughput sequencing and characterization of expressed human genes, has created new opportunities for drug discovery. Knowledge of all the human genes and their functions may allow effective preventive measures, and change drug research strategy and drug discovery development processes. Pharmacogenomics is the application of genomic technologies such as gene sequencing, statistical genetics, and gene expression analysis to drugs in clinical development and on the market. It applies the large scale systematic approaches of genomics to speed the discovery of drug response markers, whether they act at the level of the drug target, drug metabolism, or disease pathways. The potential implication of genomics and pharmacogenomics in clinical research and clinical medicine is that disease could be treated according to genetic and specific individual markers, selecting medications and dosages that are optimized for individual patients. The possibility of defining patient populations genetically may improve outcomes by predicting individual responses to drugs, and could improve safety and efficacy in therapeutic areas such as neuropsychiatry, cardiovascular medicine, endocrinology (diabetes and obesity) and oncology. Ethical questions need to be addressed and guidelines established for the use of genomics in clinical research and clinical medicine. Significant achievements are possible with an interdisciplinary approach that includes genetic, technological and therapeutic measures. PMID- 10874051 TI - QT dispersion in medicine: electrophysiological holy grail or fool's gold? PMID- 10874052 TI - Intravenous immunoglobulin for ANCA-associated systemic vasculitis with persistent disease activity. AB - Intravenous immunoglobulin (IVIg) is a potential alternative treatment for anti neutrophil cytoplasm antibody (ANCA)-associated systemic vasculitis (AASV) with less toxicity than conventional immunosuppressive agents. This randomized, placebo-controlled trial aimed to investigate the efficacy of a single course of IVIg (total dose 2 g/kg) in previously-treated AASV with persistent disease activity in whom there was an intention to escalate therapy. Vasculitic activity was monitored by the Birmingham vasculitis activity score (BVAS), C-reactive protein (CRP) and ANCA levels. Treatment response was defined as a reduction in BVAS of more than 50% after 3 months, and there was an intention to keep doses of concurrent immunosuppressive drugs unchanged during this period; follow-up continued to 12 months. Seventeen patients were randomized to receive IVIg and 17 to receive placebo. Treatment responses were found in 14/17 and 6/17 of the IVIg and placebo groups, respectively (p=0.015, OR 8.56, 95%CI 1.74-42.2). Following infusion of trial medication, greater falls in CRP were seen at 2 weeks (p=0.02) and 1 month (p=0.04) in the IVIg group. No differences were observed between ANCA levels or cumulative exposure to immunosuppressive drugs, and after 3 months there were no differences in CRP levels or disease activity between the IVIg and placebo groups. Seventeen adverse effects occurred after IVIg and six after placebo: they were mostly mild, although reversible rises in serum creatinine occurred in four from the IVIg group. A single course of IVIg reduced disease activity in persistent AASV, but this effect was not maintained beyond 3 months; mild, reversible side-effects following IVIg were frequent. IVIg is an alternative treatment for AASV with persistent disease activity after standard therapy. PMID- 10874053 TI - Increased augmentation index and systolic stress in type 1 diabetes mellitus. AB - Type 1 diabetes mellitus is associated with endothelial dysfunction and increased arterial stiffness, both of which may contribute to the excess cardiovascular mortality in such patients. Arterial stiffening increases pulse wave velocity and wave reflection, which augments central systolic pressure and stress. Using the non-invasive technique of pulse wave analysis, we investigated aortic augmentation and central pressure in 35 patients with type 1 diabetes and 35 matched controls. Peripheral pulse waveforms were recorded from the radial artery. Central aortic waveforms were then generated, and augmentation index (AIx), ascending aortic pressure and tension time index (TTI), a measure of systolic load, were calculated. Peripheral and central blood pressure did not differ between the two groups. AIx was significantly elevated in the diabetic patients compared with controls (7.1+/-1.6% vs. 0.4+/-2.0%; p=0.01), as was the TTI (2307+/-51 mmHg x s x min(-1) vs. 2010+/-61 mmHg. s x min(-1); p<0.001). Estimated pulse wave velocity was also higher in the diabetic group. Type 1 diabetes is associated with an increased AIx and rate of wave travel, indicating enhanced wave reflection and increased systemic arterial stiffness, and elevation of the TTI. Such haemodynamic effects may contribute to the increased left ventricular mass and risk of cardiovascular disease associated with type 1 diabetes mellitus. PMID- 10874054 TI - High ethanol intake and malnutrition in alcoholic cerebellar shrinkage. AB - To determine the influence of chronic ethanol intake and nutritional status on cerebellar shrinkage in alcoholism, we studied 12 undernourished patients with acute Wernicke's encephalopathy (WE), 12 undernourished and 24 well-nourished asymptomatic chronic alcoholics, and 24 age-matched well-nourished controls, using morphometric analysis of MRI scans with volumetry of the cerebellum. Alcoholics reported a mean daily intake of ethanol of 177+/-8 g over a period of 27+/-1 years. Most undernourished alcoholics and half of the well-nourished alcoholics, compared to one-tenth of the controls, showed a significant reduction in cerebellar volume (p< or =0.01, both). Alcoholics with cerebellar shrinkage (n=33) were older (p=0.05) and tended to report greater daily ethanol intake than alcoholics without cerebellar shrinkage (n=15), although not significantly so (p=0.09). Cerebellar volume correlated negatively with age in controls and asymptomatic alcoholics (r> or =0.52, p< or =0.01, both), with a significantly greater shrinkage for age in the latter (p=0.003). Logistic regression analysis showed that malnutrition (OR 6.6 [95%CI 1.7-25.6], p=0.005) and a daily ethanol intake of more than 140 g over ten years (OR 6.1 [95%CI 1.8-20.5], p=0.003) were independently associated with the development of cerebellar shrinkage. PMID- 10874055 TI - The renal pathology of primary antiphospholipid syndrome: a distinctive form of endothelial injury. AB - Some features of the vascular and glomerular pathology of primary antiphospholipid syndrome (APS) are well recognized, but we describe novel glomerular ultrastructural changes that we consider to be pathognomonic of APS. Renal biopsies from eight patients with APS were examined by light and electron microscopy. All had anti-cardiolipin antibodies, and the clinical presentation ranged from fulminant multi-system disease to isolated proteinuria. By light microscopy, the hexamine silver stain showed a combination of glomerular basement membrane wrinkling and reduplication. By electron microscopy, redundant, wrinkled segments of basement membrane were accompanied by a 'new' straighter thin basement membrane adjacent to the endothelium. In two cases the presence of these antibodies was not suspected clinically, and there was no clinical history or evidence of a thrombotic microangiopathy. We describe a distinctive glomerular lesion that represents an unexplained form of endothelial injury in this syndrome. PMID- 10874056 TI - Mast cell: pivotal player in lethal acute pancreatitis. PMID- 10874058 TI - Contributions by German emigres to British medical science. PMID- 10874057 TI - The thyroid, blood flow and atheroma. PMID- 10874059 TI - Mass spectrometric analysis of rhodopsin from light damaged rats. AB - PURPOSE: It is well established that the retina is damaged by intense visible light. Rhodopsin has been proposed to be involved in this process. We therefore undertook to examine whether rhodopsin isolated from light damaged animals is structurally altered at the molecular level. METHODS: Dark reared and dim cyclic light reared 8 week old Sprague-Dawley rats were exposed to intense visible light and sacrificed immediately or 24 h after exposure together with unexposed control animals reared under the same conditions. Rod outer segments were isolated by sucrose gradient ultracentrifugation, their membranes treated with urea, then washed with Tris buffer. The rhodopsin preparations were then reduced, pyridylethylated, delipidated, and cleaved with CNBr. Reversed phase HPLC was used to separate the fragments, and the effluent was analyzed online with a Finnigan LCQ ion trap mass spectrometer. C-terminal phosphorylation was investigated following Asp-N cleavage. MALDI-TOF mass spectrometry was used for the identification of glycosylation. RESULTS: The rat rhodopsin protein was mapped with the exception of two single amino acid fragments. The reported sequence was confirmed with the exception of the controversial T/S320 residue, which was found to be a threonine. Mono-, di-, tri-, and tetraphosphorylated forms of rhodopsin were found in the light damaged animals. Three sites of phosphorylation were confirmed with MS/MS (tandem mass spectral) data. Single or double phosphorylations were found among these three sites, in various combinations. Dark adaptation completely reversed the phosphorylation in all light damaged animals. Other posttranslational modifications were as previously reported. CONCLUSIONS: Our results indicate that intense visible light exposure of rats does not lead to oxidative or other primary structural alterations in the rhodopsin protein of rod outer segments. We also report that the mutated rhodopsin (P23H) is present in rat rod outer segments from heterozygous animals and that residue 320 in both normal and mutated rhodopsins is threonine, not serine. PMID- 10874060 TI - The risk of the hemolytic-uremic syndrome after antibiotic treatment of Escherichia coli O157:H7 infections. AB - BACKGROUND: Children with gastrointestinal infections caused by Escherichia coli O157:H7 are at risk for the hemolytic-uremic syndrome. Whether antibiotics alter this risk is unknown. METHODS: We conducted a prospective cohort study of 71 children younger than 10 years of age who had diarrhea caused by E. coli O157:H7 to assess whether antibiotic treatment in these children affects the risk of the hemolytic-uremic syndrome and to assess the influence of confounding factors on this outcome. Estimates of relative risks were adjusted for possible confounding effects with the use of logistic-regression analysis. RESULTS: Among the 71 children, 9 (13 percent) received antibiotics and the hemolytic-uremic syndrome developed in 10 (14 percent). Five of these 10 children had received antibiotics. Factors significantly associated with the hemolytic-uremic syndrome were a higher initial white-cell count (relative risk, 1.3; 95 percent confidence interval, 1.1 to 1.5), evaluation with stool culture soon after the onset of illness (relative risk, 0.3; 95 percent confidence interval, 0.2 to 0.8), and treatment with antibiotics (relative risk, 14.3; 95 percent confidence interval, 2.9 to 70.7). The clinical and laboratory characteristics of the 9 children who received antibiotics and the 62 who did not receive antibiotics were similar. In a multivariate analysis that was adjusted for the initial white-cell count and the day of illness on which stool was obtained for culture, antibiotic administration remained a risk factor for the development of the hemolytic uremic syndrome (relative risk, 17.3; 95 percent confidence interval, 2.2 to 137). CONCLUSIONS: Antibiotic treatment of children with E. coli O157:H7 infection increases the risk of the hemolytic-uremic syndrome. PMID- 10874061 TI - Electrophysiologic testing to identify patients with coronary artery disease who are at risk for sudden death. Multicenter Unsustained Tachycardia Trial Investigators. AB - BACKGROUND: The mortality rate among patients with coronary artery disease, abnormal ventricular function, and unsustained ventricular tachycardia is high. The usefulness of electrophysiologic testing for risk stratification in these patients is unclear. METHODS: We performed electrophysiologic testing in patients who had coronary artery disease, a left ventricular ejection fraction of 40 percent or less, and asymptomatic, unsustained ventricular tachycardia. Patients in whom sustained ventricular tachyarrhythmias could be induced were randomly assigned to receive either antiarrhythmic therapy guided by electrophysiologic testing or no antiarrhythmic therapy. The primary end point was cardiac arrest or death from arrhythmia. Patients without inducible tachyarrhythmias were followed in a registry. We compared the outcomes of 1397 patients in the registry with those of 353 patients with inducible tachyarrhythmias who were randomly assigned to receive no antiarrhythmic therapy in order to assess the prognostic value of electrophysiologic testing. RESULTS: Patients were followed for a median of 39 months. In a Kaplan-Meier analysis, two-year and five-year rates of cardiac arrest or death due to arrhythmia were 12 and 24 percent, respectively, among the patients in the registry, as compared with 18 and 32 percent among the patients with inducible tachyarrhythmias who were assigned to no antiarrhythmic therapy (adjusted P<0.001). Overall mortality after five years was 48 percent among the patients with inducible tachyarrhythmias, as compared with 44 percent among the patients in the registry (adjusted P=0.005). Deaths among patients without inducible tachyarrhythmias were less likely to be classified as due to arrhythmia than those among patients with inducible tachyarrhythmias (45 and 54 percent, respectively; P=0.06). CONCLUSIONS: Patients with coronary artery disease, left ventricular dysfunction, and asymptomatic, unsustained ventricular tachycardia in whom sustained ventricular tachyarrhythmias cannot be induced have a significantly lower risk of sudden death or cardiac arrest and lower overall mortality than similar patients with inducible sustained tachyarrhythmias. PMID- 10874062 TI - The effect of celecoxib, a cyclooxygenase-2 inhibitor, in familial adenomatous polyposis. AB - BACKGROUND: Patients with familial adenomatous polyposis have a nearly 100 percent risk of colorectal cancer. In this disease, the chemopreventive effects of nonsteroidal antiinflammatory drugs may be related to their inhibition of cyclooxygenase-2. METHODS: We studied the effect of celecoxib, a selective cyclooxygenase-2 inhibitor, on colorectal polyps in patients with familial adenomatous polyposis. In a double-blind, placebo-controlled study, we randomly assigned 77 patients to treatment with celecoxib (100 or 400 mg twice daily) or placebo for six months. Patients underwent endoscopy at the beginning and end of the study. We determined the number and size of polyps from photographs and videotapes; the response to treatment was expressed as the mean percent change from base line. RESULTS: At base line, the mean (+/-SD) number of polyps in focal areas where polyps were counted was 15.5+/-13.4 in the 15 patients assigned to placebo, 11.5+/-8.5 in the 32 patients assigned to 100 mg of celecoxib twice a day, and 12.3+/-8.2 in the 30 patients assigned to 400 mg of celecoxib twice a day (P=0.66 for the comparison among groups). After six months, the patients receiving 400 mg of celecoxib twice a day had a 28.0 percent reduction in the mean number of colorectal polyps (P=0.003 for the comparison with placebo) and a 30.7 percent reduction in the polyp burden (the sum of polyp diameters) (P=0.001), as compared with reductions of 4.5 and 4.9 percent, respectively, in the placebo group. The improvement in the extent of colorectal polyposis in the group receiving 400 mg twice a day was confirmed by a panel of endoscopists who reviewed the videotapes. The reductions in the group receiving 100 mg of celecoxib twice a day were 11.9 percent (P=0.33 for the comparison with placebo) and 14.6 percent (P=0.09), respectively. The incidence of adverse events was similar among the groups. CONCLUSIONS: In patients with familial adenomatous polyposis, six months of twice-daily treatment with 400 mg of celecoxib, a cyclooxygenase-2 inhibitor, leads to a significant reduction in the number of colorectal polyps. PMID- 10874063 TI - Incidence of cancer after prophylaxis with warfarin against recurrent venous thromboembolism. Duration of Anticoagulation Trial. AB - BACKGROUND: The length of time after an episode of venous thromboembolism during which the risk of newly diagnosed cancer is increased is not known, and whether vitamin K antagonists have an antineoplastic effect is controversial. METHODS: In a prospective, randomized study of the duration of oral anticoagulation (six weeks or six months) after a first episode of venous thromboembolism, patients were questioned annually about any newly diagnosed cancer. After a mean follow-up of 8.1 years, we used the Swedish Cancer Registry to identify all diagnoses of cancer and causes of death in the study population. The observed numbers of cases of cancer were compared with expected numbers based on national incidence rates, and the standardized incidence ratios were calculated. RESULTS: A first cancer was diagnosed in 111 of 854 patients (13.0 percent) during follow-up. The standardized incidence ratio for newly diagnosed cancer was 3.4 (95 percent confidence interval, 2.2 to 4.6) during the first year after the thromboembolic event and remained between 1.3 and 2.2 for the following five years. Cancer was diagnosed in 66 of 419 patients (15.8 percent) who were treated for six weeks with oral anticoagulants, as compared with 45 of 435 patients (10.3 percent) who were treated for six months (odds ratio, 1.6; 95 percent confidence interval, 1.1 to 2.4). The difference was mainly due to the occurrence of new urogenital cancers, of which there were 28 cases in the six-week group (6.7 percent) and 12 cases in the six-month group (2.8 percent) (odds ratio, 2.5; 95 percent confidence interval, 1.3 to 5.0). The difference in the incidence of cancer between the treatment groups became evident only after two years of follow-up, and it remained significant after adjustment for sex, age, and whether the thromboembolism was idiopathic or nonidiopathic. Older age at the time of the venous thrombosis and an idiopathic thromboembolism were also independent risk factors for a diagnosis of cancer. No difference in the incidence of cancer related deaths was detected. CONCLUSIONS: The risk of newly diagnosed cancer after a first episode of venous thromboembolism is elevated during at least the following two years. Subsequently, the risk seems to be lower among patients treated with oral anticoagulants for six months than among those treated for six weeks. PMID- 10874064 TI - Images in clinical medicine. Pulmonary Langerhans'-cell granulomatosis (histiocytosis X). PMID- 10874065 TI - Chemoprevention of colorectal cancer. PMID- 10874067 TI - A farewell. PMID- 10874066 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 20-2000. A 61-year-old man with a wide-complex tachycardia. PMID- 10874068 TI - Marcia Angell and the Journal. PMID- 10874069 TI - E. coli, antibiotics, and the hemolytic-uremic syndrome. PMID- 10874070 TI - Warfarin for cancer prevention. PMID- 10874071 TI - Revisiting the Canadian health care system. PMID- 10874072 TI - Improved rapid analytical method for the urinary determination of 3, 5,6 trichloro-2-pyridinol, a metabolite of chlorpyrifos. PMID- 10874073 TI - An evaluation of glutathione S-transferase activity in the Tasmanian lacewing (Micromus tasmaniae) as a biomarker of organophosphate contamination. PMID- 10874074 TI - New method for the assessment of contaminant uptake routes in the oligochaete Lumbriculus variegatus. PMID- 10874075 TI - Metabolic conversion of N'-(2,4-dimethylphenyl)-N-methylformamidine pesticide and the analysis of the metabolites. PMID- 10874076 TI - Mineralization of trichloroethylene (TCE) by the white rot fungus Phanerochaete chrysosporium. PMID- 10874077 TI - Organochlorine residues in the waters of Keoladeo National Park, Bharatpur, Rajasthan. PMID- 10874078 TI - Contaminants in eggs of western snowy plovers and California least terns: is there a link to population decline? PMID- 10874079 TI - Comparative analysis of heavy metals in two species of ichthyophagous bats Myotis vivesi and Noctilio leporinus. PMID- 10874080 TI - Accumulation of chromium and copper in three different soils and bioaccumulation in an aquatic plant, Alternanthera philoxeroides. PMID- 10874081 TI - Procedure for upgrading contaminant-detection networks in aquifers. PMID- 10874082 TI - Effect of chlorination on microfauna communities in activated sludge plants. PMID- 10874083 TI - Comparison of tissue lesions in four species of benthic fish sampled in 1972-1973 and 1997-1998 on the Grand Banks off Newfoundland. PMID- 10874084 TI - Chlorpyrifos in catfish (Ictalarus punctatus) tissue. PMID- 10874085 TI - Observations on the experimental assessment of optimal exposure time for mercury detoxification by an integrated aquatic macrophyte base system. PMID- 10874086 TI - Effects of ammonia on mortality and feeding of postlarvae shrimp Litopenaeus vannamei. PMID- 10874087 TI - Acute toxicity to the golden apple snail and estimated bioconcentration potential of triphenylphosphine oxide and series of related compounds. PMID- 10874088 TI - Screening methods for drugs and heavy metals in Chinese patent medicines. PMID- 10874089 TI - Defluoridation of groundwater in Agra city using low cost adsorbents. PMID- 10874091 TI - Contact dermatitis due to student's clothing. PMID- 10874090 TI - Sex ratio at birth associated with petrochemical air pollution in Taiwan. PMID- 10874100 TI - Importance, morbidity, and mortality of pneumonia in the surgical intensive care unit. AB - Surgical patients are at high risk to develop nosocomial pneumonia, although an accurate diagnosis is difficult to make. Staphylococcus aureus and Pseudomonas aeruginosa are the most common pathogens, but Acinetobacteris emerging as an important pathogen. Because affected patients are often critically ill with multisystem pathology, it can be difficult to ascribe morbidity or mortality directly to the infection. PMID- 10874101 TI - Discussion. PMID- 10874102 TI - Identification of patients at highest risk for ventilator-associated pneumonia in the surgical intensive care unit. AB - Ventilator-acquired pneumonia (VAP) is a major problem for patients admitted to the surgical intensive care unit mechanically ventilated. Recently, we have identified both clinical and immunologic factors associated with the development of VAP. The clinical risk factors are associated with the severity of the injury and the length of mechanical ventilation. The immunologic risk factors are associated with the local lung inflammatory response that is unchecked and affects local cell function. The combination of the severity of injury, the length of mechanical ventilation, and the failure to "auto-regulate" the lung response places the host at risk of VAP. In the next millennium, if we are to make significant advances in the management of VAP, we will need to understand the pathophysiology of the disease process. Then we can develop preventive strategies that will reduce the morbidity and the associated cost of VAP. PMID- 10874103 TI - Discussion. PMID- 10874104 TI - Prevention of ventilator-associated pneumonia. AB - Ventilator-associated pneumonia (VAP) remains a common complication within the intensive care unit. Despite identification of the patients at highest risk for developing VAP, the actual incidence appears to be unchanged over the past two decades. Colonization of the oral pharynx with pathogenic organisms and ongoing aspiration appear to be necessary in the development of VAP. Thus, prevention strategies targeted at interrupting these factors should decrease VAP. In the few randomized prospective studies in the literature, both continuous aspiration of the subglottic space and the prevention colonization of the oral pharynx have been proven successful in decreasing VAP. The main problem in interpreting these results is that they have been generally single reports and need to be replicated in larger multicenter trials. Studies of "early" tracheostomy have been unable to define both the optimal timing of tracheostomy and its effect in decreasing VAP. Other modalities, such as rotating or kinetic beds, early bronchoscopy, and changes in ventilator management, have not been shown to be useful. PMID- 10874105 TI - Discussion. PMID- 10874106 TI - Empiric therapy for pneumonia in the surgical intensive care unit. AB - Empiri c therapy of ventilator-associated pneumonia (VAP) in surgical patients should be based on intensive care unit (ICU)-specific surveillance data, because microbial flora patterns vary widely between geographic regions as well as within hospitals. Surgical ICUs have higher VAP rates than other units. Data from the National Nosocomial Infection Surveillance (NNIS) System report Pseudomonas aeruginosa and Staphylococcus aureus to be the most frequent isolates (each 17.4%). Data from the NNIS documents high resistance patterns in ICUs compared with hospitals at large, as well as unit-specific patterns. VAP risk factors for surgical patients include thoracoabdominal surgery, altered level of consciousness, advanced age, diabetes mellitus, malnutrition, chronic obstructive pulmonary disease, and prior antibiotic administration. Promising prevention strategies include restricting ventilator circuit changes, in-line heat moisture exchange filters, semi-recumbant positioning, and continuous subglottic aspiration. Pharmacodynamics should be considered when choosing antibiotic regimens. Postantibiotic effect and time-dependent versus concentration-dependent killing should be studied in clinical trials. Current guidelines for choosing regimens have been well developed by the American Thoracic Society. PMID- 10874107 TI - Discussion. PMID- 10874108 TI - Diagnosis of acute respiratory distress syndrome and differentiation from ventilator-associated pneumonia. AB - Acute respiratory distress syndrome (ARDS) remains a significant cause of morbidity and mortality in surgical intensive care unit patients. A relatively predictable sequence of pathophysiologic events occur in the lung, which involve inflammatory mediators and neutrophils. The characteristic radiographic findings of ARDS may mimic ventilator-associated pneumonia (VAP), making the diagnosis of VAP difficult. The standard clinical criteria of fever, leukocytosis, purulent sputum, and infiltrate on chest radiograph are not specific for pneumonia in the surgical patient. The use of bronchoscopy with bronchoalveolar lavage and quantitative cultures can differentiate local and systemic inflammatory response to injury or blood loss from invasive bacterial pneumonia. Basing antibiotic therapy solely on the results of quantitative cultures is safe, because quantitative cultures identify VAP in less than half the patients with clinical evidence of pneumonia. Empiric therapy should be based on the microbiology of the intensive care unit rather than the results of the Gram stain. PMID- 10874109 TI - Discussion. PMID- 10874110 TI - Multifactorial analyses in the diagnosis of pneumonia arising in the surgical intensive care unit. AB - The diagnosis of ventilator-associated pneumonia in the surgical intensive care unit continues to be problematic. The majority of intensive care units use clinical criteria based on chest x-ray; fever; leukocytosis; alterations in the pulse oximeter observations; the need to alter modes and amounts of ventilatory support; and more specific microbiologic studies, such as appropriate sputum, Gram stain, and culture to identify pneumonia. Diagnosing pneumonia based on clinical criteria alone is often difficult and inaccurate, which may lead to inappropriate use and choice of antibiotics. Invasive diagnostic techniques, such as protected specimen brush and bronchoalveolar lavage, provide an important microbiologic diagnosis. However, the cost and inconvenience limit broad usage. Furthermore, those results that return positive are often too late to dictate the need for, or direction of, therapy. Our use of a "pneumonia grid" may help identify patients likely to have a poor outcome. Until a readily available and cost-effective diagnostic study for pneumonia is developed, clinical criteria remain vital in routine practice. PMID- 10874111 TI - Pneumonia in the surgical patient: duration of therapy and does the organism matter? AB - Pneumonia is a frequent complication in the critically ill surgical or trauma patient. Despite its common occurrence and increased attention recently, many basic issues, such as how long to treat and whether the exact causative organism even matters, remain unanswered. Currently, cessation of treatment is based on clinical response, although some data suggest that both the microbiologic and immunologic response within the lung are also important. A thorough knowledge of the likely causative organisms for both early and late pneumonia should allow safe and efficacious empiric antibiotic selection. PMID- 10874112 TI - Discussion. PMID- 10874113 TI - Immunologic responses to pneumonia. AB - Pneumonia in the critically ill surgical patient often results from the bombardment of a previously normal pulmonary system with therapeutic foreign bodies, hospital pathogens, and impairment of the host defenses. Despite its long history as a significant clinical problem, a woefully inadequate amount of study has been directed toward therapy. We created an experimental model of a differential pulmonary infection using a strain of Klebsiella pneumoniae. We then compared the progressively affected pneumonic process versus the normal parenchyma. We measured neutrophil and monocyte complement antibody receptor expression and monocyte and macrophage class II major histocompatibility antigens (HLA-DR) via percent of cells and mean fluorescent intensity outcomes from flow cytometry. The main difference between infected versus noninfected tissues was monocyte DR expression, which was consistently depressed in cells from infected parenchyma. What follows is a discussion of the implications of this work as well as other work in the immunology of pneumonia and cytokine expression. Possible therapeutic modalities are included. PMID- 10874114 TI - Role of aztreonam in the treatment of nosocomial pneumonia in the critically ill surgical patient. AB - In 1995 the American Thoracic Society issued an official consensus statement on the treatment of hospital-acquired pneumonia (HAP). Classes of antimicrobials included in the list of antimicrobials deemed to be suitable for the empiric treatment of severe HAP were the aminoglycosides, quinolones, antipseudomonal penicillins, carbapenems, and beta-lactam/beta-lactamase inhibitor combinations. Aztreonam, a monobactam, was also listed and is unique among these agents based on its spectrum of activity being limited to the gram-negative bacillary bacteria combined with an excellent safety profile. This review focuses on the role of aztreonam in the treatment of nosocomial pneumonia in the critically ill patient.A review of the literature was performed using PubMed and secondary literature sources as to the clinical efficacy of aztreonam in the treatment of lower respiratory tract infections as well as its pharmacokinetic and safety profiles. An analysis of aztreonam's potential pharmacoeconomic advantages compared with other agents was also performed.Numerous studies have documented that aztreonam has effectiveness that is equal or superior to that of other suitable antibiotics in the treatment of nosocomial pneumonia. Its excellent safety profile makes it a particularly attractive agent compared with the aminoglycosides. Considering the potential costs of bacterial resistance from the use of broader-spectrum alternatives, a case can be made that aztreonam is a pharmacoeconomically sound choice as well. PMID- 10874115 TI - Discussion. PMID- 10874116 TI - Pharmacoeconomics of pneumonia. AB - Because diagnosis and treatment are so intimately linked, the pharmacoeconomics of treatment of ventilator-associated pneumonia (VAP) is impossible to discuss without discussing the cost-effectiveness of VAP diagnosis. The cost of VAP treatment is more complex than simply drug acquisition and administration costs. The critical factor in cost-effective therapy is the avoidance of inappropriate or ineffective therapy. The second most important benefit of a more accurate diagnostic strategy, such as the use of quantitative cultures, is the ability either to stop or to withhold antibiotics if the quantitative culture is negative. Therefore, the benefit of any diagnostic strategy must be evaluated principally from the aspect of these resultant changes in management. Reassurance or concern about an alternative site of infection or cause of fever will also add to the benefit or cost of more accurate diagnosis of VAP. The baseline antibiotic treatment strategy of the specific intensive care unit (ICU) will determine, to a large degree, the cost of antibiotics and the efficacy of empiric regimens. In the final analysis, pharmacy costs and cost of diagnostic testing for VAP must be based on outcome analysis, including comparison of the more expensive aspects of care, such as mortality, length of mechanical ventilation, and length of ICU stay. PMID- 10874117 TI - Single versus combination antimicrobial therapy for ventilator-associated pneumonia. AB - The appropriate selection of definitive antimicrobial therapy is a necessary component of the overall treatment for ventilator-associated pneumonia. When possible, single-agent therapy is preferable. A combination of antibiotics is necessary to treat multiple organisms not susceptible to a single appropriate antibiotic and when antibiotic-resistant gram-negative bacteria are present. Treatment failure is more commonly the result of persistent pneumonia and the development of antibiotic resistance than to recurrence after successful antimicrobial therapy. The duration of treatment will vary depending on the severity of the underlying illness and the pneumonic process. PMID- 10874118 TI - Complications of nosocomial pneumonia in the surgical patient. AB - Nosocomial pneumonia is a leading cause of morbidity and mortality in the surgical and trauma patient. Inadequate treatment can lead to the complications of acute respiratory distress syndrome (ARDS), empyema, and lung abscess. The prevention and treatment of these complications revolve around several key principles. Complete treatment of pneumonia requires appropriate antimicrobial therapy, as well as mechanical pulmonary hygiene and proper airway management. Despite advances in treatment of pneumonia, complications arise necessitating treatment. This article reviews the treatment of ARDS, empyema, and lung abscesses. In particular, the many options for treatment of empyema are discussed in detail. Additionally, the treatment of pulmonary contusion and hemopneumothorax in the trauma patient is discussed. The understanding of sound treatment principles in the critically ill postsurgical patient helps prevent complicated or recurrent pneumonia and allows the surgeon to intervene effectively when such complications occur. PMID- 10874119 TI - Discussion. PMID- 10874120 TI - Antibiotic efflux pumps. AB - Active efflux from procaryotic as well as eucaryotic cells strongly modulates the activity of a large number of antibiotics. Effective antibiotic transport has now been observed for many classes of drug efflux pumps. Thus, within the group of primary active transporters, predominant in eucaryotes, six families belonging to the ATP-binding cassette superfamily, and including the P-glycoprotein in the MDR (Multi Drug Resistance) group and the MRP (Multidrug Resistance Protein), have been recognized as being responsible for antibiotic efflux. Within the class of secondary active transporters (antiports, symports, and uniports), ten families of antibiotic efflux pumps have been described, distributed in five superfamilies [SMR (Small Multidrug Resistance), MET (Multidrug Endosomal Transporter), MAR (Multi Antimicrobial Resistance), RND (Resistance Nodulation Division), and MFS (Major Facilitator Superfamily)]. Nowadays antibiotic efflux pumps are believed to contribute significantly to acquired bacterial resistance because of the very broad variety of substrates they recognize, their expression in important pathogens, and their cooperation with other mechanisms of resistance. Their presence also explains high-level intrinsic resistances found in specific organisms. Stable mutations in regulatory genes can produce phenotypes of irreversible multidrug resistance. In eucaryotes, antibiotic efflux pumps modulate the accumulation of antimicrobials in phagocytic cells and play major roles in their transepithelial transport. The existence of antibiotic efflux pumps, and their impact on therapy, must now be taken fully into account for the selection of novel antimicrobials. The design of specific, potent inhibitors appears to be an important goal for the improved control of infectious diseases in the near future. PMID- 10874121 TI - Antioxidative properties of natural coelenterazine and synthetic methyl coelenterazine in rat hepatocytes subjected to tert-butyl hydroperoxide-induced oxidative stress. AB - Coelenterazine (CLZn; 3, 7-dihydro-2-(p-hydroxybenzyl)-6-(p-hydroxyphenyl)-8 benzylimidazolo++ +[1 ,2-a]pyrazin-3-one), the substrate for bioluminescence reactions in many marine animals, is endowed with high antioxidant properties. This work investigated the antioxidative properties of CLZn in primary cultures of rat hepatocytes subjected to the oxidant tert-butyl hydroperoxide (t-BHP). Micromolar concentrations of CLZn increased survival and decreased lipid peroxidation in rat hepatocytes subjected for 6 hr to 2.5 x 10(-4) M t-BHP. However, the extent of protection was limited by a strong toxicity of CLZn (IC(50) = 6.9 x 10(-5) M). The presence of t-BHP increased the cellular toxicity of CLZn. Methyl coelenterazine (CLZm, 3, 7-dihydro-2-methyl-6-(p-hydroxyphenyl)-8 benzylimidazolo[1, 2-a]pyrazin-3-one), a synthetic analogue of CLZn, demonstrated excellent antioxidant properties, even at very low (3 x 10(-6) M) concentrations and was not toxic throughout most of its effective concentration range. CLZm proved far more effective than reference antioxidants such as Trolox C(R), alpha tocopherol, BHT, and probucol. The assay of thiobarbituric reactive substances (TBARS) associated with cells and in the culture medium indicated that 10(-5) M CLZm provided a total protection against t-BHP-induced lipid peroxidation. This coelenterazine analogue could be used as a model compound for investigating the action mechanism of imidazolopyrazinones in mammalian hepatocytes. PMID- 10874122 TI - Determination of the DNA sequences of acetylcholinesterase and butyrylcholinesterase from cat and demonstration of the existence of both in cat plasma. AB - Cat serum contains 0.5 mg/L of butyrylcholinesterase (BChE, EC 3.1.1. 8) and 0.3 mg/L of acetylcholinesterase (AChE, EC 3.1.1.7); this can be compared with 5 mg/mL and < 0.01 mg/L, respectively, in human serum. Cat BChE differed from human BChE in the steady-state turnover of butyrylthiocholine, having a 3-fold higher k(cat) and 2-fold higher K(m) and K(ss) values. Sequencing of the cat BCHE cDNA revealed 70 amino acid differences between cat and human BChE, three of which could account for these kinetic differences. These amino acids, which were located in the region of the active site, were Phe398Ile, Pro285Leu, and Ala277Leu (where the first amino acid was found in human and the second in cat). Sequencing genomic DNA for cat and human ACHE demonstrated that there were 33 amino acid differences between the cat and human AChE enzymes, but that there were no differences in the active site region. In addition, a polymorphism in intron 3 of the human ACHE gene was detected, as well as a silent polymorphism at Y116 of the cat ACHE gene. PMID- 10874124 TI - Inhibition of a phosphodiesterase III in the lysis-sensitive target-induced elevation of cyclic AMP (cAMP) in human natural killer cells. AB - Natural killer (NK) cells are lymphocytes that are capable of destroying tumor cells and virally infected cells (cytolysis) without prior sensitization. When cyclic AMP (cAMP) is elevated artificially in NK cells, it is a potent inhibitor of their cytolytic function. Recently, we have shown that when NK cells are exposed to a range of lysis-sensitive (LS) tumor target cells, there is an increase in intracellular cAMP levels in the NK cells over a 60-min period. There is no increase in NK-cell cAMP in response to lysis-resistant (LR) tumor target cells. We determined that this cAMP elevation is due, in part, to an LS target induced activation of adenylyl cyclase (AC), and that the AC-activation component appears to require a protein tyrosine kinase (PTK) activity. In the present study, we demonstrated that an LS target-induced inhibition of phosphodiesterase (PDE) is also contributing to the overall elevation of cAMP. Direct measurement of PDE activity showed an inhibition in lymphocytes that were exposed to LS targets but not in those exposed to LR targets. The inhibition of PDE activity was maximal by 30 min. Lymphocytes were exposed to targets and then lysed, so that PDE activity could be measured. Addition of class-selective inhibitors of PDE (at levels sufficient to completely block that class of PDE) to the lysate focused the measurement of PDE activity on those classes of PDE that were unaffected by the selective inhibitor. Using the PDE IV selective inhibitor rolipram and the PDE III selective inhibitors trequinsin and milrinone, we showed that a PDE III is being inhibited in lymphocytes by exposure to LS targets. As PDE III is known to be inhibited by elevated cyclic GMP (cGMP) levels, increased cGMP in NK cells following exposure to LS targets was a possible mechanism by which a PDE III in NK cells might be inhibited. However, when we measured cGMP levels in control and LS target-stimulated lymphocytes, we saw no change. PMID- 10874123 TI - Multiple levels of regulation of selenoprotein biosynthesis revealed from the analysis of human glioma cell lines. AB - To gain a better understanding of the biological consequences of the exposure of tumor cells to selenium, we evaluated the selenium-dependent responses of two selenoproteins (glutathione peroxidase and the recently characterized 15-kDa selenoprotein) in three human glioma cell lines. Protein levels, mRNA levels, and the relative distribution of the two selenocysteine tRNA isoacceptors (designated mcm(5)U and mcm(5)Um) were determined for standard as well as selenium supplemented conditions. The human malignant glioma cell lines D54, U251, and U87 were maintained in normal or selenium-supplemented (30 nM sodium selenite) conditions. Northern blot analysis demonstrated only minor increases in steady state GSHPx-1 mRNA in response to selenium addition. Baseline glutathione peroxidase activity was 10.7 +/- 0.7, 7.6 +/- 0.7, and 4.3 +/- 0.7 nmol NADPH oxidized/min/mg protein for D54, U251, and U87, respectively, as determined by the standard coupled spectrophotometric assay. Glutathione peroxidase activity increased in a cell line-specific manner to 19.7 +/- 1.4, 15.6 +/- 2.1, and 6. 7 +/- 0.5 nmol NADPH oxidized/min/mg protein, respectively, as did a proportional increase in cellular resistance to H(2)O(2), in response to added selenium. The 15-kDa selenoprotein mRNA levels likewise remained constant despite selenium supplementation. The selenium-dependent change in distribution between the two selenocysteine tRNA isoacceptors also occurred in a cell line-specific manner. The percentage of the methylated isoacceptor, mcm(5)Um, changed from 35.5 to 47.2 for D54, from 38.1 to 47.3 for U251, and from 49.0 to 47.6 for U87. These data represent the first time that selenium-dependent changes in selenoprotein mRNA and protein levels, as well as selenocysteine tRNA distribution, were examined in human glioma cell lines. PMID- 10874125 TI - In vitro metabolism of acitretin by human liver microsomes: evidence of an acitretinoyl-coenzyme A thioester conjugate in the transesterification to etretinate. AB - The aromatic retinoid acitretin is the primary active metabolite of etretinate, and in this study we investigated the ethyl esterification of acitretin to etretinate using [(14)C]acitretin and human liver microsomes. Samples were analysed by TLC, HPLC, and LC-MS. Essential requirements for the transesterification reaction were identified and included viable microsomal protein, ATP, CoASH, and ethanol. Human liver microsomes catalysed formation of acitretinoyl-CoA at the rate of 0.08 +/- 0.02 nmol/min/mg (mean +/- SD, N = 10). Acitretinoyl-CoA was pivotal for the transesterification to etretinate and in the presence of methanol, ethanol, n-propanol, n-butanol, and hexanol, the corresponding esters, namely methyl-, ethyl (etretinate)-, propyl-, butyl-, and hexyl-acitretinate, were formed. On average, 1.7% of the acitretin present in the incubation was converted to etretinate in the presence of ethanol. In the absence of ethanol, transesterification did not proceed. Inhibition of the ester hydrolysis of etretinate by bis-p-nitrophenylphosphate (BNPP, 1 mM) prevented futile cycling of etretinate via acitretinoyl-CoA. An additional finding was that acitretin (15-30 microM) activated significantly human liver microsomal long chain fatty acid-CoA ligase (E.C.6.2.1.3, LCL), resulting in enhanced formation of palmitoyl-CoA. This study demonstrated that in the presence of ethanol the ethyl esterification of acitretin to etretinate proceeds via formation of acitretinoyl-CoA. Predicting clearance of acitretin in vivo via this unique metabolic pathway will be a challenge, as the intracellular concentration of ethanol could never be predicted with any degree of accuracy in humans. PMID- 10874127 TI - Inhibition of topoisomerase II by the marine alkaloid ascididemin and induction of apoptosis in leukemia cells. AB - Ascididemin (ASC) is a pentacyclic DNA-intercalating agent isolated from the Mediterranean ascidian Cystodytes dellechiajei. This marine alkaloid exhibits marked cytotoxic activities against a range of tumor cells, but its mechanism of action remains poorly understood. We investigated the effects of ASC on DNA cleavage by human topoisomerases I and II. Relaxation assays using supercoiled DNA showed that ASC stimulated double-stranded cleavage of DNA by topoisomerase II, but exerted only a very weak effect on topoisomerase I. ASC is a conventional topoisomerase II poison that significantly promoted DNA cleavage, essentially at sites having a C on the 3' side of the cleaved bond (-1 position), as observed with etoposide. The stimulation of DNA cleavage by topoisomerase I in the presence of ASC was considerably weaker than that observed with camptothecin. Cytotoxicity measurements showed that ASC was even less toxic to P388 leukemia cells than to P388CPT5 cells resistant to camptothecin. In addition, the marine alkaloid was found to be equally toxic to HL-60 leukemia cells sensitive or resistant to mitoxantrone. It is therefore unlikely that topoisomerases are the main cellular targets for ASC. This alkaloid was found to strongly induce apoptosis in HL-60 and P388 leukemia cells. Cell cycle analysis showed that ASC treatment was associated with a loss of cells in the G1 phase accompanied with a large increase in the sub-G1 region. Cleavage experiments with poly(ADP-ribose) polymerase (PARP) revealed that caspase-3 was a mediator of the apoptotic pathway induced by ASC. The DNA of ASC-treated cells was severely fragmented. Collectively, these findings indicate that ASC is a potent inducer of apoptosis in leukemia cells. PMID- 10874126 TI - Identification of human cytochrome P450 isoforms that contribute to all-trans retinoic acid 4-hydroxylation. AB - The role of specific human cytochrome P450 (CYP) isoforms in the oxidative metabolism of all-trans-retinoic acid was investigated by studies in human liver microsomes using isoform-specific chemical inhibitors and inhibitory antibodies. Studies using individual isoforms expressed in lymphoblastoid cells and correlation analysis using different microsome preparations were also performed. With expressed isoforms, evidence for a role for CYP2C8, CYP3A4, CYP2C9, and CYP1A1 in 4-hydroxylation was obtained, with the highest catalytic efficiency being observed for CYP2C8. Using inhibition studies and correlation analysis, we also concluded that CYP2C8 was the major all-trans-retinoic acid 4-hydroxylating cytochrome P450 in human liver microsomes, though CYP3A4 and, to a lesser extent CYP2C9, also made a contribution. In addition, we compared the rate of retinoic acid degredation in HepG2 cells when cultured in the absence and presence of 3 methylcholanthrene or all-trans-retinoic acid. Culture in the presence of all trans-retinoic acid decreased the half-life twofold and resulted in an increased sensitivity of retinoic acid degredation to ketoconazole. Since no induction of either CYP1A1, CYP2C8, CYP2C9, or CYP3A4 was detected using immunoblotting and as mRNA encoding another cytochrome P450 enzyme, CYP26, has been previously demonstrated to be induced by retinoic acid treatment of HepG2 cells and to be highly sensitive to ketoconazole, this enzyme in addition to CYP2C8, CYP2C9 and CYP3A4 likely plays a role in all-trans-retinoic acid oxidation in the liver at high retinoic acid levels. PMID- 10874128 TI - Selective inhibition of nitric oxide synthase type I by clonidine, an anti hypertensive drug. AB - Clonidine, clinically used in the treatment of hypertension, is a central alpha(2)-adrenergic agonist that reduces blood pressure and slows heart rate by reducing sympathetic stimulation. Considering the structural similarity between clonidine and hydrophobic heterocyclic nitric oxide synthase (NOS) inhibitors, the effect of clonidine on the nitric oxide (NO) pathway was investigated. This was verified by determination of NOS activity in vitro and by analysis of inducible Ca(2+)-independent NOS (NOS-II) mRNA expression and measurement of nitrite levels in rat C6 glioma cells, taken as a cellular model. Clonidine inactivated neuronal Ca(2+)-dependent NOS (NOS-I) competitively without affecting NOS-II and endothelial Ca(2+)-dependent NOS (NOS-III) activity. However, the value of K(i) for clonidine binding to NOS-I depended on tetrahydrobiopterin (BH(4)) concentration, as reported for NOS inhibition by other nitrogen heterocyclic compounds. In particular, the value of K(i) for clonidine binding to NOS-I increased (from [7. 9 +/- 0.4] x 10(-5) M to [8.0 +/- 0.4] x 10(-3) M) as BH(4) concentration was increased (between 3.0 x 10(-7) M and 1.0 x 10(-3) M), at pH 7.5 and 37.0 degrees. In addition, clonidine (1.0 x 10(-4) M) enhanced NOS-II mRNA expression in rat C6 glioma cells, as induced by Escherichia coli lipopolysaccharide (LPS) plus interferon-gamma (IFN-gamma). Finally, clonidine (1.0 x 10(-4) M to 1.0 x 10(-3) M) dose dependently increased the levels of LPS/IFN-gamma-induced nitrites, the breakdown product of NO, in supernatants of rat C6 glioma cells. As reported for other NOS inhibitors, clonidine was also able to regulate NOS-I and NOS-II inversely. PMID- 10874129 TI - Effects of dexamethasone on mitogen-activated protein kinases in mouse macrophages: implications for the regulation of 85 kDa cytosolic phospholipase A(2). AB - In mouse macrophages, arachidonate mobilisation in response to several stimuli is severely inhibited by prolonged (16-20 hr) treatment with nanomolar dexamethasone (dex). It was shown earlier that this inhibition was accompanied by a dual effect on cPLA(2); down-regulation of the enzyme protein and inhibition of its activation. We now report that cycloheximide, a protein synthesis inhibitor, caused an almost complete reversion of the inhibitory effects of dex on cPLA(2) activation. These results indicate that the effects depend on new protein synthesis. This is consistent with other data, obtained with a glucocorticoid receptor antagonist, indicating that the effects are mediated via the glucocorticoid receptor. Northern blot results showed pronounced down-regulation of cPLA(2) at the level of its mRNA. The possibility that dex also targeted the level or activation of one or more of the three mitogen-activated protein kinases (MAP kinases), extracellular signal-regulated kinase (ERK), p38, or c-Jun N terminal kinase (JNK) was also addressed. While the level of these MAP kinases and their phorbol myristate acetate (PMA)-induced activation were unaffected by dex, there was a partial inhibition of their zymosan-induced activation. However, this inhibition was not as pronounced as the dex-mediated inhibition of cPLA(2) activation. These data were confirmed by Western blot using antibodies against the phosphorylated forms of ERK, p38, and JNK. The results suggest that dex mediated inhibition of PMA-induced cPLA(2) activation is exerted downstream of the MAP kinases, while the partial inhibition of the zymosan-induced activation may be explained by effects exerted more upstream. Thus, the MAP kinases investigated here do not appear to be main targets for the inhibitory effects of dex on cPLA(2) activation. PMID- 10874130 TI - Distribution of liposome-encapsulated iodixanol in rat liver cells. AB - Distribution of liposome-encapsulated [(125)I]iodixanol in different types of liver cells following intravenous injection was studied in rats. The data showed that liposome-encapsulated [(125)I]iodixanol was rapidly taken up by the liver; after 15 min, radioactivity corresponding to nearly 25% of the injected radioactivity could be recovered therein. After 4 hr, approximately 60% of the injected radioactivity was in the liver. One week after injection, nearly 30% of the encapsulated radioactivity could still be recovered in the liver. Liposome encapsulated [(125)I]iodixanol was taken up both by hepatocytes and the Kupffer cells. On a per cell basis, the uptake of liposome-encapsulated [(125)I]iodixanol in Kupffer cells was more than 10-fold greater than that in hepatocytes, while the contribution of liver endothelial cells to uptake was negligible. Osmotic protection studies showed that iodixanol does not readily diffuse across lysosomal membranes, indicating that loss of iodixanol from the liver probably occurred by recycling rather than by diffusion across phagolysosomal and plasma membranes. PMID- 10874131 TI - Kinetics of human acetylcholinesterase inhibition by the novel experimental Alzheimer therapeutic agent, tolserine. AB - Characterization of the kinetic parameters of tolserine, a novel acetylcholinesterase (AChE) inhibitor of potential in the therapy of Alzheimer's disease, to inhibit purified human erythrocyte AChE was undertaken for the first time. An IC(50) value was estimated by three methods. Its mean value was found to be 8.13 nM, whereas the IC(100) was observed to be 25.5 nM as calculated by single graphical method. The Michaelis-Menten constant (K(m)) for the hydrolysis of the substrate acetylthiocholine iodide was found to be 0.08 mM. Dixon as well as Lineweaver-Burk plots and their secondary replots indicated that the nature of the inhibition was of the partial non-competitive type. The value of K(i) was estimated as 4.69 nM by the primary and secondary replots of the Dixon as well as secondary replots of the Lineweaver-Burk plot. Four new kinetic constants were also investigated by polynomial regression analysis of the relationship between the apparent K(i) (K(Iapp)) and substrate concentration, which may open new avenues for the kinetic study of the inhibition of several enzymes by a wide variety of inhibitors in vitro. Tolserine proved to be a highly potent inhibitor of human AChE compared to its structural analogues physostigmine and phenserine. PMID- 10874132 TI - Reductive activation of mitomycin C by neuronal nitric oxide synthase. AB - Mitomycin C (MC) requires bioreduction prior to the generation of alkylating moieties. NADPH-cytochrome P450 reductase is predominant in metabolic activation of MC in hypoxic cancer cells. In this study, neuronal nitric oxide synthase (nNOS), whose reductase domain is structurally similar to that of NADPH cytochrome P450 reductase, was assessed for its ability to activate MC. nNOS under anaerobic conditions catalyzed the reduction of MC, which was measured as the decrease in absorbance at 375 nm. Neither the heme blocker potassium cyanide (1 mM) nor the nNOS competitive inhibitor N(G)-nitro-L-arginine methyl ester (L NAME, 1 mM) affected the bioreduction of MC, whereas 0.1 mM diphenyleneiodonium chloride, which binds to the reductase domain of nNOS, inhibited MC reduction completely. The reduction of MC by nNOS was influenced by Ca(2+)/calmodulin. In the absence of Ca(2+)/calmodulin, the rate of MC reduction decreased by 28% at pH 6.6. The formation of an alkylated complex of 4-(p-nitrobenzyl)pyridine occurred in a manner analogous to that observed in MC metabolic experiments. The rate of MC reduction and the formation of the alkylated complex of 4-(p nitrobenzyl)pyridine at pH 6.6 were increased by 43 and 54%, respectively, as compared with that at pH 7.6. nNOS-activated MC resulted in the consumption of oxygen in air. The rate of oxygen consumption decreased by 50% in the presence of 2000 U/mL of catalase. MC inhibited nNOS activity in a noncompetitive manner. These findings demonstrate that nNOS is capable of catalyzing the bioreduction of MC. PMID- 10874133 TI - Inhibition of in vitro lymphoproliferation by three novel iron chelators of the pyridoxal and salicyl aldehyde hydrazone classes. AB - The capacity of three novel iron chelators, namely 1-[N ethoxycarbonylmethylpyridoxylidenium]-2-[2'-pyridyl]hydrazine bromide (EPH), 1 [5'-bromosalicylidene]-2-[2"-pyridyl]hydrazine (BsPH), and 1-pyridoxylidene-2-[1' phthalazyl]hydrazine dihydrochloride (PPhH), to inhibit the proliferation of mitogen-stimulated murine lymph node cells was examined in vitro. All three are of the aryl hydrazone class, the prototype of which is pyridoxal isonicotinoyl hydrazone. The chelators inhibited lymphoproliferation at low micromolar concentrations. EPH and PPhH had an inhibitory capacity comparable to that of desferrioxamine (IC(50): 3 and 2 microM, respectively), whereas BsPH was more potent (IC(50) < 1 microM). The inhibitory effects of the chelator were not due to cell cytotoxicity and could be abrogated by pretreating the chelator with iron. Time-course studies established a site of action for the chelators at the G(1)/S phase transition. These agents warrant further investigation for their potential as immunosuppressants. PMID- 10874135 TI - Inhibition by parthenolide of phorbol ester-induced transcriptional activation of inducible nitric oxide synthase gene in a human monocyte cell line THP-1. AB - Excessive nitric oxide production by inducible nitric oxide synthase (iNOS) in stimulated inflammatory cells is thought to be a causative factor of cellular injury in inflammatory disease states. Compounds inhibiting iNOS transcriptional activity in inflammatory cells are potentially anti-inflammatory. An assay method for estimating iNOS transcriptional activity in the human monocyte cell line THP 1 was established using a luciferase reporter gene system. In this study, we demonstrate that parthenolide, the predominant sesquiterpene lactone in European feverfew (Tanacetum parthenium), exerts potent inhibitory effects on the promoter activity of the iNOS gene in THP-1 cells. Parthenolide effectively suppressed iNOS promoter activity in a dose-dependent manner at concentrations higher than 2. 5 microM, with an IC(50) of about 10 microM. A tumor-promoting phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), significantly increased the iNOS promoter-dependent reporter gene activity, and the TPA-induced increase in iNOS promoter activity was effectively suppressed by parthenolide, with an IC(50) of approximately 2 microM. The present findings may further explain the anti inflammatory property of parthenolide. PMID- 10874134 TI - Roles of mitogen-activated protein kinase pathways for mediator release from human cultured mast cells. AB - Human cultured mast cells (HCMC) secrete histamine, sulfidoleukotrienes (LTs), and prostaglandin D(2) (PGD(2)), and produce a variety of cytokines after aggregation of high-affinity receptors for IgE (FcepsilonRI). With respect to the mitogen-activated protein kinase (MAPK) family, extracellular signal-regulated kinases (ERKs), c-Jun NH(2)-terminal kinases (JNKs), and p38 mitogen-activated protein kinase (p38 MAPK) are known. To investigate the roles of these kinase pathways for mediator release from human mast cells, we examined the participation of the activation of these kinases in mediator release, using 1,4 diamino-2, 3-dicyano-1,4-bis(2-aminophenylthio)butadiene (U0126), an ERK pathway inhibitor, and 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imid azo le (SB203580), a p38 MAPK pathway inhibitor. U0126 inhibited ERK activation, LT and PGD(2) release, and granulocyte macrophage-colony stimulating factor (GM CSF) production after stimulation of HCMC. SB203580, on the other hand, potentiated JNK activation and GM-CSF production. The findings of the present study demonstrated that: (i) the release of arachidonic acid metabolites is mediated by the ERK pathway; (ii) GM-CSF production may be driven by both the ERK and JNK pathways; and (iii) the p38 MAPK pathway negatively regulates the JNK pathway. This suggests that MAPK pathways play important roles in mediator release from human mast cells. PMID- 10874137 TI - In this issue ellipsis PMID- 10874136 TI - Protective action of cardiac DT-diaphorase against menadione toxicity in guinea pig isolated atria. AB - In myocardial preparations isolated from guinea pigs, 2-methyl-1, 4 naphthoquinone (menadione) causes an increase in contractility that is strictly related to the generation of reactive oxygen species (ROS) as a consequence of quinone metabolism. In heart, menadione undergoes one-electron reduction to semiquinone, a reaction mainly catalysed by mitochondrial NADH: ubiquinone oxidoreductase. It is also converted to hydroquinone by the soluble two-electron reductase, DT-diaphorase, and is conjugated with GSH by glutathione S transferase. In order to assess the role of DT-diaphorase in cardiac responses to menadione, we examined the effects of both a specific inhibitor (dicoumarol) and an inducer (beta-naphthoflavone) of the enzyme on the inotropic action of the quinone. In electrically driven left atria of guinea pig, 4 microM dicoumarol significantly enhanced the positive inotropic effect of menadione, especially at the lower concentrations of the quinone. In myocardial preparations isolated from guinea pigs treated with beta-naphthoflavone (80 mg/kg i.p.for 2 days), DT diaphorase activity was enhanced (+36% with respect to control animals, P < 0. 01), whereas the activities of the other enzymes involved in menadione metabolism were not modified. In these preparations, menadione caused a significantly lower increase in the force of contraction than in atria from untreated animals; moreover, pretreatment with beta-naphthoflavone caused a significant decrease in the menadione-induced oxidative stress, as evaluated from the GSH redox index. Taken together, these results demonstrate that cardiac DT-diaphorase does not contribute to ROS generation, but represents a detoxification system. PMID- 10874138 TI - Technique for neurosurgically relevant CT image transfers using inexpensive video digital technology. AB - OBJECTIVE: To examine and document neurosurgically relevant CT image transfers using inexpensive video digital technology. METHODS: Thirty abnormal CT scans were evaluated on a personal computer monitor following their digital image creation and transfer by electronic mail (e-mail). These were compared with the radiologist's interpretation of a hard copy of the CT scan originals. Any change in diagnosis based on the CT scan or e-mail image was assessed after completion of the comparison. RESULTS: A total of 30 CT scans were successfully transferred and reviewed. On only one image was there disagreement between the neurosurgeon's and the radiologist's interpretations of the image. This resulted in a change in the radiologist's diagnosis after digital image transfer occurred and the neurosurgeon diagnosed an isodense subdural hematoma, which was later confirmed at the time of surgical decompression. CONCLUSIONS: A hand-held, inexpensive digital camera may serve neurosurgeons as a helpful alternative to expensive, labor-intensive teleradiology systems. It should be considered as an adjunctive option for small community-based hospitals unable to financially support more sophisticated teleradiology techniques, which have been shown to provide a significant benefit in the management and outcome of head trauma patients. PMID- 10874139 TI - Preliminary experience with anterior cervical microdiscectomy and interbody titanium cage fusion (Novus CT-Ti) in patients with cervical disc disease. AB - BACKGROUND: Although the use of intervertebral fusion after anterior microdiscectomy in cervical disc disease remains controversial, a new surgical device is proposed for use in intervertebral fusion instead of bone graft. METHODS: This retrospective study at the Department of Neurosurgery, Cardarelli Hospital, Naples, from January 1993 to December 1998, compares the results of surgery on 58 patients with anterior microdiscectomy and intervertebral bone graft fusion (Group A) (ADIBG) with a group of 52 patients who underwent anterior microdiscectomy and intervertebral titanium cage fusion (Group B) (ADITC) in cervical radiculopathy and spondylotic myelopathy. In both groups a "radical discectomy" was performed under the operating microscope. In group A, interbody fusion was performed with autologous tricortical bone graft. In group B, a new type of titanium device (Novus CT-Ti) was used (Sofamor Danek Group). RESULTS: There was no collapse or extrusion of the device and no complications at the donor site (the bone fragments used to fill the cage were taken from osteophytes or vertebral body fragments). The use of this device provides immediate stabilization, reduces or eliminates pain, promotes bone fusion between the vertebrae adjacent to the cage by allowing bone growth through the cage, reestablishes and maintains the intervertebral space, reduces the average hospitalization time, and allows a quicker return to work. CONCLUSIONS: Patients who underwent ADITC did well and benefited from the surgery. Those who underwent ADITC did better than those who underwent ADIBG in regard to function, relief from pain, and complications. Early and good stability of the cervical spine seems to be the main advantage of using titanium cages. PMID- 10874140 TI - Simultaneous cervical diffuse idiopathic skeletal hyperostosis and ossification of the posterior longitudinal ligament resulting in dysphagia or myelopathy in two geriatric North Americans. AB - BACKGROUND: Cervical diffuse idiopathic skeletal hyperostosis (DISH) and ossification of the posterior longitudinal ligament (OPLL) rarely coexist in the North American population. Here, different surgical strategies were used to manage simultaneous DISH and OPLL resulting in dysphagia or myelopathy in two geriatric patients. METHODS: A 74-year-old male with esophageal compression and dysphagia attributed to DISH, and cord compression with myelopathy due to OPLL, was treated with a cervical laminectomy followed by anterior DISH resection. On the other hand, an 80-year-old male with asymptomatic DISH but moderate myelopathy (Nurick Grade III) secondary to OPLL required only a cervical laminectomy. RESULTS: In the first patient, dysphagia resolved within 3 months of surgery, while in the second individual, myelopathy improved to Nurick Grade I (mild myelopathy) within 6 months postoperatively. Improvement in both patients was maintained 1 year after surgery. CONCLUSIONS: While DISH and OPLL may coexist in geriatric patients, only those with dysphagia should undergo DISH resection, while others demonstrating myelopathy should have laminectomy alone. PMID- 10874141 TI - Stereotactic biopsy for brain tumors: is it always necessary? AB - BACKGROUND: Stereotactic biopsy is currently being used in oncological neurosurgery despite its limitations. The purpose of this study is to compare its diagnostic reliability with that of the diagnosis based on clinical data and neuroimaging techniques. METHODS: We studied 200 patients (134 men and 66 women) who underwent 212 stereotactic biopsy procedures to assess brain tumors. All were subjected to CT scan and 71 patients also underwent MRI. A presumptive diagnosis of brain tumor was established in each case and the findings compared with the results of stereotactic biopsy and the clinical course. RESULTS: A clear presumptive diagnosis was established before stereotactic biopsy in 90% of the patients, and in 95% of this group, the diagnosis was confirmed by the biopsy and/or the clinical course. The stereotactic biopsy modified the presumptive diagnosis in eight patients (4%), without changing the clinical diagnosis of malignant lesion. In our present series, 10% of the procedures were of no help in determining a diagnosis and in 7% of patients, the stereotactic biopsy led to diagnostic errors that influenced the therapeutic management and resulted in an erroneous prognosis. CONCLUSIONS: These data confirm the need for clinical correlation with the findings obtained at stereotactic biopsy and suggest that stereotactic biopsy may not always be useful or necessary in the management of brain tumor patients. According to our present critical analysis, when a clear presumptive diagnosis of brain tumor can be made, it may be sufficient to base the management of the patient only on clinical and neuroimaging findings. PMID- 10874142 TI - Meningiomas of the cerebellopontine angle. AB - BACKGROUND: Meningiomas of the cerebellopontine angle (CPA), although uniform in location, are diverse with regard to the site of dural origin and displacement of neurovascular structures. A study of patients with CPA meningiomas was undertaken to gain more information regarding the relationship between site of dural attachment, clinical presentation, operative approach, and outcome. METHODS: In this report, we retrospectively review 40 patients with CPA meningiomas managed surgically. RESULTS: Common clinical presentations were hearing loss, unsteadiness, and dysequilibrium. Findings upon physical examination included hearing loss (73%), cerebellar signs (32%), trigeminal neuropathy (16%), and facial nerve dysfunction (16%). The most common site of dural origin was the petrous ridge (anterior to the IAC [26%], posterior [21%], superior [18%], and inferior [16%]). Less common sites of dural origin included the tentorium (31%), the clivus (15%), the IAC (10%), and the jugular foramen (8%). Site of dural origin determined the direction of displacement of the facial/vestibulocochlear nerve bundle. The most common microsurgical complication was facial nerve dysfunction (30%). Gross total resection was achieved in 82% of cases, whereas 18% underwent subtotal resection. Two patients died. Follow-up ranged from three months to 13 years with three recurrences. CONCLUSIONS: CPA meningiomas displace the seventh and eighth cranial nerves in various directions depending on the site of dural origin. Total surgical excision can be accomplished in the majority of cases with acceptable morbidity. PMID- 10874143 TI - Primary intracranial germinoma in the medulla oblongata. AB - BACKGROUND: Primary intracranial germ cell tumor in the medulla oblongata is very rare; only five cases, including our case, have been reported. CASE REPORT: Our patient, an 18-year-old woman, was diagnosed with a primary intracranial germinoma in the medulla oblongata by an open biopsy. She was treated successfully with chemotherapy and radiosurgery. CONCLUSION: All five tumors in this site were histologically diagnosed as germinomas. The finding of female predominance in germ cell tumors in this region is shown. PMID- 10874144 TI - Rosai-Dorfman disease mimicking parasagittal meningioma: case presentation and review of literature. AB - BACKGROUND: Sinus histiocytosis with massive lymphadenopathy was originally described by Rosai and Dorfman in 1969. It usually presents with bilateral painless cervical lymphadenopathy. In extremely rare circumstances, the CNS can be affected. Only 21 prior cases of intracranial involvement have been reported. CASE REPORT: A 33-year-old white male presented with a 2-week history of progressive cephalgia. The patient underwent MRI testing that revealed an enhancing mass in the right parasagittal region with associated edema. Preoperative diagnosis was right parasagittal meningioma. The patient underwent craniotomy with complete resection of the mass. Histopathology was compatible with Rosai-Dorfman disease (RDD). CONCLUSION: Rosai-Dorfman disease is rarely found intracranially; however, its ability to mimic meningioma as well as other pathologies underlines its importance. With so few reported cases of intracranial involvement, more experience will be necessary before this clinical presentation and prognosis can be clearly outlined. PMID- 10874145 TI - The role of interstitial BCNU chemotherapy in the treatment of malignant glioma. AB - BACKGROUND: Use of interstitial BCNU wafers in the treatment of malignant glioma is currently a controversial topic among neurosurgeons. Initial clinical studies indicated implantation of BCNU wafers into the postoperative tumor bed to be an acceptably safe, partially effective treatment for glioblastoma multiforme. Yet a more recent study has put the efficacy of this treatment in doubt, and there are potential complications associated with BCNU wafer use. OBJECTIVE: This article presents a review of the information presently available on BCNU wafers-both pro and con-to aid in the clinical decision-making process. The article focuses on studies of clinical efficacy (for initial use as well as in the setting of recurrent tumor), complications associated with BCNU wafers, and the experimental data, particularly related to BCNU penetration into the brain. RESULTS: Animal studies and computer simulations have shown that the depth of penetration of BCNU from wafers is limited. Yet in actual clinical use, the interstitial pressure within the wafer-laden tumor bed might be higher, convective flow greater, and delivery of BCNU to the brain more significant than predicted. CONCLUSION: Based on current information, use of interstitial BCNU wafers continues to be an option for treating malignant glioma. Additional clinical studies of BCNU wafers are currently underway. PMID- 10874146 TI - Complicated stent supported cerebrovascular angioplasty: case analyses and review of literature. AB - BACKGROUND: Hemodynamic lesions of the cervicocerebral vasculature are currently being treated with stent supported percutaneous transluminal angioplasty. These procedures have met with increasing success when compared to the risks and morbidity of more invasive surgical approaches. The versatility of stent supported angioplasty as a primary therapeutic modality is examined in the following complex cases. CASE DESCRIPTION: We present four cases involving cervical angioplasty with emergent or adjunctive stent placement. Two cases involved the subclavian arteries, whereas the others involved the vertebral and internal carotid arteries. In our experience, complications of cervicocerebral artery angioplasty have been successfully managed by stent placement. CONCLUSION: Our cases demonstrate the emerging role of cervical angioplasty and stent implantation as a successful therapeutic modality, highlighted in these complex cases. PMID- 10874147 TI - De novo lesions in familial form of cerebral cavernous malformations: clinical and MR features in 29 non-Hispanic families. AB - BACKGROUND: To evaluate clinical and MR features of de novo lesions (DNL) in the familial form of cerebral cavernous malformation (CCM) in 40 patients belonging to 29 unrelated non-Hispanic families. METHODS: Forty patients followed up by serial cerebral MR examinations were included in this retrospective study. First and last available MR examinations were retrospectively analyzed and compared for each patient to diagnose DNL. Gradient-echo (GRE) sequences were performed in only 11 of the 40 patients and were not considered for this study. Incidence of DNL was evaluated in terms of lesions/patient-year. All DNL were characterized by their clinical and MR features (location, size, type). Type of CCM was determined according to the classification of Zabramski (1994). Patient groups with and without DNL were compared for sex, age, number of pre-existing CCMs, and follow up. RESULTS: Twenty-three DNL were recorded in 11 patients (27.5%) and the incidence was 0.2 lesions/patient-year (mean follow-up = 3.2 years). All but one DNL were asymptomatic. Twenty DNL were supratentorial and three were infratentorial. Mean diameter was 8 mm (2-35 mm). Six DNL were classified as type 1 (subacute hemorrhage), six as type 2 (hemorrhages and thromboses of varying ages) and 11 as type 3 (chronic hemorrhage with hemosiderin staining). No statistical difference between groups was found in terms of sex, age, or number of pre-existing CCMs. On the other hand, duration of follow-up was significantly longer in the group with DNL. CONCLUSION: The occurrence of DNL seems to be a hallmark of the familial form of CCM in non-Hispanic families as well as in Hispanic families. Such DNL are usually asymptomatic and are mainly classified as type 3 (chronic hemorrhage with hemosiderin staining). Within the limits of the retrospective study design and potential selection bias introduced by the varying indications for MR scanning, it does seem that DNL may occur at any time in the lifespan of CCM patients, and occurrence does not seem to depend on age, sex, or the total number of pre-existing lesions. PMID- 10874148 TI - De novo cavernoma case report and review of literature. AB - BACKGROUND: De novo cavernoma, reported with the familial form of disease, is rare in cases with a negative family history. Cranial radiation, coexistent vascular malformation, genetic and hormonal factors, previous surgery for intracranial lesions, or other apparently unrelated intracranial lesions have been reported as risk factors. METHODS: We report a case of de novo cavernoma without a family history and without previous irradiation or any other known risk factors. The genesis of this lesion is discussed. RESULTS: To our knowledge, this is the first case, based on two separate magnetic resonance imaging (MRI) studies, demonstrating evidence of de novo cavernous malformations in the absence of familial history, brain radiation therapy, or other apparently unrelated intracranial tissue lesions. Based on previous negative computed tomography scans, other cases have been presented as de novo cavernous angiomas; thus it is possible that the newly discovered cavernoma existed previously but had been missed on previous poorer quality or lower resolution imaging studies. CONCLUSIONS: Cavernoma can arise even without an associat family history; in our case, a previous head injury could have set off either a genetic cascade with attendant endothelial proliferation or a latent virus. PMID- 10874149 TI - Catecholamine-secreting carotid body tumor and intracranial aneurysm: coincidence? AB - BACKGROUND: An extremely rare case of intracranial aneurysm associated with catecholamine-secreting carotid body tumor is presented. CASE DESCRIPTION: A 64 year-old woman suffering from hypertensive attacks was admitted first to the Otolaryngology Department with a neck swelling. Right common carotid angiography revealed a hypervascular mass at the carotid bifurcation. On the same angiogram a middle cerebral artery aneurysm was discovered incidentally and the patient was referred to the Neurosurgical Department. Because of her history the tumor was considered to be endocrinologically active and the patient underwent alpha- and beta-blockade to protect intraoperative cardiovascular instability. Despite all precautions, during the operation hypertensive crises developed and the aneurysm was clipped with difficulty. CONCLUSION: Perioperative management designed to avoid complications in treating this rare association is discussed. Although this is the first reported case of an intracranial aneurysm associated with a functional carotid body tumor, a possible etiopathogenesis of the relationship between the aneurysm and hypertensive attacks due to an acute catecholamine discharging tumor is presented. PMID- 10874150 TI - Hemifacial spasm caused by a contralateral vertebral artery: case report. AB - BACKGROUND: Hemifacial spasm is usually caused by compression of the facial nerve by ipsilateral blood vessels. Compression of the facial nerve root exit zone by a contralateral tortuous vertebral artery is very rare. METHODS: This 68-year-old woman presented with left-sided hemifacial spasm and was found to have compression of the left facial nerve by the tortuous vertebrobasilar artery, as revealed by magnetic resonance imaging and magnetic resonance angiography. Retromastoid craniectomy demonstrated compression of the left facial nerve root exit zone by the distal portion of the right vertebral artery. The vertebrobasilar junction and both vertebral arteries were moved laterally from the facial nerve and a muscle implant was interposed between the brainstem and the right vertebral artery. RESULTS: The patient has remained free of hemifacial spasm for a follow-up period of 27 months. CONCLUSIONS: Compression of the facial nerve by the contralateral tortuous vertebral artery may produce hemifacial spasm. A transposed large vessel can be secured by a sling technique or by interposing a soft implant between the brainstem and the vertebral artery. PMID- 10874151 TI - Acute complications following gamma knife radiosurgery are rare. AB - BACKGROUND: Gamma knife radiosurgery (GKR) is a safe and effective alternative to surgery for intracranial lesions. Most studies evaluating toxicity after GKR have concentrated on the delayed radiation effects. METHODS: We retrospectively reviewed 835 consecutive GKR cases for early (within 7 days) neurological complications or death. RESULTS: We identified a total of 18 patients (2.2%) who had a neurological event or death. Five (0.6%) patients developed new focal deficits, 12 (1.4%) patients experienced a seizure and there were three (0.4%) deaths. Two deaths were related to development of seizures and neurological deterioration. One death was caused by a respiratory arrest related to the patient's primary cancer. Of the five patients with neurological deficits, none had a persistent deficit. In two cases the neurological deficits were due to an increase in edema. Whether this occurred as a result of the gamma knife treatment or was the natural progression of the tumor is unclear. CONCLUSIONS: Complications after GKR are uncommon and the risk of a permanent deficit arising from an acute neurological event is exceedingly low. PMID- 10874152 TI - Selective vulnerability to radiation in the hippocampal dentate granule cells. AB - BACKGROUND: Radiation therapy is an effective approach in the treatment of highly radiosensitive brain tumors such as germinomas. However, recent studies have reported intellectual disturbances in patients who underwent whole-brain irradiation as children. We detected apoptosis in the infantile murine cerebrum after systemic X-ray irradiation. METHODS: Subjects were 100 ICR mice 4 weeks old, of which 90 were systemically exposed to 18 Gy X-rays (0.45 Gy/min); 10 each were decapitated and the cerebrums were removed 1, 3, 6, 9, 12, 18, 24, 48, and 72 hours after irradiation. Controls were 10 unirradiated mice. DNA fragmentation analysis was carried out by agarose gel electrophoresis, and morphological analysis was by the TUNEL method. RESULTS: According to agarose gel electrophoresis, the cerebral DNA ladders were detected only over 6 to 24 hr, peaking in 9 hr. Even at the peak, band intensity was nearly double that of the unirradiated normal thymus. According to the TUNEL analysis, radiation-induced apoptosis increased, with a peak at 9 hours, but decreased 24 hours after irradiation. Apoptotic cells were always localized exclusively in the hippocampal dentate granule cells. CONCLUSIONS: We found that vulnerability to radiation existed in the hippocampal dentate granule cells. Intellectual disturbances in patients who have undergone whole-brain irradiation may be caused by injury to the hippocampus. PMID- 10874153 TI - Neurosurgery of the future? PMID- 10874154 TI - Hypomethylation as a cause of homocysteine-induced cell damage in human cell lines. AB - Despite the growing evidence that plasma homocysteine is a cardiovascular risk factor, the mechanism behind the vascular injuries is still unknown. In the present study we have investigated the possible role of hypomethylation as a cause of homocysteine-induced cell damage in two human cell lines. A significant growth retardation was observed in HeLa cell cultures in the combined presence of homocysteine and adenosine, but first at concentrations of 250 micromol/l of each. A significant decrease of intracellular glutathione concentration was noted both in the presence of homocysteine (250 micromol/l) alone and in the presence of the combination of homocysteine and adenosine (250 micromol/l). Intracellular concentration of homocysteine was increased to a similar extent both in the presence of homocysteine alone and in the presence of a combination of homocysteine and adenosine. Similar findings to those described for HeLa cell cultures were observed in endothelial cell cultures. Furthermore, in the presence of copper ions together with 100 micromol/l of adenosine and homocysteine a significantly retarded cell growth was observed in HeLa cell cultures. This finding shows that a combination of two potentially cell-damaging mechanisms (formation of oxygen radicals and hypomethylation) aggravated the retardation of cell growth compared to only one of these mechanisms being present. Thus, it is likely that several mechanisms of homocysteine-induced cell damage contribute to the increased rate of the atherogenic process observed in hyperhomocysteinemia. PMID- 10874155 TI - Cytotoxic effects of cantharidin on the growth of normal and carcinoma cells. AB - Cantharidin is isolated from Mylabris phalerata Pallas and is a potent inhibitor of hepatocellular carcinoma cells (Hep 3B cells). In the present study, the IC(50) values of cantharidin on Hep 3B cells and normal Chang liver cells were found to be 2.2 and 30.2 microM for 36 h, respectively. Furthermore, cantharidin treated Hep 3B cells induced cell death within 1 h (IC(50)=52.8 microM), suggesting that cantharidin is an acute cytotoxic agent. We found that although cantharidin could induce cell death, it could not directly inhibit the activity of nucleic acid biosynthesis by the cellular incorporation of 3H-thymidine, 3H uridine or 3H-leucine. Cantharidin-treated Hep 3B cells showed no evidence of major alterations in the cell cycle distribution within 1 h. However, examination of cells after treatment for 36 h showed that cantharidin regulated the cell cycle at the G(2)/M phase. Moreover, the treated Hep 3B cells had a rounded and shrunken appearance. The microvilli of treated Hep 3B cells were reduced in number and replaced by numerous blebs. Other ultrastructural changes following cantharidin treatment included the presence of lipid droplets, swelling of the mitochondria and accumulation of glycogen particles. The findings of damaged mitochondria in the cantharidin treated Hep 3B cells in this study suggest that cantharidin can induce acute and lethal toxic effects on Hep 3B cells by inhibiting the mitochondria energy system. In conclusion, this study had demonstrated that cantharidin could inhibit progression of all phases of the Hep 3B cell cycle. PMID- 10874157 TI - The role of anion exchange in the uptake of Pb by human erythrocytes and Madin Darby canine kidney cells. AB - Anion exchange (AE) plays a critical role in regulating intracellular pH in erythrocytes and epithelial cells and has been suggested to facilitate the transport of lead (Pb) across the erythrocyte cell membrane. In this study we examined the role of AE in the uptake of Pb by human erythrocytes and by Madin Darby canine kidney (MDCK) cells, the kidney epithelial cell line. Functional AE in MDCK cells was evidenced by: increased uptake of SO(4)(2-) at pH 6.0 over pH 7.0, and inhibition of SO(4)(2-) uptake by the AE inhibitor 4, 4' diisothiocyanostilbene-2, 2'- disulfonic acid (DIDS) as well as by non-halide anions. Accumulation of Pb into MDCK cells was time and temperature dependent. DIDS inhibited uptake of Pb into human erythrocytes but not MDCK cells. In conclusion, uptake of Pb into erythrocytes but not kidney epithelial cells occurs through AE. PMID- 10874156 TI - A possible role of oxidative stress in the vanadium-induced cytotoxicity in the MC3T3E1 osteoblast and UMR106 osteosarcoma cell lines. AB - The cytotoxicity and free radical production induced by vanadium compounds were investigated in an osteoblast (MC3T3E1) and an osteosarcoma (UMR106) cell lines in culture. Vanadate induced cell toxicity, reactive oxygen species (ROS) formation and thiobarbituric acid reactive substances (TBARS) increased in a concentration-dependent manner (0.1-10 mM) after 4 h. The concentration-response curve of vanadate-induced cytotoxicity and oxidative stress in MC3T3E1 cells was shifted to the left of the UMR106 curve, suggesting a greater sensitivity of the non-transformed cells in comparison to the osteosarcoma UMR106 cells. Supplementing with vitamin E acetate (80 microM) significantly inhibited ROS and TBARS formation but did not improve the vanadate-dependent decrease in cell number. Other vanadium compounds (vanadyl, pervanadate, and VO/Aspi, a complex of vanadyl(IV) with aspirin) showed different degrees of cell toxicity and induced oxidative stress. Altogether these results suggest that oxidative stress is involved in vanadium induced osteoblastic cytotoxicity, although the mechanism is unknown. PMID- 10874158 TI - Potential health effects of drinking water disinfection by-products using quantitative structure toxicity relationship. AB - Disinfection by-products (DBPs) are produced as a result of disinfecting water using various treatment methods. Over the years, chlorine has remained the most popular disinfecting agent due to its ability to kill pathogens. However, in 1974, it was discovered that the superchlorination of drinking water resulted in the production of chloroform and other trihalomethanes. Since then hundreds of additional DBPs have been identified, including haloacetic acids and haloacetonitriles with very little or no toxicological data available, thus necessitating the use of additional methods for hazard estimation. Quantitative Structure Toxicity Relationship (QSTR) is one such method and utilizes a computer based technology to predict the toxicity of a chemical solely from its molecular attributes. The current research was conducted utilizing the TOPKAT/QSTR software package which is comprised of robust, cross-validated QSTR models for assessing mutagenicity, rodent carcinogenicity (female/male; rat/mouse), developmental toxicity, skin sensitization, lowest-observed-adverse-effect level (LOAEL), fathead minnow LC(50), rat oral LD(50) and Daphia magna EC(50). A total of 252 DBPs were analyzed for the likelihood that they would produce tumors and developmental effects using the carcinogenicity and developmental toxicity submodels of TOPKAT. The model predictions were evaluated to identify generalizations between the functional groups (e.g. alcohols, acids, etc.) and specific toxic endpoints. Developmental toxicity was identified as an endpoint common to the majority of aliphatic mono- and dicarboxylic acids, aliphatic halogenated and non-halogenated ketones, and aliphatic haloacetonitriles. In the case of the carcinogenicity submodels, most aliphatic aldehydes were identified as carcinogens only in the female mouse submodel. The majority of the aliphatic and aromatic dicarboxylic acids were identified as carcinogens in the female rat submodel. All other functional groups examined were largely predicted as non carcinogens in all the cancer submodels (i.e. male/female rats and mice). The QSTR results should aid in the prioritization for evaluation of toxic endpoints in the absence of in vivo bioassays. PMID- 10874159 TI - Comparison of respiratory responses to Metarhizium anisopliae extract using two different sensitization protocols. AB - Metarhizium anisopliae, an entomopathogenic fungus, is a prototypic microbial pesticide licensed for indoor control of cockroaches, a major source of allergens. We have previously demonstrated allergy and asthma-like responses in BALB/c mice intraperitoneally (IP) sensitized in the presence of adjuvant and intratracheally (IT) challenged with the soluble factors from M. anisopliae crude antigen (MACA) (Ward et al., 1998, 2000). This protocol has been used frequently to establish animal models of allergenicity. However, the sensitization protocol is artificial and not representative of an environmental exposure. Concern has been raised that this protocol might produce allergic responses that would not occur under normal environmental exposure conditions. The objective of this study was to compare responses in mice to MACA by two exposure protocols: (1) exclusive respiratory exposures without adjuvant (representative of environmental exposures) and (2) intraperitoneal sensitization in the presence of adjuvant followed by IT challenge (the traditional approach). The intratracheal protocol consisted of four IT exposures of 10 microg MACA in 50 microl HBSS each over a 4 week period. A vehicle control group of mice was exposed IT to HBSS. The intraperitoneal protocol consisted of IP sensitization with 25 microg MACA in 0.2 ml of 1.3% alhydrogel (aluminum hydroxide) followed 14 days later with an IT challenge (10 microg MACA/50 microl HBSS). Airway reactivity responsiveness to methacholine was assessed, serum and bronchoalveolar lavage fluid (BALF) samples were obtained, and the lungs were fixed for histopathology at 1, 3, and 8 days following the last MACA IT challenge. Both groups exhibited immune and pulmonary responses typical of allergic asthma. In general, local responses in the lung, including inflammatory responses (eosinophils, lymphocytes, and macrophages), BALF IgE, and functional responses to methacholine were greater in the IT sensitized group compared to the IP sensitized group, whereas the systemic IgE response was greater in the IP sensitized group. The BALF IL-5 cytokine levels were elevated before and throughout the eosinophil influx. IL-4 was detected in the BALF of IP sensitized, but not IT sensitized mice. Histopathologic changes in the two groups were similar in nature but more severe in the IT mice. The results suggest that the IP sensitization protocol does not induce the level of respiratory responsiveness that results from sensitization by a physiologically relevant route of exposure. Thus total serum IgE levels, which were greater following IP sensitization, may not be the best indicator of allergen potency, at least with respect to respiratory responses. PMID- 10874160 TI - Significance of egg-jelly substances in the internal fertilization of the newt, Cynops pyrrhogaster. AB - Japanese newt, Cynops pyrrhogaster, undergoes internal fertilization as do most urodeles. In this study, we focused on the roles of egg-jelly in fertilization of C. pyrrhogaster and characterized the substances associated with those roles. When dry sperm were directly inseminated onto the egg, normal fertilization occurred without the presence of water. Egg-jelly extract (JE) prepared with Steinberg's salt solution contained the activity for the initiation of sperm motility. A substance of about 50 kDa in JE was significant for this activity; an inactive form of the substance probably exists in JE. Strong activity to induce acrosome reaction was detected in JE. It was inhibited by the treatment of JE with WGA, suggesting that carbohydrate in JE may be important for the induction of the acrosome reaction. This study suggests that two significant processes of fertilization are regulated by substances in the egg-jelly of the newt, C. pyrrhogaster. PMID- 10874161 TI - Fertilization signalling and protein-tyrosine kinases. AB - Fertilization is initiated by species-specific gamete cell recognition, i.e. sperm-egg interaction, followed by a rapid and sustained activation of multiple cellular and biochemical events, collectively called 'egg activation', which is indispensable for successful formation of zygotic nucleus and later embryogenesis. It is well known that sperm-induced egg activation is mediated by a transient release of calcium ions that originates from the sperm entry point and propagates through the entire egg cytoplasm. It is unclear, however, what kind of upstream events prelude to the calcium transient after sperm-egg interaction. Recently, much attention has been paid to the role of protein tyrosine phosphorylation in egg activation process by a number of studies on some well-established model organisms. These includes marine invertebrates, frogs, and mammals. In this review, we will summarize the recent findings that begin to uncover a 'missing link' between sperm-egg interaction and egg activation with emphasis on the role of egg protein-tyrosine kinases (PTKs) in Xenopus egg fertilization. PMID- 10874162 TI - Maternal program of apoptosis activated shortly after midblastula transition by overexpression of S-adenosylmethionine decarboxylase in Xenopus early embryos. AB - When we studied polyamine metabolism in Xenopus embryos, we cloned the cDNA for Xenopus S-adenosylmethionine decarboxylase (SAMDC), which converts SAM (S adenosylmethionine), the methyl donor, into decarboxylated SAM (dcSAM), the aminopropyl donor, and microinjected its in vitro transcribed mRNA into Xenopus fertilized eggs. We found here that the mRNA injection induces a SAM deficient state in early embryos due to over-function of the overexpressed SAMDC, which in turn induces inhibition of protein synthesis. Such embryos developed quite normally until blastula stage, but stopped development at the early gastrula stage, due to induction of massive cell dissociation and cell autolysis, irrespective of the dosage and stage of the mRNA injection. We found that the dissociated cells were TUNEL-positive, contained fragmented nuclei with ladder forming DNA, and furthermore, rescued completely by coinjection of Bcl-2 mRNA. Thus, overexpression of SAMDC in Xenopus embryos appeared to switch on apoptotic program, probably via inhibition of protein synthesis. Here, we briefly review our results together with those reported from other laboratories. After discussing the general importance of this newly discovered apoptotic program, we propose that the maternal program of apoptosis serves as a surveillance mechanism to eliminate metabolically severely-damaged cells and functions as a 'fail-safe' mechanism for normal development in Xenopus embryos. PMID- 10874163 TI - Xenopus msx-1 regulates dorso-ventral axis formation by suppressing the expression of organizer genes. AB - We demonstrated previously that Xmsx-1 is involved in mesoderm patterning along the dorso-ventral axis, under the regulation of BMP-4 signaling. When Xmsx-1 RNA was injected into the dorsal blastomeres, a mass of muscle tissue formed instead of notochord. This activity was similar to that of Xwnt-8 reported previously. In this study, we investigated whether the activity of Xmsx-1 is related to the ventralizing signal and myogenesis promoting factor, Xwnt-8. Whole-mount in situ hybridization showed that Xmsx-1, Xwnt-8, and XmyoD were expressed in overlapping areas, including the ventro-lateral marginal zone at mid-gastrula stage. The expression of XmyoD was induced by the ectopic expression of either Xmsx-1 or Xwnt-8 in dorsal blastomeres, and Xwnt-8 was induced by the ectopic expression of Xmsx-1. On the other hand, the expression of Xmsx-1 was not affected by the loading of pCSKA-Xwnt-8 or dominant-negative Xwnt-8 (DN-Xwnt-8) RNA. In addition, Xmsx-1 RNA did not abrogate the formation of notochord if coinjected with DN-Xwnt 8 RNA. These results suggest that Xmsx-1 functions upstream of the Xwnt-8 signal. Furthermore, the antagonistic function of Xmsx-1 to the expression of organizer genes, such as Xlim-1 and goosecoid, was shown by in situ hybridization analysis and luciferase reporter assay using the goosecoid promoter construct. Finally if Xmsx-1/VP-16 fusion RNA, which was expected to function as a dominant-negative Xmsx-1, was injected into ventral blastomeres, a partial secondary axis formed in a significant number of embryos. In such embryos, the activity of luciferase, under the control of goosecoid promoter sequence, was significantly elevated at gastrula stage. These results led us to conclude that Xmsx-1 plays a central role in establishing dorso-ventral axis in gastrulating embryo, by suppressing the expression of organizer genes. PMID- 10874164 TI - In vitro control of organogenesis and body patterning by activin during early amphibian development. AB - In the process of amphibian development, an embryonic body plan is established through cell division, sequential gene expression, morphogenesis and cell differentiation. The mechanism of body patterning is complex and includes multiple induction events. Activin, a TGF-beta family protein, can induce several kinds of mesodermal and endodermal tissues in animal cap explants in a dose dependent manner. In a recent study of the role of activin in organogenesis, we succeeded in raising a beating heart by treating animal caps with a high concentration of activin. Activin also participates in kidney organogenesis in combination with retinoic acid. An embryonic kidney induced by activin and retinoic acid in vitro can function in vivo when it is transplanted into a larva in which pronephros rudiments have already been removed. Further, the activin treated animal caps clearly show organizer actions that are closely related to body patterning along the anteroposterior axis. These experiments will help to serve as a model system for understanding organogenesis and body patterning at the cellular and molecular levels. PMID- 10874165 TI - Work in progress: the renaissance in amphibian embryology. AB - Various historical eras in the distant as well as the recent past of amphibian embryology are briefly reviewed. The concepts which emerged from the early years matured, then were laid to rest for several decades. A resurgence, driven by key discoveries with peptide growth factors, and fueled by modern molecular biology methods, is underway. The future for several amphibian research projects should be promising since interest in basic concepts remains strong, and application of frontier methodologies is yielding novel findings. PMID- 10874166 TI - Comparative study of the molecular mechanisms of oocyte maturation in amphibians. AB - Maturation-promoting factor (MPF), a complex of Cdc2 and cyclin B, is the final inducer of oocyte maturation. Its activity is controlled by inhibitory phosphorylation of Cdc2 on Tyr15/Thr14 and activating phosphorylation on Thr161. Full-grown immature oocytes of the African clawed frog Xenopus laevis contain inactive MPF (pre-MPF) that comprises cyclin B-bound Cdc2 phosphorylated on Tyr15/Thr14 and Thr161. The synthesis of Mos, but not cyclin B, after stimulation by the maturation-inducing steroid progesterone, is believed to be necessary for initiating Xenopus oocyte maturation through Tyr15/Thr14 dephosphorylation of pre MPF. In contrast, amphibians other than Xenopus (and also fishes) employ a different mechanism. Full-grown immature oocytes of these species contain monomeric Cdc2 but not cyclin B. MPF is formed after hormonal stimulation by binding of the newly produced cyclin B to the pre-existing Cdc2 and is immediately activated through Thr161 phosphorylation. Mos/MAP kinase is neither necessary nor sufficient for initiating maturation in fishes and amphibians except for Xenopus. We propose a new model of MPF formation and activation during oocyte maturation that is applicable to all amphibians (as well as fishes), based on a novel concept that pre-MPF is an artificial molecule that is not essential for inducing oocyte maturation. PMID- 10874167 TI - Dual functions of thyroid hormone receptors during Xenopus development. AB - Thyroid hormone (TH) plays a causative role in anuran metamorphosis. This effect is presumed to be manifested through the regulation of gene expression by TH receptors (TRs). TRs can act as both activators and repressors of a TH-inducible gene depending upon the presence and absence of TH, respectively. We have been investigating the roles of TRs during Xenopus laevis development, including premetamorphic and metamorphosing stages. In this review, we summarize some of the studies on the TRs by others and us. These studies reveal that TRs have dual functions in frog development as reflected in the following two aspects. First, TRs function initially as repressors of TH-inducible genes in premetamorphic tadpoles to prevent precocious metamorphosis, thus ensuring a proper period of tadpole growth, and later as activators of these genes to activate the metamorphic process. Second, TRs can promote both cell proliferation and apoptosis during metamorphosis, depending upon the cell type in which they are expressed. PMID- 10874168 TI - In vivo evidence for the production of sulfated steroids in the frog brain. AB - It is well established that sulfated neurosteroids are potent regulators of neuronal activity but the biosynthesis of sulfate esters of steroids in the central nervous system (CNS) has received little attention. In particular, the localization of hydroxysteroid sulfotransferase (HST), the enzyme which is responsible for the formation of sulfated steroids, has never been determined in the brain. We took advantage of the availability of an antiserum raised against rat liver HST to investigate the distribution of this enzyme in the CNS of the frog Rana ridibunda. Two populations of HST-positive neurons were localized in the anterior preoptic area and the magnocellular nucleus of the hypothalamus. Numerous HST-immunoreactive fibers were visualized throughout the telencephalon and the diencephalon. Reversed-phase high performance liquid chromatography (HPLC) analysis of frog telencephalon and hypothalamus extracts combined with radioimmunoasssay (RIA) detection showed the presence of substantial amounts of DHEAS-immunoreactive material which coeluted with synthetic DHEAS. The concentrations of DHEAS detected in the telencephalon and hypothalamus were respectively eight and five times higher than in the serum. The present study demonstrates the occurrence of HST-immunoreactive material in neurons of the frog telencephalon and diencephalon. This report also provides evidence for the presence of HST bioactivity, in vivo, in the frog brain. PMID- 10874169 TI - Hormonal control of urodele reproductive behavior. AB - Hormonal control of expression of courtship behavior and of development of structures related to the reproductive behavior in two species of Japanese newts, Cynops pyrrhogaster and Cynops ensicauda, was described. Prolactin (PRL) and androgen were essential factors for eliciting courtship behavior. In addition, arginine vasotocin markedly enhanced the expression of courtship behavior. PRL induced migration to water, in which courtship and oviposition take place, and converted the integument from the terrestrial type to the aquatic one. PRL also stimulated the growth of the tail fin, which was blocked by estrogen. Cellular and nuclear size and number of synapses on the somata of Mauthner cells, which are involved in tail movement, were also increased by PRL and androgen. Synthesis of sodefrin, a female-attracting pheromone, in the abdominal gland as well as that of mucopolysaccharides constituting the sac of sperm in the lateral gland was enhanced by PRL and androgen. Structural development of oviducts was elicited by estrogen or PRL to a certain extent, and full oviducal development by the combination of these two hormones, PRL being indispensable for the oviducal jelly secretion. PMID- 10874170 TI - DDT congener effects on secondary sex coloration in the reed frog Hyperolius argus: a partial evaluation of the Hyperolius argus endocrine screen. AB - Organochlorine compounds such as o,p'DDT can mimic estrogen effects. We compared the effects of o,p'DDT and six other DDT congeners to the effects of estradiol by comparing in vivo color changes in the reed frog (Hyperolius argus). Premature female color pattern induction in H. argus is specific to estrogens and the current study suggests that this assay has potential for use in discriminating between xenobiotic estrogens and non-estrogens. Animals were treated at forelimb emergence and maintained in treated solution until final evaluation. Estradiol, o,p.DDT (0.1 microg/ml), o,p'DDE (1 microg/ml) and o,p'DDD (1 microg/ml) prematurely induced adult female coloration patterns in juvenile animals, whereas p,p'DDT, p, p'DDE and p,p'DDD did not. PMID- 10874172 TI - Batrachology of Japan and adjacent regions - a systematic review. AB - As is the case with most other organisms, studies of amphibians started with systematics as the basis for many other biological fields, but systematics itself has endured a slower progress than other fields of biology have enjoyed for a long time. Nevertheless, philosophical and methodological 'revolutions' in the past 30 years, combined with more recent progress in biochemical and molecular techniques, have been forcing current systematics into a great change. Stejneger's (1907) monumental book, the first thorough review of amphibians from Japan and adjacent regions, was published nearly a century ago, and since then, the increase in number of areas that have been closely surveyed, together with the application of new theories and methods, have greatly enlarged our knowledge of the amphibian fauna in this region. For example, extensive experiments by the Hiroshima School using artificial hybridization techniques greatly contributed to our understandings of relationships among some groups of amphibians that had been taxonomically confusing. In addition to this, new species discovered by more detailed field surveys in a wider area and cryptic species revealed by analyses of isozymes and acoustic properties have made for a great increment in the number of taxa in this region. Further, analyses of DNA sequences now enable us to infer the affinities of taxa with few phylogenetically informative phenotypic properties, and the resultant phylogenetic hypotheses further contribute to our understandings of biogeography and many other issues on amphibian evolution. From these advances, it has been clarified that the amphibian species diversity is far greater than was previously expected, which in turn means a high endemism within particular regions. It is expected that future studies will also reveal exact systematic relationships of wide-ranging species, both those occurring within this region and those extending outside of it. For that purpose, international and interdisciplinary studies are indispensable. But what is also necessary is to conserve amphibian populations so that they will not disappear before their true relationships are realized. Knowledge of systematics among young amphibian biologists is also a matter of high priority. PMID- 10874171 TI - Hormonal influence on the olfactory response to a female-attracting pheromone, sodefrin, in the newt, Cynops pyrrhogaster. AB - Sodefrin is a female-attracting pheromone isolated from the abdominal glands of male newts, Cynops pyrrhogaster. Previously, the preference of conspecific female newts for sodefrin was shown to be completely abolished by plugging the bilateral nostrils, indicating that it acts on the olfactory organ. To determine the sensitivity of the olfactory receptor cells to sodefrin, electro-olfactograms (EOGs) in response to sodefrin solution were recorded from the ventral nasal epithelium of sexually developed female newts. Sodefrin elicited marked EOG responses in a dose-dependent manner on the epithelium of the lateral nasal sinus (LNS) region, a putative vomeronasal organ. In ovariectomized females, treatment with prolactin (PRL) and estrogen markedly enhanced the EOG response to sodefrin. The EOG response to the pheromone was also enhanced considerably by treatment with either PRL or estrogen alone. A slight but significant elevation was observed in castrated males receiving PRL plus estrogen or estrogen alone. It was concluded that the main site of action of sodefrin resides in the lateral sinus region and that sensitivity to sodefrin is under the control of PRL and estrogen. The presence of a sex difference in olfactory sensitivity to the hormones and/or pheromone was also suggested. PMID- 10874173 TI - Sex-linked genes and linkage maps in amphibians. AB - This paper reviews sex-linked genes and linkage maps in amphibians. It appears that there is no common ancestral or conserved sex-linkage group in amphibians, whereas an important proportion of other linkage groups has been conserved in amphibians. Comparisons of amphibian linkage maps with those of fishes and mammals reveal several syntenic associations apparently conserved over a very long period of vertebrate divergence. PMID- 10874174 TI - Clinical management of the patient with pleural effusion. AB - Pleural effusion is a frequent medical problem with a wide span of different causes. We wish to highlight the clinical management of the patient with pleural effusions but anatomic, physiologic and diagnostic management of the main pleural diseases will also be considered. PMID- 10874175 TI - Benign pleural diseases. AB - The pleural space is a potential space under normal physiologic circumstances. It envelops the lung, the mediastinum, the diaphragm and the chest wall. A thin film of pleural fluid provides lubrication for the two pleural layers; only 2-10 ml of pleural fluid is present in healthy people. For the purposes of this review, pleural abnormalities will be divided into pleural effusion, pneumothorax, and pleural calcification. PMID- 10874176 TI - Malignant pleural disease. AB - The vast majority of pleural neoplasms invade the pleura secondarily and can be seen in patients with bronchogenic carcinoma, breast cancer, lymphoma, and ovarian or gastric carcinoma. Primary pleural neoplasms are less common, although they have developed notoriety since the up-surge of malignant mesothelioma and the knowledge of its connection to asbestos exposure. Other malignant primary tumors include localized fibrous tumor and pleural liposarcoma. In most patients with diffuse malignant pleural disease the chest radiograph shows pleural effusion with or without pleural thickening. Computed tomography (CT) usually provides precise localization and extent of the disease and may be of value in assessing chest wall and mediastinal involvement. In specific situations, magnetic resonance (MR) may be useful as a problem-solving tool when CT findings of chest wall or diaphragmatic invasion are equivocal or in patients with contraindication to intravenous administration of ionic contrast material. PMID- 10874177 TI - Intervention in the pleura. PMID- 10874178 TI - Interobserver agreement in the diagnosis of pulmonary embolism with helical CT. AB - PURPOSE: Our objective was to asses the interobserver agreement in the detection of pulmonary embolism (PE) with contrast-enhanced helical CT at the main pulmonary, lobar and segmental arteries. A prospective study was carried out in 51 patients with suspected PE. Finally, 29 patients were diagnosed of PE. SUBJECTS AND METHODS: All patients were studied with helical CT. Images (5 mm collimation, 1.5 pitch factor, 3 mm reconstruction interval) were obtained after bolus contrast injection (120 ml, 4 ml/s, 15 s delay time). All cases were blinded and independently interpreted in three ways: two radiologists with different level of expertise and two expert radiologists reading by consensus. Agreement was evaluated by means of the kappa test. RESULTS: Kappa values for thrombi detection expressed an excellent agreement at the main (between 0.802 and 0. 946), lobar (between 0.915 and 0.958) and segmental (between 0.879 and 0.718) levels. For all vessels, mean kappa values were similar and excellent for all three combinations of readers. Arteries with more discrepancies were located mainly at the anterior and posterior areas of the upper lobes. CONCLUSIONS: The high degree of agreement shown in this study indicates that helical CT is a reproducible test in the diagnosis of PE to the segmental level. Isolated readings and levels of expertise do not influence agreement. PMID- 10874191 TI - Comparison of muscle sizes and moment arms of two rotator cuff muscles measured by ultrasonography and magnetic resonance imaging. AB - OBJECTIVE: The purpose was to investigate ultrasound (US) and anthropometry (AN) as valid alternatives to magnetic resonance imaging (MRI) regarding muscle size characteristics of two rotator cuff muscles. METHODS: Eight healthy females (age 27-54 yrs.) went through MRI and US scannings and AN measurements, where muscle thickness, cross-section area (CSA), moment arm, muscle length and width were measured on supraspinatus and infraspinatus muscle. RESULTS: The agreement between the methods was very satisfactory for CSA, and satisfactory for muscle thickness, moment arm, muscle length and width, with a mean difference below 2 mm in thickness and below 5 mm in muscle length and width. Volume could be estimated satisfactory in supraspinatus muscle, but not in infraspinatus muscle, where volume had to be calculated from thickness, length and width. As a significant relation was found in the MRI measurements between thickness and CSA, thickness measurements may replace CSA in inaccessible muscles. CONCLUSIONS: US was a valid method in measuring CSA, muscle thickness and moment arm. Combined with anthropometric measures of muscle length and width, volume can be calculated, which is important when defining the physiological cross-sectional area and muscle function. Further development and validation of the method is needed, however. PMID- 10874179 TI - A case of Marchiafava-Bignami disease: MRI findings on spin-echo and fluid attenuated inversion recovery (FLAIR) images. AB - Marchiafava-Bignami disease (MBD) was diagnosed in a 56-year-old man. Spin-echo (SE) magnetic resonance imaging (MRI) at the acute phase showed normal signal areas in the central layer of the corpus callosum (CC), although the intensity of these areas revealed abnormal hyperintensity on fluid attenuated inversion recovery (FLAIR). On follow-up SE MRI at the late phase, the central layer of the CC showed fluid-like intensity. On FLAIR MRI, the lesions of the CC turned into hypointense cores surrounded by hyperintense rims indicating central necrosis and peripheral demyelination. Degenerative changes of the CC in MBD were clearly demonstrated by FLAIR MRI. PMID- 10874192 TI - Duplex ultrasonography in the diagnosis of incompetent Cockett veins. AB - OBJECTIVE: Incompetent perforating veins of the medial calf, i.e. those of the Cockett groups, play a major role in the developement of chronic venous insufficiency. The aim of the present study was to test the value of duplex ultrasonography (DUS) in the diagnosis of function and localisation of those veins. METHODS: Eighty-nine legs with incompetent perforating veins of the medial calf selected for subfascial endoscopic perforator surgery (SEPS) were included in a prospective study. Preoperative DUS was used to determine the number and localisation of the perforator veins. Findings were compared with preoperative ascending phlebography and intraoperative endoscopy during SEPS. RESULTS: Nearly equal numbers of insufficient Cockett veins at each level were detected by DUS and ascending phlebography (Cockett III: n, 76 vs. n, 76, P, 1.0; Cockett: II n=84 vs. n=82, P=0.569; Cockett I: n, 36 vs. n, 37, P=1.0; chi(2)-test). Findings were confirmed intraoperatively. CONCLUSION: The accuracy of DUS is comparable to phlebography for the diagnosis of incompetent perforating veins of the lower leg. DUS is non-invasive and avoids the potential risks of radiologic imaging. PMID- 10874193 TI - Interstitial laser photocoagulation under ultrasound guidance of liver tumors: results in 104 treated patients. AB - OBJECTIVE: To evaluate the efficacy and complications of interstitial laser photocoagulation (ILP) under ultrasound (US) guidance as a technique for focal ablation of liver tumors in patients with normal and impaired hepatic function. PATIENTS AND METHODS: A total of 104 patients, 77 with 85 nodules of hepatocellular carcinoma on cirrhosis (29 in Child-Pugh A class, 43 in B e 5 in C class) and 27 patients with hepatic metastases (25 from colon, two from lung carcinoma) underwent ILP under US guidance. Depending on tumor size up to four needles were inserted in the tumor and multiple laser illuminations were performed in one or multiple sessions. Necrosis of the nodules was evaluated with triphasic contrast-enhanced CT. RESULTS: Ninety-four patients underwent a single ILP session and nine patients two sessions. CT showed complete necrosis in 70 out of 85 HCC nodules in 65 treated patients and in 24 out of 31 patients with metastases. Three Child C class patients dropped out the control of efficacy by CT because of severe liver failure associated in one case with transient paralytic ileum. One of these patients died 2 months after treatment. Two patients with metastasis dropped the completion of the treatment because of complication occurred after the ILP session (one paralytic ileum, one gastric haemorrage). CONCLUSIONS: ILP under US guidance is effective in inducing complete necrosis in small and large liver tumors. Nevertheless, ILP can cause severe derangement of liver function in patients with advanced cirrhosis. PMID- 10874194 TI - Dyspepsia in AIDS is correlated to ultrasonographic changes of antral distension. AB - OBJECTIVES: Patients with acquired immune deficiency syndrome (AIDS) frequently complain about dyspeptic symptoms. We set out to test whether changes in antral emptying or antral distension may account for these dyspeptic symptoms in AIDS. METHODS: We studied antral emptying in ten patients with HIV infection (CDC 1993 classification stage C) by means of an established real-time ultrasonographic method. Organic abdominal lesions had been excluded. Six upper gastrointestinal symptoms were evaluated using a score ranging from 0 to 3. Fifteen subjects without any abdominal complaints and without any abdominal history served as controls. Antral cross sectional area was measured after an overnight fast and at 0, 15, 30, 45, 60, 90, 120 min after an semisolid test meal. Antral postprandial distension was expressed using an antral expansion ratio (postcibal antral area/fasting antral area). Gastric emptying of the test meal was derived from the measurement of the area under the postcibal antral distension curve (AUC). RESULTS: Fasting antral cross sectional area and AUC (gastric emptying) were similar in both groups. Antral postprandial expansion tended to be lower in AIDS patients compared to controls (mean+/-S.D.): 288+/-84 versus 397+/-156%; P=0.08. In AIDS patients the symptom score of dyspepsia showed a positive correlation (r=0.55; P<0.05) with fasting antral area and a negative correlation (r=-0.62; P<0.05) with postprandial expansion. No signs of autonomic neuropathy were to be found in the AIDS patients tested in this study. CONCLUSION: A wider fasting antral cross sectional area and an impaired antral postprandial expansion are related to dyspeptic symptoms in AIDS patients. This suggests the same relationship between dyspeptic symptoms and disturbed antral distension as seen in other patients with functional dyspepsia. PMID- 10874195 TI - Calcified epidermoid cyst in the testis: an unusual finding on ultrasound. AB - Epidermoid cysts of the testis are uncommon, benign testicular tumours. They are often seen on ultrasound as rounded, hypoechoic lesions due to high keratin contents. Calcification within epidermoid cysts is rare. We report a case of prominent calcifications within an epidermoid cyst. If a possible epidermoid cyst is identified with ultrasound, testis-sparing surgery should be considered. PMID- 10874196 TI - Sonographic detection of intestinal pneumatosis. AB - Intestinal pneumatosis is an uncommon affection characterized by the presence of gas in the wall of the gastro-intestinal tract. The prognosis of this condition, observed in benign or severe diseases, is based on the outcome of the underlying affection. The diagnosis of pneumatosis intestinalis is unusually made with sonography. We report a case of pneumatosis intestinalis due to small bowel necrosis, initially suggested with sonography and further confirmed with computed tomography (CT) and pathology. PMID- 10874197 TI - Ultrasonographic evaluation of the influence of different postures on diaphragmatic motion in mechanically ventilated patients. AB - We confirmed a diaphragmatic motion during mechanical ventilation in four patients with central nervous system damage. The right hemidiaphragm were visualized with 3.5 MHz ultrasound, and motion was measured using time-motion analysis in separate postures. The dome was most impaired during the right decubitus posture. The crural region was most improving during the left decubitus posture. The costal region was most impaired during the sitting posture. The motion of the dependent region was impaired. PMID- 10874199 TI - Guest editorial. PMID- 10874198 TI - Simultaneous occurrence of fat necrosis and carcinoma after breast injury in a traffic accident. AB - An old female patient presented after a car accident with clinical and sonographic evidence of two lesions, located along the seat-belt line. Despite the recent history of trauma and the localization in the traumatized area, the discrepancy between the sonographic size of the smaller lesion and the findings on palpation, together with the lack of typical mammographic findings for fat necrosis rose the suspicion of malignancy. A fine needle aspiration biopsy was performed to confirm the nature of the smaller lesion. PMID- 10874200 TI - Differential diagnosis of hereditary pigmentary maculopathies. AB - The hereditary pigmentary macular dystrophies rank among the most challenging diagnostic entities in optometric practice. Patients with these disorders can present with clinical characteristics along a broad spectrum, ranging from normal fundus findings to profound macular changes, with or without reduction in visual function. This paper reviews the key differentiating aspects of this group of disorders, including clinical testing and management strategies. PMID- 10874201 TI - Treatment of age-related macular degeneration. AB - Age-related macular degeneration (AMD), while rapidly becoming more prevalent due to an aging population, is still poorly understood and treatment modalities are limited. Fortunately, advances are being made in the treatment of AMD that may greatly alter the outcome of this debilitating disease. Treatments for both wet and dry AMD are reviewed. PMID- 10874202 TI - The role of submacular surgery in the treatment of choroidal neovascular membranes. AB - The growth of choroidal neovascular membranes (CNVM) beneath the macula can cause significant disturbances of central vision. Conditions such as age-related macular degeneration (AMD) and presumed ocular histoplasmosis syndrome (POHS) are the most common etiologies. The Macular Photocoagulation Study group presented data that clearly showed laser photocoagulation to be beneficial in the treatment of CNVM. Poor visual results and a high rate of recurrence have prompted clinicians to seek out alternative treatments. Experiences with CNVM removal utilizing submacular surgical techniques have shown that central visual function may be restored or preserved in POHS, multifocal choroiditis and idiopathic causes. The Submacular Surgery Trials were designed to investigate whether submacular surgery is more effective in retaining central acuity in patients with subfoveal CNVM, than observation alone. The goal of this paper is to review the role of submacular surgery in the treatment of subfoveal choroidal neovascular membranes. PMID- 10874203 TI - Update on current surgical management of idiopathic macular holes. AB - Idiopathic macular holes were generally considered an untreatable condition until 1991 when the first papers reported successful closure of macular holes and visual improvement using pars plana vitrectomy, peeling of the cortical vitreous and face-down positioning. Since that time, the original pathogenesis theory has been refined as well as surgical methods and techniques. Currently, macular hole surgery is considered the most successful vitreoretinal surgery, with a greatly improved prognosis for patients. This article will review the general characteristics of macular holes and the clinical trials and outcomes that have led to the latest techniques in idiopathic macular hole surgery. PMID- 10874204 TI - Toxoplasmosis in a patient with non-Hodgkin's lymphoma: the need for a good differential diagnosis. AB - Toxoplasma gondii is an obligate intracellular parasite responsible for toxoplasmosis. Congenital and acquired forms of the disease have now been reported. In immunocompromised patients, the disease entity may resemble other diseases. Presented for discussion is an interesting case of acquired toxoplasmosis in an immunocompromised patient with several confounding factors necessitating a good differential diagnosis list. Also provided is a comprehensive review of toxoplasmosis diagnosis and management. PMID- 10874205 TI - Stargardt's macular dystrophy. AB - Stargardt's disease is the most common hereditary macular dystrophy with an estimated incidence of 1 in 10000. The typical patient presents with visual symptoms between the first and third decades of life. This paper will discuss a patient who was diagnosed with Stargardt's macular dystrophy (SMD) at an older age. The clinical characteristics, functional testing, histopathology, differential diagnosis and recent genetics advances related to SMD will be discussed in this report. PMID- 10874206 TI - Idiopathic juxtafoveolar retinal telangiectasia: a review and case report. AB - Idiopathic juxtafoveolar retinal telangiectasis is a group of retinal vascular anomalies characterized by retinal vessel dilation and tortuosity, multiple aneurysm formations, varying degrees of vascular leakage and lipid exudate deposition. Idiopathic juxtafoveolar retinal telangiectasis may occur as a primary disorder (either congenital or acquired), or may be caused or accompanied by other systemic or ocular diseases. The visual prognosis and effectiveness of therapy is dependent upon the etiology of the retinal telangiectasis. Included in this review is a case report, as well as the classification system used to identify idiopathic juxtafoveolar retinal telangiectasia. PMID- 10874207 TI - Macula double trouble. PMID- 10874208 TI - Promotion of skin carcinogenesis by dimethylarsinic acid in keratin (K6)/ODC transgenic mice. AB - Dimethylarsinic acid (DMA) is a major metabolite of inorganic arsenicals in mammals, and arsenic exposure is associated with tumor development in a wide variety of human tissues, particularly the skin. Transgenic mice with ornithine decarboxylase (ODC) targeted to hair follicle keratinocytes are much more sensitive than littermate controls to carcinogens. In this study we investigated the promoting effect of DMA on skin carcinogenesis in such K6 / ODC transgenic mice. The back skin of female C57BL / 6J K6 / ODC transgenic mice, 10 to 14 weeks old, was initiated with topical application of 7, 12 dimethylbenz[alpha]anthracene (DMBA) at a dose of 50 microg or acetone alone on day 1 of the experiment, followed by treatment with 3.6 mg of DMA, 5 microg of 12 O-tetradecanoylphorbol-13-acetate (TPA) or neutral vehicle (control) twice a week for 18 weeks. Mice were killed 1 week after the end of the treatment. Induction of skin tumors was significantly accelerated in the DMA-treated group, as well as in the TPA-treated group, indicating that DMA has a promoting effect on skin tumorigenesis in K6 / ODC transgenic mice. PMID- 10874209 TI - Chemopreventive effects of bovine lactoferrin on N-butyl-N-(4 hydroxybutyl)nitrosamine-induced rat bladder carcinogenesis. AB - Chemopreventive effects of bovine lactoferrin (bLF), which is found at high concentrations in colostrum, on rat bladder carcinogenesis were investigated using a rat bladder medium-term bioassay. In experiment 1, a total of 80 F344 male rats, 6 weeks old, were divided into 5 groups. Groups 1 and 2 were treated with 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in the drinking water for 8 weeks and after a 1-week interval, received dietary supplementation with 2% and 0.2% bLF, respectively. Group 3 received 0.05% BBN for 8 weeks and then no treatment. Group 4 was administered 2% bLF alone from week 9, without prior carcinogen exposure. Group 5 was maintained without any treatment throughout the experiment. All rats were killed at the end of week 36. Group 1 demonstrated a significantly decreased multiplicity of the bladder tumors (carcinomas and papillomas) as compared with group 3. Maximum cut surface areas of bladder tumors were also significantly decreased in groups 1 and 2 compared with group 3. No bladder tumors were observed in groups 4 or 5. In experiment 2, a total of 60 rats were divided into two groups (30 rats each); both were treated with 0.05% BBN for 4 weeks and after a 1-week interval, one received 2% bLF (group 1) and the other, basal diet (group 2) for 4 weeks. Group 1 demonstrated a tendency for decrease of the 5-bromo-2'-deoxyuridine (BrdU) labeling index. bLF was detected in the urine of rats fed bLF by ELISA as well as western blot analysis. The findings indicate that 2% bLF can inhibit BBN-induced rat bladder carcinogenesis, and that this may be due to bLF in the urine. PMID- 10874210 TI - Three new regions on chromosome 17p13.3 distal to p53 with possible tumor suppressor gene involvement in lung cancer. AB - We investigated loss of heterozygosity (LOH) at the distal portion of the p53 gene on the short arm of chromosome 17 in lung cancers in order to search for new tumor suppressor genes. The roles of the putative genes were also studied in terms of pathological features. One hundred and forty-five resected non-small cell lung cancers were examined for LOH using 11 markers mapped on, and distal to the p53 locus, and deletion maps were constructed. Four commonly deleted regions were found: one from TP53 to ENO3, where the p53 gene resides, and three others from ENO3 to D17S1566, D17S379 to D17S1574 and distal to ABR, with LOH frequencies almost the same as, or higher than, at the TP53 locus. Examination of the relationship between LOH of the latter three regions and histopathological parameters of adenocarcinomas (genetically negative for p53 mutation) revealed allelic losses on D17S379 to be associated with advanced lesions, while D17S513 was more frequently deleted in poorly differentiated tumors. These results indicate that new tumor suppressor gene(s) may reside on these three distinctly deleted regions on chromosome 17p13.3 distal to the p53 gene in lung cancer, with possible roles in progression and differentiation of adenocarcinomas. PMID- 10874211 TI - Isolation and characterization of human NBL4, a gene involved in the beta catenin/tcf signaling pathway. AB - beta-Catenin, a key regulator of cellular proliferation, is often mutated in various types of human cancer. To investigate cellular responses related to the beta-catenin signaling pathway, we applied a differential display method using mouse cells transfected with an activated form of mutant beta-catenin. This analysis and subsequent northern-blot hybridization confirmed that expression of a murine gene encoding NBL4 (novel band 4.1-like protein 4) was up-regulated by activation of beta-catenin. To examine a possible role of NBL4 in cancer, we isolated the human homologue of the murine NBL4 gene by matching mNBL4 against the human EST (expressed sequence tag) database followed by 5' rapid amplification of cDNA ends (5'RACE). The cDNA of hNBL4 encoded a protein of 598 amino acids that shared 87% identity in amino acid sequence with murine NBL4 and 71% with zebrafish NBL4. A 2.2-kb hNBL4 transcript was expressed in all human tissues examined with high levels of expression in brain, liver, thymus and peripheral blood leukocytes and low levels of expression in heart, kidney, testis and colon. We determined its chromosomal localization at 5q22 by fluorescence in situ hybridization. Expression of hNBL4 was significantly reduced when beta catenin was depleted in SW480 cells, a human cancer cell line that constitutionally accumulates beta-catenin. The results support the view that NBL4 is an important component of the beta-catenin / Tcf pathway and is probably related to determination of cell polarity or proliferation. PMID- 10874212 TI - Spicamycin and KRN5500 induce apoptosis in myeloid and lymphoid cell lines with down-regulation of bcl-2 expression and modulation of promyelocytic leukemia protein. AB - Spicamycin is a potent inducer of differentiation of human myeloid leukemia cells (HL-60) and murine myeloid leukemia cells (M1). One of the spicamycin derivatives, KRN5500, shows a broad spectrum of antitumor activity against human tumor xenografts in nude mice. In this study, we first investigated the differentiation efficacy of spicamycin and KRN5500 in HL-60 and acute promyelocytic leukemia cell line, NB4, and found that low concentrations of both compounds induced differentiation to a small extent in both cell lines, but markedly induced apoptosis in NB4 cells. Further investigation in a myeloid leukemia cell line, NKM-1, a lymphoma cell line, Daudi, and a multiple myeloma cell line, NOP-1, showed that high concentrations of both compounds also induced apoptosis in these cells. The 50% inhibitory concentration (IC(50)) determined by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that myeloid cells were more sensitive to both compounds than lymphoid cells, and spicamycin was more potent than KRN5500. Western blot analysis of Bcl-2, Bcl-xL and Bax expression and immunofluorescence analysis of promyelocytic leukemia (PML) protein indicated that apoptosis induced by spicamycin and KRN5500 was associated with down-regulation of Bcl-2 expression and modulation of PML protein. Thus, spicamycin and KRN5500 may be useful for the treatment of myeloid and lymphoid neoplasms. PMID- 10874213 TI - A single nucleotide polymorphism in the matrix metalloproteinase-1 promoter in endometrial carcinomas. AB - Recent studies demonstrated that a single guanine insertion polymorphism in a matrix metalloprotease-1 promoter created an Ets binding site and affected the elevation of the transcriptional level of matrix metalloproteinase-1 (MMP-1). Furthermore, in tumor cell lines derived from melanoma and breast cancer, the incidence of the 2G / 2G genotype was significantly higher than that in the normal population. To evaluate the contribution of this polymorphism in endometrial carcinomas, we genotyped 100 endometrial carcinomas and then analyzed immunoexpression of MMP-1 in these carcinomas. We found that endometrial carcinoma patients showed a significantly higher rate of 1G / 2G or 2G / 2G genotype than control individuals, and that tumors containing the 2G allele(a) expressed MMP-1 protein more frequently than those with 1G / 1G genotype. Therefore, the single nucleotide polymorphism at the MMP-1 promoter affected the expression level of the MMP-1 protein, which may result in the association with more aggressive character in endometrial carcinoma. Our result suggests that the presence of 2G polymorphism at the MMP-1 promoter may be one of the risk factors for the development and / or progression of endometrial carcinoma. PMID- 10874214 TI - Induction of cytotoxic T lymphocytes from peripheral blood of human histocompatibility antigen (HLA)-A31(+) gastric cancer patients by in vitro stimulation with antigenic peptide of signet ring cell carcinoma. AB - Antigenic peptides have been used as a cancer vaccine in melanoma patients and have led to a drastic regression of metastatic tumors. However, few antigens have been identified in non-melanoma tumors. We recently purified a new natural antigenic peptide, designated F4. 2, by biochemical elution from a human gastric signet cell carcinoma cell line and showed that it is recognized by an autologous human histocompatibility antigen (HLA)-A31-restricted cytotoxic T lymphocyte (CTL) clone. Here we describe in vitro induction of F4. 2-specific CTLs from peripheral blood T lymphocytes of HLA-A31( +) gastric cancer patients. The T cells of seven HLA-A31( +) patients with gastric cancers were stimulated in vitro by F4.2-pulsed autologous dendritic cells which had been induced from peripheral blood of each patient by incubation in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF) and IL-4. We tested the cytotoxicity of the T cells against F4.2-loaded C1R-A*31012 by a 6-h (51)Cr release assay after 3 stimulations with F4.2-pulsed dendritic cells. F4.2-specific cytotoxicity was detectable in the stimulated T cells from two of the seven HLA-A31( +) patients. Further, both F4.2-specific CTLs also lysed the gastric cancer cell line, HST-2, from which F4.2 was derived. These results suggest that F4.2 peptide may be useful as an HLA-A31-restricted peptide vaccine in certain patients with gastric cancer. PMID- 10874215 TI - Avidin chase can reduce myelotoxicity associated with radioimmunotherapy of experimental liver micrometastases in mice. AB - Myelotoxicity is the main factor which decides the maximum tolerated dose (MTD) in radioimmunotherapy (RIT). Since bone marrow is mostly irradiated from blood radioactivity, enhancing the clearance of unbound circulating radiolabeled antibody is important to reduce myelotoxicity and to increase the MTD. We applied the avidin chase method, which was devised to obtain high tumor-to-background ratios in tumor-targeting, to RIT of experimental liver micrometastases and evaluated its influence on the side effects and therapeutic outcome. Seven days after intrasplenic injection of human colon cancer LS174T cells, nude mice were intravenously injected with biotinylated (131)I-labeled anti-CEA monoclonal antibody (MAb) (24 - 38 microg, 11.1 MBq). Mice of the chase group then received an intravenous injection of avidin twice (24 and 30 h, 72 - 115 microg each). Biodistribution, side effects (white blood cell counts and body weight change), and short- and long-term therapeutic effects were determined. Avidin chase markedly accelerated the clearance of radiolabeled MAb from the blood (P < 0.0001) and normal tissues, resulting in milder leukocytopenia and body weight loss, both of which recovered earlier than in the non-chase group (P < 0.01). The tumor uptake of radiolabeled MAb was also decreased by avidin chase, but the metastases-to-background ratios were increased. Avidin chase gave the therapeutic gain ratio of 1.89. Treated groups with and without avidin chase showed significant therapeutic effects compared to the non-treated group. There was no significant difference in the therapeutic effects between the two treated groups. Avidin chase effectively reduced the side effects of RIT and should increase the MTD. PMID- 10874216 TI - Human T cell leukemia cell death by apoptosis-inducing nucleosides from CD57(+) HLA-DR(bright) natural suppressor cell line. AB - Apoptosis-inducing nucleosides (AINs) released from CD57( +) HLA-DR(bright) natural suppressor (57.DR-NS) cell line, derived from human decidual tissue, were isolated from 57.DR-NS cell culture supernatant by the combination of thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). Apoptotic cell death was strongly induced in human T cell leukemia Molt4 cells treated with AINs, absolutely depending on DNA strand breaks, with activation of the caspase cascade, especially caspase-3. The administration of AINs to Molt4 tumor-bearing severe combined immunodeficiency (SCID) mice resulted in drastic suppression of tumor growth, with a decrease of tumor size and the appearance of apoptotic signals in tumor tissue. Thus, AINs are candidates for development as anticancer agents. PMID- 10874217 TI - Multidrug resistance reversal activity of taxoids from Taxus cuspidata in KB-C2 and 2780AD cells. AB - Some non-taxol-type taxoids having neither an oxetane ring at C-4 and C-5 nor an N-acylphenyl-isoserine group at C-13, such as taxuspine C, 2' desacetoxyaustrospicatine, and 2-desacetoxytaxinine J, which were isolated from the Japanese yew Taxus cuspidata, increased cellular accu-mulation of vincristine (VCR) in multidrug-resistant 2780AD cells as potently as verapamil, and efficiently inhibited [(3)H]azidopine photolabeling of P-glycoprotein (P-gp). Taxuspine C, 2'-desacetoxyaustrospicatine, and 2-desacetoxytaxinine J at 10 microM completely reversed the resistance to colchicine, VCR, and taxol in KB-C2 cells, which overexpress P-gp, while taxinine and taxinine M showed no effect. Taxuspine C, 2'-desacetoxyaustrospicatine, and 2-desacetoxytaxinine J may be candidate pharmaceuticals for reversing multidrug resistance (MDR) and also may be good modifiers of MDR in cancer chemotherapy. PMID- 10874218 TI - Suppression of tumor growth and downregulation of platelet-derived endothelial cell growth factor / thymidine phosphorylase in tumor cells by angiogenesis inhibitor TNP-470. AB - We investigated the effects of the angiogenesis inhibitor TNP-470 on human lung squamous cell carcinoma cell lines H226B and H226Br both in vivo and in vitro. H226B was established from human lung squamous cell carcinoma and H226Br was established from a brain metastatic lesion of H226B in nude mice. Nude mice inoculated with these cells were treated with 30 mg / kg of TNP-470 subcutaneously every other day. At this dose, TNP-470 only significantly suppressed the growth of H226Br tumor, but not H226B tumor. Attempts to use a high dose of TNP-470 (100 mg / kg) resulted in a severe loss of body weight. Immunohistochemical studies showed marked tumor vascularization in H226Br tumor, but the formation of new blood vessels was suppressed by 30 mg / kg of TNP-470. Investigation of the mechanism of anti-angiogenic effects of TNP-470 in vivo showed that the expression and the activity of platelet-derived endothelial cell growth factor / thymidine phosphorylase (PD-ECGF / dThdPase) in H226Br tumor was significantly suppressed by 30 mg / kg of TNP-470. Furthermore, TNP-470 inhibited cell growth of cultured H226Br dose-dependently at concentrations of 1 microg / ml. Immunoblot analysis revealed H226Br cells gave a stronger PD-ECGF signal than H226B cells, and the expression of PD-ECGF / dThdPase in H226Br was also suppressed by treatment with TNP-470 at 0.1 microg / ml. No change in basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF) was noted in these cell lines. Our results suggested that TNP-470 acts, at least in part, by downregulation of PD-ECGF / dThdPase in this cell line. PMID- 10874219 TI - Effects of 1-methyl-3-propyl-7-butylxanthine (MPBX) on idarubicin-induced antitumor activity and bone marrow suppression. AB - The effects of 1-methyl-3-propyl-7-butylxanthine (MPBX), a xanthine derivative, on idarubicin (IDA)-induced antitumor activity against P388 leukemia cells (P388) and bone marrow suppression were examined. In P388 tumor-bearing mice, the combination of MPBX with IDA increased the antitumor activity of IDA. The IDA concentration in the tumors in the MPBX combination group increased by 2.0-fold compared to the level in the IDA-alone group. On the other hand, as regards IDA induced bone marrow suppression, the combination of MPBX with IDA reduced the decrease in the bone marrow cell number by 30% compared to that in the IDA-alone group. In addition, the IDA concentration in the bone marrow cells was decreased by the combination of MPBX with IDA. An in vitro experiment showed that MPBX facilitated IDA influx and suppressed IDA efflux in P388 cells. In conclusion, the combination of MPBX with IDA increased the antitumor activity and decreased the bone marrow suppression. Therefore, we expect that the combination of MPBX with IDA will be useful for leukemia chemotherapy. PMID- 10874220 TI - Heterogeneous nuclear ribonucleoprotein B1 expressed in esophageal squamous cell carcinomas as a new biomarker for diagnosis. AB - We recently reported that heterogeneous nuclear ribonucleoprotein (hnRNP) B1 was overexpressed in most human lung cancers, especially squamous cell carcinoma (SCC), as well as human oral SCC. To find the significance of hnRNP B1 in cancer diagnosis, we studied hnRNP B1 expression in 16 paraffinized sections of esophageal SCC, using immunohistochemical staining with anti-hnRNP B1 polyclonal antibody, raised in a rabbit. We compared the expression of hnRNP B1 in cancerous and noncancerous regions of the same specimen: enhanced expression was observed in 63% of cancerous regions (10 / 16), whereas none of the noncancerous regions showed enhanced expression. The enhanced expression of hnRNP B1 in cancerous regions was compared with that in noncancerous tissue in relation to histopathological grade: 83% for well differentiated (5 / 6), 83% for moderately differentiated (5 / 6) and 0% for poorly differentiated (0 / 4). Histologically, enhanced expression of hnRNP B1 was observed around cancer pearls, as well as in the cells of nests lacking keratinization in well and moderately differentiated SCC. Western blotting analysis revealed enhanced expression in three frozen specimens of moderately differentiated SCC. Using esophageal cancer cell lines, we further confirmed the decreased expression in poorly differentiated SCC cells, compared with other differentiation types. All our results support the significance of hnRNP B1 expression in esophageal SCC as a unique diagnostic marker with regard to association between expression level and histopathological grading. PMID- 10874221 TI - Noncontact anterior cruciate ligament injuries: risk factors and prevention strategies. AB - An estimated 80,000 anterior cruciate ligament (ACL) tears occur annually in the United States. The highest incidence is in individuals 15 to 25 years old who participate in pivoting sports. With an estimated cost for these injuries of almost a billion dollars per year, the ability to identify risk factors and develop prevention strategies has widespread health and fiscal importance. Seventy percent of ACL injuries occur in noncontact situations. The risk factors for non-contact ACL injuries fall into four distinct categories: environmental, anatomic, hormonal, and biomechanical. Early data on existing neuromuscular training programs suggest that enhancing body control may decrease ACL injuries in women. Further investigation is needed prior to instituting prevention programs related to the other risk factors. PMID- 10874222 TI - Necrotizing soft-tissue infections. AB - Necrotizing fasciitis is a rare and often fatal soft-tissue infection involving the superficial fascial layers of the extremities, abdomen, or perineum. Necrotizing fasciitis typically begins with trauma; however, the inciting event may be as seemingly innocuous as a simple contusion, minor burn, or insect bite. Differentiating necrotizing infections from common soft-tissue infections, such as cellulitis and impetigo, is both challenging and critically important. A high degree of suspicion may be the most important aid in early diagnosis. Prompt diagnosis is imperative because necrotizing infections typically spread rapidly and can result in multiple-organ failure, adult respiratory distress syndrome, and death. Although group A Streptococcus is the most common bacterial isolate, a polymicrobial infection with a variety of Gram-positive, Gram-negative, aerobic, and anaerobic bacteria is more common. Orthopaedic surgeons are often the first physicians to evaluate patients with such infections and therefore need to be familiar with this potentially devastating disease and its management. Prompt diagnosis, immediate administration of broad-spectrum antibiotic coverage, and emergent aggressive surgical debridement of all compromised tissues are critical to reduce the morbidity and mortality of these rapidly progressing infections. PMID- 10874223 TI - Thoracic disk disease: diagnosis and treatment. AB - Symptomatic degenerative disk disease is much less common in the thoracic spine than in the cervical and lumbar regions. Accurate diagnosis relies on a strong clinical suspicion that is confirmed with appropriate diagnostic imaging. Presenting symptoms vary tremendously, from atypical pain patterns to myelopathy. The use of computed tomography in combination with myelography and magnetic resonance imaging have greatly increased the ability to accurately visualize thoracic spine disorders. The superior resolution of available imaging modalities has made the incidental detection of asymptomatic thoracic disk abnormalities more frequent. Most patients with symptomatic thoracic disk disease will respond favorably to nonoperative management. Surgery is indicated for the rare patient with an acute thoracic disk herniation with progressive neurologic deficit (i.e., signs or symptoms of thoracic spinal cord myelopathy). Once surgical intervention has been chosen, careful preoperative planning is necessary. The level, anatomic location, and morphology of the herniation must be precisely determined to select the optimal approach. Posterior laminectomy has largely been abandoned for the treatment of symptomatic thoracic disk protrusions. Surgeons still may choose among anterior, lateral, and posterior approaches when surgically addressing the thoracic intervertebral disk. PMID- 10874224 TI - Lunotriquetral instability: diagnosis and treatment. AB - Isolated injury of the lunotriquetral interosseous ligament complex and associated structures is less common and is poorly understood compared with the other proximal-row ligament injury, scapholunate dissociation. The spectrum of injuries ranges from isolated partial tears to frank dislocation, and from dynamic to static carpal instability. The diagnosis may be difficult to establish because of the many possible causes of ulnar-sided wrist pain and the often normal radiographic appearance. The mechanism of injury is variable and includes attrition by age, positive ulnar variance, and perilunate or reverse perilunate injury. Appropriate treatment requires assessment of the degree of instability and the chronicity of the injury. Options include corticosteroid injection, immobilization, ligament repair, ligament reconstruction with tendon grafts, limited intercarpal arthrodesis, and ulnar shortening. PMID- 10874225 TI - Surgical alternatives for treatment of articular cartilage lesions. AB - Articular cartilage injuries in the knee are common; fortunately, full-thickness articular cartilage defects constitute only a small portion of this group. These lesions may be incidentally encountered during ligament or meniscal surgery, having been silent or asymptomatic for an unknown period of time. However, when they are large and symptomatic, the surgeon may choose from a wide array of techniques available for treatment. The relatively small number of natural history studies regarding full-thickness articular surface lesions complicates the decision-making process. Accurate evaluation and classification of the anatomic defect aids in the development of a clinical algorithm for treatment. Surgical techniques are either reparative or restorative in nature. Reparative techniques fall short of complete reestablishment of the articular cartilage; however, the resultant repairs may remain quite functional for varying periods of time. Restorative techniques attempt to reestablish the native articular surface. To date, no peer-reviewed, prospective, randomized, controlled studies of operative versus nonoperative treatment for full-thickness articular cartilage lesions have been published. Even though the long-term results of surgical treatment for full-thickness articular surface lesions remain unknown, the early results are encouraging. PMID- 10874226 TI - Role of neurophysiologic evaluation in diagnosis. AB - The electrodiagnostic evaluation assesses the integrity of the lower-motor-neuron unit (i.e., peripheral nerves, neuromuscular junction, and muscle). Sensory- and motor-nerve conduction studies measure compound action potentials from nerve or muscle and are useful for assessing possible axon loss and/or demyelination. Needle electromyography measures electrical activity directly from muscle and provides information about the integrity of the motor unit; it can be used to detect loss of axons (denervation) as well as reinnervation. The electrodiagnostic examination is a useful tool for first detecting abnormalities and then distinguishing problems that affect the peripheral nervous system. In evaluating the patient with extremity trauma, it can differentiate neurapraxia from axonal transection and can be helpful in following the clinical course. In patients with complex physical findings, it is a useful adjunct that can help discriminate motor neuron disease from polyneuropathy or myeloradiculopathy due to spondylosis. PMID- 10874227 TI - Ankle arthrodesis: indications and techniques. AB - Patients with ankle arthritis and deformity can experience severe pain and functional disability. Those patients who do not respond to nonoperative treatment modalities are candidates for ankle arthrodesis, provided pathologic changes in the subtalar region can be ruled out. Several techniques are available for performing the procedure; the most successful combine an open approach with compression and internal fixation. The foot must be positioned with regard to overall limb alignment and in the optimal position for function. A nonunion rate as high as 40% has been reported. Osteonecrosis of the talus and smoking are known risk factors for nonunion. When good surgical technique is used in carefully selected patients, ankle arthrodesis can be a reliable procedure for the relief of functionally disabling ankle arthritis, deformity, and pain. PMID- 10874228 TI - A prospective, randomized controlled trial comparing the efficacy and safety of sotalol, amiodarone, and digoxin for the reversion of new-onset atrial fibrillation. AB - STUDY OBJECTIVE: A prospective, randomized controlled trial of new-onset atrial fibrillation was conducted to compare the efficacy and safety of sotalol and amiodarone (active treatment) with rate control by digoxin alone for successful reversion to sinus rhythm at 48 hours. METHODS: We prospectively randomly assigned 120 patients with atrial fibrillation of less than 24 hours' duration to treatment with sotalol, amiodarone, or digoxin using a single intravenous dose followed by 48 hours of oral treatment. Patients had ECG monitoring for 48 hours, and time of reversion, adequacy of rate control, and numbers of adverse events were compared. After 48 hours, those still in atrial fibrillation underwent cardioversion according to a standardized protocol. After 48 hours of therapy and attempted cardioversion, the number of patients whose rhythms had successfully reverted were compared. RESULTS: There was a significant reduction in the time to reversion with both sotalol (13. 0+/-2.5 hours, P <.01) and amiodarone (18.1+/ 2.9 hours, P <.05) treatment compared with digoxin only (26.9+/-3.4 hours). By 48 hours, the active treatment group was significantly more likely to have reverted to sinus rhythm than the rate control group (95% versus 78%, P <.05; risk ratio 5.4, 95% confidence interval [CI] 1.5 to 19.2 ). In those patients whose rhythms did not revert to sinus rhythm, there was superior ventricular rate control in the sotalol group at both 24 and 48 hours compared with those who received either amiodarone or digoxin. There were also fewer adverse events in the active treatment group compared with the rate control group. CONCLUSION: Immediate pharmacologic therapy for new-onset atrial fibrillation with class III antiarrhythmic drugs (sotalol or amiodarone) improves complication-free 48-hour reversion rates compared with rate control with digoxin. PMID- 10874229 TI - Interpretation of immediate exercise treadmill test: interreader reliability between cardiologist and noncardiologist in a chest pain evaluation unit. AB - STUDY OBJECTIVE: To determine whether attending physicians in a chest pain evaluation unit (CPEU) can perform and interpret exercise testing with the same accuracy as cardiologists. METHODS: Between January 1996 and November 1998, immediate exercise tests were performed and interpreted by internists with additional training in exercise testing who serve as attending physicians in a CPEU at a large university medical center. For quality assurance, all tests were overread by a cardiologist. Test results were compared for each reader, and all tests with discrepant readings were reinterpreted by an independent cardiologist who was blinded to the previous results. Patients' clinical course was monitored for at least 30 days after exercise testing. RESULTS: The study group consisted of 645 patients (347 men, 298 women). Discrepant interpretations were found in 11 (1. 7%) patients. The agreement was 98.4% (kappa value 0.9618). The majority of discrepancies were insignificant and were based on subtle differences in the definition of a nondiagnostic test or the degree of ST-segment shift. Of the 11 discordant readings, the blinded cardiologist concurred with 5 (45%) of the CPEU interpretations and 4 (36%) of the cardiologist interpretations. In 2 cases, there was disagreement by all 3 interpreters. There was no cardiac morbidity or mortality of any patient with a discrepant reading. CONCLUSION: Our results suggest that noncardiologists serving as attending physicians in a CPEU can accurately interpret exercise tests and overreading by cardiologists for quality assurance is unnecessary. PMID- 10874230 TI - Safely directing patients to appropriate levels of care: guideline-driven triage in the emergency service. AB - STUDY OBJECTIVE: We sought to develop and validate standardized clinical criteria to identify patients presenting to the emergency department whose care may be safely deferred to a later date in a nonemergency setting. METHODS: Using a modified Delphi process, a 17-member multidisciplinary physician panel developed explicit, standardized, deferred-care criteria. In a prospective cohort design, emergency nurses at a tertiary care Veterans Administration (VA) Medical Center, using the criteria, screened 1,187 consecutive ambulatory adult patients presenting with abdominal pain, musculoskeletal symptoms, or respiratory infection symptoms. Patients meeting deferred-care criteria were offered the option of an appointment within 1 week in the ambulatory care clinic at the study site; all other patients were offered same-day care. As outcome measures, we assessed nonelective hospitalizations for related conditions occurring within 7 days of evaluation at our facility or any other VA facility within a 300-mile radius, and we assessed 30-day all-cause mortality. RESULTS: Two hundred twenty six (19%) patients met screening criteria for deferred care. Patients meeting deferred-care criteria experienced zero (95% confidence interval, 0% to 1.2%) related nonelective VA hospitalizations within 7 days of evaluation, and none died within 30 days. By contrast, 68 (7%) of 961 (95% confidence interval, 5.5% to 8.9%) patients who did not meet deferred-care criteria were hospitalized nonelectively for related conditions, and 5 (0.5%) died. CONCLUSION: By using hospitalization and 30-day mortality as safety gauges, standardized clinical criteria can identify, at presentation, VA ED users who may be safely cared for at a later date in a nonemergency setting. These guidelines apply to a significant proportion of VA ED users with common ambulatory conditions. These criteria deserve testing in other ED settings. PMID- 10874231 TI - Lifetime sexual assault prevalence rates and reporting practices in an emergency department population. AB - STUDY OBJECTIVE: Studies suggest significant rates of female sexual assault (SA); the majority of SAs remain unreported, and few victims receive medical care. The purpose of this study was to determine lifetime prevalence rates of SA in an emergency department population and to assess reporting patterns to police, physicians, and social service agencies. METHODS: A verbally administered survey was given to all female patients during 4-hour randomized periods in an urban Level I trauma center. All English-speaking, noncritically ill women who presented during the study period were eligible. RESULTS: Four hundred forty-two women were eligible; 360 (81%) women agreed to participate. The lifetime prevalence rate of SA was 39% (n=139). Ninety-seven women (70%) were older than 15 years at the time of SA. Of these 97 SAs occurring in adulthood, 49 (52%) reported assault by an acquaintance, family member, or friend; 28 (30%) by a stranger; and 17 (18%) by a partner. Forty-five (46%) women reported the crime to the police, 42 (43%) sought medical care, and 23 (25%) contacted a social service agency. Reporting patterns for victims assaulted by a stranger versus those assaulted by a partner were: reported to police 79% (95% confidence interval [CI] 62 to 95) versus 18% (95% CI 0 to 38); P <.001), received medical care 70% (95% CI 46 to 95) versus 29% (95% CI 11 to 48; P<.01), contacted a social service agency 30% (95% CI 5 to 47) versus 24% (95% CI 1 to 46; P=.63). CONCLUSION: Lifetime female SA rates in ED populations are significant. Fewer than half of SA victims report the assault to the police or seek medical care. Women assaulted by a partner are significantly less likely to report the SA to police or seek medical care. PMID- 10874232 TI - Opportunistic urine ligase chain reaction screening for sexually transmitted diseases in adolescents seeking care in an urban emergency department. AB - STUDY OBJECTIVE: Neisseria gonorrhoeae and Chlamydia trachomatis are the most common bacterial sexually transmitted diseases (STDs) in sexually active youth and many infections are asymptomatic or unrecognized. This study used ligase chain reaction assays for determination of prevalence of gonococcal and chlamydial infections in adolescents seeking care at an urban emergency department. METHODS: An unlinked prevalence study was performed with ligase chain reaction tests on voided urine specimens from a convenience sample of adolescents 14 years or older who sought care at the Children's Hospital of Alabama ED. Demographic data and data on care provided in the ED were determined from retrospective chart review of those patients whose urine specimens were tested. RESULTS: Of 282 urine specimens screened, 13.5% (38) yielded positive findings on ligase chain reaction testing for either or both pathogens (20 [7%] positive for N gonorrhoeae, 23 [8%] positive for C trachomatis). In the context of acute care, gonorrhea or chlamydial infection was diagnosed in 5 (1.8%). STD prevalence did not vary significantly by age. Only 39% (15/38) of patients with infections detected by ligase chain reaction testing received potentially effective antibiotics as a result of their urgent care evaluation. CONCLUSION: Many adolescents use the ED for nonurgent care and unsuspected STDs are often missed. Urine ligase chain reaction testing is a sensitive, noninvasive means of detecting STDs by which unsuspected adolescent STD cases can be detected in an ED setting. PMID- 10874233 TI - Survey of nationally registered emergency medical services providers: pediatric education. AB - STUDY OBJECTIVE: To survey emergency medical services (EMS) providers on a national level to determine and describe their perspective regarding their initial and continuing education (CE) needs in pediatrics. METHODS: A 10-question survey was developed, pilot-tested, and sent to EMS providers as a part of their National Registry of Emergency Medical Technicians reregistration materials. RESULTS: Surveys were completed by 18,218 EMS providers, a response rate of 67%. During a typical month, 60% of emergency medical technician-paramedics (EMT-Ps), 84% of EMT-intermediates (EMT-Is), and 87% of basic EMTs (EMT-Bs) care for 0 to 3 pediatric patients. CE was identified by all provider levels as the main source of their pediatric knowledge and skills. A state or national mandate for required CE in pediatrics was supported by 76% of surveyed providers. More than 70% of all providers responded they were comfortable to some degree with their own ability and their EMS system's ability when confronted with a critical pediatric call. Cost, availability, and travel distance were identified by all levels as the primary barriers to obtaining pediatric CE. All levels identified infants as the age of greatest concern if the provider was called to manage a critical case. CONCLUSION: Surveyed practicing nationally registered EMS providers have infrequent contact with pediatric patients and have acquired most of their pediatric knowledge and skills from CE. In general, these providers are comfortable with their personal and their system's ability to care for children, but clearly support the need for required pediatric CE and identify the birth to 3-year age range as the priority for an educational focus. Cost, travel distance, and availability of pediatric CE are barriers that should be considered if pediatric CE is to be required of EMS providers. PMID- 10874234 TI - Acute appendicitis in children: emergency department diagnosis and management. AB - Early diagnosis of appendicitis in infants and children can prevent perforation, abscess formation, and postoperative complications, and can decrease cost by shortening hospitalizations. This article reviews the epidemiology, physiology, and age-specific clinical presentation of childhood appendicitis. The accuracy of diagnostic adjuncts is reviewed, as are strategies for avoiding misdiagnosis and improving emergency department evaluation and management. PMID- 10874235 TI - Endotracheal tube introducer for failed intubations: a variant of the gum elastic bougie. AB - There is no universally accepted nonsurgical adjunct for management of the difficult airway in the emergency department. The gum elastic bougie is widely accepted in the British anesthesia literature. One model of endotracheal tube introducer, the Flex-Guide ET Tube Introducer (GreenField Medical Sourcing, Inc, Northborough, MA), is a less expensive plastic version of the gum elastic bougie with the same properties, available in the United States. We present 3 cases of its use in obtaining airway control in difficult airways in the ED. The bougie facilitates intubation where the cords are not visualized or neck movement is contraindicated, allows verification of correct placement before placing the endotracheal tube, is simple to use, and inexpensive to obtain. Reports of 100% first-attempt intubation success in difficult airways are published in the anesthesia literature. We advocate use of this device in the emergency department as a nonsurgical adjunct for difficult airway management. PMID- 10874236 TI - Health policy report introduction. PMID- 10874237 TI - Human error in medicine: promise and pitfalls, part 1. PMID- 10874238 TI - Update on emerging infections from the Centers for Disease Control and Prevention.Update: surveillance for West Nile virus in overwintering mosquitoes- New York, 2000. PMID- 10874239 TI - Board scores and resident performance: is there a link? PMID- 10874240 TI - Delivering news. PMID- 10874241 TI - Deferral out of the emergency department: the wrong solution? PMID- 10874242 TI - Pediatric continuing education for out-of-hospital providers: is it time to mandate review of pediatric knowledge and skills? PMID- 10874243 TI - The endotracheal tube introducer revisited. PMID- 10874244 TI - The gravest words: sudden-death notifications and emergency care. PMID- 10874245 TI - Rectus sheath hematoma. PMID- 10874246 TI - Rectus sheath hematoma PMID- 10874247 TI - Video self-instruction for cardiopulmonary resuscitation. PMID- 10874249 TI - Endogenous mediators and sepsis PMID- 10874248 TI - Endogenous mediators and sepsis. PMID- 10874250 TI - Dual randomization in cardiovascular trials. PMID- 10874251 TI - Memory function in patients with stable, moderate to severe cardiac failure. AB - BACKGROUND: Psychologic dysfunction is common in patients with heart disease. Patients with end-stage cardiac failure have cognitive dysfunction that can affect their ability to comply with complex treatment regimes. The prevalence of cognitive dysfunction in patients with stable, moderately severe cardiac failure is not known. The purpose of this study was to evaluate the prevalence of memory dysfunction in that population. METHODS: Twenty patients with previous myocardial infarction (MI), New York Heart Association class III or IV cardiac failure symptoms, and left ventricular ejection fraction (LVEF) <40% were studied. Twenty patients with previous MI, no heart failure symptoms, and LVEF >50% were studied as a control group. Memory function was assessed with the Rivermead Behavioural Memory Test and digit span test. Results were controlled for affective state and estimated premorbid intellectual function with the National Adult Reading Test. RESULTS: Patients with cardiac failure performed as well as controls in the memory test (mean [95% confidence intervals] score 18.2 [16.5-19.9] versus 19.7 [18.1-21.4] points, P =.20) and digit span test (10.1 [9.2-11.0] versus 11.0 [9.9 12.2] points). Affective symptoms were more prevalent among patients with cardiac failure. Estimated premorbid intellectual function appeared poorer in patients with cardiac failure (reading test score 24 [7-36] versus 32 [11-46] points). Memory test outcomes were not significantly affected after adjustment for these variables. CONCLUSIONS: Patients with prior MI and stable, moderately severe cardiac failure do not have significantly memory impairment. Memory dysfunction is not likely to interfere with rehabilitation or compliance with treatment regimes. Anxiety and depression, however, are relatively common in this group. PMID- 10874252 TI - Is the emperor really wearing new clothes? Informed consent for acute coronary syndromes. PMID- 10874253 TI - Clinical evaluation of chelation therapy: is there any wheat amidst the chaff? PMID- 10874254 TI - Atorvastatin versus revascularization treatment (AVERT): fact or fancy? PMID- 10874255 TI - Improvement of exercise capacity and left ventricular diastolic function with metoprolol XL after acute myocardial infarction. AB - BACKGROUND: Left ventricular (LV) diastolic function predicts and correlates with exercise capacity. Beta-blockers improve exercise capacity and LV diastolic function in patients with severe LV systolic dysfunction in dilated cardiomyopathy. However, information on the effect of metoprolol XL on exercise capacity in relation to LV diastolic function in patients with mild to moderate LV systolic dysfunction after acute myocardial infarction is limited. METHODS: In a randomized, double-blind, placebo-controlled study of 77 patients, a subgroup of 59 patients with mild to moderate LV systolic dysfunction after acute myocardial infarction were given metoprolol XL (n = 29) or placebo (n = 30). The effects of metoprolol XL on exercise capacity in relation to effects on LV diastolic filling were studied. Two-dimensional Doppler echocardiography and maximal symptom limited bicycle test were performed on days 5 through 7 and after 3 months. RESULTS: Maximal exercise capacity increased in the metoprolol XL group (124 +/- 30 W vs 135 +/- 29 W) compared with placebo (125 +/- 31 W vs 126 +/- 34 W) (P <.01). E/A ratio decreased, A peak velocity increased, reverse pulmonary flow decreased, and deceleration time was significantly prolonged in the metoprolol XL group. A significant correlation was found between the changes of deceleration time (metoprolol XL: rho = 0.69, P <.0001; placebo: rho = 0.31, P = not significant) and A peak velocity (metoprolol XL: rho = 0.71, P <.0001; placebo: rho = -0.15, not significant) in relation to changes of exercise capacity. CONCLUSION: Metoprolol XL increases exercise capacity after 3 months, and this change seems related to improvement of LV diastolic filling after acute myocardial infarction. PMID- 10874256 TI - Should we revise our diagnostic methods for Q-wave myocardial infarction in the presence of right bundle branch block? PMID- 10874257 TI - Classification and pharmacology of antiarrhythmic drugs. AB - Despite the emergence of several forms of nonpharmacologic therapy for cardiac arrhythmias, antiarrhythmic drugs continue to play an important role in the management of patients with this common clinical problem. The key to the proper use of antiarrhythmic drugs is a thorough knowledge of their mode of action and pharmacology. The pharmacology of antiarrhythmic drugs is particularly important because patients with cardiac arrhythmias frequently have multiorgan disease, which may influence the metabolism and elimination of antiarrhythmic drugs. The accumulation of toxic amounts of these agents can lead to dire effects including, but not limited to, ventricular proarrhythmia and malignant bradycardia. The goals of pharmacologic therapy of cardiac arrhythmia are to provide the maximum benefit in terms of arrhythmia suppression while maintaining patient safety. To accomplish these goals, a knowledge of the pharmacology of several antiarrhythmic drugs is mandatory. PMID- 10874258 TI - Low-molecular-weight heparin alone versus a combination of unfractionated heparin and low-molecular-weight heparin. AB - OBJECTIVES: We analyzed the effect of the pharmacologic combination of 2 indirect antithrombin drugs--enoxaparin (low-molecular-weight heparin) and unfractionated heparin--versus enoxaparin alone on the recurrence of ischemia. BACKGROUND: Blocking some key factors of the coagulation cascade supports the concept that an antithrombin effect is needed during the acute phase of ischemia. METHODS: This was a prospective, randomized, pilot trial in patients with an acute coronary ischemic event occurring within the previous 24 hours. A total of 126 patients were allocated to receive aspirin (200 mg/day orally) plus 1 mg/kg subcutaneous enoxaparin at 8 AM and 12.500 IU of subcutaneous unfractionated heparin at 8 PM (group A) or subcutaneous enoxaparin 1 mg/kg (group B). RESULTS: Severe recurrent ischemia provoking urgent coronary revascularization occurred in 12 patients (9.5%), 3 (5%) in group A and 9 (13%) in group B (P = .1). Refractory angina was present in 27 patients (21%), 10 (17%) in group A and 17 (25%) in group B (P = .45). The combination of severe recurrent ischemia and refractory angina occurred in 23% of group A, and 37% of group B (odds ratio 0.49; 95% confidence intervals, 0.21-1.15; P = .07). A total of 7 patients (5%) had acute nonfatal myocardial infarction develop, 3 (5%) in group A and 4 (6%) in group B. Two (1.6%) deaths were observed in the study, both in group B. The incidence of the double end point (death plus nonfatal myocardial infarction) was 5% in group A versus 9% in group B (P = .5) and the triple end point (death, nonfatal myocardial infarction, and severe recurrent ischemia) was 10.5% in group A vs 22% in group B (odds ratio 0.42, 95% confidence intervals, 0.13-1.29; P = .09). CONCLUSIONS: The combination of 2 indirect antithrombin drugs capable of intermittently blocking the coagulation system is not associated with a significant loss of safety. PMID- 10874259 TI - Chronic heart failure and exercise. PMID- 10874260 TI - Hierarchy of risk based on history and location of prior myocardial infarction in the thrombolytic era. GUSTO-I Investigators. AB - BACKGROUND: Among patients receiving thrombolytic therapy for myocardial infarction, the outcome for those with a history of infarction is dramatically worse than for those with their first event. Methods And Results We performed a post hoc analysis of patients with a history of myocardial infarction enrolled in the Global Utilization of Streptokinase and TPA for Occluded arteries (GUSTO)-I trial, focusing on the impact of the location of their current and prior events on mortality rates. Within the first 24 hours, mortality rate was greatest among patients with a current infarction in a territory remote from their previous event. By 48 hours after examination, mortality rates among patients with a second anterior infarct had overtaken that among patients with a current inferior/prior anterior infarct. This hierarchy of risk persisted at both 30 days and 1 year (mortality rate at 1 year: current anterior/prior inferior 23.2% +/- 1.4%, current anterior/prior anterior 20% +/- 1.5%, current inferior/prior anterior 17% +/- 1.2%, current inferior/prior inferior 10.8% +/- 0. 9%). CONCLUSIONS: In patients with ST-elevation myocardial infarction on a background of prior infarction, the location of current and prior events predicts a hierarchy of short- and long-term risk of death. PMID- 10874261 TI - Dissociation between ACE activity and autonomic response to ACE inhibition in patients with heart failure. AB - BACKGROUND: Administration of angiotensin-converting enzyme (ACE) inhibitors to patients with congestive heart failure has been shown to increase parasympathetic tone as indicated by increases in high-frequency heart rate variability. The mechanism for this effect, including its relation to changes in baroreflex activity, blood pressure variability, and suppression of ACE activity, remains undefined. This study was designed to test the relation of these variables, which may govern changes in autonomic activity, to the previously described increase in parasympathetic tone. METHODS: Seven patients with heart failure received a 3 hour infusion of the ACE inhibitor enalaprilat. Hemodynamic variables and parameters of heart rate and blood pressure variability, baroreflex gain derived from the interaction of heart rate and blood pressure variability, and serum ACE activity were measured during and after the infusion. Measures of heart rate and blood pressure variability were also compared against a historic control group. RESULTS: Serum ACE activity was significantly suppressed throughout and after enalaprilat infusion. Hemodynamic measures did not change other than a small decline in right atrial and pulmonary capillary wedge pressures. Parasympathetic tone showed an initial significant increase with a peak at 2 hours but then declined below baseline 8 hours after initiation of enalaprilat infusion. Sympathetically influenced low-frequency heart rate variability was significantly increased above baseline in the enalaprilat treatment group 8 hours after initiation of the infusion. Baroreflex gain showed a significant trend to an increase with the maximum value coinciding with the peak in parasympathetic tone. There was no change in blood pressure variability in the enalaprilat group and no change in baroreflex gain, heart rate variability, or blood pressure variability in the control group. CONCLUSIONS: Parasympathetic tone and baroreflex gain increased with parenteral administration of an ACE inhibitor but subsequently decreased below baseline values despite continued suppression of serum ACE activity. The dissociation between ACE suppression and autonomic response to ACE inhibition indicates that enzyme systems not reflected by plasma ACE activity or independent from the classic pathways of angiotensin formation contribute to the regulation of the autonomic response to ACE inhibition in patients with heart failure. The absence of significant change in hemodynamic variables or in blood pressure variability indicates that these autonomic changes are not an indirect reflex response to ACE inhibitor-induced vasodilation or hemodynamic baroreceptor stimulation. PMID- 10874262 TI - Arrhythmia risk stratification in idiopathic dilated cardiomyopathy based on echocardiography and 12-lead, signal-averaged, and 24-hour holter electrocardiography. AB - BACKGROUND: To date, considerable controversy exists regarding noninvasive arrhythmia risk stratification in idiopathic dilated cardiomyopathy (IDC). Methods and Results Between 1992 and 1997, 202 patients with IDC without a history of sustained ventricular tachycardia (VT) underwent echocardiography, signal-averaged electrocardiogram (ECG), and 24-hour Holter ECG in the absence of antiarrhythmic drugs. During 32 +/- 15 months of prospective follow-up, major arrhythmic events, including sustained VT, ventricular fibrillation, or sudden death, occurred in 32 (16%) of 202 patients. After adjusting for baseline medical therapy and antiarrhythmic therapy during follow-up, multivariate Cox regression analysis identified a left ventricular (LV) end-diastolic diameter >/=70 mm and nonsustained VT on Holter as the only independent arrhythmia risk predictors. The combination of an LV end-diastolic diameter >/=70 mm and nonsustained VT was associated with a 14. 3-fold risk for future arrhythmic events (95% confidence interval 2. 3-90). To further elucidate the prognostic value of LV ejection fraction, multivariate Cox analysis was repeated with ejection fraction forced to remain in the model. In the latter model, an ejection fraction /=70 mm and nonsustained VT on Holter, and the combination of LV ejection fraction /=0.1 mV ST-segment depression. RESULTS: Exercise duration, blood pressure, heart rate, and rate-pressure product during exercise did not differ between the 2 groups. End-diastolic volume at rest and at peak exercise did not differ between groups D and N. In contrast, the end systolic volume in group N decreased during exercise, whereas in group D it increased. As a result, the left ventricular ejection fraction in group D decreased from 70% +/- 7% to 59% +/- 15% (P <.0001), whereas ejection fraction in group N increased from 65% +/- 8% to 71% +/- 11% (P =.0002). There was a strong correlation between exercise-induced ST-segment depression and changes in ejection fraction from rest to peak exercise (P <.0001). CONCLUSIONS: These results suggest that the exercise-induced ST-segment depression seen in patients with nonobstructive HCM is associated with systolic dysfunction during exercise. PMID- 10874264 TI - Infarct size determination by technetium 99m sestamibi single-photon emission computed tomography predicts survival in patients with chronic coronary artery disease. AB - BACKGROUND: The prognostic value of infarct size quantification by technetium 99m sestamibi single-photon emission computed tomography (SPECT) in patients with chronic coronary artery disease (CAD) has not been established. Methods And Results Between September 1994 and May 1995, 1323 patients with known or suspected CAD were referred for perfusion imaging for clinical reasons and had infarct size determined by quantitative SPECT imaging. Patients underwent exercise stress (61%), pharmacologic stress (37%), and rest imaging (3%). Patients were excluded if they had cardiomyopathy, valvular heart disease, or myocardial infarction within 3 weeks of the SPECT study. There were 1224 patients who formed the study group. Follow-up was 94% complete at a median of 1.9 +/- 0.4 years. Sixty-five percent of patients had no measurable infarct. Among the patients with measurable infarcts, the mean infarct size by sestamibi imaging was 15.0% +/- 14.5% of the left ventricle (25% of infarcts /=19% of the left ventricle). By using stepwise regression analysis, age, diabetes, and hypercholesterolemia were all clinical predictors of overall death (P <.05). For cardiac death, only age and diabetes were significant. After adjusting for these clinical variables, infarct size remained an independent predictor of overall death (P =. 001) and survival free of cardiac death (P =.0002). However, when first-pass left ventricular ejection fraction was added to the models, infarct size was no longer significant. CONCLUSIONS: Infarct size determination by SPECT (99m)Tc sestamibi can predict subsequent death in patients with chronic CAD, although ejection fraction appears to have greater prognostic value. PMID- 10874265 TI - Reduction in the need for hospitalization for recurrent ischemic events and bleeding with clopidogrel instead of aspirin. CAPRIE investigators. AB - BACKGROUND: Repeat hospitalizations of patients with atherosclerosis represent a considerable burden on the health care system. We sought to determine whether clopidogrel compared with aspirin decreases the need for rehospitalization for ischemia and bleeding. METHODS AND RESULTS: The Clopidogrel Versus Aspirin in Patients at Risk of Ischemic Events (CAPRIE) trial was a randomized, blinded, multicenter, trial of 19,185 patients with atherosclerotic disease manifested as recent ischemic stroke or myocardial infarction or symptomatic peripheral arterial disease. Without any double-counting of events, the number of rehospitalizations for ischemic events (defined as angina, transient ischemic attack, or limb ischemia) or bleeding events was determined for the entire cohort. There was a significant reduction in the total number of rehospitalizations for ischemic events or bleeding with clopidogrel use compared with aspirin (1502 vs 1673; P =.010) over an average of 1.6 years of treatment. This reduction in rehospitalization was consistent across individual outcomes of angina, transient ischemic attack, limb ischemia, and bleeding. Compared with aspirin, clopidogrel also resulted in a 7.9% relative risk reduction in a combined end point of vascular death, stroke, myocardial infarction, or rehospitalization for ischemic events or bleeding (15.1% to 13.7% at 1 year; P =.011). Adjusting for baseline prognostic variables, clopidogrel therapy was an independent predictor for reduction of vascular death, stroke, myocardial infarction, or rehospitalization for ischemic events or bleeding (P =.009). CONCLUSIONS: Treatment with clopidogrel results in a significant decrease in the need for rehospitalization for ischemic events or bleeding compared with aspirin. This meaningful end point tracks well with other, more traditional measures of outcome and has incremental value beyond such end points. PMID- 10874266 TI - Clinical correlates and course of thrombocytopenia during percutaneous coronary intervention in the era of abciximab platelet glycoprotein IIb/IIIa blockade. AB - BACKGROUND: Thrombocytopenia is infrequently associated with abciximab therapy but may contribute to hemorrhagic risk. Factors associated with development of thrombocytopenia, the role of weight-adjustment in concomitant heparin administration, and clinical outcomes in patients with thrombocytopenia are not well defined. METHODS AND RESULTS: Pooled data from 3 placebo-controlled, randomized trials (EPIC, EPILOG, and EPISTENT) of abciximab therapy during percutaneous coronary intervention identified 178 patients (2. 4% of 7290 patients) in whom thrombocytopenia (platelet count <100 x 10(9)/L) developed after enrollment. Multivariate regression analysis identified age (>65 years; P <.001), weight (<90 kg; P =. 023), baseline platelet count (<200 x 10(9)/L; P <.001), abciximab therapy (P =.002), and enrollment into the EPIC trial (P <.001) to be associated with development of thrombocytopenia. Major and minor nonsurgical hemorrhage and transfusion were more frequent (all P <. 001) in thrombocytopenic patients. Although the primary composite clinical end point of these trials (death, myocardial infarction, or urgent revascularization to 30 days) was observed with similar frequency in patients with (11.2%) and those without (7.9%; P =.114) thrombocytopenia, 30-day mortality rate was higher in thrombocytopenic patients (8.4% vs 0.6%, respectively; P <.001). This excess mortality rate persisted after excluding patients in whom thrombocytopenia was first noted after the performance of coronary bypass surgery (4.8% vs 0.6%; P <.001). Among patients in whom thrombocytopenia developed during these trials, those who received prophylactic abciximab had fewer primary end point events (7.1% vs 23.1%; P =.056) and had a lower 30-day mortality rate (3.5% vs 15.4%; P =.048) than patients with thrombocytopenia who had received prophylactic placebo. CONCLUSIONS: Thrombocytopenia associated with abciximab therapy for percutaneous coronary intervention was more frequent in older, lighter-weight patients, those with lower baseline platelet counts, and in those patients who were enrolled into the EPIC trial. Both bleeding and transfusion events occur more frequently in patients with thrombocytopenia. Patients in whom thrombocytopenia developed during these trials had increased mortality rates to 30 days not attributable to the performance of coronary bypass surgery. Among patients with thrombocytopenia, those who received prophylactic abciximab had better clinical outcomes including survival than those who did not. PMID- 10874267 TI - First report of an intravenous and oral glycoprotein IIb/IIIa inhibitor (RPR 109891) in patients with recent acute coronary syndromes: results of the TIMI 15A and 15B trials. AB - BACKGROUND: RPR 109891 is a modified tetrapeptide glycoprotein IIb/IIIa inhibitor available in intravenous and oral formulations. Two phase II dose-ranging studies were performed to investigate pharmacodynamics and safety in acute coronary syndromes. METHODS: The Thrombolysis In Myocardial Infarction (TIMI) 15A trial was a randomized, open-label, study of RPR 109891 administered intravenously for 24 to 96 hours in 91 patients. TIMI 15B was a randomized, double-blind comparison of intravenous RPR 109891 plus 4 weeks of oral RPR 109891 (n = 142) compared with placebo (n = 50). RESULTS: Intravenous RPR 109891 exhibited a dose-response inhibition of platelet aggregation; mean inhibition after a bolus ranged from 53% to 92%, and at steady state 49% to 98%. Oral RPR 109891 demonstrated less platelet inhibition (peaks, range 48% to 59%; troughs, range 18% to 39%). Mean glycoprotein IIb/IIIa receptor occupancy and platelet inhibition were highly correlated (r = 0.82, 95% confidence interval 0.74-0.88). There were trends for increased major hemorrhage (10% vs 6%, P =.57), thrombocytopenia <90,000 cells/mm(3) (13% vs 4%, P =.11), and profound thrombocytopenia <20, 000 (3.5% vs 0%, P =.33) with intravenous plus oral RPR 109891 compared with placebo. In 3 of 5 cases of profound thrombocytopenia, RPR 109891 had been interrupted because of bypass surgery, and a precipitous fall in platelet count occurred after the first postoperative oral dose. CONCLUSIONS: Intravenous RPR 109891 is a potent, predictable, dose-related platelet inhibitor. Oral RPR 109891 (17 years of age from the third National Health and Nutrition Examination Survey (NHANES III) database. Body iron stores reflected by serum ferritin levels rose in the late teens in men and after menopause in women. This rise was more rapid and maximum ferritin levels were greater for blacks than whites and Hispanics of comparable age and sex. The distribution of values for the serum ferritin differed from the percent transferrin saturation. CONCLUSIONS: Different patterns of iron accumulation exist according to age, sex, and race. Serum ferritin levels reflect graded, population-based differences in body iron stores, but the percentage of transferrin saturation does not. The hypothesis that iron accumulation may contribute to higher morbidity and mortality rates can be tested in clinical trials of calibrated reduction of body iron stores in defined disease settings. PMID- 10874270 TI - Association between depression and worse disease-specific functional status in outpatients with coronary artery disease. AB - BACKGROUND: The objective of this study was to determine if depression is associated with worse disease-specific functional status in patients with coronary artery disease. The study was designed as a cross-sectional survey and 3 month longitudinal cohort. METHODS AND RESULTS: The study took place in outpatient clinics of 3 Veterans Administration hospitals. All 7282 enrollees were surveyed and 4560 (62.6%) returned baseline questionnaires, including a screening instrument for depression. Thirty-nine percent (n = 1793) reported evidence of coronary artery disease and 1282 patients (71.5%) returned the Seattle Angina Questionnaire; 1025 patients (80%) completed a subsequent 3-month series of instruments. Main outcome measures used were the Seattle Angina Questionnaire, a valid, reliable, and responsive disease-specific functional status measure for patients with coronary disease, and the Mental Health Inventory, a mental health screening instrument from the Short Form-36. Mental Health Inventory evidence of depression was associated with significantly worse disease-specific functional status. Depressed patients had more physical limitation (mean difference in Seattle Angina Questionnaire score = 16.9, P <.001), more frequent angina (mean difference in Seattle Angina Questionnaire score = 9.5, P <.001), less satisfaction with their treatment for coronary artery disease (mean difference in Seattle Angina Questionnaire score = 9.9, P <.001), and lower perceived quality of life (mean difference in Seattle Angina Questionnaire score = 16.3, P <.001) than nondepressed patients. Frequency of depressive symptoms demonstrated an inverse relation with cardiac-specific functional status and when patients' depression status changed over time, so did their cardiac-specific health status. CONCLUSIONS: Depression is associated with significantly more physical limitation, more frequent angina, less treatment satisfaction, and lower perceived quality of life in outpatients with coronary artery disease. PMID- 10874271 TI - Heart diseases affecting the liver and liver diseases affecting the heart. AB - BACKGROUND: The association of cardiac and liver disorders has not been extensively outlined in the literature. METHODS: A survey of the MEDLINE database was performed to assess the current status of research regarding the association between cardiac and liver disorders. RESULTS: Combined cardiac and hepatic disorders occur in 3 different settings: heart diseases affecting the liver, liver diseases affecting the heart, and cardiac and hepatic disorders with joint etiology. The spectrum of heart diseases affecting the liver includes mild alterations of liver function tests in heart failure, cardiogenic ischemic hepatitis, congestive liver fibrosis, and cardiac cirrhosis. The liver diseases affecting the heart include complications of cirrhosis such as hepatopulmonary syndrome, portopulmonary hypertension, pericardial effusion, and cirrhotic cardiomyopathy as well as noncirrhotic cardiac disorders such as high-output failure caused by intrahepatic arteriovenous fistulae. Cardiac and hepatic disorders with joint etiology include infectious, metabolic, immune, vasculitic, and toxic disorders. We propose a practical approach to a diagnostic workup of combined cardiac and hepatic disorders based on recognizing the sequence of appearance of the cardiac and liver disease, presence of features of a multisystem disease, and presence of pathognomonic features. The evaluation of combined cardiac and hepatic disorders takes into consideration the expected benefit of treatment and the risks related to invasive procedures. Accordingly, investigations can be limited to ancillary tests for patients with congested liver and mild alterations of liver function tests, in cardiogenic ischemic hepatitis, patients with cardiac cirrhosis who are proposed for conservative treatment, and multisystem disease involving the heart and the liver. Conversely, comprehensive investigations are recommended when invasive therapeutic interventions are considered for the treatment of hepatopulmonary syndrome, portopulmonary hypertension, or arteriovenous fistulae. CONCLUSION: Classification of a patient to any of the 3 categories-heart diseases affecting the liver, liver diseases affecting the heart, and cardiac and hepatic disorders with joint etiology-permits the physician to narrow the span of the possible diagnoses and allows for a more simple workup. PMID- 10874272 TI - Collateral recruitment and "warm-up" after first exercise in ischemic heart disease. AB - BACKGROUND: Proposed mechanisms for "warm-up" after angina on first exercise include ischemic preconditioning and collateral recruitment. The aim of this study was to determine whether patients with ischemic heart disease and well developed coronary collateral vessels have a greater warm-up response than those with no visible collateral vessels. METHODS AND RESULTS: Fifteen patients with a total coronary occlusion and collateral vessels and 18 patients with a single coronary artery stenosis and no angiographically visible collateral vessels were studied. Warm-up was measured as the difference in ST depression on the second compared with the first of 2 sequential treadmill exercise tests separated by 10 minutes of rest. There was a trend for the duration of second exercise to increase more in patients with occlusion than in those with stenosis (+1.3 vs +0.54 minutes, respectively, P =.087). In both groups, ST depression was less on second exercise than on first exercise. The size of this decrease was greater in the occlusion group than in the stenosis group. ST depression at equivalent submaximal exercise decreased by 0.52 vs 0.19 mm, respectively (P =.049). The rate of increase in ST depression during exercise decreased by 1.08 versus 0. 55 mm/min, respectively (P =.034). These differences were less after adjustment for ST depression on first exercise (P =.11 and P =.063, respectively). CONCLUSIONS: The trend for a greater decrease in ST depression on second compared with first exercise in the patients with total coronary occlusion suggests that an increase in collateral flow is a mechanism for warm-up after first exercise in ischemic heart disease. PMID- 10874273 TI - Prodromal unstable angina in acute myocardial infarction: prognostic value of short- and long-term outcome and predictor of infarct size. AB - BACKGROUND: An estimated 50% of patients with myocardial infarction have prodromal unstable angina. There is controversy over whether prodromal unstable angina identifies a group of patients at lower risk of short- and long-term death and the clinical importance of recording this event. METHODS: Of 207 patients enrolled at a single Global Utilization of Streptokinase and Tissue Plasminogen Activator for Occluded Coronary Arteries (GUSTO-I) site, 196 survived the 24 hours after presentation, achieved peak creatine kinase MB concentrations, and were classified as having either abrupt symptom onset or prodromal unstable angina in the 2 weeks before myocardial infarction. Creatine kinase MB peak was used to categorize infarct size as aborted myocardial infarction, minor myocardial damage, or extensive myocardial injury. Follow-up was performed at 24 hours, 30 days, 1 year, and 5 years. Multiple variables, including prodromal unstable angina, time to treatment, age, sex, previous infarction and infarct location, were analyzed for predicting infarct size. Also, these variables plus peak creatine kinase MB level and a combined variable of prodromal unstable angina and peak creatine kinase MB level were examined for predicting survival. RESULTS: Mortality rate was 2.5% within 24 hours, 9.0% at 30 days, 13.5% at 1 year, and 27.1% at 5 years. Patients categorized as either aborted infarction or minor myocardial damage were significantly more likely to have prodromal unstable symptoms (81.3% vs 51.2%, P <.001) and better survival at each follow-up period. Prodromal presentation was the most significant predictor of infarct size category (P =.001). Five-year survival was predicted by age (P <.0001), peak creatine kinase MB level (P =.007), infarct location (P =.009), the combined variables (P =.029), and prodromal unstable angina (P =.017). Prodromal unstable angina had the highest odds ratio for 5-year survival at 3.83 (95% confidence interval 1.27-11.47). CONCLUSIONS: Prodromal unstable angina is a strong predictor of infarct size and survival. Recognizing prodromal unstable angina is important for clinically assessing prognosis. PMID- 10874274 TI - Histopathologic and clinical characterization of cardiac myxoma: review of 53 cases from a single institution. AB - BACKGROUND: Cardiac myxomas have varying clinical presentation, uncertain histogenesis, and debatable immunohistochemical profile. A few malignant cases have been previously reported. METHODS: Fifty-three consecutive cardiac myxomas were histologically investigated and results compared with clinical data. The main goal of the study was to investigate the immunohistochemical differentiation and the clinicopathologic correlations. RESULTS: Stromal cells were characterized by the expression of the von Willebrand factor endothelial marker (12 of 53 cases) and diffuse cytoplasmic neuropeptides such as protein gene product 9.5 (50 of 53 cases), S100 protein (47 of 53) and neuron-specific enolase (30 of 53), all of which were expressed in 30 (57%) of 53 tumors. Stromal cells did not show endocrine granules, epithelial, or smooth muscle immunoreactivity. Non-cardiac related symptoms were observed in 7 of 53 patients and promptly disappeared after tumor excision; median values and percentages of total immunoreactivity scores for neuropeptides were higher in these 7 cases, but data analysis showed no statistical significance. Glands were detected in 2 myxomas, and they showed epithelial (cytokeratins and carcinoembryonic antigen), protein S100, and neuron specific enolase immunoreactivity; this pattern has been previously detected in human gut. All tumors showed benign behavior, and no mitosis was detected. CONCLUSIONS: The results of this study support the hypothesis that stromal cells originate from multipotent mesenchyme capable of neural and endothelial differentiation; rare myxoma glands would represent entrapped foregut rests. A correlation could exist between neuroendocrine differentiation and non-cardiac related symptoms. PMID- 10874275 TI - Chelation therapy for coronary heart disease: An overview of all clinical investigations. AB - BACKGROUND: Chelation therapy is popular in the United States. The question of whether it does more good than harm remains controversial. AIM: The aim of this systematic review was to summarize all the clinical evidence for or against the effectiveness and efficacy of chelation therapy for coronary heart disease. METHODS: A thorough search strategy was implemented to retrieve all clinical investigations regardless of whether they were controlled or uncontrolled. RESULTS: The most striking finding is the almost total lack of convincing evidence for efficacy. Numerous case reports and case series were found. The majority of these publications seem to indicate that chelation therapy is effective. Only 2 controlled clinical trials were located. They provide no evidence that chelation therapy is efficacious beyond a powerful placebo effect. CONCLUSION: Given the potential of chelation therapy to cause severe adverse effects, this treatment should now be considered obsolete. PMID- 10874276 TI - Observations on the transition from intermittent to permanent atrial fibrillation. AB - BACKGROUND: Quantitative data on the frequency with which transition from intermittent to permanent atrial fibrillation occurs are lacking. We conducted this study to determine the proportion of patients with intermittent atrial fibrillation who progress to permanent atrial fibrillation and to investigate baseline clinical characteristics that might predict such a progression. METHODS: This retrospective cohort study included 231 patients who were seen with intermittent atrial fibrillation at a university hospital-based clinic from January 1978 through December 1997. Patients' medical records and electrocardiograms were reviewed and data were collected for all clinic visits through May 1998. The proportion of patients who remained free of transition from intermittent to permanent atrial fibrillation was calculated by the Kaplan-Meier method. A Cox proportional hazards model was used to determine the effect of some baseline characteristics on this transition. RESULTS: The number of patients who remained free of transition from intermittent to permanent atrial fibrillation was 92% (95% confidence interval 88%-96%) at 1 year and 82% (95% confidence interval 75%-88%) at 4 years. Among 5 baseline characteristics (age, sex, structural heart disease, atrial fibrillation at presentation, and use of an antiarrhythmic medicine before presentation), the 2 significant predictors of progression from intermittent to permanent atrial fibrillation were age (P =.0003) and being in atrial fibrillation at presentation (P =.0006). The hazard ratio associated with 10 years of advancing age was 1.82 (95% confidence interval 1.31-2.51), and the hazard ratio associated with atrial fibrillation at presentation was 3.56 (95% confidence interval 1.73-7.34). CONCLUSIONS: Approximately 18% of patients who had intermittent atrial fibrillation were permanently in atrial fibrillation after 4 years of follow-up. Age and being in atrial fibrillation at presentation were the only 2 important clinical variables identified in predicting such a progression. PMID- 10874277 TI - A novel mutation in KVLQT1, L122P, found in a family with autosomal dominant long QT syndrome. AB - BACKGROUND: Linkage and mutation analysis in long QT syndrome kindreds has demonstrated locus heterogeneity, with causative mutations reported in at least 5 different genes, including KVLQT1. METHODS AND RESULTS: A 12-year-old male proband with recurrent syncope and a prolonged QT interval underwent clinical assessment and exercise testing along with 3 affected and 3 unaffected family members. The coding regions of 5 putative transmembrane segments (S2-S6) and a putative pore region of the KVLQT1 gene for the proband were amplified with the polymerase chain reaction. DNA sequencing of the KVLQT1 gene of the proband revealed a T-->C transversion at the second position of codon 122, which predicted a substitution of proline for leucine (L122P). By using restriction analysis, the L122P was found to be co-segregated with the electrocardiographic abnormalities in the nuclear family. Although the patient's mother was heterozygous for L122P, neither maternal grandparent was a carrier, suggesting that the mutation arose spontaneously. In comparison, there was a complete absence of the mutation in 1336 alleles from 668 normal individuals of 6 different ethnic backgrounds. CONCLUSION: The KVLQT1 L122P mutation is a rare novel mutation that probably arose spontaneously in this family, leading to long QT syndrome. PMID- 10874278 TI - Efficacy of anticoagulation in resolving left atrial and left atrial appendage thrombi: A transesophageal echocardiographic study. AB - BACKGROUND: Transesophageal echocardiography (TEE) is the gold standard for evaluation of the left atrium and the left atrial appendage (LAA) for the presence of thrombi. Anticoagulation is conventionally used for patients with atrial fibrillation to prevent embolization of atrial thrombi. The mechanism of benefit and effectiveness of thrombi resolution with anticoagulation is not well defined. METHODS AND RESULTS: We used a TEE database of 9058 consecutive studies performed between January 1996 and November 1998 to identify all patients with thrombi reported in the left atrium and/or LAA. One hundred seventy-four patients with thrombi in the left atrial cavity (LAC) and LAA were identified (1.9% of transesophageal studies performed). The incidence of LAA thrombi was 6.6 times higher than LAC thrombi (151 vs 23, respectively). Almost all LAC thrombi were visualized on transthoracic echocardiography (90.5%). Mitral valve pathology was associated with LAC location of thrombi (P <.0001), whereas atrial fibrillation or flutter was present in most patients with LAA location of thrombi. Anticoagulation of 47 +/- 18 days was associated with thrombus resolution in 80.1% of the patients on follow-up TEE. Further anticoagulation resulted in limited additional benefit. CONCLUSIONS: LAC thrombi are rare and are usually associated with mitral valve pathology. Transthoracic echocardiography is effective in identifying these thrombi. LAA thrombi occur predominantly in patients with atrial fibrillation or flutter. Short-term anticoagulation achieves a high rate of resolution of LAA and LAC thrombi but does not obviate the need for follow-up TEE. PMID- 10874279 TI - Effect of exercise training on heart rate variability in patients with new-onset left ventricular dysfunction after myocardial infarction. AB - BACKGROUND: Cardiac rehabilitation with exercise training alters sympathovagal control of heart rate variability (HRV) toward parasympathetic dominance in patients after acute myocardial infarction (MI). However, its effects on HRV in patients after MI with new-onset left ventricular dysfunction are yet unknown. We aimed to investigate the effects of 8 weeks of supervised, high-intensity exercise training on time- and frequency-domain measures of HRV in this selected patient population. METHODS AND RESULTS: Twenty-five men with an acute MI and a low ejection fraction were randomly assigned to enter or not to enter a training program in a regional rehabilitation center. HRV was evaluated before and after 1 and 2 months of training and at 12 months. Maximal exercise testing with respiratory gas exchange was performed at baseline and after training. Resting heart rate decreased (P <. 01) and the percentage of R-R intervals differing >50 ms from the preceding one (pNN50) increased (P <.05) after training. The standard deviation of R-R intervals (SDRR) tended to increase, but frequency-domain indexes remained unchanged. There was a significant decrease in SDRR (P <.05) and high-frequency power (P <.01) at 12 months in untrained patients. Exercise time increased by 38% and maximal oxygen uptake increased by 29% in the training group (P <. 01). CONCLUSIONS: Despite beneficial effects on clinical variables, exercise training did not markedly alter HRV indexes. A significant decrease in SDRR and high-frequency power in the control group suggests an ongoing process of sympathovagal imbalance in favor of sympathetic dominance in untrained patients after MI with new-onset left ventricular dysfunction. PMID- 10874280 TI - Race, baseline characteristics, and clinical outcomes after coronary intervention: The New Approaches in Coronary Interventions (NACI) registry. AB - BACKGROUND: The impact of race and sex on clinical outcomes after percutaneous coronary interventions remains incompletely understood. Specific data on patient demographics, lesion characteristics, and outcomes of black versus white patients are poorly described. To further evaluate these issues, we analyzed the New Approaches in Coronary Interventions (NACI) registry. METHODS: Patients (200 black, 4279 white) undergoing coronary interventions in the NACI trial were compared. A Cox proportional hazards model was used to determine which baseline demographics were independent risk factors for the combined end point of death, Q wave myocardial infarction, and coronary artery bypass grafting at 1 year. RESULTS: Black patients were significantly younger (age 59 +/- 11 vs 63 +/- 11 years; P <.001), more often obese (29.6 +/- 6 vs 27.5 +/- 4.8 kg/m(2); P <.001), female (50% vs 34%; P <.001), diabetic (34% vs 21%; P <.001), and hypertensive (71% vs 52%; P <.001). Black patients were significantly more likely to have single-vessel disease (48% vs 40%; P <.05) and less likely to have undergone coronary artery bypass grafting (26% vs 34%; P <.05). Blacks were significantly more likely to have a discrete lesion (85% vs 62%; P <. 001) with less thrombus (7% vs 12%; P <.05), tortuosity (17% vs 25%; P <.05), and an ulcerated appearance (5% vs 10%; P <.05). Despite these significant baseline differences, no significant difference was seen in the procedural success (80% vs 82%) or major adverse events (death, Q-wave myocardial infarction, any revascularization) at 1 year (39% vs 34%). Predictors of adverse events for white patients included diabetes (relative risk [RR] = 1.24; confidence intervals [CI], 1.0-1.5) and high risk status (RR = 1.58; CI, 1.26-1. 91). Predictive characteristics of adverse events for black patients included only sex (RR = 3.45; CI, 1.27-9.35; P =.02). CONCLUSIONS: There are significant differences in baseline characteristics of black patients compared with white patients. Despite these differences in traditional risk factors, they do not affect procedural success or 1-year outcome. In black patients, only sex predicted adverse events. Additional investigation is required to understand the mechanisms for this difference. PMID- 10874281 TI - A1166C polymorphism of the angiotensin II type 1 receptor gene and risk of adverse events after coronary catheter interventions. AB - BACKGROUND: Contradictory reports exist concerning the role of the angiotensin II type 1 receptor A1166C polymorphism as a coronary risk factor. Moreover, it is unknown whether the A1166C polymorphism is associated with thrombotic complications after coronary catheter interventions. METHODS: We investigated the role of the A1166C polymorphism as a risk factor in 1000 patients with coronary artery disease (CAD) and in 1000 age- and sex-matched controls. A total of 649 patients receiving interventions (270 coronary angioplasty, 102 atherectomy, and 277 stenting) were investigated for a 30-day composite end point including target vessel revascularization, myocardial infarction, or death. RESULTS: The composite end point was reached by 42 patients (6.5%) without evidence that the C allele was associated with excess procedural risk (odds ratio 0.93; 95% confidence interval 0.79-1.75; P =.82). Further analyses by device failed to show linkage with adverse events complicating coronary angioplasty, atherectomy, and stenting. Moreover, in the entire CAD group (n = 1000), the polymorphism even showed a trend to underrepresentation (odds ratio 0.83; 95% confidence interval 0.69-1. 004, P =.054). CONCLUSIONS: These results indicate that the A1166C polymorphism neither represents a risk factor for adverse events complicating coronary interventions nor seems to have significant impact on further long-term processes such as development and severity of CAD. PMID- 10874282 TI - Esmolol versus diltiazem in the treatment of postoperative atrial fibrillation/atrial flutter after open heart surgery. AB - BACKGROUND: Supraventricular tachyarrhythmias are common after open heart surgery. Possible causative factors for these arrhythmias include operative trauma, atrial ischemia, electrolyte imbalances, pericardial irritation, and excess catecholamines. Two agents commonly used to control ventricular rate in atrial fibrillation or atrial flutter (AF/AFL) are beta-blockers and calcium channel blockers. METHODS AND RESULTS: This randomized study was designed to compare the safety and efficacy of intravenous diltiazem versus intravenous esmolol in patients with postoperative AF/AFL after coronary bypass surgery and/or valve replacement surgery. A comparative cost analysis was also performed. Thirty patients received either esmolol (n = 15) or diltiazem (n = 15) for AF/AFL. During the first 6 hours of treatment, 66.6% of esmolol-treated patients converted to sinus rhythm compared with 13.3% of the diltiazem-treated patients (P <.05). At 24 hours, 66.6% of the diltiazem group converted to SR compared with 80% of the esmolol group (not significant). Drug-induced side effects, time to rate control (<90 beats/min), number of patients requiring cardioversion, and length of hospitalization were similar for the two groups. The drug cost/successfully treated patient for esmolol versus diltiazem was $254 versus $437 at 6 hours and $529 versus $262 at 24 hours. CONCLUSIONS: Although this is a small study, it suggests that esmolol is more effective in converting patients to normal sinus rhythm than diltiazem during the initial dosing period. No differences in conversion rates were observed between the two groups after 24 hours. Additional studies are needed to confirm whether esmolol is the initial drug of choice in patients with postoperative AF/AFL after coronary bypass surgery. PMID- 10874283 TI - Highlights from the American College of Cardiology 49th Annual Scientific Sessions: March 12 to 15, 2000. PMID- 10874284 TI - Sex offenders: part two/three. Public policy. PMID- 10874285 TI - Sex crimes and the new punitiveness. AB - In the last few years, new ways of punishing sex offenders have been introduced in many modern societies. However, these sanctions have a broader significance than this: they are part of a broader set of penal arrangements-directed at the criminal population as a whole-which represents a new punitiveness. This seems to be moving the direction of legal punishment beyond the established parameters that had hitherto been set for it in modern society. This had involved punishment becoming increasingly administered by penal bureaucracies, to the exclusion of the general public, being influenced by the opinion of penal experts, and becoming more tempered, consistent and purposeful in form. Sanctions that did not fit these criteria faded out of modern penality. The indeterminate prison sentence was introduced at its outer limits as a residual measure of control to be used against those offenders-frequently sex criminals-for whom the existing penal framework was thought inappropriate. Even so, by the 1970s, these special penal measures were falling into disuse. However, the new punitiveness has not only given new life to them, but has also led to the introduction of measures which seem to reverse or move beyond modernpenal parameters. The article argues that the reasons for these shifts lie in the profound economic and social changes that have taken place in Western societies over the course of the last two decades or so. PMID- 10874286 TI - The future of involuntary civil commitment in the U.S.A. after Kansas v. Hendricks. AB - This article examines new sexual predator commitment laws enacted recently in the United States to civilly commit dangerous sex offenders after they have served their prison sentences. It then examines Kansas v. Hendricks, a Supreme Court case that upheld these laws as constitutionally permitted. The article next describes the broad parameters that demarcate the government's civil commitment authority identified by the Supreme Court in that case. The author concludes that Hendricks establishes that the state has expansive civil commitment power much greater than our previous understanding. The government may use civil commitment solely to protect the public from dangerous individuals without proving a medically recognized mental disorder, recent evidence of dangerousness, or a treatment purpose or possibility. Moreover, this quarantine system may be justified by proving the same unlawful behavior for which the individual has already been criminally punished. PMID- 10874287 TI - Getting it right: the trial of sexual assault and child molestation cases under federal rules of evidence 413-415. AB - Congressional enactment of Federal Rules of Evidence 413-415 changed centuries of the law which had excluded evidence by the state that the defendant had committed other bad acts and was therefore the sort of person who would be more likely to commit the act charged. The passage of Rules 413-415 opens the door to this type of character evidence in sexual assault and child molestation cases and requires trial judges to assess the probative value of this propensity evidence offered. Yet, neither these rules nor their legislative history offer much guidance in this assessment. This article offers guidance to trial judges and lawyers to assess the probative value of propensity evidence offered under these rules. PMID- 10874288 TI - Proving sexual assault. AB - The framework provided by therapeutic jurisprudence is used to examine the rules of evidence that govern sexual assault trials. The concern is with the impact of the trial process on the accusing witness. In the first part a personal narrative is used to put the legal rules in context. Psychological sources are used in discussing the reasons for presenting such a narrative. In the second part, the effect of statutory changes to the common law rules as effected in NSW in 1981 is analyzed. Drawing upon the Heroines of Justice report the paper suggests that the intention of reducing the trauma of the victim has not yet been achieved. It is suggested that this failure is not unique. A change in attitude on the part of those involved in the trial process is necessary, and the hope is that this paper might contribute to such a change. PMID- 10874289 TI - Psychopathy, criminal responsibility, and civil commitment as a sexual predator. AB - Recent judicial decisions regarding commitment under sexual predator statutes and commentary addressing the legal significance of psychopathy provide an interesting opportunity to reflect upon the exculpatory significance of psychopathy and the appropriate relationship between criminal conviction and police power civil commitment. This paper examines the legal significance of psychopathy for the purposes of criminal responsibility and of civil commitment under sexual predator statutes. By examining the significance of psychopathy for each of these legal institutions, it clarifies our understanding of the legal significance of psychopathy and of the relationship between these institutions. This process illuminates the defensible functions and boundaries of each institution and clarifies the nature of the impairment that should qualify an individual for confinement by each. This analysis interprets criminal conviction and police power commitment, including sexual predator commitment, as integrated institutions of social control intended to provide a coherent approach to psychopaths as well as to others who require state intervention under the police power. PMID- 10874290 TI - An examination of the assumptions of specialization, mental disorder, and dangerousness in sex offenders. AB - Sex offenders have been singled out for differential treatment by the legal and mental health systems. This article attempts to inform law reform efforts and criminal justice mental health policy by examining the assumptions underlying differential legal and mental health treatment of sex offenders. These assumptions include the theories that sex offenders are mentally disordered and in need of treatment, specialists in sex crimes, and more dangerous than other criminal offenders. Empirical findings demonstrate that sex offenders are not specialists in sex crimes and are not mentally disordered. Examination of past research suggests that sex offenders are not at more risk than other criminal offenders to commit future sex crimes. Implications of research findings for selective prosecution of sex crime cases, mental health policy, sex offender legislation, and predictions of future dangerousness are discussed. Proposals for future research needs and law reform are presented. PMID- 10874291 TI - A Brunswikian evolutionary-developmental theory of adolescent sex offending. AB - A Brunswikian Evolutionary-Developmental model was developed to relate the sex offending behavior of adolescents to other forms of social deviance, tracing a history of repeated frustration and failure in various competitive sexual strategies and escalation to more extreme means of obtaining sexual gratification. Four hypothetical constructs were proposed as stages in the development of sexual criminality: (1) Psycho-Social Deficiency (PSD); (2) Non Criminal Sexuality (NCS); (3) Non-Sexual Criminality (NSC); and (4) Sexual Criminality (SC). Significant direct and indirect pathways led from PSD to SC through both NCS and NSC, each time facilitated by an interaction with PSD. Although the causal orders between stages remain equivocal, the current results are consistent with our theory and establish the heuristic value of our theoretical approach, providing empirical support for otherwise counterintuitive predictions. This interpretation also offers hope for focusing preventative intervention at one major root cause of this unfortunate cascade of consequences, Psycho-Social Deficiency. PMID- 10874292 TI - Sentencing sex crimes against children: an empirical and policy analysis. AB - This study investigated factors that contributed to the sentencing outcomes of 387 sex crimes against children who were prosecuted in a large East Coast city. Hypothesized variables that were indexed to predict sentencing included several offense, victim, and perpetrator characteristics. The findings revealed that individual victims' experiences are generally less predictive of sentencing outcomes than perpetrators' characteristics, that sentences generally tend to be lenient, that intra-family and stranger abuse seem to be taken equally seriously, and that the criminal justice system does seem to incarcerate those society is most worried about-persistent predators who abuse several children. The article ends with suggestions for further research and policy development. PMID- 10874293 TI - Sex offender commitments in Minnesota: a descriptive study of second generation commitments. AB - Civil commitment laws intended to prevent sexual violence are being considered and enacted by many state legislatures. In their use of "preventive detention" to lock up the "most dangerous," the laws are highly controversial legally and morally. As well, critics raise questions about the extent to which the laws advance their stated goals. Considering the growing expense of these programs, evaluation efforts based on an empirical foundation are imperative. This study collected data on all 116 men committed under Minnesota's program during the period 1975-1996. The authors report descriptive summary data on admission rates and population trends, demographic characteristics of detainees and victims, pre commitment criminal and mental health histories, and current status of detainees. The article also compares the committed group with sex offenders in the state's correctional system. The article describes the history and functioning of sex offender commitment programs, and draws implications for policy-makers from the data presented. PMID- 10874294 TI - Sex offender community notification: managing high risk criminals or exacting further vengeance? AB - The current trend in dealing with convicted sex offenders is to impose long prison sentences followed by stringent release conditions. Added to this practice has been the policy of making such offenders who have been returned to society more visible to the public. In state after state, sex offender community notification laws have been passed, enabling communities to be put on notice that a convicted sex offender has become a resident. To date there has been little empirical evidence regarding the impact of these laws on managing sex offenders in the community. This study focuses on the social and psychological effects of community notification on sex offender reintegration within those communities where notification has occurred. Data are derived from face-to-face interviews with 30 convicted sex offenders, residing in various locations throughout Wisconsin, who were the subjects of community notification. The findings indicate that community notification can have a critical impact on the minimum essentials needed for the reintegration of offenders within the community. What is proposed is a reintegrative approach which suggests that stable housing and employment would mitigate the disruptive and antitherapeutic effects of community notification. PMID- 10874296 TI - John F. Edens, Ph.D., norman G. Poythress, Ph.D., and scott O. Lilienfeld, Ph.D., identifying inmates at risk for disciplinary infractions: A comparison of two measures of psychopathy. Behavioral sciences & the law 17(4) 1999, 435-443 AB - The original article to which this erratum refers was published in Behavioral Sciences & the Law 17(4) 1999, 435-443 PMID- 10874295 TI - Sex offender community notification: examining the importance of neighborhood meetings. AB - Within the last decade, federal and state laws have been passed authorizing or requiring the notification of local residents that a convicted sex offender will be released and living in their neighborhood. The community meeting method of notifying neighborhood residents, although the subject of extensive news media attention, has been largely overlooked by empirical researchers. This study focuses on the experience of residents who attend such meetings and how that experience factors into a collective response on the part of the community. Data are derived from attendee surveys and recorded observations taken at all community notification meetings held throughout Wisconsin during a nine month period. The findings suggest that community notification meetings, if properly conducted, can perform an important role in managing the behavior of known sex offenders in the community. However the decision to notify and involve the public in an informal network of neighborhood surveillance may come at the cost of increased community anxiety. This anxiety is related to how the attendees were notified of the meeting, how clearly the purpose of the meeting was conveyed, and how organized the meeting appeared to the audience. Suggestions on how to more effectively utilize the community meeting method of notification are presented. PMID- 10874297 TI - PHEXdb, a locus-specific database for mutations causing X-linked hypophosphatemia. AB - X-linked hypophosphatemia (XLH) is a dominant disorder of phosphate (Pi) homeostasis characterized by growth retardation, rachitic and osteomalacic bone disease, hypophosphatemia, and renal defects in Pi reabsorption and vitamin D metabolism. The gene responsible for XLH was identified by positional cloning and designated PHEX (formerly PEX) to depict a Phosphate regulating gene with homology to Endopeptidases on the X chromosome. To date, 131 mutations in the PHEX gene have been reported. We undertook to centralize information on mutations in the PHEX gene by establishing a database search tool, PHEXdb (http://data.mch.mcgill.ca/phexdb). This site is dedicated to the collection and distribution of information on PHEX mutations, and is accessible to the scientific community. PHEXdb provides a submission form to allow the addition of newly identified mutations in the PHEX gene. Users can search the database by mutation, phenotype, and authors who have published or submitted mutations. The PHEXdb home page includes links to information pages, which refer to recent publications on PHEX, XLH, and murine Hyp and Gy homologues, and to other web pages relevant to XLH. This resource will facilitate the identification of PHEX structure-function relationships and phenotype-genotype correlations. PMID- 10874298 TI - Mutation analysis of the GJB2 (connexin 26) gene by DGGE in Greek patients with sensorineural deafness. AB - The GJB2 (connexin 26) gene, one of the major genes responsible for autosomal recessive deafness, has been investigated previously by a variety of techniques, including PCR-SSCP and sequencing of the entire gene for screening of unknown mutations, and allele-specific PCR, ASO, and PCR-mediated site-directed mutagenesis for the detection of the common mutation 35delG. Here, we present the development of a DGGE method for the characterization of the full spectrum of mutations in the GJB2 gene. The GJB2 cDNA and flanking sequences were amplified in three overlapping segments. We screened 26 Greek patients with prelingual, sensorineural deafness, where syndromic forms and environmental causes of deafness had been excluded. The 35delG mutation was detected in 28 chromosomes (53.8%), while another three sequence variations accounted for 7.6% of the alleles. The sequence variation R127H, previously described in a few Spanish and Balkan patients, was detected in two patients as the sole mutation. A novel sequence variation, K224Q, was identified as the sole mutation in one patient. Use of this approach may contribute to the full description of mutations in this important deafness gene. PMID- 10874299 TI - Private beta- and gamma-sarcoglycan gene mutations: evidence of a founder effect in Northern Italy. AB - Autosomal recessive muscular dystrophies called "sarcoglycanopathies" result from mutations in the genes encoding alpha-, beta-, gamma-, or delta-sarcoglycan complex components. The present study involved six unrelated families from Northern Italy showing mutations in the beta- or gamma-sarcoglycan genes. An 8 bp duplication in the beta-sarcoglycan gene and 1 bp insertion in the gamma sarcoglycan gene occur with high frequency in our population. These mutations have never been reported thus far in other countries. Many patients are homozygotes for a single mutation, although they derived from non-consanguineous marriages. We suggest that these alleles are "private" mutations of this geographical region. A panel of highly informative microsatellite markers that map in the beta- and gamma-sarcoglycan gene locus was used to assess the haplotypes among affected patients and control population, in order to test the presence of linkage disequilibrium. We found that the 8 bp duplication in the beta-sarcoglycan gene and the 1 bp insertion in the gamma-sarcoglycan gene are in linkage disequilibrium with neighbouring polymorphisms. The recurrence of specific sarcoglycan mutations in Northern Italy is probably due to a founder effect, combined with a relative genetic isolation. PMID- 10874300 TI - A case of methemoglobinemia type II due to NADH-cytochrome b5 reductase deficiency: determination of the molecular basis. AB - Clinical, biochemical and molecular findings in a patient with methemoglobinemia type II are described. Furthermore, a comparison between methemoglobinemia type I and type II, both caused by a deficiency of NADH-cytochrome b5 reductase (b5R), is made. Although the clinical pictures of type I and II are strikingly different, mutations in the diaphorase (DIA1) gene located on chromosome 22 have been described in both types. In the present patient, two newly identified mutations, both leading to a stop codon in exon 4 (Gln77Ter) and in exon 6 (Arg160Ter), were found. Identification of different mutations at different positions in the DIA1 gene might shed light on the clinical and biochemical differences between methemoglobinemia type I and type II. PMID- 10874301 TI - Genetic heterogeneity in Peutz-Jeghers syndrome. AB - LKB1, the human gene encoding a serine threonine kinase, was recently identified as a susceptibility gene for Peutz-Jeghers syndrome (PJS), a disease characterized by the constellation of intestinal hamartomata, oral mucocutaneous hyperpigmentation, and an increased risk for gastrointestinal as well as extraintestinal malignancies. To date, the majority of individuals with PJS have been found to have genetic alterations in LKB1, most of which result in protein truncation. Additionally, linkage analyses have suggested a modicum of genetic heterogeneity, with the majority of PJS families showing linkage to the LKB1 locus. In this study, we evaluated five kindreds with greater than two affected family members, five PJS probands with only one other affected family member, as well as 23 individuals with sporadic PJS for mutations within the LKB1 gene. Conformation sensitive gel electrophoresis was utilized for the initial screen, followed by direct sequence analysis for characterization. Long-range PCR was used for the detection of larger genetic insertions or deletions. Mutation analysis revealed genetic alterations in LKB1 in two probands who had a family history of PJS. LKB1 mutations were detected in only four of the remaining 23 cases of sporadic PJS. These data suggest the presence of significant genetic heterogeneity for PJS and the involvement of other loci in this syndrome. PMID- 10874302 TI - The human factor IX gene as germline mutagen test: samples from Mainland China have the putatively endogenous pattern of mutation. AB - Germline mutations are the major source of genetic variation that allows a species to evolve over time but at the cost of Mendelian disease and genetic predisposition to multifactorial diseases. Previous analyses have revealed that the pattern of germline mutations in the factor IX gene (F9) is similar among a variety of ethnically and geographically diverse populations and compatible with the ancient pattern that has shaped the mammalian genome. Here, we compare the pattern of germline mutation in a population of hemophilia B patients from Mainland China (n=66) to that in U.S. Caucasians, Blacks, and Mexican Hispanics and stratify by disease severity and ethnicity. The similar pattern of germline mutation in all ethnic groups studied to date provides additional data compatible with the inference that endogenous processes predominate in germline mutations. PMID- 10874303 TI - The spectrum of mutations, including four novel ones, in the thiamine-responsive megaloblastic anemia gene SLC19A2 of eight families. AB - Thiamine responsive megaloblastic anemia (TRMA) is an autosomal recessive disorder with a triad of symptoms: megaloblastic anemia, deafness, and non-type 1 diabetes mellitus. Occasionally, cardiac abnormalities and abnormalities of the optic nerve and retina occur as well. Patients with TRMA often respond to treatment with pharmacological doses of thiamine. Recently, mutations were found in patients with TRMA in a thiamine transporter gene (SLC19A2). We here describe the mutations found in eight additional families. We found four novel mutations and three that were previously described. Of the novel ones, one is a nonsense mutation in exon 1 (E65X), two are missense mutations in exon 2 (S142F, D93H), and another is a mutation in the splicing donor site at the 5' end of intron 4 (C1223+1G>A). We also summarize the state of knowledge on all mutations found to date in TRMA patients. SLC19A2 is the first thiamine transporter gene to be described in humans. Reviewing the location and effect of the disease causing mutations can shed light on the way the protein functions and suggest ways to continue its investigation. PMID- 10874304 TI - The spectrum of patched mutations in a collection of Australian basal cell carcinomas. AB - Inactivating mutations in the human patched (PTCH) gene have been identified in both familial and sporadic basal cell carcinomas (BCCs). In some tumors mutations have been detected in both alleles thereby supporting the role of PTCH as a tumor suppressor gene. We have analyzed 22/23 coding exons of PTCH for mutations in 44 sporadic BCCs, and detected 10 novel mutations in nine tumors. In two of the mutant tumors the remaining allele was inactivated by loss of heterozygosity. Five novel PTCH polymorphisms were also identified. Most of the variations found were C>T substitutions at dipyrimidine sites, supporting previous studies which indicate a role for ultraviolet-B in the genesis of sporadic BCCs. PMID- 10874305 TI - 3' polymorphisms of ETS1 are associated with different clinical phenotypes in SLE. AB - A microsatellite repeat polymorphism was identified in the 3' flanking region of the human ETS1 gene. Sequencing revealed two CA repeat segments in close proximity. Seven different alleles comprising various combinations of CA repeat units were identified in a healthy control population. Because ETS1 plays a role in lymphocyte development and function, apoptosis, and inflammation, we examined whether any of these polymorphisms were associated with a systemic inflammatory condition, systemic lupus erythematosus (SLE). Inheritance of this disease is polygenic and a recent genome-wide screen for SLE susceptibility loci revealed linkage with chromosome 11q14-23, the region in which the ETS1 gene lies. This region has also been identified as a general autoimmune susceptibility region. None of the seven distinct ETS1 alleles appeared statistically more frequently in SLE patients than controls, however, two alleles were associated with particular clinical manifestations. Allele 1 is associated with discoid lesions and allele 7 is associated with vasculitis. While this polymorphism does not directly affect the coding region of ETS1, it may be a marker for overexpression of a particular isoform or inheritance of another polymorphism which does affect function. These data suggest that ETS1 may be involved in the phenotypic expression of systemic lupus erythematosus. PMID- 10874306 TI - Isolated central form of tetrahydrobiopterin deficiency associated with hemizygosity on chromosome 11q and a mutant allele of PTPS. AB - 6-Pyruvoyl-tetrahydropterin synthase (PTS or PTPS) is involved in tetrahydrobiopterin (BH(4)) biosynthesis, the cofactor for various enzymes including the aromatic amino acid hydroxylases. Inherited PTPS deficiency is a heterogeneous disease with different phenotypes leading to BH(4) depletion. The severe form of PTPS deficiency causes hyperphenylalaninemia and monoamine neurotransmitter deficiency, whereas the mild form gives rise to hyperphenylalaninemia only. From 228 patients with PTPS deficiency at least 32 different mutant alleles have been identified on its corresponding gene, located on chromosome 11q22.3-q23.3. Here we describe a new allele from a child with PTPS deficiency who exhibited a mild but transient form of hyperphenylalaninemia, yet was deficient in CSF monoamines. The patient was found to carry, on her genomic DNA and cDNA, a homozygous A>G transition, leading to PTPS codon alteration Tyr99 to Cys (Y99C). The mother and several members of the maternal family were carriers of the Y99C allele, also verified by the reduced PTPS enzyme activity in erythrocytes. By cytogenetic, molecular, and FISH analyses, a de novo deletion spanning from 11q14 to 11q23.3 on the patient's paternal chromosome was mapped, establishing hemizygosity of the Y99C allele. The PTPS mutation observed in this patient generates a novel phenotype with an apparently isolated central form of BH(4) deficiency. PMID- 10874307 TI - Evaluation of enzymatic mutation detectiontrade mark in hereditary nonpolyposis colorectal cancer. AB - In hereditary nonpolyposis colorectal cancer (HNPCC), the majority of reported mutations are dispersed throughout the 35 exons of the two principal susceptibility genes, MLH1 and MSH2, and because of this complexity, rapid mutation screening methods are required. The aim of this study was to evaluate the sensitivity of the Enzymatic Mutation Detection (EMD) assay in HNPCC using genomic DNA samples with known gene alterations in MLH1 and MSH2. The EMD assay relies upon the enzyme T4 Endonuclease VII recognizing and cleaving DNA mismatches, created when a PCR product containing a sequence alteration is hybridized with a wild type probe. A total of 68 different sequence variants from 30 exons were analyzed. The EMD assay was able to detect 62 of the 68 sequence variants (91%) with the majority showing strong cleavage products. One of the advantages of the EMD assay over other mutation screening techniques is that larger fragments can be analyzed in a single assay. No specialized equipment is required and one set of primers is sufficient for radioactive detection of the cleavage products. This method can be adapted to use fluorescent dye-labelled primers and may be automated to detect mutations accurately and rapidly in a large number of samples. One new MLH1 mutation (418delA) and two novel MSH2 mutations (1A>C; 227-228delAG) were also detected in HNPCC patients screened using this method. PMID- 10874308 TI - Signature-based analysis of MET proto-oncogene mutations using DHPLC. AB - Research tools which improve mutation detection, SNP discovery, and allele characterization will facilitate studies of cancer, inherited disease, and genomic evolution. Denaturing High-Performance Liquid Chromatography (DHPLC) is a recently developed methodology for detection of heteroduplexes formed in DNA samples containing mismatches between wild type and mutant strands. In an effort to develop a rapid, sensitive mutation detection method for studies of families with inherited kidney cancer, we evaluated DHPLC for detection and analysis of MET proto-oncogene mutations in papillary renal carcinomas (PRC). We found DHPLC to be 100% accurate in detecting 15 known disease-associated MET mutations. Significantly, each MET mutation and two novel SNPs generated a characteristic chromatographic profile or signature with reproducible distinguishing features. Standardization of DHPLC reagents and improved methods design were critical to the reliability and accuracy of mutation prediction. Improvements included addition of a 75% acetonitrile wash followed by a rejuvenating gradient, and detailed analysis of signature shape, retention time (RT), RT differences (DeltaRT), and temperature-dependent (melt) profiling. We used signatures to predict mutations in new PRC samples, mutation carriers in asymptomatic hereditary PRC family members, and in a blind study of previously characterized DNAs. Application to SNP discovery is discussed. Wiley-Liss, Inc. PMID- 10874309 TI - Rapid and comprehensive determination of cytochrome P450 CYP2D6 poor metabolizer genotypes by multiplex polymerase chain reaction. AB - The liver enzyme cytochrome P450 CYP2D6 (debrisoquine 4-hydroxylase) metabolizes numerous drugs, including many antidepressants, neuroleptics, antiarrhythmics, and antihypertensive agents. Variability in the gene that encodes this enzyme is an important factor underlying variable drug treatment responses. Some 5-10% of Caucasians lack functional CYP2D6, and the genetic basis of most of these "poor metabolizer" alleles is now well defined. As the CYP2D6 status of a patient can have profound effects on response to drug treatment, it is important to devise methods that permit rapid and economical determination of CYP2D6 genotype. We have developed a robust polymerase chain reaction method that simultaneously identifies the variants CYP2D6 *3, *4, *6, *8, *11, *12, *14, *15, *19, and *20. This constitutes most of the poor metabolizer alleles described in Caucasian and Asian populations. Separate PCR reactions or Southern blots are required for *7, the *5 deletion, and the hybrid alleles *13 and *16. The multiplex assay was validated on 100 individuals previously genotyped by specific polymerase chain reaction-restriction fragment length polymorphism analysis, and proved 100% accurate in this sample. The assay performed consistently with Taq DNA polymerases from various suppliers, within a broad range of temperatures and MgCl(2) concentrations, and using genomic DNA prepared by a range of methods including extraction from dried blood spots on card. This multiplexed, amplification refractory mutation system (ARMS) method is reliable, rapid, relatively cheap, amenable to automation, and offers the advantages of minimal sample handling with no requirement for restriction enzymes as in earlier CYP2D6 assays. PMID- 10874310 TI - Identification of P gene mutations in individuals with oculocutaneous albinism in sub-saharan africa; robyn kerr, gwynneth stevens, prashiela manga, sarah salm, premila john, tabitha haw, and michele ramsay; (Article was originally published in human mutation 15:166-172, 2000) AB - The authors wish to correct a mistake which occurred in the reporting of one of the mutations. The mutation reported as 683insT is actually an insertion G mutation, and should thus be called 683insG. PMID- 10874311 TI - Tuberous sclerosis type 1: three novel mutations detected in exon 15 by a combination of HDA and TGGE. AB - Tuberous sclerosis (TSC) is an autosomal dominant disorder which is genetically heterogeneous with two genes, TSC1 and TSC2. TSC1 consists of 23 exons with exon 15 being the largest one comprising 559 bp. Representing 16% of the coding region exon 15 harbors 37% of the already identified point mutations in TSC1. Mutation screening of large DNA fragments as TSC1 exon 15 by SSCP has been a problem because of the low sensitivity of this method without subdivision. Therefore, we simultaneously performed heteroduplex analysis (HDA) and temperature gradient gel electrophoresis (TGGE) which are both more suitable for evaluation of fragments of this size. DNA samples of 159 patients with the clinical diagnosis of TSC were screened and a total of seven different mutations in nine unrelated cases were identified, including the three novel mutations 1754delT, 1836delT and R500Q. Comparing the two methods applied, HDA showed a higher sensitivity in detecting frameshift mutations, while TGGE seemed to be more sensitive for the detection of base exchanges. We conclude that the combination of these two methods is appropriate to reach a high degree of sensitivity for the detection of all types of small mutations in large DNA fragments. PMID- 10874312 TI - Mutational analysis of BRCA1 and BRCA2 genes in Chinese ovarian cancer identifies 6 novel germline mutations. AB - Germline mutations in the BRCA1 and BRCA2 genes predispose women to breast and ovarian cancer. An incidence of 5% and 3.3% respectively has been reported of BRCA1 and BRCA2 mutations in women with ovarian cancer unselected for family history. The contribution of BRCA1 and BRCA2 mutations to ovarian cancer in Chinese women is unknown. A total of 60 samples of ovarian cancer diagnosed in Chinese unselected for age or family history were analyzed for BRCA mutations using the protein truncation test. The entire coding exon of BRCA1 of 53 cases and that of exon 11 of BRCA2 of 43 cases were successfully screened. Six germline (11.3%) mutations (633C>T, 1080delT, 1129delA, 2371-2372delTG, 3976-3979delGTGA, and IVS 22+7 A>G) were detected in BRCA1. One germline mutation (3337C>T) (2.1%) was detected in BRCA2. None of these seven cases were associated with strong family history of breast and/or ovarian cancer. Five out of our six BRCA1 mutations and the one BRCA2 mutation identified are novel. Our 11.3% incidence of BRCA1 mutations in ovarian cancer found amongst Chinese with insignificant family history is apparently higher than that previously reported in other populations. It suggests that BRCA1 mutation may play a significant role in the development of sporadic ovarian cancer in Chinese women. PMID- 10874313 TI - Identification of mutations in the glucose-6-phosphatase gene in Czech and Slovak patients with glycogen storage disease type ia, including novel mutations K76N, V166A and 540del5. AB - Mutations in the glucose-6-phosphatase (G6Pase) gene are responsible for glycogen storage disease type Ia (GSD Ia). A study of the molecular basis of GSD Ia was carried out in 12 Czech and Slovak GSD Ia patients from 10 unrelated families. Mutation analysis was performed for the entire coding region of G6Pase gene using DGGE, sequencing and PCR/digestion. With the strategy used, all mutant alleles were identified in this study. Three novel mutations (K76N, V166A and 540del5), six previously described mutations (W77R, R83C, G188R, R295C, Q347X and 158delC) and one known polymorphism (1176T-->C) were detected. The most common mutation identified was R83C, accounting for 8 out of 20 (40%) mutant alleles. The K76N mutation was found in a Gypsy family: two siblings with GSD Ia were homozygous for this mutation. These findings expand our knowledge of mutations responsible for glycogen storage disease type Ia. PMID- 10874314 TI - Identification of PATCHED mutations in medulloblastomas by direct sequencing. AB - Medulloblastoma is the most common malignant embryonic tumors of the central nervous system. The nevoid basal cell carcinoma syndrome (NBCCS), which is caused by mutations of PTCH gene on chromosome 9q22, accounts for about 2% of all medulloblastomas. Previous studies of PTCH in sporadic medulloblastomas using single strand conformational polymorphism (SSCP) detected mutations in about 10% of the tumors. In this study, we directly sequenced the PTCH gene in 20 sporadic medulloblastoma DNA samples. A nonsense mutation (Q694X) and a splice site alteration (2875+1G>A) were identified in two of the samples. The mutations are predicted to result in a truncated PTCH protein and aberrant splicing, respectively. In both cases, only the mutant alleles were identified, indicating that the mutations were associated with loss of the wild-type PTCH allele in the tumor cells. Several novel variants, including 1653T>C, 1672C>T, and 2292C>T, were also found in these tumor samples. One of the two mutations detected in this study had been missed by SSCP, suggesting that the true rate of PTCH mutations in sporadic medulloblastomas may be underestimated by SSCP screening. Nevertheless, the frequency of mutations in this study did not differ from previous reports. PMID- 10874316 TI - Three different premature stop codons lead to skipping of exon 7 in neurofibromatosis type I patients. AB - Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder affecting one in 3,500 individuals. The mutation rate in the NF1 gene is one of the highest known for human genes. Compared to other methods, the protein truncation test (PTT) provides improved efficiency in detecting NF1 mutations which are dispersed throughout the gene which spans 350 kilobases of genomic DNA. We have applied the PTT and subsequent sequence analysis of cloned cDNA to identify mutations in NF1 patients. We report here the identification of two novel (W336X and Q315X), and one recurrent (R304X) mutation located in exon 7 and show that all three premature termination codons lead to skipping of exon 7 in a proportion of the transcripts derived from the mutated allele. Possible mutation induced alterations of the RNA secondary structure and their impact on skipping of exon 7 of the NF1 gene are explored and discussed. PMID- 10874315 TI - Mutational analysis of the lysyl hydroxylase 1 gene (PLOD) in six unrelated patients with Ehlers-Danlos syndrome type VI: prenatal exclusion of this disorder in one family. AB - Screening of full length cDNAs for lysyl hydroxylase 1 (LH1; also PLOD) amplified from dermal fibroblasts from six unrelated patients with the autosomal recessive disorder Ehlers-Danlos syndrome type VI (EDS VI) has shown them to be both homozygous and compound heterozygous for mutations in the gene. These mutations, which were verified in genomic DNA, result in a deficiency of LH activity (<25% of normal) in the probands, who are clinically characterized by kyphoscoliosis and extensibility of skin and joints. Four novel mutations identified in these patients include a mutation of an inserted C in one homozygous patient (1702insC) and three point mutations resulting in premature termination codons (PTCs): Y142X, Q327X (in two patients), and R670X. In the family with the R670X mutation we have prenatally excluded EDS VI by the characterization of mutations and their allelic inheritance. We have identified two previously reported mutations in the new patients: a seven exon duplication (in two patients) and a point mutation that codes for a PTC, Y511X, (in two patients). Genotype analysis indicated that the Y511X mutation may originate from a common ancestral gene. Several alternative splicing pathways have been identified which bypass the PTCs and can also restore the open reading frame. PMID- 10874317 TI - Distribution of Q188R and N314D mutations in the Hungarian galactosemic population. AB - The most common causes of galactosemia are mutations of the gene coding galactose 1-phosphate uridyltransferase. Since genotype may correlate with the outcome of the disease, and probably not all of the naturally occurring disease associated mutations are described, characterization of the genotypes in different galactosemic populations are in progress. So far the most extensively examined mutations are the Q188R and the N314D. The first one is associated with the classical galactosemia producing a severe clinical picture with an early onset in homozygous patients. The N314D mutation is associated with the Duarte phenotype, which means a milder form of the disease with a later onset of the symptoms. We studied these mutations in the whole Hungarian galactosemic population by PCR and restriction analysis. We have found the frequency of the Q188R mutation at 33.3%, and the N314D mutation at 11.1%. These results differ from other published data in any other populations. Since the incidence of the disease is the same in Hungary as in other European countries, our study suggests that it is worth to investigate for other mutations in the Hungarian population, of which frequency should be consequently higher. PMID- 10874318 TI - Novel germline mutation (300-305delAGTTGA) in the human MSH2 gene in hereditary non-polyposis colorectal cancer (HNPCC). AB - Hereditary non-polyposis colorectal cancer (HNPCC) is a common hereditary syndrome characterized by the high incidence and early onset of colorectal cancer. The majority of the HNPCC families carry germline mutations in either the MSH2 or the MLH1 mismatch repair gene. A 46 year-old female patient whose family history fulfilled the Amsterdam criteria for HNPCC was diagnosed with undifferentiated adenocarcinoma of the transverse colon. Recognizing the Lynch 2 syndrome (the existance of multiple HNPCC related cancers in a pedigree), we used polymerase chain reaction followed by direct sequencing to screen the coding regions of both the MSH2 and the MLH1 genes for germline mutations in DNA from the patient. We detected a novel germline mutation (300-305delAGTTGA) in exon 2 of human MSH2. We noted microsatellite instability in four microsatellite loci. Immunohistochemistry showed a lack of expression of the MSH2 gene product in the tumor, suggesting that the mutation is a disease-causing mutation. PMID- 10874319 TI - Identification of a larger than 3 Mb deletion including JAG1 in an Alagille syndrome patient with a translocation t(3;20)(q13.3;p12.2). AB - Alagille syndrome (AGS) is an autosomal dominant, developmental disorder affecting multiple organ systems including liver, heart, vertebrae, eye and face. Recurrent deletions of the 20p12 region led to the localization, and ultimately to the identification of mutations in the Jagged1 gene (JAG1) in AGS patients. A translocation t(3;20)(q13.3;p12.2) in an AGS patient was characterized using fluorescent in situ hybridization (FISH). The involvement of 3q and 20p in this translocation was demonstrated using probes for 3q and 20p. Three overlapping YAC clones, 940D11, 953A2, and 675G11 extending to nearly 4 Mb including the JAG1 were used as probes for FISH analysis to define the translocation breakpoint. The translocated chromosome was found to have a deletion of more than 3 Mb including the entire JAG1 gene. The observation of an accompanying large deletion, revealed by molecular characterization of the t(3;20) translocation, is similar to the only other translocation reported in an AGS patient; a t(2;20) translocation was also found to have a large deletion of the JAG1 region at 20p12. PMID- 10874320 TI - Enzymatic mutation detection (EMD) of novel mutations (R565X and R1523X) in the FBN1 gene of patients with Marfan syndrome using T4 endonuclease VII. AB - The Enzymatic Mutation Detection (EMDtrade mark) method is a streamlined and improved version of the original Enzymatic Cleavage of Mismatch (EMC) method. EMD is a fully homogeneous, rapid four step procedure that allows for detection and localization of mismatched or unmatched nucleotides within heteroduplex DNA. To test the utility of EMD for use in the screening of large and complex genes, the fibrillin 1 (FBN1) gene was scanned in a cohort of six patients diagnosed with connective tissue disorders. Four of the six patients were diagnosed with classic Marfan syndrome (MFS). The results were compared with a previous MDEtrade mark scanning of the same patient cohort. Two causative mutations, R565X and R1523X, were detected by EMD that were not detected by MDE. In both cases, the mutation resulted in premature termination of translation. In addition, several polymorphisms were detected by the enzymatic approach that failed detection by heteroduplex analysis. We propose that the EMD method is a sensitive and rapid approach to mutation detection in large genes such as FBN1. PMID- 10874321 TI - A CRX mutation in a Finnish family with dominant cone-rod retinal dystrophy. PMID- 10874322 TI - A novel missense mutation (P191L) in the glucose-6-phosphate translocase gene identified in a Chinese family with glycogen storage disease 1b. PMID- 10874323 TI - Identification of a novel truncating mutation (S171X) in the Emerin gene in five members of a Caucasian American family with Emery-Dreifuss muscular dystrophy. PMID- 10874325 TI - A novel missense mutation (L198R) in the Friedreich's ataxia gene. PMID- 10874324 TI - A novel presenilin 1 missense mutation (L153V) segregating with early-onset autosomal dominant Alzheimer's disease. PMID- 10874326 TI - Identification of a novel mutation, 1087delT, in exon 7 of the CFTR gene in a patient with cystic fibrosis. PMID- 10874328 TI - Identification of two new polymorphisms (c2447-125A>G; c2532G>A) in the gamma 2 COP (COPG2) gene by screening of Silver-Russell syndrome patients. PMID- 10874327 TI - Three novel mutations (P215L, T289P, and 3811-2 A-->G) in the rhodopsin gene in autosomal dominant retinitis pigmentosa in Spanish families. PMID- 10874329 TI - Polymorphism (1339G>A; A447T) in exon 13 of human kidney chloride channel gene CLCNKA. PMID- 10874330 TI - A novel mutation (1320InsT) identified in two Argentine families with variegate porphyria. PMID- 10874331 TI - Triangulation research among culturally diverse populations. AB - There exists an ongoing challenge in the health sciences to develop research methods that effectively describe patterns of health beliefs and actions in different cultures. While the dominant framework for research has traditionally been the quantitative paradigm, qualitative methods place more emphasis on holistic descriptions of the human phenomena and thus may be more appropriate for transcultural research. Triangulation offers an alternative for investigators studying transcultural health by integrating the inherent strengths of both quantitative and qualitative data while minimizing their limitations. This article discusses six approaches for employing triangulation research in transcultural health. PMID- 10874332 TI - Barriers to the use of health services by Chinese Americans. AB - This study examined the extent to which cultural, socioeconomic, and systemic factors impeded access to and utilization of health services among a convenience sample of 52 Chinese immigrants living in metropolitan Houston. The subjects, of differing levels of socioeconomic status, were 25 years old or older. Methods used for data collection included participant observation, face-to-face interview, and case study. A semistructured interview instrument with open- and closed-ended questions was administered. A pilot study and expert reviews were conducted for content and face validity. Cultural and socioeconomic factors were found to be strongly associated with access to and utilization of health services. Mainland Chinese and Taiwanese shared similar cultural dilemmas as they sought health care, including communication difficulties, beliefs about health, health care, and illness, and mistrust in Western health care. Although families played important roles in health decisions and choices of services, social class differences also appeared to affect utilization. For example, more affluent Taiwanese than Mainland Chinese were apt to carry health insurance and use Western systems. The findings suggest a need to improve services to the Chinese community through family-centered and community-based approaches adapted to Chinese culture. PMID- 10874333 TI - Definitions of allied health services in urban community contexts: consumer perspectives. AB - This study, as part of a larger project designed to increase allied health services to underserved urban community agencies, focussed on understanding how consumers defined the services provided in community agencies. Consumers were asked to describe what they defined as service and what constituted good service provision. Qualitative methods were used to conduct key-informant interviews and focus groups with consumers and staff at six community agencies representing different underserved populations, including ethnic and racial minority groups, the homeless, and individuals with disabilities living in the community. Four major themes emerged across the data: 1) the struggle to maintain a stable life with a chronic illness or disability; 2) services need to help solve life challenges; 3) the challenge of accessing and maintaining services; and 4) the need for a peer community. Data analysis revealed that services are difficult to access within and outside community agencies, services need to address short-term as well as long-term issues, and services in the community are still driven by a professional or expert model despite the availability of peer models. Recommendations for health professionals working in community settings are provided. PMID- 10874334 TI - The multicultural sensitivity of physician assistant students. AB - Using a specially designed instrument, the authors examined physician assistant students' multicultural sensitivity at four points before, during, and after the 30 months of a master's degree program. The students (n = 19) were found to have become more multiculturally sensitive by the end of the program, even in the absence of specific relevant instruction. The greatest improvement followed the end of clerkship rotations, where the students had experiences with low-income patients of other racial/ethnic backgrounds. The authors suggest that increasing such experiences during training may enhance students' multicultural sensitivity. PMID- 10874335 TI - Job-loss insecurity versus job-feature insecurity among medical technologists. AB - Using a sample of 292 medical technologists surveyed over a three-year period, this study tested the hypothesis that job-loss insecurity and job-feature insecurity were related but distinguishable constructs. Factor analysis supported this hypothesis. In addition, each construct demonstrated a differential pattern to significant antecedents. Specifically, cross-training desire and organizational downsizing were positively related, while job satisfaction was negatively related, to job-loss insecurity. However, team participation and number of professional organization memberships were negatively related, while routine task was positively related, to job-feature insecurity. Study limitations and future research issues are discussed. PMID- 10874336 TI - Caring about learning while learning about caring: coping with change in health professional education. AB - In recent years educators of health care professionals in the United Kingdom have been faced with problems of organizational upheaval consequent on incorporation into higher education. The author considers some of the problems associated with the individual and organizational strategies adopted to confront the demands of such change. A particular focus is placed upon the concept of a caring learning organization and the organizational dilemmas that must be addressed in contemporary health professional education. PMID- 10874337 TI - Allied health outcomes research using a collaborative distance approach. AB - As part of an activity sponsored by the Coalition for Allied Health Leadership, individuals representing various allied health disciples formed a team to develop a presentation and bibliography on outcomes research in allied health. Team members worked with each other while geographically separated, and concluded that several factors contributed to the difficulty the team encountered in working together in a truly collaborative manner. However, the team's efforts did result in a viable presentation on outcomes research in allied health to include definition and parameters, a discussion of the multidimensional nature of treatment and its effects, suggested schema, and comparisons of several outcome assessment instruments. In addition, a bibliography on outcomes research in allied health was produced. PMID- 10874338 TI - Representation: a call to action for allied health professionals. AB - The Coalition of Allied Health Leadership (CAHL) Representation Project committee examined the representation of allied health professionals in political and other policy-making groups and found it both fragmented and lacking. The benefits to individuals participating in such groups, as well as to the allied health profession as a whole and to the groups themselves, are described. Individuals are urged to participate, and the means to do so are presented. PMID- 10874339 TI - Clinical education in two-year colleges: cost-benefit issues. AB - Allied health program directors and administrators need to be aware of the costs and benefits of their clinical training programs to assure continued availability of training facilities for students. In a pilot study, program directors and administrators who are members of the National Network of Health Career Programs in Two-year Colleges (NN2) were surveyed concerning items to include in a cost benefit-analysis tool, intangible and tangible costs and benefits of clinical education programs, and evaluation of a tool to analyze costs and benefits. Surveys were sent to 138 NN2 members, with 58 responding. Clinical sites were primarily in independent hospitals or health care systems. Most programs had preceptor-to-student ratios of 1:1-1:2, with few students being paid for clinical work. The respondents identified costs as staff time, materials and supplies, equipment, and others. Benefits were orientation and recruitment savings; increased professionalism, job satisfaction, and work quality of staff; ability to maintain and upgrade staff skills and knowledge; and student assistance with clinical coverage. Few programs were required to perform cost analysis. Allied health clinical education programs continue to depend on the willingness of health care facilities to accept students for clinical training. PMID- 10874340 TI - The middle school mentoring program in allied health: a proposed model. AB - This article describes the efforts of one group of participants in the Coalition for Allied Health Leadership program. Their purpose was to design a mentoring program for middle school students that will increase their awareness of the allied health professions as viable career options. A rationale for such a program and a program structure are provided as a beginning point for increasing awareness of allied health careers. PMID- 10874341 TI - Commemorating a classic, and its creator Walter Bradford Cannon. PMID- 10874342 TI - Reactive oxygen species and oxidative DNA damage. AB - Reactive oxygen species (ROS) such as the superoxide anion radical (O2.-) hydrogen peroxide (H2O2) and hydroxyl radical (.OH) have been implicated in the pathophysiology of various states, including ischemia reperfusion injury, haemorrhagic shock, atherosclerosis, heart failure, acute hypertension and cancer. The free radicals, nitric oxide (NO) and O2.- react to form peroxynitrite (ONOO-), a potent cytotoxic oxidant. A potential mechanism of oxidative damage is the nitration of tyrosine residues of protein, peroxidation of lipids, degradation of DNA and oligonucleosomal fragments. Several mechanisms are responsible for the protection of the cells from potential cytotoxic damage caused by free radicals. Cells have developed various enzymatic and nonenzymatic defense systems to control excited oxygen species, however, a certain fraction escapes the cellular defense and may cause permanent or transient damage to nucleic acids within the cells, leading to such events as DNA strand breakage and disruption of Ca2+ metabolism. There is currently great interest in the possible role of ROS in causing DNA damage that leads to cancer and spontaneous mutations. A high rate of oxidative damage to mammalian DNA has been demonstrated by measuring oxidized DNA bases excreted in urine after DNA repair. The rate of oxidative DNA damage is directly related to the metabolic rate and inversely related to life span of the organism. PMID- 10874343 TI - Integrating biostatistics and experimental physiology: students' perceptions and future scope. AB - The use of biostatistics in experimental physiology is recognised by researchers. However, there has been no concerted attempt to integrate biostatistics into the undergraduate experimental physiology programme. This paper describes one such initiative. The student's response to the exercise was largely positive; it enabled them to describe and interpret data more effectively and understand the experiments more completely. The attitudes of the students to the exercise and their performance at a statistics examination at the end of the exercise was determined by a number of parameters, including prior statistical knowledge, general academic performance and the extent to which they liked mathematics. However, even those students who disliked mathematics indicated that they appreciated the value of the exercise. The results indicate the need to integrate biostatistics into the undergraduate physiology course. PMID- 10874344 TI - Subicular lesion induced impairment in operant behaviour and altered dendritic morphology of CA1, CA3 hippocampal neurons. AB - The effects of bilateral electrolytic subicular lesions were examined on the operant behaviour for food reward on a continuous reinforcement schedule as well as the dendritic morphology of CA1 and CA3 hippocampal areas. The subjects were female Wistar rats 20 days of age and were divided into four groups. 1. Age matched control 2. Sham operated 3. Operant behaviour for food reward and 4. Subicular lesion. Animals were starved twenty-four hours prior to operant behaviour training sessions. Two trial sessions with continuous reinforcement (CRF) of 10 min duration/day were done during training sessions following which the rats were allowed CRF for ten minutes per day for ten days. On the eleventh day, the operant behaviour and sham operated animals were taken up for bilateral subicular lesion and sham surgery respectively. After seventy two hours of surgical recovery, operant behavioural testing was done as before for a further period of ten days. Later all the groups of animals were sacrificed and the hippocampi were processed for rapid Golgi staining technique. Our results suggest that subicular lesions do produce a significant reduction in operant learning behaviour for food reward. Further the Golgi studies revealed a reduction in dendritic branching points and intersections of apical and basal CA1, CA3 neurons in lesioned animals. PMID- 10874345 TI - Effect of two selected yogic breathing techniques of heart rate variability. AB - The heart rate variability (HRV) is an indicator of the cardiac autonomic control. Two spectral components are usually recorded, viz. high frequency (0.15 0.50 Hz), which is due to vagal efferent activity and a low frequency component (0.05-0.15 Hz), due to sympathetic activity. The present study was conducted to study the HRV in two yoga practices which have been previously reported to have opposite effects, viz, sympathetic stimulation (kapalabhati, breathing at high frequency, i.e., 2.0 Hz) and reduced sympathetic activity (nadisuddhi, alternate nostril breathing). Twelve male volunteers (age range, 21 to 33 years) were assessed before and after each practice on separate days. The electrocardiogram (lead I) was digitized on-line and off-line analysis was done. The results showed a significant increase in low frequency (LF) power and LF/HF ratio while high frequency (HF) power was significantly lower following kapalabhati. There were no significant changes following nadisuddhi. The results suggest that kapalabhati modifies the autonomic status by increasing sympathetic activity with reduced vagal activity. The study also suggests that HRV is a more useful psychophysiological measure than heart rate alone. PMID- 10874346 TI - Recovery from stress in two different postures and in Shavasana--a yogic relaxation posture. AB - The recovery from induced physiological stress in Shavasana (a yogic relaxation posture) and two other postures (resting in chair and resting supine posture) was compared. Twenty one males and 6 females (age 21-30 yrs) were allowed to take rest in one of the above postures immediately after completing the scheduled treadmill running. The recovery was assessed in terms of Heart Rate (HR) and Blood pressure (BP). HR and BP were measured before and every two minutes after the treadmill running till they returned to the initial level. The results revealed that the effects of stress was reversed in significantly (P < 0.01) shorter time in Shavasana, compared to the resting posture in chair and a supine posture. PMID- 10874347 TI - A comparison of the blood lipid profiles of professional sportspersons and controls. AB - Total cholesterol (TC), triglyceride (TG), high density lipoprotein-cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C) and HDL-C/TC levels are important in determining the risk of coronary heart disease. The serum lipids and lipoprotein levels of regularly training sportspersons and non-sporting controls were determined and compared with each other to investigate the effects of exercise and sex on these factors. HDL-C levels of male and female training groups were higher than those of corresponding non-sporting groups (respectively P < 0.01, P < 0.001). The sportswomen's HDL-C levels were higher (P < 0.05); and TC, TG, and LDL-C levels were lower. (P < 0.001) than those of sportsmen's levels. The non-sporting women's TC and TG levels were lower than those of non sporting men's levels (P < 0.001). HDL-C/TC ratio of active females was higher than that of control females (P < 0.01). The corresponding difference in males was also significant. We conclude that physical activity and sex have effects on risk factors for cardiovascular disease. PMID- 10874348 TI - Biological efficacy and plasma norethisterone levels of orally administered norethisterone enanthate in rat and hamster. AB - Norethisterone enanthate (NET-En) is a well known intramuscular contraceptive drug. The long acting nature of this preparation when administered orally was evaluated in female rats and hamsters using fertility inhibition test and from the plasma levels of norethisterone (NET). An oral dose of 20-60 mg NET-En was administered to random groups of six female rats and hamsters and were mated after five and ten days with males of proven fertility. The fertility inhibition rate was determined from vaginal delivery. A dose-dependent reduction in fertility was seen in rats 5 days after oral administration of NET-En. This effect was found to be less pronounced and not significant 10 days after administration of similar doses of NET-En. In hamsters, a similar but less pronounced effect was noted. The decrease in fertility was significant only at the 60 mg dose. The plasma levels of NET estimated by RIA over a period of 15 days, in a different set of treated rats, suggested rapid absorption of NET-En within a day, and drug concentration decreased slowly, the levels on the 4th day ranged from 0.9-2.3 with the 10 mg and 1.0-4.0 ng/ml with the 20 mg dose. Detection of adequate levels of NET in plasma during the estrous cycle in rats, and the fertility inhibition observed in female rats and at higher doses in hamsters, suggest that NET-En is orally active. PMID- 10874350 TI - Effects of the menstrual cycle on timing and depth of breathing at rest. AB - Volume and timing components of resting ventilation were measured serially in 40 women aged 18 to 36 yr, during menstrual, follicular and luteal phases of menstrual cycle. Resting minute ventilation (VE) was significantly higher (P < 0.001) in luteal phase than in menstrual and follicular phases; in the two latter phases VE was almost equal. This increment in VE during the luteal phase was due to a significant rise (P < 0.001) in tidal volume (VT). Respiratory frequency (f) was unchanged throughout the cycle. Although there was a mean increases in inspiratory time (T1) during the luteal phase compared to the other two phases, the difference did not reach statistical significance. Duty cycle, T1/Ttot, was also unchanged throughout menstrual cycle. However, mean inspiratory flow, VT/T1, was significantly higher (P < 0.05 and P < 0.01) during luteal phase as compared to that during menstrual or follicular phases respectively. Pulmonary mechanics, as measured by forced vital capacity (FVC), forced expiratory volume in one second (FEV1) and forced mid expiratory flow rate (FEF25%, 75%), were within normal limits and remained unaltered during the menstrual cycle. Therefore, in the absence of alteration of pulmonary mechanics, the luteal increase in ventilation and inspiratory flow suggests a possible role for progesterone in stimulating the respiratory drive, either centrally or through the peripheral chemoreceptors or by both. PMID- 10874349 TI - Influence of adenosine agonists and antiepileptic drugs on theophylline-induced seizures in rats. AB - Seizures is a major toxicity of theophylline. The mechanism of theophylline induced seizures is not known, but antagonism at adenosine receptors may be a possibility. The effect of pretreatment with different doses of adenosine (100, 500 and 1000 mg/kg, i.p.), and the adenosine A1 receptor agonist N6 cyclopentyladenosine (CPA), 1, 5 and 10 mg/kg, i.p., was studied against seizures induced by theophylline in rats. Both these drugs, at all dose levels tested, failed to protect theophylline seizures. Thus adenosinergic system is unlikely to be involved in mediating the convulsant action of theophylline. On the other hand, the conventional antiepileptic drugs, i.e. diazepam (4 mg/kg), sodium valproate (300 mg/kg) and phenobarbitone (50 mg/kg), but not carbamazepine, afforded some protection. The modification of course of seizures, by the antiepileptic drugs suggests the involvement of some other alternate mechanism in theophylline-induced seizures. PMID- 10874351 TI - Nicotine induced ovarian and uterine changes in albino mice. AB - Nicotine at the dose level of 0.3 mg/100 g body weight was administered to normal cycling mice for 15 days through oral and intraperitoneal routes. At autopsy on 16th day significant reduction in the ovarian and uterine weight was observed. Histological observations showed decrease in the number and size of Graafian follicles, corpora lutea and increase in the atretic follicles in the ovary. The uterus showed absence of endometrial glands, decrease in the height of myometrium, endometrium and its epithelial cells. The total cholesterol content of the ovary and uterus is increased whereas the protein content is decreased. This antagonistic action of nicotine to gonadotrophins is discussed. PMID- 10874352 TI - Efficacy of exogenous gonadotropins on the maintenance of spermatogenesis in pethidine treated albino rats. AB - An attempt is made to induce the pethidine suppressed gonadal activities by the administration of exogenous gonadotropins (hCG, PMSG, hCG + PMSG). Administration of 5 IU gonadotropins either separately or in combination to the rats treated with pethidine for 30 days resulted in the significant increase in the weight of testis, diameter of testis and seminiferous tubules. Gonadotropin(s) treatment stimulated the spermatogenic activity which was inhibited by pethidine. Therefore the number of spermatogonia, spermatocytes, spermatids in the seminiferous tubules and spermatozoa in cauda epididymis is increased significantly. Decreased testicular cholesterol, increased protein content and weight of accessory sex organs indicate the rejuvenation of steroidogenesis. Combination of both the gonadotropins is more effective in bringing all these activities. PMID- 10874354 TI - Peak expiratory flow rate in flour mill workers. AB - The current cross-sectional study with a comparison group was undertaken to investigate peak expiratory flow rate (PEFR) in flour mill workers and to study relationship between reduction in PEFR and age, smoking, exposure to grain dust and respiratory morbidity. The study included 286 flour mill workers and equal number of neighbourhood controls group-matched for age. PEFR was measured by using Wright's Peak Flow Meter. PEFR was significantly reduced in flour mill workers as compared to comparison group. The decline in PEFR was significantly associated with grain dust exposure, duration of exposure, tobacco smoking and presence of respiratory morbidity. PMID- 10874353 TI - Electrocardiographic changes following exercise in the congenitally deaf school children: relationship with Jervell Lange Neilsen syndrome (the Long QT syndrome). AB - The present study was conducted to test the effects of exercise stress on the ECG of the congenitally deaf children from school for deaf, in view of the occurrence of the Jervell-Lange Neilsen (Surdo Cardiac) variant of the Long QT Syndrome (LQTS) in them. An ECG Lead II was recorded at rest and after two minutes of static jogging. For comparison, the same protocol was repeated in normal healthy children from another school. ECG were analysed for the calculation of corrected QT interval (QTc) by Bazett's equation QTc = QT/square root of R-R and also for the evidence for other abnormalities. Both in the normal and deaf children, exercise did not produce significant (P > 0.05) change in QTc from their resting values. However, when pre and post exercise QTc values of deaf children were compared with normal children, the female deaf had significantly longer QTc (P < 0.01) both at rest and after exercise than normal female children. Normal children did not show significant ECG abnormality either at rest or on exercise. On the contrary many of their counter part (deaf) exhibited occasional ECG abnormality at rest but plethora of abnormalities after exercise viz., sinus arrhythmias, sinus pauses, ST depression, T-inversion, biphasic-T, notched-T, T alternans, nodal ectopics and junctional rhythm. These results lend credence to the hypothesis of sympathetic imbalance and repolarisation defects in deaf children's heart, which in more severe form could pass into frank Jervell-Lange Neilsen variant of the Long: QT Syndrome. PMID- 10874355 TI - Effect of Septilin--a herbal preparation on pharmacokinetics of carbamazepine in rabbits. AB - The study was carried out in rabbits, to see the effects of Septilin, a herbal preparation on the single and multiple dose kinetics of carbamazepine. Single dose treatment of Septilin significantly decreased t 1/2a. t 1/2e. AUCo-alpha of carbamazepine. Steady state Cmax and AUC0-24 of carbamazepine were also reduced significantly in comparison to those of control rabbits after 7 days co administration of Septilin. We conclude that Septilin decrease/hinder the absorption process of carbamazepine through an unknown mechanism. PMID- 10874356 TI - Antimicrobial prescribing pattern in an Indian tertiary hospital. AB - 550 prescriptions of the indoor patients receiving antimicrobial drugs in the Departments of Internal Medicine, Surgery, Urology and Paediatrics were analysed for drug utilization studies. The prescribing frequency of one antimicrobial per prescription was maximum in Surgery and Urology (52.52%) and Internal Medicine (50.51%) whereas prescribing frequency of two antimicrobials was maximum in Paediatrics (59.9%). In all the departments, quinolones, aminoglycosides, cephalosporins and penicillins were frequently prescribed among which amikacin, ciprofloxacin, cefotaxime and cloxacillin were most preferred drugs, with a general tendency of prescribing newer antimicrobials. In majority of cases selection of antimicrobials was not based on microbiological confirmation. It is suggested that the use of newer and expensive antimicrobials should be kept reserved only for serious and life threatening situations. PMID- 10874357 TI - A study of pulmonary profile of hypertensive patients--comparison of atenolol vs amlodipine. AB - Two groups of drugs commonly used for the treatment of hypertension are atenolol and amlodipine. These drugs are reported to have conflicting changes on pulmonary responses. In order to study the effect of hypertension and antihypertensive treatment on pulmonary responses, 40 patients with essential hypertension having diastolic blood pressure between 90-114 mmHg on three consecutive weekly visits were taken. Pulmonary responses were tested at the end of 2 weeks of placebo washout period and then at the end of 6 weeks of treatment with either atenolol or amodipine. Using a computerized autospiror along with the weekly recordings of heart rate and blood pressure, the various pulmonary and cardiac parameters were taken. Analysis of the result showed that atenolol treatment resulted in significant decline of forced vital capacity (FVC), % forced vital capacity (%FVC), and forced expiratory volume in first second (FEV1) whereas amlodipine did not show any significant change on pulmonary parameters. PMID- 10874358 TI - The influence of age, sex and obesity on blood pressure levels in a tribal population. PMID- 10874359 TI - Surface changes of the erythrocytes membrane in alcoholics--a study of lectin binding. AB - Erythrocytes from 30 alcoholic patients as well as age and sex matched healthy control were examined, for hemagglutination titre, against, a purified galactose specific lectin from the latex of Pedilanthus tithymalodies. Mean hemagglutinin titre was significantly elevated (P < 0.001) in alcoholics as compared to controls. The results indicate that alcohol abuse is associated with adoptive cell surface changes on erythrocytes, measurable by its lectin binding capacity. PMID- 10874360 TI - Involvement of nitric oxide (NO) in hypoglycaemic activity of tolbutamide. AB - The study was conducted to find the involvement of Nitric Oxide (NO) using L arginine, a NO precursor and NG-methyl L-arginine a nitric oxide synthase inhibitor on tolbutamide activity in normal rabbits. L-arginine (25-300 mg/kg, body weight, oral) produced transient and dose dependent hypoglycaemia. When combined with tolbutamide (40 mg/kg, oral) it produced early and prolonged action. The effect of tolbutamide was blocked by NG-methyl L-arginine (5 mg/kg, body weight, oral). The results confirm the involvement of NO in tolbutamide activity and the possibility of using L-arginine as a supplement to antidiabetic drugs in blood glucose control. PMID- 10874361 TI - A randomized double-blind controlled study of nimodipine in acute cerebral ischemic stroke. AB - A randomized placebo controlled double-blind clinical trial of nimodipine was conducted in 31 patients of acute cerebral infarction. Nimodipine was administered in dosage of 120 mg/day for 28 days. Treatment was begun within 48 hours of ischemic stroke. Diagnosis was confirmed by computed tomographic (CT) scan. Similar number of patients (control) received placebo. Neurological assessment was done at the time of entry into the trial, and after 4 weeks, by using Mathew's scale. After four weeks of treatment with nimodipine or placebo, Mathew's scale score improved significantly (< 0.001) in both groups, but difference in mean score between two groups was insignificant (> 0.05). However, significant difference (< 0.05) was noted in relative change in neurological deficit (mean X-value) of two groups. The nimodipine group had higher value in scores on Mathew's scale. No adverse reaction, was observed in either group. The study suggests a beneficial a effect of nimodipine in acute cerebral ischaemia. PMID- 10874363 TI - Animal use in medical research. PMID- 10874362 TI - Effect of Met-enkephalin on blood glucose level. PMID- 10874364 TI - Thyroid functions in aging men. PMID- 10874365 TI - Measurement and interpretation of peak expiratory flow. PMID- 10874366 TI - Cold pressor response in normal and malnourished children. PMID- 10874367 TI - Regression equations for prediction of normal lung function. PMID- 10874368 TI - Association of angiotensin converting enzyme (ACE) gene polymorphism with hypertension in a Bangladeshi population. AB - The association of angiotensin converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism with hypertension has not been confirmed. Inconsistencies may be due to the differences of background population characteristics. Till date, there has been no report in Bangladeshi population. This study was to examine the association of ACE (I/D) polymorphism with hypertension. Fifty-one primary hypertensives and fifty-two normotensives were recruited from a hospital in Dhaka city. Height, weight and blood pressure were measured. ACE (I/D) genotypes was established using polymerase chain reaction protocol. The genotype and allele frequencies did not differ significantly (P > 0.05) between the groups. In logistic regression analysis, adjusted for age, sex and body mass index, the genotypes were not associated with hypertension (DD vs II: Adds ratio = 2.6, P = 0.34; ID vs II: 0.4, 0.23; ID + DD vs II: 0.8, 0.69). In this hospital-based sample of Bangladeshi people, significant association of ACE I/D genotype with hypertension was not observed. PMID- 10874369 TI - Foetal compromise by spontaneous foetal heart rate deceleration in reactive non stress test and decreased amniotic fluid index. AB - One hundred and sixty-three consecutive pregnant women with > 32 weeks gestation, undergoing non stress test (NST) and amniotic fluid index (AFI) determination were divided into six groups according to the amniotic fluid index and the nature of decelerations of foetal heart rate. Foetuses with antepartum decelerations had statistically significantly increased incidence of intrapartum decelerations, caesarean section due to intrapartum foetal distress, cord complications and small for gestational age infants. More so with decreased amniotic fluid index (p < 0.001). Low 5 minute apgar score was also significantly increased (p < 0.05). Prediction of foetal compromise might be done by spontaneous decelerations in reactive non-stress tests and with an amniotic fluid index < or = 5 cm. PMID- 10874370 TI - Monitoring efficacy of commonly used antimalarials by a 14-day in-vivo test in a new settler's camp in endemic zone at Cox's Bazar. AB - The study was done in a new settler's camp "Barachara" under Sadar thana of Cox's Bazar district. It has a total population of 784 of all age groups, registered in the middle of the study period. A prospective evaluation of all cases of fever were done over 12 months, to see the pattern of febrile illness among the population and to compare the therapeutic efficacy of two alternative drug regimens for uncomplicated falciparum malaria (UM). Blood for malarial parasite (MP) was done in all cases of fever and was treated in line with the new clinical case definitions and treatment guidelines for malaria in Bangladesh. Slide positive UM cases were subjected to a "14-day in-vivo test" for therapeutic efficacy testing of antimalarial agents. The two drug regimens were randomised by lottery--a) 3 days oral chloroquine plus single dose sulphadoxin/pyrimethamine (CQ + SP) and, b) 3 days oral quinine plus single dose sulphadoxin/pyrimethamine (Q3 + SP). Drug administration was supervised by the field assistant and was followed up on days 3, 7 and 14 for blood slide examinations and clinical assessment. Sensitive response was observed in 79% of the cases in the CQ + SP group and 84% in the Q3 + SP group. Early treatment failure (persistently febrile and parasitaemic on days 3 or 7) was observed in 16% in the CQ + SP group and 9% in the Q3 + SP group. Both the evaluated drug regimens had less than 20% failures and can be used as alternative first line agents and Q3 + SP regimens can also be used as the second line agents for treatment failure (to chloroquine and/or SP) UM cases in the study area. PMID- 10874371 TI - Effect of health warning on the use of breast milk substitutes by lactating mothers. AB - This cross-sectional study was carried out with the objectives to determine the awareness, readability and understandability of health warning and its effect on the use of breast milk substitutes(BMS) by lactating mothers. A total of 400 mothers who had infant of less than 5 months of age & able to read the bangla newspaper were interviewed at urban EPI centres. The mothers were requested to locate and read the health warning on BMS container. The BMS were used by 189(47.25%) mothers but only 137(34.3%) mothers knew the health warning. The mean time taken by mothers to locate the health warning in the BMS container was 40.4 +/- 40.9 seconds(95% CI 35-45). The health warning in the container could not be located by 97(24.2%) mothers and another 51(12.2%) located it with difficulty. There were 263(65.8%) mothers who were able to read the health warning easily and the rest either read with difficulty (10.0%) or unable to read(24.24%). More than one third of the mothers did not understand the meaning of health warning. Even after the education on health warning during the course of interview, 170(42.5%) mothers responded in favour of using BMS. The low level of awareness and poor understanding of the meaning of health warning by the lactating mothers might be the cause of a high rate of BMS use which calls for strengthening of information, education and communication (IEC) activities. PMID- 10874372 TI - Acquired pure red cell aplasia--a case report. AB - A married female patient of 36 years with chronic anaemia, because of pure erythroid aplasia with a haemolytic component and hypothyroidism due to antithyroid auto-antibodies, was subsequently discovered as a case of systemic lupus erythematosus (SLE). She was treated with corticosteroid and immunosuppressive therapy and her anaemia was corrected. The response of erythroid aplasia to corticosteroid and other immunosuppressive agents suggests that immunological factors play a role in erythroid aplasia in SLE. The occurrence of red cell aplasia in association with a variety of immune phenomenon supports the concept that in SLE, erythroid aplasia may be of immune aetiology. PMID- 10874373 TI - A role for conjoint analysis in technology assessment in health care? AB - The aim of this paper is to demonstrate the use of conjoint analysis (CA) in health services research. Conjoint analysis is first explained, with emphasis on the history of the technique, followed by an explanation of how to carry out such a study and how the results from such a study can be used. The technique is demonstrated with reference to a study that looks at the benefits of in vitro fertilization. It is shown how CA can be used to estimate the relative importance of attributes, the trade-offs individuals make between these attributes, willingness to pay if cost is included as an attribute, and utility or benefit scores for different ways of providing a service. The paper then considers the potential advantages of CA over other, more commonly used benefit assessment instruments. Finally, there is discussion of the issues raised in the design and analysis of CA studies. It is concluded that these issues must be addressed before the technique becomes an established instrument for technology assessment. PMID- 10874374 TI - Combining feedback from simulated cases and prescribing. Design and implementation of an educational intervention in primary care in Sweden. AB - OBJECTIVES: To develop and evaluate a new model of continuing medical education (CME) for general practitioners (GPs). The study is part of the joint European Drug Education Project (DEP). This paper presents the Swedish part regarding the design of the evaluation study, the educational methodology, and the participants' evaluation. METHODS: An educational model was developed. Two peer group discussions (facilitated by a GP/pharmacist team), including individual feedback on the GPs' judgments of written simulated cases and prescribing, were main components. The model was tested in a parallel randomized controlled study including 36 GP groups, allocated to education on asthma or urinary tract infections. Background and outcome data were knowledge and attitudes (K/A) assessed by a questionnaire and prescribing practices for actual and written simulated cases. The GPs' evaluation of the model was captured through a questionnaire. RESULTS: All 36 groups completed the program. The mean participation rate in the group discussions was 75%. The response rates were 82 98% regarding outcome data K/A questionnaire and written cases), and 80% regarding the evaluation questionnaire. Prescribing data were captured for 99% of the GPs. Both group discussions were considered important by 84-89%. Eighty-seven percent wished to take part in similar CME activities for other conditions. About 80% reported that their purpose in participating had been fulfilled. CONCLUSIONS: It was feasible to evaluate the developed educational model by using a two-armed parallel study design. The model was well received by the participants. PMID- 10874375 TI - Utilization of percutaneous transluminal coronary angioplasty for quality assurance in health care from 1983 to 1996. AB - OBJECTIVES: To examine the distribution of interventional cardiac catheterization laboratories, their case load, the time trends, and the regional variation of percutaneous transluminal cutaneous angioplasty (PTCA) utilization in Italy. METHODS: Analysis of data was provided by the annual reports of the Italian Group of Studies and Interventional Cardiology over the period from 1983 to 1996. RESULTS: The number of PTCA facilities and their use steadily increased, mainly in the North. In 1996 the utilization rate was 34 per 100,000 population, but only 60% of labs performed 200 or more procedures. CONCLUSIONS: Dramatic time trends and regional variations often took place without an epidemiology and technology assessment-based planning process. PMID- 10874376 TI - Economic evaluation of diagnostic tests. A review of published studies. AB - OBJECTIVES: The purpose of this review was to examine whether studies from the medical literature focusing on efficiency of diagnostic facilities reported economic evaluation methods appropriately, following guidelines for conducting and reporting economic evaluations. METHODS: A Medline search was conducted, and studies that concerned a diagnostic technology and fulfilled the Drummond criteria were selected for methodological review. The reliability of selection and methodological review based on the abstracts was determined by scoring a random sample of both abstracts and full articles. Interrater reliability was determined by scoring a random sample of abstracts by both authors. Kappa values were calculated. Nine methodological aspects were reviewed: study design, the type of economic evaluation, the comparison made, the study's perspective, the cost-effectiveness ratio used, the definition of cost-effective, the types of costs analyzed, the cost calculation method, and the use of sensitivity analysis. RESULTS: Two hundred fifty studies published between 1992 and 1997 were found regarding efficiency of diagnostic facilities; 134 studies fulfilled the Drummond criteria and were selected for methodological review. Kappa values showed reliability of selection and methodological review and interrater reliability. The existing literature on the economic evaluation of diagnostic facilities does not adhere well to guidelines for economic evaluation. In 95%, no perspective was mentioned, in 50% of the cases no ratio was given, in 82% the cost calculation method was not mentioned, and in 66% no sensitivity analysis was reported. CONCLUSIONS: Our review suggests that to improve the quality of reporting economic evaluations, editorial boards could issue and enforce guidelines for standard reporting of such studies. PMID- 10874377 TI - Examining the influence of picture archiving communication systems and other factors upon the length of stay for patients with total hip and total knee replacements. AB - OBJECTIVES: To examine the influence of a picture archiving and communication system (PACS) on the length of stay (LOS) for patients receiving total hip replacement (THR) or total knee replacement (TKR) procedures. METHODS: A before and-after design was used. Data were collected on all THR and TKR procedures at Hammersmith Hospital from 1993-96. A regression approach was used to examine the influence of PACS on LOS. Factors such as patient age, sex, and physician were controlled for. RESULTS: Type of admission and discharge, month of procedure, complications, and number of procedures all significantly influenced LOS for patients undergoing THR. For patients receiving TKR, age, sex, admission, prosthetic complications, number of procedures, and PACS significantly influenced LOS. CONCLUSIONS: While this study shows an apparent reduction of 25% in the average LOS for TKR patients at the time PACS was introduced, this is unlikely to be a true PACS effect and no similar reduction in LOS was shown for THR patients. PMID- 10874378 TI - Disease management. A new technology in need of critical assessment. AB - Recently, many disease management programs, especially for patients with chronic diseases, have emerged. This paper discusses the potential benefits and disadvantages of disease management, on the basis of an extensive literature review. Disease management is an innovative technology in health care management, which is diffusing throughout the health care system without critical evaluation. Evidence on its effectiveness and costs is still very scarce, while the legal, ethical, organizational, and social implications of this practice have not been analyzed seriously. Before disease management is implemented on a broader scale in different European settings, first, empirical evidence about its alleged benefits and cost-effectiveness should be collected. PMID- 10874379 TI - Economic evaluation of the cochlear implant. AB - OBJECTIVE: To examine the economic efficiency of current cochlear implant technology under Australian conditions in profoundly deaf adults, partially deafened adults, and children. METHODS: Cost-utility study, with weights based on judgments from persons experienced with the technology, and cost data from Australian sources. RESULTS: Quality-of-life improvements due to functional consequences of hearing improvement were greater than those due to amelioration of hearing disability. Costs in Australian dollars per QALY (15-year assessment) ranged from $5,070-$11,100 for children, $11,790-$38,150 for profoundly deaf adults, and $14,410-$41,000 for partially deaf adults. CONCLUSIONS: Results suggest cochlear implantation is acceptable value for money when compared with other health programs to which resources are committed in Australia. PMID- 10874380 TI - The cost-effectiveness of prophylaxis for Mycobacterium avium complex in AIDS. AB - OBJECTIVE: To develop a simulation model to project costs, life expectancy, and cost-effectiveness in discounted dollars per quality-adjusted life-year (QALY) saved for clinical strategies to prevent Mycobacterium avium complex (MAC) in patients with AIDS. METHODS: We used natural history data from the Multicenter AIDS Cohort Study, efficacy and toxicity data from randomized clinical trials, and cost data from the AIDS Cost and Services Utilization Survey. The model permits timing of prophylaxis to be stratified by CD4 count (201-300, 101-200, 51 100, and < or = 50/mm3), and allows combinations of prophylaxis, crossover to second- and third-line agents for toxicity, and consideration of adherence, resistance, and quality of life. RESULTS: The model projects that the average HIV infected patient with a beginning CD4 count between 201 and 300/mm3 has total lifetime costs of approximately $43,150 and a quality-adjusted life expectancy of 42.35 months. If azithromycin prophylaxis for M. avium complex is begun after the CD4 declines to 50/mm3, costs and quality-adjusted survival increase to approximately $44,040 and 42.78 months, respectively, for an incremental cost effectiveness ratio of $25,000/QALY compared with no M. avium complex prophylaxis. Other prophylaxis options (i.e., rifabutin, clarithromycin, and combination therapies) either cost more but offer shorter survival, or have cost effectiveness ratios above $260,000/QALY. Sensitivity analysis reveals that, for reasonable assumptions about quality of life, risk of infection, prophylaxis cost, adherence, and resistance, azithromycin remains the most cost-effective prophylaxis option. CONCLUSIONS: Azithromycin prophylaxis, begun after the CD4 count has declined to 50/mm3, is the most cost-effective M. avium complex prophylaxis strategy. Consistent with new United States Public Health Service guidelines, it should be the first-line prophylaxis option. PMID- 10874381 TI - An economic evaluation of pain therapy after hysterectomy. Patient-controlled analgesia versus regular intramuscular opioid therapy. AB - OBJECTIVES: To assess the economics of patient-controlled analgesia (PCA) treatment versus regular intramuscular (i.m.) injections of opioid analgesia for pain management after hysterectomy. METHODS: Cost-minimization analysis was used based on the comparable pain control results achieved in the two treatment groups. Observations were taken of treatment-related events with personnel (mostly nursing) time implications during the trial. Times were then associated with these events in an independent study of personnel activity. Costs were linked by using average wage rates for the various personnel for the Montreal area during the time of the study. Drug and material costs were hospital acquisition costs for all items. The cost of the PCA pump itself was not included in the analysis. Several analyses were performed to test the sensitivity of the results to various assumptions. RESULTS: The results for total costs of the two therapies generally showed PCA to be more costly than regular i.m. injections despite no costs of the pump being included in the analyses. These results were robust with respect to changes in assumptions. Even when intentionally biasing the analysis against i.m. therapy, it was difficult to obtain results that favored PCA. CONCLUSIONS: Based upon the institutions and assumptions in this analysis, PCA offers no cost advantages over regular i.m. therapy in the pain management after hysterectomy. Regular i.m. injections provided less costly analgesia. PMID- 10874382 TI - Probabilistic sensitivity analysis in cost-effectiveness. An application from a study of vaccination against pneumococcal bacteremia in the elderly. AB - OBJECTIVES: We explore the policy implications of probabilistic sensitivity analysis in cost-effectiveness analysis by applying simulation methods to a decision model. METHODS: We present the multiway sensitivity analysis results of a study of the cost-effectiveness of vaccination against pneumococcal bacteremia in the elderly. We then execute a probabilistic sensitivity analysis of the cost effectiveness ratio by specifying posterior distributions for the uncertain parameters in our decision analysis model. In order to estimate probability intervals, we rank the numerical values of the simulated incremental cost effectiveness ratios (ICERs) to take into account preferences along the cost effectiveness plane. RESULTS: The 95% probability intervals for the ICER were generally much narrower than the difference between the best case and worst case results from a multiway sensitivity analysis. Although the multiway sensitivity analysis had indicated that, in the worst case, vaccination in the 85 and older age group was not acceptable from a policy standpoint, probabilistic methods indicated that the cost-effectiveness of vaccination was below $50,000 per quality-adjusted life-year in greater than 92% of the simulations and below $100,000 in greater than 95% of the simulations. CONCLUSIONS: Probabilistic methods can supplement multiway sensitivity analyses to provide a more comprehensive picture of the uncertainty associated with cost-effectiveness ratios and thereby inform policy decisions. PMID- 10874383 TI - Costs of health care and social services during the first year after ischemic stroke. AB - OBJECTIVES: Knowledge of resource use and costs can be useful when evaluating existing services or planning new services. This study investigates the use of health care and social services during the first year after a stroke. Total costs are calculated, costs are compared across subgroups of patients, and resource items of major importance for the total costs are identified. METHODS: The study is based on a database comprising data on all stroke patients admitted to a university hospital in Copenhagen, Denmark, over a 1-year period, 1994-95. Patients were followed for 1 year after the stroke, and data on resource use during and after hospitalization were collected prospectively at interviews. This paper focuses on a subset of 385 patients who were admitted because of cerebral infarct or unspecified stroke. RESULTS: The mean cost, based on all patients, of health care and social services during the first year was 142,900 DKK (US $25,500). The hospital care until the first discharge, including acute care and rehabilitation, cost 101,600 Danish krones (DKK) (US $18,100), i.e., 71% of the total cost. Major resource items after discharge were nursing homes, readmissions, outpatient rehabilitation, and home help. The cost during the first year varied with a number of factors, with the most important being survival and degree of disability. Patients who survived the acute phase and who had severe disability (Barthel Activities of Daily Living [ADL] Index: 0-9) 7-10 days after admission had a total cost during the first year that was five times as high as patients with no disability (Barthel ADL Index: 20). CONCLUSION: Costs of health care and social services during the first year after a stroke vary considerably. Disability as measured with the Barthel ADL Index is a stronger predictor of costs than Scandinavian Stroke Scale scores and other clinical and demographic variables. PMID- 10874384 TI - Health technology assessment and clinical decision making: which is the best evidence? AB - This paper examines the rationality of the concepts underlying evidence-based medicine and health technology assessment (HTA), which are part of a new current aimed at promoting the use of the results of scientific studies for decision making in health care. It describes the different approaches and purposes of this worldwide movement, in relation to clinical decision making, through a summarized set of specific HTA case studies from Catalonia, Spain. The examples illustrate how the systematic process of HTA can help in several types of uncertainties related to clinical decision making. PMID- 10874385 TI - Health technology assessment and the regulation of medical devices and procedures in Quebec. Synergy, collusion, or collision? AB - In this paper, we discuss the complex relationship between health technology assessment (HTA) and the regulation of medical devices and procedures. The relationship is first examined through a conceptual framework describing the itinerary from research to three levels of policy making: micro (standards of medical practice), meso (institutional rules), and macro (health policies). Four reports from the Quebec Health Technology Assessment Council (CETS) are used to illustrate how HTA activities can influence the regulatory mechanisms operating at each decision-making level. We then discuss the skillful balancing act required from HTA agencies to constantly negotiate the right distance from the regulatory process at which to operate. We propose that HTA agencies should not be incorporated into any regulatory, auditing, or monitoring process. Finally, the relationship between health technology assessment and health care reform is discussed. It is suggested that HTA activities will contribute most during the data-driven preparation and consolidation phases of a reform process. The fast pace of events and the political turmoil characteristic of the implementation phase provide a less receptive environment for HTA contributions. PMID- 10874386 TI - Impact of the consensus conference on polycythemia vera. An opportunity to change or a sign of change? AB - OBJECTIVE: To assess the impact of guidelines on drug use issued by a consensus conference on polycythemia vera held in Paris in June 1993. 32Phosphorus (32P) was recommended for patients over 70 and/or at risk, whereas pipobroman and hydroxyurea were recommended for patients under 70. METHODS: A questionnaire was sent to all 119 departments of nuclear medicine in France 1 year after the conference to find out whether and how often they measured plasma volume and red cell mass (the recommended diagnostic tests for polycythemia vera). Time-series analyses were performed on sales of 32P, pipobroman (both virtually exclusively prescribed for polycythemia), and hydroxyurea over a 4-year span (January 1992 December 1995). RESULTS: The average number of plasma volume determinations per year did not change significantly after the conference (22 +/- 26 before vs 21 +/ 25 after). 32P and pipobroman sales were stable until July 1993, when 32P sales decreased while pipobroman sales rose steadily. Hydroxyurea sales increased over the whole period with no change in trend after the guidelines were published. CONCLUSIONS: The guidelines apparently influenced clinical practice since sales of drugs that are specifically used to treat polycythemia vera showed clear changes in trend after publication of the guidelines. This type of study seems to be an effective way of assessing the impact of consensus conferences. PMID- 10874387 TI - Uncertainty of incremental cost-effectiveness ratios. A comparison of Fieller and bootstrap confidence intervals. AB - OBJECTIVE: To compare different methods to estimate the confidence interval of the incremental cost-effectiveness ratio (ICER). METHODS: The adequacy of Fieller intervals and three methods for calculating bootstrap intervals are compared based on a simulation of 10,000 trials, using data from one trial. RESULTS: Both Fieller and bootstrap methods lead to unsatisfactory results when the difference in effectiveness is approximately zero. Where this difference is significant, the four methods for calculating confidence intervals for ICER do not give very different results, but Fieller's interval performs best. CONCLUSIONS: Since Fieller's confidence limits are relatively easy to compute compared with bootstrap simulations, we recommend using this method. PMID- 10874388 TI - Tarsal tunnel syndrome: ultrasonographic and MRI features. AB - Tarsal tunnel syndrome is a well-known but rare entrapment neuropathy involving the posterior tibial nerve in the tarsal tunnel, a fibro-osseous channel extending from the medial aspect of the ankle to the midfoot. Posttraumatic fibrosis, ganglion cyst, tenosynovitis, tumor of the nerves or other structures, dilated or tortuous veins can cause significant nerve compression in this anatomic region. Herein, we present the typical ultrasonographic and magnetic resonance features of this disorder in patient with a ganglion cyst. PMID- 10874389 TI - Basithoracic pain as first manifestation of pustulotic arthro-osteitis. AB - We report the case of a woman with atypical anterior basithoracic pain as only initial symptom of pustulotic arthro-osteitis. Early diagnosis was made only after development inferior lumbar pain, some months later. At that time, the radiological investigations revealed the osteoarticular counterpart of pustulotic arthro-osteitis. PMID- 10874390 TI - [Prevalence, morphology, and pathologic implications of ossification of lumbar ligamenta flava: a large prospective CT study]. AB - Ossifications of the ligamenta flava are described in all parts of the spine, particularly in the dorsal segment where they can produce thoracic myelopathy. Rarely reported in the cervical spine, they are only occasionally mentioned in the lumbar spine where the main reported pathology of the ligamentum flavum is the arthrosynovial cyst. To study the morphological appearances, the prevalence and the pathological implications of the lumbar ossified ligamenta flava, 1021 lower lumbar CT studies are prospectively reviewed totalizing 6080 ligamenta flava. The prevalence of clearly defined ossifications is 5.44%; it appears relatively independent from the age, sex, vertebral level and presence of spondylolysis. It significant increases with the presence and the stage of articular osteoarthritis. In more than 96% of cases, the ossifications remain confined to the lateral articular portion of the ligament and more central ossifications are very rare; therefore, the essential differential diagnosis is osteophytosis. The ossifications never cause radiculopathy except rarely, in association with more classical processes such as disk protrusions and osteophytosis. Our findings favours an essentially idiopathic and--to a lessen extent--a mechanical cause to explain ossification of the ligamenta flava. PMID- 10874391 TI - Renal failure and iodinated contrast media. A review. AB - Contrast medium-induced renal failure is reported to be a very frequent adverse effect (up to 70% incidence) that often goes undetected. Consequences are sometimes very dramatic and can even lead to irreversible renal damage and renal dialysis. Basically, four groups of mechanisms will be considered: decrease of renal perfusion and increased vascular resistance, glomerular injury, tubular injury, and obstructive nephropathy. Many risk factors are discussed, but the most important is believed to be preexisting renal insufficiency. Preventive measures are discussed. Principally, maintaining good hydration of the patient and avoiding supplementary risk factors in excess of a possible preexisting renal damage are advocated. PMID- 10874392 TI - [Accurate use of imaging in ankle sprain]. AB - Imaging ankle trauma has two goals: to evaluate bone lesions and to appreciate the capsulo-ligamentous tears. Concerning bone lesions, Ottawa's criteria specify the clinical characteristics to perform radiography in the emergency conditions. These recommendations negate the need for nearly 30% of unnecessary radiographs initially performed. Four views of the tarsis have been selected: anteroposterior, lateral, internal rotation, external oblique. Immediate screening of the ligament is very accurately performed with ultrasound, which depicts the number of bundle concerned and the extent of the lesions. This requires an experienced operator and a high quality sonograph. Stress radiography should not be performed anymore as sensitivity is around 50%. Arthrography and CT arthrography are very accurate when performed immediately but are more expensive. This report highlights the complementary role of radiography and ultrasound to evaluate ankle sprain. PMID- 10874393 TI - [Fast imaging sequences]. AB - Although conventional spin echo and gradient echo sequences are still in general use, the need to improve the temporal resolution of MRI and to study organ function has prompted to develop fast imaging techniques. These techniques are based on different approaches, but present common underlying principles which allow optimize k space fitting. As a result, both contrast and spatial resolution may be improved aside from the capability to reduce scan time. The selection of the most appropriate fast imaging technique will depend on the needed temporal resolution, spatial resolution and contrast. The aim of this paper is to review and explain the basic principles of fast imaging techniques, with emphasis on their advantages, limitations, and clinical applications. PMID- 10874394 TI - Demonstration of post-traumatic hepatic hemorrhage with carbon dioxide digital subtraction angiography. PMID- 10874395 TI - The organization wonderland of environmental health. PMID- 10874396 TI - National health insurance in the United States: a drama in too many acts. AB - Responding to the question, "Universal Health Care: How Can We Get There from Here?," this paper describes the history of governmental health insurance in the United States, analyzes the factors that have proved to be crucial to its establishment in other countries, and concludes that without a strong Labor Party, there will be no national health insurance, no universal healthcare system in the United States. PMID- 10874397 TI - Moving substance-abusing women from welfare to work. AB - Substance abuse and welfare reform are among the nation's highest priorities, and research that examines linkages between the two is of extreme importance to both government policy and the community. Welfare reform will have serious implications for substance abusers as well as for the various professionals who treat them and work to move their clients into functional recovery and self sufficiency. Within the context of welfare reform and the special needs of substance-abusing populations, the present study examines current welfare status, work status, and barriers and facilitators to gaining and maintaining employment among 100 low income women who participated in a long-term residential substance abuse treatment program in Miami, Florida. Participants completed a face-to-face interview to assess a detailed employment history and current sources of income as well as the Addiction Severity Index. Results indicate that completers of the treatment program were more likely to be working post-discharge than non completers. Similarly, the longer the length of stay in the program, the more likely the client was to be working post-discharge. Multivariate analysis indicates a high-school education, participation in the treatment center's aftercare program, and treatment duration of more than one year were independently related to work status. These data suggest that as welfare reform becomes a reality, continuing support of various types, particularly drug treatment, is needed to assist substance-abusing women in gaining and maintaining employment. PMID- 10874398 TI - The global expansion of alcohol marketing: illustrative case studies and recommendations for action. AB - As evidence emerges showing alcohol's significant share of the global burden of disease, alcohol sales have flattened in the developed countries, but sales are rising in developing and post-communist countries. A three-year study sought to assess the growing impact of global alcohol transnationals in the developing and post-communist countries. Case studies in three countries--Malaysia, Zimbabwe, and Estonia--provide concrete examples of current global alcohol marketing policies and procedures. Recommendations stress the need for national and local governments, international bodies, non-governmental organizations, and the global alcohol companies to adopt specific measures designed to achieve improved monitoring of alcohol problems, greater public awareness of alcohol's impact, stronger and more effective regulation of the alcohol trade, and greater restraint on the part of the companies. Alcohol problems are too serious and too preventable for the world to be left thirsting for action. PMID- 10874399 TI - Federalist flirtations: the politics and execution of health services decentralization for the uninsured population in Mexico, 1985-1995. AB - Around the world health services delivery systems are undergoing decentralization, responding to pressure to increase equity, efficiency, participation, intersectoral collaboration and accountability. This study examines the Mexican health decentralization efforts of the past decade to discern the motivations for the reform, the context for its implementation, the politics of its downfall, and the reform's impact at subnational levels of government. Sparked by economic crisis and pressure from international creditors for fiscal reform; demands for greater democracy, equity, and quality; and technocratic impulses to rationalize health services delivery, the decentralization reform could not overcome the authoritarian centralism of the federal government and its corporatist clients. In the end, even in the most technically capable states, the reform was unable to overcome political obstacles to decentralizing fiscal power, redistributing resources in an equitable fashion, and eliminating the inefficiencies of separate but unequal health systems for social security recipients and the uninsured population. PMID- 10874400 TI - Community noise policy in Denmark. AB - This paper summarizes an analysis of Danish policy on community noise during the last three decades seen in a public health perspective. Estimates made during this period of population exposures to noise from transport are presented, followed by data on population reactions to community noise. The analysis of the main elements of Danish community noise policy in the form of remedies, preventive actions and strategies leads to the conclusion that although exposures of the population to community noise have decreased during this period, the Danish policy was not consistent in the 1970s and 1980s. Administratively and politically, noise had low priority as an environment problem. The improvements in exposures from road traffic were probably mainly due to the legislation on physical planning, but also to lower speed limits and local traffic planning measures. During recent years important new steps have been taken, including the railway sector, but air traffic noise still represents a problem. Local authorities have, generally, been slow in giving priority to noise alleviating and preventive actions. Improved coordination at the central level is needed, and local action may raise the general awareness of the importance of our noise environment for health and well-being. PMID- 10874401 TI - The right-wing health agenda. PMID- 10874402 TI - A public health approach to mitigating the negative consequences of illicit drug abuse. National Association for Public Health Policy. PMID- 10874403 TI - Minority politics in the house of medicine: the physicians forum and the New York County Medical Society, 1938-1965. AB - The late 1930s challenged laissez-faire medicine. Recognition of serious inadequacies in the distribution of medical services stirred activists who questioned fee-for-service delivery and posited a national health program, including health insurance. The AMA and its components--state and county medical societies--counterattacked, mobilizing money and their powerful political arsenal to fight government intrusion in private medicine. The Physicians Forum, initially under the leadership of Ernst P. Boas, emerged as a formidable element within the New York County Medical Society (the largest component of the AMA). The Forum provoked discussion of medical indigence and economics, upsetting the Society leadership with independent candidate slates and telling the public that doctors spoke with more than one voice. For 25 years, the Physicians Forum altered the dynamics of the Medical Society of the County of New York. While the Forum effort to institutionalize minority opinion in the councils of medicine failed, the interchange between County regulars and Forum insurgents broadened the medical reform agenda and moved the County Society in directions that historically it had avoided. Although medical economics formed an unbridgeable chasm between County regulars and rebels, Forum members demonstrated that medicine was not monolithic, that "majority opinion [was not] ... unanimous opinion," and that doctors must speak to issues of medical and social policy. PMID- 10874404 TI - The convergence of labor and public health: a natural and critical alliance. AB - The reorganization of a Labor Caucus in APHA provides an opportunity to reflect on the community of methods and interests between the public health and labor communities. The nature of both movements is examined and the factors (knowledge base, political will, and social strategy) that define the actions of both movements are addressed. The antagonism between the prevailing political dogma of market individualism and the two movements is also discussed, with special emphasis of the need for strong government involvement to support the functioning of both labor and public health. Important issues that currently affect labor and public health are discussed. Constructive disagreements on policy choices of labor on national health insurance and occupational safety and health are examined. A warm endorsement of a critical alliance of labor and public health is made, and the revitalization of a labor caucus is cordially welcomed. PMID- 10874405 TI - Before the storm: informing and involving stakeholder groups in workplace biomarker monitoring. AB - The social, legal and ethical implications of advances in biomarker indentification have been discussed by scholars and environmental researchers, but not by the "everyday" professionals and workers who may eventually make and be affected by decisions about their workplace applications. Through the use of a hypothetical scenario, this study introduced members of various professional and occupational groups to the potential uses of biomarkers research on biological monitoring in the workplace. The purpose was to obtain opinions about how events would proceed based on the scenario, leading to a broad discussion of potential uses and abuses of biomarker-based health monitoring. Six professionally homogeneous focus groups, comprised of 1) company health professionals, 2) third party payers, 3) attorneys, 4) human resource managers, 5) non-unionized workers, and 6) unionized workers, participated in focus groups presented as "think-tank" discussions in Greenville and Charleston, S.C. Participants were given a fictitious "newspaper article" about the use of biomarker-based monitoring at a chemical plant and were asked to comment on what they thought would happen next. The discussion expanded to a general consideration of biological monitoring and its legal, social and ethical ramifications. Data was analyzed through the "immersion/crystallization" method. Few participants reported any knowledge of biological monitoring prior to the focus group session. Some had initial difficulty understanding the concept and how it differs from other means of measuring environmental risk. Although biological monitoring was previously unknown to many participants, occupational groups were relatively consistent in the issues they raised about its use in the workplace. In all groups, questions about potential discrimination against employees were raised. The general consensus was that the use of biomarker-based monitoring would result in conflict and litigation without regulations to protect employees from discrimination. Although most participants saw potential health benefits resulting from the preventive advantages associated with this technology, their concerns about its misuses were paramount. Perceptions varied as a function of occupation. Non unionized workers expressed the most concern about discriminatory uses of biological monitoring. Unionized workers, who said they believed the union would support their interests, expressed much less concern. Health professionals (company physicians and nurse practitioners) were most alarmed about the "extra work" a monitoring program would create for them. Human resource managers concentrated on the company's "damage control" efforts. Attorneys emphasized that the reliable use of such tests would establish a causal relationship between exposure and personal injury. The results of this project illustrate that people who are most likely to be affected by biomarker-based biological monitoring in the workplace readily understand and are alarmed by its legal and ethical implications. It is unlikely that this technology will be fully accepted as an environmental risk assessment tool or as a prevention strategy without stringent protection of workers' rights. This study demonstrated the value of focus groups in obtaining opinion data about an environmental risk issue that it not yet well known to the general public. PMID- 10874406 TI - Fixing EMTALA: what's wrong with the Patient Transfer Act. AB - Congress primarily enacted the Emergency Medical Treatment and Active Labor Act (EMTALA) in 1986 to prevent the denial of care to uninsured patients in emergency departments. The final version of EMTALA lacks specific protection for indigent patients and saddles hospitals and physicians with more liability than Congress initially intended. Loopholes in the law allow denial of care to patients when temporarily stabilized. Congress should ameliorate these problems through amendment of the law. PMID- 10874407 TI - Ernst P. Boas (1891-1955). PMID- 10874408 TI - Allan Macy Butler (1894-1986). PMID- 10874410 TI - Resonant X-ray scattering in the presence of several anisotropic factors. AB - The general form of the X-ray susceptibility tensor near absorption edges is found when several anisotropic factors, such as the anisotropy of local atomic environment, magnetic ordering and orbital ordering, simultaneously exist in a crystal. Different phenomenological approaches are used to obtain the explicit form of the susceptibility tensor and to find the contributions from each anisotropic factor separately as well as 'combined' terms owing to their simultaneous existence. The results of the theoretical treatment are applied to the resonant diffraction by La0.5Sr1.5MnO4 below the Neel temperature, where charge and orbital ordering coexist with anisotropy of local atomic environment and magnetic ordering. PMID- 10874411 TI - Diffraction analysis of decorated Fibonacci chains in the average unit-cell approach. AB - A novel approach to diffraction analysis of decorated quasicrystals is discussed. An average unit cell has been constructed for a decorated Fibonacci chain and used for analysis of its diffraction pattern. After some transformation of the scattering vectors, all the diffraction peaks are described by a single envelope function which is characteristic of a given decoration. It has been shown that by knowing several diffraction intensities, in a limited range of the scattering vector, it is possible to reconstruct the envelope function successfully and distinguish between different decorated structures. PMID- 10874412 TI - Minimal surfaces with self-intersections along straight lines. II. Surfaces forming three-periodic labyrinths. AB - 47 families of minimal surfaces with straight self-intersections have been derived which subdivide R3 into a finite number of congruent three-periodic labyrinths. In most of these cases, namely for 42 families, the number of labyrinths is two. Four congruent labyrinths have been found three times and eight congruent labyrinths twice. Minimal surfaces with three or six labyrinths seem not to exist. Most of these minimal surfaces are non-orientable. The surfaces of three families only are orientable ones. PMID- 10874413 TI - Atoms in crystals--from experimental charge densities. AB - An implementation is described of R. F. W. Bader's [Atoms in Molecules: a Quantum Theory (1990). Oxford: Clarendon Press] virial fragmentation of the electron density as applied to experimentally determined electron densities. It is analogous to the PROMEGA method [Keith (1993), PhD thesis, McMaster University, Ontario, Canada]. Integrated atomic properties have been determined using the models from two recent accurate charge-density studies: methylammonium hydrogen succinate monohydrate and methylammonium hydrogen maleate. PMID- 10874414 TI - Structures of nanometre-size crystals determined from selected-area electron diffraction data. AB - The structure of a new modification of Ti2Se, the beta-phase, and several related inorganic crystal structures containing elements with atomic numbers between 16 and 40 have been solved by quasi-automatic direct methods from single-crystal electron diffraction patterns of nanometre-size crystals, using the kinematical approximation. The crystals were several thousand times smaller than the minimum size required for single-crystal X-ray diffraction. Atomic coordinates were found with an average accuracy of 0.2 A or better. Experimental data were obtained by standardized techniques for recording and quantifying electron diffraction patterns. The SIR97 program for solving crystal structures from three-dimensional X-ray diffraction data by direct methods was modified to work also with two dimensional electron diffraction data. PMID- 10874415 TI - The arithmetic symmetry of monoatomic 2-nets. AB - A recent paper [Pitteri & Zanzotto (1998). Acta Cryst. A54, 359-373] has proposed a framework for the study of the 'arithmetic symmetry' of multilattices (discrete triply periodic point sets in the affine space). The classical approach to multilattice symmetry considers the well known 'space groups', that is, the groups of affine isometries leaving a multilattice invariant. The ensuing classification counts 219 affine conjugacy (or isomorphism) classes of space groups in three dimensions, and 17 classes in two dimensions ('plane groups'). The arithmetic criterion gives a finer classification of multilattice symmetry than space (or plane) groups do. This paper is concerned with the systematic investigation of the arithmetic symmetry of multilattices in the simplest nontrivial case, that is, monoatomic 2-nets (planar lattices with two identical atoms in their unit cell). We show the latter to belong to five distinct arithmetic types. We also give the complete description of a fundamental domain for the action of the global symmetry group of 2-nets on the space of 2-net metrics. PMID- 10874416 TI - Icosahedral quasiperiodic packing of fibres parallel to fivefold and threefold axes. AB - Building rules are examined for an icosahedral quasiperiodic packing of fibres with axes parallel to the ten threefold axes, first employing an experimental construction and afterwards a mathematical demonstration using the cut-and project method applied in hyperspace. As a result of this latter approach, very simple two-dimensional (2D) building rules are proposed. Similar simple 2D rules have also been proposed for the case of an icosahedral quasiperiodic packing with fibre axes parallel to the six fivefold axes [Duneau & Audier (1999). Acta Cryst. A55, 746-754]. Finally, the construction of another icosahedral quasiperiodic packing resulting from a combination of two groups of fibres respectively parallel to six fivefold and ten threefold axes is reported. A brief discussion is given on different particular mechanical behaviours which might a priori be expected from the various enantiomorphic properties of these packings. PMID- 10874417 TI - Prospects for kinematical least-squares refinement in polymer electron crystallography. AB - Least-squares refinement is unusual in the context of electron crystallography because of the sparsity of the measured intensity data set and the problems of systematic errors due to multiple dynamical scattering. With 120 unique hkl electron diffraction intensities measured from polymorphic form III of isotactic poly(1-butene), conditions for improving an existing structural model derived from initial direct structure analysis have been evaluated. The polymer crystallizes in space group P2(1)2(1)2(1) with a = 12.38, b = 8.88, c = 7.56 A and there are 8 unique atoms in the asymmetric unit. Starting with atomic positions resulting from Fourier refinement, four cycles of least-squares refinement, where the positional shifts of atomic positions were constrained, produced better bonding parameters than found before while lowering the conventional crystallographic residual, based on absolute value(F), from an overall value of R = 0.26 to R = 0.185 for the 58 most intense reflections where magnitude of absolute value(Fh(obs)) > or = 4sigma (Fh(obs)) or 0.216 for the complete data set of 120 reflections. The weighted residuals based on magnitude of absolute value(F)2 fell from 0.50 to 0.41 for the complete data set. This refinement was not improved however when attempts were made to fill in very weak intensities by default values. Also, effects of multiple-scattering perturbations were found in the irregularity of the final isotropic thermal parameters. PMID- 10874418 TI - Distribution and interference functions for two-dimensional hexagonal paracrystals. AB - The notion of a paracrystal is particularly well adapted to the calculation of the scattering interference function of distorted crystallographic lattices in which the long-range order does not exist. However, classical paracrystal modelling cannot be used directly for hexagonal lattices because it does not respect the hexagonal symmetry. Here an analytical determination of the distribution and interference functions for two-dimensional hexagonal paracrystals is presented. PMID- 10874419 TI - Fixed-scale wavelet-type approximation of periodic density distributions. AB - For a chosen unit cell, a function defined in real space (a standard signal) is considered as a crystallographic wavelet-type function if it is localized in a small region of the real space, if its Fourier transform is likewise localized in reciprocal space, and if it is a periodical function which possesses a symmetry. The fixed-scale analysis consists in the decomposition of a studied distribution into a sum of copies of the same standard signal, but shifted into nodes of a grid in the unit cell. For a specified standard signal and grid of the permitted shifts in the unit cell, the following questions are discussed: whether an arbitrary function may be represented as the sum of the shifted standard signals; how the coefficients in the decomposition are calculated; what is the best fixed scale approximation in the case that the exact decomposition does not exist. The interrelations between the fixed-scale decomposition and the phase problem, automatic map interpretation and density-modification methods are pointed out. PMID- 10874420 TI - Microdomain model analysis on the negative partial intensity in a disordered ternary alloy. AB - Local atomic arrangements expected in a short-range-ordered ternary alloy system are discussed from the theoretical viewpoint of X-ray diffraction by employing a microdomain model, initially developed for a binary alloy system [Hashimoto (1974). Acta Cryst. A30, 792-798]. It is concluded that a negative partial intensity of short-range-order diffuse scattering is caused by a mixing occupation of two relevant atomic species on the sublattice in the ordered lattice within microdomains, even though there is no heterogeneity of atomic concentration in the alloy crystal, such as a segregation of particular atomic species. PMID- 10874421 TI - Double crystals. AB - The lowest-energy way to enclose and separate two planar regions of prescribed areas is found, where the energy is given by the l1 norm ('Manhattan metric'), in which horizontal and vertical directions use less energy than other directions, as in some crystals. With the assumption that interfaces carry a fraction lambda of the energy of exterior faces, it is proved that there are three possible types of energy-minimizing double crystals. The dependence of these three types of lambda as well as the ratio of the areas of the two regions is discussed, and some paths for further study are suggested. PMID- 10874422 TI - Symmetry elements in space groups and point groups. Addenda to two IUCr reports on the nomenclature of symmetry. AB - The definition of 'symmetry element' given in the Report of the IUCr Ad-Hoc Committee on the Nomenclature of Symmetry by de Wolff et al. [Acta Cryst. (1989). A45, 494-499] is shown to contain an ambiguity in the case of space groups P6/m, P6/mmm, P6/mcc and point groups 6/m and 6/mmm. The ambiguity is removed by redefining the 'geometric element' as a labelled geometric item in which the label is related to the rotation angle of the rotation or rotoinversion symmetry operation. The complete set of different types of glide plane is shown to contain three more than the 15 that are illustrated in the 1992 Report by de Wolff et al. [Acta Cryst. (1992). A48, 727-732]. PMID- 10874423 TI - The role of emotional strategies in destructive behavior. PMID- 10874424 TI - Convex and concave, Part II: Images of emptiness in men. PMID- 10874425 TI - Silences from the deep: mapping being and nonbeing in the Piano and in a schizoid young woman. PMID- 10874426 TI - Paranoid-schizoid anxiety, triangulation, and oedipal trauma. AB - The interaction of strong aggressive and libidinal drives, various primitive intrapsychic fantasies linking somatic sensations, body parts, ego, object, and the effects of early environmental stress and trauma all produce a potential crisis in the paranoid-schizoid period of development. Certain innate methods of understanding somatic experiences as well as the interaction between internal and external reality lead to an unconscious triangulation of part objects. A frustrating, stimulating, or punitive "third" that blocks, nullifies, or overgratifies certain wishes then emerges as a pivotal object in the internal landscape. During the paranoid-schizoid, triadic process, there is a fluctuation between separation/individuation and de-differentiation/fusion. If the early triangulation process has been either exceedingly frustrating or overly stimulating in regards to "reaching the third" or "warding off the third," the infantile ego is fixed by aggressive and libidinal forces to de-differentiation experiences rather than to more separate and individuated ways of relating. Therefore, the later oedipal stage will be colored by excessive oral and anal conflicts and will be weighted on the side of primitive maneuvering based on splitting, projection, and introjection. When the child (and later the adult) becomes involved in oedipal situations marked by stimulation or frustration of triadic drives, there can be a regression to the earlier paranoid-schizoid triadic period. A case study was presented in which a patient struggled with a partial working through of these conditions in dreams and in the transference. This pulled her more in the direction of a differentiated Oedipal conflict and whole object functioning. PMID- 10874427 TI - A psychoanalytic contribution to psychic vampirism: a case vignette. AB - In conclusion, the vampire is a complex symbol of the voracious, ambiguous, horrific, transformed, and transforming, early mother figure. What may cause a regressive reaction leading to loss of object constancy? In this study I have addressed a specific physical transformation of the mother that created profound anxiety in her child. This experience can be felt as horrific. An emotional deadening may be another response. We are all susceptible to anxiety stemming from some loss of object constancy. Mr. A. certainly suffered from this phenomenon: witness his need to experience me as always available and sustained. He experienced his mother as an elusive, vague entity. We can only conjecture that his very early, preverbal upbringing was similarly experienced. To some extent, we were all exposed to some element of imperfect, inconsistent mothering. This reality, along with the other previously mentioned variables, could make us all vulnerable to vampiric anxieties. Also, I have suggested that these infantile experiences of frightening changes in the transformed mother, who may be felt as draining the life from the child, can become the basis for an organizing fantasy and lead to type of object-relating that is self-sacrificing to an extreme--a masochistic yielding to a severely demanding object. Through identification with that object a demanding, insatiable quality is acquired. Finally, this form of early experience is, perhaps, so much a part of human experience that it has become the basis of a myth of universal proportions. PMID- 10874429 TI - Psychoanalytic Therapy and the Gay Man PMID- 10874428 TI - Women and desire: the Karen Horney Lecture on feminine psychology. PMID- 10874430 TI - [The toxicity of local anesthetics]. PMID- 10874431 TI - [Pediatric anesthesia practice: attendant recommendations]. PMID- 10874432 TI - [Pediatric anesthesia practice in France: a survey of 1,526 anesthesiologists]. AB - OBJECTIVES: To assess the individual activity of anaesthetists in paediatric anaesthesia (PA), and collect their wishes about continuing education and recommendations in PA. STUDY DESIGN: Transversal, prospective study. METHODS: A questionnaire of 33 items, sent to 4,360 anaesthetists, spread over 15 health districts, working in a public or private institution. RESULTS: We gathered 1,526 replies (35%) of which 34% university hospitals, 32% public institutions and 31% private institutions. 943 physicians (63%) had no specific structure, and 1,119 (87%) considered a specialized nurse to be essential for PA. 1,127 physicians (74%) had undertaken a specific session during their formation. The practice of PA depends upon age and context. Above 1 year old, the surgery that is performed weekly was ENT (38%), abdominal and urologic surgery (28%). Mask induction was performed by 60% of the physicians in children under 5 years. 63% of the anaesthetists dreaded a laryngospasm during induction. 625 physicians undertook regional anaesthesia in children under 5 years (87% caudal anaesthesia, 48% peripheral nerve blocks). 1,029 physicians (67%) wished for recommendations in PA in children under 12 months. CONCLUSIONS: This survey showed that most of the anaesthetists wished for recommendations in their paediatric anaesthesia practice. PMID- 10874433 TI - [Acute toxic accident following lumbar plexus block with bupivacaine]. AB - We report the case of a patient who experienced ventricular dysrhythmias and seizure five minutes after the injection of 30 mL of 0.5% bupivacaine with 1:200,000 epinephrine, during a lumbar plexus block performed via the posterior approach described by Winnie. The patient who underwent his total hip arthroplasty was still anaesthetised and under controlled ventilation at the time of bupivacaine administration. Aspiration test performed before injection was negative. Normal cardiac activity and stable haemodynamic condition were restored after one hour of resuscitation including 15 electric shocks and administration of epinephrine (40 mg) and clonidine (300 micrograms). The patient was discharged without neurologic sequelae after four days in the ICU. PMID- 10874435 TI - [Bromide poisoning and false hyperchloremia]. AB - A 36-year-old female patient was admitted at three different times for neuropsychiatric disorders. No diagnoses were made during the first two hospital stays. A pseudohyperchloraemia allowed the diagnosis of bromide poisoning during her third hospital stay. Chloraemia was measured over 16 days by potentiometric, colorimetric and coulometric methods, in order to assess the analytical interferences caused by bromides. Results are reported and discussed. Bromide poisoning was treated by saline diuresis. PMID- 10874434 TI - [Meningitis after spinal anesthesia]. AB - The occurrence of meningitis after spinal anaesthesia is a very rare event. We report a case of Streptococcus sanguis meningitis following spinal anaesthesia for orthopaedic material removal. The presence of Gram positive cocci (Streptococcus sanguis) in the cerebrospinal fluid was in favour of an exogenous contamination, originating either from the patient's skin or the anaesthesiologist's oropharynx. The outcome was uneventful. The responsibility of the latter can result in legal consequences. The scrupulous compliance with guidelines prevents this risk. PMID- 10874436 TI - [Duration of antibiotic treatment in intensive care: current data]. AB - OBJECTIVE: To review the current data on the duration of an antibiotic treatment. METHODS: Analysis of recent and older articles on criteria of discontinuation of an antibiotic treatment in intensive care patients. SYNTHESIS: In intensive care patients the initiation of an antibiotic therapy is more or less codified, in spite of numerous existing problems. The duration of its maintenance, although based on scientific data depends mainly on a multitude of variables. The first step is to assess the therapeutic efficiency in considering the regression of clinical manifestations, the normalization of the acute phase reactants, the sterility of bacteriological samples and the absence of relapse at therapy discontinuation. An assessment after 48 hours is essential, in order to decide the maintenance or the modification of therapy. Finally the indication of bitherapy is considered. The theoretical duration of antibiotic therapy is determined in taking into account the involved microbial agent(s), the centre of infection, the bacterial inoculum, the patient, the presence of foreign material, and the administered antibiotic. PMID- 10874437 TI - [Meningitis after locoregional spinal anesthesia]. AB - OBJECTIVE: Meningitis is a severe and an uncommon complication of both spinal and epidural anaesthesia. This review summarizes the knowledge on epidemiology, clinical and microbiological diagnosis and the ways to prevent them. DATA SOURCES: Articles published in English and French language since 1989 has been collected on Medline database, using "meningitis", "spinal anaesthesia" and "epidural anaesthesia" as keywords. DATA SYNTHESIS: Bacterial meningitis are usually in relation with Gram positive bacterias which is a clue for an exogenous contamination. Another unusual ways of contamination are blood circulating bacterias and spreading of local infection. Aseptic meningitis has been described, in relation to introduction of irritant agents in subarachnoid space. Lumbar puncture must be done each time meningitis suspected so that it can assert the diagnosis and guide antibiotherapy. Easy hygienic guidelines has been widely published to prevent meningitis. Usually, antibiotherapy alone is sufficient to treat meningitis but with an unjustified cost and sometimes severe persistent neurologic sequelae. CONCLUSION: The unexpected appearance of meningitis during the wearing-off of a spinal anesthesia is exceptional; the possibility of death or serious sequela must be taken into account. The sources of contamination are quite frequently exogenous, the germs coming most often from the patient's cutaneous flora or the anesthetist's ENT flora. Prevention of this risk involves a rigorous respect for cutaneous disinfection and hygiene procedures. The anesthetist's medico-legal responsibilities will be called upon in case of exogenous contamination. PMID- 10874438 TI - [Are pharmacoeconomic studies pertinant?]. AB - Pharmacoeconomic studies are more and more considered to be part of both medical practice and evaluation. Today they are essential to drug registration. The implementation of clinical studies includes many difficulties, as costs have to be precisely defined. Depending on the type of clinical study, the pharmacoeconomic study may be aimed at lowering the costs; it may also be either a cost-efficacy, a cost-benefit or a cost-utility analysis. Difficulties are maximal in intensive care. An analysis of two examples issuing from the nomenclature of homogeneous groups of patients shows the inadequacy of this classification actually in Intensive care and for the analyses of drug cost. In regard to intensive care, difficulties in evaluating properly the cost of nosocomial infections is illustrated through concrete examples. PMID- 10874439 TI - [The economic impact of inadequate prescriptions]. AB - The impact of antibiotic therapy has gone to considerable expense. The review of literature demonstrates that an optimal use of economical resources can be achieved by an improvement of medical prescriptions. This improvement of prescriptions can be obtained for prophylaxy and for curative therapy. Cost savings can be as high as one-year budget for the recruitment of an infectious diseases consultant. PMID- 10874440 TI - [Consequences of antibiotic therapy to the intestinal ecosystem]. AB - Ecological impact of antibiotherapy results from the interaction between microorganisms in the ecosystems and antibiotics at which they are exposed. The amount of antibiotics use in the world is continuously increasing. The fraction devoted to human care is only about half the total amount. There are multiple other fields of usage, in agriculture, breeding and veterinary medicine. Bacterial ecosystems exposed at antibiotherapy in man are mainly the skin and the gastrointestinal and respiratory tracts. The gastrointestinal system is quantitatively predominant and the consequences of the bacterial imbalance induced by antibiotics are potentially severe. It is the reason why it is the most extensively studied, in the literature and in the present review. The origin of resistant bacteria will be briefly discussed. PMID- 10874441 TI - [Regulation of the use of antibiotics: objectives, means and perspectives]. AB - Antibiotics account for an important fraction of hospital pharmaceutical expense, especially in intensive care units. Moreover, they represent a permanent threat for bacterial ecology. Induced resistance tends, in turn, to inflate the costs. In that way, economical point of view and concerns for quality and safety of care become convergent. Restrictive and(or) educational actions to regulate the prescriptions have therefore been settled in many centres. In the present review, based on published studies, we describe some examples of the results achieved from these actions. Computer-based networks and expert systems offer promising possibilities of improvement for the near future. PMID- 10874442 TI - [Antibiotic use and bacterial resistance]. AB - The relationships between antibiotic use and bacterial resistance are not so easy to demonstrate. However, the evidence is abundant and mostly consistent. In this review, we will consider and discuss the different levels of evidence models: biological, consistent associations, dose-effect relationship, and concomitant variations. Decision-making is dependent of the local conditions. In the hospital, establishing an epidemiological diagnosis is a prerequisite to any decision, including restriction of antibiotic use. PMID- 10874443 TI - [Ecological consequences of preventive antibiotic prescriptions]. AB - Assessing the ecological impact of preventive antibiotherapy in hospital practice is an important piece in the strategies aiming at circumventing the development of bacterial resistance. In the present review of the literature, two situations will be taken into account: surgical antibioprophylaxis and selective digestive decontamination. Only the consequences of these on bacterial flora will be considered. Despite some discrepancies, only partially attributable to methodological differences, data as a whole are consistent. For antibioprophylaxis, they confirm the importance of a strict observance of the right therapeutic regimen, especially the duration of treatment. Selective digestive decontamination unquestionably encounters hazards of selecting a resistant flora. Monitoring the intestinal flora under treatment is mandatory. The indications must remain strictly limited. PMID- 10874444 TI - [Pharmaceutical use and antibiotic therapy in intensive care units]. AB - The present study has involved a sample of 750 medical or medical/surgical intensive care units. The aim was to identify the variations of antibiotic (AB) use, measured in monetary terms, and to sort out explicative variables accounting for the corresponding expense. Activity and expense data have been recorded for 1997. "Second intention" antibiotics have been defined as follows: imipenem, ceftazidime, cefpirome, cefepime, piperacillin/tazobactam, amikacin, isepamicin, vancomycin and teicoplanin. Only 60 evaluable sheets have been sent back. They include data about 28,000 admissions and 183,960 hospital days. The results are presented as means +/- standard deviation (SD) and the variability has been considered to be important if the ratio SD/mean was > 1. Nosocomial infections (NI) surveillance, antibiotic advisory board and restriction of use for some molecules were present in 95%, 67% and 78% of units, respectively. The units usually had 10 beds (range: 6-24) and the mean activity was 468 +/- 184 admissions/year and 3,066 +/- 1,454 hospital days/year. Mean duration of hospitalisation (MDH) was 6.9 +/- 2.7 days and mean omega score 114 +/- 61. Mean age of patients was 56.5 years, IGS II score 35.7 +/- 7; 29 +/- 16% of patients were mechanically ventilated for more than 48 hours and mortality rate was 17 +/- 7%. The mean number of bacterial isolates per unit was 369 +/- 323: Staphylococcus sp. 30% [including 25% of meticillin-resistant Staphylococcus aureus (MRSA)]; enterobacteria 30% (including 14% of cefotaxime-resistant isolates); Pseudomonas sp. 14% (including 40% of ticarcillin R isolates); other 26%. Pharmaceutical expense was 834 +/- 364 FF per day of hospitalisation, including 536 +/- 273 FF for drugs. Antibiotics accounted for 32% of the expense and second intention molecules for nearly 50% of antibiotic expense. More than 80% of antibiotic expense was accounted for by only 10 molecules. The mean cost/hospital day for the most expensive antibiotic, whatever the molecule ranking first, was 27 FF, for the second and third ones 18 and 14 FF. The expense for the tenth molecule was only 3 FF. There was a correlation between antibiotic expense and number of beds, number of hospital days, MDH, omega score, number of patients mechanically ventilated for more than 48 hours, mortality, number of bacterial isolates and incidence of NI. Molecules thought to be active against MRSA and ticarcillin-resistant Pseudomonas sp. accounted for 7 and 20% of total antibiotic expense, as compared to 7.5 and 4.6%, respectively, of bacterial isolates. As a conclusion, in this sample of 60 intensive care units, differences were shown for the cost of antibiotics, but the variability was low, without major discrepancies. Ten molecules accounted for 83% of total antibiotic expense. The financial impact of molecules against ticarcillin-resistant Pseudomonas sp is high. PMID- 10874445 TI - [The pharmacoeconomics of treatment of severe infections in intensive care]. AB - Pharmacoeconomics play an increasing role in rational decision-making for public health and therapeutic strategies. Relevant concepts such as benefit, efficacy, efficiency and utility have to be clearly defined. Fractionating the hospital costs implies to separate direct (medical and nonmedical), indirect and intangible costs, fixed and variable costs. The costs of side-effects, biological tests, administration of the drug and therapeutic failures have to be taken into account. From the medical point of view, the main pharmacoeconomic criterium is the cost/efficacy ratio, not the acquisition costs of the drug. Examples of the translation of these concepts to antibiotherapy in the intensive care unit are given, with analysis according to the principles of evidence-based medicine. PMID- 10874446 TI - [Preventive glycopeptide antibiotic therapy. Earlier administration?]. PMID- 10874447 TI - [On the thermostability of succinylcholine]. PMID- 10874448 TI - [Recommendations for sedation, analgesia and curarization. Short text. Societe Francaise d'Anesthesie et de Reanimation]. PMID- 10874449 TI - Changing epidemiology of bacterial infection in neutropenic patients with cancer. PMID- 10874450 TI - Serum and plasma parameters in clinical evaluation of neutropenic fever. AB - Clinicians are searching for a marker which may add to exclusion or diagnosis of relevant infection underlying neutropenic fever. The rise of such a parameter should ideally precede the date of significant microbiologic findings or justify additional intensive search for a focus of infection even in patients without pyrexia. However, the literature concerning the significance of CRP, proinflammatory cytokines and soluble adhesion molecules in the clinical evaluation of neutropenic fever is surprisingly small. In the case of procalcitonin, available data look very preliminary. Furthermore, in case of CRP, it appears that the widespread view that its determination may add substantially to the clinical evaluation of neutropenic fever is not well founded by most clinical trials listed here. Most of the studies available demonstrate several limitations such as poor design and small size of the study population. Additionally, studies were heterogeneous with respect to patients recruited (children and adults, patients with leukemia and patients with solid tumors) and compared different categories of febrile episodes. None of the investigators analyzed cost-effectiveness or impact of serial measurements of these parameters on patients' outcome. To our knowledge no single multicenter trial has been published addressing this issue. Although the group of proinflammatory cytokines and known acute-phase reactants will surely grow, more data on relevance of the available parameters in the diagnosis of neutropenic fever are needed. PMID- 10874451 TI - Antimicrobial therapy in neutropenic patients. PMID- 10874452 TI - Interventional once-daily administration of ceftriaxone in leukemia and lymphoma patients with febrile neutropenia. PMID- 10874453 TI - Ceftriaxone and cefotaxime are equally effective in the treatment of neutropenic fever. PMID- 10874454 TI - Outpatient treatment of cancer patients with fever and neutropenia. PMID- 10874455 TI - Current serological and molecular methods in the diagnosis of systemic infections with Candida sp. and Aspergillus sp. in immunocompromised patients with hematological malignancies. PMID- 10874456 TI - Antifungal prophylaxis in neutropenic patients with hematologic malignancies. PMID- 10874457 TI - Treatment of systemic fungal infections in patients with hematologic malignancies. PMID- 10874458 TI - Clinical use of hematopoietic growth factors. PMID- 10874459 TI - Recombinant human erythropoietin in the treatment of cancer. PMID- 10874460 TI - New strategies in the treatment and prophylaxis of chemo- and radiotherapy induced oral mucositis. PMID- 10874461 TI - Central venous catheter-related complications. PMID- 10874462 TI - Cytotoxic drug extravasation. PMID- 10874463 TI - Anticoagulant prophylaxis and therapy in patients with cancer. AB - Due to the various reported mechanisms by which tumours may alter haemostasis directly or indirectly, a close relationship between tumour and thrombosis is convincing. As patients with cancer represent a diverse group, the establishment of general treatment guidelines concerning antithrombotic prophylaxis and therapy requires more data than are present to date. However, in cancer patients who are candidates for surgery, chemotherapy, indwelling central venous line, or prolonged immobility, primary prophylaxis is recommended. Patients with manifest thromboembolism should receive immediate treatment and a secondary prophylaxis for more than 3 months or at least as long as the cancer is active. In patients inappropriate for oral anticoagulation, the administration of LMWH is a suitable alternative. More prospective, randomized and larger studies are required to determine the optimally tailored primary prophylaxis, the best time to start and to stop treatment, to determine the best route, dosage and pharmacological antithrombotic, and to define the role of newer anticoagulants as hirudins in the anticoagulative prophylaxis and treatment of patients with cancer. PMID- 10874464 TI - Quality of life assessment--problems and goals. PMID- 10874465 TI - Treatment of chemotherapy-induced emesis. PMID- 10874466 TI - Diarrhea and constipation. PMID- 10874467 TI - Association of DNAse sensitive chromatin domains with the nuclear periphery in 3T3 cells in vitro. AB - DNAse sensitive chromatin, putative transcriptionally competent sequences, exists either as pan-nuclear speckles in cells with nuclei which exhibit a flat geometry, or as a shell apposed to the nuclear envelope in cells with spheroidal nuclei. To test the hypothesis that DNAse sensitive chromatin is similarly associated with the nuclear periphery in cell types with a very flat geometry such as 3T3 fibroblasts, cells were subjected to hypotonic expansion to change their nuclei from a flat ellipsoid to a spheriod. This was based on the assumption that such a spatial association is not resolvable due to the interdigitation at the nuclear midplane of DNAse sensitive chromatin associated with the upper and lower nuclear surfaces. In situ nick translation was used to visualize the distribution of DNAse sensitive chromatin as a function of nuclear geometry. Both unexpanded and expanded cells exhibit DNAse sensitive chromatin as a dome at the apical side of the nucleus, i.e., that aspect of the cell facing the culture medium. The results argue for a polarized association of DNAse sensitive chromatin with the nuclear envelope and indicate that the nuclear periphery may function as a compartment for the spatial coupling of transcription and nucleo-cytoplasmic transport. PMID- 10874468 TI - Site-specific independent double labeling of proteins with reporter atoms. AB - Many types of physical, spectroscopic, and biological studies of proteins and other macromolecules are facilitated by the incorporation of reporter groups. In many cases these are single atom substitutes, for example isotopes (13C for C), or light (F for H) and heavy (Se for S) atom homologs. In some circumstances the incorporation of two different labels in the same molecule would be greatly desirable. Commonly used protein engineering methods for incorporating them can rarely cope with differential double labeling, and have other limitations such as universal, non-specific, or random incorporation. Although de novo peptide synthesis has the power to achieve highly specific labeling, the difficulties inherent in creating long sequences lead us to propose protein semisynthesis as the most practical approach. By ligating combinations of natural and labeled synthetic fragments to reform holoproteins, we can overcome any of the limitations discussed. Using cytochrome c as a model protein we show that two reporter atoms, selenium and bromine, can be simultaneously and site-specifically incorporated without significant consequences to structure and (or) function. This capability opens up the prospect of advances in a number of areas in structural biology. PMID- 10874469 TI - Is ATP a substrate for 15-lipoxygenase? AB - Lipoxygenases catalyze peroxidation of polyunsaturated fatty acids containing the 1-cis, 4-cis pentadiene structure. Linoleic (18:2), linolenic (18:3), and arachidonic (20:4) acids are the predominant substrates for this class of enzymes. Effects of 15-lipoxygenase on the hydrolysis of adenosine 5' triphosphate were investigated in vitro using soybean lipoxygenase and adenosine 5'-[gamma-32P]triphosphate. The amount of inorganic phosphate released from adenosine 5'-triphosphate was dependent upon enzyme as well as substrate concentrations, pH, and the duration of incubation. The ATPase activity with a Vmax value of 3.3 mumol.mg protein-1.h-1 and a Km value of 5.9 mM was noted in the presence of different concentrations of ATP at pH = 7.4. Phenidone, a lipoxygenase inhibitor, had no effect on this reaction. These findings suggest that soybean lipoxygenase catalyzes the release of inorganic phosphate from ATP primarily via hydrolysis. PMID- 10874470 TI - The pattern of sex chromosome kinetochore phosphorylation during nonrandom segregation in a flea beetle. AB - In the flea beetle species, Alagoasa bicolor, males have two sex chromosomes, X and Y, each of which is larger than the rest of the genome combined. These large sex chromosomes do not pair at meiosis I, and are therefore not joined at metaphase I. Nevertheless, they always segregate from each other at anaphase I. As prometaphase I progresses, the unpaired X and Y undergo reorientation from a parallel to a linear configuration. Using 3F3/2, an antibody that detects the level of phosphorylation of a kinetochore protein or proteins, we have determined that this reorientation is not accompanied by a change in the level of phosphorylation of the kinetochores of either X or Y. This implies that: i) either the reorientation does not involve the loss or gain of kinetochore microtubules, or ii) if such loss or gain occurs, it does not effect a change in the tension placed on the nonrandomly segregating kinetochores, or iii) the sex chromosomes, as in some other species, have lost the ability to sense kinetochore tension changes. Evolution of nonrandom segregation may necessitate the inability of the participating chromosomes to affect the metaphase checkpoint. PMID- 10874471 TI - Novel alpha 4-integrin ligands on an endothelial cell line. AB - The unique combination of adhesion molecules expressed on endothelial cells is thought to mediate the specificity of leukocyte-endothelial cell interactions. In this study, murine endothelial cell lines were used as a model to identify novel adhesion molecules that participate in these cellular interactions. Lymphocyte adhesion to the continuous endothelial cell lines mHEVa and mHEVc required alpha 4-integrin. Interestingly, lymphocyte alpha 4-integrin bound to VCAM-1 as well as an unknown ligand on the mHEVa cell line. We have demonstrated that this VCAM-1 independent adhesion to the mHEVa cells was not mediated by other known alpha 4 integrin ligands (fibronectin, alpha 4-integrin itself, or MAdCAM-1). Two novel alpha 4-integrin ligands (p50 and p10) were isolated from the mHEVa cell line but not the mHEVc cell line by B cell alpha 4-integrin-specific ligand binding of radiolabeled mHEV cell membrane proteins. These results provide the first direct evidence that novel ligands for alpha 4-integrin exist on membranes from endothelial cells. PMID- 10874472 TI - Proteasome from cytokine-treated human cells shows stimulated BrAAP activity and depressed PGPH activity. AB - The branched chain amino acid-preferring (BrAAP) activity of multicatalytic proteinase complex isolated from human umbilical vein endothelial cells and treated with interferon-gamma was increased more than 2-fold, which was associated with a marked increase in LMP7 expression and decreased peptidylglutamyl peptide-hydrolyzing activity. Increases in BrAAP activity in supernatants from cells treated with interferon-gamma, tumor necrosis factor alpha, interleukin-1 beta, interleukin-6, or lipopolysaccharide paralleled the increases in LMP7 expression. These findings are consistent with the conclusion that the increased BrAAP activity of LMP-containing multicatalytic proteinase complex results from incorporation of LMP7 or other LMP subunits. PMID- 10874473 TI - Effects of Bcl-2 and Bcl-XL protein levels on chemoresistance of hepatoblastoma HepG2 cell line. AB - The ratio between apoptotic promoters and repressors in the Bcl-2 family determines the chemosensitivity of cells to apoptotic stimuli. This study examines the chemoresistance of a transfected human hepatoblastoma HepG2 cell line during Taxol and Doxorubicin application. Sense bcl-2, and anti-sense bcl-XL gene fragments were separately inserted into HepG2 cells via stable transfection. The expression profile of the Bcl-2 family proteins was determined by Western blot analysis. Chemosensitivity of the transfected cells was measured by Trypan blue exclusion assay and XTT reduction assay during drug application. In the absence of Bax protein, HepG2 cells with elevated Bcl-2 protein levels did not exhibit any significant increase in chemosensitivity towards the drugs. Transfected cells with reduced Bcl-XL levels became more sensitive to the drugs, and a significant difference in IC50 values was observed. The chemosensitivity of HepG2 cells to Taxol and Doxorubicin was not affected by Bcl-2 levels, while reduction of Bcl-XL levels rendered the cells more sensitive to the drugs. This suggests that the Bcl-2 protein alone could not protect HepG2 cells from drug induced apoptosis, and that the Bcl-XL protein may be a target for gene therapy in hepatoblastoma treatment. PMID- 10874475 TI - Cognitive analytic therapy: a case study in treatment development. AB - A summary of the main literature on cognitive analytic therapy (CAT) is given. Ryle first developed CAT over 20 years ago, and use of the model is increasingly widespread in diverse settings and with various conditions. CAT stands as an example of modern dialogical approaches to therapy, and the underlying theory is consistent with that stance. The developments within training stress self reflexive practice and the maintenance of a collaborative approach. In contrast, however, to the rapid development in training and practice the research summarised here is primarily descriptive with a small number of open trials and one randomized controlled study in a physical disorder (Type I diabetes). The urgent need for randomized controlled research in this treatment is highlighted. PMID- 10874474 TI - Induction of p53-independent p21 during ceramide-induced G1 arrest in human hepatocarcinoma cells. AB - Ceramide is known to induce pRb (retinoblastoma gene product) dephosphorylation through the activation of ceramide-activated protein phosphatase (CAPP) during G1 arrest, but other molecular mechanisms linked to regulation of pRb dephosphorylation during ceramide-induced G1 arrest are poorly understood. In this paper, we investigated whether p21, a cdk (cyclin-dependent kinase) inhibitor, is involved in the induction of pRb dephosphorylation during ceramide induced G1 arrest. In SK-Hep-1 cells, the addition of ceramide resulted in pRb dephosphorylation and G1 arrest. The activity of cdk2 was inhibited in response to ceramide during this process. p21 protein and mRNA were remarkably induced, while the protein level of p53, known as a transcriptional activator of p21, was not elevated at the same condition. p21 induction was also observed in the Hep3B cells lacking a functional p53 after exposure to ceramide. Although p21 is induced in ceramide-treated Hep3B cells, Hep3B cells do not induce G1 arrest, because Hep3B cells are deficient in a functional pRb protein. To confirm that pRb is a critical target for the induction of G1 arrest by inhibiting cdk2 activity through p53-independent p21, pRb-expressing vector was transfected into Hep3B cells. After treatment with ceramide, pRb-expressing cells (pRb+/+), but not pRb-/- cells, were arrested in G1 phase. In pRb+/+ cells, ceramide-mediated G1 arrest was accompanied by the accumulation of hypophosphorylated pRb and p21 associated with cdk2. Together, these results suggest that p21, induced through p53-independent pathway, participates in the induction of pRb dephosphorylation by inhibiting cdk2 activity during ceramide-mediated G1 arrest in hepatocarcinoma cells. PMID- 10874476 TI - Rescuer and rescued: applying a cognitive analytic perspective to explore the 'mis-management' of asthma. AB - Asthma is a life-threatening and disabling condition in which medication is the major form of management. However, there is widespread evidence that asthmatic patients do not comply with their medical regimens which increases the likelihood of relapse, crisis, hospital admission and can represent a threat to the patient's life. A phenomenological qualitative study was designed which integrated the framework of cognitive analytic therapy. The aim was to explore those factors that influenced the self-care regimens of asthma sufferers. Thirty five patients were interviewed and three patterns of 'non-compliance' were identified. The clinical implications are discussed. The paper concludes with comments on the contribution of psychotherapeutic tools as a means of understanding 'risky' patient behaviours. PMID- 10874477 TI - Personal constructs of male survivors of childhood sexual abuse receiving cognitive analytic therapy. AB - OBJECTIVE: This study assessed whether male survivors of sexual abuse identified with their abusers rather than their child- or victim-selves and failed to identify with men in general or with their victims, following abusive 'acting out'. Changes in both identification and symptomatology were assessed following cognitive analytic therapy. DESIGN: A patient series within-participants design was used. METHOD: A repertory grid methodology examined four male survivors self construals before and after therapy. Measures of symptomatic distress, depression, self-esteem and distorted beliefs related to childhood sexual abuse were also obtained. RESULTS: Before therapy, patients showed little identification with the child-selves, men in general and ideal-selves. Two heterosexual patients identified with their abuser(s) whereas another, homosexual, patient negatively identified with his abuser. Patients did not identify with their victims following aggressive 'acting out'. After therapy, identification with the abuser reduced markedly; identification with victim- and ideal-self increased. Depression and distorted beliefs declined. CONCLUSION: Male survivors tend to identify with abusers and disavow their victim-hood. Sexual orientation may predict identification patterns. Cognitive analytic therapy may be a promising method of enabling survivors to integrate their abuse experience and reduce their commitment to the abuser role. PMID- 10874478 TI - A CAT-derived one to three session intervention for repeated deliberate self harm: a description of the model and initial experience of trainee psychiatrists in using it. AB - We describe a new Cognitive Analytic Therapy (CAT)-based intervention for those who repeatedly self-harm. It is specifically designed to be deliverable by staff with no training in psychotherapy. The intervention is simply manualized into sequential tasks that are mediated by new CAT-style standardized tools. A particular feature of this intervention is the deliberate use of feelings elicited in the therapist ('counter-transference') as (a) a guide to how professional poise is being threatened or lost and (b) an indicator of the appropriate focus for this very brief therapy. The psychiatrists' reflection on their elicited feelings is mediated by a new CAT tool, the 'Assessor's Response File' developed in this project. Audiotape analysis suggested that following a very brief learning period, trainee psychiatrists were able to adhere to the structure of the model and arrive at an appropriate reformulation in the first session but tended to be collusive in reciprocating the patients' dysfunctional coping styles. PMID- 10874479 TI - Effectiveness of time-limited cognitive analytic therapy of borderline personality disorder: factors associated with outcome. AB - Most patients with borderline personality disorder receive no formal treatment for their personality disorder and psychotherapy is widely believed to be necessarily intensive, of long duration and of uncertain effect. This study seeks to demonstrate the scope and limits of time-limited outpatient cognitive analytic psychotherapy. Cases were identified by standard diagnostic procedures. Most were referred from psychiatrists and were typical of inner city patient populations. At an assessment 6 months after therapy the 27 patients completing therapy were divided into 'improved' and 'unimproved' groups. The patients classified as improved no longer met diagnostic criteria for borderline personality disorder. The two-thirds still traceable were re-tested at 18 months. These groups were compared in terms of a number of pre-therapy measures and features. Poorer outcome was associated with greater severity of borderline features, a history of self-cutting, alcohol abuse and unemployment. PMID- 10874480 TI - Correspondence between delusions and personal goals: a qualitative analysis. AB - OBJECTIVES: This pilot study describes a qualitative method for exploring delusions in terms of motivational themes. DESIGN: A semi-structured interview schedule was developed on the basis of an elementary conceptual frame specifying research questions. The analysis of each case uses a structured format. Triangulation was used to check: (i) reliability of motive categories; (ii) their consistent application to delusions. METHODS OF ANALYSIS: All patients had delusions and were diagnosed as having a psychotic disorder. Two types of analysis were used: (i) Interpretative phenomenological analysis with features of grounded analysis was used to classify motives. Data from 14 participants was used for this. (ii) The second phase was an examination of a possible correspondence of themes and involved: (a) a category-led thematic analysis of the delusion in terms of motivations; (b) a category-led thematic analysis of life goals and problems again in terms of motivations; and (c) an examination of correspondence between (a) and (b). RESULTS: The classification of goals and difficulties suggested six main categories: social connection; competence; experiential base (i.e. states of mind and body); material base (e.g. housing); direction; and evaluation (i.e. how a person evaluates himself or believes others evaluate him). Four cases are presented, each exploring the correspondence of themes. CONCLUSION: The methods of analysis seemed coherent and useful. In the cases presented, the delusions appeared to relate to fundamental concerns in a person's life. PMID- 10874481 TI - Reality and discourse: a critical analysis of the category of 'delusions'. AB - Delusions are seen in psychiatric research and practice as central indicators of the psychotic loss of contact with reality. In this paper the psychiatric concept of 'delusions' is critically examined both theoretically and through the analysis of extracts from interviews with individuals diagnosed as 'delusional'. The diagnostic criteria for delusions, implausibility, idiosyncrasy, conviction and incorrigibility, are scrutinized, and the notion of reality that underlies the concept of delusions is deconstructed through the use of social constructionist approaches. The main arguments pursued in the paper are: first, that delusions are meaningful, not because they express something about the world or the speaker, but rather because they employ culturally available discourses and discursive strategies for their construction; second, that delusions are claims which are argued and negotiated in speech with similar strategies to those used by 'non-delusional' individuals; and third, that delusions are statements about the self and the world whose truth and falsity cannot be definitely settled in speech. All three claims are discussed through the analysis of extracts from two interviews with 'delusional' individuals, which focuses on the assumptions underlying the negotiation of claims on reality, the discursive strategies adopted in this negotiation and the way disputes on reality are conversationally produced and settled. PMID- 10874482 TI - The personality and cognitive-epistemological traits of cognitive-behavioural and psychoanalytic psychotherapists. AB - This is an investigation into the personality and cognitive-epistemological traits of psychotherapists from two major psychotherapeutic orientations. The purpose is to examine whether there are distinctive trait patterns associated with each orientation. Two hundred and forty-seven psychotherapists from the psychoanalytic and cognitive-behavioural orientations completed standardized personality and epistemological trait inventories. The results reveal significant differences on these measures between the two orientation groups, and suggest that different patterns of personality and cognitive-epistemological traits are associated with practitioners from these two orientations. These trait findings are summarized into two comparative descriptions, which illustrate their differences. PMID- 10874483 TI - Hospital adjustment in personality disorder patients admitted to a therapeutic community milieu. AB - This study investigated aspects of adjustment to the therapeutic community milieu in a group of personality disorder patients. Eighty-one patients consecutively admitted to the Cassel Hospital for medium/long-term residential treatment between April 1994 and October 1997 comprised the sample. The adjustment to the milieu was rated on the Hospital Adjustment Scale, while outcome was evaluated at 6-monthly intervals using a battery of self-rated and rater-based instruments. The aims of the study were: (a) to identify demographic and pre-admission clinical variables that would predict adjustment to the therapeutic regime; (b) to study the relationship between hospital adjustment and external social adjustment; and (c) to investigate the hypothesis that hospital adjustment and internally and externally directed aggression do not predict clinical outcome. The results showed that pre-admission level of global functioning and a previous history of substance misuse are the most significant predictors of hospital adjustment. No significant association between hospital adjustment and external social adjustment was found. Neither hospital adjustment not level of aggressiveness predicted outcome at 12 months. The implications of these results for therapeutic community treatment of personality disorder are discussed. PMID- 10874484 TI - Childhood trauma, dissociation and self-harming behaviour: a pilot study. AB - OBJECTIVE: Childhood trauma is known to be an important antecedent in those who engage in deliberate self-harm (DSH). We aimed to explore the mediating mechanisms between childhood trauma and subsequent DSH in a sample of women detained in a high secure setting. METHOD: From a previous incidence study into DSH, we subdivided a group of 50 women as follows: non-harmers (N = 13), infrequent harmers (N = 22) and frequent harmers (N = 15). These three groups were then compared on several measures believed to be associated with DSH. RESULTS: The frequency of DSH was related to low self-esteem, increased dissociation, anger (both inwardly and outwardly directed), impulsivity, and a history of sexual and physical abuse. When these variables were entered into a path analytic model exploring the relationship between childhood trauma and subsequent DSH, two paths emerged: one major path which linked childhood sexual abuse to DSH via increased dissociation and another, more minor association, linking childhood sexual abuse via reduced self-esteem. CONCLUSION: This study shows a strong association between high levels of dissociation and an increased frequency of self-harming behaviour. This association is theoretically plausible and has therapeutic implications. PMID- 10874485 TI - Assessing adult attachment status with clinically-orientated interviews: a brief report. AB - The object of this study was to examine the possibility of using clinically orientated interviews to gain a similar attachment classification to the Adult Attachment Interview. Little agreement on classifications was shown between the two interviews, showing insufficient evidence to suggest that it is possible to assess adult attachment status using clinically-orientated interviews. PMID- 10874486 TI - Restructuring and reallocation. PMID- 10874487 TI - E-cadherin, estrogens and cancer: is there a connection? AB - E-cadherin is a calcium-dependent, epithelial cell adhesion molecule. It has recently been implicated as a tumor suppressor. This review article contains a description of the structure, function, and regulation of E-cadherin and other members comprising the cadherin family. In particular, we discuss studies concerning the ability of estrogens to modulate E-cadherin levels in vivo. Finally, we consider the hypothesis that estrogens may promote breast, uterine and ovarian cancer by down-regulating E-cadherin levels in these tissues. PMID- 10874488 TI - HIV-1 associated Kaposi's sarcoma in an African population. AB - Eighty-seven patients with Kaposi's sarcoma were studied for human immunodeficiency virus (HIV-1) status, age and gender pattern during a three year period from 1990 to 1992. The results of this prospective study were compared with other Tanzanian series. The mean age in males decreased from 44.9 to 37.2 years for the periods of 1980-82 and 1990-92 respectively (p = 0.0001). No significant change in mean age was observed in females. The gender distribution was altered significantly: the present study recorded a male-female ratio of 2.6:1 compared with that of the pre-AIDS era 1980-82 which was 4:1. The role of HIV-1 infection as a potential cofactor of KS is discussed. PMID- 10874489 TI - Identification of cancer antigens: impact on development of cancer immunotherapies. PMID- 10874490 TI - Cancer vaccine: identification of human tumor antigens by SEREX. PMID- 10874491 TI - Immunotoxins and vascular leak syndrome. PMID- 10874492 TI - Critical determinants of neoplastic angiogenesis. PMID- 10874493 TI - Tumor infarction by targeting tissue factor to tumor vasculature. PMID- 10874494 TI - Role of alpha v integrins during angiogenesis. AB - Angiogenesis depends on specific molecular interactions between vascular cells and components of the extracellular matrix. This review focuses on the recent advances in the understanding of the mechanism of action of integrins and integrin antagonists during angiogenesis. For example, angiogenesis induced with vascular endothelial growth/permeability factor but not with basic fibroblast growth factor (bFGF) depends on integrin avb5 and Src kinase activity. In contrast, bFGF-induced angiogenesis requires integrin avb3 and functions independently of Src. Recent studies document a role for integrins and growth factor regulation of Src family kinases during angiogenesis. We also discuss the effect of av integrin antagonists on angiogenesis during tumor growth, inflammatory disease, and retinopathy and summarize recent clinical progress in using av integrin antagonists. PMID- 10874495 TI - Immunotherapy of human viral and malignant diseases with genetically modified T cell clones. PMID- 10874496 TI - Adoptive therapy of posttransplant lymphoma. PMID- 10874497 TI - Bispecific antibodies in cancer therapy. PMID- 10874498 TI - Direct identification of human tumor-associated peptide antigens and a preclinical model to evaluate their use. AB - Although the arsenal of a healthy immune system includes both circulating antibodies and cellular components such as T cells, the latter seem to be particularly important in tumor immunology. Under normal conditions, the immune system does not react to the body's cells, which may be described as expressing "self" antigens on the cell surface. When a cell becomes cancerous, however, novel antigens are expressed on the cell surface. These novel "tumor" antigens are recognized as foreign by the body's immune system, and the cells that express them are destroyed or incapacitated. Whereas antibodies may react directly with protein antigens, T cells instead recognize peptide antigens presented by class I and class II molecules of the major histocompatibility complex (MHC). All cells normally break down proteins that they have made. The class I antigen-processing pathway has evolved to display peptides produced by this breakdown process as a way to provide information to cytotoxic T cells about what the cell is making. The display of new peptides as a result of infection or transformation can stimulate cytotoxic T cells to kill the cell. In addition, antigen-processing cells such as dendritic cells engulf dead or dying cells and degradeproteins into peptide fragments. These peptides are then displayed by the MHC class II molecules and presented to T helper cells, which augment the activity of the cytotoxic T cells. Cytotoxic T lymphocytes have recently been isolated from human tumors (especially melanoma) and are critical to the development of promising immunotherapeutic agents. As we shall discuss, these cells can recognize antigens that are common to tumors from different patients. We shall also explore how advances in instrumentation and the use of transgenic mice have increased our understanding of tumor-associated peptides to the point where we can begin to strive for a peptide-based therapeutic vaccine. The caveats for such therapy will also be addressed. PMID- 10874499 TI - Augmentation of host immune responses to cancer: overcoming the barrier of tumor antigen-specific T-cell tolerance. PMID- 10874500 TI - Renomedullary interstitial cells: a target for endocrine and paracrine actions of vasoactive peptides in the renal medulla. AB - 1. The renal medulla plays an important role in regulating body sodium and fluid balance and blood pressure homeostasis through its unique structural relationships and interactions between renomedullary interstitial cells (RMIC), renal tubules and medullary vasculature. 2. Several endocrine and/or paracrine factors, including angiotensin (Ang)II, endothelin (ET), bradykinin (BK), atrial natriuretic peptide (ANP) and vasopressin (AVP), are implicated in the regulation of renal medullary function and blood pressure by acting on RMIC, tubules and medullary blood vessels. 3. Renomedullary interstitial cells express multiple vasoactive peptide receptors (AT1, ETA, ETB, BK B2, NPRA and NPRB and V1a) in culture and in tissue. 4. In cultured RMIC, AngII, ET, BK, ANP and AVP act on their respective receptors to induce various cellular responses, including contraction, prostaglandin synthesis, cell proliferation and/or extracellular matrix synthesis. 5. Infusion of vasoactive peptides or their antagonists systemically or directly into the medullary interstitium modulates medullary blood flow, sodium excretion and urine osmolarity. 6. Overall, expression of multiple vasoactive peptide receptors in RMIC, which respond to various vasoactive peptides and paracrine factors in vitro and in vivo, supports the hypothesis that RMIC may be an important paracrine target of various vasoactive peptides in the regulation of renal medullary function and long-term blood pressure homeostasis. PMID- 10874501 TI - Enalapril prevents aortic hyperreactivity and remodelling in one-kidney, one-clip hypertensive rats without reducing arterial pressure. AB - 1. The present study was designed to evaluate the blood pressure-independent effects of angiotensin-converting enzyme (ACE) inhibition on cardiovascular structure and function in one-kidney, one-clip (1K1C) hypertensive rats. 2. The study was conducted in four groups of rats: (i) uninephrectomized normotensive rats (1K); (ii) 1K1C hypertensive rats; (iii) 1K rats treated with enalapril; and (iv) 1K1C rats treated with enalapril. Enalapril treatment (20 mg/kg per day, p.o.) was started after surgery to induce hypertension or nephrectomy and continued for 5 weeks. 3. The increase in blood pressure of 1K1C rats was associated with activation of cardiac and aortic, but not plasma, ACE activity and with hypertrophy of both heart and aorta. No difference in cardiac output and in vitro systolic function was observed among the groups. Hypertrophied aorta isolated from 1K1C rats displayed increased sensitivity to phenylephrine (PE) and unaltered responses to both acetylcholine (ACh) and sodium nitroprusside compared with the 1K group. 4. Enalapril treatment effectively inhibited plasma and tissue ACE activity in 1K1C and 1K rats. Enalapril did not prevent the development of hypertension and cardiac hypertrophy nor did it change haemodynamic parameters in 1K1C rats. However, enalapril prevented the increase in aortic media thickness and cross-sectional area and restored the hypersensitivity to PE in aortic rings of 1K1C rats. The endothelium-dependent response to ACh was enhanced by enalapril in the aorta of 1K but not 1K1C rats. 5. These results suggest a role for activated local angiotensin II generation in aortic but not cardiac hypertrophy secondary to 1K1C hypertension. PMID- 10874502 TI - Nicotine administration in rabbits using Habitrol nicotine patches and nicotine nasal spray. AB - 1. Several methods are used to provide predictable and effective nicotine to experimental animals in scientific studies. Due to the expense and technical challenges of these methods, we sought suitable alternatives. Consequently, the purpose of the present study was to develop a reliable experimental nicotine protocol in rabbits that included either Habitrol nicotine patches (Novartis Consumer Health Inc., Summit, NJ, USA) or nicotine nasal spray. 2. Administration of one of three doses of nicotine (2.5, 5, or 10 mg) was accomplished daily on 13 rabbits divided between either the patch or spray groups. Systemic nicotine and cotinine levels at 0 h, 15 min and 8 and 24 h were assayed. Data were analysed by a Fisher's protected least significant difference test at P = 0.05. 3. Rabbits treated with Habitrol patches exhibited consistent and predictable systemic nicotine levels. The nicotine nasal spray produced an immediate dose-dependent response with no measurable nicotine serum levels at 8 h. 4. For nicotine administration in rabbits, nicotine patches are easy to administer and provide a nicotine serum level between 5 and 25 ng/mL, which is consistent with the average daily level found in a patient who smokes cigarettes. PMID- 10874503 TI - Deletion polymorphism of angiotensin-converting enzyme gene is associated with postprandial hyperglycaemia in individuals undergoing general check-up. AB - 1. Deletion polymorphism, DD, of the angiotensin-converting enzyme (ACE) gene is reported to be related to cardiovascular disease, which is frequently based on insulin resistance. 2. To clarify the relationship between the ACE genotype DD and plasma glucose increases after an oral glucose load, we performed 75 g oral glucose tolerance test (OGTT) in 301 nondiabetic men (age range 30-60 years) undergoing general check-up. 3. Insertion/deletion (I/D) polymorphism of the ACE gene was explored using a polymerase chain reaction. The frequency of the II, ID and DD genotypes was 0.43, 0.43 and 0.14, respectively. 4. There were no differences in baseline clinical characteristics between subjects with each ACE genotype. 5. The mean (+/- SEM) plasma glucose level at 60 min of the OGTT was significantly higher in subjects with the DD genotype (170.8 +/- 6.9 mg/dL) than in subjects with either the II or ID genotype (mean value for two groups 156.6 +/ 2.7 mg/dL; P < 0.05). Moreover, the mean percentage change of plasma glucose after 60 min of the OGTT, a marker of plasma glucose increase, was significantly higher in individuals with the DD genotype than in individuals with either the II or ID genotypes. 6. In contrast, the mean fasting plasma glucose level, the plasma glucose level at 120 min, the glucose response area and the fasting insulin level were not different between individuals with the DD genotype and individuals with other genotypes. 7. In conclusion, subjects with the DD genotype showed transiently higher levels of plasma glucose after an oral glucose load than subjects with other genotypes. Further studies are required to determine whether the association between ACE genotype and postprandial hyperglycaemia influences the incidence of cardiovascular disease and diabetes mellitus. PMID- 10874504 TI - Cardiovascular effects of chronic nitric oxide synthase inhibition in genetically hypertensive rats. AB - 1. The possible role of an endothelial defect in the hypertension of the New Zealand genetically hypertensive (GH) rat strain was assessed by examining cardiovascular responses to the nitric oxide synthase (NOS) inhibitor N omega nitro-L-arginine methyl ester (L-NAME) and the endothelium-dependent depressor agent acetylcholine (ACh). The vascular sensitivity of the hindquarter to nitric oxide (NO) was examined using the NO donor sodium nitroprusside (SNP). 2. NG Nitro-L-arginine methyl ester (10 mg/kg per day in drinking water) was given to GH and normotensive (N) rats from age 7-9 weeks, with GH and N untreated control groups. Systolic blood pressure (tail-cuff) was monitored weekly from age 5-9 weeks. At age 9 weeks, pressure responses to various vasoactive agents were measured in vivo and in the rat isolated hindquarter. Left ventricular (LV) mass was measured at the time of death. 3. NG-Nitro-L-arginine methyl ester induced a greater hypertensive effect in GH (P < 0.001) compared with N (P < 0.05) rats and caused a significant increase in hindquarter perfusion pressure in GH rats only (P < 0.01). 4. Genetically hypertensive rats had LV hypertrophy that was exacerbated by L-NAME (P < 0.01). Left ventricular hypertrophy was not induced by L-NAME in N rats. 5. The normalized response to ACh did not differ between GH and N control rats and was unaffected by L-NAME treatment in vivo and in vitro except at the highest ACh dose (3 micrograms/kg) in GH hindquarters (P < 0.01). The response to SNP was similar in GH and N hindquarters and enhanced by L-NAME in GH (0.1 microgram; P < 0.05) and N rats (0.01 microgram, P < 0.01; 0.01 microgram, P < 0.001). 6. These results suggest that the L-arginine/NO system is not deficient in GH rats and that endothelial function in the GH hindquarter is preserved. They confirm that NO is involved in mediating blood pressure in GH and N rats and raise the possibility that a non-NO-mediated mechanism may underlie ACh-induced vasodilation in GH and N. PMID- 10874505 TI - Differential secretion of adrenaline and noradrenaline in response to various secretagogues from bovine chromaffin cells. AB - 1. Differential secretion of adrenaline (Adr) and noradrenaline (NA) in response to various secretagogues was studied in bovine adrenal chromaffin cells. 2. Acetylcholine (ACh; 3-300 mumol/L), 1,1-dimethyl-4-phenyl-piperazinum (DMPP; 1 100 mumol/L), high K+ (20-60 mmol/L), calcimycin (1-100 mumol/L), histamine (0.3 30 mumol/L) and angiotensin (Ang)II (0.3-30 mumol/L) induced the secretion of a 1.3-2-fold greater percentage of NA stores than Adr stores in intact cells. 3. In beta-escin-permeabilized cells, Ca2+ (0.1-30 mumol/L) induced a greater secretion of Adr and NA in the presence of MgATP (2 mmol/L) than in the absence of MgATP. The percentage of NA secreted was 1.4- and 1.5-fold greater than that of Adr in the presence and absence of MgATP, respectively. 4. The ATP-independent phase of the Ca(2+)-dependent exocytosis is thought to be associated with the final step that ultimately leads to fusion, while the ATP-dependent phase is thought to be associated with the vesicle priming reaction. Therefore, the preferential secretion of NA in response to ACh, DMPP, high K+, calcimycin, histamine and AngII may be due, at least in part, to the greater effectiveness of Ca2+ in producing exocytosis in NA-containing cells. PMID- 10874506 TI - Endothelium-independent relaxation induced by histamine in human dorsal penile artery. AB - 1. In vitro preparations of human dorsal penile arteries were used to evaluate the effect of histamine and to characterize the histamine receptors involved in the response. 2. Cumulative administration of histamine induced a concentration dependent relaxation in precontracted arteries. The H1 receptor agonist 2 pyridylethylamine induced a biphasic response: contraction followed by dilation. The H2 receptor agonist dimaprit produced a marked relaxation. Mepyramine, a histamine H1 receptor antagonist, led to a slight but statistically significant change in the pD2 value corresponding to the relaxant phase of the H1 receptor agonist and the histamine curve. The H2 receptor antagonist cimetidine induced a marked shift in the dimaprit concentration-response curve without affecting the maximum response. Incubation with cimetidine led to a considerable loss in the sensitivity of the arteries to histamine and in the maximum relaxation. Combined treatment with histamine H1 and H2 receptor antagonists resulted in an additional displacement compared with the effect of each antagonist alone on the histamine response. The effects observed using a histamine H3 receptor agonist and antagonist suggest that the involvement of this receptor is unlikely. 3. Removal of the endothelium was unable to reverse the histamine response. Pretreatment with NG-nitro-L-arginine methyl ester, L-arginine and indomethacin had no effect on the histamine control curve. 4. In conclusion, the vasodilation of human dorsal penile artery induced by histamine seems to be mainly mediated by muscular histamine H2 receptors, without the intervention of key intracellular mediators, such nitric oxide or relaxant prostanoids. A minor population of relaxant histamine H1 receptors cannot be excluded. PMID- 10874507 TI - Presynaptic inhibitory actions of lignocaine in canine isolated, blood-perfused atrial preparations. AB - 1. Cardiac effects of lignocaine on sinoatrial nodal pacemaker activity and atrial contractility were investigated in five canine isolated, blood-perfused right atria that were perfused with heparinized blood from support dogs. The effects of lignocaine on responses to intracardiac nerve stimulation and administered acetylcholine and noradrenaline were also examined. 2. Lignocaine was injected into the support dog intravenously or administered selectively to the sinus node artery of the isolated atrium. At doses that did not produce significant depressor action (0.3, 1.0 and 3.0 mg/kg), lignocaine produced no significant changes in heart rate. A large dose of 10 mg/kg lignocaine caused significant depressor effects and slight bradycardia. Direct administration of lignocaine (0.3, 1.0, 3.0, 10.0 and 30.0 mumol) into the sinus node artery of the isolated atrium consistently caused slight negative chronotropic and rather marked negative inotropic effects. 3. After treatment with a relatively large dose of lignocaine, electrical stimulation-induced negative chronotropic and inotropic responses were significantly inhibited in a dose-related manner, but positive chronotropic and inotropic responses were slightly depressed only at an extremely high dose of lignocaine (10.0 mumol). 4. Noradrenaline-induced positive chronotropic and inotropic effects were not modified by any doses of lignocaine used (0.3, 1.0, 3.0 and 10.0 mumol). Acetylcholine-induced negative chronotropic and inotropic effects were slightly, but significantly, depressed by 10 mumol lignocaine. 5. These results suggest that a relatively large dose of lignocaine has a dominant presynaptic inhibitory action, particularly on the parasympathetic component. PMID- 10874509 TI - Quality use of medicines: where does pharmacology 'end'? Invited ASCEPT Lecture. Australasian Society of Clinic and Experimental Pharmacologists and Toxicologists. AB - 1. Pharmacology can be defined very narrowly (e.g. as solely a laboratory based specialty) or more broadly to include issues surrounding rational drug use in the community. 2. While the hypothetio-deductive approach is more clearly in evidence in laboratory and clinical pharmacological experimentation, it is no less important in the comparatively less-well charted area of drug use in the community. 3. Just as laboratory pharmacology has built new partnerships with molecular biology and genetics, the exploration of community drug use brings pharmacologists into partnership with other disciplines (epidemiology, economics, behavioural science) and other sectors (consumers, government, health professionals and industry). 4. Implementing Australia's Quality Use of Medicines Policy (1992) has led to vigorous exploration of issues and problems in medicinal drug use in the community and much scientific study of causes and potential interventions to improve the situation. 5. The Australian National Prescribing Service (1998) is attempting to bring an evidence-based approach to the many facets of prescribing and drug use, by prescribers and consumers alike. 6. Innovation commonly occurs at the interface between the boundaries of traditional disciplines. PMID- 10874508 TI - Effects of NKH477 on endothelin-1-induced renal responses in anaesthetized dogs. AB - 1. Intrarenal arterial infusion of a direct adenylate cyclase activator (NKH477; 300 ng/kg per min) increased renal blood flow, urine flow rate and urinary sodium excretion in anaesthetized dogs. 2. Intrarenal arterial infusion of endothelin (ET)-1 (2 ng/kg per min) reduced basal values of these parameters and glomerular filtration rate, which were recovered by the addition of NKH477 during ET-1 infusion. 3. These results demonstrate that NKH477 can counteract ET-1-induced antinatriuresis, mainly by restoring glomerular filtration. PMID- 10874510 TI - Cellular actions of opioids and other analgesics: implications for synergism in pain relief. AB - 1. mu-Opioid receptor agonists mediate their central analgesic effects by actions on neurons within brain regions such as the mid-brain periaqueductal grey (PAG). Within the PAG, mu-opioid receptor-mediated analgesia results from inhibition of GABAergic influences on output projection neurons. We have established that mu opioid receptor activation in the PAG causes a presynaptic inhibition of GABA release that is mediated by activation of a voltage-dependent K+ channel via 12 lipoxygenase (LOX) metabolites of arachidonic acid. 2. At a cellular level, mu opioid agonists have also been shown to open inwardly rectifying K+ channels, close voltage-gated Ca2+ channels and presynaptically inhibit glutamatergic synaptic transmission in the PAG. 3. The mu-opioid receptor-mediated presynaptic inhibition of GABAergic transmission was abolished by phospholipase A2 inhibitors and non-specific LOX and specific 12-LOX inhibitors. Cyclo-oxygenase (COX) and specific 5-LOX inhibitors did not reduce the inhibitory effects of mu-opioid agonists. 4. The opioid actions on GABAergic transmission were mimicked by arachidonic acid and 12-LOX metabolites, but not 5-LOX metabolites. The efficacy of mu-opioids was enhanced synergistically by treatment of PAG neurons with inhibitors of the other major enzymes responsible for arachidonic acid metabolism, COX and 5-LOX. 5. These results explain a previously described analgesic action of COX inhibitors in the central nervous system that was both independent of prostanoid release and inhibited by opioid receptor antagonists and they also explain the synergistic interaction of opioids with COX inhibitors. These findings also suggest new avenues for the development of centrally active analgesic agents involving combinations of lowered doses of opioids and specific 5-LOX inhibitors. PMID- 10874511 TI - Neuroexcitatory effects of morphine and hydromorphone: evidence implicating the 3 glucuronide metabolites. AB - 1. Morphine is recommended by the World Health Organization as the drug of choice for the management of moderate to severe cancer pain. 2. Education of health professionals in the past decade has resulted in a large increase in the prescribing of opioids, such as morphine, and in the magnitude of the doses administered, resulting in an improvement in the quality of pain relief available for many cancer patients. 3. However, the reported incidence of neuroexcitatory side effects (allodynia, myoclonus, seizures) in patients administered large doses of systemic morphine or its structural analogue, hydromorphone (HMOR), has also increased. 4. Clinically, increasing the magnitude of the morphine or HMOR dose administered to patients already exhibiting neuroexcitatory opioid related side effects, results in an exacerbation rather than an attenuation of the excitatory behaviours. 5. In contrast, cessation of the opioid or rotation to a structurally dissimilar opioid (e.g. from morphine/HMOR to methadone or fentanyl), usually results in a restoration of analgesia and resolution of the neuroexcitatory opioid side effects over a period of hours to days. 6. To explain the clinical success of 'opioid rotation', it is essential to understand the in vivo metabolic fate of morphine and HMOR. 7. Following systemic administration, morphine and HMOR are metabolized primarily to the corresponding 3-glucuronide metabolites, morphine-3-glucuronide (M3G) and hydromorphone-3-glucuronide (H3G), which are not only devoid of analgesic activity but evoke a range of dose dependent excitatory behaviours, including allodynia, myoclonus and seizures, following intracerebroventricular (i.c.v.) administration to rats. 8. Several studies have shown that, following chronic oral or subcutaneous morphine administration to patients with cancer pain, the cerebrospinal fluid (CSF) concentrations of M3G exceed those of morphine and morphine-6-glucuronide (analgesically active morphine metabolite) by approximately two- and five-fold, respectively. 9. These findings suggest that when the M3G concentration (or H3G by analogy) in the CSF exceeds the neuroexcitatory threshold, excitatory behaviours will be evoked in patients. 10. Thus, rotation of the opioid from morphine/HMOR to a structurally dissimilar opioid, such as methadone or fentanyl, will allow clearance of M3G/H3G from the patient central nervous system over hours to days, thereby producing a time-dependent resolution of the neuroexcitatory behaviours while maintaining analgesia with methadone or fentanyl. PMID- 10874512 TI - Bedside perspectives on the use of opioids: transferring results of clinical research into practice. AB - 1. Transference of research findings to clinical practice has been a challenge for those managing chronic pain. Generally, pain is not well controlled in hospitals and steps need to be taken to make pain control more effective. 2. Clinical trials of opioids have shown that pain can be controlled in the great majority of patients. Apart from the use of the World Health Organization Analgesic Ladder, a 'pain diagnosis' should be made and a comprehensive view of pain needs to be considered by the clinician. This would include pain and other physical symptoms, psychological issues and social and spiritual stresses. 3. Respiratory depression and tolerance for opioids are often seen as negative aspects of opioids and, therefore, may lead to inadequate control of pain. The evidence cited suggests that, in the long-term treatment of cancer pain, respiratory depression almost never occurs. The only situation that warrants caution is when an anaesthetic block or similar procedure relieves pain treated by opioids, when that patient has been receiving large doses of opioids. Long term studies with opioids show that tolerance may occur, but is not a clinical problem and should not impair their use in adequate doses to relieve the patient's pain. 4. The active morphine metabolites, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) do need to be considered when administering morphine. There seems to be considerable interindividual variation in the production and elimination of metabolites. In cases of renal failure or in the elderly, the ratios of M3G and M6G to morphine accumulate exponentially, making opioid toxicity more likely. Even different routes of administration seem to be associated with different ratios of metabolites. A knowledge of these sources of pharmacokinetic variability may lead to more effective use of the opioids in clinical practice. PMID- 10874513 TI - Pain control by immune-derived opioids. AB - 1. The nervous and immune systems communicate with each other by use of cytokines and neuropeptides. 2. Interactions between immune cell-derived opioid peptides and opioid receptors located in peripheral inflamed tissue lead to endogenous analgesia. 3. In addition to their immunological functions, immunocytes are involved in intrinsic pain inhibition. This provides new insights into pain associated with a compromised immune system, as in AIDS or in cancer. 4. The activation of opioid production and release from immune cells may be a novel approach to the development of peripherally acting analgesics. Because such drugs would be targeted towards events in peripheral injured tissue, these analgesics should lack unwanted central side effects typically associated with opioids. PMID- 10874514 TI - Role of neurotrophin receptor p75NTR in mediating neuronal cell death following injury. AB - 1. The neurotrophin receptor p75NTR has been shown to mediate neuronal cell death after nerve injury. 2. Down-regulation of p75NTR by antisense oligonucleotides is able to inhibit both sensory and motor neuron death and this treatment is more effective than treatment with growth factors. 3. p75NTR induces cell death by a unique death signalling pathway involving transcription factors (nuclear factor kappa B and c-jun), Bcl-2 family members and caspases. PMID- 10874515 TI - Spinal repair in immature animals: a novel approach using the South American opossum Monodelphis domestica. AB - 1. The adult mammalian central nervous system (CNS) is unable to regenerate following injury and repair has only been seen when implants of peripheral nervous tissue, fetal tissue or Schwann cells are used, or antibodies or trophic molecules applied. However, the immature mammalian CNS has revealed a capacity to repair without extrinsic influence. 2. The marsupial mammal provides a unique opportunity to access the immature CNS without invasive in utero surgery. In particular, the South American opossum Monodelphis domestica is an ideal animal for spinal cord injury studies examining the ability of the immature CNS to repair after injury. 3. The Monodelphis spinal cord may be examined for its response to injury either as an in vitro or in vivo system and, therefore, is a flexible model, allowing many different questions to be addressed by the most suitable approach. 4. The immature Monodelphis CNS was able to support fibre growth that reappeared 4 days after a crush at P3-P8 in vitro. Conduction was also restored at this time, accompanied by synaptic connections. 5. A cut lesion performed in vivo on Monodelphis spinal cords at P7 took longer to repair, with fibres reappearing across the injury site 2 weeks after the lesion; greater disruption to structure was noted both during early stages of repair and in adulthood. 6. Neural pathway tracing with dextran amine from the lumbar cord to the brain in adult Monodelphis, which received spinal lesions at P7, revealed a similar distribution of labelled cells in brainstem and mid-brain nuclei to that of control animals. 7. Studies of the locomotor behaviour of adult Monodelphis that had received either a cut or crush lesion at P7-P8 showed remarkably similar abilities to control animals when performing complex tasks. 8. The results of spinal cord injury studies with the immature Monodelphis CNS may help in the development of treatments for spinal injury patients. PMID- 10874516 TI - Neuronal response to physical injury and its relationship to the pathology of Alzheimer's disease. AB - 1. Central nerve cells undergo a stereotyped regenerative response following physical injury. 2. This reaction involves adaptive changes within the axon and cell body of origin, directed at sprouting and synaptogenesis. 3. Intimately associated with the regenerative response are specific alterations to cytoskeletal proteins, including the neurofilament (NF) triplet. 4. The morphological and neurochemical alterations to NF within axons following injury are reminiscent of plaque-associated dystrophic neurites (DN) in early Alzheimer's disease (AD). 5. Associated changes in perikaryal NF resemble Alzheimer neurofibrillary tangle pathology, while growth-associated sprouting markers are localized to the abnormal neurites of AD. 6. The present review postulates that beta-amyloid plaques in AD cause physical damage to local nerve cell processes and it is the chronic stimulation of the stereotyped response to injury that results in the end-stage pathology and neurodegeneration associated with AD. PMID- 10874518 TI - Biopharmaceutical considerations in topical ocular drug delivery. AB - 1. Despite the accessibility of the front of the eye, efficient delivery of drug to treat various ocular disorders is a challenge to the formulation scientist. The majority of ophthalmic medications are formulated as eye drops. Due to anatomical constraints, the volume that can be administered is limited to approximately 30 microL. This, together with the efficient clearance system that exists in the front of the eye, makes it difficult to maintain an effective pre ocular drug concentration for a desired length of time. Various formulation strategies have been used to increase pre-ocular retention of eye drops. The most successful of these has been the inclusion of viscosity enhancing polymers, particularly those able to interact with the mucous layer on the eye surface or those that can undergo a transition from a solution to a gel under the conditions of the pre-ocular area. 2. When the target site is intra-ocular, drug must be absorbed from the pre-ocular region into the eye. The main route for absorption is across the cornea. However, absorption of drug across the cornea is inefficient due to its impermeable nature and small surface area. Thus, the intra ocular bioavailability of topically administered medications is typically less than 10%. 3. Corneal permeability favours moderately lipophilic compounds. These compounds often have a low aqueous solubility. Problems in ocular drug delivery and formulation are compounded for poorly soluble drugs that must be formulated as suspensions. 4. Reformulation of ophthalmic suspensions as solutions has many advantages. This may be achieved by complexation using cyclodextrins. Solubilization using cyclodextrins can overcome many of the formulation problems. However, it is unclear as to their potential for improving ocular bioavailability, which is seemingly drug dependent and may be influenced by both the physicochemical properties of the drug and the complex formed. PMID- 10874517 TI - AM424: history of a novel drug candidate. AB - 1. Leukaemia inhibitory factor (LIF) is a 180 amino acid single-chain protein, named after its effect on haematopoietic cells. Leukaemia inhibitory factor belongs to a group of cytokines that includes ciliary neurotrophic factor, interleukin (IL)-6, IL-11, cardiotrophin-1 and oncostatin M. All group members use the gp130 signal transducing subunit for intracellular signalling, but show differences in biological effect. 2. Research over the past 6-8 years has shown LIF to have potent neuromuscular activity. In vitro and in vivo studies on axotomy and nerve crush models demonstrate a powerful effect of LIF in enhancing the survival of both motor and sensory neurons, while reducing denervation induced muscle atrophy. In models of both axotomy induced neuronal death and in the wobbler mouse, LIF is active at doses as low as 1 microgram/kg delivered systemically. 3. In muscle, LIF will increase the rate of muscle regeneration in vivo when applied exogenously after injury and will stimulate intrinsic muscle repair following its targeted release to dystrophic muscle in the mdx mouse. Leukaemia inhibitory factor may also have a role as an adjunct to myoblast transfer therapy, with studies showing that the transplantation of genetically competent myoblasts into mdx mouse muscle is enhanced when cells are injected with LIF. 4. Distribution and pharmacokinetic studies have been conducted in primates with doses of 20 micrograms/kg recombinant human LIF given subcutaneously over 2 weeks tolerated without major side effects. 5. A pharmaceutical form of recombinant human LIF (AM424; AMRAD Operations, Richmond, Victoria, Australia) entered human clinical trials during 1997 and a phase I clinical trial in healthy volunteers has been completed. A phase I repeat dose study has also been completed in cancer patients undergoing chemotherapy. The primary indication for a phase II study is the treatment of chemotherapy induced peripheral neuropathy. Other potential indications include muscle wasting diseases, acute nerve trauma and motor neuron disease. 6. The role of LIF in modulating nerve loss should make it an ideal candidate for the treatment of a number of neurological conditions. The phase I study represents the first trial in a programme for the clinical development of AM424. PMID- 10874519 TI - Tris lipidation: a chemically flexible technology for modifying the delivery of drugs and genes. AB - 1. One of the major challenges in the development of pharmaceuticals is their formulation with other materials to give them the desired bioavailability profile when administered into the body. 2. We have developed a flexible platform technology (Tris lipidation) to simply and effectively alter the lipophilicity of drugs. As implied by the name, the technology uses the common buffer Tris as a linker between the drugs of interest and a domain of variable hydrophobicity. 3. We demonstrate, using a mouse melanoma model, that Tris-lipidated conjugates of the widely used cytotoxic and anti-inflammatory drug methotrexate (MTX) display enhanced potency in the local treatment of tumours and reduced systemic toxicity when compared with the unconjugated drug. 4. With genes now being predicted to be the pharmaceuticals of the future, we show that Tris-lipidated cationic peptides can efficiently deliver DNA into (transfect) cells in culture. Furthermore, by comparing the abilities of variants of these Tris-based cationic lipids to transfect cultured cells, we demonstrate that modifications made to variable regions of Tris-lipidated compounds can dramatically alter their delivery profiles. PMID- 10874520 TI - Clinical relevance of hyperhomocysteinaemia in atherothrombotic disease. AB - High fasting plasma homocysteine levels (> 12 to 15 mumol/L) are commonly encountered in clinical practice and are associated with increased risk of atherothrombotic disease. Treatment with folic acid (1 to 5 mg/day) is inexpensive and effective in normalising plasma homocysteine levels. High plasma homocysteine levels after methionine loading (> 40 to 50 mumol/L) are also common and can be treated with pyridoxine-based regimens (50 to 250 mg/day). As compared with fasting plasma homocysteine levels, the association between high postmethionine loading plasma homocysteine levels and atherothrombotic disease has been less extensively studied. There is reasonable, but not clearly definitive, evidence that high plasma homocysteine levels are causally related to atherothrombotic disease. Results of randomised trials of homocysteine-lowering treatment with clinical end-points will be available in 4 to 6 years. At present, a reasonable policy for the practising clinician would be to consider homocysteine-lowering treatment in individuals at very high risk of atherothrombotic disease, such as patients with clinically manifest atherothrombotic disease with onset before 55 years of age, patients with end stage renal disease, and healthy subjects with a strong family history of early onset atherothrombotic disease. Such a policy should be reassessed as the results of randomised trials become available. PMID- 10874521 TI - Complete estrogen blockade for the treatment of metastatic and early stage breast cancer. AB - Complete estrogen blockade has long been sought as a more effective means of controlling breast cancer compared with single agent endocrine therapy. This approach may be accomplished through the use of agents which reduce estrogen production combined with agents that prevent the activity of estrogen at the cellular level. For prostate cancer, another hormonally responsive malignancy, this approach has not been successful at improving survival compared with that achieved with single agent therapy. Preclinical information is contradictory for many promising combinations and may not reflect the true nature of in vivo interaction between agents. For premenopausal patients with metastatic breast cancer, the combination of a luteinising hormone-releasing hormone (LHRH) agonist and tamoxifen is clearly effective, but whether the combination is more effective than either single agent is still controversial. Similar response rates and overall survival were reported with goserelin or goserelin plus tamoxifen by Jonat et al. in 1 randomised, prospective study, but the addition of tamoxifen improved time to progression. A second trial comparing buserelin plus tamoxifen with either single agent reported superior efficacy in terms of response rates, disease-free survival and overall survival with combination therapy. A meta analysis of 4 randomised trials making similar comparisons, demonstrated significant improvement in median overall survival, progression-free survival, response rate, and duration of response with the combination of a LHRH agonist (goserelin or buserelin) and tamoxifen in premenopausal breast cancer patients with metastatic disease. For postmenopausal women with metastatic breast cancer, the addition of an aromatase inhibitor to tamoxifen has yet to be prospectively compared to single agent therapy. Use of endocrine combinations in the treatment of early stage breast cancer is under investigation. Preliminary results of some of the ongoing adjuvant therapy trials indicate that the combination of a LHRH agonist and tamoxifen may have similar efficacy to cyclophosphamide, methotrexate, and fluorouracil chemotherapy in premenopausal women with estrogen receptor-positive tumour. Addition of LHRH agonist therapy in premenopausal patients with estrogen receptor-positive tumour who had maintained the ovarian function following chemotherapy [cyclophosphamide, doxorubicin (adriamycin), fluorouracil, and tamoxifen], also led to a reduction in the risk of recurrence. These studies have identified a sub-population of patients who may benefit from the addition of combination endocrine therapy. Overall, the issue is quite complex and the data from many ongoing trials are still awaited with anticipation to further delineate the role of complete estrogen deprivation in this disease. PMID- 10874522 TI - Tremor-predominant Parkinson's disease. Approaches to treatment. AB - Parkinson's disease is a neurodegenerative disorder that manifests clinically with variable degrees of tremor, muscle rigidity, bradykinesia and postural instability. Tremor-predominant Parkinson's disease is characterised by prominent tremor of one or more limbs with a relative lack of significant rigidity and bradykinesia. Despite the lack of other disabling motor symptoms, the tremor of tremor-predominant Parkinson's disease can be very disabling, especially if a postural and kinetic component exists. A wide variety of treatments for Parkinson's disease tremor are currently available and include use of oral medications, injections with botulinum toxin and neurosurgical procedures. Some of the first line medications (levodopa, dopamine agonists, anticholinergics) are very effective in controlling tremor. However, some patients with Parkinson's disease tremors are unresponsive to first line drugs and treatment with second line medications (clozapine, amantadine, clonazepam, propranolol, neurontin) should be attempted. In the small number of patients with disabling tremor that is refractory to all medications, neurosurgical intervention should be considered. Both thermocoagulation and deep brain stimulation at several different neuroanatomical sites (thalamus, globus pallidus, subthalamic nucleus) offer good to excellent tremor control with relatively low risk to the patient. PMID- 10874523 TI - Immunosuppression in older renal transplant patients. AB - Renal transplantation procedures in patients older than 60 years of age have clearly improved in recent years. In the cyclosporin era, graft and patient survival are good. However, older patients exhibit a higher mortality, especially from infectious and cardiovascular causes, than young patients. In this article we review the immunosuppressive treatment in older patients, analyse what drugs can be used and finally propose several immunosuppressive combinations to treat this group of patients. Currently, new immunosuppressive drugs enable more flexible immunosuppressive protocols. Nevertheless, to avoid overimmunosuppression, elderly patients should be treated with lower doses and fewer immunosuppressive drugs. PMID- 10874524 TI - Diuretic therapy in elderly heart failure patients with and without left ventricular systolic dysfunction. AB - Long term prescription of diuretics for heart failure is very prevalent among elderly patients, although the rationale for such a treatment strategy is often unclear, as diuretics are not indicated if volume overload is absent. The concept of diastolic heart failure in the elderly might particularly change the role of diuretic therapy, since diuretics may have additional adverse effects in these patients. This paper reviews the effects of diuretic therapy in elderly patients with heart failure, emphasising the differences between patients with normal and decreased left ventricular systolic function. Studies on diuretic withdrawal in elderly patients with heart failure are discussed, with emphasis on issues involved in decision making such as diuretic dose reduction and withdrawal in elderly patients and factors that have been established to predict successful withdrawal. Existing guidelines on the prescription of diuretics in elderly patients with heart failure with normal and decreased left ventricular systolic function and in those with diastolic heart failure are also discussed. By reducing intravascular volume, diuretics may further impair ventricular diastolic filling in patients with diastolic heart failure and thus reduce stroke volume. Indeed, preliminary studies demonstrate that diuretics may provoke or aggravate hypotension on standing and after meals in these patients. Therefore, it is suggested that elderly patients with heart failure with intact left ventricular systolic function should not receive long term diuretic therapy, unless proven necessary to treat or prevent congestive heart failure. This implies that physicians should carefully evaluate the opportunities for diuretic dose tapering or withdrawal in all of these patients, and that a cautiously guided intermittent diuretic treatment modality may be critical in the care for older patients with heart failure with intact left ventricular systolic function. PMID- 10874527 TI - Population screening for autoimmune thyroid disease. AB - Whether or not healthy adults in the community would benefit from screening for autoimmune thyroid disease is controversial. Although the prevalence of unsuspected overt thyroid disease is low, a significant proportion of subjects tested will have evidence of mild thyroid failure or excess. This article assesses whether subclinical thyroid disease is of sufficient clinical importance to warrant screening and, once detected and confirmed, to justify therapy. Population screening for autoimmune thyroid disease is assessed against recently revised screening criteria, using data from epidemiologic studies. Recommendations are proposed that may be applied in any iodine-replete community. PMID- 10874528 TI - The genetics of Graves' disease. AB - Graves' disease is a complex autoimmune disorder in which several genetic susceptibility loci and environmental factors are likely to contribute to the development of disease. HLA and the CTLA-4 gene region have been established as susceptibility loci, although the magnitude of their contributions seems to vary between data sets and geographic populations. Genome-wide searches are beginning to identify new loci, including GD-1, GD-2, and GD-3, although these loci have only been found in one data set. Additional loci are likely to be identified via a combination of genome-wide linkage analysis and allelic association analysis of candidate genes. PMID- 10874526 TI - Subcutaneous hydration by hypodermoclysis. A practical and low cost treatment for elderly patients. AB - As the world's population ages, chronic and degenerative diseases are rising. This scenario demands the development of new treatment techniques with lower costs, which are as efficient as the existing ones. Hypodermoclysis is the infusion of fluids into the subcutaneous tissue with a butterfly needle. This technique may be used for isotonic fluid replacement and to administer cytosine arabinoside, clodronate, antibiotics and narcotic analgesics. This review evaluates the evidence supporting the use of hypodermoclysis to treat elderly patients with dehydration and patients with terminal cancer, and discusses its indications, adverse effects and perspectives. A MEDLINE search of the last 30 years was done to recover all available literature. Hypodermoclysis therapy is a safe and effective method to provide fluids and narcotic analgesic therapy in elderly patients that are mild and moderate dehydrated and in patients with cancer. It seems a good option to provide antibiotics, but there is a need for more studies to evaluate this indication. PMID- 10874525 TI - Thrombolytic therapy in acute myocardial infarction. AB - Early reperfusion of thrombotically occluded coronary arteries by thrombolytic therapy has become a routine option in initial therapy of acute myocardial infarction. Many efforts have been made to improve the biological properties of thrombolytic agents in terms of fibrin specificity, plasma half-life and resistance to natural plasma inhibitors, to improve adjuvant therapy and to shorten the 'pain to reperfusion' time. Numerous randomised, multicentre trials have analysed the benefit of the various thrombolytic agents and regimens, which has enabled the creation of a 'current standard of therapy'. This review presents an update on available thrombolytic agents, their biochemical and pharmacological properties and results from clinical trials. PMID- 10874529 TI - New insights into the thyroid-stimulating hormone receptor. The major antigen of Graves' disease. AB - The receptor for thyroid-stimulating hormone is one of the most interesting hormone-binding sites because of its close association with common human diseases, including thyroid nodules and Graves' hyperthyroidism. This article discusses the structure and biosynthetic processing of this elusive glycoprotein, whose paucity and instability have impeded its isolation from natural sources. Topics include cleavage and subunit structure, variant species, and structural modeling, the thyroid-stimulating hormone receptor as the major autoantigen in Graves' disease, and a summary of recent efforts to replicate the symptoms of this uniquely human disease in animal models. PMID- 10874530 TI - Understanding the immunology of Graves' ophthalmopathy. Is it an autoimmune disease? AB - The notion that Graves' ophthalmopathy is an autoimmune disease has been based primarily on the clinical associations between it and Graves' hyperthyroidism, and on the frequently beneficial response of the disease to immunosuppressive therapy. Recent advances in molecular biology have led to new insights into the pathogenesis that support an autoimmune basis for this condition. In particular, there is now compelling--although not definitive--evidence that, if verified, would represent disease transfer by thyroid-secreting hormone receptor-sensitized T cells. Future studies using animal models and in vitro systems will allow definitive identification of the immune cell types, autoantigens, and autoantibodies involved in the pathogenesis of Graves' ophthalmopathy. PMID- 10874531 TI - An evidence-based approach to the treatment of Graves' ophthalmopathy. AB - A recent survey demonstrated considerable controversy about the therapeutic approach to a patient with Graves' ophthalmopathy. Among various reasons for this disagreement is the lack of an accepted way to assess the outcome of a given therapeutic intervention. An evidence-based approach requires an objective and meaningful assessment of therapeutic outcomes in prospective, randomized studies in which confounding factors (e.g., smoking, disease activity, and state of the thyroid) are accounted for and controlled. On the basis of the available studies, a treatment approach can be selected. PMID- 10874532 TI - Treatment of Graves' disease: the advantages of surgery. AB - The authors and others believe that surgery (thyroidectomy) is underused in the treatment for patients with Graves' disease. It is the most rapid and consistent method of making the patient euthyroid; it avoids the possible long-term risks of radioactive iodine; and it provides tissue for histologic examination. Children, young women, pregnant women, and patients with coexistent thyroid nodules are ideal candidates for thyroidectomy. It also is the treatment of choice for patients with Graves' ophthalmopathy. Patients should be rendered euthyroid before thyroidectomy. Although the operation is technically more difficult than operating on patients with nontoxic goiter or thyroid neoplasms because of the vascularity of the thyroid gland, this difference is small, and the complication rates are low. The authors recommend the Hartley-Dunhill operation (total thyroidectomy on one side and subtotal thyroidectomy on the other side, leaving about 4 to 5 g of thyroid tissue) for most patients and total thyroidectomy for patients with Graves' ophthalmopathy. In patients with recurrent or persistent thyroid cancer who fail to respond to surgery and radioactive iodine ablation, immunosuppressive therapy should be considered. PMID- 10874533 TI - Anti-TSH receptor antibodies in clinical practice. AB - Anti-thyroid stimulating hormone receptor antibodies are pathophysiologic and clinical indicators in autoimmune thyroid diseases, not only in Graves' disease. The detection of these antibodies is useful for diagnostic and management purposes. The presence and titers of anti-TSH receptor antibodies, however, have to be interpreted in light of the clinical and other biological characteristics of each patient. Newer, more sensitive assays of anti-TSH receptor antibodies may increase their significance in the diagnosis and management of autoimmune thyroid diseases and Graves' disease. PMID- 10874534 TI - The genetics of Hashimoto's disease. AB - Despite strong epidemiologic evidence in favor of a genetic component in the etiology of HT, few hereditary risk factors have been consistently identified. These factors include the HLA and CTLA-4 genes. The mechanisms by which these genes confer increased susceptibility to HT are unclear. The identification of these genes has failed to explain completely the large hereditary effect observed in families of patients. More substantial genetic determinants must be hidden in the folds of the human genome and will most likely be detected in the near future. The powerful approach of linkage analysis will be supported by advancements in the description of the human genome and by technologic improvements in the ability to process large amounts of biologic data. Knowledge of such determinants will provide predictive tools to be used on clinical grounds and invaluable insight into the pathogenesis of this puzzling disease. PMID- 10874535 TI - Thyroid cell apoptosis. A new understanding of thyroid autoimmunity. AB - Apoptosis is a highly regulated mechanism of cell death involved in normal development, immune regulation, and homeostasis. Abnormal apoptotic activity has been implicated in a variety of diseases including cancer, autoimmunity, and degenerative disorders. In the thyroid, altered cell death may play a role in the pathogenesis of autoimmune disorders such as Hashimoto's thyroiditis and Graves' disease. Apoptosis-signaling pathways can be initiated through activation of death receptors or in response to cellular damage, such as in gamma irradiation. It has been demonstrated that Fas, tumor necrosis factor, and tumor necrosis factor-related apoptosis-inducing ligand pathways are present and functional in the thyroid, although the expression of these molecules and their roles in thyroid autoimmunity have been debated. Thyroid apoptosis is regulated at multiple levels, including receptor and ligand expression, and the expression of antiapoptotic proteins, such as FAP-1 and Bcl-2. These factors may provide potential mechanisms for modifying the pathogenesis of autoimmune thyroid disease. PMID- 10874536 TI - The immune response to the iodide transporter. AB - In addition to physiologic, diagnostic, and therapeutic implications, the recently cloned and characterized sodium iodide symporter (NIS) also may play an important role in the pathogenesis of autoimmune thyroid disease. Sodium iodide symporter expression patterns characteristically are changed in autoimmune thyroid disease, including Graves' disease and Hashimoto's thyroiditis, which may be caused, in part, by the regulation of sodium iodide symporter expression of cytokines involved in the pathogenesis of autoimmune thyroid disease. Further, there is increasing evidence that NIS-directed antibodies are present in sera from patients with autoimmune thyroid disease, and these antibodies also may affect NIS functional activity. PMID- 10874538 TI - Recognizing, understanding, and treating postpartum thyroiditis. AB - Postpartum thyroiditis is the most common endocrinologic disorder, with an incidence that varies geographically from 5% to 10%. It has important clinical sequelae including symptoms of hyperthyroidism, hypothyroidism, and depression. Long-term follow-up of women who experience postpartum thyroiditis reveals a high recurrence rate in subsequent pregnancies. Postpartum thyroiditis is an autoimmune disorder, and thyroid antibody-positive women in the first trimester have a 33% to 50% chance of developing thyroiditis in the postpartum period. Whether or not to screen for postpartum thyroiditis remains controversial. PMID- 10874537 TI - When to treat mild hypothyroidism. AB - The availability and wide acceptance of TSH assays for primary assessment of thyroid function has led to the recognition that mild thyroid hormone deficiency is characterized by elevation of the serum TSH concentration despite a normal free thyroxine level. Other conditions can also cause isolated serum TSH elevation, and these conditions can be distinguished from mild thyroid failure usually based-on clinical and circumstantial observations alone. Thyroxine treatment of patients with mild hypothyroidism has been shown in most, but not all, clinical trials to lower atherogenic lipid levels and relieve certain somatic and neuropsychiatric symptoms. Such treatment also prevents the progression to overt hypothyroidism, which is particularly likely in patients who are older, who have circulating thyroid autoantibodies, or who have a serum TSH greater than 10 mU/L. After the optimal thyroxine dose has been defined, long term monitoring of patients with an annual clinical evaluation and serum TSH measurement is appropriate. The high prevalence of mild hypothyroidism, particularly in older women, and its subtle clinical presentation have led some authorities to recommend a low threshold for case-finding or routine population screening for the disorder. PMID- 10874539 TI - Growing an interest in autoimmune thyroid disease. An interview with Robert Volpe, MD, FRCP(C) PMID- 10874540 TI - Beyond willow bark: aspirin in the prevention of chronic disease. PMID- 10874541 TI - Consumption of contaminated lake fish and reproduction. PMID- 10874542 TI - Reduced incidence of colorectal adenoma among long-term users of nonsteroidal antiinflammatory drugs: a pooled analysis of published studies and a new population-based study. AB - Chronic treatment with nonsteroidal antiinflammatory drugs (NSAIDs) has been associated with a reduced risk of colorectal cancer, but less information is available on the relationship between NSAIDs and colorectal adenoma. We carried out a population-based cohort study with nested case-control analysis to determine the association between the use of aspirin and individual NSAIDs and the risk of colorectal adenoma. The General Practice Research Database in the United Kingdom was the source population. We followed 943,903 persons who were 40 79 years of age and free of colorectal adenoma or other cancer at baseline, which varied between January 1994 and September 1997. There were 1,864 incident cases of colorectal adenoma, for an incidence rate of 6.8 per 10,000 person-years. Compared with non-users, long-term users (1 year and more) of nonaspirin NSAIDs had a 40% decreased risk of colorectal adenoma (relative risk = 0.6; 95% confidence interval = 0.4-0.9). Long-term NSAID use was still associated with a reduced risk 1 year after stopping NSAID treatment. Use of most individual NSAIDs conferred a reduced risk. The risk of developing colorectal adenoma was reduced in long-term users of aspirin at doses of 300 mg daily (relative risk = 0.6; 95% confidence interval = 0.4-1.0), but reduced risk was not evident with daily doses of 75 and 150 mg aspirin. These results add further support to the value of NSAIDs as a candidate for primary prevention of colorectal tumors. PMID- 10874543 TI - Differential effects of aspirin and non-aspirin nonsteroidal antiinflammatory drugs in the primary prevention of myocardial infarction in postmenopausal women. AB - The antiplatelet effect of aspirin reduces the risk of clinical manifestations of atherothrombosis by approximately 25% in secondary prevention settings. Data are limited in primary prevention of coronary heart disease, and even more in women. Here, we estimate the effects of aspirin and non-aspirin nonsteroidal antiinflammatory drugs in the primary prevention of myocardial infarction in postmenopausal women. We followed a cohort of 164,769 women, 50-74 years of age, registered in the General Practice Research Database in the United Kingdom, from January 1991 through December 1995. For aspirin and non-aspirin nonsteroidal antiinflammatory drugs, the risk of myocardial infarction associated with current use was compared with risk in non-users, using a nested case-control analysis. Overall, the relative risk of myocardial infarction associated with current use of aspirin of more than 1 month's duration was 0.56 [95% confidence interval (95% CI) = 0.26-1.21], and that of nonfatal myocardial infarction was 0.28 (95% CI = 0.08-0.91). Chronic use of nonsteroidal antiinflammatory drugs was not associated with a protective effect (relative risk = 1.32; 95% CI = 0.97-1.81). These findings indicate that incomplete and reversible inhibition of platelet cyclooxygenase by non-aspirin nonsteroidal antiinflammatory drugs is not sufficient to produce clinically detectable cardiovascular protection comparable with that achieved by low-dose aspirin through irreversible inactivation of platelet cyclooxygenase. PMID- 10874544 TI - Parental consumption of contaminated sport fish from Lake Ontario and predicted fecundability. AB - Wildlife studies suggest that consumption of contaminated fish from the Great Lakes may expose humans to polychlorinated biphenyls and persistent chlorinated pesticides. To assess whether time to pregnancy or fecundability is affected, we conducted a telephone survey in 1993 with female members of the New York State Angler Cohort Study who were considering pregnancy between 1991 and 1994 (N = 2,445). Among the 1,234 (50%) women who became pregnant, 895 (73%) had a known time to pregnancy. Upon enrollment into the cohort in 1991, both partners reported duration and frequency of Lake Ontario sport fish consumption. We estimated lifetime exposure to polychlorinated biphenyls from recent consumption and used a discrete-time analog of Cox proportional hazards analysis to estimate conditional fecundability ratios and 95% confidence intervals (CIs) for fish consumption among couples with complete exposure data who discontinued birth control to become pregnant (N = 575). Maternal consumption of fish for 3-6 years was associated with reduced fecundability (fecundability ratio = 0.75; 95% CI = 0.59-0.91), as was more than a monthly fish meal in 1991 (fecundability ratio = 0.73; 95% CI = 0.54-0.98). Our findings suggest that maternal but not paternal consumption of contaminated fish may reduce fecundability among couples attempting pregnancy. PMID- 10874546 TI - Serum selenium, serum alpha-tocopherol, and the risk of rheumatoid arthritis. AB - Antioxidant micronutrients have been hypothesized to provide protection against rheumatoid arthritis. We investigated serum selenium and serum alpha-tocopherol for their prediction of subsequent development of rheumatoid arthritis in a case control study nested within a Finnish cohort of 18,709 adult men and women who had neither arthritis nor a history of it at the baseline examination in 1973 1978; by late 1989, 122 had developed rheumatoid arthritis. Of the incident cases, 34 were rheumatoid factor-negative. Three controls per each incident case were individually matched for sex, age, and municipality. Serum selenium and alpha-tocopherol concentrations were measured from stored serum samples collected at baseline. Serum selenium was inversely related to subsequent occurrence of rheumatoid factor-negative but not rheumatoid factor-positive rheumatoid arthritis. The relative risks, adjusted for smoking and serum total cholesterol, for the highest relative to the lowest tertile of serum selenium, were 0.16 [95% confidence interval (CI) = 0.04-0.69] for rheumatoid factor-negative and 0.96 (CI = 0.49-1.90) for rheumatoid factor-positive rheumatoid arthritis. During the first 10 years of follow-up, the relative risk for rheumatoid arthritis for the highest compared with the lowest tertile of serum alpha-tocopherol was 0.44 (CI = 0.19-0.99). No association was found for longer follow-up periods. Low selenium status may be a risk factor for rheumatoid factor-negative rheumatoid arthritis, and low alpha-tocopherol status may be a risk factor for rheumatoid arthritis independently of rheumatoid factor status. PMID- 10874545 TI - The contribution of recently acquired Mycobacterium tuberculosis infection to the New York City tuberculosis epidemic, 1989-1993. AB - Current theory in the molecular epidemiology of tuberculosis holds that tuberculosis cases harboring Mycobacterium tuberculosis strains with a common deoxyribonucleic acid (DNA) fingerprint are the result of recent M. tuberculosis transmission. Here we propose a mathematical approach independent of DNA fingerprinting to estimating the percentage of recent transmissions responsible for current tuberculosis incidence. The "short-term reproductive number" of tuberculosis is defined as the average number of tuberculosis cases developing within 1 year of infection. Multiplying the short-term reproductive number by the number of tuberculosis cases in each year and dividing by the subsequent year's tuberculosis case burden equals the proportion of tuberculosis cases in the subsequent year that are due to recent transmission. We carried out separate calculations for human immunodeficiency virus (HIV)-negative and HIV-positive tuberculosis cases. We applied the model to pulmonary (infectious) tuberculosis cases diagnosed in New York City during 1989-1993, using tuberculosis and AIDS surveillance data. Model-based estimates of the proportion of tuberculosis due to recent transmission were lower than estimates based on DNA fingerprints. Reconciliation of these divergent estimates may require the re-estimation of model parameters from data collected de novo, additional model development, and further advances in DNA fingerprinting methods. PMID- 10874547 TI - The use of electric bed heaters and the risk of clinically recognized spontaneous abortion. AB - We conducted a prospective cohort study to evaluate the relation of spontaneous abortion and electric bed heater use during the first trimester of pregnancy. Compared with non-users, rates of spontaneous abortion were lower for women who used electric bed heaters. The adjusted odds ratio and 95% confidence interval (CI) for the two major devices used, electric blankets (N = 524) and waterbeds (N = 796), were, respectively, 0.8 (95% CI = 0.5-1.1) and 0.9 (95% CI = 0.7-1.2). An increase of risk with increasing intensity (setting-duration combination) of use was not observed. Users of electric blankets at low settings for most of the night (N = 171) had lower risks of spontaneous abortion than non-users (adjusted odds ratio = 0.5; 95% CI = 0.3-1.0). Twenty women who used electric blankets at a high setting for 1 hour or less had an adjusted odds ratio of 3.0 (95% CI = 1.1 8.3), but we found no spontaneous abortions among the few women (N = 13) who used a high setting for 2 or more hours. We found that exposure rankings of the magnetic field time-weighted average and a rate of change metric did not correspond monotonically to the pattern of spontaneous abortion risks and that electric blankets contribute less to overnight time-weighted average magnetic fields than has been thought. PMID- 10874548 TI - Magnesium in drinking water in relation to morbidity and mortality from acute myocardial infarction. AB - We investigated the importance of magnesium and calcium in drinking water in relation to morbidity and mortality from acute myocardial infarction. Cases were men and women 50-74 years of age living in 18 Swedish municipalities who had suffered an acute myocardial infarction some time between October 1, 1994, and June 30, 1996. Controls were randomly selected from the same study base. We interviewed the surviving cases (N = 823) and controls (N = 853), focusing on risk factors for acute myocardial infarction. We collected individual data on drinking water levels of magnesium and calcium. We classified subjects by quartile of water magnesium or calcium levels. The total number of cases was similar in the four quartiles. The risk of death was 7.6% (95% confidence interval = 2.1-13.1) lower in the quartile with high magnesium levels (> or = 8.3 mg/liter). The odds ratio for death from acute myocardial infarction in relation to water magnesium was 0.64 (95% confidence interval = 0.42-0.97) for the highest quartile relative to the three lower ones. Multivariate analyses showed that other risk factors were not important confounders. For calcium, this study was inconclusive. The data suggest that magnesium in drinking water is associated with lower mortality from acute myocardial infarction, but not with the total incidence. PMID- 10874549 TI - Estimating mortality due to cigarette smoking: two methods, same result. AB - We estimated the mortality from various diseases caused by cigarette smoking using two methods and compared the results. In one method, the "Prevent" model is used to simulate the effect on mortality of the prevalence of cigarette smoking derived retrospectively. The other method, suggested by R. Peto et al (Lancet 1992;339:1268-1278), requires data on mortality from lung cancer among people who have never smoked and among smokers, but it does not require data on the prevalence of smoking. In the Prevent model, 33% of deaths among men and 23% of those among women in 1993 from lung cancer, chronic bronchitis, emphysema, ischemic heart disease, and stroke were caused by cigarette smoking. In the method proposed by Peto et al, 35% of deaths among men and 25% of deaths among women from these causes were estimated to be attributable to cigarette smoking. The differences between the two methods are small and appear to be explicable. The Prevent model can be used for more general scenarios of effective health promotion, but it requires more data than the Peto et al method, which can be used only to estimate mortality related to smoking. PMID- 10874550 TI - Prenatal active or passive tobacco smoke exposure and the risk of preterm delivery or low birth weight. AB - We examined the association of exposure to environmental tobacco smoke with birth weight and gestational age in a large, prospective study. We also compared these endpoints between infants of active maternal smokers and those of non-smoking, non-ETS exposed women. Pregnant women were interviewed by telephone during the first trimester, and pregnancy outcome was determined for 99%. Among the 4,454 singleton live births that could be linked to their birth certificate, we confirmed increased risks of low birth weight and small for gestational age with heavier maternal smoking (> 10 cigarettes/day), as well as noting an increased risk for "very preterm" birth (< 35 weeks). These associations were generally stronger among infants of older (> or = 30 years) than those of younger mothers, as well as among non-whites. High environmental tobacco smoke exposure (> or = 7 hours/day in non-smokers) was moderately associated with low birth weight (adjusted odds ratio (AOR) 1.8, 95% confidence limits (95% CL) = 0.82, 4.1), preterm birth (AOR 1.6, 95% CL = 0.87, 2.9), and most strongly with very preterm birth (AOR 2.4, 95% CL = 1.0, 5.3). These associations were generally greater among non-whites than whites. The data support earlier studies suggesting that prenatal environmental tobacco smoke exposure, in addition to maternal smoking, affects infant health. PMID- 10874551 TI - Smoking and alcohol intake as risk factors for bleeding and perforated peptic ulcers: a population-based cohort study. AB - Both the incidence of and mortality from bleeding and perforated peptic ulcers are increasing. We assessed the association between smoking, intake of alcohol (including type of alcoholic beverage), and risk of a complicated peptic ulcer in a population-based study of 26,518 Danish subjects followed up for an average of 13.4 years. There were 214 cases of incident bleeding and 107 cases with perforated ulcers. We estimated relative risks (RRs) for incident bleeding and perforated peptic ulcers using Poisson regression analysis. Smoking more than 15 cigarettes per day compared with never smoking increased the risk of a perforated ulcer more than threefold [RR = 3.5; 95% confidence interval (CI) = 1.7-7.1)]. Drinking more than 42 drinks per week increased the risk of a bleeding ulcer fourfold (RR = 4.4; 95% CI = 2.3-8.3) compared with drinking less than one drink per week. Using the same comparison group, subjects who drank more than 21 drinks per week but no wine were at a higher risk of a bleeding ulcer (RR = 8.8; 95% CI = 2.2-35) than drinkers of the same amount of alcohol, but with more than 25% of their intake as wine (RR = 2.4; 95% CI = 1.0-6.0). PMID- 10874552 TI - Diet and overall survival in a cohort of very elderly people. AB - We conducted a 5-year cohort study among 162 self-sufficient residents in a public home for the elderly in Rome, Italy, to evaluate the association between the consumption of specific food groups and nutrients and overall 5-year survival. We used a validated, semiquantitative food-frequency questionnaire to assess diet at baseline. Individuals consuming citrus fruit at least twice a week had an adjusted risk of dying that was half that of individuals who consumed citrus fruit less than once a week [relative risk (RR) = 0.52; 95% confidence interval (CI) = 0.28-0.95] (with adjustment for gender, age, education, body mass index, smoking status, cognitive function, and chronic diseases). The adjusted RRs of mortality were 0.38 (95% CI = 0.14-1.01) for consumption of milk and yogurt at least three times a week vs less than once a week; 0.21 (95% CI = 0.08 0.35) for moderate consumption of espresso coffee (1-2 cups weekly) vs less than once a week; and 0.35 (95% CI = 0.17-0.69) for > 2 cups a week of espresso coffee vs less than once a week. High levels of intake of ascorbic acid, riboflavin, and linoleic acid were associated with 50-60% decreases in mortality risk. High consumption of meat was associated with a higher risk of mortality (RR = 9.72; 95% CI = 2.68-35.1) among subjects with chronic diseases. Our findings indicate that frequent consumption of citrus fruit, milk, and yogurt; low consumption of meat; and high intake of vitamin C, riboflavin, and linoleic acid are associated with longevity. PMID- 10874553 TI - Myocardial infarction and acute cholecystitis: an application of sequence symmetry analysis. AB - Using a statewide hospital discharge database and a novel epidemiology method, sequence symmetry analysis (Epidemiology. 1996;7:478-84), I examined the relative risk for hospital admission for acute cholecystitis after admission for myocardial infarction. In sequence symmetry analysis, the ratio of the number of subjects in a fixed population who experienced two events in a "causal" vs "noncausal" temporal sequence estimates the incidence rate ratio (IRR). Of 514 patients admitted for both myocardial infarction and acute cholecystitis during a 3-year window period, 295 were admitted for myocardial infarction first and 219 for acute cholecystitis first, yielding a null sequence-adjusted IRR of 1.45 [95% confidence interval (CI) = 1.28-1.64]. A similar analysis for a known relation (myocardial infarction-->congestive heart failure, N = 27,850) showed the expected association [adjusted IRR = 1.92 (95% CI = 1.88-1.95)], whereas an analysis for a relation hypothesized not to be strong (congestive heart failure- >acute cholecystitis, N = 775) showed only a small association [adjusted IRR = 1.16 (95% CI = 1.05-1.28)]. Subgroup analysis revealed time courses that supported each relation as causal. Hospitalization for myocardial infarction may increase the risk for subsequent hospitalization for acute cholecystitis. PMID- 10874555 TI - Second primary cancer after in situ and invasive cervical cancer. AB - The Swedish Family-Cancer Database was used to analyze 9,426 second primary cancers in 117,830 subjects diagnosed with in situ and 17,556 subjects with invasive cervical cancer from the years 1958-1996. We calculated standardized incidence ratios (SIRs) from age- and period-specific rates for all women. SIRs were elevated after both in situ and invasive cervical cancer for cancers of the upper aerodigestive tract, anus, pancreas, lung, other female genitals, and urinary bladder. Anus and other female genitals, known targets of human papilloma virus, showed SIRs exceeding 3.0 and 10 or more within the year of diagnosis of cervical cancer, probably implying the effects of diagnostic intensity or transient faltering of host immunosurveillance. Among the remaining sites, smoking appeared to be the major cause, but for urinary bladder cancer it only explained one-half of the excess; human papilloma virus infection, possibly through immunosuppression, could account for the remaining excess. Although urinary bladder cancer showed a relatively small SIR compared with anal cancer, because it is more common, the number of attributable cases was about equal for the two sites. Invasive cervical cancer showed an SIR of 2.3 after in situ cancer. On follow-up, we also observed increased SIRs at many radiosensitive sites 10 or more years after diagnosis of invasive cervical cancer. PMID- 10874554 TI - Exposure to motor vehicle traffic and allergic sensitization. The Swiss Study on Air Pollution and Lung Diseases in Adults (SAPALDIA) Team. AB - We examined the association between the presence of an allergic sensitization and seasonal allergic diseases or symptoms and the exposure to road traffic in Basel, Switzerland. Traffic counts at the domiciles of subjects ranged from 24 to 32,504 cars per 24 hours, with a median of 1,624. To investigate the relation of road traffic and allergies, we matched the data of the traffic inventory of Basel with those of the 820 participants of the SAPALDIA study (Swiss Study on Air Pollution and Lung Diseases in Adults), ages 18-60 years, who had completed a detailed respiratory health questionnaire and had undergone allergy testing (skin prick tests and serologic examinations). We observed a positive association with a sensitization to pollen that was most pronounced among persons with a duration of residence of at least 10 years. The odds ratios (adjusted for educational level, smoking behavior, number of siblings, age, sex, and family history of atopy) for cars, contrasting four exposure categories with the lowest quartile as referent category, were 1.99 [95% confidence interval (CI) = 0.91-4.38], 2.47 (95% CI = 1.06-5.73), and 2.83 (95% CI = 1.26-6.31). These results suggest that living on busy roads is associated with a higher risk for a sensitization to pollen and could possibly be interpreted as an indication for interactions between pollen and air pollutants. We did not, however, find a similar relation between motor vehicle traffic and hay fever or seasonal allergic symptoms, and we saw no trend that increasing traffic exposure was associated with a rise in sensitization rates to indoor allergens. PMID- 10874556 TI - Validation of the American Cancer Society Cancer Prevention Study II Nutrition Survey Cohort Food Frequency Questionnaire. AB - We assessed the validity and reproducibility of a self-administered 68-item food frequency questionnaire completed in 1992-1993 by approximately 185,000 adults. Four hundred forty-one participants completed four 24-hour dietary recall interviews over a 1-year period and a repeat administration of the food frequency questionnaire. For 20 nutrients and 10 food groups, measured nutrient intakes, but not food group intakes, were consistently lower by food frequency questionnaire than by recall. Energy-adjusted, attenuation-corrected Pearson validity correlations ranged from 0.12 to 0.80, with a median of 0.58. Reproducibility measures were generally high, with a median of 0.69. The food frequency questionnaire performed similarly to food frequency questionnaires used in other cohort studies, indicating similar ability to examine diet-disease relations. PMID- 10874557 TI - A second look at the relation between colorectal adenomas and consumption of foods containing partially hydrogenated oils. AB - The trans fatty acids in partially hydrogenated vegetable oil may cause colorectal neoplasia by interfering with cell membrane function or eicosanoid metabolism. This possibility provided a rationale for looking at the relation between colorectal adenomas and consumption of foods containing partially hydrogenated vegetable oils in 234 cases and 407 controls recruited from referrals for colonoscopy at University of North Carolina Hospitals in Chapel Hill, between 1988 and 1990. Foods containing partially hydrogenated vegetable oils were divided into four groups: sweetened baked goods, chocolate candy, oils and condiments, and french fries and chips. We observed no evidence of increased adenoma prevalence associated with consumption of fries and chips (200+ vs 0 kcals/day: odds ratio (OR) = 0.70; 95% confidence limits (CL) = 0.27, 1.8) or chocolate candy (50+ vs 0 kcals/day: OR = 0.49; 95% CL = 0.23, 1.1). We did, however, find evidence of increased adenoma prevalence associated with consumption of sweetened baked goods (400+ vs < 100 kcals/day: OR = 1.9; 95% CL = 0.95, 3.8) and oils and condiments (200+ vs < 100 kcals/day: OR = 2.4; 95% CL = 1.3, 4.2). PMID- 10874558 TI - Yesterday's science and policy: diet and disease revisited. PMID- 10874559 TI - Moneychangers in the temple. PMID- 10874560 TI - Air pollution and childhood wheezing in a small Italian town. PMID- 10874561 TI - Trends in sexual behavior among German medical students, 1991-1997. PMID- 10874562 TI - Modeling the effects of age at and time since delivery on subsequent risk of cancer. PMID- 10874563 TI - Lung cancer and indoor air pollution arising from Chinese-style cooking among nonsmoking women living in Shanghai, China. PMID- 10874564 TI - Familial occurrence of preeclampsia. PMID- 10874565 TI - Modulation of the oestrogen receptor: a process with distinct susceptible steps. AB - The selective oestrogen receptor modulators (SERMs) constitute a group of substances which are capable of regulating the agonistic/antagonistic profile of the oestrogen receptor in distinct tissues. Their potential utility is considerable since, among the pleiotropic range of effects that oestrogens exert on their target tissues, they may provide a selective profile that better suits each clinical necessity. This review summarizes the principal steps of oestrogen action where modifications have resulted in changes of the effect profile. Three different steps of oestrogen action have been highlighted as being susceptible to modulation: type of ligand, particular species of oestrogen receptor, and particulars at the target tissue. Two main families of SERMs, the triphenylethylene derivatives, with tamoxifen as the main actor, and the benzothiophene derivatives, mainly represented by raloxifene, provide much of the basic and clinical knowledge on the influence of the type of ligand. Two types of oestrogen receptor, alpha and beta, add the second variable susceptible to modulating the response to receptor activation. Finally, the ligand-receptor complex may define particular events in its interaction with DNA, such as binding to promoters other than the oestrogen response element, recruitment of concrete sets of local transcription factors, or other options. PMID- 10874566 TI - A pharmacological review of selective oestrogen receptor modulators. AB - Selective oestrogen receptor modulators (SERMs) are structurally diverse non steroidal compounds that bind to oestrogen receptors and produce oestrogen agonist effects in some tissues and oestrogen antagonist effects in others. SERMs are being evaluated for a number of oestrogen-related diseases, including post menopausal osteoporosis, hormone-dependent cancers, and cardiovascular disease. Several compounds that exhibit a SERM profile are currently available for clinical use, including clomiphene, tamoxifen, and toremifene (which are triphenylethylenes) and raloxifene (a benzothiophene). Clomiphene is used for the induction of ovulation in sub-fertile women attempting pregnancy. Tamoxifen and toremifene are both used to treat breast cancer. Tamoxifen may have beneficial effects on bone mineral density and serum lipids. The effects of toremifene on serum lipids are similar to that of tamoxifen. Both compounds have stimulatory effects on the endometrium. Raloxifene, indicated for the treatment and prevention of post-menopausal osteoporosis, has beneficial effects on bone mineral density and serum lipids, but does not increase the risk of endometrial hyperplasia or endometrial cancer. Recently, raloxifene was shown to reduce the incidence of vertebral fractures in otherwise healthy women with osteoporosis; in the same study, a reduced incidence of breast cancer was also observed. Similar to oestrogens, SERMs increase the incidence of venous thromboembolism. Several newer compounds that exhibit a SERM profile are also in clinical development, including other triphenylethylenes (droloxifene, idoxifene) and benzothiophenes (LY353381.HCl), benzopyrans (EM-800), and naphthalenes (CP-336,156). PMID- 10874567 TI - Steroid hormone receptors: an update. AB - Steroid hormones (SHs) are lipophilic molecules derived from cholesterol and synthesized in the adrenal cortex (glucocorticoids, mineralocorticoids, and adrenal androgens), the testes (testicular androgens, oestrogen), and the ovary and placenta (oestrogens and progestagens or progestins). SHs reach their target cells via the blood, where they are bound to carrier proteins, and because of their lipophilic nature pass the cell membrane by simple diffusion. Within the target cells SHs bind to steroid hormone receptors (SHRs), the key mediators of SH action, which are complexed to chaperones, e.g. heat shock protein 90 (Hsp90), that help other proteins to fold and prevent aggregation. SHRs are intracellular transcription factors that can be activated, among other possibilities, by the specific and high affinity binding of ligand to exert positive or negative effects on the expression of target genes. Binding of agonistic or antagonistic ligands leads to different allosteric changes of SHRs making them competent to exert positive or negative effects on the expression of target genes by different mechanisms. (i) After dissociation of chaperones the liganded SHR-complexes can bind to chromatin organized DNA sequences in the vicinity of target genes, termed hormone response elements (HREs). The HRE-recruited hormone-receptor-complexes are then able to initiate chromatin remodelling and to relay activating or repressing signals to the target genes transcription machinery; (ii) through protein-protein interactions with other sequence-specific transcription factors, SHRs can also regulate the activity of many genes that are switched on, for instance, during stress or an inflammatory response; (iii) the SH response can also be integrated in the intracellular signalling network via cross-talk of SHRs with signal transduction pathways that transmit extracellular signals via membrane receptors and activation of protein kinase cascades to nuclear transcription factors that activate various target genes. By all these different mechanisms SHRs modulate numerous and specific responses in a large variety of cells, whereby their particular effect depends on the physiological, cellular and genetic context. PMID- 10874568 TI - Pure anti-oestrogens. AB - Pure anti-oestrogens are a group of at least five new compounds which are able to antagonize the effects of oestrogen in all tissues and species studied. The mechanism by which the pure anti-oestrogens produce their effects remains in question, but all of them are competitive antagonists of the oestrogen receptors and, moreover, have been proposed to block the shuttling of oestrogen receptors into the cell nucleus. When studied in vitro, these compounds are able to block the oestrogen-stimulated growth of breast cancer cells. In animals, their ability to block the effects of oestrogen on breast, uterus, bone, cardiovascular system and other reproductive-associated tissues has been demonstrated. ICI 182780 has been used in preliminary clinical trials in women with advanced, tamoxifen resistant breast cancer with promising results. Clinical trials with EM-800 are under way to assess the safety and tolerance and to obtain information on its efficacy in patients who have already been treated with tamoxifen. It seems reasonable to assume that pure anti-oestrogens will be a good alternative to tamoxifen in the treatment of breast cancer and also in some non-malignant gynaecological diseases. PMID- 10874569 TI - The endometrial effects of SERMs. AB - The ideal selective oestrogen receptor modulator (SERM) would retain an oestrogen like effect on the bones, the heart and cardiovascular apparatus, and the central nervous system, while acting as an anti-oestrogen on the breast and the genital tract. It seems, however, that such a compound is not available for clinical use yet. The uterine tissue, and particularly the endometrium, defines an area of special interest in the SERM action, since endometrial hyperplasia and cancer has been linked to agonistic oestrogen effects. Additionally, tamoxifen, the SERM which accumulates most of the clinical experience, has been associated with stimulatory effects on endometrium, including the development of cancer. In contrast, the more recent benzothiophenes, led by raloxifene, seem to operate as endometrial antagonists, thus providing an interesting alternative for clinical use. This review analyses the endometrial action of tamoxifen, including the information gathered from laboratory models, the observed endometrial effects in women using tamoxifen, and the epidemiological and molecular data which link the use of tamoxifen with endometrial cancer. A parallel examination of the raloxifene data presents the available experimental and clinical information, suggesting the endometrial neutrality of this compound. PMID- 10874570 TI - Skeletal effects of selective oestrogen receptor modulators (SERMs). AB - Women suffer a higher incidence of osteoporosis than men, in part due to oestrogen deficiency after menopause. In fact, the administration of oestrogen to post-menopausal women is associated with a decrease of bone resorption. Tamoxifen is a widely used selective oestrogen receptor modulator in women with breast cancer, which has been shown an agonistic profile in bone. However, tamoxifen seems less effective than oestradiol as an anti-resorptive agent and has no effect when the endogenous production of oestrogen is normal. Additionally, tamoxifen exhibits agonistic activity on the endometrium and has been suggested an oncogenic potential on that tissue. Raloxifene, a selective oestrogen receptor modulator from the benzothiophene family, behaves also as an agonist on the bone in both laboratory and clinical studies. Ongoing clinical trials confirm a protective effect of raloxifene similar to oestrogens. The Multiple Outcome of Raloxifene Evaluation (MORE) study is a prospective, randomized trial which, in a recent 2 year interim analysis, has shown that women suffering from osteoporosis receiving raloxifene had 42% fewer vertebral fractures than women receiving a placebo. PMID- 10874571 TI - Cochrane review: post-operative procedures for improving fertility following pelvic reproductive surgery. AB - The objectives of the study was to determine the effectiveness of post-operative procedures following female pelvic reproductive surgery. A systematic review employing the principles of the Cochrane Menstrual Disorders and Subfertility Group was used. Five randomized controlled trials were included. Participants were women undergoing pelvic reproductive surgery; interventions were any post operative procedure designed to improve fertility; outcomes were pregnancy, live birth, ectopic pregnancy and miscarriage rates and the rates of tubal patency and procedure-related complications. Summary statistics were expressed as odds ratios. The results showed that the odds of pregnancy, live birth, ectopic pregnancy and miscarriage were not significantly altered by post-operative hydrotubation nor second-look laparoscopy with adhesiolysis. Whether hydrotubation was early or late and whether hydrotubation fluid contained steroid or not had no significant impact on the odds of pregnancy, live birth, ectopic pregnancy or miscarriage. The odds of pregnancy and live birth were significantly increased and infective complications significantly decreased by hydrotubation with fluid containing antibiotic compared with hydrotubation with fluid containing no antibiotic, in late hydrotubation following tubal stent removal 6 weeks after tubal surgery. The odds of at least one patent Fallopian tube were significantly increased with late hydrotubation following tubal stent removal compared with early hydrotubation in women who had no tubal stenting, but this intervention had no significant impact on the odds of pregnancy, live birth, ectopic pregnancy or miscarriage. In conclusion, there is insufficient evidence to support the routine practice of hydrotubation or second-look laparoscopy following female pelvic reproductive surgery. The studies on which this conclusion is based were either poor quality or underpowered. Post-operative hydrotubation with fluid containing antibiotic following tubal surgery may offer benefit over hydrotubation fluid without antibiotic. A randomized controlled trial of post-operative hydrotubation with antibiotic-containing fluid versus no hydrotubation for improving fertility following tubal surgery is justified. PMID- 10874572 TI - Fluid compartments of the embryonic environment. AB - The exocoelomic cavity was probably the last remaining physiological body fluid cavity to be explored in the human embryo. Its unique anatomical position has enabled us to study the protein metabolism of the early placenta and secondary yolk sac and to explore materno-embryonic transfer pathways. The exocoelomic cavity forms inside the extraembryonic mesoderm alongside the placental chorionic plate and is now believed to be an important transfer interface and a reservoir of nutrients for the embryo. Maternal or placental proteins filtered in the extraembryonic coelomic cavity are probably absorbed by the secondary yolk sac which is directly connected with the primitive digestive system throughout embryonic development. Protein electrophoresis has shown that the coelomic fluid results from an ultrafiltrate of maternal serum with the addition of specific placental and secondary yolk sac bioproducts demonstrating that the exocoelomic cavity is a physiological liquid extension of the early placenta. The selective sampling of fluid from the exocoelomic cavity has also offered a novel approach to the study of drug and toxin transfer across the early human placenta and as a unique tool to explore embryonic physiology in vivo. Further investigation should include a comparison between the coelomic fluid values of a molecule and its quantifiable presence in decidual, placental and fetal tissues. PMID- 10874573 TI - Oocyte--granulosa cell interactions. AB - In the past, different protocols of ovulation induction, aimed to overcome problems of anovulatory infertility in humans, have been developed during IVF programmes. However, administration of exogenous hormones may cause severe health problems, e.g. ovarian hyperstimulation syndrome. To overcome this problem an attractive alternative is to develop in-vitro systems that allow follicle and oocyte growth and maturation. This paper reviews the current status of research on oocyte-granulosa cell interactions and on the autocrine and paracrine factors involved in follicle development. The ovarian follicle is a morphological and functional unit in which the somatic and germ cell components are intimately associated and interdependent. The co-ordinate development of follicle and oocyte leads to a number of modifications in the growing oocyte necessary for the acquisition of competence to mature correctly and to undergo fertilization and embryo development. The search for the optimal culture conditions and the correct balance of hormones necessary to obtain a fertilizable oocyte in vitro is extremely important for clinical and agricultural applications. PMID- 10874575 TI - The impact of the factor V Leiden mutation on pregnancy. AB - A resistance to the anticoagulant activity of activated protein C (APC), most frequently due to a point mutation in the Factor V gene (the Leiden mutation), represents the most common genetic cause of thrombophilia. The Leiden mutation has been significantly related to pregnancy complications associated with hypercoagulation, e.g. deep vein thrombosis during pregnancy (8-fold increased risk), pre-eclampsia (prevalence of the mutation up to 26%), placental infarction extending to > 10% of the placenta (10-fold increased risk), abruptio placentae (prevalence of the mutation up to 29.6%), and second- and third-trimester pregnancy failure (prevalence of the mutation up to 31.3%). An association of the maternal mutation with recurrent first-trimester miscarriage does not emerge from the literature, although fetal mutation (frequency higher than twice compared with that of the general population) has been related to early spontaneous miscarriage. Although some evidence suggests an association between APC resistance and intrauterine growth retardation, no significant relationship emerges currently from the literature. Screening for the Leiden mutation would seem advisable in women with previous pregnancy complications amongst those associated with APC resistance. Carriers of the mutation should be given appropriate counselling. The screening of asymptomatic women is not recommended at present. PMID- 10874574 TI - Leptin and reproduction. AB - Leptin, the product of the ob gene, is a small peptide molecule synthesized by white adipocytes with an important role in the regulation of body fat and food intake. Leptin and leptin receptor mRNA were first detected in the brain and hypothalamus but now their ubiquitous presence has been demonstrated. Leptin receptor signal transduction involves the activation of signal transducer and activator of transcription (STAT)-3, a member of the transcription family of proteins. Leptin is regulated by hormones and cytokines, interleukin-1, tumour necrosis factor-alpha and transforming growth factor-beta, linking this molecule with the inflammatory response. In addition, emerging evidence has demonstrated that this molecule is related to reproductive function. This small protein is present in the ovary and decidua, in mature oocytes and during embryonic development and trophoblast invasion. Animal models have demonstrated that leptin deficient ob/ob mice are sterile; however, fertility can be restored by exogenous leptin. In addition, embryos implanted in STAT-3-deficient mice degenerate rapidly and are the target disruption of STAT-3-provoked embryonic lethality. Leptin acts as a novel placental hormone participating in the control of fetal growth and development. Leptin could be a modulator for invasive features of cytotrophoblast cells. We postulate that leptin may have an autocrine/paracrine role in human implantation and placentation. PMID- 10874576 TI - Cellular and molecular biology of capacitation and acrosome reaction in spermatozoa. AB - A comparative account is given of advances in cellular and molecular biology of capacitation and acrosome reaction in spermatozoa by comparing and contrasting their biochemical and physiological changes in response to various factors in vivo and in vitro. It can now be stated that phenomena of sperm capacitation and acrosome reaction are endogenous molecular events occurring at the membrane level which can be modulated by external environmental factors. The molecular mechanisms and the signal transduction pathways mediating the process of capacitation and acrosome reaction are only partially defined and appear to involve modification of intracellular Ca2+ and other ions, lipid transfer, and phospholipid remodeling in the sperm plasma membrane as well as changes in protein phosphorylation. Evidences for the involvement of cAMP-dependent kinase pathway in the acrosome reaction are discussed. The mediation of one or more external signals by the sperm plasma membrane appears to activate this pathway after or simultaneously with the influx of Ca2+. Concurrent with or following entry of Ca2+, adenylate cyclase is activated, leading to increased concentrations of cAMP-activation of cAMP-dependent kinase and protein phosphorylation; the identity of such proteins and their role in the acrosome reaction must be determined. The roles of biological effectors of the acrosome reaction, such as ZP3 and follicular fluid are still to be defined at the molecular level. The gaps in our knowledge about the cellular and molecular aspects of capacitation and acrosome reaction are emphasized. PMID- 10874577 TI - Growth factors and epithelial-stromal interactions in prostate cancer development. AB - Epithelial-stromal interactions are important not only in growth, development, and functional cytodifferentiation of the prostate but also in derangements of prostate gland such as BPH and prostate carcinoma. This chapter explores the roles of epithelium and stroma during this delicate process and highlights the role and mutual influence of each on the other. It also examines the importance of ECM in mediating the effects of androgens and drawn attention to estrogen and genetic factors in the process. During this process of epithelial-stromal interaction, growth factors play a central role in mediating the interactions. This chapter focuses on the role of several growth factors including epidermal growth factor, fibroblast growth factor, transforming growth factor alpha, transforming growth factor beta, insulin-like growth factor-1, vascular endothelial growth factor, nerve growth factor, platelet-derived growth factor, and hepatocyte growth factor. This chapter emphasizes the importance of epithelial-stromal interactions in tumorigenesis and highlights the switch of paracrine to autocrine mode during the process of carcinogenesis. PMID- 10874578 TI - PCR-detected genome polymorphism in malignant cell growth. AB - In this chapter, we analyze the problem of genetic polymorphism in tumorigenesis, which determines basic capacities of tumors. The study of genome polymorphism with modified PCR methods allows the detection of various forms of polymorphism in tumor cells. This method has made it possible to determine association of DNA polymorphism with conditions of oncogenes, antioncogenes, and genes of apoptosis and with their allelic states. A special type of nonspecific DNA polymorphism that resulted from an increase in the mutation number in the cancer cell genome was discovered. This phenomenon was called the microsatellite mutator phenotype. Because the type of DNA polymorphism correlates with various biological capacities of malignant tumors and has an important prognostic significance, the analysis of DNA polymorphism in benign and malignant tumors of different histogenesis will play an important role both in theoretical studies of cancer and in oncological practice. A modified B1-PCR was used to study the genome polymorphism in the mouse tumor cells. The gain of the band 470 bp and the loss of the band 600 bp were revealed in the hepatoma cell line MH-22a as compared with liver cells of C3HA mice. The differentiation of teratocarcinoma EC F9 cells to endoderm-like cells was not accompanied by any changes in the B1-AF DNA fingerprint. PMID- 10874579 TI - Cellulose microfibrils in plants: biosynthesis, deposition, and integration into the cell wall. AB - Cellulose occurs in all higher plants and some algae, fungi, bacteria, and animals. It forms microfibrils containing the crystalline allomorphs, cellulose I alpha and I beta. Cellulose molecules are 500-15,000 glucose units long. What controls molecular size is unknown. Microfibrils are elongated by particle rosettes in the plasma membrane (cellulose synthase complexes). The precursor, UDP-glucose, may be generated from sucrose at the site of synthesis. The biosynthetic mechanism may involve lipid-linked intermediates. Cellulose synthase has been purified from bacteria, but not from plants. In plants, disrupted cellulose synthase may form callose. Cellulose synthase genes have been isolated from bacteria and plants. Cellulose-deficient mutants have been characterised. The deduced amino acid sequence suggests possible catalytic mechanisms. It is not known whether synthesis occurs at the reducing or nonreducing end. Endoglucanase may play a role in synthesis. Nascent cellulose molecules associate by Van der Waals and hydrogen bonds to form microfibrils. Cortical microtubules control microfibril orientation, thus determining the direction of cell growth. Self assembly mechanisms may operate. Microfibril integration into the wall occurs by interactions with matrix polymers during microfibril formation. PMID- 10874580 TI - Cell biology of peroxisomes and their characteristics in aquatic organisms. AB - The general characteristics of peroxisomes in different organisms, including aquatic organisms such as fish, crustaceans, and mollusks, are reviewed, with special emphasis on different aspects of the organelle biogenesis and mechanistic aspects of peroxisome proliferation. Peroxisome proliferation and peroxisomal enzyme inductions elicited by xenobiotics or physiological conditions have become useful tools to study the mechanisms of peroxisome biogenesis. During peroxisome proliferation, the induction of peroxisomal proteins is heterogeneous, enzymes that show increased activity being involved in different aspects of lipid homeostasis. The process of peroxisome biogenesis is coordinately triggered by a whole array of structurally dissimilar compounds known as peroxisome proliferators, and investigating the effect of some of these compounds that commonly appear as pollutants in the environment on the peroxisomes of aquatic animals inhabiting marine and estuarine habitats seems interesting. It is also important to determine whether peroxisome proliferation in these animals is a phenomenon that might occur under normal physiological or season-related conditions and plays a metabolic or functional role. This would help set the basis for understanding the process of peroxisome biogenesis in aquatic animals. PMID- 10874581 TI - Cellular responses to vasectomy. AB - A number of cell populations in the reproductive tract show a response to vasectomy. Some cell types show similar responses in man and all laboratory species, whereas others show marked species variations. This chapter describes these effects in a broadly chronological order and, in a general way, considers changes close to the site of vasectomy first and the longer term effects on the testis itself later. Following vasectomy, epididymal distension and sperm granuloma formation result from raised intraluminal pressure. The sperm granuloma is a dynamic structure and a site of much spermatozoal phagocytosis by its macrophage population. In many species, spermatozoa in the obstructed ducts are destroyed by intraluminal macrophages, and degradation products, rather than whole sperm, are absorbed by the epididymal epithelium. Humoral immunity against spermatozoal antigens following vasectomy is well established and there is evidence of modest T-lymphocyte activity. The role of lymphocytes in the reproductive tract epithelium and interstitium following vasectomy is poorly defined. In laboratory animals, there is evidence that pressure-mediated damage to the seminiferous epithelium can follow sperm granuloma formation and obstruction in the epididymal head. However, the contribution of lymphocytes and antisperm antibodies to testicular damage after vasectomy is far from clear. A number of studies have suggested that testicular changes may follow vasectomy in man but their validity and mechanism of occurrence require further study. PMID- 10874582 TI - Dopamine D4 receptors. AB - Initial investigations on dopamine D4 receptors generated much interest in the role of this receptor in schizophrenia and other aspects of human behavior, as well as new opportunities for novel therapeutics. However, attempts to treat patients suffering from schizophrenia with dopamine D4 agents have failed to yield satisfactory results so far. An examination of the dopamine D4 literature shows that contrasting and conflicting data seemed to be the norm in this field of research. This paper reviews the literature on the dopamine D4 receptor and discusses many of the associated methodological problems that might have contributed to the paradoxical findings. PMID- 10874583 TI - Molecular basis of spatio-temporal dynamics in inositol 1,4,5-trisphosphate mediated Ca2+ signalling. AB - Inositol 1,4,5-trisphosphate (IP3)-mediated Ca2+ signalling regulates many important cell functions, and the spatio-temporal dynamics of the Ca2+ signalling is a crucial factor for its versatility. The molecular mechanisms that control Ca2+ signalling are now being investigated, and I here describe the subtypes of IP3 receptors that have distinct functional properties and contribute to the diversity of Ca2+ signalling patterns. I also discuss the spatio-temporal dynamics of intracellular IP3 concentration, describing recent methodological advances in monitoring intracellular IP3 concentration. These findings highlight the potential importance of the spatio-temporal information of any signalling molecule. PMID- 10874585 TI - Anti-type I allergic mechanisms of mao-bushi-saishin-to in mice. AB - We investigated the anti-allergic effect of mao-bushi-saishin-to (MBS) on the type I allergy model in mice. When MBS was administered orally at a dose of 0.5 or 1.0 g/kg, edema of the footpad, the amount of plasma IgE and the ratio of eosinophilic leukocytes in peritoneal exudate cells were all dose-relatedly suppressed. Moreover to investigate the anti-type I allergic mechanisms of MBS, enzyme-linked immunosorbent assay was performed to determine the interleukin (IL) 4, IL-5 and interferon (IFN)-gamma production from splenocytes that were stimulated by pokeweed mitogen for 48 h. In addition, we assayed IgE production from splenic B cells stimulated with the lipopolysaccharide and IL-4 for 7 days. MBS inhibited the IL-4 and IFN-gamma production, but IL-5 and IgE production were not affected. Thus possibly, the inhibition of IL-4 production may partially be involved in the expression of the anti-type I allergic effects of MBS. PMID- 10874584 TI - Studies on the mechanism of action of the gastric H+,K(+)-ATPase inhibitor SPI 447. AB - 3-Amino-5-methyl-2(2-methyl-3-thienyl)- imidazo[1,2-a]thieno[3,2-c]pyridine, SPI 447, is a potent gastric H+,K(+)-ATPase inhibitor, but a detailed mechanism of the inhibition is unknown. This study was designed to investigate the mechanism by which SPI-447 inhibits gastric H+,K(+)-ATPase. For this purpose, the inhibitory action of SPI-447 on gastric H+,K(+)-ATPase from porcine gastric mucosa was compared with that of omeprazole (an irreversible inhibitor) and SCH28080 (a reversible inhibitor). All compounds produced dose-dependent inhibition of gastric H+,K(+)-ATPase, and the inhibitory intensities were increased under acidic conditions. The anti-H+,K(+)-ATPase actions of SPI-447 and SCH28080 were attenuated by dilution, but not influenced by glutathione pretreatment. In contrast, that of omeprazole was not influenced by dilution, but was suppressed by glutathione pretreatment. KCl addition reversed the inhibition of H+,K(+)-ATPase-mediated H(+)-transport by SPI-447 and SCH28080, but had no effect on that by omeprazole. The anti-gastric H+,K(+)-ATPase action of SPI-447 was additive with that of SCH28080. SPI-447 and SCH28080 had no effect on Na+,K(+)-ATPase activity. These findings indicated that the inhibitory mechanism of SPI-447 on gastric H+,K(+)-ATPase was similar to that of SCH28080, but different from that of omeprazole; i.e., 1) reversible, 2) SH-group independent, 3) K(+)-competitive, and 4) highly specific against gastric H+,K(+)-ATPase. PMID- 10874586 TI - Inhibition by naloxone of promoter activity of the neurofilament gene in SK-N-SH cells. AB - Chronic administration of morphine is known to decrease the levels of neurofilaments (NFs) in the ventral tegmental area. We ligated a promoter region of the mouse 68-KDa neurofilament (NF-68) gene to the pGL3-enhancer vector containing a luciferase gene, transfected it into SK-N-SH cells and then analyzed transcriptional activity in the cells treated with agonists or antagonists of opiate receptors. The activity of the NF-68 promoter was suppressed by naloxone about 55% at 10(-5) M and 30% at 10(-7) M at 48 h, but suppressed not by morphine. Naltrexone at 10(-5) M suppressed the promoter activity about 20%, but levallorphan, DAMGO, DPDPE and U50488 did not. The inhibition by naloxone was dose-dependent and not reversed by morphine. The inhibitory effect of naloxone was not observed in N18TG-2 cells and PC12 cells. Experiments with various deletion mutants revealed that a region responsible for naloxone suppression spans from -328 to -101 in the gene. These results suggest that naloxone has the ability to suppress transcriptional activity in some neurons. PMID- 10874587 TI - L-DOPA cyclohexyl ester is a novel potent and relatively stable competitive antagonist against L-DOPA among several L-DOPA ester compounds. AB - We explored L-DOPA esters with chemically bulky structures to find a potent stable competitive antagonist against L-DOPA, compared to DOPA methyl ester (DOPA ME). In anesthetized rats, DOPA cyclohexyl ester (DOPA CHE), DOPA cyclopentyl ester (DOPA CPE) and DOPA cyclopentyldimethyl ester (DOPA CPDME) at 1 microgram microinjected into depressor sites of the nucleus tractus solitarii elicited or tended to elicit more marked antagonism against depressor responses to 60 ng L DOPA, compared to DOPA ME. At 100 ng, DOPA CHE elicited the most potent antagonism. At 1 microgram, duration of the antagonistic activity of DOPA CHE was approximately three times longer than that of DOPA ME. During microdialysis of the nucleus accumbens, conversion from DOPA CHE at 1 microM perfused via probes to extracellular L-DOPA was the lowest among these compounds and less than one half of that from DOPA ME. Binding studies showed that the recognition site for L DOPA differs from ionotropic glutamatergic, dopaminergic D1 and D2 receptors. We recently found that L-DOPA evoked by transient ischemia may act as a DOPA CHE sensitive causal factor for glutamate release and resultant neuronal cell death. DOPA CHE is the most potent, relatively stable competitive antagonist against L DOPA and is a useful mother compound to develop neuroprotective drugs. PMID- 10874588 TI - Participation of GABAergic and histaminergic systems in inhibiting amygdaloid kindled seizures. AB - The effects of GABAmimetic drugs on inhibition of amygdaloid kindled seizures induced by clobenpropit were investigated to clarify the relationship between histaminergic and GABAergic systems in seizures. I.p. injection of clobenpropit caused dose-dependent inhibition of amygdaloid kindled seizures. GABAmimetic drugs such as diazepam, sodium valproate and muscimol also inhibited amygdaloid kindled seizures in a dose-dependent manner. Diazepam at doses of 0.2 and 0.5 mg/kg, which showed no significant effect on amygdaloid kindled seizures when used separately, significantly potentiated the effect of clobenpropit. Similar findings were observed with sodium valproate and muscimol at doses of 100 mg/kg and 5 ng, respectively, although neither showed any significant effects when administered separately. Bicuculline caused significant antagonism of the inhibition of amygdaloid kindled seizures induced by clobenpropit, while the effect of diazepam was not antagonized by diphenhydramine. These results suggested that inhibition of amygdaloid kindled seizures induced by histamine is closely associated with the actions of GABA. PMID- 10874589 TI - Dietary fructooligosaccharides prevent a reduction of cortical and trabecular bone following total gastrectomy in rats. AB - Fructooligosaccharides (FOS) have been shown to stimulate the absorption of several minerals in the intestine. In the present study, the effects of FOS on osteopenia induced by total gastrectomy were examined. Twenty eight male Sprague Dawley rats were divided into 2 groups: sham-operated (SH) and gastrectomized (GX). After a one-week adaptation period following surgery, the rats were fed synthetic diets with or without 7.5% FOS for 5 weeks. The right femur was then examined by soft X-ray, and the bone mineral density (BMD) was measured. Based on the soft X-ray findings, both cancellous and cortical bone were markedly decreased in GX rats, but not in GX + FOS rats. GX rats showed a 30% lower BMD in the metaphysis and a 20% lower BMD in the diaphysis, compared with SH rats (P < 0.01). As assessed by morphometry, significant decreases were observed in cortical bone in the diaphysis and trabecular bone in the distal metaphysis (P < 0.01). On the other hand, dietary FOS completely prevented these changes following gastrectomy. These findings indicate that dietary FOS might contribute to the prevention of bone diseases following gastrectomy. PMID- 10874590 TI - Stimulation of noradrenaline release by T-588, a cognitive enhancer, in PC12 cells. AB - Previously, we reported that (R)-(-)-1-(benzo[b]thiophen-5-yl)-2-[2-(N,N- diethylamino)ethoxy] ethanol hydrochloride (T-588), a novel putative cognitive enhancer, stimulated noradrenaline (NA) release from rat cerebral cortical slices. In this study, we investigated the effects of T-588 compared to other secretagogues on NA release from PC12 cells. Addition of as little as 10 microM T 588 stimulated [3H]NA release in a dose-dependent and an extracellular Ca(2+) independent manner from PC12 cells. Ten micromolar ionomycin-, 300 microM adenosine-5'-O-(gamma-thiotriphosphate)- and 10 microM forskolin-induced extracellular Ca(2+)-dependent [3H]-NA release was further enhanced by 30 microM T-588. Cytosolic synaptophysin and 25-kDa synaptosome-associated protein immunoreactivity was increased by addition of T-588 in a dose-dependent manner. Interestingly, increases in synaptic vesicle-related proteins triggered by T-588 had a 4-min lag time and were completely dependent on extracellular CaCl2. These findings suggest that T-588 stimulates NA release from PC12 cells in a Ca(2+) independent manner. T-588 also induced the translocation of synaptic vesicles in a Ca(2+)-dependent manner. PMID- 10874591 TI - Effects of neuropeptide Y on double-peaked constrictor responses to periarterial nerve stimulation in isolated, perfused canine splenic arteries. AB - The periarterial electrical nerve stimulation readily induced a double-peaked vasoconstriction in the isolated, perfused canine splenic artery. P2X Purinoceptors have previously been shown to be involved mainly in the 1st-phase response and alpha 1-adrenoceptors, mostly in the 2nd-one. The dose used of neuropeptide Y (NPY) (0.01-0.1 microM) given into the preparation caused a slight but insignificant vasoconstriction. The treatment with NPY at concentrations of 0.01-0.1 microM produced a parallel inhibition on the 1st- and 2nd-phase responses following nerve stimulation at the frequencies used (1-10 Hz) in a dose dependent manner. The vasoconstrictor responses to administered ATP (0.01-1 mumol) or noradrenaline (0.03-3 nmol) were slightly but not significantly potentiated by 0.1 microM NPY. The results indicate that NPY predominantly exerts a prejunctionally inhibitory modulation on the purinergic and adrenergic transmission in peripheral sympathetic nerves innervating the canine splenic artery. PMID- 10874592 TI - Perivascular purinergic nerve-induced vasoconstrictions in canine isolated splenic arteries. AB - We tried to induce selective perivascular purinergic nerve stimulation in isolated canine splenic arterial preparations, using the cannula insertion method. Under the conditions of periarterial electrical stimulation (ES), i.e., trains of 1, 3 and 10 pulses, 1-ms pulse duration and 10-V amplitude at 1 Hz, monophasic vasoconstriction was consistently induced. The ES-induced vasoconstriction was not influenced by prazosin in doses that completely inhibited noradrenaline-induced vasoconstrictions, but it was suppressed by alpha,beta-methylene ATP, a P2X purinoceptor desensitizer. Thus, it is indicated that a selective purinergic transmitter release is readily obtained in the isolated splenic arterial preparation. PMID- 10874593 TI - Characterization of protease-activated receptors in rat peritoneal mast cells. AB - Activation of protease-activated receptor (PAR)-1 or PAR-2 elicits inflammation most probably via mast cell degranulation in vivo. The present study aimed at characterizing PARs in rat peritoneal mast cells (PMC). Messenger RNA for PAR-1, but not for PAR-2, was detected in PMC. Thrombin, the PAR-1 agonist SFLLR-NH2 or the PAR-2 agonist SLIGRL-NH2 failed to induce histamine release from PMC. Surprisingly, the PAR-2-inactive control peptide LSIGRL-NH2 triggered histamine release from PMC. Thus, PAR-1, but not PAR-2, are expressed in PMC, whereas neither PAR-1 nor PAR-2 are considered to be involved in degranulation of PMC. LSIGRL-NH2 does not appear to be appropriate as a control peptide for PAR-2 in inflammation studies. PMID- 10874594 TI - Peroxynitrite-generating species: good candidate oxidants in aqueous extracts of cigarette smoke. AB - Cigarette smoking is a well-known risk factor for atherosclerosis, but the mechanism of the adverse biological effect of smoking remains to be established. Cigarette smoke contains high concentrations of free radicals and oxidants. We show here that cigarette smoke extracts (CSE), prepared by bubbling the gas phase of smoke into phosphate-buffered saline, could convert tyrosine to 3 nitrotyrosine. The tyrosine nitration terminated 6 h after incubating tyrosine with CSE at 37 degrees C. These results indicate that the active oxidants in CSE are peroxynitrite-generating species like 3-morpholinosydnonimine (SIN-1), suggesting that they modify plasma lipoproteins and contribute to the pathogenesis of atherosclerosis. PMID- 10874595 TI - Phenytoin and its metabolite, 5-(4-hydroxyphenyl)-5-phenylhydantoin, show bone resorption in cultured neonatal mouse calvaria. AB - The effects of phenytoin and its major metabolite, 5-(4-hydroxyphenyl)-5 phenylhydantoin (HPPH), on bone resorption of neonatal mouse calvaria were examined in vitro. Both phenytoin and HPPH induced significant bone resorption as compared to the controls after 72 h in culture. This effect may be the cause of phenytoin-induced bone loss in vivo. PMID- 10874596 TI - Roles of macrophage-colony stimulating factor and osteoclast differentiation factor in osteoclastogenesis. AB - We have isolated osteoclast precursors (OCPs) from cocultures of mouse calvarial cells and bone marrow cells without adding any osteotropic factors. OCPs expressed Mac-1, Mac-2, and Gr-1 antigens but not osteoclast markers such as tartrate-resistant acid phosphatase (TRAP) and calcitonin receptors, and they differentiated into TRAP-positive cells within 48 h on a fixed calvarial cell layer pretreated with osteotropic factors such as 1 alpha, 25-dihydroxyvitamin D3. In the present study, we investigated the regulatory mechanisms of OCP formation from hemopoietic cells and TRAP-positive cell formation from OCPs. Calvarial osteoblasts obtained from macrophage-colony stimulating factor (M-CSF) deficient op/op mice failed to support OCP formation or the differentiation of OCPs into TRAP-positive cells. Both OCP formation and TRAP-positive cell formation supported by osteoblasts were completely inhibited by osteoclastogenesis inhibitory factor (OCIF, also called OPG), which is a decoy receptor of osteoclast differentiation factor (ODF; also called TRANCE, RANKL, and OPGL). When bone marrow cells were cultured for 4 days with soluble ODF (sODF/sRANKL) together with M-CSF, OCPs were formed even in the absence of osteoblasts. When OCPs were treated with sODF/sRANKL and M-CSF in the absence of osteoblasts, they differentiated into TRAP-positive cells within 48 h even in the presence of hydroxyurea. Northern blotting analysis revealed that osteoblasts constitutively expressed a certain level of ODF/RANKL mRNA. These results indicated that M-CSF and sODF/sRANKL produced by osteoblasts are two essential factors for both OCP formation and TRAP-positive osteoclast formation. PMID- 10874597 TI - Spatiotemporal change of rat collagenase (MMP-13) mRNA expression in the development of the rat femoral neck. AB - The interepiphyseal region between the greater trochanter and the capital femoral epiphysis and the medioproximal portion of the femoral neck exhibit extensive morphological changes during the first 4 weeks after birth in rats. Previous reports show that matrix metalloproteinase-13 (MMP-13, rat collagenase) mRNA is expressed in bone and cartilage during embryonal development and fracture healing. We examined MMP-13 mRNA expression and compared it with the distribution of osteopontin and osteocalcine mRNA in the femoral neck. Moreover, we examined histomorphometric analysis in the femoral neck where the morphology changes rapidly. Histomorphometric analysis of the 4-week-old rat femoral neck showed a high rate of bone formation and resorption in the region where shape changed rapidly. Osteopontin mRNA was expressed diffusely along the endosteum. In contrast, MMP-13 mRNA expression was restricted to the medial endosteal portion near the cartilage-bone interface of the femoral neck in 15- and 28-day-old rats and in the deepest endosteal interepiphyseal region of 15-day-old rats. MMP-13 mRNA-expressing osteoblastic cells were also expressing osteopontin but not osteocalcin mRNA. MMP-13 mRNA-expressing cells differ from tartrate-resistant acid phosphatase (TRAP)-positive cells, and MMP-13 mRNA-positive cells are located adjacent to TRAP-positive cells. The results of the site- and cell specific expression of MMP-13, taken together with its enzymatic property, suggest that MMP-13 plays an important role in morphological changes in the rat femur, at least during the third and fourth week after birth, and that MMP-13 itself is involved in the interaction between osteoblastic and TRAP-positive cells. PMID- 10874598 TI - Effect of salmon calcitonin on experimental osteoporosis induced by ovariectomy and low-calcium diet in the rat. AB - To investigate the action and effect of calcitonin on osteoporosis, this study was performed in osteoporotic rats that had been ovariectomized and maintained on a low-calcium diet. Significant bone loss was noted in ovariectomized rats compared with those that underwent sham surgeries, and histological findings proved bone turnover to be increased. In comparison with this osteoporotic rat group, those given calcitonin showed less bone loss; the reduction in bone loss was obvious in the vertebral body, and rats given a high dose of calcitonin over a long duration showed even less bone loss, but this was hardly seen in the proximal tibial metaphysis. Histological findings proved that calcitonin inhibited the bone resorption that was stimulated by the ovariectomy. Values for osteoblast surfaces were relatively higher while those of reversal surfaces were lower. The results confirm that salmon calcitonin has an inhibitory action on bone resorption as well as being effective in maintaining and stimulating bone formation in vivo. These effects of calcitonin prevented progress of the osteoporosis that had been induced by ovariectomy, and the effects appeared differently in each region. PMID- 10874599 TI - Morphologically extracted trabecular skeleton superimposed upon digital radiograph structure. AB - The purpose of this study was to employ a morphological filter to digital X-ray images to extract the morphology of trabecular structures in a clearly understandable visual format. This study compares the trabecular skeleton extracted by a morphological filter to the original digital radiographic image by superimposing the images. A morphological filter (a combination of a single structuring element and a skeleton operation) based on a mathematical morphology theory was used to extract the skeletal pattern of trabecular bone from a digital X-ray image of a human femoral neck. Subset images with different operation numbers (n = 1, n = 2) were obtained, and then each image was superimposed on the original digital radiographic image using the superimpose function of a workstation. The extracted trabecular skeleton pattern was fairly consistent with the trabecular structure seen on the digital image according to the opinion of seven dentomaxillofacial radiologists. In their opinion, the majority of the structural elements were reproduced on the extracted skeleton. However, accurate skeleton elements were not extracted in the region of dense trabecular structure. The morphological filter was able to extract a large portion of the bone trabecular structure as a binary skeletal pattern image from trabecular bone on digital X-ray image, but more work is needed to improve the assessment of dense trabeculae. PMID- 10874600 TI - Peripheral computed tomography (pQCT) detected short-term effect of AAACa (heated oyster shell with heated algal ingredient HAI): a double-blind comparison with CaCO3 and placebo. AB - Trabecular bone density at the distal radius and cortical bone density at the midradius were measured in four randomized groups of women before and after 4 months administration of AAACa, oyster shell heated under reduced pressure with addition of heated algal ingredient (HAI) (group A); AACa, the same preparation without HAI (group B); CaCO3 (group C); and placebo (group D) in a double-blind system using peripheral quantitative computed tomography (pQCT) with lumbar spine density measurement by dual-energy X-ray absorptiometry (DXA). Groups A, B, and C received 900 mg/day elemental calcium and D received none. In subjects of group A, but not B, C, and D, radial trabecular bone density increased significantly, to 106.2% +/- 2.1% of the initial value (mean +/- SEM). The increase of trabecular bone density was significantly different from the placebo group (D) only in AAACa (group A) and not in AACa (group B) and the calcium carbonate (group C). Cortical bone density increase was also greater in group A (but not in B and C) than in D. Lumbar spine density did not change significantly. AAACa was apparently more effective, increasing trabecular bone density more than AACa and CaCO3 containing the same amount of elemental calcium. PMID- 10874601 TI - Intake of fermented soybean (natto) increases circulating vitamin K2 (menaquinone 7) and gamma-carboxylated osteocalcin concentration in normal individuals. AB - Changes in circulating vitamin K2 (menaquinone-7, MK-7) and gamma-carboxylated osteocalcin concentrations in normal individuals with the intake of fermented soybeans (natto) were investigated. Eight male volunteers were given sequentially fermented soybeans (natto) containing three different contents of MK-7 at an interval of 7 days as follows: regular natto including 775 micrograms/100 g (MK-7 x 1) or reinforced natto containing 1298 micrograms/100 g (MK-7 x 1.5) or 1765 micrograms/100 g (MK-7 x 2). Subsequently, it was found that serum MK-7 and gamma carboxylated osteocalcin concentrations were significantly elevated following the start of dietary intake of MK-7 (1298 or 1765 micrograms/100 g). Serum undercarboxylated osteocalcin concentrations were significantly decreased by dietary MK-7 (1765 micrograms/100 g) supplementation. Moreover, the changes in serum MK-7 level with the frequency of dietary natto intake were examined in 134 healthy adults (85 men and 39 women) without and with occasional (a few times per month), and frequent (a few times per week) dietary intake of regular natto including MK-7 (775 micrograms/100 g). Serum MK-7 and gamma-carboxylated osteocalcin concentrations in men with the occasional or frequent dietary intake of natto were significantly higher than those without any intake. The present study suggests that intake of fermented soybean (natto) increases serum levels of MK-7 and gamma-carboxylated osteocalcin in normal individuals. PMID- 10874602 TI - Osteoblastic bone metastasis with unusual synthesis of trabeculae in the bone marrow space: a case report with a node-strut analysis. PMID- 10874603 TI - Calcitonin and parathyroid hormone: an evening seminar of the 17th Annual Meeting of the Japanese Society for Bone and Mineral Research, July 29, 1999. PMID- 10874604 TI - Analgesic mechanism of calcitonin. AB - Calcitonin is employed in clinical treatment to improve bone mass in osteoporosis and to relieve its accompanying pain, but its analgesic mechanism is still unclear. Using ovariectomized (OVX) rats that are well known as an animal model of osteoporosis, the antinociceptive effect of elcatonin ([Asu] eel calcitonin, ECT), a synthetic derivative of eel calcitonin, was examined with the tail withdrawal nociceptive test. Prolonged hyperalgesia was induced by OVX, and the antinociceptive effect of ECT was proved because peripherally and repeatedly administered ECT improved hyperalgesia. This effect of ECT was completely abolished by injections of p-chlorophenylalanine (PCPA), an inhibitor of serotonin biosynthesis, suggesting that the serotonergic system may be involved in antinociception. Spinal cord slices that retained an attached dorsal root were prepared, and blind whole-cell recordings were made from substantia gelatinosa (SG, lamina II) for electrophysiological analyses of a relationship between the effect of ECT and the serotonergic system. The descending serotonergic fibers, one of the inhibitory systems for nociceptive transmission, originate from the nucleus raphe magnus and terminate preferentially on SG of the spinal dorsal horn. Excitatory postsynaptic currents (EPSCs) evoked by dorsal root stimulation were then recorded from SG neurons, and the effects of serotonin on the EPSCs were compared in sham-operated and OVX rats. In addition, influence of ECT administration to OVX rats on the effects of serotonin was also examined. Glutamatergic short- and long-latency EPSCs, mediated by A delta and C afferent fibers, respectively, were observed after stimulation of the dorsal root, and both were depressed in amplitude by serotonin in sham rats, whereas only A delta mediated EPSCs but not C-mediated were inhibited in OVX rats. Interestingly, C mediated EPSCs were inhibited by serotonin in ECT-treated OVX as well as sham rats. A relationship between the fact that OVX induced hyperalgesia and that ECT alleviated this hyperalgesia was well correlated with changes in serotonergic inhibition of C-mediated EPSCs. These results suggest an alteration in the spinal serotonergic receptors that control the nociceptive transmission. Receptor binding assay using spinal membranes and [3H]-8-OH-DPAT as a radioactive ligand was used to assess the change in the receptors. Although the level of [3H]-8-OH DPAT-binding sites was decreased in OVX rats compared with sham rats, this reversed to the normal level in ECT-treated OVX rats. All the results strongly suggest that the behavioral and electrophysiological changes may be caused by an alteration in the level of spinal serotonergic receptor expression. PMID- 10874605 TI - Calcium paradox: consequences of calcium deficiency manifested by a wide variety of diseases. AB - Calcium deficiency is a global problem, especially in the aging population. Among various nutrients, calcium is one of the few that is still deficient in industrialized countries such as Japan and many Western countries. Calcium deficiency is readily connected with osteoporosis, which is a decrease of bone calcium content. Less well known is the calcium outflow from bone that occurs to prevent decrease of blood calcium in calcium deficiency caused by the parathyroid hormone, with consequent calcium overflow into soft tissues and the intracellular compartment. Such intracellular paradoxical Ca overload as a consequence of nutritional calcium deficiency may give rise to a number of diseases common in old age: hypertension, arteriosclerosis, diabetes mellitus, neurodegenerative diseases, malignancy, and degenerative joint disease. PMID- 10874606 TI - Minimally invasive coronary artery bypass grafting: one-year follow-up. AB - BACKGROUND: Use of the minimally invasive direct coronary artery bypass grafting (MIDCAB) technique has been associated with excellent primary results, and sparing of resources has been assumed. There is, however, a limited amount of information available concerning the results of mid-term follow-up. The purpose of this study was to present 1-year follow-up results of our first 130 consecutive MIDCAB patients. METHODS: MIDCAB operations, defined as no sternotomy, no cardiopulmonary bypass, and no aortic manipulation were started in our clinic in February 1996. One hundred thirty patients requiring invasive treatment of coronary artery disease who were not suitable for percutaneous transluminal angioplasty were included in this series. The main outcome measures were mortality, the need for subsequent invasive treatment, and 1-year NYHA classification. RESULTS: There was one hospital death, but during the first-year follow-up, four additional deaths occurred and three patients were reoperated on with conventional techniques. Five percutaneous transluminal coronary angioplasties (PTCAs) had to be performed, two because of anastomosic stenosis. Additionally, cardiac- or operation-related symptoms caused a total of 27 hospital visits among 23 patients during the first-year follow-up. Angiographic left internal thoracic artery (LITA)-left anterior descending artery (LAD) patency was 97.4% (37/38) (confidence interval [CI] ranged from 86.2% to 99.9%) at 3 months. After 1 year, 86.9% (113/130) of the patients were without symptoms. A clear improvement of the follow-up results was observed to be associated with increased experience during the study period. CONCLUSIONS: MIDCAB operations, after some experience, can be performed with relatively good outcome. However, special attention should be directed to determination of correct anastomosic site and to avoiding anastomosic stenosis. We also recommend extended mobilization of the ITA and use of specific stabilizers. PMID- 10874607 TI - Complications of port-access cardiac surgery. AB - Port-Access cardiac surgery is a recent technology that is undergoing rapid development. The learning curve associated with this technique is a challenge even for the skilled and experienced cardiac surgeon. Mainly because of femoral cannulation, the use of guidewires, and working through small incisions, Port Access cardiac surgery contains certain pitfalls that are clearly associated with the technology involved. These pitfalls currently require troubleshooting, but as the technology progresses, this may become less of an issue. Communicating these pitfalls to others is important to help others to avoid or better manage complications and to contribute to improving the technology of Port-Access techniques. PMID- 10874608 TI - Extended follow-up after atrial repair for transposition of the great arteries: a younger age at surgery improves late survival. AB - BACKGROUND: Surgical treatment for transposition of the great arteries (TGA) usually involves anatomic repair, although atrial repair is used in cases with special coronary artery patterns. METHODS: Records of all 239 patients surviving at least 30 days after atrial correction of TGA between 1962 and 1987 at the University Hospital in Zurich were reviewed. The mean length of follow-up time was 13.7 years (median 14.9 years; range from 0.05 to 30.1 years). The average age at surgery was 45.7 months (range from 7 days to 24.4 years). One hundred twenty-one patients had a simple TGA, whereas the remaining 118 had a complex TGA. RESULTS: The surviving patients were in NYHA class I at time of follow-up. The most common reasons for death were systemic right ventricular dysfunction and sudden rhythm disturbances. There was a major risk for late cardiac event in patients over 3 years old at operation (p = 0.02) and also in patients with complex TGA (p = 0.03). However, date of surgery, previous surgery, or the postoperative requirement for a pacemaker did not greatly affect late cardiac mortality. CONCLUSIONS: Although the procedure of choice for TGA is the arterial switch operation, the promising findings of the current long-term study, which reports the longest follow-up to date, indicate that the atrial switch operation is a good alternative procedure for the rare cases where the use of arterial switch procedures is limited. PMID- 10874609 TI - Late results with bioprosthetic valves in the elderly. AB - AIM: We report the long-term outcome of aortic and mitral bioprostheses in patients over 65 years of age at the time of implantation. The aim was to determine actuarial patient survival, causes of death, and the rate of documented primary structural deterioration. METHODS: One hundred ten patients > or = 65 years of age (mean, 73.4; range, 65-82) underwent successful bioprosthetic valve replacement (aortic, n = 71; mitral, n = 32; both, n = 7) from 1979 to 1985. The valve was pericardial in 39 cases and porcine in 78. The mean follow-up was 8.5 years (101.9 months-total; 934 patient-years; range, 2 months to 15 years). RESULTS: Actuarial patient survival was 79.6% (71-86) at 5 years and 62.4% (52 71) at 10 years. Forty-four patients died, 21 from valve-related causes and 23 from other causes. Thirteen patients (11.8%) had reoperation for valve-related complications: 10 structural deteriorations, 2 paravalvular leaks, and 1 case of endocarditis. One surgical death occurred (7.7%). Twenty-six percent of the patients were receiving anticoagulants because of atrial fibrillation, and 6.4% developed severe bleeding (2.9% patient-years). CONCLUSIONS: Long-term follow-up of these patients > 65 years of age, undergoing bioprosthetic value replacement surgery revealed a low rate of documented primary structural deterioration (0.95% per patient-year), a low mortality rate on reoperation (7.7%), and a high mortality rate due to non-value-related causes (52.3%). PMID- 10874610 TI - New indicator for the Fontan operation: diameters of the pulmonary veins in patients with univentricular heart. AB - BACKGROUND: Operative survival after the Fontan procedure is good; however, there are some patients with disappointing results, especially those with atrial isomerism. OBJECTIVES: We tested whether the diameter of the pulmonary veins, which is reported as a useful indicator of pulmonary blood flow, predicts operative results after the Fontan operation. PATIENTS AND METHODS: We evaluated 30 consecutive patients undergoing either the bidirectional Glenn anastomosis (BDG) or the Fontan operation. Age at operation ranged from 3 to 81 months (mean 30). Diagnosis was right or left isomeric heart in 15 patients, double-outlet right ventricle in 4 and various other malformations in 11. BDG was performed in 16 patients and the Fontan operation in 14 patients. The diameters of the pulmonary veins were measured proximal to the entrance into the atrium in the late phase of a pulmonary arteriogram. The pulmonary vein (PV) index (in mm2/m2) was calculated from the sum of the cross-sectional areas of these veins divided by the body surface area. RESULTS: Of the patients undergoing BDG (+/- ancillary procedures), 12 had successful results and 4 had unsuccessful results. The PV index for hemodynamically successful patients was 361 +/- 153 and 275 +/- 60 mm2/m2 (mean +/- SD) for unsuccessful patients (p = 0.30). Of the patients who underwent the Fontan operation, 13 had successful and 1 had unsuccessful results. The PV index for successful patients was > 285 mm2/m2 and 137 mm2/m2 for the nonsuccessful patients. The new pulmonary vascular resistance (PVR) calculated by using the PV index (mean pressure difference between the pulmonary artery and the atrium/PV index) for BDG patients with successful or unsuccessful results was 2.0 +/- 0.5 or 3.5 +/- 0.2 mmHg/mm2 per m2, respectively (p < 0.01). The new PVR for Fontan patients with successful results was < 2.0 mmHg/mm2 per m2, while that for the patient with an unsuccessful result was 4.4. The new PVR completely separated patients into successful and unsuccessful groups, while conventionally calculated PVR did not (p = 0.63). CONCLUSIONS: PV index appears to be a useful morphological indicator of pulmonary blood flow and "new" PVR may improve the decision-making strategy for patients presenting with univentricular heart, especially those associated with isomeric heart. PMID- 10874611 TI - Mitral valve replacement in the presence of massive posterior annular calcification. AB - Replacement of the mitral valve in the presence of extensive calcification of the posterior annulus is a technical challenge. The heavily calcified annulus often results in difficulties of seating the prosthesis and later periprosthetic leakage. A radical calcium debridement may leave a friable and thin annulus that contributes to the risks of prosthesis dehiscence and ventricular perforation. To avoid technical difficulties and associated catastrophic complications, we devised a new technique of mitral valve replacement that allows a surgeon to implant a prosthesis securely. This technique involves inserting a larger single tilting disc mechanical valve (Medtronic Hall disc) with intra-atrial anchorage over the posterior sector of the calcified annulus, orienting the working (major) orifice of the mechanical valve anteriorly, and thereby tilting the lesser occluder segment of the disc upward into the atrium and away from the calcification in diastole. By utilizing this method, we have successfully performed mitral valve replacement in two patients who exhibited massive calcification of the posterior mitral annulus. Postoperative transeosophageal echocardiography showed excellent hemodynamic performance of the implanted valves. We therefore recommend this simple, safe, and time-saving procedure as a feasible method to deal with this surgical dilemma. PMID- 10874612 TI - New technique of posterior mitral annuloplasty. AB - We have developed a new technique of mitral annuloplasty applicable in patients with rheumatic mitral regurgitation. This technique is designed to advance anteriorly the posterior annulus and leaflet and to gain more coaptation area. Technical details of this procedure are presented. PMID- 10874613 TI - Operative treatment of hypertrophic obstructive cardiomyopathy and aortic valve disease in infants. AB - BACKGROUND: The indications for operative intervention for symptomatic hypertrophic obstructive cardiomyopathy (HOCM) in infancy and childhood are not well defined because of the rarity of the lesion. The traditional surgical procedure consists of septal myectomy. In an attempt to further improve the outcome of HOCM associated with concentric left ventricular hypertrophy and aortic valve disease in infancy, we have combined resection of the left ventricular septum and free wall with a Ross-Konno procedure. METHODS: Three infants (aged 3, 4, and 10 months) with HOCM (left ventricular aortic gradients of 75, 95, and 110 mmHg), associated concentric left ventricular hypertrophy, and valvar aortic stenosis (n = 1) or combined valvar aortic stenosis and regurgitation (n = 2) underwent extensive resection of fibroelastosis and subendocardial myocardium of the left ventricular septum and free wall in combination with a Ross-Konno operation. All three patients had marked systolic anterior motion of the mitral valve. The length of the incision into the ventricular septum was 1.8, 2.0, and 2.3 cm. RESULTS: In all three patients this procedure resulted in a marked reduction of width of the left ventricular septum (median 9 mm vs 14 mm preoperatively) and the left ventricular posterior free wall (median 8 mm vs 12 mm preoperatively) and an almost twofold increase of the left ventricular end-diastolic volume (median 13.5 cm3 vs 7.0 cm3 preoperatively). The neo-aortic valve functioned normally. Systolic anterior motion of the anterior leaflet of the mitral valve had completely resolved in two patients and had markedly regressed in the remaining patient. At follow-up of 15, 17, and 26 months, two patients had absence of a left ventricular outflow tract gradient and the third patient had a residual sub-valvar gradient of 15 mmHg. CONCLUSIONS: The reported procedure may be a valuable technique in severe forms of hypertrophic cardiomyopathy associated with aortic valve disease. The operation results in enlargement of the left ventricular stroke volume and improvement of the left ventricular diastolic function, restores aortic valve anatomy and function, and abolishes or decreases systolic anterior motion of the mitral valve. PMID- 10874614 TI - Pentastarch versus albumin in cardiopulmonary bypass prime: impact on blood loss. AB - BACKGROUND: Albumin is commonly used as a volume expander in cardiopulmonary bypass (CPB) prime. Pentastarch, a low molecular weight hetastarch, may provide similar efficacy at decreased cost but is known to alter coagulation profiles. Infectious concerns forced the temporary withdrawal of albumin in our institution. Therefore we evaluated pentastarch as an alternative with regards to perioperative hemostasis and blood loss. METHODS: One hundred consecutive adult patients undergoing first-time aorto-coronary bypass were given 750 mL of 10% pentastarch (represented as P in calculations) diluted in 1000 mL of Ringer's solution added in their CPB prime. A similar control group of 100 consecutive patients had received 200 mL of 25% albumin (represented as A in calculations) diluted in 1500 mL of Ringer's solution. RESULTS: Postoperative prothrombin time (PT) was slightly higher with pentastarch (P: 14.9 +/- 1.5 seconds, A: 14.2 +/- 1.3 seconds, p = 0.003). Postoperative bleeding was also increased (P: 2337 +/- 1242 mL, A: 1981 +/- 1121 mL, p = 0.034), mostly because of recirculated shed mediastinal blood (P: 834 +/- 499 mL, A: 640 +/- 388, p = 0.002) rather than lost pleural tube blood (P: 1503 +/- 821 mL, A: 1341 +/- 824 mL, p = 0.16). Overall net blood loss (P: 2014 +/- 914 mL, A: 2061 +/- 1015, p = 0.73) was similar. Blood-product transfusion requirements and postoperative daily hematocrits did not differ. CONCLUSION: The diminished coagulability associated with this dose of pentastarch resulted in increased postoperative bleeding. However, with recirculation of shed mediastinal blood, there was no net increase in blood loss. In this setting, pentastarch may serve as a suitable alternative to albumin. PMID- 10874615 TI - Pharmacological criteria for ventricular assist device insertion following postcardiotomy shock: experience with the Abiomed BVS system. AB - BACKGROUND/AIM: The traditional approach to postcardiotomy shock includes inotropic support followed by the application of an intra-aortic balloon pump (IABP). Consideration toward insertion of a ventricular assist device (VAD) becomes necessary when these maneuvers fail to restore hemodynamic stability. The definition of maximal inotropic support, however, is lacking such that a standard formula for VAD insertion remains problematic. The purpose of this paper is to define the pharmacological thresholds for VAD implantation in the setting of postcardiotomy cardiogenic shock. METHODS: The medical records of all adult open heart operations performed at Hahnemann University Hospital, Philadelphia, PA, from 1 July 1996 through 1 July 1999 were reviewed. Specific data were collected on the hemodynamics and inotrope levels upon separation from cardiopulmonary bypass (CPB). The hospital course was reviewed with attention toward documenting hospital mortality. Cardiogenic shock was defined as systolic blood pressure (SBP) < 100 mmHg, mean pulmonary artery blood pressure (mPAP) > 25 mmHg, central venous pressure (CVP) > 15 mmHg, and cardiac index (CI) < 2.0 L/min/per m2. Inotrope dosages were defined as low, moderate, and high according to assigned values. A formula for VAD insertion was established if cardiogenic shock parameters were present in the setting of two or more high dose inotropes. Early VAD insertion was defined as implantation within three hours of the first attempt to wean from CPB. The VAD recipients were divided into two groups. Group A were VADs placed in conjunction with the formula. Group B was VADs placed in violation (excess) of the formula. The results of these two groups were compared. [table: see text] RESULTS: From 1 July 1996 to 1 July 1999, there were 3462 adult open heart operations performed at Hahnemann University Hospital, Philadelphia, Pa. The hospital mortality for patients successfully separating from CPB on no inotropes, low-dose, moderate-dose, one high-dose, two high-dose, and three high dose inotropes were approximately 2.0%, 3.0%, 7.5%, 21%, 42%, and 80% respectively. During this time there were 29 patients supported with the Abiomed BVS (Danvers, Mass) system for postcardiotomy cardiogenic shock. For the entire group of VAD recipients, there were 18 (62%) who were successfully weaned and 8 (28%) who were discharged from the hospital. For the 20 VAD recipients in group A, there were 16 (80%) who were successfully weaned and 8 (40%) who were discharged from the hospital. For the nine VAD recipients in group B, there were two (22%) who were successfully weaned and zero (0%) who were discharged from the hospital. Multiple organ system failure occurred in three (15%) in group A versus seven (78%) in group B patients, respectively. Early VAD insertion was accomplished in 17 (85%) group A patients and 2 (22%) group B patients. CONCLUSIONS: Hospital mortality correlates with the number and level of inotropic support necessary to separate from CPB following adult open heart surgery. The application of a standard pharmacological formula together with hemodynamic criteria for VAD insertion after postcardiotomy cardiogenic shock results in earlier insertion, lower incidence of postoperative MOSF, and improved wean and discharge rates. PMID- 10874616 TI - Mid-term outcome of surgical coronary ostial plasty: our experience. AB - The conventional coronary artery bypass procedure that uses venous or arterial conduit for isolated critical stenosis of the left main coronary artery (LMCA) restores a less physiological perfusion of the myocardium and uses an appreciable length of bypass material. Coronary ostial plasty has been described as an alternative surgical technique in proximal obstructive coronary artery disease without calcifications. Here we report 23 patients (15 males and 8 females aged 37-78 years; mean age 57 years) who underwent surgical ostial plasty. Ostial reconstruction with fresh pericardial patch was performed in all patients: 15 patients with LMCA stenosis, 6 patients with right coronary (RC) ostial stenosis, and 2 patients with both RC artery and LMCA stenosis. In seven cases, coronary artery bypass grafting was added for contralateral distal stenosis with a total of five arterial conduits and six venous grafts. One patient died; the ostial plasty and grafts were patent at necropsy. Thallium-201 myocardial scintigraphy under stress at 30 days to 6 months after operation demonstrated good myocardial perfusion in 21 of 22 patients. Coronary angiography at follow-up (49 +/- 8 months) demonstrated good surgical ostial plasty results in 21 of 22 patients and good coronary flow in 19 of 22 patients; angiographic study at mid-term follow-up revealed only one failure of the surgical ostial plasty technique associated with venous graft obstruction. In 2 other patients CABG failure due to venous graft obstruction (1 patient) or distal stenotic lesions of the left coronary artery (1 patient) was noted. The overall successful outcome of the surgical ostial plasty was 22 of 23. We believe that surgical angioplasty of the coronary ostia may be used in the presence of proximal noncalcified obstructive lesions as an alternative technique, which offers a more physiological revascularization; it also spares grafting material and allows subsequent percutaneous transluminal angioplasty or coronary artery bypass surgery. PMID- 10874617 TI - Coronary ostial aneurysms after composite graft replacement. AB - Coronary ostial aneurysms after composite graft replacement of the ascending aorta and aortic valve is a rare complication. We report two patients with Marfan syndrome who developed coronary ostial aneurysms at the sites of the coronary anastomosis, presumably because of oversized windows made in the graft. They were successfully treated by redo composite graft replacement. To prevent this complication, it is important to consider that the hole made in the tube graft should not be larger than the diameter of the respective coronary ostium to avoid exposure of the diseased aortic wall to the circulating blood as much as possible, and that the suture used to anastomose the coronary buttons should pass through the rim of the ostium rather than through the aortic wall surrounding it. PMID- 10874618 TI - One-stage repair of interrupted aortic arch and aortopulmonary window with an autologous arterial flap. AB - Interrupted aortic arch associated with an aortopulmonary window is a rare congenital malformation that requires an early diagnosis and surgical treatment to avoid irreversible pulmonary vascular changes. We describe herein successful one-stage repair in a neonate without the use of pericardium or other prosthetic material. Use of a transaortic approach permitted both accurate planning of the appropriate size of pulmonary trunk flap and also facilitated easy repair of the aortopulmonary septal defect. The growth potential of both great arteries was optimized. PMID- 10874619 TI - Dermal exposure to pesticides modifies antioxidant enzymes in tissues of rats. AB - Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) activities were determined in rat tissues after dermal exposure to pesticides. Two experiments were conducted in male SD rats, 190-210 g body weight. Acephate (ACP), methamidophos (MAP) and nicotine (NIC) were dissolved either individually or together in 0.25 mL of 50% ethanol, which contained: AP = 12.6 or MAP 1.3 or NIC = 9.6 mg; EXP 1--individual pesticide exposure; 64 rats, 16/group; EXP 2- mixture of AP + MAP + NIC at levels of 1X, 2X, 3X; 48 rats, 12/group; 0.25 mL of solution or ethanol (Controls) was applied to 25 mm2 area of shaved skin 3 times a week. Half the rats were terminated after 4 weeks and the rest after 4 weeks of stopping exposure. Single pesticides decreased erythrocyte (RBC) SOD by 17% after exposure and in the NIC group after post exposure (P#0.05). Increasing concentrations of AP + MAP + NIC mixture elevated RBC SOD by 22% in the 2X and 3X groups and CAT by 13% in the 3X group (P#0.05); post exposure increased RBC SOD by 2-3 fold and CAT activity by 13% in all 3 groups. Liver GPX increased by 30 40% and CAT decreased by 12% in all exposed and post exposed groups (P#0.05). The results suggest that dermal exposure to mixtures of pesticides can selectively induce SOD, CAT and GPX activities in RBC and liver. PMID- 10874620 TI - Degradation of atrazine, metolachlor, and pendimethalin in pesticide-contaminated soils: effects of aged residues on soil respiration and plant survival. AB - This study was conducted to determine the effects of pesticide mixtures on degradation patterns of parent compounds as well as effects on soil microbial respiration. Bioavailability of residues to sensitive plant species was also determined. Soil for this study was obtained from a pesticide-contaminated area within an agrochemical dealer site. Degradation patterns were not affected by the presence or absence of other herbicides in this study. Atrazine concentrations were significantly lower at 21 through 160 days aging time compared to day 0 concentrations. Metolachlor and pendimethalin concentrations were not significantly different over time and remained high throughout the study. Microbial respiration was suppressed in treated soils from day 21 to day 160. Soybean and canola were the most successful plant species in the germination and survival tests. Generally, with increased aging of pesticides in soil, germination time decreased. Survival time of plants increased over time for some treatments indicating possible decreased bioavailability of pesticide residues. In some cases, survival time decreased at the longer 160-day aging period, possibly indicating a change in bioavailability, perhaps as the result of formation of more bioavailable and phytotoxic metabolites. No interactive effects were noted for mixtures of pesticides compared to individually applied pesticides in terms of degradation of the parent compound or on seed germination, plant survival, or microbial respiration. PMID- 10874621 TI - Kinetics of simazine advanced oxidation in water. AB - Comparison of the effects and kinetics of UV photolysis and four advanced oxidation systems (ozone, ozone/hydrogen peroxide, ozone/UV radiation and UV radiation/hydrogen peroxide) for the removal of simazine from water has been investigated. At the conditions applied, the order of reactivity was ozone < ozone/hydrogen peroxide < UV radiation < ozone/UV radiation and UV radiation/hydrogen peroxide. Rate constants of the reactions between ozone and simazine and hydroxyl radical and simazine were found to be 8.7 M-1s-1 and 2.1 x 10(9) M-1s-1, respectively. Also, a quantum yield of 0.06 mol.photon-1 was found for simazine at 254 nm UV radiation. The high value of the quantum yield corroborated the importance of the direct photolysis process. Percentage contributions of direct reaction with ozone, reaction with hydroxyl radicals and direct photolysis were also quantified. PMID- 10874622 TI - The influence of metam sodium on soil respiration. AB - A laboratory experiment was performed in order to evaluate the extent to which metam sodium (MS) applied at two different recommended rates and its degradation product, methyl isothiocyanate (MITC), affect soil respiration. Results suggest that MS degradation to MITC was complete within 4 hours and that MITC decomposed quickly in a few days, except in the soil containing high organic matter where it was still present after 15 days. Following the addition of MS, a lag phase appeared in CO2-C evolution in the soil. It was longer for the higher dose of MS added and for the two soils with low organic C content. The dynamics of the process was described by the Bonde and Rosswall model and by the Gompertz RS E model for the untreated and the MS-treated soils, respectively. PMID- 10874623 TI - Analytical methods for the determination of alachlor, metolachlor, simazine and atrazine mobility in soils. AB - A method for the determination of the mobility of the herbicides, alachlor, metolachlor, simazine and atrazine in soil is described. The method is based on the use of soil thin-layer chromatography (TLC) and does not require the use of radiolabelled compounds. Soil on the TLC plate after development was separated into various bands, the material in each band was extracted with solvents and analyzed by gas chromatography. PMID- 10874624 TI - A preliminary examination of the translocation of microencapsulated cyfluthrin following applications to the perimeter of residential dwellings. AB - Methods have been developed to monitor the translocation of microencapsulated cyfluthrin following perimeter applications to residential dwellings. A pilot study was implemented to determine both the potential for application spray to drift away from dwellings and the intrusion of residues into homes following perimeter treatments. Residential monitoring included measuring spray drift using cellulose filter paper and the collection of soil samples from within the spray zone. In addition, interior air was monitored using fiberglass filter paper as a sorbent medium and cotton ball swabs were used to collect surface wipes. Fortification of matrixes resulted in recoveries of > 90%. Spray drift was highest at the point of application and declined to low but measurable levels 9.1 m from the foundations of dwellings. Soil residues declined to low, but measurable levels by 45 days post-application. No cyfluthrin was measured from indoor air; however, some interior surfaces had detectable levels of cyfluthrin until three days post-application. Findings indicate that spray drift resulting from perimeter applications might contaminate non-target surfaces outside the spray zone. Soil borne residues may serve as persistent sources for human exposure and potentially intrude into dwellings through the activities of occupants and pets. Residues do not appreciably translocate through air and consequently inhalation is not a likely route for human exposure. Surface residues detected indoors suggest that the physical movement of residues from the exterior to the interior might be a viable route of movement of residues following this type of application. PMID- 10874625 TI - Effect of fatty acids and oils on photodegradation of azadirachtin-A. AB - Azadirachtin-A on exposure to UV-light (254 nm) as a thin film on glass surface gave a isomerised (Z)-2-methylbut-2-enoate product. Half-life of azadirachtin-A as thin film under UV light was found to be 48 min. Azadirachtin-A was irradiated along with saturated and unsaturated fatty acids, and fatty oils under ultra violet light as thin film. Saturated fatty acid increased the rate of photodegradation of azadirachtin-A, whereas unsaturated fatty acids such as oleic, linoleic and elaidic acid reduced the rate of degradation. Castor, linseed and olive oil accelerated the rate of degradation, whereas neem oil showed no or little change in the rate of degradation of azadirachtin-A. None of these fatty acids and fatty oils were effective in controlling the rate of degradation of azadirachtin-A under UV-light as thin film. PMID- 10874626 TI - Response of selected poultry cecal probiotic bacteria and a primary poultry Salmonella typhimurium isolate grown with or without glucose in liquid batch culture. AB - The objective of this study was to determine whether fermentation by a cecal probiotic co-culture of an Enterococcus sp. and Veillonella sp. would inhibit the in vitro growth of a S. typhimurium poultry isolate. The growth rates of S. typhimurium and Enterococcus were significantly reduced at pH 5. At the two pH levels, there was a significant (p < 0.001) increase at 24 h in colony forming units for each of the bacteria enumerated from the mixed culture compared to the respective pure culture enumerations. S. typhimurium was not inhibited in mixed cultures. The mixed cultures produced more acetate than any of the pure cultures and lactate produced by Enterococcus appeared to be utilized by Veillonella. PMID- 10874627 TI - Recovery of a marker strain of Salmonella typhimurium in litter and aerosols from isolation rooms containing infected chickens. AB - Screening of poultry flocks for foodborne pathogen Salmonella contamination is critical for Salmonella control in preharvest stages of poultry production. In this study, two sampling methods (litter and air filter) were compared for detection of S. typhimurium from experimentally infected chicks some of which had received either a probiotic competitive exclusion culture or transfer of cecal contents from salmonellae-free adult birds. At 4, 9, and 11 days after inoculation, S. typhimurium samples were enumerated by selective plating. For both types of sampling, the control birds yielded the greatest levels of environmental contamination followed by the samples from the probiotic inoculated birds with the birds receiving the cecal transfer culture having the lowest levels of contamination. Although the two sampling methods responded in a similar fashion, detection sensitivity needs to be increased for air filter sampling. PMID- 10874628 TI - Effect of common food preservatives on mycelial growth and spore germination of Fusarium oxysporum. AB - The growth and spore germination inhibition of Fusarium oxysporum f.sp. radicis cucumerinum by the common food additives: acetic acid, formic acid potassium sorbate, propionic acid, sorbic acid, and the fungistatic agent sec-butylamine was examined in vitro. The inhibitory efficacy of these chemicals decreased in the following order: sorbic acid, potassium sorbate, propionic acid, acetic acid, sec-butylamine and formic acid. At pH 6.4, the ED50 value for mycelium growth was: 976 ppm for sorbic acid, 1292 ppm for potassium sorbate, 2435 ppm for propionic acid, 3805 ppm for acetic acid, 3962 ppm for sec butylamine and 4668 ppm for formic acid. The ED50 value for spore germination was: 225 ppm for potassium sorbate, 1201 ppm for sorbic acid, 1402 ppm for propionic acid, 1600 ppm for sec-butylamine, 1957 ppm for acetic acid and 2485 ppm for formic acid. PMID- 10874630 TI - New criteria for improved diagnosis of Bardet-Biedl syndrome: results of a population survey. AB - Bardet-Biedl syndrome (BBS) is an autosomal recessive condition characterised by rod-cone dystrophy, postaxial polydactyly, central obesity, mental retardation, hypogonadism, and renal dysfunction. BBS expression varies both within and between families and diagnosis is often difficult. We sought to define the condition more clearly by studying 109 BBS patients and their families, the largest population surveyed to date. The average age at diagnosis was 9 years, which is late for such a debilitating condition, but the slow development of the clinical features of BBS probably accounts for this. Postaxial polydactyly had been present in 69% of patients at birth, but obesity had only begun to develop at around 2-3 years, and retinal degeneration had not become apparent until a mean age of 8.5 years. Our study identified some novel clinical features, including neurological, speech, and language deficits, behavioural traits, facial dysmorphism, and dental anomalies. In the light of these features we propose a revision of the diagnostic criteria, which may facilitate earlier diagnosis of this disorder. We present evidence for an overlapping phenotype with the Laurence Moon syndrome and propose a unifying, descriptive label be adopted (polydactyly obesity-kidney-eye syndrome). We report an increased prevalence of renal malformations and renal cell carcinoma in the unaffected relatives of BBS patients and suggest that these may be a consequence of heterozygosity for BBS genes. Our findings have important implications for the care of BBS patients and their unaffected relatives. PMID- 10874631 TI - Mutations of the retinal specific ATP binding transporter gene (ABCR) in a single family segregating both autosomal recessive retinitis pigmentosa RP19 and Stargardt disease: evidence of clinical heterogeneity at this locus. AB - Stargardt disease (STGD) is an autosomal recessive macular dystrophy of childhood characterised by bilateral loss of central vision over a period of several months. STGD has been mapped to chromosome 1p22.1 and recently ascribed to mutations in the retinal specific ATP binding transporter gene (ABCR). The fundus flavimaculatus with macular dystrophy (FFM), an autosomal recessive condition responsible for gradual loss of visual acuity in adulthood (second to third decade) has also been mapped to the same locus. However, a gene for autosomal recessive retinitis pigmentosa with distinctive features of choriocapillaris atrophy at an advanced stage (RP19) has been mapped to the genetic interval encompassing the STGD gene on chromosome 1p (D1S435-D1S236), raising the question of whether, despite striking differences in clinical course and presentation, RP19 and STGD might be allelic disorders at the ABCR locus. In a family segregating RP and STGD in two first cousins, we found that heterozygosity for a splicing mutation in the ABCR gene (1938-1 G-->A) resulted in STGD while hemizygosity for this splice mutation resulted in RP, and when studying the RP patient's parents, we found a maternal non-contribution with apparent segregation of a null allele ascribed to a partial deletion of the ABCR gene. The present study shows that, despite striking clinical differences, RP19 and STGD are allelic disorders at the ABCR locus. PMID- 10874629 TI - Clinical mitochondrial genetics. PMID- 10874632 TI - Incomplete masculinisation of XX subjects carrying the SRY gene on an inactive X chromosome. AB - 46,XX subjects carrying the testis determining SRY gene usually have a completely male phenotype. In this study, five very rare cases of SRY carrying subjects (two XX males and three XX true hermaphrodites) with various degrees of incomplete masculinisation were analysed in order to elucidate the cause of sexual ambiguity despite the presence of the SRY gene. PCR amplification of 20 Y chromosome specific sequences showed the Yp fragment to be much longer in XX males than in true hermaphrodites. FISH analysis combined with RBG banding of metaphase chromosomes of four patients showed that in all three true hermaphrodites and in one XX male the Yp fragment was translocated onto a late replicating inactive X chromosome in over 90% of their blood lymphocytes. However, in a control classical XX male with no ambiguous features, the Yp fragment (significantly shorter than in the XX male with sexual ambiguity and only slightly longer than in XX hermaphrodites) was translocated onto the active X chromosome in over 90% of cells. These studies strongly indicate that inactivation on the X chromosome spreading into a translocated Yp fragment could be the major mechanism causing a sexually ambiguous phenotype in XX (SRY+) subjects. PMID- 10874633 TI - An analysis of the distribution of hetero- and isodisomic regions of chromosome 7 in five mUPD7 Silver-Russell syndrome probands. AB - Silver-Russell syndrome (SRS) shares common features of intrauterine growth retardation (IUGR) and a number of dysmorphic features including lateral asymmetry in about 50% of subjects. Its genetic aetiology is complex and most probably heterogeneous. Approximately 7% of patients with SRS have been found to have maternal uniparental disomy of chromosome 7 (mUPD7). Genomic DNA samples from five SRS patients with mUPD7 have been analysed for common regions of isodisomy using 40 polymorphic markers distributed along the length of chromosome 7. No regions of common isodisomy were found among the five patients. It is most likely that imprinted gene(s) rather than recessive mutations cause the common phenotype. Heterodisomy of markers around the centromere indicated that the underlying cause of the mUPD7 is a maternal meiosis I non-disjunction error in these five subjects. PMID- 10874634 TI - Complex consanguinity associated with short rib-polydactyly syndrome III and congenital infection-like syndrome: a diagnostic problem in dysmorphic syndromes. AB - Short rib-polydactyly syndromes (SRPS) are a heterogeneous group of recessively inherited lethal skeletal dysplasias. Four types have been recognised. However, overlap in the clinical and radiological features of the four types has led to difficulties in distinguishing between them. The congenital infection-like syndrome is an autosomal recessive syndrome characterised by mental retardation, microcephaly, seizures, and intracranial calcifications. We report a complex consanguineous family of Baluchi origin in whom short rib-polydactyly type III and congenital infection-like syndrome are segregating. Four children inherited SRPS III, one inherited congenital infection-like syndrome, and one inherited both. Although the radiological features in all the children with SRPS in this report were typical of type III, there was overlap in the clinical features with the other types of SRP syndromes. Furthermore, the child who inherited both SRPS III and congenital infection-like syndrome had CNS malformations in addition to periventricular calcification. CNS malformations have been described in SRPS types II and IV but not type III. This report further highlights the overlap between the different types of SRP syndrome. Moreover, it draws attention to the importance of considering the possibility of two recessive syndromes in the same child in complex consanguineous families when features overlap two syndromes. PMID- 10874635 TI - Screening for the fragile X syndrome among the mentally retarded: a clinical study. The Collaborative Fragile X Study Group. AB - The fragile X syndrome is characterised by mental retardation with other features such as a long face with large, protruding ears, macro-orchidism, and eye gaze avoidance. This X linked disorder is caused by an expanded CGG repeat in the first exon of the fragile X mental retardation (FMR1) gene which is associated with shut down of transcription and absence of the fragile X mental retardation protein (FMRP). Molecular testing is used for detection of patients and carriers of the fragile X syndrome. In a screening programme for the fragile X syndrome in the south west of The Netherlands, 896 males and 685 females with an unknown cause for their mental retardation were scored on seven fragile X features. All were tested by DNA analysis and 11 new cases were diagnosed. The seven item checklist allowed exclusion from further testing in 86% of the retarded males (95% CI 0.83-0.88) without missing either any of the newly diagnosed cases or, in retrospect, any of the 50 previously diagnosed cases known to our department. These results showed that clinical preselection for DNA testing in mentally retarded males is feasible using a simple scoring list, which will increase the efficiency of further testing eightfold. PMID- 10874636 TI - A new simple enzyme assay for pre- and postnatal diagnosis of infantile neuronal ceroid lipofuscinosis (INCL) and its variants. AB - Palmitoyl-protein thioesterase (PPT) deficiency was recently shown to be the primary defect in infantile neuronal ceroid lipofuscinosis (INCL). The available enzyme assay is complicated and impractical for diagnostic use and is, in practice, unavailable. We have developed a new fluorimetric assay for PPT based on the sensitive fluorochrome 4-methylumbelliferone. This PPT assay is simple, sensitive, and robust and will facilitate the definition of the full clinical spectrum associated with a deficiency of PPT. PPT activity was readily detectable in fibroblasts, leucocytes, lymphoblasts, amniotic fluid cells, and chorionic villi, but was profoundly deficient in these tissues from INCL patients. Similarly, a deficiency of PPT was shown in patients with the variant juvenile NCL with GROD. These results show that rapid pre- and postnatal diagnosis can be performed with this new enzyme assay for PPT. PMID- 10874637 TI - Identification of two different mutations in the PDS gene in an inbred family with Pendred syndrome. AB - Recently the gene responsible for Pendred syndrome (PDS) was isolated and several mutations in the PDS gene have been identified in Pendred patients. Here we report the occurrence of two different PDS mutations in an extended inbred Turkish family. The majority of patients in this family are homozygous for a splice site mutation (1143-2A-->G) affecting the 3' splice site consensus sequence of intron 7. However, two affected sibs with non-consanguineous parents are compound heterozygotes for the splice site mutation and a missense mutation (1558T-->G), substituting an evolutionarily conserved amino acid. The latter mutation has been found previously in two Pendred families originating from The Netherlands, indicating that the 1558T-->G mutation may be a common mutation. PMID- 10874638 TI - Detection of an atypical 7q11.23 deletion in Williams syndrome patients which does not include the STX1A and FZD3 genes. AB - We present two patients with the full Williams syndrome (WS) phenotype carrying a smaller deletion than typically observed. The deleted region spans from the elastin gene to marker D7S1870. This observation narrows the minimal region of deletion in WS and suggests that the syntaxin 1A and frizzled genes are not responsible for the major features of this developmental disorder and provides important insight into understanding the genotype-phenotype correlation in WS. PMID- 10874639 TI - Two sibs with microcephaly, hygroma colli, renal dysplasia, and cutaneous syndactyly: a new lethal MCA syndrome? AB - We report two sibs of Turkish descent with multiple congenital anomalies including severe microcephaly, hygroma colli, cystic renal dysplasia, and bilateral cutaneous syndactyly of toes IV-V. In addition, the second sib presented with bilateral fusion of the eyelids, a bicornuate uterus, and clitoromegaly. The parents are first cousins, which suggests autosomal recessive inheritance. In reviewing previously published reports, several cases were found with cerebral, renal, and digital anomalies as the main features. Several of the additional symptoms present in the second sib were suggestive of Fraser syndrome, but the severe microcephaly in both sibs is unusual. The differential diagnosis is discussed, including the possibility of an entirely new entity in the broad spectrum of syndromes with cerebral, renal, and digital anomalies. PMID- 10874640 TI - A consanguineous family with Hirschsprung disease, microcephaly, and mental retardation (Goldberg-Shprintzen syndrome). AB - Hirschsprung disease, mental retardation, microcephaly, and specific craniofacial dysmorphism were observed in three children from a large, consanguineous, Moroccan family. A fourth child showed similar clinical features, with the exception of Hirschsprung disease. The association of these abnormalities in these children represents the Goldberg-Shprintzen syndrome (OMIM 235730). Mutation scanning of genes potentially involved in Hirschsprung disease, RET, GDNF, EDN3, and EDNRB, showed a sequence variant, Ser305Asn, in exon 4 of the EDNRB gene in the index patient of this family. The Ser305Asn substitution present in two of the four patients and four healthy relatives and absent in one of the remaining two patients illustrates the difficulties in interpreting the presence of mutations in families with Hirschsprung disease. It is unlikely that the EDNRB variant contributes to the phenotype. This consanguineous family might be useful for the identification of a Goldberg-Shprintzen locus. PMID- 10874641 TI - CDKN2A mutations in Spanish cutaneous malignant melanoma families and patients with multiple melanomas and other neoplasia. AB - The CDKN2A gene has been implicated in cutaneous malignant melanoma (CMM) in about 40% of families with linkage to chromosome 9p21, while a small proportion of families have mutations in the CDK4 gene. In order to estimate the importance of these genes in the predisposition to CMM in Spanish families and patients we have analysed, by SSCA, a total of 56 subjects belonging to 34 CMM families, and nine patients with multiple CMM and other neoplasia. We have detected germline CDKN2A mutations in six out of the 34 families (17%). A frameshift mutation (358delG) and four missense mutations (G59V, G101W (two cases), D84Y, and R87W) were identified. Five CMM patients from different families (14%) carried the A148T variant, which is known not to affect p16 activity. No mutations were detected in the patients with multiple CMM or other neoplasms. We have not found mutations either in exon 1 beta of the CDKN2A gene or in exon 2A of CDK4. Linkage analysis of the 9p21 region showed exclusion for one of the families for CMM and for four families for CMM/dysplastic naevi. This study indicates a small role for CDKN2A in Spanish CMM families and suggests that other genes are also responsible for CMM predisposition. PMID- 10874642 TI - Simultaneous decrease of telomere length and telomerase activity with ageing of human amniotic fluid cells. PMID- 10874643 TI - 45,X/47,XX, + 18 constitutional mosaicism: clinical presentation and evidence for a somatic origin of the aneuploid cell lines. PMID- 10874644 TI - Prevalence of Prader-Willi and Angelman syndromes among mentally retarded boys in Brazil. PMID- 10874645 TI - Mutation of FGFR2 (cys278phe) in craniolacunia and pansynostosis. PMID- 10874646 TI - Smith-Magenis syndrome and tetralogy of Fallot. PMID- 10874647 TI - Use of the alkaline comet assay to monitor DNA damage in technicians exposed to low-dose radiation. PMID- 10874648 TI - An inappropriate title: "Why most workers with occupational repetitive trauma do not file for workers' compensation". PMID- 10874649 TI - Epidemiology of compensable work-related ocular injuries and illnesses: incidence and risk factors. AB - Incidence rates of work-related compensable ocular injuries/illnesses and associated risk factors were estimated by using a state-managed workers' compensation database. The annual incidence rate was estimated to be 537 per 100,000 employees. The majority of the ocular injuries and illnesses resulted from foreign bodies in the external eye (incidence rate 194 per 100,000 employees). Incidence rates for superficial eye injury, atopic conjunctivitis, burn, keratitis, chronic conjunctivitis, and contusion were 168.3, 30.9, 28.0, 23.4, 17.9, and 15.3 per 100,000 employees, respectively. The highest incidence rate was observed in the agricultural sector, with male employees having higher rates than female employees. Cooks, housekeepers, and food service workers had higher risk of atopic conjunctivitis (relative risk, 3.2 to 7.3) compared with other workers. The majority of the atopic conjunctivitis illnesses and burn injuries were associated with chemical exposures. Reduction of exposures and targeted intervention among high-risk workers should reduce the incidence of work related ocular injuries and illnesses. PMID- 10874650 TI - Lysyl oxidase activity in the cells of flexor retinaculum of individuals with carpal tunnel syndrome. AB - Lysyl oxidase (LO) is produced by myofibroblast cells in some tissues and can be influenced by transforming growth factor beta 1 (TGF beta 1). Myofibroblast-like cells are present in the flexor reticulum of patients with carpal tunnel syndrome (CTS). The goal of the current study was to determine LO activity and the effects of TGF beta 1 on LO expression in the cells from patients with CTS. Tissues from both hands of five individuals with CTS were used for this study. LO activity with and without TGF beta 1 stimulation was assayed in 7-day cell culture specimens. A significant difference in LO activity among individual patients, but not between right and left hands of the same patient, was observed. There was no correlation between the severity of CTS determined by nerve conduction studies and LO activity. Addition of TGF beta 1 significantly increased LO in all cell lines. PMID- 10874651 TI - Direct and indirect costs of rheumatoid arthritis to an employer. AB - This study is among the first to estimate the overall economic burden of rheumatoid arthritis (RA) from an employer perspective. The annual, per capita cost of RA was determined for beneficiaries of a major employer by analyzing medical, pharmaceutical, and disability claims data. The incremental costs related to RA were determined by matching RA patients to a case-control group of individuals with no recorded RA treatment. The utilization of health care services as well as the rate of disability among RA patients was substantially higher than among the controls. For example, annual, per capita employer expenditures for RA employees with disability were almost 3 times those for their controls ($17,822 vs $6131, respectively). Treatment to address not only the severity but also the progression of RA may substantially reduce overall employer expenditures for this disease. PMID- 10874652 TI - Disability case management: an impact assessment in an automotive manufacturing organization. AB - A multifaceted disability management program was instituted at an automotive manufacturing organization to control rising workers' compensation costs. A pilot program showed major cost savings over a 9-month period. When total and component disability leave rates were calculated as a percentage of the available workforce and tracked on a weekly basis over the subsequent 3 years, total disability leave rates fell by nearly 50%. This was largely attributable to an approximately 50% decrease in the extended (> 1-year) disability leave rate and a 75% decrease in the workers' compensation leave rate. A novel approach to biostatistical analysis showed a good fit of weekly disability leave rates to a Poisson random variable distribution with an identifiable break point at about 1 1/2 years after observation for extended disability leaves and at 2 years for workers' compensation leaves. This biostatistical approach may prove generalizable to tracking leave rates in other organizations. PMID- 10874653 TI - Does perchlorate in drinking water affect thyroid function in newborns or school age children? AB - Perchlorate is known to suppress thyroid function by inhibiting uptake of iodide by the human thyroid at doses of 200 mg/day or greater. A study was conducted to investigate the potential effects of perchlorate in drinking water on thyroid function in newborns and school-age children. A total of 162 school-age children and 9784 newborns were studied in three proximate cities in northern Chile that have different concentrations of perchlorate in drinking water: Taltal (100 to 120 micrograms/L), Chanaral (5 to 7 micrograms/L), and Antofagasta (non detectable: < 4 micrograms/L). Among schoolchildren, no difference was found in thyroid-stimulating hormone levels or goiter prevalence among lifelong residents of Taltal or Chanaral compared with those of Antofagasta, after adjusting for age, sex, and urinary iodine. No presumptive cases of congenital hypothyroidism were detected in Taltal or Chanaral; seven cases were detected in Antofagasta. Neonatal thyroid-stimulating hormone levels were significantly lower in Taltal compared with Antofagasta; this is opposite to the known pharmacological effect of perchlorate, and the magnitude of difference did not seem to be clinically significant. These findings do not support the hypothesis that perchlorate in drinking water at concentrations as high as 100 to 120 micrograms/L suppresses thyroid function in newborns or school-age children. PMID- 10874654 TI - Natural rubber latex: glove use, sensitization, and airborne and latent dust concentrations at a Denver hospital. AB - Exposure to natural rubber latex may cause immediate hypersensitivity reactions. Published latex sensitization prevalence rates range from 2.9% to 22% among health care workers, and from 0.12% to about 20% of occupationally unexposed populations. In this study, self-administered questionnaires addressed job and personal characteristics, glove use, and symptoms in two groups of hospital workers: those who regularly used latex gloves and those who did not. Serum was tested for latex-specific immunoglobulin E. Air, surface, and air-filter dust samples for natural rubber latex were collected. The prevalence of latex sensitization was 6.3% in the non-users and 6.1% in the latex glove users (P = 0.9); 81.3% of sensitized workers were atopic compared with 59.5% of non sensitized workers (P < 0.05). Reporting of work-related hand dermatitis was more common in the latex glove users (23.4%) than in the non-users (4.9%), as were rhino-conjunctivitis (16.3% and 7.9%, respectively, [P < 0.01]), and hand urticaria (9.9% and 2.1%, respectively, [P < 0.01]). There was no significant difference in work-related symptoms between the sensitized and non-sensitized workers. Environmental concentrations of latex were higher in the work areas of the non-sensitized workers, but higher in the clinical than in the non-clinical areas. Occupational latex glove use was not a risk factor for sensitization. PMID- 10874655 TI - Compensation for occupational injury and disease in Norway: ranking of job groups. AB - The health risk of various job groups in Norway was estimated by ranking them according to the annual occupational insurance costs per capita. This was done by dividing the costs of work-related injury and disease from 1991 to 1996 in various job groups by the number of workers in these groups. Occupational groups were also ranked according to total annual costs. The five occupational groups with the highest total costs were metalworkers, woodworkers, nursing-related workers, fisheries workers, and teachers. The groups with the highest annual cost per worker were shoe and leather workers, oil and gas extractors, fisheries workers, miners and quarry workers, and ship's officers. Fisheries workers and ship's officers were ranked among the top 10 positions on both lists and deserve priority in preventive measures. PMID- 10874656 TI - Odor perception: multiple chemical sensitivities, chronic fatigue, and asthma. AB - Patients with multiple chemical sensitivities (MCS) often report heightened sensitivity to odors. Odor detection thresholds to phenyl ethyl alcohol (PEA) and pyridine (PYR) were evaluated as a measure of odor sensitivity for 33 MCS subjects, 13 chronic fatigue syndrome subjects, 16 asthmatic subjects, and 27 healthy controls. Odor identification ability (based on University of Pennsylvania Smell Identification Test results) and ratings in response to four suprathreshold levels of PEA and PYR were also assessed. Odor detection thresholds for PEA and PYR and odor identification ability were equivalent for all groups; however, when exposed to suprathreshold concentrations of PEA, MCS subjects reported significantly more trigeminal symptoms and lower esthetic ratings of PEA. No group differences were found in response to suprathreshold concentrations of PYR. In summary, MCS subjects did not demonstrate lower olfactory threshold sensitivity or enhanced ability to identify odors accurately. Furthermore, they were differentiated from the other groups in their symptomatic and esthetic ratings of PEA, but not PYR. PMID- 10874657 TI - Non-malignant respiratory diseases and lung cancer among Chinese workers exposed to silica. AB - The objective of this study was to explore whether a medical history for non malignant respiratory disease contributes to an increased lung cancer risk among workers exposed to silica. We analyzed data from a nested case-control study in 29 dusty workplaces in China. The study population consisted of 316 lung cancer cases and 1356 controls matched to cases by facility type and decade of birth who were alive at the time of diagnosis of the index case and who were identified in a follow-up study of about 68,000 workers. Age at first exposure and cigarette smoking were accounted for in the analysis. Smoking was the main risk factor for both lung cancer and chronic bronchitis. Lung cancer risk showed a modest association with silicosis and with cumulative silica exposure, which did not vary by history of previous pulmonary tuberculosis. Among subjects without a medical history for chronic bronchitis or asthma, lung cancer risk was associated with silicosis (odds ratio [OR], 1.6; 95% confidence interval [CI], 1.1 to 2.2), and it was increased in each quartile of cumulative silica exposure. However, risk was not elevated in the highest quartile (OR, 1.3, 1.6, 1.8, 1.4). Among subjects with a medical history for chronic bronchitis or asthma, lung cancer risk was associated with neither silicosis (subjects with chronic bronchitis: OR, 0.6; subjects with asthma: OR, 0.4) nor with silica exposure. In this study population, we observed a modest association of both silicosis and cumulative exposure to silica with lung cancer among subjects who were not previously diagnosed with chronic bronchitis or asthma, but not among subjects who had a medical history for either disease. Risk of lung cancer associated with silicosis or cumulative exposure to silica did not vary by previous medical history of pulmonary tuberculosis. PMID- 10874658 TI - Prioritizing back injury risk in hospital employees: application and comparison of different injury rates. AB - To identify high risk areas for back injury in a large teaching hospital, we calculated standard injury rates and newly developed composite statistics for nursing and non-nursing work groups. Data were extracted from the hospital's workers' compensation database. The hospital-wide total injury rate was 4.6 reports per 100 full-time equivalents (FTE); Compensation Case Rate, 1.4 cases per 100 FTE; Compensation Severity Rate, 76 days lost per 100 FTE; and the Cost Rate, $3742 per 100 FTE. The Total Injury Reports Rate for nursing varied from 14.2 per 100 FTE for Intensive Care Unit (ICU) Nursing to 3.8 per 100 FTE for Pediatric Nursing. Non-nursing areas also demonstrated increased rates for back injury. Individual statistical rates ranked areas differently in risk, whereas composite statistical measures consistently ranked ICU Nursing, Buildings and Grounds, and Orthopedics/Neurological Nursing as the top three. Patient handling was the precipitating event in the majority of nursing back injuries, indicating the need for ergonomic intervention. The use of combined statistical measures provided a more integrative measure for describing and following back injury risk over time. PMID- 10874659 TI - Occupational exposures and lung cancer in Buenos Aires, Argentina. AB - The main objective of this study was to analyze the risks associated with occupational exposures in an industrializing country where lung cancer is the primary neoplastic cause of death in men. A full occupational history was collected through interviewing 199 men with lung cancer and 393 control subjects. Exposure to arsenic, asbestos, chromium, dust, nickel, and polynuclear aromatic hydrocarbons was assessed by means of a job-exposure matrix. Elevated odds ratios were observed for employment in the alcoholic beverages industry (5.2; 95% confidence interval [CI], 1.1 to 23.1), sawmills and wood mills (4.8; 95% CI, 1.2 to 19.0), water transport (3.3; 95% CI, 1.1 to 12.1), and chemicals/plastics manufacturers (1.9; 95% CI, 1.1 to 3.3). A small, non-significant increased risk was observed after long-term exposure to arsenic and chromium, with a dose response for chromium. Although some of the present results may result from chance, most are consistent with those of previous investigations in other countries. PMID- 10874660 TI - Chronic low back pain assessment using surface electromyography. AB - This investigation examined surface electromyography as an additional tool in the comprehensive clinical evaluation of patients with chronic low back pain (CLBP). Electromyographic signals from electrodes placed in the lumbar area of 30 CLBP patients and 30 non-pain control subjects were compared. Patients and controls were matched for age, gender, and body mass index. Paired t test showed a statistically significant difference between the two groups. The muscle activity mean values were threefold higher in CLBP patients than in controls (P < 0.00001) in the static testing, and twofold higher in CLBP patients than in controls (P < 0.00001) in the dynamic testing. Our findings indicate that surface electromyography assessment of the paraspinal muscle activity may be a useful objective diagnostic tool in the comprehensive evaluation of CLBP. PMID- 10874661 TI - Stress management in men with solvent-induced chronic toxic encephalopathy. AB - Stress management was studied in male patients with solvent-induced chronic toxic encephalopathy (TE) of types 2A (TE 2A, n = 31) and 2B (TE 2B, n = 26). The patients were compared with a healthy reference group (n = 57). Self-reported symptoms (90-item Symptoms Checklist [SCL-90]), sense of coherence, coping strategies, and level of mastery were measured. As expected, both TE groups reported highly deviating symptoms on most SCL-90 scales. The TE 2B patients, who had objectified cognitive dysfunction, reported more use of passive, less situationally adequate coping strategies; a weaker sense of coherence; and a lower degree of mastery. In contrast, the TE 2A cases showed only minor deviations from the reference group in these respects. The results suggest that having a strong sense of coherence, a sense of mastery, and flexible resources for stress management could be dependent on intact brain functions. PMID- 10874662 TI - T-cell lymphoma presenting in the breast: a histologic, immunophenotypic and molecular genetic study of four cases. AB - Primary non-Hodgkin's lymphoma of the breast is uncommon. Most primary breast lymphomas are of B-cell phenotype, with only rare cases showing a T-cell phenotype. In this study, we report the clinicopathologic features of four cases of T-cell lymphoma in the breast. The patients all were female with a mean age of 48 years (range, 13 to 77 years). All cases showed immunoreactivity in paraffin embedded tissue for T-cell markers CD3, CD45RO, and CD43. beta F1 was positive in three of four cases. The four cases were further subclassified as anaplastic large cell lymphoma (CD30 positive) of T-immunophenotype; natural killer/T-cell lymphoma; peripheral T-cell (CD4 positive), large cell type; and peripheral T cell (CD8 positive, T-cell intracellular antigen positive), medium cell type. Three of the four cases were monoclonal for T-cell receptor beta and/or T-cell receptor gamma. The one case of natural killer/T-cell lymphoma was negative for monoclonality with both T-cell receptor beta and gamma by molecular diagnostic studies. In all cases, IgH was negative. Follow-up was obtained in three cases. Two patients died within less than 1 year after the diagnosis. The third patient died within 18 months of the diagnosis. Our results suggest an aggressive clinical course for T-cell lymphomas that present in the breast. PMID- 10874663 TI - Clonality of precursors of cervical cancer and their genetical links to invasive cancer. AB - Two problems were the focus of this study. (1) Is precancer and/or invasive cancer of the human cervix a poly- or monoclonal proliferation of neoplastic cells? (2) Are simultaneously present precancers and cancers of the cervix clonally related, or do they arise independently? Microdissection of 37 neoplastic lesions with different degrees of histologic severity in 22 patients followed by polymerase chain reaction-based analysis of X-chromosome inactivation was used as a principal method. Invasive cancers were interpreted as monoclonal because samples invariably showed monoclonal signals. In two thirds of these cases, simultaneously present precursors had the identical X-chromosome inactivation pattern, but in one third the pattern was different. Polyclonality was seen in a subgroup of precursors, where there was no simultaneous presence of invasive cancer. In contrast, when invasive cancer was present, no precursor signaled polyclonality. Data taken together indicate that the pathogenesis of cervical cancer is probably even more complicated than that of other cancers involving selection of subclones from originally polyclonal precursors and possibility of coexistence of precursors of different monoclonal composition. The study also observed that a large field of normal cervical squamous epithelium (approximately 500 basal squamous epithelial cells) with nonrandom X-chromosome inactivation was present. It remains to be further investigated whether this phenomenon represents an embryologic lyonization pattern of X-chromosome inactivation or postembryologic clonal expansion of submorphologically transformed cells. PMID- 10874664 TI - The location and frequency of intestinal metaplasia at the esophagogastric junction in 223 consecutive autopsies: implications for patient treatment and preventive strategies in Barrett's esophagus. AB - The frequency of intestinal metaplasia at the esophagogastric junction is as high as 36% in endoscopy studies; the majority of cases (approximately 67%) occur in short segments of esophageal columnar mucosa. The validity of these studies has been questioned, however, because of heterogenous underlying diseases prompting endoscopy. To determine the frequency and origin of intestinal metaplasia at the esophagogastric junction, we histologically evaluated the entire esophagogastric junction for the presence of intestinal metaplasia using Alcian blue/periodic acid-Schiff mucin stains in 223 consecutive autopsies. Precise localization of the Z line in relation to the esophagogastric junction and tongues of esophageal columnar-appearing mucosa were noted in each case. Mean patient age was 47 years; 69% of patients were male, and 63% were white. Twenty five of 223 cases (11%) had intestinal metaplasia at the esophagogastric junction. Only 2 of 25 cases (8%) had intestinal metaplasia in the esophagus; the remaining 23 cases (92%) had intestinal metaplasia in the gastric cardia. Male gender, advanced age, white ethnic origin, and short tongues of esophageal columnar mucosa were not associated with gastric cardia intestinal metaplasia. An association of distal gastric intestinal metaplasia (P < .01) and chronic gastritis (P < .01) with gastric cardia intestinal metaplasia suggests a role for Helicobacter pylori infection in this process. The frequency of intestinal metaplasia at the esophagogastric junction in an unselected autopsy population is low (11%) even after exhaustive histologic evaluation using Alcian blue mucin stains. Furthermore, intestinal metaplasia is confined to the gastric cardia in more than 90% of cases with no association to male gender, white ethnic origin, advanced age, or the presence of short segments of esophageal columnar-appearing mucosa at endoscopy. These results demonstrate that caution is warranted when applying the findings of endoscopy studies to the development of preventive and screening strategies aimed at identifying Barrett's esophagus in an asymptomatic general population. PMID- 10874665 TI - Mdm2 gene amplification in gastric cancer correlation with expression of Mdm2 protein and p53 alterations. AB - Mdm2, localized on chromosome 12, is considered a negative regulator of p53 function and seems to play a role in the pathogenesis of a variety of tumors. The mdm2 amplification in advanced-stage gastric carcinoma has not yet been investigated. Mdm2 amplification was determined in 43 gastric carcinomas, and the genetic results were correlated with mdm2 protein expression, p53 alterations, and clinicopathologic data. The tumors were classified according to Lauren: 20 intestinal-type tumors, 19 tumors of diffuse growth inclusive of a primary small cell carcinoma, and 4 carcinomas with mixed differentiation. Staging was based on the pTNM classification system. Mdm2 and p53 were demonstrated by immunohistology on formalin-fixed and paraffin-embedded tumor tissue. The mdm2 oncogene was amplified by nonradioactive hybridization of tumor DNA with an mdm2 cDNA probe. The Southern blots were evaluated densitometrically. For p53 mutation screening, we analyzed the highly conservative regions of the p53 gene (exons 4 to 8) with the use of the polymerase chain reaction-single-strand conformation polymorphism technique. Polymerase chain reaction products with band shifting were directly sequenced. Mdm2 amplification was demonstrated in 18 tumors (41.8%). The mdm2 gene was amplified more frequently in carcinomas with a diffuse growth pattern. Gastric carcinomas of the intestinal type, however, showed a higher frequency of p53 alterations. There was no statistical significance of the molecular genetic and immunohistologic results of the mdm2/p53 status to staging as well as to age and sex of the patients. The mdm2/p53 pathway is a part of the carcinogenesis of gastric carcinoma. Only approximately 20% of gastric carcinomas failed to show mdm2 and/or p53 alterations. The upregulation of the mdm2 oncogene and the accompanying inactivation of the tumor suppressor gene 53 seem to play a role above all in carcinomas of the diffuse type. PMID- 10874666 TI - Allelotype analysis of intrahepatic cholangiocarcinoma. AB - To identify the chromosomal loci of allelic loss in intrahepatic cholangiocarcinoma (ICC), we performed an allelotype study of 36 ICCs using 55 genome-wide microsatellite markers. Loss of heterozygosity was found most frequently on 8p (65.6%), 17p (64.7%), and 9p (64.5%), followed by 18q (54.2%), 1p (48.5%), 3p (44.8%), 9q (42.1%), 14q (41.7%), 6q (41.7%), and 1q (40.6%). The fractional allelic loss (FAL) values ranged from 0 to 0.731 (mean, 0.322). Analysis of the relationship between FAL values and clinicopathologic parameters disclosed significantly higher FAL values in moderately to poorly differentiated ICCs than in well-differentiated ones (P < .05). In summary, this study defined for the first time the overall number of chromosomes having allelic loss and the chromosomal arms and/or regions potentially involved in the development of ICC. PMID- 10874667 TI - Platelet-derived growth factor-AA and -alpha receptor expression suggests an autocrine and/or paracrine loop in osteosarcoma. AB - Platelet-derived growth factor (PDGF) is a major mitogen and chemotactic factor for mesenchymal cells such as fibroblasts, smooth muscle cells, and osteoblasts. PDGF exists as disulfide-linked homo- or heterodimers composed of two polypeptide chains encoded by distinct genes, designated PDGF-A and PDGF-B. Upon binding to its tyrosine kinase receptor PDGF-alpha, especially PDGF-AA stimulates the proliferation of osteoblastic cells and may exert autocrine and paracrine effects in regulating bone-forming processes. The purpose of this immunohistochemical study was to determine the expression of PDGF-AA and PDGF-alpha receptor in benign and malignant neoplastic bone lesions. Polyclonal antibodies to PDGF-AA and PDGF-alpha receptor were used on paraffin sections of 23 osteosarcomas and 17 osteoblastomas. Immunostaining was assessed quantitatively by evaluating the percentage of reactive tumor cells. In osteosarcomas, the mean expression of PDGF AA and PDGF-alpha receptor was 33.97% (range, 2 to 80%; SD, 24.26%) and 27.13% (range, 3.2 to 72%; SD, 18.38%), respectively. Osteoblastomas showed significantly lower expression of PDGF-AA than osteosarcomas (mean, 15.71%; range, 5 to 34%; SD, 9.43%; P = .019). Although the mean expression of PDGF-alpha receptor in osteoblastomas was much lower than in osteosarcomas (mean, 17.55%; range, 3.6 to 26.8%; SD, 6.47%), the difference was not significant (P = .122). For osteosarcomas, Spearman correlation coefficient (two-tailed) revealed a significant correlation between the expression of PDGF-AA and PDGF-alpha receptor (r = .688), which was not the case for osteoblastomas (r = .267). These data suggest that in contrast to osteoblastoma, the growth of osteosarcoma may be supported by the coordinate expression of the potent mitogenic growth factor and its receptor that exert their functions by autocrine and paracrine mechanisms. PMID- 10874668 TI - Cytokeratin 7 and 20 expression in choroid plexus tumors: utility in differentiating these neoplasms from metastatic carcinomas. AB - Tumors derived from choroid plexus epithelium are uncommon and may exhibit a wide variety of histologic patterns. They often are difficult to distinguish from metastatic carcinomas. Previous studies that addressed this issue yielded conflicting results. Recent reports have demonstrated that evaluation of coordinate expression of cytokeratin (CK) 7 and CK20 aids in distinguishing primary from metastatic lesions in a number of anatomic sites and that tumors that commonly are metastatic to the brain retain their CK7/CK20 immunophenotype in this location. We examined 35 choroid plexus tumors with a panel of antibodies to determine their CK7/CK20 immunophenotype. Tumors from 35 patients (7 male, 28 female; mean age, 25 years), including 31 choroid plexus papillomas and 4 atypical papillomas, were evaluated. All tumors were intraventricular or within the cerebellopontine angle and composed predominantly of orderly columnar epithelial cells resting on distinct fibrovascular cores. Atypical papillomas contained combinations of focal loss of architectural pattern, increased mitotic activity, necrosis, and brain parenchymal invasion. No lesion was unequivocally malignant. Twenty-six tumors (74%), including all atypical papillomas, were CK7 positive and CK20 negative. Two tumors stained with both markers, one stained with CK20 only, and six stained with neither marker. Other findings included expression of glial fibrillary acidic protein in 24 tumors, S-100 protein in 19 tumors, transthyretin in 31 tumors, Ber EP4 in 1 tumor, CAM5.2 in 33 tumors, epithelial membrane antigen in 4 tumors, and pancytokeratin in 27 tumors. Our results indicate that the majority of choroid plexus tumors have a CK7 positive/CK20-negative immunophenotype. This finding may be useful in differentiating these lesions from metastatic carcinomas that have differing CK7/CK20 profiles. PMID- 10874669 TI - "High risk" HPV types are frequently detected in potentially malignant and malignant oral lesions, but not in normal oral mucosa. AB - Studies on the involvement of the human papillomavirus (HPV) in initiation and progression of oral neoplasia have generated conflicting results. The observed discrepancy is attributable mainly to the varying sensitivity of the applied methodologies and to epidemiologic factors of the examined patient groups. To evaluate the role of HPV in oral carcinogenesis, we analyzed 53 potentially neoplastic and neoplastic oral lesions consisting of 29 cases of hyperplasia, 5 cases of dysplasia, and 19 cases of squamous cell carcinomas, as well as 16 oral specimens derived from healthy individuals. A highly sensitive nested polymerase chain reaction (PCR) assay was used, along with type-specific PCR, restriction fragment length polymorphism analysis, dot blotting, and nonisotopic in situ hybridization. Nested PCR revealed the presence of HPV DNA in 48 of the 53 (91%) pathologic samples analyzed, whereas none (0%) of the normal specimens was found to be infected. Positivity for HPV was independent of histology and the smoking habits of the analyzed group of patients. At least one "high risk" type, such as HPV 16, 18, and 33, was detected by type-specific PCR in 47 (98%) infected specimens, whereas only 1 (2%) squamous cell carcinoma was solely infected by a "low risk" type (HPV 6). HPV 16 was the prevailing viral type, being present in 71% of infected cases. Single HPV 16 and HPV 18 infections were confirmed by restriction fragment length polymorphism. HPV 58 was detected by dot blotting in three hyperplastic lesions. HPV positivity and genotyping were further confirmed, and the physical status of this virus was evaluated by nonisotopic in situ hybridization. Diffuse and punctate signals, indicative of the episomal and integrative pattern of HPV infection, were observed for low- and high-risk types, respectively. Our findings are suggestive of an early involvement of high-risk HPV types in oral carcinogenesis. PMID- 10874670 TI - Metastasizing fibrous histiocytoma of the skin: a clinicopathologic and immunohistochemical analysis of three cases. AB - The clinicopathologic and immunohistochemical features of three metastasizing fibrous histiocytomas of the skin are presented. The first patient had a 1.3-cm nodule in the right thigh, with right inguinal lymph node metastases 19 years later. The second patient, who had a 3-cm nodule excised from his left thigh and inguinal lymph node metastasis after 4 months, had a favorable outcome 14 years after local radiotherapy and chemotherapy. The third had a 2-cm nodule in his neck, which recurred 16 months later. Four months later, cervical lymph node metastases were found. The patient was alive and well 26 months after initial surgery. All three primary skin tumors involved the dermis and subcutis, appeared well-delineated but nonencapsulated, were associated with some degree of epidermal hyperplasia, and showed features of aneurysmal/atypical or cellular fibrous histiocytoma. The number of mitoses ranged from 6 to 11 per 10 high-power fields. Recurrences and metastases showed morphologic features similar to primary lesions. Tumor cells were positive, at least focally, for CD 68, Ki-M1p, and Factor XIIIa, and occasionally for smooth muscle actin. Desmin, CD 34, S-100 protein, and cytokeratin stainings were negative. Primary neoplasms, recurrences, and metastases showed a Mib-1 labeling index of 10% or less. Cellular, aneurysmal, and atypical (pseudosarcomatous) fibrous histiocytomas of the skin can metastasize, yet they often show a protracted clinical course. Risk factors for metastatic dissemination include large size, high cellularity, aneurysmal changes, marked cellular pleomorphism, high mitotic activity, tumor necrosis, and repeated local recurrences. PMID- 10874671 TI - 5'-->3' exonuclease-based real-time PCR assays for detecting the t(14;18)(q32;21): a survey of 162 malignant lymphomas and reactive specimens. AB - We describe our experience using two real-time polymerase chain reaction (PCR) assays for detecting the t(14;18)(q32;q21) in a large series of non-Hodgkin's lymphomas (NHLs). These assays utilize the 5'-->3' exonuclease activity of Taq polymerase, which cleaves a probe labeled with a fluorescent reporter dye at its 5' end and a quencher dye at its 3' end during the extension phase of PCR. In a previous study, Luthra and colleagues developed these real-time PCR assays for detecting the t(14;18) involving the major and minor breakpoint cluster regions of the bcl-2 gene and assessed a small number of NHLs. In this larger study, we analyzed 135 NHLs, 6 Hodgkin's disease, 10 reactive biopsy specimens, and 11 peripheral blood specimens. The NHL group included 46 of 70 (65.7%) follicular NHLs, 1 of 2 (50%) diffuse small cleaved cell NHLs, and 13 of 24 (54.2%) diffuse large B-cell NHLs with the t(14;18) detected by conventional PCR methods. There was excellent agreement between the real-time and conventional PCR assays with overall concordance in 160 of 162 (98.8%) specimens. For the NHLs, concordance was found in 134 of 135 (99.3%) specimens. Disagreement was observed in one case of follicular NHL in which the real-time PCR assay detected bcl-2 minor breakpoint cluster region/JH DNA fusion sequences and the conventional method was negative. The overall concordance for 10 benign biopsy specimens and 11 normal peripheral blood samples was 20 of 21 (95.2%). One lymph node biopsy specimen that showed reactive follicular hyperplasia was positive for the bcl-2 minor breakpoint cluster region/JH DNA fusion sequences detected by the real-time PCR assay but was negative by conventional PCR methods. This patient had no clinical evidence of NHL. We conclude that real-time PCR assays for detecting the t(14;18) are sensitive, specific, and more convenient than conventional PCR methods. PMID- 10874672 TI - Evaluation of cyclin expression in testicular germ cell tumors: cyclin E correlates with tumor type, advanced clinical stage, and pulmonary metastasis. AB - The measurement of proliferative index has yielded promising yet conflicting results in the evaluation of testicular tumors. We have examined the role of Ki 67, along with the cyclins A and E in testicular tumorigenesis. We compared the immunoreactivity of 20 pure seminomas with 20 mixed germ cell tumors composed predominantly of embryonal carcinoma with a variety of proliferation markers, including Ki-67, cyclin A, and cyclin E. All 40 tumors stained for Ki-67, and 19 of 20 (95%) seminomas and 18 of 20 (90%) embryonal carcinomas stained positively for cyclin A. Cyclin E stained 14 of 19 (74%) of the embryonal carcinomas and only 4 of 20 (20%) of the seminomas (Fisher's exact two-tailed test, P = .0012). There was a trend toward larger tumor size for cyclin E-positive seminomas (median, 5.92 cm versus 3.96 cm; P = .08), although the same correlation was not significant in embryonal carcinomas. For both seminomas and embryonal carcinomas, staining with cyclin E did not correlate with the presence of lymphovascular invasion or capsular invasion. However, patients who had cyclin E-positive tumors presented with higher clinical stage (P = .0015). In addition, pulmonary spread in embryonal carcinomas (four patients) and seminomas (one patient) occurred only in patients whose tumors were cyclin E positive (P = .014). Although Ki-67 and cyclin A offer little prognostic information in testicular germ cell tumors, cyclin E immunoreactivity correlates with tumor type and is strongly predictive of distant tumor spread. PMID- 10874673 TI - Trichoblastic carcinoma ("malignant trichoblastoma") with lymphatic and hematogenous metastases. AB - We report an aggressively behaving malignant trichogenic tumor arising in a trichoblastoma (TB) with widespread lymphatic and hematogenous metastases in a 55 year-old man with a concomitant B-cell chronic lymphocytic leukemia. The primary tumor had been present and unchanged for as long as 40 years before excision. Typical trichogenic TB with dystrophic calcification and even ossification was still present peripheral to the malignant transformation. The malignant neoplasm consisted of basaloid cells, spindle cells arranged in fascicles and densely packed rounded nests or "cell balls." The metastases consisted of immature basaloid cells and cell balls, and the recurrences became successively more undifferentiated. The residual TB reacted with antibodies to cytokeratin (CK) 6, 8, 14, and 17 and focally to S-100; the malignant primary tumor reacted uniformly with antibodies to vimentin and only focally with antibodies to CK and S-100. The metastatic tumor had lost epidermal CK expression but maintained expression of S 100 in paraffin-embedded tissues. Trichoblastic differentiation was confirmed in frozen tissues with antibodies to hair keratins. No expression of p53 or bcl-2 was identified, but p-glycoprotein (MDR-1 gene related) was expressed by primary and metastatic tumor cells. We believe that this neoplasm is best classified as a trichoblastic carcinoma arising in a TB in association with a B-cell chronic lymphocytic leukemia. This case illustrates that TBs have the potential for malignant transformation and aggressive behavior. PMID- 10874674 TI - Liver pathology: cirrhosis, hepatitis, and primary liver tumors. Update and diagnostic problems. PMID- 10874675 TI - Laser capture microdissection-guided fluorescence in situ hybridization and flow cytometric cell cycle analysis of purified nuclei from paraffin sections. AB - Laser capture microdissection (LCM) has recently been identified as a quick, simple, and effective method by which microdissection of complex tissue specimens for molecular analysis can be routinely performed. Assessment of gene copy number by fluorescence in situ hybridization (FISH) is useful for the analysis of molecular genetic alterations in cancer. Unfortunately, the application of FISH to paraffin sections of tumor specimens is fraught with technical difficulty and potential artifacts. Our results demonstrate that LCM-microdissected nuclei are suitable for FISH gene copy analysis. Amplification of genes in cancer specimens can be detected as easily in LCM-prepared nuclei as in fresh nuclei from cancer tissue specimens. Furthermore, contamination of tumor specimens by normal cells can make interpretation of flow cytometric cell cycle analysis difficult. Our results show that LCM-microdissected nuclei can also be used for flow cytometric cell cycle and ploidy analysis. LCM/FISH offers the advantages of multicolor FISH in a morphologically defined cell population, without the technical problems of FISH performed on paraffin sections. This technique should further simplify the methodology required to perform copy number analysis of tumor suppressor or protooncogenes in archived cancer specimens. The use of LCM specimens will also improve the specificity and simplify the interpretation of flow cytometric cell cycle and ploidy analysis of breast cancer specimens. PMID- 10874676 TI - Correspondence Re: Chibbar R, Leung K, McCormick S, Ritzkalla K, Strickler J, Staggs R, et al. bcl-1 gene rearrangements in mantle cell lymphoma: a comprehensive analysis of 118 cases, including B-5-fixed tissue, by polymerase chain reaction and southern transfer analysis. Mod Pathol 1998;11:1089-97. PMID- 10874677 TI - Correspondence Re: Gutmann EJ. "No pictures from summer vacation": portrayals of pathologists in the printed media. Mod Pathol 1998;11:686-91. PMID- 10874678 TI - Apolipoprotein D in the aging brain and in Alzheimer's dementia. AB - Apolipoprotein D (apoD) levels were examined in the temporal cortex as well as an assessment of the location of apoD positive cells within the brain by immunohistochemical and biochemical methods in young control (YC), aged control (AC), and Alzheimer's demented (AD) probands. Scattered apoD positive astrocytes and oligodendrocytes were found throughout the white matter by immunohistochemistry. ApoD immunoreactivity was also observed in the cerebellar oligodendrocytes of the YC group. There was faint positive apoD staining in scattered cortical astrocytes and a few neurons in the same group. In contrast, some of the AC and all of the AD probands had intense and frequent apoD immunostained cortical astrocytes and pyramidal neurons. The cortical senile plaques and neurofibrillary tangles were apoD immunonegative. No quantitative differences were found between the cortical apoD levels in the AC and AD groups, determined by immunoblotting. ApoD detected in the brain tissue was different in molecular weight (29 kDal) from that seen in CSF or in the serum (32 kDal). Our results indicate apoD is present in the human brain, especially in glial cells, and has increased abundance in the elderly and AD subjects. PMID- 10874679 TI - Middle cerebral artery blood flow velocity in elite power athletes during maximal weight-lifting. AB - Cerebral blood flow velocity (CBFV) has been shown to significantly increase during dynamic exercise (running) secondary to increases in cardiac output. Static exercise (weight-lifting) induces supraphysiological arterial pressures up to 450/380 mmHg, and thus may alter CBFV. Catastrophic brain injuries such as stroke, cerebral hemorrhage, subarachnoid hemorrhage, retinal hemorrhage and retinal detachment have been associated with weight-lifting. A recent study has shown that intra-ocular pressure (IOP), which is an indirect measure of intracranial pressure, elevates to pathophysiologic levels during weight-lifting. Recent CBFV studies instituting Valsalva have demonstrated decreases in CBFV from 21%-52%. To date, no studies have examined CBFV during maximal weight-lifting to elucidate the cerebrovascular responses to extreme pressure alterations. We recruited nine elite power athletes, including a multi-world record holder in powerlifting, for a transcranial Doppler study of middle cerebral artery blood flow velocity at rest and during maximal weight-lifting. All subjects' resting blood flow velocities were within normal ranges (mean 64.4 +/- 9.5 cm sec2). Blood flow velocities were significantly (p < 0.0001) decreased in all subjects during maximal lifting (mean 48.4 +/- 10.1 cm sec2). Linear regression analysis demonstrated a significant inverse linear relationship in the net change of blood velocities from rest to maximal lift for each subject (r = 0.8585, p < 0.001). This study demonstrates that blood flow velocities are significantly decreased during heavy resistance training. The drop in CBFV during weight-lifting was significantly less than previous Valsalva studies, which likely reveals the cardiovascular, baroreflex, and cerebrovascular system adaptations occurring in these elite power athletes. PMID- 10874680 TI - The effect of encephalo-myo-synangiosis on abnormal collateral vessels in childhood moyamoya disease. AB - Child patients with Moyamoya disease initially present with ischemic symptoms. However, the long-term risk of intracranial hemorrhage for childhood Moyamoya disease is unknown. Hemodynamic overload to the fragile collateral vessels has been considered to cause hemorrhage. We reviewed angiograms to evaluate the effect of encephalo-myo-synangiosis (EMS) on abnormally dilated collateral vessels in 13 child patients with Moyamoya disease. EMS was performed on 24 sides in 13 patients ranging from 5 to 14 years of age. Post-operative angiography (6 88 months after surgery) revealed good revascularizations through EMS (larger than one-third of the middle cerebral artery (MCA) distribution) in 18 sides (75%) and smaller revascularizations in 6 sides (25%). In cases with a good revascularization through EMS, reduction of the abnormal collateral vessels was observed not only in the basal Moyamoya vessels (94% of sides) but also in the medullary arteries derived from the choroidal arteries (62% of sides), which are considered to cause intraventricular hemorrhages in adult patients. It is suggested that EMS may reduce the hemodynamic load on dilated collateral vessels and, subsequently, the long-term risk of intracranial hemorrhage in childhood Moyamoya disease. PMID- 10874681 TI - Isolated oculomotor nerve palsy in lymphoma. AB - We report a patient with non-Hodgkin's lymphoma who developed a unilateral left oculomotor nerve palsy. Only eyelid lifting and vertical gaze were involved. Lateral gaze or sizes and light reactions of pupils were not involved. Magnetic resonance imaging revealed an enhancement of an upper part of left cavernous sinus and the posterior clinoid process. It was conceivable that lymphoma invaded the upper branch of oculomotor nerve. Such neurological symptoms in cases of oculomotor nerve palsy by lymphoma have not been reported previously. Because cranial neuropathy could occur as the first sign of lymphoma, lymphoma is an important differential diagnosis for the partial oculomotor palsy such as our present case. PMID- 10874682 TI - Electromagnetic therapeutic angiogenesis: the next step. AB - Therapeutic angiogenesis, in the form of growth factor protein administration or gene therapy, is a new method of treatment for patients with severe coronary and peripheral artery disease not amenable to conventional methods of revascularization. Furthermore, a new experimental strategy increases endogenous angiogenesis in ischemic tissue to induce local 'angiogens' by means of electromagnetic stimulation. Further studies examining the molecular basis and clinical efficacy of electromagnetic angiogenesis are necessary. PMID- 10874683 TI - Extradural meningioma en-plaque of the cervical cord. AB - A rare case of cervical extradural en-plaque meningothelial meningioma is reported. The magnetic resonance imaging revealed an extradural sheet of tumor encasing the cervical cord from anterior, posterior and right lateral aspects and emerging from the right C3-4 intervertebral foramina. Though a differential diagnosis of lymphoma and tubercular granulation tissue were considered, its isointense intensity patterns on T1 and T2 weighted images and the intratumoral calcification on intrathecal contrast computed tomographic scan suggested a meningioma. At surgery, the lesion was fibrous, avascular and densely adherent to the dura. The radiological features and management options of the lesion are discussed. PMID- 10874684 TI - The benefit of neuronavigation for neurosurgery analyzed by its impact on glioblastoma surgery. AB - Neuronavigation, today a routine method in neurosurgery, has not yet been systematically assessed in direct comparison with conventional microsurgical techniques. The aim of the present study was the direct comparison of the impact of neuronavigation on glioblastoma surgery regarding time consumption, extent of tumor removal and survival. For each of 52 patients operated for primary glioblastoma with neuronavigation, a patient operated on without navigation was matched. Completeness of tumor resection, including volumetric analysis, was examined by early post-operative MRI. Operating and survival times were obtained for all patients. At a rate of 86.5%, surgeons' opinions about neuronavigation were positive. Operating times were identical in the two groups, while preparation times were 30.4 min longer with navigation. Radiological radicality was achieved in 31% of navigation cases vs. 19% in conventional operations. The absolute and relative residual tumor volumes were significantly lower with neuronavigation. Radical tumor resection was associated with a highly significant prolongation in survival (median 18.3 vs. 10.3 months, p < 0.0001). Survival was longer in patients operated on using neuronavigation (median 13.4 vs. 11.1 months). Neuronavigation increases radicality in glioblastoma resection without prolonging operating time. Regarding the problem of brain shift, neuronavigation should be optimized by intraoperative real-time imaging. PMID- 10874685 TI - Heterogeneity of delta-aminolevulinic acid-induced protoporphyrin IX fluorescence in human glioma cells and leukemic lymphocytes. AB - Delta-aminolevulinic acid (ALA)-PDT efficacy is particularly dependent on the quality of protoporphyrin IX (PpIX)-induced synthesis. The purpose of this study was to determine the ability of cells from two human cancer types to synthesise PpIX after ALA administration. Biopsies of glioma cells have been obtained from patients with glioblastomas that have or have not been given ALA IV (ex vivo incubation). Peripheral blood lymphocytes, obtained from leukemic patients, have also been ALA-incubated in vitro. In glioma cells, fluorescence heterogeneity was extensive either in ALA infused patients or in ex vivo ALA incubated cells. Mean intensities after 3 h were 110 cts (range 0-340) and 1000 cts (range 0-3600). Similar results were found in leukemic lymphocytes where cell fluorescence varied from 0 to 480 cts with a percentage of fluorescent cells varying with time and from one patient to another. Furthermore, PpIX was not detectable in two patients with CLL. These observations suggest that a marked heterogeneity of ALA uptake and/or PpIX synthesis exists in a given human cancer cell population particularly after systemic administration. Improvements for ALA transformation into PpIX are strongly recommended to ensure the efficacy of ALA/PpIX-PDT. PMID- 10874686 TI - Role of protein kinase C in cerebral vasospasm: past and future. AB - The possible role of protein kinase C in chronic cerebral vasospasm after subarachnoid hemorrhage has been suggested for a decade. Experimental results in vitro or in animal models support that protein kinase C is involved in the prolonged contraction of cerebral arteries similar to cerebral vasospasm. Activation of protein kinase C may interact with other signaling pathways such as myosin-light chain kinase, nitric oxide, intracellular Ca2+, and more recently protein tyrosine kinase and its substrates such as mitogen-activated protein kinase. A protein kinase C network may be activated during cerebral vasospasm. PMID- 10874687 TI - Pharmacological and molecular characterization of glutamate receptors in the MIN6 pancreatic beta-cell line. AB - The MIN6 pancreatic beta-cell line responds to glutamate, alpha-amino-3-hydroxy-5 methyl-4-isoxazolepropionic acid (AMPA) and kainate, but not N-methyl-D-aspartate (NMDA) or 1S,3R-trans-ACPD, with increases in [Ca2+]i. This correlates with MIN6 expression of AMPA receptor subunits (GluR1-4) but only weak expression of NMDA NR2 receptor subunits, as determined by reverse transcriptase polymerase chain reaction (RT-PCR). Pharmacological characterization of the MIN6 AMPA receptors showed that AMPA-triggered [Ca2+]i responses were blocked by GYKI 52466, 6-cyano 7-nitroquinoxaline-2,3-dione (CNQX) and pentobarbital. AMPA-triggered [Ca2+]i responses were also blocked in Na(+)-free medium and by the voltage-sensitive Ca2+ channel antagonist La3+. Unlike cortical neuronal cultures, which show a loss of membrane-associated protein kinase C (PKC) activity and die in response to excitatory amino acid exposure, glutamate was not toxic to MIN6 cells and it did not decrease PKC activity. These studies indicate that MIN6 cells possess Ca(2+)-impermeable AMPA receptors that secondarily allow Ca2+ influx following AMPA-induced depolarization and that, despite elevating [Ca2+]i, AMPA is not toxic to these cells. The effects of glutamate and glutamate receptor antagonists on pancreatic cells needs to be better understood if these compounds are to be used as therapeutic agents to treat stroke. PMID- 10874688 TI - Effect of stimulation of the dorsal aspect of the cervical spinal cord on local cerebral blood flow and EEG in the cat. AB - Currently there is considerable interest in electrical stimulation of the dorsal aspect of the cervical spinal cord as a potentially effective therapy for persistent vegetative patients. The authors assessed change in the local cerebral blood flow (LCBF) and electroencephalogram (EEG) in the cat following spinal cord stimulation (SCS). In 31 adult cats under isoflurane anesthesia, an electrode for SCS was introduced epidurally to the midline of the C2-C3 segment. Stimulation was performed at 25 Hz and 0.1 msec for 30 min. These animals were divided into five groups by the voltage: (1) 2V (n = 7), (2) 4V (n = 7), (3) 6V (n = 7), (4) 4V with intravenous injection of muscarinic cholinergic agents--atropine sulfate (n = 5), and (5) sham-operated control (n = 5) without stimulation. LCBF was measured by laser Doppler flowmetry through bilateral small burr holes at the parietal area during and 60 min after stimulation. At 2V, LCBF increased only during SCS, then returned to the pre-stimulated level, while the increase continued until the end of the experiment at 4V and 6V. The increase in LCBF was not affected by atropine sulfate. EEG showed spike and wave or polyspikes after SCS in two animals of the 6V group, but not in the 2V and 4V groups, and moreover a moderate increase of the background activity at only 4V. The present data suggested that SCS at 4V can provide the appropriate microcirculatory enhancement with less harmful influence which continues to increase 30 min after SCS, although the exact mechanism should be elucidated continuously. Within the limitation of animal experiments, this study could provide the logical basis for determining the condition of SCS. PMID- 10874689 TI - Therapeutic time window in the penumbra during permanent focal ischemia in rats: changes of free fatty acids and glycerophospholipids. AB - To better define a therapeutic time window for reducing the extent of damage in ischemic penumbra, the time courses of changes in the glycerophospholipid and free fatty acid (FFA) levels were determined in the rat cerebral cortex following induction of the permanent focal ischemia. Focal ischemia induced a biphasic increase in FFA levels in the cerebral cortex, which had been recognized as the ischemic penumbra during the early stages after permanent occlusion of the middle cerebral artery (MCA). The first increase in FFA levels, in which the polyunsaturated fatty acid (PUFA) contained a large number of arachidonic acid (C20:4) molecules, began at 30 min and reached a peak at 1 h, followed by transient return to each sham level 2-6 h after the onset of MCA occlusion. Thereafter, the delayed increase in FFA levels, showing more increases of docosahexaenoic acid (C22:6) molecules than the C20:4 in PUFA compositions, occurred at 24 h. In contrast, the levels of phosphatidylinositol 4-phosphate (PIP) and phosphatidylinositol 4,5-bisphosphate (PIP2) decreased rapidly at 30 min of ischemia and returned transiently to each sham level at 1-6 h. The levels of phosphatidylcholine (PC) and phosphatidylethanolamine (PE), including polyphosphoinositides (PIPs), began to decrease significantly during the late stages, i.e., 24 h after induction of ischemia. These results suggest that the time-dependent changes in FFA and PIPs levels during the early stages of ischemia (until 6 h after induction) might be an important determinant of the subsequent neuronal death in the ischemic penumbra and that the breakdown of glycerophospholipids in the later stages after the induction of focal ischemia was associated with the development of infarction in the cerebral cortex. PMID- 10874690 TI - Thyrotropin-releasing hormone and its receptor in the cerebellum of inferior olive destroyed rat brain. AB - To study the pathophysiology of olivopontocerebellar atrophy (OPCA), we destroyed inferior olive nuclei of male Wistar rats using 3-acetyl pyridine (3-AP) + harmaline + niacinamide. These rats showed a sluggish and ataxic gait. To elucidate the relationship between thyrotropin releasing hormone (TRH) in the Purkinje cell of cerebellum and the inferior olive nucleus, we investigated the concentrations of TRH in the cerebellar cortex, nuclei, and medulla oblongata including the inferior olive nuclei using radioimmunoassay method as well as TRH receptor in the Purkinje cells of cerebellum using immunohistochemical method. All statistical comparisons were done using non-parametric tests (Mann-Whitney U test). We found that two weeks after the treatment, TRH concentrations in the cerebellar cortex as well as nuclei were significantly lower than in the controls but no significant difference in the medulla oblongata was observed between 3-AP treated rats and controls. Moreover, four weeks after the treatment, TRH-receptor positive Purkinje cell counts were significantly fewer than in the controls. These results suggest that TRH in the Purkinje cell of cerebellum may play a role in the ataxic gait observed in the rats whose inferior olive were destroyed. PMID- 10874691 TI - The effect of hyperbaric hyperoxia on brain function in the newborn dog in vivo. AB - Age is a natural factor that has been found to significantly affect sensitivity to hyperbaric hyperoxia (HBO). Exposure to HBO may lead to damages in the energy metabolism of the brain cells. The aim of this study was to test the effect of HBO on the metabolic, hemodynamic and electrical activities in the newborn dog. The study was performed using one-day- to 70-day-old puppies. The puppies were placed in a pressure chamber. The pressure of pure O2 in the chamber was raised by 5 atmospheres (ATA, 75 psi = 6 ATA) within 10 min. The first biochemical change to take place during HBO was oxidation of mitochondrial NADH. The age of the puppy was found to affect the time to the initiation of seizures. In the puppies under the age of 24 days, the average time was 35.1 +/- 5.9 min. In the puppies of 24 days old and older, the average time was 5.1 +/- 0.8 min. In the younger puppies, there was a later occurrence of blood vessel contractions and a longer life span compared to the older puppies. The comparison between the puppies of different ages during exposure to HBO showed differences in the metabolic response, hemodynamic changes and electrical activity. These differences can partially explain the higher resistance in the younger puppies to HBO. PMID- 10874692 TI - Temporal changes in expression of neuronal nitric oxide synthase mRNA in the rat hippocampus associated with kainate-induced seizures. AB - We studied the temporal changes in expression of neuronal nitric oxide (NO) synthase (nNOS) mRNA in the hippocampus of rats treated with kainic acid by use of in situ hybridization technique. Intraperitoneal injection of 10 mg kg-1 kainic acid decreased expression of nNOS mRNAs in the dentate gyrus and CA3 region of the hippocampus at 3 h and 8 h and increased it in the dentate gyrus and CA1 at one week after treatment. Although our previous study indicated that administration of kainic acid increased NO generation in the rat hippocampus, present results suggest that the injection of kainic acid results in differential regulation of nNOS mRNA and NO formation in the rat hippocampus. PMID- 10874693 TI - Elevation of mRNA levels of tissue-type plasminogen activator and urokinase-type plasminogen activator in hippocampus and cerebral cortex following middle cerebral artery occlusion in rats. AB - Tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA) have been used for thrombolitic therapy. In contrast, it is suggested that these compounds might be involved in neuronal cell damage. The activation and the function of tPA and uPA are less understood in ischemic brain tissue. Therefore, changes in tPA and uPA mRNA in rat brain tissue after MCA occlusion and in the neuronal cell line, PC12 cells, during the hypoxic stimulation were examined. Permanent middle cerebral artery (MCA) occlusion was induced by advancing a filament into the internal carotid artery in 36 adult male Sprague-Dawley rats. The ischemic cerebral cortex and contralateral cortex of MCA area, and bilateral hippocampus were collected at 0 (controls), 1, 3, 6, 12 and 24 h after occlusion. Hypoxia was induced in PC12 cells with a multigas incubator (set to 1% O2). The quantitative reverse transcription-polymerase chain reaction acted as a measurement of alteration in mRNA levels. The mRNA levels of tPA and uPA were significantly increased after MCA occlusion in the ischemic cerebral cortex. The magnitude of the increase in tPA and uPA mRNA in 24 h after occlusion was twice the value in sham-operated rat (0 h). The increases of tPA mRNA were time dependent in insult and contralateral hippocampus. The increase of uPA mRNA was also seen in the hippocampus bilaterally, although the increase was more significant on the ipsilateral side. In PC12 cells, necrotic (approximately 35%) and apoptotic cells (approximately 65%) could be distinguished by hypoxic stimulus for 24 h, and the mRNA for tPA was significantly increased for 6 h-12 h, while the mRNA for uPA was not detected at any point in the study. Our results suggest that focal ischemia might result in the activation of these proteases not only in the insult but also in the contralateral brain tissue. PMID- 10874694 TI - Histopathological examination of chemo-sympathectomy in cats. AB - In recent decades, there has been an increase in both the number of sympathectomy techniques, as well as the surgical findings of sympathetic anatomy. Currently the advanced technique of C-arm guided percutaneous thoracic chemo-sympathectomy is widely used for the treatment of palmar hyperhidrosis. However, a better understanding of chemical agents in sympathectomy is required. In this study, chemo-sympathectomy was performed in cats, using alcohol, glycerol and various concentrations of phenol, to determine the chronic neurotoxic effects of these chemical agents on the stellate ganglia. The stellate ganglia of 24 cats were exposed under endotracheal general anesthesia, then injected with about 0.02 ml of absolute alcohol, glycerol and phenol (10%, 25%, 50%, and 75% concentration) solutions, respectively. The stellate ganglia were taken for histological examination three weeks after the chemical injection. The results showed that the degenerative changes in the cytoplasm and nucleus of ganglionic cells and intercellular tissue were moderate and relatively moderate after the injection of alcohol and glycerol, respectively. Meanwhile, the stellate ganglia revealed mild, relatively moderate, serious and extremely serious degeneration after injection of 10%, 25%, 50%, and 75% phenol, respectively. In conclusion, we recommend a high concentration of phenol, in the least volume, as a chemical agent for clinical injection in the upper thoracic sympathetic ganglion. PMID- 10874695 TI - Metabolic changes after injection of quinolinic acid or 6-hydroxydopamine in the rat striatum: a time-course study using cytochrome oxidase and glycogene phosphorylase a histochemistry. AB - Injection of excitotoxins, such as quinolinic acid (QA), into the striatum has been extensively used as an experimental model of Huntington's disease, while injection of 6-hydroxydopamine (6-OHDA) into the dopaminergic nigrostriatal pathway provides a well established model of Parkinson's disease. In the present study, we have examined the metabolic changes induced by an intrastriatal injection of QA or 6-OHDA using histochemical staining for the metabolic markers cytochrome oxidase (COx) and active glycogene phosphorylase (GPa). Intrastriatal injection of QA produced major changes in COx (decrease of staining) and GPa (increase of staining, except in the core of the lesion where the staining was virtually absent) histochemistry at the level of the striatum and of most of the other basal ganglia nuclei. Although attenuated over time, these changes persisted up to one year after the lesion. On the contrary, after the intrastriatal injection of 6-OHDA (which induces only a partial lesion of the nigrostriatal pathway), we did not observe any remarkable changes in COx or GPa staining. This study illustrates the discrepancies between the morphological changes and metabolic changes that are induced when using these experimental models of neurodegenerative disorders. PMID- 10874696 TI - Nuclear medicine imaging and drug delivery. PMID- 10874697 TI - [123I] beta-CIT binding and SPET compared with clinical diagnosis in parkinsonism. AB - The largest group of neurodegenerative disorders are extrapyramidal diseases, especially parkinsonism. The development of the cocaine derivative [123I] beta CIT and single photon emission tomography (SPET) may help in the diagnosis of these patients. The aim of this study was to demonstrate the diagnostic value of this method and its relationship with clinical data. Ninety-eight individuals were investigated: 11 healthy volunteers, 58 patients with idiopathic Parkinson's disease (IPD) and 29 patients with symptomatic parkinsonism (SPD). All patients with parkinsonism were staged according to the clinical classification of Hoehn and Yahr. [123I] beta-CIT was injected intravenously and a triple-headed camera was used to obtain images 20 h later. The images were evaluated visually and semi quantitatively to obtain comparable values (ratio: specific to non-displaceable binding). The ratios differed significantly between controls and IPD patients. A significant correlation also existed between the ratios and clinical stages. In 11 hemiparkinsonian patients, a significantly diminished ratio was demonstrated not only contralateral to the affected side, but also in the clinically silent striatum. A clinical threshold at a reduction of 34% [123I] beta-CIT binding was calculated in this group. The ratios of all SPD patients in our study did not differ significantly from those of the healthy volunteers. According to the clinical degree of symptoms, the more severe subgroup showed a diminished mean ratio of 22% and therefore could not be clearly differentiated from mild IPD. In contrast, ratios were significantly different when comparing groups of the same clinical severity. We conclude that this method is not only a powerful diagnostic tool in IPD patients, but it is also possible to differentiate between IPD and SPD patients, if clinical aspects are also included. PMID- 10874698 TI - Early serial SPET in acute middle cerebral artery infarction. AB - The size and severity of perfusion defects in acute cerebral ischaemia on single photon emission tomographic (SPET) images may provide useful information regarding long-term (> 3 month) stroke outcome. A decreased predictive value has been reported with delayed SPET more than 24 h after stroke onset. We examined 20 patients with acute middle cerebral artery (MCA) infarctions using serial 99Tcm ECD or 99Tcm-HMPAO SPET (SPET 1 one day and SPET 2 three days after stroke onset). Neurological (NIH, SSS) and functional (Barthel, Rankin) scores were calculated simultaneously and 3 months poststroke. The two SPET scans correlated equally well with the severity of functional and neurological deficits evaluated 3 months after stroke onset. In comparison to clinical assessment, the prognostic value of SPET was relatively better on the first day than the third day. Crossed cerebellar diaschisis correlated with early SPET deficits, but did not predict functional outcome. Our results suggest that SPET, either with 99Tcm-ECD or 99Tcm HMPAO, can be used to predict stroke outcome in acute MCA infarction up to 72 h poststroke without significant interference from luxury perfusion. PMID- 10874699 TI - Comparison of 201Tl SPET and treadmill exercise testing in patients with Kawasaki disease. AB - About 4% of children with Kawasaki disease ultimately develop ischaemic heart disease. Therefore, the early detection, non-invasive monitoring and long-term follow-up of myocardial ischaemia are essential. We compared the sensitivity and specificity of 201Tl single photon emission tomography (SPET) and treadmill exercise in the detection of myocardial ischaemia in 23 patients (19 boys, 4 girls) with Kawasaki disease. They were divided into two groups according to the results of coronary angiography. Group I consisted of 11 patients with coronary abnormalities; Group II consisted of 12 patients with no coronary abnormalities. The sensitivity, specificity, false-positive and false-negative rates for detecting coronary arterial lesions were 72.7% (8/11), 58.3% (7/12), 38.5% (5/13) and 30% (3/10) for 201Tl SPET, and 45.5% (5/11), 100% (12/12), 0% (0/5) and 33.3% (6/18) for treadmill exercise, respectively. We conclude that 201Tl SPET is more sensitive than treadmill exercise for the detection of coronary arterial abnormalities, but that the specificity of treadmill exercise is better than that of 201Tl scintigraphy. Coronary artery lesions detected by coronary angiography have good concordance of ischaemic areas with perfusion defects detected by 201Tl SPET. When ischaemic findings on 201Tl SPET and/or positive treadmill exercise testing are noted, coronary angiography is strongly indicated to detect possible stenotic lesions in the coronary arteries. PMID- 10874700 TI - No-carrier-added 123I-MIBG: an initial clinical study in patients with phaeochromocytoma. AB - Radioiodinated meta-iodobenzylguanidine (MIBG) is used routinely for imaging and targeted radiotherapy of tumours derived from the neural crest. Since active uptake of MIBG by the noradrenaline transporter (NAT) makes a greater contribution to total drug accumulation than passive uptake when MIBG is present at low concentrations, tumour-specific uptake should be enhanced by the administration of lower molar amounts of MIBG. This could be achieved through the use of MIBG with a high specific activity. Commercially available preparations of 123I-MIBG have specific activities of approximately 200 MBq.mg-1. We have synthesized and used no-carrier-added (n.c.a.) 123I-MIBG produced by an iododesilylation reaction (specific activity 0.7 TBq.mg-1). We report here the first clinical studies comparing the commercially available and n.c.a. MIBG diagnostic preparations. Five patients with known phaeochromocytoma were studied. Unlike studies in animal models, no consistent improvement in tumour uptake was observed with the n.c.a. material. A larger patient group is required to determine whether there are significant differences between the two preparations, before proceeding to studies at therapeutic activity levels of n.c.a. 131I-MIBG. Even with no improvement in tumour uptake, n.c.a. MIBG may be the favoured formulation for therapeutic applications to reduce the molar amount of drug injected. PMID- 10874701 TI - Normalized spleen/liver ratios on 111In-labelled platelet scintigraphy to predict the outcome of partial splenic embolization in patients with idiopathic thrombocytopenic purpura. AB - In this study, we examined the use of 111In-labelled platelet imaging to predict the outcome of partial splenic embolization (PSE) in patients with idiopathic thrombocytopenic purpura (ITP). Thirty-eight patients with a clinical diagnosis of ITP underwent 111In-labelled platelet scintigraphy. Twenty-four patients with intractable ITP underwent PSE after 111In-labelled platelet scintigraphy. The conventional spleen/liver ratio at 1 h and 192 h and the normalized spleen/liver ratio [(spleen uptake at 192 h/liver uptake at 192 h)/(spleen uptake at 1 h/liver uptake at 1 h)] were compared between responders and non-responders to PSE. Patients with ITP showed a significant reduction in platelet counts, increased platelet associated IgG, decreased platelet survival, and an increased conventional spleen/liver ratio at 192 h. No significant difference was found between patients who had and who had not undergone previous medical treatment. A significant difference was observed in the mean conventional spleen/liver ratio at 192 h between responders and non-responders, but there was substantial overlap among individuals. The mean normalized spleen/liver ratio was significantly higher in responders than non-responders; there was less overlap between the two groups with the normalized spleen/liver ratio than the conventional spleen/liver ratio. The therapeutic outcome of PSE is predicted more accurately using a normalized spleen/liver uptake ratio of 111In-labelled platelets in patients with idiopathic thrombocytopenic purpura than a conventional splenic/hepatic uptake ratio on delayed images. PMID- 10874702 TI - Inter-observer reproducibility of relative 99Tcm-DMSA uptake. AB - 99Tcm-DMSA planar images of 49 randomly selected patients (10 adults, 39 children) were sent to 15 physicians at various centres in Belgium. They were asked to calculate, using their own routine program, the relative uptake (expressed as a percentage) of each kidney. The data were sent on disks formatted so that they could be read by all participants, using their own computer systems. For each scan, the inter-observer variability was expressed using the maximum difference and the standard deviation of left renal uptake. Left renal uptake measured by the 15 observers in the 49 patients was 29.0-72.0% (mean +/- s = 49.8 +/- 6.4%). The maximum differences in left renal uptake ranged between 1.7% and 12.0% (4.5 +/- 2.6%); however, the maximum difference did not exceed 8% in about 90% of the patients. The standard deviations of the individual left renal uptake were between 0.6 and 3.9 (1.3 +/- 0.8). The standard deviations were significantly higher in adults (mean standard deviation = 2.05) than in children (mean standard deviation = 1.12) (P < 0.001); this was probably related to the high background observed in three adults with severe renal impairment. Indeed there was a significant correlation (P < 0.001) between the standard deviation and both the signal-to-noise ratio and the degree of asymmetry between the right and left kidneys. The differences between right and left kidney uptake were systematically lower for some observers, suggesting an influence of the calculation programs. PMID- 10874703 TI - How good is the slope on the second exponential for estimating 51Cr-EDTA renal clearance? A Monte Carlo simulation. AB - It has been suggested that the slope of the second exponential of the plasma disappearance curve may be used to monitor changes in renal function instead of plasma clearance calculated using the slope-intercept method. The purpose of this study was to evaluate the magnitude of error in the slope induced by errors in sampling time and in activity measurement, and to compare it with the error observed in clearance. A model of mono-exponential curves based on two blood samples, taken at 120 and 240 min, was created. Normally distributed random errors were introduced into the sampling times and activity measurements. For each setting, the random errors were successively introduced 200 times and the coefficients of variation of the calculated slopes and clearances were determined. Variable errors in slope and clearance were induced by errors in sampling time and activity measurement. In general, the observed errors in the slope were high in the case of low slope values, decreasing progressively for increasing slope values. The errors in clearance followed a different pattern: highest errors were observed in the case of very low clearance, decreasing progressively for higher clearance values and attaining the minimal value at a lambda around 0.006 min-1, which corresponds to clearance of about 90 ml.min-1. The magnitude of the errors then started to increase again for higher clearance. For a large range of clearance values, the errors in the slope were higher than the errors in clearance. The only exceptions were cases with very high clearance rates. In conclusion, clearance calculation using the slope-intercept method should be preferred to that using the slope alone for monitoring changes in renal function. PMID- 10874704 TI - Evaluation of radioiodinated histamine as isotope carrier in vivo. AB - The iodo derivative of histamine labelled with 125I has been used for many years to prepare tracers used in RIA systems. The aim of this study was to evaluate radioiodinated histamine as a potential isotope carrier for in vivo applications. The biological behaviour of radioiodinated histamine has been investigated in rodents. The observed absence of any specific iodohistamine uptake by a critical organ or tissue promises a very quick distribution of the iodohistamine in soft tissues, and a rapid rate of whole-body clearance via the urinary tract (e.g. over 50% of the injected dose (ID) during the first hour after administration). In spite of moderately low in vitro stability of iodohistamine in serum, biodistribution studies in rodents have not shown any significant release of iodine from the parent molecule in the whole animal. Low uptake was observed in the thyroid (e.g. 0.22 and 0.11% ID at 1 and 2 h after administration to rats), and not more than 3% of injected activity was detected in the stomach in all of the biodistribution experiments. Moreover, our results refute any possibility of competition between histamine and iodohistamine for receptor binding sites, and suggest that radioactive mono-iodohistamine may be used successfully to develop some new radiolabelled bioactive molecules with potential application in vivo. PMID- 10874705 TI - 99Tcm-tetrofosmin: evaluation of fractionated cold kits and two new methods of quality control. AB - The fractionation of Myoview into 1:5 was followed by storage at -80 degrees C for 3 months. The kits were reconstituted with 3.5 GBq 99Tcm-pertechnetate and the average radiochemical purity (RCP) was maintained at 96.5 +/- 1.6% (n = 24) for 24 h. There was no difference in biodistribution between the 1:5 fraction and the full kit, especially with respect to myocardial uptake. These observations show that it is possible to fractionate and store Myoview kits at -80 degrees C and produce a product that is stable over 3 months. This provides a cost effective method for using 99Tcm-tetrofosmin. Two new methods of determining RCP were also examined and found to give results that were not significantly different from the recommended manufacturer's method. A mini paper chromatography (MPC) method using Whatman 17 paper and a solvent mixture of ethyl acetate: acetone (1:9) and an Amprep SAX cartridge method using absolute ethanol as the solvent were found to be more rapid and reliable methods of quality control for this product. PMID- 10874706 TI - Simulating Dictyocaulus viviparus infection in calves: the parasitic phase. AB - A model simulating Dictyocaulus viviparus infection in calves is described. The present paper only deals with the parasitic phase of the life-cycle. Descriptions are given for establishment, development rate of juvenile stages, mortality rates of both juvenile and adult stages, and fecundity. Literature data were used to to develop parameter functions and to estimate initial values for constants. Development of acquired immunity, defined as the proportional ability of the host to reduce the number of parasite individuals in some stage or about to move into a next stage, against establishment (protection) or affecting mortality rates of juvenile or adult parasite stages has been included. The effect of immunity on one parameter or process is viewed as distinctly separate from the effect on another. Preliminary comparisons between model prediction and observations gives encouraging results, indicating that the model simulates experimental D. viviparus infection in calves reasonably well. Some quantitative discrepancies between prediction and observation make clear however, that not all parts of the model are accurate. Further experimentation is needed to re-evaluate current model description and to improve model simulation. PMID- 10874707 TI - Vaccine development and diagnostics of Dictyocaulus viviparus. AB - Parasitic bronchitis is a serious disease of cattle and is caused by the nematode, Dictyocaulus viviparus. For over 30 years, a radiation-attenuated larval vaccine has been used for prevention of this disease. This vaccine has been used with considerable success in the UK and parts of Western Europe, however, it has several disadvantages. It has a short shelf-life and the vaccine has to be produced annually necessitating the use of donor calves. Following vaccination, calves must receive further boosting from natural challenge to maintain protective immunity. Sales of the irradiated larval vaccine have decreased dramatically since the 1970s. This is thought to be due to increased reliance of farmers on anthelmintic programmes to control lungworm infection. It is possible that, under certain circumstances, these programmes do not allow sufficient parasite exposure to stimulate protective immunity to further Dictyocaulus challenge. This is borne out by the recent documented increase in the number of outbreaks of parasitic bronchitis in the UK. A stable vaccine against D. viviparus that is capable of stimulating a more prolonged immunity would be beneficial. Recent research has been directed at identification and isolation of components thought to be involved in parasite survival in the host and examination of their potential as vaccine candidates. One of these components is acetylcholinesterase (AChE), an enzyme secreted by adult worms. This review describes the development of the secreted AChE as a vaccine candidate, as well as documenting recent developments in the immunodiagnosis of D. viviparus. PMID- 10874708 TI - The immune response and the evaluation of acquired immunity against gastrointestinal nematodes in cattle: a review. AB - The present review discusses the immune responses to gastrointestinal nematodes in cattle and the different immunological and parasitological parameters used to assess acquired immunity. Measuring acquired immunity to gastrointestinal nematodes in cattle (e.g. for the evaluation of candidate parasite vaccines) is hampered by the limited understanding of bovine immune responses against gastrointestinal parasites. In this paper the available data on protective immunity against gastrointestinal nematodes, and especially Ostertagia ostertagi, in cattle are compared with the current knowledge of protective immune responses against gastrointestinal nematodes in rodent models and small ruminants. In contrast to the immune response in mice, which is controlled by T helper 2 (Th2) lymphocytes and results in mast cell- or goblet cell-mediated expulsion of adult worms, bovine immune responses to O. ostertagi do not show a clear Th2 cytokine profile, nor do they result in rapid expulsion of the parasite. The first manifestation of immunity to O. ostertagi in calves is a reduction of worm fecundity, possibly regulated by the local IgA response. Worm numbers are only reduced after a prolonged period of host-parasite contact, and there are indications that O. ostertagi actively suppresses the host's immune response. Until the mechanisms of protective immunity against O. ostertagi are revealed, the use of immunological parameters to estimate acquired immunity in cattle is based on their correlation with parasitological parameters and on extrapolation from rodent and small ruminant models. Assessing the resistance of calves against a challenge infection by means of parasitological parameters is probably still the most accurate way to measure acquired immunity against gastrointestinal nematodes. PMID- 10874709 TI - Development of vaccines against gastrointestinal nematodes. AB - Vaccination against complex metazoan parasites has become a reality with the development and registration of recombinant protein-based vaccines against the cattle tick Boophilus microplus and the sheep cestode Taenia ovis. Progress towards the development of similar vaccines against gastrointestinal nematodes, primarily of ruminants, is outlined within a framework of defining the practical requirements for successful vaccination, antigen selection, recombinant protein production and antigen delivery, be it mucosal delivery or DNA vaccination. Antigen selection strategies include the fractionation of complex, but protective, parasite extracts, the use of antibody probes, evaluation of excretory-secretory components and gut-expressed hidden antigens as well as antigens targeted on the basis of function such as enzyme activity. The difficulties being encountered in recombinant protein production and their solution are discussed as are the requirements for successful antigen delivery. Recent technological developments such as the use of functional genomics to identify new vaccine candidates and DNA vaccination to present the selected antigen to the host immune system are discussed and are anticipated to have a profound effect on vaccine development in the future. PMID- 10874710 TI - Immunological responses of sheep to Haemonchus contortus. AB - Infections with Haemonchus contortus are a major constraint on ruminant health world-wide. Young lambs are very sensitive to Haemonchus infection. Older lambs and sheep acquire immunity after a continuous or seasonal exposure to the parasite. The mechanisms underlying immunity are still not completely understood. Antibodies, in particular local IgA and IgE, certainly play a role. The role of IgG is less clear. Lymphocyte proliferation responses seem to correlate to immunity. Sheep that have high antigen-induced lymphocyte responses have a low susceptibility to infection. Furthermore, several studies have demonstrated that immunity against H. contortus is associated with mastocytosis and hyper sensitivity reactions. More recently, increasing attention is being paid to the role of cytokines (interleukins and gamma-interferon) in the activation of specific defence mechanisms. Reverse transcriptase-polymerase chain reaction (RT PCR) assays to study cytokine mRNA expression have become available. The inability of young lambs to mount a significant Th2 response, which is normally characterized by high IgE levels, mastocytosis and eosinophilia, may account for the phenomenon of unresponsiveness in these animals. PMID- 10874711 TI - Impact of nutrition on the pathophysiology of bovine trypanosomiasis. AB - Trypanosomiasis is a major veterinary problem over much of sub-Saharan Africa and is frequently associated with under-nutrition. There is growing evidence that nutrition can have a profound effect on the pathophysiological features of animal trypanosomiasis. These features include anaemia, pyrexia, body weight changes, reduced feed intake and diminished productivity including reduced draught work output, milk yield and reproductive capacity. Anaemia is a principal characteristic of trypanosomiasis and the rate at which it develops is influenced by both protein and energy intakes. Pyrexia is associated with increased energy demands for maintenance which is ultimately manifested by reductions in voluntary activity levels and productivity. Weight changes in trypanosomiasis are markedly influenced by the levels of protein intake. High intakes allow infected animals to grow at the same rate as uninfected controls providing energy intake is adequate whilst low energy levels can exacerbate the adverse effects of trypanosomiasis on body weight. Reductions in feed intake are less apparent in animals which are provided with high protein diets and where intake is limited by the disease animals will often exhibit preferential selection of higher quality browse. Further studies are required to evaluate the minimum levels of protein and energy supplementation required to ameliorate the adverse effect of trypanosomiasis, the nature and quality of protein supplement to achieve these benefits and the influence these have on digestive physiology. PMID- 10874712 TI - Electrophysiological investigation of anthelmintic resistance. AB - It is pointed out that two of the three major groups of anthelmintic act by opening membrane ion-channels. It is appropriate, therefore, to use electrophysiological methods to study the properties of the sites of action of these drugs and the changes in the properties of these receptor sites associated with resistance. This paper describes the use of the patch-clamp technique to observe the currents that flow through the levamisole-activated channels as they open and close in levamisole-sensitive and levamisole-resistant isolates. It was found that, on average, the proportion of time the channels are open, is less in the resistant isolate. The patch-clamp technique also showed that the ion channels are heterogeneous and that one of the subtypes is lost with the appearance of resistance. The use of the current clamp technique is illustrated to record a site of action of ivermectin in the pharyngeal muscle of Ascaris. PMID- 10874713 TI - The development of anthelmintic resistance in sheep nematodes. AB - Anthelmintic resistance now poses problems to sheep farmers throughout the world. In some Southern hemisphere countries multiple resistance has reached levels which make sheep farming non-sustainable. Evidence from studies in the UK and Europe suggests (a) that the selection process occurs over a longer time frame than in Southern tropical/temperate regions and (b) that for some of the key ovine species little or no reversion to susceptibility may occur for many years after the withdrawal of the selecting agent. The dynamics of the selection process are influenced by a number of host, parasite, drug, management and environment-dependent factors. Recent mechanistic studies on resistance against avermectins and milbemycins (AM) suggest that there may be a number of mechanisms associated with resistance at the different target sites for these drugs. Within Europe endectocides within the AM drug group have now become the crucial element in strategies aimed at controlling important diseases such as sheep scab and nematodoses. Given that there is little likelihood of a series of novel action compounds emerging in the immediate future to replace this family the conservation of efficacy of the AM group should be accorded the highest priority for research in this area. PMID- 10874715 TI - Prospects for controlling animal parasitic nematodes by predacious micro fungi. AB - Resistance against anthelmintics is widespread, particularly in parasitic nematode populations of small ruminants. Several new techniques or supplements have been developed or are under investigation. Biological control (BC) is one of these new methods. The net-trapping predacious fungus Duddingtonia flagrans produces thick walled resting spores, chlamydospores, which are able to survive passage through the gastrointestinal tract of cattle, horses, sheep and pigs. Under Danish climatic conditions it has been shown that the number of parasite larvae on pasture and the worm burden of the grazing animals is significantly reduced when animals are fed spores during the initial 2-3 months of the grazing season. Work with D. flagrans in France, Australia, USA, and Mexico has confirmed the strong BC potential of this fungus. Today much work is going into development of suitable delivery systems for grazing livestock worldwide. Ultimately, BC should be implemented in integrated parasite control strategies, both in conventional and organic livestock production. PMID- 10874714 TI - Value of present diagnostic methods for gastrointestinal nematode infections in ruminants. AB - In this paper the different options for the diagnosis of gastrointestinal nematode infections are discussed. Diagnostic tests have a role in confirming the clinical diagnosis of parasitic gastroenteritis, but are more important for herd health monitoring of nematode infections, in particular for cattle. Therefore, emphasis is placed on discussing the available diagnostic parameters on their usefulness for that purpose. For clinical diagnosis the clinical signs, combined with the history of the animals is usually sufficient and a laboratory confirmation is not required. Faecal egg counts are, with two exceptions, not suitable for confirmation of the clinical diagnosis, because correlation between faecal egg counts and infection levels is usually low. These exceptions are the diagnosis of haemonchosis in small ruminants and the detection of anthelmintic resistance. This also limits the value of DNA-based tests of faecal material; even quantitative tests of nematode species specific DNA will have little value for diagnosis and monitoring. Pasture larval counts and worm counts are useful parameters for basic epidemiological studies on nematode infections. However, they are too laborious to be used for either routine diagnosis or monitoring. Blood parameters, such as gastrin and pepsinogen and serology are valuable tools for diagnosis. Pepsinogen and ELISAs based on recombinant proteins show most promise as parameters for herd health monitoring. However, extensive epidemiological studies are still needed before these parameters can be implemented in routine herd health monitoring schemes for parasitic gastroenteritis. PMID- 10874716 TI - Onchocerca ochengi infections in cattle as a model for human onchocerciasis: recent developments. AB - The bovine parasite Onchocerca ochengi is a nodule-dwelling filarial nematode, closely related to O. volvulus, the causal agent of human River Blindness, and, sharing with it, the same vector. This brief review, based on a presentation at the BSP Autumn Symposium 1999, describes recent work supported by the WHO Drug Development Research Macrofil programme and the Edna McConnell Clark Foundation vaccine development programme, to research the chemotherapy and immunology of onchocerciasis utilising this model system, with experimental infections in Liverpool and field infections in northern Cameroon. In a series of chemotherapeutic trials involving 10 compounds in 20 treatment regimes, the comparability of drug efficacy against O. ochengi with that described against O. volvulus has been demonstrated. Repeated, long-term treatment with oxytetracycline has been shown to be macrofilaricidal and the effect is hypothesized to be related to action on Wolbachia endobacteria, abundant in O. ochengi. Avermectins/milbemycins are not macrofilaricidal (even in high and repeated long-term treatments) but induce sustained abrogation of embryogenesis. In prospective, field exposure experiments with naive calves, prophylactic treatments with ivermectin and moxidectin prevented the development of adult worm infection, raising the possibility that drug-attenuated larval challenge infections may induce immunity. Putatively immune adult cattle exist in endemically exposed populations, and these have been shown to be significantly less susceptible to challenge than age-matched naive controls, whereas radically drug-cured, previously patently-infected cattle were not. Experimental infections with O. ochengi have revealed the kinetics of the immune response in relation to parasite development and demonstrate analogous responses to those reported in O. volvulus infection in humans and chimpanzees. In an immunization experiment with irradiated L3 larvae, cattle were significantly protected against experimental challenge--the first such demonstration of the experimental induction of immunity in a natural Onchocerca host-parasite system. Taken collectively, these studies not only demonstrate the similarity between the host-parasite relationships of O. ochengi in cattle and O. volvulus in humans, but promise to advance options for the control of human onchocerciasis. PMID- 10874717 TI - Determination of mosquito bloodmeal pH in situ by ion-selective microelectrode measurement: implications for the regulation of malarial gametogenesis. AB - Malarial gametocytes circulate in the peripheral blood of the vertebrate host as developmentally arrested intra-erythrocytic cells, which only resume development into gametes when ingested into the bloodmeal of the female mosquito vector. The ensuing development encompasses sexual reproduction and mediates parasite transmission to the insect. In vitro the induction of gametogenesis requires a drop in temperature and either a pH increase from physiological blood pH (ca pH 7.4) to about pH 8.0, or the presence of a gametocyte-activating factor recently identified as xanthurenic acid (XA). However, it is unclear whether either the pH increase or XA act as natural triggers in the mosquito bloodmeal. We here use pH sensitive microelectrodes to determine bloodmeal pH in intact mosquitoes. Measurements taken in the first 30 min after ingestion, when malarial gametogenesis is induced in vivo, revealed small pH increases from 7.40 (mouse blood) to 7.52 in Aedes aegypti and to 7.58 in Anopheles stephensi. However, bloodmeal pH was clearly suboptimal if compared to values required to induce gametogenesis in vitro. Xanthurenic acid is shown to extend the pH-range of exflagellation in vitro in a dose-dependent manner to values that we have observed in the bloodmeal, suggesting that in vivo malarial gametogenesis could be further regulated by both these factors. PMID- 10874718 TI - A 32 kDa surface antigen of Theileria parva: characterization and immunization studies. AB - Previous studies using monoclonal antibody (mAb) 4C9 specific for a 32 kDa antigen (p32) of Theileria parva demonstrated expression of the antigen on the surface of the sporozoite, making it a potential antigen for sporozoite neutralization. A full-length cDNA encoding the major merozoite/piroplasm surface antigen (mMPSA) of T. parva was cloned and expressed in bacteria. The expressed product reacted strongly with mAb 4C9, demonstrating identity between the p32 and mMPSA of T. parva. Using immunoblot analysis and immunoelectron microscopy with mAb 4C9 it was shown that the mMPSA is a major antigen of the merozoite and piroplasm at the cell surface, while lower levels of antigen are expressed in the sporozoite and schizont stages. Upregulation of the mMPSA occurs at merogony and can be induced by culturing schizont-infected lymphocytes at 42 degrees C. Recombinant mMPSA of T. parva induced high titres of specific antibodies in cattle but failed to confer protection against a T. parva sporozoite stabilate challenge. The pre-challenge sera also failed to neutralize infectivity of sporozoites in an in vitro assay. Possible reasons for the lack of parasite neutralization in vivo and in vitro are discussed. PMID- 10874719 TI - Immune responses during the acute stages of infection with the intestinal trematode Echinostoma caproni. AB - This study investigated the nature of the immune response of C57BL/6 mice infected with the trematode Echinostoma caproni. To determine the preferential development of either a Th1 or Th2 cytokine pattern during early stages of infection, cytokine production by spleen and mesenteric lymph node (MLN) cells during the first 3 weeks of infection was followed. Whereas spleen cells failed to respond to antigen stimulation, MLN cells produced IFN-gamma and to a lesser extent IL-4. IL-5 levels were elevated throughout the period studied. The humoral response was consistent with a Th1 cytokine pattern as antigen-specific IgG2a antibodies were preferentially developed. We investigated whether IFN-gamma is critical for establishment of E. caproni infection. Worm burden in infected mice treated with a single injection of anti-IFN-gamma mAb was significantly reduced compared to that of animals treated with a control antibody. PMID- 10874721 TI - Grouping of trypanosome species in mixed infections in Glossina pallidipes. AB - Trypanosomes in the dissection-positive proboscis of Glossina pallidipes were identified by PCR using species-specific primers. Of the 3741 flies dissected 643 were proboscis positive. PCR was performed on 406 dissection-positive probosces giving positive identifications in 352 (86.7%) and infection rates of 14.8% for congolense-type infections, 2.8% for vivax-type infections and 1.4% for the unidentified group. Of the 352 PCR identified infections 225 were single, 111 were double, 13 were triple infections and there were 3 quadruple infections. Statistical analysis suggests that mixed infections group into 3 largely separate divisions among the tsetse population (i) Trypanosoma congolense savannah and T. congolense Kenya coast, (ii) T. simiae, T. congolense Tsavo and T. godfreyi and (iii) T. vivax. We conclude that either differing feeding patterns among members of the fly population or the ability of the trypanosomes in each of the infection categories to significantly influence the maturation of trypanosomes in the other categories are the most likely causes of the groupings noted. Chi-squared analysis of dissection and PCR methods of trypanosome identification revealed profound differences (chi 2 = 19.1; D.F. = 1; P > 0.05). If confirmed in other studies these findings have serious implications for our understanding of trypanosome epidemiology in tsetse flies, much of which is founded on data from dissection-based trypanosome identifications. PMID- 10874720 TI - The parasitism of Schistosoma mansoni (Digenea-Trematoda) in a naturally infected population of water rats, Nectomys squamipes (Rodentia-Sigmodontinae) in Brazil. AB - Schistosomiasis is a health problem in Brazil and the role of rodents in maintaining the schistosome life-cycle requires further clarification. The influence of Schistosoma mansoni on a population of Nectomys squamipes was studied by capture-recapture (1st phase, from June 1991 to November 1995) and removal (2nd phase, from April 1997 to March 1999) studies at Sumidouro, Rio de Janeiro, Brazil. During both phases coproscopic examinations were performed. At the 2nd phase the rodents were perfused and worms were counted. The population dynamics of parasites was studied. During the 1st phase, female reproductive parameters, longevity, recruitment and survivorship rates and migration patterns were studied in relation to schistosome prevalence. Water contamination (source of miracidia), abundance intermediate host and rodent migration were related to prevalence. The N. squamipes population was not obviously influenced by the infection, as shown by the high number of reproductive infected females, high longevity of infected individuals and the absence of a relationship between recruitment or survivorship rates and the intensity of schistosome infection. The data indicate that N. squamipes can increase transmission of S. mansoni in endemic areas and carry it to non-infected areas. Furthermore, this rodent can be used as an indicator of a transmission focus. PMID- 10874722 TI - Trichinella spiralis and Trichinella pseudospiralis: developmental patterns of enzymes involved in thymidylate biosynthesis and pyrimidine salvage. AB - Thymidylate synthase, dihydrofolate reductase and dUTPase specific activities were found to remain at a high and constant level in crude extracts from adult worms of Trichinella spiralis, as well as from muscle larvae of both Trichinella spiralis (isolated 1-24 months after infection) and Trichinella pseudospiralis (isolated 5.5-13 months after infection). The results obtained with Trichinella pseudospiralis muscle larvae isolated with the use of pepsin did not differ from those obtained when pepsin was not used. No thymidine kinase activity could be detected in muscle larvae of either species and thymidine phosphorylase could be found only in T. pseudospiralis larvae isolated without the use of pepsin. Muscle larvae of both species contained orotidylate phosphoribosyl transferase activity, pointing to a possibility of 5-fluorouracil activation. Uridine phosphorylase, another enzyme involved in 5-fluorouracil anabolism, was also present in T. pseudospiralis muscle larvae. Results of comparative studies on inhibition of purified T. spiralis and rat thymidylate synthases by substrate (4-thio-5-fluoro dUMP, 2-thio-5-fluoro-dCMP and N4-hydroxy-dCMP) and cofactor (ZD 9331) analogues indicated only dUMP analogues to show feeble selectivity towards the parasite enzyme. A hypothesis is discussed, assuming high expression of thymidylate synthase in muscle larvae to be connected with their cells being arrested in the cell cycle. PMID- 10874723 TI - Transmission of Orientia tsutsugamushi, the aetiological agent for scrub typhus, to co-feeding mites. AB - Experiments were conducted to investigate the potential for transmission of Orientia tsutsugamushi, the aetiological agent for scrub typhus, when naturally infected mite larvae were co-feeding with uninfected larvae. Larvae from colonies of Leptotrombidium deliense and L. imphalum infected with O. tsutsugamushi were used. Transmission of O. tsutsugamushi to previously uninfected L. deliense and Blankaartia acuscutellaris co-fed with infected L. deliense was shown to occur. The overall minimum rate of acquisition was 1.6% (4/258) for L. deliense and 2.5% (3/119) for B. acuscutellaris. When individual infected L. deliense were co-fed with B. acuscutellaris acquisition of O. tsutsugamushi was not detected. However, when 4 and 8 infected larvae were co-fed with B. acuscutellaris acquisition of O. tsutsugamushi was detected. Transmission of O. tsutsugamushi was not observed when uninfected L. deliense were co-fed with infected L. imphalum. This novel transmission route may explain the occurrence of rickettsiae in genera other than Leptotrombidium spp, which are considered to be the main vectors of O. tsutsugamushi. PMID- 10874724 TI - Richness patterns in the parasite communities of exotic poeciliid fishes. AB - Three species of poeciliids (Gambusia holbrooki, Xiphophorus helleri and X. maculatus) and 15 species of ecologically similar native freshwater fishes (mainly eleotrids, ambassids, melanotaeniids and retropinnids) were examined for parasite richness to investigate parasite flux, qualitative differences, quantitative differences and the structuring factors in parasite communities in the 2 fish types in Queensland, Australia. Theory suggests that poeciliids would harbour depauperate parasite communities. Results supported this hypothesis; poeciliids harboured more species-poor parasite infracommunities and regional faunas than natives (P < 0.0001), despite greater sampling effort for the former. Cluster analysis of presence/absence data for poeciliids and the 6 most-sampled native fishes revealed that parasite communities of the 2 fish groups are qualitatively distinct; the proportion of parasite species with complex life cycles was lower in poeciliids than in native species, and Myxosporea, Microspora, Coccidia and parasitic Crustacea were all absent from poeciliids. Limited exchange of parasite species has occurred between natives and poeciliids. Logistic ordinal regression analysis revealed that fish origin (exotic or native), environmental disturbance and host sex were all significant determinants of parasite community richness (P < 0.05). Theoretical modelling suggests that poeciliids are at a competitive advantage over native fishes because of their lack of parasites. PMID- 10874725 TI - Conflict between co-occurring manipulative parasites? An experimental study of the joint influence of two acanthocephalan parasites on the behaviour of Gammarus pulex. AB - When two parasite species are manipulators and have different definitive hosts, there is a potential for conflict between them. Selection may then exist for either avoiding hosts infected with conflicting parasites, or for hijacking, i.e. competitive processes to gain control of the intermediate host. The evidence for both phenomena depends largely on the study of the relative competitive abilities of parasites within their common intermediate host. We studied the effects of simultaneous infection by a fish acanthocephalan parasite, Pomphorhynchus laevis, and a bird acanthocephalan parasite, Polymorphus minutus, on the behaviour of their common intermediate host, the amphipod Gammarus pulex. We compared the reaction to light and vertical distribution of individuals infected with both parasites to those of individuals harbouring a single parasite species and uninfected ones under controlled conditions. Compared to uninfected gammarids that were photophobic and tended to remain at the bottom of the water column, P. laevis-infected gammarids were attracted to light, whereas P. minutus-infected individuals showed a modified vertical distribution and were swimming closer to the water surface. The effects of both P. laevis and P. minutus appeared to be dependent only on their presence, not on their intensity. Depending on the behavioural trait under study, however, the outcome of the antagonism between P. laevis and P. minutus differed. The vertical distribution of gammarids harbouring both parasites was half-way between those of P. laevis- and P. minutus-infected individuals, whereas P. laevis was able to induce altered reaction to light even in the presence of P. minutus. We discuss our results in relation to the occurrence of active avoidance or hijacking between conflicting manipulative parasites and provide some recommendations for future research. PMID- 10874726 TI - Requirements for in vivo IFN-gamma induction by live microfilariae of the parasitic nematode, Brugia malayi. AB - Lymphatic filariasis caused by the parasitic nematode, Brugia malayi, is a chronic human disease immunologically characterized by stimulation of Th2 cells and reduced antigen-specific T cell responses. Single stage intra-peritoneal infections with infective larvae (L3) or adult nematodes induce Th2 cells, while the microfilarial stage (Mf) stimulates IFN-gamma and Mf-specific IgG1, IgG2a, IgG2b, IgG3 and IgM, but not IgE. To investigate whether IFN-gamma is elicited by live Mf in their natural site of infected, mice were infected intravenously. Intravenous infection had a striking effect on the response to Mf and high levels of IgE were induced even in the presence of IFN-gamma. Indeed IgE levels to Mf increased markedly with the number of immunizations, higher doses of Mf and prolonged exposure to Mf suggesting that under conditions of chronic antigen exposure, typical of human disease, Mf will stimulate high levels of IgE. The ability of Mf-induced IFN-gamma to modulate or regulate a pre-existing Th2 response, was investigated by infecting mice initially with adult male worms to induce a Th2 response, followed 14 days later by infection with Mf. Although Mf stimulated IFN-gamma in the presence of male adults, the antibody isotypes elicited did not reflect IFN-gamma induction and IgG1 and IgE dominated the response. Although it cannot be discounted that IFN-gamma induction by Mf may act locally as an inflammatory mediator or modulator of Th2 cells, these data suggest that Mf-stimulated IFN-gamma does not have a profound effect overall on progression of the Th2-dominated immune response to filarial infection. PMID- 10874727 TI - Anorexia in rats infected with the nematode, Nippostrongylus brasiliensis: experimental manipulations. AB - Nippostrongylus brasiliensis induces a biphasic anorexia in laboratory rats, the first phase coincident with lung invasion (ca day 2) and the second when the worms mature in the intestine (ca day 8). Using the anthelminthic, mebendazole (MBZ), N. brasiliensis infections of the rat were eliminated between the first and second anorexic episodes. This intervention prevented the expression of the second phase of anorexia. Rats exposed to a second infection with N. brasiliensis, 3 weeks after the primary infection, exhibited only a first phase anorexic response which was not influenced by MBZ termination of the primary infection. The lower cumulative food intake and weight gain of all infected rats after 8 days of infection were accompanied by elevated plasma insulin and, in some individuals, by elevated plasma leptin, compared with uninfected controls and previously-infected MBZ-treated rats. Messenger RNA levels for neuropeptide Y were higher in the hypothalamic arcuate nucleus of 8-day infected rats than in recovering MBZ-treated animals. Inoculation of rats with heat-killed N. brasiliensis larvae failed to induce anorexia and did not alter the severity of biphasic anorexia on subsequent injection of viable larvae. The first anorexic episode is therefore dependent upon viable migrating larvae. The second phase of anorexia clearly requires the continuing presence of the parasite beyond the lung phase. Viable migrating larvae are also required to confer 'resistance' to reinfection. PMID- 10874728 TI - Apoptosis in human Jurkat T cells after culture with live Taenia crassiceps cysticerci in vitro. AB - This study examines the effects of Taenia crassiceps cysticerci on the viability of a human T lymphocyte cell line (Jurkat). Both budding and non-budding T. crassiceps metacestodes were cultured over 24 and 48 h in the presence of Jurkat cells. Cell viability decreased with increasing numbers of cysticerci, particularly budding cysticerci. Single cell gel electrophoresis (comet) analysis, which grades DNA damage, showed a significant increase in apoptosis at 24 and 48 h. The morphology of treated cells was determined using acridine orange with the classical morphology of apoptotic bodies seen to increase with increasing cysticerci numbers over time. These results indicate that parasite induced apoptosis occurs during murine cysticercosis. Such a mechanism may be important in survival of other metacestode infections of medical or veterinary importance. PMID- 10874729 TI - Questions about the behaviour of bacterial pathogens in vivo. AB - Bacterial pathogens cause disease in man and animals. They have unique biological properties, which enable them to colonize mucous surfaces, penetrate them, grow in the environment of the host, inhibit or avoid host defences and damage the host. The bacterial products responsible for these five biological requirements are the determinants of pathogenicity (virulence determinants). Current knowledge comes from studies in vitro, but now interest is increasing in how bacteria behave and produce virulence determinants within the infected host. There are three aspects to elucidate: bacterial activities, the host factors that affect them and the metabolic interactions between the two. The first is relatively easy to accomplish and, recently, new methods for doing this have been devised. The second is not easy because of the complexity of the environment in vivo and its ever-changing face. Nevertheless, some information can be gained from the literature and by new methodology. The third aspect is very difficult to study effectively unless some events in vivo can be simulated in vitro. The objectives of the Discussion Meeting were to describe the new methods and to show how they, and conventional studies, are revealing the activities of bacterial pathogens in vivo. This paper sets the scene by raising some questions and suggesting, with examples, how they might be answered. Bacterial growth in vivo is the primary requirement for pathogenicity. Without growth, determinants of the other four requirements are not formed. Results from the new methods are underlining this point. The important questions are as follows. What is the pattern of a developing infection and the growth rates and population sizes of the bacteria at different stages? What nutrients are present in vivo and how do they change as infection progresses and relate to growth rates and population sizes? How are these nutrients metabolized and by what bacterial mechanisms? Which bacterial processes handle nutrient deficiencies and antagonistic conditions that may arise? Conventional and new methods can answer the first question and part of the second; examples are described. The difficulties of trying to answer the last two are discussed. Turning to production in vivo of determinants of mucosal colonization, penetration, interference with host defence and damage to the host, here are the crucial questions. Are putative determinants, which have been recognized by studies in vitro, produced in vivo and are they relevant to virulence? Can hitherto unknown virulence determinants be recognized by examining bacteria grown in vivo? Does the complement of virulence determinants change as infection proceeds? Are regulatory processes recognized in vitro, such as ToxR/ToxS, PhoP/PhoQ, quorum sensing and type III secretion, operative in vivo? What environmental factors affect virulence determinant production in vivo and by what metabolic processes? Examples indicate that the answers to the first four questions are 'yes' in most but not all cases. Attempts to answer the last, and most difficult, question are also described. Finally, sialylation of the lipopolysaccharide of gonococci in vivo by host-derived cytidine 5'-mono-phospho N-acetyl neuraminic acid, and the effect of host lactate are described. This investigation revealed a new bacterial component important in pathogenicity, the host factors responsible for its production and the metabolism involved. PMID- 10874730 TI - DNA topology and adaptation of Salmonella typhimurium to an intracellular environment. AB - The expression of genes coding for determinants of DNA topology in the facultative intracellular pathogen Salmonella typhimurium was studied during adaptation by the bacteria to the intracellular environment of J774A.1 macrophage like cells. A reporter plasmid was used to monitor changes in DNA supercoiling during intracellular growth. Induction of the dps and spv genes, previously shown to be induced in the macrophage, was detected, as was expression of genes coding for DNA gyrase, integration host factor and the nucleoid-associated protein H-NS. The topA gene, coding for the DNA relaxing enzyme topoisomerase I, was not induced. Reporter plasmid data showed that bacterial DNA became relaxed following uptake of S. typhimurium cells by the macrophage. These data indicate that DNA topology in S. typhimurium undergoes significant changes during adaptation to the intracellular environment. A model describing how this process may operate is discussed. PMID- 10874731 TI - The pathogenesis of Shigella flexneri infection: lessons from in vitro and in vivo studies. AB - Shigella flexneri is a Gram-negative facultatively intracellular pathogen responsible for bacillary dysentery in humans. More than one million deaths occur yearly due to infections with Shigella spp. and the victims are mostly children of the developing world. The pathogenesis of Shigella centres on the ability of this organism to invade the colonic epithelium where it induces severe mucosal inflammation. Much information that we have gained concerning the pathogenesis of Shigella has been derived from the study of in vitro models of infection. Using these techniques, a number of the molecular mechanisms by which Shigella invades epithelial cells and macrophages have been identified. In vivo models of shigellosis have been hampered since humans are the only natural hosts of Shigella. However, experimental infection of macaques as well as the murine lung and rabbit ligated ileal loop models have been important in defining some of the immune and inflammatory components of the disease. In particular, the murine lung model has shed light on the development of systemic and local immune protection against Shigella infection. It would be naive to believe that any one model of Shigella infection could adequately represent the complexity of the disease in humans, and more sophisticated in vivo models are now necessary. These models require the use of human cells and tissue, but at present such models remain in the developmental stage. Ultimately, however, it is with such studies that novel treatments and vaccine candidates for the treatment and prevention of shigellosis will be designed. PMID- 10874732 TI - Detection and analysis of gene expression during infection by in vivo expression technology. AB - Many limitations associated with the use of in vitro models for study of bacterial pathogenesis can be overcome by the use of technologies that detect pathogen gene expression during the course of infection within an intact animal. In vivo expression technology (IVET) accomplishes this with versatility: it has been developed with a variety of reporter systems which allow for either in vivo selection or ex vivo screening. Selectable gene fusion systems generally allow for the complementation of a bacterial metabolic defect that is lethal in vivo, or for antibiotic resistance during the course of in vivo antibiotic challenge. In contrast, the screenable gene fusion system uses a site-specific DNA recombinase that, when expressed in vivo, excises a selectable gene cassette from the bacterial chromosome. Loss of this cassette can then be either screened or selected for ex vivo. The recombinase-based IVET can be used to detect genes that are transcriptionally induced during infection, including those expressed transiently or at low levels and, in addition, can be used to monitor the spatial and temporal expression of specific genes during the course of infection. PMID- 10874733 TI - Measurement of bacterial gene expression in vivo. AB - The complexities of bacterial gene expression during mammalian infection cannot be addressed by in vitro experiments. We know that the infected host represents a complex and dynamic environment, which is modified during the infection process, presenting a variety of stimuli to which the pathogen must respond if it is to be successful. This response involves hundreds of ivi (in vivo-induced) genes which have recently been identified in animal and cell culture models using a variety of technologies including in vivo expression technology, differential fluorescence induction, subtractive hybridization and differential display. Proteomic analysis is beginning to be used to identify IVI proteins, and has benefited from the availability of genome sequences for increasing numbers of bacterial pathogens. The patterns of bacterial gene expression during infection remain to be investigated. Are ivi genes expressed in an organ-specific or cell type-specific fashion? New approaches are required to answer these questions. The uses of the immunologically based in vivo antigen technology system, in situ PCR and DNA microarray analysis are considered. This review considers existing methods for examining bacterial gene expression in vivo, and describes emerging approaches that should further our understanding in the future. PMID- 10874734 TI - Identification and analysis of bacterial virulence genes in vivo. AB - Signature-tagged mutagenesis is a mutation-based screening method for the identification of virulence genes of microbial pathogens. Genes isolated by this approach fall into three classes: those with known biochemical function, those of suspected function and some whose functions cannot be predicted from database searches. A variety of in vitro and in vivo methods are available to elucidate the function of genes of the second and third classes. We describe the use of some of these approaches to study the function of the Salmonella pathogenicity island 2 type III secretion system of Salmonella typhimurium. This virulence determinant is required for intracellular survival. Secretion by this system is induced by an acidic pH, and its function may be to alter trafficking of the Salmonella-containing vacuole. Use of a temperature-sensitive non-replicating plasmid and competitive index tests with other genes show that in vivo phenotypes do not always correspond to those predicted from in vitro studies. PMID- 10874735 TI - Salmonella interactions with host cells: in vitro to in vivo. AB - Salmonellosis (diseases caused by Salmonella species) have several clinical manifestations, ranging from gastroenteritis (food poisoning) to typhoid (enteric) fever and bacteraemia. Salmonella species (especially Salmonella typhimurium) also represent organisms that can be readily used to investigate the complex interplay that occurs between a pathogen and its host, both in vitro and in vivo. The ease with which S. typhimurium can be cultivated and genetically manipulated, in combination with the availability of tissue culture models and animal models, has made S. typhimurium a desirable organism for such studies. In this review, we focus on Salmonella interactions with its host cells, both in tissue culture (in vitro) and in relevant animal models (in vivo), and compare results obtained using these different models. The recent advent of sophisticated imaging and molecular genetic tools has facilitated studying the events that occur in disease, thereby confirming tissue culture results, yet identifying new questions that need to be addressed in relevant disease settings. PMID- 10874736 TI - In vivo gene expression and the adaptive response: from pathogenesis to vaccines and antimicrobials. AB - Microbial pathogens possess a repertoire of virulence determinants that each make unique contributions to fitness during infection. Analysis of these in vivo expressed functions reveals the biology of the infection process, encompassing the bacterial infection strategies and the host ecological and environmental retaliatory strategies designed to combat them (e.g. thermal, osmotic, oxygen, nutrient and acid stress). Many of the bacterial virulence functions that contribute to a successful infection are normally only expressed during infection. A genetic approach was used to isolate mutants that ectopically expressed many of these functions in a laboratory setting. Lack of DNA adenine methylase (Dam) in Salmonella typhimurium abolishes the preferential expression of many bacterial virulence genes in host tissues. Dam- Salmonella were proficient in colonization of mucosal sites but were defective in colonization of deeper tissue sites. Additionally, Dam- mutants were totally avirulent and effective as live vaccines against murine typhoid fever. Since dam is highly conserved in many pathogenic bacteria that cause significant morbidity and mortality worldwide, Dams are potentially excellent targets for both vaccines and antimicrobials. PMID- 10874737 TI - Challenge of investigating biologically relevant functions of virulence factors in bacterial pathogens. AB - Recent innovations have increased enormously the opportunities for investigating the molecular basis of bacterial pathogenicity, including the availability of whole-genome sequences, techniques for identifying key virulence genes, and the use of microarrays and proteomics. These methods should provide powerful tools for analysing the patterns of gene expression and function required for investigating host-microbe interactions in vivo. But, the challenge is exacting. Pathogenicity is a complex phenotype and the reductionist approach does not adequately address the eclectic and variable outcomes of host-microbe interactions, including evolutionary dynamics and ecological factors. There are difficulties in distinguishing bacterial 'virulence' factors from the many determinants that are permissive for pathogenicity, for example those promoting general fitness. A further practical problem for some of the major bacterial pathogens is that there are no satisfactory animal models or experimental assays that adequately reflect the infection under investigation. In this review, we give a personal perspective on the challenge of characterizing how bacterial pathogens behave in vivo and discuss some of the methods that might be most relevant for understanding the molecular basis of the diseases for which they are responsible. Despite the powerful genomic, molecular, cellular and structural technologies available to us, we are still struggling to come to grips with the question of 'What is a pathogen?' PMID- 10874738 TI - Virulence gene regulation inside and outside. AB - Much knowledge about microbial gene regulation and virulence is derived from genetic and biochemical studies done outside of hosts. The aim of this review is to correlate observations made in vitro and in vivo with two different bacterial pathogens in which the nature of regulated gene expression leading to virulence is quite different. The first is Vibrio cholerae, in which the concerted action of a complicated regulatory cascade involving several transcription activators leads ultimately to expression of cholera toxin and the toxin-coregulated pilus. The regulatory cascade is active in vivo and is also required for maintenance of V. cholerae in the intestinal tract during experimental infection. Nevertheless, specific signals predicted to be generated in vivo, such as bile and a temperature of 37 degrees C, have a severe down-modulating effect on activation of toxin and pilus expression. Another unusual aspect of gene regulation in this system is the role played by inner membrane proteins that activate transcription. Although the topology of these proteins suggests an appealing model for signal transduction leading to virulence gene expression, experimental evidence suggests that such a model may be simplistic. In Streptococcus pyogenes, capsule production is critical for virulence in an animal model of necrotizing skin infection. Yet capsule is apparently produced to high levels only from mutation in a two-component regulatory system, CsrR and CsrS. Thus it seems that in V. cholerae a complex regulatory pathway has evolved to control virulence by induction of gene expression in vivo, whereas in S. pyogenes at least one mode of pathogenicity is potentiated by the absence of regulation. PMID- 10874739 TI - Quorum sensing and the population-dependent control of virulence. AB - One crucial feature of almost all bacterial infections is the need for the invading pathogen to reach a critical cell population density sufficient to overcome host defences and establish the infection. Controlling the expression of virulence determinants in concert with cell population density may therefore confer a significant survival advantage on the pathogen such that the host is overwhelmed before a defence response can be fully initiated. Many different bacterial pathogens are now known to regulate diverse physiological processes including virulence in a cell-density-dependent manner through cell-cell communication. This phenomenon, which relies on the interaction of a diffusible signal molecule (e.g. an N-acylhomoserine lactone) with a sensor or transcriptional activator to couple gene expression with cell population density, has become known as 'quorum sensing'. Although the size of the 'quorum' is likely to be highly variable and influenced by the diffusibility of the signal molecule within infected tissues, nevertheless quorum-sensing signal molecules can be detected in vivo in both experimental animal model and human infections. Furthermore, certain quorum-sensing molecules have been shown to possess pharmacological and immunomodulatory activity such that they may function as virulence determinants per se. As a consequence, quorum sensing constitutes a novel therapeutic target for the design of small molecular antagonists capable of attenuating virulence through the blockade of bacterial cell-cell communication. PMID- 10874740 TI - Type III secretion: a bacterial device for close combat with cells of their eukaryotic host. AB - Salmonella, Shigella, Yersinia, Pseudomonas aeruginosa, enteropathogenic Escherichia coli and several plant-pathogenic Gram-negative bacteria use a new type of systems called 'type III secretion' to attack their host. These systems are activated by contact with a eukaryotic cell membrane and they allow bacteria to inject bacterial proteins across the two bacterial membranes and the eukaryotic cell membrane to reach a given compartment and destroy or subvert the target cell. These systems consist of a secretion apparatus made up of about 25 individual proteins and a set of proteins released by this apparatus. Some of these released proteins are 'effectors' that are delivered by extracellular bacteria into the cytosol of the target cell while the others are 'translocators' that help the 'effectors' to cross the membrane of the eukaryotic cell. Most of the 'effectors' act on the cytoskeleton or on intracellular signalling cascades. One of the proteins injected by the enteropathogenic E. coli serves as a membrane receptor for the docking of the bacterium itself at the surface of the cell. PMID- 10874741 TI - Evolution of microbial pathogens. AB - Various genetic mechanisms including point mutations, genetic rearrangements and lateral gene transfer processes contribute to the evolution of microbes. Long term processes leading to the development of new species or subspecies are termed macroevolution, and short-term developments, which occur during days or weeks, are considered as microevolution. Both processes, macro- and microevolution need horizontal gene transfer, which is particularly important for the development of pathogenic microorganisms. Plasmids, bacteriophages and so-called pathogenicity islands (PAIs) play a crucial role in the evolution of pathogens. During microevolution, genome variability of pathogenic microbes leads to new phenotypes, which play an important role in the acute development of an infectious disease. Infections due to Staphylococcus epidermidis, Candida albicans and Escherichia coli will be described with special emphasis on processes of microevolution. In contrast, the development of PAIs is a process involved in macroevolution. PAIs are especially important in processes leading to new pathotypes or even species. In this review, particular attention will be given to the fact that the evolution of pathogenic microbes can be considered as a specific example for microbial evolution in general. PMID- 10874743 TI - The relationship between the dissolved inorganic carbon concentration and growth rate in marine phytoplankton. AB - A range of marine phytoplankton was grown in closed systems in order to investigate the kinetics of dissolved inorganic carbon (DIC) use and the influence of the nitrogen source under conditions of constant pH. The kinetics of DIC use could be described by a rectangular hyperbolic curve, yielding estimations of KG(DIC) (the half saturation constant for carbon-specific growth, i.e. C mu) and mu max (the theoretical maximum C mu). All species attained a KG(DIC) within the range of 30-750 microM DIC. For most species, NH4+ use enabled growth with a lower KG(DIC) and/or, for two species, an increase in mu max. At DIC concentrations of > 1.6 mM, C mu was > 90% saturated for all species relative to the rate at the natural seawater DIC concentration of 2.0 mM. The results suggest that neither the rate nor the extent of primary productivity will be significantly limited by the DIC in the quasi-steady-state conditions associated with oligotrophic oceans. The method needs to be applied in the conditions associated with dynamic coastal (eutrophic) systems for clarification of a potential DIC rate limitation where cells may grow to higher densities and under variable pH and nitrogen supply. PMID- 10874742 TI - The immune responses to bacterial antigens encountered in vivo at mucosal surfaces. AB - Mammals have evolved a sophisticated immune system for handling antigens encountered at their mucosal surfaces. The way in which mucosally delivered antigens are handled influences our ability to design effective mucosal vaccines. Live attenuated derivatives of pathogens are one route towards the development of mucosal vaccines. However, some molecules, described as mucosal immunogens, are inherently immunogenic at mucosal surfaces. Studies on mucosal immunogens may facilitate the identification of common characteristics that contribute to mucosal immunogenicity and aid the development of novel, non-living mucosal vaccines and immunostimulators. PMID- 10874744 TI - Two spatial memories for honeybee navigation. AB - Insect navigation is thought to be based on an egocentric reference system which relates vector information derived from path integration to views of landmarks experienced en route and at the goal. Here we show that honeybees also possess an allocentric form of spatial memory which allows localization of multiple places relative to the intended goal, the hive. The egocentric route memory, which is called the specialized route memory (SRM) here, initially dominates navigation when an animal is first trained to a feeding site and then released at an unexpected site and this is why it is the only reference system detected so far in experiments with bees. However, the SRM can be replaced by an allocentric spatial memory called the general landscape memory (GLM). The GLM is directly accessible to the honeybee (and to the experimenter) if no SRM exists, for example, if bees were not trained along a route before testing. Under these conditions bees return to the hive from all directions around the hive at a speed comparable to that of an equally long flight along a trained route. The flexible use of the GLM indicates that bees may store relational information on places, connections between landmarks and the hive and/or views of landmarks from different directions and, thus, the GLM may have a graph structure, at least with respect to one goal, i.e. the hive. PMID- 10874745 TI - A selective trade-off for territoriality and non-territoriality in the polymorphic damselfly Mnais costalis. AB - Males of the damselfly Mnais costalis occur as territorial orange-winged 'fighter' males or non-territorial clear-winged 'sneaker' males. Their morph life histories differ considerably but the estimated lifetime reproductive success is the same for the two morphs. In this study we compared the developmental and reproductive costs associated with the two morphs. Orange-winged male and female reproductive costs resulted in a decline in adult fat reserves with increasing age. In contrast, the fat reserves of clear-winged males remained constant with adult age. Body size was positively correlated with mating success in orange winged males, but had no influence on the mating success of clear-winged males. The orange-winged male flight muscle ratios (FMRs) were significantly higher than the clear-winged male and female FMRs. However, there was no difference in the size-corrected fat reserves of the two morphs; both had higher fat reserves than females. The gain in mass between eclosion and reproduction in orange-winged males and females was almost double the mass gained by clear-winged males, suggesting that clear-winged male development is less costly. An experiment in which pre-reproductive levels of nutrition were manipulated confirmed this. PMID- 10874746 TI - Invading predatory crustacean Dikerogammarus villosus eliminates both native and exotic species. AB - As the tempo of biological invasions increases, explanations and predictions of their impacts become more crucial. Particularly with regard to biodiversity, we require elucidation of interspecific behavioural interactions among invaders and natives. In freshwaters in The Netherlands, we show that the invasive Ponto Caspian crustacean amphipod Dikerogammarus villosus is rapidly eliminating Gammarus duebeni, a native European amphipod, and Gammarus tigrinus, until now a spectacularly successful invader from North America. In the laboratory, survival of single (unguarded) female G. duebeni was significantly lower when male D. villosus were free to roam as compared with isolated within microcosms. In addition, survival of paired (guarded) female G. duebeni was significantly lower when male D. villosus as compared with male G. duebeni were present. D. villosus killed and consumed both recently moulted and, unusually, intermoult victims. Survival of G. tigrinus was significantly lower when D. villosus were free to roam as compared with isolated within microcosms and, again, both moulted and intermoult victims were preyed upon. Male D. villosus were significantly more predatory than were females, while female G. tigrinus were significantly more often preyed upon than were males. Predation by D. villosus on both species occurred over a range of water conductivities, an environmental feature previously shown to promote amphipod coexistence. This predatory invader is predicted to reduce further the amphipod diversity in a range of freshwater habitats in Europe and North America. PMID- 10874747 TI - Evolution of parasite virulence against qualitative or quantitative host resistance. AB - We analysed the effects of two different modes of host resistance on the evolution of parasite virulence. Hosts can either adopt an all-or-nothing qualitative response (i.e. resistant hosts cannot be infected) or a quantitative form of resistance (i.e. which reduces the within-host growth rate of the parasite). We show that the mode of host resistance greatly affects the evolutionary outcome. Specifically, a qualitative form of resistance reduces parasite virulence, while a quantitative form of resistance generally selects for higher virulence. PMID- 10874748 TI - Avoiding the cost of males in obligately asexual Daphnia pulex (Leydig). AB - Asexual organisms are thought to gain an advantage by avoiding the cost of producing males. In the cladoceran Daphnia pulex (Leydig), male production is determined by the environment and is independent of the origin of the asexual obligate parthenogens from the sexual cyclical parthenogens. If there is a cost to producing males, successful obligate parthenogens should have reduced or eliminated male production. Field and laboratory observations showed that obligate parthenogens have much-reduced male production compared to cyclical parthenogens. Although the reduction or elimination of males in the obligate parthenogens suggests that the cost of males is avoided, the coexistence of sexual and asexual forms of D. pulex may be partially explained by cyclical parthenogens compensating for the cost of males by having greater fecundity. In addition, the absence of a mating constraint for the obligate parthenogens may favour an increased allocation to asexual diapausing eggs earlier in the season compared to the cyclical parthenogens which require mating with males to produce sexual diapausing eggs. No difference in the production of diapausing eggs was observed, probably because males were abundant in populations of cyclical parthenogens and do not appear to limit the production of sexual diapausing eggs. D. pulex is a useful system for determining the ecological consequences of abandoning sexual reproduction and explaining the coexistence of sexual and asexual forms of a species. PMID- 10874749 TI - Pectoral fins and paternal quality in sticklebacks. AB - Sexual selection through female mate choice exerts a strong selection pressure on males' sexual traits, particularly when direct benefits are involved. In species with male parental care, one would expect sexual selection to favour paternal quality, for instance through selection on morphological structures which promote quality. We experimentally studied the influence of pectoral fins on paternal quality in male three-spined sticklebacks (Gasterosteus aculeatus L.). After reductions of fin area to different degrees, similar-sized males had to perform a complete reproductive cycle in enclosures in the field. The collected data on fanning behaviour and egg development showed that a reduction in pectoral fin size affected paternal quality probably through an increased beat frequency of the pectorals. Thus, pectoral fins can potentially signal paternal quality to choosy females. PMID- 10874750 TI - Adaptive significance of a circadian clock: temporal segregation of activities reduces intrinsic competitive inferiority in Drosophila parasitoids. AB - Most organisms show self-sustained circadian oscillations or biological clocks which control their daily fluctuations in behavioural and physiological activities. While extensive progress has been made in understanding the molecular mechanisms of biological clocks, there have been few clear demonstrations of the fitness value of endogenous rhythms. This study investigated the adaptive significance of circadian rhythms in a Drosophila parasitoid community. The activity rhythms of three sympatric Drosophila parasitoids are out of phase, the competitively inferior parasitoid species being active earlier than the superior competitor. This temporal segregation appears at least partially determined by endogenous periods of the clock which also vary between species and which correlate the time of activity. This earlier activity of the inferior competitor significantly reduces its intrinsic competitive disadvantage when multiparasitism occurs, thus suggesting that natural selection acting on the phase of the rhythm could substantially deviate the endogenous period from the optimal ca. 24 h period. This study demonstrates that temporal segregation of competing species could be endogenously controlled, which undoubtedly favours their coexistence in nature and also shows how natural selection can act on biological clocks to shape daily activity patterns. PMID- 10874751 TI - Phylogenetic analysis of arthropods using two nuclear protein-encoding genes supports a crustacean + hexapod clade. AB - Recent phylogenetic analyses using molecular data suggest that hexapods are more closely related to crustaceans than to myriapods, a result that conflicts with long-held morphology-based hypotheses. Here we contribute additional information to this debate by conducting phylogenetic analyses on two nuclear protein encoding genes, elongation factor-1 alpha (EF-1 alpha) and the largest subunit of RNA polymerase II (Pol II), from an extensive sample of arthropod taxa. Results were obtained from two data sets. One data set comprised 1092 nucleotides (364 amino acids) of EF-1 alpha and 372 nucleotides (124 amino acids) of Pol II from 30 arthropods and three lobopods. The other data set contained the same EF-1 alpha fragment and an expanded 1038-nucleotide (346-amino-acid) sample of Pol II from 17 arthropod taxa. Results from maximum-parsimony and maximum-likelihood analyses strongly supported the existence of a Crustacea + Hexapoda clade (Pancrustacea) over a Myriapoda + Hexapoda clade (Atelocerata). The apparent incompatibility between the molecule-based Pancrustacea hypothesis and morphology based Atelocerata hypothesis is discussed. PMID- 10874752 TI - Evidence of sex-linked effects on the inheritance of human longevity: a population-based study in the Valserine valley (French Jura), 18-20th centuries. AB - A long-standing puzzle in gerontology is the sex dependence of human longevity and its inheritance. We have analysed the sex-linked pattern of inheritance of longevity from 643 nuclear families on the historical population register of a French valley. We have focused on mean conditional life expectancy at a minimum age of 50 years, thus, in the present study, longevity refers to late or post reproductive survival. A comparison of parents' and offspring's longevity has shown the existence of a heritable component of late survival in this population. We have found that the heritable component was substantially larger for daughters compared to sons. Moreover, this result appeared to be specific to late survival, that is, when only post-reproductive mortality for parental and offspring generations is taken into account. The stronger resemblance of parents to their daughters was no longer observed when considering younger ages at death for the offspring. This observation explains the hitherto unaccountable diversity of data in previous studies. PMID- 10874753 TI - Sperm competition games between related males. AB - Three sperm competition games against relatives are examined. In the first, a male has no information at the time of mating as to whether or not his ejaculate will face sperm competition from a related or unrelated male. Sperm expenditure increases with overall sperm competition risk q and declines with the probability rho that the competitor shares the same allele for sperm expenditure. In the second game, males have almost perfect information: they 'know' whether there will be sperm competition and, if so, whether this involves a related or unrelated male. Sperm expenditure is reduced by a factor rho when competing with a relative. In the third game, males 'know' when they compete with relatives, but have no information for other matings whether they will face sperm competition from unrelated males. A male without information expends less on his ejaculate than a male competing with a close relative if the overall risk of sperm competition is low, but more if the overall risk is high. The average relative ejaculate expenditure is the same in all three games so that, if this determines testis size, data is required only on the overall sperm competition risk, the probability of competing with a relative and the average rho in order to perform comparative analyses. PMID- 10874754 TI - Genetic evidence for the effect of a postglacial population expansion on the phylogeography of a North American songbird. AB - Phylogeographical studies of Nearctic songbirds conducted to date have yielded unexpectedly low levels of genetic differentiation and weak phylogeographical structure in mitochondrial DNA lineages as compared with species studied in Neotropical areas. Factors leading to this pattern may include (i) gene flow, (ii) population expansions from bottlenecked populations, and (iii) selective sweeps. Here we provide evidence for the role played by Pleistocene postglacial population expansions on the phylogeography of MacGillivray's warbler (Oporornis tolmiei), a long-distance migratory bird. Samples from 12 breeding localities in the temperate USA were compared with those from two localities in north-eastern Mexico. The former showed evidence of a Late Pleistocene population expansion as indicated by low haplotype and nucleotide diversity, a star-like phylogeny of alleles, and a mismatch distribution indicating a sudden increase in effective population size. By contrast, the Mexican population showed high levels of genetic diversity and a mismatch distribution as expected for a population unaffected by sudden demographic change. Haplotypes from the two regions formed two distinct phylogroups which separated roughly one million years ago according to a conventional molecular clock for songbirds. This study provides support for the Pleistocene expansion hypothesis in MacGillivray's warbler and suggests that postglacial expansion of bottlenecked populations is responsible for the lack of variation and structure reported for most North American songbird species. PMID- 10874756 TI - The origin and age of haplochromine fishes in Lake Victoria, east Africa. AB - According to a widely held view, the more than 300 species of haplochromine cichlid fishes in Lake Victoria (LV), East Africa, originated from a single founder species in less than 12,000 years. This view, however, does not follow from the published geological and molecular evidence. The former does indeed suggest that the LV basin dried out less than 15,000 years ago, but it does not provide any information about the species that re-colonized the new lake or that remained in the rivers draining the area. The molecular evidence is inconclusive with respect to the origin of the LV haplochromines because cichlids from critical regions around LV were not adequately sampled; and as far as the age of the LV haplochromines is concerned, it in fact led to an estimate of 250,000 750,000 years old. In the present study, mitochondrial DNA (control region) variation was determined by heteroduplex and sequencing analyses of more than 670 specimens collected at widely distributed East African riverine and lacustrine localities. The analyses revealed the existence of seven haplogroups (I-VII) distinguishable by characteristic substitutions. All endemic LV samples tested fell into one of these haplogroups (V) which, however, was also found to be present at various other localities, both riverine and lacustrine, outside LV. Within this haplogroup, four subgroups (VA through VD) could be distinguished, two of which (VB and VC) were represented in LV and at other localities. The great majority of the LV haplochromine species could be classified as belonging to the VC subgroup, which was found only in LV and in the rivers draining into it. Hence, while the endemic haplochromine species of LV could not have originated from a single founding population, the lake does harbour a large species flock which probably arose in situ. PMID- 10874755 TI - Can fast early rates reconcile molecular dates with the Cambrian explosion? AB - Molecular dates consistently place the divergence of major metazoan lineages in the Precambrian, leading to the suggestion that the 'Cambrian explosion' is an artefact of preservation which left earlier forms unrecorded in the fossil record. While criticisms of molecular analyses for failing to deal with variation in the rate of molecular evolution adequately have been countered by analyses which allow both site-to-site and lineage-specific rate variation, no analysis to date has allowed the rates to vary temporally. If the rates of molecular evolution were much higher early in the metazoan radiation, molecular dates could consistently overestimate the divergence times of lineages. Here, we use a new method which uses multiple calibration dates and an empirically determined range of possible substitution rates to place bounds on the basal date of divergence of lineages in order to ask whether faster rates of molecular evolution early in the metazoan radiation could possibly account for the discrepancy between molecular and palaeontological date estimates. We find that allowing basal (interphylum) lineages the fastest observed substitution rate brings the minimum possible divergence date (586 million years ago) to the Vendian period, just before the first multicellular animal fossils, but excludes divergence of the major metazoan lineages in a Cambrian explosion. PMID- 10874757 TI - [Estrogens still and always?]. PMID- 10874758 TI - [Hormone replacement therapy in menopause in question?]. PMID- 10874759 TI - [Histoplasmosis due to Histoplasma capsulatum capsulatum and HIV infection]. AB - PURPOSE: Histoplasmosis due to Histoplasma capsulatum is a granulomatous fungic infection which appears opportunistic and disseminated in immunocompromised patients, especially among HIV patients in whom it can lead to death. Histoplasmosis is endemic in numerous areas worldwide, but in Europe most of the cases reported are imported. We describe the clinical features and the available diagnosis methods issued from our experience in French Guyana. METHODS: Contamination occurs by inhalation of spores contained in dust. Most endemic areas are located on the American continent, including the French West Indies, where the incidence of histoplasmosis among HIV patients in French Guyana varies from 1.2 to 2.2% per year. In non-immunocompromised patients, histoplasmosis is asymptomatic most of the time. In HIV patients, the disseminated form is common and may occur many years after exposure to the fungus. RESULTS: Non-specific symptoms, similar to those of either tuberculosis or other opportunistic infections, may reveal disseminated histoplasmosis in patients with AIDS. Early treatment (amphotericin B or itraconazole) is effective; however, it should be followed by a lifelong antifungic treatment (itraconazole) to prevent relapse. CONCLUSION: The infection should be suspected in any febrile HIV-infected patient with CD4 blood cell count < 200/mm3, if he/she ever travelled in an endemic zone. Direct examination of smear relating to clinical symptoms help guide diagnosis, while culture will confirm it after at least 4 weeks. Efficient serologic techniques for HIV-infected patients are not available in Europe. PMID- 10874760 TI - [Self health evaluation using a visual analog scale for elderly patients presenting with pain or early dementia]. AB - PURPOSE AND METHODS: Sixteen elderly patients with early dementia and 20 elderly patients with pain, all hospitalized in a geriatric hospital, evaluated their memory, health and mood, using a visual analog scale. Observations of their families and of the medical staff were also recorded for subsequent comparison. The patients' complaints were also compared to those of a control group including 16 healthy elderly subjects. RESULTS: Patients with early dementia complained more often than control subjects of memory loss and health and mood disorders, while patients with pain complained more often of only health and mood disorders. However, only patients with dementia complained of memory loss. Complaints noticed by their families and medical staff were in good agreement with theirs. Regarding patients with early dementia, the medical staff underestimated their complaint of memory loss. CONCLUSION: Visual analog scales appear to be valuable tools regarding not only evaluation of pain but also the patients' general condition. Complaints of memory loss could be of value as they would suggest the existence of early dementia in elderly patients. However, our study emphasizes the need for training medical staff to listen to such a complaint. PMID- 10874761 TI - [Heat shock proteins or "stress proteins"]. AB - INTRODUCTION: Heat shock proteins (HSP) are molecular chaperones which facilitate the biosynthesis and maturation of proteins within cells (protein folding). They promote assembly and disassembly of polypeptides and play a major role in cellular function, not only during the stress response but also at basal state. CURRENT KNOWLEDGE AND KEY POINTS: As HSP are immunogenic molecules and can be expressed on cellular membranes, their role in auto-immune and inflammatory diseases, particularly in systemic lupus erythematosus, Behcet's disease and rheumatoid arthritis, has been studied. Cellular immune response of T cells and humoral response with antibodies production against HSP occurring in the course of those diseases have been observed. FUTURE PROSPECTS AND PROJECTS: Anti-HSP immune response might provide better understanding of the pathogenic mechanisms involved in those diseases. At present it is not known whether HSP can trigger them. Indeed, anti-HSP immunity could be induced by the immunological process or be part of a normal immunoregulatory response. PMID- 10874762 TI - [Hemoperitoneum of ovarian origin complicating antivitamin K treatment]. AB - INTRODUCTION: Ovarian hemorrhage with hemoperitoneum is a rare but serious complication of ovulation related to rupture of either the corpus luteum or functional cyst. It is due to treatment using oral indirect anticoagulant and specifically affects young women. CURRENT KNOWLEDGE AND KEY POINTS: We review cases that were reported since the initial description by Weseley in 1957. The main indications for oral indirect anticoagulant are thrombophlebitis and valvular cardiac prosthesis. Pelvic pain with peritoneal irritation is the most common symptom in more than one third of the patients. An initial collapse is reported in 22% of the cases. Surgery is the main treatment. Mortality is 3% and recurrences occur in nearly 25% of the patients. FUTURE PROSPECTS AND PROJECTS: Potential ovarian hemorrhage should be investigated when a woman taking oral indirect anticoagulant develops acute abdominal pain. Surgery should be conservative and whenever possible, should include celioscopy. Systematic ovarian blockade should be discussed in women taking long-term oral indirect anticoagulant. PMID- 10874764 TI - [Emergency coronary angioplasty following treatment with 5-fluorouracil]. AB - INTRODUCTION: The incidence of cardiac toxicity due to 5-fluorouracil (5-FU) ranges from 1.2 to 18%. Most complications occur at the time of the first cure. Their mechanisms have not yet been clearly defined. EXEGESIS: The authors report a case of unstable angina induced by 5-FU. A coronary angioplasty was performed on a previously ignored coronary lesion. CONCLUSION: Recent studies support the hypothesis that 5-FU has endothelial toxicity resulting in thrombogenic effect and release of vasoactive substances. Unstable angina pectoris would be related to plaque rupture caused by 5-FU. Patients with previous history of coronary disease are at significantly increased risk for 5-FU-induced cardiotoxicity. They probably would benefit from continuous electrocardiographic monitoring. Rechallenge with 5-FU after cardiotoxicity problems should include only those patients for whom there is no alternative treatment. PMID- 10874765 TI - [Menopause and hormone replacement]. AB - INTRODUCTION: Considered until recently as a biological fate, menopause has evolved within a decade into a major public health issue at stake. Such an evolution results from various factors that deserve an exhaustive critical approach because the situation is much more complex than it appears to be at first analysis. CURRENT KNOWLEDGE AND KEY POINTS: 1) Ninety-five percent of the available epidemiological information relies on observation or cohort studies, such as case-control studies that do not allow any certitude in regard to therapy. 2) Chronic estrogen deficiency probably plays a pathogenic role in various symptoms or pathological conditions that are associated to menopause. 3) However, behind the paradigm of menopause, there is a whole psychosocial construct that classifies what remains a physiological condition within major risk factors, just as for the most serious chronic diseases. 4) Indeed, menopause cannot be considered as a well-characterized disease, and hormone replacement therapy is undeniably a complex therapeutic intervention that requires proper prescription and careful evaluation of the benefit:risk ratio. 5) While such a treatment is now considered as the gold standard for the prevention of post menopausal osteoporosis, this does not hold true regarding cardiovascular diseases. 6) Although hormone replacement therapy may lead to the relief of various symptoms associated with menopause, it may also result in side-effects that extreme medication cannot prevent, especially since some of them are not fully known, particularly in the case of either long-term or very long-term treatments. FUTURE PROSPECTS AND PROJECTS: A more rigorous evaluation of side effects of hormone replacement therapy in the framework of long-term controlled trials is therefore clearly required. The indications of such a treatment should only rely on objective data and not on questionable studies or impressions at clinical examination. PMID- 10874763 TI - [Is there a place for interferon-alpha in the treatment strategy of multicentric Castleman's disease?]. AB - INTRODUCTION: Castleman's disease is an unusual condition of unknown cause, consisting of massive proliferation of lymphoid tissue. Two forms (localized and multicentric) have been described. Interleukin-6 (IL-6) is at the core of the disease, being responsible for most of the clinical and biological signs that may be observed. Despite the benignancy of this pre-lymphoma condition, its course is usually aggressive and of poor prognosis in regard to the multicentric form. No consensus regarding treatment has been defined. Available data on the multicentric form of the disease are to scarce to allow any conclusion about the treatment timing and type of chemotherapy best suited to this condition. We report the case of a patient in whom interferon alpha (IFN-alpha) was used as first line treatment. EXEGESIS: The case of a 52-year-old man with multicentric Castleman's disease combined with high IL-6, in whom, however, testing for human herpes virus-8 proved to be negative, is described. Interferon alpha (4.5 MU/m2 three times per week during 18 months) administered as first line treatment induced dramatic improvement in the patient's general condition and normalization of the tumoral syndrome. Moreover, biological parameters and IL-6 returned to normal. Two years after interferon disruption, complete remission is still present. CONCLUSION: On the basis of the present data and those of two previous observations, anti-IL-6 and anti-infective properties of IFN-alpha are discussed. Treatment of multicentric Castleman's disease is based on corticosteroids and drugs derived from those pertaining to treatment of malignant lymphomas. Our results indicate that IFN-alpha is truly directed against Castleman's disease and has less toxicity than drugs usually prescribed. This argues for early use of IFN alpha in Castleman's disease, in association or not with corticosteroids. PMID- 10874766 TI - [A good profile. Coccygeal metastasis]. PMID- 10874767 TI - [Polyarthritis accompanying alveolar echinococcosis or the icing on the cake]. PMID- 10874768 TI - [Complete heart block following chronic chloroquine treatment]. PMID- 10874769 TI - [Hemoperitonitis revealing an isolated splenic metastasis secondarily attributed to a caecal adenocarcinoma]. PMID- 10874770 TI - [Pneumococcal cellulitis revealing a myeloma]. PMID- 10874772 TI - [Mechanisms of cytokine-induced cholestasis]. AB - INTRODUCTION: Cholestasis is a clinical, biological and histological syndrome that is secondary to the decrease in or disruption of biliary secretion. Clinical or biological cholestasis is common in the course of either infections (parainfectious cholestasis), various cancers (paraneoplastic cholestasis), granulomatosis or in the syndrome accompanying macrophage activation. Cytokine (IL-1, IL-6 and tumor necrosis factor-alpha [TNF-alpha]) synthesis by Kupffer's cells (intrahepatic macrophages) in response to the release of endotoxins occurs in the course of the various clinical syndromes, particularly in the course of sepsis. EXEGESIS: Recently, it has been demonstrated that proinflammatory cytokines and endotoxins lead to cholestasis through modulation of the activity of bile acid transporters and other organic anions. CONCLUSION: Cholestasis observed in various clinical syndromes in response to either proinflammatory cytokines or endotoxins might be related to a decrease in the flow which is secondary to a decrease in the activity of organic anion transporters. PMID- 10874771 TI - [Clinicobiological study of 100 symptomatic patients with factor V Leiden mutation]. PMID- 10874773 TI - Ocular aspects of myasthenia gravis. AB - Ocular myasthenia gravis is a not uncommon autoimmune disorder causing diplopia, ptosis, and weakness of lid closure. The predilection of myasthenia for the ocular muscles may be related to differences between limb and extraocular muscles in either physiological function or antigenicity. Clinically, ocular myasthenia can mimic any form of pupil-sparing ocular motility disorder. Dynamic abnormalities of myasthenic eye movements may reflect the primary hallmarks of the disease, which are fatigability and variability in strength, or secondary adaptive effects by the central nervous system. Tests to confirm the diagnosis include edrophonium challenge, repetitive nerve stimulation, single-fiber electromyography (EMG) of the frontalis, and assays for antibody directed against the acetylcholine receptor: all are less sensitive for ocular myasthenia than for generalized myasthenia. There is a higher incidence of other autoimmune conditions in myasthenia, notably thymoma and thyroid dysfunction. The differential diagnosis includes other diseases of the neuromuscular junction, such as Lambert-Eaton syndrome and botulism. Treatment consists of symptomatic use of acetylcholinesterase inhibitors and immunosuppression with steroids or azathioprine. Between 50 and 70% of patients with ocular myasthenia will eventually develop generalized disease: there is some retrospective data that steroids or azathioprine may reduce this by about 75%. The role of thymectomy in ocular myasthenia remains unclear. PMID- 10874774 TI - Vertical diplopia. AB - The diagnosis of an acquired vertical strabismus is not always straightforward. There is no one specific test that will diagnose a vertical deviation. The clinical presentation, signs, and symptoms are the driving forces that will help lead to the correct diagnosis. Patients with binocular vertical diplopia may have symptoms of recent onset or that have been long-standing. Others may not even be completely aware that their ocular symptoms are attributable to a doubled vertical image. The differential diagnosis for vertical diplopia includes oculomotor nerve palsy, superior oblique palsy, restrictive ophthalmopathies, myasthenia gravis, and skew deviation. This differential diagnosis is best used to sort out signs and symptoms in a patient with a vertical misalignment and diplopia. Because most clinicians feel more comfortable addressing the patient with complaints of horizontal diplopia, this paper will discuss the causes of vertical diplopia so that recognition will be easier, thus leading to more accurate diagnoses. PMID- 10874775 TI - Abnormalities of eyelid position and function. AB - Evaluation of the eyelids is an important part of the neuro-ophthalmic examination. Abnormal eyelid position and function can be caused by disorders involving the third cranial nerve, the oculosympathetic pathway, and the seventh cranial nerve, as well as supranuclear pathways, or as a result of neuromuscular diseases. To avoid unwarranted neurological investigations, it is also important for the clinician to recognize non-neurological eyelid abnormalities (such as ptosis from levator dehiscence or eyelid edema). PMID- 10874776 TI - Dysthyroid orbitopathy. AB - Dysthyroid orbitopathy (DO) is an autoimmune disorder usually associated with Graves' disease. The extra-ocular muscles are the target of the autoimmune attack. As a result, they become enlarged, producing restrictive ophthalmoplegia and proptosis. Other cardinal signs of DO include upper eyelid retraction and lag, conjunctival injection and chemosis, and periorbital edema. Visual loss may occur if the enlarged extra-ocular muscles compress the optic nerve in the orbital apex. These cardinal signs may mimic neurological conditions, including Parinaud's syndrome, sixth nerve palsy, carotid-cavernous fistula, and spheno orbital meningioma. Treatment is directed at the cardinal signs and is largely palliative. Under certain clinical conditions, judicious use of corticosteroids, radiation therapy, or surgery designed to decompress the orbit, correct strabismus, or restore normal lid position may help to restore visual function. PMID- 10874777 TI - Third nerve palsies. AB - The diagnosis and management of third nerve dysfunction varies according to the age of the patient, characteristics of the third nerve palsy, and presence of associated symptoms and signs. Indeed, third nerve palsies may be partial or complete, congenital or acquired, isolated or accompanied by signs of more extensive neurological involvement. They can result from lesions located anywhere from the oculomotor nucleus to the termination of the third nerve in the extra ocular muscles within the orbit. Recent advances in noninvasive neuroimaging facilitate early diagnosis; however, management of a patient presenting with an isolated third nerve palsy remains a challenge. PMID- 10874778 TI - Clinical assessment of complex visual dysfunction. AB - The study of patients with lesions of the central visual system has shown that certain complex visual disturbances are generally associated with lesions in the ventral occipital lobe and adjoining temporal lobe, while other disturbances are more commonly associated with lesions of the dorsal occipital cortices and adjoining parietal lobe. Cerebrovascular lesions in the distribution of the posterior cerebral artery, tumor, trauma, and neurodegenerative processes such as Alzheimer's disease are common etiologies of these disturbances and can be assessed using modern neuroimaging techniques. The corresponding visual function deficits can be separated out by a systematic clinical approach incorporating visual sensory and cognitive testing procedures, as outlined below. PMID- 10874779 TI - Idiopathic intracranial hypertension: mechanisms of visual loss and disease management. AB - Idiopathic intracranial hypertension (IIH) is a disorder of increased intracranial pressure of unknown cause. It is a disorder, predominantly of overweight women in the childbearing years. The major morbidity of the disease is visual loss. Damage to the visual system occurs at the optic nerve head. This damage is most likely due to axoplasm flow stasis and resultant intraneuronal ischemia. Management of IIH begins with educating the patient about the disease and its potential outcomes. I recommend modest dieting and following a low-salt regimen with caution against overuse of fluids. Acetazolamide and Lasix appear to be efficacious. Patients failing medical therapy have optic nerve sheath fenestration performed if visual loss is the main morbidity. Shunting procedures are considered if headache is the main symptom. Most patients respond well to therapy, but idiopathic intracranial hypertension may recur throughout life. PMID- 10874780 TI - Optic neuropathies for the neurologist. AB - Before embarking on expensive ancillary testing, it is crucial for the neurologist to distinguish visual loss due to optic nerve dysfunction from other causes of visual loss. This can usually be accomplished based on specific features of the history and bedside examination. Once it has been established that a patient has some form of optic neuropathy, several clinical features are helpful in determining the etiology. The most important of these is the time course. Other factors include presence or absence of pain, pattern of visual loss (particularly visual field defects), and funduscopic appearance. In most cases, by using this information it is possible to differentiate among the common forms of optic neuropathy: papilledema, ischemic optic neuropathy, optic neuritis, compressive lesions, toxic/nutritional deficiencies, and hereditary forms. This article also reviews recent information concerning the evaluation and treatment of optic neuritis, how to recognize conditions that mimic optic neuritis (e.g., neuroretinitis, papillophlebitis), distinguishing arteritic from non-arteritic AION, and new developments in the genetics of Leber's Hereditary Optic Neuropathy. There is also a discussion of various forms of toxic/nutritional visual loss including Cuban Epidemic Optic Neuropathy and visual loss due to commonly prescribed medications. PMID- 10874781 TI - Visual perceptual abnormalities: hallucinations and illusions. AB - Visual perceptual abnormalities may be caused by diverse etiologies which span the fields of psychiatry and neurology. This article reviews the differential diagnosis of visual perceptual abnormalities from both a neurological and a psychiatric perspective. Psychiatric etiologies include mania, depression, substance dependence, and schizophrenia. Common neurological causes include migraine, epilepsy, delirium, dementia, tumor, and stroke. The phenomena of palinopsia, oscillopsia, dysmetropsia, and polyopia among others are also reviewed. A systematic approach to the many causes of illusions and hallucinations may help to achieve an accurate diagnosis, and a more focused evaluation and treatment plan for patients who develop visual perceptual abnormalities. This article provides the practicing neurologist with a practical understanding and approach to patients with these clinical symptoms. PMID- 10874782 TI - Neuro-ophthalmic complications of sarcoidosis. AB - Sarcoidosis is a systemic disorder that is caused by granulomatous changes of uncertain etiology and commonly has multiorgan involvement. Ocular involvement may occur in up to 32% of persons afflicted by sarcoid. The neuro-ophthalmic manifestations of sarcoid, in particular, are varied and may affect any portion of the visual system, including neural structures. Diagnosis is often difficult due to the fact that the clinical presentation can mimic other disorders, such as Multiple Sclerosis, and therefore a systematic approach to testing must be used once the diagnosis has been considered. The importance of diagnosing neuro ophthalmic sarcoid lies in the fact that it is a treatable disease. The mainstay of treatment is corticosteroids although other immunosuppressive agents may be used. The long-term prognosis of neuro-ophthalmic sarcoid has not been studied in large patient populations, but the data that is available suggests that remission may occur in up to 47%. PMID- 10874784 TI - Random comments: neurologists and neuro-ophthalmology; the "ocular motor" system; and update on ophthalmoplegic migraine. AB - In this brief commentary, I recount the entry of neurologists into the field of neuro-ophthalmology, which was previously regarded as exclusively within the purview of ophthalmology. I then quickly describe the difference between "oculomotor" and "ocular motor," before summarizing "ophthalmoplegic migraine," a rare condition with a rich history that engendered a heated debate among legendary neurological luminaries in 1946. PMID- 10874783 TI - Visual syndromes as the presenting feature of degenerative brain disease. AB - The symptoms of a degenerative brain disease are dictated by its topography. Visuo-spatial impairment may be a severe and early feature of degenerative dementia. Visual symptoms in such patients are broadly divisible into dorsal and ventral visual syndromes, which result from a degenerative focus in occipito parietal and occipito-temporal visual association cortices, respectively. The dorsal visual syndrome includes asimultanagnosia and Balint's syndrome. The ventral visual syndrome includes alexia and visual agnosia (prosopagnosia). Less often, hemineglect or visual field defects result. When Alzheimer's disease and Creutzfeldt-Jakob disease present in this way there is a topographic shift of neurodegenerative changes to posteriorly situated cortices. Patients with corticobasal ganglionic degeneration often develop symptomatic involvement of contiguous sensorimotor cortices causing mixed perceptual-motor syndromes. Even in patients with more typical patterns of dementia, the degree of visuo-spatial impairment may hinder driving skills, and the issue of driving should be addressed early in the clinical course. PMID- 10874785 TI - [Intraocular foreign bodies in the posterior eye segment]. AB - BACKGROUND: A penetrating eye injury due to an intraocular foreign body (IOFB) may result in a poor vision and even in loss of the eye. A proper analysis of the cause of injury and of the injured eye enables a correct decision to be made concerning timing and method to be used not only for foreign body removal but also concerning of all sight saving surgical procedures. PATIENTS AND METHODS: Over the years 1989-1993, 51 patients with an IOFB in the posterior segment of the eye were treated at the Department of Ophthalmology of the Comenius University in Bratislava. One patient was lost from the long-term observation and in two patients was an intraocular foreign body not removed. We evaluate results in 48 patients. The operative techniques used by foreign body removal and by reoperations are mentioned. The occurrence of peroperative and postoperative complications and the final anatomical and functional results are evaluated. An average follow-up period was 40.7 months. The value of the following prognostic factors was considered in relation to the final visual acuity: size and location of the laceration of the eye, size of IOFB and time of IOFB removal. The statistical significance was tested by Chi-square. Fischer's coefficient for tetrachoric tables was used for the calculation of power of dependence. RESULTS: The foreign body was removed from 64.6% of the eyes after pars-plana vitrectomy. Foreign body caused serious damage of the intraocular structures in 37.5% of the eyes. The poor anatomical result was achieved in 18 (37.5%) and good in 30 eyes (62.5%). In twelve eyes (25%) was implanted an IOL. The final visual acuity of 6/9-6/6 was achieved in 18 (37.5%) and visual acuity lower than 2/60 was recorded in 21 (43.7%) of the eyes. The prediction of bad visual acuity (lower than 2/60) was significant related to: all lacerations except of corneal wounds smaller than 4 mm, IOFB size exceeding 3 x 2 mm. CONCLUSIONS: Management of retained intraocular foreign bodies should be individual and is dependent on the extent of the initial injury and the characteristics and location of the IOFB. The final outcome depends mostly on the extent of the primary injury and of the occurrence of the peroperative and postoperative complications. PMID- 10874786 TI - [Problems in diagnosis of intraocular foreign bodies in the posterior eye segment]. AB - BACKGROUND: To evaluate the application of the available diagnostic procedures in intraocular foreign bodies impacted in the posterior segment of the eye. MATERIALS AND METHODS: Between 1989-1993 were treated 51 patients and between 1994-1998 were treated 74 patients with intraocular foreign bodies in the posterior eye segment at the Department of Ophthalmology, Medical faculty of the Comenius University in Bratislava. It was evaluated: time of the first medical examination, cause of the injury, time of the right diagnostic and used ancillary testings. In the second period (prospective study) was evaluated also foreign body trajectory in the posterior segment of the eye. The influence on the final visual acuity was evaluated in no proper diagnosed patients. RESULTS: 50.9% of patients in the first period and 55.9% of patients in the second period were injured while at work. Foreign body was metallic in 96% and 94.7% respectively. In the first period 12 patients were injured by various explosions and in the second period 16 patients were injured by working with some circulating tools. The most common ancillary testing was plain film testing and Comberg localisation. In the second period was intraocular foreign body trajectory defined as indirect in 24 eyes (32%). The foreign body achieved its final intraocular position after simple or double contact with the retina. Foreign body was unproperly diagnosed in 14 patients. The final visual acuity was negative influenced by this fact in 8 patients. CONCLUSIONS: A history and ocular examinationare are still most important in diagnosis of the penetrating eye injury with intraocular foreign body. The basic ancilary testing is simple plain film. Ultrasonography is helpful in diagnosis of other intraocular pathological changes. In justified cases is also possible to perform computed tomography. The peroperative intraocular diagnostics is very important for treatment of the all injured intraocular structures. PMID- 10874787 TI - [Ultrastructure of the lens capsule in pseudoexfoliation syndrome]. AB - The authors examined, using transmission electron microscopy, the lenticular capsule obtained during phacoemulsification by circular capsulorrhexy in pseudoexfoliation syndrome. The assembled findings indicate degenerative changes of the lenticular capsule, manifested by destruction of the surface layer. PMID- 10874788 TI - [Refractive procedures--LASIK and intraocular pressure in myopic eyes]. AB - PURPOSE: To evaluate the effect of central corneal thickness (CCT) on the measurement of intraocular pressure (IOP) in normal myopic eyes before and after LASIK refractive surgery. METHODS: To determine CCT (by ultrasonic pachymeter NIDEC) and IOP (by Goldmann tonometer) in 27 patients (48 myopic eyes from -4 to 14 D) before LASIK (on Chiron Technolas 117) and 3 months after LASIK surgery. RESULTS: Mean CCT before LASIK was 543.8 microns (480-612), 3 months after was 430.6 microns (371-511), the difference--113.2 microns (48-157) is highly significant (p < 0.0001). Similar significant is also the decrease of mean IOP 3 months after surgery--about 5.4 mmHg (p < 0.0001). After LASIK surgery decrease IOP is 4.8 mmHg/100 microns. CCT is an important factor for accuracy of the Goldmann measurement. Myopic eyes need more attention, especially after photorefractive surgery, when CCT is decreased and therefore Goldmann IOP measurement is permanently underestimated. PMID- 10874789 TI - [Evaluation of results of cataract surgery in patients 90 years and older (in a group of long-lived individuals)]. AB - In a group of 71 patients with cataract aged 90 years or more operated between July 1 1997 and December 31 1998 at the Ophthalmological Clinic, Faculty Hospital Hradec Kralove (operations of 79 eyes) a modified questionnaire was used based on VF 14, to assess the benefit of cataract surgery in patients of this age. Replies were obtained only from 38 questionnaires, completed by the patient with the assistance of relatives. Fourteen questionnaires were supplemented by a letter where the relatives described their view on the benefit of the operation for the patient. To the remainder the questionnaire could not be delivered. Improvement of the quality of life after surgery was recorded in 32 instances, once deterioration and four times the condition did not change. The majority of patients (33, i.e. 87%) would, if there was an opportunity have the operation again, while four patients would not have it and one patient did not know. Two of four patients who would not have another operation had severe senile macular degeneration (SMD) of the fundus, another two suffer in addition to SMD from glaucoma (G) with loss of visual functions already before surgery. The questionnaire method gives a better idea of the benefit of surgery for the patient than mere evaluation of visual acuity. From the results ensues that it is necessary to pay special attention to indications for surgery in patients with severe SMD and indicate surgery very carefully also in other complicating diseases in view of the higher risk of the operation. Patients of this age take a poor effect of cataract surgery very badly, while, on the other hand, a successful operation leads to general improvement of their condition and they respond very favourably. PMID- 10874790 TI - [Results of treatment of retinal detachment with macular holes]. AB - We analysed the results of PPV combined with the silicone oil or expansive gas endotamponade of the retina by 12 patients. The etiology of the retinal detachment with a macular hole were: myopia in 8 cases, emetropia in 4 cases. Reattachment and functional results with visual acuity from 0.5/50 to 6/24 we found by all emetropic eyes. By myopic patients we found 87.5% anatomical success during 40 months, functional result of these patients advises on the degenerative myopic changes. PMID- 10874791 TI - [Use of permanent intracanalicular silicone plugs in the lacrimal ducts in the treatment of dry eye syndrome]. AB - The authors describe longer than two-year experience with the use of permanent intracanalicular silicone plugs in the treatment of the dry eye syndrome. Between August 1997 and November 1999 the authors selected by means of the test with dissolvable collagen plugs 50 patients to whom permanent silicone intracanalicular plugs were inserted. During the follow up period (1-28 months) no complications were recorded associated with the insertion of permanent intracanalicular silicone plugs. PMID- 10874792 TI - [Surgical treatment of strabismus in the Czech Republic in 1998]. AB - For the first time an anonymous survey by means of questionnaires on the scope and surgical treatment of strabism was used in the Czech Republic in 1998. In 54 departments of the Czech Republic 3167 operations of strabism were made. 80% of the operated patients were children under 15 years of age. The most frequent primary surgical operation was modification of the strength of horizontal muscles. This surgical procedure was implemented in 81.7%. The ratio of primary operations and re-operations in child patients was 10:1. The mean age of child patients during the primary operation of strabism was 4.3 years, on re-operation 7.2 years, the mean period of hospitalization during the primary operation was 4.4 days. Two surgical centres performed more than 20 operations per month. As a matter of routine a surgical microscope was used only by some departments which made less than 5 operations of children suffering from strabism per month. In order to follow up national trends of surgery of strabism the authors will repeat similar investigations in future. PMID- 10874793 TI - [Latex allergy]. AB - The authors describe a case of an allergic affection in a patient with occupational exposure to latex allergens with a history of anaphylactic reaction to poppy seed and reaction to the antigens of apples, oranges, tangerines, peanuts and bananas, revealed by the method CAP Phadiatop. A marked reaction was initiated after the use of a shampoo containing volatile banana oil. The authors emphasize the high incidence of latex allergy, the manifestations of which may be encountered also in clinical ophthalmology. PMID- 10874794 TI - [Clinical experience with the preparation Uniflox Unimed Pharma in ophthalmology]. AB - The authors investigated in a group of 186 adults and 46 children the effect of antibiotic drops UNIFLOX UNIMED PHARMA. The drops contain the effective substance ofloxacine and were used in different inflammations of the eye and adnexa as well as before and after surgery of the eye. Of 110 patients where before surgery from the conjunctival sac pathogenic microorganisms were cultivated, after 7 days following administration of the drops in 105 patients the finding was negative. The marked effect on staphylococci was remarkable. During the postoperative period, in particular after operations of the cornea, a positive prophylactic effect was found and the authors did not observe any effect on healing of the surgical wounds or epithelization of the cornea. The drug had a high antimicrobial effect also in children where it was administered in suppurative dacrocystitis of neonates, acute inflammations of the conjunctiva or bacterial superficial keratitis. All patients tolerated the drug very well, no side-effects were observed. PMID- 10874795 TI - When knowledge is bad. PMID- 10874796 TI - Reflections on the war in Korea. Part I. PMID- 10874797 TI - The role of Delaware physicians. Part. 1. Physicians A-H. PMID- 10874798 TI - Suggested guidelines for the topographic evaluation of implant surfaces. AB - The bone anchorage components of commercially available oral implant systems differ in surface roughness by at least sixfold. Correct reporting of the surface roughness of implant systems is important, since one cannot exclude the possibility that surface roughness will influence clinical results. However, many confusing statements are found in the literature when the surface topography of implants is described. Different measuring instruments and techniques strongly influence the outcome of a topographic characterization. Furthermore, a screw type design introduces problems for most measuring instruments. Without a standard procedure, it is generally impossible to compare values from one study with another. The aim of the present study was to suggest standards for topographic evaluation of oral implants in terms of measuring equipment, filtering process, and selection of parameters. It is suggested that the measuring instrument be able to measure all parts of a threaded implant if the investigation relates to such a design. Preferably, 3-dimensional measurements should be performed. On screw-type implants, tops, valleys, and flanks should be evaluated. At least 3 samples in a batch should be evaluated, filter size must be specified, and at least one of each height, spatial, and hybrid parameter should be presented. PMID- 10874799 TI - Bone response to titanium alloy implants placed in diabetic rats. AB - Although dental implants continue to provide consistent and predictable treatment options for most patients, some people with uncontrolled systemic disease may be denied implant treatment. Diabetes is one such disease. According to the U.S. Centers for Disease Control and Prevention, diabetes is a leading cause of blindness, kidney failure, and amputations of the lower extremities. These complications result from microvascular disturbances associated with diabetes. The effect of diabetes on the healing of titanium implants has not been well established. In this study of 32 rats, diabetes was induced in 16 animals by injection of streptozotocin (65 mg/kg); the remaining 16 animals served as controls. Titanium alloy implants were placed in the tibiae of all 32 rats using standard surgical techniques. Implants healed for 14 days. Blood samples were obtained for serum glucose, osteocalcin, and alkaline phosphatase analyses. Implants were retrieved and processed for histomorphometric analyses. Three quantities were measured using light microscopy, video capture, and computer analysis: percent osseointegration (i.e., linear bone interface), associated bone volume percent, and contact frequency. Diabetic animals demonstrated significantly less osseointegration than controls. However, bone volume percent in diabetic animals was about 4 times greater than controls. Biochemical analyses were mixed; diabetic animals demonstrated increased serum osteocalcin levels compared to controls but decreased alkaline phosphatase. Based on the results of this study, it was concluded that the bone response associated with titanium alloy implants in the tibiae of diabetic rats is uniquely different from controls. PMID- 10874800 TI - Surface characterization of titanium-based implant materials. AB - This study examined the effects of different treatments (polished, electropolished, and grit-blasted) on the surface morphology and chemistry of commercially pure titanium and titanium-6% aluminum-4% vanadium. The structure and composition of the surfaces were evaluated using scanning electron microscopy, atomic force microscopy, energy dispersive spectroscopy, Auger microprobe analysis, and x-ray photoelectron spectroscopy. Surface roughness values at large scales were nearly identical for grit-blasted and electropolished samples, while at smaller scales, electropolished and polished samples had nearly identical quantitative roughness values. The surface oxide compositions were found to be primarily titanium dioxide on both materials for all surface treatments. No vanadium was seen with either x-ray photoelectron spectroscopy or Auger microprobe analysis for the alloy, indicating a possible surface depletion. Calcium was present on the grit-blasted samples, and calcium and chlorine were detected on the electropolished samples. PMID- 10874801 TI - Alveolar ridge repair using resorbable membranes and autogenous bone particles with simultaneous placement of implants: an experimental pilot study in dogs. AB - The aim of this experimental study was to evaluate the use of autogenous bone harvested during preparation of implant sites in combination with resorbable membranes for vertical ridge augmentation under 2 different defect site conditions. Combined vertical/horizontal alveolar bone defects were created by experimentally induced periodontal infections around all premolar teeth in the mandibles of 3 dogs (group 1). In another 3 dogs, fresh surgical defects were created after extraction of all premolar teeth in the mandibles (group 2). In all dogs, 2 implants were placed on each side of the mandible into the defect areas. One implant on each side of the mandible received augmentation with autogenous bone particles, and both implants on one side of the mandible were covered with polylactic acid membranes. After 5 months, the material was evaluated histologically. There was a small but significant increase in bone regeneration in the defects augmented with bone particles with and without membrane coverage in group 1. In group 2, no significant difference was seen between the controls and the augmented sites. The major limiting effect for bone regeneration appeared to be insufficient stability of the bone material to withstand the overlying soft tissue pressure. It was concluded that the placement of autogenous bone particles, either with or without membrane coverage, had little effect on the regeneration of peri-implant bone defects. PMID- 10874802 TI - Lethal photosensitization, autogenous bone, and e-PTFE membrane for the treatment of peri-implantitis: preliminary results. AB - This clinical study reports on the results of a new method in the treatment of peri-implantitis. The surfaces of 24 plasma flame-sprayed cylindric implants in 17 patients who were diagnosed with peri-implantitis were decontaminated with a combination of toluidine blue (100 micrograms/mL) and laser irradiation at a wavelength of 906 nm. Bone defects were filled with autogenous bone using e-PTFE membranes for retention of the grafting material. Premature membrane exposure occurred in all patients after an average of 3 weeks (+/- 10 days), which required immediate removal of the exposed membrane in 1 patient. Since the soft tissue showed minimal signs of inflammation, the membranes were left in situ for another 6 weeks in all other patients. The mean radiographic peri-implant bone gain was 2 mm +/- 1.90 mm after 9.5 months (maxilla 2.5 mm +/- 2.38 mm; mandible 1.9 mm +/- 1.87 mm). Two implants around which the initial bone defect had already reached the basket had to be removed after 10 months and 35 months, respectively, despite radiographic evidence of improvement of the peri-implant defect. The longer the membrane stayed in situ, the more bone was gained, as long as the membrane was covered by soft tissue (P = .01). However, the longer an exposed membrane was left in place, the smaller the resultant bone gain (P = .0001). Therefore, despite the absence of clinical signs of inflammation, exposed membranes should be removed immediately. The short-term results of this study corroborate the efficacy of the applied treatment method in prolonging the service time of dental implants involved with peri-implantitis. PMID- 10874803 TI - Implant-retained mandibular overdentures with immediate loading: a prospective study of ITI implants. AB - A prospective study was conducted in which 21 patients received a mandibular implant-supported overdenture. Eighty-four ITI screw-type implants were placed in the interforaminal area of the mental symphysis (4 implants per patient). Immediately after implant placement, a U-shaped gold or titanium bar was fabricated and implants were loaded with an implant-retained overdenture. Of 21 patients treated, 19 were followed for a minimum of 25 months to a maximum of 60 months, with a mean follow-up of 37 months. Two patients dropped out during the follow-up. The overall failure rate of implants (according to Albrektsson criteria) was 4% (3/76 implants), but all implants, bars, and prostheses remained in function. Results from this study demonstrated that the success rate for immediately loaded mandibular implants is similar to that obtained in cases of delayed loading, after osseointegration has taken place. This method shortens dental rehabilitation time with relevant satisfaction for patients. PMID- 10874804 TI - A prospective study to assess osseointegration of dental endosseous implants with the Periotest instrument. AB - Long-term studies have documented the successful treatment of edentulous and partially edentulous patients with titanium implants. However, the inability to identify some non-osseointegrated implants before occlusal loading is costly to practitioners and patients. This study followed all patients (n = 40) who had implants placed over a 6-month period. The Periotest instrument was used at Stage II surgery, final impression, prosthesis placement, and 6 and 12 months after occlusal loading to quantify mobility/lack of mobility of implants with conventional 1-piece temporary healing abutments in place. The positive predictive value was 64%. The Periotest instrument was able to identify non integrated implants only when measured at Stage II surgery and 12 months after occlusal loading, 64% of the time. However, Periotest values recorded at Stage II surgery are not valid predictors of non-osseointegrated implants 12 months post occlusal loading. PMID- 10874805 TI - Evaluation of a single-tooth implant. AB - Fifty-nine commercially pure titanium implants in 59 subjects were compared with internal control teeth for 3 years. Nineteen coated implants of identical design were placed in 17 of the subjects and compared with the titanium implants. Demographic data, microbial DNA, aspartate aminotransferase levels, Plaque Index, width of adjacent keratinized tissue, probing depths, bleeding on probing, relative attachment levels, mobility, and radiographic bone height were studied. The only statistically significant changes over time were improved plaque scores in the subjects and slight bone loss around the implants. There were no differences between the 2 types of implants. Mobility was less and probing depth and bleeding on probing were greater in the implant sites than in the control sites. PMID- 10874806 TI - Tilting of posterior mandibular and maxillary implants for improved prosthesis support. AB - Rehabilitation of atrophied edentulous arches with endosseous implants in the posterior regions is often associated with anatomic problems such as jaw shape and location of the mental loop, mandibular canal, and maxillary sinuses. The purpose of this investigation was to modify the method for implant placement in the posterior part of the jaws to extend fixed implant-connected prostheses further distally, and to reduce the length of cantilevers in complete-arch prostheses without transpositioning the mandibular nerve or performing bone grafting in the maxilla. Forty-seven consecutive patients were treated with implants (25 patients/36 mandibular implants, 22 patients/30 maxillary implants) placed in tilted positions. They were followed a mean of 40 months (mandibles) and 53 months (maxillae). In the mandible, implants close to the mental foramina were tilted posteriorly approximately 25 to 35 degrees. In the maxilla, the posterior implants were placed close to and parallel with the sinus walls and were titled anteriorly/posteriorly approximately 30 to 35 degrees. Patients gained a mean distance of 6.5 mm of prosthesis support in the mandible and 9.3 mm in the maxilla, as a result of implant tilting. There were no implant failures in mandibles. The cumulative success rates in the maxilla at 5 years were 98% for tilted implants and 93% for non-tilted implants. Paresthesias of the mental nerve were observed on 4 sides during the first 2 to 3 weeks after implant placement. Analysis of the load distribution in one mandibular case showed no significant difference between tilted and the non-tilted implants, and the improved prosthesis support was confirmed. Satisfactory medium-term results concerning osseointegration and significant extension of prosthesis support show that the method can be recommended. This technique may allow for longer implants to be placed with improved bone anchorage. PMID- 10874807 TI - The palatal subepithelial connective tissue flap method for soft tissue management to cover maxillary defects: a clinical report. AB - This article presents a technique for soft tissue reconstruction and covering defects associated with maxillary implant-supported restorations. A pedicle subepithelial connective tissue flap is prepared from the palatal mucosa near the area to be treated and is displaced into the receptor site. The donor site remains primarily covered. An increase in soft tissue volume is achieved at the receptor site, which is advantageous for various reasons. The pedicle graft has been used for different indications: closure of the alveolus after immediate implant placement, papilla reconstruction, defect and dehiscence repair, and multiple-layer closures after bone grafting and treatment of peri-implantitis. Over a 32-month period, 103 patients were treated with this method. Partial flap necrosis occurred in only 2 patients. All other patients showed significant improvement over the preoperative condition. PMID- 10874808 TI - Osseous proliferation of the mandible after placement of endosseous implants. AB - Spontaneous alveolar ridge growth in the posterior region of the mandible following placement of endosseous implants is reported. The study included 27 patients with totally edentulous mandibles and fixed prostheses supported by osseointegrated implants placed between the mental foramina. In 5 patients, an increase in the height of the alveolar crest was observed in the molar region; the increase ranged from 3.3% to 8.6%. This osseous proliferation may be a physiologic response to stress distribution in the molar region. PMID- 10874809 TI - The influence of controlled occlusal overload on peri-implant tissue. Part 3: A histologic study in monkeys. AB - The influence of experimental occlusal overload on peri-implantitis in monkeys (Macaca fascicularis) has been examined to explain the pathology of the disease that develops in the tissue around osseointegrated implants. In the first article of this series, it was reported that bone resorption was not observed around implants when occlusal trauma was produced by a super-structure that was in supraocclusal contact with an excess occlusal height of approximately 100 microns, provided there was no inflammation in the peri-implant tissue. In the second part of the study, experimental inflammation was created in the peri implant tissue, and occlusal overload was produced by a superstructure with an excess occlusal height of 100 microns. Notable bone resorption was observed around the implant with the passage of time. These results suggested that, in addition to the control of inflammation in peri-implant tissue, traumatic occlusion may play a role in bone breakdown around the implant. In the present study, while the peri-implant tissue was kept in an inflammation-free state, bone level changes around the implants were investigated when various levels of traumatic force were exerted. The supraoccluding prostheses were defined as excessively high by 100 microns, 180 microns, and 250 microns, respectively. The heights were determined with an image analysis device, and the bone responses around the implants induced by the traumatic forces were investigated. The results showed that bone resorption around implants tended to increase with 180 microns or more excessive height of the superstructure. This suggests that the threshold of excessive height of the superstructures at which peri-implant tissue breakdown may start is approximately 180 microns. It is also suggested that there is a possibility of bone resorption around the implants caused by excess occlusal trauma, even when there is no inflammation in peri-implant tissue. PMID- 10874810 TI - Immediate one-stage postextraction implant: a human clinical and histologic case report. AB - The placement of an implant immediately after tooth extraction may have the following advantages: reduction in morbidity, treatment time, and treatment costs; preservation of the residual ridge width and height; optimal esthetic result; and easier definition of implant position. The aim of the present study was the presentation of a human clinical and histologic report involving a nonsubmerged implant placed in a mandibular postextraction site and removed because of persistent pain. At low-power magnification, it was possible to see that newly formed bone with wide osteocyte lacunae was present around the implant. A 1.5-mm sulcular epithelium was visible on one side of the implant, with a 0.5-mm epithelial attachment. The thickness of the supracrestal connective tissue was 3.2 mm. This connective tissue was dense, had few cells, was well vascularized, and showed no evidence of an inflammatory infiltrate. Under polarized light, it was possible to observe that the connective fibers were arranged perpendicular to the implant surface and that these fibers became parallel near the implant. These results show that human immediate postextraction implants can have a high percentage of bone-implant contact. PMID- 10874811 TI - Histologic evaluation of hydroxyapatite-coated root-form implant retrieved after 7 years in function: a case report. AB - This case report presents a clinical, radiographic, and histologic evaluation of 2 non-adjacent, hydroxyapatite-coated, root-form implants retrieved from the maxillary canine area of a patient after 7 years in function. Clinical examination revealed immobile implants with no sign of pathosis. Radiographic examination indicated close proximity of the bone to the implant surface without evidence of radiolucency. Histologically, the 2 implants appeared to be well integrated with the surrounding bone; 84% of the surface of the first implant and 79% of the surface of the second implant had close bone apposition at the interface. There was no evidence of dissolution of the hydroxyapatite coating. The bone appeared to be in immediate contact with the coating. These observations suggest that a particular hydroxyapatite coating on root-form implants can resist degradation during long-term function. PMID- 10874812 TI - The Rhode Island Department of Health. PMID- 10874813 TI - The Office of Primary Care at the Rhode Island Department of Health. PMID- 10874814 TI - Assessing and responding to pre/periconception risks--early experience with the Rhode Island Women's Health Screening & Referral Program. PMID- 10874815 TI - Water systems to report drinking water quality to all customers: how can health professionals prepare for the questions that these reports will generate? AB - The ultimate safety of drinking water depends upon protection of source waters and construction and maintenance of reliable drinking water treatment and distribution systems. These objectives require public support. Physicians can encourage their patients to call their water suppliers and advocate for investment in effective treatment systems and support zoning that will protect water supply watersheds and wellheads. The Consumer Confidence Reports are meant to inform consumers about their drinking water supply. Consumers should use the reports to verify that their drinking water meets all health standards and to understand some of the potential threats to their drinking water quality. Physicians may use the reports as an opportunity to discuss the many types of environmental exposures and ways to reduce these exposures. As a crucial component of the public health community, this is your opportunity to encourage your patients to become more aware of their environment and its impact on their health. PMID- 10874816 TI - Utilization review: a practical approach for physicians. AB - Rhode Island has rules and regulations for UR, as well as an appeals process. When the appeals process is used, it is more often than not successful. (Table 1) If questions arise concerning the UR Act, the UR process, the appeals process, the rights of patients, physicians, and reviewers, or the authority of the HEALTH, please contact OMCR. For a copy of the Rules and Regulations for the Utilization Review of Health Care Services (R23-17.12-1-UR), please write: Rhode Island Department of Health Office of Managed Care Regulation 3 Capitol Hill Cannon Building, Room 410 Providence, RI 02908. PMID- 10874817 TI - Communicating public health via the Internet in RI: www.health.state.ri.us. PMID- 10874818 TI - Emphysematous pyelonephritis. PMID- 10874819 TI - Fulminant liver failure due to neuroleptic malignant syndrome. PMID- 10874820 TI - Treatment of atrial fibrillation with warfarin in Rhode Island. PMID- 10874821 TI - The physician's role in public health statistics and surveillance. PMID- 10874822 TI - West Nile virus surveillance and prevention. PMID- 10874823 TI - [The role of preventive medicine in the preservation of health among the population]. AB - Medicine in Russia is based on a prophylactic concept. Preventive medicine develops on firm scientific basis assuming unity of organism and environment and close relationship between the social and the biologic. Present priorities are to solve up-to-date problems in economics and public health management under changing social and economic conditions, to specify fundamental problems in human ecology and environmental hygiene as a basic science for governmental measures in public health care of Russia. PMID- 10874824 TI - [Work conditions and occupational diseases among the workers of brass and bronze production industry]. AB - Zinc oxide is a main occupational hazard in electrosmelting workshops of brass and bronze alloys. Chronic occupational pulmonary diseases in the workshops are zinc exogenous fibrosing alveolitis among smelters and operators of bridge cranes, silicosis among fettlers of electric ovens. PMID- 10874825 TI - [Prognostic evaluation of occupational adaptation progress among miners at the coal mines]. AB - Gradual factorial analysis of heart rhythm regulation parameters revealed individual and typological peculiarities of adaptational phases in miners during 20 years. The studies determined dysadaptation time in individuals with cortex limbic and limbic-stem domination. PMID- 10874826 TI - [Ecological-geographical mapping of ultra-high voltage electric network objects in united energy systems of Russia ("UES of Russia")]. AB - Negatively influencing environment, electric network objects are simultaneously subjected to antropogenous (ecologic) pressure from other industrial objects. Ecologic and geographic mapping (ecologic maps formation) enables the most complete (rational) analysis of ecologic situation in electric network objects and locate them with economical and ecologic reasons. This method is highly informational and demonstrative. PMID- 10874827 TI - [The evaluation of the risk in the development of general pathological syndromes among operators of chemical production industry according to gender]. AB - The authors evaluate risks of general pathologic syndromes in operators of chemical production apparatus. The female operators demonstrated higher risk of arterial hypertension, coronary heart disease, liver and digestive dysfunction, neurologic syndrome, warning signs of boundary psychic disorders. The article proves higher influence of household intoxications on female organism. PMID- 10874828 TI - [Remote sequelae of conditionally small dosages of irradiation (literature review)]. AB - In the article the information concerning epidemiological aspects of radiation (Study the further consequences of low doses of irradiation) at the human population. Here was demonstrated that the difficulty of interpretation of resultates is conditioned by outside and inside causes in answer to the influence of radiation. Also was shown that the human biology, condition of environmental, unhealthy habites, etc. have got the great importance. PMID- 10874829 TI - [Ecological epidemiology in pediatrics]. AB - The authors necessitate precise determination of methodology for epidemiological research in ecologically unfavorable regions. The article deals with standardization principles of epidemiological research protocol, select formation, approaches to detection of regional health problems, quantitative evaluation of ecological risk factors (attributive, relative risks) and their comparative significance (calculation of standardized coefficients of discrimination) in disease formation. PMID- 10874830 TI - [Analysis of dependence of occupational diseases on length of service on the basis of monitoring]. AB - The authors tested a method determining increase in occupational diseases risk according to length of service under occupational hazards. Nonparametric approach proved to be the most informative. According to the findings, to decrease occupational morbidity significantly, the length of service under local vibration and physical overload should be reduced to 5-8 years and that under intensive noise and industrial aerosols--to 9-10 years. PMID- 10874831 TI - [Radiation safety in work with chemicals containing admixtures of natural radionuclides]. AB - The authors analyze results obtained in their own research of radiation situation in several enterprises of Nuclear Energy Department. Those enterprises treat chemicals containing admixtures of natural radionuclides in production of substances previously considered safe--tantalum, zirconium, strontium carbonate. The article necessitates regulation of radiation factor in working environment and control over products and waste materials. The authors base methods providing radiation safety during work with substances containing higher levels of natural radionuclides admixtures. PMID- 10874833 TI - Open to interpretation. PMID- 10874832 TI - [On embryotropic action of XTS-1 plant growth stimulator]. AB - XTC-1 level near 10.1 mg/m3 appeared to approximate to embryotropic activity threshold, but that near 1.3 mg/m3 is inactive. The embryotropic activity occurred on the background of general toxic effects, so the substance does not posses specific embryotropic activity. PMID- 10874834 TI - Advise and consent. PMID- 10874836 TI - Embracing technology. Computers aren't just for billing anymore. PMID- 10874835 TI - Polymorphous low-grade adenocarcinoma of the oral cavity. AB - Polymorphous low-grade adenocarcinoma (PLGA) is a neoplasm arising most commonly within the minor salivary glands of the oral cavity. Not recognized as a distinct entity until 1983, PLGA was often misdiagnosed as adenoid cystic carcinoma or pleomorphic adenoma. PLGA is thought to be the second most common salivary gland tumor after mucoepidermoid carcinoma. Affecting individuals later in life, PLGA often presents as a firm, painless nodule that exhibits a locally aggressive, infiltrative pattern. Because of PLGA's slow growth rate and low rate of metastasis, differentiation from other disease entities is crucial for treatment modalities. The study presented here reviews three cases of PLGA, their treatment and follow-up. PMID- 10874837 TI - Shade selection. Communicating with the laboratory technician. AB - Shade selection for anterior crowns has always set up a communications problem between the dentist and laboratory technician. Over the years, many different techniques have been formulated to help overcome the problem. These techniques include picture taking, drawing diagrams and using multiple porcelain shade guides. However, they have not completely erased the difficulty of communicating the choice of the proper shade of an anterior crown. This was especially true in the 1990's when all-ceramic crowns were introduced. Popular techniques dentists use for communicating shade selections will be reviewed, along with guidelines for making the proper selection. Many dentists are familiar only with the techniques they were taught in dental school and/or residency program and are unaware of the superior methods that can be used. This type of review can be extremely helpful to restorative dentists. PMID- 10874838 TI - Prognostic value of cardiac troponin I release kinetics in unstable angina. AB - BACKGROUND: Cardiac Troponins (cTn) are useful in unstable angina (UA). Moreover the different elevation patterns that can be observed in this condition seem to have different prognostic implications. AIM: To study cTn kinetics and cTn nadir in patients with UA, defined as angina at rest within the last 24 hours before admission accompanied by ischemic ECG changes and no myocardial infarction (MI) enzymatic criteria. POPULATION AND METHODS: Samples were collected from 156 patients for cardiac enzymes and cTnI at admission and at 6, 12, 18 and 24 hours. The chemilluminescence method (Access/Sanofi Pasteur) was used for cTnI. The primary end-point at 30 days was the combined occurrence of death, MI and recurrent ischemia. RESULTS: All determinations were below 0.10 ng/ml (group N) in 114 patients and the other 42 pts (group P) had at least one value equal to or above 0.10 ng/ml. The primary endpoint was observed in 24.6% of group N pts compared with 45.2% of group P pts (p = 0.02). Three different patterns of cTnI kinetics were observed. This enabled the identification of a subgroup--group N pts with increasing cTnI values within the first 12 hours and a total differential value > or = 0.03 ng/ml--with an increased risk (50.0% versus 21.4%- p = 0.02--Kaplan-Meier test). CONCLUSION: Besides the prognostic value conferred by cTnI elevation, cTnI kinetics analysis established another sub-group of patients with an adverse prognosis at 30 days follow-up, despite having a negative cTnI. PMID- 10874839 TI - Prognostic value of cardiac troponins in patients with acute coronary disease. PMID- 10874840 TI - Identification of an Arg403Gln beta myosin heavy chain gene mutation in a Portuguese family with hypertrophic cardiomyopathy. AB - INTRODUCTION: The etiology of Familial Hypertrophic Cardiomyopathy (HCM) is attributed to the mutation of genes that encode sarcomeric proteins in the heart. Until now no gene mutations had been identified in Portuguese families with HCM. OBJECTIVE: The main objective of this study is to describe a Portuguese family with HCM carrying an Arg403Gln mutation in the beta myosin heavy chain gene. METHODS: With the help of several Molecular Biology tools, 40 families with HCM were studied. In all these families, one member was identified as carrying an Arg403Gln mutation in the beta myosin heavy chain gene. All family members were submitted to a physical exam, EKG and echocardiography. Those carrying a gene mutation were also submitted to Holter monitoring and to magnetic ressonance imaging. RESULTS: Molecular biology techniques are extremely important for the diagnosis of HCM, particularly in healthy carriers. CONCLUSION: The use of molecular diagnostic tools in HCM is very useful because it allows us to identify the healthy carriers and establish earlier clinical and prevention programs for these individuals. PMID- 10874841 TI - Tissue Doppler imaging: clinical topics for the new millenium. AB - INTRODUCTION: Tissue Doppler imaging is an echocardiographic technique that allows the selective visualization and quantification of myocardial signals. Its aim is to complement the conventional Doppler study, adding more and better information on specific topics of cardiovascular diseases. During the first seven years of the technique, much scientific work was produced and some clinical applications of the method have emerged. CLINICAL APPLICATIONS: I--non ischemic heart disease: The technique has been widely used in diastology, in hypertrophic cardiomyopathy, in heart transplant patients and in arrhythmology. II--ischemic heart disease: The quantitative assessment of regional diastolic and systolic function makes the technique very promising during stress echocardiography and during percutaneous transluminal coronary angioplasty. CONCLUSIONS: In 2000, tissue Doppler echocardiography is still a young, exciting and promising technique. Despite the fact that much has already been done, there is still a long way to go, implying a great amount of time and personal investment. How often do we feel that we are building a small part of the future? PMID- 10874842 TI - [Plasma total anti-oxidant status in young survivors of myocardial infarction]. AB - Free oxygen radicals are involved in the endothelial lesion process which leads to the formation of the atheroma plaque and thrombosis. There is some evidence that antioxidant therapy may be beneficial in coronary heart disease prevention. Our objective was to study the plasma total anti-oxidant status in young survivors of acute myocardial infarction. POPULATION: 23 patients, mean age 35.2 years (22-40) admitted for acute myocardial infarction from January 1995 to June 1998 (20 males). RISK FACTORS: Tobacco smoking 22/23, systemic arterial hypertension 4/23, hypercholesterolemia 17/23, positive family history for coronary heart disease 5 patients, previous history of angina 4 patients, none of these patients had diabetes mellitus. The location of the infarct was anterior in 12 patients, inferior in 10 patients and non-Q wave in one patient. Blood samples were drawn after overnight fasting and the plasma total antioxidant status (TAS) was determined by a colorymethric method (Trolox equivalent). The mean time elapsed since the acute myocardial infarction until sample collection was 16.5 +/ 10.7 months. RESULTS: 18 patients had low TAS values, mean 1.23 +/- 0.11 mmol/L (below the reference values: 1.3-1.77 mmol/L). CONCLUSIONS: In this group of patients, the plasma total antioxidant capacity was globally decreased, which may constitute a risk factor for coronary heart disease. PMID- 10874843 TI - [Influence of smoking on homocysteinemia at baseline and after methionine load]. AB - INTRODUCTION AND AIMS: Homocysteinemia (HC) and smoking are both important risk factors for vascular disease. In the present study, we intend to evaluate the influence of smoking habits on HC values as well as on vitamins B6, B12 and folic acid, co-factors of HC metabolism. METHODS: We measured fasting homocysteinemia (basal) and homocysteinemia 6 hours after an overload with 0.1 g methionine/kg body weight in 279 subjects. We also performed the dosage of plasma levels of B6 and B12 vitamins and of red cells folates. Smoking habits were inquired and the subjects were classified as non-smokers, current smokers or ex-smokers (if they had stopped smoking more than 1 month before the study). According to the smoking status, smokers were classified in three groups: less than 20 cigarettes a day, between 20 and 39 and 40 or more cigarettes a day. We studied basal and after methionine load homocysteinemia, B6, B12 and folic acid levels in each group. RESULTS: Smokers presented significantly higher levels of basal and after methionine load homocysteinemia then non-smokers (10.6 +/- 4.9 vs 9.4 +/- 2.6, and 26.8 +/- 10.0 vs 24.3 +/- 7.4 mumol/L, respectively, p < 0.05 for both and B6 levels (29.2 +/- 12.0 versus 32.6 +/- 12.0 mumol/L, p < 0.05). B12 and folic levels were similar in the two groups. These results were quite similar either in the normal subjects or in the subjects with a history of a cardiovascular event. The subjects who smoked 40 or more cigarettes per day, compared with those who smoked less then 20 cigarettes per day, presented higher levels of basal homocysteinemia (12.4 +/- 2.9 vs 10.0 +/- 5.5 mumol/L, p < 0.05) and lower levels of B6 (24.7 +/- 8.1 vs 31.7 +/- 12.6 mumol/L, p < 0.05). CONCLUSIONS: Smoking habits are related with the increase of basal and after methionine load homocysteinemia, probably because of a decrease in B6 vitamin levels. There is a proportional effect between the number of cigarettes smoked, B6 depletion and basal homocysteinemia increase. This study suggests that B6 vitamin supplements for smokers could decrease the vascular risk related with smoking habits. PMID- 10874844 TI - [Diastolic dysfunction and left ventricular hypertrophy in familial amyloidotic polyneuropathy: a cause-effect relationship?]. AB - TTR Met30 Familial Amyloidotic Polyneuropathy of the Portuguese type (FAP) is an incapacitating and lethal hereditary disorder that affects predominantly young adults of both genders. Portuguese type FAP patients have sensory, motor and autonomic polyneuropathy. The generalised systemic amyloid infiltration involves the heart, leading to the characteristic granular bright sparkling echocardiographic pattern. LV wall thickening occurs in the late phases of the disease. LV diastolic dysfunction has been reported in the absence of systolic dysfunction; an abnormal diastolic transmitral flow pattern assessed by pulsed wave Doppler (PW) was described. PW is very much dependent on load conditions. Tissue Doppler imaging (TDI) has been used as a more reliable method to assess long axis diastolic function. OBJECTIVE: 1--To identify the incremental value of TDI in the assessment of diastolic function in FAP. 2--To correlate diastolic pattern abnormalities and left ventricular mass index (LVMI) in FAP patients. METHODS: We performed a prospective evaluation of 24 consecutive FAP patients and selected 14 (sinus rhythm, age < 45 years). Diastolic function was assessed by PW and classified as normal (GI-E/A > 1) or abnormal (GII-E/A < 1). TDI was performed in 4 sites of the mitral annulus (septum, lateral, inferior, anterior). Velocities of the rapid filling wave (E') and atrial contraction wave (A') were measured and E'/A' calculated. In each site we considered the TDI as normal (E'/A' > 1) or abnormal (E'/A' < 1). The LVMI was calculated by Devereux's formula. RESULTS: Age, gender and heart rate were similar in both groups. TDI at the septal mitral annulus was normal in all of the GI patients (E'/A': 1.29 +/- 0.19) and suggestive of abnormal LV relaxation in all of the GII patients (E'/A': 0.82 +/- 0.11, p < 0.0001). TDI revealed abnormal diastolic pattern when a restricted number of sites of the mitral annulus were assessed, even in GI patients and before PW abnormalities occurred. Fractional shortening (FS) and LVMI were similar in GI and GII (FS-GI: 45.5 +/- 5.3, GII 43.5 +/- 8.1%, p: NS; LVMI--GI: 66 +/- 9.3, GII: 67 +/- 3.0 g/m2 p: NS). CONCLUSION: The assessment of mitral annulus motion has introduced new data in the study of diastolic function of FAP patients. An abnormal LV relaxation pattern occurred early in the evolution of the disease in patients with normal LVMI and systolic function. PMID- 10874845 TI - [Non-infectious thrombotic endocarditis: a case report]. AB - A 48 year-old female patient, admitted with an ischemic stroke, had a transesophageal echocardiogram (TEE) that revealed a dense vegetation in the noncoronary aortic cusp, which was considered the probable source of embolism. The clinical investigation did not identify any other disease or infectious process. However the lack of histologic proof, the absence of fever, the fact that the blood cultures were persistently negative, the patient recovery with no need of antibiotics, and the results of the TEE, are highly suggestive of the presence of a non infectious thrombotic endocarditis. After 18 months of anti platelet treatment, the patient showed no new embolic episodes and TEE demonstrated the resolution of the vegetation. PMID- 10874846 TI - [Transient cortical blindness after cardiac catheterization in a post-surgical coronary patient]. AB - Transient cortical blindness is a known complication after cerebral and vertebral angiography although is a rare complication following cardiac catheterization. In this report, we describe a seventy-six-year-old male patient who had cortical blindness after coronary bypass graft angiography. The development of cortical blindness, in this setting, appears to be an osmotic disruption of the blood brain barrier which is selective for the occipital cortex. Other hypotheses, such as an immunological reaction to the contrast agent or a direct injection of dye into the vertebral artery system, were considered. The clinical outcome is usually favourable, with vision completely restored within 24-48 hours, as was the case of the patient described in the present report. PMID- 10874847 TI - Aortic ring abscess. PMID- 10874848 TI - Restrictive cardiomyopathy due to hypereosinophilic syndrome. PMID- 10874849 TI - [Aortic valve replacement after 80 years of age: short and medium-term results in a series of 140 patients]. PMID- 10874850 TI - Potential benefits of imidazoline receptors agonist drugs in treatment of systemic hypertension. PMID- 10874851 TI - The sympathetic system in essential hypertension. PMID- 10874852 TI - The sympathetic nervous system in human hypertension. PMID- 10874853 TI - [New perspectives in the treatment of hypertension: the contribution of rilmenidine]. PMID- 10874854 TI - Clinical benefits of rilmenidine. PMID- 10874855 TI - The contribution of I1-selective agents in the management of syndrome X. PMID- 10874856 TI - [Nutraceuticals: marketing concept of new science]. PMID- 10874857 TI - [Diabetes mellitus in dogs]. PMID- 10874858 TI - [Clinical trials: principles of the method]. AB - Comparative judgement, which is seminal to any kind of science performing measurements, has been applied to clinical reasoning for many centuries. The need for systematizing the observational methods used in medicine in order to draw more reliable inferences about the effects of therapies has been active all along the 19th century. This has resulted in controlled studies which yielded important advances in clinical and therapeutic knowledge, although their designs were not fully satisfactory. Clinical trials have gained their status of "hard science", methodology allowing causal inference, by the end of the 1940s after having adopted the statistical theories developed in the 1930s by Fisher for experimental design in agronomy. A long way has been run since the first controlled randomized trial. However, half a century later, modern clinical trial remains essentially a controlled randomized prospective study using methods to limit potential biases and to establish statistical significance. PMID- 10874859 TI - [Limitations of clinical trials]. AB - The major limitation of randomized clinical trials is their restriction to interventions that are supposed to have a positive effect. Another limit is related to the difficulty to interpret or generalize the results because the studied population is very different from the population treated in normal life. Participating in a trial may also influence the results. Limitation also includes the specificity of the questions answered; the narrow perspective of many trials leaves aside important information related to the consequences of the intervention on quality of life, satisfaction or costs. Clinical trials usually do not provide the answers to the questions asked by practitioners and deciders. A solution consists in developing disease management approach which may include thousands of patients with the implementation of trials in real life with a long duration of follow-up. PMID- 10874860 TI - [Reading a clinical trial report]. AB - To improve medical knowledge by reading clinical trial reports it is necessary to check for the respect of the methodological rules, and to analyze and criticize the results. A control group and a randomisation are always necessary. Double blind assessment, sample size calculation, intention to treat analysis, a unique primary end point are also important. The conclusions of the trial are valid only for the population included and the clinical signification of the results, depending on the control treatment, has to be evaluated. Respect of the reading rules is necessary to assess the reliability of the conclusions, in order to promote evidence-based practice. PMID- 10874861 TI - [The results of a single clinical trial-is it sufficient information?]. AB - Meta-analysis is a systematic and quantified synthesis of all clinical trials studying the same question. It is systematic because it implies an exhaustive search for all trials of the studied treatment, favourable or not, published and unpublished. It is quantified because it is based on statistical calculations leading to an estimation of the degree of the effect of the treatment. The use of statistical techniques allows to take into account the fact that different results can be obtained in different studies only by chance. Meta-analysis can be used to assess the efficacy of a treatment or to study the factors that modify this efficacy. PMID- 10874862 TI - [In a clinical trial, method is not sufficient, the purpose must be relevant]. AB - The aim of a trial is relevant if it is well formulated, has a strong scientific basis and if anticipated results are useful for patients' management. The aim must be simple and specific, clearly expressing the treatments to be compared, the study population and the primary endpoint. Moreover, the formulation must include the working hypothesis: superiority or equivalence/non inferiority. Endpoints must be valid and have either a pathogenic relevance (explanatory attitude in early phases of development) or a clinical relevance (pragmatic attitude in later development). A synthesis of available knowledge is crucial to avoid useless trials. The aim of a trial must be the result of a consensual and multidisciplinary reflection prior to the beginning of the study. PMID- 10874863 TI - [Good clinical practice:impediment of source of progress?]. AB - On the bases of the Declaration of Helsinki (1964), the law called Huriet Serusclat (1988) and several subsequent decrees have clearly defined the appropriate organization and management of clinical trials in France. The law on the Commission de l'informatique et des libertes (1978) and several European regulations have also to be taken into account. This extensive legal and regulatory system has been the source of a major improvement in the scientific quality of clinical trials and in the security of participating persons. Indirectly, it has also improved daily clinical care, as physicians have learned how to inform their patients and have been trained to use protocols and standard operative procedures. However, the legal and regulatory pressure has increased the cost and complexity of trials to such an extent that no clinician with his own resources is now able to be the sponsor of a trial involving more than a few patients. In addition to trials organized by pharmaceutical companies for drug development, health authorities in France and Europe should consider to support major clinical trials on therapeutic strategies with public health implications. A few examples can be acknowledged thus far, but a real policy remains to be defined. PMID- 10874864 TI - [Randomize the first patient?]. AB - Therapeutic innovation requires rigorous assessment, to fulfil regulatory and ethical constraints. The ideal time for this assessment in the therapeutic progress story is a complex question, mixing individual and collective ethical principles. Randomisation of the first patient is always, at least theoretically, a current issue, extending beyond drug research to reach the wider field of technical innovation. Broad information of consumers, their involvement in clinical research policy, and specific training for prescribers, could be the conditions to make clinical research ethically acceptable. PMID- 10874865 TI - [From clinical trial to prescription]. AB - Clinical trials produce factual data that should be put in practice. This requires that information drawn form their results be transferred to doctors without bias, under a format suited to their training and meeting the constraints of their daily practice. Today this critical step is poorly efficient, despite the availability of techniques and methods appropriate for accessing such information and to tailor it to each patient. Furthermore, regarding many conditions encountered in medical practice, evidence is lacking or is of low strength. Doctors should revise their practice according to the information yielded by clinical trials. PMID- 10874866 TI - [Managed care: definition(s) and suspicions]. PMID- 10874867 TI - [Hematuria. Diagnostic orientation]. PMID- 10874868 TI - [Inhalation corticosteroids. Utilization principles and rules]. PMID- 10874869 TI - [Myogenous syndrome. Diagnostic orientation]. PMID- 10874870 TI - [Normal and pathologic scarring. Physiopathology and anatomic pathology]. PMID- 10874871 TI - [Fever on return from a tropical country. Diagnostic orientation]. PMID- 10874872 TI - [Analgesics. Utilization principles and rules, posology of morphine and its derivatives]. PMID- 10874873 TI - [Auricular fibrillation. Diagnosis, complications, treatment]. PMID- 10874874 TI - Kikuchi-Fujimoto disease. Is Epstein-Barr virus the culprit? PMID- 10874875 TI - Probabilities in the cytologic diagnosis of thyroid gland lesions. PMID- 10874876 TI - Lowenstein-Jensen media. No longer necessary for mycobacterial isolation. AB - Isolation of mycobacteria on Lowenstein-Jensen media (LJ) and in the BACTEC MB9000 (MB) system was compared. Of 2,271 specimens, 317 were positive for 331 mycobacteria isolated in 1 or both media. The MB was positive in 238 isolates, and LJ was positive for 239 isolates; 92 isolates were detected by MB only and 93 by LJ only. Of the 331 isolates, 146 were recovered by both media. MB recovered 38 of 38 Mycobacterium tuberculosis complex isolates, while LJ recovered 23. MB recovered 94.1% (96/102) of Mycobacterium avium complex isolates and LJ 69.6% (71/102). The MB recovered 81% (65/80) Mycobacterium fortuitum-chelonae isolates and LJ 68% (54/80). Of the remaining species, MB isolated 39, while LJ isolated 91. Only 1 organism that was isolated on LJ alone was medically significant based on medical record review. The addition of LJ media to the MB9000 system is not warranted, as it causes clinically irrelevant workload, increased expenditures for the laboratory, and could cause the inappropriate treatment of patients. PMID- 10874878 TI - Primary marginal zone lymphoma of the thymus. AB - Primary low-grade B-cell lymphomas of the thymus are rare, with only 7 reported cases in the literature. We describe 3 cases of primary low-grade thymic lymphoma. All had histologic features of extranodal marginal zone lymphoma and were composed predominantly of small lymphocytes with variable components of monocytoid cells and plasma cells. Overt transformation to large cell lymphoma occurred in 1 case. The neoplastic cells were immunoreactive for the B-cell marker CD20 and were positive for bcl-2 in 2 cases. Two of 3 patients had a long standing history of autoimmune disease. Based on these findings and those of previously reported cases, marginal zone lymphoma is the predominant type of low grade thymic B-cell lymphoma. These tumors seem to be more common in patients with autoimmune disorders, and as observed with marginal zone lymphoma arising at other anatomic sites, they may undergo transformation to a higher grade lymphoma. PMID- 10874877 TI - Virus infection in patients with histiocytic necrotizing lymphadenitis in Taiwan. Detection of Epstein-Barr virus, type I human T-cell lymphotropic virus, and parvovirus B19. AB - The relationship of Epstein-Barr virus (EBV), type I human T-cell lymphotropic virus (HTLV-I), and parvovirus B19 to histiocytic necrotizing lymphadenitis was studied prospectively in 10 Taiwanese patients using materials obtained by fine needle aspiration and lymph node biopsy. The presence of EBV was detected by in situ hybridization for EBV-encoded RNA expression. Immunocytochemistry was used to detect virus-encoded protein for EBV and parvovirus B19. DNA in situ hybridization and polymerase chain reaction were performed to determine the existence of HTLV-I provirus. Expressions of EBV-encoded RNA and Fas ligand were detected in all cases. Expression of EBV-encoded protein was identified in only 1 case. Neither HTLV-I nor parvovirus B19 was detected in any case. PMID- 10874879 TI - Large cell transformation of Sezary syndrome. A conventional and molecular cytogenetic study. AB - Hyperdiploidy sometimes is found in mycosis fungoides-Sezary syndrome, but its diagnostic significance remains undefined. We report an unusual case of Sezary syndrome manifesting with leukemic large cell transformation. Conventional karyotypic analysis showed the presence of a near-tetraploid neoplastic clone. With dual-color cytometric analysis, we showed that the large Sezary cells were near-tetraploid with a DNA index of 1.86, thereby demonstrating a direct relationship between cell size and ploidy. Comparative genomic hybridization further showed chromosomal imbalances that were not revealed on conventional karyotyping. Our findings suggest that hyperdiploidy may be a marker of large cell transformation, so that when this karyotypic abnormality is found in mycosis fungoides-Sezary syndrome, a search for such a complication is indicated. PMID- 10874880 TI - Biclonal chronic lymphocytic leukemia. AB - Chronic lymphocytic leukemia (CLL) is well characterized clinically and immunophenotypically. Demonstration of a monotypic CD19+, CD5+ B-cell population is central to the diagnosis. We report 2 cases of biclonal CLL. Two elderly men were encountered with an absolute lymphocytosis consisting of the typical CD5+, CD19+, CD23+ B-cell population seen in CLL; however, immunoglobulin light chain restriction by flow cytometry was not apparent as B cells expressed kappa or lambda light chains without a clear monotypic population. Molecular genetic analysis of flow cytometry-sorted cells (kappa and lambda populations) revealed in both cases 2 monoclonal B-cell populations. The characterization of these cases and a review of the issues surrounding biclonal CLL are presented. PMID- 10874881 TI - Usefulness of CD79b expression in the diagnosis of B-cell chronic lymphoproliferative disorders. AB - We evaluated anti-CD79b for its usefulness in the diagnosis of B-cell chronic lymphoproliferative disorders (BCLPDs), particularly chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL). We analyzed 100 BCLPDs for CD5, CD19, CD20, CD23, CD79b, and surface immunoglobulin light chain (sIg) expression by 4-color flow cytometry. CD20, CD79b, and sIg expression were quantified. Correlational analysis and univariable and multivariable logistic regression models were used to determine the best combination of antigens for the immunophenotypic classification of CLL vs other BCLPDs. Positive and statistically significant Spearman pairwise correlations between CD20, CD79b, and sIg fluorescence intensity were demonstrated. In the simplest models in which a single variable was considered, cutoff points were chosen that gave misclassification rates for CLL of 16% for CD79b, 19% for sIg, and 18% for CD20. Low-intensity CD79b, CD20, and sIg are associated highly with CLL. A panel containing CD5, CD19, CD23, and sIg allowed correct classification of most cases. Addition of CD20 or CD79b improved diagnostic accuracy; CD79b was slightly better than CD20. CD79b seems to be a useful addition to a standard flow cytometry panel for the evaluation of BCLPDs. PMID- 10874882 TI - Immunohistochemistry can be used to subtype acute myeloid leukemia in routinely processed bone marrow biopsy specimens. Comparison with flow cytometry. AB - Flow cytometry (FC) is the preferred method of immunophenotyping acute myeloid leukemia (AML). However, there are situations in which FC is unavailable and in which immunohistologic staining of bone marrow biopsy specimens can be used to provide immunophenotypic information. To evaluate immunohistologic staining and to confirm its value, we selected 80 newly diagnosed cases of AML that were classified according to French-American-British (FAB) criteria and confirmed by flow cytometric analysis for this study. Paraffin-embedded bone marrow specimens were stained using a panel of antibodies that included CD34 (QBEND10), antimyeloperoxidase (anti-MPO), antihemoglobin, factor VIII-related antigen, and 3 epitopes of CD68 (HAM56, KP1, and PG-M1). Our findings suggest that with the use of the paraffin-reactive antibodies CD34 (QBEND10), MPO, CD68 (PG-M1), antihemoglobin, and factor VIII-related antigen, immunohistochemistry can be used to subclassify AML. Comparison of immunohistochemical results with FC immunophenotyping suggests that there is significant concordance in the results for markers that can be used with both techniques, indicating that the sensitivity and specificity of both methods is comparable (P > .53 in all cases). PMID- 10874883 TI - Flow cytometric detection of CD79a expression in T-cell acute lymphoblastic leukemias. AB - We evaluated the lineage specificity of CD79a in acute leukemias using 3-color flow cytometry in 58 consecutive cases. A panel of cell-surface antigens, including myeloid-associated markers, B-cell-associated markers, and T-cell associated markers, was used. All cases of acute myeloid leukemia were CD79a-, whereas all cases of B-lineage acute lymphoblastic leukemia (ALL) were CD79a+. Three of 8 cases of T-cell ALL showed variable CD79a expression, indicating the presence of a blast subset expressing a relatively high level of CD79a. We investigated the clinical and pathologic characteristics of these 3 cases. All 3 cases had L1 or L2 morphology and expressed surface CD3. None of the other B-cell associated markers were positive, although 1 case expressed CD13 and CD33. Uncommon random karyotypic abnormalities were identified in all 3 cases. Molecular studies demonstrated monoclonal gene rearrangement of T-cell receptor gamma in 2 of 3 cases. All 3 patients were 18 years old or younger; 1 patient did not enter remission, and 1 had disease relapse in 8 months. Our findings provide further support for the existence of a subset of T-cell ALL coexpressing CD3 and CD79a. Further study of the clinical and biologic significance of this subset may be warranted. PMID- 10874884 TI - The (11;14)(q13;q32) translocation in multiple myeloma. A morphologic and immunohistochemical study. AB - We identified 24 cases of multiple myeloma with the t(11;14)(q13;q32). In 22 cases, the t(11;14)(q13;q32) was part of a complex karyotype, and in 2 cases it was an isolated abnormality. All patients had clinical and laboratory features consistent with multiple myeloma. The median degree of plasma cell involvement in the bone marrow was 60%, and in 10 cases, the plasma cells had a lymphoplasmacytoid appearance. Of the 24 cases, 21 had intermediate or high proliferative rates based on labeling index studies. Immunohistochemical studies performed on all bone marrow biopsy specimens showed strong cyclin D1 nuclear positivity in 19 cases. There also was strong cyclin D1 nuclear positivity found in 6 of 30 additional cases without the t(11;14)(q13;q32) demonstrated by routine cytogenetics. The t(11;14)(q13;q32) in multiple myeloma results in overexpression of the cyclin D1 protein, which can be demonstrated by immunohistochemical stain. The cyclin D1 stain results in the additional cases of multiple myeloma suggest that the t(11;14)(q13;q32) may be more common than previously thought and may be missed by routine cytogenetics, particularly if the proliferative rate is low. PMID- 10874885 TI - T-cell clonality determination using polymerase chain reaction (PCR) amplification of the T-cell receptor gamma-chain gene and capillary electrophoresis of fluorescently labeled PCR products. AB - We compared the effectiveness of polymerase chain reaction (PCR) and DNA blot analysis (DBA) for detecting clonal T-cell populations and investigated whether a nonradioactive PCR method could be used in routine clinical diagnosis. We analyzed DNA from 117 cases for T-cell clonality by PCR amplification. DBA was performed on 77 of these cases. Denaturing polyacrylamide gel electrophoresis (PCR-PAGE) of radiolabeled PCR products and capillary electrophoresis (PCR-CE) of fluorescently labeled PCR products were used for PCR product separation and quantitation. Complete agreement was obtained between PCR-PAGE and DBA in 67 of 77 cases. One case was positive by DBA and negative by PCR-PAGE, and 3 cases were positive by PAGE and negative by DBA. Five cases indeterminate by DBA were positive by PCR-PAGE, and 1 indeterminate case was negative by PCR-PAGE. In the comparison of PCR-PAGE and PCR-CE, of 63 cases with height ratios less than 2.0, all were negative by PCR-PAGE. Of 52 cases with height ratios of 2.0 or more, 50 were positive by PCR-PAGE. We conclude that PCR-CE is analytically equivalent to DBA and PCR-PAGE for detecting clonal T-cell populations. The PCR-CE method is semiquantitative and, therefore, may be more objective than gel-based methods. PMID- 10874887 TI - Mucocele of the appendix secondary to endometriosis. Report of two cases, one with localized pseudomyxoma peritonei. AB - This report documents 2 cases of obstructive mucocele of the appendix secondary to endometriosis of the appendix. In 1 case, the tip of the mucocele was ruptured and associated with localized pseudomyxoma peritonei. Mucoceles of the appendix usually are associated with hyperplastic or neoplastic mucosal proliferation; obstruction, particularly that due to endometriosis, is an infrequent cause. Occurrence of localized pseudomyxoma peritonei associated with appendiceal endometriosis and mucocele has not been reported previously. PMID- 10874886 TI - HER-2/neu gene amplification compared with HER-2/neu protein overexpression and interobserver reproducibility in invasive breast carcinoma. AB - We compared the detection of HER-2/neu gene amplification by fluorescence in situ hybridization (FISH) with detection of HER-2/neu protein overexpression by immunohistochemistry using 2 antibodies on 100 archival invasive breast carcinomas. Protein overexpression for each marker was scored independently by 4 pathologists using standardized criteria, and consensus was compared with results obtained from gene amplification. The concordance rate between FISH and immunohistochemistry was 76% for e2-4001 and 91% for the HercepTest. Of the 37 cases positive by e2-4001, 21 demonstrated no gene amplification; 7 of 24 cases positive by the HercepTest demonstrated no gene amplification. However, 1 of 61 cases negative by e2-4001 showed gene amplification; none of the cases negative by the HercepTest showed amplification. The predictive values of gene amplification based on 0-1+, 2+, and 3+ immunohistochemical staining were best for cases scored as 3+ (75% for e2-4001 and 89% for the HercepTest). Complete agreement among observers for immunohistochemical scoring of e2-4001 and the HercepTest was achieved in 75 and 85 cases, respectively. The pairwise kappa agreement values were substantial for e2-4001 and substantial to almost perfect for the HercepTest. Immunohistochemical staining may be considered a useful screening test. While negative staining almost always correlated with a lack of gene amplification, positive membranous staining, especially 2+, did not predict gene amplification. The low interobserver reproducibility in separating 2+ from 3+ cases necessitates further confirmation by FISH before treatment decisions are made. PMID- 10874888 TI - Fine-needle aspiration biopsy in the evaluation of lymphadenopathy associated with cutaneous T-cell lymphoma (mycosis fungoides/Sezary syndrome). AB - We studied the role of fine-needle aspiration (FNA) in the evaluation of lymphadenopathy associated with cutaneous T-cell lymphoma (CTCL) in 11 patients with lymphadenopathy and compared findings with corresponding histologic material. Molecular genetic analysis for T-cell clonality by polymerase chain reaction (PCR) was performed on all aspirates. Immunophenotyping was successful in 4 of 7 cases in which flow cytometry was attempted from the aspirated material. Cytologic evaluation of FNA samples correlated strongly with histologic rating of involvement based on numbers of atypical cerebriform lymphocytes in the nodal specimen. Of 7 nodal specimens with scattered or small groups of atypical cells in the background of dermatopathic lymphadenopathy (LN1-2), the cytologic diagnosis was interpreted as reactive in all instances. Of 4 specimens with highly suspect (LN3) or definite histologic involvement (LN4), the cytologic diagnosis was likewise suspect or malignant. The correlation between molecular genetic studies on FNA samples and studies on tissue was not significant; in 2 cases, a T-cell clone was detected in the nodal tissue sample but not in the FNA sample, suggesting undersampling. A T-cell clone was detected by PCR in 5 of 7 nodal specimens judged reactive by FNA biopsy or histologic assessment. FNA for cytologic and molecular genetic analysis is a useful method to evaluate lymphadenopathy associated with CTCL and may obviate the need for surgical biopsy. PMID- 10874889 TI - Large-needle aspiration biopsy for the preoperative selection of palpable thyroid nodules diagnosed by fine-needle aspiration as a microfollicular nodule or suspected cancer. AB - The palpable thyroid nodules with a fine-needle aspiration (FNA) diagnosis of microfollicular nodule or suspected cancer usually are excised; however, most of them are proved benign by postoperative histologic examination. We reviewed the clinical and pathologic data for patients with thyroid nodules with an FNA diagnosis of microfollicular nodule or suspected cancer; nodules also were examined by large-needle aspiration biopsy (LNAB) to assess whether the distinction achieved by LNAB into pure microfollicular or mixed microfollicular macrofollicular nodules could be used preoperatively to better predict malignancy. One hundred fourteen nodules of this type were excised. The prevalence of cancer was 22% (14/63) among the microfollicular and 4% (2/51) among the microfollicular-macrofollicular nodules at LNAB. These data indicate that histologic examination of the LNAB specimen can be used for preoperative selection of thyroid nodules diagnosed by FNA as a microfollicular nodule or suspected cancer. PMID- 10874890 TI - False-negative frozen section results. PMID- 10874891 TI - Mechanical forces on cancer cells. PMID- 10874892 TI - Cost minization analysis of telepathology: a critical review. PMID- 10874893 TI - Role of HER-2/neu oncogene amplification in determination of outcome of patients with cancer. PMID- 10874894 TI - [Risk factors for the development of vision disorders in polymyalgia rheumatica with giant cell arteritis]. AB - BACKGROUND AND OBJECTIVE: Visual disorders, even blindness, are serious complications of polymyalgia rheumatica (PMR) associated with temporal arteritis. Their early recognition in patients at high risk is essential to avoid the development of such visual disorders. It was the aim of this study to identify these risk factors. PATIENTS AND METHODS: Clinical and laboratory data and biopsy findings in 131 patients (94 women, 37 men; mean age 74 years) with PMR and concomitant temporal arteritis were analysed retrospectively. RESULTS: Visual disorders occurred in 61 of the 131 patients. Temporal artery biopsy was not sufficient to detect those at high risk. But this was possible by identifying a typical clinical pattern in that most patients with visual disorders had severe cerebral symptoms, while they had only minor forms of PMR and few generalized symptoms. There was no correlation between any of the laboratory tests and high risk. CONCLUSIONS: A typical pattern of clinical manifestations can provide early identification of those patients who have PMR associated with temporal arteritis and are at high risk of developing visual disorders. PMID- 10874895 TI - [Prevention of thrombosis with subcutaneous recombinant hirudin in heparin induced thrombocytopenia type II. A pilot study]. AB - BACKGROUND: Heparin-induced thrombocytopenia (HIT) type II is a severe complication of heparin therapy with a high incidence of thromboembolic events. AIM: The aim of this prospective study was to evaluate efficacy and safety of prophylaxis of thromboembolism with subcutaneous r-hirudin (25 mg twice daily) in patients with HIT type II. PATIENTS AND METHODS: From 01/06/1997 until 01/08/1999, 19 patients were prospectively included into the study. During subcutaneous r-hirudin application (25 mg twice daily) the activated partial thromboplastin time (aPTT) and ecarin clotting time (ECT) were measured twice daily prior to and 2 hours after the morning injection. RESULTS: Ten patients (mean age: 68 years; two men, eight women) with thromboembolic events were intravenously treated with r-hirudin (mean 19.3 days) with a target aPTT of 1.5 to 2.5 times normal values followed by subcutaneous r-hirudin (mean 22.5 days). Five Patients without thromboembolism immediately received subcutaneous r-hirudin (mean 25.9 days; mean age: 61 jahre; two men, three women) after cessation of heparin. Four patients requiring prophylaxis of thromboembolism received subcutaneous r-hirudin (mean 32 days; mean age: 68 years; four women) because of HIT type II in the past. Mean aPTT-values prior to and 1.5-2.5 hours after the morning injection were 1.2 to 1.7 and 2.0 to 2.3 times normal values, respectively. The ECT was prolonged by 1.2 to 1.7 and 2.3 to 2.5 times the upper normal value, respectively. Thromboembolic or bleeding events were not observed during the study. CONCLUSION: The subcutaneous application of r-hirudin provides an alternative for primary and secondary prophylaxis of thromboembolism in HIT type II patients. PMID- 10874896 TI - [Idiopathic right ventricular tachycardia or arrhythmogenic right ventricular tachycardia?]. AB - HISTORY: While cycling a 38-year-old man suddenly experienced palpitations associated with marked weakness. 90 min later his general practitioner, having diagnosed a ventricular tachycardia (VT) with a rate of 218/min, terminated it by a drug injection. INVESTIGATIONS: Electrocardiography (ECG), echocardiography and biventricular cardiac catheterization with right ventricular contrast injection failed to provide any evidence of structural abnormality. However, ergometry and EPS with programmed ventricular stimulation induced VT of identical morphology (left bundle branch bloc [LBBB] with right axis deviation [RAD]). TREATMENT AND COURSE: Idiopathic right-ventricular outflow tract tachycardia (IRVT) having been diagnosed, the patient was put on a maintenance dose of 50 mg/d atenolol. After 6 months without symptoms he again experienced several attacks of tachycardia. Resting ECG merely revealed an epsilon potential and negative T waves in V1-V3. Right ventricular contrast injection revealed inferolateral dyskinesia. EPS demonstrated both the known VT and a second, morphologically different one (LBBB with LAD). These findings indicated arrhythmogenic right-ventricular cardiomyopathy (ARCV). A cardioverter/defibrillator was implanted (ICD) and over the subsequent 8 months he had six episodes of VT which were quickly terminated by the ICD. CONCLUSION: At first presentation of right-ventricular outflow tract tachycardia it is often not possible to differentiate between IRVT and arrhythmogenic RV cardiomyopathy. The two being significantly different in prognosis and treatment, follow-up monitoring is essential to establish the definitive diagnosis. PMID- 10874897 TI - [Hyper- and hypocalcemia. 2: Hypocalcemia]. PMID- 10874898 TI - [Drug-induced long QT syndrome and torsade de pointes arrhythmias]. PMID- 10874900 TI - [Treatment of hepatorenal syndrome in liver cirrhosis]. PMID- 10874899 TI - [Ambulatory treatment of pulmonary artery embolism with low molecular weight heparin]. PMID- 10874901 TI - [Diabetology and endocrinology: agreement in proceeding]. PMID- 10874902 TI - [Subclinical arteriosclerosis in patients with newly diagnosed type 2 diabetes mellitus. Demonstration by high-resolution ultrasound measurements of intima media thickness of the common carotid and femoral arteries]. AB - BACKGROUND AND OBJECTIVE: Type 2 diabetes mellitus is an important risk factor for the development of atherosclerosis. Early subclinical manifestation of atherosclerosis can be reliably recognized by measuring the thickness of the intima and media (IMT). The aim of this study was to examine vessel changes and the extent of possible risk factors in patients with newly diagnosed type 2 diabetes (interval since diagnosis < or = 1 year) and control persons without DM. PATIENTS AND METHODS: Maximal IMT was measured by high resolution ultrasound of the common carotid (CCA) and femoral arteries (FA) in 51 type 2 diabetics and 18 controls. Various clinical and laboratory data (urinary excretion of albumin and protein, blood lipids) as well as amount of smoking, frequency of eating fruit and amount of sport activity were obtained in a standardized manner. RESULTS: The patients' body-weight was markedly greater and blood pressure significantly higher than that of the controls. Hypertension was present in 43% of patients (control group 11%) microalbuminuria in 26%. Mean IMT of the CCA was 0.76 +/- 0.20 mm in the patients and 0.64 +/- 0.16 mm in the controls (p < 0.01). The IMT of the FA, however, was not significantly different in the two groups (patients: 0.80 + 0.30 mm, controls: 0.75 +/- 0.31 mm). The IMT of the CCA was correlated with the patients' age (r = 0.55; p < 0.001), with the level of total cholesterol (r = 0.39; p < 0.01), and with the presence of hypertension (r = 0.38; p < 0.01). Patients who daily eat fruit had a significantly lower IMT of the FA than those who did not eat fruit regularly (no such difference was found regarding the CCA). Linear multiple regression analysis indicated that these variables were factors that independently affected the IMD of the CAA and the FA. CONCLUSIONS: An increase in subclinical atherosclerosis was demonstrated in type 2 diabetics already during the first year after diagnosis. The risk factors for the development of atherosclerosis in newly diagnosed diabetics exert a greater effect on the CCA than the FA. Regular eating of fruit seems to have a favourable effect on the progression of atherosclerosis of the FA. PMID- 10874903 TI - [Care of elderly diabetics by ambulatory nursing in the Heinsberg region]. AB - OBJECTIVE: The number of elderly people with chronic diseases receiving ambulant nursing care in the German population is continually increasing. However, little information is available on the quality of care of elderly diabetic patients. The aim of this study was to investigate the pattern of care provided for this group by such ambulatory nursing services. PATIENTS AND METHODS: All ambulant nursing services in the county of Heinsberg, North-Rhine-Westphalia, were contacted and asked to answer a standardized questionnaire. 23 of the 24 services participated in the study and provided suitable answers. The study was conducted between October 1998 and March 1999. RESULTS: All services together took care of 337 persons with known diabetes mellitus, of whom 79.6% were female and 20.4% were male. The total diabetes prevalence among all patients was 27.2%. 231 diabetic persons (68.5%) were treated with insulin. Nurses administered insulin to 84.8% of the patients. As expected many diabetics had severe late complications (7.4% amputation, 20.8% blindness, 4.2% renal failure). In the majority of patients, blood glucose was monitored, but only occasionally, whereas urinary glucose was checked in only a few cases. Foot inspection and care were provided regularly. 115 diabetics (34.1%) had at least one hospital stay during the previous 12 months period. CONCLUSION: The results of this study indicate that ambulatory nursing services care mainly for insulin-treated diabetic patients in late stages of the disease. The current organisational structure not meet the requirements of modern diabetes management. PMID- 10874904 TI - [Female pseudohermaphroditism in congenital adrenogenital syndrome as an incidental intraoperative finding in a 68 year old patient]. AB - HISTORY AND CLINICAL FINDINGS: A 68-year-old man of small stature, previously always healthy and with a grown-up daughter, was suspected of having carcinoma of the colon with metastasis to the right kidney. At laparotomy internal female genitalia with cancerous changes were unexpectedly discovered in the left adnexae. The colon carcinoma, the right adrenocortical tumour and left adnexae were resected. INVESTIGATIONS: Histological examination revealed adenocarcinoma of the colon, right adrenocortical adenoma and a Brenner tumour of the left female adnexae. Postoperative tests showed increased levels of 17-OH-progesterone (3192 ng/dl), 21-desoxycortisol (1856 ng/dl) and of adrostenedione (745 ng/dl), while the concentrations of 17-OH pregnenolone, testosterone and mineralocorticoids were within normal limits. Chromosome analysis demonstrated karyotype 46 XX. DIAGNOSIS, TREATMENT AND COURSE: As far as could be ascertained, this is the first documented case in the German-speaking region of female pseudohermaphroditism diagnosed in an elderly person with uncomplicated virilizing congenital adrenogrenital syndrome due to 21-hydroxylase deficiency (deletion of CYP21 gene). To avoid a cortisone deficiency crisis the patient was regularly given hydrocortisone and he quickly recovered. But he died 6 months later of sequelae of the carcinoma of the colon. CONCLUSION: An adrenogenital syndrome should be excluded in a case of bilateral adrenocortical tumour. As this is usually benign, conservative treatment should be attempted. This case demonstrates the necessity of thorough examination which could have given an early indication of the underlying condition. PMID- 10874905 TI - [Hyper- and hypocalcemia. 1. Hypercalcemia]. PMID- 10874906 TI - [Risks of iodine prophylaxis]. PMID- 10874907 TI - [Risk factor hyperinsulinemia? Risk predictor insulin resistance!]. PMID- 10874908 TI - [Risk factor hyperinsulinemia? Contra!]. PMID- 10874909 TI - Rehabilitation of persons with traumatic brain injury. AB - OBJECTIVE: The objective of this NIH Consensus Statement is to inform the biomedical research and clinical practice communities of the results of the NIH Consensus Development Conference on Rehabilitation of Persons with Traumatic Brain Injury. The statement provides state-of-the-art information regarding effective rehabilitation measures for persons who have suffered a traumatic brain injury (TBI) and presents the conclusions and recommendations of the consensus panel regarding these issues. In addition, the statement identifies those areas that deserve further investigation. Upon completion of this educational activity, the reader should possess a clear working clinical knowledge of the state of the art regarding this topic. The target audience for this statement includes, but is not limited to, pediatricians, family practitioners, internists, neurologists, physiatrists, psychologists, and behavioral medicine specialists. PARTICIPANTS: Participants were a non-Federal, nonadvocate, 16-member panel representing the fields of neuropsychology, neurology, psychiatry, behavioral medicine, family medicine, pediatrics, physical medicine and rehabilitation, speech and hearing, occupational therapy, nursing, epidemiology, biostatistics and the public. In addition, 23 experts from these same fields presented data to the panel and a conference audience of 883. EVIDENCE: The literature was searched through Medline and an extensive bibliography of references was provided to the panel and the conference audience. Experts prepared abstracts with relevant citations from the literature. A compendium of evidence was prepared by the panel which included a contribution from a patient with TBI, a report from an Evidence Based Practice Center of the Agency for Health Care Policy and Research, and a report from the National Center for Injury Prevention and Control at the Centers for Disease Control and Prevention. Scientific evidence was given precedence over clinical anecdotal experience. CONSENSUS PROCESS: The panel, answering predefined questions, developed their conclusions based on the scientific evidence presented in open forum and the scientific literature. The panel composed a draft statement that was read in its entirety and circulated to the experts and the audience for comment. Thereafter, the panel resolved conflicting recommendations and released a revised statement at the end of the conference. The panel finalized the revisions within a few weeks after the conference. The draft statement was made available on the World Wide Web immediately following its release at the conference and was updated with the panel's final revisions. CONCLUSIONS: Traumatic Brain Injury (TBI) results principally from vehicular incidents, falls, acts of violence, and sports injuries, and is more than twice as likely in males as in females. The estimated incidence rate is 100 per 100,000 persons with 52,000 annual deaths. The highest incidence is among persons 15 to 24 years of age and 75 years and older, with an additional less striking peak in incidence in children ages 5 and younger. Since TBI may result in lifelong impairment of an individual's physical, cognitive, and psychosocial functioning and prevalence is estimated to be 2.5 million to 6.5 million individuals, TBI is a disorder of major public health significance. Furthermore, mild TBI is significantly under diagnosed and the likely societal burden therefore even greater. Given the large toll of TBI and absence of a cure, prevention is of paramount importance. However, the focus of this conference was the evaluation of rehabilitative measures available for the cognitive and behavioral consequences of TBI. Although studies are relatively limited, available evidence supports the use of certain cognitive and behavioral rehabilitation strategies for individuals with TBI. This research needs to be replicated in larger, more definitive clinical trials. (ABSTRACT TRUNCATED) PMID- 10874911 TI - [Myasthenic syndrome induced by lithium]. AB - BACKGROUND: Lithium salts can induce a dysfunction of the neuromuscular junction. CASE REPORT: A patient given lithium for bipolar psychosis developed a state of major fatigue revealing a myasthenia syndrome which resolved progressively after lithium withdrawal. DISCUSSION: In the literature, we found 4 cases of neuromuscular junction disorders in patients treated with lithium. A myasthenia syndrome was induced by lithium in 3 cases and underlying myasthenia was disclosed by lithium in the last one. The long-term course in our patient was in favor of an induced syndrome rather than underlying myasthenia. We discuss the pathophysiological mechanisms. Lithium is a commonly prescribed drug. Clinicians should be aware of the possibility of neuromuscular junction disorder in patients taking lithium who develop muscle fatigue. PMID- 10874910 TI - [Cutaneous, hepatic and hematologic manifestations due to nevirapine: DRESS syndrome?]. AB - BACKGROUND: Manifestations similar to DRESS syndrome (drug rash with eosinophilia and systemic symptoms) may be induced by nevirapine. CASE REPORTS: Three patients developed skin rash and general signs of liver dysfunction during the first 5 weeks after starting nevirapine. Laboratory tests showed elevated eosinophil counts and signs of inflammation simulating severe infection. DISCUSSION: The incidence of DRESS syndrome is probably underestimated. No standard treatment has been proposed. In our 3 patients, parenteral corticosteroid therapy was successful, leading to a rapidly favorable clinical course although liver tests took longer to return to normal. PMID- 10874912 TI - [Chronic cough under treatment with paroxetine]. PMID- 10874913 TI - [Could vitamin C be useful in smoking cessation?]. PMID- 10874914 TI - [Localized scleroderma after hepatitis B vaccination]. PMID- 10874915 TI - [Current therapeutics in infectious dermatology]. AB - NEW AGENTS: Among new treatments used for infectious dermatology diseases, new agents for genital herpes, valaciclovir and famciclovir, have greatly simplified therapeutic schemes. Cidofovir has also been shown to be effective against aciclovir-resistant cutaneous and mucosal herpetic lesions and for the treatment of molluscum contagiosum. NEW ADMINISTRATION ROUTES: For genital papillomavirus infections, trials using systemic or intralesional administered interferon have not provided conclusive evidence but imiquimode appears to be quite promising. Itaconazole and fluconazole are effective for onchomycoses. NEW POSSIBILITIES: Ivermectine is effective against scabies, but must be reserved for particularly severe forms. Finally, the emergence of Neisseria gonorrhoeae strains resistant to fluoroquinolones is disquieting. PMID- 10874916 TI - [Drug prescriptions outside the marketing product license and its responsibilities]. AB - In France, drugs are commercialized after obtaining marketing approval. There are some situations however in which clinicians are authorized to prescribe drugs outside the limits set by the approval document. Clinicians must comply with certain number of regulations. Compliance with prescription modalities, particularly precautions concerning prescription writing or the patient's informed consent, is mandatory. Although patients cannot be reimbursed for drugs prescribed outside the approval limitations, they may nevertheless benefit from special dispositions: temporary approval for use and experimentation within the framework of the Huriet law. We analyze here the question of liability in case of drug prescriptions outside approval limitations, discussing the issues of civil and penal law and patient reimbursement. PMID- 10874917 TI - [Rapaglinide: Novonorm, an alternative in type 2 diabetes]. PMID- 10874918 TI - [Treatment of community-acquired pneumonia with levofloxacin: 500 mg once a day or 500 mg twice a day?]. AB - LEVOFLOXACIN: A new anti-pneumococcal fluoroquinolone, levofloxacin, has received approval in France for the treatment of community-acquired pneumonia at the dose of 500 mg once or twice a day, depending on the severity of the disease, the germ susceptibility and the patient's weight. Levofloxacin has a powerful and rapid bactericidal activity, particularly against pneumococci, whatever the level of penicillin resistance. The pharmacokinetic properties of the compound allow once daily dosage. Pharmacodynamically, it has been clinically demonstrated that the most predictive parameter of efficacy is the Cmax/MIC ratio. PNEUMOCOCCAL PNEUMONIA: Because of the potential gravity of pneumococcal pneumonia, it might be preferrable to use levofloxacin at the dose of 500 mg twice daily. The efficacy of the two levofloxacin doses for the treatment of pneumococcal pneumonia was thus analyzed. Five clinical studies including 4 comparative trials, enrolling nearly 2,000 patients with community-acquired pneumonia were reported in the international approval document. Among these patients, 310 had documented pneumococcal pneumonia including 31% with bacteriemia. TASK FORCE REPORT: On the basis of available data, the level of proof is sufficient to prescribe levofloxacin at the dose of 500 mg once daily for the treatment of mild to moderately severe community-acquired pneumonia in ambulatory patients, including those with suspected pneumococcal pneumonia, with or without bacteriemia. It would be reasonable to propose the 500 mg twice daily dosage for severe community-acquired pneumonia warranting intensive care hospitalization in accordance with the criteria of the ERS Task Force Report. The well-founded rationale of this therapeutic strategy should be validated by the results of ongoing studies and by following the evolution of germ susceptibility to these new compounds. PMID- 10874919 TI - [Blood monitoring at the dawn of the 21st century]. AB - A NEW DISCIPLINE: On January 4, 1993, the French parliament voted a law on Blood Transfusion Safety, creating a monitoring and warning system operating through a complex network and aimed at guaranteeing permanent efficacy, safety and efficiency of blood transfusion in France. A new discipline, blood monitoring was born. FUNDAMENTAL PRINCIPLES: Basically, blood monitoring is organized around two fundamental principles: blood products trackability from the donor to the recipient and mandatory reporting of all transfusion incidents. After a laborious development phase, the early results point out the immunological risk, particularly by ABO error, and the risk of bacterial contamination. AN EARLY WARNING SYSTEM: This blood monitoring system, whose primary purpose is to serve as an early warning device, must have the necessary tools to monitor, verify and analyze all available epidemiological data. Coordination of the blood monitoring system within the framework of the AFSSaPS (the French Agency for Sanitary Safety of Health Products) should enable it to fully reach this goal. PMID- 10874920 TI - [Metastatic colorectal cancer: new therapeutics]. AB - NEED FOR NEW CHEMOTHERAPEUTIC AGENTS: The use of 5-FU in combination with leucoverin (LV) in the treatment of advanced colorectal cancer has consistently provided antitumoral response. The antitumoral activity of the 5-FU/LV combination is associated with inhibition of thymidylate synthetase, an essential enzyme in pyrimidine de novo biosynthesis of thymidilates as precursors of DNA synthesis. However, dose-limiting toxicities and the limited impact on survival point to the need to develop new drugs and approaches to improve therapeutic efficacy and survival. CLINICAL RESULTS: In advanced colorecal cancer, clinical results have confirmed the therapeutic efficacy of direct and indirect thymidylate synthetase inhibitors. Promising new agents: 5-FU prodrugs and inhibitors of dihydropyrimidine dehydrogenase (DPD) are promising chemotherapeutic agents with good oral bioavailability which can be combined with other drugs (leucoverin) with acceptable toxicity. Oxaliplatin and topoisomerase inhibitors such as irinotecan (CPT-11) have demonstrated activity in previously treated and newly diagnosed patients. COMBINATIONS: The best combination between DNA interaction drugs based on potentiation of DNA damage induced by thymidylate synthetase inhibitors and inhibition of DNA repair by DNA interactions remains to be evaluated. PMID- 10874921 TI - [Current status of anticoagulants]. AB - INDICATIONS: Direct inhibitors of thrombin, such as hirudin, are directed against the active site and the recognition site of thrombin. Because of their low molecular-weight, they can inactivate thrombin bound to fibrin. Prevention of thromboembolic complications in patients undergoing primary total hip or knee replacement is now an authorized indication of desirudin in France. The recommended treatment for heparin-induced thrombocytopenia is lepirudin when there is a clinically evident thrombosis and danaparoid sodium, a mixture of anticoagulant glycosaminoglycans in an antithrombotic prophylaxis setting. LMWH: Low-molecular weight heparins are not yet authorized in France for the treatment of pulmonary embolism. However, deep venous thrombosis can be securely treated with one daily fixed dose of nadroparin or tinzaparin. ORAL ANTICOAGULATION: The duration of anticoagulation therapy in patients with venous thromboembolism remains controversial. Three to six months of therapy is recommended after a first episode of venous thromboembolism; the shorter regimen may be chosen when there is an identifiable and transient risk factor, and the longer when the thrombosis is idiopathic. In the context of primary prevention of ischaemic heart disease low intensity oral anticoagulation could be recommended in men at high risk. PMID- 10874922 TI - [Treatment of sleep apnea syndromes]. AB - NECESSARY TREATMENT: Sleep apnea syndrome requires treatment because it affects cardiovascular and cerebrovascular morbidity and mortality and has important neuropsychological consequences with the risk of accidents due to impaired wakefulness. The patientis quality of life is greatly altered. GENERAL MEASURES: Patients should be informed of the risk due to the lack of sleep, advised that alcohol tranquilizers and hypnotic drugs are contraindicated, and counseled about loosing weight, the most difficult problem for obese patients. POSITIVE PRESSURE VENTILATION: Continuous positive pressure ventilation with a facial mask acts like a pneumatic prosthesis holding the airways open during sleep. Sleep can be reconstructed by eliminating the recorded pathological nocturnal events and thus reducing diurnal hypersomnia. Quality of life is improved and accidents related to diminished wakefulness are avoided. Death rate in treated patients is significantly lower than in non-treated patients. In France, the national health care system will reimburse positive pressure ventilation for sleep apnea syndromes recognized to cause more than 30 events per hour of recording or fragmented sleep due to respiratory impairment. OTHER TREATMENTS: Indications for other treatments in case of moderately severe sleep apnea syndrome (or if health care benefits are not recognized for positive pressure ventilation) are currently debated. No medication has been proven to be effective. Mandibular advancement ortheses are in the development stage and require multidisiplinary cooperation to verify their efficacy. Velar surgery has been proposed but is usually disappointing except for young patients actively participating in an integrated surgical treatment strategy. PMID- 10874923 TI - [Acne today. What's new?]. AB - WELL-CONDUCTED TREATMENT: Acne is a common condition in adolescents and requires careful management both in terms of therapeutic care and psychological support. Treatment is long and requires strict compliance. A well-conducted treatment can be expected to provide major improvement in most patients. LOCAL CARE: Local care is often sufficient for retentional or discretely inflammatory acne. A retinoid and/or a benzoyl peroxide can be associated with a local antibiotic. SEVERE FORMS: An oral antibiotic regimen for at least 3 months is proposed in association with the local treatment (retinoid, benzoyl peroxide, local antibiotic) in case of severe acne. ISOTRETINOIN: In case of unsuccessful treatment for nodular or conglobata acne, isotretinoin can be proposed in an oral preparation. This highly teratogenic drug must not be prescribed for women of reproductive age unless a well proven contraception has been instituted for more than 1 month and maintained for the entire duration of the treatment and 1 month after discontinuation. The patient must be informed of the risk in case of pregnancy and consent to regular monitoring of beta hCG less than 3 days before treatment onset, every 2 months during treatment and 5 weeks after treatment withdrawal). PMID- 10874924 TI - [Hemorrhage after biopsy or colonic polypectomy: placing a clip would be an ideal treatment]. PMID- 10874925 TI - Infrared spectroscopy of the photo- and radiobiology of DNA bases and their derivatives. PMID- 10874926 TI - Human tissues: chemical composition and photon dosimetry data. PMID- 10874927 TI - Identification of radioproducts resulting from the breakage of thymine moiety by gamma irradiation of E. coli DNA in an aerated aqueous solution. PMID- 10874928 TI - Leukemia in irradiated parabiotic rats. PMID- 10874929 TI - Radiation survival and regeneration characteristics of spermatogenic stem cells of mouse testis. PMID- 10874930 TI - Delayed brain swelling and functional derangement after X-irradiation of the right visual cortex in the Macaca mulatta. PMID- 10874931 TI - Dose protraction and acute radiation mortality in the chicken: a reappraisal of injury accumulation kinetics. PMID- 10874933 TI - Effects of single-dose partial-body x-irradiation on cell proliferation in the mouse small intestinal epithelium. PMID- 10874932 TI - Cyclic radiation-induced variations in cellular radiosensitivity in a mouse mammary tumor. PMID- 10874934 TI - Relative effects of whole-body sublethal doses of 60-MeV protons and 300-kVp X rays on disease incidences in RF mice. PMID- 10874935 TI - Metaphase chromosome aberrations in the Chinese hamster liver in vivo after either acute or fractionated 60Co irradiation. PMID- 10874936 TI - A study of the biological effectiveness of high-energy electrons at ultra-high dose rates using dry eggs of Artemia. PMID- 10874937 TI - Sedimentation of DNA from human fibroblasts irradiated with ultraviolet light: possible detection of excision breaks in normal and repair-deficient xeroderma pigmentosum cells. PMID- 10874939 TI - Haptoglobin levels in plasma of irradiated mice. PMID- 10874940 TI - Chemical radiosensitization of anoxic mammalian cells: effect of cell concentration. PMID- 10874938 TI - Dose-response relation of chromosome aberrations in human lymphocytes after in vitro irradiation with 3-MeV electrons. PMID- 10874941 TI - The repair of single-strand breaks in a radiosensitive mutant of Micrococcus radiodurans. PMID- 10874942 TI - Effects of irradiation on experimental autoallergic sialadenitis. PMID- 10874943 TI - Damage of rat thyroid by 131I and evidence against immunologic transferability. PMID- 10874944 TI - H2O and D2O sorption studies on spores of Bacillus megaterium. PMID- 10874945 TI - The influence of added H2O and D2O on anoxic radiation sensitivity in bacterial spores. PMID- 10874946 TI - Detection of pulmonary irradiation injury by determining plasma lactic dehydrogenase (LDH) activity. PMID- 10874947 TI - Near-UV effects on the induction of prophage. PMID- 10874948 TI - Interstrain variation in gamma-ray induction of hindlimb deformities in chick embryos. PMID- 10874949 TI - The effect of time between fractions on the response of tumors to irradiation. PMID- 10874950 TI - Effects of gamma irradiation on the function and conformation of ribonuclease A in dilute solution. PMID- 10874951 TI - Radiolysis of an alkaline phosphatase. PMID- 10874952 TI - A marker for mammalian DNA sedimentation. PMID- 10874953 TI - Gamma-radiolysis of trypsin, chymotrypsin, and chymotrypsinogen when associated with DNA. PMID- 10874954 TI - Studies on lysosomes after irradiation. I. A quantitative histochemical method for the study of lysosomal membrane permeability and acid phosphatase activity. PMID- 10874955 TI - Studies on lysosomes after irradiation. II. Lysosomal membrane permeability and acid phosphatase activity of lymphoid and other tissues after whole-body irradiation. PMID- 10874956 TI - Effects of X-irradiation on cell proliferation and DNA synthesis induced by administration of isoproterenol in salivary glands of the mouse. PMID- 10874957 TI - Comparative long-term effects of liver damage in the rat after (a) localized X irradiation and (b) localized X-irradiation in the presence of a strong homogeneous magnetic field. PMID- 10874958 TI - Rapid radiation cell death and cell proliferation in intestinal epithelium after 1000-rad irradiation. PMID- 10874959 TI - Early mortality in the young chicken after exposure to fast neutrons or gamma rays. PMID- 10874960 TI - An approach to statistical factors in radiation damage through the analysis of the 3.5-day effect of whole-body X-rayed chick embryos. PMID- 10874961 TI - Toxicity of 90Sr-90Y in Chinese hamsters. PMID- 10874962 TI - Chromosome breakage in human peripheral lymphocytes after radioactive iodine (125I) treatment. PMID- 10874963 TI - Mammary tumorigenesis through irradiation of mice. PMID- 10874964 TI - Effects of low-dose prenatal irradiation in humans: analysis of Chicago lying-in data and comparison with other studies. PMID- 10874966 TI - [Radiology of pulmonary ventilation by means of magnetic resonance tomography]. PMID- 10874965 TI - Effect of gamma-irradiation on the protein metabolism in Drosophilidae. PMID- 10874967 TI - [Detection and quantification of coronary calcification: an update]. AB - The demonstration of calcification of the coronary arterial wall indicates the presence of coronary heart disease (CHD). The prevalence of coronary calcifications increases with age. The extent of the calcifications correlates with the total coronary plaque burden and with the probability of a future myocardial infarction in symptomatic patients. In asymptomatic subjects with risk factors for a myocardial event demonstration of coronary calcifications is diagnostic for coronary atherosclerotic disease before clinical manifestation of the disease. Exact quantification of coronary arterial calcifications (calcium scoring) has become possible with electron beam computed tomography (EBCT) or ECG triggered subsecond CT scanners. Further improvements in the detection of coronary calcifications can be expected with the introduction of multi-slice CT. The prognostic relevance of coronary calcium scoring in asymptomatic high-risk patients is not yet clearly defined. It remains to be clarified whether newer, volume based methods of calcium quantification will be superior to the classic calcium score (Agatston-Score) for risk assessment and follow-up in this patient group. PMID- 10874968 TI - [Typical and unusual findings in MR mammography]. AB - In recent years, MR mammography has gained increasing importance in breast diagnostics. The main advantage of this technique is its high sensitivity for invasive breast cancer. The two main indications for MR mammography are preoperative staging of breast cancer and the differentiation between postoperative changes and recurrent tumor. In addition, MR mammography is increasingly used for problem solving in cases of questionable clinical, mammographic or sonographic findings. At this it is important to know not only the different manifestations of breast cancer, but also important benign and malignant diagnostic alternatives. Furthermore, it is necessary to be familiar with therapy-related changes. The most important criterion to differentiate between benign and malignant lesions in MR mammography is the extent and temporal course of contrast enhancement. In addition, the lesion morphology and the signal intensity on T2-weighted images can be used to distinguish between different disease entities. The following review article will discuss typical and unusual findings in breast MRI, including rare entities as well as changes after breast conserving therapy and chemotherapy. PMID- 10874969 TI - [Cardiac multidetector-row CT: first clinical results of retrospectively ECG gated spiral with optimized temporal and spatial resolution]. AB - PURPOSE: The significantly improved temporal and spatial resolution of Multidetector-Row CT opens up new possibilities for cardiac imaging. A method with retrospectively ECG-gated spiral acquisition is presented. MATERIALS AND METHODS: A total of 10 patients underwent cardiac CT on a fast multi-slice CT system with 4 simultaneously acquired slices and 0.5 s rotation time (Siemens Somatom Volume Zoom). Continuous spiral data of the entire heart volume (5 studies precontrast for calcium scoring, 5 studies with contrast) were acquired together with the patient's ECG and reconstructed with dedicated spiral algorithms providing 250 ms temporal resolution. Three-dimensional image data sets were built up from overlapping slices that were reconstructed in an arbitrary, user-defined phase of the heart cycle (e.g., diastolic phase). To evaluate the capability of the method for functional imaging, complete image volumes were reconstructed from the same spiral data set in different phases of the heart cycle. RESULTS: Within a single breath-hold, a spiral data set of the entire heart volume could be acquired. Typical scan times for standard examinations with 3-mm slice width were 12-17 s, and for high-resolution CT angiographies of the coronary arteries with 1.25-mm slice width about 25-35 s. Motion-free reconstruction of the heart and coronary arteries with high spatial resolution were possible in the diastolic phase of the heart cycle. Multiphase reconstructions from the same spiral scan data set were possible. CONCLUSIONS: Fast multi-slice spiral CT with retrospectively ECG-gated spiral reconstruction is well suited for three-dimensional and functional imaging of the heart, especially for high-resolution imaging of calcified coronary plaques and CT angiography of the coronary arteries. PMID- 10874970 TI - [Pre- and postoperative evaluation of ventricular function, muscle mass and valve morphology by magnetic resonance tomography in Ebstein's anomaly]. AB - PURPOSE: To evaluate the value of MRT with spin echo (SE) and CINE gradient echo (GE) sequences for the pre- and postoperative assessment of patients with Ebstein's anomaly. METHODS: Twelve patients within the ages of four to 49 years (mean 22 +/- 12 years) were examined pre- (n = 5) or postoperatively (n = 7) after tricuspid valve reconstruction with a 1.5 T scanner. For the anatomical assessment, an ECG-gated transverse SE-sequence, for the assessment of valve morphology and function as well as for volumetry a CINE GE-sequence with retrospective gating was used. With the use of the multislice-multiphase technique, after summing up the manually outlined epi- and endocardial areas, endsystolic (ESV) and enddiastolic volumes (EDV), ejection fraction (EF), stroke volume (SV), and muscle mass (MM) were calculated for both ventricles. RESULTS: The differentiation of the displaced parts of the tricuspid valve (TV) was insufficient with static SE, but was possible in all patients with CINE-MRT. Like in Doppler echocardiography, a qualitative assessment of tricuspid insufficiency was possible in CINE-MRT, the mean incompetence grade preoperative was 1.8 (+/- 0.8), postoperative 0.7 (+/- 0.5). The mean RV-EF in the preoperative group was 41.8% (+/- 6.4), in the postoperative group 47.9% (+/- 10.6), the mean LV-EF preoperative 47.4% (+/- 8.5%), postoperative 63.0% (+/- 9.4). CONCLUSION: CINE MRT should rather be used than SE for the assessment of valve morphology. EF, muscle mass and tricuspid incompetence can also be calculated pre- and postoperative with CINE-MRT. PMID- 10874971 TI - [Contrast enhanced power Doppler sonography: comparison of various administration forms of the ultrasound contrast agent Levovist]. AB - PURPOSE: Objective of the present study was the comparison of various administration forms of the ultrasound contrast medium Levovist with regard to duration and intensity of contrast enhancement in patients with tumors of the liver or pancreas. PATIENTS AND METHODS: Seven patients with tumors of the liver or pancreas were examined prospectively using power Doppler sonography. Ultrasound contrast enhancement was achieved using Levovist (8 ml, 400 mg/ml) in three different administration forms: 1st as a bolus injection through the main channel, 2nd through the injection valve of an intravenous cannula, or 3rd as a continuous infusion. Semiquantitative evaluation of the degree of contrast enhancement over the course of the examination was conducted by three independent examiners. RESULTS: Levovist, administered by continuous infusion, resulted in a significantly longer average period of contrast enhancement (9:43 min (extratumoral), 7:34 min (intratumoral)) than did the same dosage administered as a bolus injection through the main channel (6:01 min (extratumoral), 4:54 min (intratumoral), p = 0.0156 (extratumoral); p = 0.0313 (intratumoral), but contrast intensity was decreased. Bolus injection through the injection valve of the i.v. cannula was associated with decreased duration and intensity of contrast enhancement compared with injection through the main channel. CONCLUSION: Compared with bolus injection, the continuous infusion of Levovist resulted in a significant prolongation of the duration but in a decreased intensity of contrast enhancement. Administration of Levovist through the injection valve does not result in optimal contrast enhancement and is therefore not recommended. PMID- 10874972 TI - [Preoperative examination of potential kidney transplantation donors: value of gadolinium-enhanced 3D MR angiography in comparison with DSA and urography]. AB - PURPOSE: To assess a contrast-enhanced standardized MRA protocol for the presurgical evaluation of potential renal transplant donors. METHODS: Twenty three potential donors for renal transplantations were examined with gadolinium enhanced, two-phase MR angiograms (1.5 T) and DSA/urography for the number of renal arteries, the presence of aberrant arterial and venous branches, renal artery stenoses and anatomy of the renal collecting system and ureters. The diagnostic value was assessed by evaluating different image processing modalities and interobserver variability. RESULTS: Using maximum intensity projections (MIP) together with multiplanar reformatting (MPR), accessory arteries were detected with a sensitivity/specificity of 100%/98%. Depending on diagnostic experience, exclusive evaluation of MIP yielded a sensitivity/specificity of 67-100%/95-100%. Using MIP/MPR, venous depiction was good in 80%, with MIP solely in 30-40%. At least the proximal third of the ureter was visible in 67%. CONCLUSION: MPR/MIP evaluation of two-phase, contrast-enhanced MRA provides an excellent depiction of renal vessel anatomy for presurgical evaluation of renal transplant donors. Exclusive MIP assessment is less reliable and depends strongly on the examiner's experience. For sufficient visualization of the ureters, either additional measurements or low-dose diuretic injection have to performed. PMID- 10874973 TI - [CT- and ultrasound-guided biopsies: prospective comparison of fine-needle aspiration with true-cut biopsy in 103 patients]. AB - PURPOSE: A prospective comparison of FNAB and TCB was performed in an identical set of patients and lesions. METHODS: In 103 patients focal lesions were biopsied by US- or CT-guidance with at least one FNAB and TCB each. Complications were registered and documented. Cytological and histological specimens were evaluated independently. A final diagnosis was then attempted by consensus. RESULTS: A total of 253 punctures was performed with sufficient tissue in 72.8% (FNAB) and 85.4% (TCB) of the cases. Diagnosis was possible in 68% (FNAB) and 80.6% (TCB). After combined evaluation of both specimens the diagnostic yield increased to 91.3%. Minor complications without clinical sequelae were observed in 21 patients. CONCLUSION: A diagnostic strategy with the combined use of FNAB and TCB increases the diagnostic yield in image-guided punctures. Even though this approach needs at least two separate punctures, the complication rate does not increase. PMID- 10874974 TI - [Locoregional chemoperfusion with mitoxantrone for palliative therapy in bleeding bladder cancer compared with embolization]. AB - PURPOSE: To assess the efficacy of intraarterial chemoperfusion (CP) with mitoxantrone in patients with bleeding bladder cancer; comparison with the results of intraarterial embolization therapy (ET). MATERIALS AND METHODS: Thirty patients with urinary bladder cancer and intractable bladder hemorrhage were treated with intraarterial (i.a.) CP (15 patients) using Mitoxantron 820 mg/m2/1 2 h) and i.a. ET (15 patients) using Histoacryl or Ethibloc. Bleeding control rate, recurrence of hemorrhage, survival rate and complications were evaluated. RESULTS: Complete control of the hemorrhage was achieved in 14/15 and 12/15 of the patients with CP and ET, respectively. Hemorrhage stopped in CP patients after an interval of (4 to 15) 10 days, and within 24 hours in ET patients. Recurrence of hemorrhage was observed in 3/14 of CP and 4/13 of ET patients. The survival rate was 4-5 months in both groups. Complications were observed in ET patients only (7/22). Posttherapeutic pain occurred significantly more often in ET patients (20/22) than in CP patients (6/31 versus 20/22). CONCLUSION: Intra arterial chemoperfusion using Mitoxantron is an effective therapy in patients with intractable urinary bladder hemorrhage. Due to the delayed effect in CP, ET should be used in patients with life-threatening bleeding. PMID- 10874975 TI - [Temporary implantation of a metal stent in Vater's ampulla. Evaluation of early changes of the bile ducts in an animal study]. AB - PURPOSE: Evaluation of temporary stent implantation in the papilla of vater and the subsequent alterations of the bile ducts in a controlled animal study. MATERIAL AND METHODS: In seven domestic pigs the papilla of vater was stented with a nitinolelastalloy-strecker stent over a period of three days. The microscopic and macroscopic changes of the biliary tract and the serologic parameters were judged. RESULTS: In three cases a slight widening of the peripheric bile ducts was found. With the exception of a mild elevation of lactate dehydrogenase the serological parameters remained normal. The explantation of the stent two possible in six cases. A thickening of the outer layers of the bile ducts and a low to middle grade injury of the mucosa were found histologically. CONCLUSION: A temporary stent implantation in the papilla of vater is feasible. The mucosal injuries caused by the temporary stent placement may be fully repairable. The method seems to be suitable for prolonged dilatation of benign stenoses. PMID- 10874976 TI - [Diagnostic methods for the quantification of radiation injuries of the lungs]. AB - PURPOSE: The aim of this paper is to find techniques for quantifying radiation lung injury after irradiation with lung involvement to improve an early diagnosis. METHODS: The case of a patient with NSCLC was used to demonstrate different methods in order to quantify a developing pneumopathy after radiation treatment. By means of HRCT studies in the follow-up, a procedure was developed by defining a test-ROI in high-dose areas of the lung and evaluating the corresponding HU-histogramm for the parameters of the lung peak. Changes during the follow-up can be derived from the differential HU-histogram by the determination of a parameter called delta HUrel, which quantifies the shift to higher HU values. Alternatively, a Fourier analysis of the lung pattern within the test-ROI results in a Fourier amplitude distribution, which reacts sensitively to changes during the follow-up. Furthermore, a Fourier-frequency histogram can be derived which is independent of the spatial orientation of the density pattern. RESULTS: From the HRCT follow-up study, values for delta HUrel can be derived to be 0.24, 0.44, and 0.50 (56, 100 and 422 days after beginning the treatment). The differential Fourier frequency-histogram presentations demonstrate pronounced pattern changes. CONCLUSION: The presented methods demonstrate possibilities to quantify radiation lung injury. The proven sensitivity can possibly be improved after the introduction of a breath triggered HRCT technique. PMID- 10874977 TI - [A hybrid technique for the automatic floating table MRA of peripheral arteries using a dedicated phased-array coil combination]. AB - PURPOSE: We introduce a hybrid technique which allows a high resolution MRA of the peripheral arteries with a dedicated phased-array coil using the floating table technique. MATERIALS AND METHODS: Five patients with peripheral arterial occlusive disease were examined within one week with i.a. DSA and MRA using the hybrid technique. MRA examinations were done on a 1.5 T system. At first, pelvic arteries were examined in a single step mode applying the CareBolus technique. Subsequently, thighs and lower legs were examined using the floating table mode. 125 vascular segments were evaluated. RESULTS: The hybrid technique proved to be robust and could be performed in each case. Mean examination time was about 30 min. For 117 vascular segments no difference was found between i.a. DSA and MRA. Three segments revealed a higher grade of stenosis in DSA than in MRA, five segments were graded higher in MRA than in DSA. Occlusions were visualized identically in both methods. Venous overlap had no relevant effects on image evaluation. CONCLUSIONS: This hybrid technique in combination with phased-array coils allows a high resolution MRA of the peripheral arteries with very good image quality. If future studies confirm reduced venous overlap, this method may be an alternative also for users of the floating table MRA with the body resonator. PMID- 10874978 TI - [Ganglioneuromas in childhood: CT and MRI characteristics]. AB - PURPOSE: The aim of this study was to demonstrate the typical appearance of ganglioneuromas in computer-assisted tomography (CT), and magnetic resonance imaging (MRI). MATERIAL AND METHODS: Retrospective analysis of diagnostic imaging (9 CT, 6 MRI) in 9 children aged 3 to 15 years with the histological diagnosis of ganglioneuroma. RESULTS: The tomographies showed large (max. 13.4 cm in diameter) round or oval tumors with sharp delineation. The sites of the tumors were the retroperitoneum (5), the mediastinum (3), and the adrenal gland (1). Intraspinal tumor involvement occurred in 4 cases. On comparing CT with MRI, MRI was more accurate in defining the intraspinal involvement. The ganglioneuromas were of hypodense appearance in the native CT scan and showed moderate enhancement upon administration of contrast media. In five patients tumor calcifications with a disseminated sprinkled pattern were seen in CT. In MRI T1-weighted scans the tumors were homogeneous and hypointense, after gadolinium application a marked enhancement was evident. In T2-weighted scans the tumors were hyperintense. CONCLUSION: At the time of diagnosis ganglioneuromas are generally large tumors which can be well detected by CT and MRI. Information towards the diagnosis is given by the appearance of the ganglioneuromas in CT and MRI. However, MRI is the modality of choice due to its superiority in documenting intraspinal tumor expansion. PMID- 10874979 TI - [Metastatic calcinosis of the lung]. PMID- 10874980 TI - [Myocardial heart metastasis in rapidly progressing renal cell carcinoma]. PMID- 10874981 TI - [Acute heart arrest in spiral CT]. PMID- 10874982 TI - Characterization of thin layers of glucose oxidase. AB - Glucose Oxidase (GOD) has been covalently bound to functionalized glass cover slips. The surface density of immobilized GOD molecules was measured by a method based on the amperometric determination of Flavin Adenine Dinucleotide (FAD). Atomic Force Microscopy (AFM) images, obtained in aqueous solution for the covalently bound enzyme, show a monomolecular layer of the enzyme on a functionalized glass surface. The catalytic constants were measured for the immobilized GOD and compared with those of the free enzyme. PMID- 10874983 TI - Modulation between aerobic and anaerobic metabolism in the mutant cell line CdtR Q. AB - It has recently been shown that the cell line Don Q obtained by mutagenesis of wild type Chinese hamster lung fibroblasts (Don wt), presents a point mutation in the gene coding for UDP-glucose pyrophosphorylase. The persistent low level of UDP-glucose makes Don Q clone resistant to Clostridium difficile toxin B. Starting from the observation that Don Q cells exhibit many large hydrophobic cytoplasmic inclusions, that we have found to be made of neutral lipids, the aerobic metabolism of the two cell lines has been examined. The specific activity of cytochrome oxidase in Don Q cells is more than 5 times lower than that found in Don wt. Also, the activity of Complexes II + III, expressed by the activity of succinate-cytochrome c oxido-reductase, has been found to be lower in Don Q compared to wt cells. On the other hand, NADH-cytochrome c oxido-reductase activity, insensitive to rotenone, is more than doubled in Don Q. In these cells the activity of lactate dehydrogenase is very high, being able to oxidise more than 3,000 nmoles of NADH/min/mg of protein. The results obtained indicate that Don Q cells, in addition to a decreased ability to synthesise UDP-glucose, have an impairment in the respiratory chain. Such an impairment could be correlated to the increased capacity to generate a higher amount of reducing equivalents through the glycolytic activity, which can then be utilised for the synthesis of fatty acids stored in lipid droplets. PMID- 10874984 TI - Using scapular measurements in regression formulae for the estimation of stature. AB - There are very few papers in forensic literature in which scapular dimensions have been used for estimation of living stature. Allowing the forensic duty to estimate the living stature of skeletal remains, using intact or fragmented scapulae, the Authors have performed multiple regression analysis between the measurements taken from 80 scapula (40 male and 40 female) belonging to a skeletal collection with anthropometric known data. Seven parameters (max length, max breadth, max acrocoracoid distance, length of acromion, max length of coracoid, length of glenoid cavity, width of glenoid cavity) have been recorded. By statistical analysis multiple and linear regressions have been obtained. The results show that living stature may be determined by using regression formulae of single or associated parameters taken from whole or fragmented scapulae. In absence of intact or fragmented long limb bones, scapula sample can be reliably employed for the estimation of stature in forensic practice. PMID- 10874985 TI - Newborn screening for human immunodeficiency virus infection in the Bronx, NY, and evolving public health policy. AB - New advances in the diagnosis and treatment of HIV infection continue to propel changes in public understanding of HIV infection and the administration of public health law. Over the past decade, New York State has moved from a policy of blind newborn screening for seroprevalence data to mandatory HIV testing as part of the statewide Newborn Screening (NBS) Program. A new statewide program of expedited HIV testing (48-hr turnaround results) of pregnant women and newborns (whose HIV status is unknown at the time of delivery) began in the summer of 1999. To better understand the impact this program might have on the patients who receive health care at Lincoln Medical and Mental Health Center (Lincoln Hospital), we evaluated our experience with the current NBS program prior to inauguration of the new expedited testing program. We evaluated the NBS program from February 1, 1997, to January 31, 1999, including total number of HIV-exposed/infected infants born, mother's HIV status (if known) at the time of delivery, amount of time between blood sampling and return of the test results to the Hospital, and medical follow up of infants with positive newborn screening test results. This was a retrospective study of the NBS registry and the medical records of patients who receive primary health care from the Pediatric Immunology Service of the Department of Pediatrics at Lincoln Hospital. One hundred and four newborns were identified with positive-HIV antibody (HIV-Ab-positive), and 13 (12.5%) were confirmed to be HIV-infected by positive polymerase chain reaction (PCR) test of viral DNA. Sixty-five (62.5%) of the newborns with positive NBS screening test results were born to mothers who were known to be HIV-infected prior to delivery; 39 (37.5%) were unanticipated. Four (30%) of the 13 HIV-infected babies were born to mothers who were known to be HIV-infected prior to delivery, and 9 (70%) were born to mothers whose HIV status was unknown at the time of delivery. Eighty percent (80%) of HIV-Ab-positive infants continued to receive follow-up care at Lincoln Hospital. Relocation to other health-care facilities occurred as a result of parental choice or due to foster care placement. No babies were "lost" from the NBS program. The average time between sampling and receipt of results for all blood tests was 16 days (range 10 to 141). Nearly 40% of newborns who acquired HIV infection from their mothers were unanticipated because the mother's HIV status was unknown at the time of delivery. These unanticipated HIV-infected infants represent missed opportunities for prevention of maternal-to-child transmission of HIV infection and early therapeutic intervention for HIV-infected infants. The new expedited HIV-testing program for New York State will facilitate early diagnosis, prevention and treatment of the HIV-exposed/infected infant for whom maternal HIV status is unknown at the time of delivery. PMID- 10874986 TI - Congenital intrapulmonary bronchogenic cyst in the neonate--perinatal management. AB - Approximately 50% of all congenital lung malformations are pulmonary and mediastinal bronchogenic cysts (BC). Therefore, their diagnosis and management is of clinical importance. Usually asymptomatic in the first months of life, bronchogenic cysts are frequently clinically inapparent even adulthood. Early diagnosis and elective surgery can prevent late complications such as pneumothorax, pulmonary hypertension, and recurrent infections; prognosis after surgery is excellent. If mediastinal shifting is present, fetal thoracocentesis is indicated to prevent cardiovascular insufficiency. We report a case of a prenatally diagnosed intrapulmonary BC of the right lung. Following in utero thoracocentesis of the cyst and transient spontaneous regression postnatal onset of severe clinical symptoms due to rapidly developing hyperinflation and mediastinal shifting within the first days of life required early surgical intervention. PMID- 10874987 TI - Disorders of maternal calcium metabolism implicated by abnormal calcium metabolism in the neonate. AB - Normal fetal and neonatal calcium homeostasis is dependent upon an adequate supply of calcium from maternal sources. Both maternal hypercalcemia and hypocalcemia can cause metabolic bone disease or disorders of calcium homeostasis in neonates. Maternal hypercalcemia can suppress fetal parathyroid function and cause neonatal hypocalcemia. Conversely, maternal hypocalcemia can stimulate fetal parathyroid tissue causing bone demineralization. We report two asymptomatic women, one with previously unrecognized hypoparathyroidism and the other with unrecognized familial benign hypercalcemia, who were diagnosed when their newborn infants presented with abnormalities of calcium metabolism. J.B. was born at 34 weeks' gestation with transient hyperbilirubinemia and thrombocytopenia. At 1 month of age he had severe bone demineralization, cortical irregularities, widening and cupping of the metaphyses, and lucent bands in the scapulae. The total serum calcium and phosphorus were normal with an ionized calcium of 5.4 mg/dL (4.6-5.4). His alkaline phosphatase, parathyroid hormone, and 1,25-dihydroxyvitamin D levels were all increased. P.B., mother of J.B., had no symptoms of hypocalcemia either prior to, or during this pregnancy. She had severe hypocalcemia and hyperphosphatemia, laboratory values typical of hypoparathyroidism. J.N. presented at 6 weeks of age with new onset of seizures and tetany secondary to severe hypocalcemia. The serum phosphorus, creatinine, alkaline phosphatase, and parathyroid hormone levels were normal. At 15 weeks of age his calcium was slightly elevated with a low fractional excretion of calcium. P.N., mother of J.N., had no symptoms of hypercalcemia either prior to, or during this pregnancy. Her serum calcium was 12.7 mg/dL and urine calcium was 66.5 mg/24 hr, with a low fractional excretion of calcium ranging from 0.0064 to 0.0073. P.N. has a brother who previously had parathyroid surgery. Both J.N. and P.N. meet the diagnostic criteria for familial benign hypercalcemia. These cases illustrate the important relationships between maternal serum calcium levels and neonatal calcium homeostasis. They emphasize the need to assess maternal calcium levels when infants are born with abnormal serum calcium levels or metabolic bone disease. PMID- 10874988 TI - Transient marked atelectasis: an unusual complication of asthma in pregnancy. AB - We present an unusual case in which a patient with asthma presented with acute respiratory distress of acute onset, secondary to marked atelectasis of the right middle and lower lobes, which resolved within 24 hr following administration of increased doses of intravenous steroids, inhalation therapy (beta-agonists and steroids), and pulmonary physiotherapy. This transient occurrence responding to basic therapeutic measures was considered consistent with the release of a mucous plug which had caused the above obstruction and associated symptomatology and radiographic findings. This case illustrates and supports the practice of chest imaging in patients with atypical presentations of asthma and stresses the importance of pulmonary physiotherapy and bronchodilatory therapy as primary therapeutic agents in cases of mucous plug-associated atelectasis. PMID- 10874989 TI - Assessment of cord blood IL-6 levels as an indicator of neonatal sepsis. AB - Based on the recognition that interleukin-6 (IL-6) is produced early in infection, IL-6 determinations have been used to identify infants with early onset bacterial sepsis. This study intended to assess the value of IL-6 in maternal, cord and infant peripheral blood as an index of sepsis, and examine the relationships of its values in mother and infants. The population consisted of 17 mother/infant pairs at high risk for neonatal infection. Eight of these infants had clinical signs of possible sepsis. Cord blood IL-6 levels in infants of mothers considered to be noninfected were lower than those born to women with chorioamnionitis. There was also a positive correlation between maternal and cord blood IL-6 values. There were no differences in maternal blood IL-6, whether they had infections or not. Also, peripheral infant blood obtained after birth did not differentiate between those born to women with or without chorioamnionitis, nor did it correlate with maternal blood IL-6 levels. Clinical symptoms of the infants did not correlate with either cord or peripheral blood IL-6 values. Although maternal prepartum treatment with antibiotics and/or steroids may influence their own and their infants' blood IL-6 levels, there is insufficient evidence to consider low infant blood IL-6 level a reliable predictor to rule out early newborn sepsis. PMID- 10874990 TI - Aplasia cutis congenita in an infant of an initial triplet gestation: a case report. AB - Aplasia cutis congenita (ACC) is the clinical manifestation of an uncommon group of skin disorders. One postulated etiology is disseminated intravascular coagulation from release of thrombogenic maternal arising from placental injury or fetal demise. This leads to disruption of the ectodermal blood supply responsible for the skin defects. We present a neonate with group V ACC, one of an initial triplet gestation, associated with fetal demise at 14 weeks and formation of a fetus papyraceus. The practice of selective fetal reduction as a result of multiple gestation seen with the use of fertility drugs may in theory increase the incidence of group V ACC. PMID- 10874991 TI - Pattern analysis for ultrasound anomalies in fetuses with normal karyotype. AB - Two hundred cases of double or multiple anomalies (up to five in the same fetus) out of 828 karyotypically normal fetuses with at least one ultrasound finding were enrolled in the present study. One hundred and sixty patterns belonging to 200 fetuses were found. Then, we included in the final analysis only those double or triple patterns of anomalies with frequency more then 1. Thus, 123 cases and 83 patterns were analyzed. A double pattern was intended as any combination of ultrasound findings present in the same fetus. Exact Chi-square test was used to evaluate the specificity of the association of the most frequent patterns. The association of patterns was expressed as ratio between observed and expected frequency (O/E) ratio. Pattern analysis was used as statistical tool to calculate the number of possible associations of ultrasound finding. Anomalies of fluid distribution, as well as Central Nervous System malformations, and facial malformations were among the most significant associations. Among the triple patterns, Cleft Lip + Cleft Palate + Holoprosencephaly showed the highest degree of association. Antenatal diagnosis of ultrasound finding can benefits by considering the present results. PMID- 10874992 TI - Size at birth in an inner-city population. AB - Recent comparisons of growth curves and incidence of low birth weight are available for Caucasians and African-Americans or Hispanics. To compare size at birth in Hispanics with African-Americans in an inner-city population, we analyzed data on mother-infant pairs admitted to Jacobi Medical Center from January 1, 1995 until May 31, 1997 and those with a gestational age less than 34 weeks admitted from June 1, 1997 until December 31, 1997. The population mostly included mothers covered by Medicaid. The study sample included 2852 infants (1641 Hispanics and 1211 African-Americans). Among singletons, the incidence of teenage pregnancy was higher and those of premature delivery and low birth weight were lower in Hispanics than in African-Americans. Among full-term singletons, boys were heavier than girls (difference 116 g, confidence interval (CI) 57,174, p < 0.001 after adjusting for gestational age, gender, and teenage pregnancy), and Hispanics were heavier than African-Americans (adjusted difference 70 g, Confidence Interval (CI) 11,128, p = 0.019). Additional studies are needed to assess the effect of race on neonatal size after adjusting for differences in prepregnancy weight, weight gain, parity, social class, or other factors. PMID- 10874993 TI - Variability of tidal breathing flow-volume loops in healthy and sick newborns. AB - Measurement of tidal breathing flow-volume loops (TBFVL) is a frequently used noninvasive method to investigate ventilation and pulmonary mechanics in newborns. To investigate their intrasubject and intersubject variability shapes of averaged TBFVLs in 56 healthy newborns (group 1: median age and weight 7 days, 3100 g), 19 infants recovering from respiratory diseases after neonatal care (group 2: 16 days, 2770 g), and 38 infants with bronchopulmonary dysplasia (BPD) (group 3: 80 days, 2465 g) were analyzed using the dead space free flow-through technique, which permits pneumotachographic long-term measurements. We found a low intrasubject but a high intersubject variability of shapes in all groups. The incidence of normal TBFVLs was similar in all groups (group 1: 66%, group 2: 53%, group 3: 61%). The shape of the expiratory limbs in infants with BPD did not differ significantly from healthy newborns with exception of the incidence of linear or concave shapes (92 vs. 73%, p < 0.05). Nevertheless, the shape of the TBFVL has a significant (p < 0.001) influence on commonly used tidal breathing parameters which must be considered in the clinical interpretation. Unless the shape of the TBFVL illustrates certain respiratory behaviors (e.g., flow limitation, grunting) the high inter-subject variability of TBFVLs limits the diagnostic value of a shape analysis during tidal breathing. PMID- 10874994 TI - Outcome of infants with a diagnosis of hydrops fetalis in the 1990s. AB - The diagnosis of Hydrops fetalis still carries a grave prognosis with reported mortality ranging from 50 to 100%. With the advent of more aggressive therapy, improvement of survival is undetermined. The study population of this outcome case series was gathered from all cases of hydrops fetalis admitted to our Loyola University Medical Center Neonatal Intensive Care Unit (NICU) from 1990 to 1997. Forty-one patients were eligible for inclusion. Only four had a diagnosis of immune hydrops fetalis, while the remainder had varied nonimmune causes. Models predicting survival were constructed with various neonatal and maternal factors as explanatory variables using Cox proportional Hazards technique. Kaplan-Meier estimates of median survival times for different stratifying variables were likewise computed. The overall mortality rate was 49% with an overall median survival time of 15 days (95% CI 8-38). Median survival time estimates differed significantly between patients who had (a) proven infection or not and (b) had less than or greater than two fluid-filled cavities. The use of steroids, surfactant, or high-frequency ventilation did not improve survival. Stratifying the study base into those treated in early or late 1990s likewise failed to show difference in survival times. Infection remains a significant problem (46%). In our series of 41 infants with hydrops fetalis, survival rates remain comparable to those reported in the literature, despite aggressive therapy. Although the use of surfactant, steroids, and high-frequency ventilation appear to prolong survival times, these treatments failed to alter overall survival outcome. PMID- 10874995 TI - Fuzzy control of bioprocess in Japan. AB - Process control of bioprocess has been carried out by the judgment of the experts, who are the skilled operators and have lots of experiences for the control of the process. In almost all cases, those experiences are described linguistic IF-THEN rules. Fussy inference is one of the powerful tools to incorporate the linguistic rules to the computer for process control. Fuzzy control are divided into two types; one is the direct fuzzy control of process variables such as sugar feed rate in fed-batch culture and fermentation temperature in batch operation. The other is the indirect control of bioprocess, in which at first the phase recognition is carried out by fuzzy inference and the control strategies constructed in each phase are used for the control of process variables. In Japan, the fuzzy control has already been applied to practical industrial productions, such as pravastatin precursor, vitamin B2, and Japanese sake mashing process. In this review, these industrial approaches of fuzzy control are introduced. PMID- 10874996 TI - Rapid in silico cloning of genes using expressed sequence tags (ESTs). AB - Expressed sequence tags (ESTs) are short single-pass DNA sequences obtained from either end of cDNA clones. These ESTs are derived from a vast number of cDNA libraries obtained from different species. Human ESTs are the bulk of the data and have been widely used to identify new members of gene families, as markers on the human chromosomes, to discover polymorphism sites and to compare expression patterns in different tissues or pathologies states. Information strategies have been devised to query EST databases. Since most of the analysis is performed with a computer, the term "in silico" strategy has been coined. In this chapter we will review the current status of EST databases, the pros and cons of EST-type data and describe possible strategies to retrieve meaningful information. PMID- 10874997 TI - Functional genomics with protein-protein interactions. AB - Knowing the sequence of a gene does not mean knowing its function. Although, information stored at the DNA level can be used to predict biological processes, proteins are the final executors of the various response programs of a cell. Transient information, like posttranslational modifications or interactions among proteins, cannot be deduced from DNA sequences. The rapid accumulation of large amounts of DNA sequence data in genomics projects has led to an increasing demand for powerful tools to analyze proteins and their behaviour at a large scale. This review aims to compare different technologies used for identification of interacting proteins and discusses recent developments in the field of high throughput protein-protein interaction mapping. PMID- 10874998 TI - Biotechnical use of polymerase chain reaction for microbiological analysis of biological samples. AB - Since its introduction in the mid-80s, polymerase chain reaction (PCR) technology has been recognised as a rapid, sensitive and specific molecular diagnostic tool for the analysis of micro-organisms in clinical, environmental and food samples. Although this technique can be extremely effective with pure solutions of nucleic acids, it's sensitivity may be reduced dramatically when applied directly to biological samples. This review describes PCR technology as a microbial detection method, PCR inhibitors in biological samples and various sample preparation techniques that can be used to facilitate PCR detection, by either separating the micro-organisms from PCR inhibitors and/or by concentrating the micro-organisms to detectable concentrations. Parts of this review are updated and based on a doctoral thesis by Lantz [1] and on a review discussing methods to overcome PCR inhibition in foods [2]. PMID- 10874999 TI - Transcription of the insecticidal crystal protein genes of Bacillus thuringiensis. AB - Production of a large amount of insecticidal crystal proteins encoded on large plasmids is largely dependent upon the mother cell, Bacillus thuringiensis (B. thuringiensis, also Bt), specific transcription systems attributable to sporulation. In the middle stages of sporulation, cry4A is most actively transcribed from the promoter cry4A-P1. The proximal transcriptional start point of cry4A, which is under the control of the promoter P1, is used in Bacillus subtilis (B. subtilis) in the middle stage of sporulation. The nucleotide sequence that determines the cry4A-P1 promoter is homologous to the consensus sequence for the promoter of sigma E-specific genes in B. subtilis, and to those promoters of the insecticidal protein genes that are efficiently transcribed in vitro with the RNA polymerase E sigma 35 isolated from B. thuringiensis. The sigma factor sigma 35 of B. thuringiensis is highly homologous and functionally equivalent to sigma E of B. subtilis. These results suggest that the cry4A transcription from P1 is under the control of sigma E in B. subtilis, and under the control of sigma 35 in B. thuringiensis. PMID- 10875000 TI - Synthetic oligonucleotides as therapeutics: the coming of age. AB - Synthetic oligonucleotides (ODNs) are short nucleic acid chains that can act in a sequence specific manner to control gene expression. Significant progress has been made in the development of synthetic ODN therapeutics since the first demonstration of gene inhibition by antisense ODNs in a cell culture system two decades ago. This new class of therapeutic agents can potentially target any abnormally expressed genes in a broad range of diseases from viral infections to psychoneurological disorders. A number of "first" generation synthetic ODNs have entered into human clinical trials in the last few years. The eminent approval of the first ODN for the treatment of cytomaglovirus retinitis by the FDA in USA will provide much excitement that this new class of compounds holds great promise as a therapeutic "magic bullet". However, many obstacles still exist in the development of this technology. In this review, the current status of synthetic ODN chemistry, drug delivery methods, mechanisms of ODN action, potential clinical applications and its limitations in a wide range of human disorders will be described. PMID- 10875001 TI - Eukaryotic gene transfer with liposomes: effect of differences in lipid structure. AB - Liposome mediated gene transfer has a great potential in gene therapy. In this review we discuss the physical and chemical properties of cationic liposomes that affect their abilities to mediate gene transfer into eukaryotic cells. The specific focus is on functional domains of cationic lipids. We address polar head variations, counterions, linker bonds, acyl chain variations, as well as composition of liposomes. We additionally discuss different functional groups of lipids affecting lipid bilayer packing, lipid association with DNA, fusion with the cellular membranes and the release of transferred DNA from endosomes into the cytoplasm. PMID- 10875002 TI - Nonclinical safety evaluation of biotechnologically derived pharmaceuticals. AB - The primary objectives of nonclinical safety evaluation for pharmaceutical products are to identify potential target organ toxicity, provide a safe starting dose for clinical trials, and establish dose-response relationships. These objectives do not differ in concept for either small molecular weight compounds or biotechnologically derived pharmaceuticals; they are important for both. The complex structural and biological characteristics of biotechnologically derived pharmaceuticals, however, dictate that different approaches to their safety evaluation are needed. Although their novel mode of production initially raised concerns about their safety, improvements in analytical and manufacturing procedures have largely minimized the perceived risks. It is primarily their exaggerated pharmacodynamic properties that produce the toxicity observed in nonclinical studies. Even though most of these products will require a case-by case, scientifically based approach, knowledge gained from both nonclinical and clinical evaluation of these novel products have highlighted some general principles with regards to their safety evaluation. These principles include the importance of evaluating species in which the biotechnologically derived pharmaceutical is biologically active, the potential impact of immunogenicity on the interpretation of multiple dose toxicity study results, and the need for both highly sensitive and specific analytical methods to measure their pharmacodynamic properties. An understanding of these principles forms the basis for the development of a scientifically sound nonclinical safety evaluation program. PMID- 10875003 TI - Clinical research strategies in biotechnology. AB - Clinical drug development involves many steps and is extremely costly to the sponsoring company. There is intense pressure on sponsors to be faster, more efficient, and less costly. Sponsors also need to be globally oriented in their drug-development processes. There are several ways in which clinical drug development may be done more quickly and at less cost. These strategies include the use of large contract research organizations (CRO) and site management organizations (SMO). Although there is an estimated 2000 CRO worldwide, the use of SMO is quite limited but growing rapidly. Changes in the US Food and Drug Administration (FDA), especially harmonization between its two divisions Center for Biologics Evaluation and Research (CBER) and Center for Drug Evaluation and Research (CDER), and the use of CRO and SMO will make for interesting and challenging times for sponsors. PMID- 10875004 TI - Transgenic farm animals: applications in agriculture and biomedicine. AB - During the last decade, tremendous progress has been made in the area of transgenic farm animals. While there are many important transgenic farm animal applications in agriculture, funding has been very limited and progress has been rather slow in this area. Encouragingly, the potential applications of transgenic farm animals as bioreactors for producing human therapeutic proteins and as organ donors for transplantations in humans have attracted vast funding from the private sectors. Several transgenic animal products are already in various phases of clinical trials. Estimates are, that in the near future, the worlds demands on human pharmaceutical proteins may largely be met by transgenic farm animals. While there are still major challenges ahead in the area of xenotransplantation using transgenic animal organs, transgenic tissues or cells have demonstrated promising results as a potential tool for gene therapy. Recent development on cloning, embryonic stem cells and alternative transgenic methods may further expand the transgenic applications in both agriculture and biomedicine. PMID- 10875005 TI - [Current status of diagnosis and nonoperative therapy of small bowel ileus]. AB - In obstructed small bowel the number of stercoral bacteria increases due to stasis, inducing a liberation of mediators that are leading to ileus disease with facultative sepsis and breakdown of the circulatory system. The therapeutic procedures have to cure the stasis. In the case of mechanical obstruction cure is achieved by operation, while functional ileus may be healed by conservative treatment. Implantation of a gastric tube or Dennis' tube can evacuate the small intestine. Several peristalsis-inducing drugs are in use, although their clinical relevance remaines to be demonstrated. Clinical examination and anamnesis offer initial information; X-ray of the abdomen confirms the diagnosis of ileus and is the baseline examination. Addition of oral contrast medium, ultrasonography, CT and endoscopy may help to evaluate the underlying cause of small-bowel paralysis. PMID- 10875006 TI - [Prevention of adhesions. Wish and reality]. AB - For more than a century peritoneal adhesions are being recognized as frequent sequelae following abdominal surgery. Intraoperative lesions of the mesothelial lining by abrasion, ischemia, dissication, and foreign bodies result in complaints, intestinal obstruction, female infertility, and problems during reoperations. The global increase of life expectancy and surgical procedures are leading to rising incidences of adhesion-related complications and subsequent socio-economic implications. As of today, there is no safe and efficient prophylaxis available. Scientific efforts should be aimed at a liquid substance for single intraperitoneal application which significantly reduces postoperative adhesions at reasonable cost without adverse effects on blood coagulation and wound healing. PMID- 10875007 TI - [Laparoscopy in small bowel ileus]. AB - Today laparoscopic procedures are routinely performed in patients with intestinal adhesions from previous abdominal surgery. Does laparoscopy have a potential benefit in acute small-bowel obstruction? Theoretically, a lower rate of wound complications and incisional hernias, as well as less subsequent adhesions with a lower incidence of recurrent intestinal obstruction, can be expected. However, laparoscopy is successful in only 50-70% of selected patients, thereby representing the highest rate of conversion in minimally invasive surgery. Laparoscopic management of severe abdominal distension with massively dilated and fragile small-bowel or dense adhesions is extremely difficult even when performed by experienced surgeons. Significantly prolonged operating time, the high risk of bowel injury (> 6-10%) and an increased frequency of early reoperations jeopardize the patient's safe outcome. However, in strictly selected patients the laparoscopic approach may be promising. In acute intestinal obstruction without a history of previous abdominal surgery, laparoscopy is--in the absence of adhesions--an excellent diagnostic tool and may also be a successful therapeutic modality in a variety of bowel-obstruction etiologies. Furthermore, the laparoscopic option should be considered in patients who previously had undergone small laparotomies (e.g., appendectomy) or laparoscopic surgery. We recommend "postlaparoscopic" intestinal obstruction as the ideal case for laparoscopic reexploration. Incarcerated hernias at the site of trocar insertion or adhesions due to peritoneal tears are easily identified as the cause of obstruction and successfully cured with the laparoscope. In conclusion, we advocate the laparoscopic approach in acute small-bowel obstruction exclusively for selected patients. Clinical studies are required to define appropriate surgical indications objectively. PMID- 10875008 TI - [Abandoned, effective and current aspects of surgical small bowel ileus therapy]. AB - Because of the marked heterogeneity of patient groups and the diverse causes of obstruction, surgical therapy of the small-bowel ileus is difficult to standardize. A homogeneous strategy of intraoperative handling of the obstructed bowel (decompression, endoluminal splinting) as well as abdominal closure would be desirable. Adhesiolysis remains the most common procedure. Small bowel plication is still used by a minority of surgeons with the Noble's technique replaced by the Child-Phillips' technique and its modifications. However, as a result of the lack of controlled studies, there is no large body of evidence available to support the use of plication, long tubes or any other specific surgical intervention. This is an important task for the future, especially in view of incidence and economic importance of small bowel-ileus. PMID- 10875009 TI - [Identification of the recurrent laryngeal nerve. How invasive? How valuable?]. PMID- 10875010 TI - [Intraoperative electromyographic identification of the recurrent laryngeal nerve]. AB - BACKGROUND: Recurrent nerve palsy is the most important complication of thyroid surgery. Correct identification of the nerve during the surgical procedure is essential to avoid damage. METHOD: We evaluated the intraoperative electromyographic identification procedure of the recurrent laryngeal nerves in 77 consecutive patients. The operations performed were 63 subtotal resections, 6 hemithyroidectomies, 4 hemithyroidectomies in combination with subtotal resection of the opposite side and 4 thyroidectomies. RESULTS: Identification of the recurrent laryngeal nerve by electrophysiologic nerve stimulation was successful in 97.4%. Recurrent nerve palsy was not seen in postoperative laryngoscopic control. CONCLUSIONS: Intraoperative electromyography improves the identification rate of the recurrent laryngeal nerve and allows electrophysiologic verification of nerve function as a result of injury to the thyroid. PMID- 10875011 TI - [Recurrent laryngeal nerve paralysis as intubation injury?]. AB - INTRODUCTION: Vocal cord paralysis is a important complication in thyroid gland surgery. A prospective study was conducted ascertain the frequency of laryngeal nerve palsy not caused by surgical trauma. PATIENTS AND METHODS: Two hundred and ten patients were investigated laryngoscopically pre- and postoperatively after short-term intubation in the course of operations far removed from thorax or neck region. We noted the inner diameter of the tube, intubation problems, the qualification of the anaesthesiologist and the positioning of the patient. RESULTS: Preoperatively we found previously unknown unilateral laryngeal nerve palsy in 1.9% of cases. Postoperatively there were pathological findings of vocal cords in 13 patients (6.2%). In 10 patients a direct lesion (oedema, rubor, haematoma, granuloma) was established. Three patients (1.4%) were found to have a movement disorder caused by a neural lesion without morphological findings, leading to restitutio ad integrum in two of three cases after 6 months. CONCLUSION: The frequency of intubation-related recurrent nerve palsy is 1.4% transiently and 0.5% permanently. The reasons are discussed. Preoperative laryngoscopic investigation of vocal cords should be carried out before intubation. PMID- 10875012 TI - [Intraoperative electromyographic recurrent laryngeal nerve identification as a routine measure]. AB - In recent years, two methods of intraoperative monitoring of the laryngeal nerve have mostly been used: evoked electromyographic responses via endscopically applied needle electrodes inserted into the adducting laryngeal muscles, and non invasive electrodes like special tubes with integrated electrodes or separately insertable electrodes like the postcricoid electrode or disposable electrodes attached to the tube, as used in this study. The incidence of recurrent nerve paresis or paralysis during the IRM period was 1/174 nerves (0.6%). The advantage of the IRM is the quick and certain identification of the nerve; intraoperative monitoring cannot replace a proper surgical technique. We conclude that the IRM, using a laryngeal surface electrode attached to the tube, is a safe and reliable method. PMID- 10875013 TI - [First continuous nerve monitoring in thyroid gland surgery]. AB - A new "all in one" sensing device was developed for continuous transtracheal intraoperative monitoring and in situ detection of the recurrent laryngeal nerve (RLN) during thyroid surgery. PATIENTS AND METHODS: The new system is based on a double-balloon endotracheal tube with integrated atraumatic stimulating and tracing electrodes. The recurrent laryngeal nerve is stimulated transtracheally and compound action potentials are recorded from the laryngeal muscles. Fifty five patients were introduced into a phase-one clinical trial. Thirty-five patients with primary thyroid operations, 20 patients with reoperations, 10 of whom had neck dissections. All patients were evaluated laryngoscopically and phoniatrically by an ENT specialist before and after surgery. RESULTS: Compound muscle action potentials were recorded continuously during the whole operation and responded sensitively to tension and pressure to the nerve. There were no accidental permanent RLN palsies. CONCLUSION: The new system offers five advantages: (1) it is atraumatic; (2) it is easy to use; (3) it can monitor continuously with an audio feedback to the surgeon; (4) it works outside the operation field; and (5) it is highly sensitive, even indicating reversible irritation to the nerve. PMID- 10875014 TI - [Administration of Integra on primary burn wounds and unstable secondary scars]. AB - We report on our experience with Integra application in 22 patients treated from February 1996 through April 1999. Nine patients suffered a primary burn injury and 13 patients had secondary unstable scars. Integra is artificial skin that consists of a collagen/glycosaminoglycane matrix and a silicon layer. It was first used by Burke and Yannas in 1980 in the USA. The mechanical and biological features of Integra protect the tissue from fluid loss and bacterial invasion in the acutely burned patient. The integra-induced formation of a neodermis leads to functionally and aesthetically highly acceptable scar formation. We present our clinical experience with Integra in long-term follow-up. PMID- 10875015 TI - [Repair of inguinal hernia in the elderly. Results of the plug-and-patch repair with special reference to quality of life]. AB - In a prospective study the perioperative results of plug-and-patch repair were investigated in patients > or = 65 years, and quality of life was assessed using the SF36 preoperatively and 3 months after the procedure in 34 consecutive patients. From August 1994 to February 1999 147 patients with a mean age of 73 +/ 5 years (65-92 years) were operated on using the plug-and-patch technique, mostly under local anesthesia (LA: n = 124, 84%, ITN: n = 23, 16%). Preoperative risk factors were alcohol consumption, hypertonus, diabetes mellitus, ischemic heart disease, smoking, cerebrovascular disease, hyperlipidaemia and pulmonary disease. Most of the patients were ASA II (ASA I: n = 14, 9%, ASA II: n = 82, 56%, ASA III: n = 51, 35%). No intraoperative complications occurred, postoperative complications consisted of superficial wound hematoma (n = 6, 3.7%) and infection (n = 1, 0.6%), seroma (n = 7, 3.8%), urinary retention (n = 3, 1.8%) and ilioguinal pain syndrome (n = 3, 3.8%). The total amount of postoperative analgesic consumption was 4.9 +/- 1.8 g Novalgin for about 4 +/- 3 days. The duration of postoperative hospitalization was 2 +/- 1 days and limitation of daily activities 6 +/- 3 days. Clinical examinations after 3 months revealed no recurrence or late complications. Investigation of quality of life showed a significant improvement in the SF36 domains of physical activity, pain, vitality, and social functioning after the operation. No significant change was observed for physical, emotional, and global health. PMID- 10875016 TI - [Iatrogenic colon perforation from the viewpoint of the surgeon. Experiences with 11 patients]. AB - Iatrogenic colonic perforation is a rare but very dangerous complication of colonoscopy. We report on eleven prospectively recorded patients treated between 1994 and 1998. In seven cases the colonoscopy had been done for diagnostic and in four cases for therapeutic reason. Nine women and two men (age range 56 to 92 years) were affected. In 77% the perforation was found in the sigmoid colon. All patients were operated on and five sutures, five resections and one colostomy were performed. No significant differences could be shown in six patients with classical perforation compared with five patients presenting with prolonged abdominal pain. Only duration of symptoms was significantly different in the two groups. Three patients (85, 90 and 92 years) died from late complications. PMID- 10875017 TI - [Perforation of mucocele-like transformed Hartmann stump]. AB - We report an unusual complication in a 53-year-old woman following ileostomy for Crohn's disease 22 years previously. A stenosis of the distal colonic segment was the reason for the formation and subsequent rupture of a huge colonic mucocele. To our knowledge, this is the first report of a ruptured mucocele of colonic origin after ileostomy. PMID- 10875018 TI - [Multiple metachronous metastases of renal cell carcinoma in the pancreas. Case report and review of the literature]. AB - Isolated pancreatic metastases from renal-cell carcinomas (RCC) are extremely rare. Only 96 cases of clinically diagnosed renal-cell carcinoma metastatic to the pancreas have been reported in the world literature, and 70 of the patients (including ours) underwent a definitive surgical resection. In many cases the time between the nephrectomy and diagnosis of metachronous metastases is reported to exceed 10 years. Therefore, the initial diagnosis may be overlooked when there is a prolonged disease-free interval. When it does occur simultaneously or metachronously, aggressive surgical resection, when possible, seems to be the most effective treatment for this metastatic lesion. Surgical resection can provide long-term survival in selected cases. We present the case of a 69-year old woman in whom two pancreatic metastases were treated by a left-sided subtotal pancreatectomy with splenectomy, 12 years after radical nephrectomy for a RCC. The patient simultaneously presented with both a mass in the body of the pancreas and a right-sided colon cancer. Thus, the diagnosis of pancreatic metastasis of colon cancer was suspected initially. Both tumors were radically resected, and histological examination revealed two pancreatic metastases from the previous RCC. In the world literature this report represents the eighth case of multiple pancreatic metastases due to RCC. It illustrates a rare indication of pancreatic resection because of pancreatic metastasis. The need for prolonged follow-up of patients is emphasized. The few reports on surgically treated pancreatic metastases from RCC are reviewed after the presentation of this case. PMID- 10875020 TI - [Werner Wachsmuth. Memories of the 1946 to 1947 new beginning]. PMID- 10875019 TI - [Image tracking system. A new technique for safe and cost-saving laparoscopic operation]. AB - The potential for improvement of the results of laparoscopic operations as well as necessity of enhanced efficiency in the health-care systems are the main reasons for development and practical use of robotic systems in the field of laparoscopic surgery. While robotic systems imitate the human camera-holder the Image Tracking System (ImagTrac, Olympus, Tokio) is based on another principle: A voice-activated zoom function allows change between overview and detailed view. In the zoom-in position it is possible to select four different fields of view. The results of a clinical trial with control group show that the system: 1. Makes it possible to dispense with the human camera-holder without compromising patient safety, sometimes at greater convenience to the surgeon. 2. Makes it possible for routine laparoscopic operations such as laparoscopic cholecystectomy and laparoscopic hernia repair to be performed (as solo surgery) by a team of a surgeon and a nurse only. 3. Is more cost-effective than robotic systems with a similar range of features. PMID- 10875021 TI - [Comment on B. Klosterhalfen et al.: Pathology of traditional surgical mesh for hernia repair after long-term implantation in the human]. PMID- 10875022 TI - [Atypical lung parenchyma resection with the Hydro-Jet--initial experimental and clinical experiences]. PMID- 10875023 TI - [Anastomosis techniques in the gastrointestinal tract]. PMID- 10875024 TI - [Qualifications profile of specialty graduate education for surgical assistant physicians from the viewpoint of surgically active senior surgeons]. PMID- 10875025 TI - [Necessary reference to the life threatening state of an infection. On the surgeon's management of hesitant patients]. PMID- 10875026 TI - What is structural heart disease? PMID- 10875027 TI - On time to heal: observations and a review. PMID- 10875028 TI - Hyperthyroid heart disease. AB - The heart is an organ sensitive to the action of thyroid hormone, and measurable changes in cardiac performance are detected with small variations in thyroid hormone serum concentrations. Most patients with hyperthyroidism experience cardiovascular manifestations, and the most serious complications of hyperthyroidism occur as a result of cardiac involvement. Recent studies provide important insights into the molecular pathways that mediate the action of thyroid hormone on the heart and allow a better understanding of the mechanisms that underlie the hemodynamic and clinical manifestations of hyperthyroidism. Several cardiovascular conditions and drugs can interfere with thyroid hormone levels and may pose a difficulty in interpretation of laboratory data in patients with suspected thyroid heart disease. The focus of this report is a review of the current knowledge of thyroid hormone action on the heart and the clinical and hemodynamic laboratory findings as well as therapeutic management of patients with hyperthyroid heart disease. PMID- 10875029 TI - Low-dose dobutamine radionuclide ventriculography for prediction of myocardial viability: quantitative analysis of regional left ventricular function. AB - BACKGROUND: It is important to distinguish viable myocardium from necrotic tissue in order to decide upon therapy in patients with ischemic heart disease. HYPOTHESIS: We verified the hypothesis that quantitative analysis of regional left ventricular function using low-dose dobutamine radionuclide ventriculography (RNV) can sensitively predict myocardial viability and compared its usefulness with thallium-201 (201Tl) single-photon emission computed tomography (201Tl SPECT). METHODS: Radionuclide ventriculography at rest and during low-dose dobutamine infusion (5 micrograms/kg/min), 201Tl-SPECT, and coronary angiography were performed in 51 subjects with severe ischemia-related stenosis of coronary arteries and 3 subjects without coronary artery disease. 201Tl uptake was assessed as normal (control), low perfusion (LP), or defect. We compared the response of regional function to dobutamine with the regional 201Tl uptake. The accuracy of both methods for identifying viable myocardium was investigated in 17 patients who underwent successful coronary revascularization, with a resulting improvement in wall motion. RESULTS: The increase in regional ejection fraction (delta r-EF) in response to dobutamine was significantly greater in the control (12 +/- 6%) and LP (16 +/- 11%) regions than in the defect (5 +/- 10%) regions. The increase in one-third regional ejection fraction (delta r-1/3EF) was also significantly higher in the control (14 +/- 7%) and LP (10 +/- 8%) regions than in the defect regions (5 +/- 6%). We defined myocardial viability as a delta r-EF > 5% or a delta r-1/3EF > 2%. The sensitivity and specificity of the delta r-EF for identification of myocardial viability were 91.4 and 55.5%, respectively. The sensitivity and specificity of the delta r-1/3EF were 91.4 and 66.6%, respectively; the corresponding values for 201Tl SPECT were 74.2 and 77.8%. CONCLUSION: Low-dose dobutamine RNV with quantitative analysis of regional left ventricular function was more sensitive for identification of viable myocardium than 201Tl-SPECT. PMID- 10875030 TI - Effects of intravenous dofetilide in patients with frequent premature ventricular contractions: a clinical trial. AB - BACKGROUND: Although suppression of premature ventricular contractions (PVCs) is not a predictor of mortality over the long term, the extent of PVC suppression is an important characteristic of any antiarrhythmic drug. HYPOTHESIS: This study was undertaken to determine whether intravenous (i.v.) dofetilide has the ability to suppress PVCs in patients who have frequent occurrences. METHODS: Subjects were men and women, aged 18 to 75 years, with > 30 PVCs/h on two consecutive 24-h Holter recordings while drug free, and > 50 PVCs/h during a 2-hour telemetric electrocardiogram. The study was randomized, double-blind, and placebo controlled. Subjects received a single-blind, i.v. infusion of placebo and were randomized (3:1) to receive a double-blind second infusion of placebo or an infusion of dofetilide (a 15-min loading infusion of 4 g/kg followed by a 60-min maintenance infusion of 3.5 g/kg, for a total dose of 7.5 g/kg). RESULTS: Dofetilide produced an 82.6% and placebo a 2.9% median reduction in PVCs. Drug responder rate, defined as 80% reduction in PVCs, was 50% in the dofetilide group and 0% in the placebo group. CONCLUSION: Intravenous dofetilide significantly reduced PVCs in patients who had > 30 PVCs/h at baseline, and it produced > or = 80% reduction in PVCs in 50% of all subjects. PMID- 10875031 TI - Comparison of echocardiography and electron beam tomography in differentiating the etiology of heart failure. AB - BACKGROUND: The clinical manifestations in patients with ischemic cardiomyopathy are often indistinguishable from those in patients with primary dilated cardiomyopathy (DCM). Clinicians often base work-up of patients with heart failure on echocardiographic wall motion abnormalities; however misclassification can lead to unnecessary coronary angiography. HYPOTHESIS: The study was undertaken to evaluate the diagnostic ability of echocardiography and electron beam tomography (EBT) to differentiate between ischemic and nonischemic cardiomyopathy. METHODS: The accuracy of EBT and echocardiography was compared in 111 patients undergoing coronary angiography for the evaluation of heart failure. The presence of coronary calcification (CC) by EBT or segmental wall motion abnormalities by echocardiography was used as evidence of coronary-induced cardiomyopathy. RESULTS: Of 63 patients, 61 (97%) with obstructive coronary artery disease had CC by EBT. This sensitivity was significantly higher compared with 43 of 63 patients (68%) with segmental wall motion abnormalities by echocardiography (p < 0.001). Of 48 patients without obstructive coronary artery disease by angiography, 39 (81%) had no CC by EBT and 35 (73%) had no segmental wall motion (global hypokinesis) by echocardiography (p = 0.33). The overall accuracy of EBT to differentiate ischemic from nonischemic cardiomyopathy was 90%, significantly higher than echocardiography (70%, p < 0.001). CONCLUSION: This double-blind study demonstrates that the presence of CC by EBT is superior to that of segmental wall motion abnormalities by echocardiography to distinguish ischemic from nonischemic cardiomyopathy. This modality may prove to be an important diagnostic tool when the etiology of the cardiomyopathy is not clinically evident. PMID- 10875033 TI - Response to changing plasma concentrations of isosorbide-bound NO2 during acute and sustained treatment with isosorbide dinitrate in patients with coronary artery disease. AB - BACKGROUND: The mechanisms behind development of tolerance to nitrate effects during sustained, asymmetric isosorbide dinitrate (ISDN) therapy are not fully understood. HYPOTHESIS: The study was undertaken to investigate the changes of the relationships between left ventricular (LV) function and plasma concentrations of ISDN and its vasoactive metabolites (2- and 5-ISMO) during acute and sustained, asymmetric ISDN therapy. METHODS: Left ventricular function and plasma concentrations of ISDN, 2- and 5-isosorbide mononitrates (P-ISDN, P-2- and 5-ISMO) were measured at rest and at supine exercise before and for 4 h after peroral 30 mg ISDN in 15 patients with coronary artery disease, all with initial exercise pulmonary artery wedge pressure (PAWP) > 25 mmHg. Seven patients were untreated (acute group), while eight received 30 mg ISDN b.i.d. for 2 weeks before the invasive study. P-ISDN and the concentration of available isosorbide bound nitrate (NO2) in plasma (P-ISDN.2 + P-2-ISMO + P-5-ISMO) (P-NO2) were used as measures of the nitric oxide (NO) offer to the tissues. RESULTS: Throughout the study, after administration of medication, all plasma concentrations, in particular P-ISDN, were higher in the chronic than in the acute group. Peak P ISDN was reached after 15 min in the chronic group and after 25 min in the acute group, while P-2- and 5-ISMO reached maximum only after 40 min in both groups. At rest, the full effect on PAWP was observed after 10 min in both groups, but at markedly higher levels of P-ISDN and P-NO2 in the chronic group. Afterward, no further changes in PAWP were observed. During exercise, 1 h after medication, PAWP and stroke index to PAWP ratio (SI/PAWP) were normal in both groups. Thereafter, at slowly declining P-NO2, PAWP rose and SI/PAWP declined toward the initial level in the chronic group, but remained unchanged in the acute group, in spite of higher P-NO2 and greater NO release in the former. CONCLUSIONS: Patients receiving sustained, asymmetric 30 mg ISDN b.i.d. dosing had the same immediate beneficial effects on LV function during exercise after a morning dose as did untreated patients. However, in spite of higher P-NO2 and higher rate of NO release during sustained treatment, the effects deteriorated gradually 2 to 3 h after medication. The changes in metabolism and/or distribution of isosorbide bound NO2 may possibly be part of the tolerance induced by long-term treatment, even with asymmetric dosing. PMID- 10875032 TI - Mortality, risk indicators for death, and mode of death in younger and elderly patients during five years after coronary artery bypass graft. AB - BACKGROUND: The number of elderly patients who may be candidates for coronary artery bypass graft (CABG) for severe coronary artery disease has increased. Cardiac surgery in the elderly is a high-risk procedure because many of these patients have concomitant systemic disease and other disabilities. HYPOTHESIS: The study was undertaken to evaluate mortality, risk indicators for death, and mode of death in younger and elderly patients during 5 years after CABG. METHODS: The study included all patients in western Sweden who underwent CABG without concomitant valve surgery and without previously performed CABG between June 1988 and June 1991. In all, 2,000 patients, of whom 953 (48%) were > or = 65 years, were divided into two age groups (< 65 years and > or = 65 years). RESULTS: Compared with the younger patients, the elderly had a relative risk of death of 2.3 (95% confidence interval 1.8-3.0). The increased risk of death in the elderly was significantly more marked in men, in patients with more severe angina pectoris, and in patients without a history of cerebrovascular diseases. The mode and place of death appeared similar regardless of age; neither was there marked difference in symptoms of angina pectoris among survivors 5 years after CABG. CONCLUSION: Compared with patients < 65 years, the elderly have more than twice as high a risk of death during the subsequent 5 years, and this risk is higher in men, in patients with severe symptoms of angina pectoris, and in those with no history of cerebrovascular disease. PMID- 10875034 TI - Metabolic and clinical effects of oral magnesium supplementation in furosemide treated patients with severe congestive heart failure. AB - BACKGROUND: Magnesium depletion and hypomagnesemia are common among furosemide treated patients with chronic congestive heart failure. HYPOTHESIS: This investigation evaluated clinical and metabolic effects of oral magnesium supplementation. METHODS: Ten patients with severe congestive heart failure maintained on high dose furosemide (> or = 80 mg/day) received a supplement of oral magnesium citrate 300 mg/daily for 30 days. Clinical parameters were followed, and peripheral blood mononuclear cell magnesium and zinc content, serum and urine magnesium, potassium, zinc, calcium, phosphorus, and creatinine were assessed. RESULTS: Peripheral blood mononuclear cell magnesium content and serum potassium rose significantly at the end of the study (2.09 +/- 1.89 to 3.99 +/- 2.26 micrograms/mg cell protein, p < 0.05, and 4.17 +/- 0.38 to 4.39 +/- 0.27 mEq/l, p < 0.05, respectively), while the other parameters remained unchanged. CONCLUSION: In some of these patients, oral magnesium supplementation is effective in achieving substantial increments in intracellular magnesium and serum potassium which, in turn, may have cardioprotective effects. PMID- 10875035 TI - Treatment of hypertension with perindopril reduces plasma atrial natriuretic peptide levels, left ventricular mass, and improves echocardiographic parameters of diastolic function. AB - BACKGROUND: Hypertension is a major independent risk factor for cardiac deaths, and diastolic dysfunction is a usual finding during the course of this disease. HYPOTHESIS: This study was designed to investigate the effects of chronic therapy with perindopril on left ventricular (LV) mass, left atrial size, diastolic function, and plasma level of atrial natriuretic peptide (ANP) in patients with hypertension. METHODS: Twenty four patients who had not been previously taking any antihypertensive medication and without prior history of angina pectoris, myocardial infarction, congestive heart failure, dysrhythmias, valvular heart disease, or systemic illnesses received 4-8 mg/day of perindopril orally. Echocardiographic studies were acquired at baseline and 6 months after the initiation of therapy. RESULTS: Systolic and diastolic blood pressure decreased from 174 +/- 19.7 and 107.5 +/- 7.8 mmHg to 134 +/- 10.6 and 82 +/- 6.7 mmHg, respectively (p < 0.001). Left ventricular mass decreased from 252.4 +/- 8.3 to 205.7 +/- 7.08 g and left atrial volume from 20.4 +/- 5.1 to 17.6 +/- 5.2 ml, respectively (p < 0.001). Transmitral Doppler early and atrial filling velocity ratio (E/A) increased from 0.69 +/- 0.06 to 0.92 +/- 0.05 m/s and plasma ANP level decreased from 71.9 +/- 11.7 to 35.3 +/- 7.8 pg/ml (p < 0.001). Reduction of LV mass correlated positively with a reduction in ANP levels (r = 0.66, p < 0.0005). CONCLUSIONS: Perindopril caused a significant reduction of LV mass, left atrial volume, and plasma ANP levels, as well as improvement in Doppler parameters of LV filling in this group of patients with hypertension. PMID- 10875036 TI - Normalization of left ventricular dysfunction in systemic hypertension. AB - BACKGROUND: In hypertensive heart disease, it is uncertain whether the impairment of left ventricular (LV) systolic function might be reverted by antihypertensive treatment. HYPOTHESIS: This study was undertaken to address the likelihood of recovery of LV dysfunction and to identify factors potentially related. METHODS: Twenty-six patients with primary (n = 16) and renovascular (n = 10) hypertension participated in the study and were classified into Groups A (n = 12) and B (n = 14) according to normalization or persistent left ventricular dysfunction (fractional shortening < 0.30) after 36 weeks of follow-up. All patients received standard medical therapy and appropriate procedures for renovascular disease correction. Logistic regression analysis was used to identify variables associated with recovery. RESULTS: Patients in Group A compared with those in Group B were younger (41 +/- 14 vs. 52 +/- 10 years; p < 0.05), had a greater frequency of renovascular hypertension (8 vs. 2; p < 0.05), showed shorter LV end diastolic (54 +/- 5 vs. 61 +/- 8 mm; p < 0.05) and end-systolic dimensions (41 +/ 6 vs. 49 +/- 9 mm; p < 0.05), and lower mass index (215 +/- 64 vs. 261 +/- 47 g.m-2; p < 0.05) before treatment, whereas fractional shortening (0.24 +/- 0.4 vs. 0.20 +/- 0.5; p > 0.05) and diastolic blood pressure (116 +/- 12 vs. 122 +/- 19 mmHg; p > 0.05) were similar. On follow-up, Group A patients showed lower diastolic blood pressure (89 +/- 15 vs. 105 +/- 20 mmHg; p < 0.05) and mass index (142 +/- 34 vs. 222 +/- 40 g.m-2; p < 0.05). Logistic regression analysis identified systolic dimension and renovascular hypertension as factors associated with fractional shortening normalization. CONCLUSION: The recovery of LV dysfunction is expected to occur most likely in patients with renovascular hypertension and the shortest systolic dimensions. PMID- 10875037 TI - Is ventricular repolarization heterogeneity a cause of serious ventricular tachyarrhythmias in aortic valve stenosis? AB - BACKGROUND: It is well known that there is a close relation between sudden cardiac death and serious ventricular tachyarrhythmias in patients with aortic valve stenosis (AS). QT dispersion (QTd) reflects the ventricular repolarization heterogeneity and has been proposed as an indicator for ventricular arrhythmias. HYPOTHESIS: This study investigated the QTd and its relevance to the clinical and echocardiographic variables. METHODS: In all, 51 patients (33 men, 18 women, mean age 56 +/- 12) with isolated AS and 51 age- and gender-matched healthy controls comprised the study group. Left ventricular mass index (LVMI) was calculated by the Devereux formula, and we used continuous-wave Doppler (n = 15) and cardiac catheterization (n = 36) for the determination of the maximum aortic valve pressure gradient (PG). RESULTS: Corrected QTd (QTcd) (89 +/- 39 vs. 49 +/- 15 ms, p < 0.001) and LVMI (176 +/- 69 g/m2 vs. 101 +/- 28 g/m2, p < 0.001) in patients with AS were significantly different from those in the control group. The group of 21 patients had a significantly greater number of 24-h mean ventricular premature beats (VPB) and mean number of couplet VT episodes than did the control group (p < 0.05). QTcd also correlated significantly well with LVMI (r = 0.58, p < 0.001), PG (r = 0.41, p = 0.003), and number of 24-h VPB (r = 0.56, p = 0.008). With respect to symptoms (e.g., angina, syncope, and dyspnea) patients without symptoms (n = 19) displayed less QTcd (71 +/- 31 vs. 100 +/- 39 ms, p = 0.007) and less LVMI (144 +/- 80 g/m2 vs. 195 +/- 57 g/m2, p = 0.01) than patients with symptoms. Statistical analysis was similar for all variables with uncorrected QTd values. CONCLUSION: We found that ventricular repolarization heterogeneity was greater in patients with AS than in controls. Our findings also showed that QTd in the patient group correlates well with LVMI, severity of AS, and PG. The present results suggest that serious ventricular arrhythmias in patients with AS may be due to spatial ventricular repolarization abnormality. PMID- 10875038 TI - The prediction of coronary atherosclerosis employing artificial neural networks. AB - BACKGROUND: Atherosclerosis is a complex histopathologic process that is analogous to chronic inflammatory conditions. Several factors have been shown to correlate with the extent of atherosclerosis. Whereas hypertension, obesity, hyperlipidemia, diabetes, smoking, and family history are all well documented, recent literature points to additional associated factors. Thus, antibodies to oxidized low-density lipoprotein (oxLDL), cytomegalovirus (CMV), Chlamydia pneumonia, Helicobacter pylori, as well as homocysteine and C-reactive protein (CRP) levels have all been implicated as independent markers of accelerated atherosclerosis. HYPOTHESIS: In the current study we attempted to formulate a system by which to predict the extent of coronary atherosclerosis as assessed by angiographic vessel occlusion. METHODS: The 81 patients were categorized as having single-, double-, triple-, or no vessel involvement. The clinical data concerning the "classic" risk factors were obtained from clinical records, and sera were drawn from the patients for determination of the various parameters that are thought to be associated with atherosclerosis. RESULTS: Using four artificial neural networks, we have found the most effective parameters predictive of coronary vessel involvement were (in decreasing order of importance) antibodies to oxLDL, to cardiolipin, to CMV, to Chlamydia pneumonia, and to beta 2-glycoprotein I (beta 2GPI). Although important in the prediction of vessel occlusion, hyperlipidemia, hypertension, CRP levels, and diabetes were less accurate. CONCLUSION: The results of the current study, if reproduced in a larger population, may establish an integrated system based on the creation of artificial neural networks by which to predict the extent of atherosclerosis in a given subject fairly and noninvasively. PMID- 10875039 TI - Double-outlet right ventricle after operative correction. PMID- 10875040 TI - Angina induced by excessive bradycardia. AB - Angina pectoris usually occurs in patients with significant coronary artery disease during periods of increased coronary demand such as increased heart rate. Bradycardia is a rare and unusal cause of angina pectoris that should be considered during the evaluation of patients with agnia. This paper reports three cases of bradycardia-induced angina. PMID- 10875041 TI - Norman E. Shumway. AB - Dr. Norman E. Shumway has had the privilege to live through the entire evolution of cardiac surgery. He is an intelligent, imaginative individual who possesses a true pioneering spirit. He has been blessed with the fundamental qualities of leadership and continues to be capable of inspiring greatness from his associates. Through his skill, perseverance, and courage he was able to affect the course of cardiac surgery. Despite the enormous accomplishments that he has achieved, he retains the qualities of humility, humor, and generosity. Dr. Shumway possesses a keen sense of intuition and charming quick wit that he uses to place everyone he meets at ease. PMID- 10875042 TI - Sleep physiology and pathophysiology. AB - Sleep is a complex, highly organized state that is fundamental to life. And yet, many functions of sleep remain to be discovered and understood. The past 50 years of modern sleep research clearly indicate what sleep is not--simply a resting brain, as popular notions and behavioral observations might suggest. This review will describe normal sleep physiology and its regulation as well as the major disorders of sleep and their underlying pathophysiology. PMID- 10875043 TI - Effective treatment of sleep disturbances in older adults. AB - Sleep disturbance is a common and complex clinical problem, particularly in older adults. With advancing age, the normal sleep cycle begins to break down, resulting in a reduction in the deeper stages of sleep and an often-profound increase in the fragmentation of nighttime sleep by periods of intrusive wakefulness. Sleep disorders exacerbate these age-related changes, leading to reports of daytime fatigue, sleepiness, and impaired daily function. The resultant chronic sleep deprivation invariably leads to often-unsuccessful attempts by the patient to overcome the problem. Patients may stay in bed longer, take more naps during the day, or seek out agents that putatively restore normal sleep patterns. Paradoxically, these efforts often result in exacerbation of the sleep disturbance. This review delineates the common causes of disordered sleep in older persons. Further, it reviews effective diagnostic approaches and treatments for these conditions, including limitations of hypnotic medications and melatonin. PMID- 10875044 TI - Diagnosis and management of insomnia. AB - Insomnia is a significant public health issue. Good sleep is essential for emotional and physical wellbeing. The importance of adequate sleep is evidenced by the fact that insomnia can adversely affect physical and mental health. The National Heart, Lung, and Blood Institute Working Group on Insomnia defines insomnia as an experience of inadequate or poor-quality sleep characterized by one or more of the following: difficulty falling asleep, difficulty maintaining sleep, waking up too early in the morning, or unrefreshing sleep. The symptoms of insomnia also include daytime consequences such as tiredness, lack of energy, difficulty concentrating, or irritability. Insomnia can be a symptom of an underlying medical, psychiatric, sleep, or circadian disorder or a disorder in itself (i.e., primary insomnia). This paper will present information about the prevalence, morbidity, causes, and diagnoses of insomnia, and the behavioral and pharmacologic management of this disorder. PMID- 10875045 TI - Diagnosis and management of sleep apnea syndrome. AB - Sleep apnea is the cessation of breathing during sleep. These episodes result in hypoxemia and sleep disruption; thus the consequences are both cardiorespiratory and neural. Sleep apnea syndrome is defined by a constellation of signs and symptoms, with the main presenting symptom being excessive daytime sleepiness. A diagnosis requires documentation of episodes of abnormal breathing during sleep. This disorder, once thought to be very rare, is so common that it is unlikely that any busy clinician has not encountered a case. Facilities for the evaluation of sleep breathing disorders are now available in most communities. With the introduction of continuous positive airway pressure and other treatments, most patients have complete resolution of their disabling symptoms. PMID- 10875046 TI - Diagnosis and management of parasomnias. AB - Parasomnias are unpleasant or undesirable behavioral or experiential phenomena that occur predominately or exclusively during sleep. These phenomena were initially thought to represent a unitary event, often attributed to psychiatric disease. Recent clinical and polygraphic analysis has revealed that they are, in fact, the result of a large number of very different conditions, most of which are diagnosable and treatable. In fact, most are not the manifestation of psychiatric disorders, and they are far more prevalent than previously suspected. Although there are many parasomnias (1,2), from a practical standpoint only the few that comprise the overwhelming majority will be discussed in this review. These include disorders of arousal, rapid-eye-movement sleep behavior disorder, nocturnal seizures, and restless legs syndrome. Most parasomnias are readily diagnosable and, more importantly, are treatable. PMID- 10875047 TI - Tumors of the liver, bile duct, and pancreas. PMID- 10875048 TI - Why take social anxiety disorder seriously? AB - Social anxiety disorder (social phobia) is a disabling psychiatric condition, characterized by a fear of negative evaluation by others. Epidemiological studies have shown a high prevalence of the condition in the general population; the disorder is more common in women than in men. Social anxiety disorder has a typical onset during adolescence and a chronic course; remission rarely occurs without therapeutic intervention. Comorbid psychiatric conditions such as depression and alcoholism commonly occur in patients with preexisting social anxiety disorder, and increase the burden of the condition. Two subtypes of social anxiety disorder have been identified: "nongeneralized" and "generalized"; the latter form causes greater disability and is more often associated with comorbidity. The socioeconomic impact of social anxiety disorder on both sufferers and the community is considerable. For a person with social anxiety disorder, quality of life is greatly reduced; work, social, and personal relationships are all affected. Social anxiety disorder demands increased recognition, so that sufferers receive the treatment they need, in order to improve their quality of life through better social functioning. PMID- 10875049 TI - Social anxiety disorder under scrutiny. AB - Social anxiety disorder (social phobia) is a chronic disabling condition. As with many psychiatric disorders, the condition is likely to have several causes, including genetic and familial factors, early experiences, and cognitive mechanisms. This review will briefly examine the etiology of social anxiety disorder. The approach required during diagnosis of the condition will also be addressed, in particular, the differentiation between social anxiety disorder and other anxiety disorders with similar presentation. The main focus of this article is to review available treatment options for social anxiety disorder, both psychosocial and pharmacotherapeutic. A number of management strategies have shown promise for the treatment of the condition. The International Consensus Group on Depression and Anxiety recently recommended that a selective serotonin reuptake inhibitor (SSRI) should be considered as first-line pharmacotherapy. To date, however, paroxetine is the only SSRI that is licensed for the treatment of social anxiety disorder. PMID- 10875050 TI - Therapeutic advances: paroxetine for the treatment of social anxiety disorder. AB - Data from early studies of selective serotonin reuptake inhibitors have shown that these agents are effective in the treatment of social anxiety disorder (social phobia). This review highlights the outcomes of three large clinical trials of paroxetine in patients with social anxiety disorder. In two of the studies, patients received a flexible dose of paroxetine (20-50 mg/day) or placebo; the third trial was a fixed-dose study, in which patients received paroxetine 20, 40, or 60 mg/day, or placebo. A total of 861 subjects were randomized to treatment for 12 weeks, in centers across the U.S.A., Canada, Europe, and South Africa. The primary outcome measures were the Clinical Global Impressions (CGI) Global Improvement item and Liebowitz Social Anxiety Scale (LSAS) Total Score. In each of the studies, 45-66% of patients receiving paroxetine were rated as responders (very much or much improved on the CGI scale). Paroxetine treatment improved symptoms of social anxiety, as measured by the LSAS, compared with placebo. Differences between paroxetine and placebo groups were statistically significant and were clinically relevant within each study. In general, paroxetine was well tolerated. Paroxetine is effective for the treatment of social anxiety disorder. Based on the findings from these studies, a starting dose of 20 mg/day is recommended. The range of efficacy appears to be 20 50 mg/day for most patients. PMID- 10875051 TI - Longitudinal study of the influence of life events and personality status on diagnostic change in three neurotic disorders. AB - It has been known for many years that diagnosis within the neurotic spectrum of disorders is temporally unstable and also that life events can be major precipitants of change in symptoms. Reasons for this instability could include inherent inadequacy of current diagnostic practice, the influence of life events as an agent of diagnostic shift, and an innate course of disorder with features dependent on the stage at which disorder presents (e.g., development of panic to agoraphobia). These possibilities were examined in a prospective study that was initially a randomised controlled trial. Two hundred ten patients recruited from primary care psychiatric clinics with DSM-III diagnosed dysthymic, generalised anxiety, and panic disorders were randomly allocated to either drug treatment (mainly antidepressants), cognitive-behaviour therapy, or self-help therapy over a 2 year period, irrespective of original diagnosis. Life events were recorded by using a standard procedure over the period 6 months before starting treatment and at five occasions over 2 years; 181 (86%) of the patients had follow-up data and 76% maintained compliance with the original treatment allocated over the 2 years; and 155 of the 181 patients (86%) had at least one diagnostic change in this period. There was no difference in the number of diagnostic changes between the three original diagnostic groups, but dysthymic disorder changed more frequently to major depressive episode than did GAD or panic disorder (20; 11; 12) (%) and panic disorder changed more frequently to agoraphobia (with or without panic) than did dysthymia or GAD (18; 8; 6) (%). There was no relationship between loss events and depressive diagnoses or between addition events and anxiety diagnoses, but greater numbers of conflict events were associated with diagnostic change. More life events were associated with the flamboyant and dependent personality disorders, reinforcing other evidence that many life events are internally generated by personality characteristics and cannot be regarded as truly independent. PMID- 10875052 TI - Measures of depression in older adults with generalized anxiety disorder: a psychometric evaluation. AB - Generalized anxiety disorder (GAD) is the most common of the pervasive anxiety disorders among older adults, with lifetime prevalence estimates of 6%. Because of this high prevalence rate, it is important to establish the utility of assessment tools with this population. Preliminary data exist with regard to the use of anxiety measures with older anxious adults; however, no similar information is available for self-report measures of depression in this population. This study examined the psychometric properties of the Beck Depression Inventory (BDI) and the Geriatric Depression Scale (GDS) with a sample of 54 older adults diagnosed with GAD, 22 of whom were diagnosed with a coexistent depressive disorder. Internal consistency, convergent and divergent validity, construct validity, and discriminative validity of the BDI and the GDS were examined. Overall, the results support the reliability and validity of these self-report measures of depressive symptoms in a sample of older anxious adults. PMID- 10875053 TI - Over-representation of Myers Briggs Type Indicator introversion in social phobia patients. AB - The purpose of this study is to profile the personalities of patients with social phobia. Sixteen patients with social phobia were compared with a normative population of 55,971, and with 24 hospitalized Major Depressive Disorder inpatients, using the Myers Briggs Type Indicator. The Myers Briggs Type Indicator, a popular personality survey, divides individuals into eight categories: Extroverts versus Introverts, Sensors versus Intuitives, Thinkers versus Feelers, and Judgers versus Perceivers. Social phobia patients were significantly more often Introverts (93.7%) than were subjects in the normative population (46.2%). In addition, using continuous scores, the social phobia patients scored as significantly more introverted than did the patients with Major Depressive Disorder, who also scored as Introverted. Introversion is a major component of social phobia, and this observation may have both etiological and therapeutic significance. PMID- 10875054 TI - Effects of nortriptyline and paroxetine on QT variability in patients with panic disorder. AB - This study investigated beat-to-beat QT variability in patients with panic disorder before and after treatment with nortriptyline (n = 13) and paroxetine (n = 16), using an automated algorithm to compute QT intervals. An increase in QT variability appears to be associated with symptomatic patients with dilated cardiomyopathy and also with an increased risk for sudden cardiac death. QTvi (QT variability index: a log ratio of QT variance normalized for mean QT over heart rate variability normalized for mean heart rate) was significantly higher in supine posture in patients with panic disorder treated with nortriptyline (P = 0.006) but not paroxetine. Thus paroxetine may be a better drug of choice especially in patients with coexisting cardiac disease. These findings are important especially in view of the recent reports of increased risk for cardiovascular mortality and sudden death in patients with anxiety and depression. QTvi can be a valuable noninvasive measure of temporal repolarization lability, especially to study the side effects of medications which affect cardiac autonomic function. PMID- 10875055 TI - PTSD in different treatment settings: a preliminary investigation of PTSD symptomatology in substance abuse and chronic pain patients. PMID- 10875056 TI - Anxiety contributes to suicidality in depressed adolescents. AB - Several studies have suggested a positive association between anxiety symptoms and suicidality in adults. However, relatively little is known about this topic in adolescents. To investigate this issue, we examined a group of adolescents admitted to our psychiatric inpatient unit. Fifty-six adolescents (mean age = 14.8 +/- 1.4; females = 34, males = 22; race = 95% Caucasians) participated in the study. Diagnoses were made using the DSM-III-R criteria and a diagnostic interview. Anxiety was found to significantly correlate with depression (r = .60; P = < .05) and suicidality (r = .72; P < .05). A multiple regression analysis revealed that anxiety and depression together accounted for more than half (55%) of the variance in suicidal ideation [F(2,46) = 28.4; P < .0001]. In addition, anxiety had an independent ability to predict suicidality (t = 5.01; P < .0001). Self-rated but not clinician-rated suicidality was positively correlated with both anxiety and depression. Clinical and research implications of these findings are discussed. PMID- 10875057 TI - [Predictive factors of suicide? an 8-year-long prospective longitudinal study of 200 psychiatric inpatients]. AB - Suicide is the most dramatic complication of psychiatric disorders. Certain risk factors are generally accepted by practitioners. Mental disorders increase (tenfold) suicidal risk. However, this "statistically rare event" renders very difficult the definition of predictive factors. A personal prospective longitudinal study of 200 psychiatric inpatients followed up during an 8-year period found 5% of deaths by suicide. Amongst the various risk factors reputed predictive for suicide, only 2 were found statistically more frequent in the suicidal group: familial antecedents (1st degree relatives) of suicide and hospitalization in psychiatry. Impulsivity was also more frequent but could be imputed to the younger age of the suicide victims. Therefore, it was impossible to find determinants of suicide. This makes difficult preventive measures, excepted that psychiatric patients are at a much greater risk and should be diagnosed and correctly treated. There are also increasing legal aspects of responsibility for psychiatrists and psychiatric institutions in charge of these patients. PMID- 10875058 TI - [Actuality of Wallon's emotional model: toward a "body-psychosocial" model of emotions]. AB - Author focuses on qualitative approach of emotions with their human function. She postulates that emotions would be one of the way of the mind's body-inscription. A short presentation of the actual discussion, within psychoneurology and cognitive psychology, shows the lack of a developmental perspective. From the Wallon's emotions theory, the author presents a model that allows to go beyond this limit and to redefine the primary function of emotions: the "body-psycho social Wallon's model" of emotions. Wallon's emotional model focuses on the interaction between body-image and psycho-social construction. This model: a) fixes the emotions into automatisms, but these automatisms are already in link with the social world (through the sense of the mother's language), b) establishes the integrative function of antagonisms (between mind and automatisms, between emotions and mind, between emotions and automatisms). This model shows that emotion's function cannot be reduce to the adaptative response to an unpredictable situation (cognitive or motor). The initial function of emotions concerns the communication system, or better, the first function of emotions is a search for action on family circle, by means of mimicry with ambient and emotional contagion. This emotional system is completely dependent (addicted) on the environment. It gives a "tool" to put in coordination with its environment: a) it favours the setting up of an instant empathy within infant and family circle, b) it makes easier the mind and motor accommodation, c) it put in the necessary plasticity for the emergence of the consciousness. PMID- 10875059 TI - [Magnetic resonance spectroscopy in schizophrenia]. AB - Numerous studies have shown alterations of some structures and/or cerebral functions in patients with schizophrenia. However, the nature of the neurobiological process which could be at the origin of schizophrenic symptoms is still unknown. Magnetic resonance spectroscopy (MRS) is a unique technique which allows us to estimate the concentrations of endogenous substances which contain natural paramagnetic nuclei such as phosphorus (31P) and hydrogen (proton or 1H). The non invasive character of this technique, the absence of side effects, and the possibility of repetitive evaluations allowing for longitudinal studies, make possible MRS studies on the in vivo cerebral metabolism in schizophrenia. The prefrontal cortex, the hippocampus and the basal ganglia have all been implicated in the pathophysiology of schizophrenia. Therefore these brain regions have been frequently studied using MRS. Both proton and phosphorus spectroscopy have been used to study schizophrenia. Compounds that are detectable by 1H-MRS include N acetyl aspartate (NAA), choline (Cho), creatine (Cr) and myo-inositol (ml). A deficit in NAA has been consistently shown in both the frontal and temporal lobes suggesting neuronal loss in these areas. Compounds detectable by 31P-MRS include phosphomonoesters (PMEs) and phosphodiesters (PDEs), which largely represent metabolites generated by lipid turnover. 31P-MRS can also detect certain energy containing phosphorus metabolites such as phosphocreatine (PCr) and nucleotide triphosphates. Decreased levels of PMEs and increased levels of PDEs have been consistently described in the prefrontal lobes suggesting an alteration of phospholipid metabolism. The purpose of this review is to summarize the research on schizophrenia using MRS, to show the utility of this technique in understanding schizophrenia. PMID- 10875060 TI - [Causes and consequences of elderly's agitated and aggressive behavior]. AB - Agitation and aggressiveness are frequent in the elderly and often related to dementia. As a result of the ageing of the general population this is becoming a major public health concern. No or little epidemiological data, during primary health care, about symptoms, co-morbidity, nor medical and social consequences of elderlys' disruptive behavior have been gathered or published in the French literature. Thus, in order to describe these disorders, a survey in cooperation with general practitioners (GP) was conducted. A representative sample of 212 French GP's, all with preferential geriatric activity were asked to conduct a study by including retrospectively their two most recent patients older than 65, who had exhibited agitation and/or aggressiveness. From this cross sectional study, 410 patients (female: 61%, male: 39%) were included. The mean age was 81 years (sd: 7.65). The patients suffered from change in verbal behavior (80%), verbal aggressiveness (71%), physical agitation (60%), wandering (48%), and/or physical aggressiveness (31%). The average of disruptive behavior symptoms per patient was 2.9. The symptoms appeared progressively in 81% of patients, the mean duration was two years and it was the first episode in 40% of patients. Disruptive behaviors may be explained in view of organic illness in 62% of patients (cardiovascular disease: 37%, neurologic: 12%, diabetes: 7%, dehydratation: 5%), dementia (Alzheimer disease: 20%, vascular dementia: 18%, mixed dementia: 14%). In 54% of patients disruptive behavior may be explained in view of depression: 34%, and anxiety disorder: 31%. A triggering factor was observed in 57% of cases (psychosocial stress: 39%). Somatic consequences of the symptoms were frequently identified: decrease of alimentary intake: 39%, weight loss: 27%, dehydratation: 11%, falls: 32%, and irregular medication intake: 31%. Limitation of daily life activities: 85%, and family life: 97% were also noted. Acceptability of patient's symptoms by the family was good (no discomfort or transitory and mild irritability) in 61% of cases, and very bad (reactions of exhaustion, hospitalization requirement) in 13%. This study carried out during primary care, showed that the elderly's disruptive behaviors cause severe medical consequences and familial and social distress. PMID- 10875061 TI - Subjective and autonomic responses to emotion induction in psychopaths. AB - It has been theorized that psychopaths have a fundamental deficit with respect to emotions (Cleckley, 1941). This study compares psychopaths with control subjects (Hare, 1991) in terms of their physiological and subjective responses to video clips depicting different emotions: joy, fear, anger, sadness, and disgust. Physiological measures assessed cardiovascular, respiratory, electrodermal, electromyographic responding, as well as skin temperature. Subjective measures assessed bodily sensations and subjective emotional appraisal. The principal physiological difference was a lower blood pressure among psychopaths before and during emotional stimulation. Thus, while psychopaths's autonomic base-line may be generally hyporeactive, they do not seem to have any emotion-specific physiological deficit. Globally, the results suggested no specific psychophysiological deficits among psychopaths in a context using daily emotions. Subjective measures also revealed that, compared to control subjects, psychopaths reported less intense bodily sensations during emotional stimulation. There was no difference between the two groups in terms of emotional appraisal. PMID- 10875062 TI - [Psychometric properties of French version of the Calgary depression scale for schizophrenics (CDSS)]. AB - The Calgary Depression scale for schizophrenia (CDSS) is a 9 items scale, simple, quick and easy to use. It allows a quantitative approach of the subjective (or cognitive) dimension of the depression, and was developed by Addington et al. (1 5). In this work, we studied the psychometric properties of the CDSS in a population of 95 schizophrenic patients, and 41 non schizophrenic depressed patients. The CDSS was compared with commonly used hetero-questionnaires as the Hamilton Depression Scale (HDRS), the Montgomery-Asberg Depression Scale (MADRS), the Widlocher depressive slowness scale (ERD), and auto-questionnaires as the Beck Depression Inventory (BDI), and the Beck Hopelessness scale (H). In the schizophrenic group, psychotic symptoms were evaluated with the Positive and Negative Symptoms Scale (PANSS), and the extrapyramidal symptoms with the Extrapyramidal symptoms scale (EPRS). In the two populations, the CDSS has similar psychometric properties. The principal component analyses accounts for a unifactorial structure in both groups. In schizophrenics the total score of the CDSS is strongly correlated with the total scores of the HDRS, the MADRS, the ERD, and the G6 item of the PANSS. In non schizophrenic depressed patients, the total score of the CDSS is highly correlated to the total scores of the HDRS, the MADRS and the BDI, with a weaker correlation with the ERD and the H total scores. In these patients, a cut-point strictly superior to 13 may be proposed as a severity criterion for depression in these patients. The internal consistency is satisfactory in both groups, with a Cronbach's alpha of 0.82 in schizophrenics and 0.59 in non schizophrenic depressed subjects. In schizophrenics, items C4 (guilty ideas of reference) and C7 (early awakening) are not necessary to the constitution of the scale. In depressed patients, the deletion of item C6 (morning depression) might increase the internal consistency. Inte-raters agreement is high, with weighted kappas all superior to 0.75 in schizophrenics and to 0.61 in depressed patients. Stability over time is good, and the 72 hours test-retest total score of the CDSS is independent from negative and extrapyramidal symptoms. On the other hand, the positive sub-score and the positive factor of the PANSS are correlated to the CDSS total score. The validation of the CDSS is still not complete: sensitivity to change and stability of the factorial structure remain to be explored. Nevertheless, the CDSS is an interesting tool for a quantitative approach of the subjective dimension of depression in both schizophrenic and non schizophrenic patients. PMID- 10875063 TI - [Extrapyramidal side effects of neuroleptic and antidepressant treatment: assessment of potential risk factors through CYP2D6 genetic polymorphism]. AB - The objective of this study was to assign metabolizer phenotype (cytochrome P450 2D6 or CYP2D6) to drug treated psychiatric adult patients to assess if the CYP2D6 polymorphism could be a potential risk factor for the development of extrapyramidal side effects of psychotropic drugs. Twenty-eight unrelated in patients (16 men and 12 women) treated with antidepressants and/or antipsychotic drug were phenotyped using dextromethorphan. Two groups of patients were considered depending on the presence (n = 14) or not (n = 14) of extrapyramidal side effects. The mean dextromethorphan/dextrorphan metabolic ratio (log10) did not differ between the two groups of patients (-1.13 +/- 0.9 and -1.56 +/- 0.5, NS). But significantly more patients with extrapyramidal side effects (n = 4) than patients without side effects (n = 0) were poor metabolizers. This result could be due to a quantitative difference between the 2 groups of drug treatment cosegregated with dextromethorphan, but several authors reported that extrapyramidal side effects seemed not to be always related to high plasma drug levels. So the authors concluded that the 2D6 polymorphism could be a risk factor of poor neurologic tolerance of psychotropic drugs, but not only through pharmacokinetic consequences. CYP 2D6 is indeed expressed in brain and seems to interfer with the metabolism of dopamine and other related neurotransmitters. PMID- 10875064 TI - [Addiction and personality]. AB - Within the field of substance abuse, it is now widely admitted that the addictive personality does not exist. No one personality type is predisposed to addiction. The predisposition to drug dependence involves many different factors: psychological, social, familial, biological. None of these factors can be the sole determinant of drug dependence. Keeping that in mind, it is of interest to review the recent data on the relationship between personality traits or disorders and opiate and cocaine dependence. Using DSM and ICD categorical assessment, no single personality disorder emerged, instead a range of personality disorders has been evaluated in opiate and cocaine dependent subjects. Every type of personality disorders (PD) existed but cluster BPD were the most common (especially antisocial personality disorder in opiate addicts). However, it is noteworthy that a large minority to a majority of subjects did not display any king of PD. The implication of these results is that antisocial PD is probably over-diagnosed in drug dependence clinical settings. The studies reviewed failed to demonstrate that personality disorders were strong predictors of outcome in opiate or cocaine dependence. However, opiate dependent PD subjects entering treatment had more severe problems and lower retention rate than non PD subjects. But the amount of improvement was not significantly different between PD subjects and non PD subjects. This demonstrated that substance dependent PD patients could benefit from treatment whose intensity and duration must be adjusted. There is good support for the idea that Sensation Seeking trait is a vulnerability factor to substance abuse. But after dependence develops, sensation seeking is probably irrelevant to continued use of the drugs. This break between the psychopathology of vulnerability of substance abuse and the psychopathology of dependence raises the question of the existence of dramatically different factors involved in both phases of addiction. PMID- 10875065 TI - [Current status of the interdisciplinary model project "Neurodermatitis Education for Children and Adolescents"]. AB - With the help of the German Ministry of Health and the health insurance companies a national interdisciplinary project for children and adolescents with atopic eczema will be developed. The aim of the project is to generate standardized teaching concepts for parents and children covering medical, psychological and social aspect of the disease. 800 children and adolescents of different age groups will be enrolled in this project. Furthermore, during the three years of the study the efficacy of the teaching concept will be evaluated. PMID- 10875066 TI - [Urticaria. New developments and perspectives]. AB - Although the etiology of urticaria is mostly unclear, there are a number of recent new insights into the underlying pathomechanisms and causes. Dermal mast cell numbers and endothelial cell adhesion molecule and cytokine expression are also increased in uninvolved skin, while serum P-selectin levels are elevated, suggesting a systemic activation of the cutaneous inflammatory system in urticaria. In all adults, but not children with indolent mastocytosis, we found activating point mutations of c-kit, the mast cell growth factor receptor, in lesional cutaneous mast cells. In acute urticaria, IgE-dependent mechanisms are only rarely involved (0.9%), in contrast to generally held beliefs, whereas acute upper respiratory infections (39.3%) and drug intolerance (9.2%) are more frequent. In chronic urticaria, pseudoallergies to food (73%) and more rarely chronic inflammatory gastrointestinal diseases (11%) play a major role, with avoidance or elimination of the eliciting factors leading to long term remission. On the basis of recent findings, autoantibodies must be viewed mostly as secondary rather than causative in chronic urticaria. PMID- 10875067 TI - [Pemphigus. Loss of desmosomal cell-cell contact]. AB - Pemphigus diseases comprise a group of autoimmune disorders which are characterized by intraepidermal blisters and autoantibodies to components of desmosomes. Desmosomes mediate adhesion between neighbouring keratinocytes. A common feature of pemphigus diseases are intercellular deposits of IgG or, less frequently, of IgA within the epidermis. The group of pemphigus diseases includes pemphigus vulgaris, pemphigus foliaceus, pemphigus vegetans, pemphigus herpetiformis, pemphigus erythematosus, paraneoplastic pemphigus, drug-induced pemphigus, and IgA pemphigus. Using molecular tools, some of the autoantigens in these diseases have been characterized. In pemphigus vulgaris, autoantibodies are directed to desmoglein 3 and in pemphigus foliaceus to desmoglein 1. Target antigens in IgA pemphigus are desmocollin 1 and desmoglein 3. In paraneoplastic pemphigus, autoantibodies react with a complex of various proteins, including desmoplakin 1 and 2, BP230, envoplakin, periplakin, plectin, desmoglein 3, and a yet uncharacterized 170 kD protein. This review summarizes new insights into the immunopathogenesis and diagnosis of pemphigus diseases. PMID- 10875068 TI - [Prospective detection of important bacterial pathogens in pyoderma and their in vitro antibiotic susceptibility]. AB - BACKGROUND AND OBJECTIVE: For rational therapeutic recommendations the spectrum and resistance of bacteria in skin diseases were investigated. PATIENTS/METHODS: Within 4 months 911 swabs of dematoses possibly caused by bacteria were taken prospectively (481 outpatients, 430 inpatients) and the material cultured on standard media. The positive cultures including resistance screening of 210 swabs of 168 outpatients and of 175 swabs of 85 inpatients could be evaluated, the remaining cultures were sterile. RESULTS: Staphylococcus aureus was the most frequent pathogen (outpatient 67%, inpatient 61% of all positive cultures), followed by streptococci (groups A and B; 25%). In patients Pseudomonas aeruginosa was the most common pathogen in leg ulcers and between the toes (45% and 70% respectively). 13% of the Staphylococcus aureus isolates were resistant to tetracyclines and erythromycin; one strain proved to be methicillin resistant. CONCLUSIONS: Cephalosporins (I. generation), penicillins with beta-lactamase inhibitors, and to a lesser extent isoxazoyl-penicillin and clindamycin can be recommended for the treatment of skin infections. Oral quinolones are suited for infections with gram-negative bacteria (such as bewteen the toes). The indications for systemic antibiotic therapy of leg ulcers should be restricted. PMID- 10875069 TI - [Symplastic hemangioma]. AB - BACKGROUND AND OBJECTIVE: Endothelial nuclear atypia is the hallmark of malignant vascular tumors. Pleomorphic nuclei of the muscular vessel wall and the adventitia are manifestations of degenerative phenomena and should not be misinterpreted as signs of malignancy. PATIENTS/METHODS: Three long-standing solitary superficial vascular tumors (61-year-old woman, 48- and 63-year-old men) were removed by primary excision. Sections were stained according to standard histologic and immunohistologic protocols. RESULTS: Symplastic hemangiomas show the silhouette of a small superficial angioleiomyoma or capillary hemangioma. Characteristic features are multinucleate cells of the muscular vessel wall and the adventitia with pleomorphic nuclei, broad hyalinized vessel walls, and the distinctive lack of endothelial nuclear atypia. Recurrences or metastases were not reported (follow-up of 9, 45 and 90 months). CONCLUSIONS: Symplastic hemangioma is a benign superficial hemangioma with histological features of a pseudomalignancy. The distinctive lack of endothelial nuclear atypia allows distinction from malignant vascular tumors. PMID- 10875070 TI - [Acral necroses after therapy with quinine sulfate for calf cramps]. AB - The most common cause of acral cyanosis is vascular spasm which can be induced by several drugs. An 87-year-old woman developed red and livid skin lesions on the fingers of both hands and several toes one month after beginning treatment with quinine sulfate 200 mg daily. The skin lesions progressed to necrosis in some areas. Quinine sulfate is a widely prescribed drug for nocturnal cramps. The following side effects may develop, particularly in the elderly: exanthems, pruritus, urticaria, erythema multiforme, purpura and photosensitivity. Our case points to the possibility of acral necrosis and demonstrates the efficacy of vasodilator treatment. PMID- 10875071 TI - [Successful long-term therapy of Stewart-Bluefarb syndrome]. AB - The Stewart-Bluefarb syndrome is defined as an unilateral angiodermatitis due to multiple arterio-venous fistules accompanied by acroangiodermatitis resembling Kaposi sarcoma (pseudo-kaposi sarcoma). The acroangiodermatitis is most common on the lower limb. It leads to ulcerated nodules with a high risk of bleeding and infection, as well as edema, pain and seldom limb hypertrophy. Curative therapy requires elimination of the arteriovenous shunts. Surgical destruction of the multiple small fistulae is a limitating factor. A better alternative is embolisation, but this approach carries the risk of ischemia and necrosis. A 32 year old female patient with Stewart-Bluefarb syndrome is presented; she has been successfully treated with embolisation on eight occasions. PMID- 10875072 TI - [Subepidermal bullous autoimmune dermatosis with linear deposition of IgA amd IgG]. AB - A 29-year-old female patient with an autoimmune subepidermal blistering disease had linear deposits of both IgA and IgG at the basement membrane zone. Clinically, the patient presented with tense blisters on the face, trunk, extremities and oral mucosa. Histologically, we found a subepidermal blister formation and a predominantly neutrophilic infiltrate. Direct immunofluorescence showed linear deposits of IgA along the basement membrane zone, as well as linear deposits of IgG and C3 as typically found in bullous pemphigoid. Indirect immunofluorescence demonstrated circulating IgA and IgG autoantibodies. This case extends previous reports on a subgroup of patients with subepidermal blistering diseases characterized by the presence of both IgA and IgG anti-basement membrane antibodies. These patients reveal clinical, histological and immunopathological features of linear IgA disease and bullous pemphigoid. PMID- 10875073 TI - [Juxta-articular fibroid nodules and acrodermatitis chronica atrophicans in late stage Lyme borreliosis]. AB - A 60-years old female patient developed juxta-articular fibroid nodules and erythrocyanotic lesions of acrodermatitis chronica atrophicans after several tick bites. The woman was treated with ceftriaxon (Rocephin) 2 g daily parenterally without adverse reactions. PMID- 10875075 TI - [Determination of the azoospermia factor on the Y chromosome]. PMID- 10875074 TI - [Sporotrichoid infection with Mycobacterium marinum: successful therapy with oral tetracycline administration]. AB - Atypical mycobacterial infections of the skin present a diagnostic and therapeutic challenge to dermatologists in many instances. We report on a patient who was diagnosed with atypical mycobacteriosis in its rarer, sporotrichoid form. Possible differential diagnoses are discussed. Efficient therapy using minocycline is demonstrated. PMID- 10875076 TI - [Sarajevo, 1903]. AB - The current conflicts on the Balkans are used as an opportunity to call to mind the VIII. conference of the DDG in Sarajevo in 1903. For the first time, it then took place in a city which did not have a university and therefore no university clinic. Possible political but also dermato-venereological aspects concerning the choice of conference venue are discussed. Furthermore, highlights of the situation of the DDG in those days are presented and some central points regarding the professional topics of the conference and presentation are considered. PMID- 10875077 TI - [CO2 laser vaporization in cheilitis actinica. Current comment on the contribution by S. Hohenleutner, M. Landthaler and U. Hohenleutner]. PMID- 10875079 TI - [In Process Citation] PMID- 10875078 TI - [Current position of the Professional Group of Dermatologic Oncology on high dosage therapy with interferon-alpha-2b]. PMID- 10875080 TI - [Cutaneous B-cell lymphoma]. PMID- 10875081 TI - Costenbader Lecture. Idiopathic infantile nystagmus: diagnosis and treatment. AB - Current concepts of idiopathic infantile nystagmus are summarized, with special attention to treatment and differential diagnosis of this condition. Advantages of the Anderson procedure over the Kestenbaum procedure are suggested for head turn associated with this condition, and the need for further studies is acknowledged. The importance of the extended slow phase in understanding the waveforms of infantile nystagmus is stressed. Our studies reinforce what I believe to be the natural history of infantile nystagmus, as well as the history of periodic alternating nystagmus. The critical delineation of diagnosing periodic alternating nystagmus is emphasized with respect to the type of operation to avoid overcorrection of head turns in patients with nystagmus. Continued searches for manifest latent nystagmus are important, because that condition is currently the only truly treatable nystagmus. I thank the American Association for Pediatric Ophthalmology and Strabismus for the privilege and honor of presenting the 1997 Costenbader Lecture. PMID- 10875082 TI - Results of anterior transposition of the inferior oblique muscle in incomitant dissociated vertical deviation. AB - PURPOSE: Recession with anterior transposition of the inferior oblique muscle has been shown to effectively decrease dissociated vertical deviation in primary position. However, studies to date have not addressed the long-term postoperative results with respect to residual deviation in lateral gaze, development of A pattern strabismus, and the effect of the procedure on upgaze. METHODS: Twenty three eyes in 12 patients were treated with recession with anterior transposition of the inferior oblique muscle for dissociated vertical deviation greater in adduction than in abduction (termed incomitant dissociated vertical deviation) associated with inferior oblique muscle overaction. Before the operation, dissociated vertical deviation was measured in primary position and lateral gaze, oblique muscle dysfunction was graded, and A or V patterns were measured. Similar measurements were made after the operation. All patients have been followed up for a minimum of 4 years after the operation. RESULTS: Recession with anterior transposition of the inferior oblique muscle effectively eliminated the dissociated vertical deviation in primary position and in adduction. The operation was less effective in reducing small amounts of dissociated vertical deviation in abduction. No significant A patterns developed after the operation. Postoperative inferior oblique muscle function ranged from -1 underaction to +2 overaction, and postoperative upgaze in abduction was normal to mildly deficient. CONCLUSIONS: Recession with anterior transposition of the inferior oblique muscle results in long-term improvement of incomitant dissociated vertical deviation, with a low incidence of late development of A patterns and upgaze deficiency. PMID- 10875083 TI - Initial versus subsequent postoperative motor alignment in intermittent exotropia. AB - BACKGROUND: Although initial overcorrection is believed to be important after bilateral lateral rectus muscle recessions for intermittent exotropia, not all patients with desirable amounts of initial overcorrection have good final outcomes. The purpose of this study is to evaluate the relationship between initial postoperative and subsequent postoperative motor outcomes in a group of patients operated on for intermittent exotropia. METHODS: All patients on whom I performed bilateral lateral rectus muscle recessions as the initial surgical procedure for intermittent exotropia and who had at least 6 months of postoperative follow-up were included in this study. RESULTS: Of the 60 patients in this study, 38 (63%) had good outcomes (< or = 10 PD exophoria or < or = 5 PD esophoria), 15 (25%) had undercorrection (> 10 PD exodeviation), and seven (12%) had overcorrection (> 5 PD esodeviation). The chance of a good outcome was highest with initial postoperative alignment between orthotropia and 9 PD of esotropia, but 22% of patients with alignment in this range after the operation ended up overcorrected or undercorrected. Most patients had an exotropic drift after the operation, but seven patients had a drift in an esotropic direction. CONCLUSIONS: Although an initial alignment within the range of orthotropia to 9 PD of esotropia during the first few days after the operation is desirable for patients with intermittent exotropia, alignment within this range does not guarantee a good final outcome, nor does alignment outside this range guarantee a bad outcome. Little predictability exists with respect to the amount and occasionally even the direction of postoperative drift. This unpredictability may in part reflect the artifactual nature of the initial postoperative measurement. PMID- 10875084 TI - Intraoperative sponge 5-fluorouracil to reduce postoperative scarring in strabismus surgery. AB - INTRODUCTION: To determine whether 5-fluorouracil is effective in reducing scarring after strabismus surgery we used rectus muscle surgery in experimental animals to compare a single intraoperative dose of 5-fluorouracil with mitomycin C and to compare results in similarly treated controls not receiving these antimetabolites. METHODS: Muscle resections were performed on eight rabbits (16 eyes). Four eyes had 5-fluorouracil (50 mg/ml), and four eyes received mitomycin C (0.2 mg/ml), each of which was applied during surgery on an ophthalmic sponge for 5 minutes. Eight eyes served as controls. Six weeks after surgery conjunctival vascularity, muscle length-tension curves, muscle disinsertion force, and the histologic degree of scarring were assessed. RESULTS: The mitomycin C-treated eyes clearly had more conjunctival avascularity and a lower disinsertion force. Both treated groups had flatter length-tension curves and less scarring on histologic examination than the control eyes. CONCLUSIONS: Antifibroproliferative therapy with intraoperative sponge 5-fluorouracil appears as effective as, and is possibly safer than, mitomycin C. It may be a useful adjunct in recurrent strabismus surgery or in other situations where a risk of excessive postoperative scarring exists. PMID- 10875086 TI - Corneal edema after pediatric cataract surgery. AB - INTRODUCTION: We have encountered idiopathic corneal edema in four patients (five eyes) after pediatric lensectomy. This problem has not been previously described in the pediatric ophthalmology literature. METHODS: Clinical and operative records were reviewed. The children, who ranged in age from 15 months to 6 years, underwent apparently uncomplicated limbal lensectomy without lens implantation. After surgery, all received subconjunctival hydrocortisone (12.5 mg) and 2 to 4 drops daily of topical prednisolone acetate. The corneal edema developed between 2 and 14 days after surgery. RESULTS: The condition cleared in all patients during a 5- to 14-day course of intensive topical steroids. No sequelae have been apparent. Final visual acuities are 20/30 or better in the three children (four eyes) old enough for recognition acuity testing. The fifth eye has excellent central fixation. CONCLUSIONS: We suspect that the corneal decompensation was a manifestation of sterile inflammation. Two of the children had a history of iritis. Difficulty measuring cellular response at the slit-lamp examination and instilling eyedrops at home may have contributed to the complication. Postoperative corneal decompensation can be responsive to topical steroids, which we now prescribe more intensively even in apparently quiet eyes. PMID- 10875085 TI - Glaucoma and ocular hypertension in pediatric patients with cataracts. AB - BACKGROUND: The pathogenesis of open-angle glaucoma and ocular hypertension in patients who have undergone surgical correction of their congenital cataracts remains undetermined. This study examines the prevalence of glaucoma and ocular hypertension in a population of patients who did not undergo surgical correction of their pediatric cataracts. METHODS: Fifty-eight eyes of 41 patients had cataracts before 2.5 years of age and were followed up until at least 5 years of age without operative correction. The patients were studied for the following parameters: age at diagnosis, type of cataract, etiology, bilaterality, optic nerve head cup-to-disc ratio, intraocular pressures, and reason why the patient did not undergo an operation. Glaucoma was defined as the presence of glaucomatous optic nerve head cupping with intraocular pressures of greater than 22 mm Hg. Ocular hypertension was defined as intraocular pressures greater than 22 mm Hg with no optic nerve changes. RESULTS: Nine of the 58 eyes had cataracts caused by persistent hyperplastic primary vitreous. The average age to the last intraocular pressure measurement was 19 years (range 5 to 48 years). Closed-angle glaucoma developed in two patients with persistent hyperplastic primary vitreous. Neither open-angle glaucoma nor ocular hypertension developed in any patients. CONCLUSION: Pediatric cataracts not of the persistent hyperplastic primary vitreous type were not associated with ocular hypertension or glaucoma in the absence of surgical cataract correction. In eyes with persistent hyperplastic primary vitreous cataracts, spontaneous closed-angle glaucoma developed in two of nine patients and open-angle glaucoma developed in none. Surgical cataract correction, or the aphakic state that follows such operations, may be responsible for pediatric aphakic glaucoma. PMID- 10875087 TI - Histopathology and vascular endothelial growth factor in untreated and diode laser-treated retinopathy of prematurity. AB - OBJECTIVES: We had the unique opportunity to compare the eyes of a premature infant with stage 3 retinopathy of prematurity (ROP) in both eyes after the condition was treated by diode laser photocoagulation in one eye only. After the infant's death, we investigated the extent of structural damage incurred with the diode laser and examined the effect of treatment on vascular endothelial growth factor (VEGF) expression. METHODS: The eyes were fixed and embedded in paraffin. Adjacent 6 microns sections were either stained for histopathologic analysis or used for in situ hybridization. VEGF messenger RNA (mRNA) was detected by using radiolabeled antisense riboprobes. RESULTS: In the treated eye, histopathologic results demonstrated the clinically evident dose-response effect, with sparing of inner retinal elements with mild laser burns and full-thickness retinal cell disruption with severe burns. Scleral and ciliary nerve effects were absent. VEGF mRNA was localized primarily in the ganglion cell layer but was also found in the inner nuclear layer. In the untreated eye, an increase in VEGF mRNA was detected at the peripheral edge of the vascularized retina anterior to the ridge. In the laser-treated eye, VEGF mRNA expression was dramatically upregulated in the ganglion cell layer in areas adjacent to laser burns. CONCLUSIONS: VEGF mRNA was found to be elevated in the peripheral, avascular retina of the untreated eye, consistent with the hypothesis that retinal hypoxia stimulates VEGF expression. In the treated eye with recurrent ROP, VEGF mRNA was not detected in the photocoagulated areas of retina but was increased between laser scars. This finding confirms the results of prior animal studies and validates the use of these models. PMID- 10875088 TI - Asymptomatic uveitis in young people with inflammatory bowel disease. AB - PURPOSE: To ascertain the prevalence of uveitis in a population of pediatric patients with inflammatory bowel disease without ocular symptoms. METHODS: We prospectively evaluated all young people who came to the pediatric gastroenterology clinic with endoscopically proven inflammatory bowel disease between March 1994 and June 1995. All the patients were examined for evidence of ocular manifestations of inflammatory bowel disease. The examination consisted of slit-lamp examination, tonometry, and indirect ophthalmoscopy. None of the patients had visual or ocular symptoms. Eighteen patients had Crohn's disease and 14 had ulcerative colitis. RESULTS: Of the 32 patients evaluated, four (12.5%) had evidence of asymptomatic ocular inflammation, defined as anterior chamber cell and flare. All patients with ocular inflammation were male. Three of these four male patients had Crohn's disease; the other had ulcerative colitis. Five patients had posterior subcapsular cataract, one had esotropia and amblyopia, and one had unilateral high myopia. CONCLUSIONS: The prevalence of asymptomatic uveitis in our population of young people with inflammatory bowel disease was 12.5%. These findings suggest the need for a screening ophthalmologic examination to rule out occult eye disease in young people with inflammatory bowel disease. PMID- 10875089 TI - Mitochondrial diseases in pediatric ophthalmology. PMID- 10875090 TI - Ocular involvement in hand, foot, and mouth disease. PMID- 10875091 TI - Extraocular muscle involvement in sarcoidosis: a clinicopathologic report. PMID- 10875092 TI - Fetal venous circulation--an update on hemodynamics. AB - The refinements of modern ultrasound techniques permit a renewed examination of old concepts of fetal circulation. The concept of preferential streaming of umbilical blood through the foramen ovale is verified by animal experiments, and ultrasound studies have confirmed that a similar mechanism operates in human fetuses. However, the normalized umbilical flow appears to be less in the human than in fetal sheep, and decreases with advancing gestational age (115 ml min-1 kg-1 at 20 and 64 mL min-1 kg-1 at 40 weeks). Compared to the 50% shunting of umbilical blood through the ductus venosus found in animal experiments, the degree of shunting in the human fetus under physiological conditions is considerably less, 30% at 20 weeks, which decreases to 18% at 32 weeks, suggesting a higher priority of the fetal liver than previously realized. Augmented pulsatility in the precordial veins, ductus venosus, and umbilical vein is an important clinical sign that is poorly understood. Recent fluid dynamic studies show that, apart from the pressure generated in the atria, it is the stiffness of the vessel wall, compliance, and notably, impedance which modify these waves. Particularly the substantial shift in impedance at the ductus venosus-umbilical vein junction causes wave reflection and reduced transmission of waves, the result being diminished or absent pulsation in the umbilical vein. PMID- 10875093 TI - Soluble intercellular adhesion molecule and C-reactive protein as early markers of infection in newborns. AB - In order to find a reliable early marker of infection in newborns a study with simultaneous determination of soluble Intercellular Adhesion Molecule-1 (sICAM-1) and C-Reactive Protein (CRP) was planned. Prospectively 90 babies < 5 days of age suspect of infection were included. Retrospectively this population was classified into an "infected" group (n = 45) and a "non-infected" group (n = 45). For each of these two groups we calculated the sensitivity, specificity and predictive values of sICAM-1 and CRP as early markers of infection. We determined the best cut-off level for sICAM-1 to be 300 micrograms/l and for CRP 5 mg/l. As a biochemical test for infection in the newborns the sensitivity and negative predictive value for CRP were 0.69 and 0.73 respectively. When sICAM-1 was added and CRP and s-ICAM-1 were used in combination the sensitivity improved significantly to 0.93, p < 0.01 and the negative predictive value improved to 0.92, p < 0.05. In normal 5-8 days old babies' sICAM-1 was significantly higher than at birth (cord blood), p < 0.0001. In conclusion, sICAM-1 and CRP in combination are better than CRP as a primary test for identification of infection in babies < 5 days of age. PMID- 10875094 TI - Risk factors for intraventricular hemorrhage in a birth cohort of 3721 premature infants. AB - AIMS: In our study we determined possible risk factors for intraventricular hemorrhage grade III to IV (IVH) based on a regional German neonatal data base and tried to build a logistic-regression model to predict the risk of IVH according to gestational age. MATERIALS: We identified 3721 premature infants, 22 to 36 completed weeks of gestational age, born from 1994 through 1997. 136 (3.7%) IVH were diagnosed sonographically. 60 (44%) infants with IVH died. We examined the following variables as risk factors for IVH: gestational age, sex, blood pH of 7.2 or less, body temperature of 35 degrees C or less, multiple birth, small for-gestational age, intubation after birth, transport to another hospital. RESULTS: In the full logistic regression model sex, blood pH of 7.2 or less, multiple birth, and small-for-gestational age were not associated with a significant risk of IVH. Body temperature of 35 degrees C or less was associated with an increased risk of IVH (adjusted odds ratio, 1.92; 95% confidence interval, 1.09 to 3.40). Intubation after birth increased the risk of IVH in neonates under 28 weeks of gestational age (OR, 3.72; 95% CI, 1.65 to 8.38) only to a moderate extent, but significantly in neonates 32 to 36 weeks of gestational age (OR, 16.51; 95% CI: 7.35 to 36.18). The risk of IVH was mainly related to gestational age. Neonates delivered before 28 weeks of gestation (OR, 75.72; 95% CI, 46.14 to 124.30) faced the highest risk of IVH. Transport to another hospital was connected with an increased risk of IVH regardless of gestational age (adjusted OR, 1.95; 95% CI, 1.07 to 2.56). CONCLUSION: The frequency of IVH could be reduced significantly, if extremely premature infants, the vast majority of patients suffering from IVH, did not have to be transferred postnatally to another hospital. PMID- 10875095 TI - Choice of cesarean section and perception of legal pressure. AB - OBJECTIVE: To evaluate the perception of "Defensive Medicine" by hospital based obstetricians and the influence of this attitude on the choice of cesarean delivery. SUBJECTS AND METHODS: Questionnaire sent by mail to a sample (76) of obstetricians of general district, teaching and university hospitals in a region of southern Italy (Puglia). Doctors were selected as the head, the most senior and the most junior specialist of each department. Independent variables of the study were considered as demographic data of the subjects, years of service, interest in private practice, size of the hospital, background cesarean section rate, personal and site of work exposure to legal claims. Outcome measures were experience and confidence in training for operative vaginal and breech delivery, use of the partogram in labor, opinion about a trial of labor after a previous cesarean section and about cesarean section on request, personal perception of defensive medicine. Univariate and multivariate analysis of data were performed. RESULTS: The response rate was 83%. According to our data, seniority in service meant confidence in and request of more teaching of obstetrics manoeuvres, size of hospitals was positively related to a more rationale approach of the diagnosis of dystocia, heads of units were keener to accept the patient's wish for a cesarean section. Doctors with large private practices were less likely to be sued and the perception of legal pressure was directly related to the rate of cesarean section in each unit. DISCUSSION: Defensive Medicine is a reality that encompasses all categories of doctors in this survey. The only differences were in the rate of perception of legal pressure. We believe that residential programs should be modified in order to improve specialists' understanding of malpractice problems and that the patient-doctor relationship should be ameliorated in public hospitals. PMID- 10875096 TI - Detection of C-type natriuretic peptide in fetal circulation. AB - C-type natriuretic peptide (CNP) belongs to the natriuretic peptide family that plays an important role in the control of blood pressure, renal function and volume homeostasis. In contrast to the atrial natriuretic peptide and brain natriuretic peptide, CNP acts in an autocrine/paracrine fashion and is considered to be the endothelial component of the natriuretic peptide system. CNP has a high expression and tissue-specific regulation in reproductive organs. Using a radio immunoassy for CNP-22 we measured for the first time CNP in fetal blood. Samples were taken by cordocentesis in a group of fetuses with rhesus isoimmunisation (10.74 +/- 2.81 pg/ml), fetuses with rhesus isoimmunisation after intravascular transfusion (10.03 +/- 4.01 pg/ml) and a group with structural anomalies (12.9 +/ 5.67 pg/ml). A group of healthy fetuses was used as controls (11.64 +/- 4.32 pg/ml). In contrast to ANP, the fetal CNP-plasma concentrations remain stable in the investigated fetal diseases and after volume load during intravascular transfusion. Moreover, fetal CNP-plasma levels are higher than previously measured maternal concentrations in normal pregnancies. Therefore, the fetus expresses CNP independently of the maternal circulation. PMID- 10875097 TI - Evaluation of a strategy to limit blood donor exposure in high risk premature newborns based on clinical estimation of transfusion need. AB - OBJECTIVES: Reservation of dedicated series of pedipacks, consisting of 3 to 4 units of 70 ml filtered red cell concentrate in additive solution SAGM from 1 donor, may reduce donor exposure. In this prospective efficacy study the benefits, release and expiration of pedipacks (PP) assigned to preterm infants requiring neonatal intensive care are analyzed. METHODS: On the basis of clinical assessment of the need for multiple transfusions, 96 preterm neonates (gestational age < 32 wks and/or birth weight < 1500 g) were assigned to either the high risk group (HRG), who were to receive dedicated donor blood units, or the low risk group (LRG). Inclusion criteria for HRG were 1) estimated time of admission > 21 days and 2) expected need for multiple transfusions due to clinical cardiorespiratory instability, prolonged parental feeding or frequent blood sampling. To reduce wastage of donor blood, dedication of donor blood units was limited to 21 days. RESULTS: 50 series (192 PP) were assigned to 42 HRG infants. Two HRG infants received 3 series, 4 received 2 series and 36 received 1 series of PP. Mean transfusion rate was 3.1 PP in the HRG and 0.4 in the LRG. In the LRG 35 of 54 were not transfused, 19 received 1 to 2 PP. In both groups transfused newborns were exposed to 1.1 donors in average. In the HRG of 192 PP, 137 PP (71%) were used within 21 days, and another 30 (16%) before the expiration date < 35 days. Twenty five PP (13%) expired, mainly because of logistical problems in the introduction phase. CONCLUSION: Assignment of dedicated PP on the basis of clinical parameters at entry considerably reduces donor exposure in HRG. Wastage of dedicated blood transfusions was reduced by limitation of the dedicated period (21 days). In terms of efficacy, reservation and use of PP can be optimized by standardized administrative measures. PMID- 10875098 TI - The significance of interleukin-6 concentrations in cervicovaginal fluid: its relation to umbilical cord plasma and the influence of antibiotic treatment. AB - This study was performed to correlate cervicovaginal fluid and umbilical cord plasma level of IL-6 and IL-8 in patients with premature rupture of the membranes (PROM) and to see the effect of antibiotics on those concentrations. As a part of a randomized controlled trial of treatment in PROM with antibiotics, cervicovaginal fluid was sampled before delivery for measurement of IL-6 and IL-8 and for bacteria from 36 patients less than 36 weeks of gestation. Umbilical cord plasma was also collected. Concentrations of IL-6 and IL-8 were measured by an ELISA. Neonatal infections were noted in a total of 9 cases, including bacteria detection (Escherichia coli 2 cases, GBS and Streptococcus constellata) in 4 cases. Correlation between IL-6 in cervicovaginal fluid and in cord plasma (r = 0.881, p < 0.0001) was stronger than that of IL-8 (r = 0.469, p < 0.01). The difference of concentrations in IL-6 and IL-8 was not significant between cases with (n = 20) and without (n = 16) ampicillin. Our observation indicates that the measurement of IL-6 concentrations in cervicovaginal fluid is a useful marker for PROM patients who are more likely to develop neonatal infection and the antibiotic treatment does not necessarily produce their beneficial effects on fetuses at the risk of infection. PMID- 10875099 TI - Maternal serum interleukin 6 levels in preterm labor: prediction of admission-to delivery interval. AB - OBJECTIVE: To evaluate the diagnostic value of maternal serum interleukin 6 (IL 6), C-reactive protein (CRP) levels and white blood cell (WBC) count for the prediction of preterm labor and length of admission-to-delivery interval in patients with preterm labor. METHODS: Maternal serum IL-6, CRP and WBC count were prospectively determined in eighty-two patients in preterm labor and 21 controls. Data was analyzed in study and control groups, and for the assessment of clinical and laboratory risk factors in the prediction of admission-to-delivery interval in the study group. RESULTS: Maternal serum IL-6 levels were significantly higher in the study group than controls. The IL-6 value associated with the highest percent of true positives and true negatives for the prediction of preterm labor was 5 pg/ml. The area under curve of maternal IL-6 was significantly higher than the area under curve of of CRP and WBC count. In the study group maternal serum IL-6 levels were significantly higher in patients delivered within 2 and 7 days than the nondelivering ones and a cut off value of 8.3 pg/ml was determined for estimation of preterm delivery. CONCLUSION: Maternal serum IL-6 is a reliable marker in the prediction and management of preterm labor and delivery. PMID- 10875100 TI - Extracellular matrix components of the wall of umbilical cord vein and their alterations in pre-eclampsia. AB - Pre-eclampsia--edema, proteinuria, hypertension (EPH-gestosis) is one of the more common complications observed during pregnancy. The umbilical cord vein walls were taken from newborns delivered by healthy mothers (control material) and by mothers with polysymptomatic pre-eclampsia (investigated material). Normal saphenous vein walls were collected from adult subjects undergoing varicose vein surgery. The collagen content was measured by the assay of hydroxyproline. Elastin was determined according to Fastin Elastin Assay and gravimetrically. Glycosaminoglycans content was determined by uronic acids assay. The collagen content decreased in the pre-eclampsia material. The amount of soluble elastin increased in the investigated material. The insoluble elastin content decreased in the umbilical cord veins of newborns delivered by mothers with pre-eclampsia. Reconstructing the umbilical cord vein wall may disturb fetal blood flow and affect the vascular system in adulthood. PMID- 10875101 TI - Legionnaire's disease complicating pregnancy: a case report with intrauterine fetal demise. AB - OBJECTIVE: Legionnaire's disease complicating pregnancy is an unusual event that can seriously compromise both the mother and the fetus. CASE REPORT: We describe one case of such association, with an unfavourable intrauterine fetal outcome, secondary to acute placental insufficiency, related to infection. DISCUSSION: It is important in these high risk pregnancies complicated by acute pneumonia to take into consideration the diagnosis, as early as possible, and the appropriate treatment or the careful monitoring of fetal wellbeing. PMID- 10875102 TI - Intrauterine MRI with single-shot fast-spin echo imaging showed different signal intensities in hypoplastic lungs. AB - Ultrasonography is used for the prenatal diagnosis of hypoplastic lungs. However, ultrasound poses problems because of difficulties in getting the entire lung in perspective and the results depend on the skill of the examiner. When the alveolar formation of the fetal lung is retarded, the fetus is predicted to show an altered density on MRI using an SSFSE sequence due to a varied amount of alveolar lung fluid. We present a case of twins who showed a marked difference in signal intensity of the lung on MRI, which was useful for predicting the fetal pathophysiology. Intrauterine MRI provides the possibility of diagnosing hypoplastic lungs prenatally. PMID- 10875103 TI - Congenital lobar emphysema occurring in twins. AB - Twins born with congenital lobar emphysema are reported. This has not previously been described. Diagnosis, surgical management and subsequent course is high lighted. These twins may well be the smallest to have operative treatment. PMID- 10875104 TI - Fetal intrathoracic injuries following mild maternal motor vehicle accident. AB - Reported herein are the cases of three infants who were born with serious intrathoracic injuries, apparently sustained at the time of the mother's involvement in a motor vehicle accident. The accidents occurred at 26th, 29th and 36th weeks of pregnancy and resulted in minimal injuries to the mothers themselves. The infants were born four weeks, three hours and two days later, respectively. Their injuries were manifested (singly) by hemothorax, pneumothorax and contusion of lung, the latter in a setting of multi-organ trauma. We suggest that chest x-ray, in addition to brain ultrasound, be routinely included in the evaluation of neonates whose mothers were involved in a motor vehicle accident during pregnancy, not excluding cases wherein the mother's injuries were negligible or inapparent and regardless of the time elapsed between accident and delivery. PMID- 10875105 TI - [Intraaneurysmal embolization for cerebral aneurysms]. PMID- 10875106 TI - [Acute surgical and endovascular therapy for stroke: especially patients with brain infarction]. AB - From January 1994 to December 1997, 845 patients with stroke were admitted to Hakodate Municipal Hospital. They consisted of 514 patients with brain infarction, 206 with brain hemorrhage, 121 with subarachnoid hemorrhage and 4 with intracranial hemorrhage from arteriovenous malformation. The clinical categories of brain infarction were as follows; atherothrombotic recognized in 158 patients, cardioembolic in 114, lacunar in 217 and other categories in 25. With regard to the cures of brain infarction in the acute phase, direct percutaneous transluminal angioplasty (direct PTA) was carried out on three patients with atherothrombotic infarction, immediate PTA on two, superselective fibrinolytic therapy on two, and STA-MCA anastomosis on three. In all, ten atherothrombotic patients (6.3%) were treated by acute surgical or endovascular therapy. On the other hand, superselective fibrinolytic therapy was carried out on 35 patients (30.7%) with cardioembolic infarction. There were no patients in the lacunar infarction group who were given acute surgical treatment. Neurological improvement after 24 hours was recognized in 4 patients (40%) of 10 with atherothrombotic infarction, and in 9 patients (25.7%) of 35 with cardioembolic infarction. However, symptomatic intracerebral hematoma was recognized in 4 patients (11.4%) with cardioembolic infarction. Indication for acute surgical or endovascular treatment for brain infarction was very limited because of the time factor from the onset to admission. It is suggested that neurosurgeons might enlighten citizens about the necessity for acute surgical or endovascular therapy for stroke. PMID- 10875107 TI - [Clinical characteristics and indications for shunting in patients with idiopathic normal pressure hydrocephalus with brain atrophy (atypical idiopathic normal pressure hydrocephalus)]. AB - PURPOSE: To clarify clinical characteristics of atypical idiopathic normal pressure hydrocephalus (AINPH) and indications for shunt operations. SUBJECTS AND METHODS: Subjects examined in the present study included 65 patients who satisfied the following 4 diagnostic criteria of AINPH and underwent V.P shunt with Medos type shunt system; set pressure: epidural standard pressure x 13.6 - 20 mmH2O (omission of a figure in the first place). The diagnostic criteria were: 1) no apparent history of intra- or extra-cranial disease; 2) dementia was present as a main complaint; 3) the presence of moderate to severe cerebral atrophy and ventricular enlargement and PVL around the anterior horn on CT scans; 4) normal cerebrospinal pressure and filling of ventricles or subarachnoid space with contrast medium at 24 hours on cisternography. The patients were aged 49-83 with the mean age of 62.9 years; the ratio of male to female was 37:28. They were categorized as shunt-effective (group E: 36 cases) or non-shunt-effective (group NE: 29 cases), and the following parameters in both groups were compared: 1. clinical characteristics: 2. the presence or absence of pressure wave (PW) during preoperative continuous epidural pressure measurement (EDPM) 3. CSF outflow resistance (Ro) 4. preoperative serum alpha-1-antichymotrpsin (alpha-1-ACT) 5. cerebral arteriovenous difference of oxygen content (c-AVDO2) before and after surgery 6. mean cerebral blood flow (mCBF; 99mTc-HMPAO-SPECT) before and after surgery. RESULTS AND CONCLUSIONS: 1. Group E had a shorter duration between symptom onset and hospital visit (within 16 months and showed hyporoluntary and hyporeactivity as their main complaints associated with gait disturbance; the time course of symptoms was classified as suddenly progressing and fluctuating in many cases. Group NE had a relatively longer duration between symptom onset and hospital visit and showed activeness, wandering, nervousness and quick temper as their main complaints; the time course of symptoms was classified as progressing in many cases. 2. PW-positive cases were all included in group E. but some PW negative cases were also observed in group E. 3. Ro was significantly higher in group E (p < 0.01), and cases with a Ro value over 20 mmHg/ml/min. were all included in group E. 4. alpha-1-ACT was significantly lower in group E (p < 0.05), and cases with an alpha-1-ACT value over 55 mg/dl were all included in group NE. 5. Although preoperative c-AVDO2 was significantly higher in group E (p < 0.05), cases with a c-AVDO2 value over 8.5 ml% were all included in group NE. c AVDO2 values were within 5-8.5 ml% in all cases of group E. 6. mCBF significantly increased after surgery in group E (p < 0.001). 7. It was confirmed that cerebral atrophy in group E on AINPH is caused by a cerebral circulation disturbance defined as a cerebral blood flow of penumbra or more due to cerebral arteriosclerosis, etc. 8. A flowchart of indications for shunt surgery for AINPH was prepared based on the results of the present study. PMID- 10875108 TI - [Microsurgery of cervical intramedullary ependymomas extending into the medulla oblongata]. AB - Intramedullary spinal cord ependymomas are well-demarcated tumors, which can be totally resected without significant morbidity. However, it is challenging to remove intramedullary ependymomas extending into the medulla oblongata, because the medulla oblongata has such important roles as regulation of respiration, circulation, and digestion. We describe here three cases of intramedullary ependymomas extending into the medulla oblongata. The tumors were localized in the dorsal part of the caudal medulla oblongata and were surrounded by the nuclei and tracts related to the dorsal columns. The tumors were safely exposed by dividing the medulla oblongata along the posterior median sulcus and resected from the surrounding normal tissues. Although patients demonstrated transient dorsal column dysfunction postoperatively, the symptom finally recovered. PMID- 10875109 TI - [Avascular necrosis of the femoral head caused by steroid treatment in neurosurgery]. AB - Steroid induced avascular necrosis of the femoral head is a well known disease, but, there are few reports about the disease in neurosurgical patients. In the neurosurgical field, the use of steroids has become prevalent since the 1960's. Recently, the adverse effect of steroids and the limitation of its effect have been highlighted, but its use against neurosurgical diseases is still a common treatment to prevent cerebral edema or to counteract hypo-pituitarism caused by hypophyseal lesions. We reviewed 250 patients of avascular necrosis treated between 1985 and 1997 in our institute. Within these patients, 11 (4.4%) were treated with steroid during neurosurgical treatment. Six patients were treated for brain tumors near hypophyseal lesions, and 5 patients were treated for head injury or cerebro-vascular disease. It is concluded that total steroid dose over 5000 mg such as hydrocortisone may become a high risk for causing avascular necrosis of the femoral had in neurosurgical disease, and it may occur even with the supplemental steroid treatment against hypo-pituitarism. The onset is usually 2 or 3 years after the neurosurgical treatment, when neurosurgical care is no longer needed. Therefore, it tends to be ignored in the neurosurgical field. The treatments against avascular necrosis of the femoral head were femoral head osteotomy or conservative management, and good results were obtained. Early diagnosis and early treatment is essential. Further consideration concerning steroid treatment in neurosurgical patients may be required. PMID- 10875110 TI - [Systemic and cerebral hemodynamic changes during craniotomy under mild hypothermia]. AB - Systemic and cerebral hemodynamics were evaluated through pulmonary artery and jugular bulb catheters in 31 patients with cerebral aneurysms. The temperature of the pulmonary artery was lowered to 33-34 degrees C during surgeries. Systemic circulation deteriorated during mild hypothermia, but cerebral circulation showed a little improvement. Cardiac function recovered by rewarming, but the balance between systemic oxygen supply and demand did not always improve. Therefore, a strict monitoring of systemic circulation is necessary even when the hypothermia is only mild and vasodilator or inotropics would be indicated. Rapid rewarming should be avoided, especially when surgical retraction of the brain is strong or temporary occlusion is prolonged. PMID- 10875111 TI - [A case of multiple metastatic brain tumors with repeated intracerebral hemorrhages]. AB - We report a case of multiple metastatic brain tumors with repeated intracerebral hemorrhages. A 73-year-old man suffered from a cerebellar hemorrhage. Subsequent hemorrhages repeatedly occurred in the right temporal lobe, the 4th ventricle, the midbrain, and the septum pellucidum. Three months after admission, CT revealed enhanced masses with surrounding edema in the cerebellar vermis and midbrain, suggesting brain tumors. We eventually diagnosed these masses in an autopsy as metastatic brain tumors of lung adenocarcinoma. Intravascular embolization with tumor cells was a probable cause of the multiple repeated intracerebral hemorrhages. PMID- 10875112 TI - [Agenesis of the right internal carotid artery associated with complicated anastomosis of middle cerebral artery: a case report]. AB - A 57-year-old female was admitted to our hospital because of headache, nausea, and vomiting. Head CT scan demonstrated subarachnoid hemorrhage. Cerebral angiography showed the absence of the right internal carotid artery, and skull base CT of the bone window level revealed the absence of the right carotid canal. The right middle cerebral artery (MCA) and anterior cerebral artery (ACA) were opacified from the left internal carotid artery. The right A1 portion was hypoplastic and the distal portion of the right M1 portion was replaced by several minute complicated anastomotic vessels connected to the right M2 portion. The right MCA territory was mainly supplied by collateral flow from the right ACA and the right posterior cerebral artery via the leptomeningeal anastomosis. These was neither aneurysm nor arteriovenous malformation. The second angiography, 1 week after the initial angiography, showed the same hemodynamic pattern and aneurysms were not found. We diagnosed the patient as agenesis of the right internal carotid artery and the etiology of subarachnoid hemorrhage was suspected to be a rupture of the anastomotic vessels between the right M1 and M2. She was discharged on the 21st hospital day without any neurological deficit. PMID- 10875113 TI - [Prolactinoma in a child showing high MIB-1 labeling index: a case report]. AB - We report a very rare case of a prolactin secreting pituitary tumor (prolactinoma) which occurred in a 12-year-old boy. The tumor showed an extremely high MIB-1 index. The clinical implication in the postoperative management of childhood prolactinoma is discussed. The patient showed right third nerve palsy, and MRI revealed a pituitary tumor invading the right cavernous sinus. Preoperative hormonal evaluation showed a very high prolactin level (2800 ng/ml). The patient underwent transsphenoidal surgery, and the third nerve palsy disappeared just after the procedure. MIB-1 index obtained by using immunostaining was 18.9%. Postoperative prolactin level remained high (2200 ng/ml), and the patient was treated with 10 mg/day of bromocriptine. Prolactinomas in children with high MIB-1 index show resistance to treatment with bromocriptine. In the postoperative management of a childhood prolactinoma, it should be considered how to control sufficiently high serum prolactin level to expect sexual development while preserving other normal residual pituitary functions. If control with bromocriptine, fails radiation treatment should be adopted with careful observation of the increase in height and the progress of sexual development of the patient. PMID- 10875114 TI - [Coil embolization for incidental aneurysms in patients with chronic renal failure: midterm clinical results of two cases]. AB - In spite of recent advances in perioperative management, the risk of neurosurgical intervention for patients with chronic renal failure is still considered too high. In this study, coil embolization for incidental aneurysms in such patients is demonstrated in reference to midterm results. A 42-year-old woman with a history of hemodialisis for 7 years presented with subcortical hemorrhage in her right frontal lobe. The magnetic resonance angiography (MRA) demonstrated a distal anterior cerebral artery aneurysm, but it was considered to be unrelated to the hemorrhage. Two and a half months after the hemorrhage the aneurysm was embolized with interlocking detachable coils. Thirty months after embolization, the angiogram revealed the coil compaction and the recanalization of the aneurysm neck. However, 54 months after embolization, the figure of the embolized aneurysm and neck remnant was the same as the previous findings. A 69 year-old woman with a history of hemodialisis for 5 years suddenly experienced left hemiparesis. Computed tomography revealed cerebral infarction in the right frontoparietal white matter. In addition, a left middle cerebral artery aneurysm was unexpectedly found on the MRA. Five months after the onset of the attack, the aneurysm was embolized with a Guglielmi detachable coli. An angiogram obtained 24 months after the embolization showed the aneurysm to be almost completely obliterated. In considering the therapeutic risks and benefits for incidental aneurysms of patients with chronic renal failure, intra-vascular surgery could be recommended as a less invasive treatment. PMID- 10875115 TI - [Spontaneous occlusion of a dissecting aneurysm in the shape of "two dumplings on a skewer" at righ posterior inferior cerebellar artery (PICA): report of a case and neuroradiological findings]. AB - We present a case of a spontaneous dissecting aneurysm at the vertebrobasilar artery including the right PICA in a 44-year-old man, who suffered from headache, hiccup and ataxic gait. The arteriograms showed an irregular narrowing and dilatation in the right PICA and in the vertebrobasilar artery, and showed fusiform dilatations in the bilateral middle cerebral arteries. We observed intramural hematoma and true lumen at the right PICA dissecting aneurysm on T1 weighted images on magnetic resonance imaging (1.5T, MRI), and the intimal flap was enhanced on T1-weighted image after intravenous injection of Gd-DTPA. The shape of the intramural hematoma showed a unique "two dumplings on a skewer" appearance, and the intensity of its hematoma in the false lumen decreased in gradient from adventitia to intimal flap on T1-weighted image on MRI. The dissecting aneurysm of the PICA was occluded spontaneously 1 month later, and it caused cerebellar infarction. However, the patient has been left only with the symptom of slight trunkal ataxia. Various shapes of intramural hematomas on MRI have been reported by Kitanaka in association with intracranial vertebrobasilar dissections. We suggest that "two dumplings on a skewer" shape which corresponds to the flow void of the true lumen, accompanied by intramural hematoma and enhanced intimal flap, on contrast-enhanced T1-weighted image, should be regarded as a true "diagnostic sign" of a dissecting aneurysm. PMID- 10875116 TI - [Pathogenesis of cerebral vasospasm after subarachnoid hemorrhage]. PMID- 10875117 TI - [Histopathological changes in cerebral vasospasm after subarachnoid hemorrhage]. PMID- 10875118 TI - [Drug therapy]. PMID- 10875119 TI - [Endovascular treatment for cerebral vasospasm]. PMID- 10875120 TI - [Genetic factors involved in the development of bacterial CNS infections]. PMID- 10875121 TI - [Cardiac function estimated by Doppler echocardiography in patients with hypertensive intracerebral hemorrhage]. AB - We carried out transthoracic Doppler echocardiography on 38 patients with hypertensive intracerebral hemorrhage(ICH) to examine cardiac functions. The ratio of myocardial hypertrophy and the prevalence of valvular regurgitation were investigated. The E/A ratio which reflects left ventricular diastolic function and the ejection fraction(EF) of each patient were also studied to demonstrate cardiac functions in patients with hypertensive ICH. Myocardial hypertrophy was found in 39.5% of patients with hypertensive ICH. The aortic regurgitation showed a trend to have a higher prevalence in patients with hypertensive ICH when compared to normal volunteer group. Although there was no significant differences in the ejection fraction between groups, the E/A ratio in patients with hypertensive ICH showed a significantly lower value than in normal volunteer group. Therefore, transthoracic Doppler echocardiography demonstrated that patients with intracerebral ICH showed a trend to have a myocardial hypertrophy, aortic valvular regurgitation, and the lower ventricular diastolic function. Thus, transthoracic Doppler echocardiography is useful to evaluate cardiac functions in patients with hypertensive ICH. PMID- 10875122 TI - [The Japanese version of the serotonin syndrome scale (JSSS)]. AB - The serotonin(5-HT) syndrome(SS) is a condition of both the central and peripheral 5-HTergic hyperstimulation, characterized by a constellation of 5-HT related side effects(confusion, agitation, restlessness, myoclonus, hyperreflexia, diaphoresis, shivering or tremor in the setting of the recent addition of 5-HTergic agents. The SS is produced most often by the concurrent use of monoamine oxidase inhibitors and other 5-HTergic agents. However, more recent reports suggest that the tricyclic antidepressant or selective serotonin reuptake inhibitor(SSRI) monotherapy induces the SS. Recently, for the operationalized assessment of both the presence and the severity of the core symptoms of the SS, Hegerl et al. developed the Serotonin Syndrome Scale(SSS) as a modification of the diagnostic criteria of the SS proposed by Sternbach. Since, in Japan, some novel 5-HTergic agents have been, and will be in use, recognition of the SS is quite important. Therefore, for clinical application of SSS, we prepared the Japanese-language version (JSSS). PMID- 10875123 TI - [Occipital encephalocele and epilepsy]. AB - We report the incidence of epilepsy in 7 patients with occipital encephalocele (meningocele: 6 cases, meningoencephalocystocele: 1 case). Two cases had epilepsy and a case without the history of epileptic seizure had an epileptic pattern on the EEG. All these 3 cases had cortical dysplasia(CD) such as schizencephaly and subcortical gray matter, and two of them were associated with mental retardation. The location of the prominent CD was correlated with the EEG abnormalities and clinical epileptic pattern in each case. Associated hydrocephalus and placement of VP shunt had no significant influence on the occurrence of the epilepsy. Thus, complicated CD is thought to be epileptogenic in patients with occipital encephalocele. PMID- 10875124 TI - [Hemifacial spasm due to a compression of the facial nerve by a fusiform aneurysm of the vertebral artery: case report]. AB - We report a rare case of symptomatic hemifacial spasm caused by a fusiform vertebral artery aneurysm and by a branch of the anterior inferior cerebellar artery compressing the facial nerve at the root exit zone (REZ). A 71-year-old female had an 11-year history of right hemifacial spasm. MRIs demonstrated an aneurysm compressing the facial nerve at the REZ. Angiography disclosed a fusiform aneurysm of the right vertebral artery at the origin of the posterior inferior cerebellar artery. After the vertebral aneurysm was clipped distal to the origin of the posterior inferior cerebellar artery, a branch of the right anterior inferior cerebellar artery was also observed compressing the facial nerve at the REZ. Both the clipped aneurysm and the branch of the anterior inferior cerebellar artery were mobilized away from the REZ of the facial nerve, and a prosthesis was inserted between the branch of the anterior inferior cerebellar artery and the brain stem to keep the aneurysm away from its original position. The patient's hemifacial spasm immediately disappeared without any neurological deficits just after the surgery. Hemifacial spasm, especially caused by an aneurysm, is quite rare. In a review of the literature, we found only 4 cases of symptomatic hemifacial spasm caused by an aneurysm of the vertebral artery. This case is the first reported case of hemifacial spasm caused by both a fusiform vertebral artery aneurysm and a branch of the anterior inferior cerebellar artery compressing the facial nerve at the REZ. PMID- 10875125 TI - [A case of idiopathic cranial hypertrophic pachymeningitis presenting Tolosa-Hunt syndrome]. AB - A 48-year-old female was seen because of left orbital pain. The neurological findings were normal at her first visit. She presented temporary double vision during conservative period. Plain CT revealed no mass around the sellar region. Enhanced CT revealed enhanced mass in the left cavernous sinus. MRI revealed low intensity lesion on both T1 and T2 weighted images. Enhanced MRI showed strongly enhanced mass extended from the left cavernous sinus to the dura of sellar floor, the contralateral cavernous sinus, and cerebellar tentorium. Angiography showed stenosis of the left internal cerebral artery. Biopsy from the dura of the tentorium was performed via anterior temporal approach. Abundant collagen fibers with hyalinization were seen in the thickened dura. A final diagnosis of idiopathic cranial hypertrophic pachymeningitis presenting Tolosa-Hunt syndrome was made. Symptoms were free just after the surgery, however, the regrowth of the tentorial lesion was found on MRI after one year. The size of the lesion decreased after administration of steroid. PMID- 10875127 TI - [Motor weakness distal to the wrist joint]. PMID- 10875126 TI - [Dissociation of voluntary eye closure--to keep the eyes closed and to blink- following right hemisphere stroke]. AB - We report a case of a unique eye sign following right hemispheric infarction. This patient was a 78 year old right-handed woman. There was a history of a left hemispheric stroke 1 year previously. On admission, she showed left hemianopia, dysarthria, mild left central facial paresis, bilateral sensory deficit and quadriparesis which were marked on the left side. Babinski sign was elicited on the left. She did not have anosognosia or visual neglect. She had mild orofacial apraxia, but ideomotor and ideational apraxia was absent. There was no motor impersistence. Magnetic resonance imaging of the brain revealed a recent infarction in the territory of the right middle cerebral artery and an old infarction in the left tempro-parietal lobe. The patient could not open her eyelids to verbal command or voluntarily until about two weeks later, when she became able to open her eyes but showed difficulty keeping her left eye closed. She was aware of this problem and could repeat the command and comprehend what was requested to her. On verbal command to close the eyes, her right eye would be closed continuously and excessively and the left eye would only blink. When requested to blink, however, she could blink correctly without excessive eye closure. Spontaneous, reflex and voluntary blinking were normal. Her eyes were closed normally during sleep. Blepharospasm was not seen. The patient showed a striking dissociation between a failure to close her eyes continuously and a preserved ability to blink voluntarily. We suggest that her ability to contract palpebral portion of her left orbicularis oculi muscle is preserved. Regarding the mechanism of the voluntary eye closure system, separate control mechanisms should exist on closing eyes continuously and blinking. PMID- 10875128 TI - [Extrapontine myelinolysis]. PMID- 10875130 TI - [Osteonecrosis--an unsolved problem]. PMID- 10875129 TI - [A 59-year-old woman with recurrent convulsive seizures, cerebral infarctions, dementia, and intracranial calcifications]. AB - A 59-year-old woman with recurrent seizures and progressive dementia is reported. Her past history and familial history were unremarkable. She became short tempered at 56 years old (Oct. 1991). She had the first seizure attack and was admitted to a hospital at March 4, 1993, with prolonged disturbance of consciousness and subsequent mental deterioration. Her brain CT showed multiple small calcifications in the subcortical white matter and pons. The laboratory data including blood count, serum chemistry, serological studies and CSF was normal. MRI and digital subtraction angiography of the cranial vessels were unremarkable. There was a decrease in accumulation in the right cerebral hemisphere on 123I IMP SPECT. Despite anti-convulsant therapy, she had recurrent seizures several times, with gradual worsening of her mental state. She had the latest seizure attack and was transferred to Matsusaka Chuo Hospital, on August 29, 1993. After the attack she had been in the apallic state, and died on Nov. 13, 1995. This case was discussed in a neurological CPC. The discussants suggested that the isolated angiitis of the central nervous system caused secondary seizures and cerebral infarctions. Post-mortem examination revealed the CNS findings of vasculitis at various stages, calcification or mineralization mainly in the subcortical white matter and pons, massive cerebral infarctions with massive exudate, fresh and old small bleedings and exudate around the inflamed or calcified vessels. The white matter degeneration resembled that of Binswanger leukoencephalopathy. The final pathological diagnosis was isolated angiitis of the central nervous system since there was no inflammatory changes or atherosclerotic change of the blood vessels in the extracranial organs. PMID- 10875131 TI - [Epidemiological risk factors for non-traumatic osteonecrosis]. AB - Certain fractures and/or dislocations of the femoral head are known to cause arterial injury and result in post-traumatic osteonecrosis. However, the more complex etiology of non-traumatic osteonecrosis is multifactorial and includes chemotherapy, radiotherapy, thermal injuries, and especially coagulopathies, which are now commonly observed in these patients. Intravascular coagulation with fibrin thrombosis begins in the capillaries and sinusoids of the intraosseous microcirculation, and residual venous thrombosis is more likely to occur if there is coexistent hypofibrinolysis. Coagulopathies are intermediary events, which are always activated by some underlying etiologic risk factor(s). Conditions capable of triggering intravascular coagulation include familial thrombophilia (resistance to activated protein C, decreased protein C, protein S, or antithrombin III, and hyperhomocystinemia), hyperlipemia and embolic lipid (alcoholism and hypercortisonism), hypersensitivity reactions (allograft organ rejection, immune complexes, and antiphospholipid antibodies), bacterial endotoxic (Shwartzman) reactions and various viral infections, proteolytic enzymes (pancreatitis), tissue factor release (inflammatory bowel disease, malignancies, neurotrauma, and pregnancy), and other thrombophilic and hypofibrinolytic disorders. Currently known risk factors for non-traumatic osteonecrosis of the femoral head are described briefly in this review article. PMID- 10875132 TI - [Diagnostic imaging in femur head necrosis]. AB - Diagnosis of avascular necrosis (AVN) of the hip has been improved by the technical progress of imaging modalities during the last decade. For a long period, only plain radiographs had been available. Scintigraphy and computed tomography contributed to differential diagnosis and early detection of bone necrosis. In the meantime, MR imaging has gained special value in the evaluation of AVN. It is now the method of choice for early detection as well as for assessment in later stage disorders. Using the ARCO system, all imaging modalities and their diagnostic viability are described. Findings regarding the different stages of AVN are correlated to tissue-specific changes. PMID- 10875133 TI - [Pathomorphological aspects and repair mechanisms of femur head necrosis]. AB - The pathomorphologies of non-traumatic femoral head osteonecroses (ON) are usually similar, despite various known pathogenetic factors. The size and position of the subchondral bone and marrow segment, becoming necrotic after the ischemic event(s), and the kind of repair processes determine the time course and thus the fate of this hip joint disease. Four cases of conservatively or core decompression-treated femoral head ON were selected to demonstrate differently effective repair mechanisms which are discussed in respect to existing therapeutic concepts. Diagnostic criteria from magnetic resonance imaging follow ups were correlated with light microscopy findings on undecalcified ground and microtome sections from femoral heads retrieved at total joint replacement. Initial stage (ARCO 0) and reversible early stage ON (ARCO 1) after incomplete ischemias can apparently show spontaneous sufficient repair. After extensive and complete ischemia, however, ON progresses without detectable changes on plain radiographs into irreversible early stage ON (ARCO stage 2). Only in exceptional cases (with small, medially located necroses), a spontaneous sufficient repair seems possible. Usually, early ARCO stage 2 ON with intact articular surface shows no remodeling of the subchondral necrotic bone and fatty marrow, but only ineffective repair with fibrovascular tissue invasion and bone resorption at the vital bone border. Repeated bone appositions on partly resorbed necrotic trabeculae form the sclerotic rim in this pathognomonic reactive interface. New bone formation can also be increased underneath the necrotic area and reactive interface when surrounded by accompanying bone marrow edema. Core decompression in ARCO stage 2 ON, even if it reaches the necrotic lesion, can at best delay progression of the disease, but never leads to complete reconstruction of the necrotic area. More likely, after both conservative and operative treatment, destructive resorption without effective consecutive bone formation will lead sooner or later to collapse of the articular surface and thus to mechanical instability of transition stage ON (ARCO stage 3). On the other hand, this subchondral fracture can apparently also cause reconstructive repair which, by involving chondral and membranous ossification in this "creeping substitution", can reduce the necrotic area. However, it cannot prevent progression into late stage ON (ARCO stage 4) with secondary joint destructions. Principally, besides the rare sufficient repair in initial and certain early ON, three forms of insufficient repair in the necrotic area can be distinguished: lack of remodeling, destructive remodeling, and reconstructive remodeling. To date, no therapeutical intervention exists which leads to complete healing of irreversible ON stages by reconstructive repair. Improved understanding of pathomorphology and repair mechanisms, however, could be the basis for future therapeutical concepts which should aim at the complete regeneration of the osteonecrotic area. PMID- 10875134 TI - [Osteonecrosis: natural course and conservative therapy]. AB - The natural course of Osteonecrosis (ON) of the femoral head has yet not been evaluated sufficiently. Especially in the early forms of the disease (ARCO 0 to II) without collapse of the femoral head, useful information on the natural course could only be collected since the routine use of MR-imaging. The unspecific findings in ARCO stage I with negative radiographs are potential reversible. The "point of no return" already lies in the irreversible ARCO stage II in almost all cases. Prognosis for further progression for both early stages depends primarily on the extension and location of the lesion. Only the rare, small to medium sized lesions in the medial or central location may have a good prognostic course over a period of more than five years. The much more common large sized and lateral located lesions will have a probability of about 80% to progress to femoral head collapse within two years. Conservative therapy with single protected weight bearing has shown bad results not significantly different from the natural course of the disease. Pulsed electromagnetic fields are still discussed controversially. Until now there is no MRI controlled study available in early ON, showing superior results compared to the easy and cost-effective core decompression therapy. The extra corporal shock wave therapy has shown spontaneous pain relief in early ON. Midterm results of this new therapeutical concept are not available yet. Hyperbaric oxygen therapy has shown to stimulate the repair process in an animal experiment. The preliminary clinical results of this time and cost consuming therapy are not convincing. Vasoactive drugs in combination with limited weight bearing may play a role in the conservative management of early ON (ARCO I and part of ARCO II) in the future. For the conservative therapy in ON several other substance are currently in clinical testing. The benefit of conservative therapeutical concepts in ON in the future can only be evaluated with prospective MRI controlled clinical studies. The use of cytokines in combination with surgical debridement and bone grafting of the necrotic area may be a possible therapeutical concept for the future. PMID- 10875136 TI - [The value of core decompression in treatment of femur head necrosis]. AB - Core decompression of the necrotic area for treatment of idiopathic osteonecrosis of the femoral head was developed and published by Ficat and Arlet in 1962 within the scope of their "Functional exploration of bone". The mode of action is attributed to a reduction of the intramedullary pressure in the bony compartment of the femoral head. The possibilities of repair and bone regeneration following core decompression are still discussed controversially. Core decompression is a common but not generally accepted procedure in the treatment of idiopathic osteonecrosis of the femoral head. After first publications of positive mid- and long-term effects, some subsequent studies judged it as an ineffective and high risk method. Analysis of the literature shows that the effectiveness of core decompression depends on the stage of osteonecrosis at the time of surgical intervention. Prognosis is influenced by the extent and location of the necrotic area, the presence and amount of head depression, and continued risk factors- mainly corticoid medication. The best prognosis can be given for patients with a small, medial-centrally located necrosis without head depression. The classification according to Ficat appears to be insufficient, as the extent and localization of the necrotic area are not assessed. Magnetic resonance imaging has become a diagnostic gold standard, as radiographic diagnosis showed poor sensitivity and specificity, especially in the early stages of the disease. As an essential part, MRI was integrated into the new classification of the "Association Internationale de Recherche sur la Circulation Osseuse" (ARCO). On account of the literature and our own experience, treatment by core decompression can be recommended in cases of reversible early stages of osteonecrosis (ARCO 1), as well as in those cases of irreversible early stages (ARCO 2) that show a medial or central location of the necrosis with an extent of less than 30% of the femoral head. Once the disease reaches the irreversible early stage, complete recovery cannot be expected. In these cases only reduction of pain and retardation of the natural course of the osteonecrosis are possible to gain time until total hip replacement is unavoidable. PMID- 10875135 TI - ["Transient osteoporosis" as a special reversible form of femur head necrosis]. AB - There is still controversy whether transient osteoporosis of the hip joint represents a distinct self-limiting disease, or reflects only an early, reversible subtype of non-traumatic osteonecrosis (ON). Transient osteoporosis has several synonyms: algodystrophy of the hip; transient marrow oedema; or bone marrow oedema syndrome--BMOES. Clinical presentation of BMOES shows mechanical hip joint pain, ON risk factors, and a diffuse bone marrow oedema in MR imaging. Histomorphological changes resemble early ON, but with diffuse sufficient repair in BMOES and focal and insufficient repair only at the border of the necrotic lesion in ON. Therefore the clinical course and outcome are significant different, with restitution occurring in BMOES, while progressive destruction of the joint takes place in ON. So far, the preferred treatment strategies are protected weight bearing for BMOES, but operative treatment for ON. In a prospective study of patients with BMOES, the clinical, radiographic, and MRI course of 43 hip joints after core decompression treatment were investigated. All patients showed immediate relief of pain after surgery and the average duration of symptoms with conservative treatment could be dramatically reduced by core decompression from 6 months down to 2 months. There were no perioperative complications. Based on our experience with over 100 BMOES patients, we are convinced that this syndrome represents not a distinct disease but an early reversible subtype of non-traumatic ON. Due to the excellent clinical results of core decompression, we recommend this operative therapeutical concept in patients with painful BMOES. PMID- 10875137 TI - [Osteonecrosis of the hip joint in adulthood. Significance of various corrective osteotomies]. AB - The importance of osteotomies in treatment of osteonecrosis of the femoral head has decreased. However, with proper selection of the patient, osteotomies are still useful for small stage III or IV lesions in patients younger than 45 years, and with no ongoing causes for progressive osteonecrosis such as the use of high doses of corticosteroids, chemotherapy, and chronic alcoholism. For preservation of natural hip joint function, the necrotic segment is moved away from the load transmitting area of the acetabulum, and weight-bearing forces are redistributed to articular cartilage that is supported by healthy bone. In addition to radiographic evaluation, MRI is most accurate for imaging of the stage, localization, and extent of osteonecrosis in planning and selecting osteotomy. PMID- 10875138 TI - [Potential uses of cytokines and growth factors in treatment of osteonecrosis]. AB - Osteonecrosis of the femoral head remains a devastating disease for young patients. As the normal process of bone formation, bone destruction, and fracture healing becomes more clearly understood, molecular agents--including cytokines, bone morphogenetic proteins, and angiogenic factors--will be identified as potential therapeutic agents for the treatment of osteonecrosis. As the pathology of osteonecrosis and repair of osteonecrotic lesions becomes clear, the potential combination of these molecular factors to influence the outcome of the disease in its repair process should become evident. With the myriad of agents and combinations of agents which may be beneficial in the treatment of osteonecrosis, a reproducible animal model is urgently needed to determine which of these combinations is most effective. Despite the lack of an animal model, progress in the use of cytokines for osteonecrosis treatment in conjunction with traditional treatment methods is possible in human subjects. This is due to the extremely low incidence of adverse reactions when these cytokines are used locally in nanogram to microgram quantities. PMID- 10875139 TI - [Partial and total joint replacement in femur head necrosis]. AB - In literature, the results of hip arthroplasty in patients with avascular osteonecrosis of the femoral head vary. The main reason may be the nonhomogeneous patient groups concerning etiology of the femoral head necrosis (FHN). Analyzing the results of hip endoprosthesis in relation to the etiology of FHN leads to the assumption that steroid-induced FHN and FHN with underlying systemic bone diseases (renal osteodystrophy, sickle-cell hemoglobinopathy) have the highest loosening rates. Diseases with immunosuppressive medication and sickle-cell hemoglobinopathy have the highest risk of joint infection. Therefore etiology plays an important role in the long-term results of hip endoprostheses in FHN. Modern cement techniques of the second generation and new non-cemented total hip endoprostheses seem to have better results than older prostheses and cement techniques. We followed-up 52 non-cemented thrust plate prostheses in 45 patients with FHN, prospectively, for at least 2 years (3.7 +/- 1.6 years). The revision rate was 9.6% (two aseptic loosenings in one patient with renal osteodystrophy and one patient with alcohol abuse, as well as three late infections in one patient with alcohol abuse and two patients with renal osteodystrophy). Additionally, five prostheses showed radiologic lines of a minimum of 2 mm. Future studies with longer follow-up are needed to find out whether these prosthetic designs with proximal fixation of the femoral component preserving the diaphysial bone have advantages in young FHN patients. PMID- 10875140 TI - [Therapy of osteonecrosis. Basic principles and decision aids]. AB - The treatment of osteonecrosis of the femoral head involves a continuum based on a radiographic spectrum of disease. Core decompression or pharmacological agents can be utilized for the earliest small or medium-sized pre-collapsed lesions. For these types of lesions, osteotomy has been tried by various authors with moderate success. For small or medium lesions that are post-collapse, various bone grafting procedures have been used. This approach should be tempered with a look at the articular cartilage if this is damaged or the lesion is large. Limited femoral resurfacing can be used for hips that do not have acetabular involvement. If there is acetabular involvement, total hip replacement remains the treatment of choice. There are present innovations in total hip arthroplasty that hopefully will lead to increased longevity of these prostheses with newer polyethylenes as well as the use of ceramic and other types of interfaces. Another possible advance for this disease would be the use of metal on metal standard prostheses, as well as metal on metal resurfacing arthroplasties. In terms of a salvage of the femoral head, all of the different procedures--core decompression, osteotomy, bone grafting--can be enhanced by new advances in the development of the utilization of bioactive factors. These range from osteoinductive agents such as cytokines, angiogenic stimulating factors, and bone morphogenetic proteins. In addition, osteoconductive substances may be helpful and can be combined with osteoinductive substances. These bioactive factors are described in detail in another chapter of this issue. With the ushering in of the millennium, there is hope for better results with this disease. PMID- 10875141 TI - [Extracorporeal shockwave therapy in treatment of epicondylitis humeri radialis. A current overview]. AB - In the past, extracorporeal shock-wave therapy (ESWT) has been used increasingly as a treatment for conservatively unsuccessfully treated radiohumeral epicondylitis. However, published reviews of clinical trials on the efficacy of ESWT have led to inconsistent results and are outdated or methodologically inadequate. As a consequence, a systematic literature search was conducted which yielded 20 relevant papers that described trials on the efficacy of ESWT in the treatment of radiohumeral epicondylitis. These were rated according to biometrical criteria for the conduct of therapeutic trials. None of the rated trials fulfilled all of the criteria, and it is concluded that the efficacy of ESWT in the treatment of epidondylitis can presently be neither confirmed nor excluded. PMID- 10875142 TI - [Metatarsalgia. Treatment of the dorsally dislocated metatarsophalangeal joint]. PMID- 10875143 TI - Are guidelines followed in Helicobacter pylori diagnosis and therapy? An inquiry among gastroenterologists, referring physicians and patients in Munich. AB - BACKGROUND AND STUDY AIMS: Attempts to standardize Helicobacter pylori (Hp) diagnosis and therapy have led to the publication of guidelines by various national gastroenterological societies in Europe and the USA. However, little information is available either regarding the compliance of gastroenterologists and referring physicians with these guidelines, or regarding the patients' perspective. PATIENTS AND METHODS: A retrospective analysis was conducted of all outpatient upper gastrointestinal endoscopy reports for a one-month period in eleven different centers (two university hospitals and nine private practice gastroenterology offices) with a total of 24 gastroenterologists. Endoscopy reports from patients wit the indications of reflux, diarrhea, and tumors were excluded. Diagnoses and treatment recommendations given by gastroenterologists were recorded. Questionnaires concerning Hp diagnosis, treatment indications and performance, and follow-up were sent to referring physicians and patients. RESULTS: A total of 772 endoscopy reports were included in the study; analyzable questionnaires were returned by 287 referring physicians (47%) and by 265 patients (59%). Gastroenterologists recommended Hp eradication in all ulcers and in 29% of gastritis/nonulcer dyspepsia (NUD) cases. Referring physicians thought that 94% of ulcers should be treated by Hp eradication, which was also considered to be an absolute and relative indication in NUD by 15% and 53% of the referring physicians, respectively. Among the patients who replied, 52% had received Hp eradication regimens; ulcers were found in 22% of the total patient group. Check up examinations after Hp therapy were considered necessary by 75% of the referring physicians, but only 22% of the responding patients actually underwent some form of check-up (upper gastrointestinal endoscopy in 91%). CONCLUSIONS: Gastroenterologists and (to a somewhat lesser extent) referring physician appear to be following the current guidelines for Hp treatment. As expected, two thirds of referring physicians consider NUD to be absolute or relative indication for Hp eradication. Check-up examinations are apparently being performed less frequently than recommended. PMID- 10875144 TI - Comparison of a tissue transglutaminase ELISA with the endomysium antibody test in the diagnosis of gluten-sensitive enteropathy. AB - BACKGROUND: Tissue transglutaminase (tTG) has recently been found to be the major if not the only autoantigen of gluten-sensitive enteropathy (GSE). OBJECTIVES: To further determine the significance of this finding for diagnostic (screening) and follow-up purposes, we performed tTG-based ELISAs, and compared the results to the endomysium antibody test (EMA). PATIENTS: We examined 120 serum samples from patients with celiac disease (CD) including 72 on a gluten-free diet (GFD) and eleven on a gluten challenge, 47 with dermatitis herpetiformis (DH) including 16 on a GFD and one on a gluten challenge, 96 with non-CD gastrointestinal diseases, and 117 with others; i.e. 380 serum samples altogether. Follow-up sera from 13 patients were included. METHODS: Results of an ELISA with guinea pig liver tTG were compared with the EMA test using monkey esophagus. Inhibition of endomysial staining was performed with sera positive on the EMA test but negative with the guinea pig tTG ELISA. RESULTS: The specificity and sensitivity of the tTG ELISA are high (98.6% and 92.5%). The serum IgA antibody titers against tTG decrease after introduction of a GFD. In one case, endomysial staining could not be inhibited. CONCLUSIONS: Our results show that the guinea pig tTG ELISA is suitable for use as a simple diagnostic screening and follow-up method for GSE. Further studies are necessary to identify possible additional minor antigens in GSE. PMID- 10875145 TI - [Pancreatic duct stenting in chronic pancreatitis--the controversies]. PMID- 10875146 TI - [Stenting in chronic pancreatitis--the mistakes and limitations]. AB - Endoscopic treatment of chronic pancreatitis using pancreatic duct stents was first described 15 years ago. Considering our own experience and the data of the literature we describe indications, contraindications, risks and limitations of the procedure and on the other hand its therapeutic effects. According to the actual experience an indication for pancreatic duct stenting can be seen in patients with a solitary prepapillary stenosis without stenosis of side branches or as success control for a planned surgical intervention. Contraindications are suspected malignancy, multiple pancreatic duct stenosis in the main duct or stenosis in small ducts and chronic calcifying pancreatitis with pancreatic duct stones. From 6/92 until 5/97 189 patients were operated for chronic pancreatitis in the Ulm University Hospital. Of these patients 35 (18.7%) were unsuccessfully treated preoperatively in other hospitals by pancreatic duct stent. Because of frequent complications like stent dislocation and stent occlusion repeated ERCPs (4.5/patient) and stent exchanges (3.7/patient) were performed. A therapeutical long-term benefit of pancreatic duct stenting is questionable, a definitive therapy can only be achieved in a small group of patients. However stent-induced changes of the pancreatic duct similar to chronic pancreatitis can be observed in up to 80% of all patients. Long-term observations of the reversibility of these stent-induced changes are missing, persisting chronic pancreatitis in the stented region is reported in animal models and in humans. The rates in the literature for stent dislocation and stent occlusion rate are 10-18% and 39-100% respectively. Induction of acute pancreatitis (up to 10%), duodenal reflux into the pancreatic duct, and bacterial infection with abscess formation are further severe and frequent complications of pancreatic duct stenting (1, 2). Lethal courses are reported (3, 4). Endoscopic pancreatic duct stenting in chronic pancreatitis at present is not indicated because of low success rate and a substantial risk of complications. PMID- 10875147 TI - [Actinomycosis of the pelvis with an indwelling IUD]. AB - Infection with actinomycosis israeli (an anaerobic, gram-positive bacterium) presents as chronic inflammation with tendency to fibrosis and suppuration with formation of external sinuses. Cervicofacial, thoracic and abdominal forms of the disease made up 95% of cases of actinomycosis. A 53-year-old woman was admitted to the hospital because of a pelvic mass which was thought to be malignant. A laparotomy was performed and the histologic examination showed actinomycosis. The patient first received penicillin followed by tetracyclin and the pelvic mass shrunk. One year later no more mass was detectable. We think that the IUP in place over years is the source for this infection. PMID- 10875148 TI - [Therapy-refractory thrombocytopenia in chronic hepatitis C]. AB - HISTORY AND CLINICAL FINDINGS: A 74-year-old man known to be suffering from a chronic hepatitis C infection was hospitalized because of intestinal hemorrhage, multiple petechiae and suggillations. The patient had received oral anticoagulant medication after replacement of the mitral valve. The intake of oral vitamin K antagonist had been discontinued eight days before admission. EXAMINATION FINDINGS: On admission, the platelet count was 3 G/l, the Quick's test 72%. Colonoscopy revealed diffuse mucosal bleeding in the proximal colon. Bone marrow examination showed moderate hyperplasia of erythropoiesis, as well as a marked increase in megakaryocytes. DIAGNOSIS, THERAPY AND COURSE: The diagnosis was hepatitis C-associated idiopathic thrombocytopenic purpura. The administration of both immunoglobulins and prednisone failed to increase platelets sufficiently. There was also no improvement after administration of Danazol. However, six days after a single application of cyclophosphamide (2,000 mg on day 1), a continuous increase of platelets to 100 G/l was obtained, a level which has remained stable for the last 18 months. CONCLUSION: The differential diagnosis of a hepatitis C associated autoimmune thrombocytopenia must be considered in patients with chronic hepatitis C infection and severe thrombocytopenia. In cases of refractory disease, treatment with cyclophosphamide may be successful. This is particularly appropriate if, in the case of an insufficient increase in thrombocytes following administration of immunoglobulins, the success of a splenectomy is improbable, or when a splenectomy must be excluded because of relative or absolute contraindications. PMID- 10875149 TI - [The course and therapy of acute hepatitis C viral infection. Is a prevention of its becoming chronic possible?]. AB - Due to the large number of patients chronically infected with hepatitis C virus and not responding to combination therapy with interferon-alfa 2 and ribavirin new therapeutic regimens are required. Early treatment of the viral infection might improve the response, as seen in treatment of HIV infection, thereby preventing progression to chronicity. The article reviews the natural course of an acute HCV infection after different modes of transmission like i.v.-drug abuse, transfusion, needle stick injury and blood products. As there are no good animal models for HCV infection, models of an acute infection with other noncytopathic viruses might improve our understanding of the mechanisms of viral clearance. Results from an acute infection of mice with the lymphocytic chorionmeningitis virus are demonstrating the development of a T-cell tolerance by anergy or deletion of virus specific T-cells as possible mechanisms for the failure of the immune system to clear the virus. These findings are compared to the results of CD4+ and CD8+ T-cell responses in patients with acute HCV infection. Several clinical trials have demonstrated a benefit of an early treatment of HCV infection. Although the natural course of acute HCV is changing during the last few years, even recent trials indicate that progression to chronicity might be prevented by early therapy. The studies show that therapy could be improved by daily dosing, higher single doses of interferon compared and prolongation of therapy up to six month. As most patients with acute HCV infection are rather seen in an outpatient practise than in hospitals cases of acute infections should be collected and treatment protocols be standardized to confirm these results in prospective trials. First results in 21 patients show that viral clearance under therapy was achievable in all of the patients. PMID- 10875150 TI - [Eosinophilic esophagitis: a largely unknown entity?]. PMID- 10875151 TI - [Concerning: a report with commentary: Endoscopic or surgical treatment of recurrent bleeding ulcer. By H. Messmann and A. Holstege. Z Gastroenterol 1999; 37: 1125-7]. PMID- 10875152 TI - [Vertical nystagmus induced by the head-shaking test: 10 cases]. AB - In cases of central and/or peripheral vestibular system asymmetry, Head-Shaking test-induced Nystagmus (H.S.-Ny) can appear after a cycle of 20 horizontal head oscillations. Four types of H.S.-Ny have been described, all of which are horizontal: 1) deficit Ny; 2) recovery Ny; 3) biphasic Ny; 4) triphasic Ny. None of these forms are specific for any given vestibular asymmetry site, whether central or peripheral. The authors report another low vertical type of H.S.-Ny found in 13 of the 1500 cases where the test was performed. Ten of these cases are discussed here. Vertical H.S.-Ny was found in 7 cases of N.M.R.-confirmed central pathologies, in 1 case of complications from epidemic parotitis and in 2 cases for which N.M.R. did not confirm the presence of an organic pathology. Vertical H.S.-Ny was often associated with other vestibular signs (Gaze-Ny, Rebound Ny, variable direction Ny, down-beat positional Ny, labyrinthine hyper reflexia). The authors consider this form of nystagmus a simple, easily determined signal of great importance in diagnosing the presence of a central, tronco-encephalic and cerebellar pathologies. PMID- 10875153 TI - [Surgical revision of the lateral nasal wall]. AB - The purpose of rhinosinus revision through endoscopic surgery is to resolve recurrent, residual pathologies in patients with drug-resistant symptoms and to prevent ocular and endocranial complications. Surgical revision proves quite complex because of the destruction of the anatomical structures. More over the presence of tougher scare tissue which bleeds easily increases the risk of iatrogenous complications. Axial and coronal-projection tomography without contrast medium must be performed prior to surgery. Endonasal orientation is ensured by six main reference points. Three of these points--nasal septum, upper edge of the choana and upper edge of the nasolacrymal duct--are nearly always present while the others may or may not be, depending on what sort surgery has been performed. The present work gives the guidelines for a correct approach to patients who have undergone many previous procedures. PMID- 10875154 TI - [Orbital decompression in Grave's disease: comparison of techniques]. AB - Grave's ophthalmopathy is an inflammatory, autoimmune disorder often associated with Grave's disease. The inflammatory infiltration involves the retrobulbar fatty tissue and the extrinsic eye muscles, causing proptosis, extraocular muscle dysfunction and often diplopia. Orbital decompression is an effective treatment in such cases, particularly when resistant to drugs and external radiation therapy. This work compares the results of orbital decompression performed by removing: a) the medial and lateral walls (Mourits technique) in 10 patients (19 orbits) and b) the medial and lower walls (Walsh-Ogura technique) in 17 patients (31 orbits). The results show that removing the floor of the orbit enables better reduction of proptosis but more easily leads to post-operative diplopia. Thus it proves necessary to combine the two techniques, modifying the surgical approach on a case-by-case basis. PMID- 10875155 TI - [Multi-center study of recurrent nasal sinus polyposis: prognostic factors and possibility of prophylaxis]. AB - Today, surgery is the treatment of choice for nasal sinus polyposis. Nevertheless, although meticulous surgery does "per se" reduce the percentage of recurrences, there are cases where even the most painstaking removal of the entire pathology cannot prevent recurrence. Therefore recurrences do not appear linked to the type of surgery; rather onset appears linked to intrinsic, only partially recognizable factors responsible for the primary and secondary polypogenesis. In order to identify negative prognostic factors which might be implicated in recurrences, the present study extrapolated the data from forms on 181 patients who had undergone surgery for nasal sinus polyposis and subjected it to multivariance analysis. These patients were recruited during the course of a multicenter study with the participation of 12 ENT Centers in Piemont and Liguria. The recurrence rate was 13%. In analyzing unfavorable factors prognosticating recurrence, thirteen parameters were examined. Nine of these (age, sex, severe deviation of the septum causing restriction, severe turbinate hypertrophy, surgery or repeat surgery for recurrence, type of macro-micro endoscopic surgery, allergy to seasonal inhalants, allergy to perennial inhalants, mixed allergies) did not prove to have any significance in recurrences. The presence of bilateral involvement of the sinus system presented a negative trend as regards recurrences while involvement of more than one subsite (anterior ethmoid, posterior ethmoid, maxillary sinus, sphenoid), ASA and NSAID intolerance and abundant eosinophilic infiltration in the mucous chorion proved statistically significant (p < 0.05 for all three parameters) for recurrence. Post-operative topic prophylactic treatment with steroids (beclomethasone) or anti-H1 drugs (azelastin, HCl) did not appear to affect the onset of recurrence although it did have a positive effect on subjective symptoms. PMID- 10875156 TI - [Evaluation of resection margins as a prognostic factor in the surgical treatment of laryngeal carcinoma]. AB - A series of 564 patients who underwent conservative or radical surgery for laryngeal carcinoma were studied in a postoperative follow-up. In particular the onset of local recurrences was studied in relation to the histopathological typing of the resection margin. Histology was performed both on the surgical sample and performing an additional biopsy on the surgical margin. The incidence of local recurrences--in relation to the positive or negative tumor infiltration at the resection margins of the surgical sample--was analogous throughout the entire case study (11.1% vs. 11.7%). Only as light increase in the incidence of recurrence was found in those patients with partial laryngectomy (12% vs. 8.9%) although this was not statistically significant. On the contrary, when the marginal biopsy tested positive for neoplasm, it was clearly and significantly related to a higher incidence of T recurrences. This held true for all cases as a whole (36.4% vs. 10.1%, p < 0.04) and, in particular for those treated with partial laryngectomy (46.2% vs. 3%, p < 0.02). On the basis of these observations it can be concluded that, where possible, additional biopsy should be performed to integrate the traditional evaluation of the margins of the surgical piece in order to gain greater prognostic reliability, particularly in cases of partial surgery. From the therapeutic point of view, the authors assert that scheduling supplementary radiation therapy and shorter intervals between follow-up examinations is timely if the resection margin tests positive. PMID- 10875157 TI - [The role of p53 tumor suppressor gene as prognostic factor in laryngeal squamous cell carcinoma]. AB - Mutations in the p53 gene--which codifies anuclear phosphoprotein that acts as a tumor suppressor gene--is the most common genetic alteration in head and neck cancers. The aim of the present study was to investigate the prognostic significance of p53 protein over expression in squamous cell laryngeal carcinoma. To do so we analyzed 31 patients affected by precancerous lesions of the larynx who had undergone multiple biopsy between 1980 and 1995. Twenty-five of these patients later developed laryngeal carcinoma. In this group of patients, 51 biopsies were performed for precancerous lesions (17 hyperplasia, 3 light dysplasia, 23 moderate dysplasia, 8 severe dysplasia) prior to evidence of laryngeal cancer (2.04 biopsies/patient). In the group of patients who did not develop laryngeal cancer, 18 biopsy were performed (2.2 biopsies/patient) and histology revealed: 5 keratosis, 5 light dysplasia, 4 moderate dysplasia and 4 grave dysplasia. Using the immunohistopathological staining technique, 69 formalin-fixed, paraffin-embedded precancerous samples and 25 laryngeal carcinomas were examined for p53 over expression. The monoclonal antibody Pab 1801 was used with the avidinbiotin immunoperoxidase technique; p53 intensity of expression was assessed and correlated with clinical-pathological parameters. Over expression of the p53 protein was found in 56.8% of the precancerous lesions (41% of the hyperplastic lesions, 66% of light dysplastic lesions, 60% of moderate dysplastic lesions and 75% of severe dysplastic lesions) in the group patients who did develop laryngeal cancer and in 22.2% of the precancerous lesions in the group of patients that did not. The transformed lesions showed a strong correlation between intensity of positivity and grade of cellular atypia. Further in 93.3% of the patients with p53 positive precancerous lesions which later developed into laryngeal cancer, p53 over expression was present in the cancerous lesions. There was no significant correlation between p53 immuno reactivity and such clinico pathological tumor parameters as TNM staging and tumorrecurrence. On the other hand, there was a correlation between p53 overexpression and differentiation grading: p53 overexpression was found in 75% of the poorly differentiated tumors, 58.3% of moderately differentiated and 44.4% of well differentiated tumors. The fact that p53 is detected in preneoplastic lesions suggests that p53 gene alteration takes place very early in laryngeal carcinoma and moderate-to-high p53 expression constitutes a high risk of transformation into cancer; on the other hand low expression may reflect reversible changes that can be attributed to the genotoxic effects of tobacco smoking. In conclusion the present data suggest that p53 over expression could be a good prognostic marker in predicting which precancerous laryngeal lesions will progress into cancer. PMID- 10875158 TI - [Oropharyngeal angiolipoma: a case study]. AB - Angiolipoma is a histological variation of lipoma. It occurs in 17% of the cases of lipoma and the cervico-facial localization is quite rate. Indeed, in the literature 17 cases of angiolipoma have been presented in the head and neck region and none in the oropharygeal area. The present work reports a case of pedunculate angiolipoma in a 44-year-old male: the red-violaceous growth resting on the upper surface of the tongue--was 13 cm long and 1 cm in diameter. The implantation base corresponded to the left posterior-lateral wall of the oropharnyx, 1 cm below the lower tonsilar pole. A serreneoud loop was used to remove the angiolipoma in direct view, the patients mouth held open with an autostatic gag. Histologically it was a non infiltrating variant for which simple removal is curative and recurrences are rare. Viceversa, removal of the infiltrating type requires expanding there section edges to include surrounding tissues in an attempt to preventre currences which are quite frequent (occurring in approximately 50% of the cases). PMID- 10875160 TI - [Cryopreservation. State of the art]. AB - Cryopreservation was mainly introduced in order to face the increasing number of stimulation cycles per patient and multiple pregnancy during procedures of in vitro fertilization and embryo transfer, accumulating excess embryos and transferring it in a later cycle. Nevertheless embryo storage has several implications because of moral and legal problems. Starting from 1984, date of the first transfer of a cryopreserved embryo, techniques have evolved and cryopreservation is now a reality in many sterility Centers. The aim of this review is then to point out the state of the art of cryopreservation with regard to embryos and oocyte storage and to have a look to possible futures developments in this technique. PMID- 10875159 TI - [Management of infertility following treatment of testicular tumors]. AB - Testis cancer mainly affects patients in a reproductive age. Since at the present time recovery can be achieved in 84% of cases within ten years, there has been an increasing demand for paternity by these patients. In most cases, the preventive cryopreservation of spermatozoa and their subsequent use for assisted reproductive techniques can solve the problems connected with secretory infertility (no always reversible) caused by chemotherapy or radiotherapeutic treatment. Cryopreservation also allows to avoid the surgical taking of germinal cells either from testes or from seminal ducts in cases of anajeculation due to retroperineal lymphoadenectomy (present in 23.5-80% of cases). Failing the preventive cryopreservation, the surgical or microsurgical extraction of testis spermatozoa can be performed with success rates equal to 50%. Their use for intracytoplasmic sperm injection (ICSI) procedures presently seems to provide some encouraging results both for fertilization and pregnancy rates. PMID- 10875161 TI - Comparison of single preoperative oral rufloxacin versus perioperative ciprofloxacin as prophylactic agents in transurethral surgery. AB - The aim of this prospective clinical trial was to compare the efficacy of rufloxacin, a once-daily fluoroquinolone administered as a single pre-operative dose versus the perioperative prophylaxis with ciprofloxacin in transurethral surgery. Two hundred and two patients undergoing transurethral resection of bladder tumors (132) or transurethral resection of the prostate (70) were selected for the study between January 1997 and June 1998. Patients were randomized to two treatment groups. Group A received a single oral 200 mg dose of rufloxacin three hours before surgery and group B was administered oral ciprofloxacin 250 mg bid until catheter removal. The two treatment groups were homogeneous with respect to patient characteristics. One hundred and seventy three patients (89 rufloxacin and 84 ciprofloxacin) were assessed and 29 were excluded from the statistical analysis. The incidence of postoperative infection was similar in both treatment groups (5.7% rufloxacin, 4.7% ciprofloxacin). On the other hand, single-dose pre-operative prophylaxis with rufloxacin significantly reduced the cost of antibiotic prophylaxis. Results of the present study show that single dose oral rufloxacin may be used in routine clinical practice as a preoperative prophylactic antibiotic due to its low cost, its documented efficacy and its simple once daily dosage regimen. PMID- 10875162 TI - Bacterial prostatitis: urine and spermatic fluid culture. AB - To evaluate in a prospective study the sensitivity of urine and spermatic fluid cultures in identifying the presence of infection compared to Meares-Stamey's test (MSt) results. Fourty patients were diagnosed having bacterial prostatitis following MSt. They underwent both urine and spermatic fluid cultures after MSt results and immediately before antibiotic treatment. All the patients were asked which of the three examens was the least tolerable. Urine and spermatic fluid culture were negative in 36 and 4 cases respectively. Spermatic fluid culture identified infection in 36 out of 40 patients who underwent MSt (90%) and was more acceptable for the patients. Urine culture is a less accurate way of identifying the infective agent in prostatitis, compared to spermatic fluid culture. The latter procedure is similar to the MSt. PMID- 10875163 TI - Valsalva leak point pressure: how to chose the best method. AB - Urinary continence is ensured as long as the urethral closure pressure remains greater than the intravesical pressure, in the presence of adequate support to the bladder and the proximal urethra. In order to select the appropriate surgical treatment, a correct diagnosis must be made; recently urodynamic evaluation has assumed a central role. In particular, the introduction of a new urodynamic parameter, the Valsalva Leak Point Pressure (VLPP), has provided new impetus to research in this area, even if different technical approaches have limited universal acceptance. The aim of the work is to describe the reasons why the authors have been led to prefer double measurement, both at partial filling and at maximum cystometric capacity, considering that this does not involve any additional economic burden. The authors underline the promising potential of the method, once standardized, in the urodynamic evaluation of urinary incontinence. PMID- 10875164 TI - [Hyperhydration with low mineral Rocchetta water after extracorporeal lithotripsy]. AB - Both prophylaxis and stone-free status after ESWL are most important goals in treating urinary stone disease, because his high social cost. In order to this situation, we matched two homogeneous groups of patients that underwent ESWL because renal stones: during a one year follow-up with several US controls, daily 1.5 litres of low mineral content water was drank by I group patients; vice versa, daily 3 litres (1st ten days) and afterwards 2 litres of Rocchetta low mineral content water was drank by II group patients. This last kind of approach led to a significant improvement in stone fragments elimination time, in inferior calix stone cure and in stone recurrences rate. So we conclude that hyperhydration using right low mineral content water, is a simple and cheap way to improve both treatment and prophylaxis of urinary stones. PMID- 10875165 TI - [Sarcomatoid carcinoma of the kidney associated with urothelial carcinoma: report of a case with unusual clinical presentation]. AB - We report a case of sarcomatoid renal cell carcinoma and simultaneous transitional cell carcinoma of the renal pelvis in a 77 year old man admitted for uroseptic fever persisting for two months. Seven years earlier he underwent cystectomy with ureterosigmoidostomy for transitional cell carcinoma of the bladder. CT scan described a severe hydronephrosis with dilated pelvis, several pseudocystic formations with renal parenchima thinning and absence of contrast excretion. Radical nephrectomy was performed consequent to a clinical diagnosis of uroseptic fever in secondary hydronephrosis due to stenosis of ureterosigmoidostomy. Tumors were suspected on cut section and confirmed by histological examination. PMID- 10875166 TI - Tell me, what did you see? The stimulus on computers. AB - Most psychology experiments start with a stimulus, and, for an increasing number of studies, the stimulus is presented on a computer monitor. Usually, that monitor is a CRT, although other technologies are becoming available. The monitor is a sampling device; the sampling occurs in four dimensions: spatial, temporal, luminance, and chromatic. This paper reviews some of the important issues in each of these sampling dimensions and gives some recommendations for how to use the monitor effectively to present the stimulus. In general, the position is taken that to understand what the stimulus actually is requires a clear specification of the physical properties of the stimulus, since the actual experience of the stimulus is determined both by the physical variables and by the psychophysical variables of how the stimulus is handled by our sensory systems. PMID- 10875167 TI - Evaluation of a Web-based introductory psychology course: I. Learning and satisfaction in on-line versus lecture courses. AB - We offered introductory psychology on the World-Wide Web (WWW) and evaluated the on-line format relative to the traditional lecture-test format, using a pretest posttest nonequivalent control group design. Multiple sections of the introductory course were offered each semester; on-line and lecture sections were taught by the same instructor, the same textbook was used, and the same in-class examinations were taken. For on-line sections, mastery quizzes, interactive individual exercises, and weekly laboratory meetings replaced lectures. Increased content knowledge was greater for the students in the Web sections, as was in class examination performance. Use of the WWW and computers for academic purposes increased more in the on-line sections, and the on-line students showed a greater decrease in computer anxiety. The students in the on-line sections expressed appreciation for course components and the convenience of the course, but the lecture sections received higher ratings on course evaluations than did the on line sections. Learning and course satisfaction were dissociated in the two course formats. PMID- 10875168 TI - Evaluation of a Web-based introductory psychology course: II. Contingency management to increase use of on-line study aids. AB - In a Web-based general psychology course, students were observed to postpone use of on-line study aids until 2 days prior to examinations, thus negating any influence of advance organizers (Taraban, Maki, & Rynearson, 1999). We attempted to modify this behavior by providing course credit in the form of short quizzes as rewards for using on-line study aids to preview each chapter. Some students received quizzes after previewing frequently asked questions (FAQ); other students received quizzes after previewing chapter outlines. Students who received quizzes for previewing FAQ pages accessed those pages more frequently than did students who received quizzes for previewing outline pages. Increased access to FAQs was associated with higher scores on FAQ-related midterm examination questions. However, the advantage on examination items was not apparent on a cumulative final examination. Navigational structures and reward values need to be considered when one is managing contingencies in Web courses. PMID- 10875170 TI - Quiz-o-matic: a free Web-based tool for construction of self-scoring on-line quizzes. AB - Increasingly, educators are using the World-Wide Web (the Web) and related technology to supplement their traditional teaching materials. In this paper, we present a free tool that has been placed on the Web to enable educators to create Web pages that present self-scoring quizzes. Such Web-based practice quizzes provide students with an opportunity for interactive assessment and review of course material. The tool, Quiz-o-Matic, uses active server pages to build the HTML and JavaScript code for each quiz, without requiring the user to edit any code. We present evidence that students judge the self-scoring quizzes to be useful and worthwhile additions to a course. PMID- 10875169 TI - Using cognitive learning theory to design effective on-line statistics tutorials. AB - Careful attention to principles of learning can improve the design of Web-based lessons and tutorials. Tutorials from the Web Interface for Statistics Education (WISE; http:?wise.cgu.edu) demonstrate how specific principles can be integrated into Web design to enhance learning in two areas. First, the impact of students' poor self-regulation abilities on Web-based learning is considered. Second, evidence that specific types of visual presentations improve learning is discussed. Finally, the need for empirical evaluation is emphasized. Specific research and examples from the WISE project are used to illustrate each of these points. PMID- 10875171 TI - QUAID: a questionnaire evaluation aid for survey methodologists. AB - QUAID (question-understanding aid) is a software tool that assists survey methodologists, social scientists, and designers of questionnaires in improving the wording, syntax, and semantics of questions. The tool identifies potential problems that respondents might have in comprehending the meaning of questions on questionnaires. These problems can be scrutinized by researchers when they revise questions to improve question comprehension and, thereby, enhance the reliability and validity of answers. QUAID was designed to identify nine classes of problems, but only five of these problems are addressed in this article: unfamiliar technical term, vague or imprecise relative term, vague or ambiguous noun phrase, complex syntax, and working memory overload. We compared the output of QUAID with ratings of language experts who evaluated a corpus of questions on the five classes of problems. The corpus consisted of 505 questions on 11 surveys developed by the U.S. Census Bureau. Analyses of hit rates, false alarm rates, d' scores, recall scores, and precision scores revealed that QUAID was able to identify these five problems with questions, although improvements in QUAID's performance are anticipated in future research and development. PMID- 10875172 TI - OBSERVE: a multimedia course on the observational analysis of behavior. AB - OBSERVE is a preliminary release of a multimedia course for teaching undergraduate and graduate students how and why to study behavior by direct observation. The instructional text and commentary and the self-test and examination materials are built around a series of exercises in which the student observes and categorizes film clips of the behavior of several different species in several different ways. Incorporation of elements from The Observer software for computer recording and video analysis implements fully computerized continuous recording. At present, the text, together with check sheets that the program generates, enables a comparison between one-zero, instantaneous, and continuous sampling of the same behavioral excerpts. Matrices are printed out for an exercise in calculating interobserver reliability. Another section supports carrying out and writing up a small observational project on human behavior in the field. Plans for the future development of OBSERVE are briefly described. PMID- 10875173 TI - Regression analysis of correlated binary outcomes. AB - The purpose of this paper is to describe and illustrate a regression approach to the analysis of correlated binary outcomes (Liang & Zeger, 1986). Ignoring the correlations between repeated observations can lead to invalid inferences. This approach extends logistic regression to account for repeated observations in each of a series of individuals. In this paper, I present a nontechnical introduction to the generalized estimating equations (GEE) approach. A fictitious example is used to demonstrate that GEE regression correctly adjusts for the correlations between repeated binary observations. The approach is illustrated with an analysis of safer sex practices among high-risk teenagers. PMID- 10875174 TI - A computational approach to modeling population differences. AB - Four experiments were conducted to determine whether the Hyperspace Analogue to Language (HAL) model of semantic memory could differentiate between two different populations. An analysis of the differences in densities (or average distances between word neighbors in semantic space) in HAL matrices--generated from text corpora derived from younger and older adults--confirmed that HAL was able to distinguish between the two age groups. This difference was again detected when structured interview data were used to build the corpora. A third experiment, designed to test the specificity of HAL in detecting differences between groups, did not detect any difference in the densities of the memory representations when older adults generated both the test corpora. The final experiment, conducted on the language of adults with Alzheimer's and normal adults, again demonstrated that HAL could discriminate between the two populations. These results suggest that HAL is capable of modeling, on the basis of changes in mean density, some of the differences between populations without modifying the model itself but, rather, by changing the text corpus from which the model creates its representations in semantic space. PMID- 10875175 TI - Detecting nonlinearity in psychological data: techniques and applications. AB - Modern graphical and computational techniques for detecting nonlinearity in psychological data sets are presented. These procedures allow researchers to determine the information complexity of temporal data, using physiological and psychological measurements, and to provide evidence for chaos in time series contaminated by measurement noise. Problems with noise reduction and appropriate experimental control, using surrogate time series, are discussed, and applications of the technology are illustrated, using response time, handwriting, and typing data sets. In an experimental application of appropriate nonlinear analysis procedures, the results of a time series prediction experiment confirm that some subjects are sensitive to chaos. In contrast to previous attempts demonstrating sensitivity to chaos, the experiment reported here employs surrogate series to control for linear stochastic aspects of the stimulus sequences, such as autocorrelation. Recommendations for the selection of appropriate software for performing nonlinear analyses are presented, including a comprehensive list of World-Wide Web sites offering such software. PMID- 10875176 TI - MATLAB and graphical user interfaces: tools for experimental management. AB - MATLAB is a convenient platform for the development and management of psychological experiments because of its easy-to-use programming language, sophisticated graphics features, and statistics and optimization tools. Through implementation of the Brainard-Pelli Psychophysics Toolbox, the MATLAB user gains close temporal and spatial control over the CRT, while retaining the simplicity of an interpreted language conductive to rapid program development. MATLAB's abilities can be further utilized through easily programmable graphical user interfaces (GUIs). We illustrate how a GUI can serve as a powerful and intuitive tool for organizing and controlling all aspects of a psychological experiment, including design, data collection, data analysis, and theory fitting. PMID- 10875177 TI - Computerized games to study the development of attention in childhood. AB - Children enjoy playing games. We can take advantage of this in the designs of computerized tasks that will engage their interest. These designs also serve to advance the study of chronometric measures, such as manual and saccadic reaction times and event related potentials, with young children. The goals of our method development are (1) to allow for comparable tasks across a wide variety of ages, (2) to make possible comparisons of child performance with data gathered in adult cognitive studies, and (3) to help to support inferences about the development of underlying mechanisms. We have designed a battery of computerized tasks in order to study the development of attention functions of alertness, orienting, and executive control during childhood. Our purpose is to describe each of these tasks in detail and present the results that have been obtained so far. The battery was tested using a sample of 5-year-old children as subjects. PMID- 10875178 TI - EPA2000: a multilingual, programmable computer assessment of off-line metacognition in children with mathematical-learning disabilities. AB - EPA2000 is a program for the assessment of off-line measured metacognitive skills and arithmetical performances in primary school children with mathematical learning disabilities. The program is designed as a script engine. The concept makes it possible to modify and translate the test into different languages without reprogramming. A user-friendly script editor is built-in, with which all of the parameters of the test can be modified and translated in different languages. PMID- 10875179 TI - An unobtrusive measurement method of the horizontal gaze angle. AB - A horizontal gaze angle measurement device is introduced. By combining a photoelectric viewing device to measure the horizontal eye angle with a similar head angle measurement device, it is possible to measure the horizontal gaze angle without using a headrest. After discussion of circuit diagrams and measurement principles, it is shown that the measurements made with the device yield a reasonable precision. The mean absolute measurement error is below 1 degree. This inexpensive and unobtrusive device covers a visual field of about 20 degrees and can be used in parallel with many tasks. Further, data on the successful application of the device in a driving simulation setting are discussed. PMID- 10875180 TI - The electronic mood device: design, construction, and application. AB - The electronic mood device (EMD) is designed to help answer questions about the variability and dynamics of emotions. It is a small, portable instrument used for repeated recording of moods and feelings. Both construction and operation of the EMD are described. The EMD can best be conceived of as an electronic mood adjective checklist. Persons using the EMD are signaled at designated (e.g., hourly) or random intervals to register their mood or feelings. Paper and pencil are not required. An application is given. It shows how feelings vary within and between persons and during the day. It is concluded that the EMD offers several advantages over paper-and-pencil instruments. Retrospective use is impossible. Timing and registration are accurate. Data handling is fast. Potential future applications are suggested. PMID- 10875181 TI - Evolving faces from principal components. AB - A system that uses an underlying genetic algorithm to evolve faces in response to user selection is described. The descriptions of faces used by the system are derived from a statistical analysis of a set of faces. The faces used for generation are transformed to an average shape by defining locations around each face and morphing. The shape-free images and shape vectors are then separately subjected to principal components analysis. Novel faces are generated by recombining the image components (eigenfaces) and then morphing their shape according to the principal components of the shape vectors (eigenshapes). The prototype system indicates that such a statistical analysis of a set of faces can produce plausible, randomly generated photographic images. PMID- 10875182 TI - A laser-based technique for measuring accuracy and distortion in judgments of linear dimensions. AB - A new technique is described that permits precise measurement of accuracy and distortion in judgments of linear dimensions based on either perception or memory. This technique involves the use of a single laser beam and a reference line placed on a projection surface. By rotating a laser device, the distance between the reference line and the point created by the beam may be continuously varied. This procedure avoids unintentional distortion from misjudgment of standard metrics, while the semicircular movement required by this technique eliminates body-referenced estimation and some other potential confounds. Potential applications to research in visual perception, spatial memory, and body image are discussed. PMID- 10875183 TI - SurveyWiz and factorWiz: JavaScript Web pages that make HTML forms for research on the Internet. AB - SurveyWiz and factorWiz are Web pages that act as wizards to create HTML forms that enable one to collect data via the Web. SurveyWiz allows the user to enter survey questions or personality test items with a mixture of text boxes and scales of radio buttons. One can add demographic questions of age, sex, education, and nationality with the push of a button. FactorWiz creates the HTML for within-subjects, two-factor designs as large as 9 x 9, or higher order factorial designs up to 81 cells. The user enters levels of the row and column factors, which can be text, images, or other multimedia. FactorWiz generates the stimulus combinations, randomizes their order, and creates the page. In both programs HTML is displayed in a window, and the user copies it to a text editor to save it. When uploaded to a Web server and supported by a CGI script, the created Web pages allow data to be collected, coded, and saved on the server. These programs are intended to assist researchers and students in quickly creating studies that can be administered via the Web. PMID- 10875184 TI - Statistical power for the two-factor repeated measures ANOVA. AB - Determining a priori power for univariate repeated measures (RM) ANOVA designs with two or more within-subjects factors that have different correlational patterns between the factors is currently difficult due to the unavailability of accurate methods to estimate the error variances used in power calculations. The main objective of this study was to determine the effect of the correlation between the levels in one RM factor on the power of the other RM factor. Monte Carlo simulation procedures were used to estimate power for the A, B, and AB tests of a 2 x 3, a 2 x 6, a 2 x 9, a 3 x 3, a 3 x 6, and a 3 x 9 design under varying experimental conditions of effect size (small, medium, and large), average correlation (.4 and .8), alpha (.01 and .05), and sample size (n = 5, 10, 15, 20, 25, and 30). Results indicated that the greater the magnitude of the differences between the average correlation among the levels of Factor A and the average correlation in the AB matrix, the lower the power for Factor B (and vice versa). Equations for estimating the error variance of each test of the two-way model were constructed by examining power and mean square error trends across different correlation matrices. Support for the accuracy of these formulae is given, thus allowing for direct analytic power calculations in future studies. PMID- 10875185 TI - Standardization and decomposition of rates: useful analytic techniques for behavior and health studies. AB - Standardization and decomposition are widely used analytic techniques in population studies for adjusting the impact of compositional factors on rates. This study demonstrates the application of these methods to behavior and health studies. Bootstrapping is used to estimate standard errors of the component effects and to conduct significance tests for them. The authors have developed a Windows-based computer program that is demonstrated in the study for standardization and decomposition analysis by using empirical data on HIV seropositivity rates in two injection-drug-using populations in the northeastern United States. PMID- 10875186 TI - RegRand: statistical software for the multiple-baseline design. AB - RegRand (Version 1.0) is Macintosh-based software that enables a researcher to conduct a non-parametric statistical analysis of the data from Koehler and Levin's (1998) recently described regulated randomization single-case multiple baseline design. Regulated randomization design and analysis principles are reviewed in relation to an educational research application and a step-by-step illustration of them in relation to the RegRand program is presented. PMID- 10875187 TI - UCS expectancy biases in spider phobics: underestimation of aversive consequences following fear-irrelevant stimuli. AB - This paper reports the results of two studies investigating judgements made by spider phobics about the potential threatening consequences (unconditioned stimulus, UCS, expectancies) associated with their phobic stimulus, fear-relevant (FR) stimuli, and fear-irrelevant (FI) stimuli. Using a 'thought experiment' UCS expectancy paradigm, the studies reported found that (1) spider phobics reported significantly higher UCS expectancies to spider stimuli than nonphobics, (2) spider phobics consistently underestimated the probability of aversive consequences following FI stimuli and (3) this underestimation of UCS expectancies to FI stimuli in phobics was not the result of a contrast effect resulting from sequential FR and FI judgements. This differential effect may have important implications for the kind of mechanism which mediates judgements about phobic consequences. These findings suggest that the dimensions on which phobic stimuli are categorised may be 'stretched' in the case of phobics and that this gives rise to the comparative underestimation of threat associated with FI stimuli but also makes phobics more vulnerable to acquiring other phobias. PMID- 10875188 TI - Prediction and control: operational definitions for the experimental analysis of anxiety. AB - Prediction and/or control of threatening events generally results in less pronounced anxiety-related responding compared to when those same events are unpredictable or uncontrollable. For this reason, researchers have suggested that predictability and controllability may modulate anxiety-related responding, thereby serving an important role in the development and progression of anxiety pathology. Despite the recognized importance of prediction and control for anxiety, these variables have not been defined or operationalized in a uniform and unambiguous manner. In this article, we propose an operational definition that defines and distinguishes prediction and control in terms of the onset and offset of an aversive event. This operationalization is aimed at facilitating experimental-based efforts to explore the independent and interactive effects of the prediction and control on anxious responding. PMID- 10875189 TI - Relaxation therapy for insomnia: nighttime and day time effects. AB - We compared day time functioning in college students with and without insomnia and explored changes in day time functioning after progressive relaxation (PR) treatment for insomnia. Students with insomnia (SWI; n = 57) were compared to a control group of students not complaining of insomnia (SNI; n = 61) on self reported sleep variables and five questionnaires: Insomnia Impact Scale (IIS), Dysfunctional Beliefs and Attitudes About Sleep Scale (DBAS), Epworth Sleepiness Scale (ESS), Fatigue Severity Scale (FSS), and Penn State Worry Questionnaire (PSWQ). SWI demonstrated significant impairment on all day time functioning and sleep measures compared to SNI. To investigate treatment effects on day time functioning, 28 SWI were randomly assigned to PR. Treated SWI were compared to untreated SWI and SNI at posttreatment. Treated participants improved sleep in comparison to untreated SWI, but failed to show significant improvements in day time functioning. Insomnia treatments focused on improving sleep may not improve day time functioning, or day time gains may emerge more slowly than sleep gains. This study documents the wide range of day time functioning complaints in young adults with insomnia and suggests that the goal of insomnia treatment should be to not only improve sleep but also to improve the subjective experience of day time functioning. PMID- 10875190 TI - Intrusive thoughts and their relationship to actigraphic measurement of sleep: towards a cognitive model of insomnia. AB - Although cognitive over-arousal has been hypothesised as a causal factor in sleep onset insomnia, relatively little is known about the specific pre-sleep intrusions which delay sleep. To investigate this relationship adequately 'live', verifiable, unobtrusive and independent monitoring of thought process and sleep pattern is essential. This study was designed with these requirements in mind. Voice-activated audiotape recordings of spontaneous thoughts, and actigraphic data from which to estimate sleep parameters, were obtained over three consecutive nights from 21 participants (63 subject nights). Content analysis of transcribed audiotapes yielded eight categories of pre-sleep intrusion. Results from correlational and regression analyses indicate that thinking about sleep and the anticipated consequences of poor sleep, along with general problem-solving are the strongest predictors of objective sleep latency. Principal Components Analysis suggests that intrusions can be subsumed under one of three factors: 'active problem-solving', 'present state monitoring' and 'environmental reactivity'. Implications for cognitive models and treatments of insomnia are discussed. PMID- 10875191 TI - Neurological and neuropsychological signs in obsessive compulsive disorder: interaction with behavioural treatment. AB - The main aim of this study was to test the hypothesis that neurological soft signs and neuropsychological abnormalities associated with obsessive-compulsive disorder (OCD) predict poor response to behavioural treatment. The design permitted investigation of secondary hypotheses, regarding correlations among these neurological markers and levels of symptomatology, and their stability in relation to changes in levels of symptomatology. Thirty-five participants satisfying DSM-IV diagnostic criteria for OCD were assessed pre- and postbehavioural treatment using a scaled measure of symptom severity, and a battery of tests sensitive to neuropsychological deficits associated with OCD. Eighteen of the participants were also assessed on an inventory of neurological soft signs. Neither neuropsychological test deficits nor neurological soft signs pretreatment predicted response to behavioural treatment. Lower performance on neuropsychological tasks and symptom severity were both significantly correlated with levels of soft signs. Some neurological markers were less severe posttreatment, but these changes were not related to treatment response. PMID- 10875192 TI - The generality of cognitive bias in acute stress disorder. AB - Cognitive bias was investigated in acutely traumatised civilians with either acute stress disorder (ASD; n = 26) or no ASD (n = 24). Participants completed the Acute Stress Disorder Interview, the Beck Depression Inventory, the Beck Anxiety Inventory (BAI), the Impact of Event Scale (IES), and an Event Probability Questionnaire and an Event Cost Questionnaire that comprised items pertaining to (a) external harm, (b) somatic sensations and (c) social events. ASD participants exaggerated both the probability of negative external harm, somatic and social events occurring, and the adverse cost of those events more than non-ASD participants. Elevated probability and cost estimates were predicted by BAI and IES-Avoidance scores, respectively. These findings are discussed in the context of different patterns observed in other anxiety disorders, and interpreted in terms of network theories of posttraumatic stress. PMID- 10875193 TI - Self-focused attention before and after treatment of social phobia. AB - It has been hypothesized that effective psychological treatment for social phobia changes the person's representation of the self in a more positive direction. In order to test this hypothesis, we analyzed 506 thoughts that were endorsed by 23 social phobic individuals while anticipating socially stressful situations before and after exposure therapy. Treatment efficacy was assessed with the Social Phobia and Anxiety Inventory (SPAI) [Turner, S. M., Beidel, D. C., Dancu, C. V., & Stanley M. A. (1989) An empirically derived inventory to measure social fears and anxiety: the Social Phobia and Anxiety Inventory. Psychological Assessment, 1, 35-40)]. Subjects endorsed significantly fewer negative self-focused thoughts after treatment (on average 8.7% of the thoughts) than before treatment (26.5%, p < 0.005). These changes were highly correlated with pre-post difference scores in the social phobia subscale of the SPAI (r = 0.74, p < 0.0001). Implications of the results for the cognitive model of social phobia will be discussed. PMID- 10875194 TI - Self-appraised social problem solving abilities, emotional reactions and actual problem solving performance. AB - Self-report measures of social problem solving abilities have yet to be associated with objective problem solving performance in any consistent manner. In the present study, we investigated the relation of social problem solving abilities--as measured by the Social Problem Solving Skills Inventory--Revised (SPSI-R [Maydeu-Olivares, A. & D'Zurilla, T. J. (1996). A factor analytic study of the Social Problem Solving Inventory: an integration of theory and data. Cognitive Therapy and Research, 20, 115-133])--to performance on a structured problem solving task. Unlike previous studies, we examined the relation of problem solving skills to performance curves observed in repeated trials, while controlling for affective reactions to each trial. Using hierarchical modeling techniques, a negative problem orientation was significantly predictive of performance and this effect was not mediated by negative affectivity. Results are discussed as they pertain to contemporary models of social problem solving. PMID- 10875195 TI - Combination effect of photodynamic and sonodynamic therapy on experimental skin squamous cell carcinoma in C3H/HeN mice. AB - We studied a combination of photodynamic therapy (PDT) and sonodynamic therapy (SDT) for improving tumoricidal effects in a transplantable mouse squamous cell carcinoma (SCC) model. Two sensitizers were utilized: the pheophorbide-a derivative PH-1126, which is a newly developed photosensitizer, and the gallium porphyrin analogue ATX-70, a commonly used sonosensitizer. Mice were injected with either PH-1126 or ATX-70 i.p. at doses of 5 or 10 mg/kg.bw. At 24 (ATX-70) or 36 hr (PH-1126) (time of optimum drug concentration in the tumor) after injection, SCCs underwent laser light irradiation (88 J/cm2 of 575 nm for ATX-70; 44J/cm2 of 650 nm for PH-1126) (PDT), ultrasound irradiation (0.51 W/cm2 at 1.0 MHz for 10 minutes) (SDT), or a combination of the two treatments. The combination of PDT and SDT using either PH-1126 or ATX-70 as a sensitizer resulted in significantly improved inhibition of tumor growth (92-98%) (additive effect) as compared to either single treatment (27-77%). The combination using PH 1126 resulted in 25% of the treated mice being tumor free at 20 days after treatment. Moreover, the median survival period (from irradiation to death) of PDT + SDT-treated mice (> 120 days) was significantly greater than that in single treatment groups (77-95 days). Histological changes revealed that combination therapy could induce tumor necrosis 2-3 times as deep as in either of the single modalities. The combination of PDT and SDT could be very useful for treatment of non-superficial or nodular tumors. PMID- 10875196 TI - E-cadherin expression in skin tumors using an AMeX immunohistostaining method. AB - The AMeX (acetone-methylbenzoate-xylene) method results in good preservation of tissue and morphological details, almost equivalent to that of routinely processed formalin-fixed and paraffin-embedded tissue specimens, and of antigenicity equivalent to that of fresh frozen tissue specimens. It has been reported that the expression of the cell-cell adhesion molecule E-cadherin is often decreased in some types of carcinomas. A decrease in E-cadherin expression is associated with the invasive or metastatic potential of tumor cells. We immunohistochemically examined the expression of E-cadherin with anti-E-cadherin monoclonal antibody in various skin tumors (25 basal cell carcinomas, 11 squamous cell carcinomas, 9 keratoacanthomas, and 11 Bowen's disease) using the AMeX method and found that this method preserved antigenicity well without pretreatment. E-cadherin expression was decreased in 18.2% of squamous cell carcinomas and 33.3% of keratoacanthomas. On the other hand, it was preserved in almost all Bowen's disease and basal cell carcinomas. From the results of our study, we suggest that Bowen's disease and basal cell carcinoma do not have much metastatic potential due to retention of high levels of E-cadherin expression. We hope to apply the AMeX method to other immunohistochemical examinations because this is a very useful staining method. PMID- 10875197 TI - Inhibitory effects of Alpinia speciosa K. SCHUM on the porphyrin photooxidative reaction. AB - It is thought that the beta-carotene defense mechanism against photosensitivity involves the inhibition of singlet oxygen formation, a kind of active oxygen. When we screened chemical substances obtained from plants indigenous to Okinawa, known to have residents with the longest life span in Japan, we found that Alpinia speciosa K. SCHUM (Japanese name: gettou), which is used as a food preservative, has an activity similar to that of beta-carotene. We measured the amount of lipid peroxide (LPO) formed from a hematoporphyrin-containing rat liver microsomal suspension irradiated with visible light. The inhibitory effect of Alpinia speciosa on LPO formation was confirmed when the addition of increasing concentrations of Alpinia speciosa extract led to a decrease in the amount of LPO formed. Moreover, the reaction mechanism that affects the amount of singlet oxygen formed was measured, and the effect of the extract was determined by the ESR trapping technique. It was found that the extract effectively inhibited the formation of singlet oxygen. The extract of Alpinia speciosa contains dihydro-5,6 dehydrokawain. It was confirmed that dihydro-5,6-dehydrokawain, which is a water soluble compound, has singlet oxygen quenching activity. We synthesized five derivatives of kawain and found that dimethyl [6-(2-phenylethyl)-2-oxo-2H-pyran-4 yl] phosphorothionate has the strongest singlet oxygen quenching activity. The use of the compound from Alpinia speciosa that exhibits singlet oxygen quenching activity as an inhibitory agent of the phototoxic reaction in porphyria is expected. PMID- 10875199 TI - Vitiligo skin types in Koreans. AB - There is uncertainty and controversy about the relationship between skin type and development of vitiligo. The present study was undertaken to study whether vitiligo patients have a different skin type than the control group. We investigated the skin types of 201 Korean vitiligo patients and 70 healthy Korean volunteers. Skin type was determined by the skin phototyping method proposed by Fitzpatrick. Compared to normal controls, skin type II was significantly less frequent and skin type V was quite common in the vitiligo group. These results suggest that people with dark skin have a higher probability of developing vitiligo than people with light skin. PMID- 10875198 TI - The bacteriology of acne vulgaris and antimicrobial susceptibility of Propionibacterium acnes and Staphylococcus epidermidis isolated from acne lesions. AB - We examined the species of bacteria aerobically and anaerobically isolated from 30 acne lesions and determined antimicrobial susceptibilities of Propionibacterium acnes (P. acnes) and Staphylococcus epidermidis (S. epidermidis) using nine antimicrobial agents. Among the bacteria isolated, S. epidermidis was most dominant. Both P. acnes and S. epidermidis were isolated from half of the acne lesions. The MIC of seven antimicrobials (ampicillin, erythromycin, roxithromycin, clindamycin, tetracycline, minocycline, nadifloxacin) against P. acnes was under 3.13 micrograms/ml. There were very few resistant strains of P. acnes, but many of S. epidermidis. More than 30% of the S. epidermidis isolates were resistant to erythromycin, roxithromycin, and clindamycin. After long-term systemic antibiotic therapy, the resistant strains of S. epidermidis increased, but P. acnes resistance was still limited. When we use antimicrobial agents for the treatment of acne, it should be noticed that not only P. acnes but also S. epidermidis in the acne lesions may acquire resistance to antimicrobials. PMID- 10875200 TI - A case of giant pencil-core granuloma. AB - Although pencil injury is a common occurrence, only six cases of so-called pencil core granuloma have been reported. All of these granulomas were relatively small, and most of them simulated malignant melanoma. We report a giant pencil-core granuloma that developed on a patient's right palm. Its gross appearance was quite different from that of other cases previously reported; it was much larger and resembled a hemangioma. PMID- 10875201 TI - Spitz nevus on the palmar surface. AB - Relatively little is known about the incidence of Spitz nevus on palmar surfaces. This report places a case study in the context of the Japanese literature regarding the occurrence of Spitz nevus on palmar surfaces. Although the proportion of palms and soles in relation to the body surface is about 5%, the incidence of the Spitz nevus was 2%. The mean age at onset was 17.8 years, and all 4 cases were women. The clinical features were a black macule or flatly elevated small modules. The size of the lesions was relatively small, extending from 3.5 mm to 8.0 mm. Although the backs of the hands and insteps have almost the same area as the palms and soles, the incidence of onset in these regions was 6.3% (13 cases). We thus concluded that Spitz nevus tends to be rare on palms and soles. PMID- 10875202 TI - A case of anaphylaxis due to ibuprofen. AB - A forty-four-year-old Japanese female, who had persistant rhinorrhea, was administered Benza block tablets orally along with two other medicines. Immediately after ingestion, the patient displayed itching of the right upper eyelid, followed by coughing, sneezing, nasal discharge, nasal obstruction, nausea, vomiting, swelling of the face, and dyspnea. She had edema, a wheal extending from the face to the neck, and swelling of the eyelids and lips. Her symptoms subsided after treatment. Her reaction to ibuprofen, which was contained in the Benza Block tablets, was confirmed by a positive reaction to prick testing. From the results of these examinations, our patient was diagnosed as having anaphylaxis due to the ibuprofen in the Benza Block tablets. A review of the literature revealed no previous reports of anaphylaxis due to ibuprofen, although a few cases of ibuprofen urticaria have been reported. PMID- 10875203 TI - Multiple fibroepithelial basal cell carcinoma of Pinkus associated with seborrheic keratosis in a nevoid distribution. AB - We describe a patient with multiple fibroepithelial basal-cell carcinoma (FEBCC) associated with seborrheic keratosis distributed in a neviform fashion on the left side of the body and clinically resembling skin tags. PMID- 10875204 TI - A rare association of systemic sclerosis with psoriasis vulgaris. AB - Three patients with systemic sclerosis (SSc) and psoriasis vulgaris were evaluated. The onset of psoriasis preceded by 3-7 years the onset of SSc in all cases. All the patients presented diffuse scleroderma, accompanied by lung and esophageal involvement in two cases. Sjogren's syndrome occurred in one case. Myalgia developed with the onset of SSc in two patients. The PASI score was not high (mean; 5.2), implying that the degree of psoriasis was not severe in these cases. However, this might have been due to the systemic prednisolone administered for myalgia. PMID- 10875205 TI - A case of pigmented fungiform papillae of the tongue in an Asian male. PMID- 10875206 TI - Erythema multiforme caused by the H2-blocker, roxatidine. PMID- 10875207 TI - ECG of the month. Not so obvious. Complete AV block. PMID- 10875208 TI - External laryngeal trauma. AB - External laryngeal trauma, blunt or penetrating, is a rare but potentially life threatening injury. This is frequently seen in multiple-trauma patients and can go unrecognized in the absence of astute clinical awareness. Injuries may range from small endolaryngeal hematomas or lacerations to complete laryngotracheal separation. Proper airway management is of utmost importance and is one of the most controversial aspects of treatment of laryngeal trauma. Flexible fiberoptic laryngoscopy and high resolution computed tomography scanning of the larynx has greatly enhanced the evaluation of these injuries. Treatment options range from conservative, nonsurgical observation to evaluation in the operating room. Surgical intervention may involve endoscopy, open surgical exploration, and possibly laryngeal stenting. Long-term goals are aimed at maintaining voice, airway, and swallowing ability. A systematic approach to this condition often results in predictable and acceptable outcomes. PMID- 10875209 TI - Radiology case of the month. My aching hip. Intraarticular osteoid osteoma. PMID- 10875210 TI - A grits mill: the story of Field Memorial Hospital. PMID- 10875211 TI - It is more than just the heart for the American Heart Association: new interventions in the vascular tree. PMID- 10875212 TI - Carotid stenting: a technology in evolution. AB - Interest in stenting lesions involving the carotid artery bifurcation has grown during the past 10 years. Techniques first utilized by Theron and Mathias in Europe and Roubin and colleagues in the United States have evolved to where the technique has been extensively refined and its safety and efficacy firmly established. The only prospective randomized study comparing carotid stenting with carotid endarterectomy, CAVATAS (Carotid and Vertebral Artery Transluminal Angioplasty Stenting), showed similar safety profiles and long-term results for both techniques. A large scale NIH-sponsored trial is now in progress, CREST (Carotid Revascularization Endarterectomy versus Stent Trial), but the results are 5-6 years away. In the interim, one approach toward instituting a carotid stent program is described. PMID- 10875213 TI - Role of the catheter in the treatment of cardiac arrhythmias. AB - During the last decade, there has been a remarkable shift away from drug therapy toward catheter-based treatment of many tachyarrhythmias. Catheter ablation using radiofrequency energy has been shown to provide a cure for many supraventricular and ventricular tachycardias with excellent safety and has now become the first line of treatment. A review of biophysics and biology of radiofrequency energy, the technique of catheter ablation, and its application in the treatment of specific tachycardias encountered in clinical practice is presented. PMID- 10875214 TI - Renovascular hypertension: screening and therapeutic options. AB - Renovascular hypertension is part of the spectrum of hypertensive disease. Although uncommon (1% to 5% of the cases) in comparison to essential hypertension, it is a potentially curable form of the disease. We review the different tools available for the evaluation and treatment of this condition. PMID- 10875215 TI - Thrombolytic therapy for acute ischemic stroke. AB - Thrombolytic therapy with recombinant tissue plasminogen activator (rt-RA) is now an accepted treatment for acute ischemic stroke if the patient can be treated within 3 hours of onset of symptoms, and if the clinical presentation justifies use of the medication, and if there are no contraindications to the use of rt-PA. The non-contrast CT brain scan is mandatory to rule out an intracerebral hemorrhage, evidence of subarachnoid hemorrhage, or significant evolution of a large cerebral infarction. The later the patient is treated, within the 3-hour therapeutic window, the less likely there is to be clinical benefit of the treatment and the greater risk of hemorrhagic transformation of the cerebral infarction with potentially catastrophic consequences. There is approximately a 30% greater chance of full recovery from the stroke, at 3 months out from the infarct, with rt-PA compared to no rt-PA. On the other hand, there is a 6.4% risk of symptomatic intracerebral hemorrhage, within 36 hours, associated with the use of rt-PA compared to a 0.6% risk in the placebo group. The greater the neurological deficit at the time of presentation and the greater the evolution of the infarct by the admission CT brain scan, the greater the risk of intracerebral hemorrhage complicating the use of rt-PA. PMID- 10875216 TI - Percutaneous interventional approaches to diseases of the aorta. AB - Diseases of the aorta are prevalent in the Western world. Pathophysiology is influenced by hypertension, atherosclerosis, and genetic factors. The rupture of an aortic aneurysm or the dissection of a hematoma into the aortic wall causes significant mortality in this country. Physicians have long wrestled with therapies to prevent this fatal natural history. It was not until the surgical insertion of aortic grafts in the 20th century that effective therapy was developed. However, surgical mortality and morbidity still remain quite significant in the higher risk population in which these procedures often must be performed. During the 1990s, techniques and devices have been developed which allow placement of endovascular grafts into the aorta percutaneously, without traditional surgery. Two recently FDA-approved devices require a team approach for optimal deployment and care. In the initial experience, these endovascular devices appear to offer the promise of effective treatment with low procedural complications. PMID- 10875217 TI - Historical prologue: why endovascular abdominal aortic aneurysm repair? PMID- 10875218 TI - Mortality and morbidity rates after conventional abdominal aortic aneurysm repair. AB - AIM: To grade and analyse by levels of evidence the mortality and morbidity rates of elective abdominal aortic aneurysm (AAA) surgery as reported over the past 12 years. METHODS: Articles on elective AAA surgery published between 1985 and 1996 were retrieved and classified into 5 levels of evidence. Level 1 contains prospective studies and is subdivided into population-based (Level 1a) and hospital-based (Level 1b) studies. Level 2 includes retrospective studies, subdivided into population-based (Level 2a), hospital-based (Level 2b), and hospital-based studies concerning a specified group of selected patients (Level 2c). Operative mortality and systemic and local/vascular complication rates and 95% confidence intervals were calculated per level of evidence. RESULTS: Seventy two articles describing a total of 37,654 patients could be included: 2 level 1a studies (patient total: 692), 9 Level 1b studies (patient total: 1,677), 13 Level 2a studies (patient total 21,409), 32 Level 2b studies (patient total: 12,019), and 16 Level 2c studies (patient total: 1,857). The mean 30-day mortality rates of the two population-based levels were similar: 8.2% (6.4%-10.6%) for the prospective (1a) and 7.4% (7.0%-7.7%) for the retrospective series (2a). These figures were significantly higher than the remarkably similar hospital-based mortality rates: 3.8% (3.0%-4.8%) for the prospective (1b), 3.8% (3.5%-4.2%) for the retrospective (2b), and 3.5% (2.8%-4.4%) for selected patient group studies (2c). The most frequent complication was of cardiac origin. In the population based series the cardiac complication rate was 10.6% (8.5%-13.2%) and 11.1% (9.1% 13.6%) for Levels 1a and 2a respectively. This compared well with the 12.0% (10.5%-13.9%) for the prospective, hospital-based series (Level 1b). The cardiac complication rates in the retrospective, hospital-based studies was significantly lower: 8.9% (8.4%-9.5%) and 6.1% (4.9%-7.6%) for Levels 2b and 2c respectively. CONCLUSION: There is a clear and consistent disagreement in reported mortality rates between hospital-based and population-based studies of elective AAA surgery. Prospective studies give the best documentation of postoperative morbidity. PMID- 10875219 TI - When not to operate for abdominal aortic aneurysms. AB - Refinement in anaesthetic and surgical techniques for repair of abdominal aortic aneurysms has significantly reduced the mortality associated with treating this condition. Endovascular techniques have further pushed back the frontiers for the treatment of aortic aneurysms, and higher risk patients are now being treated under local or regional anaesthesia. The question of when not to offer intervention is becoming more and more difficult. Age is not a bar to aneurysm surgery in a patient who is physically fit; but the risk and benefit of intervention must be carefully evaluated for each patient on an individual basis, and risk calculation must be evidence based. Contraindications to aneurysm surgery are relative and few and include: small aneurysms (<5.5 cm), a co morbidity that increases surgical risk by >10% and a life expectancy of <1 year. Endovascular graft technology is rapidly advancing, but until the long term results of endovascular repair of aortic aneurysms are proven, the indications for intervention should be the same as for open repair. PMID- 10875220 TI - Overview of techniques and devices for endovascular abdominal aortic aneurysm repair. AB - The endovascular treatment of abdominal aortic aneurysms (AAAs) is rapidly evolving. Since the onset of clinical investigations in 1990 there has been a rapid proliferation in the number of available devices, both surgeon-made and industry-made. This chapter reviews endovascular AAA repair with regard to available devices, patient selection for each device based on anatomic criteria, and techniques for graft deployment. PMID- 10875221 TI - Endovascular repair of abdominal aortic aneurysms. Results from the EUROSTAR registry. EUROpean collaborators on Stent-graft Techniques for abdominal aortic Aneurysm Repair. AB - EUROSTAR (EUROpean collaborators on Stent-graft Techniques for abdominal aortic Aneurysm Repair) was established for the purpose of combining and studying data on endovascular abdominal aortic aneurysm (AAA) repair. EUROSTAR is independent of any commercial interest, and aims to provide scientifically reliable assessment of endovascular AAA grafting. The results of 2,016 patients from 98 European institutions have been collected and analysed. Despite the minimally invasive nature of endovascular aneurysm repair, a variety of complications do occur with considerable frequency. Complications are more often encountered in patients with large aneurysms, advanced age, and if adjuvant procedures are required. In addition, a compromised cardiac and general medical status has adverse effects on the risk of systemic complications. The experience of the operating team is an important factor, influencing device- and procedure-related complications. The observed 18-month endoleak-free survival reflects a satisfactory mid-term result. PMID- 10875222 TI - Specific complications of endovascular aortic repair. PMID- 10875223 TI - Future perspectives of endovascular abdominal aortic aneurysms repair. PMID- 10875225 TI - Aortic endoprosthesis. Closing comments. PMID- 10875224 TI - Endovascular repair of thoracic aortic aneurysms. AB - The standard technique for the treatment of descending thoracic aortic aneurysms is elective open surgical repair with graft interposition. This standard approach, although steadily improving, is associated with high morbidity and substantial mortality rates and implies a major surgical procedure with lateral thoracotomy, use of cardiopulmonary bypass, long operation times and a variety of peri- and postoperative complications. This and the success of the first endoluminal treatment of abdominal aortic aneurysms by Parodi et al. prompted the attention to be thrown on the treatment of descending thoracic aortic aneurysms with endoluminal stent-grafts in many large centres. The aim of this new minimally invasive technique is to exclude the aneurysm from blood flow and in consequence to avoid pressure stress on the aneurysmatic aortic wall, by avoiding a large open operation with significant perioperative morbidity. The potentially beneficial effect of this new treatment approach was evaluated in the course of this study. PMID- 10875226 TI - In search of binding--identification of inhibin receptors. PMID- 10875227 TI - Regulation of corticotropin-releasing factor receptor type 2 beta messenger ribonucleic acid in the rat cardiovascular system by urocortin, glucocorticoids, and cytokines. AB - CRF receptor type 2 (CRF R2) messenger RNA (mRNA) expression in the rodent heart is modulated by exposure to both the bacterial endotoxin lipopolysaccharide (LPS) and glucocorticoids. In this study we examined the roles of glucocorticoids, cytokines, and CRF R2beta ligands in the regulation of CRF R2beta expression in the cardiovascular system both in vivo and in vitro. Using ribonuclease protection assays, we found that, in addition to the injection of LPS or corticosterone, physical restraint caused a decrease in CRF R2beta mRNA levels in the rat heart and aorta. Adrenalectomy with corticosterone replacement at constant levels partially blocked LPS-induced decreases in CRF R2beta mRNA expression in the heart. Thus, elevations of endogenous circulating corticosterone could contribute to the down-regulation of CRF R2beta mRNA expression in heart. To identify other putative modulating factors, we examined CRF R2beta expression in the aorta-derived A7R5 cell line. Incubation with CRF R2 ligands or dexamethasone reduced CRF R2beta mRNA levels. In addition, incubation with a variety of cytokines, proteins released during immune challenge, also reduced CRF R2beta mRNA expression. The multifactorial regulation of CRF R2beta mRNA expression in the cardiovascular system may serve to limit the inotropic and chronotropic effects of CRF R2 agonists such as urocortin during prolonged physical or immune challenge. PMID- 10875229 TI - Transgenic analysis of the response of the rat calbindin-D 9k gene to vitamin D. AB - The promoter of the calbindin-D 9k (CaBP9k) gene, previously analyzed in transgenic mice, contains all of the information necessary for expression of a transgene similar to the endogenous gene and also for an appropriate response to vitamin D. In the present study we first investigated the role of a putative vitamin D-responsive element (9k/VDRE), located at nucleotides -489 to -445 on the rat CaBP9k promoter gene, using transgenic mice. As expected, the pattern of transgene expression in mice carrying this putative VDRE mutated in its whole promoter context was similar to that in mice bearing the wild-type sequence. These transgenic mice also responded to 1,25-dihydroxyvitamin D3 in the same way as those bearing the wild-type transgene and as those carrying a transgene with a large deletion (from -2894 to -117) eliminating the putative 9k/VDRE. Thus, the putative 9k/VDRE is not required for the control of rat CaBP9k gene expression by vitamin D in vivo. We also found that responsiveness to 1,25-dihydroxyvitamin D3 depends on the site at which the transgene is integrated into the host genome, in a tissue-specific manner. These data together with the fact that vitamin D responsive sequences are present in a two-module region (from -3731 to -2894 and/or -117 to +365) and that this region does not contain any classical VDRE show that the CaBP9k gene is submitted to a non-conventional control by vitamin D. PMID- 10875228 TI - A subset of kappa opioid ligands bind to the membrane glucocorticoid receptor in an amphibian brain. AB - Previous studies demonstrated that a membrane receptor for glucocorticoids (mGR) exists in neuronal membranes from the roughskin newt (Taricha granulosa) and that this receptor appears to be a G protein-coupled receptor (GPCR). The present study investigated the question of whether this mGR recognizes nonsteroid ligands that bind to cognate receptors in the GPCR superfamily. To address this question, ligand-binding competition studies evaluated the potencies of various ligands to displace [3H]corticosterone (CORT) binding to neuronal membranes. Initial screening studies tested 21 different competitors and found that [3H]CORT binding was displaced only by dynorphin 1-13 amide (an endogenous kappa-selective opioid peptide), U50,488 (a synthetic kappa-specific agonist) and naloxone (a nonselective opioid antagonist). Follow-up studies revealed that the kappa agonists bremazocine (BRE) and ethylketocyclazocine (EKC) also displaced [3H]CORT binding to neuronal membranes, but that U69,593 (a kappa specific agonist) and nor-BNI (a kappa specific antagonist) were ineffective. The Ki values measured for the opioid competitors were in the subnanomolar to low micromolar range and had the following rank-order: dynorphin > U50,488 > naloxone > BRE > EKC. Because these ligands displaced, at most, only 70% of [3H]CORT specific binding, it appears that some [3H]CORT binding sites are opioid insensitive. Kinetic analysis of [3H]CORT off-rates in the presence of U50,488 and/or CORT revealed no differences in dissociation rate constants, suggesting that there is a direct, rather than allosteric, interaction with the [3H]CORT binding site. In summary, these results are consistent with the hypothesis that the high-affinity membrane binding site for [3H] CORT is located on a kappa opioid-like receptor. PMID- 10875230 TI - Role of estrogen receptor alpha in hematopoietic stem cell development and B lymphocyte maturation in the male mouse. AB - Although estrogens and estrogen receptors (ERs) are known to function in the male brain and reproductive tract, few studies have evaluated their involvement in the male hematopoietic and immune systems. This study was undertaken to determine the role of ERalpha in hematopoietic progenitor and B lymphocyte maturation. ERalpha knockout (ER-/-), wild-type (ER+/+), and radiation chimeric (ERalpha positive or negative in either nonhematopoietic or hematopoietic elements, or both) male mice were used to determine target tissues. ER-/- and ER+/+ animals showed similar hematopoietic progenitor profiles, but the ER-/- animals had fewer cells in all bone marrow B lymphocyte subpopulations. Animals receiving a pharmacological dose (5 mg/kg BW) of 17beta-estradiol (E2) with both elements, ER+/+, had decreased early hematopoietic progenitors and a shift toward a mature B cell subpopulation, whereas animals with both elements, ER-/-, showed changes only in early hematopoietic progenitors. Hematopoietic element ER+/+ animals exhibited greater E2-induced hematopoietic progenitor and B lymphocyte alterations than those having only nonhematopoietic ERalpha. These data indicate that 1) ERalpha is not necessary for regulating male mouse normal hematopoietic progenitor cell proportions, but is involved in B cell regulation; and 2) ERalpha in hematopoietic elements is predominantly responsible for mediating E2-induced hematopoietic and B cell changes. PMID- 10875231 TI - Follistatin is a modulator of gonadal tumor progression and the activin-induced wasting syndrome in inhibin-deficient mice. AB - Inhibins and activins are dimeric proteins belonging to the transforming growth factor-beta superfamily. Follistatin is an activin-binding protein that antagonizes the function of activin via binding to its beta-subunits. Previously, we demonstrated that mice deficient in inhibin develop ovarian and testicular sex cord-stromal tumors of granulosa and Sertoli cell origin, with 100% penetrance as early as 4 weeks of age. Overproduction of activins in the serum directly causes a cachexia-like wasting syndrome that results in lethality of these mice at an early stage after the onset of the tumors. In an independent set of studies, overexpression of mouse follistatin using the mouse metallothionein I promoter in transgenic mice led to gonadal defects and eventual infertility, primarily due to local effects of follistatin in these tissues. Activin has a positive growth effect on gonadal tumor cells in culture and directly causes the cancer cachexia like syndrome in inhibin-deficient mice via interaction with activin receptor type IIA in livers and stomachs. We therefore hypothesized that an activin antagonist such as follistatin can act as a physiological modifier, either locally or via the serum, to block the activin-mediated cancer cachexia-like syndrome in inhibin-deficient mice and/or slow the progression of gonadal cancers in these mice. To test this hypothesis, we generated mice that are homozygous mutant for the inhibin alpha null allele (i.e. inham1/inham1) and carry the mouse metallothionein I follistatin (MT-FS) transgene. Our results show that gonadal tumors that are histologically similar in most, but not all, cases to the tumors in inhibin-deficient mice develop in these inham1/inham1, MT-FS+ mice. However, inham1/inham1, MT-FS+ mice exhibit a less severe wasting syndrome, lower serum activin levels, and a statistically significant prolonged survival in a number of cases compared with mice deficient in inhibin alone. Thus, follistatin can act as a modulator of tumor growth and the activin-induced cancer cachexia-like syndrome in inhibin-deficient mice. PMID- 10875233 TI - Regulation of human gonadotropin-releasing hormone receptor gene expression in placental cells. AB - GnRH has been suggested to regulate hCG secretion in the placenta. In the present study, we report isolation of full-length GnRH receptor (GnRHR) complementary DNA from human placental cells, including a choriocarcinoma cell line (JEG-3), immortalized extravillous trophoblasts (IEVT), and first trimester cytotrophoblast cells in primary culture. Sequence analysis of the placental GnRHR complementary DNA revealed a 100% similarity to its pituitary counterpart. Northern blot analysis using polyadenylated RNA isolated from JEG-3 and IEVT cells revealed a 2.5- and 1.2-kb GnRHR transcripts. Using semiquantitative RT PCR, regulation ofplacental GnRHR gene expression was examined. In contrast to pituitary gonadotrope alphaT3-1 cells, down-regulation of GnRHR messenger RNA (mRNA) levels was not observed in placental cells after 24 h of 0.1-microM GnRH agonist (GnRHa) treatment. Instead, a 43% (P < 0.01) and 30% (P < 0.05) increase in GnRHR mRNA levels was observed in JEG-3 and IEVT cells, respectively. In addition, 10 microM phorbol ester or forskolin treatments resulted in a significant increase in GnRHR expression in both JEG-3 and IEVT cells. The GnRHa induced increase in GnRHR expression was shown to be a receptor-mediated process, as cotreatment of GnRH antagonist abolished the effect. It has also been demonstrated that these stimulatory effects on GnRHR gene expression were regulated at least in part at the transcriptional level. Pretreatment of JEG-3 cells with a specific protein kinase C inhibitor (GF109203X), adenylate cyclase inhibitor (SQ22536), or protein kinase A inhibitor [PKI-(14-22) amide, myristylated] reversed GnRHa-induced GnRHR gene expression, suggesting that the placental GnRHR couples to the protein kinase C (PKC) and cAMP/ protein kinase A (PKA) pathways. By Northern blot analysis, we observed a 100% (P < 0.001) increase in hCGbeta mRNA levels after 0.1 microM GnRHa treatment in JEG-3 cells. Again, this effect was prevented in the presence of either protein kinase C inhibitor or adenylate cyclase inhibitor, further supporting the role of the PKC and PKA pathways in GnRHR-coupled signaling in placental cells. In summary, these data strongly support the idea that 1) GnRH plays an autocrine/paracrine role in regulating placental function through a receptor-mediated mechanism; and 2) the placental GnRHR couples to both the PKC and PKA pathways. PMID- 10875232 TI - In vivo administration of leptin activates signal transduction directly in insulin-sensitive tissues: overlapping but distinct pathways from insulin. AB - To determine whether leptin signal transduction is exerted directly upon insulin sensitive tissues in vivo, we examined the ability of iv leptin to acutely stimulate phosphorylation of STAT3, STAT1, and MAPK, and activities of PI 3 kinase and Akt, in insulin-sensitive tissues of normal rats. Both leptin (1 mg/kg iv x 3 min) and insulin (10 U/kg iv x 3 min) stimulated tyrosine phosphorylation of STAT3 5.6- to 6.0-fold and of STAT1 4.0-fold in adipose tissue. Leptin tended to increase STAT3 phosphorylation in liver and muscle. Both hormones also increased MAPK phosphorylation: leptin increased it 3.2- to 3.8-fold in adipose tissue and liver, whereas insulin stimulated MAPK phosphorylation 5.0-fold in adipose tissue, 6.8-fold in liver, and 2.5-fold in muscle. Leptin was much less effective than insulin at stimulating IRS pathways. Leptin increased IRS-1 associated PI 3-kinase activity in adipose tissue only 2.0-fold (P < 0.01) compared with the 10-fold effect of insulin. IRS-2-associated PI 3-kinase activity was increased 1.7-fold (P < 0.01) by leptin in liver and 6-fold by insulin. Akt phosphorylation and activity were not changed by leptin but increased with insulin. Lower concentrations of leptin (10 and 50 microg/kg) also stimulated STAT3 phosphorylation in fat. These effects appear to be direct because 3 min after leptin intracerebroventricular injection, phosphorylation of STAT3, STAT1, and MAPK were not stimulated in hypothalamus or adipose tissue. Furthermore, leptin activated STAT3 and MAPK in adipose tissue explants ex vivo and in 3T3-L1 adipocytes. Leptin did not activate STAT3 or MAPK in adipose tissue of db/db mice. Thus, leptin rapidly activates signaling pathways directly at the level of insulin sensitive tissues through the long-form leptin receptor, and these pathways overlap with, but are distinct from, those engaged by insulin. PMID- 10875234 TI - Regulation of insulin secretion by overexpression of Ca2+/calmodulin-dependent protein kinase II in insulinoma MIN6 cells. AB - Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) may play a key role in Ca2+-induced insulin secretion. We have previously reported that treatment of insulinoma MIN6 cells with secretagogues activated CaM kinase II and increased the phosphorylation of synapsin I, followed by insulin secretion. Here, we identified isoforms of CaM kinase II in MIN6 cells and rat islets. Immunoblot analysis suggested that the major isoforms of CaM kinase II were beta'e and delta2 at the protein level in MIN6 cells. Only the beta'e isoform was detected in rat islets by both RT-PCR and immunoblot analysis. We transiently overexpressed beta'e and delta2 isoforms in MIN6 cells and confirmed that treatment of cells with tolbutamide and glucose activated the isoforms. Immunoblot analysis with an antibody against synapsin I phosphorylated by CaM kinase II demonstrated that treatment with tolbutamide and glucose rapidly increased phosphorylation of synapsin I and that phosphorylation was potentiated by overexpression of the isoforms. The secretagogue-induced insulin secretion was potentiated by overexpression of the isoforms. Our results further support our conclusion that activation of CaM kinase II and the concomitant phosphorylation of synapsin I contribute to insulin secretion from pancreatic beta-cells. PMID- 10875235 TI - A histone deacetylase inhibitor potentiates estrogen receptor activation of a stably integrated vitellogenin promoter in HepG2 cells. AB - To compare the role of histone deactylation in estrogen activation of a transiently transfected vitellogenin (VIT) promoter and an integrated VIT promoter in the same cells, we produced three HepG2, human hepatoma, cell lines (HepG2ERV cells) stably expressing human estrogen receptor alpha (hERalpha) and containing an integrated VIT promoter-chloramphenicol acetyltransferase (VIT-CAT) reporter gene. The three ER-positive HepG2ERV cell lines and wild-type, ER negative, HepG2 cells cotransfected with cytomegalovirus-hERalpha exhibited similar MOX-dependent inductions of 20- to 50-fold with a transiently transfected VIT-luciferase reporter and 15- to 50-fold with a transfected 4-estrogen response element-TATA-luciferase reporter gene. The histone deacetylase inhibitor, trichostatin A, did not enhance MOX induction of the transiently transfected VIT promoter in the HepG2ERV cells. In contrast, trichostatin A dramatically potentiated MOX induction of the stably integrated VIT-CAT reporter gene, resulting in MOX-ER-dependent increases in CAT activity of up to 600-fold. These data demonstrate that although liganded ER exhibits the capacity to fully activate a transiently transfected VIT promoter, under some circumstances the ability to reorganize a repressive chromatin structure may be limiting for steroid receptor action. PMID- 10875236 TI - Activation of signal transducer and activator of transcription-3 during proliferative phases of 3T3-L1 adipogenesis. AB - Signal transducer and activator of transcription-3 (STAT3) is abundantly expressed in preadipocytes and adipocytes, but little is known about its activation status or functional role during adipogenesis. In this report we investigate STAT3 activation in 3T3-L1 preadipocytes before and after differentiation into adipocytes. STAT3 was highly tyrosine phosphorylated and bound to DNA in proliferating preadipocytes, but not in growth-arrested preadipocytes or adipocytes. In growth-arrested confluent preadipocytes, induction of differentiation with methylisobutylxanthine, dexamethasone, and high dose insulin led to a delayed, but prolonged (3-day), increase in STAT3 tyrosine phosphorylation. This increase in STAT3 phosphorylation coincided temporally with postconfluent preadipocyte mitotic clonal expansion. Insulin and methylisobutylxanthine alone, but not dexamethasone, induced STAT3 tyrosine phosphorylation in postconfluent cells. Diminution of endogenous STAT3 expression by antisense morpholino oligonucleotides significantly decreased preconfluent preadipocyte proliferation. Collectively, these findings suggest a regulatory role for STAT3 during the proliferative phases of adipogenesis. PMID- 10875237 TI - Calcium ions positively modulate follicle-stimulating hormone- and exogenous cyclic 3',5'-adenosine monophosphate-driven transcription of the P450(scc) gene in porcine granulosa cells. AB - Given the evident modulation of FSH-induced steroidogenesis by Ca2+ in granulosa cells, we here test the hypothesis that Ca2+ controls expression of the enzymatically rate-limiting cytochrome P450(scc) (CYP11A) gene. To test this postulate, we quantitated the ability of Ca2+ to regulate: 1) transcriptional activity of a transiently transfected luciferase reporter gene driven by a 2.32 kb 5'-upstream fragment of the porcine P450(scc) gene promoter region; and 2) accumulation of endogenous P450(scc) transcripts in primary monolayer cultures of porcine granulosa cells. To this end, granulosa cells were stimulated for 4 h with FSH (15 ng/ml, NIDDK-oFSH-20) or 8-Bromo-cAMP (8 Br-cAMP, 1 mM) in serum free medium containing either 1.8 mM Ca2- or no added Ca2+ with 100 microM EGTA or 100 microM CoCl2. In the presence of extracellular Ca2+, FSH and 8 Br-cAMP stimulated expression of the transfected P450(scc) promoter-reporter fusion construct by 5.6 +/- 1.1 and 3.6 +/- 0.67-fold, respectively over Ca2+-containing unstimulated control (P < or = 0.04, n = 5-6 experiments). The foregoing two agonists augmented 4-h progesterone production by cultured granulosa cells by 1.8 +/- 0.11 and 1.6 +/- 0.16-fold, respectively (P < or = 0.001 for FSH and P < or = 0.01 for 8 Br-cAMP). FSH and 8 Br-cAMP also significantly elevated endogenous P450(scc) transcript levels as measured by homologous solution-hybridization RNase protection assay; i.e. by 3.1 +/- 0.49 and 2.9 +/- 0.45-fold, respectively (P < or = 0.001). In Ca2+-free/EGTA-supplemented medium, basal luciferase reporter-gene activity and endogenous P450(scc) messenger RNA accumulation in granulosa cells declined to 34 +/- 12% and 78 +/- 12%, respectively, of corresponding values in control (unstimulated Ca2+-containing) cultures. Extracellular Ca2+ deprivation inhibited the stimulatory effect of FSH (and 8 Br cAMP) on P450(scc) promoter-luciferase reporter expression to 58 +/- 30% (and 58 +/- 23%), and restrained endogenous P450(scc) message accumulation to 86 +/- 15% (and 96 +/- 18%) of the value in Ca2+-containing control. Extracellular Ca2+ withdrawal suppressed FSH (and 8 Br-cAMP)-driven progesterone production over 4 h to basal levels but did not alter FSH-stimulated cAMP accumulation by granulosa cells. Ca2+-deprived cells exposed to serum-containing media regained P450(scc) responsiveness to both agonists. Antagonism of cellular uptake of Ca2+ and other divalent cations via administration of cobalt chloride (100 microM) inhibited FSH and 8 Br-cAMP's stimulation of endogenous (but not exogenous promoter-driven) P450(scc) gene expression. In contrast, granulosa-cell concentrations of messenger RNA's encoding sterol-carrier protein-2 (SCP-2) and the low density lipoprotein receptor were not altered by Ca2+ withdrawal. In summary, uptake of extracellular Ca2+ by porcine granulosa cells significantly potentiates transactivation of the endogenously expressed and exogenously transfected P450(scc) gene by FSH and 8 Br-cAMP. The agonistic impact of Ca2+ on P450(scc) promoter activity is requisite downstream of FSH-induced cAMP second-messenger signaling. PMID- 10875239 TI - Nuclear receptors Nor1 and NGFI-B/Nur77 play similar, albeit distinct, roles in the hypothalamo-pituitary-adrenal axis. AB - Studies in Nur77-deficient mice have shown that the basal regulation of hypothalamic and pituitary functions as well as the adrenocortical steroidogenesis in these animals is normal. This indicates that Nur77-related orphan receptors may substitute Nur77 functions in the hypothalamo-pituitary adrenal axis by a compensatory mechanism. Nor1 is the most recently cloned member of the NGFI-B/Nur77 subfamily, and its properties are still largely unknown. We demonstrate here that Nor1 is expressed in the pituitary gland and adrenal cortex, and that ACTH and angiotensin II (AngII) treatment of adrenal fasciculata cells induces Nor1 expression. Time-course analysis with both hormones on steroidogenic capacity and the specific gene expression in adrenal cells strongly suggest that Nor1 is an intermediate in the long-term consequences of ACTH or AngII treatment. The Nor1 and NGFI-B/Nur77 amino acid sequence homology and the analysis of the trans-activation properties of Nor1 show that the overall structural and functional organization of the two proteins is similar. As observed with NGFI-B/Nur77, Nor1 activates the expression of genes encoding steroidogenic enzymes as P450c21, through its interaction with NGFI-B response element promoter sequences. In contrast, binding experiments of Nor1 with the palindromic NurRE sequence suggest that Nor1 is not an efficient substitute for the NGFI-B/Nur77 activation of the POMC gene expression in pituitary glands. All these results indicate that Nor1 and NGFI-B/Nur77 may play similar albeit distinct roles in the hypothalamo-pituitary-adrenal axis. Further experiments also show that the mechanisms responsible for the transcriptional regulation of Nor1 in adrenal cells appear to depend on the protein kinase A and protein kinase C cascades. PMID- 10875238 TI - Induction of early growth response protein-1 gene expression in the rat ovary in response to an ovulatory dose of human chorionic gonadotropin. AB - Granulosa cells in a mature ovarian follicle have an abundance of LH/hCG receptors that respond rapidly to an ovulatory surge in gonadotropins. Within minutes, membrane signal transduction sets in motion metabolic changes that lead to follicular rupture. This study provides evidence that the initial ovarian response to such an ovulatory stimulus includes induction of the immediate-early transcription factor gene for early growth response protein-1 (Egr-1). Immature Wistar rats were primed with 10 IU equine CG (eCG), sc, and 48 h later the 12-h ovulatory process was initiated by 10 IU hCG, sc. Ovarian RNA was extracted at 0, 0.5, 1, 2, 4, 8, 12, and 24 h after the primed animals were injected with hCG. The RNA extracts were used for RT-PCR differential display for random detection of gene expression in the stimulated ovarian tissue. Northern analysis of one of the differentially amplified complementary DNAs confirmed that it was part of a gene that was significantly up-regulated within 1 h after the ovaries had been stimulated by hCG. Maximum transcription was at 4 h after hCG, and expression declined to 0 h control levels by 24 h after hCG. Subcloning and sequence analysis revealed that the complementary DNA matched the gene for Egr-1. In situ hybridization indicated that the Egr-1 messenger RNA was in the granulosa layer of mature follicles. Western blotting confirmed the temporal pattern of Egr-1 expression detected by differential display, Northern analysis and in situ hybridization. The Egr-1 protein is approximately 84 kDa. In conclusion, the data show that expression of the zinc finger transcription factor Egr-1 is an early event in the cascade of inflammatory-like changes that occur in an ovulatory follicle in response to a trophic hormone. PMID- 10875240 TI - Insulin-like growth factor binding protein-3 is regulated by dihydrotestosterone and stimulates deoxyribonucleic acid synthesis and cell proliferation in LNCaP prostate carcinoma cells. AB - We have investigated the production and actions of a growth regulatory protein, insulin-like growth factor binding protein (IGFBP)-3, in the androgen-responsive prostate carcinoma cell line LNCaP. Confluent monolayers of cells secreted approximately 0.7 ng/ml IGFBP-3 over 24 h. Dihydrotestosterone (DHT, 10 nM) and 1,25-dihydroxyvitamin D3 (vitamin D, 10 nM) increased IGFBP-3 in media to 149 +/- 15% and 206 +/- 18% of control, respectively, when added separately, and to 453 +/- 28% of control when used in combination. IGFBP-2, secreted at approximately 25-fold higher concentrations than IGFBP-3, was increased 50% by 10 nM DHT, but there was no effect of vitamin D on IGFBP-2 production in the absence or presence of DHT. Cell-associated IGFBP-3, and immunoreactive IGFBP-3 species of 20 kDa and 30 kDa were also increased in response to vitamin D plus DHT. A combination of vitamin D and DHT increased DNA synthesis in LNCaP cells 3-fold, and this was at least partly mediated by endogenous IGFBP-3 because anti-IGFBP-3 IgG, but not nonimmune serum IgG, reduced the stimulatory effect of vitamin D and DHT from 293 +/- 11.6% to 161 +/- 30.7% of control levels (P < 0.0001). Basal and DHT plus vitamin D-stimulated thymidine incorporation was significantly increased by 50 ng/ml human plasma-derived purified IGFBP-3. After 4 days treatment with vitamin D plus DHT, or pure IGFBP-3, LNCaP cell numbers were increased relative to control. These results indicate a role for IGFBP-3 in the proliferation of androgen-responsive prostate carcinoma cells. PMID- 10875241 TI - Dissection of differentially regulated (G+C)-rich promoters of the human parathyroid hormone (PTH)/PTH-related peptide receptor gene. AB - The PTH/PTH-related peptide (PTHrP) receptor (PTHR) is required for normal skeletal development, and a wide array of physiological responses mediated by PTH and PTHrP. We have previously identified three promoters, P1-P3, which control human PTHR gene transcription. P2 and P3 are (G+C)-rich, function in a number of tissues, lie within the same CpG island, and display many hallmarks of housekeeping promoters. However, they are differentially regulated during development as P2, but not P3, functions in fetal tissues. Here, we have used both stably and transiently transfected human osteoblast-like cells to delineate regions of P2 and P3 required for promoter activity. Deletion analyses performed in stably transfected cells indicated that sequences extending from -91 to -12 relative to the transcription start site were required for function of the P2 promoter. No negative regulatory elements were detected in P2. In contrast, deletion of an A-rich region of P3 extending from -147 to -115 was required for optimal basal activity, suggesting that this sequence acts as a repressor of P3. Strikingly, however, whereas the A-rich region also functioned as a negative element when inserted upstream of the (G+C)-rich P2 promoter, it enhanced expression from the thymidine kinase promoter, suggesting that its function depends on other transcription factors bound to promoter sequences. Fine deletion of P3 sequences proximal to -115 implicated Spl motifs and downstream initiation sites in P3 function. These studies indicate that function of P2 and P3 is controlled by ubiquitously expressed transcription factors and raise the possibility that P3 activity is repressed during fetal development. PMID- 10875242 TI - Increased fetal glucocorticoid exposure delays puberty onset in postnatal life. AB - The fetal environment is now recognized as a key determinant of the adult phenotype, being linked to development of diseases, including hypertension, as well as the timing of puberty. Such links may be related, in part, to the level of fetal exposure to maternal glucocorticoids in utero, which is normally regulated by placental expression of the enzyme 11beta-hydroxysteroid dehydrogenase (11beta-HSD). The present study examined whether manipulation of fetal glucocorticoid exposure, either directly or indirectly via 11beta-HSD inhibition, influences the subsequent timing of puberty. Administration of dexamethasone acetate at low (LDEX, 0.25 microg/ml drinking water) or high doses (HDEX, 1 microg/ml) or carbenoxolone (CBX, 2 x 10 mg/day, sc; an inhibitor of 11beta-HSD) to pregnant rats from day 13 to term (day 23) reduced offspring birthweight (LDEX: 9%; HDEX: 27%; CBX: 8%) and resulted in a subsequent delay in the onset of puberty in females (control: 41.4 +/- 0.5; LDEX: 44.8 +/- 0.7; HDEX: 48.5 +/- 0.4; CBX: 43.6 +/- 0.5 days). Importantly, the effects of CBX were not observed in the absence of maternal adrenals, indicating that they were mediated by increased fetal exposure to endogenous maternal glucocorticoids. In contrast, maternal treatment with metyrapone (MET; an inhibitor of glucocorticoid synthesis; 500 microg/ml drinking water from day 13) increased birthweight by 5% and advanced puberty onset in male offspring (control: 48.8 +/- 1.0; MET: 45.7 +/ 0.8 days). Changes in the timing of puberty onset were not attributable to changes in either bodyweight at puberty or peripubertal plasma leptin concentrations. Peripubertal plasma LH was also unaffected in animals with delayed puberty but was elevated in male offspring of MET-treated mothers. Collectively, these results demonstrate that fetal glucocorticoid exposure is an important determinant of the timing of puberty onset in postnatal life, and that this effect is operable within the normal physiological range of glucocorticoid concentrations. PMID- 10875243 TI - Tyrosine kinase and phosphatidylinositol 3-kinase activation are required for cyclic adenosine 3',5'-monophosphate-dependent potentiation of deoxyribonucleic acid synthesis induced by insulin-like growth factor-I in FRTL-5 cells. AB - In previous studies, we showed that pretreatment of rat FRTL-5 thyroid cells with TSH, or other agents that increased intracellular cAMP, markedly potentiated DNA synthesis in response to insulin-like growth factor-I (IGF-I). In addition, we found that TSH pretreatment caused an increase in tyrosine phosphorylation of intracellular proteins including an unidentified 125-kDa protein that was well correlated with the TSH-potentiating effect on DNA synthesis induced by IGF-I. These results suggested that cAMP amplified IGF-I-dependent signals for cell growth through changes of cAMP-dependent tyrosine phosphorylation. The present studies were undertaken to determine how tyrosine kinase activation followed by an increase in tyrosine phosphorylation is required for cAMP-dependent potentiation of DNA synthesis induced by IGF-I in this cell line. First of all, we measured tyrosine kinase or protein-tyrosine phosphatase activities in the cell lysates by the in vitro assay. Chronic treatment with TSH or (Bu)2-cAMP stimulated tyrosine kinase activity in the particulate fraction and protein tyrosine phosphatase activity in the soluble fraction, suggesting that tyrosine kinase plays more important roles for a cAMP-dependent increase in tyrosine phosphorylation of intracellular proteins. The increased tyrosine kinase activity was sensitive to genistein, a potent tyrosine kinase inhibitor. Genistein abolished both the cAMP-dependent increase in tyrosine phosphorylation of the 125 kDa protein and the enhanced DNA synthesis induced by IGF-I in a similar concentration-dependent manner. The only tyrosine-phosphorylated protein associated with the p85 regulatory subunit of phosphatidylinositol (PI) 3-kinase in response to cAMP was 125 kDa. In addition, we found that PI 3-kinase activity bound to p85 subunit significantly increased after (Bu)2cAMP treatment. These results suggested that cAMP stimulates PI 3-kinase through tyrosine phosphorylation of the 125-kDa protein. We then measured DNA synthesis in cells pretreated for 24 h with TSH or (Bu)2cAMP in the absence or presence of LY294002, a PI 3-kinase inhibitor, followed by treatment with IGF-I for 24 h. Presence of LY294002 during TSH or (Bu)2cAMP pretreatment completely abolished cAMP-dependent potentiation of DNA synthesis induced by IGF-I. These results suggest that in FRTL-5 cells cAMP activates genistein-sensitive tyrosine kinases that in turn activate PI 3-kinase activity. These mechanisms appear to be necessary for cAMP dependent potentiation of the DNA synthesis induced by IGF-I. PMID- 10875244 TI - Transcriptional activation of thymidylate synthase by 17beta-estradiol in MCF-7 human breast cancer cells. AB - Thymidylate synthase (TS) catalyzes methylation of deoxyuridine phosphate to give deoxythymidine phosphate, and 17beta-estradiol (E2) induces TS gene expression in MCF-7 human breast cancer cells. Analysis of the TS gene promoter showed that E2 responsiveness required the -229 to -140 promoter region containing a G-rich sequence and CACCC box. Subsequent mutational analysis of this region indicated that only the G-rich motif (-150 to -142) was required for E2 action. Results of gel mobility shift and in vitro DNA footprinting assays showed that both estrogen receptor alpha (ERalpha) and Sp1 proteins were required for hormone-induced trans activation that involved ERalpha/Sp1 binding to the G-rich site in which only Sp1 protein bound DNA. Both proteins also interacted in Drosophila cells in functional assays, confirming the transcriptional activation of TS-involved ERalpha/Sp1, and this adds to the increasing number of genes that are activated through this pathway in breast cancer cells. PMID- 10875245 TI - Immunohistochemical localization, biochemical characterization, and biological activity of neurotensin in the frog adrenal gland. AB - The primary structure of neurotensin has been recently determined for the frog Rana ridibunda (Endocrinology 139: 4140-4146, 1998). In the present study, we have investigated the distribution and biochemical characterization of neurotensin-like immunoreactivity in the frog adrenal gland, using an antiserum directed against the conserved C-terminal region of the peptide. Neurotensin-like immunoreactivity was detected in two populations of nerve fibers: numerous varicose fibers coursing between adrenal cells, and a few processes located in the walls of blood vessels irrigating the gland. Reversed-phase HPLC analysis of frog adrenal gland extracts revealed the existence of a major peak of neurotensin like immunoreactivity that exhibited the same retention time as synthetic frog neurotensin. The possible involvement of neurotensin in the regulation of steroid secretion was studied in vitro using perifused frog adrenal slices. For concentrations ranging from 10(-10) to 10(-5) M, synthetic frog neurotensin increased corticosterone and aldosterone production in a dose-dependent manner (EC50 = 1.2 x 10(-9) M and 5.8 x 10(-10) M, respectively). Repeated administration of neurotensin induced a reproducible stimulation of steroid output without any tachyphylaxis. Prolonged administration (3 h) of frog neurotensin caused a transient increase in corticosterone and aldosterone secretion followed by a decline of corticosteroid secretion. Neurotensin also produced a significant stimulation of corticosteroid secretion from dispersed frog adrenal cells. This study demonstrates that neurotensin is located in nerve processes innervating the adrenal gland of amphibians. The results also show that synthetic frog neurotensin exerts a direct stimulatory effect on corticosteroid output. Taken together, these data support the view that neurotensin, released by nerve fibers, may act as a local regulator of corticosteroid secretion. PMID- 10875246 TI - Inhibition of nitric oxide synthase activity diminishes the acute effects of relaxin on growth, but not softening, of the cervix in the rat. AB - Relaxin promotes growth and softening of the cervix during pregnancy in the rat. This study examined the hypothesis that nitric oxide (NO) mediates the effects of relaxin on the rat cervix. To test that hypothesis, N omega-nitro-L-arginine methyl ester (L-NAME) was used to inhibit NO synthase, the enzyme that converts arginine to NO and L-citrulline. Nonpregnant rats were ovariectomized when they were 78 days old (day 1 of treatment). At ovariectomy each animal was fitted with silicon tubing implants containing progesterone (P) and estrogen (E) in doses that provide blood levels similar to those during late pregnancy. Rats were assigned to three treatment groups. The control group OPE (n = 6 rats) received 0.5 ml L-NAME vehicle (PBS) sc at 6-h intervals from 0600 h on day 7 through 1200 h on day 8 and 0.5 ml relaxin vehicle (PBS) sc at 0600 and 1200 h on day 8. Group OPER (n = 6 rats) was treated in the same way as group OPE, except that 20 microg porcine relaxin were administered. Group OPERI (n = 7 rats) was treated in the same way as group OPER, except that L-NAME was administered at a dose of 100 mg/kg x 6 h. Between 1400-1500 h on day 8, the cervices were removed and weighed. Cervical wet weight and extensibility were markedly greater (P < 0.01) in relaxin treated group OPER rats than in group OPE controls. Treatment with L-NAME diminished relaxin's effects on cervical wet weight, but not cervical extensibility. In conclusion, this study provides evidence that NO contributes to the acute effects of relaxin on the growth, but not the softening, of the rat cervix. PMID- 10875247 TI - Effects of leptin receptor mutation on Agrp gene expression in fed and fasted lean and obese (LA/N-faf) rats. AB - Agouti-related protein provides an orexigenic signal, probably through interaction with central melanocortin receptors. Expression of Agrp is markedly increased in the hypothalamus of mice deficient in leptin (Lep(ob)/Lep(ob)) or its receptor (Lepr(db)/Lepr(db)), suggesting that leptin mediates signals suppressing Agouti-related protein production. The regulation of Agrp expression in the rat hypothalamus has not been reported. We, therefore, analyzed the expression of Agrp in the medial basal hypothalamus of lean (+/+, +/fa(f)) and obese leptin receptor-deficient (fa(f)/fa(f)) LA/N rats. Using a sensitive solution hybridization/S1 nuclease protection assay, we found no significant difference in Agrp messenger RNA (mRNA) levels (pg/microg total RNA +/- SEM) in obese rats (n = 5), compared with lean controls (n = 5): 0.46 +/- 0.06 vs. 0.47 +/- 0.06 (P = 0.9). Similarly, no difference in Agrp expression was found using in situ hybridization or semiquantitative RT-PCR. In contrast to Agrp, Pomc mRNA levels were significantly suppressed in the obese, compared with the lean, rats (P = 0.001). Thus, the ratio of Pomc to Agrp mRNA is decreased in the obese rats and may be an important modulator of food intake. To assess the physiological regulation of Agrp in rats, we examined the effect of food deprivation in lean Sprague Dawley (SD) rats. There was a 273% increase in medial basal hypothalamus Agrp mRNA in SD rats fasted for 48 h (n = 8), compared with rats fed ad libitum (n = 8): 0.82 +/- 0.23 vs. 0.30 +/- 0.08 (P = 0.0001). Lean LA/N rats (n = 7) fasted for 48 h also showed a 231% increase in Agrp expression, compared with fed lean controls (n = 8): 0.74 +/- 0.11 vs. 0.32 +/- 0.03 (P = 0.002), whereas Pomc expression was decreased by 32% in fasted animals from the same experiment (0.34 +/- 0.05 vs. 0.50 +/- 0.07; P = 0.03). There were no significant differences in Agrp or Pomc mRNA levels between fasted and fed obese LA/N-fa(f) rats. These results suggest that, in the rat, the Agrp response to fasting may involve leptin mediated phenomena, but factors in addition to leptin must also be involved in the regulation of Agrp gene expression. PMID- 10875248 TI - Distribution of uridine diphosphate-glucuronosyltransferase (UGT) expression and activity in cynomolgus monkey tissues: evidence for differential expression of steroid-conjugating UGT enzymes in steroid target tissues. AB - Based on the similarity of pathways and enzymes involved in steroid metabolism, simians represent a relevant animal model to study steroid elimination by glucuronidation. In this study the tissue distribution of UDP glucuronosyltransferase (UGT) transcripts, proteins, and enzymatic activities were examined in 24 different cynomolgus monkey tissues. RT-PCR and Western blot analysis on total RNA and microsomal proteins demonstrated the presence of UGT1A and UGT2B transcripts and proteins in a wide range of tissues including steroid target tissues. Glucuronidation activity on eugenol, 5alpha-androstane 3alpha,17beta-diol, androsterone, and 4-hydroxyestradiol was measured using tissue homogenates and radiolabeled [14C]UDP-glucuronic acid. All tissues contained conjugation activity on these substrates, but glucuronidation rates were significantly lower in steroid target tissues than in liver, kidney, or gut. However, the ratio of steroid glucuronidation vs. eugenol glucuronidation was higher in steroid target tissues, suggesting a differential expression of steroid conjugating enzymes in these tissues. Taken together, these results clearly demonstrate the presence of steroid glucuronidation enzymes in extrahepatic steroid target tissues and support the hypothesis that steroid glucuronidation is an important intracrine pathway involved in termination of steroid signaling. PMID- 10875249 TI - Cell density influences insulin-like growth factor I gene expression in a cell type-specific manner. AB - The effect of cellular density on insulin-like growth factor I (IGF-I) gene expression was characterized in several tumor-derived cell lines. IGF-I messenger RNA (mRNA) transcripts increased more than 200-fold when C6 glioma cells grew to postconfluence. IGF-I receptor and beta-actin mRNAs were induced by 6- and 2 fold, respectively, as a function of confluence. IGF-I mRNA transcripts in GH3 and SK-N-MC cells increased about 4- to 5-fold in confluent cultures compared with sparse cultures. In OVCAR-3 cells, the IGF-I mRNA level remained constant as the cell density increased. Transient transfection experiments were performed with IGF-I exon 1 promoter/luciferase fusion constructs in C6 cells. The luciferase activity of a construct containing exon 1 sequence between +75 and +282 (the most 5' transcription initiation site was designated +1) was stimulated by 2.5-fold in dense cultures compared with that in sparse cultures of C6 cells. Luciferase activities of other constructs containing at least 282 bp of exon 1 sequence were also stimulated about 2- to 4-fold by cell density. However, 3' deletion to +192 led to loss of the cell density stimulatory effect. In contrast, luciferase activities of IGF-I promoter constructs were not altered by cell density in SK-N-MC cells. When the conditioned medium of low density C6 cultures was exchanged with that of high density cultures, the IGF-I mRNA level remained the same. In summary, cell density has a cell type- and gene type-specific effect on IGF-I gene expression. A cell density response element(s) may be located between +192 and +282 of the exon 1 promoter region in C6 cells. PMID- 10875250 TI - Effects of selenium deficiency on tissue selenium content, deiodinase activity, and thyroid hormone economy in the rat during development. AB - The iodothyronine deiodinases, D1, D2, and D3, all contain selenium (Se) in the form of selenocysteine at their active sites, and they play crucial roles in determining the circulating and intracellular levels of the active thyroid hormone (TH), T3. However, not only are serum T3 levels normal in Se-deficient rats but phenotypic and reproductive abnormalities are minimal, and it has been suggested that regulatory mechanisms exist to conserve Se in critical tissues. The present study was designed to determine, in rats: 1) whether the effects of Se-deficiency are greater in the fetus and neonate than in the adult; 2) whether there are tissues other than brain and thyroid in which deiodinase activities are maintained; 3) whether the maintenance of deiodinase activity in a specific tissue is associated with a concomitant preservation of Se level in that tissue; and 4) whether TH economy and general health is maintained over several generations. The tissues studied included liver, cerebrum, thyroid, pituitary, skin, brown adipose tissue, uterus, ovary, testis, placenta, and the implantation site (uterus plus contents) at E9. The results have revealed that, with the exception of liver, skin, and nonpregnant uterus, all of the tissues studied maintained substantial deiodinase activity (>50%) during prolonged Se-deficiency. Second, although the ability of a tissue to maintain deiodinase activity in the face of dietary Se deprivation was associated in some tissues with a concomitant local preservation of Se concentration, this was not the case for all tissues. Only when Se levels were decreased by more than 80% was deiodinase activity markedly decreased. Third, the effects of Se-deficiency were no greater in the fetus than in the adult; and fourth, at the level of Se-deficiency employed in this study, TH economy and general health were successfully maintained over six generations of Se-deficient rats. How Se levels are maintained in specific tissues, whether Se is sequestered in specific cells of a tissue or organ during dietary Se deprivation, and the precise mechanisms by which plasma T3 levels are maintained in Se-deficient animals remain unanswered. Further insights may be gained by using diets that are even lower in Se than those that were used herein and/or by conducting studies using radioactive forms of Se and thyroid hormones. PMID- 10875251 TI - Design of a synthetic leptin agonist: effects on energy balance, glucose homeostasis, and thermoregulation. AB - We have previously reported that a synthetic peptide amide corresponding to amino acid residues 116-130 of mouse leptin, LEP-(116-130), reduces body weight gain, food intake, and blood glucose levels in ob/ob and db/db mice. In the present study we show that the activity of LEP-(116-130) resides in a restricted sequence between amino acid residues 116-122. A synthetic peptide corresponding to this sequence (Ser-Cys-Ser-Leu-Pro-Gln-Thr) has been named OB3. Single point D-amino acid substitution was used to study the structure-function relationship of each residue in OB3. D-Amino acid analogs of OB3 were synthesized by the solid phase method, purified to 98+%, and administered (1 mg/day, ip) for 7 days to female C57BL/6J ob/ob mice. The effects of the peptides on body weight gain, food and water intake, glucose homeostasis, and thermoregulation were assessed. In most cases, the efficacy of OB3 on all parameters tested was reduced by substitution of an L-amino acid with its corresponding D-isoform. A statistically significant increase (2.6-fold) in the weight-reducing effect of OB3, however, was observed by inversion of the configuration of the leucine residue at position 4 (Leu-4) of OB3 by substitution with its D-amino acid isoform [D-Leu-4]. Compared with OB3, mice treated with [D-Leu-4]-OB3 consumed 7.9% less food and 16.5% less water. Blood glucose was normalized to levels comparable to those in wild-type control mice within 2 days after initiation of [D-Leu-4]-OB3 treatment. Unlike native leptin, however, neither OB3 nor any of its D-amino acid-substituted analogs had any apparent effect on thermogenesis. Our results indicate that synthetic peptide strategies may be useful in the development of potent and stabile pharmacophores with potential therapeutic significance in the treatment of human obesity and its related metabolic dysfunctions. PMID- 10875252 TI - Insulin inhibits the ubiquitin-dependent degrading activity of the 26S proteasome. AB - A major metabolic effect of insulin is inhibition of cellular proteolysis, but the proteolytic systems involved are unclear. Tissues have multiple proteolytic systems, including the ATP- and ubiquitin-dependent proteasome pathway. The effect of insulin on this pathway was examined in vitro and in cultured cells. Insulin inhibited ATP- and ubiquitin-dependent lysozyme degradation more than 90% by reticulocyte extract, in a dose-dependent manner (IC50 approximately 50 nM). Insulin did not reduce the conjugation of ubiquitin to lysozyme and was not itself ubiquitin-conjugated. In HepG2 cells, insulin increased ubiquitin conjugate accumulation 80%. The association between the 26S proteasome and an intracellular protease, the insulin-degrading enzyme (IDE), was examined by a purification scheme designed to enrich for the 26S proteasome. Copurification of IDE activity and immunoreactivity with the proteasome were detected through several chromatographic steps. Glycerol gradient analysis revealed cosedimentation of IDE with the 20S proteasome and possibly with the 26S proteasome. The proteasome-associated IDE was displaced when the samples were treated with insulin. These results suggest that insulin regulates protein catabolism, at least in part, by decreasing ubiquitin-mediated proteasomal activity, and provides a new target for insulin action. The displacement of IDE from the proteasome provides a mechanism for this insulin action. PMID- 10875253 TI - Production of immunoreactive thyroglobulin C-terminal fragments during thyroid hormone synthesis. AB - Here, we studied the fragmentation of the prothyroid hormone, thyroglobulin (Tg), which occurs during thyroid hormone synthesis, a process which involves iodide, thyroperoxidase, and the H2O2-generating system, consisting of glucose and glucose oxidase. Various peptides were found to be immunoreactive to autoantibodies to Tg from patients and monoclonal antibodies directed against the immunodominant region of Tg. The smallest peptide (40 kDa) bore thyroid hormones and was identified at the C-terminal end of the Tg molecule, which shows homologies with acetylcholinesterase. Similar peptides were obtained by performing metal-mediated oxidation of Tg via a Fenton reaction. It was concluded that the oxidative stress induced during hormone synthesis generates free radicals, which, in turn, cleave Tg into immunoreactive peptides. PMID- 10875254 TI - Estrogen modulates parathyroid hormone-induced interleukin-6 production in vivo and in vitro. AB - Interleukin (IL)-6 promotes osteoclastogenesis and is thought to play a role in the bone loss that follows estrogen withdrawal. In vitro studies have demonstrated that IL-6 is produced in response to PTH by cells in the osteoblast lineage and that PTH-induced bone resorption is inhibited by a neutralizing antibody to the IL-6 receptor. In addition, we have recently reported that IL-6 plays a role in PTH-induced bone resorption in humans with chronic PTH excess and in experimental animals during the short-term infusion of PTH. In the current study, we examined whether estrogen withdrawal augments PTH-induced IL-6 production. When cultured in the absence of estrogen, human osteosarcoma cells (Saos-2) treated with PTH demonstrated significantly greater release of IL-6 than cells grown under estrogen-replete conditions, 30-fold vs. 15-fold (P = 0.005). A similar effect but of lesser magnitude was seen with primary human osteoblasts. In vivo, PTH induced IL-6 production was also increased in the estrogen-deficient state (ovx) such that at the end of a 5-day PTH infusion, the mean circulating level of IL-6 was significantly higher in ovx vs. sham/ovx mice (60.1 vs. 16.9 pg/ml; P < 0.0001). The greater increase in circulating levels of IL-6 in PTH treated ovx mice was paralleled by a greater rise in bone resorption markers with the mean level of urine collagen cross-links in the PTH-treated ovx group being more than 2.5-fold higher than in the PTH-treated sham/ovx animals (236 vs. 88.5 microg/mmol creatinine, P < 0.0001). Mean serum collagen cross-link values were 17.4 microg/liter in PTH-treated ovx vs. 7.4 microg/liter in PTH-treated sham/ovx animals (P < 0.0001). Treatment of animals with estrogen prevented the exaggerated response to PTH infusion such that the increase in both circulating levels of IL-6 and bone turnover markers in estrogen-treated animals were similar to those observed in sham/ovx animals and significantly lower than those in PTH treated ovx animals. These findings may help to explain the increased skeletal sensitivity to the resorbing effects of PTH seen in the estrogen-deficient state. PMID- 10875255 TI - Autocrine interactions of keratinocyte growth factor, hepatocyte growth factor, and kit-ligand in the regulation of normal ovarian surface epithelial cells. AB - Ovarian tumors are primarily derived from the layer of epithelium surrounding the ovary termed the ovarian surface epithelium (OSE). Although extensive research has focused on established ovarian tumors, relatively little is known about the normal biology of the OSE that gives rise to ovarian cancer. The local expression and actions of growth factors are likely involved in both normal and tumorigenic OSE biology. The current study investigates the expression and action of keratinocyte growth factor (KGF), hepatocyte growth factor (HGF), and kit-ligand (KL) in normal ovarian surface epithelium (OSE). The actions of various growth factors on KGF, HGF, and KL expression are examined. Observations indicate that freshly isolated normal OSE express the genes for KGF, HGF, and KL and expression is maintained in vitro. KGF messenger RNA expression in OSE was found to be stimulated by KGF and HGF, but not KL. HGF expression in OSE was found to be stimulated by KGF, HGF, and KL. KL expression in OSE was also found to be stimulated by KGF, HGF, and KL. Therefore, the various growth factors can regulate the mRNA expression of each other in OSE. Effects of growth factors on OSE growth were examined. KGF, HGF, and KL stimulated OSE growth to similar levels as the positive control epidermal growth factor. Observations suggest that KGF, HGF, and KL interact to promote OSE growth and growth factor expression. The ability of these growth factors to interact in a positive autocrine feedback loop is postulated to be important for normal OSE biology. Paracrine interactions with the adjacent stromal cells will also be a factor in OSE biology. Abnormal interactions of these growth factors may be involved in the onset and progression of ovarian cancer. PMID- 10875256 TI - Negative regulation of the antimetastatic gene Nm23-H1 by thyroid hormone receptors. AB - Metastasis of various malignant cells is inversely related to the abundance of the Nm23-H1 protein. The possible role of thyroid hormones in tumor metastasis has now been investigated by examining the effect of T3 on the expression of the Nm23-H1 gene. Human hepatoma HepG2 cells, in which endogenous thyroid hormone receptor subtype alpha1 (TRalpha1) is expressed at a low level, were stably transfected, either with expression plasmids encoding wild-type TRalpha1 or a dominant negative mutant of TRalpha1, or with the empty vector (yielding HepG2 Wt, HepG2-Mt, and HepG2-Neo cells, respectively). Immunoblot analysis revealed that exposure of HepG2-Wt and HepG2-Neo cells, but not HepG2-Mt cells, to T3 induced time-dependent decreases in the abundance of Nm23-H1 messenger RNA and protein, with the extent of these effects correlating with the level of expression of TRalpha1. An in vitro assay also revealed that T3 induced a marked increase in the invasive activity of HepG2-Wt cells; it induced a smaller increase in that of HepG2-Neo cells but had no effect on that of HepG2-Mt cells. Finally, the promoter region of Nm23-H1 spanning nucleotides -471 to -437 (relative to the transcriptional initiation site) inhibited the expression of a downstream reporter gene, in a T3-dependent manner, in COS-1 cells also transfected with an expression plasmid encoding TRalpha1 or TRbeta1. The DNA binding domain of TRbeta1 was required for this inhibitory effect. These results indicate that T3, acting through TRs, inhibits transcription of Nm23-H1, and that this effect is mediated by a negative regulatory element in the promoter region of the gene. Thus, it is possible that T3 promotes tumor metastasis by inducing down-regulation of Nm23-H1 expression. PMID- 10875257 TI - 1alpha,25-dihydroxyvitamin D3 inhibits prostate cancer cell growth by androgen dependent and androgen-independent mechanisms. AB - We recently reported that 1alpha,25-dihydroxyvitamin D3 [1,25-(OH)2D3] inhibits the growth of the LNCaP human prostate cancer cell line by an androgen-dependent mechanism. In the present study we examined the actions and interactions of 1,25 (OH)2D3 and the androgen 5alpha-dihydrotestosterone (DHT) on two new human prostate cancer cell lines (MDA), MDA PCa 2a and MDA PCa 2b. Scatchard analyses revealed that both cell lines express high affinity vitamin D receptors (VDRs) with a binding affinity (Kd) for [3H]1,25-(OH)2D3 of 0.1 nM. However, the MDA cell lines contain low affinity androgen receptors (ARs) with a Kd of 25 nM for [3H]DHT binding. This is 50-fold lower than the AR in LNCaP cells (Kd = 0.5 nM). Their response to DHT is greatly reduced; 2a cells do not respond to 100 nM DHT, and 2b cells show a modest response at that high concentration. 1,25-(OH)2D3 causes significant growth inhibition in both MDA cell lines, greater (for 2b cells) or lesser (for 2a cells) than that in the LNCaP cell line. Moreover, 1,25 (OH)2D3 significantly up-regulates AR messenger RNA in all three cell lines, as shown by Northern blot analysis. The growth inhibitory effect of 1,25-(OH)2D3 on LNCaP cells is blocked by the pure antiandrogen, Casodex, as we previously reported. However, Casodex (at 1 microM) did not block the antiproliferative activity of 1,25-(OH)2D3 in MDA cells. In conclusion, the growth inhibitory action of 1,25-(OH)2D3 in the MDA cell lines appears to be androgen independent, whereas the actions of 1,25-(OH)2D3 in LNCaP cells are androgen dependent. Most importantly, the MDA cell lines, derived from a bone metastasis of human prostate carcinoma, remain sensitive to 1,25-(OH)2D3, a finding relevant to the therapeutic application of vitamin D and its low calcemic analogs in the treatment of advanced prostate cancer. PMID- 10875258 TI - A targeted partial invalidation of the insulin-like growth factor I receptor gene in mice causes a postnatal growth deficit. AB - The insulin-like growth factor (IGF) system is a major regulator of somatic growth in vertebrates. Both ligands (IGF-I and IGF-II) signal via the same IGF receptor (IGF-IR). Classical IGF-IR invalidation is lethal at birth, so that conditional models are needed to study the postnatal role of this receptor. To establish a genetically inducible invalidation of IGF-IR, we targeted the IGF-IR gene using a construct that introduced a neomycin resistance cassette into intron 2, leaving the rest of the gene intact. This neomycin resistance cassette interfered with the processing of the primary transcript, resulting in there being 12% fewer IGF-binding sites at the cell surface in heterozygous mice and 41% fewer in homozygous mice. Hetero- and homozygous offspring grew more slowly than their wild-type littermates. This difference was noticeable from 4 weeks after birth and was significant from 5 weeks after birth in males. In females, the effect on postnatal growth of insertion of the neo cassette was not significant. In males, IGF-I levels increased moderately (+26%) but significantly, indicating effective feedback regulation of the IGF system. IGF binding protein-4 (IGFBP-4) levels, estimated by Western ligand blotting, were low in homozygotes (-38%), whereas IGFBP-1, -2, and -3 levels were unaffected. In females, IGF-I and IGFBP-1, -2, -3, and -4 levels did not differ significantly among heterozygous, homozygous, and wild-type animals. We investigated the molecular mechanism involved and characterized two RNA-splicing events that could account for the decrease in IGF-IR. The phenotype of these mice developed exclusively postnatally, and body proportions were maintained. IGF-IRneo mice constitute a new model for human postnatal growth deficiency. PMID- 10875260 TI - Luteinizing hormone induces mouse vas deferens protein expression in the murine ovary. AB - The aim of our study was to isolate and identify novel proteins that are involved in the process of ovulation. To achieve this goal we used the technique of proteome analysis. Comparison of ovary protein patterns, obtained by high resolution two-dimensional gel electrophoresis from recombinant FSH (rFSH)- and rFSH + human CG (hCG)-treated mice, showed significant differences in protein spot positions and intensities. Subsequent analysis of one of these proteins was performed by mass spectrometry, resulting in the identification of the mouse vas deferens protein (MVDP). MVDP, which was absent in the two-dimensional gel electrophoresis protein pattern of rFSH-primed mice and appeared 3 h after the hCG surge, is a member of the aldo-keto reductase superfamily and was originally identified in the mouse vas deferens. This is the first study describing MVDP expression and regulation by LH in the ovary. Northern blot analysis of female mice tissues showed that mvdp messenger RNA (mRNA) was only present in adrenal glands and in hCG-treated ovaries. In situ hybridization studies localized the mvdp mRNA unequivocally to ovarian thecal and interstitial cells with an expression profile starting already 1.5 h, and decreasing 24 h, after LH treatment. In the adrenal glands, mvdp mRNA was not regulated by LH and localized in the cells of the zona fasciculata. In murine adrenocortical cells, a recent study proposed a detoxifying role of MVDP. MVDP might fulfill the same function in the ovary; however, because of its strong and early transcriptional induction by LH, it is also possible that MVDP catalyses another important step during the cascade of events occurring at the time of ovulation. PMID- 10875259 TI - Hepatocyte growth factor and vitamin D cooperatively inhibit androgen unresponsive prostate cancer cell lines. AB - Expression of MET, the receptor for hepatocyte growth factor (HGF), has been associated with androgen-insensitive prostate cancer. In this study we evaluated MET activation by HGF and HGF action in prostate cancer cell lines. HGF causes phosphorylation (activation) of the MET receptor in three androgen-unresponsive cell lines (DU 145, PC-3, and ALVA-31) together with morphological change. Although HGF is known to stimulate the growth of normal epithelial cells, including those from prostate, we found that HGF inhibited ALVA-31 and DU 145 (hormone-refractory) cell lines. Moreover, HGF and vitamin D additively inhibited growth in each androgen-unresponsive cell line, with the greatest growth inhibition in ALVA-31 cells. Further studies in ALVA-31 cells revealed distinct cooperative actions of HGF and vitamin D. In contrast to the accumulation of cells in G1 seen during vitamin D inhibition of androgen-responsive cells (LNCaP), growth inhibition of the androgen-unresponsive ALVA-31 cell line with the HGF and vitamin D combination decreased, rather than increased, the fraction of cells in G1, with a corresponding increase in the later cell cycle phases. This cell cycle redistribution suggests that in androgen-unresponsive prostate cancer cells, HGF and vitamin D act together to slow cell cycle progression via control at sites beyond the G1/S checkpoint, the major regulatory locus of growth control in androgen-sensitive prostate cells. PMID- 10875261 TI - The two tumor necrosis factor receptors mediate opposite effects on differentiation and glucose metabolism in human adipocytes in primary culture. AB - Tumor necrosis factor-alpha (TNF) inhibits fat cell differentiation and may also mediate insulin resistance in adipocytes. Both TNF receptors are expressed in adipose tissue, but it is unknown how both receptors are involved in these biological functions. We therefore studied the effect of receptor-specific TNF muteins on adipose differentiation and insulin-stimulated glucose transport of in vitro differentiated human adipocytes in primary culture. Adipocyte precursor cells exposed to the 60-kDa TNF receptor (p60-TNFR)-specific TNF(R32W-S86T) showed a marked decrease in the percentage of differentiating cells in response to adipogenic factors as well as a reduction in peroxisome proliferator-activated receptor-gamma2 (PPARgamma2) messenger RNA (mRNA) and glycerophosphate dehydrogenase (GPDH) activity, but increased endogenous TNF mRNA expression. When cells were incubated with the p80-TNFR-specific TNF(D143N-A145R), adipogenesis and PPARgamma2 mRNA expression were stimulated, GPDH activity was unchanged, and TNF mRNA was completely suppressed. Insulin-stimulated 2-deoxy-D-glucose transport was inhibited by both muteins. The p60-TNFR-mediated inhibition increased continuously during 6 h of treatment and was associated with a down regulation of glucose transporter-4 (GLUT4) mRNA and GLUT4 protein, whereas the p80-TNFR-specific mutein caused a transient increase in GLUT4 mRNA, but did not alter GLUT4 protein expression after a 24-h incubation. We conclude that p60-TNFR mediates the antiadipogenic effect as well as the down-regulation of GLUT4 by TNF, thereby leading to long-term inhibition of insulin-stimulated glucose transport. In contrast, activation of the p80-TNFR induces an adipogenic effect and transiently up-regulates GLUT4 expression. Here, the acute inhibition of insulin-stimulated glucose transport may be induced by interference with the insulin signaling pathway. PMID- 10875262 TI - Subcellular distribution of phospholipase C isoforms in rodent pancreas and gastric mucosa. AB - Phosphoinositide-specific phospholipase C (PLC) has been implicated as a participant in cell proliferation as well as enzyme and hormone secretion. Defining the subcellular distribution of PLC isoforms would possibly contribute to further understanding of their function. We investigated the intracellular distribution of four PLCs (beta1, beta2, beta3, and gamma1) in mouse pancreatic cells as well as mouse and rat gastric mucosa cells by ultrastructural immunocytochemistry. In pancreatic acinar cells, PLCbeta1 and PLCgamma1 were demonstrated in the zymogen granules while PLCbeta2 was present in the granulae as well as the endoplasmic reticulum (ER), and PLCbeta3 was prominent in the ER. In the endocrine pancreas, PLCbeta2 immunolabeling was expressed in the secretory granulae of alpha, beta, delta, and pancreatic polypeptide cells. PLCbeta3 showed a slight labeling in the nucleus and ER of all four pancreatic endocrine cell types while PLCgamma1 was prominent in alpha cell granulae. In the gastric mucosa cells, PLCbeta2 was highly expressed in the heterochromatin areas and in the ER of parietal, chief, mucous, and enterochromaffin-like cells. PLCbeta3 were expressed in a manner similar to PLCbeta2 in those cells; however, no immunoreaction was seen in the ER of parietal cell. PLCgamma1 was demonstrated in the chief cell granulae. One possible, although yet speculative, interpretation of our results is that the studied PLC isoforms may be involved in processing in pancreatic secretory granulae and that nuclear PLCbeta2 and PLCbeta3 signaling pathways may be operative in the cells of the gastric mucosa. PMID- 10875263 TI - Release of guanylin immunoreactivity from the isolated vascularly perfused rat colon. AB - The intestinal peptide guanylin regulates the electrolyte/water transport in the intestinal epithelium. The aim of the present study was to investigate the mechanisms that modulate its secretion in the isolated vascularly perfused rat colon by using a specific guanylin RIA. Intraarterial infusion of bethanechol (10(-4) M) or bombesin (10(-7) M) elicited a significant 6-fold increase in the release of guanylin immunoreactivity (G-IR) in the lumen. Bombesin-stimulated G IR secretion was strongly reduced by tetrodotoxin, whereas atropine had no effect. VIP (10(-7) M) induced a moderate release of G-IR, whereas substance P, calcitonin gene-related peptide, peptide YY, somatostatin, and neurotensin were without effect. Dimethyl-PGE2 (1.4 x 10(-5) M) or interleukin-1beta (2.5 x 10( 10) M) induced a 3-fold increase in G-IR in the lumen, whereas the degranulator compound bromolasalocid did not stimulate guanylin secretion. Forskolin (10(-5) M) or sodium nitroprusside (10(-4)-10(-3) M) induced a significant release of G IR. In contrast, PMA (10(-7) M) or ionophore A23187 (10(-6) M) did not modify basal secretion of G-IR. Upon stimulation of guanylin release with bombesin or bethanechol, an increase in G-IR in the portal effluent was also detected. The release of G-IR in the portal effluent was 40-fold lower than that of G-IR into the luminal perfusate. Additionally, analysis with gel chromatography revealed that the immunoreactive material released in the lumen or in the portal effluent coeluted with the 15-amino acid peptide originally isolated from rat intestine. In conclusion, the present data suggest that the enteric nervous system and immune cells may modulate guanylin release from the rat colon. The release of guanylin in the lumen and portal effluent suggests that this peptide may exert both luminal/paracrine and hormonal effects. PMID- 10875264 TI - Structure and expression of a membrane component of the inhibin receptor system. AB - The purification and cloning of a membrane-anchored proteoglycan with affinity for inhibin A are described. Bovine pituitary membranes were isolated, and membrane-anchored proteins were solubilized and used as an enriched source of inhibin binding protein. The extract was passed over an inhibin A affinity column, and a protein, designated p120, was identified as an inhibin-binding moiety. A partial amino acid sequence was determined for the protein, which matched two human complementary DNAs (cDNAs) in the database. The full-length cDNA predicts a 1336-amino acid glycoprotein. Full-length p120-encoding cDNAs were isolated from human testis RNA and cloned into expression vectors. Two p120 messenger RNA transcripts of 4.6 kb and 2 kb are detected in rat pituitary by RNA blot analysis. Similar analysis of rat testis RNA revealed transcripts of identical molecular mass, albeit at lower abundance. To determine the cellular localization of p120 in pituitary and testis, an antibody directed against the predicted extracellular domain of the protein was generated and used in an immunohistochemical analysis of thin tissue sections. p120 immunostaining is coincident with FSHbeta immunopositive gonadotrope cells in rat pituitary. p120 staining is intense in the testicular Leydig cells, which bind iodinated inhibin but not iodinated activin. In summary, an inhibin-binding protein has been isolated that is produced in tissues that are targets of inhibin action. PMID- 10875265 TI - Assessment of growth parameters and life span of GHR/BP gene-disrupted mice. AB - GH has many biological roles, including promotion of growth. Most, if not all, of its roles are achieved through interaction with its receptor. We chose to study the effects of loss of GH signaling on growth and aging in a mouse model for Laron Syndrome (LS) in which the GHR/BP gene has been disrupted. We observed that mice homozygous for the disruption (-/-) were significantly smaller than normal wild-type (+/+) mice as well as mice heterozygous for the disruption, even at 1.5 yr of age. IGF-I levels were also significantly lower in the -/- mice and remained low as the mice aged. IGFBP-3 levels were severely reduced in the -/- mice, whereas IGFBP-1, -2, and -4 levels remained unchanged. Finally, the -/- mice lived significantly longer than +/+ and +/- mice. The latter result contradicts the anti-aging GH data and suggests the need for further analysis of GH and aging. PMID- 10875266 TI - An age-related ovarian phenotype in mice with targeted disruption of the Cyp 19 (aromatase) gene. AB - With the development of a mouse model of estrogen insufficiency due to targeted disruption of the aromatase gene [the aromatase knockout (ArKO) mouse], a new opportunity exists to examine the role of estrogen in ovarian follicular development. Ovaries and serum were collected from wild-type, heterozygous, and ArKO mice at 10-12 and 21-23 weeks and 1 yr of age. The ovaries were assessed histologically and stereologically, with primary, secondary, and antral follicles and corpora lutea counted. The uteri were hypoestrogenic, and serum levels of LH and FSH in ArKO females were elevated above those in heterozygote and wild-type animals at all ages studied. Although estrogen was not a prerequisite for reinitiation of follicle growth, there was a block of follicular development, and no corpora lutea were present in ArKO ovaries. Thus, the ArKO mouse was infertile as a consequence of disrupted folliculogenesis and a failure to ovulate. Hemorrhagic cystic follicles were present by 21-23 weeks of age. The ovarian phenotype degenerated with age, such that by 1 yr there were no secondary or antral follicles, and the primary follicles present were atretic. Extensive interstitial tissue remodeling occurred, exemplified by an influx of macrophages and collagen deposition, coincident with the loss of follicles. In conclusion, the ovarian environment in ArKO mice does not allow the characteristic development of follicles that culminates in ovulation and demonstrates an in vivo requirement of estrogen for normal ovarian function in the mouse. PMID- 10875267 TI - Expression of the leptin receptor during germ cell development in the mouse testis. AB - Leptin, a recently identified hormonal product of the ob gene, is known to regulate appetite, body metabolism, and reproductive functions. We investigated the expression of the leptin receptor (Ob-R) in testes from different age groups. The messenger RNA for Ob-R was found in testes from all age groups using RT-PCR. Using immunohistochemistry, we observed age- and stage-dependent distribution of the Ob-R in mouse testis. In testis of 5-day-old mice, its expression was mainly in type A spermatogonia. In the 20- and 30-day-old testis, Ob-R expression was in the spermatocytes; in the adult testis, it was specific to spermatocytes in stages IX and X of the cycle of the seminiferous epithelium. Five main immunoreactive proteins were detected using Western blot (220, 120, 90, 66, and 46 kDa). The 120-kDa protein was evident only in 20-day-old and older testes, whereas the 90-kDa band was present only in the 5- and 10-day-old testis. Leptin treatment induced phosphorylation of signal transducer and activator of transcription-3 in cultured seminiferous tubules from adult and 5-day-old testes. Our results show for the first time age- and stage-specific localization of a functional Ob-R in testicular germ cells. We hypothesize a direct role for leptin, through phosphorylation of signal transducer and activator of transcription-3, in proliferation and differentiation of germ cells, which may partially explain the infertility observed in leptin-deficient mice. PMID- 10875268 TI - Localization of 17beta-hydroxysteroid dehydrogenase/17-ketosteroid reductase isoform expression in the developing mouse testis--androstenedione is the major androgen secreted by fetal/neonatal leydig cells. AB - The final step in the biosynthesis of testosterone is reduction of androstenedione by the enzyme 17beta-hydroxysteroid dehydrogenase/ 17-ketosteroid reductase (17betaHSD/17KSR). In this study, we have examined expression of the four known reductive isoforms of 17betaHSD/ 17KSR (types 1, 3, 5, and 7) in the developing mouse testis and have determined changes in the localization of isoform expression and testosterone secretion during development. Using RT-PCR isoforms 1, 3, and 7 were shown to be expressed in the seminiferous tubules of neonatal testis, whereas isoforms 3 and 7 were expressed in the interstitial tissue of the adult testis. The type 7 isoform is unlikely to be involved in androgen synthesis and further study concentrated on the type 3 isoform. Developmentally, isoform type 3 was expressed in the seminiferous tubules up to day 10, showed little or no expression on day 20 and from day 30 was confined to the interstitial tissue. In situ hybridization confirmed that the type 3 isoform was expressed only in the seminiferous tubules in fetal testes and in the interstitial tissue in adult testes. In accordance with the localization of enzyme messenger RNA expression 17-ketosteroid reductase enzyme activity was very low in isolated interstitial tissue from neonatal testes while interstitial tissue from adult testes showed high activity. Seminiferous tubules from both neonatal and adult testes showed high levels of enzyme activity. The major androgen secreted by the interstitial tissue of prepubertal animals was androstenedione up to day 20 while 5alpha-androstanediol and/or testosterone were the major androgens secreted from day 30 onwards. These results show that fetal Leydig cells do not express significant levels of a reductive isoform of 17betaHSD/ 17KSR and that androstenedione is the major androgen secreted by these cells. Production of testosterone up until puberty is dependent upon 17betaHSD/17KSR activity in the seminiferous tubules--a "two cell" requirement for testosterone synthesis. Expression of the 17betaHSD/17KSR type 3 isoform (the main reductive isoform in the testis) declines in the seminiferous tubules before puberty but then reappears in the developing adult Leydig cell population. PMID- 10875269 TI - Porcine ovarian cells express messenger ribonucleic acids for the acid-labile subunit and insulin-like growth factor binding protein-3 during follicular and luteal phases of the estrous cycle. AB - It has recently been shown that, in follicular fluid, as in the circulation, insulin-like growth factors (IGFs)-I and -II exist in a ternary complex with IGF binding protein-3 (IGFBP-3) and the acid-labile subunit (ALS). The current study was designed to determine whether ovarian follicular and luteal cells could synthesize IGFBP-3 and ALS. Ovaries were collected, during the follicular and early luteal phases, from mature pigs whose cycles were synchronized with PGF2alpha. We studied IGFBP-3 and ALS messenger RNA (mRNA) by in situ hybridization. These transcripts were colocalized with aromatase mRNA, a marker of healthy granulosa cells. IGFBP-3 mRNA was equally expressed in granulosa cells of all growing follicles. In contrast, granulosa cell ALS mRNA levels were higher (P < 0.05) in preantral and small antral follicles than in large antral follicles. In thecal cells, expression of mRNA for IGFBP-3, ALS and cyclin D1 (a marker of cell proliferation) was restricted to healthy (aromatase-expressing) follicles. In those follicles, thecal expression of IGFBP-3 mRNA was low or absent in preantral follicles but increased (P < 0.05) in antral follicles. Thecal cell ALS transcripts peaked in small antral follicles (P < 0.05) and then declined. In granulosa cells of atretic follicles, transcripts for aromatase were greatly reduced, whereas IGFBP-3 mRNA levels remained high. In contrast, ALS transcript levels were greatly reduced in both granulosa (P < 0.05) and thecal cells (P < 0.001) of atretic follicles. After ovulation, IGFBP-3 mRNA was moderately expressed in granulosa luteins but strongly detected in a few theca derived cells and in vascular endothelial cells. This study demonstrates that follicular fluid IGFBP-3 and ALS, like the IGFs, originate (at least in part) from the ovary. The ability of follicular cells to synthesize, assemble, and store all components of the ternary complex may be critical in determining the bioavailability of follicular IGF-I and -II. PMID- 10875270 TI - Messenger ribonucleic acids for MAC25 and connective tissue growth factor (CTGF) are inversely regulated during folliculogenesis and early luteogenesis. AB - Cell proliferation, terminal differentiation, and angiogenesis occur during cycles of follicular and luteal development. In other paradigms, mac25, a potent tumor inhibitor is strongly induced in senescent epithelial cells, whereas CTGF stimulates angiogenesis and wound healing. Using in situ hybridization and immunohistochemistry, we have examined the possibilities that mac25 is inhibited, whereas CTGF is induced during active periods of follicular development and luteogenesis. Ovaries were collected during the follicular and early luteal phases from prostaglandin F2alpha-treated mature pigs and from slaughterhouse sows. CTGF transcripts were induced during the late preantral stage in granulosa and theca cells concomitantly with the appearance of endothelial cells in the theca. CTGF mRNA expression increased in granulosa cells to a maximum (P < 0.01) in mid-antral follicles but was down regulated (P < 0.01) in preovulatory follicles. In contrast, granulosa cell mac25 mRNA expression was undetectable between the preantral and mid-antral stage but was strongly induced in terminally differentiated granulosa cells of preovulatory follicles. CTGF mRNA and peptide were also detected in the theca externa/interstitium and in vascular endothelial cells of ovarian blood vessels, whereas mac25 transcripts, which were also abundant in ovarian blood vessels increased in the theca interna with follicular development. Transcripts of cyclin D 1, a marker of cell proliferation, appeared during the early antral stage and were moderate in granulosa cells but abundant in capillary endothelial cells in the theca interna, underneath the basement membrane. Following ovulation, CTGF and cyclin D1 mRNAs were associated with the migration of endothelial cells into the CL. Subsequently, there was a marked up regulation of CTGF mRNA expression in granulosa luteins concomitantly with an increase in endothelial cell proliferation within the CL. We hypothesize that CTGF may promote ovarian cell growth and blood vessel formation during follicular and luteal development whereas mac25, a tumor inhibitor, may promote terminal differentiation of granulosa cells in preovulatory follicles. PMID- 10875271 TI - Deficient mineralization of intramembranous bone in vitamin D-24-hydroxylase ablated mice is due to elevated 1,25-dihydroxyvitamin D and not to the absence of 24,25-dihydroxyvitamin D. AB - The 25-hydroxyvitamin D-24-hydroxylase enzyme (24-OHase) is responsible for the catabolic breakdown of 1,25-dihydroxyvitamin D [1,25(OH)2D], the active form of vitamin D. The 24-OHase enzyme can also act on the 25-hydroxyvitamin D substrate to generate 24,25-dihydroxyvitamin D, a metabolite whose physiological importance remains unclear. We report that mice with a targeted inactivating mutation of the 24-OHase gene had impaired 1,25(OH)2D catabolism. Surprisingly, complete absence of 24-OHase activity during development leads to impaired intramembranous bone mineralization. This phenotype was rescued by crossing the 24-OHase mutant mice to mice harboring a targeted mutation in the vitamin D receptor gene, confirming that the elevated 1,25(OH)2D levels, acting through the vitamin D receptor, were responsible for the observed accumulation of osteoid. Our results confirm the physiological importance of the 24-OHase enzyme for maintaining vitamin D homeostasis, and they reveal that 24,25-dihydroxyvitamin D is a dispensable metabolite during bone development. PMID- 10875272 TI - Maternal exposure to octylphenol suppresses ovine fetal follicle-stimulating hormone secretion, testis size, and sertoli cell number. AB - We have tested the hypothesis that maternal exposure to octylphenol, a putative endocrine disrupting chemical, will suppress gonadotropin secretion with a concomitant decrease in testis size and Sertoli cell number during fetal life in the lamb. In Exp 1, pregnant ewes received a continuous iv infusion of diethylstilbestrol (DES; 50 microg/kg x day), octylphenol (1000 microg/kg x day), or vehicle (1:4, alcohol-saline) from days 110-115 of gestation. The fetuses were chronically catheterized in utero, and blood samples were collected every 8 h to monitor gonadotropin secretion. In Exp 2, pregnant ewes received twice weekly sc injections of DES (0.5 microg/kg x day), octylphenol (1000 microg/kg x day), or corn oil from day 70 of gestation to birth. The pituitary gland and testes were collected from the lambs at the end of the treatment period. In Exp 1, maternal exposure to octylphenol suppressed (P < 0.05) FSH concentrations without any effect (P > 0.05) on LH concentrations compared with those in control fetuses. In Exp 2, long-term maternal exposure to octylphenol or a 1000-fold lower dose of DES suppressed (P < 0.05) FSH, messenger RNA levels and the number of FSHbeta immunopositive cells in the pituitary gland and reduced testis weight and the number of Sertoli cells in the testis compared with those in control lambs. We conclude that maternal exposure to octylphenol inhibits the secretion of FSH in the fetus with a concomitant decrease in testis size and Sertoli cell number at birth. PMID- 10875273 TI - Targeted overexpression of insulin-like growth factor I to osteoblasts of transgenic mice: increased trabecular bone volume without increased osteoblast proliferation. AB - Insulin-like growth factor I (IGF-I) is an important growth factor for bone, yet the mechanisms that mediate its anabolic activity in the skeleton are poorly understood. To examine the effects of locally produced IGF-I in bone in vivo, we targeted expression IGF-I to osteoblasts of transgenic mice using a human osteocalcin promoter. The IGF-I transgene was expressed in bone osteoblasts in OC IGF-I transgenic mice at high levels in the absence of any change in serum IGF-I levels, or of total body growth. Bone formation rate at the distal femur in 3 week-old OC-IGF-I transgenic mice was approximately twice that of controls. By 6 weeks, bone mineral density as measured by dual energy x-ray, and quantitative computed tomography was significantly greater in OC-IGF-I transgenic mice compared with controls. Histomorphometric measurements revealed a marked (30%) increase femoral cancellous bone volume in the OC-IGF-I transgenic mice, but no change in the total number of osteoblasts or osteoclasts. Transgenic mice also demonstrated an increase in the osteocyte lacunea occupancy, suggesting that IGF I may extend the osteocyte life span. We conclude that IGF-I produced locally in bone osteoblasts exerts its anabolic effect primarily by increasing the activity of resident osteoblasts. PMID- 10875274 TI - Ontogeny of corticosterone-inducible growth hormone-secreting cells during chick embryonic development. AB - We reported that corticosterone administration into the albumen of fertile chicken eggs on embryonic day (e) 11 induces an increase in the population of GH secreting cells. The present study evaluated the ontogeny, dose response, localization, and persistence of the glucocorticoid-induced increase in the somatotroph population during chicken embryonic development. Corticosterone (0, 0.02, 0.2, and 2 microg in 300 microl saline) was injected into separate eggs on e9, e10, e11, and e12, and the population of GH-secreting cells was assessed 2 days later using reverse hemolytic plaque assays. Corticosterone treatment on e9 or e10 was unable to increase the population of GH-secreting cells on e11 or e12. In contrast, 0.2 and 2 microg of corticosterone on e11 increased the population of GH-secreting cells on e13 (P < 0.05, n = 3 experiments) to 8.2 +/- 0.6 and 6.4 +/- 0.5% of all cells, respectively, relative to controls (2.4 +/- 0.2%). For e14 embryos treated on e12, only the 2 microg dose increased the proportion of GH secreting cells (6.4 +/- 0.6%) relative to controls (3.6 +/- 0.4%). In a second experiment, 0, 0.02, 0.2, 2, and 20 microg of corticosterone were injected on e0, e8, e9, e10, e11, and e12, and the population of GH-secreting cells was assessed on e13 in all groups. No dose of corticosterone was effective when given on e0, e8, e9, or e10. The 0.2 microg and 2 microg doses increased the population of GH secreting cells (7.6 +/- 0.9% and 6.7 +/- 0.8%, respectively) relative to controls (2.3 +/- 0.4%) when injected on e11 (P < 0.05, n = 4 experiments). The 2 microg dose also increased GH cell abundance when injected on e12 (5.6 +/- 0.4%), relative to controls (2.7 +/- 0.5%). Treatment with 20 microg on e11 and e12 induced the greatest responses (10.3 +/- 1.1% and 8.7 +/- 0.9%, respectively). However, in subsequent experiments, administration of 20 microg on e11 resulted in embryonic death by e18. In a third set of experiments, two groups of eggs were injected either with 2 microg of corticosterone in saline or saline alone on e11, and the number of GH-secreting cells was estimated on e13, e16, e19, and the day of hatch (d1). The population of GH-secreting cells in corticosterone treated embryos was significantly higher than in saline treated embryos only on e13 (7.1 +/- 0.8% and 2.7 +/- 0.3%, respectively). No significant differences were observed on e16 (12.4 +/- 1.5% and 13.6 +/- 1.2%), e19 (19.0 +/- 1.0% and 18.2 +/ 1.7%) and d1 (23.8 +/- 2.1% and 25.1 +/- 1.8%) between corticosterone treated and control embryos, respectively. In a fourth set of experiments, whole mount in situ hybridization indicated that injection of corticosterone on e11 induced GH messenger RNA expression in the caudal part of the pituitary gland on e13, where somatotrophs are located normally later in development. We conclude that corticosterone administration in ovo can increase the population of GH-secreting cells in the caudal anterior pituitary only during a small window of development between e11 and e13 and that this premature increase of GH-secreting cells does not affect the percentage of GH-secreting cells later in development. PMID- 10875275 TI - Rescue of preimplantatory egg development and embryo implantation in prolactin receptor-deficient mice after progesterone administration. AB - PRL, a hormone secreted essentially by the pituitary and other extrapituitary sources such as decidua, has been attributed regulatory roles in reproduction and cell growth in mammals. These effects are mediated by a membrane PRL receptor belonging to the cytokine receptor superfamily. Null mutation of the PRL receptor gene leads to female sterility due to a severely compromised preimplantation development and a complete failure of the implantation of the few embryos reaching the blastocyst stage, strongly implicating PRL in the maternal control of implantation. We measured the hormonal status of -/- mice, which confirmed that the corpus luteum is unable to produce progesterone. Progesterone administration to -/- mice completely rescued the development of preimplantatory eggs and embryo implantation. Pregnancy could be maintained to 19.5 days postcoitum, with about 22% of resulting embryos reaching adulthood. Although progesterone and perhaps PRL appear to facilitate mouse preembryo development throughout the preimplantation stages, other factors as well as a possible direct effect of PRL on the uterus are probably necessary to fully maintain pregnancy. Finally, reduced ductal side-branching in the mammary gland can be rescued by progesterone treatment, but females exhibit reduced alveolar formation. Our model establishes the PRL receptor as a key regulator of reproduction and provides novel insights into the function of lactogenic hormones and their receptor. PMID- 10875276 TI - Dissociation of locomotor activation and suppression of food intake induced by CRF in CRFR1-deficient mice. AB - Corticotropin-releasing factor (CRF) systems are involved in locomotor and feeding behaviors. Two distinct CRF receptor subtypes, CRFR1 and CRFR2, are thought to mediate CRF actions in the central nervous system. However, the role for each receptor in locomotor activity and feeding remains to be determined. Using CRFR1 null mutant mice, the present study examined the functional significance of this receptor in ambulation and feeding. CRF treatment of wild type mice resulted in increased levels of locomotion whereas no change was observed in CRFR1-deficient mice as compared to vehicle-treated mutant mice. In contrast, CRF decreased food-water intake in both wild type and CRFR1-deficient mice equally. These results support an important role for CRFR1 in mediating CRF induced locomotor activation, whereas other receptor subtypes, likely CRFR2, may mediate the appetite-suppressing effects of CRF-like peptides. PMID- 10875277 TI - Galanin-like peptide (GALP) is a target for regulation by leptin in the hypothalamus of the rat. AB - Galanin-like peptide (GALP), which was recently isolated from the porcine hypothalamus, shares sequence homology with galanin and binds with high affinity to galanin receptors. To study the distribution and regulation of GALP-expressing cells in the brain, we cloned a 120 base-pair cDNA fragment of rat GALP and produced an antisense riboprobe. In situ hybridization for GALP mRNA was then performed on tissue sections throughout the forebrain of adult ovariectomized female rats. We found GALP mRNA-containing cells in the arcuate nucleus (Arc), caudal dorsomedial nucleus, median eminence and the pituitary. Because GALP mRNA in the Arc appeared to overlap with the known distribution of leptin receptor mRNA, we tested the hypothesis that GALP expression is regulated by leptin. Using in situ hybridization, we compared the number of GALP mRNA-containing cells among groups of rats that were fed ad lib or fasted for 48 h and treated with either leptin or vehicle. Fasting reduced the number of identifiable cells containing GALP mRNA in the Arc, whereas the treatment of fasted animals with leptin produced a 4-fold increase in the number of cells expressing GALP message. The presence of GALP mRNA in the hypothalamus and pituitary and its regulation by leptin suggests that GALP may have important neuroendocrine functions, including the physiological regulation of feeding, metabolism, and reproduction. PMID- 10875278 TI - Symmetrical black plaques on the toes. PMID- 10875279 TI - Diffuse and progressive papules and nodules. PMID- 10875280 TI - Vegetative incompatibility in filamentous fungi: a roundabout way of understanding the phenomenon. AB - Vegetative incompatibility limits heterokaryon formation in fungi. It results from genetic differences at specific loci (het loci). Characterization of het genes and, more recently, of incompatibility reaction suppressors has, provided insight into the mechanisms involved. A link between development, vegetative incompatibility and signaling pathways has been established in Podospora anserina. PMID- 10875281 TI - Nuclear migration in fungi--different motors at work. AB - Lower fungi such as Saccharomyces cerevisiae and Aspergillus nidulans are ideal organisms for studying the molecular biology underlying nuclear migration in eukaryotic cells. In this review, the role of different motor proteins such as dynein, kinesin and myosin will be discussed. PMID- 10875283 TI - Genetic relatedness analysis of nocardia strains by random amplification polymorphic Dna: validation and applications. AB - Until now, no simple and rapid technique existed for epidemiological study of strains belonging to the Nocardia genus. The application of the arbitrarily primed PCR procedure to generate randomly amplified polymorphic DNA (RAPD) fingerprints for such analysis of Nocardia isolates was investigated. Fifty-one unrelated clinical isolates of N. asteroides were tested. Two conditions of RAPD using two different primers generated RAPD fingerprints that allowed the differentiation of all strains. The patterns were reproducible and discriminating. The results highlight the diversity of N. asteroides species and confirm that RAPD analysis is a highly valuable tool for studying the epidemiology of the Nocardia genus. Several examples describe the advantage of RAPD analysis for establishing the relationship between isolates from a given patient (long-term infections, coinfections) and from different patients (i.e. during an outbreak). In the future, this technique will help us to investigate the source of infection in cases of nosocomial transmission, to understand the outcome of nocardiosis, and to follow the evolution and acquisition of resistance to Nocardia strains. PMID- 10875282 TI - The presence of intragenically located REP-like elements in Bacillus sporothermodurans is sufficient for REP-PCR typing. AB - The technique of repetitive extragenic palindromic polymerase chain reaction (REP PCR) enables the identification and discrimination of clonally related Bacillus sporothermodurans isolates from ultra-high temperature and sterilized milk. The aim of this study was to investigate the genetic basis for the generation of these highly informative REP-PCR patterns. The major 947-bp REP-PCR fragment of B. sporothermodurans was cloned, together with its 5' and 3' flanking sequences. Only partial homology with the REP consensus sequence was established at the borders of the REP-PCR fragment. Moreover, these border sequences were located within two distinct open reading frames, with great homology to the uvrA and uvrB genes of Escherichia coli. The presence of these REP-like elements in B. sporothermodurans was thus sufficient for high resolution REP-PCR typing of this Gram-positive organism. In some cases (and especially with Gram-positives), REP PCR could thus be considered more as an arbitrary primed PCR, albeit at a somewhat higher annealing temperature and using conserved primers. The random priming effect at the less stringent annealing conditions of REP-PCR was also demonstrated for enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) on another Gram-positive organism, Listeria monocytogenes. PMID- 10875284 TI - Genetic diversity of mycobacterium avium recovered from AIDS patients in the caribbean as studied by a consensus IS1245-RFLP method and pulsed-field gel electrophoresis. AB - A total of 45 strains of Mycobacterium avium from 31 human immunodeficiency virus (HIV)-positive patients in the French Caribbean islands and Guiana were subjected to DNA fingerprinting using a recently described consensus IS1245 restriction fragment length polymorphism (RFLP) method, and pulsed-field gel electrophoresis (PFGE). The IS1245-RFLP resulted in three distinct clusters composed of three 27 banded isolates from two patients (cluster A), nine two-banded isolates from six patients (cluster B), and three 20-banded isolates from three patients (cluster C). PFGE results obtained after XbaI and DraI digestions gave similar clustering results irrespective of the enzyme used, and confirmed the molecular clonality for high IS1245 copy number isolates (clusters A and C). However, PFGE further discriminated the low IS1245 copy number cluster B into two distinct subclusters: subcluster I containing six isolates from four patients during the same time period from a single hospital in Guadeloupe, and subcluster IV composed of four isolates from two patients, out of which three isolates were from a single patient (patient 19). Interestingly, in the latter case, PFGE grouped together all three isolates from patient 19 despite the fact that IS1245 fingerprinting permitted grouping only two of the three isolates (the remaining unclustered isolate contained two additional bands of 3.5 and 5 kbp, and was initially considered as evidence of polyclonal infection). A combined numerical analysis of the IS1245-RFLP and PFGE results corroborated the existence of four instead of three clusters. A comparison of IS1245-RFLP versus PFGE results suggested that the standardized RFLP procedure is compatible with PFGE only for M. avium isolates containing > or = 5 copies of IS1245. Consequently, the typing results for low IS1245 copy number isolates (31% of isolates in this study) should be reconsidered for secondary typing using PFGE. Lastly, the absence of a predominant genotype of M. avium infecting HIV-positive patients over a 5-year period in this tropical region argues in favor of a lack of a privileged ecological niche for M. avium, and instead suggests that microepidemics of M. avium may prevail during limited periods of time. PMID- 10875285 TI - Lack of GATC sites in the genome of Streptococcus bovis bacteriophage F4. AB - A strong bias against GATC sites was observed in the genome of phage F4, a lytic Streptococcus bovis bacteriophage. Only three GATC sites were found within the 60.4-kbp genome of this phage. The comparative lack of GATC sequences within the F4 genome was probably not due to dam methylation, as no modification within this site was detected using methylation-sensitive isoschizomer pair restriction endonuclease analysis. The short oligonucleotide composition of available S. bovis DNA sequences suggested the existence of an unknown mechanism for counterselection of GATC sites in S. bovis bacteriophages. PMID- 10875286 TI - Tn5044, a novel Tn3 family transposon coding for temperature-sensitive mercury resistance. AB - We report the discovery and characterization of the mercury resistance transposon, Tn5044, from a Xanthomonas strain from the Kamchatka peninsula. In addition to the standard set of merRTPCAD genes, the mer operon of Tn5044 contains a gene named sigY that encodes the RNA polymerase sigma factor-like protein. Mercury resistance determined by Tn5044 is expressed at low (30 degrees C) but not at elevated temperatures (37 degrees C). None of the mer operon genes downstream of merA is responsible for the temperature-sensitive mercury resistance. The transposition module of Tn5044 is closely related to those of Tn1412 isolated from medical sources and to Tn5563 and ISXc5 from environmental sources. However, Tn5044 differs from these transposons in that it has unusually long terminal inverted repeats. Sequence analysis of the transposase (tnpA) genes places Tn5044 and its close relatives into the Tn3 subgroup of the Tn3 family. However, the orientation of their resolvase and transposase genes is unusual for the Tn3 family: tnpR is proximal to the end of the transposon, while divergently transcribed tnpA is oriented inwardly. The region between tnpA and tnpR genes is unusually large and contains two short conserved open reading frames. In addition to the complete set of sequence motifs common to true resolvases, the resolvase of Tn5044 and its close relatives possesses a C-terminal extension showing no homology to known proteins. Despite this peculiarity, Tn5044 resolvase can resolve cointegrates formed during Tn5044 transposition controlled by tnpA. Genetic data suggest that the extension is essential for TnpR functioning. PMID- 10875287 TI - estA, a gene coding for a cell-bound esterase from Paenibacillus sp. BP-23, is a new member of the bacterial subclass of type B carboxylesterases. AB - Screening of a gene library from Paenibacillus sp. BP-23 generated in Escherichia coli led to identification of a clone that directed the production of lipolytic activity. From the sequencing data, we found an open reading frame encoding a protein of 485 amino acids with an estimated molecular mass of 53 kDa and a pI of 5.1. Absence of a signal peptide indicated that it was a cell-bound protein. Sequence analysis showed that the protein contained the signature G-XI-S-X2-G included in most serine-esterases and lipases. The cloned protein showed high homology with enzymes belonging to the bacterial subclass of type B carboxylesterases. The enzyme had a significant preference for esters of short chain fatty acids and showed the kinetics behaviour of a true esterase. Maximum activity was found at pH 7.5 and 37 degrees C, although the enzyme was active in the pH range 6.0- 9.0 and at temperatures up to 45 degrees C. As expected for a serine-esterase, activity was inhibited by phenylmethylsulphonyl fluoride. PMID- 10875288 TI - Etiology and diagnostic evaluation of macrocytosis. AB - BACKGROUND: Elevation of mean cell volume (MCV) is a common clinical problem, but the etiologic spectrum and optimal diagnostic evaluation of macrocytosis are not well defined. METHODS: We studied 300 consecutive hospitalized adult patients with MCV values > or = 100 fL. Assessment included complete blood counts, morphologic review, liver function tests, and levels of serum cobalamin (Cbl), methylmalonic acid, and total homocysteine. RESULTS: The most common cause of macrocytosis was drug therapy, followed by alcohol, liver disease, and reticulocytosis. Megaloblastic hematopoiesis accounted for less than 10% of cases. MCV values > 120 fL were usually caused by Cbl deficiency. Anisocytosis, macro-ovalocytosis, and teardrop erythrocytes were most prominent in megaloblastic hematopoiesis. Elevated levels of serum methylmalonic acid and total homocysteine were useful in the diagnosis of Cbl deficiency. CONCLUSIONS: Drugs and alcohol are the most common causes of macrocytosis in hospitalized patients in a New York City teaching hospital. We have formulated tentative guidelines for the evaluation of high MCV values in this setting. PMID- 10875289 TI - Prevalence of hepatitis C virus seropositivity among hospitalized US veterans. AB - BACKGROUND: Hepatitis C virus (HCV) is a major cause of acute and chronic hepatitis in the United States and abroad. HCV antibody prevalences ranging from 10 to 90% have been reported in intravenous drug abusers, hemodialysis patients, and persons suffering from other liver diseases, whereas HCV seropositivity rates for volunteer-blood donor populations are generally under 1%. However no information has been available concerning the prevalence of HCV in general hospital populations in the United States. METHODS: We examined the rate of HCV seropositivity in 530 patients admitted to the Atlanta VA Medical Center between November 1993 and November 1994. The test population consisted of 400 random hospital admissions, 100 successive admissions to the surgical service, and 30 random admissions to the gastrointestinal service. Serum samples were assayed for HCV antibodies by a second generation EIA, and all repeat reactives were re examined using a supplemental research assay to confirm the presence of HCV antibodies. Complete chart reviews were carried out on all HCV seropositive patients and on 100 HCV seronegative patients. RESULTS: Sixty-two of the 530 patients tested (11.7%) were repeatedly positive for HCV antibodies. Of these 62 repeat reactives, 56 (90.3%) were positive and 3 others (4.8%) indeterminate by the supplemental assay. The HCV seropositivity rate after supplemental testing was 11.8% for random admissions, 5.0% for surgical admissions, and 13.3% for patients admitted to the gastroenterology service. HCV-associated risk factors in HCV seropositive patients included a history of intravenous drug abuse, current or previous alcohol abuse, previous or concurrent liver disease, previous blood transfusions, hemodialysis, and multiple sex partners or unsafe sex. CONCLUSIONS: HCV infection may be more prevalent among hospitalized VA patients (and among other US hospital populations) than previously expected. PMID- 10875290 TI - Antibiotics and return visits for respiratory illness: a comparison of pooled versus hierarchical statistical methods. AB - BACKGROUND: Antibiotic prescribing for respiratory illness has been associated with small reductions in return visits in an analysis of a large practice-based network. In this study, we apply hierarchical analytical methods that account for the clustering of patients by practices to identify whether antibiotic prescribing by primary care physicians reduces subsequent visits for 6 acute respiratory illnesses-upper respiratory infection, pharyngitis, bronchitis, otitis media, sinusitis, and cough. METHODS: The study data came from 318 family physicians and internists in 45 practices in the Practice Partner Research Network from January 1995 through December 1996, with 255,564 active patients. Patients treated with antibiotics were compared with those who were not on the frequency of revisit within the next 14 days. A simple pooling model and 3 hierarchical statistical models (fixed-effects, random-effects, and Bayesian) were used to compare the odds-ratios for return visits. RESULTS: Statistically significant results were found only for bronchitis and sinusitis by the hierarchical models, but the simple pooling model produced statistically significant results for all study conditions. CONCLUSION: We conclude that antibiotics may reduce return visits for patients with bronchitis and sinusitis, but not for patients with other respiratory illness (upper respiratory infection, pharyngitis, otitis media, or cough). Studies of large clinical databases should use methods of analysis that account for the grouping of patients by practice to avoid false positive associations (type I errors.) PMID- 10875291 TI - Echocardiographic parameters in reversible idiopathic dilated cardiomyopathy. AB - BACKGROUND: Reversible idiopathic dilated cardiomyopathy (IDCM) is a rare entity. It has been hypothesized that the degree of left ventricular end diastolic dilation is an important independent predictor of prognosis. We undertook a study to identify cases of reversible IDCM and to evaluate the echocardiographic findings in these patients. METHODS: We identified 5 patients with IDCM who showed normalization of left ventricular function over a follow up period of 5 months. The findings were compared with those of 10 patients with IDCM who did not show improvement of left ventricular function. The mean (+/-SE) left atrial and left ventricular (LV) dimensions and ejection fraction at baseline and follow up were compared in both groups. RESULTS: There were no statistical differences between the mean (+/-SE) left atrium sizes in the cases and control group at baseline [4.52 (+/-0.24) cm v4.6 (+/-0.13) cm; P = 0.758]. Also, no differences were observed between mean (+/-SE) LV dimensions in diastole and systole in both groups at baseline [LV diastole, 6.72 (+/-0.35) cm versus 6.56 (+/-0.22) cm; P = 0.711; LV systole, 5.6 (+/-0.27) cm versus 5.59 (+/-0.29) cm; P = 0.712] as well as in mean (+/-SE) ejection fraction [24% (+/-3.96) versus 21.7% (+/-3.30); P = 0.623]. CONCLUSION: Based on initial echocardiographic parameters, chamber dimensions and baseline LV ejection fraction are not predictors of reversibility. PMID- 10875292 TI - Safety, feasibility, and efficacy of a resistance training program in preadolescent obese children. AB - BACKGROUND: Safe and effective exercise programs are needed to prevent and treat chronic diseases in childhood. In particular, preadolescent obese children should participate in activities that are specific to their special needs. Resistance or strength training has been prescribed for adult obese persons. Research is limited concerning the use of resistance training in programs that treat obese preadolescents. METHODS: Nineteen treatment subjects (7-12 years of age) were enrolled in a 10-week weight management program which included diet, behavior modification, and aerobic and flexibility exercises. Forty-eight control subjects (7-12 years of age) participated in the diet, behavior modification program, and a thrice-a-week walking program. The efficacy of the overall weight management program was examined by anthropometry at 10 weeks and 1 year. RESULTS: Fifteen treatment subjects completed the 10-week program (retention rate, 78.9%). Thereafter compliance decreased by approximately 33% for the long-term study. Seventeen control subjects completed the program (retention rate, 35%). Weight, percent of ideal body weight, and body mass index were reduced significantly at 10 weeks (P<0.0001) and did not increase significantly at 1-year follow-up in both treatment and control groups. Height increased significantly at 1 year in both treatment and control subjects. In the treatment subjects, percent fat decreased significantly (P<0.001), whereas fat-free mass did not change significantly (P>0.05). CONCLUSIONS: A resistance-training program may be included safely in a multidisciplinary weight management program for obese preadolescent male and female children. The addition of specific exercise regimes such as resistance training may improve program retention especially in severely obese youth. PMID- 10875293 TI - Delayed recovery of thyrotropin responsiveness after radioactive iodine therapy for hyperthyroidism. AB - BACKGROUND: After radioactive iodine therapy for hyperthyroidism, an expected lag in the responsiveness of thyrotropin (TSH) is 60 to 90 days. In our experience, however, many patients seemed to have a more prolonged lag in TSH recovery. METHODS: A retrospective chart review was performed in 58 patients who underwent radioactive iodine therapy for hyperthyroidism (52 with Graves disease, 5 with toxic nodular goiters, and 1 with a toxic adenoma). RESULTS: Forty-nine patients (84%) had appropriate responses of TSH for their level of serum thyroid hormone. Thirty-one became hypothyroid, 12 became euthyroid, and 6 remained hyperthyroid. Nine patients (16%) had a lag in their TSH responsiveness. The TSH remained low for 3 months in 5 patients, for 9 months in 3 patients, and 1 patient had low levels of serum TSH for at least 12 months. CONCLUSIONS: After radioactive iodine therapy for hyperthyroidism, decisions upon further therapy must be based upon the clinical status as well as the serum levels of TSH and thyroid hormones. PMID- 10875294 TI - Normocalcemic hyperparathyroidism in Vietnamese immigrants living in southern California. AB - OBJECTIVE: To characterize the normocalcemic hyperparathyroidism in Vietnamese immigrants living in southern California. METHODS: Of 14 Vietnamese patients with primary hyperparathyroidism who were observed between 1991 and 1996, 50% (7 patients; 2 men and 5 women) had normal and/or fluctuating levels of serum total calcium. When the serum calcium was corrected for the albumin, the "corrected" calcium was lower than the measured serum total calcium. Their mean age was 56.4 +/- 11.4 years. All patients had normal serum levels of albumin and serum phosphate. RESULTS: Women were affected more often than men by a ratio of 5:2. The serum-ionized calcium as well as intact PTH were increased in all patients. Five patients underwent surgery with confirmation of parathyroid adenomas. Two patients refused surgery. They did not have osteitis fibrosa cystica by radiological examination. One patient had low plasma levels of 25-hydroxyvitamin D. Five of 7 normocalcemic patients (70%) were born in the month of December compared with 2 of seven hypercalcemic patients (30%). CONCLUSION: The blood ionized calcium and intact parathyroid hormone are necessary for confirmation of normocalcemic hyperparathyroidism. Most of our normocalcemic hyperparathyroid patients (70%) were born in the month of December. We postulate that a combination of exposure to solar ultraviolet light during the formation of the fetal parathyroid glands and stimulation from low vitamin D levels in the wintertime may be related to the development of hyperparathyroidism. However, it is difficult to prove a definite correlation between normocalcemic hyperparathyroidism and their month of birth (December), especially when these observations were seen in a small group of patients. PMID- 10875295 TI - Acute renal failure in HIV-infected patients: a brief review of common causes. AB - Acute renal failure is a well-described renal syndrome observed in patients infected with the human immunodeficiency virus (HIV). Underlying glomerular disease and disturbances in renal tubular function predispose these patients to a number of hemodynamic and nephrotoxic insults. Prerenal azotemia from both "true" and "effective" depletion of intravascular volume is the most common cause of acute renal insufficiency in patients infected with HIV. Direct damage to the renal tubules from both nephrotoxic medications and prolonged ischemic processes occurs frequently in hospitalized patients. Injury to the tubulointerstitium of the kidney may also result from allergic reactions to medications prescribed to patients. Deposition of crystals in the tubular lumens, and rarely in the glomerular capillaries, will cause acute renal failure in the setting of tumor lysis syndrome or during therapy with medications associated with crystal nephropathy. Finally, obstruction of the urinary system will rarely cause postrenal azotemia in patients infected with HIV. PMID- 10875296 TI - A review of Epstein-Barr virus infection in patients with immunodeficiency disorders. AB - Epstein-Barr virus (EBV), one of 8 known human herpesviruses, infects the vast majority of mankind and infections are generally subclinical. However, EBV infection has been associated with a spectrum of diseases, lymphoproliferative diseases (EBV-LPD) in particular, including malignant lymphoma. EBV-LPD are frequently observed in patients with primary or secondary immunodeficiencies. The incidence of EBV-LPD is on the rise, partly because of increasing numbers and success of hematopoietic stem cell and solid organ transplants and partly because many patients with immunodeficiencies, both primary and secondary, including AIDS, live longer, with improvements in supportive care. Herein, a spectrum of EBV-associated diseases in patients with immunodeficiency are summarized and discussed mainly focusing on their pathogenetic mechanism(s). PMID- 10875297 TI - Restless legs syndrome: clinical features and treatment. AB - Restless legs syndrome (RLS), widely recognized as a definite clinical entity, has an estimated prevalence of 1 to 15% in different ethnic populations. However, it remains an underdiagnosed condition and its symptoms are frequently ascribed to stress and anxiety. Advancement in modern imaging techniques and clinical drug trials provide evidence of an impaired dopaminergic system in RLS. Management involves investigating and correcting treatable secondary causes, avoidance of aggravating factors, and pharmacologic therapy. Recent controlled trials have demonstrated the effectiveness of dopamine agonists such as pramipexole and pergolide. Additional research is needed to further elucidate the pathophysiology of RLS, through obtaining post-mortem specimens and refinement of neuroimaging and neurophysiologic techniques. Isolation of specific genetic loci in familial cases would enable better characterization of distinct clinical and genetic subsets of RLS and result in better understanding of this disease at the molecular level. PMID- 10875298 TI - A 78-year-old woman with leg pain and weakness. PMID- 10875299 TI - Idiopathic pulmonary hemosiderosis with cystic lesions: a rare presentation. AB - This report describes a case of a 49-year-old man with cough, recurrent hemoptysis, and dyspnea during 18 months, presenting with radiological findings of alveolar infiltrate and cystic lesions in left upper lobe. Laboratory studies revealed normocytic hypochromic anemia and normal coagulation tests. C-reactive protein and mucoproteins were negative. Serum protein electrophoresis and complement, urinalysis, serum creatinine, creatinine clearance, and 24-hour urine protein were normal. Tests for antineutrophil cytoplasmic antibodies and anti glomerular-basement membrane antibodies were negative. Tests for connective tissue diseases were all negative. Histological findings were consistent with those of idiopathic pulmonary hemosiderosis. Radiological findings are discussed. PMID- 10875300 TI - Hypoplasia of the inferior vena cava with azygous continuation presenting as recurrent leg deep vein thrombosis. AB - Azygous continuation of the inferior vena cava is a rare congenital malformation. It occurs because of atresia of the retrohepatic segment of the inferior vena cava. The disorder has been described mainly as an incidental finding associated with a number of congenital malformations, including cardiovascular, tracheobronchial, and other visceral malformations. Therapeutic intervention is aimed primarily at accompanying malformations. We report a case of hypoplasia of the inferior vena cava with azygous continuation in a 30-year-old man who presented with recurrent deep venous thrombosis. PMID- 10875301 TI - Acute hepatitis A: combination of the relapsing and the cholestatic forms, two rare variants. AB - Here we present an unusual case of a 23-year-old, otherwise healthy man who had a biphasic form of viral hepatitis A with a combination of two variants, the relapsing and cholestatic forms. One month after resolution of the first phase of acute hepatitis A, he was readmitted with jaundice and intense pruritus. During hospitalization, his serum bilirubin level increased to 50.2 mg/dL, with a slight increase in the other levels of liver enzymes. He was treated with ursodeoxycholic acid and later with corticosteroid therapy, resulting in resolution of symptoms and improvement of his liver function tests after 2 weeks. Medication therapy seems to be justified in markedly symptomatic patients with relapsing hepatitis. PMID- 10875302 TI - Is chromium a trace essential metal? AB - If chromium is an essential metal it must have a specific role in an enzyme or cofactor, and a deficiency should produce a disease or impairment of function. To date, no chromium-containing glucose tolerance factor has been characterized, the purpose of the low-molecular-weight chromium-binding protein is questionable, and no direct interaction between chromium and insulin has been found. Furthermore, chromium3+ is treated like the toxic metals arsenic, cadmium, lead and mercury in animals. Chromium3+ may be involved in chromium6+-induced cancers because chromium6+ is converted to chromium3+ in vivo, and chromium3+ is genotoxic and mutagenic. Although there is no direct evidence of chromium deficiencies in humans, dietary supplements exist to provide supraphysiological doses of absorbable chromium3+. Chromium3+ may act clinically by interfering with iron absorption, decreasing the high iron stores that are linked to diabetes and heart disease. If so, this would make chromium3+ a pharmacological agent, not an essential metal. PMID- 10875303 TI - Gender influences the effect of perinatal copper deficiency on cerebellar PKC gamma content. AB - Change in cerebellar protein kinase C gamma (PKCgamma) content caused by perinatal copper (Cu) deficiency was determined in 22-day old rats. The offspring of dams with low Cu intake during gestation and lactation exhibited signs characteristic of Cu deficiency including anemia, greater than 90% reduction in liver Cu concentration, and undetectable serum ceruloplasmin. In addition, brain Cu concentrations were reduced 80%. No differences in the signs of Cu deficiency were observed between female and male offspring. However, cerebellar PKCgamma content was reduced 54% (P < 0.05, Tukey's test) in female offspring but only 18% (P > 0.05) in male offspring. Following 6 weeks of Cu supplementation, brain Cu concentrations remained depressed in female and male rats that experienced perinatal Cu deficiency, but cerebellar PKCgamma content was completely restored to control levels. Postnatal expression of PKCgamma in the cerebellum coincides with and regulates cerebellar maturation. The results of the present study indicate perinatal Cu deficiency may impair cerebellar maturation to a greater extent in females than in males. However, it is not clear whether suppression of PKCgamma by perinatal Cu deficiency produces permanent neuropathology in the cerebellum because the effects were reversed by Cu supplementation. PMID- 10875304 TI - Phytate prevents tissue calcifications in female rats. AB - The AIN-76 A, a purified rodent diet, has a propensity to cause kidney calcifications in female rats which is not observed with non-purified rodent diets, suggesting a nutritional factor that avoids these calcifications. One candidate is phytate, which inhibits crystallisation of calcium salts and is practically absent in purified diets. Therefore, the effects on calcification of kidney tissue of phytate addition to the AIN-76 A diet using female Wistar rats were studied. The rats were assigned to three groups: AIN-76 A, AIN-76 A + 1% phytate and standard nonpurified chow. Urinary phytate of the AIN-76 A fed group was undetectable. Urinary phytate of AIN-76 A + 1% phytate and standard fed groups did not differ and was significantly higher than in the AIN-76 A group. The concentrations of calcium and phosphorus in kidneys were greater in the AIN 76 A group than in AIN-76 A + 1% phytate and standard groups. Only rats of the AIN-76 A group displayed mineral deposits at the corticomedullary junction. These findings demonstrated that the absence of phytate in the AIN-76 A diet is one of the causes of renal calcification in female rats. PMID- 10875305 TI - Ecto-diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A)-hydrolase is expressed as an ectoenzyme in a variety of mammalian and human cells and adds new aspects to the turnover of Ap4A. AB - Ap4A and other dinucleotides participate in the regulation of hemostasis and blood pressure control. With the exception of two previously reported surface anchored ectoAp4A-hydrolases on bovine aortic endothelial and chromaffine cells, all Ap4A-hydrolases reported are intracellular or freely soluble. We demonstrated that ectoAp4A-hydrolases are present on a broad variety of cell types of different species: rat mesangial, bovine corneal epithelial, human Hep-G2 and peridontal cells. Ectoenzyme properties were evaluated on rat mesangium cells. Chromatography of purified plasma membranes on Sephacel 300 resulted in enrichment of ectoAp4A-hydrolase and in separation from ectoATPase. In contrast to ATPase, Ap4A-hydrolase was stable at room temperature. EctoAp4A-hydrolase also recognized ATP as substrate, and therefore is not highly specific. The molecular weight was 180 kD. Unlike ectoAMPase ectoAp4A-hydrolase was not attached via a glycosyl-phosphatidylinositol (GPI)-moiety. Concentrations of PI-PLC 10-100-fold higher than effective for ectoAMPase cleavage (10-100 mU/ml) plus extensively extended incubation times up to eight hours did not result in cleavage of ectoAp4A-hydrolase. The enzyme ectoAp4A-hydrolase might presage a direction for pharmaceutical manipulation in the control of blood pressure and hemostasis. PMID- 10875307 TI - Standards for the Ph.D. degree in the molecular biosciences. Recommendations of the Committee on Education of The International Union of Biochemistry and Molecular Biology. PMID- 10875306 TI - Scavenger receptor regulation and atherosclerosis. AB - Atherosclerosis and its complications, such as coronary heart disease, heart infarction and stroke, are the leading causes of death in the developed world. High blood pressure, diabetes, smoking and a diet high in cholesterol and lipids clearly increase the likelihood of premature atherosclerosis, albeit other factors, such as the individual genetic makeup, may play an additional role. During atherosclerosis, uncontrolled cholesterol and lipid accumulation in macrophages and smooth muscle cells leads to foam cell formation and to the progression of the atherosclerotic plaque. This review will focus on foam cell formation within the atherosclerotic lesion, the involvement of the scavenger receptor genes in this process, and the possibility to interfere with scavenger receptor function to reduce the progression of atherosclerosis. To date, the regulatory mechanisms for the expression of scavenger receptor genes and their role in atherosclerosis are not well characterized. Knowledge on this subject could lead to a better understanding of the process, prevention and therapy of this disease. PMID- 10875308 TI - Bone loss prevention and breast cancer risk. PMID- 10875309 TI - Does traction still have a role in nonspecific low back disorders? PMID- 10875310 TI - New factors controlling bone remodeling. AB - Two factors of crucial importance in bone cell differentiation were discovered within the last two years. One is the transcription factor Osf2/Cbfa1, which allows mesenchymal stem cells to differentiate into osteoblasts. Soluble factors, including bone morphogenetic proteins (BMPs), leptin, and TGF-beta, can modulate differentiation of mesenchymal stem cells to osteoblasts or to other cell types such as chondrocytes or adipocytes. The other recent discovery is osteoclast differentiating factor (ODF), which is specific for and indispensable to osteoclast differentiation. ODF belongs to the TNF family. Its soluble receptor, osteoprotegerin, prevents it from binding to osteoclasts, thus inhibiting its activity. A role of lymphocytes in bone remodeling has long been suspected, and it has now been shown that ODF is produced by activated T lymphocytes, which may therefore be implicated in bone loss accompanying inflammation. Finally, recent evidence supports a role for B lymphocytes in bone loss secondary to estrogen deprivation. In conclusion, these recent data may have important applications. Osteoprotegerin is a potent antiosteoclast agent that may prove useful in the treatment of bone disorders. Osf2/Cbfa1 and ODF are major targets in the treatment of osteoporosis. PMID- 10875311 TI - Pain, distress and joint hyperlaxity. AB - Pain dominates the lives of many patients with hyperlaxity syndromes, most commonly the Benign Joint Hypermobility Syndrome (BJHS/EDS). As a result they may experience psychosocial problems, which in many cases severely affects their healthy functioning. Above all is the overriding chronic pain in joints, muscles and ligaments, which arises from an inherent predisposition to the effects of everyday trauma, but other factors such as associated osteoarthritis or fibromyalgia are also important. There may also be neurophysiological factors at play producing nociceptive enhancement. Pain and distress of visceral origin can result from laxity of connective tissue within or providing support for the abdominal, thoracic or pelvic viscera leading to hernia, uterine and/or rectal prolapse, mitral valve prolapse or spontaneous pneumothorax. In children joint hyperlaxity is an important (and often unrecognised) source of rheumatic symptoms, which may be ignored or erroneously ascribed to juvenile idiopathic arthritis. The management of pain and distress in the hyperlaxity syndromes requires skill, patience, compassion and understanding. Often the results of conventional anti-rheumatic therapy (including anti-rheumatic drugs and surgery) as applied to other rheumatic diseases are disappointing and innovative approaches are required. Amongst these, for which evidence of efficacy is available, are physiotherapeutic and orthotic stabilisation of hyperlax joints, proprioceptive enhancement and the newer pain management techniques including cognitive behavioural therapy. PMID- 10875312 TI - Amyloidosis physiopathology. AB - Amyloidoses belong to the emerging group of conformational diseases. In the case of amyloid, the conformational change leads to pathologic deposition in tissues. Amyloid and amyloidosis are especially associated with inflammatory disorders and ageing. Twenty animal and human proteins with various primary structures are involved in amyloid deposits formation. Thermodynamic and kinetic aspects of the abnormal protein folding of amyloid proteins are better known. Several molecules of the extracellular matrix, especially amyloid P component and proteoglycans, are tightly linked to amyloid proteins within the deposits. However, their roles in amyloid deposits formation is not yet thoroughly established. Common treatments of amyloidoses are based on decreasing the availability of the amyloid protein precursor. Novel treatments will aim at blocking conformational changes of the amyloid protein and inhibiting amyloid protein-extracellular matrix interactions. PMID- 10875313 TI - Phlebotomy: a treatment of ischaemic necrosis of the femoral head? Analysis of a hypothesis. AB - The treatment of non-traumatic ischaemic osteonecrosis of the femoral head (ONFH) remains problematical and there is evidently scope to seek for a medical treatment of this disease which often leads to a surgical procedure for hip prosthesis. If we exclude the context of hemoglobin disorders, necrosis appears mainly in adults, when their limb bones contain a fatty marrow. Investigations such as intramedullary pressure measurement and transosseous phlebography suggest a disorder of the intraosseous blood circulation. Various studies in animals and man have provided arguments indicating that a kind of intraosseous 'obesity' due to hyperplasia and/or hypertrophy of the fatty tissue of the femoral marrow play a role in the development of necrosis. In this respect, the blood flow of the yellow marrow is close to ischemia, whereas red marrow flow can be very high. In various conditions, mainly hemolytic anemias, the femoral yellow marrow can convert to red marrow, which has also been described in anemias induced by blood loss. If it is not thought unreasonable to consider ONFH an 'ischemic' disease, these observations are an encouragement to attempt treatment by repeated phlebotomies. This procedure may locally restore red marrow, and then an adequate blood flow which could stabilize or even reverse the lesions if they are diagnosed early. PMID- 10875314 TI - Compliance with drug therapy in rheumatoid arthritis. A longitudinal European study. AB - OBJECTIVE: To delineate compliance with drug therapy in rheumatoid arthritis patients, determine specific characteristics of compliant and noncompliant patients, and look for changes in compliance over time. PATIENTS AND METHODS: A prospective European cohort study (EURIDISS) recruited 556 patients in four countries over three years. Compliance with drug dosages and dosing times was evaluated yearly using a questionnaire. RESULTS: Of the 556 patients, 429 (77.2%) were on drug therapy at all three evaluation time points. Use of steroids, nonsteroidal anti-inflammatory drugs, and second-line drugs varied significantly across countries. The compliance behavior was stable over time in 59.5% of them (35.7% of patients were consistently compliant and 23.8% consistently noncompliant); it was independent of disease duration and from the clinical features of the disease. Older patients and women were more likely to be compliant (odds ratios, 2.5 and 2, respectively). CONCLUSION: Compliance with drug therapy can be measured using two simple questions. Compliance is more closely dependent on individual behavior than on responses to specific features of rheumatoid arthritis. PMID- 10875315 TI - Anti-Ro(SS-A) 52 kDa and 60 kDa specificities in undifferentiated connective tissue disease. AB - Autoantibodies to Ro(SS-A) may recognize two different polypeptides, of 52 kDa and 60 kDa, respectively. We used an ELISA with purified human recombinant antigens to conduct a detailed analysis of the specificities of anti-Ro(SS-A) antibodies from 170 patients with definite diagnoses (systemic lupus erythematosus [SLE], n = 55; primary Sjogren's syndrome [PSS], n = 39; systemic sclerosis, n = 9; rheumatoid arthritis [RA], n = 10) or undifferentiated connective tissue disease (UCTD, n = 57). Most of the patients with SLE or PSS had both anti-52 kDa and -60 antibodies; isolated anti-60 kDa antibodies were found in 13% of the SLE patients and in none of the PSS patients, whereas high titers of anti-52 kDa were more common in the PSS than in the SLE patients. In the UCTD patients, the anti-Ro(SS-A) profile showed no significant correlations with clinical features but was associated with the clinical outcome. Over the mean follow-up of five years, definite SLE developed in four of the five UCTD patients with isolated anti-60 kDa vs only one of the remaining 52 patients (P < 0.0001); progression to PSS was seen in seven of the 34 patients with both anti 52 kDa and anti-60 kDa vs none of the remaining 23 patients (P = 0.03); none of the 12 patients with isolated anti-52 kDa developed a definite connective tissue disease. CONCLUSION: Our study suggests that analysis of anti-Ro(SS-A) specificity may provide useful information for predicting the course of UCTD. PMID- 10875316 TI - Interleukin-16 in synovial fluids from cases of various types of arthritis. AB - OBJECTIVES: To examine the characteristic relationship between interleukin-16 (IL 16) and clinical data in various types of arthritis. METHODS: We measured IL-16 levels of the synovial fluids (SF) of patients with various types of arthritis, which included rheumatoid arthritis, traumatic arthritis, pseudogouty arthritis, gouty arthritis, and osteoarthritis, by an enzyme immunosorbent assay, and examined their correlations with clinical parameters. RESULTS AND CONCLUSIONS: Higher levels of IL-16 in synovial fluid from patients with rheumatoid arthritis, traumatic arthritis, and pseudogouty arthritis, compared to those with osteoarthritis, and gouty arthritis were indicated. Also, synovial IL-16 levels in patients with rheumatoid arthritis correlated significantly, especially with synovial matrix metalloproteinase-3 levels. But the IL-16 levels of both synovial fluid and peripheral blood did not correlate with conventional inflammatory parameters such as C-reactive protein, erythrocyte sedimentation rate, or rheumatoid factor. Although the function of IL-16 in inflammatory arthritis has not yet been defined, these data indicated some essential features of IL-16. PMID- 10875317 TI - Clinical, radiological, and laboratory findings in Lebanese spondylarthropathy patients according to HLA-B27 status. AB - To evaluate clinical, radiologic, and laboratory features in Lebanese spondylarthropathy patients according to HLA-B27 status. METHODS: We retrospectively compared demographic, clinical, radiologic, and severity data in 40 HLA-B27-positive and 58 HLA-B27-negative patients. All 98 patients met Amor's or European Spondylarthropathy Study Group criteria for spondylarthropathy, and 51.7% met New York modified criteria for ankylosing spondylitis. RESULTS: Onset before 16 years of age, hip involvement, and an elevated mean erythrocyte sedimentation rate were significantly associated with the presence of the HLA-B27 (32.5 vs 13.8%, P=0.02; 45 vs 7.5%, P=0.001; and 47.7 vs 25.4, P=0.02; respectively). The two groups were comparable for age, sex ratio, prevalence and distribution of spondylarthropathy types, family history, sacroiliitis, bamboo spine, syndesmophytes, peripheral joint involvement, enthesopathies, extra articular involvement, response to nonsteroidal anti-inflammatory drugs, and need for other medications. CONCLUSION: In Lebanon, spondylarthropathy patients positive for HLA-B27 experience disease onset at an earlier age, are more likely to develop hip involvement, and have laboratory evidence of more severe inflammation than their HLA-B27-negative counterparts. None of the other clinical and radiologic parameters are modified by HLA-B27 status. PMID- 10875318 TI - Weak androgen levels, glucocorticoid therapy, and bone mineral density in postmenopausal women with rheumatoid arthritis. AB - OBJECTIVE: To study dehydroepiandrosterone sulfate (DHEAS) and androstenedione (AND) status in postmenopausal women with rheumatoid arthritis (RA), the effects of glucocorticoid therapy on DHEAS and AND levels, and their relationship with bone mineral density (BMD). METHODS: Forty-six postmenopausal women with RA were separated into two groups based on whether they had a negative history for glucocorticoid therapy (n = 24) or were currently on glucocorticoid therapy (n = 22). The control group was composed of 39 postmenopausal women who had never received hormone replacement therapy. Serum DHEAS and AND levels were measured using a radioimmunoassay. BMD was determined at the lumbar spine (L2-L4) and femoral neck using a DEXA Hologic QDR-1000 densitometer. Results. RA patients and controls were similar in age, weight, body mass index, and years since menopause. DHEAS and AND levels were lower in the glucocorticoid-treated RA group than in the other two groups. The glucocorticoid-treated RA group also had a significantly lower femoral BMD value than the nonglucocorticoid-treated RA group. Lumbar BMD was similar in the two RA groups and in the controls. CONCLUSION: Decreases in DHEAS and AND levels in postmenopausal women with RA are probably related to glucocorticoid therapy rather than to the disease itself. PMID- 10875319 TI - Long-term efficacy of percutaneous drill-biopsy under computed tomography guidance of osteoid osteomas of the hip and femur. A review of seven cases. AB - En bloc surgical excision of osteoid osteomas of the hip and femur is difficult because of problems with tumor boundary identification and the need to use internal fixation or bone grafting. We report our experience with seven cases treated by percutaneous drill-biopsy under computed tomography guidance. PATIENTS: Six adults and one child with a mean age of 22 years treated since 1995. All seven patients reported insomnia due to pain during a mean of 10 months, improved by aspirin or other nonsteroidal anti-inflammatory agents. The tumor was seen on plain radiographs in four cases and on computed tomography scans in all seven cases. Magnetic resonance imaging was diagnostic in only two cases. The tumor was in the femoral neck in four patients, in the acetabulum in one, in the proximal femur in one, and at the distal diaphyseal-metaphyseal junction of the femur in one. METHOD: A trephine was used to remove the tumor under computed tomography guidance during a short general anesthesia. RESULTS: No serious complications were recorded. Full weight bearing was allowed starting at the forty-eighth hour in six of the seven patients. Immediate pain relief and a full recovery were obtained in every case. Results were excellent in four patients and good in three after a follow-up of 14 to 44 months. CONCLUSION: Percutaneous drill-biopsy of osteoid osteomas is a valuable alternative to conventional surgery in patients with tumors at sites that are difficult to access. The technique allows early weight bearing and ensures a full recovery. PMID- 10875320 TI - A paleoepidemiologic study of diffuse idiopathic skeletal hyperostosis. AB - OBJECTIVE: To investigate the prevalence of diffuse idiopathic skeletal hyperostosis (DISH) in ancient populations. MATERIAL AND METHODS: We studied 38 women and 61 men aged at least 30 years at death, whose remains were recovered from four cemeteries of the High Middle Ages (VIth to VIIIth century) located in Lorraine (eastern France). Criteria previously developed by Crubezy and Crubezy Ibanez for paleopathological studies were used to determine whether the skeletal remains exhibited definite, possible, probable, or no DISH. RESULTS: Definite DISH with flowing ossification along the thoracic spine and ossification of peripheral enthesopathies was seen in 2.8 to 3.7% of the subjects from two cemeteries and in none from the other two cemeteries. Identical figures were found for possible DISH. The prevalence and male predominance s (nine of 13 subjects in any DISH category were males) were similar to those reported in contemporary studies and in other paleopathological studies. CONCLUSION: In the absence of clinical data on the metabolic status of the study subjects, our results suggest that DISH in our subjects was a marker for older age at death. PMID- 10875321 TI - Bone mineral density in patients with type 1 diabetes mellitus. AB - Although osteopenia is often reported as a complication of type 1 diabetes mellitus, its frequency and severity remain unclear, and studies of bone mineral density in type 1 diabetics have yielded conflicting results. We measured bone mineral density at the lumbar spine and femoral neck in 88 Spanish adults with type 1 diabetes mellitus responsible for moderately severe complications. Mean age (+/- SD) was 28.9 +/- 8.8 years, and mean disease duration was 11.2 +/- 6.4 years. As compared to normal Spanish adults, bone mineral density was decreased in the patients at the lumbar spine (Z-score, -0.32 +/- 1.08; P < 0.001) but not at the femoral neck (Z-score, -0.21 +/- 1.03; P non-significant). The magnitude of bone loss in the diabetics was small (T-score, -0.38 +/- 1.13 at the lumbar spine and -0.37 +/- 1.08 at the femoral neck). Only three patients met WHO criteria for osteoporosis at one or both measurement sites. Patients with retinopathy (n = 37) had lower lumbar spine bone mineral density values than patients without retinopathy; however, this difference was no longer present after adjustment for age and disease duration. Bone mineral density values were similar in patients with (n = 13) and without microalbuminuria. Our findings suggest that bone loss is not a major problem in younger type 1 diabetics with short disease durations and no severe diabetic complications. PMID- 10875322 TI - Prompt regression of paravertebral and epidural abscesses in patients with pyogenic discitis. Sixteen cases evaluated using magnetic resonance imaging. AB - OBJECTIVE: To determine whether spinal magnetic resonance imaging performed one month into anti-microbial therapy for pyogenic discitis demonstrated changes of value for predicting outcomes and making therapeutic decisions. METHODS: Prospective study of 16 patients with discitis. A physical evaluation, laboratory tests for inflammation, plain radiographs centered on the affected vertebral level, and magnetic resonance imaging with gadolinium injection were performed at baseline and on day 30. All 16 patients were reevaluated after three months and 15 after six months. RESULTS: The 12 men and four women had a mean age of 59 years. Fourteen patients had a paravertebral abscess (n=12) and/or an epidural abscess (n=6). On day 30, 14 of the 16 patients were clinically improved and ten were radiologically improved. The C-reactive protein level was still elevated in five cases. All reevaluated patients were improved after three months (16/16) and six months (15/15). Only two magnetic resonance imaging features improved during the first month: the size of paravertebral abscesses decreased in 11 of 12 patients, and the size of epidural abscesses decreased in four of six patients. CONCLUSION: Paravertebral and epidural abscesses improve promptly under antimicrobial therapy. However, the presence of these lesions have no prognostic significance. Magnetic resonance imaging does not add significantly to the follow up of patients who respond clinically to antimicrobial therapy. However, in patients with an unsatisfactory clinical response, follow-up magnetic resonance imaging can provide useful therapeutic orientation by showing whether the paravertebral and/or epidural abscesses have decreased in size. PMID- 10875323 TI - Septic arthiritis due to Prevotella bivia in a patient with rheumatoid arthritis. AB - Prevotella bivia is a gram-negative anaerobic bacteria traditionally classified in the genus Bacteroides, and usually reported in obstetric and gynecologic infections. To date, there has been only one description of infectious arthritis secondary to this germ. We report the first case of septic arthritis due to Prevotella bivia in a patient with severe, long-lasting rheumatoid arthritis (RA) treated with low doses of corticosteroids. RA is a well-known predisposing factor to septic arthritis, whose causes are multifactorial (general immunosuppression induced by the disease and its therapy, presence of prosthetic joints, etc.). However, infectious arthritis due to anaerobic bacteria is rarely observed. In general, clinical presentation varies widely: insidious onset, apyrexia, and lack of inflammatory signs or systemic disturbance are frequent features. Joint infection is generally secondary to hematogenous spread, the Bacteroides fragilis group being the most commonly isolated pathogens. Early diagnosis and prompt treatment, with drainage and debridement of the affected joint and specific antimicrobial treatment with Metronidazole are essential for a successful outcome. PMID- 10875324 TI - Brown tumor of the spine revealing secondary hyperparathyroidism. Report of a case. AB - Brown tumors are classic skeletal manifestations of hyperparathyroidism usually seen in severe forms with subperiosteal bone resorption. They are exceedingly rare at the spine. We report the case of a woman on hemodialysis who developed two brown tumors, including one at the spine, as the first manifestations of secondary hyperparathyroidism. There were no neurologic manifestations. The clinical and radiologic abnormalities resolved after parathyroidectomy preceded by a bisphosphonate infusion to prevent possible spinal lysis progression during the immediate postoperative period. In addition to the location of one of the brown tumors at the spine, unusual features in this case were the explosive onset of hyperparathyroidism and the absence of detectable subperiosteal resorption. We also suggest that bisphosphonate therapy given before parathyroidectomy may be useful in patients with spinal brown tumors. PMID- 10875325 TI - Septic arthritis of the spine facet joint: early positive diagnosis on magnetic resonance imaging. Review of two cases. AB - We report two cases of septic arthritis of the spine facet joint in two patients with no previous medical history. Clinical symptoms were consistent with a spondylodiscitis. Blood cultures were positive for Staphylococcus aureus. The infection was initially shown and precisely localised with magnetic resonance imaging, despite an initially negative or aspecific bone scintigraphy. Magnetic resonance imaging of the spine demonstrated infection involving the epidural space and paraspinal musculature and enhancement of the infected thoracic and lumbar facet joint after gadolinium injection. PMID- 10875326 TI - Chondritis dissecans of concave articular surfaces (acetabulum and glenoid fossa). Two case reports. AB - We report two patients who had chronic mechanical pain due to substantial cartilage defects documented by arthrography. One of these patients had pain in the hip and declined surgical treatment. The other had pain in the glenohumeral joint; surgery found a fissure in the glenoid fossa cartilage and provided prompt pain relief. These two cases serve as reminders that investigation of the cartilage and subchondral bone in young patients with mechanical pain should not be confined to the convex articular surfaces, although these are more likely to sustain trauma-related damage than concave surfaces. In both our patients, the diagnosis required thin-section computed arthrotomography. PMID- 10875327 TI - Concomitant rheumatoid arthritis and amyotrophic lateral sclerosis. A puzzle illustrated by a new case. AB - We report a case of amyotrophic lateral sclerosis in a patient with rheumatoid arthritis. Only three similar cases have been reported. Our case illustrates the diagnostic difficulties raised by early amyotrophic lateral sclerosis responsible for localized or unusual manifestations. Occurrence of the two diseases in the same patient is probably due to chance alone. PMID- 10875328 TI - Frozen shoulder and fluoroquinones. Two case reports. PMID- 10875329 TI - Carpal tunnel syndrome complicating hand flexor tenosynovitis due to Mycobacterium szulgai. PMID- 10875330 TI - Munchhausen's syndrome. A case-report. PMID- 10875331 TI - The endo-beta-1,4-glucanase CelA of Clavibacter michiganensis subsp. michiganensis is a pathogenicity determinant required for induction of bacterial wilt of tomato. AB - The phytopathogenic bacterium Clavibacter michiganensis subsp. michiganensis NCPPB382, which causes bacterial wilt and canker of tomato, harbors two plasmids, pCM1 (27.35 kb) and pCM2 (72 kb), encoding genes involved in virulence (D. Meletzus, A. Bermpohl, J. Dreier, and R. Eichenlaub, 1993, J. Bacteriol. 175:2131 2136; J. Dreier, D. Meletzus, and R. Eichenlaub, 1997, Mol. Plant-Microbe Interact. 10:195-206). The region of pCM1 carrying the endoglucanase gene celA was mapped by deletion analysis and complementation. RNA hybridization identified a 2.4-knt (kilonucleotide) transcript of the celA structural gene and the transcriptional initiation site was mapped. The celA gene encodes CelA, a protein of 78 kDa (746 amino acids) with similarity to endo-beta-1,4-glucanases of family A1 cellulases. CelA has a three-domain structure with a catalytic domain, a type IIa-like cellulose-binding domain, and a C-terminal domain. We present evidence that CelA plays a major role in pathogenicity, since wilt induction capability is obtained by endoglucanase expression in plasmid-free, nonvirulent strains and by complementation of the CelA- gene-replacement mutant CMM-H4 with the wild-type celA gene. PMID- 10875332 TI - Nodule-expressed Cyp15a cysteine protease genes map to syntenic genome regions in Pisum and Medicago spp. AB - PsCyp15a is a gene that encodes a vacuolar cysteine protease expressed in wilt induced shoots of Pisum sativum (pea) and in root nodules. To further the understanding of nodular PsCyp15a expression, a region 5' to the coding sequence of the gene was cloned. Varying lengths of 5' untranslated sequence were fused with the uidA coding region and introduced from Agrobacterium rhizogenes into "hairy roots" of Vicia hirsuta. In this transgenic root nodulation assay, a promoter sequence of 900 bp was sufficient to give an expression pattern indistinguishable from that obtained in pea nodules by in situ hybridization. An orthologue of PsCyp15a was cloned from nodule mRNA of Medicago sativa and a corresponding gene identified in M. truncatula was also shown to express strongly in nodules. With molecular mapping techniques, it was demonstrated that these genes map to a syntenic genome location in pea and Medicago spp., but the map positions of the Cyp15a genes cannot be correlated with existing nodulation mutants. PMID- 10875333 TI - The BMP1 gene is essential for pathogenicity in the gray mold fungus Botrytis cinerea. AB - In Magnaporthe grisea, a well-conserved mitogen-activated protein (MAP) kinase gene, PMK1, is essential for fungal pathogenesis. In this study, we tested whether the same MAP kinase is essential for plant infection in the gray mold fungus Botrytis cinerea, a necrotrophic pathogen that employs infection mechanisms different from those of M. grisea. We used a polymerase chain reaction based approach to isolate MAP kinase homologues from B. cinerea. The Botrytis MAP kinase required for pathogenesis (BMP) MAP kinase gene is highly homologous to the M. grisea PMK1. BMP1 is a single-copy gene. bmp1 gene replacement mutants produced normal conidia and mycelia but were reduced in growth rate on nutrient rich medium. bmp1 mutants were nonpathogenic on carnation flowers and tomato leaves. Re-introduction of the wild-type BMP1 allele into the bmp1 mutant restored both normal growth rate and pathogenicity. Further studies indicated that conidia from bmp1 mutants germinated on plant surfaces but failed to penetrate and macerate plant tissues. bmp1 mutants also appeared to be defective in infecting through wounds. These results indicated that BMP1 is essential for plant infection in B. cinerea, and this MAP kinase pathway may be widely conserved in pathogenic fungi for regulating infection processes. PMID- 10875334 TI - Nodule invasion and symbiosome differentiation during Rhizobium etli-Phaseolus vulgaris symbiosis. AB - By means of a detailed ultrastructural analysis of nodules induced by Rhizobium etli on the roots of Phaseolus vulgaris, we observe that the development of host invaded cells is not synchronous. An accumulation of mitochondria was found in freshly invaded host cells, containing only a few symbiosomes (SBs) that are released from highly branched intracellular ramification of the infection threads. Moreover, besides the fusion between the SB membrane with host secretory vesicles, we observe also a great number of fusions between the outer leaflets of adjoining SB membranes, thus resulting in structures that resemble the tight junction network (zona occludens with a five-layered structure) of epithelian cells. This process was found to be induced strongly and earlier both in the invaded host cells of ineffective nodules (elicited by Fix- mutant strains of R. etli) and in the older (senescence) invaded cells of effective nodules, whereas bacteroid division is seldom if ever observed. Our observations strongly suggest that multiple-occupancy SBs also arise by fusion of single-occupancy SBs and the physiological consequence of this process is discussed. PMID- 10875335 TI - Cloning and characterization of a cyclic peptide synthetase gene from Alternaria alternata apple pathotype whose product is involved in AM-toxin synthesis and pathogenicity. AB - Afternaria afternata apple pathotype causes Alternaria blotch of susceptible apple cultivars through the production of a cyclic peptide host-specific toxin, AM-toxin. PCR (polymerase chain reaction), with primers designed to conserved domains of peptide synthetase genes, amplified several products from A. alternata apple pathotype that showed high similarity to other fungal peptide synthetases and were specific to the apple pathotype. Screening of a Lambda Zap genomic library with these PCR-generated probes identified overlapping clones containing a complete cyclic peptide synthetase gene of 13.1 kb in length with no introns. Disruption of this gene, designated AM-toxin synthetase (AMT), by transformation of wild-type A. afternata apple pathotype with disruption vectors resulted in toxin-minus mutants, which were also unable to cause disease symptoms on susceptible apple cultivars. AM-toxin synthetase is therefore a primary determinant of virulence and specificity in the A. alternata apple pathotype/apple interaction. PMID- 10875336 TI - Entry of Rhizobium leguminosarum bv.viciae into root hairs requires minimal Nod factor specificity, but subsequent infection thread growth requires nodO or nodE. AB - Using various mutant strains of Rhizobium leguminosarum bv. viciae, we have investigated the role of nodO in stimulating infection thread development in vetch and pea. Analysis of R. leguminosarum bv. viciae nodE and nodO mutants revealed no significant difference from the wild-type infection phenotype. Conversely, an R. leguminosarum bv. viciae nodE nodO double mutant was severely impaired in its ability to form normal infection threads. This strain displayed a number of novel infection-related events, including intracellular accumulations of bacteria at the base of root hairs, distended and enlarged infection threads, and reversed threads growing up root hairs. Since normal infection was seen in a nodE mutant, nodO must suppress these abnormal infection phenomena A deletion mutant, retaining only the nodD and nodABCIJ genes, also formed intracellular accumulations at the base of root hairs. Addition of R. leguminosarum bv. viciae nodO could alleviate this phenotype and restore some infection thread formation, although these threads appeared to be abnormal. Exogenous application of R. leguminosarum bv. viciae Nod factors could not alleviate the aberrant infection phenotype. Our results show that the most basic Nod factor structure can allow bacterial entry into the root hair, and that nodO can promote subsequent infection thread development. PMID- 10875337 TI - Differential expression of eight chitinase genes in Medicago truncatula roots during mycorrhiza formation, nodulation, and pathogen infection. AB - Expression of eight different chitinase genes, representing members of five chitinase classes, was studied in Medicago truncatula roots during formation of arbuscular mycorrhiza with Glomus intraradices, nodulation with Rhizobium meliloti, and pathogen attack by Phytophthora megasperma f. sp. medicaginis, Fusarium solani f. sp. phaseoli (compatible interactions with root rot symptoms), Ascochyta pisi (compatible, symptomless), and F. solani f. sp. pisi (incompatible, nonhost interaction). In the compatible plant-pathogen interactions, expression of class I, II, and IV chitinase genes was enhanced. The same genes were induced during nodulation. Transcripts of class I and II chitinase genes accumulated transiently during early stages of the interaction, and transcripts of the class IV chitinase gene accumulated in mature nodules. The pattern of chitinase gene expression in mycorrhizal roots was markedly different: Expression of class I, II, and IV chitinase genes was not enhanced, whereas expression of three class III chitinase genes, with almost no basal expression, was strongly induced. Two of these three (Mtchitinase III-2 and Mtchitinase III 3) were not induced at all in interactions with pathogens and rhizobia. Thus, the expression of two mycorrhiza-specific class III chitinase genes can be considered a hallmark for the establishment of arbuscular mycorrhiza in Medicago truncatula. PMID- 10875338 TI - Localized expression of cathepsin B-like sequences from root nodules of pea (Pisum sativum). AB - Cathepsin B is an ancient family of eukaryotic cysteine proteases. We describe PsCat1, a plant cathepsin B-like transcript, identified as an expressed sequence in Rhizobium-induced, nitrogen-fixing root nodules of pea. In situ hybridization studies in root nodules showed strong, extremely localized expression of PsCat1 in individual cells associated with the central infected tissue. Restriction fragment polymorphism mapping of the PsCat1 locus in pea shows no correlation with existing mutant lines defective in symbiosis. PMID- 10875339 TI - The Ustilago maydis ubc4 and ubc5 genes encode members of a MAP kinase cascade required for filamentous growth. AB - Ustilago maydis, the causal agent of corn smut disease, displays dimorphic growth in which it alternates between a budding haploid saprophyte and a filamentous dikaryotic pathogen. We are interested in identifying the genetic determinants of filamentous growth and pathogenicity in U. maydis. To do this we have taken a forward genetic approach. Earlier, we showed that haploid adenylate cyclase (uac1) mutants display a constitutively filamentous phenotype. Mutagenesis of a uac1 disruption strain allowed the isolation of a large number of budding suppressor mutants. These mutants are named ubc, for Ustilago bypass of cyclase, as they no longer require the production of cyclic AMP (cAMP) to grow in the budding morphology. Complementation of a subset of these suppressor mutants led to the identification of the ubc4 and ubc5 genes, which are required for filamentous growth and encode a MAP (mitogen-activated protein) kinase kinase kinase and a MAP kinase kinase, respectively. Evidence suggests that they are important in the pheromone response pathway and in pathogenicity. These results further support an important interplay of the cAMP and MAP kinase signal transduction pathways in the control of morphogenesis and pathogenicity in U. maydis. PMID- 10875340 TI - The ORF8 gene product of Agrobacterium rhizogenes TL-DNA has tryptophan 2 monooxygenase activity. AB - The open reading frame 8 (ORF8) is located on the TL-DNA of the phytopathogenic soil bacterium Agrobacterium rhizogenes strain A4. The predicted ORF8 protein has a particular structure and is possibly a natural fusion protein. The N-terminal domain shows homology to the A. rhizogenes rolB protein and may modulate the auxin responsiveness of host cells. The C terminus has up to 38% homology to tryptophan 2-monooxygenases (t2m). We show that ORF8 overexpressing plants contain a fivefold higher concentration of indole-3-acetamide (IAM) than untransformed plants. Protein extracts from seedlings and Escherichia coli overexpressing ORF8 show significantly higher turnover rates of tryptophan to IAM than negative controls. We conclude that the ORF8 gene product has tryptophan 2 monooxygenase activity. PMID- 10875341 TI - Experimental validations of the HELS method for reconstructing acoustic radiation from a complex vibrating structure AB - This paper presents experimental validations of the Helmholtz Equation Least Squares (HELS) method [Wang and Wu, J. Acoust. Soc. Am. 102, 2020-2032 (1997); Wu and Wang, U.S. Patent Number 5712805 (1998); Wu, J. Acoust. Soc. Am. 107, 2511 2522 (2000)] on reconstruction of the radiated acoustic pressures from a complex vibrating structure. The structure under consideration has geometry and dimensions similar to those of a real passenger vehicle front end. To simulate noise radiation from a vehicle, a high fidelity loudspeaker installed inside the structure at the location of the engine is employed to generate both random and harmonic acoustic excitations. The radiated acoustic pressures are measured over a finite planar surface above the structure by a microphone. The measured data are taken as input to the HELS formulation to reconstruct the acoustic pressures on the top surface of the structure as well as in the field. The reconstructed acoustic pressures are then compared with measured ones at the same locations. Also shown are comparisons of the reconstructed and measured acoustic pressure spectra at various locations on the surface. Results show that satisfactory reconstruction can be obtained on the top surface of the structure subject to both random and harmonic excitations. Moreover, the more measurements and the closer their distances to the source surface, the more accurate the reconstruction. The efficiency of the HELS method may decrease with increasing of the excitation frequency. This high frequency difficulty is inherent in all expansion theories. PMID- 10875342 TI - Wave propagation in micro-heterogeneous porous media: a model based on an integro differential wave equation AB - A model hyperbolic partial differential equation with singular convolution operators and infinitely smooth solutions is studied. It is shown that short pulses, including finite-bandwidth pulses, propagate with a delay with respect to the wavefront. For a two-parameter family of such equations Green's functions are obtained in a simple self-similar form. As an application, it is demonstrated that the Gurevich-Lopatnikov dispersion law for a thin-layered porous medium can be approximated by a hyperbolic equation with singular memory. PMID- 10875343 TI - Directive line source model: a new model for sound diffraction by half planes and wedges AB - A new method termed Directive Line Source Model (DLSM) is presented for predicting the diffracted field produced by a sound wave incident on a rigid or pressure release half plane. In the new method the edge of the half plane is modeled as an infinite set of directive point sources continuously distributed along the edge. Because DLSM is fast, simple and intuitive, it is a promising tool for the study of diffraction. It can be applied for several types of incident radiation: omnidirectional cylindrical and spherical waves, plane waves, as well as waves from directional sources. Wedges may also be treated. Finally, DLSM can handle diffraction by an arbitrarily shaped edge profile, for example, a half plane having an edge that is jagged instead of straight. Results for plane, cylindrical and spherical incident waves, as well as for arrays of line and point sources, are presented and their agreement with known analytical solutions is demonstrated. Predictions based on DLSM compare favorably with experimental data. PMID- 10875344 TI - Limits of time-reversal focusing through multiple scattering: long-range correlation AB - Experimental results of time-reversal focusing in a high-order multiple scattering medium are presented and compared to theoretical predictions based on a statistical model. The medium consists of a random collection of parallel steel rods. An ultrasonic source (3.2 MHz) transmits a pulse that undergoes multiple scattering and is recorded on an array. The time-reversed waves are sent by the array back to the source through the scattering medium. The quality of temporal focusing is very well predicted by a simple statistical model. However, for thicker samples, persistent temporal side-lobes appear. We interpret these side lobes as a consequence of the growing number of crossing paths in the sample due to high-order multiple scattering. As to spatial focusing, the resolution is practically independent from the array's aperture. With a 16-element array, the resolution was found to be 30 times finer than in a homogeneous medium. Resolutions of the order of the wavelength (0.5 mm) were attained. These results are discussed in relation with the statistical properties of time-reversal mirrors in a random medium. PMID- 10875345 TI - Analytical regularization based analysis of a spherical reflector symmetrically illuminated by an acoustic beam AB - A mathematically accurate and numerically efficient method of analysis of a spherical reflector, fed by a scalar beam produced by a complex source-point feed, is presented. Two cases, soft and hard reflector surface, are considered. In each case the solution of the full-wave integral equation is reduced to dual series equations and then further to a regularized infinite-matrix equation. The latter procedure is based on the analytical inversion of the static part of the problem. Sample numerical results for 50-lambda reflectors demonstrate features that escape a high-frequency asymptotic analysis. PMID- 10875346 TI - Effects of interaction between two bubble scatterers AB - The backscattering of sound from two regularly arranged bubbles is studied theoretically and experimentally. In well-controlled laboratory experiments a bistatic acoustic system is used to interrogate the scatterers, which are placed on a very fine thread at the same distance d from the combined beam axis of the set of transmitting and receiving transducers. The radius of each bubble is 585 microm. The frequency range is 80-140 kHz, and d is varied so that the variable kd spans the range 0.2-21, where k is the acoustic wave number. Scattering calculations are carried out using an exact, closed-form solution derived from the multiple scattering series. Several experiments are performed, and the results are in close agreement with the calculations. It is verified that multiple scattering induces an oscillatory behavior about the exact coherent scattering level, with decreasing amplitude for increasing kd. For interbubble distance 2d approximately lambda/2 the backscattered radiation is maximized, while for 2d